PMID- 11098212 TI - Events PMID- 11098213 TI - Similia PMID- 11098214 TI - Epidermal growth factor receptor signaling activates orthodenticle expression during Drosophila head development. AB - The relation between signal transduction pathways and the genes that specify regional identity remains poorly understood. We investigated the interaction between the epidermal growth factor receptor (EGFR) pathway and the homeobox gene orthodenticle (otd), which specifies cell fate during head development. Previous studies of head formation in Drosophila melanogaster demonstrated that reducing either EGFR signaling or otd expression in the imaginal primordium of the dorsal head capsule eliminates the ocelli and other dorsal head structures. Here, we show that blocking EGFR signaling reduces otd expression and that activating EGFR signaling outside this primordium induces ectopic otd expression. We also demonstrate that loss of EGFR can be rescued by constitutive otd expression. Our results indicate that otd is a downstream target of the EGFR pathway during head development. PMID- 11098215 TI - Predominant transgene expression in exocrine pancreas directed by the CMV promoter. AB - The enhancer/promoter of the human cytomegalovirus gene encoding the major immediate-early protein (CMVp) is reputed to be one of the strongest and most promiscuous regulatory elements for directing transcription of heterologous genes in vitro. However, transgene expression under the promoter in adult transgenic mice is often more restricted. We selected a CMVp segment from position -350 to +59 to control expression of transgenes for two secretory fusion proteins. Expression was analyzed by immunohistology staining and quantified by Northern blot, Western blot, and ELISA of secretions from explanted tissues. In all six lines of transgenic mice, the highest expression of transgenes at the mRNA and protein level was observed in the exocrine tissue of the pancreas, although the levels of expression varied among the lines. The results indicate not only that CMVp is not a universal promoter in vivo but indeed that it can be relatively specific for the exocrine pancreas, where expression of the gene it controlled was consistently very high. PMID- 11098216 TI - Intermediate filaments reconstituted from vimentin, desmin, and glial fibrillary acidic protein selectively bind repetitive and mobile DNA sequences from a mixture of mouse genomic DNA fragments. AB - Employing the whole-genome PCR technique, intermediate filaments (IFs) reconstituted from vimentin, desmin, and glial fibrillary acidic protein were shown to select repetitive and mobile DNA sequence elements from a mixture of mouse genomic DNA fragments. The bound fragments included major and minor satellite DNA, telomere DNA, minisatellites, microsatellites, short and long interspersed nucleotide elements (SINEs and LINEs), A-type particle elements, members of the mammalian retrotransposon-like (MaLR) family, and a series of repeats not assignable to major repetitive DNA families. The latter sequences were either similar to flanking regions of genes; possessed recombinogenic elements such as polypurine/polypyrimidine stretches, GT-rich arrays, or GGNNGG signals; or were characterized by the distribution of oligopurine and pyrimidine motifs whose sequential and vertical alignment resulted in patterns indicative of high recombination potentials of the respective sequences. The different IF species exhibited distinct quantitative differences in DNA selectivities. Complexes consisting of vimentin IFs and DNA fragments containing LINE, (GT)(n) microsatellite, and major satellite DNA sequences were saturable and dynamic and were formed with high efficiency only when the DNAs were partially denatured. The major-groove binder methyl green exerted a stronger inhibitory effect on the binding reaction than did the minor-groove binder distamycin A; the effects of the two compounds were additive. In addition, DNA footprinting studies revealed significant configurational changes in the DNA fragments on interaction with vimentin IFs. In the case of major satellite DNA, vimentin IFs provided protection of the T-rich strand from cleavage by DNase I, whereas the A-rich strand was totally degraded. Taken together, these observations suggest that IF protein(s) bind to double-stranded DNAs at existing single-stranded sites and, taking advantage of their helix-destabilizing potential, further unwind them via a cooperative effort of their N-terminal DNA-binding regions. A comparison of the present results with literature data, as well as a search in the NCBI database, showed that IF proteins are related to nuclear matrix attachment region (MAR) binding proteins, and the DNA sequences they interact with are very similar or even identical to those involved in a plethora of DNA recombination and related repair events. On the basis of these comparisons, IF proteins are proposed to contribute in a global fashion, not only to genetic diversity, but also to genomic integrity, in addition to their role in gene expression. PMID- 11098217 TI - Genomic organization, expression, and chromosome localization of a third aurora related kinase gene, Aie1. AB - We previously reported two novel testis-specific serine/threonine kinases, Aie1 (mouse) and AIE2 (human), that share high amino acid identities with the kinase domains of fly aurora and yeast Ipl1. Here, we report the entire intron-exon organization of the Aie1 gene and analyze the expression patterns of Aie1 mRNA during testis development. The mouse Aie1 gene spans approximately 14 kb and contains seven exons. The sequences of the exon-intron boundaries of the Aie1 gene conform to the consensus sequences (GT/AG) of the splicing donor and acceptor sites of most eukaryotic genes. Comparative genomic sequencing revealed that the gene structure is highly conserved between mouse Aie1 and human AIE2. However, much less homology was found in the sequence outside the kinase-coding domains. The Aie1 locus was mapped to mouse chromosome 7A2-A3 by fluorescent in situ hybridization. Northern blot analysis indicates that Aie1 mRNA likely is expressed at a low level on day 14 and reaches its plateau on day 21 in the developing postnatal testis. RNA in situ hybridization indicated that the expression of the Aie1 transcript was restricted to meiotically active germ cells, with the highest levels detected in spermatocytes at the late pachytene stage. These findings suggest that Aie1 plays a role in spermatogenesis. PMID- 11098218 TI - Structure of the gene for uteroferrin. AB - The published structure of the gene for uteroferrin differs from that of the human and mouse tartrate-resistant acid phosphatase (TRAP) genes. Polymerase chain reaction using genomic DNA as template and primers designed from exon 2 of the porcine uteroferrin gene amplified a product containing two previously undescribed introns. Because of these discrepancies, we cloned an EcoRI fragment from a porcine genomic BAC library containing the uteroferrin gene, and the region containing the uteroferrin gene was completely sequenced. The uteroferrin gene spanned 2.5 kb and contained five exons, which is similar to the structure previously reported for human and mouse TRAP genes but different from the published structure of the uteroferrin gene. Southern blotting of porcine genomic DNA digested with a variety of enzymes was consistent with the sequence that we obtained. The most likely explanation for the differing results is that the previously reported structure for the uteroferrin gene was the result of artifactual elimination of introns 2 and 3 by bacteria and artifactual recombination of the region upstream of the transcription start site of this gene. PMID- 11098220 TI - A new arrival: evidence about differential diagnosis. PMID- 11098219 TI - Purification, cloning, and characterization of a second arylalkylamine N acetyltransferase from Drosophila melanogaster. AB - In insects, amine acetylation by the enzyme arylalkylamine N-acetyltransferase (AANAT) is involved in melatonin formation, sclerotization, and neurotransmitter inactivation. This wide spectrum of activities suggests that several AANAT enzymes are present. We recently purified a protein fraction with AANAT activity from Drosophila melanogaster and cloned the corresponding gene, aaNAT1. Following the same strategy, we now report the purification of an additional AANAT from D. melanogaster, AANAT2, and the cloning of the corresponding cDNA. The isolated protein differs from AANAT1a and AANAT1b in its molecular weight and isoelectric point. The AANAT2 shares about 30% identity with the products of the aaNAT1 gene. The enzyme does not follow one-site Michaelis-Menten kinetics when assayed with various concentrations of the arylalkylamine tryptamine and a constant concentration (0.5 mM) of the cofactor acetyl coenzyme A. The data can be interpreted in terms of an enzyme with two kinetic regimes (K(m1) = 7.2 microM, K(m2) = 0.6 mM, and v(max2) = 2.7 v(max1)) that are governed by binding of the substrate to a regulatory site (K(r) = 6.2 mM). These findings demonstrate the presence of a second expressed gene encoding an AANAT in D. melanogaster. Northern blot analysis revealed no diurnal variation of aaNAT2 transcription, similar to the results obtained for aaNAT1a and aaNAT1b. PMID- 11098221 TI - [Radiobiology]. PMID- 11098222 TI - [Mechanisms of repair and radiation-induced mutagenesis in higher eukaryotes]. AB - Cells of higher eukaryotes possess several very efficient systems for the repair of radiation-induced lesions in DNA. Different strategies have been adopted at the cellular level to remove or even tolerate various types of lesions in order to assure survival and limit the mutagenic consequences. In mammalian cells, the main DNA repair systems comprise direct reversion of damage, excision of damage and exchange mechanisms with intact DNA. Among these, the direct ligation of single strand breaks (SSB) by a DNA ligase and the multi-enzymatic repair systems of mismatch repair, base and nucleotide excision repair as well as the repair of double strand breaks (DSB) by homologous recombination or non homologous end joining are the most important systems. Most of these processes are error-free except the non homologous end-joining pathway used mainly for the repair of DSB. Moreover, certain lesions can be tolerated by more or less accurately acting polymerases capable of performing translesional DNA syntheses. The DNA repair systems are intimately integrated in the network of cellular regulation. Some of their components are DNA damage inducible. Radiation-induced mutagenesis is largely due to unrepaired DNA damage but also involves error-prone repair processes like the repair of DSB by non-homologous end-joining. Generally, mammalian cells are well prepared to repair radiation-induced lesions. However, some questions remain to be asked about mechanistic details and efficiencies of the systems for removing certain types of radiation-damage and about their order and timing of action. The answers to these questions would be important for radioprotection as well as radiotherapy. PMID- 11098223 TI - [Regulation of cell cycle and radiation-induced cell death]. AB - Tight control of cell proliferation is mandatory to prevent cancer formation as well as to normal organ development and homeostasis. This occurs through checkpoints that operate in both time and space and are involved in the control of numerous pathways including DNA replication and transcription, cell cycle progression, signal transduction and differentiation. Moreover, evidence has accumulated to show that apoptosis is tightly connected with the regulation of cell cycle progression. In this paper we describe the main pathways that determine checkpoints in the cell cycle and apoptosis. It is also recalled that in solid tumors radiation-induced cell death occurs most frequently through non apoptotic mechanisms involving oncosis, and mitotic or delayed cell death. PMID- 11098224 TI - [Radiation-induced superficial fibrosis and TGF-alpha 1]. AB - Radiation-induced fibrosis is a late sequela of both therapeutic and accidental irradiations, which has been described in various tissues, including the lung, liver, kidney and skin. This review presents different aspects of superficial radiation-induced fibrosis, such as clinical observations, histological changes, cellular and molecular regulations, and medical management. Recent evidence on the critical role played by TGF-beta 1 in the initiation, development and persistence of fibrosis are discussed, as well as the possibility that this cytokine may constitute a specific target for antifibrotic agents. PMID- 11098225 TI - [Clinical aspects of research in radiobiology. Past and future directions]. AB - Over the last ten years the impact of fundamental radiation biology into daily radiotherapy has been of concern chiefly to fractionation, prediction of radiation response, tumour oxygenation, intrinsic radiosensitivity including genetic approaches, and the determinants of the outcome of chemoradiotherapy combinations. Future goals will rely on sophisticated approaches, based on the progress of molecular and cellular biology and the characterisation of new targets for radiation. Some of these novel advances will be discussed. PMID- 11098226 TI - [Peritoneal carcinosis in appendiceal carcinoma--at what price can the prognosis be improved?]. AB - BACKGROUND AND AIM: Many patients with appendix carcinoma develop peritoneal carcinomatosis with poor prognosis. The purpose of this study was to look for results of multimodality treatment in these patients. PATIENTS AND METHODS: Data of 13 patients (11 men, 2 women, median age 58 years) with proven peritoneal carcinomatosis from appendiceal carcinoma, operated between 07/1995 and 10/1998 were analysed retrospectively with special regard to extent of resection, postoperative morbidity and mortality, survival. RESULTS: A macroscopically complete cytoreduction could be achieved in all patients by multivisceral resection with peritonectomy. Intraoperatively, after resection, an open, hyperthermic intraperitoneal chemotherapy with Cisplatin was performed. Morbidity and 90-days-mortality were 62% and 15%, respectively. After a median follow-up of 36 months all patients survived the therapy (excepting two postoperative deaths). CONCLUSIONS: In selected patients with advanced appendix carcinoma and peritoneal carcinomatosis prognosis can be improved by peritonectomy with intraperitoneal hyperthermic chemotherapy. This results in high mortality and morbidity. PMID- 11098227 TI - [How safe is premedication in ambulatory endoscopy in Germany? A prospective study in gastroenterology specialty practices]. AB - BACKGROUND: To date there is no prospective large-scale study of the risk of premedication on the complication rate of outpatient gastrointestinal endoscopy in Germany. PATIENTS AND METHODS: In 67 gastroenterological practises in Germany the number of esophagogastroduodenoscopies (EGD) and colonoscopies was recorded from april 1998 until march 1999. All serious complications had to be recorded in a structured protocol. RESULTS: The overall complication rate for EGD (n = 110,469) was low (0.009%) and about two third of the adverse effects were due to premedication (0.006%). The overall complication rate for diagnostic colonoscopy (n = 82,416) was 0.02% and the complication rate associated to premedication was 0.01%. An individual dosage of premedication for EGD and colonoscopy was given by all gastroenterologists. Most of the gastroenterologists applied premedication in 10 to 50 percent of the patients for EGD and in 70 to 100 percent of the patients for colonoscopy. Most of the cardiorespiratory adverse effects in colonoscopy occurred if sedation was performed by combination of benzodiazepines and opioids and in upper gastrointestinal endoscopy with high dosages of diazepam. In 6 of 45 practises disoprivan (propofol) was used for premedication in 2-50 percent of the colonoscopies. Two of four perforations in diagnostic colonoscopy occurred under sedation with disoprivan. CONCLUSIONS: Outpatient gastrointestinal endoscopy performed by German gastroenterologists in a safe. The complication rate is low compared to the international literature and could be further decreased by avoiding a combination of benzodiazepines and opioids for sedation in colonoscopy and high dosages of diazepam in upper gastrointestinal endoscopy. PMID- 11098228 TI - [Hemiparesis as a sequela of traumatic pseudoaneurysm of the carotid bifurcation]. AB - HISTORY AND CLINICAL FINDINGS: A 20-year-old patient was referred to our clinic after sudden onset of a left-sided hemiparesis. His past history revealed a severe trauma 4 years ago, including multiple bone fractures, rupture of the spleen, as well as renal failure, and an acute respiratory distress syndrome, from which he showed good recovery. At that time no central-nervous symptoms could be found. However, eighteen months ago, he complained about a transient weakness of his left arm and leg. INVESTIGATIONS: Examination of the cerebral arteries by duplex-sonography showed an aneurysm on the bifurcation of the right carotid artery with a peripheral flow-reduction. This could be confirmed by CT- and MR-based angiography, which also revealed a reopened embolic occlusion of the M1-segment of the right middle cerebral artery. On CT and diffusion weighted imaging there was evidence of an ischemic infarction pattern. TREATMENT AND COURSE: Under anticoagulation therapy with heparin the patient showed complete recovery from his symptoms. Duplex-sonography as well as MR-angiography documented a complete reopening of the primarily occluded middle cerebral artery. Finally, surgical reconstruction of the aneurysmatic part of the vessel was done. CONCLUSION: This case illustrates the potential risk of a traumatic aneurysm as a potential source of ischemic brain infarctions. We emphasize the importance of imaging the cerebral arteries in traumatic patients, even in the absence of initial neurological symptom. PMID- 11098229 TI - [Glycoprotein IIb/IIIa receptor antagonists for improving thrombolytic and interventional therapy in patients with acute myocardial infarct]. PMID- 11098230 TI - [Fitness for diving, diving accidents and diving-related illnesses--an overview]. PMID- 11098231 TI - [Procaine in pain therapy of acute pancreatitis]. PMID- 11098232 TI - [International consensus on reflux disease of the esophagus]. PMID- 11098233 TI - [Clinical significance of Helicobacter pylori: pathogen or saprophyte?]. PMID- 11098234 TI - [Do we need nutrition medicine?]. PMID- 11098235 TI - [Value of color-coded duplex ultrasound in patients with polymyalgia rheumatica without signs of temporal arteritis]. AB - BACKGROUND AND OBJECTIVE: Recent studies have described characteristic sonographic signs in patients with manifest temporal arteritis (TA). It was the aim of this study to determine whether sonography can identify TA without clinical signs but biopsy evidence of giant-cell arteritis in patients with rheumatic polymyalgia (RP). PATIENTS AND METHODS: 22 patients (14 women, 8 men; average age 67.4 +/- 9.1 years) with RP but no clinical signs of TA were prospectively examined for TA by colour-coded duplex sonography (ATL HDI 3000, linear 12-5 Mhz) before temporal artery biopsy was taken. If there was clinical suspicion of extratemporal involvement, other vessels were also examined selectively. The biopsy was taken from a site identified by the sonography. A definitive diagnosis of TA was made only if there was a positive biopsy. RESULTS: In seven of the 22 patients (32%) sonography showed an echo-poor halo around the lumen of the temporal artery. Five of these seven patients also had histological evidence of giant-cell arteritis. Conversely, all of the five patients had abnormal sonographic findings, namely a marked halo with a minimal thickness of 0.7 mm. Two of the five patients also had temporal artery stenosis, i.e. there was a 100% sensitivity and 80% specificity with respect to the halo sign in conformance with the histology. Two patients with TA were also shown sonographically to have stenosis in arteries of the shoulder girdle and arm. Stenoses in the renal and mesenteric arteries as well of the coeliac trunk were demonstrated in one patient. CONCLUSIONS: Colour-coded sonography with a high frequency transducer head probably provides reliable diagnosis of TA in patients with RP, even in the absence of clinical signs of vascular inflammation. It remains to be proven whether sonography without biopsy is reliable enough for the diagnosis and treatment of asymptomatic TA. PMID- 11098236 TI - [Contagious impetigo--pathogen spectrum and therapeutic consequences]. AB - OBJECTIVE: The aim of this prospective study was to compare the clinical picture of contagious impetigo (C.I.) with the causative organism and to generate data of the susceptibility of bacteria as the basis for adequate therapy. PATIENTS AND METHODS: In 126 patients with C.I. (86 children, 66 of them younger than 10 years) bacterial swabs were taken and antibiotic sensitivity testing for isolated organisms was tested. RESULTS: In all cases in which contents of vesicles or pustules were analysed, Staphylococcus aureus was the only pathogen isolated. In non-bullous variants of C.I. Staphylococcus aureus was the most often isolated organism as well. Both staphylococci and streptococci were isolated in 12 cases, whereas in just 9 cases streptococci were the only pathogen detected. All Staphylococcus aureus isolates were sensitive to flucloxacillin and cefotaxime. Erythromycin-resistance amounted to more than 20 percent. The percentage of resistant staphylococci against the predominantly topically applied antibiotics fusidinic acid and mupirocin was 2 and 0 per cent, respectively. CONCLUSION: For all manifestations of C.I. Staphylococcus aureus is at present the leading organism which has to be taken into consideration for treatment. If oral antibiotic therapy is indicated, penicillinase-stable penicillins or cephalosporins, preferably of the cefalexin-type, are the drugs of choice. Macrolides are no longer recommended for initiating of C.I. treatment. PMID- 11098237 TI - [Multilocular fixed drug reaction simulating intertrigo in a diabetic patient]. AB - HISTORY AND CLINICAL FINDINGS: A 57-year-old man with diabetes and hypertension was treated with amoxycillin, clarithromycin and pantoprazole for a gastric ulcer positive for Helicobacter pylori. On the second treatment day he developed inguinal pruritus with erythema. He presented at out-patient clinic on the 5th day suspected of having Candida intertrigo. He had bright red, relatively well circumscribed erythema, most marked at the edges, mainly over the inguinal region and the inside of the thigh. There were no other symptoms. INVESTIGATIONS AND DIAGNOSIS: Bacteriological and mycological tests of the affected skin were unremarkable. Immunological tests showed a normal total IgE but were negative in the CAP-FEIA test for penicilloyl G, penicilloyl V, amoxycilloyl and ampicilloyl. An epifocal epicutaneous test with amoxycillin and ampicillin (5% each in vaseline and doritin) gave a +2 positive reaction and confirmed a suspected fixed drug reaction. TREATMENT AND COURSE: After amoxycillin had been discontinued and local class III steroids had been administered (mometasone furoate, Ecural) for one week the cutaneous changes disappeared without complication, except for slight hyperpigmentation. H. pylori eradication was continued without further complications using clarithromycin, metronidazole and pantoprazole. The patient was issued with an "allergic to penicillin" card. CONCLUSION: Intertriginous changes during antibiotic treatment may not be due to Candida intertrigo, which is fairly common, but to a prognostically much more important drug reaction. PMID- 11098238 TI - [Autoantibodies in hepatology and gastroenterology]. PMID- 11098239 TI - [Anticoagulation in atrial fibrillation]. PMID- 11098240 TI - [Adjuvant drug therapy of malignant melanoma. Current knowledge and multi-center studies in German-speaking countries]. PMID- 11098241 TI - [Effectiveness and efficacy of ambulatory diabetes patient education]. PMID- 11098242 TI - [Effectiveness and efficacy of ambulatory diabetes patient education]. PMID- 11098243 TI - [Efficacy and efficiency of ambulatory diabetes patient education]. PMID- 11098244 TI - Relative effectiveness and cost-effectiveness of methods of androgen suppression in the treatment of advanced prostate cancer. AB - OBJECTIVES: With 184,500 new cases and 39,200 deaths anticipated in 1998, prostate cancer is second only to lung cancer in cancer mortality for men. This report is a systematic review of the evidence from randomized controlled trials on the relative effectiveness of alternative strategies for androgen suppression as treatment of advanced prostate cancer. Three key issues are addressed: (1) the relative effectiveness of the available methods for monotherapy (orchiectomy, luteinizing hormone-releasing hormone [LHRH] agonists, and antiandrogens), (2) the effectiveness of combined androgen blockade compared to monotherapy, and (3) the effectiveness of immediate androgen suppression compared to androgen suppression deferred until clinical progression. Outcomes of interest are overall, cancer-specific, and progression-free survival; time to treatment failure; adverse effects; and quality of life. Two supplementary analyses were conducted for each key question: (1) meta-analysis of overall survival at 2 years (questions 1 and 2) and 5 years (questions 2 and 3), and (2) cost-effectiveness analysis. SEARCH STRATEGY: The MEDLINE, CANCERLIT, and EMBASE databases were searched from 1966 to March 1998, and Current Contents to August 24, 1998, for the terms: leuprolide (Lupron); goserelin (Zoladex); buserelin (Suprefact); flutamide (Eulexin); nilutamide (Anandron, Nilandron); bicalutamide (Casodex); cyproterone acetate (Androcur); diethylstilbestrol (DES); and orchiectomy (castration, orchidectomy). The search was then limited to human studies indexed under the MeSH term "prostatic neoplasms" and by the UK Cochrane Center search strategy for randomized controlled trials. Total yield was 1,477 references. SELECTION CRITERIA: We Reports of efficacy outcomes were limited to randomized controlled trials. Phase II studies that reported on withdrawals from therapy and all studies reporting on quality of life were also included. DATA COLLECTION AND ANALYSIS: The systematic review used a prospectively designed protocol conducted by two independent reviewers, with disagreements resolved by consensus. The meta analysis combined data on overall survival using a random effects model. The cost effectiveness analysis used a decision analysis model of advanced prostate cancer with health states and transitions derived from the literature and estimates of effectiveness derived from the meta-analysis. The cost-effectiveness analysis is conducted from a societal perspective, consistent with the guidelines of the U.S. Public Health Service Panel on Cost-Effectiveness in Health and Medicine. MAIN RESULTS: Survival after treatment with an LHRH agonist is equivalent to survival after orchiectomy. The available LHRH agonists are equally effective, and no LHRH agonist is superior to the other when adverse effects are considered. Survival may be somewhat lower with use of a nonsteroidal antiandrogen. There is no statistically significant difference in survival at 2 years between patients treated with combined androgen blockade or monotherapy. Meta-analysis of the limited data available shows a statistically significant difference in survival at 5 years that favors combined androgen blockade. However, the magnitude of this difference is of questionable clinical significance. For the subgroup of patients with good prognosis, there is no statistically significant difference in survival. Adverse effects leading to withdrawal from therapy occurred more often with combined androgen blockade. No evidence is yet available from randomized controlled trials of androgen suppression initiated at prostate-specific antigen (PSA) rise after definitive therapy for clinically localized disease. For patients who are newly diagnosed with locally advanced or asymptomatic metastatic disease, the evidence is insufficient to determine whether primary androgen suppression initiated at diagnosis improves outcomes. (ABSTRACT TRUNCATED) PMID- 11098245 TI - Goldilocks on management. PMID- 11098246 TI - Patient safety: pieces of a puzzle. PMID- 11098247 TI - CGEAN: a resource for nursing administration. AB - In the last decade, CGEAN has expanded its membership to include more nurse executives, nurse researchers, and international members. CGEAN is also responding to changes in healthcare administration as it becomes more evidence based and in faculty roles, which are increasingly emphasizing external research funding as a role expectation. This year 's forum on research development held during the business meeting at the NARC conference yielded several new initiatives to support faculty development and student training in research. We invite readers to join with us to implement these new initiatives. PMID- 11098248 TI - Nurses and assistive personnel. Do patients know the difference? PMID- 11098249 TI - Managed healthcare in South Africa. PMID- 11098250 TI - Tel-eNurse Practice: a practice model for role expansion. AB - RNs are expanding their role in the continuum of care, incorporating telecommunication into their practice to provide efficient and effective patient care. However, role expansion requires the definition and articulation of nursing practice and related outcomes. The Tel-eNurse Practice Model is a theoretical framework nurses can use to define and guide the complex process of care in the telephone encounter. Application of the model gives nurse administrators a way to identify competencies and develop standards of practice and quality and outcomes measurement tools. PMID- 11098251 TI - Building community in the healthcare workplace, Part 4. Partnering with union members to create community. AB - Squeezed between mounting budget pressures and staffing demands, healthcare managers have little room to maneuver as they are buffeted by incessant new strategies. So what is the constant that they can use to keep a steady course through all the chaos? We suggest the overarching constant is "community in the workplace." Although community building may have both short- and long-term benefits, it is not a strategy but a career-long philosophy to help managers tie together all the facets of their daily work into a more meaningful and satisfying purpose. This article, part 4 in a 4-part series, discusses partnering with union leaders and members to create a sense of community in the organization. Using a case study of an organization that had significant success implementing the community philosophy, this article describes concepts, structures, and processes that work, and a few that do not, in a unionized environment. Part 1 (July/August) described the negative dynamics that obstruct sustainable change and reasons for moving to community at work. Part 2 (September) discussed the basic characteristics and principles of community building. Part 3 (October) illustrated community implementation options and obstacles. PMID- 11098252 TI - Assessing the reported financial benefits of unlicensed assistive personnel in nursing. AB - Use of unlicensed assistive personnel in nursing care is thought to reduce labor expense with a neutral or improved effect on quality. A review of the literature during the past decade suggests that there have been few long-term studies that either support or contradict this assumption. PMID- 11098253 TI - The RUG-III case mix classification system for long-term care nursing facilities: is it adequate for nurse staffing? AB - The purpose of this study was to explore the validity of the nursing time associated with the Resource Utilization Group Version III (RUG-III) case mix classification system by determining its potential usefulness for making nurse staffing decisions in long-term care (LTC) facilities. Experts in LTC nursing administration were asked to determine the amount of nursing time required by nursing home residents. The estimates were significantly higher than the RUG-III nursing time for all case mix groups. This finding suggests that the nursing time associated with the RUG-III system may not meet the needs of nursing home residents if used as a basis for nurse staffing. Further analysis questions the wisdom of mandating minimum staffing standards for LTC facilities without taking into account the individual needs of residents. PMID- 11098254 TI - The use of information generated by a patient classification system. AB - Patient classification systems (PCS) generate information that can be used to make decisions about the provision of nursing services. Expenditures for information technology are significant, but no previous research had been conducted regarding the actual use of information generated by an automated PCS. This article reports the findings of a study describing how and why information obtained from a PCS is used in hospital decision making and discusses the implications for nurse administrators. PMID- 11098255 TI - Market, hospital, and nursing unit characteristics as predictors of nursing unit skill mix: a contextual analysis. AB - The objective was to examine the relative contributions of market, hospital, and nursing unit characteristics to the prediction of nursing unit skill mix. The strongest predictors of nursing skill mix were found to be the technological sophistication of the hospital, the volume of services offered, and the percentage of patients covered by managed care payors. The results suggest that skill mix on nursing units is buffered from market forces but is highly sensitive to hospital characteristics. These factors need to be taken into account when decisions are made about the allocation of personnel resources. In this article, the terms "nursing skill mix" and "nurse staffing" are used interchangeably. PMID- 11098256 TI - Should medicine reach out to the spirit? PMID- 11098257 TI - SSRIs for vasodepressor syncope? PMID- 11098258 TI - Difficult-to-manage asthma. How to pinpoint the exacerbating factors. AB - In difficult-to-manage asthma, effective control depends on identification and alleviation of exacerbating factors, such as ongoing allergen exposure, chronic sinusitis, GERD, and emotional stress. Level of compliance with the prescribed medication regimen should be evaluated in all patients. Hormonal factors (i.e., menses, use of exogenous hormones by female patients, and hyperthyroidism) also can exacerbate asthma. When aggressive management fails, the possibility of a misdiagnosis should be considered. Other conditions that can mimic asthma include COPD, congestive heart failure, airway obstruction due to various causes, vocal cord dysfunction, and esophageal spasm. Referral to an asthma specialist is advised in severe or resistant cases. PMID- 11098259 TI - The many faces of confusion. Timing and collateral history often hold the key to diagnosis. AB - Recognition of a patient's state of confusion is only the beginning of a clinical odyssey that can implicate a huge spectrum of diagnostic possibilities. Among these are delirium, depression, dementia, and sensory deprivation. However, with appropriate physical examination and laboratory studies, collateral history, and clarification of time course for the symptom complex, the cause of confusion need not remain confusing. PMID- 11098260 TI - Suicide prevention in primary care. Careful questioning, prompt treatment can save lives. AB - Suicide is a major public health problem in the United States. Psychiatric disorders that feature or lead to suicidal ideation may be overlooked in the primary care setting because patients are reluctant to broach the subject. In this article, Dr Hamilton describes how physicians can open a dialogue with patients so that those who need treatment receive it before they resort to suicide. PMID- 11098261 TI - Simplified approach to somatization disorder. When less may prove to be more. AB - Somatization disorders--manifestations of mental pain--take many forms and can be difficult to diagnose and treat. By definition, most patients with this condition do not want to be cured. Antidepressant or antianxiety medication and referral to a support group or psychiatrist can help patients who are willing to participate in their treatment. Other patients may receive the most benefit from primary care physicians who accept the limitations of treatment, listen to their patient's concerns, and provide reassurance. PMID- 11098262 TI - Coming to terms with grief and loss. Can skills for dealing with bereavement be learned? AB - To provide optimum patient care, physicians often need to actively work with persons who experience an acute loss or terminal illness. Physicians who are aware of and open to the issues involved in the grief process are likely to maintain healthier lives and be more available to their patients and their patients' loved ones at times of loss. However, physicians must also have a way to process the losses they themselves face. Thus, self-care is an important aspect of grieving for both physician and patients. Although the experience of loss is often painful and emotionally charged, it can provide an opportunity for growth and healing. Staying open to the process and the complex feelings involved in grieving promotes greater connection with ourselves and with our patients and their families. PMID- 11098263 TI - Slowly resolving and nonresolving pneumonia. Questions to ask when response is delayed. AB - What is the proper course of action when pneumonia symptoms seem to linger? Is the disease running a typical course, or is further evaluation needed? Dr Johnson addresses these important questions with an outline of risk factors for delayed resolution and a detailed listing of diagnostic considerations. He also discusses reasonable timing of further studies and the usefulness of radiography and fiberoptic bronchoscopy. PMID- 11098264 TI - Perioperative cardiovascular evaluation. Step-by-step approach to risk assessment and follow-up care. AB - One of the integral responsibilities in primary care medicine is medical consultation, especially when a surgical procedure is contemplated. A good grasp of the elements of multiple specialties and an understanding of the complexity of a proposed operation are essential for an appropriate preoperative evaluation. This article covers the steps in assessing cardiovascular risk in candidates for noncardiac surgery, including consideration of patient-specific and surgery specific factors. The authors also recommend close postoperative follow-up to reduce morbidity and mortality. PMID- 11098265 TI - Underlying causes of erythema nodosum. Lesions may provide clue to systemic disease. PMID- 11098266 TI - A rare cause of annular papules. Disseminated superficial actinic porokeratosis. PMID- 11098267 TI - Suicide prevention. PMID- 11098268 TI - [Effectiveness of combined vancomycin and pefloxacine in gastrointestinal decontamination for preventing infections after chemotherapy-induced bone marrow aplasia. A randomized double-blind study]. AB - OBJECTIVE: To test the value of the combination of pefloxacin and vancomycin as gastro-intestinal tract decontamination for the prevention of infections in patients with chemotherapy-induced neutropenia. PATIENTS AND METHODS: Oral pefloxacin plus vancomycin (48 patients), pefloxacin alone (51 patients), or placebo (52 patients) were administered in a randomized double-blind study. Evaluation was done by determining site and documentation of infections, organisms responsible for bacteriologically documented infections, organisms acquired in surveillance cultures and number of days with fever during aplasia. RESULTS: Patients receiving pefloxacin had significantly fewer episodes of bacteremia with enterobacteriacae. No differences were noted between patients treated by pefloxacin and those who received a combination of pefloxacin with vancomycin regarding gram-positive (Gram+) infections and infections with gram negative (Gram-) organisms usually resistant to pefloxacin. However, placebo gave similar results. There was no induction of resistance to pefloxacin during the study. Tolerance of treatment was excellent. Only a prolonged aplasia has been observed in patients receiving pefloxacin. CONCLUSION: Thus, the combination of vancomycin with pefloxacin was not more efficacious than pefloxacin only for the prevention of Gram+ infections in the neutropenic patient. The systematic use of antibiotics as gastrointestinal tract decontamination for the prevention of infections in patients with aplasia may be questionable. PMID- 11098269 TI - [Development of bioethics. Analysis of practices in 20 hospital departments]. AB - OBJECTIVE: In order to analyze the constitution and management of DNA banks and the limitations on procedures currently used in France, we conducted a study in a sample of French hospital units. A DNA bank was defined as any facility where individual samples of DNA, irrespective of the form, are stored for ongoing or future gene studies. The aim of this work was to focus attention on the need for in-depth thought on the ethical issues involved in storing and using DNA as part of everyday clinical practice and research and to provide elements for a debate on the interest and limitations of the French bioethics laws whose revision is currently being considered. METHODS: A questionnaire was sent to the heads of departments in two university hospitals. Five areas were retained for evaluation: the nature and degree of DNA storage activities, procedures followed for reporting the DNA bank, procedures used to protect confidentiality, information given to patients and procedures used for informed consent, and finally, internal rules governing the bank's operation. The answers to the questionnaires were analyzed anonymously. RESULTS: Among 20 hospital units collecting DNA samples, 70% also stored other samples (DNA, blood, tissue, cell lines) including a large proportion of tissue samples. These samples were collected for purposes of fundamental research and clinical practice. The number of samples stored was quite variable, ranging from a few dozen samples to more than 40,000. Only 55% of the units had reported the facility to a control body, in compliance with current law. Seventy percent maintained computerized data bases but only 50% used an anonymous code. Seventy-five percent obtained written consent but patients were not always informed of the site of the storage or the transfer of their DNA. In addition, the validity of the consent over time, the duration of storage and the types of studies that could be conducted on the DNA were poorly detailed. Internal rules governing the bank's activities were not implemented by most of the units (65%). CONCLUSION: The responses to our questionnaire survey show that there is currently a gap between everyday practice and regulatory procedures concerning DNA banks in France. Further analysis of current practices would appear to be necessary so professionals could become more aware of the human and social issues involved with the use of DNA banks. These data on everyday practices should be made available to health care workers, public officials and law makers in order to promote ethical practices that, as has been observed, are not dictated solely by legislation which often lags behind everyday activities. All those involved in the management of DNA banks must be aware of their responsibilities in protecting patients' rights within the framework of a system based on information, consent, and over-the-board trustworthiness regularly submitted to short and long term assessments. A correct response to ethical issues is the only means of developing a real process of social interaction which cannot be achieved by revision of the bioethics laws alone. PMID- 11098270 TI - [Cystic dystrophia of the duodenal wall with aberrant pancreas. Value of magnetic resonance imaging for diagnosis]. AB - BACKGROUND: Cystic dystrophy of the duodenal wall is characterized by the development of cysts in heterotopic pancreatic tissue localized in the duodenal wall. Diagnosis is difficult and endosonography is considered as the most useful investigation. CASE REPORTS: A 40-year-old man was hospitalized for abdominal pain triggered by ingestion of alcoholic beverages and for vomiting after food intake. The diagnosis of acute atypical cephalic pancreatitis was initially made. Magnetic resonance cholangiography then magnetic resonance imaging suggested the diagnosis of cystic dystrophy of the duodenal wall in a heterotopic pancreas which was confirmed by endosonography. DISCUSSION: Magnetic resonance imaging evidenced several cystic formations within a thickened duodenal wall leading to the correct diagnosis. MRI appears to provide all the elements necessary for the diagnosis of cystic dystrophy of the duodenal wall, avoiding the need for invasive investigations. PMID- 11098271 TI - [Rhino-cerebral mucormycosis complicated by intracerebral hemorrhage with favorable outcome]. PMID- 11098272 TI - [HBs antigenemia and vaccination against hepatitis B virus: attention to chronology of markers]. PMID- 11098273 TI - [Atypical presentation of carcinomatous meningitis in neuroendocrine tumor of the cervix uteri]. PMID- 11098274 TI - [Opiates and food habits: a pitfall in drug screening]. PMID- 11098275 TI - [What is the "best" treatment of depression in general practice?]. PMID- 11098276 TI - [Asymptomatic stenosis of the internal carotid artery: causes and risks of cerebrovascular accident]. PMID- 11098277 TI - [Psychiatry and clinical research. A dialogue with hospital directors]. AB - Clinical research must be considered as a main and compulsory medical activity which has to be promoted on a European level. Some difficulties appear to be specific to mental diseases and may represent obstacles or a reason not to participate in clinical research. In fact, financing, grants, organizations and research centers do exist and should push forward clinical research in psychiatry. Human and cultural factors may explain why so few breakthroughs have occurred in France in the considered field. Incentive politics as well as avoidance of technical and frequently definitive errors must be proned and issue from a deep analysis of how leading teams are organized and operate. PMID- 11098278 TI - [Chemicals toxic to the olfactory system. Analysis and description]. AB - AN IMPORTANT PROBLEM: Occupational exposure to chemical products can have toxic effects on the olfactory system. An important number of patients have experienced olfactory disorders subsequent to the development of the chemical industry and atmospheric pollution. EPIDEMIOLOGY DATA: Straightforward data are difficult to collect because several cofactors other than the toxic product are involved. Two lists of toxic products can be made. The first list includes products for which scientific data is available and the second products for which data is lacking. Olfactory tests also differ between authors and countries. TWO TYPES OF TOXICITY: Acute, accidental toxicity is evidenced by the lesions caused by inhalation of high-doses of strongly toxic agents. Chronic intoxication caused by lower concentrations of these inhaled agents does not produce a trigeminal reflex leading to a modified respiratory rate reducing the airborne aggression. APPROXIMATIONS: Clinical data describing the olfactory toxicity of certain industrial and chemical compounds are very significant but often cannot prove a cause and effect relationship. Data obtained with experimental models in rodents are difficult to extrapolate to humans. PMID- 11098279 TI - [Medico-economic analysis of nosocomial infections]. AB - OBJECTIVES: The purpose of this study was to examine analyses of the micro economic and macro-economic consequences of hospital-acquired nosocomial infections. MATERIAL AND METHODS: We performed a critical analysis of the methods reported in the literature for economic estimations of the cost of nosocomial infections and determined the objectives that can be assigned to them. DISCUSSION: Macro-economic estimations of the consequences of nosocomial infections clearly have methodological limitations but nevertheless have an important impact since concern about global cost of nosocomial infections is a persuasive argument in favor of prevention programs. Studies based on calculating the estimated mean cost by type of nosocomial infection cannot alone provide a sufficiently reliable data to determine the pertinence of preventive strategies. Cost-efficiency studies (cost per year of saved life, or cost per nosocomial infection avoided) appear to be more adapted. CONCLUSION: One must carefully examine the underlying methodology when using cost-efficacy ratios associated with different strategies as a decision-making tool. PMID- 11098280 TI - [Left ventricular stimulation in treatment of heart failure]. AB - SEARCH FOR AN ALTERNATIVE TREATMENT: The concept of stimulating the ventricle to improve heart function in patients with severe heart failure is an old one, but the first published series was reported about 10 years ago and provided encouraging results that lead to numerous other studies. A PROMISING TECHNIQUE: Excepting a few patients with a long PR, stimulation of the right ventricle is ineffective or even deleterious. Stimulation of the lateral region of the left ventricle has produced some undeniably favorable and sometimes even spectacular improvement in hemodynamic performances. Permanent stimulation of both ventricles or the left ventricle produces an overall clinical improvement in patients with severe heart failure (NYHA III or IV) and major left intraventricular conduction disorders (QRS > 140 ms). These results have been recently confirmed in a prospective randomized trial. CAREFUL OPEN QUESTIONS: There is no fully satisfactory explanation for the improvement which, it is important to note, does not occur in all patients. A more homogeneous contraction of the left ventricle certainly plays a fundamental role. Resynchronization of the two ventricles with dual stimulation is more complex and costly and remains to be evaluated. One crucial question is currently being examined: what is the effect of stimulation on the high mortality in these patients? PMID- 11098281 TI - [Colonic nigrosis, a reversible manifestation?]. PMID- 11098283 TI - [In Process Citation] PMID- 11098284 TI - International Symposium on Neuromuscular Disorders state-of-the-art at the edge of the millennium Brussels, December 11th, 1999. PMID- 11098282 TI - [Tumor of the ampulla of Vater]. PMID- 11098285 TI - Dysferlin and muscular dystrophy. AB - The limb-girdle muscular dystrophies are a highly heterogeneous group of muscle disorders with many different genetic causes now known. Amongst the causes of LGMD, the dysferlin gene stands out as novel for several reasons. It is the first known example of a C2 domain containing protein involved in a muscular dystrophy, mutations in the gene can be involved in a variable phenotype, and a naturally occurring mouse model for dysferlin deficiency has recently been identified. This article reviews the progress made in understanding this form of limb-girdle muscular dystrophy to date. PMID- 11098286 TI - An attempt of gene therapy in Duchenne muscular dystrophy: overexpression of utrophin in transgenic mdx mice. AB - Dystrophin, its functions and the consequences of its absence are briefly reviewed. The animal model of Duchenne myopathy, the mdx mouse, was used to over express utrophin by transgenesis technology. A battery of functional tests, including mechanical responses (force development and resistance to imposed stretch), intracellular calcium homeostasis and metabolic reaction to muscle activity were applied to check the functional recovery obtained by over expression of utrophin. For most parameters tested, recovery amounted to 80%, demonstrating that utrophin can very efficiently act as a surrogate for dystrophin. PMID- 11098287 TI - The expanding clinical and genetic spectrum of the myotonic dystrophies. AB - Core features of the dominantly inherited myotonic dystrophies are myotonia, muscle weakness and cataract. Classic myotonic dystrophy (Steinert's disease) has been defined as a genetic entity by the underlying CTG repeat expansion on chromosome 19q13.3 (= DM1 locus). Later on, another disorder similar to but different from myotonic dystrophy was described as proximal myotonic myopathy (PROMM). The majority of PROMM families have been linked to a recently discovered locus on chromosome 3q21 (= DM2 locus).--This article analyses the clinical features of 70 patients from 14 German PROMM families linked to the 3q locus. In contrast to Steinert's disease, these patients did not reveal mental deficiency; no congenital type was found; weakness was mainly located in the proximal leg muscles; clinical myotonia was very mild and sometimes absent; episodes of pain occurred. In the majority of patients, the disorder seems to be more benign compared to Steinert's disease. However, life threatening cardiac involvement is possible; rarely, muscle weakness may progress until the patient is bedridden.- Some families with a PROMM-like phenotype do not link to the locus on 3q. The group of the myotonic dystrophies will get new members in the future. PMID- 11098288 TI - Mitochondrial cytopathies and neuromuscular disorders. AB - In the last decade, mitochondrial diseases were shown not to be rare but to represent an important group of metabolic disorders. Defects are caused by mutations either located in nuclear genes or in mitochondrial genes. Nuclear gene defects are found in complex IV deficient and complex I deficient patients. Deficiencies of complex II are extremely rare. Different phenotypes are associated with complex IV deficiency, including a neonatal form, cardio encephalomyopathy in young infants, Leigh syndrome, and pure myopathy. Mutations can be found in the complex IV assembly genes, such as the SURF-1 gene and the SCO2 gene. Different phenotypes are also found in complex I deficient patients and include a neonatal form, Leigh syndrome, pure myopathy, pure cardiomyopathy or multiple-system involvement. In some disorders, the mitochondrial DNA abnormalities are caused by a nuclear gene defect (Alpers-Huttenlocher syndrome, autosomal dominant multiple mitochondrial DNA deletion syndrome, and MNGIE syndrome). Since 1988, more then 70 different mutations were reported in the mitochondrial DNA. Some point mutations are associated with a specific phenotype, others have a wide range of clinical symptoms. We expect that many more mitochondrial DNA mutations will be identified in the future. The number of mutations in nuclear genes will also increase, especially since progress has been made in techniques used for identification of nuclear genes (microcell transfer). PMID- 11098289 TI - Nerve biopsy: indications and contribution to the diagnosis of peripheral neuropathy. The experience of the Born Bunge Foundation University of Antwerp and University of Liege between 1987 and 1997. AB - We reviewed 355 nerve biopsies analysed at the Laboratories of Neuropathology of the Born-Bunge Foundation/University of Antwerp (BBF/UIA) and University of Liege (ULg) between 1987 and 1997. We examined the indications for nerve biopsy, the yield of the procedure, and the influence of clinical and neuropathological parameters. Contributory biopsies accounted for 35.5% and 47.3% respectively at ULg and BBF/UIA laboratories: of these, one third showed specific histological findings, the majority being informative only when combined with the relevant clinical data. The profile of indications for nerve biopsy was roughly comparable in both laboratories. The search for an inflammatory neuropathy prompted 35-40% of all biopsies with more than 50% of specimens being informative in this indication. The lowest yield (20%) was obtained among the nerve biopsies performed in the absence of any presumptive aetiology. These accounted for 22-33% of all cases. Inadequate surgical resection, delays in transport or processing errors precluded histological study of 4% (BBF/UIA) to 8% (ULg) of the specimens. We conclude that nerve biopsies should be performed by experienced surgeons and handled in specialised laboratories. Only a relatively small number of causes of neuropathy can be diagnosed on the basis of histology alone. More often, contributory biopsies will result from the combination of non-specific suggestive histological features with relevant clinical information. The diagnostic yield of nerve biopsy is function of careful patient selection and close collaboration between the clinician and the neuropathologist. PMID- 11098290 TI - Pathogenesis and treatment of inflammatory demyelinating polyradiculoneuropathy. AB - Inflammatory demyelinating polyradiculoneuropathy causes a spectrum of conditions ranging from acute (Guillain-Barre syndrome), through subacute to chronic forms. The pathogenesis of acute forms is related to antibody responses against glycolipid epitopes which mimic bacterial, especially Campylobacter jejuni, structures but T cells are also involved. The pathogenesis of chronic forms is poorly understood. Different forms differ in their responses to steroids. Chronic inflammatory demyelinating polyradiculoneuropathy responds to steroids but a variant multifocal motor neuropathy and the acute forms of inflammatory demyelinating polyradiculoneuropathy do not. Acute and chronic forms respond to plasma exchange and intravenous immunoglobulin. PMID- 11098291 TI - Molecular genetics of inherited peripheral neuropathies: who are the actors? AB - Charcot-Marie-Tooth disease, the most common variant of the inherited peripheral neuropathies, has a prevalence of 1/2500. Clinical, electrophysiological, neuropathological and molecular genetic studies have demonstrated extensive heterogeneity. Currently, 30 genetic loci are known for distinct CMT types and related inherited peripheral neuropathies, while many other types have been excluded for linkage to these loci. Recent molecular genetic studies have demonstrated the involvement of 8 genes that encode proteins with very diverse functions. These include a structural protein confined to the compact myelin, a cytoskeletal protein, an adhesion molecule, a gap-junction protein, a transcription factor, a receptor for a neurotrophic factor, a phosphatase and a protein involved in signal transduction and cell cycle regulation. PMID- 11098292 TI - Amyotrophic lateral sclerosis: pathogenesis. PMID- 11098293 TI - Electrodiagnosis of demyelinating neuropathies. AB - The inflammatory demyelinating neuropathies constitute a significant proportion of the acquired polyneuropathies. Major progress in finding the causes and in the treatment of these neuropathies has been made over the last decade. Early recognition is of paramount importance, because timely and appropriate treatment can largely reduce morbidity and disability. Electrodiagnosis plays a key role in the detection and characterization of the inflammatory demyelinating neuropathies. Electrodiagnostic criteria for primary demyelination have therefore been developed. They are empirically based on changes of nerve conduction parameters in populations of patients with a confirmed clinical and laboratory diagnosis of inflammatory demyelinating neuropathy. The challenge consists of defining criteria sets that are highly specific but also as sensitive as possible. Most of the hereditary demyelinating neuropathies are part of Charcot Marie-Tooth disease type 1. The pattern of nerve conduction abnormalities usually provides valuable clues for the distinction from chronic inflammatory demyelinating neuropathies. PMID- 11098294 TI - Effect of stimulus contrast uncertainty on simple reaction time. AB - Both reaction time (RT) and the latency of the visually evoked potentials (VEP) to grating onset increase with increasing stimulus spatial frequency (SF). At SF higher than 5 c/deg RT increases faster than VEP latency, the difference resulting in a "central delay" (Mihaylova et al., 1999). Due to the equipment limitations, RT and VEP experiments of Mihaylova et al. (1999) differed in constancy of stimulus contrast within a sequence of trials. The present experiments were aimed at testing the assumption that the central delay is a result of contrast uncertainty effect on RT. To this end, RT were measured in condition of both constant and variable grating contrast. The stimuli were sinusoidal gratings ranging in SF from 0.5 to 16 c/deg and in contrast from 2.5 to 50%. In addition, VEP were recorded to the same stimuli in blocks of fixed contrast and the latencies of the early VEP wave were subtracted from RT. Contrast uncertainty did not affect RT at low SF, 0.5 and 2 c/deg, while increasing RT at SF higher than 5 c/deg both at low and high stimulus contrast. The results showed that the central component of RT increase at high SF is reduced but not eliminated under constant contrast condition. The uncertainty effect at high SF might be due to contrast sensitivity reduction, reduced subjective stimulus probability and differences in response strategy adopted by the subjects when contrast was constant or variable. An alternative explanation is a larger ability of low SF stimuli compared to high SF stimuli to attract visual attention. PMID- 11098295 TI - Spectral sensitivity of the visual system as revealed by evoked potentials in normal and anomalous trichomats. AB - The spectral sensitivity of the visual system upon adaptation by white light at intensity of 10,500 td was studied in Maxwellian view. The monochromatic stimuli had a size of 8 degrees and duration of 700 ms. They were superimposed on an adaptation field of 14 degrees. The spectral sensitivity was determined by the off-VEP amplitude measured from the negative peak with a latency of 75-80 ms to the positive peak with a latency of 100-110 ms. The EEG was led off monopolarly from the occipital scalp area. Eight normal trichomats, 2 protanomalous and 2 deuteranomalous subjects were investigated. Some specificity of the spectral sensitivity curve, determined by the VEP amplitude, were studied. Special attention was placed on the sensitivity decrease between lambda = 550 nm and lambda = 590 nm, which was much pronounced in the normal trichomats. Maximum sensitivity was observed at lambda = 527 nm. The sensitivity at lambda = 578 nm was lower than the maximum one by 0.8 logarithmic units or more. It is assumed that this sensitivity decrease is a manifestation of opponency between the middle wavelength and long wavelength mechanisms. In the anomalous trichomats the sensitivity decrease between lambda = 550 nm and lambda = 590 nm was much less pronounced. Maximum sensitivity was obtained at lambda = 551 nm. In the long wavelength range of the spectrum the deuteranomalous subjects showed a higher sensitivity as compared to the protanomalous ones. The obtained results suggest disturbances of color opponency in the anomalous trichomats. PMID- 11098296 TI - Experimental setup and instrumentation for measurement and preprocessing of EEG during voluntary goal-directed movements. AB - The widely used practice in the study of human complex voluntary movement organization is to record, measure and analyze EEG activity covering the period of both motor preparation and performance. The main strategy is to reveal EEG characteristics related to both cognitive and motor aspects of the action. To this end a special-purpose experimental set-ups are required providing precise enough measure of timing and characteristics of the movement in order to synchronize EEG changes time-locked to the phases of task performance. We describe an experimental set-up including a special-purpose device, which was designed for study of slow, continuous goal-directed movements. The implementation was aimed to provide (i) performance of complicated enough task in order to force the subject to concentrate his mental activity predominantly on the task; (ii) to control the successive stages of task performance. The advantages of the presented instrumentation was demonstrated by comparing power spectra of EEG segments long before and immediately prior to the movement performance. The instrumentation is flexible enough to be used in a large scale psychophysiological experiments. PMID- 11098297 TI - Studies on paraquat-induced oxidative stress in rat liver. AB - Oxidative injury of liver was studied 20 hr after a single oral administration of 150 mg/kg paraquat (PQ) to rats. PQ exerted no effect on cytosolic superoxide dismutase (SOD) activity but increased mitochondrial SOD activity by 14%. The level of GSH was decreased by 30%, and GSH/GSSG ratio was diminished almost twice. The correlation between the enhancement of mitochondrial SOD activity and the diminution of GSH level by PQ implicates O2- in the liver toxicity of the drug. Mitochondrial aconitase activity was slightly decreased (by 9%) while cytosolic aconitase activity was not affected. The results cast additional light on the responses of both aconitases to oxidative stress. PMID- 11098298 TI - Effect of CPAP on breathlessness perception in healthy subjects during methacholine induced bronchoconstriction. AB - Application of continuous positive airway pressure (CPAP) in asthmatic patients decreases breathlessness (B). The effect of CPAP on induced bronchoconstriction was studied in healthy subjects. The changes in B were related to changes in lung function indices. In nine healthy volunteers, males aged 20-27 years, acute bronchoconstriction was induced by inhalation of 1 to 128 mg/ml methacholine (M). CPAP (0.5 kPa) was then applied for 1 min. It was followed by inhalation of albuterol. Forced expiratory volume in 1 s (FEV1) and vital capacity (VC) were measured by spirometry and end expiratory lung level (EELL), to derive inspiratory capacity (IC), by inductive plethysmography. B was assessed by Borg scale. After the maximal concentration of M, FEV1 decreased by 14% (p < 0.01) as compared to the control values and Borg score (BS) increased to 2.4 (p < 0.01). In 7 out of 9 subjects we found a significant (p < 0.05) correlation between the changes in FEV1 and BS. BS decreased during CPAP (p < 0.01) and it further decreased significantly after albuterol. There was no correlation between the changes in IC and FEV1 during bronchoconstriction, or between IC and BS during CPAP. In conclusion, in healthy subjects with induced bronchoconstriction CPAP decreased significantly BS, which was further improved by inhalation of albuterol. B was related to changes in FEV1 but not in IC. PMID- 11098299 TI - Better care needed for Medicare patients, says HCFA. Collaboration will be key to success. PMID- 11098300 TI - Texas Society launches exchange program with Mexican pharmacists. PMID- 11098301 TI - Mitoxantrone receives multiple-sclerosis indication. PMID- 11098302 TI - Computerized systems can change prescribing. Evidence-based standards needed for prescribing, drug-use monitoring. PMID- 11098303 TI - Physicians care about drug costs but lack information. PMID- 11098304 TI - Studies of garlic's health benefits are inconclusive, says report. PMID- 11098305 TI - DHEA: dehydroepiandrosterone. PMID- 11098306 TI - Pricing of blood products. PMID- 11098307 TI - Pharmacology and therapeutic use of trastuzumab in breast cancer. AB - The development, pharmacology, safety, efficacy, and dosage and administration of trastuzumab are reviewed. The discovery of HER2 gene amplification in up to 30% of women with breast cancer led to the development of trastuzumab, a humanized recombinant monoclonal antibody directed against the HER2-receptor protein on breast cancer cells. In large, multicenter trials of trastuzumab as a single agent or in combination with chemotherapy as first-line or second-line therapy for metastatic breast cancer (MBC), response rates have ranged from 12% to 23% for single-agent trastuzumab and from 25% to 62% for trastuzumab plus chemotherapy. Trastuzumab increased time to disease progression and survival time when administered in combination with chemotherapy. The National Comprehensive Cancer Network guidelines for the treatment of breast cancer now include trastuzumab and paclitaxel as an option for patients with MBC or recurrent breast cancer in which the HER2-receptor protein is overexpressed. Trastuzumab is administered weekly, with an initial i.v. dose of 4 mg/kg followed by weekly doses of 2 mg/kg. Most clinical trials continued treatment until disease progression occurred. Adverse effects include infusion-related reactions manifested by fever and chills, exacerbation of chemotherapy-induced gastrointestinal toxicity and myelosuppression, and cardiotoxicity. Trastuzumab, either as a single agent or in combination with chemotherapy, can be an effective therapeutic option for MBC patients who overexpress the HER2-receptor protein and has changed the standard of care. PMID- 11098308 TI - Survey of ASHP-accredited pharmacy residency programs. AB - The results of a survey of the requirements and features of pharmacy residency programs are reported. The survey was mailed on March 5, 1999, to the directors of 414 pharmacy practice and specialty residency programs with ASHP accreditation as of March 1, 1999. Included were questions on stipends, staffing requirements, and benefits. Information not typically included in the ASHP residency directories was also sought. Three hundred surveys were returned, for a 72% response rate. Of the responding programs, 174 (58%) were pharmacy practice programs and the rest specialty programs. The specialties with the largest number of respondents were primary care (30 of 126, or 24%) and drug information (23, or 18%). The practice setting varied widely, but a university-based practice site was most frequent (122 of 300 programs, or 41%). The overall median stipend range was $28,000-$28,999; stipends varied relatively little by geographic region but were highest in New York and New Jersey. Most residency programs had a staffing requirement, which averaged eight hours per week. A substantial part of residents' time was spent on drug distribution and patient care activities, such as medical rounds and patient counseling. The most common benefits reported were paid time off, health and medical coverage, and reimbursement for specific professional expenses. Ultimately, the creation of an Internet-accessible electronic database will maximize the availability and timeliness of such information and minimize the cost and labor involved in updating currently available resources. A survey of ASHP-accredited residency programs yielded data that provide a valuable supplement to the information in ASHP residency directories. PMID- 11098309 TI - Medication sample labeling practices. PMID- 11098310 TI - Re-engineering an ambulatory care pharmacy practice. PMID- 11098311 TI - Responsible use of blood products in response to supply and demand. PMID- 11098312 TI - Implementation of a care process model for the treatment of pneumonia. PMID- 11098313 TI - Development of a best-practice model at a university hospital to increase efficiency in the management of patients with community-acquired pneumonia. AB - An economic evaluation of drug acquisition, nursing care, pharmacy time, laboratory costs, supplies, and ancillary care was conducted as a first step toward developing a pneumonia management plan at the University of Kentucky Medical Center (UKMC). UKMC costs were compared with costs at other hospitals treating pneumonia patients on Medicare. Overall costs for pneumonia at the 25% of hospitals nationwide with the lowest costs for Medicare patients with this condition were determined and compared with costs at UKMC. Against nationwide benchmarks, efficiencies at UKMC for treating simple pneumonia ranged from 45% for pharmacy expenses to 81% for nursing costs. Efficiencies for complicated pneumonia ranged from 47% for laboratory costs to 67% for nursing costs. The most cost-efficient antimicrobial treatment options were promoted and integrated into a pneumonia management plan based on Infectious Diseases Society of America treatment guidelines. A comparison of pneumonia treatment costs at UKMC with those at the 25% of hospitals nationwide with the lowest treatment costs for Medicare patients with pneumonia revealed that UKMC pharmacy costs could be optimized. Strategies for standardizing the care of patients with community acquired pneumonia (CAP) are being implemented. PMID- 11098314 TI - Experiences at a large teaching hospital with levofloxacin for the treatment of community-acquired pneumonia. AB - Costs, patient outcomes, and susceptibility patterns of selected organisms after the implementation of guidelines for the treatment of community-acquired pneumonia (CAP) at a large community teaching hospital were analyzed to assess the benefit of the guidelines. The guidelines, implemented in September 1998, included recommendations for the use of levofloxacin as the preferred antimicrobial, with rapid intravenous (i.v.) to oral (p.o.) conversion. Purchase data for levofloxacin, ceftriaxone, and azithromycin were analyzed, as well as susceptibilities and demographic and outcome data for patients admitted in 1999 in diagnosis-related groups (DRGs) 89 and 90 (simple pneumonia with and without comorbidities, respectively). Patients in each DRG were divided into a levofloxacin use only (LUO) group and an all other therapies (AOT) group. Average length of stay (LOS), hospital costs, death rate, age, and ratio of oral to intravenous dosage administration were analyzed. A total of 571 patients in DRG 89 and 110 patients in DRG 90 were included. The average LOS for DRG 89 was not significantly different between LUO patients and AOT patients (3.56 +/- 2.23 days and 3.88 +/- 2.65 days, respectively). Average total costs were significantly higher for AOT patients ($3385 +/- $2937 versus $2892 +/- $2397 for LUO patients); similar trends but no significant differences were found in the DRG 90 group. In the LUO groups in both DRGs, patients were more than five times as likely to receive an oral dosage form than patients in the AOT group. For DRG 89, the death rate was significantly lower for the LUO group (1.29%) than the AOT group (7.1%). Susceptibility data for all organisms remained stable from 1998 to 1999. The average costs in the AOT groups suggest that total hospital costs for 1999 in the LUO group were $241,516 less than costs would have been before guideline implementation. Combined drug acquisition cost savings in 1999 for levofloxacin, ceftriaxone, and azithromycin were $21,115. The use of CAP treatment guidelines was associated with reductions in antimicrobial costs, total hospitalization costs, LOS, and death rate, without a detrimental effect on organism susceptibility. PMID- 11098315 TI - Use of a physician order entry system to identify opportunities for intravenous to oral levofloxacin conversion. AB - Experience with a program for identifying, through a physician order entry system, potentially inappropriate use of expensive intravenous (i.v.) drugs, including levofloxacin, is described. The program identifies patients who are receiving i.v. levofloxacin and simultaneously receiving oral medications or an oral diet. A computerized report of such patients is printed every morning at the hospital and distributed to pharmacists according to their nursing station assignments. The pharmacists document their daily interventions to encourage switching from i.v. to oral therapy, and data are compiled and posted at the end of each month. From January 1 through September 30, 1999, the first 10 months of the program, the cost per day of levofloxacin therapy per patient decreased substantially. The program helped the institution decrease the number of patients receiving i.v. levofloxacin. PMID- 11098316 TI - Approaches to drug therapy, formulary, and pathway management in a large community hospital. AB - Use of a clinical pathway for the management of community-acquired pneumonia (CAP) at a large community hospital is described. The pathway was developed and implemented by a multidisciplinary team. Fluoroquinolone therapeutic interchange and intravenous (i.v.) to oral (p.o.) conversion were part of the pathway; through literature review, levofloxacin was chosen as the preferred quinolone. Outcomes (length of stay [LOS] and readmission rates for pathway and nonpathway patients with CAP, economic impact of the fluoroquinolone interchange protocol, and resistance patterns) were evaluated. For pathway patients in 1998, LOS was 1.2 days shorter and the readmission rate was lower. Projected drug cost savings as a result of the fluoroquinolone interchange protocol were more than $22,000 annually. Pharmacists' interventions in antimicrobial prescribing for CAP patients can lead to cost efficiency and positively affect patient outcomes. PMID- 11098317 TI - [Does information given about the anesthesia satisfy the patient?]. PMID- 11098318 TI - [Information and anesthesia: what does the patient desire?]. AB - OBJECTIVE: The aim of the study was to assess the patient's desire for information regarding their preoperative care and to assess the anaesthetists' perception of that desire. STUDY DESIGN: Questionnaire. METHODS: The question: "Would you like to be fully informed about" 13 topics of the perioperative management was asked to 106 patients at the time of the preoperative visit. Two answers were possible: Yes I want to know; No I don't want to know. 22 senior anaesthesists were also interviewed and were asked to speculate about the patients response to each item. Data were compared with those of a similar questionnaire used in different countries. RESULTS: One hundred patients who underwent general, orthopaedic, urologic surgery were interviewed. Patients sought information most frequently concerning: postoperative pain and postoperative recovery (88%), time for ambulation (83%), duration of anaesthesia (77%) and different methods of anaesthesia (77%). Only 63% patients desired to be informed about all possible complications of anaesthesia. Senior anaesthesists had a correct perception of patients desire for information about the 4 important items but not for the complications of anaesthesia. CONCLUSION: Our study suggests that an exhaustive information about anaesthesia is not wished by every patients. PMID- 11098319 TI - [The value of monitoring the bispectral index of the EEG for the management of hypertension during laparoscopic surgery]. AB - OBJECTIVE: Assessment of the usefulness of Bispectral Index of the EEG (BIS) for the management of hypertension during laparoscopic surgery. STUDY DESIGN: Preliminary, non-randomized study. PATIENTS: 15 patients undergoing laparoscopic surgery. METHODS: Anaesthesia by TCI of propofol and boluses of fentanyl in order to maintain fentanyl effect site concentration above 2 ng.mL-1 according to Scott kinetics model. Mean arterial pressure (MAP), heart rate (HR) and BIS were recorded. BIS values between 40 and 60 and MAP between 80 to 120% of preinduction values were maintained using a Gurman like decision matrix. RESULTS: MAP rose significantly after insufflation while variations of HR and BIS were not significant. DISCUSSION: During laparoscopy, factors other than pain can be responsible for hypertension. Nevertheless in clinical practice, inadequate anaesthesia should be ruled out before considering other mechanisms for hypertension. In this study, hypnosis remained adequate with a BIS under 60, and analgesia was considered sufficient as demonstrated by a good stability of the BIS despite nociceptive stimuli. This suggests that more specific haemodynamic factors were responsible for the observed rise in arterial pressure. CONCLUSION: Associating BIS monitoring and MAP in a modified Gurman decision matrix may allow more judicious therapeutic choices for hypertension during laparoscopic surgery. PMID- 11098320 TI - [Acute neurotoxic organophosphate poisoning: insecticides and chemical weapons]. AB - OBJECTIVE: To review clinical and therapeutic bases of an organophosphate poisoning, either with insecticide or nerve agent. DATA SOURCES: References were obtained from computerized bibliographic research (Medline), from personal data (academic memoir, documents under approbation of the National Defense Office), from Internet's data. DATA SYNTHESIS: Generally, organophosphate poisoning occurs during accidental exposure with agricultural insecticide or suicide. The effects of organophosphate compounds are due to the inhibition of the enzyme acetylcholinesterase. The intoxication symptoms can be divided into muscarine like, nicotine-like effects, effects on the central nervous system and symptoms related to the dysfunction of the neuromuscular junction. The interest of biological acetylcholinesterase's measuring is minimal because it is weakly specific or sensitive. The immediate severity is due to hypoxia. Respiratory failure results from the lack of central drive inflated with excessive bronchial secretions, bronchospasm and respiratory muscles paralysis. The secondary complications are early myopathies whose gravity is correlated with the decrease of acetylcholinesterases, or later neuropathies induced by a different mechanism. Beside the symptomatic measures, atropine is the specific anticholinergic treatment. When promptly used, oximes can regenerate cholinesterases. The attempted effects of the treatment are mouth dryness, pupilar dilatation and flushing of the skin. Nerve agents are lethal toxics which have a short onset time and produce severe neurological pathology. In a terrorist incident, it is as important to identify rapidly the toxic agent and provide emergency decontamination as to manage medical care. An effective response must be multidisciplinary, involving clinicians, toxicologists, Emergency Medical Service and public's health personnel. PMID- 11098321 TI - [Apical hypertrophic cardiomyopathy: a pitfall in preoperative electrocardiography]. AB - We submit two case reports of apical hypertrophic cardiomyopathy knowing that the diagnosis of one of them has been very intricate at the time of preoperative evaluation. This disease, unfrequent besides Japan, is either silent or induces cardio-vascular symptoms which are often poorly typical. The diagnosis relies on echocardiography using a high frequency probe to reveal an apical hypertrophy. In one case, a coronarography has been necessary since echocardiography failed to establish a diagnosis. Anaesthestic perioperative management should take into account the risk of apical ischaemia and the impairment of the left ventricle compliance. PMID- 11098322 TI - [Thrombus of the aortic arch: an unusual pathology in intensive care]. AB - A 54-year-old patient was admitted to the intensive care unit for voluntary drug intoxication with zolipidem (Stilnox), dimenhydrinate (Mercalm), and oestradiol 17 beta (Oromone). Four hours after the admission the patient was comatose. Cerebral computerized tomodensitometry demonstrated multiple zones of ischaemia. Transoesophageal echocardiography was performed 12 hours after the arrival of the patient and revealed a mobile thrombus of the aortic arch. The remainder of the visualized aortic arch did not present atherosclerotic plaque. Secondarily, ischaemia of the right superior limb was diagnosed probably cause by emboli originating in the aortic thrombus appeared. The patient died three days later after her arrival, because of neurologic sequelae of the cerebral embolic events. This clinical case underlines the concept that the diagnosis of drug intoxication must remain a diagnosis of elimination. The thrombosis of the aortic arch is a rare pathology in intensive care units. In the presence of unexplained ischaemic stroke and an peripheral emboli, the thrombosis of the aortic arch should be suspected. PMID- 11098323 TI - [Acute verapamil poisoning. A proposal of a strategy for therapeutic management]. AB - Verapamil poisonings are rare, but are associated with a high proportion of deaths. The authors present a case of acute poisoning by verapamil associated with collapsus and a high degree of heart block. They propose a strategy for therapeutic management based on the study of calcium-channel blocker mechanisms of toxicity. PMID- 11098324 TI - [Anesthetic management of obstetrical labor in a parturient with muscular carnitine palmitoyl transferase deficiency]. AB - We report a case of a patient with carnitine palmityl deficiency in active labour. We discuss the metabolic and energetic implications of obstetrical labour in regard with the mitochondrial myopathy and we propose an optimal management. Neuroaxial analgesia and glucose infusion are indicated in early labour because it is necessary to alleviate stress and pain in order to avoid rhabdomyolysis associated with CPT deficiency. Combined spinal epidural analgesia using intrathecal opioid alone then epidural naropein should be a relevant choice because of a minimal motor blockage. Monitoring of myolysis using serum creatinine phosphokinase levels must take in account CK and MB fractions releases to the circulation during obstetrical labour. PMID- 11098325 TI - [Prone position and severe pneumopathy in a patient with head injuries and intracranial hypertension]. AB - The treatment of hypoxaemia is one of the main goals of intensive care to patients with severe head injury. In the case reported here, the appearance of early pneumonia was accompanied by a severe deterioration of blood gases with worsening of intracranial hypertension. Prone position allowed rapid improvement of blood gases which contributed to the control of intracranial hypertension. PMID- 11098326 TI - [Acute hepatic insufficiency and exercised-induced heat stroke]. PMID- 11098327 TI - [Recommendation of the SOS ALR Group on the use of locoregional anesthesia]. PMID- 11098328 TI - [An incident during percutaneous tracheotomy using the Fantoni technic]. PMID- 11098329 TI - [The use of a CO-tester for the diagnostic elimination of collective carbon monoxide poisoning]. PMID- 11098330 TI - [Meningitis after spinal anesthesia]. PMID- 11098331 TI - [Is the systematic preanesthetic consultation unnecessary or disturbing?]. PMID- 11098332 TI - [A response to the letter from A. Benichou]. PMID- 11098333 TI - Efficacy and cost analysis of ibutilide. AB - OBJECTIVE: To determine the efficacy and safety of ibutilide in atrial fibrillation (AF) and atrial flutter (AFl) in a clinical setting and to compare the cost of first-line ibutilide with that of projected first-line electrical cardioversion (EC) from a hospital and third-party payer perspective. METHODS: Medical records of all patients (n = 60) who received ibutilide from August 1996 to March 1998 were reviewed. Efficacy was defined as successful conversion to sinus rhythm within 60 minutes of the end of the infusion, and the maintenance of sinus rhythm until hospital discharge. Safety was evaluated by determining the incidence of torsade de pointes. Charges for EC and drug administration were obtained from the hospital database and converted to costs using cost/charge ratios. Hospital costs included drug, drug administration, cardiac intensive care laboratory fee, and the cost of managing torsade de pointes. The third-party payer calculation included all of the above plus the cardiologist and anesthesiologist fees. RESULTS: Fifty percent of patients with AF or AFl were successfully converted with ibutilide; 67% of these remained in sinus rhythm at hospital discharge. Three patients experienced nonsustained torsade de pointes; all resolved with pharmacologic management. From a hospital perspective, the cost of first-line ibutilide was greater than the cost of first-line EC ($280 vs. $138 per patient). However, from a third-party payer perspective, the use of ibutilide saved approximately $324 per patient ($718 vs. $1042). CONCLUSIONS: The efficacy and safety of ibutilide in the clinical setting are consistent with data reported in clinical trials. In contrast to a previous decision analysis, ibutilide was not associated with cost savings from a hospital perspective, but was from a payer perspective. PMID- 11098334 TI - Safety and tolerability of bolus intravenous colistin in acute respiratory exacerbations in adults with cystic fibrosis. AB - OBJECTIVE: To assess the safety and tolerability of bolus intravenous doses of colistin during acute respiratory exacerbations in adults with cystic fibrosis and chronic Pseudomonas aeruginosa infection. METHODS: Twelve patients with acute exacerbations of cystic fibrosis were enrolled in a Phase I open-label study. On day 1, patients received three doses of colistin 2 mega-units (160 mg), reconstituted in 50 mL of NaCl 0.9%, by infusion over 30 minutes three times daily. On days 2, 3, and 4, the same dose of colistin was administered by bolus injection three times a day over five minutes after reconstitution in 20, 15, and 10 mL of NaCl 0.9%, respectively. The injection was given by a nurse or physician using a hand-held syringe. If the latter dose was tolerated, it was continued for the remaining eight days of the study. If any dose was not tolerated, treatment reverted to the previously tolerated concentration, which was continued throughout the remainder of the study. RESULTS: No serious adverse events occurred during the course of the trial. Patients without total indwelling venous access systems experienced mild to moderate injection pain. There were no clinically significant changes in renal function. CONCLUSIONS: This study indicates that the administration of bolus intravenous colistin as 2 mega-units (160 mg) in 10 mL of NaCl 0.9% three times a day is safe. It is well-tolerated by patients with total indwelling venous access systems. PMID- 11098335 TI - Characteristics of tranquilizer use among Australian Vietnam War veterans. AB - OBJECTIVE: To examine characteristics of tranquilizer use in a cohort of Australian Vietnam War veterans. DESIGN: Prospective analysis of medication use and assessment of social and clinical variables, including tranquilizer dependence. PATIENTS: Fifty-one Australian Vietnam War veterans were recruited from the department of psychiatry of an Australian teaching hospital. All subjects were men, with a mean +/- SD age of 52.2 +/- 3.3 years. MAIN OUTCOME MEASURES: A structured interview was used to obtain details of medical and psychiatric history, medication use, substance use, forensic history, and health service utilization data. Anxiety was assessed using the Hamilton Anxiety Rating Scale (Ham-A). A validated tranquilizer dependence rating scale was administered for each patient. RESULTS: Commonly used tranquilizers included diazepam (n = 19 patients) and zopiclone (26). Most patients (44) reported the use of one or more drugs for the purpose of nighttime sedation, while exclusive daytime use of tranquilizers for anxiolytic effect was uncommon. The median time spent in the hospital during the preceding year was 21.0 +/- 56.8 days. Symptoms of anxiety were prevalent, with a mean Ham-A score of 35.5 +/- 7.8. Screening criteria suggestive of tranquilizer dependence were met in 34 subjects. Health service utilization was correlated with tranquilizer intake and overall medication use. Tranquilizer dependence was independently associated with cigarette smoking (p = 0.039; odds ratio = 5.13, 95% CI 1.08 to 24.33). CONCLUSIONS: This study provides insight into the nature of tranquilizer use in an Australian population of Vietnam War veterans. The extensive use of these drugs suggests that further research and possibly intervention in this area is needed. PMID- 11098336 TI - Pharmacists' understanding of patient education on metered-dose inhaler technique. AB - OBJECTIVE: To assess pharmacists' attitudes, beliefs, and knowledge about assessing and educating patients regarding metered-dose inhaler (MDI) technique; to determine frequency of MDI assessment and teaching behavior; and to assess the effect of an asthma pharmaceutical care educational program on the same variables six months later. DESIGN: Questionnaire completed before and six months after the educational intervention. SETTING: Pharmacies based in clinics owned by a healthcare system located in communities of a large metropolitan area. MAIN OUTCOMES MEASURES: Pharmacists' self-reported frequency of assessment and education; attitudes and beliefs about assessing and educating patients using MDIs; and knowledge of MDI technique. RESULTS: The survey response rate was 53.7% (n = 39) for baseline and 43% (n = 32) for follow-up. Most pharmacists (85.4% at baseline, 87.5% at follow-up; p = 0.79) indicated that they educate patients receiving new MDI prescriptions. In addition, 47.4% at baseline and 68.8% at follow-up indicated they educate patients using inhalers for three months (p = 0.07). Only 21.1% at baseline and 18.8% at follow-up (p = 0.81) indicated that they follow up with long-term users. The mean +/- SD MDI technique knowledge score (steps correct out of 9 possible) at baseline was 7.2 +/- 1.1 and 7.5 +/- 1.3 at follow-up (p = 0.29). Significant changes in level of agreement with some beliefs/attitudes were observed, including the importance of frequently assessing/educating patients, confidence and comfort when assessing/educating patients, and that assessing/educating patients is not the role of only the physician. Respondents continued to acknowledge that MDI education and assessment are important to improving and maintaining control of disease. However, the respondents thought that barriers exist that inhibit this activity, such as not enough time for education and assessment. CONCLUSIONS: Pharmacists reported they frequently educate patients and assess MDI technique for new prescriptions but not very often for patients recently started, as well as for long-term users. Six months after an educational program, attitudes and beliefs toward this activity were either not changed or, in some, improved. Pharmacists perceive that there is not enough time to assess and educate patients who use MDIs. PMID- 11098337 TI - Stability of filgrastim and epoetin alfa in a system designed for enteral administration in neonates. AB - OBJECTIVE: To determine the stability of recombinant granulocyte colony stimulating factor (rG-CSF, filgrastim) and recombinant erythropoietin (rEpo, epoetin alfa) in a solution designed for enteral administration in the neonatal intensive care unit. DESIGN: Filgrastim and epoetin alfa were added to a solution with NaCl 0.9%, sodium acetate, potassium chloride, and human albumin in concentrations designed to mimic human amniotic fluid. Additionally, the solution was dripped through polyvinyl chloride feeding tubes to simulate feedings, and aliquots were collected before, during, and after priming of the tube. Other aliquots were either frozen immediately, stored at room temperature, or refrigerated for 0, 6, 12, 18, and 24 hours. MAIN OUTCOME MEASURES: Filgrastim and epoetin alfa concentrations in the various aliquots were compared with the concentrations in the original solution. RESULTS: Filgrastim and epoetin alfa concentrations were stable for at least 24 hours when refrigerated and for at least three weeks when frozen. At room temperature, filgrastim was stable for 18 hours and epoetin alfa for 24 hours. Filgrastim concentrations did not vary significantly before, during, or after priming of the feeding tube, whereas epoetin alfa concentrations decreased significantly unless the feeding tube was primed with 10 mL of solution. CONCLUSIONS: Filgrastim and epoetin alfa were stable in our amniotic fluid-like solution. In this respect, our solution is suitable for enteral administration to patients in the neonatal intensive care unit. PMID- 11098338 TI - Treatment of affective disorder and obesity with topiramate. AB - OBJECTIVE: To report a case of weight loss and mood stabilization in a patient being treated with the antiepileptic drug topiramate. CASE SUMMARY: A 37-year-old obese white woman with affective instability and obesity was being treated with adjunctive topiramate therapy. The patient lost 10 kg over 10 weeks of treatment with topiramate and improved clinically, as evidenced by a reduction in the number of times that she had to be admitted to a management unit for constant observation, and a decrease in the number of times that mechanical restraints or medication interventions were required for aggressive outbursts. Furthermore, the patient successfully completed two home visits while receiving topiramate therapy and was out of the hospital on her third home visit at the time of this writing. DISCUSSION: This case further strengthens previous reports that topiramate may be useful in treating affective disorders as well as inducing weight loss in a patient population in which weight gain is common. The patient discussed in this case report had no acute illnesses or changes in health status, no changes in diet, and no changes in her medications that could have accounted for the sudden weight loss. In addition, the patient's behavior did not improve until topiramate was added as adjunctive therapy of valproic acid, citalopram, and chlorpromazine during an adequate trial period. CONCLUSIONS: Controlled studies need to be performed to evaluate the use of topiramate in the psychiatric population and, in particular, the benefits of topiramate therapy in psychiatric patients with an additional diagnosis of obesity. PMID- 11098339 TI - Warfarin resistance due to sulfasalazine. AB - OBJECTIVE: To report a case of warfarin resistance associated with the use of sulfasalazine. CASE SUMMARY: A 37-year-old white woman on oral anticoagulant therapy with warfarin was being evaluated for complaints of joint pains. Her past medical history consisted of recurrent deep-vein thrombosis, asthma, and ulcerative colitis. Warfarin concentrations had consistently remained within the therapeutic range with dosages of approximately 30 mg per week. In an attempt to treat arthritis, her gastroenterologist replaced 5-aminosalicylic acid (5-ASA) with sulfasalazine 1000 mg four times daily. Subsequent to the medication changes, the international normalized ratio (INR) decreased and remained at subtherapeutic concentrations despite increases in the warfarin dosage. During this period, the patient developed a deep-vein thrombosis in the right popliteal vein. The INR did not return to an acceptable level until six weeks after sulfasalazine was started. The new warfarin dosage was 75 mg per week, a 250% dosage increase. When sulfasalazine was discontinued and 5-ASA reinstituted, the warfarin dosage requirements to achieve therapeutic INRs returned to weekly dosages of approximately 45 mg. DISCUSSION: Sulfonamides have been well documented to enhance the anticoagulant effect of warfarin. This patient developed a new deep-vein thrombosis due to failure in maintaining therapeutic INR levels after the recent introduction of sulfasalazine. We suspect that she developed warfarin resistance secondary to concomitant use of sulfasalazine. This patient demonstrated warfarin resistance as opposed to enhanced anticoagulant effect with sulfasalazine. CONCLUSIONS: Clinicians managing warfarin therapy should exercise caution when sulfasalazine is added to a patient's medical regimen. This case suggests a possible warfarin-sulfasalazine interaction that resulted in warfarin resistance. PMID- 11098340 TI - Drowsiness and poor feeding in a breast-fed infant: association with nefazodone and its metabolites. AB - OBJECTIVE: To investigate whether adverse effects in a premature neonate could be attributed to nefazodone exposure via breast milk. CASE SUMMARY: The breast-fed white infant (female, 2.1 kg, 36 weeks corrected gestational age) of a 35-year old woman (60 kg) taking nefazodone 300 mg/d was admitted to the hospital because she was drowsy, lethargic, unable to maintain normal body temperature, and was feeding poorly. A diagnosis of exposure to nefazodone via breast milk was considered only after other more likely diagnoses had been excluded. After breast feeding was discontinued, the infant's symptoms resolved slowly over a period of 72 hours. The maternal plasma and milk concentration-time profiles for nefazodone and its metabolites, triazoledione, HO-nefazodone, and m-chlorphenylpiperazine, were quantified by HPLC. The calculated infant dose for nefazodone and its active metabolites (as nefazodone equivalents) via the milk was only 0.45% of the weight adjusted maternal nefazodone daily dose. DISCUSSION: Our data suggest a putative association between maternal nefazodone ingestion and adverse effects in a premature breast-fed neonate. The measured amount of drug exposure would normally be considered safe in a full-term infant. However, there was a temporal relationship between resolution of adverse effects in the infant and cessation of breastfeeding. CONCLUSIONS: This case highlights the importance of individualizing the risk-benefit analysis for exposure to antidepressants in breast milk, especially when dealing with premature neonates. PMID- 11098341 TI - Toxic metronidazole-induced MRI changes. AB - OBJECTIVE: To report a case of changes documented by magnetic resonance imaging (MRI) of the head probably due to accumulation of metronidazole in a patient with liver dysfunction. CASE SUMMARY: A 34-year-old Hispanic man with cirrhosis and hepatitis C being treated with metronidazole for Bacteroides fragilis meningitis and bacteremia developed ataxia, disorientation, and peripheral neuropathy. An MRI at the time meningitis was diagnosed was negative. After the patient received > 60 g of metronidazole, an MRI revealed increased signal intensity below, behind, and lateral to the fourth ventricle. Concomitant metronidazole serum concentration was toxic at 35.1 micrograms/mL. DISCUSSION: This is the second reported case of metronidazole-induced MRI changes. Metronidazole is known to accumulate in patients with liver dysfunction and can cause peripheral neuropathy and central nervous system (CNS) dysfunction; these effects may take up to two years to completely resolve. CONCLUSIONS: Metronidazole dosages should be reduced in patients with liver dysfunction to prevent the accumulation of metronidazole, which can lead to CNS dysfunction and peripheral neuropathy. PMID- 11098342 TI - Seizure possibly associated with fluvoxamine. AB - OBJECTIVE: To inform clinicians of the possibility that seizures due to therapeutic doses of fluvoxamine may not be as rare as previously considered. CASE SUMMARY: A 49-year-old white man with schizoaffective disorder and a past history of seizures secondary to head trauma had been seizure-free for approximately 10 years. Fluvoxamine therapy was begun due to increasing obsessive compulsive behavior. Despite receiving anticonvulsants for his mood disorder, the patient had a breakthrough seizure. There were no underlying medical conditions that might have induced this seizure. No further seizures occurred after he was placed on a higher dosage of the anticonvulsants. The obsessive-compulsive behavior improved considerably as a result of fluvoxamine treatment. DISCUSSION: The patient presented here developed a seizure with a therapeutic dosage of fluvoxamine; seizures associated with this agent have occurred more often with overdose. Multiple factors such as a prior history of seizures, head trauma, and concurrent treatment with other psychotropic agents are considered in this case report. CONCLUSIONS: Despite the relatively safe and benign adverse effect profile of the selective serotonin-reuptake inhibitors such as fluvoxamine, clinicians should be cautious about seizures as an adverse effect, especially when the patient has even a remote history of seizure or head trauma. PMID- 11098343 TI - Fatal systemic reaction after multiple doses of intravesical bacillus Calmette Guerin for polyposis. AB - OBJECTIVE: To report a case of fatal systemic reaction after intravesical administrations of bacillus Calmette-Guerin (BCG) for polyposis. CASE SUMMARY: A 72-year-old white man was treated by monthly injections of intravesical BCG immunotherapy for polyposis of the urinary bladder. He received a total of eight injections; four hours after the seventh injection, he presented with pyrexia associated with chills, sweating, headache, and vomiting, which quickly resolved. Four hours after the eighth injection, the patient presented with the same symptoms plus a left-hemisphere deficiency. Results of a cerebral scan performed at this time were normal. The clinical status of the patient quickly worsened, with the appearance of disseminated intravascular coagulation, acute anuric renal insufficiency, rhabdomyolysis, hemolysis, and cytolytic and cholestatic hepatitis. The patient required hemodialysis and symptomatic treatment. Lactic acidosis with hemolytic-uremic syndrome appeared, and he died as the result of a multivisceral (respiratory, renal, hepatic) deficiency. DISCUSSION: The patient presented with symptoms compatible with a severe systemic reaction to BCG therapy, a rare but possible adverse effect. CONCLUSIONS: BCG instillation is a valuable tool in the therapy of bladder carcinoma, but increasing reports of severe adverse reactions should continue to remind practicing urologists to be alert to the possibility of common and uncommon reactions after its use. PMID- 11098344 TI - Aggrenox: a fixed-dose combination of aspirin and dipyridamole. AB - OBJECTIVE: To describe the pharmacology, pharmacokinetics, efficacy, and safety of a fixed-dose combination of aspirin and extended-release (ER) dipyridamole indicated for the secondary prevention of stroke. DATA SOURCES: Published articles and abstracts were identified from a MEDLINE search (1966-December 1999) using the search terms dipyridamole, aspirin, antiplatelet, antiaggregation, and stroke prevention. Pertinent articles written in English were considered for review. Additional articles were identified from the references of retrieved literature. STUDY SELECTION AND DATA EXTRACTION: Studies including a combination of aspirin/dipyridamole in human subjects were evaluated. Emphasis was placed on randomized, controlled trials. DATA SYNTHESIS: Aspirin is a platelet inhibitor that works by inhibiting platelet cyclooxygenase, which reduces the production of thromboxane A2. Dipyridamole is a platelet inhibitor that is thought to work in part by inhibiting platelet cyclic-3',5'-adenosine monophosphate and cyclic-3',5' guanosine monophosphate phosphodiesterase. The active metabolite of aspirin, salicylic acid, is highly bound to plasma protein and has a plasma half-life of two to three hours. Dipyridamole is also highly bound to plasma proteins, and the ER formulation has a plasma half-life of 13 hours. The first European Stroke Prevention Study (ESPS-1) found the combination of aspirin/dipyridamole to be superior to placebo in the prevention of stroke and transient ischemic attack (TIA). The ESPS-1, however, did not include an aspirin-only treatment arm. Therefore, it was unclear whether the combination of aspirin/dipyridamole was superior to aspirin alone. As a result, a second trial was conducted that included treatment arms of aspirin alone, ER dipyridamole alone, combination therapy, and placebo. The combination of aspirin 25 mg plus ER dipyridamole 200 mg twice daily was shown in the ESPS-2 to be significantly better than either agent given individually in preventing stroke and TIAs (p < 0.001). CONCLUSIONS: The American College of Chest Physicians (ACCP) recommends aspirin 50-325 mg/d to be the initial antiplatelet of choice for the prevention of atherothrombotic cerebral ischemic events. However, with the favorable results of the ESPS-2, it may be appropriate to substitute aspirin/ER dipyridamole for aspirin alone as the drug of choice. This combination appears to have a favorable adverse effect profile. The relative effectiveness of aspirin/ER dipyridamole compared with clopidogrel and ticlopidine has yet to be determined. If alternative antiplatelet therapy is needed, the ACCP recommends clopidogrel rather than ticlopidine because of its lower incidence of adverse effects. The ACCP further states that the combination of aspirin plus dipyridamole may be more effective than clopidogrel; these agents have a similarly favorable adverse effect profile. PMID- 11098345 TI - Miglitol: assessment of its role in the treatment of patients with diabetes mellitus. AB - OBJECTIVE: To evaluate miglitol, a new oral alpha-glucosidase inhibitor, and discuss its pharmacology, therapeutics, pharmacokinetics, dosing guidelines, adverse effects, drug interactions, and clinical efficacy. DATA SOURCES: A MEDLINE English-language only database search using the keywords miglitol, glyset, and Bay m 1099 (1985 to December 1999), was completed to identify relevant articles including reviews, recent studies, and abstracts; American Diabetes Association 1999 Annual Meeting abstracts; Pharmacia & Upjohn data on file and product information. STUDY SELECTION: The clinical trials that were selected to be reviewed in detail were randomized, double-blind studies with at least 100 patients in the intention-to-treat group. DATA EXTRACTION: All articles and abstracts were reviewed along with the product labeling from Pharmacia & Upjohn. DATA SYNTHESIS: Miglitol is an alpha-glucosidase inhibitor that exerts its effect through the delayed absorption of complex carbohydrates in the small intestine, resulting in a decrease in postprandial glucose concentrations that are directly correlated with the dietary carbohydrate content. Both small, short term trials and large, clinical trials show a decrease in postprandial glucose concentrations and a modest decrease in glycosylated hemoglobin of approximately 0.5-1.0% as a result of miglitol's action. The adverse effects of miglitol are mild and transitory and include flatulence, diarrhea, and abdominal pain. The incidence of gastrointestinal problems may be reduced with a small initial dose, which is slowly titrated as tolerated. CONCLUSIONS: Miglitol is an effective and safe treatment option in patients with type 2 diabetes mellitus who are inadequately controlled with diet or oral sulfonylurea therapy. Miglitol is a good choice of therapy in Hispanic, African-American, and elderly patients, or any patients in whom hypoglycemia, weight gain, or lactic acidosis are risks. No published studies comparing miglitol with acarbose have been published, but there appears to be no major clinical or financial advantages to using one agent over the other. PMID- 11098346 TI - Reboxetine: a selective norepinephrine reuptake inhibitor for the treatment of depression. AB - OBJECTIVE: To review the pharmacology, pharmacokinetics, efficacy, and tolerability of reboxetine in the treatment of major depressive illness. DATA SOURCES: A MEDLINE search restricted to English-language literature was conducted (1966 to July 1999). Abstracts and posters presented at meetings were also reviewed. DATA EXTRACTION/STUDY SELECTION: Studies currently available were conducted in Europe. Data from clinical trials were reviewed to gather information specific to efficacy analysis, pharmacokinetic parameters, tolerability profiles, and drug-drug interactions. Information on reboxetine was compared with other antidepressant therapies when appropriate data were available. DATA SYNTHESIS: Reboxetine is a selective norepinephrine reuptake inhibitor shown to be an effective agent in the treatment of major depressive illness. In clinical trials, reboxetine was effective in decreasing mean total scores of the Hamilton Rating Scale for Depression in adult populations. Improvements were similar between reboxetine and desipramine and imipramine, as well as fluoxetine. Reboxetine is relatively well tolerated, with insomnia, sweating, constipation, and dry mouth being commonly reported adverse events. Hypotension and urinary hesitancy occur at lower rates than with the tricyclics, and compared with fluoxetine, reboxetine is associated with lower rates of nausea, somnolence, and diarrhea. Dosage adjustments may be appropriate in elderly patients and those with renal and hepatic impairment. CONCLUSIONS: Reboxetine has received two letters of approval from the Food and Drug Administration for the treatment of major depression in adults. Upon completion of an additional US clinical study, reboxetine is likely to become a first-line agent in the management of depressive illness and a promising alternative for patients who have failed treatment with or do not tolerate serotonergic antidepressants. PMID- 11098347 TI - A chronobiologic approach to the pharmacotherapy of hypertension and angina. AB - OBJECTIVE: To review the chronobiology of hypertension and coronary artery disease and the application of chronotherapeutics to their treatment and prevention. DATA SOURCES: Clinical trials and review articles (English-language) on the topic of chronotherapy and cardiovascular disease were identified via a MEDLINE search from 1990 to March 2000, using the search terms chronotherapy, circadian rhythm, cardiovascular disease, hypertension, and angina. DATA EXTRACTION: Search and evaluation focused on published clinical trials and review articles of circadian variation associated with pharmacotherapy for cardiovascular disease. DATA SYNTHESIS: The existence of circadian rhythm in cardiovascular disease is well established. Heart rate and blood pressure peak during the morning hours and reach a nadir at bedtime. The incidence of myocardial infarction, stroke, sudden cardiac death, and myocardial ischemia also increases during the early-morning hours. Based on these relationships, researchers have begun to apply the science of chronotherapeutics, or timing of drug effect with biologic need, to improve cardiovascular outcomes. This includes administering traditional agents at specific times throughout the day and developing new agents--chronotherapeutic formulations with special release mechanisms--targeted at inducing the greatest effect during the morning surges. Chronotherapeutic agents are specifically designed to provide peak plasma concentrations during the early-morning hours, when effect appears most needed; lowest concentrations occur at night, when heart rate and blood pressure are lowest and, consequently, cardiovascular events are least likely to occur. CONCLUSIONS: Whether chronotherapy of cardiovascular disease offers an advantage in long-term outcomes over traditional therapy must be studied in clinical trials. PMID- 11098348 TI - Impact of pharmacists providing a prescription review and monitoring service in ambulatory care or community practice. AB - OBJECTIVE: To systematically review the impact, on patient outcomes and costs to the healthcare system, of pharmacists reviewing and monitoring prescribing in ambulatory care or community practice. DATA SOURCES: We conducted a systematic search of published literature, up to and including 1998, on outcomes of prescription monitoring and review by pharmacists. Thirteen electronic databases were reviewed, along with a hand search of 11 journals known to publish pharmacy practice research. Fifty-five articles describing 50 comparative studies were identified. DATA EXTRACTION: Data were extracted including study design, clinical site, and results. A qualitative synthesis of the findings was conducted and methodological quality was appraised. DATA SYNTHESIS: Pharmacist-run services may be accompanied by improvements in clinical outcomes. Inconsistent definitions used in the research evaluated meant that an overall interpretation of a change in the incidence of compliance and adverse drug reactions was impossible. Other outcomes such as knowledge and satisfaction showed equivocal results overall. There was little or no change in quality of life where this was assessed. Savings in drug acquisition costs may have accrued, but it was impossible to calculate the magnitude. Pharmacist involvement produced a positive impact on cost-benefit and cost-effectiveness. CONCLUSIONS: The heterogeneity of the studies found and the variety in quality of much of the research design prevent the rigorous assessment of the direction and magnitude of any changes reviewed. Further studies are required, which must be rigorously designed, with blind and independent assessment of clearly defined outcomes. In particular, there is a need to investigate the effect of such services on the incidence of adverse drug reactions and quality of life and to conduct robust cost-benefit analyses. PMID- 11098349 TI - Docetaxel as an alternative to paclitaxel after acute hypersensitivity reactions. AB - OBJECTIVE: To review the mechanism involved in paclitaxel-induced hypersensitivity reactions and to evaluate the potential use of docetaxel after acute hypersensitivity reactions (HSRs) to paclitaxel. DATA SOURCES: Literature identified through a MEDLINE search (1966-September 2000) and through secondary sources. DATA SYNTHESIS: HSRs to paclitaxel can be life-threatening. The exact etiology involved in paclitaxel-induced HSRs has not been fully elucidated; the reactions may be due to the Cremophor EL vehicle or to paclitaxel itself. Options for treatment following HSRs are limited. A rechallenge attempt can be made, but is not always successful. Docetaxel, a semisynthetic taxane, may have a role in therapy for patients unable to tolerate paclitaxel therapy. This review examines the etiology of paclitaxel-induced HSRs and the potential role of docetaxel following these acute reactions. CONCLUSIONS: Docetaxel may be a viable alternative for patients who experience HSRs to paclitaxel. PMID- 11098350 TI - Spironolactone in the treatment of congestive heart failure. AB - OBJECTIVE: To evaluate evidence supporting the use of spironolactone in managing congestive heart failure. DATA SOURCES: Literature accessed through MEDLINE (January 1966-December 1999) and cross-referencing of selected articles. Search terms included spironolactone and heart failure. DATA SYNTHESIS: Heart failure is a leading cause of morbidity and mortality. Through aldosterone antagonism, spironolactone may be an effective pharmacotherapeutic addition to patients not responding to standard drug therapy for heart failure. RESULTS: Clinical trials have demonstrated that, in patients with heart failure, spironolactone improves laboratory indices, quality of life, and morbidity. Recently, spironolactone has been demonstrated to improve the survival of patients with New York Heart Association (NYHA) III or IV heart failure. CONCLUSIONS: Spironolactone use should be considered in patients with NYHA Class III or IV heart failure. PMID- 11098351 TI - Direct-to-consumer advertising. AB - These issues will be debated and should be, even as DTC advertising of prescription drugs becomes further entrenched. I hope that physicians will see DTC advertising not as a nuisance, but rather as an opportunity to enhance health care by enabling more people to understand diseases and treatments, treat more broadly diseases that now are underdiagnosed or undertreated, and become better communicators and deliver more caring medical attention to their patients. PMID- 11098352 TI - Abnormal movements with donepezil in Alzheimer disease. PMID- 11098353 TI - Metrorrhagia and ginseng. PMID- 11098354 TI - Comment: potential risk of valproic acid therapy in patients who are HIV positive. PMID- 11098355 TI - The importance of international initiatives. PMID- 11098356 TI - Stop smoke campaign begins with you. PMID- 11098357 TI - Team dynamics. PMID- 11098359 TI - More basic procedures PMID- 11098358 TI - CBPN criteria. PMID- 11098360 TI - Clinical nurse specialists. PMID- 11098361 TI - Herbs and the perioperative patient. AB - Herbs are claimed to cure or correct many ills, and in the United States, they are classified as dietary supplements. The Federal Trade Commission, rather than the US Food and Drug Administration, has primary responsibility for dietary supplements, so companies may make claims about herbs that are unsubstantiated by research. Perioperative nurses should be aware of herb usage, interactions with other traditional medications, and herbs effects on perioperative patients. PMID- 11098362 TI - "Don't destroy the evidence!". AB - As nurses, we interact with both victims and perpetrators of violence. Our goal is to save the patient; however, we also play a role in the legal outcome of that violence. What we do in caring for these patients is important. Means of preserving and documenting evidence while caring for the victims of violence are discussed and guidelines for the nurses legal role are given. PMID- 11098363 TI - Minimizing preoperative anxiety with alternative caring-healing therapies. AB - This article reviews holistic caring-healing therapies that may decrease preoperative anxiety for the surgical patient, based on the philosophy and science of caring developed by Jean Watson, RN, PhD, FAAN. Dr Watson reveals a new paradigm emerging in health care that blends the compassion and caring of nursing in harmony with the curative therapies of medicine. Hypnosis, aromatherapy, music, guided imagery, and massage are integrative caring-healing therapies that may minimize preoperative anxiety. Alternative therapies offer a high-touch balance when integrated with high-tech conventional surgical treatments. PMID- 11098364 TI - Challenges of measuring intraoperative patient outcomes. AB - The Perioperative Patient Focused Model incorporates AORN s patient outcomes standards of perioperative care. Although the outcomes were designed for perioperative practice, it is difficult to judge which outcome standards an intraoperative nurse might appropriately use to determine if the patient has achieve the desired result at the time of discharge from the OR. The purpose of this article is to explore whether intraoperative nurses can implement the new model by substituting intermediate patient outcomes to evaluate the effect of nursing care delivered during a surgical procedure. PMID- 11098365 TI - Redesign of medication and supply distribution in a day surgery center. AB - The day surgery center at NYU Hospitals Center installed an automated medication and supply distribution system as part of its facility renovation. A constricted Manhattan site prohibited physical expansion, so space efficiency became top priority. Goals also included increasing revenue, positioning for managed care contracting, improving access to supplies and medications, use tracking and inventory adjustment, documenting and tracking, accurate charging, and enhancing patient and staff member satisfaction. Space efficiencies achieved via automated distribution enabled planners to save one OR, which generates approximately $1 million in revenue annually. Demonstrated quarterly cost savings total $35,615, for an expected $142,460 in annual savings. The number of full-time employees did not increase from the amount budgeted for the prior system in any department participating in implementing the new system. PMID- 11098366 TI - Perioperative documentation in Finland. AB - In Finland, research studies about perioperative documentation are few, and there are no professional recommendations for perioperative documentation, such as AORN s Standards, Recommended Practices, and Guidelines. Exploring current documentation practices and contents used in Finland is the first step to establishing a standard for perioperative documentation. The need for this type of exploration resulted in a study that found that the aim of nursing documentation is not always clear, and current documentation practice does not necessarily reveal the decision making that directs patient care, demonstrate nursing resources needed, or provide data for evaluating and developing perioperative practice. Education, motivation, and computerization generally were mentioned as a means to develop documentation. PMID- 11098367 TI - A phenomenologic study of how perioperative nurses perceive their work world. AB - As health care facilities restructure and downsize, many are evaluating the value of RNs as members of the health care team. While many studies have been done about how health care systems measure value, no studies have been found that focus on areas of concern from the perspective of nurses who actually work in the OR. This article details the results of a phenomenologic study that used one-on one interviews to ask five perioperative nurses how they perceive their work world. By telling the nurses stories, five essential themes of being a nurse in the perioperative work world emerged. PMID- 11098368 TI - Evaluating evidence found on the Internet. PMID- 11098369 TI - Developing a nurse-to-patient ratio policy. PMID- 11098370 TI - AVJ: lying author exposed. PMID- 11098371 TI - Plan for projectile dart restraint. PMID- 11098372 TI - Pity the poor animals--save them from our foibles. PMID- 11098373 TI - Cages criticised. PMID- 11098375 TI - AVA, ACAC and PIAS. PMID- 11098374 TI - AVA and welfare. PMID- 11098376 TI - No intrigue. PMID- 11098377 TI - Lonsdale 'unreasonable'. PMID- 11098378 TI - 'Unbalanced diet'. PMID- 11098379 TI - Policy needs review. PMID- 11098380 TI - Hemihyperaesthesia and hyperresponsiveness resembling central pain syndrome in a dog with a forebrain oligodendroglioma. AB - A 4-year-old male Boxer was presented with neurological signs referable to a right forebrain lesion that was confirmed with computed tomography. Whilst characteristic signs of a unilateral forebrain lesion were observed, the dominant and striking finding was a right-sided hemisensory disturbance characterised by hyperaesthesia and hyperresponsiveness. Necropsy revealed a gelatinous mass confined to the right forebrain that was identified histologically as an oligodendroglioma. The lesion was centred on the internal capsule and involved ventral frontal and temporal lobes and the ventrolateral thalamus, including lateral and medial parts of the ventrocaudal nuclear region (ventrobasilar complex) of the thalamus. On clinical and neuroanatomical grounds, the case exhibited features in common with central pain syndrome in human patients with thalamic lesions. These included a somatosensory disorder of hyperaesthesia affecting an entire side of the head and body, behavioural manifestations consistent with spontaneous pain and a lesion involving the ventrobasilar complex. Of interest, the hemisensory abnormality was ipsilateral to the lesion, contrasting with central pain in humans, in which clinical signs are contralateral to analogous lesions. It is suggested that species-specific differences in spinal cord organisation of pain pathways, particularly the greater bilateral projection of nociceptive afferents to thalamic relay nuclei in carnivores, may account for this disparity. Notably, central pain is rare in human patients with brain tumours, even those affecting the thalamus, and this may also be the case in dogs. PMID- 11098381 TI - Clinical findings associated with chronic ischial fracture in a gelding. AB - An 8-year-old show-jumper gelding was referred for examination as a result of a purchase dispute for reported back pain. Clinical examination identified back pain and atrophy of the left semimembranosus and semitendinosus muscles, but no lameness. Standing pelvic radiography demonstrated a chronic nonunion fracture of the left ischium, the clinical significance of which was uncertain. The apparent back pain was thought to be probably unrelated to the pelvic lesion. We conclude that chronic ischial fracture in the horse can lead to specific atrophy of the semimembranosus and semitendinosus muscles, which originate from this bone. PMID- 11098382 TI - Surgical treatment of tarsometatarsal joint luxation in a miniature horse foal. AB - A 2-week-old Miniature Horse foal was referred for evaluation and treatment of a luxated right tarsometatarsal joint. Treatment consisted of closed reduction and internal fixation using two partially threaded Steinmann pins placed in normograde fashion through the tuber calcis into the proximal third metatarsus. Traumatic luxation has been reported to occur in the tarsocrural, proximal intertarsal and tarsometatarsal joints within the equine tarsus. Treatment for luxation of the distal intertarsal joint has not been documented. The treatment method most commonly suggested for tarsal luxation is closed reduction and cast immobilisation. Internal fixation using lag screws and plating has also been described. A combination of internal fixation and external coaptation is thought to achieve maximal stability and allow faster convalescence in cases of tarsal luxation. This case report describes for the first time a technique using two Steinmann pins to achieve successful internal fixation of a traumatic tarsometatarsal joint luxation in a 2-week-old Miniature Horse foal. PMID- 11098384 TI - Transient hypoparathyroidism following successful treatment of hypercalcaemia of malignancy in a dog. AB - A 4-year-old, entire female, English Cocker Spaniel was presented for treatment of lymphosarcoma and secondary hypercalcaemia. After induction chemotherapy the dog became severely hypocalcaemic and showed signs of weakness, muscle fasciculation and facial pruritus. Hormone analysis confirmed inadequate production of parathyroid hormone. Although hypocalcaemia has been previously reported as a component of tumour lysis syndrome, it has not been associated with transient parathyroid hormone deficiency. PMID- 11098383 TI - Mycobacterium genavense infection in two aged ferrets with conjunctival lesions. AB - Mycobacterium genavense infection was diagnosed in two adult ferrets. Disseminated mycobacteriosis was diagnosed in a castrated 5-year-old sable ferret with generalised peripheral lymph node enlargement and a proliferative lesion of the conjunctiva of the nictitating membrane. The diagnosis was based on characteristic cytology and sequence analysis of the 16S rRNA gene amplified using the polymerase chain reaction from fresh biopsy material. Therapy with rifampicin, clofazimine and clarithromycin probably cured the infection. An entire 4-year-old female ferret with conjunctival swelling, serous ocular discharge and swelling of the subcutaneous tissues of the nasal bridge was diagnosed as having M genavense infection on the basis of typical cytology, histopathology and sequence analysis of 16S rRNA amplicons from formalin-fixed paraffin-embedded tissue. This patient was treated successfully using rifampicin. Both ferrets subsequently died as a result of other disease conditions, 10 and 4 months following initiation of therapy, respectively. This is the first report documenting M genavense as a cause of disseminated mycobacterial disease in ferrets. Conjunctival involvement may be a feature of disseminated mycobacteriosis in the ferret. The possibility that these infections were the consequence of a ferret retrovirus infection should be considered further. PMID- 11098385 TI - Cloning and transgenesis in farm animals--an Australian perspective. PMID- 11098386 TI - Trichinellosis in Papua New Guinea. AB - OBJECTIVES: To describe the discovery in a domestic pig of the first case of trichinellosis in Papua New Guinea, caused by a new taxon within the genus Trichinella (T papuae). Also, to establish if the disease occurred in the local wild pig population and in domestic pigs elsewhere in the country, and to test if the worm was infective to some other animals. PROCEDURE: Fresh and fixed tissue samples were examined by the digestion method and histologically, respectively, for the non-encapsulated larvae of T papuae. Feeding trials were conducted, using infected tissues and infective larvae, on animals under laboratory conditions. RESULTS: About 8.8% of a wild pig population in Western Province, adjacent to Irian Jaya, Indonesia, was found to be infected. Infection was not found in other local and feral animals or in domestic pigs from other parts of the country. Infection was experimentally established in cats, pigs and laboratory bred mice and rats. CONCLUSION: Trichinellosis is confined to one remote locality in PNG. Domestic pigs in the initial case became infected, probably, by eating infected wild pig meat. PMID- 11098387 TI - The role of lectures in veterinary education. AB - OBJECTIVE: To study the role of lectures from the perspective of staff and students involved in the veterinary course at The University of Queensland. METHODS: The Nominal Group Technique of Delbecq et al, which provides the maximum opportunity for group members to put forward points, was used to help develop a questionnaire which was completed by 351 students (a response rate of 84%) and 35 staff (76%) from the five years of the veterinary course, and was analysed using the SAS System for Windows. RESULTS: Almost all the staff and students agreed that lectures should fulfil many roles including stimulating and motivating students and encouraging them to think, as well as presenting ideas and concepts and an indication of the structure and relevance of the material. They should provide a guide for effective deep learning, but not encourage rote (or superficial) learning. A smaller percentage of staff and even fewer students agreed that lectures did fulfil these roles, especially those directed at encouraging students to look beyond simple memorisation of facts. The perceived disparity between reality and the ideal was greater amongst the older, clinical students than amongst their more junior colleagues. CONCLUSIONS: The focus of attention in lectures needs to change from the superficial, rote learning of information to deep, active learning directed at using information to solve problems that are perceived by the students to be relevant. If done in a stimulating and interesting way, this should develop skills in reasoning and critical analysis as well as providing a framework for storage and recall. It should also increase the motivation towards learning both during the veterinary course, and over the professional lifetime. Furthermore, the place of the lecture in veterinary education needs to be reassessed regularly in the face of newly emerging educational technology. PMID- 11098388 TI - Evaluation of a laboratory test to detect resistance to closantel in Haemonchus contortus. AB - OBJECTIVE: To evaluate a laboratory test for closantel resistance in Haemonchus contortus. PROCEDURE: Field isolates of H contortus, known to be resistant to closantel, were tested in the assay. In addition, mixtures of closantel susceptible and closantel-resistant laboratory reference strains were tested to develop a method of predicting the proportion of resistant worms in a sample from the field. RESULTS: The assay correctly identified as resistant all of the closantel-resistant field isolates of H contortus. It also identified one isolate with an in vivo efficacy of 98% as having emerging resistance. Testing of the mixtures of laboratory reference strains revealed that an isolate would be classified as resistant when it consists of about 25% or more resistant worms. Test samples that are not fully susceptible yet contain less than 25% resistant worms may be classified as emerging resistance. CONCLUSION: The in vitro migration assay is a sensitive method of detecting closantel resistance in H contortus. PMID- 11098389 TI - Laparoscopic intrauterine insemination in Barbary sheep (Ammotragus lervia). PMID- 11098390 TI - ST65: supporting quality in reusable textile reprocessing. PMID- 11098391 TI - A perfluorochemical loss/restoration (L/R) system for tidal liquid ventilation. AB - Tidal liquid ventilation is the transport of dissolved respiratory gases via volume exchange of perfluorochemical (PFC) liquid to and from the PFC-filled lung. All gas-liquid surface tension is eliminated, increasing compliance and providing lung protection due to lower inflation pressures. Tidal liquid ventilation is achieved by cycling fluid from a reservoir to and from the lung by a ventilator. Current approaches are microprocessor-based with feedback control. During inspiration, warmed oxygenated PFC liquid is pumped from a fluid reservoir/gas exchanger into the lung. PFC fluid is conserved by condensing (60 80% efficiency) vapor in the expired gas. A feedback-control system was developed to automatically replace PFC lost due to condenser inefficiency. This loss/restoration (L/R) system consists of a PFC-vapor thermal detector (+/- 2.5%), pneumatics, amplifiers, a gas flow detector (+/- 1%), a PFC pump (+/- 5%), and a controller. Gravimetric studies of perflubron loss from a flask due to evaporation were compared with experimental L/R results and found to be within +/ 1.4%. In addition, when L/R studies were conducted with a previously reported liquid ventilation system over a four-hour period, the L/R system maintained system perflubron volume to within +/- 1% of prime volume and 11.5% of replacement volume, and the difference between experimental PFC loss and that of the L/R system was 1.8 mL/hr. These studies suggest that the PFC L/R system may have significant economic (appropriate dosing for PFC loss) as well as physiologic (maintenance of PFC inventory in the lungs and liquid ventilator) impact on liquid ventilation procedures. PMID- 11098392 TI - Electromagnetic interference by GSM cellular phones and UHF radios with intensive care and operating-room ventilators. AB - The aim of this study was to evaluate the risks deriving from the interference by radio handsets (GSM cellular phones and UHF radios) with intensive-care and operating-room ventilators. Tests were conducted in three hospitals in Rome on 22 lung ventilators in accordance with the recommended practice ANSI C63.18-1997. When electromagnetic interference (EMI) effects occurred, the authors determined maximum interference distances. They also evaluated the distances at which the use of a given handset would result in a 5% and a 95% probability of interference. The degree of risk posed by each observed event was estimated, and safe distances are suggested. EMI events of varying degrees and natures were observed even with transmitters placed at a considerable distance. All observed effects were temporary. Only three ventilators of a certain model stopped working altogether and had to be reset. PMID- 11098393 TI - Eliminating the culture of blame; a new challenge for clinical engineers and BMETs. PMID- 11098394 TI - The OSI reference model for computer network communication. PMID- 11098395 TI - Science and pseudoscience in the development of eye movement desensitization and reprocessing: implications for clinical psychology. AB - The enormous popularity recently achieved by Eye Movement Desensitization and Reprocessing (EMDR) as a treatment for anxiety disorders appears to have greatly outstripped the evidence for its efficacy from controlled research studies. The disparity raises disturbing questions concerning EMDR's aggressive commercial promotion and its rapid acceptance among practitioners. In this article, we: (1) summarize the evidence concerning EMDR's efficacy; (2) describe the dissemination and promotion of EMDR; (3) delineate the features of pseudoscience and explicate their relevance to EMDR; (4) describe the pseudoscientific marketing practices used to promote EMDR; (5) analyze factors contributing to the acceptance of EMDR by professional psychologists; and (6) discuss practical considerations for professional psychologists regarding the adoption of EMDR into professional practice. We argue that EMDR provides an excellent vehicle for illustrating the differences between scientific and pseudoscientific therapeutic techniques. Such distinctions are of critical importance for clinical psychologists who intend to base their practice on the best available research. PMID- 11098396 TI - Paradoxical and less paradoxical effects of thought suppression: a critical review. AB - The process of consciously trying to avoid certain thoughts is referred to as thought suppression. Experimental research has documented that thought suppression may have paradoxical effects in that it leads to an increased frequency of the to-be-suppressed thought intruding consciousness. It has also been claimed that suppression has disruptive effects on episodic memory (i.e., a less paradoxical effect). The present article critically evaluates studies on the paradoxical and less paradoxical effects of thought suppression. More specifically, the issue of whether thought suppression plays a causative role in the development of various psychopathological symptoms is addressed. While laboratory studies have come up with highly consistent findings about the paradoxical effects of thought suppression, there is, as yet, little reason to believe that such effects are implicated in the etiology of obsessions, phobias, or other psychopathological conditions. Relatively little work has been done on the alleged memory effects of thought suppression. The studies that have examined this issue have found mixed results. Accordingly, the case for the amnestic power of thought suppression is weak. Alternative explanations and competing theories are discussed, and it is concluded that research concerned with the psychopathological consequences of thought suppression would benefit from development of better taxonomies of intrusive thinking and cognitive avoidance strategies. PMID- 11098397 TI - Childhood maltreatment and malevolence: quantitative research findings. AB - This article examines the quantitative research literature that focuses on the object relations sequelae of childhood maltreatment. A review of 12 studies indicates strong support for a relationship between childhood maltreatment and a malevolent object world. This relationship holds despite variations in methodology across studies, including differences in sampling and measurement techniques. This paper defines the construct of malevolence, and proposes that malevolent representations are a central feature of the experience of childhood maltreatment. Implications for diagnosis and treatment are considered, and directions for future research are outlined. PMID- 11098398 TI - Attachment security: a meta-analysis of maternal mental health correlates. AB - This meta-analysis addresses the association between attachment security and each of three maternal mental health correlates. The meta-analysis is based on 35 studies, 39 samples, and 2,064 mother-child pairs. Social-marital support (r = .14; based on 16 studies involving 17 samples and 902 dyads), stress (r = .19; 13 studies, 14 samples, and 768 dyads), and depression (r = .18; 15 studies, 19 samples, and 953 dyads) each proved significantly related to attachment security. All constructs showed substantial variance in effect size. Ecological factors and approach to measuring support may explain the heterogeneity of effect sizes within the social-marital support literature. Effect sizes for stress varied according to the time between assessment of stress and assessment of attachment security. Among studies of depression, clinical samples yielded significantly larger effect sizes than community samples. We discuss these results in terms of measurement issues (specifically, overreliance on self-report inventories) and in terms of the need to study the correlates of change in attachment security, rather than just the correlates of attachment security per se. PMID- 11098399 TI - Information processing and PTSD: a review of the empirical literature. AB - This article reviews a series of studies that have utilized information processing paradigms with posttraumatic stress disorder (PTSD) populations. The review suggests that pretrauma measures of intelligence (IQ) are predictive of the development of PTSD symptoms following trauma. There is also evidence of impaired performance on standardized tests of memory (independent of IQ) in PTSD populations. PTSD populations are found to exhibit deficits in memory function that may be due to hippocampus damage secondary to excessive neuroendocrine responses to conditioned stimuli. In addition, individuals with PTSD evince an attentional bias towards trauma-related stimuli at postrecognition stages of information processing. The review also includes that there is insufficient evidence to either support, or reject, the theoretical proposition that PTSD patients are sensitive to global valence effects at the earliest stages of information processing relative to traumatized non-PTSD populations. Finally, there is some evidence to suggest that the processes associated with autobiographical memory in PTSD populations are similar to those seen in depression. The implications of these findings for the behavioral and cognitive treatment of PTSD are discussed. Directions for future research with such paradigms are also discussed in light of contemporary information processing theories of PTSD. PMID- 11098400 TI - Mechanisms to account for maintenance of the soluble methionine pool in transgenic Arabidopsis plants expressing antisense cystathionine gamma-synthase cDNA. AB - To investigate the role of cystathionine gamma-synthase (CGS) in the regulation of methionine synthesis Arabidopsis plants were transformed with a full-length antisense CGS cDNA and transformants analysed. Plants that were heterozygous for the transgene showed a 20-fold reduction of CGS activity that was accompanied by severe growth retardation and morphological abnormalities, from germination to flowering. Application of exogenous methionine to the transgenic lines restored normal growth. Surprisingly, transformed Arabidopsis plants exhibited a modest decrease in methionine content (35% reduction of the wild-type level) but a seven fold decrease in the soluble pool of S-methylmethionine (SMM), a compound that plays a major role in storage and transport of reduced sulphur and labile methyl moieties. Several mechanisms can account for the maintenance of the soluble pool of methionine. First, the observed 20-fold increase in O-phosphohomoserine, a substrate of CGS, could compensate for the depressed level of CGS polypeptide by increasing the net rate of catalysis supported by the remaining enzyme. Second, the transgenic plants exhibited a two-fold increased level of cystathionine beta lyase, the second enzyme in the methionine biosynthetic pathway. This indicates that enzymes other than CGS are subjected to a regulatory control by methionine or one of its metabolites. In addition to these mechanisms affecting de novo methionine synthesis, the recruitment of SMM to produce methionine may account for the small change of methionine levels in transgenic lines. PMID- 11098401 TI - [Massive mortality of marine invertebrates: an unprecedented event in northwestern Mediterranean]. AB - An unprecedented mass mortality event has been observed at the end of the summer 1999 along the coasts of Provence (France) and Ligury (Italy). This event has severely affected a wide array of sessile filter-feeder invertebrates from hard substratum communities, such as sponges (particularly the keratose sponges Hippospongia and Spongia), cnidarians (particularly the anthozoans Corallium, Paramuricea, Eunicella and Cladocora), bivalves, ascidians and bryozoans. Along the Provence coasts, the outbreak spread from east to west. Exceptionally high and constant temperatures of the whole water column (23-24 degrees C, for over one month, down to 40 m) could have determined an environmental context favourable to the mass mortality event. Like the thermal anomaly, the mortality is limited in depth. However, we cannot ascertain whether temperature had a direct effect on organisms or acted in synergy with a latent and/or waterborne agent (microbiological or chemical). Taking into account the global warming context in the NW-Mediterranean, monitoring programs of physical-chemical parameters and vulnerable populations should rapidly be set up. PMID- 11098402 TI - [Morphofunctional analysis and trophic adaptations: the case of bill apparatus of Old World flycatchers]. AB - A morphofunctional analysis of the bill apparatus was conducted on some African forest flycatchers (Muscicapidae, Platysteiridae, Monarchidae) in relation to detailed eco-ethological data available on these species. The aim was to evaluate relationships between anatomical structures and habitat constraints and also identify the most pertinent trophic adaptations. If forest Muscicapidae have essentially conserved the generalized passerine structures and occupy open habitat niches in the forest, Platysteiridae and Monarchidae have adapted to forest conditions and show a key-adaptation based on specific changes in the structure of the bill apparatus in response to particular light patterns and habitat clogginess, constraints that require speed and precision for the capture of prey and protection of the skull against collisions. PMID- 11098403 TI - [A new genus of Malagasy scorpion, possible link between the Microcharmidae and the Buthidae]. AB - A new genus belonging to the family Microcharmidae is described from the northern range of Madagascar. The new genus appears as a possible link between the Microcharmidae and the Buthidae within the Buthoidea. Phylogenetic considerations are proposed in relation to the observed morphological characters. PMID- 11098404 TI - 1H-13C nuclear magnetic resonance assignment and structural characterization of HIV-1 Tat protein. AB - Tat is a viral protein essential for activation of the HIV genes and plays an important role in the HIV-induced immunodeficiency. We chemically synthesized a Tat protein (86 residues) with its six glycines C alpha labelled with 13C. This synthetic protein has the full Tat activity. Heteronuclear nuclear magnetic resonance (NMR) spectra and NOE back-calculation made possible the sequential assignment of the 86 spin systems. Consequently, 915 NMR restraints were identified and 272 of them turned out to be long range ([i-j] > 4), providing structural information on the whole Tat protein. The poor spectral dispersion of Tat NMR spectra does not allow an accurate structure to be determined as for other proteins studied by 2D NMR. Nevertheless, we were able to determine the folding for Tat protein at a 1-mM protein concentration in a 100 mM, pH 4.5 phosphate buffer. The two main Tat functional regions, the basic region and the cysteine-rich region, are well exposed to solvent while a part of the N-terminal region and the C-terminal region constitute the core of Tat Bru. The basic region adopts an extended structure while the cysteine-rich region is made up of two loops. Resolution of this structure was determinant to develop a drug design approach against Tat. The chemical synthesis of the drugs allowed the specific binding and the inhibition of Tat to be verified. PMID- 11098405 TI - The differential effects of pelvic and vagal inputs on the supraspinal cystitis viscero(noci)ceptive-related axis. AB - Recently the development of the cyclophosphamide (CP, 100 mg/kg/i.p.) model has added an important element to the study of neural activities accompanying cystitis genesis. CP cystitis genesis results in the dual activation of the pelvic and vagal sensory afferent systems, which in turn activate a supraspinal network comprising the ventrocaudal bulbar reticular formation (vcBRF), the sensory subdivisions of the dorsal vagal complex (DVC) and its subcortical telencephalic targets, the dorsolateral subdivision of the bed nucleus of the stria terminalis (BSTLd) and the nucleus centralis of the amygdala (CeL). Altogether these structures form the sensory neural axis of the CP cystitis. However, both clinical and experimental observations have given evidence that only the pelvic afferents are at the origin of the painful sensation and related behaviour. Because of this, and for a better understanding of the nervous network that subserves cystitis painful information, we sought to determine whether the structures that constitute the cystitis sensory neural pathway have the same reactivity depending on the origin of the sensory afferent inputs, either pelvic or vagal. Using c-fos expression, which permits quantitative analysis of neural activity, we have demonstrated that the supraspinal CP cystitis responding structures do not form an homogeneous population in terms of sources of inputs. Although all structures are predominantly driven by vagal inputs, only the vcBRF, the DVC and the BSTLd respond to pelvic inputs. Consequently, and by referring to clinical observations, we have concluded that, it is these three areas, excluding the CeL, which constitute the main framework of the supraspinal pain sensory neural pathway of CP-induced cystitis. The activation of the vagus nerve would more probably relate to the other side effects that accompany CP injections such as nausea and headache attacks. PMID- 11098406 TI - [Influence of biotic and abiotic factors on the morphology and reproduction of Suaeda maritima on a salt marsh]. AB - The aim of this study was to estimate the influence of biotic and abiotic factors on Suaeda maritima reproduction on a salt marsh. Individuals of Suaeda maritima were submitted in natural conditions to four series of densities (100, 1,000, 4,000 and 8,000 plants/m2). When density increases, individuals tend to be less or non-branched, while individual biomass decreases. Consequently, individual seed production decreases as density increases. Despite morphological modifications, Suaeda maritima present density-dependent mortality. For a unit area, total biomass and seed production are higher at intermediate density (1,000 plants/m2). Environmental factors could interfere with self-thinning. They seem to limit the effect of competition on mortality and to have an influence on individual and total seed production. This experiment stressed the importance of a biotic factor such as intra-specific competition, which occurs at the same time as abiotic factors, in Suaeda maritima dynamics in the field. PMID- 11098407 TI - [Impact of the climatic scenario of global warming on the growth of trees]. AB - The climatic impact on tree radial growth resulting from an atmospheric CO2 doubling was studied for 24 populations of five tree species in the French Alps and the French Mediterranean area. The Arpege AGCM, which predicts a 3 degrees C increase in mean temperature and a light increase of precipitation, is used to estimate the climatic perturbation. The method is based on the integration of estimated climate in an empirical tree-ring to climate model, involving artificial neural networks. Only a few populations are sensitive to the climatic change; all are located on the boundaries of their ecological area and can be divided in two groups. The first one is composed of high altitude populations which show a growth increase induced by the warmer climate during the growing season. The second one, composed of a single Mediterranean Scots pine population, reacts with a severe growth reduction induced by the stronger water stress in summer. PMID- 11098408 TI - Performance of 18S rDNA helix E23 for phylogenetic relationships within and between the Rotifera-Acanthocephala clades. AB - The species diversity of the phylum Rotifera has been largely studied on the basis of morphological characters. However, cladistic relationships within this group are poorly resolved due to extensive homoplasy in morphological traits, substantial phenotypic plasticity and a poor fossil record. We undertook this study to determine if a phylogeny based on partial 18S rDNA, which included the helix E23 of 18S rDNA sequence, was concordant with established taxonomic relationships within the order Ploimida (class: Monogononta). We also estimated the level of polymorphism within clones and populations of Ploimida 'species'. Finally, we included the Cycliophora Symbion pandora as outgroup and the variable helix E23 region to examine the influence of their signal on the evolutionary relationships among Acanthocephala, Bdelloidea and Ploimida. Phylogenetic reconstruction was performed using maximum parsimony, neighbour joining and maximum likelihood methods. We found 1) that morphologically similar Ploimida 'species' show vastly different 18S E23 rDNA sequences; 2) inclusion of the helix E23 of 18S rDNA and its secondary structure analysis results in better resolution of family level relationships within the Ploimida; 3) an impact of Symbion pandora as an outgroup with inclusion of the helix E23 on the relationships between the Rotifera and the Acanthocephala; and 4) partial incongruence and differential substitution rate between conserved region and helix E23 region of the 18S rDNA gene depending on the taxomic group studied. PMID- 11098409 TI - Evaluation of 3D imaging. AB - Interactive computer-based simulation is gaining acceptance for craniofacial surgical planning. Subjective visualization without objective measurement capability, however, severely limits the value of simulation since spatial accuracy must be maintained. This study investigated the error sources involved in one method of surgical simulation evaluation. Linear and angular measurement errors were found to be within +/- 1 mm and 1 degree. Surface match of scanned objects was slightly less accurate, with errors up to 3 voxels and 4 degrees, and Boolean subtraction methods were 93 to 99% accurate. Once validated, these testing methods were applied to objectively compare craniofacial surgical simulations to post-operative outcomes, and verified that the form of simulation used in this study yields accurate depictions of surgical outcome. However, to fully evaluate surgical simulation, future work is still required to test the new methods in sufficient numbers of patients to achieve statistically significant results. Once completely validated, simulation cannot only be used in pre operative surgical planning, but also as a post-operative descriptor of surgical and traumatic physical changes. Validated image comparison methods can also show discrepancy of surgical outcome to surgical plan, thus allowing evaluation of surgical technique. PMID- 11098410 TI - Role of serotonergic and noradrenergic systems in the pathophysiology of depression and anxiety disorders. AB - There is abundant evidence for abnormalities of the norepinephrine (NE) and serotonin (5HT) neurotransmitter systems in depression and anxiety disorders. The majority of evidence supports underactivation of serotonergic function and complex dysregulation of noradrenergic function, most consistent with overactivation of this system. Treatment for these disorders requires perturbation of these systems. Reproducible increases in serotonergic function and decreases in noradrenergic function accompany treatment with antidepressants, and these alterations may be necessary for antidepressant efficacy. Dysregulation of these systems clearly mediates many symptoms of depression and anxiety. The underlying causes of these disorders, however, are less likely to be found within the NE and 5HT systems, per se. Rather their dysfunction is likely due to their role in modulating, and being modulated by, other neurobiologic systems that together mediate the symptoms of affective illness. Clarification of noradrenergic and serotonergic modulation of various brain regions may yield a greater understanding of specific symptomatology, as well as the underlying circuitry involved in euthymic and abnormal mood and anxiety states. Disrupted cortical regulation may mediate impaired concentration and memory, together with uncontrollable worry. Hypothalamic abnormalities likely contribute to altered appetite, libido, and autonomic symptoms. Thalamic and brainstem dysregulation contributes to altered sleep and arousal states. Finally, abnormal modulation of cortical-hippocampal-amygdala pathways may contribute to chronically hypersensitive stress and fear responses, possibly mediating features of anxiety, anhedonia, aggression, and affective dyscontrol. The continued appreciation of the neural circuitry mediating affective states and their modulation by neurotransmitter systems should further the understanding of the pathophysiology of affective and anxiety disorders. PMID- 11098411 TI - Neuropharmacology of venlafaxine. AB - Venlafaxine (Effexor) is an effective antidepressant and has also been approved for the treatment of generalized anxiety disorder. Venlafaxine was initially characterized as an inhibitor of both serotonin (5HT) and norepinephrine (NE) uptake and was therefore termed a "dual uptake inhibitor." This chapter reviews data from both in vitro and in vivo studies regarding its effects on 5HT and NE neurotransmission. In addition, the effects of venlafaxine on other systems that may play a role in its therapeutic efficacy effects are described. The data indicate that venlafaxine is a relatively weak inhibitor of NE transport in vitro. In vivo studies indicate that venlafaxine selectively inhibits 5HT uptake at low therapeutic doses and inhibits both 5HT and NE uptake at higher therapeutic doses. This chapter concludes with a discussion of the effects of venlafaxine on various aspects of physiology. PMID- 11098412 TI - Review of the pharmacokinetics, pharmacogenetics, and drug interaction potential of antidepressants: focus on venlafaxine. AB - Improving outcomes for patients with depression involves selecting the best possible drug therapy. Considerations relevant to drug product selection include: 1) pharmacokinetic issues such as half-life and time to steady-state, and protein binding; 2) pharmacodynamic drug-drug interactions; and 3) drug metabolism related drug interactions. A comparison of selected antidepressants with an emphasis on venlafaxine's similarities and differences is presented. Based on these parameters, selecting an antidepressant medication, such as venlafaxine, that has a low potential for drug interactions at the Cytochrome P450 (CYP) enzyme system, and is easy to monitor and dose, facilitate successful treatment of patients. Venlafaxine has been evaluated in clinical studies that demonstrate low to negligible drug interaction potential at CYP2D6, CYP1A2, CYP2C19, and CYP3A4. Its short half-life and time to steady-state, when coupled with the extended release characteristics of the preferred dosage formulation allow for once daily dosing and rapid attainment of therapeutic effects. The CYP3A4 system is involved in both first-pass metabolism and systemic clearance of medications. Drug interactions at this isoenzyme have proven to be of high clinical relevance ranging from cardiovascular toxicity and death with commonly used drugs such as cisapride, to subtherapeutic levels of cyclosporine or protease inhibitors leading to transplant rejection or HIV relapse. Reasons for the under detection and reporting of drug interaction mediated adverse events include healthcare system structure, the poor return to follow up of non-adherent patients, the need for greater education and training of clinicians to recognize drug-related adverse events, and the reluctance of patients to spontaneously communicate about the unpleasant effects of their medication. PMID- 11098413 TI - Antidepressant efficacy of venlafaxine. AB - Venlafaxine is a unique antidepressant medication with well documented efficacy and safety in the acute treatment of major depressive disorder. Reports suggest that it may also be effective in the treatment of dysthymic disorder and bipolar II depression, but the available data for these conditions are more limited compared to major depressive disorder. Several studies suggest that there may be a more rapid onset of action for venlafaxine in the treatment of major depression compared to other antidepressant pharmacotherapies, but this has not been fully established. Venlafaxine is also effective in the important long term continuation and maintenance phases of the treatment of depression. PMID- 11098414 TI - Efficacy of venlafaxine in the treatment of severe depression. AB - Although the efficacy of available antidepressants has been well established in the treatment of mild to moderate depression, clinical research literature on severe depression is more limited, due to lack of a standardized definition for the condition and the resulting inconsistent data. Given the heterogeneous nature of severe depression, reports suggesting noradrenergic as well as serotonergic system involvement in depressive disorders, and the substantive capability of both clomipramine and TCA-SSRI combination to treat severe depression, investigation of dual-action antidepressant agent efficacy in the treatment of severe depression is warranted. The merit of one such combined-action agent, venlafaxine, is reviewed. Efficacy findings from the limited number of comparative clinical trials conducted in the severely depressed patient population suggest that, while venlafaxine has been evaluated in a broad range of depressed patients, this compound may be particularly effective for the severely ill. Pharmacological features of venlafaxine, which may benefit the patient with severe depression, include the possibility of a rapid onset of action and a dose response curve. Based upon studies comparing venlafaxine with both placebo and other first-line antidepressants, it is concluded that venlafaxine is safe, tolerable, and effective for the treatment of severe depression. PMID- 11098415 TI - Venlafaxine and treatment-resistant depression. AB - Treatment-resistant depression (TRD) is an important clinical problem. This paper briefly reviews the definition of TRD and summarizes methodological issues that pertain to treatment research. Recent studies of venlafaxine treatment for TRD also are reviewed. It is concluded that venlafaxine at higher doses is a reasonably well-tolerated and an effective alternative for patients with TRD and typically should be used before tricyclic antidepressants or monoamine oxidase inhibitors. Further research is needed to confirm the prediction that switching a SSRI nonresponder to venlafaxine is a more effective strategy than switching to a second SSRI. The relative merits of switching from a SSRI to venlafaxine versus adding a norepinephrine reuptake inhibitor also warrant careful study. PMID- 11098416 TI - Efficacy of venlafaxine in geriatric depression. AB - Geriatric patients with major depression present clinical challenges not encountered in younger individuals, including a greater incidence of medical comorbidity, higher rates of multiple medication use, changes in drug metabolism due to age or physical illness, and increased sensitivity to antidepressant side effects. Nevertheless, successful treatment of depressive disorders in the elderly improves mental and physical functioning, decreases morbidity and perhaps mortality, and enhances quality of life. Recent research indicates that newer antidepressants are effective for late life depression and safer for older individuals. Among newer antidepressants, venlafaxine has a pharmacological profile that makes it an attractive choice for geriatric patients. It has limited potential to interact with other medications because it only weakly inhibits the cytochrome P450 system and binds to plasma proteins at a low level. Dosing may have to be adjusted for patients with renal failure, but typically not for those with liver disease or other medical conditions. Data from three double-blind and four open clinical trials support the safety and efficacy of venlafaxine for geriatric depression. Patients may experience transient, generally tolerable side effects such as insomnia, nausea, agitation, or dry mouth early in treatment, but more serious problems such as falls or cardiac rhythm disturbances seem to be rare. Treatment emergent hypertension occurs in a small percentage of older patients, generally at doses above 150 mg/day. Finally, emerging data suggest that venlafaxine may be effective for conditions such as stroke, anxiety, and neuropathic pain that frequently accompany depressive disorders in the elderly. PMID- 11098417 TI - Comorbidity of mood and anxiety disorders. AB - This article reviews data on the prevalence of panic, social phobia, generalized anxiety, and posttraumatic stress disorder, and research documenting the comorbidity of these disorders with major depression (MDD). These anxiety disorders are frequently comorbid with MDD, and 50-60% of individuals with MDD report a lifetime history of one or more of these anxiety disorders. The anxiety disorders are also highly correlated with one another, and approximately one quarter to one-half of individuals with each of the anxiety disorders report a lifetime history of an alcohol or substance use disorder. Anxiety disorders rarely exist in isolation, with several studies reporting that over 90% of individuals with anxiety disorders have a lifetime history of other psychiatric problems. Implications for research are discussed, including the potential benefit of using combined categorical and dimensional rating scale approaches in future genetic, biochemical, neuroimaging, and treatment studies. The clinical implications of the findings are also discussed, and the results of recent clinical trials summarized. Available data suggests selective serotonin reuptake inhibitors are the first-line pharmacological treatment for these disorders, and that newer serotonin and norepinephrine reuptake inhibitors show significant promise, especially for comorbid cases. Comorbidity among depression and anxiety disorders is associated with greater symptom severity, and a considerably higher incidence of suicidality. Increased public awareness about these disorders and the availability of effective treatments is sorely needed. PMID- 11098418 TI - Efficacy of venlafaxine in mixed depression-anxiety states. AB - Patients with depression almost always suffer from comorbid anxiety or anxiety disorder. It is commonly stated that comorbid depression and anxiety has a worse prognosis, even with adequate therapy, than depression alone. An accumulation of data now make clear that the antidepressants venlafaxine and venlafaxine XR are effective in reducing anxiety in patients with depression. Several of the studies supporting this are reviewed here. Venlafaxine and venlafaxine XR have also been shown to be effective in treating anxiety disorders and venlafaxine XR is presently the only antidepressant approved by the FDA for the specific treatment of generalized anxiety disorder. The effectiveness of venlafaxine in treating anxiety associated with depression and anxiety disorders supports theories implicating abnormal noradrenergic activity as a component of pathological anxiety. PMID- 11098419 TI - Efficacy, safety, and tolerability of venlafaxine XR in generalized anxiety disorder. AB - Generalized anxiety disorder (GAD) is a common and chronic disorder with a low rate of spontaneous remission. A complication in treatment selection is the high rates of co-morbid major depressive disorder in this population. A number of treatments exist to treat GAD. The most recent medication to gain an indication for GAD is venlafaxine XR, a serotonin/norepinephrine reuptake inhibitor that is also approved for the treatment of major depressive disorder. More than 2,000 patients with GAD have been studied in outpatient trials of venlafaxine XR with demonstrated efficacy, tolerability and safety of this compound. This article reviews these studies, both short term and longer (6 month) continuation trials. The response to venlafaxine XR in this population, combined with good tolerability, makes this agent an appropriate first-line medication for GAD. In general, treatment with antidepressants, though associated with a longer onset of action than benzodiazepines, does not produce physiological dependency, and is useful in a patient population with a high prevalence of mood disorders. PMID- 11098420 TI - Use of venlafaxine in children and adolescents: a review of current literature. AB - Pediatric psychopharmacology is hindered by the relative lack of controlled, empirical clinical trials [Shatzberg and Nemeroff, 1998: Washington, D.C.: The American Psychiatric Press, Inc. p 301-306]. Psychiatric disorders in children and adolescents carry considerable morbidity, impede development, and carry a significant mortality by suicide. Therefore, there is a need for studies of antidepressants and other psychotropics in children and adolescents. This article reviews the preliminary evidence that venlafaxine (Effexor), a novel antidepressant, may be useful in children and adolescents with a variety of psychiatric disorders. PMID- 11098421 TI - Use of venlafaxine in other psychiatric disorders. AB - Venlafaxine is a medication available by prescription in the U.S. both in an immediate release and an extended release formulation. Preclinical studies indicate it has the effect of potently blocking the serotonin and norepinephrine transporters. Venlafaxine is approved by the FDA for the treatment of major depressive disorder and generalized anxiety disorder. Suggestive evidence, mostly from open label case series, indicates efficacy of venlafaxine in several other conditions including panic disorder, social anxiety disorder, obsessive compulsive disorder, trichotillomania, ADHD, chronic pain, and fibromyalgia. The limited evidence supporting efficacy in these conditions is reviewed. Additional randomized clinical trials with placebo controls are indicated. PMID- 11098422 TI - Completeness of response and quality of life in mood and anxiety disorders. AB - Mental health care has traditionally focused on the need to document relief of specific symptoms of a psychiatric disorder, as well as how the patient functions in social roles. Recently, there has been increased attention paid to the issue of quality of life (QOL) and psychiatric illness. There has been a growing recognition that different treatment options may vary in their effects on the patient's ability to function in multiple life domains. Studies focusing on the QOL in patients suffering from mood and anxiety disorders have become more prevalent. Depression and anxiety disorders impose a substantial cost on society in terms of both psychiatric service costs as well as the loss of the individual to society through lost work production. However, a change in the severity of depression or anxiety often correlates with a change in disability and health service utilization. Lately, there have been a number of treatment studies of anxiety and depressive disorders that have examined the effect of treatment on QOL. Although treatment may reduce the severity and frequency of target symptoms, the patient's assessment of QOL helps to differentiate a true treatment response and remission from a partial response. The evaluation of what constitutes an adequate treatment response or remission is complicated and likely requires multiple assessment instruments in order to develop a complete understanding. In both anxiety and depressive disorders, the patient suffers from impaired functioning, which results in increased healthcare utilization. Because these patients do respond to treatment, the idea of "wellness" as a high end state treatment outcome should be an important consideration when selecting a treatment option. PMID- 11098423 TI - Pharmacoeconomic studies of antidepressants: focus on venlafaxine. AB - Newer antidepressants are more expensive in terms of acquisition costs than older drugs. However, cost effectiveness simulations and retrospective analyses of administrative databases of newer antidepressants, including venlafaxine, suggest that the higher acquisition costs may be offset or more than offset by savings of other treatment costs. Because simulations and retrospective studies are vulnerable to multiple methodologic uncertainties, large scale randomized "real world" cost effectiveness experiments are needed. If venlafaxine in actual practice is more effective or has a more rapid onset of action than SSRIs as suggested by efficacy studies and existing meta-analyses, these effects could translate into pharmacoeconomic advantages. PMID- 11098424 TI - Improving diabetes care in a large health care system: an enhanced primary care approach. AB - OBJECTIVE: The objective of this study was to evaluate the impact of a multifaceted improvement strategy on diabetes quality of care in a defined population of patients. STUDY DESIGN: A multifaceted improvement strategy to enhance diabetes care was deployed to 18 primary care clinics serving 170,000 adults. Interventions empowered patient self-management, supported care team decision making, redesigned office systems, and maximized use of available information technology. Specific goals were to improve glycemic control and reduce cardiovascular risk in all adult diabetes patients. DATA SOURCE AND COLLECTION: Diabetes was identified through pharmacy and diagnostic data (estimated sensitivity 0.91, positive predictive value 0.94), and the target population ranged from 6,542 to 7,037 members over time. Trends in glycosylated hemoglobin (HbA1c) and low-density lipid LDL-cholesterol were analyzed monthly throughout 1999 in both cohorts and serial cross-sections. RESULTS: During 12 months, mean HbA1c improved from 7.86% to 7.47%, and the proportion of patients with HbA1c levels < 8% rose from 60.5% to 68.3%, and the proportion with HbA1c > 10% fell from 10.3% to 7.2%. The LDL test rate rose from 47.4% to 57.4%, and mean LDL fell from 120 mg/dl to 116 mg/dl. The proportion with acceptable lipid control (LDL < 130 mg/dl, or < 100 mg/dl with coronary artery disease) rose from 48.9% to 57.7%. All changes were significant at p < 0.01 or less. CONCLUSION: Clinically significant population-based improvements in diabetes care were observed during a 1-year period using a multifaceted "enhanced primary care" strategy. PMID- 11098425 TI - Improving hypertension control: a team approach in a primary care setting. AB - BACKGROUND: Blood pressure (BP) control rates in the United States have not improved significantly during the past decade. There has been limited study of improvement efforts focusing on guideline implementation and changes in the model of care to address hypertension. METHODS: Five physician (MD)/registered nurse (RN)/licensed practical nurse (LPN) teams in a large community practice modified their care model in 1997 to manage hypertensive patients as part of guideline implementation efforts. The other 25 MD teams in the same setting practiced in the usual model, but were exposed to the guideline recommendations. BP control rates of patients in each group were assessed monthly. After nine months of testing the new care model, 10 additional teams adopted the model. RESULTS: In the pilot group, hypertension control rates showed statistically significant improvement from pre- (33.1%) to postimplementation (49.7%). After adjusting for age, this was significantly greater than the improvement in the control group (p = 0.033). Medication changes were more frequent in the pilot group (32.3%) than in the control group (27.6%); however, the differences were not statistically significant. A longitudinal examination of the hypertension patients in the study showed that improved BP control was sustained for at least 12 months. DISCUSSION: A change in the model of care for hypertensive patients within a primary care practice resulted in significant, sustainable improvement in BP control rates. These changes are consistent with the chronic care model developed by Wagner; practice redesign appeared to be the most important change. PMID- 11098426 TI - Developing performance measures for alcohol and other drug services in managed care plans. Washington Circle Group. AB - BACKGROUND: Monitoring the quality and availability of alcohol and other drug (AOD) services must be a central tenet of any health-related performance measurement system. The Washington Circle Group (WCG), which was convened by the Center for Substance Abuse Treatment Office of Managed Care in March 1998, has developed a core set of performance measures for AOD services for public- and private-sector health plans. It is also collaborating with a broad range of stakeholders to ensure widespread adoption of these performance measures by health plans, private employers, public payers, and accrediting organizations. CORE PERFORMANCE MEASURES: Four domains were identified, with specific measures developed for each domain: (1) prevention/education, (2) recognition, (3) treatment (including initiation of alcohol and other plan services, linkage of detoxification and AOD plan services, treatment engagement, and interventions for family members/significant others), and (4) maintenance of treatment effects. CONTINUING EFFORTS: Four measures that are based on administrative information from health plans and two measures that require a consumer survey of behavioral health care are undergoing extensive pilot testing. The WCG has reached out to a broad range of stakeholders in performance measurement and managed care to acquaint them with the measures and to promote their investigation and adoption. As results of pilot testing become available, these outreach efforts will continue. CONCLUSIONS: Performance measures for AOD services need to become an integral part of a comprehensive set of behavioral and physical health performance measures for managed care plans. PMID- 11098427 TI - Using standardized patients to measure quality: evidence from the literature and a prospective study. AB - BACKGROUND: Use of standardized patients for evaluating the clinical skills of medical students and medical trainees is commonplace. This has encouraged the use of standardized patients to evaluate the quality of physician practice in outpatient settings. However, there may be substantive differences between observing student performance and evaluating whether the provision of care meets defined quality criteria. OBJECTIVES: This study had two primary objectives: (1) to review studies that use standardized patients to evaluate physician performance and (2) to ascertain directly whether standardized patients could be useful in assessing quality of outpatient care. METHODS: A comprehensive literature review of studies that used standardized patients to assess physician performance was conducted. A prospective study that included 20 physicians at two outpatient settings and 27 actor patients assessed quality of care using eight clinical cases divided into five clinical domains, each of which had explicit criteria checklists based on widely accepted guidelines. RESULTS: The literature review identified five important issues: developing scenarios, selecting explicit criteria, standardizing standardized patient training, creating subterfuges, and ensuring reliability and validity of measures. In the study, trained standardized patients were able to assess physician practice accurately for common medical conditions, using proven criteria linked to health outcomes. The detection rate was 3%. There was no performance variation between actors for seven of the eight cases. CONCLUSIONS: Using standardized patients to measure the quality of care is practical and feasible. The major methodological challenge is incorporating observable evidence-based criteria into realistic scripts and objective checklists. The major logistical challenge is obtaining and maintaining undetected entry into physicians' offices. PMID- 11098428 TI - Applying workflow analysis tools to assess immunization delivery in outpatient primary care settings. AB - BACKGROUND: As health care organizations face increasing pressure to institute quality assurance activities, the already-underfunded community clinics that treat the poor and underserved are challenged to perform these activities within tight constraints of human and financial resources. With pediatric immunizations as a marker, a workflow observation tool was used to identify causal processes affecting immunization delivery. METHODS: Ten clinics and five private practices, located in areas designated as health professional shortage areas, participated in the study, gaining access to a tool that would have been unaffordable to them from the private sector. Trained observers followed families through the clinic, using a 127-item workflow observation form--the Observational Checklist of Patient Encounters (OCPE)--assessing discrete activities that families encountered during the checkin/pre-exam, exam, discharge, and billing processes. A convenience sampling of the targeted population--children younger than three years of age--included observations of scheduled acute, scheduled well-child, follow-up, and walk-in visits. In the feedback session, a summary of each clinic's immunization delivery patterns was presented, with an emphasis on the individual health center's operational issues. RESULTS: The workflow observation tool was used to identify operational errors affecting both clinical and fiscal processes in each of the clinics that had not been previously apparent to either clinic management or the quality improvement (QI) teams. DISCUSSION: Feedback addressed and encouraged process-oriented improvements in response to the workflow observations, which were incorporated into the clinics' QI procedures. Twelve of the 15 clinics have formed process action teams to address QI issues on an ongoing basis. PMID- 11098429 TI - [Lipids from fossil plants and their relation to modern plants. Example s of Cenomanian flora from Anjou and Bohemia]. AB - Comparative analyses of lipids from fossil plants and from their extant counterparts were undertaken in order to test the taxonomic significance of lipids in palaeobotany. The comparison between lipids from a fossil Ginkgoaceae, Eretmophyllum andegavense, and its extant counterpart, Ginkgo biloba, revealed the presence of original molecules, dimethoxyalkylcoumarins, in lipids from both plants. Such compounds confirm, on chemical grounds the relationship between these extant and fossil Ginkgoaceaes. Moreover, differences in n-alkane distribution between E. andegavense and E. obtusum which are very similar morphologically, confirm that these fossil plants do not belong to the same species. Furthermore, comparative analyses of a fossil Cheirolepidiaceae, Frenelopsis alata, and its extant counterpart, the Cupressaceae Tetraclinis articulata, revealed some similarities between these two species although they do not belong to the same family. Otherwise, comparative analyses of fungi-infected and uninfected samples of F. alata demonstrated that these micro-organisms can significantly affect the chemical composition of fossil plant lipids. In conclusion, even if chemical analyses alone are not sufficient to determine the genus or species of a given fossil plant, they can precise the taxonomy of some specimens that have been previously studied by palaeobotanists. PMID- 11098430 TI - [The first trees. The Archaeopteris model]. AB - The earliest self-supporting organisms exceeding 2 m in height evolved about 370 million years ago, approximately 100 million years after the rise of the first land plants. Evidence for the tree habit is usually indirect and assessed from the diameter of the available stem fragments. Four systematic groups of Devonian plants evolved the tree habit independantly: the Lycopsida, Cladoxylopsida, and progymnosperms in the Middle Devonian, the Equisetopsida in the Late Devonian. All share a free-sporing life cycle which limits their habitats to wet areas. Their branching pattern involves the strict division of their apices, whether equally or unequally. The progymnosperm genus Archaeopteris was widespread worldwide and evolved the highest trees of the Devonian (maximum height estimated at 40 m). Besides it ecological significance as the dominant component of the earliest forests, Archaeopteris currently represents the closest known relative to the seed plants with which it shares two derived characters, the heterosporous life cycle, and the possession of leaves. Another distinctive feature of Archaeopteris trees is represented by the double function of their wood for both support and conduction. New analyses involving vascular trace analysis in anatomically preserved specimens have demonstrated that Archaeopteris is not the simple tree reconstructed by Beck (1962). In this fate model, Archaeopteris consisted of an erect trunk bearing short-lived, flattened, leaf-like branch systems forming a terminal crown. New evidence indicates that laterally to these appendages of apical origin, a new type of branches, of adventitious origin, evolved which development compares to that of the axillary branches of the seed plants. These branches which were large and long-lived represent major architectural components of the tree. Evidence for vascular structures comparable to those produced on stem cuttings in modern plants suggest that Archaeopteris may have evolved vegetative strategies for propagation. The set of "modern" characters of Archaeopteris may explain its success until the Devonian/Carboniferous boundary when its extinction is correlated to the radiation of the earliest seed plants. PMID- 11098431 TI - [The neuronal zootype]. AB - We present an hypothesis, derived from the zootype concept of Slack, Holland and Graham. The main point of this hypothesis is to postulate that the primordial function of the zootype genes is to design an appropriate neuronal network in bilaterian animals, by controlling the genes involved in the specificity of the axon pathways. This would be the primary function of the zootype genes in development and their primitive function in evolution. The hypothesis is discussed in view of the current knowledge on the Hox genes, their evolution, their genomic organisation, their expression and their targets. PMID- 11098432 TI - [Structural and functional diversity of homeodomain genes of the orthodenticle and empty spiracles classes in Craniata]. AB - Despite extensively divergent morphologies, the patterning of the embryonic brain is controlled by highly conserved genetic networks. Orthodenticle and empty spiracles-related homeodomain genes, which are expressed in insects as in vertebrates in anteriormost neuromeres of the embryonic brain, provide examples of such conservations. In gnathostomes, they form small multigene families, each containing three well-characterised orthology classes. In mice, paralogous genes play very different roles in the development of cephalic regions. Some of these roles are probably ancient and conserved in all chordates, while others, like the morphogenesis of gnathostomespecific characters, correspond to much more diversified functions. Genetic analyses in mice together with comparative analyses in a broad range of vertebrates provide new possibilities to investigate the molecular mechanisms which underlie these functional diversifications. PMID- 11098433 TI - [Evolution and development of dopaminergic neurotransmitter systems in vertebrates]. AB - Dopamine is a widespread neurotransmitter which exerts numerous neuromodulatory actions in the vertebrate central nervous system. This pleiotropic activity relies on the organisation of dopamine-synthesizing neuronal pathways and on a multiplicity of specific membrane receptors. A comparative approach has been undertaken to gain clues on the genetic events which took place during evolution to devise the dopamine systems of modern vertebrates. The localisation and phenotype of dopamine-synthesizing neurones is determined by different gene networks in each of the dopaminergic nuclei. However, despite this amazing diversity, the overall organisation of the dopaminergic nuclei is strinkingly conserved in the main vertebrates groups. In sharp contrast, the number of dopamine receptors subtypes has been multiplied by two major steps of gene duplications during vertebrates evolution. The first one occurred in the lineage leading to agnathans, whereas the second was concomitant to the emergence of cartilaginous fish. Accordingly, three subtypes exist in D1 receptor class (D1A, D1B, D1C) in all the jawed vertebrates, with two exceptions: eutherian mammals where only two D1 subtypes are found (D1A, D1B) and archosaurs where a fourth subtype is present (D1D). Comparisons of the pharmacological and biochemical characteristics of the dopamine receptors in vertebrate groups revealed homologous features that define each of the receptor subtypes and that have been fixed after gene duplications. The comparison of the distribution of the D1 receptor transcripts in the brain of teleosts and mammals points to significant conserved or derived expression territories, revealing previously neglected aspects of dopamine physiology in vertebrates. PMID- 11098434 TI - [Genistein represses the induction of prostatic buds by testosterone] . AB - Genistein, a phytoestrogen and a kind of endocrine disrupters, inhibits tyrosine specific protein kinase activity of the epidermal growth factor (EGF) receptor. It is also effective both in the suppression of the prostatic cell proliferation and the prostate carcinogenesis. We have recently demonstrated that several growth factors, like EGF, transforming growth factor-alpha (TGF-alpha), or keratinocyte growth factor (KGF), can induce prostatic bud formation in the absence of androgen. The present study was performed to investigate whether genistein can suppress testosterone-induced prostatic bud formation. Urogenital sinuses of 16.5-day male rat fetuses were cultured organotypically for 5 days in a serum-free medium containing 10 or 100 ng/ml genistein and 50 ng/ml testosterone. The number and total volume of prostatic buds were analyzed by laser scanning microscopy and computerized. We found that genistein inhibits significantly testosterone-induced prostatic bud formation. In the presence of genistein, cell proliferation of the sinus epithelium was suppressed and the number of prostatic buds and total volume of the buds were reduced as compared with those in the sinuses cultured with testosterone alone. Genistein did not appear to cause necrosis of the sinus. These results support our hypothesis that growth factors like EGF secreted from the sinus mesenchyme activated by testosterone are involved in the induction and stimulation of growth of the prostatic buds. PMID- 11098435 TI - [In vivo studies of the inhbitory effect of various food components on aflatoxin B1 metabolism]. AB - Possible interferences with aflatoxin B1 metabolism, of some compounds naturally present in food (quercetin, beta-naphthoflavone), resulting from way of cooking method (2-aminodipyrido [1,2-a; 1',2'-d] imidazole (Glu-P-2), norharmane; NH) or used as food additives (butylated hydroxytoluene; BHT) have been studied in vivo by evaluating the production of adducts to glutathione and adducts to serum proteins in laboratory rats. Glu-P-2 and norharmane inhibit strongly the production of adducts to glutathione whereas quercetin and beta-naphthoflavone have only a low effect. BHT is completely ineffective. The adducts to proteins are inhibited by the five compounds, norharmane being the most efficient. PMID- 11098436 TI - [In vitro study of the interference of certain food components with the metabolism of aflatoxin B1]. AB - Some compounds naturally present in food (quercetin, beta-naphthoflavone), used as food additives (butylated hydroxytoluene, sodium sulfite) or resulting from the way they were cooked (2-aminodipyrido [1,2-a; 3', 2'-d] imidazole, norharmane) can interfere with AFB1 metabolism. These interferences have been studied in vitro by evaluating the production of adducts to glutathione and by the Ames test on Salmonella typhimurium. Whereas all compounds produced a drastic decrease of the mutagenic activity, the first three only (quercetin, beta naphthoflavone, butylated hydroxytoluene) interfered with the production of the adducts to glutathione. PMID- 11098437 TI - Coupled electromagnetic and thermal modeling of microwave oven heating of foods. PMID- 11098438 TI - Dielectric properties of wood from 2 to 3 GHz. AB - Many applications of microwave energy to wooden materials have been developed in the last few decades, both for treatment and for diagnostic purposes. All these applications require a reliable estimation of the permittivity of the wood species of interest, which is the physical parameter of crucial importance in the absorption of electromagnetic energy. This paper presents results obtained in the dielectric characterization of five wood species in the frequency range from 2 to 3 GHz, including the ISM frequency of 2.45 GHz. Permittivity was measured by an open-ended coaxial-line probe of new design on wood samples conditioned at several moisture levels. The influence of the natural variability of wood characteristics on the measured permittivity was also investigated by a suitable experimental setup consisting of a poplar table including both sapwood and heartwood regions. Finally, a theoretical discussion on the meaning of a scalar measurement on anisotropic dielectrics is conducted in terms of an isotropic equivalent permittivity, which is related to the permittivity tensor of the dielectric material. PMID- 11098439 TI - The effect of microwave radiation on the magnetic properties of minerals. AB - The effects of microwave radiation on the magnetic properties of common ore minerals are discussed. The effects of varying microwave power levels on heating rates are presented along with comparative magnetic susceptibility surveys for both treated and non treated minerals. Various chemical and physical analysis techniques are considered to quantify any changes in mineral phases during heating. Conclusions are made as to the possible impact of microwave pretreatment on the downstream magnetic processing of minerals and ores. PMID- 11098440 TI - Simulation of RF coronas using the FE-FCT method. AB - The precursor of arcs in radio frequency heating systems, namely corona discharges, are examined numerically in this paper, using the finite element-flux corrected transport technique. A point-plane configuration is used as a first approximation for a radio frequency applicator and the effects of the operating frequency and pressure on the current output, charge densities, corona onset voltage and light output are investigated. It was found that an increase in the operating frequency around the radio frequency part of the spectrum, resulted in an increase in the corona onset voltage, which accounts for the less arc-prone behavior of microwave systems. Also, a decrease in the pressure resulted in a reduction of the corona onset voltage, and thus the increased possibility of arc formation in radio frequency systems operating under vacuum conditions. PMID- 11098441 TI - Thermal modeling of microwave heated packed and fluidized bed catalytic reactors. AB - Thermal models of small-scale, microwave-heated, packed-bed and fluidized-bed catalytic chemical reactors were developed to investigate the possibility of selectively heating the catalyst sites or the catalyst pellets with microwaves. Results indicate catalyst sites may be selectively heated under special conditions in a packed or fluidized bed, and catalyst pellets may be heated above the temperature of the cooling(and reacting) gas under certain conditions in a fluidized bed. PMID- 11098442 TI - New method for moisture content measurement using phase shifts at two microwave frequencies. AB - A new method to measure moisture content of material is described in this paper. The method is based on two phase shifts of transmitted microwave signal at two different frequencies. The result using this method for moisture measurement of timber indicates this method being effective. The maximum error and mean value of error in moisture range from 2 to 30% on dry basis are 4.1% and 1.9%, respectively. PMID- 11098443 TI - Complex high-frequency technology for protection of grain against pests. AB - The results of experimental investigation of physical methods are presented for suppressing of biological activity of grain and grain product pests: harmful insects at each developmental stage except eggs (Insecta), mites (Arachnida, Acariformes) and microscopic fungi and bacteria. The technologies under development for disinfestation and disinfection of grain are based on irradiation of grain by modulated pulses of high-frequency (HF) electromagnetic fields and on simultaneous action of a complex of factors: vacuum and HF-field induced plasma. The threshold value of the electric field intensity for total insect mortality was found to be E = 4.0-5.0 kV/cm in the pulse mode at the base frequency of 47.5 MHz. When the combined technology is used, conditions are created in the irradiation chamber for HF-discharge and plasma formation, which are very strong factors influencing the biological organisms. These raise the energy (and cost) efficiency (approximately $2-3 per tonne of grain) of the combined technology for destruction of grain pests with complete environmental safety. PMID- 11098444 TI - Dielectric properties of emulsions and suspensions: mixture equations and measurement comparisons. AB - Dielectric properties of water-in-oil emulsions, oil in water emulsions and limestone-in-water suspensions have been measured at 2.45 GHz by an open-ended coaxial-line probe. The results were compared to various equations for the dielectric properties of mixtures. The equation by Fricke and Mudgett describes best the behavior of oil-in-water emulsions and limestone in water suspensions. For water-in-oil emulsions the equation by Lichtenecker and Rother gives the best results. PMID- 11098445 TI - Simplified theory of microwave drying of alkali metal silicate solutions with arbitrary values of SiO2/M2O mole ratio. AB - In this work an extension of the simplified theory of microwave drying of silicate solutions to arbitrary values of SiO2/M2O mole ratio was introduced, and good agreement between theoretical results and new experimental data for a broad range of values of SiO2/M2O mole ratio was obtained. PMID- 11098446 TI - Another perspective on child advocacy. PMID- 11098447 TI - Insights on immunizations from caregivers of children receiving Medicaid-funded services. AB - ISSUES AND PURPOSE: Despite numerous programs aimed at improving immunization rates among American children, under-immunization remains a significant problem. This study was conducted to gain insight into parents'/guardians' knowledge and attitudes regarding childhood immunizations. DESIGN AND METHODS: Thirteen African American mothers and grandmothers participated in semistructured, audiotaped focus-group interviews. RESULTS: Four major themes emerged: health knowledge and beliefs about immunizations, system barriers that impede obtaining immunizations, facilitators that enhance obtaining immunizations, and suggestions for change. PRACTICE IMPLICATIONS: Immunizations are one of the most important health advantages available to children. Therefore, nurses must become aware of the problem of underimmunization and work to address some of the concerns caregivers have identified in this study. The health and lives of the nation's children depend on it. PMID- 11098448 TI - "Love is all you need"? Student nurses' interest in working with children. AB - ISSUES AND PURPOSE: To investigate the impact of comprehensive nursing education on the future career interests of undergraduate nursing students, with particular emphasis on attitudes toward working with children. DESIGN AND METHODS: Descriptive design with surveys of 793 undergraduates in university schools of nursing throughout Victoria, Australia. RESULTS: Working with children is the most popular of nine possible career choices. Student explanations reflected a love of, and desire to work with, children, suggesting a relatively naive and uninformed view. PRACTICE IMPLICATIONS: In view of the problems experienced in the recruitment and retention of adequate numbers of pediatric nurses, there is an urgent need to promote a more realistic view of this area of practice. PMID- 11098449 TI - Chronic intestinal pseudo-obstruction: pediatric case presentations and review of the literature. AB - ISSUES AND PURPOSE: Chronic intestinal pseudo-obstruction (CIP) is a rare condition characterized by small bowel dysmotility. Its effects are severe and disabling in pediatric clients. The purpose of this article is to provide an overview of CIP and summarize information useful to pediatric nurses. CONCLUSION: Nursing management of the pediatric client with CIP is challenging, not only in terms of direct care provided to the child, but also in ongoing support and education of the child and family. PRACTICE IMPLICATIONS: Nurses practicing in either inpatient or outpatient settings may encounter children and families dealing with this disorder. Nurses are in a key position to educate others and influence the outcomes of care provided to children with CIP and their families. PMID- 11098450 TI - The healthcare needs of children in foster care. PMID- 11098451 TI - Benchmarking: what's in it for nurses? PMID- 11098452 TI - [Dispute regarding prescription of prisms]. PMID- 11098453 TI - [Combined pars-plana-vitrectomy and tectonic keratoplasty; indications for and results from 15 patients]. AB - BACKGROUND: Acute endophthalmitis requires a vitrectomy. Vitrectomy and autokeratoplasty has been reported, if the infection originates from a stromal keratitis in an aphakic eye. This retrospective non-randomized cohort study points out the requirements, indications and results of combined keratoplasty and vitrectomy in keratitis and endophthalmitis compared with noninfectious corneal and vitreoretinal problems. PATIENTS AND SURGERY: In 1995-1999, a vitrectomy and keratoplasty was performed on 15 patients (16 eyes), 10 of these with an endophthalmitis (median 71 years) and a follow-up of 2-60 months (median 19.3 months). 14 of 15 patients had had multiple prior surgery. Stromal keratitis as a sequela of keratoplasty was seen in 5 eyes (3x ruptured suture), 5x diffuse infiltration in compromised corneas (1x with a perforation, 2x with Fuchs' corneal dystrophy, 3x postoperative). In the patients without endophthalmitis 6 eyes were aphakic with corneal scars and no fundus visualization. Five eyes had a retinal detachment, one had an intraocular foreign body. An allogeneic keratoplasty was done in 14, and an allogeneic sclerokeratoplasty and an autologous sclerokeratoplasty in one eye each. RESULTS: Keratoplasty without keratoprosthesis allowed for fundus visualization, and a pars plana vitrectomy was done with a wide angle contact lens, 8x with C2F6-, 1x with silicone oil 5000 cs instillation, and gentamicin and 15 micrograms r-tPA added. In 5 vitrectomy specimens (50%) pathogenic bacteria were found. No recurrences of infection were seen. Conservation of the eyes and postoperative fundus visualization was possible in each case. The postoperative increase in visual acuity of 0.1 or better was significant in both patient groups. 2 eyes remained at preoperative levels, 14 ameliorated by > 1 lines. Complications were 1x directly postoperative graft decompensation, 1x rejection after 40 months, 6x persisting secondary glaucomas, 2x hypotony syndromes, 1x with phthisis and enucleation, 1x epiretinal gliosis. CONCLUSIONS: Curative surgery of acute keratitis and endophthalmitis by vitrectomy and keratoplasty may result in similarly successful outcomes as in noninfectious corneal scars and vitreoretinal pathology, if some requirements (e.g. adequate antibiotic treatment, graft material, skilled anterior and posterior segment surgeon) are fulfilled. PMID- 11098454 TI - [Clinical and histologic features of 141 primary basal cell carcinomas of the periocular region and their rate of recurrence after surgical excision]. AB - BACKGROUND: The aim of this study was to assess clinical and histologic features of primary basal cell carcinomas of the periocular region and to calculate the rate of recurrence of these tumors after surgical excision without intraoperative frozen section control. METHODS: All primary basal cell carcinomas of the periocular region which were treated by surgical excision at the University Eye Hospital and Clinic of Ulm between 1993 and 1998 were reviewed and special attention was paid to age and sex of the patient and localization and histology of the tumor. The recurrence rates were calculated depending on whether the margins were free or involved on histologic examination of the excised specimens. RESULTS: We could include 137 patients with 141 tumors into this study. In 92 patients with 95 tumors, follow-up data of at least 3 months was available with a mean follow-up period of 31.3 months. The mean age of the patients was 68.9 years, range 28.7-91.3 years. The tumor was located at the lower eyelid, medial canthus, upper eyelid and lateral canthus in 63.1%, 29.8%, 5.7% and 1.4%, respectively. The solid (53.9%) and the solid-cystic (17.7%) type were the most common encountered histology followed by the mixed type with sclerosing foci (12.1%) and the morphea or sclerosing type (10.6%). A free margin on histologic examination was achieved in only 47.6% of cases. The overall recurrence rate was 9.5% [definite recurrences: 6 cases (6.3%); questionable recurrences: 3 cases (3.2%)]. With histologically complete excision the recurrence rate was only 2.3% (1 case) and in the case of an incomplete excision definite recurrent tumors were observed in 11.8% (6 cases) and questionable recurrences in 3.9% (2 cases) of cases during the follow-up period. CONCLUSIONS: Surgical excision of basal cell carcinomas of the periocular region without utilizing intraoperative frozen section control or Mohs' micrographic surgery is associated with a high rate of positive tumor margin on histologic examination. However, our study showed that this is only infrequently followed by local recurrences. This result may raise the question whether simple surgical excision is still an option in the case that more cumbersome methods such as Mohs' micrographic surgery or intraoperative frozen section control are not available. However, with positive tumor margin on histologic examination of the excised tissue or with the presence of a sclerosing type regular follow-up is mandatory. PMID- 11098455 TI - [Blood-aqueous barrier after phacoemulsification with posterior chamber lens implantation; foldable acrylate lens vs PMMA lens-- a clinical study on 46 eyes]. AB - BACKGROUND: Cataract surgery leads to a more or less pronounced postoperative inflammation due to breakdown of the blood-aqueous barrier. This alteration of the blood-aqueous barrier can be reduced by minimally invasive surgery. The purpose of this study was to quantify the early course of the postoperative alteration of the blood-aqueous barrier following phacoemulsification with implantation of conventional PMMA posterior chamber lens (IOL) in comparison with foldable acrylic lens implantation. PATIENTS AND METHODS: Forty-six eyes of 46 patients (age 63 +/- 8.8 years) without preexisting deficiences of the blood aqueous-barrier or previous intraocular surgeries were divided into two groups: group 1 (24 patients): phacoemulsification with one-piece-PMMA-IOL implantation (6.5 mm corneoscleral tunnel incision); group 2 (22 patients): phacoemulsification with foldable acrylic-IOL implantation (3.5 mm incision, 15 patients with corneoscleral tunnel and 7 patients with clear cornea incision). All surgical procedures were performed by one surgeon. The postoperative treatment was standardized. Alteration of the blood-aqueous barrier was quantified by the laser flare-cell meter (Kowa, FC-1000) preoperatively and on the first and the second day after surgery. RESULTS: Preoperative aqueous flare values (photon counts/ms) were comparable in both groups (6.7 +/- 2.7 versus 5.6 +/- 2.7 respectively, p = 0.1). On day 1, aqueous flare in group 1 (9.7 +/- 2.9) was not statistically significantly higher than in group 2 (9.2 +/- 2.2, p = 0.2) and remained relatively constant on day 2 after surgery (9.3 +/- 3.3), whereas the aqueous flare values in group 2 decreased statistically significant (6.7 +/- 2.3, p = 0.01). Postoperatively, there was no statistically significant difference of aqueous flare values between eyes with corneoscleral tunnel incision and eyes with clear corneal incision (p = 0.7) in group 2. CONCLUSIONS: Our study shows that phacoemulsification with foldable IOL implantation leads to a mild and short-lasting alteration of the blood-aqueous barrier. Thus, implantation of foldable IOL may be useful in eyes especially with preexisting alteration of the blood-aqueous-barrier. PMID- 11098456 TI - [Central corneal thickness in normal eyes, patients with ocular hypertension, normal-pressure and open-angle glaucomas--a clinical study]. AB - BACKGROUND: The relation between Goldmann applanation tonometry and central corneal thickness (CCT) was investigated in several studies during the last thirty years. It was the aim of the present study to evaluate CCT in normals, patients with ocular hypertension, low-tension, and open-angle glaucomas. PATIENTS AND METHODS: CCT was measured in 135 normal eyes, 137 with ocular hypertension, 65 with low-tension, and 94 with primary and secondary open-angle glaucomas using the AL-11000-pachymeter (Tomey). The results were compared using the unpaired t-test. RESULTS: CCT was significantly higher in the patients with ocular hypertension (586 +/- 43 microns) than in the normal group (566 +/- 37 microns, p < 0.0001), in low-tension glaucomas (555 +/- 46 microns, p < 0.0001), and in open-angle glaucomas (558 +/- 31 microns, p < 0.0001). The latter three groups did not differ significantly. There was no significant correlation between CCT and age, the actually measured IOP, the highest IOP in the patient's history, or the spherical equivalent. CONCLUSIONS: Only patients with ocular hypertension showed a significant difference in CCT compared with normals. Pachymetry thus should be conducted in those patients to avoid overestimation of the IOP by applanation tonometry. In most of the patients with low-tension and open-angle glaucomas however, CCT regarded without other parameters (e.g. corneal or scleral rigidity) plays a minor role in detection of elevated IOP according to the results of this study. PMID- 11098457 TI - [Age-correlation of blood pressure induced myogenic autoregulation of human retinal arterioles in 40 volunteers]. AB - PURPOSE: For the first time noninvasive measurement of the myogenic response of retinal arterioles is possible by the use of the Retinal-Vessel-Analyzer (RVA). This study investigates the influence of age on retinal autoregulation. PATIENTS AND METHODS: In 40 healthy volunteers continuous measurement of a retinal arteriole over a 9-min period was performed. After a 3-min baseline measurement isometric exercise caused a rise in blood pressure of 15-30 mm Hg. During the last 3 minutes recovery was documented. Volunteers were split into a group of 20 younger (18-34 yrs.) and 20 older subjects (35-57 yrs.). For statistical analysis the Mann-Whitney test was used. RESULTS: All volunteers showed a rise in blood pressure. Significant differences in the blood-pressure rise were followed by a myogenic response of 1% up to 10% vasoconstriction. An individual autoregulation curve was defined with negative gradient. Significant differences between the younger and the older volunteers can be identified by this methodology. CONCLUSION: Age has a significant influence on the myogenic response in human retinal arterioles. Age should be considered as a factor in clinical studies on retinal autoregulation. PMID- 11098458 TI - [Retinal vessel reaction to 100% O2-breathing--functional imaging using the retinal vessel analyzer with 10 volunteers]. AB - BACKGROUND: Retinal vessel diameter assessment is complicated by various components among them dynamic changes due to vasomotoric effects. Measurement of these diameters was usually obtained from fundus photographs. Functional diameter changes induced by external stimuli were difficult to evaluate because of their dynamic nature. The Retinal Vessel Analyzer (RVA) allows continuous on-line measurement of those dynamic changes. Whether functional changes due to 100% O2 breathing can be assessed by RVA is investigated in this study. MATERIALS AND METHODS: Continuous on-line registration of retinal arterial and venous branch vessels was obtained in 10 healthy volunteers. A baseline was taken during the first minute. Then for 5 minutes 100% O2 was delivered by mask. Further recording ensued for 4 minutes, while breathing room air. Vessel diameter change in percent to baseline was calculated for each individual and for a mean of the group. RESULTS: Each individual demonstrated vasoconstriction. The mean diameter reduction for the group was 6.5% for arteries and 15% for veins. CONCLUSIONS: RVA allows assessment of functional retinal branch vessel reactions. Retinal branch vessels diameters are denominators for capillary perfusion. RVA might be able to demonstrate an individual vessel's regulation potential by purposeful stimulation to constrict and dilate. This property could be helpful in understanding pathophysiologic processes as well as improving diagnosis and therapeutic effects in diseases influencing ocular perfusion such as diabetes, retinal vessel occlusion or even glaucoma. Further evaluation of effects of systemic diseases might be an additional application of functional retinal vessel diameter assessment by RVA. PMID- 11098459 TI - [Circumscribed choroidal granulomatous inflammation after perforating injury. A histopathological study of four eyes]. AB - BACKGROUND: This study was performed to describe the clinicopathologic features of patients wo showed circumscribed choroidal granulomatous inflammation after trauma. METHODS: We examined histologically 4 eyes which had been enucleated within 4 weeks after treatment for perforating trauma. The second eyes of the patients were not affected. RESULTS: Four enucleated eyes with perforating injuries had focal uveal granulomatous inflammation of the posterior choroid. None of these eyes contained Dalen-Fuchs' nodules. One eye had a disrupted lens without lens-induced inflammation. All eyes exhibited choroidal ruptures. Foreign material could be detected in one of these eyes. CONCLUSIONS: Focal choroidal granulomatous inflammation may occur as a result of penetrating ocular trauma. The origin of this condition is unknown, although a foreign body reaction and choroidal rupture may be involved in the pathogenesis of the granulomatous inflammation. PMID- 11098460 TI - [Surgical transvenous embolization of spontaneous carotid cavernous sinus fistulas in two patients]. AB - BACKGROUND: Arteriovenous communications in which blood flows from meningeal branches of the internal and external carotid arteries into the venous circulation around and in the cavernous sinus are termed spontaneous (dural) carotid sinus cavernous fistulas. Due to their mostly low shunt volume they are rarely life threatening, but without treatment they may cause severe ocular complications like episcleral secondary glaucoma, central vein occlusion or exudative retinal detachment. Traditional therapy is the transarterial approach by an interventional neuroradiologist. If such an approach is not possible or unsuccessful a transvenous route has to be considered. PATIENTS AND METHODS: Two patients underwent anterior orbitotomy via sub brow incision or infraciliary incision with cannulation of the superior ophthalmic vein or the inferior ophthalmic vein and embolization of the cavernous sinus with platinum coils. RESULTS: Successful closure was achieved on angiography and normalisation of clinical symptoms after a short period of progressive venous congestion. CONCLUSIONS: For arteriovenous fistulas that cannot be embolized arterially the surgical transvenous orbital route may work as a method of second choice. When performed by an interdisciplinary team (orbital surgeon, interventionell neuroradiologist) it is a technically straightforward, effective and promising approach. PMID- 11098461 TI - [Hereditary occlusive cerebroretinal vasculopathy in two sisters]. AB - BACKGROUND: Retinal microvascular abnormalities associated with multiorgan disease may be observed in a number of systemic illnesses and syndromes. PATIENTS: Two sisters with identical signs of a hereditary cerebroretinal vasculopathy (occlusive retinal angiopathy--cerebral vasculopathy--microcephalus) were treated at the University Hospital of Saarland. COURSE: Photocoagulation for treatment of neovascular complications secondary to retinal ischemia was performed. In one eye a vitrectomy became necessary because of persistent vitreal hemorrhage. One sister died because of non treatable intracranial hypertension at the age of 22 years. CONCLUSIONS: Interdisciplinary work-up is important in patients with cerebroretinal disease. Neuropathologic evaluation including brain biopsy and neuroimaging plus ophthalmoscopy and pedriatic findings lead to the diagnosis of this rare hereditary, in this case most likely autosomal recessive condition. Photocoagulation may limit neovascular complications secondary to retinal ischemia. A specific form of treatment has, however, not yet been found. PMID- 11098462 TI - [Walker-Warburg syndrome in adulthood?]. PMID- 11098463 TI - [Instrument and pump for relief of retinal detachment in rotation of macula]. PMID- 11098464 TI - Intercept-PCR, an improvement for elevating performance to find a new member of a certain gene family. AB - We have established a method by which the performance of reverse transcriptase coupled polymerase chain reaction (RT-PCR) for seeking a new gene is improved. The actual procedure is quite easy: it is only to add several specific oligonucleotides into the reaction mixture of the usual RT-PCR. To verify the effectiveness of this method is also easy: it is only to detect the PCR products in the preliminary experiment. The finding in the present study provides valuable information for gene cloning tactics. PMID- 11098465 TI - Substitution of Glu-59 by Val in amidase from Pseudomonas aeruginosa results in a catalytically inactive enzyme. AB - A mutant strain, KLAM59, of Pseudomonas aeruginosa has been isolated that synthesizes a catalytically inactive amidase. The mutation in the amidase gene has been identified (Glu59Val) by direct sequencing of PCR-amplified mutant gene and confirmed by sequencing the cloned PCR-amplified gene. The wild-type and altered amidase genes were cloned into an expression vector and both enzymes were purified by affinity chromatography on epoxy-activated Sepharose 6B-acetamide followed by gel filtration chromatography. The mutant enzyme was catalytically inactive, and it was detected in column fractions by monoclonal antibodies previously raised against the wild-type enzyme using an ELISA sandwich method. The recombinant wild-type and mutant enzymes were purified with a final recovery of enzyme in the range of 70-80%. The wild-type and mutant enzymes behaved differently on the affinity column as shown by their elution profiles. The molecular weights of the purified wild-type and mutant amidases were found to be 210,000 and 78,000 Dalton, respectively, by gel filtration chromatography. On the other hand, the mutant enzyme ran as a single protein band on SDS-PAGE and native PAGE with a M(r) of 38,000 and 78,000 Dalton, respectively. These data suggest that the substitution Glu59Val was responsible for the dimeric structure of the mutant enzyme as opposed to the hexameric form of the wild-type enzyme. Therefore, the Glu59 seems to be a critical residue in the maintenance of the native quaternary structure of amidase. PMID- 11098466 TI - Extraction of total RNA from a high pigment content plant. Marigold (Tagetes erecta). AB - The Mexican marigold (Tagetes erecta) produces inflorescences of intense yellow color that contain high levels of xanthophylls, particularly lutein, which makes it a suitable model for the study of carotenoid biosynthesis and regulation throughout the development of the inflorescences. However, these studies require the recovery of total RNA from floral buds and inflorescences at different developmental stages, each of which presents specific extraction problems. Four protocols were tested, but only through the modification of one of them was it possible to obtain total RNA of sufficient quality and quantity to perform RT-PCR and Northern blots and to construct a cDNA library. This article presents the modified protocol for the recovery of total RNA from carotenoid-rich plant tissues. PMID- 11098469 TI - Transcriptional activation analysis using bioluminescent reporter assays. AB - Analysis of promoter and enhancer DNA sequences provides the researcher with valuable information regarding the expression patterns of genes. Insertion of small DNA fragments containing the regulatory sequence of interest into vectors carrying reporter genes allows for the accurate quantitative analysis of the gene's expression patterns and responses to various stimuli. The use of bioluminescent reporter genes provides a simple, rapid, and inexpensive system that generates virtually no toxic or radioactive waste products. In addition, bioluminescent reporter vectors are more sensitive than previous methods such as the chloramphenicol acetyl transferase (CAT) systems, that require the use of hazardous chemicals and isotopically labeled reagents. PMID- 11098468 TI - Plant transformation technology. Developments and applications. AB - Plant transformation has its roots in the research on Agrobacterium that was being undertaken in the early 1980s. The last two decades have seen significant developments in plant transformation technology, such that a large number of transgenic crop plants have now been released for commercial production. Advances in the technology have been due to development of a range of Agrobacterium mediated and direct DNA delivery techniques, along with appropriate tissue culture techniques for regenerating whole plants from plant cells or tissues in a large number of species. In addition, parallel developments in molecular biology have greatly extended the range of investigations to which plant transformation technology can be applied. Research in plant transformation is concentrating now not so much on the introduction of DNA into plant cells, but rather more on the problems associated with stable integration and reliable expression of the DNA once it has been integrated. PMID- 11098470 TI - New insights into protein-DNA interactions obtained by electron microscopy. AB - The electron microscopic study of DNA-protein complexes can yield valuable information that is often not easily available by other methods. In this article we give a number of examples that were chosen to illustrate the utility of electron microscopy. Along with the strategy used are protocols that allow such experiments to be carried out. The first example employs the following strategy. Points of close proximity between nucleic acid and protein within a bacteriophage or virus are made permanent by crosslinking. Bacteriophage or virus are then partially disrupted so that individual components can be visualized. With bacteriophages, such experiments show which DNA end first enters the host on infection and therefore can in principle indicate which phage genes would be first available for transcription. This type of experiment can also show which DNA end is first to be encapsulated during formation of the bacteriophage. Information on direction of encapsulation and indirectly, direction of replication of the rolling circles that lead to concatermeric DNA to be encapsulated, can also be derived. Such experiments can additionally accurately define the degree of DNA permutation, if present, within a bacteriophage population. Finally, examples are shown for in vitro reactions involving DNA, RecA, RecO, RecF, RecR, and SSB that lead to a further understanding of recombinational repair. Additionally antibody-gold labeling is used to locate various proteins in such complexes. PMID- 11098471 TI - Fighting the 'silent epidemic'. PMID- 11098472 TI - We shouldn't minimize vision concerns. PMID- 11098473 TI - Keeping a virtual eye on employees. PMID- 11098467 TI - Recombinant protein expression in Pichia pastoris. AB - The methylotrophic yeast Pichia pastoris is now one of the standard tools used in molecular biology for the generation of recombinant protein. P. pastoris has demonstrated its most powerful success as a large-scale (fermentation) recombinant protein production tool. What began more than 20 years ago as a program to convert abundant methanol to a protein source for animal feed has been developed into what is today two important biological tools: a model eukaryote used in cell biology research and a recombinant protein production system. To date well over 200 heterologous proteins have been expressed in P. pastoris. Significant advances in the development of new strains and vectors, improved techniques, and the commercial availability of these tools coupled with a better understanding of the biology of Pichia species have led to this microbe's value and power in commercial and research labs alike. PMID- 11098474 TI - Practical application of an Aaberg hood. PMID- 11098475 TI - The NIOSH QA rule. PMID- 11098476 TI - Safety comes from the top at Mollenberg-Betz. PMID- 11098477 TI - How to select a full-body harness. PMID- 11098478 TI - A recipe for training. PMID- 11098479 TI - Faceshield fundamentals. PMID- 11098480 TI - Software utilization. Getting your money's worth. PMID- 11098481 TI - Why GFCIs fail. PMID- 11098482 TI - Meeting the refrigerant challenge. PMID- 11098483 TI - Brownfield remediation. PMID- 11098484 TI - UFT: mechanism of drug action. AB - The mechanism of action of fluorouracil (5-FU) and the oral fluoropyrimidines and the importance of biochemical modulation and inhibition of dihydropyrimidine dehydrogenase for oral application of the prodrugs of 5-FU are discussed. The regulation of thymidylate synthase, as well as its roles as a target for antineoplastic activity and in drug resistance, are also mentioned. The more downstream events of 5-FU action and their implication in 5-FU resistance, with the possible involvement of the Fas/FasL system and the proteins associated with apoptosis regulation, are highlighted. Also discussed is the suggestion that the oral 5-FU prodrug tegafur and uracil (UFT) and its metabolites function as an angiogenesis inhibitor. PMID- 11098485 TI - Oral DPD-inhibitory fluoropyrimidine drugs. AB - Over the past decade, increasing data have emphasized both the importance of dihydropyrimidine dehydrogenase (DPD), the initial, rate-limiting enzyme in the catabolism of fluorouracil (5-FU), and its role as a control step in 5-FU metabolism, regulating the availability of 5-FU for anabolism. It is now clear that DPD also accounts for much of the variability observed with therapeutic use of 5-FU, including variabilities in 5-FU levels over a 24-hour infusion, interindividual pharmacokinetics, bioavailability, toxicity, and drug response (resistance). This variability makes effective dosing of 5-FU and related drugs difficult. In order to lessen this variability, and potentially improve 5-FU pharmacology, the pharmaceutical industry has made an effort to develop DPD inhibitors to modulate 5-FU metabolism, which has resulted in the creation of a new subclass of orally administered fluoropyrimidines, known as DPD-inhibiting fluoropyrimidines (DIF). Four drugs--uracil and tegafur (UFT) or the combination of UFT and leucovorin, ethynyluracil (eniluracil), S-1, and BOF-A2--have recently undergone clinical evaluation in the United States. The biochemical basis for using these drugs is reviewed. PMID- 11098486 TI - UFT/leucovorin vs 5-FU/leucovorin in colon cancer. AB - Adjuvant chemotherapy has been shown to alter the natural history of resected colon cancer. Two regimens (fluorouracil [5-FU] plus leucovorin and 5-FU plus levamisole) have been found to prolong disease-free survival and overall survival in affected patients. Previous comparisons of these two regimens indicate that 5 FU plus leucovorin may offer a small disease-free survival and overall survival advantage. Evidence that UFT (uracil and tegafur) plus oral leucovorin is associated with significant antitumor activity and has an acceptable toxicity profile makes this a logical formulation for the adjuvant treatment of colon cancer. The National Surgical Adjuvant Breast and Bowel Project Protocol C-06 is a randomized comparison of the relative efficacies of 5-FU plus leucovorin vs UFT plus leucovorin. Preliminary analysis of the toxicity findings among 1,530 evaluable patients indicates that both regimens are well tolerated and have similar toxicity profiles. PMID- 11098487 TI - UFT/leucovorin plus irinotecan in advanced or metastatic colorectal cancer. AB - UFT (with leucovorin) and irinotecan both have single-agent activity in colorectal cancer, with non-cross-resistant mechanisms of action. Combining these drugs would be anticipated to increase response rates while maintaining the advantages of a regimen based on an orally administered fluoropyrimidine. This trial aims to determine the maximum tolerated dose, side-effect profile, dose limiting toxicity, and response rate for the combination of UFT plus leucovorin, and irinotecan. The design is for an initial phase I study investigating escalating doses of UFT (250 to 350 mg/m2/d) and irinotecan (200 to 300 mg/m2) with fixed doses of leucovorin at 90 mg/d, in up to five cohorts of six patients each. UFT and leucovorin are given orally on days 1 to 14 of a 21-day cycle and irinotecan is given intravenously on day 1. The maximum tolerated dose is defined as the dose at which three of six patients in a cohort have a dose-limiting toxicity in the first cycle of treatment. In the second part of the study, the cohort below that experiencing maximum tolerated dose will be expanded up to a total of 20 patients in order to assess further the toxicity and response rate (phase II). To date, 17 assessable patients have been recruited, with a median age of 64 years (range: 35-80 years). At the first dose level (UFT 250 mg/m2/d and irinotecan 200 mg/m2), no dose-limiting toxicities were seen, and in cohort 2 (UFT 250 mg/m2/d and irinotecan 250 mg/m2), there was one grade 3 diarrhea. In cohort 3 (UFT 250 mg/m2/d and irinotecan 300 mg/m2), currently there have been two grade 3/4 toxicities (both neutropenic fever). Thus, the maximum tolerated dose has not yet been reached. Response has been assessed in the first six patients, with three having partial response, two having stable disease, and one having progressive disease (response rate 50%). We conclude that this regimen appears feasible with acceptable toxicity at these dose levels. There is also evidence of significant antitumor activity similar to that seen with other fluoropyrimidine-based combination regimens but without the requirement for indwelling catheters or inpatient admission. PMID- 11098488 TI - UFT/leucovorin combined with mitomycin-C in metastatic colorectal Ca. AB - UFT is an investigational agent that may be useful treatment for colorectal carcinoma and other cancers for which fluorinated pyrimidines are a useful treatment. The combination of continuous-infusion fluorouracil (5-FU) with leucovorin and mitomycin-C can significantly delay recurrence and may prolong life. UFT may be an attractive alternative to continuous-infusion 5-FU because three-times-daily administration of UFT results in comparable concentrations of 5 FU in the plasma. Recently, a clinical trial investigated the combination of UFT and mitomycin-C and found it demonstrated significant synergy in the treatment of advanced colorectal cancer. We describe the design of a phase II study to investigate the safety and efficacy of UFT/leucovorin combined with mitomycin-C in a larger group of previously untreated patients with metastatic colorectal cancer. PMID- 11098489 TI - Impact of UFT on tumoral TS and DPD levels in colorectal cancer. AB - This was an open lable, pilot translational clinical pharmacology study of a brief (7 day) course of UFT, 300 mg/m2/day, in combination with leucovorin, 90 mg/day, in six patients with newly diagnosed advanced colorectal cancer. The primary objectives of the study were to examine the impact of this treatment course on the UFT targets which are thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD). The rate of tumoral TS inhibition after the 7-day UFT treatment sequence varied from 5% up to 31%. UFT treatment induced a constant and variable decrease in tumor DPD activity ranging from 13% to 60%. UFT treatment induced a constant increase in uracil concentrations both in plasma and tumors. FT, 5-FU and the metabolite fluoro-beta-alanine (FBAL) were found in plasma and tumors at variable concentrations; the highest drug concentrations were those of FBAL in plasma. The present translational clinical study provides data related to the in vivo pharmacological effects of UFT with a description of its impact on cellular targets. PMID- 11098490 TI - UFT/leucovorin plus weekly irinotecan in advanced or metastatic colorectal cancer. AB - This is an open-label, nonrandomized phase I trial to determine the safety and maximum tolerated dose of irinotecan with a fixed dose of UFT plus oral leucovorin in patients with advanced or metastatic colorectal cancer. A secondary objective of the study is to determine the response rate in this patient population. Adult patients with histologically confirmed advanced or metastatic colorectal carcinoma and no prior chemotherapy for advanced or metastatic disease, or those patients who have received adjuvant chemotherapy (> 6 months prior to the study entry) will be eligible to participate. All patients must have measurable or evaluable lesions. Symptoms will be evaluated at baseline and every 6 weeks thereafter. Computed tomography scans will be performed at baseline and every three cycles of treatment to assess tumor response. A total of six cycles of treatment may be given, depending on patient tolerance. Patients will be followed for a maximum of 12 months for time to progression, following the last dose of the study drug. Tumor response will be defined using standard World Health Organization criteria. PMID- 11098491 TI - UFT plus leucovorin for metastatic colorectal cancer: Japanese experience. AB - In the United States and Europe, the combination of oral UFT plus leucovorin has been reported to produce objective responses and survival rates similar to those achieved with standard intravenous 5-fluorouracil plus leucovorin in patients with metastatic colorectal cancer, with reduced toxicity. However, because knowledge and experience with UFT plus leucovorin are relatively limited in Japan, we conducted a phase II study to evaluate the safety and efficacy of this combination in Japanese patients with metastatic colorectal cancer. For the purposes of this study, 20 patients received oral UFT 400 mg/m2/day in two divided doses (q 12 h) and a 5-mg tablet of leucovorin (q 8 h). Treatment was administered for 5 days, followed by a 2-day rest period, for a 28-day cycle. There were six partial responses (30%) and one complete response (5%) (overall response rate, 35%; 95% confidence interval, 14.1% to 55.9%). Greater efficacy of UFT plus leucovorin was demonstrated in patients with lung metastases, with a response rate of 63% (five of eight patients). Patients received a median of 4.5 courses (range, 2 to 12) of therapy. The median duration of survival was 228+ days (range, 81 to 540; six patients remain alive). Grade 3 or 4 toxicities occurred in three patients: diarrhea in two and mucositis in one. No toxicity related hospitalization was reported. In summary, this combination showed promising activity and an acceptable toxicity profile in the treatment of Japanese patients with metastatic colorectal cancer. PMID- 11098492 TI - A phase I study. UFT/leucovorin combined with paclitaxel for anthracycline pretreated advanced breast cancer. AB - Taxanes are the most active drugs in the treatment of metastatic breast and ovarian cancer. Weekly therapy with paclitaxel produces notable activity, with remarkably low toxicity. Moreover, combination therapy with paclitaxel and fluorouracil (5-FU) exhibits high activity in anthracycline-pretreated breast cancer patients: recent studies report response rates of 54% to 69%. UFT plus oral leucovorin constitutes an orally administered compound that provides activity comparable to that of intravenously administered 5-FU plus leucovorin. An open-label phase I study was initiated to determine the maximum tolerated dose and dose-limiting toxicities of the combination of weekly paclitaxel and UFT plus leucovorin administered to patients with anthracycline-resistant metastatic breast cancer. PMID- 11098493 TI - UFT/leucovorin plus bolus epirubicin and cyclophosphamide in advanced/metastatic breast cancer. AB - This article describes the design and early results of an open-label, nonrandomized phase I/II trial of oral UFT plus leucovorin therapy in combination with bolus injections of epirubicin and cyclophosphamide in patients with advanced or metastatic breast cancer. This study was designed as a cohort dose escalation study with the principal aims being to determine dose-limiting toxicity, overall toxicity, maximum tolerated dose, tumor response, and time to disease progression. Currently there are 22 patients randomized in the study. Overall response rate to date is 56% (66% in the locally advanced group). Based on the preliminary data, the dose-limiting toxicity appears to be neutropenia and the optimum dose of UFT for use in a phase II study appears to be 300 mg/m2. PMID- 11098494 TI - UFT/leucovorin plus vinorelbine combination for advanced breast cancer. AB - This phase I study was undertaken to define the maximum tolerated dose, the dose limiting toxicity, and the recommended dose of UFT plus leucovorin and vinorelbine in combination treatment of patients with metastatic breast cancer previously treated with one chemotherapy regimen. The pharmacokinetics of UFT and vinorelbine were also evaluated. Patients were treated with escalating doses of UFT and vinorelbine, given every 4 weeks. At least three patients were treated at each dose level before escalating to the next level. As of September 1, 1999, 22 patients have been treated. Eighteen patients were evaluable for antitumor response. One patient had a complete response (which was obtained after three cycles); four patients had a partial response. The area under the concentration time curve (AUC0-6 h) of 5-fluorouracil increase was more than dose-proportional. The AUC0-6 h values of fluorouracil were significantly higher than those noted for the four patients who developed dose-limiting toxicity at day 15. The removal of one administration of vinorelbine at dose levels 3 and 4 has allowed for increased UFT dosage and AUC0-6 h of fluorouracil, with no dose-limiting toxicity reported for these patients. No pharmacokinetic interaction between UFT and vinorelbine was observed. PMID- 11098495 TI - A phase I study. Preoperative UFT/leucovorin and radiation therapy in rectal cancer. AB - The use of combined modality regimens has been well established in the treatment of stages II and III rectal cancer. The most common chemotherapy regimens used include continuous-infusion 5-FU delivered with the help of a central venous catheter and the use of portable pumps. These regimens are expensive and can potentially develop line problems. The availability of the oral agent UFT in combination with oral leucovorin prompted the development of an all-oral chemotherapy regimen that could be combined with radiotherapy. At The University of Texas M. D. Anderson Cancer Center, we routinely use combined chemotherapy and radiotherapy preoperatively for the treatment of rectal cancers, and decided to conduct a phase I trial in which UFT and leucovorin was used instead of the conventional 5-FU. The preliminary results are encouraging and seem to demonstrate the feasibility of this approach. PMID- 11098496 TI - Postoperative radiation therapy for rectal cancer combined with UFT/leucovorin. AB - Postoperative combined-modality therapy with fluorouracil (5-FU) and radiation therapy is accepted practice for high-risk rectal cancer. Postoperative pelvic radiotherapy alone may improve pelvic control, but is not associated with an improvement in survival. Protracted infusional 5-FU has been associated with decreased tumor recurrence and improved survival when combined with postoperative adjuvant pelvic radiotherapy. The use of new drugs and alternative ways of administering 5-FU is desirable. UFT plus leucovorin is an oral 5-FU prodrug with efficacy equal to 5-FU plus leucovorin in metastatic colorectal cancer. A combination of postoperative adjuvant UFT plus leucovorin concurrent with radiation therapy should be feasible, and the design of an ongoing phase I trial is presented. PMID- 11098497 TI - UFT/leucovorin plus weekly paclitaxel in the treatment of solid tumors. AB - The palliation of symptoms and improvement of quality of life are important aspects of therapy in patients with incurable metastatic cancer. This article describes the preliminary results of a phase I study of uracil and tegafur, an orally available fluorouracil (5-FU) derivative combined with oral leucovorin plus weekly intravenous paclitaxel. While the daily oral dose of UFT is fixed at 300 mg/m2 plus 90 mg leucovorin on days 1 to 28, paclitaxel is escalated in 10 mg/m2 steps starting with 50 mg/m2 weekly as a 1-hour infusion. To date, 26 patients with a median age of 57 years have been entered into the protocol and have received a median 2.2 cycles of therapy. Dose level 4 (paclitaxel 80 mg/m2) has been recently completed. Major dose-limiting toxicities were fatigue syndrome (two patients) and diarrhea (five patients). Preliminary responses have been observed in three of 14 currently evaluable patients. This protocol is taking the development of protracted 5-FU administration--given orally as UFT--in combination with paclitaxel one step further, using paclitaxel in a dose-dense, weekly schedule. It is hoped that an active regimen for the outpatient treatment of solid tumors will be developed. PMID- 11098498 TI - UFT and its metabolites inhibit cancer-induced angiogenesis. Via a VEGF-related pathway. AB - Treatment with UFT for spontaneous lung metastasis of murine renal carcinoma (RENCA) after resection of the primary tumor has resulted in significant prolongation of the life span of tumor-bearing animals. UFT inhibited the growth of metastatic nodules in the lung, apparently via decreased density of microvessels in the metastatic foci. Subsequent experiments used dorsal air sac assay to directly trace newly forming microvessels. UFT abrogated the process of angiogenesis, induced by the RENCA cells, in a dose-dependent manner. The inhibitory effect appeared to originate from tegafur, a component of UFT, and from its known metabolites: fluorouracil (5-FU), gamma-hydroxybutyric acid (GHB), and gamma-butyrolactone (GBL). The inhibition of angiogenesis by UFT appeared to be a common phenomenon, also observed in other human cancer cell lines characterized by an excessive production of vascular endothelial growth factor (VEGF)--such as gastric, lung, and colon cancers. In vitro analysis revealed that 5-FU and gamma-hydroxybutyric acid regulated VEGF-dependent responses of human umbilical vein endothelial cells. Dorsal air sac assay revealed that UFT, 5-FU, and gamma-hydroxybutyric acid strongly inhibited the angiogenesis induced by recombinant human VEGF. These data suggest that the antiangiogenic activity of UFT is at least partially associated with an ability of the metabolites of UFT to interfere with VEGF-dependent responses of vascular endothelial cells. PMID- 11098499 TI - UFT plus carboplatin for head and neck cancer. AB - Cisplatin plus fluorouracil (5-FU) is widely accepted as neoadjuvant and adjuvant chemotherapy in the treatment of head and neck squamous cell carcinoma; UFT is also an active agent against this disease. In the first retrospective study, we examined the efficacy of UFT as adjuvant chemotherapy in patients with maxillary cancer. The 5-year survival rate of those treated with UFT vs those not treated was 71.4% vs 23.8%, respectively. In the second study we developed the carboplatin plus UFT regimen--as a modification of cisplatin plus 5-FU--and studied its efficacy and toxicity in patients with advanced head and neck squamous cell carcinoma. These patients received UFT plus carboplatin. The objective response rate was 53.1%; grade > or = 3 leukopenia, anemia, and thrombocytopenia were rare. These findings suggest that UFT plus carboplatin in the outpatient setting is feasible for patients with head and neck squamous cell carcinoma. PMID- 11098500 TI - A phase I study of paclitaxel, UFT, and leucovorin. AB - This phase I study examines the dose escalation of UFT given in combination with fixed doses of oral leucovorin and weekly doses of paclitaxel in patients with metastatic solid tumor malignancies (excluding colorectal cancer). There are two main objectives for this study. The first is to determine both the maximum tolerated dose and the dose-limiting toxicities of UFT when administered with leucovorin in combination with weekly paclitaxel (1-hour infusions of 80 mg/m2 for 4 weeks every 6 weeks). The second is to define the appropriate dose for phase II studies. Both UFT and leucovorin combinations, as well as paclitaxel, are known to be effective as single agents in heavily pretreated patients with a variety of solid tumor malignancies. UFT plus leucovorin provide activity comparable to that of intravenously administered 5-fluorouracil plus leucovorin, and weekly schedules of paclitaxel offer high dose intensity with limited hematologic toxicity. This combination is advantageous in its ease of administration and could be a tolerated outpatient regimen in patients with metastatic solid tumor malignancies. PMID- 11098501 TI - UFT/leucovorin in advanced squamous cell carcinoma of the head and neck administered with radiotherapy. AB - This is an open-label, nonrandomized phase I study to determine the maximum tolerated dose and dose-limiting toxicity of UFT plus leucovorin when given concomitantly with hyperfractionated radiotherapy in patients with head and neck cancer. The study period is determined by the course of radiotherapy, which is given as 1.7 Gy per fraction twice daily for 5 days (Monday to Friday) in 2 consecutive weeks, followed by 1 week of rest, and subsequently another 2 weeks of radiotherapy (Monday to Friday plus Monday to Thursday). Total duration of therapy will be 5 weeks. PMID- 11098502 TI - Evaluation of adjuvant UFT for gastric cancer. AB - In a trial of adjuvant chemotherapy with mitomycin and 5-FU followed by oral UFT for T1 and T2 gastric cancer after curative gastrectomy, there was no significant difference in survival between the treated and control (surgery alone) groups (5 year survival rate, 82.9% control vs 85.8% treated). Although not significantly different, 5-year survival for patients with T2 cancer was slightly higher in the treated group than in the control group (76.9% control vs 83.0% treated). This finding suggests that T2 cancer should remain the target of further trials of adjuvant chemotherapy. PMID- 11098503 TI - A novel weekday-on/weekend-off UFT schedule. AB - In a step toward a clinical trial, the tumor response and survival of a weekday on/weekend-off schedule of UFT was compared with its conventional daily schedule in a cancer-bearing rat model. The dose-intensive schedule--600 mg of UFT for 5 days followed by 2 drug-free days--amounts to a weekly dose similar to the conventional schedule of 400 mg/day. The weekday-on/weekend-off schedule provided increased survival and significantly greater antitumor activity than the conventional daily schedule, with no difference in adverse reactions. A study was also conducted in human subjects to measure fluorouracil (5-FU) concentrations that identified the pharmacokinetic activity during the 2 drug-free days of the weekday-on/weekend-off schedule. The plasma 5-FU concentration declined markedly after 24 hours, but the concentration in the tumor remained at a relatively high level after 2 days off the drug. A one-year clinical study evaluated the compliance and toxicity of the weekday-on/weekend-off UFT schedule as adjuvant chemotherapy for colorectal cancer. Based on the findings of all these studies, the weekday-on/weekend-off schedule for UFT as adjuvant chemotherapy for colorectal cancer can be recommended for a clinical trial. PMID- 11098504 TI - Irinotecan and UFT/leucovorin in patients with advanced cancers. AB - The combination of irinotecan and fluorouracil (5-FU) is synergistic when applied to human colon cancer cell lines in vitro and appears to be schedule-dependent: maximal activity occurs when irinotecan is administered prior to 5-FU. In this phase I study, irinotecan is administered in combination with UFT and leucovorin in patients with advanced solid tumors. Irinotecan is given as a 90-minute intravenous infusion on day 1 followed by twice-daily UFT plus oral leucovorin on days 2 through 15. Cycles are repeated every 21 days. Five patients have been treated to date; four are evaluable for toxicity. Starting doses were irinotecan 200 mg/m2/day, UFT 200 mg/m2/day, and leucovorin 60 mg/day. Preliminary results indicate that irinotecan in combination with UFT plus leucovorin is well tolerated at the initial doses (described in this article). PMID- 11098505 TI - Limited small-cell lung cancer: a potentially curable disease. AB - Patients with limited-stage small-cell carcinoma of the lung are treated with combined-modality therapy with the intent to cure. Standard therapy consists of platinum-based combination chemotherapy, thoracic irradiation, and for responders, prophylactic cranial irradiation. Despite this aggressive approach, too few patients achieve 5-year survival. In the past several years, new chemotherapeutic agents, including the taxanes and the topoisomerase I inhibitors, have demonstrated substantial activity against small-cell carcinoma. These agents are now being incorporated into clinical trials for patients with limited-stage disease. The best combination of these agents with platinum-based regimens is yet to be determined, and data supporting increased survival are awaited. Other studies are exploring thoracic radiation issues. Questions remain regarding optimal timing, dose, volume, and fractionation schemes. The most effective combination of thoracic irradiation and the newer chemotherapy agents also remains to be determined. The current approach to limited-stage small-cell carcinoma is reviewed, ongoing trials are described, and future directions are explored. PMID- 11098506 TI - Using the body's anticancer defenses to guide and improve breast cancer treatment. PMID- 11098507 TI - Vaccine studies demonstrate promise of immunotherapy to treat breast cancer and prevent recurrence. PMID- 11098508 TI - High-dose chemotherapy in poor-risk germ-cell tumors. AB - Testicular cancer is a highly curable cancer. However, 30% of patients are refractory to standard therapy and will need additional therapy. This article focuses on the use of high-dose chemotherapy in germ-cell tumors. High-dose chemotherapy use is discussed both in the refractory setting and as either first salvage or first-line therapy. Various criteria for risk assessment are also discussed. PMID- 11098509 TI - Clinical trials referral resource. Current clinical trials of the anti-CD20 monoclonal antibody rituximab. PMID- 11098510 TI - Computer technology helps radiologists spot overlooked small breast cancers. PMID- 11098511 TI - Indications for selective neck dissection: when, how, and why. AB - Selective neck dissection is a procedure that is primarily indicated in patients with clinically negative nodal disease in which there is a high risk of occult metastases. Others have advocated its use for patients with positive nodes, although under very specific circumstances and in combination with postoperative radiation therapy. The type of selective neck dissection performed varies according to the site of the primary, because the pattern of metastases is unique in each case. This review presents the author's philosophy on when, how, and why to employ the procedure, based on the location of primary cancers at oral, pharyngeal, laryngeal, cutaneous, thyroid, and salivary gland sites. PMID- 11098512 TI - Multidisciplinary management of pediatric soft-tissue sarcoma. AB - The management of pediatric soft-tissue sarcomas has improved drastically through the use of multimodal therapy. These tumors include rhabdomyosarcomas and nonrhabdomyosarcomas. Both are staged using physical, radiographic, and histologic evaluation, and both have intricate staging and grouping systems that correlate closely with prognosis. However, approaches to therapy for the two tumor types remain somewhat different. Rhabdomyosarcomas are treated primarily with chemotherapy. Surgical intervention is limited to initial biopsy, wide local excision when clear margins are feasible, and resection of residual disease. Radiation therapy is reserved for patients with persistent or recurrent disease and may be delivered by external beam or brachytherapy. Nonrhabdomyosarcomas are best treated primarily by surgical resection, although radiation and chemotherapy are now being used with some success. Another major difference concerns evaluation of lymphatics. Nonrhabdomyosarcomas in children frequently behave similarly to adult sarcomas, and less commonly involve regional lymph nodes, whereas pediatric patients with rhabdomyosarcomas often have nodal involvement necessitating surgical evaluation of regional lymph nodes as part of the staging protocol. Multimodal therapy has led to improved survival as well as better functional and cosmetic results. With further clinical trials and improved techniques such as brachytherapy and lymphatic mapping with sentinel node biopsy, we expect to continue to optimize therapy for pediatric patients with soft-tissue sarcomas. PMID- 11098513 TI - Follow-up care for cancer: making the benefits equal the cost. AB - Posttreatment follow-up is a staple of oncologic practice. Clinicians have traditionally presumed that close surveillance improves clinical outcome. However, new evidence reveals that frequent, procedure-intensive follow-up may provide no more significant benefit to patients than simpler approaches. Several recent consensus recommendations from major oncology organizations support this theory. Published surveys of clinician and institutional follow-up policies reveal significant variations in practice, with many providers continuing to use costly, unproven regimens. This review highlights current data on follow-up care for three common cancers--breast, colorectal, and prostate. These data suggest an acute need for changes leading to more rational, consistent, and efficient follow up practices. PMID- 11098514 TI - Lipid profile changes in young children treated for hypothyroidism. PMID- 11098515 TI - Children and influenza outbreaks. PMID- 11098516 TI - Influenza: a pediatrician's perspective. PMID- 11098517 TI - The epidemiology of influenza in children. PMID- 11098518 TI - Complications of influenza infection in children. PMID- 11098519 TI - Influenza vaccines. PMID- 11098520 TI - Should health care workers be immunized against influenza? PMID- 11098521 TI - Antiviral agents for influenza. PMID- 11098522 TI - Evaluation of novel influenza A viruses and their pandemic potential. PMID- 11098523 TI - Resident's column. Influenza. PMID- 11098524 TI - Heart failure in women. AB - Heart failure affects more than 5 million Americans. Each year, about 400,000 individuals develop heart failure, making it the nation's most rapidly growing cardiac problem. Almost one third of these individuals have New York Heart Association Functional Class III or IV heart failure and are faced with progressive clinical deterioration and frequent hospital admissions. These figures will continue to escalate as the population ages. The success of interventional procedures and pharmacologic therapies in the management of coronary artery disease has enabled this population to survive acute events, at the same time creating a population with chronic disease. Common etiologies of heart failure in women include coronary artery disease, myocardial infarction, and valvular disease. However, women are at especially high risk for developing heart failure due to diastolic dysfunction associated with hypertension and diabetes. Heart failure in women is best managed across the care continuum, incorporating pharmacologic agents, interventional procedures when appropriate, dietary restrictions, self-monitoring, and psychosocial support. Much of the recent literature has focused on women and heart disease. This emphasis is partly due to public misconception about women's health problems and the growing body of research distinguishing gender differences. Significant advances in therapy have been made to improve the quality and span of life for people with heart disease. Despite therapeutic advances, however, women have high mortality rates from heart disease, including heart failure due to ischemic causes. In fact, women with heart failure present differently than men and have different etiologies and treatment options. As we learn more about women and heart disease, the distinguishing differences unfold and become helpful in establishing a plan of care. PMID- 11098525 TI - Optimum bedside cardiac monitoring. AB - Correct electrode placement is critical to obtaining accurate information from any monitoring lead. The choice of lead should be based on the goals of monitoring for a specific patient population and on the individual patient's clinical situation. When using a 5-wire monitoring cable, arm electrodes should be placed on the shoulders; leg electrodes, on the lower thorax or hip area; and the chest electrode, in the desired V lead position. When using a 3-wire system, lead placement depends on which lead is desired for monitoring. If arrhythmia diagnosis is the goal of monitoring, lead V1 is the best lead; lead V6 is the next best lead. If ST segment monitoring for ischemia or reocclusion following percutaneous coronary interventions is the goal, the best lead depends on the coronary artery involved. Multiple lead monitoring is superior to single lead monitoring. If two leads are available, V1 and lead III or aVF (or a limb lead with maximal ST segment displacement) are good choices. If three leads are available, leads V1, III, and aVF are the best choices. Continuous 12-lead monitoring is available and offers several advantages. PMID- 11098526 TI - Salt sensitivity and hypertension in African Americans: implications for cardiovascular nurses. AB - Hypertension is a major public health problem in the U.S. Salt sensitivity is an important factor associated with hypertension and its complications, yet it has not been addressed in the nursing literature. Salt sensitivity is a directionally appropriate rise or fall in blood pressure when salt is added or removed, respectively. The change in blood pressure in salt-sensitive subjects occurs to a degree exceeding random blood pressure fluctuations. Salt sensitivity is present in 30% of normotensive and over 50% of hypertensive persons. It is more prevalent among African Americans, older persons, and individuals with renal insufficiency or diabetes. This paper provides nurses with an overview of salt sensitivity and its significance in hypertension. It presents conceptual and operational definitions of salt sensitivity, identifies factors contributing to its development, and describes implications for nursing practice. PMID- 11098527 TI - Spironolactone in the management of congestive heart failure. PMID- 11098528 TI - Committee of Nursing and Social Sciences of the ISHLT: an opportunity for global collaboration. International Society of Heart and Lung Transplantation. PMID- 11098529 TI - How is atrial tachycardia differentiated from atrial flutter? PMID- 11098530 TI - High sensitivity C-reactive protein: can it predict cardiovascular risk in postmenopausal women? PMID- 11098531 TI - The National Morbidity, Mortality, and Air Pollution Study. Part I: Methods and methodologic issues. AB - The Health Effects Institute, established in 1980, is an independent and unbiased source of information on the health effects of motor vehicle emissions. HEI supports research on all major pollutants, including regulated pollutants (such as carbon monoxide, ozone, nitrogen dioxide, and particulate matter) and unregulated pollutants (such as diesel engine exhaust, methanol, and aldehydes). To date, HEI has supported more than 200 projects at institutions in North America and Europe and has published over 100 research reports. Typically, HEI receives half its funds from the US Environmental Protection Agency and half from 28 manufacturers and marketers of motor vehicles and engines in the US. Occasionally, funds from other public and private organizations either support special projects or provide resources for a portion of an HEI study. Regardless of funding sources, HEI exercises complete autonomy in setting its research priorities and in reaching its conclusions. An independent Board of Directors governs HEI. The Institute's Research and Review Committees serve complementary scientific purposes and draw distinguished scientists as members. The results of HEI-funded studies are made available as Research Reports, which contain both the Investigators' Report and the Review Committee's evaluation of the work's scientific quality and regulatory relevance. PMID- 11098532 TI - Ectopic breast cancer. PMID- 11098533 TI - [Therapeutic potential of thrombopoietin]. AB - Thrombocytopenia induced by disease and/or chemotherapy is one of the major complications in the treatment of patients with hemato-oncological malignancies. The discovery of Mpl ligand, a hematopoietic growth factor that stimulates megakaryopoiesis and thrombopoiesis lineage-specifically both in vitro and in preclinical models, is an important step in the development of new treatment concepts for patients with thrombocytopenia. Two recombinant forms of Mpl ligand, recombinant human thrombopoietin (rHuTPO) and pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), are currently being tested in clinical trials. Development of these thrombopoietins has recently been constrained due to the observation of anti-thrombopoietin antibodies in some patients following thrombopoietin treatment. However, clinical trials performed so far provide significant insight into the therapeutic potential of these molecules. TPO/MGDF stimulates thrombopoiesis in cancer patients before chemotherapy and reduces the duration and sometimes the degree of thrombocytopenia following myelosuppressive chemotherapy. Despite marked thrombocytosis in some of these patients an increased risk for thromboembolic complications was not observed. Although administration of TPO/MGDF has provided some clinical benefit after autologous bone marrow transplantation, such effects have not been found in the settings of peripheral blood stem cell transplantation and aplasiogenic AML chemotherapy. The potential role of Mpl ligand in other indications such as thrombocytopenia in liver disease, plateletpheresis and stem cell expansion remains to be determined. Recent efforts in the development of thrombopoietins focus on the generation of molecules with maintained thrombopoietic activity which are not immunogenic. PMID- 11098534 TI - Admission for syncope: evaluation, cost and prognosis. AB - BACKGROUND: Despite extensive in-hospital evaluation the cause of syncope remains unexplained in up to 40% of patients. AIMS: To determine the application and cost of diagnostic tests, cost of hospital stay, success of evaluation and prognosis of patients admitted via the emergency department after syncope. METHODS: A retrospective cohort study including all consecutive patients admitted via the emergency department for evaluation of syncope between 1 January 1994 and 31 December 1998. The findings obtained from clinical history, physical examination and diagnostic tests were reviewed systematically. The costs of specific tests and hospital stay were analysed. Patients were followed until 31 December 1998. RESULTS: 127 patients underwent a median of 4 diagnostic tests (interquartile range, 3 to 6) over 12 days (IQR 8 to 17). The overall median cost of syncope evaluation was 106,728 ATS/7,756 EUR (IQR 70,860 to 143,583 ATS) per patient; the cost of diagnostic tests per patient was 6,863 ATS/499 EUR (IQR 3,345 to 11,969 ATS); hospital maintenance and in-hospital care accounted for the major part of these costs [median 97,680 ATS/7,099 EUR (IQR 65,120 to 138,380 ATS)]. At the time of hospital discharge, syncope remained unexplained in 48 patients (38%). The strength of agreement between the emergency department diagnosis and the discharge diagnosis was moderate (kappa = 0.49, 95% confidence interval 0.36 to 0.61). None of the patients had recurrent syncope or died during the hospital stay. Within the first 30 days after the index event 2 patients (2%) died due to known pre-existing diseases. CONCLUSION: The emergency department diagnosis markedly influenced the work-up of syncope, but not the cost of evaluation. The moderate diagnostic yield, high cost of in-hospital evaluation and good short term prognosis indicate the need for alternative strategies of in-hospital evaluation. PMID- 11098535 TI - Seroprevalence of ehrlichiosis, Lyme borreliosis and tick-borne encephalitis infections in children and young adults in Slovenia. AB - BACKGROUND: Little is known about the seroprevalence of ehrlichiosis in adults and much less about the same in children. METHODS: One hundred and forty-three healthy children and young adults (6-24 years of age, male to female ratio, 1:1) were assessed for the presence of antibodies to the agents of human granulocytic ehrlichiosis (HGE), human monocytic ehrlichiosis (HME), Borrelia burgdorferi sensu stricto (BB), and tick-borne encephalitis (TBE) virus in Slovenia, where tick-related infections are endemic. Antibodies to HGE and HME agents were assayed by indirect immunofluorescence, and antibodies to BB and TBE by enzyme linked immunosorbent assay. A questionnaire about tick exposure was answered by all subjects. In the event of a positive result, a detailed interview was conducted. RESULTS: Of 143 study subjects, 22 (15.4%) had detectable antibodies to HGE agent, 22 (15.4%) were positive to BB, 18 (12.6%) were positive to TBE virus (12 of these were vaccinated) and 4 (2.8%) were positive to the HME agent. The history of persons seropositive to an HGE agent had been uneventful. CONCLUSIONS: Our study documents a high seroprevalence of HGE in children and young adults in Slovenia, similar to the seroprevalence of LB and higher than that of TBE and HME. Although the majority of these infections are probably asymptomatic or mild, active surveillance for acute HGE infections in children in areas endemic for tick-related infections is necessary. PMID- 11098536 TI - Nontoxigenic sorbitol-fermenting Escherichia coli O157:H- associated with a family outbreak of diarrhoea. AB - A recent study from Germany reported the isolation of E. coli O157:H7/H- from patients with non-bloody diarrhoea and hemolytic uremic syndrome, questioning the role of Shiga toxin as the main trait of virulence for human disease. We isolated 6 sorbitol-fermenting E. coli O157:H- strains that do not contain Shiga toxin genes. The isolates originated from an outbreak (3 patients, 3 asymptomatic contacts) of non-bloody diarrhoea affecting two families sharing one household. Two children (age 10 months and 2 years) suffered severe diarrhoea over 30 and 10 days, respectively. Their uncle had moderate diarrhoea for 2 weeks. In contrast to the other isolates, the uncle's strain (EH109) did not harbour a chromosomal eae gene encoding gamma-intimin nor the plasmid gene E-hly; it also showed a PFGE pattern that was different from the unique pattern of the other isolates. Employing PFGE, phage typing, and P-gene typing, five of the six stx negative isolates were indistinguishable from the stx 2 positive "Bavarian outbreak strain". The only human serum tested, obtained from one asymptomatic contact, contained antibodies to the O157 lipopolysaccharide antigen. Our finding of five stx negative sorbitol-fermenting E. coli O157:H- isolates (harbouring eae and E hly) associated with an outbreak of non-bloody diarrhoea supports the hypothesis that Stx production is not obligatory for the pathogenicity of E. coli O157 for humans. PMID- 11098537 TI - [Importance of computer tomography in preoperative diagnostics of polyposis nasi]. AB - Computed tomography (CT) is a very important diagnostic tool prior to endoscopic nasal or paranasal sinus surgery. However, it is frequently notice during endonsal surgery of the paranasal sinuses, that the intraoperative results do not completely agree with the radiological ones. The objective of the present study was to compare clinical and operative findings with those of CT investigations. We collected 200 cases treated during the past 2 years and studied their CT reports. Paranasal sinuses were separated into six regions, and the pathological changes due to sinusitis found by CT were classified into four groups. Altogether, we found a highly significant relationship between CT and intraoperative findings (r = 0.44; P < 0.0001), but differences were found in several regions. The highest correlation was found in the anterior ethmoid bone area (r = 0.98), posterior ethmoid bone area (r = 0.53), maxillary sinus (r = 0.36) and sphenoid sinus (r = 0.35). There was only a low agreement in the case of frontal sinus and recessus frontalis. Possible factors such as time between CT and operation or inflammation are discussed. While CT is the image modality of choice in evaluating patients with chronic paranasal sinusitis, agreement to intraoperative findings is not perfect. This should be taken into consideration when planning functional endoscopic sinus surgery. PMID- 11098538 TI - Primary breast cancer of the vulva: a case report and review of the literature. AB - Since 1872, 40 cases of ectopic mammary gland tissue in the vulva have been reported in the literature. Out of these, 12 had a primary cancer in the ectopic breast tissue. Seven metastases of an orthotopic breast cancer have been found in this location. We are presenting the 20th case of cancerous breast tissue in the vulva whom we classified as the 13th case of primary cancer based on clinical and histopathological criteria of primary and metastatic malignant disease. Because of the advanced age of the patient, wide local excision followed by adjuvant hormonal therapy was opted for. Nineteen months after surgery, there is no evidence of recurrent disease. Due to the rarity of this entity, its management presents therapeutic dilemmas, and variable treatment strategies are being found in the literature. In our opinion, the same basic principles used for treatment of cancers of the orthotopic breast should be applied in ectopic breast carcinoma. PMID- 11098539 TI - [Heterotopic cystic pancreas as a growing submucous tumor close to the cardia of the stomach]. AB - Ectopic pancreas, presenting as a growing submucosal tumor, is a rarity in stomach surgery. Up to now, only about 250 cases of ectopic pancreas have been described. To our knowledge this is the first report of ectopic pancreas found near the cardia. A 52 year old female patient suffered from unclear upper abdominal distress. Gastroscopy revealed a submucosal growing tumor near the cardia. The tumor was locally exceeded and histologically examined showing ectopic, cystic pancreatic tissue without signs of malignancy. Wound healing was without complications and the patient is now, two years later, free of symptoms. The preoperative diagnosis of ectopic pancreas still is rarely conclusive. Nevertheless, new techniques such as endoscopic sonography or transmucosal biopsies may provide a non-invasive alternative to surgery. However, currently, local excision with intraoperative frozen section still is the therapy of choice. PMID- 11098540 TI - [Myocardial contrast echocardiography. Research instrument or already clinical application?]. AB - Increasing interest has been focused on myocardial contrast echocardiography (MCE) since latest imaging technologies allow the detection of echocontrast agents within myocardial structures following intravenous injections. Although numerous studies on different aspects of pre-clinical and clinical applications have been published, MCE has not yet become a standard clinical application. This paper summarises and estimates the clinical value of the recent developments in this evolving field of research, particularly with regard to the new real-time perfusion imaging technologies. PMID- 11098541 TI - [Echocardiographic online quantification of left ventricular systolic function in children: comparison with conventional off-line determination]. AB - Accurate and efficient echocardiographic on-line determination of left ventricular volume would be advantageous in the care of children with congenital heart disease and children with hemodynamic instability. The prospective study was performed to evaluate the clinical usefulness of the on-line automatic border detection system (acoustic quantification: AQ) for determination of left ventricular volumes and ejection fraction in comparison to the conventional off line method (manual tracing). 107 patients were enrolled in the study. The ages ranged from 0.1 to 18.8 years (mean 8.3 +/- 5.6). All patients were studied in the apical four-chamber plane for acoustic quantification (AQ) and manual tracing as well. Left ventricular volumes were determined using the mono-plane Simpson's rule. Left ventricular end-diastolic volumes obtained by AQ correlated well but were slightly underestimated compared to those determined by manual tracing (r = 0.99). Left ventricular endsystolic volumes by AQ correlated well but were also slightly underestimated compared to those obtained by manual tracing (r = 0.98). Mean ejection fraction was 61.1 +/- 6.8% by AQ compared with 61.5 +/- 5.9% by manual tracing. Linear regression analysis demonstrated good correlation: y = 0.77x + 14.1, r = 0.89; p < 0.001. Measurement of left ventricular volumes and ejection fraction by AQ using automatic border detection compares well with measurements done by manual tracing. However, AQ tends to underestimate to some degree. The time necessary for acquisition of data was similar in both methods. AQ seems to be a promising method for real-time estimation of left ventricular volume, even in children. PMID- 11098542 TI - [Myocardial contrast echocardiography with harmonic power Doppler and lung traversing SHU 563A ultrasound contrast medium for imaging myocardial perfusion disorders]. AB - The current approach for the assessment of myocardial perfusion using contrast echocardiography involves black-and-white gray scale imaging in b-mode. For better appreciation of perfusion abnormalities, off-line postprocessing techniques including color encoding are used. In this study, we examined whether we could exploit the contrast microbubble response to high ultrasound amplitude- the phenomenon of stimulated acoustic emission--that could be recorded with harmonic power Doppler (HPD) in color to identify myocardial perfusion defects. To assess the potential of HPD, we occluded branches of the left coronary artery for 2-3 h followed by 1 h reperfusion in 10 dogs. After transvenous administration of the new air-filled contrast agent SHU 563A, echocardiographic imaging was performed with ECG-triggered harmonic b-mode (HBM) and the harmonic power Doppler (HPD) approach in different short (SAX) and long axis (LAX) views. Post-mortem TTC staining was performed to verify infarction. HBM, HPD and TTC data were analyzed by independent observers. During coronary occlusion, HPD with SHU 563A showed perfusion defects in 10 dogs in all SAX and LAX views. HBM demonstrated perfusion defects in all dogs in SAX and in 8 dogs in LAX. The correlation of perfusion defect size between HPD and HBM images was good (SAX: r = 0.9, p < 0.001, LAX: r = 0.7, p < 0.01). One hour after reperfusion, both HPD and HBM showed perfusion defects with SHU 563A in 7 dogs. Five dogs showed TTC evidence of infarction. Perfusion defect size by HPD correlated well with residual infarct size (r = 0.8, p < 0.01), while defect size by HBM showed poor correlation (r = 0.3, p = ns). Myocardial contrast echocardiography with HPD and contrast agent SHU 563A identifies perfusion defects in acute coronary occlusion as reliably as HBM. After reperfusion HPD and SHU 563A accurately portray the site and size of residual myocardial infarction on line, in color. This approach has excellent potential for clinical application. PMID- 11098543 TI - [Assessment of myocardial vitality with dobutamine echocardiography: current review]. AB - Myocardial stunning (contractile dysfunction in the presence of normalized perfusion) and myocardial hibernation (contractile dysfunction matching reduced perfusion) have represented separate concepts of viable, but dyssynergic myocardium in the past. However, in vivo experimental and clinical work suggests that repetitive ischemia due to coronary artery disease may induce a gradual transition between stunned and hibernating myocardium. Myocardial hibernation itself can result from a spectrum of ischemic conditions ranging from impaired myocardial blood flow reserve to frank hypoperfusion. With increasing severity and duration of ischemia, degeneration of cardiac myocytes, accumulation of glycogen and cell death ensue. Additionally, there is an increase of extracellular matrix protein content leading to reparative fibrosis, which in turn limits functional recovery. In the light of these structural features, the available methods for detection of viable myocardium, in particular dobutamine echocardiography and nuclear imaging techniques, offer complementary rather than contradictory information. Dobutamine echo has satisfactory sensitivity, excellent specificity, and high diagnostic accuracy for the detection of viable dyssynergic myocardium. While in the past only its predictive accuracy for segmental recovery has been validated, newer data show an improved survival after revascularization if at least four viable dyssynergic left ventricular segments in a 16 segment model can be identified by dobutamine echocardiography. The complete (low and high dose) dobutamine protocol can elicit several types of contractile responses (sustained improvement in contraction or monophasic response, biphasic response, new wall motion abnormality) which should be interpreted in view of other clinical data including a previous infarction. The test protocol can be used safely at the end of the first week after myocardial infarction. If ischemia or viability is documented, revascularization should be performed promptly. A similar strategy should be followed in the setting of chronic coronary heart disease with left ventricular dysfunction. Since the structural changes of hibernating myocardium are progressive, time to revascularization is critical. On the other hand, responsible therapeutic planning requires proof of ischemia or viability before initiating a potentially hazardous revascularization procedure. PMID- 11098544 TI - [Reconstruction of the bicuspid aortic valve]. AB - Reconstruction of a regurgitant bicuspid aortic valve is a new alternative to aortic valve replacement. With concomitant aortic root dilatation adequate reconstruction is feasible by valve-sparing aortic replacement. Between 10/95 and 02/00, 30 patients underwent reconstruction of a regurgitant bicuspid aortic valve. Additional aortic replacement was performed in 23 cases. Valve reconstruction was performed by plication of the prolapsing leaflet. No patient died peri- or postoperatively. Freedom from aortic valve regurgitation > or = II as well as freedom from reoperation were 100% after 48 months. Reconstruction of a regurgitant bicuspid aortic valve is feasible with encouraging mid-term results. With concomitant dilatation of the ascending aorta, a combination of aortic replacement and valve reconstruction can achieve stable results even in bicuspid valve anatomy. PMID- 11098545 TI - [Indirect genotype analysis as a diagnostic procedure in Marfan syndrome]. AB - Marfan syndrome (MFS) is an autosomal dominant disorder of connective tissue characterized by skeletal, ocular and cardiovascular manifestations. The disease is caused by mutations in the FBN1 gene, encoding fibrillin, an important component of elastic fibers. Diagnosis of Marfan syndrome is currently based on detailed clinical examination and/or mutation analysis in the fibrillin gene. Clinical expression varies widely both among and within families, rendering clinical diagnosis extremely difficult. In this study, we performed segregation analysis of allelic DNA polymorphisms to support diagnosis of Marfan syndrome. This type of genotype analysis is a useful, additional diagnostic tool for families with Marfan syndrome and provides incremental information of diagnosis or exclusion of Marfan syndrome based on clinical findings. PMID- 11098546 TI - [Significance of matrix metalloproteinases in cardiovascular diseases]. AB - The cardiovascular interstitial space is largely composed of type I and III fibrillar collagens. Tissue structure, form and function are determined not only by the collagen content but also by the ratio of different collagens to each other. Matrix metalloproteinases are members of a family of secreted and membrane bound enzymes that are capable of degrading highly proteolytic resistant fibrillar type I and III collagens. Collagen tissue content is determined by balanced collagen synthesis and degradation. MMP activity and adverse tissue remodeling have been identified in coronary plaques in unstable angina. It has also been linked with the progression of aortic aneurysms and with left ventricular dilatation in congestive heart failure in patients with ischemic and non-ischemic cardiomyopathy. The role of MMPs in these cardiovascular diseases and possible therapeutic options are the focus of this review. PMID- 11098547 TI - [Aortic isthmus stenosis with open ductus Botalli as etiology of severe heart failure in a 36-year-old patient--case report of successful surgical treatment]. AB - We report on the history of a 36-year-old woman with untreated coarctation of the aorta and patent ductus arteriosus who developed refractory heart failure due to severely impaired left ventricular function. After coarctation repair and duct resection, left ventricular function improved to normal. Even in the presence of longstanding left ventricular pressure and volume overload, subsequent severe myocardial failure may be reversible by surgical repair. PMID- 11098548 TI - [Interventional therapy of aortic isthmus stenosis with concomitant thoracic aortic aneurysm with a stent graft]. AB - A 23 year old male presented with hypertension and a systolic murmur. ECG and echo revealed signs of left ventricular hypertrophy. Aortography showed aortic coarctation (delta p max. 40 mmHg) in combination with an aneurysm originating from the coarcted segment distal from the origin of the left subclavian artery. Coarctation and aneurysm were treated using direct placement of a stent graft and subsequent balloon dilatation. On final aortography, the outline of the stented segment was smooth with good stent apposition and the aneurysm entry was closed. The residual gradient was 10 mmHg (max). Six months later, the patient has no symptoms and a blood pressure of 120/80 mmHg on both arms. PMID- 11098549 TI - [Progression of an intramural hematoma to dissection]. AB - Intramural hematoma is a spontaneous, localized bleeding in the wall of the thoracic aorta without evidence of intimal tear. Clinically, intramural hematoma manifests itself as an acute thoracical pain in patients with hypertension and therefore shows parallels with the classical aortic dissection. In the literature there is controversial discussion whether intramural hematoma can be regarded as its own aortic pathology or a precursor in the development of classical aortic dissection. We present the case of a 66 year old male who showed an intramural hematoma of the descending aorta which rapidly progressed to classical dissection within 3 months. This finding was secondarily accompanied by a dilatation of the descending aorta which leads to operative treatment with replacement of the proximal aorta descendens. This case supports the hypothesis that intramural hematoma is a potential early manifestation of aortic dissection and at the same time stresses the necessity for frequent follow-up investigations and, if necessary, early operative therapy. PMID- 11098550 TI - [Comment on the contribution by Wagdi P., "Selective kidney angiography within the scope of heart catheterization; pathological findings and therapeutic consequences"]. PMID- 11098551 TI - [Is everything really so simple...? Or after all more intense than we thought?]. PMID- 11098552 TI - An overview of the evolution of the atherosclerotic plaque: from fatty streak to plaque rupture and thrombosis. AB - Today, there is a wealth of information concerning the chronology of the cellular and molecular events associated with the development of cardiovascular diseases. It is now clear that atherosclerosis is largely a result of a dysregulated fibroproliferative inflammatory response [1]. Treatment of cardiovascular disease is rapidly adapting to this new knowledge and the future holds great potential for preventing the disease by blocking the process at multiple stages. This short review provides an overview of the evolution of the atherosclerotic plaque. It focuses on three basic stages of the disease processes: initiation of the fatty streak, transition of the fatty streak to an atheroma, and progression and destabilization of the lesions leading to plaque rupture and occlusive thrombosis. It includes a discussion of the molecular and cellular biology of the atherogenic process with an emphasis on key molecular mediators of the disease process and provides three tables which list examples of some of the key molecular mediators and the stages in which they are purported to play a role. PMID- 11098553 TI - [Pathogenesis of coronary disease]. AB - Being overweight (OW) was recognized very early as a risk factor for coronary heart disease (CHD). Its significance in the pathogenesis of CHD has been strengthened by observations showing that OW is responsible for the development of diabetes, hypertension and lipid disorders due to its induction of insulin resistance (IR). Its key role has been underlined further by recent studies indicating that OW causes endothelial dysfunction via elevated serum fatty acids, which initiates the molecular events that further the process of CHD. It is, therefore, of the utmost importance to determine its roots. The most probable reason for its high incidence is due to the genetic outfit of most people which does not permit adequate adaptation of the cerebral cortex according to the environmental changes which have occurred since the early days. PMID- 11098554 TI - [New ways of imaging diagnosis of the heart with electron beam tomography]. AB - Electron beam computed tomography is a noninvasive imaging tool for diagnosing heart disease. The method offers new insights into coronary heart disease, because the quantification of coronary artery calcification provides a fast, low risk and cost efficient estimation of coronary risk with greater prognostic power than traditional risk estimates. With intravenous administration of an x-ray contrast agent, EBCT can detect coronary artery stenoses more accurately than other noninvasive methods. The quantification of myocardial blood flow by EBCT represents a realistic alternative to other methods of assessing myocardial perfusion. High temporal resolution allows for detailed analysis of cardiac dynamics. Other applications include the work-up of cardiac masses, pericardial disease, and investigations of the great vessels. As a dedicated cardiac imaging tool, EBCT provides new insights into cardiac physiology and disease, and offers a quantitative approach to many diagnostic problems. Limited availability, radiation exposure, and the need for a iodinated contrast agent still make EBCT an accessory modality at this time. PMID- 11098555 TI - [New strategies in the treatment of restenosis]. AB - Twenty-three years after introduction of coronary angioplasty (PTCA), the inhibition of restenosis formation continues to be the major challenge for the interventional cardiologist. About 35-50% of all patients undergoing PTCA develop a renarrowing of the intravascular lumen within the following six months. The use of specific systemic drug therapy as well as different angioplastic methods (rotablation, atherectomy, laser angioplasty) all failed to significantly reduce restenosis. Local drug delivery and local gene therapy have only shown to be effective in animal experiments. Restenosis can be reduced by the use of stents; however restenosis can also develop within the stents. The treatment of choice for severe in-stent restenosis may become radiotherapy, which seems to be a promising tool also for other forms of restenosis. PMID- 11098556 TI - [Neoangiogenesis by local gene therapy: a new therapeutic concept in the treatment of coronary disease]. AB - This article presents the results of our initial clinical experience with the human growth factor FGF-1 as applied to the ischemic human myocardium. After the completion of extensive preliminary animal experiments, the human angiogenetic growth factor FGF-1, obtained from a genetically transformed strain of E. coli was introduced into aortocoronary bypass surgery as an additional therapeutic agent. A double-blind study was carried out in a total of 40 patients with coronary heart disease. The patients were randomized into growth-factor and control groups, each containing 20 patients. All patients underwent aortocoronary bypass surgery because of their coronary multivessel disease, in each case with an IMA bypass for the LAD and single venous bypasses for the RCX and/or RCA. In order to bridge over additional stenosis of the LAD or one of its branches, the human growth factor FGF-1 was injected into the myocardium during the operation. In the control group heat-denatured growth factor was substituted for FGF-1. After three months as well as three years postoperatively, the IMA bypasses were selectively depicted by intraarterial DSA. These angiographies were then quantitatively evaluated. After the application of the growth factor, the formation of new vessels could be demonstrated after three months as well as three years postoperatively. A capillary network initiating from the coronary artery could be found and the computer-supported evaluation of the angiographs revealed a significant increase in the blood supply of the region of the myocardium injected. According to the angiographic findings there was also a clinical improvement of the patients with FGF-1 application compared to the patients of control group, concerning the NYHA classification as well as the need for antiangina drug therapy. In the meantime, the results of other research groups support the evidence that the induction of neoangiogenesis by human growth factor could become a therapeutic approach especially in patients with diffuse coronary artery disease. Nevertheless further studies have to be carried out in order to prove the long-term clinical profit of those patients after the growth factor treatment. PMID- 11098557 TI - [Percutaneous myocardial laser revascularization (PMR), a new therapeutic procedure for patients with refractory angina pectoris]. AB - In patients with severe angina pectoris due to coronary artery disease, who are not candidates for either percutaneous coronary angioplasty or coronary artery bypass surgery, transmyocardial laser revascularization (TMR) often leads to improvement of clinical symptoms and increased exercise capacity. One drawback of TMR is the need for surgical thoracotomy in order to gain access to the epicardial surface of the heart. Therefore, a catheter-based system has been developed, which allows creation of laser channels into the myocardium from the left ventricular cavity. Between January 1997 and November 1999, this "percutaneous myocardial laser-revascularization" (PMR) was performed in 85 patients at the Herzzentrum Leipzig. In 43 patients, only one region of the heart (anterior, lateral, inferior or septal) was treated with PMR; in 42 patients two or three regions were treated in one session. 12.3 +/- 4.3 (range 4-22) channels/region were created into the myocardium. Six months after PMR, the majority of patients reported significant improvement of clinical symptoms (CCS class at baseline: 3.3 +/- 0.4; after 6 months: 1.6 +/- 0.9) (p < 0.001) and an increased exercise capacity (baseline: 349 +/- 138 s; after 6 months: 470 +/- 193 s) (p < 0.05); however, thallium scintigraphy failed to show increased perfusion in the PMR treated regions. PMR seems to be a safe and feasible new therapeutic option for patients with refractory angina pectoris due to end-stage coronary artery disease. The first results indicate improvement of clinical symptoms and increased exercise capacity; evidence of increased perfusion in the laser-treated regions is still lacking. PMID- 11098558 TI - [Multiple myocardial revascularization on the beating heart: risks, benefits and outlooks]. AB - INTRODUCTION: Avoidance of extracorporeal circulation during beating heart surgery (OP CAB = Off-Pump Coronary Artery Bypass Surgery) for aortocoronary bypass grafting (ACBG) is gaining increasing importance in modern cardiac surgery. With the development of new mechanical stabilization devices, the revascularization of the posterior wall of the heart and the distal right coronary artery became feasible. Especially high-risk patients with multiple risk factors for open heart surgery will profit from this approach, because the negative effects of the extracorporeal circulation are avoided. METHODS: From 7/97 until 12/99 a total of 158 patients with multivessel-coronary artery disease were operated on the beating heart. In the same time course, a total of 2869 patients were operated conventionally using the extracorporeal circulation as a standard procedure. RESULTS: Due to patient selection, the two groups differ in the preoperative data concerning previous neurologic deficits. None of the patients in the OP CAB group suffered a permanent neurologic deficit after the operation, whereas in the CAB group the rate of stroke was 1.3%. The rate of temporary neurologic deficits, such as TIA (temporary ischemic attack) or PRIND (prolonged reversible neurologic deficit), was higher in the OP CAB group due to patient selection. No surgcially associated aortic dissection was seen in the OP CAB group. CONCLUSIONS: This and other studies have shown the effectiveness of OP CAB surgery concerning perioperative complications and survival rates. Especially high-risk patients with multiple risk factors for a cardiac operation profit from a beating-heart operation avoiding extracorporeal circulation. The question in how far the higher rate of reoccurring angina and the larger number of interventional treatments in OP CAB patients reported in the literature are due to the learning curve remains unanswered. Long-term studies will show whether the result of beating heart surgery is as good as the result of conventional "on bypass" surgery. PMID- 11098559 TI - Surgery of acute type A dissection: what have we learned during the past 25 years? AB - Every acute dissection involving the ascending aorta (Stanford type A) must undergo emergency surgical repair. However, the surgical techniques must vary according to the clinical presentation of the patients or the anatomical patterns observed. Furthermore, surgery is generally difficult because of the poor condition of the aortic tissues. To reduce those difficulties many technical artifacts have been described. In 1977, we proposed the use of gelatin-resorcin formalin (GRF) biological glue to reinforce the suture areas. From January 1977 to July 1999, 212 patients (pts) (152 males and 60 females) aged from 15 to 80 years (mean age: 54 +/- 11 years) underwent an emergency operation for type A aortic dissection. One-hundred-seventy-eight pts (84%) were operated on within 4 hours after being referred to the hospital. Twenty-eight pts (13.2%) had Marfan's syndrome. In 44 patients (20.7%), the aortic valve was replaced either independently (6 cases--2.8%) or by means of a composite graft (38 cases--17.9%). Because of the location of the intimal tear, the aortic replacement was extended to the transverse arch in 61 pts (28.7%). Hospital mortality amounts to 21.6% (46 pts), 25% in pts with arch replacement and 19.4% in pts without arch replacement (n.s). Analysis of hospital mortality demonstrates that the main causes of death were cardiac tamponade, neurologic disorders and visceral malperfusion. One hundred-sixty-six pts were discharged and surveyed from 5 months to 22 years postoperatively (mean follow-up: 85 +/- 66 months). During this period of time, 25 pts (15%) had to be reoperated for a total of 33 reoperations. Seven pts (28%) died at reoperation. Using univariate analysis, the presence of Marfan's syndrome (p < 0.05) and absence of arch replacement (p < 0.02) were determinant risk factors for reoperation. Emergency (p < 0.01) and thoraco-abdominal replacement (p < 0.04) were determinant riskfactors for death at reoperation. The freedom from reoperation (Kaplan-Meier, CI: 95%) is 96% (90-98), 87% (79-92), 80% (70 88), 66% (51-78) at 1, 5, 10 and 15 years respectively. A total of 39 pts (24.3%) died during follow-up. The presence of Marfan's syndrome (p < 0.01), reoperation (p < 0.02), stroke (p < 0.05), and cardiac failure (p < 0.05) were determinant risk factors of late mortality. The late survival rate (K-M. C.I.: 95%), including hospital mortality, is 71% (64-77), 66% (58-73), 56% (47-64), 46% (36 56), 37% (28-44) at 1, 10, 15 and 20 years, respectively. From our experience extending over more than 23 years, GRF glue has proved to be extremely useful, making the procedure much easier and safer. Nevertheless, many factors are of importance in the pre-, intra- and postoperative management of the patients. Cardiac tamponade and visceral malperfusion must be properly diagnosed and treated. During aortic repair, the main intimal tear must be resected. The transverse arch must be checked and replaced whenever necessary. The aortic valve should be preserved whenever possible. During CPB, perfusing the aorta in the regular antegrade manner seems to dramatically reduce the rate of malperfusion. The quality of the first emergency operation seems to have a major influence on the late results, especially concerning the rate of late reoperations and aortic ruptures. However, those late results depend also on the patient's basic condition, particularly in Marfan patients. PMID- 11098560 TI - [Heart failure and sudden cardiac death: pharmacological and nonpharmacological treatment possibilities from the viewpoint of the rhythmologist]. AB - Recent multicenter studies have shown that the implantable cardioverter defibrillator (ICD) is superior compared to antiarrhythmic agents after sudden cardiac death (SCD) in patients with congestive heart failure. Further ICD studies have to be performed for primary prevention of SCD in patients with heart failure. Primary prevention studies of SCD with Amiodarone or new class III agents (e.g., Dofetilide) were not able to lower cardiac mortality in these patients. How much the new method of biventricular pacing in patients with heart failure and left bundle branch block will reduce cardiac mortality has to be proven in future prospective trials. PMID- 11098561 TI - [Therapeutic inhibition of platelets in a acute coronary syndrome and in coronary intervention: mechanisms and clinical results]. AB - Platelets play a crucial role in the pathogenesis of atherosclerosis and especially in the final ischemic consequences such as acute coronary syndromes. Furthermore, platelets are central mediators of acute or subacute complications of coronary interventions. Therefore, therapeutic inhibition of platelet function is of major interest in cardiology. The following review describes three different therapeutic strategies for platelet inhibition and provides a representative overview on the clinical results of studies based on these strategies. First, the mechanism of acetylsalicylic acid is described and the strong meta-analytic data demonstrating a convincing positive clinical effect is discussed. Second, the mode of action of the thienopyridines is described and initial clinical results are discussed. Third, the inhibition of the platelet integrin receptor GP IIb/IIIa is described as a potent way to block the final common pathway of platelet stimulation. The structural description of GP IIb/IIIa is followed by a structural classification of the available GP IIb/IIIa inhibitors. Clinical studies, meanwhile including several thousands of patients, are discussed based on representative examples. Finally, unresolved issues regarding the various GP IIb/IIIa inhibitors, such as differences in receptor affinity and specificity, intrinsic activation and GP IIb/IIIa inhibitor induced thrombocytopenia are, discussed. PMID- 11098562 TI - Left ventricular restoration by endoventricular circular patch plasty (EVCPP). AB - The Endoventricular Circular Patch Plasty technique, developed to reorganize the left ventricular cavity after post-ischemic modification, is described as it has been used since 1984. Experience of more than 900 cases demonstrates the efficiency of the technique in terms of left ventricular shape, left ventricular performances and long-term clinical results. Particular attention is focused on very large asynergia with congestive heart failure. PMID- 11098563 TI - [Reverse remodeling by surgery--fact or fiction?]. AB - Mortality of chronic heart failure in industrial countries remains unacceptably high despite advances in medical therapy. Heart transplantation, the gold standard in the treatment of end-stage heart failure is reserved for only a few patients because of the shortage of donor hearts. Surgical alternatives to transplantation include dynamic cardiomyoplasty (CMP), mitral valve reconstruction, left ventricular reduction surgery (PLVR) and ventricular assist devices (VAD). Improved survival and objective physiologic improvement have not been documented for CMP in the treatment of dilative cardiomyopathy. Mitral valve reconstruction on the other hand shows promising results. PLVR is an innovative procedure in which the heart is surgically reduced in size and cardiac function is dramatically improved immediately after surgery. The presence of long-term effects is still unknown. VAD have been shown to be extremely effective as a short- and long-term "bridge" to heart transplantation. They are not approved for permanent support. A randomized trial in the U.S. is underway to compare the efficacy of these devices with the efficacy of medical therapy in NYHA functional class IV patients in quality of life, survival and costs. PMID- 11098564 TI - [Advances and perspectives of heart-lung transplantation]. AB - Thoracic organ transplantation has evolved as standard therapy in end-stage cardiac and pulmonary failure. Meanwhile more than 50,000 heart, 2500 heart-lung and 10,000 lung transplantations have been performed worldwide. Improvements in patient selection as well as in the postoperative management have led to 1-year survival rates of 70-90%. Long-term results are still influenced by the development of graft vessel disease, bronchiolitis obliterans syndrome and malignancies. Furthermore, indications and results of different procedures will be presented and new developments discussed. PMID- 11098565 TI - [Advances and perspectives in mechanical heart assist]. AB - Mechanical circulatory support devices were first developed to permanently replace the failing heart. Today, however, the majority of these devices are used as a mechanical bridge in patients awaiting heart transplantation. With this indication, important information on using mechanical assist devices has been assembled. We present our experience, which has been gained since 1987 in the area of patient selection, post-implant patient care and device maintenance. More than 450 patients have since been implanted with assist devices at our institution. Mechanical circulatory support may not only lead to recovery from secondary organ failure, but also to myocardial remodeling and recovery of the heart function in some patients. Additionally we report our experience with a newly developed implantable axial flow pump and discuss the possibility and costs of permanent support in some patients. PMID- 11098566 TI - [Robotic-guided techniques in heart surgery]. AB - The rigid design of conventional endoscopic instruments with limited degrees of freedom has not allowed for endoscopic cardiac surgery. Using a surgical telemanipulation system that is operated from a master console, endoscopic coronary artery bypass grafting and mitral valve repair can be performed with high precision. Closed-chest cardiopulmonary bypass systems that allow cardioplegic cardiac arrest are required. After extensive experimental testing a telemanipulation system was used clinically. PMID- 11098567 TI - Loss of sinus compliance, cause of degenerative aortic valve disease, and early failure of biological valves. AB - Loss of compliance of aortic sinuses due to aging or graft replacement causes significant aortic valve leaflet dysfunction. This event leads to stress overload and induces degenerative changes in the patient's (natural or bioprosthetic) valve and unfavorably influences leaflet longevity. PMID- 11098568 TI - [Reconstructive surgery of the aortic root]. AB - Surgical treatment of proximal aortic disease traditionally consists of composite replacement of valve and aorta. Recent reconstructive procedures on the aortic root allow for treatment of aortic dilatation and concomitant aortic valve regurgitation without the associated disadvantages of mechanical heart valves. From 10/95 to 09/99 we treated 84 patients for regurgitation of the aortic valve and dilatation of the aortic root. Valve preserving replacement of the root consisted of root remodeling (n = 68) or reimplantation of the aortic valve (n = 16). Operative mortality in valve-preserving surgery was not elevated compared to overall results of proximal aortic replacement (3.6% vs 5.6%); this applied to elective procedures (1.8% vs 2.3%) as well as emergency operations (9.3% vs 16.3%). Initial aortic valve function was adequate in all cases. Actuarial freedom from regurgitation grade II or higher was 98% after root remodeling and 92% after valve reimplantation. Freedom from reoperation at two years was 96% in remodeling and 100% in valve reimplantation. Hemodynamic function of the reconstructed valve was investigated in 17 patients under conditions of rest and exercise. These were compared to 9 patients with a mechanical composite valve. Patients with reconstructed valves had almost physiologic gradients during rest and exercise. These gradients were thus significantly lower than the increased gradients of patients after composite replacement. Application of reconstructive procedures to the aortic root allows for restoration of aortic valve function in the majority of patients. Disadvantages of heart valve prostheses can be avoided, and the hemodynamic performance of the reconstructed valve appears almost physiologic. PMID- 11098569 TI - [Progress in Oncologic Surgery: The importance of lymph node dissection and blood transfusion in oncologic surgery. Symposium proceedings. 25-26 May 2001. Biel, Switzerland]. PMID- 11098570 TI - [Lymph nodes and malignant tumors]. AB - The planning of locoregional tumor therapy (radical surgical resection, curative radiotherapy) is based on the knowledge of locoregional tumor spread, in particular lymph node metastasis. In general, lymphatic spread follows anatomic rules, skipping of nodes is observed maximally in 3%. According to tumor site, uni- or multidirectional lymph drainage is found. In some tumors (carcinoma of penis, malignant melanoma, breast carcinoma) the concept of detection and examination of sentinel node increasingly is of importance. Lymph node metastasis is to be distinguished from the finding of isolated tumor cells in the sinus of lymph nodes (tumor cell emboli). A definite diagnosis of lymph node metastasis requires a careful histopathologic examination. The incidence of regional lymph node metastasis predominantly depends on tumor type, histological grade of differentiation, lymphatic invasion and depth of invasion/tumor size/tumor volume. A careful histopathologic examination of tumor resection specimens in regard of lymph node metastasis is important for indication to additional postoperative treatment, estimation of prognosis and analysis of treatment results. Adequate quality assurance is necessary. PMID- 11098571 TI - [Significance of immunohistochemically detectable tumor cells in lymph nodes]. AB - Distant metastasis is mainly determined by the tumor biology, whereas local relapse after complete (R0) resection of solid tumors is largely determined by the effectiveness of the surgeon. The detection of a minimal tumor cell spread in lymph nodes became recently possible by the introduction of sensitive immunohistochemical and molecular methods and is increasingly considered as clinically relevant because of its independent prognostic significance. Furthermore, these tumor cells, compared to solid metastases, are appropriate targets for intravenously applied anti-cancer therapeutics because they are easily accessible for macromolecules and immunologic effector cells. Double staining analyses have demonstrated that the majority of these tumor cells stay in a non proliferating phase of the cell cycle. This phenomenon could be an explanation for the extended latency period ("dormancy") between their primary diagnosis and the occurrence of a subsequent metastatic relapse, and it may furthermore explain the failure of anti-proliferative chemotherapy. Consequently, adjuvant therapeutic strategies, which are directed also against these "dormant" tumor cells are of increasing importance after radical local tumor resection (R0). PMID- 11098572 TI - [Significance of lymph nodes in tumor surgery. Value of minimally invasive staging]. AB - Malignant tumors require an exact staging in order to initiate individual tumor related therapeutic concepts to avoid unnecessary explorative laparotomy and to compare different treatment regimes. The assessment of the lymph node status with regard to tumor involvement using any of the actual imaging methods is quite unsatisfactory. For the improvement of the pretherapeutic tumor staging including N-classification the diagnostic laparoscopy and laparoscopic sonography are presently being evaluated. Both methods should be carried out according to a standardized investigation record. When limited to the pure diagnostic aspect, the morbidity is approx. 2%. Low patient figures with different tumor entities, insufficient information on the simultaneous occurrence of lymph node and distant metastases and/or of a peritoneal carcinomatosis as well as on the extent of the lymphadenectomy and histopathologic outcomes restrict the signifying value of many studies. It seems to be only clear that, when using the laparoscopic sonography, the sensitivity of the evidence of lymph node metastases increases in comparison with the sole laparoscopy. Definite recommendations based upon the outcomes with the required evidence, can presently neither be made with regard to the use of the method in general nor for the laparoscopic lymph node staging in particular. The use with regard to a lymph node assessment from today's point of view seems to be appropriate above all in case of: Suspect of an advanced tumor stage (existence of M1 lymphomas) For the indication in case of justified application of multimodal therapeutic concepts (exact tumor staging/N classification). Beyond this, the laparoscopy for lymph node staging should only be used in conjunction with prospective randomized studies. Sufficient experience in the field of laparoscopic surgery and sonography as well as compliance with the rules of action for the prevention of tumor cell conveyance should be demanded. PMID- 11098573 TI - [Immunotherapy in malignant melanoma]. AB - The malignant melanoma is because of its ability to form metastasis even at early stages of disease the deadliest of all skin tumors. Its incidence rises faster than that of any other human tumor. Today, treatment of melanoma is based on surgical removal, and depending on the stage, chemotherapy and/or biological response modifiers. The response and cure rates are, however, not satisfactory and there is, therefore, ongoing research for other approaches. The identification of melanoma antigenic peptides like Melan-A, gp100 and ras peptides has opened new possibilities in the treatment of malignant melanoma. The research on generating mainly T cell mediated immune responses against malignant melanoma using many different approaches like injection of dendritic cells pulsed with melanoma specific peptides, injection of in vitro with cytokines stimulated T cells, immunization with peptides in the presence of different adjuvants, immunization with genetically modified melanoma cells etc. has produced many encouraging results. However, the future of tumor-vaccine development still lies in generation of more potent vaccination protocols. PMID- 11098574 TI - [Melanoma. Results with "sentinel node" sampling]. AB - Since June 1995 we have practised a gamma probe guided sentinel lymphadenectomy (SLNE) in 274 patients after injecting a colloidal 99 m-Tc labelled solution around the tumor. By this technique the detection and excision of the SLN succeeded in 99.3% of all cases. We found micrometastases in about 53.1% of patients with pT3 and pT4 melanomas. The specimen of the radical lymph node dissection contained in 30% further metastases. A regional recurrence after SLNE occurred only in one case, a SLN-negative patient. PMID- 11098575 TI - [Importance of lymph node involvement in breast carcinoma for the oncologist]. AB - Axillary node status is the most important prognostic information regarding treatment and prognosis for breast cancer patients. Localisation and number of involved lymph nodes add further information and are of prognostic relevance as well. The problem of the involvement of different axilla levels as well as skip metastases are discussed. Sentinel lymph node dissection is still considered investigational as well as axilla downstaging by preoperative chemotherapy. New prognostic markers are helpful in individual situations but are not yet considered as standard procedures. Conclusions of practical relevance are summarised. PMID- 11098576 TI - ["Sentinel" lymph node biopsy in breast carcinoma. Current experiences]. AB - Recent studies show the sentinel lymph node biopsy (SNB) as a reliable method for the determination of the nodal status in patients with breast cancer. We present our experience with this method during 3 years and discuss its potential and limitations. From 11/95 to 3/99 we performed a sentinel node detection in 146 patients with breast cancer stage I to III. We used the raionuclide method including preoperative lymphscintigraphy and intraoperative gamma.probe detection. The detection rate varied with the tumor size between 94% for tumors with a diameter < 1 cm, 85% (1-3 cm), 70% (3-5 cm) and 63% (> 5 cm). The accuracy of the SNB in the prediction of the nodal status varied also with the tumor diameter between 100% for very small tumors (< 1 cm), 97% (1-3 cm), 88% (3-5 cm) and 67% (> 5 cm). In the subgroup of patients restricted to T1-2-tumors (n = 106), 57 patients (53%) showed true negative, 4 (4%) false negative SNB. 38 (36%) revealed tumor cells in the HE-staining and an additional 7 patients (7%) solely in the immunohistochemical staining. The presented results show, that SNB is a reliable method for the evaluation of the nodal status in early breast cancer patients with a tumor size up to ca. 3 cm. While in about 50% of these patients a surgical intervention could be avoided after a negative SNB, an additional 5-10% of conventionally nodal negative patients can be found by the immunohistochemical examination of the sentinel node. The sn-concept can also identify parasternal metastasis and can be applied in patients after neoadjuvant therapy and patients with recurrent tumor. Indications and contraindications of this method, however, still remain to be determined. PMID- 11098577 TI - [Surgical axilla dissection. Technical standard or obsolete method?]. AB - Axillary lymph node status remains the single most important prognostic parameter and has crucial therapeutic implications in patients with breast carcinoma. Surgical dissection of the axilla is commonly regarded as the standard procedure of axillary staging, its sensitivity and specificity being 99% and 100%, respectively. Apart from giving reliable information on the individual prognosis axillary dissection also contributes to efficient local tumor control in the axilla, as it reduces the risk of local recurrence to less than 1.4% if more than 10 lymph nodes are removed. Alternative, less or non-invasive axillary staging methods have either not yet been sufficiently standardized (immunoscintigraphy, PET-scan, prediction of axillary lymph node status by means of individual risk factors) or are associated with a considerable risk of false-negative staging (up to 50% of patients with positive axillary lymph nodes are not detected by palpation alone, ultrasonography or CT-scan). The basic principles of axillary sampling and axilloscopic dissection are questionable because the number of lymph nodes removed during these procedures is commonly less than 10. With its sensitivity/specificity being comparable to that of standard axillary dissection sentinel lymph node biopsy represents a highly promising approach which will in the future potentially lead to significant optimization of the clinical management of patients with breast cancer, especially those diagnosed in early stages (T1 a, T1 b and T1 c). PMID- 11098578 TI - [The significance of lymph node status in papillary and follicular thyroid gland carcinoma for the nuclear medicine physician]. AB - The impact of lymph node metastases on prognosis of differentiated thyroid cancer is discussed controversially. Therefore the data of 596 patients with papillary or follicular thyroid cancer are analysed retrospectively, which have been treated between 1980 and 1995 at the Clinic and Policlinic for Nuclear Medicine of the University of Wurzburg. The influence of lymph node metastases on prognosis with respect to survival is analysed with the univariate Kaplan-Meier method and with the multivariate discriminant analysis. In addition, the influence of the prognostic factor "lymph node involvement" on distant metastases is analysed by a stratified comparison and an univariate test. In papillary thyroid cancer, the 15 year-survival-rate for stage pN1 is significantly lower (p < 0.001) with 88.7% as compared to stage pN0 (99.4%). In patients with follicular thyroid cancer this difference is even more pronounced (64.7% versus 97.2%, p < 0.001). However, the multivariate discriminant analysis shows that the only prognostic factors are tumour stage and distant metastases, and--in papillary thyroid cancer--patient's age. So lymph node metastases are not an independent prognostic factor concerning survival. However, lymph node metastases have a prognostic unfavourable influence with respect to distant metastases especially in papillary thyroid cancer stage pT4 (distant metastases in patients with negative lymph nodes 0% and in patients with positive lymph nodes 35.3% [p < 0.001]). PMID- 11098579 TI - [Pro and contra lymphadenectomy in papillary and follicular thyroid gland carcinoma]. AB - Patients with papillary and follicular thyroid carcinomas are said to have an excellent long-term prognosis. However, over 90% of papillary carcinomas have synchrone lymph node metastases. Based on clinical, histopathologic and molecular biological factors, different patient groups can be defined with an increased recurrence rate and disease related mortality-rate of up to 50% in the long-term follow-up of 30 years. Therefore, not only the extent of the surgical resection of the thyroid cancer but also the lymph node dissection as an oncologic correct surgical procedure is of great importance. Although the position of surgical lymphadenectomy is still discussed controversially there are increasing reasons to support the concept of a radical initial operation following the rules of oncologic surgery. We consider total thyroidectomy and modified neck-dissection as the standard operation in well differentiated thyroid carcinoma. In unilateral carcinoma both the central and the ipsilateral cervico-lateral lymph node compartments are dissected. In multicentric bilateral carcinomas a bilateral cervico-central and cervico-lateral lymphadenectomy has to be performed. PMID- 11098580 TI - [The value of blood transfusion in tumor surgery. Experiences from randomized trials]. AB - Since the first publication by Tartter in 1982 [3] multiple studies were performed concerning the association between blood transfusion and outcome in surgical oncology. The hypothesis was that blood transfusion-associated immunosuppression influences postoperative infection rates and long-term prognosis. Due to the fact that most of these studies were uncontrolled observational the decisive question about the causal relationship could not be answered so long. In randomized studies different methodological approaches were performed. Jeekel from Rotterdam and our group tried to control the allogeneic transfusion-associated immunosuppression by use of autologous transfusions. We found in our monocentric study with a very homogenous study population a significantly reduced infection rate in the autologous blood group which was not observed in the multicentric Dutch trial. The second approach was chosen by Jensen from Denmark and van de Velde from the Netherlands. They used leucocyte depletion to create a less immunosuppressive blood product, which was then randomized and compared with conventional allogeneic transfusions. Whereas the monocentric Dutch study showed a significant effect in the pilot study and also in the later larger confirmational study, again this was not found in the multicentric Dutch trial. Concerning the association between prognosis and transfusion the results of two randomized studies are available. The multicentric study from Jeekel from Rotterdam and our monocentric study found also in this endpoint different results. Whereas in our study allogeneic transfusions were an independent risk factor demonstrated in the intention-to-treat comparison by a clear trend (p = 0.067), this could not be detected in the multicentric Dutch study. In conclusion, the results from actually available randomized studies are not compulsive. Whereas the homogeneous monocentric studies showed a significant transfusion effect for the postoperative infection rate and also for long-term prognosis, this could not be verified in two multicentric trials. Also the cumulative meta-analyses are not able to answer the decisive question concerning the causal relationship between transfusion and long-term prognosis. Therefore new and innovative study designs are mandatory. PMID- 11098581 TI - [Transfusion use in curatively operated patients with colorectal carcinomas. Swiss Study Group for Clinical Cancer Research]. AB - OBJECTIVE: Is there an improvement of the behaviour for restrective blood transfusions after the data in the literature and especially the preliminary data of the SAKK 40/81 study have been published? They have shown a worsening of the prognosis in patients with colorectal cancer after pre-/postoperative blood transfusions have been given. MATERIAL AND METHODS: Analysis of the retrospective transfusion data of the SAKK 40/81 study in comparison with the prospective transfusion data of the study SAKK 40/87. RESULTS: The analysis of the data showed that in the SAKK 40/81 study more patients received blood transfusions than in the SAKK 40/87 study (77% versus 49%). Especially there was a diminution from 90% in the SAKK 40/81 to 59% in the SAKK 40/87 study for the rectal cancer patients respectively from 70% to 44% in the colon cancer patients having received blood transfusions. The mean value of hemoglobin of the patients not having received transfusions has decreased from 11.2 (7.8-15) g/100 ml in the SAKK 40/81 to 10.6 (5.4-15) g/100 ml in the SAKK 40/87 study (p = 0.0001). CONCLUSION: The data of the two SAKK studies showed that in Switzerland the donation of blood transfusions in patients with colorectal cancer has continuously been handled more and more restrictive. An even more restrective use may be possible in future due to new operation techniques and the possibility of preoperative administration of erythropoetin to increase the hemoglobin level. PMID- 11098582 TI - [Role of lymph nodes in rectal carcinoma]. AB - The significance of the lymph nodes for the therapy and the prognostics of the rectal cancer are reviewed. Not only the depth of penetrations, but the probability of lymph node involvement (in our series tumors staged T1 had in 12% positive lymph nodes, T2 in 20% respectively, T3 in 37% and T4 in 80%) determine the possible surgical treatment. The available data of different techniques in lymphadenectomy and lymph node staging are compared and discussed. The aim of this paper is to give the oncologically interested the opportunity to pursue the actual questions and to inform themselves about the current standards in regard to the lymphatics in rectal cancer. PMID- 11098583 TI - [The role of lymph nodes in colon carcinoma]. AB - Metastatic spread of colon cancer to intermediate lymph nodes is found, depending on tumor stage, in up to 44% of patients and to central lymph nodes in about 10%. The incidence of skip metastases is estimated not to exceed 5%. Tumorous involvement of pericolic lymph nodes occurs almost only within a 10 cm distance from the primary. There is only one prospective randomized controlled trial available comparing hemicolectomy versus radical segmental colectomy in patients with left colonic cancer. In this limited study there was no statistical difference regarding survival, mortality and morbidity between the two groups. However, several large retrospective studies are in favor of extended colon resection with radical clearance of lymphatic tissue. In 198 patients with colon cancer excluding rectal cancer in a 6-year period we performed 151 radical (76%) and 47 segmental (24%) colonic resections. The median length of the specimens in an unstretched and formalin-fixed state was 39 cm and 19 cm, respectively. The mean number of investigated lymph nodes was 16 and 12, respectively. 40% of our patients showed lymph node metastases. In-hospital mortality was 4/198 patients (2%) and surgical morbidity occurred in 29/198 patients (15%). We recommend radical colonic resections in all potentially curable patients with colonic cancer. PMID- 11098584 TI - [Prostate cancer: at the beginning of a new millennium]. PMID- 11098585 TI - [Bladder cancers]. PMID- 11098586 TI - [EAU highlights in pediatrics]. PMID- 11098587 TI - [Incontinence and urodynamics]. PMID- 11098588 TI - [Erectile dysfunction: an update on new therapeutic options]. PMID- 11098589 TI - [In Process Citation] PMID- 11098590 TI - [In Process Citation] PMID- 11098591 TI - [In Process Citation] PMID- 11098592 TI - [Aging of the working population in the European Union]. AB - The working population over 50 years of age will grow considerably during the next 15 years. After 2010, the number of retired people over 65 years of age will be almost double that of 1995, with a strong impact also on working conditions and the labour market. Work ability is a dynamic process that changes, through its components, throughout life and is the result of the interaction between individual resources (including health, functional capacity, education and skills), working conditions, and the surrounding society. Work ability creates the basis for the employability of an individual, which can be supported by a number of actions (e.g. legislation on work and retirement) and social attitudes (e.g. age discrimination). Consequently, the prevalence of limitations in work ability varies significantly according to how it is evaluated and the frequency of work disability can vary considerably in different times, locations and populations. The Work Ability Index, created and used in a Finnish 11-year longitudinal study, has been proved a useful practical tool for the assessment of workers' fitness and a good predictor of work disability. Measures able to restore, maintain or promote work ability depend on the current work status and the needs of the target groups, and must concentrate on work content, physical work environment and the work community. The actions targeted towards the individual, on the other hand, concentrate on strengthening the health status and functional resources of the workers and developing professional expertise and skills. Correctly targeted and integrated measures improve work ability of ageing workers and therefore lead to improved work quality, increased productivity and also improved quality of life and well-being. They also have positive long-term effects on the "third age", when the worker retires. PMID- 11098593 TI - [Biology and needs of the aged]. AB - The elderly age group is growing rapidly as a proportion of the total population of Italy. By the year 2020 the population aged 65 and over is expected to increase from 17% to 23%. The biological process of aging leads to a loss of functional reserves, and so elderly people are frail and affected by multiple diseases. However, health in the elderly is synonymous with self-sufficiency, which may be preserved even in the presence of a disabling disease. According to the new WHO International Classification of Functioning and Disability (ICIDH-2) the important elements in defining health status include personal activity and social participation, as well as body functions. As people age in different ways, the response to the needs of the elderly must be flexible and multidimensional. One of the priority objectives of modern medicine is to push disability to the end of life as much as possible. PMID- 11098594 TI - [Work capacity and aging]. AB - Maintaining a good work ability depends on satisfactory health and employment status, which is supported by suitable working conditions and correct life styles. From the biological perspective, ageing means a foreseeable progressive and overall deterioration of the various physiological systems, but not of such a kind and severity to consider most people over 50 as too old or unfit for work, as has been shown by several studies that assessed work ability not only in terms of biological age, but of functional age and actual work output. From the physio pathological perspective, we can observe either illnesses associated with the passage of time or age-related changes that might precipitate diseases, as well as environmental changes that modulate ageing and developmental changes that accelerate or retard ageing. From the practical point of view, it should taken into account that job demands often do not follow the natural biological and functional changes of the individual, consequently the relative work load can be higher in older workers. On the other hand, ageing also means a professional growth in terms of strategic ability, shrewdness, wisdom and experience. The high interindividual variability of physical, mental and social conditions that is observed with the increase in age makes it necessary to adopt flexible and personally tailored measures, as shown by recent surveys in some European countries aimed at reducing age discrimination and work disability, and at promoting work ability by means of actions directed towards both improvement of work organisation and support of psycho-physical conditions of older workers. PMID- 11098595 TI - [Deficiencies and resources of working population in relation to age: a multidisciplinary approach]. AB - The aging of the population as a whole and the later age at which young people start work are increasing the percentage of older employees. In situations where the working conditions are highly demanding, as in shiftwork, time-pressure jobs, and adaptation to modern technology or skill diversification, this demographic trend may cause serious problems. The way in which job constraints and demands are withstood at various ages should be considered in relation to health, which is often, whether implicitly or explicitly, a selection criterion in the work place. The connection between work and health can rarely be described by a single causal relationship and requires specific epidemiological methods. Moreover, a health problem linked to age can have a feedback effect on the manner in which a job is performed. While these problems do indeed arise in the areas of work and health, they are nonetheless usually symptoms of modifications that have taken place in the work activity itself. The ergonomic approach nevertheless allows us to improve our understanding of changes in work behavior as age increases, as experience is gained, and as skills are acquired. Men and women on the job are not passive spectators of the good or poor fit between the characteristics of their jobs and their own functional state. Consciously or unconsciously, they modify their operating modes (movements, work pace, posture, etc.), reduce their effort level in some subtasks, make more plans to avoid emergency situations, check the outcome of their actions so as to reduce errors that would be costly to correct, and adjust the distribution of tasks in cooperative and collective work situations. But these strategies can only be implemented if the work conditions and organization foster and promote them. PMID- 11098596 TI - [Psychological and psychosocial aspects in the elderly workers]. AB - The international literature is producing information about job performance of elderly workers that is in contrast with current prejudice. Objectively, the elderly worker exhibits changes in performance but these changes do not necessarily interfere in a negative way. On the contrary, they sometimes have an extremely positive value: it is up to social partners and entrepreneurs to attribute a role and a value to the elderly workers. The psychological data on populations exposed to different occupational risks carried out by the Department of Occupational Health of the University of Milan are revised according to age and seniority. Some suggestions are given to encourage elderly workers to stay at work. PMID- 11098597 TI - [Occupational aging and cardiovascular diseases]. AB - A short review of occupational aging and cardiovascular disease is presented. Coronary and stroke events are typical diseases of aging, in which rare monogenic disorders and common polymorphisms interact with environmental factors. Among these factors, particular attention has been given to social and occupational organizations. Cardiovascular mortality apparently increases with heavy industrialization processes and lower socioeconomic and educational status. Similar data are now available for incidence and case fatality. Since traditional risk factors--e.g., cholesterol, blood pressure and smoking--follow the same pattern, but with a weaker association than expected, observational and preventive studies are considering risk possibilities beyond such factors. Job demand/job decision latitude is the most widely used paradigm to evaluate specific organizational risk at work. A few cohort or population studied and many cross-sectional studies have shown an independent and inverse relationship between decreasing control of the job, coronary disease and blood pressure levels. While such results need further confirmation, they suggest the need for more accurate research on the adverse or mutually protective influences of long lasting adaptation processes required by prevailing work organization. PMID- 11098598 TI - [Aging at work and musculoskeletal disorders]. AB - By means of a critical review of the international literature and of their own published experiences, the Authors discuss the influence of the "age" factor on work related musculoskeletal disorders of the spine and upper limbs. Regarding the spine, the lumbosacral spine in particular, there is evidence (both in relation to pathways and from epidemiological data) of the influence of age in determining a progressive increase in the occurrence of spondyloarthropathy with clear radiological signs. For upper limb disorders the influence of the "age" factor is still under debate and in any case does not seem of great importance. As far prevention is concerned for elderly workers subject to fixed postures and repetitive movements of the upper limbs it seems sufficient, to adopt the general measures used for the whole working population. However, specific measures should be adopted for elderly workers exposed to manual material handling (MMH). These consist in using reference values for the recommended weight that are lower than those adopted for younger workers (aged 18-45 years) and in implementing specific programs of active health surveillance. PMID- 11098599 TI - [Fatiguing work, aging and health: a cross-sectional study of a group of anesthesiologists]. AB - A cross-sectional study was carried out to verify if work as an anaesthetician reanimator can be considered as a fatiguing job. An anonymous self-administered questionnaire was utilized to obtain information about the general characteristics of the subjects, job organization, human relations, perceived risks and previous diseases; 1438 questionnaires were examined. The results of the study showed that in anaesthetist-reanimator: organic diseases do not occur earlier than expected; perception of the emotional aspects related to the particular occupation is low; the group relationships are difficult, on both horizontal and vertical levels, and these difficulties increase work-stress and hinder its management; age influences the occurrence of both arthritic and stress related diseases; there is an association between jobs dissatisfaction and stress disorders; all the causal variables considered, evaluated as a group, constitute a heavy physical load, that leads to risk of stress related diseases. PMID- 11098600 TI - [ESTEV study on relationship between health, work and aging in Italy]. AB - A longitudinal epidemiological study into the relationships between age/health/work is currently under way in different geographical areas throughout Italy. The research is co-ordinated by INRCA (Italian National Research Centres on Aging) in Ancona with the collaboration of the Universities of Ancona, Verona and Bari. This study concerns a population of approximately 2,000 employees from a variety of production sectors. The sample is made up of 5 groups of workers selected according to the year they were born and aged: 32, 37, 42, 47 and 52 years. The chosen research tool is modelled on ESTEV and VISAT researches, the former conducted on a sample of 20,000 French workers, the latter on approximately 3,000 workers and still under way. It involves a set of three questionnaires which allow for a number of variables to be taken into consideration: the first questionnaire concentrates on information regarding the occupation, both past and present; the second on the life style and self-assessed health according to the Nottingham Health Profile (NHP); the third, completed by the occupational physician, contains information on current and previous illnesses, the presence of disorders of the musculo-skeletal apparatus, the taking of any drugs and some anthropometrical and clinical-instrumental parameters (Respiratory Functionality Test, Visiotest and Audiogram). The study with be carried out in two phases: a first survey (under way) and a second one five years later on the same subjects. The results of the analysis will be compared with those of other European countries. PMID- 11098601 TI - [Physical exercise in the prevention of musculoskeletal diseases in the elderly worker]. AB - This paper reviews the impact of ageing on physical work capacity in the elderly worker and the benefits of exercise programmes designed for the prevention of musculo-skeletal complaints. Physical work capacity generally declines significantly after 50 years of age as a consequence of a reduction in aerobic capacity and muscle strength. The latter is accompanied by a decline in the fatigue threshold. This, in combination with a reduced recovery capacity after exercise, may lead to chronic overload of muscles and tendons in the elderly worker. Thus, it has been proposed that physical exercise programmes may prevent the onset of musculo-skeletal disorders. The effectiveness of different physical exercise programmes proposed in the literature for the prevention of musculo skeletal disorders is reviewed here. The adoption of an active life-style and the rational use of ergonomic interventions, such as automatization of certain work processes, also appear to be crucial in minimizing the impact of ageing on work ability and capacity in the elderly worker. PMID- 11098602 TI - [Aging and work: health aspects in cleaners]. AB - Over the last few years, studies on the relationship between ageing and work have attracted growing interest due to the increased probability among workers of developing major health problems as a consequence of ageing workers. Negative outcomes for health are possible when an age-related imbalance appears between physical workload and physical work capacity. Cleaning could be considered as a paradigm for studying the relationship between ageing and physically demanding jobs. Cleaning workers show a high proportion of ageing women who are at the bottom level of the social-status class and are generally poorly educated with low income and social support level. This study, particularly aimed at highlighting the presence of musculo-skeletal disorders, was conducted by means of standardized questionnaires and protocols for collecting anamnestic and clinical data. The results show an increased prevalence of disorders of elbow, wrist/hand and cervical spine which could be caused by work organization and non ergonomic tools. On the basis of the experience gained it was possible to propose solutions for the ageing working population concerning the workload, health surveillance methods and the ergonomic measures as regards organization, work place and tools. PMID- 11098604 TI - [Violence against physicians. every 6th colleague is assaulted]. PMID- 11098603 TI - [Aging and occupational accidents]. AB - Ageing and occupational accidents is an underestimated issue in Italy. This deals both with shortcomings of data and proper studies in the matter and with safety culture and interventions. Nevertheless, some tables are presented showing a relation between ageing and occupational accidents both in frequency and in severity. These results are discussed in the light of a review of the literature on age-related accidents risks of three decades published in 1995. A framework is reported outlined for the purpose of identifying jobs in which ageing has either a preventive or an aggravating effect on accident occurrence. It is concluded that the safety problems of older workers may well be restricted to activities that are specifically "age-impaired". Age-related accident problems can also be specific in terms of injury type, as also demonstrated by some Italian data. Finally, it is emphasized that further research would benefit greatly from longitudinal designs, proper exposure measurements, intra-occupational investigations, consideration of the positive effect of relevant experience on occupational safety and greater precision about the type of accidents in focus. On the prevention side, a better attention to a global approach to safety problems (not only to technological aspects but also to immaterial ones, like work organization, psychosocial factors and so on) is pointed out. PMID- 11098605 TI - [The patient becomes dangerous. Controlling the situation]. PMID- 11098607 TI - [A new effective principle in stress headache. Bacterial toxin for the head?. Interview by Dr. Beate Schumacher]. PMID- 11098606 TI - [Reflux after Helicobacter pylori eradication. What is the ambivalence about?]. PMID- 11098608 TI - [News in the anorectal area. Heart salve and stapler against anal diseases]. PMID- 11098609 TI - [Incidental findings in gastroenterology. Preventing overdiagnosis]. AB - The use of diagnostic imaging procedures and laboratory investigations in gastroenterology can sometimes lead to findings of questionable value. This highlights the problem--also from the point of view of costs--of deciding whether further diagnostic investigation is necessary or not. No requirement for action is given, for example, in the case of shallow diverticula in the esophagus, hiatal hernia in the absence of reflux symptoms, in the case of small cysts or hemangiomas in the liver, polyps on the wall of the gallbladder, hyperplasia of Brunner's glands in the duodenum or fungi in the stools of immunocompetent persons. Of a controversial nature are the recommendations with regard to Helicobacter pylori gastritis, and non-symptomatic gallstones. Further diagnostic clarification is certainly required when such findings as fatty liver, diverticula in the colon or pancreas divisum are encountered. PMID- 11098610 TI - [Incidental findings in cardiology. Diagnosing today what can cause illness tomorrow]. AB - Among the anatomical and functional findings in cardiology, congenitally corrected transposition of the major vessels (ventricular inversion), bicuspid aortic valve and prolapse of the mitral valve with simultaneous mitral insufficiency have at least a potential for causing future problems. In such cases, regular cardiological checks or a further diagnostic work-up is to be recommended. The assessment of cardiac sounds can usually be correctly interpreted on the basis of the history, physical examination and auscultation. Among the electrocardiological findings, complete left bundle-branch block and prolongation of the QT segment, mandate clarification of a structural heart condition. Furthermore, in the event of ventricular extrasystoles, right ventricular cardiomyopathy needs to be excluded. Such isolated conduction disorders as left-anterior fascicular block or right bundle-branch block are of no prognostic significance. PMID- 11098611 TI - [Incidental findings in endocrinology. When to observe and when intervention is necessary]. AB - Hirsutism, gynecomastia or delayed puberty are usually benign conditions. However, underlying serious disorders must be excluded. Abnormalities in thyroid hormone measurements with contraceptives, glucocorticoids or severe nonthyroid disease must be differentiated from "true" impairments of thyroid function that require specific treatment. Increasingly, abnormalities are detected incidentally (incidentaloma) by diagnostic imaging procedures in organs such as the pituitary or the adrenal gland. Apart from judgements based on the original size of the tumor or the progression in size during observation, functional evaluation is always necessary to decide whether surgery should be recommended. PMID- 11098612 TI - [Puzzling urticaria. Allergies, pseudo-allergy, bacteria, fungi, parasites?]. PMID- 11098613 TI - [The "diabetes health record". Patient education series, 1]. PMID- 11098614 TI - [Intensified therapy with insulin analogs in type 2 diabetes: normalizing metabolism and maintaining body weight?]. PMID- 11098615 TI - [Diagnostic quiz. Lung shadow of uncertain origin. Silicosis]. PMID- 11098616 TI - [Chronic hepatitis. New therapeutic approaches on trial]. PMID- 11098617 TI - [Mycologic commonalities in veterinary and human medicine]. PMID- 11098618 TI - [Epidemiology, clinic and treatment of dermatomycoses caused by zoophilic dermatophytes] . AB - In the last 50 years the spectrum of agents of dermatophytoses changed. The change is remarkable especially in the case of zoophilic dermatophytes. Microsporum canis has displaced Trichophyton verrucosum. In the period from 1966 to 1970 2822 dermatophytoses were registered as occupational dermatoses in the former German Democratic Republic. In the following years the number of these diseases was continuously reduced, between 1981 and 1985 down to 995 cases. This reduction is predominantly the result of active immunoprophylaxis by means of inactivated, hydrolized vaccines, mainly by the Soviet live vaccine LTF-130. The scientific basis of this vaccination was established by Kielstein and co-workers. The treatment of Microsporum canis infection of the scalp requires the administration of systemically acting antimycotics over a long period (6-8 weeks). PMID- 11098619 TI - [Incidence and control of cattle ringworm in Scandinavia]. AB - Clinical aspects, prophylactic as well as therapeutical vaccination with a live attenuated vaccine against Trichophyton verrucosum are dealt with in this article. Further, the epidemical and zoonotical view of the disease is described. The farmers' motives to vaccinate differ depending on type of herd. Hygienic and disinfectant measures in connection with the start of a vaccination program are emphasized. In Sweden, vaccination against ringworm is included in the "Faultless Hide" scheme as one amongst other preventative measures aiming to improve the hide quality. A beneficial effect on both frequency of diseased herds and quality of the hides is obtained. PMID- 11098620 TI - [Molecular biological methods and their consequences in taxonomy and diagnosis of dermatophytes]. AB - Trichophyton rubrum, T. mentagrophytes, T. verrucosum, Microsporum canis, M. gypseum and Epidermophyton flocoosum represent the cause of human dermatomycoses isolated most often at the University Hospital of Dermatology in Graz, Austria, between 1991 and 1998. So far, identification was mainly based on the cultivation of fungal isolates on special media as well as on the analysis of their microscopic characters. Restriction enzyme length polymorphisms (RFLPs) were now used to identify these human fungal parasites. For this purpose, internal transcribed spacers (ITS) which are located between ribosomal RNA genes have been amplified by using PCR and have afterwards been used to generate species specific RFLP patterns. By this method, a fast and reliable identification of these species was made possible. Nucleotide sequence data of this region not needed for identification have been worked out to show RFLPs in more detail. PMID- 11098621 TI - [Differentiation of yeasts in mastitis milk] . AB - Differentiating 174 yeasts from clinical samples to get a survey of those species found in mastitis milk and milking machines we identified five genera with 29 species. Species of the genus Candida dominated with 77% of all samples. More than 25% of yeasts from milk were identified as C. rugosa and more than 20% as C. catenulata. The six species found most often represented 74% of all yeasts in this group. Yeasts from milking machines were more heterogeneous. PMID- 11098622 TI - [Mycozoonoses with special regard to ringworm of cattle]. AB - Ringworm of cattle is nearly exclusively caused by Trichophyton verrucosum. This skin disease is worldwide present in cattle and is responsible for high economic losses in cattle farming. T. verrucosum may also be responsible for severe skin diseases in man. For economic and epidemiological causes a control of ringworm in cattle is still necessary. Preventive measures consist in hygiene and vaccination of cattle with live vaccines against ringworm in cattle. PMID- 11098623 TI - [Evidence of Cryptococcus neoformans in domestic and sports pigeons in Thyringia]. AB - 19 strains of Cryptococcus neoformans var. neoformans were isolated from 17 (= 40.5%) of 42 investigated pigeon breeder flocks in Thuringia. PMID- 11098624 TI - [Asteroid bodies: host-pathogen reactions in mycoses]. AB - A review of the literature on asteroid bodies (Splendore-Hoeppli phenomenon) as well as an immunoelectronmicroscopic study on asteroid bodies with Candida albicans and electronmicroscopic observations on asteroid bodies with Aspergillus are presented. The following definitions are proposed: Asteroid bodies with Candida albicans: precipitate consisting of fungal antigen and antibodies of host origin in light microscopically visible dimension deposited on the cell wall surface of Candida cells in parasitic condition. Asteroid bodies with Aspergillus: precipitate consisting of fungal antigen, antibodies of host origin and necrotic cellular detritus originated from cells of host defense, in light microscopically visible dimension, deposited on the cell wall surface of abortive Aspergillus cells, reduced in longitudinal growth, in parasitic condition. PMID- 11098625 TI - [Host-pathogen relations in yeasts]. AB - Allogenic and autogenic factors are decisive for a colonization of warmblooded hosts by yeasts where they can live inside as symbiont, commensal or pathogen. To develop pathogenicity a capacity of yeasts for multiplication inside of humans or homoiothermic animals and resistance to environmental influences is not sufficient. Pathogenicity rather depends on the ability of structural morphological changes, phenomena of adherence and of microbial competition as well as the presence of genes for down-regulation of the host defense and for enzyme production enabling a parasitic life style. Among the yeasts Cryptococcus neoformans and some Candida species meet these requirements. PMID- 11098626 TI - [Molecular biological differentiation of yeasts]. AB - Regarding the rise and the epidemiological shifts in Candida infections, accurate and rapid identification of yeasts to species level is exceedingly important. Furthermore, non-albicans species (e.g. C. glabrata, C. dubliniensis, C. inconspicua, C. krusei and C. norvegensis) have recently emerged as azole resistant pathogens. Standard methods for cultivation and differentiation of Candida species are time-consuming, often insensitive and can fail to distinguish non-albicans species. To overcome low sensitivity, poor specificity and untolerable delay, molecular tools to the diagnosis of Candida infection have been adopted, particularly the polymerase chain reaction (PCR). Several genus and species specific amplification-based diagnostic methods for detection of medically important yeasts have been published. As the incidence of nosocomial Candida infections has been risen significantly, typing has become increasingly important in candidal epidemiology to define infectious processes as well as sources and modes of transmission. For the development of rational infection control measures, modern genotyping methods based on genomic DNA polymorphism (e.g. pulsed-field gel electrophoresis, DNA amplification-based methods) have replaced conventional physiological techniques. However, reliability, sensitivity and efficiency of genotyping methods have been controversially discussed and these DNA-based methods are mostly insufficiently standardized. Criteria as typeability, reproducibility and discriminatory power should be considered in evaluating typing systems. PMID- 11098627 TI - [Cryptococcus species--etiological agents of zoonoses or sapronosis?]. AB - Cryptococcus strains are doubtless the cause of a sapronosis which occurs worldwide as cryptococcosis in humans and warm-blooded animals. The etiological agent is Cryptococcus neoformans with the varieties neoformans and gattii. The infection proceeds from environmental sources and not from animals suffering from cryptococcosis. Therefore the designation as zooanthroponosis is not right. There is a correlation between the geographical distribution of the varieties of Cryptococcus neoformans in the environments and the clinical manifestation of cryptococcosis. The reservoir of var. neoformans, serotypes A and D, are worldwide pigeons and pet birds which are frequent healthy carriers. They excrete these yeasts with their excrements. Bird droppings favour the propagation of cryptococci in natural substrates. The cryptococcal infection arises from inhalation of contaminated dust. Variety gattii, serotype B, occurs mainly in tropical and subtropical climates and its environmental niche has been identified in Eucalyptus trees. Var. gattii was detected in the air and in decaying wood debris under the canopies of these trees. Serotype C was isolated once from almond trees in Colombia. Cryptococcoses in men caused by var. gattii are endemic in Australia, California, Brazil and other tropical and subtropical regions. Modern molecular typing procedures are available for studying ecological problems of cryptococci and epidemiological questions of cryptococcosis. Immunosuppressed individuals, especially HIV infected persons and AIDS patients, should avoid contact with pigeons and pet birds. PMID- 11098628 TI - [Fourier transform infrared spectroscopy, molecular biologic methods and antimyocotic susceptibility patterns for identification and differentiation of cryptococcus species]. AB - Molecular biological methods as well as the FTIR method allows the rapid, reliable and reproducible determination and identification of Cryptococcus species from human, veterinary and environmental origin and their serovars. The results obtained by FTIR could be verified by the molecular methods. In addition, with the PCR and FTIR fingerprinting methods it is possible to distinctly group the serovars and differentiate the different Cryptococcus strains. PMID- 11098629 TI - [Yeasts of the genus Malassezia: taxonomic classification and significance in (veterinary and) clinical medicine]. AB - The historical development of the taxonomic classification of Malassezia yeasts until today yielded the description of seven different species based upon molecularbiological, morphological and biochemical parameters (M. furfur, M. pachydermatis, M. sympodialis, M. globosa, M. obtusa, M. restricta, M. slooffiae). Tests like Cremophor EL assimilation, esculin degradation, catalase test, pigment production and determination of polidocanol sensitivity appear to be suitable procedures for routine identification of the different species. Their colonization of clinically healthy humans (in particular M. sympodialis, M. globosa, M. obtusa) and animals (M. pachydermatis) skin renders interpretation difficulties, when isolating Malassezia spp. from clinical specimens. Out of the seven species, in man the clinical significance of M. furfur for pityriasis versicolor and systemic infections appears to be accepted largely. In dogs and cats M. pachydermatis has been regarded as cause of otitis externa and seborrhoeic dermatitis. But, due to geno- and phenotypic variabilities described for M. pachydermatis field isolates further investigations concerning the taxonomic uniformity of the species are necessary. PMID- 11098630 TI - [Mycologic investigation of foodstuff]. AB - This short communication presents a review on the work of the mycological laboratory of the Institute of Microbiology and Epidemics, Hannover, Germany, concerning the mycological investigation on feeding stuff. Out of the routine diagnostic work, examining more than 4000 probes in the last 10 years, some selected investigations on scientific questions will be mentioned. Different subjects as the influence of homogenisation of feeding stuff on the mycological evidence rate, the microbiological status of germinating feeding stuff for pet birds the contamination of wheat with Fusarium spp. after fungicidal treatment will be discussed. Out of the great number of diagnostic investigations the most often examined feeding stuffs, those for pigs, will be looked at more closely. The evidence rate of yeasts and moulds and the number of isolated fungi are demonstrated. PMID- 11098631 TI - [Mycoses in fish]. AB - Over 4 years 1.241 fish were examined mycologically. In 182 (14.7%) of them positive results were obtained. Most of the isolates belonged to the following genera: Cladosporium, Saprolegnia, Candida and Penicillium. Fungal infections were correlated with findings of Saprolegnia, Branchiomyces, Pythium, Ichtyophonus and sometimes Cladosporium. Most fungal isolates were etiologically irrelevant for diseases or death. PMID- 11098632 TI - [Relevance of mycotoxin contaminated feed for farm animals and carryover of mycotoxins to food of animal origin]. AB - Contaminated feed is the main source for mycotoxin infestation of farm animals. The oral intake of fungal metabolites with feed results in a negative impact on all relevant parameters of animal production. Moreover, under experimental conditions mycotoxins and/or their metabolites can be traced in meat, edible tissues, milk and eggs. However due to the high concentrations of toxins involved, such findings are rare in the daily practice. In Germany today only aflatoxins (aflatoxin M1 in milk) and ochratoxin A (in blood, meat and edible tissues from swine) are of practical relevance from the view of food hygiene and food safety. Other mycotoxins at present discussed like toxins of Fusaria (trichothecenes, zearaleone, fumonisins) and ergot alkaloids are of no importance as possible contaminants in food from animal origin although they could have a negative impact on animal production. PMID- 11098633 TI - Radiation-induced diffuse brain injury in the neonatal rat model--radiation induced apoptosis of oligodendrocytes. AB - The mechanism of radiation-induced diffuse brain injury was investigated using a model of delayed myelination in the irradiated neonatal rat brain in which the number of oligodendrocytes decreases without associated necrosis of the cerebral white matter. Immunohistochemical analysis using antibody against the large myelin-associated glycoprotein, a specific marker of oligodendrocytes at an early stage of development, showed that the number of the oligodendrocytes associated with myelination decreased in the irradiated hemisphere 1 day after irradiation and remained low until 5 days after irradiation. In situ terminal deoxynucleotidyl transferase-mediated nick end-labeling assay revealed that apoptosis mainly occurred in the cerebral white matter of the irradiated hemisphere. Three hours after irradiation, apoptotic cells were found in the subcortical white matter and the periventricular white matter. Six hours after irradiation, apoptotic cells were found in the internal capsule, and the numbers of apoptotic cells in the periventricular white matter and subcortical white matter increased. One day after irradiation, the number of apoptotic cells in the periventricular white matter decreased. Three days after irradiation, apoptotic cells were not observed in the cerebral white matter. These results suggest that the oligodendrocytes associated with myelination may be damaged via radiation induced apoptosis, and depletion of the oligodendrocytes may cause delay of myelination. PMID- 11098634 TI - Can continuous intraoperative facial electromyography predict facial nerve function following cerebellopontine angle surgery? AB - Intraoperative cranial nerve monitoring has significantly improved the preservation of facial nerve function following surgery in the cerebellopontine angle (CPA). Facial electromyography (EMG) was performed in 60 patients during CPA surgery. Pairs of needle electrodes were placed subdermally in the orbicularis oris and orbicularis oculi muscles. The duration of facial EMG activity was noted. Facial EMG potentials occurring in response to mechanical or metabolic irritation of the corresponding nerve were made audible by a loudspeaker. Immediate (4-7 days after tumor excision) and late (6 months after surgery) facial nerve function was assessed on a modified House-Brackmann scale. Late facial nerve function was good (House-Brackmann 1-2) in 29 of 60 patients, fair (House-Brackmann 3-4) in 14, and poor (House-Brackmann 5-6) in 17. Postmanipulation facial EMG activity exceeding 5 minutes in 15 patients was associated with poor late function in five, fair function in six, and good function in four cases. Postmanipulation facial EMG activity of 2-5 minutes in 30 patients was associated with good late facial nerve function in 20, fair in eight, and poor in two. The loss of facial EMG activity observed in 10 patients was always followed by poor function. Facial nerve function was preserved postoperatively in all five patients in whom facial EMG activity lasted less than 2 minutes. Facial EMG is a sensitive method for identifying the facial nerve during surgery in the CPA. EMG bursts are a very reliable indicator of intraoperative facial nerve manipulation, but the duration of these bursts do not necessarily correlate with short- or long-term facial nerve function despite the fact that burst duration reflects the severity of mechanical aggression to the facial nerve. PMID- 11098635 TI - Recovery from Duret hemorrhage: a rare complication after craniotomy--case report. AB - A 44-year-old female presented with Duret hemorrhage due to transtentorial herniation by extradural hematoma as a complication after craniotomy for treatment of spontaneous middle cranial fossa cerebrospinal fluid leakage through the oval window. Brain computed tomography revealed linear hemorrhage in the midbrain and the rostral pons. She awoke after 2 weeks in a coma, despite showing ocular bobbing and bilateral intranuclear ophthalmoplegia. She was discharged from the hospital with minimal neurological defects. Duret hemorrhage is usually fatal, but this case shows that early surgical decompression is the most important factor to avoid the worst sequelae. PMID- 11098636 TI - Ruptured aneurysm at the origin of the median artery of the corpus callosum associated with accessory middle cerebral artery--case report. AB - A 62-year-old male presented with ruptured anterior communicating artery (ACoA) aneurysm manifesting as severe headache associated with the rare combination of median artery of the corpus callosum (MACC) and accessory middle cerebral artery (MCA). Computed tomography demonstrated diffuse subarachnoid hemorrhage. Left carotid angiography demonstrated an anomalous vessel originating from the ACoA complex and passing forward in the interhemispheric fissure between the two companion A2 segments. This vessel was identified as the MACC. Another anomalous vessel originated from the left A1-A2 segment and passed into the sylvian fissure. This vessel was identified as the accessory MCA. Left frontotemporal craniotomy was performed to clip the neck of the aneurysm. After identifying both A1 and A2 segments, accessory MCA, and the MACC, the aneurysm neck was occluded successfully. The ACoA complex is one of the most frequent sites of vascular anomalies. Preoperative and intraoperative care is required to identify the presence of anomalies of the ACoA complex prior to clip placement, to avoid accidental damage or clipping, which may result in severe neurological deficits. PMID- 11098637 TI - Endovascular treatment of basilar tip aneurysm associated with moyamoya disease- case report. AB - A 58-year-old female was admitted unconscious to a local hospital. Computed tomography demonstrated subarachnoid hemorrhage. Cerebral angiography revealed evidence of moyamoya disease and a saccular aneurysm at the tip of the basilar artery. The patient was transferred to our hospital for embolization of the basilar tip aneurysm. Endovascular embolization was performed using Guglielmi detachable coils (GDCs), and the aneurysm was completely occluded with preservation of the parent artery. No change in the patient's neurological status was seen during and after the procedure. Endovascular treatment using GDCs appears to be particularly suitable for ruptured cerebral saccular aneurysms in patients with moyamoya disease. PMID- 11098638 TI - Correlation of magnetic resonance imaging and histological findings in a large basilar tip aneurysm after coil embolization--case report. AB - A 73-year-old female with a large basilar tip aneurysm underwent endovascular coil embolization with interlocking detachable coils (IDCs). The patient subsequently died of pneumonia 25 days after the embolization, and the aneurysm specimen was obtained at autopsy. Histological examination showed that the intraaneurysmal structure consisted of three layers. The outer layer was mildly organized thrombus surrounding the coils, the middle layer was disorganized clot, and the inner layer consisted of fresh blood clot. Gradient-echo magnetic resonance (MR) imaging taken before death had demonstrated a central region of high intensity and a peripheral low intensity region corresponding to the inner and middle-outer layers of the aneurysm, respectively. Intraaneurysmal placement of IDCs leads to the formation of a clot surrounding the coils. However, clot formation may be inadequate where the packing of the IDCs is incomplete. Comparison of the MR imaging and histological findings in this case show that gradient-echo MR imaging can assess thrombus and residual blood flow within the aneurysm. PMID- 11098639 TI - Restoration of hearing with an auditory brainstem implant in a patient with neurofibromatosis type 2--case report. AB - A 25-year-old male with neurofibromatosis type 2 had hearing restored with an auditory brainstem implant (ABI) after removal of an acoustic schwannoma. The ABI allows the patient to discern many different environment sounds and is a significant adjunct to lip-reading, enabling conversation with people who have clear pronunciation without the necessity for writing. PMID- 11098640 TI - Huge facial schwannoma extending into the middle cranial fossa and cerebellopontine angle without facial nerve palsy--case report. AB - A 46-year-old male presented with a huge facial schwannoma extending into both the middle cranial fossa and the cerebellopontine angle but without manifesting facial nerve palsy. Neurological examination on admission revealed no deficits except for speech disturbance. Computed tomography showed a multicystic tumor extending into the middle cranial fossa and the cerebellopontine angle, with destruction of the petrous bone. The tumor was totally grossly removed. Histological examination identified schwannoma. Total facial nerve palsy appeared postoperatively, but hearing acuity was preserved at a useful level. Facial nerve palsy is one of the most typical symptoms in patients with facial schwannoma, but is not always manifested even if the tumor extends into both the middle cranial fossa and the cerebellopontine angle. PMID- 11098641 TI - Langerhans cell histiocytosis limited to the pituitary-hypothalamic axis--two case reports. AB - Langerhans cell histiocytosis rarely presents as a solitary lesion in the pituitary-hypothalamic region, and is indistinguishable from germinoma, which occurs much more frequently, especially in Japanese. A 14-year-old girl and a 9 year-old girl presented with polydipsia and polyuria as the initial symptoms. Magnetic resonance (MR) imaging demonstrated a round mass at the pituitary stalk appearing as isointense on T1-weighted imaging and hyperintense on T2-weighted imaging. Endocrinological examination revealed mild hypopituitarism with central diabetes insipidus. Both patients underwent open craniotomy. Histological examination revealed granulomatous tissue with eosinophil infiltration and frequent Langerhans histiocyte clustering, compatible with the diagnosis of Langerhans cell histiocytosis. Low-dose local irradiation of 20 Gy was administered. First patient was followed up for 8 years, and her hypopituitarism gradually improved to a minimal level with only amenorrhea as the residual symptom. Recent MR imaging showed no residual mass at the region. Second patient was followed up for 15 months, and her diabetes insipidus is stable. MR imaging performed 5 months after the treatment showed marked reduction of the mass. These cases reemphasize the importance of histological diagnosis for lesions with similar neuroimaging appearances. Biopsy and low-dose irradiation are an effective treatment for this rare and essentially benign lesion, as opposed to attempting total removal of the mass. PMID- 11098642 TI - Interhemispheric glioependymal cyst associated with agenesis of the corpus callosum--case report. AB - A male neonate was admitted because prenatal ultrasonography indicated central nervous system abnormalities. Neurological examination showed no abnormality except for electroencephalographic spike activities. Magnetic resonance imaging revealed a cystic lesion in the left interhemispheric fissure, agenesis of the corpus callosum, and microgyria in the left frontotemporal lobes. Cerebral blood flow (CBF) was diffusely reduced. The cyst wall was partially removed and a cyst peritoneal shunt procedure was performed. The histological diagnosis was glioependymal cyst. The spike activity disappeared and CBF dramatically improved after the operation. PMID- 11098643 TI - [Counseling of parents of infants and toddlers]. PMID- 11098644 TI - ["Reading-your-baby lessons" for parents of excessively crying infants--the concept of "guided parent-infant training sessions"]. AB - New findings indicate that persistent crying (infantile "colic") may be closely interlinked with regulatory difficulties in the organization of sleep and waking states and in sustaining prolonged episodes of uninterrupted sleep. It has been shown that preverbal communication plays a significant role in the regulation of infant states and in the prevention of infantile persistent crying. However, parents are usually unaware of these explanations, and tend to attribute the crying to inappropriate causes or have no understanding at all about why their child cries so much. Therefore, the baby's signals are not answered in an effective way. In this article "guided parent-infant-training sessions", a new treatment method for parents with excessively crying babies, will be introduced. It is the goal of this approach to help parents to improve the "reading" of their child's signals and to act according to the baby's needs. In order to provide an opportunity for training, parents are given their appointment at a time, when it is most likely for the child to cry. In the context of a therapeutic relationship they may be experiencing for the first time calming a baby they previously thought to be inconsolable. PMID- 11098645 TI - [Early appearance and intergenerational transmission of maternal traumatization within the context of mother-infant interaction]. AB - At the age of 9 weeks a girl was presented by her mother because of intolerable hysterical attacks' triggering maternal impulses of abuse. Maternal perception of her infant was distorted to the extent that the mother was reexperiencing encounters with her own intrusive and traumatizing mother in the face of her screaming child. She also perceived the infant's motor impulses as physical transgressions against herself and expressed intense anxieties about her daughter's future aggressive potential. The infant was viewed by her mother as extraordinarily and dangerously greedy. Even neutral infantile vocalizations were perceived as manipulating and sadistic. She tried to ward off these anxieties by employing a rigid scheme of rules and obsessively controlling the father's and grandmother's interaction with the child. The mother feared to be overwhelmed by the infant's needs if she'd yield to them in a flexible way. A background of early neglect and trauma experienced by the mother is illustrated considering recent literature about early intergenerational transmission of traumatic experiences in order to demonstrate treatment modalities in view of protective and risk factors. PMID- 11098646 TI - [Counseling of families with two- to three-year old children]. AB - In this paper, we present the counselling practice in parents with two- or three year-old children. In most of the cases, the presented children are suffering from separation anxiety or behavioral disturbances. It is either a problem of infants who cling to their mothers and who avoid unfamiliar situations or of children who come into conflicts with their parents because of their oppositional behavior. The basis for our diagnostic and therapeutic procedure is a model of the relationship disturbances in the parents-child-system which is concentrated on the dimensions attachment/intimacy and autonomy/differentiation. These two dimensions are to be understood as a pair of scales loaded by the parents with burdens of conscious and unconscious expectations and projections, thus influencing the balance of the two dimensions. Based on their own experiences parents may encourage or hinder attachment or autonomy in a manner which is inadequately stressing or too early. We present the typology of relationship disturbances and illustrate the diagnostic and therapeutic procedure by case examples. PMID- 11098647 TI - [Clinical relevance of attachment theory for the psychotherapy of small children and their parents]. AB - Since the beginning of the 1990s, the relevance of the attachment theory has gained increasing significance for psychotherapeutic practice. Due to the lack of empirical proof, the only feasible approach, for the time being, are conceptional deliberations, which, however, offer useful ideas for the practical work. Based on an case record obtained from the psychotherapeutic treatment of a 4-month-old suckling and his parents, this article attempts to discuss the incorporation of attachment-theoretical concepts into infant-parent psychotherapy. Infant-parent psychotherapy constitutes a treatment method which, on the initiative of Mechthild Papousek, founder of the "Munich interdisciplinary intervention Program", has enjoyed larger-scale dissemination and deals with regulatory disturbances in infancy. Irrespective of core symptoms and their etiology, regulatory disturbances often lead to impaired mother-child interactions and formation of secure attachment organization of the child. The observation of essential, attachment-relevant contents, such es, for instance, the child's attachment and exploratory behavior an the maternal sensitivity to signals emanating from the child gives rise to novel viewpoints regarding therapeutic interventions. PMID- 11098648 TI - [Fathers in parent-infant psychotherapy]. AB - This paper discusses the relative neglect of the role of the father by researchers and parent-infant psychotherapists. It goes on to describe reasons why the father's role is of developmental significance. Using clinical examples, it is suggested that it is often critical to ensure that fathers are included if the therapeutic work is to have the best chance of success. Two particular aspects of the father's contribution are considered, namely the direct influence that his own "ghosts" may exercise on the parent-infant relationship, and a slightly more indirect role that he may play as the other half of the parental couple. PMID- 11098649 TI - [Use of video feedback in parent-infant counseling and psychotherapy]. AB - Video-based observation and behavioral microanalyses have critically contributed to the international boom of infancy research in the early seventies. Video techniques have opened up a direct scientific access to the broad domains of intuitive behavior and thus to early integrative processes, self regulation, parenting, and preverbal communication. The exploratory potential of video based microanalyses may similarly promote innovative clinical approaches to preverbal processes and mechanisms of developmental psychopathology and disordered parent infant relationships. The article review currently available video-based methods that are used in clinical diagnostics and parent-infant consultation and psychotherapy. The article is based on relevant international publications as well as on the Papouseks' use of behavioral microanalyses during three decades of joint basic research and almost ten years clinical experience from the Munich Interdisciplinary Research and Intervention Program for Fussy Baby's. PMID- 11098650 TI - Volume transmission as a key feature of information handling in the central nervous system possible new interpretative value of the Turing's B-type machine. PMID- 11098651 TI - Integration of wiring transmission and volume transmission. PMID- 11098652 TI - Ultrastructural evidence for diffuse transmission by monoamine and acetylcholine neurons of the central nervous system. PMID- 11098653 TI - Comparative aspects of volume transmission, with sidelight on other forms of intercellular communication. PMID- 11098654 TI - Diffusion of molecules in brain extracellular space: theory and experiment. AB - Volume transmission depends on the migration of informational substances through brain extracellular space (ECS) and almost always involves diffusion; basic concepts of diffusion are outlined from both the microscopic viewpoint based on random walks and the macroscopic viewpoint based on the solution of equations embodying Fick's Laws. In a complex medium like the brain, diffusing molecules are constrained by the local volume fraction of the ECS and tortuosity, a measure of the hindrance imposed by cellular obstacles. Molecules can also experience varying degrees of uptake or clearance. Bulk flow and the extracellular matrix may also play a role. Examples of recent work on diffusion of tetramethylammonium (molecular weight, 74) in brain slices, using iontophoretic application and ion selective microelectrodes, are reviewed. In slices, the volume fraction is about 20% and tortuosity about 1.6, both similar to values found in the intact brain. Using integrative optical imaging, results obtained with dextrans and albumins up to a molecular weight of 70,000 are summarized, for such large molecules the tortuosity is about 2.3. Experiments using synthetic long-chain PHPMA polymers up to 1,000,000 molecular weight show that these molecules also diffuse in the ECS but with a tortuosity of about 1.6. Studies with osmotic challenge show that volume fraction and tortuosity do not vary together as expected when the size of the ECS changes; a model is presented that explains the osmotic-challenge on the basis of changes in cell shape. Finally, new analytical insights are provided into the complex movement of potassium in the brain. PMID- 11098655 TI - Extracellular space diffusion and pathological states. PMID- 11098656 TI - Diffusion of radiolabeled dopamine, its metabolites and mannitol in the rat striatum studied by dual-probe microdialysis. PMID- 11098657 TI - Relationship between glia and the perineuronal nets of extracellular matrix in the rat cerebral cortex: importance for volume transmission in the brain. PMID- 11098658 TI - Glial influence on neuronal signaling. PMID- 11098659 TI - Glial modulation of neural excitability mediated by extracellular pH: a hypothesis revisited. PMID- 11098660 TI - The astrocyte-mediated coupling between synaptic activity and energy metabolism operates through volume transmission. PMID- 11098661 TI - Metabolic trafficking between cells in nervous tissue. PMID- 11098662 TI - Cell volume and water exchange in neural cells monitored by diffusion weighted 1H NMR spectroscopy. PMID- 11098663 TI - Extrasynaptic distribution of monoamine transporters and receptors. PMID- 11098664 TI - Distinct regional differences in dopamine-mediated volume transmission. PMID- 11098665 TI - Geometry and kinetics of dopaminergic transmission in the rat striatum and in mice lacking the dopamine transporter. PMID- 11098666 TI - Evidence for the existence of pulses of dopamine in the extracellular space of the rat striatum. PMID- 11098667 TI - Restoration of dopamine transmission in graft reinnervated striatum. Evidence for regulation of dopamine D2 receptor function in regions lacking dopamine. PMID- 11098668 TI - When it comes to communications between neurons, synapses are over-rated: insights from an animal model of parkinsonism. PMID- 11098669 TI - GABAergic excitation and K(+)-mediated volume transmission in the hippocampus. PMID- 11098670 TI - Spillover and synaptic cross talk mediated by glutamate and GABA in the mammalian brain. PMID- 11098671 TI - Adenosine as a volume transmission signal. A feedback detector of neuronal activation. PMID- 11098672 TI - Dynorphins are endogenous opioid peptides released from granule cells to act neurohumorly and inhibit excitatory neurotransmission in the hippocampus. PMID- 11098673 TI - Neuropeptide spread in the brain and spinal cord. PMID- 11098674 TI - Neuronal mechanisms of synaptic and network plasticity in the lamprey spinal cord. PMID- 11098675 TI - Long distance signalling in volume transmission. Focus on clearance mechanisms. PMID- 11098676 TI - CSF signaling in physiology and behavior. PMID- 11098677 TI - Volume transmission in the year 2000. PMID- 11098678 TI - Sodium channels and the molecular pathophysiology of pain. PMID- 11098679 TI - Chemical mediators of pain due to tissue damage and ischemia. PMID- 11098680 TI - Nociceptor excitation by thermal sensitization--a hypothesis. PMID- 11098681 TI - Protein kinase subtypes involved in injury-induced nociception. PMID- 11098682 TI - Synaptic transmission and plasticity in the superficial dorsal horn. PMID- 11098683 TI - Synaptic mechanisms of hyperalgesia. PMID- 11098684 TI - Silent glutamatergic synapses and long-term facilitation in spinal dorsal horn neurons. PMID- 11098685 TI - Spinal dorsal horn synaptic plasticity: involvement of group I metabotropic glutamate receptors. PMID- 11098686 TI - The functional anatomy of lamina I and its role in post-stroke central pain. PMID- 11098687 TI - Long-term potentiation in single wide dynamic range neurons induced by noxious stimulation in intact and spinalized rats. PMID- 11098688 TI - The role of spinal nitric oxide in the control of spontaneous pain following nociceptive input. PMID- 11098690 TI - Neuroplasticity in the spinal cord of monoarthritic rats: from metabolic changes to the detection of interleukin-6 using mRNA differential display. PMID- 11098689 TI - The role of high-threshold calcium channels in spinal neuron hyperexcitability induced by knee inflammation. PMID- 11098691 TI - Central roles of nociceptin/orphanin FQ and nocistatin: allodynia as a model of neural plasticity. PMID- 11098692 TI - The biological role of galanin in normal and neuropathic states. PMID- 11098693 TI - Plasticity in descending pain modulatory systems. PMID- 11098694 TI - Role of thalamus in pain. PMID- 11098695 TI - Plasticity of pain-related neuronal activity in the human thalamus. PMID- 11098697 TI - Cortex areas involved in the processing of normal and altered pain. PMID- 11098696 TI - Concepts of pain mechanisms: the contribution of functional imaging of the human brain. AB - Functional imaging of the conscious human brain has a solid physiological basis in synaptically induced rCBF responses. We still do not know how these responses are generated, but recent studies have shown that the rCBF response is parametrically positively correlated with functional measures of neuronal activity. Technical advances in both fMRI and PET imaging have improved the spatial and temporal resolution of imaging methods. Further advances may be expected in the near future. Consequently, we now have an important tool to apply to the study of normal and, most importantly, pathological pain. There is a tendency to expect too much of this exciting technique, but the problems we wish to address are complex and will require considerable time, effort, and patience. We now know that the CNS adapts to both peripheral and central nervous system injury, sometimes in beneficial ways, but sometimes with reorganization that is maladaptive. An understanding of the pathophysiology of neuropathic pain is further complicated by the new knowledge, emphasized by functional brain imaging, that pain and pain modulation is mediated, not by a simple pathway with one or a few central targets, but by a network of multiple interacting modules of neuronal activity. Simplified phrenological thinking, with complete psychological functions separate and localized, is appealing, but wildly misleading. It is far more realistic and productive to apply qualitative and quantitative spatial and temporal analyses to the distributed activity of the conscious, communicating human brain. This will not be quick and easy, but there is every reason for optimism in our search for a thorough and useful understanding of both normal and pathological pain. PMID- 11098698 TI - Regional brain oxygenation during phasic and tonic painful stimulation. PMID- 11098699 TI - The functional organization of the brain in chronic pain. PMID- 11098700 TI - Neuroplasticity and clinical pain. PMID- 11098701 TI - Multiple mechanisms of secondary hyperalgesia. PMID- 11098702 TI - Referred pain as an indicator for neural plasticity. PMID- 11098704 TI - Modelling the prolonged effects of neonatal pain. PMID- 11098703 TI - Neurochemical plasticity in persistent inflammatory pain. PMID- 11098705 TI - Role of sensitized pelvic nerve afferents from the inflamed rat colon in the maintenance of visceral hyperalgesia. PMID- 11098706 TI - Cellular and neurochemical remodeling of the spinal cord in bone cancer pain. PMID- 11098707 TI - Altered spinal processing in animal models of radicular and neuropathic pain. PMID- 11098708 TI - Allodynia and hyperalgesia within dermatomes caudal to a spinal cord injury in primates and rodents. PMID- 11098709 TI - Pain following spinal cord injury: pathophysiology and central mechanisms. PMID- 11098710 TI - Sympathetic nervous system: contribution to chronic pain. PMID- 11098711 TI - A role for the endogenous cannabinoid system in the peripheral control of pain initiation. PMID- 11098712 TI - Cellular and molecular mechanisms of opioid action. PMID- 11098713 TI - Pre-emptive analgesia in postamputation pain: an update. PMID- 11098714 TI - Pre-emptive analgesia and surgical pain. PMID- 11098715 TI - Attentional control of pain and the process of chronification. PMID- 11098716 TI - Jorgenson vs. Vener, statistics run amok? PMID- 11098717 TI - Teamwork in healthcare--go for the gold! PMID- 11098718 TI - More than cholesterol: the complexity of coronary artery disease. AB - There have been many recent articles published that emphasize the fact that cholesterol deposits are only one of many mechanisms through which acute coronary artery disease develops. Recently, a meta-analysis shows that only 14% of acute coronary events occur in stenotic lesions in coronary arteries that are greater than 70% occluded. The majority of acute coronary events (68%) occur in coronary arteries that have less than 50% stenotic lesions. The acute coronary syndrome is a very complex and unpredictable disease. Recent information now points to the endothelium as a modulating factor in the pathogenesis of coronary artery disease through the production of nitric oxide and angiotensin-II which maintain the homeostatic environment influencing the progression of coronary artery disease. With dysfunctional endothelium there seems to be an imbalance in terms of angiotensin production with regards to the nitric oxide production. This imbalance tends to promote coronary artery disease in individuals who have multiple risk factors. Furthermore, it has been suggested that certain inflammatory compounds are produced in a very dysfunctional endothelium, thereby propagating or leading to acute coronary syndromes. Specifically, this includes C reactive protein which promotes chronic inflammation at various sites. There are also other acute phase reactants, such as fibrinogen, which may play a role in atherogenesis. Certain statin drugs, as they are called, tend to ameliorate the levels of the above acute phase reactants, while other statins do not. This reduction of coronary events by statins is independent of the LDL lowering benefits from statin drugs. This article delineates some of the beneficial effects of the different statin agents and points out that all statins are not equal in terms of their known lipid beneficial effects. For the practicing physician, choosing a particular statin agent is important. Some have more drug/drug interaction potential as compared with the others because of their inability to be metabolized through the cytochrome P450 system. There are also some, because of their lipophilic and hydrophilic nature, that tend to enter cells more readily than other statin agents. The effects conferred by these subtle differences among the currently available statins tend to be beneficial in patients with low to moderate levels of LDL cholesterol. PMID- 11098719 TI - [What is memory?]. PMID- 11098720 TI - [The assessment of episodic memory: theory and practice]. AB - Episodic memory refers to a system which stores personally experienced events located in time and in space. It is characterized by autonoetic consciousness allowing a subject to be aware of his/her own identity throughout subjective time and to perceive the present as both a continuation of his/her past and as a prelude to his/her future. Current studies of episodic memory should take into account all these features. However, most episodic memory tests are restricted to memory performance and do not really measure episodic memory. In this article, after defining the terms of context and states of awareness, we describe two original tasks designed specially to investigate episodic memory: the 'Quoi-Ou Quand' (What-Where-When) and the 'TEMPau'. The first task allows the study of anterograde amnesia whereas the second consists of an assessment of retrograde amnesia. These two tasks include both scores of memory performance and measures of states of awareness using a procedure derived from the Remember/Know paradigm. PMID- 11098721 TI - [Working memory: a neuropsychologic and cerebral imagery approach]. AB - The working memory model proposed by Baddeley and Hitch has frequently been used in cognitive psychology and neuropsychological studies. This model consists of several interacting components, responsible for the storage and processing of the information stored in working memory. Many neuropsychological and functional imagery data are consistent with that formulation of working memory. Moreover, many cognitive tasks have been specifically designed to explore particular aspects of working memory functioning. Taken as a whole, these data confirm that the working memory model remains a useful theoretical framework to explore memory functioning both in normal subjects and in pathological conditions. PMID- 11098722 TI - Cellular and molecular approaches to memory storage. AB - There has been nearly a century of interest in the idea that information is stored in the brain as changes in the efficacy of synaptic connections on neurons that are activated during learning. The discovery and detailed report of the phenomenon generally known as long-term potentiation opened a new chapter in the study of synaptic plasticity in the vertebrate brain, and this form of synaptic plasticity has now become the dominant model in the search for the cellular bases of learning and memory. To date, considerable progress has been made in understanding the cellular and molecular mechanisms underlying synaptic plasticity and in identifying the neural systems which express it. In parallel, the hypothesis that the mechanisms underlying synaptic plasticity are activated during learning and serve learning and memory has gained much empirical support. Accumulating evidence suggests that the rapid activation of the genetic machinery is a key mechanism underlying the enduring modification of neural networks required for the laying down of memory. These advances are reviewed below. PMID- 11098723 TI - [Memory: a functional imaging approach]. AB - Functional imaging permits us to tackle in a better way one of the most noble functions of the human brain: memory. First of all, it is a way to validate the wide subdivisions proposed by cognitive psychology and clinical neuropsychology such as short-term memory/long-term memory dissociation, episodic memory/semantic memory dissociation and the distinction among working memory subcomponents. Moreover, functional imaging yields a new perspective on the global physiology of the brain. It makes it possible to propose new relationships that are more or less reciprocal between cerebral structures and cognitive functions and raises new fundamental questions. Thus, its first goal is to answer precise questions in the framework of definite cognitive models and it offers the possibility of elaborating a new modelling conception. Finally, functional imaging is a potential tool for a predictive approach to memory functions in both normal subjects and patients. PMID- 11098724 TI - [An animal model of human declarative (relational) memory and of its dysfunction]. AB - The present work was aimed at determining, both at the psychological and at the neurobiological levels, aspects of rodent memory that fall into line with human declarative memory which is known to be selectively impaired in amnesic subjects and during the course of ageing. The ability to compare and to contrast items in memory, and to support inferential use of memories in novel situations (flexibility), were considered to be the two key psychological features of human declarative memory that were altered by both hippocampal lesions and hippocampal dysfunction. Adult and aged mice were trained on learning tasks using two-stage paradigms, the aim of which was to assess memory performance through these two psychological aspects in the same subjects. Results suggest that ageing specifically impairs the ability to both compare and contrast items in memory (declarative/relational memory based on complex associations), without altering memory based on simple S-R associations (procedural memory). Hippocampal lesions in adult mice produced the same dissociation between relational memory (impaired) and procedural memory (spared). Pharmacological experiments showed that, depending on the drug used, the relational memory deficit of aged mice may be selectively reversed (i.e. without changes in procedural memory) and that the behavioural efficacy of certain treatments was shown to parallel their potency in re-establishing normal (i.e. adult) levels of hippocampal plasticity-related mechanisms. Together with previous findings, these results suggest that the storage and use of relational representations would critically depend on the plasticity of hippocampal synapses, which via their connections with cortical areas, would support the storage of associations between perceptual, behavioral and cognitive events. PMID- 11098725 TI - [Pharmacologic models of memory disorders in pathology]. AB - Some drugs induce memory impairments that resemble those observed in neurological and psychiatric disorders. It has therefore been suggested that these drugs might be used as models of these disorders. The concepts and methods developed by cognitive neuropsychology to investigate memory impairments are analysed. The interest and limitations of pharmacological models are discussed and illustrated by the benzodiazepine model of organic amnesia. It is concluded that the validity of a pharmacological model has to be defined in terms of face, construct and predictive validity and assessed according to the aims of modelization. PMID- 11098726 TI - [Animal models of amnesia of alcoholic origin: an amnesia without memory impairment]. AB - Our studies show that chronic alcohol consumption (CAC) in Balb/c mice induces (1) a deficit of spontaneous but not effortful retrieval processes, and (2) a concomitant reduction of anxiety, suggesting a potential interaction between emotional and memory disorders. We have shown that the benzodiazepine agonist, diazepam, induces memory deficits similar to those produced by CAC, whereas administering beta CCM (an inverse agonist of the benzodiazepine receptor) alleviated the memory deficits of alcohol-treated subjects. Parallel neuroanatomical studies have shown that CAC produced cell damage in the mamillary bodies, whereas no major changes were observed in the hippocampus or the frontal cortex, which is involved in long-term consolidation processes. Overall data show that CAC induced amnesia is not due to a dysfunction of the neural networks underlying memory storage processes, but rather results from a difficulty in activating the neural substrates engaged in retrieval processes which depend on emotional, motivational or environmental factors. PMID- 11098727 TI - [Memory, aging and risk factors. Lessons from clinical trials and epidemiologic studies]. AB - Memory troubles associated with age justify treatment and medical attention because they lead to real disabilities in daily living even without dementia, and also because they are early predictors of dementia in Alzheimer's Disease (AD). However, the treatment of these troubles remains uncertain, badly codified and extremely variable depending on the health systems concerned. Moreover, the dominant idea of many physicians, in the general population and even in health authorities, is that it is normal to have memory troubles in the elderly. This fact explains the common attitude of therapeutic nihilism. However, recent progress in the treatment of AD demonstrates that this disease should not be considered as an irremediable phenomenon. Moreover, on the basis of the follow up of an elderly cohort, it is possible to determine subjects at high risk of dementia where preventive interventions could be undertaken. PMID- 11098728 TI - [Pharmacologic treatment for cognitive disorders: an update]. AB - Today, the memory is regarded as a network of memories which is sustained by distinct interconnected neuronal pathways. These diverse neuronal pathways are differentially impaired during cerebral ageing. Consequently, according to their molecular and cellular targets, cognitive enhancing therapies will be specifically applied to some neurodegenerative diseases or to age-related cognitive disorders. During recent decades, cognition enhancers have been devised in order to facilitate or to serve as substitutes for the effects of deficient neurotransmitters, particularly acetylcholine. These pharmacological approaches have allowed the proposition of acetylcholinesterase inhibitors and muscarinic agents. At present, these targets remain in progress but research is orientated on the discovery of nicotinic agents and acetylcholine releasers. Simultaneously, intense research activity has been undertaken around the other major systems of neurotransmission and/or neuromodulation such as the glutamatergic, monoaminergic and peptidergic pathways. Although research has always given precedence to symptomatic treatments (cognition enhancers) over organic treatments (neuroprotective agents), those are the subject of intensive research activity as they could be capable of counteracting the neuronal death and decrease of synaptic plasticity that occur during neurodegenerative diseases and ageing, respectively. PMID- 11098730 TI - [Rehabilitation of memory disorders: objectives, strategies and evaluation]. AB - Memory rehabilitation can be proposed to amnesic patients with mild or moderate isolated memory deficit and with some preserved memory components. It is necessary to study memory with global psychometric tests, specific memory examination and pragmatic study, and to have definite memory aims. The facilitating effects of mental imagery and mnemonic strategies, preserved learning and compensatory aids were analysed. PMID- 11098729 TI - [Memory: therapeutic approach. Clinical evaluation]. AB - Memory assessment in pharmaceutical trials has to date been mainly performed in Alzheimer's disease. North American and European Medical Evaluating Agencies recommend guidelines. Tests need to be simple and short, and sensitive enough to assess changes over a wide range of severity to avoid floor or ceiling effect. They require inter-rater reliability and face validity. They should be at least appropriate for different languages and culture, correlate well with universally accepted estimates of a function and have several parallel forms available to avoid a training effect. Memory is assessed with sub-tests of composite scales assessing a number of cognitive functions including episodic memory. The ADAS-Cog has become the standard instrument since it was used for the approval of tacrine and other cholinesterase inhibitors. Some studies have assessed more specifically different sub-systems of memory. However, autobiographical, remote or prospective memory, as well as metamemory, has not been explored in pharmaceutical trials. PMID- 11098731 TI - [Connectionist models of memory]. AB - Even if computer science, at its birth, had strong links with the neurosciences, it is today mainly oriented toward efficiency and robustness. For example, memory in a computer has few relationships with memory in a living being. Nevertheless, some domains in computer science are interested in this kind of modelling. In particular, connectionism, whose goal is to elaborate artificial neural networks, uses a formalism for its calculus inspired from calculus in the brain. Different kinds of memory that can be emulated by artificial neural networks, inspired by statistics or biology, are presented here. Their relationships with human memory are discussed together with their tentative interest for the biologist or the therapist. PMID- 11098732 TI - [Global cardiovascular risk: definition, evaluation and management strategies. Round table no. 1. XV]. AB - The global cardiovascular risk is the probability of developing a cardiovascular disease within a defined period of time, taking into account several risk factors simultaneously. Born 50 years ago with the analytical epidemiology of cardiovascular diseases, this concept may be considered today as a potential prevention tool integrating the multifactorial aspect of these diseases. Its implementation in clinical practice raises numerous questions related to its estimation, practical use and consequences for the health system. This new approach to cardiovascular diseases does not aim to cure a complaint but to try to avoid its appearance by means of restricting measures. This will modify the relationships between physician and patient and will imply major modifications of our treatment habits. For this strategy to be effective three main prerequisites are needed: management choices will have to be the result of a deal between the patient and the physician, the responsibility for decisions will be endorsed by both of them, and the actions should be maintained over time. PMID- 11098733 TI - [Public health networks: practical aspects and contribution to clinical research. Round table no. 2. XV]. AB - A public health network involves several and different health professionals who work in synergy to achieve a common objective: to improve the management of a given disease. Per se, this objective could not be achieved by each of these professionals if working separately. In practice, existing public health networks ensue from very different legal frameworks and come up against various impediments. They have been developed mainly for the management of chronic and serious diseases (e.g. asthma, diabetes, virus C hepatitis, Parkinson's disease, drug abuse). Public health networks can be a very valuable source of data for clinical and epidemiological research, mainly because patients are followed up over a very long period and information coming from various health professionals (general practitioners, specialists, nurses, etc.) is centralized and recorded in a common database. It can also be useful for pharmacovigilance purposes (assessment of Type A and delayed reactions). In any case, the relative interest of such networks should be regularly assessed by an ad hoc methodology, e.g. an experimental or pseudo-experimental design vs. a reference. Despite the fact that there are relatively few operational networks, they can be of great interest for clinical research. PMID- 11098734 TI - [Self-medication: Is regulation needed...from whom?]. AB - The participants attending the 1999 French Clinical Pharmacological meeting in Giens eventually reached a consensus about the necessity to set up a clear policy regarding self-medication products in France. Despite a wish by French people to take more responsibility for their own health care, self-medication products' use and development in France have been held back for many years for the reasons reported below. However, self-medication could be developed while respecting or even improving public health care by applying the following key principles: (1) the acknowledgement of a consensual definition. 'A self-medication drug is a drug (the guarantees for which are provided by marketing authorization approval and dispensation and advice of a pharmacist) specifically suitable for use without any physician's prescription'; (2) the advertising of self-medication products to the general public is authorized; (3) the safe use of self-medication products is linked to the selected indications and the relevance of the information provided to the general public; (4) the self-medication drug approval procedure could be simplified, managed by a specific AFSSAPS unit; (5) the role of the pharmacist involved, as well as suitable pharmacovigilance, is mentioned; (6) more than 1000 previously marketed drugs without any clear status, the so called 'grey zone drugs', should be taken into consideration. According to the participants at the meeting, it is possible to set up a new clear and dynamic policy for self medication products in France in conformity with EU requirements. PMID- 11098735 TI - [Health foods: definitions, status, public health. Round table no. 4. XV]. AB - 'Alicaments' ('ali' for 'aliment' = 'food' in French, and 'cament' for medicament = drug), is an inappropriate term to designate a developing domain: functional foods, in other words health claims in nutrition. It is not easy to delineate the frontier between foods and drugs; this results in some regulatory problems. The key question is the evaluation of health claims. In this report, we synthesize the discussions of a workshop bringing together specialists from the food industry and nutritionists, scientists and representatives of public health departments. PMID- 11098736 TI - [Evaluation of analgesics in children. Round table no. 5. XV]. AB - Despite recent progress, childhood pain in France is still underevaluated and undertreated and its treatment very heterogeneous among clinicians and health care providers. Pain assessment is often a difficult task in this population. The ontogeny of nociceptive pathways, their maturation, and their functionality during development are not completely understood. Painful sensations primarily involve the child's own experience but can be modified, influenced by his/her parents and the psychosocial environment with their own experience of pain and anxiety. The use of analgesics cannot be precisely quantified using retail sales data. However, the 'Pediadol' survey has demonstrated that their use in children experiencing painful situations is definitely insufficient. Also, another survey conducted among pharmacists showed that at least one-third of analgesics approved for adults are not licensed for paediatric use, sometimes due to inadequate drug formulation. The situation is even more critical for young infants and neonates for whom there are even fewer available approved analgesics. A number of issues need to be addressed in order to improve pain management in children. First of all, evaluation of pain should be generalized. Auto-evaluation should be used in children over 6 years of age. Under 6 years, the initial step would be to further evaluate and validate hetero-evaluation scores. Second, analgesics should be evaluated in order to be licensed for paediatric use. When possible, data obtained in adults or older children should be used and studies in younger infants performed only when necessary. Finally it is of paramount importance to continue to increase paediatricians' and health care providers' awareness of the existence of pain in children and to train them to evaluate pain and treat it appropriately. PMID- 11098737 TI - [Drug-drug interactions: predictive in vitro models of in vivo interactions. Round table no. 6.XV]. AB - Liver drug metabolism is a major source of drug interactions. Three major human in vitro models are employed to detect drug interactions in the preclinical phases of drug development: recombinant enzymes, human liver microsomes and primary human hepatocyte cell cultures. Results obtained from these models may vary during the different phases of drug development. Identification of the metabolic pathways, enzymes involved in drug metabolism (mainly cytochrome P450s), enzyme induction or inhibition allow us to detect the major pharmacokinetic drug interactions which can occur in man and to identify specific populations at risk for such interactions. In vitro models are essential to decide if and which future drug interaction studies should be performed in man. However, the clinical relevance of the potential pharmacokinetic drug interactions detected by these in vitro models remains to be determined and confirmed by human studies. PMID- 11098738 TI - [Scaphoid bone fractures]. PMID- 11098739 TI - [Scaphoid fractures--diagnosis, classification and therapy]. AB - Herbert's classification of scaphoid fractures provides the underlying rationale for treatment according to the fracture type. A CT bone scan in the long axis of the scaphoid is the best means of differentiating between stable and unstable fractures. This is difficult from conventional X-rays due to the particular three dimensional anatomy of the scaphoid. To avoid long-term plaster immobilization and to diminish the risk of a nonunion, unstable fractures of type B should be fixed operatively. With headless screws such as the Herbert screw, which are now available in a cannulated shape, the majority of scaphoid fractures of type B1 and B2 can be stabilized using minimally invasive procedures. Severely displaced fractures require the classical open palmar approach. Proximal pole fractures (B3) are best managed from the dorsal approach, using the Mini-Herbert screw. Stable fractures of type A2 can be treated conservatively in a below-elbow cast or, alternatively, stabilized percutaneously, which allows early mobilization. PMID- 11098740 TI - [Implant positioning in epiphysiolysis capitis femoris. Sequelae of poor outcome and consequences]. AB - Which consequences can be ascribed to the intraarticular position of devices in the operative treatment of a slipped capital femoral epiphysis? Which steps are to define as a standard of a careful procedure? The clinical and radiological results of five cases of a pin or nail penetration after the operative treatment of a slipped capital femoral epiphysis are described, a possible connection of casualties is investigated. The intraarticular position of devices in most cases goes along with an unfortunate clinical outcome and leads to a higher risk of developing chondrolysis. Because of the radiologic overprojection with the femoral head it is possible to oversee the malposition of the pin. Any operative treatment of a slipped capital femoral epiphysis requires a careful intraoperative X-ray examination combined with documentation. With this procedure the bad results of an intraarticular implant position must be ascribed to the reminding risk of a fateful development. PMID- 11098741 TI - [Distal radius fracture--is non-bridging articular external fixator a therapeutic alternative? A prospective randomized study]. AB - A randomised prospective study was carried out to compare non-bridging external fixation using a small A0 external fixator with percutaneous Kirschner wire fixation and plaster in the treatment of distal radial fractures (A2/A3 in the A0 classification). The study involved 40 patients, 20 in each group. The advantages of the non-bridging fixation are: (1) early functional therapy of the wrist, (2) simplified reduction of the fracture, and (3) considerable less restriction of wrist mobility in day-to-day situations. Although the final examination 6 months after treatment showed almost identical functional results, the patients treated with the external fixator benefited from the fact that use of the wrist was virtually free throughout the entire treatment period. PMID- 11098742 TI - [Traumatic sternoclavicular instability. A therapeutic alternative]. AB - Traumatic instability of the sternoclavicular joint is a rare diagnosis. It is usually treated by different bandaging techniques without the possibility of early functional aftercare. In the period between 1 January 1996 and 31 December 1998, a total of eight patients with unstable sternoclavicular joints requiring surgical treatment were treated with Balser plates. The population comprised seven anterior and one posterior dislocations. The results achieved with this alternative treatment option, which offers the advantage of enabling early functional aftercare, are presented. Seven of eight patients were available for follow-up. The eighth patient moved from the area. The Constant Score ranged between 84 and 100 points (average 89 +/- 6.6). PMID- 11098743 TI - [Mechanical behavior of Ilizarov ring fixators. Effect of frame parameters on stiffness and consequences for clinical use]. AB - Using a mechanical testing procedure, various fixator constructs were tested in vitro. In addition, the influence of the passive soft tissue structures on the fixation stiffness was determined. An increased number of Schanz' screws or Kirschner wires led to a comparable increase in stiffness than that observed with an increasing screw or wire diameter. In consequence, larger diameters should be preferred over an additional screw or wire where clinically applicable. With diaphyseal telescoping rods only the axial stiffness decreased. As expected, large ring diameters as well as titanium wires reduced stiffness components. Bracing the outer rings caused a reduction of the overall stiffness. Asymmetric pre-tensioning of the K-wires resulted in a significant reduction of tension in the neighboring wire. Removal of the soft tissues reduced stiffness to a similar extend as experienced in a fibula defect situation. The study demonstrates the correlation between design parameters, passive soft tissues and fixation stiffness and presents guidelines for an optimized fixator design. PMID- 11098744 TI - [5- to 8-year results of total hip endoprosthesis implantation with the Muller straight shaft prosthesis (cemented TiAlNb shaft)]. AB - The aim of this study was to obtain mid-term results after total hip arthroplasty (THA) with cemented titanium stems. In all, 184 patients with a total of 202 THAs (cemented titanium stem) were clinically and radiologically examined after an average follow-up of 6 years (5-8). The recruitment was 86%. The Harris score was determined clinically. Radiologically, the directly postoperative radiographs were compared to the control radiographs according to the recommendations of Gruen et al. and Johnston et al. In 2 cases (1%) septical complications appeared after 2 years, which were treated in two-stage surgery. To date, revisions after loosening have been carried out in 3 cases (1.5%). This is equivalent to a revision rate of 2.5%. Three further cases showed evidence of loosening in more than 5 radiolucent lines (RLL), according to Gruen, making close-meshed controls necessary. Clinically, in all of the 6 cases of aseptic loosening, the Harris score remained above 80 points. In 36 cases, more than one RLL, compared to the postoperative radiographs, was ascertained and mainly found in zones 1, 7, 8, and 14. Substantial subsidence or varus could only be observed in one case. The clinical results in the Harris score were good or excellent in 78% and satisfactory in 20%. With an average of 75 at the time of follow-up, the age of the patients was, according to the indication that only patients above age 60 are to receive cemented-stem prostheses, clearly advanced. The body weight was significantly higher (82 kg; d = 2.4) in those 6 patients having evident RLL, than in patients without RLL. The ratio body weight to surface of the stem was especially different (1.5 kg/cm2 versus 1 kg/cm2; P < 0.005) in the two groups. This did not apply to sex or activity of the patient, size or kind of stem, Harris score, ectopic ossification, or body weight index. The biggest possible stem should be implanted. Not all cemented titanium stem prostheses are necessarily affected with a high rate of loosening at a mid-term follow-up. PMID- 11098745 TI - [Conservative management of articular metacarpophalangeal joint fractures]. AB - We present a splint system for a protected mobilization program after closed reduction of articular proximal phalangeal base fractures. This therapy consists of the periarticular soft tissue and functional anatomy. The soft-tissue around the base of the proximal phalanx leads to a circular stabilization effect. This so called Zancolli Complex (Metacarpophalangeal Retention Apparatus) can be used with the effect of a brace treatment. Treating 31 patients with articular fractures of the proximal phalanx way we found good functional results within a mean follow up period of 2 years after the accident. PMID- 11098746 TI - [Arthroscopy of the upper ankle joint. A retrospective analysis of complications]. AB - The purpose of this retrospective study was to analyse the complications of our arthroscopic procedures at the ankle joint. In all patients we evaluated the treatment protocols and in 111 patients (79.3%) we also evaluated the clinical and radiologic results. All in all we have found complications in 14 patients. In 4 cases we have seen a delayed wound healing, in 2 cases a superficial infection, in 3 cases a deep infection, in 3 cases a neural damage and in 2 cases a phlebothrombosis. Our study show, that the complication rate could be minimized by detailed knowledge of the anatomy, exact preoperative diagnostic and planing of the operation and careful preparation of the portals. The use of a laser also shows a tendency to a lower complication rate. PMID- 11098748 TI - [Thoracic trauma]. PMID- 11098747 TI - [Proprioception after reconstruction of the anterior cruciate ligament. Endoscopic vs. 2 tunnel technique]. AB - ACL reconstruction with patellar tendon autograft is a standard procedure which can be performed arthroscopically with a femoral half tunnel drilled from the joint or using the two-tunnel technique with medial miniarthrotomy and additional femoral approach. The arthroscopic procedure with single incision was supposed to increase stability and to improve proprioception by earlier rehabilitation. 29 patients with ACL deficiency were included in the study. 15 patients were treated endoscopically, 14 patients by using the two-tunnel technique. Proprioception, Lysholm and Tegner scores as well as stability (KT-1000) were assessed preoperatively, 3 and 6 months postoperatively. A deficit of proprioception was assessed in either group preoperatively, which could be improved after 3 months only in the arthroscopic group. 6 months postoperatively, either group showed a restitution of proprioception near full extension and full flexion of the knee. In the mid range position, the proprioception could not be restored. There were no differences in stability and the Lysholm and Tegner scores between the groups after 6 months postoperatively. The comparison between the patients with acute and chronic instability shows a better proprioception in the acute group in the mid range position 6 months after the reconstruction. PMID- 11098749 TI - [Soft tissue reconstruction after scalping injury caused by a dog bite]. AB - The reconstruction of soft-tissue damage on the part of the head which is covered with hair is aimed not only at repairing the damage in such a way that it will withstand mechanical strain but also at the restoring the hair on the scalp. In a 5-year-old boy with a scalping injury involving second- and third-degree soft tissue damage over an area of 20 x 12 cm after a dog bite, surgery was initially performed to transform the damage into purely second-degree damage, which was then repaired with a split skin graft. One year later, the scalp containing the hair was stretched with the aid of two skin expanders over a period of 3 months, so that the split skin graft area could be removed and the hair on the scalp restored. PMID- 11098750 TI - [Delayed diagnosis of odontoid fracture after whiplash trauma of the cervical spine]. AB - Fractures of the odontoid process represent about 10-20% of all diagnosed cervical spine fractures. Approximately 35% of these fractures are classified as Type II according to Anderson and D'Alonzo. They can be potentially unstable especially if combined with a dens displacement of over 6 mm. In severe cervical spine trauma, these fractures do not usually cause difficulties in diagnosis. However, in whiplash injuries, which are very common and only rarely associated with such fractures, the surgical management can be complicated if they are underestimated. These patients can present without significant neurological deficits or the situation can be complicated due to intoxication or additional trauma. Under these circumstances in particular, the diagnosis can be delayed or missed, if no strict protocols for diagnostic effort in all whiplash injuries are employed. A case of delayed diagnosis of an odontoid fracture in a neurological asymptomatic patient after whiplash injury is presented. PMID- 11098751 TI - [Chondroblastoma of the patella with pathological fracture]. AB - Chondroblastoma is a rare benign bone tumor of cartilaginous origin. The typical localization of a chondroblastoma is the epiphysis of long tubular bones--the patella is a very unusual site with an estimated occurrence of 2%. We report a case of a 16-year-old patient with a chondroblastoma of the patella associated with a pathologic fracture. Partial resection of the patella was performed. This is the sixth case in the literature that associates patellar chondroblastoma with fracture. PMID- 11098752 TI - [Mediator modulation in infection and multiple organ failure]. PMID- 11098753 TI - [Comment on the contribution by Wick et al.: "Intrathoracic displacement of the transverse colon as a late complication after abdominal knife stab wound"]. PMID- 11098754 TI - [D. Smrke, J. Princic: Plate and screw osteosynthesis in femur shaft fractures]. PMID- 11098755 TI - [Professional politics]. PMID- 11098756 TI - [Traumatology in Switzerland]. PMID- 11098757 TI - [The new specialty "trauma surgery, orthopedics and orthopedic surgery" assumes definition]. PMID- 11098758 TI - Renal anatomy. Endourologic considerations. AB - The general anatomy of the kidney as applied to minimally invasive surgery is described. The kidney morphometry, the spatial planes, and the perirenal coverings are presented. The kidney's relationship with the diaphragm, ribs, pleura, liver, spleen, and colon is described in importance of intrarenal access. The intrarenal anatomy is also described, based on a large series of three dimensional endocasts. The anatomic relationships of the intrarenal vessels (arteries and veins) with the kidney collecting system are presented and discussed with respect to intrarenal access by puncture, for endopyelotomy and for nephron-sparing operations. PMID- 11098759 TI - Renal physiology. Laparoscopic considerations. AB - Oliguria is a recognized component of the physiologic effect of increased intra abdominal or retroperitoneal pressure. The cause is multifactorial, emanating from vascular and parenchymal compression, and is associated with systemic hormonal effects. Ureteral obstruction does not play a significant role. These changes are pressure-dependent and are usually not apparent until pressures reach 15 mm Hg or more. This effect is not associated with any histologic pathology or evidence of renal tubular damage. After the release of the pneumoperitoneum or pneumoretroperitoneum, the renal function and urine output return to normal with no long-term sequelae, even in patients with pre-existing renal disease. The entire operative team must understand the physiologic effects of CO2 insufflation, which allows appropriate intraoperative monitoring and management and minimizes intraoperative and postoperative complications. PMID- 11098760 TI - Renal calculi. Percutaneous management. AB - Percutaneous nephrolithotomy has established indications and is performed with high success and minimal morbidity. Patients who have large or hard stones or stones associated with urinary obstruction are candidates for a percutaneous procedure. When the certainty of the final result is important, the patient should have a PNL. In general, the best treatment for SWL failure is not more SWL; such patients usually should have an endoscopic procedure. PMID- 11098761 TI - Ureteropyeloscopic treatment of ureteral and intrarenal calculi. AB - Endoscopic lithotripsy is an essential part of the armamentarium at any complete stone treatment center. It is first-line therapy for complicated upper urinary tract calculi and for patients who fail primary extracorporeal shock wave lithotripsy. In the presented series, ureteroscopy is defined as a safe and particularly effective treatment for upper urinary tract calculi. Endoscope miniaturization, the Holmium laser, and refined surgical technique have positive results. Complications are less frequent today, even with in the most complex large stone burdens being addressed in a retrograde ureteroscopic way. PMID- 11098762 TI - Percutaneous management of caliceal diverticula. AB - The diagnosis of a caliceal diverticulum may be serendipitous or established owing to patient symptoms. Once the decision to treat a diverticulum has been made, a percutaneous approach should be considered. If stones are present, complete stone removal and obliteration of the diverticulum should be the goals of surgery. The authors prefer the direct puncture technique whenever possible to limit the risk for bleeding and to facilitate stone removal. Use of a percutaneous approach in properly selected patients affords high success rates and results in few complications. PMID- 11098763 TI - Calyceal diverticula. Ureteroscopic management. AB - In selected cases, RIRS management of calyceal diverticula and its related problems has been shown to be more efficacious than ESWL monotherapy and avoids the potential complications and discomfort of percutaneous and laparoscopic procedures. The advent of the Holmium laser energy source, with innovations such as the flexible ureteroscope and the tipless stone basket, have expanded the role of RIRS in the management of calyceal diverticula and associated problems. Presently, RIRS is the initial treatment choice for the management of low to moderate stone burdens that are contained in calyceal diverticula or trapped behind any narrowed intrarenal segment (e.g., infundibular stenosis). If repair of the stenotic segment is not successful, thereby excluding stone treatment, then under the same anesthesia, the patient undergoes a percutaneous antegrade renal surgery. The authors feel that the percutaneous approach as a first choice is more suitable for posteriorly located diverticula with a large stone burden. In selected cases of anteriorly located diverticula with large stone burdens, the laparoscopic approach is more expedient. PMID- 11098764 TI - Caliceal diverticulum and hydrocalyx. Laparoscopic management. AB - Laparoscopic treatment of caliceal diverticula or hydrocalyces is an excellent choice for anterior cavities without significant overlying renal parenchyma that are large or that have an endoscopically inaccessible neck and either a narrow neck or large stone burden. PMID- 11098765 TI - Renal cystic disease. Laparoscopic management. AB - Laparoscopy offers a safe and efficacious means of ablating symptomatic simple renal cysts while conferring the usual benefits of shorter hospital stay, quicker convalescence, and reduced postoperative pain, although no direct comparison with open surgery has been performed. For indeterminate, complex renal cysts, laparoscopic exploration may spare the patient a morbid open operation to assess a cystic lesion of indeterminant risk. Although laparoscopic removal of kidneys with ADPKD remains a technically challenging exercise, centers of laparoscopic expertise have demonstrated the safety and feasibility of the procedure, thereby expanding the benefits of laparoscopic surgery to patients traditionally relegated to open surgical management. PMID- 11098766 TI - Ureteropelvic junction obstruction. Retrograde endopyelotomy. AB - The modern day treatment of UPJO with retrograde endopyelotomy continues to evolve as experience and knowledge progress. Use of the straight lateral incision and selective use of spiral CT angiogram has refined treatment decisions with retrograde endopyelotomy further. The authors' decision-oriented approach offers guidelines for the practicing urologist. Ultimately, it is up to the urologist and the patient to select the best approach for each clinical scenario. PMID- 11098767 TI - Percutaneous endopyelotomy. AB - Percutaneous endopyelotomy, introduced over 15 years ago, is a well-established alternative to open operative pyeloplasty for management of ureteropelvic junction (UPJ) obstruction. Although several variations of the technique have been described, the goal in all cases is to develop a full thickness incision though the obstructing proximal uretra that extends out to the peripyeloureteral fat and heals over an internal stent. Though a percutaneous endopyelotomy can be considered for almost any patient with primary or secondary UPJ obstruction, it is particularly valuable in the setting of upper tract stones that can then be managed simultaneously. This article reviews the indications, techniques, and outcomes of percutaneous endopyelotomy. PMID- 11098768 TI - Laparoscopic pyeloplasty. AB - Laparoscopic pyeloplasty must be compared with open surgery in terms of efficacy and with endopyelotomy in terms of morbidity. All of the series published so far show that the results of laparoscopic pyeloplasty equal those of open surgery. Laparoscopy is associated with a lower morbidity; therefore, it is preferable to open surgery. The morbidity of endopyelotomy is also low, at least when it is performed in a retrograde fashion. The results of endopyelotomy are poor if UPJ obstruction is caused by crossing vessels. In addition, endopyelotomy in this clinical setting carries the risk of hemorrhage. Most adults with symptomatic UPJ obstruction present with crossing vessels at the UPJ. These patients benefit from laparoscopy, and endopyelotomy should be reserved for patients with true intrinsic stenoses. For this reason, preoperative investigation using contemporary imaging techniques is of crucial importance to be able to select the most appropriate surgical method for a given patient. Laparoscopic dismembered pyeloplasty is technically feasible but difficult. The authors prefer nondismembered techniques that yield equally good results in selected patients. Nondismembered pyeloplasty as described by Fenger is easy to perform and well suited for laparoscopy. PMID- 11098769 TI - Laparoscopic radical nephrectomy for cancer. AB - Laparoscopic radical nephrectomy is a rapidly emerging technique for the treatment of renal cell carcinoma. Surgeons at multiple institutions have reported excellent technical results with this procedure, with encouraging safety and efficacy data and low complication rates comparable with the rates in open radical nephrectomy. Although debate continues regarding the pros and cons of the transperitoneal versus retroperitoneal approach and regarding morcellation versus intact specimen extraction, laparoscopic radical nephrectomy is beginning to approach standard-of-care status at select institutions for tumors less than 8 cm in size. Although generally accepted indications for laparoscopic radical nephrectomy include T1-T2N0M0 tumors, increasing experience and operator confidence have allowed expansion of these indications to include select patients with nodal disease, preoperatively staged level I renal vein thrombus, cytoreductive surgery before immunotherapy protocols, and the rare patient with a laterally directed locally invasive (pT4N0M0) renal cell carcinoma. PMID- 11098770 TI - Laparoscopic partial nephrectomy. The European experience. AB - Laparoscopic partial nephrectomy is technically difficult but oncologically effective. The operation should be performed in centers with expertise. Hemostasis can be achieved using bipolar coagulation and fibrin glue-coated cellulose. Further studies will determine whether less invasive alternatives (focused ultrasound, cryotherapy) will meet the high standard of open (or laparoscopic) nephron-sparing surgery for small renal cell carcinoma. PMID- 11098771 TI - Primary percutaneous approach to upper urinary tract transitional cell carcinoma. AB - The optimal approach to upper tract TCC remains to be redefined. A routine nephroureterectomy for every filling defect in the upper urinary system, even in the case of a normal contralateral kidney, constitutes an unnecessary mutilation in more than two thirds of the cases. Nephroureterectomy does not reduce the need for a long-term cystoscopic follow-up because of the high rate of bladder tumor recurrence that may happen years later after nephroureterectomy. Relying solely on radiography and cytology, lacking sensitivity and specificity, to recommend a nephroureterectomy is against the principles of oncologic surgery, especially now that preoperative histologic proof is easy to obtain endoscopically without compromising cancer control. Ureteroscopy, rigid and flexible, provides a complete assessment of the upper urinary system. Biopsy specimens taken with ureteroscopy may be sufficient for grading but less adequate for staging of the tumor. The authors reserve ureteroscopy for ureteral tumors and small (< 1.5 cm) single tumors of the renal pelvis. They approach large or multiple tumors of the renal pelvis percutaneously, in which a full histologic assessment is possible along with a complete resection of the tumor. The decision on the therapeutic approach is made only after the final pathologic report is reviewed. Grade I and grade II superficial disease (Ta, T1) can be treated endoscopically with minimal morbidity and with an efficiency comparable with the standard more invasive nephroureterectomy (Table 5). The indications for endourologic treatment in these cases can be extended safely beyond a solitary kidney or a high surgical risk to include any healthy individual with a normal contralateral kidney who is willing to commit to a rigorous lifelong follow-up. Patients with grade II T1 lesions require a more vigilant follow-up. For grade III Ta disease, more caution should be exercised in selecting these patients for elective endourologic management. When criteria of good prognosis are found, such as absence of carcinoma in situ, presence of diploidy, low p53 expression and a single-tumor, endoscopic management can be offered [table: see text] with a closer follow-up and resorting always to immediate nephroureterectomy at the first evidence of upstaging. Because of the high incidence of recurrence and progression, elective endourologic management for grade III T1 tumors is not recommended. Endoscopic conservative surgery still can be offered in the cases of a solitary kidney or chronic renal insufficiency or for poor surgical candidates. Patients with localized stages (T2, T3) TCC should be offered immediate nephroureterectomy. The authors do not expect adequate endoscopic extirpation with muscle invasive tumors. Although the tissue removed may include deep layers, deep resection is precluded by the thin renal pelvic wall and the associated risk for perforation. Patients with more extensive disease (T3, T4) have a bad prognosis regardless of the form of therapy. Achieving local control percutaneously while preserving as many nephrons as possible for the future chemotherapy can be a reasonable option. PMID- 11098772 TI - Ureteroscopic management of patients with upper tract transitional cell carcinoma. AB - Endoscopic therapy for the management of upper urinary tract TCC is mainly indicated for patients with an anatomically or functionally solitary kidney, renal insufficiency, bilateral tumors, or severe medical comorbidity. It may be a reasonable alternative to distal ureterectomy with bladder-cuff resection in individuals with low-grade superficial distal ureteral tumors. Although use of this approach has been suggested for treating standard patients with low-grade, low-stage collecting system tumors, this recommendation should not be embraced until more supporting evidence is generated. The efficacy of adjuvant therapy for the prevention of recurrent or progressive disease needs to be defined. If current adjuvant strategies prove ineffective, alternative ones will need to be developed. It is anticipated that advancements in endoscopic technology will facilitate the performance of this type of surgery in the future. PMID- 11098773 TI - Laparoscopic nephroureterectomy. A new standard for the surgical management of upper tract transitional cell cancer. AB - Laparoscopic nephroureterectomy for upper tract TCC still remains somewhat controversial. Unlike laparoscopic radical nephrectomy, which has become widely accepted, LNU is still in its earliest stages. Although there are obvious benefits for the patient who has LNU--less pulmonary complications, less postoperative discomfort, a shorter hospital stay, a better cosmetic result, and a brief convalescence--there are significant concerns. The longer operative time creates a negative financial and professional inducement to learn this technique. Operative times need to fall into the 4-hour range or less to make the procedure cost-effective. Analysis of the efficacy of laparoscopic nephroureterectomy as a curative treatment modality is important. In the short-run, LNU seems to provide similar results to open nephroureterectomy for upper TCC. Although concerns over port site seeding, bladder recurrence, and intraperitoneal seeding have been voiced, these problems have not occurred. The higher incidence of local recurrence noted in the authors' series, however, is of concern and remains an unsettled issue. Despite these local recurrences, the overall cancer survival for a given grade and stage of upper tract TCC seem to be similar to survivals recorded after open nephroureterectomy. Still, the number of LNU cases remains small, and follow-up is brief. These patients need to be monitored closely, with follow-up CT scans over the next decade. The authors believe that there are still several significant hurdles standing in the path of LNU before it can become a widely accepted procedure. Issues of cost, training, and long-term efficacy must be answered definitively. To obtain these types of data, it will be necessary to create a multi-institutional, cooperative study to obtain sufficient numbers of patients with a more than 5-year follow-up on which to base future recommendations. PMID- 11098774 TI - Renal transplantation: laparoscopic live donor nephrectomy. AB - Laparoscopic nephrectomy is now performed at many centers worldwide. This technique for organ harvesting offers less postoperative pain, quicker convalescence, and an optimal cosmetic result when compared with the traditional open approach. With experience, the laparoscopic technique is accomplished without compromise to donor safety or allograft function, and complications are comparable with the rates in open historic series. Longer operative times and the need for disposable equipment result in greater hospital costs; however, the quicker convalescence permits patients to resume activity sooner, allowing marked cost savings for patients and employers. The laparoscopic technique is associated with a steep learning curve. Launching a successful laparoscopic living donor program requires a dedicated coordinated effort involving physicians, nurses, and hospital administration. The ultimate impact of this technique on the willingness of individuals to donate has not yet been determined. PMID- 11098775 TI - Renal transplant complications. Minimally invasive management. AB - Advancements in endourology, laparoscopic urology, and interventional radiology continue to influence the contemporary management of renal transplant complications. The successful implementation of these minimally invasive therapies significantly relies on careful patient selection; not all renal transplantation complications are suitable or amenable for this form of management--true for transplant ureteral complications and less so for other potential complications. With such a strategy, renal transplant complications can be managed efficiently and effectively with these minimally invasive modalities to minimize further recipient morbidity while also minimizing potential risk to the recipient and for the renal allograft. PMID- 11098776 TI - Renal surgery in the new millennium. AB - In the not too distant future, the minimally invasive renal surgeon will be able to practice an operation on a difficult case on a three-dimensional virtual reality simulator, providing all attributes of the real procedure. The patient's imaging studies will be imported into the simulator to better mimic particular anatomy. When confident enough of his or her skills, the surgeon will start operating on the patient using the same virtual reality simulator/telepresence surgery console system, which will permit the live surgery to be conducted by robots hundreds of miles away. The robots will manipulate miniature endoscopes or control minimally or noninvasive ablative technologies. Endoscopic/laparoscopic footage of the surgical procedure will be stored digitally in optical disks to be used later in telementoring of a surgery resident. All this and more will be possible in the not so distant third millennium. PMID- 11098777 TI - Laparoscopic cryoablation of renal masses. AB - Laparoscopic cryoablation seems to be an effective treatment modality for small peripheral renal tumors. The technique is minimally invasive, has a rapid learning curve, results in minimal blood loss and morbidity, and, to date, has demonstrated precise reliable ablation of small renal neoplasms. Long-term follow up is necessary to confirm the absence of local tumor recurrence or distant or port-site metastases. PMID- 11098778 TI - Public health consequences among first responders to emergency events associated with illicit methamphetamine laboratories--selected states, 1996-1999. AB - Methamphetamine, a central nervous system stimulant, is manufactured in illicit laboratories using over-the-counter ingredients. Many of these ingredients are hazardous substances that when released from active or abandoned methamphetamine laboratories can place first responders at risk for serious injuries and death. In 16 states, the Agency for Toxic Substances and Disease Registry maintains the Hazardous Substances Emergency Events Surveillance (HSEES) system to collect and analyze data about the morbidity and mortality associated with hazardous substance-release events. Based on events reported to HSEES during 1996-1999, this report describes examples of events associated with illicit methamphetamine laboratories that resulted in injuries to first responders in three states, summarizes methamphetamine-laboratory events involving injured first responders, and suggests injury prevention methods to protect first responders. PMID- 11098779 TI - Progress toward poliomyelitis eradication--Eastern Mediterranean Region, 1999 September 2000. AB - In 1988, the Regional Committee for the Eastern Mediterranean Region (EMR) of the World Health Organization (WHO) adopted a resolution to eradicate poliomyelitis from the region by 2000. Since then, substantial progress has been made in vaccination and surveillance and, by the end of the year, 19 of the 23 EMR countries are expected to have interrupted poliovirus transmission. This report summarizes progress toward this goal from January 1999 through September 2009. PMID- 11098780 TI - Shortage of tetanus and diphtheria toxoids. AB - A temporary shortage of adult tetanus and diphtheria toxoids (Td) in the United States has resulted from two coincident situations: 1) a decrease in the number of lots released by Wyeth Lederle (Pearl River, New York), and 2) a temporary decrease in inventory of vaccine following routine maintenance activities at the production facilities by Aventis Pasteur (Swiftware, Pennsylvania) that lasted longer than anticipated. Approximately one half of the usual number of Td doses has been distributed this year. Although there have been no decreases in production of tetanus toxoid (TT), availability is low because of increased use during the Td shortage. On the basis of information provided by Aventis Pasteur, the Public Health Service expects vaccine supplies to be restored early in 2001. Until then, Aventis Pasteur will be limiting orders to assure the widest possible distribution of available doses. PMID- 11098781 TI - Ischemic bandage injuries: a case series and review of the literature. AB - OBJECTIVE: To determine the prognosis and distribution of ischemic injuries caused by inappropriate bandaging of the lower limb in dogs and cats. STUDY DESIGN: Retrospective clinical study. ANIMAL POPULATION: Eleven client-owned animals, including 9 dogs and 2 cats with a history of injuries consistent with incorrect application of a bandage. METHODS: Medical records for dogs and cats referred to the Queen's Veterinary School Hospital with limb wounds between 1995 and 1999 were reviewed for clinical history and referring veterinary surgeons' reports, indicating that the injury was directly related to the application of a bandage to the limb. RESULTS: The indications for bandage application included 2 postoperative cruciate ligament ruptures, 2 lacerations, 3 internal fixations, an onychectomy, a shoulder dislocation, a dog bite, and a tendon strain. None of the patients had other body systems involved or underlying or concurrent diseases. Five different types of bandage were described (support, Robert Jones, pressure, splint, and Velpeau), and no particular age or breed was overrepresented. Of the 11 animals, 5 required full-thickness skin grafts, 3 had to have digits amputated, and 2 required limb amputations. Nine animals survived, but only 4 became fully functional on the affected limb. CONCLUSIONS: Bandage-related injuries are potentially serious sequelae to a routine procedure. A guarded prognosis should be given when there is loss of deeper structures. CLINICAL RELEVANCE: A review of bandaging principles is presented. Method of application, choice of materials, and close monitoring of the bandage are important factors in preventing iatrogenic injury. PMID- 11098782 TI - Ureteral obstruction after ureteroneocystostomy in dogs assessed by technetium TC 99m diethylenetriamine pentaacetic acid (DTPA) scintigraphy. AB - OBJECTIVE: To use technetium Tc 99m diethylenetriamine pentaacetic acid (99mTc DTPA) renal scintigraphy to monitor ureteral obstruction after ureteroneocystostomy in a canine model of partial ureteral obstruction. STUDY DESIGN: Experimental study. ANIMALS: Eight normal adult dogs. METHODS: Partial ureteral obstruction was created in 8 dogs by incomplete ligation of the terminal right ureter. Two weeks later, ureteroneocystostomy was performed in 7 dogs with unilateral partial ureteral obstruction and in 1 dog that had developed bilateral partial ureteral obstruction. 99mTc-DTPA scintigraphy was performed intermittently for 2 weeks after ureteroneocystostomy. Renal transit time of each kidney, as assessed by the time to maximal uptake (time of peak), and glomerular filtration rate, as assessed by percentage of kidney uptake of the radiopharmaceutical between 1 and 3 minutes, were estimated. Comparison between affected and nonaffected kidneys was performed with the Wilcoxon rank sum test. RESULTS: Unilateral partial ureteral obstruction was induced successfully in 7 dogs. In 1 dog, bilateral partial obstruction was induced inadvertently. After ureteroneocystostomy, percentage of kidney uptake of 99mTc-DTPA was low in 4 affected kidneys. The uptake returned to within normal limits in 2 of the kidneys during the observation period. The time activity curve had a more rounded appearance or was increasing continuously for all affected kidneys. A significant increase in renal transit time was observed 2 and 4 days after ureteroneocystostomy. Transit time progressively returned to normal by 4 to 11 days for all affected kidneys except 1. CONCLUSION: Ureteroneocystostomy resulted in persistent partial ureteral obstruction for 4 to 11 days as determined by 99mTc-DTPA scintigraphy. CLINICAL RELEVANCE: 99mTc-DTPA scintigraphy may be a useful procedure for monitoring renal function and ureteral obstruction after ureteroneocystostomy. Persistent partial ureteral obstruction may be seen 1 to 2 weeks after ureteral reimplantation in dogs with previously existing dilated ureters. PMID- 11098783 TI - Definition and determination of acetabular component orientation in cemented total hip arthroplasty. AB - OBJECTIVE: To describe the spatial orientation of the cemented acetabular component in cemented total hip arthroplasty, based on a ventrodorsal and lateral radiographic projection of the pelvis. METHODS: Equations were derived by using trigonometric relationships that describe the radiographic rotation about the longitudinal pelvic axis (alpha), transverse pelvic axis (beta), acetabular inclination (phi), acetabular inclination corrected for longitudinal pelvic rotation, version (phiC), acetabular version (theta), acetabular version corrected for longitudinal pelvic rotation (thetaC), acetabular inclination corrected for transverse pelvic rotation (phi(beta)), and acetabular version corrected for transverse pelvic rotation (theta(beta)) RESULTS: Alpha was calculated by using the equation alpha = sin(-1) (x/y) where x is the transverse distance between the dorsal spinous processes and the center of the pubis on a ventrodorsal radiograph and y is the distance from the pubis to the dorsal aspect of the first coccygeal vertebra perpendicular to the long axis of the pelvis on a lateral radiograph. Phi was calculated from the long axis (LA) and short axis (SA) of the ellipse formed by the radiopaque acetabular marker ring by using the equation phi = sin(-1) (SA/LA). phiC was calculated by using the equation phiC = phi +/- (alpha - tan(-1) (tan alpha cos thetaC)). Theta was determined as previously described. ThetaC was calculated by using the equation thetaC = tan( 1) (tan theta cos alpha). Theta(beta) and theta(beta) were calculated with the equations phi(beta) = tan(-1) (tan theta cos beta) and theta(beta) = theta - tan( 1) (sin beta), respectively. Beta could not be accurately determined from ventrodorsal and lateral pelvic radiographs. CONCLUSIONS AND CLINICAL RELEVANCE: These techniques allow for more accurate postoperative radiographic assessment of acetabular component positioning. This information can then be used in retrospective or prospective analyses examining that effects of implant positioning on clinical outcome. PMID- 11098784 TI - Acetabular component orientation as an indicator of implant luxation in cemented total hip arthroplasty. AB - OBJECTIVE: To determine the sensitivity and specificity with which acetabular component angles of inclination and version could be used, alone or in combination, to predict luxation of cemented total hip arthroplasties (THA). STUDY DESIGN: Comparison of retrospectively selected cases and controls SAMPLE POPULATION: All THA performed at the University of Florida between 1991 and 1998 with the BioMedtrix system and for which at least 2 months of radiographic follow up were available. All THA performed at the University of Georgia with the BioMedtrix system which subsequently luxated. METHODS: Acetabular component inclination angle (IA) and acetabular version angle (VA) were determined for each THA. Data were grouped according to outcome - luxation or no luxation - with the luxated cases from the 2 institutions pooled. Receiver operator characteristic (ROC) analysis was used to evaluate decision rules for using IA and VA as tests for detecting postoperative luxation. Sensitivity and specificity for luxation and 95% confidence bounds were computed with selected values of IA and VA as cut points. RESULTS: The nonluxation group consisted of 68 THA with a median follow up time of 5 months (range, 2-60 months). The luxation group consisted of 12 THA with a mean time to luxation of 36 days. The nonluxation group had a mean +/- standard deviation (SD) IA and VA of 40.3 degrees +/- 8.9 degrees and 71.1 +/- 13.6 degrees, respectively, whereas the luxation group had a mean +/- SD IA and VA of 34.7 degrees +/- 12.6 degrees and 72.9 degrees +/- 16.6 degrees, respectively. An IA cut-point of 37.8 degrees achieved 58.3% sensitivity and 57.4% specificity. A VA cut-point of 73 degrees achieved 75.0% sensitivity and 51.5% specificity. IA and VA considered simultaneously achieved a 50.0% sensitivity and 88.2% specificity. CONCLUSIONS AND CLINICAL RELEVANCE: ROC analysis indicated that both IA and VA considered individually or simultaneously were poor indicators of luxation. Although extreme values of IA may predict luxation with high specificity, the potential for luxation cannot be excluded based on apparently appropriate values of IA and VA. The results of this study also indicate that a successful outcome is possible with a wide range of acetabular component positions. PMID- 11098785 TI - Evaluation of risk factors for luxation after total hip replacement in dogs. AB - OBJECTIVE: To identify risk factors for luxation after canine total hip replacement (THR). STUDY DESIGN: Retrospective study. SAMPLE POPULATION: 256 client-owned dogs that underwent THR. METHODS: Patient data surveyed included signalment, body weight, diagnosis, prior hip surgery, implant size, intraoperative complications, and angle of lateral opening of the acetabular component. RESULTS: Postoperative complications were recorded in 20 cases (7.8%). The most common complication was dorsal luxation which occurred in 12 dogs (4.7%). The interval between joint replacement and luxation ranged from 1 to 116 days (mean, 44 days). In 1 case, luxation was attributable to failure of the repair of an intraoperative fracture of the greater trochanter. Excluding this case, the mean angle of lateral opening in those dogs that sustained luxation was 62 degrees (range, 46 degrees - 75 degrees). The mean angle of lateral opening overall was 48 degrees (range, 18 degrees - 76 degrees). The angle of lateral opening was the only factor that had a statistically significant effect on whether luxation occurred (P = .035). Acetabular revision, performed primarily to reduce the angle of lateral opening, was performed in 8 dogs and successfully prevented subsequent luxation. CONCLUSION: Luxation of the prosthesis is substantially under the control of the surgeon. It is recommended that the acetabular cup be inserted at an angle of lateral opening of 35 degrees to 45 degrees. In those cases of THR luxation in which an inappropriate angle of lateral opening is identified, acetabular revision arthroplasty generally results in a good clinical outcome. PMID- 11098786 TI - Ventral surgical approach to the caudal brain stem in dogs. AB - OBJECTIVE: To develop a safe neurosurgical procedure that accessed the ventral pons and medulla of the dog primarily for the removal of brain stem neoplasms. STUDY DESIGN: In vivo study. METHODS: A cadaver study was performed on mesocephalic dog heads to develop a neurosurgical approach to the ventral brain stem. Based on this study, an approach to the ventral brain stem was developed by basioccipital craniectomy. This procedure was performed on 4 young neurologically normal Beagle dogs to assess morbidity and mortality associated with the procedure. Morbidity was evaluated by subjective criteria, daily complete neurologic examinations, comparison of preoperative and postoperative brain stem auditory evoked response (BAER) tests, and postmortem examinations. RESULTS: Three dogs developed a transient cough but were neurologically normal after surgery. One dog was euthanatized within 12 hours of surgery because of severe postoperative morbidity associated with basilar artery disruption due to improper development of the craniectomy. Prolongations of postoperative BAER latencies were observed in 2 dogs but did not appear to be associated with clinical deficits or histopathologic changes in the brain stem. Minimal histopathologic changes were observed except in the dog with basilar artery disruption. Results of this study indicate that, although technically challenging, this procedure can be performed with minimal morbidity. CLINICAL IMPLICATIONS: The main indication for this procedure is surgical reduction or biopsy of ventrally located brain stem neoplasms in dogs. The major disadvantage is anatomic restrictions that prevent access to laterally oriented ventral brain stem masses. PMID- 11098787 TI - Fertility of mares after unilateral laparoscopic tubal ligation. AB - OBJECTIVE: To develop a technique for laparoscopic tubal (oviductal) ligation and to evaluate pregnancy rates for mares that ovulated ipsilateral or contralateral to the ligated oviduct. STUDY DESIGN: Randomized prospective clinical trial comparing pregnancy rates after unilateral laparoscopic tubal ligation. ANIMALS: Twelve mares of light horse breeds. METHODS: One oviduct in each of 6 mares was surgically ligated with a laparoscopic technique; 6 other mares served as nonligated controls. Mares with unilateral tubal ligations (UTL) were inseminated with 500 million progressively motile sperm during 1 cycle when the dominant follicle was ipsilateral to the ligation site and 1 cycle when the dominant follicle was contralateral to the ligation site. Control mares were bred during 2 cycles regardless of the side of the dominant follicle. Pregnancy examinations were performed on days 12, 14, and 16 after ovulation by transrectal ultrasonography. RESULTS: None of the mares became pregnant when ovulations occurred from the ovary adjacent to the ligated oviduct. All 6 mares became pregnant on the first cycle when an ovulation occurred from the opposite ovary. Control mares became pregnant on 10 of 12 cycles (83.3 %). CONCLUSIONS: UTL was completely effective in preventing pregnancy when ovulation occurred ipsilateral to the ligation site. The surgical procedure did not interfere with the establishment of pregnancy when ovulation occurred from the contralateral ovary. CLINICAL RELEVANCE: UTL may be a clinically useful procedure for preparing a recipient mare for gamete intrafallopian transfer. The recipient mare could be allowed to ovulate and UTL would prevent fertilization of her oocyte but would not interfere with normal corpus luteum formation. The donor oocyte could be placed into the oviduct contralateral to the UTL site. PMID- 11098788 TI - The effects of methylprednisolone on normal and monocyte-conditioned medium treated articular cartilage from dogs and horses. AB - OBJECTIVE: To study in vitro (1) the dose-response relationships between proteoglycan metabolism in normal and corticosteroid-treated articular cartilage; (2) long-term proteoglycan metabolism after treatment of articular cartilage with corticosteroids; and (3) the effect of corticosteroids on proteoglycan metabolism in articular cartilage treated with monocyte-conditioned medium (MCM). STUDY DESIGN: Equine and canine articular cartilage explants were treated with corticosteroids and MCM. Proteoglycan synthesis and degradation were measured by radioactive labeling in short-term culture, and the long-term effect of corticosteroid treatment on proteoglycan metabolism was studied in normal explants. ANIMALS: Two young cross-breed horses and 3 young Labrador retrievers. METHODS: Equine articular cartilage explants were incubated in medium containing methylprednisolone sodium succinate (MPS) at 0, .001, .01, .1, 1, and 10 mg/mL (final concentration) for 1 day and then in fresh medium without MPS. Proteoglycan synthesis was measured by incorporation of sodium [35S]sulfate at 1, 3, 7, 10, and 13 days after initial treatment with MPS. Proteoglycan release was measured from separate explants prelabeled with sodium [35S]sulfate and treated similarly. Equine articular cartilage explants were treated with equine MCM simultaneously with, and 24 hours before MPS, at 0, 0.01, 0.1, 1, or 5 mg/mL for 72 hours. Proteoglycan synthesis and degradation in these explants was compared. Proteoglycan synthesis and degradation were measured similarly in canine articular cartilage explants treated simultaneously with canine MCM and MPS at 0, 0.001, 0.01, 0.1, 1 and 10 mg/mL for 72 hours. Equine articular cartilage explants treated with 0, 0.01, 0.1, 1, and 5 mg/mL of MPS for 72 hours were evaluated histologically. RESULTS: Proteoglycan synthesis in normal equine articular cartilage was severely depressed by 10 mg/mL MPS for 24 hours, and proteoglycan synthesis failed to recover after 13 days of culture in medium without MPS. Cartilage treated with 5 mg/mL MPS had pyknotic chondrocyte nuclei and empty lacunae. Concentrations of 1 and 0.1 mg/mL MPS depressed proteoglycan synthesis in normal equine cartilage explants. For these 2 concentrations, proteoglycan synthesis recovered 2 days after MPS removal and increased significantly (P < .05) 7 days after treatment with MPS compared with controls without MPS. Concentrations of 0.001 and 0.01 mg/mL MPS did not significantly affect proteoglycan synthesis in normal equine cartilage explants. Cumulative proteoglycan loss over 13 days in culture from normal equine explants treated for 24 hours with different concentrations of MPS was not significantly different between treatment groups at any time point. MCM significantly depressed proteoglycan synthesis in both canine and equine articular cartilage explants and significantly increased proteoglycan release. These effects were prevented in the canine explants by simultaneous treatment with MPS at 1 and 0.1 mg/mL, and proteoglycan release induced by MCM in equine articular cartilage was inhibited by 1 mg/mL MPS. CONCLUSIONS: Concentrations of 1.0 and 0.1 mg/mL MPS alleviated articular cartilage degradation in MCM-treated articular cartilage in vitro. These concentrations of MPS in contact with normal cartilage explants for 24 hours are unlikely to be detrimental in the long term to proteoglycan synthesis. The response of articular cartilage to MPS was affected by treatment with MCM so that results of experiments with normal articular cartilage explants may not reflect results obtained with abnormal cartilage. CLINICAL RELEVANCE: It may be possible to find an intraarticular concentration of corticosteroid that protects articular cartilage against cytokine-induced matrix degradation yet not have prolonged or permanent detrimental effects on chondrocyte matrix synthesis. PMID- 11098789 TI - Realignment of the radius in canine antebrachial growth deformities treated with corrective osteotomy and bilateral (type II) external fixation. AB - OBJECTIVE: To identify factors affecting radial alignment after oblique corrective osteotomy stabilized with a type II external fixator and to evaluate the results of this treatment for antebrachial growth deformities. STUDY DESIGN: Retrospective study SAMPLE POPULATION: Twenty-eight dogs with unilateral antebrachial growth deformities treated with acute corrective osteotomy stabilized with a type II external fixator. METHODS: Medical records and preoperative and postoperative radiographs of the affected and contralateral limb were reviewed. Cause of deformity, age, weight, and gender were recorded. Radial length, varus/valgus angulation, and cranial/caudal angulation were measured from radiographs of the treated and contralateral limbs. Preoperative and postoperative angulation and length discrepancy were compared between affected and contralateral limbs. RESULTS: Correction of varus/valgus angle discrepancy was achieved by using acute corrective osteotomy stabilized with type II external skeletal fixation. No significant change was noted for correction of cranial/caudal angle discrepancy or length discrepancy between the affected and control limb. CLINICAL RELEVANCE: Varus/valgus angle deformities can be treated successfully with type III external fixation after oblique corrective osteotomy. Patients with significant length or cranial/caudal angle discrepancies or both that negatively impact function may require the use of hinged circular fixators or other dynamic techniques to achieve adequate correction. PMID- 11098790 TI - Screw fixation in lag fashion of equine cadaveric metacarpal and metatarsal condylar bone specimens: a biomechanical comparison of shaft and cortex screws. AB - OBJECTIVE: To compare acute fixation stability and insertion effort of cortex bone screws with and without a shaft inserted in lag fashion in equine metacarpal (metatarsal, MC(T)III) bone. METHODS: Screw types with independent variables of screw diameter (4.5 or 5.5 mm) and shaft type (without shaft, with 20-mm shaft, or with 25-mm shaft) were studied. Bone specimens cut from distal equine MC(T)III condyles were used. After screw insertion in lag fashion into 2 bone blocks with an instrumented device, shear tests were conducted in a mechanical testing machine. Outcome variables of peak insertion torque, insertion energy, stiffness. yield strength, and displacement at 3 kN of load were compared. RESULTS: The effects of screw design were substantial. Screws with shaft were 30% to 40% stiffer and 60% to 70% stronger than screws without shaft. Screws with shaft could tolerate 80 to 95 kg more force than screws without shaft before yielding. At 3 kN load, the displacement with screws with shaft was 55% to 60% of that with screws without shaft. Screws with a long shaft tended to perform better than those with a short shaft. There was no difference in the shear stiffness, shear yield strength, or shear displacement between the 2 screw diameters. Although larger diameter screws required more insertion effort, and screws with a short shaft required the most insertion energy, these differences were small. CONCLUSIONS: Cortex screws with a long shaft of 4.5- or 5.5-mm diameter provide better stability in equine MC(T)III condyle bone with less insertion effort compared with those with a short shaft or no shaft. CLINICAL SIGNIFICANCE: Cortex bone screws with a shaft inserted in lag fashion should be considered for the fixation of equine MC(T)III condylar fractures. PMID- 11098791 TI - Use of pelvic flexure biopsies to predict survival after large colon torsion in horses. AB - OBJECTIVE: To determine if morphologic evaluation of intraoperative biopsies of the large colon could be used to accurately predict outcome in horses with large colon torsion. STUDY DESIGN: Clinical study. ANIMALS: Fifty-four horses with large colon torsion. METHODS: A full-thickness biopsy was collected from the pelvic flexure of the ascending colon after correction of naturally occurring colonic torsion. Morphologic changes were evaluated and graded for interstitial tissue to crypt ratio (I:C ratio), percentage loss of superficial and glandular epithelium, and the degree of hemorrhage and edema. These variables were then used to predict survival. RESULTS: Morphologic variables could be used to correctly predict survival or death in 51 horses (P < .0001). This corresponded to a sensitivity of 95.1% (82.2%-99.2%; 95% CI) and a specificity of 92.3% (62.0% 99.6%; 95% CI). Of 6 horses that had colonic resection, 5 survived; an accurate prediction of outcome based on morphologic criteria was made for each horse. CONCLUSIONS: Interpretation of changes in colonic morphology can be used to accurately predict postoperative survival in horses with large colon torsion. CLINICAL RELEVANCE: Use of frozen colonic tissue sections is a rapid, reliable, and relatively inexpensive method for assessing morphologic damage associated with large colon torsion during surgery. Intraoperative evaluation of pelvic flexure biopsies can aid in the prediction of survival and guide surgical judgment as to the need for colonic resection. PMID- 11098792 TI - Application of arthroplasty principles to canine cemented total hip replacement. PMID- 11098793 TI - Disturbed functional brain interactions underlying deficient tactile object discrimination in Parkinson's disease. AB - Somatosensory discrimination of cuboid objects was studied in a group of healthy volunteers and patients with Parkinson's disease using regional cerebral blood flow (rCBF) measurements obtained with positron emission tomography (PET) and 15O labeled water [H2 15O]. A 6-[18F]-fluoro-L-dopa (FDOPA) PET scan demonstrated that the patients may be grouped into those with normal and those with abnormally lowA FDOPA uptake in the caudate nucleus. The categorical group comparisons revealed that task-induced rCBF increases were deficient in bilateral motor and sensory cortical areas in the Parkinson patients. Moreover, deficient rCBF increases were evident in the mesial and right dorsolateral prefrontal cortex for patients in a more advanced disease state, who showed low FDOPA uptake in the caudate nucleus. A principal component analysis (PCA), performed on the rCBF data, identified three patterns (principal components, PCs) that differentiated patients from normals. The first PC represented a right-hemisphere dominant, bilateral group of brain areas known to be involved in tactile exploration. A second PC reflected a cortical-subcortical pattern of functional interactions, comprising cortical areas important for working memory processes. The third group differentiating PC revealed a pattern of functional interactions involving bilateral temporo-parieto-occipital association cortices, which was consistent with a hypothesized supramodal network necessary for object discrimination. In an additional subgroup analysis, greater expression of the third PC pattern predicted greater caudate FDOPA uptake in patients. Our neuroimaging data revealed a disturbance of distinct patterns of brain functional interactions related to the sensorimotor deficit in Parkinson's disease and to deficits of cognitive information processing deficits in the more advanced stage of Parkinson's disease. PMID- 11098794 TI - What have Klingon letters and faces in common? An fMRI study on content-specific working memory systems. AB - Neuroimaging studies show that prefrontal, premotor, and parietal cortical regions are part of a working memory network that supports the active retention of information. In two experiments we used fMRI to examine whether prefrontal and posterior cortical areas are organized in a content-specific way for object and spatial working memory. Subjects performed a delayed matching-to-sample task modified to allow the examination of content-specific retention processes, independent of perceptual and decision-related processes. In Experiment 1, either unfamiliar geometrical objects (Klingon letters from an artificial alphabet unknown to the participants) or their spatial locations had to be memorized, whereas in Experiment 2, either unfamiliar faces or biological objects (butterflies) were actively memorized. All tasks activated a similar cortical network including posterior parietal (banks of the intraparietal sulcus), premotor (banks of the inferior precentral sulcus) and prefrontal regions (banks of the inferior frontal sulcus), and the presupplementary motor area (pre-SMA). For geometrical objects and faces for which strategic semantic processing can be assumed, this activation was larger in the left than in the right hemisphere, whereas a bilateral or right dominant distribution was obtained for butterflies and spatial locations. The present results do not support the process-specific or content-specific view of the role of the prefrontal cortex in working memory task. Rather, they suggest that the inferior prefrontal cortex houses nonmemonic strategic processing systems required for response selection and task management that can flexibly be used across a variety of tasks and informational domains. PMID- 11098795 TI - Brain processing of visual sexual stimuli in human males. AB - Despite its critical sociobiological importance, the brain processing of visual sexual stimuli has not been characterized precisely in human beings. We used Positron Emission Tomography (PET) to investigate responses of regional cerebral blood flow (rCBF) in nine healthy males presented with visual sexual stimuli of graded intensity. Statistical Parametric Mapping was used to locate brain regions whose activation was associated with the presentation of the sexual stimuli and was correlated with markers of sexual arousal. The claustrum, a region whose function had been unclear, displayed one of the highest activations. Additionally, activations were recorded in paralimbic areas (anterior cingulate gyrus, orbito-frontal cortex), in the striatum (head of caudate nucleus, putamen), and in the posterior hypothalamus. By contrast, decreased rCBF was observed in several temporal areas. Based on these results, we propose a model of the brain processes mediating the cognitive, emotional, motivational, and autonomic components of human male sexual arousal. PMID- 11098796 TI - Individual cortical current density reconstructions of the semantic N400 effect: using a generalized minimum norm model with different constraints (L1 and L2 norm). AB - Event-related brain potentials were recorded to study whether verbs and nouns activate topographically distinct cortical generators. Fifteen subjects performed a primed lexical decision task with verb/verb and noun/noun pairs. The relatedness between prime and target items was varied in three steps (unrelated, moderately, and strongly related) and the EEG was recorded from 124 scalp electrodes. The topography of cortical sources of the N400 effect was evaluated by standardized differences scores and by cortical current source estimates which were constrained by the individual MRI-determined cortex anatomy. A behavioral priming effect and a substantial N400 effect was found for both word categories. However, the topography of the grand average N400 effect of verbs and nouns did not differ, neither for raw nor for standardized amplitudes. Cortical current source estimates of the N400 effect revealed a very broad and scattered distribution of active locations with pronounced interindividual differences. Cortical current source estimates obtained with the L1-norm and L2-norm model, respectively, differed in the distribution of sources over the cortex but converged on the same "hot spots." The data give no indication that the N400 effect is generated by word category-specific networks which have a different topography. The marked individual differences are discussed with respect to the involved processes and the current source estimation procedures. PMID- 11098797 TI - Evaluation of octree regional spatial normalization method for regional anatomical matching. AB - The goal of regional spatial normalization is to remove anatomical differences between individual three-dimensional (3D) brain images by warping them to match features of a standard brain atlas. Processing to fit features at the limiting resolution of a 3D MR image volume is computationally intensive, limiting the broad use of full-resolution regional spatial normalization. In Kochunov et al. (1999: Neuro-Image 10:724-737), we proposed a regional spatial normalization algorithm called octree spatial normalization (OSN) that reduces processing time to minutes while targeting the accuracy of previous methods. In the current study, modifications of the OSN algorithm for use in human brain images are described and tested. An automated brain tissue segmentation procedure was adopted to create anatomical templates to drive feature matching in white matter, gray matter, and cerebral-spinal fluid. Three similarity measurement functions (fast-cross correlation (CC), sum-square error, and centroid) were evaluated in a group of six subjects. A combination of fast-CC and centroid was found to provide the best feature matching and speed. Multiple iterations and multiple applications of the OSN algorithm were evaluated to improve fit quality. Two applications of the OSN algorithm with two iterations per application were found to significantly reduce volumetric mismatch (up to six times for lateral ventricle) while keeping processing time under 30 min. The refined version of OSN was tested with anatomical landmarks from several major sulci in a group of nine subjects. Anatomical variability was appreciably reduced for every sulcus investigated, and mean sulcal tracings accurately followed sulcal tracings in the target brain. PMID- 11098798 TI - Quantification of fMRI artifact reduction by a novel plaster cast head holder. AB - In light of artifact-induced high variability of activation in fMRI repeat studies, we developed and tested a clinically useful plaster cast head holder (PCH) with improved immobilization, repositioning, and comfort. With PCH, there were considerably lower levels of translational and rotational head motion components compared to head fixation with conventional restraining straps (CRS). Rotational components cannot be fully compensated by realignment and lead to "false activations." In addition, task-correlated head motion, which highly increases the risk of artifacts, was considerably reduced with PCH, especially in a motion prone subject. Compared with PCH, head motion was 133% larger with CRS in a highly cooperative subject. With a motion prone subject, head motion range was increased by 769% (PCH: 0.9 mm, CRS: 7.8 mm), which may indicate the usefulness of PCH for restless patients. In functional activation maps, PCH alone yielded fewer residual motion artifacts than CRS + image registration. Subject tolerance of the head holder during the long measurement times of up to 2.5 hr was good, and slice orientation on different days confirmed the quality of repositioning. PMID- 11098799 TI - The impact of increased mean airway pressure on contrast-enhanced MRI measurement of regional cerebral blood flow (rCBF), regional cerebral blood volume (rCBV), regional mean transit time (rMTT), and regional cerebrovascular resistance (rCVR) in human volunteers. AB - Contrast-enhanced magnetic resonance imaging (MRI) measurement of cerebral perfusion is a diagnostic procedure increasingly gaining access to clinical practice not only in spontaneously breathing patients but also in mechanically ventilated patients. Effects of increased mean airway pressure on cerebral perfusion are entirely possible. Therefore, the present study used continuous positive airway pressure (CPAP) (12 cm H2O) to study the effects of increased mean airway pressure on cerebral perfusion in volunteers. CPAP significantly reduced regional cerebral blood flow (rCBF) and regional cerebral blood volume (rCBV) but increased regional mean transit time (rMTT) and regional cerebrovascular resistance (rCVR). Active vasoconstriction (e.g., arterial) and/or passive compression of capillary and/or venous vessel areas are the most likely underlying mechanisms. The number of interhemispheric differences in rCBF, rCBV, rMTT, and rCVR found at baseline rose when mean airway pressure was increased. These results, although obtained in volunteers, should be taken into consideration for the interpretation of contrast-enhanced MRI perfusion measurements in mechanically ventilated patients with an increased positive airway pressure. PMID- 11098800 TI - Detecting bilateral abnormalities with voxel-based morphometry. AB - In this article we describe a new method, using SPM99, that searches explicitly for bilateral structural abnormalities. Children with bilateral pathology have a poorer prognosis than children with unilateral damage. After brain injury or disease in childhood, it is thought that rescue of function is only possible if the neuronal substrates of that function are preserved and operational in at least one hemisphere [Vargha-Khadem and Mishkin, 1997]. If this is the case, the detection of bilateral abnormalities would greatly facilitate more accurate prognosis in children with brain injury or developmental disorders. We have therefore developed a technique to detect bilateral abnormalities that uses conjunction analysis with voxel based morphometry. It is illustrated using a group of patients with bilateral hypoxic-ischaemic damage to the hippocampus. The approach is shown to have enhanced specificity and sensitivity relative to conventional unilateral characterisations. PMID- 11098801 TI - The case of dementia: psychiatry or neurology? PMID- 11098802 TI - Autism and Asperger syndrome: coexistence with other clinical disorders. AB - OBJECTIVE: : To provide a clinically useful analysis of the extent to which autism and Asperger syndrome coexist with other disorders. METHOD: Selective review of the literature detailing data pertaining to symptoms and disorders sometimes encountered in connection with autism or Asperger syndrome. RESULTS: A large number of medical conditions, psychiatric disorders and behavioural and motor dyscontrol symptoms are associated with autism and Asperger syndrome. CONCLUSION: Comorbidity is to be expected in autism spectrum disorders -directly or indirectly. Comorbid conditions may be markers for underlying pathophysiology and suggest a more varied treatment approach. There is a great need for in-depth research into this area, meaning that the exclusion criteria of current diagnostic manuals, i.e. those that rule out a diagnosis of autism in some disorders, and a diagnosis of certain other disorders in autism may have to be revised. PMID- 11098803 TI - CAMCOG as a screening instrument for dementia: the Odense study. Cambridge Cognitive Examination. AB - OBJECTIVE: The Cambridge Cognitive Examination (CAMCOG) score is correlated with age and sociodemographic variables. The aim of the study was to determine an individualized CAMCOG cut-off score for dementia, taking such correlates into account. METHOD: From the general population 150 people aged 65-84 years were examined with CAMDEX which includes CAMCOG, and a neuropsychological test battery. Data from 130 non-demented people were entered in setwise multiple regression analyses to identify variables predicting the CAMCOG score. RESULTS: The variables age, social class, matrimonial status, general knowledge and intellectual level explained 66% of the variance of the CAMCOG scores. A cut-off defined as a difference between actual and predicted CAMCOG score < or = -4.41 resulted in a sensitivity of 88.2% and a specificity of 89.2% for very mild to moderate dementia. CONCLUSION: It is possible to determine an individualized CAMCOG cut-off score for dementia in its early stages. PMID- 11098804 TI - Assessment of dysfunctional working models of self and others in schizophrenic patients: a summary of data collected in nine nations. International Research Group. AB - OBJECTIVE: To investigate the cross-cultural feasibility of a new scale for assessing dysfunctional working models of self and others, and to evaluate its discriminative power. METHOD: Schizophrenic patients (N=351), non-psychotic patients (N= 86) and non-clinical subjects (N= 511) collected in 10 centres completed the DWM-S. Current psychopathology was assessed by means of the BPRS. RESULTS: Alpha coefficients were high in all samples. Mean scores on the DWM-S appeared to be comparable in all countries, suggesting cross-national generalizability. No significant correlation was found with sex, age, levels of psychopathology and duration of illness. Discriminant analyses showed that more than 70% of the schizophrenic patients are correctly classified. CONCLUSION: The DWM-S is an easily administered self-report instrument which allows to pinpoint internal dysfunctional working models of self and others in various types of patients. It is a useful tool for case conceptualization, especially when psychotherapeutic interventions are part of the treatment programme. PMID- 11098805 TI - Progressive deterioration of soft neurological signs in chronic schizophrenic patients. AB - OBJECTIVE: Neurological signs are found to be increased in schizophrenia in cross sectional studies. Whether they progress with time is an important issue in addressing the course of the illness. METHOD: The current study investigated different groups of neurological signs in 43 stable chronic schizophrenic patients over a 3-year period using an operationalized instrument. RESULTS: While symptoms and medication have remained largely unchanged in the 3-year period, significant increase in soft neurological signs (SNS) ('motor coordination', 'sensory integration' and 'disinhibition') has been observed. This contrasted with the stability of 'pyramidal', 'extrapyramidal', 'dyskinesia' and 'catatonia' signs. The increase in SNS appears not to be related to age, illness duration, symptoms or medication. CONCLUSION: This finding suggests that SNS represent a marker sensitive to a possible late deterioration process in the course of a schizophrenic illness. PMID- 11098806 TI - Emerging homosexual conduct during hospitalization among chronic schizophrenia patients. AB - OBJECTIVE: The study investigated the emerging homosexual conduct during hospitalization among chronic schizophrenia patients. METHOD: We interviewed 55 male and 58 female chronic schizophrenic patients to investigate their sexual history before and after admission. Those patients were under 45 years old, without significant deteriorated cognitive function, lived in a homogeneous gender chronic ward and did not demonstrate homosexual behaviour before admission. RESULTS: Nineteen patients (16.8%) reported having homosexual conducts during hospitalization. Their characteristics were: 1) having sexual experience before admission; 2) with younger age at first sexual experience; and 3) female patients having more sexual partners before admission. CONCLUSION: A significant proportion of patients need a sexual outlet during long-term hospitalization. Sexual education and counselling are greatly needed. PMID- 11098807 TI - Differences between patients with identified and not identified psychiatric disorders in primary care. AB - OBJECTIVE: The aim of this study was to discover the differences between the primary care patients with a psychiatric disorder whose illness was detected and the patients whose disorder was not detected. METHOD: We collected 1000 randomly selected PC patients. We used SCL-25 as a screening method and PSE as a diagnostic tool. RESULTS: Ninety-one (89.2%) of the interviewed patients received a psychiatric diagnosis. The physicians detected a disorder in 36 (36.9%). A larger part of the undetected group belonged to the highest social groups. Also the SCL-25 mean scores differed significantly, indicating that the symptoms of the undetected cases were milder. The detected cases had higher levels of anxiety and depression, but the difference in anxiety symptoms was greater between the groups. Detection was associated with treatment. CONCLUSION: The GPs should also be aware of psychiatric morbidity in patients with a higher social status, a good level of education and milder symptoms. PMID- 11098808 TI - The relationship between depressive and vital exhaustion symptomatology post myocardial infarction. AB - OBJECTIVE: Many peri-myocardial infarction patients experience decreased wellbeing, which is either conceptualized as depression or as vital exhaustion. The objective of the present study is to investigate whether or not depression and vital exhaustion are separate entities. It was hypothesized that, if depression and vital exhaustion are separate phenomena, the correlation between two depression questionnaires would be higher than those between either of the two depression questionnaires and a vital exhaustion questionnaire. METHOD: Subjects were 143 patients who had recently experienced a first acute myocardial infarction (MI). At 1, 3, 6 and 12 months post-MI, patients completed two self report depression questionnaires (the Zung-SDS and the Depression scale of the SCL-90), and a vital exhaustion questionnaire (the Maastricht Questionnaire). Correlation coefficients were calculated for the two depression questionnaires and the vital exhaustion questionnaire. Furthermore, an exploratory principal component analysis was performed on the combined items of the three questionnaires. RESULTS: Strong and virtually identical correlations were found between the three measures at all four time-points. A one-factor model was the best fit in the exploratory principal component analysis. CONCLUSION: The present results do not support the hypothesized separate conceptual identity of depression and vital exhaustion. PMID- 11098809 TI - The perception of needs for care in staff and patients in community-based mental health services. The South-Verona Outcome Project 3. AB - OBJECTIVE: The present study aims to assess needs for care rated by patients and staff and their agreement on needs assessment in a community-based mental health service by using the Camberwell Assessment of Need (CAN). METHOD: The Italian version of the CAN was used in a sample of 247 patient-staff pairs. RESULTS: Patients and staff showed poor agreement on both the presence of a need and on whether need had been met or not. Higher disability predicted a higher number of patient-rated needs, while higher disability, higher number of service contacts and patient unemployment predicted a higher number of staff-rated needs. Lower global functioning predicted higher disagreement in patients and staff ratings of needs. CONCLUSION: Patients and staff show different perceptions of needs for care and therefore multiple perspectives should be taken into account for planning and providing effective needs-led mental health care. PMID- 11098810 TI - Social networks and functional status in patients with psychosis. AB - OBJECTIVE: To investigate whether large social networks are associated with better functional outcome in psychotic disorders. METHOD: A prospective cohort study of epidemiologically representative patients with psychosis was analysed to examine the relationship between social networks and functional outcome. Standardized validated measures were used for diagnosis, social networks and outcome. RESULTS: There was strong evidence of cross-sectional associations between the size of the social network and the number of active interactions, and functional outcome. Structural equation modelling found weak evidence of an additional longitudinal association. CONCLUSION: There was consistent evidence of a positive correlation between the total size of patients' social networks and the number of active interactions, and functional outcome. The direction of causality cannot be determined by this analysis, although it is suggested that the effect of social networks on functioning is stronger than vice versa. Patients' functioning may improve with interventions that improve social networks. PMID- 11098811 TI - Prolactin and beta-endorphin serum elevations after ECT in manic patients. AB - OBJECTIVE: Alterations in prolactin and beta-endorphin serum levels after ECT are well-established findings in depression. The present study focuses on electroconvulsive therapy (ECT) response patterns of the mentioned parameters in patients suffering from acute mania. METHOD: Following the first three ECTs of a treatment series in 19 patients diagnosed according to DSM-III-R criteria as suffering from mania, blood samples were drawn before, and 20, 30 and 40 minutes after ECT. Serum prolactin and beta-endorphin levels were established in order to gain information about the effects of ECT on different neurotransmitter systems. RESULTS: A significant transient increase in serum prolactin after ECT was found. Furthermore, in females but not males, delta(max)prolactin diminished over the course of treatment as prolactin baseline levels increased. beta-endorphin levels showed a stable transient increase after ECT stimulus regardless from sex or treatment. CONCLUSION: The reported findings reflect those established in depression. This suggests that they are epiphenomenal to ECT. PMID- 11098813 TI - Italy's mad law. PMID- 11098812 TI - Augmentation with sulpiride for a schizophrenic patient partially responsive to clozapine. AB - OBJECTIVE: Schizophrenic patients who are only partially responsive to clozapine pose a therapeutic challenge. In these circumstances some clinicians would consider adding in a second antipsychotic. We present a case report and review evidence for the efficacy of such augmentation strategies. METHOD: Single case report and literature review. RESULTS: The total number of patients in studies and case reports of combining clozapine with other antipsychotics is small. There has been only one randomized controlled trial. This found the addition of sulpiride to clozapine resulted in clinical improvement in some patients. CONCLUSION: Further randomized controlled studies of augmentation of clozapine therapy are needed to provide scientific justification for this clinical practice. PMID- 11098814 TI - The siderophores of Pseudomonas fluorescens 18.1 and the importance of cyclopeptidic substructures for the recognition at the cell surface. AB - The structure of the pyoverdin siderophore of Pseudomonas fluorescens 18.1 was elucidated by spectroscopic methods and chemical degradation. By cross feeding studies structurally closely related pyoverdins containing a C-terminal cyclopeptidic substructure were tested regarding the mutual recognition by the producing strains. Partial recognition of foreign pyoverdins was observed. PMID- 11098815 TI - Anachelin, the siderophore of the cyanobacterium Anabaena cylindrica CCAP 1403/2A. AB - A catecholate siderophore - anachelin - has been isolated from the cyanobacterium Anabaena cylindrica CCAP 1403/2A. The central part of the siderophore is a tripeptide consisting of L-Thr, D-Ser and L-Ser. Its C-terminus is linked amidically to a 1,1-dimethyl-3-amino-1,2,3,4-tetrahydro-7,8-dihydroxyquinolinium system and its N-terminus to 6-amino-3,5,7-trihydroxyheptanoic acid. The 7 hydroxyl group of the latter is esterified with salicylic acid whose carboxyl group is condensed with the 6-amino group to an oxazoline ring. Anachelin is the first genuine siderophore of a cyanobacterium whose structure has been elucidated. PMID- 11098816 TI - Major constituents of the foliar epicuticular waxes of species from the Caatinga and Cerrado. AB - The epicuticular waxes of leaves of four species (Aspidosperma pyrifolium, Capparis yco, Maytenus rigida and Ziziphus joazeiro) from the Caatinga, (a semi arid ecosystem of Northeast Brazil) and four species (Aristolochia esperanzae, Didymopanax vinosum, Strychnos pseudoquina and Tocoyena formosa) from the Cerrado, (a savanna ecosystem covering one third of the Brazilian territory), were analyzed. Six species contained a high content (above 60 microg x cm(-2)) of wax, four of them from the Caatinga. Triterpenoids and n-alkanes were the most frequent and abundant constituents found in the species from both habitats. The distribution of n-alkanes predominated by homologues with 27, 29, 31 and 33 carbon atoms, displayed no consistent differences between species from the two habitats. Lupeol, beta-amyrin, epifriedelinol and ursolic acid were the triterpenoids found. Triterpenoids clearly predominate over alkanes in the waxes from the Cerrado species. The waxes of two evergreen species from the Caatinga yielded n-alkanes as predominant constituents. A comparison of foliar epicuticular waxes of native plants from ecosystems with different hydric constraints is discussed. PMID- 11098817 TI - Two new eudesmane alcohols from Jasonia glutinosa. AB - Two new sesquiterpene alcohols have been isolated from the aerial parts of Jasonia glutinosa D. C. The structure of these sesquiterpenes were characterized by 1D and 2D NMR techniques (DQCOSY. TOCSY, NOESY, HMQC and HMBC) as (11R)-eudesm 4-en-11,12-diol and (11R)-eudesmane-5alpha, 11,12-triol. PMID- 11098818 TI - Sesquiterpene lactones from Inula montana L. AB - Aerial parts of Inula montana were investigated for its sesquiterpenoid composition. Five sesquiterpene lactones, isoinuviscolide, gaillardin, 1beta hydroxy-3beta-acetoxy-eudesm-4(15), 11(13)-dien-12-8beta-olide, pulchellin-C and pulchellin-E were identified for the first time in this plant. One of them, 1beta hydroxy-3beta-acetoxy-eudesm-4(15),11(13)-dien-1 2-8beta-olide, is a novel natural product. The structures of this compounds were established by 1D and 2D NMR spectroscopy. PMID- 11098819 TI - Occurrence and characterization of a UDP-glucose:hydroxamic acid glucosyltransferase isolated from wheat (Triticum aestivum) seedlings. AB - Cyclic hydroxamic acid glucosides are present at high concentrations immediately after germination in wheat (Triticum aestivum L.). Changes in the activity of UDP Glucose:cyclic hydroxamic acid glucosyltransferase (EC 2.4.1.-) in wheat were investigated using the cyclic hydroxamic acids 2.4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) and its 7-methoxy derivative (DIMBOA) as sugar acceptors. Glucosyltransferase activity on both substrates was detected in dry seeds, with activity increasing after imbibition, peaking in shoots and roots 36-48 hours after imbibition and decreasing thereafter. The transience of glucosyltransferase activity was concurrent with the transient occurrence of the hydroxamic acid glucosides [Nakagawa E., Amano T., Hirai N., and Iwamura H. (1995) Phytochemistry 38, 1349-1354], suggesting that glucosyltransferases regulate the accumulation of hydroxamic acid glucosides in wheat seedlings. Two peaks in activity of UDP Glucose:DIMBOA glucosyltransferase were detected using a Mono Q column, indicating the presence of at least two isozymes of this glucosyltransferase. The enzyme in the major peak was purified about 1500-fold and shown to be in a monomeric form with a molecular mass of 47 or 49 kDa. The enzyme reacted strongly with DIMBOA, less so with DIBOA. The enzyme of the minor peak on the Mono Q chromatogram, which was also a monomeric enzyme with a molecular mass of 47 kDa, showed similar substrate specificity to that of the major peak enzyme. PMID- 11098820 TI - Copper accumulation and phosphatase activities of Aspergillus and Rhizopus. AB - Copper accumulation and phosphatase activities of three Aspergillus species resistant to copper were compared to three copper-sensitive Rhizopus species. High level of acid phosphatases and decreased Cu2+-uptake were found with resistant in contrast to sensitive strains. The presence of copper(II) ions in the medium increased the production of acid phosphatases in the resistant A. niger and decreased their activity in the sensitive R. delemar. Copper ions inhibited the activity of A. niger cellular acid phosphatase with a Ki of 8.9x10( 4) M and slightly activated the R. delemar enzyme. PMID- 11098821 TI - Biotransformation of two cytotoxic terpenes, alpha-santonin and sclareol by Botrytis cinerea. AB - Two cytotoxic terpenes, alpha-santonin (1) and sclareol (3) were biotransformed by a plant pathogenic fungus Botrytis cinerea to produce oxidized metabolites in high yields. Alpha-Santonin (1) on fermentation with the fungus for ten days afforded a hydroxylated metabolite identified as 11beta-hydroxy-alpha-santonin (2) in a high yield (83%), while sclareol (3) was metabolized to epoxysclareol (4) (64%) and a new compound 8-deoxy-14,15-dihydro-15-chloro-14-hydroxy-8,9 dehydrosclareol (5) (7%), representing a rare example of microbial halogenation. PMID- 11098822 TI - Elicitor-induced defence reactions in cell suspension cultures of soybean cultivars. AB - Suspension cultured soybean (Glycine max [L.] Merr.) cells of four cultivars (Wilis, Lumut, Kalmit, Doko RC) were compared for their response to different fungal and bacterial elicitors. Cells were treated either with crude cell wall extracts of the fungal pathogens Phytophthora sojae (Pmg-elicitor) and Rhizoctonia solani (Riso-elicitor) or with two isolates of the bacterial pathogen Pseudomonas syringae pv. glycinea (Psg01/02) and a broad spectrum of antimicrobial defence reactions was measured. Cells of all four cultivars showed the same elicitor-induced rapid (H2O2 accumulation, alkalinization of the culture medium, peroxidative cross-linking of cell wall proteins) and slow (activation of phenylpropanoid metabolism, accumulation of phenolic compounds, induction of PR proteins) defence responses. However, the reactivity of the cultivars was not identical in terms of time courses and intensities. Furthermore, the ability of the various elicitors to induce defence responses varied markedly. These differences indicate that (1) cells of the same species but of different cultivars are equipped with the same array of perception systems to recognise various stimuli but (2) the sensitivity of these perception systems or later steps in the signal transduction seem to be stimulated to a different extent in the analysed cultivars. PMID- 11098823 TI - Induction of DNA-protein cross-links by platinum compounds. AB - The differences between cis- and trans-diamminedichloroplatinum II (DDP) in forming DNA-protein cross-links in isolated human lymphocytes were investigated. Both cis- and trans-DDP can induce DNA-protein cross-links. We show that cis-DDP forms complexes between DNA and proteins faster than trans-DDP. This results from an increase in the quantity of DNA and platinum together with an increase in drug concentration. Under the same conditions trans-DDP causes a decrease in DNA forming complexes with proteins. After a 12 h incubation of lymphocytes we observe a similar level of DNA in DNA-protein cross-links induced by DDP isomers, but more platinum appears in complexes induced by trans-DDP. The results obtained demonstrate that the antitumor drug - cis-DDP and the clinically ineffective trans-DDP induce links between DNA and proteins in a different manner. We suggest that the therapeutic activity of cis-DDP can in part arise from rapidly forming DNA-protein complexes which can destroy the most important cellular processes, such as replication and transcription. PMID- 11098824 TI - The influence of formamide on thermal denaturation profiles of DNA and metaphase chromosomes in suspension. AB - Systematic photometric studies are presented to analyze the thermal denaturation behaviour with and without formamide of metaphase chromosome suspensions in comparison to DNA solutions. Temperature dependent hyperchromicity measurements at 256 nm and 313 nm were performed using an appropriately designed computer controlled photometer device. Due to an upright optical axis, this allowed absorbance measurements with negligible sedimentation effects not only for solutions of pure DNA, but also for particle suspensions of isolated metaphase chromosomes. This device has a temperature resolution of +/- 0.5 degrees C and an optical sensitivity of 10(-3) to 10(-4) optical density. For calf thymus DNA the reduction of the melting point with the increase of formamide in the solution was measured at pH 7.0 and pH 3.2. The good correlation of the theoretical approximation to experimental data indicated the suitability of the apparatus to quantitatively describe DNA conformation changes induced by thermal denaturation. For metaphase chromosome preparations of Chinese hamster culture cells, absorbance changes were measured between 20 degrees C and 95 degrees C with a temperature gradient of 1 degrees C/min. These measurements were performed at pH 7.0 and at pH 3.2. The denaturation profiles (= first derivative of the absorbance curve) resulted in a highly variable peak pattern at 256 nm and 313 nm indicating complex conformation changes. A statistical evaluation of the temperature values of the peak maxima resulted in temperature ranges typical for chromosomal conformation changes during thermal treatment. Especially the range of highest temperature values was independent from pH modifications. For pH 3.2 the influence of formamide on the denaturation behaviour of metaphase chromosome preparations was analyzed. In contrast to pure DNA solutions, a reduction of the "melting point" (i.e. the maximum temperature at which a conformation change takes place) was not found. However, the denaturation behaviour depended on the duration of formamide treatment before the measurement. PMID- 11098825 TI - Complexation of membrane-bound enzyme systems. AB - The effect of changes in the N-terminal membrane-binding domain of cytochrome P450 forms and NADPH-cytochrome P450 reductase types on the cytochrome P450 dependent monooxygenase activities, has been examined. The nifedipine oxidase activity of two human P450 forms (CYP3A4, CYP3A4NF14) which differ only in their primary structure by ten amino acid residues in the N-terminal membrane-binding domain, yields nearly the same catalytic cycle time tau =2.65 +/- 0.15 s, due to their identical cytosolic catalytic protein structure. In contrast, the complex formation process ([P450]+[reductase] <--> [complex]) described by the dissociation constant KD, at high substrate concentration ([S]>>KS) and low product concentration ([P]< approximately 1 mT) on chemical reactions involving free radicals, in the context of possible effects of environmental electromagnetic radiation on biological systems. MATERIALS AND METHODS: Transient absorption, flash photolysis experiments have been performed to study the kinetics and yields of radical reactions. The triplet state of benzophenone has been used as a convenient source of radical pairs, whose identity is largely immaterial to the investigation of the so-called Low Field Effect. Hydrogen abstraction from surfactant molecules in micelles yields a pair of neutral radicals, one large and one small, in a region of restricted translational and rotational motion. RESULTS: In alkyl sulphate and sulphonate micelles a weak field increases the concentration of free radicals that escape from the micelle to an extent that depends on the structure, dynamics and volume of the space in which the radical pairs are confined. The effect (up to 10%) is typically largest at 1-2 mrT. Smaller effects are found for Brij and TX100 micelles. CONCLUSIONS: Low Field Effects depend strongly on the local environment of the radical pair. Larger effects than observed here might be expected for radicals formed from singlet (rather than triplet) precursors, as would be the case in biological reactions. PMID- 11098855 TI - Oxidation of cellular thiols by hydroxyethyldisulphide inhibits DNA double-strand break rejoining in G6PD deficient mammalian cells. AB - PURPOSE: We investigated the effect of protein- and non protein-thiol oxidation on DNA double-strand-break (DSB) rejoining after irradiation and its relevance in the survival of CHO cells. MATERIALS AND METHODS: We used mutant cells null for glucose 6 phosphate dehydrogenase (G6PD) activity since reducing equivalents, required for reduction of oxidized thiols, are typically generated through G6PD regulated production of NADPH. Cellular thiols were oxidized by pre-incubating the cells with hydroxyethyldisulphide (HEDS), the oxidized form of mercaptoethanol (ME). The concentrations of the intracellular and extracellular non-protein thiols (NPSH), glutathione, cysteine and mercaptoethanol were quantitated by HPLC. Protein thiols (PSH) were estimated using Ellman's reagent. Cell survival was determined by clonogenic assay. The induction and rejoining of DSB in cells was quantitated by Pulse Field Gel Electrophoresis after exposure to ionizing radiation. RESULTS: Much lower bioreduction of HEDS was found in the G6PD deficient mutants (E89) than in the wild-type cells (K1). A 1 h treatment of E89 cells with HEDS produced almost complete depletion of non-protein thiol (NPSH) and a 26% decrease in protein thiols. Only minor changes were found under similar conditions with K1 cells. When exposed to gamma radiation in the presence of HEDS, the G6PD null mutants exhibited a higher cell killing and decreased rate and extent of rejoining of DSB than were observed in K1 cells. Moreover, when the G6PD deficient cells were transfected with the gene encoding wild-type G6PD (A1A), they recovered close to wild-type cellular thiol status, cell survival and DSB rejoining. CONCLUSIONS: These results suggest that a functioning oxidative pentose phosphate pathway is required for DSB rejoining in cells exposed to a mild thiol oxidant. PMID- 11098856 TI - Coming of age: 'dysgenetics'--a theory connecting induction of persistent delayed genomic instability with disturbed cellular ageing. AB - INTRODUCTION: In recent years a new phenomenon has manifested itself: delayed, persistent genomic instability. When cells are treated with carcinogens not only direct induction of chromosome aberrations and mutations takes place, but there is also an indirect induction: in the distant progeny of treated cells persistently enhanced levels of new chromosome aberrations and enhanced mutation rates are found. This persistent enhanced genomic instability is not due to the presence of lesions in the DNA induced by the treatment because the response can be transmitted to untreated cells. Apparently it is caused by a persistent dysfunctioning of the cell as a whole. Due to these findings a new model for multistep carcinogenesis emerges. According to this model the initiation of carcinogenesis is the induction of a state of persistent genomic instability that not only is responsible for enhanced mutation rates of oncogenes and tumor suppressor genes, but also predisposes to immortalization. This view could lead to a radical change in our views on carcinogenesis. Therefore understanding the mechanism is of utmost importance. PURPOSE: Up to now, the mechanism responsible for this persistent delayed genomic instability remains completely elusive and has only been described as 'unknown'. In this review the phenomenon is connected with a recent theory on cellular ageing and immortalization. CONCLUSION: Although highly speculative this review provides a framework for further experimental approaches that will contribute to our understanding of delayed genomic instability and possibly even to a better understanding of cellular ageing also. PMID- 11098857 TI - Mathematical methods in biological dosimetry: the 1996 Iranian accident. AB - PURPOSE: To report 18 months of cytogenetic follow-up for an Iranian worker accidentally overexposed to 192Ir, the mathematical extrapolation and comparison with clinical data. MATERIAL AND METHODS: Unstable chromosome aberrations were measured using conventional cytogenetic tests by French and Iranian biological dosimetry laboratories on five occasions after the exposure. The decrease in dicentrics over time was analysed mathematically. In addition, Dolphin and Qdr extrapolations were applied to the data to check the exposure estimates. FISH determination of translocation yields was performed twice by the French laboratory and the results compared with the Dolphin and Qdr corrected values. RESULTS: Dose estimates based on dicentrics decreased from 3.1 +/- 0.4 Gy at 5 days after the accident to 0.8 +/- 0.2 Gy at 529 days. This could be fitted by double-exponential regression with an inflexion point between rapid and slow decrease of dicentrics after about 40 days. Dose estimates of 3.4 +/- 0.4 Gy for the Qdr model and 3.6 +/- 0.5 Gy for the Dolphin model were calculated during the post-exposure period and were remarkably stable. FISH translocation data at 26 and 61 days appeared consistent with the Dolphin and Qdr estimates. CONCLUSION: Dose correction by the Qdr and Dolphin models and translocation scoring appeared consistent with the clinical data and provided better information about the radiation injury than did crude estimates from dicentric scoring alone. Estimation by the Dolphin model of the irradiated fraction of the body seemed unreliable: it correlated better with the fraction of originally irradiated lymphocytes. PMID- 11098858 TI - Radiation effects in spheroids of cells exposed to alpha emitters. AB - PURPOSE: To simulate spheroids of cells and use the spheroid models to establish methods of calculating cell survival following exposure to alpha irradiation from decays of 211At and 213Bi. METHODS: Using a Monte Carlo technique, two models of spheroids of cells were generated in which cells either did not or were allowed to overlap. At 40% packing of the volume it is shown that the two models are equivalent for longer-ranged alpha particles. The overlapping model was used to examine the effects of spheroid size and cell packing on cell survival. Cell survival was also calculated for different distributions of alpha decays such as uniform distribution within the spheroid, external to the spheroid and as a shell on the periphery of the spheroid. RESULTS: Three examples of the cell structure of the spheroids are shown. Detailed calculations show that cell survival decreases from 57% to 37% as the spheroid diameter increases from 75 microm to 225 microm for an average of one 211At decay per cell and 50% packing. Increasing the packing of the cells in the spheroid from 40% to 70% reduces survival from 46% to 26% for 200 microm diameter spheroids for one 211At or 213Bi decay per cell. The presence of small regions of unlabelled cells within the spheroids does not significantly change cell survival. CONCLUSIONS: Monte Carlo programmes generating spheroids of cells and their subsequent use to score cell survival for alpha irradiation should be useful in the design and interpretation of work on spheroids of cells in vitro and the application of such modelling to the study of very small tumours in vivo. Copies of the programmes are available from the author on request. PMID- 11098859 TI - Influence of acetylsalicylic acid on development of radiation-induced nephropathy. AB - PURPOSE: Previous studies have demonstrated that long-term treatment with acetylsalicylic acid (ASA) can significantly reduce the renal functional impairment that develops after high doses of irradiation. The effect is hypothesized to be mediated by selective inhibition of thromboxane A2 synthesis and inhibition of platelet aggregation. The present study was undertaken to investigate this phenomenon further using more clinically relevant fractionated and re-irradiation schedules. METHODS AND MATERIALS: Groups of mice were given bilateral renal irradiation with a series of four or 20 daily fractions of X rays, or 10 daily fractions with a single dose of re-irradiation (0-10 Gy) after 27 weeks. Half the mice received ASA in drinking water (2.4 g x l(-1)) from 1 week before the start of irradiation and continuously throughout the follow-up period. Renal function was assessed by clearance of [51Cr]EDTA, about every 4 weeks for up to 80 weeks after the start of treatment. Histological damage in representative groups of mice was also assessed. RESULTS: Oral administration of ASA caused inhibition of thromboxane A2 synthesis (to < 36% of controls) and a strong inhibition of platelet aggregation in whole mouse blood (ex vivo). Prolonged treatment with ASA also resulted in a small, non-significant reduction of radiation-induced renal functional damage. No reduction in histological damage was seen in the ASA treated mice. CONCLUSION: Long-term oral administration of ASA gave only a modest, non-significant reduction of renal radiation injury after clinically relevant fractionated irradiation schedules. PMID- 11098860 TI - State-specific prevalence of current cigarette smoking among adults and the proportion of adults who work in a smoke-free environment--United States, 1999. AB - Tobacco use in the United States causes approximately 430,000 deaths each year, including an estimated 3000 deaths from lung cancer among nonsmokers exposed to environmental tobacco smoke (ETS). In addition, an estimated 62,000 coronary heart disease deaths annually among nonsmokers exposed to ETS. The detrimental health effects of exposure to ETS are well documented and include, in addition to lung cancer and coronary heart disease among adults, low birthweight and sudden infant death syndrome from exposure during and after pregnancy and asthma, bronchitis, and pneumonia in children. This report summarizes the 1999 prevalence of current cigarette smoking among adults by state and the proportion of persons who work indoors and who report that their workplaces have smoke-free policies. The findings indicate that in 1999, adult smoking prevalence differed more than two-fold across states (13.9%-31.5%) and that the proportion of persons who reported that their workplace had an official smoke-free policy ranged from 61.3% 82.1%. As the respondents' level of education increased, they were more likely to report working under a smoke-free policy. PMID- 11098861 TI - Update: outbreak of Rift Valley Fever--Saudi Arabia, August-November 2000. AB - On September 10, 2000, the Ministry of Health (MOH), Kingdom of Saudi Arabia and subsequently, the MOH of Yemen began receiving reports of unexplained hemorrhagic fever in humans and associated animal deaths and abortions from the far western Saudi-Yemeni border region. These cases subsequently were confirmed as Rift Valley fever (RVF), the first such cases on the Arabian peninsula. This report updates the findings of the ongoing investigation conducted by the Saudi Arabian MOH in collaboration with CDC and the National Institute of Virology, South Africa. PMID- 11098862 TI - Progress toward interrupting indigenous measles transmission--Region of the Americas, January 1999-September 2000. AB - In 1994, countries in the Region of the Americas set a goal of interrupting indigenous measles transmission by the end of 2000. From 1990 to 1996, measles cases declined from approximately 250,000 to an all-time low of 2109 confirmed cases. However, a resurgence began in 1997, with 52,284 confirmed cases reported from Brazil and in 1998, with 14,330 confirmed cases reported from 16 (39%) of the 41 countries that report to the Pan American Health Organization (PAHO). This report summarizes the measles control strategies implemented in the region and measles incidence during 1999-2000 and indicates that the region has made important progress towards interrupting indigenous measles transmission and that achieving this goal is within reach. PMID- 11098863 TI - Introduction: Listening to the Voice of the C/S/X: Consumer/Survivor/Expatient. AB - People who are receiving services for professionally diagnosed psychological disabilities often are not consulted about the nature of those services, or their willingness to participate in them. This issue of the journal presents the autobiographical accounts of four such people, followed by commentaries about those accounts by three professional service-givers. This collection emphasizes the need to obtain informed consent for any psychological services that are offered, for ethical, humane, and professional reasons. PMID- 11098864 TI - Agents, not objects: our fights to be. AB - People who have been diagnosed and treated for major mental illness have an insider expertise that can provide invaluable insight into the mysteries of people's often inexplicable movement in and out of madness. The author vividly describes his passages from identity-seeking young adult to mental hospital patient to psychologist and mental health system critic. The harmful, life threatening treatments he experienced are examined as part of our society's propensity to treat people who are different as deviant and relegate them to ineffective and harmful medical interventions. Alternatives to psychiatric hospitalizations are promoted as more positive options for people going through confusing and frightening life-changing experiences. The article concludes with the author's ideas about what is necessary and helpful when working with someone who has been labeled with a major mental illness and what is counterproductive and harmful. PMID- 11098865 TI - Psychology practitioners and schizophrenia: a view from both sides. AB - In recent years some few psychologists, psychiatrists, and other mental health professionals have begun to reveal their own experiences as persons who have been diagnosed with various forms of serious mental illnesses. This article gives a brief background of one such person, a practicing psychologist who was diagnosed early in his adult life with schizophrenia. Despite numerous breakdowns and hospitalizations he was able to establish a career as a practitioner and as an advocate. The article also offers a number of recommendations for professionals in this field based on the author's experiences as both a recipient and a provider of psychological services to persons with schizophrenia and other serious mental illnesses. PMID- 11098866 TI - The long road back. AB - "The Long Road Back" captures the undulating path of the recovery process from sexual abuse and bipolar disorder in the public mental heath system. It highlights ways the traditional mental health system can help and harm. Even more so. it highlights for the author the importance of involvement in the peer self help movement. Role modeling, hope, and recovery by fellow recipients of services enabled movement toward life-engaging functions such as volunteerism, school and career possibilities. and a commitment to advocacy on behalf of her peers still in the public mental health system. The peer perspective can especially enhance care. Utilizing peer self-help and carefully listening to the voices of those in the system can make mental health services more responsive to our needs. PMID- 11098867 TI - It has to be about choice. AB - My time in the mental health system began a little over 20 years ago, when I was seven. Over the past eight years I have been fighting for, demanding that, respect and ethics play a part in psychology and all that falls under it. I realized that for all the reasons that have been given, for all the excuses that have been made, for a long time I acted as little more than an apologist for a system that refused to change. I learned that it is impossible to make any great systemic changes without the change being wanted-no matter how warranted it is. A system may desperately need to be overhauled, gutted out, and rebuilt. However, inside of that system are the people who work for it, who need the consistency that they have become used to. Psychology, to me, is a self-perpetuating institution. It creates the laws and then defines who breaks them. I am no longer an apologist for the mental health system. Candidly, I have not yet learned how to put a pretty bow on all of my views, which are a direct result of my voluntary and involuntary time in the system. both personally and professionally. All I know is that it has to be about choice. PMID- 11098868 TI - Personal accounts of consumer/survivors: insights and implications. AB - The personal accounts of people diagnosed with mental illness have much to teach practitioners. Based on the four articles in this issue, the expanding literature of personal accounts, and the relevant professional literature, some of the salient themes are discussed. Initially, the human context of mental illness is explored, including the diversity among those diagnosed with mental illness and the cataclysmic impact of such a diagnosis on individuals. The recovery process also is examined, including the role of hope, locus of control, coping skills and strategies, and social support and self-help. Finally, the mental health system is considered, including the limitations of the system, patienthood, professional roles and relationships, and psychotherapy. Implications for practitioners are noted. PMID- 11098869 TI - Commentary: listening to the message. AB - The first-person accounts of c/s/x experiences with the mental health system have several common threads that run through them. These are: (1) labels are stigmatizing; (2) pathology-based models are destructive: (3) it is important to value individuality; (4) alternative treatments are valuable: (5) peer support and self-help are important; (6) therapist training must be reconsidered; and (7) services should be recovery-oriented. In order to be as helpful as possible, mental health professionals must enter into creative partnerships with those in the c/s/x movement, and must consider incorporating these issues in their own functioning and in the training of future professionals. PMID- 11098870 TI - Consumers' perspectives on helpful and hindering factors in mental health treatment. AB - We summarize the four papers in this issue by consumers evaluating their mental health treatment, focusing on aspects of their treatment that they collectively found helpful and hindering. These factors include the context of treatment, the therapy relationship, interventions used and issues addressed, helpful experiences outside the mental health system, and hindering views of mental illness and treatment. We then present comments by two clients in outpatient therapy on the same topic. Research on clients' perceptions of their outpatient psychotherapy is discussed in reference to these six consumer perspectives, and future directions for therapists and researchers are suggested. PMID- 11098871 TI - Intensive behavioral/psychoeducational treatments for autism: research needs and future directions. AB - It is widely acknowledged that, to date, the forms of treatment enjoying the broadest empirical validation for effectiveness with individuals with autism are those treatments based upon a behavioral model and that such treatments are best implemented intensively and early in the child's development. This paper describes several features important in the success of this model and presents remaining issues to be addressed for improving treatment effectiveness. While it is appreciated that there is no "one size fits all" treatment for children with autism, there is as yet no established protocol for relating specific child, family, target behavior, and treatment variables to individualized treatment regimens. Future research needs to include well-conceived and methodologically rigorous investigations allowing for the determination of these important variables. PMID- 11098872 TI - Commentary: the environment as a source of variability: implications for research with individuals who have autism. PMID- 11098873 TI - Interventions to facilitate communication in autism. AB - The purpose of this article is to discuss research opportunities arising from the current literature in the area of communication. Six general themes are discussed, including (a) increasing spontaneity, initiations, and the variety of functions of language verbal and nonverbal children with autism exhibit; (b) assessing and teaching precursors relating to positive outcome; (c) the importance of family involvement in intervention programs; (d) best practices for implementation of communicative interventions; (e) the interrelationship between language and other behavioral symptoms of autism; and (f) the social and pragmatic use of language. These areas are discussed in terms of improving assessment and intervention practices to produce greater long-term communicative outcomes for individuals with autism. PMID- 11098874 TI - Commentary: achievements and future directions for intervention research in communication and autism spectrum disorders. PMID- 11098875 TI - Interventions that facilitate socialization in children with autism. AB - Social dysfunction is perhaps the most defining and handicapping feature of autism. Improved social functioning has long been considered one of the most important intervention outcomes. A variety of social interventions have been designed, empirically examined, and published in the autism literature. Children with autism have been found to be responsive to a wide variety of interventions aimed at increasing their social engagement with others, both adults and peers. Successful strategies employing peer-mediated approaches and peer tutoring have involved typically developing peers. Furthermore, several studies have demonstrated that social engagement directly affects other important behaviors like language, even when these behaviors are not specifically targeted by the teaching program. Thus, while an area of severe involvement, social behavior is also responsive to intervention. PMID- 11098876 TI - Commentary: interventions to facilitate socialization. PMID- 11098878 TI - Commentary: interventions to facilitate auditory, visual, and motor integration: "show me the data". PMID- 11098877 TI - Interventions to facilitate auditory, visual, and motor integration in autism: a review of the evidence. AB - Evidence is reviewed on the prevalence of sensory and motor abnormalities in autism and the effectiveness of three interventions designed to address such abnormalities--sensory integration therapy, traditional occupational therapy, and auditory integration training. Although sensory processing and motor abnormalities are neither universal nor specific to autism, the prevalence of such abnormalities in autism is relatively high. There is, however, little controlled research on the effectiveness of interventions designed to address these abnormalities. Four objective outcome studies of sensory integration therapy were identified. These were of such small scale that no firm conclusions regarding efficacy could be made. No empirical studies of traditional occupational therapy in autism were found. Five studies of auditory integration training were found. Results of these studies provided no, or at best equivocal, support for the use of auditory integration training in autism. PMID- 11098879 TI - Repetitive thoughts and behavior in pervasive developmental disorders: treatment with serotonin reuptake inhibitors. AB - Repetitive thoughts and behavior are considered integral and core components of autistic disorder. Results from recent studies suggest that the types of repetitive thoughts and behavior of adults with autism and those with obsessive compulsive disorder (OCD) may be different. Serotonin reuptake inhibitors (SRIs), the primary drug treatment for patients with OCD, may reduce the repetitive phenomena of some autistic patients. Two controlled studies of the nonselective SRI clomipramine have shown the drug to be more efficacious than the relatively selective norepinephrine reuptake inhibitor desipramine and placebo in children with autism. One controlled study of the selective SRI fluvoxamine found it to be significantly better than placebo for reducing repetitive phenomena and aggression in adults with autistic disorder. Additional research is needed. PMID- 11098880 TI - Commentary: considerations on the pharmacological treatment of compulsions and stereotypies with serotonin reuptake inhibitors in pervasive developmental disorders. PMID- 11098881 TI - Pharmacological treatment of mood disturbances, aggression, and self-injury in persons with pervasive developmental disorders. AB - Aggression, self-injury, and mood disturbances in persons with autistic disorders, while not uncommon, do not constitute core features of autism. Moreover, these problems can occur for a variety of reasons, which need to be assessed in order to plan appropriate and frequently combined (behavioral pharmacological) treatments. Drugs acting primarily in the dopaminergic, serotonergic, adrenergic, opioidergic, and glutamatergic systems all have been explored in the treatment of aggression and self-injury. While no single drug or class of medication has yet emerged as consistently effective, a number of drugs appear promising. Advances in the assessment of aggressive behaviors, the identification of predictors of drug response, and additional controlled clinical drug trials specifically aimed at these target behaviors are essential in improving the approach to these problematic behaviors in the context of autistic disorder. PMID- 11098882 TI - Commentary: considerations on the characterization and treatment of self injurious behavior. PMID- 11098883 TI - Pharmacotherapy for hyperactivity in children with autism and other pervasive developmental disorders. AB - We reviewed pharmacological treatments used in children with autism and PDD-NOS who present with hyperactive symptoms. Some 41 studies were identified from the following drug categories: antipsychotics (n = 13), serotonin reuptake inhibitors (n = 3), antianxiety drugs (n = 4), psychostimulants (n = 10), alpha adrenergic agonists (n = 2), opiate blockers (n = 7), and other drugs (n = 2). Empirical evidence for significant reductions in hyperactive symptoms was strongest for the antipsychotics, psychostimulants, and naltrexone. Most studies have focused on the reduction of overactivity, and more emphasis needs to be placed on distractibility and attentional variables. A theoretical model was proposed in which participants' attentional performance may be used to predict clinical response to psychostimulants. More carefully controlled and comprehensive studies of hyperactivity are badly needed in these children. PMID- 11098884 TI - Commentary: considerations on the pharmacotherapy of attention deficits and hyperactivity in children with autism and other pervasive developmental disorders. PMID- 11098886 TI - Commentary: potential neurobiologic mechanisms through which metabolic disorders could relate to autism. PMID- 11098885 TI - Metabolic approaches to the treatment of autism spectrum disorders. AB - Although the exact prevalence of metabolic abnormalities in autism spectrum disorders is unknown, several metabolic defects have been associated with autistic symptoms. These include phenylketonuria, histidinemia, adenylosuccinate lyase deficiency, dihydropyrimidine dehydrogenase deficiency, 5'-nucleotidase superactivity, and phosphoribosylpyrophosphate synthetase deficiency. When the metabolic consequences of an enzyme defect are well defined (e.g., phenylketonuria, 5'-nucleotidase superactivity), treatment with diet, drugs, or nutritional supplements may bring about a dramatic reduction in autistic symptoms. This review evaluates evidence for metabolic etiologies in autism spectrum disorders, as well as for the efficacy of dietary and vitamin treatments. The relationship between gastrointestinal abnormalities and autism spectrum disorders is also considered. PMID- 11098887 TI - Immunological treatments for autism. AB - Several investigators, including ourselves, have reported significant changes in various immune responses in children with autism. These changes demonstrate dysregulation of the immune system (deficiency in some components of the immune system and excesses in others). In addition, certain genes in the major histocompatibility complex (that regulates immune responses) appear to be involved in autism. Based upon immunological abnormalities, various treatment modalities have been applied to children with autism. In this brief review, these immunological changes and various biological therapies are analyzed and summarized. PMID- 11098888 TI - Commentary: immunological treatments for autism: in search of reasons for promising approaches. PMID- 11098889 TI - Treatment of seizure disorders and EEG abnormalities in children with autism spectrum disorders. AB - The treatment of seizure disorders EEG epileptiform abnormalities without epilepsy in children with autism spectrum disorders (ASD) is considered within the context of the relationship of epilepsy and epileptiform disorders to language, behavior, and cognition. There is an increased prevalence of both epilepsy and abnormal potentially epileptogenic activity in children with ASD. Anecdotal evidence suggests that the use of anticonvulsants to treat epileptiform discharges thought to be producing dysfunction in selected aspects of cognition, language, or behavior makes a positive difference in a subgroup of children with ASD, but there is inadequate evidence on which to base specific recommendations. There is, at present, no scientific justification for considering epilepsy surgery in children with ASD in the absence of intractable clinical seizures. PMID- 11098890 TI - Commentary: the treatment of seizure disorders and EEG abnormalities in children with autistic spectrum disorders: are we getting ahead of ourselves? PMID- 11098891 TI - The basis of hyperspecificity in autism: a preliminary suggestion based on properties of neural nets. AB - This article reviews a few key ideas about the representation of information in neural networks and uses these ideas to address one aspect of autism, namely, the apparent hyperspecificity that is often seen in autistic children's application of previously acquired information. Hyperspecificity is seen as reflecting a possible feature of the neural codes used to represent concepts in the autistic brain. PMID- 11098892 TI - Commentary: a neural systems perspective for improving behavioral treatments for autism. PMID- 11098893 TI - High-resolution image reconstruction method for time-of-flight positron emission tomography. AB - We present a new image reconstruction method for time-of-flight positron emission tomography (TOF-PET). The TOF-PET measurement system is modelled using the continuous-discrete mapping model, and images are reconstructed using an algebraic technique. The proposed method can produce images with better spatial resolution than conventional methods based on the filtered backprojection method. Numerical simulation results show that accurate modelling of the measurement system improves the spatial resolution and the contrast recovery, while the utilization of TOF information improves the signal-to-noise ratios of images. PMID- 11098894 TI - EPR imaging from projections: errors due to misalignment of projection centres and their rectification by a novel acquisition modality. AB - Continuous wave and pulsed wave electron paramagnetic resonance imaging (EPRI) makes use of classical methods of acquisition of projections. Acquisition/reconstruction techniques, such as spin-echo, gradient-echo, etc, cannot be applied to EPRI because they would require very short switching times for the gradient coils. Due to the use of the polar acquisition technique, it is necessary to define a centre of rotation about which the measured projections are rotated during the reconstruction process. This centre represents the point at which the field gradient coils must produce zero magnetic field. Due to the presence of a magnetic field control system that serves to compensate for field variations, principally due to heating, some interference can occur in the control system between the main magnetic field and the magnetic field produced by the gradient coils. The effect changes as the orientation changes. This results in a shift of the centres of the projections as a function of the variation of magnetic field produced by the gradient coils on the control Hall probe. If this condition is present, some artefacts can appear on the reconstructed image. This effect is irrelevant when EPR is used for imaging of paramagnetic probes whose linewidths are of the order of 10(-4) T, while it can be significant in the case of linewidths of the order of 10(-5) T or lower or when EPR is used in microimaging applications (i.e. for high values of magnetic field gradient). We describe the effects that misalignments of the projections have on the reconstructed images. We present a useful method for estimating the real position of the centre and correcting the measured projections before the application of the reconstruction algorithm. Moreover, we demonstrate the functioning of our technique by presenting some examples of EPR reconstruction collected by an X band EPR imaging apparatus. PMID- 11098896 TI - Backscatter towards the monitor ion chamber in high-energy photon and electron beams: charge integration versus Monte Carlo simulation. AB - In some linear accelerators, the charge collected by the monitor ion chamber is partly caused by backscattered particles from accelerator components downstream from the chamber. This influences the output of the accelerator and also has to be taken into account when output factors are derived from Monte Carlo simulations. In this work, the contribution of backscattered particles to the monitor ion chamber response of a Varian 2100C linac was determined for photon beams (6, 10 MV) and for electron beams (6, 12, 20 MeV). The experimental procedure consisted of charge integration from the target in a photon beam or from the monitor ion chamber in electron beams. The Monte Carlo code EGS4/BEAM was used to study the contribution of backscattered particles to the dose deposited in the monitor ion chamber. Both measurements and simulations showed a linear increase in backscatter fraction with decreasing field size for photon and electron beams. For 6 MV and 10 MV photon beams, a 2-3% increase in backscatter was obtained for a 0.5 x 0.5 cm2 field compared to a 40 x 40 cm2 field. The results for the 6 MV beam were slightly higher than for the 10 MV beam. For electron beams (6, 12, 20 MeV), an increase of similar magnitude was obtained from measurements and simulations for 6 MeV electrons. For higher energy electron beams a smaller increase in backscatter fraction was found. The problem is of less importance for electron beams since large variations of field size for a single electron energy usually do not occur. PMID- 11098895 TI - Brain perfusion monitoring with frequency-domain and continuous-wave near infrared spectroscopy: a cross-correlation study in newborn piglets. AB - The newborn piglet brain model was used to correlate continuous-wave (CW) and frequency-domain (FD) near-infrared spectroscopy. Six ventilated and instrumented newborn piglets were subjected to a series of manipulations in blood oxygenation with the effects on brain perfusion known to be associated with brain hypoxia ischaemia. An excellent agreement between the CW and FD was demonstrated. This agreement improved when the scattering properties (determined by the FD device) were employed to calculate the differential pathlength factor, an important step in CW data processing. PMID- 11098897 TI - A Monte Carlo track structure code for electrons (approximately 10 eV-10 keV) and protons (approximately 0.3-10 MeV) in water: partitioning of energy and collision events. AB - An event-by-event Monte Carlo simulation code for track structure studies is described. In the present form the code transports protons (approximately 0.3-10 MeV) and electrons (approximately 10 eV-10 keV) in a water medium in the gas phase approximation. For the type of particles and energy range considered, ionization, electronic excitation and electron elastic scattering are the most important collision events accounted for in the transport simulation. Efforts were made to ensure that the analytic representation of the various interaction cross sections rests on well established experimental data and theory. For example, the secondary-electron spectrum as well as partial and total ionization cross sections are represented by a semitheoretical formulation combining Bethe's asymptotic expansion and binary-encounter theory. Binding effects for five levels of ionization and eight levels of electronic excitation of the water molecule are explicitly considered. The validity of the model cross sections is examined against available experimental data and theoretical predictions from other similar studies. Results pertaining to the partitioning of energy loss and interaction events for the first-collision probability and nanometre-size track segments are presented. PMID- 11098898 TI - Comparison of dosimetry recommendations for clinical proton beams. AB - The formalism and data in the two most recent dosimetry recommendations for clinical proton beams, ICRU Report 59 and the forthcoming IAEA Code of Practice, are compared. Chamber calibrations in terms of air kerma and absorbed dose to water are considered, including five different cylindrical ionization chamber types commonly used in proton beam dosimetry. The methodology for both types of calibration for ionization chambers is described in ICRU Report 59. The procedure based on air kerma calibrations is compared with an alternative formalism based on IAEA Codes of Practice (TRS-277, TRS-381), modified for proton beams. The new IAEA Code of Practice is exclusively based on calibrations in terms of absorbed dose to water and a direct comparison with ICRU Report 59 recommendations is made. Common to the two formalisms are the fundamental quantities Wair and w(air) and their atmospheric conditions of applicability. The difference in the recommended values of the ratio w(air)/Wair (protons to 60Co) is as large as 2.3%. The use of Wair and w(air) values for dry air (IAEA) and for ambient air (ICRU) is a contribution to the discrepancy, and the ICRU usage is questioned. For air kerma based chamber calibrations, ICRU Report 59 does not take into account the effect of different compositions of the build-up cap and chamber wall on the calibration beam quality. For the chamber types included in the study, this introduces discrepancies of up to 1.1%. Combined with differences in the recommended basic data, discrepancies in absorbed dose determination in proton beams of up to 2.1% are found. For the absorbed dose to water based formalism, differences in the formalism, notably the omission of perturbation factors for 60Co in ICRU 59, and data yield discrepancies in calculated kQ factors, and in absorbed dose determinations, between -1.5% and +2.6%, depending on the chamber type and the proton beam quality. PMID- 11098899 TI - A modified polymer gel for radiotherapy dosimetry: assessment by MRI and MRS. AB - The characteristics of a new formulation of polymer gel are assessed for MRI based radiotherapy dosimetry. The gel, based on the first BANG gel formulation, replaces acrylamide with the less toxic monomer sodium methacrylate. The relationship between MR T2 relaxation time and radiation dose for the gel formulation was studied using spin-echo imaging. Proton magnetic resonance spectroscopy was also used to assess the gel composition as a function of dose. The effect of gel pH on the dose-response and baseline R2 was then investigated. A calibration performed on gel without pH modulation (pH = 6.6) revealed a dose response of 0.14 s(-1) Gy(-1) within the range 0-8 Gy. The baseline R2 increases with pH above neutrality, rising from 1.2 s(-1) at pH = 5.1 to 5.0 s(-1) at pH = 10.1. The dose-response is also pH dependent, having a minimum value of 0.09 s( 1) Gy(-1) at pH = 10.1 and peaking at 0.21 s(-1) Gy(-1) at pH = 7.7. Undertaking proton spectroscopy on the gels enabled resonances associated with the monomer and co-monomer to be studied. By integrating the peaks from the respective monomers and normalizing to the signal at 0 Gy it was shown that only 50% of the methacrylate monomer was used at 10 Gy, whereas 80% of the co-monomer was used at this dose. The data indicate that this gel has a reduced toxicity and a comparable dose response to the previously reported BANG gel. In addition, the performance of the gel can be optimized by controlling the pH. MR spectroscopy revealed that the crosslinking co-monomer is consumed more readily than the monomer, which is in agreement with previous compositional studies. PMID- 11098900 TI - Additional factors for the estimation of mean glandular breast dose using the UK mammography dosimetry protocol. AB - The UK and European protocols for mammographic dosimetry use conversion factors that relate incident air kerma to the mean glandular dose (MGD) within the breast. The conversion factors currently used were obtained by computer simulation of a model breast with a composition of 50% adipose and 50% glandular tissues by weight (50% glandularity). Relative conversion factors have been calculated which allow the extension of the protocols to breasts of varying glandularity and for a wider range of mammographic x-ray spectra. The data have also been extended to breasts of a compressed thickness of 11 cm. To facilitate the calculation of MGD in patient surveys, typical breast glandularities are tabulated for women in the age ranges 40-49 and 50-64 years, and for breasts in the thickness range 2-11 cm. In addition, tables of equivalent thickness of polymethyl methacrylate have been provided to allow the simulation for dosimetric purposes of typical breasts of various thicknesses. PMID- 11098901 TI - Estimation of mean glandular dose for mammography of augmented breasts. AB - The standard quantity used to relate breast surface exposure to radiation risk is the mean dose received by the radiation sensitive tissue contained within the female breast, the mean glandular dose (MGD). At present, little is known about the MGD received by women with breast implants as there is no technique available to facilitate its calculation. The present work has involved modification of the conventional method for MGD estimation to make it applicable to women with augmented breasts. The technique was used to calculate MGDs for a cohort of 80 women with breast implants, which were compared with similar data calculated for a total of 1258 non-augmented women. Little difference was found in median MGD at low compressed breast thickness. At high breast thickness, however, the MGDs received by women with augmented breasts were found to be considerably lower than those relating to their non-augmented counterparts. PMID- 11098902 TI - A comparison of normalization effects on three whole-body cylindrical 3D PET systems. AB - Normalization coefficients in three-dimensional positron emission tomography (3D PET) are affected by parameters such as camera geometry and the design and arrangement of the block detectors. In this work, normalization components for three whole-body 3D-capable tomographs (the GE Advance, the Siemens/CTI962/HR+ and the Siemens/CTI951R) are compared by means of a series of scans using uniform cylindrical and rotating line sources. Where applicable, the manufacturers' normalization methods are validated, and it is shown that these methods can be improved upon by using previously published normalization protocols. Those architectural differences between the three tomographs that affect normalization are discussed with a view to drawing more general conclusions about the effect of machine architecture on normalization. The data presented suggest that uniformity of system response becomes easier to achieve as the uniformity of crystal response within the detector block is improved. PMID- 11098903 TI - Simultaneous reconstruction of internal tissue region boundaries and coefficients in optical diffusion tomography. AB - In this paper we propose a new numerical method to the inverse problem in optical diffusion tomography. We consider the reconstruction of the diffusion and absorption coefficients (kappa, mu(a)) within a domain omega which is known to consist of a set of disjoint regions of distinct tissue types. The assumption is that the regions of different tissues are bounded by smooth boundary curves and have constant absorption and diffusion coefficients. The goal in the proposed method is to reconstruct simultaneously the boundaries of the tissue regions together with the absorption and diffusion coefficients within these regions. The solution of the problem is based on the finite element method and subdivision of the elements. The performance of the proposed method is evaluated by simulations in which the optical parameters (kappa, mu(a)) are relevant in medical applications of optical tomography. It is shown that the proposed method is able to recover both the boundaries and the coefficients with good accuracy. PMID- 11098904 TI - Dielectrospectroscopic monitoring of early embryogenesis in single frog embryos. AB - Dielectric spectroscopy has been used to monitor the early embryogenesis of frog (Xenopus laevis) eggs. The dielectric spectra of a single egg in suspension over the frequency range 10 Hz to 10 MHz were collected at various stages of its development. The uncleaved egg showed a dielectric dispersion with a narrow distribution of relaxation times. After the first cleavage, the dielectric spectra were mainly composed of two subdispersions. In the cleavage process, up to the morula stage, changes in the spectra were quantitatively simulated by the 'cell-aggregate' model in which the embryo is regarded as a concentrated suspension of shell-spheres that correspond to the blastomeres (i.e. the cells within the embryo). In the stages from the morula to the blastula, the changes in the dielectric spectra were explained as due to a reduction in the size of the blastomere accompanied by an expansion of the blastocoel (i.e. the central cavity in the embryo) using the 'vesicle-inclusion' model that is a cell aggregate covered with a less conducting shell corresponding to the outermost layer of tightly interconnected cells. PMID- 11098905 TI - Treatment planning for heavy-ion radiotherapy: physical beam model and dose optimization. AB - We describe a novel code system, TRiP, dedicated to the planning of radiotherapy with energetic ions, in particular 12C. The software is designed to cooperate with three-dimensional active dose shaping devices like the GSI raster scan system. This unique beam delivery system allows us to select any combination from a list of 253 individual beam energies, 7 different beam spot sizes and 15 intensity levels. The software includes a beam model adapted to and verified for carbon ions. Inverse planning techniques are implemented in order to obtain a uniform target dose distribution from clinical input data, i.e. CT images and patient contours. This implies the automatic generation of intensity modulated fields of heavy ions with as many as 40000 raster points, where each point corresponds to a specific beam position, energy and particle fluence. This set of data is directly passed to the beam delivery and control system. The treatment planning code has been in clinical use since the start of the GSI pilot project in December 1997. Forty-eight patients have been successfully planned and treated. PMID- 11098906 TI - Treatment planning for heavy-ion radiotherapy: calculation and optimization of biologically effective dose. AB - We describe a novel approach to treatment planning for heavy-ion radiotherapy based on the local effect model (LEM) which allows us to calculate the biologically effective dose not only for the target region but also for the entire irradiation volume. LEM is ideally suited for use as an integral part of treatment planning code systems for active dose shaping devices like the GSI raster scan system. Thus it has been incorporated into our standard treatment planning system for ion therapy (TRiP). Single intensity modulated fields can be optimized with respect to a homogeneous biologically effective dose. The relative biological effectiveness (RBE) is calculated separately for each voxel of the patient CT. Our radiobiologically oriented code system has been used since 1995 for the planning of irradiation experiments with cell cultures and animals such as rats and minipigs. It has been in regular and successful use for patient treatment planning since 1997. PMID- 11098907 TI - The width of margins in radiotherapy treatment plans. AB - Publication of ICRU Reports 50 and 62 has highlighted the need to devise protocols for the process of drawing the planning target volume (PTV) around the clinical target volume (CTV). The margin surrounding the CTV should be wide enough to account for all geometric errors so that no part of the CTV accumulates a dose less than, for instance, 95% of that prescribed. One approach to the problem has been to draw a margin around the CTV delineated at the treatment preparation stage which is sufficiently wide that the mean position of the CTV will be encompassed in a specific percentage of cases, for example 90%. This accounts for the systematic errors. A further margin is then drawn to account for random set-up and organ-motion uncertainties during treatment. The width of this second margin has previously been shown to be 1.64(sigma - sigmap). Here sigma, a vector quantity, is the standard deviation which results from convolving the penumbra spread function of standard deviation sigmap with the Gaussian distributions of the daily positional uncertainties of organ motion and set-up error. However, it is shown in this paper that the calculation should take into account the beam configuration of the treatment plan. In a typical coplanar multibeam plan, usually in the transverse plane, any given edge of the target volume is normally defined by a single beam or two parallel and opposed beams. However, because of the presence of the other beams, the effect of the blurring of the edge-defining beam(s) is reduced, which changes the value of the required margin to beta (sigma - sigmap) where, for example, beta can be as low as 1.04 in the transverse plane of a three-beam plan. The width of the required margins is calculated for up to six beams and presented in a table. It is shown that, while the table was derived using an idealized plan of equally weighted plane beams irradiating a spherical target, it is also valid for non-uniform beam weightings, wedged-beam plans, target volumes of general shape and intensity-modulated radiotherapy (IMRT). PMID- 11098908 TI - A new concept of multileaf collimator (the shuttling MLC)--an interpreter for high-efficiency IMRT. AB - This paper proposes a radically new concept for a multileaf collimator for a photon linear accelerator for delivering IMRT with high monitor-unit efficiency. The concept is to consider each M (rows) x N (columns) two-dimensional intensity modulated beam (2D IMB) as a set of N/2 M (rows) x 2 (columns) areas of modulation. Each area is then delivered by a set of M shuttling attenuating elements (called here the shuttling MLC) with a very high monitor-unit efficiency. The elements shuttle between each of the two columns comprising the M x 2 area and the modulation is provided by the variation in dwell time of the elements. The principles of this shuttling multileaf collimator are discussed and examples illustrating its operation are given. The main achievement reported in this paper is the development and robust testing of an interpreter which describes the position-time course of movement of the elements as a function of monitor units. This interpreter fully accounts for leakage transmission through the elements. It completely avoids the across-the-rows tongue-and-groove underdose. A large number of ID and 2D IMBs have been subjected to this interpreter and it is shown that for random patterns of fluence the SMLC is more monitor-unit efficient than the Bortfeld-Boyer technique (the most efficient with a conventional MLC) when the modulation is highly structured. PMID- 11098909 TI - Elimination of field size dependence of enhanced dynamic wedge factors. AB - Enhanced dynamic wedge factors (EDWF) are characterized by a strong field size dependence. In contrast to physical wedge factors, the EDWF decrease as the field size is increased: for 6 MV 60 degrees wedge, the EDWF decreases by 50% when the field size is increased from 4 x 4 cm2 to 20 x 20 cm2. A method that eliminates the field size dependence of EDWF was developed and investigated in this work. In this method, the wedged field shape is determined by a multileaf collimator. The initial position of the moving Y jaw is determined by the field size and the stationary Y jaw is kept fixed at 10 cm for field sizes < or = 20 cm in the wedged direction. For all other fields, the stationary Y jaw setting is determined by the field size. The modified method results in EDWF that are independent of field size, with no change in the wedge dose distribution when compared with the conventional use of EDW. PMID- 11098910 TI - Modified sector-integration method for predicting the output factors of electron beams including extended source to surface distance. AB - A modified sector-integration method is presented that can predict the output factors of irregular shaped electron fields even in the case of extended source to surface distance (SSD). The model takes as input measured output factors for circular inserts of various radii. These circular fields were measured at SSDs of 100, 105 and 110 cm to determine the effective source distance as a function of radius (ESD(r)). For an arbitrary electron field at any SSD, the shape is divided into small sectors, and the contribution calculated from the radius and ESD(r). The calculated output factors were verified by direct measurements of various types of electron fields mainly based on clinical use. The energies modelled were 8, 10 and 12 MeV for applicator sizes of 10 cm x 10 cm and 14 cm x 14 cm (defined at 95 cm). The calculated values agreed with the measured data within 1% for the various rectangular cutouts including extended source to surface distance. We retrospectively modelled 97 patient inserts of irregular shape, and found agreement within 2% of measured values. PMID- 11098911 TI - The feasibility of MRI-guided whole prostate ablation with a linear aperiodic intracavitary ultrasound phased array. AB - Over the past decade, numerous minimally invasive thermal procedures have been investigated to treat benign prostate hyperplasia and prostate cancer. Of these methods, ultrasound has shown considerable promise due to its ability to produce more precise and deeper thermal foci. In this study, a linear, transrectal ultrasound phased array capable of ablating large tissue volumes was fabricated and evaluated. The device was designed to be compatible for use with MRI guidance and thermometry. The intracavitary applicator increases treatable tissue volume by using an ultrasonic motor to provide a mechanical rotation angle of up to 100 degrees to a 62-element 1D ultrasound array. An aperiodic array geometry was used to reduce grating lobes. In addition, a specially designed Kapton interconnect was used to reduce cable crosstalk and hence also improve the acoustic efficiency of the array. MRI-guided in vivo and ex vivo experiments were performed to verify the array's large-volume ablative capabilities. Ex vivo bovine experiments were performed to assess the focusing range of the applicator. The array generated foci in a 3 cm (2 to 5 cm from the array surface along the axis normal to the array) by 5.5 cm (along the long axis of the array) by 6 cm (along the transverse axis of the array at a depth of 4 cm) volume. In vivo rabbit thigh experiments were performed to evaluate the lesion producing capabilities in perfused tissue. The array generated 3 cm x 2 cm x 2 cm lesions with 8 to 12 half-minute sonications equally spaced in the volume. The results indicate that transrectal ultrasound coagulation of the whole prostate is feasible with the developed device. PMID- 11098912 TI - How to apply a discrete vessel model in thermal simulations when only incomplete vessel data are available. AB - For accurate predictions of the temperature distribution during hyperthermia treatment a thermal model should incorporate the individual impact of discrete vessels. In clinical practice not all vessels can be reconstructed individually. This paper investigates five possible strategies to model the thermal impact of these missing vessels. A tissue volume with a detailed, realistic, counter current discrete vasculature is heated and the steady-state temperature distribution is calculated using our Discrete Vasculature (DIVA) thermal model. To mimic incomplete discrete vasculatures the full tree is gradually stripped, that is, the number of discretely described vessels is reduced in four steps until no discrete vessels are left. At each strip level the steady state temperature distribution is calculated for five different strategies to model the missing vessels. The strategies all use a local or global heat sink model in addition to the discrete vasculature. The resulting temperature distributions are compared with the full tree simulation. With increasing strip level the correspondence with the full tree simulation deteriorated for all strategies. An optimal strategy was found to model the missing vessels depending on the available angiographic data. It was also found that simulations with a decreased number of discrete vessels, or no vessels at all, yield temperatures which are too high. Theoretically this can be compensated by increasing the thermal conductivity; finding the optimal value is done empirically. PMID- 11098913 TI - Decreased entropy of symbolic heart rate dynamics during daily activity as a predictor of positive head-up tilt test in patients with alleged neurocardiogenic syncope. AB - Entropy measures of RR interval variability during daily activity over a 24h period were compared in 30 patients with a positive head-up tilt (HUT) test and 30 patients with a negative HUT test who had a history of alleged neurocardiogenic syncope. Two different entropies, approximate entropy (ApEn) and entropy of symbolic dynamics (SymEn), were employed. In patients showing a positive HUT test, the entropies were significantly decreased when compared with the patients with a negative HUT test. In addition, SymEn in the patients with a negative HUT test was significantly lower than in the normal controls. Discriminant analysis using SymEn could correctly identify 89.3% (520/582) of the 1 h RR interval data of the patients with a positive HUT test regardless of the time of day. Baseline entropies of heart rate dynamics during daily activity were found to be significantly lower in patients with alleged neurocardiogenic syncope and a positive HUT test than in those with the same history but with a negative HUT test. The decreased entropy of symbolic heart rate dynamics may be of predictive value of a positive HUT test in patients with alleged neurocardiogenic syncope. PMID- 11098914 TI - Direct recovery of regional tracer kinetics from temporally inconsistent dynamic ECT projections using dimension-reduced time-activity basis. AB - We present an algorithm of reduced computational cost which is able to estimate kinetic model parameters directly from dynamic ECT sinograms made up of temporally inconsistent projections. The algorithm exploits the extreme degree of parameter redundancy inherent in linear combinations of the exponential functions which represent the modes of first-order compartmental systems. The singular value decomposition is employed to find a small set of orthogonal functions, the linear combinations of which are able to accurately represent all modes within the physiologically anticipated range in a given study. The reduced dimension basis is formed as the convolution of this orthogonal set with a measured input function. The Moore-Penrose pseudoinverse is used to find coefficients of this basis. Algorithm performance is evaluated at realistic count rates using MCAT phantom and clinical 99mTc-teboroxime myocardial study data. Phantom data are modelled as originating from a Poisson process. For estimates recovered from a single slice projection set containing 2.5 x 10(5) total counts, recovered tissue responses compare favourably with those obtained using more computationally intensive methods. The corresponding kinetic parameter estimates (coefficients of the new basis) exhibit negligible bias, while parameter variances are low, falling within 30% of the Cramer-Rao lower bound. PMID- 11098916 TI - Ring filter to locate external boundaries in SPECT using scattered radiation. AB - A novel form of filter for SPECT is described, in which, after back projection and summation, the reconstructed signal is a measure of the total activity within a ring of specified radius, centre and width. The filter is applied to the problem of using Compton scattered radiation to locate external boundaries. In the simple case of the determination of the radius of a circular scattering body of known centre, the filter output would identify a transition region and define an appropriate threshold as the boundary was crossed. However it can also be applied to locate the boundaries seen in individual SPECT projections and hence trace out the envelope of the scattering body. Monte Carlo simulation based on 99mTc is used to test the performance of the filter in a range of situations, with encouraging results. PMID- 11098915 TI - Dual-window scatter correction and energy window setting in cerebral blood flow SPECT: a Monte Carlo study. AB - The image quality in SPECT studies of the regional cerebral blood flow (rCBF) performed with 99mTc-HMPAO is degraded by scattered photons. The finite energy resolution of the gamma camera makes the detection of scattered photons unavoidable, and this is observed in the image as an impaired contrast between grey and white matter structures. In this work, a Monte Carlo simulated SPECT study of a realistic voxel-based brain phantom was used to evaluate the resulting contrast-to-noise ratio for a number of energy window settings, with and without the dual-window scatter correction. Values of the scaling factor k, used to obtain the fraction of scattered photons in the photopeak window, were estimated for each energy window. The use of a narrower, asymmetric, energy discrimination window improved the contrast, with a subsequent increase in statistical noise due to the lower number of counts. The photopeak-window setting giving the best contrast-to-noise ratio was found to be the same whether or not scatter correction was applied. Its value was 17% centred at 142 keV. At the optimum photopeak-window setting, the contrast was improved by using scatter correction, but the contrast-to-noise ratio was made worse. PMID- 11098917 TI - A three-dimensional ray-driven attenuation, scatter and geometric response correction technique for SPECT in inhomogeneous media. AB - The qualitative and quantitative accuracy of SPECT images is degraded by physical factors of attenuation, Compton scatter and spatially varying collimator geometric response. This paper presents a 3D ray-tracing technique for modelling attenuation, scatter and geometric response for SPECT imaging in an inhomogeneous attenuating medium. The model is incorporated into a three-dimensional projector backprojector and used with the maximum-likelihood expectation-maximization algorithm for reconstruction of parallel-beam data. A transmission map is used to define the inhomogeneous attenuating and scattering object being imaged. The attenuation map defines the probability of photon attenuation between the source and the scattering site, the scattering angle at the scattering site and the probability of attenuation of the scattered photon between the scattering site and the detector. The probability of a photon being scattered through a given angle and being detected in the emission energy window is approximated using a Gaussian function. The parameters of this Gaussian function are determined using physical measurements of parallel-beam scatter line spread functions from a non uniformly attenuating phantom. The 3D ray-tracing scatter projector-backprojector produces the scatter and primary components. Then, a 3D ray-tracing projector backprojector is used to model the geometric response of the collimator. From Monte Carlo and physical phantom experiments, it is shown that the best results are obtained by simultaneously correcting attenuation, scatter and geometric response, compared with results obtained with only one or two of the three corrections. It is also shown that a 3D scatter model is more accurate than a 2D model. A transmission map is useful for obtaining measurements of attenuation and scatter in SPECT data, which can be used together with a model of the geometric response of the collimator to obtain corrected images with quantitative and diagnostically accurate information. PMID- 11098918 TI - Development of a miniature scintillation camera using an NaI(Tl) scintillator and PSPMT for scintimammography. AB - We have developed a small scintillation camera dedicated to breast imaging and have evaluated the performance of the system. In order to increase the limited field of view (FOV) determined by the size of a position-sensitive photomultiplier tube (PSPMT), the imaging characteristics of a diverging hole collimator (DHC) were also investigated. The small scintillation camera system consists of an NaI(Tl) crystal (60 mm x 60 mm x 6 mm) coupled to a Hamamatsu R3941 PSPMT, a resistor chain circuit, preamplifiers, nuclear instrument modules, an analogue to digital converter and a PC for control and display. The intrinsic energy resolution of the system was 12.9% FWHM at 140 keV. The spatial resolution was measured using a line-slit mask and 99mTc point sources and was 3.1 mm FWHM. The intrinsic sensitivity of the system was approximately 162 counts/s kBq(-1). The DHC made it possible to image a larger FOV (75 x 75 mm2 at the surface of collimator) than a parallel-hole collimator (60 x 60 mm2). The system sensitivity obtained using the DHC gradually decreased with distance (3% at 1 cm, 6% at 2 cm and 9% at 3 cm). The results demonstrate that the system developed in this study could be utilized clinically to image malignant breast tumours. A DHC can be employed to expand the FOV of the system confined by the size of PSPMT with a modest compromise in the performance of the system. PMID- 11098919 TI - Analytic method based on identification of ellipse parameters for scanner calibration in cone-beam tomography. AB - This paper is about calibration of cone-beam (CB) scanners for both x-ray computed tomography and single-photon emission computed tomography. Scanner calibration refers here to the estimation of a set of parameters which fully describe the geometry of data acquisition. Such parameters are needed for the tomographic reconstruction step. The discussion is limited to the usual case where the cone vertex and planar detector move along a circular path relative to the object. It is also assumed that the detector does not have spatial distortions. We propose a new method which requires a small set of measurements of a simple calibration object consisting of two spherical objects, that can be considered as 'point' objects. This object traces two ellipses on the detector and from the parametric description of these ellipses, the calibration geometry can be determined analytically using explicit formulae. The method is robust and easy to implement. However, it is not fully general as it is assumed that the detector is parallel to the rotation axis of the scanner. Implementation details are given for an experimental x-ray CB scanner. PMID- 11098920 TI - Blood irradiation with accelerator produced electron beams. AB - Blood and blood products are irradiated with gamma rays to reduce the risk of graft versus host disease (GVHD). A simple technique using electron beams produced by a medical linear accelerator has been studied to evaluate irradiation of blood and blood products. Variations in applied doses for a single field 20 MeV electron beam are measured in a phantom study. Doses have been verified with ionization chambers and commercial diode detectors. Results show that the blood product volume can be given a relatively homogeneous dose to within 6% using 20 MeV electrons without the need to rotate the blood bags or the beam entry point. The irradiation process takes approximately 6.5 minutes for 30 Gy applied dose to complete as opposed to 12 minutes for a dual field x-ray field irradiation at our centre. Electron beams can be used to satisfactorily irradiate blood and blood products in a minimal amount of time. PMID- 11098921 TI - Verification of lung dose in an anthropomorphic phantom calculated by the collapsed cone convolution method. AB - Verification of calculated lung dose in an anthropomorphic phantom is performed using two dosimetry media. Dosimetry is complicated by factors such as variations in density at slice interfaces and appropriate position on CT scanning slice to accommodate these factors. Dose in lung for a 6 MV and 10 MV anterior-posterior field was calculated with a collapsed cone convolution method using an ADAC Pinnacle, 3D planning system. Up to 5% variations between doses calculated at the centre and near the edge of the 2 cm phantom slice positioned at the beam central axis were seen, due to the composition of each phantom slice. Validation of dose was performed with LiF thermoluminescent dosimeters (TLDs) and X-Omat V radiographic film. Both dosimetry media produced dose results which agreed closely with calculated results nearest their physical positioning in the phantom. The collapsed cone convolution method accurately calculates dose within inhomogeneous lung regions at 6 MV and 10 MV x-ray energy. PMID- 11098922 TI - Potential application of PET in quality assurance of proton therapy. AB - Our investigation supporting the feasibility of in situ PET monitoring in proton therapy is presented. We simulated by means of the FLUKA code the number and the spatial distribution of the main beta+ emitters created in PMMA targets by protons at typical therapeutic energies. The quantitative comparison with the activation induced by 12C ions of energies corresponding to the same range shows that the available signal at the same physical dose level should be up to twice as intense for protons than that actually successfully used for the control of carbon ion therapy at GSI Darmstadt. The spatial correlation between the activity and the dose profile for protons is poorer than for 12C nuclei. However, an important check of the particle range, dose localization and stability of the treatment during all the fractions seems to be possible. PMID- 11098923 TI - Hydrodynamic effects on the solute transport across endothelial pores and hepatocyte membranes. AB - In this short note we propose a simple and rapid procedure to calculate the net quantity of metabolites absorbed by hepatocytes from blood plasma. The blood movement through sinusoids determines an opposed circulation of plasma through the space of Disse. Hydrodynamic considerations lead to the conclusion that hepatocytes absorb for their own synthesis processes a quantity of metabolites in a volume flow of the order of 10(-12) nl s(-1) through a sieve plate surface with an area of 1 microm2. At pathological temperature (40 degrees C), the excess of the net absorbed volume flow for the entire sinusoidal surface of the mammalian liver may be as high as 1.9 nl s(-1). Some observations on the effect of red and white blood cells on the chylomicron traffic through endothelial pores are made. PMID- 11098924 TI - Comparison of correction techniques for simultaneous 201Tl/99mTc myocardial perfusion SPECT imaging: a dog study. AB - We compared two correction methods for simultaneous 201Tl/99mTc dual-isotope single-photon emission computed tomography (SPECT). Both approaches use the information from the third energy window placed between the photopeak windows of the 201Tl and 99mTc. The first approach, described by Moore et al, corrects only for the contribution of the 99mTc to the 201Tl primary 70 keV window. We developed the three-window transformation dual isotope correction method, which is a simultaneous cross-talk correction. The two correction methods were compared in a simultaneous 201Tl/99mTc sestamibi cardiac dog study. Three separate acquisitions were performed in this dog study: two single-isotope and one dual isotope acquisition. The 201Tl single-isotope images were used as references. The total number of counts, and the contrast between the left ventricular cavity (LVC) and the myocardium, were used in 70 keV short axis slices as parameters for evaluating the results of the dual-isotope correction methods. Three consecutive short-axis slices were used to calculate averaged contrast and the averaged total number of counts. The total number of the counts was 667000+/-500 and 414500+/ 400 counts for the dual isotope (201Tl+/-99mTc) and single-isotope (201Tl-only) 70 keV images, respectively. The corrected dual-isotope images had 514700+/-700 and 368000+/-600 counts for Moore's correction and our approach, respectively. Moore's method improved contrast in the dual isotope 70 keV image to 0.14+/-0.03 from 0.11+/-0.02, which was the value in the 70 keV non-corrected dual-isotope image. Our method improved the same contrast to 0.22+/-0.03. The contrast in the 201Tl single-isotope 70 keV image was 0.28+/-0.02. Both methods improved the 70 keV dual-isotope images. However, our approach provided slightly better images than Moore's correction when compared with 201Tl-only 70 keV images. PMID- 11098925 TI - Response to 'Comparison of dosimetry recommendations for clinical proton beams'. PMID- 11098926 TI - Evaluation and management of ultrafiltration problems in peritoneal dialysis. International Society for Peritoneal Dialysis Ad Hoc Committee on Ultrafiltration Management in Peritoneal Dialysis. PMID- 11098927 TI - Pathophysiology of peritoneal membrane failure. PMID- 11098928 TI - Encapsulating peritoneal sclerosis: definition, etiology, diagnosis, and treatment. International Society for Peritoneal Dialysis Ad Hoc Committee on Ultrafiltration Management in Peritoneal Dialysis. AB - Current definitions of encapsulating peritoneal sclerosis are practical and clinically relevant. It is important to adhere to a more uniform use of the proper terminology, and it is the recommendation of the authors that EPS be adopted as the more appropriate term. The best literal definition of EPS is based on clinical-pathologic criteria. Differentiation of EPS from the general category of ultrafiltration failure is required. Further, better appreciation of the diverse pathways that can lead to the same final common clinical-pathologic picture should not be overshadowed by the requirement of uniform terminology. Incidence and prevalence of the syndrome have been defined in some large populations and a few single-center experiences. The former show an incidence of less than 1%, while higher percentages are reported in the latter. The reported increased incidence with duration on therapy requires validation. The epidemiology of the syndrome offers limited insight into its pathogenesis. A list of factors, both dialysis-related and non dialysis-related. has been accumulated. Except in a few categories where agents are clearly related to the development of EPS, the majority of the listed factors for dialysis-related BPS remain, at best, associations and at worst, simple conjecture. The same limitations that plague the issue of etiology apply in the area of pathogenesis. More basic, focused work is required. The diagnosis of EPS remains based on clinical suspicion confirmed with, primarily, radiologic findings. Pathologic confirmation is obtained in cases that come to surgery for management or for catheter removal. Radiologic studies are precise enough for confirmation, but none have been evaluated for early diagnosis for possible early intervention or prevention. Studies based on transport characteristics or effluent dialysate constituents are not useful for EPS. At present, there are no reliable predictive tests for BPS that can be used in individual patients. Therapy of BPS is based on anecdotal evidence. The possible variable etiologies and probable distinct pathways leading to the syndrome may make a uniform therapeutic approach unlikely. Further, the limited number of cases and the sporadic pattern of occurrences make therapeutic trials not readily feasible. This is distinct from the case of ultrafiltration failure, where significant advances in mechanism elucidation and rationale-based interventions have been made. PMID- 11098929 TI - Diversity is demonstrated in class I HLA-A and HLA-B alleles in Cameroon, Africa: description of HLA-A*03012, *2612, *3006 and HLA-B*1403, *4016, *4703. AB - To examine the genetic diversity in west Africa, class I HLA-A and HLA-B alleles of 92 unrelated individuals from two areas in the Cameroon, the capital Yaounde and the village of Etoa, were identified by direct automated DNA sequencing of exons 2 and 3 of the HLA-B locus alleles and sequence-specific oligonucleotide probe (SSOP) and/or sequencing of the HLA-A locus alleles. HLA-A*2301 (18.7%), A*2902 (10.4%), B*5301 (10.9%), and B*5802 (10.9%) were the most frequently detected alleles, present in at least 10% of the population. A total of 30 HLA-A locus and 33 HLA-B locus alleles, including six novel alleles, were detected. The novel alleles were HLA-A*03012, A*2612, A*3006 and HLA-B*1403, B*4016, and B*4703. HLA-B*4703 contains a novel amino acid sequence that is a combination of the first 5 amino acids of the Bw6 epitope and the last 2 residues of the Bw4 epitope. The addition of 6 alleles to the ever-expanding number of known class I HLA alleles supports our hypothesis that extensive genetic diversity, including previously undescribed alleles, would be observed in this African population. In the Yaounde population, the allele frequency distribution at the HLA-A locus is consistent with distributions indicative of balancing selection. Extensive HLA-A B haplotypes were observed in this population suggesting that only a fraction of the Cameroon HLA-A-B haplotype diversity has been observed. PMID- 11098930 TI - The molecular determination of HLA-Cw alleles in the Mandenka (West Africa) reveals a close genetic relationship between Africans and Europeans. AB - HLA-Cw alleles were determined by high-resolution polymerase chain reaction sequence-specific oligonucleotide probe (PCR-SSOP) oligotyping in a sample of 165 Mandenka, a population from Eastern Senegal previously analysed for A/B and DRB/DQB polymorphisms. A total of 18 Cw alleles were identified, with Cw*0401/5 and 1601 accounting for a combined frequency of 36%. A comparison of Cw allele frequencies among several populations of different origins, Mandenka, Swiss, English, Ashkenazi Jews from the UK and Japanese, reveals a high genetic heterogeneity among them, but also a much closer relationship between Mandenka, Europeans and Ashkenazi than between any of these populations and Japanese. Cw*0501, Cw*0701 and Cw*1601, among others, appear to be restricted to the European and African populations. Many B-Cw haplotypes exhibit a significant linkage disequilibrium in the Mandenka, among which B*3501-Cw*0401 and B*7801 Cw*1601, formed by the most frequent B and Cw alleles, and B*5201-Cw*1601, B*5702 Cw*18 and B*4410-Cw*0401, not yet observed in other populations. B*3501-Cw*0401 is found with similar frequencies in Europeans. The results possibly support a close historical relationship between Africans and Europeans as compared to East Asiatics. However, the HLA-Cw frequency distributions are characterised by an excess of heterozygotes, indicating that balancing selection may have played a role in the evolution of this polymorphism. PMID- 11098931 TI - Development of a PCR-SSOP approach capable of defining the natural killer cell inhibitory receptor (KIR) gene sequence repertoires. AB - A molecular typing method based on polymerase chain reaction (PCR) amplification of three different target domains (immunoglobulin domains 1 and 3, and the transmembrane-cytoplasmic domain), followed by hybridisation with 26 digoxigenin labelled sequence-specific oligonucleotide probes (SSOP) has been established for the polymorphic killer inhibitory receptor (KIR) genes. In addition to identifying the 12 KIR subfamilies, our PCR-SSOP typing approach could also distinguish the putative alleles, NKB1 and NKAT3, that comprise the KIR3DL1 subfamily. Ninety unrelated blood donors and 13 families (52 individuals), including both parents, were subjected to our KIR PCR-SSOP typing approach. All 12 KIR subfamilies, including a 2DS5 variant sequence, were present in the 90 individuals and displayed varied phenotype frequencies: 2DL1 (0.96), 2DL2 (0.31), 2DL3 (0.95), 2DS1 (0.56) 2DS2 (0.51), 2DS3 (0.27), 2DS4 (0.96), 2DS5v (0.35), 3DS1 (0.47), 3DL1 (0.96), 3DL2 (1.0) and 2DL4 (1.0). A total of 23 different KIR phenotypes were defined in this study, and 10 of these were only found on one occasion in one individual, indicating considerable diversity in the KIR phenotype profiles within the Irish population. Most individuals (93%) possessed the complement of inhibitory KIR specificities for the three well-defined HLA-B and -C ligands. An unusual probe pattern for 3DS1 was observed in 3 individuals indicating a variant 3DS1 gene sequence with changes at nucleotide positions 1185 1186, within the cytoplasmic domain. Sequencing analysis revealed a new single nucleotide polymorphism in exon 3 of 3DL1 NKB1(195, G-A) and a 22-bp deletion polymorphism in exon 5 of 2DS4 (nucleotides 777-798 deleted). A number of strong KIR associations were observed, namely 2DL1 with 2DL3, 2DS4 with 3DL1, 2DL2 with 2DS1/2DS2/2DS3, 2DS1 with 2DS3/2DS5v/3DS1, 2DS2 with 2DS3 and 2DS5v with 3DS1. Analysis of the KIR segregation observed in the 13 families confirmed these strong associations and permitted the definition of a number of partial KIR haplotypes, e.g. 2DL2-2DS1-2DS2-2DS3-3DL1. The segregation analysis concluded that at least 3 distinct gene loci encode 2DL1-4 and at least 4 gene loci encode the non-inhibitory KIR2DS1-2DS5. In the case of 3DL1-2 and 3DS1, our data suggests 3 gene loci, one for each subfamily. PMID- 11098932 TI - Coordinate downregulation of multiple MHC class I antigen processing genes in chemical-induced murine tumor cell lines of distinct origin. AB - In murine tumor cell lines, downregulation of MHC class I surface expression has been frequently detected, but the underlying molecular mechanisms of such deficiencies have not been defined. In this study, murine tumor cell lines of different histology derived from spontaneous or from chemical-induced tumors were analyzed for the expression of multiple components of the major histocompatibility complex (MHC) class I antigen-processing machinery (APM), including the peptide transporter TAP, the interferon (IFN)-gamma inducible proteasome subunits and several chaperones. The tumor cell lines analyzed demonstrated a heterogeneous expression pattern of various APM components. In comparison to control cells an impaired coordinated expression of at least three APM components was detected. In particular, extensive APM deficiencies were found in cell lines derived from chemical-induced tumors. A strong coordinated downregulation of expression and/or function of TAP, the low molecular weight proteins (LMP) subunits, the proteasome activator PA28 and/or tapasin was found in 5 of 10 tumor cells, which was associated with impaired MHC class I surface expression. In contrast, the expression of beta2-microglobulin (beta2-m), PA28beta, the constitutive proteasome subunits X, Y, Z and of the chaperones calnexin, calreticulin, ER60 and phospho disulfide isomerase (PDI) was unaltered or only weakly decreased. The deficient expression of APM components could be corrected by IFN-gamma treatment, which also reconstituted MHC class I surface expression. However, impaired expression of APM molecules appears not to be the only cause of abnormal MHC class I expression, since it could neither be corrected by the addition of exogeneous MHC class I binding peptides nor by incubation at low temperature. These results suggest that one major mechanism of murine tumor cells, in particular chemical-induced tumors, to evade the immune system is the combined dysregulation of various APM components and other factors, which still have to be identified. PMID- 11098933 TI - HLA class II alleles in Japanese patients with immune thrombocytopenic purpura. Associations with anti-platelet glycoprotein autoantibodies and responses to splenectomy. AB - HLA class II alleles and immunoglobulin allotypes were determined in 83 Japanese patients with immune thrombocytopenic purpura (ITP) and 114 race-matched healthy controls. Distribution of DRB and DQB1 alleles as well as G1M(f, z), G2M(n+, n-), and KM(1, (1,2), 3) were not different between ITP patients and healthy controls, while DPB1*0201 was marginally increased in ITP patients vs. healthy controls (51% vs. 28%, Pc= 0.04, OR=2.6 [1.4-4.8]). In contrast, strong associations between anti-glycoprotein autoantibodies and HLA class II genes were found as follows: antiGPIIb-IIIa antibody with DRB1*0405 and DQB1*0401; and anti-GPIb-IX antibody with DRB1*0803 and DQB1*0601. When factors influencing therapeutic responses to splenectomy were examined, a poor response was correlated with the presence of DRB1*0405, DQB1*0401 and anti-GPIIb-IIIa antibody (P=0.01, 0.002, and 0.03, respectively). Our results indicate that HLA class II genes influence the production of anti-glycoprotein antibody specificities rather than the development of ITP. In addition, HLA class II genotyping could be useful in predicting therapeutic responses to splenectomy in Japanese patients with ITP. PMID- 11098934 TI - Screening for the IDDM high-risk genotype. A rapid microtitre plate method using serum as source of DNA. AB - Norwegian babies born with the HLA-DRB1*0401-DQA1*03-DQB1*0302/DRB1*03-DQA1+ ++*05-DQB1*0201 genotype have an estimated 17% lifetime risk of developing insulin-dependent diabetes mellitus (IDDM). Identifying these children is important for future prevention, and for studies of the non-genetic factors involved in IDDM. The aim of the study was to develop a rapid screening method for this high-risk genotype. DNA was extracted from serum collected during routine newborn screening for phenylketonuria and hypothyreosis. The second exons of HLA-DQA1 and DQB1 were co-amplified using biotinylated primers, amplicons were hybridized to a set of seven probes immobilized on a microtitre plate using a single hybridisation temperature, and detected colorimetrically by streptavidin HRP conjugate and tetramethylbenzidine substrate. The DRB1*04 subtyping was performed using six different probes at identical conditions. The prevalence of the DRB1*0401-DQA1*03-DQB1*0302/DRB1*03-DQA1*0 5-DQB1*0201 genotype among 1,026 Norwegian babies was 2.7% (CI 95%: 1.7-3.7%). The new high-throughput genetic screening method for IDDM risk can easily be automated and included in newborn screening programs. PMID- 11098935 TI - The CTLA4/CD28 gene region on chromosome 2q33 confers susceptibility to celiac disease in a way possibly distinct from that of type 1 diabetes and other chronic inflammatory disorders. AB - The effect of the gene region on chromosome 2q33 containing the CD28 and the cytotoxic T-lymphocyte associated (CTLA4) genes has been investigated in several diseases with chronic inflammatory nature. In addition to celiac disease (CD), type I diabetes, Grave's disease, rheumatoid arthritis and multiple sclerosis have all demonstrated associations to the A/G single nucleotide polymorphism (SNP) in exon 1, position +49 of the CTLA4 gene. The purpose of this study was to investigate this gene region in a genetically homogeneous population consisting of 107 Swedish and Norwegian families with CD using genetic association and linkage methods. We found a significant association with preferential transmission of the A-allele of the exon 1 +49 polymorphism by using the transmission disequilibrium test (TDT). Suggestive linkage of this region to CD was moreover demonstrated by non-parametric linkage (NPL) analysis giving a NPL score of 2.1. These data strongly indicates that the CTLA4 region is a susceptibility region in CD. Interestingly, of the several chronic inflammatory diseases that exhibit associations to the CTLA4 +49 A/G dimorphism, CD appears to be the only disease associated to the A allele. This suggests that the +49 alleles of the CTLA4 gene are in linkage disequilibrium with two distinct disease predisposing alleles with separate effects. The peculiar association found in the gut disorder CD may possibly relate to the fact that the gastrointestinal immune system, in contrast to the rest of the immune system, aims to establish tolerance to foreign proteins. PMID- 11098936 TI - Sequence-based typing of HLA-B: the B7 cross-reacting group. AB - The large number of polymorphic sites in the HLA-B locus makes sequencing an efficient way of detecting and analysing them. Most polymorphic sites are located in the alpha1 and alpha2 domains of the molecule, encoded by exons 2 and 3 of the gene. An HLA-B-specific sequence-based typing (SBT) strategy was designed for routine application identifying the polymorphic sites in these domains. Exons 2 and 3 were amplified separately using amplification primers located in intron 1, intron 2 and intron 3. Separate amplification of exons 2 and 3 resulted in short polymerase chain reacting (PCR) products and enabled a solid-phase sequencing approach, which made correct assignment of heterozygous positions possible due to low background. A one-step sequencing reaction was performed using fluorescent dye-labelled sequencing primers. One forward sequencing reaction was performed for exon 2, whereas for exon 3, two forward sequencing reactions were needed using two different sequencing primers located in intron 2 and exon 3. The combined sequences of exon 2 and 3 were used for automatic alignment to an HLA-B sequence database and automatic allele assignment. A total of 355 individuals with at least one allele belonging to the B7 cross-reacting group (B7, 13, 22, 27, 40, 41, 42, 47, 48, 81 and 82) were typed for HLA-B by SBT. In the B7 group 48 different alleles were identified, in the non-B7 group a further 59 alleles were sequenced, 9 new alleles were identified. The sequencing strategy described has proven to be reliable and efficient for high-resolution HLA-B typing. PMID- 11098937 TI - Unexpected Bw4 and Bw6 reactivity patterns in new alleles. AB - The Bw4 and Bw6 epitopes were the first HLA-B differences to be recognized by serological methods. Since then 44 serological groups have been identified and more than 250 alleles assigned by molecular typing methods. In general each serological HLA-B group is associated with the presence of either the Bw4 or the Bw6 epitope. There are several exceptions to this rule. Four alleles, B*4601, *7301, *5503 and *1806, show no serological reactivity with either Bw4 or Bw6. Although the Bw6 motif at residues 77-83 is present in these alleles the Bw6 epitope is modified by a valine at residue 76. One or more alleles from the B8, B40 and B62 groups are identified as Bw4 positive, whereas all others are Bw6 positive. In the groups B27, B44 and B47 several alleles are found to be Bw6 positive, while the majority is Bw4 positive. Histocompatibility testing of dialysis patients and their families revealed the serological presence of an unexpected Bw4 epitope associated with B18 in one patient and B56 in another. Allele-specific amplification and sequencing of exons 2 and 3 of these HLA-B alleles revealed the presence of the Bw4 sequence motif for both. The new alleles were assigned B*1809 and B*5607, respectively. In 2 other patients the presence of a new B*07 allele was determined by sequence based typing. Although the new allele, B*0715, showed the Bw6 sequence motif at positions 77 to 83, a substitution of amino acid 76 from glutamic acid to valine was identified. This change resulted in an aberrant Bw6 serological reaction pattern. PMID- 11098938 TI - Increased diversity within the HLA-B*07 group: identification of the two novel alleles B*0709 and B*0710. AB - The identification of the two novel alleles, HLA-B*0709 and B*0710, is described. B*0709 differs from B*07021 by a nucleotide exchange at position 419 (A>C) which is located in exon 3. At the protein level the nucleotide exchange results in an amino acid residue difference (Tyr116Ser). The other newly detected allele, B*0710, differs from B*07021 by a nucleotide exchange at position 272 (A>G) which is located in exon 2. This nucleotide exchange also leads to an amino acid substitution (Tyr67Cys). The allogeneic potential in case of mismatch with other alleles of the B*07 group at bone marrow transplantation was assessed. The peptide motifs between B*0709 and B*0711 may be very similar and thus the alloreactive potential in case of mismatch may be low. In contrast, mismatches of B*0709 and B*0710 with other B*07 alleles are likely to stimulate alloreactive T cells. PMID- 11098939 TI - Characterization of HLA-B*3921 and confirmation of HLA-B*4415, two variant HLA-B alleles identified in the Omani population. AB - We describe a variant HLA-B*39 allele present in two individuals from Oman, which has been officially named HLA-B*3921. In addition we confirm the existence of HLA B*4415, an allele closely related to HLA-B*4501 differing only at the Bw4/Bw6 epitope. PMID- 11098940 TI - Identification by sequencing based typing and complete coding region analysis of three new HLA class II alleles: DRB3*0210, DRB3*0211 and DQB1*0310. AB - The study of HLA class II polymorphism by direct exon 2 DNA sequencing analysis has been established to be a reliable and accurate high-resolution typing procedure. This approach shows some advantages in relation to previous methods, polymerase chain reaction using sequence-specific oligonucleotides (PCR-SSO) and sequence-specific primers (PCR-SSP), basically due to the capability of analysis for the complete sequenced genomic region, including non-polymorphic motifs. DRB3 and DQB1 sequencing based typing (SBT) in unrelated bone marrow donor searching allowed us to detect three new alleles. The complete coding region sequences were characterised from cDNA. Two new DRB3 alleles, DRB3*0210 and DRB3*0211, were described in two Caucasian bone marrow donors. Both sequences showed single point mutations regarding DRB3*0202, producing amino acid replacements at positions 51 (Asp to Thr) and 67 (Leu to Ile), respectively. These two point mutations can be found in other DRB alleles, and suggest that gene conversion would be involved in the origin of both alleles. A new DQB1 sequence was found in a Spanish patient that showed two nucleotide differences, positions 134 and 141, with regard to its close similar DQB1*03011 allele. Only substitution at position 134 provoked amino acid replacement at residue 45, Glu to Gly. This single amino acid change would be involved in the lack of serologic recognition of this new molecule by DQ7 specific reagents. PMID- 11098941 TI - The carotid pulse check revisited: what if there is no pulse? AB - This study was undertaken to evaluate the diagnostic accuracy and time required by first responders to assess the carotid pulse in potentially pulseless patients. We conducted a prospective, randomized study of first responders (n = 206; four different training levels) and were blinded as to the patients' conditions in the cardiac operating rooms of a university hospital. Sixteen patients underwent coronary artery bypass surgery on nonpulsatile cardiopulmonary bypasses. Carotid pulse check was performed either during pulsatile (spontaneous) or during nonpulsatile (extracorporeal) circulation. Patients' hemodynamic status at the time of assessment, diagnostic accuracy of the first responders, and the time required to diagnose carotid pulsatility or pulselessness were documented. Within 10 secs, only 16.5% of the participants (34 of 206) were able to reach any decision about their patients' pulse status. Assessments that were both rapid and correct (15%, i.e., 31 of 206) occurred almost exclusively in pulsatile patients. Advanced training level shortened the delay to decision and improved its accuracy. However, merely 2% of the participants (1 of 59) correctly recognized a truly pulseless patient within 10 secs. Recognition of pulselessness of the carotid artery by rescuers with basic cardiopulmonary resuscitation training is time-consuming and highly inaccurate. Although the carotid pulse check needs to be taught, its importance in the context of layperson basic life support should be de-emphasized. PMID- 11098942 TI - Cardiopulmonary resuscitation without ventilation. AB - Current resuscitation methods, although occasionally effective, rarely perform as well as initially anticipated. Some of the disappointment can be attributed to the difficulty of the task for many, including both professional and lay first responders. Significant attention has been paid recently to the need to simplify both the technique and the teaching of resuscitation. In considering simplification of the current resuscitation scheme, a logical start is an honest reappraisal of the importance and priorities of each of the once sacrosanct ABCs, specifically, establishment of an Airway, artificial Breathing (mouth-to-mouth breathing), and chest compressions for temporary Circulation. Experimental data continue to accumulate indicating that most important within this triad is circulation. Adequate oxygen exists within the blood during at least the first 10 mins of cardiac arrest. If circulation is provided to distribute such oxygen, no survival disadvantage results with chest compression-only basic life support (BLS) efforts. Even a totally occluded airway during the first 6 mins of cardiac arrest does not compromise survival if reasonable circulation is provided with chest compressions. Clinical studies support the same conclusion that what most influences survival in any BLS effort is circulation, not ventilation. Belgium investigators have shown equal survival rates among those treated with chest compressions plus ventilation and those who received chest compressions alone. Telephone dispatcher-guided BLS cardiopulmonary resuscitation (CPR) has likewise shown no survival disadvantage to chest compression-only CPR when compared with telephone-guided standard BLS CPR. Based on this reasoning, a new simplified BLS method has been proposed. "Staged" CPR consists of a strategy to initially teach laypersons a simplified approach to BLS, which requires only chest compressions and not mouth-to-mouth breathing. "Bronze" CPR, in which chest compression-only BLS is taught, was compared with the standard European Resuscitation Council BLS course for laypersons. Manikin "exit testing" at course completion has revealed significant advantages of the simplified approach compared with standard CPR courses for the lay public. PMID- 11098943 TI - Dispatcher-assisted "phone" cardiopulmonary resuscitation by chest compression alone or with mouth-to-mouth ventilation. AB - Based on both animal studies and field studies of the process and intermediate outcomes related to cardiopulmonary resuscitation (CPR), we initiated a randomized trial of dispatcher-assisted CPR, with the intervention arm receiving instructions for chest compression only and the control arm receiving standard instructions for airway maintenance ventilation, and chest compression. Of 241 patents randomized to chest compression instructions only, 35 survived (14.6%) compared with 29 of 279 (10.4%) patients in the control arm (p = .09). These results may have implications for future guidelines and teaching CPR. PMID- 11098944 TI - Role of mouth-to-mouth rescue breathing in bystander cardiopulmonary resuscitation for asphyxial cardiac arrest. AB - There is increasing evidence that mouth-to-mouth rescue breathing may not be necessary during brief periods of bystander cardiopulmonary resuscitation (CPR) for ventricular fibrillation. In contrast to ventricular fibrillation cardiac arrests, it has been assumed that rescue breathing is essential for treatment of asphyxial cardiac arrests because the cardiac arrests result from inadequate ventilation. This review explores the role of mouth-to-mouth rescue breathing during bystander CPR for asphyxial cardiac arrests. Clinical data suggest that survival from apparent asphyxial cardiac arrest can occur after CPR consisting of chest compressions alone, without rescue breathing. Two randomized, controlled swine investigations using models of bystander CPR for asphyxial cardiac arrest establish the following: a) that prompt initiation of bystander CPR is a crucially important intervention; and b) that chest compressions plus mouth-to mouth rescue breathing is markedly superior to either technique alone. One of these studies further demonstrates that early in the asphyxial pulseless arrest process doing something (mouth-to-mouth rescue breathing or chest compressions) is better than doing nothing. PMID- 11098945 TI - Effects of inspired gas content during respiratory arrest and cardiopulmonary resuscitation. AB - Mouth-to-mouth and bag-valve-mask ventilation have been an indispensable part of cardiopulmonary resuscitation (CPR). However, only recently have the effects of different tidal volumes on arterial oxygenation been reported for mouth-to-mouth and bag-valve-mask ventilation. Currently recommended tidal volumes (10-15 mL/kg) are associated with an increased risk of gastric inflation because they produce high peak inspiratory pressures. An animal model of ventilation with an unprotected airway showed that a smaller tidal volume (6 mL/kg) is as effective as a larger tidal volume (12 mL/kg) in maintaining Sao2 at >96%. However, a smaller tidal volume with exhaled gas ventilation produced a mean Sao2 of 48%, which is ineffective. Ventilation gas mixtures have been studied in models of cardiac arrest and CPR. One study showed that ventilation with air during 6 mins of CPR resulted in a return of spontaneous circulation in 10 of 12 animals compared with only 5 of 12 animals ventilated with exhaled gas (p<.04). Arterial and mixed-venous Po2 were significantly higher, and Pco2 was significantly lower in the air ventilation group. Investigations of the cardiovascular effects of mouth-to-mouth ventilation during CPR suggest that there are adverse effects during low blood flow states. However, mouth-to-mouth ventilation during respiratory arrest is lifesaving and should continue to be taught and emphasized in basic life support courses. PMID- 11098946 TI - Efficacy of interposed abdominal compression-cardiopulmonary resuscitation (CPR), active compression and decompression-CPR and Lifestick CPR: basic physiology in a spreadsheet model. AB - This study was undertaken to understand and predict results of experimental cardiopulmonary resuscitation (CPR) techniques involving compression and decompression of either the chest or the abdomen. Simple mathematical models of the adult human circulation were used. Assumptions of the models are limited to normal human anatomy and physiology, the definition of compliance (volume change/pressure change), and Ohm's law (flow = pressure/resistance). Interposed abdominal compression-CPR, active compression and decompression of the chest, and Lifestick CPR, which combines interposed abdominal compression and active compression and decompression, produce, respectively, 1.9-, 1.2-, and 2.4-fold greater blood flow than standard CPR and systemic perfusion pressures of 45, 30, and 58 mm Hg, respectively. These positive effects are explained by improved pump priming and are consequences of fundamental principles of cardiovascular physiology. PMID- 11098947 TI - Cardiopulmonary resuscitation with a hydraulic-pneumatic band. AB - Improved blood flow during cardiopulmonary resuscitation (CPR) has been shown to enhance survival from cardiac arrest. Chest compression with a circumferential pneumatic vest enhances blood flow, but the size, weight, and energy consumption of the inflation system limit its portability and, thereby, have made clinical studies difficult. The purpose of this investigation was to study an improved circumferential chest compression device that uses a constricting band that is pneumatically actuated. The constricting band applies its force to a hydraulic cushion that contacts the anterior and lateral aspects of the chest. The hydraulic cushion transfers the circumferential constriction to inward force. CPR was performed on subjects 5 mins after induction of ventricular fibrillation, with the hydraulic-pneumatic band system (HB-CPR), with a pneumatic vest system (PV-CPR), and with standard manual CPR (S-CPR), each done for 2 mins in randomized order. Aortic and right atrial pressures were measured with micromanometers. Coronary perfusion pressure was calculated as the mean difference between the aortic and right atrial pressures during the release phase of chest compression. Aortic pressure and coronary perfusion pressure with HB-CPR and PV-CPR were improved over S-CPR, and HB-CPR produced comparable pressures to those of PV-CPR. The system for performing HB-CPR, however, was substantially lighter (10 vs. 50 kg) and consumed less energy (300 vs. 1000 watts) than that for PV-CPR. Thus, HB-CPR appears to produce a similar improvement in hemodynamics over S-CPR as PV-CPR but may be more portable than PV-CPR. Therefore, HB-CPR may allow larger scale testing of circumferential chest compression approaches. PMID- 11098948 TI - Improving the efficiency of cardiopulmonary resuscitation with an inspiratory impedance threshold valve. AB - In an effort to improve the efficiency of cardiopulmonary resuscitation (CPR), a new inspiratory impedance threshold valve has been developed to enhance the return of blood to the thorax during the chest decompression phase. This new device enhances negative intrathoracic pressure during chest wall recoil or the decompression phase, leading to improved vital organ perfusion during both standard CPR and active compression-decompression CPR. With active compression decompression CPR, addition of the impedance threshold valve results in sustained diastolic pressures of >55 mm Hg in patients in cardiac arrest. The new valve shows promise for patients in asystole or shock refractory ventricular fibrillation, when enhanced return of blood flow to the chest is needed to "prime the pump." The potential long-term benefits of this new valve remain under study. PMID- 11098949 TI - Electrocardiographic waveform analysis for predicting the success of defibrillation. AB - A new electrocardiographic predictor of the likelihood that an electrical shock would restore a perfusing rhythm is described. The intent was to develop a prognosticator that would be displayed during precordial compression. We anticipated that such a predictor would allow more selective timing of electrical shocks and reduce electrical injury to the myocardium caused by repetitive shocks. In a porcine model of cardiac arrest because of ventricular fibrillation, electrocardiographic recordings of ventricular fibrillation wavelets were analyzed and transformed into an amplitude spectrum area (AMSA). An AMSA value of 21 mV x Hz predicted restoration of perfusing rhythm with a positive predictive value equivalent to that of coronary perfusion pressure. More important, the negative predictive value that a shock would fail to reestablish spontaneous circulation was 96%. AMSA, therefore, has the potential for guiding optimal timing of defibrillation. PMID- 11098950 TI - Suspended animation for delayed resuscitation from prolonged cardiac arrest that is unresuscitable by standard cardiopulmonary-cerebral resuscitation. AB - Standard cardiopulmonary-cerebral resuscitation fails to achieve restoration of spontaneous circulation in approximately 50% of normovolemic sudden cardiac arrests outside hospitals and in essentially all victims of penetrating truncal trauma who exsanguinate rapidly to cardiac arrest. Among cardiopulmonary-cerebral resuscitation innovations since the 1960s, automatic external defibrillation, mild hypothermia, emergency (portable) cardiopulmonary bypass, and suspended animation have potentials for clinical breakthrough effects. Suspended animation has been suggested for presently unresuscitable conditions and consists of the rapid induction of preservation (using hypothermia with or without drugs) of viability of the brain, heart, and organism (within 5 mins of normothermic cardiac arrest no-flow), which increases the time available for transport and resuscitative surgery, followed by delayed resuscitation. Since 1988, we have developed and used novel dog models of exsanguination cardiac arrest to explore suspended animation potentials with hypothermic and pharmacologic strategies using aortic cold flush and emergency portable cardiopulmonary bypass. Outcome evaluation was at 72 or 96 hrs after cardiac arrest. Cardiopulmonary bypass cannot be initiated rapidly. A single aortic flush of cold saline (4 degrees C) at the start of cardiac arrest rapidly induced (depending on flush volume) mild to-deep cerebral hypothermia (35 degrees to 10 degrees C), without cardiopulmonary bypass, and preserved viability during a cardiac arrest no-flow period of up to 120 mins. In contrast, except for one antioxidant (Tempol), explorations of 14 different drugs added to the aortic flush at room temperature (24 degrees C) have thus far had disappointing outcome results. Profound hypothermia (10 degrees C) during 60-min cardiac arrest induced and reversed with cardiopulmonary bypass achieved survival without functional or histologic brain damage. Further plans for the systematic development of suspended animation include the following: a) aortic flush, combining hypothermia with mechanism specific drugs and novel fluids; b) extension of suspended animation by ultraprofound hypothermic preservation (0 degrees to 5 degrees C) with cardiopulmonary bypass; c) development of the most effective suspended animation protocol for clinical trials in trauma patients with cardiac arrest; and d) modification of suspended animation protocols for possible use in normovolemic ventricular fibrillation cardiac arrest, in which attempts to achieve restoration of spontaneous circulation by standard external cardiopulmonary resuscitation advanced life support have failed. PMID- 11098951 TI - Electrophysiology of ventricular fibrillation and defibrillation. AB - The survival rate from ventricular fibrillation is very high for short-duration fibrillation (<30 secs) but decreases to approximately 3% to 30% in out-of hospital conditions. During short-duration fibrillation, action potentials occur rapidly with no intervening period of electrical diastole; a shock defibrillates by interacting with the fibrillation action potential to produce a uniformly long postshock extension of refractoriness. In contrast, during long-duration fibrillation, ischemia-induced degradation of cellular electrophysiology occurs, which causes intervening periods of electrical diastole between fibrillation action potentials and, thus, slowing of fibrillation frequency. A successful defibrillation shock must now not only prolong refractoriness when delivered during the action potential but must also excite cells during the periods of depolarized diastole. Biphasic waveforms enhance both effects by causing premature membrane repolarization with the first pulse, thereby allowing sodium channel recovery from inactivation so that the second pulse produces better formed responses both during the cellular action potential and during the depolarized diastole. Therefore, biphasic waveforms remain superior to monophasic waveforms for treatment of long-duration fibrillation. Improved understanding of the ischemia-induced changes in cellular electrophysiology will suggest further improvements in both defibrillator waveforms and resuscitation techniques. PMID- 11098952 TI - Low-energy biphasic waveform defibrillation reduces the severity of postresuscitation myocardial dysfunction. AB - Both clinical and experimental studies have demonstrated substantial impairment of ventricular function after resuscitation from cardiac arrest. Indeed, postresuscitation myocardial dysfunction has been implicated as a potentially important mechanism, accounting for fatal outcomes after successful resuscitation in 70% of victims within the first 72 hrs. Recent experimental studies implicated the total electrical energy delivered during defibrillation as an important correlate with the severity of postresuscitation myocardial dysfunction and postresuscitation survival. This prompted us to investigate the option of using lower electrical energy biphasic waveform defibrillation. We compared the effects of low-energy biphasic waveform defibrillation with conventional monophasic waveform defibrillation after a short (4 mins), intermediate (7 mins), or prolonged (10 mins) interval of untreated ventricular fibrillation. Biphasic waveform defibrillation with a fixed energy of 150 joules proved to be as effective as conventional monophasic damped sine waveform defibrillation for restoration of spontaneous circulation, with significantly lower delivered energy. This was associated with significantly less severity of postresuscitation myocardial dysfunction. The low-energy biphasic waveform defibrillation is, therefore, likely to be the future direction of transthoracic defibrillation in settings of cardiopulmonary resuscitation. PMID- 11098953 TI - Experimental studies on precordial compression or defibrillation as initial interventions for ventricular fibrillation. AB - Countershock of prolonged ventricular fibrillation is usually followed by asystole or a nonperfusing rhythm. Data from three laboratory investigations indicate that administration of epinephrine and cardiopulmonary resuscitation (CPR) preceding countershock of prolonged ventricular fibrillation significantly improves cardiac resuscitation outcome compared with immediate countershock (relative risk reduction of failed resuscitation, 0.61). Preliminary investigations indicate that a similar improvement is not observed when the ventricular fibrillation period is of shorter duration, e.g., 5 mins. This time interval is probably at the lower limit at which CPR preceding shock of ventricular fibrillation provides benefit in terms of cardiac resuscitation. A single clinical trial of "CPR first" supports the use of a brief period of CPR before countershock of prolonged ventricular fibrillation. Additional trials with and without epinephrine are anticipated. PMID- 11098954 TI - Effects of repetitive electrical shocks on postresuscitation myocardial function. AB - Whereas myocardial cell injury can occur during electrical defibrillation proportional to the energy level of individual shocks, only minimal (or no) injury seems to develop when the energy is limited to the levels typically required to terminate ventricular fibrillation. During cardiac arrest, however, multiple shocks are often required to terminate ventricular fibrillation or to treat episodes that appear subsequently during the resuscitation effort or the postresuscitation interval. Concern exists because an inverse relationship has been reported between the number of electrical shocks delivered during cardiac resuscitation and both resuscitability and survival. Repetitive electrical shocks can alter diastolic function and prompt leftward shifts of the end-diastolic pressure-volume curves. Repetitive shocks may, therefore, contribute to the recently recognized phenomenon of postresuscitation myocardial dysfunction and hamper efforts to reestablish competent myocardial function after resuscitation. Thus, strategies aimed at limiting the number of electrical shocks during cardiopulmonary resuscitation are highly desirable. These may include real-time ventricular fibrillation waveform analysis to improve targeting of individual shocks and efforts (using mechanical and pharmacologic means) to render the myocardium more responsive to individual shocks and to promote greater electrical stability after successful defibrillation. PMID- 11098955 TI - Vasopressin and endothelin during cardiopulmonary resuscitation. AB - Vital organ blood flow during cardiopulmonary resuscitation (CPR) and neurologic recovery after CPR were significantly better in pigs treated with vasopressin compared with epinephrine. Furthermore, two clinical studies evaluating both out of-hospital and inhospital cardiac arrest patients found higher 24-hr survival rates in patients who were resuscitated with vasopressin compared with epinephrine. Scientists at the Leopold Franzens University in Innsbruck, Austria, are currently coordinating a multicenter, randomized clinical trial under the aegis of the European Resuscitation Council to study the effects of vasopressin vs. epinephrine in out-of-hospital cardiac arrest patients. Results of anticipated 1,500 enrolled patients may be available in 2001 and may help to determine the role of vasopressin during CPR. Another new, recently studied vasopressor for CPR is endothelin-1. To date, this vasopressor has only been studied as an intervention in animal CPR models, although plasma levels have been investigated in cardiac arrest patients. Initial reports found improved coronary perfusion pressure when combined with epinephrine. However, the CPR research group of the University of Arizona Sarver Heart Center found excessive vasoconstriction and worse survival than with epinephrine alone. PMID- 11098956 TI - Echo-Doppler observations during cardiac arrest and cardiopulmonary resuscitation. AB - We describe a series of investigations that used transesophageal echo-Doppler observations during cardiac arrest and cardiopulmonary resuscitation. Regular contractions of the left atrium persisted during the initial 7 mins of untreated ventricular fibrillation. Ventricular chamber deformation and mitral valve closing and opening followed precordial compression and relaxation. Stroke volumes computed from differences between diastolic and systolic areas of the left ventricle were predictive of the success of the resuscitation. Progressive decreases in left ventricular compliance were associated with decreases in left ventricular diastolic and stroke volumes and progressed to a stone heart. PMID- 11098957 TI - Prospective evaluation of short-term, high-volume isovolemic hemofiltration on the hemodynamic course and outcome in patients with intractable circulatory failure resulting from septic shock. AB - OBJECTIVE: To evaluate the effects of short-term, high-volume hemofiltration (STHVH) on hemodynamic and metabolic status and 28-day survival in patients with refractory septic shock. DESIGN: Prospective, interventional. SETTING: Intensive care unit (ICU), tertiary institution. PATIENTS: Twenty patients with intractable cardiocirculatory failure complicating septic shock, who had failed to respond to conventional therapy. INTERVENTIONS: STHVH, followed by conventional continuous venovenous hemofiltration. STHVH consisted of a 4-hr period during which 35 L of ultrafiltrate is removed and neutral fluid balance is maintained. Subsequent conventional continuous venovenous hemofiltration continued for at least 4 days. MEASUREMENTS AND MAIN RESULTS: Cardiac index, systemic vascular resistance, pulmonary vascular resistance, oxygen delivery, mixed venous oxygen saturation, arterial pH, and lactate were measured serially. Fluid and inotropic support were managed by protocol. Therapeutic endpoints were as follows during STHVH: a) by 2 hrs, a > or =50% increase in cardiac index; b) by 2 hrs, a > or =25% increase in mixed venous saturation; c) by 4 hrs, an increase in arterial pH to >7.3; d) by 4 hrs, a > or =50% reduction in epinephrine dose. Patients who attained all four goals (11 of 20) were considered hemodynamic "responders"; patients who did not (9 of 20) were considered hemodynamic "nonresponders." There were no differences in baseline hemodynamic, metabolic, and Acute Physiology and Chronic Health Evaluation and Simplified Acute Physiology Scores between responders and nonresponders. Survival to 28 days was better among responders (9 of 11 patients) than among nonresponders (0 of 9). Factors associated with survival were hemodynamic-metabolic response status, time interval from ICU admission to initiation of STHVH, and body weight. CONCLUSIONS: These data suggest that STHVH may be of major therapeutic value in the treatment of intractable cardiocirculatory failure complicating septic shock. Early initiation of therapy and adequate dose may improve hemodynamic and metabolic responses and 28-day survival. PMID- 11098958 TI - Effectiveness of end-tidal carbon dioxide tension for monitoring thrombolytic therapy in acute pulmonary embolism. AB - OBJECTIVE: In acute massive pulmonary embolism with hemodynamic instability, monitoring of pulmonary artery pressure can be used to assess the efficacy of thrombolytic therapy. As a noninvasive alternative to pulmonary artery catheterization, we investigated the efficacy of continuous monitoring of end tidal CO2 tension. DESIGN: In 12 patients with massive pulmonary embolism who required mechanical ventilation, mean pulmonary arterial pressure (MPAP) and end tidal carbon dioxide tension (ETCO2) were registered continuously during thrombolytic therapy. PaCO2, cardiac index as estimated by thermodilution catheter and respiratory ratio of arterial oxygen tension and inhaled oxygen concentration (PaO2/FIO2) were determined every 60 mins. MEASUREMENTS AND MAIN RESULTS: Before thrombolysis, MPAP (34.5+/-9.8 mm Hg) and the difference between PaCO2 and ETCO2 (10.1+/-4.7 mm Hg) were markedly increased compared with normal values. Continuously monitored MPAP was related to ETCO2 for both all patients (r2 = .42; p < .001) and individually (mean r2 = .92; range, .79-.98; p < .001). In ten survivors, the mean cardiac index and PaO2/FIO2 increased during therapy from 1.7+/-0.4 to 2.8+/-0.6 L/min x m2 and 125+/-27 to 285+/-50 mm Hg (p < .01, respectively). In these patients, the difference between PaCO2 and ETCO2 decreased from 9.8+/-4.5 to 2.8+/-0.9 mm Hg (p < .001). Recurrent embolism was detected in two patients by sudden reduction of ETCO2. CONCLUSIONS: Analysis of ETCO2 allows monitoring of the efficacy of thrombolysis and may reflect recurrent embolism. Thus, on the basis of this small study, analysis of ETCO2 appears to be useful for noninvasive monitoring in mechanically ventilated patients with massive pulmonary embolism. PMID- 11098959 TI - Stimulation of beta-adrenergic receptors inhibits the release of tumor necrosis factor-alpha from the isolated rat heart. AB - OBJECTIVES: Beta-adrenergic receptor agonists such as isoproterenol inhibit production of tumor necrosis factor (TNF)-alpha in a number of cell types. Because the heart is a source of TNF-alpha, we hypothesized that isoproterenol would inhibit cardiac production of the cytokine. DESIGN: Analysis of cardiac release of TNF-alpha. SETTING: Medical research laboratory. SUBJECTS: Rats. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: With the approval of the Institutional Animal Care and Use Committee, rats were anesthetized and hearts were removed and perfused. After 30 mins, bacterial lipopolysaccharide (LPS) with or without isoproterenol was infused for 60 mins. At 30, 60, 90, 120, and 150 mins, coronary flow was measured and coronary effluent was analyzed for TNF alpha. Cardiac production of TNF-alpha was expressed as pg/min. Cyclic adenosine monophosphate (AMP) in the coronary effluent was measured. TNF-alpha messenger RNA was determined in ventricular tissue. After 30 mins, TNF-alpha was undetectable in the coronary effluent However, 60 mins after the initiation of LPS infusion, TNF-alpha release was 875+/-255 pg/min and increased to 2164+/-721 pg/min at 150 mins. Simultaneous infusion of isoproterenol with LPS stimulated cyclic AMP release and inhibited TNF-alpha production. For instance, at 60 and 150 mins, TNF-alpha release was 75+/-38 and 58+/-29 pg/min, respectively (p < .05 vs. LPS alone). Simultaneous infusion of isoproterenol with LPS blocked the induction of TNF-alpha messenger RNA by LPS. Isoproterenol, begun 30 mins after the initiation of LPS infusion, still suppressed LPS-stimulated TNF-alpha release by 95% at 150 mins. Similar results were obtained with norepinephrine. CONCLUSIONS: Activation of beta-adrenergic receptors inhibits cardiac TNF-alpha release. This implies that cytokine production by the heart is inhibited by the sympathetic nervous system. In heart failure, the cardiac response to the sympathetic nervous system is impaired. This impairment may play a role in the high plasma levels of TNF-alpha found in heart failure. PMID- 11098960 TI - Long-term health-related quality of life in survivors of sepsis. Short Form 36: a valid and reliable measure of health-related quality of life. AB - OBJECTIVE: To describe the long-term health-related quality of life (HRQL) of survivors of sepsis and to evaluate the reliability and validity of the medical outcomes study Short Form-36 (SF-36) in this population. STUDY DESIGN: Cross sectional survey. SETTING: University intensive care unit. PATIENTS: Surviving patients over the age of 17 yrs who met the criteria for the Society of Critical Care Medicine/American College of Chest Physicians definition of sepsis identified through a review of patients admitted to the intensive care unit from 1994 to 1998. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Baseline demographics and clinical characteristics were abstracted from the medical chart. After hospital discharge, the SF-36 and Patrick's Perceived Quality of Life scale were administered by telephone. The SF-36 was readministered 2 wks later. We screened the charts of 109 patients; 78 had a diagnosis of sepsis. Of these, 31 had died, 3 had severe communication problems, 9 refused to participate, and 5 patients could not be located. A total of 30 patients completed the first interview; 26 completed the second. Compared with established norms for the U.S. general population, survivors of sepsis scored significantly lower on the physical functioning, role physical, general health, vitality, and social functioning domains, as well as on the Physical Health Summary Scale. Mean scores on the Mental Health Summary Scale were very similar between the survivors of sepsis and U.S. norms. The SF-36 demonstrated high internal consistency (Cronbach's alpha ranged from 0.65 to 0.94) and excellent test-retest stability (intraclass correlation coefficient ranged from 0.75 to 0.97). Both the Physical Health Summary Scale and the Mental Health Summary Scale correlated well with overall Perceived Quality of Life scores (Pearson correlation coefficients 0.45 and 0.56, respectively). CONCLUSIONS: The long-term HRQL of survivors of sepsis is significantly lower than that of the general U.S. population. The SF-36 demonstrated good reliability and validity when used to measure HRQL in survivors of sepsis. PMID- 11098961 TI - Hypocaloric total parenteral nutrition: effectiveness in prevention of hyperglycemia and infectious complications--a randomized clinical trial. AB - OBJECTIVE: To determine whether the frequency rate of hyperglycemia and infectious complications can be reduced by an underfeeding strategy in patients requiring total parenteral nutrition (TPN), without deleterious effects on nitrogen balance. DESIGN: Prospective, randomized, controlled nonblinded trial. SETTING: A university-affiliated teaching hospital with a dedicated TPN service. PATIENTS: TPN was initiated in 40 adult patients and continued for > or =5 days. INTERVENTION: Two different TPN feeding strategies were compared: hypocaloric feeding (1 L containing 70 g protein and 1000 kcal) and standard weight-based regimen, begun in similar amounts initially, but advanced in increments toward 25 kcal and 1.5 g protein/kg dry (or adjusted ideal) weight. MEASUREMENTS AND MAIN RESULTS: We evaluated the frequency rate of hyperglycemia, average blood glucose, numbers and types of infections while receiving nutritional support and nitrogen balance after 5 days of TPN. There were significant differences between the quantities of calories, dextrose, fat, and protein provided to the two groups. However, average blood glucose, frequency rate of hyperglycemia, and infection rates (from intravenous catheter, pneumonia, and wound/abdominal collection) were similar in each group. The control group showed a trend toward a higher insulin requirement. Nitrogen balance, only available as a subset, was significantly more negative in the hypocaloric group. CONCLUSIONS: Provision of TPN to a goal of 25 kcal/kg was not associated with more hyperglycemia or infections than a deliberate underfeeding strategy. A regimen of 1.5 g/kg protein in conjunction with 25 kcal/kg did, however, provide significant nutritional benefit in terms of nitrogen balance in comparison with hypocaloric TPN. PMID- 11098962 TI - Midazolam and 2% propofol in long-term sedation of traumatized critically ill patients: efficacy and safety comparison. AB - OBJECTIVE: We proposed to compare the efficacy and safety of midazolam and propofol in its new preparation (2% propofol) when used for prolonged, deep sedation in traumatized, critically ill patients. We also retrospectively compared 2% propofol with its original preparation, 1% propofol, used in a previous study in a similar and contemporary set of patients. DESIGN: A prospective, randomized, unblinded trial (midazolam and 2% propofol) and a retrospective, contemporary trial (2% propofol and 1% propofol). SETTINGS: A trauma intensive care unit in a tertiary university hospital. PATIENTS: A total of 63 consecutive trauma patients, admitted within a period of 5 months and requiring mechanical ventilatory support for >48 hrs, 43 of whom (73%) suffered severe head trauma. We also retrospectively compared the 2% propofol group with a series of patients in whom 1% propofol was used. INTERVENTIONS: For the prospective trial, we randomized two groups--a midazolam group with continuous administration of midazolam at dosages 0.1-0.35 mg/kg/hr, and a 2% propofol group with continuous infusion at dosages 1.5-6 mg/kg/hr. Equal dosages of analgesics were administered. Similar management protocols were applied in the 1% propofol group, used in the retrospective analysis with 2% propofol. MEASUREMENTS AND MAIN RESULTS: Epidemiologic and efficacy variables were recorded. Hemodynamic and biochemical variables were also monitored on a regular basis. Neuromonitoring was also performed on those patients with head trauma. Sedation adequacy was similar and patient behavior after drug discontinuation was not different in either prospective group (midazolam and 2% propofol). Hemodynamic or neuromonitoring variables were also similar for both groups. Triglyceride levels were significantly higher in the 2% propofol group compared with the midazolam group. A higher number of therapeutic failures because of sedative inefficacy was seen in the 2% propofol group compared with the midazolam group, especially during the first sedation days. When comparing 2% propofol and 1% propofol, a significantly higher number of therapeutic failures because of hypertriglyceridemia were found in the 1% propofol group, as opposed to a major number of therapeutic failures because of inefficacy, found in the 2% propofol group. CONCLUSIONS: Propofol's new preparation is safe when used in severely traumatized patients. Its more concentrated formula improves the lipid overload problem seen with the prolonged use of the previous preparation. Nevertheless, a major number of therapeutic failures were detected with 2% propofol because of the need for dosage increase. This fact could be caused by a different disposition and tissue distribution pattern of both propofol preparations. New studies will be needed to confirm these results. PMID- 11098963 TI - Safety of granulocyte colony-stimulating factor (filgrastim) in intubated patients in the intensive care unit: interim analysis of a prospective, placebo controlled, double-blind study. AB - OBJECTIVE: To investigate the safety of the granulocyte colony-stimulating factor filgrastim in the prevention of nosocomial infections in intubated patients in the intensive care unit (ICU), with special emphasis on the possible deleterious effect on acute respiratory distress syndrome (ARDS) and the development of multiple organ dysfunction (MOD). DESIGN: Predetermined, interim analysis of a prospective, randomized, placebo-controlled, double-blind trial. SETTING: University hospital medical-surgical ICU. PATIENTS: A total of 59 consecutive ICU patients, aged >18 yrs, admitted to the ICU no more than 12 hrs before the study, intubated because of ventilatory insufficiency no more than 48 hrs before the study, expected to stay in the ICU for >48 hrs, and had informed consent from the next relative. INTERVENTIONS: Patients were randomized to receive either placebo or 300 microg of filgrastim subcutaneously once daily for 7 days. MEASUREMENTS AND MAIN RESULTS: No significant differences were found in the number of patients developing ARDS (2 of 20 in the placebo group vs. 0 of 22 in the filgrastim group), disseminated intravascular coagulation (3 of 27 vs. 3 of 29), acute renal failure (1 of 27 vs. 1 of 23), or change in MOD. Data analysis showed nosocomial infections in 11 of 29 patients in the placebo group and in 7 of 30 patients in the filgrastim group (p = .266). The median (range) length of ICU stay was 8 (1 34) days in the placebo group and 6 days (1-28) in the filgrastim group. The day 28 mortality rate was 17% (5 of 29) in the placebo group and 13% (4 of 30) in the filgrastim group. No drug-related adverse events occurred. CONCLUSION: Filgrastim is safe in intubated ICU patients, with no excess risk for development of ARDS or MOD. PMID- 11098964 TI - Intensive care unit drug use and subsequent quality of life in acute lung injury patients. AB - OBJECTIVE: To examine the relationship between the use of sedative and neuromuscular blocking agents during a patient's intensive care unit (ICU) stay and subsequent measures of health-related quality of life. DESIGN: Cross sectional mail survey and retrospective medical record abstraction of a prospectively identified cohort of lung injury patients. SETTING: ICUs in three teaching hospitals in a major metropolitan area. PATIENTS: Patients with acute lung injury (n = 24). INTERVENTIONS: None--observational study. MEASUREMENTS AND MAIN RESULTS: Patients' charts were reviewed for those patients returning postdischarge quality-of-life questionnaires. Duration, daily dose, and route of administration for sedatives and neuromuscular blocking agents were abstracted from ICU flow sheets. Relationships among ICU variables (days of sedation, days of neuromuscular blockade, and severity of illness as measured by Acute Physiology and Chronic Health Evaluation III score) and outcomes (symptoms of depression and symptoms of posttraumatic stress disorder) were assessed. Depressive symptoms at follow-up were correlated with days of sedation (p = .007), but not with days of neuromuscular blockade or initial severity of illness. The composite posttraumatic stress disorder symptom impact score was correlated with days of sedation (p = .006) and days of neuromuscular blockade (p = .035), but not with initial severity of illness. There were no significant differences between the frequency of patients reporting a specific posttraumatic stress disorder symptom in the high sedation group and the low sedation group, and there were no significant differences in specific posttraumatic stress disorder symptoms between the group that had received neuromuscular blockade and those who had not. CONCLUSIONS: The use of sedatives and neuromuscular blocking agents in the ICU is positively associated with subsequent measures of depression and posttraumatic stress disorder symptoms 6-41 months after ICU treatment for acute lung injury. PMID- 11098965 TI - Estimating cardiac filling pressure in mechanically ventilated patients with hyperinflation. AB - OBJECTIVE: When positive end-expiratory pressure (PEEP) is applied, the intracavitary left ventricular end-diastolic pressure (LVEDP) exceeds the LV filling pressure because pericardial pressure exceeds 0 at end-expiration. Under those conditions, the LV filling pressure is itself better reflected by the transmural LVEDP (tLVEDP) (LVEDP minus pericardial pressure). By extension, end expiratory pulmonary artery occlusion pressure (eePAOP), as an estimate of end expiratory LVEDP, overestimates LV filling pressure when pericardial pressure is >0, because it occurs when PEEP is present. We hypothesized that LV filling pressure could be measured from eePAOP by also knowing the proportional transmission of alveolar pressure to pulmonary vessels calculated as index of transmission = (end-inspiratory PAOP--eePAOP)/(plateau pressure--total PEEP). We calculated transmural pulmonary artery occlusion pressure (tPAOP) with this equation: tPAOP = eePAOP--(index of transmission x total PEEP). We compared tPAOP with airway disconnection nadir PAOP measured during rapid airway disconnection in subjects undergoing PEEP with and without evidence of dynamic pulmonary hyperinflation. DESIGN: Prospective study. SETTING: Medical intensive care unit of a university hospital. PATIENTS: We studied 107 patients mechanically ventilated with PEEP for acute respiratory failure. Patients without dynamic pulmonary hyperinflation (group A; n = 58) were analyzed separately from patients with dynamic pulmonary hyperinflation (group B; n = 49). INTERVENTION: Transient airway disconnection. MEASUREMENTS AND MAIN RESULTS: In group A, tPAOP (8.5+/-6.0 mm Hg) and nadir PAOP (8.6+/-6.0 mm Hg) did not differ from each other but were lower than eePAOP (12.4+/-5.6 mm Hg; p < .05). The agreement between tPAOP and nadir PAOP was good (bias, 0.15 mm Hg; limits of agreement, -1.5-1.8 mm Hg). In group B, tPAOP (9.7+/-5.4 mm Hg) was lower than both nadir PAOP and eePAOP (12.1+/-5.4 and 13.9+/-5.2 mm Hg, respectively; p < .05 for both comparisons). The agreement between tPAOP and nadir PAOP was poor (bias, 2.3 mm Hg; limits of agreement, -0.2-4.8 mm Hg). CONCLUSIONS: Indexing the transmission of proportional alveolar pressure to PAOP in the estimation of LV filling pressure is equivalent to the nadir method in patients without dynamic pulmonary hyperinflation and may be more reliable than the nadir PAOP method in patients with dynamic pulmonary hyperinflation. PMID- 11098966 TI - Total parenteral nutrition enriched with arginine and glutamate generates glutamine and limits protein catabolism in surgical patients hospitalized in intensive care units. AB - OBJECTIVES: To study the effect of a parenteral nutrition solution enriched with potential precursors of glutamine, i.e., arginine and glutamate, on plasma glutamine concentrations and protein metabolism. DESIGN: Prospective, randomized, single-blind, comparative study. SETTING: Two intensive care units in two different hospitals. PATIENTS: Fifteen surgical patients. INTERVENTIONS: Patients were randomized to receive total parenteral nutrition for 5 days with the enriched glutamine precursor solution (GlnP+ group) or a conventional solution (control group), both total parenteral nutrition providing 0.25 gN/kg per day and 35 kcal/kg per day (glucose/lipids, 70%:30%). MEASUREMENTS AND MAIN RESULTS: Plasma amino acid concentrations before (T0) and after 3 hrs (T3) of perfusion, nitrogen balance (daily and cumulated), and urinary excretion of 3 methylhistidine were measured daily from day 1 to day 5. The two groups were identical for age, weight, severity score, and nitrogen and energy intakes. After a 3-hr perfusion, plasma concentrations of arginine, ornithine, and glutamine increased, and the differences (T3 - T0) were significantly higher in the GlnP+ group: arginine, 107.6+/-7.0 vs. 51.9+/-3.3 (mean over 5 days; p < .001); ornithine, 78.9+/-7.1 vs. 43.6+/-3.1 (p < .001); and glutamine, 32.4+/-8.6 vs. 6.7+/-5.0 micromol/L (p < .05), respectively. A positive correlation was found between arginine and glutamine plasma increases only in the GlnP+ group: r = .45; p < .01 (Spearman's rank-correlation test). Daily and cumulated nitrogen balances were not significantly different between the two groups but were positive (difference from 0) only in the GlnP+ group. The urinary 3 methylhistidine/creatinine ratio decreased significantly from day 1 to day 5 only in the GlnP+ group: 24.5+/-2.7 vs. 18.8+/-2.7 micromol/mmol (p < .05). CONCLUSIONS: Total parenteral nutrition enriched with arginine and glutamate promotes a better nitrogen balance, limits protein myofibrillar catabolism, and generates glutamine, with arginine (not glutamate) probably being the main contributor to the glutamine-generating effect of the solution through the formation of ornithine. PMID- 11098967 TI - The comfort of breathing: a study with volunteers assessing the influence of various modes of assisted ventilation. AB - OBJECTIVE: To assess the subjective feeling of comfort of healthy volunteers breathing on various modes of ventilation used in intensive care. DESIGN: A randomized, prospective, double-blinded, crossover trial using volunteers. SETTING: An intensive care unit (ICU) in a teaching hospital. INTERVENTIONS: We compared, by using healthy volunteers, the subjective feeling of comfort of three modes of ventilation used during the weaning phase of critical illness. We used healthy volunteers to avoid other distracting influences of intensive care that may confound the primary feeling of comfort. The modes we compared were synchronized intermittent mandatory ventilation, assisted spontaneous breathing, and biphasic positive airway pressure. The imposed ventilation was comparable with 50% of the volunteers' normal respiratory effort. The volunteers breathed via a mouthpiece through a ventilator circuit, and the modes of ventilation were introduced in a randomized manner. MEASUREMENTS AND MAIN RESULTS: We measured visual analog scores for comfort for the three modes of ventilation and collected a ranking order and open-ended comments. We demonstrated that at the level of support we imposed, assisted spontaneous breathing was the most comfortable mode of ventilation and that synchronized intermittent mandatory ventilation was the most uncomfortable. These results were strongly supported by both the ranking scale and comments of the volunteers. CONCLUSIONS: Assisted spontaneous breathing was the most comfortable mode of ventilation because the pattern was primarily determined by the volunteer. Synchronized intermittent mandatory ventilation was the most uncomfortable because the ventilatory pattern was imposed on the volunteers, leading to ventilator-volunteer dyssynchrony. We also conclude there is wide individual variation in the subjective feeling of comfort. Whereas the mode of ventilation in ICUs is based primarily on the physiologic needs of the patient, the feeling of comfort may be considered when choosing an appropriate mode of ventilation during the weaning phase of critical illness. PMID- 11098968 TI - Jejunal and gastric mucosal perfusion versus splanchnic blood flow and metabolism: an observational study on postcardiac surgical patients. AB - OBJECTIVES: To evaluate the association between changes in total splanchnic and mucosal perfusion, assessed either by gastric tonometry or jejunal laser Doppler flowmetry in postcardiac surgical patients. DESIGN: A prospective, observational study. SETTINGS: A general intensive care unit in a tertiary care center. PATIENTS: Twelve, postoperative cardiac surgery patients were studied. INTERVENTIONS: Patients were treated according to clinical routine. Total splanchnic blood flow (indocyanine green extraction), jejunal mucosal perfusion (laser Doppler flowmetry), gastric mucosal-arterial PCO2 gradients, and splanchnic lactate uptake were studied during four 30-min measurements periods, each separated by a period of 1 hr. MEASUREMENTS AND MAIN RESULTS: There was no consistent association between either total splanchnic and local mucosal perfusion or between gastric and jejunal perfusion as assessed by two different techniques. The PCO2 gradient increased from 0.73+/-0.21 kPa to 1.15+/-0.30 kPa (p < .05), and splanchnic oxygen extraction increased from 40%+/-9% to 49%+/-14% (p < .01). CONCLUSIONS: In this observational study on postcardiac surgical patients, local mucosal perfusion did not reflect total splanchnic blood flow and vice versa. Either changes in gastric and jejunal mucosal perfusion were different or increasing tissue metabolism was responsible for the observed lack of association between tonometry, laser Doppler flowmetry, and total splanchnic blood flow. Increasing mucosal arterial PCO2 gradient and splanchnic oxygen extraction may reflect a mismatch between splanchnic perfusion and metabolic demands. PMID- 11098969 TI - Cysteinyl-leukotriene generation as a biomarker for survival in the critically ill. AB - OBJECTIVE: To evaluate the capacity of cysteinyl-leukotriene generation in the progression of critical illness compared with that in healthy volunteers and to clarify interrelationships between the rate of leukotriene generation, severity of the disease, and clinical outcome. DESIGN: Prospective, observational study. SETTING: Surgical intensive care unit (ICU) in a German university hospital. PATIENTS: We studied 14 ICU patients (nine men, five women; aged 42-82 yrs) suffering from systemic inflammatory response syndrome, sepsis, or sepsis syndrome, with a calculated sepsis severity score of 17.7+/-4.2 and a Simplified Acute Physiology score of 17.6+/-3.0. In addition, five healthy volunteers (three men, two women; aged 34-38 yrs) were included in the study. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Blood samples were obtained every second day from septic patients until discharge from the ICU or death. Leukotriene C4 (LTC4) synthesizing capacity was assessed in isolated and stimulated leukocytes (Ca ionophore) by using combined reversed-phase, high-pressure liquid chromatography and radioimmunoassay methods. Initially, all patients synthesized less LTC4 than the healthy subjects. In patients who did not survive, the low LTC4 generation persisted throughout the observation period, whereas in surviving patients, its formation was normalized during convalescence. In surviving patients, LTC4 concentrations correlated with sepsis severity score. CONCLUSIONS: LTC4 generation is impaired in sepsis and may serve as a biomarker for survival in the critical ill. PMID- 11098970 TI - Procalcitonin release patterns in a baboon model of trauma and sepsis: relationship to cytokines and neopterin. AB - OBJECTIVES: Procalcitonin (PCT) has been described as an early, discriminating marker of bacteria-associated sepsis in patients. However, little is known of its source and actions, in part because no appropriate animal models have been available. We tested the hypothesis that plasma PCT increases during various pathophysiological conditions, such as hemorrhagic shock and sepsis, which differ with regard to the degree of associated endotoxemia. We further hypothesized that in sepsis, PCT would be significantly different in survivors vs. nonsurvivors. DESIGN: Prospective, blinded analysis of previously collected plasma of experimental animals. SETTING: Independent nonprofit research laboratory in a trauma hospital and a contract research institute. SUBJECTS: A total of 22 male baboons (17.5-31 kg). INTERVENTIONS: Hemorrhagic-traumatic shock was induced by hemorrhage for up to 3 hrs, reperfusion with shed blood and infusion of cobra venom factor (n = 7). By using a similar experimental setup, severe hyperdynamic sepsis was induced (n = 15) by intravenous infusion of live Escherichia coli (2 x 10(9) colony-forming units/kg) over 2 hrs, followed by antibiotic therapy (gentamicin 4 mg/kg twice a day). MEASUREMENTS AND MAIN RESULTS: Plasma PCT at baseline was barely detectable, but levels increased significantly (p < .05) to 2+/-1.8 pg/mL 2 hrs after the start of reperfusion in the shock group, and to 987+/-230 pg/mL at 4 hrs after E. coli in the sepsis group. Levels were maximal between 6 and 32 hrs and had returned nearly to baseline levels at 72 hrs. Interleukin-6 levels paralleled the course of PCT measurements, whereas a significant increase in neopterin was seen at 24 hrs. PCT levels were approximately three times higher in the sepsis group than in the shock group, corresponding to endotoxin levels (at the end of hemorrhage, 286+/-144 pg/mL vs. 3576+/-979 pg/mL at the end of E. coli infusion; p = .003). PCT levels were significantly different at 24 hrs between survivors (2360+/-620 pg/mL) and nonsurvivors (4776+/-563 pg/mL) in the sepsis group (p = .032), as were interleukin-6 (1562+/-267 vs. 4903+/-608 pg/mL; p = .01) and neopterin/creatinine ratio (0.400+/-0.038 vs. 0.508+/-0.037; p = .032). CONCLUSIONS: PCT is detectable in the baboon as in humans, both in hemorrhagic shock and sepsis. PCT levels are significantly higher in sepsis than in hemorrhage, a finding that is probably related to the differences in endotoxin. The baboon can be used for the study of PCT kinetics in both models; PCT kinetics are clearly different from other markers of sepsis, either IL-6 or neopterin, in both models. There are significant differences between survivors and nonsurvivors in the sepsis model. PMID- 11098971 TI - Pediatric cardiac output measurement using surface integration of velocity vectors: an in vivo validation study. AB - OBJECTIVE: To test the accuracy and reproducibility of systemic cardiac output (CO) measurements using surface integration of velocity vectors (SIVV) in a pediatric animal model with hemodynamic instability and to compare SIVV with traditional pulsed-wave Doppler measurements. DESIGN: Prospective, comparative study. SETTING: Animal research laboratory at a university medical center. SUBJECTS: Eight piglets weighing 10-15 kg. INTERVENTIONS: Hemodynamic instability was induced by using inhalation of isoflurane and infusions of colloid and dobutamine. MEASUREMENTS: SIVV CO was measured at the left ventricular outflow tract, the aortic valve, and ascending aorta. Transit time CO was used as the reference standard. RESULTS: There was good agreement between SIVV and transit time CO. At high frame rates, the mean difference +/- 2 SD between the two methods was 0.01+/-0.27 L/min for measurements at the left ventricular outflow tract, 0.08+/-0.26 L/min for the ascending aorta, and 0.06+/-0.25 L/min for the aortic valve. At low frame rates, measurements were 0.06+/-0.25, 0.19+/-0.32, and 0.14+/-0.30 L/min for the left ventricular outflow tract, ascending aorta, and aortic valve, respectively. There were no differences between the three sites at high frame rates. Agreement between pulsed-wave Doppler and transit time CO was poorer, with a mean difference +/- 2 SD of 0.09+/-0.93 L/min. Repeated SIVV measurements taken at a period of relative hemodynamic stability differed by a mean difference +/-2 SD of 0.01+/-0.22 L/min, with a coefficient of variation = 7.6%. Intraobserver coefficients of variation were 5.7%, 4.9%, and 4.1% at the left ventricular outflow tract, ascending aorta, and aortic valve, respectively. Interobserver variability was also small, with a coefficient of variation = 8.5%. CONCLUSIONS: SIVV is an accurate and reproducible flow measurement technique. It is a considerable improvement over currently used methods and is applicable to pediatric critical care. PMID- 11098972 TI - A single endotoxin injection in the rabbit causes prolonged blood vessel dysfunction and a procoagulant state. AB - OBJECTIVES: To determine the duration of vascular blood vessel dysfunction and coagulation abnormalities after administration of endotoxin in a nonlethal septic rabbit model. DESIGN: Randomized, controlled, interventional trial. SETTING: University animal laboratory. SUBJECTS: A total of 30 male New Zealand White rabbits, randomly assigned to one of two groups. INTERVENTIONS: Male New Zealand White rabbits were randomly divided into control or lipopolysaccharide (LPS) (0.5 mg/kg iv bolus Escherichia coli endotoxin)-treated groups. Metabolic acidosis and coagulation activation confirmed the presence of septic shock. The abdominal aorta was removed at 24 hrs (day 1), day 5, or day 21 after LPS injection. Immunohistochemical staining for an endothelial cell marker (PECAM-1/CD31) was performed to assess endothelial injury. Endothelium-dependent vascular relaxation was analyzed by in vitro vascular reactivity studies. Responses to acetylcholine, to calcium ionophore (A-23187), and to sodium nitroprusside were studied. In addition, arterial blood samples were collected on day 1, day 5, and day 21 for measurement of clotting factors and tissue factor activity. MEASUREMENTS AND MAIN RESULTS: LPS injection resulted in endothelial injury, with loss of approximately 25% of the endothelial area on day 5, which disappeared on day 21. LPS injection also caused a significantly reduced relaxation response to acetylcholine (44.9% +/-9.9% vs. 76.5%+/-5.4% for the control group; p < .005), which was restored on day 21. In contrast, vascular relaxation in response to A-23187 and sodium nitroprusside was not altered. A significant decrease in the platelet count was observed on day 1, associated with a decrease in factors II and V. On day 5, platelet count and factors II and V were corrected in conjunction with an elevated monocyte tissue factor activity in LPS-injected rabbits. On day 21, coagulation abnormalities were corrected. CONCLUSIONS: A single endotoxin injection in the rabbit was responsible for prolonged aortic endothelial cell dysfunction, as well as a prolonged procoagulant state. The latter is a potential trigger for disseminated intravascular coagulation. Importantly, these features are associated with normalization of conventional biological evidence of septic shock. PMID- 11098973 TI - Inhibition of nitric oxide synthase enhances the myocardial toxicity of phenylpropanolamine. AB - OBJECTIVE: To investigate the direct and indirect effects of the anorexic agent phenylpropanolamine (PPA) on the heart and to determine whether nitric oxide deficiency exacerbates the myocardial toxicity of PPA. DESIGN: Dose response effects using sequential drug administration. SETTING: Animal research laboratory of a large tertiary academic medical center. SUBJECTS: Isolated hearts (n = 8) from male Sprague-Dawley rats weighing 300-400 g. INTERVENTIONS: Measurement of heart rate, maximal change in pressure over time (dP/dtmax), -dP/dtmax, and coronary blood flow in isolated hearts perfused on a Langendorff apparatus. PPA was infused through the aortic cannula at 0.05, 0.125, 0.25, 0.5, and 1.25 mmol/L before and after inhibition of nitric oxide synthesis with N-nitro-L-arginine methyl ester (L-NAME). RESULTS: PPA had little effect on myocardial contractility of normal hearts until the highest dose of PPA (1.25 mmol/L). However, after L NAME, PPA significantly depressed contractility at a dose of 0.25 mmol/L. PPA had no significant effects on coronary blood flow. PPA failed to induce arrhythmias in normal hearts. However, after L-NAME, PPA induced ventricular fibrillation in 50% of the hearts. CONCLUSION: PPA causes myocardial contractile depression without altering global coronary artery blood flow. Inhibition of nitric oxide synthesis sensitizes the heart to the myocardial depressant effects of PPA and increases the risk for ventricular fibrillation. PMID- 11098974 TI - Contact activation in shock caused by invasive group A Streptococcus pyogenes. AB - OBJECTIVES: The aim of this study was to characterize abnormalities of coagulation in mice with experimental, invasive group A, streptococcal shock, in an attempt to explain the prolongation of the activated partial thromboplastin time identified in patients with streptococcal toxic shock syndrome. DESIGN: A longitudinal descriptive animal model study of coagulation times and single coagulation factors in mice infected with Streptococcus pyogenes. This was followed by an experimental study to determine whether streptococci or streptococcal products could activate the human contact system in vitro. SETTING: University infectious diseases and hemostasis molecular biology laboratories. SUBJECTS: CD1 outbred mice. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Coagulation times, single factor assays, and bradykinin assays were conducted on murine plasma at different times after streptococcal infection and compared with uninfected mice. In experiments in which streptococcal products were co-incubated with human plasma, we compared coagulation times, single factor assays, and activities against a range of chromogenic substrates with control plasma. In a murine model of streptococcal necrotizing fasciitis, the activated partial thromboplastin times were significantly prolonged in infected mice compared with controls, whereas prothrombin times were normal, suggesting an isolated abnormality of the intrinsic pathway. Bleeding was not seen. Prolongation of activated partial thromboplastin time was associated with reduced factor XII and prekallikrein, whereas levels of factors VIII, IX, XI, and high molecular weight kininogen were elevated. In vitro studies suggested that streptococcal supernatants can activate prekallikrein, in addition to causing plasminogen activation through the action of streptokinase. CONCLUSIONS: Prolongation of activated partial thromboplastin time in streptococcal toxic shock syndrome is associated with activation of the contact system, possibly contributing to the profound shock associated with streptococcal toxic shock syndrome. PMID- 11098975 TI - Inducible nitric oxide synthase gene knockout mice have increased resistance to gut injury and bacterial translocation after an intestinal ischemia-reperfusion injury. AB - OBJECTIVE: Intestinal ischemia-reperfusion after severe shock states is often associated with bacterial translocation and intestinal barrier dysfunction. Our previous studies showed that inducible nitric oxide synthase (iNOS) gene knockout mice were resistant to endotoxin-induced bacterial translocation and ileal mucosal damage. The goal of this study was to test whether iNOS mediates bacterial translocation after intestinal ischemia-reperfusion, using iNOS knockout mice (iNOS-/-) and their wild-type littermates (iNOS+/+). DESIGN: Prospective animal study with concurrent controls. SETTING: Small animal laboratory. SUBJECTS: Thirty-eight iNOS knockout mice and 51 wild-type littermates. INTERVENTIONS: iNOS+/+ mice or iNOS-/- mice were subjected to a sham operation or 30 mins of superior mesenteric artery occlusion followed by reperfusion. Twenty-four hours after reperfusion, bacterial translocation to mesenteric lymph nodes, ileal villous damage, and cecal bacterial population were evaluated. MEASUREMENTS AND MAIN RESULTS: Sham operation did not induce bacterial translocation, change cecal bacterial population levels, or cause ileal villous damage. Intestinal ischemia-reperfusion caused bacterial translocation in 72% of the iNOS+/+ mice but only 28% of the iNOS-/- mice. Both iNOS+/+ and iNOS-/- mice subjected to superior mesenteric artery occlusion (SMAO) in which bacterial translocation occurred had cecal bacterial population levels that were three logs higher than mice subjected to sham SMAO or mice subjected to SMAO in which bacterial translocation did not occur. The magnitude of villous injury was less in the iNOS-/- mice than the iNOS+/+ mice after SMAO, although the incidence of ileal villous damage was significantly higher in both the iNOS+/+ and iNOS-/- mice in which bacterial translocation occurred after SMAO than in the mice in which bacterial translocation did not occur after SMAO. iNOS+/+ mice subjected to SMAO had increased plasma concentrations of nitrite (NO2-) and nitrate (NO3-), and the plasma concentrations of NO2- and NO3- were highest in the mice in which bacterial translocation had occurred. CONCLUSION: iNOS knockout mice were more resistant to intestinal ischemia-reperfusion-induced bacterial translocation and mucosal injury than wild-type mice, suggesting that iNOS might play a role in intestinal ischemia-reperfusion-induced loss of gut barrier function. PMID- 11098976 TI - An improved in vivo rat model for the study of mechanical ventilatory support effects on organs distal to the lung. AB - OBJECTIVE: To study the influence of different mechanical ventilatory support strategies on organs distal to the lung, we developed an in vivo rat model, in which the effects of different tidal volume values can be studied while maintaining other indexes. DESIGN: Prospective, randomized animal laboratory investigation. SETTING: University laboratory of Ospedale Maggiore di Milano Instituto di Ricovero e Cura a Carattere Scientifico. SUBJECTS: Anesthetized, paralyzed, and mechanically ventilated male Sprague-Dawley rats. INTERVENTIONS: Two groups of seven rats each were randomized to receive tidal volumes of either 25% or 75% of inspiratory capacity (IC), calculated from a preliminary estimation of total lung capacity. Ventilation strategies for the two groups were as follows: a) 25% IC, 9.9+/-0.8 mL/kg; frequency, 59+/-4 beats/min; positive end expiratory pressure, 3.6+/-0.8 cm H2O; and peak inspiratory airway pressure (Paw), 13.2+/-2 cm H20; and b) 75% IC, 29.8+/-2.9; frequency, 23+/-13; positive end-expiratory pressure, 0; peak inspiratory Paw, 29.0+/-3. MEASUREMENTS AND MAIN RESULTS: Mean arterial pressure (invasively monitored) remained well above adequate perfusion pressure values throughout, and no significant difference was seen between the two groups. PaO2, pHa, and PaCO2 values were compared after 60 mins of ventilation and again, no significant difference was seen between the two groups (PaO2, 269+/-25 and 260+/-55 torr; pHa, 7.432+/-0.09 and 7.415+/-0.03; PaCO2, 35.4+/-8 and 32.5+/-2 torr, for the 25% IC and 75% IC groups, respectively). Mean Paws were not different (6.4+/-0.8 cm H2O in the 25% IC groups, and 6.1+/-1.2 in the 75% IC groups, respectively). At the end of the experiment, animals were killed and the liver and kidney isolated, fixed in 4% formalin, cut, and stained for optic microscopy. Kidneys from rats ventilated with 75% IC showed increased Bowman's space with collapse of the glomerular capillaries. This occurred in a greater percentage of rats ventilated with 75% IC (0.67+/-0.2 vs. 0.29+/-0.2, 75% IC vs. 25% IC, respectively; p < .05). Perivascular edema was also present in rats ventilated with 75% IC (p < .05). Morphometric determinations of the empty zones (index of edema) demonstrated a trend toward differences between 75% IC livers and 25% IC (0.14+/-0.05 vs. 0.11+/ 0.02, respectively). CONCLUSION: We conclude that it is possible to study the effects of mechanical ventilatory support on organs distal to the lung by means of an in vivo rat model. PMID- 11098977 TI - Bacterial translocation and tumor necrosis factor-alpha gene expression in experimental hemorrhagic shock. AB - OBJECTIVE: To investigate whether bacterial translocation is the causative mechanism underlying cytokine production during hemorrhagic shock. DESIGN: Prospective, randomized, unblinded animal study. SETTING: Surgical research laboratories of Shiga University of Medical Science. SUBJECTS: Male Sprague Dawley rats. INTERVENTIONS: The rats were randomly divided into three groups. Each animal was anesthetized with pentobarbital, given a continuous infusion of 0.9% saline, and monitored for blood pressure. The normoxic and sham shock groups breathed room air, whereas the hyperoxic shock group was administered 100% oxygen. Except in the sham shock group, blood was withdrawn to induce a hemorrhagic shock state, then the shed blood was reinfused. Sixty minutes after the induction of hemorrhagic shock, arterial blood cultures were performed in all three groups. The animals were then killed, and their mesenteric lymph nodes (MLNs) were harvested for bacterial culture. The terminal ileum, liver, spleen, kidney, lung, and MLNs were also collected for histologic study by in situ hybridization. MEASUREMENTS AND MAIN RESULTS: In the bacteriologic study, the prevalence of bacterial translocation was 0% (0/11) in the hyperoxic shock group, 55% (6/11) in the normoxic shock group, and 0% (0/9) in the sham shock group. In the in situ hybridization study, tumor necrosis factor-alpha gene expression was detected only in the ileal tissue, MLNs, and spleens of the normoxic shock group. Blood cultures were sterile in all three groups. CONCLUSIONS: Bacterial translocation occurred in MLNs within 1 hr of hemorrhage. Hemorrhagic shock causes tumor necrosis factor-alpha gene expression as well as bacterial translocation in MLNs, but not in the liver, in this model. Bacterial translocation was prevented by hyperoxia early in the course of hemorrhagic shock. Hyperoxia also prevented tumor necrosis factor-alpha gene expression along the bacterial invasion route. PMID- 11098978 TI - Effect of neutropenia and granulocyte colony stimulating factor-induced neutrophilia on blood-brain barrier permeability and brain edema after traumatic brain injury in rats. AB - OBJECTIVE: Granulocyte colony stimulating factor (GCSF) has been used to increase systemic absolute neutrophil count (ANC) in patients with severe traumatic brain injury to reduce nosocomial infection risk. However, the effect of increasing systemic ANC on the pathogenesis of experimental traumatic brain injury has not been studied. Thus, we evaluated the effect of systemic ANC on blood-brain barrier (BBB) damage and brain edema after traumatic brain injury in rats. DESIGN: Experimental study. SETTING: Research laboratory at the University of Pittsburgh, PA. SUBJECTS: Forty-three adult male Sprague-Dawley rats. INTERVENTIONS: Protocol I: rats were randomized to receive either vinblastine sulfate to reduce ANC, GCSF to increase ANC, or saline before controlled cortical impact (CCI) of moderate overall severity. Evans blue was used to assess BBB damage at 4-24 hrs after CCI. Protocol II: rats received GCSF or saline before CCI. Brain edema was estimated at 24 hrs using wet - dry) / wet weight method. Protocol III: rats received GCSF or saline before CCI. Brain neutrophil accumulation was estimated at 24 hrs using a myeloperoxidase assay. MEASUREMENTS AND MAIN RESULTS: Physiologic variables were controlled before CCI was maintained at normal in all animals before traumatic brain injury. No rats were anemic, hypoglycemic, or hypotensive before CCI. Protocol I: compared with control, systemic ANC decreased in vinblastine-treated rats and increased in GCSF-treated rats. BBB damage correlated with systemic ANC. Protocol II: mean systemic ANC before traumatic brain injury increased 15-fold in rats given GCSF vs. control; however no difference in brain edema was observed at 24 hrs after injury between groups. Protocol III: median systemic ANC at the time of CCI was increased ten fold in rats given GCSF vs. control. No difference in brain myeloperoxidase activity 24 hrs after CCI was observed in rats treated with GCSF vs. control. CONCLUSIONS: Systemic ANC influences BBB damage after traumatic brain injury produced by CCI. Because BBB damage and brain edema are discordant, mechanisms other than BBB damage likely predominate in the pathogenesis of brain edema after contusion. The implications of increased BBB permeability with the administration of GCSF in our model remains to be determined. Increasing systemic ANC before CCI with GCSF administration does not increase posttraumatic brain neutrophil accumulation or brain edema after CCI in rats. The finding that neutrophil infiltration is not enhanced by systemic neutrophilia suggests that the ability of GCSF-stimulated neutrophils to migrate into injured tissue may be impaired. Further studies are needed to evaluate the effects of GCSF administration on secondary injury and functional outcome in experimental models of traumatic brain injury. PMID- 11098979 TI - Improved outcomes of children with malignancy admitted to a pediatric intensive care unit. AB - OBJECTIVE: To assess the acute and long-term outcomes of children admitted to the intensive care unit with cancer or complications after bone marrow transplantation. DESIGN: Retrospective analysis of databases from a prospective pediatric intensive care unit (PICU) database supplemented by case notes review. SETTING: A PICU in a tertiary pediatric hospital. PATIENTS: All children with malignancy admitted to the PICU between May 1, 1987, and April 30, 1996. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: There were 206 admissions to the PICU during a 9-yr study period of 150 children with malignancies or complications after bone marrow transplantation. Forty patents died in the PICU (27% mortality rate). The most frequent indications for PICU admission were shock and respiratory disease. Of 56 children admitted with shock, there were 16 deaths (29% mortality rate). In 24 episodes of sepsis, inotropic and ventilatory support were required and 13 patients (54%) survived. Analysis of long-term survival gave estimates of 50% survival for all oncology patients admitted to the PICU and 42% for those admitted for shock. CONCLUSIONS: A high proportion of oncology patients admitted to the PICU requiring intensive intervention survive and go on to be cured of their malignancy. Our study suggests the PICU outcome for these patients has improved. PMID- 11098980 TI - Inhaled nitric oxide reduces the need for extracorporeal membrane oxygenation in infants with persistent pulmonary hypertension of the newborn. AB - OBJECTIVE: We previously reported improved oxygenation, but no change, in rates of extracorporeal membrane oxygenation (ECMO) use or death among infants with persistent pulmonary hypertension of the newborn who received inhaled nitric oxide (NO) with conventional ventilation, irrespective of lung disease. The goal of our study was to determine whether treatment with inhaled NO improves oxygenation and clinical outcomes in infants with persistent pulmonary hypertension of the newborn and associated lung disease who are ventilated with high-frequency oscillatory ventilation (HFOV). DESIGN: Single-center, prospective, randomized, controlled trial. SETTING: Newborn intensive care unit of a tertiary care teaching hospital. PATIENTS: We studied infants with a gestational age of > or =34 wks who were receiving mechanical ventilatory support and had echocardiographic and clinical evidence of pulmonary hypertension and hypoxemia (PaO2 < or =100 mm Hg on FIO2 = 1.0), despite optimal medical management Infants with congenital heart disease, diaphragmatic hernia, or other major anomalies were excluded. INTERVENTIONS: The treatment group received inhaled NO, whereas the control group did not. Adjunct therapies and ECMO criteria were the same in the two groups of patients. Investigators and clinicians were not masked as to treatment assignment, and no crossover of patients was permitted. MEASUREMENTS AND MAIN RESULTS: Primary outcome variables were mortality and use of ECMO. Secondary outcomes included change in oxygenation and duration of mechanical ventilatory support and supplemental oxygen therapy. Forty-two patients were enrolled. Baseline oxygenation and clinical characteristics were similar in the two groups of patients. Infants in the inhaled NO group (n = 21) had improved measures of oxygenation at 15 mins and 1 hr after enrollment compared with infants in the control group (n = 20). Fewer infants in the inhaled NO group compared with the control group were treated with ECMO (14% vs. 55%, respectively; p = .007). Mortality did not differ with treatment assignment. CONCLUSIONS: Among infants ventilated by HFOV, those receiving inhaled NO had a reduced need for ECMO. We speculate that HFOV enhances the effectiveness of inhaled NO treatment in infants with persistent pulmonary hypertension of the newborn and associated lung disease. PMID- 11098981 TI - Comparison of the response of saline tonometry and an automated gas tonometry device to a change in CO2. AB - OBJECTIVE: To examine the speed of response of saline tonometry and an automated gas tonometry system by using standard tonometry catheters. DESIGN: In vitro validation study. SETTING: Experimental research laboratory. INTERVENTIONS: Tonometry catheters were placed in a test chamber designed to simulate the lumen of a hollow viscus and were exposed to a rapid change in CO2 from 0% to 5% or 10%. Measured CO2 over time was fit to a mathematical model to determine the response time constant (the time to reach 63% of the final value) for each system. MEASUREMENTS AND MAIN RESULTS: Response time to a change in CO2 was significantly faster with the automated gas system than with traditional saline tonometry. The mathematical time constant for a 5% change in CO2 in a gas environment was 2.8 mins (95% confidence interval, 2.6-3.0 mins) for the gas and 6.3 mins (95% confidence interval, 5.8-7.3 mins) for the saline technique. These times were longer for the CO2 change in a liquid environment: The time constant was 4.6 mins (95% confidence interval, 4.5-4.7 mins) for the gas system and 7.8 mins (95% confidence interval, 7.15-8.6 mins) for the saline tonometry. There was a significantly lower final equilibration value for the CO2 measurement with saline tonometry. There was essentially no difference in time constants for each system for a 5% change compared with a 10% CO2 change, except for a slightly faster time constant for the gas tonometry system with a 5% change in the gas environment (5%: 2.8 mins vs. 10%: 3.3 mins). CONCLUSIONS: The automated gas tonometry system has a significantly faster response to a change in CO2 than conventional saline tonometry. PMID- 11098982 TI - Percutaneous tracheostomy in critically ill patients: a prospective, randomized comparison of two techniques. AB - OBJECTIVE: To prospectively compare two commonly used methods for percutaneous dilational tracheostomy (PDT) in critically ill patients. DESIGN: Prospective, randomized, clinical trial. SETTING: Trauma and general intensive care units of a university tertiary teaching hospital, which is also a level 1 trauma center. PATIENTS: One hundred critically ill patients with an indication for PDT. INTERVENTIONS: PDT with the Ciaglia technique using the Ciaglia PDT introducer set and the Griggs technique using a Griggs PDT kit and guidewire dilating forceps. MEASUREMENTS AND MAIN RESULTS: Surgical time, difficulties, and surgical and anesthesia complications were measured at 0-2 hrs, 24 hrs, and 7 days postprocedure. Groups were well matched, and there were no differences between the two methods in surgical time or in anesthesia complications. Major bleeding complications were 4.4 times more frequent with the Griggs PDT kit. With the Ciaglia PDT kit, both intraoperative and at 2 and 24 hrs, surgical complications were less common (p = .023) and the procedure was more often completed without expert assistance (p = .013). Tracheostomy bleeding was not associated with either anticoagulant therapy or an abnormal clotting profile. Multivariate analysis identified the predictors of PDT complications as the Griggs PDT kit (p = .027) and the Acute Physiology and Chronic Health Evaluation (APACHE) II score (p = .041). The significant predictors of time required to complete PDT were the APACHE II score (p = .041), a less experienced operator (p = .0001), and a female patient (p = .013). CONCLUSIONS: Patients experiencing PDT with the Ciaglia PDT kit had a lower surgical complication rate (2% vs. 25%), less operative and postoperative bleeding, and less overall technical difficulties than did patients undergoing PDT with the Griggs PDT kit. Ciaglia PDT is, therefore, the preferred technique for percutaneous tracheostomy in critically ill patients. PMID- 11098983 TI - Longitudinal study of pediatric house officers' attitudes toward death and dying. AB - OBJECTIVE: To investigate pediatric residents' attitudes toward end-of-life issues and their education in dealing with these issues. DESIGN: Exploratory survey. SETTING: Department of Pediatrics at the University of California, Los Angeles, Center for Health Sciences. SUBJECTS: Volunteer sample. A total of 182 of 203 pediatric residents at all levels of training completed anonymous questionnaires. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Data on residents' attitudes toward issues of death and dying and the efficacy of educational interventions were collected over a 4-yr period. When entering training, house officers are uncomfortable dealing with death and dying issues (mean, 3.3 of 5; 5 = not comfortable). By the end of their training, these house officers become comfortable dealing with these issues (mean, 2.2; p < .05). During their first 2 yrs of training, house officers report that their medical education is not helping them to deal with the issues of death and dying (mean, 3.3). At the end of their third year of training, residents report that their education is helping them to deal with these issues (mean, 2.5; p < .05). Strikingly, as house officers progress through their residency, they become less comfortable with the idea of administering pain medication to a dying patient, because the pain medication might hasten the patient's death (p < .05). CONCLUSIONS: Pediatric residents may benefit from more formal training in the practical aspects of death and dying issues. Residency education should do more to address these issues systematically for the benefit of both the residents and the patients and family members. PMID- 11098984 TI - Position paper on critical care pharmacy services. Society of Critical Care Medicine and American College of Clinical Pharmacy Task Force on Critical Care Pharmacy Services. AB - OBJECTIVE: The goal of the Task Force on Critical Care Pharmacy Services was to identify and describe the scope of practice that characterizes the critical care pharmacist and critical care pharmacy services. Specifically, the aims were to define the level of clinical practice and specialized skills characterizing the critical care pharmacist as clinician, educator, researcher, and manager; and to recommend fundamental, desirable, and optimal pharmacy services and personnel requirements for the provision of pharmaceutical care to critically ill patients. Hospitals having comprehensive resources as well as those with more limited resources were considered. DATA SOURCES: Consensus opinion of critical care pharmacists from institutions of various sizes providing critical care services within several types of pharmacy practice models was obtained, including community-based and academic practice settings. Existing guidelines and literature describing pharmacy practice and medication use processes were reviewed and adapted for the critical care setting. CONCLUSIONS: By combining the strengths and expertise of critical care pharmacy specialists with existing supporting literature, these recommendations define the level of clinical practice and specialized skills that characterize the critical care pharmacist as clinician, educator, researcher, and manager. This Position Paper recommends fundamental, desirable, and optimal pharmacy services as well as personnel requirements for the provision of pharmaceutical care to critically ill patients. PMID- 11098985 TI - Hemofiltration in sepsis: is removal of "bad humors" the answer? PMID- 11098986 TI - A new niche for end-tidal CO2 in pulmonary embolism. PMID- 11098987 TI - Regulating cardiac cytokine expression: the role of the adrenergic nervous system. PMID- 11098988 TI - Quality of life in intensive care unit survivors: a place for outcomes research in critical care. PMID- 11098989 TI - Total parenteral nutrition: can we decrease infectious complications? PMID- 11098990 TI - The use of granulocyte colony-stimulating factor in critically ill patients. PMID- 11098991 TI - A step forward--but the path is still unclear. PMID- 11098992 TI - An alternative method to supplement glutamine in parenteral nutrition? PMID- 11098993 TI - Prognostic markers in sepsis: the role of leukotrienes. PMID- 11098994 TI - Lose weight...NO problem. PMID- 11098995 TI - Keep in contact: the role of the contact system in infection and sepsis. PMID- 11098996 TI - X's and O's. PMID- 11098997 TI - Oncology patients in the pediatric intensive care unit: room for optimism? PMID- 11098998 TI - Teaching physicians about end-of-life issues: we need to do a better job. PMID- 11098999 TI - Critical care pharmacists: an essential or a luxury? PMID- 11099000 TI - Severity of illness scoring in obstetrical patients. PMID- 11099001 TI - Where's the beef? PMID- 11099002 TI - Venous thromboembolism D-dimer and the intensive care unit. PMID- 11099003 TI - Ethics can boost science. PMID- 11099004 TI - European panel rejects creation of human embryos for research. PMID- 11099005 TI - Political uncertainty halts bioprospecting in Mexico. PMID- 11099006 TI - Online naming of species opens digital age for taxonomy. PMID- 11099007 TI - Election impasse leaves science in the dark. PMID- 11099008 TI - Climate talks face uncertainty over US strategy. PMID- 11099009 TI - Row over fate of endangered monkeys. PMID- 11099010 TI - Fake finds reveal critical deficiency. PMID- 11099011 TI - Promise of Higgs fails to save CERN collider. PMID- 11099012 TI - Medical institute opens amid hopes of Kansas rebirth. PMID- 11099013 TI - Bacterial biofilms and infections. Slimebusters. PMID- 11099014 TI - Why are AIDS dissidents still making 15-year-old, long-refuted claims? PMID- 11099015 TI - Mildest organochlorines still cause toxic pollution. PMID- 11099016 TI - Ancestors knew how to harness horsepower... PMID- 11099017 TI - So animals could pull their weight, and more. PMID- 11099018 TI - Fraud: retracted articles are still being cited. PMID- 11099019 TI - Award organizers should have noted the paper. PMID- 11099021 TI - At the zoo. PMID- 11099022 TI - Archaeoastronomy. Plotting the pyramids. PMID- 11099020 TI - Nice work--but is it science? PMID- 11099023 TI - Fish found in flagrante delicto. PMID- 11099024 TI - Nanothermodynamics. Breathing life into an old model. PMID- 11099025 TI - Global change. It's not a gas. PMID- 11099026 TI - Mitochondrial genes on the move. PMID- 11099027 TI - Real brains for real robots. PMID- 11099028 TI - Surfing the p53 network. PMID- 11099029 TI - Tracing the geographical origin of cocaine. PMID- 11099030 TI - Detecting milk proteins in ancient pots. PMID- 11099031 TI - Feedback control of intercellular signalling in development. AB - The intercellular communication that regulates cell fate during animal development must be precisely controlled to avoid dangerous errors. How is this achieved? Recent work has highlighted the importance of positive and negative feedback loops in the dynamic regulation of developmental signalling. These feedback interactions can impart precision, robustness and versatility to intercellular signals. Feedback failure can cause disease. PMID- 11099032 TI - Ancient Egyptian chronology and the astronomical orientation of pyramids. AB - The ancient Egyptian pyramids at Giza have never been accurately dated, although we know that they were built approximately around the middle of the third millennium BC. The chronologies of this period have been reconstructed from surviving lists of kings and the lengths of their reigns, but the lists are rare, seldom complete and contain known inconsistencies and errors. As a result, the existing chronologies for that period (the Old Kingdom) can be considered accurate only to about +/-100 years, a figure that radiocarbon dating cannot at present improve. Here I use trends in the orientation of Old Kingdom pyramids to demonstrate that the Egyptians aligned them to north by using the simultaneous transit of two circumpolar stars. Modelling the precession of these stars yields a date for the start of construction of the Great Pyramid that is accurate to +/ 5 yr, thereby providing an anchor for the Old Kingdom chronologies. PMID- 11099033 TI - Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. AB - Complete genomic sequence is known for two multicellular eukaryotes, the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster, and it will soon be known for humans. However, biological function has been assigned to only a small proportion of the predicted genes in any animal. Here we have used RNA mediated interference (RNAi) to target nearly 90% of predicted genes on C. elegans chromosome I by feeding worms with bacteria that express double-stranded RNA. We have assigned function to 13.9% of the genes analysed, increasing the number of sequenced genes with known phenotypes on chromosome I from 70 to 378. Although most genes with sterile or embryonic lethal RNAi phenotypes are involved in basal cell metabolism, many genes giving post-embryonic phenotypes have conserved sequences but unknown function. In addition, conserved genes are significantly more likely to have an RNAi phenotype than are genes with no conservation. We have constructed a reusable library of bacterial clones that will permit unlimited RNAi screens in the future; this should help develop a more complete view of the relationships between the genome, gene function and the environment. PMID- 11099034 TI - Functional genomic analysis of cell division in C. elegans using RNAi of genes on chromosome III. AB - Genome sequencing projects generate a wealth of information; however, the ultimate goal of such projects is to accelerate the identification of the biological function of genes. This creates a need for comprehensive studies to fill the gap between sequence and function. Here we report the results of a functional genomic screen to identify genes required for cell division in Caenorhabditis elegans. We inhibited the expression of approximately 96% of the approximately 2,300 predicted open reading frames on chromosome III using RNA mediated interference (RNAi). By using an in vivo time-lapse differential interference contrast microscopy assay, we identified 133 genes (approximately 6%) necessary for distinct cellular processes in early embryos. Our results indicate that these genes represent most of the genes on chromosome III that are required for proper cell division in C. elegans embryos. The complete data set, including sample time-lapse recordings, has been deposited in an open access database. We found that approximately 47% of the genes associated with a differential interference contrast phenotype have clear orthologues in other eukaryotes, indicating that this screen provides putative gene functions for other species as well. PMID- 11099035 TI - Mean-field cluster model for the critical behaviour of ferromagnets. AB - Two separate theories are often used to characterize the paramagnetic properties of ferromagnetic materials. At temperatures T well above the Curie temperature, Tc (where the transition from paramagnetic to ferromagnetic behaviour occurs), classical mean-field theory yields the Curie-Weiss law for the magnetic susceptibility: X(T) infinity 1/(T - Weiss constant), where Weiss constant is the Weiss constant. Close to Tc, however, the standard mean-field approach breaks down so that better agreement with experimental data is provided by critical scaling theory: X(T) infinity 1/(T - Tc)gamma, where gamma is a scaling exponent. But there is no known model capable of predicting the measured values of gamma nor its variation among different substances. Here I use a mean-field cluster model based on finite-size thermostatistics to extend the range of mean-field theory, thereby eliminating the need for a separate scaling regime. The mean field approximation is justified by using a kinetic-energy term to maintain the microcanonical ensembles. The model reproduces the Curie-Weiss law at high temperatures, but the classical Weiss transition at Tc = Weiss constant is suppressed by finite-size effects. Instead, the fraction of clusters with a specific amount of order diverges at Tc, yielding a transition that is mathematically similar to Bose-Einstein condensation. At all temperatures above Tc, the model matches the measured magnetic susceptibilities of crystalline EuO, Gd, Co and Ni, thus providing a unified picture for both the critical-scaling and Curie-Weiss regimes. PMID- 11099036 TI - Universal quantum computation with the exchange interaction. AB - Various physical implementations of quantum computers are being investigated, although the requirements that must be met to make such devices a reality in the laboratory at present involve capabilities well beyond the state of the art. Recent solid-state approaches have used quantum dots, donor-atom nuclear spins or electron spins; in these architectures, the basic two-qubit quantum gate is generated by a tunable exchange interaction between spins (a Heisenberg interaction), whereas the one-qubit gates require control over a local magnetic field. Compared to the Heisenberg operation, the one-qubit operations are significantly slower, requiring substantially greater materials and device complexity--potentially contributing to a detrimental increase in the decoherence rate. Here we introduced an explicit scheme in which the Heisenberg interaction alone suffices to implement exactly any quantum computer circuit. This capability comes at a price of a factor of three in additional qubits, and about a factor of ten in additional two-qubit operations. Even at this cost, the ability to eliminate the complexity of one-qubit operations should accelerate progress towards solid-state implementations of quantum computation. PMID- 11099037 TI - Kondo physics in carbon nanotubes. AB - The connection of electrical leads to wire-like molecules is a logical step in the development of molecular electronics, but also allows studies of fundamental physics. For example, metallic carbon nanotubes are quantum wires that have been found to act as one-dimensional quantum dots, Luttinger liquids, proximity induced superconductors and ballistic and diffusive one-dimensional metals. Here we report that electrically contacted single-walled carbon nanotubes can serve as powerful probes of Kondo physics, demonstrating the universality of the Kondo effect. Arising in the prototypical case from the interaction between a localized impurity magnetic moment and delocalized electrons in a metallic host, the Kondo effect has been used to explain enhanced low-temperature scattering from magnetic impurities in metals, and also occurs in transport through semiconductor quantum dots. The far greater tunability of dots (in our case, nanotubes) compared with atomic impurities renders new classes of Kondo-like effects accessible. Our nanotube devices differ from previous systems in which Kondo effects have been observed, in that they are one-dimensional quantum dots with three-dimensional metal (gold) reservoirs. This allows us to observe Kondo resonances for very large electron numbers (N) in the dot, and approaching the unitary limit (where the transmission reaches its maximum possible value). Moreover, we detect a previously unobserved Kondo effect, occurring for even values of N in a magnetic field. PMID- 11099038 TI - Increased marine production of N2O due to intensifying anoxia on the Indian continental shelf. AB - Eutrophication of surface waters and hypoxia in bottom waters has been increasing in many coastal areas, leading to very large depletions of marine life in the affected regions. These areas of high surface productivity and low bottom-water oxygen concentration are caused by increasing runoff of nutrients from land. Although the local ecological and socio-economic effects have received much attention, the potential contribution of increasing hypoxia to global-change phenomena is unknown. Here we report the intensification of one of the largest low-oxygen zones in the ocean, which develops naturally over the western Indian continental shelf during late summer and autumn. We also report the highest accumulations yet observed of hydrogen sulphide (H2S) and nitrous oxide (N2O) in open coastal waters. Increased N2O production is probably caused by the addition of anthropogenic nitrate and its subsequent denitrification, which is favoured by hypoxic conditions. We suggest that a global expansion of hypoxic zones may lead to an increase in marine production and emission of N2O, which, as a potent greenhouse gas, could contribute significantly to the accumulation of radiatively active trace gases in the atmosphere. PMID- 11099039 TI - Microseismological evidence for a changing wave climate in the northeast Atlantic Ocean. AB - One possible consequence of a change in climate over the past several decades is an increase in wave heights, potentially threatening coastal areas as well as the marine industry. But the difficulties in observing wave heights exacerbates a general problem of climate-change detection: inhomogeneities in long-term observational records owing to changes in the instruments or techniques used, which may cause artificial trends. Ground movements with periods of 4-16 seconds, known as microseisms, are associated with ocean waves and coastal surf, and have been recorded continuously since the early days of seismology. Here we use such a 40-year record of wintertime microseisms from Hamburg, Germany, to reconstruct the wave climate in the northeast Atlantic Ocean. For the period 1954-77, we detect an average of seven days per month with strong microseismic activity, without a significant trend. This number increases significantly in the second half of the record, reaching approximately 14 days of strong microseisms per month. The implied increase in northeast Atlantic wave height over the past 20 years parallels increased surface air temperatures and storminess in this region, suggesting a common forcing. PMID- 11099040 TI - Fine-scale genetic structuring on Manacus manacus leks. AB - Leks have traditionally been considered as arenas where males compete to attract females and secure matings. Thus, direct fitness benefits mediated through competition between males to fertilize females have been considered to be the primary force driving the evolution of lekking behaviour. Inclusive fitness benefits mediated through kin selection may also be involved in lek formation and evolution, but to date this theory has been largely ignored. According to kin selection theory, both reproducing and non-reproducing males may gain indirect inclusive fitness benefits. If females are attracted to larger leks, non reproducing males add attractiveness to a lek, and therefore, in a genetically structured population, boost the reproductive success of kin. Theory predicts that the attractiveness of leks is plastic, and that males establish themselves on a lek in which the top male, in terms of reproductive success, is a close relative. Here we show that in white-bearded manakins (Manacus manacus), for which larger leks are more attractive to females and so secure the maximum number of matings, there is extraordinary fine-scale genetic structure, with leks being composed of clusters of related kin. We propose that males establish themselves where they find relatives to such an extent that they form groups within leks, and that such behaviour is consistent with kin-selection theory to maximize reproductive success of the group. PMID- 11099041 TI - Repeated, recent and diverse transfers of a mitochondrial gene to the nucleus in flowering plants. AB - A central component of the endosymbiotic theory for the bacterial origin of the mitochondrion is that many of its genes were transferred to the nucleus. Most of this transfer occurred early in mitochondrial evolution; functional transfer of mitochondrial genes has ceased in animals. Although mitochondrial gene transfer continues to occur in plants, no comprehensive study of the frequency and timing of transfers during plant evolution has been conducted. Here we report frequent loss (26 times) and transfer to the nucleus of the mitochondrial gene rps10 among 277 diverse angiosperms. Characterization of nuclear rps10 genes from 16 out of 26 loss lineages implies that many independent, RNA-mediated rps10 transfers occurred during recent angiosperm evolution; each of the genes may represent a separate functional gene transfer. Thus, rps10 has been transferred to the nucleus at a surprisingly high rate during angiosperm evolution. The structures of several nuclear rps10 genes reveal diverse mechanisms by which transferred genes become activated, including parasitism of pre-existing nuclear genes for mitochondrial or cytoplasmic proteins, and activation without gain of a mitochondrial targeting sequence. PMID- 11099042 TI - Imagery neurons in the human brain. AB - Vivid visual images can be voluntarily generated in our minds in the absence of simultaneous visual input. While trying to count the number of flowers in Van Gogh's Sunflowers, understanding a description or recalling a path, subjects report forming an image in their "mind's eye". Whether this process is accomplished by the same neuronal mechanisms as visual perception has long been a matter of debate. Evidence from functional imaging, psychophysics, neurological studies and monkey electrophysiology suggests a common process, yet there are patients with deficits in one but not the other. Here we directly investigated the neuronal substrates of visual recall by recording from single neurons in the human medial temporal lobe while the subjects were asked to imagine previously viewed images. We found single neurons in the hippocampus, amygdala, entorhinal cortex and parahippocampal gyrus that selectively altered their firing rates depending on the stimulus the subjects were imagining. Of the neurons that fired selectively during both vision and imagery, the majority (88%) had identical selectivity. Our study reveals single neuron correlates of volitional visual imagery in humans and suggests a common substrate for the processing of incoming visual information and visual recall. PMID- 11099043 TI - Real-time prediction of hand trajectory by ensembles of cortical neurons in primates. AB - Signals derived from the rat motor cortex can be used for controlling one dimensional movements of a robot arm. It remains unknown, however, whether real time processing of cortical signals can be employed to reproduce, in a robotic device, the kind of complex arm movements used by primates to reach objects in space. Here we recorded the simultaneous activity of large populations of neurons, distributed in the premotor, primary motor and posterior parietal cortical areas, as non-human primates performed two distinct motor tasks. Accurate real-time predictions of one- and three-dimensional arm movement trajectories were obtained by applying both linear and nonlinear algorithms to cortical neuronal ensemble activity recorded from each animal. In addition, cortically derived signals were successfully used for real-time control of robotic devices, both locally and through the Internet. These results suggest that long-term control of complex prosthetic robot arm movements can be achieved by simple real-time transformations of neuronal population signals derived from multiple cortical areas in primates. PMID- 11099044 TI - A regulator of transcriptional elongation controls vertebrate neuronal development. AB - The development of distinct vertebrate neurons is defined by the unique profiles of genes that neurons express. It is accepted that neural genes are regulated at the point of transcription initiation, but the role of messenger RNA elongation in neural gene regulation has not been examined. Here we describe the mutant foggy, identified in a genetic screen for mutations that affect neuronal development in zebrafish, that displayed a reduction of dopamine-containing neurons and a corresponding surplus of serotonin-containing neurons in the hypothalamus. Positional cloning disclosed that Foggy is a brain-enriched nuclear protein that is structurally related to the transcription elongation factor Spt5 (refs 5-12). Foggy is not part of the basic transcription apparatus but a phosphorylation-dependent, dual regulator of transcription elongation. The mutation disrupts its repressive but not its stimulatory activity. Our results provide molecular, genetic and biochemical evidence that negative regulators of transcription elongation control key aspects of neuronal development. PMID- 11099045 TI - ClC-5 Cl- -channel disruption impairs endocytosis in a mouse model for Dent's disease. AB - Dent's disease is an X-linked disorder associated with the urinary loss of low molecular-weight proteins, phosphate and calcium, which often leads to kidney stones. It is caused by mutations in ClC-5, a renal chloride channel that is expressed in endosomes of the proximal tubule. Here we show that disruption of the mouse clcn5 gene causes proteinuria by strongly reducing apical proximal tubular endocytosis. Both receptor-mediated and fluid-phase endocytosis are affected, and the internalization of the apical transporters NaPi-2 and NHE3 is slowed. At steady state, however, both proteins are redistributed from the plasma membrane to intracellular vesicles. This may be caused by an increased stimulation of luminal parathyroid hormone (PTH) receptors owing to the observed decreased tubular endocytosis of PTH. The rise in luminal PTH concentration should also stimulate the hydroxylation of 25(OH) vitamin D3 to the active hormone. However, this is counteracted by a urinary loss of the precursor 25(OH) vitamin D3. The balance between these opposing effects, both of which are secondary to the defect in proximal tubular endocytosis, probably determines whether there will be hypercalciuria and kidney stones. PMID- 11099046 TI - The Eps8 protein coordinates EGF receptor signalling through Rac and trafficking through Rab5. AB - How epidermal growth factor receptor (EGFR) signalling is linked to EGFR trafficking is largely unknown. Signalling and trafficking involve small GTPases of the Rho and Rab families, respectively. But it remains unknown whether the signalling relying on these two classes of GTPases is integrated, and, if it is, what molecular machinery is involved. Here we report that the protein Eps8 connects these signalling pathways. Eps8 is a substrate of the EGFR, which is held in a complex with Sos1 by the adaptor protein E3bl (ref. 2), thereby mediating activation of Rac. Through its src homology-3 domain, Eps8 interacts with RN-tre. We show that RN-tre is a Rab5 GTPase-activating protein, whose activity is regulated by the EGFR. By entering in a complex with Eps8, RN-tre acts on Rab5 and inhibits internalization of the EGFR. Furthermore, RN-tre diverts Eps8 from its Rac-activating function, resulting in the attenuation of Rac signalling. Thus, depending on its state of association with E3b1 or RN-tre, Eps8 participates in both EGFR signalling through Rac, and trafficking through Rab5. PMID- 11099047 TI - Deacetylation of p53 modulates its effect on cell growth and apoptosis. AB - The p53 tumour suppressor is a transcriptional factor whose activity is modulated by protein stability and post-translational modifications including acetylation. The mechanism by which acetylated p53 is maintained in vivo remains unclear. Here we show that the deacetylation of p53 is mediated by an histone deacetylase-1 (HDAC1)-containing complex. We have also purified a p53 target protein in the deacetylase complexes (designated PID; but identical to metastasis-associated protein 2 (MTA2)), which has been identified as a component of the NuRD complex. PID specifically interacts with p53 both in vitro and in vivo, and its expression reduces significantly the steady-state levels of acetylated p53. PID expression strongly represses p53-dependent transcriptional activation, and, notably, it modulates p53-mediated cell growth arrest and apoptosis. These results show that deacetylation and functional interactions by the PID/MTA2-associated NuRD complex may represent an important pathway to regulate p53 function. PMID- 11099048 TI - Insights into SCF ubiquitin ligases from the structure of the Skp1-Skp2 complex. AB - F-box proteins are members of a large family that regulates the cell cycle, the immune response, signalling cascades and developmental programmes by targeting proteins, such as cyclins, cyclin-dependent kinase inhibitors, IkappaBalpha and beta-catenin, for ubiquitination (reviewed in refs 1-3). F-box proteins are the substrate-recognition components of SCF (Skp1-Cullin-F-box protein) ubiquitin protein ligases. They bind the SCF constant catalytic core by means of the F-box motif interacting with Skp1, and they bind substrates through their variable protein-protein interaction domains. The large number of F-box proteins is thought to allow ubiquitination of numerous, diverse substrates. Most organisms have several Skp1 family members, but the function of these Skp1 homologues and the rules of recognition between different F-box and Skp1 proteins remain unknown. Here we describe the crystal structure of the human F-box protein Skp2 bound to Skp1. Skp1 recruits the F-box protein through a bipartite interface involving both the F-box and the substrate-recognition domain. The structure raises the possibility that different Skp1 family members evolved to function with different subsets of F-box proteins, and suggests that the F-box protein may not only recruit substrate, but may also position it optimally for the ubiquitination reaction. PMID- 11099049 TI - The production of pyrethrins by plant cell and tissue cultures of Chrysanthemum cinerariaefolium and Tagetes species. AB - Pyrethrins, the most economically important natural insecticide, comprise a group of six closely related monoterpene esters. The industrial production is based on their extraction from Chrysanthemum cinerariaefolium (Pyrethrum) capitula. The world production of natural pyrethrins still falls short of global market demand stimulating the research in in vitro production as an alternative to conventional cultivation methods. The different biotechnological alternatives such as callus cultures, shoot and root cultures, plant cell suspension cultures, and bioconversion of precursors by means of enzymatic synthesis or genetically engineered microorganisms, as well as the progress achieved in methods for the identification and quantitation of insecticidal compounds have been reviewed. Although technology for plant cell culture exists, industrial applications have, to date, been limited due to both the low economical viability and technological feasibility at large scale. Bioconversion of readily available precursors looks more attractive, but more research is needed before this technology is used for the industrial production of pyrethrins. PMID- 11099050 TI - Computer-intensive statistical procedures. PMID- 11099051 TI - Beyond eyeballing: fitting models to experimental data. PMID- 11099052 TI - Nutritional support in pancreatitis. PMID- 11099053 TI - A multicenter, randomized, double-blind clinical trial examining the effect of oral human recombinant epidermal growth factor on the healing of duodenal ulcers. AB - BACKGROUND: Our aim was to study the efficacy of oral human recombinant epidermal growth factor (EGF) in the treatment of duodenal ulcers, on the basis of its repairing actions in the gastrointestinal tract. METHODS: A placebo-controlled, multicenter, randomized, and double-blind study was conducted. Treatment groups were A) placebo solution, B) 10 microg/ml of human recombinant (hr)-EGF, and C) 50 microg/ml of hr-EGF, three times daily during 6 weeks. Patients, 15-65 years old, with a duodenal ulcer >4 mm, who gave their written informed consent to participate were eligible. Exclusion criteria were gastric ulcer and more than one duodenal ulcer, ulcer-related complications, and previous treatment with oral EGF or other specific anti-ulcer drugs in the previous 2 weeks. The main outcome variable was ulcer healing, evaluated by endoscopy after the 2nd, 4th, and 6th week. RESULTS: One hundred and three patients were included. The groups were comparable with regard to age, sex, toxic habits, antecedents of ulcerous disease, initial size and depth or the ulcer, initial symptoms, and positivity for Helicobacter pylori. The ulcers were healed in a larger proportion of patients treated with hr-EGF at the highest dose (70.6% in group C versus 40.0% and 35.3% in placebo and low-dose groups, respectively (P = 0.007)). The difference was significant from week 4 on. Groups A and B did not differ. Eighty eight percent of group C patients were cured or improved versus 57% and 56% in groups A and B, respectively. No adverse reactions were reported. CONCLUSIONS: Oral hr-EGF was effective in the treatment of duodenal ulcer at a 50-microg/ml dose every 8 h but not at 10 microg/ml. PMID- 11099054 TI - Eradication of Helicobacter pylori compared with long-term acid suppression in duodenal ulcer disease. A randomized trial with 2-year follow-up. The Danish Ulcer Study Group. AB - BACKGROUND: Trials evaluating long-term management of duodenal ulcer disease have mainly been focused on recurrence of ulcers, disregarding effects on dyspeptic and reflux symptoms. Profound acid inhibition with a proton pump inhibitor is the gold standard therapy in acid-related diseases. We aimed to compare the symptomatic effects of eradication therapy with those of long-term omeprazole treatment in a design with periods both with and without acid inhibition. METHODS: Patients with active duodenal ulcer were randomized either to omeprazole, 20 mg twice daily until healing, followed by omeprazole, 20 mg/ day for 1 year, or to eradication therapy (metronidazole, amoxicillin, and omeprazole for 2 weeks) followed by placebo for 1 year. All patients were followed up passively for an additional year. Clinical controls were performed every 2 months the 1st year (maintenance phase) and every 6 months during the passive follow-up phase. The study was multicentric and double-blind. The primary end-point was discontinuation of treatment, irrespective of reason. RESULTS: Two hundred and seventy-six patients were randomized (139 in the eradication treatment group). In the maintenance phase there were no differences in the reporting of dyspeptic symptoms or in premature withdrawal. In the passive follow-up phase only five patients in the eradication therapy group discontinued owing to relapse of dyspeptic symptoms or ulcer, compared with 51 patients initially randomized to long-term omeprazole. There were no differences in reflux symptoms or in the development of reflux oesophagitis. CONCLUSIONS: Eradication therapy and long term omeprazole are equally effective in controlling dyspeptic symptoms and reflux in duodenal ulcer patients with healed ulcers. One-quarter of the duodenal ulcer patients who start eradication therapy continue to be symptomatic or fail therapy for other reasons over a 2-year period. Eradication therapy does not increase the risk of reflux in ulcer patients. PMID- 11099055 TI - Different patterns of Helicobacter pylori adherence to gastric mucosa cells in children and adults. An ultrastructural study. AB - BACKGROUND: Infection with Helicobacter pylori in childhood may be the initiation of a lifelong coexistence between microorganisms and epithelial cells resulting in chronic inflammation. The adhesion pattern of H. pylori found in antral biopsies from a group of H. pylori-infected children with recurrent abdominal pain was compared with a group of H. pylori-infected adults suffering from dyspepsia, in an attempt to reveal differences in the type of adhesion. METHODS: The histology of antrum biopsies and the ultrastructure of adherent H. pylori in biopsies from 26 children (median age, 10.1 years) were compared with organisms in biopsies from 19 adults (median age, 54.4 years). RESULTS: More than 1000 adherent H. pylori were studied and divided into four types of adhesion: 1) contact to microvilli; 2) connection to the plasma membrane via filamentous material; 3) adhesive pedestal formation; and 4) abutting or making a depression in the plasma membrane. Contact to microvilli was significantly higher (69% versus 39%; P = 0.002) in children compared with adults and comprised two-thirds of all adherent organisms in children. The more intimate adhesion types as abutting or adhesive pedestals dominated in adults. CONCLUSIONS: These results indicate a change in contact types between H. pylori and gastric epithelial cells in adults compared with children and this may be a natural development in the lifelong infection of humans. PMID- 11099056 TI - Effects of age on proximal gastric motor and sensory function. AB - BACKGROUND: Healthy aging is associated with a reduction in appetite and food intake, which may predispose to pathologic weight loss and malnutrition. Changes in intragastric mechanisms mediating satiation in the elderly have not been studied. The aim of this study was to evaluate the effects of aging on i) fasting gastric compliance and the perception of gastric distension, and ii) food intake and gastric accommodation to a meal. METHODS: Five healthy older (aged 68-73 years) and five healthy young (aged 22-27 years) men, matched for body mass index, were each studied on three occasions after an overnight fast. On one day ('barostat day'), isovolumetric and isobaric distensions of the proximal stomach were performed, and meal-induced changes in intrabag volume were measured with an electronic barostat. On another day ('tube-only day') subjects were intubated with a nasogastric tube without an intragastric bag before the meal. On the 3rd day (control day) subjects were given the meal without intubation. Energy intake from the buffet meal was quantified, and perceptions assessed using visual analogue questionnaires. RESULTS: During both isobaric and isovolumetric distensions the pressure-volume relationship did not differ significantly between older and young subjects. During gastric distensions perceptions of fullness (P < 0.01), abdominal discomfort (P < 0.05), and bloating (P < 0.05) were less in older than young subjects, whereas the perception of hunger (P < 0.05) was less in the young than in older subjects. There was no difference in energy intake (P = 0.44) between young and older subjects. Food intake was less on the barostat day (P < 0.01) and the tube-only day (P < 0.01) than on the control day in young subjects but was not affected by the different study conditions in the older subjects. After the meal the maximum intrabag volume occurred later in the older than in the young subjects (105 +/- 4 min versus 36 +/- 8 min; P < 0.05), and the intrabag volume change was greater (P = 0.05) in the older than the young subjects later in the postprandial period. CONCLUSIONS: Healthy aging is associated with decreased perception of gastric distension without any change in fasting gastric compliance and with reduced gastric tone late in the postprandial period when compared with the young. Control of food intake is less sensitive to external stimuli in older than in young subjects. PMID- 11099057 TI - Fructose- and sorbitol-reduced diet improves mood and gastrointestinal disturbances in fructose malabsorbers. AB - BACKGROUND: Fructose malabsorption is characterized by the inability to absorb fructose efficiently. As a consequence fructose reaches the colon where it is broken down by bacteria to short fatty acids, CO2 and H2. Bloating, cramps, osmotic diarrhea and other symptoms of irritable bowel syndrome are the consequences and can be seen in about 50% of fructose malabsorbers. We have previously shown that fructose malabsorption is associated with early signs of mental depression and low serum tryptophan concentrations. It was therefore of interest whether a fructose-reduced diet could not only improve gastrointestinal complaints but also depressive signs seen in fructose malabsorbers. METHODS: Fifty-three adults (12 males, 41 females), who were identified as fructose malabsorbers according to their breath-H2 concentrations, filled out a Beck's depression inventory-questionnaire, and a questionnaire with arbitrary scales for measurement of meteorism, stool frequency and quality of life for a 4-week period before dietary intervention and 4 weeks after dietary change as for fructose- and sorbitol-reduced diet. RESULTS: Depression scores were reduced by 65.2% after 4 weeks of diet (P < 0.0001), and there was a significant reduction of meteorism (P < 0.0001) and stool frequency (P < 0.01). Improvement of signs of depression and of meteorism was more pronounced in females than in males. CONCLUSION: Fructose- and sorbitol-reduced diet in subjects with fructose malabsorption does not only reduce gastrointestinal symptoms but also improves mood and early signs of depression. PMID- 11099058 TI - Increased permeability in dextran sulphate colitis in rats: time course of development and effect of butyrate. AB - BACKGROUND: Increased mucosal permeability is an important factor in the genesis of mucosal inflammation in inflammatory bowel disease. This study examined the time course of increased permeability and the effect of butyrate on permeability in experimental colitis in rats. METHODS: Colitis was induced in albino rats by administration of 4% dextran sulphate sodium (DSS) orally for up to 7 days. Rats were killed sequentially after 1-7 days of DSS feeding and compared with control animals. Distal colon sheets, from normal and DSS rats, were mounted in Ussing chambers. Electric resistance and passive permeation of 14C-mannitol were measured over 90 min. In control and 5-day DSS rats additional permeability measurements were made in the presence of butyrate (25 mmol/l) in the bathing solutions. The permeability of the normal distal colon was measured after addition of DSS in vitro. Sections of colon were examined by light microscopy. The viability of colonocytes, from normal and DSS rat colon, was measured by release of lactate dehydrogenase immediately and during a 60-min incubation after isolation. RESULTS: Focal mild inflammation and shedding of epithelium were noted after 2 days of DSS administration; crypt loss with flattened epithelium in adjacent areas after 5 days; and fibrosis after 7 days. Decreased epithelial cell survival after 60 min of incubation was noted after 1 day of DSS administration, whereas decreased viability at the time of isolation was noted after 2 days of DSS administration (viability, 72.7% +/- 1.4%; mean +/- standard error) compared with control (89.3% +/- 0.8%) (P < 0.01). Increased permeability was noted after 1 day of DSS administration. Electric resistance (mu omega/cm2/h) was significantly reduced after 1 day of DSS administration to 85.9 +/- 4.6 (mean +/- standard error) compared with control animals (117.2 +/- 2.2; P < 0.001). Serosa mucosa flux of mannitol (micromol/cm2/h) was also significantly increased after 1 day of DSS feeding (0.169 +/- 0.01) compared with control (0.061 +/- 0.08) (P < 0.01). Electric resistance and mannitol permeability were significantly returned towards normal by the presence of butyrate. DSS added directly to the bathing solution did not significantly alter the colon permeability in vitro. CONCLUSIONS: Increased mucosal permeability is a very early change in colitis induced by DSS, is accompanied by decreased cell survival, and precedes detectable changes in histology. Reversal of increased mucosal permeability by butyrate may explain its utility in the therapy of inflammatory disease of the colon. PMID- 11099059 TI - The dietary combination of germinated barley foodstuff plus Clostridium butyricum suppresses the dextran sulfate sodium-induced experimental colitis in rats. AB - BACKGROUND: Recent studies have suggested that dietary fiber exerts a therapeutic effect on IBD patients. The aim of this study was to evaluate the effects of a dietary combination of germinated barley foodstuff (GBF), derived from the aleurone and scutellum fraction of germinated barley, plus Clostridium butyricum against dextran sulfate sodium (DSS)-induced experimental colitis in rats. METHODS: Sprague-Dawley rats were fed a 3% DSS diet containing GBF only, GBF plus C. butyricum, cellulose only (control) or cellulose plus C. butyricum for 8 days. The mucosal damage (macroscopic and microscopic inflammation) and fecal short chain fatty acid (SCFA) levels were then determined. RESULTS: The combination of GBF plus C. butyricum most effectively prevented bloody diarrhea and mucosal damage. The GBF-only diet also showed some preventive effects. With respect to fecal SCFAs, the combination of GBF plus C. butyricum most effectively increased the fecal SCFA level. CONCLUSION: The dietary combination of GBF plus C. butyricum most effectively suppressed DSS-induced experimental colitis in rats. These effects may be closely associated with its high activity to increase SCFA levels in the gut lumen. The potential clinical efficacy of GBF in IBD patients is also discussed. PMID- 11099060 TI - Quality of life study in a regional group of patients with Crohn disease. A structured interview study. AB - BACKGROUND: The course and prognosis of Crohn disease has previously been described in a regional group of patients in Copenhagen County. The aim of the present study was to reveal the quality of life. as judged by the patients, and compared to age- and sex-matched healthy controls. METHODS: Out of 100 consecutive out-patients with Crohn disease, 94 patients accepted to participate together with 94 age- and sex-matched healthy controls. A modified McMaster Inflammatory Bowel Disease Questionnaire (IBDQ23) was used, excluding bowel related questions. Medical students conducted interviews without knowing who were Crohn disease patients and who were controls. The bowel-related questions and Crohn's Disease Activity Index (CDAI) were assessed by gastroenterologists at inclusion in the study. Responses were indicated on a seven-point scale (7 best/1 worst). Mean numeric score was calculated as well as a delta score, i.e. the difference in score between a patient and the matched control. RESULTS: In 21 of 23 questions the median delta score was zero, indicating no difference between patient and control. The median total delta score was 0.4 in favour of healthy controls (P < 0.001), and significantly higher in patients in relapse, 0.9, than in patients in remission, 0.3 (P < 0.01). The median total numeric score was 5.7 for patients and 6.1 for controls. CONCLUSIONS: Although patients with Crohn disease scored significantly lower on the quality of life scale than matched healthy controls, the differences were smaller than could be expected, taking the chronic disease into consideration. Disease activity correlated with the quality of life score. PMID- 11099061 TI - Concordance of inflammatory bowel disease among Danish twins. Results of a nationwide study. AB - BACKGROUND: Previous studies have shown an increased risk of inflammatory bowel disease (IBD) among relatives of patients with Crohn disease and ulcerative colitis. In the present study the probandwise concordance rates for ulcerative colitis and Crohn disease among mono- and dizygotic twins were estimated. Further we aimed to evaluate whether smoking habits might influence the concordance, and to look for clinical characteristics of concordant versus discordant twin pairs. METHODS: Among the 38,507 identified twins born in Denmark from 1953 to 1982, a questionnaire was sent to the 34,076 who previously had accepted to participate in studies. For twins reporting IBD, the diagnosis was verified by applying standard criteria to records requested from hospitals or practitioners. RESULTS: Among the 29,421 (86.3%) twins answering the questionnaire, 103 pairs had at least one twin who suffered from IBD. In the Crohn disease group five of 10 monozygotic pairs, but none of 27 dizygotic pairs were concordant. In the ulcerative colitis group three of 21 monozygotic, and two of 44 dizygotic pairs were concordant. The probandwise concordance rate among monozygotic pairs was 58.3% for Crohn disease and 18.2% for ulcerative colitis; among the dizygotic pairs the rates were 0 and 4.5%, respectively. The frequency of smokers was higher among twins with Crohn disease and lower among twins with ulcerative colitis compared to the frequency in the twin register. Furthermore, smoking habits were found to be of significance for discordance for disease. Regarding the clinical characteristics no homogenous pattern was observed within the concordant pairs and the differences between concordant and discordant pairs were not significant. CONCLUSION: The observation of a significantly higher concordance rate among monozygotic than among dizygotic twin pairs strongly points to a genetic influence on occurrence of IBD, which seems to be more pronounced with regard to Crohn disease than to ulcerative colitis. Differences in smoking habits among the members of the discordant twin pairs may influence the discordance. PMID- 11099062 TI - The epidemiology of oesophageal adenocarcinoma: has the cancer of gastric cardia an influence on the rising incidence of oesophageal adenocarcinoma? AB - BACKGROUND: Owing to overgrowth and definitional problems in classification, the cancer of gastric cardia may affect significantly the epidemiological analysis of oesophageal adenocarcinoma. The purpose of the present study was to evaluate the changes in the incidence of all the adenocarcinomas near the gastrooesophageal junction. METHODS: Trends in the incidence rates of adenocarcinoma of the oesophagus and the gastric cardia were described through the Finnish Cancer Registry. The annual age-standardized incidence rates during 1976-95 were analysed by a linear regression technique. RESULTS: The total incidence of oesophageal carcinoma remained around 3.5/100,000 in men, and decreased from 2.8 to 1.3/100,000 in women. The incidence of oesophageal adenocarcinoma increased from 0.28 to 0.77/100,000 (nearly 300%) in males, and from 0.08 to 0.11 per 100,000 in females. There were no significant changes with time in the incidence rate of gastric cardia cancer in either sex. Combined gastric cardia and oesophageal adenocarcinoma incidence rates remained stable in women, and increased slightly, but significantly, from 2.4 to 2.9/100,000 in men. CONCLUSION: Oesophageal adenocarcinoma has increased significantly in men in Finland, but the combined incidence of cancers of the gastro-oesophageal junction has increased only slightly. To overcome the difficulties in classification of oesophageal adenocarcinoma and the cancer of gastric cardia in the epidemiological studies, the focus should be on all adenocarcinomas at or near the oesophagogastric junction. PMID- 11099063 TI - N-acetyltransferase 2 (NAT2) genotype and colorectal carcinoma: risk variability according to tumour site? AB - BACKGROUND: Dietary heterocyclic aromatic amines (HAAs) are members of a family of chemicals that comprise highly mutagenic compounds related to colon cancer. The polymorphic N-acetyltransferase 2 enzyme (NAT2, E.C. 2.3.1.5) plays a key role in the transformation of HAAs to ultimate carcinogens. NAT2 enzyme activity is expressed in a genotype-dependent manner in colon epithelium. Therefore local activation of HAAs in colon, and hence increased risk to develop colon cancer, is likely to be related to high NAT2 enzyme activity. This study is aimed at analysing the association between genotypes leading to high NAT2 activity and colorectal cancer risk. METHODS: Genomic DNA from 120 colorectal cancer patients and 258 healthy individuals were analysed for enzyme-inactivating mutations at the coding region of the NAT2 gene by means of a mutation-specific polymerase chain reaction. RESULTS: Among patients with sigmoid colon cancer, a significant excess of individuals with genotypes leading to high NAT2 activity was observed as compared both to controls and to the rest of patients with colorectal cancer (P < 0.05). CONCLUSIONS: Our findings, which require independent confirmation, suggest that the NAT2 genotype constitutes a secondary risk factor to develop sigmoid colon cancer. PMID- 11099064 TI - C5b-9 and interleukin-6 in chronic hepatitis C. Surrogate markers predicting short-term response to interferon alpha-2b. AB - BACKGROUND: Available data and our observations suggest that elevated levels of interleukin (IL)-6 and -10 and some complement parameters may be associated with a poor response to IFN alpha. We evaluated how baseline levels of C5b-9, IL-6, and IL-10 influence the outcome of IFN alpha treatment. METHODS: Fifty-one patients with established chronic hepatitis C were enrolled and treated with IFN alpha-2b. Before and after a 12-week-IFN-treatment (3 MU or 5 MU tiw) serum levels of IL-6, IL-10, C5b-9 and RNA of hepatitis C virus (HCV) were assessed. Sera of 46 sex- and age-matched, healthy blood donors served as control. RESULTS: While two-thirds of patients was considered 'responder', 14 patients had no significant decrease either in HCV RNA or in ALT levels. In the responder's group lower baseline levels of IL-6 and C5b-9 were found than those in the 'non responder' group. As a result of IFN therapy HCV RNA and C5b-9 levels significantly decreased. While the serum concentration of IL-6 increased during the follow-up period, regarding IL-10, no change was observed. In patients with 'low' baseline levels of C5b-9 (<2053 ng/ml) IFN alpha resulted in a significantly (P = 0.0005) higher decrease in HCV RNA level. Regarding 'low' IL-6 values (< 1.47 pg/ml) similar but somewhat less significant (P = 0.0039) difference was found if the change of HCV RNA was investigated. The odds ratio of patients with low IL-6 and/or C5b-9 to responding to IFN alpha treatment was almost 10 times (CI: 9.1 (1.8-50.9)) higher as compared with patients without 'low' levels of these parameters. CONCLUSION: Our data suggest that serum level(s) of IL-6 and/or C5b-9 taken prior to the initiation of IFN treatment may serve as surrogate marker(s) in evaluating patients with chronic hepatitis C whether to get IFN alpha in monotherapy or to consider having combination therapy in the form of IFN alpha-ribavirin. PMID- 11099065 TI - Role of nitric oxide and endothelin-1 in a portal hypertensive rat model. AB - BACKGROUND: Portal hypertension is often accompanied by a hyperdynamic circulation state. Some reports have suggested that nitric oxide (NO) plays an important role in this hyperdynamic state. On the other hand, although endothelin (ET)-1, a powerful vasoconstrictor, was recently identified, little is known about its role in portal hypertension or about the interaction between NO and ET 1. The aim of this study was therefore to investigate whether or not the inhibitor of NO synthase (NOS) might improve portal hypertension, and also to clarify the relationship between NO and ET-1. METHODS: Portal hypertensive (PHT) rats, in which hypertension was induced by a two-step ligation of the portal vein (PVL), were used. The mean arterial pressure (MAP), portal pressure (PP), visceral blood flow volume (BFV), and serum levels of NO and ET-1 were determined in PVL rats treated with two NOS inhibitors with different functions: N(G)-nitro L-arginine methyl ester (L-NAME) and aminoguanidine (AG). Control (CTR) rats. treated by a sham operation (SO), were also studied. RESULTS: Two-step PVL treatment induced a significant increase in the serum level of NO3-and ET-1 in the portal vein. L-NAME and AG administration significantly decreased PP at doses of 50 mg/kg in PHT rats after 60 min administration, while no inhibitor effected any modification in the CTBR rats. Both NOS inhibitors increased MAP and decreased PP and BFV in the portal vein, gastric mucosa, and spleen, in addition to decreasing the serum levels of NO3- and ET-1 in the PHT rats, while neither blockade modified any parameters in the CTR rats. In PHT rats, L-arginine, a NO substance, reversed the effect of L-NAME, while it did not induce any recovery from the AG effect. CONCLUSIONS: In PHT rats, NO seems to contribute to portal hypertension. PVL increases not only the serum level of NO3-, but also that of ET 1 in the portal vein. Both L-NAME and AG reduce PP and BFV of the portal vein, spleen, gastric mucosa. and liver. In addition, the inhibition of NOS diminishes the serum level not only of NO, but also of ET-1. Use of an appropriate NOS inhibitor may therefore positively affect the hyperdynamic state in portal hypertension. PMID- 11099066 TI - Electrogenic ion transport in duodenum, an aid in cystic fibrosis diagnosis. AB - BACKGROUND: Abnormality in chloride transport across epithelial tissues is a basic defect in cystic fibrosis (CF). Our aim was to compare the induced chloride secretion in duodenum in CF patients with different mutations. METHODS: Duodenal biopsies from 9 patients were investigated in a modified Ussing chamber and the secretory response to prostaglandin E2 (PGE2) and acetylcholine (ACh) were measured. RESULTS: PGE2 and ACh induced no changes in chloride secretion in the AF508 homozygotes. In heterozygotes the induced change in chloride secretion corresponded to the severity of the known mutations. CONCLUSION: The secretory response in duodenum in CF is influenced by the patients genotype and mainly related to sweat chloride secretion. PMID- 11099067 TI - Incidence and mortality of acute pancreatitis between 1985 and 1995. AB - BACKGROUND: The incidence of acute pancreatitis seems to have increased in Western countries. It has been suggested that this increase can be explained by improved diagnostic procedures. We performed a nationwide study to assess the annual sex- and age-specific incidence and mortality rates of acute pancreatitis in the Netherlands between 1985 and 1995, a period in which diagnostic procedures did not change considerably. METHODS: We conducted a population-based retrospective follow-up study in which we used automated hospital discharge data accumulated by Prismant Health Care Information. All patients admitted with acute pancreatitis (ICD-9CM, 577.0) in the Netherlands were identified. We accounted for referrals to other hospitals to avoid double counting and for miscoding of chronic pancreatitis as acute pancreatitis. The annual population size was retrieved from the Netherlands Central Statistics Office. RESULTS: The observed incidence of acute pancreatitis increased from 12.4/100,000 person-years (95% confidence interval (CI), 11.8-12.9) in 1985 to 15.9/100,000 person-years (95% CI, 15.3-16.5) in 1995. The annual mortality rate of acute pancreatitis remained fairly stable at 1.5/100,000 person-years. The incidence and mortality rate of acute pancreatitis increased considerably with age. The case-fatality proportion of first admissions for acute pancreatitis decreased from 14.3% to 10.7%. The case-fatality for relapses remained stable at 3.2%. CONCLUSIONS: In this retrospective study the observed incidence of acute pancreatitis increased by 28% between 1985 and 1995. Due to a decrease in the case-fatality proportion, the mortality remained stable during this period. PMID- 11099068 TI - Eradication of hepatitis C virus 1b by interferon in a health care worker with acute hepatitis following needlestick transmission from a patient with chronic hepatitis C unresponsive to interferon. AB - Hepatitis C virus (HCV) was successfully eradicated by a short course of interferon (IFN) therapy in a nurse with acute HCV infection from a needlestick accident. The source patient had chronic hepatitis C and was a nonresponder to IFN therapy. The HCV genotype was 1b in patients, and a single point mutation (H- >R in amino acid 2218) was observed in the IFN sensitivity-determining region of the nonstructural 5A gene, in comparison with sequences of HCV-J, in HCV RNA from both the source patient (before and after IFN therapy) and the recipient (before IFN therapy). Though the strain transmitted was believed to be IFN-resistant in the patient with chronic hepatitis, the patient with acute hepatitis had a sustained response. PMID- 11099069 TI - A tribute to Thomas Bourne Turner, MD. PMID- 11099071 TI - Some methodological issues in the study of sexual networks: from model to data to model. AB - BACKGROUND: Mixing between sexual activity classes is an important determinant of sexually transmitted disease transmission. However, attempts to estimate sexual mixing patterns in the field remain limited partly because of practical and methodological difficulties. GOAL: To evaluate and identify appropriate sampling schemes to estimate the mixing pattern between sexual activity classes from large population networks with one or more components. STUDY DESIGN: The study is based on simulations of large population networks with various structural characteristics. A variety of snowball sampling schemes are applied to these networks and are evaluated by the quality of the mixing matrix estimates that they produce. RESULTS AND CONCLUSIONS: Unbiased estimation of mixing patterns (global assortativity, within-group mixing of the lowest activity classes, within group mixing of the highest activity classes) from large population networks is possible with a snowball sampling design in which the initial sample of index cases is drawn from the general population, all partners of the index case are recruited, and only one generation of partners are traced (one cycle). Simulation techniques proved useful in addressing complex methodological issues in situations where analytic results are difficult to obtain. PMID- 11099070 TI - Mathematical modeling as a tool in STD prevention and control: a decade of progress, a millennium of opportunities. PMID- 11099072 TI - Influence of mathematical modeling of HIV and AIDS on policies and programs in the developing world. AB - BACKGROUND: A number of simulation models have been developed to explore the dynamics of the AIDS epidemic. There seems to be minimal impact of these models on policies and programs. GOAL: To describe the major findings from simulation modeling and the impact of these findings on policies and programs. STUDY DESIGN: A literature review to summarize the major findings that are supported by more than one modeling group. RESULTS: Simulation modeling has contributed to improved understanding of a number of issues including the demographic impact of AIDS, the value of targeting prevention efforts to high-risk behavior, the importance of controlling STDs, the benefits of early intervention, and the need for combined interventions. CONCLUSIONS: Modeling has played a major role in increasing our understanding of the dynamics of the epidemic and in demonstrating how much we still need to learn. Its impact on policies and programs has been limited. The need for better translation of modeling findings to policy action will be even greater in the future. PMID- 11099073 TI - Risks of acquiring and transmitting sexually transmitted diseases in sexual partner networks. AB - BACKGROUND: A person's risk for acquiring infection and their role in continued transmission has traditionally been assessed on the basis of individual characteristics. Recently, network studies have attempted to relate individual risks to position in the wider network. GOAL: To assess the importance of local and global network structures in assessing the risk of acquiring and transmitting infection. STUDY DESIGN: An individual-based simulation model was used to construct a variety of potential network structures and track the transmission of infection over time. Logistic and Poisson regression were used to identify which measures of network position influence a person's risk for acquiring and transmitting infection. RESULTS: Measures of local centrality were more important to risk of acquisition, whereas global centrality mattered more to transmission. Continuous snowball sampling, rather than a fixed number of waves, better estimates a person's risks. CONCLUSIONS: There is an asymmetry regarding the risk of acquiring and transmitting infection. PMID- 11099075 TI - More realistic models of sexually transmitted disease transmission dynamics: sexual partnership networks, pair models, and moment closure. AB - BACKGROUND: Mathematical models of sexually transmitted disease transmission have proven powerful tools for interpreting observed epidemiologic pattern. However, the most commonly used formulation of such models largely fail to capture the effect of partnership concurrency and contact network structure on transmission. GOAL: The development of a compartmental model of partnership formation and dissolution that includes approximations for the influence of the sexual-partner network. STUDY DESIGN: Theoretical analysis of ordinary differential equation models for sexually transmitted disease transmission within sex-partner networks. RESULTS: The approach developed advances earlier pair models, allows for the influence of concurrent sexual partnerships, and illustrates the importance of concurrency to the persistence of diseases with relatively short durations of infectiousness. The authors also illustrate that heterogeneity in risk is possible even in model populations in which all individuals follow the same behavioral rules. CONCLUSION: Deterministic extended pair models offer a powerful approach to modelling sexually transmitted disease transmission that usefully complement computationally intensive microsimulation models. PMID- 11099074 TI - Epidemiology and control and curable sexually transmitted diseases: opportunities and problems. AB - BACKGROUND: Despite the availability of safe and effective treatment, infection with bacterial sexually transmitted diseases persists at a high prevalence in many populations. GOAL: To review the difficulties of parameter estimation when a cure is readily available and to explore the impact of different treatment and screening strategies that might maximize the benefits of using available treatments. STUDY DESIGN: A standard deterministic model for the spread of a bacterial sexually transmitted disease that causes symptomatic and asymptomatic infections, in which the population is stratified according to sex and sexual activity, is further stratified into two host groups to enable the modeling of different treatment and screening strategies. RESULTS: In the presence of a core group, if an infection has a high transmission probability, then screening for asymptomatic infections has a short-lived benefit. Repeated screening is slightly better if it is not restricted to a fraction of the at-risk population, but targeting of high-risk groups should be effective. Screening to treat asymptomatic infections in men could be beneficial if a substantial fraction of cases remain asymptomatic. CONCLUSIONS: After the initial gains achieved through treating symptomatic infections, further reductions in the prevalence of infections can be achieved by finding asymptomatic infections. However, these gains are difficult to achieve, especially in the case of gonorrhea. Because men are likely to have an asymptomatic chlamydial infection, screening of men for chlamydia should be worthwhile. PMID- 11099076 TI - Remarks on the role of economic modeling. AB - BACKGROUND: Despite the growing scholarly acceptance of quantitative evaluation methods, modelers still struggle to define an appropriate role for their work at the decision-making level. GOAL: To make observations about the policy relevance of mathematical and economic policy modeling in HIV and sexually transmitted disease prevention and treatment. RESULTS: The debate is framed within the context of the inevitability of decision making. Viewed in this light, models can inform choices between competing alternatives by leveraging existing information, integrating data from multiple sources, addressing "what if" questions, evaluating future scenarios, and providing a uniform metric for evaluation. CONCLUSIONS: The goal of model-based evaluation should not be to supplant the decision maker, but rather to redefine the terms of the debate using quantitative methods that make the beliefs and values that lie at the heart of all difficult choices explicit. PMID- 11099077 TI - Modeling contact networks and infection transmission in geographic and social space using GERMS. AB - BACKGROUND: Stochastic models of discrete individuals and deterministic models of continuous populations may give different answers to questions about infectious diseases. GOAL: Discrete individual model formulations are sought that extend deterministic models of infection transmission systems so that both model forms contribute cooperatively to model-based decision making. STUDY DESIGN: GERMS models are defined as stochastic processes in continuous time with parameters analogous to those in deterministic models. A GERMS model simulator was developed that insured that the rate of events depended only on the current state of model. RESULTS: The confidence intervals of long-term averages of infection level in simulated GERMS models were shown to contain the deterministic model means. CONCLUSION: GERMS models provide a convenient framework for testing the sensitivity of model-based decisions to a variety of unrealistic assumptions that are characteristic of differential equation models. GERMS especially facilitates making more realistic assumptions about contact patterns in geographic and social space. PMID- 11099078 TI - Sexual network structure and sexually transmitted disease prevention: a modeling perspective. AB - BACKGROUND: Mathematical modeling techniques are being used increasingly to plan public health policy for the prevention of sexually transmitted diseases. GOAL: An introduction to various aspects of network structure and sexually transmitted disease prevention using mathematical and simulation models. STUDY DESIGN: A short overview is given about some modeling approaches that are presently available. The effect of partnership duration on the spread of sexually transmitted diseases is discussed in the context of pair-formation models. Network models, which take concurrent partnerships into account, are discussed as a natural extension of pair-formation models. RESULTS: Results obtained by applying different modeling approaches to analyzing the efficacy of contact tracing in reducing the prevalence of sexually transmitted diseases are summarized. CONCLUSION: Partnership duration and network structure should be taken into account when estimating the impact of sexually transmitted disease prevention. Contact tracing can contribute significantly to reducing incidence and prevalence of sexually transmitted diseases. modeling has to be developed further to gain a better understanding of the relationship of network structure and the spread of specific sexually transmitted diseases. PMID- 11099079 TI - Mathematical models of the transmission and control of sexually transmitted diseases. AB - BACKGROUND: The development of mathematical models to describe and interpret the epidemiology of sexually transmitted infections has involved the incremental addition of various forms of biological and behavioral complexity to simple mathematical templates. GOAL: To review simple and complex models used in study of observed epidemiologic pattern. STUDY DESIGN: An overview of modeling in sexually transmitted disease epidemiology identifies the function of different types of models. RESULTS: Simple models have the advantage of transparency and analytical tractability and can illustrate the relative merits of different intervention options. However, real life is replete with complexities that can have effects that are difficult to predict in the absence of a mathematical framework. CONCLUSIONS: Research should increasingly be based on robust parameterization of model structures and try to capture individual behaviors. Progress will be most rapid by interdisciplinary work where the clinician, epidemiologist, and mathematician work collaboratively to help improve our knowledge of how to best control infection and disease. PMID- 11099081 TI - European Society for Paediatric Infectious Diseases Annual Awards, 2000. PMID- 11099080 TI - Evidence undermining the adequacy of the HIV reproduction number formula. PMID- 11099083 TI - Bacteriologic and clinical efficacy of one day vs. three day intramuscular ceftriaxone for treatment of nonresponsive acute otitis media in children. AB - BACKGROUND: One dose of intramuscular ceftriaxone has been recently licensed in the United States for the treatment of acute otitis media. However, data regarding the bacteriologic and clinical efficacy of this regimen in the treatment of nonresponsive acute otitis media are incomplete. OBJECTIVES: To determine the bacteriologic and clinical efficacy of a 1-day 50-mg/kg vs. a 3-day 50-mg/kg/day intramuscular ceftriaxone regimen in the treatment of nonresponsive acute otitis media in children. PATIENTS AND METHODS: In an open, prospective study 109 patients ages 3 to 36 months with culture-proved, nonresponsive acute otitis media were randomized to receive 1 (n = 49) or 3 (n = 60) 50-mg/kg/day intramuscular ceftriaxone doses, respectively. Middle ear fluid was aspirated for culture by tympanocentesis on the day of enrollment (Day 1); a second tympanocentesis with middle ear fluid culture was performed on Days 4 to 5. Additional middle ear fluid cultures were obtained if clinical relapse occurred after completion of therapy. Bacteriologic failure was defined by positive cultures on Days 4 to 5. Patients were followed until Day 28 after completion of therapy. Susceptibility of the middle ear pathogens was measured by E-test. RESULTS: Organisms recovered (n = 133) were Streptococcus pneumoniae (30 and 35 isolates for the 1-day and 3-day treatment group, respectively), Haemophilus influenzae (26 and 38, respectively) and Moraxella catarrhalis (n = 4). Of the 30 S. pneumoniae isolated from the 1-day group, 27 (90%) and 6 (20%) were nonsusceptible to penicillin and ceftriaxone, respectively; 9 of 27 (33%) were fully resistant to penicillin. Thirty-four (97%) and 6 (17%) of the 35 S. pneumoniae isolated from the 3-day group were nonsusceptible to penicillin and ceftriaxone, respectively; 16 of 34 (47%) were fully resistant to penicillin. Bacterial eradication of all H. influenzae and penicillin-susceptible S. pneumoniae was achieved in both treatment groups. Bacterial eradication of 14 of 27 (52%) and 33 of 34 (97%) penicillin-nonsusceptible S. pneumoniae was achieved in the 1-day and 3-day group, respectively. Seven (50%) of the 14 patients from the 2 groups who did not achieve bacterial eradication did not improve clinically on Days 4 to 5 and required additional ceftriaxone treatment. CONCLUSION: The 3 day intramuscular ceftriaxone regimen was significantly superior to the 1-day intramuscular ceftriaxone regimen in the treatment of nonresponsive acute otitis media caused by penicillin-resistant S. pneumoniae. PMID- 11099082 TI - Viral strain identification in varicella vaccinees with disseminated rashes. AB - BACKGROUND: Approximately 15% of recipients of live attenuated varicella vaccine may develop mild breakthrough varicella months to years after immunization. Although some vaccinees will develop zoster, it is less common in recipients of vaccine than in those who have had natural varicella. OBJECTIVE: To determine the varicella-zoster virus (VZV) strain responsible for breakthrough varicella and zoster in recipients of varicella vaccine. METHODS: A PCR assay capable of distinguishing wild-type from vaccine strain VZV was performed on samples from skin lesions from vaccinees with breakthrough varicella and zoster. RESULTS: All of 57 vaccinees with breakthrough varicella, clinically diagnosed on the basis of a generalized maculopapular or vesicular rash, in which there was amplifiable DNA [corrected], had wild-type VZV infection based on analysis of viral DNA. The Oka vaccine strain of VZV was not identified in any of these cases. In contrast, in 32 patients with zosteriform rashes, the vaccine strain was identified in 22 samples, and the wild-type strain was identified in 10 samples. CONCLUSIONS: Wild type virus was identified in all generalized rashes occurring after the immediate 6-week postvaccination period. When reactivation of vaccine strain occurred, it presented as typical zoster. We find no evidence that reactivation of vaccine virus occurs with the clinical picture of generalized rash. PMID- 11099084 TI - Immunization against hepatitis A in the first year of life: priming despite the presence of maternal antibody. AB - BACKGROUND: Maternal antibodies interfere with hepatitis A vaccination in young infants. We examined the response to a high dose hepatitis A vaccine administered concomitantly with a combination of diphtheria-tetanus toxoids-acellular pertussis-inactivated poliovirus vaccine/Haemophilus influenzae type b vaccine to initially seropositive vs. initially seronegative infants. METHODS: Three hundred subjects were originally planned to be enrolled at age 6 to 10 weeks and received hepatitis A vaccine (formalin-inactivated vaccine, SB-Bio, 720 enzyme-linked immunosorbent assay units) at 2, 4 and 6 months concomitantly with a diphtheria tetanus toxoids-acellular pertussis-inactivated poliovirus vaccine/H. influenzae type b vaccine. Children initially seropositive received a booster dose at 12 months of age. An additional 100 twelve-month-old infants previously not vaccinated with hepatitis A vaccine were given 1 dose, to observe the primary response at that age. Reactogenicity was recorded on diary cards for the 3 subsequent days. Immunogenicity was measured at Months 2, 4, 5, 10 and 11 after administration of the first vaccine dose. For the subjects enrolled at 12 months, blood was drawn before and 1 month after the first vaccination. RESULTS: Of 297 initially enrolled infants 36% were seronegative before vaccination (Group A). The geometric mean concentration (GMC) (milli-International Units/ml) of the seropositive infants (Group B) before immunization was 2587. The GMCs of Group A infants 1 month after each dose and at 12 months of age were 93, 518, 1656 and 786, respectively. For Group B infants, the respective GMCs were 1165, 460, 508 and 167. One hundred subjects of Group B received a booster dose at age 12 months; at Month 13 all were seropositive with a GMC of 1902. For comparison, a third group of 100 not previously immunized 12-month-old infants (Group C) were enrolled and received 1 dose of hepatitis A vaccine with pre- and postimmunization GMCs of 52 and 120, respectively. CONCLUSIONS: Our results suggest that the initially seropositive infants were primed despite maternal antibody interference. The hepatitis A vaccine was well-tolerated in this population of young infants. PMID- 11099085 TI - The Immunization Monitoring Program Active (IMPACT) prospective five year study of Canadian children hospitalized for chickenpox or an associated complication. AB - BACKGROUND: Varicella vaccine was approved for use in Canada in 1998. A major goal of universal varicella vaccine programs is to reduce severe infection and associated complications. Baseline data are essential against which to judge the effectiveness of routine childhood immunization. OBJECTIVE: To describe morbidity and mortality among children hospitalized for chickenpox. Methods. From January 1, 1991, to March 31, 1996, chickenpox admissions to 11 pediatric referral centers were actively identified. Patient and illness characteristics were compared for 3 subgroups defined by prior health: healthy; unhealthy but immunocompetent; immunocompromised. RESULTS: Of 861 cases 488 (56.7%) were healthy, 75(8.7%) were unhealthy and 298 (34.6%) were immunocompromised. The immunocompromised children differed from healthy/unhealthy cases in mean age (6.4 vs. 4.0/4.6 years, respectively, P < 0.0001); median interval from rash onset to admission (2 vs. 5/5 days, P < 0.0001); complication rate (20% vs. 90%/79%; P = 0.001); and rate of acyclovir therapy (98% vs. 24%/39%; P = 0.001). Unhealthy vs. healthy cases had a higher frequency (P < 0.01) of intensive care (13.3% vs. 4.7%), ventilation (9.3% vs. 2.0%) and death (4% vs. 0.2%). CONCLUSION: These data provide a baseline for morbidity/mortality resulting from chickenpox before varicella vaccine use in Canada. PMID- 11099087 TI - Effectiveness of palivizumab: evaluation of outcomes from the 1998 to 1999 respiratory syncytial virus season. The Palivizumab Outcomes Study Group. AB - BACKGROUND: Respiratory syncytial virus (RSV) remains a significant cause of morbidity, especially in premature infants and immunocompromised children, resulting in approximately 100 000 hospitalizations annually. A study was performed to evaluate the outcomes of those given palivizumab (Synagis; MedImmune, Inc., Gaithersburg, MD) during the 1998 to 1999 RSV season, its first season in general use. METHODS: A retrospective chart review of 1839 patients from 9 United States sites was conducted, representing all patients given palivizumab at each site. Those evaluated were to have a gestational age of < or =35 weeks, were to be <2 years old at their first injection and were to have received at least one dose of palivizumab (humanized monoclonal antibody against RSV) between September, 1998, and May, 1999. Gestational age, comorbidities, frequency of injections, hospitalizations and length of hospital stays were assessed. RESULTS: The antigen- or culture-positive RSV hospitalization rates for those given prophylaxis were 2.3% (42 of 1839) overall, 16/399 (4.0%) with chronic lung disease of infancy and 26 of 1227 (2.1%) born prematurely without chronic lung disease of infancy. Twenty-six patients had a gestational age of >35 weeks and were included in the analysis. CONCLUSIONS: Only 2.3% of children receiving palivizumab prophylaxis were hospitalized with RSV lower respiratory infection. This compares favorably with the rates observed in the pivotal trial (IMpact-RSV trial in 1996 to 1997), in which prophylaxis reduced hospitalization from 10.6% in the placebo group to 4.8% in those children receiving prophylaxis. PMID- 11099086 TI - Oral ciprofloxacin vs. intramuscular ceftriaxone as empiric treatment of acute invasive diarrhea in children. AB - BACKGROUND: Acute invasive diarrhea is a potentially serious condition in children. Because of the increasing resistance of enteric pathogens to commonly used oral antibiotics, intramuscular ceftriaxone has become the routine drug in the treatment of acute invasive diarrhea requiring an emergency visit in southern Israel. The inconvenience of this parenteral regimen created an increased need for oral pediatric formulations for the treatment of invasive diarrhea. OBJECTIVES: To evaluate the efficacy and safety of a suspension formulation of ciprofloxacin in the treatment of acute invasive diarrhea in infants and children. PATIENTS AND METHODS: From July 1996 through December 1997, 201 evaluable children ages 6 months to 10 years (35% <1 year; 70% <3 years) presenting with acute invasive diarrhea at the Pediatric Emergency Room were randomized to receive either ciprofloxacin suspension (10 mg/kg twice a day + im placebo; n = 95) or im ceftriaxone (50 mg/kg/day + placebo suspension; n = 106) for 3 days in a double blind manner. Stool cultures for Shigella, Salmonella, Campylobacter spp. and diarrheagenic Escherichia coli were obtained on Days 1, 3, 4 to 5 and 21 +/- 5. Clinical response and safety were assessed on Days 1, 2, 3, 4 to 5 and 21 +/- 5. RESULTS: We isolated 127 pathogens from 121 (60%) patients: 73 (57%) Shigella; 23 (18%) Salmonella; 18 (14%) E. coli; and 13 (10%) Campylobacter. Overall bacteriologic eradication on Day 4 to 5 was 99% for Shigella, 77% for Salmonella and 77% for Campylobacter, with no difference between the 2 groups. Clinical cure or improvement was observed in 100 and 99% of the ciprofloxacin and ceftriaxone groups, respectively. Serum ciprofloxacin values determined on Day 3 of the treatment were higher in the majority of patients than were the MIC50 and MIC90 values for the Shigella and Salmonella spp. isolated. Possible drug-related adverse events occurred in 13 patients [ciprofloxacin, 8 (8%); ceftriaxone, 5 (4.7%)] and were mild and transient. Joint examination was normal during and after completion of therapy in all patients. CONCLUSION: Oral ciprofloxacin was as safe and effective as intramuscular ceftriaxone for the empiric treatment of acute invasive diarrhea in ambulatory pediatric patients requiring an emergency room visit. PMID- 11099088 TI - Clinical sinusitis in children attending primary care centers. AB - OBJECTIVES: To determine the proportion of children who meet the clinical criteria for the diagnosis of sinusitis among all children attending primary care pediatric practices, to explore the relationship between passive smoking and the occurrence of sinusitis and to study the role of antibiotics in the management of sinusitis. DESIGN: A prospective observational cohort study. SETTING: Outpatient Pediatric Clinics of Jordan University of Science and Technology and Princess Rahma Teaching Hospital. Patients. All children ages 1 to 10 years presenting for any reason to participating practices. METHODS: Physicians participating in this study completed a questionnaire on all children attending the primary care centers, detailing the presence of nasal congestion or discharge, the duration of symptoms, daytime cough and whether symptoms were improving. The presence or absence of smokers in the family was also recorded. Children meeting our clinical criteria for sinusitis were further evaluated for other signs and symptoms including the type of medication prescribed. The severity of symptoms was reassessed at 10-day follow-up after the first visit. RESULTS: The study population was composed of 3001 children, of whom 249 met our clinical criteria for diagnosis of sinusitis (8.3%; 95% confidence interval, 7.3 to 9.3%). The prevalence rate of clinical sinusitis was greater among children age 5 years and older than among those younger (9.3% vs. 7.2%, P = 0.04). Children exposed to passive smoking in the household had clinical sinusitis significantly more than those not exposed (68.8% vs. 1.2%, P = 0.00). Antibiotics were prescribed for 80% of children who fulfilled the clinical criteria for diagnosis of sinusitis. Marked improvement of symptoms at the 10-day follow-up visit was reported among those who received antibiotics compared with those who did not (91% vs. 21.4%, P = 0.00). CONCLUSIONS: Sinusitis is not an uncommon problem in children, passive smoking might be a contributing factor and a course of antibiotic therapy is beneficial. PMID- 11099090 TI - Bacteriology of histopathologically defined appendicitis in children. AB - BACKGROUND: Acute appendicitis is the most common surgical emergency in childhood. However, the pathogenesis and detailed microbiology are obscure. OBJECTIVE: To determine in detail the bacterial etiology of appendicitis in children in relation to the histologic tissue pathology. STUDY DESIGN: Tissue samples obtained at surgery from 41 children with suspected acute appendicitis were examined histologically and by culture for aerobic and anaerobic bacteria. The patients were analyzed according to histopathologic and clinical findings. RESULTS: Aerobic and anaerobic species were isolated from 40 of 41 (98%) samples; on average, 14.1 isolates per specimen (10.4 anaerobes and 3.7 aerobes). Specimens from patients with gangrenous appendices yielded significantly higher numbers of anaerobic isolates per specimen than did specimens from patients with healthy appendices (11.7 vs. 7.7; P < 0.01). Bacteria belonging to the Bacteroides fragilis group were the most frequently isolated anaerobic microorganisms (95%). Other organisms frequently isolated in all histology groups were Peptostreptococcus micros (66%), Bilophila wadsworthia (63%), Fusobacterium nucleatum (44%), Eggerthella lenta (44%) and a hitherto undescribed bile resistant, pigment-producing Gram-negative rod (41%). Of the aerobes Escherichia coli (88%) and Streptococcus anginosus group (former Streptococcus "milleri" group) organisms (61%) were the most frequent findings. CONCLUSIONS: The shift from histologically normal toward gangrenous appendices was clearly associated with markedly elevated anaerobic bacterial counts in terms of species. The unusually high frequencies of B. wadsworthia (75%) and the hitherto undescribed bile-resistant, pigment-producing Gram-negative rod (56%) in gangrenous appendices represent unique and different findings from those reported in adults. PMID- 11099089 TI - Mother-to-infant transmission of TT virus: prevalence, extent and mechanism of vertical transmission. AB - OBJECTIVE: It is currently unknown which mechanisms are responsible for TT virus (TTV) infection in early childhood and whether it may be transmitted in utero from mother to infant. METHODS: The prevalence, mode and extent of maternal TTV transmission was investigated by testing blood, cord blood and breast milk samples from mother-infant pairs for the existence of the novel DNA virus. RESULTS: By means of polymerase chain reaction, TTV DNA was detected in 57 (41.3%) of 138 mothers and in 19 (13.8%) of 138 cord blood samples; therefore 33.3% of infants are likely to be infected by their mothers during the fetal period. Direct sequencing of TTV DNA from 2 mother-child pairs showed identical isolates. Follow-up sera from 3 TTV infected babies showed persistence of viremia. In blood samples from newborns older than 1 week 9 (27.3%) of 33 sera were TTV-positive. Viral sequences were also detected in 2 of 2 breast milk samples. In none of the infected subjects were biochemical or clinical signs of hepatitis observed. CONCLUSIONS: Our data prove that TT virus is efficiently transmitted transplacentally. The increase of its prevalence in the group of newborns older than 1 week suggests that it may be furthermore transmitted postnatally. Therefore in our Caucasian population, vertical transmission, particularly in utero transmission, of TTV is likely to account for a major part of TTV infection in early childhood. However, no disease activity could be established for the novel virus by this infection route. PMID- 11099091 TI - Pilot study of hydroxyurea in human immunodeficiency virus-infected children receiving didanosine and/or stavudine. AB - OBJECTIVE: To evaluate the safety and antiviral and immunologic effects of hydroxyurea given with didanosine (ddI) and/or stavudine (d4T) to symptomatic HIV infected children. METHODS: HIV-infected children with a history of long term nucleoside antiretroviral therapy were treated orally with hydroxyurea (initial dose, 10 to 20 mg/kg once daily; final dose, 30 mg/kg once daily), added to existing therapy that included ddI and/or d4T. RESULTS: Sixteen children were enrolled (mean age, 6.7 years; range, 1.8 to 13.4 years). Antiretroviral therapy used with hydroxyurea included d4T/ddI (12), ddI (2), d4T (1) and d4T/lamivudine (1). Children received between 24 and 48 weeks of therapy, which was well tolerated. Hydroxyurea was held temporarily during the first month of therapy in 4 cases because of neutropenia; all patients resumed hydroxyurea at full dosage without recurrence of neutropenia. No patient discontinued therapy permanently because of intolerance or toxicity. For the 13 children who completed 48 weeks of study treatment, the mean plasma HIV RNA concentration decreased from 4.6 log10 copies/ml at baseline to 4.2 log10 copies/ml at study Week 48 (P = 0.035, paired t test). Eight of these 13 children experienced a 0.5-log10 copies/ml or greater drop in HIV RNA concentration in the 48 weeks of study treatment. Appreciable changes in CD4+ lymphocyte percentage were not noted. CONCLUSIONS: Hydroxyurea, added to existing therapy with ddI and/or d4T, was well-tolerated and safe in HIV infected children. Evidence of antiviral activity was observed in some cases. PMID- 11099092 TI - Relationship of climate, ethnicity and socioeconomic status to Kawasaki disease in San Diego County, 1994 through 1998. AB - BACKGROUND: Kawasaki disease (KD) is the most common cause of acquired heart disease in children in the United States. By monitoring trends in patient numbers and demographics during a 5-year period, we were able to explore the relationship between climate, ethnicity, socioeconomic status and susceptibility to KD. METHODS: We conducted active surveillance for all patients hospitalized with KD in San Diego County from 1994 through 1998. Data on seasonal variation in monthly rainfall and temperature were obtained from the US Meteorological Service. Patient sex, age, date of admission and self-reported ethnicity were identified from patient medical records. Socioeconomic status was assessed on the basis of insurance status among patients hospitalized at a single institution. RESULTS: During the 5-year period there were 169 cases of KD in San Diego County. The overall annual incidence of KD in children < 5 years of age ranged from 8.0 to 15.4/100 000. KD incidence was inversely associated with average monthly temperature (r = -0.47, P < 0.001) and positively associated with average monthly precipitation (r = -0.52, P < 0.001). Asian/Pacific Islanders < 5 years of age were 2.7 times as likely and Hispanics were one-third as likely to be hospitalized for KD than children from all other ethnic groups combined. Children with private or military insurance in all ethnic groups were more likely to have a diagnosis of KD than children with government assistance or no insurance. After controlling for insurance status, only Asian/Pacific Islanders remained at increased risk (rate ratio, 2.14) for KD relative to all other ethnic groups combined. CONCLUSION: KD is a common childhood vasculitis of unknown etiology. The skewed ethnic distribution and seasonality are consistent with the hypothesis that KD is an infectious disease that is influenced by environmental and genetic factors. PMID- 11099094 TI - Diagnosis of tuberculosis in children. PMID- 11099095 TI - Clearance of adenoviral hepatitis with ribavirin therapy in a pediatric liver transplant recipient. PMID- 11099093 TI - Staphylococcus aureus with reduced susceptibility to glycopeptide antibiotics. PMID- 11099096 TI - Hepatitis B core promoter or precore gene mutations related to age at seroconversion from hepatitis B e antigen to anti-hepatitis B e in Japanese children. PMID- 11099097 TI - Rickettsia typhi seroprevalence among children in the Southeast United States. Tick-Borne Infections in Children Study (TICKS) Group. PMID- 11099098 TI - Coxiella burnetii myopericarditis and rhabdomyolysis in a child. PMID- 11099099 TI - Enteroviral meningoencephalitis in X-linked agammaglobulinemia: intensive immunoglobulin therapy and sequential viral detection in cerebrospinal fluid by polymerase chain reaction. PMID- 11099100 TI - Epidermodysplasia verruciformis associated with human papillomavirus 20 in a Korean child. PMID- 11099101 TI - Congenital tuberculosis manifesting as cutaneous disease. PMID- 11099102 TI - Fulminant amebic colitis in a ten-day-old infant. PMID- 11099103 TI - Brucellosis with acute acalculous cholecystitis. PMID- 11099104 TI - Fever and jaw pain in a five-year-old. TMJ septic arthritis. PMID- 11099105 TI - Treatment of hepatitis C infection. PMID- 11099106 TI - False positive strep A antigen test. PMID- 11099108 TI - Zinc, copper and manganese enhanced keratinocyte migration through a functional modulation of keratinocyte integrins. AB - The migration of keratinocytes plays an important role in the re epithelialization of cutaneous wounds. Zinc, copper and manganese are used in vivo for their healing properties and their mechanism of action is still only partially known. Thus, they have been shown both to promote keratinocyte proliferation and to modulate integrins expression. The aim of this study was to determine if trace elements induce an increase of the migration of keratinocytes and if this effect is related to the modulation of integrins. Two independent migration assays were used to study keratinocyte migration: the scratch assay using normal human keratinocytes and the modified Boyden chamber using HaCaT cells. Inhibition studies using function-blocking antibodies directed to alpha3, alpha6, alpha(v) and beta1 subunits were performed to investigate the modulator effect of trace elements on integrin function. In this way, zinc and copper gluconates increased alpha3, alpha(v) and beta1 function whereas manganese gluconate seems mainly able to modulate the function of alpha3 and beta1. The stimulating effect of these trace elements on keratinocyte migration does not appear related to alpha6 subunit. Thus, zinc, copper and manganese enhanced keratinocyte migration and one of the mechanisms was going through a modulation of integrin functions. PMID- 11099107 TI - Modulation of endothelin-1 in normal human keratinocytes by UVA1/B radiations, prostaglandin E2 and peptidase inhibitors. AB - In the skin, keratinocytes synthesize and secrete endothelin-(ET-1), a potent vasoconstrictor peptide which acts also as a growth factor for most skin cells. The aim of the study was to test the effects of UVA1 and the associations UVA1/B on the expression of ET-1 in normal human keratinocytes and to determine whether exogenously added prostaglandin E2 (PGE2) regulated ET-1 expression. As ET-1 is susceptible to degradation, we also evaluated whether ET-1 secretion was modulated by peptidase inhibitors. Our results showed that UVA1 (365 nm) did not modify the levels of preproET-1 mRNA and protein. Moreover, the associations UVA1+UVB or UVB+UVA1 down-regulated the overexpression of secreted ET-1 induced by UVB alone. PGE2 at 10(-5) M reduced the expression of ET-1 at the mRNA and protein levels but did not exert any significant modification at lower concentrations from 10(-10) to 10(6) M. Phosphoramidon, an endothelin converting enzyme (ECE) inhibitor, drastically decreased the amount of ET-1 accumulating in the culture medium in basal conditions or after UVB irradiation. Conversely, thiorphan, a specific inhibitor of neutral endopeptidase (NEP), rather increased the levels of ET-1 secretion mainly after UVB irradiation. Taken together, the results showed that normal human keratinocytes secrete and partly degrade ET-1 through ECE and NEP pathways and pointed out a differential regulation of ET-1 by UVB and UVA1 radiations without any noticeable role for PGE2. PMID- 11099109 TI - The effect of ultraviolet B irradiation on nitric oxide synthase expression in murine keratinocytes. AB - Nitric oxide (NO), which has several physiological functions in skin, is generated by NO synthase (NOS). NOS has at least three isoforms; endothelial NOS (eNOS), brain NOS (bNOS), and inducible NOS (iNOS). Ultraviolet B (UVB) irradiation has been reported to stimulate NO production in skin via induction or activation of NOS, however, the exact mechanism of NOS induction by UVB irradiation remains obscure. In this study, we investigated the direct effect of UVB on the expression of NOS isoforms in murine keratinocytes, and found a significant increase in NO production within 48 h. mRNA and protein expressions of bNOS were both enhanced by UVB irradiation in murine keratinocytes, whereas iNOS mRNA expression was suppressed at 4 and 12 h after UVB irradiation. These results suggest that the enhancement of NO production observed after UVB irradiation in murine keratinocytes may be explained in part by the upregulation of bNOS expression, but not iNOS expression. PMID- 11099110 TI - Inhibition of intercellular adhesio nmolecule-1 (ICAM-1) expression in ultraviolet B-irradiated human antigen-presenting cells is restored after repair of cyclobutane pyrimidine dimers. AB - The present study assessed the molecular mechanism underlying ultraviolet (UV) B radiation-induced inhibition of the expression of the adhesion molecule ICAM-1 in human antigen-presenting cells (APC). UVB radiation-induced inhibition of ICAM-I expression in human peripheral blood monocytes was associated with the generation of cyclobutane pyrimidine dimers (CPD). CPD were reduced by 60% after treatment with liposomal packed photolyase, an enzyme which removes CPD after absorption of photoreactivating light. Although incomplete, reduction of CPD was associated with complete restoration of ICAM-1 expression at the mRNA and protein level. Neither reduction of CPD level nor restoration of ICAM-1 expression were observed, if monocytes were treated with empty liposomes, or if they were irradiated with photoreactivating light prior to application of photolyase. DNA damage might also induce soluble mediators capable of autocrine inhibition of ICAM-1 expression. UVB irradiation of monocytes did not induce IL-10 production, but resulted in release of prostaglandin (PG) E2. Treatment of unirradiated monocytes with PGE2 completely inhibited ICAM-1 expression, thus mimicking the UVB effect. Inhibition of monocytic PGE2 production by indomethacin, however, did not restore ICAM-1 expression. These results suggest that formation of CPD is necessary and sufficient for UVB radiation-induced inhibition of ICAM-1 expression. In contrast, PGE2 might serve a paracrine role in UVB radiation induced immunosuppression. PMID- 11099111 TI - Expression of involucrin in normal, hyperproliferative and neoplastic mouse keratinocytes. AB - Involucrin is a protein precursor of the epidermal cornified envelope. Although expression of the human protein has been documented extensively, studies in the mouse have been hampered by a shortage of good antibodies. We describe the production of recombinant mouse involucrin and preparation of rabbit antisera to the protein that work well by immunohistochemistry and Western blotting. We confirm that in normal mouse epidermis the onset of involucrin expression is in the upper spinous layers and inner root sheath of the hair follicle. Involucrin was also detected in the differentiating epithelial cells of normal tongue, oesophagus and bladder. Involucrin was expressed in a subpopulation of mouse keratinocytes cultured in standard or low calcium medium and the proportion of involucrin-positive cells increased during suspension-induced terminal differentiation. Western blotting of keratinocytes from several inbred mouse strains revealed a remarkable heterogeneity in the electrophoretic mobility of involucrin, reflecting inter-strain variation in the number of tandem repeats in the protein. In the hyperproliferative epidermis of healing wounds involucrin was expressed in most of the suprabasal layers. In epidermal papillomas and carcinomas involucrin expression correlated well with degree of histological differentiation. The sites of expression of the mouse protein were thus the same as those previously reported for human involucrin. With the development of the new antibodies we anticipate that involucrin will become as widely used a marker of keratinocyte differentiation in the mouse as it is in the human. PMID- 11099112 TI - What is the most promising strategy for the treatment of metastasizing melanoma? AB - The treatment of patients with metastasizing melanoma, still one of the most deadly diseases in modern medicine, ranks among the greatest challenges that a clinician has to face. Metastatic melanoma also is one of the most profound sources of clinical frustration, since it provides far more ultimately defeating experiences than clinical victories. At the same time, the fascinating biology of melanoma has invited the study of this neuroectodermal tumor as a model system for dissecting many of the key problems of modern oncology, ranging from molecular oncogenesis via the controls of tumor proliferation, apoptosis, invasion, metastasis, and angiogenesis to tumor immunosurveillance and tumor drug resistance. Together with the dire need to develop more effective treatment modalities for improving both life expectancy and quality of life of affected patients, this has made metastatic melanoma a favorite model for the exploration of innovative strategies for tumor management. Encouragingly, many of these have already generated very promising results in animal models. However, this impressive level of research progress in conquering melanoma in the animal room contrasts rather pitifully with the actual progress made on the ward. This CONTROVERSIES feature, therefore, critically and soberly reviews the state of the art of treating metastatic melanoma today (distinguishing between nodal and distant metastases), and sharply defines unresolved or comparatively neglected key problems. In addition, this feature highlights several novel, provocative, hitherto underappreciated, yet potentially promising treatment approaches that deserve systematic exploration. Hopefully, this will offer further inspiration for the design and pursuit of innovative anti-melanoma strategies off-the-beaten track. PMID- 11099113 TI - Parents' and partners' life course and family experiences: links with parent child relationships in different family settings. AB - Life course and current family factors associated with individual differences in parent-child relationships were investigated in a sample of 467 children from 192 families, including stepfather, single-parent, stepmother, and complex stepfamilies; informants were fathers, mothers, and children. Both positive and negative dimensions of father-child and mother-child relationships were linked to earlier life course experiences of parent and of partner, to current family factors, and to the quality of partner's relationship with the child. The pattern of associations between the adults' life course experiences meant that children were at risk for a "double dose" of less affectionate relationships in families in which parents had experienced early adversities. The significance of biological relatedness, family setting, and child partner relationships was highlighted. PMID- 11099114 TI - Risk factors for long-term psychological effects of a disaster experienced in adolescence: predictors of post traumatic stress disorder. AB - This paper examines risk factors for the development of Post Traumatic Stress Disorder (PTSD), and its severity and chronicity, in a group of 217 young adults who survived a shipping disaster in adolescence. The survivors were followed up 5 to 8 years after the disaster. Risk factors examined fell into three main categories: pre-disaster child and family vulnerability factors, including childhood psychopathology; objective and subjective disaster-related experiences; and post-disaster factors, including results from screening questionnaires administered 5 months post-disaster, coping mechanisms adopted subsequently, life events, and availability of social supports. Developing PTSD following the disaster was significantly associated with being female, with pre-disaster factors of learning and psychological difficulties in the child and violence in the home, with severity of exposure to the disaster, survivors' subjective appraisal of the experience, adjustment in the early post-disaster period, and life events and social supports subsequently. When all these factors were considered together, measures of the degree of exposure to the disaster and of subjective appraisal of life threat, and ratings of anxiety obtained 5 months post-disaster, best predicted whether survivors developed PTSD. For those survivors who developed PTSD, its duration and severity were best predicted not by objective and subjective disaster-related factors, but by pre-disaster vulnerability factors of social, physical, and psychological difficulties in childhood together with ratings of depression obtained 5 months post-disaster, and whether survivors received post-disaster support at school. The implications of these findings are considered for targeting assessment and intervention efforts at survivors most at risk of developing difficulties in adjustment following similar traumatic experiences. PMID- 11099115 TI - Judgements about emotional events in children and adolescents with post-traumatic stress disorder and controls. AB - Research with clinically anxious adults has revealed that they estimate future negative events as far more likely to occur, relative to healthy controls. In addition, anxious adults estimate that such events are more likely to happen to themselves than to others. Previous research with anxious children and adolescents, in contrast, has revealed no increased probability estimates for negative events, relative to controls, and the events were rated as more likely to happen to others than to the self. The present study followed up these discrepant findings by investigating probability judgements concerning future negative events generated by children and adolescents who had actually experienced an extreme negative event and who met criteria for a diagnosis of Post-traumatic Stress Disorder (PTSD). Control groups comprised a group of healthy participants, and a group of healthy participants whose parents had experienced a trauma and who met criteria for PTSD. The results revealed no overall differences between the clinical group and the controls. However, children and adolescents with PTSD estimated all negative events as significantly more likely to happen to others than to themselves, with this other-referent bias being strongest for events matched to their trauma. In contrast, the two control groups exhibited an other-referent bias for physically threatening events but not for socially threatening ones. Developmental analyses indicated that the strength of the relationship between anxiety and elevated judgements about future negative events declined with age in the control participants but that there was no significant relationship in the groups who had been exposed to trauma. The findings are discussed in the context of the literature on information processing biases and PTSD. PMID- 11099116 TI - The association between direct and relational bullying and behaviour problems among primary school children. AB - The prevalence of direct and relational bullying and their differential relationship to behaviour problems in young primary school children was investigated. Individual interviews were conducted with 1982 children aged 6 9 years (mean age 7.6 years) and 1639 parents completed the Strength and Difficulties Questionnaire regarding behaviour problems of their children. Of the 1639 children with both data sets, 4.3% were direct bullies, 39.8 % victims, and 10.2% both bullied and were victimised frequently (bully/victims). The rates for relational bullying were 1.1% bullies, 37.9% victims, and 5.9% bully/victims. All children involved in direct bullying had significantly increased total behaviour problems, hyperactivity, conduct problems, and peer problem scores, and lower prosocial behaviour scores compared to those not involved in bullying (neutrals). Findings were similar for relational bullying involvement and behaviour problems for bully/victims and victims but less pronounced. Relational bullies had the lowest behaviour problem scores while being rated the least prosocially inclined children, consistent with the concept of a cool manipulator. Overall, direct bully/victims and children who were involved in both direct and relational bullying behaviour had the highest rates of behaviour problems. No relationship between victimisation and increased emotional problems were found. Those involved in bullying behaviour who show externalising and hyperactivity problems in primary school may be at increased risk for persistent conduct problems. Different interventions may be needed for those involved in relational bullying only, both direct and relational bullying, and those with additional behaviour problems. PMID- 11099117 TI - The developmental sequelae of nonorganic failure to thrive. AB - The developmental sequelae of infant failure to thrive (FTT) were examined in an unreferred group of 6-year-olds with a history of severe nonorganic growth retardation, sampled from a 1-year birth cohort in an inner-city area of South London. Children who failed to thrive in infancy (weight below the third centile for at least 3 months) and their pairwise matched comparisons were originally studied at 15 months, and 42 cases and 42 controls (89.5% of the sample) were followed up. At 6 years, previously growth-retarded children were considerably smaller than matched comparisons, in terms of body mass index (BMI), and height and weight for age Z scores. History of FTT explained substantial variance in weight and BMI at 6 years, with maternal height also contributing to variation in height for age. Child cognitive functioning at 6 years was examined using the McCarthy Scales: cases had more limited quantitative and memory skills than comparisons, but there was no intergroup variation in general cognitive performance. In contrast to analyses of physical development, failure to thrive did not account for cognitive functioning; maternal IQ was the sole significant predictor of performance on all indices of child cognitive abilities. At 15 months, earlier growth faltering was linked to limitations in mental development, but these findings were not confirmed by the follow-up data: the timing of FTT was not related to cognitive abilities at 6 years. Results correspond to past research indicating that nonorganic failure to thrive is associated with persistent limitations in physical stature. There was little evidence of cognitive disadvantage for case group children at school age, suggesting that the adverse effects of early malnutrition on cognitive functioning appear to diminish over time. PMID- 11099119 TI - The development and adjustment of 7-year-old children adopted in infancy. AB - The present study (N = 159) provides evidence of an increased risk for behavior problems of infant-placed 7-year-old internationally, transracially adopted children in the Netherlands. However, parents reported more behavior problems for adopted boys than for adopted girls. Notably, about 30% of the adopted children were classified as clinical on the CBCL scale for total problems, which is a much larger percentage than the 10% found in the normative population. It was suggested that these results could be explained by the operation of multiple risk factors before and after adoption placement, e.g. the child's genetic disposition, pre-natal and pre-adoption care, or the child's cognitive understanding of adoption in middle childhood. Also, results suggest that maternal sensitive responsiveness in adoptive families declines in the transition from early to middle childhood. In contrast to the home setting, the adopted children showed favorable behavioral and socioemotional adjustment at school, while their academic achievement and intelligence were in the normal range or above average. In particular Korean children had high IQs: 31% of these children obtained an intelligence score above 120. It was suggested that adoptive parents seem to offer their children sufficient or even more than average cognitive stimulation. Furthermore, adopted girls scored higher in optimal ego-control, social competence, and peer group popularity than nonadopted girls from the general population: 30% of the adopted girls were rated as popular by their classmates, which compares favorably to the 13% found in the general school population. PMID- 11099118 TI - The mother-child relationship following in vitro fertilisation (IVF): infant attachment, responsivity, and maternal sensitivity. AB - Infant attachment and mother-child interaction were evaluated for 65 primiparous women and their singleton infants conceived through in vitro fertilisation (IVF) and a control group of 61 women and their infants conceived naturally. The sample was enrolled during pregnancy as part of a longitudinal study. At 12 months postpartum, security of infant attachment was assessed using the Strange Situation procedure, and mother-child interaction was assessed in a free play context using the Emotional Availability Scales. IVF children demonstrated predominantly secure attachment relationships with their mothers (64.6% IVF, 55.9% controls), and there were no significant between-group differences in the proportion of IVF compared to control group children classified in any of the secure or insecure attachment groups. Furthermore, there were no significant group differences on maternal (sensitivity, structuring, hostility) or child (responsivity, involving) dimensions of interaction during play. The majority of IVF mothers (86%) were sensitive and their infants responsive (91%). Contrary to expectation, mother's ratings of greater anticipated infant difficultness assessed during pregnancy and higher ratings of infant temperament and behaviour difficulty assessed at 4 and 12 months postpartum were associated with secure attachment relationships and more optimal mother-child interaction in both the IVF and control groups. PMID- 11099120 TI - Psychiatric comorbidity in children and adolescents with reading disability. AB - This study investigated the association between reading disability (RD) and internalizing and externalizing psychopathology in a large community sample of twins with (N = 209) and without RD (N = 192). The primary goals were to clarify the relation between RD and comorbid psychopathology, to test for gender differences in the behavioral correlates of RD, and to test if common familial influences contributed to the association between RD and other disorders. Results indicated that individuals with RD exhibited significantly higher rates of all internalizing and externalizing disorders than individuals without RD. However, logistic regression analyses indicated that RD was not significantly associated with symptoms of aggression, delinquency, oppositional defiant disorder, or conduct disorder after controlling for the significant relation between RD and ADHD. In contrast, relations between RD and symptoms of anxiety and depression remained significant even after controlling for comorbid ADHD, suggesting that internalizing difficulties may be specifically associated with RD. Analyses of gender differences indicated that the significant relation between RD and internalizing symptoms was largely restricted to girls, whereas the association between RD and externalizing psychopathology was stronger for boys. Finally, preliminary etiological analyses suggested that common familial factors predispose both probands with RD and their non-RD siblings to exhibit externalizing behaviors, whereas elevations of internalizing symptomatology are restricted to individuals with RD. PMID- 11099121 TI - Assessing exposure to violence using multiple informants: application of hierarchical linear model. AB - The present study assesses the effects of demographic risk factors on children's exposure to violence (ETV) and how these effects vary by informants. Data on exposure to violence of 9-, 12-, and 15-year-olds were collected from both child participants (N = 1880) and parents (N = 1776), as part of the assessment of the Project on Human Development in Chicago Neighborhoods (PHDCN). A two-level hierarchical linear model (HLM) with multivariate outcomes was employed to analyze information obtained from these two different groups of informants. The findings indicate that parents generally report less ETV than do their children and that associations of age, gender, and parent education with ETV are stronger in the self-reports than in the parent reports. The findings support a multivariate approach when information obtained from different sources is being integrated. The application of HLM allows an assessment of interactions between risk factors and informants and uses all available data, including data from one informant when data from the other informant is missing. PMID- 11099122 TI - Local and global processing of music in high-functioning persons with autism: beyond central coherence? AB - A multi-modal abnormality in the integration of parts and whole has been proposed to account for a bias toward local stimuli in individuals with autism (Frith, 1989; Mottron & Belleville, 1993). In the current experiment, we examined the utility of hierarchical models in characterising musical information processing in autistic individuals. Participants were 13 high-functioning individuals with autism and 13 individuals of normal intelligence matched on chronological age, nonverbal IQ, and laterality, and without musical experience. The task consisted of same-different judgements of pairs of melodies. Differential local and global processing was assessed by manipulating the level, local or global, at which modifications occurred. No deficit was found in the two measures of global processing. In contrast, the clinical group performed better than the comparison group in the detection of change in nontransposed, contour-preserved melodies that tap local processing. These findings confirm the existence of a "local bias" in music perception in individuals with autism, but challenge the notion that it is accounted for by a deficit in global music processing. The present study suggests that enhanced processing of elementary physical properties of incoming stimuli, as found previously in the visual modality, may also exist in the auditory modality. PMID- 11099123 TI - Understanding theory of mind in children who are deaf. AB - Research on theory of mind began in the context of determining whether chimpanzees are aware that individuals experience cognitive and emotional states. More recently, this research has involved various groups of children and various tasks, including the false belief task. Based almost exclusively on that paradigm, investigators have concluded that although "normal" hearing children develop theory of mind by age 5, children who are autistic or deaf do not do so until much later, perhaps not until their teenage years. The present study explored theory of mind by examining stories told by children who are deaf and hearing (age 9-15 years) for statements ascribing behaviour-relevant states of mind to themselves and others. Both groups produced such attributions, although there were reliable differences between them. Results are discussed in terms of the cognitive abilities assumed to underlie false belief and narrative paradigms and the implications of attributing theory of mind solely on the basis of performance on the false belief task. PMID- 11099124 TI - The academic cup: Part I. PMID- 11099125 TI - Comments on the article "Marburg and Ebola virus infections in laboratory non human primates: a literature review". Soren Schou and Axel Kornerup Hansen. Comparative Medicine, 2000. 50:108-123. PMID- 11099126 TI - Improving the scientific basis for regulation of animal care and use. PMID- 11099128 TI - Can rabbits tell humans apart?: Discrimination of individual humans and its implications for animal research. AB - To predict when food reward was available, 12 New Zealand White rabbits were trained to discriminate between two humans. All subjects had significantly higher response rates and greater behavioral arousal in the presence of the positive stimulus person. The ability to discriminate between individual humans sets the stage for unanticipated Pavlovian conditioning, which may have considerable implications for animal research in behavioral and biomedical settings. PMID- 11099127 TI - Health care for genetically altered animals. PMID- 11099129 TI - Modified procedure for implantation of subcutaneous central venous access devices in macaques (Macaca mulatta). AB - A nonhuman primate model comprising adult male rhesus monkeys (Macaca mulatta) with chronically indwelling subcutaneous central venous access devices provides a unique opportunity to determine plasma pharmacokinetics of new drugs such as anticancer and anti- retroviral agents. The central venous access we use is a low profile, single-septum, titanium port that is attached to a radiopaque, indwelling catheter; the catheter is implanted in an internal jugular vein. A common complication following placement of the venous access device was migration of the catheter tip. We therefore modified the standard procedure by cutting the silicone catheter and introducing the rigid connector to secure the catheter to the vessel at the insertion site (approximately 9 to 13 cm from the distal end of the catheter). Prior to the use of the connector, three of five catheters migrated within 4 weeks after placement. In contrast, all 13 internal jugular catheters with connectors have remained patent without migration of the catheter tip. Therefore, incorporation of the catheter connector appears to have eliminated the problem of catheter migration. PMID- 11099130 TI - Evaluation of insulin resistance in two kinds of South American camelids: llamas and alpacas. AB - Insulin resistance was evaluated in South American camelids, llamas and alpacas, by use of the minimal model test and the insulin tolerance test. Animals were catheterized for long-term studies and tamed to minimize stress during evaluation. Results indicated a low insulin sensitivity index (SI) = 0 to 0.97, median = 0.39 x 10(-4) min/uIU x ml, about a fifth the value in other mammals and humans. The KITT was between 1.43 and 3.19 %/min, also significantly lower than that reported for humans. Glycosylated hemoglobin concentration was 6%, and HbAlc concentration was 5.5%; red blood cell lifetime, as measured by use of the 51Cr method, was 120 days, similar to the value in humans. We concluded that llamas and alpacas have naturally higher blood glucose concentration than do humans and other mammals during the glucose tolerance test. Using the same mathematical tools to evaluate glucose metabolism as those used in people, South American camelids appear to be resistant to insulin. Thus, the South American camelid may be a useful new animal model for the study of sugar metabolism and various facets of diabetes mellitus, especially protection from the deleterious effects of glycosylation. PMID- 11099131 TI - Leukocyte mobilization induced by hypervolemia is due to a combined alpha- and beta-adrenoceptor activation. AB - A phenomenon of leukocytosis induced by hypervolemic stress was discovered. Although a single injection of 350 microl of saline (equivalent to approx. 70 ml in humans, 1 ml/kg of body weight) did not have an effect on the leukocyte counts in long-term intravenously cannulated, freely behaving rats, a single injection of 750 microl of saline (equivalent to approx. 150 ml in humans, 2.1 ml/kg) induced rapid leukocytosis of 160% within 1 minute followed by a gradual increase up to 180% after 1 hour. Measurement of serum norepinephrine concentration revealed a significant increase in rats of the hypervolemic group, compared with those of the low volume group. Pretreatment with either the beta-adrenoceptor antagonist nadolol or the selective alpha2-adrenoceptor antagonist yohimbine prevented both leukocyte peaks in the high volume group, suggesting a combined receptor activation. This critical dependence of leukocyte counts on changes in blood volume should be taken into consideration in experiments with laboratory animals (the quantity of volume applications can falsify results of experiments). PMID- 11099132 TI - Effect of mouse strain and age on detection of mouse parvovirus 1 by use of serologic testing and polymerase chain reaction analysis. AB - BACKGROUND AND PURPOSE: Detection of mouse parvovirus 1 (MPV) depends on use of serologic and polymerase chain reaction (PCR) assays. These assays were evaluated for their ability to detect virus-specific antibodies or viral DNA in multiple strains and ages of mice inoculated with MPV. METHODS: Twelve-week-old ICR, BALB/c, C3H, C57BL/6, and DBA/2 mice and four- and eight-week-old ICR mice were inoculated with MPV. Serum was harvested four weeks after inoculation and analyzed by use of recombinant non structural protein 1 (rNS1) enzyme-linked immunosorbent assay (ELISA), minute virus of mice (MVM) ELISA, and MPV indirect fluorescent antibody (IFA), MVM IFA, and MPV hemagglutination inhibition (HAI) assays. Select tissues were harvested and analyzed by use of an MPV-specific PCR assay. RESULTS: The number of mice in each group with detectable MPV-specific antibodies or MPV DNA varied with mouse strain, mouse age when inoculated, and viral dose. Seroconversion in mice inoculated at 12 weeks of age was detected almost exclusively by use of the MPV IFA and MPV HAI assays, whereas seroconversion in almost all mice inoculated at 4 and 8 weeks of age was detected by use of all immunoassays except the MVM ELISA. Viral DNA was detected by use of PCR analysis in all strains and ages of mice except DBA/2 mice. CONCLUSIONS: Mouse strain and age have important roles in seroconversion to nonstructural and structural MPV antigens and persistence of viral DNA in mouse tissues. Therefore, diagnostic serologic testing and PCR analysis should be considered within the context of mouse strain and age at the time of MPV exposure, especially when sentinel mice are used for surveillance. PMID- 11099133 TI - Neuropathology of bouncer Long Evans, a novel dysmyelinated rat. AB - BACKGROUND AND PURPOSE: Spontaneous animal mutants affected by abnormal formation of myelin in the central nervous system (CNS) are useful in studies on myelinogenesis and remyelination leading to better understanding of cellular and molecular interactions involved in myelin repair. A novel rat mutant, Bouncer Long Evans (LE-bo) is severely dysmyelinated, but with exceptional longevity, and its clinical and pathologic phenotype are described. METHODS: Clinical observations, genetic studies, and determination of longevity were performed in a colony of rats, including carriers of LE-bo phenotype producing the mutant animals. Comprehensive histologic studies were performed on all perfusion-fixed tissues, and ultrastructural examination of the optic nerve and thoracic part of the spinal cord also was done in rats 1 to 14 weeks old. RESULTS: The LE-bo phenotype is characterized by whole body tremor, progressively severe ataxia, and severe seizure activity. The LE-bo phenotype is transferred as an autosomal recessive trait and is stable. The LE-bo rat can survive in good health beyond 45 weeks. Neuropathologic changes include severe global dysmyelination, with thin uncompacted myelin sheaths in young rats forming no major dense line, whereas the myelin sheaths of the peripheral nervous system appear normal. Oligodendrocytes degenerate with apparently progressing accumulation of membranous material in the perikaryon. Large numbers of immature glial cells were detected in the CNS of LE bo rats at 4 to 14 weeks. CONCLUSION: The LE-bo rat is severely dysmyelinated due to inability of its oligodendrocytes to form myelin sheaths. Similarities of the LE-bo rat and Long Evans Shaker (les) rat neuropathologic features, such as severe dysmyelination, lack of major dense line in uncompacted myelin sheaths, apparent proliferation of oligodendroglial cells, and considerable longevity, are striking and suggest that a LE-bo mutation may functionally affect the myelin basic protein gene. PMID- 11099134 TI - Variant form of diffuse corporal gastritis in NHE2 knockout mice. AB - Mice lacking the NHE2 Na+/H+ gene develop gastritis of the glandular mucosa as early as the tenth day of life, achieving maximal intensity of inflammation from 17 to 19 days after birth and maximal atrophy at one year. We assessed the effects of this process in such mice to 16 months of age. The stomach of NHE2 null mutants was examined at 10, 17 to 20, 24 to 35 and 49 to 70 days, and at 12 to 16 months. The NHE2 wild-type (+/+) and NHE2 heterozygous (+/-) mice were compared with the NHE2 homozygous mutant mice (-/-). The stomach of the mutant mice at all ages was characterized by a substantially reduced number of parietal cells. The 10-day-old mouse stomach had a transmural infiltrate of primarily neutrophils. With increasing age, neutrophils were replaced by lymphocytes and plasma cells in the glandular mucosa of the mutant mice. Young adult 49- to 70 day-old mice had surface cell hyperplasia and expansion of the replicating cell population. Hyperplasia of enterochromaffin-like cells and antral gastrin cells accompanied profound fundic gland and surface cell hyperplasia, and became progressively more severe with increasing age of the NHE2-/- mice. Neoplasms were not found in the mutant or control mice. This gastritis differs from that of autoimmune gastritis in that it is transmural, begins in infancy, and is associated with a predominantly neutrophilic infiltrate in its early stages. Some of the histologic changes in the adult mice can be explained on the basis of prolonged achlorhydria. This mouse may be a suitable model for prolonged effects of achlorhydria. PMID- 11099135 TI - Behavioral, clinical, and physiologic analysis of mice used for ascites monoclonal antibody production. AB - BACKGROUND AND PURPOSE: The effects of pristane inoculation, ascites accumulation, peritoneocentesis, and analgesics on the well-being of mice used in monoclonal antibody (MAb) production protocols were investigated. METHODS: Four experiments, each containing 17 to 21, 6- to 8-week-old male Balb/c mice, were conducted. Each experiment involved a period in which baseline data were collected, followed by intraperitoneal injections of pristane or phosphate buffered saline (PBS) inoculations into each mouse. One week later mice received intraperitoneal inoculations of either hybridoma cells or PBS. Parameters used to assess well-being throughout each of these periods included: wheel-running activity, food and water consumption, open-field box activity, clinical observation, and plasma corticosterone concentration. RESULTS: Compared to controls, pristane inoculation had slight to no affect on mice. There was no evidence of distress in cell-inoculated mice prior to their gaining 25% of their baseline body weight. The number of times (up to three) that peritoneocentesis was performed did not have a significant impact on mice's well-being, but ascites yields were greater when multiple harvests were performed. Cell-inoculated mice that gained weight slowly or developed high-particulate ascites were at higher risk of being distressed. CONCLUSION: Ascites yields can be maximized by performing multiple harvests; however, the well-being of mice used in such protocols should be closely monitored, as suggested here. PMID- 11099136 TI - Procedure for laryngotracheal separation in the goat: development of a new animal model for analysis of craniofacial growth. AB - Most animal models used to study the process of postnatal craniofacial growth require direct manipulation of the craniofacial area, a growth period, then evaluation of the area. However, the scar tissue associated with direct manipulation of the craniofacial structures can produce growth abnormalities that are unrelated to the manipulation itself. To avoid this confounding variable in the study of craniofacial growth, we developed an animal model that involves laryngotracheal separation in a young animal. Our procedure completely separates the trachea from the upper aerodigestive tract and removes the site of scar tissue formation from the region of investigation. The tracheal stomas of the goats we describe were maintained for as long as 9 months. Unlike human patients, goats with laryngotracheal separation require laryngectomy tubes to prevent life threatening stenosis of the tracheal stoma. Here we describe the operative procedure and post-operative care required for this new animal model. PMID- 11099137 TI - Isolation and characterization of Corynebacterium ulcerans from cephalic implants in macaques. AB - To determine the prevalence of colonization by Corynebacterium ulcerans, we cultured samples from the cephalic implant-skin margin and pharynx of 26 rhesus macaques and one pig-tailed macaque. All but one of the samples from the cephalic implants yielded a mixed population of bacteria. C. ulcerans grew from the cephalic implants in 56% and from the pharynx in 3% of the implanted animals. We screened nine of these isolates for diphtheria toxin (DT) and phospholipase D (PLD). Polymerase chain reactions (PCR) failed to identify DT in any of the tested isolates, which also lacked DT activity in Elek tests. However, all nine isolates tested had PLD toxin activity as determined by conjoint hemolysis on sheep blood agar plates in the presence of equi factor (Rhodococcus equi). In addition, PCR assays and Southern blot hybridization confirmed the presence of pld in the isolates. The role of the PLD toxin in promoting colonization of cephalic implants by C. ulcerans is unknown. We found C. ulcerans to be a frequent contaminant of the cephalic implant-skin margin. Further studies are necessary to investigate the relative clinical importance of this organism and the efficacy of various implant maintenance protocols in preventing infection. PMID- 11099138 TI - Ringtail in suckling Munich Wistar Fromter rats: a histopathologic study. AB - Ringtail is a pathologic condition of the tail of rats and other rodents that is traditionally attributed to low environmental humidity, although dietary deficiencies, genetic susceptibility, environmental temperature, and degree of hydration of the animal also have been suggested as possible causes. To the authors' knowledge, a detailed histopathologic study that may serve to shed light on the etiopathogenesis of this disease has not yet been published. We describe the histologic findings of ringtail observed in 12 suckling Munich Wistar Fromter (MWF) rats from two litters. Epidermal hyperplasia characterized by orthokeratotic and parakeratotic hyperkeratosis and acanthosis was observed in all affected rats. Numerous often dilated vessels were present in the dermis of tails that appeared of red/brown color at gross examination. In severe cases, the dilated vascular structures were thrombotic and accompanied by dermal hemorrhages and focal coagulative necrosis of the overlying epidermis. These findings suggest that epidermal acanthosis and hyperkeratosis are the main and primary events in the development of ringtail. To clarify the cause of this disease, future studies should be focused on the numerous factors that can induce such epidermal changes. PMID- 11099139 TI - Spontaneous intestinal adenocarcinoma in geriatric macaques (Macaca sp.). AB - BACKGROUND AND PURPOSE: Intestinal adenocarcinoma appears to be the most common malignant neoplasm in macaques, and is a substantial cause of morbidity and mortality in the elderly. METHODS: A retrospective review of 32 cases was done. RESULTS: Thirty-two cases were reviewed. Clinical examination had revealed severe weight loss, anorexia, and palpable abdominal mass. Microcytic hypochromic anemia, intermittent fecal occult blood positive test results, hypoproteinemia, and hypoalbuminemia were the predominant clinical laboratory findings. Carcinoembryogenic antigen serologic testing and single-strand conformational polymorphism analysis were performed in selected cases. The most common sites of the intestinal adenocarcinoma were ileocecal junction, colon, ileum, jejunum, and cecum. Metastases were evident in 34% of the cases and involved peripheral nodes, liver, lungs, pancreas, and adrenal gland. Overall survival of 12 macaques that underwent surgical excision was 83% at 6 months, 58% at 1 year, 50% at 1.5 years, 33% at 2 years, and 8% at 4 years. The overall mean survival rate (MSR) was > 483 postoperative days. CONCLUSION: Intestinal adenocarcinomas should be amenable to surgical resection. Early detection of localized, non-invasive neoplasms will increase surgical cure rate. Survivability could be potentially improved by use of adjuvant therapies. PMID- 11099140 TI - Use of injectable potassium chloride for euthanasia of American lobsters (Homarus americanus). AB - Potassium chloride (KCl: 330 mg/ml) was assessed as an euthanasia agent in American lobsters (Homarus americanus). Two groups of 10 lobsters (408.2 to 849.9 g) were maintained at 11.9 to 12.1 degrees C ('warm') and 1.5 to 2.5 degrees C ('cold') to evaluate the possible effect of ambient temperature on response to KCl. Death was defined as time of cardiac arrest, as viewed and measured by use of ultrasound. The KCl solution was injected (100 mg of KCl/100 g of body weight) at the base of the second walking leg to flood the hemolymph sinus containing the ventral nerve cord with potassium. Disruption of this 'central nervous system' was immediate, followed by cardiac arrest within 60 to 90 seconds. Group median ( +/- SD) baseline heart rate was 42 +/- 14 'warm' and 36 +/- 5 'cold' beats per minute. Time until cardiac arrest ranged from 35 to 90 (57 +/- 18) seconds in the 'warm' group and from 40 to 132 (53 +/- 34) seconds in the 'cold' group. There was no significant difference between group medians for either parameter. Histologic lesions were limited to mild to moderate acute degeneration, characterized by cell swelling, loss of contraction bands, and occasional mild cytoplasmic vacuolation of skeletal muscle at the injection site. Injectable KCl solution was an effective, reliable method for euthanasia of H. americanus. PMID- 11099141 TI - Serial laparoscopic biopsies of liver and spleen from Schistosoma-infected baboons (Papio spp.). AB - PURPOSE: To obtain large, serial biopsy samples from the liver and spleen by using laparoscopy. Large samples were needed for measurement of inflammatory mediators during various stages of schistosomiasis. METHODS: Each of the seven female baboons (Papio sp.) underwent as many as three laparoscopies, for a total of 19 laparoscopic procedures. This process permitted sampling of the liver, spleen, and mesenteric lymph nodes before and at 6 and 9 weeks after infection with Schistosoma mansoni. All surgery was performed through three trocar sites. Postoperative care included preemptive analgesia. After surgery, we monitored the animals' appetite and measured the core body temperature and activity by using implanted radiofrequency transmitters. RESULTS: We obtained samples of the liver and splenic biopsies during all 19 laparoscopic procedures. The mean weight of the liver biopsies was 3.7 g and that of the spleen samples was 5.3 g. We encountered small adhesions during 5 of the 12 reoperations. Eating and activity rapidly returned after surgery. CONCLUSIONS: Laparoscopy permitted collection of large, serial biopsies with apparently limited stress to the animals. Laparoscopy can be used for biopsies in studies to characterize disease response, confirm normal organ histology prior to drug toxicity studies, determine target-organ drug concentrations in pharmacokinetic studies, and measure drug residues. This refinement likely will reduce required animal numbers by decreasing the effect of surgery compared to that of the experimental conditions, enhance animal well being, and permit repeated measurements in an animal that serves as its own control. PMID- 11099143 TI - Laryngeal chondrosarcoma: a 24-year experience at the Royal National Throat, Nose and Ear Hospital. AB - This paper presents a review of the experience of 12 patients with chondrosarcoma of the larynx treated at The Royal National Throat, Nose and Ear Hospital, London, over the last 24 years. This represents the largest European series and the third largest in the world. Chondrosarcoma is the most frequent non epithelial malignant tumor of the larynx, with 83.3% of cases arising from the cricoid cartilage in our series. We have shown that deep wedge biopsy with a CO2 laser, aided by computerized tomography scanning, gives the most reliable diagnosis. Laryngeal chondrosarcoma is characterized by indolent growth, a potential for local recurrence and, infrequently, by metastases. The treatment details of our patients are discussed. Adequate partial resection is often successful and use of CO2 laser as the initial treatment in five of these cases is presented. PMID- 11099142 TI - Environmental enrichment-related injury in a macaque (Macaca fascicularis): intestinal linear foreign body. AB - A three-year old male cynomolgus macaque (Macaca fascicularis) presented with clinical signs of anorexia and depression that decreased over a 48-hour period. Results of abdominal radiography abdominocentesis, blood biochemical analysis and CBC suggested septic peritonitis. Exploratory laparotomy revealed multiple perforations along the mesenteric border of the small intestine. Necropsy revealed masses of fibrous material in the stomach and cecum. Multiple mucosal ulcerations, as well as linear fibrous material, were found in the small intestine. The ulceration, perforations, and septic peritonitis were attributed to the ingestion of rope that had been attached to the animal's cage as an environmental-enrichment device. PMID- 11099144 TI - Auditory dysfunction in stroke. AB - The auditory and vestibular systems share the same end organ and cranial nerve, yet vestibular signs and symptoms are common with stroke, whereas hearing disturbances are much less frequent. Several reasons would appear to account for this striking dissimilarity. One is that the auditory pathway is less ubiquitous than the vestibular pathways. The likelihood that a stroke involves the auditory pathway is, therefore, less on this basis alone. A second difference, to our knowledge not previously reported, is that the auditory pathway is often spared by the most common strokes. This is because major parts of the auditory pathway, such as the cochlear nucleus, inferior colliculus and medial geniculate body, have multiple sources of blood supply. A third well-recognized factor is the redundancy of the central auditory system and its strong bilateral representation above the level of the cochlear nuclei. Consequently, rostral to the cochlear nuclei gross deficits in hearing, such as those measured by standard pure-tone audiometry and speech discrimination, only occur if lesions are bilateral. Furthermore, widespread bilateral lesions of the auditory system typically render the patient unable to respond or are incompatible with life. In contrast, language disorders are more frequent because language is usually unilaterally represented in the cortex. Certainly, cerebral stroke often includes the auditory system, resulting in various types of auditory disorders, but most hemispherical lesions produce subtle hearing dysfunctions that can only be detected with sophisticated psychoacoustic and electrophysiological testing. The purpose of this review is to provide an overview of the auditory system and its blood supply and to review how auditory processing can be affected by stroke. Psychoacoustic and electrophysiological test procedures for identifying lesions in the central auditory system are described. The literature of hearing disorders due to stroke is reviewed and illustrative cases are presented. PMID- 11099145 TI - Extracellular adenosine 5'-ATP-induced calcium signaling in isolated vestibular ganglion cells of the guinea pig. AB - Extracellular adenosine 5'-triphosphate (ATP)-induced intracellular calcium concentration ([Ca2+]i) changes in acutely isolated vestibular ganglion cells (VGCs) of the guinea pig were investigated using the Ca2+ -sensitive dye Fura-2. Extracellular ATP induced an increase in [Ca2+]i in VGCs in a dose-dependent manner. ATP induced an increase in [Ca2+]i even in the absence of extracellular Ca2+ (1 mM Ethylene Glycol-bis (beta-aminoethyl Ether) N,N,N',N'-Tetraacetic Acid (EGTA)), thus suggesting that ATP induces Ca2+ release from the intracellular stores. The P2-receptor antagonists suramin and reactive blue 2 inhibited the ATP induced [Ca2+]i increase in a dose-dependent manner. The P1-receptor agonist adenosine did not induce any changes in [Ca2+]i. These results suggest that VGCs may possess a P2-purinergic receptor but not a P1-purinergic receptor. La3+, a receptor-mediated calcium channel blocker, inhibited the ATP-induced [Ca2+]i increase but, in contrast, nifedipine, a L-type calcium channel blocker, did not. These results suggest that ATP induces both a Ca2+ -release from the intracellular stores and a Ca2+ influx from the extracellular space through La3+ sensitive and nifedipine-insensitive Ca2+ channels in VGCs. Our results also suggest that extracellular ATP may act as a neurotransmitter or neuromodulator of the vestibular peripheral system in the guinea pig. PMID- 11099146 TI - Distribution of HLA-A, -B and -DRB1 alleles in patients with sudden sensorineural hearing loss. AB - This study was performed to investigate the association between human leukocyte antigen (HLA) and susceptibility to sudden sensorineural hearing loss in the Korean population. HLA-A and HLA-B typing using a standard microlymphocytotoxicity technique and HLA-DRB1 genotyping were performed in 35 patients with sudden sensorineural hearing loss and in 206 healthy controls. Prednisone (usual dose 60 mg/day) was administered for 6 days and tapered for an additional 4-6 days. Both initial hearing levels at the onset of deafness and final hearing levels after treatment were examined and evaluated for association with HLA alleles. The frequency of HLA-DRB1*14 was increased in patients with sudden sensorineural hearing loss compared with controls (relative risk [RR] = 2.7, p = 0.016). The frequencies of HLA-A2, -A31, -B52, -B61, -DRB1*04, -DRB1*11 and -DRB1*12 were slightly higher than in the controls, but did not reach statistical significance. When an association between the treatment results and HLA alleles was also evaluated, the frequency of HLA-DRB1*04 was found to be increased in the patients who did not respond to steroid treatment compared with both patients who responded well to steroid (50%, vs 16%, p = 0.034) and controls (RR = 3.0, p = 0.046). These results suggest that there is an association between HLA-DRB1*14 and disease susceptibility and that the presence of HLA-DRB1*04 may be an useful marker for predicting a poor prognosis in Korean patients with sudden sensorineural hearing loss. PMID- 11099147 TI - Eustachian tube function varies over time in children with secretory otitis media. AB - Impaired opening and closing functions of the Eustachian tube are considered to be pathogenic factors in secretory otitis media (SOM). As the clinical course of SOM is variable, the variability of tubal function is of interest. We aimed to explore the short- and long-term variability of tubal opening and closing functions in SOM. The study comprised 42 ears in 21 children (13 males and 8 females) with tympanostomy tubes due to SOM. The middle ear pressure was recorded during repeated passive forced openings, equalization of + 100 daPa and - 100 daPa by swallowing, Valsalva inflation and forceful sniffing. Test sessions were performed twice (separated by 30 min) on each of 2 days, with a mean interval of 3.7 months in between. In the forced opening test there was a considerable intra individual variability over time. Expressed as SD of the mean, the variability of the forced opening and closing pressures in individual ears was on average 15% and 23%, respectively, between sessions and 20% and 30% respectively, between test days. In the equalization, Valsalva and sniff tests the rates of responses that changed from positive to negative between sessions and test days ranged from 12% to 33%. Female gender and retraction pockets were related to poorer opening function in the forced opening test. Ears with serous effusion (in contrast to mucoid) showed a similar trend and also a lower occurrence of positive equalization, Valsalva and sniff tests. It was concluded that Eustachian tube opening and closing functions are highly variable in ears with SOM. Consequently, single tubal function tests have low value when used as a prognostic tool in individual ears. PMID- 11099148 TI - Computerized electronystagmography: normative data revisited. AB - This study provides normative data from computerized electronystagmography (ENG) testing of 40 healthy subjects with an average age of 45 years. The clinical test protocol comprises an extensive vestibular examination with oculomotor, positional, rotary chair and caloric tests. The results show that with a computerized ENG set-up considerable variabilities of 22% for rotary chair asymmetry and 19% for caloric labyrinth asymmetry remain. PMID- 11099149 TI - Bone-conducted evoked myogenic potentials from the sternocleidomastoid muscle. AB - The aim of this study was to show that bone-conducted clicks and short tone bursts (STBs) can evoke myogenic potentials from the sternocleidomastoid muscle (SCM) and that these responses are of vestibular origin. Evoked potential responses to bone-conducted auditory stimuli were recorded from the SCMs of 20 normal volunteers and from 12 patients with well-defined lesions of the middle or inner ear or the VIIIth cranial nerve. The subjects, who had various labyrinthine and retro-labyrinthine pathologies, included five patients with bilateral profound conductive hearing loss, two with bilateral acoustic neuroma post-total neurectomy and five with bilateral sensorineural hearing loss. Air- and bone conducted evoked myogenic potentials in response to clicks and STBs were recorded with surface electrodes over each SCM of each subject. In normal subjects, bone- and air-conducted clicks and STBs evoked biphasic responses from the SCM ipsilateral to the stimulated ear. The bone-conducted clicks evoked short-latency vestibular-evoked myogenic potential (VEMP) responses only in young subjects or in subjects with conductive hearing loss. STBs evoked VEMPs with higher amplitude and better waveform morphology than clicks with the same acoustic intensity. Patients with total VIIIth cranial nerve neurectomy showed no responses to air- or bone-conducted click or STB stimuli. Clear VEMP responses were evoked from patients with conductive or sensorineural hearing loss. It is concluded that loud auditory stimuli delivered by bone- as well as air conduction can evoke myogenic potentials from the SCM. These responses seem to be of vestibular origin. PMID- 11099150 TI - The role of neck afferents in subjective orientation in the visual and tactile sensory modalities. AB - We studied the influence of neck afferents on the perception of orientation. In Experiment 1, we investigated the effect of head tilt on the subjective vertical in both the visual and tactile modalities. The results showed that head tilt triggers an Aubert effect in the visual modality and a Muller effect in the tactile modality. Significant positive correlations between the two adjustment modalities were restricted to head tilt to the left. In Experiment 2, we investigated the role of neck afferents on tactile orientation in seated and supine positions. The results showed that, in the supine position, the tactile E effect was twice as large as in the seated position. These experiments confirm that tactile perception of orientation is affected by neck afferents, and show that the influence of neck afferents is limited by relevant gravitational cues. PMID- 11099151 TI - Analysis of magnetic resonance imaging acoustic noise generated by a 4.7 T experimental system. AB - High intensity acoustic noise is an undesirable side-effect in magnetic resonance imaging (MRI) that can cause discomfort and hearing loss in patients and may be an impediment in functional MRI (fMRI) studies of the auditory system. Experimental MRI systems with high magnetic field strengths may generate acoustic noise of higher sound pressure levels (SPLs) than conventional 1.0 and 1.5 T clinical systems. We measured the SPL and spectral content of the acoustic noise generated by the Bruker Biospect 47/40 4.7 T experimental MRI system during scanning sequences commonly used in animal testing. Each sequence generated acoustic noise of high SPL, rapid pulse rates, amplitude-modulated pulse envelopes and multi-peaked spectra. The rapid acquisition with enhancement sequence with a 0.25 mm slice thickness generated SPLs of up to 129 dB peak SPL and 130 dB (A). Fourier analysis of the spectral content of the acoustic noise generated by each MRI sequence showed a wide band of acoustic energy with spectral peaks from 0.2-5 kHz. The intense MRI acoustic impulse noise generated by the 4.7 T system may cause masking of stimuli used in fMRI of the auditory cortex, reduce the hearing acuity of experimental animals and present a risk for unprotected human ears. PMID- 11099152 TI - Long-term changes in middle latency response and evidence of retrograde degeneration in the medial geniculate body after auditory cortical ablation in cats. AB - Short- and long-term changes in the middle latency response (MLR) after bilateral ablation of the auditory cortices were studied in awake cats. The amplitude of the negative peak with a latency of about 15 ms (NA) decreased to 60% of the original value 1 week after ablation (short-term change). In the long term, i.e. 11-30 months, NA either decreased further (decreased group) or remained unchanged (non-decreased group). A histological study with light microscopy revealed degeneration of neurons in the ventral nucleus of the medial geniculate body (MGv) in the decreased group, whereas the neurons in this region were preserved in the non-decreased group. This study suggests that long-term changes in NA reflect retrograde degeneration in the MGv after auditory cortical ablation. PMID- 11099153 TI - Spontaneous firing activity of cortical neurons in adult cats with reorganized tonotopic map following pure-tone trauma. AB - We hypothesized that moderate sensorineural hearing loss resulting from acoustic trauma would cause (i) a change in the cortical tonotopic map, (ii) an increase in spontaneous activity in the reorganized region and (iii) increased inter neuronal synchrony within the reorganized part of the cortex. Five kittens were exposed to a 126 dB sound pressure limit tone of 6 kHz for 1 h at both 5 and 6 weeks of age. Recordings were performed 7-16 weeks after the exposure. Auditory brainstem response thresholds for frequencies above 12 kHz were increased by 30 dB on average relative to those in normal cats. Tonotopic maps in the primary auditory cortex were reorganized in such a way that the area normally tuned to frequencies of 10-40 kHz was now entirely tuned to 10 kHz. Spontaneous firing rates were significantly higher in reorganized areas than in normal areas. In order to test for changes in inter-neuronal synchrony, cross-correlation analysis was done on 225 single-unit pairs recorded in the traumatized cats. For the single- and dual-electrode pairs there was no significant difference in peak cross-correlation coefficients for the firings of simultaneously recorded cells between normal and reorganized areas. However, the percentage of correlations that differed significantly from zero was higher in the reorganized area than in the normal area. This suggests a potential correlation between cortical reorganization, increased spontaneous firing rate and inter-neuronal synchrony that might be related to tinnitus found in high-frequency hearing loss induced by acoustic trauma. PMID- 11099154 TI - Nasal secretion in ragweed-sensitized dogs: effect of leukotriene synthesis inhibition. AB - Allergic rhinitis is an inflammatory disease associated with local leukotriene release during periods of symptoms. Zileuton, a leukotriene synthesis inhibitor, is known to inhibit the release of leukotriene B4. Because we previously showed that leukotriene B4 is a potent mediator of neutrophil-dependent nasal secretion, we investigated whether Zileuton inhibited allergen-induced nasal secretion. Using a newly developed method for isolating and superfusing a nasal segment, we examined the effect of Zileuton on nasal secretion and neutrophil recruitment in ragweed-sensitized dogs. Instillation of ragweed into the nasal segment caused time-dependent increases in the volume of airway fluid and the recruitment of neutrophils. Zileuton prevented ragweed-induced neutrophil recruitment and nasal secretion. These results indicate that leukotrienes are important mediators of allergy-induced nasal secretion in dogs. Future clinical studies in allergic patients will determine whether there is a therapeutic role for leukotriene synthesis inhibitors in modulating neutrophil recruitment and hypersecretion in the nose. PMID- 11099155 TI - Effects of anti-VLA-4 monoclonal antibody treatment in murine model of allergic rhinitis. AB - In order to study the role of VLA-4 in allergic rhinitis, the effects of anti mouse VLA-4 monoclonal antibody (mAb) were evaluated in a murine model. BALB/c mice were sensitized first by i.p. injections (general sensitization) and then by daily nasal dripping of antigen (local sensitization) before performing a nasal antigen challenge. The mAb was applied either before the antigen challenge (BC group), before the local sensitization (BL group) or before the general sensitization (BS group). The effects were evaluated in terms of antigen-induced early-phase nasal symptoms (sneezing), late-phase nasal eosinophilia and the serum level of antigen-specific IgE. Antigen-induced nasal eosinophilia was significantly (p = 0.009) reduced in the BL group but not in the BC group (number of eosinophils = 114 +/- 15.1, 244 +/- 52.8 and 347 +/- 50.5 in the BL, BC and control groups, respectively). The serum level of the specific IgE was also significantly (p = 0.038) reduced in the BL group but not in the BC group (optical density = 1.18 +/- 0.07, 1.28 +/- 0.13 and 1.58 +/- 0.14 in the BL, BC and control groups, respectively). The suppressive effect on sneezing was not significant in either the BL or BC groups. In the BS group, suppressive effects on antigen-induced nasal responses and the specific IgE level were not statistically significant. These findings suggest that VLA-4 plays an important role in the topical booster or priming effects during repeated nasal antigen exposures in pre-sensitized animals. PMID- 11099156 TI - Scanning electron microscopic study of the neuromuscular junctions of the cricothyroid and thyroarytenoid muscles in rats. AB - Neuromuscular junctions were observed in the cricothyroid (CT) and thyroarytenoid (TA) muscles of adult rats by scanning electron microscopy after removing the intramuscular connective tissue components using the HCI hydrolysis method. Morphologically, the junctions were classified into three types in the CT muscle and two types in the TA muscle, based on the structural characteristics of the subneural apparatuses, including junctional folds. In the CT muscle, type 1 junctions (32%) consisted of more than 15 cup-like depressions with slit-like junctional folds. Type 2 junctions (20%) were characterized by approximately 10 cup-like depressions with a small number of pit- or slit-like junctional folds. Type 3 junctions (48%) had irregular labyrinthine gutters with slit-like junctional folds. In the TA muscle, type 1 (82%) and 2 (18%) junctions had similar structures to type 1 and 2 junctions in the CT muscle, respectively. Histochemical studies using myosin adenosine triphosphatase staining showed that both CT and TA muscles predominantly consisted of type II muscle fibers (78% and 82%, respectively), and that the diameter of type II fibers was larger than that of type I fibers. These findings suggest that the type 2 junction belongs to type I muscle fibers, while both type 1 and type 3 junctions belong to type II fibers, and that the type 3 junction is a structural variation of the type 1 junction. The significance of the structural differences of the subneural apparatuses in the intrinsic laryngeal muscles is discussed briefly. PMID- 11099157 TI - Acoustic analyses clarify voiced-voiceless distinction in tracheoesophageal speech. AB - In order to clarify the ability of the voice to achieve voiced voiceless distinction in [ce1]tracheoesophageal (TE) speech, acoustic cues such as closure duration, onset and offset of vibration during closure period and voice onset time (VOT), in conjunction with intraoral pressure, were analyzed in 40 TE speakers. Both closure period and VOT during [p] production were longer in TE speakers with high intelligibility compared with laryngeal speakers; during [b] production these parameters were similar between the two groups. TE speakers with high intelligibility and laryngeal speakers showed significant differences between [p] and [b] production in terms of both closure duration and VOT. TE speakers with low intelligibility of [b] had higher values of VOT during [b] production compared with those with high intelligibility. TE speakers with low intelligibility of [p] had lower values of VOT during [p] production compared with those with high intelligibility. It is concluded that these characteristic acoustic cues reflect voicing ability in TE speech. PMID- 11099158 TI - Correlation between glottal area and photoglottographic signal in normal subjects. AB - Photoglottography (PGG) is an established technique for depicting the vibratory patterns of the vocal folds. The present study investigates the correlation between the glottal area and the corresponding PGG signal. Six normal (five male, one female) subjects who did not use their voices professionally were investigated during constantly sustained phonation at spontaneous pitches. Laryngostroboscopy was performed in combination with PGG. The simultaneously recorded laryngostroboscopic images and PGG signals were directly digitized and stored on a computer. The correlation between the glottal area and the corresponding PGG amplitudes across each vibratory cycle of the vocal folds was calculated and they were found to be highly and positively correlated (r = 0.973, p < 0.001). The PGG signal reflects changes in the glottal area during the vibration cycle of the vocal folds. The proposed simultaneous laryngostroboscopic and PGG technique has proved to be useful for facilitating the interpretation of changes in glottal area. PMID- 11099159 TI - Preoperative virtual endoscopy and three-dimensional imaging of the surface landmarks of the internal carotid arteries in trans-sphenoidal pituitary surgery. AB - In trans-sphenoidal pituitary adenoma surgery the sella turcica is opened between the internal carotid arteries. Three-dimensional image processing methods were applied in this study to avoid the risk of damaging the arteries during the opening of the anterior wall of the sella. By using graphical software it was possible to combine the anatomies of the carotid arteries and the sellar wall into one non-perspective three-dimensional image. With a perspective image (virtual endoscopy), the sphenoid sinus landmarks were presented as if looking through a nasoendoscope. This also facilitated preoperative planning but the non perspective images, with the carotid arteries marked, were found to be the most useful and suitable for clinical routine. The pituitary tumor itself and its relations with the adjacent structures were best evaluated from magnetic resonance imaging scans but, for the opening of the sellar wall and in the three dimensional orientation with endoscopy, three-dimensional computerized tomography imaging with the carotid arteries marked was found to be helpful. PMID- 11099160 TI - Inhibition of carnitine synthesis modulates protein contents of the cardiac sarcoplasmic reticulum Ca2+-ATPase and hexokinase type I in rat hearts with myocardial infarction. AB - It was previously reported that inhibition of carnitine synthesis by 3-(2,2,2 trimethyl-hydrazinium) propionate (MET-88) restores left ventricular (LV) systolic and diastolic function in rats with myocardial infarction (MI). Preservation of the calcium uptake function of sarcoplasmic reticulum Ca2+-ATPase (SERCA2) is one of the possible mechanisms by which MET-88 alleviates hemodynamic dysfunction. To test this hypothesis, the effects of MET-88 on protein content of SERCA2 were evaluated using the same rat model of heart failure. Myocardial protein content of hexokinase, which is one of the key enzymes of glucose utilization, was also measured. Either MET-88 (MET-88 group) or a placebo (MI group) was administered for 20 days to rats with MI induced by coronary artery ligation. The control group underwent sham surgery (no ligation) and received placebo. In LV myocardial homogenates, the myocardial SERCA2 protein content was 32% lower (p<0.05) in the MI group than in the control group. However, in the MET 88 group myocardial SERCA2 content was the same as in the control group. Hexokinase I protein content was 29 % lower (p<0.05) in the MI group compared with the control. In contrast, hexokinase II protein content did not differ significantly among the three groups. Consequently, inhibition of carnitine synthesis ameliorates depression of SERCA2 and hexokinase I protein content which may reduce tissue damage caused by MI. PMID- 11099161 TI - Diminished posttetanic potentiation in failing human myocardium. AB - In heart failure a decreased function of SERCA 2 has been demonstrated. The present study aimed at investigating the relation between sarcoplasmic reticulum Ca2+-load (SR-Ca2+-load) and the activity of the SERCA 2. SR-Ca2+ load was evaluated by measuring posttetanic potentiation (PTP) in human nonfailing (NF, n = 10) and endstage failing myocardium (DCM, n = 11). In addition, the effect of cyclopiazonic acid (CPA), a specific inhibitor of SERCA 2, on PTP was studied in both NF and DCM. In crude membrane preparations from the same hearts the maximal SERCA 2 activity was determined and correlated with the PTP. In failing myocardium the PTP was significantly reduced compared to nonfailing myocardium (13.7+/-0.75 mN/mm2 vs. 17.1+/-1.55 mN/mm2, p<0.05, +/- SEM). When PTP was studied in the presence of increased extracellular Ca2+-concentrations, the difference between NF and DCM was further pronounced. CPA decreased PTP in both nonfailing and failing human tissue. The maximal SERCA 2 activity was significantly reduced in failing myocardium (NF 267+/-18.5 nmol ATP/mg protein x min(-1) vs. DCM 191+/-13.4 nmol ATP/mg protein x min(-1), p<0.05, +/- SEM). Correlation of the PTP and maximal SERCA 2 activity revealed a close correlation between both parameters in NF and DCM. In summary, the presented results suggest that reduced SERCA 2 activity in DCM influences posttetanic force potentiation probably through a reduced SR-Ca2+-load. PMID- 11099162 TI - Ventricular fibrillation threshold and local dispersion of refractoriness in isolated rabbit hearts with left ventricular dysfunction. AB - Patients with congestive heart failure or left ventricular dysfunction (LVD) have a high incidence of ventricular tachyarrhythmias and sudden cardiac death. In addition to structural, metabolic and neuroendocrine changes, mechanoelectrical feedback may play a role in arrhythmogenesis in heart failure. Three groups of rabbits (n = 10 for each) were studied: chronic coronary ligation with ejection fraction (EF) > or = 0.45 or < 0.45, and sham-operated controls. Ventricular fibrillation (VF) thresholds were measured at LV pressures of 0 and 40 mm Hg during modified Langendorff perfusion. Intervals between local activations during VF (VFI) were used as an index of refractoriness. Global dispersion was expressed as coefficient of variation of VFI; local dispersion by maximum difference in VFI between adjacent sites. Median VF threshold was lower at 0 mm Hg in the lower EF group compared to controls (30 vs. 67.5 mA, P<0.05). VF threshold in control hearts was lower at 40 mm Hg than at 0 mm Hg (P<0.01), but there was no further reduction in threshold in LVD hearts at 40 mm Hg. Global dispersion of VFI did not differ significantly between groups. Local dispersion of VFI in the lower EF group was greater than in controls at 0 mm Hg in the infarct border zone (P<0.05). At 40 mm Hg, local dispersion of VFI in zones bordering and remote from the infarct were greater in both LVD groups than in controls (P<0.05). Local inhomogeneity of refractoriness is more marked in the infarct border zone, but latent abnormalities are evident in normal myocardium of rabbits with left ventricular dysfunction and are revealed by left ventricular distension. PMID- 11099163 TI - Complementary cardioprotective effects of flavonoid metabolites and terpenoid constituents of Ginkgo biloba extract (EGb 761) during ischemia and reperfusion. AB - Hemodynamic and electron spin resonance (ESR) analyses were performed on isolated ischemic and reperfused rat hearts to assess the cardioprotective and antioxidant effects of therapeutically relevant concentrations of Ginkgo biloba extract (EGb 761; 5, 50 or 200 microg/ml), its terpenoid constituents (ginkgolide A; 0.05 microg/ml and ginkgolide B; 0.05, 0.25 or 0.50 microg/ml), and a terpene-free fraction of EGb 761 (CP 205; 5 or 50 microg/ml). Hearts underwent 10 min of low flow ischemia, 30 min of no-flow global ischemia, and 60 min of reperfusion. Test substances were added to the perfusion fluid during the last 10 min of control perfusion, low-flow ischemia and the first 10 min of reperfusion. A separate group of rats was treated with CP 205 (60 mg/kg/day; p.o.) for 15 days, after which the hearts were perfused with plain buffer. In ESR experiments, the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was added to the perfusate to determine the effects of treatments on post-ischemic myocardial free radical generation. Results showed that in vitro exposure of hearts to EGb 761 (5 or 50 microg/ml) or to ginkgolides A and B (both at 0.05 microg/ml), or in vivo pretreatment of the rats with CP 205 delayed the onset of contracture during ischemia. The strong reperfusion-induced elevation of left ventricular end diastolic pressure observed in untreated hearts was significantly reduced by in vitro exposure to the lowest concentrations of EGb 761, by ginkgolide A, and to a lesser extent by ginkgolide B, or by prior oral treatment with CP 205. Post ischemic functional recovery.was significantly improved by in vivo administration of CP 205, by perfusion with 5 microg/ml of EGb 761 or with both terpenoids as compared to untreated group but in vitro CP 205 was not effective. ESR analyses revealed that DMPO-OH (the DMPO/hydroxyl radical spin-adduct) concentrations in coronary effluents were markedly decreased by all treatments, except for the lowest concentration of ginkgolide B. Perfusing 5 microg/ml EGb 761 resulted in a better inhibition of baseline DMPO-OH concentration than 5 microg/ml CP 205 (-70 % and -48 % vs. control, respectively), indicating that both terpenoid and flavonoid constituents of EGb 761 are required to produce this effect. CP 205 was significantly more efficient in reducing DMPO-OH concentration when administered in vivo than when applied in vitro, indicating that the antioxidant effect of flavonoid metabolites (formed in vivo) is superior to that of intact flavonol glycosides (present in vitro). Collectively, these findings provide the first evidence that part of the cardioprotection afforded by EGb 761 is due to a specific action of its terpenoid constituents and that this effect involves a mechanism independent of direct free radical-scavenging. Thus, the terpenoid constituents of EGb 761 and the flavonoid metabolites that are formed after in vivo administration of the extract act in a complementary manner to protect against myocardial ischemia-reperfusion injury. PMID- 11099164 TI - Species-dependent changes in mechano-energetics of isolated cardiac muscle during hypoxia. AB - The present study investigates the mechanical and energetic changes induced by hypoxia in isolated cardiac muscles of different species characterized by different myosin isoforms. Classic mechanical parameters of contraction and energetic parameters derived from the tension-velocity relationship were studied in rat and guinea pig left ventricular papillary muscles and in frog ventricular strips before and after 15 min hypoxia (n = 8 in each group). The isomyosin pattern is predominantly V1 with high ATPase activity in rat and V3 with low ATPase activity in guinea pig and frog heart ventricles. At baseline, cardiac mechanical performance was greater in rat than in guinea pig and frog muscle, but the economy of tension generation did not differ significantly between the three species. Hypoxia significantly decreased myocardial mechanical performance in all three groups. Mechanical impairment was more marked in rat than in the other two species and was intermediate in guinea pig. The energetic consequences of hypoxia differed according to species and in a different manner from the mechanical parameters. Hypoxia decreased the economy of tension generation in rat heart, in contrast to no change in guinea pig and frog muscle. These results suggest that in terms of mechano-energetic properties, cardiac muscles with V1 isomyosin were more sensitive to hypoxia than those containing V3 isomyosin. PMID- 11099165 TI - Recovery from sarafotoxin-b induced cardiopathological effects in mice following low energy laser irradiation. AB - OBJECTIVE: Low energy laser irradiation has been shown to accelerate various biological processes, including regeneration of injured tissues. In the present work we studied the effect of low energy laser irradiation on ischemic mice hearts, following administration of sarafotoxin-b, a powerful vasoconstrictor peptide of the endothelin/sarafotoxin family. METHODS: Immediately after injection of the toxin and two days later, hearts were exposed to low energy laser irradiation. Eight days after the injection the mice were sacrificed and their hearts were processed for light and electron microscopy. RESULTS: Sarafotoxin-b induced an approximate 2-fold increase in the relative cavity volume of the left ventricle. Low energy laser irradiation of the sarafotoxin injected mice caused a significant decrease (62%) in the left ventricular dilatation. Electron microscopy of the sarafotoxin-injected mice hearts revealed that the extent of formation of large vacuoles in the cytoplasm of the cardiomyocytes as well as disorganization of the myofibrils were much higher than in the laser irradiated mice. CONCLUSIONS: Our study indicates that low energy laser irradiation enhanced recovery of the damaged cardiomyocytes from the ischemia induced by sarafotoxin-b. PMID- 11099166 TI - Inhibition of protein phosphatase 1 induces apoptosis in neonatal rat cardiac myocytes: role of adrenergic receptor stimulation. AB - The mechanisms that regulate cardiac myocyte apoptosis are not well understood. To study the role of protein phosphatase 1 (PP1) and 2A (PP2A) in apoptosis, we exposed cultured neonatal rat cardiac myocytes to the phosphatase inhibitor okadaic acid (OA). Exposure (18 h) to 100 nM OA (a concentration which inhibits both PP1 and PP2A) decreased the number of adherent cells, caused genomic DNA fragmentation, and increased the percentage of apoptotic cells. These effects did not occur at a lower concentration of OA (1 nM) which is relatively specific for PP2A. Stimulation of alpha1- or beta-adrenergic receptors with norepinephrine (NE) in the presence of propranolol or prazosin partially blocked OA-induced apoptosis as measured by flow cytometry. Likewise, stimulation of adenylyl cyclase with forskolin reduced OA-induced apoptosis. Conversely, inhibition of protein kinase A with H89 or protein kinase C with chelerethrine potentiated OA induced apoptosis. OA increased caspase-3 activity, and this effect was reduced by NE. Thus, inhibition of PP1 stimulates apoptosis in NRVM and stimulation of adrenergic receptors protects against OA-induced apoptosis. PMID- 11099167 TI - Reduction in myocardial apoptosis associated with overexpression of heat shock protein 70. AB - It is reported that ischemia-reperfusion induces apoptotic cell death in myocardium. It is also demonstrated that heat shock protein 70 (HSP70) enhances myocardial tolerance. Therefore, it is hypothesized that HSP70 may play a role in the attenuation of myocardial apoptosis. To elucidate this goal, HSP70 overexpressing and control-transfected rat hearts were prepared using gene transfection by intra-coronary infusion of the hemagglutinating virus of Japan liposome. In vivo experiment Hearts of both groups were subjected to global ischemia, followed by reperfusion in situ. Shorter recovery time to spontaneous beating (HSP70-transfected vs. control-transfected; 46.7+/-4.6 vs. 67.5+/-7.0 s, p = 0.033) and lower serum CPK levels (415+/-27 vs. 533+/-36 IU, p = 0.027) were observed in the HSP70-transfected group. The HSP70-transfected group also showed a lower percentage of cardiac myocytes positively stained by nick end labeling after ischemia-reperfusion (17.5+/-4.9 vs. 40.0+/-5.1%, p = 0.010). In vitro experiment Cardiac myocytes isolated from the hearts of both groups (prepared separately from the in vivo experiment) were subjected to hypoxia-reoxygenation. Flow cytometry was used to identify the cells that showed sub-G1 DNA content as apoptotic cells. Apoptotic cells as a percentage of viable cells increased more in the control-transfected group after hypoxia-reoxygenation (13.0+/-0.77 vs. 21.9+/-1.18%, p<0.0001). In conclusion, we demonstrated that apoptosis after ischemia-reperfusion was decreased in the HSP70-overexpressing heart in vivo and in vitro, leading to the suggestion that HSP70 could be associated with the reduction in myocardial apoptosis. PMID- 11099168 TI - Influence of the angiotensin II AT1 receptor antagonist irbesartan on ischemia/reperfusion injury in the dog heart. AB - The aim of the present study was to investigate whether the non-peptide angiotensin II type 1 (AT1) receptor antagonist irbesartan (SR 47436, BMS 186295, 2-n-butyl-3 [2'-(1H-tetrazol-5-yl)-biphenyl-4-yl)methyl]-1,3-diaza-spiro [4,4]non 1-en-4-one) has myocardial protective effects during regional myocardial ischemia/reperfusion in vivo. Eighteen anesthetized open-chest dogs were instrumented for measurement of left ventricular and aortic pressure (tip manometer and pressure transducer, respectively), and coronary flow (ultrasonic flowprobes). Regional myocardial function was assessed by Doppler displacement transducers as systolic wall thickening (sWT) in the antero-apical and the postero-basal wall. The animals underwent 1 h of left anterior descending coronary artery (LAD) occlusion and subsequent reperfusion for 3 hours. Irbesartan (10 mg kg(-1), n = 9) or the vehicle (KOH, control, n = 9) was injected intravenously 30 min before LAD occlusion. Regional myocardial blood flow (RMBF) was measured after irbesartan injection and at 30 min LAD occlusion using colored microspheres. Infarct size was determined by triphenyltetrazolium chloride staining after 3 h of reperfusion. There was no recovery of sWT in the LAD perfused area in both groups at the end of the experiments (systolic bulging, -15.1+/-6.1% of baseline (irbesartan) vs. -12.3+/-3.0% (control), mean+/-SEM). Irbesartan led to an increase in RMBF in normal myocardium (2.47+/-0.40 vs. 1.35+/-0.28 ml min(-1) g(-1), p<0.05), and also to an increase in collateral blood flow to the ischemic area (0.27+/-0.04 vs. 0.17+/-0.02 ml min(-1) g(-1), P = <0.05). Infarct size (percent of area at risk) was 24.8+/-3.2 % in the treatment group compared with 26.9+/-4.8% in the control group (P = 0.72). These results indicate that a blockade of angiotensin II AT1 receptors with irbesartan before coronary artery occlusion led to an increase in RMBF, but did not result in a significant reduction of myocardial infarct size. PMID- 11099169 TI - Level of circulating phospholipase A2 in prediction of the prognosis of patients with suspected myocardial infarction. AB - OBJECTIVES: Atherosclerotic lesions result from inflammatory-proliferative responses of the endothelium and smooth muscle of the arterial wall. Poor prognosis of acute myocardial infarction (AMI) patients has been associated with elevated levels of acute phase proteins including C-reactive protein. We investigated the significance of circulating phospholipase A2 in the long-term prognosis of suspected AMI patients. METHODS: The concentration of phospholipase A2 was measured by an immunoassay in sera of 100 suspected AMI patients. Admission phospholipase A2 95 % fractile outliers were excluded to eliminate the effect of acute infectious diseases. The total and atherosclerotic mortalities were followed for a 4-year period. RESULTS: The most powerful prognostic limit for both admission (p = 0.02, RR = 2.6 and 95% CI = 1.2 to 5.6) and maximal (p = 0.06, RR = 2.4 and 95% CI = 0.96 to 5.9) phospholipase A2 groups was > or =8 microg/l. The admission phospholipase A2 level had an independent prognostic significance for atherosclerotic mortality (p = 0.04, RR = 2.4 and CI = 1.02 to 5.8) in multivariate analysis with CK-MB and age. CONCLUSIONS: The elevated serum phospholipase A2 level at admission is an independent predictor of long-term atherosclerotic mortality in patients with suspected AMI. The prognostic significance of phospholipase A2 weakens during hospitalisation concomitant to the onset of the acute inflammatory response to myocardial injury. PMID- 11099170 TI - Delayed attenuation of myocardial ischemia with repeated exercise in subjects with stable angina: a possible model for the second window of protection? AB - AIMS: A delayed myocardial protection extends between 24 and 96 h after ischemic preconditioning in animals. To test for this phenomenon in humans, subjects with stable angina were subjected to exercise test-induced myocardial ischemia and the effect of this "preconditioning" ischemic insult on the exercise-induced myocardial ischemia with the re-exercise after 24-96 hours was studied. METHODS AND RESULTS: Forty-eight males with a history of infarction and positive exercise test were recruited to the study. After baseline symptom-limited exercise test, the subjects were randomized to four experimental groups (n = 12/group). The groups were allowed to recover for 24 h, 48 h, 72 h or 96 h before performing the second exercise test. Variables analyzed were heart rate-systolic blood pressure product at 1 mm ST segment depression, time to 1 mm ST segment depression, maximum ST segment depression, exercise duration, and the total ischemic time. There were no intergroup differences in baseline values for these variables. All variables were significantly improved at 24 h, the improvement peaked usually at 48 h (maximum increase in the variables by 31-46%), and the variables returned to baseline by 96 h after the first test. CONCLUSIONS: The exercise-induced ischemia caused transient attenuation of myocardial ischemia with re-exercise. Although the time-window and the time-course of this effect shows striking resemblance to those of the delayed preconditioning in animals, its mechanism remains speculative. The most probable mechanisms that may be involved include increased myocardial perfusion and/or some adaptive changes in the myocardium, the delayed preconditioning being one possibility. PMID- 11099171 TI - Magnetocardiographic QT dispersion during cardiovascular autonomic function tests. AB - QT dispersion is considered to reflect nonhomogeneity of ventricular repolarization. The autonomic nervous system modulates QT interval duration, but the effect may not be spatially homogenous. Magnetocardiography (MCG) registers the weak magnetic fields generated by myocardial electric currents with high localizing accuracy. We studied the effects of rapid cardiovascular autonomic nervous adjustment on QT dispersion in MCG. Ten healthy male volunteers were monitored during deep breathing, the Valsalva maneuver, sustained handgrip, hyperventilation, the cold pressor test and mental stress. 67 MCG channels and 12 ECG leads were recorded simultaneously. A computer algorithm was used for QT interval measurements. QT dispersion was defined as maximum - minimum or standard deviation of the QTpeak and QTend intervals. In MCG the QT(end) dispersion increased during deep inspiration compared with deep expiration (96+/-19 ms v. 73+/-27 ms, p = 0.05). Magnetic QT dispersion tended to increase during the bradycardia phase of the Valsalva maneuver, but the change was obvious only for QT(end) (55+/-26 ms v. 76+/-29 ms, p<0.05). Other tests had no significant effect on QT dispersion, not even the cold pressor test, although it causes strong sympathetic activation. Magnetic and electric QT(peak) and QT(end) intervals correlated closely (r = 0.93 and 0.91), whereas the QT dispersion measures showed no correlation. In conclusion, magnetic QT dispersion is not modified by rapid changes in autonomic tone, but maneuvers involving deep respiratory efforts and changes in ventricular loading affect QT dispersion measurements. PMID- 11099173 TI - Conformational transition of DNA in electroreduction studied by in situ UV and CD thin layer spectroelectrochemistry. AB - Electrochemically induced three conformational transitions of calf thymus DNA from B10.4 to Z10.2-DNA and from B10.2 to B10.4 and to C-DNA in 10 mM phosphate buffer solution (pH 7.21) at glassy carbon electrode are found and studied by in situ circular dichroism (CD) thin layer spectroelectrochemistry with singular value decomposition least square (SVDLS) analysis. It indicates that the so called B10.2 form and the C-form of DNA may be composed of B10.4 and left-A DNA and of B10.4 and right-A DNA, respectively. The irreversible electrochemical reduction of adenine and cytosine groups in the DNA molecule is studied by UV-Vis spectroelectrochemistry. Some electrochemical parameters alpha n = 0.17, E0' = 0.70 V (vs. Ag/AgCl), and the standard heterogeneous electron transfer rate constant, k0 = 1.8 x 10(-5) cm s(-1), are obtained by double logarithmic analysis and non-linear regression. PMID- 11099172 TI - Correlation between the effects of a cationic peptide on the hydration and fluidity of anionic lipid bilayers: a comparative study with sodium ions and cholesterol. AB - The cationic tridecapeptide alpha-melanocyte stimulating hormone (alpha-MSH) is known to interact with anionic vesicles of 1,2-dimyristoyl-sn-glycero-3 phosphoglycerol (DMPG), partially penetrating the lipid membrane. In the lipid liquid crystal phase, phospholipid derivatives spin labeled at the different C atoms along the acyl chain, show that the peptide increases the bilayer packing at all depths. Parallel to that, there is an increase in the probe's isotropic hyperfine splittings, indicating that the peptide significantly decreases the membrane hydrophobic barrier. Accordingly, it is suggested that the increase in membrane packing yielded by alpha-MSH is partly due to a greater level of interchain hydration. This result is compared to the increase in packing and decrease in polarity yielded by cholesterol, and the absence of structural or polar alterations with Na+. The latter result shows that the peptide effect is not related to an increase of positive charges at the anionic vesicle surface. Alterations on the lipid bilayer polar profile measured by the nitroxide hyperfine splitting z component in frozen samples are shown to be different from those obtained at room temperature. However, it is shown here that a certain correlation can be drawn between the increase in polarity measured in frozen samples and the packing effect caused by the different molecules in the lipid gel phase. PMID- 11099174 TI - A spectroscopic analysis of thermal stability of the Chromobacterium viscosum lipase. AB - The thermal stability of the lipase from Chromobacterium viscosum was assessed by deactivation (loss of activity), fluorescence, circular dichroism (CD) and static light scattering (SLS) measurements. Lipase fluorescence emission is dominated by the tryptophyl contribution. An increase in the tyrosyl contribution from 2 to 16% was only observed upon prolonged incubation at 60 degrees C. The effect of temperature on the tryptophyl quantum yield was studied and two activation energies were calculated. Tryptophan residues in the native structure have an activation energy of 1.9 kcal mol(-1) for temperature-dependent non-radiative deactivation of the excited state. A structural change occurs at approximately 66.7 degrees C and the activation energy increases to 10.2 kcal mol(-1). This structural change is not characterized by tryptophan exposure on the surface of the protein. The deactivation and the evolution of structural changes with time after lipase incubation at 60 degrees C were assessed by fluorescence, CD and SLS measurements. CD spectra show that both secondary and tertiary structures remain native-like after incubation at 60 degrees C in spite of the fluorescence changes observed (red-shift from 330 to 336 nm on the trytophyl emission). SLS measurements together with the CD data show that deactivation may be due to protein association between native molecules. Deactivation and the decrease on the fraction of non-associated native lipase evaluated by changes in fluorescence intensity with time, show apparent first order kinetics. According to the rate constants, fluorescence changes precede deactivation pointing to an underestimation of the deactivation. Reactivation upon dilution during the activity assay and substrate-induced reactivation due to lipase interfacial adsorption are possible causes for this underestimation. PMID- 11099175 TI - A chemical flow system mimics waves of gene expression during segmentation. AB - The early vertebrate developmental process of somitogenesis involves bands of gene expression that form periodically at the posterior end of the presomitic mesoderm (PSM) and traverse it with decreasing width and velocity. We have constructed a chemical flow system that, based on the novel flow-distributed oscillator (FDO) mechanism of wave pattern formation, reproduces key physical features of the PSM and observe concentration waves having similar spatio temporal behavior. This suggests that the gene expression waves can be understood qualitatively in terms of phase dynamics in an open flow of a self-oscillating medium and that chemical flow systems can be used to mimic and model biological pattern formation during axial growth. In fact, expressions for wavelength and wave velocity derived from phase dynamics are found to be in quantitative agreement with measurements from both the biological and the chemical systems. This indicates that they, despite their significant differences, have common dynamics. PMID- 11099176 TI - pH-sensitive liposomes as a carrier for oligonucleotides: a physico-chemical study of the interaction between DOPE and a 15-mer oligonucleotide in excess water. AB - The cytoplasmic delivery of drugs encapsulated into pH-sensitive liposomes is under the control of a lamellar-to-hexagonal transition. In a previous study, under anhydrous conditions, oligonucleotides (ODN) encapsulated in pH-sensitive liposomes composed of dioleoylphosphatidylethanolamine (DOPE)/oleic acid (OA)/cholesterol (CHOL) were shown to modify the phase behaviour of DOPE. In the present study, the lipid/ODN interactions were evaluated in fully hydrated samples by surface tension measurements, differential scanning calorimetry, X-ray diffraction and turbidimetry. Concerning the lipids, it was shown that OA provoked a disorganisation of DOPE lamellar phases and led to the complete disappearance of hexagonal transition along with heating. The addition of CHOL further decreased the lipid packing in the bilayers. Concerning ODN, these molecules provoked an increase in the surface pressure of a DOPE/OA/CHOL monolayer, indicating the existence of molecular interactions with the lipids. At a supramolecular level, ODN induced a more ordered organisation of DOPE molecules in the lamellar and hexagonal phases, and completely abolished the disorganisational effect of OA and CHOL. PMID- 11099177 TI - Step-wise formation of helical structure and side-chain packing in a peptide from scorpion neurotoxin support hierarchic model of protein folding. AB - The mechanism of protein folding has been the subject of extensive investigation during the last decade, both because of its academic challenge and because of its relation to many diseases which are known to occur due to misfolding of proteins. In this context, we report here a systematic investigation on the step-wise formation of a helical structure by the addition of hexafluoroacetone, in a 14 residue peptide derived from a part of the scorpion neurotoxin protein. The NMR and circular dichroism results indicate that the peptide has an inherent propensity for helix formation and this is limited to the internal few residues in aqueous solution. With the addition of the fluorosolvent, the helical content progressively increases and spans the whole sequence. This is accompanied by concomitant packing of the side chains. These results provide support to the so called hierarchic model of protein folding which dictates that the local sequence determines the secondary structures in the protein and the side chains play an important role in this process. PMID- 11099178 TI - Binding of human immunodeficiency virus type 1 nucleocapsid protein to psi-RNA SL3. AB - The interaction of the nucleocapsid protein NCp7, from the pNL4-3 isolate of HIV 1, with psi-RNA-SL3, with the sequence 5'-GGACUAGCGGAGGCUAGUCC, was studied using non-denaturing gel electrophoresis. Two kinds of experiments were performed, using buffered solutions of radiolabeled RNA and unlabeled protein. In the 'dilution' experiments, the total RNA concentration, RT, was varied for a series of solutions, but kept equal to the total protein concentration, PT, In the 'titration' experiments, solutions having RT constant but with varying PT were analyzed. The solutions were electrophoresed and the autoradiographic spot intensities, proportional to the amounts of the different species present, were measured. The intensities were fit to a number of equilibrium models, differing in species stoichiometries, by finding the best values of the binding constants. It was shown that NCp7 protein and SL3 RNA combine to form at least two complexes. When PT is below approximately 10 microM, a complex that contains two RNAs and one protein forms. Increasing PT to approximately 100 microM causes the 2:1 complex to oligomerize, forming a species having eight RNAs and four proteins. For the dilution experiments, run at 5 degrees C at an ionic strength of 31 mM, we found K1 for the 2:1 complex is approximately 10(11) M(-2) and K2 for the 8:4 complex is approximately 10(16) M(-3). The titration experiments returned K1 approximately 10(7) M(-2) (poorly determined) and K2 approximately 10(19) M(-3). The analysis was complicated by the loss of RNA at higher protein concentrations, due to formation of an insoluble species containing both RNA and protein, which does not enter the gel. Correcting for this changes the calculated values of equilibrium constants, but not the molecularities determined by our analysis. The observation that a small complex can oligomerize to form a larger species is consistent with the fact that NCp7 organizes and condenses the genome in the virus particle. PMID- 11099180 TI - Voltammetric and spectroscopic studies on methyl green and cationic lipid bound to calf thymus DNA. AB - DNA interaction with cationic lipids promises to be a versatile and effective synthetic transfection agent. This paper presents the study on binding of a simple artificial cationic lipid, cetyltrimethylammonium bromide (CTAB), to calf thymus DNA (CT DNA) prior to the condensation process, taking methyl green (MG) as a probe. The results show that the CTAB binds to DNA through electrostatic interaction forming a hydrophobic complex, thus changing the micro-environment of duplex of DNA, so the binding state of MG and DNA is changed, and a complex CTAB CT DNA-MG is formed. This fact suggests a new way to mediate the conformation of molecular assemblies of DNA and lipids. PMID- 11099181 TI - Analysis of negative cooperativity for glutamate dehydrogenase. AB - The empirical equation, which describes negative cooperativity in the enzyme kinetics, has been proposed. The equation is obtained from the Michaelis-Menten equation where the Michaelis constant is replaced by the effective Michaelis constant, which is a linear function of the v/Vmax ratio (v is the rate of the enzymatic reaction and Vmax is the limiting value of v at saturating concentrations of substrate). The equation allows the limiting values of the Michaelis constant at v/Vmax --> 0 and V/Vmax --> 1 to be estimated, K0 and Klim, respectively. The Klim/K0 ratio is considered as a quantitative characteristic of negative cooperativity. The applicability of the equation has been demonstrated for the kinetic data obtained for glutamate dehydrogenases from various sources (negative kinetic cooperativity for coenzyme). The negative cooperativity for the functions of saturation of protein by ligand is also analyzed. The data on binding of spin-labeled NAD, NADH, and NADPH by beef liver glutamate dehydrogenase are used as examples. PMID- 11099179 TI - Electromigration of polyion homopolymers across biomembranes: a biophysical model. AB - The analysis of polyion transmembrane translocation was performed using membrane electrical equivalent circuit. The dependence of polyion flux across membranes on time, membrane electrical conductance, membrane electrical capacitance, degree of polymerization, water solution conductance and applied transmembrane potential is discussed. The changes in polyion flux were up to 88% after 1 ms. Both the increase of polyion chain length and the decrease of membrane conductance resulted in the diminution of this effect. Inversion of flux direction was observed as a result of external potential changes. Reversal curves, representing the values of considered parameters for zero-flux were also shown. The replacement of a polyanion by a polycation of the same chain length resulted in the same shape of the surface plot but with opposite orientation. The analysis describes the effect of transmembrane potential on the translocation rate of polyanionic polysialic acid and polynucleotides, and polycationic peptides across membranes. PMID- 11099182 TI - Dynamics of compact denatured states of glutaminyl-tRNA synthetase probed by bis ANS binding kinetics. AB - Bis-ANS binds to native glutaminyl-tRNA synthetase (GlnRS) with a fast and a slow phase. The rate constant of the slow phase is independent of bis-ANS concentration suggesting a slow conformational change in the pathway of bis-ANS binding. Aging of GlnRS causes a large decrease of the slow phase amplitude with concomitant increase of the fast phase amplitude. Several other large, multi domain proteins show similar patterns upon aging. The near UV-CD spectra of the native and the aged GlnRS remain similar. Significant changes in far UV-CD, acrylamide quenching and sulfhydryl reactivity, are seen upon aging, suggesting disruptions in native interactions. Refolding of GlnRS from the urea-denatured state rapidly produces a state that is very similar to the equilibrium molten globule state. Bis-ANS binds to the molten globule state with kinetics similar to that of the aged state and unlike that of the native state. This suggests that the slow binding phase of bis-ANS, seen in native proteins, originate from relatively high energy barriers between the native and the more open states. Thus bis-ANS can be used as a powerful probe for large amplitude, low-frequency motions of proteins. PMID- 11099183 TI - Interaction of benzocaine with model membranes. AB - We measured the absorption properties, water solubility and partition coefficients (P) between n-octanol, egg phosphatidylcholine (EPC) liposomes and erythrocyte ghosts/water for benzocaine (BZC), an ester-type always uncharged local anesthetic. The interaction of BZC with EPC liposomes was followed using Electron Paramagnetic Resonance, with spin labels at different positions in the acyl chain (5, 7, 12, 16-doxylstearic acid methyl ester). Changes in lipid organization upon BZC addition allowed the determination of P values, without phase separation. The effect of BZC in decreasing membrane organization (maximum of 11.6% at approx. 0.8:1 BZC:EPC) was compared to those caused by the local anesthetics tetracaine and lidocaine. Hemolytic tests revealed a biphasic (protective/inductive) concentration-dependent hemolytic effect for BZC upon rat erythrocytes, with an effective BZC:lipid molar ratio in the membrane for protection (RePROT), onset of hemolysis (ReSAT) and 100% membrane solubilization (ReSOL) of 1.0:1, 1.1:1 and 1.3:1, respectively. The results presented here reinforce the importance of considering hydrophobic interactions in the interpretation of the effects of anesthetics on membranes. PMID- 11099184 TI - Involvement of two groups in reversal of the bathochromic shift of pharaonis phoborhodopsin by chloride at low pH. AB - Pharaonis phoborhodopsin (ppR; or pharaonis sensory rhodopsin II, psRII) is a photophobic receptor of the halobacterium Natronobacterium pharaonis. Its lambdamax is at 496 nm, but upon acidification in the absence of chloride, lambdamax shifted to 522 nm. This bathochromic shift is thought to be caused by the protonation of Asp75, which corresponds to Asp85 of bacteriorhodopsin (bR). The D75N mutant, in which Asp75 was replaced by Asn, had its lambdamax at approximately 520 nm, supporting this mechanism for the bathochromic shift. A titration of the shift yielded a pKa of 3.5 for Asp75. In the presence of chloride, the spectral shifts were different: with a decrease in pH, a bathochromic shift was first observed, followed by a hypsochromic shift on further acidification. This was interpreted as: the disappearance of a negative charge by the protonation of Asp75 was compensated by the binding of chloride, but it is worthy to note that the binding requires the protonation of another proton-associable group other than Asp75. This is supported by the observation that in the presence of chloride, upon acidification, the lambdamax of D75N even showed a blue shift, showing that the protonation of a proton-associable group (pKa = 1.2) leads to the chloride binding that gives rise to a blue shift. PMID- 11099185 TI - Medical physicists in developed countries should actively help medical physicists in developing countries. PMID- 11099186 TI - Performance of a fluorescent screen and CCD camera as a two-dimensional dosimetry system for dynamic treatment techniques. AB - A two-dimensionally position sensitive dosimetry system has been tested for different dosimetric applications in a radiation therapy facility with a scanning proton beam. The system consists of a scintillating (fluorescent) screen, mounted at the beam-exit side of a phantom and it is observed by a charge coupled device (CCD) camera. The observed light distribution at the screen is equivalent to the two-dimensional (2D)-dose distribution at the screen position. It has been found that the dosimetric properties of the system, measured in a scanning proton beam, are equal to those measured in a proton beam broadened by a scattering system. Measurements of the transversal dose distribution of a single pencil beam are consistent with dose measurements as well as with dose calculations in clinically relevant fields made with multiple pencil beams. Measurements of inhomogeneous dose distributions have shown to be of sufficient accuracy to be suitable for the verification of dose calculation algorithms. The good sensitivity and sub-mm spatial resolution of the system allows for the detection of deviations of a few percent in dose from the expected (intended or calculated) dose distribution. Its dosimetric properties and the immediate availability of the data make this device a useful tool in the quality control of scanning proton beams. PMID- 11099187 TI - Adaptive portal CT reconstruction: a simulation study. AB - In radiotherapy, radiation treatment beams contain valuable information for patient setup verification. These beams may be used for portal CT reconstruction. However, direct use of the beam data for reconstruction may yield inadequate CT images simply because these beams cover only a part of the patient body. In this study, we use the treatment beams in addition to a set of regular CT projection beams to reconstruct a locally enhanced portal CT image. This approach is called adaptive portal CT reconstruction. A computer simulation demonstrated the advantages of the approach. The image reconstruction was carried out by the multilevel scheme algebraic reconstruction technique. Results indicated that the image quality of adaptive portal CT reconstruction is equivalent to that obtained from a full set of projections. This proposed technique should be not only valuable for three-dimensional radiotherapy verification, but also applicable to diagnostic CT imaging. PMID- 11099188 TI - Dependence of linac output on the switch rate of an intensity-modulated tomotherapy collimator. AB - The electro-mechanical, multivane intensity modulated collimator ("MIMiC") slit collimator with 40 vanes has been applied in the delivery of inversely planned sequential tomotherapy to over 4,000 patients. The collimator is binary in that each vane switches between fully open or closed status. Resulting beamlet patterns provide the intensity distributions imparting dose to the patient. The bouncing and damping of vanes at the two ends of their travel cause transient dose perturbations near and at the borders of the treatment field. These perturbations are not explicitly modeled by the planning system. Clinical beamlet profiles and output factors may then differ from those in the planning system and as a function of the vane switch period. A mechanical model of vane switching was developed to describe this dependency. Dose output and distribution of seven simple vane patterns with different switch times were measured with ionization chambers and radiographic films in polystyrene and anthropomorphic phantoms. Linac output dependence on switch time relative to vane open time was determined for four intensity modulated radiotherapy (IMRT) patients from measurements of an ionization chamber embedded in a cylindrical polystyrene phantom. Results demonstrate output dependence on switch time and, accordingly, on the servo mechanism for monitor units, arc length, dose rate, and gantry speed. In conclusion, the output dependence borders on clinical significance-improvements to collimator, dose calculation, commissioning, and quality assurance (QA) are suggested. PMID- 11099189 TI - A monitor unit verification calculation in intensity modulated radiotherapy as a dosimetry quality assurance. AB - In standard teletherapy, a treatment plan is generated with the aid of a treatment planning system, but it is common to perform an independent monitor unit verification calculation (MUVC). In exact analogy, we propose and demonstrate that a simple and accurate MUVC in intensity modulated radiotherapy (IMRT) is possible. We introduce the concept of modified Clarkson integration (MCI). In MCI, we exploit the rotational symmetry of scattering to simplify the dose calculation. For dose calculation along a central axis (CAX), we first replace the incident IMRT fluence by an azimuthally averaged fluence. Second, the Clarkson integration is carried over annular sectors instead of over pie sectors. We wrote a computer code, implementing the MCI technique, in order to perform a MUVC for IMRT purposes. We applied the code to IMRT plans generated by CORVUS. The input to the code consists of CORVUS plan data (e.g., DMLC files, jaw settings, MU for each IMRT field, depth to isocenter for each IMRT field), and the output is dose contribution by individual IMRTs field to the isocenter. The code uses measured beam data for Sc, Sp, TPR, (D/MU)ref and includes effects from multileaf collimator transmission, and radiation field offset. On a 266 MHz desktop computer, the code takes less than 15 to calculate a dose. The doses calculated with the MCI algorithm agreed within +/-3% with the doses calculated by CORVUS, which uses a 1 cm x 1 cm pencil beam in dose calculation. In the present version of MCI, skin contour variations and inhomogeneities were neglected. PMID- 11099191 TI - A design for a dual assembly multileaf collimator. AB - A multileaf collimator for radiation therapy has been designed that splits each leaf bank into two vertically displaced levels with each level consisting of alternate leaves and leaf spaces. The leaves in the upper level shield the spaces in the lower level. Each level can move laterally, in the direction perpendicular to leaf motion by one leaf width. Following lateral movement of one level, the leaves align with the other level and radiation is transmitted through the collimator as multiple slit fields in a grid pattern. This transmission can be used to form an image of the external anatomy and would enable double-exposure portal images to be acquired much more rapidly than at present. These could potentially be acquired during the treatment delivery. The radiation profiles transmitted for image formation through the collimator design were investigated. Individual and grid pattern slit field profiles formed by tungsten and lead alloy collimators were measured with varying slit width, source-collimator distance, collimator-detector distance, and collimation thickness. The slit width was found to have the major influence on the transmitted profiles. As the slit width decreases the profiles become broader than the geometric slit projection resulting in increasing overlap of adjacent profiles. The overlap results in a modulated image of the external anatomy for small slit widths, rather than a sampled or "grid" image for larger widths. The shielding of this design was found to be adequate provided the leaf faces of the adjacent vertically displaced leaves are at least aligned, therefore an overlap or tongue and groove is not required. PMID- 11099190 TI - A method for determining multileaf collimator transmission and scatter for dynamic intensity modulated radiotherapy. AB - The main purpose of this work is to demonstrate a practical means of determining the leaf transmission and scatter characteristics of a multileaf collimator (MLC) pertinent to the commissioning of dynamic intensity modulated radiotherapy, especially for the sweeping window technique. The data are necessary for the conversion of intensity distributions produced by intensity-modulated radiotherapy optimization systems into trajectories of MLC leaves for dynamic delivery. Measurements are described for two, tungsten alloy MLCs: a Mark II 80 leaf MLC on a Varian 2100C accelerator and a Millenium 120-leaf MLC on a Varian 2100EX accelerator. MLC leakage was measured by film for a series of field sizes. Measured MLC leakage was 1.68% for a 10 x 10 cm2 field for both 6 and 18 MV for the 80-leaf MLC. For the 6 MV field, the 1.68% leakage consisted of 1.48% direct transmission and 0.20% leaf scatter. Direct transmission through the 80-leaf MLC, including the rounded leaf tip, was calculated analytically taking into account the detailed leaf geometry and a Monte Carlo-generated energy spectrum of the accelerator. The integrated fluence under the leaf tip was equivalent to an inward shift of 0.06 cm of a hypothetical leaf with a flat, focused tip. Monte Carlo calculations of the dose to phantom beyond a closed 80-leaf MLC showed excellent agreement with the analytic results. The transmission depends on the density of the MLC alloy, which may differ among individual MLCs. Thus, it is important to measure the transmission of any particular MLC. Calculated doses for a series of uniform fields produced by dynamic sweeping windows of various widths agree with measurements within 2%. PMID- 11099192 TI - Comparison of measured and Monte Carlo calculated dose distributions from the NRC linac. AB - We have benchmarked photon beam simulations with the EGS4 user code BEAM [Rogers et al., Med. Phys. 22, 503-524 (1995)] by comparing calculated and measured relative ionization distributions in water from the 10 and 20 MV photon beams of the NRC linac. Unlike previous calculations, the incident electron energy is known independently to 1%, the entire extra-focal radiation is simulated, and electron contamination is accounted for. The full Monte Carlo simulation of the linac includes the electron exit window, target, flattening filter, monitor chambers, collimators, as well as the PMMA walls of the water phantom. Dose distributions are calculated using a modified version of the EGS4 user code DOSXYZ which additionally allows scoring of average energy and energy fluence in the phantom. Dose is converted to ionization by accounting for the (L/rho)water(air) variation in the phantom, calculated in an identical geometry for the realistic beams using a new EGS4 user code, SPRXYZ. The variation of (L/rho)water(air) with depth is a 1.25% correction at 10 MV and a 2% correction at 20 MV. At both energies, the calculated and the measured values of ionization on the central axis in the buildup region agree within 1% of maximum ionization relative to the ionization at 10 cm depth. The agreement is well within statistics elsewhere. The electron contamination contributes 0.35(+/- 0.02) to 1.37(+/- 0.03)% of the maximum dose in the buildup region at 10 MV and 0.26(+/- 0.03) to 3.14(+/- 0.07)% of the maximum dose at 20 MV. The penumbrae at 3 depths in each beam (in g/cm2), 1.99 (dmax, 10 MV only), 3.29 (dmax, 20 MV only), 9.79 and 19.79, agree with ionization chamber measurements to better than 1 mm. Possible causes for the discrepancy between calculations and measurements are analyzed and discussed in detail. PMID- 11099194 TI - Dosimetric characteristics of a new 125I brachytherapy source. AB - 125I brachytherapy sources are being used for interstitial implants in tumor sites such as the prostate. Recently, a new 125I source has been introduced, which has a design different from that of other sources presently on the market. Dosimetric characteristics of this source, including dose rate constant, radial dose function, and anisotropy function, were determined experimentally following the AAPM Task Group 43 recommendations. The characteristics were related to the 1999 NIST calibration assigned to this source [SK,99std]. Measurements were performed in a solid water phantom using LiF thermoluminescent dosimeters. For these measurements, slabs of solid water phantom material were machined to accommodate the source and LiF TLD chips of dimensions (3.1 x 3.1 x 0.8 mm3) and (1.0 x 1.0 x 1.0 mm3). The TLD chips were surrounded by at least 10 cm of solid water phantom material to provide full scattering conditions. The results indicated a dose rate constant, lambda, of 0.88 +/- 0.07cGyh(-1)U(-1) for the new 1251 source as compared to 0.98 and 1.04 cGy h(-1)U(-1) for the Nycomed/Amersham model 6711 and 6702 seeds, respectively. Per TG-43, the values reported here represent the dose absorbed by water at 1 cm from the source in a water medium. The radial dose function, g(r), of the new 125I source was measured at distances ranging from 0.5 to 10 cm. The anisotropy function, F(r,theta), of the new 125I source was measured at distances of 2 and 5 cm from the source center. Calculations of anisotropy and radial dose function were also made using a Monte Carlo code. These calculations were made for both solid water and liquid water, the former to validate the Monte Carlo code and the latter to provide results in liquid water for clinical use. All data compared favorably with those from the Nycomed/Amersham models 6711 and 6702 sources. PMID- 11099193 TI - Comparison of dosimetric characteristics of Siemens virtual and physical wedges. AB - Dosimetric properties of Virtual Wedge (VW) and physical wedge (PW) in 6 and 23 MV photon beams from a Siemens Primus linear accelerator, including wedge factors, depth doses, dose profiles, peripheral doses and surface doses, are compared. While there is a great difference in absolute values of wedge factors, VW factors (VWFs) and PW factors (PWFs) have a similar trend as a function of field size. PWFs have a stronger depth dependence than VWF due to beam hardening in PW fields. VW dose profiles in the wedge direction, in general, match very well with PW, except in the toe area of large wedge angles with large field sizes. Dose profiles in the nonwedge direction show a significant reduction in PW fields due to off-axis beam softening and oblique filtration. PW fields have significantly higher peripheral doses than open and VW fields. VW fields have similar surface doses as the open fields while PW fields have lower surface doses. Surface doses for both VW and PW increase with field size and slightly with wedge angle. For VW fields with wedge angles 45 degrees and less, the initial gap up to 3 cm is dosimetrically acceptable when compared to dose profiles of PW. VW fields in general use less monitor units than PW fields. PMID- 11099195 TI - A dose-volume histogram based optimization algorithm for ultrasound guided prostate implants. AB - The task of treatment planning for prostate implants is to find an optimal seed configuration, comprising the target coverage and dosimetric consideration of critical structures such as the rectum and urethra. An efficient method to accomplish this is to use an inverse planning technique that derives the optimized solution from a prescribed treatment goal. The goal can be specified in the voxel domain as the desired doses to the voxels of the target and critical structures, or in the dose volume representation as the desired dose volume histograms (DVHs) of the target and critical structures. The DVH based optimization has been successfully used in plan optimization for intensity modulated radiation therapy (IMRT) but little attention has been paid to its application in prostate implants. Clinically, it has long been known that some normal structure tolerances are more accurately assessed by volumetric information. Dose-volume histograms are also widely used for plan evaluation. When working in the DVH domain for optimization one has more control over the final DVHs. We have constructed an objective function sensitive to the DVHs of the target and critical structures. The objective function is minimized using an iterative algorithm, starting from a randomly selected initial seed configuration. At each iteration step, a trial position is given to a randomly selected source and the trial position is accepted if the objective function is decreased. To avoid being trapped in a less optimal local minimum, the optimization process is repeated. The final plan is selected from a pool of optimized plans obtained from a series of randomized initial seed configurations. PMID- 11099196 TI - Well-ionization chamber response relative to NIST air-kerma strength standard for prostate brachytherapy seeds. AB - The response of well-ionization chambers to the emissions of 103Pd and 125I radioactive seed sources used in prostate cancer brachytherapy has been measured. Calibration factors relating chamber response (current or dial setting) to measured air-kerma strength have been determined for seeds from nine manufacturers, each with different designs. Variations in well-ionization chamber response relative to measured air-kerma strength have been observed because of differences in the emitted energy spectrum due to both the radionuclide support material (125I seeds) and the mass ratio of 103Pd to 102Pd (103Pd seeds). Obtaining accurate results from quality assurance measurements using well ionization chambers at a therapy clinic requires knowledge of such differences in chamber response as a function of seed design. PMID- 11099197 TI - In vivo urethral dose measurements: a method to verify high dose rate prostate treatments. AB - Radiation doses delivered in high dose rate (HDR) brachytherapy are susceptible to many inaccuracies and errors, including imaging, planning and delivery. Consequently, the dose delivered to the patient may deviate substantially from the treatment plan. We investigated the feasibility of using TLD measurements in the urethra to estimate the discrepancy in treatments for prostate cancer. The dose response of the 1 mm diam, 6 mm long LiF rods that we used for the in vivo measurements was calibrated with the 192Ir HDR source, as well as a 60Co teletherapy unit. A train of 20 rods contained in a sterile plastic tube was inserted into the urethral (Foley) catheter for the duration of a treatment fraction, and the measured doses were compared to the treatment plan. Initial results from a total of seven treatments in four patients show good agreement between theory and experiment. Analysis of any one treatment showed agreement within 11.7% +/- 6.2% for the highest dose encountered in the central prostatic urethra, and within 10.4% +/- 4.4% for the mean dose. Taking the average over all seven treatments shows agreement within 1.7% for the maximum urethral dose, and within 1.5% for the mean urethral dose. Based on these initial findings it seems that planned prostate doses can be accurately reproduced in the clinic. PMID- 11099198 TI - Fitting and benchmarking of dosimetry data for new brachytherapy sources. AB - New source designs of encapsulated low-energy gamma emitting nuclides for permanent implants require dosimetric analysis and calibration standardization. The dosimetry measurements can be incorporated into a treatment planning system by fitting the data. The use of a fitting function whose behavior at range limits mimics the physical phenomena, using as few parameters as possible, eliminates noisy outliers and lends credence to calculations beyond the measured range. Clinical implementation of the new sources also requires benchmarking against existing sources, where the current clinical experience lies. We present an analysis of measured dosimetry data for three brachytherapy sources recently available from North American Scientific, Inc. (North Hollywood, CA): 103Pd source model MED3633 ("PdGold"), 125I source models MED3631-A/M ("IoGold-AM") and MED3631-A/S ("IoGold-AS"). Using the formalism of the Interstitial Collaborative Working Group (ICWG) the radial dose function, g(r), the anisotropy function, F(r, theta), and the anisotropy factor, phi an(r), were previously evaluated from measurements of each source design. In this report we use fitting functions whose forms are chosen to approach reasonable values at data limits. These forms are quite similar to those used in a previous analysis of TG43 Iodine and palladium compendium data. Fitting parameter results for each function are provided for each brachytherapy source model. Fit-data discrepancies are smaller than measurement uncertainties, meaning that incorporation into treatment planning systems will not introduce significant errors in clinical use. Current clinical experience is based on the Theragenics (Norcross, GA) 103Pd seed ("PdThera"), and the Nycomed-Amersham (Arlington Heights, IL) 125I seed models 6711 ("6711") and 6702 ("6702"). The new sources are benchmarked against these seeds. PMID- 11099199 TI - Dosimetric modeling of the microselectron high-dose rate 192Ir source by the multigroup discrete ordinates method. AB - The DANTSYS multigroup discrete ordinates computer code is applied to quantitatively estimate the absorbed dose rate distributions in the vicinity of a microSelectron 192Ir high-dose-rate (HDR) source in two-dimensional cylindrical R Z geometry. The source is modeled in a cylindrical water phantom of diameter 20 cm and height 20 cm. The results are also used for evaluation of the Task Group 43 (TG-43) dosimetric quantities. The DANTSYS accuracy is estimated by direct comparisons with corresponding Monte Carlo results. Our 210-group photon cross section library developed previously, together with angular quadratures consisting of 36 (S16) to 210 (S40) directions and associated weights per octant, are used in the DANTSYS simulations. Strong ray effects are observed but are significantly mitigated through the use of DANTSYS's stochastic ray-tracing first collision source algorithm. The DANTSYS simulations closely approximate Monte Carlo estimates of both direct dose calculations and TG-43 dosimetric quantities. The discrepancies with S20 angular quadrature (55 directions and weights per octant) or higher are shown to be less than +/- 5% (about 2.5 standard deviations of Monte Carlo calculations) everywhere except for limited regions along the Z axis of rotational symmetry, where technical limitations in the DANTSYS first collision source implementation makes adequate suppression of ray effects difficult to achieve. The efficiency of DANTSYS simulations is compared with that of the EGS4 Monte Carlo code. It is demonstrated that even with the 210-group cross section library, DANTSYS achieves two-fold efficiency gains using the the S20 quadrature set. The potential of discrete ordinates method for further efficiency improvements is also discussed. PMID- 11099200 TI - The collapsed cone superposition algorithm applied to scatter dose calculations in brachytherapy. AB - Methods for scatter dose calculations in brachytherapy have been developed based on the collapsed cone superposition algorithm. The methods account for effects on the scatter dose caused by the three-dimensional distribution of heterogeneities in the irradiated volume and are considerably faster than methods based on straightforward superposition of kernels or direct Monte Carlo simulations. Use of a successive-scattering approach, in which the dose contribution from once- and multiply scattered photons are calculated separately, was found superior to conventional superposition using a single point kernel for all scatter generations. Use of the successive-scattering approach significantly reduces artifacts stemming from steep fluence gradients, typical of the brachytherapy geometry and critical for the collapsed cone approximation. The algorithm is tested versus Monte Carlo simulations for point sources of energies 28.4, 100, 350, and 662 keV. Results agree well for both a homogeneous water phantom and an air-water half-phantom. PMID- 11099201 TI - Optimized bounding boxes for three-dimensional treatment planning in brachytherapy. AB - It is sometimes necessary to determine the optimal value for a direction dependent quantity. Using a search technique based on Powell's quadratic convergent method such an optimal direction can be approximated. The necessary geometric transformations in n-dimensional space are introduced. As an example we consider the approximation of the minimum bounding box of a set of three dimensional points. Minimum bounding boxes can significantly improve accuracy and efficiency of the calculations in modern brachytherapy treatment planning of the volumes of objects or the dose distribution inside an object. A covariance matrix based approximation method for the minimum bounding box is compared with the results of the search method. The benefits of the use of optimal oriented bounding boxes in brachytherapy treatment planning systems are demonstrated and discussed. PMID- 11099202 TI - Dose-volume histograms computation comparisons using conventional methods and optimized fast Fourier transforms algorithms for brachytherapy. AB - In anatomy based optimization procedures for large volume implants the calculation of dose-volume histograms (DVH) accounts for the major part of the time involved and can be as long as a few hours. This time is proportional to the number of seeds or source dwell positions required for the implant. A procedure for the calculation of brachytherapy seed dose distribution calculation employing fast Fourier transforms (FFT) and the convolution theorem has been described by others and was supposed to significantly improve the speed of the dose distribution computation. Using new significantly improved FFT algorithms and various other optimization techniques we have compared the calculated differential and integral DVHs in high dose rate (HDR) brachytherapy with a single stepping source using actual clinical implants. This is so that we could assess the efficiency and accuracy of the FFT method with that of conventional methods. Our results showed that the FFT based method of calculating DVHs in brachytherapy is comparable in speed with conventional dose calculation methods, but only for implants with more than 287 sources. It is therefore of limited practical use even for large implants. This result is in direct opposition to the claim by other authors. PMID- 11099203 TI - Measurement of the dose components of fast and thermal neutrons and photons from a 0.1 mg 252Cf source in water for brachytherapy treatment planning. AB - The use of 252Cf in brachytherapy is expected to be more effective with the therapy of bulky tumors than the conventional therapy with photons. For treatment planning a code developed for calculation of gamma dose was used to generate the dose distributions of fast and 10B enhanced thermal neutrons and photons. Dose distributions of these components measured with ionization chambers and a GM counter were fitted to analytical functions as required by the modified treatment planning program. A comparison of these experimental results and the treatment planning output indicate good agreement. Therefore, the program may be used to optimize the brachytherapy procedure considering all three dose components. A realistic case of a patient being treated with conventional brachytherapy has been used to calculate the dose distribution that would be obtained by use of the 252Cf source. PMID- 11099204 TI - Determination of the air w-value in proton beams using ionization chambers with gas flow capability. AB - The purpose of this work was to determine the w-value of air for protons using the paired gas method. Several plastic- and magnesium-walled chambers were used with air, synthetic air, nitrogen, and argon flowing gases. Using argon as a reference gas, the w-value of air was measured and ranged from 32.7 to 34.5 J/C for protons with energies encountered in radiotherapy. Using nitrogen as a reference gas, the w-value of air ranged from 35.2 to 35.4 J/C over the same range of proton energies. The w-value was found, at a given energy, to be independent of the ion chamber used. The uncertainty in these measurements was estimated at 5.2% at the 2sigma level. This uncertainty was dominated by the 4.4% uncertainty in the w-value of the reference gas. PMID- 11099206 TI - Determination of effective electron source size using multislit and pinhole cameras. AB - Two independent methods have been utilized for determination of effective source sizes for 6, 12, and 20 MeV electron beams generated by a Varian 2100C linear accelerator. First, a multislit camera has been constructed using parallel aluminum plates and plastic strip spacers, similar to the beam-spot camera for the photon source imaging. Second, pinhole imaging was performed using a lead plate with a small hole on the central axis of the beam. The plate thickness and the hole diameter varied with electron energy. The cameras were positioned directly at the opening of the movable photon collimator. The size of the source distribution from each camera was characterized by its full width at half-maximum (FWHM) value. The measured values of FWHM are different for each camera because of their different imaging principles. For the multislit camera, the measured FWHM values were (6.3 +/- 0.4) cm for the 6 MeV beam, (3.6 +/- 0.4) cm for 12 MeV, and (2.7 +/- 0.4) cm for 20 MeV. For the pinhole camera the measured values of FWHM were (7.9 +/- 0.6) cm for 6 MeV, (4.5 +/- 0.4) cm for 12 MeV, and (3.0 +/ 0.4) cm for the 20 MeV beam. Additionally, the effective source position was derived from output measurements at different values of the SSD, which were fitted to the inverse square law. PMID- 11099205 TI - Confining proton beams with longitudinal magnetic fields: Monte Carlo calculations. AB - The problem of the lateral containment of a 160 MeV proton beam interacting with a medium simulating biological material was studied. The confining action of a longitudinal magnetic field was calculated by means of Monte Carlo simulation, where scattering and motion in the magnetic field were treated simultaneously. Appreciable compression of the beam could only be achieved using fields of the order at least 50 T, much beyond the realm of practical feasibility. PMID- 11099207 TI - A mathematical study of the area over perimeter rule using the sector-integration equation. AB - The equivalence between rectangular photon fields and square fields is currently used to simplify tabulation and handling of data as well as to reduce measurement time. Widely used for routine calculation in the case of rectangular fields of moderate elongation, the so-called "area over perimeter rule" (A/P rule) has a remarkable accuracy. Several approaches have been developed to determine the physical and mathematical grounds of this rule, yet the statement that it is independent of depth and energy was not fully clarified. By means of the Clarkson sector-integration equation and Taylor expansion, this work demonstrates that the A/P rule is a first-order approximation on the elongation variable for all cases, i.e., for moderate rectangular field elongation presents a quadratic deviation. To appreciate the degree of approximation of this rule, a model scatter air ratio for Co60 y-rays at 10.0 cm depth was used to compute rectangular radiation fields and the results were compared to those given by the A/P rule. The model scatter air ratio was the proposed by Day and Aird [Br. J. Radiol. 25, 138-151 (1996)]. PMID- 11099208 TI - Shielding considerations for tomotherapy. AB - Tomotherapy presents an evolutionary modality that holds forth the promise of better dose conformation to tumor volumes with a concomitant reduction in radiation-induced damage to surrounding normal structures. This delivery technique also presents a new set of radiation protection challenges that impact upon the design of the shielding vault required to house such a unit. A formalism is presented to determine the requisite amounts of shielding for both the primary beam and leakage radiation associated with a generic tomotherapy unit. A comparison is made with the shielding requirements for a conventional linear accelerator operated in a standard manner. Substantial differences in the amount of both primary and secondary shielding are indicated. A tomotherapy primary beam shield is both reduced in width by a factor of almost 10 and increased in thickness by more than a tenth value layer in comparison to a conventional accelerator. Furthermore, the secondary shielding requirements are enhanced by more than two tenth value layers with respect to conventional shielding demands. PMID- 11099209 TI - Predictions of a stochastic model of bone marrow cell survival in high dose rate radiation fields with arbitrary neutron to gamma-ray absorbed dose rate ratios. AB - In this paper, a stochastic model of cell survival, which was developed by Cotlet and Blue, based on the work of Jones, is extended to describe bone marrow cell survival in high dose rate radiation fields with arbitrary neutron to gamma-ray absorbed dose rate ratios. Mathematical formulas are obtained that describe the interaction of the neutron and gamma-ray components of the absorbed dose, for radiation fields with arbitrary neutron to gamma-ray dose rate ratios, for exposures of cells to various absorbed doses, at various high dose rates. PMID- 11099210 TI - Monte Carlo assessment of computed tomography dose to tissue adjacent to the scanned volume. AB - The assessment of the radiation dose to internal organs or to an embryo or fetus is required on occasion for risk assessment or for comparing imaging studies. Limited resources hinder the ability to accurately assess the radiation dose received to locations outside the tissue volume actually scanned during computed tomography (CT). The purpose of this study was to assess peripheral doses and provide tabular data for dose evaluation. Validated Monte Carlo simulation techniques were used to compute the dose distribution along the length of water equivalent cylindrical phantoms, 16 and 32 cm in diameter. For further validation, comparisons between physically measured and Monte Carlo-derived air kerma profiles were performed and showed excellent (1% to 2%) agreement. Polyenergetic x-ray spectra at 80, 100, 120, and 140 kVp with beam shaping filters were studied. Using 10(8) simulated photons input to the cylinders perpendicular to their long axis, line spread functions (LSF) of the dose distribution were determined at three depths in the cylinders (center, mid-depth, and surface). The LSF data were then used with appropriate mathematics to compute dose distributions along the long axis of the cylinder. The dose distributions resulting from helical (pitch = 1.0) scans and axial scans were approximately equivalent. Beyond about 3 cm from the edge of the CT scanned tissue volume, the fall-off of radiation dose was exponential. A series of tables normalized at 100 milliampere seconds (mAs) were produced which allow the straight-forward assessment of dose within and peripheral to the CT scanned volume. The tables should be useful for medical physicists and radiologists in the estimation of dose to sites beyond the edge of the CT scanned volume. PMID- 11099211 TI - Scatter/primary in mammography: comprehensive results. AB - Monte Carlo procedures using the SIERRA code (validated in a companion article) were used to investigate the scatter properties in mammography. The scatter to primary ratio (SPR) was used for quantifying scatter levels as a function of beam spectrum, position in the field, air gap, breast thickness, tissue composition, and the area of the field of view (FOV). The geometry of slot scan mammography was also simulated, and SPR values were calculated as a function of slot width. The influence of large air gaps (to 30 cm) was also studied in the context of magnification mammography. X-ray energy and tissue composition from 100% adipose to 100% glandular demonstrated little effect on the SPR. Air gaps over a range from 0 to 30 mm showed only slight effects. The SPR increased with increased breast thickness and with larger fields of view. Measurements from 82 mammograms provided estimates of the range of compressed breast thickness (median: 5.2 cm, 95% range: 2.4 cm to 7.9 cm) and projected breast area onto the film (left craniocaudal view, median: 146 cm2, 95% range: 58 cm2 to 298 cm2). SPR values for semicircular breast shapes, Mo/Mo spectra, and a 15 mm air gap were parametrized as a function of breast thickness and (semicircular) breast diameter. With the coefficients a = - 2.35452817439093, b = 22.3960980055927, and c = 8.85064260299289, the equation SPR= [a + b x (diameter in cm)--(-1.5) + c x (thickness in cm) --(-0.5)]-- -1 produces SPR data from 2 to 8 cm and from 3 to 30 cm breast diameters with an average error of about 1%. PMID- 11099212 TI - Linear response theory for detectors consisting of discrete arrays. AB - The optical transfer function (OTF) and the noise power or Wiener spectrum are defined for detectors consisting of a lattice of discrete elements with the assumptions of linear response, Gaussian statistics, and stationarity under the discrete group of translations which leave the lattice fixed. For the idealized classification task of determining the presence or absence of a signal under signal known exactly/background known exactly (SKE/BKE) conditions, the Wiener spectrum, the OTF, along with an analog of the gray-scale transfer characteristic, determine the signal-to-noise ratio (SNR), which quantifies the ability of an ideal observer to perform this task. While this result is similar to the established result for continuous detectors, such as screen-film systems, the theory of discrete lattices of detectors must take into account the fact that the lattice only supports a bounded but (in the limit of a detector of arbitrarily great extent) continuous range of frequencies. Incident signals with higher spatial frequencies appear in the data at lower aliased frequencies, and there are pairs of signals which are not distinguishable by the detector (the SNR vanishes for the task of distinguishing such signals). Further, the SNR will in general change if the signal is spatially displaced by a fraction of the lattice spacing, although this change will be small for objects larger than a single pixel. Some of the trade-offs involved in detectors of this sort, particularly in dealing with signal frequencies above those supported by the lattice, are studied in a simple model. PMID- 11099214 TI - Simulating coronary arteries in x-ray angiograms. AB - Clinical validation of quantitative coronary angiography (QCA) algorithms is difficult due to the lack of a simple alternative method for accurately measuring in vivo vessel dimensions. We address this problem by embedding simulated coronary artery segments with known geometry in clinical angiograms. Our vessel model accounts for the profile of the vessel, x-ray attenuation in the original background, and noise in the imaging system. We have compared diameter measurements of our computer simulated arteries with measurements of an x-ray Telescopic-Shaped Phantom (XTSP) with the same diameters. The results show that for both uniform and anthropomorphic backgrounds there is good agreement in the measured diameters of XTSP compared to the simulated arteries (Pearson's correlation coefficient 0.99). In addition, the difference in accuracy and precision of the true diameter measures compared to the XTSP and simulated artery diameters was small (mean absolute error across all diameters was < or = 0.11 mm +/- 0.09 mm). PMID- 11099213 TI - Design of a dual CCD configuration to improve the signal-to-noise ratio. AB - The noise of a digital charge coupled device (CCD) detector increases with the readout speed, causing problems in a number of important applications, such as x ray fluoroscopy and micro-CT. In this paper, we present an approach for the design of a dual-CCD configuration to improve the average signal-to-noise ratio, and hence provide an inexpensive solution within the constraint of the current technology. PMID- 11099216 TI - Comparison of rigid and elastic matching of dynamic magnetic resonance mammographic images by mutual information. AB - The present study aims at (i) the evaluation of the performance of a rigid and of an elastic matching algorithm for the coregistration of dynamic magnetic resonance (MR) images visualizing the female breast, and (ii) the evaluation of the mutual information (MI) as a matching criterion. To this end, ten patient data sets were analyzed. The comparison was performed with respect to the achieved increase in the MI and by visual inspection of the dynamic image series in a continuous film sequence ((cine mode). In most cases, the achieved increase in MI by elastic image registration is much higher than that achieved by rigid registration. Only for three of the ten data sets could the MI be increased by rigid image registration to a similar or even larger degree than by elastic image registration. Taking into account the results of the visual inspection of the rigid and elastic matched data sets, however, the elastic match leads to equal or better results for all data sets. Therefore, elastic matching is the gold standard for the registration of dynamic MR mammographic images of the female breast. The comparison of the increase in MI and the visual inspection further shows that the visual impression agrees in most cases with the result of the calculation of the MI. Therefore, the MI proves to be a suitable matching criterion for the type of data sets studied. PMID- 11099215 TI - Automatic segmentation of lung fields in chest radiographs. AB - The delineation of important structures in chest radiographs is an essential preprocessing step in order to automatically analyze these images, e.g., for tuberculosis screening support or in computer assisted diagnosis. We present algorithms for the automatic segmentation of lung fields in chest radiographs. We compare several segmentation techniques: a matching approach; pixel classifiers based on several combinations of features; a new rule-based scheme that detects lung contours using a general framework for the detection of oriented edges and ridges in images; and a hybrid scheme. Each approach is discussed and the performance of nine systems is compared with interobserver variability and results available from the literature. The best performance is obtained by the hybrid scheme that combines the rule-based segmentation algorithm with a pixel classification approach. The combinations of two complementary techniques leads to robust performance; the accuracy is above 94% for all 115 images in the test set. The average accuracy of the scheme is 0.969 +/- 0.0080, which is close to the interobserver variability of 0.984 +/- 0.0048. The methods are fast, and implemented on a standard PC platform. PMID- 11099217 TI - Validation of a precision radiochromic film dosimetry system for quantitative two dimensional imaging of acute exposure dose distributions. AB - We present an evaluation of the precision and accuracy of image-based radiochromic film (RCF) dosimetry performed using a commercial RCF product (Gafchromic MD-55-2, Nuclear Associates, Inc.) and a commercial high-spatial resolution (100 microm pixel size) He-Ne scanning-laser film-digitizer (Personal Densitometer, Molecular Dynamics, Inc.) as an optical density (OD) imaging system. The precision and accuracy of this dosimetry system are evaluated by performing RCF imaging dosimetry in well characterized conformal external beam and brachytherapy high dose-rate (HDR) radiation fields. Benchmarking of image based RCF dosimetry is necessary due to many potential errors inherent to RCF dosimetry including: a temperature-dependent time evolution of RCF dose response; nonuniform response of RCF; and optical-polarization artifacts. In addition, laser-densitometer imaging artifacts can produce systematic OD measurement errors as large as 35% in the presence of high OD gradients. We present a RCF exposure and readout protocol that was developed for the accurate dosimetry of high dose rate (HDR) radiation sources. This protocol follows and expands upon the guidelines set forth by the American Association of Physicists in Medicine (AAPM) Task Group 55 report. Particular attention is focused on the OD imaging system, a scanning-laser film digitizer, modified to eliminate OD artifacts that were not addressed in the AAPM Task Group 55 report. RCF precision using this technique was evaluated with films given uniform 6 MV x-ray doses between 1 and 200 Gy. RCF absolute dose accuracy using this technique was evaluated by comparing RCF measurements to small volume ionization chamber measurements for conformal external-beam sources and an experimentally validated Monte Carlo photon transport simulation code for a 192Ir brachytherapy source. Pixel-to-pixel standard deviations of uniformly irradiated films were less than 1% for doses between 10 and 150 Gy; between 1% and 5% for lower doses down to 1 Gy and 1% and 1.5% for higher doses up to 200 Gy. Pixel averaging to form 200-800 microm pixels reduces these standard deviations by a factor of 2 to 5. Comparisons of absolute dose show agreement within 1.5%-4% of dose benchmarks, consistent with a highly accurate dosimeter limited by its observed precision and the precision of the dose standards to which it is compared. These results provide a comprehensive benchmarking of RCF, enabling its use in the commissioning of novel HDR therapy sources. PMID- 11099219 TI - Novel human topoisomerase I inhibitors, topopyrones A, B, C and D. II. Structure elucidation. AB - The structures of novel topoisomerase I inhibitors, topopyrones A, B, C and D were elucidated by spectral analysis of the chemical derivatives. These compounds are an anthraquinone type containing a fused 1,4-pyrone moiety. Topopyrones A and B contain a chlorine atom, however C and D do not. It was suggested that topopyrones B and D are converted from topopyrones A and C, respectively by Wessely-Moser type rearrangement. PMID- 11099218 TI - Novel human topoisomerase I inhibitors, topopyrones A, B, C and D. I. Producing strain, fermentation, isolation, physico-chemical properties and biological activity. AB - In the course of a screening program for specific inhibitors of human topoisomerase I using a recombinant yeast, we have discovered four new active compounds. All four compounds were isolated from the culture broth of a fungus, Phoma sp. BAUA2861, and two of them were isolated from the culture broth of a fungus, Penicillium sp. BAUA4206. We designated these compounds as topopyrones A, B, C and D. Topopyrones A, B, C and D selectively inhibited recombinant yeast growth dependent on expression of human topoisomerase I with IC50 values of 1.22, 0.15, 4.88 and 19.63 ng/ml, respectively. The activity and selectivity of topopyrone B were comparable to those of camptothecin. The relaxation of supercoiled pBR322 DNA by human DNA topoisomerase I was inhibited by these compounds, however they did not inhibit human DNA topoisomerase II. Topopyrones A, B, C and D were cytotoxic to all tumor cell lines when tested in vitro. Topopyrone B has potent inhibitory activity against herpesvirus, especially varicella zoster virus (VZV). It inhibited VZV growth with EC50 value of 0.038 microg/ml, which is 24-fold stronger than that of acyclovir (0.9 microg/ml). Topopyrones A, B, and C were inhibitory to Gram-positive bacteria. PMID- 11099224 TI - FR901469, a novel antifungal antibiotic from an unidentified fungus No.11243. I. Taxonomy, fermentation, isolation, physico-chemical properties and biological properties. AB - FR901469 is a novel antifungal antibiotic produced by an unidentified fungus No.11243. This compound was isolated from the culture broth by solvent extraction, HP-20 and YMC ODS gel column chromatography, and lyophilization. FR901469 is a white powder which melts at 182 approximately 187 degrees C and possesses the molecular formula C71H116N14O23. This compound has good water solubility. FR901469 inhibited the activity of 1,3-beta-glucan synthase from Candida albicans with an IC50 value of 0.05 microg/ml, and displayed greater inhibitory activity than other 1,3-beta-glucan synthase inhibitors such as, WF11899A, echinocandin B, aculeacin A, and papulacandin B. PMID- 11099220 TI - Tubulysins, new cytostatic peptides from myxobacteria acting on microtubuli. Production, isolation, physico-chemical and biological properties. AB - New cytostatic compounds, tubulysins, were isolated from the culture broth of strains of the myxobacteria Archangium gephyra and Angiococcus disciformis. The compounds are peptides partly consisting of unusual amino acids and are distantly related to the dolastatins. The tubulysins were not active against bacteria and only little against fungi, but showed high cytostatic activity against mammalian cell lines with IC50 values in the picomolar range. An incubation with 50 ng/ml tubulysin A led to a complete disappearance of the microtubuli network of the cells within 24 hours. The more active tubulysin D induced multipolar spindles: At 0.5 ng/ml all mitotic cells showed more than four spindle poles. PMID- 11099221 TI - New triene-ansamycins, thiazinotrienomycins F and G and a diene-ansamycin, benzoxazomycin. AB - New triene-ansamycins designated thiazinotrienomycins F (TT-F) and G (TT-G) and a new diene-ansamycin, benzoxazomycin, were isolated from a culture broth of Streptomyces sp. MJ672-m3 and their structures were elucidated by spectroscopic analyses. The Mean Graphs of TT-G suggests that the tumor growth inhibitory activities are almost as strong as TT-B, in respect of GI50 and TGI against several human cancer cell lines. PMID- 11099222 TI - 4'-N-methyl-5'-hydroxystaurosporine and 5'-hydroxystaurosporine, new indolocarbazole alkaloids from a marine Micromonospora sp. strain. AB - Two new indolocarbazole alkaloids, 4'-N-methyl-5'-hydroxystaurosporine (2) and 5' hydroxystaurosporine (3), were isolated together with the known staurosporine (1) from the culture broth of a marine Micromonospora sp. (strain L-31-CLCO-002). The fermentation, structural data and cytotoxic activities of these compounds against various tumor cell lines are given. PMID- 11099225 TI - FR901469, a novel antifungal antibiotic from an unidentified fungus No.11243. II. In vitro and in vivo activities. AB - FR901469 is a water-soluble macrocyclic lipopeptidolactone (C71H116N14O23) that has inhibitory activity against 1,3-beta-glucan synthase and exhibits in vitro and in vivo antifungal activity against both Candida albicans and Aspergillus fumigatus. The MICs of FR901469 against Candida albicans FP633 and Aspergillus fumigatus FP1305 in a micro-broth dilution test were 0.63 and 0.16 microg/ml, respectively. FR901469 showed excellent efficacy by subcutaneous injection against both Candida albicans and Aspergillus fumigatus in a murine systemic infection mode, with ED50s of 0.32 and 0.2 mg/kg, respectively. This compound also showed potent anti-Pneumocystis activity in the nude mice model with experimental Pneumocystis pneumonia. The hemolytic activity of FR901469 towards mouse red blood cells, is about 30-fold weaker than that of amphotericin B. PMID- 11099223 TI - Phellinsin A, a novel chitin synthases inhibitor produced by Phellinus sp. PL3. AB - Phellinsin A, a novel chitin synthases inhibitor was isolated from the cultured broth of fungus PL3, which was identified as Phellinus sp. PL3. Phellinsin A was purified by solvent partition, silica gel, ODS column chromatographies, and preparative HPLC, consecutively. The structure of phellinsin A was assigned as a phenolic compound on the basis of various spectroscopic analyses including UV, IR, Mass, and NMR. Its molecular weight and formula were found to be 358 and C18H14O8, respectively. Phellinsin A selectively inhibited chitin synthase I and II of Saccharomyces cerevisiae with an IC50 value of 76 and 28 microg/ml, respectively, in our cell free assay system. This compound showed antifungal activity against Colletotrichum lagenarium, Pyricularia oryzae, Rhizoctonia solani, Aspergillus fumigatus, and Trichophyton mentagrophytes. PMID- 11099226 TI - Xanthoepocin, a new antibiotic from Penicillium simplicissimum IFO5762. AB - A new antifungal antibiotic xanthoepocin was isolated from the culture broth of Penicillium simplicissimum IFO5762. Xanthoepocin was obtained from the culture fluid by solvent extraction and chromatographic purification. It showed antibiotic activity against Gram-positive bacteria and yeasts. PMID- 11099227 TI - Feigrisolides A, B, C and D, new lactones with antibacterial activities from Streptomyces griseus. AB - Four new lactone compounds, named feigrisolides A to D (1 to 4), have been isolated from Streptomyces griseus. The chemical structures were determined by detail analysis of their spectroscopic data and chemical transformations. Structurally, the feigrisolides A (1) and B (2) are hepta-lactones, feigrisolide C (3) and D (4) are 16-membered macrodiolides. Biological studies showed that feigrisolide B (2) exhibited strong antibacterial, as well as medium cyctotoxic, and antiviral activities. Feigrisolides A (1), C (3) and D (4) are medium inhibitors of 3alpha-hydroxysteroid-dehydrogenase (3alpha-HSD) inhibiting activity. PMID- 11099228 TI - New 1-O-acyl alpha-L-rhamnopyranosides and rhamnosylated lactones from Streptomyces sp., inhibitors of 3 alpha-hydroxysteroid-dehydrogenase (3alpha HSD). AB - Chemical screening with extracts of Streptomyces sp. (strain GT 61150) resulted in the detection, isolation, and structure elucidation of two new acyl alpha-L rhamnopyranosides (1 and 2) and three new rhamnosyllactones A, B1 and B2 (3 approximately 5). Rhamnosyllactones B1 and B2 were obtained as a 5:1 mixture. The structures were confirmed by spectroscopic analysis, especially 2D-NMR techniques. The rhamnosyltransferase of our strain is able to connect the sugar moiety to heteroaromatic carboxylic acids and enols. The metabolites 1 and 4/5 as well as previously reported acylrhamnosides 6 approximately 11 inhibit the enzyme 3alpha-hydroxysteroid-dehydrogenase (3alpha-HSD). PMID- 11099229 TI - Cyclo(dehydroala-L-Leu), an alpha-glucosidase inhibitor from Penicillium sp. F70614. AB - A diketopiperazine (1) has been isolated from the culture broth of Penicillium sp. F70614 and its structure has been determined to be cyclo(dehydroala-L-Leu) by various spectroscopic analyses. This compound selectively inhibited yeast alpha glucosidase and porcine intestinal alpha-glucosidase with IC50 values of 35 and 50 microg/ml, respectively. However, it did not show significant inhibitory effects against almond beta3-glucosidase, Aspergillus alpha-galactosidase, Escherichia coli beta-galactosidase and jack bean alpha-mannosidase. PMID- 11099230 TI - Structure determination and total synthesis of a novel antibacterial substance, AB0022A, produced by a cellular slime mold. AB - A novel antibacterial substance, AB0022A, was isolated from the cellular slime mold Dictyostelium purpureum K1001. It inhibited the growth of Gram-positive bacteria, and its MICs ranged from 0.39 to 50 microg/ml. Because AB0022A was a highly substituted aromatic compound, we could not determine its structure based on only its physico-chemical and spectral data. We therefore used a dehalogenated derivative from AB0022A and deduced that its structure was 1,9-dihydroxy-3,7 dimethoxy-2-hexanoyl-4,6,8-trichlorodibenzofuran . To confirm this structure, we synthesized the compound having the deduced structure. The synthetic compound was identical to naturally occurring AB0022A. PMID- 11099231 TI - TMC-89A and B, new proteasome inhibitors from streptomyces sp. TC 1087. PMID- 11099232 TI - Piptamine, a new antibiotic produced by Piptoporus betulinus Lu 9-1. PMID- 11099233 TI - Inhibitory effect of laidlomycin on human immunodeficiency virus replication. PMID- 11099234 TI - Wide distribution of interspecific stimulatory events on antibiotic production and sporulation among Streptomyces species. PMID- 11099235 TI - Chemoembolization of liver tumor in a rabbit model: assessment of tumor cell death with diffusion-weighted MR imaging and histologic analysis. AB - PURPOSE: To assess the efficacy of chemoembolization of liver tumors by determining the fraction of viable tumor cells remaining after treatment with use of diffusion magnetic resonance (MR) imaging and histologic analysis. MATERIALS AND METHODS: VX2 tumor was grown in the livers of 12 rabbits. Animals were divided into a chemoembolization group and an untreated group. Conventional, perfusion, and diffusion MR imaging was performed on all rabbits. Histopathologic analysis of explanted livers was performed to document tumor cell death and measure Bcl-2 levels (inhibitor of apoptosis). RESULTS: Diffusion-weighted MR imaging delineated zones of tumor cell death as regions of lower signal intensity in both groups. Apparent diffusion coefficients were significantly greater in the area of tumor necrosis than in the area of viable tumor. Histologic analysis demonstrated a significantly lower percentage of viable cells in the treated group (<1%) than in the control group (55%). Bcl-2 expression detected within the viable areas of the tumor was greater in the treated group than in the control group. CONCLUSIONS: Chemoembolization causes extensive tumor cell destruction. Diffusion MR imaging can detect tumor cell death and can be used to assess the efficacy of chemoembolization. Bcl-2 was expressed in the treated group, suggesting an apoptotic pathway of cell death. PMID- 11099236 TI - Noninvasive vascular laboratory for evaluation of peripheral arterial occlusive disease: Part II--clinical applications: chronic, usually atherosclerotic, lower extremity ischemia. PMID- 11099237 TI - Determination of optimal injection parameters for intraarterial gadolinium enhanced MR angiography. AB - PURPOSE: Rapid vascular depiction with use of a minimum of gadolinium (Gd) contrast agent will be required to generate road-map vascular images for magnetic resonance (MR) imaging-guided endovascular interventions. The objective of this study was to optimize intraarterial injections of MR contrast agent during magnetic resonance angiography (MRA), obtained during interventions, by determining the optimal Gd vascular concentration ([Gd]) for vessel depiction. MATERIALS AND METHODS: The authors derived theoretical expressions to estimate the [Gd] resulting in maximal signal in blood. A model was developed to account for flow dilution to estimate [Gd] given the injected Gd concentration, injection rate, and the blood flow rate. Experiments in four animals (three dogs, one pig) were conducted to verify this model with use of both time-resolved two dimensional (2D) thick-slab and single-phase three-dimensional (3D) MRA acquisitions. The authors also determined the optimal [Gd] required for vessel depiction in animal models. RESULTS: The theoretical expressions yielded optimal [Gd] of 10.2 mmol/L in blood. The animal experiments used the flow dilution model and examined signal enhancement in the aorta and the renal and iliac arteries. Maximal enhancement occurred at [Gd] = 16.2 +/- 4.0 mmol/L (mean +/- SE). CONCLUSIONS: The theoretically predicted values for [Gd]optimal and the flow dilution model were successfully validated. The relationship between injected [Gd], injection rate, and blood flow rate permits rapid intraarterial administration of contrast material, using less overall contrast material than with standard intravenous Gd-enhanced MRA. PMID- 11099238 TI - Long-term outcome of embolotherapy and surgery for high-flow extremity arteriovenous malformations. AB - PURPOSE: To assess the long-term efficacy of embolotherapy in combination with surgery for management of symptomatic high-flow arteriovenous malformations (HFAVMs) of the lower and upper extremities. MATERIALS AND METHODS: Twenty consecutive patients with symptomatic high-flow lower extremity AVMs (LE-AVMs; n = 9) and upper extremity AVMs (UE-AVMs; n = 11) were treated from 1982 to 1999. All nine patients with LE-AVM had pain and seven had ulceration of the skin. All 11 patients with UE-AVM had debilitating pain, seven had weakness of the affected hand, and two had bony erosion. Embolization of the nidus beneath the site of maximum pain or ulceration was performed percutaneously from the femoral artery through coaxially placed microcatheters (n = 18) or surgical cutdown (n = 2). Cyanoacrylate (isobutyl or n-butyl) diluted with iophendylate or ethiodized oil was used in 19 of 20 patients. RESULTS: Follow-up was completed in eight of nine patients with LE-AVM (mean, 8.6 y) and nine of 11 patients with UE-AVM (mean, 7.4 y) after treatment. One patient with localized LE-AVM was functioning well 13 years after embolotherapy and another was functioning well 16 years after undergoing three embolotherapy procedures and two skin grafts. Five of nine patients with LE-AVM required below-the-knee (n = 4) or above-the-knee (n = 1) amputation 1-6 years after technically and clinically successful embolotherapy. All three trifurcation arteries were diffusely involved in HFAVM in patients requiring amputation. Healing of the two amputation sites, involved by AVM at the knee, was excellent after preoperative geniculate artery embolotherapy. All 11 patients with UE-AVM experienced marked symptomatic improvement; seven after embolotherapy alone and the other four after resection of AVM. One complication of digital spasm was reversed by administration of nerve blocks. CONCLUSIONS: LE AVM with diffuse involvement of all three trifurcation arteries ultimately required amputation because of recurrence of symptoms after technically and clinically successful embolotherapy. Cyanoacrylate embolotherapy alone or in combination with surgical resection of the AVM provided excellent long-term palliation in patients with UE-AVM. PMID- 11099239 TI - Endovascular management of acute extensive iliofemoral deep venous thrombosis caused by May-Thurner syndrome. AB - PURPOSE: The authors report their experience on the treatment of acute extensive iliofemoral deep venous thrombosis (DVT) due to May-Thurner syndrome using endovascular techniques. MATERIALS AND METHODS: During a 1-year period, 10 symptomatic women (age range, 22-52 years; mean, 35.5 years) were referred for treatment. After ascending venography, an infusion catheter system was placed and urokinase was infused locally into the thrombus burden. After near complete clot dissolution (> or = 95%) or lytic stagnation, the residual left common iliac vein narrowing was treated by means of angioplasty and/or placement of Wallstent endoprosthesis. All patients continued to receive oral warfarin. Patients were followed-up by means of clinic visits, and stent patency was assessed by means of duplex Doppler sonography performed at 1, 3, 6, and 12 months, and then yearly thereafter. RESULTS: The total dose of urokinase used and the duration of infusion were 5.87 +/- 2.57 million units (range, 3.18-10.7) and 51.95 +/- 21.57 hours (range, 26.5-89), respectively. After completion of thrombolytic therapy, the iliac vein narrowing was successfully treated by deployment of a Wallstent endoprosthesis in all 10 patients because of failure of angioplasty. No major bleeding complications occurred. Initial clinical success was 100%, with complete resolution of symptoms in all patients. One patient, who was hypercoagulable and was receiving chemotherapy for metastatic adenocarcinoma, had recurrent symptomatic acute DVT 1 month after therapy. She underwent successful repeated lysis. The remaining nine patients were asymptomatic, with a mean follow-up of 15.2 months (range, 6-36 months). One asymptomatic patient, at 36-month follow-up ultrasound, had iliac vein occlusion and well-developed venous collaterals. Serial ultrasonography in all 10 patients showed no evidence of valvular insufficiency in the femoral and popliteal veins. CONCLUSION: Catheter-directed thrombolytic therapy for the treatment of acute extensive iliofemoral DVT due to May-Thurner syndrome is an effective method for restoring venous patency and provides relief of the acute symptoms. The underlying left common iliac vein lesion invariably needs to undergo stent placement. PMID- 11099240 TI - Percutaneous treatment of brachiocephalic ischemic complications of a Stanford type A aortic dissection with use of endovascular stents. PMID- 11099241 TI - Venous thrombosis associated with the placement of peripherally inserted central catheters. AB - PURPOSE: Peripherally inserted central catheters (PICCs) have become an essential component of the management of an increasing number of patients, including patients who may require hemodialysis. Reported symptomatic venous thrombosis rates associated with PICC lines are based on clinical signs and symptoms and range from 1% to 4%. The purpose of this study is to evaluate the true rate of thrombosis of upper extremity veins after the placement of PICCs and the potential impact on future access in hemodialysis patients. MATERIALS AND METHODS: A retrospective analysis was performed. Patients who had (i) normal findings during initial venography, (ii) PICC placement, and (iii) who underwent subsequent repeated venography were included. Age, sex, vein cannulated, catheter size, location, and incidence of thrombosis were analyzed. RESULTS: Three hundred fifty-four PICCs were placed in 119 patients. Of the 144 extremities, 137 had normal findings during initial venography. Of the 137 extremities, 32 developed thrombosis of the cannulated vein (or central veins) after initial PICC placement (23.3%). When all extremities with multiple PICC lines placed were considered, 52 developed thrombosis, for an overall thrombosis rate of 38%. The incidence of thrombosis by site was cephalic 57%, basilic 14%, and brachial 10%. No significant differences were noted in the rates of thrombosis by age, sex, or catheter size. CONCLUSIONS: There is a relatively high rate of venous thrombosis associated with PICCs, particularly cephalic thrombus. Because of the high rate of thrombosis associated with these catheters, an alternative mode of access should be considered in current or potential hemodialysis patients. All patients with a history of PICC line placement requiring dialysis access should undergo upper extremity venography prior to the placement of permanent access. PMID- 11099242 TI - Change in peripherally inserted central catheter tip position with abduction and adduction of the upper extremity. AB - PURPOSE: This study examines whether the tip of peripherally inserted central catheters (PICCs) moves significantly with changes in arm position from abduction to adduction. MATERIALS AND METHODS: The catheters were inserted in the brachial or basilic veins under ultrasonographic guidance with the upper extremity in a 90 degrees abducted position. A flexible, radiopaque ruler was then placed on the anterior chest and digital images were obtained with the arm abducted and adducted in a similar phase of quiet respiration. Catheter tip movement was measured with use of the radiopaque ruler and fixed, bony anatomic landmarks. RESULTS: Sixty-one consecutive PICCs were placed and evaluated during the study period (eight patients were excluded). Thirty-three catheters were placed from the right arm and 20 from the left. Overall, 43 moved caudally, seven moved cephalad, and three did not move with movement of the arm from abduction to adduction. Of those that moved caudal, the mean distance of movement was 21 mm (range, 2-53 mm). Right arm PICCs tended to move more than left arm PICCs, but this did not attain significance (P = .29). CONCLUSIONS: There is a tendency for the PICC tip to move in a caudal direction with the change in arm position from abduction to adduction; 58% of PICCs moved 20 mm or more. This change in position should be considered during final catheter tip positioning. PMID- 11099244 TI - Small intestinal submucosa covered expandable Z stents for treatment of tracheal injury: an experimental pilot study in swine. AB - PURPOSE: To evaluate efficacy of small intestinal submucosa (SIS) as a stent covering in healing experimentally created tracheal defects and to explore the trachea's reaction to placement of SIS-covered stents. MATERIAL AND METHODS: A tracheal defect with a diameter of approximately 10 mm was created in six swine with use of a blade or electrocauterization. A double-body, self-expandable SIS covered Gianturco Rosch Z stent was placed into the trachea to cover the defect. The animals were observed, and were killed when they developed respiratory problems. Autopsy and histologic studies were performed. RESULTS: The SIS-covered stents were accurately placed without immediate complications related to placement. All animals developed respiratory problems on follow-up. One animal died 9 days after procedure because of pneumonia, the others five were killed at 12, 17, 18, 28, and 56 days because of stridor, wheezing, and cough. At autopsy and histology, the tracheal defects were found to be completely healed, with epithelial lining and regeneration of submucosal glands. Animals whose defects were created with a blade demonstrated cartilage remodeling between 9 and 18 days, and apparent deposition of new cartilage at 28 days after SIS placement. The defects made by electrocauterization showed only fibrous tissue with no cartilage regeneration. The tracheal lumen was narrowed by overgrowth of granulation tissue, particularly at the end wires of the stents. In three animals, polypoid masses caused 60%, 70%, and 80% tracheal obstruction, respectively. CONCLUSION: Placement of SIS-covered stents contributed to rapid and effective healing of large tracheal defects. Rigidity and oversizing of Gianturco Rosch Z stents led to secondary changes of the tracheal wall, causing significant airway obstructions. Smaller size and flexible stents should be selected for future work. PMID- 11099243 TI - Congenital lacrimal system obstruction: treatment with balloon dilation. AB - PURPOSE: To evaluate the safety and effectiveness of balloon dilation for the treatment of congenital lacrimal system obstruction. MATERIALS AND METHODS: Fluoroscopically guided balloon dilation was attempted in 20 eyes of 16 patients with an age range of 12-78 months (mean, 33 mo) for congenital lacrimal system obstruction. Fifteen eyes had complete obstruction at the valve of Hasner, three eyes had completely obstruction at the junction between the lacrimal sac and the nasolacrimal duct, and two eyes had partial obstruction at the nasolacrimal duct. Under general anesthesia, a ball-tipped guide wire was introduced through the superior punctum into the inferior meatus of the nasal cavity and pulled out through the naris with use of a hook. A deflated 3-mm-diameter balloon catheter was then advanced in a retrograde direction and the balloon was dilated. Every patient underwent an ophthalmic evaluation before the procedure and was scheduled to be followed with office examination at 1, 3, and 6 months after the procedure. RESULTS: There were no major complications. "Technical success" was defined as free passage of contrast medium through the entire lacrimal system to the nasal cavity. The procedure failed in one eye. After balloon dilation, all 19 eyes in which technical success was achieved showed improvement of epiphora. During the follow-up period of 2-33 months (mean, 16 mo), all eyes maintained improvement of epiphora and needed no further treatment. CONCLUSION: Balloon dilation is a safe and effective therapeutic technique for the treatment of congenital lacrimal system obstruction. PMID- 11099245 TI - Management of acute lower extremity embolus with use of the oasis thrombectomy device and suction embolectomy. PMID- 11099246 TI - Use of histoacryl and a covered nitinol stent to treat a bronchobiliary fistula. PMID- 11099247 TI - Successful retrieval of infected Gunther Tulip IVC filter. PMID- 11099248 TI - Elevated plasma levels of matrix metalloproteinase-9 in patients with abdominal aortic aneurysms: a circulating marker of degenerative aneurysm disease. AB - PURPOSE: Matrix metalloproteinase-9 (MMP-9) is abundantly expressed in abdominal aortic aneurysms (AAAs), where it plays a pivotal role in connective tissue destruction. Elevated plasma concentrations of MMP-9 (MMP-9PL) also have been reported in patients with AAAs, but it is unclear if this can distinguish patients with AAAs from those with atherosclerotic occlusive disease (AOD). The purpose of this study was to further define the utility of elevated MMP-9PL levels in the diagnosis and evaluation of AAAs, and to examine if changes in MMP 9PL can be used as a functional biomarker of degenerative aneurysm disease. MATERIALS AND METHODS: Peripheral venous blood was obtained from 25 patients with AAAs, 15 patients with AOD, and five normal control subjects. MMP-9PL levels were determined by an enzyme-linked immunosorbent assay. In four patients undergoing open AAA repair, MMP-9PL levels were directly compared with the amount of MMP-9 produced in aortic tissue. Six additional patients undergoing operative AAA repair were followed for 3-10 months to determine how treatment affected elevated MMP-9PL concentrations. RESULTS: Mean (+/- SE) MMP-9PL was 36.1 +/- 7.7 ng/mL in normal control subjects, 54.7 +/- 10.5 ng/mL in patients with AOD, and 99.4 +/- 17.4 ng/mL in patients with AAAs (P < .05 versus normal control subjects and patients with AOD). Elevated MMP-9PL levels (> 87.8 ng/mL) were found in 12 of 25 (48%) patients with AAA but in only one of 15 (7%) patients with AOD (P < .05). MMP-9PL levels did not correlate significantly with either age, gender, or aneurysm diameter, although there was a trend toward the highest values in male patients with large AAAs. Production of MMP-9 in aneurysm tissues paralleled MMP 9PL levels, and elevated MMP-9PL levels decreased by 92.7% +/- 3.2% after surgical AAA repair. CONCLUSIONS: Elevated MMP-9PL levels were observed in approximately one half of patients with AAAs and less than 10% of those with AOD (positive predictive value of 92.3%), but normal MMP-9PL levels had limited utility in excluding the presence of an aortic aneurysm (negative predictive value, 52%). MMP-9PL levels in patients with AAAs appeared to directly reflect the amount of MMP-9 produced within aneurysm tissue, and MMP-9PL levels decreased substantially after aneurysm repair. Measures of circulating MMP-9 may provide a biologically relevant marker of connective tissue metabolism in patients with AAAs. PMID- 11099249 TI - Plasminogen-enriched pulse-spray thrombolysis with tPA: further developments. AB - PURPOSE: To further improve methods for pulsed plasminogen-enriched thrombolysis and to compare results with the best obtainable with use of tissue plasminogen activator (tPA) alone. MATERIALS AND METHODS: Parameters of plasminogen-enriched pulse-spray thrombolysis were manipulated in groups of rabbits with inferior vena cava thrombosis, and weights of 1-hour residual thrombus were compared. Variables evaluated were (i) tPA pulse frequency, (ii) amount of plasminogen used for enrichment, (iii) tPA concentration and amount, (iv) pulsed versus infused tPA, and (v) admixture versus separation of plasminogen and tPA. RESULTS: With use of 3 mg of tPA and approximately 0.9 mg plasminogen enrichment, efficacy varied directly with pulse frequency over a pulse range of every 15 minutes to every 30 seconds. With use of 30-second pulses of tPA at a concentration 0.125 mg/mL, efficacy also correlated directly with increasing plasminogen enrichment up to, but not beyond, approximately 1.8 mg per 1.24 g of clot. Optimized methodology yielded 89% lysis in 1 hour, as compared to 74% lysis previously reported with use of optimized low-concentration (0.01 mg/ mL) tPA alone. Plasminogen enrichment in conjunction with low concentrations of tPA, admixture of tPA and plasminogen, and fractionation of the plasminogen enrichment all proved to be nonproductive or counterproductive. CONCLUSION: Optimized in vivo postthrombotic plasminogen enrichment significantly accelerated thrombolysis of experimental clots compared to use of optimized tPA alone. PMID- 11099250 TI - Re: Current practice of temporary vena cava filter insertion: a multicenter registry. PMID- 11099251 TI - Re: External beam irradiation as an adjunctive treatment in failing dialysis shunts. PMID- 11099252 TI - Bioinformatics--the necessity of the quest for 'first principles' in life. PMID- 11099253 TI - A systematic approach to dynamic programming in bioinformatics. AB - MOTIVATION: Dynamic programming is probably the most popular programming method in bioinformatics. Sequence comparison, gene recognition, RNA structure prediction and hundreds of other problems are solved by ever new variants of dynamic programming. Currently, the development of a successful dynamic programming algorithm is a matter of experience, talent and luck. The typical matrix recurrence relations that make up a dynamic programming algorithm are intricate to construct, and difficult to implement reliably. No general problem independent guidance is available. RESULTS: This article introduces a systematic method for constructing dynamic programming solutions to problems in biosequence analysis. By a conceptual splitting of the algorithm into a recognition and an evaluation phase, algorithm development is simplified considerably, and correct recurrences can be derived systematically. Without additional effort, the method produces an early, executable prototype expressed in a functional programming language. The method is quite generally applicable, and, while programming effort decreases, no overhead in terms of ultimate program efficiency is incurred. PMID- 11099254 TI - ComboScreen facilitates the multiplex hybridization-based screening of high density clone arrays. AB - MOTIVATION: The construction of physical maps based on bacterial clones [e.g. bacterial artificial chromosomes (BACs)] is valuable for a number of molecular genetics applications, including the high-resolution mapping of genomic regions of interest and the identification of clones suitable for systematic sequencing. A common approach for large-scale screening of bacterial clone libraries involves the hybridization of high-density arrays of immobilized, lysed colonies with collections of DNA probes. The use of a multiplex hybridization screening strategy, whereby pooled probes are analysed en masse, simplifies the effort by reducing the total number of parallel experiments required. However, this approach generates large amounts of hybridization-based data that must be carefully analysed, assimilated, and disambiguated in a careful but efficient manner. RESULTS: To facilitate the screening of high-density clone arrays by a multiplex hybridization approach, we have written a program called ComboScreen. This program provides an organizational framework and analytical tools required for the high-throughput hybridization screening of clone arrays with pools of probes. We have used this program extensively for constructing mouse sequence ready BAC contig maps. PMID- 11099255 TI - Generation of patterns from gene expression data by assigning confidence to differentially expressed genes. AB - MOTIVATION: A protocol is described to attach expression patterns to genes represented in a collection of hybridization array experiments. Discrete values are used to provide an easily interpretable description of differential expression. Binning cutoffs for each sample type are chosen automatically, depending on the desired false-positive rate for the predictions of differential expression. Confidence levels are derived for the statement that changes in observed levels represent true changes in expression. We have a novel method for calculating this confidence, which gives better results than the standard methods. Our method reflects the broader change of focus in the field from studying a few genes with many replicates to studying many (possibly thousands) of genes simultaneously, but with relatively few replicates. Our approach differs from standard methods in that it exploits the fact that there are many genes on the arrays. These are used to estimate for each sample type an appropriate distribution that is employed to control the false-positive rate of the predictions made. Satisfactory results can be obtained using this method with as few as two replicates. RESULTS: The method is illustrated through applications to macroarray and microarray datasets. The first is an erythroid development dataset that we have generated using nylon filter arrays. Clones for genes whose expression is known in these cells were assigned expression patterns which are in accordance with what was expected and which are not picked up by the standards methods. Moreover, genes differentially expressed between normal and leukemic cells were identified. These included genes whose expression was altered upon induction of the leukemic cells to differentiate. The second application is to the microarray data by Alizadeh et al. (2000). Our results are in accordance with their major findings and offer confidence measures for the predictions made. They also provide new insights for further analysis. PMID- 11099256 TI - Six-fold speed-up of Smith-Waterman sequence database searches using parallel processing on common microprocessors. AB - MOTIVATION: Sequence database searching is among the most important and challenging tasks in bioinformatics. The ultimate choice of sequence-search algorithm is that of Smith-Waterman. However, because of the computationally demanding nature of this method, heuristic programs or special-purpose hardware alternatives have been developed. Increased speed has been obtained at the cost of reduced sensitivity or very expensive hardware. RESULTS: A fast implementation of the Smith-Waterman sequence-alignment algorithm using Single-Instruction, Multiple-Data (SIMD) technology is presented. This implementation is based on the MultiMedia eXtensions (MMX) and Streaming SIMD Extensions (SSE) technology that is embedded in Intel's latest microprocessors. Similar technology exists also in other modern microprocessors. Six-fold speed-up relative to the fastest previously known Smith-Waterman implementation on the same hardware was achieved by an optimized 8-way parallel processing approach. A speed of more than 150 million cell updates per second was obtained on a single Intel Pentium III 500 MHz microprocessor. This is probably the fastest implementation of this algorithm on a single general-purpose microprocessor described to date. PMID- 11099257 TI - Genetic network inference: from co-expression clustering to reverse engineering. AB - MOTIVATION: Advances in molecular biological, analytical and computational technologies are enabling us to systematically investigate the complex molecular processes underlying biological systems. In particular, using high-throughput gene expression assays, we are able to measure the output of the gene regulatory network. We aim here to review datamining and modeling approaches for conceptualizing and unraveling the functional relationships implicit in these datasets. Clustering of co-expression profiles allows us to infer shared regulatory inputs and functional pathways. We discuss various aspects of clustering, ranging from distance measures to clustering algorithms and multiple cluster memberships. More advanced analysis aims to infer causal connections between genes directly, i.e. who is regulating whom and how. We discuss several approaches to the problem of reverse engineering of genetic networks, from discrete Boolean networks, to continuous linear and non-linear models. We conclude that the combination of predictive modeling with systematic experimental verification will be required to gain a deeper insight into living organisms, therapeutic targeting and bioengineering. PMID- 11099258 TI - Inferring qualitative relations in genetic networks and metabolic pathways. AB - MOTIVATION: Inferring genetic network architecture from time series data of gene expression patterns is an important topic in bioinformatics. Although inference algorithms based on the Boolean network were proposed, the Boolean network was not sufficient as a model of a genetic network. RESULTS: First, a Boolean network model with noise is proposed, together with an inference algorithm for it. Next, a qualitative network model is proposed, in which regulation rules are represented as qualitative rules and embedded in the network structure. Algorithms are also presented for inferring qualitative relations from time series data. Then, an algorithm for inferring S-systems (synergistic and saturable systems) from time series data is presented, where S-systems are based on a particular kind of nonlinear differential equation and have been applied to the analysis of various biological systems. Theoretical results are shown for Boolean networks with noises and simple qualitative networks. Computational results are shown for Boolean networks with noises and S-systems, where real data are not used because the proposed models are still conceptual and the quantity and quality of currently available data are not enough for the application of the proposed methods. PMID- 11099259 TI - In silico identification of transcripts and SNPs from a region of 4p linked with bipolar affective disorder. PMID- 11099260 TI - TRES: comparative promoter sequence analysis. AB - Comparative promoter analysis is a promising strategy for elucidation of common regulatory modules conserved in evolutionarily related sequences or in genes showing common expression profiles. To facilitate such analysis, we have developed a software tool that detects conserved transcription factor binding sites, cis-elements, palindromes and k-tuples simultaneously in a set of sequences, and thus helps to identify putative motifs for designing further experiments. PMID- 11099261 TI - A comparison of signal sequence prediction methods using a test set of signal peptides. AB - We describe the creation of a test set containing secretory and non-secretory proteins. Five existing prediction programs for signal sequences and their cleavage sites are compared on the basis of this test set: SPScan, SigCleave, SignalP V1.1, SignalP V2.0. b2-HMM and SignalP V2.0.b2-NN. PMID- 11099262 TI - gff2ps: visualizing genomic annotations. AB - gff2psis a program for visualizing annotations of genomic sequences. The program takes the annotated features on a genomic sequence in GFF format as input, and produces a visual output in PostScript. While it can be used in a very simple way, it also allows for a great degree of customization through a number of options and/or customization files. PMID- 11099263 TI - BBID: the biological biochemical image database. AB - The Biological Biochemical Image Database is a WWW accessible relational database of archived images from research articles that describe regulatory pathways of higher eukaryotes. Pathway information is annotated and can be queried in the study of complex gene expression. In this way, complex regulatory pathways can be tested empirically in an efficient manner in the context of large-scale gene expression systems. PMID- 11099264 TI - PASS: prediction of activity spectra for biologically active substances. AB - The concept of the biological activity spectrum was introduced to describe the properties of biologically active substances. The PASS (prediction of activity spectra for substances) software product, which predicts more than 300 pharmacological effects and biochemical mechanisms on the basis of the structural formula of a substance, may be efficiently used to find new targets (mechanisms) for some ligands and, conversely, to reveal new ligands for some biological targets. We have developed a WWW interface for the PASS software. A WWW server for the on-line prediction of the biological activity spectra of substances has been constructed. PMID- 11099265 TI - Personal medical services: a barometer for the NHS? PMID- 11099266 TI - Atypical antipsychotics. PMID- 11099267 TI - The People's Health Assembly. PMID- 11099268 TI - Economic evaluation and clinical trials: size matters. PMID- 11099269 TI - The failings of NICE. PMID- 11099270 TI - Lords call for regulation of complementary medicine. PMID- 11099271 TI - MPs recommend tightening the law on female circumcision. PMID- 11099272 TI - In brief PMID- 11099274 TI - Doctors question new test for Down's syndrome PMID- 11099273 TI - English GP suspended for misleading drug companies. PMID- 11099275 TI - Talks on climate change collapse in acrimony. PMID- 11099277 TI - US calls for research into research integrity PMID- 11099276 TI - Reducing greenhouse gases will have good short term effect. PMID- 11099278 TI - Data collection system needed for gun injuries. PMID- 11099279 TI - Hospital acquired infections kill 5000 patients a year in England. PMID- 11099280 TI - Atypical antipsychotics in the treatment of schizophrenia: systematic overview and meta-regression analysis. AB - OBJECTIVE: To develop an evidence base for recommendations on the use of atypical antipsychotics for patients with schizophrenia. DESIGN: Systematic overview and meta-regression analyses of randomised controlled trials, as a basis for formal development of guidelines. SUBJECTS: 12 649 patients in 52 randomised trials comparing atypical antipsychotics (amisulpride, clozapine, olanzapine, quetiapine, risperidone, and sertindole) with conventional antipsychotics (usually haloperidol or chlorpromazine) or alternative atypical antipsychotics. MAIN OUTCOME MEASURES: Overall symptom scores. Rate of drop out (as a proxy for tolerability) and of side effects, notably extrapyramidal side effects. RESULTS: For both symptom reduction and drop out, there was substantial heterogeneity between the results of trials, including those evaluating the same atypical antipsychotic and comparator drugs. Meta-regression suggested that dose of conventional antipsychotic explained the heterogeneity. When the dose was /= grade 2 acute GVHD (aGVHD) and two developed extensive chronic GVHD (cGVHD). Three patients who received the highest number of donor lymphocyte infusions (DLIs) developed grade 3 GVHD (two patients) and extensive cGVHD (one patient). Ten patients are currently alive, and five are in continuous CR. Seven patients received DLI, with five CRs. Five patients died: one of progressive disease, two of progressive disease combined with aGVHD or cGVHD, one of extensive cGVHD, and one of infection. CONCLUSION: Fludarabine/cyclophosphamide was well tolerated and allowed consistent engraftment in lymphoma allografted patients. Response rates were high in this group of refractory and heavily pretreated patients. This dual procedure seems to be most promising in patients with end-stage malignant lymphomas. PMID- 11099322 TI - Identifying breast cancer patients at high risk for bone metastases. AB - PURPOSE: To identify patient populations at high risk for bone metastases at any time after diagnosis of operable breast cancer, because these patients are potential beneficiaries of treatment with bisphosphonates. PATIENTS AND METHODS: We evaluated data from 6,792 patients who were randomized in International Breast Cancer Study Group clinical trials between 1978 and 1993. Median follow-up was 10. 7 years. A total of 1,275 patients (18.7%) presented with node-negative disease, whereas 3,354 patients (49.4%) had one to three and 2,163 patients (31.9%) had four or more involved axillary lymph nodes. We also assessed the incidence of subsequent bone metastases in the cohort of 1,220 patients who had a first event in local or regional sites or soft tissue alone. Median follow-up for this cohort was 7.7 years from first recurrence. RESULTS: For the entire population with operable disease, the cumulative incidence of bone metastases at any time was 8.2% at 2 years from randomization and 27.3% at 10 years. The highest cumulative incidences of bone metastases at any time were among patients who had four or more involved axillary nodes at the time of diagnosis (14.9% at 2 years and 40.8% at 10 years) and among patients who had as their first event a local or regional recurrence or a recurrence in soft tissue, without any other overt metastases (21.1% at 2 years from first recurrence and 36.7% at 10 years). CONCLUSION: Treatments to prevent bone metastases may have a major impact on the course of breast cancer and may be most efficiently studied in populations with several involved axillary nodes at the time of presentation and in populations with local or regional recurrence or recurrence in soft tissue. PMID- 11099323 TI - p53 gene status and response to platinum/paclitaxel-based chemotherapy in advanced ovarian carcinoma. AB - PURPOSE: The p53 gene plays a critical role in cellular response to DNA damage and has been implicated in the response to platinum compounds in ovarian carcinoma patients. Because taxanes could induce p53-independent apoptosis, we assessed the relevance of p53 gene status to response in ovarian carcinoma patients receiving paclitaxel and platinum-containing chemotherapy. PATIENTS AND METHODS: Forty-eight previously untreated patients with advanced disease received standard paclitaxel/platinum-based chemotherapy. In tumor specimens collected at the time of initial surgery, before therapy, p53 gene status and expression were examined by single-strand conformation polymorphism, sequence analysis, and immunohistochemical analysis. Microsatellite instability analysis was performed on available samples from 30 patients. RESULTS: Thirty-four (71%) of the 48 patients had a clinical response. Pathologic complete remission was documented in 13 (27%) of 48 patients. p53 mutations were detected in 29 (60%) of 48 tumors. Among the patients with mutant p53 tumors, 25 patients (86%) responded to chemotherapy. Only nine (47%) of 19 patients with wild-type p53 tumors responded to the same treatment. The overall response rate and the complete remission rate were significantly higher among patients with mutant p53 tumors than among patients with wild-type p53 tumors (P: =.008). Most of the tested tumors not associated with complete remission (10 of 12 tumors) were also characterized by microsatellite instability. The complete remission rate was higher among patients with tumors without microsatellite instability (five of seven patients). CONCLUSION: In contrast to the limited efficacy of treatment with paclitaxel in combination with standard platinum doses against wild-type p53 ovarian tumors, patients with mutant p53 ovarian tumors were more responsive to paclitaxel-based chemotherapy. The pattern of response to chemotherapy containing paclitaxel is different from that associated with high-dose cisplatin therapy. Determining p53 mutational status can be useful in predicting therapeutic response to drugs effective in ovarian carcinoma. PMID- 11099324 TI - Gene expression for dihydropyrimidine dehydrogenase and thymidine phosphorylase influences outcome in epithelial ovarian cancer. AB - PURPOSE: Dihydropyrimidine dehydrogenase (DPD) is a pyrimidine salvage enzyme responsible for degradation of thymine, which is produced from thymidine by thymidine phosphorylase (TP). Our purpose was to determine whether DPD affects prognosis in patients with epithelial ovarian cancer and how the two enzymes may interact in such effects. PATIENTS AND METHODS: DPD gene expression was analyzed by reverse transcription-polymerase chain reaction in 27 samples from normal ovaries and the 85 epithelial ovarian cancers previously studied with regard to TP gene expression. RESULTS: DPD gene expression was significantly lower in epithelial ovarian cancers than in normal ovaries (P: <.0001), whereas TP gene expression and the ratio of TP to DPD gene expression (TP:DPD) were significantly higher in epithelial ovarian cancer (P: <.0001 for both). In patients with epithelial ovarian cancer, DPD gene expression and the TP:DPD ratio did not significantly correlate with any clinicopathologic factors. Patients with a high TP:DPD ratio (higher than the median) had significantly poorer outcomes than those with lower ratios (P: =.0002). The difference in survival between groups with high and low TP:DPD ratios was greater than the difference between groups with high and low TP gene expression. Multivariate analysis showed the TP:DPD ratio to be the independent prognostic factor (P: =.0002). In tumors with high TP gene expression, low DPD gene expression significantly correlated with poor survival (P: =. 04). CONCLUSION: Downregulation of DPD gene expression may enhance the negative prognostic effect of high TP gene expression in patients with epithelial ovarian cancer. Certain newly available chemotherapeutic choices may take the TP:DPD ratio into consideration. PMID- 11099326 TI - Phase I study in advanced cancer patients of a diversified prime-and-boost vaccination protocol using recombinant vaccinia virus and recombinant nonreplicating avipox virus to elicit anti-carcinoembryonic antigen immune responses. AB - PURPOSE: This trial sought to determine, for the first time, the validity in human vaccinations of using two different recombinant vaccines in diversified prime-and-boost regimens to enhance T-cell responses to a tumor antigen. PATIENTS AND METHODS: Eighteen patients with advanced tumors expressing carcinoembryonic antigen (CEA) were randomized to receive either recombinant vaccinia (rV)-CEA followed by three avipox-CEA vaccinations, or avipox-CEA (three times) followed by one rV-CEA vaccination. Subsequent vaccinations in both cohorts were with avipox-CEA. Immunologic monitoring was performed using a CEA peptide and the enzyme-linked immunospot assay for interferon gamma production. RESULTS: rV-CEA followed by avipox-CEA was superior to the reverse order in the generation of CEA specific T-cell responses. Further increases in CEA-specific T-cell precursors were seen when local granulocyte-macrophage colony-stimulating factor (GM-CSF) and low-dose interleukin (IL)-2 were given with subsequent vaccinations. The treatment was extremely well tolerated. Limited clinical activity was seen using vaccines alone in this patient population. Antibody production against CEA was also observed in some of the treated patients. CONCLUSION: rV-CEA was more effective in its role as a primer of the immune system; avipox-CEA could be given up to eight times with continued increases in CEA T-cell precursors. Future trials should use rV-CEA first followed by avipox-CEA. Vaccines specific to CEA are able to generate CEA-specific T-cell responses in patients without significant toxicity. T-cell responses using vaccines alone may be inadequate to generate significant anticancer objective responses in patients with advanced disease. Cytokines such as GM-CSF and IL-2 may play a key role in generating such responses. PMID- 11099325 TI - Phase I and pharmacokinetic trial of weekly oral fluorouracil given with eniluracil and low-dose leucovorin to patients with solid tumors. AB - PURPOSE: Fluorouracil (5-FU) given as a weekly, high-dose 24-hour infusion is active and tolerable. We evaluated an oral regimen of eniluracil (which inactivates dihydropyrimidine dehydrogenase [DPD]), 5-FU, and leucovorin to simulate this schedule. PATIENTS AND METHODS: Patients received a single 24-hour infusion of 5-FU (2,300 mg/m(2) on day 2) with leucovorin (15 mg orally [PO] bid on days 1 through 3) to provide reference pharmacokinetic data. Two weeks later, patients began treatment with eniluracil (20 mg) and leucovorin (15 mg) (PO bid on days 1 through 3) and 5-FU (10 to 15 mg/m(2) PO bid on day 2). RESULTS: Dose limiting toxicity (diarrhea, neutropenia, and fatigue) was seen with 5-FU 15 mg/m(2) PO bid on day 2 given weekly for either 6 of 8 weeks or 3 of 4 weeks, whereas five of seven patients tolerated 5-FU 10 mg/m(2) PO bid given weekly for 3 of 4 weeks. Eniluracil led to a 35-fold reduction in 5-FU clearance. Fluoro beta-alanine, a 5-FU catabolite, was not detected in plasma during oral 5-FU eniluracil therapy. DPD activity was markedly suppressed in all patients during eniluracil therapy; the inactivation persisted after the last eniluracil dose; percentages of baseline values were 1.8% on day 5, 4.5% on day 12, and 23.6% on day 19. CONCLUSION: The recommended oral dosage of 5-FU (10 mg/m(2) PO bid) given with eniluracil and leucovorin is approximately 115-fold lower than the reference dosage for 24-hour infusional 5-FU. This difference is greater than expected given the reduction in 5-FU clearance. DPD inactivation persisted for several weeks after completion of eniluracil therapy. PMID- 11099327 TI - Phase I and pharmacokinetic study of the differentiating agent vesnarinone in combination with gemcitabine in patients with advanced cancer. AB - PURPOSE: To evaluate the maximum-tolerated dose, dose-limiting toxicities (DLTs), and pharmacokinetic profile of vesnarinone given once daily in combination with gemcitabine. PATIENTS AND METHODS: Twenty-six patients were treated with oral vesnarinone once daily on a continuous schedule at doses of 60, 90, 120, 150, and 180 mg in combination with intravenous (IV) gemcitabine at a dose of 1,000 mg/m(2) on days 1, 8, and 15 every 4 weeks. To determine whether biologically relevant concentrations were being achieved, predose concentrations (C(min)) of vesnarinone were obtained weekly. Plasma gemcitabine and 2',2' difluorodeoxyuridine concentrations were obtained during courses 1 and 2. RESULTS: Twenty-six patients were treated with 92 courses of vesnarinone/gemcitabine. The principal toxicities of the regimen consisted of neutropenia and thrombocytopenia, which were dose-limiting in two of eight heavily pretreated new patients treated at the 90 mg/1,000 mg/m(2) dose level and one of 10 minimally pretreated new patients at the 120 mg/1,000 mg/m(2) dose level. None of three patients treated with 15 courses at the vesnarinone/gemcitabine dose levels of 60 mg/1,000 mg/m(2) experienced DLT. Pharmacokinetic studies of vesnarinone revealed significant interpatient variability at any given dose level. There was evidence of a linear relationship between vesnarinone dose and mean C(min) at dosages of vesnarinone less than 150 mg, with plateauing of mean C(min) values at higher dosages. There was no impact of vesnarinone on gemcitabine concentrations, and the vesnarinone pharmacokinetics did not change with gemcitabine between weeks 1 and 2. Two partial responses occurred in patients with refractory breast and non-small-cell lung carcinoma. CONCLUSION: When combined with gemcitabine, the recommended dose of vesnarinone for phase II evaluations is 90 mg orally once daily with gemcitabine 1,000 mg/m(2) IV on days 1, 8, and 15 every 4 weeks. There is no evidence of pharmacokinetic interaction between vesnarinone and gemcitabine. Further studies of vesnarinone as a single agent or in combination with gemcitabine and other antineoplastic agents are warranted. PMID- 11099329 TI - Myeloblastoma (chloroma) in leukemia: case 1. Granulocytic sarcoma (chloroma) of the breast. PMID- 11099328 TI - Phase I and pharmacokinetic study of the camptothecin analog DX-8951f administered as a 30-minute infusion every 3 weeks in patients with advanced cancer. AB - PURPOSE: DX-8951f is a totally synthetic derivative of camptothecin with greater cytotoxicity and more potent topoisomerase I inhibition than SN-38, topotecan, and camptothecin in preclinical studies. This phase I study aimed to describe the toxicity and to determine the maximum-tolerated dose (MTD) and pharmacokinetics of DX-8951f given as a 30-minute intravenous infusion every 3 weeks. PATIENTS AND METHODS: Twelve patients with refractory solid malignancies were treated with DX 8951f at dose levels ranging from 4 to 7.1 mg/m(2). All but one patient had received previous chemotherapy, and eight patients were considered heavily pretreated. Total DX-8951f plasma concentrations were assayed using high performance liquid chromatography. RESULTS: Thirty-six cycles of DX-8951f were administered. Neutropenia was the dose-limiting toxicity, and it was dose related, reversible, and noncumulative. Other toxicities included nausea and vomiting, alopecia, asthenia, fever, and anemia. Grade 1 or 2 diarrhea was observed in seven patients but was transient and resolved without requiring treatment. Pharmacokinetic analysis showed that DX-8951f had a half-life of 7.15 hours and a clearance rate of 1.65 L/h.m(2). The DX-8951f area under the plasma concentration curve increased linearly with the dose. We defined the MTD of DX 8951f administered as a 30-minute intravenous infusion every 3 weeks as 7.1 mg/m(2). CONCLUSION: The dose-limiting toxicity of DX-8951f is neutropenia. The recommended dose for phase II studies is 5.33 mg/m(2) every 3 weeks in patients previously treated with chemotherapy. PMID- 11099330 TI - Myeloblastoma (chloroma) in leukemia: case 2. Meningeal granulocytic sarcoma (chloroma) in essential thrombocythemia. PMID- 11099331 TI - Death denial. PMID- 11099332 TI - Breast cancer radiotherapy: safe for all? PMID- 11099334 TI - Radiation dose escalation for prostate cancer PMID- 11099333 TI - Standard chemotherapy for gastric carcinoma: is it a myth? PMID- 11099335 TI - Why stop at using high-dose-rate brachytherapy as a boost for the prostate? PMID- 11099336 TI - Radiation endocrine action in prostate cancer PMID- 11099339 TI - Why stop at using high-dose-rate brachytherapy as a boost for the prostate? PMID- 11099337 TI - BR96-doxorubicin: been there, done that! PMID- 11099340 TI - Radiation dose escalation for prostate cancer. PMID- 11099341 TI - Radiation endocrine action in prostate cancer. PMID- 11099342 TI - Announcements PMID- 11099344 TI - Fraction of the dark current carried by Ca(2+) through cGMP-gated ion channels of intact rod and cone photoreceptors. AB - The selectivity for Ca(2+) over Na(+), PCa/PNa, is higher in cGMP-gated (CNG) ion channels of retinal cone photoreceptors than in those of rods. To ascertain the physiological significance of this fact, we determined the fraction of the cyclic nucleotide-gated current specifically carried by Ca(2+) in intact rods and cones. We activated CNG channels by suddenly (<5 ms) increasing free 8Br-cGMP in the cytoplasm of rods or cones loaded with a caged ester of the cyclic nucleotide. Simultaneous with the uncaging flash, we measured the cyclic nucleotide-dependent changes in membrane current and fluorescence of the Ca(2+)-binding dye, Fura-2, also loaded into the cells. The ratio of changes in fura-2 fluorescence and the integral of the membrane current, under a restricted set of experimental conditions, is a direct measure of the fractional Ca(2+) flux. Under normal physiological salt concentrations, the fractional Ca(2+) flux is higher in CNG channels of cones than in those of rods, but it differs little among cones (or rods) of different species. Under normal physiological conditions and for membrane currents 0.6 nm. The relative permeabilities measured under bi-ionic conditions by changes in E(rev) were as follows: C(CN)(3) > SCN > N(CN)(2) > ClO(4) > I > N(3) > Br > Cl > formate > HCO(3) > acetate = F > gluconate. The conductance sequence was as follows: N(3) > Br > Cl > N(CN)(2) > I > SCN > COOH > ClO(4) > acetate > HCO(3) = C(CN)(3) > gluconate. Permeant anions block in a voltage-dependent manner with the following affinities: C(CN)(3) > SCN = ClO(4) > N(CN)(2) > I > N(3) > Br > HCO(3) > Cl > gluconate > formate > acetate. Although these data suggest that anionic selectivity is determined by ionic hydration energy, other factors contribute, because the energy barrier for permeation is exponentially related to anion hydration energy. Cl(Ca)Cs exhibit weak anomalous mole fraction behavior, implying that the channel may be a multi-ion pore, but that ions interact weakly in the pore. The affinity of the channel for Ca(2+) depended on the permeant anion at low [Ca(2+)] (100-500 nM). Apparently, occupancy of the pore by a permeant anion increased the affinity of the channel for Ca(2+). The current was strongly dependent on pH. Increasing pH on the cytoplasmic side decreased the inward current, whereas increasing pH on the external side decreased the outward current. In both cases, the apparent pKa was voltage-dependent with apparent pKa at 0 mV = approximately 9.2. The channel may be blocked by OH(-) ions, or protons may titrate a site in the pore necessary for ion permeation. These data demonstrate that the permeation properties of Cl(Ca)Cs are different from those of CFTR or ClC-1, and provide insights into the nature of the Cl(Ca)C pore. PMID- 11099351 TI - Dynamics of signaling between Ca(2+) sparks and Ca(2+)- activated K(+) channels studied with a novel image-based method for direct intracellular measurement of ryanodine receptor Ca(2+) current. AB - Ca(2+) sparks are highly localized cytosolic Ca(2+) transients caused by a release of Ca(2+) from the sarcoplasmic reticulum via ryanodine receptors (RyRs); they are the elementary events underlying global changes in Ca(2+) in skeletal and cardiac muscle. In smooth muscle and some neurons, Ca(2+) sparks activate large conductance Ca(2+)-activated K(+) channels (BK channels) in the spark microdomain, causing spontaneous transient outward currents (STOCs) that regulate membrane potential and, hence, voltage-gated channels. Using the fluorescent Ca(2+) indicator fluo-3 and a high speed widefield digital imaging system, it was possible to capture the total increase in fluorescence (i.e., the signal mass) during a spark in smooth muscle cells, which is the first time such a direct approach has been used in any system. The signal mass is proportional to the total quantity of Ca(2+) released into the cytosol, and its rate of rise is proportional to the Ca(2+) current flowing through the RyRs during a spark (I(Ca(spark))). Thus, Ca(2+) currents through RyRs can be monitored inside the cell under physiological conditions. Since the magnitude of I(Ca(spark)) in different sparks varies more than fivefold, Ca(2+) sparks appear to be caused by the concerted opening of a number of RyRs. Sparks with the same underlying Ca(2+) current cause STOCs, whose amplitudes vary more than threefold, a finding that is best explained by variability in coupling ratio (i.e., the ratio of RyRs to BK channels in the spark microdomain). The time course of STOC decay is approximated by a single exponential that is independent of the magnitude of signal mass and has a time constant close to the value of the mean open time of the BK channels, suggesting that STOC decay reflects BK channel kinetics, rather than the time course of [Ca(2+)] decline at the membrane. Computer simulations were carried out to determine the spatiotemporal distribution of the Ca(2+) concentration resulting from the measured range of I(Ca(spark)). At the onset of a spark, the Ca(2+) concentration within 200 nm of the release site reaches a plateau or exceeds the [Ca(2+)](EC50) for the BK channels rapidly in comparison to the rate of rise of STOCs. These findings suggest a model in which the BK channels lie close to the release site and are exposed to a saturating [Ca(2+)] with the rise and fall of the STOCs determined by BK channel kinetics. The mechanism of signaling between RyRs and BK channels may provide a model for Ca(2+) action on a variety of molecular targets within cellular microdomains. PMID- 11099352 TI - Do inactivation mechanisms rather than adaptation hold the key to understanding ryanodine receptor channel gating? PMID- 11099353 TI - Ryanodine receptor adaptation. PMID- 11099354 TI - Questions about adaptation in ryanodine receptors. PMID- 11099355 TI - Early repolarization syndrome: clinical characteristics and possible cellular and ionic mechanisms. AB - Early repolarization syndrome (ERS) has traditionally been regarded as benign. In the electrocardiogram (ECG), it is characterized by a diffuse upward ST-segment concavity ending in a positive T wave in leads V2-V4 (5). Clinical interest in this ECG phenomenon has recently been rekindled because of similarities with the electrocardiographic manifestations of the highly arrhythmogenic Brugada syndrome and the potential for misdiagnosis. This article addresses the clinical characteristics and cellular and ionic basis for ERS. In experimental models, the ECG signature of ERS can be converted to that of the Brugada syndrome, raising the possibility that ERS may not be as benign as generally thought, and that under certain conditions known to predispose to ST-segment elevation, patients with ERS may be at greater risk. Further clinical and experimental data are clearly required to test these hypotheses, and the characteristics of ERS need to be more fully delineated within the framework of what has been learned about the Brugada syndrome in recent years. PMID- 11099356 TI - Mechanisms in T-wave alternans caused by intraventricular block. AB - It is recognized that 2:1 intraventricular (IV) block can result in T-wave alternans but is usually assumed that it would also affect QRS waveform. Block in a local region is not, however, varied activation sequence of the same muscle mass because the blocked region is not activated and is not part of the mass that is activated in cycles without block. Also, the block region may have electrocardiogram (ECG) effects when its state differs from other regions. In view of those considerations, the ECG effects of IV block were evaluated by using a computer model of excitation and recovery. ECGs were calculated from differences between the excited state and various degrees of recovery. Results provided evidence that boundaries associated with regions of block rather than regions having varied activation sequence were the major factors in T-wave alternans caused by IV block. Effects of the boundaries included cancellation of the effects of IV block on QRS complexes. Findings suggest that IV block cannot be excluded as a mechanism of T-wave alternans in the absence of QRS alternans. PMID- 11099357 TI - Effects of cardiovascular autonomic function tests on QT dispersion in the 12 lead electrocardiogram of healthy patients. AB - Changes in autonomic tone modulate QT interval duration. How cardiovascular autonomic reflexes affect QT dispersion, a suggested marker of arrhythmia risk, is not well established. We studied 10 healthy young adult volunteer men during quiet and deep breathing, the Valsalva maneuver, sustained handgrip, hyperventilation, the cold pressor test, and mental stress. An automated method was used for measurement of QT-peak and QT-end intervals, and QT dispersion was defined as maximum-minimum of the measured intervals. QT-peak dispersion was greater on deep expiration than deep inspiration (49 +/- 20 ms vs 37 +/- 14 ms, P < .05). QT-end dispersion decreased in the tachycardia phase of the Valsalva maneuver (45 +/- 23 ms vs 35 +/- 21 ms, P < .05), but QT dispersion did not change during the other interventions. Rapid cardiovascular autonomic reflex adjustment does not change QT dispersion in healthy young adult men. However, large intrathoracic volume and intrathoracic pressure changes during forced respiratory movements might confound QT dispersion measurements. PMID- 11099358 TI - The added diagnostic value of automated QT-dispersion measurements and automated ST-segment deviations in the electrocardiographic diagnosis of acute cardiac ischemia. AB - The purpose of this study was to determine the added value of automated QT dispersion and ST-segment measurements to physician interpretation of 12-lead electrocardiograms (ECGs) in patients with chest pain. To date, poor reproducibility of manual measurements and lack of shown added value have limited the clinical use of QT dispersion. Twelve-lead ECGs (n = 1,161) from the Milwaukee Prehospital Chest Pain Database were independently classified by 2 physicians into 3 groups (acute myocardial infarction (AMI), acute cardiac ischemia (ACI), or nonischemic), and their consensus was obtained. QT-end and QT peak dispersions were measured by a computerized system. The computer also identified ST-segment deviations. Sensitivity, specificity, and positive predictive values (PPVs) and negative predictive values (NPV) for AMI and ACI were evaluated independently and in combinations. For AMI, physicians' consensus classification was remarkably good (sensitivity, 48%, specificity, 99%). Independent classification by QT-end and QT-peak dispersions or ST deviations was not superior to the physicians' consensus. Optimal classification occurred by combining automated QT-end dispersion and ST deviations with physicians' consensus. This combination increased sensitivity for the diagnoses of AMI by 35% (65% vs 48%, P < .001) and ACI by 55% (62% vs 40%, P < .001) compared with physicians' consensus, while maintaining comparable specificity. This study supports a potential clinical role for automated QT dispersion when combined with other diagnostic methods for detecting AMI and ACI. PMID- 11099359 TI - The ability of a computer program based on the Marquette Matrix-12 short measurement matrix to replicate coding by the Minnesota ECG coding laboratory. AB - The study was undertaken to determine whether a computer program that uses "short measurement matrix" data from the Marquette Matrix-12 system can replicate Minnesota electrocardiogram (ECG) coding laboratory interpretations. An agreement was found between coding of median complex ECGs at the Minnesota ECG coding laboratory and coding based on Marquette Matrix-12 short measurement matrix. The comparison was based on 763 ECGs plus chest pain history and serum enzyme values for a stratified random sample of 141 patients hospitalized in 1990 or 1991 for an event coded as HICDA 410.x (acute myocardial infarction), 411 (other acute and subacute forms of ischemic heart disease), 413 (angina pectoris), or 796.9 (other ill defined and unknown causes of morbidity and mortality). The population was reconstructed from the stratified random sample to enable population-based inferences. Exact agreement between Matrix-12 and Minnesota coding laboratory interpretation on 4 ECG patterns (evolving diagnostic, diagnostic, equivocal, or other ECG pattern) was 74.5% (Kappa = 0.63 +/- 0.05) for the stratified random sample and 78.8% (Kappa = 0.66 +/- 0.05) for the reconstructed population. For coding myocardial infarction based on the ECG, serum enzyme levels, and ischemic chest pain, agreement was 91.5% (Kappa = 0.85 +/- 0.04) for the stratified random sample and 90% (Kappa = 0.83 +/- 0.04) for the reconstructed population. Although ECG interpretation by a computer program based on the short measurement matrix of the Matrix 12 system results in better agreement than prior attempts to replicate the Minnesota coding laboratory, interpretation remains unacceptably discordant. PMID- 11099360 TI - Onset dynamics of reentrant tachycardia and rate-dependent conduction changes in canine ventricular muscle: effects of Na+ and Ca2+ channel blockade. AB - To show that cycle-length (CL) prolongation occurring at the onset of reentrant tachycardias may be associated with an increase in conduction time (CT), and to investigate the involvement of Na+ and Ca2+ channel activity, reentrant activity was induced by programmed stimulation in thin ventricular muscle slices with a central cryothermal lesion, as documented with 7 to 12 bipolar recordings. We studied the course of the CL measured in successive tachycardia beats, as well as the course of conduction times after abrupt transition from a pacing CL of 1,000 to 400 ms (pacing paradigm). The majority of the tachycardias displayed a dynamic behavior in which CL increased progressively, with an exponential rate constant of 37 +/- 35 beats (mean +/- SD), stabilizing at 325 +/- 67 ms after a total increase of 17 +/- 17 ms. In the pacing paradigm, CT was prolonged from 68 +/- 21 ms to 79 +/- 24 ms according to a biphasic course consisting of an abrupt increase in the first response to 400 ms, followed up by an exponential increase, stabilizing with a rate constant of 18 +/- 23 beats. Lidocaine 5 x 10(-5) mol/L induced an increase in steady-state CT, which was not further modified by adding verapamil 10(-5) mol/L. However, verapamil prolonged the rate constant of the exponential course by 60 +/- 40 beats. Thus, the onset dynamics of reentrant tachycardias share common features with the dynamic behavior of CT in the pacing paradigm, in which both Na+ channel activity and Ca2+-modulated cellular coupling appear to be involved. PMID- 11099361 TI - Effects of intracellular calcium elevation on action potential and L-type calcium current of normal and chronically infarcted rat ventricles. AB - The present work investigated the effects of raising [Ca+2]i levels on action potential (AP) and L-type calcium current (I(Ca.L)) of normal and chronically infarcted rat ventricles. Experiments were performed by conventional electrophysiology and whole-cell patch-clamp techniques. In the former, APs were recorded in ventricular strips subjected to different pacing rates or elevation of [Ca+2]o levels. In the latter, I(Ca.L) was studied in isolated myocytes in the absence of an intracellular Ca+2 chelator. The acceleration of heart rate (6 to 240 beats/min) reduced AP duration measured at 20%, 50%, and 90% repolarization (APD20, APD50, and APD90) in the infarcted group, and increased APD20 and APD50 in the control group. Rising [Ca+]o (1.25 to 5.0 mmol/L) induced a decrease of APD20 and APD50 in both groups. Voltage clamp revealed a smaller I(Ca.L) density at approximately -17 mV in myocytes from infarcted ventricles (-1.86 +/- 0.37 vs 3.98 +/- 0.65 pA/pF, P < .05), and the appearance of a non-K+ outward current coupled to I(Ca.L). The results suggest the participation of a Ca+2-activated outward current in the repolarization of normal and infarcted rat ventricles. PMID- 11099362 TI - Apparent bradycardia-dependent sinoatrial block associated with respiration. AB - In our previous patients, apparent bradycardia-dependent block has been shown in the atrioventricular (AV) junction and in the accessory pathway. It was suggested that these previous cases were not of true bradycardia-dependent block; namely, that, as a result of periodic increases in vagal tone associated with respiration, conductivity in the AV junction or in the accessory pathway was depressed to a greater degree than automaticity in the sinus node. In the present article, 3 patients with frequent sinoatrial (SA) block were reported. In 1 patient, sinus escape-capture bigeminy caused by SA block was found. In these present patients, when the sinus cycle lengthened, SA block occurred. The purpose of the present article is to show that the patients have apparent bradycardia dependent SA block, namely, not true bradycardia-dependent SA block. In all patients, the respiration curve was recorded simultaneously with the electrocardiogram. In all patients, during inspiration, the sinus cycle gradually shortened; on the other hand, during expiration, the sinus cycle gradually lengthened, and then a sinus impulse was blocked in the SA junction. These findings suggested that increased vagal tone during expiration depressed conductivity in the SA junction to a greater degree than automaticity in the sinus node. PMID- 11099363 TI - Electrophysiologic findings of a patient with inappropriate sinus tachycardia cured by selective radiofrequency catheter ablation. AB - Radiofrequency catheter ablation (RFCA) for inappropriate sinus tachycardia (IST) is associated with a high recurrence rate and sometimes requires pacemaker implantation, especially after extensive ablation. We report a patient with drug refractory IST who was successfully treated by selective RFCA to the 2 earliest activation sites. During tachycardia, the earliest atrial activation preceded the surface P wave by 50 ms or more, whereas it was only 27 ms for the rest of the right atrium after ablation. Our patient had the longest activation period during tachycardia among the reported patients. In IST patients, a longer activation time at the site of the earliest atrial activation may imply that the abnormality is confined to a small area within the sinus node and may predict the efficacy of selective RFCA. PMID- 11099364 TI - Right bundle branch block-induced Q waves simulating anterior myocardial infarction extension. AB - A concept generally accepted in clinical electrocardiography is the assumption that a right bundle branch block (RBBB) does not alter significantly the initial portion of the QRS complex and because the left bundle branch is intact, the septum is activated normally in a left-to-right direction. We report a woman with an acute anterior myocardial infarction (MI) in which a small R wave was present in leads V1 and V4, but with the development of RBBB associated with PR-interval prolongation, these R waves were replaced by Q waves. Subsequently, the electrocardiographic features of anterior MI disappeared concomitantly with the loss of RBBB. The clinical and electrophysiologic implications of these findings are discussed. PMID- 11099365 TI - Intravenous versus oral rehydration during a brief period: stress hormone responses to subsequent exhaustive exercise in the heat. AB - The purpose of this study was to determine if intravenous fluid rehydration, versus oral rehydration, during a brief period (20 min) differentially affects plasma ACTH, cortisol, and norepinephrine concentrations during subsequent exhaustive exercise in the heat. Following dehydration (DHY) to Eth 4% of body weight, 8 nonacclimated highly trained males (age = 23.5 +/- 1.2 years, VaO2peak = 61.4 +/- 0.8 ml a kg a min-1, % body fat = 13.5 +/- 0. 6%) cycled to exhaustion at 74% VaO2peak in 36.8 C on three different occasions. These included: (a) no fluid (NF), where no fluid was provided during the rehydration period; (b) DRINK, where oral rehydration (0.45% NaCl) was provided equal to 50% of the prior DHY; and (c) IV, where intravenous infusion (0.45% NaCl) was provided equal to 50% of the prior DHY. Exercise time to exhaustion was not different (p =.07) between the DRINK (34.86 +/- 4.01) and IV (29.48 +/- 3.50) trials, but both were significantly (p <.05) longer than the NF (18.95 +/- 2.73) trial. No differences (p >.05) were found for any of the hormone measures among trials. The endocrine responses at exhaustion were similar regardless of hydration state and mode of rehydration, but rehydration prolonged the exercise time to exhaustion. PMID- 11099366 TI - Effects of ingesting a large volume of carbohydrate-electrolyte solution on rehydration during recovery and subsequent exercise capacity. AB - This randomized, double-blind study examined the effects of rehydration per se and rehydration plus carbohydrate (CHO) ingestion during recovery (REC) on subsequent endurance running capacity. Nine men ran at 70% VaO2max on a level treadmill for 90 min (T1) on two occasions, followed by a 4 hour REC and a further exhaustive run at the same speed (T2). During the first 3 hours of REC, subjects drank either a 6.9% CHO-electrolyte solution (CE) or a CHO- and electrolyte-free sweetened placebo (PL) every 30 min. Volumes prescribed were 200% of the fluid lost after T1, but the actual volume of fluid ingested during the REC ranged from 113-200% and 88. 5-200% of the body mass lost for the CE and PL trials (NS). However, positive fluid balance was found in both trials after REC. During T2, run time was 24.3 +/- 4.4 min longer in the CE trial (69.3 +/- 5.5 vs. 45.0 +/- 4.2 min; p <.05). Higher blood glucose concentrations were observed throughout REC in the CE trial. These results suggest that ingesting a CHO-electrolyte solution is more effective in restoring endurance capacity compared to the same large volume of placebo, even though complete rehydration was achieved in both trials. PMID- 11099367 TI - Active recovery counteracts the post-exercise rise in plasma-free fatty acids. AB - The present study investigated the effect of active recovery (AR) as compared to rest recovery (RR) upon FFA concentrations following moderate- (MI) or high intensity (HI) running. Fourteen well-trained males (23.7 +/- 6 years, VaO2max = 69.5 +/- 1.8ml a min-1 kg-1) were randomly assigned into two trials (HI = 30 min at 82% of VaO2max; MI = 60 min at 75% of VaO2max). Within each group, the subject completed two sets of experiments of running followed by either AR (15 min running at 50% of VaO2max) or RR (complete rest in the supine position). Plasma volume changes after the exercise did not deviate between the AR or RR trials. In both the HI and MI trials, AR resulted in lower FFA peaks and lower overall FFA concentrations while performing AR (p <.05). However, upon discontinuing AR, there was a rise in the FFA concentration. At 120-min post-exercise, the FFA concentrations after AR and RR were not significantly different. The changes in the FFA/albumin ratio were similar to the FFA responses. It is concluded that AR may counteract the rise in FFA 5-15 minutes after exercise. PMID- 11099368 TI - Effects of meal form and composition on plasma testosterone, cortisol, and insulin following resistance exercise. AB - The purpose of this study was to examine the effects of postexercise feeding on plasma levels of insulin, testosterone, cortisol, and testosterone:cortisol (T:C). Ten experienced, resistance trained males (20.7 +/- 0.95 years) were given whole food (WF: protein 38 g; carbohydrate 70 g; fat 7 g), a supplemental drink (SD; isocaloric and isonitrogenous to WF), an isocaloric carbohydrate beverage (C), or a placebo beverage (P) immediately, 2 and 4 hours after a standardized weight training protocol on 4 days, each separated by 1 week, in a repeated measures design. Subjects also received a standardized meal at 7 and 12 hours postexercise. Insulin, testosterone, and cortisol were measured pre-exercise and during 24 hours of recovery (at 0.5, 2.5, 4.5, 8, and 24 hours) using venous blood samples. Significant (condition 3 time) interactions were found for insulin, testosterone, and T:C, but not for cortisol (p <. 05). The SD yielded a greater response for insulin than all other conditions. Conversely, P demonstrated the greatest values for testosterone and T:C at 2.5 and 4.5 hours postexercise. Cortisol did not vary between conditions and there were no condition effects for insulin, testosterone, cortisol, and T:C at 8 or 24 hours. In conclusion, the efficacy of postexercise feeding for optimizing T:C and muscle growth is unclear; however, consumption of SD appears to maximize circulating insulin for several hours following resistance exercise. PMID- 11099369 TI - Iron status and resting immune function in female collegiate swimmers. AB - Iron deficiency may lead to anemia and may result in compromised endurance exercise performance. Iron deficiency has also been reported to adversely affect the immune system and has been associated with attenuation of natural killer cell (NK) activity. This study was conducted to examine the relationship between iron status and NK activity in highly conditioned female athletes. Ten collegiate female swimmers (SWM) and 9 inactive females (SED) participated in this investigation. Resting blood samples were obtained and analyzed for serum iron and ferritin. NK activity (% lysis) was determined using a whole blood method (51Cr release assay). No significant relationship was found between iron and NK activity (r = 0.55, p =.09), nor between serum ferritin and NK activity (r = 0.33, p =.35) for SWM. ANOVA revealed significantly greater NK activity for SWM (51.63 +/- 15.79%) versus SED (30.34 +/- 13.67%). Serum ferritin levels were not significantly different between SWM (20.38 +/- 8.62 hg a ml-1) and SED (16.79 +/- 10.53 hg a ml-1), nor were iron values different between groups (16.54 +/- 2. 17 mmol a L-1 SWM; 11.92 +/- 2.61 mmol a L-1 SED). A significant relationship between iron status and resting immune function could not be established. Exercise training may affect NK activity; however, the influence of iron status on immune function requires further evaluation. PMID- 11099370 TI - Effect of iron supplementation on thyroid hormone levels and resting metabolic rate in two college female athletes: a case study. AB - Iron plays an important role in thyroid hormone metabolism; thus, iron deficiency anemia may lead to alterations in resting metabolic rate (RMR). Based on this premise, two iron-deficient-anemic female athletes, 18 (A1) and 21 (A2) years of age, were supplemented with 23 mg/day of elemental iron to assess its effects on iron and thyroid hormone status and RMR at 0, 8, and 16 weeks. Anemia was clinically corrected in both subjects (hemoglobin: A1 = 11.0 to 13.0 to 12.6 g/dL and A2 = 11.5 to 13.9 to 12.6 g/dL, 0 to 8 to 16 weeks, respectively). Serum ferritin (SF) concentration also improved in both subjects (A1: 5.0 to 11.0 to 15.0 ng/dL and A2: 5.0 to 16.0 to 20.0 ng/dL; 0 to 8 to 16 weeks, respectively); however, 16 weeks of iron supplementation did not fully replete iron stores. A2 increased dietary iron and ascorbic acid intakes from 8 to 16 weeks, possibly accounting for her higher SF concentrations. RMR and total thyroxine changed over time: A1 increased while A2 decreased in these variables. Although clinical correction of iron deficiency anemia occurred after 16 weeks of low-level iron supplementation, RMR and thyroid hormone metabolism were oppositely affected in the two subjects. PMID- 11099371 TI - The effect of siberian ginseng (Eleutherococcus senticosus) on substrate utilization and performance. AB - It has been suggested that Eleutherococcus senticosus (ES), also known as Siberian ginseng or ciwuija, increases fat utilization in humans. The purpose of this study was to examine the physiological responses to supplementation with ES in endurance cyclists. Using a randomized, double-blind crossover design, 9 highly-trained men (28 +/- 2 years, VaO2max 57.3 +/- 2.0 ml a kg-1 a min-1) cycled for 120 min at '60% VaO2max followed by a simulated 10-km time trial. Diet was controlled, and ES (1,200 mg a day-1) or a placebo (P) were administered for 7 days prior to each of the two trials. Oxygen consumption, respiratory exchange ratio, and heart rate were recorded every 30 min, and rating of perceived exertion, plasma [lactate], and plasma [glucose] were recorded every 20 min during the 120 min of steady state cycling. There were no significant differences (p >.05) between the ES and P groups at any steady-state time interval or during the cycling time trial (ES = 18.10 +/- 0.42, P = 17.83 +/- 0.47 min). In contrast with previous reports, the results of this study suggest that ES supplementation does not alter steady-state substrate utilization or 10-km cycling performance time. PMID- 11099372 TI - Creatine monohydrate supplementation enhances high-intensity exercise performance in males and females. AB - Creatine monohydrate supplementation has been shown to enhance high-intensity exercise performance in some but not all studies. Part of the controversy surrounding the ergogenic effect(s) of creatine monohydrate supplementation may relate to design issues that result in low statistical power. A further question that remains unresolved in the creatine literature is whether or not males and females respond in a similar manner to supplementation. We studied the effect of creatine supplementation upon high intensity exercise performance in 24 subjects (n = 12 males, n = 12 females). Creatine monohydrate (Cr; 5g, 4x/d 3 4d) and placebo (Pl; glucose polymer 3 4d) were provided using a randomized, double-blind crossover design (7 week washout). Outcome measures included: 2 3 30-s anaerobic cycle test, with plasma lactate pre- and post-test; dorsi-flexor: maximal voluntary contraction (MVC), 2-min fatigue test, and electrically stimulated peak and tetanic torque; isokinetic knee extension torque and 1-min ischemic handgrip strength. Significant main effects of Cr treatment included: increased peak and relative peak anaerobic cycling power ( 3.7%; p <. 05), dorsi-flexion MVC torque ( 6.6%; p <.05), and increased lactate ( 20.8%; p <.05) with no gender specific responses. We concluded that short-term Cr supplementation can increase indices of high-intensity exercise performance for both males and females. PMID- 11099373 TI - Improved 2000-meter rowing performance in competitive oarswomen after caffeine ingestion. AB - Eight competitive oarswomen (age, 22 +/- 3 years; mass, 64.4 +/- 3.8 kg) performed three simulated 2,000-m time trials on a rowing ergometer. The trials, which were preceded by a 24-hour dietary and training control and 72 hours of caffeine abstinence, were conducted 1 hour after ingesting caffeine (6 or 9 mg a kg-1 body mass) or placebo. Plasma free fatty acid concentrations before exercise were higher with caffeine than placebo (0.67 +/- 0.34 vs. 0.72 +/- 0.36 vs. 0.30 +/- 0.10 mM for 6 and 9 mg a kg-1 caffeine and placebo, respectively; p <.05). Performance time improved 0.7% (95% confidence interval [CI] 0 to 1.5%) with 6 mg a kg-1 caffeine and 1. 3% (95% CI 0.5 to 2.0%) with 9 mg a kg-1 caffeine. The first 500 m of the 2,000 m was faster with the higher caffeine dose compared with placebo or the lower dose (1.53 +/- 0.52 vs.1.55 +/- 0.62 and 1. 56 +/- 0.43 min; p =.02). We concluded that caffeine produces a worthwhile enhancement of performance in a controlled laboratory setting, primarily by improving the first 500 m of a 2,000-m row. PMID- 11099374 TI - Effects of carbohydrate and chromium ingestion during intermittent high-intensity exercise to fatigue. AB - PURPOSE: This study was designed to test the hypothesis that addition of chromium (Cr) to a carbohydrate-electrolyte drink would enhance the reported benefits of carbohydrate on exercise capacity during intermittent high-intensity shuttle running. METHODS: Eight physically active men performed 3 exercise trials while ingesting 6% carbohydrate-electrolyte (CHO), CHO plus chromium picolinate (400 mg) (CHO + Cr+3), or placebo (P) using a double-blind, counterbalanced design. Each trial consisted of 5 3 15 min bouts of shuttle running (walk, sprint, and run at 95 and 55% of estimated VaO2max, separated by 3-min rest). This was followed by a fatigue test (running alternating 20-m lengths at 55 and 95% of estimated VaO2 until fatigue). RESULTS: During the standardized shuttle running, blood glucose was higher with both CHO and CHO + Cr+3 than P. Plasma free fatty acid was higher in P than both CHO and CHO + Cr+3 at 75 min of exercise and at fatigue. In the fatigue test, subjects ran longer with both CHO and CHO + Cr+3 than P. CONCLUSIONS: The data confirm an ergogenic benefit of ingesting CHO during exercise designed to imitate sports like basketball, soccer, and hockey, but do not support the hypothesis that the addition of Cr would enhance this effect. PMID- 11099375 TI - Ligand-induced strain in hydrogen bonds of the c-Src SH3 domain detected by NMR. AB - Changes in the molecular conformation of proteins can result from a variety of perturbations, and can play crucial roles in the regulation of biological activity. A new solution NMR method has been applied to monitor ligand-induced changes in hydrogen bond geometry in the chicken c-Src SH3 domain. The structural response of this domain to ligand binding has been investigated by measuring trans-hydrogen bond (15)N-(13)C' scalar couplings in the free state and when bound to the high affinity class I ligand RLP2, containing residues RALPPLPRY. A comparison between hydrogen bonds in high resolution X-ray structures of this domain and those observed via (h3)J(NC') couplings in solution shows remarkable agreement. Two backbone-to-side-chain hydrogen bonds are observed in solution, and each appears to play a role in stabilization of loop structure. Reproducible ligand-induced changes in trans-hydrogen bond scalar couplings are observed across the domain that translate into changes in hydrogen bond length ranging between 0.02 to 0.12 A. The observed changes can be rationalized by an induced fit mechanism in which hydrogen bonds across the protein participate in a compensatory response to forces imparted at the protein-ligand interface. Upon ligand binding, mutual intercalation of the two Leu-Pro segments of the ligand between three aromatic side-chains protruding from the SH3 surface wedges apart secondary structural elements within the SH3 domain. This disruption is transmitted in a domino-like effect across the domain through networks of hydrogen bonded peptide planes. The unprecedented resolution obtained demonstrates the ability to characterize subtle structural rearrangements within a protein upon perturbation, and represents a new step in the endeavor to understand how hydrogen bonds contribute to the stabilization and function of biological macromolecules. PMID- 11099376 TI - Movement of the decoding region of the 16 S ribosomal RNA accompanies tRNA translocation. AB - The ribosome undergoes pronounced periodic conformational changes during protein synthesis. Of particular importance are those occurring around the decoding site, the region of the 16 S rRNA interacting with the mRNA-(tRNA)(2) complex. We have incorporated structural information from X-ray crystallography and nuclear magnetic resonance into cryo-electron microscopic maps of ribosomal complexes designed to capture structural changes at the translocation step of the polypeptide elongation cycle. The A-site region of the decoding site actively participates in the translocation of the tRNA from the A to the P-site upon GTP hydrolysis by elongation factor G, shifting approximately 8 A toward the P-site. This implies that elongation factor G actively pushes both the decoding site and the mRNA/tRNA complex during translocation. PMID- 11099377 TI - Molecular cloning and expression of human UDP-d-Xylose:proteoglycan core protein beta-d-xylosyltransferase and its first isoform XT-II. AB - Human UDP-d-xylose:proteoglycan core protein beta-d-xylosyltransferase (EC 2.4.2.26, XT-I) initiates the biosynthesis of glycosaminoglycan chains in proteoglycans by transferring xylose from UDP-xylose to specific serine residues of the core protein. Based on the partial amino acid sequence of the purified enzyme from human JAR choriocarcinoma cell culture supernatant we isolated a cDNA encoding XT-I using the degenerate reverse transcriptase-polymerase chain reaction method. This enzyme, which is involved in chondroitin sulfate, heparan sulfate, heparin and dermatan sulfate biosynthesis, belongs to a novel family of glycosyltransferases with no homology to proteins known so far. 5' and 3'-RACE were performed to isolate a novel cDNA fragment of 3726 bp with a single open reading frame encoding at least 827 amino acid residues with a molecular mass of 91 kDa. The human XT-I gene was located on chromosome 16p13.1 using radiation hybrid mapping, and extracts from CHO-K1 cells transfected with the XT-I cDNA in an expression vector exhibited marked XT activity. A new 3608 bp cDNA fragment encoding a protein of 865 amino acid residues was also isolated by PCR using degenerate primers based on the amino acid sequence of human XT-I. The amino acid sequence of this XT-II isoform displayed 55% identity to the human XT-I. The XT II gene was located on chromosome 17q21.3-17q22, and the exon/intron structure of the 15 kb gene was determined. RT-PCR analyses of XT-I and XT-II mRNA from various tissues confirmed that both XT-I and XT-II transcripts are ubiquitously expressed in the human tissues, although with different levels of transcription. Furthermore, the cDNAs encoding XT-I and XT-II from rat were cloned. The deduced amino acid sequences of rat xylosyltransferases displayed 94% identity to the corresponding human enzyme. PMID- 11099378 TI - Phage display screening reveals an association between germline-specific transcription factor Oct-4 and multiple cellular proteins. AB - Oct-4 is a transcription factor that is specifically expressed in mouse embryonic stem cells and in cell lines derived thereof. In these cells, Oct-4 activates transcription from remote binding sites due to as of yet unknown co-activators. Expression of Oct-4 in differentiated cells is not sufficient to activate transcription from a distance, rather it requires the co-expression of co activators such as the adenoviral oncoprotein E1A. In this paper, we used phage display to identify Oct-4-interacting proteins. We first analyzed the interaction between Oct-4 and E1A in order to optimize the biochemical conditions that enable Oct-4-specific interactions with other interacting proteins. A panning approach was used to enrich Oct-4 interacting phages from a pool of excess unspecific phages. The biochemical conditions established in our interaction assays were then used to screen a P19 EC cell cDNA expression library in M13 filamentous phage. A number of phage clones displaying portions of unknown and known transcription factors were obtained, from which the HMG-1 transcription factor was identified. HMG-1, and the closely related factor HMG-2, interact with Oct-4 when co-expressed in mammalian cells. In addition, HMG-1 was found to cooperate with Oct-4 in P19 EC cells. These results provide the first evidence of a non viral factor that enhances Oct-4 distance-dependent transactivation in stem cells. PMID- 11099379 TI - BII nucleotides in the B and C forms of natural-sequence polymeric DNA: A new model for the C form of DNA. AB - A combination of solid-state (31)P and (13)C NMR, X-ray diffraction, and model building is used to show that the B and C forms of fibrous macromolecular DNA consist of two distinct nucleotide conformations, which correspond closely to the BI and BII nucleotide conformations known from oligonucleotide crystals. The proportion of the BII conformation is higher in the C form than in the B form. We show structural models for a 10(1) double helix involving BI nucleotides and a 9(1) double helix involving BII nucleotides. The 10(1) BI model is similar to a previous model of B-form DNA, while the 9(1) BII model is novel. The BII model has a very deep and narrow minor groove, a shallow and wide major groove, and highly inclined bases. This work shows that the B to C transition in fibers corresponds to BI to BII conformational changes of the individual nucleotides. PMID- 11099380 TI - Structure of membrane-bound annexin A5 trimers: a hybrid cryo-EM - X-ray crystallography study. AB - Annexins constitute a family of phospholipid- and Ca(2+)-binding proteins involved in a variety of membrane-related processes. The property of several annexins, including annexin A5, to self-organize at the surface of lipid membranes into 2D ordered arrays has been proposed to be functionally relevant in cellular contexts. To further address this question, we investigated the high resolution structure of annexin A5 trimers in membrane-bound 2D crystals by cryo electron microscopy (Cryo-EM). A new 2D crystal form was discovered, with p32(1) symmetry, which is significantly better ordered than the 2D crystals reported before. A 2D projection map was obtained at 6.5 A resolution, revealing protein densities within each of the four domains characteristic of annexins. A quantitative comparison was performed between this structure and models generated from the structure of the soluble form of annexin A5 in pseudo-R3 3D crystals. This analysis indicated that both structures are essentially identical, except for small local changes attributed to membrane binding. As a consequence, and contrary to the common view, annexin A5 molecules maintain their bent shape and do not flatten upon membrane binding, which implies either that the four putative Ca(2+) and membrane-binding loops present different types of interaction with the membrane surface, or that the membrane surface is locally perturbed. We propose that the trimerization of annexin A5 molecules is the relevant structural change occurring upon membrane binding. The evidence that 2D arrays of annexin A5 trimers are responsible for its in vitro property of blood coagulation inhibition supports this conclusion. PMID- 11099381 TI - Crystal structure of a truncated mutant of glucose-fructose oxidoreductase shows that an N-terminal arm controls tetramer formation. AB - N-terminal or C-terminal arms that extend from folded protein domains can play a critical role in quaternary structure and other intermolecular associations and/or in controlling biological activity. We have tested the role of an extended N-terminal arm in the structure and function of a periplasmic enzyme glucose fructose oxidoreductase (GFOR) from Zymomonas mobilis. We have determined the crystal structure of the NAD(+) complex of a truncated form of the enzyme, GFORDelta, in which the first 22 residues of the N-terminal arm of the mature protein have been deleted. The structure, refined at 2.7 A resolution (R(cryst)=24.1%, R(free)=28.4%), shows that the truncated form of the enzyme forms a dimer and implies that the N-terminal arm is essential for tetramer formation by wild-type GFOR. Truncation of the N-terminal arm also greatly increases the solvent exposure of the cofactor; since GFOR activity is dependent on retention of the cofactor during the catalytic cycle we conclude that the absence of GFOR activity in this mutant results from dissociation of the cofactor. The N-terminal arm thus determines the quaternary structure and the retention of the cofactor for GFOR activity and during translocation into the periplasm. The structure of GFORDelta also shows how an additional mutation, Ser64Asp, converts the strict NADP(+) specificity of wild-type GFOR to a dual NADP(+)/NAD(+) specificity. PMID- 11099382 TI - Crystal structures of a Rab protein in its inactive and active conformations. AB - We have determined crystal structures of Sec4, a member of the Rab family in the G protein superfamily, in two states: bound to GDP, and to a non-hydrolyzable GTP analog, guanosine-5'-(beta, gamma)-imidotriphosphate (GppNHp). This represents the first structure of a Rab protein bound to GDP. Sec4 in both states grossly resembles other G proteins bound to GDP and GppNHp. In Sec4-GppNHp, structural features common to active Rab proteins are observed. In Sec4-GDP, the switch I region is highly disordered and displaced relative to the switch I region of Ras GDP. In two of the four molecules of Sec4-GDP in the asymmetric unit of the Sec4 GDP crystals, the switch II region adopts a conformation similar to that seen in the structure of the small G protein Ran bound to GDP. This allows residues threonine 76, glutamate 80, and arginine 81 of Sec4 to make contacts with other conserved residues and water molecules important for nucleotide binding. In the other two molecules in the asymmetric unit, these interactions do not take place. This structural variability in both the switch I and switch II regions of GDP bound Sec4 provides a possible explanation for the high off-rate of GDP bound to Sec4, and suggests a mechanism for regulation of the GTPase cycle of Rab proteins by GDI proteins. PMID- 11099383 TI - Searching the protein structure databank with weak sequence patterns and structural constraints. AB - A method is described in which proteins that match PROSITE patterns are filtered by the root-mean-square deviation of the local 3D structures of the probe and target over the pattern components. This was found to increase the discrimination between true and false members of the protein family but was dependent on how unique the structural features in the pattern were compared to equivalent fragments extracted from the structure databank (for example; if the pattern fell in an alpha-helix, then discrimination was poor.) We then generalised the sequence patterns (by widening the range of amino acid residues allowed at each position) and monitored how well the structural information helped retain specificity. While the discrimination of the pure sequence pattern had generally disappeared at information content values less than ten bits, the discrimination of the combined sequence structure probe remained high at this point before following a similar decay. The displacement between these curves indicates that the structural component is, on average, equivalent to about ten bits. The sequence patterns were also filtered using the structure comparison program SAP, giving a global, rather than local "view" of the proteins. This allowed the information content of the sequence patterns to become even less specific but raised problems of whether some proteins encountered with the same fold but no PROSITE pattern should constitute family members. PMID- 11099384 TI - The TolA-recognition site of colicin N. ITC, SPR and stopped-flow fluorescence define a crucial 27-residue segment. AB - Colicins translocate across the Escherichia coli outer membrane and periplasm by interacting with several receptors. After first binding to the outer membrane surface receptors via their central region, they interact with TolA or TonB proteins via their N-terminal region. Colicin N residues critical to TolA binding have been discovered, but the full extent of any colicin TolA site is unknown. We present, for the first time, a fully mapped TolA binding site for a colicin. It was determined through the use of alanine-scanning mutants, glutathione S transferase fusion peptides and Biacore/fluorescence binding studies. The minimal TolA binding region is 27 residues and of similar size to the TolA binding region of bacteriophage g3p-D1 protein. Stopped-flow kinetic studies show that the binding to TolA follows slow association kinetics. The role of other E. coli Tol proteins in colicin translocation was also investigated. Isothermal titration microcalorimetry (ITC) and in vivo studies conclusively show that colicin N translocation does not require the presence of TolB. ITC also demonstrated colicin A interaction with TolB, and that colicin A in its native state does not interact with TolAII-III. Colicin N does not bind TolR-II. The TolA protein is shown to be unsuitable for direct immobilisation in Biacore analysis. PMID- 11099385 TI - Killing of Trypanosoma brucei by concanavalin A: structural basis of resistance in glycosylation mutants. AB - Concanavalin A (Con A) kills procyclic (insect) forms of Trypanosoma brucei by binding to N-glycans on EP-procyclin, a major surface glycosyl phosphatidylinositol (GPI)-anchored protein which is rich in Glu-Pro repeats. We have previously isolated and studied two procyclic mutants (ConA 1-1 and ConA 4 1) that are more resistant than wild-type (WT) to Con A killing. Although both mutants express the same altered oligosaccharides compared to WT cells, ConA 4-1 is considerably more resistant to lectin killing than is ConA 1-1. Thus, we looked for other alterations to account for the differences in sensitivity. Using mass spectrometry, together with chemical and enzymatic treatments, we found that both mutants express types of EP-procyclin that are either poorly expressed or not found at all in WT cells. ConA 1-1 expresses mainly EP1-3, a novel procyclin that contains 18 EP repeats, is partially N-glycosylated, and bears hybrid-type glycans. On the other hand, ConA 4-1 cells express almost exclusively EP2-3, a novel non-glycosylated procyclin isoform with 23 EP repeats and no site for glycosylation. The predominance of EP2-3 in ConA 4-1 cells explains their high resistance to ConA killing. Thus, switching the procyclin repertoire, a process that could be relevant to parasite development in the insect vector, modulates the sensitivity of trypanosomes to cytotoxic lectins. PMID- 11099386 TI - Characterization of heterodimeric alkaline phosphatases from Escherichia coli: an investigation of intragenic complementation. AB - Escherichia coli alkaline phosphatase (EC 3.1.3.1) belongs to a rare group of enzymes that exhibit intragenic complementation. When certain mutant versions of alkaline phosphatase are combined, the resulting heterodimeric enzymes exhibit a higher level of activity than would be expected based upon the relative activities of the parental enzymes. Nine previously identified alkaline phosphatase complementation mutants were re-examined in this work in order to determine a molecular explanation of intragenic complementation in this experimental system. The locations of these mutations were determined by DNA sequence analysis after PCR amplification of the phosphatase-negative phoA gene. Most of the mutations involved ligands to metal-binding sites. Each of the mutant enzymes was re-created by site-specific mutagenesis, expressed, purified, and kinetically characterized. To investigate cooperativity between the two subunits, we analyzed heterodimeric forms of some of the site-specific mutant enzymes. To enable the isolation of the heterodimeric alkaline phosphatase in pure form, the overall charge of one subunit was altered by replacing the C-terminal Lys residue with three Asp residues. This modification had no effect on the kinetic properties of the enzyme. Heterodimeric alkaline phosphatases were created using two methods: (1) in vitro formation by dissociation at acid pH followed by reassociation at slightly alkaline pH conditions in the presence of zinc and magnesium ions; and (2) in vivo expression from a plasmid carrying two different phoA genes. Increases in k(cat), as well as a large reduction in the p nitrophenyl phosphate K(m) were observed for certain combinations of mutant enzymes. These results suggest that the structural assembly of E. coli alkaline phosphatase into the dimer induces cooperative interactions between the monomers necessary for the formation of the functional form of the holoenzyme. PMID- 11099387 TI - Thermostability and thermoactivity of citrate synthases from the thermophilic and hyperthermophilic archaea, Thermoplasma acidophilum and Pyrococcus furiosus. AB - Citrate synthases from Thermoplasma acidophilum (optimal growth at 55 degrees C) and Pyrococcus furiosus (100 degrees C) are homo-dimeric enzymes that show a high degree of structural homology with each other, and thermostabilities commensurate with the environmental temperatures in which their host cells are found. A comparison of their atomic structures with citrate synthases from mesophilic and psychrophilic organisms has indicated the potential importance of inter-subunit contacts for thermostability, and here we report the construction and analysis of site-directed mutants of the two citrate synthases to investigate the contribution of these interactions. Three sets of mutants were made: (a) chimeric mutants where the large (inter-subunit contact) and small (catalytic) domains of the T. acidophilum and P. furiosus enzymes were swapped; (b) mutants of the P. furiosus citrate synthase where the inter-subunit ionic network is disrupted; and (c) P. furiosus citrate synthase mutants in which the C-terminal arms that wrap around their partner subunits have been deleted. All three sets of mutant enzymes were expressed as recombinant proteins in Escherichia coli and were found to be catalytically active. Kinetic parameters and the dependence of catalytic activity on temperature were determined, and the stability of each enzyme was analysed by irreversible thermal inactivation experiments. The chimeric mutants indicate that the thermostability of the whole enzyme is largely determined by the origin of the large, inter-subunit domain, whereas the dependence of catalytic activity on temperature is a function of the small domain. Disruption of the inter-subunit ionic network and prevention of the C-terminal interactions both generated enzymes that were substantially less thermostable. Taken together, these data demonstrate the crucial importance of the subunit contacts to the stability of these oligomeric enzymes. Additionally, they also provide a clear distinction between thermostability and thermoactivity, showing that stability is necessary for, but does not guarantee, catalytic activity at elevated temperatures. PMID- 11099388 TI - Macrophages secrete matrix metalloproteinase 9 covalently linked to the core protein of chondroitin sulphate proteoglycans. AB - Matrix metalloproteinases (MMPs) secreted from the leukemic macrophage cell-line THP-1 have been investigated. Under serum-free conditions, this cell-line synthesizes and secretes proMMP-9, which was detected in the culture medium as a monomer of 92 kDa, and in dimeric forms, including a homodimer of approximately 225 kDa. In addition, a new heterodimer complex is described, in which proMMP-9 is covalently linked to the core protein of chondroitin sulphate proteoglycan (CSPG) through one or more disulphide bridges. After SDS-PAGE electrophoresis, at least two forms of this complex were detected, a large form in the stacking gel and a smaller form with an estimated size of 300 kDa. When the CS chains were removed by chondroitin ABC lyase treatment, heterodimers of proMMP-9/CSPG core protein of approximately 145, 127 and 109 kDa were found, based on zymography and Western blots. Since as much as 10-15 % of the total proMMP-9 secreted from THP-1 cells was covalently linked to CSPG, this association may have important implications for transport, targetting and regulation of the enzyme activity. PMID- 11099389 TI - Splanchnic hyposensitivity to glypressin in a haemorrhage/transfused rat model of portal hypertension: role of nitric oxide and bradykinin. AB - Hyposensitivity to vasopressin is a well documented phenomenon in animals with portal hypertension and patients with cirrhosis subject to haemorrhage. Haemorrhage is associated with the endogenous release of bradykinin, which may subsequently stimulate the formation of nitric oxide (NO). The present study investigated the relative contribution of NO synthase (NOS) isoforms and the role of bradykinin in the pathogenesis of splanchnic hyposensitivity to a long-acting vasopressin analogue, glypressin, in rats with portal hypertension induced by partial portal vein ligation (PVL). At 14 days after the operation, systemic and portal haemodynamics were measured in stable or bleeding PVL rats receiving an intravenous infusion of glypressin (0.07 mg/kg). In the treatment groups, N(G) nitro-L-arginine methyl ester (L-NAME; a non-selective NOS inhibitor), L canavanine (a specific inhibitor of inducible NOS) or HOE 140 (a bradykinin B(2) receptor antagonist) was administered 45 min before the infusion of glypressin. In rats with a hypotensive haemorrhage, 4.5 ml of blood was withdrawn and 50% of the withdrawn blood was re-infused before the administration of glypressin or various inhibitors. Splanchnic hyposensitivity to glypressin was demonstrated in the haemorrhage/transfused PVL rats. The infusion of L-NAME elevated the mean arterial pressure in the bleeding PVL rats without the modulation of portal pressure. The addition of L-NAME or HOE 140, but not L-canavanine, significantly and similarly potentiated the portal-hypotensive effects of glypressin. It is concluded that constitutive NOS and bradykinin are responsible, at least partly, for the splanchnic hyposensitivity to glypressin observed in the early stages of the haemorrhage/transfused rat model of portal hypertension. PMID- 11099390 TI - Acute effects of oxygen and carbon dioxide on retinal vascular network geometry in hypertensive and normotensive subjects. AB - The optimal design of vascular networks maximizes circulatory efficiency while minimizing power costs. We investigated the effects of acute changes in vascular tone on retinal arteriolar network geometry. Six hypertensive and six normotensive subjects each breathed air, 5% CO(2) (with 12% O(2)), and 100% O(2) for 5 min periods in random order. Retinal photographs were taken at the end of each test period. Bifurcation angles and arteriolar diameters were measured using operator-directed image analysis, and junction exponents were calculated. Arteriolar diameters narrowed on breathing O(2). The magnitude of this change was significantly greater in normotensive than in hypertensive subjects. Angles narrowed in normotensive subjects, but not significantly in hypertensive subjects. Arteriolar diameters increased significantly on breathing CO(2) in normotensive but not in hypertensive subjects, but there were no changes in angles. Despite changes in diameter, junction exponents did not change under any conditions. Vascular reactivity in the retinal arteriolar bed appears to be diminished in hypertensive subjects. The failure of junction exponents to change, despite alterations in diameter, suggests that arteriolar diameters at retinal bifurcations adhere to optimality principles when exposed to acute vasoactive stress. As vasoconstriction is associated with the narrowing of bifurcation angles, previous observations showing narrowed angles in hypertensive subjects could be explained by increased tone in the retinal arteriolar bed. PMID- 11099391 TI - Induction of the stress response increases interleukin-6 production in the intestinal mucosa of endotoxaemic mice. AB - Previous studies suggest that production of interleukin-6 (IL-6) is increased in the intestinal mucosa during sepsis and endotoxaemia. We tested the hypothesis that mucosal IL-6 production during endotoxaemia is increased further by the heat shock (stress) response. The stress response was induced in mice by hyperthermia (rectal temperature of 42 degrees C for 3 min) or by intraperitoneal injection of sodium arsenite (10 mg/kg). At 2 h after induction of the stress response, groups of mice were injected subcutaneously with endotoxin (10 mg/kg) or sterile saline. IL-6 mRNA and protein levels in the jejunal mucosa were determined by an RNase protection assay and an ELISA respectively, and levels of hsp72 (heat-shock protein of 72 kDa) were determined by Western blot analysis. Hyperthermia and sodium arsenite increased hsp72 levels in the intestinal mucosa. IL-6 concentrations were increased in the jejunal mucosa of endotoxaemic mice, and this effect of endotoxaemia was potentiated by the stress response. Mucosal IL-6 mRNA levels were increased in endotoxaemic mice, and were increased further by the stress response. Thus it is concluded that mucosal IL-6 production during endotoxaemia may be further stimulated by the stress response. Increased IL-6 levels in the intestinal mucosa may be a potential mechanism by which the stress response exerts a protective effect during sepsis and endotoxaemia. PMID- 11099393 TI - Resting peripheral blood flow in normal pregnancy and in pre-eclampsia. AB - Multiple organ dysfunction followed by end organ failure occurs in pre-eclampsia. While one would intuitively reason that one of the factors contributing to the end organ failure is poor nutritional blood flow, this has yet to be demonstrated. The aim of the present study was to determine whether changes in resting nutritional blood flow occur in pre-eclampsia. We used strain-gauge plethysmography to study calf blood flow in 19 women with pre-eclampsia, 13 normal pregnant women and 17 non-pregnant controls. We reasoned that, since the calf comprises mostly skeletal muscle, without anastomotic channels, blood flowing through this region would primarily reflect nutritive flow. Calf blood flow was significantly reduced in women with pre-eclampsia (1.95+/-0.9 ml.min( 1).100 ml(-1)) compared with normal pregnant (3.9+/-1.4 ml.min(-1).100 ml(-1)) and non-pregnant (3.8+/-1.0 ml.min(-1).100 ml(-1)) women (P=0.0004 and P=0.0005 respectively; ANOVA). Blood flow in pre-eclampsia was also correlated significantly with platelet count as an index of disease severity. In addition, there was a significant negative correlation between blood flow and systolic blood pressure (r=-0.69, P=0.004) in the women with pre-eclampsia. These findings support the hypothesis that nutritional blood flow is reduced in pre-eclampsia. We suggest that measurement of resting calf blood flow could give a non-invasive index of deterioration of nutritive blood flow to vital organs in pre-eclampsia. PMID- 11099392 TI - Role of the L-arginine/nitric oxide pathway in ischaemic/reoxygenation injury of the human myocardium. AB - The role of the L-arginine/nitric oxide (NO) pathway in myocardial ischaemic/reperfusion injury remains controversial in experimental animal models. The aim of the present studies was to investigate the role of this pathway in the human myocardium. Myocardial specimens from right atrial appendages of patients undergoing elective coronary bypass graft surgery were incubated in crystalloid buffer at 37 degrees C and subjected to 120 min of simulated ischaemia followed by 120 min of reoxygenation. Tested drugs were added 15 min before ischaemia, and maintained during ischaemia and throughout reoxygenation. Ischaemia resulted in severe myocardial damage, as assessed by the leakage of lactate dehydrogenase (LDH) into the incubation medium and by the capacity of the tissue to reduce 3 (4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) to formazan product. L-Arginine (10 mM), a precursor of NO, significantly decreased LDH leakage (from 9.0+/-0.6 to 5.3+/-0.3 units/g wet wt; P<0.05), but had no effect on MTT reduction or oxygen consumption. D-Arginine (10 mM), N(G)-nitro-L-arginine methyl ester (L-NAME; 0.5 mM), an NO synthase inhibitor, and S-nitroso-N acetylpenicillamine (at 1, 100, 500 and 1000 microM), an NO donor, had no significant effects on the measured indices, and L-NAME did not reverse the protection afforded by L-arginine against LDH leakage. In addition, the formation of nitrotyrosine was not influenced by ischaemia/reoxygenation alone or by the agents investigated. In conclusion, these data suggest that L-arginine affords modest protection against ischaemic/reoxygenation injury of the human myocardium, an action that is NO-independent, and that NO metabolism does not play a significant role in this model. PMID- 11099394 TI - Conjugated linoleic acid induces lipid peroxidation in men with abdominal obesity. AB - Conjugated linoleic acid (CLA) has been shown in experimental studies to have chemoprotective properties, and may decrease the deposition of body fat. CLA is prone to oxidation, and it has been suggested that increased lipid oxidation may contribute to the anti-tumorigenic effects of this agent. The present study investigates the urinary levels of 8-iso-prostaglandin F(2alpha) (8-iso PGF(2alpha)), a major isoprostane, and of 15-oxo-dihydro-PGF(2alpha), a major metabolite of PGF(2alpha), as indicators of non-enzymic and enzymic arachidonic acid oxidation respectively after dietary supplementation with CLA in middle-aged men (mean age 53 years) with abdominal obesity for 1 month in a randomized controlled trial. Significant increases in the levels of both 8-iso-PGF(2alpha) and 15-oxo-dihydro-PGF(2alpha) in urine (P<0. 0001 and P=0.0013 respectively) were observed after 1 month of daily CLA intake (4.2 g/day) as compared with the control group. The lipid peroxidation parameters had returned to their basal levels at 2 weeks after the cessation of CLA intake, and remained at the same levels for a further 2 weeks until the end of the study. CLA had no effect on serum alpha-tocopherol and gamma-tocopherol levels, or on the urinary levels of 2,3-dinor-thromboxane B(2). Thus CLA may induce both non-enzymic and enzymic lipid peroxidation in vivo in middle-aged men with abdominal obesity, without any side effects. The consequences of the increased lipid peroxidation after CLA supplementation are unknown. PMID- 11099395 TI - Non-invasive quantification of liver perfusion with dynamic computed tomography and a dual-input one-compartmental model. AB - Various liver diseases lead to significant alterations of the hepatic microcirculation. Therefore, quantification of hepatic perfusion has the potential to improve the assessment and management of liver diseases. Most methods used to quantify liver perfusion are invasive or controversial. This paper describes and validates a non-invasive method for the quantification of liver perfusion using computed tomography (CT). Dynamic single-section CT of the liver was performed after intravenous bolus administration of a low-molecular mass iodinated contrast agent. Hepatic, aortic and portal-venous time-density curves were fitted with a dual-input one-compartmental model to calculate liver perfusion. Validation studies consisted of simultaneous measurements of hepatic perfusion with CT and with radiolabelled microspheres in rabbits at rest and after adenosine infusion. The feasibility and reproducibility of the CT method in humans was assessed by three observers in 10 patients without liver disease. In rabbits, significant correlations were observed between perfusion measurements obtained with CT and with microspheres (r=0.92 for total liver perfusion, r=0.81 for arterial perfusion and r=0.85 for portal perfusion). In patients, total liver plasma perfusion measured with CT was 112+/-28 ml.min(-1).100 ml(-1), arterial plasma perfusion was 18+/-12 ml.min(-1).100 ml(-1) and portal plasma perfusion was 93+/-31 ml.min(-1).100 ml(-1). The measurements obtained by the three observers were not significantly different from each other (P>0.1). Our results indicate that dynamic CT combined with a dual-input one-compartmental model provides a valid and reliable method for the non-invasive quantification of perfusion in the normal liver. PMID- 11099396 TI - Elevated circulating leptin levels in arterial hypertension: relationship to arteriovenous overflow and extraction of leptin. AB - Leptin, a peptide hormone produced mainly in fat cells, appears to be important for the regulation of metabolism, insulin secretion/sensitivity and body weight. Recently, elevated plasma leptin levels have been reported in patients with arterial hypertension. Because a change in circulating leptin concentrations in such patients could be caused by altered rates of production or disposal, or both, the aim of the present study was to identify regions of leptin overflow into the bloodstream and of leptin extraction. Patients with arterial hypertension (n=12) and normotensive controls (n=20) were studied during catheterization with elective blood sampling from different vascular beds (artery, and renal, hepatic, iliac and cubital veins). Plasma leptin was determined by a radioimmunoassay. Patients with hypertension had significantly elevated levels of circulating leptin (12.8 ng/l, compared with 4.1 ng/l in the controls; P<0.001), and this was also the case when adjusted for body mass index (BMI) [0.435 and 0.167 ng/l per unit BMI (kg/m(2)) respectively; P<0.001]. Circulating leptin was directly related to arterial blood pressure (r=0.38-0.62, P>Edg-4) by reverse transcription-PCR in PT cells. Our data suggest that LPA may behave as a local growth factor in PT cells following tubular injury. PMID- 11099402 TI - A compound heterozygote for a novel missense mutation (G105R) in exon 3 and a missense mutation (D204E) in exon 5 of the lipoprotein lipase gene in a Japanese infant with hyperchylomicronaemia. AB - We systematically investigated the molecular defects resulting in primary lipoprotein lipase (LPL) deficiency in a Japanese male infant (hereafter called 'the patient') with severe fasting hypertriglyceridaemia (type I hyperlipoproteinaemia). The primary LPL deficiency was diagnosed on the basis of the findings that no LPL activity was detected in post-heparin plasma (PHP) and that the immunoreactive LPL mass in PHP was less than 2% of the control level. The patient was a compound heterozygote for a novel missense mutation (G(568)GA- >AGA/Gly(105)-->Arg; G105R) in exon 3 and a missense mutation (GAC(867)- >GAG/Asp(204)-->Glu; D204E) in exon 5 of the LPL gene. The biological significance of both missense mutations was examined by an in vitro study of the expression of the mutant G105R LPL cDNA and D204E LPL cDNA in COS-1 cells. Both mutant LPLs were catalytically inactive and were barely released by heparin from the expressing COS-1 cells. These findings explain the failure to detect LPL activity and immunoreactive LPL mass in the patient's PHP. The G105R allele could be detected by digestion with the BsmAI restriction enzyme, and the D204E allele by digestion with HincII. The patient inherited the G105R allele from his mother and the D204E allele from his father. His parents were heterozygotes for the corresponding mutant allele, but normolipidaemic. The novel G105R missense mutation could not be detected by conventional analysis of single-strand conformation polymorphism, but it was identified by extensive sequencing of the entire exons and their flanking regions in the LPL gene. PMID- 11099403 TI - Selenium supplementation and selenoenzyme activity. PMID- 11099405 TI - How cells handle cholesterol. AB - Cholesterol plays an indispensable role in regulating the properties of cell membranes in mammalian cells. Recent advances suggest that cholesterol exerts many of its actions mainly by maintaining sphingolipid rafts in a functional state. How rafts contribute to cholesterol metabolism and transport in the cell is still an open issue. It has long been known that cellular cholesterol levels are precisely controlled by biosynthesis, efflux from cells, and influx of lipoprotein cholesterol into cells. The regulation of cholesterol homeostasis is now receiving a new focus, and this changed perspective may throw light on diseases caused by cholesterol excess, the prime example being atherosclerosis. PMID- 11099406 TI - Self-mode-locking of quantum cascade lasers with giant ultrafast optical nonlinearities. AB - We report on the generation of picosecond self-mode-locked pulses from midinfrared quantum cascade lasers, at wavelengths within the important molecular fingerprint region. These devices are based on intersubband electron transitions in semiconductor nanostructures, which are characterized by some of the largest optical nonlinearities observed in nature and by picosecond relaxation lifetimes. Our results are interpreted with a model in which one of these nonlinearities, the intensity-dependent refractive index of the lasing transition, creates a nonlinear waveguide where the optical losses decrease with increasing intensity. This favors the generation of ultrashort pulses, because of their larger instantaneous intensity relative to continuous-wave emission. PMID- 11099404 TI - Autophagy as a regulated pathway of cellular degradation. AB - Macroautophagy is a dynamic process involving the rearrangement of subcellular membranes to sequester cytoplasm and organelles for delivery to the lysosome or vacuole where the sequestered cargo is degraded and recycled. This process takes place in all eukaryotic cells. It is highly regulated through the action of various kinases, phosphatases, and guanosine triphosphatases (GTPases). The core protein machinery that is necessary to drive formation and consumption of intermediates in the macroautophagy pathway includes a ubiquitin-like protein conjugation system and a protein complex that directs membrane docking and fusion at the lysosome or vacuole. Macroautophagy plays an important role in developmental processes, human disease, and cellular response to nutrient deprivation. PMID- 11099407 TI - Tunable resistance of a carbon nanotube-graphite interface. AB - The transfer of electrons from one material to another is usually described in terms of energy conservation, with no attention being paid to momentum conservation. Here we present results on the junction resistance between a carbon nanotube and a graphite substrate and show that details of momentum conservation also can change the contact resistance. By changing the angular alignment of the atomic lattices, we found that contact resistance varied by more than an order of magnitude in a controlled and reproducible fashion, indicating that momentum conservation, in addition to energy conservation, can dictate the junction resistance in graphene systems such as carbon nanotube junctions and devices. PMID- 11099409 TI - Tropical climate at the last glacial maximum inferred from glacier mass-balance modeling. AB - Model-derived equilibrium line altitudes (ELAs) of former tropical glaciers support arguments, based on other paleoclimate data, for both the magnitude and spatial pattern of terrestrial cooling in the tropics at the last glacial maximum (LGM). Relative to the present, LGM ELAs were maintained by air temperatures that were 3.5 degrees to 6.6 degrees C lower and precipitation that ranged from 63% wetter in Hawaii to 25% drier on Mt. Kenya, Africa. Our results imply the need for a approximately 3 degrees C cooling of LGM sea surface temperatures in the western Pacific warm pool. Sensitivity tests suggest that LGM ELAs could have persisted until 16,000 years before the present in the Peruvian Andes and on Papua, New Guinea. PMID- 11099408 TI - Formation of sphalerite (ZnS) deposits in natural biofilms of sulfate-reducing bacteria. AB - Abundant, micrometer-scale, spherical aggregates of 2- to 5-nanometer-diameter sphalerite (ZnS) particles formed within natural biofilms dominated by relatively aerotolerant sulfate-reducing bacteria of the family Desulfobacteriaceae. The biofilm zinc concentration is about 10(6) times that of associated groundwater (0.09 to 1.1 parts per million zinc). Sphalerite also concentrates arsenic (0.01 weight %) and selenium (0.004 weight %). The almost monomineralic product results from buffering of sulfide concentrations at low values by sphalerite precipitation. These results show how microbes control metal concentrations in groundwater- and wetland-based remediation systems and suggest biological routes for formation of some low-temperature ZnS deposits. PMID- 11099410 TI - The formation of chondrules at high gas pressures in the solar nebula. AB - High-precision magnesium isotope measurements of whole chondrules from the Allende carbonaceous chondrite meteorite show that some aluminum-rich Allende chondrules formed at or near the time of formation of calcium-aluminum-rich inclusions and that some others formed later and incorporated precursors previously enriched in magnesium-26. Chondrule magnesium-25/magnesium-24 correlates with [magnesium]/[aluminum] and size, the aluminum-rich, smaller chondrules being the most enriched in the heavy isotopes of magnesium. These relations imply that high gas pressures prevailed during chondrule formation in the solar nebula. PMID- 11099411 TI - Support for the lunar cataclysm hypothesis from lunar meteorite impact melt ages. AB - Lunar meteorites represent a more random sampling of lunar material than the Apollo or Luna collections and, as such, lunar meteorite impact melt ages are the most important data in nearly 30 years with which to reexamine the lunar cataclysm hypothesis. Within the lunar meteorite breccias MAC 88105, QUE 93069, DaG 262, and DaG 400, seven to nine different impact events are represented with 40Ar-39Ar ages between 2.76 and 3.92 billion years ago (Ga). The lack of impact melt older than 3.92 Ga supports the concept of a short, intense period of bombardment in the Earth-moon system at approximately 3.9 Ga. This was an anomalous spike of impact activity on the otherwise declining impact- frequency curve. PMID- 11099412 TI - Nitric acid trihydrate (NAT) in polar stratospheric clouds. AB - A comprehensive investigation of polar stratospheric clouds was performed on 25 January 2000 with instruments onboard a balloon gondola flown from Kiruna, Sweden. Cloud layers were repeatedly encountered at altitudes between 20 and 24 kilometers over a wide range of atmospheric temperatures (185 to 197 kelvin). Particle composition analysis showed that a large fraction of the cloud layers was composed of nitric acid trihydrate (NAT) particles, containing water and nitric acid at a molar ratio of 3:1; this confirmed that these long-sought solid crystals exist well above ice formation temperatures. The presence of NAT particles enhances the potential for chlorine activation with subsequent ozone destruction in polar regions, particularly in early and late winter. PMID- 11099413 TI - Reinterpreting space, time lags, and functional responses in ecological models. AB - Natural enemy-victim interactions are of major applied importance and of fundamental interest to ecologists. A key question is what stabilizes these interactions, allowing the long-term coexistence of the two species. Three main theoretical explanations have been proposed: behavioral responses, time-dependent factors such as delayed density dependence, and spatial heterogeneity. Here, using the powerful moment-closure technique, we show a fundamental equivalence between these three elements. Limited movement by organisms is a ubiquitous feature of ecological systems, allowing spatial structure to develop; we show that the effects of this can be naturally described in terms of time lags or within-generation functional responses. PMID- 11099414 TI - Posttranslational N-myristoylation of BID as a molecular switch for targeting mitochondria and apoptosis. AB - Many apoptotic molecules relocate subcellularly in cells undergoing apoptosis. The pro-apoptotic protein BID underwent posttranslational (rather than classic cotranslational) N-myristoylation when cleavage by caspase 8 caused exposure of a glycine residue. N-myristoylation enabled the targeting of a complex of p7 and myristoylated p15 fragments of BID to artificial membranes bearing the lipid composition of mitochondria, as well as to intact mitochondria. This post proteolytic N-myristoylation serves as an activating switch, enhancing BID induced release of cytochrome c and cell death. PMID- 11099415 TI - Requirement of the RNA editing deaminase ADAR1 gene for embryonic erythropoiesis. AB - The members of the ADAR (adenosine deaminase acting on RNA) gene family are involved in site-selective RNA editing that changes adenosine residues of target substrate RNAs to inosine. Analysis of staged chimeric mouse embryos with a high contribution from embryonic stem cells with a functional null allele for ADAR1 revealed a heterozygous embryonic-lethal phenotype. Most ADAR1+/- chimeric embryos died before embryonic day 14 with defects in the hematopoietic system. Our results suggest the importance of regulated levels of ADAR1 expression, which is critical for embryonic erythropoiesis in the liver. PMID- 11099416 TI - Down-regulation of the macrophage lineage through interaction with OX2 (CD200). AB - OX2 (CD200) is a broadly expressed membrane glycoprotein, shown here to be important for regulation of the macrophage lineage. In mice lacking CD200, macrophage lineage cells, including brain microglia, exhibited an activated phenotype and were more numerous. Upon facial nerve transection, damaged CD200 deficient neurons elicited an accelerated microglial response. Lack of CD200 resulted in a more rapid onset of experimental autoimmune encephalomyelitis (EAE). Outside the brain, disruption of CD200-CD200 receptor interaction precipitated susceptibility to collagen-induced arthritis (CIA) in mice normally resistant to this disease. Thus, in diverse tissues OX2 delivers an inhibitory signal for the macrophage lineage. PMID- 11099417 TI - Accumulation of dietary cholesterol in sitosterolemia caused by mutations in adjacent ABC transporters. AB - In healthy individuals, acute changes in cholesterol intake produce modest changes in plasma cholesterol levels. A striking exception occurs in sitosterolemia, an autosomal recessive disorder characterized by increased intestinal absorption and decreased biliary excretion of dietary sterols, hypercholesterolemia, and premature coronary atherosclerosis. We identified seven different mutations in two adjacent, oppositely oriented genes that encode new members of the adenosine triphosphate (ATP)-binding cassette (ABC) transporter family (six mutations in ABCG8 and one in ABCG5) in nine patients with sitosterolemia. The two genes are expressed at highest levels in liver and intestine and, in mice, cholesterol feeding up-regulates expressions of both genes. These data suggest that ABCG5 and ABCG8 normally cooperate to limit intestinal absorption and to promote biliary excretion of sterols, and that mutated forms of these transporters predispose to sterol accumulation and atherosclerosis. PMID- 11099418 TI - From marrow to brain: expression of neuronal phenotypes in adult mice. AB - After intravascular delivery of genetically marked adult mouse bone marrow into lethally irradiated normal adult hosts, donor-derived cells expressing neuronal proteins (neuronal phenotypes) developed in the central nervous system. Flow cytometry revealed a population of donor-derived cells in the brain with characteristics distinct from bone marrow. Confocal microscopy of individual cells showed that hundreds of marrow-derived cells in brain sections expressed gene products typical of neurons (NeuN, 200-kilodalton neurofilament, and class III beta-tubulin) and were able to activate the transcription factor cAMP response element-binding protein (CREB). The generation of neuronal phenotypes in the adult brain 1 to 6 months after an adult bone marrow transplant demonstrates a remarkable plasticity of adult tissues with potential clinical applications. PMID- 11099419 TI - Turning blood into brain: cells bearing neuronal antigens generated in vivo from bone marrow. AB - Bone marrow stem cells give rise to a variety of hematopoietic lineages and repopulate the blood throughout adult life. We show that, in a strain of mice incapable of developing cells of the myeloid and lymphoid lineages, transplanted adult bone marrow cells migrated into the brain and differentiated into cells that expressed neuron-specific antigens. These findings raise the possibility that bone marrow-derived cells may provide an alternative source of neurons in patients with neurodegenerative diseases or central nervous system injury. PMID- 11099420 TI - Coding the location of the arm by sight. AB - Area 5 in the parietal lobe of the primate brain is thought to be involved in monitoring the posture and movement of the body. In this study, neurons in monkey area 5 were found to encode the position of the monkey's arm while it was covered from view. The same neurons also responded to the position of a visible, realistic false arm. The neurons were not sensitive to the sight of unrealistic substitutes for the arm and were able to distinguish a right from a left arm. These neurons appear to combine visual and somatosensory signals in order to monitor the configuration of the limbs. They could form the basis of the complex body schema that we constantly use to adjust posture and guide movement. PMID- 11099421 TI - Neurons in monkey prefrontal cortex that track past or predict future performance. AB - Although frontal cortex is thought to be important in controlling behavior across long periods of time, most studies of this area concentrate on neuronal responses instantaneously relevant to the current task. In order to investigate the relationship of frontal activity to behavior over longer time periods, we trained rhesus monkeys on a difficult oculomotor task. Their performance fluctuated during the day, and the activity of prefrontal neurons, even measured while the monkeys waited for the targets to appear at the beginning of each set of trials, correlated with performance in a probabilistic rather than a determinist manner: neurons reflected past or predicted future performance, much more than they reflected current performance. We suggest that this activity is related to processes such as arousal or motivation that set the tone for behavior rather than controlling it on a millisecond basis, and could result from ascending pathways that utilize slow, second-messenger synaptic processes. PMID- 11099422 TI - Rapid induction of mild cerebral hypothermia by cold aortic flush achieves normal recovery in a dog outcome model with 20-minute exsanguination cardiac arrest. AB - OBJECTIVES: Resuscitation attempts in trauma victims who suffer cardiac arrest (CA) from exsanguination almost always fail. The authors hypothesized that an aortic arch flush with cold normal saline solution (NSS) at the start of exsanguination CA can preserve cerebral viability during 20-minute no-flow. METHODS: Twelve dogs were exsanguinated over 5 minutes to CA of 20-minute no flow, resuscitated by cardiopulmonary bypass, followed by post-CA mild hypothermia (34 degrees C) continued to 12 hours, controlled ventilation to 20 hours, and intensive care to 72 hours. At CA 2 minutes, the dogs received a 500 mL flush of NSS at either 24 degrees C (group 1, n = 6) or 4 degrees C (group 2, n = 6), using a balloon-tipped catheter inserted via the femoral artery into the descending thoracic aorta. RESULTS: The flush at 24 degrees C (group 1) decreased tympanic membrane temperature [mean (+/-SD)] from 37.5 degrees C (+/-0.1) to 35.7 degrees C (+/-0.2); the flush at 4 degrees C (group 2) to 34.0 degrees C (+/-1.1) (p = 0.005). In group 1, one dog achieved overall performance category (OPC) 2 (moderate disability), one OPC 3 (severe disability), and four OPC 4 (coma). In group 2, four dogs achieved OPC 1 (normal), one OPC 2, and one OPC 3 (p = 0.008). Neurologic deficit scores (0-10% normal, 100% brain death) [median (25th-75th percentile)] were 62% (40-66) in group 1 and 5% (0-19) in group 2 (p = 0.01). Total brain histologic damage scores were 130 (62-137) in group 1 and 24 (10-55) in group 2 (p = 0.008). CONCLUSIONS: Aortic arch flush of 4 degrees C at the start of CA of 20 minutes rapidly induces mild cerebral hypothermia and can lead to normal functional recovery with minimal histologic brain damage. The same model with aortic arch flush of 24 degrees C results in survival with brain damage in all dogs, which makes it suitable for testing other (e.g., pharmacologic) preservation potentials. PMID- 11099423 TI - Histopathologic effects of cutaneous tape stripping in pigs. AB - OBJECTIVE: The stratum corneum (SC) is the major barrier to topical absorption of medications. Skin tape stripping (TS) removes the stratum corneum, allowing more rapid absorption of drugs such as local anesthetics. Prior to evaluating TS in human clinical trials, this study was performed to evaluate its immediate and delayed histopathologic effects in swine. METHODS: This was a prospective, interventional, longitudinal, experimental animal study using two isoflurane anesthetized young swine. Cellophane tape was applied to the skin of clipped swine flanks and peeled away 10, 15, 20, 25, or 30 times. Each level of tape stripping was performed three times in each pig. Full-thickness biopsies were taken at 30 minutes and two weeks later for blinded histopathologic evaluation by a dermatopathologist using randomly ordered hematoxylin-and-eosin-stained tissue sections and conventional light microscopy. The absolute thickness of the cornified layer was measured and compared with normal (unstripped) cornified layer. RESULTS: Tape stripping up to 30 times produces thinning of the SC without detectable changes in the underlying epidermis and dermis at 30 minutes and 14 days post-stripping. The degree of thinning was proportional to the number of tape applications. Complete recovery of the cornified layer was noted at two weeks. There were no adverse effects, such as infection, scarring, or inflammatory cell infiltrates. CONCLUSIONS: Under the conditions studied, TS not only appears safe, but appears to produce no long-term sequelae. PMID- 11099424 TI - Luminarias PMID- 11099425 TI - Critical care in the emergency department: A physiologic assessment and outcome evaluation. AB - OBJECTIVES: The changing landscape of health care in this country has seen an increase in the delivery of care to critically ill patients in the emergency department (ED). However, methodologies to assess care and outcomes similar to those used in the intensive care unit (ICU) are currently lacking in this setting. This study examined the impact of ED intervention on morbidity and mortality using the Acute Physiology and Chronic Health Evaluation (APACHE II), the Simplified Acute Physiology Score (SAPS II), and the Multiple Organ Dysfunction Score (MODS). METHODS: This was a prospective, observational cohort study over a three-month period. Critically ill adult patients presenting to a large urban ED and requiring ICU admission were enrolled. APACHE II, SAPS II, and MODS scores and predicted mortality were obtained at ED admission, ED discharge, and 24, 48, and 72 hours in the ICU. In-hospital mortality was recorded. RESULTS: Eighty-one patients aged 64 +/- 18 years were enrolled during the study period, with a 30.9% in-hospital mortality. The ED length of stay was 5.9 +/- 2.7 hours and the hospital length of stay was 12.2 +/- 16.6 days. Nine (11.1%) patients initially accepted for ICU admission were later admitted to the general ward after ED intervention. Septic shock was the predominant admitting diagnosis. At ED admission, there was a significantly higher APACHE II score in nonsurvivors (23.0 +/- 6.0) vs survivors (19.8 +/- 6.5, p = 0.04), while there was no significant difference in SAPS II or MODS scores. The APACHE II, SAPS II, and MODS scores were significantly lower in survivors than nonsurvivors throughout the hospital stay (p 90% yet <94%, was seen in only four patients. All four patients responded to blow-by oxygen and required no other intervention. CONCLUSION: Intravenous pentobarbital is more effective than IV midazolam for sedation of children requiring CT imaging. PMID- 11099428 TI - Sports-related injuries in children. AB - OBJECTIVE: To describe the demographics and types of sports-related injuries (SRIs) in children. METHODS: The authors performed a retrospective chart review of children 5-18 years of age diagnosed as having an SRI in a pediatric emergency department (ED) during a two-year period. Patients were identified by ICD-9 codes. Data collected were age, sex, sport, ED interventions, consultations, mechanism, location, and injury type. Pairwise comparisons were reported as odds ratios with 95% confidence intervals. RESULTS: Six hundred seventy-seven SRIs fit the inclusion criteria; 480 of the patients were male (71%). The mean ages of the males and females were 13.0 years (SD +/- 3.0 yr) and 12.4 years (SD +/- 2.9 yr), respectively. The six most common sports implicated were basketball (19.5%), football (17.1%), baseball/softball (14.9%), soccer (14.2%), in-line skating (Rollerblading)/skating (5.7%), and hockey (4.6%). Sprains/strains (32.0%), fractures (29.4%), contusions/abrasions (19. 3%), and lacerations (9.7%) accounted for 90% of injury types. Pairwise comparison of the four injury types in the six sports listed showed significant associations for contusions/abrasions in baseball, sprains/strains in basketball, fractures in Rollerblading/skating, and lacerations in hockey. Age variance, including all sports, of the younger group (5-11 yr) in fractures and the older group (12-18 yr) in sprains was significant. The most common injury location was wrist/hand (28%), followed by head/face (22%) and ankle/foot (18%). Each had significant sport-specific predilections. Contact with person or object was the mechanism for >50% of the SRIs. Sport-specific mechanisms followed lines drawn from the sport-specific injury types and locations. CONCLUSIONS: The pediatric age group incurs a variety of injuries in numerous sports with diverse sex, age, mechanism, location, injury type, and sport-specific differences. PMID- 11099429 TI - Acceptability of emergency department-based screening and brief intervention for alcohol problems. AB - OBJECTIVES: To adapt screening and brief intervention for alcohol problems (SBI) to a high-volume emergency department (ED) setting and evaluate its acceptability to patients. METHODS: Patients at a large public-hospital ED were screened with the Alcohol Use Disorders Identification Test (AUDIT). Screen-positive drinkers (AUDIT score >/= 6) were provided brief, on-site counseling and referral as needed. Three months later, project staff blinded to baseline measures reassessed alcohol intake, alcohol-related harm, alcohol dependence symptoms, and readiness to change. RESULTS: Of 1, 034 patients approached, 78.3% (810) consented to participate (95% CI = 75.5% to 81.2%), and 21.2% (172) screened positive (95% CI = 18.4% to 24.0%). Of 88 patients with complete intervention data, 94.3% (83) accepted an intervention (95% CI = 89.5% to 99.2%), with acceptance rates ranging from 93% to 100% across four alcohol-problem-severity levels (p = 0.7). A majority (59.0%) set goals to decrease or stop drinking (95% CI = 48.4% to 69.6%). The group recontacted (n = 23) experienced statistically significant decreases in alcohol intake, alcohol-related harm, and dependence symptoms, with measures decreasing for 68%, 52%, and 61% of the patients. Readiness to change also showed statistically significant improvement, with scores increasing for 43% of the patients. Moreover, two-thirds of the patients (15/23) reported at follow up that SBI was a helpful part of their ED visit. CONCLUSIONS: High rates of consent and acceptance of counseling for alcohol problems by patients across a wide range of problem severity indicate that this protocol was acceptable to at risk patients in a public-hospital ED. Improvements in alcohol-related outcome measures at follow-up were strong enough to warrant controlled studies of intervention efficacy. PMID- 11099430 TI - Breast cancer knowledge and preventive behaviors: An urban emergency department based survey. AB - OBJECTIVE: To assess general knowledge and preventive behaviors regarding breast cancer among women who present to an urban emergency department. METHODS: During a six-month study period, a convenience sampling of women aged 21 years and older who were in treatment and waiting areas was surveyed. The anonymous written survey asked about demographic variables, knowledge, and preventive behaviors regarding breast cancer. Knowledge was assessed with questions about the recommended frequency of breast self-examination and the recommended age for first mammography. Performance was assessed by questions about breast self-exam and mammography. Subgroup analysis was done by age (above and below 40 years old), race, income (above and below the median), insurance type, history of breast lump, and family history (FH) of breast cancer. RESULTS: Four hundred women completed surveys. Two hundred twelve (53%) correctly knew the answers to the two knowledge questions. Knowledge was greater in women with private insurance. Knowledge of the frequency of breast self-exam was significantly greater among whites and Native Americans than among African Americans, Asians, or Hispanics. Stated performance of preventive behaviors was 72% (288) for breast self-exam and for mammography. Preventive behaviors were significantly more likely to be performed by higher-income and privately-insured women. Breast self exam was more likely to be done in older women, those with a history of a breast lump, and those with a FH of breast cancer. CONCLUSIONS: Women with lower income and without private insurance were less likely to be knowledgeable and practice preventive measures for detecting breast disease. PMID- 11099431 TI - Emergency medicine residency applicant educational debt: relationship with attitude toward training and moonlighting. AB - OBJECTIVES: Heated debate persists regarding the role of resident moonlighting in emergency medicine (EM). The attitudes of EM residency applicants have not been assessed. The objectives of this study were to assess: 1) the level of educational debt among EM residency applicants, 2) their perception of increased risk potential to patients from unsupervised EM resident practice, and 3) their opposition to laws restricting moonlighting. The authors then report the relationship between the degree of indebtedness and these stated positions. METHODS: Fifty-four EM residency programs returned 393 responses to a 1996 anonymous survey. Applicants recorded: 1) their indebtedness, 2) whether they believed that EDs should hire only physicians who have completed full training in an EM residency, and 3) whether they believed that unsupervised EM practice prior to completing EM training carries a higher risk of adverse patient outcomes. The authors used a t-test and logistic regression to determine whether there was any significant difference in debt between responders who answered yes and those who answered no to the various questions. A p-value < 0.05 was considered significant. RESULTS: The mean +/- SD debt was $72,290 +/- 48,683 (median $70,000). Most EM applicants (84.8%) agreed that unsupervised medical care by EM residents carries a higher risk of adverse patient outcomes. Paradoxically, only half the applicants opposed a moonlighting ban. Responses did not statistically correlate with educational debt. CONCLUSIONS: Emergency medicine residency applicant debt is large. The EM applicants' opposition to laws that would restrict moonlighting was mixed. This was inconsistent with the majority acknowledging an increased risk potential to patients. Nearly all EM applicants would still select EM as a career, even if moonlighting were to be banned. PMID- 11099432 TI - Resident perception of academic skills training and impact on academic career choice. AB - OBJECTIVES: 1) To evaluate residents' perceptions of the quality of training in basic academic skills and the availability and quality of research resources during residency; 2) to evaluate the association between these attitudes and choice of an academic career; and 3) to assess residents' attitudes toward the importance of postgraduate fellowship training for success in an academic career. METHODS: A 15-item survey was administered to all U.S. emergency medicine (EM) residents in conjunction with the February 1997 American Board of Emergency Medicine (ABEM) In-service Examination. The survey assessed resident interest in a career in academic EM, and resident perception of the general quality of training in academic (research and teaching) skills. Residents were also asked to rate the quality of their training in the following specific academic skills: medical and grant writing, bedside teaching, lecturing, the use of computers, study design, statistics, and the use of audiovisual aids. Resident perceptions of the availability of the following resources were also assessed: teaching and research role models, data collection and analysis support, laboratory facilities, financial support of research, research fundamentals lectures, and computers. RESULTS: The response rate was 93%. Forty-four percent of the respondents were interested in academic EM, 36.6% were undecided, and 19.6% were not interested in an academic career. On a scale of 1 (unprepared) to 5 (well prepared), the residents rated their overall preparedness for an academic career fairly high (3.97 [0.86]). In contrast, they perceived the quality of their training in the specific academic skill areas assessed and research resource availability to be only fair. Despite resident perception of relatively inadequate training in basic academic skills, only 24% of the respondents indicated that they believed fellowship training was important for success in an academic career. Logistic regression analyses demonstrated that participation in a research project in medical school, the length of the training program (4- vs 3 year), being a first-year resident, and a better perception of one's overall academic skill preparation were factors independently associated with having a greater interest in an academic career. CONCLUSIONS: A relatively high percentage of residents initially express an interest in an academic career, but this interest wanes as residency progresses. A minority of residents believe that their training provides them with the specific skills needed to succeed in academics, or with adequate exposure to research resources or mentors. Emergency medicine may be able to increase the number of qualified academic faculty by recruiting medical students with prior research experience, and providing residents with better research training and role models. PMID- 11099433 TI - Emergency department use by adult medicaid patients after implementation of managed care. AB - OBJECTIVE: To determine whether the advent of a mandatory Medicaid managed care (MMC) plan had any effect on emergency department (ED) utilization by adult Medicaid patients at an urban teaching hospital. METHODS: This was a retrospective cohort study using four years of ED records encompassing the year prior to initiation of MMC (1994-95), the enrollment year (1995-96), and two years after the program had matured (1996-98). RESULTS: Total ED census declined slightly, then returned to 1995 levels. Emergency department use by MMC patients declined steadily, with the 1998 figure of 5,888 representing a 40% decline over the pre-MMC volume of 9,849. Visits by MMC patients with acute illness or injury declined by 29%; MMC low-acuity visits decreased by 43%. Medicaid managed care low-acuity after-hours/weekend visits declined by 19%, then leveled off. The MMC enrollment was stable throughout the study period. CONCLUSIONS: Mandatory managed care can be associated with considerable diminution in ED use by Medicaid patients. This decline is most pronounced in low-acuity triage categories, and least evident after hours and on weekends. PMID- 11099434 TI - Public health preventive services, surveillance, and screening: the emergency Department's potential. PMID- 11099435 TI - Injury prevention, emergency medicine, and sports medicine on the same playing field. PMID- 11099436 TI - The rationale for developing public health surveillance systems based on emergency department data. AB - Emergency departments (EDs) are well positioned to provide national data on several aspects of public health. The large volume of patients seen annually, improving medical record technology, and emergency uniform data sets make the development of public health surveillance systems a realistic opportunity for emergency medicine. Such data could identify public health concerns and suggest interventions to improve the health of the nation. This article describes current concepts and status of ED surveillance systems, their advantages and disadvantages, the rationale for their existence, and recommendations to allow their continued consideration and development. PMID- 11099437 TI - Teaching patient empathy: the ED visit program. AB - OBJECTIVE: Residency programs only are not challenged with developing competent emergency clinicians, but should strive to develop caring, empathetic, and community-minded physicians. An exercise was designed to help residents experience emergency department (ED) visits from the patient's perspective. METHODS: This study occurred in emergency medicine residency program at an urban teaching institution with an annual ED census of 94,000. On the first day of residency orientation, each resident was given a clinical scenario and registered through triage into the ED. Nurses were blinded to the study. The study concluded when the examining physician entered the exam room. Residents were then presented with a simulated bill based on their scenario. Residents completed a survey initially and at six months. Survey ratings were measured using a 100-mm visual analog scale (VAS) (0 = not at all; 100 = a great deal). RESULTS: Twenty-five residents participated over two years. Sixty-four percent had never been an ED patient before. Median length of stay was 139 minutes. This exercise was found to improve resident empathy for patients on initial survey, 66 mm (range 16-71), and at follow-up, 66 mm (range 23-91). Residents found the exercise useful both initially, 50 mm (range 4-86), and at follow-up, 49 mm (range 15-81). Ninety-two percent of the residents thought the goals of the exercise had been met. Residents also stated the study changed their approach to patient care (45 mm, range 4-76) and made them a better physician (49 mm, range 5-80). CONCLUSIONS: The ED visit study enhanced patient empathy within residents and was useful in improving patient care attitude. PMID- 11099438 TI - Follow-up program for emergency department patients with gonorrhea or chlamydia. AB - OBJECTIVES: To study the performance of a centralized regional follow-up program organized by a municipal department of health (DH) for female patients presenting to the emergency department (ED) with Neisseria gonorrhoeae and/or Chlamydia trachomatis, who are not diagnosed or treated at the time of presentation. METHODS: This was a retrospective observational study of female patients seen in the ED with positive cervical specimens, and their subsequent treatment and follow-up by the DH. Medical records were reviewed to determine the female patients seen in the ED who had positive specimens for N. gonorrhoeae or C. trachomatis. The DH followed up those not treated in the ED. Analysis of how long it took for these patients to be treated and the proportion lost to follow-up was performed. RESULTS: Of 2,121 specimens, 342 were positive for N. gonorrhoeae or C. trachomatis. Of the 342, 154 (45%) were recognized and appropriately treated in the ED. One hundred fifty-nine of the 342 (46.5%) patients were discharged from the ED without treatment but were contacted by the DH and appropriate treatment was provided. The DH could not locate 23 (6.7%) patients, and four (1.2%) refused treatment. One died before treatment. Only 21 of the 159 were treated within nine days. Median time to treatment was 36 days. CONCLUSIONS: Centralized laboratory analysis and follow-up by the DH for N. gonorrhoeae and C. trachomatis identified many female patients undiagnosed and untreated in the ED. The DH follow-up program provided appropriate treatment to most female patients. PMID- 11099439 TI - Computer-assisted instruction. PMID- 11099440 TI - Reflections quotes on EM PMID- 11099441 TI - The new neuroanatomy of speech perception. PMID- 11099442 TI - The neurological basis of developmental dyslexia: an overview and working hypothesis. AB - Five to ten per cent of school-age children fail to learn to read in spite of normal intelligence, adequate environment and educational opportunities. Thus defined, developmental dyslexia (hereafter referred to as dyslexia) is usually considered of constitutional origin, but its actual mechanisms are still mysterious and currently remain the subject of intense research endeavour in various neuroscientific areas and along several theoretical frameworks. This article reviews evidence accumulated to date that favours a dysfunction of neural systems known to participate in the normal acquisition and achievement of reading and other related cognitive functions. Historically, the first arguments for a neurological basis of dyslexia came from neuropathological studies of brains from dyslexic individuals. These early studies, although open to criticism, for the first time drew attention towards a possible abnormality in specific stages of prenatal maturation of the cerebral cortex and suggested a role of atypical development of brain asymmetries. This has prompted a large amount of subsequent work using in vivo imaging methods in the same vein. These latter studies, however, have yielded less clear-cut results than expected, but have globally confirmed some subtle differences in brain anatomy whose exact significance is still under investigation. Neuropsychological studies have provided considerable evidence that the main mechanism leading to these children's learning difficulties is phonological in nature, namely a basic defect in segmenting and manipulating the phoneme constituents of speech. A case has also been made for impairment in brain visual mechanisms of reading as a possible contributing factor. This approach has led to an important conceptual advance with the suggestion of a specific involvement of one subsystem of vision pathways (the so called magnosystem hypothesis). Both phonological and visual hypotheses have received valuable contribution from modern functional imaging techniques. Results of recent PET and functional MRI studies are reported here in some detail. Finally, one attractive interpretation of available evidence points to dyslexia as a multi-system deficit possibly based on a fundamental incapacity of the brain in performing tasks requiring processing of brief stimuli in rapid temporal succession. It is proposed that this so-called 'temporal processing impairment' theory of dyslexia could also account for at least some of the perceptual, motor and cognitive symptoms very often associated with the learning disorder, a coincidence that has remained unexplained so far. PMID- 11099443 TI - Identification of a pathway for intelligible speech in the left temporal lobe. AB - It has been proposed that the identification of sounds, including species specific vocalizations, by primates depends on anterior projections from the primary auditory cortex, an auditory pathway analogous to the ventral route proposed for the visual identification of objects. We have identified a similar route in the human for understanding intelligible speech. Using PET imaging to identify separable neural subsystems within the human auditory cortex, we used a variety of speech and speech-like stimuli with equivalent acoustic complexity but varying intelligibility. We have demonstrated that the left superior temporal sulcus responds to the presence of phonetic information, but its anterior part only responds if the stimulus is also intelligible. This novel observation demonstrates a left anterior temporal pathway for speech comprehension. PMID- 11099444 TI - Acute disseminated encephalomyelitis, multiphasic disseminated encephalomyelitis and multiple sclerosis in children. AB - Forty-eight children with disseminated demyelination of the CNS, 28 with acute disseminated encephalomyelitis (ADEM), seven with multiphasic disseminated encephalomyelitis (MDEM) and 13 with multiple sclerosis were studied for a mean follow-up period of 5.64 years. The presentation findings of the ADEM/MDEM group were compared with those of the multiple sclerosis group. The following findings were more commonly seen in ADEM/MDEM presentation compared with the multiple sclerosis presentations: predemyelinating infectious disease (74 versus 38%, P: < 0.05); polysymptomatic presentation (91 versus 38%, P: < 0.002); pyramidal signs (71 versus 23%, P: < 0.01); encephalopathy (69 versus 15%, P: < 0.002); and bilateral optic neuritis (23 versus 8%, not significant). Seizures occurred only in the ADEM/MDEM group (17 versus 0%, not significant). Unilateral optic neuritis occurred only in the multiple sclerosis patients (23 versus 0%, P: < 0.01). There were no differences in the frequencies of transverse myelitis, brainstem involvement, cerebellar signs and sensory disturbance between the two groups. ADEM/MDEM patients were more likely to have blood leucocytosis (64 versus 22%, P: < 0.05), CSF lymphocytosis (64 versus 42%, not significant) and CSF protein elevation (60 versus 33%, not significant). Patients presenting with multiple sclerosis were more likely to have intrathecal synthesis of oligoclonal bands on presentation (64 versus 29%, not significant). MRI showed that subcortical white matter lesions were almost universal in both groups, though periventricular lesions were more common in multiple sclerosis (92 versus 44%, P: < 0.01). By contrast, in ADEM/MDEM there was absolute and relative periventricular sparing in 56 and 78% of patients, respectively. Follow-up MRI revealed complete or partial lesion resolution in 90% and no new lesions in the ADEM/MDEM group. All of the multiple sclerosis patients had new lesions on repeat MRI (five during relapse and six during asymptomatic convalescent phases). The outcome in the ADEM patients was mixed; 57% of patients made a complete recovery. The mean follow-up for the 35 ADEM/MDEM patients was 5.78 years (range 1.0-15.4 years). Eight of the 13 multiple sclerosis patients relapsed within the first year; 11 had a relapsing remitting course, one a primary progressive course and one a secondary progressive course. These differences in the presentation of ADEM/MDEM compared with multiple sclerosis may help in the prognosis given to families regarding the possibility of later development of multiple sclerosis. PMID- 11099445 TI - Brain iron deposition in Parkinson's disease imaged using the PRIME magnetic resonance sequence. AB - Iron content of the basal ganglia was investigated in 25 patients with idiopathic Parkinson's disease and 14 matched healthy control subjects using a partially refocused interleaved multiple echo sequence on a 1.5 Tesla MRI system. R(2)* (1/T(2)*) and R(2)' (1/T(2)') relaxation rates were higher in the substantia nigra of patients with Parkinson's disease, which indicates that iron content is elevated in this region. R(2)' was lower in the putamen, indicating reduced iron levels; reduction in this region was positively correlated with disease duration. Iron-related oxidative stress may contribute to the neurodegeneration of Parkinson's disease, which may lead to alterations in the iron levels of the striatum. We describe a simple, non-invasive technique for measuring iron content. PMID- 11099446 TI - The reorganization of sensorimotor function in children after hemispherectomy. A functional MRI and somatosensory evoked potential study. AB - Children who have suffered extensive unilateral brain injury early in life may show a remarkable degree of residual sensorimotor function. It is generally believed that this reflects the high capacity of the immature brain for cerebral reorganization. In this study, we investigated 17 patients who had undergone hemispherectomy for relief from seizures; eight of the patients had congenital brain damage and nine had sustained their initial insult at the age of 1 year or older. Sensorimotor functions of the hand were investigated using functional MRI (fMRI) during a passive movement task, somatosensory evoked potentials (SEPs) arising from electrical and vibration stimulation, and behavioural tests including grip strength, double simultaneous stimulation and joint position sense. On fMRI, two of the eight patients studied with this technique (one with congenital damage and one with damage acquired at the age of 3 years) showed activation in the sensorimotor cortex of the remaining hemisphere with passive movement of the hemiplegic hand. The location of the ipsilateral brain activation was similar to that found on movement of the normal contralateral hand, although the latter was greater in spatial extent. In one of these patients, a greater role was demonstrated for the ipsilateral secondary sensorimotor area (compared with the ipsilateral primary sensorimotor area) for movement of the hemiplegic hand than for movement of the normal hand. Median nerve stimulation of the hemiplegic hand showed reproducible early-latency ipsilateral SEP components in the remaining sensorimotor cortex in 10 of the 17 patients (five with congenital and five with acquired disease). Five of the patients who demonstrated ipsilateral electrical SEPs also showed ipsilateral vibration SEPs (two with congenital and three with acquired disease). The behavioural tests revealed residual sensorimotor function in 14 of the patients; however, not all of the patients who exhibited ipsilateral SEP or fMRI responses had residual sensorimotor function in the hemiplegic hand. Ipsilateral sensorimotor responses were demonstrated both in patients with congenital disease and those with acquired disease, suggesting that factors additional to aetiology and age at injury may influence the degree of residual sensorimotor function and cerebral reorganization. PMID- 11099447 TI - Failed surgery for epilepsy. A study of persistence and recurrence of seizures following temporal resection. AB - From a series of 282 consecutive temporal resections for medically intractable epilepsy associated with mesial temporal sclerosis (MTS), dysembryoplastic neuroepithelial tumour (DNT) or non-specific pathology (NSP), 51 patients had persistent or recurrent seizures occurring at least monthly. Of these patients, 44 underwent detailed assessment of their postoperative seizures, which included clinical evaluation, interictal and ictal EEG and high-resolution MRI. Of the 20 patients with MTS in the original pathology, 14 (70%) had postoperative seizures arising in the hemisphere of the resection, the majority (12 patients) in the temporal region. Although MRI demonstrated residual hippocampus in five of these 12 patients, only one patient was considered to have seizures arising there, whilst the remainder had electroclinical evidence of seizure onset in the neocortex. In contrast, five of the MTS relapses (25%) had seizure onset exclusively in the contralateral temporal region. Among the 14 patients with non specific pathology, relapse was also predominantly from the ipsilateral hemisphere (64%), but more relapsed from extratemporal sites compared with the MTS cases, including two with NSP who had occipital structural abnormalities. Although 70% of the 10 patients with DNT had postoperative partial seizures arising in the ipsilateral hemisphere, many (60%) had evidence of a more diffuse disorder with additional generalized seizures, cognitive and behavioural disturbance and multifocal and generalized EEG abnormalities. Nine patients (20%) had immediate postoperative seizure-free periods of at least 1 year, and seven of these had MTS in the operative specimen. Of these seven patients, four had ipsilateral temporal seizures and three had contralateral temporal seizures. Overall, few missed lesions were discovered on postoperative MRI and reoperations were performed or considered possible in a minority of cases. Despite well defined preoperative electroclinical syndromes of temporal lobe epilepsy, many patients relapsed unexpectedly, either immediately or remotely from the time of surgery. Maturing epileptogenicity in a surgical scar was not, however, considered to be a significant primary mechanism in patients who relapsed after a seizure-free interval. PMID- 11099448 TI - Pathology of early-onset Alzheimer's disease cases bearing the Thr113-114ins presenilin-1 mutation. AB - Most cases of familial presenile Alzheimer's disease are caused by mutations in the presenilin-1 (PSEN-1) gene, most of these mutations being missense mutations. A mutation in the splice donor site of intron 4 of PSEN-1 has been described recently which results in aberrant splicing of PSEN-1 mRNA, causing insertion of an additional amino acid, Thr113-114ins, into the protein. We studied the neuropathology of four cases bearing this mutation in an attempt to clarify the pathology of this hereditary form of Alzheimer's disease and to determine whether it differs from other familial forms of the disease. The disease presented as a progressive cognitive decline, myoclonus and seizures developing later in the disease, a feature common to PSEN-1-linked Alzheimer's disease. The course of the disease was relatively rapid, death occurring approximately 6 years after onset. Pathology in the intron 4 cases demonstrated a severe Alzheimer's disease pathology with abundant deposition of ss-amyloid (Ass) 1-42 senile plaques and the formation of neurofibrillary tangles. Amyloid angiopathy was present in these cases and was readily demonstrated by Ass 1-40 staining, particularly in the cerebellum. Cases with the intron 4 mutation appear clinically and pathologically similar to other cases of early-onset Alzheimer's disease bearing PSEN-1 mutations. PMID- 11099449 TI - Multimodal EEG analysis in man suggests impairment-specific changes in movement related electric brain activity after stroke. AB - Movement-related slow cortical potentials and event-related desynchronization of alpha (alpha-ERD) and beta (beta-ERD) activity after self-paced voluntary triangular finger movements were studied in 13 ischaemic supratentorial stroke patients and 10 age-matched control subjects during movement preparation and actual performance. The stroke patients suffered from central arm paresis (n = 8), somatosensory deficits (n = 3) or ideomotor apraxia (n = 2). The multimodal EEG analysis suggested impairment-specific changes in the movement-related electrical activity of the brain. The readiness potential of paretic subjects was centred more anteriorly and laterally; during movement, they showed increased beta-ERD at left lateral frontal recording sites. Patients with somatosensory deficits showed reduced alpha-ERD and beta-ERD during both movement preparation and actual performance. Patients with ideomotor apraxia showed more lateralized frontal movement-related slow cortical potentials during both movement preparation and performance, and reduced left parietal beta-ERD during movement preparation. We conclude that (i) disturbed motor efference is associated with an increased need for excitatory drive of pyramidal cells in motor and premotor areas or an attempt to drive movements through projections from these areas to brainstem motor systems during movement preparation; (ii) an undisturbed somatosensory afference might contribute to the release of relevant cortical areas from their 'idling' state when movements are prepared and performed; and (iii) apraxic patients have a relative lack of activity of the mesial frontal motor system and the left parietal cortex, which is believed to be part of a network subserving ideomotor praxis. PMID- 11099450 TI - Evidence of functional somatotopy in GPi from results of pallidotomy. AB - The objective of this study was to explore the functional anatomy of the globus pallidus internus (GPi) by studying the effects of unilateral pallidotomy on parkinsonian 'off' signs and levodopa-induced dyskinesias (LID). We found significant positive correlations between the preoperative levodopa responsiveness of motor signs and the levodopa responsiveness of scores in timed tests (Core Assessment Program for Intracerebral Transplantations) in the contralateral limbs and the improvement in these scores after surgery, whereas there was no correlation with the improvement in LID. We also found a highly significant correlation (P: < 0.0001, r = 0.8) between the volume of the ventral lesion in the GPi and the improvement in LID in the contralateral limbs, whereas there was no correlation between the ventral volume and the improvement in parkinsonian 'off' signs. The volumes of the total lesion cylinder and the dorsal lesion did not correlate with the outcome of either dyskinesias or parkinsonian 'off' signs. The differential predictive value of levodopa responsiveness for the outcome of parkinsonian 'off' signs and LID and the different correlations of ventral lesion volume with dyskinesias and parkinsonian 'off' signs indicate that different anatomical or pathophysiological substrates may be responsible for the generation of parkinsonian 'off' signs and dyskinesias. Whereas cells in a wider area of the GPi may be implicated in parkinsonism, the ventral GPi seems to be crucial for the manifestation of LID. We suggest that our observations are additional proof of the functional somatotopy of the systems within the GPi that mediate parkinsonism and dyskinesias, especially along the dorsoventral trajectory used in pallidotomy. The outcome of pallidotomy in which the lesion involves the ventral and dorsal GPi could be the net effect of alteration in the activity of pathways which mediate different symptoms, and hence could be variable. PMID- 11099451 TI - Anterior and posterior callosal contributions to simultaneous bimanual movements of the hands and fingers. AB - In order to study the role of the corpus callosum in two-handed coordination we tested callosotomy subjects while they attempted to initiate simultaneous discrete movements with both hands. We observed four split-brain patients, including one pre- and post-operatively, as well as normal and epileptic control subjects. Split-brain patients made button presses that were less synchronous than either normal or epileptic controls. Although split-brain patients' average performance did not always differ from control subjects, callosotomy resulted in a 3-fold increase in the variability with which 'simultaneous' movements were initiated. The one subject tested pre- and post-callosotomy showed distinct changes in movement initiation synchrony after both the anterior and the posterior stages of the surgery. These changes suggest that anterior and posterior callosal fibres may make unique contributions to bimanual synchronization, depending on whether responses are self-initiated or in reaction to a visual stimulus. This study demonstrates that neural communication across anterior and posterior fibres of the corpus callosum strongly influences the temporal precision of bimanual coordination. Specifically, callosal transmission affects the degree of bilateral synchrony with which simple simultaneous hand and finger movements are initiated. PMID- 11099452 TI - Handedness and hemispheric language dominance in healthy humans. AB - In most people the left hemisphere of the brain is dominant for language. Because of the increased incidence of atypical right-hemispheric language in left-handed neurological patients, a systematic association between handedness and dominance has long been suspected. To clarify the relationship between handedness and language dominance in healthy subjects, we measured lateralization directly by functional transcranial Doppler sonography in 326 healthy individuals using a word-generation task. The incidence of right-hemisphere language dominance was found to increase linearly with the degree of left-handedness, from 4% in strong right-handers (handedness = 100) to 15% in ambidextrous individuals and 27% in strong left-handers (handedness = -100). The relationship could be approximated by the formula: f1.gif" BORDER="0">. These results clearly demonstrate that the relationship between handedness and language dominance is not an artefact of cerebral pathology but a natural phenomenon. PMID- 11099453 TI - Distribution of ataxin-7 in normal human brain and retina. AB - Spinocerebellar ataxia 7 (SCA7) is a neurodegenerative disease caused by the expansion of a CAG repeat encoding a polyglutamine tract in the protein ataxin-7. We developed antibodies directed against two different parts of the ataxin-7 protein and studied its distribution in brain and peripheral tissue from healthy subjects. Normal ataxin-7 was widely expressed in brain, retina and peripheral tissues, including striated muscle, testis and thyroid gland. In the brain, expression of ataxin-7 was not limited to areas in which neurones degenerate, and the level of expression was not related to the severity of neuronal loss. Immunoreactivity was low in some vulnerable populations of neurones, such as Purkinje cells. In neurones, ataxin-7 was found in the cell bodies and in processes. Nuclear labelling was also observed in some neurones, but was not related to the distribution of intranuclear inclusions observed in an SCA7 patient. In this patient, the proportion of neurones with nuclear labelling was higher, on average, in regions with neuronal loss. Double immunolabelling coupled with confocal microscopy showed that ataxin-7 colocalized with BiP, a marker of the endoplasmic reticulum, but not with markers of mitochondria or the trans Golgi network. PMID- 11099454 TI - Neural consequences of acting in near versus far space: a physiological basis for clinical dissociations. AB - We used PET to determine which brain regions are implicated when normal volunteers bisect horizontal lines and point to dots in near (peripersonal) or far (extrapersonal) space. Studies of line bisection in patients with right hemisphere lesions have shown that bisection performance can be severely impaired in either near or far space while remaining within normal limits in the other spatial domain. Likewise, clinical dissociations between pointing to objects in near and far space have been reported. The normal functional anatomy of these dissociations has not been demonstrated convincingly. Regional cerebral blood flow measurements using PET were carried out in 12 healthy right-handed male volunteers who bisected lines or pointed to dots in near or far space, using a laser pen. Subjects performing either task in near space showed neural activity in the left dorsal occipital cortex, left intraparietal cortex, left ventral premotor cortex and left thalamus. In far space, subjects performing either task showed activation of the ventral occipital cortex bilaterally and the right medial temporal cortex. These data provide physiological support for the clinically observed dissociations demonstrating that attending to and acting in near space differentially employs dorsal visuomotor processing areas, whereas attending to and acting in far space differentially draws on ventral visuoperceptual processing areas, even when the motor components of the tasks are identical when performed in the two spaces. PMID- 11099455 TI - Effects of membrane polarization and ischaemia on the excitability properties of human motor axons. AB - Multiple nerve excitability measurements have been proposed for clinical testing of nerve function, since excitability measures can provide evidence of altered axonal membrane properties and are complementary to conventional nerve conduction studies. An important determinant of excitability is membrane potential, and this study was undertaken to determine the changes in a range of excitability properties associated with alterations in membrane potential. Membrane potential was varied directly using DC polarizing currents and indirectly by ischaemia. The median nerve was stimulated at the wrist and the resultant compound muscle action potentials recorded from abductor pollicis brevis. Stimulus-response behaviour, strength-duration time constant (tau(SD)), threshold electrotonus to 100-ms polarizing currents, a current-threshold relationship and the recovery of excitability following supramaximal activation were each followed in four normal subjects during the two manoeuvres, using a recently described protocol. Membrane depolarization and ischaemia produced an increase in axonal excitability, an increase in the slope of the current-threshold relationship, a 'fanning in' of responses during threshold electrotonus, a decrease in super-excitability, and increases in both tau(SD) and the refractory period. Changes in the opposite direction occurred with membrane hyperpolarization and during the post-ischaemic period. One excitability parameter differentiated between the direct and indirect changes in membrane potential: late subexcitability was sensitive to polarizing currents but relatively insensitive to ischaemia, probably because of compensatory changes in extracellular potassium ions. These results should enable multiple excitability measurements to be used as a tool to identify changes in axonal membrane potential in neuropathy. PMID- 11099456 TI - Brain responses to nouns, verbs and class-ambiguous words in context. AB - Recent neuropsychological and imaging data have implicated different brain networks in the processing of different word classes, nouns being linked primarily to posterior, visual object-processing regions and verbs to frontal, motor-processing areas. However, as most of these studies have examined words in isolation, the consequences of such anatomically based representational differences, if any, for the processing of these items in sentences remains unclear. Additionally, in some languages many words (e.g. 'drink') are class ambiguous, i.e. they can play either role depending on context, and it is not yet known how the brain stores and uses information associated with such lexical items in context. We examined these issues by recording event-related potentials (ERPs) in response to unambiguous nouns (e.g. 'beer'), unambiguous verbs (e. g. 'eat'), class-ambiguous words and pseudowords used as nouns or verbs within two types of minimally contrastive sentence contexts: noun-predicting (e.g. 'John wanted THE [target] but.') and verb-predicting ('John wanted TO [target] but.'). Our results indicate that the nature of neural processing for nouns and verbs is a function of both the type of stimulus and the role it is playing. Even when the context completely specifies their role, word class-ambiguous items differ from unambiguous ones over frontal regions by approximately 150 ms. Moreover, whereas pseudowords elicit larger N400s when used as verbs than when used as nouns, unambiguous nouns and ambiguous words used as nouns elicit more frontocentral negativity than unambiguous verbs and ambiguous words used as verbs, respectively. Additionally, unambiguous verbs elicit a left-lateralized, anterior positivity (approximately 200 ms) not observed for any other stimulus type, though only when these items are used appropriately as verbs (i.e. in verb predicting contexts). In summary, the pattern of neural activity observed in response to lexical items depends on their general probability of being a verb or a noun and on the particular role they are playing in any given sentence. This implicates more than a simple two-way distinction of the brain networks involved in their storage and processing. Experience, as well as context during on-line language processing, clearly shapes the neural representations of nouns and verbs, such that there is no single neural marker of word class. Our results further suggest that the presence and nature of the word class-based dissociations observed after brain damage are similarly likely to be a function of both the type of stimulus and the context in which it occurs, and thus must be assessed accordingly. PMID- 11099457 TI - The vesicular neurotransmitter transporters: current perspectives and future prospects. PMID- 11099458 TI - In vivo imaging of the vesicular acetylcholine transporter and the vesicular monoamine transporter. AB - Validation of the vesicular acetylcholine transporter (VAChT) and the neuronal vesicular monoamine transporter (VMAT2) as important molecular targets in the cholinergic and dopamine neurons, respectively, has sparked interest in the development of radiotracers for studying these markers in vitro and in vivo. Currently, a number of selective high-affinity radiotracers are available for studying these targets in vivo with positron emission tomography (PET) or single photon emission computed tomography (SPECT). PET studies of VMAT2 in neuropathology reveal changes in the density of this marker that can be verified independently. Similarly, in vivo studies with VAChT ligands suggest that the latter are potentially useful in detecting cholinergic lesions in vivo; however, additional development is required to fully realize the potential of these radioligands. PMID- 11099459 TI - Neurogenetics of vesicular transporters in C. elegans. AB - The nematode Caenorhabditis elegans has a number of advantages for the analysis of synaptic molecules. These include a simple nervous system in which all cells are identified and synaptic connectivity is known and reproducible, a large collection of mutants and powerful methods of genetic analysis, simple methods for the generation and analysis of transgenic animals, and a number of relatively simple quantifiable behaviors. Studies in C. elegans have made major contributions to our understanding of vesicular transmitter transporters. Two of the four classes of vesicular transporters so far identified (VAChT and VGAT) were first described and cloned in C. elegans; in both cases, the genes were first identified and cloned by means of mutations causing a suggestive phenotype (1, 2). The phenotypes of eat-4 mutants and the cell biology of the EAT-4 protein were critical in the identification of this protein as the vesicular glutamate transporter (3, 4). In addition, the unusual gene structure associated with the cholinergic locus was first described in C. elegans (5). The biochemical properties of the nematode transporters are surprisingly similar to their vertebrate counterparts, and they can be assayed under similar conditions using the same types of mammalian cells (6, 7). In addition, mild and severe mutants (including knockouts) are available for each of the four C. elegans vesicular transporters, which has permitted a careful evaluation of the role(s) of vesicular transport in transmitter-specific behaviors. Accordingly, it seems appropriate at this time to present the current status of the field. In this review, we will first discuss the properties of C. elegans vesicular transporters and transporter mutants, and then explore some of the lessons and insights C. elegans research has provided to the field of vesicular transport. PMID- 11099460 TI - Transport mechanisms in acetylcholine and monoamine storage. AB - Sequence-related vesicular acetylcholine transporter (VAChT) and vesicular monoamine transporter (VMAT) transport neurotransmitter substrates into secretory vesicles. This review seeks to identify shared and differentiated aspects of the transport mechanisms. VAChT and VMAT exchange two protons per substrate molecule with very similar initial velocity kinetics and pH dependencies. However, vesicular gradients of ACh in vivo are much smaller than the driving force for uptake and vesicular gradients of monoamines, suggesting the existence of a regulatory mechanism in ACh storage not found in monoamine storage. The importance of microscopic rather than macroscopic kinetics in structure-function analysis is described. Transporter regions affecting binding or translocation of substrates, inhibitors, and protons have been found with photoaffinity labeling, chimeras, and single-site mutations. VAChT and VMAT exhibit partial structural and mechanistic homology with lactose permease, which belongs to the same sequence-defined superfamily, despite opposite directions of substrate transport. The vesicular transporters translocate the first proton using homologous aspartates in putative transmembrane domain X (ten), but they translocate the second proton using unknown residues that might not be conserved between them. Comparative analysis of the VAChT and VMAT transport mechanisms will aid understanding of regulation in neurotransmitter storage. PMID- 11099461 TI - Chemical neuroanatomy of the vesicular amine transporters. AB - Acetylcholine, catecholamines, serotonin, and histamine are classical neurotransmitters. These small molecules also play important roles in the endocrine and immune/inflammatory systems. Serotonin secreted from enterochromaffin cells of the gut epithelium regulates gut motility; histamine secreted from basophils and mast cells is a major regulator of vascular permeability and skin inflammatory responses; epinephrine is a classical hormone released from the adrenal medulla. Each of these molecules is released from neural, endocrine, or immune/inflammatory cells only in response to specific physiological stimuli. Regulated secretion is possible because amines are stored in secretory vesicles and released via a stimulus-dependent exocytotic event. Amine storage-at concentrations orders of magnitude higher than in the cytoplasm is accomplished in turn by specific secretory vesicle transporters that recognize the amines and move them from the cytosol into the vesicle. Immunohistochemical visualization of specific vesicular amine transporters (VATs) in neuronal, endocrine, and inflammatory cells provides important new information about how amine-handling cell phenotypes arise during development and how vesicular transport is regulated during homeostatic response events. Comparison of the chemical neuroanatomy of VATs and amine biosynthetic enzymes has also revealed cell groups that express vesicular transporters but not enzymes for monoamine synthesis, and vice versa: their function and regulation is a new topic of investigation in mammalian neurobiology. The chemical neuroanatomy of the vesicular amine transporters is reviewed here. These and similar data emerging from the study of the localization of the recently characterized vesicular inhibitory and excitatory amino acid transporters will contribute to understanding chemically coded synaptic circuitry in the brain, and amine handling neuroendocrine and immune/inflammatory cell regulation. PMID- 11099462 TI - Molecular analysis of vesicular amine transporter function and targeting to secretory organelles. AB - Vesicular transporters are responsible for the loading of neurotransmitters into specialized secretory organelles in neurons and neuroendocrine cells to make them available for regulated neurosecretion. The exocytotic release of neurotransmitter therefore depends on the functional activity of the vesicular transporters and their efficient sorting to these secretory organelles. Molecular analysis of vesicular transport proteins has revealed important information regarding structural domains responsible for their functional properties, including substrate specificity and trafficking to various classes of secretory vesicles. These studies have established the existence of an important functional relationship between transporter activity and presynaptic quantal neurosecretion. PMID- 11099463 TI - The VMAT2 gene in mice and humans: amphetamine responses, locomotion, cardiac arrhythmias, aging, and vulnerability to dopaminergic toxins. AB - Monoamine compartmentalization in monoaminergic neurons uses serial action of the plasma membrane and vesicular monoamine (VAMT2) transporters. We can now define the sequences of the genes encoding these transporters in mice and humans, examine influences of deletions of this gene and alteration in its expression levels in transgenic mice, and identify sequence polymorphisms in the human VMAT2 gene. Examination of VMAT2 variants can provide potential insights into roles for allelic variants at these loci in variant drug responses and in diseases linked to monoaminergic systems, including substance abuse and Parkinson's disease. PMID- 11099464 TI - The P2 purinergic receptors of human dendritic cells: identification and coupling to cytokine release. AB - We investigated the expression of purinoceptors in human dendritic cells, providing functional, pharmacological, and biochemical evidence that immature and mature cells express P2Y and P2X subtypes, coupled to increase in the intracellular Ca(2+), membrane depolarization, and secretion of inflammatory cytokines. The ATP-activated Ca(2+) change was biphasic, with a fast release from intracellular stores and a delayed influx across the plasma membrane. A prolonged exposure to ATP was toxic to dendritic cells that swelled, lost typical dendrites, became phase lucent, detached from the substrate, and eventually died. These changes were highly suggestive of expression of the cytotoxic receptor P2X(7), as confirmed by ability of dendritic cells to become permeant to membrane impermeant dyes such as Lucifer yellow or ethidium bromide. The P2X(7) receptor ligand 2',3'-(4-benzoylbenzoyl)-ATP was a better agonist then ATP for Ca(2+) increase and plasma membrane depolarization. Oxidized ATP, a covalent blocker of P2X receptors, and the selective P2X(7) antagonist KN-62 inhibited both permeabilization and Ca(2+) changes induced by ATP. The following purinoceptors were expressed by immature and mature dendritic cells: P2Y(1), P2Y(2), P2Y(5), P2Y(11) and P2X(1), P2X(4), P2X(7). Finally, stimulation of LPS-matured cells with ATP triggered release of IL-1 beta and TNF-alpha. Purinoceptors may provide a new avenue to modulation of dendritic cells function. PMID- 11099465 TI - Halofuginone: a potent inhibitor of critical steps in angiogenesis progression. AB - We have previously demonstrated that halofuginone, a low molecular weight quinazolinone alkaloid, is a potent inhibitor of collagen alpha1(I) and matrix metalloproteinase 2 (MMP-2) gene expression. Halofuginone also effectively suppresses tumor progression and metastasis in mice. These results together with the well-documented role of extracellular matrix (ECM) components and matrix degrading enzymes in formation of new blood vessels led us to investigate the effect of halofuginone on the angiogenic process. In a variety of experimental system, representing sequential events in the angiogenic cascade, halofuginone treatment resulted in profound inhibitory effect. Among these are the abrogation of endothelial cell MMP-2 expression and basement membrane invasion, capillary tube formation, and vascular sprouting, as well as deposition of subendothelial ECM. The most conclusive anti-angiogenic activity of halofuginone was demonstrated in vivo (mouse corneal micropocket assay) by showing a marked inhibition of basic fibroblast growth factor (bFGF) -induced neovascularization in response to systemic administration of halofuginone, either i.p. or in the diet. The ability of halofuginone to interfere with key events in neovascularization, together with its oral bioavailability and safe use as an anti-parasitic agent, make it a promising drug for further evaluation in the treatment of a wide range of diseases associated with pathological angiogenesis. PMID- 11099466 TI - High-efficiency transient transfection of endothelial cells for functional analysis. AB - The definition of signaling pathways in endothelial cells has been hampered by the difficulty of transiently transfecting these cells with high efficiency. This investigation was undertaken to develop an efficient technique for the transfection of endothelial cells for functional analyses. Cells cotransfected with plasmid expressing green fluorescent protein (GFP) and the plasmid of interest were isolated by fluorescence-activated cell sorting (FACS) based on GFP expression. In the sorted cell population, a 2.5-fold enhancement in the number of cells expressing the gene of interest was observed, as confirmed by FACS analysis and Western blotting. Sorted cells retained functional properties, as demonstrated by chemotaxis to the agonist sphingosine 1-phosphate (SPP). To demonstrate the usefulness of this method for defining cellular signaling pathways, cells were cotransfected with plasmids encoding GFP and the carboxyl terminal domain of the beta-adrenergic receptor kinase (beta ARKct), which inhibits signaling through the beta gamma dimer of heterotrimeric G-proteins. SPP induced chemotaxis in sorted cells coexpressing beta ARKct was inhibited by 80%, demonstrating that chemotaxis was driven by a beta gamma-dependent pathway. However, no significant inhibition was observed in cells transfected with betaARKct but not enriched by sorting. Thus, we have developed a method for enriching transfected cells that allows the elucidation of crucial mechanisms of endothelial cell activation and function. This method should find wide applicability in studies designed to define pathways responsible for regulation of motility and other functions in these dynamic cells. PMID- 11099467 TI - Protein oxidation and degradation during cellular senescence of human BJ fibroblasts: part I--effects of proliferative senescence. AB - Oxidized and cross-linked proteins tend to accumulate in aging cells. Declining activity of proteolytic enzymes, particularly the proteasome, has been proposed as a possible explanation for this phenomenon, and direct inhibition of the proteasome by oxidized and cross-linked proteins has been demonstrated in vitro. We have further examined this hypothesis during both proliferative senescence (this paper) and postmitotic senescence (see the accompanying paper, ref 1 ) of human BJ fibroblasts. During proliferative senescence, we found a marked decline in all proteasome activities (trypsin-like activity, chymotrypsin-like activity, and peptidyl-glutamyl-hydrolyzing activity) and in lysosomal cathepsin activity. Despite the loss of proteasome activity, there was no concomitant change in cellular levels of actual proteasome protein (immunoassays) or in the steady state levels of mRNAs for essential proteasome subunits. The decline in proteasome activities and lysosomal cathepsin activities was accompanied by dramatic increases in the accumulation of oxidized and cross-linked proteins. Furthermore, as proliferation stage increased, cells exhibited a decreasing ability to degrade the oxidatively damaged proteins generated by an acute, experimentally applied oxidative stress. Thus, oxidized and cross-linked proteins accumulated rapidly in cells of higher proliferation stages. Our data are consistent with the hypothesis that proteasome is progressively inhibited by small accumulations of oxidized and cross-linked proteins during proliferative senescence until late proliferation stages, when so much proteasome activity has been lost that oxidized proteins accumulate at ever-increasing rates. Lysosomes attempt to deal with the accumulating oxidized and cross-linked proteins, but declining lysosomal cathepsin activity apparently limits their effectiveness. This hypothesis, which may explain the progressive intracellular accumulation of oxidized and cross-linked proteins in aging, is further explored during postmitotic senescence in the accompanying paper (1). PMID- 11099468 TI - Protein oxidation and degradation during cellular senescence of human BJ fibroblasts: part II--aging of nondividing cells. AB - Oxidized/cross-linked intracellular protein materials, known as ceroid pigment, age pigment, or lipofuscin, accumulate in postmitotic tissues. It is unclear, however, whether diminishing proteolytic capacities play a role in the accumulation of such oxidized intracellular proteins. Previous studies revealed that the proteasome is responsible for the degradation of most oxidized soluble cytoplasmic and nuclear proteins and, we propose, for the prevention of such damage accumulations. The present investigation was undertaken to test the changes in protein turnover, proteasome activity, lysosome activity, and protein oxidation status during the aging of nondividing cells. Since the companion paper shows that both proteasome activity and the overall protein turnover decline during proliferative senescence whereas the accumulation of oxidized proteins increases significantly, we decided to use the same human BJ fibroblasts, this time at confluency, at different PD levels (including those that are essentially postmitotic) to investigate the same parameters under conditions where cells do not divide. We find that the activity of the cytosolic proteasome declines dramatically during senescence of nondividing BJ fibroblasts. The peptidyl glutamyl-hydrolyzing activity was particularly affected. This decline in proteasome activity was accompanied by a decrease in the overall turnover of short-lived (radiolabeled) proteins in the nondividing BJ fibroblasts. On the other hand, no decrease in the actual cellular proteasome content, as judged by immunoblots, was found. The decline in the activity of the proteasome was also accompanied by an increased accumulation of oxidized proteins, especially of oxidized and cross-linked material. Unlike the loss of lysosomal function seen in our accompanying studies of proliferative senescence (1), however, the present study of hyperoxic senescence in nondividing cells actually revealed marked increases in lysosomal cathepsin activity in all but the very 'oldest' postmitotic cells. Our comparative studies of proliferating (1) and nonproliferating (this paper) human BJ fibroblasts reveal a good correlation between the accumulation of oxidized/cross-linked proteins and the decline in proteasome activity and overall cellular protein turnover during in vitro senescence, which may predict a causal relationship during actual cellular aging. PMID- 11099469 TI - S-adenosylmethionine regulates MAT1A and MAT2A gene expression in cultured rat hepatocytes: a new role for S-adenosylmethionine in the maintenance of the differentiated status of the liver. AB - Methionine metabolism starts with the formation of S-adenosylmethionine (AdoMet), the most important biological methyl donor. This reaction is catalyzed by methionine adenosyltransferase (MAT). MAT is the product of two different genes: MAT1A, which is expressed only in the adult liver, and MAT2A, which is widely distributed, expressed in the fetal liver, and replaces MAT1A in hepatocarcinoma. In the liver, preservation of high expression of MAT1A and low expression of MAT2A is critical for the maintenance of a functional and differentiated organ. Here we describe that in cultured rat hepatocytes MAT1A expression progressively decreased, as described for other liver-specific genes, and MAT2A expression was induced. We find that this switch in gene expression was prevented by adding AdoMet to the culture medium. We also show that in cultured hepatocytes with decreased MAT1A expression AdoMet addition markedly increased MAT1A transcription in a dose-dependent fashion. This effect of AdoMet was mimicked by methionine, and blocked by 3-deazaadenosine and L-ethionine, but not D-ethionine, indicating that the effect was specific and mediated probably by a methylation reaction. These findings identify AdoMet as a key molecule that differentially regulates MAT1A and MAT2A expression and helps to maintain the differentiated status of the hepatocyte. PMID- 11099470 TI - Mechanism of the antimicrobial drug trimethoprim revisited. AB - We tested the hypothesis that the mechanism of action of the antifolate drug trimethoprim is through accumulation of bacterial dihydrofolate resulting in depletion of tetrahydrofolate coenzymes required for purine and pyrimidine biosynthesis. The folate pool of a strain of Escherichia coli (NCIMB 8879) was prelabeled with the folate biosynthetic precursor [(3)H]-p-aminobenzoic acid before treatment with trimethoprim. Folates in untreated E. coli were present as tetrahydrofolate coenzymes. In trimethoprim-treated cells, however, a rapid transient accumulation of dihydrofolate occurred, followed by complete conversion of all forms of folate to cleaved catabolites (pteridines and para aminobenzoylglutamate) and the stable nonreduced form of the vitamin, folic acid. Both para-aminobenzoylglutamate and folic acid were present in the cell in the form of polyglutamates. Removal of trimethoprim resulted in the reconversion of the accumulated folic acid to tetrahydrofolate cofactors for subsequent participation in the one-carbon cycle. Whereas irreversible catabolism is probably bactericidal, conversion to folic acid may constitute a bacteriostatic mechanism since, as we show, folic acid can be used by the bacteria and proliferation is resumed once trimethoprim is removed. Thus, the clinical effectiveness of this important drug may depend on the extent to which the processes of either catabolism or folic acid production occur in different bacteria or during different therapeutic regimes. PMID- 11099471 TI - Impaired cutaneous wound healing in interleukin-6-deficient and immunosuppressed mice. AB - It has been postulated that an inflammatory response after cutaneous wounding is a prerequisite for healing, and inflammatory cytokines, such as interleukin-6 (IL 6), might be intimately involved in this process. IL-6-deficient transgenic mice (IL-6 KO) displayed significantly delayed cutaneous wound healing compared with wild-type control animals, requiring up to threefold longer to heal. This was characterized by minimal epithelial bridge formation, decreased inflammation, and granulation tissue formation. Using electrophoretic mobility shift assays of wound tissue from IL-6 KO mice, decreased AP-1 transcription factor activation was shown compared with wild-type mice 16 h after wounding. In situ hybridization of wound tissue from wild-type mice revealed IL-6 mRNA expression primarily in the epidermis at the leading edge of the wound. Delayed wound healing in IL-6 KO mice was reversed with a single dose of recombinant murine IL-6 or intradermal injection of an expression plasmid containing the full-length murine IL-6 cDNA. Treatment with rmIL-6 also reconstituted wound healing in dexamethasone-treated immunosuppressed mice. The results of this study may indicate a potential use for IL-6 therapeutically where cutaneous wound healing is impaired. PMID- 11099472 TI - Neuropilin-1 expression by tumor cells promotes tumor angiogenesis and progression. AB - Neuropilin-1 (NRP1) is a VEGF(165) and semaphorin receptor expressed by vascular endothelial cells (EC) and tumor cells. The function of NRP1 in tumor cells is unknown. NRP1 was overexpressed in Dunning rat prostate carcinoma AT2.1 cells using a tetracycline-inducible promoter. Concomitant with increased NRP1 expression in response to a tetracycline homologue, doxycycline (Dox), basal cell motility, and VEGF(165) binding were increased three- to fourfold in vitro. However, induction of NRP1 did not affect tumor cell proliferation. When rats injected with AT2.1/NRP1 tumor cells were fed Dox, NRP1 synthesis was induced in vivo and AT2.1 cell tumor size was increased 2.5- to 7-fold in a 3-4 wk period compared to controls. The larger tumors with induced NRP1 expression were characterized by markedly increased microvessel density, increased proliferating EC, dilated blood vessels, and notably less tumor cell apoptosis compared to noninduced controls. It was concluded that NRP1 expression results in enlarged tumors associated with substantially enhanced tumor angiogenesis. PMID- 11099473 TI - Selection of ventricular-like cardiomyocytes from ES cells in vitro. AB - Ischemic disorders of the heart can cause an irreversible loss of cardiomyocytes resulting in a substantial decrease of cardiac output. The therapy of choice is heart transplantation, a technique that is hampered by the low number of donor organs. In the present study, we describe the specific labeling, rapid but gentle purification and characterization of cardiomyocytes derived from mouse pluripotent embryonic stem (ES) cells. To isolate the subpopulation of ventricular-like cardiomyocytes, ES cells were stable transfected with the enhanced green fluorescent protein (EGFP) under transcriptional control of the ventricular-specific 2.1 kb myosin light chain-2v (MLC-2v) promoter and the 0.5 kb enhancer element of the cytomegalovirus (CMV(enh).). First fluorescent cells were detected at day 6 + 8 of differentiation within EBs. Four weeks after initiation of differentiation 25% of the cardiomyocyte population displayed fluorescence. Immunohistochemistry revealed the exclusive cardiomyogenic nature of EGFP-positive cells. This was further corroborated by electrophysiological studies where preferentially ventricular phenotypes, but no pacemaker-like cardiomyocytes, were detected among the EGFP-positive population. The enzymatic digestion of EBs, followed by Percoll gradient centrifugation and fluorescence activated cell sorting, resulted in a 97% pure population of cardiomyocytes. Based on this study, ventricular-like cardiomyocytes can be generated in vitro from EBs and labeled using CMV(enh)./MLC-2v-driven marker genes facilitating an efficient purification. This method may become an important tool for future cell replacement therapy of ischemic cardiomyopathy especially after the proof of somatic differentiation of human ES cells in vitro. PMID- 11099474 TI - RACK1 is up-regulated in angiogenesis and human carcinomas. AB - Angiogenesis is crucial for many biological and pathological processes including the ovarian cycle and tumor growth. To identify molecules relevant for angiogenesis, we performed mRNA fingerprinting and subsequent Northern blot analysis using bovine cord-forming vs. monolayer-forming endothelial cells (EC) in vitro and staged bovine corpora lutea in vivo. We detected the receptor for activated C kinase 1 (RACK1), the specific receptor for activated protein kinase C beta (PKC beta), to be up-regulated in bovine cord-forming EC in vitro and in angiogenically active stages of bovine corpora lutea in vivo. Thereafter we established and determined the complete bovine RACK1 cDNA sequence. RACK1 was massively induced in subconfluent vs. contact-inhibited bovine EC, during angiogenesis in vitro, active phases of the murine ovarian cycle, human tumor angiogenesis, and in cancer cells in vivo as assessed by quantitative PCR and in situ hybridization. RACK1 transcripts were localized to proliferating EC in vitro and the endothelium of tumor neovascularizations in vivo by in situ hybridization. PKC beta plays an important role in angiogenesis and cancer growth. Our data suggest that downstream signaling of PKC beta in angiogenically active vs. inactive tissues and endothelium is affected by the availability of RACK1. PMID- 11099475 TI - Functional characterization of MT3-MMP in transfected MDCK cells: progelatinase A activation and tubulogenesis in 3-D collagen lattice. AB - MT3-MMP, a membrane-anchored matrix metalloproteinase, has been proposed to participate in the remodeling of extracellular matrix either by direct proteolysis or via activating other enzymes such as progelatinase A. To test this hypothesis, we analyzed the effect of exogenously transfected MT3-MMP in a tissue remodeling system: growth and tubulogenesis of Madin-Darby canine kidney (MDCK) cells in collagen gels. Although the parental cells require MMP activities for both growth and tubulogenesis, over-expression of wild-type MT3-MMP, but not its catalytically inactive mutant, leads to further enhancement of both processes, independent of its downstream substrate, progelatinase A. Mechanistically, MT3 MMP accomplishes such an effect by displaying on cell surfaces as active species, ready to activate progelatinase A or degrade ECM molecules. These data strongly suggest that MT3-MMP possesses the potential to directly enhance the growth and invasiveness of cells in vivo, two critical processes for development and carcinogenesis. PMID- 11099476 TI - The G-protein-coupled receptor kinase GRK4 mediates homologous desensitization of metabotropic glutamate receptor 1. AB - G-protein-coupled receptor kinases (GRKs) are involved in the regulation of many G-protein-coupled receptors. As opposed to the other GRKs, such as rhodopsin kinase (GRK1) or beta-adrenergic receptor kinase (beta ARK, GRK2), no receptor substrate for GRK4 has been so far identified. Here we show that GRK4 is expressed in cerebellar Purkinje cells, where it regulates mGlu(1) metabotropic glutamate receptors, as indicated by the following: 1) When coexpressed in heterologous cells (HEK293), mGlu(1) receptor signaling was desensitized by GRK4 in an agonist-dependent manner (homologous desensitization). 2) In transfected HEK293 and in cultured Purkinje cells, the exposure to glutamate agonists induced internalization of the receptor and redistribution of GRK4. There was a substantial colocalization of the receptor and kinase both under basal condition and after internalization. 3) Kinase activity was necessary for desensitizing mGlu(1a) receptor and agonist-dependent phosphorylation of this receptor was also documented. 4) Antisense treatment of cultured Purkinje cells, which significantly reduced the levels of GRK4 expression, induced a marked modification of the mGlu(1)-mediated functional response, consistent with an impaired receptor desensitization. The critical role for GRK4 in regulating mGlu(1) receptors implicates a major involvement of this kinase in the physiology of Purkinje cell and in motor learning. PMID- 11099477 TI - Quantitation of signal transduction. AB - Conventional qualitative approaches to signal transduction provide powerful ways to explore the architecture and function of signaling pathways. However, at the level of the complete system, they do not fully depict the interactions between signaling and metabolic pathways and fail to give a manageable overview of the complexity that is often a feature of cellular signal transduction. Here, we introduce a quantitative experimental approach to signal transduction that helps to overcome these difficulties. We present a quantitative analysis of signal transduction during early mitogen stimulation of lymphocytes, with steady-state respiration rate as a convenient marker of metabolic stimulation. First, by inhibiting various key signaling pathways, we measure their relative importance in regulating respiration. About 80% of the input signal is conveyed via identifiable routes: 50% through pathways sensitive to inhibitors of protein kinase C and MAP kinase and 30% through pathways sensitive to an inhibitor of calcineurin. Second, we quantify how each of these pathways differentially stimulates functional units of reactions that produce and consume a key intermediate in respiration: the mitochondrial membrane potential. Both the PKC and calcineurin routes stimulate consumption more strongly than production, whereas the unidentified signaling routes stimulate production more than consumption, leading to no change in membrane potential despite increased respiration rate. The approach allows a quantitative description of the relative importance of signal transduction pathways and the routes by which they activate a specific cellular process. It should be widely applicable. PMID- 11099478 TI - Two-chain high molecular weight kininogen induces endothelial cell apoptosis and inhibits angiogenesis: partial activity within domain 5. AB - We previously reported that the binding of two-chain high molecular weight kininogen (HKa) to endothelial cells may occur through interactions with endothelial urokinase receptors. Since the binding of urokinase to urokinase receptors activates signaling responses and may stimulate mitogenesis, we assessed the effect of HKa binding on endothelial cell proliferation. Unexpectedly, HKa inhibited proliferation in response to several growth factors, with 50% inhibition caused by approximately 10 nM HKa. This activity was Zn(2+) dependent and not shared by either single-chain high molecular weight kininogen (HK) or low molecular weight kininogen. HKa selectively inhibited the proliferation of human umbilical vein and dermal microvascular endothelial cells, but did not affect that of umbilical vein or human aortic smooth muscle cells, trophoblasts, fibroblasts, or carcinoma cells. Inhibition of endothelial proliferation by HKa was associated with endothelial cell apoptosis and unaffected by antibodies that block the binding of HK or HKa to any of their known endothelial receptors. Recombinant HK domain 5 displayed activity similar to that of HKa. In vivo, HKa inhibited neovascularization of subcutaneously implanted Matrigel plugs, as well as rat corneal angiogenesis. These results demonstrate that HKa is a novel inhibitor of angiogenesis, whose activity is dependent on the unique conformation of the two-chain molecule. PMID- 11099479 TI - Glucose- and arginine-induced insulin secretion by human pancreatic beta-cells: the role of HERG K(+) channels in firing and release. AB - The human ether-a-go-go-related genes (herg) are expressed in tissues other than heart and brain where the HERG K(+) channels are known to regulate the repolarization of the heart action potential and the neuronal spike-frequency accommodation. We provide evidence that herg1 transcripts are present in human pancreatic islets that were used to study both insulin secretion and electrical activity with radioimmunoassay and single cell perforated patch-clamp techniques, respectively. Glucose- and arginine-induced islets insulin secretion data suggested a net increase of release under perfusion with antiarrhythmic drugs known to selectively block HERG channels. Indeed we could routinely isolate a K(+) current that was recognized as biophysically and pharmacologically similar to the HERG current. An analysis of the glucose- and arginine-induced electrical activity (several applications during 30 min) in terms of firing frequency and putative insulin release was done in control and in the presence of selective blockers of HERG channels: the firing frequency and the release increased by 32% and 77%, respectively. It is concluded that HERG channels have a crucial role in regulating insulin secretion and firing of human beta-cells. This raises the possibility that some genetically characterized hyperinsulinemic diseases of unknown origin might involve mutations in the HERG channels. PMID- 11099480 TI - Impaired beta-cell regeneration in perinatally malnourished rats: a study with STZ. AB - We investigated the mechanisms implicated in beta-cell mass reduction observed during late fetal and early postnatal malnutrition in the rat. Beta-cell regeneration, including proliferation and neogenesis, was studied after neonatal beta-cell destruction by streptozotocin (STZ). STZ was injected at birth and maternal food restriction was continued until weaning. Beta-cell mass, proliferation, and islet number were quantified by morphometrical measurements on pancreatic sections in STZ-injected normal (C-STZ) and malnourished (R-STZ) rats, with noninjected C and R rats as controls. At day 4, only 20% of the beta cell mass remained in C-STZ rats. It regenerated to 50% that of noninjected controls, mainly through active neogenesis, as shown by the entire recovery of islet number/cm(2), and also through moderately increased beta-cell proliferation. In contrast, beta-cell mass from R-STZ animals poorly regenerated, despite a dramatic increase of beta-cell proliferation, because islet number/cm(2) recovered insufficiently. In conclusion, perinatal malnutrition impairs neogenesis and the capacity of beta-cell regeneration by neogenesis but preserves beta-cell proliferation, which remains the elective choice to increase beta-cell mass. These results provide an explanation for the impaired capacity of malnourished animals to adapt their beta-cell mass during aging or pregnancy, which aggravate glucose tolerance. PMID- 11099481 TI - Patterns within protein/polyphosphoinositide interactions provide specific targets for therapeutic intervention. AB - Signaling pathways involving the inositol polyphosphates and the polyphosphoinositides have become intricately linked with a number of disease states. More recently, this has principally involved the 3-phosphorylated products of phosphoinositide 3-kinase, an enzyme that itself shows oncogenic activity and has hence become of interest in the design of antitumorigenic drugs. The downstream effectors of phosphoinositide 3-kinase are involved in different aspects of cellular signaling and cytoskeleton and trafficking events that are linked to specific polyphosphoinositide binding properties of specific protein domains, which themselves have emerging roles in specific disease states. Our recent findings have demonstrated that there is a selectivity of the intracellular effects of extracellularly applied inositol polyphosphates in their abilities to inhibit a range of growth-related in vivo assay conditions, and that these can themselves be linked to the inhibition of the membrane localization of a green fluorescent protein (GFP) -tagged PH domain. We propose that GFP fusions of the polyphosphoinositides binding domains of specific proteins of interest can be used in high-throughput investigations of the therapeutic value of specific inositol polyphosphates analogs. Inhibition of in vivo membrane targeting of these domains from proteins involved in cell growth and tumorigenesis can thus be used in the search for new anticancer drugs. PMID- 11099482 TI - Inhibition of NF-kappaB and AP-1 activation by R- and S-flurbiprofen. AB - R-flurbiprofen is considered the 'inactive' isomer of the nonsteroidal anti inflammatory drug (NSAID), flurbiprofen, because it does not inhibit cyclooxygenase (COX) activity. However, previous studies have revealed that it has antinociceptive and antitum or effects not due to epimerization to the cyclooxygenase-inhibiting S-isomer. Here, we show that R-flurbiprofen has additional anti-inflammatory activity comparable with that of dexamethasone in the zymosan-induced paw inflammation model in rats. Different criteria suggest that the observed effects are mediated at least in part through inhibition of NF kB activation: R-flurbiprofen inhibited i) LPS-induced NF-kB DNA binding activity in RAW 264.7 macrophages, ii) translocation of the p65 subunit of NF-kB into the nucleus of these cells, and iii) zymosan-induced NF-kB-dependent gene transcription in the inflamed paw and spinal cord of rats. S-flurbiprofen produced similar effects but was less potent. In addition, R-flurbiprofen inhibited DNA binding activity of AP-1, another key regulatory transcription factor in inflammatory processes. Because R-flurbiprofen does not cause gastrointestinal mucosal damage or other side effects associated with long-term NSAID or glucocorticoid use, it might be a useful drug in inflammatory or other diseases in which increased or constitutive NF-kB and AP-1 activation are involved in the pathophysiological processes. PMID- 11099483 TI - Retinoids as ligands and coactivators of protein kinase C alpha. AB - Whereas retinoic acids control nuclear events, a second class of retinol metabolites, that is, the hydroxylated forms exemplified by 14-hydroxy-retro retinol (HRR), operate primarily in the cytoplasm. They function as regulatory cofactors for cell survival/cell death decisions. In accordance with these biological aspects, we demonstrate that these retinoids bound protein kinase C (PKC) alpha with nanomolar affinity and markedly enhance the activation of PKC alpha and the entire downstream MAP kinase pathway by reactive oxygen species. HRR was 10 times more efficient than retinol, and the optimum doses are 10-7 and 10-6 M, respectively. PKC alpha activation was reversed rapidly by imposition of reducing conditions. The retinoid binding site was mapped to the first cysteine rich region in the regulatory domain, C1A, yet was distinct from the binding sites of diacylglycerol and phorbol esters. The C1B domain bound retinoids poorly. The emerging theme is that retinoids serve as redox regulators of protein kinase C. PMID- 11099484 TI - HIV induces lymphocyte apoptosis by a p53-initiated, mitochondrial-mediated mechanism. AB - HIV-1 induces apoptosis and leads to CD4+ T-lymphocyte depletion in humans. It is still unclear whether HIV-1 kills infected cells directly or indirectly. To elucidate the mechanisms of HIV-1-induced apoptosis, we infected human CD4+ T cells with HIV-1. Enzymatic analysis with fluorometric substrates showed that caspase 2, 3, and 9 were activated in CD4+ T cells with peak levels 48 h after infection. Immunoblotting analysis confirmed the cleavage of pro-caspase 3 and 9, and of specific caspase substrates. Release of cytochrome c and apoptosis inducing factor (AIF) from mitochondria was observed in HIV-infected cells. The cytochrome c and AIF release preceded the reduction of the mitochondrial transmembrane potential and nuclear chromatin condensation. H IV infection led to phosphorylation of p53 at the Ser15 residue, detectable as early as 24 h after infection. The p53 phosphorylation was followed by increased mRNA and protein expression of p21, Bax, HDM2, and p53. Up-regulation of surface FasL expression, accompanied by a down-regulation of Fas-associated proteins (FADD, DAXX, and RIP), was observed 72 h after infection. Our results suggest that HIV activates the p53 pathway, leading to cytochrome c and AIF release with ensuing caspase activation. PMID- 11099485 TI - Ceramide binds to the CaLB domain of cytosolic phospholipase A2 and facilitates its membrane docking and arachidonic acid release. AB - Excessive production of eicosanoids is characteristic of many inflammatory diseases. In this study we show that ceramide, which is an early messenger of inflammatory cytokine action, exerts a dual effect on the cytosolic phospholipase A2 (cPLA2), the rate-limiting enzyme in arachidonic acid release and subsequent eicosanoid formation. Stimulation of renal mesangial cells with exogenous short chain ceramide analogs for 30 and 60 min leads to a concentration-dependent increase in arachidonic acid release that is not blocked by specific inhibitors of mitogen-activated protein kinase pathways. This suggests that these established upstream activators of cPLA2 are not involved in ceramide-induced arachidonic acid release. By use of photoactivatable ceramide analogs, D- and L [125I]3-trifluoromethyl-3-(m-iodophenyl)diazirine-ceramides (TID-ceramides), we observed a direct interaction of ceramide with cPLA2. This interaction was independent of the absolute configuration as D- and L-TID-ceramide were equally effective in binding to cPLA2. Moreover, recombinant CaLB domain of cPLA2 as well as a mutant deficient in the connecting 'hinge' domain of cPLA2, efficiently bound D- and L-TID-ceramides, whereas the catalytic domain did not interact with TID-ceramides. In vitro binding assays reveal that stearoyl-arachidonyl phosphatidylcholine (SAPC)-liposomes containing increasing mol% of ceramide lead to an increased association of recombinant cPLA2 to the liposomes. Furthermore, measurement of cPLA2 activity in vitro shows that the presence of SAPC-liposomes resulted in only weak cPLA2 activity. However, the activity dramatically increases by addition of ceramide to the liposomes. Furthermore, liposomes containing SAPC and sphingomyelin resulted in no better substrate than SAPC liposomes, unless bacterial sphingomyelinase was added to generate ceramide, which then causes a marked increase in cPLA2 activity. These results demonstrate that ceramide can interact directly with cPLA2 via the CaLB domain and thereby serves as a membrane-docking device that facilitates cPLA2 action in inflammatory diseases. PMID- 11099486 TI - The common Arg972 polymorphism in insulin receptor substrate-1 causes apoptosis of human pancreatic islets. AB - Molecular scanning of human IRS-1 gene revealed a common polymorphism causing Gly ->Arg972 change. Diabetic and pre-diabetic carriers of Arg972 IRS-1 are characterized by low fasting levels of insulin and C-peptide. To investigate directly whether the Arg 972 IRS-1 affects human islet cells survival, we took advantage of the unique opportunity to analyze pancreatic islets isolated from three donors heterozygous for the Arg972 and six donors carrying wild-type IRS-1. Islets from carriers of Arg972 IRS-1 showed a two-fold increase in the number of apoptotic cells as compared with wild-type. IRS-1-associated PI3-kinase activity was decreased in islets from carriers of Arg972 IRS-1. Same results were reproduced in RIN rat b-cell lines stably expressing wild-type IRS-1 or Arg972 IRS-1. Using these cells, we characterized the downstream pathway by which Arg972 IRS-1 impairs b-cell survival. RIN-Arg972 cells exhibited a marked impairment in the sequential activation of PI3-kinase, Akt, and BAD as compared with RI N-WT. Impaired BAD phosphorylation resulted in increased binding to Bcl-XL instead of 14-3-3 protein, thus sequestering the Bcl-XL antiapoptotic protein to promote survival. Both caspase-9 and caspase-3 activities were increased in RIN-Arg972 cells. The results show that the common Arg972 polymorphism in IRS-1 impairs human b-cell survival and causes resistance to antiapoptotic effects of insulin by affecting the PI3-kinase/Akt survival pathway. These findings establish an important role for the insulin signaling in human b-cell survival and suggest that genetic defects in early steps of insulin signaling may contribute to b-cell failure. PMID- 11099487 TI - A retro-inverso peptide homologous to helix 1 of c-Myc is a potent and specific inhibitor of proliferation in different cellular systems. AB - In 1998 we reported that an L-peptide derived from H1 of c-Myc (Int-H1-S6A,F8A), linked to an internalization sequence from the third a-helix of Antennapedia, was endowed with an antiproliferative and proapoptotic activity toward a human mammary cancer cell line: The activity apparently depends upon the presence of the Myc motif. In the present work we have added new dimensions to our original findings. It is known that short retro-inverso (RI-) peptides can assume a 3D conformation very close to their corresponding L-forms and can be recognized by the same monoclonal antibody. We synthesized a RI-peptide form of our original L peptide: It was much more resistant to serum peptidases than the original molecule (a half life of days rather than hours); in addition, the RI-form of the original Antennapedia internalization sequence was perfectly capable of carrying a D-peptide into human cells. We have studied three different potentially active peptides. L-peptides: Int-H1wt, Int-H1-S6A,F8A. D-peptides: RI-Int -H1-S6A,F8A. We have also studied three presumed control peptides: Int and RI-Int (no H1 motif), H1-S6A,F8A (no internalization sequence). Both 'active' and 'control' peptides have essentially confirmed our expectations, however, in cells treated with the higher concentration (10 mM) of the control peptide RI-Int, non-Myc related side effects could be detected. In order to investigate whether the antiproliferative activities displayed by some of our molecules were indeed related to an interference with the role of c-Myc (and molecules of the family), we chose an iso-amphipathic modified peptide of the H1 motif, with a proximity coefficient >50% and where the major change was at position 7 (F-->A). From a family of 73 H1 motifs belonging to (H1-Loop-H2) hu man sequences, the smallest evolutionary distance from our reference peptide was observed for the H1 of N Myc, L-Myc, c-Myc, H1-S6A,F8A of c-Myc, and Max, in that order. Our reference peptide was therefore appropriate as a check of whether we were indeed observing activities related to Myc functions. Both Int-H1isoamph and the corresponding RI Int-H1isoamph peptide were synthesized and studied. In terms of biological targets, we added to the human mammary cancer line of our previous work (MCF-7 cells) a colon cancer line (HCT-116 cells) and also a system of normal cells: human peripheral blood lymphocytes (PBLs) stimulated with phytohemoagglutinin (PHA). Peptides carrying an iso-amphipathic-modified H1 sequence were always very clearly (3-10 times) less active than the corresponding peptides carrying a conserved "H1 of Myc" motif. This finding was noted in five independent situations (all the cellular models considered at the present time): MCF-7 cells treated with L-peptides; MCF-7 cells treated with RI-peptides; HCT-116 cells treated with L-peptides; PBLs treated with L-peptides; PBLs treated with RI peptides. Modulation of transcription levels of ornithine decarboxylase (ODC), p53, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in PBLs treated with our different molecules, was well compatible with an interference by our active peptides at the level of Myc transcriptional activity. We had already reported a similar observation in MCF-7 cells. On a molar basis, RI-peptides were about 5-10 times more potent and 30-35 times more stable in complete culture medium, than their corresponding L-forms. RI-Int can probably internalize longer peptido mimetic molecules (for instance molecules mimetic of (H1-Loop-H2), or even more. These possibilities open the way to rodent studies and to more potent/selective Myc inhibitors-two steps closer to a potential drug. PMID- 11099488 TI - Mechanism and significance of increased glutathione level in human hepatocellular carcinoma and liver regeneration. AB - Increased glutathione (GSH) level occurs early during liver regeneration and in many drug and/or radiation-resistant tumors. Whether GSH level is elevated in liver cancer is unknown. GSH levels and expression of GSH synthetic enzymes were measured in hepatocellular carcinoma (HCC) and normal liver. GSH levels doubled in HCC. The mRNA levels of g-glutamylcysteine synthetase heavy subunit (GCS-HS) and GSH synthetase (GS) doubled, whereas the expression of GCS light subunit was unchanged. Nuclear run-on assay showed that the rate of gene transcription doubled for both GCS-HS and GS. In HCC, there is increased binding to anti oxidant response, AP-1 and NF-kB, three cis-acting elements in the 5'-flanking region of the human GCS-HS important for its transcriptional regulation. The role of GSH in cell growth was examined by using HepG2 cells. Cell GSH level was varied by treating cells with cystine (0 to 0.2 mM) with or without GSH ester or buthionine sulfoximine. Cell GSH level correlated directly with growth rate. Finally, preventing the increase in GSH after two-thirds partial hepatectomy blunted liver regeneration. Thus, GSH level is increased during liver growth as a result of up-regulation of GCS-HS and GS. This increase, in turn, facilitates growth. PMID- 11099489 TI - Triiodothyronine-mediated up-regulation of UCP2 and UCP3 mRNA expression in human skeletal muscle without coordinated induction of mitochondrial respiratory chain genes. AB - Triiodothyronine (T3) increases mitochondrial respiration and promotes the uncoupling between oxygen consumption and ATP synthesis. T3 effect is mediated partly through transcriptional control of genes encoding mitochondrial proteins. We determined the effect of T3 on mRNA levels of uncoupling proteins (UCP) and proteins involved in the biogenesis of the respiratory chain in human skeletal muscle and on UCP2 mRNA expression in adipose tissue. Ten young, healthy males received 75 to 100 5g of T3 per day for 14 days. The increase in plasma-free T3 levels was associated with an increase of resting metabolic rate and a decrease of respiratory quotient. In skeletal muscle, treatment with T3 induced a twofold increase of both UCP2 and UCP3 mRNA levels (p c oxidase subunits 2 and 4, nuclear respiratory factor 1, mitochondrial transcription factor A, and the co-activator PGC1 did not change during the treatment. In adipose tissue, UCP2 mRNA levels increased threefold. The direct effect of T3 on skeletal muscle an d adipose tissue UCP2 and UCP3 mRNA expression was demonstrated in vitro in human primary cultures. Our data show that T3 induces UCP2 and UCP3 mRNA expression in humans. In skeletal muscle, UCP regulation by T3 is not associated with the transcriptional regulation of respiratory chain proteins. PMID- 11099490 TI - Peroxynitrite induces integrin-dependent adhesion of human neutrophils to endothelial cells via activation of the Raf-1/MEK/Erk pathway. AB - Accumulating evidence suggests that enhanced peroxynitrite (ONOO-) formation occurs during inflammation. We have studied the impact and the mechanisms of ONOO action on expression of adhesion molecules on human neutrophils and coronary artery endothelial cells (HCAEC) and binding of neutrophils to HCAEC. Addition of ONOO- (0.1 to 200 5M) to isolated neutrophils resulted in a concentration dependent down-regulation of L-selectin expression, and up-regulation of CD11b/CD18 expression. ONOO- stimulation of Erk activity was accompanied by activation of Ras, Raf-1 and MEK (mitogen-activated protein kinase kinase), and was sensitive to the MEK inhibitor PD 98059. We have observed a tight association between Erk activation and changes in CD11b/CD18 expression. ONOO- also evoked activation of neutrophil p38 MAPK. Neither ONOO--induced up-regulation of CD11b/CD18 expression nor Erk activation was affected by SB 203580, a selective inhibitor of p38 MAPK. ONOO- by itself had little effect on expression of ICAM-1 and E-selectin on HCAEC, whereas it markedly enhanced attachment of neutrophils to lipopolysaccharide-activated HCAEC only when it was added together with neutrophils. Increases in neutrophil adhesion evoked by ONOO- were blocked by an anti-CD18 monoclonal antibody. These data suggest that ONOO- activates Erk in neutrophils via the Ras/Raf-1/MEK signal transduction pathway, leading to up regulation of surface expression of CD11b/CD18 and consequently to increased neutrophil adhesion to endothelial cells. PMID- 11099491 TI - Reduction of Abeta accumulation in the Tg2576 animal model of Alzheimer's disease after oral administration of the phosphatidyl-inositol kinase inhibitor wortmannin. AB - The abnormal accumulation of the amyloid beta protein (Abeta) has been implicated as an early and critical event in the etiology and pathogenesis of Alzheimer's disease (AD). Compounds that reduce Abeta accumulation may therefore be useful therapeutically. In cell-based screens we detected a significant reduction in Abeta concentration after treatment with the phosphatidylinositol kinase inhibitors wortmannin and LY294002. To determine the effect of this class of compounds on in vivo Abeta accumulation, we administered wortmannin to the Tg2576 mouse model of AD. Oral administration of wortmannin over four months resulted in a significant, non-overlapping 40%-50% reduction in the number of senile plaques, one of the pathological hallmarks of AD. Sandwich ELISA analysis of formic acid extractable Abeta in the brain of treated animals indicates that both Abeta40 and the longer, more amyloidogenic form of the peptide, Abeta42, were significantly reduced. These data provide the first direct evidence that compounds identified by their ability to reduce Abeta concentration in vitro can reduce Abeta accumulation and deposition in the brain, thus establishing a basic paradigm for the identification and evaluation of additional compounds that lower Abeta accumulation. PMID- 11099492 TI - Connexin 43 hemi channels mediate Ca2+-regulated transmembrane NAD+ fluxes in intact cells. AB - A previously unrecognized passive transport for pyridine dinucleotides has been described recently in the plasmamembrane of several mammalian cells. Despite elucidation of some functional and kinetic properties of this transport system, it is still undefined at the molecular level. Therefore, we have addressed the molecular characterization of the NAD+ transporter and identified it as connexin 43 (Cx43). This is a structural component of hexameric hemichannels that, when juxtaposed on adjacent cells, builds up intercellular gap junctions and mediates exchange of molecules between cells. However, the role of connexin hemichannels as potential pores in individual, noncoupled cells remains elusive. Bidirectional NAD+ transport in isolated Cx43-expressing mur ine 3T3 fibroblasts was affected by known modulators of connexin-mediated intercellular coupling and was completely inhibited by treatment of the cells with a Cx43-antisense oligonucleotide. NAD+ transport in proteoliposomes reconstituted with 3T3 membrane proteins was inhibited in the presence of a monoclonal anti-Cx43 antibody. Finally, Cx43 immunopurified to homogeneity was reconstituted in unilamellar proteoliposomes, which displayed full NAD+-transporting activity. This finding is the first evidence that connexin hemichannels can mediate transmembrane fluxes of a nucleotide in whole cells: The pleiotropy of NAD+ dependent cellular events, including redox reactions, signaling, and DNA repair, implicates Cx43 hemichannels in intercellular NAD+ trafficking, which suggests new paracrine functions of NAD. PMID- 11099493 TI - Translocation of jellyfish green fluorescent protein via the Tat system of Escherichia coli and change of its periplasmic localization in response to osmotic up-shock. AB - The bacterial twin arginine translocation (Tat) pathway is capable of exporting cofactor-containing enzymes into the periplasm. To assess the capacity of the Tat pathway to export heterologous proteins and to gain information about the property of the periplasm, we fused the twin arginine signal peptide of the trimethylamine N-oxide reductase to the jellyfish green fluorescent protein (GFP). Unlike the Sec pathway, the Tat system successfully exported correctly folded GFP into the periplasm of Escherichia coli. Interestingly, GFP appeared as a halo in most cells and occasionally showed a polar localization in wild type strains. When subjected to a mild osmotic up-shock, GFP relocalized very quickly at the two poles of the cells. The conversion from the halo structure to a periplasmic gathering at particular locations was also observed with spherical cells of the DeltarodA-pbpA mutant or of the wild type strain treated with lysozyme. Therefore, the periplasm is not a uniform compartment and the polarization of GFP is unlikely to be caused by simple invagination of the cytoplasmic membrane at the poles. Moreover, the polar gathering of GFP is reversible; the reversion was accelerated by glucose and inhibited by azide and carbonyl cyanide m-chlorophenylhydrazone, indicating an active adaptation of the bacteria to the osmolarity in the medium. These results strongly suggest a relocalization of periplasmic substances in response to environmental changes. The polar area might be the preferential zone where bacteria sense the change in the environment. PMID- 11099494 TI - X protein of hepatitis B virus inhibits Fas-mediated apoptosis and is associated with up-regulation of the SAPK/JNK pathway. AB - The X protein from a chronic strain of hepatitis B virus (HBx) was determined to inhibit Fas-mediated apoptosis and promote cell survival. Fas-mediated apoptosis is the major cause of hepatocyte damage during liver disease. Experiments demonstrated that cell death caused by anti-Fas antibodies was blocked by the expression of HBx in human primary hepatocytes and mouse embryo fibroblasts. This effect was also observed in mouse erythroleukemia cells that lacked p53, indicating that protection against Fas-mediated apoptosis was independent of p53. Components of the signal transduction pathways involved in this protection were studied. The SAPK/JNK pathway has previously been suggested to be a survival pathway for some cells undergoing Fas-mediated apoptosis, and kinase assays showed that SAPK activity was highly up-regulated in cells expressing the HBx protein. Normal mouse fibroblasts expressing HBx were protected from death, whereas identical fibroblasts lacking the SEK1 component from the SAPK pathway succumbed to Fas-mediated apoptosis, whether HBx was present or not. Assays showed that caspase 3 and 8 activities and the release of cytochrome c from mitochondria were inhibited, in the presence of HBx, following stimulation with anti-Fas antibodies. Coprecipitation and confocal immunofluorescence microscopy experiments demonstrated that HBx localizes with a cytoplasmic complex containing MEKK1, SEK1, SAPK, and 14-3-3 proteins. Finally, mutational analysis of HBx demonstrated that a potential binding region for 14-3-3 proteins was essential for induction of SAPK/JNK activity and protection from Fas-mediated apoptosis. PMID- 11099495 TI - Mouse ribonucleotide reductase control: influence of substrate binding upon interactions with allosteric effectors. AB - Using ribonucleotide reductase encoded by vaccinia virus as a model for the mammalian enzyme, our laboratory developed an assay that allows simultaneous monitoring of the reduction of ADP, CDP, GDP, and UDP. That study found ADP reduction to be specifically inhibited by ADP itself. To learn whether this effect is significant for cellular regulation, we have analyzed recombinant mouse ribonucleotide reductase. We report that allosteric control properties originally described in single-substrate assays operate also under our four-substrate assay conditions. Three distinctions from the vaccinia enzyme were seen: 1) higher sensitivity to allosteric modifiers; 2) higher activity with UDP as substrate; and 3) significant inhibition by ADP of GDP reduction as well as that of ADP itself. Studies of the effects of ADP and other substrates upon binding of effectors indicate that binding of ribonucleoside diphosphates at the catalytic site influences dNTP binding at the specificity site. We also examined the activities of hybrid ribonucleotide reductases, composed of a mouse subunit combined with a vaccinia subunit. As previously reported, a vaccinia R1/mouse R2 hybrid has low but significant activity. Surprisingly, a mouse R1/vaccinia R2 hybrid was more active than either mouse R1/R2 or vaccinia R1/R2, possibly explaining why mutations affecting vaccinia ribonucleotide reductase have only small effects upon viral DNA replication. PMID- 11099496 TI - Insufficient phosphorylation prevents fc gamma RIIB from recruiting the SH2 domain-containing protein-tyrosine phosphatase SHP-1. AB - Fc gamma RIIB are IgG receptors that inhibit immunoreceptor tyrosine-based activation motif (ITAM)-dependent cell activation. Inhibition depends on an immunoreceptor tyrosine-based inhibition motif (ITIM) that is phosphorylated upon Fc gamma RIIB coaggregation with ITAM-bearing receptors and recruits SH2 domain containing phosphatases. Agarose bead-coated phosphorylated ITIM peptides (pITIMs) bind in vitro the single-SH2 inositol 5-phosphatases (SHIP1 and SHIP2) and the two-SH2 protein tyrosine phosphatases (SHP-1 and SHP-2). Phosphorylated Fc gamma RIIB, however, recruit selectively SHIP1/2 in vivo. We aimed here at explaining this discordance. We found that beads coated with low amounts of pITIM bound in vitro SHIP1, but not SHP-1, i.e. behaved as phosphorylated Fc gamma RIIB in vivo. The reason is that SHP-1 requires its two SH2 domains to bind on adjacent pITIMs. Consequently, the binding of SHP-1, but not of SHIP1, increased with pITIM density on beads. When trying to increase Fc gamma RIIB phosphorylation in B cells and mast cells, we found that concentrations of ligands optimal for Fc gamma RIIB phosphorylation failed to induce SHP-1 recruitment. SHP-1 was, however, recruited by Fc gamma RIIB when hyperphosphorylated following cell treatment with pervanadate. Our data suggest that Fc gamma RIIB phosphorylation may not be sufficient in vivo to enable the recruitment of SHP-1 but that (pathological?) conditions that would hyperphosphorylate Fc gamma RIIB might enable SHP-1 recruitment. PMID- 11099497 TI - Reaction mechanism and stereochemistry of gamma-hexachlorocyclohexane dehydrochlorinase LinA. AB - gamma-Hexachlorocyclohexane dehydrochlorinase (LinA) catalyzes the initial steps in the biotransformation of the important insecticide gamma-hexachlorocyclohexane (gamma-HCH) by the soil bacterium Sphingomonas paucimobilis UT26. Stereochemical analysis of the reaction products formed during conversion of gamma-HCH by LinA was investigated by GC-MS, NMR, CD, and molecular modeling. The NMR spectra of 1,3,4,5,6-pentachlorocyclohexene (PCCH) produced from gamma-HCH using either enzymatic dehydrochlorination or alkaline dehydrochlorination were compared and found to be identical. Both enantiomers present in the racemate of synthetic gamma-PCCH were converted by LinA, each at a different rate. 1,2,4 trichlorobenzene (1,2,4-TCB) was detected as the only product of the biotransformation of biosynthetic gamma-PCCH. 1,2,4-TCB and 1,2,3-TCB were identified as the dehydrochlorination products of racemic gamma-PCCH. delta-PCCH was detected as the only product of dehydrochlorination of delta-HCH. LinA requires the presence of a 1,2-biaxial HCl pair on a substrate molecule. LinA enantiotopologically differentiates two 1,2-biaxial HCl pairs present on gamma HCH and gives rise to a single PCCH enantiomer 1,3(R),4(S),5(S),6(R)-PCCH. Furthermore, LinA enantiomerically differentiates 1,3(S),4(R),5(R),6(S)-PCCH and 1,3(R),4(S),5(S),6(R)-PCCH. The proposed mechanism of enzymatic biotransformation of gamma-HCH to 1,2,4-TCB by LinA consists of two 1,2-anti conformationally dependent dehydrochlorinations followed by 1,4-anti dehydrochlorination. PMID- 11099498 TI - Regulation of Rac and Cdc42 pathways by G(i) during lysophosphatidic acid-induced cell spreading. AB - The pertussis toxin-sensitive G protein, G(i), has been implicated in lysophosphatidic acid-induced cell mitogenesis and migration, but the mechanisms remain to be detailed. In the present study, we found that pertussis toxin blocks lysophosphatidic acid-induced cell spreading of NIH 3T3 fibroblasts on fibronectin. This prevention of cell spreading was eliminated by the expression of constitutively active mutants of Rho family small GTP-binding proteins, Rac and Cdc42, but not by Rho. In addition, activation of the endogenous forms was suppressed by pertussis toxin, indicating that G(i)-induced cell spreading is mediated through the Rac and Cdc42 pathway. Transfection of constitutively active mutants of G alpha(i) and G alpha(11) and G beta gamma subunits enhanced spreading of pertussis toxin-treated cells. G beta(1) with G gamma(12), a major G gamma form in fibroblasts, was more effective for increasing cell spreading than G beta(1)gamma(2) or G beta(1) plus G gamma(12)S2A, a mutant in which Ser-2, a phosphorylation site for protein kinase C, is replaced with alanine. In addition, a protein kinase C inhibitor diminished G beta(1)gamma(12)-induced cell spreading, suggesting a role for phosphorylation of the protein. These findings indicate that both G alpha(i) and G beta gamma stimulate Rac and Cdc42 pathways with lysophosphatidic acid-induced cell spreading on fibronectin. PMID- 11099499 TI - Characterization of histatin 5 with respect to amphipathicity, hydrophobicity, and effects on cell and mitochondrial membrane integrity excludes a candidacidal mechanism of pore formation. AB - Histatin 5 is a 24-residue peptide from human saliva with antifungal properties. We recently demonstrated that histatin 5 translocates across the yeast membrane and targets to the mitochondria, suggesting an unusual antifungal mechanism (Helmerhorst, E. J., Breeuwer, P., van't Hof, W., Walgreen-Weterings, E., Oomen, L. C. J. M., Veerman, E. C. I., Nieuw Amerongen, A. V., and Abee, T. (1999) J. Biol. Chem. 274, 7286-7291). The present study used specifically designed synthetic analogs of histatin 5 to elucidate the role of peptide amphipathicity, hydrophobicity, and the propensity to adopt alpha-helical structures in relation to membrane permeabilization and fungicidal activity. Studies included circular dichroism measurements, evaluation of the effects on the cytoplasmic transmembrane potential and on the respiration of isolated mitochondria, and analysis of the peptide hydrophobicity/amphipathicity relationship (Eisenberg, D. (1984) Annu. Rev. Biochem. 53, 595-623). The 14-residue synthetic peptides used were dh-5, comprising the functional domain of histatin 5, and dhvar1 and dhvar4, both designed to maximize amphipathic characteristics. The results obtained show that the amphipathic analogs exhibited a high fungicidal activity, a high propensity to form an alpha-helix, dissipated the cytoplasmic transmembrane potential, and uncoupled the respiration of isolated mitochondria, similar to the pore-forming peptide PGLa (Peptide with N-terminal Glycine and C-terminal Leucine amide). In contrast, histatin 5 and dh-5 showed fewer or none of these features. The difference in these functional characteristics between histatin 5 and dh-5 on the one hand and dhvar1, dhvar4, and PGLa on the other hand correlated well with their predicted affinity for membranes based on hydrophobicity/amphipathicity analysis. These data indicate that the salivary protein histatin 5 exerts its antifungal function through a mechanism other than pore formation. PMID- 11099500 TI - Interaction of the type IIa Na/Pi cotransporter with PDZ proteins. AB - The type IIa Na(+)-dependent inorganic phosphate (Na/P(i)) cotransporter is localized in the apical membrane of proximal tubular cells and is regulated by an endocytotic pathway. Because molecular processes such as apical sorting, internalization, or subsequent degradation might be assisted by associated proteins, a yeast two-hybrid screen against the C-terminal, cytosolic tail of type IIa cotransporter was designed. Most of the potential proteins found belonged to proteins with multiple PDZ modules and were either identical/related to PDZK1 or identical to NHERF-1. Yeast trap truncation assays confined the peptide-protein association to the C-terminal amino acid residues TRL of type IIa cotransporter and to single PDZ domains of each identified protein, respectively. The specificity of these interactions were confirmed in yeast by testing other apical localized transmembraneous proteins. Moreover, the type IIa protein was recovered in vitro by glutathione S-transferase-fused PDZ proteins from isolated renal brush border membranes or from type IIa-expressing oocytes. Further, these PDZ proteins are immunohistochemically detected either in the microvilli or in the subapical compartment of proximal tubular cells. Our results suggest that the type IIa Na/P(i) cotransporter interacts with various PDZ proteins that might be responsible for the apical sorting, parathyroid hormone controlled endocytosis or the lysosomal sorting of internalized type IIa cotransporter. PMID- 11099501 TI - Overexpression and characterization of carboxyl-terminal processing protease for precursor D1 protein: regulation of enzyme-substrate interaction by molecular environments. AB - CtpA, which is classified as a novel type of serine protease with a Ser/Lys catalytic dyad, is responsible for the C-terminal processing of precursor D1 protein (pD1) of the photosystem II reaction center, a process that is indispensable for the integration of water-splitting machinery in photosynthesis. In this study, overexpression in Escherichia coli and one-step purification of spinach CtpA were carried out to analyze the characteristics of this new type of protease and to elucidate the molecular interactions in the C-terminal processing of pD1 on the thylakoid membrane. The successful accumulation of functional CtpA in E. coli may argue against the possibility, based on homology to E. coli Tsp, that the enzyme is involved in the degradation of incomplete proteins in chloroplasts, e.g. by utilizing the ssrA-tagging system. Analysis using a synthetic pD1 oligopeptide demonstrated that the enzymatic properties (including substrate recognition) of overexpressed CtpA with an extra sequence of GSHMLE at the N terminus were indistinguishable from those of the native enzyme. CtpA was insensitive to penem, which has been shown to inhibit some Ser/Lys-type proteases, suggesting that the catalytic center of CtpA is quite unique. By using the substrate in different molecular environments (i.e. synthetic pD1 oligopeptide in solution and pD1 in photosystem II-enriched thylakoid membrane), we observed a dramatic difference in the pH profile and affinity for the substrate, suggesting the presence of a specific interaction of CtpA with a factor(s) that modulates the pH dependence of proteolytic action in response to physiological conditions. PMID- 11099502 TI - Thermal denaturation of the Na,K-ATPase provides evidence for alpha-alpha oligomeric interaction and gamma subunit association with the C-terminal domain. AB - Thermal denaturation can help elucidate protein domain substructure. We previously showed that the Na,K-ATPase partially unfolded when heated to 55 degrees C (Arystarkhova, E., Gibbons, D. L., and Sweadner, K. J. (1995) J. Biol. Chem. 270, 8785-8796). The beta subunit unfolded without leaving the membrane, but three transmembrane spans (M8-M10) and the C terminus of the alpha subunit were extruded, while the rest of alpha retained its normal topology with respect to the lipid bilayer. Here we investigated thermal denaturation further, with several salient results. First, trypsin sensitivity at both surfaces of alpha was increased, but not sensitivity to V8 protease, suggesting that the cytoplasmic domains and extruded domain were less tightly packed but still retained secondary structure. Second, thermal denaturation was accompanied by SDS-resistant aggregation of alpha subunits as dimers, trimers, and tetramers without beta or gamma subunits. This implies specific alpha-alpha contact. Third, the gamma subunit, like the C-terminal spans of alpha, was selectively lost from the membrane. This suggests its association with M8-M10 rather than the more firmly anchored transmembrane spans. The picture that emerges is of a Na,K-ATPase complex of alpha, beta, and gamma subunits in which alpha can associate in assemblies as large as tetramers via its cytoplasmic domain, while beta and gamma subunits associate with alpha primarily in its C-terminal portion, which has a unique structure and thermal instability. PMID- 11099503 TI - Effects of anoxia and the mitochondrion on expression of aerobic nuclear COX genes in yeast: evidence for a signaling pathway from the mitochondrial genome to the nucleus. AB - Eucaryotic cells contain at least two general classes of oxygen-regulated nuclear genes: aerobic genes and hypoxic genes. Hypoxic genes are induced upon exposure to anoxia while aerobic genes are down-regulated. Recently, it has been reported that induction of some hypoxic nuclear genes in mammals and yeast requires mitochondrial respiration and that cytochrome-c oxidase functions as an oxygen sensor during this process. In this study, we have examined the role of the mitochondrion and cytochrome-c oxidase in the expression of yeast aerobic nuclear COX genes. We have found that the down-regulation of these genes in anoxic cells is reflected in reduced levels of their subunit polypeptides and that cytochrome c oxidase subunits I, II, III, Vb, VI, VII, and VIIa are present in promitochondria from anoxic cells. By using nuclear cox mutants and mitochondrial rho(0) and mit(-) mutants, we have found that neither respiration nor cytochrome c oxidase is required for the down-regulation of these genes in cells exposed to anoxia but that a mitochondrial genome is required for their full expression under both normoxic and anoxic conditions. This requirement for a mitochondrial genome is unrelated to the presence or absence of a functional holocytochrome-c oxidase. We have also found that the down-regulation of these genes in cells exposed to anoxia and the down-regulation that results from the absence of a mitochondrial genome are independent of one another. These findings indicate that the mitochondrial genome, acting independently of respiration and oxidative phosphorylation, affects the expression of the aerobic nuclear COX genes and suggest the existence of a signaling pathway from the mitochondrial genome to the nucleus. PMID- 11099504 TI - Walker A lysine mutations of TAP1 and TAP2 interfere with peptide translocation but not peptide binding. AB - We generated mutants of the transporter associated with antigen-processing subunits TAP1 and TAP2 that were altered at the conserved lysine residue in the Walker A motifs of the nucleotide binding domains (NBD). In other ATP binding cassette transporters, mutations of the lysine have been shown to reduce or abrogate the ATP hydrolysis activity and in some cases impair nucleotide binding. Mutants TAP1(K544M) and TAP2(K509M) were expressed in insect cells, and the effects of the mutations on nucleotide binding, peptide binding, and peptide translocation were assessed. The mutant TAP1 subunit is significantly impaired for nucleotide binding relative to wild type TAP1. The identical mutation in TAP2 does not significantly impair nucleotide binding relative to wild type TAP2. Using fluorescence quenching assays to measure the binding of fluorescent peptides, we show that both mutants, in combination with their wild type partners, can bind peptides. Since the mutant TAP1 is significantly impaired for nucleotide binding, these results indicate that nucleotide binding to TAP1 is not a requirement for peptide binding to TAP complexes. Peptide translocation is undetectable for TAP1.TAP2(K509M) complexes, but low levels of translocation are detectable with TAP1(K544M).TAP2 complexes. These results suggest an impairment in nucleotide hydrolysis by TAP complexes containing either mutant TAP subunit and indicate that the presence of one intact TAP NBD is insufficient for efficient catalysis of peptide translocation. Taken together, these results also suggest the possibility of distinct functions for TAP1 and TAP2 NBD during a single translocation cycle. PMID- 11099505 TI - Peptide leucine arginine, a potent immunomodulatory peptide isolated and structurally characterized from the skin of the Northern Leopard frog, Rana pipiens. AB - On the basis of histamine release from rat peritoneal mast cells, an octadecapeptide was isolated from the skin extract of the Northern Leopard frog (Rana pipiens). This peptide was purified to homogeneity using reversed-phase high performance liquid chromatography and found to have the following primary structure by Edman degradation and pyridylethylation: LVRGCWTKSYPPKPCFVR, in which Cys(5) and Cys(15) are disulfide bridged. The peptide was named peptide leucine-arginine (pLR), reflecting the N- and C-terminal residues. Molecular modeling predicted that pLR possessed a rigid tertiary loop structure with flexible end regions. pLR was synthesized and elicited rapid, noncytolytic histamine release that had a 2-fold greater potency when compared with one of the most active histamine-liberating peptides, namely melittin. pLR was able to permeabilize negatively charged unilamellar lipid vesicles but not neutral vesicles, a finding that was consistent with its nonhemolytic action. pLR inhibited the early development of granulocyte macrophage colonies from bone marrow stem cells but did not induce apoptosis of the end stage granulocytes, i.e. mature neutrophils. pLR therefore displays biological activity with both granulopoietic progenitor cells and mast cells and thus represents a novel bioactive peptide from frog skin. PMID- 11099506 TI - Identification of the protein C/activated protein C binding sites on the endothelial cell protein C receptor. Implications for a novel mode of ligand recognition by a major histocompatibility complex class 1-type receptor. AB - The endothelial cell protein C receptor (EPCR) is an endothelial cell-specific transmembrane protein that binds both protein C and activated protein C (APC). EPCR regulates the protein C anticoagulant pathway by binding protein C and augmenting protein C activation by the thrombin-thrombomodulin complex. EPCR is homologous to the MHC class 1/CD1 family, members of which contain two alpha helices that sit upon an 8-stranded beta-sheet platform. In this study, we identified 10 residues that, when mutated to alanine, result in the loss of protein C/APC binding (Arg-81, Leu-82, Val-83, Glu-86, Arg-87, Phe-146, Tyr-154, Thr-157, Arg-158, and Glu-160). Glutamine substitutions at the four N-linked carbohydrate attachment sites of EPCR have little affect on APC binding, suggesting that the carbohydrate moieties of EPCR are not critical for ligand recognition. We then mapped the epitopes for four anti-human EPCR monoclonal antibodies (mAbs), two of which block EPCR/Fl-APC (APC labeled at the active site with fluorescein) interactions, whereas two do not. These epitopes were localized by generating human-mouse EPCR chimeric proteins, since the mAbs under investigation do not recognize mouse EPCR. We found that 5 of the 10 candidate residues for protein C/APC binding (Arg-81, Leu-82, Val-83, Glu-86, Arg-87) colocalize with the epitope for one of the blocking mAbs. Three-dimensional molecular modeling of EPCR indicates that the 10 protein C/APC binding candidate residues are clustered at the distal end of the two alpha-helical segments. Protein C activation studies on 293 cells that coexpress EPCR variants and thrombomodulin demonstrate that protein C binding to EPCR is necessary for the EPCR-dependent enhancement in protein activation by the thrombin-thrombomodulin complex. These studies indicate that EPCR has exploited the MHC class 1 fold for an alternative and possibly novel mode of ligand recognition. These studies are also the first to identify the protein C/APC binding region of EPCR and may provide useful information about molecular defects in EPCR that could contribute to cardiovascular disease susceptibility. PMID- 11099553 TI - Class II combination therapy (distal jet and Jasper Jumpers): a case report. AB - Class II combination therapy is a method that combines orthodontic and orthopedic mechanics in a single stage of treatment. Molar distalization is followed by fixed functional mechanics to reduce the dependence upon patient compliance while seeking more predictable completion of Class II correction. PMID- 11099554 TI - Class II correction-reducing patient compliance: a review of the available techniques. AB - The correction of Class II malocclusions has been hampered by the use of appliances which require the patient to co-operate with headgear, elastics, or the wearing of a removable appliance. 'Non-compliance therapy' involves the use of appliances which minimize the need for such co-operation and attempt to maximize the predictability of results. This article reviews and describes the types of appliances used, and their mode of action-based on the current available research. PMID- 11099555 TI - Matrix turnover. AB - This review concentrates on how the major component of extracellular matrix, collagen, is catabolized. This process is important in a number of aspects of orthodontics since matrix is constantly turning over, the rate of which differs in embryogenesis, ageing, disease, and physiological processes, such as orthodontic tooth movement. It is not the purpose of this review to consider each process in detail. The aim is to give a clear account of the matrix metalloproteinases (a major family of proteinases) including their classification, properties, and functions. PMID- 11099556 TI - The effects of orthognathic surgery on pharyngeal airway dimensions and quality of sleep. AB - Orthognathic surgery has been associated with airway narrowing and induction of sleep-related breathing disorders. Therefore, the pharyngeal airway dimensions of 32 orthognathic surgery cases were prospectively investigated, and the relationship between the surgery and sleep quality assessed. Digitized lateral cephalometric radiographs were used to compare oropharyngeal airway morphologies before and after surgery. Patients were assessed in two main surgical groups based on sagittal jaw relationship. A questionnaire was used to assess changes in daytime sleepiness. The mandibular surgery cases were also assessed by overnight domiciliary sleep monitoring. A significant decrease in the retrolingual airway dimension was found in all patients after mandibular setback surgery and a significant increase in this dimension after mandibular advancement. The questionnaire and sleep study revealed no significant changes in snoring incidence or apnoeic events after mandibular setback surgery. For the mandibular advancement group, a change in sleep quality was found, but only in cases with signs of a pre-existing sleep disorder. PMID- 11099557 TI - The influence of bracket base design on the strength of the bracket-cement interface. AB - The objectives of the study were to isolate the bracket-cement interface, and to determine the influence of bracket base morphology and orthodontic bonding agent chosen on strength of adhesion. The bracket bases evaluated included 60, 80, and 100 single mesh bases, a double mesh base, and the Dynalock, and Mini Twin bases. The strength of interface provided by each of these bases with Concise, Transbond, Right On, and non-encapsulated Fuji Ortho LC cements, was measured in tension and recorded in Mega Pascals. The single-mesh bases performed well with either Concise or Right On (11*88-22*72 MPa) and, other than the 80-mesh bracket, relatively poorly with Transbond (2*18-5*15 MPa). With Fuji Ortho LC, the single mesh bases performed well (6*05-12*19 MPa). The double mesh base performed well with Right On (13*75 MPa), and reasonably well with Concise, Transbond, and Fuji Ortho LC (6*00-9*20 MPa). The Dynalock and Mini Twin Bases performed fairly well with all cements (8*87-17*16 MPa). It was concluded that the orthodontic bonding agent selected would appear to largely determine the bond strength achieved with a particular bracket base design. A definite trend was difficult to identify in this study, and it appeared that certain combinations of bracket base and bonding agent performed optimally. Particular base designs may allow improved adhesive penetration or improved penetration of curing light. Alternatively, the dimension and distribution of resin/cement tags prescribed by one base could promote a stress distribution that is better resisted by a particular adhesive. PMID- 11099558 TI - The use of a cyanoacrylate adhesive for bonding orthodontic brackets: an ex-vivo study. AB - The purpose of this study was to evaluate the performance of a cyanoacrylate orthodontic adhesive with regard to tensile bond strength and bond failure location in comparison with a conventional no-mix orthodontic composite adhesive using stainless steel and ceramic brackets. One-hundred-and-twenty caries-free extracted premolar teeth were used in this study. There were 30 specimens for each tooth, adhesive, and bracket combination, and of these half were tested at 24 hours and half at 3 months. Hence, there were 15 samples in each test group. Bond strengths were assessed using an Instron Universal Testing Machine after storage for 24 hours and for 3 months at 37 degrees C in distilled water. Analysis of variance showed the mean bond strength of specimens bonded with cyanoacrylate was significantly lower than for those bonded with Right-on (P < 0.001). Weibull analysis showed that at a given stress the probability of failure significantly increased after 3 months for brackets bonded with cyanoacrylate. A Chi-square test of the ARI scores revealed no significant difference among the groups tested. This study showed that cyanoacrylate adhesives are unsuitable for use as a bonding agent in routine orthodontic practice. PMID- 11099559 TI - Evaluation of a reproduction technique for the study of the enamel composite/bracket base area. AB - The objective of the study was to evaluate a reproduction method that would enable the study of the enamel/ bracket/composite interface in vivo, and consisted of in vitro assessment of two different impression materials to compare reproduction of brackets bonded to extracted teeth followed by in vivo assessment of the superior material. In vitro standard edgewise brackets were bonded to two extracted teeth and impressions were taken using two different types of low viscosity silicone-based impression materials. A medium viscosity silicone impression material was used to support the original impression. Three impressions of both the gingival and occlusal aspect of the bracket base region were obtained using each of the impression materials. Replicas were then prepared for SEM viewing and these compared to SEMs of the real teeth for reproduction of detail. A 3-point Reproducibility Index was used to compare the SEM photographs of the comparable replicas. One impression material was clearly superior to the other and produced an acceptably accurate representation of the true clinical situation in three out of four samples. This material also performed well in the in vivo situation. The technique described is satisfactory for the production and analysis of SEM pictures of the enamel/composite/ bracket base interface in vivo. PMID- 11099560 TI - The need for cost-effectiveness. PMID- 11099561 TI - Statistical applications in orthodontics. Part I. Confidence intervals: an introduction. PMID- 11099562 TI - A survey of continuing professional education for orthodontists in 23 European countries. AB - This paper reports on a survey of the organization, forms and methods of funding continuing professional education (CPE) for those providing orthodontics in 23 European countries in 1997. A postal questionnaire was sent to all members of the EURO-QUAL II BIOMED project, who came from 28 countries, together with an explanatory letter. Answers were validated during a meeting of project participants and by further correspondence, when necessary. Completed questionnaires, which were subsequently validated, were returned by orthodontists from 23 countries and indicated that orthodontic CPE took place in 22 of the 23 countries surveyed. A number of different bodies were reported as organizing orthodontic CPE. This task was most frequently performed by orthodontic societies (in 22 out of 23 countries), but a number of other bodies were also involved. Practical technique courses were reported as taking place in 20 countries. Other frequently occurring forms of orthodontic CPE were lectures (in 18 countries) and study groups (in 15 countries). Orthodontists were reported as financing their CPE in 22 countries; others, who contributed to some or all of the costs, were the Government (in six countries), employers (in four countries), universities (in four countries), and a dental company (in one country). It was concluded that some orthodontic CPE took place in the vast majority of the countries surveyed, and was invariably organized by and paid for, wholly or in part by orthodontists themselves. PMID- 11099563 TI - A survey of the delegation of orthodontic tasks and the training of chairside support staff in 22 European countries. AB - This paper reports on a survey which was undertaken to investigate the delegation of orthodontic tasks and the training of chairside support staff in Europe. Two questionnaires were posted to all members of the EURO-QUAL BIOMED II project together with an explanatory letter. The first dealt with the delegation of nine clinical tasks during orthodontic treatment. The second with the types of chairside assistant employed in each country and the training that they are given. Completed questionnaires, which were subsequently validated, were returned by orthodontists from 22 countries. They indicated that there was no delegation of clinical tasks in six of the 22 countries and delegation of all nine tasks in five countries. The most commonly delegated tasks were taking radiographs (in 14 of the 22 countries) and taking impressions (in 13 of the 22 countries). The least commonly delegated tasks were cementing bands (in five of the 22 countries) and trying on bands (in six of the 22 countries). Seven of the 22 countries provided chairside assistants with training in some clinical orthodontic tasks. Eighteen of the 22 countries provided general training for chairside assistants and offered a qualification for chairside assistants. Four of these 18 countries reported that they only employed qualified chairside assistants. Of the four countries which reported that they did not provide a qualification for chairside assistants, two indicated that they employed chairside assistants with no formal training and two that they did not employ chairside assistants. It was concluded that there were wide variations within Europe as far as the training and employment of chairside assistants, with or without formal qualifications, and in the delegation of clinical orthodontic tasks to auxiliaries was concerned. PMID- 11099564 TI - Orthodontic banding cements. PMID- 11099565 TI - The B.S.S.O. M.Orth. Prize of the Royal College of Surgeons of England 1998. PMID- 11099566 TI - Hazards of orthodontics appliances and the oropharynx. AB - Occasionally orthodontic appliances or parts of orthodontic appliances have caused problems with either the airway or the gastrointestinal tract. The type of appliances that have caused problems and their clinical management are discussed. A case is described in which an upper removable appliance with inadequate retention became lodged in a patient's pharynx lacerating the palatine tonsils. Suggestions are made to try and avoid the problems that were encountered in this case and others reported in the literature in patients undergoing orthodontic treatment. PMID- 11099568 TI - Construction for the modern head: current concepts in craniofacial development. AB - The vertebrate head is a highly complex composite structure whose morphological characteristics are controlled at the level of the gene. There is now increasing evidence for the role of gene families that encode transcription factors in determining the embryonic plan of the developing craniofacial complex. These genes act as regulators of gene transcription being intimately involved with the control of complex interactions between multiple downstream genes. Combinatorial expression of the Hox genes (a family of highly conserved master regulatory genes related to the homeotic genes of the fruitfly Drosophila) have been shown to play a definitive role in patterning distinct regions of the craniofacial complex. In the vertebrate, Hox genes pattern the hindbrain and branchial regions of the developing head up to and including structures derived from the second branchial arch. The first branchial arch and more rostral regions of the head are patterned by groups of homeobox genes more diverged from the original Hox clusters. Transgenic mice, with targeted disruptions in many of these genes, are now providing insights into the molecular mechanisms that lie behind a number of craniofacial defects seen in man. PMID- 11099567 TI - Unusual indelible enamel staining following fixed appliance treatment. AB - Two cases are described of indelible enamel staining following fixed appliance therapy. The acquired pigmentation occurred in patients with an identifiable enamel defect prior to treatment. The interaction of factors to cause the staining is discussed and it's prevention in future cases highlighted. Subsequent restoration of the affected teeth is shown. PMID- 11099569 TI - An analysis of the skeletal relationships in a group of young people with hypodontia. AB - The objective of this investigation was to examine the dentofacial features of a group of patients with hypodontia, in particular assessing whether cephalometric analysis confirmed the clinical assumption of a reduced lower face height, and to determine the relationship of these facial features with different numbers of missing teeth. It took the form of a cephalometric study, undertaken in a dedicated Dental Hospital clinic for patients with hypodontia. The study group comprised 59 patients seen on the Hypodontia Clinic: 32 females, 27 males, mean age 13.1+/-3.1 years (range 6-23 years). The average number of missing teeth was 7 (SD 5), ranging from 1 to 21. The mean SNA, SNB, and MMA angles were within normal limits, but there was a statistically significant reduction in the MMA when more than one tooth type was missing (P = 0.007) and the ANB angle decreased as the number of missing tooth types increased (P = 0.034). The mean values for the whole sample were within the normal range and did not demonstrate any feature specific to the group, but patients with more severe hypodontia showed tendencies to a Class III skeletal relationship and a reduced maxillary-mandibular planes angle. PMID- 11099570 TI - An audit of the Yorkshire Regional Cleft Database. AB - This study assessed the validity of the Yorkshire regional orofacial cleft database by comparing the computer-based records with locally collated records of primary surgical events for babies born over a 2-year period (1994-1995). One hundred-and-thirty-two infants with clefts (excluding submucous cleft palate) were identified from the latter source with an equal proportion of unilateral cleft lip/palate and isolated cleft palate births. However, only 62 per cent of cases were recorded on the database and the reporting rate of individual cleft units was highly variable (43-85 per cent). In addition, there was a significant under-reporting of both cleft lip and isolated cleft palate cases (42 and 50 per cent ascertainment, respectively). Consequently, the database figures understated the prevalence of all cleft births, but especially of these two cleft subtypes. Conversely, the relative frequency of combined cleft lip and palate cases was exaggerated. The reasons for such discrepancies and possible improvements to data collection are discussed. PMID- 11099571 TI - A comparison between written, verbal, and videotape oral hygiene instruction for patients with fixed appliances. AB - The objective of the study was to compare the effectiveness of written, videotape, and one-to-one instruction upon the knowledge, oral hygiene standard, and gingival health of subjects undergoing orthodontic treatment with a lower fixed appliance. Subjects for whom fixed appliances had been fitted recently were divided randomly into three groups of 21, 22, and 22, respectively. Group 1 received written oral hygiene instruction, group 2 a specially made videotape, and group 3 saw a hygienist for one-to-one instruction. Results were assessed in terms of improvement in knowledge concerning oral hygiene procedures, and of plaque and gingival index scores. Analysis of variance revealed no significant main effects or interactions at P = 0.05, although the difference in the plaque index scores before and after instruction was close to significance. PMID- 11099572 TI - An ex vivo assessment of resin-modified glass ionomer bonding systems in relation to ceramic bracket debond. AB - This ex vivo study assessed three new resin-modified glass ionomer cements (Fuji ORTHO LC, Vitremer, and Dyract-Cem) in relation to ceramic bracket removal. It was hypothesized that the use of these cements would facilitate bracket removal and eliminate debond complications Eighty extracted premolar teeth were divided into four groups of 20 teeth and bonded with Intrigue brackets using each of the resin-modified cements (groups 1, 2, and 3), the control group 4 was bonded with Concise chemically-cured adhesive. The teeth were debonded by applying a shear load using an Instron universal testing machine. The mean force to debond was calculated for each group and each tooth was examined under the stereomicroscope to record the site of bond failure and the Adhesive Remnant Index (ARI). The results showed that the resin-modified cements were very effective at eliminating ceramic bracket debond problems. Bracket fracture was eliminated compared with a 40 per cent fracture rate with the control and the ARI scores were all reduced. The elimination of debond problems appears to be related to the significantly reduced (P < 0.001 using ANOVA and Tukey tests) mean and maximal debond forces compared with the control. PMID- 11099574 TI - A precis of the Proceedings of the First International Orthodontic Editors' Symposium, 28 April 2000. PMID- 11099575 TI - Specialist training and Europe. PMID- 11099576 TI - Statistical applications in orthodontics: part II. Confidence intervals for proportions and their differences. PMID- 11099578 TI - Buttons and elastics for the conservative treatment of the fractured mandible. PMID- 11099579 TI - A survey of perceived problems in orthodontic education in 23 European countries. AB - This paper reports on a survey of perceived problems in the provision of orthodontic education at the stages of undergraduate, postgraduate, and continuing professional education (CPE) in 23 European countries in 1997. A questionnaire, together with an explanatory letter, was mailed to all members of the EUROQUAL II BIOMED project. Answers were validated during a meeting of project participants and by further correspondence, when necessary. The topics covered in the questionnaire were adequacy of funding, numbers of orthodontic teachers, availability of equipment, regulations, training centres, numbers of orthodontists, availability of books, journals, and information technology. Completed questionnaires were returned by orthodontists from all 23 countries. Respondents from seven countries did not answer all questions. Respondents reported a perceived almost universal lack of adequate funding for postgraduate orthodontic training (from 18 out of 20 countries) and, to a lesser extent, at undergraduate (13 out of 20 countries) and CPE levels (17 out of 21 countries). Respondents from 12 of the 20 countries reported adequate numbers of qualified teachers at undergraduate level, but only seven out of 18 at postgraduate level and eight out of 19 for CPE. Lack of suitable equipment was reported as a more frequent problem by central and eastern European countries (six out of 20 countries at undergraduate level, eight out of 20 countries at postgraduate level, and 12 out of 19 at CPE level). Too few or too many regulations were only perceived to be a problem by the respondent from one country out of 19 at undergraduate level, by seven out of 19 at postgraduate level, and by eight out of 16 at CPE level). Lack of training centres was more frequently reported as a problem by respondents from central and eastern European countries, but was generally not perceived as a problem by respondents from west European countries. Respondents from seven countries reported a lack of training centres for CPE. Respondents from six countries reported that they perceived there to be too many orthodontists at postgraduate level, from seven countries that there were an appropriate number, and from seven that there were too few. A lack of books, journals, and information technology was reported to be a problem by respondents from four out of 19 countries at undergraduate level, eight out of 20 at postgraduate level, and 10 out of 20 at CPE level. At both undergraduate and postgraduate level, the majority of respondents from central and eastern European countries reported problems with books, journals, and information technology. The results of the survey confirmed many anecdotal impressions and provided an extremely useful background against which to formulate quality guidelines for orthodontic education in Europe. PMID- 11099580 TI - Suggested guidelines for the provision and assessment of orthodontic education in Europe. A report from the Professional Development Group of the EURO-QUAL BIOMED II Project. AB - The suggested guidelines for the provision and assessment of Orthodontic education in Europe, which are introduced, set out, and discussed in this paper, resulted from the work of the Professional Development Group (PDG) of the EURO QUAL BIOMED II project. They were published in the final report of the project, after comments had been received from a range of national and European bodies and societies, including the British and the European Orthodontic Societies, Royal Colleges, and the General Dental Council. PMID- 11099582 TI - Annual summary of vital statistics: trends in the health of Americans during the 20th century. AB - The overall improvement in the health of Americans over the 20th century is best exemplified by dramatic changes in 2 trends: 1) the age-adjusted death rate declined by about 74%, while 2) life expectancy increased 56%. Leading causes of death shifted from infectious to chronic diseases. In 1900, infectious respiratory diseases accounted for nearly a quarter of all deaths. In 1998, the 10 leading causes of death in the United States were, respectively, heart disease and cancer followed by stroke, chronic obstructive pulmonary disease, accidents (unintentional injuries), pneumonia and influenza, diabetes, suicide, kidney diseases, and chronic liver disease and cirrhosis. Together these leading causes accounted for 84% of all deaths. The size and composition of the American population is fundamentally affected by the fertility rate and the number of births. From the beginning of the century there was a steady decline in the fertility rate to a low point in 1936. The postwar baby boom peaked in 1957, when 123 of every 1000 women aged 15 to 44 years gave birth. Thereafter, fertility rates began a steady decline. Trends in the number of births parallel the trends in the fertility rate. Beginning in 1936 and continuing to 1956, there was precipitous decline in maternal mortality from 582 deaths per 100 000 live births in 1935 to 40 in 1956. Since 1950 the maternal mortality ratio dropped by 90% to 7.1 in 1998. The infant mortality rate has shown an exponential decline during the 20th century. In 1915, approximately 100 white infants per 1000 live births died in the first year of life; the rate for black infants was almost twice as high. In 1998, the infant mortality rate was 7.2 overall, 6.0 for white infants, and 14.3 for black infants. For children older than 1 year of age, the overall decline in mortality during the 20th century has been spectacular. In 1900, >3 in 100 children died between their first and 20th birthday; today, <2 in 1000 die. At the beginning of the 20th century, the leading causes of child mortality were infectious diseases, including diarrheal diseases, diphtheria, measles, pneumonia and influenza, scarlet fever, tuberculosis, typhoid and paratyphoid fevers, and whooping cough. Between 1900 and 1998, the percentage of child deaths attributable to infectious diseases declined from 61.6% to 2%. Accidents accounted for 6.3% of child deaths in 1900, but 43.9% in 1998. Between 1900 and 1998, the death rate from accidents, now usually called unintentional injuries, declined two-thirds, from 47. 5 to 15.9 deaths per 100 000. The child dependency ratio far exceeded the elderly dependency ratio during most of the 20th century, particularly during the first 70 years. The elderly ratio has gained incrementally since then and the large increase expected beginning in 2010 indicates that the difference in the 2 ratios will become considerably less by 2030. The challenge for the 21st century is how to balance the needs of children with the growing demands for a large aging population of elderly persons. PMID- 11099583 TI - Baby CareLink: using the internet and telemedicine to improve care for high-risk infants. AB - OBJECTIVE: To evaluate an Internet-based telemedicine program designed to reduce the costs of care, to provide enhanced medical, informational, and emotional support to families of very low birth weight (VLBW) infants during and after their neonatal intensive care unit (NICU) stay. BACKGROUND: Baby CareLink is a multifaceted telemedicine program that incorporates videoconferencing and World Wide Web (WWW) technologies to enhance interactions between families, staff, and community providers. The videoconferencing module allows virtual visits and distance learning from a family's home during an infant's hospitalization as well as virtual house calls and remote monitoring after discharge. Baby CareLink's WWW site contains information on issues that confront these families. In addition, its security architecture allows efficient and confidential sharing of patient based data and communications among authorized hospital and community users. DESIGN/METHODS: A randomized trial of Baby CareLink was conducted in a cohort of VLBW infants born between November 1997 and April 1999. Eligible infants were randomized within 10 days of birth. Families of intervention group infants were given access to the Baby CareLink telemedicine application. A multimedia computer with WWW browser and videoconferencing equipment was installed in their home within 3 weeks of birth. The control group received care as usually practiced in this NICU. Quality of care was assessed using a standardized family satisfaction survey administered after discharge. In addition, the effect of Baby CareLink on hospital length of stay as well as family visitation and interactions with infant and staff were measured. RESULTS: Of the 176 VLBW infants admitted during the study period, 30 control and 26 study patients were enrolled. The groups were similar in patient and family characteristics as well as rates of inpatient morbidity. The CareLink group reported higher overall quality of care. Families in the CareLink group reported significantly fewer problems with the overall quality of care received by their family (mean problem score: 3% vs 13%). In addition, CareLink families also reported greater satisfaction with the unit's physical environment and visitation policies (mean problem score: 13% vs 50%). The frequency of family visits, telephone calls to the NICU, and holding of the infant did not differ between groups. The duration of hospitalization until ultimate discharge home was similar in the 2 groups (68.5 +/- 28.3 vs 70.6 +/- 35.6 days). Among infants born weighing <1000 g (n = 31) there was a tendency toward shorter lengths of stay (77.4 +/- 26.2 vs 93.1 +/- 35.6 days). All infants in the CareLink group were discharged directly to home whereas 6/30 (20%) of control infants were transferred to community hospitals before ultimate discharge home. CONCLUSIONS: CareLink significantly improves family satisfaction with inpatient VLBW care and definitively lowers costs associated with hospital to hospital transfer. Our data suggest the use of telemedicine and the Internet support the educational and emotional needs of families facilitating earlier discharge to home of VLBW infants. We believe that further extension of the Baby CareLink model to the postdischarge period will significantly improve the coordination and efficiency of care. PMID- 11099584 TI - Providing pediatric subspecialty care: A workforce analysis. AAP Committee on Pediatric Workforce Subcommittee on Subspecialty Workforce. AB - OBJECTIVE: To provide a snapshot of pediatric subspecialty practice, examine issues pertaining to the subspecialty workforce, and analyze subspecialists' perspective on the health care market. BACKGROUND: Before the effort of the Future of Pediatric Education II (FOPE II) Project, very little information existed regarding the characteristics of the pediatric subspecialty workforce. This need was addressed through a comprehensive initiative involving cooperation between subspecialty sections of the American Academy of Pediatrics and other specialty societies. METHODS: Questionnaires were sent to all individuals, identified through exhaustive searches, who practiced in 17 pediatric medical and surgical subspecialty areas in 1997 and 1998. The survey elicited information about education and practice issues, including main practice setting, major professional activity, referrals, perceived competition, and local workforce requirements. The number of respondents used in the analyses ranged from 120 (plastic surgery) to 2034 (neonatology). In total, responses from 10 010 pediatric subspecialists were analyzed. RESULTS: For 13 of the subspecialties, a medical school setting was specified by the largest number of respondents within each subspecialty as their main employment site. Direct patient care was the major professional activity of the majority of respondents in all the subspecialties, with the exception of infectious diseases. Large numbers of subspecialists reported increases in the complexity of referral cases, ranging between 20% (cardiology) and 44% (critical care), with an average of 33% across the entire sample. In all subspecialties, a majority of respondents indicated that they faced competition for services in their area (range: 55%-90%; 71% across the entire sample); yet in none of the subspecialties did a majority report that they had modified their practice as a result of competition. In 15 of the 17 subspecialties, a majority stated that there would be no need in their community over the next 3 to 5 years for additional pediatric subspecialists in their discipline. Across the entire sample, 42% of respondents indicated that they or their employer would not be hiring additional, nonreplacement pediatric subspecialists in their field in the next 3 to 5 years (range: 20%-63%). CONCLUSION: This survey provides the first comprehensive analysis to date on how market forces are perceived to be affecting physicians in the pediatric subspecialty workforce. The data indicate that pediatric subspecialists in most areas are facing strong competitive pressures in the market, and that the market's ability to support additional subspecialists in many areas may be diminishing. PMID- 11099585 TI - The changing demographics of neonatal extracorporeal membrane oxygenation patients reported to the Extracorporeal Life Support Organization (ELSO) Registry. AB - BACKGROUND: Extracorporeal membrane oxygenation (ECMO) is an important treatment tool in the management of near-term and term neonates with severe hypoxemic respiratory failure. To better understand how health care for patients treated with ECMO has changed, we studied the demographic and treatment data reported to the Extracorporeal Life Support Organization (ELSO) registry from January 1, 1988, through January 1, 1998. METHODS: We used data stored in the ELSO registry and evaluated the changes in demographics, use of alternate therapies before ECMO, severity of illness, duration of ECMO therapy, and mortality over a 10-year period. All data on neonates reported between January 1, 1988, and January 1, 1998 were used. Verification checks were performed on all fields to eliminate nonsense outliers. We separated the neonates into 2 groups-those with and those without a congenital diaphragmatic hernia (CDH). All analyses were performed on the total group and each subgroup separately. Changes in continuous data were analyzed by year using analysis of variance. Year differences in categorical data were evaluated with chi(2) analysis. We also used the linear trend test and the Cochran-Armitage trend test to evaluate time-related changes. RESULTS: We reviewed 12 175 neonates. Over the decade, there were no changes in mean gestational age, gender, age at which ECMO was started, pH, or PaCO(2) just before ECMO. The proportion of neonates with CDH increased from 18% to 26%, while the proportion with respiratory distress syndrome decreased from 15% to 4%. Other diagnostic categories remained constant. The use of surfactant, high-frequency ventilation, and inhaled nitric oxide increased from 0% in 1988 to 36%, 46%, and 24%, respectively, in 1997. The mean peak pressure being used just before ECMO decreased (47 +/- 10 in 1988 to 39 +/- 12 in 1997), and the mean PaO(2)/FIO(2) ratio increased (38 +/- 23 in 1988 to 48 +/- 36 in 1997). The primary mode of ECMO remains venoarterial; however, the use of venovenous ECMO increased from 1% to 32% over the decade. Duration of ECMO treatment increased overall, and this trend was seen for patients with and without CDH (124 +/- 67 to 141 +/- 104 hours for the non-CDH group, 161 +/- 99 to 238 +/- 141 hours for the CDH group). The number of centers reporting neonatal data to the ELSO registry increased from 52 in 1988 to a peak of 100 in 1993. In 1997, 96 centers reported data to ELSO. The average number of neonatal patients reported from each site decreased from a peak of 18 in 1991 to 9 in 1997. Mortality increased from 18% to 22%; however, when corrected for the relative increase in neonates with CDH, this trend disappeared. Diagnoses-specific mortality rates remained constant. The occurrence of intracranial hemorrhage and/or infarct also stayed constant at 16%. CONCLUSIONS: The population of neonates treated with ECMO in 1997 was very different from patients treated in the 1980s and early 1990s. They were exposed to an ever expanding group of new therapies, appeared to be healthier based on indices of gas exchange, and were cared for at centers that reported fewer cases per year. PMID- 11099586 TI - Decreased use of neonatal extracorporeal membrane oxygenation (ECMO): how new treatment modalities have affected ECMO utilization. AB - OBJECTIVE: Over the last decade, several new therapies, including high-frequency oscillatory ventilation (HFOV), exogenous surfactant therapy, and inhaled nitric oxide (iNO), have become available for the treatment of neonatal hypoxemic respiratory failure. The purpose of this retrospective study was to ascertain to what extent these modalities have impacted the use of neonatal extracorporeal membrane oxygenation (ECMO) at our institution. METHODS: Patients from 2 time periods were evaluated: May 1, 1993 to November 1, 1994 (group 1) and May 1, 1996 to November 1, 1997 (group 2). During the first time period (group 1), HFOV was not consistently used; beractant (Survanta) use for meconium aspiration syndrome (MAS), persistent pulmonary hypertension of the newborn (PPHN), and pneumonia was under investigation; and iNO was not yet available. During the second time period (group 2), HFOV and beractant treatment were considered to be standard therapies, and iNO was available to patients with oxygenation index (OI) >/=25 x 2 at least 30 minutes apart, or on compassionate use basis. Patients were included in the data collection if they met the following entry criteria: 1) OI >15 x 1 within the first 72 hours of admission; 2) EGA >/=35 weeks; 3) diagnosis of MAS, PPHN or sepsis/pneumonia; 4) <5 days of age on admission; and 5) no congenital heart disease, diaphragmatic hernia, or lethal congenital anomaly. RESULTS: Of the 49 patient in group 1, 21 (42.8%) required ECMO therapy. Of these ECMO patients, 14 (66.6%) had received diagnoses of MAS or PPHN. Only 3 of the patients that went on to ECMO received beractant before the initiation of bypass (14.3%). All ECMO patients in group 1 would have met criteria for iNO had it been available. Of all patients in group 1, 18 (36.7%) were treated with HFOV, and 13 (26.5%) received beractant. Of the 47 patients in group 2, only 13 (27.7%) required ECMO therapy (compared with group 1). Of these ECMO patients, only 5 (38.5%) had diagnoses of MAS or PPHN, with the majority of patients (61.5%) requiring ECMO for sepsis/pneumonia, with significant cardiovascular compromise. Only 5 of these ECMO patients, all outborn, did not receive iNO before cannulation because of the severity of their clinical status on admission. Of all patients in group 2, 41 (87.2%) were treated with HFOV (compared with group 1), 42 (89.3%) received beractant (compared with group 1), and 18 (44.7%) received iNO. CONCLUSIONS: The results indicate that ECMO was used less frequently when HFOV, beractant and iNO was more commonly used. The differences in treatment modalities used and subsequent use of ECMO were statistically significant. We speculate that, in this patient population, the diagnostic composition of neonatal ECMO patients has changed over time. PMID- 11099587 TI - Outpatient treatment of moderate croup with dexamethasone: intramuscular versus oral dosing. AB - OBJECTIVE: Steroid use for the treatment of croup has been supported by several studies, although few have addressed the use of oral dexamethasone for outpatient management. The efficacy of oral (PO) versus intramuscular (IM) dosing of dexamethasone in the outpatient treatment of moderate croup are compared in this study. METHODS: Patients between the ages of 3 months and 12 years with moderate croup (history or presence of stridor, cyanosis, or retractions) were eligible for enrollment in this single-blind, prospective study. Patients were randomized to receive a single dose (0.6 mg/kg, maximum 8 mg) of IM or PO dexamethasone. Parents were contacted by phone to assess resolution of symptoms and need for further evaluation. RESULTS: Two hundred seventy-seven patients were enrolled (median age: 2.1 years). One hundred thirty-nine patients received IM dexamethasone, and 138 received PO. At phone follow-up, 141 (51%) had total resolution of symptoms (75 in IM, 66 in PO). Eighty patients (29%) returned for further evaluation (45 in IM, 35 in PO). Twenty-three (8%) received either more steroids, racemic epinephrine, or admission (11 in IM, 12 in PO). CONCLUSION: No statistically significant difference was found in the need for subsequent interventions after a single dose of either IM or PO dexamethasone. A single PO dose of dexamethasone can be effectively and safely used for the outpatient treatment of moderate croup. PMID- 11099588 TI - Infantile colic: crying time reduction with a whey hydrolysate: A double-blind, randomized, placebo-controlled trial. AB - OBJECTIVE: To determine the effectiveness of whey hydrolysate formula in the treatment of infantile colic in a primary care setting in the Netherlands. STUDY DESIGN: Randomized, double-blind, parallel trial with a 1-week qualification period and a 1-week intervention period. Participants. Forty-three healthy, thriving, formula-fed infants, <6 months old, crying >3 hours per day on at least 3 days per week. Infants were randomized to whey hydrolysate formula (n = 23) or standard formula (n = 20). MAIN OUTCOME MEASURE: Difference in duration of crying (minutes per day) between qualification week and intervention week. RESULTS: Analysis according to the intention to treat principle showed a difference in the decrease of crying duration of 63 minutes per day [95% confidence interval: 1-127 minutes per day] in favor of the whey hydrolysate formula. Five infants did not complete the trial. The scope of the study was not sufficient to expect significant differences in the subgroup analyses. CONCLUSIONS: An extensively hydrolyzed whey formula is effective in reducing the duration of crying in a primary care setting. PMID- 11099589 TI - Infant feeding mode affects early growth and body composition. AB - BACKGROUND: Differences in the growth pattern of breastfed (BF) and formula-fed (FF) infants are well-recognized and have been attributed to differences in nutrient intake. However, the impact of qualitative and quantitative differences in nutrient intake on the body composition of BF and FF infants has been unclear. Furthermore, it is unknown whether putative differences in body composition persist beyond weaning. DESIGN: Prospective cohort study. METHODS: Repeated anthropometric and body composition measurements were performed on 40 BF and 36 FF infants at 0.5, 3, 6, 9, 12, 18, and 24 months of age. A multicomponent body composition model based on total body water by deuterium dilution, total body potassium by whole body counting, and bone mineral content by dual-energy x-ray absorptiometry was used to estimate fat-free mass (FFM) and fat mass (FM). Independent measurements of FFM and FM were made using total body electrical conductivity and dual-energy x-ray absorptiometry. By design, infants were either exclusively BF or FF from birth to 4 months of age; thereafter, the feeding mode was at the discretion of the parents. Infant food intake was measured at 3, 6, 12, and 24 months of age using 3-day weighed-intake records. Data were analyzed by repeated measures analysis of variance. RESULTS: Weight velocity was higher in FF than BF infants age 3 to 6 months, and higher in FF than BF girls 6 to 9 months of age. Adjusted for gender and baseline values, BF infants had lower total body water at 3 months, lower total body potassium at 3 to 24 months, and lower bone mineral content at 12 months. The multicomponent model indicated that FFM was lower in BF than FF infants at 3 months, and FM and %FM were higher in BF than FF infants at 3 and 6 months (boys only). Total body electric conductivity confirmed lower FFM in BF than FF infants at 3 months, as well as at 6 and 9 months; FM and %FM were higher in BF than FF at 3 and 6 months, and 9 months (boys only). Intakes of energy, protein, fat, and carbohydrate were lower in BF than FF infants at 3 and 6 months, and were positively correlated with weight gain and FFM gain, but not FM gain. No differences in nutrient intakes were observed at 12 or 24 months. CONCLUSION: Infant feeding mode is associated with differences in body composition in early infancy which do not persist into the second year of life. PMID- 11099590 TI - Do pediatricians recognize mothers with depressive symptoms? AB - OBJECTIVE: To determine whether pediatric health care providers recognize maternal depressive symptoms and to explore whether maternal, provider, and visit characteristics affect pediatric providers' ability to recognize inner-city mothers with depressive symptoms. DESIGN: A cross-sectional study was conducted at a hospital-based, inner-city, general pediatric clinic. Two groups of participants completed questionnaires, each unaware of the other's responses: 1) mothers who brought their children ages 6 months to 3 years for health care maintenance or a minor acute illness and 2) pediatric health care providers (attending pediatricians, pediatric trainees, and nurse practitioners). The mothers' questionnaire consisted of sociodemographic items and a self administered assessment of depressive symptoms using the Psychiatric Symptom Index (PSI). Pediatric providers assessed child, maternal, and family functioning and documented maternal depressive symptoms. Criteria for positive identification of a mother by the pediatric health care provider were met if the provider reported one or more maternal symptoms (from a 10-item list of depressive symptoms), a rating of 4 or less on a scale of functioning, a yes response to the question of whether the mother was acting depressed, or a response that the mother was somewhat to very likely to receive a diagnosis of depression. RESULTS: Of 338 mothers who completed the questionnaire, 214 (63%) were assessed by 1 of 60 pediatric providers. Seventy-seven percent of surveys were completed by the child's designated pediatric provider. The mean visit length was 23 minutes. Mothers primarily were single, were black or Hispanic, and had a mean age of 26 years (15-45 years). Almost 25% of mothers were living alone with their children. Eighty-six (40%) mothers scored >/=20 on the PSI, representing high symptom levels. Of these, 25 were identified by pediatric providers (sensitivity = 29%). A total of 104 of 128 mothers with a PSI score <20 were identified as such by providers (specificity = 81%). Pediatric providers were more likely to identify mothers who were <30 years old, living alone, and on public assistance. Also, mothers who were assessed by the child's own primary provider or by an attending pediatrician were more likely to be identified accurately than were mothers whose children were seen by a pediatric trainee or a nurse practitioner. CONCLUSIONS: Pediatric health care providers did not recognize most mothers with high levels of self-reported depressive symptoms. Pediatricians may benefit from asking directly about maternal functioning or by using a structured screening tool to identify mothers who are at risk for developing depressive symptoms. In addition, training pediatric providers to identify mothers with depressive symptoms may be beneficial. PMID- 11099591 TI - Teething and tooth eruption in infants: A cohort study. AB - OBJECTIVE: Many symptoms are attributed to teething in infants. There is little evidence to support these beliefs, despite their implications for clinical management. We investigated relationships between tooth eruption, fever, and teething symptoms. METHODS: Prospective cohort study. PARTICIPANTS: Twenty-one children 6 to 24 months old attending 3 suburban long-day care centers >/=3 days/week. Measures. 1) Daily temperature recording and examination of alveolar ridges for tooth eruption (dental therapist). 2) Daily questionnaires-symptoms over preceding 24 hours (staff and parents independently). 3) Final questionnaire beliefs/experiences related to teething (parents). Definitions. Eruption day-the first day a tooth could be seen or felt. Non-toothdays-more than 28 days clear of any eruption day. Toothdays-the 5 days preceding eruption days. RESULTS: Data were collected for 236 toothdays and 895 non-toothdays pertaining to 90 teeth. Child temperatures were similar on toothdays and non-toothdays (36.21 vs 36.18, paired t test). Logistic regression adjusted for age did not show an association between toothdays and temperature (odds ratio [OR] = 1.35, 95% confidence interval [CI] = 0.80, 2.27 for high fever; OR = 1.34, 95% CI = 0.48, 3.77 for low fever). Logistic regression models allowing for within-child cluster effects and age were fitted to daily staff and parent reports of mood, wellness/illness, drooling/dribbling, sleep, diarrhea, strong diapers, red cheeks, and rashes/flushing. Only parent-reported (but not staff-reported) loose stools were significantly associated with tooth eruption (OR = 1.86, 95% CI = 1. 26, 2.73). When the toothday definition was varied to 10 days preceding or 5 days surrounding tooth eruption, this single significant association was no longer apparent (OR = 1.42, 95% CI = 0.98, 2.05 and OR = 1.47, 95% CI = 0.97, 2.21, respectively). All parents retrospectively reported that their own children had suffered a range of teething symptoms. CONCLUSIONS: This study did not confirm the expected strong associations between tooth eruption and a range of teething symptoms in children 6 to 30 months old, although we cannot rule out the possibility that weak associations may exist (Type II error). These findings contrast with strong parent and professional beliefs to the contrary. Such beliefs may preclude optimal management of common patterns of illness and behavior in young children.teething, infants, symptoms, tooth eruption, illness. PMID- 11099592 TI - Maternal perceptions of overweight preschool children. AB - CONTEXT: Childhood obesity is a major public health problem, and prevention efforts should begin early in life and involve parents. OBJECTIVE: To determine what factors are associated with mothers' failure to perceive when their preschool children are overweight. DESIGN: Cross-sectional survey. SETTINGS: Offices of private pediatricians and clinics of the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC). PARTICIPANTS: Six hundred twenty-two mothers with children 23 to 60 months of age. MAIN OUTCOME MEASURES: Maternal demographic variables, maternal self-reported height and weight, and children's measured height and weight. Mothers were asked whether they considered themselves or their children overweight. RESULTS: Forty-five percent of mothers had low education (high school degree or less) and 55% had high education (some college or more). Obesity (body mass index: >/=30 kg/m(2)) was more common in the low education group of mothers (30% vs 17%), and their children tended to be more overweight (weight-for-height percentile: >/=90th; 19% vs 14%). Ninety-five percent of obese mothers believed that they were overweight, with no difference between education groups. However, 79% of mothers failed to perceive their overweight child as overweight. Among the 99 mothers with overweight children, low maternal education was associated with a failure to perceive their children as overweight after adjusting for low family income (3 days old need to effectively treat Gram negative pathogens, particularly Pseudomonas sp., because these organisms, although less frequent, are strongly associated with fulminant late-onset sepsis in the NICU. Avoiding empiric vancomycin therapy seemed to be a reasonable approach to late-onset sepsis, because of the very low frequency of fulminant sepsis caused by coagulase-negative staphylococci. PMID- 11099594 TI - Pediatric office emergencies and emergency preparedness in a small rural state. AB - OBJECTIVE: Although the frequency of pediatric office medical emergencies has been investigated in a retrospective manner, there have been no prospective studies. We examined how often pediatricians in a small rural state encountered medical emergencies in the office setting. This study included an in-office educational program and the donation of resuscitation equipment to study participants. DESIGN AND INTERVENTION: Thirty-eight of the 40 active primary care pediatric practices in the state of Vermont participated in this study. Thirty seven sites were surveyed retrospectively regarding office preparedness for emergencies and frequency of office emergencies. At each practice site, an educational session was provided and an office resuscitation kit was donated. Thirty-seven sites were followed prospectively for a 12-month period evaluating the incidence of office medical emergencies and the adequacy of the donated resuscitation kit. RESULTS: Three hundred twenty-seven individuals from 38 Vermont pediatric practice sites participated. Forty-nine percent had basic life support training and 26% had pediatric advanced life support training. Sixty seven percent of practices had a plan for office medical emergencies. Forty-six percent of practices had called local emergency medical services providers to their offices in the past year. Emergency preparedness ranged from a high of 95% of sites with oxygen to a low of 27% of sites with intraosseous needles. The estimate of the frequency of medical emergencies was.9 (standard deviation =.8) per office in the previous 12 months. In the 12-month study, there were 28 medical emergencies reported, averaging.8 (standard deviation = 1.5) emergencies per office per year. Sixty-five percent of participating sites had no emergencies in the study. Of the emergencies reported, 75% were respiratory in origin. The donated resuscitation kits proved sufficient for all of the emergencies reported. CONCLUSIONS: Serious medical emergencies are rare events in the primary care pediatric office, occurring less than once per office per year. The most common emergency situations encountered are respiratory. All of the emergencies in this study were managed effectively using a simple and relatively inexpensive resuscitation kit. We provided an emergency preparedness program for pediatric practices in a small rural state. PMID- 11099595 TI - Longitudinal neurologic follow-up in neonatal intensive care unit survivors with various neonatal morbidities. AB - OBJECTIVE: The purpose of this prospective longitudinal study was to examine neurocognitive and school performance outcomes of low birth weight infants with reference to neonatal morbidity and socioeconomic status. We further evaluated the cognition and school performance based on their neurologic status at the time of assessment. METHODS: One hundred eighty-eight children (39 healthy full-term and 149 preterm infants) were classified into 4 subgroups based on their neonatal medical status: healthy, sick (without neurologic complications), small for gestational age, and neurologically compromised infants. Neurologic status was classified as normal, suspect, or abnormal at hospital discharge, 18 months, 30 months, 4 years, and 8 years of age. Socioeconomic status, cognitive, and school performances were assessed. RESULTS: Neurologically, both full-term and healthy preterm groups did well during the 8-year period. There were significant fluctuations between suspect and abnormal neurologic classifications among the 3 preterm groups with neonatal complications. Preterms with neurologic abnormality during the neonatal period did the poorest with 45% of the group remaining abnormal at 8 years of age. Children who were neurologically normal had higher cognitive scores at ages 4 and 8 than those categorized as suspect or abnormal. Preterm infants with neurologic abnormality required significantly more academic resources in the school. Reading and math achievement scores were the lowest for the preterm groups classified as neurologically suspect or abnormal. CONCLUSIONS: Neonatal morbidities exert a significant impact in neurologic outcomes among preterm children during the 8 years of assessment. Compromised neurologic status adversely affects cognitive and school performances. Neonatal medical status is an important variable indicating neurocognitive and school performance outcomes in low birth weight infants. PMID- 11099596 TI - Predictors of asthma three years after hospital admission for wheezing in infancy. AB - OBJECTIVE: To evaluate the influence of early antiinflammatory therapy in the development of asthma 3 years after hospitalization for wheezing in infancy. In addition, the effects of allergic sensitization and respiratory syncytial virus (RSV) infection on the development of asthma were investigated. DESIGN AND SETTING: A randomized, controlled follow-up study in a university hospital that provides primary hospital care for all pediatric patients in a defined area. PATIENTS: Eighty-nine infants under 2 years of age who had been hospitalized for infection associated with wheezing and followed up for 3 years. INTERVENTION: Early antiinflammatory therapy was given for 16 weeks; 29 patients received cromolyn sodium and 31 received budesonide. Twenty-nine control patients received no therapy. OUTCOME MEASURES: Clinical diagnosis of current asthma, defined as having at least 3 episodes of physician-diagnosed wheezing and either a wheezing episode during the preceding year or ongoing antiinflammatory medication for asthma. RESULTS: Fourteen (48%) patients in the former cromolyn group, 15 (48%) in the former budesonide group, and 16 (55%) in the control group had current asthma. The significant predictors of asthma were age over 12 months (risk ratio [RR] 4.1; 95% confidence interval [CI] = 1.59-10.35), history of wheezing (RR 6.8; CI = 1.35-34.43), and atopic dermatitis on study entry (RR 3.4; CI = 1.17 9.39). Skin prick test positivity at the age of 16 months significantly predicted asthma (RR 9.5; CI = 2.45-36.72). In addition, all of the 18 (20%) children sensitized with furred pet developed asthma. RSV identification (RR 0.3; CI = 0.08-0.80) and early furred pet contact at home (RR 0.3; CI 0.10-0.79) were associated with the decreased occurrence of asthma. CONCLUSIONS: Antiinflammatory therapy for 4 months has no influence on the occurrence of asthma 3 years after wheezing in infancy. Early sensitization to indoor allergens, especially to pets, and atopic dermatitis predict subsequent development of asthma. RSV infection in wheezing infants may have a better outcome than other infections. PMID- 11099597 TI - Trends in intussusception-associated hospitalizations and deaths among US infants. AB - CONTEXT: The newly licensed tetravalent rhesus-human reassortant rotavirus vaccine has been withdrawn following reports of intussusception among vaccinated infants. OBJECTIVE: To describe the epidemiology of intussusception-associated hospitalizations and deaths among US infants. DESIGN: This retrospective cohort study examined hospital discharge data from the National Hospital Discharge Survey for 1988-1997, Indian Health Service (IHS) for 1980-1997, California for 1990-1997, Indiana for 1994-1998, Georgia for 1997-1998, and MarketScan for 1993 1996, and mortality data from the national multiple cause-of-death data for 1979 1997 and linked birth/infant death data for 1995-1997. PATIENTS: Infants (<1 year old) with an International Classification of Diseases, Ninth Revision, Clinical Modification code for intussusception (560.0) listed on their hospital discharge or mortality record, respectively. RESULTS: During 1994-1996, annual rates for intussusception-associated infant hospitalization varied among the data sets, being lowest for the IHS (18 per 100 000; 95% confidence interval [CI] = 9-35 per 100 000) and greatest for the National Hospital Discharge Survey (56 per 100 000; 95% CI = 33-79 per 100 000) data sets. Rates among IHS infants declined from 87 per 100 000 during 1980-1982 to 12 per 100 000 during 1995-1997 (relative risk =7.6, 95% CI = 3.2-18.2). Intussusception-associated hospitalizations were uncommon in the first 2 months of life, peaked from 5 to 7 months old, and showed no consistent seasonality. Intussusception-associated infant mortality rates declined from 6.4 per 1 000 000 live births during 1979-1981 to 2.3 per 1 000 000 live births during 1995-1997 (relative risk = 2.8, 95% CI = 1.8-4.3). Infants whose mothers were <20 years old, nonwhite, unmarried, and had an education level below grade 12 years were at an increased risk for intussusception-associated death. CONCLUSIONS: Intussusception-associated hospitalization rates varied among the data sets and decreased substantially over time in the IHS data. Although intussusception-associated infant deaths in the United States have declined substantially over the past 2 decades, some deaths seem to be related to reduced access to, or delays in seeking, health care and are potentially preventable.intussusception, hospitalizations, deaths, risk factors, infants. PMID- 11099598 TI - Influence of parental gender and self-reported health and illness on parent reported child health. AB - BACKGROUND: Although there is clear evidence of the influence of parental factors on child health outcomes, the influence of parental perceptions of their health and illness on the reporting of child health remains primarily unknown. OBJECTIVES: To examine relationships between parents' reporting of their own health and illness with the reporting of their children's health and illness. METHOD: We surveyed parents of a representative population-based sample of children aged 5 to 18 years. One parent of each child completed a written questionnaire including the Child Health Questionnaire, a subjective measure of functional health and well-being, and an assessment of self-reported parental health and illness. Logistic regression models were used to examine relationships between parent and child health and illness. MAIN RESULTS: 5340 parents responded (86% mothers, 14% fathers). After adjusting for confounding effects, parents self reporting poor health had increased odds of reporting their children with poor health (odds ratio: 7.5), although the effect was modified by parent gender. There were increased odds of mothers with self-reporting poor global health reporting their children with poor global health and illness (odds ratio: 9.0 and 2.5, respectively) that were not observed for fathers. CONCLUSIONS: A mother's self-reported health is strongly associated with her reporting of her child's health; this was not observed for fathers. These results suggest that parental gender should be considered as a mediating factor in the reporting of child health. PMID- 11099599 TI - Improving Preschoolers' self-regulation of energy intake. AB - BACKGROUND: Children exhibit individual differences in their ability to self regulate energy intake. Feeding strategies that focus on external signals, like the time of day or amount of food left on a plate, tend to diminish children's ability to respond to internal cues of hunger and fullness. OBJECTIVES: We investigated whether children could be taught to focus on internal cues of hunger and satiety, and consequently improve their self-regulation of energy intake. We explored whether parents' eating behaviors and adiposity were related to their children's self-regulation skills and adiposity. DESIGN: In a pretest and posttest design, preschoolers participated in single-meal protocols to assess their individual ability to self-regulate food intake. During a 6-week intervention period, children took part in individual and group activities designed to help them recognize internal cues. Parents completed questionnaires regarding adult dietary restraint and disinhibition. RESULTS: At baseline, we found a large individual variability in children's regulation: some children overate, some regulated accurately, and others underate. At baseline, children's eating related to their adiposity and to mothers' disinhibition: heavier children and children whose mothers' reported difficulty controlling food intake showed less evidence of self-regulation. Both overeaters and undereaters responded to the intervention, improving their ability to self-regulate, and children's eating was no longer significantly related to mother's eating. CONCLUSIONS: Children's disregulated energy intake is related to mothers' weight status and mothers' perceived control over eating. Cues can be provided that help children to focus on internal signals and improve their ability to self-regulate energy intake. PMID- 11099600 TI - Do obese inner-city children with asthma have more symptoms than nonobese children with asthma? AB - OBJECTIVE: To test whether obesity is associated with decreased peak expiratory flow rates (PEFR), increased asthma symptoms, and increased health service use. DESIGN/METHODS: Secondary analysis of data from a cross-sectional convenience sample. SETTING: Emergency departments (EDs) and primary care clinics in 8 inner city areas in 7 cities. PARTICIPANTS: One thousand three hundred twenty-two children aged 4 to 9 years with asthma. MEASURES: Obesity was defined as a body mass index (BMI, weight/height(2)) >95th percentile. Nonobese children were those with a BMI between the 5th and 95th percentile. Underweight children with a BMI <5th percentile were eliminated from the study. Demographic and anthropometric data were obtained during a baseline interview with the primary caretaker and the child. Symptoms, health service use data and measurements of PEFR were obtained by parental report during the baseline interview and at 3-month intervals by telephone interview over the following 9-month period. RESULTS: Obese (n = 249) and nonobese (n = 1073) children did not differ in terms of age, gender, family income, passive smoke exposure, caretaker's mental health, and skin test reactivity to indoor allergens. Obese children were more often Latino (28% vs 17%) and, in the 3 months before the baseline interview, were more likely to have used oral steroids (30% vs 24%). There were no differences between groups in terms of baseline PEFR scores. During the 9 months after baseline assessment, the obese group had a higher mean number of days of wheeze per 2-week period (4.0 vs 3.4), and a greater proportion of obese individuals had unscheduled ED visits (39% vs 31%). There were no differences between the groups in terms of frequency of hospitalization, or in nocturnal awakening. CONCLUSIONS: In our sample of inner-city children with asthma, obese children used more medicine, wheezed more, and a greater proportion had unscheduled ED visits than the nonobese children. PMID- 11099601 TI - Bronchial hyperresponsiveness before and after the diagnosis of bronchial asthma in children. AB - OBJECTIVE: To assess at what age bronchial hyperresponsiveness (BHR) is acquired in children with asthma. BACKGROUND: A relationship between BHR and infantile wheezing diseases has been reported. Infants with a genetic predisposition to atopy are more likely to wheeze with respiratory viral infection or bronchiolitis, and it is suspected that the continued BHR after the first attack of asthma may be induced or triggered by some viral infections. Also, recent studies have reported the existence of atopic and BHR-related genes. However, whether BHR is congenital or acquired after asthma attacks, and when BHR in children with asthma is established or acquired remain unclear. METHODS: We performed methacholine inhalation challenge using a transcutaneous oxygen pressure (tcPO(2)) monitoring system in 205 children without asthma from 6 months to 6 years of age. During follow-up, 18 of these participants were diagnosed with asthma (group N-A). This group and 15 age-matched children without asthma (group N-N) were tested twice using methacholine inhalation challenge. For comparison, 39 age-matched atopic-type asthmatic children (group A-A) were also given the inhalation challenge twice. Methacholine inhalation challenge using a tcPO(2) monitoring system was performed while the participants were asleep in the supine position. Sequential doses of inhaled methacholine delivered by oxygen mask were doubled until a 10% decrease in tcPO(2) from the baseline was reached. The cumulative dose of methacholine at the inflection point of tcPO(2) (minimal dose of methacholine [Dmin]-PO(2)) was considered to represent BHR. RESULTS: In groups N-N and A-A, there was no difference in Dmin-PO(2) between the first and second challenge. However, the Dmin-PO(2) in group N-A significantly decreased from the first challenge to the second challenge. There was no significant difference between the Dmin-PO(2) in group N-N and the first Dmin-PO(2) in group N-A; or between the Dmin-PO(2) in group A-A and the second Dmin-PO(2) in group N-A. CONCLUSIONS: These data suggest that BHR in many infants with asthma is acquired after several asthma attacks.bronchial hyperresponsiveness, childhood asthma, methacholine inhalation challenge, transcutaneous oxygen pressure. PMID- 11099602 TI - Role of carbon monoxide and nitric oxide in newborn infants with postasphyxial hypoxic-ischemic encephalopathy. AB - OBJECTIVE: To investigate the role of carbon monoxide (CO) and nitric oxide (NO) in the pathogenesis of neonatal hypoxic-ischemic encephalopathy (HIE). METHODS: Plasma CO and NO levels were studied in 33 asphyxiated term neonates, and 30 healthy neonates served as controls. RESULTS: Among the 33 asphyxiated term neonates, plasma CO and NO levels in 28 neonates with HIE were significantly higher than those in the 5 infants without HIE and in the normal controls. The plasma CO and NO levels in the newborn infants with HIE stage 3 were found to be significantly higher than those in the neonates with HIE stage 1 and 2. Moreover, plasma CO and NO levels were significantly increased in neonates with brain damage and adverse outcome as compared with those in patients with normal neuroimaging and normal outcome. CONCLUSION: Plasma CO and NO levels after perinatal asphyxia are related to the severity of neonatal HIE, brain damage, and neurologic outcome. The present study suggests that CO and NO might play important roles in the pathogenesis of neonatal HIE. PMID- 11099603 TI - Lung elastic tissue maturation and perturbations during the evolution of chronic lung disease. AB - BACKGROUND: Infants <30 weeks' gestation have difficulty maintaining adequate functional residual capacity after the first week of life without positive end expiratory pressure. We hypothesized that this is caused, in part, by increased lung elastic recoil. Our aims were to quantitate parenchymal elastic tissue during normal fetal development and in infants born at 23 to 30 weeks' gestation with prolonged survival at risk for chronic lung disease (CLD). METHODS: The controls were 22 to 42 weeks' gestation (n = 71), received ventilator care, and died within 48 hours of birth, plus 7 term infants who died at 43 to 50 weeks' postconceptional age from nonpulmonary causes. Infants who were 23 to 30 weeks' gestation, at risk for CLD, and who lived 5 to 59 days (n = 44), were separated into groups based on respiratory score (SCORE; The integrated area under the curve of the average daily fraction of inspired oxygen x mean airway pressure (cm H(2)O) over the number of days lived). The SCORE groups, <20, 21 to 69 and 70 to 200, related clinically to mild to severe lung disease. The lungs were tracheally perfused and formalin-fixed and total lung volume (TLV) was measured by water displacement. The paraffin-embedded lung blocks were stained with Miller's elastic stain. The parenchyma and parenchymal elastic tissue were point-counted. The absolute elastic tissue was calculated by multiplying TLV by the parenchymal and elastic fractions. Septal width, alveoli and alveolar duct diameters, and internal surface area (ISA) were also measured. RESULTS: In the controls, the volume density of parenchymal elastic tissue and absolute quantity of elastic tissue increased progressively from 22 to 50 weeks. In infants with CLD and SCORE >/=20, the volume density and absolute quantity of elastic tissue increased significantly. Mean absolute elastic tissue in the 20 to 69 group was 0.76 +/- 0.20 cm(3) greater than in the <20 group (0.46 +/- 0.10 cm(3)) who were similar to the controls, and the 70 to 200 group was 1.32 +/- 0.56 cm(3) greater than the 20 to 69 group. Elastic tissue for infants at risk for CLD, as a percent of predicted for same-age controls, rose linearly with increasing SCORE (r = 0.73; r(2) = 0.55). Control TLV and ISA were linearly related to age. Thirty-nine of the 44 CLD-risk infants had TLVs greater than controls. However, 77% with SCORE 20 to 200 had ISAs less than or equal to the control 95% confidence interval. Control septal width decreased sharply from 23 to 30 weeks, then gradually decreased to term. All infants with SCORE 70 to 200 and 80% of those with SCORE 20 to 69 had widths more than the control 95% confidence interval. Control alveolar and duct diameters doubled from 23 to 50 weeks and were significantly greater in infants with SCORES 20 to 200. DISCUSSION: Lung elastic tissue maturation is tightly controlled during fetal development. With increasing SCORE, elastic tissue increased >200%, accounting, in part, for the positive end expiratory pressure needed to maintain end-expiratory lung volume in infants at risk for CLD. Saccule and duct diameters more than doubled, and septa thickened significantly in CLD. We propose the following sequence to be operative in CLD: at birth, the preterm infant (/=38 degrees C to an urban pediatric emergency department. Positive cultures and times to detection were noted. Patients were categorized as being at high risk for serious bacterial illness (SBI) based on clinical and laboratory criteria. RESULTS: Of the 3166 febrile infants seen in the emergency department during the study, 2733 had blood (86%), 2517 had urine (80%), and 2361 had CSF (75%) specimens obtained for culture, and 2190 had all 3 cultures (69%) sent. There were 224 positive cultures in 214 patients; of these, 191 had all 3 cultures (89%) sent. Subsequent analyses were confined to those who had all 3 cultures sent. The detected rate of SBI was 8.7% (191/2190). There were 28 positive blood cultures (1. 3%), 165 positive urine cultures (7.5%), and 8 positive CSF cultures (.4%). Median time to detection of positive cultures was 16 hours for blood, 16 hours for urine, and 18 hours for CSF. Four blood cultures (.1%), 20 urine cultures (.9%), and 0 CSF cultures were noted to have growth of a pathogen >24 hours after the specimen was obtained. All 4 blood cultures and 17 of 20 urine cultures with growth noted after 24 hours occurred among high-risk patients. CONCLUSIONS: The risk of identifying SBI after 24 hours is 1.1% among all 28- to 90-day-old febrile infants and.3% in low-risk infants. PMID- 11099618 TI - Missed opportunities for preventing tuberculosis among children younger than five years of age. AB - OBJECTIVES: Childhood tuberculosis (TB) is an important indicator of public health success in interrupting and preventing TB transmission. To determine the frequency and types of missed opportunities for preventing TB among children <5 years of age. METHODS: We collected data from the public health records of child TB cases and their adult source cases. These children were from health jurisdictions where TB case rates in children were higher than the California average for this age group. RESULTS: We reviewed the records for 165 children reported with TB (20% confirmed by culture). These children were evaluated for TB because of signs or symptoms of illness (32%), a contact investigation (26%), screening (22%), a source case investigation (4%), and unknown reasons (16%). Excluding 4 children infected by Mycobacterium bovis, only 59 of 161 children (37%) had a source case found. Children found in a contact investigation, born in the United States, <1 year of age, or who were black were more likely to have a source case found than children who did not have one of these characteristics. Of 43 children found in a contact investigation, improvements in contact investigations may have prevented TB in 17 of these children (40%). Among the 43 adult source cases, factors that may have facilitated transmission include delayed reporting in 23%, a delayed contact investigation in 21%, and delayed or nondocumented bacteriologic sputum conversion in 42% of culture-positive cases. CONCLUSIONS: Important missed opportunities to prevent TB in children include the failure to find and appropriately manage adult source cases and failure to completely evaluate and properly treat children exposed to TB. Improvements in case detection, case management, and contact investigations are necessary to eliminate TB in children. PMID- 11099620 TI - Morbidity among human immunodeficiency virus-1-infected and -uninfected African children. AB - OBJECTIVE: To assess patterns of morbidity and associated factors in late infancy and early childhood among human immunodeficiency virus (HIV)-infected and uninfected African children. DESIGN: Prospective study. SETTING: The Queen Elizabeth Central Hospital, Blantyre, Malawi. PARTICIPANTS: Children with known HIV status from an earlier perinatal intervention trial were enrolled during the first year of life and followed to approximately 36 months of age. OUTCOME MEASURES: Morbidity and mortality information was collected every 3 months by a questionnaire. A physical examination was conducted every 6 months. Blood to determine CD4(+) values was also collected. Age-adjusted and Kaplan-Meier analyses were performed to compare rates of morbidity and mortality among infected and uninfected children. RESULTS: Overall, 808 children (190 HIV infected, 499 HIV-uninfected but born to infected mothers, and 119 born to HIV uninfected mothers) were included in this study. Of these, 109 died during a median follow-up of 18 months. Rates of childhood immunizations were high among all children (eg, lowest was measles vaccination [87%] among HIV-infected children). Age-adjusted morbidity rates were significantly higher among HIV infected than among HIV-uninfected children. HIV-infected children were more immunosuppressed than were uninfected children. By 3 years of age, 89% of the infected children died, 10% were in HIV disease category B or C, and only approximately 1% were without HIV symptoms. Among HIV-infected children, median survival after the first occurrence of acquired immunodeficiency syndrome-related conditions, such as splenomegaly, oral thrush, and developmental delay, was <10 months. These same conditions, in addition to frequent bouts of fever, were the main morbidity predictors of mortality. CONCLUSIONS: The frequency of diseases was high, and progression from asymptomatic or symptomatic HIV disease to death was rapid. Management strategies that effectively reduce morbidity for HIV infected children are needed. PMID- 11099619 TI - Predicting HIV disease progression in children using measures of neuropsychological and neurological functioning. Pediatric AIDS clinical trials 152 study team. AB - BACKGROUND: Neuropsychological testing and 2 measures of neurological status, cortical atrophy, and motor dysfunction were assessed for their usefulness in predicting human immunodeficiency virus (HIV) disease progression in infants, children, and adolescents who participated in Pediatric AIDS Clinical Trials Group Protocol 152 (PACTG 152). METHODS: A cohort of 722 antiretroviral therapy naive children with symptomatic HIV infection were assessed at study entry and at later intervals. Assessments included neurodevelopmental testing, neuroradiologic imaging, and neurological examination of motor function. CD4 cell count and plasma RNA viral load also were measured. RESULTS: Children with the lowest neuropsychological functioning (IQ < 70) at baseline had the highest risk for later HIV disease progression (56%), compared with those with borderline/low (IQ = 70-89) functioning (26%), or with average or above (IQ > 90) functioning (18%). This was also true of week 48 neuropsychological functioning. Motor dysfunction (especially reduced muscle mass) at entry also predicted disease progression. Furthermore, motor dysfunction and week 48 neuropsychological functioning provided predictive information beyond that obtainable from surrogate markers of HIV disease status (eg, CD4 count, HIV RNA level). Children with cortical atrophy also were at higher risk for later disease progression, but when CD4 count and RNA viral load were known, cortical atrophy information provided no additional predictive information. CONCLUSIONS: Measures of neuropsychological and motor function status provide unique information regarding pediatric HIV disease progression. As such, these findings have important implications for predicting long-term outcomes (eg, longevity) in pediatric patients. PMID- 11099621 TI - Hepatitis B immunization coverage among Vietnamese-American children 3 to 18 years old. AB - OBJECTIVE: Persons with chronic hepatitis B virus (HBV) infection are at increased risk of chronic hepatitis, cirrhosis, and liver cancer. Although HBV infection is relatively uncommon in the United States, the disease is endemic in persons born in Southeast Asia, including Vietnamese-Americans. Current US infant immunization recommendations and state-mandated school-entry programs have left many nontargeted age-cohorts unvaccinated and at risk of infection. To assess the need for catch-up hepatitis B immunizations, this study reports the hepatitis B immunization rates of Vietnamese-American children 3 to 18 years old living in the metropolitan areas of Houston and Dallas, Texas, and the Washington, DC, area. DESIGN: We conducted 1508 telephone interviews with random samples of Vietnamese households in each of the 3 study sites. We asked for hepatitis B immunization dates for a randomly selected child in each household. Attempts were made to verify immunization dates through direct contact with each child's providers. Low and high estimates of coverage were calculated using reports from providers when reached (n = 720) and for the entire sample (n = 1508). RESULTS: Rates of having 3 hepatitis B vaccinations ranged from 13.6% (entire sample) to 24.1% (provider reports, Dallas), 10. 3% to 26.4% (Houston), and 18.1% to 37.8% (Washington, DC). Children living in the Texas sites, older children, children whose families had lived in the United States for a longer time, and children whose provider was Vietnamese or who had an institutional provider were less likely to have been immunized. The odds of being immunized were greater, however, for children who had had at least 1 diphtheria, tetanus toxoid, and pertussis shot, and whose parents had heard about HBV infection, and were married. CONCLUSIONS: The low rates of hepatitis B vaccine coverage among children and adolescents portend a generation which, too old to benefit from infant programs and school entry laws, will grow into adulthood without the protection of immunization. Increased efforts are needed to design successful catch-up campaigns for this population. PMID- 11099622 TI - Elevated blood lead levels and blood lead screening among US children aged one to five years: 1988-1994. AB - OBJECTIVES: To estimate the proportion of children 1 to 5 years of age who received blood lead testing during 1988-1994 and to assess whether predictors of testing coincided with predictors of elevated blood lead levels. DESIGN: Cross sectional analysis of data from the Third National Health and Nutrition Examination Survey. Participants. US children 1 to 5 years of age. Outcome Measures. Prevalence of blood lead testing and elevated blood lead levels among children 1 to 5 years of age and odds ratios for factors predicting blood lead testing and elevated blood lead levels. RESULTS: Overall, 6.3% had elevated blood lead levels and 10.2% had undergone previous blood lead tests. Being of minority race/ethnicity, living in an older home, residing in the Northeast or Midwest regions of the United States, being on Medicaid, having a head of household with <12 years of education, and having a history of anemia were significant factors in both models. Additional independent risk factors for an elevated blood lead level included being sampled in phase 1 of the survey, being 1 to 2 years of age, not having a regular doctor, and being sampled during the summer months. Additional independent correlates of a previous blood lead test included having moved less than twice in one's lifetime, having a female head of household, and having parents whose home language was English. Of an estimated 564 000 children 1 to 5 years of age who had elevated blood lead levels and no previous screening test in 1993, 62% were receiving Medicaid, 40% lived in homes built before 1946, and 34% were black, non-Hispanic. CONCLUSIONS: Lead screening was more frequent among children with risk factors for lead exposure. However, among children with elevated blood lead levels, only one third had been tested previously. In 1993 an estimated 564 000 children 1 to 5 years of age had elevated blood lead levels but were never screened. Physicians should screen Medicaid-eligible children and should follow state or local health department recommendations about identifying and screening other at-risk children. In areas where no health department guidelines exist, physicians should screen all children or screen based on known risk factors. PMID- 11099623 TI - Hyperbaric oxygen therapy for cerebral palsy: two complications of treatment. AB - There is growing interest in the use of hyperbaric oxygen therapy (HBO(2)) for children with cerebral palsy. Although there is no rigorous evidence to support this management, private hyperbaric centers have been established throughout the United States and Canada. There is likely to be increasing pressure on pediatricians and other health professionals to prescribe HBO(2). We describe 2 children with cerebral palsy who suffered significant morbidity immediately after treatment with hyperbaric oxygen. Both the temporal association and pathologic findings suggest that the hyperbaric treatment is likely to have been responsible for the resulting complications. As with any new therapy, we suggest waiting for the results of a randomized, controlled trial before recommending this treatment. PMID- 11099624 TI - Hypnosis as a diagnostic modality for vocal cord dysfunction. AB - Vocal cord dysfunction (VCD) is a condition of paradoxical adduction of the vocal cords during the inspiratory phase of the respiratory cycle. VCD often presents as stridorous breathing, which may be misdiagnosed as asthma. The mismanagement of this disorder may result in unnecessary treatment and iatrogenic morbidity. An association with psychogenic factors has been reported, and a higher incidence of anxiety-related illness has been demonstrated in patients with VCD. Definitive diagnosis of VCD is made by visualization of adducted cords during an acute episode using nasopharyngeal fiber-optic laryngoscopy. Diagnosis can be problematic, because it may be difficult to reproduce an attack in a controlled setting. To maximize diagnostic yield during laryngoscopy, provocation of symptoms using methacholine, histamine, or exercise challenges have been used. We report a case of an 11-year-old boy, wherein hypnotic suggestion was used as an alternative method to achieve a diagnosis of VCD. The patient was admitted to the pediatric intensive care unit for elective fiber-optic laryngoscopy to confirm a diagnosis of VCD. The patient had a 4-year history of refractory asthma, severe gastroesophageal reflux disease (GERD) for which he had undergone a Nissen fundoplication, and suspected VCD. At 9 years of age the patient began manifesting monthly respiratory distress episodes of a severe character different from those that had been attributed to his asthma. Typically, he awoke from sleep with shortness of breath and difficulty with inhalation. He described a "neck attack" during which he felt as if the walls of his throat were "beating together." The patient was at times noted by his mother to exhibit a "suckling" behavior before onset of his respiratory distress episodes. On 4 occasions the patient became unconscious during an attack and then spontaneously regained consciousness after a few minutes. On these occasions, he was transported by ambulance to the hospital and the severe difficulty with inhalation resolved within a few minutes on treatment with oxygen and bronchodilators. Sometimes he was noted to manifest wheezing for several hours, which was responsive to bronchodilator therapy. Given the severity of the patient's disease, it was imperative to determine whether VCD was a complicating factor. It was proposed that an attempt be made to induce VCD by hypnotic suggestion while the patient underwent a fiberscopic laryngoscopy to establish a definitive diagnosis. The patient and his mother gave written consent for this procedure. He was admitted for observation to the pediatric intensive care unit for the induction attempt. The patient requested that no local anesthesia be applied in his nose before passage of the laryngoscope because he wanted to eat right after the procedure. Therefore, the nasopharyngeal laryngoscope was inserted while he used self hypnosis as the sole form of anesthesia. He demonstrated no discomfort during its passing. Once the vocal cords were visualized, the patient was instructed to develop an episode of respiratory distress while in a state of hypnosis by recalling a recent "neck attack." His vocal cords then were observed to adduct anteriorly with each inspiration. The patient then was asked to relax his neck. When he did, the vocal cords immediately abducted with inspiration, and he breathed easily. After removal of the laryngoscope, the patient alerted from hypnosis and said he felt well. He reported no recollection of the procedure, thus demonstrating spontaneous amnesia that sometimes is associated with hypnosis. Because the diagnosis of VCD was confirmed, the patient was encouraged to use self-hypnosis and speech therapy techniques to control his symptoms. He also was referred for counseling. To our knowledge this is the first description in the medical literature of the use of hypnotic suggestion for making a diagnosis of VCD. (ABSTRACT TRUNCATED) PMID- 11099625 TI - Attitudes toward secondhand smoke, smoking, and quitting among young people. AB - OBJECTIVE: To assess the impact of attitudes toward secondhand smoke among young people. METHODS: Three hundred nonsmokers and 300 smokers (smoked a cigarette in last 30 days) 14 through 22 years of age in the United States were surveyed with random-digit dialing. The results of this cross-sectional survey were analyzed using logistic regression to determine predictors of nonsmoking and intent to stop among current smokers. RESULTS: Controlling for age, ethnicity, and education, nonsmokers were more likely to consider smoking risky than smokers (odds ratio [OR] = 3.46). Nonsmokers were twice as likely to consider secondhand smoke dangerous than smokers (OR = 1.47). Among the variables in our model, the only statistically significant predictor of planning to stop smoking or having actually stopped was believing that secondhand smoke harmed nonsmokers, which more than doubled the chances of planning to stop or having stopped smoking (relative risk = 2.17). CONCLUSIONS: Educating young people about the dangers of secondhand smoke and empowering nonsmokers to speak out should be a strong element of any tobacco control program. PMID- 11099626 TI - Welfare reform consequences for children: the Wisconsin experience. AB - BACKGROUND: The Temporary Assistance to Needy Families, enacted under the Personal Responsibility and Work Opportunity Reconciliation Act of 1996, is a reality for many working families. As public policies are enacted, unintended consequences for infants/children must be minimized. Child advocates in Wisconsin, leading this nation in reforming Aid to Families with Dependent Children (AFDC), are concerned about supporting eligible infants/children as safety-net programs are unlinked. OBJECTIVE: This study reviews the enrollment status of 4 linked programs over time in Wisconsin, from January 1995 to August 1998. Eligible infants/children in programs, such as Medicaid/AFDC, Medicaid/Healthy Start, and Food Stamps, were analyzed and compared with enrollment in Special Supplemental Nutrition Program for Women, Infants/children (WIC), a nonlinked program. DESIGN: A cross-sectional analysis of monthly enrollment for infants/children was subdivided into 3 periods: prewelfare reform or AFDC (January 1, 1995 to December 31, 1995), the welfare reform pilot or Pay For Performance (January 1, 1996 to August 31, 1997), and welfare reform better known as Wisconsin Works (W-2), (September 1, 1998 to August 31, 1998), periods 1, 2, and 3, respectively. PARTICIPANTS: Infants/children in Wisconsin from birth to 18 years of age enrolled in W-2 and/or other safety-net programs were monitored: AFDC or W-2, WIC, Food Stamps, Medicaid/AFDC, and Medicaid/Healthy Start. RESULTS: The average number of infants/children removed from AFDC and Medicaid/AFDC during periods l and 2 were -1210 increasing to -3128 per month, respectively, almost tripling the rates of decline during the pilot period (see ). By the end of this study, >100 000 (111 198) infants/children were removed from AFDC/W-2 enrollment and 51 559 fewer infants/children benefited from Medicaid. This rate of decline slowed during period 3, averaging -687 per month, while W-2 enrollment continued to decline significantly at a rate of -2692 per month. In contrast, Medicaid/Healthy Start enrollment, targeted to infants/children <6 years of age, increased significantly over all periods by +332, +1327, and +266, respectively. Food Stamps enrollment also declined throughout all 3 consecutive periods, -603, -2462, and -1450, respectively. However, enrollment in the WIC program did not decline significantly to the same degree as other certification-linked programs with AFDC or W-2, as indicated by the consecutive slopes of -60, -111, and -183, respectively. CONCLUSION: Wisconsin infants/children were rapidly removed from welfare rolls in unprecedented numbers during the periods January 1995 and August 1998. Comparisons of periods before W-2 implementation and 1 year after implementation support the fact that certification-linked programs, such as Medicaid and Food Stamps, were sufficiently aligned to AFDC/W-2 to significantly impact infants/children enrollment. Historically, WIC certification in Wisconsin has not been linked to AFDC, and infants/children traditionally eligible for Medicaid and Food Stamps are also eligible for WIC. Yet, contrary to the AFDC-linked safety net programs, declines in WIC enrollment were not statistically significant during all study periods. Statewide and local interventions within Wisconsin, such as outreach activities, targeted to Medicaid/Healthy Start and more recently Title XXI (State Children Health Insurance Program), slowed the reductions of Medicaid enrollment for Wisconsin infants/children. These findings support that altering safety-net programs can result in unintended consequences if not carefully transitioned as demonstrated in Wisconsin welfare reform. PMID- 11099627 TI - The role of the pediatrician in the oral health of children: A national survey. AB - OBJECTIVES: To assess pediatricians' knowledge, attitudes, and professional experience regarding oral health, and to determine willingness to incorporate fluoride varnish into their practices. BACKGROUND: Poor and minority children suffer disproportionately from dental caries and have limited access to dental care. In a recent analysis of national survey data, the General Accounting Office reported that poor children had 5 times more untreated decay than did children from higher income families. Untreated decay can lead to problems with eating, speaking, and attending to learning. Children who are poor suffer 12 times the number of restricted activity days because of dental problems, compared with more affluent children. Despite higher rates of dental decay, poor children had one half the number of dental visits compared with higher income children in 1996. Medicaid's Early Periodic Screening Diagnosis and Treatment (EPSDT) program is intended to provide regular dental screenings and appropriate treatment but has apparently played a limited role in improving access to dental care for poor children. According to a report by the Office of the Inspector General of the Department of Health and Human Services, only 20% of children under 21 years of age, who were enrolled in Medicaid and eligible for EPSDT, actually received preventive dental services. By increasing their involvement in oral health prevention during well-child care visits, pediatricians may be able to play an important role in improving the dental health of their patients who have difficulty obtaining access to professional dental care. However, it is unclear to what degree pediatricians are knowledgeable about preventive oral health and the extent to which they may already be participating in prevention and assessment. Also, little is known about the incidence of dental problems in pediatric practice, and whether pediatricians perceive barriers to their patients' receiving professional dental care. Finally, it is important to know how pediatricians value the promotion of oral health and whether they would be willing to take on additional activities aimed at its improvement. We addressed these questions in a national survey of pediatricians. DESIGN: We surveyed a national sample of 1600 pediatricians randomly selected from the American Medical Association Master File to assess their knowledge, current practice, and opinion on their role in the promotion of oral health; experience with dental decay among patients and in referring patients for professional dental care; and willingness to apply fluoride varnish. RESULTS: Of 1386 eligible survey recipients, 862 returned surveys for a response rate of 62%. Respondents reported seeing dental problems regularly. Two thirds of respondents observed caries in their school aged patients at least once a month. Of the respondents, 55% reported difficulty achieving successful dental referrals for their uninsured patients and 38% reported difficulty referring their Medicaid patients. More than 90% of the respondents agreed that they had an important role in identifying dental problems and counseling families on the prevention of caries. Moreover, respondents were interested in increasing their involvement: 74% expressed a willingness to apply fluoride varnish in their practices. One half of the respondents, however, reported no previous training in dental health issues during medical school or residency, and only 9% correctly answered all 4 knowledge questions. CONCLUSION: Access to dental care and unmet dental health needs are serious, under addressed problems for poor and minority children in the United States. In promoting preventive oral health, pediatricians benefit all children and particularly the underserved. We know of 2 states, Washington and North Carolina, that have acknowledged, through the provision of reimbursement, that pediatricians have a unique opportunity at well-child care visits to provide caries prevention c PMID- 11099628 TI - Effects of cisapride on corrected QT interval, heart rate, and rhythm in infants undergoing polysomnography. AB - OBJECTIVE: To evaluate the effects of cisapride, a prokinetic gastrointestinal drug, on the electrocardiographic QT interval, heart rate, and rhythm in infants during routine 8-hour polysomnography. Reported electrocardiogram (ECG) and rhythm disturbances in a small number of patients with the use of cisapride provided the impetus for this prospective study. STUDY DESIGN: Two hundred fifty two infants born at term were enrolled. Of these, 134 were on cisapride therapy for suspected gastroesophageal reflux and 118 were not on cisapride and served as controls. Cisapride-treated and control infants were from the outset divided into 3 age groups; group 1: under 3 months of age; group 2: between 3 and 6 months of age; and group 3: >6 months of age. Continuous ECG bipolar limb lead I recording, saturation monitoring, and electroencephalography were conducted. QT intervals and heart rate were measured at hourly intervals. RESULTS: Cisapride doses were: group 1 mean, 0.80 mg/kg/day (range: 0.38-1.55); group 2 mean, 0.80 mg/kg/day (range: 0. 23-1.38); and group 3 mean, 0.72 mg/kg/day (range: 0.32-1.41). Heart rate was higher in the younger infants, with a gradual decrease with age. No difference in heart rate was detected between the cisapride and control groups. The QTc interval in patients in group 1 was statistically longer than the controls, when applying both Bazett's and Hodges' formulae for QT correction. The other age groups did not differ. No arrhythmia or atrioventricular conduction abnormalities were observed. CONCLUSION: Infants under 3 months of age on cisapride treatment had significantly longer QTc intervals (with Bazett's formula, the 98th percentile was 504 ms in the cisapride group vs 447 ms in controls). The clinical significance and risk of the increased QTc interval in these infants are unclear and need further evaluation and risk stratification. Meanwhile, cisapride should be judiciously prescribed in infants <3 months of age. PMID- 11099629 TI - Circumstances leading to a change to prone sleeping in sudden infant death syndrome victims. AB - CONTEXT: In addition to usual prone sleeping, unaccustomed prone sleeping represents a significant risk factor for sudden infant death syndrome (SIDS). However, little information is available regarding the circumstances leading caretakers to change the infant's sleep position to prone position in SIDS victims. OBJECTIVE: To determine, in a population of SIDS victims, the timing of a change to prone sleeping and the reason for that change in infants who were originally nonprone sleepers. DESIGN AND SETTING: Case series analysis from a questionnaire administered between 1991 and 1997 to parents and other caretakers of SIDS victims in the province of Quebec (Canada). SUBJECTS: One hundred fifty seven SIDS cases occurring in the province during the study. RESULTS: Of the 157 SIDS cases studied, 139 were found in the prone position, although only 93 infants usually slept prone. Of the 64 nonprone sleepers, 34 had been changed to prone by the parents or another caretaker before death, and 18 had apparently turned to prone for the first time. In the 34 cases changed to prone, the change occurred <1 week before death for 21 infants; for 16 of those infants, death occurred the first or second time that they slept prone. In 56% of the cases changed from a nonprone to prone sleeping position, a caretaker other than the parents had precipitated the change. CONCLUSIONS: Ongoing campaigns to decrease the risk of SIDS should emphasize the risk of unaccustomed prone sleeping to both parents and secondary caretakers. PMID- 11099630 TI - The pivotal role of deep vein thrombophlebitis in the development of acute disseminated staphylococcal disease in children. AB - Deep vein thrombophlebitis (DVT) and septic pulmonary emboli (PE) are rare in children. The association of DVT and acute disseminated staphylococcal disease (DSD) during childhood has not been previously reported. We report 3 children who developed a triad of DVT, septic PE, and acute osteomyelitis with Staphylococcus aureus cultured from blood and bone. One child succumbed, while 2 survived following prolonged, morbid hospitalizations. The rapid clinical deterioration observed in these patients might be caused by the aggressiveness of staphylococcal infection combined with an ongoing showering of septic emboli from the ileo-femoral DVT. We suggest that infected DVT with septic PE had a pivotal role in the development of DSD in these children. The presence of this triad should prompt aggressive treatment with the appropriate antibiotics, anticoagulation, surgical drainage, and assisted ventilation when indicated. PMID- 11099631 TI - Technical report: perinatal human immunodeficiency virus testing and prevention of transmission. Committee on Pediatric Aids. AB - In 1994, the US Public Health Service published guidelines for the use of zidovudine to decrease the risk of perinatal transmission of human immunodeficiency virus (HIV). In 1995, the American Academy of Pediatrics and the US Public Health Service recommended documented, routine HIV education and testing with consent for all pregnant women in the United States. Widespread incorporation of these guidelines into clinical practice has resulted in a dramatic decrease in the rate of perinatal HIV transmission and has contributed to more than a 75% decrease in reported cases of pediatric acquired immunodeficiency syndrome (AIDS) since 1992. Substantial advances have been made in the treatment and monitoring of HIV infection; combination antiretroviral regimens that maximally suppress virus replication are now available. These regimens are recommended for pregnant and nonpregnant individuals who require treatment. Risk factors associated with perinatal HIV transmission are now better understood, and recent results from trials to decrease the rate of mother-to child HIV transmission have contributed new strategies with established efficacy. However, perinatal HIV transmission still occurs; the Centers for Disease Control and Prevention estimates that 300 to 400 infected infants are born annually. Full implementation of recommendations for universal, routine prenatal HIV testing and evaluation of missed prevention opportunities will be critical to further decrease the incidence of pediatric HIV infection in the United States. This technical report summarizes recent advances in the prevention of perinatal transmission of HIV relevant to screening of pregnant women and their infants. PMID- 11099632 TI - Technical report: precautions regarding the use of aerosolized antibiotics. Committee on Infectious Diseases and Committee on Drugs. AB - In 1998, the Food and Drug Administration (FDA) approved the licensure of tobramycin solution for inhalation (TOBI). Although a number of additional antibiotics, including other aminoglycosides, beta-lactams, antibiotics in the polymyxin class, and vancomycin, have been administered as aerosols for many years, none are approved by the FDA for administration by inhalation. TOBI was approved by the FDA for the maintenance therapy of patients 6 years or older with cystic fibrosis (CF) who have between 25% and 75% of predicted forced expiratory volume in 1 second (FEV(1)), are colonized with Pseudomonas aeruginosa, and are able to comply with the prescribed medical regimen. TOBI was not approved for the therapy of acute pulmonary exacerbations in patients with CF nor was it approved for use in patients without CF. Currently, no other antibiotics are approved for administration by inhalation to patients with or without CF. The purpose of this statement is to briefly summarize the data that supported approval for licensure of TOBI and to provide recommendations for its safe use. The pharmacokinetics of inhaled aminoglycosides and problems associated with aerosolized antibiotic treatment, including environmental contamination, selection of resistant microbes, and airway exposure to excipients in intravenous formulations, will be discussed. PMID- 11099633 TI - Toward construction of a transcript profile database predictive of chemical toxicity. PMID- 11099634 TI - Josef Warkany. PMID- 11099635 TI - Cognitive tests: interpretation for neurotoxicity? (Workshop summary). AB - The appropriate use and interpretation of cognitive tests presents important challenges to the toxicologist and to the risk assessor. For example, intelligence cannot be measured directly; rather intelligence is quantified indirectly by scoring responses (i.e., behaviors) to specific situations (problems). This workshop, "Cognitive Tests: Interpretation for Neurotoxicity?" provided an overview on the types of cognitive tests available and described approaches by which the validity of such tests can be assessed. Unlike many tools available to the toxicologist, cognitive tests have a particular advantage. Being noninvasive and species-neutral, the same test can be performed in different mammalian species. This enhances one's ability to assess the validity of test results. Criteria for test validity include comparable responses across species as well as similar disruption by the same neurotoxicant across species. Test batteries, such as the Operant Test Battery, have indicated remarkable similarity between monkeys and children with respect to performance of certain tasks involving, for example, short-term memory. Still, there is a need for caution in interpretation of such tests. In particular, cognitive tests, especially when performed in humans, are subject to confounding by a range of factors, including age, gender, and, in particular, education. Moreover, the ability of such tests to reflect intelligence must be considered. Certain aspects of intelligence, such as the ability to plan or carry out specific tasks, are not well reflected by many of the standard tests of cognition. Nonetheless, although still under development, cognitive tests do hold promise for reliably predicting neurotoxicity in humans. PMID- 11099636 TI - 3',4'-dimethoxyflavone as an aryl hydrocarbon receptor antagonist in human breast cancer cells. AB - Treatment of MCF-7 and T47D human breast cancer cells with 3', 4' dimethoxyflavone (3',4'-DMF) alone did not induce CYP1A1-dependent ethoxyresorufin O:-deethylase (EROD) activity or reporter gene activity in cells transfected with an aryl hydrocarbon (Ah)-responsive construct (pRNH11c). In contrast, 1 nM 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) induced up to a 50- to 80-fold increase in EROD and reporter gene activity in MCF-7 and T47D cells. In cells cotreated with 1 nM TCDD plus 0.1-10 microM 3',4'-DMF, there was a concentration-dependent decrease in the TCDD-induced responses, with 100% inhibition observed at the 10 microM concentration. Gel mobility shift assays using rat liver cytosol and breast cancer cell nuclear extracts showed that 3',4' DMF alone did not transform the AhR to its nuclear binding form, but inhibited TCDD-induced AhR transformation in rat liver cytosol and blocked TCDD-induced formation of the nuclear AhR complex in MCF-7 and T47D cells. TCDD also inhibited estrogen-induced transactivation in MCF-7 cells, and this response was also blocked by 3',4'-DMF, confirming the AhR antagonist activity of this compound in breast cancer cells. PMID- 11099637 TI - Effect of trihalomethanes on cell proliferation and DNA methylation in female B6C3F1 mouse liver. AB - Trihalomethanes (chloroform, bromodichloromethane, chlorodibromomethane, and bromoform) are regulated organic contaminants in chlorinated drinking water. In female B6C3F1 mouse liver, the 4 trihalomethanes have demonstrated carcinogenic activity when administered by oral gavage; however, chloroform was not carcinogenic when administered in drinking water. Female B6C3F1 mice were administered the trihalomethanes for 11 days by gavage at 2 dose levels or in the drinking water at approximately 75% saturation. When administered by gavage, the trihalomethanes were toxic to the liver, increased the liver:body weight (bw) ratio, and increased the proliferating cell nuclear antigen-labeling index (PCNA LI). Chloroform and bromodichloromethane were the most toxic, and they increased the liver:bw ratio the most, while bromoform and chloroform increased the PCNA-LI the most. When administered in drinking water, the toxicity of the trihalomethanes was similar to their low gavage-dose. Furthermore, only chloroform significantly increased the liver:bw ratio and bromoform and chloroform increased the PCNA-LI. Chloroform and bromodichloromethane decreased the level of 5-methylcytosine in hepatic DNA. Methylation in the promoter region of the c-myc gene was reduced by the trihalomethanes. Chloroform administered by gavage was more efficacious than given in drinking water; the efficacy of the other trihalomethanes did not differ for the 2 routes. Thus, in mouse liver, the trihalomethanes administered by gavage enhanced cell proliferation and decreased the methylation of the c-myc gene, consistent with their carcinogenic activity. Furthermore, the more modest toxicity, enhancement of cell proliferation, and decreased methylation induced by chloroform administered in drinking water correlated with its lack of carcinogenic activity. Hence, the activity of the trihalomethanes was dependent on the rate of delivery, i.e. rapid by oral gavage and more slowly in drinking water. PMID- 11099638 TI - A computationally based identification algorithm for estrogen receptor ligands: part 1. Predicting hERalpha binding affinity. AB - The common reactivity pattern (COREPA) approach is a 3-dimensional, quantitative structure activity relationship (3-D QSAR) technique that permits identification and quantification of specific global and local stereoelectronic characteristics associated with a chemical's biological activity. It goes beyond conventional 3-D QSAR approaches by incorporating dynamic chemical conformational flexibility in ligand-receptor interactions. The approach provides flexibility in screening chemical data sets in that it helps establish criteria for identifying false positives and false negatives, and is not dependent upon a predetermined and specified toxicophore or an alignment of conformers to a lead compound. The algorithm was recently used to screen chemical data sets for rat androgen receptor binding affinity. To further explore the potential application of the algorithm in establishing reactivity patterns for human estrogen receptor alpha (hERalpha) binding affinity, the stereoelectronic requirements associated with the binding affinity of 45 steroidal and nonsteroidal ligands to the receptor were defined. Reactivity patterns for relative hERalpha binding affinity (RBA; 17ss-estradiol = 100%) were established based on global nucleophilicity, interatomic distances between electronegative heteroatoms, and electron donor capability of heteroatoms. These reactivity patterns were used to establish descriptor profiles for identifying and ranking compounds with RBA of > 150%, 100 10%, 10-1%, and 1-0.1%. Increasing specificity of reactivity patterns was detected for ligand data sets with RBAs above 10%. Using the results of this analysis, an exploratory expert system was developed for use in ranking relative ER binding affinity potential for large chemical data sets. PMID- 11099639 TI - A computationally based identification algorithm for estrogen receptor ligands: part 2. Evaluation of a hERalpha binding affinity model. AB - The objective of this study was to evaluate the capability of an expert system described in the previous paper (S. Bradbury et al., Toxicol. Sci. 58, 253-269) to identify the potential for chemicals to act as ligands of mammalian estrogen receptors (ERs). The basis of the expert system was a structure activity relationship (SAR) model, based on relative binding affinity (RBA) values for steroidal and nonsteroidal chemicals derived from human ERalpha (hERalpha) competitive binding assays. The expert system enables categorization of chemicals into (RBA ranges of < 0.1, 0.1 to 1, 1 to 10, 10 to 100, and >150% relative to 17ss-estradiol. In the current analysis, the algorithm was evaluated with respect to predicting RBAs of chemicals assayed with ERs from MCF7 cells, and mouse and rat uterine preparations. The best correspondence between predicted and observed RBA ranges was obtained with MCF7 cells. The agreement between predictions from the expert system and data from binding assays with mouse and rat ER(s) were less reliable, especially for chemicals with RBAs less than 10%. Prediction errors often were false positives, i.e., predictions of greater than observed RBA values. While discrepancies were likely due, in part, to species-specific variations in ER structure and ligand binding affinity, a systematic bias in structural characteristics of chemicals in the hERalpha training set, compared to the rodent evaluation data sets, also contributed to prediction errors. False positive predictions were typically associated with ligands that had shielded electronegative sites. Ligands with these structural characteristics were not well represented in the training set used to derive the expert system. Inclusion of a shielding criterion into the original expert system significantly increased the accuracy of RBA predictions. With this additional structural requirement, 38 of 46 compounds with measured RBA values greater than 10% in hERalpha, MCF7, and rodent uterine preparations were correctly categorized. Of the remaining 129 compounds in the combined data sets, RBA values for 65 compounds were correctly predicted, with 47 of the incorrect predictions being false positives. Based upon this exploratory analysis, the modeling approach, combined with a high-quality training set of RBA values derived from a diverse set of chemical structures, could provide a credible tool for prioritizing chemicals with moderate to high ER binding affinity for subsequent in vitro or in vivo assessments. PMID- 11099640 TI - Carboxylesterase and A-esterase activities during maturation and aging: relationship to the toxicity of chlorpyrifos and parathion in rats. AB - Chlorpyrifos (CPF) and parathion (PS), two common organophosphorus (OP) pesticides, exhibit higher acute toxicity in younger animals compared to adults. Maturational differences in detoxification via carboxylesterases (CEs) and A esterases (AEs) have been suggested as contributors to the higher sensitivity of younger animals to OP toxicants. AEs (e.g., chlorpyrifos oxonase and paraoxonase) catalytically inactivate while CEs stoichiometrically eliminate OP anticholinesterases. While earlier studies have reported a relationship between the toxicity of some OP pesticides and the maturational profile of AEs and CEs, little information exists on the relative OP-toxicant sensitivity and detoxification capacities of aged animals. In the present study, we investigated the relationship between toxicity of CPF and PS and the activity of CEs and AEs in liver, plasma, and lung of neonatal (7 day), juvenile (21-day), adult (3 month), and aged (24-month) Sprague Dawley rats. CE sensitivity in vitro to chlopyrifos oxon and paraoxon was also evaluated across age groups. Neonatal and juvenile rats were more sensitive than adults to the acute lethality of both CPF and PS. Aged rats exhibited similar sensitivity to CPF but were markedly more sensitive than adults to PS. Levels of CEs and AEs in neonatal and juvenile rats were significantly lower than in adult tissues. Aged rats showed similar levels of AEs in all tissues and CEs in liver and lung, but plasma CE levels were significantly lower (50%) when compared to the adult rats. There were no significant age-related differences in in vitro sensitivity of CEs to either chlorpyrifos oxon or paraoxon in any tissues. In general, acute sensitivity (MTD) was highly correlated with age-related differences in both esterase activities across all 3 tissues with CPF, but only plasma carboxylesterase activity was highly correlated with sensitivity to parathion. The results suggest that both carboxylesterase and A-esterase activities can be correlated with acute sensitivity to CPF and PS, but that age-related differences in CE activity are probably more important in differential toxicity. Furthermore, plasma carboxylesterase activity may play a more pivotal role in the differential sensitivity to PS. PMID- 11099641 TI - Mechanisms involved in the immunotoxicity induced by dermal application of JP-8 jet fuel. AB - Dermal application of JP-8 jet fuel induces immune suppression. Classic delayed type hypersensitivity as well as the induction of contact hypersensitivity to allergens applied to the shaved skin of JP-8-treated mice is suppressed. In addition, the ability of T cells isolated from JP-8-treated mice to proliferate in vitro is suppressed. Here we focused on further characterizing the immunotoxicity induced by JP-8 exposure and determining the mechanism involved. Suppression of T-cell proliferation was first noted 3 to 4 days after a single JP 8 treatment and lasted for approximately 3 weeks, at which time T-cell proliferation returned to normal. Cellular immune reactions appear to be more susceptible to the immunosuppressive effects of JP-8, as antibody production in JP-8-treated mice was identical to that found in normal controls. The mechanism through which dermal application of JP-8 suppresses cell-mediated immune reactions appears to be via the release of immune biological-response modifiers. Blocking the production of prostaglandin E(2) with a selective cyclooxygenase-2 inhibitor abrogated JP-8-induced immune suppression. Neutralizing the activity of interleukin-10 with a highly specific monoclonal antibody also blocked JP-8 induced immune suppression. Furthermore, injecting JP-8-treated mice with recombinant interleukin-12, a cytokine that drives cell-mediated immune reactions in vivo, overcame the immunotoxic effects of JP-8 and restored immune function. These data indicate that immune suppressive cytokines, presumably produced by JP 8-treated epidermal cells, are responsible for immune suppression in JP-8-treated mice and that blocking and/or neutralizing their production in vivo overcomes the immunotoxic effects of JP-8. PMID- 11099642 TI - Influence of MHC background on the antibody response to detergent enzymes in the mouse intranasal test. AB - The mouse intranasal test (MINT) was developed to assess the immunogenic potential of detergent enzymes. The BDF1 mouse (H-2(b/d)), a cross of C57Bl/6 (H 2(b)) x DBA/2 (H-2(d)) has been used for most of the development work. Preliminary data in the CB6F1(H-2(d/b)), a cross of Balb/c (H-2(d)) x C57Bl/6 showed that this strain was similar to the BDF1 in its response to enzymes. These data also showed that the parental strains responded differently to the enzymes. To understand better the influence the major histocompatibility complex (MHC) background has on immune responses to enzymes, 3 different enzymes were tested in 4 inbred strains (C57Bl/6, DBA/2, Balb/c, and C57Bl/10), 2 hybrid strains (BDF1 and CB6F1), and 2 congenic strains (Balb.B10 and B10.D2). BDF1 mice rank enzymes the same as the guinea pig, which in turn correlates with sensitization in occupationally exposed humans. The ranking is based upon the dose of enzyme needed to give one-half maximal IgG1 antibody response (ED(50)) where Termamyl is more potent than Alcalase, which is equipotent to Savinase. The H-2(d) strains ranked the enzymes the same as the BDF1 but generated ED(50)s for the proteases that were one order of magnitude greater than the BDF1 ED(50)s. The response to Termamyl was the same in the two F1 strains and the H-2(d) strains. The H-2(b) strains did not rank the enzymes the same as BDF1 but the ED(50)s for the proteases were similar to the ED(50)s in the F1 strains. The response to Termamyl in the H-2(b) strains was lower than the response in the F1 and H-2(d) strains. Initial data show that the inbred strains will make enzyme-specific IgE antibody to high doses of enzyme with DBA/2 > Balb/c > C57Bl/6 in terms of the robustness of the response. The IgG1 responses in the congenic strains were similar to the responses in the H-2 matched strains. In addition, the antibody response to enzymes was consistent regardless of the source of BDF1 mice. The responses to these enzymes are clearly MHC linked with a role for Class II I-E molecules indicated. The current data strongly support the use of the F1 hybrid as an appropriate strain for evaluating allergic responses to enzymes. PMID- 11099643 TI - Corneal organ culture model for assessing epithelial responses to surfactants. AB - The main goal of the present study was to investigate the response of cultured bovine corneas to the application of irritant substances and its potential use for predicting ocular irritancy in humans. We hypothesized that chemicals causing eye irritation may induce disruption of epithelial tight junctions and trigger cell stress responses modulated via transcription factors such as AP-1 and NF kappaB. A simple air-lifted corneal organ culture system was used as an ex vivo model for ocular irritancy test. The effects of two surfactants, sodium dodecyl sulfate (SDS) and benzalkonium chloride (BAK), on corneal epithelial permeability and DNA-binding activity of AP-1 and NF-kappaB were studied in cultured bovine corneas. Both SDS and BAK induced tight junction disruption and increased permeability of corneal epithelium assessed using surface biotinylation in a concentration- and time-dependent manner. An increase in DNA-binding activity measured using electrophoretic mobility shift assay was observed when cultured corneas were treated with surfactants at concentrations causing minimal to mild ocular irritation, indicating epithelial cell stress response. Furthermore, exposure of cultured corneas to SDS or BAK at concentrations causing severe ocular irritancy resulted in a decrease in DNA-binding activity of these transcription factors in epithelial cells. These results indicate that the combination of corneal organ culture and measurements of corneal epithelial permeability and DNA-binding activity of stress-response transcription factors following chemical exposure has the potential to be used as a mechanistically based alternative to in vivo animal testing. PMID- 11099644 TI - Predicting exposure conditions that facilitate the potentiation of noise-induced hearing loss by carbon monoxide. AB - Hearing loss is the most common occupational disease in the United States, with noise serving as the presumed causative agent in most instances. This investigation characterizes the exposure conditions that facilitate the potentiation of noise-induced hearing loss (NIHL) by carbon monoxide (CO). Auditory function was compared in rats exposed 4 weeks earlier to noise alone, CO alone, combined exposure, and air in the exposure chamber. This interval between exposure and auditory threshold assessment was selected to permit recovery of temporary threshold shifts. The compound action potential (CAP) threshold evoked by pure tone stimuli was used as a measure of auditory sensitivity. The no adverse effect level (NOAEL) with respect to potentiation of NIHL was found to be 300 ppm CO. Potentiation of NIHL by CO increases linearly as CO concentration increases between 500 -1500 ppm. Benchmark dose software (version 1. 1B) published by the U.S. EPA National Center for Environmental Assessment was employed to determine a benchmark concentration of CO that produced either a 5-dB potentiation of NIHL or an increase in auditory threshold equivalent to 10% of the effect of noise alone. The lower bound for these benchmark concentrations were 320 and 194 ppm CO, respectively. Unlike CO dose, the relationship between noise severity and potentiation of NIHL by CO shows a nonlinear relationship. The greatest potentiation was observed at moderate noise exposures (100 dB, 2-h, octave band-limited noise, or OBN) that produce limited permanent threshold shifts. Repeated exposures to 95-dB noise for 2-h periods in combination with 1200 ppm CO also yielded potentiation of NIHL, though such effects were not observed following a single combined exposure. These results underscore the potential risk of hearing loss from combined exposure to noise and CO, and the risks associated with repeated exposure. PMID- 11099645 TI - 2,3,7,8-tetrachlorodibenzo-p-dioxin inhibits luminal cell differentiation and androgen responsiveness of the ventral prostate without inhibiting prostatic 5alpha-dihydrotestosterone formation or testicular androgen production in rat offspring. AB - In utero and lactational exposure to a single maternal dose of 1 microg 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD)/kg causes some overt toxicity and impairs prostate growth in male offspring. As similar effects on the ventral prostate can be caused by decreased testosterone production during perinatal development, we determined whether intratesticular testosterone content, testicular responsiveness to gonadotropin stimulation, or plasma testosterone concentrations were reduced in fetal and newborn rats. Because these endpoints were not affected, the ability of TCDD exposure to inhibit synthesis of the proximal androgen in prostate development, 5alpha-dihydrotestosterone (DHT), from the circulating precursor testosterone and 5alpha-androstane-3alpha,17ss-diol (3alpha Diol), was studied on postnatal days (PNDs) 14, 21, and 32. The ability of the ventral prostate to form DHT from 3alpha-Diol was slightly impaired on PND 14, but this transient effect was not statistically significant, and recovery was evident by PND 21. Subsequent experiments used organ culture to study the effects of in vivo TCDD exposure on androgen metabolism, androgen responsiveness, androgen receptor expression, and luminal epithelial cell differentiation after in vitro exposure to graded androgen concentrations. In utero and lactational TCDD exposure had no effect on DHT formation in organ culture, but transiently reduced the androgen -induced expression of prostatic-binding protein subunit C3, decreased ventral prostate epithelial cell androgen receptor expression, and inhibited the formation of androgen responsive luminal epithelial cells. These results suggest that TCDD exposure impairs prostate growth and androgen responsiveness by inhibiting prostatic epithelial cell differentiation. PMID- 11099646 TI - The plasticizer diethylhexyl phthalate induces malformations by decreasing fetal testosterone synthesis during sexual differentiation in the male rat. AB - Phthalate esters (PE) such as DEHP are high production volume plasticizers used in vinyl floors, food wraps, cosmetics, medical products, and toys. In spite of their widespread and long-term use, most PE have not been adequately tested for transgenerational reproductive toxicity. This is cause for concern, because several recent investigations have shown that DEHP, BBP, DBP, and DINP disrupt reproductive tract development of the male rat in an antiandrogenic manner. The present study explored whether the antiandrogenic action of DEHP occurs by (1) inhibiting testosterone (T) production, or by (2) inhibiting androgen action by binding to the androgen receptor (AR). Maternal DEHP treatment at 750 mg/kg/day from gestational day (GD) 14 to postnatal day (PND) 3 caused a reduction in T production, and reduced testicular and whole-body T levels in fetal and neonatal male rats from GD 17 to PND 2. As a consequence, anogenital distance (AGD) on PND 2 was reduced by 36% in exposed male, but not female, offspring. By GD 20, DEHP treatment also reduced testis weight. Histopathological evaluations revealed that testes in the DEHP treatment group displayed enhanced 3ss-HSD staining and increased numbers of multifocal areas of Leydig cell hyperplasia as well as multinucleated gonocytes as compared to controls at GD 20 and PND 3. In contrast to the effects of DEHP on T levels in vivo, neither DEHP nor its metabolite MEHP displayed affinity for the human androgen receptor at concentrations up to 10 microM in vitro. These data indicate that DEHP disrupts male rat sexual differentiation by reducing T to female levels in the fetal male rat during a critical stage of reproductive tract differentiation. PMID- 11099647 TI - Perinatal exposure to the phthalates DEHP, BBP, and DINP, but not DEP, DMP, or DOTP, alters sexual differentiation of the male rat. AB - In mammals, exposure to antiandrogenic chemicals during sexual differentiation can produce malformations of the reproductive tract. Perinatal administration of AR antagonists like vinclozolin and procymidone or chemicals like di(2 ethylhexyl) phthalate (DEHP) that inhibit fetal testicular testosterone production demasculinize the males such that they display reduced anogenital distance (AGD), retained nipples, cleft phallus with hypospadias, undescended testes, a vaginal pouch, epididymal agenesis, and small to absent sex accessory glands as adults. In addition to DEHP, di-n-butyl (DBP) also has been shown to display antiandrogenic activity and induce malformations in male rats. In the current investigation, we examined several phthalate esters to determine if they altered sexual differentiation in an antiandrogenic manner. We hypothesized that the phthalate esters that altered testis function in the pubertal male rat would also alter testis function in the fetal male and produce malformations of androgen-dependent tissues. In this regard, we expected that benzyl butyl (BBP) and diethylhexyl (DEHP) phthalate would alter sexual differentiation, while dioctyl tere- (DOTP or DEHT), diethyl (DEP), and dimethyl (DMP) phthalate would not. We expected that the phthalate mixture diisononyl phthalate (DINP) would be weakly active due to the presence of some phthalates with a 6-7 ester group. DEHP, BBP, DINP, DEP, DMP, or DOTP were administered orally to the dam at 0.75 g/kg from gestational day (GD) 14 to postnatal day (PND) 3. None of the treatments induced overt maternal toxicity or reduced litter sizes. While only DEHP treatment reduced maternal weight gain during the entire dosing period by about 15 g, both DEHP and DINP reduced pregnancy weight gain to GD 21 by 24 g and 14 g, respectively. DEHP and BBP treatments reduced pup weight at birth (15%). Male (but not female) pups from the DEHP and BBP groups displayed shortened AGDs (about 30%) and reduced testis weights (about 35%). As infants, males in the DEHP, BBP, and DINP groups displayed femalelike areolas/nipples (87, 70, and 22% (p < 0.01), respectively, versus 0% in other groups). All three of the phthalate treatments that induced areolas also induced a significant incidence of reproductive malformations. The percentages of males with malformations were 82% (p < 0.0001) for DEHP, 84% (p < 0.0001) for BBP, and 7.7% (p < 0.04) in the DINP group. In summary, DEHP, BBP, and DINP all altered sexual differentiation, whereas DOTP, DEP, and DMP were ineffective at this dose. Whereas DEHP and BBP were of equivalent potency, DINP was about an order of magnitude less active. PMID- 11099648 TI - The effects of atrazine on female wistar rats: an evaluation of the protocol for assessing pubertal development and thyroid function. AB - The effects of atrazine (ATR), a chlorotriazine herbicide, on the onset of puberty were evaluated in Wistar rats. Female rats were dosed by oral gavage from postnatal day(s) (PND) 22 through PND 41 with 0, 12.5, 25, 50, 100, or 200 mg ATR/kg. Vaginal opening (VO) was significantly delayed 3.4, 4.5, or greater than 6.8 days by 50, 100, and 200 mg/kg, respectively. VO had not occurred in 4 of 15 females in the 200 mg/kg group by the time of necropsies (PND 41). Body weight (bw) at necropsy was reduced in the 200 mg/kg group by 11.6%, but was not different from the control (0) in the 50 and 100 mg/kg groups. To examine the influence of reduced bw on pubertal development, a group of pair-fed controls was included whose daily food intake was dependent upon the amount consumed by their counterpart in the 200 mg/kg group. Although necropsy bw was reduced to the same extent as the ATR females, VO in the pair-fed controls was not significantly delayed. Adrenal, kidney, pituitary, ovary, and uterine weights were reduced by 200 mg/kg ATR. Serum T(3), T(4), and TSH were unaltered by ATR, which was consistent with no histopathologic/morphologic changes in the thyroid. Estrous cyclicity was monitored in a second group of females from VO to PND 149. The number of females displaying regular 4- or 5-day estrous cycles during the first 15-day interval after VO was lower in the 100 and 200 mg/kg ATR and pair-fed controls. Irregular cycles were characterized by extended periods of diestrus. By the end of the second 15-day interval (PND 57-71), no effects on estrous cyclicity were observed. These data show that ATR can delay the onset of puberty and alter estrous cyclicity in the female Wistar rat ( NOAEL of 25 mg/kg). Reduced food consumption and bw did not account for the delay in VO, because this effect was not observed in the pair-fed controls. In addition, the effect on estrous cyclicity was observed in the 100 mg/kg ATR group where no significant reduction in bw was observed. PMID- 11099649 TI - Chronic toxicity of di(2-ethylhexyl)phthalate in mice. AB - B6C3F1 mice were treated with 0, 100, 500, 1500, or 6000 ppm di(2 ethylhexyl)phthalate (DEHP) in the diet for up to 104 weeks. Blood and urine were analyzed at Weeks 26, 52, 78, and 104 from 10 animals per sex per group. Body weights and food consumption were measured weekly for the first 16 weeks, then monthly thereafter. Survival was reduced for mice receiving 6000 ppm DEHP. Overall weight gains were significantly lower for the 6000-ppm male group, but there was no difference among female groups. Food consumption was not affected by exposure. No biologically significant changes in clinical chemistry, hematology, or urinalysis were observed. After 104 weeks of exposure, kidney weights for the 500- and 1500-ppm male, and 6000-ppm male/female groups were significantly lower than for the controls. Significantly higher liver weight was seen for the 500-, 1500-, and 6000-ppm male groups and the 6000-ppm female group of mice. Testis weights for the 500-, 1500-, and 6000-ppm males were significantly lower than for the controls. Uterine weights for the 6000-ppm group were significantly lower than for the controls. All organs were examined for histopathology. The incidence of hepatocellular lesions has been reported separately (R. M. David et al., 1999. Toxicol. Sci. 50, 195-205). Tumors were observed at > or = 500-ppm dosages, where peroxisome proliferation was significantly increased. A NOEL for both tumors and peroxisome proliferation was 100 ppm. In the study presented here, bilateral hypospermia in the testes of male mice, hepatocyte pigmentation and cytoplasmic eosinophilia in the liver, and chronic progressive nephropathy of male and female mice were observed at 6000 ppm. Hypospermia and chronic progressive nephropathy were also observed at 1500 ppm, where peroxisome proliferation was 2.7-6.8-fold higher than controls. Many lesions observed in rats were not seen in mice. A dose level of 500 ppm (98.5-116.8 mg/kg/day) was identified as a no-observed-adverse effect level (NOAEL) for noncarcinogenic effects. PMID- 11099650 TI - trans-Nonachlor and cis-nonachlor toxicity in Sprague-Dawley rats: comparison with technical chlordane. AB - cis-Nonachlor and trans-nonachlor are bioaccumulating components of the pesticide chlordane, which can be detected in various environmental biota and in humans. Existing studies have focused on the potential adverse health effects of the parent chlordane mixture. Comparable toxicity data are nonexistent for individual chlordane constituents such as trans-nonachlor, cis-nonachlor, or oxychlordane, which are among the most common chlordane-related environmental contaminants and tissue residues. In this study, rats were administered cis-nonachlor, trans nonachlor, or technical chlordane by gavage for 28 days at doses of 0.25 to 25 mg/kg body weight. Residue analyses indicated that trans-nonachlor accumulation in adipose was greater than cis-nonachlor when rats were administered each chemical under identical conditions of dose and exposure. For all test chemicals, the major metabolite oxychlordane accumulated in adipose tissue. Adipose tissue residue levels of all test chemicals and the major metabolite were higher in female rats. The liver was a target organ in male and female rats, indicated by increased liver weight and histopathological changes consistent with microsomal enzyme induction. Hepatic changes were most pronounced in rats treated with trans nonachlor. Elevated kidney weights and depressed organic ion transport were observed in males treated with trans-nonachlor and chlordane. Although in general, changes in target organs and clinical chemistry endpoints were similar for all 3 test chemicals, the approximate toxicity ranking from most to least toxic was trans-nonachlor > technical chlordane > cis-nonachlor. PMID- 11099651 TI - Toxicogenomics-based discrimination of toxic mechanism in HepG2 human hepatoma cells. AB - The rapid discovery of sequence information from the Human Genome Project has exponentially increased the amount of data that can be retrieved from biomedical experiments. Gene expression profiling, through the use of microarray technology, is rapidly contributing to an improved understanding of global, coordinated cellular events in a variety of paradigms. In the field of toxicology, the potential application of toxicogenomics to indicate the toxicity of unknown compounds has been suggested but remains largely unsubstantiated to date. A major supposition of toxicogenomics is that global changes in the expression of individual mRNAs (i.e., the transcriptional responses of cells to toxicants) will be sufficiently distinct, robust, and reproducible to allow discrimination of toxicants from different classes. Definitive demonstration is still lacking for such specific "genetic fingerprints," as opposed to nonspecific general stress responses that may be indistinguishable between compounds and therefore not suitable as probes of toxic mechanisms. The present studies demonstrate a general application of toxicogenomics that distinguishes two mechanistically unrelated classes of toxicants (cytotoxic anti-inflammatory drugs and DNA-damaging agents) based solely upon a cluster-type analysis of genes differentially induced or repressed in cultured cells during exposure to these compounds. Initial comparisons of the expression patterns for 100 toxic compounds, using all approximately 250 genes on a DNA microarray ( approximately 2.5 million data points), failed to discriminate between toxicant classes. A major obstacle encountered in these studies was the lack of reproducible gene responses, presumably due to biological variability and technological limitations. Thus multiple replicate observations for the prototypical DNA damaging agent, cisplatin, and the non-steroidal anti-inflammatory drugs (NSAIDs) diflunisal and flufenamic acid were made, and a subset of genes yielding reproducible inductions/repressions was selected for comparison. Many of the "fingerprint genes" identified in these studies were consistent with previous observations reported in the literature (e. g., the well-characterized induction by cisplatin of p53-regulated transcripts such as p21(waf1/cip1) and PCNA [proliferating cell nuclear antigen]). These gene subsets not only discriminated among the three compounds in the learning set but also showed predictive value for the rest of the database ( approximately 100 compounds of various toxic mechanisms). Further refinement of the clustering strategy, using a computer-based optimization algorithm, yielded even better results and demonstrated that genes that ultimately best discriminated between DNA damage and NSAIDs were involved in such diverse processes as DNA repair, xenobiotic metabolism, transcriptional activation, structural maintenance, cell cycle control, signal transduction, and apoptosis. The determination of genes whose responses appropriately group and dissociate anti-inflammatory versus DNA-damaging agents provides an initial paradigm upon which to build for future, higher throughput-based identification of toxic compounds using gene expression patterns alone. PMID- 11099652 TI - Does secondhand smoke activate platelets? PMID- 11099653 TI - Re: "Reevaluation of the cancer potency factor of toxaphene: recommendation from a peer review". PMID- 11099654 TI - Universal RBCs. PMID- 11099655 TI - Transfusion to blood group A and O patients of group B RBCs that have been enzymatically converted to group O. AB - BACKGROUND: The transfusion of ABO-incompatible RBCs is the leading cause of fatal transfusion reactions. Group O RBCs, lacking terminal immunodominant A and B sugars to which humans are immunized, are safe for transfusion to persons of any ABO blood group. With the use of a recombinant alpha-galactosidase to remove terminal galactose from group B RBCs, the safety and efficacy of enzyme-converted group-B-to-group-O (ECO) RBC components were studied in transfusion-dependent patients. STUDY DESIGN AND METHODS: Twenty-four patients (blood groups A and O) were randomly assigned to receive transfusion(s) of either ECO or control group O RBCs. If a second transfusion was given, the other blood component was administered. RESULTS: Twenty-one patients were given ECO RBCs; 18 also underwent control transfusions. One patient received only a small aliquot for RBC survival studies, instead of a full-unit transfusion, because his serum was incompatible with ECO RBCs. No adverse events occurred. Both ECO and control transfusions resulted in appropriate Hb increments and comparable (51)Cr-labeled RBC survival studies. One patient developed a transient, weak-positive DAT, without hemolysis. Two weeks after transfusion, 5 of 19 evaluable ECO RBC recipients had increases in anti-B titers. CONCLUSION: ECO RBCs were comparable to group O cells for safety and efficacy in this study. The clinical significance of the increase in anti-B and of occasional serologic incompatibilities with ECO RBCs is unclear. If strategies can be developed to remove A epitopes, enzymatic conversion could be used to create a universal (group O) donor blood supply. PMID- 11099656 TI - Serum supplement, inoculum cell density, and accessory cell effects are dependent on the cytokine combination selected to expand human HPCs ex vivo. AB - BACKGROUND: The prolonged periods of pancytopenia associated with cord blood transplants suggest that in some cases cell numbers may be limiting. The possibility that limiting cell numbers may be overcome and prolonged periods of pancytopenia abrogated by the transplantation of human umbilical cord blood cells expanded ex vivo has led to efforts to define optimal culture conditions for these cells. STUDY DESIGN AND METHODS: Cord blood CD34+ cells were cultured with three cytokine combinations: SCF+G-CSF+GM-CSF+MGDF (SGGM); IL-6+ SCF+MGDF+Flt3 ligand (6SMF); and IL-1+IL-3+IL-6+G-CSF+GM-CSF+SCF+Epo (GFmix). Serum effects, inoculum concentration (cells/mL) seeding density (cell/cm(2)) and accessory cell effects on the expansion of CD34+ cells were determined. RESULTS: Cellular outputs were significantly higher with fetal calf serum (FCS) than with cord blood serum (CBS) or adult group AB serum (ABS) in the presence of 6SMF, however, CBS was as effective as FCS. The best seeding concentrations varied for each of the cytokine combinations, and inoculum densities exceeding 1000 cells per cm(2) proved detrimental for cultures containing GFmix and SGGM. Accessory cell studies indicated that populations expressing the CD33 antigen inhibited the expansion of purified CD34+ cells in the presence of GFmix or SGGM, but not in the presence of 6SMF. CONCLUSION: Serum supplement, inoculum cell concentration, seeding densities, and accessory cell effects are dependent upon the cytokine combination selected to expand cord blood HPCs ex vivo. Thus, each of these measures should be assessed to establish reproducible and reliable conditions for the selection of different cytokine combinations to culture cord blood HPCs. PMID- 11099657 TI - Growth of bacteria in inoculated platelets: implications for bacteria detection and the extension of platelet storage. AB - BACKGROUND: Recent reports from Europe have advocated the use of bacterial culturing of platelets on Day 2 or 3 of storage to extend the shelf life of platelets to 7 days, thereby reducing the outdating of platelets and preserving a limited medical resource. To assess the optimal timing, the necessary sensitivity, and the possible efficacy of bacterial detection, the bacterial growth characteristics were reviewed in 165 platelet units, each inoculated on the day of collection with one of the following organisms: Bacillus cereus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Serratia marcescens, Staphylococcus aureus, and Staphylococcus epidermidis from four previously published studies. STUDY DESIGN AND METHODS: Quantitative culture data from inoculated platelet concentrates from five sites and four studies were combined into one database and analyzed for bacterial concentration thresholds (> or =10(1), > or =10(2), > or =10(3), > or =10(4), > or =10(5) CFU/mL) by day of storage. RESULTS: All examples of B. cereus, P. aeruginosa, K. pneumoniae, S. marcescens, and S. aureus had concentrations > or =10(2) CFU per mL by Day 3 after inoculation. By Day 4, all units with these organisms contained > or =10(5) CFU per mL. Units contaminated with S. epidermidis showed slower and more varied growth. By Day 3 after inoculation, 81.3 percent had 10(2) CFU per mL. By Day 4 after inoculation, 46 (95.8%) of 48 units had concentrations > or =10(2) CFU per mL. CONCLUSION: These experiments suggest that an assay capable of detecting 10(2) CFU per mL on Day 3 of storage would detect the vast majority of bacterially contaminated platelet units, prevent many cases of platelet associated bacterial sepsis, and provide a scientific basis for the extension of the current platelet storage time. It would be expected that a rare, slow-growing organism could escape such a detection scheme. PMID- 11099658 TI - L-carnitine decreases glycolysis in liquid-stored platelets. AB - BACKGROUND: The platelet storage lesion is characterized metabolically by a pH decrease associated with lactic acid generation; a change in platelet morphology from discoid to spherical; a diminished response to in vitro challenge tests, such as the hypotonic shock response (HSR) and extent of shape change (ESC); increased surface P-selectin expression; and decreased in vivo recovery and survival. Altering storage conditions to improve these measures could allow for extension of the duration of in vitro storage. STUDY DESIGN AND METHODS: ABO identical paired platelet concentrates were pooled and then equally divided into two plastic bags. Either L-carnitine (LC) or an equal volume of saline (control) was added to one container of each pair. Platelets were stored at 20 to 24 degrees C for 5 to 10 days or at 1 to 6 degrees C for 5 days at various concentrations of LC between 0.1 and 15 mM: At the end of storage, pH, glucose consumption, lactate generation, HSR, ESC, and surface P-selectin expression were measured. In different experiments, paired platelet concentrates were spiked with a Staphylococcus epidermidis suspension in the presence and absence of L carnitine at a concentration of 5 mM: RESULTS: At 20 to 24 degrees C and concentrations of LC between 0.1 and 5 mM:, there was evidence of better pH preservation, less glucose consumption, and less lactate generation. Only with storage beyond 5 days was a difference present in either surface P-selectin expression or HSR. An L-carnitine concentration of 5 mM: appeared optimal. L carnitine did not enhance the growth of bacteria after 7 to 8 days of storage. CONCLUSION: LC at 5 mM: may improve the quality of platelet concentrates that are stored beyond 5 days. There was no indication that LC at this concentration would promote bacterial growth. It may be a useful additive to platelet preservation. PMID- 11099659 TI - The mechanism of apoptosis in human platelets during storage. AB - BACKGROUND: Although it is usually involved only in nucleated cells (NCs), artificially enucleated cells also lose viability by a programmed process of cell death called apoptosis. Because platelets undergo loss of viability during storage, an attempt was made to determine whether platelets contained the apoptotic mechanisms and whether it was activated during platelet storage. STUDY DESIGN AND METHODS: Platelet viability was measured by reduction of a tetrazolium dye (MTS) and annexin V binding. Members of the death receptor, caspase, and Bcl 2 families were detected by RNase protection assay and Western blotting. Caspase 3 activation was measured by enzyme and Western blot assays and by cleavage of gelsolin. RESULTS: After 5 days of storage under standard blood banking conditions, platelets display biochemical signs of apoptosis by losing MTS activity and increasing the amount of phosphatidylserine on their surface. The mRNA and the proenzyme for several members of the caspase, death receptor, and Bcl-2 families are expressed at high levels in platelets. An increase in caspase 3 activity and the amount of the biologically active p17 subunit of active caspase 3 were observed to coincide with the appearance of apoptotic markers during storage. These effects were not due to platelet activation. The caspase 3 substrate, gelsolin, began to undergo proteolysis after 3 to 4 days of storage, and the addition of the caspase inhibitor z-VAD-fmt substantially inhibited this process. CONCLUSION: Platelets contain many of the components of the apoptotic mechanism and show activation of caspase 3 and consequent cleavage of gelsolin during storage, independent of platelet activation. Evaluation of the mechanism of apoptosis in platelets may provide a basis for developing novel strategies to enhance platelet viability during storage. PMID- 11099660 TI - Selective protection of RBCs against photodynamic damage by the band 3 ligand dipyridamole. AB - BACKGROUND: All studied photosensitizers for virus inactivation impair RBCs. To reduce damage to the RBCs without affecting virucidal activity, selective protection of the RBCs is necessary. The ability of the band 3 ligand, dipyridamole, to react with singlet oxygen and to increase the selectivity of photosterilization was investigated. STUDY DESIGN AND METHODS: Solutions of dipyridamole were illuminated in the presence of tetrasulfonated aluminum phthalocyanine (AlPcS(4)) and dimethylmethylene blue (DMMB). Solutions of amino acids, RBCs, and vesicular stomatitis virus (VSV) in RBC suspensions were photodynamically treated in the presence or absence of dipyridamole. RESULTS: Illumination of a solution of dipyridamole in the presence of AlPcS(4) or DMMB resulted in changes in the optical spectrum of dipyridamole. The photooxidation of dipyridamole was inhibited by azide and augmented by D(2)O, which suggests the involvement of singlet oxygen. Photooxidation of amino acids and photodamage to RBCs was strongly reduced in the presence of dipyridamole. In contrast, photoinactivation of VSV in RBC suspensions was only slightly affected by dipyridamole. CONCLUSION: Dipyridamole can improve the specificity of photodynamic sterilization of RBC concentrates, thereby increasing the practical applicability of this photodecontamination method. PMID- 11099661 TI - Posttransfusion survival (24-hour) and hemolysis of previously frozen, deglycerolized RBCs after storage at 4 degrees C for up to 14 days in sodium chloride alone or sodium chloride supplemented with additive solutions. AB - BACKGROUND: Previously frozen human RBCs currently are glycerolized and deglycerolized by the use of open systems that limit storage of the deglycerolized RBCs at 4 degrees C to only 24 hours. STUDY DESIGN AND METHODS: Healthy male volunteers who met AABB requirements for blood donors (n = 38) were studied. A volume of 450 mL of blood was collected into CPDA-1. The RBC concentrates were stored at 4 degrees C for 3 to 6 days before being frozen with 40-percent (wt/vol) glycerol and stored at -80 degrees C. The RBCs were deglycerolized, resuspended in 0.9-percent sodium chloride and 0.2-percent glucose (SG) solution or SG solution supplemented with AS-1, AS-3, or AS-5, and stored in the resuspension medium at 4 degrees C for 14 days. RESULTS: The mean +/- SD freeze-thaw-wash process recovery was 90.0 +/- 4.0 percent for all 38 units. The mean 24-hour posttransfusion survival value was 79 percent for deglycerolized RBC stored at 4 degrees C for 7 days in SG alone, SG plus AS-3, or SG plus AS-5. Deglycerolized RBC that were stored at 4 C for 14 days in SG supplemented with AS-1, AS-3, or AS-5 had a mean 24-hour posttransfusion survival of 74 percent. After 7 days of storage of deglycerolized RBCs in SG alone, the mean hemolysis was 3. 7 percent. After 14 days of storage of deglycerolized RBCs in SG supplemented with AS-1, AS-3, or AS-5, the mean hemolysis was 2.5 percent. CONCLUSIONS: The levels of hemolysis did not correlate with the 24-hour posttransfusion survival values. PMID- 11099662 TI - The survival, function, and hemolysis of human RBCs stored at 4 degrees C in additive solution (AS-1, AS-3, or AS-5) for 42 days and then biochemically modified, frozen, thawed, washed, and stored at 4 degrees C in sodium chloride and glucose solution for 24 hours. AB - BACKGROUND: A study was done to assess the quality of RBCs stored at 4 degrees C in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. STUDY DESIGN AND METHODS: RBCs were stored at 4 degrees C for 42 days in AS-1, AS 3, or AS-5 and then biochemically modified with pyruvate, inosine, phosphate, and adenine solution (Rejuvesol), frozen with 40-percent (wt/vol) glycerol, and stored at -80 degrees C for at least 2 months. The RBCs were deglycerolized by the use of a cell washer (Haemonetics 115), and stored for 24 hours at 4 degrees C in a 0.9-percent sodium chloride and 0.2-percent glucose solution before the autologous transfusion. RESULTS: The mean freeze-thaw-wash recovery process produced RBC recovery values of 85 percent, with the mean 24-hour posttransfusion survival at 75 percent, and the mean index of therapeutic effectiveness at 64 percent for the RBCs stored at 4 degrees C in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. All the units exhibited normal or slightly higher than normal 2,3 DPG levels after deglycerolization and postwash storage at 4 degrees C for 24 hours. CONCLUSION: RBCs stored in AS-1, AS-3, or AS 5 at 4 degrees C for 42 days and then biochemically modified with pyruvate, inosine, phosphate, and adenine and glycerolized, frozen, washed, and stored at 4 degrees C for 24 hours before autologous transfusion had acceptable in vitro and in vivo measurements. PMID- 11099663 TI - The capacity of different infusion fluids to lower the prooxidant activity of plasma iron: an important factor in resuscitation? AB - BACKGROUND: Prooxidant activity of non-protein-bound iron (NPBI) is an important contributor to reactive oxygen species-induced injury after the resuscitation of critically ill patients. Plasma NPBI occurs in critically ill adults, children, and newborn babies, who often require resuscitation. The ability of the resuscitation fluids to bind iron and lower the patients' NPBI levels in vitro has not previously been studied. STUDY DESIGN AND METHODS: In an in vitro model, highly iron-saturated cord blood plasma from 10 preterm and 10 term babies was mixed with FFP, pasteurized plasma protein solution, and 0.9-percent saline. Plasma from 10 healthy adult volunteers was used as a control. Before and after the mixing with any resuscitation fluid, NPBI levels and ceruloplasmin iron oxidizing and transferrin iron-binding antioxidant capacities were measured. RESULTS: After the in vitro mixing with FFP, the incidence and concentration of NPBI were markedly decreased and the iron-binding antioxidant capacity was increased in the plasma of the preterm and term babies. Being mixed with pasteurized plasma protein solution and 0.9-percent saline did not influence the iron-binding antioxidant capacity of newborn babies' plasma. In the control plasma, results were not changed after the mixing with any resuscitation fluid. In every group, the iron-oxidizing antioxidant capacity was not changed after the mixing with any fluid. CONCLUSION: Iron-induced oxidative tissue damage may be influenced by resuscitation fluids. In the ongoing debate over the choice of crystalloid or colloid resuscitation fluids, the influence of each fluid on the patient's antioxidant capacity warrants more attention. PMID- 11099664 TI - Transfusion of washed and centrifuged shed RBCs during maxillofacial surgery affects cytokine concentrations. AB - BACKGROUND: In patients undergoing elective maxillofacial surgery, hyperthermic reactions have been observed after the transfusion of autologous washed and centrifuged shed blood. It was the aim of this study to correlate the clinical features with changes in cytokine levels. STUDY DESIGN AND METHODS: In 24 consecutive patients, TNFalpha, IL-1, and IL-6 levels were determined in washed and centrifuged shed RBCs (CS RBCs) and in the patient's serum before, as well as 15 and 120 minutes after transfusion. At the same time, blood was drawn for culture. Patients in whom whole blood was saved through the use of acute normovolemic hemodilution served as a control group (n = 6). RESULTS: After the transfusion of CS RBCs, patients had not only elevated cytokine levels but also transient bacteremia involving the pathogens previously detected in CS RBCs. No rise in body temperature occurred. CONCLUSION: In the light of these results, the use of CS RBCs in patients undergoing maxillofacial surgery should be restricted to those patients with no primary bacterial contamination. PMID- 11099665 TI - Apparent rejuvenation of transfused donor blood in the fetus is due to accelerated removal of the older RBCs. AB - BACKGROUND: In severely anemic fetuses of women alloimmunized to RBC antigens, transfused donor RBCs disappear faster than in adults. This may result from an accelerated linear or nonlinear decline with time. It was investigated whether changes in donor RBC age characteristics after circulation in the fetus may reflect the main type of cellular decline. STUDY DESIGN AND METHODS: Donor RBC age characteristics (density, mean cell volume [MCV], and mean cell Hb content [MCHC]) were determined before intrauterine transfusions. Density gradient centrifugation was used to obtain RBCs of different ages. The results from gradient centrifugation were used to calculate mean values for the density, MCV, and MCHC to be expected after the transfusion interval, assuming a linear decline in RBCs of 1 percent per day. Donor and fetal RBCs, taken just before the second transfusion, were separated by agglutination with IgM D MoAb. For these donor cells, the observed mean values for density, MCV, and MCHC were compared with the calculated, expected values (n = 12). RESULTS: The mean +/- SD transfusion interval was 17.9 +/- 3.6 days. The Hb declined by 1.75 +/- 0.62 percent per day (n = 9). After the transfusion interval and contrary to the expected changes, cell density and MCHC decreased and MCV increased significantly (0. 001C). CONCLUSION: This is the first report describing the molecular alteration in a non-CGD McLeod patient who has made anti-Kx. The immune response of people with the McLeod phenotype can vary, and K(O) blood may not always be compatible. PMID- 11099668 TI - Analysis of factors affecting quantification of fetomaternal hemorrhage by flow cytometry. AB - BACKGROUND: An analysis was carried out to determine the sources and extent of errors encountered in the quantitation of the volume of fetomaternal hemorrhage (FMH) by flow cytometry. Different assay conditions were compared, to define the simplest, most accurate protocol. STUDY DESIGN AND METHODS: D-, D+, and artificial FMH (mixtures of D+ and D- RBCs) were stained either by a direct method (using FITC-conjugated IgG3 D MoAb [BRAD-3]), with or without dual labeling with PE-conjugated anti-GPA, or by indirect methods (using polyclonal anti-D followed by FITC- or biotin-conjugated anti-IgG reagents). Cells were selected for flow cytometric analysis on the basis of either forward or side scatter (log FSC/log SSC) characteristics or of GPA+ labeling or were unselected. The numbers of events labeled with anti-D were determined from histograms. For some samples, 10 replicates of 500,000 events each were analyzed. RESULTS: Background fluorescent events in 10 directly labeled gated D- samples ranged from 0.007 to 0.023 percent, equivalent to 0.15- to 0.51-mL FMH. Both the use of a gate on log FSC/SSC or the selection of GPA+ events only resulted in a reduction in FMH of 0.3 mL or less. The intra-assay variation in FMH, or sampling error, was found to be approximately 10 percent at low artificial FMH (<10 mL) but greater (< or =50% with a CV of 15%) with D- samples. Direct staining was quicker and produced a lower background than indirect staining. CONCLUSION: The inherent sampling error that is due to the random distribution of rare events throughout the blood sample contributed greatly to the variation in the volume of FMH calculated by flow cytometry. The FMH should not be underestimated. For a routine assay, a simplified protocol and calculation will be sufficiently accurate to determine the dose of prophylactic anti-D that should be given to the patient. PMID- 11099669 TI - In vitro studies of the impact of transfusion on the detection of alloantibodies after autoadsorption. AB - BACKGROUND: In a patient with warm autoantibodies who has recently received a transfusion, it is not recommended to perform adsorptions using autologous RBCs to detect alloantibodies. Although not scientifically documented, this position is based on the theory that transfused RBCs in the patient's circulation would be capable of adsorbing alloantibodies that may be present. This in vitro study was designed to determine what percentage of transfused RBCs might completely remove alloantibodies in vivo. STUDY DESIGN AND METHODS: Selected D, E, K, Fy(a), and Jk(a) antibodies were adsorbed with mixtures of antigen-positive and antigen negative RBCs to determine the lowest concentration of antigen-positive RBCs capable of removing all alloantibody reactivity. The percentage of antigen positive RBCs in each mixture was determined by flow cytometry. RESULTS: Small amounts of antigen-positive RBCs (2-6%, as determined by flow cytometry) completely removed anti-D, -E, and -Fy(a) reactivity. Reactivity of two examples of anti-K was removed by 11 percent and 17 percent of K+ RBCs, respectively. Anti Jk(a) reactivity was completely removed by 4 to 5 percent Jk(a+) RBCs using a PEG adsorption; the endpoint (>11%) was estimated, but complete adsorption with ZZAP treated RBCs was not performed. CONCLUSION: Small amounts of antigen-positive RBCs are generally capable of removing all alloantibody reactivity. Thus, waiting for 3 months after transfusion before performing autologous adsorptions is a prudent policy. PMID- 11099670 TI - Allelic polymorphisms of human fcgamma receptor IIa and Fcgamma receptor IIIb among distinct groups in Brazil. AB - BACKGROUND: The FcgammaRIIA gene is expressed in two polymorphic forms, R131 and H131, which differ by the replacement of histidine by arginine at position 131. The FCGR3B (FcgammaRIIIB) gene exists in two allelic isoforms, known as FCGR3B1 (FcgammaRIIIB-NA1) and FCGR3B2 (FcgammaRIIIB-NA2), which differ in nucleotides 141, 147, 227, 277, and 349. An additional polymorphism is the SH antigen that is associated with the FCGR3B3 (FcgammaRIIIB-SH) allele. STUDY DESIGN AND METHODS: By use of a PCR with allele-specific primers, the allelic polymorphisms of FcgammaRIIA and FcgammaRIIIB were determined among 263 unrelated Brazilian subjects, including Amazon Indians (n = 92), blood donors (n = 85), and patients with sickle cell disease (SCD) (n = 86). RESULTS: Amazon Indians had a significantly higher frequency of the R131 allele than did blood donors and SCD patients (0.91 vs. 0.55 vs. 0.55; p<0.001). NA1 and NA2 gene frequencies were found to be 0.67 and 0.21 for Amazon Indians, 0.58 and 0.42 for blood donors, and 0.61 and 0.39 for SCD patients, respectively. The FcgammaRIIIB-SH allele was absent from the Amazon Indians, but 9 (10.6%) blood donors and 10 (11.6%) SCD patients expressed this allele. CONCLUSION: Overall, the data indicate that the distribution of the FcgammaRIIIB alleles is significantly different in Amazon Indians from the distribution in Brazilian blood donors or African Brazilian patients with SCD, but that it is similar to the distributions reported in Asian populations. Moreover, the distribution of the FcgammaRIIA and FcgammaRIIIB alleles among Brazilian blood donors and SCD patients is comparable to the distributions reported in whites from the United States and Europe. PMID- 11099671 TI - HCV lookback in the United States: effectiveness of an extended lookback program. AB - BACKGROUND: In 1998, the FDA recommended look-back for HCV. The recommendation was initially limited, however, to donors who reacted on a multiantigen HCV screening test and to components collected since January 1, 1988. A lookback program was extended to include donors who reacted on the first-generation (single-antigen) HCV screening test and who were positive on a supplemental assay (RIBA-1 or -2) and all components for which transfusion records could be found (back to 1978). STUDY DESIGN AND METHODS: The yield of the incremental lookback programs was compared to that originally recommended by the FDA by comparing the number of newly identified HCV-positive recipients in each program. The results of lookbacks were reviewed on 385 blood components for which 314 transfusion recipients were identified. RESULTS: Of the 135 recipients in the FDA program, 70 percent were dead, 28 percent were living and notified, and 2 percent could not be located. In the incremental programs, there were 179 recipients, of whom 80.4 percent were dead, 16.2 percent were living and notified, and 3.4 percent could not be located. Most adult recipients were dead (81%), but the majority of pediatric recipients were alive (57%); 76 percent of tested recipients were HCV seropositive, with no significant difference between programs. One-half of test positive recipients in each program were newly identified through the lookback program. Seven of the 20 newly identified HCV-positive recipients were found through the incremental programs. The yield, defined as newly detected HCV cases per total number of recipients, was 9.6 percent for the FDA and 3.9 percent for the incremental programs. This difference was significant (p = 0.04). CONCLUSION: The yield of both programs was limited by the high percentage of recipients who had died. Pediatric recipients were more likely to be living at the time of notification. The incremental program was less efficacious than the FDA program in identifying newly HCV-positive recipients, but one-third of the newly detected HCV cases were identified through the incremental program. PMID- 11099672 TI - Surveillance of HIV-1 genetic subtypesand diversity in the US blood supply. AB - BACKGROUND: Recent reports of variant (non-subtype B) HIV infections in US populations have raised concerns about the sensitivity of subtype B virus-based donor screening and diagnostic assays. This study was designed to determine the prevalence and genetic diversity of HIV subtypes in US blood donors over the last two decades. STUDY DESIGN AND METHODS: Three groups were studied: hemophiliacs infected by clotting factor concentrates in the early 1980s (n = 49), blood donors retrospectively identified as being seropositive in 1985 (n = 97), and blood donors identified as seropositive between 1993 and 1996 (n = 405). Subtype assignment was based primarily on heteroduplex mobility analysis (HMA) of HIV-1 env, with DNA sequence confirmation of selected specimens. HIV peptide-based EIA serotyping was used to rule out HIV-2 and group O infections and to serotype HMA refractory specimens. RESULTS: Of 551 specimens, 535 (97%) were assigned subtypes; 532 (99%) of these were subtype B. Three postscreening donations (1%) were assigned non-B subtypes (2 A, 1 C). Two of these three donors were born in Africa; the third was born in the United States and reported no risk factors other than heterosexual activity. HMA distribution plots showed an increase in env diversity among HIV-1 group B strains over time. CONCLUSION: The results support the need for continued surveillance of HIV subtype diversity and ongoing validation of the sensitivity of HIV diagnostic assays to non-B subtype infections. PMID- 11099673 TI - Validation of selected donor-screening questions: structure, content, and comprehension. AB - BACKGROUND: Donor questioning is an integral component of blood safety, designed to identify and interdict donors considered to be at elevated risk of transfusion transmissible infection. Questions are frequently added to this screening procedure, but they are rarely evaluated for comprehension. STUDY DESIGN AND METHODS: Seven American Red Cross blood donor history questions were selected for evaluation. Focus groups were conducted using individuals from high school/college, church, business, and other sources, who had never donated blood. Both sexes and various age and racial groups were represented. The donation process and regulatory requirements or guidelines were explained. Participants were asked to consider 1) content, 2) clarity 3) likelihood of asking for more information, 4) suggestions for improvement, and 5) how a need for clarifying information would best be met. RESULTS: Constructive, helpful, and consistent comments obtained might be utilized in revising questions. CONCLUSION: Focus group discussions can be useful in providing guidance for developing screening questions that can be easily understood. PMID- 11099674 TI - Donor neutrophil function after plateletpheresis. AB - BACKGROUND: Neutrophils are important mediators of inflammation and may be activated by foreign surfaces in apheresis systems. Because most of the WBCs are returned to the donor, it was investigated whether artificial activation leads to altered donor neutrophil function. STUDY DESIGN AND METHODS: Three apheresis systems (Amicus, Autopheresis-C, and CS-3000; all: Baxter Fenwal) were investigated. Preapheresis and postapheresis blood samples were drawn from 10 volunteer donors, with all three apheresis systems used in random order for each donor. Changes in neutrophil phagocytic ability, oxidative burst, and expression of L-selectin and CD11b were measured by flow cytometry, and plasma concentrations of myeloperoxidase and lactoferrin were measured by EIA. Complement activation was evaluated by quantification of C3bc and the terminal complement complex by EIA. RESULTS: Neutrophil expression of L-selectin increased after apheresis (p = 0.02), and the production of oxygen radicals was reduced (p = 0.01). This effect was possibly a result of priming. Complement was not activated. There were no significant differences in neutrophil function after apheresis with any of the three apheresis systems. CONCLUSIONS: Neutrophil function was altered after apheresis, although to a very small extent, and contact between neutrophils and the foreign surface in the apheresis systems is found to be a biotolerant procedure. PMID- 11099675 TI - A novel closed system utilizing styrene copolymer bead adherence for the production of human dendritic cells. PMID- 11099676 TI - Application of 16S ribosomal RNA gene amplification to the rapid identification of bacteria from blood culture bottles. PMID- 11099677 TI - Transmission of HTLV-I by kidney transplant. PMID- 11099678 TI - The first case of HCV seroconversion after 3 years of HCV NAT screening in Baden Wurttemberg. PMID- 11099679 TI - Systolic versus diastolic heart failure in community practice: clinical features, outcomes, and the use of angiotensin-converting enzyme inhibitors. AB - BACKGROUND: Among patients with heart failure, there is controversy about whether there are clinical features and laboratory tests that can differentiate patients who have low ejection fractions from those with normal ejection fractions. The usefulness of angiotensin-converting enzyme (ACE) inhibitors among heart failure patients who have normal left ventricular ejection fractions is also not known. METHODS: From a registry of 2,906 unselected consecutive patients with heart failure who were admitted to 10 acute-care community hospitals during 1995 and 1997, we identified 1291 who had a quantitative measurement of their left ventricular ejection fraction. Patients were separated into three groups based on ejection fraction: < or =0.39 (n = 741, 57%), 0.40 to 0.49 (n = 238, 18%), and > or =0.50 (n = 312, 24%). In-hospital mortality, prescription of ACE inhibitors at discharge, subsequent rehospitalization, quality of life, and survival were measured; survivors were observed for at least 6 months after hospitalization. RESULTS: The mean (+/- SD) age of the sample was 75+/-11 years; the majority (55%) of patients were women. In multivariate models, age >75 years, female sex, weight >72.7 kg, and a valvular etiology for heart failure were associated with an increased probability of having an ejection fraction > or =0.50; a prior history of heart failure, an ischemic or idiopathic cause of heart failure, and radiographic cardiomegaly were associated with a lower probability of having an ejection fraction > or =0.50. Total mortality was lower in patients with an ejection fraction > or =0.50 than in those with an ejection fraction < or =0.39 (odds ratio [OR] = 0.69, 95% confidence interval [CI 0.49 to 0.98, P = 0.04). Among hospital survivors with an ejection fraction of 0.40 to 0.49, the 65% who were prescribed ACE inhibitors at discharge had better mean adjusted quality-of life scores (7.0 versus 6.2, P = 0.02), and lower adjusted mortality (OR = 0.34, 95% CI: 0.17 to 0.70, P = 0.01) during follow-up than those who were not prescribed ACE inhibitors. Among hospital survivors with an ejection fraction > or =0.50, the 45% who were prescribed ACE inhibitors at discharge had better (lower) adjusted New York Heart Association (NYHA) functional class (2.1 versus 2.4, P = 0.04) although there was no significant improvement in survival. CONCLUSIONS: Among patients treated for heart failure in community hospitals, 42% of those whose ejection fraction was measured had a relatively normal systolic function (ejection fraction > or 0.40). The clinical characteristics and mortality of these patients differed from those in patients with low ejection fractions. Among the patients with ejection fractions > or =0.40, the prescription of ACE inhibitors at discharge was associated favorable effects. PMID- 11099680 TI - Outcomes and cost-effectiveness of ventilator support and aggressive care for patients with acute respiratory failure due to pneumonia or acute respiratory distress syndrome. AB - PURPOSE: Many patients with acute respiratory failure die despite prolonged and costly treatment. Our objective was to estimate the cost-effectiveness of providing rather than withholding mechanical ventilation and intensive care for patients with acute respiratory failure due to pneumonia or acute respiratory distress syndrome. SUBJECTS AND METHODS: We studied 1,005 patients enrolled in a five-center study of seriously ill patients (the Study to Understand Prognoses and Preferences for Outcomes and Risks of Treatments [SUPPORT]) with acute respiratory failure (pneumonia or acute respiratory distress syndrome and an Acute Physiology Score > or =10) who required ventilator support. We estimated life expectancy based on long-term follow-up of SUPPORT patients. Utilities were estimated using time-tradeoff questions. Costs (in 1998 dollars) were based on hospital fiscal data and Medicare data. RESULTS: Of the 963 patients who received ventilator support, 48% survived for at least 6 months. At 6 months, survivors reported a median of 1 dependence in activities of daily living, and 72% rated their quality of life as good, very good, or excellent. Among the 42 patients in whom ventilator support was withheld, the median survival was 3 days. Among patients whose estimated probability of surviving at least 2 months from the time of ventilator support ("prognostic estimate") was 70% or more, the incremental cost per quality-adjusted life-year (QALY) saved by providing rather than withholding ventilator support and aggressive care was $29,000. For medium-risk patients (prognostic estimate 51% to 70%), the incremental cost-effectiveness was $44,000 per QALY, and for high-risk patients (prognostic estimate < or =50%), it was $110,000 per QALY. When assumptions were varied from 50% to 200% of baseline estimates, the results ranged from $19,000 to $48,000 for low-risk patients, from $29,000 to $76, 000 for medium-risk patients, and from $67,000 to $200,000 for high-risk patients. CONCLUSIONS: Ventilator support and intensive care for acute respiratory failure due to pneumonia or acute respiratory distress syndrome are relatively cost-effective for patients with >50% probability of surviving 2 months. However, for patients with an expected 2-month survival < or =50%, the cost per QALY is more than threefold greater at >$100,000. PMID- 11099681 TI - Mechanisms of hypertension in patients with chronic obstructive pulmonary disease and acute respiratory failure. AB - PURPOSE: To investigate the effects of hypoxemia, hypercapnia, and cardiovascular hormones (norepinephrine, endothelin-1, and atrial natriuretic factor) on blood pressure during acute respiratory failure. PATIENTS AND METHODS: Patients with chronic obstructive pulmonary disease and acute respiratory failure were divided into four groups of 10 patients each: hypoxemia-normocapnia, hypoxemia hypercapnia, hypoxemia-hypocapnia, and normoxemia-hypercapnia. Plasma norepinephrine levels were determined by high-performance liquid chromatography with electrochemical detection. Plasma endothelin-1 and atrial natriuretic factor levels were radioimmunoassayed after chromatographic preextraction. RESULTS: Systolic blood pressure and cardiovascular hormone levels were greater in patients with hypercapnia (whether or not they also had hypoxemia) than in those with normocapnia and hypoxemia. For example, in patients with hypercapnia and normoxemia, the mean (+/- SD) systolic blood pressure was 183+/-31 mm Hg and the mean norepinephrine level was 494+/-107 pg/mL, as compared with 150+/- 6 mm Hg and 243+/-58 pg/mL in those with normocapnia and hypoxemia (both P<0.05). Similar results were seen for endothelin-1 and atrial natriuretic factor levels, and for the comparisons of hypoxemic patients who were hypercapnic with those who were normocapnic. CONCLUSIONS: These results suggest that blood carbon dioxide levels, rather than oxygen levels, are responsible for hypertension during acute respiratory failure, perhaps as a result of enhanced sympatho-adrenergic activity. PMID- 11099682 TI - Predictors of clinical outcome and radiologic progression in patients with neuropsychiatric manifestations of systemic lupus erythematosus. AB - PURPOSE: We sought to identify the predictors of clinical outcome and of the evolution of cerebral abnormalities in patients with neuropsychiatric systemic lupus erythematosus (SLE). SUBJECTS AND METHODS: Thirty-two patients with SLE (including 14 with the antiphospholipid syndrome) who had been hospitalized with primary neuropsychiatric disease were observed prospectively for at least 2 years. Laboratory and clinical characteristics and data from magnetic resonance imaging (MRI) studies obtained during the hospitalization and 2 years later were evaluated. We ascertained nonreversible or new MRI changes and clinical outcomes, including neuropsychiatric events, during follow-up. RESULTS: Cranial MRI scans on admission were abnormal in 26 (81%) of the 32 patients. Patients with the antiphospholipid syndrome were more likely to have focal cerebral white matter lesions (odds ratio [OR] = 12, 95% confidence interval [CI]: 2.0 to 72). After 2 years, neuropsychiatric deficits substantially improved in 22 (69%) of the patients, stabilized in 6 (19%), and deteriorated in 4 (12%). The number of prior neuropsychiatric events was associated with persistent MRI lesions (OR = 4.8 per each event, 95% CI: 1.1 to 21) and unfavorable clinical outcome (OR = 4.3 per each event, 95% CI: 1.4 to 13) at 2 years. The antiphospholipid syndrome also predicted an unfavorable clinical outcome at 2 years (OR = 11, 95% CI: 1.7 to 65). CONCLUSIONS: Among patients with SLE who have neuropsychiatric disease, prior neuropsychiatric events and the antiphospholipid syndrome increase the risk of adverse outcomes. PMID- 11099683 TI - Association between anticardiolipin antibodies and mortality in patients with peripheral arterial disease. AB - PURPOSE: Anticardiolipin antibodies may be associated with recurrent thromboembolic events in patients with myocardial infarction or stroke. We sought to determine the prevalence of anticardiolipin antibodies in patients with peripheral arterial disease and their association with subsequent thromboembolic events and mortality. METHODS: We ascertained anticardiolipin antibodies using a standardized enzyme-linked immunosorbent assay (immunoglobulin G [IgG] anticardiolipin > or =15 GPL units or IgM anticardiolipin > or =15 MPL units) in 232 patients with peripheral arterial disease and 100 control subjects. Patients were observed to determine overall and cardiovascular mortality, and incident thromboembolic events. RESULTS: IgG anticardiolipin antibodies were significantly more common in the patients with peripheral arterial disease (36 of 232 [16%]) than in the controls (7 of 100 [7%], P = 0.03). During a median follow-up of 3.5 years, 3 of the 232 patients were lost to follow-up and 56 (24%) died. Overall mortality was significantly greater in the IgG anticardiolipin-positive patients (16 of 35 [46%]) compared with those who were IgG anticardiolipin-negative (40 of 194 [21%], P = 0.0003), largely due to an increase in cardiovascular mortality among the IgG anticardiolipin-positive patients. In a multivariate proportional hazards analysis, IgG anticardiolipin antibodies were an independent risk factor for overall mortality (hazard ratio [HR] = 2.1, 95% confidence interval [CI]: 1.2 to 4.0) and cardiovascular mortality (HR = 4.4, 95% CI: 1.6 to 12). CONCLUSIONS: IgG anticardiolipin antibodies are common in patients with peripheral arterial disease and are associated with an increased risk of overall and cardiovascular mortality. PMID- 11099684 TI - The natural history of incidental renal artery stenosis in patients with aortoiliac vascular disease. AB - PURPOSE: To examine the association between incidentally discovered renal artery stenosis and deterioration of renal function as determined by the change in serum creatinine concentration over time. SUBJECTS AND METHODS: We performed a retrospective review of consecutive patients who underwent aortography for aortoiliac vascular disease. Angiograms were reviewed for renal artery stenosis, defined as a narrowing of at least 20% compared with adjacent normal renal artery. For patients with at least 180 days of subsequent follow-up, the change in serum creatinine concentration per year was compared in patients who had or did not have renal artery stenosis. RESULTS: Of the 201 patients, 96 (48%) had some degree of renal artery stenosis in one or both renal arteries, including 53 (26%) who had at least one stenosis > or= 50% and 40 (20%) who had bilateral stenoses. The only clinical predictor of renal artery stenosis was a history of coronary artery disease (odds ratio = 2.0, 95% confidence interval: 1.2 to 3.8, P = 0.001). Among the 174 patients with > or =180 days of follow-up, there was no statistically significant difference (P = 0.88) in the mean change in serum creatinine concentration per year in the 78 patients with renal artery stenosis (0.06+/-0.33 mg/dL per year) as compared with the 96 patients without renal artery stenosis (0.06+/-0.22 mg/dL per year). Grouping the patients by the maximal percentage of stenosis did not reveal any difference in the mean changes in serum creatinine concentration per year. CONCLUSIONS: Although renal artery stenosis is a common incidental finding in patients with atherosclerotic vascular disease, it is an uncommon cause of progressive renal disease. PMID- 11099685 TI - Physician attitudes toward and prevalence of the hospitalist model of care: results of a national survey. AB - PURPOSE: We sought to determine the availability and utilization of, as well as physician attitudes toward, the hospitalist model in the United States. SUBJECTS AND METHODS: Using a telephone survey, we asked physicians who were board certified in internal medicine about their inpatient practice arrangements, the availability of hospitalist services, and their attitudes toward the hospitalist model. All physicians were generalists in active clinical practice. Using multivariable methods, we determined factors associated with attitudes toward the hospitalist model. RESULTS: We were able to contact 787 of 2,829 physicians who were randomly selected from a national list of board-certified internists, of whom 400 agreed to participate. Most respondents were familiar with the term "hospitalist" and had hospitalist services available in their community, and 28% used hospitalists for their inpatients. Few (2%) reported the presence of the "mandatory" hospitalist model. Physicians reported that the model was more commonly available in Western states (84% vs. 55% to 63% in other regions, P<0.0001). Seventy-three percent thought hospitalist systems would reduce continuity of care. Only 28% thought that patients would prefer care from an inpatient specialist, but 51% thought patients might get better care, and 47% thought patients might get more cost-effective care in a hospitalist system. In multivariable models, physicians who were in solo practice, those in specialties with more inpatient practice, and those who had more patients hospitalized each month responded more negatively about the model, whereas those with hospitalists in their community were more positive. CONCLUSIONS: Although agreeing that quality of care and efficiency might be improved, physicians were concerned about patient-doctor relationships and patient satisfaction in a hospitalist model. Future studies should determine the effect of the hospitalist model on these outcomes. PMID- 11099686 TI - Pygeum africanum for the treatment of patients with benign prostatic hyperplasia: a systematic review and quantitative meta-analysis. AB - PURPOSE: To conduct a systematic review and quantitative meta-analysis of the therapeutic efficacy and tolerability of Pygeum africanum in men with symptomatic benign prostatic hyperplasia. METHODS: Studies were identified through the search of Medline (1966 to 2000), Embase, Phytodok, the Cochrane Library, bibliographies of identified trials and review articles, and contact with relevant authors and drug companies. Randomized trials were included if participants had symptomatic benign prostatic hyperplasia, the intervention was a preparation of P. africanum alone or in combination with other phytotherapeutic agents, a control group received placebo or other pharmacologic therapies for benign prostatic hyperplasia, and treatment duration was at least 30 days. Two investigators independently extracted key data on design features, subject characteristics, and therapy allocation. RESULTS: A total of 18 randomized controlled trials involving 1,562 men met the inclusion criteria and were analyzed. Many studies did not report results in a method that permitted meta-analysis. Only 1 of the studies reported a method of treatment allocation concealment, although 17 were double blinded. The mean study duration was 64 days (range 30 to 122). Compared with placebo in 6 studies, P. africanum provided a moderately large improvement in the combined outcome of urologic symptoms and flow measures as assessed by an effect size defined by the difference of the mean change for each outcome divided by the pooled standard deviation for each outcome (-0.8 SD [95% confidence interval (CI): -1.4 to -0.3]). Summary estimates of individual outcomes were also improved by P. africanum. Men were more than twice as likely to report an improvement in overall symptoms (risk ratio = 2.1, 95% CI: 1.40 to 3.1). Nocturia was reduced by 19% and residual urine volume by 24%; peak urine flow was increased by 23%. Adverse effects due to P. africanum were mild and similar to placebo. The overall dropout rate was 12% and was similar for P. africanum (13%), placebo (11%), and other controls (8%; P = 0.4 versus placebo and P = 0.5 versus other controls). CONCLUSIONS: The literature on P. africanum for the treatment of benign prostatic hyperplasia is limited by the short duration of studies and the variability in study design, the use of phytotherapeutic preparations, and the types of reported outcomes. However, the evidence suggests that P. africanum modestly, but significantly, improves urologic symptoms and flow measures. Further research is needed using standardized preparations of P. africanum to determine its long-term effectiveness and ability to prevent complications associated with benign prostatic hyperplasia. PMID- 11099687 TI - Reactive oxygen species and acute renal failure. AB - Acute renal failure is commonly due to acute tubular necrosis (ATN), the latter representing an acute, usually reversible loss of renal function incurred from ischemic or nephrotoxic insults occurring singly or in combination. Such insults instigate a number of processes-hemodynamic alterations, aberrant vascular responses, sublethal and lethal cell damage, inflammatory responses, and nephron obstruction-that initiate and maintain ATN. Eventually, reparative and regenerative processes facilitate the resolution of renal injury and the recovery of renal function. Focusing mainly on ischemic ATN, this article reviews evidence indicating that the inordinate or aberrant generation of reactive oxygen species (ROS) may contribute to the initiation and maintenance of ATN. This review also discusses the possibility that ROS may instigate adaptive as well as maladaptive responses in the kidney with ATN, and raises the possibility that ROS may participate in the recovery phase of ATN. PMID- 11099688 TI - Extreme hyponatremia of 87 mmol/l without neurologic complications in a severely hypovolemic patient. PMID- 11099689 TI - Case of the month AB - Each month, we will present a challenging Case of the Month for Green Journal readers, who must use their clinical acumen to arrive at the correct answer. We will also post the case each month on the Journal's web site (http://www.ajmselect.com). Several possible answers may be consistent with the case presentation; use your best judgment. Please send your answer (one per respondent) to the Green Journal at editors@amjmed.org or via FAX to (415) 447 2799. Indicate the case to which you are responding, and include your complete address. The correct answer will appear in the next issue of the Journal. The first five persons who submit correct answers will receive a free one-year subscription to the Journal. Because of the volume of answers we receive, neither correct nor incorrect answers can be individually acknowledged. Colleagues of Drs. Chow, Chan, and Szeto at the Chinese University of Hong Kong are not eligible for this month's case. If you would like to contribute a case, please submit a brief synopsis (<250 words) to the editorial office. PMID- 11099690 TI - Diastolic heart failure: miles to go before we sleep. PMID- 11099691 TI - Another perspective on SUPPORT. PMID- 11099692 TI - Pathogenesis and management of hyponatremia. PMID- 11099693 TI - Preparing residents to become more effective teachers: a priority for internal medicine. PMID- 11099694 TI - Cilostazol prevents impairment of slow axonal transport in streptozotocin diabetic rats. AB - We studied the effects of cilostazol, an antiplatelet and vasodilating agent, on axonal transport patterns of cytoskeletal proteins in the motor fibers of sciatic nerve of streptozotocin-induced diabetic rats. Proteins labeled with L [35S]methionine in 6-mm consecutive segments of the nerve were analyzed electrophoretically following fractionation into Triton-soluble and-insoluble subpopulations. Transport rates of proteins (particularly neurofilaments) in slow component a were reduced by 50% 2 weeks after labeling (4 weeks after streptozotocin). An apparent reduction of tubulin and actin was observed at later intervals after induction of diabetes. Actin transported in slow component b was also impaired, though to a lesser extent than in component a. Cilostazol prevented transport impairment of both slow components a and b without affecting hyperglycemia or reduction in body weight gain. These results suggest that in sciatic motor fibers early defects in slowly transported proteins are more marked in slow component a, and that impairment may be caused primarily by hemodynamic abnormalities. PMID- 11099695 TI - Cytotoxicity and cell cycle effects of the plant alkaloids cryptolepine and neocryptolepine: relation to drug-induced apoptosis. AB - Cryptolepine and neocryptolepine are two indoloquinoline derivatives isolated from the roots of the african plant Cryptolepis sanguinolenta. These two alkaloids, which only differ by the respective orientation of their indole and quinoline rings, display potent cytotoxic activities against tumour cells and present antibacterial and antiparasitic properties. Our previous molecular studies indicated that these two natural products intercalate into DNA and interfere with the catalytic activity of human topoisomerase II. Here we have extended the study of their mechanism of action at the cellular level. Murine and human leukemia cells were used to evaluate the cytotoxicity of the drugs and their effects on the cell cycle were measured by flow cytometry. Cryptolepine, and to a lesser extent neocryptolepine, provoke a massive accumulation of P388 murine leukemia cells in the G2/M phase. With HL-60 human leukemia cells, the treatment with cryptolepine leads to the appearance of a hypo-diploid DNA content peak (sub-G1) characteristic of the apoptotic cell population. With both P388 and HL-60 cells, cryptolepine proved about four times more toxic than its isomer. But the use of the HL-60/MX2 cell line resistant to the anticancer drug mitoxantrone suggests that topoisomerase II may not represent the essential cellular target for the alkaloids, which are both only two times less toxic to the resistant HL 60/MX2 cells compared to the parental cells. The capacity of the drugs to induce apoptosis of HL-60 human leukemia cells was examined by complementary biochemical techniques. Western blotting analysis revealed that cryptolepine, but not neocryptolepine, induces cleavage of poly(ADP-ribose) polymerase but both alkaloids induce the release of cytochrome c from the mitochondria. The cleavage of poly(ADP-ribose) polymerase observed with cryptolepine correlates with the appearance of a marked sub-G1 peak in the cell cycle experiments. The proteolytic activity of Asp-Glu-Val-Asp- or Ile-Glu-Thr-Asp-caspases was found to be enhanced much more strongly with cryptolepine than with its isomer, as expected from their different cytotoxic potential. Despite the activation of the caspase cascade, we did not detect internucleosomal cleavage of DNA in the HL-60 cells treated with the alkaloids. Altogether, the results shed light on the mechanism of action of these two plant alkaloids. PMID- 11099696 TI - Role of glucose and insulin in thiazolidinedione-induced alterations in hepatic gluconeogenesis. AB - Previous studies from our laboratory as well as from others have suggested that the thiazolidinediones have the capacity to act as insulinomimetic agents, especially in the liver. In order to further characterize this insulinomimetic action, we evaluated the effect of troglitazone, a representative thiazolidinedione, on lactate- and glucagon-stimulated gluconeogenesis, in the presence or absence of insulin (10 nM) in isolated rat hepatocytes. The antigluconeogenic effect of troglitazone under basal (lactate-stimulated) conditions was found to be due to an elevation in the fructose 2,6-bisphosphate content, which was, however, not mediated by an activation of 6-phosphofructo 2 kinase. Troglitazone (125 and 250 microM) in the absence of insulin, produced a dose-dependent reduction in glucagon-stimulated gluconeogenesis, thereby suggesting an insulinomimetic effect. In addition, troglitazone (125 and 250 microM), in combination with insulin, produced an additive inhibition of gluconeogenesis during glucagon-stimulated conditions. However, unlike insulin, the metabolic mechanism responsible for these effects (in the presence or absence of insulin) does not involve fructose 2,6-bisphosphate. PMID- 11099697 TI - Interaction between methotrexate and nonsteroidal anti-inflammatory drugs in organic anion transporter. AB - The antifolate drug methotrexate is mainly eliminated from the body by renal tubular secretion via organic anion transporters. In clinical situations, severe methotrexate toxicity, due to an increase in serum concentrations, was observed after coadministration with nonsteroidal anti-inflammatory drugs (NSAIDs) or probenecid. In this study, we examined the effects of NSAIDs and probenecid on methotrexate transport via the rat renal organic anion transporter rOAT1, using Xenopus laevis oocytes. [3H]Methotrexate uptake was markedly stimulated in the rOAT1 cRNA-injected oocytes, and this uptake was inhibited by probenecid and various NSAIDs, whereas the influence of salicylate was less. The Dixon plots showed that probenecid, indomethacin and salicylate competitively inhibited rOAT1 with apparent K(i) values of 15.8 microM, 4.2 microM and 1.0 mM, respectively. These findings demonstrate that rOAT1 is the major site of the transporter mediated interaction between methotrexate and NSAIDs and/or probenecid, leading to a decrease in renal excretion of methotrexate. PMID- 11099698 TI - Effect of intrathecal administration of serotonin in chronic pain models in rats. AB - The present study examined the effects of intrathecal (i.t.) administration of 5 hydroxytryptamine (5-HT; 0.1-100 microg) on mechanical hyperalgesia associated with neuropathic pain (chronic constriction of the sciatic nerve model and diabetic model) and inflammatory pain (carrageenan and polyarthritic models) in rats. Results demonstrated that the hyperalgesia observed in the mononeuropathic and diabetic rats was attenuated by 5-HT; the active dose, however, was 100- to 1000-fold higher than that required in normal rats, and was moderately effective. In the two experimental models of inflammatory pain, 5-HT was not markedly or similarly active. In the carrageenan model, 5-HT at the highest dose was only weakly effective whereas in the polyarthritic model it was inactive. Together, these results show that 5-HT has antinociceptive effects in several rat pain models, except in the model of diffuse pain (polyarthritic rats). Its antinociceptive effects in these models, however, are slight and differ from those observed in normal rats. PMID- 11099699 TI - TC-2559: a novel orally active ligand selective at neuronal acetylcholine receptors. AB - TC-2559 [(E)-N-Methyl-4-[3-(5-ethoxypyridin)yl]-3-buten-1-amine] is a novel nicotinic agonist markedly more selective than recently reported novel nicotinic receptor ligands (selectivity ratio for central nervous system (CNS) to peripheral nervous system (PNS)>4000). TC-2559 competes effectively with [3H] nicotine binding (K(i)=5 nM) but not with [125I]-bungarotoxin (>50,000 nM). Dopamine release from striatal synaptosomes and ion flux from thalamic synaptosomes indicate that TC-2559 is potent and efficacious in the activation of CNS receptors and significantly reduced glutamate-induced neurotoxicity in vitro. TC-2559 has no detectable effects on muscle and ganglion-type nicotinic acetylcholine receptors at concentrations up to 1 mM. TC-2559 significantly attenuates scopolamine-induced cognitive deficits in a step-through passive avoidance task. Acute and repeated oral dosing of TC-2559 enhances performance in a radial arm maze task. In contrast to the effects of equimolar concentrations of (-) nicotine, TC-2559 does not induce hypothermia and locomotor activity is not enhanced following repeated daily administration of 14 days. TC-2559 has a markedly enhanced CNS-PNS selectivity ratio and an intra-CNS selectivity as evidenced by the improved cognition without increased locomotor activity. The in vitro and in vivo studies in the present study suggest that TC-2559 has the desired profile to be further evaluated as a potential therapeutic agent for neurodegenerative diseases. PMID- 11099700 TI - Effects of nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester on phencyclidine-induced effects in rats. AB - Phencyclidine (PCP) is widely used as an animal model of schizophrenia. In rats, acute PCP treatment increased locomotor activity and induced stereotyped behaviours consisting of head weaving, turning and backpedalling. PCP had differential regional effects on c-fos expression in rat brain, suggesting different patterns of neuronal activity. The most prominent immunostaining was observed in the cortical regions. To elucidate the role of nitric oxide, an important intracellular messenger, in the mechanism of action of PCP the effects of nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) were studied in PCP-treated animals. L-NAME potentiated PCP-induced behaviours and c-fos expression in many brain regions. The greatest increases were observed in the frontal, retrosplenial granular cortex, cerebellum, thalamic and subthalamic nuclei. While PCP alone induced low c-fos expression in the entorhinal cortex, with almost no expression in the rostral part of caudate putamen, animals pretreated with L-NAME showed marked activation in these brain areas. These results strongly indicate the involvement of the nitric oxide system in the mechanism of action of PCP. PMID- 11099701 TI - Increased plasma corticosterone, aggressiveness and brain monoamine changes induced by central injection of pertussis toxin. AB - The effects of intracerebroventricular (i.c.v.) injection of pertussis toxin, a specific inhibitor of G(i)/G(o) proteins, on plasma corticosterone levels, aggressiveness, and hypothalamic and hippocampal monoamines and their metabolites levels were examined in mice. Plasma corticosterone level was markedly increased at 3 h after pertussis toxin injection (0.03 and 0.2 microg/mouse), peaked at 6 h and was still increased for up to 6 days after injection. Mice injected with pertussis toxin (0.2 microg/mouse) did not show weight gain between day 0 and day 6 after injection. In addition, pertussis toxin (0.2 microg/mouse) induced a progressive increase in aggressiveness, i.e. a decrease in attack latency and an increase in number of attacks, on day 1 and 6 after injection. Brain monoamines and their metabolites levels were changed on day 1 and 6 after pertussis toxin injection (0.2 microg/mouse): in the hypothalamus, levels of dopamine and 3,4 dihydroxyphenylacetic acid were increased, norepinephrine level decreased, and 5 hydroxyindole acetic acid (5-HIAA) level was markedly increased, with no changes in 5-hydroxytryptamine (5-HT) level, whereas in the hippocampus, 5-HT level was significantly decreased, with no changes in 5-HIAA and catecholamines. These results suggest that signal transduction through G(i)/G(o) proteins in the brain is involved in the modulation of hypothalamo-pituitary-adrenal axis, aggressiveness, and monoamine levels in vivo. PMID- 11099702 TI - The effect of the dopamine D2 receptor antagonist raclopride on the pattern of licking microstructure induced by midazolam in the rat. AB - The role of dopamine in the effects of midazolam on ingestive behaviour was investigated using microstructural analysis of licking behaviour in the rat. Midazolam (1.8 mg/kg i.p.) was administered alone or in combination with the dopamine D2 receptor antagonist raclopride (0.1 and 0.3 mg/kg i.p.). The effect on licking patterns during 60 s exposure to a range of concentrations of sucrose solution was recorded using an automated lickometer. Midazolam increased the total number of licks via an increase in mean bout duration, an effect consistent with the proposal that these drugs enhance palatability. Midazolam also decreased the intrabout lick rate, probably because of muscle relaxant effects. Pre treatment with raclopride blocked midazolam-induced increases in mean bout duration, at doses that by themselves were ineffective, but did not reverse the decrease in intrabout lick rate. These data point to the interdependence of benzodiazepine and dopamine substrates in the mediation of palatability. PMID- 11099703 TI - Pretreatment with diazepam suppresses the reduction in defensive freezing behavior induced by fluvoxamine in the conditioned fear stress paradigm in mice. AB - The effects of the selective serotonin (5-hydroxytryptamine (5-HT)) reuptake inhibitor fluvoxamine, given alone or in combination with the benzodiazepine anxiolytic diazepam on the defensive freezing behavior of mice in the conditioned fear stress paradigm were examined. Fluvoxamine (5-20 mg/kg, i.p.) induced a dose dependent reduction in freezing behavior. In contrast, while low doses of diazepam (0.125 and 0.25 mg/kg, i.p.) reduced the freezing behavior, such effects were not observed with high doses of diazepam (0.5 and 1 mg/kg, i.p.). In the combination study, fluvoxamine (20 mg/kg, i.p. ) did not reduce the freezing behavior in mice that had been pretreated with diazepam (0.125-1 mg/kg, i.p.). None of the doses of fluvoxamine and diazepam used in the present study had any effects on motor activity under non-stressed conditions. These results suggest that benzodiazepines may negatively influence the clinical efficacy of selective 5-HT reuptake inhibitors in the treatment of anxiety disorders. PMID- 11099704 TI - Nitric oxide synthase-independent release of nitric oxide induced by KCl in the perfused mesenteric bed of the rat. AB - The aim of the present study was to test whether the contractile responses elicited by KCl in the rat mesenteric bed are coupled to the release of nitric oxide (NO). Contractions induced by 70 mM KCl were coincident with the release of NO to the perfusate. The in vitro exposure to the nitric oxide synthase (NOS) inhibitor L-N(omega)-nitro-L-arginine methyl ester, L-NAME (1-100 microM) potentiated the vascular responses to 70 mM KCl and, unexpectedly, increased the KCl-stimulated release of NO. Moreover, even after the chronic treatment with L NAME (70 mg/kg/day during 4 weeks), the KCl-induced release of NO was not reduced, whereas the potentiation of contractile responses was indeed achieved. The possibility that NOS had not been completely inhibited under our experimental conditions can be precluded because NOS activity was significantly inhibited after both L-NAME treatments. After the in vitro treatment with 1 to 100 microM L NAME, the inhibition of NOS was concentration-dependent (from 50% to 90%). With regard to the basal release of NO, the inhibition caused by L-NAME was not concentration-dependent and reached a maximum of 40%, suggesting that basal NO outflow is only partially dependent on NOS activity. An eventual enhancement of NOS activity caused by KCl was disregarded because the activity of this enzyme measured in homogenates from mesenteric beds perfused with 70 mM KCl was significantly reduced. On the other hand, endothelium removal, employed as a negative control, almost abolished NOS activity, whereas the incubation with the Ca(2+) ionophore A23187, employed as a positive control, induced an increase in NOS activity. It is concluded that in the mesenteric arterial bed of the rat, the contractile responses elicited by depolarization through KCl are coincident with a NOS-independent release of NO. This observation, which differs from the results obtained with noradrenaline, do not support the use of KCl as an alternative contractile agent whenever the participation of NO is under study. PMID- 11099705 TI - Primary CNS lymphoma: clinical presentation, pathological classification, molecular pathogenesis and treatment. AB - Primary CNS lymphomas (PCNSL) represent malignant non-Hodgkin's B cell lymphomas, which are confined to the central nervous system. They show a dramatic increase in frequency in the immunocompromised as well as in the immunocompetent population. Recent studies have identified germinal center B cells as the cellular origin of PCNSL; however, the details of their molecular pathogenesis still remain to be elucidated. Treatment recommendations are not clearly established. Radiotherapy (RT) is efficient in terms of tumor response, but not curative. Median survival after RT alone is about 1 year. According to the results of uncontrolled studies the combination of RT and chemotherapy based on high-dose methotrexate (HD-MTX) is most efficient in terms of survival rates. However, long-term neurotoxicity overshadows treatment efficacy, especially in patients over 60 years of age. The authors favor the systematic evaluation of chemotherapy alone with protocols including HD MTX, because unicenter results are promising in terms of both survival as well as quality of life in long term survivors. PMID- 11099706 TI - Cobalt-55 positron emission tomography in late-onset epileptic seizures after thrombo-embolic middle cerebral artery infarction. AB - The pathogenesis of late-onset epileptic seizures after thrombo-embolic cerebral infarction is poorly understood. Our previous positron emission tomographic (PET) studies with 15O have demonstrated that post-apoplectic epilepsy is associated with more severe brain ischemia, but we were unable to determine if this was the cause or the consequence of the seizures. Using cobalt-55 (55Co) as PET tracer we can now distinguish recurrent, recent infarction in patients with a previous old infarct in the same vascular territory. In seven out of twelve patients with post apoplectic seizures an increased uptake of 55Co was observed in the border area and in two of them also within the old infarct core. In the control group, composed of eight seizure-free patients with also an old infarct involving the cortical territory of the middle cerebral artery, no increase in 55Co uptake was observed on PET examination. The present study indicates that in a significant number of patients late-onset epilepsy is the clinical expression of recurrent strokes, occurring in the same vascular territory. PMID- 11099707 TI - Comparison of regional cerebral blood flow and glucose metabolism in the normal brain: effect of aging. AB - The regional cerebral blood flow (rCBF) and metabolic rate for glucose (rCMRGlc) are associated with functional activity of the neural cells. The present work reports a comparison study between rCBF and rCMRGlc in a normal population as a function of age. 10 young (25.9+/-5.6 years) and 10 old (65.4+/-6.1 years) volunteers were similarly studied at rest. In each subject, rCBF and rCMRGlc were measured in sequence, during the same session. Both rCBF and rCMRGlc values were found to decrease from young (mean rCBF=43.7 ml/100 g per min; mean rCMRGlc=40.6 micromol/100 g per min) to old age (mean rCBF=37.3 ml/100 g per min; mean rCMRGlc=35.2 micromol/100 g per min), resulting in a drop over 40 years of 14.8% (0.37%/year) and 13.3% (0.34%/year), respectively. On a regional basis, the frontal and the visual cortices were observed to have, respectively, the highest and the lowest reduction in rCBF, while, for rCMRGlc, these extremes were observed in striatum and cerebellum. Despite these differences, the ratio of rCBF to rCMRGlc was found to have a similar behavior in all brain regions for young and old subjects as shown by a correlation coefficient of 88%. This comparative study indicates a decline in rCBF and rCMRGlc values and a coupling between CBF and CMRGlc as a function of age. PMID- 11099708 TI - Use of the auditory brainstem response testing in the clinical evaluation of the patients with diabetes mellitus. AB - The objective of the study was to assess whether a relationship exists between the auditory brain stem response (ABR) results and diabetes mellitus with and without complications. In the clinical and audiometry laboratory settings, diabetic patients with and without complications (retinopathy and/or nephropathy) were examined using ABR testing, and the results were interpreted for their applicability in clinical practice. Fifty-nine patients with diabetic retinopathy or nephropathy (study group) and 20 diabetic patients without any known diabetic complication (control group) were assessed with audiometry and ABR testing. ABR revealed that the absolute latencies and interwave intervals of the waves I through V were prolonged significantly in the study group when compared to the control group. The amplitudes of waves I through V were diminished in the study group when compared to the control group, but a statistical significance was present only for wave V amplitude. Quantitative (wave I to wave V amplitude ratio) and qualitative analyses of the ABR waves showed abnormal waveforms in the study and control groups by 55.2 and 27.6%, respectively. There is a brain stem neuropathy in diabetes mellitus which can be assessed with ABR testing. The likelihood of encountering a diabetic complication increases as the ABR results become abnormal. PMID- 11099709 TI - Benefits of glatiramer acetate on disability in relapsing-remitting multiple sclerosis. An analysis by area under disability/time curves. The Copolymer 1 Multiple Sclerosis Study Group. AB - New immunomodulatory therapies for relapsing-remitting multiple sclerosis (RRMS) have well-documented effects in reducing relapses, but it has been difficult to demonstrate their benefits on disability in relatively short treatment trials. Commonly utilised disability outcome measures are problematic both in usefulness and clinical interpretation when applied to MS subjects with fluctuating and variable disease courses. An alternative technique is to use the summary measure 'area under the disability/time curve' (AUC) to index the total in-trial morbidity experienced by patients. In this study, we applied AUC analyses to the serial Expanded Disability Status Scale (EDSS) scores from the U.S. multicentre, Phase III, two-year core study of glatiramer acetate in 251 RRMS patients. When all available EDSS evaluations were analysed with AUC(CHANGE) ('combined data', including relapse-related assessments), active treatment was significantly superior to placebo (P=0.018). The benefits of glatiramer acetate persisted when transient relapse effects were reduced by using 'scheduled visit data' only (P=0.021). With the more conservative AUC(SUM) measure, significant active treatment effects remained (P=0.029 and 0.046, for both 'combined' and 'scheduled visit' data, respectively). Subgroup calculations performed with baseline disability stratified at EDSS 3.5 also showed benefits of treatment over placebo, but statistical significance was not reached. This analysis of data from a Phase III treatment trial illustrates the AUC summary measure technique and provides further evidence of the efficacy of glatiramer acetate in RRMS. PMID- 11099710 TI - Neuromyotonia: autoimmune pathogenesis and response to immune modulating therapy. AB - BACKGROUND: Neuromyotonia (NMT) has been postulated to be an autoimmune channelopathy, probably by affecting voltage gated potassium channels (VGKC) leading to excitation and abnormal discharges [Sinha et al., Lancet 338 (1991) 75]. OBJECTIVE: To report three patients with NMT who had other associated immune mediated conditions, i.e., myasthenia gravis, thymoma and various types of peripheral neuropathies. One patient had peripheral neuropathy and involvement of pre- and post-synaptic neuromuscular junction. RESULTS: All three patients had evidence of polyneuropathy and neuromyotonic discharges on electrodiagnostic studies. Elevated acetylcholine receptor antibodies were noted in all patients and malignant thymoma was found in two patients with metastasis. All three patients showed moderate to marked response to plasma exchange. CONCLUSIONS: These findings strongly suggest a humoral autoimmune pathogenesis of NMT, probably by K(+) channel involvement, affecting acetylcholine quantal release and postsynaptic membrane. Clinicians should be aware of this association of immune mediated conditions in NMT patients and marked improvement with plasma exchange. PMID- 11099711 TI - Inhibition of glutamate uptake into synaptic vesicles of rat brain by the metabolites accumulating in maple syrup urine disease. AB - Maple syrup urine disease is an inherited metabolic disorder characterized by tissue accumulation of branched-chain amino acids and their corresponding keto acids in the affected children. Although this disorder is predominantly characterized by neurological symptoms, only few studies were carried out to investigate its neuropathology. In this study we investigated the effect of the metabolites accumulating in maple syrup urine disease on the in vitro uptake of [3H]glutamate by synaptic vesicles of rat brain. Synaptic vesicle preparations from whole brain of male adult Wistar rats (200-250 g) were incubated with the branched-chain amino acids and their corresponding keto acids at final concentrations ranging from 0.25 to 10 mM for the determination of glutamate uptake. Glutamate uptake was significantly inhibited by L-leucine, L-isoleucine, L-2-ketoisocaproic acid and L-2-keto-3-methylvaleric acid by approximately 60%, whereas L-valine and L-2-ketoisovaleric acid showed no effect. We also verified that the metabolites probably act by competitive inhibition. Therefore, it is possible that extracellular glutamate levels may be increased in maple syrup urine disease and that excitotoxicity may be involved in the neuropathology of this disorder. PMID- 11099712 TI - Visual working memory revealed by repetitive transcranial magnetic stimulation. AB - We evaluated whether repetitive transcranial magnetic stimulation (rTMS) could be utilized for studying the hemispheric lateralization and anatomical localization of the cortical areas of the visual system that are concerned with object-related visual working memory. In eight normal volunteers, visual working memory was tested during rTMS delivery over nine regions in each hemisphere. Visual working memory was significantly disturbed by rTMS over the right hemisphere compared with the left (P<0.05). The disturbance in visual working memory by rTMS was significant over the right inferior frontal (F8), inferior temporal (T8), and middle parietal (P4) areas compared with the control region (P<0.05). This study suggests that visual working memory is lateralized to the right hemisphere and localized in the right inferior frontal, inferior temporal, and middle-parietal areas. As a non-invasive tool, rTMS may be useful for the functional localization of the working memory system. PMID- 11099713 TI - Seasonal patterns in optic neuritis and multiple sclerosis: a meta-analysis. AB - To quantify and characterize seasonal variation in monosymptomatic optic neuritis (MON) onsets, multiple sclerosis (MS) onsets and MS exacerbations (MSE), a meta analysis was performed, using established methods and pooling weighted information obtained from nine reports on MON, six reports on MS onsets and nine reports on MSE, which fulfilled specific criteria for report quality and data homogeneity. The results suggested that MON, MS onsets and MSE in the Northern hemisphere present a similar pattern with highest frequencies in spring and lowest in winter. These differences were highest for MS onsets, 45% with 95% CI 36-55%, and lowest for MSE, 10% with 95% CI 7-13%, statistically significant and robust, insensitive to an alternative seasonal definition, not unduly influenced by any single primary study, and supported by fail-safe N calculations. Random variation, misclassification and publication bias were less likely to account for the reported generalized seasonal patterns. PMID- 11099714 TI - Piracetam versus acetylsalicylic acid in secondary stroke prophylaxis. A double blind, randomized, parallel group, 2 year follow-up study. AB - Piracetam has been shown to inhibit platelet aggregation. Therefore, we performed a double-blind, randomized, parallel group study to compare the efficacy of daily 1600 mg piracetam t.i.d. vs. 200 mg acetylsalicylic acid (ASA) t.i.d. in secondary stroke prophylaxis. 563 patients after stroke as confirmed by computed tomography (CT) or magnetic resonance imaging (MRI) were enrolled and received either piracetam or ASA during a 2 year follow-up period. The primary endpoint was the rate of stroke, transient ischaemic attack (TIA), or death from vascular cause. The secondary endpoint was the rate of adverse events leading to a premature discontinuation of the study medication. Patients were visited at home every 3 months and were examined in hospital after 1 and 2 years. At every visit, the platelet function was evaluated. No significant difference and no significant equivalence could be shown for the primary endpoint between the piracetam and the ASA group both in the intention-to-treat and in the per-protocol analysis. However, there was a not significant trend in favor of ASA (11.7 vs. 15.2%). After excluding those patients who did not respond to antiplatelet medication in vitro, however, piracetam and ASA were equivalent in secondary stroke prophylaxis (stroke, TIA, or vascular death 10.1% in the piracetam group vs. 9.7% in the ASA group). Piracetam was significantly superior to ASA in the secondary endpoint (P=0.0039). The data suggest that the overall efficacy of piracetam in secondary stroke prophylaxis is not as good as that of ASA but that piracetam is better tolerated. However, our data furthermore show that nonresponders to pharmacological inhibition of platelet function are more frequent under piracetam therapy and that they may influence the results of large studies on secondary prophylaxis in vascular diseases. PMID- 11099715 TI - Does diffusion-weighted magnetic resonance imaging enable detection of early ischemic change following transient cerebral ischemia? AB - To examine the usefulness of diffusion-weighted imaging for detecting neuronal damage following ischemia, dynamic changes in diffusion-, T1- and T2-weighted images of rats subjected to 10 min of 4-vessel occlusion and of humans who had suffered 10-20 min of cardiac arrest were observed. In rats, no remarkable alteration was observed on day 1. On day 3, however, diffusion-weighted images showed high signal intensity in the hippocampal area, in which the apparent diffusion coefficient was significantly lower than that of the control (760+/ 28x10(-6) mm(2)/s in control vs. 480+/-29x10(-6) mm(2)/s on day 3, P<0.0001). Histological observation revealed microvacuolation in 92+/-4% of pyramidal neurons in the CA1 region. On day 7, the hyperintensity in diffusion-weighted images had disappeared and microvacuolation had also disappeared in the CA1 region, but severely disrupted pyramidal neurons containing pyknotic nuclei had appeared in the CA1 region instead. In humans, diffusion-weighted images did not show any apparent abnormality in the cerebral cortex on the day of resuscitation. On day 3, however, diffusion-weighted images consistently showed hyperintensities in the temporal or occipital cortex, and these hyperintensities had disappeared in images obtained on days 7 and 14. From day 14, T1-weighted images showed laminar hyperintensity, suggesting laminar necrosis, along the cortex, where diffusion-weighted images showed high signal intensity on day 3. These results suggested that diffusion-weighted imaging has a potential for detection of the occurrence of microvacuolation and is useful for detecting the progression of ischemic changes in humans following global ischemia. PMID- 11099716 TI - Activation of thrombosis and fibrinolysis following brain infarction. AB - To clarify the sequence of alterations in the thrombotic and fibrinolytic systems after acute brain infarction, we prospectively examined sequential changes in coagulatory markers in 38 patients suffering from cardioembolic infarcts (CEI), 41 patients with atherothrombotic infarcts (ATI), 58 patients with lacunar infarcts (LI), and 32 age-matched controls. The plasma level of thrombin antithrombin III complex (TAT), fibrinopeptide A (FpA), D-dimer, fibrin degradation products-E (FDP-E), fibrinogen, alpha2-plasmin inhibitor-plasmin complex (PIC), and percent activity of antithrombin III (AT-III) were measured within 48 h, at 1 week, and at 3 weeks after the stroke onset. Significantly elevated levels of TAT and FpA, which are both markers of thrombin formation, were observed in CEI patients, and these elevated levels were associated with increasing D-dimer levels for 3 weeks (P<0.0001). D-Dimer in CEI patients was significantly elevated compared to control, LI and ATI levels within 48 h (P<0.001). Percent activity of AT-III was significantly decreased in CEI patients for 3 weeks compared to this activity in controls, LI and ATI (P<0.001). TAT and FpA also increased significantly within 48 h in ATI subjects and declined thereafter. A significant elevation of FDP-E (P<0.001) and D-dimer (P<0.05, P<0.01) was detected in parallel with increasing fibrinogen for 3 weeks. However, there was no significant depletion of percent activity of AT-III in ATI. In LI subjects, no significant elevation of TAT, D-dimer or FDP-E were observed within 1 week. PIC increased significantly in three subtypes of brain infarcts, but did not differ significantly among the three subtypes for 3 weeks. An accurate assessment of sequential alterations in thrombotic and fibrinolytic markers in the acute stage of brain infarct should contribute to the clinical diagnosis of brain infarct subtype. Alterations in these markers in response to activation of the coagulatory system are attributable to the different pathogenesis of ischemic stroke. PMID- 11099717 TI - Botulinum toxin type-A treatment in spastic paraparesis: a neurophysiological study. AB - OBJECTIVE: The aim of this study was to verify the action of Botulinum toxin type A (BoNT-A) by means of neurophysiological techniques, in patients presenting lower limb spasticity and requiring BoNT-A injections in the calf muscles, due to the poor response to medical antispastic treatment. SUBJECTS AND METHOD: Patients presenting paraparesis were enrolled. They underwent clinical evaluation for spasticity according to the Ashworth scale and neurophysiological recordings including: motor evoked potentials (MEPs) to transcranial magnetic stimulation (TMS) of the leg area; compound motor action potential (cMAP) to tibial nerve stimulation, F-wave, and H-reflex before the treatment and 24 h, 2 weeks and 1 month after the injection of BoNT-A. In all patients, gastrocnemius was treated and in some cases soleus or tibialis posterior muscles were also injected. RESULTS: In all patients, BoNT-A injections induced a clear clinical improvement as showed by the reduced spasticity values of the Ashworth scale. A significant increment of MEP latency and central conduction time (CCT) duration were observed 2 weeks after the treatment only in the injected muscles. CONCLUSIONS: Prolonged MEP latencies and CCT after BoNT-A injections is probably due to a central alteration in responsiveness of spinal motor neurons to descending impulses from the corticospinal tracts. Such changes represent objective parameters heralding clinical efficacy of treatment. PMID- 11099718 TI - Choice reaction time after levodopa challenge in parkinsonian patients. AB - Various types of choice reaction time paradigms demonstrated deficits in the preparation and execution of movements in parkinsonian subjects. These studies showed controversial results, since they included parkinsonian individuals being: (i) previously untreated; (ii) off; or (iii) on anti-parkinsonian medication. Moreover, these trials do not take into consideration the acute effects of levodopa administration. Objective of this study was to determine the effect of long-term dopaminergic substitution therapy within a standardized levodopa challenge test in combination with a repeatedly performed choice reaction time task in parkinsonian individuals. Parkinsonian participants consisted of previously untreated, so-called "de-novo" patients and of individuals, who were chronically substituted with dopaminergic drugs, but were taken off medication for at least 12 h. All participants took 250 mg levodopa/benserazide after assessment of baseline data. Then we repeatedly measured choice reaction- and movement time within the next 90 min. No significant change of the assessed task data appeared in the "de-novo" group, but reaction- and movement time significantly shortened in previously treated subjects. Sedative effects of levodopa and/or dopaminergic overstimulation hypothetically explain the results of the previously untreated patients, whereas long-term dopaminergic substitution therapy hypothetically causes tolerance to these phenomena in treated parkinsonian individuals. Future studies on parkinsonian subjects should discuss their results on the basic pathophysiology or basal ganglia dysfunction in the light of a putative impact of long-term anti-parkinsonian drug therapy. PMID- 11099719 TI - Comparison of serum S-100 protein levels following stroke and traumatic brain injury. AB - Temporal changes in serum S-100 protein levels were compared between patients with ischemic stroke, transient ischemic attack (TIA) and traumatic brain injury (TBI). In addition, S-100 levels were correlated with clinical severity and outcome. Measurements were done with a LIA-mat((R)) Sangtec((R)) 100 using an automated immunoluminometric assay. Serum S-100 was measured in 21 stroke patients, 18 TIA patients and ten TBI patients on days 1 (0-24 h), 2, 3, 4, 5 or 6 and 8 or 9. In a control group of 28 healthy volunteers one measurement was done. For the stroke and TIA patients, National Institutes of Health Stroke Scale (NIHSS) scores were obtained on admission and on day 10. For the TBI patients, Glasgow Coma Scale (GCS) scores were obtained on admission and Glasgow Outcome Scale (GOS) scores were obtained after 6 months. Changes in serum S-100 levels over the first 3 days were significantly different between stroke and TBI patients (P=0.014) and between stroke and TIA patients (P=0.006). Peak concentrations of S-100 were most often observed on day 3 or 4 after stroke and on day 1 or 2 after TBI. In the stroke patients individual S-100 peak levels correlated well with the NIHSS score on admission (r=0.58 P=0.014) and the change in NIHSS score between day 10 and day 1 (r=0.65, P=0. 005). In the TBI patients a good correlation between individual peak levels of S-100 and the GCS score on admission (r=-0.81, P=0.010) and the GOS score 6 months after the trauma was found (r=-0.87, P=0. 004). We conclude that there is a significant difference in temporal changes of S-100 levels between ischemic stroke and TBI patients. This suggests different pathophysiological mechanisms. The results of this study further confirm that peak levels of serum S-100 correlate with neurological deficit resulting from either stroke or TBI. PMID- 11099720 TI - MRI correlates of dementia after first clinical ischemic stroke. AB - BACKGROUND AND PURPOSE: Dementia after first clinical stroke frequently has been found, but the clinical and radiological correlates have not been fully detailed. We examined magnetic resonance imaging (MRI) correlates of dementia in a large well-defined series of patients with first clinical ischemic stroke. METHODS: Detailed medical, neurological and neuropsychological examination was conducted 3 months after ischemic stroke for 273 patients with first clinical stroke from a consecutive series of 486 patients aged 55-85 years. MRI of the head categorised infarcts (type, site, side, number, volume), extent of white matter lesions (WMLs) and degree of atrophy. The DSM-III definition for dementia was used. RESULTS: Dementia was diagnosed in 79 (28.9%) of the patients with first clinical stroke. Volumes, numbers, distinct sites of infarcts, extent of WMLs and degree of atrophy were different for the demented and nondemented subjects. Logistic regression analysis showed that the correlates of dementia included the combination of infarct features (volume of infarcts in left-sided anterior corona radiata; OR 1.86), extent of WMLs (OR 1. 37), medial temporal lobe atrophy (OR 3.4) and host factors (low education; OR 1.11). The additive effect of having more than one correlate was detected (OR 2.53). CONCLUSIONS: Dementia occurring after first clinical stroke is frequent and not solely due to a single stroke, but contain a combination of infarcts features, extent of WMLs, medial temporal lobe atrophy and host factors reflecting more than one underlying pathology. PMID- 11099721 TI - The clinical, radiological and pathological profile of tuberculous meningitis in patients with and without human immunodeficiency virus infection. AB - BACKGROUND: As human immunodeficiency virus (HIV) infection primarily impairs cellular immunity, the immune responses of HIV-infected individuals to tuberculous bacilli may be inadequate. The features of pulmonary and abdominal tuberculosis evident in HIV-positive (HIV-P) patients with severe immunosuppression are markedly different from those seen in HIV-negative (HIV-N) patients. However, such differences have not been reported in tuberculous meningitis (TBM). Here, we therefore compared the clinical, radiological and pathological features of TBM in patients with and without HIV infection. METHODS AND RESULTS: Twenty-two HIV-P patients with TBM, seen over 5 years, were studied and compared with 31 HIV-N patients with TBM. Although clinical features were similar, cognitive dysfunction was more common amongst the HIV-P group. Pathological features were markedly different in the HIV-P group reflecting severely reduced and atypical inflammatory response, and extensive vasculopathy. This manifested as absence or minimal meningeal enhancement and absence of communicating hydrocephalus on CT scan in HIV-P patients. Mortality was higher within the HIV-P group and depressed levels of consciousness and hemiplegia were associated with poor prognosis. CONCLUSION: The clinical, radiological and pathological features of TBM in HIV-P patients are distinctly different from those without HIV infection; a finding previously unreported. PMID- 11099722 TI - Genetic risk factors of sporadic Alzheimer's disease among Chinese in Taiwan. AB - To evaluate the genetic factors for AD among a Chinese population in Taiwan, we studied the polymorphisms of six candidate genes of Alzheimer's disease (AD), including the regulatory region of apolipoprotein E (Apo-E, G-186T), the promoter of apolipoprotein E (Apo-E, A-491T), the bleomycin hydrolase gene (BH, A1450G), a mutation of alpha(2)-macroglobulin gene (A2M G2998A), low-density lipoprotein receptor-related protein gene (LRP, C766T), and alpha(1)-antichymotrypsin gene (ACT, -15Ala/Thr) in AD patients and non-affected elder individuals among Taiwanese Chinese. Eighty-two AD patients and 110 non-affected individuals were recruited for this study. We used polymerase chain reaction (PCR) and restriction enzyme digestion to identify their genotypes. The statistical examination was performed by combining the results of our previous reports - apolipoprotein E epsilon4 (ApoE-4), presenilin-1 intronic polymorphism (PS-1, allele 1/2), and the five-nucleotide deletion of alpha(2)-macroglobulin gene (A2M). Among these nine candidate genes of AD, the ApoE-4 allele is the only independent genetic risk factor for AD. The other candidate genes in this study were not associated with the occurrence of AD. In addition, there are no gene-gene interactions. PMID- 11099723 TI - Liposuction and ischemic optic neuropathy. Case report and review of literature. AB - Ischemic optic neuropathy occurred in a patient following liposuction. Perioperative anemia and hypotension may be the cause of this complication. Correction of anemia with transfusion improved the hemoglobin and hematocrit but the right eye remained blind. Liposuction should be added to the list of the surgical procedures that may produce ischemic optic neuropathy as an isolated complication. PMID- 11099724 TI - Simultaneous hypertensive intracerebral hematomas: two case reports. AB - We describe two patients (76- and 54-year-old females) with multiple hypertensive intracerebral hematomas occurring simultaneously. One patient had a right thalamic hematoma extending into the internal capsule and basal ganglia together with an other one in the left putamen. The other patient had two hematomas located ipsilaterally in the left putamen and thalamus. Their neurological examinations showed only unilateral deficits. Their magnetic resonance angiograms revealed no vascular malformations. Neuroradiological procedures are essential for the diagnosis of these multiple brain events. PMID- 11099725 TI - Electrophysiological dissociation between verbal and nonverbal semantic processing in learning disabled adults. AB - Event-related potentials (ERPs) were recorded as 16 adults with learning disabilities (LD) and 16 controls were presented with two sets of stimuli. The first set comprised pairs of line drawings and environmental sounds (nonverbal condition); the second consisted of printed and spoken words (verbal condition). In the controls, semantically related items elicited smaller N400s than unrelated items in both conditions, with opposing hemispheric asymmetries for spoken words and environmental sounds. The LD group did not show a significant difference between related and unrelated words, despite a robust context effect for nonspeech sounds. The results suggest anomalous processing limited to the verbal domain in a simple semantic association task in the LD group. Semantic deficits in this group may reflect a relatively specific deficit in forming verbal associations rather than a more general difficulty that spans both verbal and nonverbal domains. PMID- 11099726 TI - Manual laterality in haptic and visual reaching tasks by tufted capuchin monkeys (Cebus apella). An association between hand preference and hand accuracy for food discrimination. AB - Manual laterality was examined in 26 tufted capuchins (Cebus apella) in three tasks differing in their sensorimotor demands and the availability of visual cues. The Haptic discrimination task required the monkeys to discriminate haptically between two pumpkin seeds and two tinfoil items stuck into a tray inside an opaque box. The other two tasks required the monkeys to reach for two pumpkin seeds stuck into the tray within a transparent box with vision (Visually guided reaching task) or without vision (Visual-Tactual reaching task) during reaching. A significant group-level left hand bias was found for food retrieval in both the Haptic discrimination and Visual-Tactual tasks, and a significant group-level right hand bias in the Visually guided reaching task. The strength of hand preferences did not differ among the tasks. It was found that the accuracy of food recognition in the Haptic discrimination task was greater for the left than the right hand. The results suggest that the differences in the manipulo spatial requirements of the tasks and in the availability of visual cues can variously affect manual laterality in capuchins. The left-hand preferences for the Haptic discrimination and Visual-Tactual tasks as well as the left-hand advantage for food discrimination may reflect a greater involvement of the right hemisphere in processing haptic information. PMID- 11099727 TI - Categorization and category effects in normal object recognition: a PET study. AB - To investigate the neural correlates of the structural and semantic stages of visual object recognition and to see whether any effects of category could be found at these stages, we compared the rCBF associated with two categorization tasks (subjects decided whether pictures represented artefacts or natural objects), and two object decision tasks (subjects decided whether pictures represented real objects or nonobjects). The categorization tasks differed from each other in that the items presented in the critical scan window were drawn primarily from the category of artefacts in the one task and from the category of natural objects in the other. The same was true for the object decision tasks. The experiment thus comprised a two-by-two factorial design. The factors were Task Type with two levels (object decision vs. categorization) and Category also with two levels (natural objects vs. artefacts). The object decision tasks were associated with activation of areas involved in structural processing (fusiform gyri, right inferior frontal gyrus). In contrast, the categorization tasks were associated with activation of the left inferior temporal gyrus, a structure believed to be involved in semantic processing. In addition, activation of the left premotor cortex was found during the categorization of artefacts compared with both the categorization of natural objects and object decision to artefacts. These findings suggest that the structural and semantic stages are dissociable and that the categorization of artefacts, as opposed to the categorization of natural objects, is based, in part, on action knowledge mediated by the left premotor cortex. However, because artefacts and natural objects often caused activation in the same regions within tasks, processing of these categories is not totally segregated. Rather, the categories differ in their weight on different forms of knowledge in particular tasks. PMID- 11099728 TI - Planum temporale, planum parietale and dichotic listening in dyslexia. AB - A reduction or reversal of the normal leftward asymmetry of the planum temporale (PT) has been claimed to be typical of dyslexia, although some recent studies have challenged this view. In a population-based study of 20 right-handed dyslexic boys and 20 matched controls, we have measured the PT and the adjacent planum parietale (PP) region in sagittal magnetic resonance images. For the PT, mean left and right areas and asymmetry coefficients were compared. Since a PP area often could not be identified in one or both hemispheres, a qualitative comparison was used for this region. The total planar area (sum of PT and PP) was also compared between the two groups. A dichotic listening (DL) test with consonant-vowel syllables was administered to assess functional asymmetry of language. The results showed a mean leftward PT asymmetry in both the dyslexic and the control group, with no significant difference for the degree of PT asymmetry. Planned comparisons revealed however, a trend towards smaller left PT in the dyslexic group. In control children, but not in the dyslexic children, a significant correlation between PT asymmetry and reading was observed. A mean leftward asymmetry was also found for the total planar area, with no difference between the groups for the degree of asymmetry. Significantly fewer dyslexic children than control children showed a rightward asymmetry for the PP region. Both groups showed a normal right ear advantage on the DL task, with no significant difference for DL asymmetry. No significant correlation was observed between PT asymmetry and DL asymmetry. The present population-based study adds to recent reports of normal PT asymmetry in dyslexia, but indicates that subtle morphological abnormalities in the left planar area may be present in this condition. PMID- 11099729 TI - Bimanual coordination and limb-specific parameterization in patients with Parkinson's disease. AB - Bimanual coordination and the capability to parameterize the individual limb movements were examined in patients with Parkinson's disease (PD) as compared to healthy control subjects. In-phase and anti-phase patterns were performed while the individual limb movements were subjected to amplitude and loading manipulations. Findings showed that PD patients produced the bimanual configurations with lower degrees of phasing accuracy and consistency than control subjects, indicating an impairment at the global (coordinative) level of simultaneously produced movements. At the local (limb-specific) level, the imposed distances with and without loading were unaffected in PD patients as compared to control subjects, whereas cycle times were prolonged and depended on the task requirements. This illustrates a disturbance at the limb-specific level in complying with the execution of the submovements. The finding that movement slowness only became evident in the more complex conditions, suggests that it did not mainly represent a deficit in the execution of coordinated movements, but rather an inability to accommodate the motor output during stringent spatiotemporal task constraints. PMID- 11099730 TI - Prefrontal brain electrical asymmetry predicts the evaluation of affective stimuli. AB - Measures of left-right asymmetry in resting brain activity were derived from spectral estimates of electroencephalogram (EEG) alpha-band power density in 13 homologous scalp electrode pairs from 81 right-handed individuals (43 F) on two occasions separated by 6 weeks. At a third, later session, these individuals completed a cognitive task, comparing word-pairs that systematically differed in affective tone. For an extended series of paired-comparisons, the subject chose the one word-pair that 'went together best'. Objectively, associative strength was comparable for both word-pairs. Individuals with relatively greater left sided anterior frontal resting activity were more likely to select the more pleasant word-pair. Relations between word-pair selection and asymmetry in resting brain activity at central and posterior sites were not significant. PMID- 11099731 TI - Emotional curiosity: modulation of visuospatial attention by arousal is preserved in aging and early-stage Alzheimer's disease. AB - Previous studies have shown that Alzheimer's disease, even in its early stages, decreases novelty-seeking behaviors (curiosity) and impairs the shifting of spatial attention to extrapersonal targets. In this study, early-stage probable Alzheimer's disease patients (PRAD) and young and aging controls were shown pairs of visual scenes, some of which contained emotionally-arousing material, while eye movements were recorded under free viewing conditions. In all three subject groups, emotionally-arousing scenes attracted more viewing time and also became the preferential target of the initial visual orientation. Our findings suggest that the arousing properties of sensory stimuli may overcome some of the AD related impairments in the distribution of attention to extrapersonal targets. These results may have implications for interventions aimed at improving the cognitive symptoms of PRAD. PMID- 11099732 TI - 'Where' depends on 'what': a differential functional anatomy for position discrimination in one- versus two-dimensions. AB - Line bisection is widely used as a clinical test of spatial cognition in patients with left visuospatial neglect after right hemisphere lesion. Surprisingly, many neglect patients who show severe impairment on marking the center of horizontal lines can accurately mark the center of squares. That these patients with left neglect are also typically poor at judging whether lines are correctly prebisected implies that the deficit can be perceptual rather than motoric. These findings suggest a differential neural basis for one- and two-dimensional visual position discrimination that we investigated with functional neuroimaging (fMRI). Normal subjects judged whether, in premarked lines or squares, the mark was placed centrally. Line center judgements differentially activated right parietal cortex, while square center judgements differentially activated the lingual gyrus bilaterally. These distinct neural bases for one- and two-dimensional visuospatial judgements help explain the observed clinical dissociations by showing that as a stimulus becomes a better, more 'object-like' gestalt, the ventral visuoperceptive route assumes more responsibility for assessing position within the object. PMID- 11099733 TI - Atypical organisation of the auditory cortex in dyslexia as revealed by MEG. AB - Neuroanatomical and -radiological studies have converged to suggest an atypical organisation in the temporal bank of the left-hemispheric Sylvian fissure for dyslexia. Against the background of this finding, we applied high temporal resolution magnetoencephalography (MEG) to investigate functional aspects of the left-hemispheric auditory cortex in 11 right-handed dyslexic children (aged 8-13 years) and nine matched normal subjects (aged 8-14 years). Event-related field components during a passive oddball paradigm with pure tones and consonant-vowel syllables were evaluated. The first major peak of the auditory evoked response, the M80, showed identical topographical distributions in both groups. In contrast, the generating brain structures of the later M210 component were located more anterior to the earlier response in children with dyslexia only. Control children exhibited the expected activation of more posterior source locations of the component that appeared later in the processing stream. Since the group difference in the relative location of the M210 source seemed to be independent of stimulus category, it is concluded that dyslexics and normally literate children differ as to the organisation of their left-hemispheric auditory cortex. PMID- 11099734 TI - Apraxic disturbances in patients with mild to moderate Alzheimer's disease. AB - Twenty-two patients meeting the NINCDS-ADRDA diagnostic criteria for probable AD were included in the study, along with 10 matched controls. Praxic disturbances were investigated using eight tasks and the results were interpreted according to the neuropsychological model of Roy and Square modified by Rothi et al. (1988) [Aphasiology 21:381-388] which distinguishes a conceptual system concerned with knowledge of the action and function of gestures and a production system that effects gestures in the environment. Disturbance of the production system was found only in 17 patients. Disturbance of the production system was correlated to disturbance of verbal comprehension. The patients scored lower using the left hand than the right. Disturbance of the conceptual system was found in all patients and was not significantly correlated with other cognitive deficits. No significant difference in results was found according to the type of input to the conceptual system (visual or verbal). Deficits in tasks using real objects were correlated to disturbances of both the production and conceptual systems. Most patients performed poorly both in tasks exploring the conceptual system and in those exploring the production system. However, two patients performed badly in production tasks but had performances in the range of controls for conceptual tasks and one patient had the opposite pattern of dissociation. This provides evidence that the production and the conceptual system are independent. Impairment in the ability to perform everyday activities was correlated to disturbances of the conceptual system whereas poor performances in tasks exploring the production system or in using real objects were not. PMID- 11099735 TI - Changes in depression and physical decline in older adults: a longitudinal perspective. AB - BACKGROUND: The impact of chronicity and changes in depression on physical decline over time in older persons has not been elucidated. METHODS: This prospective cohort study of 2121 community-dwelling persons aged 55-85 years uses two measurement occasions of depression (CES-D scale) over 3 years to distinguish persons with chronic, remitted, or emerging depression and persons who were never depressed. Physical function is assessed by self-reported physical ability as well as by observed performance on a short battery of tests. RESULTS: After adjustment for baseline physical function, health status and sociodemographic factors, chronic depression was associated with significantly greater decline in self-reported physical ability over 3 years when compared to never depressed persons (odds ratio (OR)=2.83, 95% confidence interval (CI)=1.86-4. 30). In the oldest old, but not in the youngest old, chronic depression was also significantly predictive of greater decline in observed physical performance over 3 years (OR=2.22, 95% CI=1.43-3. 79). Comparable effects were found for older persons with emerging depression. Persons with remitted depression did not have greater decline in reported physical ability or observed performance than persons who were never depressed. CONCLUSIONS: Our findings among community-dwelling older persons show that chronicity of depression has a large impact on physical decline over time. Since persons with remitted depression did not have greater physical decline than never depressed persons, these findings suggest that early recognition and treatment of depression in older persons could be protective for subsequent physical decline. PMID- 11099736 TI - Effects of rapid cycling on response to lithium maintenance treatment in 360 bipolar I and II disorder patients. AB - INTRODUCTION: Rapid cycling (RC) in bipolar disorders is widely believed to predict future morbidity and poor treatment response, although empirical testing of its predictive utility remains limited. METHODS: In 360 DSM-IV bipolar I (N=218) and II (N=142) disorder subjects (64% women) followed over an average of 13.3 years, we evaluated factors associated with RC status with bivariate and multivariate techniques, and response to lithium maintenance treatment (recurrence rates, time ill, survival analysis of time to recurrence on lithium). RESULTS: RC risk (15.6% of cases) was 5. 1-times greater in bipolar II vs. I subjects (30.3%/6.0%), in minor excess in women vs. men (17.9%/11.5%), and associated with premorbid cyclothymia, depressive first episodes, older onset age, and being employed or married. Before lithium, RC vs. non-RC cases had more mean total (3.9/1.2), manic, and depressive episodes/year, and greater percent time ill (60%/38%). During treatment, prior RC status was unrelated to time to first recurrence and other measures of morbidity and improvement including percent time ill, although depressive episodes were 2.7-times more frequent, and there was 13.7% less chance of full protection from all recurrences in RC cases. LIMITATIONS: The study is naturalistic, without random assignment or blind assessment. CONCLUSIONS: The RC bipolar subtype was strongly associated with type II diagnosis, higher average prelithium episode frequency and percent time ill, and weakly with female sex, but not with greater overall morbidity during treatment. PMID- 11099737 TI - Failure to detect amphetamine or 1-amino-3-phenylpropane in humans or rats receiving the MAO inhibitor tranylcypromine. AB - BACKGROUND: There have been conflicting reports in the literature about whether or not tranylcypromine is metabolized to amphetamine. In the current report, we investigated this possible route of metabolism in both rats and humans. Body fluid samples from patients and rats and brain, liver and heart samples from rats were analyzed for levels of amphetamine and 1-amino-3-phenylpropane, another potential product of cleavage of the cyclopropyl ring of tranylcypromine after administration of tranylcypromine. Extracted samples were reacted with pentofluorobenzenesulfonyl chloride and analyzed using electron-capture gas chromatography. RESULTS: Amphetamine or 1-amino-3-phenylpropane were not found in any of the samples, indicating that opening of the cyclopropyl ring of tranylcypromine is not a significant route of metabolism for this drug at usual doses. LIMITATIONS: The assay procedure did not permit analysis of 1-amino-2 phenylpropane (another possible product of cleavage of the cyclopropyl ring of tranylcypromine) or of N-methylamphetamine. CONCLUSIONS: These studies support the growing body of evidence indicating that opening of the cyclopropyl ring of tranylcypromine to form amphetamine, a drug of abuse, is not significant at usual doses of tranylcypromine. PMID- 11099738 TI - Protective effect of pregnancy in women with lithium-responsive bipolar disorder. AB - BACKGROUND: Recent psychiatric literature, while indicating a high incidence of postpartum depression, contains a few clinical reports which support our observations that women with episodic bipolar disorder often remain well without treatment during pregnancy. Our retrospective study statistically examines the clinical course of 28 women with RDC typical bipolar disorder, type I, who became pregnant prior to receiving successful lithium prophylaxis. METHODS: We derived all data from the International Group for the Study of Lithium-treated Patients (IGSLI) database of excellent lithium responders. Data were compared both intraindividually, using data from three 9-month periods - immediately prior to pregnancy, pregnancy and postpartum - and interindividually, using never-pregnant women as controls. RESULTS: Intraindividual data show that women with typical bipolar disorder, type I, experience significantly fewer and shorter recurrences during pregnancy than either before or after. Interindividual comparisons indicate that the recurrence risk during pregnancy is markedly lower than the clinical course would predict. Moreover, the few recurrences observed during pregnancy all took place in the last 5 weeks. LIMITATIONS: Limiting cases to lithium responsive patients could have reduced heterogeneity and perhaps generalizability. CONCLUSIONS: The findings, nonetheless, indicate a marked improvement of the clinical course of typical bipolar disorder, type I, lithium responsive, during pregnancy. Exploring the underlying protective mechanisms may lead to new understanding of the pathophysiology of mood disorders and to new approaches to treatment and prevention. PMID- 11099739 TI - Relation of subjective and received social support to clinical and self-report assessments of depressive symptoms in an elderly population. AB - BACKGROUND: The authors sought to evaluate the associations between depressive symptoms and social support in a sample drawn from a relatively understudied population - depressed elderly patients. The present study also used a multi measure approach to assess both depressive symptomatology and social support. METHODS: In this prospective study of 115 patients we examined: (1) the baseline relations among a self-report measure of depressive symptoms, two clinical assessments of depressive symptoms, and subjective and received social support, and (2) the ability of social support to predict changes in clinical assessments of depressive symptoms at 6 months and 1 year. Education level, financial concerns, activities of daily living ratings, and gender were controlled for. RESULTS: Baseline subjective support was negatively related to self-reports of depressive symptoms, but unrelated to clinical assessments at baseline or follow up. Conversely, received support was unrelated to self-reported depressive symptoms, but positively related to both clinical assessments at baseline. However, higher ratings of received support at baseline predicted decreases in clinical ratings of depressive symptoms at 6 months and 1 year. LIMITATIONS: These data were gathered in a primarily Caucasian sample, thus the findings may not generalize to more diverse ethnic populations. Potential confounding due to treatment mode and setting was not controlled in the present analyses. CONCLUSIONS: These results have important implications for interpreting clinical data in elderly depressed patients. Specifically, when depressive symptoms are assessed using clinician ratings, the most informative aspect of social support with respect to future clinical status appears to be received, rather than perceived, support measures. PMID- 11099740 TI - Human fibroblasts as a relevant model to study signal transduction in affective disorders. AB - BACKGROUND: Previous studies have demonstrated a blunted beta adrenoceptor-linked protein kinase A (PKA) response in the 900xg supernatant fraction of human fibroblasts cultured from patients with major depression. RESULTS: Results of the present studies demonstrate a significant reduction in the B(max) value of [3H]cyclic AMP binding to the regulatory subunit of PKA in the supernatant fraction of fibroblasts from patients with major depression with no change in the K(d) values. The data are consistent with the previous observation that the maximal stimulation of PKA by cyclic AMP is reduced without a change in the EC(50) value. The blunted beta adrenoceptor-mediated PKA response in fibroblasts from patients with major depression is reflected in a significant reduction in the isoproterenol-stimulated phosphorylation of the nuclear transcription factor CREB. Both, the isoproterenol-mediated phosphorylation of nuclear CREB and the activation of the stably transfected luciferase reporter gene, pAD neo2-C12-BGL, were inhibited by the beta(2) adrenoceptor antagonist ICI 118551, thus indicating that the gene activating action of isoproterenol in human fibroblasts is mediated via the beta(2) adrenoceptor cascade. The low EC(50) value of 1 nM isoproterenol for activation of gene expression in stably transfected human fibroblasts appears to be a reflection of the amplification mechanism occurring via the beta adrenoceptor-cyclic AMP-PKA-CREB transduction cascade. CONCLUSIONS: The results support the notion that human fibroblasts represent a relevant model for studying processes of signal transduction in patients with affective disorders. PMID- 11099741 TI - Gender differences in the presentation of depressed outpatients: a comparison of descriptive variables. AB - BACKGROUND: Gender differences in the clinical manifestation of depression and related variables were examined in 170 depressed outpatients. METHOD: Age of onset of depression, chronicity, recurrence, subtype of depression, self-harm history and prior treatment history were assessed with structured clinical interviews. Depression symptom profile, family psychiatric history and social, occupational and interpersonal functioning were assessed with self-report and clinician ratings. RESULTS: Overall, males and females were remarkably similar. Significant findings were that depressed females reported significantly more appetite increase, weight gain and carbohydrate craving, and in general, expressed their depression in a more emotional manner, than depressed males. CONCLUSION: Psychosocial and biological explanations for these results are explored. LIMITATIONS: Descriptive study and multiple testing PMID- 11099742 TI - Further evidence for low serum cholesterol and suicidal behaviour. AB - OBJECTIVE: To examine for a relationship between serum cholesterol and suicidal behavior. METHODS: Patients admitted after an overdose (N=120) were compared with controls (N=120) for their serum cholesterol levels. RESULTS: Patients who had overdosed had significantly lower serum cholesterol levels than controls (mean+/ S.D. 171+/-31 vs. 196+/-30 mg/dl, P<0.0001). CONCLUSION: These results add to a grouping literature reporting that low serum cholesterol is associated with suicidal behavior. PMID- 11099744 TI - The correlates of community functioning in patients with bipolar disorder. AB - BACKGROUND: Although bipolar disorder is associated with substantial levels of disability, efforts to investigate the correlates of impairment have been meagre. METHODS: Sixty-one euthymic patients with a diagnosis of bipolar disorder were administered a variety of quality of life measures, including a measure of community functioning entitled the Occupational Performance Questionnaire (OPQ). This measure included a Community Functioning Scale (CFS) that provides a rating of adaptive level of functioning that was compared with other clinical and functional indices. RESULTS: The OPQ was found responsive to the assessment of community functioning among euthymic patients, as about one third of the patients did not meet the criteria for adequate level of community functioning. Moreover, a positive history for alcoholism or alcohol abuse and reported current levels of high anxiety were associated with the impairment in community functioning. LIMITATIONS: This study relied on self-report data derived from a measure of community functioning developed specifically for this study. CONCLUSION: Despite the preliminary nature of these findings, further investigation of the functional impairments associated with bipolar disorder is needed. PMID- 11099743 TI - Short-term cotherapy with clonazepam and fluoxetine: anxiety, sleep disturbance and core symptoms of depression. AB - BACKGROUND: SSRIs resolve depression slowly and may increase anxiety or insomnia. Adding clonazepam to fluoxetine sped response, raising the question of mechanism of action: reducing symptoms co-existing with depression, suppressing side effects, and/or alleviating core depressive symptoms. METHOD: Adult outpatients randomly assigned to double-blind treatment with fluoxetine 20 mg+placebo or fluoxetine+clonazepam 0.5-1.0 mg were assessed by a HAM-D anxiety cluster, sleep disturbance cluster, and core symptoms cluster. RESULTS: No serious AEs were noted; no cotherapy patients dropped for AEs. Cotherapy proved superior (HAM-D total, anxiety cluster, sleep disturbance cluster ANOVA P<0.001; core symptoms P<0.011). Treatment-emergent anxiety was reported for 25% of placebo patients and 7% of cotherapy patients (P<0.037); sleep disturbance for 10% of placebo patients and no cotherapy patients (P<0.055). Sedation and dry mouth were more common for cotherapy treatment (P>0.20). LIMITATIONS: Extended treatment and refractory depression were not addressed. CONCLUSIONS: Low-dose cotherapy of fluoxetine with clonazepam was safe and accelerated response over 21 days of treatment, decreasing anxiety and sleep disturbance as symptoms and partially suppressed them as SSRI side-effects; it also modestly reduced core symptoms of low mood and loss of interest. PMID- 11099745 TI - Is impaired outcome following a first manic episode due to mood-incongruent psychosis? AB - BACKGROUND: Mood-incongruent psychosis during the course of bipolar disorder has been associated with poor outcome. However, it remains unknown whether this is secondary to persistent affective or psychotic symptoms or both. METHOD: Fifty patients with bipolar disorder between the ages of 16 and 45 years were recruited during their first psychiatric hospitalization for mania. These patients were evaluated using structured and semi-structured clinical instruments then followed longitudinally. Outcomes during the first eight months of follow-up were compared between patients with mood-incongruent psychosis and those without (i.e., patients with mood-congruent psychosis or no psychosis) during the index manic episode. Specifically, ratings of the percent of weeks during follow-up with psychosis and affective syndromes and symptoms, as well as ratings of global outcome (GAF), were compared. RESULTS: Patients with mood-incongruent psychosis at the index hospitalization exhibited significantly more weeks during follow-up with both mood-incongruent and mood-congruent psychotic symptoms than patients without mood-incongruent psychosis. Mood-incongruent psychosis was also associated with poorer overall functioning during the outcome interval. The groups did not differ in the percent of weeks with affective syndromes or symptoms. Treatment during follow-up did not differ between groups and was not associated with outcome variables in general. CONCLUSION: Mood-incongruent psychosis that occurs during the first manic episode appears to predict an increased likelihood of persistent psychotic symptoms during the subsequent eight months. This persistence of psychosis is associated with a worse overall course of illness as compared to patients without mood-incongruent psychosis. LIMITATIONS: These results apply to a relatively short outcome period and are from a single center. PMID- 11099746 TI - Early-onset versus late-onset atypical depression: unipolar and bipolar II. AB - BACKGROUND: To find differences between early- and late-onset atypical depression (AD). METHODS: 211 unipolar/bipolar II AD outpatients, interviewed with DSM-IV Structured Clinical Interview and depression rating scales. Logistic regression was used. RESULTS: Early-onset AD was significantly associated with age, female gender, duration of illness, recurrences, chronicity, MADRS, bipolar II and unipolar. Early-onset bipolar II AD was significantly associated with age, female gender, duration of illness, recurrences and chronicity. Early-onset unipolar AD was significantly associated with age. LIMITATIONS: Age at onset recall bias, single interviewer, non-blind, cross-sectional assessment, bipolar II diagnosis reliability. CONCLUSIONS: Bipolar II AD is more likely to be chronic if early onset. PMID- 11099747 TI - Obstetric complications in patients with depression--a population-based case control study. AB - BACKGROUND: To examine whether sufferers of affective disorders are more likely to be subject to obstetric complications than normal healthy people. METHOD: Data based on prospectively recorded birth case-notes for patients with a diagnosis of depression (or related disorders) with early onset were compared to those of normal healthy controls, individually matched by gender, time and parity of birth, maternal age and marital status. RESULTS: Forty-one case-controls pairs born between 1964 and 1978 were compared. No differences between cases and controls in gestational age or birthweight were significant, though depressive patients on average weighed 200 g less than controls at birth. Patients were more likely than controls to be small for their gestational age (22 vs. 1: chi(2)=4.34, P=0.03). They were significantly more likely than controls to have suffered at least one obstetric complication: 35 (85%) vs. 25 (60%), chi(2)=5.03, P=0.02; or more than one (two on average, as opposed to one on average among controls). No obstetric complication was seen significantly more among cases than controls, apart from bleeding during gestation, which was observed for four cases and no controls. The prevalence of complications with a clear brain damaging potential did not differ significantly between cases and controls: 11 (26%) vs. 8 (19%). CONCLUSIONS: A developmental deficit, as indicated by lower birthweight and gestational age, may contribute to the risk of depressive breakdowns and affective disorders in later life. Severe, brain damaging obstetric complications are unlikely to be a significant risk factor for affective disorders, though some early onset cases may be accounted for by prenatal brain lesions. LIMITATIONS: Sample size limits statistical power for isolation of a rare, single risk factor. PMID- 11099748 TI - The Edinburgh Postnatal Depression Scale (EPDS) and the detection of major depressive disorders in early postpartum: some concerns about false negatives. AB - This paper presents a phenomenological study of three false negative cases according to the Edinburgh Postnatal Depression Scale (EPDS) of major depressive disorder identified by a semi-structured clinical interview. In a study of 87 unselected women with 23 of them suffering from a major depressive disorder (according to the Research Diagnostic Criteria), three cases of major depressive disorders were not identified as potential cases by the EPDS. The symptomatology of these three false negative cases was also assessed by a semi-structured interview (Present State Examination). Comparisons between EPDS scores and the scores of two other self report questionnaires (the General Health Questionnaire 28 and the Center for Epidemiologic Studies-Depression Scale) suggest that EPDS is better at identifying depressed postnatal women with anhedonic and anxious symptomatology rather than those whose depression presents mainly with psychomotor retardation. PMID- 11099749 TI - Diminished perception of ambient light: a symptom of clinical depression? AB - OBJECTIVE: In a non-randomized, uncontrolled pilot study, the authors investigated whether depressed patients were more likely to perceive the lighting in their environment as being dimmer than usual. METHOD: 120 patients (46 males, 74 females) who presented for possible admission for depression at a psychiatric facility were administered a Diagnostic and Statistical Manual of disorders (DSM IV) based questionnaire and underwent psychiatric evaluation. A question asking whether 'the lights in my surroundings seem dimmer than usual' was included in the 15-point question survey. Statistical analyses were performed to determine whether an affirmative response to this dimness question was correlated with the depth of depression (mild, moderate, severe) and also whether significant correlation was present between the percentage of patients answering yes to the dimness question versus the number of yes responses to the core symptoms of depression. RESULTS: Two thirds of the patients categorized as severely depressed responded that their ambient environment appeared dimmer than usual compared to 21% of moderately and 14% of mildly depressed patients. This difference was statistically significant (P<0.05). The degree of depression as determined by the number of core questions answered affirmatively and the presence of this 'dimness' symptom were highly correlated (P=0.002, R=0.87). LIMITATIONS: The specificity of the finding has not been tested in reference to non-affective psychiatric patient groups. CONCLUSION: A patient's perception of the ambient light in the environment being dimmer than usual may be an important symptom of a major depressive disorder. Further replication and objective testing of visual function in depressed patients appears warranted. PMID- 11099750 TI - A double-blind, placebo-controlled, efficacy, safety, and pharmacokinetic study of INN 00835, a novel antidepressant peptide, in the treatment of major depression. AB - BACKGROUND: INN 00835 is a synthetic pentapeptide with a potential for rapid onset of action as an antidepressant. Its efficacy was investigated in a pilot study in patients diagnosed with major depression. METHODS: Fifty two patients received either active drug - INN 00835 (26 patients) - or placebo (26 patients), subcutaneously at 0.2 mg/kg for 5 consecutive days. The patients were evaluated for an additional 4 weeks after treatment. Efficacy was evaluated by the following psychiatric rating scales: HAMD, MADRS, CSRS, CGI, and VAS. The effect of treatment was also evaluated by using a biochemical marker: changes in blood platelet serotonin (5HT) uptake rates in drug-treated patients compared to those in the placebo group. Plasma concentrations of INN 00835 were measured by LC/MS. RESULTS: Statistical analysis indicated a strong pharmacodynamic correlation between plasma drug concentrations at 1 h after dosing and the reduction in the severity of depression as measured by the psychiatric rating scales. A minimum effective plasma concentration (MEC) of INN 00835 was 5 ng/ml. Statistically significant differences in response to treatment (P<0.05) were found between patients with plasma concentrations above MEC and those in the placebo group, as well as between subjects with plasma concentrations above and below the MEC. The peak effect was observed after the 5-day treatment and the response to treatment persisted during the 4-week follow-up period. The change of 5HT uptake rates after treatment was significantly larger in the drug-treated group than in the placebo group. LIMITATIONS: This was a pilot study conducted in a relatively small population (52 patients) and the limited number of blood sampling times did not allow a comprehensive pharmacokinetic analysis. There was a relatively large placebo response. The results have to be confirmed in future, large scale studies. CONCLUSIONS: INN 00835 appears to be a promising drug for the treatment of major depression. PMID- 11099751 TI - Open pergolide treatment of tricyclic and heterocyclic antidepressant-resistant depression. AB - BACKGROUND: Recently, a dopamine hypothesis of depression was put forward, and several studies have demonstrated that direct and indirect dopamine agonists have antidepressant effects. METHODS: Using Clinical Global Impressions, we evaluated the efficacy of 4-week treatment of pergolide as an antidepressant adjuvant involving 20 unipolar depressed patients who were refractory to standard treatment with antidepressants. RESULTS: One patients (5%) were very much improved, seven (35%) much improved, four (20%) minimally improved, six (30%) no change or worse, and two (10%) not assessed. There was no significant difference in any clinical factors between the pergolide responder and non-responder group. LIMITATIONS: This study was a non-blind open trial, and pergolide was added to tricyclic and heterocyclic antidepressants. CONCLUSION: Pergolide may be useful as an antidepressant adjuvant, suggesting a potential role for dopamine-2 stimulation in the antidepressant response. PMID- 11099752 TI - Graded sparing of visually-guided orienting following primary visual cortex ablations within the first postnatal month. AB - We compared the abilities of intact cats and cats that incurred lesions of areas 17 and 18 in adulthood, at one month of age (P28), or on the day of birth (P1), to detect and orient towards visual stimuli either moved into or illuminated in the periphery of the visual field, and to detect and orient towards a stationary, broad-band white-noise auditory stimulus. For all groups of cats, movement of a stimulus into the visual field was a more potent stimulus for evoking visually guided orienting movements than illumination of a static light-emitting diode (LED). The potency of the auditory stimulus was also extremely high. Proficiency on both visual tasks was graded according to the age at which areas 17 and 18 were ablated in the sequence: adult, P1, P28 and intact in the sequence worst- >best performance. The superior performance of the P1- and P28-groups provided evidence for sparing of visually-guided orienting, but the sparing was incomplete because it did not match performance of intact cats. Lesions of areas 17 and 18 incurred in adulthood had no significant impact on orienting to auditory white noise stimuli. However, orienting performance to auditory stimuli presented in the peripheral quadrants was slightly superior in the P28 group and reduced in the P1 group. Thus, the visual sparing exhibited by the P1 group may be at the expense of highly proficient orienting to auditory cues. Overall, these results extend our knowledge by showing that in addition to P1-cats, cats that incur lesions of areas 17 and 18 at one month-of-age also exhibit sparing of visually guided orienting, and that the sparing is not confined to a single stimulation paradigm. Finally, the covariation in the magnitude of pathway modifications with the scale of the orienting proficiency in P1- and P28 cats helps to solidify the linkage between rewired brain pathways and spared visually-guided behaviors. PMID- 11099753 TI - The failure of some rats to acquire intravenous cocaine self-administration is attributable to conditioned place aversion. AB - Although cocaine administration in humans includes euphoric and anxiogenic effects, the latter are less well understood. Acute cocaine administration produces aversive effects including anxiogenic effects as well as appetitive effects in rats and mice. In the present study the self-administration and conditioned place preference paradigms were used to determine whether the failure of some rats to acquire intravenous cocaine self-administration is attributable to either an interference with learning or an aversion to cocaine. Rats were classified as self-administrators or non-self-administrators based on the mean number of cocaine self-infusions per session and whether or not rats exhibited either a stable high level of responding or a stable low level of responding. Intravenously administered cocaine produced place preference for the self administrators, while intravenously administered cocaine produced place aversion for the non-self-administrators. The fact that the non-self-administrators showed place aversion is inconsistent with the interpretation that the failure of these rats to readily self-administer is attributable to cocaine-mediated interference of learning. This is the first study in which both the self-administration and the conditioned place preference paradigms have been used in the same animals to demonstrate that the effects of cocaine are appetitive for some rats and aversive for others, and are not an artifact of cocaine's interference with learning. PMID- 11099754 TI - Diazepam modifies the effect of pedunculopontine lesions on morphine but not on amphetamine conditioned place preference. AB - We have previously shown that T-maze learning impairments caused by lesions to the pedunculopontine tegmental nucleus (PPTg) can be reversed by the anxiolytic diazepam. We now report that diazepam also reverses the effect of PPTg lesions on conditioned place preference (CPP) to morphine but not to amphetamine. Rats with bilateral sham or N-methyl-D-aspartate lesions (0.1 or 0.05 M) to the PPTg were trained in a unbiased CPP paradigm with 2 mg/kg morphine or 2 mg/kg D-amphetamine associated with one compartment of the apparatus and vehicle injections in the alternative compartment. After three drug/saline-compartment pairings, the preference of the animals was assessed by allowing them to explore the entire apparatus for 20 min. In contrast to sham-lesioned subjects, the rats with PPTg lesions did not show a preference for the compartment paired with morphine or amphetamine. In two experiments the expression of a morphine CPP was restored by injecting the lesioned animals with 1 mg/kg of diazepam 30 min before the test session. Diazepam pre-treatment did not restore the expression of amphetamine CPP. PMID- 11099755 TI - Nonconvulsive status epilepticus in rats: impaired responsiveness to exteroceptive stimuli. AB - An animal model of human complex partial status epilepticus induced by lithium chloride and pilocarpine administration was developed in our laboratory. The objective of the study was to provide a detailed analysis of both ictal and postictal behavior and to quantify seizure-related morphological damage. In order to determine the animal's responsiveness to either visual or olfactory stimuli, adult male rats were submitted to the following behavioral paradigms: the object response test, the social interaction test, and the elevated plus-maze test. The rotorod test was used to evaluate motor performance. Two weeks after status epilepticus, brains were morphologically examined and quantification of the brain damage was performed. Profound impairment of behavior as well as responsiveness to exteroceptive stimuli correlated with the occurrence of epileptic EEG activity. When the epileptic EEG activity ceased, responsiveness of the pilocarpine-treated animals was renewed. However, remarkable morphological damage persisted in the cortical regions two weeks later. This experimental study provides support for the clinical evidence that even nonconvulsive epileptic activity may cause brain damage. We suggest that the model can be used for the study of both functional and morphological consequences of prolonged nonconvulsive seizures. PMID- 11099756 TI - Memory of an operant response and of depressed mood retained in activation states of 5-HT(1A) receptors: evidence from rodent models. AB - Three series of studies were conducted to specify the role of 5-HT(1A) receptors in memory; using selective ligands that differentially activate 5-HT(1A) receptors, it was determined whether a change in the activation state of these receptors can lead to deficient retrieval, and whether a so-produced deficit can occur in an animal model of depression. First, in vitro studies of [35S]GTPgammaS binding responses identified ligands that differentially activate 5-HT(1A) receptors in rat hippocampus. WAY 100635, 8-OH-DPAT and flesinoxan induced 5 HT(1A) receptor activation that amounted to -2, +50 and +63%, respectively, of that produced by 5-HT. Second, we determined whether changes in the activation state of 5-HT(1A) receptors could impair the retrieval of an operant response in vivo. Rats treated with either a 5-HT(1A) receptor ligand or saline were trained to lever press for milk reward, and were then tested for retrieval with either the same or another treatment. Animals trained with 8-OH-DPAT retrieved the response when tested in the same state, but not when tested in the saline state, and vice versa. Rats trained with 0.16 mg/kg of 8-OH-DPAT also retrieved the response when tested with the other intermediate-efficacy ligand flesinoxan (0.63 mg/kg), but not when tested in a state of lower-magnitude activation (i.e. with 0.16 mg/kg of WAY 100635). Animals trained with 0.16 mg/kg of WAY 100635 retrieved the response when tested in this same state or with saline, but not when tested in a state of intermediate-magnitude activation (i.e. with 0.16 mg/kg of 8-OH-DPAT). Finally, studies using the forced swimming paradigm indicated that the retrieval of learned immobility was similarly dependent upon the activation state of 5-HT(1A) receptors. The findings indicate that changes in activation states of 5-HT(1A) receptors can impair the retrieval of learned responses. It is suggested that depression may in part be acquired in the course of ontogeny and may be available for retrieval in the same but not in other states; various biological rhythms conceivably define such states. PMID- 11099757 TI - Associative learning and latent inhibition in a conditioned suppression paradigm in humans. AB - A paradigm based on conditioned suppression of ongoing motor activity, sensitive to latent inhibition (LI), was developed and tested in healthy volunteers. Subjects were trained to move disks from one peg to another with a high degree of regularity in the Tower of Toronto puzzle, a well-known cognitive skill learning task. Once this was achieved, they were submitted to a Pavlovian conditioning procedure. The conditioned stimulus (CS) was a pure tone and the unconditioned stimulus (US) a loud white noise. The resulting response suppression was assessed by a transient increase in latency of the hand movements. In control subjects, there was non-contingent CS and US presentation. The results evidenced conditioning after a single CS-US pairing. Following five preexposures to the to be-conditioned CS, however, conditioning was abolished, seemingly expressing LI. Because a weak unconditioned response to the tone was observed after its first two presentations, an additional experiment was performed with two preexposures to the to-be-conditioned CS. With such procedure, conditioning was obtained, supporting the existence of LI in the preceding experiment. These results indicate that the present paradigm may be useful for the study of LI in human subjects, having the advantage of being similar to the experimental conditions used in the majority of LI studies in experimental animals. PMID- 11099758 TI - Reduction of latent inhibition by D-amphetamine in a conditioned suppression paradigm in humans. AB - The sensitivity of latent inhibition (LI) to amphetamine has been tested in humans with a paradigm close to the conditioned emotional response suppression currently used in experimental animals. The conditioned stimulus (CS) was a tone, the unconditioned stimulus (US) a strong white noise, and the response a transient delay in a regular sequence of hand movements in the resolution of the Tower of Toronto puzzle. The aim of this study was to verify whether the previously reported, disruptive effect of CS preexposure on conditioning really represents LI, by examining its sensitivity to amphetamine. Three groups of healthy volunteers received placebo, 5 or 10 mg of dexamphetamine sulphate, respectively, in a double-blind experimental design. The preexposure, conditioning and test phases were carried out under either amphetamine or placebo. The non preexposed groups treated with amphetamine were not different from the non preexposed placebo group, indicating that amphetamine did not affect conditioning. Among the preexposed groups, those receiving 10 mg of amphetamine showed normal rates of conditioning, whereas those treated with either 5 mg of amphetamine or placebo showed LI. Similar results have been reported in experimental animals. This sensitivity to amphetamine suggests that the present paradigm may be used to study LI in humans. PMID- 11099759 TI - Increased noradrenergic activity in prefrontal cortex slices of an animal model for attention-deficit hyperactivity disorder--the spontaneously hypertensive rat. AB - Spontaneously hypertensive rats (SHR) are used as a model for attention deficit/hyperactivity disorder (ADHD) since SHR are hyperactive and they show defective sustained attention in behavioral tasks. Using an in vitro superfusion technique we showed that norepinephrine (NE) release from prefrontal cortex slices of SHR was not different from that of their Wistar-Kyoto (WKY) control rats when stimulated either electrically or by exposure to buffer containing 25 mM K(+). The monoamine vesicle transporter is, therefore, unlikely to be responsible for the deficiency in DA observed in SHR, since, in contrast to DA, vesicle stores of NE do not appear to be depleted in SHR. In addition, alpha(2) adrenoceptor mediated inhibition of NE release was reduced in SHR, suggesting that autoreceptor function was deficient in prefrontal cortex of SHR. So, while DA neurotransmission appears to be down-regulated in SHR, the NE system appears to be under less inhibitory control than in WKY suggesting hypodopaminergic and hypernoradrenergic activity in prefrontal cortex of SHR. These findings are consistent with the hypothesis that the behavioral disturbances of ADHD are the result of an imbalance between NE and DA systems in the prefrontal cortex, with inhibitory DA activity being decreased and NE activity increased relative to controls. PMID- 11099760 TI - Hand kinematics during reaching and grasping in the macaque monkey. AB - In this paper, we develop an animal model of prehension movements by examining the kinematics of reaching and grasping in monkeys and by comparing the results to published data on humans. Hand movements were recorded in three dimensions in monkeys who were trained to either point at visual targets under unperturbed and perturbed conditions, or to reach and grasp 3-D objects. The results revealed the following three similarities in the hand kinematics of monkey and man. (1) Pointing movements showed an asymmetry depending on target location relative to the hand used; in particular, movements to an ipsilateral target took longer than those to a contralateral one. (2) Perturbation of target location decreased the magnitude of the velocity peak and increased the duration of pointing movements. (3) Reaching to grasp movements displayed a bell-shaped wrist velocity profile and the maximum grip aperture was correlated with object size. These similarities indicate that the macaque monkey can be a useful model for understanding human motor control. PMID- 11099761 TI - Event-related potentials as a function of movement parameter variations during motor imagery and isometric action. AB - Neuroimaging and electrophysiological studies have shown that executed action and motor imagery activate common neuronal substrates, leading to the hypothesis that movement preparation and motor imagery are functionally equivalent processes. This study further tested the functional equivalence hypothesis by determining whether electrocortical patterns associated with variations in motor control parameters are similar during imagined and executed actions. Event-related potentials (ERPs) were recorded from the supplementary motor/premotor area (SMA/PMA; FCz site) and primary motor area (M1; C3, C4 sites) during an executed and an imagined, cued, discrete isometric contraction task while target force (TF; low, moderate) and rate of force development (RFD; slow, rapid) were varied. For M1, the correlation of ERPs between moderate- and low force-executions was near zero and N2 amplitude was greater for moderate than low force executions, indicating that M1 activity is related to TF. Rapid executions were greater in amplitude and longer in latency than slow executions and the ERPs for rapid- and slow-executions were negatively correlated, indicating that M1 activity is also related to RFD. There were no differences in N2 amplitude and a zero correlation between execution and imagined actions of similar TF and RFD, indicating that neither TF or RFD are represented in M1 activity during imagery. For SMA/PMA, there was a moderate correlation between moderate- and low force-executions and larger N2 amplitude for moderate- than for low force-executions, indicating that TF may be related to SMA/PMA electrocortical activity. ERP patterns were uncorrelated between rapid- and slow-execution at FCz, but N2 amplitude was the same, making it unclear whether the RFD parameter is represented in FCz activity. The correlational and N2 amplitude analyses demonstrate that patterns of electrocortical activity at SMA/PMA are nearly isomorphic during executed and imagined actions as TF and RFD are varied. These results provide evidence that patterns of electrocortical activity associated with variations in the parameters of executed action are similar during motor imagery at SMA/PMA but not at M1. PMID- 11099762 TI - Dissociation of memory and anxiety in a repeated elevated plus maze paradigm: forebrain cholinergic mechanisms. AB - The effect of intraseptal injection of the cholinergic immunotoxin 192-IgG saporin on behavior in the elevated plus maze was investigated. A 5-min test retest paradigm, with minute-by-minute analysis of the first session, was used to evaluate both anxiety and memory in this task. Biochemical analyses revealed a decrease in acetylcholinesterase (AChE) activity in the hippocampus (HPC), septum, and frontal cortex of animals injected with IgG-192 saporin (237.5 ng) when compared with controls. No statistical differences were found between groups in terms of behaviors associated with locomotor activity, conventional measures of anxiety, or ethological behaviors during either session 1 or 2. During test session 2 the controls exhibited decreased exploratory activity and increased indices of anxiety. In contrast, the saporin-treated rats did not exhibit these experience-dependent behavioral changes from session 1 to 2. The minute-by-minute analysis showed a significant decrease in exploratory as well in anxiety associated behaviors during the first session for the control group, but not for the saporin-treated group. These results suggest that the cholinergic innervation of the HPC, the frontal cortex, or both forebrain structures, modulate the initiation of exploratory activity which, results in the acquisition and retention of spatial information, but does not affect the expression of anxiety in the elevated plus-maze. PMID- 11099763 TI - Post-weaning social isolation of male rats reduces the volume of the medial amygdala and leads to deficits in adult sexual behavior. AB - At 21 days of age, gonadally intact male Long Evans rats were weaned and placed into standard laboratory conditions (three per cage) or housed singly. They were tested for noncontact erections and sexual performance at 90 and 220 days of age. Rats raised in isolation displayed significantly fewer noncontact erections in response to sensory cues from an estrous female and fewer intromissions when allowed to mate with a female than did males raised in groups. The volume of the posterodorsal component of the medial amygdala (MePD) and the size of neurons within the MePD were significantly smaller in the isolated males than in socially housed males. Similarly, neurons in the sexually dimorphic nucleus of the preoptic area (SDN-POA) were smaller in isolate animals than in controls. As both MePD volume and SDN-POA soma size are responsive to sex steroids, these differences could result if the isolates experienced lower testosterone levels. Finally, the volume of the overall medial amygdala (MeA) correlated significantly with the number of noncontact erections, a relationship that was not explained by housing condition. These findings highlight the role of social experience as a factor in the sexual differentiation of the brain and suggest a positive relationship between the volume of a brain structure and the display of sexual behaviors. PMID- 11099764 TI - BALB/c mice are not so bad in the Morris water maze. AB - We compared the learning performances of BALB/c mice subjected to the Morris water spatial task under two different lighting conditions. In the first one, the experimental room was lit by neon tubes (direct and bright illumination) and in the second one by a halogen lamp directed to the roof (diffuse illumination). The scores of BALB/c mice in the diffuse illumination condition clearly demonstrated that these mice could learn to escape to a hidden platform while they could not under direct illumination condition. Moreover, they were able to acquire the task by means of spatial cues. These results are interpreted in terms of a decrease of anxiety levels. PMID- 11099765 TI - Yawning responses induced by local hypoxia in the paraventricular nucleus of the rat. AB - Yawing was induced by microinjections of L-glutamate, cyanide and a nitric oxide releasing compound (NOC12) into the paraventricular nucleus of the hypothalamus (PVN) in anesthetized, spontaneously breathing rats. To evaluate physiological aspects of yawning, we monitored intercostal electromyogram (EMG) as an index of inspiratory activity, digastric EMG, blood pressure and electrocorticogram (ECoG). Microinjection of L-glutamate in the medial parvocellular subdivision (mp) elicited a stereotyped yawning response, i.e. an initial depressor response and an arousal shift in ECoG followed by a single large inspiration with mouth opening. The same sequential events were observed during spontaneous yawning, indicating that the mp is responsible for triggering yawning. Microinjection of cyanide into the mp caused the same yawning responses as the ones elicited by microinjection of L-glutamate, suggesting that the mp is sensitive to chemical hypoxia or ischemia within the PVN. Microinjection of NOC12 into the mp elicited a single large inspiration with a variable onset delay, suggesting that diffusible nitric oxide (NO) within the mp may act as a paracrine agent to cause a yawning response. We hypothesize that the mp of the PVN contains an oxygen sensor that causes a yawning response. PMID- 11099766 TI - Spatial learning, contextual fear conditioning and conditioned emotional response in Fmr1 knockout mice. AB - Fmr1 knockout mice are an animal model for fragile X syndrome, the most common form of heritable mental retardation in humans. Fmr1 knockout mice exhibit macro orchidism and cognitive and behavioural deficits reminiscent of the human phenotype. In the present study additional behavioural and cognitive testing was performed. Knockouts and control littermates were subjected to a spatial learning test using a plus-shaped water maze. Animals had to learn the position of a hidden escape platform during training trials. The position of this platform was changed during subsequent reversal trials. Previously reported deficits in reversal learning were replicated, but we also observed significant differences during the acquisition trials. A plus-shaped water maze experiment with daily changing platform positions failed to provide clear evidence for a working memory impairment, putatively underlying the spatial learning deficits. Two different test settings were used to examine the reported deficit of Fmr1 knockout mice in fear conditioning. Conditioned fear responses were observed in a contextual fear test, and the ability to acquire an emotional response was tested by means of response suppression in a conditioned emotional response procedure. Neither protocol revealed significant differences between controls and knockouts. PMID- 11099767 TI - Defeat followed by individual housing results in long-term impaired reward- and cognition-related behaviours in rats. AB - In contrast to the well-documented acute effects on behavioural sensitivity, chronic effects that persist for weeks or even months after the cessation of the stressor received relatively little attention. This study aimed at the long-term effects of a severe stressor, i.e. social defeat followed by individual housing. Defeated and subsequently individually housed animals displayed impaired social memory, decreased social interaction and diminished anticipation for a sucrose solution for up until a period of 3 months after defeat. Remarkably, social housing counteracted the defeat-induced effects. The impaired capability to anticipate for a reward was discussed in relation to anhedonia, an important symptom of human depression. Moreover, the disturbed memory, the chronic nature of the effects, and the therapeutic effects of social housing, suggest that the defeat model may serve as a potential model for human psychopathology. PMID- 11099768 TI - Oxiracetam prevents the MK-801 induced amnesia for the elevated plus-maze in mice. AB - We investigated the effect of the nootropic substance oxiracetam on the impairment of memory induced in mice by the non-competitive NMDA antagonist MK 801. Memory capacities of animals having different experience were evaluated using the elevated plus-maze test. Oxiracetam was injected immediately after the acquisition session(s), MK-801 was given 30 min before the retention session which followed 24 h after the acquisition session(s). In slightly experienced animals (Section 3.1), oxiracetam (3 and 30 mg/kg, s.c.) prevented MK-801 (0.15 mg/kg, i.p.) induced memory deficits characterized by a prolongation of the transfer latency. In well-trained animals (Section 3.2), oxiracetam (30 mg/kg, s.c.) attenuated MK-801 (0.15,0. 25 and 0.4 mg/kg, i.p.) induced amnesia for a spatial orientation in the elevated plus-maze. These results show that oxiracetam interacted with the glutamatergic NMDA receptor system and forestalled the impairment of retrieval of long-term memory. The results also justify the usage of the elevated plus-maze method in the evaluation of potential anti-amnesic or nootropic drugs. PMID- 11099769 TI - Role of the substantia nigra pars reticulata in sensorimotor gating, measured by prepulse inhibition of startle in rats. AB - The substantia nigra pars reticulata (SNR) is one of the major output nuclei of the basal ganglia. It connects the dorsal and ventral striatum with the thalamus, superior colliculus and pontomedullary brainstem. The SNR is therefore in a strategic position to regulate sensorimotor behavior. We here assessed the effects of SNR lesions on prepulse inhibition (PPI) of the acoustic startle response (ASR), stereotypy and locomotion in drug-free rats, as well as after systemic administration of the dopamine agonist DL-amphetamine (2 mg/kg), and the NMDA receptor antagonists dizocilpine (0.16 mg/kg) and CGP 40116 (2 mg/kg). SNR lesions reduced PPI, enhanced spontaneous sniffing and potentiated the locomotor stimulation by dizocilpine and CGP 40116. PPI was impaired by dizocilpine and CGP 40116 in controls. The ASR was enhanced in controls by dizocilpine and amphetamine. SNR lesions prevented the enhancement of the ASR by amphetamine. A second experiment tested the hypothesis that the SNR mediates PPI via a GABAergic inhibition of the startle pathway. Infusion of the GABA(B) antagonist phaclofen but not the GABA(A) antagonist picrotoxin into the caudal pontine reticular nucleus reduced PPI. Hence, lesion of the SNR reduces sensorimotor gating possibly by elimination of a nigroreticular GABAergic projection interacting with GABA(B) receptors. Moreover, destruction of the SNR enhances the motor stimulatory effects of amphetamine and of the NMDA antagonists dizocilpine and CGP 40116. We conclude that the SNR exerts a tonic GABAergic inhibition on sensorimotor behavior that is regulated by the dorsal and the ventral striatum. PMID- 11099770 TI - Early exposure to chronic variable stress facilitates the occurrence of anhedonia and enhanced emotional reactions to novel stressors: reversal by naltrexone pretreatment. AB - The present research studied the influence of an early chronic variable stress (CVS) paradigm - an animal model of depression - on behavioral responses to subsequent environmental challenges suggested to model anhedonia and emotional reactions such as anxiety and fear. In order to explore a potential involvement of an endogenous opiate mechanism - presumably activated during CVS exposure - in the development of such behavioral reactions, in all experiments rats were administered naltrexone (NAL, 2 mg/kg, i.p.) or vehicle (VH) prior to each daily stressor of the CVS procedure. Animals were exposed to CVS and 1 week later tested for sucrose preference (1%) in a free choice paradigm after the presentation or not of a 90-min restraint period. Only CVS treated animals that were later exposed to restraint showed a reduction of sucrose preference, this reduction was absent when CVS rats were pretreated previously with NAL. Moreover, CVS rats were one week later tested on the elevated plus maze (EPM) and in their conditioned and unconditioned freezing response to a single shock session. Early chronic stress resulted in an anxiogenic behavior in the EPM and in an enhanced conditioned and unconditioned freezing which were all abolished by NAL pretreatment. These behavioral findings suggest that the potential activation of an endogenous opiate mechanism during CVS participates in the development of anhedonia and exaggerated emotional reactions in response to subsequent stressful experiences. PMID- 11099771 TI - Lesions of the medial shell of the nucleus accumbens impair rats in finding larger rewards, but spare reward-seeking behavior. AB - The goal of this study was to help better understand the importance of the nucleus accumbens (Nacc) in the processing of position and reward value information for goal-directed orientation behaviors. Sixteen male Long-Evans rats, under partial water deprivation, were trained in a plus-maze to find water rewards in the respective arms which were lit in pseudo-random sequence (training trials). Each day one reward arm was selected to deliver six drops of water (at 1 s intervals) the others provided only one drop per visit. After 32 visits, probe trials were intermittently presented among training trials. Here, all four arms were lit and offered the previously assigned reward. The rats rapidly learned to go to the highly rewarded arm. Six trained rats were given bilateral electrolytic lesions in the Nacc shell, two others had unilateral lesions and eight had sham operations (with approved protocols). Field potentials evoked by fornix stimulation were recorded in lesion electrodes to guide placements. Only the lesioned rats showed significant impairments (P<0.05) in selecting the greater reward on probe trials. However on training trials, lesioned (and sham-operated) rats made only rare errors. While the motivation to drink and the capacity for cue-guided goal-directed orientation behavior was spared, lesioned rats were impaired in learning the location of the larger reward. The accumbens lesions apparently impaired integration of position and reward value information, consistent with anatomical and electrophysiological data showing the convergence of hippocampal, amygdalar, ventral tegmental area (VTA) and prefrontal cortical inputs there. PMID- 11099772 TI - Balance control and posture differences in the anxious BALB/cByJ mice compared to the non anxious C57BL/6J mice. AB - A relation between anxiety disorders and balance control dysfunctions has been observed in many studies in humans. A mismatch in the integration of sensory inputs could trigger these disturbances. Very few experimental animal procedures have been designed to study the functional link between anxiety and balance control. A task was therefore developed, challenging the visual, vestibular and somesthesic sensory systems in mice. The test, called the 'rotating beam', gave an accurate assessment of balance control and the posture, using sensitive measures (number of falls and imbalances, position of tail and trunk). Striking differences were observed between the two inbred strains of mice known to have radically different anxiety-related behaviour. The highly anxious strain, BALB/cByJ, performed poorly compared to the non anxious strain, C57BL/6J. Balance control and postural abilities of anxious mice were improved by acute anxiolytic diazepam treatment. Lower behavioural performance level was registered in non anxious mice given anxiogenic beta-CCM treatment. The findings account for a strong relationship between anxiety and balance control in mice. Finally, the highly sensitive procedure proved to be well suited to the study of functional links between anxiety and sensorimotor processes. PMID- 11099773 TI - Effect of subtype selective nicotinic compounds on attention as assessed by the five-choice serial reaction time task. AB - Nicotine can improve attentional functioning in humans, and a number of studies have recently demonstrated that under specific task conditions, nicotine can also improve attention in the rat. Neuronal nicotinic receptors comprise combinations of alpha(2-9) and beta(2-4) subunits, arranged to form a pentameric receptor, with the principal CNS subtypes currently believed to be alpha(4)beta(2) and a homomeric alpha(7) receptor. In the present studies, we attempted to delineate the particular nicotinic receptor subtype(s) contributing to the effects of nicotine on attention by assessing various nicotinic ligands on performance in the five-choice serial reaction time task (5-CSRTT). In rats performing below criterion (<80% correct, >20% omissions to a 1-s visual stimulus), subchronic dosing with nicotine (0.2 mg/kg sc) and the alpha(4)beta(2) agonist SIB 1765F (5 mg/kg sc) increased correct responding and decreased response latencies across the treatment week; whereas the alpha(7) agonist AR-R 17779 (20 mg/kg sc) was without effect. In subjects meeting the criterion, the competitive high affinity (including alpha(4)beta(2)) nicotine receptor antagonist DHbetaE (1-10 mg/kg sc) and the alpha(7) antagonist methyllycaconitine (MLA: 5-10 mg/kg i.p.) did not disrupt performance, whereas at the highest dose, the non-competitive antagonist mecamylamine (0.3-3 mg/kg sc) decreased accuracy and increased response latencies. These changes bore some similarities to those of pre-feeding and the non-competitive NMDA antagonist dizocilpine (0.03-0.06 mg/kg sc), suggesting that mecamylamine-induced performance disruption may relate to non-nicotinic receptor effects. In subjects chronically treated with nicotine, acute nicotine challenge (0.4 mg/kg sc) significantly increased accuracy whilst having no effect on any other performance measures. Finally, in these same nicotine pre-treated rats, the decrease in latency and increase in premature responses induced by nicotine (0.2 mg/kg sc) to a target stimulus of 150 ms was fully antagonised by DHbetaE (3 mg/kg sc) but not MLA (5 mg/kg i.p.). These results suggest that alpha(7) receptors do not play a role in any of the behavioural effects of nicotine observed in the 5-CSRTT, whereas a high affinity site, perhaps alpha(4)beta(2), is more likely involved. PMID- 11099774 TI - Auditory cues support place navigation in rats when associated with a visual cue. AB - Rats, like other crepuscular animals, have excellent auditory capacities and they discriminate well between different sounds [Heffner HE, Heffner RS, Hearing in two cricetid rodents: wood rats (Neotoma floridana) and grasshopper mouse (Onychomys leucogaster). J Comp Psychol 1985;99(3):275-88]. However, most experimental literature concerning spatial orientation almost exclusively emphasizes the use of visual landmarks [Cressant A, Muller RU, Poucet B. Failure of centrally placed objects to control the firing fields of hippocampal place cells. J Neurosci 1997;17(7):2531-42; and Goodridge JP, Taube JS. Preferential use of the landmark navigational system by head direction cells in rats. Behav Neurosci 1995;109(1):49-61]. To address the important issue of whether rats are able to achieve a place navigation task relative to auditory beacons, we designed a place learning task in the water maze. We controlled cue availability by conducting the experiment in total darkness. Three auditory cues did not allow place navigation whereas three visual cues in the same positions did support place navigation. One auditory beacon directly associated with the goal location did not support taxon navigation (a beacon strategy allowing the animal to find the goal just by swimming toward the cue). Replacing the auditory beacons by one single visual beacon did support taxon navigation. A multimodal configuration of two auditory cues and one visual cue allowed correct place navigation. The deletion of the two auditory or of the one visual cue did disrupt the spatial performance. Thus rats can combine information from different sensory modalities to achieve a place navigation task. In particular, auditory cues support place navigation when associated with a visual one. PMID- 11099775 TI - Novelty enhances retrieval of one-trial avoidance learning in rats 1 or 31 days after training unless the hippocampus is inactivated by different receptor antagonists and enzyme inhibitors. AB - Rats were implanted bilaterally with cannulae in the CA1 region of the dorsal hippocampus. The animals were trained in one-trial step-down inhibitory avoidance and tested either 1 or 31 days later. Some of the animals were exposed, 1 h prior to retention testing, to a novel environment. This was a 50-cm high, 50-cm wide and 39-cm high wooden box covered on the inside with black plastic. Through the cannulae, 10 min prior to the retention test, the rats received 0.5-microl infusions of saline, of a vehicle (2% dimethylsulfoxide in saline), or of the following drugs: the glutamate NMDA receptor blocker, aminophosphonopentanoic acid (AP5, 5.0 microg), the AMPA receptor blocker, 6,7-cyanonitroquinoxaline-2,3 dione (CNQX, 1.25 microg), the generic glutamate metabotropic receptor antagonist, alpha-methyl-(4-carboxyphenyl)glycine (MCPG), the inhibitor of cAMP dependent protein kinase (PKA), Rp-cAMPs (0.1 or 0.5 microg), or the inhibitor of the mitogen-activated protein kinase (MAPK), PD098059 (10 or 50 microM). CNQX and PD098059 were dissolved in the vehicle; AP5 and Rp-cAMPs were dissolved in saline. All these drugs except AP5 had been previously found to alter retrieval of this task. Novelty markedly enhanced retention test performance of the avoidance task. The drugs, in accordance with previous results, and with the exception of AP5 at any of the two training-test intervals and of CNQX at the 31 day interval, hindered retention test performance. The results indicate that the effect of novelty on retrieval can not be observed if the major biochemical mechanisms of retrieval (AMPA receptors, PKA, MAPK) are blocked, i.e. if the hippocampus was temporarily inactivated by drugs that inhibit those mechanisms. PMID- 11099776 TI - Glucose deprivation and chemical hypoxia: neuroprotection by P2 receptor antagonists. AB - In this work we investigate cell survival after glucose deprivation and/or chemical hypoxia and we analyse the neuroprotective properties of selected antagonists of P2 ATP receptors. We find that in rat cerebellar granule neurones, the antagonist basilen blue prevents neuronal death under hypoglycaemia. Basilen blue acts through a wide temporal range and it retains its efficacy under chemically induced hypoxic conditions, in the presence of the respiratory inhibitors of mitochondria electron transport chain complexes II (3 nitropropionic acid) and III (antimycin A). In spite of the presence of these compounds, basilen blue maintains normal intracellular ATP levels. It furthermore prevents neuronal death caused by agents blocking the mitochondrial calcium uptake (ruthenium red) or discharging the mitochondrial membrane potential (carbonyl cyanide m-chlorophenylhydrazone). Inhibition of poly (ADP-ribose) polymerase, modulation of the enzyme GAPDH and mitochondrial transport of mono carboxylic acids are not conceivable targets for the action of basilen blue. Survival is sustained by basilen blue also in CNS primary cultures from hippocampus and in PNS sympathetic-like neurones. Partial neuroprotection is furthermore provided by three additional P2 receptor antagonists: suramin, pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid 4-sodium and 4,4' diisothiocyanatostilbene-2,2'disulphonic acid. Our data suggest the exploitation of selected P2 receptor antagonists as potential neuroprotective agents. PMID- 11099777 TI - Hypoglycaemia-induced cell death: features of neuroprotection by the P2 receptor antagonist basilen blue. AB - Our previous work in neuronal cultures has shown that several antagonists of P2 ATP receptors prevent cell death evoked by hypoglycaemia, chemical hypoxia, mitochondria dysfunction, as well as glutamate-dependent excitotoxicity and low potassium-induced apoptosis. Experiments are now designed to examine which biological pathway contributes to cell death/survival under glucose starvation. We show here that, consequently to hypoglycaemic insults, cerebellar granule neurones undergo a combination of apoptosis and necrosis both inhibited by the P2 receptor antagonist basilen blue. This is demonstrated by morphological and biochemical features, such as TdT-mediated dUTP-biotin nick end-labelling, fluorescent staining of nuclear chromatin using Hoechst 33258, direct counting of intact viable nuclei and extracellular releasing of the cytosolic enzyme LDH. Furthermore, we show that hypoglycaemia induces outflow of cytochrome c from mitochondria and it up-regulates heat-shock proteins HSP70, but not HSP90, glucose-regulated proteins GRP75 and GRP78, as well as expression and activity of the enzyme caspase-2. Basilen blue can modulate only some of these effects. Our data contribute to dissect the role played by P2 receptor antagonism in sustaining neuroprotection against metabolic stresses. PMID- 11099778 TI - Pharmacological evidence for a role of gamma-aminobutyric acid A receptor mechanism in modulating nitric oxide synthase activity in rat brain. AB - The role of gamma-aminobutyric acid (GABA) mechanism on the synthesis of nitric oxide (NO) has been investigated by measuring the activity of nitric oxide synthase (NOS) and the concentration of NO in rat brain 15 min after administration of anticonvulsant doses of diazepam (0.25 and 0.5 mg/kg) which is known to activate GABA A receptor for its anticonvulsant action. Diazepam enhanced both NOS activity and the concentration of NO in a dose-dependent manner. A reversal has been observed in animals treated with a convulsant dose of picrotoxin (5 mg/kg) which is known to produce convulsions by blocking GABA A receptor mechanism. These results suggest that a functional interaction occurs between GABA A receptor activity and NO synthesis in the brain. PMID- 11099779 TI - Inhibition of adenylate cyclase activity by 5-aminolevulinic acid in rat and human brain. AB - The effect of the haem precursor 5-aminolevulinic acid (ALA) on the production of cyclic adenosine-monophosphate (cAMP) by rat cerebellar membranes was investigated. It was found that ALA dose-dependently decreased cAMP levels (maximal inhibition of 38%, at 1 mM), due to an inhibition of basal adenylate cyclase activity. ALA also inhibited fluoride- and Gpp(NH)p-stimulated, but not the forskolin-stimulated adenylate cyclase activity. 5-Aminovaleric acid (an inhibitor of GABA(B) receptors) did not prevent the inhibition, indicating that it was not mediated by the activation of the G(i)-protein coupled GABA(B) receptor. In addition, the nucleotide binding site of G-protein appeared not to be affected by ALA since it did not inhibit [3H]Gpp(NH)p binding to our membrane preparation. Antioxidants (glutathione, ascorbate and trolox) completely prevented the inhibition indicating that ALA effect was mediated by an oxidative damage of adenylate cyclase. ALA also inhibited the activity of adenylate cyclase in membranes isolated from rat cortex and striatum and from human cortex. These results may be of value in understanding the neurochemical mechanisms underlying the neurotoxic effects of ALA. PMID- 11099780 TI - Reversible and irreversible acetylcholinesterase inhibitors cause changes in neuronal amyloid precursor protein processing and protein kinase C level in vitro. AB - The alternative routes of cleavage of the amyloid precursor protein (APP) result in the generation and secretion of both soluble APP and beta-amyloid, the latter being the main component of the amyloid deposits in the brains of individuals with Alzheimer's disease (AD). This study examined the question of whether acetylcholinesterase (AChE) inhibitors can alter the processing of APP and the level of protein kinase C (PKC) in primary rat basal forebrain cultures. Western blotting was used to test two AChE inhibitors (reversible and irreversible) for their ability to enhance the release of APP and PKC content. These inhibitors were ambenonium (AMB) and metrifonate (MTF), at different concentrations. A significant increase was found in the cell-associated APP level in a basal forebrain neuronal culture, and there was an elevation of the APP release into the medium. Increases were similarly observed in the PKC levels after AMB or MTF treatment. The results suggest that these AChE inhibitors promote the non amyloidogenic route of APP processing, which may be due to their stimulatory effects on PKC. The PKC activation may enhance the alpha-secretase activity and consequently the production of the N-terminal APP. Since both a decreased level of APP secretion and a low activity and level of PKC may be involved in the pathogenesis of AD, it is concluded that the administration of AChE inhibitors to AD patients may facilitate the memory processes and exert a neuroprotective effect. PMID- 11099781 TI - TGF-beta1 inhibits caspase-3 activation and neuronal apoptosis in rat hippocampal cultures. AB - The effect of TGF-beta1 on apoptosis varies depending on the cell type, the kind of stimulus and the experimental conditions. The present study attempted to identify whether TGF-beta1 can prevent neuronal apoptosis and interrupt caspase-3 activation in rat primary hippocampal cultures after staurosporine treatment. TGF beta1 at the concentration of 1 and 10 ng/ml significantly reduced neuronal damage as detected by trypan blue exclusion. Nuclear staining with Hoechst 33258 and TUNEL-staining further demonstrated that TGF-beta1 at the same concentration range effectively diminished neuronal apoptosis 24 h after staurosporine treatment, whereas 0.1 ng/ml of TGF-beta1 did not. Furthermore, TGF-beta1 (1 and 10 ng/ml) markedly inhibited the activation of caspase-3 induced by staurosporine as demonstrated by both caspase-3 activity assay and Western blotting. This study provides evidence that TGF-beta1 is able to efficiently inhibit caspase-3 activation, and thereby protects cultured hippocampal neurons against apoptosis. PMID- 11099782 TI - Low temperature prevents potentiation of norepinephrine release by phenylephrine. AB - The effect of 1-phenylephrine (1-PE), an alpha(1)-receptor agonist, was investigated on the release of tritiated norepinephrine ([3H]NE). Pairs of guinea pig vasa deferentia were loaded with [3H]NE, superfused continuously, and stimulated electrically. 1-PE (10, 100 microM) enhanced the basal release of tritium in concentration-dependent manner. The stimulation-evoked release of radioactivity was significantly increased by 100 microM 1-PE. Both basal and stimulation-evoked release by 1-PE were reduced by desipramine (10 microM), a monoamine uptake inhibitor. The effect of 1-PE on basal release was independent on extracellular Ca(2+) concentration ([Ca(2+)](o)) and alpha(1)-adrenoceptor blockade. However, the 1-PE-induced release was temperature dependent: at low temperature 1-PE failed to increase either basal or stimulation-evoked release of NE. Using three different temperatures (7, 12, 17 degrees C, respectively), it was found that basal release was blocked at all three temperature values but the stimulation-evoked release was inhibited only at the lower values. The effect of 1-PE on the NE release appears to involve a desipramine-, and temperature sensitive process. These results suggest that a non-receptorial and direct carrier-mediated mechanism is involved in NE releasing effect of 1-PE. PMID- 11099783 TI - A new in vitro approach for investigating the MPTP effect on DA uptake. AB - Previous studies have shown that dopamine (DA) uptake was decreased after preincubation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or 1-methyl 4-phenylpyridinium (MPP(+)) in in vitro slice and synaptosome models. The present study, conducted with and without preincubation, attempted to determine whether inhibition results from a direct effect of neurotoxins on neuronal DA transporter or from an alteration of the transporter secondary to other toxic events. DA uptake was inhibited about 50% in the presence of MPTP+O(2) or MPP(+) (0.1, 1 and 5 mM) in rat striatal slices and synaptosomes. Such inhibition was obtained in synaptosomes preincubated for 150 min with MPP(+) and then washed. Inhibition of DA uptake was lower in slices preincubated with MPTP (5 mM)+O(2) and then washed (30%). Experiments in synaptosomes prepared from slices preincubated with MPTP or MPP(+) showed greater inhibition of DA uptake with MPTP. The results suggest that the inhibition of DA uptake in vitro by MPTP or MPP(+) results initially from a direct effect on the transporter during its penetration in nerve endings and subsequently from a transporter alteration related to toxic events. Thus, the preincubation of striatal slices followed by DA uptake measurement in synaptosomes would appear to be a good in vitro model for studying the dopaminergic toxicity of MPTP. PMID- 11099784 TI - Effects of tumor necrosis factor alpha on taurine uptake in cultured rat astrocytes. AB - Taurine is known to play a major role in volume regulation in astrocytic swelling associated with stroke and brain trauma. Apart from brain edema, the severity of brain injury is related to the levels of inflammatory cytokines such as tumor necrosis factor alpha (TNFalpha). TNFalpha had been shown to be closely associated with brain edema formation since the neutralization of TNFalpha reduced brain edema. Considering taurine has osmoregulatory functions in astrocytes, experiments were performed to study the effects of TNFalpha on taurine uptake in cultured astrocytes. Astrocytes exposed to 20 ng/ml of TNFalpha for 48 h showed a 91% increase in taurine uptake and significant increase was observed after 24 h exposure. This cytokine caused neither significant changes in cell volume nor taurine release. The increased in taurine uptake induced by TNFalpha was unlikely resulted from the modification of Na(+) movement because TNFalpha decreased tyrosine uptake, Na(+)-dependent transport system. In contrast to TNFalpha, interferon-gamma (IFNgamma) did not significantly affect taurine uptake. Taken together, our results did not support a suggestion that TNFalpha affects cell volume regulation via modulating taurine uptake in astrocytes. Increasing lines of evidence have demonstrated that taurine has anti-inflammatory and anti-oxidative effects, these findings therefore suggested that the increase in taurine uptake might be an adaptive response or a tool for astrocytes against oxidative stress. PMID- 11099785 TI - Characterization of electrically evoked [3H]-D-aspartate release from hippocampal slices. AB - Electrical stimulation has certain advantages over chemical stimulation methods for the study of neurotransmitter release in brain slices. However, measuring detectable quantities of electrically evoked release of endogenous or radiolabeled markers of excitatory amino acid neurotransmitters has required current intensities or frequencies much higher than those usually required to study other transmitter systems. We demonstrate here that [3H]-D-aspartate (D ASP) release can be detected from hippocampal slices at lower stimulation intensities in the presence of a glutamate reuptake inhibitor. Subsequently, we optimized the electrical stimulus parameters for characterizing electrically evoked D-ASP release. Under the experimental conditions described, greater than 90% of electrically evoked D-ASP release is calcium-dependent. Evoked D-ASP release is markedly reduced by pre-treating slices with the synaptic vesicle toxin bafilomycin A1 (BAF A1) or in the presence of 10-mM magnesium. Evoked D-ASP release is also reduced to variable degrees by N- and P/Q type voltage-sensitive calcium channel antagonists. Neither spontaneous efflux nor evoked D-ASP release were affected by NMDA, AMPA or group I metabotropic glutamate receptor (mGluR) antagonists. Evoked D-ASP release was reduced in the presence of an adenosine A1 receptor agonist and potentiated by treatment with a group I mGluR5 agonist. Evoked [3H]-D-ASP release was similar in magnitude to evoked [3H]-L-glutamate (L GLU) release. Finally, in separate experiments using the same electrical stimulus parameters, more than 90% of electrically evoked endogenous L-GLU release was calcium dependent, a pattern similar to that observed for evoked [3H]-D-ASP release. Taken together, these results indicate that electrically evoked [3H]-D ASP release mimics evoked glutamate release in brain slices under the experimental conditions employed in these studies. PMID- 11099786 TI - Activation of beta-adrenoceptors opens calcium-activated potassium channels in astroglial cells. AB - In the present study, effects of the alpha(2)- and beta-adrenoceptor agonists clonidine and isoproterenol on astrocytes in astroglial/neuronal cocultures from rat cerebral cortex were evaluated. The calcium- and potassium-sensitive dyes fura-2 and potassium-binding benzofuran isophtalate (PBFI) were used to study alterations in intracellular concentrations of calcium ([Ca(2+)](i)) and potassium ([K(+)](i)), respectively, while the perforated patch clamp technique was used to analyze transmembrane currents. Exposure to isoproterenol or clonidine elicited an immediate increase in [Ca(2+)](i) that was totally abolished in calcium-free extracellular media. Isoproterenol also decreased [K(+)](i), but clonidine did not. The reduction in [K(+)](i) was inhibited in Ca(2+)-free media. As evaluated with the perforated patch technique, isoproterenol (10(-6)-10(-4) M) induced a slowly developing and long lasting outward current that also was totally abolished in calcium-free buffer. This current was blocked by external tetraethylammonium (TEA, 10 mM) and charybdotoxin (ChTX, 10 nM), but was not affected by apamin (50 nM). The current-to-voltage (I V) relationships for the isoproterenol-induced currents showed a markedly negative reversal potential, -96 mV+/-7, (mean+/-S.D., n=5). These results suggest that the stimulation of astroglial beta-adrenoceptors by isoproterenol opens calcium-activated potassium channels (K((Ca))). Preincubation with forskolin significantly increased the isoproterenol-induced currents compared with controls, indicating that the opening of astroglial K((Ca)) channels after beta-adrenergic stimulation not only depends on [Ca(2+)](i) but also synergistically involves the cAMP transduction system to which beta-adrenoceptors are known to be positively coupled. PMID- 11099787 TI - Characterization with [3H]quisqualate of group I metabotropic glutamate receptor subtype in rat central and peripheral excitable tissues. AB - Radioligand binding studies were performed to label metabotropic glutamate receptor (mGluR) in rat brain synaptic membranes using [3H]quisqualic acid (QA) synthesized in our laboratory as a radioligand. In the presence of ionotropic glutamate receptor (iGluR) agonists, including N-methyl-D-aspartic (NMDA), DL alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic (AMPA) and kainic acids (KA), at concentrations maximally effective in displacing each receptor binding, the agonists for group I mGluR subtype (+/-)-1-aminocyclopentane-trans-1,3 dicarboxylic acid (trans-ACPD) and (S)-3,5-dihydroxyphenylglycine ((S)-3,5-DHPG) more potently displaced [3H]QA binding in a concentration-dependent manner than their absence. The addition of these three iGluR agonists did not significantly affect potencies of (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG-IV) and L-(+)-2-amino-4-phosphonobutyric acid (L-AP4) to displace [3H]QA binding. Scatchard analysis revealed that [3H]QA binding consisted of a single component with a maximal number of binding sites (B(max)) of 431.6 fmol/mg protein and a dissociation constant (K(d)) of 50.9 nM, in the presence of the three iGluR agonists. [3H]QA binding was markedly inhibited by GTP and its analogues; but not by GDP, GMP and ATP, under these conditions. Inhibition by GTP was seen in all central structures examined, but [3H]QA binding was not detectable in peripheral tissues, such as pituitary and adrenal glands. Neither reverses transcription polymerase chain reaction nor immunoblotting analysis demonstrated the expression of mGluR1 and mGluR5 subunits in the aforementioned two peripheral tissues. These results suggest that [3H]QA indeed labels group I mGluR subtype functionally coupled to GTP binding protein in rat brain synaptic membranes under the experimental conditions employed. Group I mGluR subtype seems to be selectively distributed in central structures but not in pituitary and adrenal glands. PMID- 11099788 TI - Dialysate dihydroxyphenylglycol as a window for in situ axoplasmic norepinephrine disposition. AB - To examine basal axoplasmic norepinephrine (NE) kinetics at the in situ cardiac sympathetic nerve ending, we applied a dialysis technique to the heart of anesthetized cats and performed the dialysate sampling with local administration of a pharmacological tool through a dialysis probe. The dialysis probe was implanted in the left ventricular wall, and dihydroxyphenylglycol (DHPG, an index of axoplasmic NE) levels were measured by liquid chromatogram-electrochemical detection. Control dialysate DHPG levels were 161+/-19 pg/ml. Pargyline (monoamine oxidase inhibitor, 1 mM) decreased the dialysate DHPG levels to 38+/ 10 pg/ml. Further alpha-methyl-para-tyrosine, omega-conotoxin GVIA, desipramine (NE synthesis, release and uptake blockers) decreased the dialysate DHPG levels to 64+/-19, 106+/-15, 110+/-22 pg/ml, respectively. In contrast, reserpine (vesicle NE transport inhibitor, 10 microM) increased the dialysate DHPG levels to 690+/-42 pg/ml. Thus, NE synthesis, metabolism and recycling (release, uptake and vesicle transport) affected basal intraneuronal NE disposition at the nerve endings. Measurement of DHPG levels through a dialysis probe provides information about basal intraneuronal NE disposition at the cardiac sympathetic nerve endings. Yohimbine (alpha(2)-adrenoreceptor blocker, 10 microM) and U-521 (catechol-O-methyltransferase blocker, 100 microM) did not alter the dialysate DHPG levels. Furthermore, there were no significant differences in the reserpine induced DHPG increment between the presence and absence of desipramine (10 microM) or alpha-methyl-para-tyrosine (100 mg/kg i.p.). These results may be explained by the presence of two axoplasmic pools of NE, filled by NE taken up and synthesized, and by NE overflow from vesicle. The latter pool of NE may be closed to the monoamine oxidase system in the axoplasma. PMID- 11099789 TI - Relief of dyspeptic symptoms by colloidal bismuth subcitrate in Helicobacter negative and -positive patients: results of a study in general practice. AB - BACKGROUND: The role of H. pylori in non-ulcer dyspepsia is controversial. Colloidal bismuth subcitrate (CBS) is known to suppress H. pylori. We hypothesized that if H. pylori is a causal factor in dyspepsia, then suppression of H. pylori would lead to a decrease in symptoms. AIM: To assess the relationship between H. pylori status and the effect of CBS on dyspeptic symptoms in patients visiting their general practitioner for dyspeptic complaints. METHODS: In total 446 patients between 17 and 81 years of age (median 44 years) were included. All patients were treated with CBS (240 mg Bi2O3) twice a day for 4 weeks. Symptoms were scored at baseline, and after 2 and 4 weeks of treatment. At the first visit, blood was taken for serological H. pylori testing. RESULTS: During follow up, 65 patients were lost due to violation of protocol. Positive H. pylori serology was found in 110 (24.7%) of the 446 initially selected patients, and in 90 (23.6%) of the 381 patients who completed the protocol (NS). The mean overall symptom score decreased significantly after 4 weeks of CBS (P<0.001). This reduction in overall symptom score was not significantly different between the H. pylori-positive and -negative groups. CONCLUSIONS: The H. pylori status does not influence the outcome of CBS therapy in patients who consult their general practitioner for dyspepsia. This finding suggests that H. pylori does not play an important role in the etiology of dyspepsia in patients seen by the general practitioner. PMID- 11099790 TI - Totally implantable venous access devices: evaluation of complications and a prospective comparative study of two different port systems. AB - BACKGROUND: Totally implantable venous access devices (TIVADs) are valuable instruments in case prolonged intravenous therapy is required, but implantation and use of these devices are associated with complications. The purpose of this study was to evaluate perioperative and long-term complications associated with TIVADs. In addition, we compared two different types of TIVADs with respect to implantation, care protocol and patients' comfort. METHODS: In a retrospective study perioperative and long-term complications in a general oncology population were analysed. In a prospective randomized study comparison of two types of TIVADs was carried out. RESULTS: Perioperative complications occurred in 27 (21.4%) of 126 implanted TIVADs: catheter malposition (16.7%) in 21 patients, pneumothorax (0.8%) in one and haemorrhage (4.0%) in five. Long-term complications appeared in 31 (25.2%) out of 123 TIVADs: thrombosis in 9 (7.3%), especially associated with malposition of the tip of the catheter; infection in 10 (8.1%); extravasation in 2 (1.6%); migration of the catheter tip in 6 (4.8%); pain at reservoir in 3 (2.4%) and inaccessibility of the port in 1 (0.8%). No significant differences were found with respect to implantation, care accessibility and patients' comfort between the two TIVADs. CONCLUSIONS: The use of TIVADs is associated with some risk of serious perioperative and long-term complications. In case of thrombotic complications these systems can be saved with appropriate treatment. Correct positioning of the catheter tip is essential to prevent thrombotic complications. In case of TIVAD-related infectious complications, the possibility of saving the TIVAD depends on the causative microorganism and type of infection. Furthermore, to increase patients' satisfaction with TIVADs they should be well informed about the surgical procedure and possible disadvantages of these devices. PMID- 11099791 TI - The difficult diagnostic approach of the intraperitoneal mesothelioma. AB - The malignant mesothelioma is a rare tumour with a rising incidence during the past decades [Lancet 1 (1986) 1275; Br J Int Med 28 (1971) 59; Am J Int Med 9 (1986) 397]. The tumour is derived from the mesothelial lining of the pleural cavity, the pericardium or the peritoneum. Mesotheliomas solely involving the peritoneum are particularly rare. Two cases of malignant peritoneal mesotheliomas are described to demonstrate the difficulties in diagnosing this disease. Surgical exploration, either by laparotomy, or preferably by laparoscopy, has been proven valuable as a diagnostic method. PMID- 11099792 TI - Pulmonary hypertension with limited cutaneous scleroderma (CREST syndrome). AB - A patient is described with a typical manifestation of pulmonary hypertension associated with limited cutaneous scleroderma, also known as CREST syndrome. The patient was treated with a calcium antagonist, oral anticoagulation and, because of evidence for parenchymal inflammation of the lung, with low-dose prednisone and cyclophosphamide. This treatment resulted in initial improvement of diffusion capacity and exercise tolerance, however, 1 year after diagnosis the patient died of progressive pulmonary hypertension. PMID- 11099793 TI - Mycophenolate mofetil, Cellcept, a new immunosuppressive drug with great potential in internal medicine. AB - Mycophenolate mofetil is a new immunosuppressive drug, exhibiting its effect through inhibition of proliferation of T- and B-lymphocytes. Superior efficacy of mycophenolate mofetil compared to azathioprine, in combination with cyclosporine and prednisone, in the prevention of acute rejection in organ transplantation has made mycophenolate mofetil one of the standard immunosuppressive drugs after transplantation. Mycophenolate mofetil also is an interesting candidate drug for many other, mainly auto-immune mediated diseases. The use of mycophenolate mofetil in several of these diseases is discussed. The definitive place of mycophenolate mofetil will depend on the results of randomised trials currently under way. PMID- 11099794 TI - Digestive lipases: from three-dimensional structure to physiology. AB - Human gastric lipase (HGL) is a lipolytic enzyme that is secreted by the chief cells located in the fundic part of the stomach. HGL plays an important role in lipid digestion, since it promotes the subsequent hydrolytic action of pancreatic lipase in duodenal lumen. Physiological studies have shown that HGL is able of acting not only in the highly acid stomach environment but also in the duodenum in synergy with human pancreatic lipase (HPL). Recombinant HGL (r-HGL) was expressed in the baculovirus/insect cell system in the form of an active protein with a molecular mass of 45 kDa. The specific activities of r-HGL were found to be similar to that of the native enzyme when tested on various triacylglycerol (TG) substrates. The 3-D structure of r-HGL was the first solved within the mammalian acid lipase family. This globular enzyme (379 residues) shows a new feature, different from the other known lipases structures, which consists of a core domain having the alpha/beta hydrolase fold and a cap domain including a putative 'lid' of 30 residues covering the active site of the lipase (closed conformation). HPL is the major lipolytic enzyme involved in the digestion of dietary TG. HPL is a 50 kDa glycoprotein which is directly secreted as an active enzyme. HPL was the first mammalian lipase to be solved structurally, and it revealed the presence of two structural domains: a large N-terminal domain (residues 1-336) and a smaller C-terminal domain (residues 337-449). The large N terminal domain belongs to the alpha/beta hydrolase fold and contains the active site. A surface loop called the lid domain (C237-C261) covers the active site in the closed conformation of the lipase. The 3-D structure of the lipase procolipase complex illustrates how the procolipase might anchor the lipase at the interface in the presence of bile salts: procolipase binds to the C-terminal domain of HPL and exposes the hydrophobic tips of its fingers at the opposite site of its lipase-binding domain. These hydrophobic tips help to bring N terminal domain into close conformation with the interface where the opening of the lid domain probably occurs. As a result of all these conformational changes, the open lid and the extremities of the procolipase form an impressive continuous hydrophobic plateau, extending over more than 50 A. This surface might able to interact strongly with a lipid-water interface. The biochemical, histochemical and clinical studies as well as the 3-D structures obtained will be a great help for a better understanding of the structure-function relationships of digestive lipases. PMID- 11099795 TI - Kinetic behavior of the pancreatic lipase-colipase-lipid system. AB - Pancreatic lipase is a surface-active protein that binds avidly to interfaces comprised of the substrates and products of lipolysis. However, both lipase binding to substrate-containing particles and subsequent interfacial catalysis are inhibited by a number of amphipathic molecules. The most thoroughly studied of these, phosphatidylcholine, is a common constituent of membranes and intestinal lipid contents. Colipase, a surface-active cofactor of lipase, relieves inhibition by phosphatidylcholine in several ways. Through protein protein interactions, colipase helps anchor lipase to surfaces and stabilizes it in the open conformation. Within the interface, colipase packs more efficiently with substrates and products of lipolysis than with phosphatidylcholine, thereby concentrating these reactants in the vicinity of colipase. This enrichment of lipase substrates and products in the vicinity of colipase enhances lipase-lipid interactions. The result is that colipase facilitates the adsorption of lipase to the interface and, possibly, increases the availability of substrate to the enzyme. Thus, the functional unit in intestinal lipolysis appears to be a lipase colipase-reactant complex. PMID- 11099796 TI - Properties and function of pancreatic lipase related protein 2. AB - The lipase gene family includes pancreatic triglyceride lipase and two pancreatic proteins, pancreatic lipase related proteins 1 and 2, with strong nucleotide and amino acid sequence homology to pancreatic triglyceride lipase. All three proteins have virtually identical three-dimensional structures. Of the pancreatic triglyceride lipase homologues, only pancreatic lipase related protein 2 has lipase activity. Like pancreatic triglyceride lipase, related protein 2 cleaves triglycerides, but it has broader substrate specificity. Pancreatic lipase related protein 2 also hydrolyzes phospholipids and galactolipids, two fats that are not substrates for pancreatic triglyceride lipase. The rat-related protein 2 also differs from pancreatic triglyceride lipase in sensitivity to bile salts and in response to colipase. Although the pancreas expresses both lipases, their temporal pattern of expression differs. Pancreatic lipase-related protein 2 mRNA appears before birth and persists into adulthood, whereas PTL mRNA first appears at the suckling-weanling transition. Additionally, intestinal enterocytes, paneth cells and cultured cytotoxic T-cells express mRNA encoding pancreatic lipase related protein 2. A physiological function for pancreatic lipase related protein 2 was demonstrated in mice that did not express this protein. Pancreatic lipase related protein 2 deficient mice malabsorbed fat in the suckling period, but not after weaning. They also had a defect in T-cell mediated cytotoxicity. Thus, pancreatic lipase related protein 2 is a lipase that participates in the cytotoxic activity of T-cells and plays a critical role in the digestion of breast milk fats. PMID- 11099797 TI - Staphylococcal lipases: biochemical and molecular characterization. AB - To date, the nucleotide sequences of nine different lipase genes from six Staphylococcus species, three from S. epidermidis, two from S. aureus, and one each from S. haemolyticus, S. hyicus, S. warneri, and S. xylosus, have been determined. All deduced lipase proteins are similarly organized as pre-pro proteins, with pre-regions corresponding to a signal peptide of 35 to 38 amino acids, a pro-peptide of 207 to 321 amino acids with an overall hydrophilic character, and a mature peptide comprising 383 to 396 amino acids. The lipases are secreted in the pro-form and are afterwards processed to the mature form by specific proteases. The pro-peptide of the S. hyicus lipase is necessary for efficient translocation and for protection against proteolytic degradation. Despite being very similar in their primary structures the staphylococcal lipases show significant differences in their biochemical and catalytic properties, such as substrate selectivity, pH optimum and interfacial activation. The lipase from S. hyicus is unique among the staphylococcal and bacterial lipases in that it has not only lipase activity, but also a high phospho-lipase activity. All staphylococcal lipases are dependent on Ca(2+), which is thought to have a function in stabilizing the tertiary structure of the lipases. Evidence exists that staphylococcal lipases like other bacterial lipases, possess a lid-like domain that might be involved in the interfacial activation of these enzymes. PMID- 11099798 TI - Fusarium solani pisi cutinase. AB - Cutinase from Fusarium solani pisi has been studied extensively with respect to its structural and functional properties. The crystal structure of the enzyme was solved to high atomic resolution (1 angstrom), while data on structural dynamics have been obtained from detailed NMR studies. Functional data were mainly derived from kinetic studies using substrate analogues that simplify the kinetic behaviour. The properties of wild-type cutinase are reviewed and discussed in relation with the effects brought about by site-directed variants of the enzyme. PMID- 11099799 TI - Bacterial lipases from Pseudomonas: regulation of gene expression and mechanisms of secretion. AB - Lipases from Pseudomonas bacteria are widely used for a variety of biotechnological applications. Overexpression in heterologous hosts like Escherichia coli failed to produce enzymatically active lipase prompting to study the molecular mechanisms underlying the regulation of lipase gene expression and secretion. The prototype lipase from P. aeruginosa is encoded in a bicistronic operon which is transcribed from two different promotors, one of which depends on the alternative sigma factor RpoN (sigma(54)). Recently, a two-component regulatory system was identified as an element controlling transcription of the lipase operon. P. aeruginosa lipase is secreted via a type II pathway. The cytoplasmic prelipase contains a 26 amino acid N-terminal signal sequence mediating secretion across the inner membrane via the Sec-machinery. In the periplasm, lipase folds into an enzymatically active conformation assisted by its specific intermolecular chaperone Lif and by unspecific accessory folding catalysts including Dsb-proteins which catalyze the formation of a disulfide bond. Enzymatically active and secretion-competent lipase is finally transported through a complex secretion machinery consisting of 12 different Xcp-proteins of which XcpQ forms a pore-like structure in the outer membrane allowing the release of lipase into the extracellular medium. Biotechnologically important lipases from Burkholderia glumae and P. alcaligenes also use such a type II secretion pathway whereas lipases from P. fluorescens and Serratia marcescens, which lack a typical signal sequence are secreted via a type I pathway. Future challenges to produce Pseudomonas lipases may include artificial up-regulation of lipase gene transcription and construction of more efficient expression strains in which both folding and secretion of lipase are optimized. PMID- 11099800 TI - What distinguishes an esterase from a lipase: a novel structural approach. AB - Esterases and lipases both hydrolyse ester bonds. Whereas the lipases display high activity towards the aggregated state of its substrate, the esterases typically show highest activity towards the soluble state of its substrate. We have compared the amino acid sequence, the 3D-structure as well as the pH dependent electrostatic signature of selected members of the two families, for which 3D-structural information is publicly available. Lipases display a statistically significant enhanced occurrence of non-polar residues close to the surface, clustering around the active-site. Lid opening appears to strengthen this pattern further. As we have proposed earlier the active site of lipases displays negative potential in the pH-range associated with their maximum activity, typically at pH values above 8. The esterases show a very similar pattern, however, at pH values around 6 correlated with their usually lower pH activity optimum. PMID- 11099801 TI - The His gap motif in microbial lipases: a determinant of stereoselectivity toward triacylglycerols and analogs. AB - Lipases preferably hydrolyze the sn-1 and sn-3 acyl chain of triacylglycerols and sn-2 substituted analogs. Molecular modeling studies of the stereopreference of microbial lipases from Rhizopus oryzae, Rhizomucor miehei, Candida rugosa, and lipase B from Candida antarctica toward the hydrolysis of triacylglycerols and analogs revealed that sterical interactions occurring between the sn-2 substituent and the His gap affect substrate geometry, which can be monitored by a single torsion angle. This torsion angle correlates to the experimentally determined stereopreference and is, therefore, suitable to predict stereopreference by molecular modeling. For a given microbial lipase, stereopreference can be estimated by measuring the distance between the side chains of the His gap residues: a narrow His gap cleft implies sn-3 stereopreference for all investigated substrates; a medium-sized His gap discriminates by flexibility of the substrates: flexible substrates are hydrolyzed in sn-1, while rigid substrates are hydrolyzed in sn-3. A wide open His gap implies sn-1 stereopreference for all substrates. This rule holds for all investigated microbial wild type lipases and mutants. PMID- 11099802 TI - Effect of the lipid interface on the catalytic activity and spectroscopic properties of a fungal lipase. AB - Lipase from the fungi Thermomyces (formerly Humicola) lanuginosa (TlL) is widely used in industry. This interfacial enzyme is inactive under aqueous conditions, but catalytic activation is induced on binding to a lipid-water interface. In order for protein engineering to design more efficient mutants of TlL for specific applications, it is important to characterize its interfacial catalysis. A complete analysis of steady-state kinetics for the hydrolysis of a soluble substrate by TlL has been developed using an interface different from the substrate. Small vesicles of 1-palmitoyl-2-oleoylglycero-sn-3-phosphoglycerol (POPG) or other anionic phospholipids are a neutral diluent interface for the partitioning of substrate and enzyme. TlL binds to these interfaces in an active or open form, thus implying a displacement of the helical lid away from the active site. A study of the influence of substrate and diluent concentration dependence of the rate of hydrolysis provides a basis for the determination of the primary interfacial catalytic parameters. The interfacial activation is not supported by zwitterionic vesicles or by large anionic vesicles of 100 nm diameter, although TlL binds to these interfaces. Using a combination of fluorescence-based techniques applied to several mutants of TlL with different tryptophan residues we have shown that TlL binds to phospholipid vesicles in different forms rendering different catalytic activities, and that the open lid conformation is achieved and stabilized by a combination of electrostatic and hydrophobic interactions between the enzyme's lipid-binding face and the interface. PMID- 11099803 TI - Reverse micelles as reaction media for lipases. AB - Reversed micelles are at the present time faced as common organic media to perform biocatalysis. They have been associated to the idea of a microreactor where the enzyme can be sheltered and protected from solvent detrimental effects. This simplistic idea led some investigators to ignore some basic understanding, such as the recognition of the enzyme-specific microenvironment and what the enzyme experiences inside the reversed micelle. To date the number of reactions catalyzed by lipases in reversed micelles is large. This review aims to highlight some of the fundamental aspects of the lipase microencapsulation as well as to resume the outstanding progress of the reversed micellar systems. The properties of the micellar microenvironment are reviewed and related to the lipases' performance both in terms of activity and stability. The heterogeneity of reversed micellar systems is discussed in relation to component distribution models and also to enzymatic kinetics. The new trends and the practical aspects where efforts should be centralized in order to spread out the micellar bioreactor technology over industrial processes are also discussed. PMID- 11099804 TI - Change of editorship PMID- 11099805 TI - Frontal electrocortical and cardiovascular reactivity during happiness and anger. AB - The present study investigated electrocortical and cardiovascular reactivity during positive and negative emotion, and examined the relation of asymmetric frontal lobe activation to cardiovascular responses. Participants were 30 healthy, right-handed university students (mean age, 23.9; 60% female; 76% Caucasian). Electroencephalographic (EEG), blood pressure (BP), and heart rate (HR) responses were assessed while subjects engaged in laboratory tasks (personally-relevant recall tasks and film clips) designed to elicit happiness or anger. Happiness-inducing tasks evoked more prominent left than right frontal EEG activation, and greater left frontal EEG activation than anger-inducing tasks. However, anger-inducing tasks were, on average, associated with comparable left and right frontal EEG activation. Irrespective of emotional valence, cardiovascular activation was more pronounced during personally-relevant recall tasks than during the viewing of film clips. During anger recall, both greater left frontal EEG response (r=-0.46, P<0.02) and greater right frontal EEG response (r=-0.45, P<0.02) were correlated significantly with increased HR reactivity during the task. In addition, a right lateralized frontal EEG response during anger-inducing tasks was associated with greater concomitant systolic BP (P<0.03) and diastolic BP (P<0.008) reactivity. Exploratory analyses also indicated that men who displayed a left lateralized frontal EEG response during happiness-inducing tasks showed the greatest concomitant systolic BP and HR reactivity (P's<0.03). These findings suggest that asymmetric frontal EEG responses to emotional arousal may elicit different patterns of cardiovascular reactivity in healthy adults. PMID- 11099806 TI - The gap effect in pro-saccades and anti-saccades in psychometric schizotypes. AB - Schizophrenic patients and their first-degree relatives exhibit deficits in the anti-saccade task. In the present study, anti-saccade task performance was examined in subjects with 'high' and 'low' expressions of the schizotypal personality trait. For that purpose, the SPQ-G, the German adaptation of the Schizotypal Personality Questionnaire (SPQ; Raine, 1991), was filled in by 489 university students. Twenty and 21 participants with 'high' and 'low' SPQ-G scores, respectively, were compared with respect to saccadic eye movements elicited under the overlap and 200 ms gap conditions of the pro- and anti-saccade tasks. Each task block comprised 150 trials, 75 to either side in random order. The order of presentation of the task blocks was counterbalanced across the participants of each group. Saccadic reaction times were slower during the anti- as compared to the pro-saccade task and under the overlap as compared to the gap condition. Direction errors occurred almost exclusively during the anti-saccade task, express saccades mainly under the pro-saccadic gap condition. High schizotypal participants did not differ significantly from low-schizotypal participants in any of these measures. While these results might suggest normal anti-saccade task performance in schizotypal personality as defined by the SPQ-G, the sampling strategy adopted in the present study is the more plausible explanation for the lack of group differences. PMID- 11099807 TI - Cortical plasticity, contingent negative variation, and transcendent experiences during practice of the Transcendental Meditation technique. AB - This study investigated effects of transcendent experiences on contingent negative variation (CNV) amplitude, CNV rebound, and distraction effects. Three groups of age-matched subjects with few (<1 per year), more frequent (10-20 per year), or daily self-reported transcendent experiences received 31 simple RT trials (flash (S(1))/tone (S(2))/button press) followed by 31 divided-attention trials - randomly intermixed trials with or without a three-letter memory task in the S(1)-S(2) interval). Late CNV amplitudes in the simple trials were smallest in the group with fewest, and largest in the group with most frequent transcendent experiences. Conversely, CNV distraction effects were largest in the group with fewest and smallest in the group with most frequent transcendent experiences (the second group's values were in the middle in each case). These data suggest cumulative effects of transcendent experiences on cortical preparatory response (heightened late CNV amplitude in simple trials) and executive functioning (diminished distraction effects in letter trials). PMID- 11099808 TI - Further investigations of stimulus coding in nonlinear discrimination problems. AB - Two human Pavlovian SCR conditioning experiments are reported, investigating CS coding in negative patterning (NP). In Experiment 1, NP was run with two pairs of letter stimuli (C, N, and M, J), reinforced by shock when alone but not in compound. Controls with the same paired-unpaired sequence saw two additional nonreinforced letters (X and H) instead of compounds. The NP group learned the element-compound differentiation, but the controls did not discriminate reinforced from nonreinforced letters. The availability of an abstract rule (such as stimulus number) for distinguishing between reinforced and nonreinforced CSs led to discrimination in NP, but its unavailability resulted in no discrimination in the controls. In Experiment 2, NP was run with one pair of letters (reinforced as elements but not in compound). A control group had the same paired-unpaired sequence, but their compound contained two different letters from the reinforced ones. The NP discrimination was learned, but the controls failed to differentiate the reinforced elements from the nonreinforced compound. It was concluded that the NP discrimination was not based on the number of reinforced and nonreinforced stimuli, because this dimension was available to both groups in Experiment 2. The abstract dimension of separate versus together, on the other hand, was available only in NP, suggesting that it is the SINE QUA NON for the acquisition of the kind of NP discriminations we have been studying. PMID- 11099820 TI - Peroxynitrite-induced modification of the N-methyl-D-aspartate receptor in the cerebral cortex of the guinea pig fetus at term. AB - The present study tests the hypothesis that nitration is a potential mechanism of N-methyl-D-aspartate (NMDA) receptor modification, by assessing the effect of peroxynitrite in vitro on the glutamate and ion-channel sites of the NMDA receptor in the fetal guinea pig. Nitration of NMDA receptor subunits was confirmed by Western blot. Following peroxynitrite exposure, (3)H-MK-801 bindings show an increase in the B(max) and a decrease in the K(d), while (3)H-glutamate bindings show a decrease in the K(d) with no change in the B(max). We conclude that peroxynitrite regulates the NMDA receptor function by increasing the affinity of the ion-channel and glutamate sites, and by exposing additional ion channel sites. We propose that nitration of the NMDA receptor is a potential mechanism for the regulation of the receptor during hypoxia. PMID- 11099819 TI - Population code for tracking velocity based on cerebellar Purkinje cell simple spike firing in monkeys. AB - Velocity is an important determinant of the simple spike discharge of cerebellar Purkinje cells. In a previous study, Purkinje cells in the intermediate and lateral cerebellum recorded during manual tracking were found to be tuned to a combination of direction and speed, (i.e. preferred velocity). In this study a population analysis of this simple spike discharge was used to determine whether the velocity of tracking could be predicted. For the majority (30/32) of direction-speed combinations, the population response accurately specified the target velocity. A temporal analysis showed how the population response gradually converged to the required velocity 200 ms prior to the onset of tracking. Therefore, the simple spike discharge of a Purkinje cell ensemble contains sufficient information to reconstruct target velocity, providing support for the hypothesis that the cerebellum controls or signals movement velocity. PMID- 11099821 TI - The reversal potential of Ca(2+)-activated Cl(-) currents indicates that chick sensory neurons accumulate intracellular Cl(-). AB - In order to establish the physiological role of the Ca(2+)-activated Cl(-) current (I(Cl(Ca))) of chick primary afferent neurons, I measured the reversal potential of this current using either the amphotericin perforated patch technique (that alters intracellular Cl(-)) or the gramicidin perforated patch technique (that does not perturb intracellular Cl(-)). In the amphotericin experiments at 35 degrees C, I(Cl(Ca)) reversed at the Cl(-) equilibrium potential (E(Cl)=-24 mV) set by the superfusate (147 mM Cl(-)) and the pipette solution (60 mM Cl(-)). In contrast, in the gramicidin experiments at 35 degrees C, I(Cl(Ca)) reversed at -42+/-2 mV, midway between E(Cl) of the solutions and E(Cl) expected if Cl(-) were passively distributed. Thus the gramicidin perforated patch technique monitors Cl(-) currents without perturbing intracellular Cl(-). Further, the data imply that chick dorsal root ganglia (DRG) neurons actively accumulate Cl(-). I(Cl(Ca)) reversed at the same potential ( 46+/-3 mV) at 20 degrees C indicating that the non-equilibrium distribution of Cl(-) is maintained at the lower temperature. Thus, I(Cl(Ca)) is a depolarizing current that can contribute to the after-depolarization in chick DRG neurons and thereby alter Ca(2+) influx. PMID- 11099822 TI - Weightlessness during spaceflight results in enhanced synapse formation in a fish brain vestibular nucleus. AB - Synapse counts were undertaken by conventional electron microscopy in primary vestibular integration centers, (i.e. nucleus descendens and nucleus magnocellularis of the brainstem area octavolateralis) and in the diencephalic visual nucleus corticalis of spaceflown neonate swordtail fish Xiphophorus helleri as well as in 1 g control siblings. Spaceflight (16 days microgravity, (microg), STS-90 Neurolab Mission) yielded an increase in synaptic contacts within the vestibular nucleus descendens indicating that lack of input resulted in compensation processes. No effect of microg, however, was observed in the visual nucleus corticalis and in the vestibular nucleus magnocellularis which is situated in the close vicinity of the nucleus descendens. In contrast to the latter, the nucleus magnocellularis does not receive exclusively vestibular input, but inputs from the lateral line as well, possibly providing sufficient input at microgravity. PMID- 11099823 TI - Evidence against association of the FE65 gene (APBB1) intron 13 polymorphism in Alzheimer's patients. AB - A genetic polymorphism in intron 13 of the FE65 gene (APBB1) was reported to be associated with Alzheimer's disease (AD). Our analyses of this polymorphism, both in a family-based or a case-control sample, fail to support the association between the FE65 intron 13 polymorphism and AD. We performed the sibship disequilibrium test (SDT, P=0.77) and the sib transmission/disequilibrium test (Sib-TDT, P=0.56) in a family-based study which included 526 subjects from 158 sibships. In addition, we compared the genotype and allele frequencies of this biallelic polymorphism in 311 AD patients to those of a control group consisting of 260 subjects and found no significant difference (chi(2), P=0.847 and P=0.586, respectively). Furthermore, our two-point linkage analysis in a family-based sample was in agreement with a genome wide scan for linkage to AD and showed no evidence for linkage to the short arm of chromosome 11 where the FE65 gene is located. We conclude that the association of the FE65 intron 13 polymorphism with AD, if any, is smaller than previously reported. PMID- 11099824 TI - Subthreshold 5-Hz repetitive transcranial magnetic stimulation of the human primary motor cortex reduces intracortical paired-pulse inhibition. AB - Paired-pulse transcranial magnetic stimulation (TMS) at short interstimulus intervals was employed to investigate short-term effects of 5-Hz repetitive TMS (rTMS) over the primary motor hand area (M1(HAND)) on intracortical excitability. In ten healthy individuals, 1250 pulses of 5-Hz rTMS were applied at 90% of motor resting threshold over the left M1(HAND). Ten minutes after 5-Hz rTMS, paired pulse inhibition was significantly reduced, whereas paired-pulse facilitation was not modified. Sham-rTMS had no lasting effect on intracortical excitability. These findings suggest that subthreshold 5-Hz rTMS causes a short-term modulation of the excitability of intracortical circuitry in the stimulated M1(HAND). The lasting effect of subthreshold 5-Hz rTMS on intracortical inhibition provides a useful probe for studying short-term plasticity of the human M1(HAND). PMID- 11099825 TI - Antinociceptive effect of a group II metabotropic glutamate receptor antagonist in the thalamus of monoarthritic rats. AB - Adult rats were rendered monoarthritic (MA) by injection of 50 microl of complete Freund's adjuvant (CFA) into the tibiotarsal joint. The ankle-bend (AB) test of nociception was performed in those animals before and during 60 min after the stereotaxic injection of 2 microl of either saline (controls) or (2S)-alpha ethylglutamic acid (EGLU, 80 nmol in 2 microl), a group II metabotropic glutamate receptors (mGluR) antagonist, in the reticular thalamic nucleus (Rt) contralateral to the arthritic joint. AB scores reached near maximum values before the stereotaxic injections (18.7+/-0.8), and remained constant throughout the entire experimental period in the control group, denoting marked allodynia. In the EGLU-treated group, AB scores gradually decreased after EGLU injection, with minimum values at 10 min (7.7+/-1.6), recovering to scores near maximum at 60 min (19.7+/-0.3). The data point to an activation of group II mGluR by noxious inputs in the Rt of MA rats, suggesting their participation in inhibiting local gamma-aminobutyric acid (GABA)ergic inhibitory neurones. PMID- 11099826 TI - Tryptamine derivatives as non-competitive N-methyl-D-aspartate receptor blockers: studies using [(3)H]MK-801 binding in rat hippocampal membranes. AB - Derivatives of phenylethylamine and tryptamine share structural features with the non-competitive N-methyl-D-aspartate (NMDA) receptor blockers phencyclidine, ketamine, and MK-801. Tryptamine and phenylethylamine inhibited the specific binding of [(3)H]MK-801 to rat hippocampal membranes with IC(50)'s 190 and 905 microM, respectively. The corresponding amino acids phenylalanine and tryptophan were inactive, their methyl esters, however, were slightly more potent than the amines. The methyl ester of the naturally occurring L-tryptophan was 12 times more potent than the methyl ester of the D-isomer, whereas the corresponding isomers derived from phenylalanine exhibited no stereoselectivity. The potency of tryptamine was increased by substitutions as, e.g. in 5-methyltryptamine (IC(50) 12 microM) and tryptophan octylester (IC(50) 5.2 microM). Compounds formed in vivo from L-tryptophan and a lipophilic counterpart may function as endogenous non-competitive NMDA receptor blockers. PMID- 11099827 TI - Naming of animals and tools: a functional magnetic resonance imaging study of categorical differences in the human brain areas commonly used for naming visually presented objects. AB - To investigate the neural substrates for naming objects and their category dependency, we performed functional magnetic resonance imaging (fMRI) with naming of animals and tools. Naming objects, irrespective of their category, activated left frontal to bilateral parietal regions and occipital to posterior temporal regions. Within these areas, naming animals caused more activation of the primary visual cortex bilaterally and the ventral occipital cortex to the inferior temporal area on the right side. Naming tools caused more activation of the posterior part of the left middle temporal area, the rostral part of the left inferior parietal lobule, and the left inferior frontal cortex. These findings suggest that the neural network for naming objects has discrete category dependent nodes through which pertinent conceptual knowledge may be mediated. PMID- 11099828 TI - Superoxide radical scavenging and attenuation of hypoxia-reoxygenation injury by neurotransmitter ferric complexes in isolated rat hepatocytes. AB - Reactive oxygen species have been implicated in the pathogenesis of hypoxia reoxygenation injury. Previously, we demonstrated that 2:1 catecholic iron complexes were more effective than uncomplexed catechols at (a) scavenging superoxide radicals generated enzymatically, and (b) protecting hepatocytes against hypoxia-reoxygenation injury [25]. Based on these findings, we sought to demonstrate similar effects using catecholamine neurotransmitters. Various catecholamine-iron complexes were shown to be more effective than uncomplexed catecholamines at scavenging superoxide radicals and could be used to protect cells from hypoxia-reoxygenation injury. alpha-Methyl-3, 4-dihydroxyphenylalanine (alpha-methylDOPA) complexed with ferric ion (2:1) showed the greatest superoxide scavenging potency amongst the catecholamine-iron complexes. The uncomplexed catecholamines were much less effective at scavenging superoxide radicals than the iron-catecholamine complexes. Dopamine was the most effective superoxide scavenger among the uncomplexed catecholamines. The superoxide scavenging effectiveness of the latter seemed to correlate with their reduction potentials, but not directly to their pK(a) values. Furthermore, dopamine:iron(III) complex protected isolated hepatocytes against hypoxia-reoxygenation injury at concentrations four-fold lower than that required for protection by dopamine alone. PMID- 11099829 TI - Formation of hydroxy radicals by environmental estrogen-like chemicals in rat striatum. AB - We investigated effects of environmental estrogen-like chemicals, para nonylphenol and bisphenol A, on hydroxy radical formation in the striatum of adult rats, using an in vivo microdialysis system. Para-nonylphenol significantly stimulated hydroxy radical formation in the striatum. Bisphenol A also increased hydroxy radical formation, albeit effect being slight. The formation of hydroxy radicals induced by para-nonylphenol was dose-dependently inhibited by tamoxifen, which suggests that the effect of this chemical was an estrogenic action via estrogen receptors. The results of the present study are the first demonstration on hydroxy radical formation induced by environmental estrogen-like chemicals and suggest that the in vivo microdialysis may be useful for evaluating toxic effects of environmental chemicals on nervous tissues. PMID- 11099830 TI - Changes in electromyogram during upper limb muscle contraction induced by resistive loaded breathing in humans. AB - Expiratory only resistive loaded breathing (RL) reduces high energy electromyogram (EMG) power (EH) during an isometric contraction of a leg extensor muscle, but not an arm flexor. An interaction between afferent activity during expiratory RL and inspiratory non-loaded phases of breathing, which the contraction spanned, could have accounted for the reduced EH in these long contractions. Therefore this study tested the hypothesis that brief arm extensor muscle contractions (70% of maximal force), performed during a single phase of expiratory RL, would also exhibit reduced EH. Surprisingly, EH in triceps, but not biceps brachii was reduced significantly when the contraction was performed during inspiratory RL rather than expiratory RL. The results suggest that either (a) short and prolonged contractions or (b) motor drive to arm and leg extensors are affected differently by RL. PMID- 11099831 TI - Testosterone stimulates rapid secretory amyloid precursor protein release from rat hypothalamic cells via the activation of the mitogen-activated protein kinase pathway. AB - The processing of the amyloid precursor protein (APP) has become a major focus of research into Alzheimer's disease (AD). Recently, repeated doses of testosterone have been shown to enhance the secretion of the product of the alpha-cleavage pathway of APP (sAPPalpha) over a period of days. Here, the time course of secretion of sAPPalpha after a single physiological dose of testosterone using an immortalized rat hypothalamic cell line (GT1-7) and the signalling pathways involved was analyzed. Testosterone was found to increase the amount of APP secretion rapidly after treatment without effecting the overall amount of cellular APP. The species of APP secreted was found to be predominantly the product of the non-amyloidogenic alpha-secretory pathway. Further, this event is regulated via aromatase-mediated conversion of testosterone to estrogen and the mitogen-activated protein kinase (MAP kinase) signalling pathway. Taken together these data partially elucidates the cellular cascade by which testosterone stimulates sAPP secretion. PMID- 11099832 TI - Increased protein levels of serotonin transporter in frontal cortex of patients with Down syndrome. AB - Serotonin transporters (SERTs) are presynaptic proteins specialized for the clearance of serotonin from the synaptic cleft. A large body of evidence exists on altered platelet serotonin uptake and metabolism in Down syndrome (DS). Besides, dysregulation of SERTs expression have been reported in various complex behavioural traits and disorders including, neurodegenerative disorders. This prompted us to investigate SERT protein levels in adult brain specimens. Western blot analyses were performed in frontal cortex and cerebellum of aged controls and patients with DS and Alzheimer's disease (AD). The result revealed that frontal cortex SERT was significantly increased (P<0.05) in DS, whereas in AD it was comparable to controls. In cerebellum, no significant difference was observed among the study populations. A remarkable difference was noted when SERT was normalized vs. neuron specific enolase (NSE), a neuronal marker. The increase in SERT/NSE was highly significant (P<0.01) in DS frontal cortex compared to controls. Neither AD frontal cortex nor DS and AD cerebellum did show any significant difference. These findings indicate that a region specific alteration in SERT expression may exist in DS with AD-like pathology. As little is known about the status of serotenergic synaptic markers in DS brain, the findings may contribute to an effort made to delineate the underlying causes of serotonergic dysfunction in DS and the quest for therapeutic strategies. The study also suggest caution should be taken in applying data obtained from DS to model AD biochemical defects. PMID- 11099833 TI - Evidence for existence of immobilization stress-inducible semicarbazide-sensitive amine oxidase inhibitor in rat brain cytosol. AB - An endogenous inhibitor of semicarbazide-sensitive amine oxidase (SSAO) was separated by gel filtration from 105000xg supernate in rat brain cytosol following immobilization stress (IMMO). The molecular weight of this inhibitor was estimated to be 500-700 by gel filtration. This inhibitor was proved to be heat-stable resistant to protease treatment. These results suggest that this inhibitor is induced by IMMO. SSAO activity in rat brain might be regulated by the level of this inhibitor. PMID- 11099834 TI - Diminution of training-induced transient motor cortex plasticity by weak transcranial direct current stimulation in the human. AB - Training of a thumb movement in the opposite direction of a twitch in response to transcranial magnetic stimulation (TMS) induces a transient directional change of post-training TMS-evoked movements towards the trained direction. Functional synaptic mechanisms seem to underlie this rapid training-induced plasticity. Transcranial direct current stimulation (tDCS) induces outlasting changes of cerebral excitability, thus presenting as promising tool for neuroplasticity research. We studied the influence of tDCS, applied over the motorcortex during training, on angular deviation of post-training to pre-training TMS-evoked thumb movements. With tDCS of anodal and cathodal polarity the training-induced directional change of thumb movements was significantly reduced during a 10 min post-training interval, indicating an interference of tDCS with mechanisms of rapid training-induced plasticity. PMID- 11099835 TI - Preface PMID- 11099836 TI - Historical perspectives and current global challenges of Trichinella and trichinellosis. AB - Trichinella spiralis and related species of Trichinella have had a long history of causing human disease, and as a foodborne pathogen have had a major impact on international commerce of pork and other meat animal species which are known to transmit the parasite. Our knowledge of Trichinella has increased substantially over the past few years particularly in the areas of phylogeny, host diversity, epidemiology and control. In this paper, we provide a brief overview of our understanding of Trichinella from its discovery to present time. Past and current challenges of the control of Trichinella and trichinellosis are summarized. As editors of this special issue of Veterinary Parasitology, we introduce a series of invited review articles prepared by experts from around the world, summarizing recent knowledge in Trichinella and trichinellosis. PMID- 11099837 TI - Trichinellosis: a worldwide zoonosis. AB - Trichinella spp. are some of the most widespread parasites infecting people and other mammals all over the world, regardless of climate. This paper attempts to describe the present status of trichinellosis worldwide and to determine if and why trichinellosis is emerging or re-emerging. The global prevalence of the disease is difficult to evaluate but as many as 11 million people may be infected. More than 10000 cases of human trichinellosis were reported by the International Commission on Trichinellosis from 1995 to June 1997 and about 10000 porcine infections were reported by the Office International des Epizooties in 1998. The disease is particularly worrisome in the Balkans, Russia, the Baltic republics, in some parts of China and Argentina. Horsemeat-related outbreaks have been reported in France and Italy and have involved about 3000 patients in the past 25 years. The emergence of trichinellosis in some countries is explained by a better knowledge of the disease (formerly often misdiagnosed as influenza), modifications of consumer habits, re-forestation in Europe and increase of wild game, importation of meats from countries where trichinellosis is endemic and failure of veterinary control due to human error or to social upheavals. This disease linked to meat-consumption which is theoretically easy to prevent by adequate cooking, freezing and veterinary controls, should deserve the attention of all persons involved in public health and it could be eradicated at least from domestic pigs. PMID- 11099838 TI - Epidemiology of trichinellosis in Mexico, Central and South America. AB - Trichinella species are widely distributed throughout the world and are found in a large number of carnivorous animals, humans and incidental hosts. The data presented in this review show that Trichinella infection has been reported in both humans and animals in Mexico, Argentina and Chile since the end of the 19th century, and more recently in Bolivia. This parasitic infection is still a public health problem in countries such as Argentina and Chile. Although efforts have focused on the control and prevention of trichinellosis in these countries, there were still human cases and outbreaks of trichinellosis reported. Diagnosis of infection in animals such as pigs still includes, in many endemic areas, the use of trichinoscopy. In Argentina, however, artificial digestion has been recently introduced in slaughterhouses to detect Trichinella infection in pigs. In some endemic areas in Mexico, the use of serological assays for human trichinellosis and pig infections have resulted in improved detection. Most of the outbreaks of human trichinellosis in Mexico, Argentina and Chile have occurred as a result of the consumption of undercooked pork or pork products from animals raised under poor hygienic conditions and which are clandestinely slaughtered. In several studies, rats, dogs and cats have been found to be infected with Trichinella and may be considered a risk for transmission of the infection to pigs or other animals intended for human consumption. Another potential source of transmission of Trichinella to humans is horsemeat; however, information related to horse trichinellosis in Latin-American countries is scarce. In most cases the etiological agent of human trichinellosis in Central and South America has been reported to be Trichinella spiralis; however, only in a few cases has the parasite species been properly identified. Based on the reports available, it is clear that there is a need to carry out better controlled epidemiological studies to determine the true prevalence of the infection in this region of the world. Also, more sensitive methods of diagnosis and improvements in conditions for pig production as well as better sanitary inspection systems, are needed for controlling and preventing transmission of trichinellosis in these countries. PMID- 11099839 TI - Epidemiology of trichinellosis in Asia and the Pacific Rim. AB - The epidemiology of trichinellosis, species of Trichinella present and the food and eating habits of people affected in Asia and the Pacific Rim are reviewed with emphasis on Japan, China and Thailand. Trichinella seems to be prevalent throughout this region although outbreaks of trichinellosis have not been reported in some areas. Major outbreaks of the disease have been reported primarily in China and Thailand. This is the result of three factors: (1) China and Thailand are highly endemic areas for this parasite; (2) the two countries are well-organized and there is a public health system that enables precise reporting of disease outbreaks and (3) culinary habits provide many opportunities to eat undercooked meats. Trichinella found in Asia and the Pacific Rim includes both encapsulated species (Trichinella spiralis, Trichinella britovi, Trichinella nativa) and noncapsulated species (Trichinella pseudospiralis, Trichinella papuae). T. britovi, isolated in Japan, is a different genotype from the European strain. Therefore, the Japanese strain of T. britovi is designated Trichinella T9. Human trichinellosis caused by T. pseudospiralis has occurred in New Zealand and Thailand. Tasmania has had animal cases of T. pseudospiralis infection and animals with T. papuae infection have been found in Papua New Guinea. Economic losses due to Trichinella infection are not negligible in China, where there have been more than 500 outbreaks of human trichinellosis, affecting more than 20,000 people and causing more than 200 deaths. In Thailand, over the past 27 years, 120 outbreaks were reported involving nearly 6700 patients and 97 deaths. Japan has had fewer outbreaks and some sporadic cases have been attributed to imported infection. PMID- 11099840 TI - Factors affecting the flow among domestic, synanthropic and sylvatic cycles of Trichinella. AB - Nematodes of the genus Trichinella are maintained in nature by sylvatic or domestic cycles. The sylvatic cycle is widespread on all continents, from frigid to torrid zones, and it is maintained by cannibalism and scavenging behavior of carnivores. Trichinella is primarily a parasite of carnivorous mammals, although one non-encapsulated species, Trichinella pseudospiralis, has also been detected in birds. The anaerobic metabolism of larvae in nurse cells allows their survival in extremely decayed meat. Encapsulated larvae in the decomposing carcass function similarly to the species-dispersing population of eggs or larvae of other nematodes, suggesting that the natural cycle of Trichinella includes a free living stage when the parasite is no longer protected by the homeothermy of the host. Consequently, environmental temperature and humidity play an important role in the transmission of Trichinella among wildlife. Of the 10 recognized genotypes of Trichinella, only Trichinella spiralis is transmitted and maintained in a domestic cycle, although it can be present also in wildlife. All other genotypes (Trichinella nativa, Trichinella britovi, T. pseudospiralis, Trichinella murrelli, Trichinella nelsoni and Trichinella papuae, Trichinella T6, T8, and T9) are transmitted and maintained only in a sylvatic cycle. This generalization does not preclude sylvatic species of Trichinella from invading the domestic habitat, and T. spiralis may return to this habitat when humans fail in the management of wildlife and domestic animals. However, the presence of sylvatic genotypes of Trichinella in the domestic habitat represents a "dead-end" for the sylvatic cycle. Synanthropic animals (rats, foxes, mustelids, cats, dogs, etc.) contribute to the flow of sylvatic Trichinella genotypes from wildlife to domestic animals and of T. spiralis from domestic to sylvatic animals. Furthermore, human behavior not only influences the transmission patterns of Trichinella genotypes in the domestic habitat, but also it can contribute to the transmission and spread of this infection among wildlife, for example by improper hunting practices. PMID- 11099841 TI - Host diversity and biological characteristics of the Trichinella genotypes and their effect on transmission. AB - The host spectra and biological diversity of the Trichinella genotypes are reviewed. While all genotypes appear to reproduce equally well in carnivore hosts, their infectivity and persistence in omnivores and herbivores show remarkable differences. Most of the genotypes found in wildlife have low infectivity for pigs and some persist only for a few weeks; in herbivores this tendency is even more profound, but malnourished, environmentally stressed, or otherwise immuno-suppressed hosts are likely to be more susceptible to Trichinella genotypes that would otherwise cause no, or only low level infection in that particular host species. In the domestic habitat (e.g. domestic pig farms), Trichinella spiralis is found almost exclusively, but in the sylvatic habitat the other Trichinella genotypes have found individual ecological niches. Thus, when environmental stress is limited in the domestic habitat, the high reproductive capacity of T. spiralis has a selective advantage, but in nature, the tolerance of other (sylvatic) genotypes to high and low temperatures and decomposition of host tissue might be more important. Parasite distribution according to muscle appear to be independent of the genotype of Trichinella and predilection sites are primarily determined by host species and secondarily by the age and level of infection. The biological diversity of the Trichinella genotypes should definitely be considered when planning experimental studies, as the uniform high infectivity of all genotypes in carnivores probably make them more suited for comparative studies than rodents. PMID- 11099842 TI - Molecular and biochemical methods for parasite differentiation within the genus Trichinella. AB - Delineation of the genus Trichinella into a more complex group of parasites has substantially motivated investigators to better identify and characterize the species and genotypes that form the basis of their investigations. Because of the cosmopolitan geographical distribution and broad host range that typify this genus, assigning unique biological, immunological and biochemical characters to each species and genotype has been essential for researchers to further advance this field. Numerous groups have developed simple methods to differentiate the genotypes, and by so doing, have generated diagnostic keys that accurately reflect the distinct differences among parasites of this group. Throughout the years, many methods have been used to accomplish this task, beginning with isoenzyme analyses and the use of repetitive DNA probes, to employing the polymerase chain reaction (PCR) and more state-of-the-art technologies. This review article summarizes the development of these methods with emphasis on molecular techniques and the ultimate goal of providing a simple, rapid and reproducible test to differentiate Trichinella parasites at the highest level of sensitivity, i.e. single parasite. PMID- 11099843 TI - The systematics of the genus Trichinella with a key to species. AB - The authors review the major biological, biochemical, and molecular characters that are used to distinguish the seven Trichinella species (T. spiralis, T. nativa, T. britovi, T. pseudospiralis, T. murrelli, T. nelsoni, T. papuae) and three genotypes whose taxonomic status is yet uncertain (T-6, T-8, T-9). A comparison of host specificity, morphology, reproductive abilities, nurse cell development and freeze resistance is presented, along with useful biochemical and molecular markers. Finally, this information is used to construct a diagnostic key for the species. A phylogenetic classification of the species is needed. PMID- 11099844 TI - Trichinella in horses: a low frequency infection with high human risk. AB - After the initial report in 1976 of a trichinellosis epidemic caused by the consumption of infected horsemeat, 12 other outbreaks have been described in Europe. Since the first serious human outbreak several experiments have confirmed the susceptibility of horses to Trichinella species and the rapid disappearance of specific antibodies in this host that prevents the use of serological methods for routine screening. A review of the distribution of parasite burdens in muscles of naturally or experimentally infected horses indicates that the tongue is the most likely sample to contain detectable numbers of Trichinella larvae in low level infections. Requirements for testing of horsemeat are specified in legislation of the European Union, and other recommendations are published elsewhere. The EEC directives have evolved into very specific requirements which specify the testing of at least 5g of tongue, masseter or diaphragm per horse using a pooled digestion assay. More recently, France has revised the requirement for sample size to 10g for horsemeat originating from countries with high prevalence of Trichinella. To address the continuing outbreaks of human trichinellosis due to infected horsemeat, the development and implementation of a quality assurance system for testing is being considered. PMID- 11099845 TI - The occurrence and ecology of Trichinella in marine mammals. AB - Trichinella in marine mammals has a circumpolar arctic distribution and a narrow range of host species. It is commonly found in polar bears (Ursus maritimus), and increasingly in walruses (Odobenus rosmarus) where it presents a significant zoonotic hazard. This has resulted in the implementation of food safety programs in some arctic communities to test harvested walrus meat for Trichinella larvae prior to consumption. Trichinella has been reported infrequently in bearded seals (Erignathus barbatus) and ringed seals (Phoca hispida), and was once observed in a Beluga whale (Delphinapterus leucas). Cannibalism is probably the most important factor in maintaining a Trichinella cycle in polar bears. Arctic carnivores such as polar bears, arctic foxes (Alopex lagopus) and domestic dogs (Canis familiaris) have a high prevalence of Trichinella infection and the carcasses of at least some of these animals are deposited in the ocean. Scavenging of these carcasses by walruses probably occurs, but may not account for the high prevalence the parasite seen in this host species. Predation, carrion feeding and cannibalism have been documented for walruses and a sylvatic cycle similar to that of bears may exist in walrus populations. Seals and whales are likely infected through infrequent exposure to infected carcasses, either directly by scavenging or indirectly by consuming amphipods or fish that have fed on infected carcasses. The inefficiency of this mechanism may account for the low prevalence of Trichinella in seals and whales. It is known that isolates from marine mammals are cold tolerant, and infectious for man, and have been identified as Trichinella nativa (T2). Molecular and other phylogenetic studies would be useful to facilitate studies on the inter-relationship of Trichinella cycles involving marine and terrestrial mammals in the arctic and subarctic, and in the investigation of human outbreaks of trichinellosis in these areas. PMID- 11099846 TI - Detection of Trichinella infection in food animals. AB - The first part of this review article deals with classical methods used for the detection of Trichinella larvae in muscle samples of those animal species which are recognized as traditional sources of trichinellosis for human beings, as well as those species which are important for epidemiological reasons. Special consideration is given to the main applications of these methods (routine slaughter inspection, and epidemiological studies in reservoir animals), and to the major factors that may influence detection methods (sampling site, sample size). Historical, current and future aspects concerning national and EU legislation for Trichinella inspection are also presented. The latter part of this review is directed at serodiagnostic methods for the detection of Trichinella-specific antibodies in different animal species. Classical methods of serodiagnosis such as the complement fixation test and immunofluorescence antibody test are reviewed and the characteristics and performance of the ELISA are discussed. Factors dependent upon the animal species being tested or on components of the ELISA test system are considered. This paper also reviews systematic development of the ELISA in relation to improvements in test specificity and sensitivity. Additionally, remarks are made on implementing this test for surveillance and control programs in domestic pigs and wildlife. PMID- 11099847 TI - National occurrence reporting of Trichinella and trichinellosis using a computerized database. AB - Historical and contemporary national data on the occurrence and distribution of Trichinella and trichinellosis in people, domestic animals and wildlife in Canada have been incorporated into a computerized database, the Canada Database of Animal Parasites (CDAP). This database was established in 1998, and contains similar information on several other helminth and protozoan parasites which are of importance in animal health, human health, food safety or trade. The CDAP is a unique assemblage of national information on parasite occurrences, not available from any other single source. This paper describes the CDAP, with emphasis on sources of data, database structure and outputs, logistical issues associated with database development and maintenance, and the application of the CDAP to Trichinella surveillance at a national level. It is suggested that the CDAP, or a similar approach, could be applied in other countries for assembling data on Trichinella and trichinellosis. PMID- 11099848 TI - Trichinellosis: human disease, diagnosis and treatment. AB - In this review, the pathological mechanisms of human trichinellosis are presented, including a discussion on organ pathology, with particular attention paid to intestinal and muscular invasion. The clinical pattern in the acute stage of trichinellosis is presented, together with a classification of trichinellosis relative to severity of the disease. In turn, complications and diagnostic criteria are discussed. Drugs employed in the contemporary treatment of trichinellosis are presented (mainly those of the benzimidazole group and glucocorticosteroids) as well as indications for administering them, as related to severity of the disease. PMID- 11099849 TI - Control of trichinellosis by inspection and farm management practices. AB - The prevention of human trichinellosis by proper meat inspection is a classic example of successful veterinary public health measures. The microscopic methods which have been used for more than a century to test pigs for trichinae were intended to prevent human disease. However, the value of these relatively insensitive direct detection methods, including trichinoscopy and pooled sample digestion, was debated as soon as more sensitive indirect (serological) methods became available. Two issues related to testing were discussed. First, should public health authorities endeavour to prevent all infections of humans rather than simply prevent the occurrence of disease, and second, would epidemiological surveillance and monitoring of the pig population on farms not provide a better control system to prevent human infection. This latter issue is of particular importance for those countries in the world where human trichinellosis acquired from farmed animals is absent and examination of pigs at the abattoir only results in negative findings. In countries where domestic pig infections are virtually non-existent, monitoring of Trichinella infection in wildlife could also contribute to understanding the infection pressure from nature to livestock. Trichinella-free pig farming is a feasible option for controlling this zoonosis, even in endemic areas. This approach provides an opportunity to combine good veterinary practice, in order to prevent animal diseases, with the prevention of Trichinella infection. All animals with access to the environment, or animals which are fed with potentially Trichinella-infected feed (swill, carcasses) will always constitute a public health threat, and must be inspected individually at slaughter (swine, horses, wild boars). Finally, it is important to recognize that trichinellosis is a world-wide problem that needs continuous public health attention. If no control system exists, for whatever reason, the public should be educated not to consume improperly cooked meat. PMID- 11099850 TI - International Commission on Trichinellosis: recommendations on methods for the control of Trichinella in domestic and wild animals intended for human consumption. AB - This document provides a uniform set of recommendations for the control of Trichinella at all levels (on the farm, at slaughter and in processed meats). These recommendations are based on the best scientific information available and represent the official position of the International Commission on Trichinellosis regarding acceptable control methods. These recommendations are subject to change as new scientific information becomes available. PMID- 11099851 TI - Electron spin resonance of copper(II) as a tool for the determination of asparagine concentration in Bacillus subtilis cultures. AB - A procedure is presented that is based on the detection of Cu(II)-asparagine complexes by quantitative ESR, and allows in a very simple and rapid manner to evaluate the changes of asparagine concentration during the entire time range of the growth of Bacillus subtilis in a typical growth medium. The analysis is carried out in terms of the decrease of the intensity of the ESR-active mono- and di-asparagine copper(II) complexes. It is resulted that at the end of the exponential growth the asparagine concentration was reduced to values as low as 2% of the initial value. The procedure here reported may be the basis of similar methods to be used for other amino acids and prokaryote systems. PMID- 11099852 TI - New long-wavelength perylenequinones: synthesis and phototoxicity of hypocrellin B derivatives. AB - Five new derivatives of hypocrellin B were obtained from the reactions of hypocrellin B with ammonia and ethanolamine. Their photophysical and photochemical properties were investigated. The phototoxicity of one compound on AH cells irradiated with red light (lambda = 600-700 nm) was also studied. Their significantly enhanced red absorptivities at wavelengths longer than 600 nm and singlet oxygen-generating function qualify them as promising photodynamic therapy agents. PMID- 11099853 TI - Relative position of the hexahistidine tag effects binding properties of a tumor associated single-chain Fv construct. AB - Hexahistidine tag (His-tag) is the most widely used tag for affinity purification of recombinant proteins for their structural and functional analysis. In the present study, single chain Fv (scFv) constructs were engineered form the monoclonal antibody (MAb) CC49 which is among the most extensively studied MAb for cancer therapy. For achieving efficient purification of scFvs by immobilized metal-ion affinity chromatography (IMAC), a His-tag was placed either at the C terminal (scFv-His6) or N-terminal (His6-scFv) of the coding sequence. Solid phase radioimmunoassay for scFv-His6 showed only 20-25% binding whereas both His6 scFv and scFv (no His-tag) showed 60-65% binding. Surface plasmon resonance studies by BIAcore revealed the binding affinity constant (KA) for His6-scFv and scFv as 1.19 x 10(6) M(-1) and 3.27 x 10(6) M(-1), respectively. No K(A) value could be calculated for scFv-His6 due to poor binding kinetics (kon and koff). Comparative homology modeling for scFv and scFv-His6 showed that the C-terminal position of the His-tag partially covered the antigen-binding site of the protein. The study demonstrates that the C-terminal position of His-tag on the CC49 scFv adversely affects the binding properties of the construct. The results emphasize the importance of functional characterization of recombinant proteins expressed with purification tags. PMID- 11099854 TI - In vivo chitin-cadmium complexation in cell wall of Neurospora crassa. AB - Fungal cell wall, mainly composed of chitin, an N-acetylglucosamine polymer, is known to participate in heavy metal detoxification. In the present study, an effort was made to elucidate the sites involved in complexation of cadmium by the chitin material of cell wall of Neurospora crassa. Based on the results of physical techniques, such as solid-state 13C-NMR, X-ray diffraction, IR and molecular modeling, a structure was proposed for the chitin-cadmium complex. The ring and C-3 hydroxyl oxygens of N-acetylglucosamine were implicated in the complexation of cadmium by the chitin of the fungal cell wall. The studies further revealed that the conformation of chitin did not alter after cadmium complexation. PMID- 11099855 TI - Photosensitized damage to calf thymus DNA by a hypocrellin derivative: mechanisms under aerobic and anaerobic conditions. AB - The di-cysteine substituted hypocrellin B (DCHB) derivative has been found to be a potential phototherapeutic agent and exhibit photosensitized damage to DNA. Electronic paramagnetic resonance (EPR) and spectrophotometry demonstrate that one-electron transfer from calf thymus DNA to triplet DCHB induces the generation of the reduced form of DCHB (DCHB*- radical), followed by the second electron transfer from DNA to DCHB*- or the disproportionation of DCHB*- to form the hydroquinone of DCHB (DCHBH2) in anaerobic conditions. This electron transfer process induces the direct damage to DNA in oxygen-free media and contributes partly to the damage of DNA in aerobic media. Superoxide radical and hydroxyl radical are formed with enhanced efficiencies while singlet oxygen is generated with a reduced efficiency from irradiation of DCHB and DNA solution under aerobic conditions as compared with the case in the absence of DNA. All of three reactive oxygen species play an evident role in the photosensitized damage to DNA in aerobic system in addition to the direct electron-transfer damage. PMID- 11099856 TI - Induction of hepatic antioxidants in freshwater catfish (Channa punctatus Bloch) is a biomarker of paper mill effluent exposure. AB - Enzymatic and non-enzymatic antioxidants serve as an important biological defense against environmental oxidative stress. Information on antioxidant defense in fish is meager despite that fish are constantly exposed to a myriad of environmental stress including the oxidants. This study, therefore, assesses the activities of antioxidant enzymes viz., glutathione peroxidase, catalase and glutathione S-transferase and the non-enzymatic antioxidants viz., glutathione and metallothionein in various tissues of freshwater fish Channa punctatus (Bloch), in response to short-term and long-term exposures to paper mill effluent. The fish were exposed to the effluent at a concentration of 1.0% (v/v) for 15, 30, 60 and 90 days. The exposure caused a time-dependent increase in glutathione level (P < 0.001), activities of glutathione peroxidase (P < 0.05 to P < 0.001), glutathione S-transferase (P < 0.001) and a marginal initial decrease in catalase activity in the liver (P < 0.01 to P < 0.001). Metallothionein was induced in liver after 60 days of exposure. Two isoforms of metallothionein were detected. Catalase activity also increased 60 days afterwards. Antioxidant pattern was different in gill and kidney showing that liver was more resistant to oxidative damage as compared to gills and kidney. Our results demonstrate a pollutant-induced adaptive response in fish. In addition, levels of enzymatic and non-enzymatic tissue antioxidants may serve as surrogate markers of exposure to oxidant pollutants in fish. PMID- 11099857 TI - Role of intestinal surfactant-like particles as a potential reservoir of uropathogenic Escherichia coli. AB - The binding of uropathogenic Escherichia coli is mediated at the tips of pili by the PapG adhesin, which recognizes the Galalpha(1-4)Gal disaccharide on the uroepithelial surface. These receptors have been identified unequivocally in the human and murine urinary tracts but not in intestinal epithelium, yet uropathogenic E. coli strains are commonly found in normal colonic microflora. The gastrointestinal tract from duodenum to rectum elaborates a phospholipid-rich membrane particle with surfactant-like properties. In these studies, we report that purified murine particles contain a receptor recognized by the class I PapG adhesin because: (1) PapD-PapG complexes and class I pili bound to surfactant like particles in a solid-phase assay, whereas binding was not detected in microvillous membranes derived from the same tissues, (2) purified PapD-PapG complex bound to a glycolipid receptor detectable in lipid extracts from the particles, and (3) soluble Galalpha(1-4)Gal inhibited the adhesin by 72% from binding to surfactant-like particles. The Galalpha(1-4)Gal receptor present in the intestinal surfactant-like particle which overlies the intestinal mucosa could provide one means to establish an intestinal habitat for uropathogenic E. coli. PMID- 11099858 TI - Spectroscopic detection of adrenaline-quinone formation in micelles. AB - Spectral changes, from 200 nm to 600 nm, of the oxidation of adrenaline to adrenochrome induced by periodate in electrically charged and neutral micelles at pH 3.77 were studied. The observed variations of the peak position, intensity and shape of the fluorescence spectra indicated that depending on the charge of the micelle adrenaline ion is partially embedded into the micellar core. Fluorescence lifetime measurements using Omnilyzer allowed to calculate partition coefficients of 0.36, 0.05 and 0.01 in sodium dodecyl sulphate, tetradodecyltrimethylammonium bromide and Triton X-100, respectively. Kinetics of adrenaline decay during oxidation were followed by its fluorescence what overcame spectral interference in the absorption spectra of adrenaline from the formed intermediates. Scanning absorption spectroscopy, with 100 ms resolution, allowed the recording of spectral changes during the transformation. With this method, the formation of adrenaline-quinone with absorption maxima at 388 nm and 274 nm was detected. The calculated rate constants of the observed kinetics during oxidation were significantly lowered in both charged micelles compared to buffer solution and in Triton X-100 neutral micelles. The observed phenomena are discussed in terms of the electrostatic forces mechanism and in the frame of the Raper-Mason scheme of adrenaline transformation. PMID- 11099859 TI - Differential regulation of [Ca2+]i oscillations in mouse pancreatic islets by glucose, alpha-ketoisocaproic acid, glyceraldehyde and glycolytic intermediates. AB - Glucose induces slow oscillations of the cytoplasmic Ca2+ concentration in pancreatic beta-cells. In order to elucidate the mechanisms responsible for the slow [Ca2+]i oscillations the effects of various nutrient insulin secretagogues on glucose-induced [Ca2+]i oscillations in intact mouse pancreatic islets and single beta-cells were studied. These were the glycolytic intermediates, glyceraldehyde and pyruvate, and the mitochondrial substrate, alpha ketoisocaproic acid (KIC). Glucose, at a 10 or 15 mM concentration, induced the typical slow oscillations of [Ca2+]i (0.4 min(-1)). At higher glucose concentrations the frequency of these oscillations decreased further (0.2 min( 1)). Glyceraldehyde, an insulin secretagogue like glucose, did not cause slow oscillations of [Ca2+]i in the absence of glucose. However, it exhibited a synergistic action with glucose. Glyceraldehyde, at 3 or 10 mM concentration, induced slow [Ca2+]i oscillations at a substimulatory concentration of 5 mM glucose (0.3-0.4 min(-1)) and reduced the frequency of the glucose-induced [Ca2+]i oscillations at stimulatory concentrations of 10 or 15 mM glucose (0.2 min(-1)). KIC (5 or 10 mM) as well as pyruvate (10 mM), the end product of glycolysis, and its ester methyl pyruvate (10 mM), did not cause slow oscillations of [Ca2+]i in the absence of glucose. In contrast to glyceraldehyde, however, all three compounds were capable of preventing the slow [Ca2+]i oscillations induced by glucose. Mannoheptulose (2 mM), an inhibitor of glucokinase and glucose-induced insulin secretion, reversibly blocked any kind of [Ca2+]i oscillation and returned the [Ca2+]i to a basal level through its ability to inhibit glycolytic flux. It can be concluded therefore that only substrates which generate a glucokinase-mediated metabolic flux through glycolysis and produce glycolytic ATP can induce slow [Ca2+]i oscillations in pancreatic beta cells. PMID- 11099860 TI - A validated model of in vivo electric field distribution in tissues for electrochemotherapy and for DNA electrotransfer for gene therapy. AB - Permeabilising electric pulses can be advantageously used for DNA electrotransfer in vivo for gene therapy, as well as for drug delivery. In both cases, it is essential to know the electric field distribution in the tissues: the targeted tissue must be submitted to electric field intensities above the reversible permeabilisation threshold (to actually permeabilise it) and below the irreversible permeabilisation threshold (to avoid toxic effects of the electric pulses). A three-dimensional finite element model was built. Needle electrodes of different diameters were modelled by applying appropriate boundary conditions in corresponding grid points of the model. The observations resulting from the numerical calculations, like the electric field distribution dependence on the diameter of the electrodes, were confirmed in appropriate experiments in rabbit liver tissue. The agreement between numerical predictions and experimental observations validated our model. Then it was possible to make the first precise determination of the magnitude of the electric field intensity for reversible (362+/-21 V/cm, mean +/- S.D.) and for irreversible (637+/-43 V/cm) permeabilisation thresholds of rabbit liver tissue in vivo. Therefore the maximum of induced transmembrane potential difference in a single cell of the rabbit liver tissue can be estimated to be 394+/-75 and 694+/-136 mV, respectively, for reversible and irreversible electroporation threshold. These results carry important practical implications. PMID- 11099861 TI - Nuclear coactivator protein p100 is present in endoplasmic reticulum and lipid droplets of milk secreting cells. AB - We have identified the p100 protein, previously known as a novel cellular coactivator, as a constituent of endoplasmic reticulum and cytosolic lipid droplets from milk secreting cells. Cytosolic lipid droplets of terminally differentiated mammary epithelial cells are secreted as milk lipid globules. However, milk lipid globules did not have detectable amounts of p100 protein. The p100 protein was found also in cytosol from lactating mammary gland, in storage lipid droplets from mouse adipocytes, and in endoplasmic reticulum from liver. Immunofluorescence microscopy of mammary epithelial cells confirmed the presence of p100 in non-nuclear regions of these cells. Partial sequence analysis of tryptic peptides from p100 from cow mammary gland showed extensive homology with the reported sequence of p100 determined from a human cDNA. Antibodies against a peptide synthesized to duplicate a sequence in human p100 recognized a protein of the size of p100 in cow, mouse and rat cell fractions. PMID- 11099862 TI - Interactions of dietary protein and carbohydrate determine blood sugar level and regulate nutrient selection in the insect Manduca sexta L. AB - The non-homeostatic regulation of blood sugar concentration in the insect Manduca sexta L. was affected by nutritional status. Larvae maintained on diets lacking sucrose displayed low concentrations of trehalose, the blood sugar of insects, which varied from 5 to 15 mM with increasing dietary casein level between 12.5 and 75 g/l. These insects were glucogenic, as demonstrated by the selective 13C enrichment of trehalose synthesized from [3-13C]alanine, and de novo synthesis was the sole source of blood sugar. The distribution of 13C in glutamine established that following transamination of the 13C substituted substrate, [3 13C]pyruvate carboxylation rather than decarboxylation was the principal pathway of Pyr metabolism. The mean blood trehalose level was higher in insects maintained on diets with sucrose. At the lowest dietary casein level blood trehalose was approximately 50 mM, and declined to 20 mM at the highest casein level. Gluconeogenesis was detected in insects maintained on sucrose-free diets at the higher protein levels examined, but [3-13C]pyruvate decarboxylation and TCA cycle metabolism was the principal fate of [3-13C]alanine following transamination, and dietary carbohydrate was the principal source of glucose for trehalose synthesis. Feeding studies established a relationship between nutritional status, blood sugar level and dietary self-selection. Insects preconditioned by feeding on diets without sucrose had low blood sugar levels regardless of dietary casein level, and when subsequently given a choice between a sucrose diet or a casein diet, selected the former. Larvae preconditioned on a diet containing sucrose and the lowest level of casein had high blood sugar levels and subsequently selected the casein diet. Larvae maintained on the sucrose diet with the highest casein level had low blood sugar and self-selected the sucrose diet. When preconditioned on diets with sucrose and intermediate levels of casein, insects selected more equally between the sucrose and the casein diets. It is concluded that blood sugar level may be intimately involved in dietary self-selection by M. sexta larvae, and that in the absence of dietary carbohydrate, gluconeogenesis provides sufficient blood sugar to ensure that larvae choose a diet or diets that produce an optimal intake of dietary protein and carbohydrate. PMID- 11099863 TI - Strength of conjugate binding to plasmid DNA affects degradation rate and expression level in vivo. AB - In vitro assays have demonstrated the capability of poly-L-lysine to protect plasmid DNA from serum nucleases and cellular lysates. Our purpose was to evaluate the stability and potency of poly-L-lysine-DNA polyplexes after intravenous injection into mice. Polyplexes consisted of 32P-radiolabeled plasmid DNA complexed with poly-L-lysine at specified charge ratios. Variations in conjugate hydrophobicity and levels of modification with polyethylene glycol were investigated. Our results show that, in contrast to in vitro studies, the systemically administered polyplexes exhibited marked DNA degradation in the vascular compartment within 5 min. Substitution of poly-L-lysine epsilon-amino sites with polyethylene glycol or hydrocarbon chains resulted in faster degradation even when complexed at higher charge (+/-) ratios. Use of excess cationic charge in the polyplexes (+/- 2.5) diminished degradation rates only slightly. An analysis was made of the strength of the poly-L-lysine:DNA interaction by competition with poly-aspartic acid. Polyplexes with the strongest binding between conjugate and DNA in the competition assay were also the most stable in blood. However, tighter binding was not enough to fully protect the polyplex in vivo and polyplex DNA was substantially degraded within 10 min. Increased polyplex stability did not correlate with improved in vivo transfection efficiency. PMID- 11099864 TI - Polycarboxylates inhibit the glucan-binding lectin of Streptococcus sobrinus. AB - Polycarboxylates, such as carboxymethylcellulose and hyaluronan, were found to be reversible inhibitors of the glucan-binding lectin of Streptococcus sobrinus. When the carboxylate groups were coupled to ethylenediamine, or reduced with carbodiimide-borohydride, inhibitory powers were lost. Similarly, N-deacetylated hyaluronan had poor inhibitory powers, probably due to the introduction of positive charges into the polymer. Other polymers, such as chondroitin sulfates, dextran sulfate, fetuin, heparin were not inhibitors. It appears that inhibition is based on repeating carboxylates, free of influence from ammonium groups. Such polymers have the property of complexing with metals. Earlier studies had concluded that the streptococcal lectin depended on manganese for activity. It is likely the carboxymethylcellulose and hyaluronan perturb essential metal coordination centers in the lectin. Polycarboxylates may have value in oral health care by acting on glucan-dependent microbial adhesion and biofilm formation. PMID- 11099865 TI - Polyphenolics enhance red blood cell resistance to oxidative stress: in vitro and in vivo. AB - In this study we investigated the potential antioxidant properties of blueberry polyphenolics in vitro and vivo, using red blood cell (RBC) resistance to reactive oxygen species (ROS) as the model. In vitro incubation with anthocyanins or hydroxycinnamic acids (HCA) (0.5 and 0.05 mg/ml) was found to enhance significantly RBC resistance to H2O2 (100 microM) induced ROS production. This protection was also observed in vivo following oral supplementation to rats at 100 mg/ml. However, only anthocyanins were found to afford protection at a significant level, this at 6 and 24 h post supplementation. This protection was not consistent with the measured plasma levels of anthocyanins. Indeed, plasma polyphenolic concentrations were highest after 1 h, declining considerably after 6 h and not detected after 24 h. The difference in absorption between anthocyanins and HCA is likely to have contributed to the observed difference in their abilities to afford protection to RBC. This protection represents a positive role following dietary consumption of polyphenolics from blueberries, against ROS formation within RBC in vivo. PMID- 11099866 TI - Involvement of arginine and tryptophan residues in catalytic activity of glutaryl 7-aminocephalosporanic acid acylase from Pseudomonas sp. strain GK16. AB - The glutaryl 7-aminocephalosporanic acid (GL-7-ACA) acylase from Pseudomonas sp. strain GK16 is an (alphabeta)2 heterotetramer of two non-identical subunits that are cleaved autoproteolytically from an enzymatically inactive precursor polypeptide. The newly formed N-terminal serine of the beta subunit plays an essential role as a nucleophile in enzyme activity. Chemical modification studies on the recombinant enzyme purified from Escherichia coli revealed the involvement of a single arginine and tryptophan residue, per alphabeta heterodimer of the enzyme, in the catalytic activity of the enzyme. Glutaric acid, 7 aminocephalosporanic acid (7-ACA) (competitive inhibitors) and GL-7-ACA (substrate) could not protect the enzyme against phenylglyoxal-mediated inactivation, whereas except for glutaric acid protection was observed in case of N-bromosuccinimide-mediated inactivation of the enzyme. Kinetic parameters of partially inactivated enzyme samples suggested that while arginine is involved in catalysis, tryptophan is involved in substrate binding. PMID- 11099867 TI - Impaired ability of glycated insulin to regulate plasma glucose and stimulate glucose transport and metabolism in mouse abdominal muscle. AB - Previous studies have shown that glycated insulin is secreted from pancreatic beta-cells under conditions of hyperglycaemia. This study has investigated the effects of monoglycated insulin on plasma glucose homeostasis and in vitro cellular glucose transport and metabolism by isolated abdominal muscle of mice. Monoglycated insulin was prepared under hyperglycaemic reducing conditions, purified by RP-HPLC and identified by electrospray ionisation mass spectrometry (5971.1 Da). When administered to mice at an intraperitoneal dose of 7 nmoles/kg body weight, insulin (non-glycated) decreased plasma glucose concentrations and substantially reduced the glycaemic excursion induced by conjoint intraperitoneal injection of 2 g glucose/kg body weight. In comparison, the same dose of monoglycated insulin decreased plasma glucose concentrations to a lesser extent (P < 0.05), corresponding to an approx. 20% reduction of glucose lowering potency. Using isolated abdominal muscle, insulin (10(-9)-10(-7) M) stimulated dose-dependent increases in cellular 2-deoxy-D-[1-3H]glucose uptake, D-[U 14C]glucose oxidation and glycogen production. Monoglycated insulin was approx. 20% less effective than native insulin in stimulating glucose uptake and both indices of metabolism, generally requiring 10-fold greater concentrations to achieve significant stimulatory effects. These data indicate that the impaired biological activity of glycated insulin may contribute to glucose intolerance of diabetes. PMID- 11099868 TI - Canadian menopause study-I: Understanding women's intentions to utilise hormone replacement therapy. AB - OBJECTIVES: This research examines women's intentions to use or to continue to use HRT, as a function of perceived advantages and disadvantages of HRT and perceived social support for utilising HRT. METHODS: A cross-national sample of 205 pre-, peri-, and postmenopausal women convened in 33 'Women's Health Discussion Groups' in small, mid-sized, and large cities across the regions of Canada. Participants completed close-ended questionnaires assessing perceptions of advantages, disadvantages, and social support for HRT, and intentions to utilise or to continue to utilise HRT in the future. RESULTS: Women who intended to utilise or to continue to utilise HRT, compared with women who did not, perceived significantly more advantages of HRT and significantly more social support for utilising HRT, and did not, in general, differ in their perceptions of the negative side effects of HRT. Regression analysis indicated that perceived advantages of HRT and perceived social support for using HRT, but not perceived negative side effects of HRT, were significantly associated with intentions to utilise or to continue to utilise therapy and explained 46% of the variance in these intentions. CONCLUSIONS: Findings suggest that the importance of perceived health benefits of HRT and perceived social support for HRT may be substantially underestimated, and that the importance of perceived negative side effects of HRT may be substantially overestimated, in understanding women's HRT decision making and in counselling women about initiation or maintenance of therapy. PMID- 11099869 TI - Heritability and risk factors of uterine fibroids--the Finnish Twin Cohort study. AB - OBJECTIVES: Our aim was to study heritability, risk factors and hospitalization for uterine fibroids. METHODS: A random sample of 80 MZ and 80 DZ twins from the Finnish Twin Cohort were invited and 51% of the eligible women (n=82, 17 MZ and 16 DZ pairs, 40-47 years, mean age 43.0), underwent a transvaginal ultrasound. The entire cohort of 13872 women was linked to the national hospital discharge registry 1972-1990. RESULTS: Prevalence of fibroids was 66% and the average number of fibroids 1.7. The casewise concordance for being hospitalized for uterine fibroids was higher in MZ (0.31, 95% CI 0.24-0.37) than in DZ pairs (0.18, 95% CI 0.14-0.22). The proportion of variance in liability to fibroid hospitalization accounted for by genetic factors was 54.8% (95% CI 46.2-62.7%). Women with fibroids had higher body mass index (23.7 vs 21.7, P=0.0086), lower age at first birth (25.7 vs 29.3, P=0.012) and higher parity (3+ children 48.2 vs 29.6%, P=0.009) than women without fibroids. Risk ratio (RR) for fibroids in a MZ twin whose sister had been diagnosed with fibroids was 1.1 (95% CI 0.08;15), for a DZ twin 1.1 (95% CI 0.16;8.8) and for all twins 1.3 (95% CI 0.3; 6.1). Intraclass correlation for the number of fibroids was 0.24 for MZ and 0.11 for DZ twins, yielding an heritability estimate of 0.26. CONCLUSION: Reproductive and anthropometric factors may have at least as large role in pathogenesis of fibroids than genetic factors. PMID- 11099870 TI - Uterine myoma in postmenopause: a comparison between two therapeutic schedules of HRT. AB - OBJECTIVES: It is still controversial whether hormone replacement therapy (HRT) can affect the onset of uterine myomas or their growth in postmenopause. It is likely that some therapeutic schedules can influence the myometrial growth differently, due to a more potent stimulation of the uterine receptors. The aim of the present study is to evaluate the effects of two different hormonal treatment schedules on the risk of uterine myoma onset or progression. METHODS: In a 2 year prospective randomised study we compared an oral cyclic association of oestradiol valerate and cyproterone acetate versus a sequential combination of transdermal E(2) and per oral medrossiprogesterone acetate on 240 postmenopausal women with and without uterine myomas. RESULTS: Among the patients without uterine myomas treated with the transdermal-oral combination we noted the onset of myomas in 5% of cases after 24 months of treatment, while no new uterine formation was observed for the orally treated women (P<0.01). Among the patients with uterine myomas at the beginning of the study, in the group transdermally treated we found a mean increase in myoma volumes of 25.3% in the following 24 months, which was significantly different compared with the initial volume of myomas. On the other hand, women treated with the oral combination showed no significant modification of myoma volumes at the end of the study. CONCLUSIONS: Percutaneous-oral schedule of HRT seems to affect the growth of uterine myomas more than a single oral combination of oestradiol valerate and cyproterone acetate. PMID- 11099871 TI - Preperitoneal lipoleiomyoma of the abdominal wall in a postmenopausal woman. AB - OBJECTIVE: Extrauterine leiomyomas are rare events. These tumors may be easily misdiagnosed as ovarian tumors at the clinical investigation. We present the first case of an otherwise healthy postmenopausal woman, hysterectomized 20 years ago, who developed a preperitoneal lipoleimoyoma in the 30-year-old scar of a Pfannenstiel incision. The patient received continuous hormone replacement therapy (HRT) for 5 years with 1.25 mg conjugated estrogen and 5 mg medrogeston per day. METHODS: In sections of the tumor, immunohistochemical reactions with antibodies against actin, desmin, vimentin, estrogen and progesterone receptors and factor VIII related antigen was performed. RESULTS: Histologic findings revealed cellular fascicles of spindle-shaped smooth muscle cells in a whorled arrangement. Mitotic figures were absent. Central degenerative changes and focal edema were observed. Between muscle fascicles, a significant amount of fat cells (20% of tumor volume) was visible. Leiomyocytes showed immunohistochemicaly positive reactions with actin, desmin, vimentin, and steroid hormone receptors. Based on these findings, the tumor was diagnosed as lipoleiomyoma. CONCLUSIONS: Origin of the tumor of smooth muscle cells of vessels located in the abdominal wall and development under influence of oral steroids seems most probable. HRT appears to promote the development of extrauterine leiomyomas in postmenopausal women. PMID- 11099872 TI - Lipid effects, effectiveness and acceptability of tibolone versus transdermic 17 beta-estradiol for hormonal replacement therapy in women with surgical menopause. AB - OBJECTIVE: To compare the effectiveness of tibolone and 17 beta-estradiol on climacteric symptoms, lipid and biochemical parameters in women with surgical menopause. METHODS: In a prospective randomised clinical trial group comparative study, the effects on the aforementioned parameters, as well as treatment compliance and side effects were studied with oral tibolone 2.5 mg per day and with transdermic 17 beta-estradiol at 50 microg per day for a period of 12 months. Statistical analysis was carried out using the Fisher-test, analysis of the variance (ANOVA) for the two factors and the Bouferoni test. RESULTS: Lipid metabolism analysis showed lower levels of HDL and triglycerides in the tibolone group. Other biochemical parameters were not affected. Similar reductions in climacteric symptoms were found in both the groups, but the tibolone group revealed a greater improvement in psychological problems and in sexual behaviour. No differences were observed with respect to compliance and side effects. CONCLUSIONS: Tibolone is as effective or more than 17 beta-estradiol in reducing climacteric symptoms, and shows greater triglyceride and total cholesterol improvements. Tibolone is a good alternative to estrogens in women with surgical menopause. PMID- 11099873 TI - The effect of estrogen replacement therapy on cardiac autonomic regulation. AB - OBJECTIVE: To study the effects of estrogen replacement therapy (ERT) and sleep stage on autonomic cardiac regulation. SRUDY DESIGN: Seventy-one healthy postmenopausal women received transdermal ERT and placebo separated by a washout in a randomized, placebo-controlled, double-blind, cross-over trial. Polysomnography was conducted at the end of each treatment. Heart rate variability (HRV) was assessed in epochs of the awake state, stage 2, slow wave and REM sleep. The effects of estradiol and sleep stages on HRV were analyzed. RESULTS: ERT decreased heart rate in the awake state and quiet sleep, but not in REM sleep. ERT did not change the heart rate variability. Heart rate decreased and HRV increased during stage 2 and slow wave sleep compared with the awake state with placebo. In REM sleep, similarly, heart rate increased above awake values and the values of HRV parameters fell back to awake levels. CONCLUSIONS: The results suggest that ERT increases vagal tone, but does not change cardiac vagal modulation. Changes in HRV suggest a strong vagal influence in non-REM and a sympathetic influence in REM sleep. PMID- 11099874 TI - The rabbit as a model for reproductive and developmental toxicity studies. AB - The rabbit has many advantages as a nonrodent and second model for assessing the effects of toxic agents on semen quality, fertility, developmental toxicity, and teratology. The male and female reproductive systems of the rabbit are described, and data on growth, sexual development and reproduction are compared with mice, rats, and humans. Techniques for semen collection and evaluation in the male, and artificial insemination, superovulation, embryo culture, and embryo transfer in the female are included as useful procedures in toxicity testing. Examples of the use of rabbits and experimental replication for toxicity testing are given. Special features of the visceral yolk sac and development of the chorioallantoic placenta of the rabbit are compared with rodents. The rabbit extraembryonic membranes more closely resemble the human than do the rodents, in some respects. The use of the rabbit in developmental toxicity and teratology studies is discussed. PMID- 11099875 TI - Concordant in situ and in vitro data show that maternal cigarette smoking negatively regulates placental cytotrophoblast passage through the cell cycle. AB - Maternal cigarette smoking is associated with fetal growth restriction and other pregnancy complications. To investigate possible mechanisms involving the placenta, we studied the morphology of first trimester chorionic villi from mothers who smoked. In mothers who smoked > 20 cigarettes/day, floating villi showed focal defects including an absence of cytotrophoblast stem cells and an abnormal thinning of the syncytium. Anchoring villi displayed a striking increase in the number of cytotrophoblast columns that failed to reach the uterus or degenerated in the intervillous space. Many samples showed a significant reduction in the number of anchoring villi. Also, the number of Ki67-positive cytotrophoblasts was dramatically decreased, indicating that fewer cells were in S phase of the mitotic cycle. Together, these results suggested premature depletion of the cytotrophoblast stem cell population. To test this hypothesis, we exposed anchoring villi from nonsmokers to nicotine in vitro and analyzed the effects on cytotrophoblast passage through the cell cycle. Nicotine (0.23 to 6.0 microM) negatively affected the expression of a number of cell cycle regulators/markers and BrdU incorporation, without discernable effects on apoptosis. These results link abnormal placental development secondary to maternal cigarette smoking to a substantial decrease in the mitotic potential of cytotrophoblasts. PMID- 11099876 TI - Effect of ginger tea on the fetal development of Sprague-Dawley rats. AB - This study investigated the effect of ginger, a common morning sickness remedy, on fetal development. Pregnant Sprague-Dawley rats were administered, from gestation day 6 to 15, 20 g/liter or 50 g/liter ginger tea via their drinking water and then sacrificed at day 20. No maternal toxicity was observed, however embryonic loss in the treatment groups was double that of the controls (P<0.05). No gross morphologic malformations were seen in the treated fetuses. Fetuses exposed to ginger tea were found to be significantly heavier than controls, an effect that was greater in female fetuses and was not correlated with increased placental size. Treated fetuses also had more advanced skeletal development as determined by measurement of sternal and metacarpal ossification centers. The results of this study suggest that in utero exposure to ginger tea results in increased early embryo loss with increased growth in surviving fetuses. PMID- 11099877 TI - Effect of butyl benzyl phthalate in Sprague-Dawley rats after gavage administration: a two-generation reproductive study. AB - Butyl benzyl phthalate (BBP), a plasticizer, has been shown in in vitro studies to be weakly estrogenic, and in in vivo studies to possess testicular toxicity and teratogenicity, but few experimental data on BBP multigeneration effects on reproduction in mammals are available. The present two-generation reproductive study was conducted in male and female Sprague-Dawley rats using oral doses of 0, 20, 100, and 500 mg/kg/day BBP. Endpoints were chosen in order to evaluate both subchronic and reproductive toxicity. In the parent animals (F(0)), a decrease in body weight gain was observed in males in the 500 mg/kg/day group, although no significant decrease in food consumption was found. No dose-related changes were observed in estrous cyclicity, fertility, or lactation. A dose-dependent increase in kidney weight in rats of both sexes, an increase in liver weight in males, and a decrease in the weight of the ovaries in females were observed. No macroscopic or microscopic changes were found in the reproductive system of males or females. Oral administration of BBP caused a decrease in the serum concentration of testosterone, and an increase in FSH. In the next generation (F(1)), the body weight of male and female offspring at birth in the 100 and 500 mg/kg groups was significantly decreased, and the body weight in the 500 mg/kg group was lower throughout the study, while viability was not affected. Anogenital distance (AGD) at birth was decreased in male pups and was increased in female pups of the 500 mg/kg/day group. Preputial separation for male offspring in the 500 mg/kg/day group was delayed, while vaginal opening for female offspring in this group was not affected. BBP did not affect reproductive ability, including delivery and lactation, at any dose whereas macroscopic and microscopic changes of the testis, and decreased serum concentrations of testosterone were observed in male offspring of the 500 mg/kg/day group after puberty. From these data, it would appear that 20 mg/kg BBP is a no observed adverse effect level (NOAEL) for reproductive effects on parent animals and the next generation. PMID- 11099878 TI - Influence of the drinking water disinfection by-product dibromoacetic acid on rat estrous cyclicity and ovarian follicular steroid release in vitro. AB - The drinking water disinfection by-product, dibromoacetic acid (DBA) has been reported to affect gonadal functions in the male rat. However, there is little information regarding the influence of DBA on female reproductive activity. Consequently, the present study investigated the effects of DBA on estrous cyclicity and the impact in vitro of DBA on ovarian follicular steroid secretion. Regularly cycling animals were dosed with DBA (0 to 270 mg/kg/day) for 14 days and estrous cyclicity was monitored during treatment and for an additional 2-week posttreatment interval. A dose-related alteration in cyclicity was observed at 90 and 270 mg/kg/day, which persisted through the posttreatment monitoring in the high dose group. An in vitro exposure of preovulatory follicles to DBA was then used to assess the influence of DBA on steroid release. To select a concentration for use, a single oral exposure to 270 mg/kg was administered, and the mean blood levels were determined over a 5-h interval. For this in vitro work, pairs of preovulatory follicles from PMSG-primed immature rats were exposed to 0 or 50 microg/mL DBA over a 24-h period and evaluated for estradiol and progesterone release under baseline and hCG-stimulated conditions. The influence of tumor necrosis factor (TNFalpha) exposures under these conditions was also determined. In the nonstimulated condition, DBA was found to increase the release of estradiol, but had no detectable effect in response to hCG. Progesterone, however, showed marked suppression under hCG stimulation following exposure to DBA, while nonstimulated secretion was unaffected. TNFalpha by itself also suppressed stimulated progesterone release, but had no additional effect in combination with DBA. The data suggest that one factor in the disruption in estrous cyclicity could be an alteration in steroid production, which was characterized by separate effects on both estradiol and progesterone secretion. PMID- 11099879 TI - Ovarian hyperstimulation and oophorectomy following accidental daily clomiphene citrate use over three consecutive months. AB - We describe the longest-known continuous use of clomiphene citrate ever reported in a human. As a result of a pharmacy error, a woman took 50 mg/day clomiphene citrate for three months. The prolonged use of this medication resulted in ovarian hyperstimulation and unilateral oophorectomy for torsion. PMID- 11099880 TI - Edinburgh high risk study--findings after four years: demographic, attainment and psychopathological issues. AB - This study reports findings of the Edinburgh High Risk Study four years after it began. This study is designed to explore the pathogenesis of schizophrenia by examining a large sample of young adults aged 16-25 years who are at enhanced risk of developing schizophrenia by having two close relatives with the disorder, and comparing them with matched controls. This paper presents comparisons of the high risk subjects, well controls and subjects with first-episode schizophrenia in terms of demographic, childhood, psychopathological, educational and employment, forensic and social work variables. High risk subjects have more psychological difficulties, poorer educational and employment attainment, and more social work contact than controls. The enhanced social work involvement related to the presence of a schizophrenic parent (especially a mother) but the other difficulties could not be attributed to that situation. Neurotic, partially held psychotic and fully held psychotic symptoms all occurred in both subjects and controls, but all were significantly more common in high risk subjects. Clinical schizophrenia has so far developed in 10 high risk subjects and in no controls. Possible confounding effects of drug or alcohol misuse were considered but were found unlikely to be important. PMID- 11099881 TI - The antecedents of psychoses: a case-control study of selected risk factors. AB - Winter birth, urban birth and/or childhood residence, and perinatal complications have each been identified as environmental risk factors for the later development of schizophrenia, schizoaffective disorder, and bipolar disorder. A preliminary case-control study also identified cat exposure in childhood as a possible risk factor. To assess selected environmental events, including childhood exposure to pets, as possible risk factors for these diseases, a case-control telephone survey was carried out by the University of Maryland Survey Research Center for 264 mothers of cases and 528 mothers of matched controls. The cases were randomly selected mothers who were members of the National Alliance for the Mentally Ill, and whose children had been diagnosed with schizophrenia, schizoaffective disorder, or bipolar disorder. The controls were mothers randomly selected from the same telephone exchanges. For five of the 19 major variables, there were statistically significant differences between cases and controls: fever during pregnancy, complications during delivery, city or suburban residence at birth, cat ownership between birth and age 13, and breast-feeding. In a multivariate logistic regression including these five variables, each variable made a significant contribution. The finding of perinatal complications, urban/suburban residence at birth, and cat ownership in childhood as risk factors for the later development of psychoses confirmed previous studies. Previous research on breast feeding as a risk factor has yielded contradictory results. Additional research is needed to ascertain how such environmental risk factors interact with genetic risk factors. Understanding these could lead to better treatments and possible prevention strategies. PMID- 11099882 TI - The quantity and quality of the social networks of young people with early psychosis compared with closely matched controls. AB - This study compares the social networks and perceived social support of 26 people with early psychosis and 26 people without a mental illness. The two groups were closely matched for age, sex, education level, and employment and relationship status, and had equivalent levels of depression. There were no differences between the two groups in the amount of perceived social support, number of family members, and number of participants with acquaintances. However, the psychosis group identified significantly smaller networks, t (50)=-2.34, P=0.024, with fewer friends, t (48)=-3.61, P=0.001, fewer people to turn to in a crisis, t (22.97)=-2.34, P=0.028, and a higher likelihood of service providers as members, chi(2)(1)=7.02, P=0.008. Given the important relationship between strong social networks and high levels of community functioning and tenure, future research needs to evaluate the type of social support most beneficial for people with early psychosis and to develop strategies to maintain and facilitate that support. PMID- 11099883 TI - First-episode schizophreniform disorder: comparisons with first-episode schizophrenia. AB - BACKGROUND: Schizophreniform disorder remains poorly understood and has been reported probably to be a heterogeneous group of psychotic disorders. METHOD: This study compared first-episode schizophreniform disorder (N=12) and schizophrenia (N=18) patients. The authors propose that schizophreniform disorder has a different type of onset and outcome than schizophrenia. Patients were given extensive assessments at initial evaluation, 6 month follow-up, and 24 month follow-up. Comparisons between the two groups were made on type of onset, demographic, clinical ratings and outcome variables. RESULTS: Patients with schizophreniform disorder compared to patients with schizophrenia were more likely to have an acute onset (P=0.003), and have recovered by 6 months (P=0.03). However, there were no differences in outcome at 24 months. Furthermore, all schizophreniform cases except for two were re-diagnosed at 24 months as having schizophrenia. CONCLUSIONS: The findings suggest that the initial differences of schizophreniform disorder compared to schizophrenia were not apparent at 24 months follow-up. Schizophreniform disorder did not emerge as a highly distinctive and stable form of psychosis that merits a diagnostic classification separate from schizophrenia. PMID- 11099884 TI - Insular cortex abnormalities in schizophrenia: a structural magnetic resonance imaging study of first-episode patients. AB - The insular cortex is a limbic integration region that is engaged in emotional and cognitive functions. To investigate possible insular cortex abnormalities in schizophrenia, we measured insular gray matter volume and cortical surface size in drug-naive first-episode patients. Magnetic resonance images were used to explore the morphology of the insular cortex of 25 healthy male volunteers, and 25 male schizophrenic patients. Groups were matched for age, sex, height, and parental socio-economic status. Clinical dimension scores were correlated with insular gray matter volume and cortical surface area. Patients had a significant reduction in cortical surface area [patients=2020 (206); controls=2142 (204); F=5.83, df=1, 47; P=0.01] and gray matter volume [patients=8.12 (0.77); controls=8.57 (0.94); F=3.93, df=1,47; P=0.05] in the left insular cortex. Insular gray matter volume and cortical surface size correlated negatively and significantly with the psychotic symptom dimension. Schizophrenic patients show morphological abnormalities in the insular cortex at early stages of the illness. These abnormalities are related to the severity of psychotic symptoms. Further investigations are needed to evaluate the role of the insula in the pathophysiology of schizophrenia. PMID- 11099885 TI - Supporting evidence for the model of cognitive dysmetria in schizophrenia--a structural magnetic resonance imaging study using deformation-based morphometry. AB - The aim of the study was to investigate whether there is any structural evidence for the model of 'cognitive dysmetria' in schizophrenia if an automatic whole brain analysis method is used. High-resolution magnetic resonance scans were obtained for 75 schizophrenic patients and 75 controls. These data were analysed using the recently developed deformation-based morphometry allowing the assessment of volumetric differences without a priori definition of regions of interest. When compared with controls, we found reduced volumes in patients with schizophrenia in the frontal lobe (gyrus frontalis superior, medius and medialis), the temporal lobe (gyrus temporalis superior and inferior), the thalamus, the left cerebellar hemisphere and the right cerebellar vermis. There was an increase in volume in the right putamen. To date, this is the first structural magnetic resonance imaging study to demonstrate that the three key elements of the model of cognitive dysmetria--frontal lobe, thalamus, and cerebellum--are reduced in volume in schizophrenic patients. This highlights the importance of this concept for future investigations. PMID- 11099886 TI - Elevated levels of cognitive-perceptual deficits in individuals with a family history of schizophrenia spectrum disorders. AB - This study finds that the relatives of schizophrenics have elevated scores on the cognitive-perceptual factor of the schizotypal personality questionnaire (SPQ), particularly for the 'unusual perceptual experiences' and 'ideas of reference' subscales. These results support recent findings by Kremen et al. (1998) and suggest that previous failures to demonstrate elevated scores on 'positive' symptoms of schizotypy may be a function of instrumentation. PMID- 11099887 TI - The 10th Biennial Winter Workshop on Schizophrenia. A report. AB - The 10th Biennial Winter Workshop on Schizophrenia (5-11 February 2000), organized by Drs Tim Crow and Steven Hirsch, attracted a wide range of contributions, not only from European countries, but from all across the world, encompassing clinical, neuropsychological, cognitive, experimental, pharmacological and neuroimaging approaches to enhance the understanding of possible causes, features and the treatments of this illness. The Kongresszentrum, Davos, Switzerland was once again the venue. We present a flavour of the variety of presentations, the news and the views emerging from these presentations, and discuss their implications for further advancement in the field of schizophrenia research. PMID- 11099888 TI - The 10th Biennial Winter Workshop on Schizophrenia, Davos, 5-11 February 2000. The genetics of cerebral asymmetry and the structure of language--what's the alternative? An organizer's viewpoint. PMID- 11099890 TI - Editorial. PMID- 11099891 TI - Single automated donor plateletpheresis increases the plasma level of proinflammatory cytokine tumor necrosis factor-alpha which does not associate with endothelial release markers von Willebrand factor and fibronectin. AB - The effect of plateletpheresis on endothelium, which has strong effects on blood coagulation, fibrinolysis and platelet function, is not known. Activation of leukocytes and subsequent generation of proinflammatory cytokines during the extracorporeal circulation may activate the endothelium. To test this hypothesis we measured plasma levels of tumor necrosis factor (TNF)-alpha as a prototype of the proinflammatory cytokines, and von Willebrand factor (vWF) and fibronectin as endothelial release/damage markers before and after a single plateletpheresis procedure on an intermittent-flow machine Haemonetics MCS 3p in 17 healthy donors. We found a significant increase in median plasma level of TNF-alpha following plateletpheresis (3.5 vs 26.5 pg/ml, P=0.02). Such increases in vWF and fibronectin were not observed. The increase in plasma TNF-alpha indicates that a single plateletpheresis procedure causes leukocyte activation which does not seemingly impair endothelial cell function. The relation of plateletpheresis induced proinflammatory cytokine release to some adverse effects observed in both donors and recipients, and the effect of repeated plateletpheresis on endothelium deserve further studies. PMID- 11099889 TI - Treatment of schizoaffective disorder with divalproex sodium. PMID- 11099892 TI - Preoperative autologous blood donation in hip surgeries. AB - We evaluated the effectiveness of preoperative autologous blood donation in reduction of the need for transfusion of homologous blood in hip surgeries at our hospital. The cases of 55 patients who had 67 hip surgeries, including 17 total hip arthroplasties (THA) and 41 rotational acetabular osteotomies (RAO), were studied. The patients predeposited an average of 995 ml of blood for each procedure. The calculated blood loss was an average of 961 ml. Ninety-seven percent of the procedures for which autologous blood had been predeposited were performed without transfusion of homologous blood. In the group for THA, an average of 981 ml of autologous blood was transfused for a blood loss of 1417 ml; hemoglobin levels after operation averaged 103 g/l. From this data, a 1000 ml donation seemed to be an optimal blood deposit for THA and a 800 ml blood deposit seemed sufficient for RAO, where the patients are younger. PMID- 11099893 TI - Primary and secondary haemochromatosis. PMID- 11099894 TI - The importance of non-transferrin bound iron in disorders of iron metabolism. AB - The concept of non-transferrin bound iron (NTBI) was introduced 22 years ago by Hershko et al. (Brit. J. Haematol. 40 (1978) 255). It stemmed from a suspicion that, in iron overloaded patients, the large amounts of excess iron released into the circulation are likely to exceed the serum transferrin (Tf) iron-binding capacity (TIBC), leading to the appearance of various forms of iron not bound to Tf. In accordance with this assumption, NTBI was initially looked for and detected in patients with > or = 100% Tf-saturation. As techniques for its detection became more sophisticated and sensitive, NTBI was also found in conditions where Tf was not fully saturated, leading to a revision of the original view of NTBI as a simple spillover phenomenon. In this review, we will discuss some of the properties of NTBI, methods for its detection, its significance and potential value as an indicator for therapeutic regimens of iron chelation and supplementation. PMID- 11099895 TI - Clinical aspects of hemochromatosis. AB - Hemochromatosis is one of the most frequent genetic diseases among the white populations, affecting one in three hundred persons. Its diagnosis has been radically transformed by the discovery of the HFE gene. In a given individual, the diagnosis can, from now on, be ascertained on the sole association of a plasma transferrin saturation (TS) over 45% and homozygosity for the C282Y mutation. Liver biopsy is only required to search for cirrhosis whenever there is hepatomegaly and/or serum ferritin >1000 ng/ml and/or elevated serum AST. Family screening is mandatory, primarily centered on the siblings. The treatment remains based on venesection therapy which improves many features of the disease (one of the most refractory, however, being the joint signs) and permits normal life expectancy provided the diagnosis is established prior to the development of cirrhosis or of insulin-dependent diabetes. In view of the prevalence, the non invasive diagnosis, the spontaneous severity and the efficacy of a very simple therapy, hemochromatosis should benefit from population screening. This screening could be based, first, on the assessment of transferrin saturation, followed - when elevated - by the search for the C282Y mutation. The discovery of the HFE gene has also paved the road for the individualization of other types of iron overload syndromes which are not HFE-related. PMID- 11099896 TI - The design and properties of 3-hydroxypyridin-4-one iron chelators with high pFe(3+) values. PMID- 11099897 TI - Transfusional iron overload and chelation therapy with deferoxamine and deferiprone (L1). AB - Iron is essential for all living organisms. Under normal conditions there is no regulatory and rapid iron excretion in humans and body iron levels are mainly regulated from the absorption of iron from the gut. Regular blood transfusions in thalassaemia and other chronic refractory anaemias can result in excessive iron deposition in tissues and organs. This excess iron is toxic, resulting in tissue and organ damage and unless it is removed it can be fatal to those chronically transfused. Iron removal in transfusional iron overload is achieved using chelation therapy with the chelating drugs deferoxamine (DF) and deferiprone (L1). Effective chelation therapy in chronically transfused patients can only be achieved if iron chelators can remove sufficient amounts of iron, equivalent to those accumulated in the body from transfusions, maintaining body iron load at a non-toxic level. In order to maintain a negative iron balance, both chelating drugs have to be administered almost daily and at high doses. This form of administration also requires that a chelator has low toxicity, good compliance and low cost. DF has been a life-saving drug for thousands of patients in the last 40 years. It is mostly administered by subcutaneous infusion (40-60 mg/kg, 8 12 h, 5 days per week), is effective in iron removal and has low toxicity. However, less than 10% of the patients requiring iron chelation therapy worldwide are able to receive DF because of its high cost, low compliance and in some cases toxicity. In the last 10 years we have witnessed the emergence of oral chelation therapy, which could potentially change the prognosis of all transfusional iron loaded patients. The only clinically available oral iron chelator is L1, which has so far been taken by over 6000 patients worldwide, in some cases daily for over 10 years, with very promising results. L1 was able to bring patients to a negative iron balance at doses of 50-120 mg/kg/day. It increases urinary iron excretion, decreases serum ferritin levels and reduces liver iron in the majority of chronically transfused iron-loaded patients. Despite earlier concerns of possible increased risk of toxicity, all the toxic side effects of L1 are currently considered reversible, controllable and manageable. These include agranulocytosis (0.6%), musculoskeletal and joint pains (15%), gastrointestinal complaints (6%) and zinc deficiency (1%). The incidence of these toxic side effects could in general be reduced by using lower doses of L1 or combination therapy with DF. Combination therapy could also benefit patients experiencing toxicity with DF and those not responding to either chelator alone. The overall efficacy and toxicity of L1 is comparable to that of DF in both animals and humans. Despite the steady progress in iron chelation therapy with DF and L1, further investigations are required for optimising their use in patients by selecting improved dose protocols, by minimising their toxicity and by identifying new applications in other diseases of iron imbalance. PMID- 11099898 TI - Regulation of intracellular iron levels in iron-acceptor and iron-donor cells. AB - In recent years many new genes and proteins were identified with crucial functions in iron metabolism. This gave an explosion of our knowledge and understanding of iron related disorders. Mutations have been found that are responsible for disturbances in iron transport, leading to either iron overload or iron deficiency. For experts in the field, these new findings clarify the sky and open new routes for exploring hitherto hidden fields of research. For the physician, however, iron metabolism may become even more complicated. In this review, we have tried to assemble all new iron related genes into the context of pathophysiology. Important results from animal experiments, mainly derived from knockout mouse models, are included in this review as they often explain the phenotype of human disease. PMID- 11099899 TI - Free radical formation and oxyhemoglobin oxidation in beta-thalassemic red blood cells in the presence of prooxidants: effects of the free radical scavenger rutin and oral chelator L1. PMID- 11099900 TI - The Sydney Children's Hospital experience with the oral iron chelator deferiprone (L1). PMID- 11099901 TI - Desferrioxamine-chelatable iron (DCI), a component of serum non-transferrin-bound iron (NTBI) used for assessing iron chelation therapy. PMID- 11099902 TI - The effect of deoxynucleosides on cell proliferation of peripheral blood lymphocytes treated with deferoxamine or hydroxyurea. PMID- 11099903 TI - Non-transferrin-bound iron, iron-related oxidative stress and lipid peroxidation in beta-thalassemia intermedia. PMID- 11099904 TI - Multiple effects of iron chelators on molecules controlling cell cycle progression. PMID- 11099905 TI - Combining iron chelators with the nucleoside analog didanosine in anti-HIV therapy. PMID- 11099906 TI - Endocrine problems in ex-thalassemic patients. PMID- 11099907 TI - L1 effects on reactive oxygen (ROS) and nitrogen species (RNS) release, hemoglobin oxidation, low molecular weight antioxidants, and antioxidant enzyme activities in red and white blood cells of thalassemic patients. PMID- 11099909 TI - Using SQUID biomagnetic liver susceptometry in the treatment of thalassemia and other iron loading diseases. PMID- 11099908 TI - Morbidity and mortality of iron intoxication in adult patients with thalassemia major, and effectiveness of chelation. PMID- 11099910 TI - Competition studies of L1-deferiprone with copper and iron. Possible implications on efficacy, toxicity and new therapeutic applications. PMID- 11099911 TI - Haematopoietic stem cell transplantation for the management of haemoglobinopathies in Greek patients. PMID- 11099912 TI - Hydroxamate analog libraries and evaluation of their iron affinity. PMID- 11099913 TI - A relationship between glucose metabolism and NO-mediated iron mobilization from cells. PMID- 11099914 TI - The design and properties of 3-hydroxypyridin-4-one iron chelators with high pFe(3+) values. PMID- 11099915 TI - Post-transfusion thrombocytopenia in recipients with anti-HLA antibody. AB - Allogeneic red cell transfusion produced a significant decrease in the platelet counts of recipients who possessed anti-HLA antibodies. PMID- 11099917 TI - The impact of limited needle and syringe availability programmes on HIV transmission-a case study in Kathmandu. PMID- 11099916 TI - Editorial. PMID- 11099918 TI - The social construction of drug-related death. AB - This article invites you to a social constructionist view on the issue of drug related death. Social constructionism is often misunderstood for denying plain facts. It sure is a fact that there are deadly doses of legal and illegal substances. In this sense it is a truism that drugs kill people. Nonetheless, it is argued that reducing the causes of death to a certain drug as the essential underlying cause of death is a social construction. A case is discussed to demonstrate that a drug-related death can just as well be seen as a free-market death. Free markets kill people at least as much as drugs do. It is argued that drug-related death is a social construction, because attributing a death to a drug is based on unfalsifiable counterfactual thinking. Counterfactual thoughts about what the world would look like if there were no drugs, are seen as expressing one's view of life. PMID- 11099919 TI - A comprehensive package of support to facilitate the treatment of problem drug users in primary care: an evaluation of the training component. AB - Since the early 1980s, government policy documents and specialist reports have encouraged the involvement of general practitioners (GPs) in the treatment of problem drug users. In spite of such policy initiatives, their involvement has been patchy and slow. In response to this apparent reluctance, the London Boroughs of Brent and Harrow established the substance misuse management project (SMP) to support and train GPs in the management of substance misuse. The SMP is a GP-led project that provides ongoing support, shared-care protocol, primary care team training, treatment audits and financial reimbursements. In 1996, the SMP worked with GPs who were not currently involved in treating problem drug users, and those who were providing only minimal interventions. This paper evaluates the training and support given to these GPs and examines changes in their practice. A pre- and post-test survey was undertaken of GP knowledge, attitudes and levels of activity. A structured questionnaire was administered to all GPs before training (n=40) and re-administered between 6 and 9 months following training. SMP audit data were also reviewed to validate any reported changes in practice. All GPs initially reported insufficient knowledge to manage problem drug users. One-fifth were unaware they could prescribe methadone, and nearly half believed drug problems should be treated by specialist services. Post training, the GPs had increased their levels of treatment activity and reported greater confidence and willingness to treat. This study demonstrates the potential to involve GPs in the treatment of problem drug users. The training was part of a package that included ongoing support sessions, team training, audits of treatment and financial reimbursements. It is proposed that, whilst training is a necessary condition, a more comprehensive package of support is needed to facilitate the treatment of problem drug users in primary care. PMID- 11099920 TI - Substance misuse in Russia: a partnership for policy change and service development. AB - The paper provides an account of a joint project of education and training of doctors and nurses in St. Petersburg, funded by the Know How Fund Health Sector Small Partnership Scheme (Russia). Contextual material on substance misuse in Russia is introduced prior to focussing on the situation in St. Petersburg. Reference is made to historical and contemporary material and attention is drawn to the reliability of statistical data on alcohol and drug misuse. The aims of the project and the work carried out are outlined. The response of politicians, policy makers and health care workers to the changes in policy, provision and practice in St. Petersburg are discussed. Evidence from two evaluations is considered - an external evaluation for the project sponsors and an 'insider' evaluation. The latter utilised a framework developed by Cranfield and Stoneman. The paper concludes with a discussion of the tentative nature of education and training programmes, which involve issues regarding the suitability and transferability of educational models and therapeutic skills to a different culture. PMID- 11099921 TI - Doing the possible: harm reduction, injecting drug use and blood borne viral infections in Australia. AB - Most surveys show that, other than among men who inject drugs and have a history of homosexual contact, the prevalence of HIV infection among injecting drug users (IDUs) in Australia is about 2%. Rates of needle sharing have also declined greatly in the last decade, although the high prevalence and incidence of hepatitis C infection suggest that existing strategies have not yet brought this epidemic under control. Harm reduction has been the major Australian approach to the reduction of blood borne viral infections (BBVIs) in IDUs. Harm reduction strategies include needle distribution schemes, drug substitution therapies and education about safe administration practices. Importantly, with IDUs as with gay men, the infected and affected communities have been brought into partnership with health educators, researchers and policy makers. This paper will review Australia's approach to the prevention of BBVI in IDUs and the effectiveness of current strategies. I will argue that while HIV/AIDS among heterosexual IDUs appears to have been successfully prevented, international experiences of rapidly emerging epidemics demonstrate there is little room for complacency. Moreover, reducing the incidence of hepatitis C and hepatitis B among IDUs remains a major challenge. PMID- 11099922 TI - Laboratory study of the effects of citric and ascorbic acids on injections prepared with brown heroin. AB - The addition of acidic substances to brown street heroin to facilitate the solubility of diamorphine in the injection preparation process is commonplace amongst UK injectors. Knowledge of the chemistry behind this process supports the need for this stage in the injection preparation process. It is currently illegal, under the Misuse of Drugs Act 1971, section 9A, to supply acidifiers and other paraphernalia to injectors in the UK. In the current climate of evidence based practice, any consideration given to changing the law would look for evidence to illustrate that the paraphernalia was necessary. Although the theory behind the use of acidifiers suggests they are essential, no previous work using street heroin has actually been reported to illustrate this fact. Anecdotal information has found that drug users are being told by some service providers that the addition of acids is unnecessary. It is important that drugs services give credible information to their clients. The provision of inaccurate information in one area may lead to a lack of trust of all information provided. The small study reported here investigated, under controlled laboratory conditions, the effects of citric and ascorbic acids (vitamin C) on injections prepared with brown heroin, in order to demonstrate the need for acidifiers in the injection preparation process. PMID- 11099923 TI - The sharing of injecting paraphernalia by intravenous drug users (IDUs) within a Worcestershire cohort, with specific reference to water and filters. AB - Britain continues to have a drug misuse health strategy that is HIV led. Because of this, little attention has been paid to other blood-borne viruses such as hepatitis. Moreover, while the provision of needle exchange schemes has been particularly successful in containing the spread of HIV, they have had less impact on the prevalence of hepatitis within IDU cohorts. Thus, it is necessary to understand more about the potential pathways through which the hepatitis viruses can be transmitted. One way of achieving this is to assess the propensity of IDUs to share other items of injecting paraphernalia such as water and filters. In addition, it is useful to gauge the level of opinion with respect to health hazards associated with sharing such items, amongst injecting drug users. This study reports on a small pilot project initiated to assess the degree of sharing of filters and water among 40 needle exchange service users in Worcestershire. Results based on questionnaires show that sharing of water and filters is very high within the sample group. Indeed, only 10% of clients reported never sharing either water or filters. The study also demonstrates that although injectors are aware of the health risks associated with sharing (including hepatitis transmission), they continue to participate in high risk sharing activities. Moreover, the majority of IDUs questioned have a mis conception with respect to the most hygienic sources of water for injecting. For example, only 10% consider sterile water to be the most hygienic source for injecting, with >70% considering tap water in one form or another to be safe. The study is important because it highlights the value of providing sterile water and filters to IDUs to meet their basic and fundamental needs. It is hoped that the findings from this small project will have a wider transferability to other IDU cohorts throughout the UK and beyond. PMID- 11099924 TI - Peer-initiated overdose resuscitation: fellow drug users could be mobilised to implement resuscitation. AB - Research interviews about overdose experiences were conducted with 115 patients attending a methadone maintenance clinic in south London, UK. While almost half (49.6%) reported having experienced overdose personally (on an average of four occasions each), almost all (97.4%) reported that they had witnessed overdoses (on an average of six occasions each). This represents a total of 706 overdoses witnessed, of which 106 had resulted in fatalities. The vast majority of patients (86/97) reported that they had taken actions when they had witnessed overdoses with those acting taking an average of nearly threee different actions on the last occasion on which they had seen someone overdosing. Most respondents reported that they would be willing to act, even if they did not know the overdose victim personally and that they had not been deterred from acting by the previous response from the emergency services. Fear of punishment was not a strong deterrent from acting certainly not for this sample, with many participants also expressing an interest in expanding their repertoire of overdose interventions, for example through training in resuscitation techniques and by keeping naloxone at home for use in overdose emergency. PMID- 11099925 TI - Presentation by scanning electron microscopy of the life cycle of microsporidia of the genus Encephalitozoon. AB - This paper presents, for the first time, documentation by detailed scanning electron microscopy of the life cycle of microsporidia of the genus Encephalitozoon. Phase 1 is represented by the extracellular phase with mature spores liberated by the rupture of host cells. To infect new cells the spores have to discharge their polar filament. Spores with everted tubes show that these are helically coiled. When the polar tubules have started to penetrate into a host cell they are incomplete in length. The infection of a host cell can also be initiated by a phagocytic process of the extruded polar filament into an invagination channel of the host cell membrane. After the penetration process, the tube length is completed by polar tube protein which passes through the tube in the shape of swellings. A completely discharged polar tube with its tip is also shown. The end of a polar tube is normally hidden in the cytoplasm of the host cell. After completion of the tube length the transfer of the sporoplasm occurs and phase 2 starts. Phase 2 is the proliferative phase, or merogony, with the intracellular development of the parasite that cannot be documented by scanning electron microscopy. The subsequent intracellular phase 3, or sporogony, starts when the meronts transform into sporonts, documented as chain-like structures which subdivide into sporoblasts. The sporoblasts finally transform directly into spores which can be seen in their host cell, forming bubble-like swellings in the cell surface. PMID- 11099926 TI - Bacterial and host-derived cationic proteins bind alpha2-laminins and enhance Mycobacterium leprae attachment to human Schwann cells. AB - It has recently been demonstrated that laminin alpha2 chains present on the surface of Schwann cells are involved in the process of attachment of Mycobacterium leprae to these cells. In this study, a protein in the M. leprae cell wall that was found to be capable of binding alpha2-containing laminins (merosin) was isolated and characterized. The M. leprae laminin-binding protein was identified as a 21-kDa histone-like protein (Hlp), a highly conserved cationic protein present in other species of mycobacteria. The gene that encodes this protein was PCR amplified, cloned, and expressed, and the recombinant protein was shown to bind alpha2-laminins. More significantly, when added exogenously, Hlp was able to greatly enhance the attachment of mycobacteria to ST88-14 human Schwann cells. The capacity to bind alpha2-laminins and to enhance mycobacterial adherence to Schwann cells was also found in other cationic proteins such as host-derived histones. Moreover, mutation in the hlp gene was shown not to affect the capacity of mycobacteria to bind to ST88-14 cells, suggesting that alternative adhesins and/or pathways might be used by mycobacteria during the process of adherence to Schwann cells. The potential role of Hlp as a fortuitous virulence factor contributing to the pathogenesis of M. leprae-mediated nerve damage is discussed. PMID- 11099927 TI - Cell-dependent gag mutants of HIV-1 are crucially defective at the stage of uncoating/reverse transcription in non-permissive cells. AB - We have previously shown that some of the human immunodeficiency virus type 1 (HIV-1) gag matrix (MA), capsid (CA), and nucleocapsid (NC) mutants display host cell-dependent replication potential, and that they are defective at the early phase of the virus replication cycle in non-permissive cells. To determine the defective replication stage of the cell-dependent mutants precisely, the processes of virus entry into cells and virus DNA synthesis were monitored by the highly sensitive enzyme-linked immunosorbent assay and polymerase chain reaction amplification analysis. The results obtained indicated that all the cell dependent MA, CA and NC mutants are defective at the stage of uncoating/reverse transcription, and that a cellular factor(s) is involved in this process. PMID- 11099928 TI - The virulence of mixed infection with Streptococcus constellatus and Fusobacterium nucleatum in a murine orofacial infection model. AB - Orofacial infections are usually polymicrobial, and it is the microbial interactions of pathogenic species that cause tissue destruction. In this study, the microbial interaction between Streptococcus constellatus and Fusobacterium nucleatum was characterized using a murine orofacial infection model. A mixture of viable S. constellatus and F. nucleatum cells (both 2 x 10(8) CFU/mouse) was injected into the submandible; as a result, all of the test mice died. In contrast, none of the experimental animals monoinjected with either S. constellatus or F. nucleatum died (P<0.001), indicating that the synergism between the two resulted in the virulence. When a mixture of viable S. constellatus cells and a culture filtrate of F. nucleatum was tested, lethality and the bacterial cell count per lesion were significantly enhanced as compared with monoinjections (P<0.02). However, the virulence of F. nucleatum was not enhanced by infection of a culture filtrate of S. constellatus. The enhancement of virulence was observed even when viable S. constellatus cells and the culture filtrate of F. nucleatum were injected at separate sites. Heat treatment of the culture filtrate of F. nucleatum did not affect the enhancement. These results indicate that a heat-stable substance(s) produced by F. nucleatum contributes to the microbial synergy of S. constellatus and F. nucleatum in orofacial infections. PMID- 11099929 TI - Detection of serum antibodies to Bartonella henselae and Coxiella burnetii from Japanese children and pregnant women. AB - The participation of Bartonella henselae and Coxiella burnetii in the pathogenesis of fever of unknown origin (FUO) and lymphadenopathy has not been completely clarified. Prevalence of these two agents in Japanese children is also unknown. Serum IgG and IgM antibodies to B. henselae and to C. burnetii were examined by the indirect fluorescence antibody assay. Enzyme immunoassay kits were used to detect serum IgG and IgA antibodies against Chlamydia trachomatis. Out of 200 healthy normal pregnant women, two (1.0%) had serum IgG antibodies to B. henselae, four (2.0%) to C. burnetii and 49 (24.5%) to C. trachomatis. Out of 29 patients with FUO, one (3.4%) had serum IgG antibodies to B. henselae, four (13.8%) to C. burnetii and none to C. trachomatis. Out of 31 patients with cervical lymphadenopathy, three (9.6%) had serum IgG antibodies to B. henselae, two (6.5%) to C. burnetii and none to C. trachomatis. Out of 22 patients with generalized lymphadenopathy, one (4.5%) had serum IgG antibodies to B. henselae, three (13.6%) to C. burnetii and none to C. trachomatis. Prevalences of serum antibodies to C. burnetii in the patients with FUO and generalized lymphadenopathy and to B. henselae in the patients with cervical lymphadenopathy were significantly higher than those of normal pregnant women (Welch's t-test; P<0.01). These two agents may have some roles in the pathogenesis of FUO and lymphadenopathy in Japanese children. PMID- 11099930 TI - Different cytokines are required for induction and maintenance of the Th2 type response in DBA/2 mice resistant to infection with Leishmania major. AB - Experimental cutaneous leishmaniasis is a useful model in studying the mechanism regulating immune responses between T helper type 1 (Th1) and Th2. Mice susceptible to Leishmania major infection such as BALB/c (H-2(d)) are associated with the induction of the disease-promoting Th2 response, while the resistant mice such as DBA/2 (H-2(d)) develop the protective Th1 response. To understand the induction mechanism of Th1 and Th2 responses, it is necessary to establish an immunization scheme by which the induction of each Th response can be easily and experimentally controlled. Adjuvants are known to enhance the immune responses through the combined effect of several factors: prolonged release of antigen, migration of cells, mitogenic effect and so forth. When the genetically resistant DBA/2 mice were immunized twice with soluble leishmanial antigen (SLA), emulsified in incomplete Freund's adjuvant (IFA) before L. major inoculation, these mice mounted a Th2 cell response and suffered from progressive infection. While IL-4 and IL-13 were upregulated early after the infection in both healer and non-healer groups of mice, IL-5 and IL-10 were upregulated only in non-healer mice. From these results, IL-5 and IL-10 appear to have an important role, at least in the early phases of the infection, rather than IL-4 and IL-13 in establishing the disease-promoting Th2 response in leishmaniasis. Further, IL-9 was found to be expressed in both BALB/c and DBA/2 mice immunized with IFA/SLA. This cytokine may support the establishment of a Th2 response in these mice. Therefore it is suggested that Th2 cytokines play different roles between priming and maintaining the Th2 immune response after the infection. PMID- 11099931 TI - Prion disease--the propagation of infectious protein topologies. AB - Prion propagation is associated with accumulation of a conformational isomer of host encoded cellular prion protein, PrP(C). Solution structures of several mammalian PrPs have now been reported and they have stimulated a significant advance in our understanding of the folding dynamics of PrP. Studies on recombinant PrP have shown the polypeptide chain is able to adopt different topologies in different solvent conditions. Concomitantly, advances in the analysis of the abnormal isoform, PrP(Sc), have expanded our knowledge on the molecular basis of prion strains and have done much to reinforce the protein-only hypothesis of prion replication. PMID- 11099932 TI - How to survive in the host: the Yersinia lesson. AB - The Yop virulon allows Yersinia spp. to resist the immune response of the host by injecting harmful proteins into host cells. It is composed of four elements: (i) type III secretion machinery called Ysc; (ii) a set of proteins required to translocate the effector proteins inside the eukaryotic cells; (iii) a control system, and (iv) six Yop effector proteins. PMID- 11099933 TI - The apical organelles of malaria merozoites: host cell selection, invasion, host immunity and immune evasion. AB - Malaria is caused by protozoan parasites belonging to the phylum Apicomplexa. These obligate intracellular parasites depend on the successful invasion of an appropriate host cell for their survival. This article is a broad overview of the molecular strategies employed by the merozoite, an invasive form of the malaria parasite, to successfully invade a suitable red blood cell. PMID- 11099934 TI - Assessing the impact of Plasmodium falciparum genome sequencing. AB - With the publication of the complete sequences for chromosomes 2 and 3 and the increasing availability of shotgun sequence covering most of its genome, Plasmodium falciparum biology is entering its post-genomic era. Analysis of the results generated to date has identified higher-order organisation of gene families involved in parasite pathology, provided information regarding the unique biology of this parasite and allowed the identification of potential chemotherapeutic drug targets. Continuing efforts to complete the P. falciparum genome and the availability of sequences from other protozoan parasites will facilitate a broader understanding of their biology, particularly with respect to their pathogenicity. PMID- 11099935 TI - Molecular mimicry between HLA-DR alleles associated with rheumatoid arthritis and Proteus mirabilis as the Aetiological basis for autoimmunity. AB - Molecular mimicry is one of the pathological mechanisms proposed to explain the association between microorganisms and autoimmune diseases. This review deals with the association between bacteria and rheumatic diseases with a special emphasis on rheumatoid arthritis where upper urinary tract infection by Proteus mirabilis is the possible cause of this severe, arthritic condition. Prospective trials involving anti-Proteus therapy should be carried out. PMID- 11099936 TI - Pathogenesis of Proteus mirabilis urinary tract infection. AB - Proteus mirabilis is a causative agent of cystitis and pyelonephritis primarily in individuals with indwelling catheters or structural abnormalities of the urinary tract. The organism produces a variety of unique virulence factors that contribute to its pathogenicity and persistence in the human host. PMID- 11099937 TI - Cell surface components and adhesion in Corynebacterium diphtheriae. AB - Main primary approaches and new developments in the study of the molecular basis of the adhesive process of Corynebacterium diphtheriae are reviewed along with a discussion of the potential importance of hemagglutinins, exposed sugar residues, hydrophobins and trans-sialidase enzymes as adhesins of strains of the sucrose fermenting and non-fermenting biotypes. PMID- 11099938 TI - Evolutionary changes in mutation rates and spectra and their influence on the adaptation of pathogens. AB - The evolutionary tuning of mutational processes may play a key role in prokaryotic evolution, particularly among pathogens. In this paper we review the evidence that genetic systems controlling the rate and spectrum of heritable mutations have evolved to optimize levels of adaptive variation and rates of genetic change. PMID- 11099939 TI - Use of fluorescent probes to assess physiological functions of bacteria at single cell level. AB - A wide diversity of fluorescent probes is currently available to assess the physiological state of microorganisms. The recent development of techniques such as solid-phase cytometry, the increasing sensitivity of fluorescence tools and multiparametric approaches combining taxonomic and physiological probes have improved the effectiveness of direct methods in environmental and industrial microbiology. PMID- 11099940 TI - 'Infectious web'. AB - Exhaustive information on the Epstein-Barr virus, a member of the herpes family, is described at the International Herpes Management Forum web-site. Cervical cancer associations, AIDS treatment projects, and the Los Alamos National Laboratories provide useful information on papillomavirus infections, as well as hyperlinks to recent international papillomavirus conferences. A private pharmaceutical company, in collaboration with the National Institutes of Health, has launched a lively web-site covering different aspects of microbial infections for the general public. PMID- 11099941 TI - Forty winks: molecular basis of sleep disorders. PMID- 11099942 TI - Reference standard for gene therapy closer. PMID- 11099943 TI - Poliovirus replicons for targeting the CNS. PMID- 11099945 TI - A new gene involved in mental retardation? PMID- 11099944 TI - An early start to ill health. PMID- 11099946 TI - Polyglutamine stretches suppress transcription PMID- 11099948 TI - Chemoprevention of intestinal neoplasia PMID- 11099947 TI - Synergism: bioinformatics and biological experimentation PMID- 11099950 TI - A new gene transfer vector for the lung PMID- 11099949 TI - Overexpression of CDC25A associated with poor prognosis in breast cancer PMID- 11099951 TI - The adenomatous polyposis coli (APC) tumour suppressor--genetics, function and disease. AB - Mutations in the adenomatous polyposis coli (APC) gene are the basis of familial adenomatous polyposis and the majority of sporadic colorectal cancer. APC is expressed in a wide variety of tissues, interacts with the cytoskeleton, is involved in regulating levels of beta-catenin and, most recently, has been shown to bind DNA, suggesting that it may possess a nuclear role. The mutation spectrum implicated in tumorigenesis and its correlation with disease phenotype is well characterized and has contributed to our understanding of important functional domains in APC. Despite these advances, APC continues to provide a fertile subject of research for both colorectal tumorigenesis and cancer in general. PMID- 11099952 TI - Mining bacterial genomes for antimicrobial targets. AB - The elucidation of whole-genome sequences is expected to have a revolutionary impact on the discovery of novel medicines. With the availability of complete genome sequences of more than 30 different species, the field of antimicrobial drug discovery has the opportunity to access a remarkable diversity of genomic information. In this review, I summarize how microbial genomics has changed strategies of drug discovery by applying bioinformatics, novel genetic approaches and genomics-based technologies, including analysis of gene expression using DNA microarrays. PMID- 11099953 TI - Genetics of drug response to immunosuppressive treatment and prospects for personalized therapy. AB - The use of immunosuppressive agents in the treatment of transplant rejection and autoimmune disorders is gaining momentum, with significant improvements of both graft and patient survival. The individual response to drugs, however, is variable and unexpected toxicity, or impaired activity might be seen, as a result of molecular determinants that eventually dictate how the individual will respond to immunosuppressive agents. This review addresses a number of issues related to pharmacogenetics, and discusses how this approach might be used to improve the clinical efficacy and tolerability of therapeutic options for the management of organ transplantation and autoimmune disorders in the next decade. PMID- 11099954 TI - Enteric viruses in HIV-related diarrhoea. AB - HIV-related diarrhoea is an important cause of morbidity and mortality in HIV infection. Cytomegalovirus is a well-established cause of diarrhoea, but the role of other enteric viruses is less clear and will be discussed here. The clinical manifestations, disease mechanisms, diagnostic techniques and current treatments for the management of these infections are reviewed. PMID- 11099955 TI - Murine gamma-herpesvirus-68: a mouse model for infectious mononucleosis? PMID- 11099956 TI - X-ray structure determination of the monoclinic (121 K) and orthorhombic (85 K) phases of langbeinite-type dithallium dicadmium sulfate AB - The structures of the monoclinic and the orthorhombic phases of type I langbeinite Tl(2)Cd(2)(SO(4))(3) have been determined at 121 and 85 K, respectively, by X-ray diffraction. A precise analysis of these structures shows the existence of some differences compared to langbeinites of type II. The monoclinic structure differs very little from the high-temperature cubic structure and the distortion relating the monoclinic structure to the cubic one is very small. SO(4) tetrahedra seem to rotate under orthorhombic symmetry in the monoclinic phase. A symmetry distortion analysis of the ferroelectric monoclinic distortion discloses the importance of the secondary modes with orthorhombic symmetry, especially for the O atoms of the SO(4) groups. PMID- 11099957 TI - Application of graph theory to detect disconnected structures in a crystallographic database: copper oxide perovskites as a case study AB - Every crystal structure can be described as a graph with atoms as vertices and bonds as edges. Although such a graph loses the space arrangement of atoms and symmetry elements, it can mathematically represent the connectivity between atoms. This topological approach was used to develop a new method for detecting disconnected structures, in which individual atoms or structural fragments are located too far from each other, forming impossibly large gaps. Approximately 2300 perovskite-related crystal structures have been extracted from the Inorganic Crystal Structure Database (in 1999) and the maximum disconnecting distances, and the relations between them and the ionic radii of elements, have been analysed. Several disconnected structures, which are erroneous by our definition, have been revealed. Conventional tests for crystallographic data checking did not detect those entries. PMID- 11099958 TI - Structure and complex twinning of dysprosium disilicate (Dy2Si2O7), type B AB - Dysprosium disilicate (Dy(2)Si(2)O(7)) is triclinic with a = 6.6158 (2), b = 6.6604 (2), c = 12.0551 (4) A, alpha = 94.373 (2), beta = 90.836 (2), gamma = 91.512 (2) degrees, V = 529.4 (1) A(3), space group P1;, Z = 4 and D(x) = 6.156 g cm(-3). The structure (single-crystal X-ray, R = 0.033, wR = 0.041) is built from a linear triple tetrahedral group [Si(3)O(10)] and isolated [SiO(4)] tetrahedron cross-linked by Dy(3+) in one sixfold and three eightfold coordinated positions, and corresponds to the presently revised type B structure of Ho(2)Si(2)O(7). The formation of the unusual linear triple tetrahedral group in the type B structure allows for a more continuous transition in the mean size of REE(3+)O(n) (REE = rare earth element) polyhedra in REE disilicates through the 4f transition metal series. The crystal of Dy(2)Si(2)O(7) investigated was complexly twinned such that the diffraction pattern was also consistent with a larger dimensionally monoclinic unit cell (a = 22.5354, b = 14.2102, c = 6.6158 A, beta = 91.788 degrees ), which resulted in an apparent superstructure of the type B structure in space group C1;. Lattice coincidence with the type B unit cell appears to have been maintained during crystal synthesis and quenching by the complex sector zoned growth twin. PMID- 11099959 TI - Localized defects in radiation-damaged zircon AB - The crystal structure of a radiation-damaged natural zircon, ZrSiO(4) (alpha decay radiation dose is ca 1.8 x 10(18) alpha-decay events g(-1)), has been determined. The anisotropic unit-cell swelling observed in the early stages of the amorphization process (0.17% along the a axis and 0.62% along the c axis compared with the undamaged material) is a consequence of the anisotropy of the expansion of ZrO(8) polyhedra. Larger anisotropic displacement parameters were found for Zr and O atoms, indicating that the distortion produced by alpha particle-induced localized defects mainly affects the ZrO(8) unit. The overall shape of SiO(4) tetrahedra remains essentially undistorted, while Si-O bonds are found to lengthen by 0.43%. PMID- 11099961 TI - Modulated structure of La2Co1.7 from neutron and X-ray diffraction data AB - An La(2)Co(1.7) crystal was investigated by single-crystal neutron and X-ray diffraction. The neutron measurement was performed with a Laue white-beam technique at 15 K and room temperature, using a large position-sensitive detector. The X-ray measurements were obtained at room temperature from a CCD detector. The average structure of La(2)Co(1.7) is hexagonal with cell parameters a = 4. 885 (1), c = 4.273 (2) A and space group P6(3)/mmc. The satellites are located at the vertices of small hexagons perpendicular to the c axis. The modulated crystal was indexed assuming a sixfold twinned 3 + 1 dimensional structure with q = (alpha, 0, gamma). The structure was solved in the pseudoorthorhombic cell, with a = 8.461 (1), b = 4. 885 (1), c = 4.273 (2) A, in the superspace group C2/m(alpha, 0, gamma). Owing to space requirements, the Co atoms cannot fit precisely into the octahedral sites of the La h.c.p. (hexagonal close packing). Instead, the Co atoms adopt a different periodicity, which is not commensurate with the periodicity of the La atoms. Two structure models have been refined in order to describe this behaviour, one using the sawtooth function for the positional modulation of cobalt and the other describing the structure as a composite system. The chemical composition calculated from the composite model is La(2)Co(1.8 (1)) with the estimated standard deviation arising from the variation of q for different samples. In both models lanthanum is incommensurately modulated, while the position of cobalt seems not to be affected by any relative periodic displacement. PMID- 11099960 TI - Structure of the beta form of calcium pyrophosphate tetrahydrate. AB - beta-Ca(2)P(2)O(7).4H(2)O is monoclinic, P2(1)/c, a = 12.287 (6), b = 7.511 (3), c = 10.775 (5) A, and beta = 112.54 (1) degrees. Five of the terminal O atoms from a pyrophosphate group bind to Ca atoms, together with O atoms from three of the water molecules. The fourth H(2)O forms only hydrogen bonds. Both Ca atoms have coordination number 7 and show characteristics between those of a capped octahedron and a pentagonal bipyramid. The analysis of coordination distortions suggests that regularity and volume efficiency of a Ca coordination polyhedron increases with the number of bound water O atoms. The structure is layered after ?100?, reflected also in the morphology of crystals which are formed as extremely thin plates. The central parts of the layers are formed by chains of Ca coordination polyhedra which run along the b axis and are interconnected by pyrophosphate groups. Water molecules form the surfaces of the layers. A peculiar auto-inhibition of growth from supersaturated solutions is supposed to be caused by a direct attachment of CaP(2)O(7)(2-) and P(2)O(7)(4-) to the water molecules on the surfaces of layers. Ca(2)P(2)O(7).4H(2)O is known in two polymorphs. The unit-cell volume of the beta form compared with that of alpha suggests that the former is a low-temperature modification. PMID- 11099962 TI - Structures and phase transitions of the A7PSe6 (A = ag, Cu) argyrodite-type ionic conductors. III. alpha-Cu7PSe6 AB - The crystal structure of the third polymorph of the Cu(7)PSe(6) argyrodite compound, alpha-Cu(7)PSe(6), heptacopper phosphorus hexaselenide, is determined by means of single-crystal diffraction from twinned crystals and X-ray powder diffraction, with the help of extensive NMR measurements. In the low-temperature form, i.e. below the last phase transition, alpha-Cu(7)PSe(6) crystallizes in orthorhombic symmetry, space group Pna2(1), with a = 14.3179 (4), b = 7.1112 (2), c = 10.1023 (3) A, V = 1028.590 (9) A(3) (deduced from powder data, T = 173 K) and Z = 4. Taking into account a twinning by reticular merohedry, the refinement of the alpha-Cu(7)PSe(6) structure leads to the residual factors R = 0.0466 and wR = 0.0486 for 127 parameters and 3714 observed, independent reflections (single crystal data, T = 173 K). A full localization of the Cu(+)d(10) element is reached with one twofold-, one threefold- and five fourfold-coordinated Cu atoms. The observation of two phase transitions for Cu(7)PSe(6), to be compared with only one for Ag(7)PSe(6), is attributed to the d(10) element stability in a low coordination environment, copper being less prone to lower coordination sites than silver, especially at low temperature. PMID- 11099963 TI - Dopant positions in strontium/chromium- and barium-doped KTP, determined with synchrotron X-radiation AB - Structure factors for strontium/chromium- (Sr/Cr) and barium- (Ba) doped potassium titanyl phosphate (KTiOPO(4), KTP) were measured with focused synchrotron X-radiation [0.75000 (9) A] using a fast avalanche photodiode counter. Space group Pna2(1), Z = 8, a = 12.786 (2), b = 6.3927 (8), c = 10.5585 (9) A, T = 293 (1) K, R = 0.028 (SrCrKTP); a = 12.851 (6), b = 6.418 (3), c = 10.620 (5) A, T = 120 (1) K, R = 0.031 (BaKTP). The refinement of the dopant positions showed that Ba(2+) is positioned in the larger of the two K cavities of KTP, while the smaller Sr(2+) ion is located in both. Split positions are found for the strontium dopant in both cavities and they are located in the positive c direction from the potassium cation. The chromium dopant has two different oxidation states, namely +III and +VI; in both states the dopant is located inside the TiO(6) octahedra. The two structures show slightly less distorted TiO(6) octahedra than pure KTP. PMID- 11099964 TI - Anharmonicity of potentials of atoms in potassium hydrogensulfide (KDS) determined by neutron single-crystal diffraction AB - Potassium hydrogensulfide (KHS) is an ionic compound with an anionic molecular group HS(-). The fast reorientational disorder of the anions was determined for the ambient temperature modification [R3;m; Jeffrey (1974). Can. J. Phys. 52, 2370-2378]. Single crystals are available now as protonated or deuterated specimens. With neutron single-crystal diffraction at room temperature, a considerable anharmonicity of the atom potential of the H or D atoms was observed. Even the thermal motions of K and S atoms show small deviations from an isotropic probability density function, which can be modelled using anharmonic temperature factors. The temperature factors of the atoms were expanded into a Gram-Charlier series [Kuhs (1992). Acta Cryst. A48, 80-98] in order to evaluate the anharmonicity quantitatively. Parameters up to a fourth-order approximation are relevant for the D atoms. Results from neutron single-crystal diffraction are compared with split-atom models extracted from neutron powder diffraction patterns of fully deuterated samples. PMID- 11099965 TI - X-ray diffraction study of copper(I) thiourea complexes formed in sulfate containing acid solutions AB - The formation of three different copper(I) thiourea complexes in sulfate containing acid solutions was observed. The ratio between Cu(I) and thiourea (tu) in these complexes depends on the amount of thiourea and copper sulfate in the solution. The crystal and molecular structure of a new complex, [Cu(2)(tu)(6)](SO(4)).H(2)O, was determined, and the formation and structures of [Cu(2)(tu)(5)](SO(4)).3H(2)O and [Cu(4)(tu)(7)](SO(4))(2).H(2)O were confirmed. The compound [Cu(2)(tu)(6)](SO(4)).H(2)O crystallizes in the P1; space group, with a = 11.079 (2), b = 11.262 (1), c = 12.195 (2) A, alpha = 64.84 (1), beta = 76.12 (1), gamma = 66.06 (1) degrees, and Z = 2. The Cu-thiourea complex is arranged as a Cu(I) tetranuclear ion, [Cu(4)(tu)(12)](4+), sited on a crystallographic inversion center. All copper ions are in a tetrahedral coordination with thiourea ligands and located at alternate sites on an eight membered, crown-like ring. PMID- 11099966 TI - Powder and single-crystal X-ray diffraction study of the structure of AB - From powder pattern indexing it has been demonstrated that [Y(H(2)O)](2)(C(2)O(4))(CO(3))(2), yttrium oxalate carbonate, crystallizes with orthorhombic symmetry, space group C222(1), a = 7. 8177 (7), b = 14.943 (1), c = 9.4845 (7) A, V = 1108.0 (1) A(3), Z = 4. This unit cell displays a doubling of the c parameter, arising from weak diffraction lines observed in the powder diffraction pattern, with respect to results reported in the literature. The crystal structure has been solved ab initio using direct methods from powder data and has been confirmed by additional single-crystal data collected with a CCD area detector. The overall crystal structure is similar for both unit cells, except that an alternation of the carbonate groups in the direction parallel to the screw axis is displayed in the larger cell, while with the suggested half unit cell (space group C2mm) the carbonate groups would show only one orientation. The unit-cell determination strategy from single-crystal diffraction, collected with Nonius CAD-4 and Nonius Kappa CCD diffractometers, is discussed with respect to the results extracted from the powder diffraction pattern. The study demonstrates the power and usefulness of the full trace of a powder pattern for the detection of subtle structure details. PMID- 11099967 TI - Orientational disorder as a function of temperature in the clathrate structure of hydroquinone and C60 AB - In the compound [C(6)H(6)O(2)](3)C(60), hydroquinone (C(6)H(6)O(2)) forms a three dimensional hydrogen-bonded network enclosing roughly spherical cages with point symmetry 3;m and a diameter of 13.2 A at 100 K. Although C(60) fits tightly into these cages, it shows threefold orientational disorder, the molecular site symmetry being 2/m. The disorder has been studied with single-crystal Mo Kalpha X ray data at four temperatures, 100, 200, 293 and 373 K. In the refinement, C(60) was restrained to the icosahedral molecular symmetry m3;5; and to rigid-body translational and librational displacements including third- and fourth-order cumulants to account for curvilinear atomic movements, R(|F|) = 3.2-4.7%. At 100 K, bond lengths in C(60) refine to the expected values 1.450 (1) and 1.388 (1) A. The ratio of these values increases with increasing temperature, but the radius of the molecule remains constant at 3. 537 (2) A. The r.m.s. libration amplitudes of C(60) are relatively small (5.5 degrees at 373 K) and the probability function of orientations of C(60) inside the cage shows large values only at the refined positions, indicating that the energy barrier of reorientation is large. Refinement of an ordered twinned structure was unsuccessful; the orientations of neighboring C(60) appear to be uncorrelated. PMID- 11099968 TI - Crystallographic and molecular mechanics investigation of an order-disorder transition and dimorphism in 5H,10H-dithiolo AB - Single-crystal X-ray structures are presented for three forms of 5H, 10H dithiolo[2,3-b]-2,5-benzodithiocine-2-thione. The alpha (at 150 K) and alpha' (at ambient) forms are very similar and differ only in the presence of crystallographic m symmetry in the molecules of alpha', which is absent in the case of alpha. This pair is related by an order-disorder transition. The beta phase (also determined at 150 K) has a different structure in terms of the molecular packing from either of the other two and therefore constitutes a true polymorph. Molecular mechanics calculations indicated that the most stable CHCl(3)-solvated conformations for the title compound were a pair of twisted U shaped enantiomers, U(R) and U(L), i.e. similar to the arrangements found in the alpha and beta phases, with the low-lying saddle point between them corresponding to the situation in the alpha' phase. These calculations also indicated that the most stable CHCl(3)-solvated conformation for the related dibromo-5H, 10H dithiolo[2,3-b]-2,5-benzodithiocine-2-thione was Z-shaped, in agreement with the crystal structure determined earlier for its DMSO solvate [Wang et al. (1998). Synthesis, pp. 1615-1618]. PMID- 11099970 TI - Structural characterization of protonated benzeneseleninic acid, the dihydroxyselenonium ion AB - The structure of the dihydroxyphenylselenonium ion (C(6)H(7)O(2)Se(+)) has been determined in its benzenesulfonate (C(6)H(5)O(3)Se(-)-) and p-toluenesulfonate (C(7)H(7)O(3)S(-)) salts. Whereas the former salt is disordered, the latter less dense salt is well defined. This difference in crystallization behaviour is attributed to a C-H.O hydrogen bond involving the methyl group of the p toluenesulfonate ion. The two salts display very similar hydrogen-bond arrangements and differ only with respect to the stacking of the phenyl groups. The dihydroxyselenonium ion is a strong acid with a pK value of -0.9 determined from the variation of the (77)Se chemical shift. A comparison with the two deprotonated species reveals a systematic increase in the Se-O bond lengths and the pyramidal configuration around Se with the number of protons attached. PMID- 11099969 TI - Supramolecular structure of 1H-pyrazoles in the solid state: a crystallographic and ab initio study. AB - The secondary structure of 1H-unsubstituted pyrazole derivatives bearing only one hydrogen donor group and one or more acceptor groups has been analyzed in terms of some descriptors representing the substituents at C3 and C5. The substituent at C4 appears to affect mainly the tertiary or quaternary structure of these compounds. The proposed semi-quantitative model, which explains most hydrogen bonded motifs as a combination of the effects of substituents at C3 and C5, has also been examined as a function of the steric and polarizability effects of these substituents represented by molar refractivity. The model also applies to other five-membered rings (1,2,4-triazoles, 1,2,4-diazaphospholes and 1,2, 4 diazaarsoles). Furthermore, ab initio calculations at RHF/6-31G* have been performed to discover the relative stability of three of the four hydrogen-bond patterns displayed by several symmetrical pyrazoles (dimers, trimers, tetramers). The fourth motif, catemers, has only been discussed geometrically. PMID- 11099971 TI - Topology of molecular packings in organic crystals AB - Using the methods of coordination sequences and of molecular Voronoi-Dirichlet polyhedra, the topological properties of molecular packings and molecular coordination numbers (MCNs) were determined in the crystal structures of 33 575 monosystem organic compounds within the first three coordination spheres. Numerous examples of disagreement between the topology of molecular packing and the system of intermolecular contacts in a crystal structure were found. It is concluded that within the first coordination sphere most of the molecules tend to arrange with MCN = 14, obeying the model of the thinnest covering of space, but molecular packings as a whole tend to be constructed according to one of the close packings. PMID- 11099972 TI - Comparison of the structural motifs of acetoacetanilides and related azo pigments AB - The structures of three methyl-substituted acetoacetanilides and of an azo pigment derived from one of them are presented and discussed together with a review of related known crystal structures. By considering the position of any aromatic substituents it is possible to predict whether the simple acetoacetanilides adopt planar structures with intramolecular hydrogen bonding or twisted structures featuring intermolecular hydrogen bonding. However, we find that the same crystal engineering rules cannot be applied to the related azo pigments: this is apparently due to the presence of an sp(2) atom which facilitates the adoption of planar conformations. The thermal properties of the acetoacetanilides were measured by DSC and are discussed with reference to their crystal structures. PMID- 11099973 TI - Adducts of meso and racemic 5,5,7,12,12,14-hexamethyl-1,4,8, 11 tetraazacyclotetradecane with trigonally trisubstituted benzene carboxylic acids: supramolecular structures in one and two dimensions AB - The meso and racemic forms of 5,5,7,12,12,14-hexamethyl-1,4,8, 11 tetraazacyclotetradecane, C(16)H(36)N(4) (tet-a and tet-b, respectively), form adducts with trigonally trisubstituted benzene carboxylic acids; tet-a-3,5 dinitrobenzoic acid (1/2) (1), tet-a-5-hydroxyisophthalic acid-water (1/1/1) (3) and tet-b-5-hydroxyisophthalic acid-water (1/1/1) (4) are all salts, [C(16)H(38)N(4)](2+).2[C(7)H(3)N(2)O(6)](-) (1) and [C(16)H(38)N(4)](2+).[C(8)H(4)O(5)](2-).H(2)O (3) and (4). The conformations of the [(tet-a)H(2)](2+) and [(tet-b)H(2)](2+) cations are entirely different: [(tet a)H(2)](2+) is precisely centrosymmetric in (1) and approximately so in (3), while [(tet-b)H(2)](2+) has approximate C(2) symmetry in (4). In each salt the cation forms two intramolecular N-H.N and four intermolecular N-H.O hydrogen bonds. In (1) the supramolecular structure is one-dimensional, a C(2)(2)(13)[R(2)(4)(16)] chain of rings. Compounds (3) and (4) crystallize in space groups P2(1)2(1)2(1) and P2(1)/c, respectively, but the supramolecular structures are very similar: in each, the anions and the water molecules form a C(7)[R(3)(3)(13)] chain of rings, generated in (3) by a 2(1) axis and in (4) by a glide plane. These chains are linked, in both (3) and (4), by cations to form sheets. Adjacent meso cations in (3) are related by a 2(1) axis and adjacent chiral cations in (4) are related by a glide plane. PMID- 11099974 TI - Crystal engineering in the gem-alkynol family: interplay between strong and weak interactions in structures of 2,3,5,6-tetrahalo AB - Structures of the title compounds are all mediated by strong cooperative arrangements of O-H.O hydrogen bonds, supported by a variety of weaker interactions which affect the type of O-H.O synthon that is formed. The tetrafluoro compound contains hexameric O-H.O synthons in a supramolecular chair conformation, together with C identical withC-H.F interactions. However, the tetrachloro and tetrabromo compounds both form tetrameric O-H.O synthons. This dominant pattern is supported by halogen.halogen interactions having one C Cl[Br].Cl[Br] angle close to 180 degrees and the other close to 90 degrees, and by C identical withC-H.Cl[Br] interactions. PMID- 11099975 TI - Crystal engineering in the gem-alkynol family; synthon repetitivity and topological similarity in diphenylethynylmethanols: structures that lack O-H.O hydrogen bonds AB - The structures of four para-substituted derivatives of diphenylethynylmethanol have been determined [ditolylethynylmethanol, di(4-chlorophenyl)ethynylmethanol, di(4-bromophenyl)ethynylmethanol and bis(4,4'-biphenylyl)ethynylmethanol]. The dimethyl, dichloro, dibromo and diphenyl compounds have been analysed using X-ray diffraction at 150 K, and the dichloro compound has also been studied using neutron diffraction at 150 K. In common with the parent diphenylethynylmethanol [Garcia, Ramos, Rodriguez & Fronczek (1995). Acta Cryst. C51, 2674-2676], all four derivatives fail to form the expected strong O-H.O hydrogen bonds due to steric hindrance. Instead, the supramolecular structural organization in this family of gem-alkynols is mediated by a variety of weaker interactions. The two most acidic protons, O-H and C[triple-bond]C-H, participate in weak hydrogen bonds to pi-acceptors, forming synthons that stabilize all five structures. These primary interactions are reinforced by a variety of other weak hydrogen bonds involving C-H donors and the hydroxy-O as an acceptor, and by halogen.halogen interactions in the dichloro and dibromo compounds. PMID- 11099976 TI - Halogen trimer synthons in crystal engineering: low-temperature X-ray and neutron diffraction study of the 1:1 complex of 2,4, 6-tris(4-chlorophenoxy)-1,3,5 triazine with tribromobenzene AB - The title complex has been studied using low-temperature X-ray (150 K) and neutron (100 K) diffraction. Molecules of the triazine host form a two dimensional hexagonal network mediated by trigonally symmetric Cl(3) synthons having Cl.Cl interactions of 3.441 (3) A, a C-Cl.Cl angle of 165 degrees and a Cl.Cl-C angle of 105 degrees, close to the ideal values of 180 and 90 degrees, respectively. The guest molecules are of an appropriate size to fit the hexagonal networks and interact with the host via C-H.pi (phenyl) and C-Br.pi (phenyl) interactions which stabilize the overall structure. Both C-donor bond vectors are directed more closely towards the mid-point (X) of an individual aromatic bond, rather than the ring centroid, with H.X 2.817 (9) A and C-H.X 174.0 (9) degrees, and Br.X 3.353 (4) A and C-Br.X 158.1 (2) degrees. PMID- 11099977 TI - Quantitative analysis of hydrogen bonding and atomic thermal motion in the organic non-linear optical material DCNP using X-ray and neutron diffraction AB - A single-crystal neutron diffraction study of the organic non-linear optical material 3-(1,1-dicyanoethenyl)-1-phenyl-4, 5-dihydro-1H-pyrazole (hereafter DCNP), space group Cc, is presented. The study was conducted in order to relate the structural characteristics of the compound to its physical properties. DCNP exhibits a very large second harmonic generation (SHG) output, an extremely large linear electro-optical effect and photoconductive and pyroelectric properties. The nature of the hydrogen-bonding revealed by the study, in part, accounts for the first two of these phenomena. The neutron study also shows that some rather atypical atomic thermal motion is present in part of the molecule. With the aid of a variable-temperature single-crystal X-ray diffraction study, in conjunction with the neutron study, this thermal motion is attributed to libration and is fully characterized. As a result, suitable corrections to the bond geometry and the anisotropic displacement parameters of DCNP are made. The libration is also shown to enhance the SHG effect. The cell parameters from the variable temperature X-ray study are also used in order to evaluate the thermal expansivity coefficients of DCNP. PMID- 11099978 TI - Selective solvent inclusion as a tool for mapping molecular properties in crystal structures: a diethylstilbestrol example. AB - Useful information about hydrogen bonding, the preferred modes of hydrophobic interaction and conformational preferences of a specific molecule can be obtained from cocrystallization of the solute with a selected series of solvent molecules. This method is used in a study of nine different crystal structures of diethylstilbestrol (DES) solvates. It is shown that solvent inclusion results not only in stronger hydrogen bonds, but usually also in a larger number of favorable C-H.pi interactions between DES molecules. Furthermore, solvent molecules such as DMSO, DMF, acetonitrile and acetone demonstrate important hydrogen-bond donating properties in addition to their more familiar role as hydrogen-bond acceptors. Molecular conformations in the crystal structures compare favorably with results from molecular mechanics calculations. PMID- 11099979 TI - Structure of CnCn+2Cn-type (n = even) beta'-triacylglycerols. AB - The crystal structures of the beta' phase of CLC (1, 3-didecanoyl-2 dodecanoylglycerol) and MPM (1, 3-ditetradecanoyl-2-hexadecanoylglycerol) have been determined from single-crystal X-ray diffraction and high-resolution X-ray powder diffraction data, respectively. Both these crystals are orthorhombic with space group Iba2 and Z = 8. The unit-cell parameters of beta'-CLC are a = 57.368 (6), b = 22.783 (2) and c = 5.6945 (6) A and the final R value is 0.175. The unit cell parameters of beta'-MPM are a = 76.21 (4), b = 22.63 (1) and c = 5.673 (2) A and the final R(p) value is 0.057. Both the beta'-CLC and beta'-MPM molecules are crystallized in a chair conformation, having a bend at the glycerol moiety. The zigzag planes of the acyl chains are orthogonally packed, as is typical for a beta' phase. Furthermore, unit-cell parameters of some other members of the C(n)C(n+2)C(n)-type triacylglycerol series have been refined on their high resolution X-ray powder diffraction pattern. Finally, the crystal structures are compared with the currently known structures and models of triacylglycerols. PMID- 11099980 TI - Conformational transformation coupled with the order-disorder phase transition in 2-methyl-1,3-cyclohexanedione crystals. erratum AB - In the recently published article by Katrusiak (2000) a misprint occurred in the name of one of the authors in a reference (Wasicki et al., 1995) in the final printing stage. The correct spelling of the name is Wasicki. PMID- 11099981 TI - Scintag X'TRA: new X-ray diffraction system PMID- 11099982 TI - Biventricular pacing for congestive heart failure: questions of who, what, where, why, how, and how much. PMID- 11099983 TI - The congestion score: a simple tool for a complicated disease? PMID- 11099984 TI - Incidence and risk factors predictive of unstable angina resulting from restenosis after percutaneous angioplasty of saphenous vein grafts. AB - BACKGROUND: The current study was designed to determine the incidence and risk factors for unstable angina resulting from restenosis in patients undergoing percutaneous transluminal coronary angioplasty (PTCA) of saphenous vein graft (SVG), about which little data are available. METHODS AND RESULTS: A retrospective analysis of a consecutive series of 212 patients undergoing PTCA of SVG was performed. Procedural success was achieved in 200 patients (94.3%) who formed the study group. During a follow-up of 16.8 +/- 10.2 months, 24.5% of patients presented with unstable angina resulting from restenosis. There was a higher prevalence of dyslipidemia (81. 6% vs 51.2%, P <.0002) and greater postprocedural residual stenosis (14.2% +/- 12.6% vs 7.1% +/- 11.0%, P =.007) in patients with unstable angina caused by restenosis compared with the remaining patient population. By multivariate analysis, dyslipidemia (odds ratio [OR] 3.55, 95% confidence interval [CI] 1.64-8.39, P <.002) and to a lesser extent postprocedural residual stenosis (OR 1.04, 95% CI 1.01-1.07, P <.05) were predictive of unstable angina resulting from restenosis. Among dyslipidemic patients, those not on lipid-lowering drugs during the index procedure had a significantly higher incidence of unstable angina caused by restenosis than did those on lipid-lowering drugs (P <.05). CONCLUSION: Unstable angina caused by restenosis presents in as many as one fourth of patients undergoing PTCA of SVG. Dyslipidemia strongly, and to a lesser extent postprocedural residual stenosis, predicts its occurrence. Scrupulous attention to these modifiable risk factors may help reduce the incidence of unstable angina after SVG angioplasty. PMID- 11099985 TI - Design and methodology of the ESPRIT trial: evaluating a novel dosing regimen of eptifibatide in percutaneous coronary intervention. AB - BACKGROUND: Clinical trials of the glycoprotein (GP) IIb/IIIa inhibitors have shown that these potent antiplatelet agents are effective in reducing the ischemic complications of percutaneous coronary interventions. However, even though stents are now implanted in >75% of percutaneous interventional procedures, only one study, a trial of the monoclonal antibody abciximab, has formally evaluated adjunctive GP IIb/IIIa inhibition in this setting. METHODS AND RESULTS: Eptifibatide, a nonimmunogenic and rapidly reversible inhibitor of the platelet receptor integrin IIb/IIIa, has also undergone evaluation as an adjunct to coronary intervention. In clinical trials performed heretofore, however, it has appeared to have less relative clinical efficacy than the monoclonal antibody abciximab. Since the early seminal trials, it has been recognized that the doses of eptifibatide previously used achieved only 30% to 50% of maximal platelet GP IIb/IIIa integrin inhibition. This is considerably less than the 80% level of receptor inhibition that has been proposed to prevent coronary thrombus formation in animal models and that has been achieved in clinical trials with abciximab. CONCLUSIONS: The Enhanced Suppression of the Platelet IIb/IIIa Receptor with Integrilin Therapy (ESPRIT) trial was designed to test the safety and efficacy of a high-dose, "180/2.0/180" double-bolus regimen of eptifibatide (a 180-microg/kg bolus followed 10 minutes later by a second 180-microg/kg bolus of eptifibatide combined with a 2.0-microg/kg per minute infusion) as an adjunct to nonacute percutaneous coronary intervention with stent implantation. In this report, we review the rationale, design, and methods of this clinical investigation. PMID- 11099986 TI - Freedom from congestion predicts good survival despite previous class IV symptoms of heart failure. AB - BACKGROUND: This study determined whether evidence of congestion after 4 to 6 weeks of heart failure management predicted outcome for patients hospitalized with chronic New York Heart Association class IV symptoms. Class IV symptoms predict high mortality rates, but outcome is not known for patients who improve to establish freedom from congestion. Revised estimates at 1 month could facilitate decisions regarding transplantation and other high-risk interventions. METHODS: At 4 to 6 weeks after hospital discharge, 146 patients were evaluated for congestion by 5 criteria (orthopnea, jugular venous distention, edema, weight gain, and new increase in baseline diuretics). Heart failure management included inpatient therapy tailored to relieve congestion, followed by adjustments to maintain fluid balance during the next 4 weeks. RESULTS: Freedom from congestion was demonstrated at 4 to 6 weeks in 80 (54%) patients, who had 87% subsequent 2 year survival compared with 67% in 40 patients with 1 or 2 criteria of congestion and 41% in 26 patients with 3 to 5 criteria. The Cox proportional hazards model identified left ventricular dimension, pulmonary wedge pressure on therapy, and freedom from congestion as independent predictors of survival. Persistence of orthopnea itself predicted 38% 2-year survival (without urgent transplantation) versus 77% in 113 without orthopnea. Serum sodium was lower and blood urea nitrogen and heart rate higher when orthopnea persisted. CONCLUSIONS: The ability to maintain freedom from congestion identifies a population with good survival despite previous class IV symptoms. At 4 to 6 weeks, patients with persistent congestion may be considered for high-risk intervention. PMID- 11099987 TI - Left ventricular hypertrophy as a predictor of coronary heart disease mortality and the effect of hypertension. AB - BACKGROUND: Although associations between hypertension, left ventricular hypertrophy (LVH), and coronary heart disease (CHD) have been described, it is less clear whether LVH is associated with increased rates of CHD in the absence of hypertension. METHODS: We examined this association with Cox regression analyses of data from 7924 adults 25 to 74 years of age from the Second National Health and Nutrition Examination Survey (NHANES II) Mortality Study (1976 to 1992). Covariates included age, race, sex, history of cardiovascular diseases and diabetes, cholesterol, body mass index, blood pressure, and smoking. RESULTS: During 16.8 follow-up years, there were 462 (26%) deaths from CHD (ICD-9 410-414) and 667 (38%) deaths from diseases of the heart (ICD-9 390-398, 402, 404, 410 414, 415-417, 420-429). LVH prevalence was 13.3 per 1000 population. Hypertension prevalence was 29.1%. LVH prevalence was higher among hypertensive adults than among normotensive adults (29.9 vs 6.4 per 1000, P <.001). Persons with LVH were twice as likely to die of CHD (relative risk, 2.0; 95% confidence interval, 1.2, 3.5) and diseases of the heart (relative risk, 1.9; 95% confidence interval, 1.1, 3.0) after adjustment for hypertension and covariates. In age-adjusted predicted survival, probability plots for CHD, and diseases of the heart, normotensives with LVH had survival similar to hypertensive adults with LVH and lower survival than normotensive and hypertensive adults with no LVH. CONCLUSIONS: Our results confirm previous findings that the presence of LVH is a strong predictor of future cardiovascular death. Although LVH appears to be rare among normotensives, clinicians should be aware that such individuals may have an increased risk for death similar to that of hypertensive adults with LVH. PMID- 11099988 TI - Characteristics of patients referred for cardiac transplantation: implications for the donor organ shortage. AB - BACKGROUND: When the decision is made to proceed with cardiac transplantation, the risk/benefit ratio for continued medical therapy in that particular patient must be weighed against the risk/benefit ratio associated with cardiac transplantation. This can only be accomplished while the patient is on maximal medical therapy. METHODS: To better define the appropriateness of patients being referred for consideration of transplant, we examined the records of 100 consecutive adult patients referred to a cardiac transplant program. RESULTS: Two of five patients referred for transplantation had at least one contraindication for transplantation. Twenty percent of the patients were not treated with angiotensin-converting enzyme inhibitors and did not have any documented reason for undertreatment. Of those deemed too well for cardiac transplantation, 84% were alive and either class I or II (mean follow-up 21 months). CONCLUSIONS: We found the majority to be undertreated or with an absolute contraindication to transplantation. Of those deemed too well for transplantation after appropriate treatment, 84% were alive and well. PMID- 11099989 TI - A pilot experience with permanent biventricular pacing to treat advanced heart failure. AB - BACKGROUND: The prognosis and quality of life of patients with advanced heart failure remain poor. The purpose of this study was to evaluate new nonpharmacologic approaches. Biventricular pacing was proposed in this indication, based on the encouraging results of acute hemodynamics studies. METHODS: Fifty patients with drug-resistant heart failure (New York Heart Association [NYHA] class III/IV, 16 of 34) were consecutively implanted with biventricular pacemakers. All patients had severe dilated cardiomyopathy and intraventricular conduction delay. Survival, NYHA class, electrocardiogram, echocardiographic data, and exercise tolerance were assessed over a mean follow up period of 15.4 +/- 10. 2 months. RESULTS: At the end of follow-up, 55% of patients were alive without heart transplantation or left ventricular assistance device. The mortality rate was significantly lower in class III (12. 5%) than in class IV patients (52.5%). In survivors, biventricular pacing significantly improved symptoms (NYHA class 2.2 +/- 0.5 at follow-up vs 3.7 +/- 0.5 at baseline) and exercise tolerance ((. )VO(2) peak 15.5 +/- 3.4 mL/min per kilogram at follow-up vs 11.1 +/- 3 mL/min per kilogram at baseline). CONCLUSIONS: Biventricular pacing appears to improve the functional status of patients with dilated cardiomyopathy with advanced heart failure. The technique appears to be attractive as an additive treatment, especially in class III patients. Controlled randomized studies are needed to validate this novel concept. PMID- 11099990 TI - Rate-control versus conversion strategy in postoperative atrial fibrillation: a prospective, randomized pilot study. AB - BACKGROUND: Atrial fibrillation remains a frequent complication after heart surgery. The optimal strategy to treat the condition has not been established. Several retrospective studies have suggested that a primary rate-control strategy may be equivalent to a strategy that restores sinus rhythm. METHODS: Fifty patients with atrial fibrillation after heart surgery were randomly assigned to a strategy of antiarrhythmic therapy with or without electrical cardioversion or ventricular rate control. Both arms received anticoagulation with heparin overlapped with warfarin. The primary end point was time to conversion to sinus rhythm analyzed by the Kaplan-Meier method. Atrial fibrillation relapse after the initial conversion was monitored in the hospital over a 2-month period. RESULTS: There was no significant difference between an antiarrhythmic conversion strategy (n = 27) and a rate-control strategy (n = 23) in time to conversion to sinus rhythm (11.2 +/- 3. 2 vs 11.8 +/- 3.9 hours; P =.8). With the use of Cox multivariate analysis to control for the effects of age, sex, beta-blocker usage, and type of surgery, the antiarrhythmic strategy showed a trend toward reducing the time from treatment to restoration of sinus rhythm (P =.08). The length of hospital stay was reduced in the antiarrhythmic arm compared with the rate control strategy (9.0 +/- 0.7 vs 13.2 +/- 2.0 days; P =.05). In-hospital relapse rates in the antiarrhythmic arm were 30% compared with 57% in the rate-control strategy (P =.24). There were no significant difference in relapse rates at 1 week (24% vs 28%), 4 weeks (6% vs 12%), and 6 to 8 weeks (4% vs 9%). At the end of the study, 91% of the patients in the rate-control arm were in sinus rhythm compared with 96% in the antiarrhythmic arm (P =.6). CONCLUSIONS: This pilot study shows little difference between a rate-control strategy and a strategy to restore sinus rhythm. Regardless of strategy, most patients will be in sinus rhythm after 2 months. A larger randomized, controlled study is needed to assess the impact of restoration of sinus rhythm on length of stay. PMID- 11099991 TI - New atrial fibrillation after acute myocardial infarction independently predicts death: the GUSTO-III experience. AB - BACKGROUND: Atrial fibrillation (AF) or flutter occurring after myocardial infarction may occur alone or in association with other complications. Whether the arrhythmia portends a poor prognosis independent of other complications with contemporary therapy is unknown. METHODS AND RESULTS: As part of the Global Use of Strategies To Open occluded coronary arteries (GUSTO-III) trial, we evaluated whether postinfarction complications were associated with the subsequent development of AF and whether AF independently predicted death over periods of 30 days and 1 year. Information including exact timing was collected on deaths and major in-hospital postinfarction complications up to 30 days. Of the 13,858 patients with sinus rhythm at enrollment, 906 later had AF or flutter and 12, 952 did not. We compared outcomes between these 2 groups, adjusting for differences in baseline characteristics and prefibrillation complications. Worsening heart failure, hypotension, third-degree heart block, and ventricular fibrillation were independent predictors of new-onset AF. The unadjusted odds ratio (OR) for death among patients with versus those without AF was 2.74 (95% confidence interval [95% CI], 2.56-3.34). After adjusting for baseline differences, the OR was reduced to 1.63 (95% CI, 1.31-2.02). Adjustment for other in-hospital complications before the onset of AF further reduced the OR to 1.49 (95% CI, 1.17 1.89). CONCLUSIONS: Atrial fibrillation or flutter occurs secondary to other postinfarction complications but independently portends a worse prognosis. Prevention and management may improve outcome. PMID- 11099992 TI - Influence of atrial fibrillation on the morbido-mortality of patients on hemodialysis. AB - BACKGROUND: The consequences of atrial fibrillation (AF) on morbido-mortality of patients on hemodialysis have not been fully explored. The objective of this study was to determine the prevalence of AF in patients on hemodialysis and to evaluate its influence on the development of thromboembolic phenomena (TEP). METHODS: The incidence of AF in 190 patients in our hemodialysis program was assessed, and the patients were followed up for 1 year. Pertinent demographic and biochemical parameters were entered into univariate and multivariate statistical analyses to evaluate associations with overall mortality and TEP such as cerebrovascular accident, transitory ischemic accident, or peripheral embolism. RESULTS: In 13.6% of patients, AF was found; 9.4% of these were of the permanent type. In the multivariate analysis, only increased age was associated with a higher probability of having arrhythmia (odds ratio, 1.10; 95% confidence interval, 1.03-1.17; P =.003). During follow-up, 23% of the patients with AF died compared with 6% of those in sinus rhythm (P <.05), although AF did not appear to be an independent predictive factor for death. Thirty-five percent of the patients with AF and 4% with sinus rhythm had TEP (P <.01). In the multivariate analysis, AF was identified as the only independent predictor for TEP (odds ratio, 8; 95% CI, 2.3-27; P =.0008). CONCLUSIONS: AF is a frequent arrhythmia in patients on hemodialysis, and approximately 1 in 3 hemodialysis patients with AF had thromboembolic complications within 1 year of follow-up. These findings suggest that the consensus contraindication of prophylactic anticoagulation therapy for this group of patients may need to be redefined. PMID- 11099993 TI - A randomized study of intravenous magnesium in acute myocardial infarction treated with direct coronary angioplasty. AB - BACKGROUND: Notwithstanding the negative result of the International Study of Infarct Survival-4 (ISIS-4), the controversy about the role of magnesium in acute myocardial infarction is still open because, according to experimental data, magnesium could decrease myocardial damage and mortality only if infusion is started before reperfusion. This randomized placebo-controlled trial was designed to evaluate the effect of intravenous magnesium, delivered before, during, and after direct coronary angioplasty, in patients with acute myocardial infarction. METHODS: One-hundred fifty patients were randomized to intravenous magnesium sulfate or placebo. The primary end point was an infarct zone wall motion score index at 30 days, as a measure of infarct size. The secondary end points included creatine kinase peak, ventricular fibrillation/tachycardia within the first 24 hours, death and congestive heart failure within the 30-day follow-up, and 30-day left ventricular ejection fraction. Analysis was by intention to treat. RESULTS: There were no significant differences between the magnesium and placebo groups in the 30-day infarct zone wall motion score index (1.93 +/- 0.61 vs 1.85 +/- 0.51, P =.39), ventricular arrhythmias (24% vs 15%, P =.15), death (0 vs 1%, P =.32), heart failure (8% vs 7%, P =.75), and 30-day left ventricular ejection fraction (49% +/- 11% vs 50% +/- 9%, P = 0.55). There was a trend toward a higher creatine kinase peak in the magnesium group (3059 +/- 2359 vs 2404 +/- 1673,P =.052). CONCLUSIONS: Intravenous magnesium delivered before, during, and after reperfusion did not decrease myocardial damage and did not improve the short-term clinical outcome in patients with acute myocardial infarction treated with direct angioplasty. PMID- 11099994 TI - Immediate and one-year outcome of intracoronary stent implantation in small coronary arteries with 2.5-mm stents. AB - BACKGROUND: The role of coronary stenting in the treatment of stenoses in small coronary arteries with use of 2.5-mm stents is not well defined. METHODS AND RESULTS: Between January 1995 and August 1999, 651 patients with stenoses in small coronary arteries were treated with 2.5-mm stents (n = 108) or 2.5-mm conventional balloon angioplasty (BA) (n = 543). Patients who received treatment with both 2.5-mm and > or =3.0-mm stent placement or balloons were excluded. Procedural success and complication rates as well as 1-year follow-up outcomes were examined. Baseline clinical characteristics were similar between the two groups, except patients in the stent group were more likely to have hypertension and a family history of coronary artery disease and less likely to have prior myocardial infarction. Angiographic success rates were higher in the stent group (97.2% vs 90.2%, P =.02). In-hospital complication rates were comparable between the two groups. Among successfully treated patients, 1-year follow-up revealed no significant differences in the survival (96.2% vs 95.2%, P =.89) or the frequency of Q-wave myocardial infarction (0% vs 0.4%, P =.60) or coronary artery bypass grafting (8.4% vs 6.8%, P =.89) between the stent and BA groups, respectively. However, patients in the stent group were more likely to have adverse cardiac events (35.4% vs 22.1%, P =.05). Stent use after excluding GR II stent use, however, was not independently associated with reduced cardiac events at follow up (relative risk 1. 3 [95% confidence interval 0.8-2.3], P =.30). CONCLUSIONS: Intracoronary stent implantation of stenoses in small coronary arteries with 2.5 mm stents can be carried out with high success and acceptable complication rates. However, compared with BA alone, stent use was not associated with improved outcome through 1 year of follow-up. PMID- 11099996 TI - Elevated cardiac troponin levels predict the risk of adverse outcome in patients with acute coronary syndromes. AB - BACKGROUND: Elevations of cardiac troponin T or I are predictive of adverse outcomes in patients with acute coronary syndromes. However, odds ratios (ORs) vary substantially between studies. This investigation refines these values by means of a meta-analysis. METHODS: Twenty-one studies were suitable. ORs were calculated for short-term (30 days) and long-term (5 months to 3 years) follow-up in patients with ST-segment elevation (ST upward arrow), in those without ST segment elevation (no ST upward arrow), and in patients with unstable angina. The primary end point was a composite of death or nonfatal myocardial infarction. RESULTS: A total of 18,982 patients were included. At 30 days, the OR for death or myocardial infarction was 3.44 (95% confidence interval [CI], 2.94-4.03; P <. 00001) for patients with positive troponin. In the ST upward arrow group, troponin elevations carried a 2.86-fold (95% CI, 2.35-3.47; P <.0001) higher risk during short-term follow-up, which was maintained long term. The no-ST upward arrow patients with troponin elevations manifested a 4.93-fold (95% CI, 3.77 6.45; P <.0001) increase of adverse outcomes. The OR for patients with unstable angina and positive troponin was 9.39 (95% CI, 6.46-13.67; P <.0001). For cardiac death alone, the results were similar. CONCLUSIONS: Patients with acute coronary syndromes who have troponin elevations show a substantial increase in risk during short and long-term follow-up. PMID- 11099995 TI - Thrombolytic therapy for prosthetic valve thrombosis: short- and long-term results. AB - BACKGROUND: Thrombolytic therapy (TT) has evolved as an alternative to surgery for prosthetic valve thrombosis (PVT), but its utility in patient management is still debated and the long-term results are not available. METHODS: From 1990 through 1999, we treated 110 consecutive patients (52 men, mean age 35.4 +/- 10.8 years) of left-sided obstructive PVT (96 mitral, 14 aortic) with TT (streptokinase in 108, urokinase in 2) according to a specified protocol of prolonged infusion. Serial echo Doppler parameters were monitored in all patients to guide the duration of TT and to quantify its efficacy. Ninety of the 102 survivors of the index episode were followed up for a mean period of 31.3 +/- 27.8 months (range 1-112 months). RESULTS: Complete hemodynamic response (on cinefluoroscopy and echo Doppler criteria) was seen in 90 (81.8%) episodes, partial response in 11 (10%), and failure in 9 (8.2%). The mean duration of TT was 42.8 +/- 20.4 hours. Five of the 7 patients who were initially seen in cardiogenic shock/overt pulmonary edema died during therapy. After these patients were excluded, the rate of complete response did not differ among patients with New York Heart Association class I/II (80%), class III (86.3%), or class IV (81.5%). The response rate also did not vary with the type, position of prosthesis, duration of symptoms, or time lag since surgery. There were 21 (19.1%) embolic episodes during therapy, including 6 strokes. These were significantly more frequent in patients with atrial fibrillation (AF) (odds ratio on multivariate analysis 2.3, 95% confidence interval 1.3-3.9, P =.01). On follow up, there were 25 recurrences of PVT, of which 20 again received TT with a complete response in 14 (70%). At 5 years the actuarial survival was 85.2% and the event-free survival was 61.5%. The presence of chronic AF was a significant predictor of recurrence of PVT (odds ratio 2.2, 95% confidence interval 1.2-3.9, P =.008). CONCLUSIONS: TT is effective in the majority of patients with PVT but is associated with a high rate of embolism, especially in patients with AF. Excluding patients with cardiogenic shock/overt pulmonary edema (in whom TT is largely ineffective), the success of TT does not vary with the New York Heart Association class, duration of symptoms, or other patient variables. The recurrence rates of PVT are high after even successful TT, especially in patients with AF. PMID- 11099997 TI - Positron emission tomography and low-dose dobutamine echocardiography in the prediction of postrevascularization improvement in left ventricular function and exercise parameters. AB - BACKGROUND: We studied the value of low-dose dobutamine echocardiography (LDDE) and positron emission tomography (PET) in predicting improvement of left ventricular function and exercise parameters after revascularization. METHODS: Forty-six consecutive patients with ischemic heart disease and an ejection fraction (EF) of 35% +/- 7% were included. Before revascularization, the patients underwent exercise testing and myocardial viability testing by LDDE and fluoride 18-fluoro-2-deoxyglucose PET. Six months after revascularization they underwent coronary angiography to study graft patency, and echocardiographic examination and the exercise test were repeated. RESULTS: In the prediction of the presence or absence of improved postrevascularization function in left ventricular regions with patent grafts, PET was more sensitive than LDDE (42/52 regions [81%] vs 27/52 regions [51%], P <.01), whereas LDDE was more specific than PET (187/209 regions [89%] vs 118/209 regions [56%], P <.001). Improvement of regional myocardial dysfunction was found in 19 patients, but their global left ventricular function did not improve significantly (EF 34% +/- 6% and 36% +/- 7%). In the remaining 27 patients with irreversible dysfunction, EF decreased (EF 36% +/- 7% vs 32% +/- 8%, P <.05). Among patients with reversible myocardial dysfunction, the rate pressure product (RPP) increased after revascularization (19,522 +/- 5474 vs 26,190 +/- 5610 mm Hg/min, P <.01), whereas the RPP did not change in patients with irreversible myocardial dysfunction (21,546 +/- 5450 and 22,774 +/- 8249 mm Hg/min). The number of PET viable segments was a predictor of the postoperative increase in the RPP in univariate (P <.04) and multivariate analyses (P <.001). In contrast, LDDE did not bear any prognostic information about improvement in the RPP. CONCLUSIONS: This study confirms earlier findings of higher sensitivity and lower specificity of PET compared with LDDE in predicting improvement of regional left ventricular function after revascularization. However, the feasibility of predicting postrevascularization improvement of exercise parameters seems unique for PET. The potential prognostic value of this finding needs further investigation. PMID- 11099998 TI - Specificity of the stress electrocardiogram during adenosine myocardial perfusion imaging in patients taking digoxin. AB - BACKGROUND: In patients taking digoxin, the exercise electrocardiogram has a lower specificity for detecting coronary artery disease. However, the effect of digoxin on adenosine-induced ST-segment depression is unknown. The purpose of this study was to evaluate the specificity of the electrocardiogram during adenosine myocardial perfusion imaging in patients taking digoxin. METHODS: Between May 1991 and September 1997, patients (n = 99) taking digoxin who underwent adenosine stress imaging with thallium-201 or technetium-99m sestamibi and coronary angiography within 3 months were retrospectively identified. Exclusion criteria included prior myocardial infarction, coronary artery angioplasty or bypass surgery, left bundle branch block, paced ventricular rhythm, or significant valvular disease. Twelve-lead electrocardiograms were visually interpreted at baseline, during adenosine infusion, and during the recovery period. The stress electrocardiogram was considered positive if there was > or =1 mm additional horizontal or downsloping ST-segment depression or elevation 0.08 seconds after the J-point compared with the baseline tracing. RESULTS: ST-segment depression and/or elevation occurred in 24 of 99 patients. There were only 2 false-positive stress electrocardiograms, yielding a specificity of 87% and positive predictive value of 92%. All 8 patients with > or =2 mm ST segment depression had multivessel disease by coronary angiography. CONCLUSIONS: ST-segment depression or elevation during adenosine myocardial perfusion imaging in patients taking digoxin is highly specific for coronary artery disease. Marked (> or =2 mm) ST-segment depression and/or ST-segment elevation is associated with a high likelihood of multivessel disease. PMID- 11099999 TI - The 24-hour rhythm of blood pressure differs from that of leg hemodynamics in orthotopic heart transplant recipients. AB - BACKGROUND: This study was aimed at investigating whether a circadian rhythm of peripheral resistance exists in patients with orthotopic cardiac transplantation (OCT) and whether it parallels that of blood pressure (BP). METHODS: BP and leg flow and resistance (plethysmography) were monitored for 24 hours in 13 denervated OCT recipients and 13 control patients with native heart, matched for casual blood pressure. RESULTS: On the basis of BP trend, control patients showed a BP reduction during sleep, whereas OCT recipients did not. Leg resistance was significantly lower and leg flow significantly higher during sleep than during waking in all patients, and the extent of the nocturnal decrease was similar in the two categories. CONCLUSIONS: The decrease in leg resistance in patients confined to bed for 24 hours is caused by peripheral mechanisms and does not depend on the autonomic control of the heart. The nocturnal decline in BP depends, on the contrary, on cardiac control and is lost in patients with denervated heart. PMID- 11100000 TI - Long-term effects of bisoprolol compared with imidapril on left ventricular remodeling after reperfusion in acute myocardial infarction: an angiographic study in patients with maintained vessel patency. AB - BACKGROUND: Although angiotensin-converting enzyme inhibitor attenuates ventricular enlargement, whether beta-blocker therapy induces regression of left ventricular remodeling is not known. The purpose of this study was to compare the effects of bisoprolol therapy with those of imidapril therapy on left ventricular remodeling after acute myocardial infarction (AMI). METHODS: Sixty patients with AMI who underwent reperfusion therapy were randomly assigned to an imidapril group (20 patients), a bisoprolol group (20 patients), or a control group (20 patients). Administration was started within 24 hours. Left ventricular function on admission and 3 months and 1 year after AMI was investigated. RESULTS: Baseline characteristics on admission were similar in the 3 groups except for sex distribution. Mean pulmonary capillary wedge pressure and left ventricular end diastolic pressure in the bisoprolol group were higher than those in the imidapril group 1 year after admission (pulmonary capillary wedge pressure: 12 +/ 7 vs 8 +/- 2 mm Hg, left ventricular end-diastolic pressure: 17 +/- 8 vs 11 +/- 4 mm Hg, P <. 01). Left ventricular end-diastolic volume index (EDVI) increased in the bisoprolol group throughout the 1-year period (P <.01), whereas EDVI in the imidapril group decreased (P <.01). The increases in EDVI during 1 year in the bisoprolol group were greater than those of the other 2 groups (bisoprolol: 12 +/- 10, imidapril: -9 +/- 7, control: 4 +/- 11 mL/m2, P <.01). CONCLUSIONS: Early treatment with bisoprolol in AMI cannot prevent left ventricular remodeling, whereas imidapril attenuates left ventricular dilation by decreasing preload. PMID- 11100001 TI - Circulating levels of cytokines and their site of production in patients with mild to severe chronic heart failure. AB - BACKGROUND: Patients with chronic heart failure have elevated levels of proinflammatory cytokines; however, the mechanism for their increased expression and the site of their production are unknown. METHODS: Twenty-two patients with heart failure, New York Heart Association functional class II to IV, underwent hemodynamic evaluation and echocardiographic study. Blood samples for cytokine evaluation were performed in the ascending aorta, coronary sinus, inferior vena cava, and hepatic vein. Levels of tumor necrosis factor-alpha (TNF-alpha), its soluble receptors sTNF-RI and sTNF-RII, interleukin-6 (IL-6), IL-6 soluble receptor, soluble gp130, interleukin-2 soluble receptor, and soluble Fas were measured with enzyme-linked immunosorbent assay kits. RESULTS: IL-6 concentrations were higher in class IV patients than in class III patients, which in turn were higher than those in class II. TNF-alpha, sTNF-RI, and sTNF-RII were higher in class IV patients than in class III and II patients. Significant correlations were found between IL-6 concentrations and left ventricular end systolic volume (r = 0.64; P <.001), pulmonary wedge pressure (r = 0.56; P <.01), and left ventricular ejection fraction (r = -0.56; P <.01). No correlation was found between TNF-alpha and its soluble receptors and left ventricular volumes or hemodynamic measures. Finally, no difference in cytokine concentrations was found among the different sample sites. CONCLUSIONS: Among inflammatory cytokines, IL-6 concentrations better reflect the hemodynamic derangement in patients with heart failure. No cardiac or gut production of cytokines occurs in patients with mild to severe heart failure. PMID- 11100003 TI - Importance of microembolization and inflammation in atherosclerotic heart disease. AB - Considerable evidence now exists that inflammation is a central component of events that initiate and propagate an acute coronary syndrome. This process evokes the potential for embolization, which occurs more often than previously suspected, and imparts poor cardiovascular prognosis. Recent development of techniques to detect inflammation and embolization represents an important advance. In addition, therapies that diminish occurrence of these phenomena such as aspirin, statins, angiotensin converting enzyme (ACE) inhibitors, and IIb/IIIa receptor antagonists have been shown to improve outlook. PMID- 11100004 TI - GUSTO IV: expanding therapeutic options in acute coronary syndromes. PMID- 11100005 TI - Glycoprotein IIb/IIIa inhibitors: therapeutic applications in acute ST-segment elevation myocardial infarction. PMID- 11100006 TI - Facilitated percutaneous coronary intervention: a novel concept in expediting and improving acute myocardial infarction care. PMID- 11100008 TI - Critical concepts in cost-effectiveness for cardiovascular specialists. PMID- 11100007 TI - Applications of monitoring platelet glycoprotein IIb/IIIa antagonism and low molecular weight heparins in cardiovascular medicine. AB - With the addition of more potent antiplatelet and antithrombin therapies to the armamentarium of the treatment of acute coronary syndromes and percutaneous coronary interventions, monitoring these therapies has become an important interest. Current and evolving technologies to monitor the extent of inhibition of platelet aggregation and activity of factor Xa caused by IIb/IIIa antagonists and low-molecular-weight heparin, respectively, will be covered in this overview. An underlying question to be considered is whether the results generated from monitoring will effect a change that will improve the efficacy (prevent thrombotic events) or reduce adverse events (bleeding) from these potent therapies. PMID- 11100009 TI - Abciximab: cost-effective survival advantage in clinical trials and clinical practice. AB - Adjunctive blockade of the platelet glycoprotein (GP) IIb/IIIa receptor during either percutaneous coronary intervention (PCI) or for patients who present with non-ST segment elevation acute coronary syndromes has demonstrated efficacy in reducing platelet-mediated adverse cardiovascular ischemic events. The three currently available agents (abciximab, eptifibatide, tirofiban) differ markedly in pharmacodynamic and pharmacokinetic profiles, receptor affinity, and cost. Although pharmacoeconomic substudies are available from placebo-controlled randomized trials of platelet GPIIb/IIIa blockade during PCI, "real-world" cost effectiveness data from high-volume practice are lacking. Therefore, in-hospital and late (6-month) clinical outcomes and cumulative cost/charge data were analyzed on 1472 consecutive PCI procedures (70% received abciximab) performed by high-volume operators at a single institution.(1) Data were adjusted for lack of randomized treatment allocation with the use of a propensity scoring technique. Adjunctive abciximab therapy for PCI was associated with a significant (3.4%) reduction in mortality to 6 months. Based on the economic cost-effectiveness concept of cost per life year gained relative to standard therapy,(2,3) abciximab provided a cost-effective survival advantage in high-volume interventional practice that compares very favorably with currently accepted standards. Clinical and procedural demographics associated with increased cost-effectiveness include multivessel coronary intervention, stent deployment, recent (<1 week) myocardial infarction (MI), and impaired left-ventricular (LV) function. PMID- 11100010 TI - Exploring the issues of appropriate dosing in the treatment of acute myocardial infarction: potential benefits of bolus fibrinolytic agents. AB - The Institute of Medicine report on the frequency and consequences of medical errors in clinical practice has stimulated physicians to evaluate current practice and means of improving medical care. In the treatment of patients with acute myocardial infarction, previous studies have found that dosing of fibrinolytic therapy is closely related to outcomes, with too low a dose associated with lower rates of infarct-related artery patency and higher doses associated with increased intracranial hemorrhage. Thus there is a narrow "therapeutic window" for fibrinolytic-antithrombotic regimens, and the potential for adverse outcomes is high if incorrect doses are administered. The first demonstration of this concept came from the GUSTO-I trial, in which 13.5% of patients treated with streptokinase and 11.5% of patients treated with tissue plasminogen activator (t-PA) had a dosing regimen that deviated from the protocol, that is, an incorrect total dose or infusion length. In patients with protocol deviations, 24-hour and 30-day mortality rates were significantly higher compared with those of patients with per-protocol dosing: for t-PA, patients who received incorrect dosing had a 30-day mortality rate of 7.7% versus 5.5% for patients who received correct t-PA dosing (P <.001), with similar findings for streptokinase. More recent data from the InTIME-II trial have shown that the use of a bolus fibrinolytic agent significantly increases the percentage of patients who receive complete and optimally dosed fibrinolysis. Thus use of the simpler bolus fibrinolytic agents may reduce medication errors and thus may optimize clinical outcomes. PMID- 11100013 TI - Cutaneous manifestations of end-stage renal disease. AB - Examination of the skin and nails can reveal many abnormalities in patients with end-stage renal disease that precede or follow initiation of dialysis treatment or kidney transplantation. This article focuses on specific and nonspecific cutaneous signs of end-stage renal disease, reviewing both banal and life threatening conditions, including pruritus, perforating disorders, calcifying disorders, and bullous dermatoses. The pathogenesis, clinical findings, histologic findings, differential diagnosis, and treatment of these diseases are discussed. Cutaneous manifestations unique to kidney transplantation will not be covered. (J Am Acad Dermatol 2000;43:975-86.) PMID- 11100014 TI - Prognostic value of Ki-67 expression in localized cutaneous malignant melanoma. AB - BACKGROUND: The proliferative activity of some tumors is related to the development of metastatic disease and survival. Thus it could be used as a potential prognostic variable. OBJECTIVE: The purpose of this study was to determine the prognostic value of the Ki-67 index and of a "proliferation-based prognostic index" (PBPI, derived as tumor thickness x Ki-67 index/100) in localized cutaneous malignant melanoma (CMM). METHODS: The Ki-67 index (percent of total tumor nuclei) was determined in a series of 84 localized CMMs, with the use of the alkaline phosphatase-antialkaline phosphatase labeling method in formalin-fixed, paraffin-embedded material, and was correlated with other prognostic variables. Survival analysis was performed to determine whether the Ki 67 index and the PBPI could be predictive of metastatic spread or recurrent disease. A stratified analysis of these two parameters according to the tumor thickness was done. RESULTS: An association among the Ki-67 index and location, Clark level, tumor thickness and stage, and prognostic index was detected. Increased Ki-67 index and PBPI were associated with poorer overall survival (P =.03 and P <.0001, respectively) and disease-free survival (P =.01 and P <.0001, respectively). However, after stratification for thickness, only the PBPI showed independent prognostic significance, restricted to tumors thicker than 4 mm (P =. 03). CONCLUSION: The determination of the PBPI in CMM conveys prognostic information for localized thick (>4 mm) CMM, identifying two groups of patients with distinct outcome. PMID- 11100015 TI - Leg ulcers in peripheral arterial disease (arterial leg ulcers): impaired wound healing above the threshold of chronic critical limb ischemia. AB - BACKGROUND: Peripheral arterial disease is the only identifiable etiology in approximately 10% of leg ulcers. Clinical data on the management of these chronic wounds are scarce. OBJECTIVE: We attempted to outline the threshold of systolic ankle pressure and ankle-brachial-index (ABI) below which arterial leg ulcers can occur and to outline the indication for revascularization in arterial leg ulcers. METHODS: Diagnostic and outcome analysis was performed for 26 consecutive patients with arterial leg ulcers. We calculated sensitivities, specificities, and receiver operating characteristic (ROC) curves for the identification of arterial leg ulcers among all 223 consecutive leg ulcer patients within a 3-year period, as well as the ROC curve for patients who required revascularization. RESULTS: The systolic ankle pressure was 88 (18-130) mm Hg (median; 95% confidence interval) and the ABI was 0.60 (0.15-0.86), respectively. Eighteen patients (69%) were subjected to revascularization. By the end of the study, 24 patients (92%) healed completely, 1 improved (90% wound closure), and 1 patient had to undergo below-knee amputation for chronic osteomyelitis. During this study, the ankle pressure and ABI were poor in distinguishing those patients who required revascularization from those who healed without revascularization. CONCLUSION: Most arterial leg ulcers do not meet the criteria of chronic critical limb ischemia, but they do not heal under conservative measures, either. A majority of these patients benefit from revascularization and should, therefore, be referred for arterial duplex ultrasound investigation or angiography. In our study, an ankle pressure below 110 mm Hg identified all patients (100%) who were subjected to revascularization procedures. However, controlled clinical studies are required to find the systolic ankle pressure and ABI below which revascularization can be recommended to speed up the healing time. PMID- 11100016 TI - In vitro and in vivo determination of the UV protection factor for lightweight cotton and viscose summer fabrics: a preliminary study. AB - BACKGROUND: One of the most important elements in the prevention of skin cancer is the use of comfortable UV-protective clothing. Owing to their low weight, cotton fabrics, and especially viscose fabrics made from filament yarns, are ideal for summer clothing and in fact enjoy a high degree of acceptance among consumers. Two methods are available for determining the UV protection factor (UPF) of textiles: the in vitro method is based on the spectrophotometric determination of the transmission of UV radiation through these fabrics; the in vivo method is based on the determination of the minimal erythema dose for a test subject with and without textiles. OBJECTIVE: This study was performed to assess the UPF of lightweight cotton and viscose fabrics and whether the use of these two methods to determine the UPF of viscose fabrics and cotton fabrics produces congruent results. METHODS: We tested 7 different viscose fabrics and 7 different cotton fabrics. Three of the viscose fabrics (ENKA SUN) had been specially treated, by depositing pigments in the fibers, to confer UV-protective properties. The determination of the in vitro and in vivo UPF was performed with a spectrophotometer and sun simulator, respectively. RESULTS: The in vivo measurements on the untreated viscose fabrics produced UPF values lower than those obtained from the in vitro measurements. For one of these untreated viscose fabrics, the difference between the in vivo UPF and the in vitro UPF was statistically significant (P <.05). In contrast, the in vivo measurements on the specially treated viscose textiles and on the cotton fabrics resulted in UPF values higher than the in vitro UPF values. For one specially treated viscose fabric and 4 cotton fabrics, this difference was statistically significant (P < .05). CONCLUSION: Our results suggest, however, that-depending on the type of fabric-determination of the UPF by the in vitro method is not in agreement with the in vivo method. In vivo measurements made with lightweight specially treated UV-protective clothing showed in contrast to the untreated viscose fabrics that these garments offer very good protection against UV radiation. These results underscore the importance of developing and refining such UV-protective materials. PMID- 11100017 TI - Double-blind, placebo-controlled study of oral calcitriol for the treatment of localized and systemic scleroderma. AB - BACKGROUND: Various treatments including corticosteroids, nonsteroidal anti inflammatory drugs, D-penicillamine, interferon gamma, cyclosporine, and cytostatic drugs have been used with limited success in both morphea and systemic sclerosis (SSc). OBJECTIVE: We investigated the effect of treatment with oral calcitriol in patients with localized or systemic scleroderma. METHODS: A randomized, double-blind, placebo-controlled study of 9 months' duration with a 6 month follow-up was performed at the Department of Dermatology. A total of 27 patients (7 patients with SSc and 20 with morphea) were selected on a minimal skin score of 3 for patients with morphea and 12 for those with SSc. Each patient received calcitriol (0.75 microg/day for 6 months plus 1.25 microg/day for 3 months) or placebo for 9 months. Efficacy parameters included skin score, measurement of serum markers of collagen synthesis and degradation and, additional for the patients with SSc, oral aperture measurements, lung function studies, and esophagus motility. RESULTS: The skin score in patients with morphea after 9 months' treatment showed no significant difference between the placebo and calcitriol groups (mean percentage reduction [SD] in skin score in the placebo group was -29.3 [57.9]; in the calcitriol group it was -19.4 [46.6]). The small group of patients with SSc was inadequate to allow us to draw any conclusions regarding efficacy. No significant change was found in the serum markers of collagen metabolism. CONCLUSION: In this study calcitriol was not more effective than placebo in patients with morphea. Because of the small group of patients with SSc treated, no conclusions regarding efficacy in SSc can be drawn. PMID- 11100018 TI - In vitro assessment of the broad-spectrum ultraviolet protection of sunscreen products. AB - BACKGROUND: There are considerable data to suggest that protection from solar ultraviolet (UV) radiation will reduce the risk of acute and chronic skin damage in humans. Whereas the sun protection factor (SPF) provides an index of protection against erythemally effective solar UV, largely confined to the UVB (290-320 nm) and short-wavelength UVA (320-340 nm) region, there is currently no agreed-upon method to measure broad-spectrum protection against long-wavelength UVA (340-400 nm). OBJECTIVE: The objective of these studies was to assess the potential of in vitro UV substrate spectrophotometry and subsequent calculation of the "critical wavelength" value as a measure of broad-spectrum UV protection and as a routine, practical procedure for classification of sunscreen products. METHODS: The spectral absorption of 59 commercially available sunscreen products and multiple experimental formulas with one or more UV filters was measured. Sunscreen product, 1 mg/cm(2), was applied to a hydrated synthetic collagen substrate, preirradiated with a solar simulator, and then subjected to UV substrate spectrophotometry. Multiple determinations from 5 independent samples per product were used to calculate the critical wavelength value, defined as the wavelength at which the integral of the spectral absorbance curve reached 90% of the integral from 290 to 400 nm. RESULTS: We found that a recognized long-wave UVA active ingredient such as titanium dioxide, zinc oxide, or avobenzone is a necessary but insufficient product requirement for achieving the highest proposed broad-spectrum classification, that is, critical wavelength of 370 nm or more. Although SPF and critical wavelength are largely independent of each other, UVA absorbance must increase commensurate with SPF to maintain the same critical wavelength value. Substrate spectrophotometry and the calculation of critical wavelength can readily account for sunscreen photostability by UV preirradiation. Finally, there is also a strong positive relationship between critical wavelength and a currently available in vivo measure of UVA protection. CONCLUSION: Determination of critical wavelength by means of UV substrate spectrophotometry provides a rapid, inexpensive, and reliable measure of broad-spectrum protection, which is largely independent of SPF, yet ensures long-wavelength UVA protection commensurate with SPF. The procedure provides a routine, sensitive means of differentiating and classifying sunscreen products and, importantly, obviates the need to subject volunteers to acute exposures of high-dose, nonterrestrial UV, the health risks of which are still poorly understood. PMID- 11100019 TI - Comparison of UVA protection afforded by high sun protection factor sunscreens. AB - UVA protection afforded by 6 different sunscreens with a sun protection factor of 21 or more was compared by means of the persistent pigmentation darkening method. Colorimetric and visual assessment showed significant differences in UV radiation induced pigmentation at 2 hours. The labeled sun protection factor of the tested sunscreens was not predictive of UVA protection level. PMID- 11100020 TI - Tinea corporis gladiatorum: a cross-sectional study. AB - BACKGROUND: Tinea corporis gladiatorum has been an infrequently reported condition in the dermatological literature. Two previous reports have investigated wrestlers during recognized epidemics of tinea corporis gladiatorum, but no study has examined wrestlers to determine the point prevalence of tinea corporis gladiatorum in a team without a known epidemic. Furthermore, no comparative study exists. OBJECTIVE: The purpose of this article is to determine the prevalence of tinea corporis gladiatorum in high school wrestlers in comparison with high school indoor track athletes. METHODS: Members of both the high school wrestling and indoor track teams were examined for the presence of tinea corporis. Students with clinical lesions of tinea corporis were evaluated with potassium hydroxide examination. The prevalence of tinea corporis in each group was compared. RESULTS: Seven of the 29 high school wrestlers (24%) had lesions of tinea corporis. No members of the track team had evidence of tinea corporis. There was a statistical difference between the two groups (P =.005). CONCLUSION: Tinea corporis gladiatorum can be found quite frequently among high school wrestlers. Because infection with dermatophytes can disqualify a wrestler from competing in matches, vigilant surveillance and rapid initiation of therapy can reduce the suspension of a team's practice and competition. PMID- 11100021 TI - Medication and medical service utilization for acne 1995-1998. AB - BACKGROUND: Acne occurs in most persons sometime during adolescence or early adulthood and is a frequent reason for visits to dermatologists and for prescription drug therapy. OBJECTIVE: The purpose of this study was to analyze changes in the utilization of physician services and medications for the treatment of acne. METHODS: An analysis was performed of data from two US federal surveys of outpatient physician services and prescribing for the years 1980 to 1997 and two commercial sources of drug prescription data for 1996 to 1998. From these data, I estimated visits for acne and drugs prescribed during these visits. RESULTS: Visits for women principally for acne are about 80% more frequent than those for men. Each year 5 million prescriptions for oral antibiotics and 1.4 million prescriptions for isotretinoin are dispensed for the treatment of acne. CONCLUSION: Substantial health care resources are devoted to the treatment of acne, with women particularly likely to continue to frequently utilize these services after 19 years of age. PMID- 11100022 TI - Analysis of current data on the use of intravenous immunoglobulins in management of pemphigus vulgaris. AB - BACKGROUND: Systemic corticosteroids, with or without the addition of immunosuppressive adjuvant agents, are frequently used in treating patients with pemphigus vulgaris (PV). The severe, catastrophic, and potentially fatal side effects of these agents highlight the need for the development of safe alternatives for PV therapy. Intravenous immunoglobulin (IVIG) therapy has recently been reported to be effective in the treatment of PV. OBJECTIVE: Our purpose was to do a retrospective analysis of the available literature on the use of IVIG in the treatment of PV. We also wished to determine whether the cumulative evidence permits making preliminary conclusions regarding the potential role of IVIG in the overall management of PV. METHODS: A review of the English-language, peer-reviewed literature was conducted for reports on IVIG use in treatment of PV. The available information on 21 patients was used to assess different dimensions of clinical efficacy. RESULTS: A minimum dose of 2 g/kg per cycle given at regular monthly intervals for a minimum of 3 cycles seems be effective in inducing a rapid clinical remission in patients with severe, recalcitrant PV. However, this should not be perceived as a "standard" dose. Tapering and eventual discontinuation of other agents were possible in many patients. Long-term follow-up was not provided to examine the influence of IVIG on the clinical course of disease, its efficacy as monotherapy, and the benefit of using it as maintenance therapy to keep the patient in prolonged clinical remission. In 4 of the 21 patients (19%), use of IVIG was of no clinical benefit. This failure of efficacy was primarily due to inadequate use. IVIG demonstrated beneficial effect in 17 of 21 patients (81%). CONCLUSION: IVIG may be a safe and effective agent in the management of severe, recalcitrant PV. Multicenter controlled studies, using different dose regimens, with lengthy follow-up periods are necessary to clearly define the emerging beneficial role of IVIG. PMID- 11100023 TI - The use of plasmapheresis and immunosuppression in the treatment of pemphigus vulgaris. AB - BACKGROUND: Pemphigus vulgaris is an autoimmune blistering disease for which the mainstay of treatment is systemic corticosteroids and immunosuppressants. This therapy had reduced the mortality of pemphigus; however, it is associated with significant morbidity. OBJECTIVE: The objective of this study was to assess the group's experience with plasmapheresis in the treatment of pemphigus vulgaris and report on its utility. METHODS: Seven patients with severe or resistant pemphigus vulgaris underwent a series of 5 plasma exchanges over an average of 8 days. Immunosuppressive drugs were administered immediately after plasmapheresis to prevent the "rebound" flare of disease that can occur after plasmapheresis. RESULTS: Remission was induced in 4 patients, partial remission was induced in 2 patients, and 1 patient continues to have active disease. CONCLUSION: This study suggests that plasmapheresis is a useful intervention in patients with pemphigus vulgaris who are not responding to standard therapy or who require unacceptably high doses of steroids or immunosuppressants. PMID- 11100024 TI - Immunoablative high-dose cyclophosphamide without stem cell rescue in a patient with pemphigus vulgaris. AB - The use of ablative intravenous cyclophosphamide (50 mg/kg per day for 4 days) without stem cell rescue has been described in patients with refractory autoimmune diseases such as paraneoplastic pemphigus, systemic lupus erythematosus, and aplastic anemia. We describe a 33-year-old patient with pemphigus vulgaris recalcitrant to multiple therapies. The patient presented with numerous oral ulcerations, erosions, and hyperpigmented crusted plaques on his face, trunk, and arms. Findings of a skin biopsy and direct immunofluorescence were consistent with pemphigus vulgaris. The circulating pemphigus vulgaris autoantibodies were present at a titer of 1:640. The patient received immunoablative therapy (50 mg/kg of cyclophosphamide for a total of 4 days) and tolerated the regimen well. Complications such as thrombocytopenia and Pseudomonas septicemia were quickly treated. Four months after the 4-day therapy, his oral and skin lesions completely healed, and his pemphigus titers have decreased to zero. He is no longer receiving prednisone and no new lesions have developed. This provides further evidence that this regimen is relatively safe and provides a potential "cure" for refractory autoimmune diseases such as pemphigus vulgaris. PMID- 11100025 TI - Hyperthermic treatment of Bowen's disease with disposable chemical pocket warmers: a report of 8 cases. AB - Bowen's disease is a form of squamous cell carcinoma in situ, in which local hyperthermia may be efficacious. We studied 8 patients with Bowen's disease to ascertain whether hyperthermia can be effective against it. As a heat source, disposable chemical pocket warmers were applied daily with pressure directly to the lesion site while the patient was awake, and the clinical course was observed for 4 to 5 months. The lesion was then excised and examined to determine the histopathologic effects. The results showed efficacy in 6 cases, in which the nodular and invasive lesions and the erythematous patches abated; then the lesions disappeared, leaving deposits of pigment (complete remission). In one case, the signs improved by at least 50% (partial remission); in the other case, there was only slight palliation (no response). As for the posttreatment histopathologic effects, tumor cells were eliminated in 3 cases, isolated tumor cells were seen in 3 cases, and there was no change in 2. Although the treatment did not yield perfect results, it represents a major improvement of hyperthermic therapy and is one effective method of treating Bowen's disease. PMID- 11100026 TI - Histopathologic features seen in Gianotti-Crosti syndrome secondary to Epstein Barr virus. AB - BACKGROUND: Gianotti-Crosti syndrome (GCS) or infantile papular acrodermatitis presents as a symmetric erythematous lichenoid papular and papulovesicular eruption of the face, extremities, and buttocks, usually occurring in young children. GCS has been associated with hepatitis B and enteroviruses, as well as Epstein-Barr virus (EBV) and, rarely, cytomegalovirus. OBJECTIVE: The purpose of this study was to use immunohistochemical studies to determine the pattern of the lymphoid infiltrate and evidence for viral antigens in cases of EBV-associated GCS. METHODS: Routine histologic and immunohistochemical stains were evaluated in 3 patients with typical GCS. All 3 patients showed serologic evidence of an acute EBV infection. The immunohistochemical studies included monoclonal antibodies for CD3, CD4, CD8, CD20, TIA, S-100 protein, KP-1, EBV latent membrane antigen-1, and EBV-encoded nuclear antigen-2. RESULTS: All biopsy specimens showed minimal epidermal spongiosis with marked papillary dermal edema. The associated inflammatory infiltrate showed a mixed mononuclear cell infiltrate with rare eosinophils. Immunohistochemical stains for latent membrane antigen-1 and EBV encoded nuclear antigen-2 were negative for EBV. The majority of mononuclear cells showed membrane staining for CD3, 30% to 40% of the CD3 mononuclear cells showed positive staining for CD4, and 50% to 60% showed positive staining with CD8. TIA(+) cells appeared to correspond to the CD8(+) cells. CONCLUSION: Although papillary dermal edema has been reported within the spectrum of histologic findings in GCS, it was marked and a consistent finding in the 3 cases in which EBV was the most likely etiologic agent. The presence of large numbers of cytotoxic T cells in the inflammatory infiltrate may have accentuated this histologic finding and may be a relatively distinctive histologic finding with GCS associated with EBV. PMID- 11100027 TI - Treatment of Bowen's disease of the digit with carbon dioxide laser. AB - BACKGROUND: Therapy for Bowen's disease is essential to prevent invasive squamous cell carcinoma and metastases. Surgical excision is preferred because of the low relapse rate, but it may result in scar contracture, especially in the digit. OBJECTIVE: This study was undertaken to evaluate the effectiveness of carbon dioxide laser for Bowen's disease of the digit. METHODS: Patients with biopsy proven Bowen's disease of the digits were recruited from the Department of Dermatology, Siriraj Hospital during the period 1992-1998. Risk factors were assessed. The lesion and 4 mm of surrounding clinically uninvolved skin were vaporized by means of the carbon dioxide laser. Wounds were managed by second intention. Patients were followed up at 1 week, 4 weeks, 6 months, and yearly thereafter if applicable. RESULTS: During this 7-year period, only 6 patients with Bowen's disease of the digits were identified. Four of the 6 had a long history of exposure to arsenic from herbal medicines. Postoperative clinical appearance was mild atrophic skin and hypopigmentation. No functional impairment was found. Biopsy specimens taken after treatment showed a slightly thinned epidermis and mild fibrosis in the papillary dermis. There has been no evidence of recurrence in the 6 months to 7.7 years since surgery. CONCLUSION: At least 67% of the patients affected by Bowen's disease of the digits had chronic arsenicism. It may be the earliest presentation of chronic arsenicism. Carbon dioxide laser provides ease of surgery, excellent cosmesis, preservation of function, and effective treatment for patients with Bowen's disease of the digits. PMID- 11100028 TI - Allergists and dermatologists have far more expertise in caring for patients with urticaria than other specialists. AB - BACKGROUND: Urticaria is a common disease for which numerous treatments have been described, yet there is little information about what agents are commonly used to treat urticaria. There may be differences in the way in which urticaria is treated by different medical specialties. OBJECTIVE: The purpose of this study was to characterize the visits and treatments of urticaria in office-based practices. METHODS: National Ambulatory Medical Care Survey data from 1990 to 1997 were analyzed to determine patient populations, medications used, and physician specialties for visits of urticaria. RESULTS: Women accounted for 69% of all patient visits, but an equal gender distribution was observed in patients 18 years of age and younger. There was a bimodal age distribution with peak visits in patients aged birth to 9 years and 30 to 40 years. H(1) antihistamines and systemic corticosteroids were used in 56% and 14% of visits, respectively. Other medications reported as useful in the treatment of urticaria were used in 12% of visits. Allergists and dermatologists had a mean of 47 and 37 visits per physician per year, respectively, compared with all other physicians who averaged fewer than 10 visits per physician per year. Allergists were the least likely to use a corticosteroid agent (6% of visits), whereas internists were the most likely (29% of visits). Dermatology and allergy recorded a relatively large percentage of visits for urticaria that were referred for their condition by other physicians (49% and 25% of visits, respectively). CONCLUSION: We observed a bimodal utilization curve for age and urticaria not previously described. H(1) antihistamines remain the mainstay in treatment of urticaria, whereas the low use of systemic corticosteroids likely reflects physicians' understanding of their secondary function in the treatment of urticaria. PMID- 11100029 TI - Nevoid basal cell carcinoma syndrome. PMID- 11100030 TI - Immunohistochemical stains in dermatopathology. PMID- 11100031 TI - A visit to Eden: living and working at the regional dermatology training center in Tanzania. PMID- 11100032 TI - Surgical pearl: repair of helical rim defects with the bipedicle advancement flap. PMID- 11100033 TI - Acral peeling skin syndrome. AB - Peeling skin syndrome is a rare autosomal recessive disease characterized by widespread painless peeling of the skin in superficial sheets. We describe a 34 year-old man with a lifelong history of spontaneous asymptomatic peeling skin limited to the acral surfaces. This patient probably represents a localized variant of peeling skin syndrome, which has previously been described as a generalized condition. Light and electron microscopic studies of biopsy specimens taken before and after immersion in water were performed. It was concluded that this patient has abnormal keratohyalin granules and inadequate aggregation of keratin filaments that caused the separation of the epidermis in the stratum corneum through the clear zone. Alternatively, unknown keratin species expressed in the clear zone may also cause the abnormality. (J Am Acad Dermatol 2000;43:1112-9.). PMID- 11100034 TI - Parotid oncocytoma in the Birt-Hogg-Dube syndrome. AB - Birt-Hogg-Dube syndrome consists of fibrofolliculomas, trichodiscomas, and acrochordons and has been associated with several noncutaneous tumors. We report the first case of Birt-Hogg-Dube syndrome in association with a parotid oncocytoma, a rare salivary gland tumor. PMID- 11100035 TI - Central white scarlike patch: a dermatoscopic clue for the diagnosis of dermatofibroma. AB - In this study, dermatoscopic examination of 24 dermatofibromas was performed to evaluate specific dermoscopic criteria. A central white scarlike patch was appreciable in 22 of 24 lesions, whereas 20 of 24 dermatofibromas exhibited a delicate pigment network at the periphery. This stereotypical dermatoscopic finding allowed the diagnosis of dermatofibroma in most instances. (J Am Acad Dermatol 2000;43:1123-5.). PMID- 11100036 TI - CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy): diagnostic skin biopsy changes determined by electron microscopy. AB - Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a familial vascular disorder associated with migraines, recurrent ischemic strokes, and early-onset multiinfarct dementia. The diagnosis of CADASIL is made ultrastructurally by finding characteristic granular, electron-dense, osmiophilic material attached to vascular smooth muscle cells. These changes have been found in brain, skeletal muscle, nerve, and skin. We describe a woman with CADASIL diagnosed on the basis of brain and skin electronmicroscopic findings. Dermatologists and dermatopathologists need to be aware of this disorder because characteristic electronmicroscopic changes on a skin biopsy specimen are diagnostic. PMID- 11100037 TI - CADASIL: the dermatologic diagnosis of a neurologic disease. Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy. AB - Cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is an increasingly recognized neurologic disease characterized by pathognomonic changes to the small vessels, particularly in the brain and skin. Although much has recently been written about this disease in the neuropathology literature, to our knowledge nothing has appeared in the dermatology literature. We wish to call attention to the unique role dermatologists and dermatopathologists can play in the diagnosis of this disease. We review the condition's clinical, histologic, and ultrastructural features. PMID- 11100038 TI - Transplacental passage of maternal pemphigus foliaceus autoantibodies induces neonatal pemphigus. AB - The association of maternal pemphigus foliaceus (PF) with neonatal PF is rare and may be secondary to transplacental passage of PF autoantibodies. We describe a 25 year-old patient with PF who was delivered of two consecutive babies, one with classic skin lesions of PF and another that was normal. The neonate with PF was born when the mother had widespread skin disease; the normal newborn was born when the mother was in partial remission. The titers of PF autoantibodies were higher in the mother's serum and the cord serum of the baby with PF than in the mother during partial remission and the unaffected baby. The mother and affected baby had autoantibodies to desmoglein 1. Furthermore, cord blood from the baby with PF induced skin disease when injected into mice. In this case, maternal PF was associated with neonatal PF when the titers of maternal anti-desmoglein 1 autoantibodies were elevated. The cutaneous disease in neonatal PF is due to anti desmoglein 1 autoantibodies. PMID- 11100039 TI - Treatment of a subungual hemangioma with flashlamp-pumped pulsed-dye laser. PMID- 11100040 TI - Plexiform neurofibroma with massive hand involvement. PMID- 11100041 TI - Photopheresis for systemic sclerosis: evidence? PMID- 11100043 TI - Evaluating laboratory performance with the six sigma scale. PMID- 11100045 TI - Evaluating laboratory performance with the six sigma scale. PMID- 11100047 TI - Hyperplastic mesothelial cells in mediastinal lymph node sinuses. PMID- 11100049 TI - The legacy of William Heberden. PMID- 11100050 TI - The legacy of william heberden PMID- 11100052 TI - Succinate dehydrogenase deficiency. AB - BACKGROUND: Partial succinate dehydrogenase deficiency (15% to 50% of normal reference enzyme activity) in skeletal muscle causes mitochondrial myopathy with various symptoms, for example, brain involvement, cardiomyopathy, and/or exercise intolerance. The deficiency may be isolated or may coexist with other respiratory chain enzyme defects. The histopathologic assessment of succinate dehydrogenase activity in muscle biopsies of patients with suspected mitochondrial myopathies has focused on the finding of increased staining, usually in ragged-red fibers, rather than on reduced staining. OBJECTIVES: To determine the prevalence of muscle succinate dehydrogenase deficiency among patients with respiratory-chain defects and to determine whether the reduced activity is present histochemically and is comparable to the quantitative reduction found in muscle homogenates. PATIENTS AND METHODS: One hundred eight muscle biopsies were evaluated from patients with suspected mitochondrial myopathies by qualitative histochemical analysis and quantitative biochemical analyses of respiratory-chain enzymes using standard methodologies. RESULTS: Fifty-two patients had defects in respiratory chain complexes; of these patients, 12 (23%) had partial deficiencies in succinate dehydrogenase activity either alone or together with reductions in other enzymes. The reduced activity was detectable histochemically in muscle biopsies with residual enzyme activity of up to 34% of the normal reference activity, while 2 biopsies with higher residual activity (49% and 68% of normal) could not be distinguished from normal biopsies. CONCLUSIONS: Of the patients with respiratory-chain enzyme defects, 23% had partial deficiencies of succinate dehydrogenase activity in muscle biopsies. This reduction could be detected histochemically in biopsies in most cases. The marked prevalence of succinate dehydrogenase deficiency among patients with respiratory-chain defects and its detection initially by histochemical analysis are important findings. PMID- 11100053 TI - Pathologic evaluation of sentinel lymph nodes in colorectal carcinoma. AB - BACKGROUND: The identification of lymph node metastases in colorectal resection specimens is necessary for accurate tumor staging. However, routine lymph node dissection by the pathologist yields only a subset of nodes removed surgically and may not include those nodes most directly in the path of lymphatic drainage from the tumor. Intraoperative mapping of such sentinel lymph nodes (SLNs) has been reported in cases of melanoma and breast cancer. We applied a similar method to cases of colorectal carcinoma, with emphasis on the pathology of the SLNs. METHODS: Eighty-three consecutive patients with colorectal carcinoma were evaluated after intraoperative injection of 1 to 2 mL of 1% isosulfan blue dye (Lymphazurin) into the peritumoral subserosa. Blue-stained lymph nodes were suture-tagged by the surgeon within minutes of the injection for identification by the pathologist, and a standard resection was performed. Designated SLNs were sectioned at 10 levels through the block; a cytokeratin immunostain (AE1) was also obtained. To evaluate the possibility that increased detection of metastases in the SLN might be solely due to increased histologic sampling, all initially negative non-SLNs in the first 25 cases were sectioned also at 10 levels. RESULTS: Sentinel lymph nodes were identified intraoperatively in 82 (99%) of 83 patients and accounted for 152 (11.9%) of 1275 lymph nodes recovered, with an average of 1.9 SLNs per patient. A total of 99 positive lymph nodes (38 positive SLNs and 61 positive non-SLNs) were identified in 34 node-positive patients. The SLNs were the only site of metastasis in 17 patients (50%), while 14 patients (41%) had both positive SLNs and non-SLNs. Three patients (9%) had positive non SLNs with negative SLNs, representing skip metastases. In patients with positive SLNs, 91 (19%) of 474 total lymph nodes and 53 (12%) of 436 non-SLNs were positive for metastasis. In patients with negative SLNs, 8 (1%) of 801 total lymph nodes and 8 (1.2%) of 687 non-SLNs were positive for metastasis. Multilevel sections of 330 initially negative non-SLNs in the first 25 patients yielded only 2 additional positive nodes (0. 6%). All patients with positive SLNs were correctly staged by a combination of 4 representative levels through the SLN(s) together with a single cytokeratin immunostain. CONCLUSIONS: Intraoperative mapping of SLNs in colorectal carcinoma identifies lymph nodes likely to contain metastases. Focused pathologic evaluation of the 1 to 4 SLNs so identified can improve the accuracy of pathologic staging. PMID- 11100054 TI - Loss of high-molecular-weight cytokeratin antigenicity in prostate tissue obtained by transurethral resections. AB - OBJECTIVE: Staining of prostatic basal cells for the expression of high-molecular weight cytokeratin has been suggested as a way of distinguishing benign from malignant prostate glands. We evaluated the utility of high-molecular-weight cytokeratin in the diagnosis of malignancy in prostate specimens obtained in various ways. DESIGN: Prostate tissues obtained from needle biopsies, transurethral resections, and total prostatectomies were immunostained with monoclonal antibody 34betaE12, an antibody directed against high-molecular-weight cytokeratins. RESULTS: Antiserum to high-molecular-weight cytokeratin only stained the basal cells in normal glands in 3 (12%) of 25 specimens obtained by transurethral resection. Other antigens, such as the alternate 10-nm filament protein vimentin, were unaffected and were detected in 100% of these specimens. However, keratin antigenicity in transurethral biopsies could be restored in these specimens by antigen retrieval in a low pH citrate buffer using a microwave heat technique. Keratin staining in needle biopsies and total prostatectomies was unaffected. CONCLUSION: In summary, our results indicate the technique of transurethral resection results in a specific loss of keratin antigenicity. This limits the utility of anticytokeratin 34betaE12 in interpreting transurethral resections without the application of antigen retrieval. PMID- 11100055 TI - M30 expression demonstrates apoptotic cells, correlates with in situ end labeling, and is associated with Ki-67 expression in large intestinal neoplasms. AB - CONTEXT: The monoclonal antibody M30 recognizes a neoepitope of cytokeratin 18 produced during apoptosis. It is reactive in formalin-fixed, paraffin-embedded tissue and has great potential in the study of apoptosis in clinical and experimental material. OBJECTIVES: To compare the results of M30 immunoexpression with a more established technique of demonstrating apoptosis in tissue sections, in situ end-labeling. A secondary objective was to compare the results with immunoexpression of the proliferation-associated antigen Ki-67. DESIGN: Retrospective analysis of adenomas and adenocarcinomas of the large intestine. INTERVENTIONS: Immunohistochemistry for M30 and Ki-67, and in situ end-labeling. Formalin-fixed, paraffin-embedded tissue was used. MAIN OUTCOME MEASURES: The number of cells positive for M30, Ki-67, and in situ end-labeling, expressed as a proportion of the total number of cells counted. RESULTS: A strong positive correlation was found between in situ end-labeling and expression of M30, although the counts were widely scattered around the regression line. Counts of Ki-67 were strongly correlated with both M30 expression and in situ end-labeling. Immunoexpression of M30 was generally easier to interpret than in situ end labeling, and the procedures for M30 immunohistochemistry were technically less exacting. CONCLUSION: These findings support the application of M30 immunoreactivity in the study of apoptosis. PMID- 11100056 TI - Characteristics of salivary diffuse infiltrative lymphocytosis syndrome in West Africa. AB - OBJECTIVE: To determine the prevalence of diffuse infiltrative lymphocytosis syndrome (DILS) in the minor salivary glands of 30 African Cameroonian adults with the acquired immunodeficiency syndrome (AIDS). DESIGN: Salivary gland tissue was analyzed using a modified classification system that was developed to aid the diagnosis of Sjogren syndrome. The advantages and disadvantages of this approach are discussed. MATERIALS AND METHODS: Formalin-fixed, paraffin-embedded, hematoxylin-eosin-stained biopsy sections were prepared for 30 patients with AIDS, 26 healthy individuals who declined human immunodeficiency virus (HIV) testing, and 4 seronegative healthy controls. Tissues were immunostained for CD4/CD8+ lymphocytes and cytomegalovirus (CMV), and transmission electron microscopy was performed to locate viral particles. Patients were tested for HIV 1 and HIV-2 by the HIV/Chek System 3 or CAMSTIX-HIV-1 and HIV-2 assay. RESULTS: Severe salivary ductal atypia (96%) was the feature most strongly associated with AIDS, and the lymphocytic focus score was the second histologic feature most strongly correlated with AIDS. Forty-eight percent of patients with HIV-1 infection had more than 1 lymphocytic focus in a minor salivary gland. These lymphocytes were primarily CD8+. We report, to the best of our knowledge, the first case of multinucleated salivary duct epithelial cells in minor salivary glands also containing enveloped virus particles. All cases were negative for CMV. CONCLUSIONS: The prevalence of DILS in West Africans with AIDS appears higher than the prevalence reported in whites from the United States and Europe and in blacks from the United States, a group that has been reported to have a greater incidence of DILS than whites. This discrepancy may be related to differences in patient selection criteria. The determination of lymphocytic focus score, as used in the diagnosis of Sjogren syndrome, with the adjunct of ductal atypia is useful for assessing DILS. The impact of patient selection, drug therapy, and parasites on salivary gland pathology is discussed. PMID- 11100057 TI - Parotid gland malignant melanomas. AB - BACKGROUND: Malignant melanomas are relatively unusual tumors in the parotid gland. The majority of previously reported cases appear to represent metastatic lesions, often from cutaneous head and neck primaries. METHODS: Retrospective clinicopathologic review of 12 cases of malignant melanoma involving the parotid gland encountered between 1980 and October 1999 at a tertiary referral center. RESULTS: Patients consisted of 9 men and 3 women ranging in age from 30 to 84 years (median, 66 years). Eleven of 12 patients presented with a neck mass or nodule. In 9 of 12 patients, a cutaneous or conjunctival primary was noted in the head region. In 2 patients, a cutaneous melanoma and the parotid gland melanoma were diagnosed at the same time. In 1 patient, melanoma was initially diagnosed in the parotid gland, and a definite primary was not uncovered. All patients underwent excision of the parotid melanoma, which was accompanied by a lymph node biopsy or dissection in 10 out of 11 patients. Four patients received adjuvant radiotherapy, and 3 patients received adjuvant chemotherapy. Four of 11 patients had ipsilateral cervical lymph node metastasis at the time of parotid tumor resection, and 5 patients had involvement of intraparotid lymph nodes by metastatic melanoma. Tumors ranged in size from 0.3 to 2.5 cm in greatest dimension. Multiple parotid nodules were noted in 4 patients. All tumors were characterized by a diffuse proliferation of cells with abundant eosinophilic cytoplasm and prominent nucleoli. Four tumors demonstrated focal spindle cell regions. Intravascular and/or lymphatic involvement by tumor within the parotid gland was noted in 3 lesions. At last known follow-up, 6 patients had died with tumor at a median follow-up period of 11 months after parotid gland surgery. Four patients were alive with evidence of tumor at follow-up intervals of 4, 17, 21, and 113 months after parotid gland surgery. Two patients were alive with no evidence of residual tumor at 20 and 148 months of follow-up. CONCLUSIONS: The majority of melanomas involving the parotid gland appeared to be associated with lymph node metastasis in and around the gland from a cutaneous primary in the head region. Prognosis is generally poor, although rare patients may survive a long period of time following surgery. PMID- 11100058 TI - Placental lesions associated with cerebral palsy and neurologic impairment following term birth. AB - OBJECTIVE: The aim of this study was to determine the association of placental findings with cerebral palsy and related forms of neurologic impairment (NI) following birth at > or =37 weeks gestation (term). DESIGN: In a retrospective comparison, placentas from 40 term infants with NI ascertained on the basis of clinicopathologic review for medicolegal consultation were compared with placentas from 176 consecutive meconium-stained term infants at low risk for NI. RESULTS: After stratification for severity, 9 lesions were significantly increased in placentas from infants with NI: 5 lesions generally considered to occur within days of the time of labor and delivery (meconium-associated vascular necrosis, severe fetal chorioamnionitis, chorionic vessel thrombi, increased nucleated red blood cells, and findings consistent with abruptio placenta) and 4 lesions generally believed to have their onset long before labor and delivery (diffuse chronic villitis, extensive avascular villi, diffuse chorioamnionic hemosiderosis, and perivillous fibrin). Findings independently associated with NI by logistic regression in this descriptive study were severe fetal chorioamnionitis (odds ratio [OR], 13.2; 95% confidence interval [CI], 1.2-144); extensive avascular villi (OR, 9.0; 95% CI, 1.6-51); and diffuse chorioamnionic hemosiderosis (OR, 74.8; 95% CI, 6.3-894). The risk of NI increased as a function of the number of lesions present (OR, 10.1; 95% CI, 5.1-20 for each additional lesion), particularly when lesions generally considered to occur near the time of labor and those believed to occur well before labor were found in the same placenta (OR, 94.2; 95% CI, 11.9-747). CONCLUSIONS: These findings suggest that placental pathology can contribute to an understanding of the mechanisms that contribute to NI at term. PMID- 11100059 TI - Utility of flow cytometry immunophenotyping for the diagnosis and classification of lymphoma in community hospital clinical needle aspiration/biopsies. AB - CONTEXT: Flow cytometry immunophenotyping (FC) of needle aspiration/biopsy (NAB) samples has been reported to be useful for the diagnosis and classification of lymphoma in university and cancer center-based settings. Nevertheless, there is no agreement on the utility of these methods. OBJECTIVE: To further define the utility of adjunctive FC of clinical NAB for the diagnosis and classification of lymphoma, and to determine if this approach is practicable in a routine clinical practice setting. SETTING: A community-based hospital. METHODS: Clinical NABs were submitted for adjunctive FC between June 1996 and September 1999 if initial smears were suspicious for lymphoma. Smears and cell block or needle core tissues were routinely processed and paraffin-section immunostains were performed if indicated. The final diagnosis was determined by correlating clinical and pathologic data, and the revised European-American classification criteria were used to subtype lymphomas. RESULTS: Needle aspiration/biopsies from 60 different patients were submitted for FC. Final diagnoses were lymphoma (n = 38), other neoplasm (n = 15), benign (n = 6), or insufficient (n = 1). For 38 lymphomas (20 primary, 18 recurrent), patients ranged in age from 32 to 86 years (mean, 62 years); samples were obtained from the retroperitoneum (n = 11), lymph node (n = 9), abdomen (n = 8), mediastinum (n = 6), or other site (n = 4); and lymphoma subtypes were indolent B-cell (n = 20; 2 small lymphocytic, 14 follicle center, 4 not subtyped), aggressive B-cell (n = 14; 3 mantle cell, 10 large cell, 1 not subtyped), B-cell not further specified (n = 2), or Hodgkin disease (n = 2). For the diagnosis of these lymphomas, FC was necessary in 20 cases, useful in 14 cases, not useful in 2 cases, and misleading in 2 cases. Thirty-two of 36 lymphoma patients with follow-up data received antitumor therapy based on the results of NAB plus FC. CONCLUSIONS: Adjunctive FC of NABs is potentially practicable in a community hospital, is necessary or useful for the diagnosis and subtyping of most B-cell lymphomas, and can help direct lymphoma therapy. Repeated NAB or surgical biopsy is necessary for diagnosis or treatment in some cases. PMID- 11100060 TI - Acute fulminant hepatic failure in a woman treated with phenytoin and trimethoprim-sulfamethoxazole. AB - Massive hepatic necrosis following exposure to phenytoin and trimethoprim sulfamethoxazole is a rare occurrence and to the best of our knowledge has not been reported previously. Acute hepatic failure following administration of trimethoprim-sulfamethoxazole has rarely been seen, and only 4 cases have been well documented pathologically. We report a case of acute liver failure in a 60 year-old woman following ingestion of phenytoin and trimethoprim-sulfamethoxazole concomitantly over a 9-day period. Autopsy findings revealed acute fulminant hepatic failure. This case demonstrates the effects of chemical-chemical interactions in the potentiation of hepatotoxicity of single agents and specifically illustrates the need for discontinuing trimethoprim-sulfamethoxazole in the presence of early liver injury. PMID- 11100061 TI - CD13-positive anaplastic large cell lymphoma of T-cell origin--a diagnostic and histogenetic problem. AB - The expression of myelomonocytic-associated antigens in anaplastic large cell lymphomas (ALCLs), particularly those presenting in extranodal sites, can make their distinction from extramedullary myeloid cell tumors (EMCTs) or histiocytic tumors problematic. Yet, this distinction is clinically significant because of its therapeutic and prognostic implications. Herein, we describe a case of extranodal anaplastic lymphoma kinase-positive CD30-positive ALCL of T-cell origin in a 12-year-old boy, which was initially called an EMCT because of the expression of CD13 and HLA-DR detected by flow cytometry and the absence of other T-cell-related surface markers. However, the detection of cytoplasmic CD3 by flow cytometry prompted further studies. The tumor was composed of large cells with abundant slightly eosinophilic vacuolated cytoplasm and ovoid or reniform nuclei with a few small nucleoli. Using immunohistochemistry, the tumor was positive for CD45, CD30, CD45RO, and CD43 with a strong cytoplasmic and nuclear anaplastic lymphoma kinase stain. The tumor cells showed a T-cell clonal genotype. Electron microscopy revealed no ultrastructural features of myelomonocytic or histiocytic origin. The patient responded well to the chemotherapy and was in complete remission for 10 months at the time of submission of this manuscript. Review of the literature showed inconsistencies regarding the diagnosis, nomenclature, and, therefore, treatment and prognosis of these tumors. In addition, the CD13 expression in ALCL raises some histogenetic questions and may indicate origin from a pluripotent stem cell, misprogramming during malignant transformation, or a microenvironmental effect on lymphoid cell expression of surface antigens. Therefore, ALCL should be considered in the differential diagnosis of EMCTs or histiocytic tumors, particularly when surface marker lineage assignment is ambiguous. PMID- 11100062 TI - Herpes simplex encephalitis occurring after chemotherapy, surgery, and stereotactic radiotherapy for medulloblastoma. AB - Reactivation of latent herpes simplex virus (HSV) in the trigeminal ganglion most commonly gives rise to recurrent herpes labialis and rarely to herpes simplex encephalitis. The mechanisms underlying reactivation of latent trigeminal HSV are complex. Here we report the case history of a 25-year-old woman who developed a fatal, bilateral necrotizing destructive temporal lobe lesion following surgical removal of a cerebellar medulloblastoma and combined radiotherapy and chemotherapy for recurrent tumor. Neuropathologic examination of the brain revealed minimal inflammatory changes, but immunohistochemistry was positive for HSV protein, and HSV deoxyribonucleic acid (DNA) was recovered from formalin fixed paraffin-embedded brain tissue. The temporal proximity of the surgery, chemotherapy, and radiotherapy to the onset of disease suggests that these factors may have acted as triggers that precipitated conversion of latent HSV to overt HSV. PMID- 11100063 TI - Cholesterol granulomas of the lungs associated with microangiopathic hemolytic anemia and thrombocytopenia in pulmonary hypertension. AB - Cholesterol granulomas unrelated to endogenous lipoid pneumonia, pulmonary alveolar proteinosis, or cholesterol pneumonia are a rare finding during pneumectomy or autopsy. They have been occasionally reported in association with pulmonary hypertension. We report a case where these lesions were associated with long-standing pulmonary hypertension and microangiopathic hemolytic anemia and thrombocytopenia. Plexiform lesions were present in the pulmonary vasculature secondary to pulmonary hypertension, causing hemolysis and thrombocytopenia. We suggest that destruction of red blood cells and platelets could provide membrane lipids that are taken up by phagocytic cells, which promotes the formation of these cholesterol deposits. PMID- 11100064 TI - Primary nodal marginal zone B-cell lymphoma arising from more than one clonal neoplastic population. AB - Primary nodal marginal zone B-cell lymphoma is an uncommon monoclonal B-cell lymphoproliferative disorder. We report a case of a 79-year-old woman who presented with generalized lymphadenopathy. Histologic and immunohistochemical examinations of biopsy sections from an axillary lymph node were consistent with nodal marginal zone B-cell lymphoma. Flow cytometry analysis showed 2 distinct clonal B-cell populations expressing lambda or kappa light chain restriction. Subsequently, genomic deoxyribonucleic acid (DNA) isolated from a paraffin embedded lymph node section was analyzed for the presence of gene rearrangements. Polymerase chain reaction (PCR) analysis of immunoglobulin heavy chain genes revealed 3 rearranged DNA bands, confirming the presence of more than one clonal B-cell population. These immunophenotypic and genotypic findings have not been previously described in association with this type of lymphoma. To our knowledge, this represents the first reported case of biclonal nodal marginal zone B-cell lymphoma. PMID- 11100065 TI - Papillary glioneuronal tumor. AB - Tumors of mixed glioneuronal type are well recognized in the central nervous system. The most common examples of these lesions include gangliogliomas and dysembryoplastic neuroepithelial tumors. Recently, unusual examples of these lesions have been described, including the papillary glioneuronal tumor. This report describes a histologically similar-appearing lesion arising in the left parieto-occipital lobe of an 18-year-old man who presented with headaches and difficulties with vision. Imaging studies noted a large cystic neoplasm marked by a peripheral rim of enhancement. The patient underwent gross total resection of the tumor, which histologically was marked by a mixture of glial (glial fibrillary acidic protein-positive) and neural (synaptophysin-positive) components. Architecturally, the tumor was notable for a focal pseudopapillary pattern. Papillae were lined by predominantly glial cells, with intervening areas occupied by neurally differentiated cells. Mitotic activity, vascular proliferation, and necrosis were not noted. A MIB-1 labeling index of 1.1% was seen. p53 immunoreactivity was not observed. This report adds further evidence supporting the existence of this unusual mixed glioneuronal tumor of the central nervous system. PMID- 11100066 TI - Primary localized extranodal hodgkin disease of the transverse colon. AB - Extranodal Hodgkin disease presenting as a primary localized neoplasm is uncommon, with rare case reports describing primary sites other than lymph nodes. The gastrointestinal tract is the most frequent site of involvement by extranodal Hodgkin disease, typically involving the stomach or small bowel. To date, we have been able to find only one fully documented case of Hodgkin disease of the sigmoid colon confirmed by immunohistochemical studies. We report a case of extranodal Hodgkin disease involving the transverse colon, presenting as inflammatory bowel disease and documented by light microscopic, immunohistochemical, cytogenetic, and molecular studies. PMID- 11100067 TI - Malakoplakia of the appendix. An unusual association with eggs of Taenia species. AB - Malakoplakia of the appendix is an unusual condition that has been reported to occur in association with tumors, infections, and immunocompromised states. We describe a case of appendicular malakoplakia associated with eggs of Taenia species. The diagnosis was made on histopathologic examination of surgically resected tissue from an appendicular mass. To the best of our knowledge, this is the first time that helminths have been documented to be associated with malakoplakia. We also discuss the implications of helminthic infestation in the pathogenesis of the lesion. PMID- 11100068 TI - Sarcomatoid renal cell carcinoma of papillary origin. A case report and cytogenic evaluation. AB - Sarcomatoid renal cell carcinoma (SRCC) is an aggressive tumor variant thought to arise predominantly from dedifferentiation of clear cell carcinoma. A few reports of SRCC associated with non-clear cell tumors led to the presumption that SRCC may arise from any renal cell carcinoma, although direct evidence of this is lacking. Cytogenetic studies on 3 previously documented SRCCs associated with papillary renal cancers showed either 3p deletions or absence of trisomy 7, 17 in the sarcomatoid tumors, suggesting origin from a coexistent clear cell tumor. The present case represents the first conclusive evidence of direct progression of non-clear cell carcinoma to SRCC with both tumor components containing multiple copies of chromosomes 7 and 17. Many genetic anomalies, including p53 mutations, frequently recognized in SRCC were not recognized in this case, highlighting the importance of cytogenetic evaluation of all SRCC. The patient is well and without evidence of tumor progression 1 year after surgery, and the sinister outlook of SRCC in association with clear cell carcinoma may not apply in SRCC of non-clear cell origin. PMID- 11100069 TI - Laryngeal metastasis 7 years after radical nephrectomy. AB - Metastasis of renal cell carcinoma to the head and neck, especially the larynx, is an extremely rare event. Most previously reported cases have involved a presenting laryngeal lesion that lead to the discovery of an underlying primary renal cell carcinoma. Even more unusual is the occurrence of an isolated laryngeal metastasis revealing itself years after nephrectomy, with an interim of undetected recurrence. We believe this case to be the first reported example of an isolated supraglottic laryngeal renal cell carcinoma metastasis occurring 7 years after radical nephrectomy. Local excision of such isolated lesions seems to offer a favorable prognosis. PMID- 11100070 TI - Pathologic quiz case. Acinic cell adenocarcinoma of accessory right parotid gland. PMID- 11100071 TI - Pathologic quiz case. Syncytial variant nodular sclerosing Hodgkin disease. PMID- 11100072 TI - Pathologic quiz case. Intra-abdominal desmoplastic small round cell tumor. PMID- 11100073 TI - Pathologic quiz case. Pulmonary manifestations of tuberous sclerosis. PMID- 11100074 TI - Pathologic quiz case. Pulmonary infestation by Paragonimus westermani. PMID- 11100075 TI - Pathologic quiz case. Acute lymphoblastic leukemia. PMID- 11100076 TI - Nasal glial heterotopia. PMID- 11100081 TI - Pathology PMID- 11100082 TI - Tumors of the testis, adnexa, spermatic cord, and scrotum PMID- 11100084 TI - Atlas of gastrointestinal endoscopy and endoscopic biopsies PMID- 11100083 TI - Viral infections of the nervous system PMID- 11100085 TI - Science and politics-a broken union. PMID- 11100086 TI - Reply to "The 'African enigma' - another explanation" PMID- 11100087 TI - The 'African enigma' - another explanation. PMID- 11100088 TI - Varicella vaccine revisited. PMID- 11100089 TI - Varicella vaccine revisited. PMID- 11100090 TI - Reply to "Varicella vaccine revisited" PMID- 11100091 TI - Europe increases precautions against BSE transmission. PMID- 11100092 TI - US monitors TSE in livestock. PMID- 11100093 TI - NIH sues BMS over missing drug royalties. PMID- 11100094 TI - Company disputes scientist's right to publication. PMID- 11100095 TI - UK tries to increase physician scientist numbers PMID- 11100096 TI - Gates foundation hires preeminent scientist PMID- 11100097 TI - Plans for stem cell research could be blocked. PMID- 11100098 TI - New database could lead to antibiotic overuse. PMID- 11100099 TI - New bioterrorism institute enters burgeoning field. PMID- 11100101 TI - Finally, a plan to reduce measles deaths PMID- 11100100 TI - Female japanese scientist wins harassment case PMID- 11100102 TI - NIH spending in developing countries increases. PMID- 11100103 TI - A crisis of trust: for science, scientists or for institutions? PMID- 11100104 TI - AIDS doctors: voices from the epidemic PMID- 11100105 TI - Immunomodulation: a new role for statins? PMID- 11100106 TI - Cholesterol esters and atherosclerosis-a game of ACAT and mouse. PMID- 11100107 TI - Targeting tumors through the HIF system. PMID- 11100108 TI - From catheters to vectors: the dawn of molecular electrophysiology. PMID- 11100109 TI - Pointing (zinc) fingers at BRCA1 targets. PMID- 11100110 TI - Oxidation versus aggregation - how do SOD1 mutants cause ALS? PMID- 11100111 TI - In living color. PMID- 11100112 TI - Will we have and why do we need an Ebola vaccine? PMID- 11100113 TI - Kissing cousins-evidence for a common vascular cell precursor. PMID- 11100114 TI - Research news PMID- 11100115 TI - Latent Mycobacterium tuberculosis-persistence, patience, and winning by waiting. AB - Mycobacterium tuberculosis is a globally successful pathogen due to its ability to persist for long periods of time unrecognized by the human immune system. The panoply of genes that allows the organism to enter latency and then re-emerge later during endogenous reinfection are now being elucidated. Novel antimicrobials and vaccines will need to target these mycobacterial pathogenic mechanisms to suceed against tuberculosis. PMID- 11100116 TI - In vivo veritas: the search for TB drug targets goes live. AB - The term 'microbial persistence' describes a phenomenon whereby microorganisms which are drug-susceptible when tested outside the body are nevertheless capable of surviving within the body despite intensive therapy with the appropriate antimicrobial drug. In clinical practice this phenomenon obviously has to do with the post-treatment 'carrier state' and with post-treatment relapse. In short, it is this phenomenon which is responsible for our inability to eradicate an infection from a person or a community by the use of drugs. - Walsh McDermott, The Yale Journal of Biology and Medicine 30, 257 (1958). PMID- 11100117 TI - Suppression of tumor growth through disruption of hypoxia-inducible transcription. AB - Chronic hypoxia, a hallmark of many tumors, is associated with angiogenesis and tumor progression. Strategies to treat tumors have been developed in which tumor cells are targeted with drugs or gene-therapy vectors specifically activated under hypoxic conditions. Here we report a different approach, in which the normal transcriptional response to hypoxia is selectively disrupted. Our data indicate that specific blockade of the interaction of hypoxia-inducible factor with the CH1 domain of its p300 and CREB binding protein transcriptional coactivators leads to attenuation of hypoxia-inducible gene expression and diminution of tumor growth. Thus, disrupting the normal co-activational response to hypoxia may be a new and useful therapeutic strategy. PMID- 11100118 TI - Resistance to diet-induced hypercholesterolemia and gallstone formation in ACAT2 deficient mice. AB - The importance of cholesterol ester synthesis by acyl CoA:cholesterol acyltransferase (ACAT) enzymes in intestinal and hepatic cholesterol metabolism has been unclear. We now demonstrate that ACAT2 is the major ACAT in mouse small intestine and liver, and suggest that ACAT2 deficiency has profound effects on cholesterol metabolism in mice fed a cholesterol-rich diet, including complete resistance to diet-induced hypercholesterolemia and cholesterol gallstone formation. The underlying mechanism involves the lack of cholesterol ester synthesis in the intestine and a resultant reduced capacity to absorb cholesterol. Our results indicate that ACAT2 has an important role in the response to dietary cholesterol, and suggest that ACAT2 inhibition may be a useful strategy for treating hypercholesterolemia or cholesterol gallstones. PMID- 11100119 TI - Efficient presentation of exogenous antigen by liver endothelial cells to CD8+ T cells results in antigen-specific T-cell tolerance. AB - Myeloid antigen-presenting cells (APC) are known to cross-present exogenous antigen on major histocompatibility class I molecules to CD8+ T cells and thereby induce protective immunity against infecting microorganisms. Here we report that liver sinusoidal endothelial cells (LSEC) are organ-resident, non-myeloid APC capable of cross-presenting soluble exogenous antigen to CD8+ T cells. Though LSEC employ similar molecular mechanisms for cross-presentation as dendritic cells, the outcome of cross-presentation by LSEC is CD8+ T cell tolerance rather than immunity. As uptake of circulating antigens into LSEC occurs efficiently in vivo, it is likely that cross-presentation by LSEC contributes to CD8+ T cell tolerance observed in situations where soluble antigen is present in the circulation. PMID- 11100120 TI - Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress. AB - Activation of the zinc-finger transcription factor early growth response (Egr)-1, initially linked to developmental processes, is shown here to function as a master switch activated by ischemia to trigger expression of pivotal regulators of inflammation, coagulation and vascular hyperpermeability. Chemokine, adhesion receptor, procoagulant and permeability-related genes are coordinately upregulated by rapid ischemia-mediated activation of Egr-1. Deletion of the gene encoding Egr-1 strikingly diminished expression of these mediators of vascular injury in a murine model of lung ischemia/reperfusion, and enhanced animal survival and organ function. Rapid activation of Egr-1 in response to oxygen deprivation primes the vasculature for dysfunction manifest during reperfusion. These studies define a central and unifying role for Egr-1 activation in the pathogenesis of ischemic tissue damage. PMID- 11100121 TI - In vivo delivery of the caveolin-1 scaffolding domain inhibits nitric oxide synthesis and reduces inflammation. AB - Caveolin-1, the primary coat protein of caveolae, has been implicated as a regulator of signal transduction through binding of its "scaffolding domain" to key signaling molecules. However, the physiological importance of caveolin-1 in regulating signaling has been difficult to distinguish from its traditional functions in caveolae assembly, transcytosis, and cholesterol transport. To directly address the importance of the caveolin scaffolding domain in vivo, we generated a chimeric peptide with a cellular internalization sequence fused to the caveolin-1 scaffolding domain (amino acids 82-101). The chimeric peptide was efficiently taken up into blood vessels and endothelial cells, resulting in selective inhibition of acetylcholine (Ach)-induced vasodilation and nitric oxide (NO) production, respectively. More importantly, systemic administration of the peptide to mice suppressed acute inflammation and vascular leak to the same extent as a glucocorticoid or an endothelial nitric oxide synthase (eNOS) inhibitor. These data imply that the caveolin-1 scaffolding domain can selectively regulate signal transduction to eNOS in endothelial cells and that small-molecule mimicry of this domain may provide a new therapeutic approach. PMID- 11100122 TI - The nuclear receptor co-repressor nrip1 (RIP140) is essential for female fertility. AB - Ovulatory dysfunction is the commonest cause of female infertility. Here we show that the co-repressor nuclear-receptor-interacting protein 1 (Nrip1; encoded by the gene Nrip1) is essential for ovulation. Mice null for this protein are viable, but female mice are infertile because of complete failure of mature follicles to release the oocyte at ovulation. In contrast, luteinization proceeds normally, resulting in a phenotype closely resembling that of luteinized unruptured follicle syndrome, often associated with infertility in women. Therefore, whereas the pre-ovulatory surge of luteinizing hormone induces both ovulation and luteinization, the ability to suppress the action of nuclear receptors is essential for the coordinated control of ovarian function with the essential process of oocyte release dependent on the activity of the transcriptional co-repressor Nrip1 (RIP40). PMID- 11100123 TI - The broad-spectrum antiviral ribonucleoside ribavirin is an RNA virus mutagen. AB - The ribonucleoside analog ribavirin (1-beta-D-ribofuranosyl-1,2, 4-triazole-3 carboxamide) shows antiviral activity against a variety of RNA viruses and is used in combination with interferon-alpha to treat hepatitis C virus infection. Here we show in vitro use of ribavirin triphosphate by a model viral RNA polymerase, poliovirus 3Dpol. Ribavirin incorporation is mutagenic, as it templates incorporation of cytidine and uridine with equal efficiency. Ribavirin reduces infectious poliovirus production to as little as 0. 00001% in cell culture. The antiviral activity of ribavirin correlates directly with its mutagenic activity. These data indicate that ribavirin forces the virus into 'error catastrophe'. Thus, mutagenic ribonucleosides may represent an important class of anti-RNA virus agents. PMID- 11100124 TI - Uptake of HIV-1 tat protein mediated by low-density lipoprotein receptor-related protein disrupts the neuronal metabolic balance of the receptor ligands. AB - Neurological disorders develop in most people infected with human immunodeficiency virus type 1 (HIV-1). However, the underlying mechanisms remain largely unknown. Here we report that binding of HIV-1 transactivator (Tat) protein to low-density lipoprotein receptor-related protein (LRP) promoted efficient uptake of Tat into neurons. LRP-mediated uptake of Tat was followed by translocation to the neuronal nucleus. Furthermore, the binding of Tat to LRP resulted in substantial inhibition of neuronal binding, uptake and degradation of physiological ligands for LRP, including alpha2-macroglobulin, apolipoprotein E4, amyloid precursor protein and amyloid beta-protein. In a model of macaques infected with a chimeric strain of simian-human immunodeficiency virus, increased staining of amyloid precursor protein was associated with Tat expression in the brains of simian-human immunodeficiency virus-infected macaques with encephalitis. These results indicate that HIV-1 Tat may mediate HIV-1-induced neuropathology through a pathway involving disruption of the metabolic balance of LRP ligands and direct activation of neuronal genes. PMID- 11100125 TI - Overexpression of the alpha1B-adrenergic receptor causes apoptotic neurodegeneration: multiple system atrophy. AB - Progress toward elucidating the function of alpha1B-adrenergic receptors (alpha1BARs) in the central nervous system has been constrained by a lack of agonists and antagonists with adequate alpha1B-specificity. We have obviated this constraint by generating transgenic mice engineered to overexpress either wild type or constitutively active alpha1BARs in tissues that normally express the receptor, including the brain. All transgenic lines showed granulovacular neurodegeneration, beginning in alpha1B-expressing domains of the brain and progressing with age to encompass all areas. The degeneration was apoptotic and did not occur in non-transgenic mice. Correspondingly, transgenic mice showed an age-progressive hindlimb disorder that was parkinsonian-like, as demonstrated by rescue of the dysfunction by 3, 4-dihydroxyphenylalanine and considerable dopaminergic-neuronal degeneration in the substantia nigra. Transgenic mice also had a grand mal seizure disorder accompanied by a corresponding dysplasia and neurodegeneration of the cerebral cortex. Both behavioral phenotypes (locomotor impairment and seizure) could be partially rescued with the alpha1AR antagonist terazosin, indicating that alpha1AR signaling participated directly in the pathology. Our results indicate that overstimulation of alpha1BAR leads to apoptotic neurodegeneration with a corresponding multiple system atrophy indicative of Shy-Drager syndrome, a disease whose etiology is unknown. PMID- 11100126 TI - Focal modification of electrical conduction in the heart by viral gene transfer. AB - Modern treatment of cardiac arrhythmias is limited to pharmacotherapy, radiofrequency ablation, or implantable devices. Antiarrhythmic medications suppress arrhythmias, but their systemic effects are often poorly tolerated and their proarrhythmic tendencies increase mortality. Radiofrequency ablation can cure only a limited number of arrhythmias. Implantable devices can be curative for bradyarrhythmias and lifesaving for tachyarrhythmias, but require a lifetime commitment to repeated procedures, are a significant expense, and may lead to severe complications. One possibility is the use of gene therapy as an antiarrhythmic strategy. As an initial attempt to explore this option, we focused on genetic modification of the atrioventricular node. First, we developed an intracoronary perfusion model for gene delivery, building on our previous work in isolated cardiac myocytes and hearts perfused ex vivo. Using this method, we infected porcine hearts with Adbetagal (recombinant adenovirus expressing Escherichia coli beta-galactosidase) or with AdGi (adenovirus encoding the Galphai2 subunit). We hypothesized that excess Galphai2 would mimic the effects of beta-adreneric antagonists, in effect creating a localized beta-blockade. Galphai2 overexpression suppressed baseline atrioventricular conduction and slowed the heart rate during atrial fibrillation without producing complete heart block. In contrast, expression of the reporter gene beta-galactosidase had no electrophysiological effects. Our results demonstrate the feasibility of using myocardial gene transfer strategies to treat common arrhythmias. PMID- 11100127 TI - Statins as a newly recognized type of immunomodulator. AB - Inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, or statins, are effective lipid-lowering agents, extensively used in medical practice. Statins have never been shown to be involved in the immune response, although a report has indicated a better outcome of cardiac transplantation in patients under Pravastatin therapy. Major histocompatibility complex class II (MHC-II) molecules are directly involved in the activation of T lymphocytes and in the control of the immune response. Whereas only a limited number of specialized cell types express MHC-II constitutively, numerous other cells become MHC-II positive upon induction by interferon gamma (IFN-gamma). This complex regulation is under the control of the transactivator CIITA (refs 6,7). Here we show that statins act as direct inhibitors of induction of MHC-II expression by IFN-gamma and thus as repressors of MHC-II-mediated T-cell activation. This effect of statins is due to inhibition of the inducible promoter IV of the transactivator CIITA and is observed in several cell types, including primary human endothelial cells (ECs) and monocyte-macrophages (Mstraight phi). It is of note that this inhibition is specific for inducible MHC-II expression and does not concern constitutive expression of CIITA and MHC-II. In repressing induction of MHC-II, and subsequent T-lymphocyte activation, statins therefore provide a new type of immunomodulation. This unexpected effect provides a scientific rationale for using statins as immunosuppressors, not only in organ transplantation but in numerous other pathologies as well. PMID- 11100128 TI - Transcutaneous immunization: a human vaccine delivery strategy using a patch. AB - Transcutaneous immunization, a topical vaccine application, combines the advantages of needle-free delivery while targeting the immunologically rich milieu of the skin. In animal studies, this simple technique induces robust systemic and mucosal antibodies against vaccine antigens. Here, we demonstrate safe application of a patch containing heat-labile enterotoxin (LT, derived from Escherichia coli) to humans, resulting in robust LT-antibody responses. These findings indicate that TCI is feasible for human immunization, and suggest that TCI may enhance efficacy as well as improve vaccine delivery. PMID- 11100130 TI - On the market PMID- 11100129 TI - Artificial antigen-presenting cells as a tool to exploit the immune 'synapse'. AB - Recent progress in molecular medicine has provided important tools to identify antigen-specific T cells. In most cases, the approach is based on oligomeric combinations of recombinant major histocompatibility complex-peptide complexes fixed to various rigid supports available for binding by the T-cell receptor. These tools have greatly increased our insight into mechanisms of immune responses mediated by CD8+ T cells. Examples of the diverse fields of application for this technology include immunization, viral infections and oral tolerance induction. PMID- 11100131 TI - Errata PMID- 11100133 TI - 2000 subject index - volume 6 PMID- 11100134 TI - Why do we sleep? PMID- 11100135 TI - A new link between pesticides and Parkinson's disease. PMID- 11100136 TI - Maximinis. PMID- 11100137 TI - Dissecting the ins and outs of excitement: glutamate receptors on the move. PMID- 11100138 TI - Closing in on a mammalian touch receptor. PMID- 11100140 TI - Keep your eye off the ball PMID- 11100139 TI - Sleep on it! PMID- 11100141 TI - Visual discrimination learning requires sleep after training. PMID- 11100142 TI - The two revolutions PMID- 11100143 TI - A guide to good behavior PMID- 11100144 TI - Light induces chromatin modification in cells of the mammalian circadian clock. AB - The mammalian circadian clock resides in neurons of the hypothalamic suprachiasmatic nucleus (SCN). Light entrains phase resetting of the clock using the retino-hypothalamic tract, via release of glutamate. Nighttime light exposure causes rapid, transient induction of clock and immediate-early genes implicated in phase-shifting the pacemaker. Here we show that a nighttime light pulse caused phosphorylation of Ser10 in histone H3's tail, in SCN clock cells. The effect of light was specific, and the kinetics of H3 phosphorylation were characteristic of the early response, paralleling c-fos and Per1 induction. Using fos-lacZ transgenic mice, we found that H3 phosphorylation and Fos induction occurRed in the same SCN neurons. Systemic treatment with the GABAB receptor agonist baclofen prevented light-induced c-fos and Per1 expression and H3 phosphorylation, indicating that one signaling pathway governs both events. Our results suggest that dynamic chromatin remodeling in the SCN occurs in response to a physiological stimulus in vivo. PMID- 11100145 TI - The molecular receptive range of an odorant receptor. AB - An odor perception is the brain's interpretation of the activation pattern of many peripheral sensory neurons that are differentially sensitive to a wide variety of odors. The sensitivity of these neurons is determined by which of the thousand or so odor receptor proteins they express on their surface. Understanding the odor code thus requires mapping the receptive range of odorant receptors. We have adopted a pharmacological approach that uses a large and diverse pool of odorous compounds to characterize the molecular receptive field of an odor receptor. We found a high specificity for certain molecular features, but high tolerance for others-a strategy that enables the olfactory apparatus to be both highly discriminating, and able to recognize several thousand odorous compounds. PMID- 11100146 TI - Presynaptic calcium stores underlie large-amplitude miniature IPSCs and spontaneous calcium transients. AB - The cellular mechanisms responsible for large miniature currents in some brain synapses remain undefined. In Purkinje cells, we found that large-amplitude miniature inhibitory postsynaptic currents (mIPSCs) were inhibited by ryanodine or by long-term removal of extracellular Ca2+. Two-photon Ca2+ imaging revealed random, ryanodine-sensitive intracellular Ca2+ transients, spatially constrained at putative presynaptic terminals. At high concentration, ryanodine decreased action-potential-evoked rises in intracellular Ca2+. Immuno-localization showed ryanodine receptors in these terminals. Our data suggest that large mIPSCs are multivesicular events regulated by Ca2+ release from ryanodine-sensitive presynaptic Ca2+ stores. PMID- 11100147 TI - Coincidence detection in single dendritic spines mediated by calcium release. AB - Cerebellar long-term depression (LTD) is a calcium-dependent process in which coincident activity of parallel fiber (PF) and climbing fiber (CF) synapses causes a long-lasting decrease in PF synaptic strength onto Purkinje cells. Here we show that pairing CF activation with bursts of PF activity triggers large (>10 microM) calcium signals in Purkinje cell dendrites. When PFs are densely activated, signals span whole dendritic branchlets and are mediated by voltage dependent calcium entry. When PFs are sparsely activated, however, signals are restricted to single spines and blocked by metabotropic glutamate receptor antagonists. Single-spine signals and sparse-stimulation LTD are also blocked by thapsigargin, indicating that calcium must be released from stores. Single-spine signals and sparse-stimulation LTD are greatest when PF activation precedes the CF activation within 50-200 ms. This timing rule matches the properties of several forms of motor learning, providing a link between behavior and functional properties of cerebellar synaptic plasticity. PMID- 11100148 TI - Benzodiazepines act on GABAA receptors via two distinct and separable mechanisms. AB - Benzodiazepines (BZs) act on gamma-aminobutyric acid type A (GABAA) receptors such as alpha1beta2gamma2 through key residues within the N-terminal region of alpha subunits, to render their sedative and anxiolytic actions. However, the molecular mechanisms underlying the BZs' other clinical actions are not known. Here we show that, with low concentrations of GABA, diazepam produces a biphasic potentiation for the alpha1beta2gamma2-receptor channel, with distinct components in the nanomolar and micromolar concentration ranges. Mutations at equivalent residues within the second transmembrane domains (TM2) of alpha, beta and gamma subunits, proven important for the action of other anesthetics, abolish the micromolar, but not the nanomolar component. Converse mutation of the corresponding TM2 residue and a TM3 residue within rho1 subunits confers diazepam sensitivity on homo-oligomeric rho1-receptor channels that are otherwise insensitive to BZs. Thus, specific and distinct residues contribute to a previously unresolved component (micromolar) of diazepam action, indicating that diazepam can modulate the GABAA-receptor channel through two separable mechanisms. PMID- 11100149 TI - Distinct molecular mechanisms and divergent endocytotic pathways of AMPA receptor internalization. AB - Internalization of postsynaptic AMPA receptors depresses excitatory transmission, but the underlying dynamics and mechanisms of this process are unclear. Using immunofluorescence and surface biotinylation, we characterized and quantified basal and regulated AMPA receptor endocytosis in cultured hippocampal neurons, in response to synaptic activity, AMPA and insulin. AMPA-induced AMPA receptor internalization is mediated in part by secondary activation of voltage-dependent calcium channels, and in part by ligand binding independent of receptor activation. Although both require dynamin, insulin- and AMPA-induced AMPA receptor internalization are differentially dependent on protein phosphatases and sequence determinants within the cytoplasmic tails of GluR1 and GluR2 subunits. AMPA receptors internalized in response to AMPA stimulation enter a recycling endosome system, whereas those internalized in response to insulin diverge into a distinct compartment. Thus, the molecular mechanisms and intracellular sorting of AMPA receptors are diverse, and depend on the internalizing stimulus. PMID- 11100150 TI - Regulation of AMPA receptor endocytosis by a signaling mechanism shared with LTD. AB - The endocytosis of AMPA receptors is thought to be important in the expression of long-term depression (LTD) triggered by NMDA receptor activation. Although signaling pathways necessary for LTD induction have been identified, those responsible for the regulated internalization of AMPA receptors are unknown. Here we show that activation of NMDA receptors alone can trigger AMPA receptor endocytosis through calcium influx and activation of the calcium-dependent protein phosphatase calcineurin. A distinct signaling mechanism mediates the AMPA receptor endocytosis stimulated by insulin. These results demonstrate that although multiple signaling pathways can induce AMPA receptor internalization, NMDA receptor activation enhances AMPA receptor endocytosis via a signaling mechanism required for the induction of LTD. PMID- 11100151 TI - Chronic systemic pesticide exposure reproduces features of Parkinson's disease. AB - The cause of Parkinson's disease (PD) is unknown, but epidemiological studies suggest an association with pesticides and other environmental toxins, and biochemical studies implicate a systemic defect in mitochondrial complex I. We report that chronic, systemic inhibition of complex I by the lipophilic pesticide, rotenone, causes highly selective nigrostriatal dopaminergic degeneration that is associated behaviorally with hypokinesia and rigidity. Nigral neurons in rotenone-treated rats accumulate fibrillar cytoplasmic inclusions that contain ubiquitin and alpha-synuclein. These results indicate that chronic exposure to a common pesticide can reproduce the anatomical, neurochemical, behavioral and neuropathological features of PD. PMID- 11100152 TI - Learning motivational significance of visual cues for reward schedules requires rhinal cortex. AB - The limbic system is necessary to associate stimuli with their motivational and emotional significance. The perirhinal cortex is directly connected to this system, and neurons in this region carry signals related to a monkey's progress through visually cued reward schedules. This task manipulates motivation by displaying different visual cues to indicate the amount of work remaining until reward delivery. We asked whether rhinal (that is, entorhinal and perirhinal) cortex is necessary to associate the visual cues with reward schedules. When faced with new visual cues in reward schedules, intact monkeys adjusted their motivation in the schedules, whereas monkeys with rhinal cortex removals failed to do so. Thus, the rhinal cortex is critical for forming associations between visual stimuli and their motivational significance. PMID- 11100153 TI - Neural events that underlie remembering something that never happened. AB - We induced people to experience a false-memory illusion by first asking them to visualize common objects when cued with the corresponding word; on some trials, a photograph of the object was presented 1800 ms after the cue word. We then tested their memory for the photographs. Posterior brain potentials in response to words at encoding were more positive if the corresponding object was later falsely remembered as a photograph. Similar brain potentials during the memory test were more positive for true than for false memories. These results implicate visual imagery in the generation of false memories and provide neural correlates of processing differences between true and false memories. PMID- 11100154 TI - A cortical area that responds specifically to optic flow, revealed by fMRI. AB - The continuously changing optic flow on the retina provides information about direction of heading and about the three-dimensional structure of the environment. Here we use functional magnetic resonance imaging (fMRI) to demonstrate that an area in human cortex responds selectively to components of optic flow, such as circular and radial motion. This area is within the region commonly referrred to as V5/MT complex, but is distinct from the part of this region that responds to translation. The functional properties of these two areas of the V5/MT complex are also different; the response to optic flow was obtained only with changing flow stimuli, whereas response to translation occurred during exposure to continuous motion. PMID- 11100155 TI - Long-lasting cortical plasticity in the object naming system. AB - A single exposure to an object can produce long-lasting behavioral change. Here, using event-related functional magnetic resonance imaging (fMRI), we provide evidence for long-lasting changes in cortical activity associated with perceiving and naming objects. In posterior regions, we observed an immediate (30-second) and long-lasting (3-day) decrease in neural activity after brief (200-ms) exposure to nameable and nonsense objects. In addition, slower-developing decreases in left inferior frontal activity were observed concurrently with increases in left insula activity, only for nameable objects. These time dependent cortical changes may reflect two distinct learning mechanisms: the formation of sparser, yet more object-form-specific, representations in posterior regions, and experience-induced reorganization of the brain circuitry underlying lexical retrieval in anterior regions. PMID- 11100156 TI - Early sleep triggers memory for early visual discrimination skills. AB - Improvement after practicing visual texture discrimination does not occur until several hours after practice has ended. We show that this improvement strongly depends on sleep. To specify the process responsible for sleep-related improvement, we compared the effects of 'early' and 'late' sleep, dominated respectively by slow-wave and rapid eye movement (REM) sleep. Discrimination skills significantly improved over early sleep, improved even more over a whole night's sleep, but did not improve after late sleep alone. These findings suggest that procedural memory formation is prompted by slow-wave sleep-related processes. Late REM sleep may promote memory formation at a second stage, only after periods of early sleep have occurred. PMID- 11100157 TI - From eye movements to actions: how batsmen hit the ball. AB - In cricket, a batsman watches a fast bowler's ball come toward him at a high and unpredictable speed, bouncing off ground of uncertain hardness. Although he views the trajectory for little more than half a second, he can accurately judge where and when the ball will reach him. Batsmen's eye movements monitor the moment when the ball is released, make a predictive saccade to the place where they expect it to hit the ground, wait for it to bounce, and follow its trajectory for 100-200 ms after the bounce. We show how information provided by these fixations may allow precise prediction of the ball's timing and placement. Comparing players with different skill levels, we found that a short latency for the first saccade distinguished good from poor batsmen, and that a cricket player's eye movement strategy contributes to his skill in the game. PMID- 11100158 TI - corrections and errata PMID- 11100160 TI - 2000 subject index - volume 3 nature neuroscience PMID- 11100161 TI - [Who am "I"? An essay on the dubious nature of the curriculum vitae]. PMID- 11100162 TI - ["Cranz" from "Das Gelehrte Osterreich" ("The Scholary Austrian")]. PMID- 11100163 TI - ["Crantz" from the "Biographie Luxembourgeoise" ("Luxembourg Biographies")]. PMID- 11100164 TI - ["Climate of knowledge, professional career and outside interests" using H.J.N. Crantz (1722-1797) as an example]. PMID- 11100165 TI - [H.J.N. Crantz. His rank, its origin and significance]. PMID- 11100166 TI - [Animal nutrition in the dietetics of the 18th Century with "Materiae Medicae" by H.J.N. Crantz as an example]. AB - H.J.N. Crantz (1722-1797), a physician and botanist born in the Duchy of Luxembourg and practising in Vienna under the Empress Maria Theresia during the 18th century, deals in his "materiae medicae et chirurgicae", written in Latin language, also with animal foodstuff. Animals from various species are commented on the utility of their meat and products to maintain health or to recover it after illness. The items of the text are described in free translation and in summary. Some more explanations are added. PMID- 11100167 TI - [Selenum carvifolia versus Peucedanum carfifolia. An historical look back on a botanical review and the first observation of the botanical work of H.J.N. Crantz]. AB - The case history of Peucedanum carvifolia Vill. and Selinum carvifolia L. (Apiaceae/Umbelliferae) is on one hand the occasion for a first attempt of a historical review of H.J.N. Crantz's (1722-1797) publications on the Umbelliferae, and on the other hand the opportunity for a discussion of the taxonomical and nomenclatural peculiarities of both species. PMID- 11100168 TI - [De duabus draconis arboribus botanicorum]. PMID- 11100169 TI - [Dragon's blood. A glance into the history of pharmacognosy]. AB - Dragon's blood, a red resin, which is often used as a dry powdered "herbal" remedy in traditional medicine, has different origins which are briefly discussed. This contribution however focuses on the species of the genus Dracaena Vand ex L., belonging to the Agavaceae and native to the Old World and the Canary Islands. H.J.N. Crantz, a botanist and medical doctor turned pharmacologist (born in 1722 near Luxembourg), had published a brief and very uncommon treatise on this group of plants, which he only knew from botanical gardens. The pharmacognostical and historical background of dragon's blood use is at the core of this contribution--the drug itself being of no major importance in today's medical care. PMID- 11100170 TI - [Science and medicine in 18th Century Luxembourg]. AB - During the 18th century science and medicine are poorly developed in Luxembourg (present part of the former duchy). Leprosy is vanishing and black death belongs to the past, whereas dysentery, typhoid fever and other epidemical diseases are thriving. In some specific diseases people claim help from protective saints such as St. Hubert (rabies) and St. Willibrord or the Blessed Virgin Mary. Besides these aspects the paper gives a concise outline of the situation in Luxembourg during the 18th century concerning hospitals, physicians, pharmacists and popular medicine. PMID- 11100171 TI - [The aging process and its effects on the digestive system]. PMID- 11100172 TI - ["The twilight of the gods" (of neurology)]. PMID- 11100173 TI - Complications and hospitalisation--duration after chemoembolisation for liver metastases. AB - In a retrospective study, all patients of the hemato-oncology department of the Centre Hospitalier who were treated from 1988 to 1997 by chemoembolisation for liver metastases were analysed for treatment-related hospitalisation duration, side effects and complications, in order to assess the treatment burden. Major side-effects were: pain in 17 of 29 patients, nausea in 8, vomiting in 7, persistent hickup in 3, fever in 12, a temporary confusional state in 4 patients. 1 patient experienced syncope, 2 patients developed homolateral pleral effusions, 1 patient suffered transient supraventricular arrhythmias. Major complications included 1 hemoperitoneum (under anticoagulant therapy), 1 hemorrhagic gastritis, 1 acute cholecystitis due to inflammatory tumoral choledochal obstruction and one iatrogenous acute pancreatic ischemic necrosis. Two patients died of post-embolic acute hepatic insufficiency, one 10 days, one 41 days after the last treatment session). In summary, chemo-embolisation of liver metastases is a complication burdened treatment in a strictly palliative setting with inestimable efficacy. The treatment modalities have to be discussed with the patient beforehand and preferably in controlled study setting. Large randomised trials may indicate patients' subgroups for benefit. PMID- 11100174 TI - [Cholesterol embolism: an often unknown disease; a report of two cases]. AB - We report on a case of post-invasive acute renal insufficiency and a case of acute necrotizing pancreatitis, both histologically proven to have been caused by cholesterol embolisms. A survey of the literature with discussion of diagnostic and therapeutic options is given. PMID- 11100175 TI - [Diagnosis of lung cancer]. AB - Due to the frequent diagnosis at a late inoperable stage and the bad prognosis of metastatic disease, lung cancer has become the first cause of cancer mortality. Early detection is thus the only way to influence mortality as there is no good treatment available for advanced disease. In the eighties, large screening studies using standard chest X Ray and sputum cytology have not been able to show a significant reduction in global lung cancer mortality. However these studies are now largely criticized for their methodological flaws. Recently, a new technique using auto-fluorescence fibroscopy has been developed, which is able to detect dysplastic and in situ neoplastic lesions which are invisible to standard fibroscopic techniques. This technique holds great promise in the detection of early lesions. In addition, molecular biology techniques are being developed which aim at detecting early invasive lesions at a stage where surgical treatment is still curative. The addition of these two techniques will probably in the future increase the efficiency of lung cancer screening. Therefore we think that new large scale screening studies are needed. PMID- 11100176 TI - [Recurrent abortion. Etiology and occurrence]. PMID- 11100177 TI - [External quality assurance in heart surgery at INCCI at the St Elizabeth Clinic using the modified Cleveland Scores]. PMID- 11100178 TI - [The story of the discovery of X rays]. PMID- 11100179 TI - [Medical liability insurance. Risk management from the view of the insurance]. PMID- 11100180 TI - [Personal risk management: private services for the aged]. PMID- 11100181 TI - [Professional incompetence: significance, definition, cost investigation]. PMID- 11100182 TI - [The place of arbitration agencies in practice]. PMID- 11100183 TI - [Risk management in gynecology and obstetrics. Forensic questions in connection with operative gynecology and obstetrics]. AB - Due to the avalanche-like increase of malpractice suits, it has become mandatory for doctors in hospitals and private practices to acquire a profound knowledge relating to modern management and to become familiar with the legal aspects of medical procedures. The article at hand reviews the standards of medical documentation and risk information within the scope of operative gynaecology and obstetrics and provides guidelines for a structured behaviour once an incident has occurred. The typical sources of complications are presented and strategies to avoid them are outlined: professional clinical management including the definition of competences for all therapists working together in a hospital. By observing these principles a professional medical level even in administrative respect is provided and thus enables an atmosphere of mutual confidence among doctors and patients. PMID- 11100184 TI - [Risk management in the USA]. AB - In the US and even within healthcare, Risk Management as a concept has existed for a long time--however, never as formally as it exists today. The change in how risk is identified and managed in hospitals and within medical care has evolved considerably in just the last 25 years. Risk management took on increased importance with the medical malpractice crisis the US experienced in the mid 1970s. Up until then, it appears its primary function had been to economically transfer risk. With the advent of managed care, risks have been recognized as generators of considerable potential cost to the healthcare system and one that must be proactively managed. Today, the successful management of risk is an essential part of healthcare administration. PMID- 11100185 TI - [Preventive aspects of risk management and quality assurance in medical training with examples in the European Union and the United States of America]. AB - The risk to be sued for clinical negligence by a patient is an integral part of modern medicine characterised by numerical evaluation of data, quality control of the results and cost containment. All three issues raised already at the beginning of the century by the American Surgeon Codman are without any doubt the basis of the spectacular achievements in the management of practically all diseases. At the same time they became a basis for compensation claims of patients who were not satisfied with the results of their treatment. The spiralling legal costs diverting resources of patients care and associated with feeling of doctors not being valued were interpreted as a profoundly negative aspect of modern medicine. Surprisingly, however, the complaints had also positive impact-revival of better documentation, involvement of the patient in the management of his disease and the realisation that a non-adversarial dispute resolution should replace the existing blame culture. PMID- 11100186 TI - [Liability risks in permanent resident physicians in physician-patient relations]. AB - Liability risks of medical doctors in their relationship to patients. Claims for compensations from patients, unallowed and non-approved treatment practices, offences against doctors, malfunctions of medical devices, risks of gynecologists, radiologists and orthopedic surgeons in private practice. PMID- 11100187 TI - [Quality practice]. AB - Risk-management, prospective and preventive quality-management and liability of doctors are important topics in daily practice and administration of justice. PMID- 11100188 TI - [The D10med patient clarification system]. AB - Doctors have to inform their patients about their disease, the procedures and risks of the treatments which will follow. This information is an essential topic in risk-management of our health care systems. DIOmed has designed over 500 checklists in different languages and for different medical treatments. These sheets facilitate the procedure and guarantee a high quality of information. PMID- 11100189 TI - ["Risk management in the practice field of ambulatory surgery in the intersection of home and hospital practice", with the practical example of the Ilmtal Clinic in Pfaffenhofen]. AB - Caring for patients in out-patients departments with ambulant surgery requires new thinking, new organizational structures and administration in hospitals. Ilmtal-Clinic in Pfaffenhofen has designed a model system to facilitate procedures with a high standard and risk management. PMID- 11100190 TI - [Risk management in an ambulatory surgery clinic]. AB - The special conditions in ambulant surgery in hospital require a preventing risk management to reach a security for patients which is comparable to stationary therapy. These requirements will be outlined concisely. At the example of organisation of ambulant surgery at the Ilmtalklinik Pfaffenhofen (Germany) a system of risk management is described. There defined running offs and developed checklists and their relation to risk management will be represented. In meaning of the authors risk-management is an essential part of general quality management. PMID- 11100191 TI - ["Health Forum". "Overstepping the limits of efficient cooperation in the field of health systems"]. PMID- 11100192 TI - [Address of the Minister of Health and Social Security Carlo Wagner at the 10th International Congress CALASS (23 September 1999)]. PMID- 11100193 TI - [Futurcare. The new definition]. PMID- 11100194 TI - [Home care. A concept and its place]. PMID- 11100195 TI - [Why should home care be theme?]. PMID- 11100196 TI - [Fresenius Kabi "From product to home care service provider"]. PMID- 11100197 TI - [Home care services in Luxembourg--innovative activity of the CPL Group]. PMID- 11100198 TI - [Activity and regeneration--modern orthopedic rehabilitation principles. Problem: epidemiology/demography]. PMID- 11100199 TI - [Rehabilitation in neighboring Germany]. PMID- 11100200 TI - [Significance of electronic security in health services in the German Republic]. PMID- 11100201 TI - [Date manipulation in the area of health services. The example of SAP R/3]. PMID- 11100202 TI - [Outsourcing. Advantages and disadvantages of these processes with regard to the management from the view of the technical services in a hospital]. AB - Cost effectiveness and improvement of service quality are the main reasons for outsourcing in hospitals. Long term relationships and customer satisfaction confirm the concept of providing complex services from external specialized companies. There is no standard guideline to determine whether outsourcing should be preferred to inhouse service. Outsourcing is not a guarantee for improved service and a lot of promoted advantages could theoretically also be provided by inhouse service departments. But the effectiveness of inhouse services has to be proven in comparison and competition to external providers. The often named disadvantages of outsourcing have to be considered. Nevertheless they are also not only subject to outsourcing and can also be a problem in inhouse organisations. Furthermore a proper controlling is an important tool to eliminate most of the difficulties in outsourcing projects. In the competition between internal and external service providers concepts of service partnerships that integrate internal and external resources will be the successful ones in the long run. The main potential for improvement is the optimization of the whole process of maintenance management, instead of the mere reduction of head count. PMID- 11100203 TI - [Integrated document management--key technology for successful undertakings and organizations in health services]. PMID- 11100204 TI - A new oral medicine journal? PMID- 11100205 TI - Electrocardiographic examination of dental patients with systemic lupus erythematosus. AB - Cardiac complications and abnormalities associated with systemic lupus erythematosus (SLE) have been well described. The most well-recognized cardiac myopathies in SLE are pericarditis and valvular problems such as verrucous endocarditis. Electrocardiogram (EKG) abnormalities and arrhythmias have also occurred and such individuals may have persistent tachycardia. To determine the prevalence of dental patients with SLE who have these EKG changes, a pilot study was designed in which a group of 13 subjects with SLE underwent EKG monitoring in the clinic. Most of these were patients referred to the oral medicine clinic. A randomly selected group of age-gender matched controls were monitored in an identical manner. All subjects first completed a cardiac screening questionnaire, were comfortably seated, and then monitored. The results indicated that the overall prevalence of EKG abnormalities among the subjects with SLE was 61.5% (n = 8). The most common arrhythmia was supraventricular extrasystoles. These were compared with a control group with a prevalence of 23% (n = 3). Of the 76.9% of the subjects with SLE who reported a positive history of cardiac abnormalities, 80% demonstrated EKG abnormalities as compared with 50% of the control group. Of the subjects with SLE who did not report a previous history, none demonstrated EKG abnormalities as was the same for the control group. These results indicate a high prevalence of EKG abnormalities in dental patients with SLE, especially those with a positive cardiac history. PMID- 11100206 TI - Multiple dental extractions using general anesthesia for a patient with Down and Eisenmenger syndromes and periodontal disease. AB - A patient with Down syndrome and severe retardation and Eisenmenger syndrome sought dental treatment at the Oregon Health Sciences University Hospital Dental Service. Eisenmenger syndrome is a form of cyanotic congenital cardiovascular pulmonary disease. The cardiac structural abnormalities associated with Eisenmenger syndrome preclude the use of conscious sedation and predispose a patient to the development of bacterial endocarditis. A prophylactic antibiotic regimen is recommended for patients with Eisenmenger syndrome who are undergoing procedures that produce a transient bacteremia. This article reviews the appropriate measures necessary for the safe anesthetic, operative, and perioperative care of a severely compromised patient. It is based on an understanding of the pathophysiology associated with Eisenmenger syndrome. PMID- 11100207 TI - Oral mucosal lichenoid reaction to sulfamethoxazole. AB - A clinical case of oral mucosal lichenoid reaction to sulfamethoxazole in a dental patient with complicating medical conditions is described. Although the relationship between oral mucosal lichenoid reaction and sulfa drugs has not been documented previously, the patient's lichenoid reaction corresponded with sulfamethoxazole use, and improved when the drug was discontinued. Reactions of this type should be monitored so that treatment and preventive measures may be instituted. PMID- 11100208 TI - Self-inflicted oral trauma: report of case. AB - Self-inflicted oral trauma is often encountered in patients who lack cerebral control over the masticatory cycle. Many factors must be considered in treatment, and a variety of appliances may be fabricated to prevent injury to oral structures. This case report documents the use of a soft acrylic appliance in treatment of a comatose patient who showed ruminatory reflex chewing. PMID- 11100209 TI - Nine years of Medicare dental benefits utilization. PMID- 11100210 TI - Physical restraints in dentistry had its origins in an era when there were no viable alternatives to forcing needed dental care on uncooperating patients. PMID- 11100211 TI - Not everyone loved Lassie. PMID- 11100212 TI - Clinical decision making in evaluating patients: a process study. AB - A volunteer patient was examined by five dentists experienced in caring for geriatric patients. The patient and dentist interaction was videotaped and reviewed to evaluate a previously generated hypothesis regarding the diagnosis process. This descriptive study suggests that dentists experienced in geriatric care did assess and evaluate a wide range of patient characteristics. However, they did not follow the previously hypothesized process in arriving at an appropriate treatment plan. Instead, they relied heavily on past experiences with similar situations. The possible implications of this process for treatment planning and teaching of geriatric dentistry are discussed. PMID- 11100213 TI - Profile and expectations of students entering general practice residency programs. AB - Although various studies have attempted to evaluate the success of general practice residency (GPR) programs in providing postdoctoral training, little work has investigated the motivation for senior dental students opting for such training compared with entering directly into private practice. Questionnaires were administered to 58 seniors at the State University of New York at Buffalo School of Dental Medicine 1 month before graduation to determine possible differences between those entering GPR programs and those planning to enter private practice. The survey also examined the goals of dentists about to enter GPR programs and compared them with the training goals determined by the Commission on Dental Accreditation. Although 75% of those entering GPR programs cited additional clinical experience as the reason for choosing this step in their careers, the most frequent response from those entering directly into private practice concerned the financial benefits. Also, of the respondents about to enter GPR programs, only one student indicated a desire for training in hospital dentistry. Although this survey was relatively limited in size, it appears that advanced instruction in the management of patients with complex health care problems is not one of the primary motivating factors for students considering residencies. This may indicate a need at the predoctoral level for greater instruction in changing health care demographics and in the training goals of general practice residencies, to continue to attract qualified dental graduates with goals compatible with those of GPR programs. PMID- 11100214 TI - Treatment of self-induced trauma in a patient with cerebral palsy. AB - Factitious illness can be a problem for disabled patients who experience recurrent trauma or irritation to a particular part of the body. In this case, the patient's recurring lip trauma combined with his overlying physical disability presented a problem not easily solved with prior methods of treating lip trauma. Modifying the standard orthodontic appliance solved the problem; after a short period of use, the patient dropped the harmful habit that caused the lip trauma. The appliance was no longer needed. PMID- 11100215 TI - Pouching of medications in the mouth: a case report. AB - Foods or medicines can be pouched or held in the mouth to prolong taste, enjoyment, or effect. This article describes a patient who pouches medications. The oral injury that occurred from this behavior is reported. PMID- 11100216 TI - Personality and dental care utilization: findings from the VA longitudinal study. AB - The personality traits of introversion/extraversion and neuroticism were investigated as determinants of the utilization of restorative dental care, controlling for socioeconomic status and restorative need. The VA Dental Longitudinal Study (DLS) provided information on the restorative treatment received by 593 healthy adult males during a 10-year period. Utilization of restorative services was measured by calculating the percentage of surfaces that needed and received treatment, as identified by the DLS examiners by clinical and radiographic examination. Oral examinations were initiated in 1969 with subsequent examinations occurring at approximately 3-year intervals. Personality measures for these individuals were obtained using the EPI-Q, a shortened form of the Eysenck Personality Inventory. A plot of neuroticism versus the utilization measure yielded a curvilinear relationship suggesting that participants seeking the most dental care scored moderately on the neuroticism scale while those scoring lowest and highest on this scale sought less treatment. In contrast, the introversion/extraversion scale showed no apparent relationship with dental care utilization. Results from the regression analysis suggest that dentition and socioeconomic status are strong determinants of utilization and that need for restorative treatment influences utilization. PMID- 11100217 TI - Effects of age and removable artificial dentition on taste. AB - Sodium chloride and sucrose gustatory recognition thresholds, suprathreshold taste intensity function, and sucrose suprathreshold taste preference in healthy adult males with removable artificial dentition were compared with persons having natural dentition. In addition, several inorganic salivary constituents were evaluated for their possible relationship with these modalities. The 75 participants studied were divided into two age and two dentition groups: less than 65 years versus 65 years and older; and natural dentition only versus either removable partial dentures or complete artificial dentition. Recognition threshold and suprathreshold psychophysical functions were assessed using aqueous solutions of sodium chloride and sucrose. Suprathreshold hedonic judgments were also determined for sucrose. Samples of stimulated parotid saliva were analyzed for sodium, potassium, and bicarbonate. As age increased, a statistically significant decrease was noted in the exponent of the psychophysical function for sodium chloride. A similar, but nonsignificant, trend was observed for sucrose, which was complicated by an interaction between age and dentition status. For sodium chloride, recognition thresholds tended to be higher for older persons with removable partial and complete dentures. However, no statistically significant effect was observed on the sucrose threshold or hedonic response as a result of dentition status. Analysis of several parotid saliva constituents showed no statistically significant effect caused by age or dentition status and correlations noted among the psychophysical measures appeared to be fortuitous. PMID- 11100218 TI - Predictors of oral health behaviors among elderly Japanese Americans. AB - As part of a series of studies on oral health behaviors of older persons, 81 elderly Japanese Americans were interviewed regarding their dental service utilization patterns, oral health attitudes, and dental status. The sample consisted of 35 Issei (first generation) and 46 Nisei (second generation) Japanese Americans. Because of the significant age difference between the Issei and Nisei, it was difficult to examine generational differences between the two groups. When age differences were controlled, there was no difference between the two generations in interval since their last dental visit. However, reasons for dental visits differed between the two groups, with the Issei more likely to seek help with dentures or general restorative work while the Nisei were more likely to schedule appointments for dental check-ups and cleaning. The best predictors of dental service utilization for the entire sample were the use of complete dentures, age, and education (R2 = .38), but when the respondent's perceived importance of oral health was entered into the equation, this variable accounted for more variance than age or education. The combination of complete dentures, importance, and age accounted for 41% of the variance in recency of dental service utilization. Locus of control, the belief component of attitudes, and overall attitude scores did not add significantly to the prediction of dental utilization. PMID- 11100219 TI - Use of intraligamentary anesthesia in a patient with severe hemophilia and factor VIII inhibitor. AB - This article reviews the therapeutic problems that arise in the dental management of hemophilia patients with inhibitor and suggests an anesthesia technique that may eliminate the need for factor replacement during restorative procedures. Intraligamentary anesthesia was used during restorative procedures that were performed throughout an 8-year period on a patient with factor VIII inhibitor. Multiple restorative procedures were accomplished without the use of replacement factor and with no postoperative bleeding. This technique, in consultation with the patient's physician, may be performed by most general practitioners in their private offices. PMID- 11100221 TI - The philosophy of dental care of the handicapped: past, present, and future. PMID- 11100220 TI - Appropriate responses by elders to oral symptoms: dentists' recommendations. AB - This paper reports the results of a study of dentists' recommendations regarding the appropriate health action of elders in response to certain oral common symptoms and problems. Previous studies have found that elders most commonly respond to oral symptoms by using nonprofessional means. This study found that, without exception, the recommendation to consult a dentist was the primary mode of response favored by dentists, even in response to symptoms and problems that are most often trivial and rarely indicative of serious health problems. The results of this pilot study suggest that the need exists for dentists to reach a more realistic consensus regarding alternatives to professional consultation in response to some symptoms and problems. PMID- 11100222 TI - Exaggerated use of statistics that refer to a 400% increase in training sites for dental geriatric programs. PMID- 11100223 TI - Detecting diabetes in an elderly dental patient population. AB - The prevalence of both diagnosed and undiagnosed diabetes, a disease which has a significant impact on dental care delivery, increases with age. Because of these two factors, the dental practitioner must pay special attention to the detection of diabetes in this age group. This paper describes a study that validates the use of a simple blood test in conjunction with a questionnaire as a method to detect undiagnosed diabetes in an elderly dental patient population. PMID- 11100224 TI - Dental conditions in patients with bipolar disorder on long-term lithium maintenance therapy. AB - Bipolar disorder (manic depressive disease) affects 1% of the United States population. These persons suffer from episodes of extreme elation followed by long periods of depression. Dental screening examinations of 40 patients consecutively admitted to the medical center with this diagnosis were performed. Poor oral hygiene, accumulations of supragingival and subgingival calculus, extensive dental caries, and numerous missing teeth were commonly identified. The majority of patients with bipolar disorder are treated with lithium carbonate. The physiological effects of lithium, and its interaction with drugs used in dentistry are reviewed, and disease-specific modifications in dental treatment are recommended. PMID- 11100225 TI - A comparison of prescription medication use by nursing home and homebound dental patients. AB - The elderly consume a disproportionate number of prescription medications dispensed in the United States. Although medication use among nursing home residents has been documented, little information is available regarding the medication use of homebound elderly. This study compared the medication use of nursing home and homebound patients treated by a mobile dental homecare program. A total of 338 dental records were reviewed, identifying the number and type of medications used. In addition, medications with xerostomic potential were identified. PMID- 11100227 TI - Drugs and the geriatric patient: a review of problems and special considerations faced by the dentist. PMID- 11100226 TI - A clinical evaluation of the restoration of root surface caries. AB - Root surface caries is of growing importance because its prevalence increases with age, and the population of the United States is growing older while edentulism and tooth loss rates have declined. Few clinical studies have evaluated materials used for the restoration of active root caries lesions. This study evaluated a Type II glass ionomer cement and a microfilled composite resin, both placed in preparations without mechanical retention or acid etching of enamel, in the restoration of root caries. Fifty adult volunteers with active root caries received one or both materials with the material chosen randomly. Patients were recalled after 24 months to evaluate restorations for retention, additional caries, marginal integrity, and overall clinical acceptability. Seventy-seven restorations were available for reevaluation. Forty-five percent of the glass ionomer and 73% of the composite restorations were clinically acceptable after 24 months. Of the glass ionomers, 39% were fully retained compared with 73% of composite restorations. Among those partially or fully retained, 25% of the glass ionomer restorations had minimal loss of marginal integrity and 30% had extensive loss while 53% and 9% of composite restorations had minimal and extensive loss, respectively. Most restorations were clinically unacceptable because of restorative material loss. Substantial numbers of glass ionomer cement and composite resin restorations were lost. This may be the result of difficulties in maintaining isolation and obtaining a proper gingival seal. Thus, routine use of mechanical retention is still highly recommended to reduce the loss of restorative material. PMID- 11100228 TI - Shared responsibility for oral health in long-term care facilities. PMID- 11100229 TI - Oral neglect in the institutionalized elderly. Part 1: The role of the institution. PMID- 11100230 TI - Referral of the handicapped, disabled, or infectious patient. PMID- 11100231 TI - A survey of the availability of dental services to developmentally disabled persons residing in the community. AB - Public Law 88-164, enacted in 1963, has led to extensive deinstitutionalization of persons with mental retardation from a peak census of 194,650 in 1967 to 91,440 by 1988. This population now depends on the community-based health care system for medical and dental care. A survey conducted to determine the availability of dental care to the developmentally disabled residing in group homes located in north central Florida indicated that 40% of caretakers experienced difficulty in locating dentists willing to provide comprehensive dental services for residents. According to the caretakers, although 75% of the residents were cooperative dental patients, dentists were reluctant to provide services for a variety of reasons, including financial disincentives, inadequate knowledge and preparation, and a lack of proper equipment necessary to treat this group of special patients. PMID- 11100232 TI - Aplastic anemia: current concepts and dental management. AB - Aplastic anemia (AA) is a rare blood dyscrasia in which the peripheral blood cells are decreased because of bone marrow failure. The clinical course reflects the severity of pancytopenia and is unpredictable for the individual. Hemorrhage and infection remain the major threats to these patients. Recent advances in transfusion medicine, infection management, bone marrow transplantation, and immunosuppressive therapy have improved survival of patients with AA. Oral manifestations of AA are common and may have serious sequelae. Two cases of acute periodontal infection associated with AA are presented. Dental management guidelines are presented in the context of interdisciplinary care. PMID- 11100233 TI - A scoring system for the quantitative evaluation of oral mucositis during bone marrow transplantation. AB - A system for assessing the severity of mucositis in patients undergoing bone marrow transplantation (BMT) is presented. Ten criteria were graded to give component scores together with a total score. The overall severity score ranged from 0 through 21. Scores were assigned three times daily by nursing staff members and verified daily by the attending dental and medical practitioners. Total scores were highly reproducible and were related to the severity of neutropenia. Variation between sequential total scores was not related to interexaminer variation but rather to changes in the severity of oral mucositis. Component scores provided a useful means for transmitting oral health information between health care personnel. Total scores were used to regulate the nature and frequency of oral hygiene procedures for patients undergoing BMT as well as other hematology/oncology patients. Application of this oral assessment system to other institutional settings may be beneficial. PMID- 11100234 TI - Oral herpes simplex virus infection with cardiac transplantation. AB - Oral herpes simplex virus infection is a common complication of cardiac transplantation. Lesions are secondary to reactivation of the virus, are atypical in appearance, and can involve any oral and perioral surface. Diagnosis on clinical grounds is difficult and should be confirmed with laboratory testing. A case report and review of the literature are presented to support the features of this infection. PMID- 11100235 TI - Oral neglect in the institutionalized elderly. Part 2: The role of the dentist and the standard of care. PMID- 11100236 TI - Legal issues in dentistry for the handicapped. PMID- 11100237 TI - Ethical issues in dental care for the compromised patient. PMID- 11100238 TI - Restraint and sedation of the dental patient with developmental disabilities. PMID- 11100239 TI - The 2000 Eleanor Clarke Slagle Lecture. Our mandate for the new millennium: evidence-based practice. AB - The health care environment of the past quarter century went through numerous evolutionary processes that affected how occupational therapy services were provided. The last iterations of these processes included requests for the evidence that supported what we were doing. This year's Eleanor Clarke Slagle Lecture (a) examines the strength of the evidence associated with occupational therapy interventions--what we do and how we do it--(b) raises dilemmas we face with our ethical principles when some of our practices are based on limited evidence, and (c) proposes a framework of continued competency to advance the evidence base of occupational therapy practice in the new millennium. PMID- 11100240 TI - A curricular renaissance: graduate education centered on occupation. AB - A 3-year project of curricular renaissance undertaken by the faculty of an entry level master's degree program is described. This project culminated in a thoroughly redesigned program of study centered around the construct of occupation and built on a foundation of knowledge in occupational science. Described herein are three developmental and highly iterative domains of activity that were crucial to the project's success: (a) environmental scanning and analysis, (b) creation of a compelling future vision of occupational therapy, and (c) curriculum planning. Also detailed are especially salient assumptions and beliefs about graduate education as well as seven themes that encompass the program's academic content and illustrate its defining emphases. These themes are (a) occupation, (b) the human as an occupational being, (c) occupation as a medium of change, (d) clinical reasoning, (e) ethical reasoning, (f) investigative reasoning, and (g) occupational therapists as scholars and change agents in systems. The article concludes with reflections on innovation in graduate education in occupational therapy today. PMID- 11100241 TI - Pressure garment adherence in adult patients with burn injuries: an analysis of patient and clinician perceptions. AB - OBJECTIVE: This study provides a descriptive analysis of the factors affecting pressure garment adherence from the perspective of adult patients with burn injuries and occupational therapy clinicians. METHOD: Questionnaires were administered to 23 adult patients with burn injuries and 10 occupational therapy clinicians selected from six provincial hospitals in a large metropolitan area in South Africa. Adherence behavior was evaluated from the perspective of both sample groups in terms of four variables: garment type, garment comfort, garment cosmesis, and garment instructions. RESULTS: Adherence behavior was negatively influenced by differences in both patient and clinician perceptions across several variables, including the types and consequences of skin problems arising from pressure garment use, levels of satisfaction with garment construction and color, and the issuing and understanding of garment instructions. Other factors compromising adherence behavior included the negative effects of visible burn disfigurement, the issuing of pressure garments after hypertrophic scarring had developed, a lack of patient choice in the selection of scar management techniques, and a lack of social support in the wearing of pressure garments. CONCLUSION: Much of what is traditionally understood as "patient nonadherence" appeared to be largely because of rational choices made by patients in the face of several difficulties they experienced with the current form and nature of their pressure garment therapy. On the basis of these findings, a range of patient-centered interventions are indicated to enhance treatment efficacy and consumer satisfaction with this treatment regimen, including horizontal rather than vertical therapist-patient communication, closer interaction among members of the health care team, the facilitation of family and social support, and interactive health education interventions. PMID- 11100242 TI - ADL performance of black Americans and white Americans on the assessment of motor and process skills. AB - OBJECTIVE: The purpose of this study was to examine whether the Assessment of Motor and Process Skills (AMPS), an assessment of personal and domestic activities of daily living (ADL) performance, can be used as a valid, nonbiased tool when assessing black Americans. METHOD: The participants were 466 blacks and 466 whites drawn from the entire sample of blacks and whites contained in the AMPS database who met the following criteria: (a) were 16 years of age and older; (b) had a notable history of a neurological, musculoskeletal, medical, developmental, cognitive, or psychiatric disorders or were healthy older persons; and (c) resided in North America. The participants were matched according to functional level, gender, diagnosis, and age. Examination for bias included between-group comparison of (a) item difficulty and task challenge hierarchies of the AMPS, (b) goodness-of-fit of the participants to the many-faceted Rasch (MFR) model, and (c) mean ADL motor and ADL process abilities. RESULTS: Both the item difficulty and the task challenge hierarchies remained stable between the two groups. On the ADL Motor scale, 95.3% of the black participants and 92.4% of the white participants demonstrated acceptable goodness-of-fit (MS < or = 1.4, z < 2) to the MFR model. On the ADL Process scale, 91.2% of the black participants and 90.1% of the white participants demonstrated acceptable goodness-of-fit. A significant difference, t(2, 930) = 3.56, p < .01, between the two groups was found in mean ADL process ability, but no significant difference, t(2, 930) = .69, p = .49) was found in mean ADL motor ability. CONCLUSION: The results of this study support the validity of the AMPS when applied to black Americans. PMID- 11100243 TI - Occupational Therapy Code of Ethics (2000). PMID- 11100244 TI - Enforcement procedure for Occupational Therapy Code of Ethics (2000). PMID- 11100245 TI - Occupational therapy and the Americans with Disabilities Act (ADA). AB - Occupational therapy practitioners play a key role in educating the public, as well as persons with disabilities, about their rights and responsibilities under the ADA. Occupational therapy practitioners' understanding of work and task analysis, knowledge of the functional limitations of disability, and experience with reasonable accommodations, adaptive equipment, and environmental adaptations place them in a unique position to serve as a resource in ADA-related matters [EEOC, Title I Technical Assistance Manual sections 3.6, 3.10(6), 4.5(4), 6.2, 6.4, 6.6 (1992)]. The AOTA supports the fundamental purposes of the ADA and encourages its members to assist the public in complying with ADA mandates to promote the inclusion of persons with disabilities into the mainstream of society and the pursuit of meaningful occupations. PMID- 11100246 TI - Occupational therapy services in facilitating work performance (statement). PMID- 11100247 TI - Specialized knowledge and skills in eating and feeding for occupational therapy practice. PMID- 11100248 TI - Specialized knowledge and skills for occupational therapy practice in the neonatal intensive care unit. PMID- 11100249 TI - [Sevoflurane, desflurane--an no end in sight!]. PMID- 11100250 TI - [Xenon--a new anesthetic]. AB - Today, xenon anaesthesia becomes an interesting object due to xenon's positive ecological aspects as well as its nearly ideal properties as an anaesthetic gas. The application of xenon provides advantages regarding intraoperative hemodynamic stability and the recovery period. Xenon increases tissue perfusion in most organ systems. This increase of tissue perfusion during an operation can be important during some surgical procedures and may improve postoperative outcome. In anaesthetic concentrations xenon leads to an increase in cerebral perfusion. For that reason, xenon should be avoided in patients with increased cerebral pressure or cerebral perfusion disorders. Therefore, further clinical investigations are necessary to improve these clinical findings. The development of accepted indications of xenon anaesthesia becomes necessary regarding a limited availability and high costs of the gas. Focussing this target, more clinical and experimental investigations will become necessary in the next years. PMID- 11100251 TI - [Influences and predictors of unanticipated admission after ambulatory surgery]. AB - In order to plan the daily routine of a surgical day care unit optimally and effectively, it is indispensable to know the causes of unanticipated admission of outpatients. The purpose of this experiment was to evaluate the influences and predictors of unanticipated admission of patients in our day care unit for ambulatory surgery. The data sets of 3152 surgical outpatients were evaluated. The duration of stay had been entered online by computers. METHOD: From January 1997 until June 1999, all clinically relevant parameters from any outpatient were entered into an anesthesia information management system (NarkoData, Imeso GmbH, Huttenberg-Rechtenbach, Germany). The correlation of potential nominal and ordinal scaled predictors of unanticipated admission was tested using the chi squared test. Univariate analysis was used in determining predictors for the occurrence of unanticipated admission. Pearson's contingency coefficient (CC) was used as a standard for the correlation rigidity in nominal and ordinal scaled parameters. The correlation standard eta was used for metrical parameters. RESULTS: Unanticipated admission occurred in 169 (5.4%) of the 3152 outpatients. The following parameters significantly influenced unanticipated admission: age, ASA status, diagnosis (ICD-9), time of admission, different anesthesia procedures and anesthetics (opioids and non-depolarizing muscle relaxants), surgical department, type of surgery (ICPM), duration of operation, blood loss, intraoperative hemoglobin values, and the administration of colloid and crystalloid solutions. The parameters blood loss, intraoperative hemoglobin values, and administration of colloid solutions were evaluated as being good predictors. CONCLUSION: The causes of unanticipated admission of patients in our day care unit for ambulatory surgery are manifold. Some relate to the patient, the anesthesia, and the organization of the day care unit, whereas lengthy operative trauma leading to intraoperative blood loss also plays a major role. PMID- 11100252 TI - [Emergency medicine online course--integrating into curriculum of computer-based training]. AB - Emergency medicine is characterized by rapid decision making to help patients in life-threatening situations. Teaching these skills requires a high level of interaction between medical students and the lecturer. We designed, implemented, and evaluated a generic computer-based training (CBT) system to provide a more active way of learning emergency medicine. The content of the training program is adapted to the knowledge of third year medical students and is focused on basic skills and real-world problems. The teacher presents the case with authentic video sequences and slides. The cases are classified into four groups: heart (e.g., myocardial infarction), respiration (e.g., asthma bronchiale), trauma (e.g., car accident), and loss of consciousness (e.g., coma). Within a realistic time frame, the students have to answer free text and multiple choice questions on a work-station. All answers given by the students are processed anonymously by the CBT system via a central server and displayed on a large video screen, thus enabling a detailed discussion without intimidation of individual students. This interactive technique allows for immediate feedback from the lecturer based on the specific knowledge of his group and his own experience. The IT concept, which is scalable to many subjects, is based on state of the art internet technology and therefore suitable for teleteaching. A major design objective for the program was a self-explaining and robust user interface. The system has been in routine use since 1998. We designed an evaluation form consisting of 21 items focused on subjective rating of learning success, acceptance of CBT, and technical feasibility. We analyzed forms from 138 students and found high scores for acceptance and learning success (median 5 on a 6-point scale). user problems with the program were denied (median 1 on a 6-point scale). Computer-based training with Internet technology can provide a successful method for interactive teaching of emergency medicine and is well accepted by students. PMID- 11100253 TI - [Diagnostic value of s-100 protein and neuron-specific enolase as serum markers for cerebral deficiency after general anesthesia. Study in patient with hip or knee replacement]. AB - S-100 protein and neuron-specific enolase (NSE) serum concentrations serve as markers of cerebral damage in cardiac surgery, neurology, or after head injury. In these circumstances, S-100 and NSE levels correspond with the results of neuropsychological tests. The present study investigated the diagnostic value in orthopaedic patients after joint replacement. METHODS: Forty patients scheduled for elective hip or knee arthroplasty were investigated. Serum values of NSE and S-100 were determined preoperatively and 30 min and 4, 18, and 36 h postoperatively. Neuropsychological tests (syndrome short test, SKT, delirium assessment according to DSM IV) were performed preoperatively and two, three, and four days following surgery. General anaesthesia was induced with fentanyl and etomidate and maintained with isoflurane in oxygen/air. FINDINGS: The S-100 increased from a median of 0.04 ng/ml (range 0.004-0.19 ng/ml) preoperatively to 1.03 ng/ml (range 0.18-3.65 ng/ml) at 30 minutes postoperatively (P < 0.0001). These levels returned to normal in the course of the following 2 days. NSE values were 8.55 ng/ml (range 4.6-14.9 ng/ml) preoperatively and 7.07 ng/ml (range 4 16.4 ng/ml) postoperatively (P = 0.167). There were no differences in serum concentrations of S-100 and NSE between normal patients and those with postoperative cognitive deficit. Furthermore, no correlation was found between the serum marker and neuropsychological tests. INTERPRETATION: Obviously, increased NSE levels seem to indicate cerebral damage only in more severe cases. S-100 does not seem to be brain-specific in patients undergoing orthopaedic surgery. Therefore, the value of S-100 in the assessment of brain disorders is limited. PMID- 11100254 TI - [Brain death and intensive care medicine. The cultural history of a medical concept]. AB - The paper focuses on the cultural history of brain death in Germany in the second half of the 20th century. It analyzes scientific and public discourses on the relevance of brain death and the importance of medical innovations in intensive care medicine. The paper examines how the public reacted when heart transplantation led to an urgent need for a new definition of death. It will be shown how the concept of brain death was accepted by the public, introduced into clinical routine, and implemented through medical and legal policies. Finally, it will be analyzed why the public consensus on brain death was definitely questioned in the last ten years. An understanding of the use of the concept of brain death by scientists, lawyers, theologians, and the public during the last three decades may help to shed light on the social role of science in modern and late-modern societies. PMID- 11100255 TI - [Pneumothorax in vertical infraclavicular block of the brachial plexus. Review of a rare complication]. AB - A 50 year old female patient received anaesthesia of the arm by the vertical infraclavicular blockade of the plexus brachialis (VIP). Postoperatively an ipsilateral pneumothorax occurred complicated by pleural effusion and a contralateral bronchopneumonia, which resolved completely after treatment. The blockade of the plexus was performed correctly, failures in determining the correct point of needle insertion could be excluded. Therefore a pneumothorax has to be regarded as a specific complication of the VIP, which might occur despite correct technique, and requires that the patient be informed of this eventuality. Nevertheless, the VIP is an important method due to its high success rate concerning blockade of the musculocutaneous nerve and tolerance of tourniquet. The risk of a pneumothorax is about 0.2 to 0.7%. PMID- 11100256 TI - [The effect of local anesthetics on blood coagulation]. PMID- 11100257 TI - [Anesthesia in cesarean section and previously existing inoperable intracerebral angioma]. PMID- 11100258 TI - [Excitation after sevoflurane: an problem in pediatric anesthesia? Comment on an article in Der Anaesthesist (1999) 48:917-918]. PMID- 11100259 TI - [Is the incidence of postoperative vomiting dependent on the operative procedures? Remarks on a letter in Der Anaesthesist (2000) 49:227-229]. PMID- 11100260 TI - [Acute craniocerebral injury. Pathophysiology, monitoring and treatment]. PMID- 11100261 TI - Exotic becomes erotic: interpreting the biological correlates of sexual orientation. AB - Although biological findings currently dominate the research literature on the determinants of sexual orientation, biological theorizing has not yet spelled out a developmental path by which any of the various biological correlates so far identified might lead to a particular sexual orientation. The Exotic-Becomes Erotic (EBE) theory of sexual orientation (Bem, 1996) attempts to do just that, by suggesting how biological variables might interact with experiential and sociocultural factors to influence an individual's sexual orientation. Evidence for the theory is reviewed, and a path analysis of data from a large sample of twins is presented which yields preliminary support for the theory's claim that correlations between genetic variables and sexual orientation are mediated by childhood gender nonconformity. PMID- 11100262 TI - The subtlety of sex-atypicality. AB - Memories of sex-atypical behavior and interests in childhood usually differ between homosexual and heterosexual people. However, variation within these broad groups has not previously been explored in detail, especially among women. We utilized data from a postal survey of a nationwide sample of Australian adult twins (n = 4,901, age range: 19-52 years). Among men, 15.2% reported homosexual behavior (ever), 11.5% said they had been sexually attracted to the same sex, and 6.4% said they were not heterosexual; the corresponding figures for women were 7.9, 10.6, and 3.5%. A continuous measure of childhood gender nonconformity (CGN) was sensitive to slight variations in homosexual attraction and behavior. In particular, among both men and women who identified as heterosexual, there were significant differences between "complete" heterosexuals and those who admitted to only one or a few same-sex behaviors but no homosexual attraction. Among men, CGN scores distinguished between heterosexuals who admitted to same-sex behavior only and those who admitted to some homosexual attraction. The sexual subgroups also differed on a measure of gender atypicality in adulthood. Implications for developmental theories of sexuality are discussed. PMID- 11100263 TI - Psychoanalysis and sexual fantasies. AB - Psychoanalysis began as a depth psychology, heavily based on the sexual experiences and memories of patients. A long-term treatment, utilizing a free association method, psychoanalysis has provided a window onto the meanings and functions of fantasy, including sexual fantasy. Although psychoanalysis has produced some scientific research, the field has tended to rely on observational data collected from individuals studied in depth. Sex research on the other hand, carried out by investigators from different disciplines is based on empirical investigation. Each field has made contributions fundamentally important to the other. In this article, we review psychoanalytic ideas about human sexuality and distinguish those that have been invalidated by systematic research from those that remain useful. Perhaps, the single most important revision of psychoanalytic theory during the past century was concerned with the psychological development of girls and women. We separately discuss the development of the sexes, and stress the need for bridge building between psychoanalysis and sex research. PMID- 11100264 TI - The magical age of 10. AB - Developmental processes of "puberty" and their cultural contexts in understanding the emergence of sexual subjectivity, especially sexual attraction, prior to gonadarche are critically examined. In particular, we consider the hypothesis that "sexual attraction" follows the onset of adrenal puberty, termed adrenarche, precipitating the development of stable and memorable attraction toward others approximately by the age of 10. In a prior study, the authors suggested that adrenarche is a significant source of this developmental change in sexuality (McClintock, M., and Herdt, G., 1996). The inferential evidence from New Guinea is compared with recent studies from the United States, including clinical findings on "precocious puberty." We conclude with the question of whether the age of 10 is a human universal in the development of attraction and sexuality. PMID- 11100265 TI - Sexual identity trajectories among sexual-minority youths: gender comparisons. AB - The present investigation explored gender differences in sexual identity development--first same-sex attractions, self-labeling, same-sex sexual contact, and disclosure--among 164 sexual-minority young adults. Based on interviews, results indicated the value of assessing gender differences in the context, timing, spacing, and sequencing of sexual identity milestones. Adolescent males had an earlier onset of all milestones except disclosure. The context for sexual identity milestones were likely to be emotionally oriented for young women and sexually oriented for young men. The gap from first same-sex attractions (8-9 years of age) to first disclosure (around 18 years) averaged 10 years for both sexes. Young women followed label-first developmental trajectories; men were more likely to pursue sex before identifying themselves as gay. In terms of achieving sexual identity milestones, gender mattered, but it was not everything. PMID- 11100266 TI - Identification and characterization of Malaysian river catfish, Mystus nemurus (C&V): RAPD and AFLP analysis. AB - This work represents the first application of the amplified fragment length polymorphism (AFLP) technique and the random amplified polymorphic DNA (RAPD) technique in the study of genetic variation within and among five geographical populations of M. nemurus. Four AFLP primer combinations and nine RAPD primers detected a total of 158 and 42 polymorphic markers, respectively. The results of AFLP and RAPD analysis provide similar conclusions as far as the population clustering analysis is concerned. The Sarawak population, which is located on Borneo Island, clustered by itself and was thus isolated from the rest of the populations located in Peninsular Malaysia. Both marker systems revealed high genetic variability within the Universiti Putra Malaysia (UPM) and Sarawak populations. Three subgroups each from the Kedah, Perak, and Sarawak populations were detected by AFLP but not by RAPD. Unique AFLP fingerprints were also observed in some unusual genotypes sampled in Sarawak. This indicates that AFLP may be a more efficient marker system than RAPD for identifying genotypes within populations. PMID- 11100267 TI - Biochemical polymorphism in yellow catfish, Mystus nemurus (C&V), from Thailand. AB - Yellow catfish, Mystus nemurus (Cuv. & Val.), is becoming one of the major freshwater species farmed by aquaculturists in Southeast Asia. It was of interest to examine levels of genetic subpopulation differentiation among samples of this species obtained from parts of its range, as well as to compare the genetics of wild and hatchery-bred fish. Horizontal starch gel electrophoresis and histochemical staining techniques were used to examine genetic variation within and among eight wild and one hatchery populations of M. nemurus from northern, northeastern, central and southern Thailand. Four tissues (heart, liver, kidney, and muscle) from individual specimens were used to analyze variations at 23 protein-coding loci. Fifteen of the 23 loci examined (65.22%), namely, ACP*, AAT 1*, EST-1*, EST-2*, GPI*, IDH-1*, IDH-2*, MDH-1*, MDH-2*, MDH-3*, ME*, PGM*, 6PGD*, SOD*, and HB*, were polymorphic at the 0.95 level. Observed heterozygosities ranged from 0.041 to 0.111, with an average of 0.068 +/- 0.028. Genetic distances ranged from 0.005 to 0.164. The greatest genetic distance was found between the Chainat and the Suratthani populations (0.164), a level indicative of subspecific differentiation in M. nemurus from within Thailand. PMID- 11100268 TI - New genetic variation in European hares, Lepus granatensis and L. europaeus. AB - Six genetic polymorphisms for the Iberian hare (Lepus granatensis) and four for the brown hare (L. europaeus) are newly described. The genetic variation of peptidases B (PEPB) and C (PEPC), hemoglobin alpha chain (HBA), hemopexin (HPX), vitamin D binding protein (GC), and properdin factor B (BF) was assessed by conventional electrophoresis and isoelectric focusing in carrier ampholytes and hybrid pH gradients. Six alleles were detected in PEPB, three in PEPC, four in HBA, six in GC, five in HPX, and six in BF. At least one allele was shared between species at all loci except HBA. The allelic overlap between the two species was medium to high in PEPB, GC, and HPX and small in PEPC and BF. PMID- 11100269 TI - Heterologous amplification of microsatellite loci from mouse and rat in oryzomyine and Thomasomyine South American rodents. AB - Eleven heterologous primers were investigated in 119 individuals of 11 species of rodents of the Oryzomyine and Thomasomyine groups. The animals were collected at four sites of the "Cerrado," a dryland biome located on the Brazilian Plateau, all of them being karyotyped and taxonomically allocated according to the karyotype. Four of these primers, R47, R65, R75 (from Rattus), and ATP (from Mus) cross-amplified in at least one of these taxa, giving products of seven, nine, one, and three bands, respectively. These values are of the same order as others obtained when heterologous primers were amplified in other orders of mammals. Of the 20 products amplified in these two rodent groups by these four primers, only 7 of the bands were seen in a heteromorphic state (one individual presenting two bands), in two species (Rhipidomys aff. leucodactylus and Oryzomys megacephalus). The others occurred as monomorphic bands. PMID- 11100270 TI - Molecular cloning and tissue expression analysis of the beta subunit of elongation factor 1 in the mouse. PMID- 11100271 TI - Genetic factors influencing the development of chronic graft-versus-host disease in a murine model. AB - Graft-versus-host disease (GVHD) is a major complication of bone marrow transplantation that can occur in either acute or chronic forms. Much of the long term pathology seen in chronic GVHD is a result of autoantibody production. In the DBA/2-->B6D2F1 murine model of chronic GVHD, anti-ssDNA autoantibodies can be detected by 14 days post cell transfer. These autoantibodies are not observed in B6D2F1 recipients of cells from C57BL/6 or B10.D2 donors, which develop acute rather than chronic GVHD. Therefore, in this model, donor genetic factors predispose to the development of chronic GVHD in recipients. We performed a genetic analysis aimed at mapping donor loci that influence the magnitude of early autoantibody production in B6D2F1 recipients of cells from DBA/2 donor mice. Linkage analysis suggested an influence of two loci: a locus on chromosome 11 linked to D11Mit278 and a locus on chromosome 4 linked to D4Mit226. The locus on chromosome 11 also appeared to influence the development of renal pathology associated with chronic GVHD. PMID- 11100272 TI - Pharmacokinetics and adverse events following 5-day repeated administration of lenograstim, a recombinant human granulocyte colony-stimulating factor, in healthy subjects. AB - Recombinant human granulocyte colony-stimulating factor (rhG-CSF) (lenograstim) was administered to healthy subjects at doses of 2, 5 and 10 micrograms/kg/day for 5 days (twice a day subcutaneously) to examine the optimal dose and schedule of lenograstim in mobilizing peripheral blood progenitor cells (PBSC) for allogeneic transplantation. Lenograstim administration significantly increased CD34+ cells in a dose-related manner. A significant correlation was observed between the maximal post-dosing counts and the pre-dosing baseline counts of CD34+ cells. Peripheral neutrophils increased markedly by seven to 13 times from the baseline to a peak of approximately 40,000/microliter on day 5 for the 5 and 10 micrograms/kg/day doses. After peak serum concentration (Cmax) was attained 4 h following administration, serum G-CSF declined with time in a log-linear fashion. The Cmax and 12 h area-under-the-curve increased dose dependently, but minimum drug level increased up to day 2 and then decreased until day 5. Clearance decreased with increasing dosage at the first dose, and increased significantly at the last dose. We found a highly significant correlation between absolute neutrophil counts and clearance for each dose. Adverse events most frequently occurred on day 6, with increases of alkaline phosphatase and lactate dehydrogenase and onset of bone pain. Increases of aspartate aminotransferase and alanine aminotransferase occurred as delayed events. Platelet count gradually decreased after the end of drug administration to 57% of the pre-dosing count on day 10, but was still within the normal range. These preliminary results suggest that repeated doses of lenograstim induce mobilization of PBSC in a dose dependent manner and the pre-dosing baseline count of PBSC may predict the post dosing maximal mobilization. The drug treatment may cause delayed-onset moderate thrombocytopenia and increased transaminase, and the drug clearance changes in a complex manner during repeated dosing. PMID- 11100273 TI - Outcomes of high-dose chemotherapy and autologous stem cell transplant in isolated locally recurrent breast cancer: a multicenter evaluation. AB - To determine the outcomes of women with isolated loco-regional recurrence (LRR) of breast cancer treated with high-dose chemotherapy (HDCT) and autologous stem cell transplantation (ASCT) following conventional therapy, we conducted a retrospective review of 58 patients from five institutions treated between 1990 and 1998. Forty-five patients (78%) had > or = 2 poor prognostic factors (PPF) (defined as disease-free interval preceding LRR < or = 2 years, hormone receptor negative/refractory disease, and incomplete resection). At median follow-up of 14.2 (0.5-72) months, 36 patients (62%) developed progressive disease. Disease progression usually occurred at local (27 patients) vs distant (nine patients) sites. Median time to disease progression following ASCT was 6.1 (1.3-31.4) months. At last follow-up, 23 patients (40%) had expired (all due to disease progression), and 13 (22%) were alive with, and 22 (38%) without progressive disease. By Kaplan-Meier analysis, the estimated median PFS and OS was 20.3 and 29.2 months, respectively. In a multivariate model, complete remission at time of HDCT and estrogen-receptor positive disease were predictive of significantly longer PFS and OS. The survival of this cohort was similar to previous reports of those treated with conventional therapy alone, and to those with distant metastases treated with HDCT. Frequent progression locally, suggests that strategies to improve local disease control are needed. PMID- 11100274 TI - Repetitive high-dose therapy with cyclophosphamide, thiotepa and docetaxel with peripheral blood progenitor cell and filgrastim support for metastatic and locally advanced breast cancer: results of a phase I study. AB - This phase I study was designed to determine the optimal dosages of a novel repetitive high-dose therapy regimen for patients with metastatic breast cancer (MBC). The planned treatment was three cycles of high-dose cyclophosphamide, thiotepa and docetaxel delivered every 35 days with progressive dose-escalation in successive cohorts. Each cycle was supported by peripheral blood progenitor cells (PBPC) and filgrastim. Eighteen patients were entered into this trial. Of the planned 54 treatment cycles, 44 were delivered and 11 patients completed all three cycles. The dose-limiting toxicities were interstitial pneumonitis and mucositis with moderately severe diarrhea (n = 3) and rash (n = 3). There were no treatment-related deaths. Of the 17 patients with evaluable disease, 16 patients responded with six patients achieving a complete remission and an additional four patients achieving no detectable disease (negative restaging including PET scan) but a persistently abnormal bone scan. At a median follow-up of 12 months, median progression-free survival was 11 months with the median overall survival not reached. The recommended doses for phase II/III studies are cyclophosphamide (4 g/m2), thiotepa (300 mg/m2) and docetaxel (100 mg/m2). PMID- 11100275 TI - Blood stem cell transplantation as therapy for primary systemic amyloidosis (AL). AB - This study investigated the response rate and toxicity of blood cell transplantation as treatment for primary amyloidosis (AL). Twenty-three patients had stem cells collected between November 1995 and September 1998. Conditioning included melphalan and total body irradiation in 16 and melphalan alone in 4. Three patients did not undergo stem cell infusion because of poor performance status. Two died of progressive amyloid at 1 and 3 months. One patient is alive on hemodialysis. Fourteen males and six females (median age, 57 years) underwent transplantation. Renal, cardiac (by echocardiography), peripheral neuropathy or liver amyloidosis occurred in 14, 12, 3, and 1, respectively. Echocardiography demonstrated an interventricular septal thickness > or = 15 mm in six patients, five of whom died post transplantation. Three patients died of progressive amyloidosis at 7, 7, and 21 months. Thirteen patients are alive with a follow-up of 3 to 26 months. Twelve (60%) fulfilled the criteria of a hematologic or organ response. Severe gastrointestinal tract toxicity was seen in five (25%). We conclude that blood cell transplantation for amyloidosis had a much higher morbidity and mortality compared with transplantation for myeloma. The best results appear to occur in patients with nephrotic syndrome as the only manifestation of their disease. PMID- 11100277 TI - Effect of complete response on outcome following autologous stem cell transplantation for myeloma. AB - We studied the effect of complete response (CR) among 126 consecutive patients who underwent stem cell transplantation (SCT) for myeloma. The CR rate with SCT was 33%. Median overall survival (OS) from diagnosis of myeloma was 56 months. OS following SCT was 22 months. Progression-free survival (PFS) was 12 months. OS was not different between patients who achieved CR and those who did not, median survival 25 vs 24 months, P = 0.5. Corresponding median times for PFS were 15 and 11 months, P = 0.2. The plasma cell labeling index (PCLI) was high (> or = 1%) in 36% (high risk group) and was associated with poor OS and PFS (P < 0.001). Achieving CR did not influence OS or PFS in either the high or the low risk group. In contrast, OS and PFS were significantly influenced by high PCLI both in patients who achieved CR and those who did not. OS was poor (< 30 months) in high risk patients regardless of CR status and in low risk patients who did not achieve CR, compared to low risk patients achieving CR (57 months), making them candidates for novel post-transplant treatment options. Outcome following SCT is dependent more on biological variables such as the PCLI than on CR status. PMID- 11100276 TI - Value of autologous stem cell transplantation with purged bone marrow as first line therapy for follicular lymphoma with high tumor burden: a GOELAMS phase II study. AB - This prospective phase II study was undertaken to evaluate the efficacy and toxicity of early intensive therapy followed by purged autologous bone marrow transplantation (ABMT) in patients with follicular lymphoma with high tumor burden. All patients received the VCAP regimen (vindesine, cyclophosphamide, doxorubicin and prednisone) as conventional chemotherapy and DHAP as second-line therapy. Twenty-nine consecutive patients were included in the study. Twenty seven patients were grafted, seven in first complete remission (CR) and 20 in first partial remission (PR). Preparative therapy consisted of cyclophosphamide and total body irradiation (TBI) in all the patients. With a median follow-up of 6 years, the actuarial overall survival is 64% and the actuarial event-free survival is 55%. Two treatment-related early deaths were observed. Eleven patients were informative for serial PCR analysis of minimal residual disease after ABMT: two relapsed, four remained disease-free with PCR positivity and five were disease-free with PCR negativity. These encouraging results lay the basis of future prospective randomized trials comparing autologous stem cell transplantation as front-line treatment with conventional chemotherapy for patients with bad prognostic factors. PMID- 11100278 TI - Clinical outcome after conversion to FK 506 (tacrolimus) therapy for acute graft versus-host disease resistant to cyclosporine or for cyclosporine-associated toxicities. AB - This retrospective study describes the outcome in 53 patients who had immunosuppressive treatment changed from cyclosporine (CSP) to tacrolimus for resistant acute GVHD (n = 23), hemolytic uremic syndrome (HUS) (n = 13) or CSP associated neurotoxicity (n = 17). Tacrolimus was administered at doses of 0.03 mg/kg/day intravenously or 0.12 mg/kg/day orally in divided doses, as tolerated. Median time of conversion to tacrolimus after transplant was day 47. Nineteen patients had treatment changed to tacrolimus for resistant acute GVHD grades III or IV, with the median day of conversion being day 49 after transplant. Two of 20 evaluable patients had a complete resolution of GVHD after changing treatment to tacrolimus, with 18 showing no improvement. Eleven evaluable patients had therapy changed to tacrolimus for CSP-associated neurotoxicity at a median of 36 days after transplant. Eight patients had resolution of neurotoxicity and three had partial improvement. Eleven evaluable patients had therapy changed to tacrolimus for HUS at a median of 46 days after transplant. One patient had complete resolution of HUS and 10 showed no response. Side-effects related to tacrolimus included renal toxicity (34%), neurotoxicity (15%) and HUS (9%). Nine (17%) patients remain alive, including six patients who had therapy changed to tacrolimus for CSP-associated neurotoxicity. While often successful for dealing with neurotoxicity, only a rare patient improved after therapy was changed from CSP to tacrolimus for HUS or resistant acute GVHD. PMID- 11100279 TI - Fungal colonization and invasive fungal infections following allogeneic BMT using metronidazole, ciprofloxacin and fluconazole or ciprofloxacin and fluconazole as intestinal decontamination. AB - Invasive fungal infections (IFI) are increasingly diagnosed in patients undergoing allogeneic BMT. We have previously shown that the addition of metronidazole to ciprofloxacin for gastrointestinal bacterial decontamination significantly reduces the incidence of grades II-IV aGVHD by reduction of the anaerobic intestinal bacterial flora. Here, we found that the combined use of ciprofloxacin, metronidazole and fluconazole as antifungal prophylaxis increased intestinal yeast colonization when compared to ciprofloxacin and fluconazole alone (P < 0.01). Based on the EORTC criteria, a total of 18 out of 134 study patients developed IFI: seven of 68 (10%) patients who received metronidazole compared to 11 of the 66 (17%) patients decontaminated without metronidazole developed IFI (log-rank P = 0.36). Lethal IFI occurred in two of seven patients receiving metronidazole and in four of 11 patients without anaerobic decontamination. In conclusion, bacterial intestinal decontamination using metronidazole as an antibiotic with activity against most anaerobic intestinal bacteria significantly increases the intestinal yeast burden without influencing the incidence of IFI in patients undergoing allogeneic BMT. PMID- 11100280 TI - Invasive fungal infections in pediatric bone marrow transplant recipients: single center experience of 10 years. AB - Invasive fungal infections (IFI) with substantial mortality constitute an increasing problem among BMT patients. From 1986 to 1996 148 children underwent BMT, and are included in a retrospective analysis of the incidence, risk factors and outcome of IFI. By histopathology or culture-proven IFI (Candida, 10; Aspergillus, 8) was documented in 12/73 (16%) allogeneic and in 6/75 (8%) autologous BMT patients. Of these 18 patients, 15 subsequently died, and in 12 (66%) IFI was regarded as the main cause of death. In addition to the patients with documented IFI, 48 had suspected and 82 no fungal infection. Invasive candidal infections were more frequent in patients with semiquantitatively estimated abundant candidal colonization as compared with those with no colonization (18% vs 3%, P = 0.015). In the allogeneic group, 50% of those with severe (grades III-IV) aGVHD had IFI as opposed to 8% of those with no or mild aGVHD (P < 0.001). Regarding cGVHD, 57% of those with extensive cGVHD vs 5% of those with absent or limited cGVHD had IFI (P < 0.001). The dose of steroids was associated with IFI: 77% of those who received high-dose steroids (methylprednisolone 0.25-1 g/day for 5 days) vs 5% of those with conventional dose (prednisone 2 mg/kg/day) had IFI (P < 0.001). Particularly for BMT patients at risk, new, quicker and better diagnostic tests and more effective anti-fungal agents, both for prophylaxis and treatment, are needed. PMID- 11100281 TI - Impact of thrombotic thrombocytopenic purpura on leukemic children undergoing bone marrow transplantation. AB - Thrombotic thrombocytopenic purpura (TTP) has emerged as one of the main transplant-related complications over the last 15 years. The current study defines the incidence and the risk factors for the occurrence of TTP in 131 consecutive leukemic children who were transplanted between January 1994 and December 1997 at four Italian pediatric centers. Patients with ALL (101), AML (21), MDS (9), underwent an HLA-identical sibling BMT (82) or an HLA-identical unrelated BMT (49), receiving a conditioning regimen consisting of high-dose chemotherapy in 24 patients and of F-TBI combined with high-dose chemotherapy in 107 patients. The diagnosis of TTP was retrospectively evaluated on the basis of parallel criteria. TTP treatment varied according to the protocol of each treatment center. Twenty-eight of 131 patients (21.4%) developed TTP at a median of 46 days (range 21-80) after BMT. Multivariate analysis demonstrated that the risk of TTP was higher in patients who underwent unrelated BMT (P value = 0.02). Acute GVHD, stage of disease at BMT, conditioning with TBI, gender, age, did not appear to be associated with the occurrence of TTP. As to the outcome, TTP resolved in 19 patients while in nine it was the principal cause of death (32.1%). In patients with TTP, LDH peak value was the only statistically significant factor (P = 0.001) related to severe TTP. In conclusion, our experience demonstrates that leukemic children undergoing BMT, especially from an unrelated donor, should be carefully assessed for TTP which appears to be a severe and relatively common transplant-related complication when strict diagnostic criteria are applied. PMID- 11100282 TI - Extramedullary relapse after allogeneic bone marrow transplantation for haematological malignancy. AB - We describe the risk factors for and the natural history and response to treatment of extramedullary (EM) relapse in 183 patients who underwent allogeneic bone marrow transplantation (alloBMT) for a variety of haematological malignancies at our institution over a 7 1/2 year period. Fifty-one patients relapsed; 15 had EM relapse either alone or in association with marrow involvement. A retrospective analysis found that the presence of chronic GVHD and a longer interval between transplant and relapse were independently associated with an increased risk of EM compared to marrow-only relapse. EM relapse was also associated with a longer post-relapse survival. Patients with EM relapse appeared to respond to cytotoxic therapy but not to DLI. EM relapse after alloBMT may be more common than previously thought and have a better prognosis than marrow-only relapse. While patients developing chronic GVHD after alloBMT have a lower overall relapse risk than those who do not, they may be more prone to delayed relapse at EM sites. PMID- 11100283 TI - Recurrent penicillin-resistant pneumococcal sepsis after matched unrelated donor (MUD) transplantation for refractory T cell lymphoma. AB - Patients who undergo splenectomy and recipients of allogeneic marrow (alloBMT) or peripheral stem cell transplantation are at increased risk of overwhelming infection from encapsulated organisms such as Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis. As prophylaxis against these pathogens splenectomised patients are immunised and may also receive antibiotics for life. We report relapsing overwhelming sepsis caused by penicillin-resistant pneumococcus in a patient who was immunised and received prophylactic phenoxymethylpenicillin for 8 months following splenectomy and matched unrelated donor (MUD) marrow transplantation for refractory T cell lymphoma. No obvious focus of sepsis was found during any of the three episodes and S. pneumoniae serogroup 6, subtype 6B was isolated from blood cultures on each occasion. He was treated with i.v. cephalosporins, as the organisms were resistant to penicillin with a minimum inhibitory concentration (MIC) of 2.0, and there was complete resolution of symptoms each time. In the light of recurrent sepsis with this penicillin-resistant organism the decision was made to give prophylactic levofloxacin for the next 12 months. This case illustrates that the choice of prophylactic regimen and the treatment of sepsis in immunocompromised patients remain difficult and challenging issues. PMID- 11100284 TI - Post-transplant lymphoproliferative disorder after autologous peripheral stem cell transplantation in a pediatric patient. AB - Post-transplant lymphoproliferative disorder (PTLD) is a complication of allogeneic bone marrow transplantation (BMT). Rare cases of PTLD after autologous BMT have been reported only in adults. This case report is the first to describe PTLD in a pediatric patient after autologous peripheral stem cell transplantation (PSCT). This 2-year-old male with stage IV neuroblastoma underwent autologous PSCT. The post-PSCT course was complicated by fever with hematochezia and a lung mass. On day 94 post PSCT, colonoscopy revealed an ulcer due to a PTLD, monomorphic type, B cell phenotype, associated with Epstein-Barr virus. Fine needle aspiration identified the lung mass as neuroblastoma. PTLD can occur in pediatric autologous PSCT recipients, and may occur more frequently in autologous grafts manipulated by T cell depletion or CD34+ cell selection. PMID- 11100285 TI - Successful treatment of invasive aspergillosis in chronic granulomatous disease by granulocyte transfusions followed by peripheral blood stem cell transplantation. AB - Chronic granulomatous disease (CGD) is a primary immunodeficiency disorder characterized by impaired microbial killing and susceptibility to bacterial and fungal infections. Cure of the disease can be achieved by stem cell transplantation when performed early in its course, and before severe infections have developed. Invasive aspergillosis constitutes a very high risk for transplantation. We report a 4-year-old boy with X-linked CGD who underwent successful HLA-identical peripheral blood stem cell (PBSC) transplantation during invasive pulmonary aspergillosis and osteomyelitis of the left fourth rib, which was unresponsive to antifungal treatment. During the 2 months prior to the transplant he received G-CSF-mobilized granulocyte transfusions (GTX) from unrelated donors three times a week in addition to the antifungal treatment. This resulted in clinical improvement in his respiratory status. He also received GTX during the aplastic period after the conditioning regimen, until he had engrafted. Post-transplant superoxide generation test revealed that neutrophil function was within normal range. One year post transplant the CT scan showed almost complete clearance of the pulmonary infiltrates and a marked improvement in the osteomyelitic process. Based on other reports and our own experience, GTX can serve as important treatment in patients with CGD who have failed conventional anti-fungal treatment and for whom stem cell transplantation is the only chance for cure. PMID- 11100286 TI - High-dose chemotherapy with autologous stem cell support in an adult with bone marrow relapse of medulloblastoma. PMID- 11100287 TI - Ethical considerations in allogeneic hematopoietic cell transplantation for children with slowly fatal conditions. PMID- 11100288 TI - Clinical problem of preterm labor. PMID- 11100289 TI - Endocrinology of preterm labor. AB - The clinical importance of preterm labor and delivery dictates that we understand the physiology and diagnostic usefulness of endocrine as well as other agents that may be helpful in this regard. Clearly, estrogen and progesterone establish the environment that allows parturition and probably preterm labor to occur. The use of salivary estriol, though not a foolproof test, is becoming more frequent and is commercially available. Fibronectin, though not an endocrine test, has a similar diagnostic usefulness. In the future, we would expect to see CRH and even the use of selective cytokines, probably IL-6, as possible diagnostic tests. Whereas all of these agents have some diagnostic usefulness, none of them can be expected to predict every case of preterm delivery and some battery of tests, not unlike the triple or quadruple tests that are used for prenatal diagnosis of Down syndrome, may be effective and should be examined in the future. The use of these tests, salivary estriol and fetal fibronectin in particular, has already had an effect on management and decision making involved in preterm labor, and the future should give us more options and hopefully, better choices to manage this most difficult condition. PMID- 11100290 TI - Risk factors for preterm labor. PMID- 11100291 TI - Cervical length and preterm labor. PMID- 11100292 TI - Use of fetal fibronectin in women at risk for preterm delivery. AB - Fetal fibronectin, a large molecular weight glycoprotein produced in the chorion, is expressed in cervical and vaginal secretions in women with disruption of the choriodecidual [table: see text] junction by labor or by inflammation. The presence of FFN in vaginal or cervical secretions before 35 weeks is a moderately good predictor of preterm delivery. The absence of FFN is a strong predictor that preterm delivery is unlikely within the next 7 to 14 days, with NPVs exceeding 99% in some studies. The predictive power of FFN is stronger at earlier gestation ages (24-28 wks) than it is later [table: see text] in pregnancy and is stronger for short-term prediction (7-14 d) than for predicting overall outcome (however, it remains statistically significant for predicting delivery < 37 wks). Although use of FFN in the clinical setting may require some changes to common protocols (e.g., performing sterile speculum examination before digital cervical examination), the use of FFN in patients with suspected preterm labor appears to have significant utility in reducing unnecessary interventions in women with symptoms suggestive of preterm labor. In women without symptoms, the use of FFN may be most beneficial in providing reassurance to some women thought to be at high-risk for preterm delivery because of past obstetric history. Screening women without symptoms at low-risk with FFN is not yet recommended because effective interventions are not demonstrated for patients found to be positive. PMID- 11100293 TI - Infection and preterm labor. AB - There are many conditions, such as non-white race, young maternal age, and uterine malformations, that have been associated with preterm birth that are not amenable to intervention. Maternal cervical and intrauterine infection and inflammation may have a primary causative role in a fraction of the cases of preterm birth and preterm rupture of membranes and may also interact adversely with a variety of maternal (shortened cervix, smoking) and fetal factors (polyhydramnios, multifetal gestation) to decrease the threshold to preterm birth. Further studies are needed to better-define the link between various maternal microbial colonizations and preterm delivery, with the possibility to establish new screening and treatment recommendations. Because of the innumerable causes of preterm birth, a new strategy of targeted treatment of cervical or vaginal infections may lead to only a modest reduction in the incidence of this devastating problem of modern obstetrics. PMID- 11100294 TI - How to diagnose preterm labor: a clinical dilemma. AB - The clinician is faced with many dilemmas in the diagnosis of preterm labor. The tools at hand (no pun intended) yield subjective information and it is difficult to distinguish true labor from false labor. Because preterm labor is multifactorial in cause and can occur in primiparas, estimating risk for the individual patient is difficult. The cause of the preterm labor in the majority of patients is idiopathic; in this group, the use of salivary estriol as a biochemical marker for preterm labor can increase the accuracy of the diagnosis of true labor. It may also lend confidence to the diagnosis of false labor and may allay anxiety and prevent unnecessary interventions. On the horizon are two noninvasive methods: the EMG, to evaluate uterine contractility, and the collascope, to evaluate the cervix. Both show promise and may provide a more objective assessment of risk for preterm delivery among women with symptoms of preterm labor. PMID- 11100295 TI - Home uterine activity monitoring: a critical review. PMID- 11100296 TI - Therapeutic agents in preterm labor: tocolytic agents. PMID- 11100297 TI - Therapeutic agents in preterm labor: steroids. PMID- 11100298 TI - Antibiotics and preterm labor. AB - In summary, a definite association has been demonstrated between preterm labor and genital tract infection. Conclusions regarding the true benefits of antibiotics as adjunctive therapy in treatment of preterm labor are inconsistent. Whereas some of the studies were able to demonstrate significant prolongation of pregnancy, no consistent reduction in either maternal or neonatal morbidity has been demonstrated. However, because the actual incidental morbidity rate is low in the populations studied, the power of this finding is also low. The potential risks for using antimicrobials has yet to be adequately addressed. It has been shown that bacterial resistance can develop when antibiotics are used without specific aim or when a specific bacteria is undertreated. It has been recently shown that prenatal and intrapartum antibiotic use is associated with an increased risk for antibiotic resistant neonatal sepsis if infection occurs. Because of these reasons, we discourage the administration of antibiotic treatment to women in preterm labor for the purpose of pregnancy prolongations. Treatment should be directed towards those with specific indications for treatment (e.g., intrapartum, group B streptococci prophylaxis, urinary tract infection, etc). The primary flaw in these many evaluations of preterm labor is the true incidence of preterm birth. The clinical diagnosis of preterm labor is a difficult one. Approximately one-half of those individuals with preterm contractions will not deliver until term. So, the use of antibiotics for all women in idiopathic preterm labor is destined to treat many women who are unlikely to benefit. If we were able to truly identify those who were in "true" labor, perhaps we could be more selective in determining who may benefit from antibiotics. Biochemical markers such as onco-fetal fibronectin could well-be a helpful marker. Goldberg et al evaluated FFN in vaginal and cervical secretions while attempting to better-predict who would have upper genital tract infection. In this large, multicenter trial, patients were tested for FFN every 2 weeks from 23 to 30 weeks gestation. In those patients who proceeded to deliver before 32 weeks gestation, increased levels of cervical FFN (> 50 ng/ml) were identified in approximately one-quarter. Fetal fibronectin was positive in 4% of their samples and was found to be twice as likely in one with bacterial vaginosis. They showed that the presence of increased FFN was associated with upper genital tract infection (clinical and histologic chorioamnionitis) as a main reason for preterm labor and delivery (increased risk 16-20-fold). Those with increased FFN levels were also shown to have an increased incidence of neonatal sepsis as well. Peaceman et al used FFN to attempt to identify those at risk for preterm delivery among women with contractions between 24 and 34 6/7 weeks gestation. Those with negative FFN were less likely to deliver within 7 days of the test. The negative predictive value was 99.7%, suggesting that this test may be helpful in identifying women who would not benefit from antibiotic treatment. However, if in the absence of prospective clinical trials demonstrating the efficacy of this approach, we discourage the use of FFN screening for this indication. PMID- 11100299 TI - Environmental factors in infertility. AB - In conclusion, several studies indicate that there is an association between cigarette smoking and adverse reproductive outcomes in women as well as men. Some studies indicate that alcohol consumption impairs the reproductive capacity of women. Exposures to PCE in the dry cleaning industry, toluene in the printing business, ethylene oxide and mixed solvents have been associated with decreased fecundity. Abnormalities in sperm production have been found in men exposed to radiant heat or heavy metals. Environmental exposure to chlorinated hydrocarbons (e.g., DDT, PCB, pentachlorophenol, hexachlorocyclohexane) has been associated with an increase in rates of miscarriage and endometriosis. Clinicians should counsel patients who are trying to achieve a successful pregnancy to stop smoking and limit alcohol intake. Clinicians can additionally counsel patients who are in contact with potentially harmful occupational and environmental toxicants to limit their exposure. It is important to recognize, however, that many of the studies to date are limited by small sample size, poor exposure assessment, poor outcome measurements, recruitment bias, or recall bias. Additional studies will be necessary to clarify the magnitude of risk associated with these factors. PMID- 11100300 TI - Immunologic factors in infertility. PMID- 11100301 TI - Evaluation of the couple with infertility in a managed care environment. AB - The classic infertility evaluation is a costly experience for many couples. In the new era of managed care, it may be possible to perform a targeted evaluation that, while significantly lowering the total cost, will not impair our ability to correctly diagnose the cause of a couple's infertility. When combined with algorithms for streamlined treatment, it may be possible to significantly reduce the cost for the diagnosis and treatment of infertility. PMID- 11100302 TI - Evaluation and treatment of male infertility. PMID- 11100303 TI - Endoscopy in the evaluation of the woman experiencing infertility. PMID- 11100304 TI - Unexplained infertility. PMID- 11100305 TI - Ovulation induction. AB - In the woman with anovulation and polycystic ovarian syndrome, there are many options for ovulation induction. Treatment should be individualized, but clomiphene citrate is an excellent first-line agent. In the woman resistant to clomiphene citrate, combination therapy often results in pregnancy. Some women with PCOS only respond to gonadotropin therapy. These women are at a higher risk for multiple pregnancy and ovarian hyperstimulation syndrome. In the woman with anovulation and hypothalamic amenorrhea, the options for ovulation induction are limited. The luteal phase must be supported. The hypothalamus is unable to support the corpus luteum or early pregnancy. PMID- 11100306 TI - Surgical management of adhesions, endometriosis, and tubal pathology in the woman with infertility. AB - There are many considerations in the surgical treatment of patients with infertility. Of prime importance is the baseline condition of the tubes and skill of the surgeon. With further advances in the understanding of the process of fertilization and implantation, it is anticipated that the use of surgical methods and application of new technologies will continue to improve fecundity for patients with infertility. PMID- 11100307 TI - Infertility: is there a role for the surgeon. AB - No matter how successful IVF and intracytoplasmic sperm injection become, there will always be a role for the infertility surgeon in the care of the woman with infertility. The challenge is to develop evidence-based protocols that will specify when surgery is clearly a better choice than IVF. It will also be important to develop training programs in pelvic surgery that will teach techniques and strategies designed to maintain or improve fertility as well as remove or destroy pelvic pathology. With a decreasing number of surgical procedures available to train residents in infertility surgery, it will become increasingly important to consider alternative models such as virtual reality and postresidency fellowships or preceptorships. PMID- 11100308 TI - An appreciation of modern ART. AB - As we have entered into the new millennium, it is difficult not to recognize ART as one of the most dynamic and rapidly emerging fields in all of medicine. What began as an experimental procedure in animals has developed into a multidisciplinary technology. A great debt is owed to the field of animal husbandry. In humans, implantation is inefficient and has been recognized as the rate-limiting step in reproduction. In vitro fertilization has allowed the observation of human gamete interaction in the laboratory. A milestone was reached when ART allowed couples with infertility to have success rates that exceeded those associated with normal human fecundity. Continued innovations are improving the rate of embryonic implantation. Blastocyst transfer will have a major role in the future of ART. It is common for the couple with infertility to battle their problem by increasing the frequency of intercourse because it is the only weapon at their disposal. So, too, did clinicians increase the number of embryos transferred to the uterus. New developments promise to challenge the long held contention that successful IVF/embryo transfer is positively correlated with the number of oocytes retrieved or the number of embryos transferred. More accurate reporting will enable us to measure the impact of technologic improvements that improve implantation rates and decrease multiple gestations. The future holds the promise of some tangible goals. The development of in vitro maturation of immature oocytes could lead to an era in which oocytes are harvested without the need for controlled ovarian hyperstimulation. This would provide for the possibility of the banking of gametes. Women could preserve or store their fertility at a young age in a manner similar to that which has been possible for men for decades. They could avoid the increased risk for aneuploidy seen with the age of 35, and theoretically, they should have a lower risk for miscarriage. The future is near. One day, ART will be perfected so that a single embryo transfer will be the standard of care. Soon it will be possible to know all about the genetic makeup of that embryo and it will be routinely selected from its cohort by virtue of those genetic traits. PMID- 11100309 TI - Cost-effective treatment for the couple with infertility. AB - Although the evaluation of cost-effective approaches to infertility treatment remains in its infancy, several important principles have emerged from the initial studies in this field. Currently, in treating couples with infertility without tubal disease or severe male-factor infertility, the most cost-effective approach is to start with IUI or superovulation-IUI treatments before resorting to IVF procedures. The woman's age and number of sperm present for insemination are significant factors influencing cost-effectiveness. The influence of certain diagnoses on the cost-effectiveness of infertility treatments requires further study. Even when accounting for the costs associated with multiple gestations and premature deliveries, the cost of IVF decreases within the range of other cost effective medical procedures and decreases to less than the willingness to pay for these procedures. Indeed, for patients with severe tubal disease, IVF has been found to be more cost-effective than surgical repair. The cost-effectiveness of IVF will likely improve as success rates show continued improvements over the course of time. In addition, usefulness of embryo selection and practices to reduce the likelihood of high-order multiple pregnancies, without reductions in pregnancy rates, will significantly impact cost-effectiveness. The exclusion of infertility treatments from insurance plans is unfortunate and accentuates the importance of physicians understanding the economics of infertility treatment with costs that are often passed directly to the patient. The erroneous economic policies and judgments that have led to inequities in access to infertility health care should not be tolerated. PMID- 11100310 TI - Intraperitoneal hemorrhage as a major complication of percutaneous ethanol injection therapy for hepatocellular carcinoma. AB - Two cases of intraperitoneal hemorrhage, which is one of the major complications of percutaneous ethanol injection therapy for hepatocellular carcinoma, are reported. A 70-year-old man was hospitalized for treatment of a small recurrent hepatocellular carcinoma located on the surface of the left lobe of the liver. Acute hemoperitoneum developed after percutaneous ethanol injection therapy, but he was treated conservatively with blood transfusion, and recovered. The other patient was a 72-year-old man who was admitted for treatment of a solitary superficial hepatocellular carcinoma on the dome of the liver. Immediately after percutaneous ethanol injection, he suffered the sudden onset of severe abdominal pain with shock and massive hemoperitoneum. His bleeding was successfully controlled by emergency transcatheter arterial embolization. Our experience suggests that care must be taken when using percutaneous ethanol injection to treat patients with superficial hepatocellular carcinomas located on the surface of the liver. Moreover, transcatheter arterial embolization should be considered the treatment of choice for the management of uncontrollable intraperitoneal hemorrhage after percutaneous ethanol injection therapy. PMID- 11100311 TI - Restoration of propulsive peristalsis of the esophagus in achalasia. AB - The set consisting of 3 patients with esophageal achalasia diagnosed by manometry, pseudoachalasia excluded by esophagoscopy and endosonography, was treated with combined conservative procedure. Botulinum toxin 250u (Dysport) was applied to the area of lower esophageal sphincter and after 7 days balloon dilatation was carried out. Treatment efficacy was evaluated by the data obtained about the subjective condition, manometrically and endoscopically. The spine condition was evaluated in all patients before treatment and functional blockades were released by manual medicine and even by acupuncture. We succeeded in restoring propulsive peristalsis of the esophagus in all of them. It is objectively proven in the longest duration of 44 months in the case of a patient treated with a balloon dilatation. PMID- 11100312 TI - Laparoscopic Sugiura procedure for conditioning of the blood stream through TIPSS in cirrhotic patient. Initial experience. AB - The Sugiura procedure or the proximal gastric devascularisation was formerly the method of choice for esophageal varicose treatment in some patients. The frequency of this operation decreased stenting after the introduction of the transjugular portosystemic shunt into clinical practice. However this method performed laparoscopically could be useful as a complementary procedure when the blood stream through the transjugular intrahepatic portosystemic stent shunt is low and an esophageal rebleeding occurs. A 40-year old patient with hepatic cirrhosis and Child stage "B" was admitted to our clinic due to recurrent esophageal varicose bleeding. He underwent a transjugular intrahepatic portosystemic stent shunt implantation 27 months before the admission and the transjugular intrahepatic portosystemic stent shunt became occluded 3 times since implantation and was repeatedly revised. After admission a color Doppler of the stent was performed. The blood stream was 15 cm/s. The laparoscopic Sugiura procedure was performed after conditioning of the general status of the patient. Five ports were introduced 5 cm above the umbilicus, under the xiphoid, the right and left hypochondrium as well as the left mesogastrium. The dilated veins between the gastric coronary vein and esophagus and the short gastric veins on the great curvature were interrupted by means of an ultrasonic scalpel. The hiatus esophagus was opened, the esophagus was prepared up to the first pulmonal vein and the dilated esophageal veins were occluded with stitch ligatures. The operation was completed with Toupet partial fundoplication. The patient was followed 6 months after the surgery. No rebleeding occurred in this time period. Immediately after surgery the blood flow increased up to 97 cm/s; at 3 and 6 months follow-up the intrahepatic shunt remained open with 82 and 80 cm/s blood flow respectively. Laparoscopic Sugiura procedure performed as a complementary surgery after transjugular intrahepatic portosystemic stent shunt implantation increased blood perfusion through the intrahepatal constructed shunt and prevented its occlusion. However this initial experience has to be confirmed with a larger number of patients. PMID- 11100313 TI - Technical dilemma in living-donor or split-liver transplant. AB - In partial liver transplantation for adults criteria for the extent of reconstruction of middle hepatic vein tributaries have not been clarified. After hepatic venous and portal anastomoses in living-donor liver transplantation using left liver graft without middle hepatic vein, color Doppler ultrasonography was applied to check venous and portal blood flow. Color Doppler ultrasonography demonstrated absent hepatic venous flow and reversed portal venous flow in the congested area of the left paramedian sector which had been drained by the divided branch of the middle hepatic vein. The area was darkly discolored before arterial reperfusion and under clamping of the artery. Reconstruction of the venous branch was added after arterial anastomosis. Color Doppler ultrasonography revealed restored normal venous outflow and portal inflow after venous reconstruction. Postoperative course of the recipient was uneventful with rapid recovery of liver function. We propose that middle hepatic vein tributaries should be reconstructed if color Doppler ultrasonography demonstrates absent venous flow and reversed portal flow, and if the liver volume excluding the discolored area under occlusion of the hepatic artery is estimated to be insufficient for postoperative metabolic demand. PMID- 11100314 TI - Hepatobiliary scintigraphy after biliary reconstruction--a comparative study on Roux-Y and ESCD. AB - BACKGROUND/AIMS: Reconstruction of extrahepatic biliary tract with benign lesion still has some unsettled problems, such as postoperative cholangitis. This study was conducted to compare bile through the remnant alimentary tract in patients undergoing end-to-side choledochoduodenostomy, and those undergoing Roux-Y choledochojejunostomy, using hepatobiliary scintigraphy. METHODOLOGY: Five normal human volunteers and 13 patients underwent end-to-side choledochoduodenostomy (n = 5), Roux-Y choledochojejunostomy (n = 8), using hepatobiliary scintigraphy. RESULTS: Postoperative acute cholangitis developed in 1 patient (12%) with Roux-Y choledochojejunostomy and none with end-to-side choledochoduodenostomy. Hepatobiliary scintigraphy showed prominent stasis of 99mTc in the proximal jejunum loop of the patients who underwent the Roux-Y choledochojejunostomy procedure, which was not found in the upper jejunum of the patients with end-to side choledochoduodenostomy. The time taken before visualization of 99mTC at the upper jejunum in the patient who underwent Roux-Y choledochojejunostomy (66.5 +/- 5 min) was significantly longer than that in the healthy controls (40 +/- 5 min). On the other hand, the time taken before visualization of 99mTc at the upper jejunum in end-to-side choledochoduodenostomy (35 +/- 5 min) was similar to that of healthy controls. CONCLUSIONS: This data suggested that end-to-side choledochoduodenostomy procedure for reconstructing the extrahepatic biliary tract was more physiological with less postoperative complication than Roux-Y choledochojejunostomy procedure. PMID- 11100315 TI - Selective use of perioperative ERCP in patients undergoing laparoscopic cholecystectomy. AB - BACKGROUND/AIMS: Management of common bile duct stones in the era of laparoscopic surgery is still controversial. The purpose of this study is to investigate the safety, feasibility, success rate and short-term results of the selective use of endoscopic retrograde cholangiopancreatography in patients undergoing laparoscopic cholecystectomy. METHODOLOGY: A prospective study comprising 300 consecutive patients with either symptomatic or complicated gallbladder stones was performed between January 1994 and November 1996. Depending on clinical, laboratory and ultrasonographic criteria, 73 patients (24.3%) underwent endoscopic retrograde cholangiopancreatography with or without endoscopic sphincterotomy. The procedure was successful in 71 patients (97%) either preoperatively in 62 patients (21%) or postoperatively in 9 patients (3%). RESULTS: Endoscopic retrograde cholangiopancreatography was positive in 37 cases (52%), endoscopic sphincterotomy and stone extraction was performed in 35 cases and endoscopic sphincterotomy alone was performed in 2 cases for benign papillary stenosis. The overall predictive value for the presence of common bile duct stone was 52%, the predictive value for patients with jaundice, dilated common bile duct together with elevated liver enzymes was 73.3%. Complications of perioperative endoscopic retrograde cholangiopancreatography were encountered in 4 patients (5.5%) with no mortality. CONCLUSIONS: We conclude that the combination of perioperative endoscopic retrograde cholangiopancreatography and laparoscopic cholecystectomy is a useful approach for the management of choledochocholelithiasis. PMID- 11100316 TI - Alterations of retinoblastoma protein and p16INK4 protein expression in extrahepatic bile duct carcinomas. AB - BACKGROUND/AIMS: Human cancer development results from dysfunction of G1-phase regulators of the cell cycle. Retinoblastoma protein and p16INK4 are the most essential links between cell cycle control and cancer. We examined the expression of p16INK4 and pRb and their possible prognostic relevance in 34 extrahepatic bile duct carcinomas. METHODOLOGY: Expression of pRb and p16INK4 was determined using immunohistochemical techniques. Associations between expression of pRb and p16INK4 and the clinicopathological features were analyzed by using the chi 2 test and survival analysis was performed by Log-rank test. RESULTS: Two (6%) extrahepatic bile duct carcinomas were pRb negative, 26 (76%) showed pRb overexpression, and 6 (18%) demonstrated moderate expression. Twenty-two (65%) tumors were p16INK4 negative and 12 (35%) were p16INK4-positive. Cases with pRb negative or pRb overexpression were significantly correlated with tumor progression (P = 0.004) and TNM stage (P = 0.009). Alterations in pRb and p16INK4 expression did not correlate with patient outcome. CONCLUSIONS: Alterations of pRb and p16INK4 expression are frequently involved in extrahepatic bile duct carcinomas, and that aberrant pRb expression significantly associates with tumor progression. PMID- 11100317 TI - Laparoscopic cholecystectomy and perioperative ERCP. AB - BACKGROUND/AIMS: Study of acceptance of simultaneous laparoscopic cholecystectomy and endoscopic retrograde cholangiopancreatography for treatment of cholelithiasis with choledocholithiasis. METHODOLOGY: There were 25 patients. Ten patients had acute pancreatitis of biliary etiology, accompanied by transitory icterus. In 15 patients, choledocholithiasis was suspected preoperatively both on ultrasonography and i.v. cholangiography. In all patients laparoscopic cholecystectomy with perioperative endoscopic retrograde cholangiopancreatography and sphincterotomy were performed for the treatment of cholelithiasis and choledocholithiasis. RESULTS: Simultaneous laparoscopic cholecystectomy and endoscopic retrograde cholangiopancreatography was successfully done in all patients. The patients were discharged home on the 4th day after the surgery. Concerning early complications, there where 3 early complications, e.g., prolonged hemorrhage after papillotomy in a patient with choledocholithiasis with stenotic papillitis. Conservative therapy (fresh frozen plasma, local hemostats) was used in this patient. In 4 patients with choledocholithiasis, transitory hyperamylasemia was observed, with no clinical symptoms of pancreatitis. The symptoms disappeared with conservative therapy 3 days after the beginning of treatment. CONCLUSIONS: Simultaneous laparoscopic cholecystectomy and endoscopic retrograde cholangiopancreatography for treatment of cholelithiasis and choledocholithiasis is a safe and acceptable treatment. PMID- 11100318 TI - Inhibited serum phospholipase A2 activity in hyperbilirubinemia. AB - BACKGROUND/AIMS: Phospholipase A2 activity is reported to be related to the physiology of various disorders, and the activity is modified by deoxycholic acid. METHODOLOGY: Serum phospholipase A2 activity was measured by counting the radioactivity of 14C-palmitic acid released from L-3-phosphatidylcholine (1 palmitoyl-2-14C-palmitoyl) in 40 patients with hyperbilirubinemia due to malignant neoplasms and benign cholestasis. Phospholipase A2 activity in serum from healthy subjects was also measured after incubation with 5-30 mg of bilirubin per dL or 0.1-5 mM of cholic acid for 60 min at 37 degrees C. RESULTS: Serum phospholipase A2 activity was significantly lower in patients with hyperbilirubinemia. There were negative correlations between serum phospholipase A2 activity and concentration of total bilirubin (r = 0.668; P < 0.0001) or total bile acids (r = 0.423; P < 0.05) in patients with hyperbilirubinemia. In 12 patients who underwent percutaneous transhepatic biliary drainage, the low serum phospholipase A2 activity was reversed while bilirubin concentrations decreased. Incubation of sera from healthy subjects with more than 3 mM cholic acid significantly reduced phospholipase A2 activity (P < 0.01), whereas incubation with bilirubin had no effect. CONCLUSIONS: Inhibition of serum phospholipase A2 activity in patients with hyperbilirubinemia may be caused by increased serum cholic acid concentration. PMID- 11100319 TI - Left hepatic duct injury and thoracobiliary fistula after abdominal blunt trauma. AB - Thoracobiliary fistula after blunt hepatic trauma is rare. We report a case of pleurobiliary fistula after a blunt hepatic trauma leading to a left hepatic lobe laceration together with a left hepatic duct injury. The management of this traumatic lesion is discussed and related to the existing literature data. The diagnosis of traumatic thoracobiliary fistula rests upon clinical suspicion in the setting of a persistent right pleural effusion. Demonstration of the presence of bile in the pleural cavity by thoracocentesis is considered a proof of pleural biliary fistula. We think that laparotomy is an appropriate route for the treatment of pleurobiliary fistulas. However, when a bronchobiliary fistula is suspected, the patient should be treated with thoracotomy and may require pulmonary resection to remove the fistulous tracts. PMID- 11100320 TI - Mass-forming inflammatory periductal fibrosis mimicking hilar bile duct carcinoma. AB - A case of a rare benign biliary lesion at the hepatic hilum mimicking hilar bile duct carcinoma is reported. A 73-year-old man was found to have gastric cancer by gastrointestinal fiberscopy. Dilated right intrahepatic bile ducts and a 2-cm mass in the right hepatic duct were demonstrated by further imaging investigations. He was finally diagnosed as having hilar bile duct and gastric carcinomas, and underwent right portal vein embolization followed by a single stage extended right hepatectomy and total gastrectomy. Pathologically, however, the lesion in the right hepatic duct showed inflammatory changes with periductal fibrosis, without any signs of malignancy. A literature search revealed 11 such cases including the present one. PMID- 11100321 TI - Undifferentiated spindle cell carcinoma of the extrahepatic bile ducts: a case report. AB - Spindle cell carcinoma is a rare tumor that generally occurs in the upper digestive tract. We report an 81-year-old man with spindle cell carcinoma located in the extrahepatic bile ducts, resulting in obstructive jaundice. The patient died 10 months after operation due to local recurrence. The literature on this rare disease is reviewed and discussed. PMID- 11100322 TI - Mucosal dysplasia in the bile duct after choledochoduodenostomy: a case report. AB - A 60-year-old woman who had undergone cholecystectomy, choledocholithotomy and choledochoduodenostomy 21 years previously for cholecystolithiasis and choledocholithiasis, presented with nausea and vomiting. With a preoperative diagnosis of recurrent common bile duct stones, the extrahepatic bile duct was excised and choledochojejunostomy was performed. Histologic examination of the resected specimen disclosed chronic cholangitis, papillary epithelial hyperplasia, and mild dysplasia. Choledochoduodenostomy predisposes to reflux of duodenal contents, resulting in chronic mechanical and chemical irritation likely to induce histopathologic alterations in the bile duct mucosa. Since bile duct dysplasia induced by chronic inflammation may be a precursor of cancer, indication for choledochoduodenostomy should be specific and limited, and careful long-term follow-up is mandatory. PMID- 11100323 TI - Use of autologous blood in general surgery. AB - BACKGROUND/AIMS: Autologous blood predonation is still not as widespread as it should be in general surgery practice, even if the method is well-known and has benefits established in international literature. Authors describe the impact of an autotransfusion program, in a general surgery university department, focusing on management and cost problems. METHODOLOGY: A description of the efficacy of the program during a yearlong activity period is presented. An analysis has been made about the quantity of predonated blood/plasma units, the quantity actually transfused and use of homologous blood. The problems which occurred and the cost are discussed. RESULTS: The most used autotransfusion method was preoperative predeposit of autologous blood. The analysis of results focused on some organizational problems that need to be avoided in order to show the methods maximum benefits. In a large number of cases (some 50%) predeposit was not made because of several managing/technical problems. In another large number of cases (38%) the quantity of units predonated did not fully supply the needs and several patients received homologous products. In another number of cases predonated blood units were not used at all (61/34%). CONCLUSIONS: Predeposit, preoperative hemodilution and intraoperative recovery, are methods that should all be available in a general surgery department to manage in the best way the single patients blood/plasma needs, reducing post-transfusion complication. To optimize the program and minimize waste some guidelines must be established, with the aim of a rational and correct use of the procedure. Despite the value of the method, and the favor encountered by the patients, we must not forget that the use of autologous blood is not costless. PMID- 11100324 TI - Intraportal administration of low-dose recombinant human hepatocyte growth factor enhances effects of hepatocellular transplantation. AB - BACKGROUND/AIMS: Although recombinant human hepatocyte growth factor (rhHGF) is a potent mitogen, the dose used for patients is still not clear and must be low to avoid untoward effects. Firstly, the optimal strategy of the dose and route of rhHGF was investigated. Secondly, low-dose rhHGF, which would induce proliferation of transplanted hepatocytes, was explored using Nagase analbuminemic rats. METHODOLOGY: 1) Concentrations of rhHGF in the portal vein were measured after continuous administration of titrated rhHGF through the jugular vein or portal vein. 2) F344 rat hepatocytes (2 x 10(7) cells) were transplanted in the liver of Nagase analbuminemic rats. On the 7th day, the rats were subjected to a low-dose rhHGF treatment. RESULTS: When the rats were given rhHGF in a dose of 50 micrograms/kg/day, the mean concentration in the portal vein (0.8 +/- 0.1 ng/mL) was almost similar to the minimum concentration which stimulated hepatocyte proliferation in vitro. When low-dose rhHGF (50 micrograms/kg/day) was administered directly into the portal vein following hepatocyte transplantation in Nagase analbuminemic rats, the serum levels of albumin were significantly higher than in other groups. It was found that the concentration of rhHGF in the portal vein were 3.1 +/- 0.5 ng/mL with continuous intraportal infusion and 0.8 +/- 0.1 ng/mL with continuous systemic infusion. CONCLUSIONS: It was found that the minimal dose of rhHGF needed to stimulate hepatocyte proliferation was 50 micrograms/kg/day. With rhHGF (50 micrograms/kg/day), continuous intraportal infusion afforded a more favorable outcome in case of proliferation of hepatocytes. PMID- 11100325 TI - Nitric oxide induction in a rat model of selective pancreatic ischemia and reperfusion. AB - BACKGROUND/AIMS: Ischemia and reperfusion of the pancreas may be important in aggravating the course of acute pancreatitis. In a rat model of selective pancreatic ischemia and reperfusion, we studied plasma levels of nitric oxide and expression of nitric oxide synthase in the pancrease and lung. METHODOLOGY: Pancreatic ischemia was achieved by occlusion of the 4 main pancreatic arteries for 40 min; this was followed by a 7-hour reperfusion period (group A, 10 rats). Outcome measures were compared with those of animals undergoing a sham operation (group B, 10 rats). RESULTS: Pancreatic damage in group A animals was demonstrated by increased serum alpha-amylase and by macroscopic and microscopic evidence. Total nitric oxide synthase activity in pancrease and lung was higher than in shams [median: 0.73 vs. 0.54 pmol/mg protein/min in the pancreas (P = 0.0082); 1.38 vs. 0.68 pmol/mg protein/min in the lung (P = 0.023)]; this was mainly due to activation of the inducible isoform of the enzyme. There was an associated 58.2% increase in plasma levels of nitric oxide metabolites [from mean 55.0 to 131.6 mumol/L (P < 0.001)]. Immunohistochemistry confirmed expression of inducible nitric oxide synthase and nitric oxide-mediated oxidative damage (nitrotyrosine) in both pancreas and lung. CONCLUSIONS: Ischemia and reperfusion of the pancreas induces pancreatic damage, overexpression of inducible nitric oxide synthase and oxidative damage within the pancreas and lung. PMID- 11100326 TI - An in-vitro model of scirrhous carcinoma of the stomach using stomach fibroblasts derived from gastric cancer patients. AB - BACKGROUND/AIMS: The mechanisms of the particular stromal changes that occur upon cancer invasion by scirrhous carcinoma of the stomach, in particular, the relationships among cancer cells, stomach fibroblasts and collagen, a major constituent of the stroma of the invasive tumor, have yet to be clarified. METHODOLOGY: Three different human fibroblast cell lines (TIG-101, MF-2, MKF-1) and a cancer cell line derived from scirrhous carcinoma of the stomach (KATO III) were cultured three-dimensionally in collagen gels to investigate collagen gel contraction by these cells as a model of scirrhous carcinoma of the stomach. RESULTS: The gels contracted and gradually decreased in size in all of the fibroblast (TIG-101, MF-2 and MKF-1) cultures, but not in the KATO III culture, and the extent of gel contraction was not uniform among the fibroblast cell lines. The extent of gel contraction when fibroblasts derived from stomach (MF-2, MKF-1) were co-cultured with KATO III cells in collagen gel was almost similar to that of fibroblasts alone. Moreover, microscopic examination following Masson's trichrome staining revealed condensation and remodeling of collagen fibrils only around the fibroblast cells. CONCLUSIONS: The extent of collagen gel contraction by fibroblasts may depend on their in vivo origin. This property appears to be characteristic of fibroblasts, but not of malignant epithelial cells, under this culture system. Furthermore, the results of the present study demonstrate that stomach fibroblasts may play an important role in the stromal changes associated with scirrhous gastric cancer. PMID- 11100327 TI - Laparoscopic subsegmentectomy for hepatocellular carcinoma with cirrhosis. AB - BACKGROUND/AIMS: Laparoscopic liver resection is feasible for both benign and malignant disease with today's laparoscopic techniques and technology. Location of the tumor at the edge of segment 3, 4, 5, or 6 of our patients makes them an ideal candidate for laparoscopic resection. METHODOLOGY: There were 9 patients who underwent laparoscopic subsegmentectomy for hepatocellular carcinoma with cirrhosis. They were classified as Child A in 6 and B in 3 patients. Hepatitis B was found in 5 and Hepatitis C in 4 cases. Preoperative diagnosis of hepatocellular carcinoma was completed in 7 and definitive histologic diagnosis from frozen section in 2 cases. All 9 patients underwent subsegmentectomy and removal of the tumor with non-tumor cirrhotic liver with a distance of 10 mm at the least margin. Laparoscopic ultrasound allows exact localization of lesions and achievement of adequate resection margin. RESULTS: Those patients resumed a full diet on the 2nd-3rd day after the operation and were discharged home on day 4-7 with no complications but one had prolonging discharge due to ascitis from a drainage tube. Finally, the ascitis was controlled by medications for 1 week. All patients had high postoperative satisfaction. CONCLUSIONS: Laparoscopic liver resection is a procedure of significant risk and technically demanding. Therefore, it should be performed only by experienced liver surgeons with a high level of laparoscopic skill and in the carefully selected patient. PMID- 11100328 TI - Right hepatic artery indwelling catheter for adjuvant chemotherapy after right hepatectomy. AB - Intraarterial hepatic adjuvant chemotherapy after radical hepatic resection for liver metastases from colorectal carcinoma lowers the rate of liver disease relapse. The technique for catheter implantation in the right hepatic artery for subsequent intraarterial hepatic adjuvant chemotherapy after right hepatectomy is herein described and recommended as an effective alternative approach to the standard catheter implantation in the gastroduodenal artery in cases of hepatectomies for liver metastases from colorectal cancer. PMID- 11100329 TI - Double cystic duct detected by endoscopic retrograde cholangiopancreatography and confirmed by intraoperative cholangiography in laparoscopic cholecystectomy: a case report. AB - A case of double cystic duct with cholecystolithiasis detected by preoperative endoscopic retrograde cholangiopancreatography and confirmed by intraoperative cholangiography which was treated successfully by laparoscopic surgery is reported. The patient was a 74-year-old woman who complained of abdominal pain in the right upper quadrant. On admission, ultrasonography revealed hyperechoic areas accompanied by obscure acoustic shadows in the gallbladder. Preoperative endoscopic retrograde cholangiopancreatography showed 2 cystic ducts; 1 branched from the common bile duct and the other from the right hepatic duct. After a diagnosis of double cystic ducts, we chose laparoscopic cholecystectomy. Intraoperative cholangiography via 1 of the cystic ducts revealed the presence of the other. We were able to perform laparoscopic cholecystectomy without any complications and the postoperative course was uneventful. This case suggests that preoperative endoscopic retrograde cholangiopancreatography and intraoperative cholangiography is required to avoid complications during laparoscopic cholecystectomy. PMID- 11100330 TI - Liver metastases of endocrine tumour associated with multiple endocrine neoplasia type 1: a sustained response to interferon therapy or a peculiar benign course? AB - The authors describe the case of a 51-year-old male with Zollinger-Ellison syndrome manifested by epigastralgia, nausea, vomiting, hypergastrinemia and multiple endocrine neoplasia type 1. History included a Billroth II procedure for a perforated duodenal ulcer. Multiple metastatic liver lesions were found that were gastrin-negative and chromogranin-positive. Endoscopy revealed a large ulcerated gastro-jejuno-colonic fistula which was surgically repaired. Pre- and postoperative imaging studies, including the highly sensitive somatostatin receptor scintigraphic scan using In-pentetreotide, have consistently failed to disclose other tumors. Recent reports indicate that most Zollinger-Ellison syndrome-associated gastrinomas are small, easily overlooked lesions located in the proximal duodenum rather than in the pancreas as formerly believed. In the present patient therapy with omeprazole and alpha-interferon has produced complete remission of the Zollinger-Ellison syndrome and a stabilization of tumor growth has occurred during the last 7 years, allowing the patient to live a normal life. This peculiar response to therapy is discussed. PMID- 11100331 TI - The effect of intraperitoneal paclitaxel administration on colonic anastomosis. AB - BACKGROUND/AIMS: The effect of intraperitoneal administration of chemotherapeutic agents on colonic anastomosis are still under investigation. In this study the effects of intraperitoneally administered paclitaxel on rat colonic anastomosis was investigated. METHODOLOGY: After colonic anastomosis, 3 mL of isotonic saline was administered intraperitoneally to rats in control group (Group 1, n = 20). The study group (Group 2, n = 20), paclitaxel 3 mg/kg diluted with isotonic saline was administered intraperitoneally after colonic anastomosis. Rats were sacrificed on 14th day and mean body weight, mean anastomosis bursting pressure and the histopathology of the anastomosis site of the two groups were compared. RESULTS: Mean body weight was approximately the same with the preoperative values at 14th day in both groups. Anastomosis bursting pressure in paclitaxel group (127 +/- 3 mm Hg) was found to be similar to control group (133 +/- 5 mm Hg) (P > 0.05). Mucosal layer formation in the anastomosis line was complete on the 14th day in both groups. CONCLUSIONS: As a result intraperitoneal paclitaxel administration was found to be safe as it did not reduce the anastomotic strength and not delay wound healing. PMID- 11100332 TI - Association of diverticulosis coli and vascular ectasias and the results of fecal occult blood test. AB - BACKGROUND/AIMS: This study was conducted to clarify the diagnostic value of an immunochemical fecal occult blood test for diverticulosis coli and vascular ectasias of the colon, and to assess the association of these diseases to the results of fecal occult blood test. METHODOLOGY: An immunochemical fecal occult blood test over 2 consecutive days was carried out on 72 patients with diverticulosis coli, on 36 patients with vascular ectasias of the colon, on 36 patients with colon cancer, and on 144 healthy subjects. RESULTS: The test was positive in 13.8% patients with diverticulosis coli, in 11.1% patients with vascular ectasias, in 83.3% patients with colon cancer, and in 6.2% healthy subjects, respectively, showing a significant difference in the detection rate between colonic diverticulosis and colon cancer, between vascular ectasias and colon cancer (P < 0.001). However, there was no significant difference in the detection rate between diverticulosis coli and healthy subjects, and between vascular ectasias and healthy subjects. In addition, there was no significant association between the degrees of diverticulosis coli and vascular ectasias to the results of immunochemical fecal occult blood test. CONCLUSIONS: These findings indicate that the immunochemical fecal occult blood is unsuitable for the diagnosis of the patients with colonic diverticulosis and patients with vascular ectasias, and these disorders have little influence on the results of fecal occult blood test. PMID- 11100333 TI - Management of sigmoid colon volvulus. AB - BACKGROUND/AIMS: Sigmoid colon is the most frequent site for a volvulus. The condition has been a formidable one, fraught with innumerable complications responsible for many deaths. In this report, we reviewed our experience with sigmoid colon volvulus. METHODOLOGY: We present our experience of 61 cases of sigmoid volvulus admitted to our department. Twenty-four patients were subjected to non-operative decompression and the others underwent emergency operation. RESULTS: Intestinal volvulus has quite a high morbidity and mortality. Mortality rate of elective resection following sigmoidoscopy was 7.6%. Mortality rate for emergency surgical detortion, primary resection and Hartman procedure were respectively 13%, 16.6% and 37.5%. Important factors such as the patient's features and frequent late diagnosis can influence the complicated outcome of the disease. Plain X-ray of the abdomen is helpful. CONCLUSIONS: Management with the conservative method of treatment in the form of detortion by sigmoidoscopy and rectal tube application is initially effective in most cases of volvulus of the sigmoid colon. On the other hand, elective or emergency sigmoid resection is the most effective treatment for the disease. PMID- 11100334 TI - Experimental study of sutureless colorectal anastomosis. AB - BACKGROUND/AIMS: The present research project has been made mainly with the idea of comparing the tensile strength values and histological answers of three types of colon anastomosis: sutured with silk 5/0; polyglycolic acid 5/0; and sutureless anastomosis with human fibrin gum. METHODOLOGY: One hundred and five (105) Wistar breath rats allocated into 3 groups of 35 animals were used to implement this experimental research project: silk, polyglycolic acid and human fibrin gum. Furthermore, each group was subdivided in 5 series respectively to carry out an experimental study on the tensile strength parameter and anatomic pathological determinations on the 10th, 20th, 30th, 40th and 50th day after the surgical intervention. The following surgical interventions were practiced on them: A cross section of the colon, followed by: group 1: an end-to-end discontinuous suture anastomosis with Silk; group 2: an end-to-end discontinuous suture anastomosis with polyglycolic acid; group 3: sutureless anastomosis with human fibrin gum. On the 10th, 20th, 30th, 40th and 50th days we proceeded to measure the anastomosis' tensile strength value for each series. We used a tensile strength apparatus and waited until the break down of the suture sample took place and wrote down the value, in g/cm, given by the voltmeter at that moment. RESULTS: The results obtained indicate that anastomosis made in group 1 (silk) lasted longer to the tensile strength apparatus; followed by those practiced in group 2 (polyglycolic acid); and finally anastomosis carried out in group 3 (human fibrin gum). However in the anastomotic process carried out with the human fibrin gum the healing started from the 10th day. In the same period of time we carried out the following anatomic-pathological determinations: a) sharp inflammation; b) edema; c) non-specific chronic inflammatory infiltrate; d) granulomatous inflammatory infiltrate to foreign bodies; e) fibrosis. CONCLUSIONS: The results show a better answer for anastomosis made with human fibrin gum than those carried out with the two other suture materials. This conclusion is based on the facts that the human fibrin gum used to carry out sutureless anastomosis during this research project generated a lower sharp inflammation and speediness in its absorption; absence of granular reaction to a foreign body; a minor or non-existent edema at all; as well as a good fibrous healing speediness process. Therefore, all these experimental results lead us to conclude that the human fibrin gum used to carry out sutureless anastomosis may be an alternative to the handmade conventional anastomosis. Moreover they are easy to be implemented. PMID- 11100335 TI - New concepts of molecular biology for colon carcinogenesis. AB - The progressive accumulation of genetic changes in both oncogenes and tumor suppressor genes parallels the clinical and histopathologic progression from normal colonic epithelium through benign adenomas to frank colon cancer. A similar progression is postulated in the transition of normal squamous epithelium to metaplastic mucosa (Barrett's esophagus) and subsequently through dysplasia to adenocarcinoma of the esophagus. A common link between colorectal cancer and Barrett's esophagus or esophageal carcinoma might be explained by either genetic predisposition or common environmental risk factors. The multistep nature of oncogenesis is most directly illustrated by molecular experimental genetic studies which demonstrate that the progression from adenoma to colon carcinoma results from the accumulation of molecular genetic alterations involving mainly 3 factors: activation of oncogenes; inactivation of tumor-suppressor genes; and abnormalities in genes involved in DNA mismatch repair. Changes in oncogenes encoding four distinct groups of proteins (peptide growth factors, protein kinases, signal transducing proteins, and nuclear transcriptional regulatory proteins) can contribute to colon carcinogenesis. In addition, various carcinogens may act at different stages of this model, affecting somatic mutations and resulting in additional genetic alterations. Other promoters, including hormones, may enhance the likelihood of these events through the stimulation of the rate of cell turnover. Diseased detoxification processes may also play a role in carcinogenesis. PMID- 11100336 TI - Benign duodenal tumors. AB - BACKGROUND/AIMS: Benign duodenal tumors are rare and less common than malignant tumors. They comprise a wide variety of pathologies. Treatment is by endoscopic excision or surgical resection. In this report, we aim to review the management of benign tumors located in the proximal duodenum. METHODOLOGY: A retrospective review of 11 patients with benign duodenal tumors treated in a single institution was performed over 10 years. Malignant tumors and periampullary tumors were excluded from the study. RESULTS: The most common presentations were abdominal pain and upper gastrointestinal bleeding. Diagnosis was established by gastroduodenoscopy with biopsy. Seven tumors were located in the first part of the duodenum. The mean size of the tumors was 2.8 cm. Three patients with bleeding tumors were treated with endoscopic hemostasis and underwent surgery because the tumors were larger than 2 cm. Four patients had endoscopic polypectomies and 5 patients had surgical excision. The histological types included 6 adenomas, 3 Brunner's gland adenomas or harmatoma, 1 schwannoma and 1 leiomyoma. The results were good with only 1 case of recurrence. CONCLUSIONS: The presentation of benign duodenal tumors is non-specific. They are diagnosed by gastroduodenoscopy and the tumors can be removed if small and pedunculated. Endoscopic ultrasound is useful in detecting submucosal involvement of sessile tumors. In such cases and large tumors (> 2 cm), surgical excision by laparotomy or laparoscopy should be undertaken. PMID- 11100337 TI - Omeprazole and regulation of cytokine profile in Helicobacter pylori-infected patients with duodenal ulcer disease. AB - BACKGROUND/AIMS: To estimate the cytokine profile in Helicobacter pylori-positive patients with duodenal ulcers treated with omeprazole. METHODOLOGY: The subjects were 22 patients with endoscopically defined active duodenal ulcers and H. pylori infection treated with omeprazole for 3 months after initial 1-week triple therapy. Peripheral blood CD3+ CD4+ and CD8+ T lymphocytes, percentage of HLA-DR+ peripheral blood lymphocytes, and cytokine levels (GM-CSF, IL-6 and IL-2) were measured in the supernatants from PHA-cultured peripheral blood lymphocytes at baseline conditions and after 3-months' omeprazole treatment. RESULTS: The H. pylori eradication triple regimen cured H. pylori infection in 86.4% of our patients. Peripheral blood CD3+ CD4+ and CD8+ T lymphocytes were not affected by omeprazole treatment (P > 0.05), whereas the percentage of HLA-DR+ peripheral blood lymphocytes increased significantly post-treatment (P < 0.05). All cytokine levels measured in the supernatants showed a significant decrease after treatment (P < 0.001). CONCLUSIONS: The significant decrease in the two pro-inflammatory cytokines (GM-CSF, IL-6) and the major inflammatory cytokine IL-2, implies that the potential cytokine synthesis of the Th1 pattern from peripheral blood lymphocytes is highly suppressed after treatment. This effect on cytokine regulation seems to be a result of the positive H. pylori infection outcome at the immunological level, which leads to "normalization" of cytokine production. PMID- 11100338 TI - Omentoplasty for cervical esophagogastrostomy following radical esophagectomy with three-field dissection. AB - BACKGROUND/AIMS: Anastomotic leakage is the main cause of postoperative mortality and incidence of which, following three-field lymph node dissection, is around 30%. The study was undertaken to investigate the role of omentoplasty to reinforce cervical esophagogastrostomy with the expectation of lowering the rate of anastomotic leakage after radical esophagectomy with three-field lymph node dissection. METHODOLOGY: Between July 1995 and Dec 1997, a total of 32 patients underwent total thoracic esophagectomy with three-field lymph node dissection and cervical esophagogastrostomy. Eleven patients were stage IIA, 3 stage IIB, 5 stage III and 13 stage IV. After radical esophagectomy and lymph node dissection, several omental branches of the gastroepiploic vessels remained to supply a gastric tube. An end-to-side cervical esophagogastrostomy was performed on the posterior wall of the gastric tube using a circular stapler. The omentoplasty- wrapping the esophagogastrostomy--was performed. A retrosternal route for reconstruction was used in 23 patients and a posterior mediastinal route in 9 patients. RESULTS: Esophageal anastomotic leakage occurred in only 1 patient, 3.1% overall. There was neither pyothorax nor mediastinitis. There was no lethal anastomotic leakage. Later, 2 patients (6.2%) developed an anastomotic stricture that required balloon dilatation. CONCLUSIONS: Omentoplasty to reinforce cervical esophagogastrostomy decreases anastomotic failure following radical esophagectomy with three-field lymph node dissection. PMID- 11100339 TI - Octreotide in acute bleeding esophageal varices: a prospective randomized study. AB - BACKGROUND/AIMS: To assess the value of octreotide in the control of acute bleeding esophageal varices, in a prospective randomized study. METHODOLOGY: One hundred and ninety-seven patients admitted for variceal bleeding confirmed at endoscopy were recruited and divided into two groups: group I (n = 111) with endoscopic stigmata of recent bleeding; and group II (n = 86) with active bleeding at emergency endoscopy. Patients in group I were randomized to receive a continuous infusion of octreotide (n = 58) or emergency sclerotherapy (n = 53). Patients in group II were assigned to sclerotherapy (n = 42) or to sclerotherapy plus octreotide (n = 44). At the end of the period of study (48 hours), patients were submitted to sclerotherapy or band ligation until variceal obliteration was achieved. RESULTS: In group I, octreotide was found to be as effective as sclerotherapy regarding hemostasis at 48 hours and on day 7 after the index bleeding episode. Transfusion needs were not significantly different for the two treatment modalities. In group II, the association of octreotide with sclerotherapy was significantly better than sclerotherapy alone either in controlling acute active bleeding (P < 0.001) or in achieving hemostasis at 48 hours (P < 0.01). Transfusion needs were significantly fewer in patients treated with this therapeutic association as compared to sclerotherapy alone. CONCLUSIONS: These results suggest that octreotide infusion is effective in the treatment of variceal bleeding. In patients with recent bleeding, octreotide infusion is as effective as emergency sclerotherapy. In active variceal bleeding, it is a valuable adjuvant treatment in association with emergency sclerotherapy. PMID- 11100340 TI - Predicting initial recurrence pattern of esophageal cancer after neoadjuvant chemotherapy. AB - BACKGROUND/AIMS: No report has reviewed which clinicopathological factors including 3-field dissection and the response to neoadjuvant chemotherapy can predict the recurrence pattern of an esophageal carcinoma. The aim of this study was to reveal clinicopathological predictors for the initial recurrence pattern of a thoracic esophageal carcinoma. METHODOLOGY: Sixteen parameters derived from 98 patients who underwent a curative esophagectomy with neoadjuvant chemotherapy for a squamous cell carcinoma of the thoracic esophagus were examined using univariate and multivariate logistic regression analyses. RESULTS: Thirty-seven (37.8%) of the 98 patients had recurrences (hematogenous; 16, lymphatic; 13, others; 8). Univariate analyses revealed that the completion of 3-field dissection was the only factor for suppressing the lymphatic recurrence (P = 0.009; odds ratio: 0.2). Multivariate analyses showed that the number of positive nodes was a significant predictor for recurrence including all modalities (P = 0.02; odds ratio: 1.2) and both the number of positive nodes (P = 0.04; odds ratio: 1.1) and the poor response to neoadjuvant chemotherapy (P = 0.02; odds ratio: 6.9) were significant predictors for the hematogenous recurrence. CONCLUSIONS: The number of positive nodes and the response to neoadjuvant chemotherapy could predict the hematogenous recurrence of esophageal carcinoma. PMID- 11100341 TI - A rare complication of endoscopic injection sclerotherapy: thrombosis of subclavian vein. AB - A 28-year-old man with compensated cirrhosis of the liver (Child B) and after 4 episodes of esophageal variceal bleeding received prophylactic endoscopic variceal sclerotherapy in our Gastroenterology Clinic for 8 consecutive months. Sclerotherapy of the esophageal varices had been performed at monthly intervals until variceal obliteration was achieved. Both the intravariceal and paravariceal injection techniques were used and injections were repeated periodically as necessary. On the 8th month, 1 week after the 4th sclerotherapy procedure, the patient complained of swelling on his right shoulder and on his right arm. There was jugular congestion and swelling of his right arm and right shoulder. The patient was hemodynamically stable. An X-ray of brachial venography revealed an obstruction of the vena subclavia dextra. During follow-up, the jugular congestion and swelling of his right arm gradually subsided spontaneously over a 6-month period without any need for medication. There has been no recurrence of his symptoms during the 1-year follow-up period. Now, he is still well clinically. This experience suggests that endoscopic injection sclerotherapy may cause thrombosis of the subclavian vein which have been never seen before. PMID- 11100342 TI - Peritoneovenous shunt in malignant ascites. The Bordet Institute experience from 1975-1998. AB - BACKGROUND/AIMS: This is the review of our experience in the treatment of malignant ascites using Le Veen and Denver peritoneovenous shunt. METHODOLOGY: From 1975-1998, 24 peritoneovenous shunts were inserted in 22 patients with malignant ascites. RESULTS: All patients benefited from the procedure. The principal cause of failure was shunt occlusion (n = 5). CONCLUSIONS: This study permits us to conclude that a preoperative appropriated selection of patients, should decrease failure rate. Patients should be sent to surgery as soon as the diagnosis of recalcitrant ascites is confirmed, to obtain a longer and more comfortable quality of life with terminal cancer. PMID- 11100343 TI - Somatostatin and ranitidine in the treatment of non-variceal upper gastrointestinal bleeding: a prospective, randomized, double-blind, controlled study. AB - BACKGROUND/AIMS: The aim of this study was to compare the efficacy of somatostatin vs. ranitidine in controlling acute non-variceal gastrointestinal bleeding. METHODOLOGY: A total of 48 patients with acute upper gastrointestinal bleeding due to duodenal or gastric ulcer were divided into 2 groups. Group I consisted of 15 patients with Forrest IB and Group II consisted of 30 patients with Forrest II. Two regimens were randomly allocated to all patients within half an hour after the endoscopic procedure: 1) somatostatin-UCB 250 mcg i.v. bolus followed by continuous i.v. infusion at a rate of 6 mg/d for 72 h, or 2) ranitidine 300 mg/d by continuous i.v. infusion for 72 h. RESULTS: In Group I, although mean blood transfusion requirements (no. of units) were lower in patients treated with somatostatin than in those treated with ranitidine, this was not statistically significant (mean +/- SD: 2.56 +/- 3.05 vs. 5.17 +/- 4.96, respectively; P > 0.05); the time of bleeding stop was shorter in the somatostatin group than in the ranitidine group (mean +/- SD: 3.24 +/- 2.45 vs. 11.25 +/- 11.63, respectively; P = 0.0383). The rebleeding and the mortality rates did not differ between the treatment groups in both Group I and Group II. CONCLUSIONS: Somatostatin is more effective than ranitidine in controlling acute non-variceal gastrointestinal bleeding in patients with Forrest IB bleeding activity. Somatostatin has no additional benefit in those with Forrest II bleeding activity. PMID- 11100344 TI - Felodipine does not increase the reflux episodes in patients with gastroesophageal reflux disease. AB - BACKGROUND/AIMS: Calcium channel blocking agents have been reported to increase the risk of gastroesophageal reflux. However, whether felodipine, a newer calcium channel blocker, increases reflux episodes and decreases esophageal acid clearance in patients with gastroesophageal reflux disease has never been studied. Therefore, the aim of this study was to evaluate whether felodipine increases the incidence of gastroesophageal reflux in patients with gastroesophageal reflux disorder. METHODOLOGY: Nine patients with gastroesophageal reflux disease, 6 men and 3 women, with a mean age of 62.6 +/- 14.4 years (range: 37-80 years) were studied. They received ambulatory esophageal pH monitoring for 45.7-48 hours (mean: 47.1 +/- 0.8 hours). Various pH parameters were evaluated during a similar interval of monitoring time before and after receiving 5 mg of felodipine. RESULTS: No significant difference was noted in any pH parameter by the Wilcoxon signed Ranks test, including reflux episodes (P = 0.552), reflux episodes longer than 5 min (P = 0.683), esophageal acid clearance (P = 0.663) and fraction time of pH < 4 (P = 0.752) before and after the use of felodipine. CONCLUSIONS: Felodipine does not increase reflux episodes or impair esophageal acid clearance in patients with gastroesophageal reflux disease. PMID- 11100345 TI - Endoscopic injection therapy vs. multipolar electrocoagulation vs. laser vs. injection + octreotide vs. injection + omeprazole in the treatment of bleeding peptic ulcers. A prospective randomized study. AB - BACKGROUND/AIMS: A prospective randomized study was performed to assess the effectiveness and safety of 5 different methods of hemostasis in selected patients with high-risk bleeding peptic ulcers. METHODOLOGY: Two hundred and eight patients (n = 208; mean age: 61.6 yrs) with endoscopic stigmata of active hemorrhage, non-bleeding vessel or adherent fresh clot were randomized during emergency endoscopy to receive one of the following modalities of endoscopic therapy (with or without pharmacological therapy): I) injection of absolute alcohol (n = 44); II) multipolar electrocoagulation (BICAP; n = 42); III) Nd-YAG laser (n = 40); IV) injection of absolute ethanol + octreotide (n = 42); V) injection of absolute ethanol + omeprazole (n = 40). RESULTS: The 5 treatment groups were clinically and endoscopically comparable. The initial hemostatic success was > 90% in every group. No significant differences between groups were found in any of the following parameters assessed during hospitalization: incidence of rebleeding (I = 14.8% vs. II = 19.0% vs. III = 16.6% vs. IV = 18.1% vs. V = 20.0%; P > 0.05 mean = 17.7%); incidence of definitive hemostasis (I = 89.3% vs. II = 85.7% vs. III = 86.6% vs. IV = 84.0% vs. V = 86.6%; P > 0.05; mean = 86.5%); incidence of emergency surgery (I = 8.5% vs. II = 11.9% vs. III = 10.0% vs. IV = 6.8% vs. V = 11.1%; P > 0.05; mean = 9.6%); mortality rate (I = 4.2% vs. II = 4.7% vs. III = 3.3% vs. IV = 13.6% vs. V = 4.4%; P > 0.05; mean = 6.2%). Mean age of deceased patients was significantly higher than living patients (71.2 +/- 13.4 vs. 60.9 +/- 14.4; P < 0.05). Approximately 2/3 of the fatal cases were strongly weakened by coexistent medical diseases. The duration of hospital stay was similar for all groups. The BICAP group required less units of blood transfusion (1.9 +/- 1.8 vs. I = 3.0 +/- 2.6; III = 3.5 +/- 3.6; IV = 2.8 +/- 2.3; V = 3.1 +/- 2.5; P < 0.05), perhaps due to the higher mean value of hemoglobin of these patients at hospital admission, compared to all other groups. No significant complications were reported. CONCLUSIONS: This study provides good evidence that injection of absolute ethanol, multipolar electrocoagulation (BICAP) and Nd-YAG laser are equally safe and effective in the endoscopic therapy of acute bleeding peptic ulcers. In contrast, no additional hemostatic benefits arose from the association of pharmacological agents (octreotide or omeprazole) to sclerosis injection. PMID- 11100346 TI - Surgery-induced decline in circulating dendritic cells in operable cancer patients: a possible explanation of postoperative immunosuppression. AB - BACKGROUND/AIMS: The recent advances in the immunobiology of tumor have demonstrated the essential role of dendritic cells in anticancer immunity. Dendritic cells activate anticancer immunity by secreting interleukin-12 and by activating T helper lymphocytes, with the following production of interleukin-2. Since surgery-induced immunosuppression has been proven to be associated with a decline in the blood levels of both interleukin-2 and interleukin-12, it could depend at least in part on a transient deficiency of dendritic cells system. Unfortunately, at present there are no data about changes in circulating dendritic cell number during the postoperative period. This preliminary study was performed to evaluate the influence of surgery on dendritic cell number in the peripheral blood. METHODOLOGY: The study included 14 consecutive operable gastrointestinal tract cancer patients, who were evaluated before and at day 7 of the postoperative period. The control group consisted of 50 healthy subjects. Immature (CD 123+) and mature (CD 11+) dendritic cell subsets were measured by FACS and monoclonal antibodies. RESULTS: Cancer patients showed a significantly lower mean number of immature dendritic cells with respect to that found in controls. The mean number of mature dendritic cells was also lower in patients than in controls, without, however, significant differences. Finally, surgery induced a statistically significant decline in the mean number of both immature and mature dendritic cells, and the decrease was particularly pronounced for immature dendritic cells. CONCLUSIONS: In addition to the well-demonstrated surgery-induced lymphocytopenia, this preliminary study shows that the surgical treatment may determine a significant decrease in circulating immature and mature dendritic cells. Because of the fundamental role of dendritic cells in regulating the immune responses, surgery-induced decline in circulating dendritic cells number could play a role in determining the immunosuppressive status, which characterizes the postoperative period. PMID- 11100347 TI - Prevalence of hepatitis G virus (HGV) infection in an endemic area of hepatitis C virus (HCV) infection. AB - BACKGROUND/AIMS: We investigated the prevalence of hepatitis G virus infection among inhabitants of a hepatitis C virus endemic area. METHODOLOGY: Two hundred and eighty-eight inhabitants, who underwent medical examinations for health screening, were enrolled in this epidemiological study. HGV RNA and HCV RNA were detected by polymerase chain reaction. We also examined anti-HGV envelope protein (E2) antibodies in all serum samples. RESULTS: In these 288 inhabitants, we found anti-HCV antibodies (HCV-Ab) and HCV RNA in 28.5% and 17.4%, respectively. HGV RNA and anti-HGV E2 were detected in 9 (3.1%) and 16 (5.5%), respectively. One patient was positive for both HGV RNA and anti-HGV E2. The exposure rate, expressed as the percentage of people with HGV RNA and/or anti-HGV E2, was 8.3%, which was significantly lower than the incidence of positive HCV-Ab. Of the 24 patients with HGV RNA and/or anti-HGV E2, 15 (62.5%) were positive for HCV-Ab, of those HCV RNA was detected in 9 (37.5%). Further, we found a higher prevalence of HGV exposure in patients with HCV-Ab than in those without (8.3% vs. 4.4%). CONCLUSIONS: HGV infection was not identical to the epidemic hepatitis C virus infection among inhabitants of this town, suggesting that hepatitis C virus might be less infectious than hepatitis C virus. PMID- 11100348 TI - Intensive care during prolonged anhepatic state after total hepatectomy and porto caval shunt (two-stage procedure) in surgical complications of liver transplantation. AB - BACKGROUND/AIMS: Recently, total hepatectomy and temporary porto-caval shunt has been indicated in surgical complications of liver transplantation. Four cases of liver transplantation which presented liver hemorrhage at the time of implant, and a 5th case with surgical trauma of hepatic hilum are presented. METHODOLOGY: The graft was removed and a porto-caval shunt was performed in all patients. Retransplantation was possible in all recipients, after an anhepatic period of 16 24 hours. RESULTS: Early persistent ionic hypocalcemia and late olyguric renal failure were the most constant and prominent complications during the anhepatic period. Two patients died of renal failure and respiratory distress syndrome at 6 and 28 days, respectively, after liver transplantation. The other 3 patients are alive and without complications at 48, 33 and 11 months of follow-up. CONCLUSIONS: Total hepatectomy with a temporary porto-caval shunt and later retransplantation must be considered as a useful procedure for surgical complications of liver transplantation which may not be treated using other techniques. Special attention should be paid to preserve renal function in the anhepatic state in order to improve survival in similar cases of two-stage liver transplantation. PMID- 11100349 TI - Early use of cyclic TPN prevents further deterioration of liver functions for the TPN patients with impaired liver function. AB - BACKGROUND/AIMS: Impaired liver function is frequently found in patients who need prolonged total parenteral nutrition. Cyclic total parenteral nutrition can minimize the adverse effects of long-term total parenteral nutrition, such as hepatic complication. The adequate timing to shift to use cyclic total parenteral nutrition for patients with impaired liver function may prevent further hepatic dysfunction. METHODOLOGY: A prospective study of 65 patients who need total parenteral nutrition and have impaired liver functions was performed. Cyclic total parenteral nutrition was used in different groups of patients, when their total bilirubin levels were just over 5 mg%, 10 mg%, or 20 mg% during the course of total parenteral nutrition. The patients of control groups received straight non-cyclic total parenteral nutrition. All the patients had stable vital signs without major stress, such as sepsis or acute bleeding. Ten patients (A2) in Group A were shifted to cyclic total parenteral nutrition when their total bilirubin was just over 5 mg%; the other 10 patients (A1) continued the non cyclic total parenteral nutrition. Eleven patients (B2) in Group B were shifted to cyclic total parenteral nutrition when their total bilirubin was just over 10 mg%; the other 11 patients (B1) continued the non-cyclic total parenteral nutrition. Ten patients (C2) in Group C were shifted to cyclic total parenteral nutrition when their total bilirubin was just over 20 mg%; the other 13 patients (C1) continued the non-cyclic total parenteral nutrition. The average energy intake among 3 groups had no difference. Their liver functions were examined each week for 2 weeks. RESULTS: The results showed that the patients with non-cyclic total parenteral nutrition had significant increase of direct-total bilirubin and alkaline phosphatase (P < 0.05) in Group A and significant decrease of albumin accompanied with increase of GOT, GPT, direct/total bilirubin (P < 0.05) in Group B. The patients either using cyclic or non-cyclic total parenteral nutrition showed significant decrease of albumin and increase of direct/total bilirubin (P < 0.05) in Group C. CONCLUSIONS: We conclude that the early use of cyclic total parenteral nutrition may prevent deterioration of liver function for the patients with jaundice and need prolonged total parenteral nutrition. PMID- 11100350 TI - Twelve-month interferon-alpha therapy induces a similar sustained response in anti-HBe-positive and HBeAg-positive chronic hepatitis B patients. AB - BACKGROUND/AIMS: We compared the response to interferon-alpha 2a in 35 patients with antibody to HBeAg (anti-HBe) and 20 patients with HBeAg in serum, and histological features of chronic hepatitis B. METHODOLOGY: Patients were treated with 4.5-6 MU of interferon-alpha 2a, three times a week for 12 months, and followed for 30.8 +/- 13.5 additional months. RESULTS: All of them had elevated serum levels of aminotransferases and positive test for hepatitis B virus-DNA in serum. Patients with anti-HBe-positive chronic hepatitis were older and had higher serum aminotransferase levels than HBeAg-positive patients, but no differences were seen between both groups with respect to sex, history of acute hepatitis, mode of transmission of the infection or histological appearance before interferon therapy. Serum levels of alanine transaminase became normal and hepatitis B virus-DNA undetectable by PCR at the end of therapy in 25 (71%) of anti-HBe-positive patients and in 10 (50%) of HBeAg-positive patients (P > 0.05). Although 10 (29%) of the anti-HBe-positive and none of the HBeAg-positive patients relapsed, no significant difference was seen in the rate of sustained response (43% vs. 50%, respectively). The histological improvement was similar in both groups. CONCLUSIONS: The results of this study indicated that biochemical, virological and histological response to 12-month interferon-alpha 2a therapy was similar in patients with anti-HBe antibody than in patients with the classical HBeAg-positive of chronic hepatitis B. PMID- 11100351 TI - Diffuse intrahepatic recurrence after resection of hepatocellular carcinoma. AB - BACKGROUND/AIMS: An early diffuse type in the pattern of the postoperative intrahepatic recurrence of hepatocellular carcinoma has been recognized. The purpose of this study was to elucidate risk factors for diffuse recurrence of hepatocellular carcinoma. METHODOLOGY: The subjects involved in the present study were 114 patients with hepatocellular carcinomas resected in Tenri Hospital during the past 12 years. Univariate analysis was used for retrospective determination of the factors related to diffuse recurrences after surgery in 10 cases among 114 patients. RESULTS: The risk factors linked to diffuse recurrence were microscopical portal infiltration (P < 0.01), elevated alpha-fetoprotein (more than 1000 ng/mL) (P < 0.05), the absence of preoperative transcatheter arterial embolization (P < 0.01), and two or more segmentectomies of the liver (P < 0.01). Six of 10 patients with microscopical portal infiltration and elevated alpha-fetoprotein (more than 1000 ng/mL) had diffuse recurrence (P < 0.01). Six of 8 patients with two or more segmentectomies without preoperative TAE had diffuse recurrence (P < 0.01). CONCLUSIONS: When patients with the diagnosis of operable hepatocellular carcinoma have portal infiltration and elevated alpha fetoprotein (more than 1000 ng/mL), two or more segmentectomies of the liver without preoperative transcatheter arterial embolization should be avoided. PMID- 11100352 TI - The value of ascitic fluid polymorphonuclear cell count determination during therapy of spontaneous bacterial peritonitis in patients with liver cirrhosis. AB - BACKGROUND/AIMS: Spontaneous bacterial peritonitis is one of the most common complications attending the onset of ascites in patients with liver cirrhosis. The aim of this study was to demonstrate whether it is possible, on the basis of ascitic fluid polymorphonuclear cell count in patients with liver cirrhosis and spontaneous bacterial peritonitis, to determine the optimal duration of cefotaxime therapy, as the most frequently applied empirical therapy, and possibly anticipate the disease recurrence. METHODOLOGY: In 16 patients with alcoholic liver cirrhosis and confirmed diagnosis of spontaneous bacterial peritonitis, cefotaxime therapy was administered 2g t.i.d. during 5 days. Before the therapy, at 48 hours, 5 days and 15-20 days after the cefotaxime therapy was started, in all patients with spontaneous bacterial peritonitis diagnostic abdominal paracentesis was performed, each time determining the ascitic fluid polymorphonuclear cell count together with microbiological analysis. RESULTS: In the course of the "primary" spontaneous bacterial peritonitis attack, 3 patients died (18.8%). In 4 patients the recurrence of spontaneous bacterial peritonitis was observed within 15-20 days after therapy was discontinued. Two patients died during the therapy of spontaneous bacterial peritonitis recurrence. After 48 hours of therapy, 11 patients with the "primary" spontaneous bacterial peritonitis attack were without any symptoms (68.8%). Out of these 11, 10 patients (62.5%) had the ascitic fluid polymorphonuclear cell count lower than 250/mm3. After 5 days of therapy, 12 patients (75%) were free of symptoms, and the number of ascitic fluid polymorphonuclear cell count < 250/mm3 was still found in 10 (62.5%) patients. No association between the presence of symptoms 48 hours after the therapy and the recurrence of spontaneous bacterial peritonitis was established. A significant association was found between the ascitic fluid polymorphonuclear cell count determined 48 hours after the therapy and the recurrence of spontaneous bacterial peritonitis. A recurrence occurred in only 1 patient with the number of ascitic fluid polymorphonuclear cell count < 250/mm3, 48 hours after the therapy was started. A recurrence of spontaneous bacterial peritonitis occurred in all the patients who had an ascitic fluid PMN cell count > or = 250/mm3, 48 hours after the therapy was started. CONCLUSIONS: By monitoring the ascitic fluid PMN cell count it seems to be possible to determine the efficacy and optimal duration of cefotaxime therapy in patients with spontaneous bacterial peritonitis when it is of most importance that the number of ascitic fluid PMN cell count should decrease below 250/mm3 during the therapy. PMID- 11100353 TI - Nitric oxide synthase inhibitor increases hepatic injury with formation of oxidative DNA damage and microcirculatory disturbance in endotoxemic rats. AB - BACKGROUND/AIMS: Nitric oxide plays important roles in the pathogenesis of endotoxin shock and multiple organ failure. Nitric oxide synthase inhibitors are used in patients to improve hemodynamics in endotoxin shock. However, the role of nitric oxide is controversial in hepatic injury with oxidative DNA damage in endotoxemia. This report investigated the role of nitric oxide on hepatic blood flow and liver injury in endotoxemic rats. METHODOLOGY: Under light ether anesthesia, male Wistar rats were given lipopolysaccharide (10 mg/kg) intravenously. Several hours (0-24 hr) later, the animals were used for experiments. In some experiments, NG-[1-iminoethyl]-L-ornithine, a potent inhibitor of nitric oxide synthase, was administered 5 mg/kg intraperitoneally every 3 hour after lipopolysaccharide injection. Hemodynamic changes, biochemical and histological analysis were determined. RESULTS: Lipopolysaccharide increased the activity of inducible nitric oxide synthase in the liver, lungs and spleen. Significant amounts of nitric oxide-hemoglobin complexes and nitrite plus nitrate appearing in the blood peaked at 8 hr after treatment. NG-[1-iminoethyl]-L ornithine completely inhibited the generation of nitric oxide metabolites, but hardly affected formation of urinary 8-hydroxydeoxyguanosine and the systemic blood pressure in normal rats. NG-[1-iminoethyl]-L-ornithine increased 8 hydroxydeoxyguanosine formation and decreased the blood flow more in the superior mesenteric artery and hepatic microvascular blood flow in endotoxemic rats. Inhibition of nitric oxide synthase markedly caused deterioration of the lipopolysaccharide-induced liver injury indicated by hepatic enzymes and histological findings. CONCLUSIONS: These results suggested that suppresion of endogenous nitric oxide might aggravate hepatic injury, partly caused by decrease in hepatic blood flow accompanied with oxidative stress in endotoxemia. PMID- 11100354 TI - Surgical complications and long-term outcome in pediatric liver transplantation. AB - BACKGROUND/AIMS: Liver transplantation has been widely accepted for the treatment of children with end-stage liver disease over the last 10 years particularly with the advent of reduced-size liver transplant technique. This study reviewed the perioperative and long-term results in the pediatric program of the Queensland Liver Transplant Service, Brisbane, Australia. METHODOLOGY: Retrospective analysis was performed in 153 children who received 176 liver grafts between 1985 and 1995, including 109 (62%) reduced-size and 67 (38%) whole liver grafts. Median follow-up period was 5.3 years. RESULTS: One-, 5-, and 10-year patient and graft survival rates were 82% and 74%, 75% and 63%, and 70% and 60%, respectively. Normal physical and intellectual development was observed in 98% of survivors. There were no significant differences in patient or graft survival rates between transplants using reduced-size and whole liver grafts. Portal vein thrombosis was the most common vascular complication, occurring in 8%. Hepatic artery thrombosis occurred in 7%, including 11% of children less than 1 year old and 8% of those under 10 kg. Biliary complication was found in 16% and posttransplant gastrointestinal perforation in 19%. CONCLUSIONS: Liver transplantation has the potential to cure and allow development in children with end-stage liver disease. PMID- 11100355 TI - Diagnostic value of 99mTc-labeled red blood cell SPET for a solitary solid liver mass in HBV carrier patients with different echogenicities. AB - BACKGROUND/AIMS: The purpose of this study is to evaluate the efficacy of technetium-99m labeled red blood cell liver single photon emission computed tomography (RBC liver SPET) in evaluating the diagnostic ability for differentiating the nature of a solitary liver tumor detected with ultrasonography in hepatitis B carrier patients. METHODOLOGY: One hundred and one hepatitis B carrier patients (56 males, 45 females, aged 13-70 years) with a solitary solid liver mass found on ultrasonography were included in this study. The final diagnosis was made after liver biopsy, aspiration with cytology and/or autopsy in 27 patients and after follow-up with both clinical and ultrasonography findings in 74 patients. RESULTS: Hemangioma was found in 79 patients, hepatocellular carcinoma in 14, focal nodular hyperplasia in 5, fatty liver in 2, and metastasis in 1. The diagnostic sensitivity of RBC liver SPET for hemangioma, with a hyperechoic, hypoechoic, or isoechoic ultrasonography pattern, was between 75-80%, while the specificity for all patterns was 100%. For mixed-echoic lesions, the sensitivity was 100%, but the specificity was only 50%. Two false positives were noted; both were mixed-echoic lesions. CONCLUSIONS: RBC liver SPET is useful for differentiating hemangioma from other liver tumors in hepatitis B carrier patients with a various sonographic patterns, especially for those who had a mixed-echoic sonographic liver mass. PMID- 11100356 TI - Multimodal therapy of hepatocarcinoma: personal experience on 90 cases. AB - BACKGROUND/AIMS: In recent years, surgical and non-surgical options have been developed in the treatment of hepatocellular carcinoma in cirrhotic patients. We review our personal series from 1995-1999, in order to assess the choice of treatment. METHODOLOGY: Of 90 cases of hepatocellular carcinoma observed in the years 1995-1999, 15 underwent curative resective surgery; in 42 cases TAE, PEI or RITA were utilized (9 of them as multimodal therapy). In the remaining 33 patients any kind of therapy was scheduled. RESULTS: The mean survival of the 15 resected patients was 18 months, non-statistically better than RITA survival, compared by Log-Rank test. Perioperative mortality calculated in all procedures was 5.2% (2 pts surgery, 1 pt TAE). CONCLUSIONS: The high percentage of not treated hepatocellular carcinomas in our series is generally due to large tumor size diagnosed in advanced Child's stage. PEI, TAE and RITA have to be considered effective and safe for palliation for HCCs. However, surgical resection represents the curative therapy in selected cirrhotic patients affected by HCC. PMID- 11100357 TI - Retrospective analysis of 29 patients surgically treated for hepatocellular adenoma or focal nodular hyperplasia. AB - BACKGROUND/AIMS: Hepatocellular adenoma resection and focal nodular hyperplasia supervision are widely recognized as the best management when these benign liver tumors are diagnosed. Differential diagnosis is thus mandatory. METHODOLOGY: Twenty-nine patients with a presumed benign liver tumor were retrospectively analyzed. RESULTS: Histopathological analysis of these resected liver tumors demonstrated hepatocellular adenoma in 16 patients and focal nodular hyperplasia in 13. One hepatocellular carcinoma was disclosed into a hepatocellular adenoma and 2 hepatocellular adenoma showed foci of liver-cell dysplasia. Seven patients with hepatocellular adenoma (43%) had evidence of intratumoral hemorrhage, among which 3 patients were admitted with intraperitoneal tumoral rupture. Computed tomography, performed in 26 patients, was the most reliable examination to characterize these presumed benign liver tumors. Magnetic resonance imaging concerned only 5 patients but 3 hepatocellular adenoma and 1 focal nodular hyperplasia were diagnosed. The indications of focal nodular hyperplasia surgical resection were chronic pain (4 pts), hepatocellular adenoma diagnosis (4 pts), undeterminate liver mass (2 pts), a liver mass of unknown origin in patients with a neoplastic history (3 pts). A diagnosis of focal nodular hyperplasia assumed by the imaging work-up was always histologically confirmed. All the patients underwent hepatic resection with no mortality. CONCLUSIONS: This report underlines the risk of hemorrhage or malignant transformation of hepatocellular adenoma that justifies a safety surgical resection. An imaging work-up in favor of focal nodular hyperplasia allows radiological observation. PMID- 11100358 TI - Hepatitis G virus infection in intravenous drug users with or without human immunodeficiency virus infection. AB - BACKGROUND/AIMS: To evaluate the HGV infection prevalence in a group of intravenous drug users with or without human immunodeficiency virus coinfection. METHODOLOGY: We studied 57 patients (48 males and 9 females) who were either previous or still ongoing intravenous drug users. Thirty-seven patients were HIV+ve, 55 patients were anti-HCV+ve and 3 patients were HBsAg chronic carriers. Patient sera were tested for HGV-RNA, anti-E2, qualitative and quantitative HCV RNA as well as for HCV genotypes. Moreover, the ALT level was checked in the serum sample of each patient. RESULTS: We found a high prevalence (35/57; 61.4%) of HGV infection in our patients. HGV-RNA was detected in 16 out of the 57 intravenous drug users (28%). In particular HGV-RNA was positive in 12 out of the 37 HIV+ve patients (32.4%) and in 4 out of the 20 HIV-ve patients (20%). Anti-E2 were detected in 19 out of the 57 patients (33.3%) with greater prevalence among HIV-ve subjects (12/20; 60%) compared to HIV+ve group (7/37; 18.9%). This resulting difference was statistically significant (P < 0.05). All HGV RNA+ve/anti-E2+ve patients were anti-HCV/HCV-RNA+ve and none of our patients were anti-E2+ve/HGV-RNA+ve at the same time. Significant differences were not found between HGV-RNA+ve and HGV-RNA-ve patients as far as clinical and virological data are concerned. CONCLUSIONS: The prevalence of HGV infection in intravenous drug users proved to be high especially in the HIV+ve group. Moreover HGV was associated with HCV in all our cases. The actual clinical impact of HGV infection remains unclear since HGV does not seems to influence the biochemical, virological or histological alterations caused by HCV infection. PMID- 11100359 TI - The hemodynamics of dysplastic nodules in the liver evaluated by CT angiography and immunohistochemistry. AB - BACKGROUND/AIMS: Dysplastic nodules diagnosed pathologically exhibit various hemodynamic patterns. To evaluate these differences in their hemodynamics, we observed basement membrane formation of sinusoids. METHODOLOGY: For 12 low-grade dysplastic nodules, 24 high-grade dysplastic nodules and 16 hepatocellular carcinomas, both computed tomography during arterial portography and CT arteriography were performed preoperatively. Resected specimens were examined immunohistochemically for COL IV (type IV collagen) and laminin to observe basement membrane formation. We compared their hemodynamics on computed tomography during arterial portography and CT arteriography with the expressions of COL IV and laminin. RESULTS: All of the low-grade dysplastic nodules were not hypervascular on CT arteriography, and negative for COL IV and laminin. All of the hepatocellular carcinomas were hypervascular on CT arteriography, and positive for COL IV and laminin. High-grade dysplastic nodules exhibit four hemodynamic patterns on CT during Arterial Portography/CT arteriography. All of 7 iso/iso and 5 iso/hypo nodules were negative for COL IV and laminin. Of the 9 hypo/hyper nodules, which exhibited hemodynamics similar to hepatocellular carcinoma, COL IV was identified in 7 and laminin identified in 8 nodules. Of the 3 hypo/hypo nodules, COL IV and laminin were identified in one. CONCLUSIONS: In high-grade dysplastic nodules, most cases exhibited a correlation between the hemodynamic patterns and the degree of the sinusoidal basement formation. PMID- 11100360 TI - Does surveillance for hepatocellular carcinoma in HCV cirrhotic patients improve treatment outcome mainly due to better clinical status at diagnosis? AB - BACKGROUND/AIMS: Cirrhotic patients with hepatitis C virus infection are a group at higher risk for hepatocellular carcinoma. Conventional screening programs detect only few early hepatocellular carcinomas that are eligible for radical treatment. Our aim was to compare characteristics of patients, modality of treatment, and outcome in anti-HCV positive cirrhotics with hepatocellular carcinoma diagnosed during follow-up, or incidentally. METHODOLOGY: Sixty-one hepatocellular carcinomas were consecutively diagnosed in cirrhotic anti-HCV patients from 1993-1998 among which 34 during biannual ultrasonographic biochemical follow-up and the others incidentally. Child-Pugh's score, alpha fetoprotein levels, uni- or multifocality of the tumor, and treatment and survival of the patients were then analyzed on the basis of modality of diagnosis. RESULTS: Surgical treatment was feasible only in a minority of patients. Radical and palliative treatment was more frequent among patients with HCC diagnosed during follow-up. Child-Pugh's score was lower in these patients, moreover their survival rate was better. Analysis of survival of patients treated with the same procedure and grouped by modality of diagnosis did not demonstrate any differences. Regression analysis showed that patients with a lower Child Pugh's score, one nodule, with a tumor diagnosed during follow-up and who were treated had a better survival rate. CONCLUSIONS: In our population surveillance did not detect a higher percentage of curable HCC. Nevertheless the results of palliative treatment and of curative treatment overlapped. Overall better outcome was observed in patients with preserved liver function whatever the treatment. Surveillance allowed us to diagnose HCC in patients with these characteristics thus leading to an improved survival rate. PMID- 11100361 TI - Comparison of CT, MRI and CT during arterial portography in the detection of malignant hepatic lesions. AB - BACKGROUND/AIMS: A prospective study was performed to compare the sensitivities of computed tomography, magnetic resonance imaging and CTAP (CT during arterial portography) in the detection of focal malignant hepatic lesions. METHODOLOGY: Twenty-eight (28) patients with primary and secondary hepatic malignant tumors were evaluated. All of these patients underwent hepatic resection and a lesion-to lesion imaging-pathological analysis was performed. RESULTS: The overall sensitivities were 53% for CT, 66% for MRI sequences and 88% for CTAP. For lesions smaller than 1 cm the sensitivities were 6% for CT, 17% for MRI and 72% for CTAP. The combination of CTAP and MRI yielded an overall detection rate of 93%. The difference between the sensitivity of CTAP and that of the other two imaging techniques was statistically significant (P < 0.04) according to the McNemar test. CTAP demonstrated four false-positive lesions, two of which were correctly characterized by MRI and one by CT. In 6 patients (21.4%) the surgical plan was modified after CTAP. CONCLUSIONS: We conclude that, CTAP has the highest sensitivity and should be part of the preoperative examination. In some instances, the addition of MR imaging must be considered a helpful adjuvant. Both techniques should be considered complementary in the preoperative diagnostic algorithm. PMID- 11100362 TI - Thrombocytopenia associated with liver cirrhosis and hepatitis C viral infection: role of thrombopoietin. AB - BACKGROUND/AIMS: Thrombocytopenia in chronic liver diseases has traditionally been considered a consequence of platelet pooling and destruction in spleen. We tried to evaluate the influence of thrombopoietin, the physiological regulator of thrombopoiesis, on the origin of this thrombocytopenia. METHODOLOGY: We determined serum thrombopoietin levels by ELISA in thrombocytopenic patients with liver cirrhosis (n = 32) and with chronic hepatitis C viral infection (n = 23). A group of 43 healthy subjects was used as a control. RESULTS: Liver cirrhosis patients presented slightly, but not significantly, lower serum thrombopoietin levels (104 +/- 56 pg/mL) than controls (121 +/- 58 pg/mL) or patients infected with chronic hepatitis C virus (125 +/- 40 pg/mL). No correlations were found between serum thrombopoietin concentrations and liver tests or hematological parameters. CONCLUSIONS: We conclude that low thrombopoietin production may play a role, along with hypersplenism, in the development of thrombocytopenia in patients with liver cirrhosis. Normal thrombopoietin levels exclude a defect in thrombopoietin production as a possible etiology for the thrombocytopenia in patients with chronic hepatitis C viral infection. However, a direct viral megakaryocyte infection or an immune mechanism could explain this thrombocytopenia, according to the thrombopoietin levels detected. PMID- 11100363 TI - Early enteral nutrition after hepatectomy to prevent postoperative infection. AB - BACKGROUND/AIMS: Enteral nutrition helps to prevent septic complications in patients with critical illness, but there are few reports on its use after hepatectomy. To evaluate its benefits, we studied the indications for it. METHODOLOGY: In a retrospective study we reviewed 67 hepatectomized patients, 19 with enteral nutrition after hepatectomy (EN group) and 48 without (TPN group). In the TPN group, the risk factors of postoperative infections were analyzed with discriminant analysis. Then we prospectively selected high-risk patients before surgery and started enteral nutrition soon after surgery. The incidences of postoperative infections were examined in the EN and TPN groups of high-risk patients. RESULTS: In high-risk patients selected using a new discriminant formula, the infection rate was decreased from 73.1% in the TPN group to 53.3% in the EN group (NS). However, in cases in which enteral nutrition was initiated within 4 days after surgery, the infection rate dropped to 30%, a significant decrease (P < 0.03). In this prospective study, the infection rate of high-risk patients was markedly decreased and the overall infection rate was decreased significantly, to 21.4% from 47.4% in the retrospective study (P < 0.02). CONCLUSIONS: We conclude that early enteral nutrition after hepatectomy is helpful for preventing septic complications, especially in patients at high risk of infection as evaluated with our new formula. PMID- 11100365 TI - An autopsy case of multilocular cystic hepatocellular carcinoma without liver cirrhosis. AB - Although simple cysts, cystadenoma and cystadenocarcinoma of the liver have been well documented as hepatic cystic diseases, cystic hepatocellular carcinoma is a curious entity. Only 3 cases have been reported in the English literature. A 70 year-old man was admitted to Nagoya University Hospital for multiple liver tumors and a thrombus in the main trunk of the portal vein. A part of the tumors contained cystic components, and were diagnosed as hepatocellular carcinoma by needle biopsy. After giving informed consent, the patient was treated with several systemic chemotherapy using doxorubicin, fluorouracil, cyclophosphamide, cisplatin and oral anticancer agent UFT, a combination of uracil and tegafur, for almost 2 years. During this time, the tumors enlarged gradually, and also underwent cyst formation, the patients then died of biliary sepsis. Autopsy confirmed the diagnosis of multilocular cystic hepatocellular carcinoma without liver cirrhosis. PMID- 11100364 TI - Intraperitoneal chemohyperthermia for peritoneal carcinomatosis: original modeling, clinical tolerance and results study about 30 patients. AB - BACKGROUND/AIMS: The authors' objective is to report their experience of the intraperitoneal chemohyperthermia after a thermal modeling study which has allowed the optimization the intraperitoneal chemohyperthermia circuit and its running parameters and to evaluate the intraperitoneal chemohyperthermia tolerance. Intraperitoneal chemohyperthermia is considered more and more as an interesting therapeutic option in cases of some abdominal carcinomatosis, particularly of digestive origin. However, the main technical problem of this treatment is the homogenization of the temperature distribution in the abdominal cavity. METHODOLOGY: A thermal modeling has allowed us to finalize a reliable and well-tolerated intraperitoneal chemohyperthermia technique. The achievement of a physical model of the abdomen has allowed us to make an experimental study of the temperature distribution in a given liquid volume. Two steps were carried out. The first step was the characterization of the model with a thermal study carried out on the physical model and which has led to dynamic data about the heat balance leading to a knowledge model. The second step was the identification of a theoretical model of the thermal behavior which would correlate best with the experimental data. Between January 1995 and January 1998, 30 patients with peritoneal carcinomatosis were studied. Twenty-six patients underwent maximal cytoreductive surgery with abdominal evisceration, intraperitoneal chemohyperthermia. Intraperitoneal chemohyperthermia was carried out for 1 hour, at 42 degrees C, with a flow rate of 0.9 L/min in the 30 patients. The thermal modeling has shown the main purpose of a high flow rate of 0.9 L/min in the homogenization of temperature distribution. RESULTS: The 2 steps are shown to converge. This coherency between the 2 models proves that the thermal aspects of the process have been properly identified. Our initial results have shown that intraperitoneal chemohyperthermia was properly tolerated. Major intraoperative complications occurred for 1 patient. CONCLUSIONS: The experimental study with thermal modeling results should help to optimize the intraperitoneal chemohyperthermia circuit and its running parameters for human treatment, with an acceptable morbidity in 30 patients. PMID- 11100366 TI - Chronic hepatitis C, common variable immunodeficiency and autoimmune hemolytic anemia. Coincidence by chance or common etiology? AB - A case of chronic viral hepatitis is presented which is accompanied by an acquired immunodeficiency from the very beginning. Further problems arise by onset of an autoimmune hemolytic anemia. This is the first report on an association of chronic hepatitis C with acquired immunodeficiency and autoimmune hemolytic anemia. We assume in accordance with the patient's history that both have occurred as complications secondary to HCV infection. This adds another facet to the list of autoimmune phenomena during hepatitis C infection probably by induction of self-reactive B-cell clones. PMID- 11100367 TI - Polymyositis as a paraneoplastic manifestation of hepatocellular carcinoma. AB - A case of hepatocellular carcinoma associated with polymyositis is reported. A 70 year-old man noticed muscular weakness mainly in the proximal limb muscles. The clinical course, a raised level of serum creatine kinase and electromyographic findings suggested polymyositis, and the pathological findings on muscle biopsy were compatible with this diagnosis. Computed tomography of the upper abdomen revealed a mass lesion in segment VII and VIII of the liver, which was diagnosed pathologically as hepatocellular carcinoma. The patient underwent systematic resection of segment VII and the dorsal part of segment VIII of the liver. After surgery, the weakness improved and the serum creatine kinase level normalized without medical treatment for the polymyositis. The relief of neurological symptoms and signs after complete resection of the tumor strongly suggests paraneoplastic polymyositis, which has been described only rarely in association with hepatocellular carcinoma. PMID- 11100368 TI - Peritoneal seeding of hepatocellular carcinoma after ethanol injection therapy. AB - The tumor seeding due to percutaneous ethanol injection therapy has been considered to be a very rare complication. Four cases of peritoneal seeding of hepatocellular carcinoma following percutaneous ethanol injection therapy are presented here. All patients had been initially treated for hepatocellular carcinomas with percutaneous ethanol injection therapy. Between 5 and 20 months after the percutaneous ethanol injection therapy, peritoneal seeding tumors were detected and resected surgically. Three patients recurred in the liver and one patient recurred in the abdominal cavity. Two died of cancer and 2 are still alive. The incidence of seeding following percutaneous ethanol injection therapy should not be so rare as considered referring to that due to fine needle biopsy, therefore careful attentions should be paid during the follow-up of those patients after percutaneous ethanol injection therapy. PMID- 11100369 TI - Pleural effusion during interferon treatment for chronic hepatitis C. AB - A 54-year-old male patient was admitted to Osaka University Medical School Hospital for interferon treatment for chronic hepatitis C and the daily administration of recombinant interferon-alpha 2a started at the dose of 9 megaunits per day. Fourteen days later, moderate right pleural effusion was detected by abdominal magnetic resonance imaging study. He had experienced no symptom to suggest pleural effusion or any pulmonary lesions during interferon treatment. The pleural fluid was serous, showing the character of slightly bloody, turbid and positive Rivalta test, and the levels of lactic dehydrogenase and adenosine deaminase were not elevated. His serum titer of anti-nuclear antibody increased to 80 in homogenous staining, but anti-DNA antibody and anti liver kidney microsome-1 antibody remained negative. Other laboratory tests or physical findings could not satisfy the criteria of any autoimmune diseases, such as systemic lupus erythematosus. After discontinuation of interferon administration, his pleural effusion resolved gradually and disappeared completely by use of no specific drugs. This is the first case that pleural effusion developed during interferon administration without any other clinical signs indicating autoimmune diseases. The increase of serum titer of anti-nuclear antibody prompted us to elucidate that pleuritis might be induced by immunological activation of interferon. PMID- 11100370 TI - A case of the development of two hepatocellular carcinomas and a cholangiocarcinoma with cirrhosis after elimination of serum hepatitis C virus RNA with interferon therapy. AB - A 64-year-old man who had exhibited abnormal transaminase levels for about 20 years was admitted to a hospital for the treatment of liver damage. Laboratory testing demonstrated that he was suffering from chronic hepatitis C, and a liver biopsy showed chronic active hepatitis with septal fibrosis. He was treated with interferon-alpha, and HCV-RNA became undetectable. Four years after the completion of interferon treatment, 3 lesions in the patient's liver were revealed by routine ultrasonography, and he was referred to Nagoya University Hospital for further examinations on November 1997. Radiographical examinations such as computed tomography and angiography demonstrated 2 hypervascular tumors (size: phi 35 mm and phi 20 mm) and 1 hypovascular tumor (phi 28 mm). Qualitative analysis for HCV-RNA by polymerase chain reaction method was negative. Partial hepatectomy was performed, and pathological examination of the tumors showed 2 moderately differentiated hepatocellular carcinomas and cholangiocarcinoma accompanied with liver cirrhosis. We propose that even patients with disappearance of HCV-RNA after interferon therapy, especially with liver cirrhosis or severe fibrosis, should be followed-up closely and examined at regular intervals because of the high risk of developing primary liver cancers. PMID- 11100371 TI - Simple (non-parasitic) liver cysts: clinical presentation and outcome. AB - Non-parasitic hepatic cysts are frequent and usually asymptomatic. Investigation must differentiate from parasitic or neoplastic cysts. Ultrasonography, computed tomography, magnetic resonance imaging and serology do not always ensure a definitive diagnosis, where as other diagnostic methods offer little assistance. Symptoms, complications or uncertain diagnosis make treatment necessary. Various techniques have been used in management of non-parasitic hepatic cysts. Both surgical and recent minimally invasive methods such as laparoscopy and percutaneous aspiration with sclerotherapy are discussed and evaluated. Treatment of choice or indications for each method remains a controversial subject that requires further study. PMID- 11100372 TI - Distal pancreatectomy without splenectomy and with preservation of splenic vessels. AB - BACKGROUND/AIMS: Conventional distal pancreatic resection routinely involves splenectomy. The awareness that spleen removal may lead to postoperative septic and hematological complications motivated the development of spleen-preserving procedures. Successful distal pancreatectomy with splenic conservation has been reported for treatment of benign pancreatic diseases of the distal pancreas. This report presents the results of spleen-preserving distal pancreatectomy with conservation of the splenic artery and vein. METHODOLOGY: Ten patients underwent distal pancreatectomy with splenic vessel preservation. In all cases, both splenic vessels were separated from the pancreas towards the spleen after transecting the body of the pancreas. RESULTS: The indications for the procedure were: neuroendocrine pancreatic tumors (n = 4), cystic neoplasm of the pancreas (n = 4) and cystic-papillary pancreatic tumors (n = 2). Four patients developed pancreatic fistulas with spontaneous healing and there was no mortality. CONCLUSIONS: Spleen-preserving distal pancreatectomy with splenic vessel conservation can be safely performed and should be indicated in the surgical management of benign pancreatic diseases of the distal pancreas. PMID- 11100373 TI - Preoperative embolization of the common hepatic artery in preparation for radical pancreatectomy for pancreas body cancer. AB - BACKGROUND/AIMS: To assess preliminary results of preoperative embolization of the common hepatic artery in preparation for distal pancreatectomy with en bloc resection of the celiac and common hepatic arteries for carcinoma of the body of the pancreas involving these arteries. METHODOLOGY: Four patients underwent the embolization with coils 1-7 (median: 5) days before surgery. A detachable coil was used to obtain the best position of the first coil as an anchor in 3 patients. RESULTS: Immediately after embolization, collateral pathways developed from the superior mesenteric artery via the pancreatoduodenal arcades to the proper hepatic and gastroduodenal arteries in all 4 patients; however, they were relatively poor in one patient. There were no complications after embolization. The pulsation of the proper hepatic and gastroduodenal arteries was well palpable during surgery, although it had been compromised sometimes in previous cases without embolization. There were no ischemia-related complications in the 2 patients who underwent radical surgery. CONCLUSIONS: Preoperative embolization of the common hepatic artery is a safe technique and has the potential to enlarge the collateral pathways by the time of distal pancreatectomy with en bloc resection of the celiac artery and prevent postoperative fatal ischemia-related complications. PMID- 11100374 TI - Gemcitabine and protracted 5-FU for advanced pancreatic cancer. A phase II study. AB - BACKGROUND/AIMS: Although chemotherapy in advanced pancreatic cancer procures dismal results, both 5-fluorouracil and gemcitabine have shown a modest activity. We report a phase II study of gemcitabine combined with protracted 5 fluorouracil. METHODOLOGY: Gemcitabine was given at 1000 mg/m2/week intravenously, in combination with concomitant 5-fluorouracil 200 mg/m2/day as a protracted venous infusion, both 3 out of 4 weeks in patients with locally advanced or metastatic pancreatic adenocarcinoma. Twenty-nine patients were enrolled, among whom 27 were metastatic. Response rate, overall and progression free survival were endpoints, as well as tolerance and clinical benefit. RESULTS: We observed 3 (10%) partial responses, and 12 (42%) stabilizations within which the median disease control was 5.6 months. The median progression-free and overall survivals were 2.8 and 4 months, respectively. A clinical benefit was observed in 39% of patients. Myelosuppression was the main toxicity, but no grade 4 was observed. Other toxicities were mild. CONCLUSIONS: This combination chemotherapy was well tolerated in advanced pancreatic cancer patients. PMID- 11100375 TI - A retrospective analysis of 88 patients with pancreaticogastrostomy after pancreaticoduodenectomy. AB - BACKGROUND/AIMS: Recently, pancreaticogastrostomy after pancreaticoduodenectomy has been reintroduced as a useful procedure with a low incidence of pancreatic leakage. We decided to retrospectively analyze the early postoperative and late follow-up complications in a large number of patients who had undergone our improved pancreaticogastrostomy. METHODOLOGY: Between August 1993 and November 1998, we performed pancreaticogastrostomy following pancreaticoduodenectomy in 88 consecutive patients including pylorus preservation in 14. Our pancreaticogastrostomy used a two-layer implantation method using a pancreatic duct stent with an anterior gastrotomy. RESULTS: The morbidity and mortality rates were 5.7% and 0%, respectively. There were no cases of pancreatic leakage and no postoperative complications directly related to the pancreaticogastrostomy. As for gastric emptying, the average time until resumption feeding was 12 days, with no significant difference between pancreaticogastrostomy and pylorus-preserving pancreaticogastrostomy. As to late follow-up complications, diabetes developed postoperatively in 6.5% (4/62) patients, and of 14 patients who were shown by magnetic resonance cholangiopancreatography, 5 (35.7%) developed pancreatic ductal dilatation after surgery. CONCLUSIONS: These results from a relatively large group conclusively prove the safety of pancreaticogastrostomy and indicate that the follow-up quality of life is good under the circumstances. PMID- 11100376 TI - Pancreatic neuroendocrine tumors. Report of nine cases. AB - BACKGROUND/AIMS: Tumors of the endocrine pancreas are infrequent, and their malignant behavior is assessed only in the presence of lymph node or hepatic metastases. We present 9 new cases from the past 11 years. METHODOLOGY: We reviewed the clinical records of 9 patients diagnosed of pancreatic neuroendocrine tumors, analyzing age, sex, past medical history, symptoms, clinical presentation, laboratory tests, imaging studies, operative findings, pathological diagnosis, mortality, morbidity and hospital stay. RESULTS: There were 5 women and 4 men, with a mean age of 48.7 years. In 4 patients the tumor presented with hypoglycemia. Malignant behavior was observed in 2 cases. We discuss the origin and classification of these tumors, as well as the best diagnostic and therapeutic approaches. CONCLUSIONS: Endocrine tumors of the pancreas affect middle aged men and women, presenting with specific signs or symptoms in less than half of the cases. In small tumors preoperative or intraoperative localization may be difficult. Most of the lesions are localized in the tail of the pancreas, and malignant behavior is seen in less than 25% of the cases. PMID- 11100377 TI - Epidermoid cyst of an intrapancreatic accessory spleen--a case report. AB - We report an extremely rare case of a splenic epidermoid cyst of the pancreas in a 51-year-old Japanese male with no clinical symptoms. A cystic tumor of the pancreatic tail was detected incidentally by abdominal ultrasonography. The patient was referred to the Gunma University Hospital for further examination of the pancreatic tumor. Upon diagnosis of a benign cystic tumor, a distal pancreatectomy with splenectomy was performed. Microscopically, the multicystic tumor, which was surrounded by the splenic tissue, was located within the pancreatic tissue. The cysts were lined by non-keratinizing squamous epithelium. The diagnosis of an epidermoid cyst occurring in an intrapancreatic accessory spleen was confirmed. To our knowledge, this is the 4th case ever reported in the English literature. PMID- 11100378 TI - Expression patterns of the E-cadherin-catenin cell-cell adhesion complex in gastric cancer. AB - BACKGROUND/AIMS: The E-cadherin-catenin complex plays a key role in intercellular adhesion of epithelial cells. Aberrant expression and/or function of its components have been implicated in tumor progression and metastasis. We evaluated the expression of the E-cadherin-catenin complex in gastric cancer by immunohistochemistry and investigated its relationship to histopathological features. METHODOLOGY: The expression of E-cadherin, alpha-, beta-, and gamma catenin, and p120 protein was evaluated by immunohistochemistry in 36 formalin fixed, paraffin-embedded specimens of gastric cancer. RESULTS: In benign gastric mucosa all five molecules co-localized at the cell membrane. Reduced E-cadherin, alpha-, beta-, and gamma-catenin, and p120 expression was found in 67%, 61%, 50%, 64%, and 56% of cases, respectively. The expression of E-cadherin and beta catenin correlated significantly with the histological type and the degree of tumor differentiation. gamma-Catenin expression correlated only with the histological type of the tumor. The expression of E-cadherin correlated significantly with alpha-, beta- and gamma-catenin, and p120 expression, respectively. The expression of E-cadherin and alpha-catenin showed the highest concordance. CONCLUSIONS: In gastric cancer, reduced E-cadherin, catenin and p120 expressions are related events with E-cadherin showing the most frequent aberrations. PMID- 11100379 TI - Does extended lymphadenectomy influence prognosis of gastric carcinoma after curative resection? AB - BACKGROUND/AIMS: It is unclear whether gastric cancer prognosis is improved by extended lymph node dissection more than by lymph node dissection limited to the contiguous N1 perigastric lymph nodes. METHODOLOGY: Four hundred and thirty-eight patients treated by curative gastrectomy were evaluated. Outcomes of D1/D1.5 lymphadenectomy, limited lymph node dissection and of D2/D2.5 lymphadenectomy, extended lymph node dissection and histopathological prognostic factors as in the 1993 TNM staging classification supplement were analyzed. RESULTS: Estimated overall 5-year survival was 54.9%. Five-year survival was 58.4% in the limited lymph node dissection group and 54% in the extended lymph node dissection (P n.s.). Stage I 5-year survival was 59% after D2.5 lymph node dissection, 58% after D1.5 and 50% after D2 dissection (P n.s.). Stage II 5-year survival was 86% in D2.5 group and 56% in D1.5 group (P = 0.041). Stage IIIa survival was 61% in the D2.5 group and 22% in the D1.5 group (P = 0.001). Stage IIIb 5-year survival was 42% after D2.5 resection and 0% in D1.5 group (P = 0.001). In the pT3 group 5 year survival was 72% after D2.5 dissection and 33% after D2 dissection (P = 0.001). In the positive N1 lymph nodes group 5-year survival was better after extended lymph node dissection than after limited lymph node dissection. In pN2a patients 5-year survival was 57% after D2.5 resection and 0% after D2 resection (P < 0.001). In pN2b and pN2c patients extended lymph node dissection did not statistically improve survival. CONCLUSIONS: Even if no statistical differences were found in overall survival, prognosis was improved by extended lymph node dissection in stage II and III, particularly in T2 and T3 subgroups and in N1 and N2a subgroups. When large numbers of positive nodes were found, improved survival was dependent upon resection of extragastric nodes distal to the uppermost echelon of positive nodes. PMID- 11100380 TI - Perigastric lymph node status can be a simple prognostic parameter in patients with gastric cancer. AB - BACKGROUND/AIMS: The number of metastatic lymph nodes has been a significant prognostic factor after curative resection of gastric cancer and adopted as a new UICC classification of nodal stages in gastric cancer. The extent of lymphadenectomy is another significant factor but has been fiercely debated. Regardless of the type of lymphadenectomy, perigastric lymph node dissection is always carried out. In this study, we examined whether the number of metastatic perigastric nodes can be a prognostic indicator of gastric cancer. METHODOLOGY: For the purpose of evaluating perigastric lymph node status, a retrospective study was carried out with 760 patients who underwent curative gastric resection from June 1994 to November 1998. RESULTS: The 4-year cumulative survival rate was 64% and the survival rate decreased significantly when the number of positive perigastric nodes exceeded 3. Comparing with the patients having 0-2 positive perigastric nodes, patients whose metastatic perigastric lymph nodes exceeded 3 or more exhibited deeper tumor invasion, larger tumor size and older age. Multivariate analysis identified the number of positive perigastric nodes, together with depth of tumor invasion, as the strongest independent prognostic factors for survival. CONCLUSIONS: We suggest that the number of metastatic perigastric nodes can be used as a simple prognostic parameter in patients with gastric cancer and that intensive follow-up and adjuvant chemotherapy should be recommended for the patients with more than 3 metastatic perigastric nodes. PMID- 11100381 TI - The resection of non-hepatic intraabdominal recurrence of gastric cancer. AB - BACKGROUND/AIMS: Surgical resection of hepatic or pulmonary metastases from gastrointestinal cancer has been recognized as a curative modality in some patients. However, the role and outcome of the surgical management of a non hepatic intraabdominal recurrence of gastrointestinal cancer have not been clearly delineated. METHODOLOGY: We treated 5 patients for non-hepatic intraabdominal recurrence of gastric carcinoma surgically. All the resected specimens were microscopically identified as recurrent gastric cancer. Three of 5 patients received postoperative chemotherapy. The clinicopathological findings were analyzed according to the general rules for gastric cancer study. RESULTS: The lymph nodes were dissected for lymph node metastases. Surgical resection of the tumors combined with total gastrectomy, esophagectomy, or colectomy was performed for the local and peritoneal recurrences. All of the recurrent tumors were macroscopically resected with curative states. One patient died of sepsis 54 days after surgery. Three patients died of recurrent gastric cancer: 2 within 1 year of surgery and 1 after 3 years. One patient still survives disease free 3 years and 6 months after the 2nd operation. CONCLUSIONS: Surgical resection for non-hepatic intraabdominal recurrence of gastric cancer is the treatment of choice for selected patients. Surgical resection followed by adjuvant chemotherapy may improve the outcome of these patients. PMID- 11100382 TI - The influence of Helicobacter pylori infection on gastrin and somatostatin values present in serum. AB - BACKGROUND/AIMS: Recent studies on the role of Helicobacter pylori in pathogenesis of duodenal ulcers have focused on the mechanism by which H. pylori infections causes exaggerated gastrin release. METHODOLOGY: We compared the gastrin and somatostatin serum values between two groups of patients; 37 H. pylori-positive ones and 29 H. pylori-negative ones. We applied radioimmunoassay technique to determine the gastrin and somatostatin values in serum. H. pylori was confirmed by urease test and by histopathological color according to Giemsa. RESULTS: The level of gastrin in the serum of Helicobacter pylori-positive patients with chronic gastritis were significantly higher in relation to H. pylori-negative patients. The somatostatin concentration in the sera of H. pylori positive patients with duodenal ulcer (16.27 +/- 9.49 pg/mL) were less in comparison with those without duodenal ulcer (23.25 +/- 13.59 pg/mL). CONCLUSIONS: The results suggest that H. pylori infection suppresses the somatostatin secretion. PMID- 11100383 TI - Clinicopathological features of long-term survivors of scirrhous gastric cancer. AB - BACKGROUND/AIMS: Prognosis of scirrhous gastric cancer remains low. To determine the clinicopathological features that are correlated with prognosis, we studied long-term survivors of scirrhous gastric cancer (survival duration more than 5 years) in comparison with patients with short survival. METHODOLOGY: Among 2719 gastric cancer patients who underwent surgery at Matsuyama Red Cross Hospital, 211 cases were diagnosed as scirrhous type gastric cancer. Seventeen patients survived more than 5 years, and the rest had short survival (less than 5 years). Comparison of clinicopathological factors was done by chi 2 analysis. Multivariate analysis was done in order to focus on the prognostic factors. RESULTS: The 5-year survival of the total 211 patients was 12%. The 5-year survival of patients who underwent curative surgery (67 cases) was 30%, which was significantly higher than that of the non-curative surgery group (144 cases, 6%). Significant differences were noted in the following variables: peritoneal dissemination, hepatic metastasis, lymph node dissection, pattern of infiltrating growth, depth of invasion, histological lymph node metastasis, histological stage, and histological curability. Patients with either hepatic metastasis or peritoneal dissemination did not survive 5 years. Multivariate analysis revealed that the most significant independent prognostic factor was histological curability, followed by peritoneal dissemination. CONCLUSIONS: There is a possibility of long-term survival for patients with scirrhous gastric cancers without hepatic metastasis, peritoneal dissemination, or extensive lymph node metastasis. Curative surgery is important, suggesting that the extended operation is rational if possible. PMID- 11100384 TI - Detection and prognosis of recurrent gastric cancer--is routine follow-up after gastrectomy worthwhile? AB - BACKGROUND/AIMS: Although routine follow-up after surgery for gastric cancer is recommended its value after gastrectomy has not been evaluated. METHODOLOGY: All patients who underwent gastrectomy for gastric cancer entering the routine follow up program between January 1987 and August 1996 were identified. The patients studied were those with either histologically proven recurrence or those in whom recurrence was highly probable from clinical course. Two groups were compared. The first group comprised the patients whose recurrence was detected by routine follow-up prior to the development of clinical signs (asymptomatic group). The second group consisted of the patients who developed clinical symptoms due to a recurrence that was detected subsequently (symptomatic group). The main parameters were the time until recurrence occurred, the pattern of recurrence, treatment and survival. RESULTS: Out of 184 patients entering the routine follow up 135 patients had undergone potentially curative gastrectomy. Sixty-seven patients (49.6%) had recurrences. Only 15 (22.3%) belonged to the asymptomatic group and 52 (77.7%) to the symptomatic one. The time until recurrence occurred was not different between the 2 groups (17.1 vs. 18.0 months). Chemotherapy was performed more frequently in the asymptomatic group and survival was longer (8.4 vs. 5.9 months). This difference was due to the time the patients remained asymptomatic (average 43 months). No effect of either early detection or chemotherapy was seen. In the asymptomatic group distant recurrence was common while recurrence in the symptomatic group was more often local but this difference did not reach statistic significance. CONCLUSIONS: Routine follow-up after gastrectomy for gastric cancer does not contribute to early detection of gastric cancer recurrence. It has no benefit with respect to treatment and survival of patients with recurrent disease and should therefore be reduced to symptomatic and psychological aftercare. PMID- 11100385 TI - New method for jejunal pouch interposition reconstruction after distal gastrectomy. AB - BACKGROUND/AIMS: Recently pouch reconstruction has been reported to improve quality of life and functional results after surgery for gastric cancer. Although jejunal pouch reconstruction after distal gastrectomy has favorable results for patients' quality of life, it is complicated and takes a long time to complete. We developed a new technique using a linear stapling device to avoid this problem. METHODOLOGY: The duodenum and the jejunum are simultaneously divided with a 100-mm linear stapler 0.5 cm distal to the pyrolus ring and 20 cm distal to the ligament of Treitz, respectively. A 100-mm linear stapler is introduced into two approximated segments of the jejunum through two small stab wounds 10 cm and 15 cm distal to the stump, respectively, and side-to-side anastomosis is performed along the antimesenteric borders. The anterior wall of the pouch is cut along the prospective line of anastomosis with the gastric remnant. The anterior wall of the stomach is cut along the planned suture line having a length similar to that of the pouch. The posterior walls of the stomach and the jejunal pouch are placed back-to-back on the planned anastomotic line. End-to-end posterior anastomosis between the gastric remnant and the jejunal pouch is simultaneously performed with gastrectomy using a 100-mm linear stapler. End-to-end anterior anastomosis is created by hand. RESULTS: This technique has been used in 4 patients, and there have been no complications related to the pouch or anastomoses. Mean operative time was 255 +/- 37 min (range: 205-290 min). CONCLUSIONS: Shortening of operative time can be attributed to adoption of end-to end posterior anastomosis between the stomach and the jejunal pouch using the linear stapling device simultaneously with gastrectomy. PMID- 11100386 TI - The efficacy of 1 alpha hydroxy vitamin D3 treatment of the metabolic bone disorder in patients who underwent gastrectomy for gastric cancer. AB - BACKGROUND/AIMS: The aim of this study was to elucidate the efficacy of 1 alpha hydroxy vitamin D3 for treatment of metabolic bone disorder after gastrectomy. METHODOLOGY: MD-MS method was performed in 16 patients who underwent gastrectomy to evaluate the metabolic bone disorder and compared before and after 1 alpha hydroxy vitamin D3 treatment. The symp-toms related to the metabolic bone disorder were also analyzed through patient interview. RESULTS: The severity of the metabolic bone disorder analyzed by the MD-MS method improved after 1 alpha hydroxy vitamin D3 treatment in 56.3% of patients. The deviation degree of the cortical bone density was also significantly improved. The symptoms related to metabolic bone disorder disappeared in 80% of patients after treatment. CONCLUSIONS: The administration of 1 alpha hydroxy vitamin D3 may be useful for the metabolic bone disorder in the treatment of patients after gastrectomy. PMID- 11100387 TI - The influence of local heating on skin microcirculation in pressure ulcers, monitored by a combined laser Doppler and transcutaneous oxygen tension probe. AB - In a prospective, controlled study the skin microcirculation and transcutaneous oxygen tension were evaluated in 28 elderly patients (85 +/- 0.8 years) with grade 2 and 3 pressure ulcer. Laser Doppler fluxmetry and transcutaneous oxymetry techniques were used together in a combined probe to simultaneously evaluate the influence of heat stress at 44 degrees C, at the same time and in the same area of the ulcer edge. Total skin microcirculation was already increased at rest before heating, and increased even more during a fast and then a more gradually heat-induced hyperaemia response compared with undamaged skin. The increase showed a biphasic manner. On the other hand, the oxygen diffusibility from the capillaries to the skin surface was significantly reduced, compared with undamaged skin. One hypothesis may be that in ageing skin the main part of the increased skin microcirculation in a pressure ulcer were passing through thermoregulatory vessels in subpapillary tissue layers. Interactions between the increased blood flow and metabolic activity at the ulcer edge might be oxygen consuming, leading to reduced oxygen content passing through the capillaries and contributing to tissue ischaemia. Disturbances of the local skin microcirculation and tissue oxygenation will influence the ulcer healing rate and may affect the healing progress. PMID- 11100388 TI - Circadian systemic haemodynamics in borderline and mild hypertension. AB - Circadian variations in blood pressure (BP), stroke volume (SV), heart rate (HR), cardiac output (CO) and total peripheral resistance (TPR) were determined by a pulse contour method from the intra-arterial pulse wave in 32 normotensive (NT), 32 borderline hypertensive (BHT) and 31 hypertensive (HT) middle-aged men. Daytime averages were used as the reference levels. The nocturnal decrease in BP and HR were similar in the three groups. In the night, SV did not change in the NT group, but was increased in the BHT and HT groups. The nocturnal increase in SV may reflect reduced venous capacity causing increased cardiac filling. As a consequence of the difference in SV, the nocturnal CO fall was diminished in the HT group as compared with the NT group. Moreover, TPR had a tendency to decrease in the HT group, which may be considered as a baroreflex response to buffer the expected rise in BP. Five years later, 25 NT, 24 BHT and 19 HT subjects were reassessed using casual BP measurements. In the NT and BHT groups, six and 17 subjects, respectively, had progressed to hypertension. In a logistic regression model for those who became HT, the nocturnal increase in SV was a significant predictor for future hypertension. In conclusion, the results suggest that circadian systemic haemodynamics may be altered before BP is markedly elevated, and that haemodynamic studies might be useful in predicting the development of sustained hypertension. PMID- 11100389 TI - Technical achievement: transthoracic Doppler echocardiography can be used to detect LAD restenosis after coronary angioplasty. AB - We investigated the capability of transthoracic Doppler echocardiography (TTE) to detect and quantify the severity of restenosis in the left anterior descending coronary artery (LAD) after percutaneous transluminal coronary angioplasty (PTCA). We studied 10 consecutive patients assigned for quantitative coronary angiography (qCA) due to a recurrent angina pectoris after PTCA of the LAD. The LAD was visualized by TTE, and the presence of local turbulence and an increase in the blood flow velocity was regarded to indicate coronary stenosis. To assess the severity of the stenosis, the increase of blood flow velocity was measured. Angiography showed stenoses of various degrees (27-100%) in all patients. All stenoses were detectable using TTE. Moreover, the ratio of maximal blood flow velocity at the site of stenosis to the pre-stenotic blood flow velocity (M/P ratio) correlated significantly with the reduction of the luminal diameter of LAD (r = 0.85, P < 0.003). A M/P-ratio higher than 3.0 predicted a diameter reduction of 50% or higher with sensitivity and specificity of 100% in patients with a subtotal stenosis (n = 9). Our results indicate that stenoses in the LAD could be found and the severity of the stenoses could be quantified reliably with TTE. This approach is totally non-invasive and less expensive than coronary angiography and can be used clinically in clarifying restenosis after coronary angioplasty. PMID- 11100390 TI - Exhaled nitric oxide and its long-term variation in healthy non-smoking subjects. AB - Exhaled nitric oxide (NOexp) is an indicator of inflammation in the airways. Reference values obtained from healthy adults or information on long-term variation of NOexp are not yet available. The aims of this pilot study were to collect values of NOexp from a selected group of healthy adults and to assess their long-term variation. We studied 26 healthy subjects (age 21-48, 16 male, 10 female) with normal findings in flow-volume spirometry, pulmonary diffusing capacity, relative amount of blood eosinophils, chest X-ray and ECG at rest. NOexp was determined according to the European Respiratory Society guidelines during slow expiration against an airflow resistance. The measurements were repeated after 7 (n = 13) and 23 days (n = 17). The mean value of NOexp (n = 26) was 6.9 ng g-1 (95% confidence interval, 6.0-7.9 ng g-1). The upper limit of intra-individual variation (+2 SD) was 11.9 ng g-1 and the lower limit (-2 SD) 1.9 ng g-1, respectively. The mean (SD) value of NO production (NO output) was 39.1 pmol s-1 (20 pmol s-1). We found no correlation between NOexp and age (r = 0.06, P = 0.78) and no association of NOexp with the gender (male vs. female, P = 0.40). The intraindividual coefficient of variation (CoV) was 15.8% of NOexp and 20.7% of NO output within the interval of 7 days. CoV was 16.8% of NOexp and 18% of NO output within the interval 23 days. The results suggest that NOexp values over 12 ng g-1 are abnormally high in healthy subjects. According to the results the change of NOexp by 30-35% or more within the interval of 1-3 weeks would be abnormal. PMID- 11100391 TI - Evaluation of endothelium-dependent vasodilation in the human peripheral circulation. AB - Flow-mediated vasodilation (FMD) in the brachial artery measured by ultrasound, and the increase in forearm blood flow (FBF) induced by local infusion of a muscarinic-receptor agonist have both frequently been used to evaluate endothelium-dependent vasodilation (EDV) in the human forearm. The present study intended to evaluate the relationship between these techniques and to investigate if vasodilation induced by the muscarinic receptor-agonist methacholine (MCh) was owing to production of nitric oxide (NO). FMD during hyperaemia was assessed by ultrasound and FBF was measured by venous occlusion plethysmography during local infusion of MCh or L-arginine in the human forearm. Both these methods were applied in 26 individuals. In another 12 individuals forearm arterial and venous plasma concentrations of nitrate/nitrite (NOx) were measured together with FBF before and during local MCh infusion. While the change in brachial artery diameter induced by sublingually given nitroglycerine and the vasodilatory response to sodium nitroprusside (SNP) given locally in the forearm were significantly correlated (r = 0.70, P < 0.01), FMD showed no relationship with the vasodilation evoked by MCh (r = -0.03) or L-arginine (r = 0.04). The five fold increase in FBF during MCh infusion was associated with a significant increase in venous plasma NOx concentrations (P < 0.05) and a more than 11-fold increase in forearm NOx-release (P < 0.01). Thus, a significant relationship between the two methods regarding the evaluation of endothelium-independent vasodilation evoked by NO-donors was found, but no relationship was found between the two methods regarding the evaluation of endothelium-dependent vasodilation. Furthermore, vasodilation induced by MCh in the forearm seems to be induced by NO release. PMID- 11100392 TI - Lack of effect of acupuncture upon signs and symptoms of delayed onset muscle soreness. AB - The effect of acupuncture upon experimentally induced delayed onset muscle soreness (DOMS) was assessed in a placebo-controlled study under blinded conditions. Volunteers (n = 48; 24 M & 24 F) were randomly allocated to one of four groups: control (20 min rest), placebo (minimal needling at non-acupuncture points), treatment group 1 (acupuncture at classic acupuncture points) and treatment group 2 (acupuncture at 'tender' points). DOMS was induced in the elbow flexors of the non-dominant arm using a standardized eccentric exercise regime. Measurements of elbow range of movement (flexion, extension, relaxed angle), and pain as well as visual analogue scores (VAS), tenderness (using a pressure algometer) were employed as indices of treatment efficacy. Measurements of elbow range of movement and tenderness were made prior to DOMS induction on the first day, and repeated pre- and post-treatment on subsequent days; pain was assessed using visual analogue scales post-induction and post-treatment on the first day, and pre- and post-treatment thereafter. For all conditions, subjects rested supine for a period of 20 min, during which treatment was delivered according to group allocation. Repeated measures and one-way analysis of variance (ANOVA) demonstrated no significant interactive (AB) effects, except for visual analogue scores (P = 0.0483); one factor ANOVA on the second day of the experiment (pre treatment) indicated significant differences between the control and all other groups. However, such differences were not found on any other day of the experiment. It is concluded that acupuncture has little effect upon the cardinal signs and symptoms of DOMS, at least under the conditions of the current experiment. PMID- 11100393 TI - Effects of low-dose adrenomedullin on cardiac function and systemic haemodynamics in man. AB - The cardiovascular effects of low-dose adrenomedullin (ADM, 1, 2 and 3 pmol kg-1 min-1 for 30 min each) were evaluated in six healthy subjects in a placebo controlled, cross-over study by determining cardiac volumes, systolic and diastolic function (echocardiography) and systemic haemodynamics before, during and after ADM or placebo. High-resolution ultrasound was used to evaluate changes in carotid artery distension. ADM caused a +85% increment in its plasma levels and significantly increased plasma cyclic adenyl monophosphate (cAMP). Compared with placebo, ADM induced significant decrements in left ventricular (LV) systolic diameter and systemic vascular resistance, and increments in LV posterior wall thickening, ejection fraction and cardiac index. Right and left atrial emptying fraction and carotid artery distention increased. LV diastolic function, heart rate, and plasma renin activity did not change, whereas packed cell volume increased. These results indicate that ADM influences cardiovascular function and systemic haemodynamics at physiological plasma levels in man mainly because of its vasodilating activity, leading to reduced afterload. PMID- 11100394 TI - Achieving haemodynamic baseline values with Finapres in elderly subjects during supine rest. AB - BACKGROUND: Clear guidelines for the resting time necessary to achieve stable blood pressure (BP) levels are scant in gerontology research. Therefore, we aimed to determine the minimum period required for obtaining haemodynamic baseline values in elderly subjects during supine rest. In addition, we evaluated the effect of cardiovascular morbidity, such as diastolic heart failure, and the effect of complex comorbidity of geriatric patients, on haemodynamic changes during supine rest. METHODS: A total of 17 healthy subjects, 18 heart failure patients with normal systolic function and 24 geriatric patients, aged 70 years and more, participated. After an overnight fast, changes in systolic BP (SBP), diastolic BP (DBP), heart rate (HR), and stroke volume (SV) were determined by Finapres beat-to-beat non-invasive BP monitoring during a 20-min supine rest. The procedure was repeated in the healthy subjects and geriatric patients on a second day. RESULTS: Complete BP stabilization was reached in each group within 5 min of supine rest, as SBP remained essentially unchanged and DBP did not change significantly anymore after the fourth minute. In the heart failure patients, HR decreased and SV increased until the twelfth minute of rest. The SBP, DBP, HR, and SV changes during supine rest showed good reproducibility. CONCLUSIONS: A span of 5 min of supine rest ensured achievement of reliable and reproducible baseline BP values by Finapres in elderly subjects. However, we recommend at least 12 min of rest to obtain full haemodynamic stability in elderly patients with diminished cardiac compliance and diastolic function. PMID- 11100395 TI - The effects of length on fatigue and twitch potentiation in human skeletal muscle. AB - Fatigue is the decrease in active force that happens after repeated muscle stimulation, and post tetanic twitch potentiation (PTP) is the increase in twitch force observed after repeated muscle stimulation. This study investigated the effects of length on the interaction between fatigue and PTP, as these two forms of force regulation are length-dependent and may coexist. A total number of 14 subjects were tested in 3 days, in which fatigue and PTP were induced in the knee extensor muscles in three different knee angles (30 degrees, 60 degrees and 90 degrees; full extension = 0 degree). PTP was evaluated in rested and fatigued muscles with twitch contractions elicited before and after 10 s maximal voluntary contraction (MVC), and fatigue was evaluated with nine 50 Hz electrically elicited contractions (5 s duration, 5 s interval between contractions). Fatigue was length-dependent, with force values that were (mean +/- SEM) 59 +/- 5, 56 +/- 3 and 38 +/- 1% of maximal force at 90 degrees, 60 degrees and 30 degrees, respectively. PTP was also length-dependent. Rested muscles showed PTP of 39 +/- 4, 47 +/- 2 and 68 +/- 5% at 90 degrees, 60 degrees and 30 degrees, respectively. Fatigued muscles showed PTP of 44 +/- 3, 55 +/- 6 and 68 +/- 5%, at 90 degrees, 60 degrees and 30 degrees, respectively. This study shows that fatigue and PTP may represent independent mechanisms, as they regulate force in opposite directions and are both enhanced in short muscle lengths. PMID- 11100396 TI - Mental stress increases right heart afterload in severe pulmonary hypertension. AB - Little is known about mental stress effects on the pulmonary circulation in health and disease. The current study was conducted to investigate whether pulmonary artery pressure (PAP) and pulmonary vascular resistance (PVR) would further increase during standardized mental stress testing in patients with severe pulmonary hypertension. The study was a prospective analysis of seven patients (average age: 40 years, range from 21 to 56 years) with severe pulmonary hypertension (primary: n = 4, secondary forms: n = 3; resting mean pulmonary artery pressure ranged between 48 and 65 mmHg). Right heart catheterization for the determination of PAP, pulmonary capillary wedge pressure (PCW) and cardiac output (CO) was clinically indicated (diagnostic workup, acute drug testing). Patients accomplished a standardized 10 min mental stress test (computer based, adaptive complex reaction-time task). Pulmonary haemodynamics during stress were compared to resting baseline. During mental stress mean PAP (+/- SEM) increased by 9.4 +/- 2.1 mmHg (P < 0.005). Pulmonary vascular resistance increased by 149 +/- 25 dyne s cm-5 (P < 0.001). Stroke volume decreased by 6.6 +/- 2.2 ml (P < 0.03). The data show that moderate mental stress increases right heart afterload in patients with severe pulmonary hypertension owing to elevation of PVR. PMID- 11100397 TI - Can stroke volume and cardiac output be determined reliably in a tilt-table test using the pulse contour method? AB - The applicability of the finger pressure-derived pulse contour (PC) technique was evaluated in the measurement of stroke volume (SV), cardiac output (CO) and their changes in different phases of the tilt-table test. The reference method was whole-body impedance cardiography (ICG). A total number of 40 physically active patients, aged 41 +/- 19 years, were randomly chosen from a pool of 230. Specifically speaking, 20 of the patients experienced (pre)syncope (tilt+ patients) during the head-up tilt (HUT), and 20 did not (tilt-). A total number of three measurement periods, 30-60 s each, were analysed: supine position, 5 min after the commencement of HUT, and 1 min before set down. SV and CO values measured by PC underestimated significantly those measured by ICG (biases +/- SD 19 +/- 14 ml and 1.55 +/- 1.14 l min-1, respectively) in agreement with earlier reports. The bias between the methods was almost the same in the different phases of the test. However, the SD of the bias was bigger for tilt+ (P < 0.05). When the bias between the methods was eliminated by scaling the first measurement to 100%, the agreement between the methods in the second and third measurements was clearly better than without scaling. Both methods showed a physiological drop in SV after the commencement of HUT. These results indicate that PC suffices in tracking the changes in CO and SV, but for absolute values it is not reliable. PMID- 11100398 TI - Cardiovascular functioning during the menstrual cycle. AB - Variations in cardiovascular functioning during the 'normal' menstrual cycle have been little researched. Resting-blood pressures, resting-heart rate, rate pressure product (RPP) and a derived index of fitness (Schneider Index) were monitored throughout natural, hormonally defined menstrual cycles. Volunteers were 26 women (20-48 years) who had regular (25-35 days) cycles. Their blood pressures and heart rate (at rest and according to Schneider's protocol) were measured at the same time daily (Monday-Friday) for 5 weeks. Daily, early morning urine samples were assayed for sex hormones enabling accurate definition of cycle phase for each woman. Resting systolic-blood pressure was significantly higher in the ovulatory phase (P < 0.05) than in the follicular or luteal phases, but resting-diastolic pressures did not differ significantly between phases. Resting heart rate was significantly higher in both ovulatory (P < 0.01) and luteal (P < 0.01) phases than in the menstrual and follicular phases. The Schneider Index was higher during the follicular phase than during the ovulatory (P < 0.005) or luteal (P < 0.01) phases, the RPP was higher during the ovulatory phase than during the bleeding (P < 0.05) and follicular (P < 0.005) phases. These findings provide a pattern of menstrual cycle-related variation in cardiovascular functioning that can be related to established actions of the ovarian steroids. PMID- 11100399 TI - Di(2-ethylhexyl) phthalate. PMID- 11100400 TI - Di(2-ethylhexyl) adipate. PMID- 11100401 TI - Cinnamyl anthranilate. PMID- 11100402 TI - Coumarin. PMID- 11100403 TI - Ethylbenzene. PMID- 11100404 TI - ortho-toluidine. PMID- 11100405 TI - 4-Chloro-ortho-toluidine. PMID- 11100406 TI - 5-Chloro-ortho-toluidine. PMID- 11100407 TI - Diethanolamine. PMID- 11100408 TI - Triethanolamine. PMID- 11100409 TI - N-nitrosodiethanolamine. PMID- 11100410 TI - 2,3-Dibromopropan-1-ol. PMID- 11100411 TI - 2,2-Bis(bromomethyl)propane-1,3-diol. PMID- 11100412 TI - Glycidol. PMID- 11100413 TI - Nitromethane. PMID- 11100414 TI - Pyridine. PMID- 11100415 TI - Management of the septic patient in the operating room. AB - Sepsis, severe sepsis, and septic shock represent the spectrum of physiological response to a variety of infecting pathogens. Multiple-organ dysfunction may result from widespread activation of inflammatory and antiinflammatory mechanisms. Intensive multiorgan support, effective antibiotic therapy, and eradication of the inciting source remain the cornerstones in the care of septic patients. Perioperative planning and management need to ensure the continuation of such care in addition to providing for the requirements of the given surgical procedure. PMID- 11100416 TI - Perioperative management of selected endocrine disorders. AB - Anesthesiologists routinely encounter patients with endocrine disorders. Good perioperative outcome depends on preoperative identification, risk stratification and optimization of the patients' endocrinopathies and their sequelae; intraoperative control of metabolic and physiological parameters; and appropriate postoperative pain management, stress modulation, and evaluation of neurological, cardiovascular, and renal function. PMID- 11100417 TI - Recent changes in the management of intracranial hypertension. PMID- 11100418 TI - The traumatic airway: the anesthesiologist's role in the emergency room. AB - An approach to the airway is addressed in Table 1. A summary of induction/NMB agents and doses is given on Table 2; indications for the different agents are noted on Table 3. The central pharmacological issue is not that any one drug is universally preferred over another. Rather, it is key that one develop a thoughtful rationale for the drugs used, and a plan to get out of trouble if one is suddenly in the sinking situation of a patient with a difficult airway who cannot breathe on his or her own. The backup plan might involve the use of BVM ventilation, blind digital intubation, fiberoptic bronchoscope-aided intubation, retrograde techniques, light wand intubation, laryngeal mask airway techniques, posterior pharyngeal endotracheal tube placement ventilation, or a surgical airway. Most of these approaches are reviewed elsewhere. PMID- 11100419 TI - Automated information systems in anesthesiology. AB - The current state of the art of anesthesia information systems remains primitive. Currently, available commercial systems focus only at automating the charting process and not the care process. Until systems are available that integrate these two functions, anesthesiologists will not truly benefit from such systems. PMID- 11100420 TI - How to get the most out of the Internet for your clinical practice. PMID- 11100421 TI - Operating room utilization: the need for data. AB - Keeping the OR scheduled to satisfy all the various constituents is a complex dynamic process. The health care environment needs to be carefully analyzed to ensure that the services the OR offers are appropriate. OR utilization is not simply ensuring that the greatest number of cases are done. The cost of doing these cases must be considered and, in order to do so, compromises must be made. Creating information systems that track all aspects of utilization, including costs and revenues, will be vital for the future management of operating rooms. PMID- 11100422 TI - Fluid management in trauma. PMID- 11100424 TI - Pulmonary artery catheter: what does the literature actually tell us? PMID- 11100423 TI - Transfusion in the operating room and the intensive care unit: current practice and future directions. PMID- 11100425 TI - Experiences with a compound method for estimating the time since death. I. Rectal temperature nomogram for time since death. AB - The temperature-based nomogram method for estimation of the time period since death was used at the scene of death as the primary method within a compound method in 72 consecutive cases. The situation and cooling conditions inspected and evaluated by the forensic pathologist at the scene are described as far as necessary to enable handling of the method. A comparison of the estimated period since death with the period determined by the police investigations demonstrates the reliability of the method. There were no contradictions in any of the 60 cases between the period of death estimated by this method and that determined by the police investigations. The criminal investigations were effectively supported in the earliest stages in 11 cases despite the fact that the period estimated was of considerable duration. PMID- 11100426 TI - Experiences with a compound method for estimating the time since death. II. Integration of non-temperature-based methods. AB - The period since death was estimated at the scene in 72 consecutive cases using the temperature-based nomogram method as the primary method and supplemented by examination of criteria such as lividity, rigor mortis, mechanical and electrical excitability of skeletal muscle and chemical excitability of the iris. A case oriented, computer-assisted selection of the non-temperature-based methods and integration of the results into a common result of the compound method was made following a special logistic. The limits of the period since death as estimated by the nomogram were improved in 49 cases by including the non-temperature-based methods and also provided results in 4 cases where the temperature method could not be used. In a further 6 cases the non-temperature-based methods confirmed the limits estimated by the temperature method but in 14 cases a useful result could not be obtained. In only one of the cases investigated was the upper limit of the period since death, as estimated by the criterion re-establishment of rigor (8 h post-mortem), in contradiction with the period determined by the police investigations (9.4 h post-mortem). PMID- 11100427 TI - Thickness of the air-blood tissue barrier in infants. AB - The harmonic mean barrier thickness of the alveolar air-blood tissue barrier was measured in nine SIDS cases, six cases of unnatural death (three with asphyxiation, three without asphyxiation) and six cases showing interstitial pneumonia (IP, three cases with lymphomonocyte infiltration of alveolar walls, three cases with peribronchiolar infiltration). Approximately 550-600 measurements were carried out in each case using micrographs with a final magnification of 11,000. The Th values ranged between 0.37 micron and 0.39 micron in the SIDS group, in deaths due to asphyxiation and IP with a peribronchiolar type of infiltration, were lowest in the unnatural deaths without asphyxiation (0.32 micron) and highest in cases showing IP with alveoloseptal infiltration (0.44 micron). The differences between the groups were significant (H-test, Hcor = 5.927). Compared to "normal" unnatural deaths (Th = 0.32 micron), cases with interstitial cell infiltration of the alveolar septa showed a nearly 40% increase of the barrier thickness which indicates a corresponding decrease of the diffusion capacity. A decreased diffusion capacity can cause hypoxemia which could be an additional trigger mechanism in the death process. PMID- 11100428 TI - Immunohistochemical expression of E-selectin in sepsis-induced lung injury. AB - The vascular endothelium controls leukocyte extravasation into tissue by the induction and modulation of endothelial cell adhesion molecules, such as E selectin (CD62E). E-selectin is not expressed by non-stimulated endothelium, but is activated by cytokines and initiates neutrophil recruitment in sepsis-induced lung injury. The aim of the present study was to assess the value of the immunohistochemical expression of endothelial E-selectin for the post-mortem differentiation between death due to sepsis and death due to other causes. The immunohistochemical expression of E-selectin was investigated in lung specimens obtained at autopsy from sepsis-associated fatalities (n = 6), possible sepsis associated fatalities (n = 7), non-sepsis group I (death due to unnatural causes, e.g. trauma, electrocution, drowning, hanging n = 17) and non-sepsis group II fatalities (death due to natural causes, e.g. myocardial infarction, intracerebral bleeding n = 7). E-selectin was detected in paraffin sections using the ABC technique and the expression was scored semiquantitatively by evaluating the intensity and incidence of positively stained endothelium of the interstitial pulmonary microvasculature. E-selectin was strongly expressed in all cases of the definite sepsis group, in 29% of the possible sepsis-associated fatalities and in only 4% of the cases in the non-sepsis groups I and II. In comparison to all other study groups, E-selectin expression in the definite sepsis group differed significantly (p < 0.05). Cases with inflammatory and mechanical lung tissue alterations from the control groups showed no positive immunohistochemical reaction for E-selectin; therefore, false positive results should not be expected in non-sepsis cases. Our findings suggest that the immunohistochemical detection of an intense expression of E-selectin in lung tissue may prove to be a valuable diagnostic tool in the forensic post-mortem elucidation of death due to sepsis. PMID- 11100429 TI - Spectrophotometric evaluation of the colour of intra- and subcutaneous bruises. AB - Spectrophotometric measurements were performed on intra- and/or subcutaneous bruises occurring in direct temporal connection with peracute fatal trauma. The purpose of these measurements was to determine whether the visual colour impression of a fresh traumatic extravasation can give information on the localisation of the haemorrhage in a certain tissue layer. After visual assessment of the colour of the bruise, the spectral reflectance curves and the CIE-L*a*b* colour values were determined with the help of a diode array spectrophotometer. The localisation and size of the haemorrhages in the cutis and/or subcutis were evaluated morphologically after incision of the skin. It was confirmed that there is a relationship between the colour impression and the localisation of the bruise. Bruises localised near the surface have a more reddish appearance while bruises in deeper layers give a more bluish colour impression. An explanation may be found in the optical characteristics of skin. Blood localised in the subcutis appears blue on the surface due to scattering processes in the dermis (Rayleigh scattering), as the blue wavelengths of the light are scattered (and thus reflected) to a greater extent than the red wavelengths. PMID- 11100430 TI - Gunshot wound to the head with full recovery. AB - A 28-year-old man was shot in the back of the head at close range by a robber who then locked him in a room assuming that he was dead. The man was discovered 2 days later. The entrance wound of the bullet was in the left occipital region and it passed into the periphery of the right temporal lobe, where it lodged. The man was transferred to a rehabilitation centre 3 weeks later in relatively good health with only slight general EEG changes. The mild clinical course in this case is attributable to two major factors: firstly, no important brain structures were injured, and secondly, the kinetic energy of the silver-tip hollow-point bullet was probably rather low. Three years after the incident, the man still has slight sensory disturbances in the fingers of the left hand and left-sided homonymous hemianopia. He is now working again at his old profession (managing director) and the projectile is still lodged in the right temporal lobe. PMID- 11100431 TI - Errors in ABO typing of blood stains using PCR. AB - Genotypes of the ABO blood group system were investigated using a multiplex PCR and subsequent restriction enzyme digestion on experimental blood stains. Differences were found when typing blood between the PCR and serological methods and one blood sample, typed as B with the agglutination test was classified as AB using the method described here. The subsequent sequencing procedure revealed the genotype to be BB. Methodological causes for errors in typing which should be taken into consideration are discussed. PMID- 11100432 TI - [Value of molecular biology in the identification of trypanosomes responsible for African trypanosomiasis or sleeping sickness]. PMID- 11100433 TI - [Laos: a landlocked country]. PMID- 11100434 TI - [Can carob powder be used with oral rehydration solutions for the treatment of acute diarrhea?]. PMID- 11100435 TI - [Benzathine penicillin]. PMID- 11100436 TI - [Tropical medicine on the Internet]. PMID- 11100437 TI - [Sleeping sickness, again and always]. PMID- 11100438 TI - [Chlamydia infection and peripartum dilated cardiomyopathy in Niger]. AB - Peripartum cardiac failure due to cardiomyopathy is common in sub-saharan Africa. The etiology is unknown. This study was performed in Niger to assess a possible relationship between peripartum cardiomyopathy and Chlamydia. A total of 50 African women presenting peripartum cardiomyopathy underwent testing for infection by Chlamydia pneumoniae, Chlamydia trachomatis, and Chlamydia psittaci. The inclusion criteria were cardiac failure during the last three months of pregnancy or first 6 months postpartum with echocardiographic evidence of dilated cardiomyopathy. Similar testing was carried out in a control group of 25 African women from the same geographical location without cardiac disease. Detection of specific IgG, IgA and IgM antibodies was performed using the microimmunofluorescence technique. The cut-off values were > or = 1/32 for specific IgG antibody and > or = 1/16 for specific IgA and IgM antibody. Statistical comparison of the patient and control groups was achieved using the chi 2 test. For Chlamydia pneumoniae, 48 patients (96 p. 100) versus 20 controls (80 p. 100) controls were positive for IgG antibodies (p < 0.025) and 39 patients (80 p. 100) versus 14 controls (56 p. 100) were positive for IgA antibodies (p < 0.05). No patient or control demonstrated IgM antibodies for Chlamydia pneumoniae. For Chlamydia trachomatis and Chlamydia psittaci, differences in positive rates were not statistically significant. This is the first study demonstrating infection in patients with peripartum cardiomyopathy. The possible role of Chlamydia pneumoniae is discussed. PMID- 11100439 TI - [Bancroftian filariasis in Madagascar: persistent endemicity]. AB - A major study was conducted to determine the prevalence of Bancroftian filariasis in 9 health districts located mainly on the east and north coast of Madagascar between 1995 and 1997. The study population included 2524 people 10 years or older. On the east and north coast, the incidence of microfilarial carriers varied depending on location from 7 p. 100 to 47 p. 100 in men and 3 p. 100 to 33 p. 100 in women. The highest incidences, i.e., around 33 p. 100 in both sexes, were observed in the southeastern districts of Ifanadiana, Manakara, and Vangaindrano. In the other districts on the east coast, the highest rates occurred mainly in men, i.e., 47 p. 100 in Vavatenina, 33 p. 100 in East Feneriva, and 33 p. 100 in Mahanoro. Only two districts on the west coast were studied, i.e., Marovoay where the incidence was zero and Ankazoabo where the prevalence was 4 p. 100 for men and 3 p. 100 for women. The results are compared with those of a study carried out in 1958. At 16.22 p. 100, chronic morbidity is relatively common in men but less disabling, i.e. mainly scrotal and member elephantiasis and hydroceles. Chronic morbidity was only 2.26 p. 100 in women, i.e. mainly member elephantiasis. Control of Bancroftian filariasis may be achievable by targeted use of drug prophylaxis and bednets in zones of high prevalence. PMID- 11100440 TI - [Seroprevalence of viral hepatitis B in the city of Mahajanga, Madagascar in 1999]. AB - A seroepidemiological study was carried out to assess the prevalence of hepatitis B virus (HBV) in the city of Mahajanga, Madagascar in July 1999. A total of 654 serum samples were collected from randomly selected subjects over the age of 2 years. The ELISA technique was used to detect serum markers of HBV infection. Prevalence rates were 14.2 p. 100 for HBs, 1.4 p. 100 for HBe antigens, and 49.5 p. 100 for HBV infection as defined by the presence of at least one serum markers. HBs antigens were detected in 20.8 p. 100 of children between the ages of 2 and 4 years and 15.3 p. 100 of women of childbearing age. In the latter age group, 5.6 p. 100 demonstrated HBe antigens, which are considered as a replication marker. The findings of this study are in agreement with those of a previous study in Madagascar and indicate that an expanded program of immunization against hepatitis B virus is warranted. PMID- 11100441 TI - [Onchocerciasis and epilepsy. Epidemiological survey in Mali]. AB - A door-to-door survey was conducted in 18 villages in Mali with a total of 5,243 inhabitants classified according to the endemicity of onchocerciasis. Each epileptic was matched with two controls. The survey protocol included the following steps in cases and controls: census taking, socioeconomic data, screening for epilepsy, clinical examination, laboratory testing to detect parasites in stools and urine, and snip-test. The crude prevalence of epilepsy was 13.35 per 1,000 (n = 70). Epidemiological study provided a number of valuable demographic insights concerning age at onset, type of seizure activity during seizure and personal and family medical history. A transverse study showed that the prevalence of epilepsy was not significantly higher (p = 9.09) in zones of high endemicity of onchocerciasis (16.1 per 1000) than in zones of low endemicity (10.8 per 1000). Case-control findings showed evidence of onchocerciasis in 22.4 p. 100 of epileptics and 21.7 p. 100 of controls (odds ratio = 1.02 IC 95 p. 100: 0.4-2.19, not significant). Various risk factors including genetic factors and low socio-economic status could explain the trend toward a higher incidence of epilepsy as well as higher morbidity rates in zones of high endemicity of onchocerciasis. PMID- 11100442 TI - [Blackwater fever in adults with sickle cell anemia. Two fatal cases]. AB - Blackwater fever is characterized by severe intravascular hemolysis with renal failure caused by recurrent use of quinine for prophylaxis. Once described in European patients, sporadic cases have been reported more and more often in autochthonous Africans and Asians. Newer antimalarials including aminoalchohol mefloquine, and halofantrine have also been implicated in Blackwater fever. In this report we describe two cases of blackwater fever involving patients with sickle cell anemia (HbSS). Symptoms including fever, acute hemolytic anemia, emesis, back pain, and hemoglobinuria were characteristic of blackwater fever. Both patients died. Although the underlying mechanism of blackwater fever remains unclear, a likely explanation is an immunoallergic reaction to quinine. Association with glucose-6-phosphate dehydrogenase deficiency has often been reported. Our cases suggest that blackwater fever may also be correlated with hemoglobinopathy such as HbSS. PMID- 11100443 TI - [Natural infestation of Lymnaea truncatula Muller by Fasciola hepatica in the Tozeur oasis in southwest Tunisia]. AB - Natural infestation of Lymnaea truncatula by Fasciola hepatica was studied at various sites in the traditional oasis of Tozeur 8 times between September 1997 and August 1998. Infestation of snails was documented in five of the eight sites with a mean level of 26.1 p. 100. Level of infestation varied depending on the site. It was highest in secondary irrigation canals (32.8 p. 100), seguias, (34.8 p. 100), and secondary drainage canals (34.8 p. 100). Lower levels were observed in the main drainage canal, (15.2 p. 100). Two infestation periods were noted at two sites in relation with spreading of manure by farmers in autumn and spring. Perennial infestation due to the permanent presence of animal hosts was noted at one. Infestation of drainage canals is enhanced by incoming parasite eggs from upstream irrigation canals. Infestation was age dependent reaching up to 100 p. 100 in older snails of 5.5 mm. Environmental factors such as climatic conditions and human intervention are important to sustaining snail infestation in the oasis of Tozeur. PMID- 11100444 TI - [Parasitologic survey of schistosomiasis due to Schistosoma mansoni in Katana, Democratic Republic of Congo]. AB - It is now widely recognized that development of irrigation projects in endemic areas for schistosomiasis invariably leads to a recrudescence of the disease by increasing the habitat of the intermediate snail host of Schistosoma mansoni. This fatality was again demonstrated by experience in the Katana region of the Democratic Republic of Congo where development of 43 fresh water reservoirs for raising Tilapia nilotica led to multiplication of Biomphalaria pfeifferi. A parasitological study was conducted in three new villages around these basins and in neighboring villages. Stool examinations were performed in a total of 787 people. Infestation rates were 8.1 p. 100 and 4 p. 100 respectively. Infestation exceeded 25 p. 100 in children between the ages of 10 and 14 years. These findings underline the need for preventive measures. PMID- 11100445 TI - [Hepatitis B, C, and E in New Caledonia. Seroepidemiologic study in military recruits]. AB - A seroepidemiological study in army conscripts was carried out to collect data on hepatitis B, C, and E in New Caledonia. All young men recruited between October 1998 and June 1999 (n = 351) were retrospectively included in study. Anamnestic data was obtained during the induction physical examination. Blood tests to detect viral markers and assessment of liver function were performed in all cases. The incidence of hepatitis B was 6.6 p. 100 of chronic carriers of HB antigens. The overall number of vaccinated subjects was low, i.e., 17.9 p. 100. The incidence of carriers presenting at least anti-HBc antibodies was higher in persons of Melanesian and Wallisian extraction, i.e. 59.5 p. 100 and 49.2 p. 100 respectively. This is logical since vaccination rates in these ethnic groups were lower. The incidence was also higher in the northern province and islands, i.e., 48.7 p. 100 and 75 p. 100. Mention of a family history of hepatitis B was a significant predictor of infection. No case of hepatitis C was found. Six carriers of anti-HVE were identified including three who had never left New Caledonia. Detection tests for viral RNA were negative in all cases. This study confirms the high incidence of hepatitis B in New Caledonia and the need for mass vaccination. Findings also suggest that the area may still be free of hepatitis C but the presence of hepatitis E cannot be ruled out. PMID- 11100447 TI - [Nutrition, urbanization and poverty in subsaharan Africa]. AB - Africa is currently the continent with the highest urbanization rate in the world. This demographic upheaval has sometimes been considered as an opportunity for modernization but as early as the 1980s experts called attention to its potential impact on nutrition. In recent decades, economic problems and structural reforms have had dire effects on urban populations. Today increasing poverty and the effects of globalization have revived concerns about urban nutrition. Retarded growth and emaciation are less common than in urban areas than in rural areas, but disparities between the rich and poor are much greater. However, in some cities, the incidence of emaciation progressed the more during the 1990s, and now equals that in rural areas. In cities the level of obesity in adult women is a sign of nutritional transition but emaciation has also increased. Despite the wide variety of urban conditions, analysis of the underlying factors reveals several constants. For most of these factors, the characteristic feature of the urban environment is a further increase of social inequality. PMID- 11100446 TI - [Dangerous sharks in tropical seas]. AB - Sightseeing travel in tropical zones is a growing industry. The risks incurred by travelers depend on the destination, duration of stay, individual behavior, and type of leisure activity. Water sports expose visitors to encounters with dangerous marine animals. Shark attacks are rare but always serious occurrences. Divers should handle any shark, regardless of size, with due precaution. Prevention of shark attack depends on avoiding encounters by not attracting the attention of the shark and knowing the proper attitude to adopt in case an encounter should occur. Active and passive protection can be used, but each method has advantages and disadvantages depending on the situation. Rescue operations are difficult due to the gravity of injuries and their occurrence in a marine environment. This along with the nature of the aggressor explain that many attacks are immediately fatal. Wounds are often deep with involvement of bone, blood vessels, and nerves. A possible source of complication in survivors is infection, which can involve uncommon microorganisms associated with bacteria in sharks mouth or marine environment. PMID- 11100448 TI - [Judicial proceedings involving sexual abuse of minors in Cameroon]. AB - The purpose of this study was to evaluate how courts in Cameroon treat cases involving sexual abuse on minors by comparing the incidence of hospital examinations and legal proceedings for sexual abuse. This retrospective study is based on a review of public records at the Yaounde Court of Justice covering the period from October 1, 1994 to January 6, 1999. Of the 2345 criminal cases recorded during the study period, 224 involved sexual abuse on minors under the age of 16 years (9.5 p. 100 of cases). The victims were all female ranging from 3 to 15 years of age (mean, 9 years) with a peak incidence between 10 and 15 years (70 p. 100). All offenders were adults between 21 and 50 years of age (mean, 30 years) at the time of the crime. This study showed that the incidence of court proceedings for sexual abuse is higher than that of hospital examinations for sexual abuse. Most offenders convicted of sexual abuse on minors received long prison sentences, i.e. 15 years or more. No reconciliatory action or mediation was initiated by the court or third parties. PMID- 11100449 TI - [Statistical analysis in the evaluation of controlled diffusion of iodine in the water in developing countries]. AB - This report describes the methodology and findings of a novel statistical technique for evaluation of the efficacy of the Rhodifuse Iode system in prevention of endemic goiter in Mali. The system involves continuous release of iodine in ground water used for drinking. Study was carried out in four villages for one year. The iodine release system was used in three villages. The fourth village served as the control. The incidence of goiter graded using the criteria of the WHO was assessed in each village according to sex. Statistical analysis consisted of correlating goiter grade with four predictors, i.e., village, sex, iodine release, and time. Since goiter grade is a dependent variable, its law of probability was modeled in function of the predictors. The Cat Mot procedure included in the SAS software package allowed both definition of the law of probability of grade of goiter and its transformation in function of predictor. The generalized linear model was obtained by either the generalized least square method or greatest likelihood method. The Proc Catmod procedure was then used to generalize analysis of variance in case of a nominal or ordinal, binary or polytomic response. The results of this novel statistical technique suggested that the Rhodifuse Iode system was effective. PMID- 11100450 TI - [Bacteriological particularities of Vibrio cholerae serotype Ogawa, biotype El Tor isolated in Burundi]. PMID- 11100451 TI - [Cold pre-sternal tuberculosis abscess: a little known form of tuberculosis]. PMID- 11100452 TI - [Male behavior with modern contraception methods in northern Togo]. PMID- 11100453 TI - [Modulator genes in a monogenic disease]. PMID- 11100454 TI - [Viral hemorrhagic fevers: history and lessons from the last forty years]. PMID- 11100455 TI - Lassa fever: immuno-epidemiological approach to the study of an endemic viral haemorrhagic fever. PMID- 11100456 TI - [Lessons from the Marburg virus epidemic in Durba, Democratic Republic of the Congo (1998-2000)]. PMID- 11100457 TI - [Impact of veterinary arboviruses. The case of Rift Valley Fever]. PMID- 11100458 TI - Realities of viral haemorrhagic fevers in Africa. PMID- 11100459 TI - [The arenavirus in the New World: a virus in evolution and emerging diseases]. PMID- 11100460 TI - [Ebola virus and virus reservoirs]. PMID- 11100461 TI - [Malaria chemoprophylaxis in French and foreign armies]. PMID- 11100462 TI - [Control of schistosomiasis: realities and futurology]. PMID- 11100463 TI - [Realities of human trypanosomiasis]. PMID- 11100464 TI - Protein kinase-mediated signaling networks. Regulation and functional characterization. PMID- 11100465 TI - Cloning protein tyrosine kinases by screening cDNA libraries with antiphosphotyrosine antibodies. PMID- 11100466 TI - Immunoprecipitation and western blotting of phosphotyrosine-containing proteins. PMID- 11100467 TI - Two-dimensional phosphoamino acid analysis. PMID- 11100468 TI - Two-dimensional phosphopeptide mapping of receptor tyrosine kinases. PMID- 11100469 TI - Identification of the sites of phosphorylation in proteins using high performance liquid chromatography and mass spectrometry. PMID- 11100470 TI - Phosphorylation of Smad signaling proteins by receptor serine/threonine kinases. PMID- 11100471 TI - Assays for mitogen-activated protein kinase (MAPK) subtypes and MAPK activating protein kinase-2 (MAPKAP K-2) using a common cell lysate. PMID- 11100472 TI - JAK-mediated phosphorylation and activation of STAT signaling proteins. Analysis by phosphotyrosine blotting and EMSA. PMID- 11100473 TI - Assays for glycogen synthase kinase-3 (GSK-3). PMID- 11100474 TI - Cyclin-dependent kinases and cyclin-dependent kinase inhibitors. Detection methods and activity measurements. PMID- 11100475 TI - Protein histidine kinase. PMID- 11100476 TI - Cloning and expression of recombinant HEXA-HIS tagged ZAP-70 using the baculovirus system. PMID- 11100477 TI - Analysis of protein kinase subcellular localization by visualization of GFP fusion proteins. PMID- 11100478 TI - Detection of phosphorylation-dependent interactions by far-western gel overlay. PMID- 11100479 TI - Purification of tyrosine-phosphorylated proteins by immunoaffinity chromatography and direct cloning of their cDNAs from bacterial expression libraries. PMID- 11100480 TI - Identification of receptor tyrosine kinase (RTK) substrates by the cloning of receptor targets (CORT) strategy. PMID- 11100481 TI - Identification of receptor tyrosine kinase associating proteins using the yeast two-hybrid system. PMID- 11100483 TI - Analysis of SH2 domain--phosphopeptide interactions by isothermal titration calorimetry and surface plasmon resonance. PMID- 11100482 TI - cDNA expression cloning and characterization of phosphorylation dependent protein interactors using the yeast tribrid system. PMID- 11100484 TI - Cloning and characterization of RTK ligands using receptor-alkaline phosphatase fusion proteins. PMID- 11100485 TI - Isolation and characterization of "orphan-RTK" ligands using an integrated biosensor approach. PMID- 11100486 TI - Imaging of the brain in the HIV-positive child. AB - The prevalence of human immune-deficiency virus (HIV) infection around the world, coupled with increasing population movement, make it likely that many physicians will treat HIV-infected patients. New treatment protocols for the specific manifestations of acquired immune-deficiency syndrome (AIDS) make distinguishing the different neurological diseases of great importance. The pattern of disease in children differs from those of adults both in its distribution and etiology. This article encapsulates the salient aspects relating to the imaging of the brain in HIV-positive children, paying particular attention to recent advances and the different features of the various pathological conditions affecting the HIV-infected brain in children. PMID- 11100487 TI - Transcranial Doppler (TCD) screening for stroke prevention in sickle cell anemia: pitfalls in technique variation. AB - BACKGROUND: The Stroke Prevention Trial in Sickle Cell Anemia (STOP) identified children as being at high stroke risk if the time-averaged maximum mean velocity (TAMMV) of the middle cerebral or intracranial internal carotid arteries measured > or = 200 cm/s. These values were obtained utilizing a 2-mHz dedicated nonimaging pulsed Doppler technique (TCD) and manual measurements. Questions have been raised as to the comparability of results obtained with different ultrasound machines and measurement techniques. OBJECTIVE: The purpose of this study was to compare nonimaging (TCD) and transcranial duplex imaging (TCDI) findings in children potentially at risk for stroke with sickle cell disease. MATERIALS AND METHODS: Twenty-two children with sickle cell disease and no history of stroke were evaluated by both TCD and TCDI. Examinations were performed on the same day without knowledge of the other modality results and read independently using manually obtained measurements. Mean velocities, peak systolic velocities, and end diastolic velocities obtained by the two techniques were compared. In a subgroup, manual measurements were compared to electronically obtained measurements. RESULTS: TCDI values were lower than TCD measurements for all vessels. TCDI TAMMV values were most similar to the TCD values in the middle cerebral artery (-9.0%) and distal internal cerebral artery (-10.8%), with greater variability in the anterior cerebral artery (-19.3%), bifurcation ( 16.3%), and basilar arteries (-23.1%). Risk group placement based on middle cerebral artery TAMMV values did not change when comparing the two techniques. Measurements obtained electronically were lower than those obtained manually. CONCLUSION: Velocities obtained by TCDI may be lower than TCD measurements, and these differences should be taken into consideration when performing screening for stroke risk and selection for prophylactic transfusion based on the STOP protocol. PMID- 11100488 TI - Changes in echogenicity of spinal subarachnoid space associated with intracranial hemorrhage: new observations. AB - PURPOSE: The role of subarachnoid blood and secondary, sterile inflammation in the pathogenesis of posthemorrhagic hydrocephalus (PHH) is not well understood. The aims of this study were to study the frequency and rate of spread of blood into the spinal subarachnoid space (SSS) and to evaluate the relationship of this finding and PHH. MATERIALS AND METHODS: Nine premature babies with major intracerebral hemorrhage (ICH, grade 3 or higher), and ten premature infants with minor ICH (grade 1) or no evidence of ICH (control group) were identified and underwent serial cranial and spinal sonography at the time of initial diagnosis, 12-24 h after the ICH and weekly thereafter for at least 9 weeks. Sagittal and axial scans of the thoracolumbar spine were obtained and evaluated for the presence of echogenic debris in the dorsal SSS. Six additional patients who had cranial and spinal sonography died within the 1st week of life and underwent post mortem examinations. RESULTS: The SSS was echo-free (normal) in all cases at the time of initial sonographic diagnosis of ICH. Within 24 h, all babies with major ICH had developed increased echogenicity of the cervical and thoracic SSS. Echogenicity of the SSS decreased gradually over several weeks. Although transient ventricular dilatation was present in every patient, only one patient had rapidly progressive PHH requiring shunt placement. Transient cysts of the cervicothoracic subarachnoid space were identified in two patients 6-7 weeks after ICH. The subarachnoid space remained echo-free in all control infants At autopsy, all four infants with echogenic spinal debris had blood or blood products in the spinal subarachnoid space, whereas two infants with echo-free spinal images did not. CONCLUSIONS: Spread of blood from the ventricular system into the spinal subarachnoid space after ICH is common and can be seen within 24 h of initial ICH. Subarachnoid blood is associated with post-hemorrhagic ventricular dilatation and transient spinal subarachnoid cyst formation. PMID- 11100489 TI - Intracranial venous anomalies associated with atretic cephalocoeles. AB - BACKGROUND: Midline scalp lesions are frequent in children. They include soft tissue masses and atretic meningocoeles. Their recognition is important as their treatment differs. Intracranial venous anomalies are known to be associated with atretic cephalocoeles. MATERIALS AND METHODS: A retrospective study was undertaken to assess the frequency of intracranial venous anomalies associated with atretic meningocoeles (AT). Thirty-one patients with AT were studied by MRI. There were 13 meningocoeles and 14 encephalocoeles; 4 have not yet received surgery. RESULTS: Venous anomalies were found when the cephalocoeles lay above the torcular. They include absence of the straight sinus and duplication of the longitudinal sinus. CONCLUSION: Venous anomalies are frequent in atretic cephalocoeles and are part of the dysraphic state. PMID- 11100490 TI - Periventricular nodular heterotopia in patients with filamin-1 gene mutations: neuroimaging findings. AB - BACKGROUND: The filamin-1 (FLN-1) gene is responsible for periventricular nodular heterotopia (PNH), which is an X-linked dominant neuronal migration disorder. OBJECTIVE: To review the clinical and imaging findings in a series of patients with documented filamin-1 mutations. MATERIALS AND METHODS: A retrospective review of the medical records and MR studies of a series of patients with PNH and confirmed FLN-1 mutations was done. There were 16 female patients (age range: .67 71 years; mean = 28.6) with filamin-1 gene mutations. RESULTS: In six of the patients the same mutation was inherited in four generations in one pedigree. In a second pedigree, a distinct mutation was found in two patients in two generations. In a third pedigree, a third mutation was found in four patients in two generations. The remaining four patients had sporadic de novo mutations that were not present in the parents. Ten patients had seizures, and all patients had normal intelligence. In all 16 patients MR demonstrated bilateral near-continuous PNH. There were no consistent radiographic or clinical differences between patients carrying different mutations. CONCLUSION: Patients with confirmed FLN-1 gene mutations are usually female and have a distinctive MR pattern of PNH. Other female patients with this same MR pattern probably harbor FLN-1 mutations and risk transmission to their progeny. This information is important for genetic counseling. PMID- 11100491 TI - MR of physeal fractures of the adolescent knee. AB - The aim of this study was to assess physeal fractures of the pediatric knee identified by MR imaging and to describe the MR findings of such fractures. The authors reviewed 315 consecutive pediatric knee MR examinations done to assess for traumatic injury. The MR images were reviewed for evidence of physeal fracture. Fractures were classified by the Salter-Harris system, and associated findings and injuries were noted. Plain radiographs and medical records were reviewed. Seven distal femoral physeal fractures (Salter II, n = 6; Salter III, n = 1) and two proximal tibia physeal fractures (Salter III, n = 1; complex Salter IV, n = 1) were identified. Magnetic resonance demonstrated widening of a portion of the physis with visualization of a metaphyseal/epiphyseal fracture line. Periosteal elevation was observed in six cases. Four patients had associated ligamentous or meniscal injuries. Plain radiographs were available for review in eight patients. Bone abnormalities suggesting fracture were evident in six of eight patients; however, the fracture was fully delineated in only one patient. The diagnosis or confirmation of fracture by MR changed clinical management in seven of eight patients in whom follow-up was available. Physeal fractures of the pediatric knee are occasionally diagnosed by MR. Magnetic resonance provides improved delineation of non-displaced physeal fractures of the knee, while simultaneously allowing for evaluation of soft tissue structures. PMID- 11100492 TI - MR imaging evaluation of subacute and chronic bone abscesses in children. AB - OBJECTIVE: The aim of the present study was to assess the value of magnetic resonance (MR) imaging in subacute and chronic bone abscesses in children. MATERIALS AND METHODS: Seventy-four patients underwent MR imaging because of suspected musculoskeletal infections between January 1996 and January 1999 in Montreal Children's Hospital. The clinical, radiographic, scintigraphic and MR imaging features of patients with a bone abscess were studied. RESULTS: Eleven patients had osteomyelitis with no bone abscess and six had osteomyelitis with a subacute or chronic bone abscess. Although the lucency was eventually seen on plain radiographs in all cases, MR imaging made a significant contribution, as it helped narrow the differential diagnosis and showed better delineated medullary involvement and extension into the epiphysis. CONCLUSION: MR imaging is valuable in the diagnostic evaluation of children with bone infection and abscess. It reveals the extent of subperiosteal and epiphyseal involvement not seen on plain radiographs. The extent of the medullary involvement around the abscess is best visualized with MR imaging, which can also distinguish between isolated soft tissue infection adjacent to bone and true bone infection. PMID- 11100493 TI - Role of embolisation in the treatment of bronchopulmonary sequestration. AB - BACKGROUND: Sequestrations represent bronchopulmonary malformations that are increasingly diagnosed antenatally. After birth, the therapeutic approach in asymptomatic children is debated, as some may spontaneously regress. OBJECTIVE: To evaluate the efficacy of embolisation of the feeding systemic artery in the treatment of bronchopulmonary sequestration. MATERIALS AND METHODS: Sixteen children with bronchopulmonary sequestration were treated by endovascular embolisation of the feeding systemic artery. RESULTS: Ten patients were considered cured by embolisation alone. One patient was operated on after unsuccessful embolisation, three had partial regression of the lung mass and two are still under follow-up. CONCLUSIONS: Our experience indicates that bronchopulmonary sequestrations in children can be treated by embolisation alone. PMID- 11100494 TI - Intralobar bronchopulmonary sequestration evaluated by contrast-enhanced three dimensional MR angiography. AB - Bronchopulmonary sequestration (PS) is characterized by non-functioning lung tissue fed from one or several aberrant systemic arteries. The condition is diagnosed by visualizing the feeding arteries using non-invasive CT, MRI, colour Doppler sonography or conventional angiography. We present a 5-year-old boy in whom intralobar sequestration was diagnosed using contrast-enhanced 3D MR angiography, which visualised fine blood vessels in the thoraco-abdominal region without arterial puncture. This technique is useful for diagnosing PS. PMID- 11100495 TI - Malignant pleural mesothelioma in a 13-year-old girl. AB - Pleural mesothelioma is an uncommon tumor in all age groups, but is especially rare in childhood. We describe the clinical and radiological features of malignant pleural mesothelioma in a 13-year-old girl. The chest radiograph showed nearly complete opacification and loss of volume in the left hemithorax. Computed tomography demonstrated a large pleural effusion centrally surrounded by a thick enhancing rind of soft tissue. The radiological features of childhood pleural mesothelioma in our case were similar to those described in adults with this disease. PMID- 11100496 TI - Intrahepatic chemoembolization in unresectable pediatric liver malignancies. AB - OBJECTIVE: To determine the effectiveness of a new multidisciplinary approach using neoadjuvant intrahepatic chemoembolization (IHCE) and liver transplant (OLTx) in patients with unresectable hepatic tumors who have failed systemic chemotherapy. MATERIALS AND METHODS: From November 1989 to April 1998, 14 children (2-15 years old) were treated with 50 courses of intra-arterial chemotherapy. Baseline and post-treatment contrast-enhanced CT and alpha fetoprotein levels were performed. Seven had hepatoblastoma, and 7 had hepatocellular carcinoma (1 fibrolamellar variant). All patients had subselective hepatic angiography and infusion of cisplatin and/or adriamycin (36 courses were followed by gelfoam embolization). The procedure was repeated every 3-4 weeks based on hepatic function and patency of the hepatic artery. RESULTS: Six of 14 children received orthotopic liver transplants (31 courses of IHC). Pretransplant, 3 of 6 showed a significant decrease in alpha-fetoprotein, while only 1 demonstrated a significant further reduction in tumor size). Three of 6 patients are disease free at this time. Three of 6 patients died of metastatic tumor 6, 38, and 58 months, respectively post-transplant. One of 14 is currently undergoing treatment, has demonstrated a positive response, and is awaiting OLTx. Three of 14 withdrew from the program and died. Four of 14 patients developed an increase in tumor size, developed metastatic disease, and were not transplant candidates. Two hepatic arteries thrombosed, and one child had a small sealed-off gastric ulcer as complications of intrahepatic chemoembolization. CONCLUSION: The results of intrahepatic chemoembolization are promising and suggest that some children who do not respond to systemic therapy can be eventually cured by a combination of intrahepatic chemoembolization orthotopic liver transplant. Alpha fetoprotein and cross-sectional imaging appear to be complementary in evaluating tumor response. IHCE does not appear to convert an anatomically unresectable lesion to a candidate for partial hepatectomy. PMID- 11100497 TI - Benign pneumatosis in children. AB - BACKGROUND: In pediatrics, pneumatosis intestinalis (PI) is usually due to necrotizing enterocolitis in premature newborns. Beyond infancy, PI is uncommon. "Benign pneumatosis" is PI in patients with few or no symptoms that resolves with conservative management. OBJECTIVE: Our goal was to better characterize benign PI in children. Our investigation focused on identifying underlying risk factors, symptoms at time of diagnosis, management and outcome. MATERIALS AND METHODS: Available medical records and radiographs of children with pneumatosis intestinalis from 1990 to 1998 were reviewed for underlying conditions, symptoms at time of radiographs, management and outcome. RESULTS: Thirty-seven children (mean age 4 years) were included. Thirty-two children had identifiable risk factors. Twenty-five children were immunocompromised by their underlying conditions or therapeutic regimen. Thirty-five children were managed conservatively with resolution of PI. Two patients, however, required surgery and one patient died. CONCLUSION: Benign pneumatosis does occur in children. The majority have underlying risk factors, most commonly related to immunosuppression. Clinical deterioration is the most useful indicator for surgical intervention. In most patients PI resolves with conservative management. PMID- 11100498 TI - Hepatoblastoma in a neonate: a hypervascular presentation mimicking hemangioendothelioma. AB - Congenital heart failure in the neonate supported by classic imaging findings may allow the implementation of medical therapy for presumed hemangioendothelioma without obtaining a tissue diagnosis. This case report describes a neonate with these classic clinical and radiographic findings but who underwent surgery for failing medical treatment and was diagnosed as having a hepatoblastoma by pathology. This case supports the need to obtain tissue confirmation before beginning medical therapy. PMID- 11100499 TI - Blunt injury of the abdomen: a plea for CT. AB - A 3-year-old boy was brought to the emergency unit 1 h following a deceleration injury. On clinical examination there were no signs of injury and US showed only free intraperitoneal fluid. The following morning, contrast-enhanced CT showed the right kidney did not enhance and delayed scans showed contrast medium in the renal vein. This is an indirect sign of post-traumatic renal artery occlusion. Failure to recognise this sign may have disastrous consequences in a patient with solitary kidney or bilateral renal artery occlusion. Contrast-enhanced CT scan remains the most widely available investigation for accurate staging of blunt renal trauma. PMID- 11100500 TI - Inflammatory pseudotumour of the pancreas in a child. AB - We describe a 4-year-old girl with an inflammatory pseudotumour of the pancreas, which was preceded by varicella-zoster infection. Inflammatory pseudotumour may involve a variety of tissues, the lungs and liver being typical sites of predilection. Imaging and laboratory tests are nonspecific, and for this reason the diagnosis of inflammatory pseudotumour is rarely made prior to surgery. These benign but locally aggressive masses simulate malignancy in the majority of cases. Inflammatory pseudotumour should, therefore, be considered when a mass arises in an unusual location in the paediatric age group. PMID- 11100501 TI - Trends in pediatric radiology in the United States. AB - Long-term trends in pediatric radiology in the United States--in the radiologic care of children, in research, in education and career development, in the Society for Pediatric Radiology, and in the field as a whole--were listed in 1995. The present article attempts to bring those trends up to date for the year 2000 and also describes trends not identified in the earlier report. PMID- 11100502 TI - Progressive liver calcifications in neonatal coxsackievirus infection. PMID- 11100503 TI - MR imaging recognition of nasopharyngeal gauze retention in a child. PMID- 11100504 TI - Policy processes--ways to improve health. PMID- 11100505 TI - Staying the course: how to stay optimistic in challenging times. PMID- 11100507 TI - 5th Global Conference on Health Promotion, Mexico 2000. PMID- 11100506 TI - The future of work and health. PMID- 11100508 TI - Effects on attitudes, knowledge, intentions and behaviour of an AIDS prevention programme targeting secondary school adolescents. PMID- 11100509 TI - Health education promotion practice in Zimbabwe. PMID- 11100510 TI - Creating a CD-ROM for health education. PMID- 11100511 TI - [Pediatric kidney transplantation and living donors--invaluable by virtue of necessity]. AB - Renal transplantation is the treatment of choice for paediatric patients with end stage renal failure. Living donor transplantation (LDT) has become an important therapeutic option due to the shortage of cadaver donors and increasingly long waiting times. METHODS: Between 1992 and 1999, a total of 48 paediatric and adolescent patients underwent renal transplantation in Zurich. Of these, 21 patients (44%) received a kidney from a living related donor. 11 patients had been dialysed before LDT over a period of 0.2-5.7 years (median 0.6), and 10 were transplanted preemptively. Triple immunosuppression consisted of cyclosporine A, azathioprine or mycophenolate mofetil (MMF; since 1998), and prednisone. The observation period was 0.5-7.3 years (median 2). RESULTS: Recipients were 2-18 (median 10.5) years old at transplantation. One third had either a congenital malformation, an inherited disease, or an acquired disorder. One patient died of an associated cardiac disease at 4 months with functioning graft, and one functional graft loss occurred after 2.8 years. 9 patients were switched from cyclosporine to tacrolimus, 7 for biopsy-proven rejection and 2 for cosmetic reasons (hypertrichosis). No antibody preparations were used. Median glomerular filtration rate (51Cr-EDTA), measured after one year in 11 donor/recipients, was 64 (55-95) and 54 (32-82) ml/min/1.73 m2, respectively. The most recent estimated renal function (Schwartz formula) of 19 functioning grafts was 37-79 ml/min/1.73 m2 (median 63). Median body height of 16 patients with no associated extrarenal disease was -0.9 SDS (standard deviation score); the remaining 3--with serious extra-renal disease--were considerably growth retarded. Main complications were reversible rejection episodes in 19 (90%), arterial hypertension (16), CMV disease (2) and asymptomatic CMV infection (3), pyelonephritis (3), and recurrence of the primary renal disease, seizures, diabetes mellitus and non compliance (one each). Actuarial patient and graft survival (Kaplan-Meier) after 3 years was 95 and 83% respectively. This was not statistically different from the cadaveric donor group (n = 27) with 100 and 80% survival respectively. Overall rehabilitation was excellent. The donors were 12 mothers, 8 fathers and one grandmother aged 31 to 50 (median 39) years; none of them experienced serious postoperative problems. CONCLUSIONS: The paediatric transplantation programme would no longer be feasible in Switzerland without LDT. The results are very encouraging; preemptive transplantation makes it possible to avoid dialysis in half of the patients. The risk for the donor is small, and careful evaluation without putting pressure on the family is essential. PMID- 11100512 TI - [CVID (common variable immunodeficiency): heterogeneous clinical manifestation of the commonest symptomatic primary immunodeficiency disease]. AB - Common variable immunodeficiency is the most common symptomatic primary immunodeficiency disease. The patients typically present with a long history of respiratory tract infections, sometimes sarcoid-like lesions and in rare cases boils. Heterogeneity of initial clinical manifestations as well as insufficient knowledge of the syndrome often delay the diagnosis. However, early therapy is important to reduce infections and in particular the development of bronchiectasis. Documenting 19 cases, we discuss initial clinical manifestations, some clinical complications, diagnostic procedures and therapeutic management. PMID- 11100513 TI - [Generalized intestinal CMV infection with protein-losing syndrome in ulcerative colitis]. AB - Infection by cytomegalovirus (CMV) in immunocompetent patients is rare, and if it occurs it is most often associated with ulcerative colitis. This case illustrates a CMV infection in a patient with an ulcerative colitis combined with CMV-induced protein losing enteropathy, a condition reported in immunocompetent individuals in only a very few cases worldwide. It demonstrates the importance of differentiating between a flare-up of ulcerative colitis and CMV colitis. The indication for antiviral therapy is discussed. A 76-years-old patient with a 23 year history of leftsided ulcerative colitis presented with acute pancolitis sparing the rectum. He showed no evidence of impaired host defence, nor has he ever had taken immunosuppressive drugs. Disseminated primary CMV infection involving of the colon, the oesophagus and the small intestine with protein losing enteropathy was diagnosed on the basis of histology, culture and serology. In view of the long duration of the illness and the highly active infection, antiviral therapy with ganciclovir was given and led to a dramatical improvement of all disease manifestations. The patient subsequently remained in remission from ulcerative colitis for three years. PMID- 11100514 TI - Giant cell arteritis "causing" AA-amyloidosis with rapid renal failure. AB - Giant cell arteritis (GCA) is a systemic vasculitic disease, which in very rare cases causes inflammatory complications such as secondary amyloidosis. We describe a well-documented case, with a clinically mild course, of biopsyproven giant cell arteritis as the only apparent cause of systemic AA-Amyloidosis. The deterioration in renal function due to amyloid deposition occurred rapidly and only a few months after manifestation of giant cell arteritis and was not reversible by steroid treatment. The renal arteries were normal and there was no glomerulonephritis due to giant cell arteritis. This unique case demonstrates that giant cell arteritis with a mild clinical course is closely associated with early-onset severe secondary amyloidosis. PMID- 11100515 TI - [Amiodarone-induced thyrotoxicosis]. AB - Amiodarone is the most important drug in the treatment of ventricular arrhythmias and is widely used for atrial fibrillation. Thyrotoxicosis, a classical side effect, was thought to be iodine induced, but recent evidence suggests that other mechanisms play an important role (toxic effect on thyreocytes, immunological effects). Thyrotoxicosis due to amiodarone is difficult to treat and is further complicated by the pro-arrhythmic potential of thyrotoxicosis and the fading antiarrhythmic effect after amiodarone withdrawal. The mechanism, diagnosis and therapy of amiodarone-induced thyrotoxicosis are discussed in the light of the available literature. PMID- 11100516 TI - [Neurogenic voiding disorders. Current status of diagnosis and therapy]. AB - Treatment of the neurogenic voiding disorders which occur after spinal cord injury represents one of the most important challenges of rehabilitation. Inadequate management of neurogenic voiding disorders, especially of urinary incontinence, results in impaired quality of life. Moreover, inadequately treated neurogenic voiding disorders may result in medium and long-term renal complications and even death. Treatment of neurogenic disorders, whatever their origin (spinal cord injury, multiple sclerosis, Parkinson's disease), must take into account the gravity of the neurological disease, the potential risks for the upper urinary tract and the expected quality of life. Therefore, each patient must be considered as a separate entity and treated individually. Recent progress in the comprehension of the neurophysiology of the lower urinary tract and the neurophysiopathology of the neurogenic voiding disorders has been followed by the development of new diagnostic and therapeutic tools aimed at improving the patients' health and quality of life. PMID- 11100517 TI - [Rendu-Osler-Weber disease]. PMID- 11100518 TI - A newborn with diffuse skin rashes and an occipital mass. PMID- 11100519 TI - Management for pneumothorax in children. PMID- 11100520 TI - Current strategy for management of meconium aspiration syndrome. AB - Meconium aspiration syndrome (MAS) is characterized by the atelectasis due to the complete airway obstruction, emphysema and air leak syndrome resulted from the partial obstruction of airway, chemical pneumonitis, and surfactant dysfunction. As far as meconium is present in the airway, exogenous surfactant will be inactivated rather quickly even given as multiple doses. MAS can easily develop persistent pulmonary hypertension of the newborn. Therefore, the removal of meconium from the airway rather than the surfactant replacement therapy should be the cardinal step for the treatment of MAS. Our previous studies revealed that the removal of meconium from airway by the tracheobronchial lavage with diluted surfactant solution, 100 mg/10 mL/Kg of Surfactant-TA, resulted in the recovery of both blood gas values and lung compliance to the normal range. Thus, the current strategies of management of MAS are the selective intubation with toileting only on infants with severe distress at birth, and the early airway lavage with diluted surfactant solution, followed by high frequency oscillatory ventilation, which may prevent the further injuries in the fragile neonatal lung. PMID- 11100521 TI - Recent trends in viral hepatitis among Japanese children. AB - The prevalence of viral hepatitis in Japan were very high as peak HBsAg positive rate of 2-3% and HBsAb positive rate of 25% of peoples born in the 1930th and 1940th. But recently in Japan hepatitis A has completely eradicated and hepatitis B and C were seldom seen in our pediatric clinics. Many strategies were done beginning from sterilized water service to introduction of HCV-Ab screening system for blood transfusion in 1989. Many efforts had combined to get final success. The historical views of epidemics of viral hepatitis and maneuvers to improve were introduced here. PMID- 11100522 TI - Diagnostic pitfalls in congenital right diaphragmatic hernia. AB - Congenital diaphragmatic hernia (CDH) is a rare disease of newborns. Right-sided diaphragmatic hernia is even rarer. The clinical and radiological presentations, which are well documented in left-sided diaphragmatic hernia, are variable in right-sided diaphragmatic hernia. This makes the diagnosis of right-sided diaphragmatic hernia more difficult. During a 12-year period, seven cases of right-sided diaphragmatic hernia were collected from a single institution. Their presentations and clinical courses have been reviewed. Low prenatal diagnostic rate, various roentgenogram expressions after birth, and absence of specific clinical presentations were noted. These expressions may become pitfalls in diagnosis and lead to inappropriate treatment. From our experience in these 7 cases and a brief literature review, we try to emphasize the characteristics and the diagnostic pitfalls of this disease. In conclusion, right-sided congenital diaphragmatic hernia has a variable clinical spectrum with high mortality and morbidity. Careful evaluation of the clinical presentations, ultrasonography and chest films is mandatory for precise diagnosis. PMID- 11100523 TI - Risk factors of pulmonary hemorrhage in very-low-birth-weight infants: a two-year retrospective study. AB - Pulmonary hemorrhage is a serious complications in very-low-birth-weight (VLBW) infants with respiratory distress syndrome (RDS). We undertook a 2-year retrospective study to investigate the predisposing factors and the incidence of pulmonary hemorrhage in VLBW infants. From January 1997 through December 1998, twenty infants were diagnosed with massive pulmonary hemorrhage (MPH) according to the following criteria: active bleeding from the endotracheal tube, acute drop in hematocrit (> or = 10%), and the development of multilobar infiltration on chest radiograph. The mean gestational age was 26.9 +/- 2.5 weeks, the mean birth weight was 909 +/- 290 g. Twenty historic controls with similar gestational age and birth weight were retrospectively identified during the study period. The incidence of MPH in VLBW infants was 5.9%(20/340). A lack of prenatal corticosteroid administration, surfactant replacement therapy for RDS, and a patent ductus arteriosus (PDA) with cardiovascular dysfunction requiring dopamine support were the significantly predisposing factors of MPH in the acute stage (< or = 7th day of life). To avoid MPH and decrease mortality and morbidity in the acute stage, prenatal corticosteroid administration, evaluation of the necessity of surfactant therapy, and early recognition and aggressive treatment of hemodynamically significant PDA were necessary. PMID- 11100524 TI - Mutation analysis of human LEFTY A and LEFTY B genes in children with Ivemark syndrome. AB - The search for genes responsible for the abnormal development of the left-right (L/R) asymmetry has been conducted but no definite results have been reported. Recently, two human homologus mouse lefty1 genes, LEFTY A and LEFTY B, were analyzed for mutations in patients with the L/R anomalies. However, only two mutations were found in a survey of 126 patients. We collected genomic DNA from 10 children with Ivemark syndrome, a disease with anomalies in L/R asymmetry. Mutation analysis of LEFTY A and LEFTY B genes using single strand conformation polymorphism and direct sequencing was performed, but no mutations were found. This indicates that the L/R asymmetry anomaly in Ivemark syndrome may not be caused by the mutation of LEFTY A and LEFTY B genes. Other genes responsible for the anomalies of L/R asymmetry should be further investigated. PMID- 11100525 TI - Spontaneous pneumothorax in children. AB - Sixteen consecutive children diagnosed with spontaneous pneumothorax (SP) and admitted to Chang Gung Children's Hospital between July 1994 and June 1999 were retrospectively studied. The presenting features, radiographic abnormalities, and indications of surgery were reviewed. All patients were exclusively adolescents and were aged between 12 and 17 years. Six patients received chest tube drainage only. Nine patients underwent ablation of blebs or bullae using video-assisted thoracoscopic surgery (VATS) with good results. One patient had a bullous emphysema removed by wedge resection. One patient with spontaneous hemopneumothorax required a blood transfusion and emergent thoracotomy for stabilization. Computed tomographic (CT) scans of the chest detected all seven (100%) cases with apical bulla formations. First episodes of spontaneous pneumothorax were treated conservatively using closed-tube thoracostomy if the plain chest radiographs and CT scans were negative for apical bleb or bulla formations. In summary, SP occurred exclusively in adolescents in this study, commonly they had no underlying pulmonary abnormalities except for bulla or bleb formations. SP can be safely and effectively managed using VATS and abrasive pleurodesis in children without recurrence at follow-up. SP may be treated surgically even in the first event of pneumothorax when subpleural blebs are demonstrated by high-resolution computed tomographic scans. PMID- 11100526 TI - Transcatheter stent treatment for congenital peripheral pulmonary arterial stenosis. AB - A total of 5 Johnson and Johnson stents were implanted in two patients with significant residual peripheral pulmonary arterial stenosis. These were a 15-year old boy with post-open heart surgery for tetralogy of Fallot and a 3 8/12 year old boy with D-transposition of great vessels. Immediately after balloon dilatation and implantation of the stents, the diameter of the narrowing pulmonary arteries increased significantly from 6.0 +/- 0.8 mm to 13.5 +/- 1.7 mm (P < 0.001) and the systolic pressure gradients across the stenosis of peripheral pulmonary artery dropped significantly from 33.0 +/- 16.0 mmHg to 10.2 +/- 4.4 mmHg (P < 0.01). One year later, repeated cardiac catheterization was performed on both patients. In the patient with tetralogy of Fallot, a 20 mmHg pressure gradient was found between the main and left pulmonary artery. This patient then received another stent implantation to release the residual stenosis. The boy with D-transposition of great vessels had only 9 mmHg gradient between main and right pulmonary artery. Transcatheter placement of the stent is a feasible and effective method to treat certain patients with significant pulmonary arterial stenosis if surgical correction can not be performed. PMID- 11100527 TI - Alstrom syndrome with hepatic dysfunction: report of one case. AB - Alstrom syndrome is a rare autosomal recessive disorder associated with early childhood retinopathy, progressive sensorineural hearing loss, truncal obesity, and acanthosis nigricans. We report a 10-year-old boy with Alstrom syndrome presenting with general malaise and abnormal liver function for 1 year. In addition to the above mentioned features, he also had hyperglycemia and hyperinsulinemia. The mechanism responsible for the persistent elevation of liver enzymes could not be identified. To the best of our knowledge, this is the first reported case of Alstrom syndrome with hepatic dysfunction in Taiwan. PMID- 11100528 TI - Pediatric eosinophilic gastroenteritis: report of one case. AB - Eosinophilic gastroenteritis is an uncommon disease of unknown cause found particularly in children. It is characterized by the infiltration of eosinophils into the wall of the gastrointestinal tract, peripheral eosinophilia, and chronic relapsing gastrointestinal disorder. This report describes a patient with serosal eosinophilic gastroenteritis presenting ascites and peripheral eosinophilia. The diagnosis of the eosinophilic gastroenteritis was supported by sonographic analysis and dramatic clinical response to prednisolone therapy. PMID- 11100529 TI - Recurrent pancreatitis secondary to type V hyperlipidemia: report of one case. AB - With the exception of cystic fibrosis and hereditary pancreatitis, case reports about pancreatitis in children have rarely been mentioned. We report here an 11 year-old boy with type V hyperlipidemia, who suffered from two episodes of acute pancreatitis. Sudden onset of severe upper abdominal pain, fever, and hypertriglyceridemia were the common presentations. Initial treatments including analgesics, fasting, parenteral nutrition support and following diet control with medium-chain triglycerides seem to be successful in our case. PMID- 11100530 TI - [Ultrasonic autopsy or ecopsy]. PMID- 11100531 TI - [Descriptive study of autopsies of internal medicine department at the hospital of Barbastro and clinico-pathological correlation]. AB - BACKGROUND: To review the autopsies of Internal Medicine Department at the Hospital of Barbastro and to compare clinical and pathological diagnosis. MATERIAL AND METHODS: Retrospective study of 51 consecutive autopsies performed between April 1989 to December 1996 is carried out. Clinico-pathological correlation is stablished with the underlying cause of death distinguishing among concordance and severe (with adverse impact on survival) or mild discrepancy. RESULTS: The autopsy rate was 6.6% in that period. 70.5% were male. The median of the age was 70. Severe yatrogenia was found in one case. Respiratory diseases (23.5%) followed by cardiovasculars (19.6%) and infections (17.6%) were the most frequents underlying cause of death. Pulmonary thromboembolism was demonstrated in 37.2%, being massive only in 7.8%. Discrepancies were found in 31% of cases, 25% mild and 6% severe. CONCLUSIONS: Our data are similar to other studies but differ essentially in a lesser number of severe discrepancies in our findings and also in the frequency of the groups of diseases (there are more infections and tumours in other series) attributed to the hospital features and to the oldness of the studied population, among others. PMID- 11100532 TI - [Risk of tuberculosis in a cohort of HIV positive IVDA patients in relation to the degree of immunodeficiency and tuberculosis skin test]. AB - OBJECTIVE: The aim of this study was to assess risk for developed active tuberculosis in HIV infected IDU oblation and relate with immunodeficiency degree. METHODS: Longitudinal follow-up was realized, the period of study was 9 years, in HIV seropositive IDU patients. Control group was HIV seronegative IDU. CD4+ lymphocytes count and cutaneous test (PPd and DHT) were performed at baseline and each three months, them screening from active tuberculosis. RESULTS: Study's population was 165 patients and control group was 69. The average age was 26 and 23.8 respectively. The medium follow-up was 25 months. Active tuberculosis was developed in 9.26 cases per one hundred persons and year from follow-up. It was between 4.92 in cases with more than 500 CD4+ lymphocytes and 3.08 for them with lymphocytes between 50 and 200 per mm3. The incidence tests was 3.99 in PPD negative normoergic patients and 15.87 in PPD negative anergic patients. CONCLUSIONS: Risk from active tuberculosis was relactioned with latent infection and immunodeficiency degree. We constated high risk in low CD4+ lymphocytes counts; and degrees from DHT let an approach to tuberculosis production mechanism. PMID- 11100533 TI - [Prognostic factors of mortality in elderly patients in the emergency department]. AB - BACKGROUND: Increasing attention has been given to the elderly patients in emergency departments. However, prevalence and predictive factors of mortality in geriatric patients in emergency rooms have not been stablished. This study was conducted to analyse predictive factors associated with mortality in patients aged more than 75 years-old. METHODS: Descriptive transversal study. Data from 1003 patients aged over 75 years were recorded: biographic data, past medical history, functional capacity, acute vs chronic diseases, discharge diagnosis, emergency room stay, mortality and their causes. Bivariant and multivariant (logistic regression) analysis were performed in an attempt to establish mortality associated factors. Prevalence odds ratio (OR) and 95% confidence intervals were estimated. RESULTS: 45 patients died (4.7% IC 95%: 3.2 a 5.7). In bivariant analysis age (p = 0.008), previous heart failure (p = 0.008) or renal failure (p = 0.05), functional disability (p < 0.001), mental disorders (p < 0.001), acute diseases (p = 0.01), and the following entities: politraumatism, sepsis, respiratory failure, pneumonia, ileus, acute abdomen, and coronary artery diseases, were associated with mortality. In multivariant analysis were detected as independent prognostic factors: circulatory disorders, coronary artery diseases, sepsis, respiratory failure, and functional disability. CONCLUSIONS: Over 75 years-old patients mortality in emergency departments is more frequently associated with diagnosis of coronary heart disease, respiratory failure, sepsis and functional disability. PMID- 11100534 TI - [Economic evaluation of the use of diclofenac/misoprostol in the treatment of osteoarticular diseases]. AB - OBJECTIVE: To carry out a economic evaluation of diclofenac/misoprostol in the treatment of rheumatoid arthritis and osteoartritis when comparing with diclofenac alone, diclofenac + omeprazol, and diclofenac + ranitidine. DESIGN: Cost effectiveness analysis using a decision analytic model, where the effectiveness unit was defined as the patient free of gastro-intestinal toxicity. MATERIAL AND METHODS: The effectiveness data of the four alternatives under evaluation have been obtaining from published clinical trials. In this analysis only direct medical costs have been included without incorporating indirect costs or intangible costs. The perspective chosen has been a primary care area and the time horizon 6 months. All costs are expressed in monetary units of 1998. MEASUREMENTS AND RESULTS: The cost/effectiveness ratio obtained with diclofenac/misoprostol has been a 37% lower compared with diclofenac alone (42,238 vs 67,214 ptas), a 39% compared with diclofenac + omeprazol (42,238 vs 69,058 ptas) and a 50% compared with diclofenac + ranitidine (42,238 vs 85,198 ptas). The sensitivity analysis performed has shown that diclofenac/misoprostol is the therapeutic alternative more efficient even when most influential variables are modified. CONCLUSIONS: Diclofenac/misoprostol has demonstrated to be an alternative with a better cost/effectiveness ratio, and therefore more efficient than diclofenac alone or the concomitant use of diclofenac either with omeprazol or ranitidine. The routinary use of this association will save important resources to the National Health Service. PMID- 11100535 TI - [RS3PE syndrome: report of 11 cases]. AB - The RS3P syndrome or remitting seronegative symmetrical synovitis with pitting oedema, was described in 1985 by McCarty. The description refers to a rheumatologic set of symptoms with an acute commence, with no erosive lesions, with seronegativity, affecting more frequently to elderly males and showing an excellent prognosis. These characteristics make it possible to difference from the rheumatoid arthritis and from the rheumatic polymyalgia. We present eleven cases which confirm the good prognosis already described, but we suggest the possibility of this syndrome could be a manifestation or the beginning of other possible diseases. PMID- 11100536 TI - [Pneumonia from Nocardia nova in lung cancer]. AB - The clinical and radiographic manifestations of pulmonary nocardiosis are nonspecific. The diagnosis depends of the clinical suspicion for alerting the microbiology laboratory to use special culture and identification methods to initiate the appropriate therapy early. A nocardia pneumonia in a locally advanced non small cell lung cancer patients is present. Even after initiate the treatment the patient died. It is important to suspect the diagnosis of uncommon infection to begin the therapy. PMID- 11100537 TI - [Primary adrenal insufficiency and primary antiphospholipid syndrome]. AB - Primary adrenal insufficiency (PAI) is a rare complication of antiphospholipid syndrome (APS). The hypercoagulable state in the APS may lead to adrenal vein thrombosis and subsequently to hemorrhagic necrosis of the adrenal glands. This complication of APS is important to recognize because it may be fatal if untreated. We describe one case of PAI and primary APS, with magnetic resonance studies consistent with hemorrhagic necrosis of the adrenal glands. PMID- 11100538 TI - [Unifocal Langerhans cell granulomatosis in a young woman]. AB - We report a 24-years-old woman who presented with thoracic pain after coughing. Physical exam revealed no abnormalities except pain after pressing under blade bone area. A rib radiography and CT of the thorax showed a lonely osteolytic lesion inside eleventh left costal arch without affecting others tissues. There were no more osteolytic lesions at other levels and the histopathological study of a resection-biopsy of the lesion was diagnosed as Langerhans' cell granulomatosis. This is an uncommon disease which diagnosis can only be made through histopathological study of suspected lesions. PMID- 11100539 TI - [Clinical approach to portal hypertension]. AB - Portal Hypertension is one of the most common and severe complication arising from hepatic cirrhosis natural history. Its development conditions the patient prognosis, and its diagnosis and correct evaluation contribute to the correct management of the patient. New techniques for the measurement of portal pressure gradient allow the study and follow-up of patients with esophageal and gastric varices and with risk of hemorrhage, analyzing the efficacy of the treatment applied in a reliable and secure way. It has been probed its utility in the study of the patient with hepatocellular carcinoma, complications after liver transplantation, portal hypertension with no hepatopathy, etc. This review analyzes, from a clinical point of view, the repercussion of the development of portal hypertension in the patient with hepatic cirrhosis, its diagnosis and interpretation, and the importance that its adequate valuation has for the clinical practice. PMID- 11100540 TI - [Ticlopidine induced cholestatic hepatitis]. PMID- 11100541 TI - [Acute renal failure associated with rifampicin]. PMID- 11100542 TI - [Reflex sympathetic dystrophy after nail surgery]. PMID- 11100543 TI - [Urinary tract infection and bacterial resistance in urine cultures of outpatients]. PMID- 11100544 TI - [HIV infection in patients over sixty years old]. PMID- 11100545 TI - [Primary non-Hodgkins lymphoma of the spleen diagnosed by percutaneous biopsy]. PMID- 11100546 TI - [Acute idiopathic tubulointersticial nephritis associated with uveitis (TINU syndrome)]. PMID- 11100547 TI - [Nodular pulmonary amyloidosis. Diagnosis by transbronchial biopsy]. PMID- 11100548 TI - [Neurileptic malignant syndrome. A case associated with halazepam and fluoxetine treatment]. PMID- 11100549 TI - [Effectiveness and appropriateness in health care]. PMID- 11100550 TI - [Management applied in hospital hygiene programs]. PMID- 11100551 TI - [Hospital infections and accreditation: presentation of a project]. PMID- 11100552 TI - [Surveillance of hospital infections: review of the literature in historical perspective]. PMID- 11100553 TI - [Methods of surveillance of hospital infections]. PMID- 11100554 TI - [Systems of epidemiological typing of microorganisms responsible for hospital infections: use and evaluation]. PMID- 11100555 TI - [How to construct a protocol of surveillance of hospital infections]. PMID- 11100556 TI - [How to approach a protocol of surveillance of urinary tract infections]. PMID- 11100557 TI - [Guidelines for nosocomial infection surveillance in heart surgery and heart surgery intensive care unit. Work Group on Surveillance]. PMID- 11100558 TI - [How to approach a protocol for surveillance of infections in burn units]. PMID- 11100559 TI - [Surveillance and prevention of invasive candidiasis in oncohematology]. PMID- 11100560 TI - [Management of an epidemic of multidrug-resistant tuberculosis in patients with HIV infection in 2 hospitals of Milan]. PMID- 11100561 TI - [How to approach management of an outbreak caused by enterococci]. PMID- 11100562 TI - [Management of an epidemic caused by Pseudomonas aeruginosa]. PMID- 11100563 TI - [Sterilization in hospital environment]. PMID- 11100564 TI - [Disinfection of endoscopes]. PMID- 11100565 TI - [Hygienic management of air conditioning systems]. PMID- 11100566 TI - [Management of water supply systems]. PMID- 11100567 TI - [Management of hazardous medical waste in Italy]. PMID- 11100568 TI - [Implementation of hygienic-medical surveillance system in hospital food service for prevention of foodborne diseases]. PMID- 11100569 TI - [Management of sanitary waste. Guidelines for sanitary operators]. PMID- 11100570 TI - [Individual and collective activities for prevention and protection from biological risk in health institutions]. PMID- 11100571 TI - [Immunization of workers in medical environment]. PMID- 11100572 TI - [Compliance of hospital personnel to vaccination policy]. PMID- 11100573 TI - [Biological risk in microbiology laboratories: epidemiology and prevention]. PMID- 11100574 TI - [Efficacy of guidelines as educational tool]. PMID- 11100575 TI - [Continuing education project for health personnel in the prevention of hospital infections]. PMID- 11100576 TI - [Guidelines for selection and use of devices protective from biological agents]. PMID- 11100577 TI - [Biological risk: applied procedure for risk assessment and planning of preventive and protective interventions]. PMID- 11100578 TI - [Ethics and esthetics of direct economic incentive systems: intersections of efficiency and equity in clinical practice]. PMID- 11100579 TI - [Variability of medical practice in sinusitis as a function of resource utilization]. AB - OBJECTIVE: To find what factors linked to the doctor affect the variability of clinical practice. DESIGN: Cross-sectional descriptive study through a clinical interview with a fictitious complaint (sinusitis). SETTING: Asturias health centres. PARTICIPANTS: 132 doctors chosen through conglomerate sampling. MEASUREMENTS AND MAIN-RESULTS: Social and professional variables of the doctor (sex, age, years in practice, postgraduate training, courses, recycling courses and distance to the referral centre) were gathered. After the case was presented, the doctor's actions were noted and the cost in pesetas of each action was noted. Cost and the consumption of resources were related to the personal variables of the doctors through multivariate analysis. Mean total expenditure was 76,592 pesetas (minimum: 8958; maximum: 244,220), of which 55,550 were for short-term time off work (average of ten days off) and 15,261 for the consultations made. Average cost of treatment was 3762 pesetas. Through multiple linear regression, the only variable that showed significant effect on expenditure was distance to the referral centre (the greater the distance, the less expenditure). On transforming the total expenditure variable into a dichotomous variable (above and below the mean), the variables with significant effect on less expenditure were courses done, greater distance from the referral centre and being a woman. CONCLUSIONS: There is a very broad variability in decision-taking before the same clinical problem. The only variables that explain (and only partially) an expenditure trend are sex, distance to the referral centre and doing courses. PMID- 11100580 TI - [Utilization of generic drugs in Spain and Catalonia: possibilities of savings]. AB - OBJECTIVES: To analyze the degree of utilization of generic drugs in Spain and Catalonia. To determine real savings and scope for further savings due to the use of generic drugs. DESIGN: Descriptive, crossover study. SETTING: Public health prescription in Spain and Catalonia in June of 1999 and in Catalonia from January to June of 1999. MEASUREMENTS AND MAIN RESULTS: In June of 1999, the market share of generic drugs in Spain was 1.15% in units and 0.91% in pesetas of the hole drug prescription. The most used were oral ranitidine, oral diclofenac and oral amoxicillin. Catalonia, in this period, has a greater relative utilization of generic drugs, specially in psychotherapeutic agents like alprazolam and fluoxetine. During the first semester of 1999 only in 9.9% of the occasions where generic drugs could be used, were they actually prescribed. Specifically, the drug most used in its generic form was oral diclofenac with 27.1% of the all possible prescriptions. The real savings generated by the generic drugs were 0.2% of the public pharmaceutical bill, and the theoretical capacity of savings with the current generic offer were 2.3%. CONCLUSIONS: There is a low but fast-rising utilization of generic drugs in Spain and Catalonia. Nevertheless the possibilities of savings are at the moment, in relative terms, small. A greater use of them and specially a diversification of the offer is necessary to achieve important relative savings in the overall pharmaceutical bill. PMID- 11100581 TI - [Impact of caregiving on the health of family caregivers]. AB - OBJECTIVES: To identify the type of care provided by informal carers of dependent persons and the repercussions this care might have on the health of the carers, and to find the characteristics of both informal carers and cared-for people. DESIGN: An observational cross-sectional study. SETTING: This study was conducted in various towns in the province of Barcelona between January and December 1997 in primary health care. PARTICIPANTS: Those taking part were 240 informal carers (IC) for dependent persons. MEASUREMENTS AND MAIN RESULTS: The ICUB 97 questionnaire was the data-gathering instrument. It was validated previously by the research team and based on the fourteen needs of the Virginia Henderson nursing model. The questionnaire was filled in at a personal interview. The level of dependence of people cared for was evaluated with the Barthel and Philadelphia Geriatric Center indices. The analysis of the results reflected that the greater the level of dependence of the person cared for, the more care is provided by the carer. The main repercussions of caring on the health of the carers were: back pain (73%), tiredness (72%), reduced leisure time (73%), insomnia (65%), anxiety (72%) and changes in family life (54%). Repercussions that correlated most closely with the fact of caring were: sleep disorders, family economy, personal development and leisure, middle age and having few educational qualifications. CONCLUSIONS: Most carers are middle-aged women performing multiple care tasks. This work causes their quality of life to deteriorate. PMID- 11100582 TI - [Breast feeding and birth of twins]. AB - OBJECTIVES: To investigate the breast-feeding (BF) situation after birth of twins and find what factors affect the decision to breast-feed, its length and the reasons for stopping it. DESIGN: Retrospective, observational study. SETTING: Sabadell Hospital, Barcelona. PARTICIPANTS: All the women who gave birth to twins between January 1994 and June 1997 (n = 72). MAIN RESULTS: 64 women (88.9%) began BF. It was exclusive in 37 cases (57.8%) and mixed in the other 27 (42.2%). The main reason for choice was "better feeding" (100% of cases). The age of mothers, the type of birth, the weight of the children, admissions to the New-born Baby Department, work situation, mother's educational qualifications and the existence of domestic help affected neither initial breast-feeding nor its length. Women with prior counselling started BF in greater numbers than those not counselled (p = 0.026). Almost half of them received the information through the matron or nurse. Mean length of BF was 102 days. After two months, half of those who began BF were still breast-feeding; and at four months, 26.5%. CONCLUSIONS: We observed a high level of starting breast-feeding after twin births, almost the same as for single births. Women with previous knowledge of the question are more likely to breast-feed. The length of BF is slightly greater in the studies reviewed. Adequate prenatal, postnatal and puerperal support from health-workers can have a positive effect on the success of BF after twin births. PMID- 11100583 TI - [Externally induced prescriptions, degree of agreement and ... possibility of change in primary care?]. AB - OBJECTIVE: To find whether externally induced prescriptions (EIP) condition attendance through their prevalence, quality, the degree of agreement of the PC doctor and his/her capacity to alter them. DESIGN: Cross-sectional study of use of indication-prescription type medicines. SETTING: Health district. PARTICIPANTS: 2656 prescriptions for 678 patients interviewed. MEASUREMENTS: Each interview recorded: type of visit, age, sex, work situation, existence or otherwise of social problems and/or psychiatric pathology; doctor-patient relationship, pharmaceutical preparations (PP) prescribed and those which the patient remembers he/she is taking, indication, origin, duration, speciality of the prescribing person, agreement of the PC doctor issuing the prescription and the possibility of his/her changing it. For each prescription the following was analysed: therapeutic group, intrinsic value, time it lasts, cost and whether it is a recently marketed PP. MAIN RESULTS: 90% of visits to the doctor end in prescription. 58% of patients remember taking one or more EIP. 72% of the prescriptions analysed were externally caused. They came mostly from the public health system (66%), private medicine (20%) and self-medication (11%). There was no PC agreement with almost half these EIPs, but only 13% could be changed. The EIPs without agreement and without possibility of change were greater in: women, the elderly, people on a pension, psychiatric pathologies and in cases of bad doctor-patient relationship. The EIPs originated in health insurance companies, pharmacies, self-medication, former GPs and private doctors. They were associated with ill-defined signs and symptoms, circulatory diseases and locomotive disease. We found no significant differences in expenditure or use of PP recently put onto the market between self-medication and EIP, though there were in quality. CONCLUSIONS: The current model of prescribing medication causes consultations to be greatly "medicinised" at the expense of EIP. Doctors only alter a small part of the EIPs they don't agree with. Longitudinal studies are needed to monitor patients to find the evolution of EIPs (withdrawal, replacement, dragging on or new external prescription). PMID- 11100584 TI - [Chronic low back pain: multispecialty assessment of 100 patients]. AB - OBJECTIVES: Chronic lower-back pain (CLP) is a common pathology and has a high social and economic impact, especially in primary care where its treatment is changing at present. The results of the multi-disciplinary assessment of 100 patients with chronic lower-back pain are given. DESIGN: Cross-sectional, observational and prospective study. SETTING: Out-patient clinics of the rheumatology service of a tertiary-level hospital (referral from base districts where there is no primary care rheumatologist). PATIENTS: 100 consecutive patients seen for back pain lasting for more than 6 months were analysed. INTERVENTIONS: There was no therapeutic intervention. MEASUREMENTS AND MAIN RESULTS: The personal, work, clinical, examination, x-ray, functional and psychological features of 100 patients with CLP were analysed. There were 38 men and 62 women, with average age of 45 +/- 10 years and low social, cultural and job levels. Pain had lasted 82 +/- 7 months and 52% had had time off work. Mean intensity of pain was 6.5 +/- 2.3 (scale of zero to 10). There was vertebral restriction in 16%, and conduct expressing pain on examination in 47%. The x-ray showed disorder in 51%. Functional incapacity was nil or light in 46% and severe in 16%. 74.5% of the patients were depressed; 57% had features of anxiety; and 44% were anxious at the time of the interview. CONCLUSIONS: Patients with CLP are middle-aged, with long-standing pain and frequent time off work. Pain intensity is high, but vertebral restriction, disorders on x-rays and functional incapacity are scant. However, anxiety and depression levels are high. This could suggest a change in how we treat CLP towards a multi-disciplinary approach and psycho affective, social and labour assessment, both at the time of assessment and in later treatment of patients. PMID- 11100585 TI - [Failure of an educational intervention to change anticholesteremic drug prescription in primary care]. AB - OBJECTIVE: To evaluate the effectiveness of an intervention for decreasing hypocholesterolemic drugs prescription in patients with low cardiovascular risk profile. DESIGN: Quasi-randomized intervention study. SETTING: Public primary health care centres in the province of Valencia (Spain). PARTICIPANTS: 238 general/family practitioners from 23 primary health care centres. INTERVENTIONS: The centres were assigned to either an experimental group that received the educational intervention (individual scientific information by outreach detailing and invitation to a workshop about treatment of hypercholesterolemia in the primary care setting with the participation of an opinion leader) or a control group. MEASUREMENTS AND MAIN RESULTS: The effectiveness of the intervention was measured through the monthly mean daily defined doses (DDD) from the 4 months prior to the intervention until 5 months thereafter. Mean DDD increased along the study period in both groups, with no detectable differences between them. Similarly, there were no between-group differences after controlling the initial prescription levels using a mixed lineal model. CONCLUSIONS: The educational intervention as it was implemented was ineffective for changing overall hypocholesterolemic drug prescription in primary care. Consequently, this intervention is not justified for reducing pharmaceutical expenditure. PMID- 11100586 TI - [Sexual activity in cardiac patients. An empirical study]. AB - OBJECTIVES: The objectives of this study are to evaluate the beliefs and views linked to sexuality and sexual behaviour, and the emotional impact of a cardiac lesion. DESIGN: A retrospective cross-sectional study of the lived experience of sexuality after the appearance of the cardiac lesion. SETTING: Hospital and primary care. PATIENTS AND OTHER PARTICIPANTS: A sample of 30 people (12 women and 18 men) with a cardiac complaint, with an average age of 69.89 for women and 60.89 for men. 89% of the men had a stable partner (with an average age of 46.25), and 50% of the women (with an average age of 71.17). MEASUREMENTS AND MAIN RESULTS: Results showed that the appearance of disease was lived as a tough blow and imposed clear limitations on sufferers' lives. Specific sexual problems linked to cardiac complaints appeared, such as difficulties with erection (50%), reduction of libido (61.1%) and of intensity of pleasure (38.9%). Sexual activity was practically non-existent in the women surveyed. CONCLUSIONS: The need to contribute a proper intervention for cardiovascular patients involves recognising the need to develop adequate programmes of sexual counselling. Health-workers must also give information on these kinds of question to cardiac patients. PMID- 11100587 TI - [Battered women: medical-forensic intervention and new legislation]. PMID- 11100588 TI - [Self-measurement of blood pressure in primary care]. PMID- 11100589 TI - [Copayment and accessibility to health services. Work Group semFYC: Copayment]. PMID- 11100590 TI - [Cost effectiveness analysis of alendronate versus placebo in the prevention of hip fractures]. PMID- 11100591 TI - [Use of opioid drugs]. PMID- 11100592 TI - [Medications of wellbeing]. PMID- 11100593 TI - [Minor surgery: a professional experience]. PMID- 11100594 TI - [Primary care and winter epidemics]. PMID- 11100595 TI - [Level of medical information on diabetes, attitude of patients towards their illness and its association with the level of blood sugar control]. AB - OBJECTIVE: To determine the level of information and attitude that it has more than enough their illness has patient with diabetes type 2 (DM2), and their association with level of glycemic control. DESIGN: Cross-sectional. SETTING: Two units of family medicine. PATIENT: 200 subject with DM2. INTERVENTIONS: Two instruments were applied validated to measure, level of knowledge and attitude was measured the average of the last 6 glycemia. MEASUREMENTS AND RESULTS: The qualification average of the instrument of knowledge was 58.6 +/- 17.9 (it scale 0-100). For the instrument of attitude it was of 18.9 +/- 2.1 (it scale 0 at 35). The qualification of knowledge of the controlled group was of 55.48 +/- 16.8, and of the uncontrolled group it was of 59.2 +/- 18.1. The qualification has more than enough attitude of the controlled group it was of 17.8 +/- 2.3, and of the uncontrolled group of 19.1 +/- 2, p = 0.001. The proportionate level of information the family doctor was of 42.9%, of the team of health of 10.2% and of other sources of 6.3%. At the analysis of the degree of attitude and the level of information, there was a better attitude when the information was provided by other sources p < 0.05. In the percentage of information and the level of glycemic control, the control level was better when the information was for the team of health p < 0.01. CONCLUSIONS: The level of medical information on diabetes provided by the family doctor and the team of health is low and it doesn't and only this last are associate to better glycemic control. The attitude is better when one receives information of other sources. PMID- 11100596 TI - [Effectiveness of the protocols on cardiovascular risks in the Basque Country]. AB - OBJECTIVES: To find the effectiveness of clinical protocols in detecting and monitoring the most important cardiovascular risk factors: tobacco dependency, high blood pressure, hyperlipaemia and diabetes mellitus. DESIGN: Cross-sectional descriptive study. SETTING: Primary care centres in the Basque Country. PARTICIPANTS: 1485 clinical histories of users aged between 40 and 75 who had attended for consultation over the previous two years, registered with general practitioners in the Basque Country who normally use clinical histories. MEASUREMENTS AND MAIN RESULTS: A questionnaire was sent to the people in charge of the 103 hierarchically organised centres requesting information on the use of protocols to detect and monitor the problems studied, and copies of protocols were requested. 77 centres (76%) replied and 66 sent 170 protocols. 990 clinical histories of doctors who used the four protocols and 495 of doctors who used none were analysed. These clinical histories belonged to lists of 45 doctors from 22 centres, chosen at random and stratified by whether their centre is registered with the PAPPS (preventive programme) or is a family and community medicine teaching centre. The centres using protocols were better at detecting tobacco dependency (24.6% vs 11.9%--p < 0.0005), hyperlipaemia (61.8% vs 53.1%--p < 0.001) and diabetes mellitus (75% vs 66.1%--p < 0.0005). The difference was less in the case of high blood pressure: 56.8% vs 52.1% (p = 0.097). No differences in quality in the monitoring of these problems were found. CONCLUSIONS: The use of cardiovascular risk protocols by primary care centres is related to an improvement in the quality of cardiovascular risk detection, although not of its control. PMID- 11100598 TI - [Weight-stature growth in pediatric patients with urinary tract infection with or without vesicoureteral reflux]. AB - OBJECTIVE: To evaluate the association between urinary tract infection (UTI), vesical-uretero-renal reflux (VUR) and renal parenchyma damage, without chronic renal failure. PATIENTS AND METHODS: Retrospective study of 85 children with UTI consulting at the ambulatory paediatric unit of a private university centre in Chile, with at least 3 anthropometric evaluations: before, during and after the UTI. The group was divided into two subgroups: with and without VUR. A descriptive and inferential analysis was made. Simple regression modelling and likelihood ratio test was done for the anthropometric measures. RESULTS: Thirty nine children had VUR (46%). Both subgroups were similar. There were no difference in the nutritional status between these. Only girls with VUR have a tendency toward lower height. There was significant association between renal damage, VUR and degree of severity of it. There was no difference in the index weight for height among children with renal damage with or without RVU. In this series, other risk factors as less age, recurrent UTI and bilateral VUR with renal damage, were not cause of malnourish and lower height. CONCLUSIONS: UTI with or without VUR, with or without renal damage, excluding chronic renal failure, might not be a cause of bad physical development in children. PMID- 11100597 TI - [Prevention of thromboembolic disease in patients with heart disease]. AB - OBJECTIVE: To evaluate the adequacy to thromboembolic disease prophylaxis protocol in patients with heart disease. DESIGN: Cross-sectional study. SUBJECTS: Patients older than 14 years affected of heart disease in a semi-urban health primary-care clinic with a population of 10,610 persons and 5582 clinical records. METHODS: Data about age, sex, cardiovascular risk factors, heart disease, prophylactic treatment and its adequacy to the protocol of the "thromboembolic disease commission" of the reference hospital were analysed. RESULTS: Age 67 +/- 13 years (mean +/- SD). Cardiovascular risk factors: hypertension 40%, diabetes 33%, dyslipemia 15%, smoking 21%. Heart disease: ischemic cardiopathy 48%, atrial fibrillation 15%, valvulopathy 19%, dilated myocardiopathy 4% and other 14%. In 20% of cases had two different affections (80% with atrial fibrillation). Prophylactic therapy: 52% of patients were under prophylactic treatment (35% antiaggregation, 18% anticoagulation). Among antiaggregants, drugs used were acetylsalicylic acid 73.5%, triflusal 14.7%, dipyridamole, 8.8% and ticlopidine 3%. In 53% of people without prophylactic treatment antiaggregation criteria were present. 15% of patient under antiaggregation therapy did not meet antiaggregation criteria, and 6% fulfilled anticoagulation criteria. 67% treatments accorded the reference protocol, without significant differences between kind of heart disease or sex. The only statistically significant difference was found in age: 46% of patients older than 80 year were correctly treated, in front 75% adequacy in younger people. CONCLUSION: Prophylactic antithrombotic therapy was according the reference protocol in 67% of cases. In older patients, with greater risk of thromboembolic disease, the adequacy is worse. PMID- 11100599 TI - [Thrombolytic delay in myocardial infarction and primary care]. AB - OBJECTIVES: To analyse the effect of a primary care consultation at a health centre or at home and to determine the effect of the use of the pre-hospital electrocardiogram on thrombolytic delay. DESIGN: Analytical cross-sectional study. SETTING: La Safor county (136,000 inhabitants), Valencia, Spain. PATIENTS: Sample of 137 patients from the area admitted for acute myocardial infarction. INTERVENTION: None. MEASUREMENTS AND RESULTS: Multivariate analysis through Cox regression models of the thrombolytic delay, comparing the patients who attended a primary care centre (40, 29.2%) and those who called out a doctor to their home (26, 19.0%) with those who attended hospital (71, 51.8%). The thrombolysis proportions in the groups were analysed with logistic regression. Patients referred from primary care arrived at hospital later than those who attended directly, although a greater thrombolytic delay was only seen in those visited at home (RR 0.25, 95% CI 0.09-0.71). A primary care electrocardiogram (14 patients, 10.2%) reduced the thrombolytic delay (RR 8.81, 95% CI 1.20-64.91) by reducing intra-hospital delay. There were no differences between the groups for the thrombolysis proportion (67 patients, 48.9%). CONCLUSIONS: Patients with infarction seen in primary care reach hospital later. Calling and waiting for the doctor at home increases the thrombolytic delay. An electrocardiogram on the infarction patients who attend a health centre reduces thrombolytic delay by reducing intra-hospital delay. PMID- 11100600 TI - [Opinions and expectations of primary care professionals concerning team work]. AB - OBJECTIVE: To find the views and expectations of professionals working at primary care centres on team-work. DESIGN: Qualitative study. Focus group. SETTING: The study was conducted during May 1997 at the following primary care centres: Terrassa North, Sant Llatzer (Terrassa) and Anton de Borja (Rubi). These PCCs depend on the Department of Health and Social Security and are managed by the Terrassa Health Consortium. PARTICIPANTS: 38 primary care professionals, distributed in 5 independent groups of 8 doctors, 9 nurses, 3 social workers, 9 clerical staff and 9 auxiliaries. MEASUREMENTS AND RESULTS: Qualitative methodology and focal group technique were used. Contents of session transcriptions were analysed and the information classified according to themes. Views on the definition of team-work, basic aspects of its proper functioning, restrictions met by professionals at present and suggestions to overcome these restrictions were all identified. CONCLUSIONS: The setting of objectives, the clear and structured definition of the functions of each professional group, the participation of all the team components in decision-taking, communication, and positive enabling approaches to team-work were all specified as basic features of the team's proper functioning. Ill-defined functions, few meetings, heterogeneity within the teams, scant team-work training, and passive and ill-adapted approaches were seen as restrictions. Training, the coordinator's activities, more meetings and changes in attitude were posed as alternatives. PMID- 11100601 TI - [Sociodemographic characteristics and use of health services by the immigrant population residing in a district of the Community of Madrid]. AB - OBJECTIVE: To describe the social and demographic characteristics of the immigrant population residing in a health district and their use of health services. PARTICIPANTS: Immigrant population with papers or otherwise living in the area under study. SETTING: Health District 6 in the Community of Madrid. MEASUREMENTS AND MAIN RESULTS: Individual interviews with 300 immigrants chosen by two-stage conglomerate sampling. Average age was 32.6 (95% CI, 31.6-33.7) with 6.6 years living in Spain. 30.9% (CI, 24.3-35.9) lived in a situation of administrative and health-coverage irregularity. One of every five people had neither running water nor rubbish collection. The main health problems arising were muscular pains (69%; CI, 63.9-74.1), upper respiratory path ailments (40.3%; CI, 33.9-46.7) and accidents (27.3%; CI, 16.9-37.7). Health centres (64.5%; CI, 56.7-72.3) were used in preference to other services, with emergency being almost the only path into hospital admission. CONCLUSIONS: The immigrant population has lived in this area for a long time, despite living in adverse conditions in administrative and health terms. The public health service through INSALUD is the principal provider of health services to immigrants. PMID- 11100602 TI - [Psychopathologic evaluation of cocaine-dependent patients]. AB - OBJECTIVE: To study the presence of psychopathology in patients seeking treatment for cocaine abuse. DESIGN: Case/control study. SETTING: Addictive conduct unit. Treatment of addiction to legal and illegal drugs. Primary care. PATIENTS: Patients seeking treatment for cocaine abuse (n = 35). Control group of people with no drug-abuse problems (n = 40). MEASUREMENTS AND RESULTS: Evaluation of psychopathology through the Brief Symptom Inventory (BSI, Derogatis, 1975). Patients who abused cocaine scored high in the BSI, especially in the paranoid ideas (mean = 1.48) and obsessive-compulsive (mean = 1.25) dimensions. When the cocaine group was compared with the control group, there were significant differences in 11 of the 12 indicators. The most relevant significance occurred in the paranoid ideas and psychotic dimensions (p = 0.000). CONCLUSIONS: The high prevalence of psychiatric symptoms in patients seeking cocaine-abuse treatment was confirmed. The high scores in the obsessive-compulsive, anxiety and general malaise dimensions hindered the process of treatment in order to stop using the drug, in that the scores drop when consumption stops but start up again intensively in the following days. Given the characteristics of these patients, the BSI seems a useful instrument for guiding the diagnosis and treatment of people who abuse cocaine. PMID- 11100603 TI - [Attitude of primary care physicians to early detection of prostatic cancer through prostate-specific antigen]. AB - OBJECTIVES: To study the attitude of primary care doctors when a high (> or = 4 ng/ml) prostate-specific antigen (PSA) is found and to examine the variables linked to a prostate biopsy and the diagnosis of prostate cancer (PC). DESIGN: Descriptive, observational study. SETTING: Urban health district. PATIENTS: Ninety-four men not previously diagnosed with PC who in 1998 had a PSA figure > or = 4 ng/ml. The list was obtained from the pertinent laboratory. MEASUREMENTS: The following variables were gathered from review of clinical records: family background of PC, age, PSA figure, reason for request for PSA (if not given, it was considered a screening), referral to the urologist, rectal touch, transrectal echography, prostate biopsy and final diagnosis. RESULTS: Average age was 70 (SD, 9.31). The reason for requesting PSA was: urine symptoms in 25 (26.6%), other signs or symptoms in 25 (26.6%), request of patient in 2 cases (2.1%) and screening in 42 (44.7%). Rectal touch took place in 16 cases. Twenty-nine people were referred for examination to the urologist. 36 patients had an echography and biopsy. Variables linked to the prostate biopsy in the logistic model were: higher value of the PSA (OR 1.1; 95% CI, 1.03-1.18), being older (OR 0.92; CI, 0.87-0.98) and rectal touch performed (OR 3.58; CI, 1.02-12.51). Ten cases of PC were diagnosed. CONCLUSIONS: The most common reason for a PSA request was screening. Prostate biopsy was not requested for 58 men. A primary care guide to action concerning PC diagnosis in cases of PSA > or = 4 ng/ml would be useful. PMID- 11100605 TI - [Our elderly people become itinerant]. PMID- 11100604 TI - [Evaluation of the Glucocard Memory 2 analyzer for measuring glucose concentration in capillary blood]. AB - OBJECTIVE: To evaluate the analytical reliability and accuracy as well as the practicability of the Glucocard Memory 2 glucose meter, intended to the control of the diabetic patient. DESIGN: Descriptive, crossover study. To validate an analytical instrument according to guidelines of the Spanish Society of Clinical Biochemistry and Molecular Pathology. SETTING: Primary health care, urban setting. PARTICIPANTS: Ninety-three blood samples from diabetic patients were used. These samples were selected by a consecutive sampling of the tubes received in the laboratory for the diabetes follow-up protocol. MEASUREMENTS AND MAIN RESULTS: Repeatability of the system was studied analysing the within-run precision at four concentrations of glucose. We obtained coefficients of variation between 2.12% (at 410 mg/dl of glucose) and 4.17% (at 37.2 mg/dl). The linearity study allowed to check experimentally the linear response of the instrument between 27 and 485 mg/dl. The accuracy was evaluated comparing the Glucocard results with the routine procedure of our laboratory (Hitachi 747, GOD PAP) and calculating the regression parameters with the Passing and Bablok method (y = 1.01 x -2.34) and the intraclass correlation (99%). To evaluate the clinical significance of possible deviations related with the reference laboratory method the "error Grid" analysis was used. This analysis showed that 100% of Glucocard Memory 2 results fell into the clinical accuracy zone. Practicability study showed that the instrument is very simple to use. CONCLUSIONS: Glucocard Memory 2 is a glucose meter intended to the measurement of glucose both on capillary and venous blood that, besides its extreme simplicity of use, shows very good analytical features. PMID- 11100606 TI - [Which statin is more efficient? Concepts and applications in economic evaluation]. PMID- 11100607 TI - La depresion en el anciano. PMID- 11100608 TI - [Response to the editorial in Atencion Primaria "Generic drug lines: all is not gold that glitters"]. PMID- 11100609 TI - [Cardiac tamponade: suspected diagnosis in primary care]. PMID- 11100610 TI - [Self-care in acute diseases in primary care. Results of a qualitative study]. PMID- 11100611 TI - Reducing child mortality. PMID- 11100612 TI - Child mortality--the challenge now. PMID- 11100613 TI - The decline in child mortality: a reappraisal. AB - The present paper examines, describes and documents country-specific trends in under-five mortality rates (i.e., mortality among children under five years of age) in the 1990s. Our analysis updates previous studies by UNICEF, the World Bank and the United Nations. It identifies countries and WHO regions where sustained improvement has occurred and those where setbacks are evident. A consistent series of estimates of under-five mortality rate is provided and an indication is given of historical trends during the period 1950-2000 for both developed and developing countries. It is estimated that 10.5 million children aged 0-4 years died in 1999, about 2.2 million or 17.5% fewer than a decade earlier. On average about 15% of newborn children in Africa are expected to die before reaching their fifth birthday. The corresponding figures for many other parts of the developing world are in the range 3-8% and that for Europe is under 2%. During the 1990s the decline in child mortality decelerated in all the WHO regions except the Western Pacific but there is no widespread evidence of rising child mortality rates. At the country level there are exceptions in southern Africa where the prevalence of HIV is extremely high and in Asia where a few countries are beset by economic difficulties. The slowdown in the rate of decline is of particular concern in Africa and South-East Asia because it is occurring at relatively high levels of mortality, and in countries experiencing severe economic dislocation. As the HIV/AIDS epidemic continues in Africa, particularly southern Africa, and in parts of Asia, further reductions in child mortality become increasingly unlikely until substantial progress in controlling the spread of HIV is achieved. PMID- 11100614 TI - Reducing child mortality in India in the new millennium. AB - Globally, child mortality rates have been halved over the last few decades, a developmental success story. Nevertheless, progress has been uneven and in recent years mortality rates have increased in some countries. The present study documents the slowing decline in infant mortality rates in india; a departure from the longer-term trends. The major causes of childhood mortality are also reviewed and strategic options for the different states of India are proposed that take into account current mortality rates and the level of progress in individual states. The slowing decline in childhood mortality rates in India calls for new approaches that go beyond disease-, programme- and sector-specific approaches. PMID- 11100615 TI - Trends in under-5 mortality rates and the HIV/AIDS epidemic. AB - INTRODUCTION: The prevalence of human immunodeficiency virus (HIV) among adults and mortality rates among under-5-year-olds have increased or stagnated in many countries. The objective of this study was to investigate whether there is a link between under-5 mortality trends and the prevalence of HIV among adults and, if so, to assess the magnitude of the effect of adult HIV prevalence on under-5 mortality rates. METHOD: Data from Demographic and Health Surveys were used to establish the trends in under-5 mortality rates for 25 countries for which there are data for at least two points in time. Countries were ranked according to the most recent adult HIV prevalence data and grouped in three categories: those with very high HIV prevalence (> or = 5%); those with moderately high prevalence (1 4.9%); and those with low prevalence (< 1%). A mathematical model was fitted to obtain an estimate of the contribution of HIV/AIDS to the level of under-5 mortality in each country. RESULTS: Under-5 mortality rates showed an increase in most countries with high adult HIV prevalence, but a decrease in almost every country with moderately high or low prevalence. The estimated contribution of adult HIV prevalence to the observed level of under-5 mortality was highest (up to 61%) in Zimbabwe (where HIV prevalence was highest) and tended to decrease with the level of HIV prevalence. DISCUSSION: The contribution of HIV/AIDS to childhood mortality therefore appears to be most noticeable in settings where the epidemic is most severe. PMID- 11100616 TI - Malnutrition as an underlying cause of childhood deaths associated with infectious diseases in developing countries. AB - INTRODUCTION: Recent estimates suggest that malnutrition (measured as poor anthropometric status) is associated with about 50% of all deaths among children. Although the association between malnutrition and all-cause mortality is well documented, the malnutrition-related risk of death associated with specific diseases is less well described. We reviewed published literature to examine the evidence for a relation between malnutrition and child mortality from diarrhoea, acute respiratory illness, malaria and measles, conditions that account for over 50% of deaths in children worldwide. METHODS: MEDLINE was searched for suitable review articles and original reports of community-based and hospital-based studies. Findings from cohort studies and case-control studies were reviewed and summarized. RESULTS: The strongest and most consistent relation between malnutrition and an increased risk of death was observed for diarrhoea and acute respiratory infection. The evidence, although limited, also suggests a potentially increased risk for death from malaria. A less consistent association was observed between nutritional status and death from measles. Although some hospital-based studies and case-control studies reported an increased risk of mortality from measles, few community-based studies reported any association. DISCUSSION: The risk of malnutrition-related mortality seems to vary for different diseases. These findings have important implications for the evaluation of nutritional intervention programmes and child survival programmes being implemented in settings with different disease profiles. PMID- 11100617 TI - Is malnutrition declining? An analysis of changes in levels of child malnutrition since 1980. AB - Nutritional status is the best global indicator of well-being in children. Although many surveys of children have been conducted since the 1970s, lack of comparability between them has made it difficult to monitor trends in child malnutrition. Cross-sectional data from 241 nationally representative surveys were analysed in a standard way to produce comparable results of low height-for age (stunting). Multilevel modelling was applied to estimate regional and global trends from 1980 to 2005. The prevalence of stunting has fallen in developing countries from 47% in 1980 to 33% in 2000 (i.e. by 40 million), although progress has been uneven according to regions. Stunting has increased in Eastern Africa, but decreased in South-eastern Asia, South-central Asia and South America; Northern Africa and the Caribbean show modest improvement; and Western Africa and Central America present very little progress. Despite an overall decrease of stunting in developing countries, child malnutrition still remains a major public health problem in these countries. In some countries rates of stunting are rising, while in many others they remain disturbingly high. The data we have presented provide a baseline for assessing progress and help identify countries and regions in need of populationwide interventions. Approaches to lower child malnutrition should be based on successful nutrition programmes and policies. PMID- 11100618 TI - The evolution of child health programmes in developing countries: from targeting diseases to targeting people. AB - Mortality rates among children and the absolute number of children dying annually in developing countries have declined considerably over the past few decades. However, the gains made have not been distributed evenly: childhood mortality remains higher among poorer people and the gap between rich and poor has grown. Several poor countries, and some poorer regions within countries, have experienced a levelling off of or even an increase in childhood mortality over the past few years. Until now, two types of programmes--short-term, disease specific initiatives and more general programmes of primary health care--have contributed to the decline in mortality. Both types of programme can contribute substantially to the strengthening of health systems and in enabling households and communities to improve their health care. In order for them to do so, and in order to complete the unfinished agenda of improving child health globally, new strategies are needed. On the one hand, greater emphasis should be placed on promoting those household behaviours that are not dependent on the performance of health systems. On the other hand, more attention should be paid to interventions that affect health at other stages of the life cycle while efforts that have been made to develop interventions that can be used during childhood continue. PMID- 11100619 TI - Reducing deaths from diarrhoea through oral rehydration therapy. AB - In 1980, diarrhoea was the leading cause of child mortality, accounting for 4.6 million deaths annually. Efforts to control diarrhoea over the past decade have been based on multiple, potentially powerful interventions implemented more or less simultaneously. Oral rehydration therapy (ORT) was introduced in 1979 and rapidly became the cornerstone of programmes for the control of diarrhoeal diseases. We report on the strategy for controlling diarrhoea through case management, with special reference to ORT, and on the relationship between its implementation and reduced mortality. Population-based data on the coverage and quality of facility-based use of ORT are scarce, despite its potential importance in reducing mortality, especially for severe cases. ORT use rates during the 1980s are available for only a few countries. An improvement in the availability of data occurred in the mid-1990s. The study of time trends is hampered by the use of several different definitions of ORT. Nevertheless, the data show positive trends in diarrhoea management in most parts of the world. ORT is now given to the majority of children with diarrhoea. The annual number of deaths attributable to diarrhoea among children aged under 5 years fell from the estimated 4.6 million in 1980 to about 1.5 million today. Case studies in Brazil, Egypt, Mexico, and the Philippines confirm increases in the use of ORT which are concomitant with marked falls in mortality. In some countries, possible alternative explanations for the observed decline in mortality have been fairly confidently ruled out. Experience with ORT can provide useful guidance for child survival programmes. With adequate political will and financial support, cost effective interventions other than that of immunization can be successfully delivered by national programmes. Furthermore, there are important lessons for evaluators. The population-based data needed to establish trends in health service delivery, outcomes and impact are not available in respect of diarrhoea, as is true for malaria, pneumonia and other major childhood conditions. Standard indicators and measurement methods should be established. Efforts to change existing global indicators should be firmly resisted. Support should be given for the continuing evaluation and documentation activities needed to guide future public health policies and programmes. PMID- 11100620 TI - Factors associated with trends in infant and child mortality in developing countries during the 1990s. AB - The 1990s have seen a remarkable decrease in mortality among infants and children in most developing countries. In some countries, particularly in sub-Saharan Africa, these declines in mortality among children have slowed and are now increasing again. Internationally comparable data derived from survey programmes, such as the Demographic and Health Survey (DHS) programme, are available both to document the changes that have occurred in mortality and to provide insight into some of the factors that may explain these trends in mortality. The factors found in repeated DHS programmes that explain these trends fall into five categories: fertility behaviour; nutritional status, breastfeeding, and infant feeding; the use of health services by mothers and for children; environmental health conditions; and socioeconomic status. Both simple analyses and multivariate analyses of changes in these factors between surveys indicate that all factors affected the mortality trends. However, to explain trends in mortality, the variables themselves had to have changed over time. During the 1990s fertility behaviour, breastfeeding, and infant feeding have changed less than other factors and so would seem to have played a smaller role in mortality trends. This study confirms that trends in mortality during the 1990s were related to more than just a handful of variables. It would, therefore, be a mistake to concentrate policy actions on one or a few of these while forsaking others. Countries with the largest decreases in mortality have had substantial improvements in most of the factors that might be used to explain these changes. In some countries mortality has risen. In part these increases can be explained by the factors included in this study, such as deterioration in seeking medical care for children with fever. Other factors that were not measured, such as the increasing resistance of malaria to drug treatment and the increased prevalence of parental HIV/AIDS, may be contributing to the increase noted. PMID- 11100621 TI - Measuring nutritional status in relation to mortality. PMID- 11100622 TI - Mortality in second and third degree malnutrition. 1956. PMID- 11100623 TI - Royalty-free licenses for genetically modified rice made available to developing countries. PMID- 11100625 TI - Adult-onset Still's disease. PMID- 11100624 TI - New fly trap may reduce prevalence of blindness from trachoma. PMID- 11100626 TI - Infectious diseases in England and Wales: October to December 1999. PMID- 11100627 TI - Outbreak of heterosexually acquired syphilis in Cambridgeshire. PMID- 11100628 TI - Public health impact of flooding. PMID- 11100629 TI - Culturing the throat to protect the heart: Dr. Milton Markowitz and the prevention of rheumatic fever. AB - The purpose of this article is to briefly review how the understanding of rheumatic fever has evolved over the last 50 years. Particular emphasis is given to the identification of the Group A streptococcus as the causative agent of rheumatic fever and the use of antibiotics to treat and prevent rheumatic fever. Throughout his 50-year career, Dr. Milton Markowitz, former chairman of the department of pediatrics at the University of Connecticut School of Medicine, has been involved in these advances as an international expert on rheumatic fever prevention. Using archival materials, including some of the earliest literature descriptions of rheumatic fever, as well as an extended interview with Dr. Markowitz, the authors present a unique personal perspective both on the history of rheumatic fever and the context within which the scientific advances that have led to contemporary treatment and prevention strategies have evolved. PMID- 11100630 TI - Unusual presentation of "extracavitary" primary effusion lymphoma in previously unknown HIV disease. AB - Human herpes virus, type 8, also called Kaposi's sarcoma-associated virus, is associated with primary effusion lymphoma, an uncommon and unusual subset of acquired immunodeficiency syndrome-related lymphomas mostly confined to body cavities, which primarily affects human immunodeficiency virus-positive men. We report the case of a 40-year-old male with primary effusion lymphoma that presented initially with generalized lymphadenopathy and hepatosplenomegaly, followed by pericardial effusion and cardiac tamponade, in a previously undiagnosed human immunodeficiency virus patient. Cytomorphological studies disclosed a large-cell lymphoma with a population of cells demonstrating intermediate CD45 expression and partial coexpression of CD20 and CD23 markers, as well as universal expression of HLA-DR, CD71, CD38, and CD-30. Molecular studies showed clonal B-cell gene rearrangements and molecular evidence of human herpes virus, type 8. This case stresses the necessity, even in the absence of the 'classical clinical features,' of molecular testing for human herpes virus, type 8 in a subset of patients with high risk for human herpes virus, type 8 associated lymphomas. PMID- 11100631 TI - Mammography utilization in a skilled nursing facility. AB - OBJECTIVE: To examine mammography utilization and subsequent clinical decisions in a skilled nursing facility in women aged 75 years and older with no history of breast cancer. METHODS: This study was a retrospective medical chart review of 95 women living in a skilled nursing facility aged 75 years and older who did not have a history of breast cancer. RESULTS: One-hundred seventy-nine mammograms were performed on 95 patients with an average of 1.9 mammograms per patient. Forty-five percent of the women had some evidence of abnormal results requiring further investigation. DISCUSSION: Determining the utility of screening mammography in long-term care populations is essential to the development of appropriate guidelines. Future research should further explore the influence of patient preferences and psychological burden. PMID- 11100633 TI - Drugs of choice for cancer chemotherapy. PMID- 11100632 TI - Dofetilide (Tikosyn). PMID- 11100635 TI - The human side. PMID- 11100634 TI - Preferences for CPR among the elderly: the influence of attitudes and values. AB - OBJECTIVES: While many older individuals wish to forgo cardiopulmonary resuscitation regardless of potential positive outcomes, others desire this intervention despite low chances of survival. This study examines the extent to which health, function, attitudes, and values influence preferences for cardiopulmonary resuscitation. DESIGN: An in-person, physician-administered survey. SETTING: Three clinical sites affiliated with a university-based geriatrics program. PARTICIPANTS: One hundred three individuals age 65 or older with ability to speak, read, and write English. MEASUREMENTS: Demographic, health and functional status, social involvement, religiosity, attitudes, and values were assessed. A standard description of cardiopulmonary resuscitation was followed by open-ended questions regarding treatment preferences. RESULTS: Mean age was 81 (+/- 7), 66% were women and 78% had an advance directive. The majority (75%) declined cardiopulmonary resuscitation. Women were more likely to decline cardiopulmonary resuscitation. Attitudes toward life were the strongest predictors for the cardiopulmonary resuscitation decision. Family issues were important, especially the prospect of becoming a burden, as well as the outcome of cardiopulmonary resuscitation. CONCLUSION: Attitudes toward life, perceived outcome of cardiopulmonary resuscitation, and family issues play a significant role in end-of-life treatment decision-making. PMID- 11100636 TI - The fruit of the vine: aid to longevity? PMID- 11100637 TI - Qualidigm's ongoing evaluation of Medicaid managed care. PMID- 11100638 TI - "I shall please". PMID- 11100639 TI - Diagnostic value of cervical discography. PMID- 11100640 TI - Malingering and symptom magnification. PMID- 11100641 TI - A study of twelve hospital ethics committees in eastern South Carolina. PMID- 11100642 TI - Trends in physical activity levels among black and white adults in South Carolina. PMID- 11100643 TI - Promoting professionalism: a primer. PMID- 11100644 TI - The hospital ethics committee: integral player or passive observer? PMID- 11100645 TI - Hospital ethics committees: good news and bad. PMID- 11100646 TI - MCN: 25 years and counting. AB - The history of the journal MCN, The American Journal of Maternal/Child Nursing, is reviewed in this article, pointing out how the journal began and the changes it has undergone. Statistics concerning types of manuscripts published throughout its 25 year history are presented, and the future plans for the journal are described. PMID- 11100647 TI - Trends in nursing education since 1976. AB - Since the first issue of MCN was published, many changes have transpired in nursing education on all levels. These trends in nursing education since 1976 are addressed for diploma, associate, baccalaureate, master's, and doctoral programs. Examples of such trends focus on enrollments, graduations, and minority students. Trends to watch for in nursing education over the next quarter century are forecasted along with recommendations for nurse educators. PMID- 11100648 TI - International nursing: the past 25 years and beyond. AB - International nursing has grown substantially in the last 25 years. Once practiced only during war time, with immigrant populations, or in peace time in consultative roles, international nursing is now common, with direct practice, education, consultation, and research collaboration as the roles in which nurses work across borders. A brief history of international nursing, forces that have influenced its growth, and an assessment of its future are discussed. PMID- 11100649 TI - A critical evaluation of the past 25 years of perinatal nursing practice: opportunities for improvement. AB - Changes in perinatal nursing over the past 25 years have been abundant. Nursing advocacy has contributed to the introduction of innovations such as single-room maternity care and family-centered care, and to the end of restrictive practices such as the use of hand and leg restraints during birth, routine use of episiotomy, and routine general anesthesia for normal births. Perinatal nursing involves complex clinical interventions, intensive patient and family education, empathetic support and evaluation of family dynamics, and a wide range of opportunities to make a difference in the lives of mothers, babies, and families. The strengths and weaknesses of perinatal nursing practice at the beginning of the new century are chronicled, and suggestions for improvements are made. PMID- 11100650 TI - Perinatal nursing research: a 25-year review--1976-2000. AB - Parent-infant nursing research from 1976-2000 is reviewed through four groups of studies: development of research instruments, studies of mothers and fathers through the childbearing years, studies of newborns (both healthy and at risk), and studies of special populations. Potential directions for maternal/parent/newborn nursing research in the 21st century are suggested. PMID- 11100651 TI - Nurses speaking up for mothers and children: 25 years of public policy involvement. AB - Public policy decisions directly affect the health care of women and children and also affect the practice of maternal and child nursing. The past quarter century has seen a shift in nursing involvement in the public policy process. Heightened awareness of the collective power of nurses, greater independence of the nursing profession, the increasing capability for generating research to guide the formulation of public policy, and nurses' better understanding of the political process have all contributed to the increasing influence of our nation's 2.6 million nurses. The passage of several significant pieces of legislation, such as expansions of the Medicaid program for pregnant women and children in the late 1980s, have opened up new opportunities for nurses to further shape the nation's health care agenda for women and children. Nurses can and should become more involved with the policy-making process at local, state, and national levels to assure that decisions are made that benefit this important population group. Leadership in the public policy arena will give nurses the best opportunities for putting forth the agendas that will accomplish these goals. PMID- 11100652 TI - Returning to our roots: 25 years of maternal/child nursing in the community. AB - In many ways the past 25 years in maternal/child nursing have brought us back to community roots. Components of community care in maternal/child nursing include self-care; prevention; family, culture, and community; and collaboration. These components are reviewed through a retrospective look at community maternal/child nursing activities during the past 25 years. In addition, maternal/child nursing for the future is examined by thinking expansively in five areas: childbirth education, community-based interventions for better health behavior, community based interventions to meet 2010 objectives, family forms, and definitions of women's health in the United States and beyond. We are birthing a new vision of maternal/child health and wellness. Central to that birthing is an awakening of the still, small, wise intuitive voice in all of us. PMID- 11100653 TI - Pediatric nursing practice: keeping pace with technological advances. AB - Over the past 25 years, extensive technological and medical advances have had a major impact on the way pediatric nursing is practiced. Pediatric nurses have expanded their nursing roles, established professional organizations and certification standards to ensure clinical competence at the bedside, and tirelessly advocated for the health care needs of children and their families. In addition, pediatric nurses have collaborated with other health care providers to institute family-centered and developmentally appropriate philosophies of care. All of these changes will assist pediatric nurses to remain focused on the most important aspect of their work: Supporting the unique needs of children and their families. PMID- 11100654 TI - Nurse-midwifery: yesterday, today, and tomorrow. AB - This article chronicles the dramatic changes in nurse-midwifery over the last 25 years. Presently, multiple models of midwifery education leading to certification exist, all within a competency-based framework. Accreditation of education programs and the certification process within nurse-midwifery remain examples to others. The consumer demand for certified nurse-midwives continues to rise, spurring the preparation for more professionals. However, the average woman in the United States still does not have access to a certified nurse midwife/certified midwife for care. Several of the barriers to practice have been dismantled during the last quarter century; however, adequate reimbursement, relationships with various groups, and managed care are among the issues that will challenge midwifery in the new century. PMID- 11100655 TI - Advanced practice in maternal/child nursing: history, current status, and thoughts about the future. AB - Advanced practice nursing has undergone dramatic growth and change in during the past quarter century. Specialization in maternal/child health started with hospital-developed postlicensure programs which evolved into formal master's level programs. The first nurse-practitioner program in pediatrics was begun in 1965 at the University of Colorado. The last 25 years has seen an increase in clinical specialization and in advanced practice roles within the specialties. The growth of credentials and types of certification available have provided opportunities for nurses, but have also created confusion for consumers and other health care professionals. Current challenges facing advanced practice nurses include issues related to legal authority for scope of practice, direct reimbursement for services, and prescriptive authority. The current health care climate provides challenges and opportunities for nurses in advanced practice. PMID- 11100656 TI - Celebrating 25 years of pediatric nursing research: progress and prospects. AB - This article was designed to celebrate and highlight contributions of nurse researchers to the care of children, adolescents, and families across healthcare settings and to provide direction for future research focused on our ultimate goal of optimizing health and healthcare for the consumers of our services. It focuses on the past 25 years of pediatric nursing research, which have witnessed substantial advances in the conceptualization, design, and methods of nursing research focused on children, adolescents, and families. PMID- 11100657 TI - Information technology in maternal/child nursing: past, present, and future. AB - This issue commemorates the remarkable 25-year span of MCN, a journal born at the dawn of the microcomputer revolution. Key events in computer history and nursing informatics are chronicled, followed with visions of future technologies. A review of the technology literature published in MCN is summarized. Clinical implications include four nursing strategies for integrating electronic technology into practice and a list of online information resources. PMID- 11100658 TI - [Hypercalcemia in granulomatous diseases]. PMID- 11100659 TI - [The White Book of Spanish nephrology III. Spanish Society of Nephrology]. PMID- 11100660 TI - [Diagnostic tests: their use in epidemiological studies]. PMID- 11100661 TI - [Mechanisms implied in aminoglycoside-induced nephrotoxicity]. PMID- 11100662 TI - [Hormonal profile and participation of nitric oxide in salt-sensitive and salt resistant essential arterial hypertension]. AB - Recent studies have shown that cardiovascular events and end-organ damage occur more frequently in patients with salt-sensitive essential hypertension (SH) than in salt-resistant essential hypertension (RH). Nitric oxide (NO) plays an important role in regulating the pressure-natriuresis relationship. Therefore impaired NO synthesis may produce or aggravate salt-sensitive hypertension. This study was conducted to determine the hormonal levels and nitric oxide metabolites in hypertensive patients. 25 patients underwent salt sensitivity testing. 24 h ambulatory blood pressure was recorded after a 5-day period on low salt diet (20 mEq/d) and after a 5-day period on a high salt diet (200 mEq/d). Subjects showing > or = 10 mmHg increase in mean BP when changing from low to high dietary salt intake were classified as salt sensitive and as salt resistant when the BP changes were < 10 mmHg. Based on BP recordings 13 patients were characterised as white coat hypertension (WC), 13 patients as salt resistant (SR) and 12 as salt sensitive (SS). A significative relationship was seen between plasma glucose insulin concentration and body mass index. The ventricular mass index was similar in SS and SR patients. The plasma uric acid, triglicerides and PAI-I were elevated in SS compared with SR, and control group (C). During low sodium intake, plasma renin and aldosterone were decreased in SS compared with SR, and C. No differences in plasma catecholamines or their changes with intake sodium modifications were seen among the patients. During high sodium intake urinary NO excretion increased in SR (38 +/- 9 vs 18 +/- 2 mg/g creat), and C (24 +/- 2 vs 16 +/- 3 mg/g creat) (p < 0.01) but not in SS patients (21 +/- 3 vs 26 +/- 4 mg/g creat). The NO excretion changes showed negative correlation with BP changes (r = 0.49, p < 0.01). During low sodium intake, SR and SS patients showed a normal nocturnal decrease of BP (dippers). During high sodium intake SS patients became non-dippers. Our results showed that patients with salt sensitive hypertension displayed a suppressed renin-aldosterone system, an attenuated nocturnal decline in blood pressure on high-salt diet and an impairment of endothelial function. The relationship between urinary nitrate excretion and arterial pressure suggest that the salt sensitivity of arterial pressure may be related bo blunted generation of endogenous nitric oxide. PMID- 11100663 TI - [Increased adrenomedullin levels in hypertensive patients on maintenance hemodialysis]. AB - Hypertension is a frequent finding in uremic patients. The pathogenesis of this complication in uremia is complex and not fully elucidated. An imbalance between the vasoconstrictor and vasodilator systems may be involved in its pathogenesis. In this study we have evaluated the state of nitric oxide (NO) and adrenomedullin (ADM) in hemodialyzed patients, especially those with hypertension. We included a group of hypertensive hemodialyzed patients (n = 9) and a group of normotensive control patients (n = 10). We measured plasma renin activity, as well as plasma catecholamines, ADM, and nitrite/nitrate levels in basal conditions before starting the hemodialysis session. Plasma volume, as well as left ventricular ejection fraction were also measured. Hemodialysis patients showed plasma levels of nitrite/nitrates and ADM higher than the reference values in the normal population. We observed no differences in the plasma levels of nitrite/nitrates, but ADM levels were higher in hypertensive (278.2 +/- 15.5 pg/ml) patients than in normotensive patients (225 +/- 9.9 pg/ml) (p < 0.05). When considering all patients together, mean arterial pressure positively correlated with plasma ADM (r = 0.468, p < 0.05). Plasma volume and left ventricular ejection fraction were similar in the two groups of patients. In summary, plasma levels of nitrite/nitrates and ADM are increased in hemodialyzed patients, although only ADM levels were further increased in hypertensive patients. Our results do not suggest that a decreased production in the vasodilator factors evaluated is involved in the pathogenesis of hypertension in uremic patients. PMID- 11100664 TI - [Interpretation of health-related quality of life of patients on replacement therapy in end-stage renal disease]. AB - STATEMENT: Older age is associated with a worse Health-Related Quality of Life (HRQOL) in patients with End-Stage Renal Disease (ESRD). The aim of this study is to demonstrate that differences on HRQOL between two groups of patients, defined according to age (< 65 years and > or = 65 years or elderly), change according to the form in which the results are analysed. METHODS: We evaluated the HRQOL of 170 patients undergoing hemodialysis and 210 transplant patients from Asturias (Spain), using the SF-36 Health Survey. Sociodemographic and clinical data, Karnofsky Scale and a Comorbidity Index were also collected. The raw scores of the SF-36 and the standardised scores according to age and gender were employed. RESULTS: The majority of elderly patients on hemodialysis lived alone, constituted a smaller percentage on the transplant waiting list, had a lower serum albumin and lower score of the Karnofsky Scale, than patients under 65 years. No differences were found in transplant patients. The raw scores on the SF 36 were less for the elderly patients on hemodialysis and transplant. The raw scores for elderly undergoing hemodialysis were less than those obtained by the general population, and raw scores for elderly transplant patients were similar or slightly greater. The standardized scores of the SF-36 were greater for the elderly in both treatment groups. CONCLUSIONS: Important differences exist in the evaluation of HRQOL differences between the two groups of age according to the method of analysing the results. The HRQOL of elderly patients is better than that of patients under 65 years of age. PMID- 11100665 TI - [Hemoglobin levels and probability of better quality of life in chronic hemodialysis patients]. AB - Hemoglobin is one of the health quality of life predictors in chronic hemodialysis patients. We were interested in knowing whether high or normal levels of hemoglobin corresponded to better quality of life and where it could be find the threshold value from which the odds of improvement would be significatively different. There were studied, transversal and retrospectively, 87 patients (38 females) in chronic hemodialysis (ages: 54 +/- 23 years--time in HD treatment 52.30 +/- 56.40 months--kt/vsp: 1.30 +/- 0.12, TACu: 94 +/- 93 mg/dl and PCR: 1.05 +/- 0.33 g/kw/d. They were gathered in G1: X Hb: 8.70 +/- 1.06 g/dl, G2: X Hb: 10.55 +/- 0.40 g/dl y G3 X Hb: 11.92 +/- 0.66 g/dl and were measured the eight SF-36 domains. There were founded significative differences among the G1 and G2 scores in general health, vitality and social functioning (table II). The hemoglobin was predictors of general health as to vitality and both showed as direct relationship. The odds differences in a better quality of life (table III) were seen in general health and vitality, respectively, and with 13.5 g/dl or more, there++ were no statistically significative differences. PMID- 11100666 TI - [Study of quality of life of patients on maintenance hemodialysis before and after the use of erythropoietin]. PMID- 11100667 TI - [Association of hypercalcemia, elevated levels of calcitriol and tuberculosis in patients on hemodialysis]. AB - Hypercalcemia is associated with numerous chronic granulomatous processes and chronic infections. Increased production of calcitriol by activated macrophages has been shown to be the cause in most cases. In this article, we describe three cases of hypercalcemia associated with inappropriately elevated calcitriol levels and suppressed PTH in hemodialysis. In addition to conventional techniques for tuberculosis diagnosis we used Ligase Chain Reaction (LCR) to detect mycobacterial DNA in pleural effusion with acid-fast stain and culture negativity. Antituberculous therapy was associated with a decrease in the levels of calcium, as well as in serum calcitriol concentrations, and a substantial increase in the levels of iPTH. The serum levels of 25(OH)D3 remained unchanged. These findings suggested ectopic production of calcitriol. The discussion reviews the previously reported cases of hypercalcemia and tuberculosis that occurred during hemodialysis, and concludes that ectopic production of calcitriol by tuberculous granulomas is extremely unusual and its demonstration requires a high index of suspicion. Molecular techniques are a potentially useful approach for early and rapid diagnosis of tuberculous infection in dialysis patients. PMID- 11100668 TI - [G-CSF versus GM-CSF in the treatment of neutropenia in a patient with Felty's syndrome on hemodialysis]. AB - We describe a patient with Felty's syndrome and chronic renal failure due to secondary amyloidosis in a periodic haemodialysis programme, who was successfully treated for neutropenia with sequential administration of colony-stimulating factors: granulocyte colony-stimulating factor and granulocyte macrophage colony stimulating factor. PMID- 11100669 TI - [Glomerulocystic kidney disease and hemolytic-uremic syndrome: clinicopathological case]. AB - Glomerulocystic kidney is a heterogeneous group of conditions morphologically characterised by multiple cortical cysts apparently originated from a cystic dilation of the filtration space with atrophy of the glomerular tufts. We report a case of glomerulocystic kidney affecting a 13-year-old boy who underwent renal transplantation for end-stage renal disease following a haemolytic-uraemic syndrome diagnosed nine years ago. The absence of other stigmas (urinary obstruction, extrarenal congenital abnormalities and family history of cystic kidney disease) suggest that our observation is apparently a sporadic and acquired glomerulocystic kidney following a haemolytic-uraemic syndrome, an infrequent association previously reported only twice. Our histological and immunohistochemical findings suggest that the cysts in this rare condition are really of glomerular origin but the pathogenesis of cyst development remains unknown. PMID- 11100670 TI - [False-negative with 123I-meta-iodobenzylguanidine in patients with pheochromocytoma]. PMID- 11100672 TI - Colorectal cancer: treatment considerations. PMID- 11100671 TI - [Chronic renal failure as health problem: a timely reflexion]. PMID- 11100673 TI - Arthritis guidelines emphasize exercise. PMID- 11100674 TI - Reduced-fat fallacy. PMID- 11100675 TI - Breast cancer follow-up treatment not always optimal. PMID- 11100676 TI - Whole grains cut stroke risk. PMID- 11100677 TI - Bone up on fluoride, diuretics? PMID- 11100678 TI - Treating MS: better sooner than later. PMID- 11100679 TI - Mis-lead about calcium supplements? PMID- 11100680 TI - Exercise away the blues. PMID- 11100681 TI - New heart-healthy dietary guidelines. PMID- 11100682 TI - Fish oil for type 2 diabetes. PMID- 11100683 TI - Exercise for COPD. PMID- 11100684 TI - Olive oil and colon cancer. PMID- 11100685 TI - What causes a detached retina, and can it be prevented? PMID- 11100686 TI - What are antioxidants and how do they help fight disease? PMID- 11100687 TI - My mother recently had a stroke and was put on Dilantin (phenytoin) to prevent seizures. I've heard Dilantin can interfere with coordination and the ability to think clearly. Is there an alternative drug? PMID- 11100688 TI - Responses to Alferov. PMID- 11100690 TI - Partners edge closer to vital step on fusion. PMID- 11100689 TI - Anthropologists in turmoil over allegations of misconduct. PMID- 11100691 TI - French target research money at allaying BSE fears. PMID- 11100692 TI - Europe boosts genome resource centres...as German genomics gets cash windfall. PMID- 11100693 TI - Placebos could improve link between medical outlooks. PMID- 11100694 TI - Researchers fail to find signs of life in 'living' particles. PMID- 11100695 TI - Biotech report dubbed 'worthless'. PMID- 11100696 TI - Governments urged to rethink dam projects. PMID- 11100698 TI - Zhores Alferov: Russia's prize fighter. PMID- 11100697 TI - Aventis gets short shrift over release of modified corn. PMID- 11100699 TI - Evolutionary genetics: fruitfly centre spreads its wings. PMID- 11100700 TI - Revolution in references: give readers a chance by putting page numbers. PMID- 11100701 TI - Bridging the quality gap. PMID- 11100702 TI - The public has (rightly) lost faith in scientists. PMID- 11100703 TI - The all-chemist. PMID- 11100704 TI - Brain drain. PMID- 11100705 TI - Gaining light from silicon. PMID- 11100706 TI - Gene regulation. Local or global? PMID- 11100707 TI - Clipping the carbon-carbon bond. PMID- 11100708 TI - Evolutionary biology. Deja vu. PMID- 11100709 TI - Developmental biology. Control by combinatorial codes. PMID- 11100710 TI - Diabetes. Gene therapy for rats and mice. PMID- 11100711 TI - Molecular electronics. A dual-action material. PMID- 11100712 TI - Olympus/Nature competition. A 1, 2, 3 in light microscopy. PMID- 11100713 TI - Older bull elephants control young males. PMID- 11100714 TI - C66 fullerene encaging a scandium dimer. PMID- 11100715 TI - A stable non-classical metallofullerene family. PMID- 11100716 TI - The 'feathers' of Longisquama. PMID- 11100718 TI - The DNA damage response: putting checkpoints in perspective. AB - The inability to repair DNA damage properly in mammals leads to various disorders and enhanced rates of tumour development. Organisms respond to chromosomal insults by activating a complex damage response pathway. This pathway regulates known responses such as cell-cycle arrest and apoptosis (programmed cell death), and has recently been shown to control additional processes including direct activation of DNA repair networks. PMID- 11100717 TI - In search of the tumour-suppressor functions of BRCA1 and BRCA2. AB - Hereditary breast and ovarian cancer syndromes can be caused by loss-of-function germline mutations in one of two tumour-suppressor genes, BRCA1 and BRCA2 (ref. 1). Each gene product interacts with recombination/DNA repair proteins in pathways that participate in preserving intact chromosome structure. However, it is unclear to what extent such functions specifically suppress breast and ovarian cancer. Here we analyse what is known of BRCA gene function and highlight some unanswered questions in the field. PMID- 11100719 TI - Optical gain in silicon nanocrystals. AB - Adding optical functionality to a silicon microelectronic chip is one of the most challenging problems of materials research. Silicon is an indirect-bandgap semiconductor and so is an inefficient emitter of light. For this reason, integration of optically functional elements with silicon microelectronic circuitry has largely been achieved through the use of direct-bandgap compound semiconductors. For optoelectronic applications, the key device is the light source--a laser. Compound semiconductor lasers exploit low-dimensional electronic systems, such as quantum wells and quantum dots, as the active optical amplifying medium. Here we demonstrate that light amplification is possible using silicon itself, in the form of quantum dots dispersed in a silicon dioxide matrix. Net optical gain is seen in both waveguide and transmission configurations, with the material gain being of the same order as that of direct-bandgap quantum dots. We explain the observations using a model based on population inversion of radiative states associated with the Si/SiO2 interface. These findings open a route to the fabrication of a silicon laser. PMID- 11100720 TI - Evolution of the Sun's large-scale magnetic field since the Maunder minimum. AB - The most striking feature of the Sun's magnetic field is its cyclic behaviour. The number of sunspots, which are dark regions of strong magnetic field on the Sun's surface, varies with a period of about 11 years. Superposed on this cycle are secular changes that occur on timescales of centuries and events like the Maunder minimum in the second half of the seventeenth century, when there were very few sunspots. A part of the Sun's magnetic field reaches out from the surface into interplanetary space, and it was recently discovered that the average strength of this interplanetary field has doubled in the past 100 years. There has hitherto been no clear explanation for this doubling. Here we present a model describing the long-term evolution of the Sun's large-scale magnetic field, which reproduces the doubling of the interplanetary field. The model indicates that there is a direct connection between the length of the sunspot cycle and the secular variations. PMID- 11100721 TI - Coexistence of ferromagnetism and metallic conductivity in a molecule-based layered compound. AB - Crystal engineering--the planning and construction of crystalline supramolecular architectures from modular building blocks--permits the rational design of functional molecular materials that exhibit technologically useful behaviour such as conductivity and superconductivity, ferromagnetism and nonlinear optical properties. Because the presence of two cooperative properties in the same crystal lattice might result in new physical phenomena and novel applications, a particularly attractive goal is the design of molecular materials with two properties that are difficult or impossible to combine in a conventional inorganic solid with a continuous lattice. A promising strategy for creating this type of 'bi-functionality' targets hybrid organic/inorganic crystals comprising two functional sub-lattices exhibiting distinct properties. In this way, the organic pi-electron donor bis(ethylenedithio)tetrathiafulvalene (BEDT-TTF) and its derivatives, which form the basis of most known molecular conductors and superconductors, have been combined with molecular magnetic anions, yielding predominantly materials with conventional semiconducting or conducting properties, but also systems that are both superconducting and paramagnetic. But interesting bulk magnetic properties fail to develop, owing to the discrete nature of the inorganic anions. Another strategy for achieving cooperative magnetism involves insertion of functional bulky cations into a polymeric magnetic anion, such as the bimetallic oxalato complex [MnIICrIII(C2O4)3]-, but only insoluble powders have been obtained in most cases. Here we report the synthesis of single crystals formed by infinite sheets of this magnetic coordination polymer interleaved with layers of conducting BEDT-TTF cations, and show that this molecule-based compound displays ferromagnetism and metallic conductivity. PMID- 11100722 TI - Direct imaging of the pores and cages of three-dimensional mesoporous materials. AB - Mesostructured composite materials, with features ranging from 20 to 500 A in size, are obtained by the kinetically controlled competitive assembly of organic and inorganic species into nanostructured domains. Short-range order is limited, and long-range order is determined by weak forces such as van der Waals or hydrogen-bonding. Three-dimensional mesoporous materials obtained by removing the organic phase are of particular interest for applications such as catalysis and chemical sensing or separation, for which structural features such as cavity shape, connectivity and ordered bimodal porosity are critical. But atomic-scale structural characterization by the usual diffraction techniques is challenging for these partially ordered materials because of the difficulty in obtaining large (> 10 microm) single crystals, and because large repeat spacings cause diffraction intensities to fall off rapidly with scattering angle so that only limited small-angle data are available. Here we present a general approach for the direct determination of three-dimensional mesoporous structures by electron microscopy. The structure solutions are obtained uniquely without pre-assumed models or parametrization. We report high-resolution details of cage and pore structures of periodically ordered mesoporous materials, which reveal a highly ordered dual micro- and mesoscale pore structure. PMID- 11100723 TI - Improved estimates of global ocean circulation, heat transport and mixing from hydrographic data. AB - Through its ability to transport large amounts of heat, fresh water and nutrients, the ocean is an essential regulator of climate. The pathways and mechanisms of this transport and its stability are critical issues in understanding the present state of climate and the possibilities of future changes. Recently, global high-quality hydrographic data have been gathered in the World Ocean Circulation Experiment (WOCE), to obtain an accurate picture of the present circulation. Here we combine the new data from high-resolution trans oceanic sections and current meters with climatological wind fields, biogeochemical balances and improved a priori error estimates in an inverse model, to improve estimates of the global circulation and heat fluxes. Our solution resolves globally vertical mixing across surfaces of equal density, with coefficients in the range (3-12) x 10(-4) m2 s(-1). Net deep-water production rates amount to (15 +/- 12) x 10(6) m3 s(-1) in the North Atlantic Ocean and (21 +/- 6) x 10(6) m3 s(-1) in the Southern Ocean. Our estimates provide a new reference state for future climate studies with rigorous estimates of the uncertainties. PMID- 11100724 TI - The use of earthquake rate changes as a stress meter at Kilauea volcano. AB - Stress changes in the Earth's crust are generally estimated from model calculations that use near-surface deformation as an observational constraint. But the widespread correlation of changes of earthquake activity with stress has led to suggestions that stress changes might be calculated from earthquake occurrence rates obtained from seismicity catalogues. Although this possibility has considerable appeal, because seismicity data are routinely collected and have good spatial and temporal resolution, the method has not yet proven successful, owing to the non-linearity of earthquake rate changes with respect to both stress and time. Here, however, we present two methods for inverting earthquake rate data to infer stress changes, using a formulation for the stress- and time dependence of earthquake rates. Application of these methods at Kilauea volcano, in Hawaii, yields good agreement with independent estimates, indicating that earthquake rates can provide a practical remote-sensing stress meter. PMID- 11100725 TI - Concurrent density dependence and independence in populations of arctic ground squirrels. AB - No population increases without limit. The processes that prevent this can operate in either a density-dependent way (acting with increasing severity to increase mortality rates or decrease reproductive rates as density increases), a density-independent way, or in both ways simultaneously. However, ecologists disagree for two main reasons about the relative roles and influences that density-dependent and density-independent processes have in determining population size. First, empirical studies showing both processes operating simultaneously are rare. Second, time-series analyses of long-term census data sometimes overestimate dependence. By using a density-perturbation experiment on arctic ground squirrels, we show concurrent density-dependent and density independent declines in weaning rates, followed by density-dependent declines in overwinter survival during hibernation. These two processes result in strong, density-dependent convergence of experimentally increased populations to those of control populations that had been at low, stable levels. PMID- 11100726 TI - Variation in the reversibility of evolution. AB - How reversible is adaptive evolution? Studies of microbes give mixed answers to this question. Reverse evolution has been little studied in sexual populations, even though the population genetics of sexual populations may be quite different. In the present study, 25 diverged replicated populations of Drosophila melanogaster are returned to a common ancestral environment for 50 generations. Here we show that reverse evolution back to the ancestral state occurs, but is not universal, instead depending on previous evolutionary history and the character studied. Hybrid populations showed no greater tendency to undergo successful reverse evolution, suggesting that insufficient genetic variation was not the factor limiting reverse evolution. Adaptive reverse evolution is a contingent process which occurs with only 50 generations of sexual reproduction. PMID- 11100727 TI - Integration of target and body-part information in the premotor cortex when planning action. AB - To plan an action, we must first select an object to act on and the body part (or parts) to use to accomplish our intention. To plan the motor task of reaching, we specify both the target to reach for and the arm to use. In the process of planning and preparing a motor task, information about the motor target and the arm to use must be integrated before a motor program can be formulated to generate the appropriate limb movement. One of the structures in the brain that is probably involved in integrating these two sets of information is the premotor area in the cerebral cortex of primates. The lateral sector of the dorsal premotor cortex is known to receive both visual and somatosensory input, and we show here that neurons in this area gather information about both the target and the body part, while subsequent activity specifies the planned action. PMID- 11100728 TI - MOD-1 is a serotonin-gated chloride channel that modulates locomotory behaviour in C. elegans. AB - The neurotransmitter and neuromodulator serotonin (5-HT) functions by binding either to metabotropic G-protein-coupled receptors (for example, 5-HT1, 5-HT2, 5 HT4 to 5-HT7), which mediate 'slow' modulatory responses through numerous second messenger pathways, or to the ionotropic 5-HT3 receptor, a non-selective cation channel that mediates 'fast' membrane depolarizations. Here we report that the gene mod-1 (for modulation of locomotion defective) from the nematode Caenorhabditis elegans encodes a new type of ionotropic 5-HT receptor, a 5-HT gated chloride channel. The predicted MOD-1 protein is similar to members of the nicotinic acetylcholine receptor family of ligand-gated ion channels, in particular to GABA (gamma-aminobutyric acid)- and glycine-gated chloride channels. The MOD-1 channel has distinctive ion selectivity and pharmacological properties. The reversal potential of the MOD-1 channel is dependent on the concentration of chloride ions but not of cations. The MOD-1 channel is not blocked by calcium ions or 5-HT3a-specific antagonists but is inhibited by the metabotropic 5-HT receptor antagonists mianserin and methiothepin. mod-1 mutant animals are defective in a 5-HT-mediated experience-dependent behaviour and are resistant to exogenous 5-HT, confirming that MOD-1 functions as a 5-HT receptor in vivo. PMID- 11100729 TI - Notch signalling and the synchronization of the somite segmentation clock. AB - In vertebrates with mutations in the Notch cell-cell communication pathway, segmentation fails: the boundaries demarcating somites, the segments of the embryonic body axis, are absent or irregular. This phenotype has prompted many investigations, but the role of Notch signalling in somitogenesis remains mysterious. Somite patterning is thought to be governed by a "clock-and wavefront" mechanism: a biochemical oscillator (the segmentation clock) operates in the cells of the presomitic mesoderm, the immature tissue from which the somites are sequentially produced, and a wavefront of maturation sweeps back through this tissue, arresting oscillation and initiating somite differentiation. Cells arrested in different phases of their cycle express different genes, defining the spatially periodic pattern of somites and controlling the physical process of segmentation. Notch signalling, one might think, must be necessary for oscillation, or to organize subsequent events that create the somite boundaries. Here we analyse a set of zebrafish mutants and arrive at a different interpretation: the essential function of Notch signalling in somite segmentation is to keep the oscillations of neighbouring presomitic mesoderm cells synchronized. PMID- 11100730 TI - Binding of disease-associated prion protein to plasminogen. AB - Transmissible spongiform encephalopathies are associated with accumulation of PrP(Sc), a conformer of a cellular protein called PrP(C). PrP(Sc) is thought to replicate by imparting its conformation onto PrP(C) (ref. 1), yet conformational discrimination between PrP(C) and PrP(Sc) has remained elusive. Because deposition of PrP(Sc) alone is not enough to cause neuropathology, PrP(Sc) probably damages the brain by interacting with other cellular constituents. Here we find activities in human and mouse blood which bind PrP(Sc) and prion infectivity, but not PrP(C). We identify plasminogen, a pro-protease implicated in neuronal excitotoxicity, as a PrP(Sc)-binding protein. Binding is abolished if the conformation of PrP(Sc) is disrupted by 6M urea or guanidine. The isolated lysine binding site 1 of plasminogen (kringles I-III) retains this binding activity, and binding can be competed for with lysine. Therefore, plasminogen represents the first endogenous factor discriminating between normal and pathological prion protein. This unexpected property may be exploited for diagnostic purposes. PMID- 11100731 TI - Remission in models of type 1 diabetes by gene therapy using a single-chain insulin analogue. AB - A cure for diabetes has long been sought using several different approaches, including islet transplantation, regeneration of beta cells and insulin gene therapy. However, permanent remission of type 1 diabetes has not yet been satisfactorily achieved. The development of type 1 diabetes results from the almost total destruction of insulin-producing pancreatic beta cells by autoimmune responses specific to beta cells. Standard insulin therapy may not maintain blood glucose concentrations within the relatively narrow range that occurs in the presence of normal pancreatic beta cells. We used a recombinant adeno-associated virus (rAAV) that expresses a single-chain insulin analogue (SIA), which possesses biologically active insulin activity without enzymatic conversion, under the control of hepatocyte-specific L-type pyruvate kinase (LPK) promoter, which regulates SIA expression in response to blood glucose levels. Here we show that SIA produced from the gene construct rAAV-LPK-SIA caused remission of diabetes in streptozotocin-induced diabetic rats and autoimmune diabetic mice for a prolonged time without any apparent side effects. This new SIA gene therapy may have potential therapeutic value for the cure of autoimmune diabetes in humans. PMID- 11100732 TI - A ubiquitin-like system mediates protein lipidation. AB - Autophagy is a dynamic membrane phenomenon for bulk protein degradation in the lysosome/vacuole. Apg8/Aut7 is an essential factor for autophagy in yeast. We previously found that the carboxy-terminal arginine of nascent Apg8 is removed by Apg4/Aut2 protease, leaving a glycine residue at the C terminus. Apg8 is then converted to a form (Apg8-X) that is tightly bound to the membrane. Here we report a new mode of protein lipidation. Apg8 is covalently conjugated to phosphatidylethanolamine through an amide bond between the C-terminal glycine and the amino group of phosphatidylethanolamine. This lipidation is mediated by a ubiquitination-like system. Apg8 is a ubiquitin-like protein that is activated by an E1 protein, Apg7 (refs 7, 8), and is transferred subsequently to the E2 enzymes Apg3/Aut1 (ref. 9). Apg7 activates two different ubiquitin-like proteins, Apg12 (ref. 10) and Apg8, and assigns them to specific E2 enzymes, Apg10 (ref. 11) and Apg3, respectively. These reactions are necessary for the formation of Apg8-phosphatidylethanolamine. This lipidation has an essential role in membrane dynamics during autophagy. PMID- 11100733 TI - G alpha(i) and G alpha(o) are target proteins of reactive oxygen species. AB - Reactive oxygen species (ROS) have been identified as central mediators in certain signalling events. In the heart, ROS have important functions in ischaemia/reperfusion-induced cardiac injury and in cytokine-stimulated hypertrophy. Extracellular signal-regulated kinase (ERK) is one of the ROS responsive serine/threonine kinases. Previous studies showed that tyrosine kinases and small G proteins are involved in the activation of ERK by ROS; however, the initial target protein of ROS that leads to ERK activation remains unknown. Here we show that inhibition of the betagamma-subunit of G protein (G betagamma) attenuates hydrogen peroxide (H2O2)-induced ERK activation in rat neonatal cardiomyocytes. The G betagamma-responsive ERK activation induced by H2O2 is independent of ligands binding to Gi-coupled receptors, but requires phosphatidylinositol-3-kinase and Src activation. In in vitro studies, however, treatment with H2O2 increases [35S]GTP-gammaS binding to cardiac membranes and directly activates purified heterotrimeric Gi and Go but not Gs. Analysis using heterotrimeric Go and its individual subunits indicates that H2O2 modifies G alpha(o) but not G betagamma, which leads to subunit dissociation. We conclude that G alpha(i) and G alpha(o) are critical targets of oxidative stress for activation of ERK. PMID- 11100734 TI - Global histone acetylation and deacetylation in yeast. AB - Histone acetyltransferases and deacetylases can be targeted to promoters to activate or repress genes. For example, the histone acetyltransferase GCN5 is part of a yeast multiprotein complex that is recruited by the DNA-binding activator protein GCN4 (refs 1-3). The histone deacetylase RPD3 complex is recruited to DNA by the repressor UME6 (refs 4, 5); similar mechanisms exist in other eukaryotes. However, deletion of RPD3 also increases expression of the PHO5 gene that is repressed by nucleosomes, and regulated by GCN5 (ref. 10) but not by UME6. We have determined whether acetylation and deacetylation are promoter specific at PHO5, by using antibodies against acetylated lysine residues and chromatin immunoprecipitation to examine the acetylation state of a 4.25-kilobase region surrounding the PHO5 gene. Here we show that this region is acetylated extensively by ESA1 and GCN5 and deacetylated by HDA1 and RPD3, and that widespread histone modification affects three separate chromosomal regions examined, which total 22kb. Our data indicate that targeted modification occurs in a background of global acetylation and deacetylation that not only reduces basal transcription, but also allows a rapid return to the initial state of acetylation when targeting is removed. PMID- 11100735 TI - Hyaluronan is essential for the expansion of the cranial base growth plates. AB - Exquisite control of chondrocyte function in the zone of hypertrophy results in expansive growth of cartilaginous growth plates, and is a prerequisite for normal skeletal lengthening. We hypothesize that hyaluronan-mediated hydrostatic pressure causes lacunae expansion in the zone of hypertrophy; an important mechanism in cartilaginous growth plate and associated skeletal expansion. The role of hyaluronan and CD44 in this mechanism was studied using organ culture of the bipolar cranial base synchondroses. Hyaluronan was present in the hypertrophic zones, pericellular to the hypertrophic chondrocytes, while no hyaluronan was detected in the resting, proliferating and maturing zones. This localization of hyaluronan was associated with increased lacunae size, suggesting that chondrocytes deposit and retain pericellular hyaluronan as they mature. In comparison, Toluidine Blue staining was associated with the territorial matrix. Hyaluronidase, the hyaluronan-degrading enzyme, and CD44, the receptor for hyaluronan which also participates in the uptake and degradation of hyaluronan, were co-localized within the zone of ossification. This pattern of expression suggests that cells in the early zone of ossification internalize and degrade hyaluronan through a CD44-mediated mechanism. Treatment of the cultured segments with either Streptomyces hyaluronidase or hyaluronan hexasaccharides inhibited lacunae expansion. These observations demonstrate that hyaluronan-mediated mechanisms play an important role in controlling normal skeletal lengthening. PMID- 11100736 TI - Vertical regulation of En-2 expression and eye development by FGFs and BMPs. AB - The formation of the midbrain region depends mainly on the activity of a signalling center located in the isthmus region, on the border between the prospective mesencephalon and metencephalon. FGF-8 has been proposed as a signalling molecule responsible for this specification because of its expression pattern and its ability to elicit duplication of the midbrain region when expressed ectopically in the neuroepithelium. Here we present evidence that members of the FGF family of growth factors when released in the cephalic mesenchyme are able to extend the expression of the mesencephalic marker En-2 to both the anterior and the posterior regions of its original landmark. This alteration in the expression pattern of En-2 is not accompanied by a significant alteration in the later development of the midbrain-cerebellar anlage, although the eye development is severely altered. Members of the bone morphogenetic protein family ectopically released from the mesenchyme down-regulate the expression of En-2 and also have an effect on the development of the eye. These results demonstrate that growth factor molecules produced in the mesenchyme (vertical signalling) participate in the correct establishment of the antero posterior patterning of the cephalic nervous system during development. PMID- 11100737 TI - Influences of osteoclast deficiency on craniofacial growth in osteopetrotic (op/op) mice. AB - It is well known that the defect in bone resorption in osteopetrotic (op/op) mice brings about deformation of the cranium and failure of tooth eruption. However, the influences on longitudinal growth of the craniofacial skeleton have not been elucidated. This study was thus conducted to examine craniofacial morphology and longitudinal changes in the op/op mice by means of morphometric analysis with lateral cephalograms. Lateral cephalograms, taken every 10 days from 10- to 90 day-old mice, were analyzed on a personal computer for 11 measurement items. For the nasal bone region, the most prominent differences were found between the op/op and normal mice. The anterior cranial base and occipital bone height presented almost equivalent growth changes in both the op/op and normal mice. The size of mandible, meanwhile, was significantly smaller in the op/op mice than in the normal controls. The gonial angle was also significantly larger in the op/op mice than in the normal mice throughout the experimental period. Thus, substantial differences in craniofacial growth were demonstrated in various areas of the craniofacial complex, which are assumed essentially due to the lack of osteoclastic bone resorption during growing period. Since the difference became more prominent in the anatomic regions relevant to the masticatory functions, it would be a reasonable assumption that reduced masticatory function is also a key determinant for the less-developed craniofacial skeleton in the op/op mouse. PMID- 11100738 TI - Merging the old skeletal biology with the new. I. Intramembranous ossification, endochondral ossification, ectopic bone, secondary cartilage, and pathologic considerations. AB - Skeletogenesis and chondrogenesis result from a sequence of events involving epithelial-mesenchymal interaction, condensation, and differentiation. Types of bone and cartilage formation include: (1) intramembranous ossification, (2) endochondral ossification, (3) combined endochondral and intramembranous ossification, (4) heterotopic bone and cartilage formation, and (5) secondary cartilage formation. Pathologic conditions with bone and cartilage include: (1) benign and malignant tumors and (2) reactive osseous and cartilaginous metaplasia. PMID- 11100739 TI - Merging the old skeletal biology with the new. II. Molecular aspects of bone formation and bone growth. AB - Molecular aspects of bone formation and bone growth are discussed together with selected genetic disorders of the involved genes. Topics covered include: collagenopathies and osteogenesis imperfecta; core binding factor transcription factors and cleidocranial dysplasia; bone morphogenetic proteins and fibrodysplasia ossificans progressiva; transforming growth factor beta, suture closure, and craniosynostosis; Indian hedgehog, parathyroid hormone-related protein together with its receptor, and Jansen metaphyseal chondrodysplasia. PMID- 11100740 TI - Subsite specificity of divalent metal ions to glucosyltransferase. AB - Glucosyltransferase from oral bacteria Streptococcus mutans is the most significant virulent factor in causing dental caries. The enzyme has two subsites. The binding specificity of divalent metal ions to glucosyl or fructosyl subsite was examined using multiple inhibition kinetics. The interaction factor "alpha" identifies whether the two subsites are exclusive or non-exclusive. PMID- 11100741 TI - Pfeiffer syndrome is not caused by haploinsufficient mutations of FGFR2. PMID- 11100742 TI - Long-term survival after induction therapy with idarubicin and cytosine arabinoside for de novo acute myeloid leukemia. AB - We treated 153 patients with de novo acute myeloid leukemia (AML) with two induction courses of conventional-dose cytosine arabinoside (ara-C) and idarubicin (AIDA) followed by either a third course of AIDA, high-dose ara-C or bone-marrow transplantation. The complete remission (CR) rate for all patients was 63.4%, with a higher CR rate for patients with a normal (versus unfavorable) karyotype (73.2% vs 52.5%; P=0.038). The probability of overall survival (OS) was 30.7% after 5 years (26.3% after 7 years). Improved OS at 5 years could be observed for patients up to 50 years old versus patients older than 50 years of age (37.6% vs 19.9%; P=0.001) and patients with a normal (versus unfavorable) karyotype (42.9% vs 14.1%; P=0.0016). Disease-free survival (DFS) after 5 years was 33.2% for all 97 CR patients and was significantly better for patients with a normal (versus unfavorable) karyotype (44.3% vs 12.3%; P= 0.003). Multivariate analysis revealed that the age for OS (P < 0.02) and the karyotype for both OS (P<0.03) and DFS (P< 0.05) were independent prognostic factors. In conclusion, AIDA is an effective and well-tolerated induction regimen (even in elderly patients) with a 5-year survival of more than 30% when combined with ara-C containing postremission therapy. The karyotype is the most powerful prognostic factor for predicting the outcome of patients treated with this protocol. PMID- 11100743 TI - Flow-cytometric detection of minimal residual disease with atypical antigen combinations in patients with de novo acute myeloid leukemia. AB - The immunophenotypic features in adult de novo acute myeloid leukemia (AML) patients at diagnosis using flow cytometry double marker analysis and the detection of minimal residual disease with atypical leukemia-associated antigen combinations during remission were investigated. Fifty adult patients with de novo AML at diagnosis were studied. Bone marrow samples from 21 patients with AML were analyzed upon achievement of complete remission and during continuous complete remission. Ten bone marrow samples of normal donors were also studied. CD34/CD13, CD34/CD33, CD33/CD7, CD33/CD10, CD33/CD19 and CD33/TdT are the leukemia-associated antigen combinations used for the detection of minimal residual disease. The outcome of 19 patients has been evaluated. Of these 19 patients, 10 were found to be in immunophenotypic remission (median follow-up after the study: 837 days, range 620-1343 days). Only one patient in this group has relapsed so far. In the other nine patients residual disease was detected. Seven of these patients developed systemic relapse following a median follow-up time of 86 days after the study (range 34-273 days), one received allogeneic bone marrow transplantation 70 days after the study, and another has been in complete remission and off chemotherapy for 36 months. The presence of cells with atypical antigen combinations identified at diagnosis in certain patients is valuable for monitoring the disease in remission. The persistence of such a population in remission has indicated the impending relapse in this study. PMID- 11100744 TI - High-dose therapy with peripheral blood stem cell transplantation for patients with relapsed or refractory Hodgkin's disease: long-term outcome and prognostic factors. AB - From March 1986 to March 1998, 82 patients with relapsed or refractory Hodgkin's disease underwent high-dose chemotherapy (HDCT) with peripheral blood stem cell (PBSC) transplantation in our center. This is a retrospective analysis of the long-term clinical outcome. There were 52 males and 30 females with a median age of 32 years (range 18-59 years). Prior to transplantation, 36 patients were in complete remission (CR), 34 in partial remission (PR), and 12 had refractory disease after salvage therapy. For HDCT, 78 patients were treated with CBV (cyclophosphamide, 6.0-6.8 g/ m2; etoposide, 1.0-1.6 g/m2; carmustine, 0.45-0.8 g/m2), while four patients received different regimens. Probability of freedom from progression (FFP), overall survival (OS), and event-free survival (EFS) at 5 years of the entire group was 63%, 61%, and 54%, respectively. Early mortality rate ( < or = 100 days) declined from 17% to 6% after 1992. Five patients died of late transplant-related complications (> 100 days), including secondary lymphoma and leukemia in two patients. None of the refractory patients survived beyond 3.5 years. Multivariate analyses identified extranodal sites of disease at relapse and refractory disease status prior to transplantation as significant prognostic factors for FFP, EFS, and OS. As we have shown in our study, remarkable progress was achieved in reducing early morbidity and mortality over time, but this was associated with only a slight, not significant improvement of long-term outcome (OS 66% vs 57% at 5 years for patients undergoing PBSC transplantation before and after 1992, P = 0.26). Although the results as a whole are encouraging for chemosensitive patients, new therapeutic strategies are needed to reduce toxicity and improve the clinical outcome of patients, especially of those with a less favorable prognosis. PMID- 11100745 TI - Fludarabine therapy in Waldenstrom's macroglobulinemia. AB - Seven patients with macroglobulinemia (six previously untreated, one with minimal pretreatment) were treated with fludarabine (25 mg/m2/day for 5 days, repeated every 4 weeks). The median age was 58 years. The time from diagnosis to treatment with fludarabine was 4.5 months to 175 months (median 32.6 months). The patients received six (n =5), five (n =1), and three (n = 1) courses of fludarabine. One patient showed only a slight decrease of immunoglobulin (Ig) M (from 5,750 mg/dl to 4,700 mg/dl) and no improvement of anemia. Therefore, treatment was stopped after three cycles. In the other six patients, a marked reduction of IgM levels (from 6,140 mg/dl to 1,220 mg/dl median), a normalization of hemoglobin (from 10.8 g/dl to 12.3 g/dl median), a reduction of lymphocyte count (from 1992/>microl to 652/microl median), and a reduction of beta2 microglobulin (from 2.3mg/l to 1.8 mg/l median) were achieved. A 50% IgM reduction was achieved 5.4 months (median) after the beginning of therapy, and the maximum response was observed 17.3 months (median) after the end of treatment. The responses were sustained without further therapy in six patients for 20.8-55.2 months. In one patient, disease progression was observed 12.5 months after the end of therapy. Fludarabine therapy was well tolerated with few side effects. In three patients, febrile episodes occurred. No opportunistic infections were recorded. We conclude that fludarabine is an effective treatment in previously untreated or in minimally pretreated patients with Waldenstrom's macroglobulinemia. PMID- 11100746 TI - Characteristics of the specific removal of lymphocytes and granulocytes from whole blood in an automated bottom-and-top processing system. AB - Characteristics of lymphocyte and granulocyte removal were studied during processing of fresh and overnight-stored human blood in a bottom-and-top semiautomated system using the buffy coat (BC) technique. Blood cells were counted in the resulting components. Removal efficiency of lymphocytes and granulocytes correlated with the loss of erythrocytes due to removal of the BC. Sigmoidal curves showed a good fit to experimental data. Variables of the equations differed substantially concerning removal of lymphocytes and granulocytes. Overnight storage of blood at 20-22 degrees C prior to processing resulted in changes in efficiency of leukocyte removal. Removal of lymphocytes decreased, while that of granulocytes increased due to overnight processing. Lymphocytes may be removed almost quantitatively with less than 10% (fresh blood) or 15% (overnight-stored blood) loss in erythrocyte content of the donated blood, corresponding to less than 28 ml packed red cells in the BC. Granulocytes seem to represent the residual amount of the final leukocyte content in the erythrocyte product. The present study may contribute to the production of erythrocyte products with fewer contaminating leukocytes and to the standardizing of semiautomated blood processing. PMID- 11100747 TI - Morphologically defined myeloid cell compartments, lymphocyte subpopulations, and histological findings of bone marrow in patients with nonimmune chronic idiopathic neutropenia of adults. AB - This report describes the morphologically defined myeloid cell compartments, lymphocyte subpopulations, and histological findings of bone marrow in 38 patients with nonimmune chronic idiopathic neutropenia of adults (NI-CINA) and in 14 controls. We found that patients had a striking shift to the left of the granulocytic series due to both an increased proportion of proliferating cells and a reduced proportion of maturating cells compared with controls (P<0.001 and P<0.001, respectively). Individual proportions of these cells strongly correlated with the number of circulating neutrophils (r = -0.462, P < 0.01 and r = 0.495, P<0.01, respectively). However, in the great majority of patients (78.9%), no significant changes in marrow cellularity or the myeloid to erythroid cell ratio could be demonstrated. Patients also had increased proportions of CD19+B cells, CD20+B cells, and plasma cells with polytypic expression relative to controls (P < 0.02, P< 0.01, and P< 0.001, respectively). Individual values of plasma cells were inversely correlated with the number of blood neutrophils (r=-0.414, P<0.01). Dispersed bcl-2+lymphocytic aggregates without germinal centers were seen in about one-third of the patients. T cells and natural killer (NK) cells did not show any significant change. Patients had increased proportions of CD57+, CD16+, and HLA-DR+ cells and, in a few cases, increased proportions of histiocytes and eosinophils. CD45RO+ cells were reduced only in patients with pronounced neutropenia. Expression of p53 protein has not been detected in any cell population. With the exception of some megaloblastoid features of erythroid lineage seen in two patients and the presence of some micromegacaryocytes seen in two others, no significant morphological abnormalities were noted. All of these findings are consistent with our previously reported suggestion for the possible existence of an underlying low-grade chronic inflammatory process in NI-CINA patients, which may be involved in the pathogenesis of neutropenia in the affected subjects. PMID- 11100748 TI - Pregnancy outcome of a transfusion-dependent thalassemic woman. AB - A clinical case concerning a normal pregnancy outcome in a transfusion-dependent woman affected by homozygous beta thalassemia, whose partner was negative with regard to the "thalassemic trait", was reported. The patient showed no iron deposit problems, viral diseases that could have made the pregnancy management difficult or any complications during the gestation. Blood transfusion was not necessary during the following caesarean delivery. The outcome was a healthy female child, born at a gestational age of 38 weeks, showing neither malformations nor problems. This was possible due to a detailed preconceptual guidance and a pre-pregnancy assessment. The patient normally would have had a blood transfusion every 20 days and a strict desferrioxamine chelating therapy; however, this treatment was suspended during her pregnancy because of the well recognised teratogenic effects of the drug. The average values of ferritin were just a little higher than before being pregnant. The foetus, due to her particular chelating activity, probably maintained these ferritin levels. A sample of 95 ml umbilical cord blood was taken during the delivery. It is well known that umbilical cord blood contains a good quantity of CD34+ stem cells, the haematopoietic progenitors. It was therefore collected for transplanting to the mother and for bone marrow reconstitution. Moreover, our experience suggests that desferrioxamine therapy during lactation does not alter iron excretion in breast milk. Therefore, women now affected by Cooley disease may possibly have a normal pregnancy without ovulation induction, intrauterine growth retardation, foetal loss and preterm labour. PMID- 11100749 TI - Bone marrow microvessel density is a prognostic factor for survival in patients with multiple myeloma. AB - The importance of neoangiogenesis for the progressive growth and viability of solid tumors is well established. Recently, there has been growing evidence that angiogenesis might also be important in hematological malignancies, but only few data are available. In this report, we have studied the impact of bone marrow microvessel density and survival in patients with multiple myeloma (MM). Immunohistochemical CD34 stained paraffin-embedded bone marrow biopsies of 44 patients with newly diagnosed MM were studied. Microvessels were counted in 400 x magnification and the mean number of vessels per area in each sample was noted as the microvessel density (MVD). The median MVD was 48 vessels/mm2, the range was 0 125 vessels/mm2. Using a cut-off value of the median MVD in the Kaplan-Meier analysis, the median survival was 22.2 months in the group with the higher MVD and was not reached in the group with the lower MVD (P< 0.01). In a multivariate Cox regression analysis, using previously identified prognostic factors beta2 microglobulin, C-reactive protein (CRP), and age, MVD remained significant as a prognostic factor (P< 0.03). PMID- 11100750 TI - Allogeneic bone marrow transplantation for refractory mantle cell lymphoma. AB - We report about a 28-year-old woman with relapsed mantle cell lymphoma (MCL, centrocytic lymphoma according to the Kiel classification) refractory to salvage chemotherapy. The patient underwent allogeneic bone marrow transplantation from a HLA-identical brother after myeloablative chemotherapy consisting of busulfan, etoposide, and cyclophosphamide. The patient experienced hepatic toxicity (grade I), mucositis (grade II) according the Bearman scale, and graft versus host disease of the skin (grade II) and showed stable engraftment with complete chimerism on day 15 after bone marrow transplantation. Eight years after transplantation, the patient is still disease free and in good condition without any late side effects. This report suggests a curative potential of allogeneic stem cell transplantation in MCL. PMID- 11100751 TI - Angiotropic B-cell lymphoma with hemophagocytic syndrome associated with syndrome of inappropriate secretion of antidiuretic hormone. AB - A case of angiotropic B-cell lymphoma associated with hemophagocytic syndrome (HPS) has been reported. In addition to fever, pancytopenia, hepatosplenomegaly, and lack of lymphadenopathy, unique clinical features, such as syndrome of inappropriate secretion of antidiuretic hormone (SIADH) and pulmonary infarction, were manifested. Both soluble interleukin-2 receptor (sIL-2R) and IL-6 were elevated in the patient's sera in addition to an increase of serum lactate dehydrogenase and ferritin. In contrast, tumor necrosis factor-alpha and interferon-gamma were within normal ranges. Serum antibodies against Epstein-Barr virus and cytomegalovirus showed a past infection pattern. An autopsy examination revealed systemic intravascular proliferation of lymphoma cells with a B-cell phenotype, confirming the diagnosis of angiotropic B-cell lymphoma. Moreover, SIADH was suggested to result from the infiltration of tumor cells into the pituitary gland. Triple association of angiotropic B-cell lymphoma, HPS and SIADH is quite rare. Therefore, the present case seems to be helpful for clarifying the mechanism for HPS of non-Hodgkin's lymphoma with B-cell origin. PMID- 11100752 TI - Sweet's syndrome and accelerated phase of chronic myelogenous leukemia. AB - Sweet's syndrome is a neutrophilic dermatosis characterized clinically by raised, erythematous, tender lesions on the face, neck, upper thorax and extremities. Several diseases have been associated with this entity. We report a case of Sweet's syndrome associated with chronic myelogenous leukemia: a 48-year-old woman who developed recurrent skin lesions 3 years after the diagnosis of chronic myelogenous leukemia. Progression to an accelerated phase of the disease was detected 3 months after the beginning of the skin lesions. This case shows the convenience of evaluation and closer follow-up of patients with chronic myelogenous leukemia who develop skin lesions, especially if these lesions are recurrent. PMID- 11100753 TI - Fatal cerebral hemorrhage associated with cyclosporin-A/FK506-related encephalopathy after allogeneic bone marrow transplantation. AB - We report a case of cerebral hemorrhage associated with cyclosporin A (CsA)/FK506 related encephalopathy that developed in a 16-year-old woman after allogeneic bone marrow transplantation. Hematopoietic engraftment occurred on day 15, and the patient developed systemic convulsions after CsA was replaced by FK506 for the treatment of acute graft-versus-host disease (GVHD). Based on magnetic resonance imaging, laboratory findings and cerebrospinal fluid studies, she was diagnosed as having CsA/FK506-related encephalopathy with cerebral hemorrhagic infarction. Although she recovered completely after discontinuation of FK506, she developed convulsions again 15 days after re-administration of FK506. A computed tomography scan showed cerebral hemorrhage. She died of respiratory failure. Vascular damage induced by immunosuppressive drugs and enhanced by acute GVHD seemed to be the cause of the cerebral hemorrhage. Since hypertension, which was present during both of the central nervous system events, seemed to have contributed to the development of the cerebral hemorrhage, it is proposed that CsA and FK506 should be reduced or discontinued when patients who have risk factors of hypertension become hypertensive even if they have no symptoms of neurotoxicity. PMID- 11100754 TI - Thrombotic thrombocytopenia purpura caused by piperacillin successfully treated with plasma infusion. AB - An 81-year-old man was admitted to the hospital with a fever and loss of appetite. After treatment with piperacillin sodium (PIPC), the patient exhibited thrombocytopenia, hemorrhagic colitis, and drug-induced skin eruption. On the fifth day after PIPC induction, he further experienced neurological abnormalities, such as disorientation and confusion, renal dysfunction, and microangiopathic hemolytic anemia (MAHA). The patient was diagnosed with thrombotic thrombocytopenic purpura (TTP) on the basis of thrombocytopenia, MAHA, renal dysfunction, fever, and neurological abnormalities. Infusion of fresh frozen plasma was initiated for treatment. His condition improved markedly after this treatment. It is rare for TTP to be accompanied with hemorrhagic colitis and skin eruption. These symptoms were induced by PIPC and were successfully treated with plasma infusion. PMID- 11100755 TI - Transient polycythemia and multi-organ failure due to acute alcohol ingestion. AB - Relative polycythemia is characterized by elevated hematocrit with normal red cell mass and results from decreased plasma volume. We present a case of a 39 year-old man who had at least two episodes of severe relative polycythemia and multi-organ failure following acute alcohol ingestion. Although the acute dehydrating effects of alcohol are well known, they usually result in an indolent course. This is the first report of recurrent severe polycythemia and multi-organ failure following acute alcohol consumption. PMID- 11100756 TI - Pathomorphological and histological analysis of the indirect revascularization method. AB - OBJECTIVE: This experimental study was initiated to determine whether TMLR may prevent porcine myocardium from ischemia and necrosis after acute myocardial infarction. In addition, the influence of TMLR on healthy myocardium was analyzed. METHODS: The short-term effectiveness of TMLR was evaluated in 38 open chest anesthetized pigs with (n = 18) or without (n = 20) acute LAD occlusion (observation period 6 hours): Six pigs served as controls (thoracotomy only). An additional six pigs had LAD occlusion only (ischemic group). A subsequent 12 pigs were treated by TMLR (CO2) prior to LAD occlusion: Six pigs received one laser channel/cm2 (group 1) and in six pigs two channels/cm2 in the LAD territory (group 2) were performed. In addition, 14 pigs underwent TMLR without ischemia: Seven pigs received 1 channel/cm2 (group 3) and seven pigs 2 channels/cm2 (group 4). Pathomorphological assessment and histology were performed. RESULTS: TMLR limits the expansion of the myocardial infarction zone: laser group 2 demonstrated a significantly smaller area of necrosis in the area at risk (ischemic group (32%) vs. laser group 1 (18%, p = ns) and 2 (8%, p = 0.0076); laser group 1 vs. 2, p = 0.0056). The amount of the area of necrosis of laser groups 3 (0.4%) and 4 (0.04%) compared to control (0%) did not differ significantly (p = ns). Furthermore, in the lased territories of laser groups 3 and 4 microscopic analysis revealed signs of ischemia in 10 +/- 30.9% of all examined histological samples (p = ns vs. control). During a short coronary occlusion the laser-induced tracks were partially filled with blue dye in 94.8 +/ 27.0/85.9 +/- 34.3/94.85 +/- 22.0%/70.21 +/- 47.0% (laser groups 1 - 4 respectively p = ns) The myocardial water content-measurements (MWC) of the ischemia and laser group 1 were not different at the end of the experiment (p = ns). In contrast, laser groups 2, 3 and 4 revealed significantly higher MWC values compared to control (p = 0.036, p < 0.001, p < 0.001; respectively). CONCLUSIONS: This prolonged acute study demonstrates that preventive CO2-laser revascularization significantly reduces the amount of necrosis in the area at risk. Histological examination supported the idea that some pigment gained access to the ischemic tissue via patent channels. In healthy myocardium, TMLR significantly increases myocardial water content and induces non-significant small ischemic and very small necrotic areas surrounding open laser channels. PMID- 11100757 TI - Uncontrolled reoxygenation by initiating cardiopulmonary bypass is associated with higher protein S100 in cyanotic versus acyanotic patients. AB - BACKGROUND: The systemic reoxygenation injury produced by initiating cardiopulmonary bypass (CPB) in infants with cyanotic heart disease may be associated with cerebral dysfunction and injury. Increased protein S100 (S100) serum levels may indicate cerebral and blood brain barrier damage as well as inflammatory changes, therefore serving to quantify these changes. The present clinical study assessed S100 in cyanotic patients undergoing CPB with normoxic versus hyperoxic paO2 in acyanotic cases and in controls without CPB. METHODS: 43 patients with congenital heart disease aged 5 days to 15 years (mean 4.4 years) were enrolled consecutively and divided in four groups: (1) Cyanotic infants undergoing controlled normoxic reoxygenation on CPB (n = 12), (2) cyanotic infants undergoing uncontrolled hyperoxic reoxygenation on CPB (n = 9), (3) acyanotic infants operated with CPB (n = 16) and (4) patients operated without CPB (n = 6). Blood samples were collected after induction of anesthesia (A), up to 4 hours after surgery (B) and at postoperative day one (C). RESULTS: Preoperative S100 serum levels [microg/l] in all groups were below clinical relevance. S100 increased markedly after surgery in groups 1 and 2. Differences in postoperative S100 levels were significant between groups 1 (0.45 +/- 0.13) and 3 (0.35 +/- 0.09; p = 0.018), between groups 2 (1.41 +/- 0.47) and 3 (p = 0.01), and between groups 2 and 4 (0.29 +/- 0.09; p = 0.045). There were no significant differences in postoperative S100 levels (B) between groups 1 and 2 (p = 0.05), groups 1 and 4 (p = 0.05), or groups 3 and 4 (p = 0.93). CONCLUSION: Uncontrolled hyperoxic reoxygenation on CPB for surgical correction of congenital heart defects is associated with higher S100 levels in cyanotic infants as compared to acyanotic patients undergoing comparable operations. PMID- 11100758 TI - Complete arterial revascularization using T-graft technique in diabetics with coronary three-vessel disease. AB - BACKGROUND: Coronary revascularization using exclusively arterial grafts holds the promise of improved long-term patency. The T-graft approach achieves this goal with only two arterial grafts in coronary 3-vessel disease. Arterial grafts in diabetics, however, exhibit more frequently atherosclerotic wall abnormalities, and higher levels of endothelin-1 were found in diabetic arterial grafts, which may be associated with a higher incidence of vasoconstriction. The objective of this prospective study was to compare functional und angiographic parameters of arterial T-grafts in diabetics and nondiabetics. METHODS: Coronary angiography was performed consecutively in 20 patients with insulin-dependent diabetes mellitus (IDDM), 20 patients with non-insulin-dependent diabetes mellitus (NIDDM), and 100 non-diabetics one week after complete arterial revascularization with T-grafts. Graft patency was assessed, and the diameter of the proximal left internal mammary artery (IMA) graft was measured using quantitative coronary analysis. Absolute flow volume in the proximal left IMA was measured using the flow-wire technique at baseline and after an adenosine injection into the graft to induce maximal hyperemia. Coronary flow reserve (CFR) was calculated as the ratio of maximal to baseline flow. RESULTS: There was no difference between patients with IDDM, patients with NIDDM and non-diabetics with respect to patency (98.3% vs. 98.8% vs. 97,8%, n.s.), graft lumen diameter (3.42 +/- 0.48 vs. 3.36 +/- 0.50 vs. 3.38 +/- 0.41 mm, n.s.), baseline flow (78.4 +/- 34.3 vs. 83.1 +/- 36.6 vs. 81.5 +/- 39.0 ml/min, n.s.), and CFR (1.85 +/- 0.37 vs. 1.89 +/- 0.44 vs. 1.90 +/- 0.40, n.s.). CONCLUSION: Baseline parameters (graft diameter and quantitative graft flow), patency and CFR are identical in diabetics and non-diabetics. Our results suggest that diabetic patients with coronary 3-vessel disease take comparable profit from complete arterial revascularization using the T-graft technique as non-diabetics. PMID- 11100759 TI - Controlled limb reperfusion with a simplified perfusion system. AB - The goal of revascularisation of an acutely ischemic limb is to prevent the loss of the extremity and to enable return of normal function. Although reperfusion of an ischemic limb is a prerequisite for the preservation or reestablishment of function, it may, in itself, cause further injury. Controlled reperfusion after revascularisation may reduce this injury and may facilitate return of normal function. Thus far, the technique of controlled limb reperfusion required the use of cardiosurgical equipment and has therefore been reserved for cardiac surgery centers. However, the majority of patients with acute limb ischemia are referred to hospitals, where the technical equipment for controlled reperfusion is not available. We modified the technique of controlled limb reperfusion using a simplified perfusion system with a pressure-cuffed bag, which allows controlled reperfusion without the use of a roller pump. The modified reperfusion technique was applied to 9 patients with acute and persistent ischemia of the limbs (mean ischemic period: 21 +/- 21 hours). Controlled reperfusion consisted of a 30 min infusion of a normothermic reperfusate solution, which was mixed with the patient's blood (6:1, blood/reperfusate ratio) distally to the proximal obstruction. RESULTS: Five patients (56%) recovered with normal function of the limb. Two patients (22%) lost the ischemic limb (ischemic periods: 77 h and 9h; creatine kinase before operation: 6230 U/I and 1045 U/I): another two patients (22%) died, who were in profound cardiogenic shock. CONCLUSION: The simplified perfusion system allows to put controlled limb reperfusion into practice in any operating room. The results support the notion that controlled limb reperfusion should be applied in any patient with acute ischemia of an extremity. PMID- 11100760 TI - A 20-year follow-up of internal carotid artery endarterectomy with bifurcation advancement. AB - BACKGROUND: Carotid artery disease is a frequent cause of transient ischemic attack and of cerebral infarction. For two last decades, we have been performing endarterectomy of the internal carotid artery with bifurcation advancement. METHODS: From January 1977 until December 1997, all records of patients who underwent internal carotid artery endarterectomy with bifurcation advancement were reviewed. Data were collected from patients charts and by a questionnaire. 160 patients (80.6% men, 19.4% women, average lifetime 65.1 year) underwent a total of 181 endarterectomies with bifurcation advancement. RESULTS: The 30-day mortality was 1.9% and the postoperative stroke plus death rate 3.1%. The incidence of reoperations was 0.6% with an average follow up of 64 months. In one patient (0.6%), a significant restenosis of the repaired carotid artery was observed. The 1, 5 and 10 years neurological death free survival (including early mortality) was 99.3%, 97.2% and 92.5% and the overall survival (including early mortality) was 96.3%, 78.9% and 59.3% (Kaplan-Meier). CONCLUSIONS: The technique of the internal carotid artery endarterectomy by bifurcation advancement is a safe and reliable method for improvement of cerebral blood supply. Or foreign material or autologous vein can thus be avoided. This method offers excellent long term patency and has a notable lack of late restenosis. PMID- 11100761 TI - Coronary vascular stunning in bypass operations during reperfusion and treatment with nitroglycerin. AB - OBJECTIVE: A compromised blood flow after ischemia and reperfusion caused by an increased coronary artery resistance can additionally jeopardize the recovery of myocytes. During routine bypass operations, we investigated the effect of various nitroglycerin doses on elevated coronary resistance before and after ischemia and after a defined reperfusion period. METHODS: 46 patients with a low-risk profile scheduled for routine coronary artery bypass grafting were investigated. During normothermic total extracorporeal circulation, the completely relieved and fibrillating heart was completely isolated from the systemic circulation and the coronary artery system was perfused at 300 ml/min and flow-controlled. The perfusion pressures were monitored continuously. This protocol was performed at three time points: I. Control (ctr) = 10 minutes after institution of extracorporeal circulation, II. Early reperfusion (early rep) = immediately after an myocardial ischemia of 46 +/- 8 minutes, and III. Late reperfusion (late rep) = after a reperfusion period of 25 +/- 4.5 minutes. In 12 randomly chosen patients in a second step, 3 microg per kg heart weight per min of nitroglycerin (low-dose NTG) was added to the perfusate at time points I and III. In another 12 patients, we applied a bolus injection of 2 mg into the aortic root instead of low-dose NTG. RESULTS: Compared to ctr, vascular resistance had decreased at early rep by 17% (0 - 48%) (p < 0.005). At late rep, resistance had increased by 46% (5 - 94%) (p < 0.001) compared to early rep and by 23% (3 - 36%) (p < 0.005) compared to ctr. Resistances had risen in particular in patients with hypertension. Application of low-dose NTG lowered resistances by 5% (0-8%) (non significant) at ctr, and by 6% (0 - 11%) (non-significant) at late rep. Bolus NTG decreased resistances at ctr by 11% (2 - 21%) (p < 0.05) and at late rep by 21% (6 - 48%) (p < 0.01). CONCLUSIONS: In routine heart surgery, coronary vascular constriction is regularly present during postischemic reperfusion despite myocardial protection measures. NTG abolishes this coronary vascular stunning only in part if systemically applicable dosages are given. High-dose intracoronary application of NTG relieves the coronary vasoconstriction completely, but the dosages needed cannot be applied systemically. In this study, vasoconstriction after reperfusion was markedly increased in patients with hypertension. PMID- 11100762 TI - Surgical outcome of mycobacterium other than mycobacterium tuberculosis pulmonary disease. AB - Between January 1995 and May 1999, MOTT were cultured from sputum, bronchoalveolar lavage or resected lung specimens in 110 cases. 17 patients with MOTT pulmonary disease underwent pulmonary resection. Preoperatively, 5 of 17 patients had been diagnosed with MOTT pulmonary disease. The diagnosis of others was based on positive cultures from surgically resected material, and organism identification was successfully performed by the microplate DNA-DNA hybridization procedure. Surgical resections performed included wedge resection in 7, lobectomy in 6, and segmentectomy in 4. Antibiotics were generally continued for 6 to 24 months postoperatively. However, postoperative antibiotics therapy was not performed for patients who were postoperatively diagnosed with foci localized at the peripheral lung. Resected specimens yielded positive cultures for MOTT in all patients. There were no patients infected with M. konsasii. Regarding postoperative complications, 1 late bronchopleural fistula developed after right upper and middle lobectomy, and was treated with omentopexy. Persistent air leaks (> 7 days) occurred in 5 patients, none of which occurred where linear stapling devices fitted with expanded polytetrafluoroethylene (ePTFE) sleeves were used. One patient diagnosed with M. szulgai postoperatively experienced reactivation 2 years after middle lobectomy despite postoperative antibiotic therapy for 6 months. Other patients have remained free of disease postoperatively. Surgical resection achieve good results for MOTT pulmonary disease, and wedge resection or segmentectomy without postoperative antibiotic therapy was enough for patients whose foci localized at the peripheral lung and whose sputum or BAL cultures revealed no MOTT. Surgical treatment should be performed as early as possible before the pulmonary disease necessitates an extensive operation, and ePTFE sleeves were effective in preventing a postoperative prolonged air leak. PMID- 11100763 TI - E-cadherin expression in surgically-resected non-small cell lung cancers--a clinicopathological study. AB - BACKGROUND: E-Cadherin is a subclass of the cadherin family that plays a major role in the maintenance of intercellular junctions in normal epithelium. Decreased expression of E-cadherin might be closely related to invasiveness and dedifferentiation in human cancers. This study is aimed at investigating the clinicopathological significance of E-cadherin expression and its impact on the prognosis in surgically resected non-small-cell lung cancer patients. METHODS: Using immunohistochemical staining, the expression of E-cadherin was studied in 207 surgically resected lung cancer specimens from January 1990 through December 1994. The clinicopathological data and survival status were recorded and analysed against the E-cadherin expression level in each tumor. RESULTS: E-cadherin expression was detected in 122 of the 207 lung tumors (59.0%), and the expression was significantly lower in tumors with poor differentiation (p < 0.001), in tumors with vascular invasion (p < 0.05), and in tumors with direct invasion into surrounding structures (p < 0.01). There was no correlation between E-cadherin expression, and tumor stage and regional lymph-node metastasis. There was no significant difference in survival rate between higher (> 40%) and lower (< 40%) E-cadherin expression groups; however, in tumors 3 cm or less, a significant difference was found between higher and lower E-cadherin expressions (p < 0.0001). CONCLUSIONS: Underexpression of E-cadherin is associated with poor differentiation and invasiveness in NSCLC. In patients with small NSCLC (< or = 3 cm), higher E-cadherin expression (> 40%) significantly had a favorable prognosis. PMID- 11100764 TI - Doppler-sonographic evidence of cerebral microembolism originating from a biventricular assist device. AB - The use of extracorporeal assist devices in heart failure is associated with the risk of thromboembolic complications [1]. Prove of thromboembolic material in the ventricles and tubes of the assist devices is difficult, and the clinical relevance of thrombotic material in the tubes is not clear. Here, we report on a patient with severe heart failure caused by endstage dilated cardiomyopathy who was bridged to transplantation using a biventricular assist device (BVAD). Five weeks after implantation, transcranial Doppler sonography (TCD) revealed high intensity transient signals (HITS) in basal cerebral arteries, suggesting continued cerebral microembolism. Apart from a correlation of these Doppler sonographic findings with neurological symptoms, macroscopic evidence of fibrin thrombi in the artificial ventricle, and post mortem confirmation of cerebral infarction could be proved. PMID- 11100765 TI - Surgical treatment of a large, highly mobile atheroma in the ascending aorta. AB - A 53-year-old woman diagnosed with chronic renal failure requiring homodialysis, and Behcet's disease was found, during a regular examination, having a large protruding mobile atheroma in the ascending aorta by transthoracic echocardiography. We performed surgery to remove the atheroma and localized debridement using moderate hypothermic circulatory arrest to prevent future stroke and embolism. She had an uneventful postoperative course. PMID- 11100766 TI - Late esophagopleural fistula after left pneumonectomy: report on one case. AB - Esophagopleural fistula is an uncommon complication of pneumonectomy. Late nonmalignant esophagopleural fistula after left pneumonectomy for lung cancer is exceedingly uncommon. We report on one patient who developed such a fistula 33 months after the operation. Signs and symptoms were first attributed to infection of the thoracotomy incision and diagnosis was made only after detection of some food coming from the pleural space. Thoracostomy, enteral feeding by a percutaneously placed gastrostomy tube and myoplasty allowed both closure of the fistula and obliteration of the pleural space. PMID- 11100767 TI - Primary left-atrial leiomyosarcoma. AB - A patient rapidly developing right heart failure due to a left-atrial leiomyosarcoma was admitted for surgery. A large tumor, originating in the left atrium and extending into both pulmonary veins, was removed. Histology showed an unusual epitheloid appearance of many cells with occasional mitoses and a strong immunexpression to desmin and actin. Six months later heart failure developed again: computer tomography demonstrated regrowth of the tumor in the left atrium with invasion of the mediastinum and the para-aortal lymph nodes. The patient died shortly after. PMID- 11100768 TI - Re-opened foramen ovale--a rare cause of postoperative dyspnea following pneumonectomy. AB - Dyspnea and hypoxemia are common postoperative problems following pneumonectomy. Platypnea, the increased dyspnea in the erect position relieved by assuming a prone position, has been reported as a result of right to left inter-atrial shunt. We report here on our experience with a patient who had severe platypnea with remarkable positional arterial desaturation following right pneumonectomy. After establishing the diagnosis with contrast-enhanced transesophageal echocardiography of the preoperatively undetected interatrial right-left shunt, cardiac surgery led to clinical improvement and resumption of platypnea. Given the rarity of the diagnosis, we think interatrial shunt, based on an open foramen ovale, should be taken into consideration when platypnea occurs in patients as a postoperative complication following lung surgery. Transesophageal echocardiography may be helpful in detecting patients with "anatomical closed but functional open" foramen ovale or genuine inter-atrial septal defect prior to lung surgery. PMID- 11100769 TI - Diagnosis, treatment and outcome of patients with Askin-tumors. AB - Askin tumors are highly malignant small-round-cell tumors of the thoracopulmonary region, which occur rarely. Therefore, we report on our experiences with eight patients (5 male, 3 females), who were treated in our department between 11'94 and 10'97 (age: 9-40 years, mean age: 20.5 years). All Askin tumors were diagnosed by histological and immunohistochemical examinations as well as molecular genetic proof of characteristic translocations. In all patients, the tumor arose from the chest wall, infiltrating adjacent ribs and parts of the lung. At the time of first diagnosis, five patients did not reveal any metastases. One patient suffered from intrapulmonary metastases and two patients from an infiltration of the diaphragm and of adjacent vertebral bodies. Treatment consisted of a pre- and postoperative (radio-) chemotherapy according to the EVAIA protocol and a radical tumor resection in all patients. The postoperative course was uneventful in seven patients, one patient suffered from pneumonia after multiple wedge resections for intrapulmonary metastases. Four patients, in whom primary tumor resection was complete, are alive 14, 20, 35 and 84 months after first diagnosis - only one patient had to undergo a second operation for a local relapse 17 months after first diagnosis. The other 4 patients, who suffered from a very extensive primary tumor, expired 13, 17, 18 and 39 months after the diagnosis was made. Our data demonstrate that Askin tumors require an aggressive multimodality treatment consisting of pre- and postoperative chemotherapy, radical surgical resection and postoperative irradiation, which may be performed preoperatively in selected cases, too. PMID- 11100770 TI - Valve preserving treatment of Ebstein's anomaly: perioperative and follow-up results. PMID- 11100771 TI - Characterization of organelles in the vacuolar-sorting pathway by visualization with GFP in tobacco BY-2 cells. AB - We have shown the localization and mobilization of modified green fluorescent proteins (GFPs) with various signals in different compartments in a vacuolar sorting system of tobacco BY-2 cells. In contrast to the efficient secretion of GFP from the transformed cells expressing SP-GFP composed of a signal peptide and GFP, accumulation of GFP in the vacuoles was observed in the cells expressing SP GFP fused with the C-terminal peptide of pumpkin 2S albumin. This indicated that this peptide is sufficient for vacuolar targeting. Interestingly, the fluorescence in the vacuoles disappeared sharply at 7 d after inoculation of the cells, but it appeared again after re-inoculation into a new culture medium. When SP-GFP was fused with the region, termed PV72C, including a transmembrane domain and a cytosolic tail of a vacuolar-sorting receptor PV72, GFP-PV72C was detected in the Golgi-complex-like small particles. Prolonged culture showed that GFP PV72C that reached the prevacuolar compartments was cleaved off the PV72C region to produce GFP, that arrived at the vacuoles to be diffused. These findings suggested that the vacuolar-sorting receptor might be recycled between the Golgi complex and prevacuolar compartments. PMID- 11100772 TI - Circadian waves of expression of the APRR1/TOC1 family of pseudo-response regulators in Arabidopsis thaliana: insight into the plant circadian clock. AB - The Arabidopsis pseudo-response regulator, APRR1, has a unique structural design containing a pseudo-receiver domain and a C-terminal CONSTANS motif. This protein was originally characterized as a presumed component of the His-to-Asp phosphorelay systems in Arabidopsis thaliana. Recently, it was reported that APRR1 is identical to the TOC1 gene product, a mutational lesion of which affects the periods of many circadian rhythms in Arabidopsis plants. TOC1 is believed to be a component of the presumed circadian clock (or central oscillator). Based on these facts, in this study four more genes, each encoding a member of the APRR1/TOC1 family of pseudo-response regulators were identified and characterized with special reference to circadian rhythms. It was found that all these members of the APRR1/TOC1 family (APRR1, APRR3, APRR5, APRR7, and APRR9) are subjected to a circadian rhythm at the level of transcription. Furthermore, in a given 24 h period, the APRR-mRNAs started accumulating sequentially after dawn with 2-3 h intervals in the order of APRR9-->APRR7-->APRR5-->APRR3-->APRR1. These sequential events of transcription, termed 'circadian waves of APRR1/TOCI', were not significantly affected by the photoperiod conditions, if any (e.g. both long and short days), and the expression of APRR9 was first boosted always after dawn. Among these APRRs, in fact, only the expression of APRR9 was rapidly and transiently induced also by white light, whereas such light responses of others were very dull, if any. These results collectively support the view that these members of the APRR1/TOC1 family are together all involved in an as yet unknown mechanism underlying the Arabidopsis circadian clock. Here we propose that the circadian waves of the APRR1/TOC1 family members are most likely a molecular basis of such a biological clock in higher plants. PMID- 11100773 TI - The current state and problems of circadian clock studies in cyanobacteria. AB - Circadian rhythms have been observed in innumerable physiological processes in most of organisms. Recent molecular and genetic studies on circadian clocks in many organisms have identified and characterized several molecular regulatory factors that contribute to generation of such rhythms. The cyanobacterium is the simplest organism known to harbor circadian clocks, and it has become one of most successful model organisms for circadian biology. In this review, we will briefly summarize physiological observations and consideration of circadian rhythms in cyanobacteria, molecular genetics of the clock using Synechococcus, and current knowledge of the input and output pathways that support the cellular circadian system. Finally, we will document some current problems in the studies on the cyanobacterial circadian clock. PMID- 11100774 TI - Deglucosidation of quercetin glucosides to the aglycone and formation of antifungal agents by peroxidase-dependent oxidation of quercetin on browning of onion scales. AB - Outer scales of yellow onion bulbs turn brown during maturing. The brown outer scales contain an antifungal component, 3,4-dihydroxybenzoic acid. An aim of the present study is to elucidate the mechanism of formation of the benzoic acid. In a browning scale, the scale was divided into three areas; fleshy, drying and dried brown areas. Levels of quercetin glucosides in dried brown areas were less than 10% of the glucosides in fleshy and drying areas, whereas levels of quercetin were high in dried brown areas. This result suggests that quercetin was formed by deglucosidation of quercetin glucosides on the border between drying and dried brown areas. Peroxidase (POX) activity of dried brown areas was about 10% of those of fleshy and drying areas. Quercetin was oxidized by autooxidation, and cell-free extracts of drying areas and POX isolated from onion scales enhanced the oxidation even in the absence of externally added hydrogen peroxide. The enhancement of quercetin oxidation was suppressed by catalase. No tyrosinase like activity was detected in the cell-free extracts and the POX preparation. These results suggest that, during the enhanced oxidation of quercetin, hydrogen peroxide is formed. 3,4-Dihydroxybenzoic acid and 2,4,6-trihydroxyphenylglyoxylic acid, which were the oxidation products of quercetin, were found in dried brown area. These results suggest that an antifungal agent 3,4-dihydroxybenzoic acid is formed by POX-dependent oxidation of quercetin on browning of onion scales. PMID- 11100775 TI - Overexpression of mitochondrial citrate synthase in Arabidopsis thaliana improved growth on a phosphorus-limited soil. AB - The gene for mitochondrial citrate synthase (CS) was isolated from Daucus carota (DcCS) and introduced into Arabidopsis thaliana (strain WS) using Agrobacterium tumefaciens-mediated transformation. Characteristics of citrate excretion were compared between T3 transgenic plants, which were derived from the initial transgenic plants by self-fertilization and homozygous for DcCS, and the control plants that had no DcCS. The highest CS activity 0.78 micromol protein min(-1) exhibited by the transgenic plants was about threefold greater than that found in the control plants (0.23-0.28 micromol protein min(-1)). Western analysis of the transgenic plants showed two CS signals corresponding to signals obtained from both D. carota and A. thaliana. Thus, it appears that the CS polypeptides by ectopic expression of DcCS were processed into the mature form and localized in the mitochondria of A. thaliana. The signal corresponding to the mature form of DcCS were greater in the transgenic plants having higher levels of CS activity. When the transgenic plants were grown in Al-phosphate media, a correlation between the levels of CS activity and the amounts of citrate excreted into the medium. The highest value (5.1 nmol per plant) was about 2.5-fold greater than that from control plants (1.9 nmol per plant). Both growth and P accumulation were greater in transgenic plants with high CS activity than that in control plants when they were grown on an acid soil where the availability of phosphate was low due to the formation of Al-phosphate. It appears that the overexpression of CS in A. thaliana improves the growth in phosphorous limited soil as a result of enhanced citrate excretion from the roots. PMID- 11100776 TI - Salicylic acid induces the expression of a number of receptor-like kinase genes in Arabidopsis thaliana. AB - Receptor-like protein kinases (RLKs) are encoded by a divergent multigene family and their functions have been implicated in a wide range of signal transduction pathways. In this study, we examined the effect of salicylic acid (SA) on the expression of RLK genes in Arabidopsis thaliana. RNA gel blot analysis revealed that transcripts of RKC1 and a number of its homologs, whose translation products contain C-X8-C-X2-C motifs in the putative extracellular domain, accumulated to a higher level in response to SA treatment of plants. The chimeric fusion between the RKC1 5'-upstream region and the beta-glucuronidase (GUS) reporter gene reproduced the SA responsiveness in transgenic plants. In addition, some of RLK genes of the leucine-rich repeat (LRR) class and those of the S-domain class were also induced by SA. We found that the upstream regions of these SA-responsive RLK genes contain the TTGAC sequence, which has been suggested to be important for induced expression of many plant defense genes. These results suggest the involvement of a number of RLKs in SA-mediated defense responses. PMID- 11100777 TI - Photosynthetic electron flow regulates transcription of the psaB gene in pea (Pisum sativum L.) chloroplasts through the redox state of the plastoquinone pool. AB - Plants respond to changing light conditions by altering the stoichiometry between components of the photosynthetic electron transport chain of chloroplast thylakoids. We measured specific run-on transcription of the chloroplast genes psaB, psbA and rbcL in pea (Pisum sativum L.) seedlings grown under three different conditions of illumination: light selective for photosystem I (PSI light); light selective for photosystem II (PSII-light); and a combination of PSI and PSII-light (mixed light, ML). The transcriptional rate of the psaB gene increased under PSII-light and decreased under PSI-light, while the transcriptional rates of the psbA and rbcL genes were affected only in a non specific way. Similar effects also occurred in plants grown under ML and switched to either PSI- or PSII-light for 4 h. Addition of the inhibitors of photosynthetic electron transport 3-(3,4 dichlorophenyl)-1,1-dimethylurea (DCMU) and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) influenced psaB transcription in isolated, illuminated chloroplasts: DCMU addition resulted in oxidation of the plastoquinone pool and decreased transcription of psaB; DBMIB addition resulted in reduction of the plastoquinone pool and increased transcription of psaB. The experimental results obtained in vivo and in vitro provide evidence for coupling between the redox state of plastoquinone and the rate of transcription of the psaB gene in pea. PMID- 11100778 TI - hosoba toge toge, a syndrome caused by a large chromosomal deletion associated with a T-DNA insertion in Arabidopsis. AB - We isolated a T-DNA-tagged mutant named hosoba toge toge (hot) in which a pleiotropic phenotype was observed in both the shoot and root throughout the life cycle. The phenotype and allelism indicated that the mutant has a defect in both the FASCIATA1 (FAS1) gene and the FT gene located on the bottom arm of chromosome 1. Analysis of the junctions between the T-DNA ends and the plant genome suggested the presence of a 75.8-kbp deletion at the insertion site. In addition to FAS1 and FT, 13 genes were predicted to exist in the region corresponding to that deleted in hot. They include homologs of genes for type II inositol-1,4,5 triphosphate 5-phosphatase (IP5Pase), the beta-chain of N-acetyl-beta glucosaminidase (NAGase), NADPH oxidoreductase of the zeta-crystallin family, polygalacturonase, and endo-1,4-beta-glucanase. Although most aspects of the hot phenotype can be explained by loss of FAS1 and FT functions, some novel phenotypic features which may represent aspects of a mutant phenotype due to loss of-function of other gene(s) were observed. One "wild-type" ecotype and a previously reported T-DNA insertion line, neither of which has any obvious phenotypic abnormality, carry a possible loss-of-function mutation in the zeta crystallin homolog and in the NAGase beta chain homolog, respectively. PMID- 11100779 TI - Quantification of Ca2+/H+ antiporter VCAX1p in vacuolar membranes and its absence in roots of mung bean. AB - Vacuolar Ca2+/H+ antiporter VCAX1p, which contributes to the Ca2+ accumulation into vacuoles, was quantified by immunochemistry. The antiporter content in vacuolar membranes was 0.14 and 1.1 microg mg(-1) of membrane protein for hypocotyls and epicotyls, respectively. The calculated turnover number was 120 s( 1). Roots lacked the antiporter protein and the transcript. PMID- 11100780 TI - Ethylene suppresses jasmonate-induced gene expression in nicotine biosynthesis. AB - In Nicotiana sylvestris, a set of nicotine biosynthesis genes were activated by exogenous application of methyl jasmonate, but the activation was effectively suppressed by simultaneous treatment with ethylene. When N. sylvestris transgenic hairy roots were treated with a natural ethylene precursor, the jasmonate responsive expression of the promoter from a nicotine pathway enzyme gene was completely suppressed, and this suppressive effect was abolished when ethylene perception was blocked with silver cation. These and additional immunoblot results suggest that ethylene signal antagonizes jasmonate signal in nicotine biosynthesis. PMID- 11100781 TI - Ultraviolet-B radiation causes tendril coiling in Pisum sativum. AB - Low dose UV-B radiation (UV-B(BE,300) = 0.1 W m(-2)), but neither UV-A radiation, ozone and NaCl stress, nor wounding, caused tendril coiling in Pisum sativum. This coiling occurred with both attached and detached tendrils and can be used as a specific UV-B stress marker in pea. PMID- 11100782 TI - cDNA cloning and expression of an aminoalcoholphosphotransferase isoform in Chinese cabbage. AB - Aminoalcoholphosphotransferase (AAPT) catalyzes the synthesis of phosphatidylcholine and phosphatidylethanolamine from diacylglycerol plus a CDP aminoalcohol such as CDP-choline or CDP-ethanolamine. Previously we reported the cloning of a cDNA encoding this enzyme from Chinese cabbage roots, and suggested the presence of possible isoforms [Min et al. (1997) J. Plant Biol. 40: 234]. We now report the cDNA cloning and expression analysis of a second AAPT from Chinese cabbage. This AAPT cDNA, AAPT2, contains an open reading frame of 1,170 bp coding for a protein of 389 amino acids. It shares 95% identity and 96% similarity with Chinese cabbage AAPT1 at the deduced amino acid level. The results from reverse transcriptase-PCR indicate that expression of AAPT2 is regulated temporally and up-regulated by low temperature. PMID- 11100783 TI - Observations on the preferential biodegradation of selected components of polyaromatic hydrocarbon mixtures. AB - The capacity of the naphthalene degrading enzyme (NAH) system of Pseudomonas fluorescens 5R and a number of other NAH system bacterial isolates to degrade mixtures of polyaromatic hydrocarbons (PAHs) and heterocyclic compounds were examined. It was found that all the examined organisms displayed similar patterns of preferential compound degradation when presented with the same mixture. Using strains that possess portions of the NAH system, this preferential degradation was localized to the activity of naphthalene dioxygenase. Comparisons of the first-order rates of compound degradation with the structures of the mixture components indicated that increased deviation from the base structure of naphthalene led to slower disappearance. Structural features that were found to decrease the rate of compound degradation include an increase in the number of methyl substituents and an increase in the size of a substituent. PMID- 11100784 TI - Photocatalytic degradation of gaseous perchloroethylene: products and pathway. AB - Batch photocatalytic degradation of 1000-ppm gaseous perchloroethylene (PCE) was conducted with UV irradiation such that nearly 100% was decomposed within 10 min. The main intermediate and final product were identified as trichloroacetylchloride (TCAC) and hydrogen chloride (HCl), respectively, and minor ones as dichloroacetic acid (DCAC), monochloroacetic acid (MCAC), carbon tetrachloride, chloroform, and phosgene. More than 90% of Cl- equivalent, i.e., the sum of the chlorine number in PCE, intermediates, and HCl, was compensated for during the time of PCE degradation; a result indicating that no other major chlorinated intermediates are present during the time of PCE degradation. In a similar experiment, 500 ppm of gaseous TCAC degraded into HCl within 3 h without producing DCAC or MCAC, where like PCE, more than 90% of Cl- equivalent, i.e., the sum of the chlorine number in TCAC and HCl, was compensated for during time of TCAC degradation. Accordingly, gaseous PCE is concluded to predominantly follow a degradation pathway of PCE --> TCAC --> HCl. PMID- 11100785 TI - Comparison of the degradation of p-hydroxybenzoic acid in aqueous solution by several oxidation processes. AB - A comparative study is made of 12 methods of chemical oxidation applied to degrading p-hydroxybenzoic acid in aqueous solution. The oxidation processes tested were: UV, O3, UV/TiO2, O3/Fe2+, O3/H2O2, O3/UV, UV/H2O2, H2O2/Fe2+, H2O2/Fe2+/O3, UV/H2O2/O3, H2O2/Fe2+/UV and O3/UV/H2O2/Fe2+. The 12 processes were ranked by reactivity. In a kinetic study, the overall kinetic rate constant was split up into three components: direct oxidation by UV irradiation (photolysis), direct oxidation by ozone (ozonation), and oxidation by free radicals (mainly OH*). PMID- 11100786 TI - 3-Chlorophenol elimination upon excitation of dilute iron(III) solution: evidence for the only involvement of Fe(OH)2+. AB - The transformation of 3-chlorophenol (3CP) photoinduced by iron(II) in aqueous solution has been investigated under monochromatic irradiation (lambda(exc) = 365 nm) representative of atmospheric solar emission. Hydroxyl radicals are formed via an intramolecular photoredox process in iron(III) excited hydroxy-complexes. Fe(OH)2+ is the most active complex in terms of HO* formation and according to our experiments and calculations, it appears that Fe(OH)2+ is the only iron(III) species involved in 3CP oxidation process. Hydroxyl radicals react very rapidly with 3CP, which is eliminated from the solution. The primary intermediates do not accumulate in the medium but rapidly degraded to non-absorbing compounds by a subsequent action of hydroxyl radicals. PMID- 11100787 TI - Rapid reductive destruction of hazardous organic compounds by nanoscale Fe0. AB - Fe0-mediated reductive destruction of hazardous organic compounds such as chlorinated organic compounds (COCs) and nitroaromatic compounds (NACs) in the aqueous phase is one of the latest innovative technologies. In this paper, rapid reductive degradation of COCs and NACs by synthesized nanoscale Fe0 in anaerobic batch systems was presented. The nanoscale Fe0, characterized by high specific surface area and high reactivity, rapidly transformed trichloroethylene (TCE), chloroform (CF), nitrobenzene (NB), nitrotoluene (NT), dinitrobenzene (DNB) and dinitrotoluene (DNT) under ambient conditions, which results in complete disappearance of the parent compounds from the aqueous phase within a few minutes. GC analysis reported that the main products of the dechlorination of TCE and CF were ethane and methane as well as that most of the nitro groups in NACs were reductively transformed to amine groups. These results suggest that the rapid reductive destruction by nanoscale Fe0 is potentially a viable in situ or aboveground treatment of groundwater contaminated with hazardous organic compounds including COCs and NACs. PMID- 11100788 TI - Flotation of metal-loaded clay anion exchangers. Part I: the case of chromates. AB - Synthetic hydrotalcite-like layered materials are known for their ability to remove anions, like the chromates. These sorbents usually exist in powder form, thereby exhibiting high surface area and rapid kinetics for adsorption, but presenting appreciable problems in the subsequent solid/liquid separation process. Almost complete removals were obtained in this paper, from batchwork dispersed-air flotation in presence of a flocculant. Due to the experienced difficulty of flotation of thermally activated (at 500 degrees C) hydrotalcite metal-loaded particles, the application of various surfactants was studied. Continuous-flow laboratory runs certified also the effectiveness of this combined process of sorptive flotation, a promising innovative treatment technology. PMID- 11100789 TI - Photodegradation of humic acids in the presence of hydrogen peroxide. AB - A batch photoreactor was used to evaluate the UV/H2O2 oxidation process for the removal of humic acids in water. A 450-W UV lamp with high-pressure mercury vapor was employed as the light source. The residues of humic acids and hydrogen peroxide were measured for assessment of process performance and understanding of process reaction behavior. The UV photolysis alone can play an important role in the degradation of humic acids. The presence of hydrogen peroxide was found to promote the degradation efficiency. However, excessive dosage of H2O2 does not further improve the degradation of humic acids. On the contrary, the lower the H2O2 dosage the higher the amount of humic acids which can be removed. Aeration with air does not favor the removal efficiency of humic acids as the oxidation lasts for a sufficiently long time. The presence of carbonate species deteriorates the humic acids' removal, whereas it results in a larger amount of H2O2 decomposition. PMID- 11100790 TI - Pyrite-assisted degradation of trichloroethene (TCE). AB - Chemical kinetics of trichloroethene (TCE) degradation by pyrite was investigated at different experimental conditions. The TCE was transformed into C2H2, 1,1 dichloroethene (DCE) and C2H4 by a slow process (240 h required to convert TCE by 80%). Thereafter, the degradation rate showed a monotonous increase with the pH. A simple kinetic model was proposed to quantify the TCE degradation by pyrite. PMID- 11100791 TI - The spinning disc reactor--studies on a novel TiO2 photocatalytic reactor. AB - A new type of photocatalytic reactor, the spinning disc reactor (SDR), was used to degrade aqueous solutions of 4-chlorophenol and salicylic acid. The efficiency of the photocatalytic process depends on the type of UV source used. Lamps supplying shorter wavelength UV radiation are more efficient than those whose emissions lay mainly in the near UV region. The method used to coat the disc of the SDR does not meet its operational requirements. The characteristics of the turbulent liquid films produced in the SDR reduce the influence of mass transfer over the overall photocatalytic process. PMID- 11100793 TI - Bisphenol A in hazardous waste landfill leachates. AB - The levels of bisphenol A in hazardous waste landfill leachates collected in Japan in 1996 were determined by gas chromatograph/mass spectrometer (GC/MS). Bisphenol A was found in seven of 10 sites investigated. All the hazardous waste landfills with leachates contaminated by bisphenol A were controlled. The concentrations of bisphenol A ranged from 1.3 to 17,200 microg/l with a median concentration of 269 microg/l. The source of bisphenol A in landfill leachates may be the waste plastics in waste landfill. The concentrations of bisphenol A in some samples exceeded the EC50 or LC50 levels for aquatic biota. Landfill leachates may be a significant source of bisphenol A found in the environment. PMID- 11100792 TI - Kinetics of oxidation of chlorobenzenes and phenyl-ureas by Fe(II)/H2O2 and Fe(III)/H2O2. Evidence of reduction and oxidation reactions of intermediates by Fe(II) or Fe(III). AB - The rates of degradation of 1,2,4-trichlorobenzene (TCB), 2,5 dichloronitrobenzene (DCNB), diuron and isoproturon by Fe(II)/H2O2 and Fe(III)/H2O2 have been investigated in dilute aqueous solution ([Organic compound]0 approximately 1 microM, at 25.0 +/- 0.2 degrees C and pH < or = 3). Using the relative rate method with atrazine as the reference compound, and the Fe(II)/H2O2 (with an excess of Fe(II)) and Fe(III)/H2O2 systems as sources of OH radicals, the rate constants for the reaction of OH* with TCB and DCNB were determined as (6.0 +/- 0.3)10(9) and (1.1 +/- 0.2)10(9) M(-1) s(-1). Relative rates of degradation of diuron and isoproturon by Fe(II)/H2O2 were about two times smaller in the absence of dissolved oxygen than in the presence of oxygen. These data indicate that radical intermediates are reduced back to the parent compound by Fe(II) in the absence of oxygen. Oxidation experiments with Fe(III)/H2O2 showed that the rate of decomposition of atrazine markedly increased in the presence of TCB and this increase has been attributed to a regeneration of Fe(II) by oxidation reactions of intermediates (radical species and dihydroxybenzenes) by Fe(III). PMID- 11100794 TI - Reductive degradation of carbaryl in water by zero-valent iron. AB - Reduction of carbaryl solution by zero-valent iron powder (ZVIP) was studied in a rotator batch system (70 rpm) in order to evaluate the utility of this reaction in remediation of carbamate contaminated water. Degradation with different amount of ZVIP: 0.01, 0.02, 0.03, 0.04 g/ml at pH 6.6 and at ambient temperature was investigated. The results show that the process exhibits a degradation rate appearing to be directly proportional to the surface contact area of ZVIP (325 mesh) with the carbaryl molecules. Three analytical techniques were used to monitor carbaryl degradation: (1) A UV-Vis diode array spectrophotometer was used to record all spectra. (2) A high performance liquid chromatography was used to separate by-products and examine the evolution of breakdown products. (3) A home built spectrophosphorimeter that uses the solid surface room temperature phosphorescence (SSRTP) was employed to observe selectively the decline of the carbaryl concentration at different amount of ZVIP on Whatman no. 1 filter paper. Results show that the reducing degradation of carbaryl with ZVIP as the source of electrons is effective with a half-life close to several minutes. PMID- 11100795 TI - Bacterial release of arsenic ions and organoarsenic compounds from soil contaminated by chemical warfare agents. AB - The objective of this paper was to investigate possible participation of microorganisms in the release of soluble arsenical compounds from organoarsenic warfare agents in contaminated soil. A number of bacterial strains were isolated with high resistance against As3+ and As5+ ions which are able to degrade the water insoluble compounds triphenylarsine (TP) and triphenylarsineoxide (TPO). These strains belong to different genera of bacteria. Release of arsenic ions and soluble organoarsenic compounds from soil by the activity of autochthonic soil bacteria and a mixture of the isolated pure cultures was demonstrated by percolation experiments with undisturbed soil samples (core drills) from the contaminated site. This release increased after addition of nutrients (mineral nitrogen and phosphorus, sodium acetate and ethanol) and is nearly independent of the percolation temperature (5 degrees C and 22 degrees C). These results show that bacteria play an important role in the release of arsenical compounds from organoarsenic warfare agent contaminated soil. This release is limited by shortage of water and, above all, of nutrients for the microorganisms in the sandy forest soil. These results are important both for the management and security and possibly for bioremediation of military waste sites containing similar contaminations. Furthermore, this is the first report on bacterial degradation of organoarsenic warfare compounds. PMID- 11100796 TI - Prediction of Fenton oxidation positions in polycyclic aromatic hydrocarbons by Frontier electron density. AB - Five recalcitrant polycyclic aromatic hydrocarbons (PAHs) in ethanol were subjected to Fenton oxidation, and following GC-MS identification of respective oxidation products, their oxidation positions were compared to those predicted by Frontier electron density. Quinone forms of oxidation products were identified in each PAH. With the exception of fluorene, oxidation positions of quinone forms of products of acenaphthylene, anthracene, benz(a)anthracene, and benzo(a)pyrene corresponded with predicted positions in which Frontier electron density was high. From these results, it appears that determining the Frontier electron density of a PAH is a promising method for predicting the Fenton oxidation position. PMID- 11100797 TI - The effect of context on mother's interaction style with Down's syndrome and typically developing children. AB - Several studies suggest that difficulties with production or comprehension of language might be associated with the number of interactions initiated by parent or child, responsiveness or ability to sustain ongoing interactional sequences, or the distribution of parental interaction, control and reinforcement strategies. In this study Down's syndrome and typically developing preschool children were observed interacting with their mothers in free play and mealtime settings. We expected interaction patterns in the mothers of Down's syndrome children to be different from those in the mothers of typically developing children. Sixteen mother-child dyads (eight with Down's syndrome children and eight with typically developing children) served as subjects. Mothers of Down's syndrome children use more teacher and helper behaviors, particularly in meal time context, and less positive verbalizations than the mothers of typically developing children. Down's syndrome children also showed higher frequency of eye gazes during mealtime context. Patterns of such differences are discussed in terms of how mothers' style interactions during home activities might be differentially affected by different types of parent training interventions. PMID- 11100798 TI - Hematological and immunological acute mental stress responses of people who are severely and profoundly mentally retarded. AB - Relocation stress may be one factor increasing the mortality rate of people who are severely and profoundly retarded (S/P MR) when they transfer from institutional to community living arrangements. However, no research exists concerning acute stress effects with groups who are S/P MR. In this project, 28 residents of a state facility for those with S/P MR were exposed to five-minute structured educational tasks. Venous blood samples were drawn before and after the stressor. Granulocytes, red blood cells, hemoglobin, and plasma protein increased while monocytes decreased after stress. Immune cell subsets did not change significantly. Hemoconcentration, an important factor in thrombosis and ischemia, may relate to relocation stress in S/P MR populations. Methodological factors limit generalization but additional research with larger samples, more indices of stress, more poststress blood samples, and additional stressors are encouraged. PMID- 11100799 TI - Say-do-report training to change chronic behaviors in mentally retarded subjects. AB - Two mentally retarded subjects with language deficits participated. A say-do report correspondence training was implemented to break the functions of present conditions given by a long history of contingencies maintaining chronic inadequate patterns of behavior. The say-do-report procedure was implemented following a multiple baseline design across two behaviors in one subject and in an AB design for the other. During baseline, all possible social contingencies maintaining the inadequate behaviors were eliminated. Promising or saying what a subject would do was then implemented and followed by the differential reinforcement of say-do correspondence reports. All behaviors changed and were maintained at an appropriate level, even after eliminating the components involved in the say-do-report procedure, that is, the reports, the extra consequences, and even the promise. Results are discussed in the context of verbal behavior altering the function of present conditions in subjects with limited verbal repertory, as well as in the context of new applications to make a difference in chronic behaviors. PMID- 11100800 TI - Perceptions of three terms to describe physical awkwardness in children. AB - A survey examined the perceptions of 125 Ss who completed a 20-item Semantic Differential ratings for three terms: developmental dyspraxia (DD); developmental coordination disorder (DCD); clumsy child syndrome (CCS). Ss included parents, teachers, physicians, occupational therapists, physiotherapists, psychologists, and speech therapists. Analyses revealed significant main effects for term and role, and interactions between them on certain items. DD is perceived as severe, complex, strong, difficult, serious and technical, whereas CCS as mild, simple, weak, easy, humorous and nontechnical. Both DD and DCD are regarded as more sensitive, positive, graceful and objective than CCS. DD is considered more permanent than DCD and CCS. The meaning of the terms varies as a function of role surrounding children with physical awkwardness. PMID- 11100801 TI - A review of "noncontingent" reinforcement as treatment for the aberrant behavior of individuals with developmental disabilities. AB - The term noncontingent reinforcement (NCR) refers to the delivery of an aberrant behavior's known reinforcer on a response-independent basis. The typical result is a decrease in responding from baseline (i.e., reinforcement) levels. NCR has become one of the most reported function-based treatments for aberrant behavior in the recent literature. The purpose of this review is to briefly discuss the history of the procedure and summarize the findings from the treatment research literature. The review is organized into the following sections: (a) basic research on NCR, (b) NCR as a control procedure, (c) NCR as a function-based treatment, (d) considerations in the programming of NCR schedules, (e) behavior change mechanisms underlying NCR effects, and (t) directions for future research. PMID- 11100802 TI - Functional analysis of aberrant behavior maintained by automatic reinforcement: assessments of specific sensory reinforcers. AB - The purpose of this study was to develop a systematic functional assessment package for aberrant behaviors maintained by nonsocial (automatic) reinforcement. The assessment package included four components: (1) functional analysis, (2) antecedent assessment of specific automatic reinforcement sources, (3) stimulus preference assessment, and (4) treatment evaluation. Functional analysis data indicated automatic reinforcement functions of the stereotypy exhibited by a 10 year-old male and the self-injury (SIB) exhibited by a 30-year-old male. Antecedent assessments of sensory classes indicated that auditory stimulation and tactile stimulation were associated with stereotypy and SIB, respectively. A multiple-stimulus-without-replacement procedure was conducted with each participant to identify the most- and least-preferred stimuli within the identified sensory classes. In an attempt to validate the assessment package for each participant, a DRO procedure was implemented using a reversal design with a multielement component. DRO procedures using stimuli within the targeted sensory classes were successful in eliminating the aberrant behaviors of both participants. The results are discussed in the context of improving the methodology for assessing and treating automatically reinforced behaviors. PMID- 11100803 TI - Effectiveness of video feedback and self-management on inappropriate social behavior of youth with mild mental retardation. AB - The effectiveness of a video feedback and self-management package was assessed with various inappropriate behaviors exhibited by six youth with mild mental retardation. The procedure consisted of (a) videotaping participants' inappropriate behavior, (b) having them self-monitor and record their behavior, (c) prompting them to evaluate their behavior against a criterion, and (d) allowing to reinforce themselves for appropriate behaviors. Data were collected within a nonconcurrent multiple baseline design across participants. Results showed a statistically significant decrease of the percentage intervals of inappropriate behavior when the procedure was in effect. The total number of interactions remained stable across the different phases of the study. Video feedback and self-management contributed to generalization across settings. PMID- 11100804 TI - Mechanisms of induction of tolerance to organ allografts. AB - The long-term acceptance of organ allografts can be induced in numerous rodent and some preclinical outbred models. Induction methods can use donor alloantigen in various forms, including spontaneous acceptance of grafts such as livers, to deviate the immune response so a subsequent graft will be accepted. Establishment of lymphohemopoietic chimerism is not essential. Short-term immunosuppressive treatments that prevent acute rejection can also induce tolerance. These include nonspecific immunosuppressive drugs and immunotherapy that blocks cell surface molecular interactions or cytokine function. There is variation in the effect of these protocols on different strain combinations that may be due to innate differences in the cell subpopulations and cytokines activated in the hosts. Th1 cytokines, although important in the mediation of rejection, are also required for induction of tolerance. Th2 cytokines may facilitate tolerance induction but are not essential. The tolerant state takes weeks to fully mature after exposure to alloantigen. Tolerance is associated with a loss or change in dendritic cells and the development of suppressor cells, which in all cases include CD4+ T cells. In the near future precise understanding of the function of these two cell types may allow diagnosis and induction of tolerance in clinical transplantation. PMID- 11100805 TI - Internalization and intracellular fate of TCR-CD3 complexes. AB - The number of surface TCR-CD3 complexes is maintained by an equilibrium between the synthesis and secretion of new polypeptides, their internalization, recycling, and degradation. The different subunits of the TCR-CD3 complex do not display the same intracellular trafficking dynamics. Thus, in the absence of stimuli, TCR and zeta chains may be degraded at a higher rate than CD3 subunits, which are mostly recycled. T-cell activation by antigen, anti-TCR-CD3 antibodies, or pharmacological activators of protein kinase C, results in increased TCR-CD3 internalization, followed by the downmodulation of TCR-CD3 surface levels. Once internalized, TCR-CD3 complexes may either enter a recycling pathway or be sorted to lysosomes and degraded. Protein serine kinases and protein tyrosine kinases may influence the internalization and intracellular sorting of TCR-CD3 complexes. In line with these results TCR-CD3 ligands stimulate both TCR-CD3 internalization and degradation, whereas protein kinase C activators stimulate internalization only. Depending on the stimulus applied, internalization motifs from one or several TCR-CD3 subunits mediate endocytic routing of the complex. The involvement of signaling molecules in the intracellular fate of TCR-CD3, the nature and location of sequences for internalization and intracellular sorting, and the role of downregulation in T-cell activation are still the main open questions. PMID- 11100806 TI - CD5 signal transduction: positive or negative modulation of antigen receptor signaling. AB - The CD5 lymphocyte surface glycoprotein is a coreceptor involved in the modulation of antigen-specific receptor-mediated activation and differentiation signals. Although first considered a costimulatory molecule in mature peripheral T cells, recent studies of CD5-/- mice have opened the possibility that CD5 may also mediate inhibitory signals that attenuate TCR/CD3- and BCR-mediated triggering in thymocytes and a subgroup of B cells (B-1a cells), respectively. The ultimate molecular basis for these differential modulatory properties of CD5, depending on the context of lymphocyte subset and differentiation stage, are presently unknown and are an issue of current intensive investigation. Here, we review recent reports, both contradictory and complementary, focused on CD5 mediated molecular intracellular signaling events that could provide the basis for its immunomodulatory properties. PMID- 11100807 TI - Non-hematopoietic cutaneous metastases in children and adolescents: thirty years experience at St. Jude Children's Research Hospital. AB - BACKGROUND: The spectrum of cutaneous metastasis of non-hematopoietic neoplasms in the pediatric population is not well documented. We report the histologic diversity of this unusual process over a 30-year period at a tertiary care center for pediatric malignancy (St. Jude Children's Research Hospital, Memphis, TN, USA). METHODS: Of 1,971 pathology accessions which included histologic material on skin (1,604 surgical cases and 367 autopsy cases) we found 40 cases (2% of total skin accessions) coded for metastatic non-hematopoietic malignancy. RESULTS: The patients (n=34) ranged in age from 1 month to 20 years (mean=9.8 years) and had a male:female ratio of 1:1. The histologic diagnoses were as follows: rhabdomyosarcoma NOS (6 cases), embryonal rhabdomyosarcoma (4 cases), alveolar rhabdomyosarcoma (4 cases), neuroblastoma (8 cases), osteosarcoma (2 cases), choriocarcinoma (2 cases), peripheral neuroepithelioma or Ewing's sarcoma (2 cases), malignant rhabdoid tumor (1 case), paraganglioma (1 case), nasopharyngeal carcinoma (1 case), sarcoma NOS (1 case), colon adenocarcinoma (1 case), and malignant melanoma (1 case). CONCLUSIONS: Cutaneous or subcutaneous metastasis of non-hematopoietic malignancies in children and adolescents is a rare occurrence but in a high percentage of cases may be the first manifestation of disease. The tumors most likely to metastasize to the skin in children are rhabdomyosarcoma and neuroblastoma and they are more likely than adult malignancies to disseminate to multiple distant sites. PMID- 11100808 TI - Transepithelial elimination of cutaneous vulval granuloma inguinale. AB - BACKGROUND: Transepithelial elimination (TEE), a distinct and well-known entity, is a process during which the skin eradicates undesirable or irritative dermal substances through intact epidermis or follicular epithelium by passive or active means. Although TEE is being described in an increasing number and range of pathological processes, to date, TEE of granuloma inguinale (GI) remains unrecorded in the English-language literature. The aims of this study were: 1) To appraise the light microscopic and ultrastructural morphological epidermal changes that are associated with TEE of cutaneous vulval GI; and 2) To determine the role of intra-epidermal leucocytes and histiocytes in the pathogenesis of TEE of vulval GI. METHODS: This is a retrospective 9-year histopathological review of all cases diagnosed and coded as vulval granuloma inguinale in the Department of Anatomical Pathology, Nelson R. Mandela School of Medicine, University of Natal, Durban, South Africa. Ultrastructural evaluation was performed on selected cases using a Jeol transmission electron microscope. RESULTS: Of 53 skin biopsies from 47 patients with vulval GI, 43 were suitable for the study. The age range of patients was 15-40 years (mean age=22 years). There were eleven papular, twelve nodular, seven verrucous and thirteen ulcerative lesions. Donovan bodies within macrophages, free-lying Donovan bodies and dense aggregates of neutrophils and plasma cells were seen in the dermis of all biopsies. There was consistent overlying pseudoepitheliomatous hyperplasia. The dermal inflammatory infiltrate hugged the dermo-epidermal junction and appeared entrapped between elongated and acanthotic epidermal rete ridges and pegs. Transepidermal neutrophil microabscesses, histiocytes containing Donovan bodies and neutrophilic and histiocytic fragmentation were present. A variable number of free-lying and intra histiocytic Donovan bodies and neutrophils were present on the surface of the epidermis. On ultrastructural investigation epidermal spongiosis, intracellular oedema, free-lying, intra-neutrophilic and intra-histiocytic Donovan bodies, and intact and degenerating neutrophils and histiocytes were evident between keratinocytes. The degenerative histiocytes demonstrated marked vacuolation, mitochondrial swelling and bacilli within phagolysosomal vacuoles, bound by intact or disrupted limiting membranes. CONCLUSION: The inflammatory infiltrate at the epitheliomesenchymal interface, pseudoepitheliomatous hyperplasia, intra epidermal accumulation and disintegration of neutrophils and histiocytes, and the associated release of lytic enzymes, play important contributory roles in TEE of GI. TEE of infectious agents is a poorly recognised mechanism of spread of infectious diseases and represents a public health hazard. In cutaneous vulval GI, TEE is highlighted as a hitherto unrecognised, potential method of spread of Calymmatobacterium granulomatis. PMID- 11100809 TI - "Tubular Spitz's nevus" an artifact of fixation? AB - BACKGROUND: Spitz's nevi may contain numerous tubule-like structures. This phenomenon has been described as "tubular Spitz's nevus" and has been attributed to either apoptosis in the center of nests of melanocytes or tubular metaplasia of melanocytes. METHODS: In order to study this peculiar phenomenon, we reviewed 31 Spitz's nevi from our teaching collection. RESULTS: Empty spaces vaguely resembling tubules were found in 15 Spitz's nevi, all of which were examples of the epithelioid-cell type of Spitz's nevus. Histopathologically and immunohistochemically, there was no evidence of apoptosis within or around tubule like structures. Empty; spaces in the center of nests ranged from distinctive tubule-like structures to only slight separation of melanocytes from one another. Furthermore, empty spaces were also seen around nests, often in combination with distinctive tubule-like changes. CONCLUSIONS: Our findings suggest that so-called "tubular" structures in epithelioid Spitz's nevi result from retraction of tissue secondary to fixation in formalin. "Tubular Spitz's nevus" seems to represent an artifact rather than a distinctive variant of Spitz's nevus. PMID- 11100810 TI - A causal role for parvovirus B19 infection in adult dermatomyositis and other autoimmune syndromes. AB - BACKGROUND: Infection with parvovirus B19 (B19) has been associated with connective tissue disease (CTD) stigmata, namely, a systemic lupus erythematosus (SLE)-like illness, seronegative polyarthritis resembling rheumatoid arthritis, and vasculitis. The dermatopathology and pathogenetic basis of such B19 associated CTD-like syndromes have not been elucidated. OBJECTIVE: We attempted to document persistence of the B19 genome in skin lesions of 7 patients with CTD like symptomatology following B19 infection and to correlate systemic manifestations to dermatopathological findings. METHOD: In 7 prospectively encountered patients in whom history, clinical signs and/or serology supported a diagnosis of CTD in the setting of B19 infection, dermatopathological and clinical features were correlated. Parvovirus B19 viral genome was sought in skin tissue using the polymerase chain reaction (PCR). RESULTS: Two patients had clinical features diagnostic of myopathic dermatomyositis (DM), 1 of whom is still symptomatic 1.5 years after the onset of her illness, and the other has had typical clinical features of DM for a duration of 3.5 years. A 3rd patient with SLE remains symptomatic 4 years after the onset of her illness. A 4th patient has persistent seronegative symmetrical polyarthritis of 6 years' duration and cutaneous lesions of granuloma annulare (GA). The 5th patient has a 1.5-year history of debilitating polyarthritis and cutaneous lesions with overlap features of DM and subacute cutaneous LE (SCLE). The 6th patient has had a persistent folliculocentric necrotizing vasculitis for 3 years. The 7th patient has a 1-year history of microscopic polyarteritis nodosa (PAN) with cutaneous vasculitis and persistent active renal disease. In 4 patients, exposure to children with fifth disease immediately preceded the onset of their CTD. Parvovirus B19 infection was documented serologically in 6 patients with antibodies of IgG subclass in 6 and of IgM subclass in 1. Four of 6 patients questioned had a history of atopy. Skin biopsies from patients with clinical features of SLE or DM demonstrated an interface dermatitis with dermal mucinosis. A necrotizing vasculitis with epithelial pustulation was seen in 2 patients. Interstitial GA-like infiltrates were seen in 5 cases. Immunofluorescent (IF) testing revealed a positive lupus band test (LBT) and epidermal nuclear and vascular staining for IgG and C5b-9 in the SLE patient. One DM patient had a negative LBT in concert with C5b-9 deposition along the dermoepidermal junction (DEJ) and within blood vessels while the other showed endomysial vascular Cs5b-9 deposition. In all patients, skin biopsy material contained B19 genome, which was absent in the serum of 4 patients analyzed. Symptomatic relief followed immunosuppressive and immunomodulatory therapy with agents including prednisone, cyclophosphamide, hydroxychloroquine, non-steroidal anti-inflammatory drugs and etanercept, but no patient has had complete symptom resolution. CONCLUSIONS: Persistent B19 infection may be of pathogenetic importance in certain prototypic CTD syndromes, to which underlying immune dysregulation associated with a blunted IgM response to viral antigen may predispose. Anti-viral therapy might be worthy of consideration since traditional immunosuppressive therapy was unsuccessful in our cases. PMID- 11100811 TI - Angiocentric primary cutaneous T-cell-rich B-cell lymphoma: a case report and review of the literature. AB - BACKGROUND: T-cell-rich B-cell lymphoma (TCRBCL) is an uncommon B-cell lymphoma, which is characterized histologically by a small number of neoplastic B cells surrounded by large numbers of nonneoplastic T cells. Diagnosis is frequently difficult because the neoplastic 'B-cell population may be quite sparse, and immunohistochemical and molecular analysis to identify the B-cell origin and clonality of the cells is necessary. METHODS: In this report we present an additional case of primary cutaneous TCRBCL with immunohistochemical and gene rearrangement studies and review the literature of this unusual entity. RESULTS: The patient was a 52-year-old woman who had a slowly growing cutaneous lymphoma which fulfills the strictest criteria initially proposed for a diagnosis of TCRBCL. This case has additional features of a prominent angiocentric/angioinvasive histology and dual clonality, showing both immunoglobulin heavy chain and T-cell receptor gene rearrangements. CONCLUSIONS: Controversy surrounds the notion of TCRBCL as a distinct clinicopathological entity. There have only been 14 reported cases of TCRBCL with skin involvement in the literature and only seven of these cases have arisen in the skin. To better understand the histological and clinical characterisitcs of TCRBCL we have reviewed previously reported cases of TCRBCL with skin involvement, and contrast them with the present case. PMID- 11100812 TI - Palisaded angioleiomyoma. AB - BACKGROUND: We report a case of a palisaded angioleiomyoma, a histopathologic variant of angioleiomyoma with prominent Verocay body formation. RESULTS: A healthy 31-year-old male requested removal of a subcutaneous nodule on the back of his head. The striking Verocay-like body formation of this tumor led to an initial frozen section diagnosis of neurilemoma. On hematoxylin and eosin staining, areas consistent with an angioleiomyoma were found as well. Immunohistochemistry gave positive reactions for actin and desmin, but was negative for S-100. CONCLUSIONS: To our knowledge this morphologic pattern in an angioleiomyoma has not been previously described. Palisaded angioleiomyoma should be added to the ever-expanding list of tumors that demonstrate nuclear palisades. PMID- 11100813 TI - Cutaneous epithelioid malignant nerve sheath tumor with rhabdoid features: a histologic, immunohistochemical, and ultrastructural study of three cases. AB - INTRODUCTION: Malignant rhabdoid tumors are morphologically defined as sheets of loosely cohesive cells with eccentric nuclei and hyaline, paranuclear inclusions. Although originally described as a distinctive renal neoplasm of childhood, these tumors have since been described in all age groups and in a variety of extrarenal sites. In the latter setting, it is thought that the rhabdoid phenotype is comprised of histogenetically unrelated tumors, that regardless of histogenesis, pursue a biologically aggressive behavior. METHODS: We report on the clinical, histologic, immunophenotypic, and ultrastructural characteristics of three cases of cutaneous malignant rhabdoid tumor. RESULTS: Each of the cases arose on the trunk or the extremity of elderly men. None of the patients had neurofibromatosis. All of the lesions histologically showed sheets of loosely cohesive polygonal cells with eccentric nuclei and hyaline paranuclear inclusions. Each of the cases showed the following immunophenotype: S-100 (+), synaptophysin(+), vimentin (+), alpha smooth muscle actin (-), CD-30 (-), HMB-45 (-), and pankeratin(-). Ultrastructure of two of the cases yielded similar results showing paranuclear filamentous aggregates of intermediate filaments, cell membrane dense plaques, and rudimentary cell junctions consistent with nerve sheath differentiation. Tonofilaments, dense bodies, microtubules, neurosecretory granules, and melanosomes were not identified. Each of the patients died of widely metastatic disease within 1 year of diagnosis. CONCLUSIONS: Cutaneous epithelioid malignant nerve sheath tumor is a potentially aggressive tumor capable of showing rhabdoid differentiation thus simulating a variety of neoplasms. Immunophenotyping and ultrastructural analysis reliably discriminates these lesions from melanoma, de-differentiated carcinoma, lymphoma, and rhabdomyosarcoma. PMID- 11100814 TI - Cellophane coverslips. PMID- 11100815 TI - Announcement of the Winner of the 1999 Munksgaard and Journal of Cutaneous Pathology Prize. PMID- 11100816 TI - Boron neutron capture therapy of brain tumors: functional and neuropathologic effects of blood-brain barrier disruption and intracarotid injection of sodium borocaptate and boronophenylalanine. AB - Sodium borocaptate (BSH) and boronophenylalanine (BPA) are two drugs that have been used clinically for boron neutron capture therapy (BNCT) of brain tumors. We previously have reported that hyperosmotic mannitol-induced disruption of the blood-brain barrier (BBB-D), followed by intracarotid (i.c.) administration of BPA or BSH, either individually or in combination, significantly enhanced tumor boron delivery and the efficacy of BNCT in F98 glioma bearing rats. The purpose of the present study was to determine the short-term neuropathologic consequences of this treatment and the long-term effects on motor and cognitive function, as well as the neuropathologic sequelae 1 year following neutron capture irradiation. BBB-D was carried out in non-tumor bearing Fischer rats by infusing a 25% solution of mannitol i.c. followed by i.c. injection of BPA or BSH, either individually or in combination, immediately thereafter. Animals were euthanized 2 days after compound administration, and their brains were processed for neuropathologic examination, which revealed sporadic, mild, focal neuronal degeneration, hemorrhage, and necrosis. To assess the long-term effects of such treatment followed by neutron capture irradiation, non-tumor bearing rats were subjected to BBB-D after which they were injected i.c. with BPA (25 mg B/kg body weight (b.w)) or BSH (30 mg B/kg b.w.) either individually or in combination (BPA 12.5 mg and BSH 14 mg B/kg b.w.). Two and a half hours later they were irradiated at the Medical Research Reactor, Brookhaven National Laboratory, Upton, NY, with the same physical radiation doses (5.79, 8.10 or 10.06 Gy), delivered to the brain, as those that previously had been used for our therapy experiments. The animals tolerated this procedure well, after which they were returned to Columbus, Ohio where their clinical status was monitored weekly. After 1 year, motor function was assessed using a sensitive and reliable locomotor rating scale for open field testing in rats and cognitive function was evaluated by their performance in the Morris water maze, the results of which were similar to those obtained with age matched controls. After functional evaluation, the rats were euthanized, their brains were removed, and then processed for neuropathologic examination. Subtle histopathologic changes were seen in the choroid plexuses of irradiated animals that had received BPA, BSH or saline. Radiation related ocular changes consisting of keratitis, blepharitis, conjunctivitis and cataract formation were seen with similar frequency in most rats in each treatment group. Based on these observations, and the previously reported significant therapeutic gain associated with BBB-D and i.c. injection of BSH and BPA, the present observations establish its safety in rats and suggest that further studies in large animals and humans are warranted. PMID- 11100818 TI - Bcl-2 expression in higher-grade human glioma: a clinical and experimental study. AB - Bcl-2 protein plays an important role in inhibiting apoptosis and protecting normal and neoplastic cells from toxicity. Bcl-2 overexpression in malignant tumors, on the other hand, may cause resistance against adjuvant treatment. Since there are subpopulations of patients with glioma that differ considerably in their treatment benefit, it is important to identify prognostic factors for outcome and to tailor adjuvant protocols in accordance with specific biological features of the respective tumor. The present study aimed at investigating the role of bcl-2 expression in higher-grade glioma (WHO grade III and IV). Bcl-2 expression was correlated with clinical and paraclinical parameters, and evaluated in univariate and multivariate statistical models. In addition, bcl-2 overexpressing human glioma cells in culture were used for modeling the in vivo findings and for investigating the importance of bcl-2 for tumor resistance against cytotoxic treatment. A group of 86 patients with higher-grade glioma were investigated. Anaplastic astrocytoma (AA; WHO G III, n = 29) showed bcl-2 expression in 48% of the cases, and immunohistochemical positivity was associated with a significantly shorter survival time (p = 0.0068). In glioblastoma patients (GBM; WHO G IV, n = 57), 51% of tumors were bcl-2 positive, but bcl-2 expression did not correlate significantly with survival (p = 0.39). In a Cox proportional hazards regression model, bcl-2 positivity was confirmed as a negative prognostic parameter in AA, but not in GBM. Bcl-2 overexpressing and control human glioma cell clones (T98MG line) were treated in culture with the cytotoxic drugs carmustine (BCNU), paclitaxel, vincristine, and doxorubicin. In addition, bcl-2 overexpressing and control cells were infected with a retrovirus carrying the herpes-simplex-virus thymidine kinase gene (HSV-tk), and then treated with ganciclovir (GCV). Bcl-2 overexpression significantly increased tumor cell resistance against all of the above cytotoxic drugs, and also against HSV-TK/GCV mediated gene therapy. PMID- 11100817 TI - Alteration in p53 modulates glial proteins in human glial tumour cells. AB - In transformed human glial cells, abnormalities of the p53 gene and altered expression of glial-specific properties (GSPs) have been observed. We therefore investigated whether (i) expression of the altered p53 protein is involved in the reduced expression of GSPs; and (ii) expression of the wild-type p53 (wt-p53) gene leads to induction of GSPs. We first determined that the p53 gene is mutated in human glioblastoma U-373MG cells. In these cells, and in human T-98G glioblastoma cells reported to possess a mutated p53 (m-p53) gene, nuclear m-p53 expression was intense while GSP expression was low in the same cell as revealed by double labelling immunocytochemistry. Conversely, glial fibrillary acidic protein (GFAP) and glutamate synthase (GS) were expressed in cells devoid of nuclear m-p53 immnunoreactivity. Therefore, a mutually exclusive relationship exists between the cytoplasmic GSPs and nuclear m-p53. Upon treatment with retinoic acid (RA) and dibutyryl cyclic AMP (dbcAMP), overall GSP staining were increased concomitant with suppression of nuclear m-p53. Their mutually exclusive expression pattern was maintained suggesting a functional relationship. This is supported by the observation of a similar mutually exclusive expression pattern for p53 and GSPs in pathologic specimens of human glioblastoma tissues. We then explored the role of the wt-p53 gene in the induction of GSPs using a wt-p53 tetracycline-regulated conditional expression system in human LN-Z308 glioblastoma cells. These cells normally express no p53 and no appreciable levels of GS or GFAP. Induced expression of wt-p53 lead to induction of GSP. These observations are consistent with the hypotheses that (i) nuclear m-p53 expression and cytoplasmic expression of GFAP and GS are inversely correlated, and (ii) expression of the wt-p53 gene leads to the expression of GSPs. PMID- 11100819 TI - Suppression of cell invasion on human malignant glioma cell lines by a novel matrix-metalloproteinase inhibitor SI-27: in vitro study. AB - Matrix metalloproteinase (MMP) has come to be highlighted by its close relation to the cell invasion of gliomas. Suppression of MMP activity in malignant glioma cells would be meriting to local delivery of genes or chemotherapeutic agents. In this study, we employed a novel MMP inhibitor, SI-27 to investigate inhibition of cell invasiveness in human malignant glioma cell lines, U87MG, U251MG, and U373MG. We evaluated with zymogram, reverse zymogram, and cell invasion assay after exposure of SI-27 for 24 h followed by preliminary MTT assay to find non cytotoxic dose range, 5, 10, 50, 100 microg/ml compared with non-treatment group as the control. Common to three glioma cell lines, zymogram disclosed that expressions of MMP-2 and -9 were suppressed in a dose-dependent fashion, meanwhile those of tissue inhibitor of MMP (TIMMP) in reverse zymogram were not. The numbers of invading cells through Boyden chamber were significantly reduced in a dose-dependent manner, while those with 5 microg/ml were not diminished common to those three lines. In conclusion, dose concentration ranging 10-100 microg/ml of SI-27 inhibited MMP-2 and -9 mediated cell invasiveness in malignant glioma cell lines. This is the first report for chemotherapeutic effect of SI-27 on glioma cells. PMID- 11100820 TI - Inhibition of the glioblastoma cell cycle by type I IFNs occurs at both the G1 and S phases and correlates with the upregulation of p21(WAF1/CIP1). AB - The antiproliferative effect of IFNalpha was tested on the human glioblastoma cell lines, U-373MG and T98G. IFNalpha significantly inhibited the growth of both cell lines, but was more effective in retarding the growth of U-373MG cells. Flow cytometry analysis indicated that synchronized IFNalpha-treated U-373MG cells showed a strong block in the progression of cells out of the S phase of the cell cycle. T98G cells, on the other hand, showed a moderate delay in the transition of cells from G1 to S phase and only a slight delay in the S phase, consistent with the decreased antiproliferative effect of IFNalpha on this cell line. IFNalpha-treated cells were then tested for the induction of the tumor suppressor gene product, p21(WAF1/CIP1). Higher levels of p21(WAF1/CIP1) were detected in lysates from IFNalpha-treated U-373MG cells as compared to media controls for as long as 18 h. In IFNalpha-treated T98G cells, p21(WAF1/CIP1) levels were slightly elevated at 4 and 6 h, but decreased to levels similar to controls thereafter, correlating with the antiproliferative effects of IFNalpha on each cell line. Immunoprecipitation studies on lysates from IFNalpha-treated U-373MG and T98G cells indicated that increased amounts of p21(WAF1/CIP1) were complexed to both cyclin D1 and cyclin E. Further, reduced cyclin-dependent kinase 2 (cdk2) activity was found in both IFNalpha-treated U-373MG and T98G cells, suggesting a mechanism by which p21(WAF1/CIP1) exerted its antiproliferative effects. Lastly, we analyzed the time-dependent production of the cyclins D1, E, and A. No differences in cyclin D1 levels were found between IFNalpha-treated and media treated U-373MG and T98G cells. However, both IFNalpha-treated U-373MG and T98G cells showed a prolonged elevation in cyclin E, correlating with the G1 to S phase delays observed in these cell lines. Further, the duration of cyclin E production corresponded with the magnitude of the cell cycle delays seen in IFNalpha-treated U-373MG and T98G cells. Prolonged elevation of cyclin A was also seen in both IFNalpha-treated U-373MG and T98G cells, the magnitude of which correlated with the S phase delay observed in these cell lines. Thus, the data indicate that IFNalpha has significant antiproliferative activity against glioblastoma cells that is mediated, at least in part, by the tumor suppressor gene product, p21(WAF1/CIP1). PMID- 11100821 TI - Quality of life following surgery for intracranial meningiomas at Brigham and Women's Hospital: a study of 164 patients using a modification of the functional assessment of cancer therapy-brain questionnaire. AB - The objective of this study was to determine the subjectively reported quality of life (QOL) of patients with meningiomas surgically treated. Demographic, medical and outcomes data on 164 patients were retrospectively analyzed with the use of the Brain Tumor Center database at the Brigham and Women's Hospital, Boston, MA. The patients were contacted via a telephone survey and were asked 26 standardized QOL questions based on a modification of the validated Functional Assessment of Cancer Therapy-Brain (FACT-BR) Study, which used only questions adjuvant to brain tumor patients. The patients' ages ranged from 23 to 87 years. The mean follow-up time after intracranial surgery was 33 months and median follow-up time was 28 months, with a range of 0 to 165 months. Of those 164 patients still living, 95% (155) participated in the telephone survey. 80% reported being satisfied with their post-treatment quality of life; 86% reported that they could write, read, drive and return to work at their pre-morbid level of functioning; 87% described themselves as 'independent' and able to act on their own initiatives. Our study found a high level of satisfaction for QOL in patients who have undergone surgery for intracranial meningiomas. Patients, by their own report, are able to lead independent, personally satisfying, meaningful and productive lives. This provides useful information to share with patients in discussions regarding surgical treatment of these lesions. PMID- 11100822 TI - Asymptomatic brain metastases (BM) in small cell lung cancer (SCLC): MR-imaging is useful at initial diagnosis. AB - PURPOSE: In this study we evaluated the usefulness of MR-imaging in the detection of asymptomatic brain metastases (BM) at the initial diagnosis in patients with small cell lung cancer (SCLC) and studied the follow-up of these patients. PATIENTS AND METHODS: One-hundred and twenty-five patients with SCLC were investigated with MR-imaging. RESULTS: In 112 patients with normal neurological findings, MR-imaging of the brain demonstrated BM in 17 patients (15%). Six of these 17 patients were therefore upgraded to extensive disease (ED). Two of these 17 patients died during chemotherapy because of progressive disease and 3 patients became neurologic symptomatic with progressive disease on MR-imaging of the brain. After completion of chemotherapy a repeated MR-imaging of the brain in the remaining 12 patients showed 1 complete remission, 4 partial remission and 7 progressive disease of the BM. CONCLUSION: This study showed that at presentation an unexpectedly high percentage of SCLC patients had asymptomatic BM on MR imaging. We propose that MR-imaging of the brain should be included in the staging of SCLC patients as well for staging, prognosis and therapy. PMID- 11100823 TI - The effect of anticonvulsant drugs on blood levels of methotrexate. AB - The reduced bioavailability of chemotherapeutic agents is one of the reasons that explains the limited efficacy of adjuvant chemotherapy in high grade glioma patients. We report how even the results of high dose sequential chemotherapy can be influenced by antiepileptic drugs. PMID- 11100824 TI - Zentralblatt fur Bakteriologie--100 years ago. First studies on the nucleus of bacteria. PMID- 11100825 TI - The evolution of human pathogens: examples and clinical implications. AB - Recent advances in sequencing of complete bacterial genomes, molecular typing of micro-organisms, and research on microbial pathogenicity factors changed our view on the evolution of human bacterial pathogens. We review current evolutionary concepts on plague and meningococcal disease to illustrate the interplay of molecular phylogeny, epidemiology, and pathogenicity research. Furthermore, examples of the tremendous velocity of bacterial evolution under changing environmental conditions will be discussed. PMID- 11100826 TI - Pathogenicity islands and phage conversion: evolutionary aspects of bacterial pathogenesis. AB - Horizontal gene transfer plays a key role in the generation of novel bacterial pathogens. Besides plasmids and bacteriophages, large genomic regions termed pathogenicity islands (PAIs) can be transferred horizontally. All three mechanisms for DNA exchange or transfer may be important for the evolution of bacterial pathogens. PMID- 11100827 TI - ShlB mutants of Serratia marcescens allow uncoupling of activation and secretion of the ShlA hemolysin. AB - The ShlB protein in the outer membrane of Serratia marcescens secretes hemolytic ShlA protein into the culture medium. In the absence of ShlB, nonhemolytic ShlA remains in the periplasm. ShlB mutants were isolated in which secretion was uncoupled from activation. Mutants with a tetrapeptide insertion after residues 136 or 224 of mature ShlB and a mutant with an insertion after residue 154 and a deletion secreted inactive ShlA. In vitro, secreted nonhemolytic ShlA was converted into hemolytic ShlA by isolated wild-type ShlB and by complementation with an N-terminal ShlA fragment of 255 residues (ShlA-255). The isolation of secretion-competent, but activation-negative mutants indicates that secretion alone is not sufficient for activation of ShlA. Rather, ShlB is required for activation and secretion, and the mutants define sites in ShlB which are involved in activation. According to a predicted transmembrane model of ShlB, the mutations that retain secretion competence but abolish activation competence are located in the most prominent surface loop and the following transmembrane loop. In one tetrapeptide insertion mutant, ShlB-332, most of the ShlA remained cell associated in an inactive form and low amounts (6%) were hemolytic. Secreted inactive ShlA(o) was completely degraded by trypsin, in contrast to hemolytic ShlA, which was cleaved into two fragments of 60 and 100 kDa. This result indicates that the conformational change from a highly trypsin-sensitive to a highly trypsin-resistant protein with only a single cleavage site in a polypeptide of 1,578 residues occurs upon activation of ShlA and not during secretion. PMID- 11100828 TI - Characterization of intestinal cnf1+ Escherichia coli from weaned pigs. AB - Escherichia coli isolated from 204 cases of porcine postweaning diarrhoea were tested by PCR for the genes of cytotoxic necrotic factors (CNF) and of cytolethal dystending toxin (CDT). Selected strains were also examined by PCR for the presence of papC-, sfa-, f17-, f18-, and afa-specific sequences encoding P, S, F17, F18 fimbriae and afimbrial adhesins. A 5.9% (12/204) of the strains had cnf1 gene, and two of them had cdt gene as well. Further six cdt+ strains were detected which were cnf-negative. Most of the cnf1+ strains belonged to serogroups O2, O6, O8, O54 characteristic of necrotoxic E. coli (NTEC) of humans. All the cnf1+ strains possessed the genes for P or S fimbriae or both, but were negative for F4, F17, or F18 or afimbrial adhesins. Results suggest that these enteric isolates may have entero- and/or uropathogenic significance in weaned pigs, and may have zoonotic potential for humans. PMID- 11100829 TI - Linkage of determinants for streptogramin A, macrolide-lincosamide-streptogramin B, and chloramphenicol resistance on a conjugative plasmid in Enterococcus faecium and dissemination of this cluster among streptogramin-resistant enterococci. AB - A new streptogramin A resistance gene, satG (= vatE), has been recently identified in Enterococcus faecium UW1965 (Werner and Witte 1999. Antimicrob. Agents Chemother. 43: 1813-1814). Further sequence analysis of this plasmid revealed that vatE is in a cluster together with other resistance genes. The identified ORFs were nearly identical with the already known genes ermB and cat. The ermB fragment exhibited more than 99% identity with a resistance region from the streptococcal plasmid pIP501, whereas the cat fragment also contained a truncated rep gene homologue with more than 99% identity to sequences in small staphylococcal plasmids. The cat-rep and the ermB-vatE segments were linked by an IS1216V insertion sequence widely distributed among enterococci. PCR analysis of additional 76 streptogramin-resistant isolates possessing vatE and ermB revealed a linkage of both genes in 45 isolates (59%); 15 of them with a gene arrangement, cat-repU-IS1216V-ermB-vatE, identical to the reference strain UW1965. An identical linkage of IS1216V-ermB-vatE was found among isolates from poultry manure, poultry meat, stool samples of humans, and hospital patients indicating a possible spread of the resistance gene cluster via the food chain to humans. PMID- 11100830 TI - Molecular characterization and influence on fungal development of ALP2, a novel serine proteinase from Aspergillus fumigatus. AB - A novel subtilisin-related serine proteinase (ALP2) [EC 3.4.21.48] with a broad range of activity between pH 4.5 and 11.0 was released from a cell wall fraction of Aspergillus fumigatus by an alkaline pH shift. The enzyme which was not detected in the culture supernatant was partially purified by phenylbutylamine agarose chromatography. The N-terminal sequence revealed that ALP2 is the same protein identified as the major allergen of A. fumigatus in patients suffering from extrinsic bronchial asthma (Shen et al. 1999, Int. Arch. Allergy Immunol. 119, 259-264). Based on this N-terminal sequence and on a conserved region of fungal subtilisins, a specific PCR probe was generated and the ALP2 genomic and cDNA were isolated from corresponding phage libraries. ALP2 shares a 49% identity with the vacuolar proteinase B (PrB) of Saccharomyces cerevisiae. In addition there is a 78% identity with PEPC, a serine proteinase which has been described in Aspergillus niger. Targeted disruption of the ALP2-encoding gene resulted in a slightly decreased speed of vegetative growth and in a more than 80% reduction of sporulation in the alp2-negative mutants, correlated with an approximately 50% reduction of the median diameter of conidiophore vesicles. The requirement of ALP2 for regular sporulation, in addition to its role in allergic asthma, raises further interest in cellular proteinases in respect to morphogenesis and pathogenesis in A. fumigatus. PMID- 11100831 TI - Detection of three genospecies of Borrelia burgdorferi sensu lato in Ixodes ricinus ticks collected in different regions of Poland. AB - Ixodes ricinus ticks, collected in 1996-1998 in different Polish woodlands, were examined to assess the frequency of the occurrence of Lyme borreliosis-associated genospecies. A total of 568 samples of individual adults and 162 samples of individual (n =48) and pooled (of 2 to 7) samples of nymphs were analysed by the polymerase chain reaction (PCR) for Borrelia burgdorferi sensu lato. Spirochetes were detected in 130 adult ticks (22.9%) and in a minimum of 32 (5.3%) nymphs. Further identification of 153 B. burgdorferi s.l.-positive samples by nested PCR using three species-specific primers revealed the occurrence of B. afzelii, B. burgdorferi sensu stricto and B. garinii. Both single-species and mixed infections were noted. Single-species infections were observed in the majority of samples (n = 83/153; 54.2%). Within this group B. afzelii was found in 38/153 samples (24.9%), followed by B. burgdorferi sensu stricto (n = 23/153; 15.0%) and B. garinii (n = 22/153; 14.4%). Dual infections with B. burgdorferi s.s. and B. afzelii were detected in 17/121 (14.0%) adults, while both B. burgdorferi s. s./B. garinii and B. afzelii/B. garinii coinfected 11/121 (9.1%) adult ticks. Triple infection with B. burgdorferi s.s., B. afzelii and B. garinii was noted twice (1.6%). In general, B. afzelii was found in 72/153 (47.1%) tick samples and was the predominant species. B. burgdorferi s. s. and B. garinii were detected in a total of 60/153 (39.2%) and 51/153 (33.3%) samples, respectively. Although, 21 (13.7%) samples were infected by B. burgdorferi s.l. genospecies undetectable by the primers used, results of our study confirm that Lyme borreliosis pathogenic genospecies are well established in tick populations throughout Poland. PMID- 11100832 TI - Signaling pathways in cell death and survival after photodynamic therapy. AB - Photodynamic therapy (PDT) is a cytotoxic treatment, which can induce cells to initiate a rescue response, or to undergo cell death, either apoptosis or necrosis. The many signaling pathways involved in these processes are the topic of this review. The subcellular localization of the photosensitizer has been shown to be a key factor in the outcome of PDT. Mitochondrial localized photosensitizers are able to induce apoptosis very rapidly. Lysosomal localized photosensitizers can elicit either a necrotic or an apoptotic response. In the plasma membrane, a target for various photosensitizers, rescue responses, apoptosis and necrosis is initiated. Several protein phosphorylation cascades are involved in the regulation of the response to PDT. Finally, a number of stress induced proteins play a role in the rescue response after PDT. Notably, the induction of apoptosis by PDT might not be crucial for an optimal outcome. Recent studies indicate that abrogation of the apoptotic pathway does alter the clonogenic survival of the cells after PDT. Further studies, both in vitro and especially in vivo could lead to more efficient combination therapies in which signaling pathways, involved in cell death or rescue, are either up- or downregulated before PDT. PMID- 11100833 TI - Photodynamic action of amino substituted hypocrellins: EPR studies on the photogenerations of active oxygen and free radical species. AB - A novel method has been employed to prepare 2-butylamino-2-demethoxy hypocrellin A (BADMHA) and 2-butylamino-2-demethoxy hypocrellin B (BADMHB). Both compounds exhibit stronger absorption at the phototherapeutic window (600-900 nm). The spin trapping and spin counteraction studies have shown that they are both efficient generators of the active oxygen (1O2, O2*-) in the aerobic condition. Under the anaerobic condition they generate non-oxygen free radical (semiquinone radical anion), and the active oxygen mechanism of photosensitization can be converted into non-oxygen free radical mechanism with the depletion of oxygen. The quantum yields of 1O2 generation of BADMHA and BADMHB are 0.46 and 0.44, respectively. Both are lower than those of their parent compounds HA and HB. But the productions of superoxide anion are enhanced significantly compared with HA and HB, indicating they are both favorable Type I phototherapeutic agents. PMID- 11100834 TI - Pharmacokinetics and pharmacodynamics of tetra(m-hydroxyphenyl)chlorin in the hamster cheek pouch tumor model: comparison with clinical measurements. AB - The pharmacokinetics (PK) of the photosensitizer tetra(m-hydroxyphenyl)chlorin (mTHPC) was measured by optical fiber-based light-induced fluorescence spectroscopy (LIFS) in the normal and tumoral cheek pouch mucosa of 29 Golden Syrian hamsters with chemically induced squamous cell carcinoma. Similar measurements were carried out on the normal oral cavity mucosa of five patients up to 30 days after injection. The drug doses were between 0.15 and 0.3 mg per kg of body weight (mg/kg), and the mTHPC fluorescence in the tissue was excited at 420 nm. The PK in both human and hamster exhibited similar behavior although the PK in the hamster mucosa was slightly delayed in comparison with that of its human counterpart. The mTHPC fluorescence signal of the hamster mucosa was smaller than that of the human mucosa by a factor of about 3 for the same injected drug dose. A linear correlation was found between the fluorescence signal and the mTHPC dose in the range from 0.075 to 0.5 mg/kg at times between 8 and 96 h after injection. No significant selectivity in mTHPC fluorescence between the tumoral and normal mucosa of the hamsters was found at any of the applied conditions. The sensitivity of the normal and tumoral hamster cheek pouch mucosa to mTHPC photodynamic therapy as a function of the light dose was determined by light irradiation at 650 nm and 150 mW/cm2, 4 days after the injection of a drug dose of 0.15 mg/kg. These results were compared with irradiations of the normal oral and normal and tumoral bronchial mucosa of 37 patients under the same conditions. The reaction to PDT of both types of human mucosae was considerably stronger than that of the hamster cheek pouch mucosa. The sensitivity to PDT became comparable between hamster and human mucosa when the drug dose for the hamster was increased to 0.5 mg/kg. A significant therapeutic selectivity between the normal and neoplastic hamster cheek pouch was observed. Less selectivity was found following irradiations of normal mucosa and early carcinomas in the human bronchi. The pharmacodynamic behavior of mTHPC was determined by test irradiations of the normal mucosa of hamsters and patients between 6 h and 8 days after injection of 0.5 and 0.15 mg/kg in the hamsters and the patients, respectively. The normal hamster cheek pouch showed a maximum response to irradiation 6 h after injection and then decreased continuously to no observable reaction at 8 days after injection. The reaction of the normal human oral mucosa, however, showed an increasing sensitivity to the applied light between 6 h and 4 days after mTHPC injection and then decreased again at 8 days. The hamster model with the chemically induced early squamous cell cancer in the cheek pouch thus showed some similarity to the early squamous cell cancer of the human oral mucosa considering the PK. However, a quantitative difference in fluorescence signal for identical mTHPC doses as well as a significant difference in pharmacodynamic behavior were also observed. The suitability of this animal model for the optimization of PDT parameters in the clinic is therefore limited. Hence great care must be taken in screening new dyes for PDT of early squamous cell cancer of the upper aerodigestive tract based upon observables in the hamster cheek pouch model. PMID- 11100835 TI - Contribution of LHC II complex to the electric properties of thylakoid membranes: an electric light scattering study of Chl b-less barley mutant. AB - Electric light scattering measurements demonstrate a strong decline in the permanent electric dipole moment and electric polarizability of both thylakoid membranes and photosystem II-enriched particles of the Chlorina f2 mutant which has severely reduced levels of light-harvesting chlorophyll a/b-binding proteins compared to the wild type barley chloroplasts. The shift in the electric polarizability relaxation to higher frequencies in thylakoids and photosystem II particles from Chlorina f2 reflects higher mobility of the interfacial charges of the mutant than that of the wild type membranes. The experimental data strongly suggest that the major light-harvesting complex of photosystem II directly contribute to the electric properties of thylakoid membranes. PMID- 11100836 TI - Phototaxis in the ciliated protozoan Ophryoglena flava: dose-effect curves and action spectrum determination. AB - The sensitivity of positive phototactic orientation of cells of the ciliated protozoan Ophryoglena flava has been measured for white light, broad-band blue and red light, and narrow-band monochromatic light, using a laboratory-developed computer aided system. The white-light fluence rate-response curve shows that there is no negative phototaxis in the fluence rate range investigated (0-15 W/m2) and no adaptation phenomena; it is very well fitted by a hyperbolic function; the fluence rate curves under broad band blue and red light (full width at half maximum, FWHM= 100 nm) can be fitted by the same model. The saturation level is, within experimental errors, the same for the three curves, indicating that there are no chromaticity effects and that if there is more than one photoreceptor pigment, they act independently of each other. The fluence rate response curves determined under narrow band monochromatic light (FWHM = 10 nm) can also be fitted by the same model and show, within experimental errors, the same saturation level. An action spectrum for positive phototaxis at 10-nm intervals has been calculated from fluence rate-response curves: it shows three maxima, at 420, 540 and 590 nm. This action spectrum is significantly different from the ones for photomotile responses in Blepharisma japonicum, Stentor coeruleus and Chlamydodon mnemosyne, whereas it resembles the ones of Paramecium bursaria and Fabrea salina. PMID- 11100837 TI - Long-range assemblies on poly(dG-dC)2 and poly(dA-dT)2: phosphonium cationic porphyrins and the importance of the charge. AB - The present paper describes synthesis and spectroscopic properties of novel cationic meso-tetraphenylporphyrins bearing two (trans) (P2) or three (P3) triphenylphosphonium substituents. The porphyrin aggregation in aqueous solutions is discussed in detail. Porphyrin binding to and self-organization onto long range assemblies on poly(dA-dT)2 or poly(dG-dC)2 were probed by combination of absorption, fluorescence, circular dichroism (CD), transient and resonance light scattering (RLS) techniques. The higher hydrophobicity of P2 is manifested by more extensive self-organization. Induced CD and intensive RLS indicate binding to the chiral environment on the nucleic acids exterior and exciton coupling between adjacent porphyrin moieties. The CD spectra of P2 on poly(dG-dC), and poly(dA-dT)2 suggest that the binding geometry is essentially independent of the base sequence. The fluorescence lifetime of about 4 ns was attributed to the long range assembly. In the case of P3 the distinctly different CD spectra induced by GC or AT base-pair regions reveal that the number of the substituents determines how closely the porphyrin can approach the specific electronic environment on the nucleic acid exterior. The fluorescence lifetime of the P3 assembly is about 2 ns. PMID- 11100838 TI - Changes of microsomal membrane properties in spring wheat leaves (Triticum aestivum L.) exposed to enhanced ultraviolet-B radiation. AB - The properties of microsomal membranes in spring wheat leaves (Triticum aestivum L. cv. Ganlong No. 92-005) exposed to (0) control, 8.64 (T1) and 11.2 kJ m(-2) day(-1) (T2) biologically effective UV-B irradiation (UV-B(BE)) were studied under greenhouse conditions. These irradiance levels correspond to a decrease in the stratospheric ozone of approximately 12.5 and 20%, respectively, for a clear solstice day at Lanzhou (36.04 degrees N, 1550 m), China. Compared with controls, the content of malondialdehyde (MDA) increased by 70.8% in T1 and 83.8% in T2 on the 7th day of the radiation, and the IUFA (index of unsaturated fatty acids) decreased, indicating peroxidation of lipid acids. Simultaneously, a drastic decrease of phospholipid content after 21 days and an increase of membrane lipid microviscosity on UV-B irradiation were also found, suggesting a reduction in the fluidity of membrane lipids. Ethylene emission by the microsomal membrane, in the presence of exogenous 1-aminocyclopropane-1-carboxylic acid was higher in the wheat seedlings after 7, 14 and 21 days' irradiation than in the controls. These changes were correlated with a rise in lipoxygenase activity. Membrane-bound enzymes (Ca2+ -ATPase and Mg2+ -ATPase) were promoted by UV radiation in the first 7 days and significantly decreased after 14 and 21 days' treatment in comparison to control. Our results suggest that UV-B radiation may cause changes in structural complexity and function of microsomal membranes in spring wheat leaves. PMID- 11100839 TI - Selective light-induced modulation of bcl-XL and bax expressions in indocyanine green-loaded U937 cells: effects of continuous or intermittent photo sensitization with low IR-light using a 805-nm diode laser. AB - The present study is concerned with the effects of an IR diode laser source emitting at 805 nm on a human leukaemic cell strain from a histiocytic lymphoma (U937) pre-loaded with the dye indocyanine green (ICG). The first aim of this work was the assessment of the earliest cellular defense events occurring upon ICG photosensitization. To this purpose, photosensitization was performed at low ICG concentration and low light energy density. The second aim was the comparative evaluation of the effects produced by continuous or fragmented irradiation. Independent of the irradiation method employed (continuous or fragmented), we could demonstrate that cells are forced to apoptosis, as indicated by the appearance in cell extracts of the 85-kDa proteolytic fragment of the poly(ADP-ribose)polymerase (PARP). This proteolysis, however, could be entirely prevented by a specific Caspase-3 inhibitor benzyloxycarbonyl-Asp-Glu Val-Asp-fluoromethylketone (Z-DEVD-fmk). Indeed the only perceptible change in the expression of pro/antiapoptotic proteins produced by continuous photostimulation was a small, albeit reproducible, increase in Bax. In contrast, when the photostimulation was achieved by means of several consecutive pulses, we observed not only a remarkable increase in Bax but also a noticeable abatement in Bcl-XL expression. The potential involvement of singlet oxygen in this process has been directly demonstrated by ICG photo-mediated oxidation of dithiothreitol in water. It has also been demonstrated that this oxidation is apparently more efficient when ICG is photostimulated by light pulses. PMID- 11100840 TI - Triplet state mechanism for electron transfer oxidation of DNA. AB - The interaction of anthraquinone-2-sulfonate with nucleotides and DNA in acetonitrile and acetonitrile water solvent mixture have been studied using KrF laser photolysis aimed at elucidation of the reaction mechanism. Laser spectroscopy directly demonstrates that the initial species from interaction of anthraquinone-2-sulfonate with nucleotides are radical cations of nucleotides and radical anion of anthraquinone-2-sulfonate. In addition, formation of ion pair from interaction of any of nucleotides with anthraquinone-2-sulfonate is synchronous with decay of triplet anthraquinone-2-sulfonate, which has provided dynamic evidence for initiation of electron transfer from DNA bases to triplet anthraquinone-2-sulfonate. Moreover, direct observation of stabilized DNA guanyl radical cation from interaction of anthraquinone-2-sulfonate with DNA has provided initial evidence for selective cleavage of DNA at guanine moiety. The solvent-separated ion pairs in acetonitrile have evidently dissociated into free ions, thereby enabling independent study of the behavior of guanyl radical cations and radical anion of anthraquinone-2-sulfonate. PMID- 11100841 TI - Standardization of capillary zone electropherograms obtained by using field enhanced sample stacking. AB - Migration times in a capillary zone electropherogram obtained by using the field enhanced sample stacking technique are strongly affected by the injected sample volume. That is, the migration times significantly decrease with the increase of the sample volume. To avoid inaccurate qualitative analysis due to the above phenomena, the time axis of the electropherograms was converted into an effective mobility axis using our conversion method taking account of the temperature increase in the separation tube and relaxation of the potential gradient of the separation field. After the conversion, accurate qualitative analysis was possible in spite of drastic change of the migration time, suggesting our conversion method could be successfully used for the standardization of electropherograms obtained even by using the stacking effect. The cause of the decrease of the migration time in the stacking process was briefly discussed. PMID- 11100842 TI - New method for standardization of electropherograms obtained in capillary zone electrophoresis. AB - A new method for standardization of electropherograms obtained by capillary zone electrophoresis was proposed, where the migration time axis was replaced by the effective mobility axis. The mobility increase due to temperature increase by Joule heating and the relaxation effect of the potential gradient were eliminated successfully by introducing a temperature coefficient for mobility expression and a delay time, respectively. The precision of the mobility evaluated by the proposed conversion methods was evaluated for a model sample. By using the conversion method, almost the same electropherograms could be obtained even from the electropherograms originally obtained by using different hardware conditions. PMID- 11100843 TI - Integration of a microextraction system solvent extraction of a Co-2-nitroso-5 dimethylaminophenol complex on a microchip. AB - A newly designed microchannel for solvent extraction was fabricated in a quartz glass chip and applied to solvent extraction of a Co-2-nitroso-5 dimethylaminophenol complex. The aqueous solution of Co complex and toluene were introduced into the microchannel, and the Co complex extracted in toluene was detected by thermal lens microscopy (TLM). The Co complex was quickly extracted into toluene when the flow was stopped. The observed extraction time, ca. 50 s, was almost equivalent to the value calculated using the diffusion distance and diffusion coefficient. The dependence of the TLM signal on the concentration of the Co complex showed good linearity in the range of 1 x 10(-7) - 1 x 10(-6) M. PMID- 11100845 TI - Life-time studies with capillary electrochromatography columns operated under different conditions. AB - A test system has been established to permit the monitoring of the life-time performance of several reversed- phase capillary electrochromatography (CEC) columns. The retention factors, k(cec), peak symmetry coefficients, lambda(sym), and column efficiencies, N, of three neutral n-alkylbenzene analytes, namely ethyl-, n-butyl- and n-pentylbenzenes, were determined for Hypersil 3 microm n octylsilica and n-octadecylsilica packed into CEC capillary columns of 100 microm I.D., with a packed length of 250 mm, and a total length of 335 mm. The performances of these CEC capillary columns were examined for a variety of eluents with pH values ranging between pH 2.0 - 8.0, similar to those employed to study the retention behaviour of peptides that we have previously reported. The relative standard deviation (RSD) of the retention factors (k(cec) values) of these n-alkylbenzenes, acquired with an eluent of (25 mM Tris-HCl, pH 8.0,) acetonitrile (1:4, v/v), when the CEC capillary columns were used for the first time (virgin values), were 4% (based on data acquired with 4 CEC capillary columns) for the n-octyl bonded silica capillary columns, and 6% (based on 8 columns) for n-octadecyl bonded silica capillary columns. The RSD values of the k(cec) values of the n-alkylbenzenes for one set of replicates (n=6) with one CEC capillary column was < 0.5%. The theoretical plate numbers, N, for the virgin CEC capillary columns were ca. 60,000, whilst the observed N values for all new CEC capillary columns were > or = 40,000 for n-octyl bonded silica capillary columns and > or = 50,000 for n-octadecyl bonded silica capillary columns. The peak symmetry coefficients, lambda(sym), of the n-alkylbenzenes for virgin CEC capillary columns and for CEC capillary columns used for more than 1,000 injections were always in the range 0.95-1.05. The experimental results clearly document that the life-time performance of the CEC capillary columns depends on the eluent composition, as well as the nature of the analytes to which the CEC capillary columns are exposed. PMID- 11100844 TI - Comparison of separation selectivity in capillary electrokinetic chromatography using a cationic linear polymeric pseudo-stationary phase or monomeric additives of similar structure. AB - The retention properties in electrically driven systems with monomeric additives were compared to an electrokinetic chromatographic system with a linear, charged polymer of similar chemical structure (all additives are quaternary tetraalkyl ammonium ions). The monomeric additives were tetramethylammonium (TMA), tetraethylammonium (TEA) and dimethylpyrrolidinium (DMP), respectively, the polymeric additive was poly(diallyldimethyl)ammonium (PDADMA). The additive concentration in the background electrolyte was 2 and 4% (w/w). The retention characteristics were based on the apparent mobilities of 10 non-charged analytes with different chemical functionality, which were transported by the anodic electroosmotic flow in the dynamically coated capillary, and retained by the counter-flowing cationic additives. From these data capacity factors were derived, which ranged up to 0.8. Association constants were calculated, and were found between 10 and 170. Roughly, the association constants increased for a given analyte in the sequence TMA 34 years and previous adverse obstetric history. Other factors that were observed in 1981 but which were not linked to preterm delivery in 1995 included the mothers being very young, single or foreign. However, parity and previous induced abortion were associated with preterm delivery in 1995, but not in 1981. These results show that the definition of high-risk groups used in prevention programmes should be brought up to date regularly. PMID- 11101019 TI - Extreme prematurity and school outcomes. AB - The purpose of this study was to assess the impact of extreme prematurity on three global measures of school outcomes. Using a matched cohort design, exposed infants comprised all surviving singleton infants < or = 28 weeks gestation born at one regional neonatal intensive care hospital between 1983 and 1986 (n = 132). Unexposed infants comprised randomly selected full-term infants (> or = 37 weeks gestation) frequency matched on date of birth, zip code and health insurance. All children were selected from a regional tertiary children's centre serving western New York population. Standardised telephone interviews elicited information on grade repetition, special education placement and use of school-based services. Unconditional logistic regression was used to estimate odds ratios (OR) and corresponding 95% confidence intervals (CI) adjusted for potential confounders for children without major handicaps. Extreme prematurity was associated with a significant increase in risk of grade repetition (OR = 3.22; 95% CI = 1.63, 6.34), special education placement (OR = 3.16; 95% CI = 1.14, 8.76) and use of school-based services (OR = 4.56; 95% CI = 1.82, 11.42) in comparison with children born at term, even after controlling for age, race, maternal education, foster care placement and the matching factors. These findings suggest that survivors of extreme prematurity remain at risk of educational underachievement. PMID- 11101020 TI - Gestational age- and birthweight-specific declines in infant mortality in Canada, 1985-94. Fetal and Infant Health Study Group of the Canadian Perinatal Surveillance System. AB - We studied infant mortality rates in Canada within specific gestational age and birthweight categories after using probabilistic techniques to link information in Statistics Canada's live births data base (1985-94) with that in the death data base (1985-95). Gestational age- and birthweight-specific mortality rates in 1992-94 were contrasted with those in 1985-87 with changes expressed in terms of relative risks with 95% confidence intervals [CI]. Statistically significant reductions in infant mortality were observed beginning at 24-25 weeks of gestation and extended across the gestational age range to post-term births. Crude infant mortality rates, infant mortality rates among those > or = 500 g and among those > or = 1000 g decreased by 22%, 25% and 26%, respectively, from 1985 87 to 1992-94. The magnitude of the reductions in infant mortality rates ranged from 14% [95% CI 7, 21%] at 24-25 weeks of gestation to 40% [95% CI 31, 47%] at 28-31 weeks. Almost all reductions in gestational age- and birthweight-specific infant mortality between 1985-87 and 1992-94 were due to approximately equal reductions in neonatal and post-neonatal mortality. Live births > or = 42 weeks of gestation did not follow this rule; post-neonatal mortality rates among such live births decreased significantly by 51% [95% CI 26, 68%], although neonatal mortality rates showed no significant change. The mortality reductions observed across the gestational age and birthweight range are probably a consequence of specific clinical interventions complementing improvements in fetal growth. Temporal changes in the outcome of post-term pregnancies need to be carefully examined, especially in relation to recent changes in the obstetric management of such pregnancies. PMID- 11101021 TI - Vaginal births after Caesarean (VBAC): a population study. AB - This paper describes delivery outcomes for women from Victoria, Australia, who gave birth in 1995 and whose immediately previous (penultimate) delivery, within a 5-year search period, was a Caesarean section. Because of the large numbers of records involved, dedicated computer software for record linkage was used to identify the previous delivery and link it with the woman's current birth in 1995. Overall, 79% of the records from multiparous women were linked successfully. Approximately 15% were not linked because the previous birth was before the search period or was an abortion that would not have been reported to the Perinatal Data Collection Unit. Reasons for not being able to link the last 6% of the records include the previous pregnancy being overseas or interstate. Women who had a vaginal birth as the penultimate birth or a multiple birth at either event were excluded, resulting in a study population of 4663 linked records. More women (68%) had a repeat Caesarean than went into labour and, of the remaining women who laboured, 56% delivered vaginally. Overall, 18% of the women delivered vaginally. For the women who went into labour, the reported number experiencing a uterine rupture was two per 1000 births. Uterine rupture was not reported in the two-thirds who did not labour but had a repeat Caesarean. A review of the perinatal deaths identified only two deaths, one baby being born by elective Caesarean and one by a vaginal birth after a previous Caesarean (VBAC) where the choice of delivery methods may have contributed to the death. This large study is one of the few in the literature to provide population-based information on vaginal births after a previous Caesarean and related outcomes. PMID- 11101022 TI - A critical evaluation of the evidence on the association between type of infant feeding and cognitive development. AB - This paper presents a critical evaluation of 24 studies on the association between type of infant feeding and cognitive development published over the past 20 years. Validity and generalisability of study findings were assessed according to three methodological standards: clearly defined outcome, specification of partial vs. exclusive breast feeding and control of confounding. Only six of the 24 investigations met all three standards. The most frequent study flaw was failure to distinguish between partial and exclusive breast feeding. Studies which made this distinction found larger IQ advantages to breast-fed infants than studies that did not. Four of the six studies meeting all three standards found an advantage in cognitive development to breast-fed infants of the order of two to five IQ points for term infants and eight points for low birthweight infants. We conclude that the question of whether breast feeding and formula feeding have differential effects on cognitive development has not yet been comprehensively answered. Research to date provides only an indication of the effect of relatively brief durations of partial breast feeding and even briefer durations of exclusive breast feeding. Future studies should measure breast feeding as a continuous dose-type variable, examine longer durations of breast feeding and control for a full range of confounders using techniques that deal appropriately with multicollinearity. PMID- 11101023 TI - Prevalence of usual-corrected distance visual acuity impairment in Hispanic and non-Hispanic children and adolescents. AB - Data from the Hispanic Health and Nutrition Examination Survey and the National Health and Nutrition Examination Survey I were analysed to determine the prevalence of visual acuity impairment among US Hispanics and non-Hispanics aged 6-19 years. The prevalence of 20/30 or worse distance visual acuity with usual or habitual correction ranged from 10.8% in non-Hispanic whites to 19.1% in Puerto Ricans. Puerto Rican boys aged 13-19 years had significantly greater rates of moderate or greater impairment (20/70 or worse) than 6-12-year-old Puerto Rican boys (5.7% vs. 0.7%). The prevalence of visual impairment was generally greater in girls than in boys. Assessment and comparison of refractive error and eye disease prevalence rates are necessary in future studies to determine factors influencing prevalence of visual acuity impairment in children. PMID- 11101024 TI - Epidemiology and predictors of infant morbidity in rural Malawi. AB - In rural Malawi, 703 newborns were visited monthly for 1 year to describe the epidemiology and health-seeking behaviour during acute episodes of diarrhoea, respiratory infections (ARI) and malaria. On average, the infants suffered from 1.3 annual episodes (11.0 illness days) of diarrhoea, 1.1 episodes (9.4 days) of ARI and 0.7 episodes (4.8 days) of malaria. Multivariate analysis with polychotomous logistic regression indicated that the amount of morbidity was associated with the child's area of residence, weight in early life, number of siblings, father's marital status and the source of drinking water. Diarrhoea and malaria were most common at 6-12 months of age and during the rainy months whereas respiratory infections peaked at 1-3 months of age and in the cold season. Ten per cent of diarrhoea, 9% of ARI and 7% of malaria episodes lasted for more than 14 days. Fifty-eight infants died, giving case fatality rates of 1% for diarrhoea, 2% for ARI and 4% for malaria. One-third (37%) of the illness episodes were managed at home without external advice. A traditional healer was consulted in 16% of episodes and a medical professional in 55% of episodes. If consulted, traditional healers were seen earlier than medical professionals (median duration after the onset of symptoms 0.7 vs. 1.8 days, P < 0.001). Traditional healers were significantly more commonly used by those families whose infants died than by those whose infants did not die (odds ratio 1.8, 95% CI 1.1, 3.0). Our results emphasise the influence of seasonality, care and living conditions on the morbidity of infants in rural Malawi. Case fatality for diarrhoea, ARI and malaria was high and associated with health-seeking behaviour among the guardians. Future interventions must aim at early and appropriate management of common childhood illnesses during infancy. PMID- 11101025 TI - Anorectal malformations and Down's syndrome. PMID- 11101026 TI - Authors' reply to the commentary Maternal birthweight and newborn status. PMID- 11101027 TI - Antenatal ultrasound exposure. PMID- 11101028 TI - Factors affecting the survivability of bovine oocytes vitrified in droplets. AB - Vitrification of bovine oocytes performed using the traditional, in straw system has not given satisfactory results. Although an alternative approach based on minimizing the volume of the vitrified sample has recently resulted in a much more promising survival rate of vitrified oocytes, we attempted to examine some additional factors influencing the survival and subsequent fertilization and development rates of bovine oocytes subjected to vitrification according to the minimum drop size approach. In total, 748 bovine, in vitro matured oocytes were vitrified using VS14 vitrification solution, containing 5.5-M ethylene glycol and 1.0-M sucrose after different pre-equilibration and equilibration protocols performed at 35 degrees to 37 degrees C. Experiment 1 showed no significant toxic effect during pre-equilibration treatments of oocytes in 2%, 4% or 6% ethylene glycol solutions, except the lower cleavage rate of oocytes exposed to 6% ethylene glycol (77.2% vs. 93.9% in control, P< 0.05). In Experiment 2, 12 to 15 min of pre-equilibration treatments in 0%, 1% or 2% ethylene glycol solutions were tested, followed by 30 or 45 sec of equilibration in VS 14 solution and vitrification in droplets of medium dropped directly into liquid nitrogen. The development rate of vitrified oocytes to the blastocyst stage tended to be higher after 30-sec equilibration treatment (9.5%, 13.9% and 13.8% in groups of oocytes pre-equilibrated in 0%, 1% or 2% ethylene glycol solutions, respectively). Experiment 3 tested pre-equilibration treatments in 0%, 1%, 2%, 3%, 4%, 5% or 6% ethylene glycol solutions, followed by 30-sec equilibration and vitrification in droplets. The highest cleavage, blastocyst and hatched blastocyst rates, which were not significantly different from control, were achieved in a group of oocytes pre-equilibrated in 3% ethylene glycol solution (76%, 30% and 15% vs. 89%, 42% and 21% in control, respectively). A healthy calf was born on Feb 22 1999, after transfer of 4 morula/blastocyst stage embryos developed from oocytes vitrified in droplets after pre-equilibration in 3% ethylene glycol solution. We conclude that gentle pre-equilibration of bovine oocytes in diluted, 3% ethylene glycol solution is an important factor improving the effectiveness of vitrification in droplets of bovine oocytes. PMID- 11101029 TI - Culture of in vitro produced bovine zygotes in vitro vs in vivo: implications for early embryo development and quality. AB - The objectives of this study were to examine the effect of culture system on bovine blastocyst formation rates and quality. Presumptive IVM/IVF bovine zygotes were cultured either in vitro in synthetic oviduct fluid (SOF, 25 embryos/25 microL in 5% CO2, 5% O2, 90% N2 at 39 degrees C) or in vivo in the ewe oviduct (approximately 100 embryos per oviduct). The recovery rate after in vivo culture was 53% (813/1,530). The blastocyst rate on Day 7 was significantly higher for the in vitro system (28%, 362/1,278 vs 17%, 37/813; P< 0.0001). However, after culture in vitro for a further 24 h, there was no difference in Day 8 yields (36%, 457/1,278 vs 32%, 258/813, for in vitro and in vivo culture, respectively). There was no difference in blastocyst cell number between treatments (Day 7: 96 vs 103; Day 8: 78 vs 85 for in vitro and in vivo culture, respectively). Irrespective of culture system, Day 7 blastocysts had a significantly higher cell number than those appearing on Day 8. There was no difference in pregnancy rate at Day 35 after fresh transfer of a single Day 7 blastocyst (37.5%, 21/56 vs 45.3%/, 24/53 for in vitro and in vivo culture, respectively). After cryopreservation by freezing in 10% glycerol, VS3a vitrification or solid surface vitrification, the survival of in vitro cultured embryos was significantly lower than survival of embryos cultured in the ewe oviduct or those produced by superovulation of donors. In conclusion, these findings demonstrate that while bovine zygotes cultured in vitro are capable of rates of development similar to those of their in vivo cultured counterparts (in terms of Day 8 blastocyst yield, cell number and early pregnancy rate), there are significant differences in embryo cryosurvival. This suggests that current in vitro culture systems need to be improved to optimize embryo quality and pregnancy rates. PMID- 11101030 TI - Nuclear transfer in cattle using colostrum-derived mammary gland epithelial cells and ear-derived fibroblast cells. AB - To assess the developmental potential of nuclear transfer embryos in cattle using mammary gland epithelial (MGE) cells derived from the colostrum, we compared the effectiveness of cloning using those cells and fibroblast cells derived from the ear. The fusion rate of the enucleated oocytes with fibroblast cells (75 +/- 4%) was significantly higher than that with MGE cells (56 +/- 7%, P<0.05). There were no significant differences in the cleavage rate (85 +/- 3% vs. 91+/- 2%) or in the developmental rate to the blastocyst stage (35 +/- 6% vs. 35 +/- 5%) using MGE cells vs. fibroblast cells as donor nuclei (P>0.05). After transfer of blastocysts derived from nuclear transfer embryos produced using MGE cells and fibroblast cells, 13% (4/31) and 16% (6/37) of recipient heifers were pregnant on Day 42 as assessed by ultrasonography, respectively. Two of the 4 and 4 of the 6 recipients of embryos with MGE cell- and fibroblast cell-derived nuclei, respectively, aborted within 150 days of pregnancy. Four live female calves were obtained from MGE cells or fibroblast cells. However, one died from internal hemorrhage of the arteria umbilicalis. The other three calves were normal and healthy. There were no differences in the pregnancy rate or calving rate when using MGE cells vs. fibroblast cells. Microsatellite DNA analyses confirmed that the cloned calves were genetically identical to the donor cows and different from the recipient heifers. We conclude that colostrum-derived MGE cells have the developmental potential to term by nuclear transfer, and the efficiency of development of those cloned embryos was the same as that of embryos obtained using fibroblast cells as donor nuclei, although there was a significant difference in the fusion rate. This method using MGE cells derived from colostrum, which is obtained easily and safely from live adult cows, is more advantageous for cloning with somatic cells. PMID- 11101031 TI - First ovulation and ketone body status in the early postpartum period of dairy cows. AB - The effect of ketone body status on occurrence of first ovulation during early lactation was assessed in 84 multiparous dairy cows under field conditions. Animals were equally distributed across 8 farms and were controlled by the same herd fertility monitoring program. Cows were visited twice antepartum and 6 times postpartum at weekly intervals between 5:30 and 8:30 AM. On these occasions, body condition scores and milk yields were measured, blood and milk samples were taken, cows were gynecologically examined, and parameters of reproduction were determined. The onset of first ovulation was specified by milk progesterone determination and rectal palpation. Cows starting postpartum ovarian cyclicity within or after 30 d were classified as early and late responders (ER and LR, respectively). Resumption of the estrous cycle within 30 d postpartum is considered optimal under practical conditions, and classification based on this threshold value resulted in groups of equal size and equal distribution of ER + LR cows within farms. Ketone bodies measured were beta-hydroxybutyrate in serum and acetoacetate and acetone in serum and milk. Blood serum and milk ketone body concentrations during the first 6 wk of lactation were higher in LR than in ER, whereas plasma glucose and nonesterified fatty acid and milk fat, protein and urea concentrations did not differ between groups. Maximal concentrations of ketone bodies from parturition to first ovulation were better predictors of the onset of the estrous cycle than mean or minimal concentrations over the same period. Milk acetone and serum beta-hydroxybutyrate concentrations provided the most reliable information with regard to resumption of ovarian activity of all ketone bodies. PMID- 11101032 TI - Effects of spermatozoal concentration and post-thaw dilution rate on survival after thawing of dog spermatozoa. AB - The objectives of this study were to evaluate the effects and interactions of freezing dog semen using 4 different sperm concentrations (50 x 10(6), 100 x 10(6), 200 x 10(6) and 400 x 10(6) spermatozoa/mL) in 0.5-mL straws and diluting the thawed semen at 4 different rates (1:0, 1:1, 1:2 and 1:4) on post-thaw survival and longevity of dog spermatozoa during incubation at 38 degrees C. Fifteen ejaculates were collected from 12 dogs and pooled. The semen pool was divided into 4 aliquots containing respectively 4,200 x 10(6), 2,100 x 10(6), 1,050 x 10(6) and 525 x 10(6) spermatozoa, which were centrifuged. Sperm pellets were rediluted with TRIS-glucose-egg yolk extender containing 5% glycerol and 0.5% of Equex STM Paste to obtain the designated sperm concentrations. The semen was frozen in 0.5-mL straws 4 cm above liquid nitrogen (LN2). The straws were thawed at 70 degrees C for 8 sec and the contents of each straw were divided into 4 aliquots and diluted with TRIS buffer at 38 degrees C at rates of 1:0, 1:1, 1:2 and 1:4 (semen:buffer), respectively, making a total of 16 treatments. Sperm motility was subjectively evaluated after thawing and at 1-h intervals during 8 h of incubation at 38 degrees C. Plasma membrane integrity and acrosomal status were evaluated at 1, 3, 6, 12 and 18 h post-thaw using a triple-staining procedure and flow cytometry. For data pooled across the post-thaw dilution rate, motility was higher (P< 0.001) in samples frozen with 200 x 10(6) spermatozoa/mu. The integrity of sperm plasma membranes after 18 h incubation was higher (P<0.05) in samples frozen with 200 x 10(6) and 400 x 10(6) spermatozoa/mL. For data pooled across sperm concentration, samples diluted at a rate of 1:2 or 1:4 had better (P<0.001) motilities after 8 h of incubation than undiluted samples or those diluted at 1:1. The integrity of the sperm plasma membranes was higher (P<0.001) at increasing dilution rates. When the 16 treatments were compared, the best longevity was obtained when semen packaged at a concentration of 200 x 10(6) spermatozoa/mL was diluted immediately after thawing at 1:4 dilution rate. PMID- 11101033 TI - Sexual maturational changes circulatory inhibin concentration in relation to FSH concentration and testicular size in Suffolk and DLS rams. AB - Developmental patterns in immunoactive inhibin and FSH concentrations in peripheral blood were determined for Suffolk and DLS (Dorset x Leicester x Suffolk) rams born in January Blood samples were taken every 3 to 4 wk when testes were developing during puberty (5 to 44 wk of age) and redeveloping in early adulthood (17 to 23 months of age). Suffolk lambs had a greater average daily gain (195 vs. 143 g/day, P<0.01), and they developed larger testes (P<0.01) than DLS lambs. Inhibin and FSH concentrations peaked at about the same pubertal (8 wk) and early adult (19 or 20 months) ages in both breeds. Elevations in FSH were greater (P< 0.05) in Suffolk than DLS rams at each stage of development. The pubertal inhibin peak was nearly 70% larger (P<0.01) in DLS than Suffolk rams, and the early adult peak was comparable in rams of both breeds, but much smaller (P<0.01) than the pubertal peak. Nonetheless, inhibin was positively correlated (r=0.48 to 0.57) with FSH in both breeds during each developmental stage. Inhibin and testicular size were negatively correlated in Suffolk (r=-0.74) and DLS (r= 0.86) rams during puberty, and positively correlated in DLS rams (r=0.46) in early adulthood. We conclude that 1) inhibin concentrations are higher in juvenile rams at the time Sertoli cell numbers are being established than in adult rams during testicular recrudescence and 2) rises in FSH concentration participate in regulating corresponding rises in inhibin concentration in both stages of testicular development. PMID- 11101034 TI - Inhibition of apoptosis in cultured porcine granulosa cells by inhibitors of caspase and serine protease activity. AB - Protease inhibitors were used to test the hypothesis that caspases and other proteases were active during apoptosis in cultured porcine granulosa cells. Cells isolated from 3 to 6 mm follicles were cultured for 24 h in Dulbecco's modified Eagles medium: Hams F12 (1:11 containing 1% fetal bovine serum. Final inhibitor concentrations, added in 10 microL of dimethylsulfoxide, were 0, 1, 5, 25 and 125 microM. Cells with compromised plasma membrane integrity, identified by uptake ethidium homodimer, increased during culture in the absence of inhibitors from 37% to 43%. Apoptotic (A0) cells, identified by DNA fluorescence flow cytometry, increased (P < 0.05) from 1.7% to 29%. The serine protease inhibitor N-tosyl-L phenylalanine chloromethyl ketone (TPCK) at 125 microM was lethal increasing (P < 0.05) cells with compromised membranes to 92%. In response to TPCK, A0 cells decreased from 55% to 1.2%; progesterone and estradiol production were decreased by 94% and 98%, respectively. The general caspase inhibitor, benzyloxycarbonyl valinyl-alaninyl-aspartyl fluoro methylketone, decreased (P < 0.05) A0 cells linearly from 33% to 3 % between 0 and 125 microM without significant effect on steroidogenesis or on the percentage of cells with compromised plasma membranes. Other inhibitors only had a marginal effect on apoptosis; concentrations of > or = 1 microM decreased (P < 0.05) A0 cells from 29% to 18% to 21% and had no significant effect on membrane integrity or steroid production. We conclude that caspases are associated with apoptosis in cultured porcine granulosa cells. Death induced by TPCK was through a non-apoptotic mechanism. PMID- 11101035 TI - Effects of oocyte quality, oxygen tension, embryo density, cumulus cells and energy substrates on cleavage and morula/blastocyst formation of bovine embryos. AB - Various factors, such as quality of the oocyte, oxygen tension, embryo density, and kind of energy substrate during in vitro production of embryos may affect the rate of preimplantation embryo development. In the present study we used 12553 bovine oocytes aspirated from slaughterhouse ovaries to evaluate various culture conditions that would increase in vitro production of advanced stages of preimplantation embryos. The morphological quality of the oocyte based on the compactness and number of layers of cumulus cells had significant positive effects on the rates of in vitro maturation, fertilization and development to the morula and blastocyst stages. None of the corona-enclosed or nude oocytes progressed beyond the 8- to 16-cell stage. The level of oxygen (5 or 20%) did not affect the proportion of one-cell embryos undergoing cleavage or progressing to morula and blastocyst stages. The rate of development of one-cell embryos originating from inferior quality oocytes was significantly improved when cultured in groups of 40 instead of 20 embryos per 0.5 mL medium. In the presence of cumulus cells, glucose had beneficial effects on in vitro maturation and subsequent development of IVM-IVF zygotes. The presence of serum improved the rate of in vitro development of one-cell embryos. Minimum Essential Medium supplemented with energy substrates according to the findings of metabolic studies was less effective in supporting in vitro maturation and subsequent development than TCM-199. In conclusion, morphological grading of immature oocytes is an appropriate selection criterion for their developmental ability. Embryo yields from low quality oocytes can be increased by culturing them in large groups. Serum is not essential for in vitro generation of embryos but its addition improves rates of success. PMID- 11101036 TI - Induction of follicular wave emergence for estrus synchronization and artificial insemination in heifers. AB - The objective was to synchronize follicular wave emergence among cattle for synchronization of estrus and ovulation, and to determine pregnancy rate after AI at observed estrus. At random stages of the estrous cycle, a controlled internal drug release device (CIDR-B) was inserted intravaginally (Day 0) in 67 cross-bred beef heifers, and they were randomly allocated to receive either no further treatment (Control; n = 18); 5 mg of estradiol-17beta and 100 mg of progesterone im (E/P; n = 16); 100 microg im of GnRH (GnRH; n = 16); or transvaginal ultrasound-guided follicular ablation of all follicles > or = 5 mm (FA; n = 17). All heifers received a luteolytic dose of PGF (repeated 12 h later), and CIDR-B were removed on Days 9, 8, 6 or 5, in Control, E/P, GnRH or FA groups, respectively, so the dominant follicle of the induced wave was exposed to exogenous progesterone for a similar period of time in each group. Mean (+/- SEM) intervals (and range, in days) from treatment to follicular wave emergence in these groups were 3.5 +/- 0.6 (-2 to 8), 3.4 +/- 0.1 (3 to 4), 1.5 +/- 0.3 (-1 to 4), and 1.0 +/- 0.1 (0 to 2), respectively. Although the interval was longest (P<0.01) in the E/P and Control groups, it was least variable (P<0.01) in the E/P and FA groups. Intervals (and range, in days) from CIDR-B removal (and first PGF treatment) to estrus were 2.3 +/- 0.2 (1.5 to 4.5), 2.2 +/- 0.2 (1.5 to 3.0), 2.1 +/- 0.1,(1.5 to 3.5), and 2.5 +/- 0.1 (2.0 to 3.5), and to ovulation were 3.5 +/- 0.2 (2.5 to 5.5), 3.4 +/- 0.1 (3.0 to 4.5), 3.5 +/- 0.1 (2.5 to 4.5), and 3.8 +/- 0.1 (3.0 to 4.5), for Control, E/P, GnRH and FA groups, respectively (ns). The proportion of heifers displaying estrus was higher in the Control than in the FA group (94% versus 65%, P<0.05) and intermediate in EP and GnRH groups (87% and 75%). Heifers were inseminated approximately 12 h prior to ovulation (based on estrous behavior and ultrasound examinations). Pregnancy rates were 78%, 80%, 69% and 65% for Control, E/P, GnRH and FA groups, respectively (P=0.73). Results support the hypothesis that synchronous follicular wave emergence results in synchronous follicle development and, following progesterone removal, synchronous estrus and ovulation with high pregnancy rates to AI. The synchrony of estrus and ovulation in the E/P, GnRH and FA groups suggest that these treatments, in combination with CIDR-B, could be adapted to fixed-time insemination programs. PMID- 11101037 TI - Altered functional and immunophenotypical properties of neutrophilic granulocytes in postpartum cows associated with fatty liver. AB - The intention of the study was to analyze the relationship between liver triacyl glycerol content (liver TAG content) and immunophenotypical and functional properties of polymorphonuclear neutrophilic granulocytes (PMN) of dairy cows in the peripartum period. We investigated characteristics of bovine PMN from the blood and uterus of clinically healthy cows in the periparturient period. The numbers of circulating leukocytes and segmented granulocytes continuously increased until parturition and declined afterwards to starting values. This was independent of the liver TAG content and mainly affected neutrophils. The liver TAG content exceeded 40 mg/g liver, the reference value, in 12 of 19 cows in the first two weeks postpartum. Increased liver TAG content, > 40 mg/g, went in parallel with a reduced expression of function-associated surface molecules on blood neutrophils (e.g. CD11b/CD18 = CR3 and CD11c/CD18 = CR4). Moreover, in cows with high liver TAG levels the antibody-independent and -dependent cellular cytotoxicity (AICC, ADCC) of blood PMN was markedly reduced. PMN also were less capable of ROS generation after stimulation with Phorbol Myristate Acetate (PMA). In comparison with contemporarily harvested blood PMN, neutrophils recovered from the uterine lumen showed a decreased expression of 4/6 examined surface structures. Only the expression densities of CR3 molecules and those detected by mAb IL-A110 were enhanced on uterine PMN. The cytotoxic capacity and the ROS generation were significantly lower for uterine PMN than for blood PMN. The results suggest that increased liver TAG content in the first and second week after calving is associated with decreased functional capacities of PMN derived from blood and uterus. This may help to explain why cows who are too fat at calving (who therefore have an increased liver TAG content) have a higher incidence of infectious diseases such as endometritis PMID- 11101039 TI - A model for economic comparison of swine insemination programs. AB - Optimal artificial insemination schedules are those that result in a high farrowing rate and litter size, while minimizing costs of semen and labor by avoiding unnecessary inseminations. A simulation model programmed in a commercial spreadsheet was developed to permit comparison of alternative schedules. Farrowing rate and litter size for a particular schedule were dependent on the timing of insemination relative to the time of ovulation. Economic return was calculated by multiplying the number of pigs born per bred sow by $33.00 and subtracting the cost of producing a litter of pigs and raising them to weaning ($222.88 per sow plus $2.44 per pig born) and the cost of detection of estrus and breeding. Seven insemination schedules combined with once versus twice per day detection of estrus were simulated in 500 herds of 100 sows each. Inseminations were simulated to occur on schedules of: 1) 0, 12, 24 and 36 h; 2) 12, 24 and 36 h; 3) 0 and 24 h; 4) 12 and 36 h; 5) 12 h; 6) 24 h; and 7) 36 h after first detection of estrus. Schedule 1 was predicted to yield the highest farrowing rate and litter size. Economic return was highest for Schedule 2 with twice per day detection of estrus followed closely by Schedule 1 with once per day detection of estrus at $14.90 and $13.75 per bred sow, respectively. High performance was dependent on insuring that inseminations occurred at an optimum time in as great a proportion of sows as possible. PMID- 11101038 TI - Development and viability of pig oocytes matured in a protein-free medium containing epidermal growth factor. AB - This study examined the ability of epidermal growth factor (EGF) to improve the developmental competence of pig oocytes matured in a protein-free (PF) in vitro maturation (IVM) system. Oocyte maturation was done in one of three media: 1. PF TCM: tissue culture medium (TCM) 199 + 0.1% polyvinylalcohol (PVA); 2. PF TCM+EGF: PF-TCM + 10 ng/ml EGF; and 3. +ve CONT: North Carolina State University (NCSU) 23 medium + 10% porcine follicular fluid. All media contained 0.57 mM cysteine. Hormonal supplements, 0.5 microg/mL LH and 0.5 microg/mL FSH, were present only for the first half (20 to 22 h) of the culture period. After maturation, oocytes were co-incubated with frozen-thawed spermatozoa for 5 to 6 h and transferred to embryo culture medium, NCSU 23 containing 0.4% BSA, for 144 h. In Experiment 1, differences in cumulus expansion were observed for oocytes matured in +ve CONT (Category 4), PF-TCM (Category 2) and PF-TCM+EGF (Category 3). However, no significant differences in nuclear maturation to metaphase II stage were observed. In Experiment 2, no differences in fertilization parameters were observed. Significant (P < 0.01) differences in cleavage rates were observed among the three media for a proportion of the oocytes matured (52, 60 and 69% in PF-TCM, PF-TCM+EGF, and +ve CONT, respectively). Oocytes matured in PF-TCM showed the lowest (P < 0.01) blastocyst development (22%). However, the same rate of blastocyst development was obtained for +ve CONT (37%) and PF-TCM+EGF (37%). Blastocyst cell numbers were significantly higher when oocytes were matured in the presence of EGF (26 vs. 37 to 41). In Experiment 3, oocytes matured in PF TCM+EGF had a significantly (P < 0.05) higher intracellular glutathione (GSH) concentration (5.9 vs. 11.4 pmol/oocyte) compared with PF-TCM. Twenty-two of 25 embryo transfer recipients became pregnant (Experiment 4). Four animals returned to estrus in within 60 days. Six pregnant animals slaughtered at 26 to 45 days had 43 fetuses (range: 4 to 12) and the remaining 12 animals farrowed 82 piglets (range: 3 to 12). These results indicate that EGF enhances the developmental competence of pig oocytes matured in a protein-free culture medium which is correlated with higher GSH level in oocytes. Birth of piglets indicate that embryos derived from oocytes matured in the presence of EGF are viable. PMID- 11101040 TI - Effect of coconut water and Braun-Collins solutions at different temperatures and incubation times on the morphology of goat preantral follicles preserved in vitro. AB - Preservation of preantral follicles becomes very important to ensure follicle quality at the onset of cryopreservation or in vitro culture. However, for domestic animals, the ovarian donor of preantral follicles for in vitro studies is commonly encountered far away from reproduction laboratories. We investigated the effectiveness of coconut water and Braun-Collins solutions on the preservation of goat preantral follicles. At the slaughterhouse, the ovarian pair of each animal was divided into 19 fragments. One ovarian fragment was immediately fixed (Control - Time 0). The other 18 fragments were randomly distributed into tubes containing 2 mL of coconut water or Braun-Collins solution at 4 degrees, 20 degrees or 39 degrees C and then stored for 4, 12 or 24 h. Histological analysis showed that the storage of ovarian fragments in coconut water and Braun-Collins solutions at 20 degrees or 39 degrees C for 12 or 24 h significantly reduced (P < 0.05) the percentage of morphologically normal preantral follicles when compared with the control. However, storage in coconut water at 20 degrees C for 4 h and in both solutions at 4 degrees C kept the percentage at control values. Ultrastructural analysis of follicles exposed to the stated conditions confirmed the integrity of preantral follicles stored at 4 degrees C in Braun-Collins and coconut water solutions for up to 12 and 24 h, respectively. Reduced cellular metabolism at 4 degrees C may explain why the best preservation of preantral follicles was at 4 degrees C, which may suggest a useful method for ovary transport in the future. PMID- 11101041 TI - Comment on the critique of the published article by Visser et al. (Theriogenology. 1999; 51:689-697). PMID- 11101042 TI - Tissue and blood levels of zinc, copper, and magnesium in nitric oxide synthase blockade-induced hypertension. AB - The aim of this study was to determine the levels of tissue and blood zinc (Zn), copper (Cu), magnesium (Mg) in nitric oxide (NO) synthase blockade-induced hypertension. A group of albino rats received a NO synthase inhibitor, N(G)-nitro L-arginine-methyl ester (L-NAME, 60 mg/kg/d) in their drinking water for 21 d. L NAME intake caused a progressive rise in this group's resting mean arterial blood pressure compared to a control group (p < 0.01). There were no differences between the groups with regard to tissue and blood levels of Zn or Cu; however, Mg concentrations were significantly lower in the hypertensive rats' erythrocytes (20.2% reduction from control levels), cerebral cortex (17.0%), heart (9.1%), renal cortex (12%), renal medulla (16.7%), and in the tissues of the caval vein (23.7%), mesenteric artery (29.8%), renal artery (18.4%), and renal vein (22.1%). There were no significant Mg concentration changes in the hypertensive group's plasma, cerebellum, liver, duodenum, or aortal tissue. These findings suggest that Mg depletion may play a role in the blood pressure rise that occurs in the model of chronic NO synthase inhibition-induced hypertension. PMID- 11101043 TI - Differences in accumulation of elements in human cardiac valves. AB - To elucidate changes of human cardiac valves with aging, the authors determined age-related changes of element contents in the four human cardiac valves by inductively coupled plasma-atomic emission spectrometry and attempted to examine the relationships in the element contents among the four cardiac valves. The subjects consisted of 10 men and 15 women, ranging in age from 65 to 102 yr. The accumulation of calcium and phosphorus was the highest in the aortic valve, and decreased in the order mitral, pulmonary, and tricuspid valves. The contents of calcium and phosphorus in the aortic valves corresponded to about 12 and 19 times the amounts of those in the tricuspid valves, in which the contents were very low. The contents of calcium and phosphorus in the aortic valves were about 2.5 fold the amounts of those in the mitral valves. An examination was attempted to determine whether or not there were relationships in element contents among the four cardiac valves. As for the aortic and mitral valves, there were no relationships in the contents of calcium and phosphorus between them, but there were relationships in the contents of sulfur and magnesium between them. Three out of 24 cases contained high contents of calcium and phosphorus in both the mitral and aortic valves, whereas 16 out of 24 cases contained high contents of calcium and phosphorus in the aortic valves alone, without the high contents in the mitral valves. Likewise, there were no relationships in the element contents, such as calcium, phosphorus, sulfur, and magnesium, between the mitral and pulmonary valves or between the mitral and tricuspid valves. It is suggested that the accumulation of calcium and phosphorus in the cardiac valve occurs independent of the other cardiac valves. PMID- 11101044 TI - Age-related variations of elements as compared among optic, radial, and sciatic nerves. AB - To elucidate changes of peripheral nerves with aging, the authors studied age related changes of element contents in the optic, radial, and sciatic nerves by inductively coupled plasma-atomic emission spectrometry. The subjects consisted of seven men and seven women, ranging in age from 61 to 97 yr. The contents of phosphorus and sulfur remained constant through ages 61 to 97 yr in three nerves, the optic, radial, and sciatic nerves. It was found that there were age-related differences in calcium content among the optic, radial, and sciatic nerves: The calcium content of the optic nerve increased progressively with aging; in the radial nerve, it was hardly changed with aging; in contrast, the calcium content of the sciatic nerve decreased gradually with aging. In addition, it was found that in the radial nerve there were moderate correlations between age and zinc or sodium content, whereas significant correlations between age and the content of silicon or iron were found in the sciatic nerve. Furthermore, there was a correlation between the silicon and iron contents in the sciatic nerves. PMID- 11101045 TI - Age-dependent changes of elements in human trachea. AB - To elucidate compositional changes of human trachea by aging, element contents in tracheae were determined by inductively coupled plasma-atomic emission spectrometry. The subjects consisted of seven men and seven women, ranging in age from 61 to 97 yr. The sulfur content of the tracheae decreased gradually with aging. In regard to calcium and phosphorus, both the contents increased to about threefold amounts in their seventies compared with those in their sixties, and decreased thereafter. The contents of calcium and phosphorus were the highest in their seventies. Therefore, it is likely that surplus calcium released from bones is deposited temporally in the trachea, and the deposits are released from the trachea at older age. Based on our results of human cartilages, there are two types in regard to calcium accumulation: The first type is that calcium accumulation occurs progressively with aging; the second one is that calcium accumulation becomes the highest in the seventies or eighties, and decreases thereafter. Therefore, the trachea belongs to the second type. Furthermore, the magnesium content remained constant through the age range. PMID- 11101046 TI - Elements in the hair of workers at a workshop, foundry, and match factory. AB - The levels of seven elements determined in the hair were compared between male controls and industrial workers from Madras (South India). Particularly, the concentrations of Cd, Cr, Cu, Mn, and Ni in the foundry workers and Cu, Mn, and Ni in the workers of a workshop and match factory were observed to be higher than that of controls working in the office. In addition, the occupation, age, and period of service of workers had an influence, but diet, smoking habits, income of family, and hair color had no influence on the element levels in the hair. PMID- 11101047 TI - Possible contaminant origins of the red cosmetics decorating ancient burial sites in Japan. AB - Marker elements were estimated from the red cosmetics collected from different ancient burials and mine ruins in three separate districts of Japan. Element levels were displayed in reference to the relative amount to sulfur (RA/S), by which the cosmetics were divided into five types: I--a low Hg/S with a low Fe/S; II--both moderate Hg/S and Fe/S; III--a moderate Hg/S with a high Fe/S; III 2--a high Hg/S with a moderate Fe/S; IV--a high Hg/S with a high Fe/S. The cosmetics can be further characterized by referring to other contaminants such as Zn, Cu, and Mn. These combined analyses with contaminant metals were capable of characterizing the origins of the cosmetics; it is useful to compare them to each other. The cosmetics were identified as being due to several groups of contaminants from ancient mines in Japan, and also with this system analysis of the markers it is possible to identify them from neighboring countries. PMID- 11101048 TI - Adverse developmental and reproductive effects of copper deficiency in Xenopus laevis. AB - The effect of copper (Cu) deficiency on the reproduction and development in Xenopus laevis was evaluated, culminating in the development of a defined concentration-response relationship. Separate groups of four adult frog pairs were fed one of three diets for 28 d: (1) low-copper (-Cu); (2) copper supplemented (+Cu); and (3) ASTM standard beef liver and lung (BLL). Embryos collected from frogs administered the -Cu diet had markedly decreased egg masses and viability rates and an increased rate of necrosis when compared to the other dietary treatments. Malformations in -Cu larvae included maldevelopment of the heart, eye, craniofacial region, brain, and notochord. Larvae from adults administered the -Cu diet showed delayed abnormal hindlimb development, characterized as selective reductive deficiencies distal to the femur, with poor cartilaginous development. A U-shaped dose-response curve characteristic of nutritional essentiality was developed for Cu. Overall, these studies indicated that embryos produced from frogs administered a -Cu diet are substantially less viable than embryos from frogs administered a +Cu or copper-adequate (BLL) diet. PMID- 11101049 TI - Chronic boron or copper deficiency induces limb teratogenesis in Xenopus. AB - Sets of adult male and female Xenopus laevis were administered a boron-deficient (-B) diet under low-boron culture conditions, a boron-supplemented (+B) diet under ambient boron culture conditions, a copper-deficient (-Cu) diet under low copper culture conditions, or a copper-supplemented (+Cu) diet under ambient copper culture conditions, for 120 d. Adults from each group were' subsequently bred, and the progeny were cultured and bred. Results from these studies indicated that although pronounced effects on adult reproduction and early embryo larval development were noted in the -B F1 generation, no effects on limb development were observed. No significant effects on reproduction, early embryogenesis, or limb development were noted in the +B group, irrespective of generation. Highly specific forelimb and hindlimb defects, including axial flexures resulting in crossed limbs and reduction deficits, were observed in -B F2 larvae, but not in the +B F2 larvae. As was noted in the boron-deficiency studies, significant effects on reproduction and early embryo development were observed in the -Cu F1 generation, but not in the +Cu F, generation. Unlike the effects associated with boron deficiency, maldevelopment of the hindlimbs (32 responders, n = 40) was found in the F1 generation. PMID- 11101050 TI - Premature lesions of the carotid vessels in HIV-1-infected patients treated with protease inhibitors. AB - OBJECTIVES: To evaluate the presence of premature atherosclerotic lesions of epiaortic vessels in HIV-1-infected protease inhibitor-(PI) treated patients compared with PI-naive patients and healthy individuals. DESIGN: One-hundred and two HIV-1-positive patients, including 55 treated with PI for at least 12 months and 47 either naive or treated with PI-sparing regimens, were subjected to epiaortic vessel ultrasonography. These data were compared with those obtained from 104 healthy individuals. METHODS: Intima characteristics, pulsation and resistance indexes, and minimal, peak and mean speed were evaluated using a colour power doppler. Atherosclerotic plaques were described. Independent risk factors and values for glycaemia, cholesterolaemia and triglyceridaemia were considered. Statistical analysis included the chi-square test, Mantel-Haenszel test, odds ratio and logistic regression analysis. RESULTS: Of the PI-treated patients, 29 out of 55 (52.7%) presented acquired lesions of the vascular wall at ultrasonography, whereas similar lesions were found in seven out of 47 (14.9%) PI naive patients. Of the 104 healthy individuals, seven cases (6.7%) of intimal medial thickness were noted. A slightly significant correlation was found between carotid lesions and age, male sex and hypercholesterolaemia, whereas cigarette smoking, hypertriglyceridaemia and Centers for Disease Control and Prevention stage significantly increased the risk of vascular lesions (P= 0.022, P= 0.017 and P= 0.079 respectively). However, the highest significance regarded use of PI (P= 0.011). These results were confirmed by logistic regression analysis. CONCLUSIONS: These data demonstrate a higher than expected prevalence of premature carotid lesions in the PI-treated compared with PI-naive patients. If confirmed, a periodic ultrasonographic study of the vascular wall should be included in the follow-up of HIV infected patients. PMID- 11101051 TI - Didanosine once daily: potential for expanded use. AB - Factors affecting patient adherence to therapy, such as frequent daily dosing and complex dosing schedules, are widely understood to be key obstacles to the durability of effective anti-HIV therapy. Didanosine, a nucleoside analogue reverse transcriptase inhibitor (NRTI) that is a core component of combination antiretroviral regimens, is currently indicated for twice-daily dosing. However, the active metabolite of didanosine (2',3'-dideoxyadenosine-5'-triphosphate) has a long intracellular half-life that supports the use of didanosine in a more patient-friendly, once-daily dosing schedule. Clinical studies in which didanosine was administered either once or twice daily, as monotherapy or in combination with another NRTI, have demonstrated the equivalence of both dosing schedules, with respect to safety and tolerability, virologic and immunologic endpoints, and short-term clinical effects (e.g., weight gain). Preliminary results from recent studies support the clinical efficacy and utility of once daily didanosine in combination antiretroviral regimens that provide maximal drug exposure, while allowing for once- or twice-daily dosing of all component drugs. PMID- 11101052 TI - Correlation of virus load and soluble L-selectin, a marker of immune activation, in pediatric HIV-1 infection. AB - OBJECTIVE: HIV infections in children are characterized by high viral load and, in some perinatally infected newborns, delayed appearance of viral markers. Both phenomena may be related to different levels of immune activation affecting viral replication. This study was designed to investigate the relationship between immune activation and viral replication in pediatric HIV infection, and the role of pre-existent immune activation in facilitating HIV transmission to the fetus/newborn. DESIGN: Plasma levels of soluble L-selectin (s-LS), an immune activation marker, were determined in 100 infants with perinatally transmitted HIV infection, compared with 106 age-matched HIV-exposed uninfected controls. Included in the analysis were samples from 31 HIV-infected (10 PCR+ and 21 PCR-) and 35 uninfected newborns aged < 2 days. METHODS: To determine s-LS levels, a solid phase ELISA was performed on plasma samples of patients and controls. RESULTS: s-LS levels in uninfected children were higher than those in normal adults. HIV-infected patients had more rapidly increasing values in the first 6 months of life compared with uninfected infants. Plasma s-LS levels correlated with HIV viral loads (r, 0.50). Among newborns in the first 2 days of life, s-LS levels were lowest in those with negative PCR tests, compared with PCR-positive or uninfected infants. CONCLUSIONS: These results suggest that higher immune activation in children contributes to higher viral loads, and that the level of pre-existent immune activation may have a role in determining which infants have detectable virus in peripheral blood at birth. PMID- 11101053 TI - The effect of treatment of schistosomiasis on blood plasma HIV-1 RNA concentration in coinfected individuals. AB - OBJECTIVE: To determine whether drug treatment of Schistosomiasis mansoni infection leads to a reduction in plasma HIV-1 RNA concentration in coinfected individuals. METHODS: Stool and plasma samples were obtained prospectively from a cohort of HIV-infected persons (n = 30) in Kisumu, Kenya, before and after treatment of schistosomiasis with praziquantel (mean follow-up, 5.6 months; range 1-15 months). Schistosomal circulating cathodic antigen (CCA) concentrations in plasma were determined by ELISA and fecal egg counts were determined by microscopy. HIV-1 RNA concentrations were measured in pre- and post-treatment plasma samples obtained from the patients whose stool samples remained free of schistosomal eggs for the great majority of the follow-up period. RESULTS: Comparison of pretreatment and follow-up samples revealed that mean +/- SD fecal egg burden was reduced by 96.7% (481.5+/-803.5 versus 16.1+/-24.4 eggs/g feces) and mean plasma CCA concentration decreased by 90.1% (3.22+/-3.26 versus 0.32+/ 0.38 microg/ml). In contrast, mean plasma HIV-1 load increased from 3.60+/-0.90 to 3.93+/-0.95 log10 RNA copies/ml (P< 0.001). Although no correlation was found between changes in HIV-1 load and changes in schistosomal burden, there was a significant correlation between changes in plasma HIV load and the time interval between pretreatment and follow-up samples (r = 0.41; P = 0.027). CONCLUSIONS: Treatment of schistosomiasis was not associated with a reduction in plasma HIV-1 load. This study does not, however, exclude the possibility of an adverse effect of helminthic infections on HIV-1 pathogenesis. PMID- 11101054 TI - Cross-protection in NYVAC-HIV-1-immunized/HIV-2-challenged but not in NYVAC-HIV-2 immunized/SHIV-challenged rhesus macaques. AB - OBJECTIVES: Immunization with attenuated poxvirus-HIV-1 recombinants followed by protein boosting had protected four of eight rhesus macaques from HIV-2SBL6669 challenge. The present study was designed to confirm this result and to conduct the reciprocal cross-protection experiment. METHODS: Twenty-four macaques were primed with NYVAC (a genetically attenuated Copenhagen vaccinia strain) recombinants with HIV-1 and HIV-2 env and gag-pol or NYVAC vector alone and boosted with homologous, oligomeric gp160 proteins or adjuvant only. Binding and neutralizing antibodies, cytotoxic T-lymphocytes (CTL) and CD8 T cell antiviral activity (CD8AA) were evaluated. One half of each immunization and control group were intravenously challenged with SHIV(HXB2) the other half was challenged with HIV-2SBL6669,. Protective outcome was assessed by monitoring virus isolation, proviral DNA and plasma viral RNA. RESULTS: Both immunization groups developed homologous binding antibodies; however, homologous neutralizing antibodies were only observed in NYVAC-HIV-2-immunized macaques. While no cross-reactive neutralizing antibodies were detected, both immunization groups displayed cross reactive CTL. Significant CD8AA was observed for only one NYVAC-HIV-2-immunized macaque. Virological assessments verified that both NYVAC-HIV-1 and NYVAC-HIV-2 immunization significantly reduced viral burdens and partially protected against HIV-2 challenge, although cross-protection was not at the level that had been previously reported. Humoral antibody and/or CTL and CD8AA were associated with protection against homologous HIV-2 challenge, while cellular immune responses seemed more important for cross-protection. No significant protection was observed in the SHIV-challenged macaques, although NYVAC-HIV-1 immunization resulted in significantly lower viral burdens compared with controls. CONCLUSIONS: Further delineation of cross-reactive mechanisms may aid in the development of a broadly protective vaccine. PMID- 11101055 TI - Targeting cell-free HIV and virally-infected cells with anti-HLA-DR immunoliposomes containing amphotericin B. AB - OBJECTIVE: To evaluate the ability of liposomes bearing anti-HLA-DR Fab' fragments (immunoliposomes) and containing amphotericin B (AmB) to target and neutralize cell-free HIV-1 particles and virally-infected cells. METHODS: The effect of AmB on the attachment and fusion of HIV-1(NL4-3) to Jurkat E6.1 cells has been evaluated using a p24 enzymatic assay. The ability of AmB to inhibit HIV 1-based luciferase reporter viruses pseudotyped with HXB2, AML-V and VSV-G envelopes has been evaluated in Jurkat E6.1 cells. The efficacy of free and immunoliposomal AmB to inhibit cell-free HIV, that have incorporated or not HLA DR molecules, has been evaluated in HLA-DR/negative (NEG) 1G5 T cells and HLA DR/positive (POS) Mono Mac 1 cells. RESULTS: AmB inhibited HIV infectivity independently of the nature of viral envelope proteins. Pretreatment of HIV with AmB had no major effect on viral attachment and fusion process to Jurkat E6.1 cells. Immunoliposomal AmB (0.5 microg/ml) led to a 77% inhibition of replication of HLA-DR/POS HIV-1 with no cell toxicity, whereas free AmB had no significant antiviral activity at this concentration. A complete inhibition of viral replication was observed following incubation of viruses with immunoliposomal AmB (2.5 microg/ml). Anti-HLA-DR immunoliposomes containing AmB had no effect on the infectivity of HLA-DR/NEG HIV-1 particles in HLA-DR/NEG T lymphoid cells but completely inhibited replication of viruses in an HLA-DR/POS monocytic cell line. CONCLUSION: The incorporation of neutralizing agents in anti-HLA-DR immunoliposomes could represent a novel therapeutic strategy to specifically target cell-free HIV particles and virally-infected cells to treat HIV infection more efficiently. PMID- 11101056 TI - Ritonavir increases the level of active ADD-1/SREBP-1 protein during adipogenesis. AB - OBJECTIVE: A novel lipodystrophy syndrome characterized by truncal adiposity, peripheral fat atrophy, type 2 diabetes mellitus, and dyslipidemia occurs in HIV infected individuals, and may be aggravated by HIV-1 protease inhibitors. The increase in truncal fat could be due to enhanced preadipocyte differentiation. Using the 3T3-L1 preadipocyte model, we reported that ritonavir enhances adipocyte differentiation in culture. The goal of this study was to characterize the molecular mechanism of ritonavir on preadipocyte differentiation. DESIGNS AND METHODS: Time course studies of 3T3-L1 preadipocytes placed in standard differentiation medium (insulin, dexamethasone, and isobutylmethylxanthine) were performed. Glycerol phosphate dehydrogenase (GPDH) was assayed enzymatically, and triacylglycerol (TG) mass was quantified. The adipogenic transcription factors adipocyte determination and differentiation-dependent factor 1 (ADD-1)/sterol regulatory element binding protein 1 (SREBP-1), CCAAT/enhancer-binding protein alpha (CEBPalpha), and peroxisome proliferator activated receptor-gamma (PPARgamma), were measured by Western analysis. RESULTS: Ritonavir (10 microg/ml) enhanced 3T3-L1 preadipocyte differentiation (30% increase in TG mass; 50% increase in GPDH activity), and transiently raised levels of the 68 kDa active mature form of ADD-1/SREBP-1 during adipogenesis by threefold, compared with standard differentiation. In contrast, ritonavir attenuated the differentiation induced increase in CEBPalpha and PPARgamma. CONCLUSIONS: Our data suggest that ritonavir enhances 3T3-L1 adipogenesis by increasing the level of active mature ADD-1/SREBP-1. This effect may be due to reduced proteolysis of ADD-1/SREBP-1, as ritonavir inhibits an N-acetyl-leucyl-leucyl-norleucinal (ALLN)-sensitive proteosomal degradation pathway in lymphocytes, and ALLN itself inhibits the breakdown of mature ADD-1/SREBP-1. As mature ADD-1/SREBP-1 regulates several lipogenic enzymes, higher levels may explain the effect of ritonavir on TG accumulation and GPDH activity. Studying ADD-1/SREBP-1 may lead to better understanding and prevention of the lipodystrophy syndrome. PMID- 11101057 TI - Cell-mediated immune responses to autologous virus in HIV-1-seropositive individuals after treatment with an HIV-1 immunogen. AB - OBJECTIVE: We hypothesized that cell mediated immune responses to an HIV-1 immunogen (whole-killed, gp120-depleted HIV-1 in IFA, REMUNE) would include those to autologous virus. METHODS: Five chronically HIV-1 infected individuals were examined for HIV-specific immune responses to their own virus (autologous viral antigen) after treatment with an HIV-1 immunogen. RESULTS: Subjects had low proliferative responses to HIV and p24 antigens prior to immunization and mounted strong lymphocyte proliferative responses to the immunizing HIV-1 virus, native p24, and autologous viral antigen post immunization. Similarly, subjects produced low amounts of interferon-gamma in response to HIV and p24 antigens prior to immunization and increased their interferon-gamma production in response to HIV 1, native p24, and to autologous antigen post-immunization. Furthermore, beta chemokine responses measured as migratory inhibitory protein-1beta production were low at baseline in response to HIV-1 and native p24 antigens and were enhanced post immunization to HIV-1, native p24, and autologous antigen. CONCLUSIONS: In this study HIV-specific immune responses to autologous virus were observed after treatment with an HIV-specific immunogen. PMID- 11101058 TI - Construction of infectious SIV/HIV-2 chimeras. AB - OBJECTIVE: To construct SIV/HIV-2 chimeras (SHIV) that replicate in vivo. These would be valuable tools to elucidate the mechanism by which HIV-2 can bypass protection conferred by live attenuated SIV vaccines. METHOD: Novel SHIV were constructed to express either the vpx, vpr, tat, rev and env genes (SHIV-2isy env) or the gag and pol genes (SHIV-2isy gag/pol) of the infectious molecular clone HIV-2isy in an SIVmac backbone. The replication of SHIV-2isy env and SHIV 2isy gag/pol were evaluated on selected cell lines and peripheral blood mononuclear cells (PBMC) in vitro. In addition, their infectivity was assessed in vivo. RESULT: Virus stocks of SHIV-2isy env and SHIV-2isy gag/pol were prepared in vitro. For SHIV-2isy gag/pol both the 5' and 3' boundaries of the chimeric construct were critical for infectivity in vitro. The growth of each chimera on T cell lines in vitro mirrors that of the parental viruses donating the envelope gene. On PBMCs SHIV-2isy env replicated well on human and simian PBMC whereas SHIV-2isy gag/pol replicated to detectable levels on human PBMC only. In vivo, SHIV-2isy env virus was isolated from one of two cynomolgus macaques challenged intravenously, SHIV-2isy gag/pol was isolated from one of two cynomolgus macaques and both rhesus macaques challenged intravenously. CONCLUSION: This is the first report of SIV/HIV-2 chimeras that are infectious in macaques. Moreover, this is the first report of an infectious chimera in which both SIV gag and pol have been replaced with the equivalent regions of an HIV isolate. PMID- 11101059 TI - Comparison of twice-daily stavudine plus once- or twice-daily didanosine and nevirapine in early stages of HIV infection: the scan study. AB - OBJECTIVES: To evaluate the safety and effectiveness of once-daily didanosine and nevirapine plus twice-daily stavudine versus twice-daily administration of all three drugs. METHODS: This open-label, randomized, multicentre study enrolled 94 antiretroviral-naive patients with chronic HIV infection, CD4+ cell counts > 500 x 10(6) cells/l, and viral loads > 5000 copies/ml. Patients were treated with either 40 mg stavudine (twice daily) plus 400 mg didanosine (once daily) and 400 mg nevirapine (once daily) or 40 mg stavudine (twice daily) plus 200 mg didanosine (twice daily) and 200 mg nevirapine (twice daily). RESULTS: After 12 months, 68% of patients who received twice-daily didanosine and nevirapine had viral loads < 200 copies/ml in the intention-to-treat and 79% in the on-treatment analysis, respectively. The corresponding values for patients treated with didanosine and nevirapine, taken once-daily, were 73 and 85%. The percentages of patients in each group with viral loads < 5 copies/ml at 12 months were 40% (once daily ) and 45% (twice daily) for the intention-to-treat analysis. Five of 11 patients (45%) with plasma viral loads < 5 copies/ml at 12 months had detectable virus in tonsillar tissue. Genotypic resistance to nevirapine was noted in seven of the 14 patients with detectable viral load at month 12. Mean changes in CD4+ cell counts for patients treated with stavudine plus once- or twice-daily didanosine and nevirapine were 154 and 132 x 10(6) cells/l, respectively. Treatment was interrupted due to adverse events in seven patients (8%) (four who received once-daily didanosine and nevirapine and three treated with twice-daily doses). CONCLUSIONS: The combination of twice-daily stavudine plus once-daily didanosine and nevirapine was as safe and well tolerated as twice-daily administration of all three agents. Both regimens were equally effective in reducing viral loads and in increasing CD4+ cell counts. PMID- 11101060 TI - Competing drug-drug interactions among multidrug antiretroviral regimens used in the treatment of HIV- infected subjects: ACTG 884. AB - OBJECTIVE: To evaluate the steady state concentrations of saquinavir, ritonavir, nelfinavir, delavirdine, and adefovir in six different three- and four-drug combination regimens. DESIGN: Randomized, partially double-blinded, multicenter study in a population of indinavir-experienced subjects with virologic failure. The first seven subjects enrolled in each of the six treatment arms from 10 participating sites were entered into this pharmacokinetic evaluation. SETTING: Multicenter study of the AIDS Clinical Trials Group (ACTG). PATIENTS: HIV infected subjects. INTERVENTIONS: A 12-hour pharmacokinetic study was conducted after 2 weeks of drug administration. MAIN OUTCOME MEASURES: Area under the concentration-time curve with statistical comparisons to evaluate the effect of the second protease inhibitor and the effect of the non-protease inhibitors. RESULTS: There was no difference in saquinavir concentrations according to whether the second protease inhibitor was ritonavir or nelfinavir. Saquinavir concentrations in the groups receiving the combination of delavirdine plus adefovir dipivoxil were reduced by approximately 50% compared with those receiving delavirdine. Delavirdine concentrations were reduced by approximately 50%, in the delavirdine plus adefovir dipivoxil arms compared with the delavirdine arms. CONCLUSIONS: Saquinavir concentrations were significantly lower in the arms containing the combination of delavirdine and adefovir dipivoxil compared with the arms containing delavirdine. Delavirdine concentrations were significantly lower when coadministered with adefovir dipivoxil. These drug-drug interactions were not expected, the mechanism(s) is (are) not clear, and additional studies are warranted. This study illustrates the need to understand more fully the pharmacokinetic characteristics of complex combination antiretroviral regimens prior to use in patient management. PMID- 11101061 TI - A multicenter, randomized, double-blind, placebo-controlled trial of recombinant human interleukin-10 in HIV-infected subjects. AB - OBJECTIVE: To determine the effect of multiple subcutaneous doses of recombinant human interleukin (rhuIL)-10 on plasma HIV RNA levels and CD4 T-cell counts, and to evaluate its safety and tolerability in HIV-infected subjects. DESIGN: Prospective, randomized, double-blind, placebo-controlled, multicenter trial. SUBJECTS: Thirty-nine HIV-infected subjects with CD4 T-cell counts > 200 x 10(6)/l, plasma HIV RNA concentrations > or = 3.18 log10 copies/ml and on stable antiretroviral therapy were recruited from six centers. INTERVENTION: Subjects received (subcutaneously) rhuIL-10 1 microg/kg daily, 4 microg/kg daily, 8 microg/kg three times per week, placebo daily or placebo three times per week for 4 weeks. MAIN OUTCOME MEASURES: Prospectively defined outcomes included safety and tolerability, plasma HIV RNA levels and CD4 T-cell counts. Outcomes were assessed at baseline, weeks 1, 2, 3 and 4 during treatment and weeks 2 and 4 following completion of therapy. RESULTS: Baseline characteristics were similar in all groups. Compared to baseline, no significant change in plasma HIV RNA concentrations or CD4 T-cell counts was observed in any of the groups. RhuIL-10 was generally well tolerated. Two patients receiving rhuIL-10 4 microg/kg required discontinuation due to thrombocytopenia. One patient receiving rhuIL-10 4 microg/kg who had chronic hepatitis B and C infections discontinued drug because of elevated liver function tests. One patient receiving placebo discontinued study drug because of depression. CONCLUSION: The lack of a demonstrable virological benefit, as assessed by plasma viral load, with 4 weeks of rhuIL-10 does not support the development of this immune-based therapy for treatment of HIV infection. PMID- 11101062 TI - A randomized, controlled 24-week study of intermittent subcutaneous interleukin-2 in HIV-1 infected patients in Thailand. AB - OBJECTIVES: To assess the immunological and virological effects, safety profile and feasibility of subcutaneous interleukin-2 (scIL-2) therapy in an HIV-infected Thai population. DESIGN: Seventy-two patients with baseline CD4 cell count of > or = 350 x 10(6)/l and no history of opportunistic infection were randomized to receive antiretroviral therapy plus scIL-2 (scIL-2 group) or antiretroviral therapy alone (control group). scIL-2 was administered at one of three doses for at least 24 weeks. The main measure of treatment efficacy was change in CD4 cell count. RESULTS: The time-weighted mean change in CD4 cell count from baseline to week 24 was + 252 x 10(6)/l for the scIL-2 group compared with + 42 x 10(6)/l for the control group (P< 0.0001). Changes in plasma HIV RNA were not significantly different between the groups over the same time period: there was a 0.83 log10 copies/ml decrease for the scIL-2 group and a 0.70 log copies/ml decrease for the control group (P= 0.362). CONCLUSIONS: This study provides the most extensive experience of scIL-2 therapy in HIV-1 infected women and Asians, and demonstrates the immunological efficacy, tolerability and feasability of scIL-2 therapy in this population. Data from this study were instrumental in guiding the selection of the scIL-2 dosing regimen for ongoing phase III trials. PMID- 11101063 TI - HIV infection in Haiti: natural history and disease progression. AB - OBJECTIVE: A study was conducted to define the natural history and disease progression of HIV infection in a developing country. DESIGN: A prospective longitudinal cohort study. METHODS: Forty-two patients with documented dates of HIV seroconversion were followed in Port-au-Prince, Haiti. Patients were seen at 3 month intervals or when ill. Patients were treated for bacterial, mycobacterial, parasitic, and fungal infections, but antiretroviral therapy was not available. Patients were followed until death or until 1 January 2000; median follow-up was 66 months. RESULTS: By Kaplan-Meier analyses, the median time to symptomatic HIV disease (CDC category B or C) was 3.0 years [95% confidence interval (CI) 2.3-5.0 years]. The median time to AIDS (CDC category C) was 5.2 years (95% CI 4.7-6.5 years), and the median time to death was 7.4 years (95% CI 6.2-10.2 years). Community-acquired infections, including respiratory tract infections, acute diarrhea, and skin infections were common in the pre-AIDS period. AIDS-defining illnesses included tuberculosis, wasting syndrome, cryptosporidiosis, cyclosporiasis, candida esophagitis, toxoplasmosis, and cryptococcal meningitis. Rapid progression to death was associated with anemia at the time of seroconversion hazards ratio (HR) 4.1 (95% CI 1.1-15.0), age greater than 35 years at seroconversion HR 4.4 (95% CI 1.1-16.6), and lymphopenia at seroconversion HR 11.0 (95% CI 2.3-53.0). CONCLUSION: This report documents rapid disease progression from HIV seroconversion until death among patients living in a developing country. Interventions, including nutritional support and prophylaxis of common community-acquired infections during the pre-AIDS period may slow disease progression and prolong life for HIV-infected individuals in less-developed countries. PMID- 11101064 TI - Management of sexually transmitted diseases and HIV prevention in men at high risk: targeting clients and non-paying sexual partners of female sex workers in Benin. AB - OBJECTIVES: Male clients of female sex workers have rarely been specific targets for HIV/sexually transmitted diseases (STD) interventions in sub-Saharan Africa. We assessed the effectiveness of outreach methodology for contacting sexual partners of female sex workers for purposes of HIV/STD prevention in Cotonou, Benin. DESIGN AND METHODS: In collaboration with owners/managers, outreach personnel and female sex workers, 404 clients were recruited on-site at prostitution venues, and provided urine samples for leukocyte esterase dipstick (LED), STD and HIV testing before having sex with female sex workers. After having sex they underwent an interview and physical examination. No payment was made for study participation. Prostitution site personnel (n = 41) and boyfriends of female sex workers (n = 56) were also recruited. RESULTS: In all 68% of the clients approached agreed to participate. On-site LED testing and free STD treatment were important factors in participation. HIV-1 prevalence was several fold higher than in the general population in Cotonou, at 8.4, 12.2 and 16.1% in clients, personnel and boyfriends respectively, and was associated with increasing age and lack of condom use with female sex workers. Condom use rates by clients with female sex workers were non-negligible but sub-optimal, and low with regular partners. Approximately one-third of clients with regular partners also had other non-female sex worker sex partners. Boyfriends of female sex workers are of particular concern due to high numbers of partners, very low condom use rates and high HIV prevalence. CONCLUSIONS: Study findings indicate that male sex partners of female sex workers form a 'bridging population' for HIV/STD transmission both to female sex workers, as well as from female sex workers to the general population of women, particularly regular female partners. PMID- 11101065 TI - Risk factors for postnatal mother-child transmission of HIV-1. AB - OBJECTIVE: To identify factors affecting HIV-1 breastfeeding transmission. DESIGN: Longitudinal observational cohort study. METHODS: HIV-1 seropositive pregnant women and seronegative controls were enrolled at a maternity hospital in Nairobi. Women and their children were followed from birth, and data on HIV-1 transmission, breastfeeding, clinical illness, and growth were collected. Specimens for HIV-1 serology and/or polymerase chain reaction were obtained at birth, 2, 6, and 14 weeks, 6, 9, 12, and 18 months, and every 6 months thereafter. Children were classified as HIV-1 uninfected, perinatally, or postnatally infected. Potentially breastfeeding transmission related risk factors were compared between postnatally infected and uninfected children. RESULTS: Among children born to seropositive or seroconverting mothers, 317 were uninfected, 51 infected perinatally and 42 infected postnatally. Identified risk factors for postnatal transmission were maternal nipple lesions (OR = 2.3, CI 95% 1.1-5.0), mastitis (OR = 2.7, CI 95% 1.1-6.7), maternal CD4 cell count < 400 mm3 (OR = 4.4, CI 95% 1.9-9.9), maternal seroconversion while breastfeeding (OR = 6.0, CI 95% 1.8-19.8), infant oral thrush at < 6 months of age (OR = 2.8, CI 95% 1.3-6.2) and breastfeeding longer than 15 months (OR = 2.4, CI 95% 1.2-5.1). All factors, except maternal seroconversion due to its rarity, were independently associated with an increased postnatal transmission risk by multivariate logistic regression analysis. CONCLUSION: In addition perinatal antiretroviral therapies, public health strategies should address: (i) prevention of maternal nipple lesions, mastitis and infant thrush; (ii) reduction of breastfeeding duration by all HIV-1-infected mothers; (iii) absolute avoidance of breastfeeding by those at high risk, and (iv) prevention of HIV-1 transmission to breastfeeding mothers. PMID- 11101066 TI - The cost-effectiveness of elective Cesarean delivery for HIV-infected women with detectable HIV RNA during pregnancy. AB - OBJECTIVES: To determine the net health consequences, costs, and cost effectiveness of alternative delivery strategies for HIV-infected pregnant women with detectable HIV RNA in the USA. DESIGN: Cost-effectiveness analysis using a probabilistic decision model. METHODS: The model compared two strategies: elective Cesarean section and vaginal delivery. Data for HIV transmission rate, maternal death rate, health-related quality of life and costs were obtained from the literature, national databases, and a tertiary hospital's cost accounting system. Model outcomes included total lifetime costs, quality-adjusted life expectancy, maternal death rate, HIV transmission rate, and incremental cost effectiveness ratios. RESULTS: Elective Cesarean section resulted in a vertical HIV transmission rate of 34.9 per 1000 births compared with 62.3 per 1000 births for vaginal delivery. Elective Cesarean section was more effective (38.7 quality adjusted life years per mother and child pair) and less costly ($10600 per delivery) than trial of labor (38.2 combined quality adjusted life years at a cost of $14500 per delivery). However, elective Cesarean section increased maternal mortality by 2.4 deaths per 100000 deliveries. The results were consistent over a wide range of the variables, but were sensitive to the risk of HIV transmission with vaginal delivery and the relative risk of HIV transmission with elective Cesarean section. CONCLUSIONS: In pregnant HIV-infected women with detectable HIV RNA, elective Cesarean section would reduce total costs and increase overall quality-adjusted life expectancy for the mother-child pair, albeit at a slight loss of quality adjusted life expectancy to the mother. PMID- 11101067 TI - Challenges in the conduct of vaginal microbicide effectiveness trials in the developing world. AB - Conducting a phase III trial of a vaginal microbicide in a developing country poses several important and complex ethical challenges. As part of a process to bridge the gap between ethical theory and practice, we share our experiences in performing a phase III trial of Col 1492 (Advantage S) among female sex workers at four sites world-wide; Durban, Abidjan, Cotonou and Hat Yai. The ethical challenges included: (i) difficulties in obtaining informed consent. Participants were unable to grasp the concepts of a clinical trial for several weeks to months. In Cotonou, 30% of the women did not know the gel was tested for HIV prevention. Only 25% understood what a placebo was. In Durban, 70% of the women did not fully understand the study after 3 months; (ii) in sustaining the use of known HIV prevention strategies. Participants at the Durban site had difficulty in sustaining condom use due to financial and client preferences. Sex without condoms was worth more ($20) than sex with condoms ($10); (iii) in maintaining the confidentiality of the subject's HIV status. Novel approaches such as role plays and emphasis on other exclusion criteria were needed to maintain the confidentiality of women not included in the trial due to their HIV status; (iv) in providing care and support to the subjects who became infected with HIV during the trial. Women could only be offered routine sexually transmitted disease treatment and counselling. Anti-retrovirals were not offered. The successes and failures of the solutions attempted are described. PMID- 11101068 TI - Epidemiology of Pneumocystis carinii pneumonia in an era of effective prophylaxis: the relative contribution of non-adherence and drug failure. AB - OBJECTIVE: To determine the relative contribution of patient non-adherence, provider failure to prescribe prophylaxis, and drug failure to the continued occurrence of Pneumocystis carinii pneumonia (PCP), and to determine correlates of non-adherence. DESIGN: Retrospective case-control study. METHODS: Patients with confirmed or presumptive PCP from May 1995 to September 1997 who had at least 6 months of prior HIV care (cases) were compared to controls matched for initial CD4 cell count and date of initial HIV care. RESULTS: The incidence of PCP declined by 85% in the 28 months of the study. Of the 118 cases of PCP identified, 59 (50%) were in HIV care for > 6 months prior to PCP diagnosis. In a multivariate logistic regression model, risk factors for PCP among patients in HIV care were patient non-adherence [odds ratio (OR), 12.4; 95% confidence interval (CI), 6.4-23.5], use of prophylaxis other than trimethoprim sulfamethoxazole (OR, 27.0; 95% CI, 13.8-52.9), and absence of antiretroviral use (OR, 7.5; 95% CI, 4.5-12.5). Provider non-adherence occurred in one out of 59 cases (2%), and five out of 106 controls (5%). Of the patients who developed PCP on prophylaxis, 18 cases (30%) appeared due to drug failure; there were no cases of apparent drug failure among patients on trimethoprim-sulfamethoxazole. In multivariate analysis, non-adherence was more common among patients of non-white race, those with a history of injecting drug use, and those with active substance abuse or psychiatric illness. CONCLUSIONS: Patient non-adherence was the most common reason for the occurrence of PCP among patients in HIV care; provider non adherence was uncommon. Drug failure occurred only among patients on prophylaxis other than trimethoprim-sulfamethoxazole. PMID- 11101070 TI - Risk behaviour change and HIV infection among injection drug users in Montreal. AB - OBJECTIVE: To investigate the independent association between changes in risk behaviour and HIV seroconversion risk among Montreal injection drug users (IDU). DESIGN: A longitudinal study of risk behaviour change and the maintenance of low risk practices. At baseline and semi-annually, subjects were tested for HIV, and questionnaires on risk behaviour were completed. RESULTS: A total of 833 IDU were recruited from January 1992 to June 1998, and completed a minimum of three visits. Large fluctuations in risk behaviour were observed, and the risk of HIV infection appeared to be dependent upon the consistency of risk behaviour practised. IDU who consistently engaged in risky behaviour were at high risk of HIV infection. IDU who attempted to practise low-risk behaviour but experienced relapses to risky behaviour were also at considerable risk of infection. IDU who managed to maintain low-risk practices were at minimal risk. Using Cox regression analysis, the hazard ratio (HR) of HIV seroconversion among IDU who consistently and inconsistently shared needles with an HIV-positive partner was 8.17 (95% CI 3.59-18.59) and 2.63 (95% CI 1.33-5.17), respectively, relative to non-needle sharers. Corresponding HIV incidence rates were 30.42 per 100 person-years (py) among consistent sharers, 13.78 per 100 py among inconsistent sharers and 2.51 per 100 py among non-sharers. CONCLUSION: Although some HIV risk reduction was evident, behaviour change seems to be effective only in IDU who adopt and maintain low-risk practices. Additional strategies may be needed to assist IDU in the maintenance of low-risk practices. PMID- 11101069 TI - Quality of life outcomes of combination zidovudine- didanosine-nevirapine and zidovudine-didanosine for antiretroviral-naive advanced HIV-infected patients. AB - OBJECTIVES: To evaluate the quality of life outcomes in antiretroviral-naive patients randomized to zidovudine plus didanosine versus zidovudine plus didanosine plus nevirapine for treatment of advanced HIV disease (the Istituto Superiore di Sanita 047 trial). DESIGN: A 48-week randomized, double-blind trial. METHODS: Sixty patients were enrolled and evaluated over 24 weeks. Quality of life was assessed using a modified version of the Medical Outcomes Study-HIV Health Survey. For analysis, we calculated two summary scores reflecting the physical (PHS) and the mental (MHS) components of health. RESULTS: Although the three-drug combination was superior at inducing immunologic and virologic responses, the two-drug regimen was superior for both PHS and MHS, especially at week 8 where differences were both statistically and clinically significant (5.8 and 9.2 points, respectively, P< 0.02 for both). Quality of life changes paralleled trends in body weight and Karnofsky performance status score. CONCLUSION: Although a three-drug antiretroviral therapy regimen was superior in terms of short term virologic/immunologic response, the two-drug regimen was better in terms of quality of life. In general, triple therapy remains the most effective treatment option. However, quality of life assessments can yield results that may be discordant with and complementary to those obtained using conventional endpoints. Comparative trials should collect a comprehensive range of outcome measures, including patient-reported quality of life, in order to provide clinicians and patients with additional information that may influence treatment decisions. PMID- 11101071 TI - Complementary hypothesis concerning the community sexually transmitted disease mass treatment puzzle in Rakai, Uganda. AB - OBJECTIVES: To study the dynamics of a mass treatment programme for sexually transmitted diseases (STD) on prevalence of STD and HIV incidence in order to help explain the results of the STD mass treatment community trial in Rakai, Uganda. METHODS: The analysis is based on simulations of STD mass treatment interventions using a deterministic model describing the course of STD and HIV transmission over time and incorporating demographic, biological and behavioural parameters. The mass intervention modelled mimics that used in the Rakai community trial. RESULTS: Mass treatment decreases STD prevalence to a very low level compared with baseline but is unsuccessful at eradicating the infection. STD prevalences return to baseline fairly rapidly after each round of mass treatment. Under different realistic scenarios, the fraction of HIV cases prevented by STD mass treatment assuming uniform 80% coverage of high- and low risk groups, over the 20-month period following the first round of treatment, was greater than 35%. If, however, differential coverage is assumed, for example that while the total coverage is still 80%, only 40 or 25% of those at high risk are treated, the HIV preventable fraction is reduced, to 19 and 15% respectively (undetectable given the statistical power of the study). The tremendous impact of differential coverage can also be observed even in the early stage of the HIV epidemic. CONCLUSIONS: In the Rakai trial, mass treatment may have had an effect, although transient, on all STD prevalences, which could have had positive repercussions for HIV incidence. This modelling exercise suggests that although an 80% coverage appears high, the differential coverage of low- and high-risk populations may seriously impair our ability to test the STD-HIV interaction hypothesis. PMID- 11101072 TI - Impact of highly active antiretroviral therapy on onset of Mycobacterium avium complex infection and cytomegalovirus disease in patients with AIDS. AB - OBJECTIVES: To assess the impact of highly active antiretroviral therapy (HAART) on the onset of first disseminated Mycobacterium avium complex (MAC) infection and first cytomegalovirus (CMV) disease episode in HIV-infected at-risk patients. METHODS: The incidence of the two infections occurring in at-risk patients was calculated for two periods (January 1995-June 1996 and July 1996-December 1997) using the database of the HIV-infected patients followed in the Infectious Diseases Department at the Pitie-Salpetriere Hospital in Paris. HAART was progressively introduced in late June 1996 in France. RESULTS: A total of 91 first disseminated MAC infections and 124 first CMV disease episodes were recorded. The incidence of first disseminated MAC infections fell from 13.4 per 100 person-years in the first 18-month period to 2.6 per 100 person-years in the second 18-month period. Similarly, the incidence of first CMV disease episodes fell from 20.9 to 3.5 per 100 person-years. Fourteen patients on HAART developed a first MAC infection, 12 (85.7%) within 2 months of starting HAART. Nineteen patients on HAART had a first CMV disease episode, 10 (52.6%) within 2 months of starting HAART. CONCLUSIONS: HAART led to a five-fold decrease in the incidence of first disseminated MAC infections and a six-fold decrease in first CMV disease episodes, although patients remain vulnerable to both diseases for approximately 2 months after starting HAART. PMID- 11101073 TI - What a test for recent infection might reveal about HIV incidence in England and Wales. AB - BACKGROUND: A laboratory method has been developed that detects recent HIV infection and allows incidence to be estimated by testing single stored antibody positive specimens. A theoretical exploration of the method's surveillance utility was carried out. METHODS: Using various data sources, HIV incidence rates were postulated. The confidence intervals (CI) for these postulated incidences were calculated using the expected number of recent infections for each postulated incidence, the actual number tested for HIV, and the known number of HIV-1 positives. A test for trend was used to determine when an important change in incidence could be recognized. RESULTS: If the incidence was 5% per annum (p.a.) in homosexual/bisexual men attending sexually transmitted diseases (STD) clinics in London, 64 recent infections would be expected in the 392 HIV seropositive specimens and, if observed, would result in a 95% CI of 3.1-7.9% p.a. for the incidence rate. An incidence of 1% p.a. in pregnant women would be most unlikely as this would require detection of 193 recent infections, 26 more than the total 167 HIV-seropositive specimens found in 1997. In African women attending STD clinics in London, 30% of prevalent infections would be classified as recent if the incidence was 5% p.a. Further, if the incidence in homosexual/bisexual men were to fall by 50% over 3 years, a decrease of this magnitude would be recognized as significant within 2 years. CONCLUSIONS: The detuned assay will increase the information from HIV serosurveys even where prevalence and incidence are relatively low. Existing surveillance systems should be redesigned to take full advantage of the method. PMID- 11101074 TI - Cervicovaginal anti-HIV antibodies in HIV-seronegative female sex workers in Abidjan, Cote d'Ivoire. AB - OBJECTIVE: To detect anti-HIV antibodies in cervicovaginal secretions of HIV seronegative female sex workers and to evaluate whether the presence of these antibodies is associated with increased sexual exposure. METHODS: A cross sectional study was carried out at a confidential clinic for female sex workers in Abidjan, Cote d'Ivoire. The participants were 342 HIV-seronegative female sex workers in whom a cervicovaginal lavage was collected. The main outcome measures were the detection of antibodies to HIV-1 in cervicovaginal lavages using an in house and a commercial (Seradyn Sentinel; Calypte Biomedical Corporation, Berkeley, California, USA) enzyme immunoassay; the detection of semen in cervicovaginal lavages; and the assessment of epidemiological and biological markers of sexual exposure to HIV. RESULTS: Cervicovaginal anti-HIV antibodies were detected in 7.3 and 29.8% of women using in-house enzyme-linked immunosorbent assay (ELISA) and Seradyn Sentinel respectively. All cervicovaginal secretions found to be positive by in-house ELISA were also positive by Seradyn Sentinel. In a minority of women, ranging from 2.9% by in-house ELISA to 12.3% by Seradyn Sentinel, the anti-HIV antibodies were present in vaginal fluids that did not contain semen. Sexual exposure to HIV was similar in women with anti-HIV antibodies in their semen-free cervicovaginal secretions compared with women without anti-HIV antibodies in their cervicovaginal secretions. CONCLUSIONS: Cervicovaginal HIV-specific antibodies were detected in a minority of sexually exposed HIV-seronegative female sex workers in Abidjan. The lack of association between increased sexual exposure to HIV and presence of cervicovaginal HIV specific antibodies suggests that the production of genital HIV-specific antibodies in exposed seronegative women depends on the ability of individual women to mount specific mucosal immunity to HIV antigens, the determinants of which are currently unknown. PMID- 11101075 TI - Fighting global AIDS: the value of cost-effectiveness analysis. PMID- 11101076 TI - Why does Pneumocystis carinii pneumonia still occur? PMID- 11101077 TI - An outbreak of the circulating recombinant form AECM240 HIV-1 in the Finnish injection drug user population. PMID- 11101078 TI - Discontinuation of secondary prophylaxis for cryptococcal meningitis in HIV infected patients responding to highly active antiretroviral therapy. PMID- 11101079 TI - Lack of significant viral load alterations of hepatitis B virus and hepatitis C virus during treatment with IL-2 and highly active antiretroviral therapy. PMID- 11101080 TI - Determination of HIV-1 circular DNA as a surrogate marker for residual virus replication in patients with undetectable virus loads. PMID- 11101081 TI - Once-daily indinavir plus ritonavir: preliminary results of the PIPO study. PMID- 11101082 TI - HIV-1 group N among HIV-1-seropositive individuals in Cameroon. PMID- 11101083 TI - Rapid quantification of SIV-specific CD8 T cell responses with recombinant vaccinia virus ELISPOT or cytokine flow cytometry. PMID- 11101084 TI - Increased levels of anti-CCR5 antibodies in sera from individuals immunized with allogeneic lymphocytes. PMID- 11101085 TI - Thresholds of CD4 cells for initiating trimethoprim-sulfamethoxazole prophylaxis in west Africa. PMID- 11101086 TI - Response to Eberly et al., Kaposi's sarcoma and central nervous system disease: a real association or an artifact of the control group? PMID- 11101087 TI - Infection by zidovudine-resistant HIV-1 compromises the virological response to stavudine in a drug-naive patient. PMID- 11101088 TI - Biological pathways to bladder carcinogenesis. AB - The transformation of normal urothelium into histologically different neoplastic states has been well characterized, and current clinical management of both superficial and invasive bladder cancer has benefited from recent scientific discoveries. The ability to define novel treatment strategies including surgical, chemotherapeutic, and gene therapies relies on our understanding of the basic mechanisms underlying human bladder carcinogenesis. Many in vitro culture systems and in vivo animal models have been developed over recent years, which have been used to define key molecular events that are associated with the development of bladder cancer. The biological pathways through which normal urothelium may progress to superficial or invasive disease will be discussed in the framework of recent advances in the field. PMID- 11101089 TI - Endoscopic fluorescence diagnosis and laser treatment of transitional cell carcinoma of the bladder. AB - Recurrent bladder cancer is due to tumor cell implantation, incomplete resection, and multicentric neoplastic changes throughout the bladder. The possibilities of 5-Aminolevulinic acid-induced fluorescence endoscopy (AFE), a highly sensitive method in detecting bladder cancer and laser energy as treatment to lower the recurrence rate in bladder cancer, are evaluated. After intravesical administration of AFE Protoporphyrin IX, a tumor-selective manner is excited by a xenon-arc lamp (wavelength 400 to 410 nm) to emit red fluorescence. Suspicious lesions can be detected by their red fluorescence and are electroresected or treated with laser energy. Complete resection or destruction of all tumors in the bladder is crucial to prevent recurrent and invasive growth of transitional cell carcinoma. AFE detects malignant lesions in the bladder with a sensitivity of 98% and Cis in 100%, respectively. Laser treatment of superficial bladder cancer lowers the local recurrence rate and reduces the risk of viable tumor cell implantation. PMID- 11101090 TI - Follow-up of patients with noninvasive and superficially invasive bladder cancer. AB - Low-grade carcinomas (pTaG1) comprise 50% of all stage pTa-pT1 carcinomas and have an almost benign course of disease. Follow-up policies may be changed so that patients with a single tumor at diagnosis and a negative cystoscopy at 3 months should be examined 9 months later. Check cystoscopies may be performed with flexible instruments and a considerable number of the recurrences could be managed with fulguration under urethral anesthesia only. Because low-grade carcinomas are so common, the seriousness of the other tumors in stages pTa-pT1 is not fully appreciated. Patients with high-grade carcinoma (pTaG2-G3, pT1G2-G3) have at least a 70% risk for recurrence and a 20% risk for stage progression. The course of disease is more unpredictable than for patients with low-grade carcinoma, and there are at present no firm data that support a change in follow up routines. Routine follow-up urographies are not necessary. PMID- 11101091 TI - Intravesical therapy for superficial bladder cancer. AB - Intravesical therapy is a classic approach to the treatment of superficial bladder cancer. Bacillus Calmette-Guerin (BCG) has assumed the role of the most commonly administered agent, and clearly the most effective form of therapy for carcinoma in situ. Recent data quantify the advantage of "booster" treatments over a 3-year interval but suggest that current regimens are not easily tolerated by most patients. Initial studies suggest a role for intravesical interferon in potentiating the effect of BCG. Classic intravesical chemotherapy can decrease tumor recurrence by 14% to 17%, but no data exist to demonstrate any positive effect on decreasing tumor progression. Recent studies suggest a growing rationale for the intravesical administration of a single dose of a chemotherapeutic agent to decrease early tumor recurrence. PMID- 11101092 TI - Indications for early cystectomy. AB - Treatment goals for superficial bladder cancer are two-fold: (1) reducing tumor recurrence and the subsequent need for additional therapies (cystoscopy, transurethral resection, intravesical therapy) and the morbidity associated with these treatments; and (2) preventing tumor progression and the subsequent need for more aggressive therapy. The administration of intravesical therapy has become an important component in these treatment goals; however, there remains a group of patients with superficial bladder cancer that are at risk of disease progression, metastases, and death from their disease. The treating urologist must be extremely active and diligent when treating patients with superficial bladder cancer. An understanding of tumor biology and current intravesical therapies is important. Furthermore, and perhaps most important, the timely decision to abandon conservative therapy and proceed with radical cystectomy and urinary diversion should be kept in mind to prevent the potentially lethal sequelae of invasive bladder cancer. In view of the potential for recurrence, stage progression, and significant clinical understaging of patients with superficial bladder cancer, and the fact that radical cystectomy provides excellent results with regard to prevention of local recurrence and overall survival, radical cystectomy should be considered in patients with high-risk factors. Radical cystectomy may also provide important prognostic information and influence the decision for adjuvant therapy based on pathologic criteria. Finally, definite improvements in lower urinary tract reconstruction and nerve sparing techniques have decreased the social implications of cystectomy for both men and women. PMID- 11101093 TI - Transurethral resection and partial cystectomy for invasive bladder cancer. AB - Bladder-preserving modalities for patients with invasive bladder cancer have become increasingly popular in recent years. Surgical-only approaches, such as transurethral resection (TUR) or partial cystectomy, are unique among a variety of bladder-preserving modalities, most of which involve combination with radiation and chemotherapy. TUR and partial cystectomy remain incompletely evaluated due to relatively small series in the literature and the lack of standardized selection criteria. The outcome as measured by long-term bladder preservation and overall survival is not dissimilar to concurrent radical cystectomy series, possibly because of positive selection of patients. PMID- 11101094 TI - Management dilemmas in bladder cancer: radical TUR +/- systemic chemotherapy in the treatment of muscle-invasive bladder cancer. AB - This case report follows the course of a highly aggressive bladder cancer that presented with carcinoma in situ and illustrates the rapid rate of progression from superficial to invasive disease that may be observed in some individuals. This case also highlights concerns about alternative management options and the dilemmas that arise when cystectomy is delayed by failed conservative strategies attempting to preserve the native bladder. This discussion focuses on the combination of radical transurethral resection (TUR) and systemic chemotherapy as a therapeutic alternative to cystectomy. The concept, technique, indications, and contraindications of radical TUR, and the rationale for combination systemic chemotherapy are discussed with reference to this case. We consider the value of radical TUR and systemic chemotherapy as an alternative to cystectomy and the reasons why this patient would not have been a suitable candidate for this bladder-sparing approach. PMID- 11101095 TI - Organ preservation by external beam and afterloading interstitial radiation in patients with muscle infiltrating bladder cancer. AB - In several European cancer centers, the combination of external beam and interstitial radiotherapy with the aim of bladder preservation is standard treatment in a selected group of patients with muscle-infiltrating bladder cancer. Three case reports are presented. The published local control rates at 5 years vary from 64% to 88% and the 5-year overall and disease-free survival range from 47% to 66% and from 62% to 81%, respectively. It is concluded that this approach is successful in preserving the bladder. Conditions for good results are careful selection of patients, excellent cooperation between urologist and radiation oncologist, and modern brachytherapy facilities. PMID- 11101096 TI - Multimodality therapy for the treatment of muscle-invasive bladder cancer. AB - In the United States, local management of muscle-invasive bladder cancer largely remains radical cystectomy with urinary diversion. However, this approach is undergoing transition. Organ-preserving approaches using a combination of multiple modalities have been successfully applied to the management of several types of cancer and clearly play an important role in the management of muscle invasive bladder cancer as well. Since the 1980s, several single and multi institutional trials have confirmed that a combined modality organ-preserving approach (chemotherapy administered in conjunction with radiation) consistently confers equivalent overall survival compared with survival following radical cystectomy. These trials are very encouraging and allow organ preservation to be considered an appropriate therapeutic option for patients with muscle-invasive bladder cancer. PMID- 11101097 TI - Presidential Address of the American Orthopaedic Society for Sports Medicine. The past, present, and future. PMID- 11101098 TI - Clinical comparison of intraarticular anterior cruciate ligament reconstruction using autogenous semitendinosus and gracilis tendons in men versus women. AB - Reconstruction of the anterior cruciate ligament using a hamstring tendon autograft has often been recommended for female athletes. We compared the results of acute, isolated, intraarticular anterior cruciate ligament reconstructions using quadruple-looped hamstring autografts in 39 female and 26 male patients. All reconstructions were performed by the same surgeon using a similar surgical technique and the same postoperative management. In each case, patients had Endobutton femoral fixation and either post or button fixation for the tibial side. The average follow-up was 40.9 months for women and 39.0 months for men. Objective analysis of results included examination for the presence of effusion and crepitus, Lachman and pivot shift testing, and KT-1000 arthrometer testing for side-to-side differences. Subjective analysis consisted of a 15-item visual analog scale completed by patients postoperatively, and pre- and postoperative Tegner and Lysholm scores. The clinical failure rate was 23% (9 of 39) for the female patients and 4% (1 of 26) for the male patients, which was statistically significant. There was also a trend toward increased laxity in female patients. Subjectively, the women also reported a higher frequency and intensity of pain. Based on Tegner activity levels, more of the men returned to their preinjury level of activity than did the women. When compared with the male patients, female patients had a significantly higher failure rate after reconstruction. PMID- 11101099 TI - In vitro structural properties of braided tendon grafts. AB - In an effort to increase strength in hamstring tendon grafts for anterior cruciate ligament reconstruction, braiding or weaving of the tendons has been suggested. The purpose of this study was to examine the biomechanical properties of two braiding techniques compared with a four-stranded tendon graft using a sheep model. Digital extensor tendons from 5 adult sheep were harvested in 28 matched pairs and randomly allocated to French plait or four-stranded weave. The grafts were tested in a hydraulic testing machine with the tendons secured in brass grips frozen with liquid carbon dioxide. The tendons were preconditioned to a distraction of 1 mm for 10 cycles followed by testing to failure at 50 mm/sec, with a data acquisition rate of 1,000 Hz. The stiffness, ultimate load to failure, and the mode of failure were recorded. All braided samples failed at the midsubstance, while the four-stranded controls failed at the grip interface. There was a significant reduction in strength and stiffness of the braided samples compared with the controls. This study demonstrated that braiding decreases the strength and stiffness of a four-stranded tendon graft by up to 54% and 85%, respectively. This finding is supported by the work of Hearle et al. (1969), who demonstrated that the decrease in strength of fiber bundles is equal to the square of the cosine of the twist angle. The twist angle in our samples was approximately 45 degrees, which equates to a decrease in strength of 50%. PMID- 11101100 TI - Recurrent shoulder instability after open reconstruction in athletes involved in collision and contact sports. AB - The purpose of this study was to evaluate the incidence of recurrent instability in a group of young athletes who underwent open shoulder stabilization with a modified Bankart repair and anterior capsulorrhaphy. Recurrent dislocation was defined as an instability episode resulting in complete dislocation requiring manual reduction. Recurrent subluxation was defined as the subjective history of the shoulder "slipping or popping out" or pain and apprehension that caused cessation of athletics for at least 1 day. Sixty-six patients (64 men and 2 women) were included in the study. A collision sport precipitated instability in 53 patients and a contact sport in 13. The average follow-up was 47 months (range, 24 to 72). The average American Shoulder and Elbow Surgeons score was 95 points (range, 71 to 100). The average Rowe score was 80 points (range, 40 to 100). Two patients had experienced recurrent dislocation after surgery (3%). Eight patients (12%) had rare (fewer than three) episodes of postsurgical subluxation. Five patients (8%) had multiple recurrent subluxations after surgery. Postsurgical recurrent instability was significantly associated with preoperative episodes of subluxation. However, all patients with rare subluxation had an excellent functional result. PMID- 11101101 TI - Cervical spine alignment in the immobilized ice hockey player. A computed tomographic analysis of the effects of helmet removal. AB - To determine if helmet removal causes a significant increase in lordosis of the cervical spine in ice hockey players, we radiographically assessed the position of the cervical spine in subjects immobilized to a standard spine backboard wearing shoulder pads both with and without a helmet. Ten adult male volunteers (ages, 18 to 28 years) with no previous history of cervical spine injuries were fitted with an appropriately sized ice hockey helmet and shoulder pads and immobilized in a supine position to a standard spine backboard. Computerized tomographic lateral scout scans were obtained of the cervical spine for three conditions: 1) no equipment (control), 2) helmet and shoulder pads, and 3) shoulder pads only (helmet removed). With the helmet removed and the shoulder pads remaining, a significant increase in C2 to C7 lordosis was found when compared with the other two conditions. Individual segmental measurements revealed a significant increase in cervical lordosis at the C6-7 level with the helmet removed compared with the helmet and shoulder pads condition. Our results demonstrate that the removal of an ice hockey helmet from a supine player causes a significant increase in lordosis (extension) of the cervical spine. We recommend that ice hockey helmets not be removed from injured players, with rare exceptions, because doing so results in unnecessary motion of the cervical spine. PMID- 11101102 TI - Immediate surgical repair of the medial patellar stabilizers for acute patellar dislocation. A review of eight cases. AB - An open surgical repair of the injured medial patellar stabilizers, including the vastus medialis obliquus muscle and the medial patellofemoral ligament, after acute patellar dislocation was studied in eight patients. At initial examination, all patients had tenderness over the adductor tubercle and a positive patellar apprehension sign. Four of eight patients had obvious ecchymosis over the adductor tubercle. Magnetic resonance imaging, diagnostic arthroscopy, and open surgical exploration documented injury to both the medial patellofemoral ligament and the origin of the vastus medialis obliquus muscle. In all patients, the torn muscle was retracted in an anterior and superior direction and an arthroscopic lateral release was performed followed by open primary repair of the medial patellofemoral ligament to the adductor tubercle and repair of the vastus medialis obliquus muscle to the adductor magnus tendon. Patients were evaluated postoperatively with the Kujala scoring questionnaire. The average follow-up was 3.0 years, with a minimum of 1.5 years. No patients experienced a recurrent dislocation. The average Kujala score was 91.9. Patients rated their return to athletic activity at an average 86% of their pre-injury level. The average subjective satisfaction was 96%. In appropriate cases of acute patellar dislocation, we recommend primary repair of the medial patellofemoral ligament and the vastus medialis obliquus muscle to avoid recurrent dislocation, chronic subluxation, pain, and disability. PMID- 11101103 TI - Resection of clinically localized segments of painful retinaculum in the treatment of selected patients with anterior knee pain. AB - We reviewed the long-term results of 25 patients who had localized soft tissue resections for refractory anterior retinacular knee pain. Patients completed visual analog scales to determine their activity and pain level changes, subjective assessment of their results, and whether they would have the surgery again under the same circumstances. Five of the 25 patients (20%) had had no knee surgery before the soft tissue excision, with the rest having had an average of two prior operations (range, 1 to 6). Subjectively, 22 patients (88%) noted moderate-to-substantial improvement after surgery; 3 patients (12%) declared no long-term benefit. All 25 patients stated that they would repeat the surgery under the same circumstances. Five patients (20%) noted a decrease in their results over time, but only two patients (8%) decreased their job level after surgery because of their knee pain. The average activity level dropped 60% after knee symptoms developed and increased 40% after surgery. Pain levels decreased 50% after surgery. The patients with the best overall results had lesions that were in the medial, inferomedial, or inferolateral retinaculum. The histologic results of the specimens included fibrosis, vascular proliferation, and small nerves with decreased myelin (neuromata). Our results show that specific soft tissue excision of painful tissue can often lead to successful clinical outcomes. PMID- 11101104 TI - The influence of functional knee bracing on the anterior cruciate ligament strain biomechanics in weightbearing and nonweightbearing knees. AB - Functional knee braces are commonly prescribed after anterior cruciate ligament injury or reconstruction; however, their ability to protect the ligament, or graft, remains unclear. Our objective was to evaluate the anterior cruciate ligament strain response in braced and unbraced knees during weightbearing and nonweightbearing in combination with three externally applied loads: 1) anterior posterior shear forces, 2) internal-external torques, and 3) varus-valgus moments. The Legend brace was tested. All external loads were applied to the tibia with the knee flexed to 20 degrees. Reproducible data were obtained from 11 subjects. For anterior shear loads up to 130 N, the brace significantly reduced strain values compared with the unbraced knee during nonweightbearing and weightbearing conditions. For internal torques of the tibia (up to 9 N x m), strain in the braced knee was significantly less than in the unbraced knee when the knee was nonweightbearing only. The brace did not reduce strain values when the knee was subjected to external torques (9 N x m) or varus-valgus moments (10 N x m) in weightbearing and nonweightbearing knees. These data indicate that a functional knee brace can protect the anterior cruciate ligament during anterior posterior shear loading in the nonweightbearing and weightbearing knee and during internal torques in the nonweightbearing knee. PMID- 11101105 TI - Upper extremity snowboarding injuries. Ten-year results from the Colorado snowboard injury survey. AB - A survey of snowboarding injuries was conducted over 10 seasons (1988 to 1998). A questionnaire evaluating 20 variables was used to collect data from 47 medical facilities near Colorado ski resorts. A total of 7430 snowboarding-related injuries were seen. A control group consisted of 3107 noninjured snowboarders. Most of those injured were 30 years of age or younger; 74% of injuries occurred in men and 26% in women; 39% of injured snowboarders were beginners and 61% were intermediate or experts. Men rode at more advanced levels than women. Injured snowboarders were more likely than noninjured snowboarders to be beginners. There were 3645 (49.06% of total) upper extremity injuries; 56.43% were fractures, 26.78% sprains, and 9.66% dislocations. The most common site of injury was the wrist (21.6% of all snowboarding injuries). Wrist fractures (except to the scaphoid) and sprains were more common in beginners, women, and younger age groups. Intermediate and expert men were more likely to sustain hand, elbow, and shoulder injuries, as well as more severe injuries. Falling was the predominant mechanism of upper extremity injuries. Snowboarders who wore protective wrist guards were half as likely to sustain wrist injuries as those who did not wear guards. PMID- 11101106 TI - Two-bundle posterior cruciate ligament reconstruction. An in vitro analysis of graft placement and tension. AB - This study had two purposes: first, to determine how femoral attachment location affects the load sharing between the two bundles of a Y-type posterior cruciate ligament reconstruction, and second, to determine how the bundles, separately and in combination, control posterior tibial translation throughout the full range of knee flexion. One and two-bundle reconstructions were performed in 12 cadaveric knees. The one-bundle reconstructions were attached within the femoral posterior cruciate ligament footprint at one of three locations, high and shallow (S1), mid and shallow (S2), or mid and deep (D). The two-bundle reconstructions comprised an S1 bundle with either an S2 or a D bundle. Posterior translation and bundle tension were measured as the knee was flexed from full extension to 1,200 of flexion while a posterior force of either 50 or 100 N was applied to the proximal tibia. The shallow one-bundle reconstruction restored posterior translation to within 2 mm of that of the intact knee over the entire range of knee flexion. The deep reconstruction did not control abnormal posterior translation above 45 degrees. The tension in the shallow bundles increased with knee flexion, and the deep bundle tension remained nearly constant throughout knee flexion. Both two bundle reconstructions controlled posterior translation, but with different load sharing characteristics. The S1-S2 configuration resisted posterior tibial translation as both bundles became taut in flexion. In contrast, the S1-D configuration resisted posterior translation in a reciprocal fashion with the D bundle tension being the greatest in extension and the S1 bundle tension being the greatest tension in flexion. PMID- 11101107 TI - Effects of interference fit screw length on tibial tunnel fixation for anterior cruciate ligament reconstruction. AB - Graft-tunnel mismatch during arthroscopically assisted anterior cruciate ligament reconstruction using the central-third patellar tendon results in less than 20 mm of bone plug remaining in the tibial tunnel. We decided to evaluate the strength of bone plug fixation using interference fit screws that were less than 20 mm in length. Biomechanical testing was performed on 48 porcine hindquarters using 9-mm diameter interference fit screws that measured 12.5, 15, and 20 mm in length. No significant difference was noted between the different-length screws for insertion torque, divergence, stiffness, displacement, or load to failure. We believe, therefore, that comparable graft fixation can be achieved in the tibial tunnel using 9-mm diameter interference fit screws that are less than 20 mm long, and that these shorter screws may be useful in cases of graft-tunnel mismatch. PMID- 11101108 TI - Comparison of score evaluations and instrumented measurement after anterior cruciate ligament reconstruction. AB - Forty-four patients who had undergone unilateral anterior cruciate ligament reconstructions were evaluated retrospectively with seven different scoring systems (International Knee Documentation Committee, Orthopadische Arbeitsgruppe Knie, Lysholm, Feagin and Blake, Zarins and Rowe, Cincinnati, and Marshall scores). The results varied between systems and therefore lacked reliability. Of the 44 patients, 32 were rated as excellent according to the Cincinnati score while only 3 patients were rated as normal according to the International Knee Documentation Committee form. Good and excellent results were found twice as frequently with the Cincinnati and Lysholm scores compared with the scores of Zarins and Rowe or the International Knee Documentation Committee form. Statistical analysis confirmed this observation and revealed significant differences between the scoring systems. Side-to-side differences using the manual maximum displacement test with the KT-1000 arthrometer revealed good correlation with the International Knee Documentation Committee and the Orthopadische Arbeitsgruppe Knie questionnaires. None of the other scoring systems, which do not measure anterior laxity, produced reasonable correlation with instrumented measurements. We found that certain population-specific factors as well as the distribution of single findings can distort the results of scoring systems. To avoid these interference factors, the patient sample should be homogeneous and selected prospectively and there should be agreement about the value of single findings. PMID- 11101109 TI - Light microscopic histology of achilles tendon ruptures. A comparison with unruptured tendons. AB - We studied biopsies from the Achilles tendons of patients undergoing open repair for a subcutaneous rupture of their Achilles tendons (27 men, 11 women; mean age, 45.3 +/- 13.8 years) and specimens of Achilles tendons from persons with no known tendon ailments (43 men, 3 women; mean age, 64.2 +/- 9.7 years). Histologic examination was performed using stained slides that were interpreted using a semiquantitative grading scale assessing fiber structure and arrangement, rounding of the nuclei, regional variations in cellularity, increased vascularity, decreased collagen stainability, hyalinization, and glycosaminoglycan. We gave up to three marks for each of these variables, with 0 being normal and 3 being maximally abnormal. All the histology slides were assessed twice in a blinded manner; the agreement between two readings ranged from 0.56 to 0.87 (kappa statistics). The score of ruptured tendons was significantly greater than the average score of control tendons (20.5 +/- 3.6 versus 6.5 +/- 2.1), and there was significantly higher degeneration in the ruptured tendons. Nonruptured Achilles tendons, even at an advanced age, and ruptured Achilles tendons are clearly part of two distinct populations. Using these staining techniques, light microscopic degeneration is not a feature of tendons from healthy, older persons. PMID- 11101110 TI - A prospective study of modified Ottawa ankle rules in a military population. Interobserver agreement between physical therapists and orthopaedic surgeons. AB - To determine the necessity of ankle and foot radiographs, we used modified Ottawa Ankle Rules to evaluate all cadets seen with an acute ankle or midfoot injury at the United States Military Academy. This scoring system determines the need for radiographs. Each patient was independently examined and the decision rules were applied by a physical therapist and an orthopaedic surgeon. Ankle and foot radiographs were obtained for all subjects. Sensitivity, specificity, and the positive predictive value were calculated in 153 patients. There were six clinically significant ankle fractures and three midfoot fractures, for a total incidence of 5.8%. For physical therapists, the sensitivity was 100%, the specificity for ankle injuries was 40%, and the specificity for foot injuries was 79%. For orthopaedic surgeons, the sensitivity was also 100%, the specificity for ankle injuries was 46%, and the specificity for foot injuries was 79%. Interobserver agreement between the orthopaedic surgeons and physical therapists regarding the overall decision to obtain radiographs was high, with a kappa coefficient value of 0.82 for ankle injuries and 0.88 for foot injuries. There were no false-negative results. Use of the modified Ottawa Ankle Rules would have reduced the necessity for ankle and foot radiographs by 46% and 79%, respectively. PMID- 11101111 TI - Anatomy and kinematics of the lateral collateral ligament of the knee. AB - The anatomy and kinematics of the lateral collateral ligament were studied in 10 unembalmed limbs and 20 isolated femurs and fibulas. The ligament's average overall length was 66 mm (range, 59 to 74) and the average greatest dimension of its thin middle portion was the anteroposterior dimension of 3.4 mm (range, 3 to 4). The center of the femoral attachment site was 3.7 mm posterior to the ridge of the lateral epicondyle, not at it apex. A potential radiographic technique for operatively locating the femoral attachment site to within 3 mm is described. During knee flexion in neutral rotation the distance between the femoral and fibular attachment sites of the lateral collateral ligament decreased to 88% of its value in full extension. With 6.5 N x m of applied external rotation force, beyond 30 degrees of flexion the attachment sites rapidly approximated. With the same internal rotation force, beyond 15 degrees of flexion the attachment sites separated. From 60 degrees to 105 degrees they were greater than 100% of the value in full extension, suggesting significant distraction between the attachment sites. These changes correlated well with the ligament's change from an 11 degrees posterior slope in extension to a 19 degrees anterior slope in flexion with no applied rotation. PMID- 11101112 TI - Boxer's knuckle in the professional athlete. AB - Injuries to the extensor mechanisms of the fingers can be career-ending in professional athletes if not treated appropriately. We identified 8 professional athletes who underwent 11 direct metacarpophalangeal joint extensor mechanism repairs including centralization of the extensor tendon and sagittal band repair between 1989 and 1994. Success of the operative procedure was determined by the athlete's attainment of full range of motion, return to professional sports, and no need for additional surgical intervention. The metacarpophalangeal joints of the long and little fingers were most commonly involved. The position of the extensor mechanism disruption and the direction of the tendon subluxation varied. Capsular tears were identified in seven joints and none were repaired. At follow up, each athlete had regained full range of motion and each had returned to professional sport an average of 5 months postoperatively. No patient required additional surgery. In this series, the principal lesion in metacarpophalangeal joint injury was extensor mechanism disruption with a predictable sagittal band tear and either a radial or ulnar subluxation of the central tendon. We recommend centralization of the extensor tendon and sagittal band repair without capsular repair as a treatment of choice for this injury, particularly in the athlete. PMID- 11101113 TI - A novel suture anchor of high-density collagen compared with a metallic anchor. Results of a 12-week study in sheep. AB - We report the early mechanical properties and histologic findings of a high density, type I collagen bone anchor. This new anchor was compared with a traditional metallic anchor in a sheep patellar tendon model. No difference in strength of the repair was noted between the two devices at any time point. The insertions on the repaired side approached the strength of the nonoperated side by 12 weeks. Histologic analysis showed that the collagen anchor integrated with the surrounding bone by 6 weeks, and there was little degradation at 12 weeks. The high-density collagen anchor supported tendon healing to bone comparable with that seen with a traditional metallic device, but it has the potential advantage of the anchor being incorporated into bone. PMID- 11101114 TI - The proliferative response of isolated human tendon fibroblasts to cyclic biaxial mechanical strain. AB - At the cellular level, dynamic strain plays a key role in cell stimulation and organization of the extracellular matrix. Although positive effects of physical strain on tendon tissue are well known, little knowledge exists on how mechanical strain affects tendon cells. In this study, human tendon fibroblasts from patellar tendon were cultured on silicone dishes. Subsequently, cyclic biaxial mechanical strain was applied to the dishes for 15, 30, and 60 minutes using a specially developed cell stretching system. After the fibroblasts were strained, cells were tested for proliferation at 6, 12, and 24 hours. As a control, cells were grown on silicone dishes but did not receive any strain. A biphasic response in proliferation was observed for the 15- and 60-minute strain periods: at 6 hours and 24 hours there was more proliferation than at 12 hours. After a strain duration time of 30 minutes, a lower proliferation rate was measured compared with control levels. This study shows that application of mechanical stress to tendon fibroblasts resulted in an alteration of cellular proliferation depending on the stress time. Our results may implicate future modifications in the treatment of ligament and tendon injuries. PMID- 11101115 TI - Voluntarily evoked positive Lachman test produced by gastrocnemius muscle contraction. A report of three cases. PMID- 11101116 TI - Rupture of both peroneal tendons in a professional athlete. A case report. PMID- 11101117 TI - Intracranial delayed epidural hematoma in a soccer player. A case report. PMID- 11101118 TI - Ipsilateral clavicle fracture, sternoclavicular joint subluxation, and long thoracic nerve injury: an unusual constellation of injuries sustained during wrestling. PMID- 11101119 TI - The pathophysiology of shoulder instability. AB - Over the last several decades there has been an improved understanding of the intricate anatomy that provides stability to the glenohumeral joint. In addition, significant advances in identifying the pathologic etiology of the unstable shoulder have occurred because of basic science glenohumeral ligament cutting studies, clinical evaluation, and the advent of arthroscopic evaluation and treatment of the unstable shoulder. This article will review the pertinent anatomy of the normal glenohumeral joint and will carefully review the pathoanatomy found in the unstable shoulder. Sports medicine specialists who treat athletes with unstable shoulders should have a firm understanding of both the normal and pathologic shoulder conditions to be able to provide the best care for these athletes. PMID- 11101120 TI - The effect of neuromuscular training on the incidence of knee injury in female athletes: a prospective study. PMID- 11101121 TI - The effect of neuromuscular training on the incidence of knee injury in female athletes: a prospective study. PMID- 11101122 TI - The effect of neuromuscular training on the incidence of knee injury in female athletes: a prospective study. PMID- 11101123 TI - The effect of reconstruction of the medial patellofemoral ligament on patellar tracking. PMID- 11101124 TI - The value of volunteering. PMID- 11101125 TI - The sun tan myth. PMID- 11101126 TI - Private offices have at least one advantage. PMID- 11101127 TI - Bilateral occipital lobe stroke with inferior altitudinal defects. AB - BACKGROUND: Cerebrovascular disease is the most common cause of neurological disability in Western countries. Patients who survive cerebrovascular accidents exclusive to the occipital lobe often have no significant neurological deficits other than visual-field loss. Visual-field defects from occipital lobe stroke typically include congruous homonymous hemianopsias or quadranopsias, with or without macular sparing. CASE REPORT: A 61-year-old white man came to us with symptoms of sudden loss of vision and difficulty reading. Visual-field testing revealed a bilateral inferior altitudinal defect with normal optic nerve and fundus appearance in both eyes. On radiological examination, he was found to have had a bioccipital lobe cerebrovascular accident secondary to complete occlusion of the left vertebral artery. An embolic event causing the artery occlusion, in combination with bilaterally compromised cerebellar and posterior cerebral arteries, presumably caused the bilateral stroke. After appropriate medical and neurological consultation, optometric management consisted of maximizing the patient's remaining vision with a prismatic spectacle correction. DISCUSSION/CONCLUSION: Patients with infarction exclusive to the occipital lobe typically have no other neurological deficits except visual-field loss and are often easier to manage than patients with infarctions to other areas of the cerebral cortex or multiple infarctions. Visual-field loss from occipital lobe damage can be successfully managed with optical systems and/or visual rehabilitation. Factors related to management include location and extent of visual-field damage, functional visual needs, and both personal and health concerns of the patient. A discussion is presented on cerebrovascular disease, occipital lobe infarction, imaging techniques, and visual rehabilitation. PMID- 11101128 TI - Case presentations of retinal artery occlusions. AB - BACKGROUND: Retinal artery occlusions typically result in sudden, unilateral painless loss of vision and may have varying presentations. They are associated with systemic diseases such as atherosclerosis, hypertension, and valvular heart disease. Additional risk factors include diabetes mellitus, cigarette smoking, giant-cell arteritis, and hyperlipidemia. They most often occur in persons 60 to 80 years of age. METHODS: Four patients have come to our clinics with varying degrees of visual loss as a result of retinal artery occlusions. The types of arteriolar occlusions presented include: precapillary arteriole occlusion, cilioretinal artery occlusion, branch retinal artery occlusion, and central retinal artery occlusion. RESULTS: Patients were followed for their ocular involvement, but also included was medical management of the underlying systemic disease condition. Workup of retinal artery occlusion included laboratory testing, carotid duplex scans, and echocardiograms to uncover the possible systemic etiologies of the artery occlusion. CONCLUSION: Optometrists should recognize the signs and symptoms of the various arterial obstructions and refer patients for systemic treatment as indicated. Patients who manifest retinal or pre-retinal artery occlusions should undergo thorough systemic evaluations for vascular disease, including: atherosclerotic disease, hypertension, and valvular heart disease. PMID- 11101129 TI - Intravitreal injection of tissue plasminogen activator and gas bubble for treatment of subretinal hemorrhage in ARMD. AB - BACKGROUND: Subretinal hemorrhage (SRH) can arise from any number of underlying etiologies and can stem from either the choroidal and/or retinal circulation. It is most commonly associated with age-related macular degeneration (ARMD), in which a choroidal neovascular membrane is the usual source of bleeding. Vision loss resulting from SRH can be secondary to toxic, tractional, and barrier effects from persistent blood. To minimize long-term visual loss from SRH, several treatment modalities have evolved over the past few years. The most recent therapeutic techniques involve treatment with the thrombolytic agent tissue plasminogen activator The value of surgical removal of subretinal hemorrhage to improve visual outcome remains unsubstantiated, as definitive studies have not been completed. CASE REPORT: A 73-year-old man manifested a 1 day history of decreased vision in his right eye. A large submacular hemorrhage had developed as a result of exudative age-related macular degeneration. Treatment included intravitreal injection of tissue plasminogen activator, followed by intravitreal injection of SF6 gas, which displaced the subretinal hemorrhage away from the fovea and resulted in clearance of the submacular blood. This case describes a new treatment for submacular hemorrhage secondary to ARMD. CONCLUSIONS: Subretinal hemorrhage secondary to ARMD can cause significant permanent visual loss. A thorough understanding of the pathogenesis of vision loss and the treatment options available are essential in successful management of these patients. Intravitreal injection of tissue plasminogen activator and gas bubble may provide an effective treatment for subretinal hemorrhage in age related macular degeneration. PMID- 11101130 TI - Duane's retraction syndrome: literature review. AB - BACKGROUND: Duane's retraction syndrome (DRS), also known as Stilling-Turk-Duane syndrome, is defined as a congenital miswiring of the lateral and medial recti muscles, resulting in an impaired ocular motility syndrome that includes palpebral fissure narrowing. The incidence of DRS is approximately 1% of the total cases of strabismus. Eighty percent of cases are unilateral and characterized by either limited abduction, limited adduction, or both. CASE REPORT: A 21-year-old man came to the clinic for a routine ocular examination without symptoms. A review of the history uncovered the presence of congenital, type I Duane's retraction syndrome. The examination demonstrated orthophoria in primary gaze, an abduction deficit O.S., and left globe retraction with palpebral fissure narrowing on right gaze O.S. MANAGEMENT: In most cases of DRS the eyes are straight in primary position and there is no amblyopia. Amblyopia, when present, is usually the result of anisometropia and not strabismus. Because our patient had no symptoms of diplopia in primary gaze (orthophoria) or in attempted right gaze (due to suppression of the left eye with abduction), prismatic and/or surgical management were not indicated, since the patient was free from binocular and cosmetic abnormalities. CONCLUSION: DRS is characterized by abnormal development of the cells in the abducens nucleus (CN VI), resulting in restricted or absent abduction and erroneous innervation of the lateral rectus by branches emanating from oculomotor nuclei (CN III). Management may include orthoptics, surgery, or monitoring. PMID- 11101131 TI - The complexities of HIPAA and administration simplification. AB - BACKGROUND: The Health Insurance Portability and Accessibility Act (HIPAA) was signed into law in 1996. Although focused on information technology issues, HIPAA will ultimately impact day-to-day operations at multiple levels within any clinical setting. Optometrists must begin to familiarize themselves with HIPAA in order to prepare themselves to practice in a technology-enriched environment. METHOD: Title II of HIPAA, entitled "Administration Simplification," is intended to reduce the costs and administrative burden of healthcare by standardizing the electronic transmission of administrative and financial transactions. The Department of Health and Human Services is expected to publish the final rules and regulations that will govern HIPAA's implementation this year. RESULTS: The rules and regulations will cover three key aspects of healthcare delivery: electronic data interchange (EDI), security and privacy. EDI will standardize the format for healthcare transactions. Health plans must accept and respond to all transactions in the EDI format. Security refers to policies and procedures that protect the accuracy and integrity of information and limit access. Privacy focuses on how the information is used and disclosure of identifiable health information. Security and privacy regulations apply to all information that is maintained and transmitted in a digital format and require administrative, physical, and technical safeguards. CONCLUSIONS: HIPAA will force the healthcare industry to adopt an e-commerce paradigm and provide opportunities to improve patient care processes. Optometrists should take advantage of the opportunity to develop more efficient and profitable practices. PMID- 11101132 TI - Web portals: supermarkets of online services. PMID- 11101133 TI - Use job analysis to hire the right employee. PMID- 11101134 TI - Notice required. PMID- 11101135 TI - Medicare to cover clinical trials. PMID- 11101136 TI - Time to tap unused employee health benefits. PMID- 11101137 TI - Early development of the secretory cavity of peltate glands in Humulus lupulus L. (Cannabaceae). AB - Early development of the secretory cavity of chemically fixed peltate glands in Humulus lupulus L. showed secretions with different densities, light, gray and dark, in the cytoplasm of disc cells and in the periplasmic space adjacent to the developing secretory cavity. Secretions were detected in the disc cell wall and subsequently in the developing secretory cavity under the subcuticular wall of the sheath. Light and gray secretions in the cavity possessed a membrane-like surface feature. Secretions were in contact with the irregular inner surface of the cuticle. Secretions contributed to the thickening of the cuticle, whereas the membrane-like surface feature contributed to a network of Cannabis striae distributed throughout the cuticle. This study supports an early development and organization of the secretory cavity in H. lupulus, parallel to those in Cannabis, and may represent common features for lipophilic glands in angiosperms. PMID- 11101138 TI - CDNA cloning of chloroplast omega-3 fatty acid desaturase from Capsicum annuum and its expression upon wounding. AB - A clone for a plastid omega-3 fatty acid desaturase (FAD) was isolated from a leaf cDNA library of hot pepper (Capsicum annuum L.). The nucleotide sequence of a 1,317 bp open reading frame in the CachFAD showed 80.9% homology with that of tobacco plant. It codes for a polypeptide of 438 amino acids with molecular mass of 50.5 kDa and a pI of 8.14. The CachFAD had a putative transit peptide for targeting the chloroplast. Genomic Southern hybridization indicated that it exists as small gene family. Northern hybridization revealed that its mRNA was present in leaves, but not in roots. Transcript levels in the leaves upon wounding increased rapidly to reach the first peak between 1-3 h, decreased thereafter and slightly increased at 24 h after wounding. The levels of linolenic acid (18:3) in wounded leaves also reached the first peak at 6 h, decreased thereafter and reached the second peak at 18 h. These results indicated that wounding not only enhanced the accumulation of the CachFAD mRNA but also increased the conversion of linoleic acid (18:2) to linolenic acid (18:3) in leaf lipids of hot pepper. PMID- 11101139 TI - Enzymatic and molecular biochemical characterizations of human neutrophil elastases and a cathepsin G-like enzyme. AB - Human neutrophil elastase (HNE, IEC 3. 4. 21. 37) is a causative factor of inflammatory diseases, including emphysema and rheumatoid arthritis. Enzymatic characterization is important for the development of new drugs involved in the regulation of this enzyme. In this study, we investigated the enzymatic and biochemical properties of five different elastolytic enzymes, with a molecular mass between 24 kDa and 72 kDa. Three elastases, molecular masses of 27, 29, 31 kDa, might be elastase isozymes that have the same NH2-terminal amino acid sequences of Ile-Val-Gly-Gly-Arg-Arg-Ala. The 24-kDa enzyme, which showed the identical NH2-terminal amino acid sequences to elastase, was a degraded fragment of native elastase. The elastolytic activity was conserved at the 6/7 domain of the NH2-terminal region. The inhibitory characteristics of PMSF, DipF were the same as those of native elastases. The 72-kDa molecule, which showed elastolytic activity, might be a trimer formed between native elastases (31 kDa and 29 kDa) and a cathepsin G-like enzyme, which did not show elastolytic activity but enhanced the elastolytic activity of neutrophil elastase. Although this cathepsin G-like enzyme showed weak cathepsin G activity, it has distinguishable NH2 terminal sequences of Ile-Val-Gly-Gly-Ser-Arg-Ala- from those of elastase or cathepsin G. The potentiation of elastolytic activity could be a result of the trimerization of native elastase with a cathepsin G-like enzyme, and was then weakly inhibited by serine protease inhibitors, such as PMSF, DipF. Therefore, we suggest the cathepsin G-like enzyme to be a novel enzyme, which has an important role in the development of inflammation. PMID- 11101140 TI - Characterization of active-site residues of the NIa protease from tobacco vein mottling virus. AB - Nuclear inclusion a (NIa) protease of tobacco vein mottling virus is responsible for the processing of the viral polyprotein into functional proteins. In order to identify the active-site residues of the TVMV NIa protease, the putative active site residues, His-46, Asp-81 and Cys-151, were mutated individually to generate H46R, H46A, D81E, D81N, C151S, and C151A, and their mutational effects on the proteolytic activities were examined. Proteolytic activity was completely abolished by the mutations of H46R, H46A, D81N, and C151A, suggesting that the three residues are crucial for catalysis. The mutation of D81E decreased kcat marginally by about 4.7-fold and increased Km by about 8-fold, suggesting that the aspartic acid at position 81 is important for substrate binding but can be substituted by glutamate without any significant decrease in catalysis. The replacement of Cys-151 by Ser to mimic the catalytic triad of chymotrypsin-like serine protease resulted in the drastic decrease in kcat by about 1,260-fold. This result might be due to the difference of the active-site geometry between the NIa protease and chymotrypsin. The protease exhibited a bell-shaped pH dependent profile with a maximum activity approximately at pH 8.3 and with the abrupt changes at the respective pKa values of approximately 6.6 and 9.2, implying the involvement of a histidine residue in catalysis. Taken together, these results demonstrate that the three residues, His-46, Asp-81, and Cys-151, play a crucial role in catalysis of the TVMV NIa protease. PMID- 11101142 TI - High temperature stress resistance of Escherichia coli induced by a tobacco class I low molecular weight heat-shock protein. AB - TLHS1 is a class I low molecular weight heat-shock protein (LMW HSP) of tobacco (Nicotiana tabacum). For a functional study of TLHS1, a recombinant DNA coding for TLHS1 with a hexahistidine tag at the amino-terminus was constructed and expressed in Escherichia coli. An expressed fusion protein, H6TLHS1, was purified using a Ni2+ affinity column and a Sephacryl S400 HR column. A polyclonal antibody against H6TLHS1 was produced to follow the fate of H6TLHS1 in E. coli. The fusion protein in E. coli maintained its solubility at a temperature of up to 90 degrees C and most of the proteins in the E. coli cell lysate with H6TLHS1 were prevented from thermally induced aggregation at up to 90 degrees C. We compared the viability of E. coli cells expressing H6TLHS1 to the E. coli cells without H6TLHS1 at a temperature of 50 degrees C. After 8 h of high temperature treatment, E. coli cells with H6TLHS1 survived about three thousand times more than the bacterial cells without H6TLHS1. These results showed that a plant class I LMW HSP, TLHS1, can protect proteins of E. coli from heat denaturation, which could lead to a higher survival rate of the bacterial cells at high temperature. PMID- 11101143 TI - Characterization of mutant HIV-1 integrase carrying amino acid changes in the catalytic domain. AB - To gain insight into the importance of conserved residues in the core domain of HIV-1 IN, we performed site-directed mutagenesis of the full-length enzyme, overexpressed the mutant proteins in E. coli, purified and analyzed their 3' processing, integration and disintegration activities in vitro. Change of E152V in the DD(35)E motif abolished all detectable activities of IN. Alteration of two highly conserved residues, P145 and K156, by isoleucine, resulted in a substantial loss or completely abolished the three activities of the enzyme. Mutant P90D weakly reduced the 3'-processing but severely affected the two other IN activities. Results obtained from double and triple mutations, P90D/P1451 and P145I/F185K/C280S, clearly suggest a crucial role of P145 in the catalytic function of IN, whereas the mutants V150E, L158F and L172M had no detectable effect on any of the IN activities. Taken together, these results allowed us to conclude that all the conserved amino acids in the core domain of IN are not equally important for catalytic functions: like D64, D116 and E152, our data suggest that P90, P145 and K156 are also essential for all three enzymatic activities of HIV-1 IN in vitro, whereas V150, L158 and L172 appear to be less critical. PMID- 11101141 TI - Evolution of the X-linked zinc finger gene and the Y-linked zinc finger gene in primates. AB - We have sequenced the partial exon of the zinc finger genes (ZFX and ZFY) in 5 hominoids, 2 Old World monkeys, 1 New World monkey, and 1 prosimian. Among these primate species, the percentage similarities of the nucleotide sequence of the ZFX gene were 96-100% and 91.2-99.7% for the ZFY gene. Of 397 sites in the ZFX and ZFY gene sequences, 20 for ZFX gene and 42 for ZFY gene were found to be variable. Substitution causes 1 amino acid change in ZFX, and 5 in ZFY, among 132 amino acids. The numbers of synonymous substitutions per site (Ks) between human and the chimpanzee, gorilla and orangutan for ZFY gene were 0.026, 0.033, and 0.085, respectively. In contrast, the Ks value between human and hominoid primates for the ZFX gene was 0.008 for each comparison. Comparison of the ZFX and ZFY genes revealed that the synonymous substitution levels were higher in hominoids than in other primates. The rates of synonymous substitution per site per year were higher in the ZFY exon than in the SRY exon, and higher in the ZFY exon than in the ZFY intron, in hominoid primates. PMID- 11101144 TI - Molecular cloning and expression analysis of 3-ketoacyl-ACP synthases in the immature seeds of Perilla frutescens. AB - We report the isolation and expression analysis of two cDNAs encoding 3-ketoacyl acyl carrier protein synthases (KAS) that are involved in the de novo synthesis of fatty acids in plastids of perilla (Perilla frutescens L.). The cDNAs, designated PfFAB1 and PfFAB24, encoded polypeptides with high sequence identities to those of KAS I and KAS II/IV, respectively, of various plants. Genomic Southern blots revealed that there was a single PfFAB1 gene but two PfFAB24 genes in the perilla genome. Of interest is that the expression of both genes was developmentally regulated in seeds. Their mRNA expression patterns in seeds were also discussed in comparison with the profile of fatty acid accumulation. PMID- 11101145 TI - Effective gene transfer into regenerating sciatic nerves by adenoviral vectors: potentials for gene therapy of peripheral nerve injury. AB - Replication defective adenoviral vectors have been demonstrated as an effective method for delivering genes into a variety of cell types and tissues both in vivo and in vitro. Transfecting genes into neuronal cells has proven to be difficult because of their lack of cell division. Since the major problem in neurological disease is the degeneration of the terminally differentiated neuronal cells, the adenoviral vector's ability to transfer genes into differentiated post-mitotic cells makes them advantageous for a gene delivery system for the nervous system. Here we showed that a replication defective recombinant adenovirus carrying the lacZ gene could infect the neuronal stem cells and even the differentiated neuronal cells derived from the central nervous system. The lacZ gene delivered into the neuronal cells was expressed efficiently. In addition, the recombinant virus also infected Schwann cells in intact and injured nerves in vivo. The expression of the lacZ gene lasted for 5 weeks, within which nerve regeneration is accomplished in the rat. Adenoviral vectors might thus be used to modulate Schwann cell gene expression for treating peripheral nerve injury or peripheral neuropathy. PMID- 11101146 TI - c-Fos expression by dopaminergic receptor activation in rat hippocampal neurons. AB - While dopamine is likely to modulate hippocampal synaptic plasticity, there has been little information about how dopamine affects synaptic transmission in the hippocampus. The expression of IEGs including c-fos has been associated with late phase LTP in the CA1 region of the hippocampus. The induction of c-fos by dopaminergic receptor activation in the rat hippocampus was investigated by using semiquantitative RT-PCR and immunocytochemistry. The hippocampal slices which were not treated with dopamine showed little expression of c-fos mRNA. However, the induction of c-fos mRNA was detected as early as 5 min after dopamine treatment, peaked at 60 min, and remained elevated 5 h after treatment. Temporal profiles of increases in c-fos mRNA by R(+)-SKF-38393 (50 microM) and forskolin (50 microM) were similar to that of dopamine. An increase in [cAMP] was observed in dopamine-, SKF-, or forskolin-treated hippocampal slices. By immunocytochemical studies, control hippocampal cells showed little expression of c-Fos immunoreactivity. However, when cells were treated with dopamine, an increase in the expression of c-Fos immunoreactivity was observed after treatment for 2 h. The treatment of hippocampal neurons with R(+)-SKF38393 (50 microM) or forskolin (50 microM) also induced a significant increase in c-Fos expression. These results indicate that the dopamine D1 receptor-mediated cAMP dependent pathway is associated with the expression of c-Fos in the hippocampal neurons. These data are consistent with the possible role of endogenous dopamine on synaptic plasticity via the regulation of gene expression. Furthermore, these results imply that dopamine might control the process of memory storage in the hippocampus through gene expression. PMID- 11101147 TI - FLIP is constitutively hyperexpressed in Fas-resistant U266 myeloma cells, but is not induced by IL-6 in Fas-sensitive RPM18226 cells. AB - Despite the expression of Fas, some clones of myeloma cells are resistant to Fas mediated apoptosis. To define a cellular factor involved in the resistance, we performed a comparative study using two clones of myeloma cells, RPM18226 and U266. These cells were reported to express cell surface Fas at similar levels, but only RPM18226 cells lost their viability upon anti-Fas treatment. The resistance of U266 cells to anti-Fas did not appear to reflect dysregulation of Bcl-2, Bcl-X(L), and Bax, because these proteins were expressed in both RPM18226 and U266 cells to similar levels. Moreover, levels of those proteins were not significantly altered by treating RPM18226 cells with IL-6, a cytokine which suppresses the Fas-mediated death of RPM18226 cells. Interestingly, mRNA levels of FLIP(L), an endogenous inhibitor of Fas signaling, were constitutively elevated in U266 cells. Consistent with this observation, U266 cells expressed both FLIPL protein and its truncated 43 kDa product which is seen in FLIP(L) overexpressing cells. The truncated form of FLIP(L) protein was not detected in RPM18226. Moreover, the levels of truncated FLIP(L) in U266 cells were considerably higher than those of pro-FLIP(L) in RPM18226. The overall data indicate that FLIPL is constitutively hyperexpressed in U266 cells. However, IL-6 failed to enhance the protein levels of FLIP molecules in either of the tested cells. It appears, therefore, that FLIP(L) plays a role in the intrinsic resistance of U266 cells to the apoptotic action of Fas, but is not involved in the protective action of IL-6. PMID- 11101148 TI - Inactivation of p16INK4a in primary tumors and cell lines of head and neck squamous cell carcinoma. AB - Inactivation of the p16INK4a gene by mutation and deletion is common in head and neck squamous cell carcinoma (HNSCC). The present study demonstrates that hypermethylation of the 5' CpG islands can serve as an alternative mechanism for the inactivation of the p16INK4a gene in this tumor. We studied 11 HNSCC cell lines and 17 oral squamous cell carcinoma (OSCC) primary tumors for p16INK4a gene status by protein/mRNA and DNA genetic/epigenetic analyses to determine the incidence of its inactivation. Our study indicates that: (1) inactivation of p16 protein is frequent in HNSCC cell lines (6/11, 54.5%) and OSCC primary tumors (15/17, 88.2%), (2) inactivation of p16INK4a protein is commonly associated with the presence of gene alteration such as mutation, homozygous deletion and especially aberrant methylation, and (3) genomic sequencing of bisulfite-modified DNA shows that the carcinoma develops a heterogeneous pattern of hypermethylation. PMID- 11101149 TI - 5'-Flanking sequence and promoter activity of the rabbit neuronal nitric oxide synthase (nNOS) gene. AB - We have isolated a rabbit neuronal nitric oxide synthase (nNOS) cDNA encoding a protein of 1,435 amino acids. Using the cDNA clones as probes, the 5'-flanking region of the nNOS gene was isolated from a rabbit genomic DNA library. 5'RACE and primer extension analysis of rabbit brain total RNA mapped multiple transcription initiation sites localized 474-487 bp upstream from the translation start codon. Analysis of 5,197 bp of the 5'-flanking sequence revealed that the rabbit nNOS gene promoter lacks canonical TATA or CCAAT boxes and, instead, contains a GC-rich region and multiple Sp1 sites. Farther from the +1start, various putative cis-elements including AP-1, AP-4, NF-kappaB, STAT, CREB, C/EBP and c-Myc were observed. The functional promoter activity of the 5'-flanking region was demonstrated by its ability to drive the expression of a beta galactosidase reporter gene in several cell types. Serial deletion analysis of the promoter region revealed that the -291 to -172 region, which contains two Sp1 sites, is essential for basal transcriptional activity. These results suggest that the rabbit nNOS promoter contains characteristics of inducible genes. PMID- 11101150 TI - A new non-radioactive method for IL-2 bioassay. AB - An oxidation-reduction (redox) indicator, alamarBlue, was used to measure the bioactivity of interleukin 2 (IL-2). This assay system has several advantages over other bioassays for measuring IL-2. It is a nonradioactive method unlike the conventional tritium-labeled thymidine ([3H]TdR) incorporation assay. The alamarBlue assay is also easier to use than other colorimetric methods, such as the MTT assay, because the alamarBlue assay does not depend on the extraction of insoluble formazan salt, which is time-consuming, error-prone, and cumbersome. Due to its solubility in culture medium and its nontoxicity to cells, alamarBlue provides an easy method to monitor cellular growth using either a fluorescence- or an absorbance-based instrument. The alamarBlue assay is not sample destructive, unlike the thymidine incorporation and MTT methods. This adds another advantage to the alamarBlue method as the measurement of cellular growth by sample-destructive methods requires as many tubes as time points whereas the alamarBlue method requires only one tube for the entire growth period. In this study, alamarBlue was used to measure the proliferation of the IL-2-dependent cytotoxic T cell line, CTLL-2. The colorimetric change of alamarBlue at 570 nm compared to the reference wavelength, 600 nm, was proportional to the number of viable cells. The sensitivity of the IL-2 assay using alamarBlue was comparable to that of the [3H]thymidine incorporation method. These results demonstrate that the alamarBlue assay is valid for the IL-2 bioassay and that alamarBlue can replace the [3H]thymidine employed in the conventional proliferation assays. PMID- 11101151 TI - Characterization of geranium (Pelargonium graveolens) chloroplast EF-Tu cDNA. AB - A geranium (Pelargonium graveolens) chloroplast translational elongation factor EF-Tu (tufA) cDNA was isolated. The geranium tufA cDNA is 1,584 bp long with 20 bp of 5' untranslated region (UTR) and 139 bp of 3' UTR. It encodes 474 amino acids including a putative chloroplast transit peptide of 65 amino acids. The deduced polypeptides of the geranium tufA cDNA contains four GTP binding sequences in its N-terminal region and two chloroplast EF-Tu signature regions in the C-terminal region. The predicted molecular weight of the mature geranium chloroplast EF-Tu protein was about 45,000 and its amino acid sequence identity with the chloroplast EF-Tu proteins of tobacco, pea, Arabidopsis, rice, and soybean ranges from 85% to 91%. The geranium tufA appears to exist as a single copy gene like Arabidopsis and rice, whereas other known dicot plants have more than one copy in their nuclear genomes. PMID- 11101152 TI - Cloning of hHRI, human heme-regulated eukaryotic initiation factor 2alpha kinase: down-regulated in epithelial ovarian cancers. AB - Protein synthesis is regulated in response to environmental stimuli by covalent modification, phosphorylating the components of the translational machinery. Phosphorylation of the alpha subunit of eIF-2 is one of the best-characterized mechanisms for down-regulating protein synthesis in higher eukaryotes in response to various stress conditions. One of mammalian eIF-2alpha kinases is a heme regulated inhibitor kinase (HRI), which is activated by heme deficiency and plays an important role in translational control. In this work, we have analyzed the differentially expressed genes between epithelial ovarian cancer and normal ovary. We have screened a total of 1,408 genes isolated from a human dermal papilla cell cDNA library by cDNA array hybridization. Among many differentially expressed genes, eIF2alpha kinase, a heme regulated inhibitor was down-regulated in ovarian epithelium cancer. The down-regulation of hHRI was also confirmed in other ovarian cancer tissues by Northern blot hybridization. The hHRI gene is 2,887 bp in length and the amino acid sequence deduced from the cDNA clone encodes a protein of 630 amino acids with molecular mass of 73 kDa. It contains all 12 catalytic domains of the protein kinases with consensus sequences of the protein-serine/threonine kinases. The expression pattern of hHRI mRNA showed approximately 3.0 kb bands which were expressed ubiquitously in all human tissues examined, which indicates that eIF-2alpha kinase could play an important role in the translational regulation of nonerythroid tissues. PMID- 11101153 TI - A physical interaction of UvrD with nucleotide excision repair protein UvrB. AB - The dual-incision nature of the reaction of UV-irradiated DNA catalyzed by the UvrABC complex potentially leads to excision of a damaged fragment. However, neither fragment release under nondenaturing conditions nor the UvrBC proteins are turned over. The addition of the UvrD protein to the incised DNA-UvrBC complex results in excision of the incised damaged strand and in the turnover of the UvrC protein. In an effort to better understand the involvement of UvrD in the excision step, immunoprecipitation was used to detect interacting proteins with UvrD in the DNA repair. In this communication, it is shown that UvrA and UvrB are precipitated with UvrD in solution but the UvrAB complex is not. In the incision complex, UvrB could be precipitated and the preincubation of UvrD with UvrB revealed an inhibitory effect on the turnover of the incision complex. These data imply that UvrB in the incision complex seems to recruit UvrD to the 3' incised site of the incised strand by protein-protein interaction and to allow initiation of unwinding by UvrD from the resulting nick in a 3' to 5' direction. PMID- 11101154 TI - Human Fas associated factor 1, hFAF1, gene maps to chromosome band 1p32. AB - Human Fas associated factor 1 protein (hFAF1) is involved in the positive regulation of Fas signaling even though it can not initiate the signal for itself. By chromosomal assignment using somatic cell hybrids (CASH), the hFAF1 gene was located on human chromosome 1 between markers D1S443 and D1S197. The hFAF1 gene was mapped to human chromosome band 1p32 by FISH utilizing a genomic PAC clone containing the gene. In genomic Southern analysis using hFAF1 cDNA as a probe, several bands appeared in three different restriction enzyme digestions. The single band appearance in FISH analysis compared to several bands in Southern blots implies that the hFAF1 gene would be rather big or that an additional hFAF1 gene isotype(s) might be present in close vicinity. PMID- 11101155 TI - Crosslinkable coatings from phosphorylcholine-based polymers. AB - 2-Methacryloyloxyethyl phosphorylcholine (MPC) was synthesised and then used in the preparation of crosslinked polymer membranes with lauryl methacrylate, hydroxypropyl methacrylate and trimethoxysilylpropyl methacrylate (crosslinker) comonomers. Some physical aspects of the membrane properties were evaluated in order to establish the basis for the synthesis of a series of post-crosslinkable polymers. These materials were made by copolymerisation of the constituent monomers via a free radical method, and characterised using NMR, FT-IR, viscometry and elemental analysis. The optimum crosslink density and conditions required for curing coatings of these polymers were investigated using atomic force microscopy (AFM) and showed the inclusion of 5 mol% silyl crosslinking agent to be ideal. A nanoindentation technique was employed to determine if the coating developed elasticity upon crosslinking. The biological properties of the coatings were evaluated using a variety of protein adsorption assays and blood contacting experiments, and an enzyme immunoassay was developed to detect E. coli in order to assess the level of bacterial adhesion to these biomaterials. Polymers of this type were shown to be very useful as coating materials for improving the biocompatibility of, or reducing the levels of adherent bacteria to medical devices. PMID- 11101156 TI - Thermo-debonding mechanisms in dentin bonding systems using finite element analysis. AB - The finite element method (FEM) has been extensively used in evaluating the interfacial status of biomaterials. We used FEM to explore the microscopic debonding mechanism of the dentin/hybrid layer/resin adhesive interface. The stress status of the local material was used as an index to judge whether the adhesive interface would develop a debonding mechanism. To generate the local stress concentration, the thermal boundary condition was applied to the model which has the phenomenon of the coefficient of thermal expansion (CTE) mismatch. The thermal boundary condition was used to emulute a previous study conducted with a laser thermoacoustic technique (LTAT). The materials, Scotchbond MP, Optibond, and Tenure bonding systems, used in the previous experiment were also tested in this study. The results show that interfacial debonding in the finite element model occurred through the hybrid layer for both the Scotchbond MP and Tenure systems, as well as within the adhesive layer itself for the Optibond system. These findings are compatible with observations by SEM obtained by LTAT. Another transformed model was created to test the "elastic cavity wall" concept. The result also confirms the importance of the elastic cavity wall concept. These compatible results between FEM and LTAT indicate that FEM can be a very useful supplement to thermoacoustic testing. PMID- 11101157 TI - Characterization of electrolytic ZrO2 coating on Co-Cr-Mo implant alloys of hip prosthesis. AB - An electrolytic Zr(OH)4 gel has been coated on ASTM F-75 Co-Cr-Mo alloy specimens in 0.0625 M ZrO(NO3)2 solution with pH = 2.2 at a current density of 2 mA/cm2. After annealing at 623-973 K for 120 min in air, the ZrO2-coated specimen was evaluated by electrochemical polarization in Hank's solution, wear tests with UHMWPE (Ultra-high molecular-weight polyethylene) under a load stress of 50 MPa, scratch tests, surface morphology observations, and XRD analysis. The ZrO2-coated specimen annealed at 773 K for 120 min revealed a good adhesion of 610 MPa on Co Cr-Mo substrate, a lower wear loss of UHMWPE and a higher protection potential than the uncoated specimen in Hank's solution. A monoclinic structure with (1 1 1) preferred orientation parallel to the sheet plane was detected at 623 K < or = T < or = 673 K and a tetragonal structure of ZrO2 was detected at T > or = 773 K. Then a monoclinic structure with random orientation and a tetragonal structure were mixed at T > or = 973 K. PMID- 11101158 TI - Resorbable bioceramics based on stabilized calcium phosphates. Part II: evaluation of biological response. AB - Synthetic materials capable of being remodelled in vivo by the same processes responsible for natural bone turnover have long been sought for use as an artificial bone substitute. These materials must ideally combine osteoinductive capacity with the ability to withstand random dissolution at normal physiological pH, while being resorbed by natural cell-mediated processes. Resorbable calcium phosphate based coatings and bulk ceramics have been developed which promote the uniform deposition of new mineralized bone matrix thus enabling rapid integration with the surrounding host bone tissue in vivo. Furthermore, a critical result of this study is the determination that the silicon-stabilized calcium phosphate ceramics are essentially insoluble in biological media but are resorbed when acted upon by osteoclasts. In vitro biological testing and preliminary in vivo testing show that the important features of this new biomaterial are a characteristic calcium phosphate phase composition and a unique microporous morphology. PMID- 11101159 TI - Immobilization of heparin to EDC/NHS-crosslinked collagen. Characterization and in vitro evaluation. AB - In the present study, heparin immobilization to a non-cytotoxic crosslinked collagen substrate for endothelial cell seeding was investigated. Crosslinking of collagen using N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) and N hydroxysuccinimide (NHS) resulted in a material containing 14 free primary amino groups per 1000 amino acid residues (E/N14C). At a fixed molar ratio NHS:EDC of 0.6, the amount of heparin covalently immobilized to E/N14C increased with increasing molar ratios of EDC to heparin carboxylic acid groups (Hep-COOH), to a maximum of approximately 5-5.5 wt% at a ratio of 2. Upon incubation in cell culture medium of endothelial cells, 4 to 7% of the immobilized heparin was released during 11 days. Immobilization of increasing amounts of heparin to E/N14C progressively reduced activation of contact activation proteases. Optimal anticoagulant activity, as measured by thrombin inhibition, was obtained after heparin immobilization using a ratio of EDC to Hep-COOH of 0.2-0.4 (14-20 mg heparin immobilized per gram of collagen). Platelets deposited to (heparinized) E/N14C showed only minor spreading and aggregation, although heparin immobilization slightly increased the number of adherent platelets. The results of this study suggest that heparin immobilization to EDC/NHS-crosslinked collagen may improve the in vivo blood compatibility of this material. PMID- 11101160 TI - Fabrication and characterization of a sponge-like asymmetric chitosan membrane as a wound dressing. AB - A novel asymmetric chitosan membrane has been prepared by immersion-precipitation phase-inversion method and evaluated as wound covering. This new type of chitosan wound dressing which consists of skin surface on top-layer supported by a macroporous sponge-like sublayer was designed. The thickness of the dense skin surface and porosity of sponge-like sublayer could be controlled by the modification of phase-separation process using per-evaporation method. The asymmetric chitosan membrane showed controlled evaporative water loss, excellent oxygen permeability and promoted fluid drainage ability but could inhibit exogenous microorganisms invasion due to the dense skin layer and inherent antimicrobial property of chitosan. Wound covered with the asymmetric chitosan membrane was hemostatic and healed quickly. Histological examination confirmed that epithelialization rate was increased and the deposition of collagen in the dermis was well organized by covering the wound with this asymmetric chitosan membrane. The results in this study indicate that the asymmetric chitosan membrane thus prepared could be adequately employed in the future as a wound dressing. PMID- 11101161 TI - Interactions of bioactive glasses with osteoblasts in vitro: effects of 45S5 Bioglass, and 58S and 77S bioactive glasses on metabolism, intracellular ion concentrations and cell viability. AB - In a cell culture model of murine osteoblasts three particulate bioactive glasses were evaluated and compared to glass (either borosilicate or soda-lime-silica) particles with respect to their effect on metabolic activity, cell viability, changes in intracellular ion concentrations, proliferation and differentiation. 45S5 Bioglass caused extra- and intracellular alkalinization, a rise in [Ca2+]i and [K+]i, a small plasma membrane hyperpolarization, and an increase in lactate production. Glycolytic activity was also stimulated when cells were not in direct contact with 45S5 Bioglass particles but communicated with them only through the medium. Similarly, raising the pH of culture medium enhanced lactate synthesis. 45S5 Bioglass had no effect on osteoblast viability and, under most conditions, did not affect either proliferation or differentiation. Bioactive glasses 58S and 77S altered neither the ion levels nor enhanced metabolic activity. It is concluded that: (1) some bioactive glasses exhibit well-defined effects in osteoblasts in culture which are accessible to experimentation; (2) 45S5 Bioglass causes marked external and internal alkalinization which is, most likely, responsible for enhanced glycolysis and, hence, cellular ATP production; (3) changes in [H+] could contribute to alternations in concentrations of other intracellular ions; and (4) the rise in [Ca2+]i may influence activities of a number of intracellular enzymes and pathways. It is postulated that the beneficial effect of 45S5 on in vivo bone growth and repair may be due to some extent to alkalinization, which in turn increases collagen synthesis and crosslinking, and hydroxyapatite formation. PMID- 11101162 TI - Collaboration between social workers and physicians: perspectives on a shared case. AB - As health care is being restructured, health care institutions are recognizing that interdisciplinary collaboration is an essential element of both effective patient care and organizational survival. This paper analyzes self-reported views of actual collaborative activities between 50 pairs of social workers and physicians on a specific shared case in an acute care hospital setting. Through examining the degree of congruence in perspectives of each pair of collaborators, we compare the two professions' views of the collaborative process and outcome. Additionally, each profession's outlook on its own and the other profession's roles and responsibilities in the case is examined. Our findings indicate that many social work and physician collaborators share similar perspectives about many aspects of their joint patient care endeavors. Where there was disagreement within a pair, almost always, it was a social worker selecting or discussing a variable when her physician counterpart did not. Physicians were less likely than their social work counterparts to identify patient/family problems related to adjustment to illness and problems connected to hospital and community resources as well. Social workers were much less satisfied with the collaboration, saw many more things that they or their collaborator could have done differently and even perceived more disagreement about the approach to the case than did their physician collaborators. It is important to understand, empirically, the dynamics of successful collaboration and to assist social workers in becoming influential and effective collaborators with other health professionals. PMID- 11101163 TI - Social work and the behavioral sciences: past history, future prospects. AB - The relationship between social work and medical sociology is analyzed, utilizing case studies of an academic institution, Harvard University, and a private foundation, the Russell Sage Foundation. Both strongly supported social work during the first decades of the twentieth century, and both changed their focus to social science. Why this change occurred, and its implications for social work and the behavioral sciences in medical education is the central theme of this discussion. PMID- 11101164 TI - Complementary alternative medicine in health and mental health: implications for social work practice. AB - This article describes the increasing use of complementary alternative medicine (CAM) in this country and its implications for social work practice, education, research and policy in the health care field. Descriptive examples of CAM treatment modalities are provided along with their underlying rationale, common uses and available empirical support. It is concluded that patients will be better served by social workers who have knowledge of CAM treatment modalities. Furthermore, the need for further research on the efficacy of many CAM treatments and the certification of CAM treatment providers is discussed. PMID- 11101165 TI - Managed care and its implications for social work curricula reform: policy and research initiatives. AB - A multitude of social, economic, and health factors are forcing the health care industry to examine the effectiveness and efficiency of health and mental health care delivery. Managed care has emerged in this environment of cost containment, as a viable distributive mechanism for the allocation of health care resources. Schools of social work must consider additions and/or revisions to their curriculum, in order to adequately prepare social workers to practice in a managed care environment. A major barrier is the negative attitude held by many practitioners and educators towards managed care. This paper discusses the evolution of managed care, and the challenges to social work education, including the necessity for developing a more balanced view of managed care. Suggested curricular revisions are provided for two major content areas: policy and research. PMID- 11101166 TI - Managed care and its implications for social work curricula reform: clinical practice and field instruction. AB - Managed care continues to be a major focus and debate within the health care field. Regardless of whether one is for or against managed care, it has become the predominant system for distributing finite health care resources. Many academicians and practitioners point to the uncertainty about the future of health care, and the role of social workers to practice within these new environments, schools of social work will need to integrate content related to managed care. Students should be exposed to a more balanced analysis of what is advantageous and problematic with managed care. This paper offers recommendations regarding ways in which the practice and field curriculum can be strengthened to more effectively prepare social workers for practice within a managed care environment. PMID- 11101167 TI - Effect of inhaled corticosteroids on intraocular pressure. PMID- 11101168 TI - Risks of penicillin skin testing. PMID- 11101169 TI - Progress towards an understanding of idiopathic anaphylaxis. PMID- 11101170 TI - Anaphylaxis to deer dander. PMID- 11101171 TI - How to report atmospheric pollen counts in journal publications. PMID- 11101172 TI - Quality of life in patients with allergic rhinitis. AB - LEARNING OBJECTIVES: Reading this article will familiarize the practitioner with ongoing efforts to measure the effects of allergic rhinitis symptoms and its treatments on the health-related quality of life (HRQOL) of patients. The reader will learn about instruments used to collect HRQOL data, results of studies showing the burden of illness and the effects of treatments on HRQOL, and efforts to interpret the clinical relevance of changes in HRQOL status. DATA SOURCES: Information was gleaned from articles listed in MEDLINE regarding HRQOL in allergic rhinitis between 1966 and 2000 (English language only), and from the personal experiences of the authors. STUDY SELECTION: Questionnaire validation studies and representative controlled trials employing measures to assess the effects of allergic rhinitis symptoms and its treatments on HRQOL are described. RESULTS FROM THIS REVIEW: Allergic rhinitis symptoms can have detrimental effects on the physical, psychologic, and social aspects of patients' lives. Clinical trial data suggest a variety of pharmacologic therapies can significantly improve HRQOL in patients with allergic rhinitis. Validated questionnaires are now available that can easily be used in clinical practice to measure the effects of interventions on HRQOL for individual patients. CONCLUSIONS: Evaluating the effects of interventions on HRQOL may be particularly important in a non-life threatening condition such as rhinitis. Health-related quality of life measures can be used to indicate the risk/benefit and the cost/benefit ratios of competing treatment options. Clinicians and policy makers are already using HRQOL data to evaluate results of medical interventions to guide patient management and reimbursement decisions. PMID- 11101173 TI - Failure to thrive and two weeks of persistent vomiting in an 11-month-old infant. PMID- 11101174 TI - Advice from your allergist: drug reactions. PMID- 11101175 TI - Association between intraocular pressure and budesonide inhalation therapy in asthmatic patients. AB - BACKGROUND: The extent to which inhaled glucocorticoids increase the risk of intraocular pressure elevation has been controversial. OBJECTIVE: The authors attempt to assess such risk attributable to budesonide, an inhaled glucocorticoid for asthma therapy. METHODS: Data were pooled from four prospective, randomized, double-blind, parallel-group, placebo-controlled, multicenter clinical trials of 12 to 20 weeks in duration. One thousand two hundred and fifty-five patients, 6 to 70 years of age whose intraocular pressures (IOPs) were less than 23 mmHg at screening were randomized to receive placebo or inhaled budesonide at doses ranging from 100 to 800 microg, administered twice daily. Intraocular pressure was measured at screening and at the end of double-blind treatment. Intraocular change was compared between budesonide and placebo, accounting for the confounding effects of gender, race, age, history of diabetes, history of hypertension, clinical trial, systemic glucocorticoid use during the trials, ophthalmic glucocorticoid use during the trials, and prior oral glucocorticoid use. RESULTS: No budesonide treatment effect on the IOP was evident either in the crude analysis or after adjustment for possible confounding factors. For patients exposed to budesonide at a total daily dose of 1600 microg for 20 weeks, there was no difference in IOP change compared with the placebo controls. CONCLUSIONS: No association with an increased IOP was observed in asthmatic patients treated with budesonide at daily doses ranging from 200 to 1600 microg for durations of 12 to 20 weeks. The subgroup analysis, which focused on the highest dose and longer term therapy was reassuring, as was the overall result. PMID- 11101176 TI - Frequency of systematic reactions to penicillin skin tests. AB - BACKGROUND: Penicillin skin testing is generally considered to be safe when performed sequentially with puncture and intradermal testing although fatalities have been reported. OBJECTIVE: We analyzed the rate of systemic reactions to penicillin skin tests for a period of seven and two-thirds years. METHOD: This retrospective study used a computerized database at the Mayo Clinic. Altogether 1710 patients were skin-tested to penicillin from January 1992 to September 1999. All patients tested had a history of penicillin allergy. Patients were tested with benzylpenicilloyl polylysine (Pre-Pen) (6.0 X 10(-5) M), freshly prepared penicillin G (10,000 units/ml), and penicilloate (0.01 M). Prick tests were done first and if negative then intradermal tests. Systemic reactions were evaluated and treated by physicians. RESULTS: Eighty-six patients had positive penicillin skin tests of which two had systemic reactions. Our systemic reaction rate for all patients tested was 0.12%; and 2.3% for the penicillin skin test-positive group, with no fatalities. CONCLUSION: The incidence of systemic reaction to penicillin skin tests is low. Skin prick tests should always be done first. If there is a history of a previous serious reaction, the skin tests-if done-should be diluted to start with. Those doing penicillin skin tests should be prepared to treat a systemic reaction. PMID- 11101177 TI - Lymphocyte subsets and activation markers in patients with acute episodes of idiopathic anaphylaxis. AB - BACKGROUND: Idiopathic anaphylaxis (IA), a type of anaphylaxis in which no external allergen can be identified, is a corticosteroid-responsive disease, that suggests that it may have an immunologic pathogenesis. OBJECTIVE: The objective of this study is to compare patients with acute episodes of IA with normals, patients with chronic idiopathic urticaria, and patients with IA in remission relative to lymphocyte subsets and activation markers. METHODS: This is a prospective cohort study of 38 adults: 5 normals, 4 idiopathic urticaria, 11 IA patients in remission, 9 IA patients with acute attacks who had not yet received prednisone, and 9 IA patients who had received prednisone. The main outcome measures were lymphocyte subset and activation markers determined by two and three color flow cytometry (CD2, CD3, CD4, CD5, CD8, CD16, CD19, CD23, CD25, CD56, and HLA-DR). RESULTS: Comparing patients with acute IA with those in remission, the only significant difference was that the acute IA patients had a significantly higher percentage of CD3+HLA-DR+ cells. Normals had a significantly lower percentage of CD3+ HLA-DR+ cells than all other groups. Patients with acute IA on prednisone as well as IA patients in remission had a significantly higher percentage of CD 19+ CD23+ cells than normals. CONCLUSIONS: These results suggest that there are more activated T cells in patients with acute episodes of IA than in patients in remission. Perhaps, these activated T cells have a role in the pathogenesis of IA. PMID- 11101178 TI - Anaphylaxis to deer dander in a child: a case report. AB - BACKGROUND: Hypersensitivity to deer dander is rarely reported, with only 26 cases in the literature. Ours is the youngest reported case and the first reported case of anaphylaxis on exposure to a live deer. OBJECTIVE: Evaluation of a case of anaphylaxis in a young boy upon exposure to a deer. METHODS AND RESULTS: A 4-year-old boy experienced hives, swelling, and shortness of breath requiring epinephrine following a deer exposure. He had one mild reaction 5 days prior to his anaphylaxis with an indirect exposure. A deer dander extract was made from fur supplied by the patient's mother. IgE-mediated reactivity was positive for deer and cattle by both selective skin prick method and RAST results. CONCLUSION: Hypersensitivity to wild animals can lead to life threatening anaphylaxis, even in children. Passive transfer of antigen may occur, but needs further investigation. PMID- 11101179 TI - Late summer and fall (March-May) pollen allergy and respiratory disease in Northern New South Wales, Australia. AB - BACKGROUND: Many people in the subtropical Northern Rivers area of New South Wales, Australia, blame the pollen of Tibouchina tree, which flowers at the same time as ragweed, Bahia grass and Bermuda grass, for hayfever and asthma exacerbations during fall between March and May. OBJECTIVES: To determine whether Tibouchina pollen is allergenic. To determine whether airborne ragweed pollen is present in this region for sufficient length of time and concentration to cause fall respiratory symptoms, and to determine if Bahia grass and Bermuda grass are associated with fall respiratory symptoms. METHODS: Pollen and Alternaria spores were monitored using a Burkard 7-day spore trap. Two hundred and six volunteers in the Northern Rivers area filled in questionnaires before skin prick tests (SPT) were performed with a panel of skin testing extracts. RESULTS: One hundred fifty-three (74.3%) subjects were atopic and reacted to one or more aeroallergens. Seventy were SPT positive to ragweed, OR 3.36 (CI 1.03 to 12.15) and 11 to Tibouchina (OR incalculable). Fifty of the 70 ragweed-positive subjects had fall hayfever or exacerbations of hayfever and/or asthma, OR 23.4 (CI 8.90 to 64.00). Eleven subjects were SPT positive to Tibouchina extract. There was a statistical association between Bermuda grass and hayfever, but not asthma OR 13.44 (CI 1.85 to 27.04). CONCLUSIONS: Ragweed pollen was present for a sufficient length of time and concentration to sensitize and provoke fall hayfever and asthma exacerbations. Tibouchina pollen is an aeroallergen causing mild-to-moderate allergic symptoms in a few people. There is an association between Bahia grass and asthma in children, and between Bermuda grass and allergic rhinitis in adults. PMID- 11101180 TI - Accuracy of patient prediction of allergy skin test results. AB - BACKGROUND: The ability of patients to self-diagnose allergy is unknown. OBJECTIVE: To estimate the ability of patients to correctly predict the results of allergy skin tests. METHODS: We conducted a structured interview of 86 patients with chronic rhinitis or asthma undergoing aeroallergen skin tests. We asked, "Do you expect the skin tests to be positive or negative?" and "What do you expect the allergy tests to be positive for?" Responses to these questions were correlated with the results of aeroallergen skin tests. Skin tests were performed using the prick technique and included cat, grass pollen, tree pollen, weed pollen, dust mites, and molds. RESULTS: Seventy-three participants provided usable responses. Of those with a positive skin test, the number (percentage) of participants who predicted correctly was 10/18 (56%) for cat, 4/14 (29%) for tree, 7/26 (27%) for weeds, 5/23 (22%) for dust mite, 2/12 (17%) for grass, and 1/8 (12%) for mold. Of those with a negative skin test, the number (percentage) of participants who predicted correctly was 47/50 (94%) for dust mite, 51/59 (86%) for trees, 56/65 (86%) for mold, 52/61 (85%) for grass, 45/55 (82%) for cat, and 38/47 (81%) for weeds. CONCLUSIONS: (1) Patients have limited ability to correctly predict positive skin tests to aeroallergen. (2) Patients are able to predict negative skin tests with reasonable accuracy. (3) "What do you think you are allergic to?" may be a good screening question for patients with asthma and rhinitis. PMID- 11101181 TI - Allergy to macadamia nut. AB - BACKGROUND: Nuts are one of the most common foods causing allergic reactions in children and adults although an allergic reaction to macadamia nut has been reported only once previously. OBJECTIVE: To report a case of macadamia nut induced allergic reaction. METHODS: Skin prick test with macadamia nut was performed in a private practice office. RESULTS: The results demonstrated type-I hypersensitivity reaction with macadamia nut. CONCLUSION: Macadamia nut ingestion can cause immediate-type allergic reactions. PMID- 11101182 TI - Exercise-induced laryngochalasia: an imitator of exercise-induced bronchospasm. AB - BACKGROUND: Patients with exercise-induced laryngochalasia present with dyspnea and stridor during exercise. Symptoms are due to a subtotal occlusion of the larynx resulting from mucosal edema from the aryepiglottic folds being drawn into the endolarynx. METHODS: We report on three patients with exercise-induced bronchospasm, refractory to standard therapy. RESULTS: Spirometry with flow volume loops revealed truncation of the inspiratory limb. Abnormal movement of the arytenoid region was visualized on laryngoscopy. A diagnosis of exercise induced laryngochalasia was made. CONCLUSIONS: Evaluation of laryngeal motion in patients with refractory exercise-induced bronchospasm is important. Surgical correction with laser laryngoplasty is effective in carefully selected cases. PMID- 11101183 TI - The effect of windspeed on pollen and spore counts collected with the Rotorod Sampler and Burkard spore trap. AB - BACKGROUND: A recent review article presented conflicting evidence concerning the relationship between the Rotorod Sampler's particle collection efficiency and windspeed. Conventional wisdom holds that its collection efficiency is not affected by windspeed; experimental data paint a mixed picture. OBJECTIVES: This brief addendum will present additional data concerning the relationship between the Rotorod's collection efficiency and windspeed. This information will be contrasted with similar data for the Burkard spore trap. DATA SOURCE: Data obtained from an obscure technical report published by Ogden et al. in 1969 will be assessed. RESULTS: The Rotorod's collection efficiency appeared to be greatest at moderate windspeeds; lower efficiencies occurred at both lower and higher windspeeds. Maximum changes in collection efficiency ranged from 29% to 39% over the six windspeeds employed. The Burkard exhibited the opposite relationship between collection efficiency and windspeed. Collection efficiency was lowest at moderate windspeeds and higher at both lower and higher windspeeds. Maximum changes in collection efficiency ranged from 38% to 142%. CONCLUSIONS: Four principal conclusions emerged. First, these newly rediscovered data demonstrate that the Rotorod's collection efficiency is affected by windspeed. These results appear to overturn the conventional wisdom which holds that this device is insensitive to wind. Second, the Rotorod is less affected by windspeed than the Burkard. The maximum change in collection efficiency uncovered for the former device was 39%; this represented the minimum maximum change for the latter device. Third, the Rotorod and Burkard are affected by windspeed in fundamentally different ways. The former device's collection efficiency is greatest at moderate windspeeds and lower at both lower and higher windspeeds. The opposite effect occurs with the latter device. Last, the previous two findings may partially explain some of the differences in particle recovery by the two devices noted in the parent publication. Figures and equations presented in this addendum indicate which circumstances are likely to produce these differences. PMID- 11101184 TI - Insulin desensitization with insulin lispro and an insulin pump in a 5-year-old child. AB - BACKGROUND AND OBJECTIVE: Allergic reactions to insulin, though rare, can have serious consequences in children with type I diabetes mellitus. We report a case of insulin allergy in a 5-year-old child in whom insulin desensitization was accomplished using an insulin pump. METHODS AND RESULTS: A 5-year-old girl with insulin-dependent diabetes mellitus (IDDM) developed progressive reactions to insulin and was found to have positive intradermal skin tests to regular and NPH insulin. Addition of oral antihistamine and co-administration of subcutaneous dexamethasone along with the insulin failed to control her symptoms. The patient was therefore hospitalized and desensitized to insulin using an insulin pump and insulin lispro. CONCLUSION: An insulin pump can be used for insulin desensitization in selected children. The slow constant infusion rate provided by the pump may be an ideal means of achieving insulin desensitization. In addition, insulin lispro, a synthetic insulin analog, should be considered for use in insulin-allergic individuals. PMID- 11101185 TI - Fluticasone propionate versus zafirlukast: effect in patients previously receiving inhaled corticosteroid therapy. AB - BACKGROUND: The use of inhaled corticosteroids compared with leukotriene modifying drugs in the treatment of persistent asthma has not been extensively studied. OBJECTIVE: To compare the efficacy and safety of a low dose of fluticasone propionate (FP) and zafirlukast in patients previously maintained on inhaled corticosteroids. METHODS: Patients (> or = 12 years old; FEV1 = 60% to 85% of predicted) with persistent asthma who were previously treated with low doses of triamcinolone acetonide (TAA) 400 to 800 microg/day or beclomethasone dipropionate (BDP) 168 to 336 microg/day were randomized to treatment with FP aerosol 88 microg BID (FP, n = 221) or zafirlukast 20 mg BID (n = 216) over 6 weeks. RESULTS: Treatment with FP significantly increased the mean change at endpoint (the last post-baseline observation) in FEV1 (0.22 L versus 0.03 L, P < .001), morning PEF (17.8 versus 3.1 L/min, P = .004), evening PEF (16.7 versus 2.6 L/min, P = .002), the percentage of symptom-free days (16.2 versus 7.1%, P = .007), and the percentage of rescue-free days (23.4 versus 9.3%, P < .001), and significantly decreased rescue albuterol use (-0.66 puffs/day versus an increase of 0.27 puffs/day, P < .001) and combined symptom scores (-0.13 versus an increase of 0.08, P < .001) compared with zafirlukast. Treatment with FP maintained the percentage of awakening-free nights (-1.0 +/- 1.0); in contrast, treatment with zafirlukast reduced the percentage of awakening-free nights (-9.0 +/- 1.6, P < .001). A clinically meaningful difference (change of > or = 0.5; P < .001) was observed between FP and zafirlukast in the Asthma Quality of Life Questionnaire (AQLQ) global score and for each domain score except activity limitation (change of 0.3, P < .001). Significantly more patients in the zafirlukast group experienced an asthma exacerbation (n = 14) compared with FP treated patients (n = 5, P = .035). Patients in the zafirlukast group were significantly more likely to be withdrawn due to lack of efficacy (P < .001). CONCLUSION: Switching patients from low doses of inhaled corticosteroids to a lower total microgram dose of FP improves pulmonary function, asthma symptoms, and quality of life, while switching to the leukotriene receptor antagonist zafirlukast may result in worsening of asthma control. This was indicated by the significant number of zafirlukast-treated patients who were dropped from the study due to lack of efficacy within 6 weeks of discontinuing inhaled corticosteroids. PMID- 11101186 TI - Efficacy and safety of dry powder fluticasone propionate in children with persistent asthma. AB - BACKGROUND: Flovent Diskus is a powder formulation of the inhaled corticosteroid fluticasone propionate (FP) delivered via a breath-actuated, multidose inhaler. OBJECTIVE: To determine the efficacy and safety of dry powder FP administered once or twice daily (200 microg per day) to children with persistent asthma. METHODS: Twelve-week, randomized, double-blind, placebo-controlled, multicenter trial with a 52-week, open-label extension. Children aged 4 to 11 were required to have pulmonary function 50% to 85% of predicted values. The population was stratified for baseline therapy (inhaled corticosteroid/cromolyn or bronchodilators only). After a 2-week placebo run-in, 242 patients received dry powder FP 200 microg each morning, dry powder FP 100 microg BID, or placebo for 12 weeks; 192 were rerandomized to the QD or BID regimen for an additional 52 weeks of open-label treatment. Primary endpoints were mean changes in FEV1 and morning PEF recorded at clinic visits. RESULTS: Both dry powder FP regimens significantly improved FEV1, evening PEF, and asthma symptoms at the double-blind phase endpoint (P < or = .017 compared with placebo). The BID regimen also significantly improved morning PEF and nighttime awakenings due to asthma (P < or = .005). Among patients previously treated with inhaled corticosteroids/cromolyn, improvements observed with the QD and BID regimens were similar. Patients switched from BID to open-label QD treatment showed additional improvements at week 52 generally comparable to patients who received the BID regimen during both phases. Fluticasone propionate was well tolerated for up to 64 weeks with few reports of drug-related adverse events or morning plasma cortisol abnormalities. CONCLUSIONS: Once daily dosing of dry powder FP 200 microg is an effective and convenient alternative for children whose asthma is controlled with a more frequent dosing regimen of inhaled corticosteroids. PMID- 11101187 TI - Measurement of children's asthma medication adherence by self report, mother report, canister weight, and Doser CT. AB - BACKGROUND: Accurate assessment of medication adherence has been difficult to achieve but is essential to drug evaluation in clinical trials and improved outcomes in clinical care. OBJECTIVE: This study was conducted to compare four adherence assessment methods: child report, mother report, canister weight, and electronic measurements of metered dose inhaler (MDI) actuation. METHODS: Participants included 27 children with mild-to-moderate asthma who were followed prospectively for 6 months. All patients used an MDI equipped with an electronic Doser attached to their inhaled steroid. At each 2-month follow-up visit, Doser and canister weight data were recorded, while child and mother were interviewed separately regarding medication use. RESULTS: Children and mothers reported, on average, over 80% adherence with the prescribed inhaled steroid. Canister weight revealed, on average, adherence of 69%, significantly lower than self-report. When adherence recorded by the electronic Doser was truncated to no more than 100% of prescribed daily use, average adherence was 50%. Older children and adolescents, nonwhite children, and those from poorer functioning families were least adherent. CONCLUSIONS: Electronic adherence monitoring was significantly more accurate than self-report or canister weight measures. Such accuracy is an essential prerequisite to increasing understanding of the treatment, setting, and patient factors that influence adherence, and to the consequent design of effective intervention strategies. PMID- 11101188 TI - Counterpoint to the use of systemic corticosteroids in the Stevens-Johnson syndrome. PMID- 11101189 TI - Counterpoint to the use of systemic corticosteroids in the Stevens-Johnson syndrome. PMID- 11101190 TI - Studies of nasal sprays for rhinitis. PMID- 11101191 TI - Shortage of IV immunoglobulin. PMID- 11101192 TI - T-type calcium channel blockade in the management of chronic ischemic heart disease. AB - The T-type calcium channel offers a new therapeutic target for treatment of patients with cardiovascular disease. Mibefradil, a T channel blocker, produces heart rate slowing and coronary vasodilatation but without the negative inotropic effect commonly seen when L-type channel blockers are used. The present study shows Mibefradil prevents ischemic episodes that are and are not preceded by an increase in heart rate. Although Mibefradil has been withdrawn because of multiple drug interactions, new T-type calcium channel blockers are under development. PMID- 11101193 TI - Effect of vanadate and insulin on glucose transport in isolated adult rat cardiomyocytes. AB - It is now widely accepted that insulin stimulation of glucose uptake by muscle cells is due to the activation of protein kinase B, leading to the recruitment of glucose transporter proteins from an intracellular compartment to the plasma membrane. Vanadate is a protein tyrosine phosphatase (PTP) inhibitor and a known insulin mimetic agent. Vanadate causes an increase of glucose transport in various tissues, but the mechanism of stimulation is not clearly understood. Hence in the present study, we have compared the mechanism of 2-deoxy-D-glucose transport induced by vanadate and insulin in isolated rat cardiomyocytes. Vanadate stimulated deoxyglucose transport in a time- and concentration-dependent manner. Insulin (100 nM) and vanadate (5 mM) stimulated 2-deoxy-D-glucose transport on an average by 3- and 2-fold respectively over basal values. The stimulation of glucose transport was accompanied by an activation of protein kinase B (PKB). This study also revealed that the activation of PKB and stimulation of 2-deoxyglucose uptake by vanadate and insulin are inhibited by treatment with wortmannin, a specific inhibitor of phoshatidylinositol 3-kinase (PI 3-kinase). Hence, we conclude that both insulin and vanadate follow the same signalling pathway downstream of PI 3-kinase to stimulate 2-deoxy-D-glucose transport. PMID- 11101194 TI - Atrial natriuretic peptide reduces the severe consequences of coronary artery occlusion in anaesthetized dogs. AB - The aim of the present study was to examine the effects of atrial natriuretic peptide (ANP) on the responses to coronary artery occlusion. In chloralose urethane anaesthetised mongrel dogs either saline (controls) or human synthetic ANP was infused intravenously (10 microg kg(-1) + 0.1 microg kg(-1) min(-1)), starting 30 min before and continuing 10 min during a 25 min occlusion of the left anterior descending coronary artery (LAD). ANP infusion resulted in a fall in mean arterial blood pressure (by 17 +/- 2 mmHg, p < 0.05), a transient (max. at 5 min) increase in coronary blood flow (by 24 +/- 5 ml min(-1), p < 0.05), and a reduction in coronary vascular resistance (by 0.27 +/- 0.05 mmHg ml(-1), p < 0.05). When the LAD coronary artery was occluded, there was a less marked elevation in left ventricular end-diastolic pressure (LVEDP) in the ANP-treated dogs than in the controls (9.0 +/- 0.9 versus 12.2 +/- 0.8 mmHg, p < 0.05). Compared to the controls, ANP reduced the number of ventricular premature beats (VPBs, 26 +/- 12 versus 416 +/- 87, p < 0.05), the number of episodes of ventricular tachycardia per dogs (VT, 0.7 +/- 0.3 versus 12.4 +/- 4.2, p < 0.05), and the incidences of VT (45% versus 100%, p < 0.05) and ventricular fibrillation (VF 18% versus 57%, p < 0.05) during occlusion. Reperfusion of the ischaemic myocardium at the end of the occlusion period led to VF in all the control dogs (survival from the combined ischaemia-reperfusion insult was therefore 0%), but VF following reperfusion was much less in the dogs given ANP (survival 64%; p < 0.05). The severity of myocardial ischaemia, as assessed from changes in the epicardial ST-segment and the degree of inhomogeneity, was significantly less marked in dogs given ANP. We conclude that ANP protects the myocardium from the consequences of myocardial ischaemia resulting from acute coronary artery occlusion and reperfusion in anaesthetized dogs. PMID- 11101195 TI - Isosorbide-2-mononitrate reduces the consequences of myocardial ischaemia, including arrhythmia severity: implications for preconditioning. AB - The effects of the intracoronary administration of isosorbide-2-mononitrate (ISMN; 3 microg kg(-1) min(-1)), a major metabolite of isosorbide dinitrate, were examined in chloralose-urethane anaesthetized dogs before and during a 25 min, acute occlusion of the left anterior descending coronary artery. The only significant haemodynamic effects of ISMN administration were a slight (-11 +/- 2 mmHg) decrease in arterial blood pressure and a decrease (< 12%) in diastolic coronary vascular resistance. Coronary occlusion in the presence of ISMN led to a markedly reduced incidence and severity of ventricular arrhythmias compared to those in control, saline-infused dogs. There were fewer ectopic beats (62 +/- 35 versus 202 +/- 72; p < 0.05), a lower incidence (25% versus 75%; p < 0.05) and number of episodes (0.7 +/- 0.4 versus 4.3 +/- 2.1; p < 0.05) of ventricular tachycardia and fewer dogs fibrillated during the ischaemic period (17% versus 82%; p < 0.05). More dogs given ISMN survived the combined ischaemia-reperfusion insult (50% versus 0%; p < 0.05). Changes in ST-segment elevation (recorded by epicardial electrodes) and in the degree of inhomogeneity of electrical activation within the ischaemic area were much less pronounced throughout the occlusion period in dogs given ISMN. These results add weight to the hypothesis that the previously reported antiarrhythmic effects of ischaemic preconditioning, and of the intracoronary administration of nicorandil, involve nitric oxide. PMID- 11101196 TI - The cardioprotective effects of a new 1,4-benzothiazepine derivative, JTV519, on ischemia/reperfusion-induced Ca2+ overload in isolated rat hearts. AB - A new 1,4-benzothiazepine derivative, JTV519 (JTV), has strong protective effects against isoproterenol-induced myocardial injury. We investigated the effects of JTV on Ca2+ overload and on functional recovery during ischemia/reperfusion in isolated coronary-perfused rat hearts. After 30 minutes of reperfusion following 30 min of global ischemia, the % recovery of LV developed pressure was improved in a concentration-dependent manner when JTV (0.3-3.0 microM) was administered either 5 min before induction of ischemia or for 5 min at the time of reperfusion only JTV showed a negative inotropic effect only at concentrations above 3.0 microM. In indol-loaded isolated heart preparations, 0.3 microM JTV did not affect the preischemic systolic or diastolic Ca2+ levels of the Ca2+ transient as measured by the ratio of 2-wavelength fluormetry (R405/500). In contrast, it significantly reduced the increase in the ratio in the postischemic reperfusion period (% change of R405/500 from baseline: JTV(-), by 42.7 +/- 3.2%; JTV(+), by 18.4 +/- 9.1%, p < 0.05). In isolated rat ventricular myocytes with a standard patch-clamp method, we further tested the interaction of JTV with the L-type Ca2+ channel (I(Ca)). The % inhibition of the peak current of I(Ca) was 6.2 +/- 0.8% at 0.3 microM (p = n.s.), 22.0 +/- 3.3% at 1.0 microM (p < 0.05), and 59.6 +/- 1.4% at 3.0 microM (p < 0.01). Thus, the marked cardioprotection due to JTV at 0.3 microM may not be solely attributed to its inhibitory effect on the transsarcolemmal Ca2+ influx through I(Ca). In conclusion, JTV519 is a novel pharmacological agent that has been demonstrated for the first time to have clinical potential for the treatment of acute coronary syndrome by its efficacy in administration at the time of reperfusion, by its suppression of reperfusion related intracellular Ca2+ overload with no significant interaction with I(Ca), and by its subsequent ability of strong myocardial protection. PMID- 11101197 TI - Infarct size limitation by bradykinin receptor activation is mediated by the mitochondrial but not by the sarcolemmal K(ATP) channel. AB - Earlier studies have shown that activation of bradykinin B2 receptor triggers protein kinase C (PKC)-mediated cardioprotective mechanism in ischemic preconditioning (PC). In the present study, we examined whether the effector in this B2-receptor triggered pathway of PC is the ATP sensitive potassium (K(ATP)) channel in the mitochondria (mito-K(ATP) channel) or K(ATP) channel in the sarcolemma (sarc-K(ATP) channel). Isolated rabbit hearts were perfused with modified Krebs-Henseleit buffer in a Langendorff mode, and regional myocardial ischemia was induced by occluding a left coronary artery for 30 min and then reperfusing for 2 hours. Infarct size was determined by triphenyltetrazolium chloride staining and expressed as a percentage of area at risk (% IS/AR). Infusion of bradykinin (500 nmol/L) for 15 min prior to ischemia significantly reduced % IS/AR from 37.4 +/- 2.9 (SE) of the untreated controls to 12.0 +/- 3.3%. This protective effect of bradykinin was completely abolished by coinfusion of 5-hydroxydecanoate (5-HD, 50 micromol/L), a selective mito-K(ATP) channel blocker (% IS/AR = 44.2 +/- 6.4). In contrast, a high dose of HMR1098 (20 micromol/L), which is a newly developed sarc-K(ATP) channel selective blocker with IC50 of 0.6 micromol/L, failed to modify the infarct size limitation by preischemic infusion of bradykinin (% IS/AR = 11.7 +/- 3.4). Neither 5-HD nor HMR1098 alone modified infarct size (% IS/AR = 37.8 +/- 3.8 and 35.1 +/- 6.2, respectively). These results suggest that opening of the mito-K(ATP) channel but not the sarc-K(ATP) channel is involved in infarct size limitation by a mechanism triggered by bradykinin B2 receptor activation. PMID- 11101198 TI - The effect of mibefradil on ischemic episodes with and without increase in heart rate. AB - Myocardial ischemia during daily life can be induced by increased demand and by increased coronary tone. The purpose of this study was to assess the mechanism of action of mibefradil, a new T-channel calcium blocker that is a vasodilator with negative chronotropic properties. Included in this study were 114 patients with chronic stable angina pectoris and ischemic episodes during baseline 48-hour ambulatory ECG monitoring (AEM). After a placebo run-in period patients received 50 mg, 100 mg, or 150 mg of mibefradil per day and repeat 48 hours AEM was performed. Ischemic episodes were divided into 2 categories: Type I is those in which an increase in heart rate > 10% preceded the development of 1 mm ST depression; Type II is those with < or = 10% increase in heart rate. Of the 625 ischemic episodes recorded at baseline, 83% were Type I and 17% were Type II. At 50 mg mibefradil dose, there was a significant decrease in the number of Type I ischemic episodes but not of Type II. At doses of 100 mg and 150 mg/day, there was a significant decrease in frequency of both types of ischemic episodes. At a low dose of 50 mg/day, mibefradil reduces ischemia predominantly by preventing an increase in heart rate, while at higher doses of 100 mg and 150 mg/day, it also acted as a vasodilator suppressing episodes associated with increased coronary tone. PMID- 11101199 TI - Hemodynamic, antiischemic, and neurohumoral effects of tedisamil and atenolol in patients with coronary artery disease. AB - Clinical drawbacks of beta-blocker treatment in stable angina have motivated researchers to provide alternative heart-rate-lowering agents, such as tedisamil, which additionally exerts antiischemic and antiarrhythmic effects by blockade of cellular repolarizing K+ currents. Forty-eight patients with stable angina pectoris were investigated (doubleblind, randomized, parallel grouped) comparing the hemodynamic, antiischemic, metabolic and neurohumoral effects of tedisamil 100 mg b.i.d. and atenolol 50 mg b.i.d. after a single dose and over 6 days of treatment. Tedisamil and atenolol produced a decrease in heart rate both at rest [day 1:-13.6 versus - 15.4 bpm; day 6: - 14.8 versus - 22.2 bpm, resp.; p > 0.05] and exercise [day 1: - 9.1 versus - 18.3 bpm; p = 0.001; day 6: - 12.0 versus - 24.8 bpm, resp.; p = 0.001], while anginal threshold increased. Cardiac output decreased with tedisamil and atenolol at rest [day 1: -1.01 versus -1.19 l/min; p > 0.05; day 6: - 0.86 versus - 1.10 l/min, resp.; p > 0.05] and exercise [day 1: 0.82 versus - 1.28 l/min; p > 0.05; day 6: - 0.65 versus - 2.68 l/min, resp.; p = 0.03], while stroke volume remained unchanged. Right atrial pressure changed during exercise only: it decreased with tedisamil (-1.7 mmHg) and increased with atenolol (+ 3.7 mmHg) (p < .001). Mean pulmonary capillary wedge pressures decreased both at rest (- 0.5 mmHg) and exercise (- 6.9 mmHg) in the tedisamil group but tended to increase with atenolol on day 6 of treatment [rest: + 1.7; exercise: + 3.7 mmHg) (p = 0.03). Arterial pressure decreased under atenolol treatment only. Exercise-induced plasma norepinephrine levels were reduced by tedisamil (- 93 pg/ml) but elevated by atenolol (+ 172 pg/ml) (p = 0.001). As compared to atenolol, tedisamil produced a prolongation of QTc interval [+ 31 versus 8 ms] at initial values of 0.408 +/- 0.018 s with PQ and QRS remaining unaltered. In patients with stable angina, tedisamil (100 mg b.i.d.) as compared to atenolol (50 mg b.i.d.) generated similar hemodynamic, neurohumoral and antiischemic effects. The antiischemic efficacy of tedisamil, as measured by ST segment depression and angina threshold, was comparable to that of atenolol. PMID- 11101200 TI - Calcium channel blockers reduce blood pressure in part by inhibiting vascular smooth muscle carbonic anhydrase I. AB - Calcium channel blockers are a group of drugs used for the treatment of hypertension. Carbonic anhydrase (CA) I detected in vascular smooth muscle and in other cells in the organism has a major role in the acid-base balance and in vascular processes. Our previous work has proven that verapamil inhibits CA activity by a direct mechanism of action. Starting from our results in this article we studied in vitro and in vivo the effect of calcium channel blockers (verapamil and amlodipine) on erythrocyte CA I, on vascular smooth muscles CA I, and on arterial blood pressure values in human and in animals. Our in vitro and in vivo results have proved that verapamil and amlodipine are strong CA I inhibitors both in human erythrocytes and also in vascular smooth muscles in animals. In humans, calcium channel blockers studied here progressively reduce arterial blood pressure in hypertensive subjects, in parallel with progressive lowering of erythrocyte CA I activity in the normal range in normotensive subjects. From our point of view verapamil and amlodipine possess a dual mechanism of action: the first well-known action consists of their action on calcium channels. The second mechanism, suggested by us, directly acts on the vascular smooth muscle CA I isozyme, so that its inhibition should ensure an adequate pH for calcium ions transport through the channels, having as result vasodilation. This double mechanism could explain the hypotensive effect of verapamil and amlodipine, with a mechanism that partially dependent on CA I inhibition. PMID- 11101201 TI - Effect of long-term losartan administration on renal haemodynamics and function in hypertensive patients. AB - In this study the efficacy and safety of long-term losartan administration on renal haemodynamics were evaluated in mild to moderate hypertension. After a run in period with placebo, 18 hypertensives without renal or cardiovascular disease were allocated to losartan (50 mg/die for one year) treatment. Renal haemodynamic measurements included renal plasma flow (ERPF) and glomerular filtration rate (GFR) by standardized radioisotope study. Effective renal blood flow (ERBF), filtration fraction (FF), and renal vascular resistance (RVR) were also calculated. Blood pressure was evaluated monthly, whereas renal haemodynamics and function were detected at baseline and after 6 and 12 months of losartan administration. Losartan induced a significant (p < 0.001) decrease in SBP, DBP, and MBP versus baseline values both at 6 months and at 12 months. In addition a significant decrease in RVR (p < 0.001) and in FF (p < 0.05) was also seen. In addition RVR values at 1 year of treatment were higher than their values at 6 months, but this difference was not significant. Our data indicated that long term control in blood pressure induced by losartan administration was associated with a maintained renal function after 6 months of treatment, but these favourable effects were attenuated after 1 year of treatment. PMID- 11101202 TI - Inhibition of nitric oxide synthesis induces coronary vascular remodeling and cardiac hypertrophy associated with the activation of p70 S6 kinase in rats. AB - Chronic inhibition of nitric oxide (NO) synthesis is reported to induce the thickening of coronary artery walls and cardiac hypertrophy in vivo via angiotensin II receptors. Increased protein synthesis is the main feature of these structural changes. Activation of 70 kD S6 kinase (p70S6K) phosphorylates the 40S ribosomal protein S6 that regulates protein synthesis. We examined the role of p70S6K in the vascular and myocardial structural changes induced by the chronic inhibition of NO synthesis. The following 5 groups were studied: untreated Wister-Kyoto rats, those treated with an inhibitor of NO synthase, Nomega-nitro-L-arginine methyl ester (L-NAME), those treated with L-NAME and an angiotensin I converting enzyme inhibitor (imidapril), those treated with L-NAME and hydralazine, and those treated with L-NAME and an inhibitor of p70S6K (rapamycin). After 8 weeks, wall-to-lumen ratio in myocardium and cardiomyocyte cross-sectional areas were quantified. L-NAME increased systolic blood pressure, wall-to-lumen ratio, and cardiomyocyte cross-sectional area compared with control animals. Imidapril or rapamycin, but not hydralazine, markedly reduced these structural changes. L-NAME increased p70S6K activity in myocardium compared with control rats. Imidapril or rapamycin prevented the activation of p70S6K activity in myocardium induced by L-NAME. These results suggest that activation of p70S6K plays an important role in coronary vascular remodeling and cardiac hypertrophy induced by the chronic inhibition of nitric oxide synthesis in vivo. PMID- 11101203 TI - Prediction of the effect of bisoprolol in dilated cardiomyopathy. AB - Survival improvement by beta-blocker treatment in patients with chronic heart failure appears to be related to the intermediate-term changes in left ventricular function. The therapeutic potential of beta blockade might be increased by early identification of patients in whom left ventricular function would deteriorate. We aimed at predicting the intermediate-term effect of bisoprolol on left ventricular systolic and diastolic function in patients with dilated cardiomyopathy. Twenty-five patients with symptomatic chronic heart failure treated with bisoprolol were investigated. As a background, tailored therapy with digitalis, diuretics and vasodilators was given. Prediction of the 6 month (intermediate-term) effect of bisoprolol was investigated, using baseline values and short-term (1-month) changes of simple, noninvasive parameters obtained at rest and during maximal exercise. Multivariate analysis resulted in reliable predictions, there was close correlation between the observed and predicted changes of left atrial filling pressure (R = 0.87) and left ventricular ejection fraction (R = 0.74). The baseline value of left ventricular ejection fraction, short-term changes of the pulse amplitude and the double product proved independent predictors of intermediate-term changes of left ventricular ejection fraction. The baseline value of mean pulmonary capillary wedge pressure, heart rate, and increase in heart rate during maximal exercise were predictors of the intermediate-term changes in mean pulmonary capillary wedge pressure. In dilated cardiomyopathy, the intermediate-term effects of bisoprolol on left ventricular ejection fraction and mean pulmonary capillary wedge pressure can be predicted reliably by simple noninvasive variables in the early treatment phase. PMID- 11101204 TI - Traumatic brain injury reduces hippocampal alpha7 nicotinic cholinergic receptor binding. AB - Changes in the expression of central nervous system (CNS) neurotransmitter receptors may contribute to behavioral and physiological deficits that occur following traumatic brain injury (TBI). Studies investigating the neurochemical basis for the protracted cognitive dysfunction that follows TBI have focused in part on cholinergic mechanisms. The present study compared the effects of mild and moderate cortical contusion injury (CCI) on the density of cholinergic receptor subtypes, NMDA-type glutamate receptors, and calcium channel expression. Quantitative autoradiography was used to determine the effects of CCI on receptor expression, 48 h following injury. The most robust and consistent change in receptor binding was in the density of alpha7 nicotinic receptors as determined by alpha-[125I]-bungarotoxin (BTX) binding. Bilateral deficits in BTX binding were present following both mild and moderate levels of injury. In contrast, changes in the density of alpha3/alpha4 nAChr's, muscarinic AChr's, NMDA-type glutamate receptors, and L-type calcium channel expression were more regionally restricted and lower in magnitude, as compared to changes in BTX binding. The high calcium permeability of the alpha7 nAChr may be related to the extensive decrease in BTX binding that occurs following TBI. PMID- 11101205 TI - Neurological recovery from closed head injury is impaired in diabetic rats. AB - Diabetes mellitus is a metabolic disorder associated with central nervous system impairments. Recent studies implicate oxidative stress mediated by reactive oxygen species (ROS) in the pathogenesis of diabetic complications. ROS have been shown to play role in the pathophysiology of brain injury. In the present study, closed head injury (CHI) was induced in diabetic rats to test the hypothesis that chronic oxidative stress exacerbates brain damage following CHI. Neurological recovery, edema, levels of low molecular weight antioxidants (LMWA), and markers of lipid peroxidation were determined at different intervals after injury. Diabetic rats (4 weeks after induction with streptozotocin) were subjected to CHI. Brain edema (percent water) and clinical status (neurological severity score) were assessed during 7 days. Brain LMWA were determined using cyclic voltammetry (CV) and HPLC-EC. In addition, conjugated dienes and thiobarbituric acid reactive substances (TBARS) were measured. Diabetic-CHI rats exhibited a lower rate of recovery and greater and more sustained edema (p < 0.01), as compared with the controls. At all times diabetic rats had higher levels of TBARS and conjugated dienes and lower concentrations of LMWA, and of vitamins C and E, suggesting chronic oxidative stress. At 5 min of CHI, the amounts of LMWA in control-CHI brains decreased (approximately 50%, p < 0.01) and returned to normal by 48 h and 7 days. In the diabetic-CHI brain only one class of LMWA slightly declined but remained low for 7 days. The present results support the hypothesis that diabetic rats are under chronic oxidative stress, and suffer greater neurological dysfunction, associated with further lipid peroxidation following CHI. PMID- 11101206 TI - Postinjury treatment with magnesium chloride attenuates cortical damage after traumatic brain injury in rats. AB - The neuroprotective effect of magnesium chloride (MgCl2), a compound previously demonstrated to improve behavioral and neurochemical outcome in several models of experimental brain injury, was evaluated in the present study. Male Sprague Dawley rats were anesthetized and subjected to lateral fluid-percussion brain injury of moderate severity (2.5-2.8 atm). A cannula was implanted in the left femoral vein and at 1 h following injury, animals randomly received a 15 min i.v. infusion of either MgCl2 (125 micromol/rat) or saline. A second group of animals received anesthesia, surgery, and either MgCl2 or vehicle to serve as uninjured (sham) controls. Two weeks following brain injury, animals were sacrificed, brains removed, and coronal sections were taken for quantitative analysis of cortical lesion volume and hippocampal CA3 cell counts. Traumatic brain injury resulted in a lesion in the ipsilateral cortex and loss of pyramidal neurons in the CA3 region of the hippocampus in vehicle-treated animals (p < 0.01 vs. uninjured animals). Administration of MgCl2 significantly reduced the injury induced damage in the cortex (p < 0.01) but did not alter posttraumatic cell loss in the CA3 region of the ipsilateral hippocampus. The present study demonstrates that, in addition to its beneficial effects on behavioral outcome, MgCl2 treatment attenuates cortical histological damage when administered following traumatic brain injury. PMID- 11101207 TI - Upregulation of neuronal amyloid precursor protein (APP) and APP mRNA following magnesium sulphate (MgSO4) therapy in traumatic brain injury. AB - The aim of this study was to assess and quantitate topographically the effects of posttraumatic intravenous magnesium sulphate (MgSO4) on neuronal perikaryal APP antigen and messenger RNA (mRNA) expression in sheep brains 2 h after a controlled focal head impact. The percentage brain area with APP immunoreactive neuronal perikarya was 71, 56, 27.5 and 5.5%, respectively, in MgSO4-treated head injured animals, head-injured animals without any treatment, MgSO4 treated nonimpacted animals, and nontreated nonimpacted control sheep. Although there was no statistically significant difference in APP immunoreactive neuronal perikarya in the MgSO4-treated HI group (mean 71%) compared to the HI group without any treatment (mean 56%), northern analysis showed that there was a 2.3-+/-0.2-fold increase in APP mRNA in the thalamus of treated impacted animals compared to untreated impacted animals (p < 0.005). However, MgSO4 treated nonimpacted control animals also showed a 1.6-+/-0.1-fold increase in APP mRNA compared to untreated nonimpacted controls (p < 0.005). MgSO4 therapy results in upregulation of neuronal APP mRNA and APP expression that is quantitatively greater following a focal head impact. PMID- 11101208 TI - Enoxaparin reduces brain edema, cerebral lesions, and improves motor and cognitive impairments induced by a traumatic brain injury in rats. AB - Traumatic brain injury (TBI) is often accompanied by secondary ischemia due, in part, to edema-induced blood vessel compression. Enoxaparin, a low-molecular weight heparin, which is efficacious in models of myocardial and brain ischemia was studied in lateral fluid percussion-induced TBI in rats. Enoxaparin was administered 2 h post-TBI at 0.5 mg/kg i.v. followed by 4 x 0.5, 4 x 1, or 4 x 2 mg/kg s.c. over 30 h. Brain edema was measured in the hippocampus, temporal cortex and parietal cortex. Edema was reduced by enoxaparin (0.5 + 4 x 0.5 mg/kg) in the hippocampus (-53%, p = 0.07) and the parietal cortex (-39%, ns). At 0.5 + 4 x 1 mg/kg edema was reduced in the hippocampus (-63%, p < 0.05) and the parietal cortex (-47%, p = 0.06). At 0.5 + 4 x 2 mg/kg, the reduction was more important in the hippocampus (-69%, p < 0.01) and in the parietal cortex (-50%, p < 0.05). No reduction was seen in the temporal cortex. The lesion size was reduced by enoxaparin at 0.5 + 4 x 1 mg/kg (-50%, p < 0.05), and at 0.5 + 4 x 2 mg/kg (-35%, ns). The neurological deficit evaluated with a 9-point scale was also improved with enoxaparin at 0.5 + 4 x 1 mg/kg 1 week post-TBI (p < 0.05). The cognitive impairment evaluated with a Lashley maze task was improved with enoxaparin (0.5 + 4 x 1 mg/kg) from 48 h (p < 0.05) to 2 weeks post-TBI (p < 0.01). Our results demonstrate for the first time that enoxaparin significantly reduces the brain contusion and edema, and improves the functional outcomes induced by a TBI. Therefore, enoxaparin could be a candidate drug to treat acute brain-injured patients. PMID- 11101209 TI - Focal brain injury, FGF-2 and the adverse effects of excessive motor demand on cortical and nigral degeneration: marked protection by delayed intermittent exposure to halothane. AB - The neuroprotective potential of halothane anesthesia was investigated following unilateral electrolytic lesions to the forelimb representation area of the sensorimotor cortex (FL-SMC). Previously, it was found that the FL-SMC lesion increases substantially in size when the intact forelimb is immobilized with a plaster of paris cast for the first 7 days postlesion, which forces extreme overuse of the impaired forelimb during a time when nonlethally damaged tissue is vulnerable to behavioral demand. Initially, the purpose of this study was to investigate whether intracisternal infusion of basic fibroblast growth factor (bFGF or FGF-2), a potent neurotrophic factor that has been shown to have neuroprotective and plasticity promoting properties in focal stroke and other injury models, could prevent this use-dependent exaggeration of injury. Although intracisternal bFGF (starting 24 h after surgery, twice per week) was not found to produce significant neuroprotective or behavioral effects, the brief exposure to halothane anesthesia (15-20 min) during bFGF or vehicle administration was found to prevent expansion of the lesion size, and to reduce delayed loss of neurons in the substantia nigra pars reticulata (SNr). The data have implications for investigations of the effects of neurotrophic factor in vivo, and other investigations requiring brief, intermittent halothane anesthesia. PMID- 11101210 TI - Efficacy of a new neuroprotective agent, gacyclidine, in a model of rat spinal cord injury. AB - Prevention of the immediate excitotoxic phase occurring in response to spinal cord injury (SCI) is a major issue to reduce the neuronal damage responsible for any ensuing motor deficits. The present study evaluated the neuroprotective efficacy of three noncompetitive NMDA receptor antagonists: Gacyclidine (GK-11), a new compound, Dizocilpine (MK-801), and Cerestat (CNS-1102) in a rat spinal cord contusion model. To mimic human SCI, a standardized model of rat spinal cord closed contusion in which animals spontaneously and progressively recover from the induced paraplegia was employed. Such model, characterized by a slow recovery of hindlimb locomotor function enables easy quantification of the neuroprotection at both the behavioral and cellular level. The animals were treated intravenously with the respective drugs 10 min after the spinal contusion. The dose range study suggested that 1 mg/kg of Gacyclidine was the most effective dose to promote functional recovery in reducing by half the time needed to reach full locomotor recovery. Racemate and enantiomers of Gacyclidine showed similar neuroprotective effects, but treatment with the enantiomers were not as efficacious in promoting full functional recovery. Similarly, a prolonged treatment with the racemate was not as efficious as a single dose, suggesting that a prolonged blockade of the amino-excitatory neurotransmission may be deleterious. Finally, Dizocilpine and Cerestat treatments induced only a partial and delayed neuroprotective effect compared to Gacyclidine. Neuroprotection characterized by a reduction of the cystic cavity and of the astrogliosis was observed with all treatments. As Gacyclidine is already in clinical trials, the present findings suggest the premise that it is a promising agent for limiting the initial neuronal damage induced by CNS trauma leading to better functional recovery. PMID- 11101211 TI - Sequence of cellular changes following localized axotomy to cortical neurons in glia-free culture. AB - This study utilizes an in vitro model of localized physical injury to axons to examine the specific responses of neocortical neurons to trauma in isolation from glia cell types. The neuronal response to axotomy was closely linked with nerve cell maturity. Cultures grown for 14 days in vitro showed no accumulation of either neurofilaments or, the axonal sprouting marker, GAP43, within injured axons following injury. In older cultures (21 days in vitro), however, temporally distinct axonal changes were evident following transection of axonal bundles. At 12 h postinjury, these included extensive accumulation of neurofilaments into ring-like structures within the cut stumps and an increase in punctate GAP43 labelling throughout the damaged area. At 24 h postinjury, bulb-like accumulations of neurofilaments were also present within the transected axons. Finally at 3 days postinjury, distinct GAP43 and neurofilament immunolabeled axons, and GAP43 immunopositive growth cones, emanated from the cut stump. These results indicate that injured axons of mature neurons undergo a defined series of reactive changes, ultimately culminating in a sprouting response, which occur independently of the presence or effects of glial cell populations. PMID- 11101212 TI - Oxygen-independent real-time monitoring of distinct biphasic glutamate release using dialysis electrode in rat striatum during anoxia: in vivo evaluation of glutamate release and reversed uptake. AB - Using a dialysis electrode, previous studies showed a clear biphasic release of glutamate during anoxia and ischemia. In this study, we examined two hypotheses: (1) glutamate is of vesicular origin and its release is thus Ca2+- and ATP dependent in the first phase, while in the second phase glutamate is derived primarily from the metabolic pool, and (2) reversed glutamate uptake, due to electrogenic stoichiometry, produces the second phase during anoxic insult in the rat brain. A dialysis electrode continuously perfused with glutamate oxidase and ferrocene-conjugated bovine serum albumin (BSA) optimized the time resolution of monitoring, allowing quantitative oxygen-independent, real-time measurement of the extracellular glutamate concentration ([Glu]e) during anoxia. [Glu]e dynamics were analyzed during anoxia by combining the dialysis electrode with focal microinjection of substances inducing glutamate release. Following anoxia in the rat brain, a sharp and rapid [Glu]e elevation took place (first phase). The [Glu]e elevation then shifted, continuing a gently sloping rise throughout the anoxic period (second phase). This first phase disappeared with intracranial administration of either Co2+ or omega-conotoxin. The second phase rise increased with focal microinjection of KCl (300 mM, 1 microL) and decreased with NaCl (300 mM, 1 microL), ultimately reaching a plateau in both cases. Preloading with a novel glutamate transporter inhibitor (tPDC) decreased both the first and second phases of [Glu]e elevation. This dialysis electrode system provides data supporting in vivo evidence that the peak of the first phase of [Glu]e elevation is derived from the "neurotransmitter pool," while the second phase is derived from the neuronal and glial "metabolic pool," which is, at least, partly related to a "reversed uptake" mechanism in the anoxic rat brain. PMID- 11101213 TI - Lipari-Szabo mapping: A graphical approach to Lipari-Szabo analysis of NMR relaxation data using reduced spectral density mapping. AB - In this paper, we explore connections between the Lipari-Szabo formalism and reduced spectral density mapping, and show how spectral density estimates can be associated with Lipari-Szabo parameters via a simple geometric construction which we call Lipari-Szabo mapping. This relationship can be used to estimate Lipari Szabo parameters from spectral density estimates without the need for nonlinear optimization, and to perform 'model selection' in a graphical manner. The Lipari Szabo map also provides insight into the Lipari-Szabo model, and allows us to determine when a given set of experimental spectral densities are inconsistent with the Lipari-Szabo formalism. Practical applications of Lipari-Szabo mapping in conjunction with more traditional analysis methods are discussed. PMID- 11101214 TI - Measurement of one-bond 15N-13C' dipolar couplings in medium sized proteins. AB - A simple and accurate method is described for measurement of 1J(C'N) splittings in isotopically enriched proteins. The method is of the quantitative J correlation type, and the 1J(C'N) splitting is derived from the relative intensity in two 3D TROSY-HNCO spectra with 1J(C'N) dephasing intervals of approximately 1/(2 1J(C'N)) (reference intensity) and approximately 1/1J(C'N) (residual intensity). If the two spectra are recorded under identical conditions and with the same number of scans, the random error in the 1J(C'N) value extracted in this manner is inversely related to the signal-to-noise (S/N) in the reference spectrum. A S/N of 30:1 in the reference spectrum yields random errors of less than 0.2 Hz in the extracted 1J(C'N) value. Dipolar couplings obtained from the difference in 1J(C'N) splitting in the isotropic and liquid crystalline phase for the C-terminal domain of calmodulin are in excellent agreement with its 1.68-A crystal structure, but agree considerably less with the 2.2-A structure. PMID- 11101215 TI - Backbone dynamics of free barnase and its complex with barstar determined by 15N NMR relaxation study. AB - Backbone dynamics of uniformly 15N-labeled free barnase and its complex with unlabelled barstar have been studied at 40 degrees C, pH 6.6, using 15N relaxation data obtained from proton-detected 2D [1H]-15N NMR spectroscopy. 15N spin-lattice relaxation rate constants (R1), spin-spin relaxation rate constants (R2), and steady-state heteronuclear [1H]-15N NOEs have been measured at a magnetic field strength of 14.1 Tesla for 91 residues of free barnase and for 90 residues out of a total of 106 in the complex (excluding three prolines and the N terminal residue) backbone amide 15N sites of barnase. The primary relaxation data for both the cases have been analyzed in the framework of the model-free formalism using both isotropic and axially symmetric models of the rotational diffusion tensor. As per the latter, the overall rotational correlation times (tau(m)) are 5.0 and 9.5 ns for the free and complexed barnase, respectively. The average order parameter is found to be 0.80 for free barnase and 0.86 for the complex. However, the changes are not uniform along the backbone and for about 5 residues near the binding interface there is actually a significant decrease in the order parameters on complex formation. These residues are not involved in the actual binding. For the residues where the order parameter increases, the magnitudes vary significantly. It is observed that the complex has much less internal mobility, compared to free barnase. From the changes in the order parameters, the entropic contribution of NH bond vector motion to the free energy of complex formation has been calculated. It is apparent that these motion's cause significant unfavorable contributions and therefore must be compensated by many other favorable contributions to effect tight complex formation. The observed variations in the motion and their different locations with regard to the binding interface may have important implications for remote effects and regulation of the enzyme action. PMID- 11101216 TI - Refined solution structure of the dimeric N-terminal HHCC domain of HIV-2 integrase. AB - The solution structure of the dimeric N-terminal domain of HIV-2 integrase (residues 1-55, named IN(1-55)) has been determined using NMR spectroscopy. The structure of the monomer, which was already reported previously [Eijkelenboom et al. (1997) Curr. Biol., 7, 739-746], consists of four alpha-helices and is well defined. Helices alpha1, alpha2 and alpha3 form a three-helix bundle that is stabilized by zinc binding to His12, His16, Cys40 and Cys43. The dimer interface is formed by the N-terminal tail and the first half of helix alpha3. The orientation of the two monomeric units with respect to each other shows considerable variation. 15N relaxation studies have been used to characterize the nature of the intermonomeric disorder. Comparison of the dimer interface with that of the well-defined dimer interface of HIV-1 IN(1-55) shows that the latter is stabilized by additional hydrophobic interactions and a potential salt bridge. Similar interactions cannot be formed in HIV-2 IN(1-55) [Cai et al. (1997) Nat. Struct. Biol., 4, 567-577], where the corresponding residues are positively charged and neutral ones. PMID- 11101217 TI - Sequence-specific NMR assignment of proteins by global fragment mapping with the program MAPPER. AB - A new program, MAPPER, for semiautomatic sequence-specific NMR assignment in proteins is introduced. The program uses an input of short fragments of sequentially neighboring residues, which have been assembled based on sequential NMR connectivities and for which either the 13C(alpha) and 13C(beta) chemical shifts or data on the amino acid type from other sources are known. MAPPER then performs an exhaustive search for self-consistent simultaneous mappings of all these fragments onto the protein sequence. Compared to using only the individual mappings of the spectroscopically connected fragments, the global mapping adds a powerful new constraint, which results in resolving many otherwise intractable ambiguities. In an initial application, virtually complete sequence-specific assignments were obtained for a 110 kDa homooctameric protein, 7,8 dihydroneopterin aldolase from Staphylococcus aureus. PMID- 11101218 TI - Selection of side-chain carbons in a high-molecular-weight, hydrophobic peptide using solid-state spectral editing methods. AB - Solid-state spectral editing techniques have been used by others to simplify 13C CPMAS spectra of small organic molecules, synthetic organic polymers, and coals. One approach utilizes experiments such as cross-polarization-with-polarization inversion and cross-polarization-with-depolarization to generate subspectra. This work shows that this particular methodology is also applicable to natural abundance 13C CPMAS NMR studies of high-molecular-weight biopolymers. The editing experiments are demonstrated first with model peptides and then with alpha elastin, a high-molecular-weight peptidyl preparation obtained from the elastic fibers in mammalian tissue. The latter has a predominance of small, nonpolar residues, which is evident in the crowded aliphatic region of typical 13C CPMAS spectra. Spectral editing is particularly useful for simplifying he aliphatic region of the NMR spectrum of this elastin preparation. PMID- 11101219 TI - Production of stable isotope enriched antimicrobial peptides in Escherichia coli: an application to the production of a 15N-enriched fragment of lactoferrin. AB - A method is described for the production of recombinant isotopically enriched peptides in E. coli. Peptides are produced in high yield as fusion proteins with ketosteroid isomerase which form insoluble inclusion bodies. This insoluble form allows easy purification, stabilizes the peptide against degradation and prevents bactericidal activity of the peptide. Cyanogen bromide cleavage released peptide which was conjugated with alkylamines to form lipopeptide. An important advantage of this system is that it allows production of peptides that are toxic to bacteria, which we have demonstrated on a dodecapeptide based on residues 21-31 of human bactericidal protein lactoferrin. PMID- 11101220 TI - Redor in IS1S2 systems. AB - An approach to the determination of the 2-(13)C' chemical shift (CS) tensor orientation in pyrimidine bases via heteronuclear MAS NMR spectroscopy is presented. Considering a dipolar coupled spin 1/2 network of the type S1-I-S2 consisting of directly bonded heteronuclear spins, we have carried out numerical simulations to assess the sensitivity of I-S REDOR spinning sidebands to the Euler angles defining the orientation of the I-S1 and I-S2 dipolar vectors in the I spin CS tensor principal axes system. Our investigations clearly demonstrate the potential of I-S REDOR studies in IS1S2 systems for obtaining with high reliability and accuracy the I spin chemical shift tensor orientation in the molecular frame spanned by the two internuclear vectors I-S1 and I-S2. The significant contribution to the observed REDOR sideband intensities from anti phase operator terms which are present at the start of the data acquisition is illustrated. The procedure for the recording and analysis of the I-S REDOR spectra in IS1S2 systems is presented and the measurement of the 2-(13)C' CS tensor orientation in a polycrystalline sample of [1,3-(15)N2, 2-(13)C] uracil, which is one of the four bases in RNA, is experimentally demonstrated. PMID- 11101221 TI - 13C NMR chemical shifts can predict disulfide bond formation. AB - The presence of disulfide bonds can be detected unambiguously only by X-ray crystallography, and otherwise must be inferred by chemical methods. In this study we demonstrate that 13C NMR chemical shifts are diagnostic of disulfide bond formation, and can discriminate between cysteine in the reduced (free) and oxidized (disulfide bonded) state. A database of cysteine 13C C(alpha) and C(beta) chemical shifts was constructed from the BMRB and Sheffield databases, and published journals. Statistical analysis indicated that the C(beta) shift is extremely sensitive to the redox state, and can predict the disulfide-bonded state. Further, chemical shifts in both states occupy distinct clusters as a function of secondary structure in the C(alpha)/C(beta) chemical shift map. On the basis of these results, we provide simple ground rules for predicting the redox state of cysteines; these rules could be used effectively in NMR structure determination, predicting new folds, and in protein folding studies. PMID- 11101222 TI - Assignment of 1H(N), 15N, 13C(alpha), 13CO and 13C(beta) resonances in a 67 kDa p53 dimer using 4D-TROSY NMR spectroscopy. AB - The p53 tumor suppressor is a transcription factor that plays a crucial role in the activation of genes in response to DNA damage. As a first step towards detailed structural studies of the molecule aimed at understanding its regulation, we have used 4D-TROSY triple resonance NMR spectroscopy to obtain nearly complete 1H(N), 15N, 13C(alpha), 13CO and 13C(beta) resonance assignments of a dimeric form of the protein comprising DNA-binding and oligomerization domains (67 kDa). A simple comparison of 4D spectra recorded on p53 molecules consisting of DNA-binding and oligomerization domains with and without the regulatory domain establishes that both constructs have essentially identical chemical shifts. Although the affinity of p53 for target DNA is decreased in constructs containing the regulatory domain, the chemical shift results reported here suggest that this decrease is not due to specific domain interactions involving the regulatory portion of the molecule, or alternatively, that such interactions require the presence of DNA. PMID- 11101223 TI - 1H, 13C, and 15N assignment of a bleomycin resistance protein in its native form and in a complex with Zn2+ ligated bleomycin. PMID- 11101224 TI - Assignment of the 1H, 15N and 13C resonances of the C-terminal domain of the TolA protein of Escherichia coli, involved in cell envelope integrity. PMID- 11101225 TI - Assignments of the 1H, 13C, and 15N resonances of the 21 kDa Vesl/Homer family protein, Vesl-1S. PMID- 11101226 TI - An electrochemical metalloimmunoassay based on a colloidal gold label. AB - A novel, sensitive electrochemical immunoassay has been developed using a colloidal gold label that, after oxidative gold metal dissolution in an acidic solution, was indirectly determined by anodic stripping voltammetry (ASV) at a single-use carbon-based screen-printed electrode (SPE). The use of disposable electrodes allows for simplified measurement in 35 microL of solution. The method was evaluated for a noncompetitive heterogeneous immunoassay of an immunoglobulin G (IgG) and a concentration as low as 3 x 10(-12) M was determined, which is competitive with colorimetric ELISA or with immunoassays based on fluorescent europium chelate labels. The high performance of the method is related to the sensitive ASV determination of gold(III) at a SPE (detection limit of 5 x 10(-9) M) and to the release of a large number of gold(III) ions from each gold particle anchored on the immunocomplex (e.g., 1.7 x 10(5) gold atoms are theoretically contained in a 18-nm spherical gold particle). PMID- 11101227 TI - Multidimensional information on the chemical composition of single bacterial cells by confocal Raman microspectroscopy. AB - In many biotechnological processes, living microorganisms are used as biocatalysts. Biochemical engineering science is becoming more aware that individual cells of an organism in a process can be fairly inhomogeneous regarding their properties and physiological status. Raman microspectroscopy is a novel approach to characterize such differentiated populations. Cells of the anaerobic bacterium Clostridium beijerinckii were dried on transparent support surfaces. The laser beam of a confocal Raman microscope was focused on individual cells viewed through the objective. Single bacterial cells in size approximately 1 microm and sample mass approximately 1 pg could be analyzed within a few minutes, when placed on a calcium fluoride support and using excitation at 632.8 nm. Spectral features could be attributed to all major cell components. Cells from a morphologically differentiated culture sample showed different compositions, indicating the presence of subpopulations. As a reference, the storage polymer granulose was detected. The multidimensional information in Raman spectra gives a global view on all major components of the cell at once, complementing other more specific information-rich methods for single-cell analysis. The method can be used, for example, to study heterogeneities in a microbial population. PMID- 11101228 TI - A fluorescence-based method for determining the surface coverage and hybridization efficiency of thiol-capped oligonucleotides bound to gold thin films and nanoparticles. AB - Using a fluorescence-based method, we have determined the number of thiol derivatized single-stranded oligonucleotides bound to gold nanoparticles and their extent of hybridization with complementary oligonucleotides in solution. Oligonucleotide surface coverages of hexanethiol 12-mer oligonucleotides on gold nanoparticles (34 +/- 1 pmol/cm2) were significantly higher than on planar gold thin films (18 +/- 3 pmol/cm2), while the percentage of hybridizable strands on the gold nanoparticles (1.3 +/- 0.3 pmol/cm2, 4%) was lower than for gold thin films (6 +/- 2 pmol/cm2, 33%). A gradual increase in electrolyte concentration over the course of oligonucleotide deposition significantly increases surface coverage and consequently particle stability. In addition, oligonucleotide spacer sequences improve the hybridization efficiency of oligonucleotide-modified nanoparticles from approximately 4 to 44%. The surface coverage of recognition strands can be tailored using coadsorbed diluent oligonucleotides. This provides a means of indirectly controlling the average number of hybridized strands per nanoparticle. The work presented here has important implications with regard to understanding interactions between modified oligonucleotides and metal nanoparticles, as well as optimizing the sensitivity of gold nanoparticle-based oligonucleotide detection methods. PMID- 11101229 TI - Detection of native amino acids and peptides utilizing sinusoidal voltammetry. AB - Native amino acids and peptides were detected at a copper microelectrode using sinusoidal voltammetry (SV). Traditionally, these molecules can only be measured after derivatization with either a fluorescent or electroactive tag. In this work, an electrocatalytic oxidation reaction at copper is used to detect underivatized peptides and amino acids. The oxidation reaction is somewhat independent of peptide structure (i.e., it is not limited to the detection of aromatic amino acids) and is therefore able to produce nanomolar detection limits for all amino acids and peptides tested. A scanning technique, sinusoidal voltammetry, is used to provide the sensitivity of constant-potential techniques but also provide selectivity gained through utilization of the frequency domain. The frequency spectrum due to the oxidation of each molecule has a unique "fingerprint" response resulting from the kinetics of oxidation at the electrode surface. Through examination of the frequency spectra, even structurally similar molecules can be easily distinguished from one another. Flow injection analysis is used to demonstrate the sensitive and selective detection of a variety of amino acids and peptides. This technique can also be easily coupled to a separation step, i.e., high-performance liquid chromatography or capillary electrophoresis without electrode fouling from the adsorption of the analytes. PMID- 11101230 TI - Spectroelectrochemical sensing based on multimode selectivity simultaneously achievable in a single device. 9. Incorporation of planar waveguide technology. AB - Incorporation of planar waveguide technology into a spectroelectrochemical sensor is described. In this sensor design, a potassium ion-exchanged BK7 glass waveguide was over-coated with a thin film of indium tin oxide (ITO) that served as an optically transparent electrode. A chemically selective film was spin coated on top of the ITO film. The sensor supported five optical modes at 442 nm and three at 633 nm. Investigations on the impact of the ITO film on the optical properties of the waveguide and on the spectroelectrochemical performance of the sensor are reported. Sensing was based on the change in attenuation of light propagated through the waveguide resulting from an optically absorbing analyte. By applying either a triangular or square wave excitation potential waveform, electromodulation of the optical signal has been demonstrated with Fe(CN)6(3-/4-) as a model electroactive couple that partitions into a PDMDAAC-SiO2 film [where PDMDAAC = poly(dimethyldiallylammonium chloride)] and absorbs at 442 nm. PMID- 11101231 TI - The effects of nonionic surfactants on the tris(2,2'-bipyridyl)ruthenium(II)- tripropylamine electrochemiluminescence system. AB - The electrochemistry and electrogenerated chemiluminescence (ECL) of Ru(bpy)3(2+) (bpy = 2,2'-bipyridyl) were studied in the presence of the nonionic surfactants Triton X-100, Thesit, and Nonidet P40. The anodic oxidation of Ru(bpy)3(2+) produces ECL in the presence of tri-n-propylamine in both aqueous and surfactant solutions. Increases in both ECL efficiency (> or =8-fold) and duration of the ECL signal were observed in surfactant media. A shift to lower energies of the Ru(bpy)3(2+) ECL emission by approximately 8 nm was also observed. The one electron oxidation of Ru(bpy)3(2+) to Ru(bpy)3(3t) occurs at + 1.03 V vs Ag/AgCl in aqueous buffered (0.2 M potassium phosphate) solution as found by square wave voltammetry. This potential did not shift in surfactant systems, indicating that the redshifts in ECL emission are due to stabilization of ligand pi* orbitals in the metal-to-ligand charge-transfer excited state. These results are consistent with hydrophobic interactions between Ru(bpy)3(2+) and the nonionic surfactants. PMID- 11101232 TI - Pulse amperometric detection of salt concentrations by flow injection analysis using ionodes. AB - A sensitive novel approach of using an amperometric ion detector for the flow injection analysis of salts has been developed. The detection methodology is based on measuring the current associated with the transfer of ions across polarized microinterfaces between the aqueous sample solution and a 2 nitrophenyloctyl ether-poly(vinyl chloride) gel phase, referred to as ionodes. Different sodium salts of fluoride, chloride, bromide, nitrate, and sulfate were investigated. It was found that by employing an amperometric pulse detection mode and pure water as eluent, the detection limit of the ionode detector could be lowered to ppt level of salt concentrations under flowing conditions. PMID- 11101234 TI - Electrochemiluminescent metallopolymer coatings: combined light and current detection in flow injection analysis. AB - The application of thin films of the metallopolymer [Ru(bpy)2PVP10]2+ for the electrochemiluminescent (ECL) detection of oxalate in a flow injection analysis system is reported, where bpy is 2,2'-bipyridyl and PVP is poly(4-vinylpyridine). Immobilization of the ECL reagent means that it can be regenerated in situ, eliminating the need to constantly deliver it to the reaction zone. Electrochemically generated Ru3+ reacts with the analyte to form the excited state [Ru2+]*, which luminesces at 610 nm. The reaction is optimal at low pH, where the layer is swollen and homogeneous charge transport through the layer is more facile. Unlike traditional approaches, we simultaneously monitor both the amperometric and luminescent response of the modified electrode. The precision of both signals is similar at approximately 2% (n = 10). However, the ECL response has a larger dynamic range extending from the low-micromolar to high-millimolar range and a lower limit of detection, approximately 0.2 microM or 4 pmol of oxalate injected. The ECL approach displays excellent selectivity for oxalate over a wide range of potential interferences including oxygen, amines, iron sulfate, ammonium nitrate, urea, and glucose. Ascorbic acid represents the most significant ECL interference. However, the signal observed for a 1 mM solution of ascorbic acid is still only 2.6% of the response observed for the injection of a similar concentration of oxalate. PMID- 11101233 TI - Spectroelectrochemical sensing based on multimode selectivity simultaneously achievable in a single device. 5. Simulation of sensor response for different excitation potential waveforms. AB - The simulation of the optical response in spectroelectrochemical sensing has been investigated. The sensor consists of a sensing film coated on an optically transparent electrode (OTE). The mode of detection is attenuated total reflection. Only species that partition into the sensing film, undergo electrochemistry at the potentials applied to the OTE, and have changes in their absorbance at the wavelength of light propagated within the glass substrate of the OTE can be sensed. A fundamental question arises regarding the excitation potential waveforms employed to initiate the electrochemical changes observed. Historically, selection has been based solely upon the effectiveness of the waveform to quickly electrolyze any analyte observable by the optical detection method employed. In this report, additional requirements by which the waveform should be selected for use in a remote sensing configuration are discussed. The effectiveness of explicit finite difference simulation as a tool for investigating the applicability of three different excitation potential waveforms (square, triangle, sinusoid) is demonstrated. The simulated response is compared to experimental results obtained from a prototype sensing platform consisting of an indium tin oxide OTE coated with a cation-selective, sol-gel-derived Nafion composite film designed for the detection of a model analyte, tris(2,2' bipyridyl)ruthenium(II) chloride. Using a diffusion coefficient determined from experimental data (5.8 x 10(-11) cm2 s for 5 x 10(-6) M Ru(bipy)3(2+)), the simulator program was able to accurately predict the magnitude of the absorbance change for each potential waveform (0.497 for square, 0.403 for triangular, and 0.421 for sinusoid), but underestimated the number of cycles required to approach steady state. The simulator program predicted 2 (square), 3 (triangle), and 5 cycles (sinusoid), while 5 (square), 15 (triangle), and 10 (sinusoid) cycles were observed experimentally. PMID- 11101235 TI - Studies of protein--DNA interactions by capillary electrophoresis/laser-induced fluorescence polarization. AB - Protein-DNA interactions were studied on the basis of capillary electrophoretic separation of bound from free fluorescent probe followed by on-line detection with laser-induced fluorescence polarization. Changes in electrophoretic mobility and fluorescence anisotropy upon complex formation were monitored for the determination of binding affinity and stoichiometry. The method was applied to study the interactions of single-stranded DNA binding protein (SSB) with synthetic oligonucleotides and single-stranded DNA. Increases in fluorescence anisotropy and decreases in electrophoretic mobility upon their binding to SSB were observed for the fluorescently labeled 11-mer and 37-mer oligonucleotide probes. Fluorescence anisotropy and electrophoretic mobility were used to determine the binding constants of the SSB with the 11-mer (5 x 10(6) M(-1)) and the 37-mer (23 x 10(6) M(-1)). Alternatively, a fluorescently labeled SSB was used as a probe, and the formation of multiple protein-DNA complexes that differ in stoichiometry was observed. The results demonstrate the applicability of the method to study complex interactions between protein and DNA. PMID- 11101236 TI - Depth-profiling and diffusion measurements in ice films using infrared laser resonant desorption. AB - A new infrared laser resonant desorption (LRD) technique has been developed that permits depth-profiling and diffusion measurements in ice. This LRD technique utilizes an Er:YAG rotary Q-switched laser with an output wavelength of lambda = 2.94 microm and a pulse duration of approximately 100 ns. The Er:YAG laser light resonantly excites O-H stretching vibrations in the H2O molecules that form the ice. This laser resonant heating induces H2O desorption at the ice surface. Control experiments were conducted on pure and isotopically mixed laminated ice films to determine the optimum experimental parameters for the LRD depth profiling and diffusion measurements. Depending on laser energy, the measured desorption depth was either less than, comparable to, or larger than the optical penetration depth of approximately 0.8 microm at lambda = 2.94 microm. LRD studies were used to analyze H2 18O/H2 16O stacked multilayers and laminate sandwich structures. These measurements revealed that the LRD technique can depth profile into ice films with submicrometer spatial resolution and high sensitivity. Two types of experiments employing LRD depth-profiling were demonstrated to monitor diffusion in ice. HCl hydrate diffusion in ice was measured versus time after depositing ice/HCl/ice sandwich structures. Na diffusion into ice was studied after adsorbing Na using a continuous Na source for a given exposure time at the diffusion temperature. PMID- 11101237 TI - Development and evaluation of a glow discharge microwave-induced-plasma tandem source for time-of-flight mass spectrometry. AB - In this work, a new glow discharge, microwave-induced-plasma (GDMIP), tandem ion source was developed, characterized, and used in conjunction with time-of-flight mass spectrometry. This tandem source was designed to be simple and compact The GD and the MIP unit are independent and demountable, providing flexibility in tuning and operating. The microwave plasma can be formed with very low power (a few watts) and overlapped with the glow discharge. The discharge current increased with the addition of microwave discharge when the GD was operated in constant-voltage mode, and the amplitude of the current increase was found to be related to microwave power, discharge pressure, and sampling distance. With the addition of microwave discharge, significant signal enhancement was achieved under a discharge pressure of 2.0 Torr and a discharge voltage of 500 V with 2.0 mm sampling distance. Enhanced factors of analyte signals ranging from approximately 1 to nearly 10 could be obtained with MIP boosting, while no significant change in noise level was observed. This new tandem source provides improved performance in direct solid sample analysis. PMID- 11101238 TI - Probing single molecules in single living cells. AB - Single-molecule detection in single living cells has been achieved by using confocal fluorescence microscopy and externally tagged probe molecules. The intracellular background fluorescence is substantially higher than that in aqueous buffer, but this background is continuous and stable and does not significantly interfere with the measurement of single-molecule photon bursts. As a result, single-molecule data have been obtained on three types of fluorescent probes at spatially resolved locations (e.g., cytoplasm and nucleus) inside human HeLa cells. First, the iron transport protein transferrin labeled with tetramethylrhodamine undergoes rapid receptor-mediated endocytosis, and single transferrin molecules are detected inside living cells. Second, the cationic dye rhodamine 6G (R6G) enters cultured cells by a potential-driven process, and single R6G molecules are observed as intense photon bursts when they move in and out of the intracellular laser beam. Third, we report results on synthetic oligonucleotides that are tagged with a fluorescent dye and are taken up by living cells via a passive, nonendocytic pathway. PMID- 11101239 TI - Three-dimensional optical random access memory materials for use as radiation dosimeters. AB - This article describes the development of the first three-dimensional optical random access memory (3D-ORAM) material and readout system for monitoring energetic neutrons. Two different photochromic dyes, 5'-chloro-6-nitro-1',3',3' trimethylspiro-[2H-1-benzopyran-2,2'-in doline] (spirobenzopyran) and anthracene, have been investigated for use in these 3-D ORAM dosimeter materials. These dyes were immobilized in a poly(methyl methacrylate) support, and the resulting dosimeter materials were irradiated with neutrons from a Cf-252 source. Fluorescence measurements from the dosimeter show a dramatic decrease in the overall fluorescence intensity of the 3D-ORAM dosimeter exposed to the Cf-252, relative to a nonirradiated dosimeter. In addition, a two-photon excitation readout system has been developed for determining characteristics of the radiation that are necessary for estimating dose. PMID- 11101240 TI - High-density fiber-optic DNA random microsphere array. AB - A high-density fiber-optic DNA microarray sensor was developed to monitor multiple DNA sequences in parallel. Microarrays were prepared by randomly distributing DNA probe-functionalized 3.1-microm-diameter microspheres in an array of wells etched in a 500-microm-diameter optical imaging fiber. Registration of the microspheres was performed using an optical encoding scheme and a custom-built imaging system. Hybridization was visualized using fluorescent labeled DNA targets with a detection limit of 10 fM. Hybridization times of seconds are required for nanomolar target concentrations, and analysis is performed in minutes. PMID- 11101241 TI - TOF-SIMS characterization and imaging of controlled-release drug delivery systems. AB - Time-of-flight secondary ion mass spectrometry (TOF-SIMS) was used for the analysis of multilayer drug beads that serve as controlled-release drug delivery systems. TOF-SIMS analysis of a cross section of each bead system allowed molecular chemical information to be gained from all of the layers simultaneously, in situ. The integrity of each of the layers was evaluated through imaging of specific ion species for the drug, excipient, and coating materials. The three beads in this study each showed a unique distribution of ingredients. Images of the parent molecular ion for each drug (theophylline, paracetamol, prednisolone) showed their distribution ranged from micrometer-sized particles in one bead cross section to almost homogeneous in another bead cross section. The chemical composition of each of the layers in the beads was evaluated through mass spectrometry; the ingredients did not always match the manufacturer's specification. In addition, many common drug bead ingredients were analyzed as pure substances, providing TOF-SIMS reference spectra of these materials for the first time. PMID- 11101242 TI - A REMPI method for the ultrasensitive detection of NO and NO2 using atmospheric pressure laser ionization mass spectrometry. AB - We report on the development of a quasi-simultaneous highly selective method for NO and NO2 detection at the ultratrace level. Atmospheric pressure laser ionization (APLI), recently introduced by our group, is used to detect both compounds at low parts per trillion by volume (pptv) mixing ratios. APLI is based on resonance-enhanced multiphoton ionization mass spectrometry. Two-color pump probe experiments employing a single excimer pumped dye laser combination allow for the ultrasensitive measurement of NO and NO2 within a narrow range of maximum pumping efficiency of the laser dye Coumarin 120. NO is detected via excitation of the long-lived A 2sigma+ (nu' = 1) level at 215.36 nm and subsequently ionized with 308-nm radiation provided by the excimer pump laser. NO2 is ionized after double resonant excitation of the A2B1 and 3psigma manifolds in a (1 + 1' + 1(')) process using 431.65 + 308 nm. The selectivity of the NO measurement exceeds 2,000 with respect to NO2 and N2O5. For NO2, a selectivity of >3,000 with respect to N2O5 and organic nitrates is observed. The current APLI detection limit of NO and NO2 is 0.5 and 5 pptv, respectively, with a 20-s integration time. PMID- 11101243 TI - An interface with a linear quadrupole ion guide for an electrospray-ion trap mass spectrometer system. AB - A new ion sampling interface for an electrospray ionization 3D ion trap mass spectrometer system is described. The interface uses linear rf quadrupoles as ion guides and ion traps to enhance the performance of the 3D trap. Trapping ions in the linear quadrupoles is demonstrated to improve the duty cycle of the system. Dipolar excitation of ions trapped in a linear quadrupole is used to eject unwanted ions. A resolution of ejection of up to 254 is demonstrated for protonated reserpine ions (m/z 609.3). A composite waveform with a notch in frequency space is used to eject a wide range of matrix ions and to isolate trace analyte ions in a linear quadrupole before ions are injected into the 3D trap. This is useful to overcome space charge problems in the 3D trap caused by excess matrix ions. For trace reserpine in a 500-fold molar excess of poly(propylene glycol) (PPG), it is demonstrated that the resolution and sensitivity of the 3D trap can be increased dramatically with ejection of the excess PPG matrix ions. In comparison to ejection of matrix ions in the 3D trap with a similar broad-band waveform, a 5-fold increase in sensitivity with a 7 times shorter acquisition time was achieved. PMID- 11101244 TI - A robust, detergent-friendly method for mass spectrometric analysis of integral membrane proteins. AB - Recent breakthroughs in the high-resolution structural elucidation of ion channels and transporters are prompting a growing interest in methods for characterizing integral membrane proteins. These methods are proving extremely valuable in facilitating the production of X-ray diffraction-grade crystals. Here we present a robust and straightforward mass spectrometric procedure that utilizes matrix-assisted laser desorption/ionization to analyze integral membrane proteins in the presence of detergents. The utility of this method is illustrated with examples of high-quality mass spectral data obtained from membrane proteins for which atomic resolution structural studies are ongoing. PMID- 11101245 TI - An automated technique for measuring deltaD and delta18O values of porewater by direct CO2 and H2 equilibration. AB - The stable-oxygen and -hydrogen isotopic values (deltaD, delta18O) of porewater in geologic media are commonly determined on water obtained by extraction techniques such as centrifugation, mechanical squeezing, vacuum heating and cryogenic microdistillation, and azeotropic distillation. Each of these techniques may cause isotopic fractionation as part the extraction process and each is laborious. Here we demonstrate a new approach to obtain automated, high precision deltaD and delta18O measurements of porewater in geologic sediments by direct H2- and CO2-porewater equilibration using a modified commercial CO2-water equilibrator. This technique provides an important and cost-effective improvement over current extraction methods, because many samples can be rapidly analyzed with minimal handling, thereby reducing errors and potential for isotopic fractionation. The precision and accuracy of direct H2- and CO2-porewater equilibration is comparable to or better than current porewater extraction methods. Finally, the direct equilibration technique allows investigators to obtain high-resolution (cm scale) porewater deltaD and delta18O profiles using cores from individual boreholes, eliminating the need for costly piezometers or conventional porewater extractions. PMID- 11101246 TI - An electrospray ionization source for the investigation of thermally initiated reactions. AB - An electrospray ionization source for mass spectrometry is described, which allows one to heat the spray capillary. Thermal dissociation of the trityl dimer 1 and tetra(p-anisyl)hydrazine (3) have been investigated, operating the ESI source as electrolytic cell to ionize neutral species, e.g. the trityl radical 2. PMID- 11101247 TI - Carbon isotope effects resulting from equilibrium sorption of dissolved VOCs. AB - To accurately interpret isotopic data obtained for volatile organic compounds (VOCs) dissolved in groundwater systems, the isotopic effects of subsurface processes must be understood. Previous work has demonstrated that volatilization and dissolution of BTEX and chlorinated ethene compounds are not significantly isotopically fractionating. This study characterized the carbon isotopic effects of equilibrium sorption of perchloroethylene, trichloroethylene, benzene, and toluene to both graphite and activated carbon directly in batch experiments over a range of 10-90% sorption. Results demonstrate that, over this range, equilibrium sorption of these VOCs to graphite and activated carbon does not result in significant carbon isotopic fractionation within the +/-0.5% accuracy and reproducibility associated with compound-specific isotope analysis. This implies that the isotopic values of dissolved VOCs will not be significantly affected by equilibrium sorption in the subsurface. Therefore, isotopic analysis has potential to be used in the field to differentiate between mass losses due to isotopically fractionating processes such as biodegradation versus mass loss due to nondegradative processes. PMID- 11101248 TI - Observation of low molecular weight poly(methylsilsesquioxane)s by graphite plate laser desorption/ionization time-of-flight mass spectrometry. AB - Mass spectra of polystyrene and poly(methylsilsesquioxane)(PMSSQ) derived from methyltriethoxysilane(MTES) were obtained in the 100-1,000 Da range by laser desorption/ionization time-of-flight mass spectrometry using a graphite plate without a matrix. Clean mass spectra were obtained without interference from carbon clusters or other low molecular weight compounds. Initial reaction products derived from condensation of partially hydrolyzed MTES were observed. Upon 30 min of heating at 30 degrees C, the ethoxy groups were fully hydrolyzed to hydroxy groups. Many PMSSQ species consistent with predictable polymerization reaction pathways involving intermolecular condensation and intramolecular dehydration were observed. Thus, laser desorption/ionization time-of-flight mass spectrometry using a graphite plate, without added matrix materials, is shown to provide valuable information on low molecular weight polymer not available by MALDI-TOF-MS. PMID- 11101249 TI - A study of the enthalpy and entropy contributions of the stationary phase in reversed-phase liquid chromatography. AB - The goal of this study was to elucidate the roles played by the stationary and mobile phases in retention in reversed-phase liquid chromatography (RPLC) in terms of their individual enthalpic and entropic contribution to the Gibbs free energy of retention. The experimental approach involved measuring standard enthalpies of transfer of alkylbenzenes from typical mobile phases used in RPLC (methanol/water and acetonitrile/water mixtures), as well as from n-hexadecane (a simple analogue of the stationary phase) to the gas phase, using high-precision headspace gas chromatography. By combining the measured enthalpies with independently measured free energies of transfer, the entropies of transfer were obtained. This allowed us to examine more fully the contribution that each phase makes to the overall retention. It was found that the standard enthalpy of retention in RPLC (i.e., solute transfer from the mobile phase to the stationary phase) is favorable, due to the large and favorable stationary-phase contribution, which actually overcomes an unfavorable mobile-phase contribution to the enthalpy of retention. Further, the net free energy of retention is favorable due to the favorable enthalpic contribution to retention, which arises from the net interactions in the stationary phase. Entropic contributions to retention are not controlling. Therefore, to a great extent, retention is due to enthalpically dominated lipophilic interaction of nonpolar solutes with the stationary phase and not from solvophobic processes in the mobile phase. Further, our enthalpy data support a "partition-like" mechanism of retention rather than an "adsorption-like" mechanism. These results indicate that the stationary phase plays a very significant role in the overall retention process. Our conclusions are in direct contrast to the solvophobic model that has been used extensively to interpret retention in RPLC. PMID- 11101250 TI - Construction of large-volume monolithic columns. AB - Monolithic supports have become the subject of extensive study in the past years. Despite their advantageous features and many successful chromatographic applications in the analytical scale, only a very few examples of larger volume monoliths were described. In the case of GMA-EDMA monoliths, this can be attributed to the fact that due to the exothermic polymerization a pronounced temperature increase inside the monolith significantly affects the structure. The temperature increase depends on the thickness of the monolith, and consequently, there is an upper limit that allows the preparation of a unit with a uniform structure. In the present work, we have analyzed a heat release during the polymerization and have derived a mathematical model for the prediction of the maximal thickness of the monolithic annulus having a uniform structure. On the basis of the calculations, two annuluses of different diameters were polymerized and merged into a single monolithic unit with a volume of 80 mL. In addition, a special housing was designed to provide a uniform flow distribution in the radial direction over the entire monolith bed. It was shown that such a monolithic column exhibits flow-independent separation efficiency and dynamic binding capacity up to flow rates higher than 100 mL/min. The separation and loading times are in the range of a few minutes. The pressure drop on the column is linearly dependent on the flow rate and does not exceed 2.5 MPa at a flow rate of 250 ml/min. PMID- 11101251 TI - Justification of statistical overlap theory in programmed temperature gas chromatography: thermodynamic origin of random distribution of retention times. AB - A specific distribution of compounds' standard-state changes of enthalpy and entropy between mobile and stationary phases in programmed temperature gas chromatography (PTGC) is shown to produce the Poisson distribution of retention times often postulated in statistical-overlap theory (SOT). A three-part model is proposed, in which the enthalpy change is Poisson distributed, the average entropy change depends on the enthalpy change, and the actual entropy change varies in a uniformly random manner about the average entropy change. To test the model, the entropy and enthalpy changes of 350 aliphatic and aromatic hydrocarbons in petroleum were calculated with commercial GC software. These changes are shown to follow the three-part model. The model then was used with Monte Carlo methods to mimic the enthalpy and entropy changes. The substitution of the mimicked enthalpy and entropy changes into an equation for the retention temperature in PTGC is shown to produce a Poisson distribution of retention times that is statistically significant. This finding establishes a scientific link between the thermodynamics governing retention in PTGC and the superficially ad hoc assumption of the Poisson retention time distribution in SOT. Similar thermodynamic distributions are found for flavors and fragrances and for tetrachlorodibenzo-p-dioxins and furans, which follow SOT based on the Poisson distribution, but not for polychloronaphthalenes, which do not follow that SOT. PMID- 11101252 TI - Determination of effective protein charge by capillary electrophoresis: effects of charge regulation in the analysis of charge ladders. AB - Protein charge ladders are an effective tool for measuring protein charge and studying electrostatic interactions. However, previous analyses have neglected the effects of charge regulation, the alteration in the extent of amino acid ionization associated with differences between the pH at the protein surface and in the bulk solution. Experimental data were obtained with charge ladders constructed from bovine carbonic anhydrase. The protein charge for each element in the ladder was calculated from the protein electrophoretic mobility as measured by capillary electrophoresis using the hindrance factor for a hard sphere with equivalent hydrodynamic radius. The protein charge was also evaluated theoretically from the amino acid sequence by assuming a Boltzmann distribution in the hydrogen ion concentration. The calculations were in excellent agreement with the data, demonstrating the importance of charge regulation on the net protein charge. These results have important implications for the use of charge ladders to evaluate effective protein charge in solution. PMID- 11101253 TI - Surface-enhanced resonance Raman spectroscopy as an identification tool in column liquid chromatography. AB - The compatibility of ion-pair reversed-phase column liquid chromatography and surface-enhanced resonance Raman spectroscopy (SERRS) for separation and identification of anionic dyes has been investigated, with emphasis on the at line coupling via a thin-layer chromatography (TLC) plate. SERR spectra using silver sols were recorded both for aqueous solutions and for samples deposited on aluminum oxide and silica TLC plates at 514.5- and 457.9-nm laser excitation. For some dyes, the shorter wavelength was needed to diminish the fluorescence background. For aqueous solutions and for samples deposited on aluminum oxide, clear SERR spectra were obtained upon addition of poly(L-lysine); for the silica plates, the addition of nitric acid was required. Upon drying the plates, the SERRS signals decreased in intensity; simply adding a drop of water could largely restore them. At-line coupling of LC and SERRS was successfully achieved when using silica, but not aluminum oxide, plates. The application of a gradient, a high water content, and the presence of ion-pair reagents needed for the separation did not adversely affect the deposition and the recording of SERR spectra. The identification limits were 10-20 ng of deposited material, depending on the dye selected, which corresponded to injected concentrations of 5-10 microg mL(-1). PMID- 11101254 TI - On-line separation and determination of bromate in drinking waters using flow injection ICP mass spectrometry. AB - A flow injection (FI) system with a microcolumn of anion exchanger has been used to effect rapid on-line separation of bromate and bromide prior to quantitation by ICP mass spectrometry. Basic performance studies are described including the effect of key FI parameters, i.e., sample injection volume, carrier stream flow rate, and eluent concentration on system response. The new approach permitted ultratrace determinations of bromate in drinking waters, the main benefits being low limit of detection (0.13 microg/L based on a 500-microL sample injection), rapid analysis time (10 min/sample), and good precision (2.8% at the 5 microg/L level). Accuracy was checked via an EC-sponsored interlaboratory trial. PMID- 11101256 TI - Automatic on-line coupling of supercritical fluid extraction and capillary electrophoresis. AB - An interface for the automatic coupling of a supercritical fluid extractor (SFE) with capillary electrophoretic (CE) equipment, both commercially available, was developed with a view to improving sample treatment, which is a crucial step in capillary electrophoresis. Extracted analytes were collected in a trap following depressurization in the SFE and were transferred to the CE equipment across the interface. The key elements of the experimental assembly are a laboratory-made programmable arm and the autosampler of the CE equipment, both of which are controlled by a built-in microprocessor using an appropriate electronic interface and customized software. This combined system was successfully used to determine cresols and chlorophenols in liquid samples (river water and human urine) with increased precision, throughput, and automatability. The proposed arrangement opens up interesting prospects for the direct determination of analyte traces in solid samples without human intervention. PMID- 11101255 TI - Electrophoretically mediated microanalysis of leucine aminopeptidase using two photon excited fluorescence detection on a microchip. AB - Two-photon excited fluorescence detection was performed on a microfabricated electrophoresis chip. A calibration curve of the fluorescent tag beta naphthylamine was performed, resulting in a sensitivity of 2.5 x 10(9) counts M( 1) corresponding to a detection limit of 60 nM. Additionally, leucine aminopeptidase was assayed on the chip using electrophoretically mediated microanalysis. The differential electroosmotic mobilities of the enzyme and substrate, L-leucine beta-naphthylamide, allowed for efficient mixing in an open channel, resulting in the detection of a 30 nM enzyme solution under constant potential. A zero potential incubation for 1 min yielded a calculated detection limit of 4 nM enzyme. PMID- 11101257 TI - Tantalum-filament pulling device for fabrication of submicrometer fused-silica tips. AB - A simple and low-cost pulling device for fused-silica capillaries was developed. By using a tantalum heating filament and the self-tension in a bent capillary, tips and constricted regions with outer diameters of approximately 1 microm and inner diameters of a few hundred nanometers could be reproducibly pulled from 50 microm-i.d., 375-microm-o.d. capillaries. The tips can be used in different applications such as microinjection, micromanipulation, and single-channel patch clamp, injection ends for CE or as electrospray tips. Constricted capillaries with optimized dimensions to minimize cylindrical lensing effects and to match the size of a diffraction-limited laser focus can be used as optical detection windows in CE and micro-HPLC. Fused silica has several advantages over other glasses such as high melting temperature and superior optical and mechanical properties. PMID- 11101258 TI - High-efficiency on-line concentration technique of capillary electrochromatography. AB - With the coexistence of the mobile phase, the stationary phase, and the electric field in capillary electrochromatography, the chromatographic zone-sharpening effect and field-enhanced sample-stacking technique were utilized to improve detection sensitivity. By the former means, with less organic modifier in the sample solution compared to that in the mobile phase, the concentration factors of neutral solutes benzoin and mephenytion were 134 and 219, respectively. Through the latter one, without electrolyte in the sample solution, the detection sensitivity of propatenene with positive charge was improved by 1600 times. While with the combination of these two methods, improvement of over 17,000 times for the sensitivity of propatenene was obtained. By the combined means, the analysis of basic pharmaceutical compounds at concentrations of nanograms per milliliter by UV detection was realized. In addition, parameters that might affect the efficiency of on-line concentration were studied and equations that described the on-line concentration procedure were deduced. PMID- 11101259 TI - Extending the reach of immunoassays to optically dense specimens by using two photon excited fluorescence polarization. AB - Fluorescence anisotropy/polarization measurements represent a powerful tool for quantifying biomolecule/ligand complexation. These types of measurements are also at the heart of a wide variety of commercial homogeneous fluoroimmunoassays. In this note, we demonstrate the power of two-photon excited fluorescence anisotropy (2-PEFA) measurements as a tool for quantifying hapten/antibody association in the presence of a strongly absorbing, nonfluorescent dye. The results of these experiments show that 2-PEFA measurements are intrinsically more sensitive when compared to traditional one-photon excited fluorescence anisotropy (1-PEFA) strategies and 2-PEFA-based measurements allow one to perform accurate hapten/antibody binding measurements in strongly absorbing samples directly under conditions where 1-PEFA measurements fail completely. Overall, the 2-PEFA approach offers significant advantages when compared to traditional 1-PEFA methods especially in strongly absorbing samples. 2-PEFA also opens the door to perform more rapid and reliable polarization/anisotropy-based measurements with minimal sample preparation. PMID- 11101260 TI - Geography and the Olympics: an evaluation of the work of Ernst Jokl. PMID- 11101261 TI - Periodization of pyschological skills training. PMID- 11101262 TI - Physical activity programs for underserved youth. AB - Underserved youth face enormous barriers in their emotional, social, and intellectual development. One such barrier is the scarcity of developmentally oriented extended day programs. Recent research and professional opinion have stimulated a reconceptualization of the field of youth development as well as the identification of specific guidelines for extended day programs in underserved communities. The Urban Youth Leader Project (UYLP) in Chicago, which has now spread to several other universities, is a specific example of these developments in practice. UYLP sponsors 1) a number of youth programs in Chicago's most underserved communities, 2) service learning and professional preparation programs linked to the community youth programs for interested university students, and 3) applied research. The personal and social responsibility model provides the template for youth programs and for university interns and is the focus of the applied research. Several alternative structures have been created to provide more compatible settings for this work. Although such activities hold promise for incremental change, they do not address the more deeply rooted systemic causes of being underserved. PMID- 11101263 TI - The role of the motor system in spinal pain: implications for rehabilitation of the athlete following lower back pain. AB - The purpose of this review is to consider the role of the motor system in spinal pain. It is well accepted that spinal stability is dependent on the contribution of the muscular system. However, the ability of this system to satisfy the requirements of stability is dependent on its controller--the central nervous system (CNS). The CNS must predict the outcome of movements to plan appropriate strategies of muscle activity to meet the demands of internal and external forces, and initiate appropriate responses to unexpected disturbances. In addition, this complex control of stability must occur in conjunction with control of the trunk muscles for other functions, such as respiration. For the CNS to cope with athletic performance the coordination of these parameters must be streamlined. Yet evidence suggests that when spinal pain is present the strategies used by the CNS to control trunk muscles may be altered. The mechanism for these changes is poorly understood but may be due to changes at many levels of the CNS. For rehabilitation of the athlete with spinal pain it is critical that the motor control of stability is optimised. Furthermore, this must be coordinated with the multiple other functions of trunk muscles, including respiration. PMID- 11101264 TI - Sport and exercise medicine: building the foundations of a new discipline. AB - New disciplines emerge as medicine evolves. But, to be recognised as a clinical discipline we must be prepared to measure ourselves against established criteria; there should be consensus on the core content of the discipline and an identifiable body of knowledge to allow us to deal with common clinical problems. At this developmental stage in our evolution we have general agreement on what defines sport and exercise medicine but, as most of our clinical skills developed empirically, research is relatively underdeveloped. The volume and quality of this research does not yet reflect a vibrant research culture. The validity of our clinical method is supported by research evaluating the sensitivity and specificity of clinical tests but the evidence supporting other areas of clinical practice is uncertain. Some aspects of prevention have become integrated into clinical practice without sufficient evidence and, screening has been introduced without critical evaluation. To ensure the future success of our discipline and achieve recognition of its value in the wider medical community, we must nurture an evaluative culture and ensure that clinical practice is built on firm foundations of research evidence. PMID- 11101265 TI - Athletic footwear: design, performance and selection issues. AB - The purpose of this paper is to assist the practitioner in understanding the various advantages and disadvantages associated with the use of athletic footwear. In addition, the various components of a typical athletic shoe are described, including the upper, the midsole/outsole, the last, as well as the lasting process. Since the various models of athletic shoes that are available to the consumer can change in a very rapid and unpredictable manner, it is extremely difficult for the clinician to maintain a database of current shoe models and features. This paper stresses the importance of the clinician providing the athlete a list of footwear features and components based on their particular foot classification or problem, rather than attempting to recommend a specific model of athletic shoe. A detailed explanation of these features is provided to assist the practitioner in helping the athlete select the most appropriate shoe. PMID- 11101266 TI - Stress fractures and bone health in track and field athletes. AB - The effect of exercise on bone health has received much attention in recent years. The problems of the female athlete triad: disordered eating, amenorrhea and osteoporosis have helped us to better understand and appreciate the important interaction of mechanical, hormonal, nutritional as well as genetic factors on bone health in the young female athlete. The relatively high stress fracture incidence of young track and field athletes can be quite disabling for the athlete's present and future running career. A number of risk factors including low bone mineral density (BMD), menstrual irregularities, dietary factors and prior history of stress fractures have been associated with an increased risk for stress fractures in the female athlete. Few studies have found risk factors for stress fractures in the male athlete. Female gender has been found to be a risk factor for stress fractures in the military population, but this finding is less apparent in athlete studies. Caucasians have been found to have a higher risk for stress fractures than African-American military recruits, but there is very limited data assessing stress fracture risk in athletes of varying ethnicity. Prevention of stress injury to bone involves maximizing peak bone mass in the pediatric and young adult age groups. Maintaining adequate calcium nutrition, caloric intake as well as hormonal and energy balance are important preventive measures, as are ensuring appropriate amounts of weight bearing exercise for optimizing bone health and preventing fractures. More research is needed to determine factors leading to improvements in bone density and fracture reduction in athletes at risk. PMID- 11101267 TI - Promoting physical activity for youth. AB - The challenge for helping others enjoy a healthy and active life is to move the focus of instruction from physical fitness toward physical activity. Participation in regular physical activity offers a number of benefits including reduction of the risk of premature mortality. coronary heart disease, diabetes mellitus, hypertension, and colon cancer. The physical fitness of American children has not declined over the years even though teachers and parents often believe it to be true. A significant amount of fitness test performance is explained by heredity. Both the response to training and genetic limitations are limiting factors outside the control of individuals. Not all people can reach a high fitness level, but all can be physically active. The Children's Lifetime Physical Activity Model (C-LPAM) offers guidance in how to prescribe activity for youth. Guidelines suggest youngsters should receive at least 60 minutes or more of physical activity on a daily basis. PMID- 11101268 TI - Gender differences in metabolism; nutrition and supplements. AB - For many decades researchers did not consider that there were any differences between the genders in the metabolic response to exercise. As a result, nutritional recommendations and exercise training prescriptions have not considered the potential for gender specific responses. More recently, we and others have demonstrated that females oxidize proportionately more lipid and less carbohydrate during endurance exercise as compared to males. The oxidation of amino acids is similarly lower in females as compared to males during exercise. These gender differences are partially mediated by a higher estrogen concentration in females. Specific areas where there are gender differences in nutritional/supplement recommendations include carbohydrate (CHO) nutrition, protein requirements and creatine (CRM) supplementation. We have shown that females do not carbohydrate load in response to an increase in dietary carbohydrate when expressed as a percentage of total energy intake (i.e., 55 75%), however if they consume >8 g CHOxkg(-1)xd(-1), they show similar increases as compared to males. Top sport male and female athletes require somewhat more dietary protein as compared to sedentary persons. The maximal increase is approximately 100% for elite male athletes and approximately 50-60% for elite female athletes. Fortunately, most athletes habitually consume this level of protein intake. We have recently demonstrated that females show a lesser increase in lean body mass following acute CRM loading as compared to males. Females also did not show reductions in protein breakdown in response to CRM loading, whereas males did. In the future I expect that there will be further research from which gender specific nutritional/supplement recommendations can be made. PMID- 11101269 TI - Altitude acclimatization, training and performance. AB - Exposure to altitude results in a reduction in partial pressure of oxygen in the arterial blood and a reduction in oxygen content. In an attempt to maintain aerobic metabolism during increased effort, a series of acclimatization responses occur. Among the most conspicuous of these responses is an increase in hemoglobin (Hb) concentration. The increase in Hb has been construed as the fundamental adaptation enabling increases in aerobic power and performance to occur on return to sea-level. However, the use of altitude to boost training adaptations and improve elite sea-level performance, although tantalizing, is largely unproven. The reasons appear to be many, ranging from the poor experimental designs employed, to the numerous strategies designed to manipulate the altitude experience and the large inter-individual differences in response patterns. However, other factors may also be important. Acclimatization has also been shown to induce alteration in selected properties of the muscle cell, some of which may be counterproductive. The processes involved in cation cycling, as an example, appear to be down-regulated. Changes in these processes could impair certain types of performance. PMID- 11101270 TI - Functional adaptation of bone to exercise and injury. AB - Bone adapts to altered physical stimuli, dietary changes, or injury. Dietary calcium and vitamins play important roles in maintaining skeletal health, but high-fat diets are pervasive in western cultures and may contribute to the increasing prevalence of osteoporosis and incidence of related hip fractures. Exercise helps maintain bone mass and counter osteoporosis, but exercise can also have detrimental effects-particularly for immature bone. Some negative exercise effects may also be linked to diet. For example, insufficient dietary protein during exercise can impair bone development and remodeling. Bone remodeling is a potent example of tissue repair. Chronically altered loading after a joint injury, however, can result in remodeling processes that can be detrimental to the joint. Anterior cruciate ligament injury, for example, commonly leads to osteoarthritis. Early changes in the periarticular cancellous bone may play a role in the development of knee osteoarthritis. Although these factors influence skeletal health, the mechanisms remain unclear by which bone interprets its environment and responds to mechanical stimuli or injury. To understand why different levels of exercise are beneficial or detrimental or why altered joint loading leads to changes in periarticular bone structure, underlying mechanisms must be understood by which bone interprets its mechanical environment. PMID- 11101271 TI - A novel approach to pain relief pre-therapeutic exercise. AB - Musculoskeletal problems are often multifactorial and consequently can be challenging to treat. This paper examines management of chronic musculoskeletal conditions in the light of Panjabi's stabilisation subsystems and Dye's concept of homeostasis and critical symptom threshold. In many circumstances treatment can aggravate symptoms. Tape may be used to unload painful structures to minimise the aggravation of the symptoms so treatment can be directed at improving the patient's 'envelope of function'. This involves specific muscle training of the dynamically unstable segment/s and increasing the mobility of the less flexible surrounding soft tissues. Three case studies of chronic low back and leg pain, patellofemoral pain and shoulder impingement secondary to multidirectional instability, are presented as examples of multifactorial musculoskeletal problems requiring unloading, stabilisation and control. PMID- 11101272 TI - Effect of serum on the in vitro maturation of canine oocytes. AB - The meiotic competence of canine oocytes cultured for 72 h in medium supplemented with three different concentrations (5, 10 and 20%) of anoestrous, oestrous or metoestrous bitch serum, or with 0.3% bovine serum albumin (BSA), was examined. The oestrous serum supplement had a positive effect on the resumption of meiosis, compared with the other supplements (P<0.05). The number of oocytes that reached metaphase I (MI) and metaphase II (MII) was significantly higher (P<0.05) with the oestrous serum supplement than with the anoestrous serum supplement. There were no significant differences among the three different concentrations in each serum type with respect to the proportion of oocytes that completed meiosis (MI to MII). The number of oocytes that resumed meiosis in the 10% oestrous serum supplement was significantly higher (P<0.05) than those of each concentration of the anoestrous and metoestrous serum supplements, and of the 0.3% BSA supplement. Moreover, a higher number of oocytes reached MII in the presence of the 10% oestrous serum supplement than with the 10% anoestrous serum supplement. These results suggest that supplementation of the culture medium with 10% oestrous serum is the optimal treatment for in vitro maturation of canine oocytes. PMID- 11101273 TI - Maternal behaviour and the survival of lambs in superfine wool sheep. AB - Superfine merino lambs from superfine and medium-wool ewes were compared to test the hypothesis that superfine ewes have poorer maternal capacity than broader wool ewes. All lambs were sired by one ram using artificial insemination in the superfine ewes and embryo transfer in the medium-wool ewes. This ensured a similar genetic background in all lambs. Single born lambs from the superfine ewes were 1.5 kg lighter than those from medium-wool ewes (3.3 v. 4.8 kg, P<0.05). Compared with the medium-wool ewes, in the first hour after birth the superfine ewes spent less time grooming their lambs (8.7 v. 25.2 min, P<0.05) and more time separated from them (21.3 v. 0.0 min, P<0.05). The lambs of superfine ewes also stood and attempted to suck for less time (17.7 and 2.4 min v. 32.4 and 10.5 min, P<0.05) than those of medium-wool ewes. Twenty-nine per cent of lambs from superfine ewes died in the first 4 weeks after birth compared with only 4% of lambs from medium-wool ewes. It was concluded that superfine ewes lost more lambs than medium-wool ewes and that this was associated with their lambs being smaller at birth and with the ewes being poorer mothers. PMID- 11101274 TI - Matrix metalloproteinases and their endogenous inhibitors during the implantation period in the rat uterus. AB - Rats were studied to determine if matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) were detectable in developing embryonic and uterine tissues during the implantation period; that is, Days 6-8 of pregnancy. Tissue extracts were studied by zymography, reverse zymography and Western blotting. Immunohistochemistry procedures using antibodies against MMPs and TIMPs were applied to tissue sections that passed through implantation sites. The MMP inhibitor, doxycycline, was injected intraluminally into uterine horns of pseudopregnant animals. MMP-2 and -9, and TIMP-2 were detected in gelatin and reverse zymographs, respectively, from extracts of implantation sites on each day studied. Western blots defined bands corresponding to MMP-1, but casein zymographs did not consistently show evidence of extractable MMP-1 or -3. In tissue sections the primary decidual tissue area stained consistently for TIMP-2 and on Days 7 and 8 this area also had positive staining for MMP-1 in close proximity to the implanting embryo. On Day 8 positive staining in the outer stromal tissue was detected using an antibody against MMP-9, and another MMP (possibly MMP-3) was localized exclusively in ectoplacental cone/trophoblastic cells. Intraluminal injection of doxycycline did not significantly prevent the growth of the uterine horns following surgical induction of pseudopregnancy on Day 5. The strong localization of TIMP-2 in the primary decidual tissue is similar to that reported previously for TIMP-3, indicating that these inhibitors have a role in decidualization, including the regulation of trophoblastic invasion. PMID- 11101275 TI - Effects of maternal captopril treatment on growth, blood glucose and plasma insulin in the fetal spontaneously hypertensive rat. AB - In the spontaneously hypertensive rat (SHR) fetal growth and metabolism are abnormal. It has been speculated that maternal hypertension may be the cause of these abnormalities. Captopril treatment, which reduces maternal blood pressure, during pregnancy and lactation, is reported to have a beneficial effect postnatally, normalizing the blood pressure of offspring in the SHR. In the present study, the effects of maternal captopril treatment on fetal growth and plasma metabolites were investigated in the fetuses of two rat strains (SHR and Wistar-Kyoto (WKY)), in order to determine whether normalizing maternal blood pressure also normalized abnormalities in fetal growth and metabolism. On fetal Day 20, SHR fetuses were lighter and placentae were heavier than for the corresponding WKY. Captopril had no effect on fetal weight in the SHR, but decreased it in the WKY. There was no effect of captopril on placental weight. Fetal plasma insulin levels were higher in the SHR than in the WKY and were decreased by captopril treatment in both strains. Fetal blood glucose was elevated and fetal blood lactate was decreased in captopril-treated litters from both strains. Captopril had no effect on fetal plasma IGF-1 but fetal plasma IGF 2 levels were lower in the captopril-treated SHR than in the captopril-treated WKY. These findings suggest that maternal captopril treatment decreases insulin secretion in the fetal rat. High levels of fetal plasma insulin suggest that the SHR fetus is insulin resistant. Fetal insulin levels may contribute to the adverse consequences of gestational captopril treatment observed in many species. The differences in the effect of captopril on the two strains suggest that there are underlying endocrine differences in the SHR. PMID- 11101276 TI - Glycolytic enzyme activity in hypotonically treated boar spermatozoa. AB - Treatment of washed boar sperm with hypotonic phosphate buffer disrupted the cytoplasmic membrane and released the soluble contents and phosphofructokinase, but the other glycolytic enzymes and lactate dehydrogenase were retained. Addition of the appropriate substrates and co-factor(s) to preparations of treated cells in phosphate-buffered saline showed that enzyme activity could be re-instated. This simple preparation should be of assistance in the investigation of specific sections of the glycolytic pathway without the use of chemical inhibitors. PMID- 11101277 TI - Chronic cold exposure has no effect on brown adipose tissue in newborn lambs born to well-fed ewes. AB - It has been previously shown in twin-bearing ewes fed only 60% of their metabolizable energy requirements for late pregnancy that chronic cold exposure induced by winter shearing of the ewes results in larger lambs with more brown adipose tissue. This effect appears to be primarily due to prevention of a decline in fetal body and tissue weights between 145 days' gestation and 2 h after birth (i.e. 147 days' gestation) in lambs born to underfed shorn ewes. The present study therefore examined the impact, in ewes that were well fed (i.e. received 100% of their metabolizable energy requirements) during the final month of gestation, of chronic cold exposure induced by winter shearing on lamb birthweight and perirenal adipose tissue composition as measured 2 h after birth. Perirenal adipose tissue was analysed for its thermogenic activity (i.e. GDP binding to mitochondria) and catecholamine content. These observations were combined with similar measurements made in near-term (i.e. 145 days' gestation) fetuses sampled from well-fed unshorn ewes. There was no difference between lambs born to shorn or unshorn ewes with respect to birthweight or perirenal adipose tissue weight and composition. Perirenal adipose tissue weight was higher in lambs born to unshorn ewes than in fetuses. The thermogenic activity of adipose tissue was 2-fold higher in lambs born to unshorn ewes compared with 145-day-old fetuses. Epinephrine was detectable only at very low levels in fetal perirenal adipose tissue, increasing 10-fold after birth, with no difference between lambs born to shorn or unshorn ewes. In newborn lambs, plasma growth hormone concentration was lower and insulin concentration higher in shorn compared with unshorn groups. In conclusion, chronic cold exposure induced by winter shearing had no effect on brown adipose tissue development in lambs born to well-fed ewes. PMID- 11101278 TI - Renal responses to angiotensin II receptor blockade in ventilated preterm newborn lambs. AB - Renal and cardiovascular immaturity has been linked with poor outcomes in the premature human newborn. Despite extensive study in the fetus, the contribution of the renin-angiotensin system to renal and cardiovascular function in the premature newborn has not been well characterized. To evaluate the angiotensin II contribution to preterm newborn renal and cardiovascular functions, preterm (120 day) and near-term (136-day) lambs were Caesarean delivered and ventilated. One hour following delivery, animals were randomized to receive angiotensin II receptor-blockade (saralasin; 20 microg kg(-1) min(-1)) or saline (CON). Prior to blockade, mean +/- SEM values for urine flow (U(Flow)), urinary sodium excretion (U(NaV)), and fractional excretion of sodium (FENa) were similar in all groups. Angiotensin II receptor-blockade decreased U(Flow), U(NaV) and FENa in the 120 day group with no changes in the 136-day animals. No changes in mean arterial pressure, or plasma angiotensin II, aldosterone, and renin activity levels were noted at either gestational age. CONCLUSIONS: (1) angiotensin II contributes to the regulation of renal function in 120-day preterm lambs without changing blood pressure and (2) angiotensin II-mediated feedback inhibition of renin release is uncoupled in preterm newborns. PMID- 11101279 TI - Role of developmental factors in the switch from pyruvate to glucose as the major exogenous energy substrate in the preimplantation mouse embryo. AB - Preimplantation mouse embryos, cultured in vitro and those freshly flushed from the reproductive tract, exhibit a switch in energy substrate preference, from pyruvate during the early preimplantation stages, to glucose at the blastocyst stage. Although the biochemical basis of this phenomenon is quite well characterized, its timing and possible association with developmental factors have not been considered. We have therefore examined the role of five developmental factors in determining the timing of the switch, namely: (1) embryo age (in hours post hCG); (2) developmental stage; (3) cytokinesis; (4) cell number; and (5) activation of the embryonic genome. One-cell embryos, which develop more slowly than 2-cell embryos in vitro, were used to investigate the role of embryo age and developmental stage. Cytochalasin D, which inhibits cytokinesis and delays the timing of compaction and cavitation, was used to investigate the role of cell division and developmental stage. Finally, transcription of the embryonic genome was examined with the inhibitor, alpha amanitin. Pyruvate and glucose consumption by single embryos were measured using a noninvasive ultramicrofluorometric technique. The results showed that the timing of the switch in energy substrate preference is precisely regulated in the mouse preimplantation embryo. Activation of the embryonic genome is a prerequisite for the switch and its timing is closely associated with developmental stage, specifically compaction and/or cavitation. Cell number, cytokinesis and embryo age appeared to be unrelated to the timing of the switch. These conclusions may well be extrapolated to other species, since an increase in net glucose uptake, if not always at the expense of pyruvate, is a feature of preimplantation embryo metabolism in all mammals studied. PMID- 11101280 TI - Effects of progesterone on expression of messenger RNA encoding oxytocin neurophysin, oxytocin receptor and prostaglandin G/H synthase-1 and -2 during the early oestrous cycle in the ovine corpus luteum. AB - This study was conducted to determine whether early progesterone treatment plays a role in the regulation of messenger RNA (mRNA) expression for oxytocin neurophysin, oxytocin receptor, prostaglandin G/H synthase (PGHS)-1 and PGHS-2 in the ovine corpus luteum. The expression of ovarian oxytocin, oxytocin receptor, PGHS-1 and PGHS-2 mRNA was investigated in control, progesterone- or RU486 treated ewes. Fifteen ewes were randomly assigned to three groups to receive intramuscular injections of progesterone (12.5 mg; n = 5), RU486, (2.5 mg kg(-1) bodyweight; n = 4) or corn oil (1 mL; n = 6) twice daily from Day 1 to Day 3 post oestrus. On the morning of Day 4 post oestrus, the corpora lutea were collected and analysed for oxytocin-neurophysin mRNA by Northern blot using a labelled cDNA probe, and for the expressions of the oxytocin receptor, PGHS-1 and PGHS-2 mRNA using the reverse transcription polymerase chain reaction. Administration of progesterone or suppression of progesterone activity with RU486 did not affect expression of oxytocin-neurophysin mRNA (P>0.05). Pretreatment of the ewes with progesterone resulted in the enhancement of luteal oxytocin receptor mRNA expression and suppression of PGHS-1 and PGHS-2 mRNA (P<0.001). These results indicate that early progesterone treatment does not control the expression of oxytocin-neurophysin mRNA in the ovine ovary but may be involved in the regulation of ovarian oxytocin receptor and PGHS expression. It is proposed, on the basis of these results, that progesterone may play a role in premature corpus luteum regression through an intra-ovarian mechanism involving the induction of ovarian oxytocin receptor mRNA expression. PMID- 11101281 TI - Role of the MAPK cascade in mammalian germ cells. AB - The mitogen-activated protein kinase (MAPK) cascade is one of the most important of the intracellular signaling pathways that play a crucial role in cell proliferation, cell differentiation and cell cycle regulation. Since the first report in 1993 of MAPK's involvement in the functional regulation of mammalian oocytes, much work has been done on the role of the MAPK cascade in germ cells in different species of mammals. This review describes the possible involvement of the MOS/MEK/MAPK/RSK cascade in spermatogenesis, sperm function, oocyte meiotic re-initiation, spindle assembly, metaphase II arrest, pronuclear formation and the entry of first mitosis, as well as the cross-talk of this cascade to maturation-promoting factor (MPF) and other signal molecules in mammals. PMID- 11101282 TI - Improved fertility through superoxide removal by Toki-shakuyaku-san in mice--a preliminary study. AB - The effect of a superoxide dismutase inhibitor on pregnancy rate and fetal survival, and the effect of concomitant treatment with Toki-shakuyaku-san, was studied. Twelve week-old IV CS female mice were given diethyldithiocarbamate (DEDC; 0.9, 9, 45, and 225 mg kg(-1)) intraperitioneally, from Day 3 to Day 5 of pregnancy, every 12 h, for a total of six times. Two additional groups received both 45 mg kg DEDC and Toki-shakuyaku-san (50 or 200 mg/100 mL), by water bottle, from 12 h before to 12 h after DEDC administration. For each group, the pregnancy rate, the number of surviving fetuses, and the number of resorbed fetuses was calculated. Pregnancy rates were 95.5, 81.8, 54.5 and 0%, respectively, in the DEDC groups and were 100% in the control group. The fetal resorption rate was 0.9 in the control group, and 1.0, 1.7, and 2.3, respectively, in the 0.9, 9, 45 mg kg(-1) DEDC groups. Administration of 200 mg Toki-shakuyaku-san significantly (P<0.05) improved the pregnancy rate to 81.8%, and improved the degree of reduced intercellular interactions and insufficient decidual reactions found on Day 5 in the endometrium of the DEDC groups. Toki-shakuyaku-san reduces the adverse effects of excess superoxides on the endometrium during implantation. PMID- 11101283 TI - Activation of in vivo- and in vitro-derived porcine oocytes by using multiple electrical pulses. AB - The current protocols used to activate pig nuclear transfer embryos are less efficient than those used for other species. To address this problem, the effect of multiple sets of electrical pulses on the parthenogenetic development of in vivo- and in vitro-derived porcine oocytes was examined. Each set of pulses consisted of two 1.5 kV cm(-1) DC pulses of 60 micros duration each, administered 1 s apart. For in vivo-derived oocytes, application of a second set of pulses 30 min after the first set increased the proportion of oocytes that developed to the blastocyst stage compared with a single treatment (51 v. 34%). Application of a third set of pulses 30 min after the second set reduced the rate of blastocyst formation compared with two sets of pulses. In contrast, the rate of blastocyst formation was greater with one set of pulses compared with two sets for in vitro matured oocytes (31 v. 16%). Additional sets of electrical pulses did not affect the number of cells in blastocysts obtained from either group of oocytes compared with a single treatment. In summary, the study demonstrates that the application of a second set of activating pulses 30 min after the first set is beneficial to in vivo-derived oocytes, but detrimental to in vitro matured oocytes, in terms of their ability to develop parthenogenetically to the blastocyst stage. PMID- 11101284 TI - Expression of bcl-2 and 3-beta hydroxysteroid dehydrogenase protein during oocyte and follicle development in fetal and post-natal pig ovaries. AB - The fetal and post-natal development of the pig ovary involves both proliferation and apoptotic loss of germ cells, follicle formation and growth, and the initiation of oocyte meiotic maturation. The present study measured the expression of the proto-oncogene Bcl-2 immunohistochemically on paraffin sections of pig ovaries to determine its relationship with folliculogenesis on Days 50 and 80 post coitum (p.c.) and on Days 1, 21, and 56 post partum (p.p.). The expression of the steroidogenic enzyme 3beta-hydroxysteroid dehydrogenase (3betaHSD) was used to determine the lineages of the cells forming the ovarian follicles, and the expression of the cell proliferation-associated nuclear antigen Ki-67 was used to determine germ cell proliferation and the initiation of follicle growth. Expression of Ki-67 showed that many oogonia were proliferating on Days 50 and 80 p.c. Granulosa cells were more proliferative on Day 56 p.p. than at any other stage; Ki-67 was expressed in 70% of growing follicles and granulosa cells had a 3% mean staining index per section. Less than 4% of germ cells and follicles had morphological signs of degeneration during the period of the study. Bcl-2 was most abundant on Days 21 p.p. and 56 p.p.; staining was localized to stromal cells among follicles and in small clusters in the cortical medullary junction (CMJ). 3BetaHSD staining on Day 50 p.c. was seen in cords of stromal cells within the medulla of the ovary, and in the stromal cells investing the oogonial nests. On Days 80 p.c., 1 p.p., 21 p.p., and 56 p.p., 3betaHSD was expressed in the granulosa cells of primary or primordial follicles at the CMJ. Production of Bcl-2 by somatic cells may support germ cell and preantral follicle survival. PMID- 11101285 TI - A model for coupling of H(+) and substrate fluxes based on "time-sharing" of a common binding site. AB - Both prokaryotic and eukaryotic cells contain an array of membrane transport systems maintaining the cellular homeostasis. Some of them (primary pumps) derive energy from redox reactions, ATP hydrolysis, or light absorption, whereas others (ion-coupled transporters) utilize ion electrochemical gradients for active transport. Remarkable progress has been made in understanding the molecular mechanism of coupling in some of these systems. In many cases carboxylic residues are essential for either binding or coupling. Here we suggest a model for the molecular mechanism of coupling in EmrE, an Escherichia coli 12-kDa multidrug transporter. EmrE confers resistance to a variety of toxic cations by removing them from the cell interior in exchange for two protons. EmrE has only one membrane-embedded charged residue, Glu-14, which is conserved in more than 50 homologous proteins. We have used mutagenesis and chemical modification to show that Glu-14 is part of the substrate-binding site. Its role in proton binding and translocation was shown by a study of the effect of pH on ligand binding, uptake, efflux, and exchange reactions. The studies suggest that Glu-14 is an essential part of a binding site, which is common to substrates and protons. The occupancy of this site by H(+) and substrate is mutually exclusive and provides the basis of the simplest coupling for two fluxes. PMID- 11101286 TI - Heterodimer formation between superoxide dismutase and its copper chaperone. AB - Copper, zinc superoxide dismutase (SOD1) is activated in vivo by the copper chaperone for superoxide dismutase (CCS). The molecular mechanisms by which CCS recognizes and docks with SOD1 for metal ion insertion are not well understood. Two models for the oligomerization state during copper transfer have been proposed: a heterodimer comprising one monomer of CCS and one monomer of SOD1 and a dimer of dimers involving interactions between the two homodimers. We have investigated protein-protein complex formation between copper-loaded and apo yeast CCS (yCCS) and yeast SOD1 for both wild-type SOD1 (wtSOD1) and a mutant SOD1 in which copper ligand His 48 has been replaced with phenylalanine (H48F SOD1). According to gel filtration chromatography, dynamic light scattering, analytical ultracentrifugation, and chemical cross-linking experiments, yCCS and this mutant SOD1 form a complex with the correct molecular mass for a heterodimer. No higher order oligomers were detected. Heterodimer formation is facilitated by the presence of zinc but does not depend on copper loading of yCCS. The complex formed with H48F-SOD1 is more stable than that formed with wtSOD1, suggesting that the latter is a more transient species. Notably, heterodimer formation between copper-loaded yCCS and wtSOD1 is accompanied by SOD1 activation only in the presence of zinc. These findings, taken together with structural, biochemical, and genetic studies, strongly suggest that in vivo copper loading of yeast SOD1 occurs via a heterodimeric intermediate. PMID- 11101287 TI - Remarkable rate enhancement of orotidine 5'-monophosphate decarboxylase is due to transition-state stabilization rather than to ground-state destabilization. AB - The remarkable rate enhancement of orotidine 5'-phosphate decarboxylase (ODCase) has been attributed to ground-state destabilization (GSD) by desolvation and more recently to GSD by electrostatic stress. Here we reiterate our previous arguments that the GSD mechanisms are not likely to play a major role in enzyme catalysis and analyze quantitatively the origin of the rate enhancement of ODCase. This analysis involves energy considerations and computer simulations. Our energy considerations show that (i) the previously proposed desolvation mechanism is based on an improper reference state; (ii) a nonpolar active site cannot account for the catalytic effect of the enzyme; (iii) the focus on the role of the negatively charged protein residues in the electrostatic stress GSD mechanism overlooks the fact that the positively charged Lys72 strongly stabilizes the substrate; (iv) although the previous calculation of the actual enzymatic reaction correctly reproduced the observed rate enhancement, it could not obtain this rate enhancement from the calculated binding energies (which are the relevant quantities for determining GSD effects); (v) the GSD mechanism is inconsistent with the observed binding energy of the phosphoribosyl part of the substrate; and (vi) the presumably unstable substrate (orotate) can be stabilized, at equilibrium, by accepting a proton from the solvent. Our computer simulation studies involve two set of calculations. First, we study the catalytic reaction by using an empirical valence bond potential surface calibrated by ab initio calculations of the reference solution reaction. This calculation reproduces the observed catalytic effect of the enzyme. Next, we use free-energy perturbation calculations and evaluated the electrostatic contributions to the binding energies of the ground state and transition state (TS). These calculations show that the rate enhancement in ODCase is due to the TS stabilization rather than to GSD. The differences between our own and the previous theoretical analyses stem from both the selection of the reacting system and the treatment of the long-range electrostatic contributions to the binding energy. The reacting system was previously assumed to encompass only the orotate. However, this selection does not allow proper description of the reaction catalyzed by the enzyme (i.e., [Orotate(-) + LysH(+)] if [uracil + Lys + CO(2)]). Therefore, the reacting system should include both orotate and the general acid in the form of the protonated Lys72 protein residue. This selection leads to a simple and consistent interpretation of the catalytic effect where the electrostatic stabilization of the transition state is due to the fact that the two negatively charged aspartic residues are already placed near the reactive lysine so that they do not have to reorganize significantly during the reaction. Interestingly, even calculations with only orotate(-) as the reacting system do not produce sufficient destabilization to account for a GSD mechanism. In summary, we conclude, in agreement with previous workers, that ODCase catalyzes its reaction by electrostatic effects. However, we show that these effects are associated with TS stabilization due to a reduction in the protein-protein reorganization energy and not with protein-substrate destabilization effects. PMID- 11101288 TI - Crystallization and the crystal properties of the oxygen-evolving photosystem II from Synechococcus vulcanus. AB - A photosystem II (PSII) complex highly active in oxygen evolution was purified and crystallized from a thermophilic cyanobacterium, Synechococcus vulcanus. The PSII complex in the crystals contained the D1/D2 reaction center subunits, CP47 and CP43 (two chlorophyll-binding core antenna proteins of photosystem II), cytochrome b-559 alpha- and beta-subunits, several low molecular weight subunits, and three extrinsic proteins, that is, 33 and 12 kDa proteins and cytochrome c 550. The PSII complex also retained a high rate of oxygen evolution. The apparent molecular mass of the PSII in the crystals was determined to be 580 kDa by gel filtration chromatography, indicating that the PSII crystallized is a dimer. The crystals diffracted to a maximum resolution of 3.5 A at a cryogenic temperature using X-rays from a synchrotron radiation source, SPring-8. The crystals belonged to an orthorhombic system, and the space group was P2(1)2(1)2(1) with unit cell dimensions of a = 129.7 A, b = 226.5 A, and c = 307.8 A. Each asymmetric unit contained one PSII dimer, which gave rise to a specific volume (V(M)) of 3.6 A(3)/Da based on the calculated molecular mass of 310 kDa for a PSII monomer and an estimated solvent content of 66%. Multiple data sets of native crystals have been collected and processed to 4.0 A, indicating that our crystals are suitable for structure analysis at this resolution. PMID- 11101289 TI - Insights into nucleotide signal transduction in nitrogenase: structure of an iron protein with MgADP bound. AB - Coupling the energy of nucleoside triphosphate binding and hydrolysis to conformational changes is a common mechanism for a number of proteins with disparate cellular functions, including those involved in DNA replication, protein synthesis, and cell differentiation. Unique to this class of proteins is the dimeric Fe protein component of nitrogenase in which the binding and hydrolysis of MgATP controls intermolecular electron transfer and reduction of nitrogen to ammonia. In the work presented here, the MgADP-bound (or "off") conformational state of the nitrogenase Fe protein has been captured and a 2.15 A resolution X-ray crystal structure is presented. The structure described herein reveals likely mechanisms for long-range communication from the nucleotide binding sites for controlling the affinity of association with the MoFe protein component. Two pathways, termed switches I and II, appear to be integral to this nucleotide signal transduction mechanism. In addition, the structure provides the basis for the changes in the biophysical properties of the [4Fe-4S] cluster observed when Fe protein binds nucleotides. The structure of the MgADP-bound Fe protein provides important insights into the respective contributions of nucleotide interaction and complex formation in defining the conformational states that are the keys to nitrogenase catalysis. PMID- 11101290 TI - Three-dimensional solution structure of oryzacystatin-I, a cysteine proteinase inhibitor of the rice, Oryza sativa L. japonica. AB - The three-dimensional structure of oryzacystatin-I, a cysteine proteinase inhibitor of the rice, Oryza sativa L. japonica, has been determined in solution at pH 6.8 and 25 degrees C by (1)H and (15)N NMR spectroscopy. The main body (Glu13-Asp97) of oryzacystatin-I is well-defined and consists of an alpha-helix and a five-stranded antiparallel beta-sheet, while the N- and C-terminal regions (Ser2-Val12 and Ala98-Ala102) are less defined. The helix-sheet architechture of oryzacystatin-I is stabilized by a hydrophobic cluster formed between the alpha helix and the beta-sheet and is considerably similar to that of monellin, a sweet tasting protein from an African berry, as well as those of the animal cystatins studied, e.g., chicken egg white cystatin and human stefins A and B (also referred to as human cystatins A and B). Detailed structural comparison indicates that oryzacystatin-I is more similar to chicken cystatin, which belongs to the type-2 animal cystatins, than to human stefins A and B, which belong to the type 1 animal cystatins, despite different loop length. PMID- 11101291 TI - Three-dimensional solution structure of omega-conotoxin TxVII, an L-type calcium channel blocker. AB - We determined the three-dimensional structure of omega-conotoxin TxVII, a 26 residue peptide that is an L-type calcium channel blocker, by (1)H NMR in aqueous solution. Twenty converged structures of this peptide were obtained on the basis of 411 distance constraints obtained from nuclear Overhauser effect connectivities, 20 torsion angle constraints, and 21 constraints associated with hydrogen bonds and disulfide bonds. The root-mean-square deviations about the averaged coordinates of the backbone atoms (N, C(alpha), C, and O) and all heavy atoms were 0.50 +/- 0.09 A and 0.99 +/- 0.13 A, respectively. The structure of omega-conotoxin TxVII is composed of a triple-stranded antiparallel beta-sheet and four turns. The three disulfide bonds in omega-conotoxin TxVII form the classical cystine knot motif of toxic or inhibitory polypeptides. The overall folding of omega-conotoxin TxVII is similar to those of the N-type calcium channel blockers, omega-conotoxin GVIA and MVIIA, despite the low amino acid sequence homology among them. omega-Conotoxin TxVII exposes many hydrophobic residues to a certain surface area. In contrast, omega-conotoxin GVIA and MVIIA expose basic residues in the same way as omega-conotoxin TxVII. The channel binding site of omega-conotoxin TxVII is different from those of omega-conotoxin GVIA and MVIIA, although the overall folding of these three peptides is similar. The gathered hydrophobic residues of omega-conotoxin TxVII probably interact with the hydrophobic cluster of the alpha(1) subunit of the L-type calcium channel, which consists of 13 residues located in segments 5 and 6 in domain III and in segment 6 in domain IV. PMID- 11101292 TI - Role for lysine 142 in the excision of adenine from A:G mispairs by MutY DNA glycosylase of Escherichia coli. AB - MutY participates in the repair of oxidatively damaged DNA by excising adenine from dA:dG and dA:8-oxodG mispairs; this DNA glycosylase can be cross-linked to DNA through Lys-142. We have investigated the properties of a mutant protein in which Lys-142 is replaced by glutamine. Using the rifampicin resistance assay, MutY K142Q was shown to complement the mutY mutator phenotype to the same extent as wild-type MutY. Although MutY K142Q does not form a Schiff base with DNA, it retains in part the catalytic properties of wild-type enzyme. The K142Q mutation selectively impairs processing of DNA containing dA:dG mispairs but not that of substrates containing dA:8-oxodG. Decreased substrate processing is mediated primarily via an increase in K(D) (21.8 nM for MutY vs 298 nM for MutY K142Q). The catalytic constant, measured in single turnover experiments, was not significantly affected. At pH < 6.0, the activity of MutY K142Q on the dA:dG mispair was approximately the same as for wild-type protein, suggesting that a dG(anti) to dG(syn) transition is effected at low pH. The three-dimensional structure of the catalytic domain of MutY K142Q, determined at 1.35 A resolution, shows no significant differences between wild-type and mutant protein, indicating that Lys-142 is not critical for maintaining the conformation of MutY. We conclude that Lys-142 recognizes guanine in the dA:dG mispair, helping position this residue in the syn conformation and facilitating binding of substrate DNA. Lys-142 is not involved in the catalytic steps of base excision. PMID- 11101293 TI - Crystal structure of human CD69: a C-type lectin-like activation marker of hematopoietic cells. AB - CD69 is a widely expressed type II transmembrane glycoprotein related to the C type animal lectins that exhibits regulated expression on a variety of cells of the hematopoietic lineage, including neutrophils, monocytes, T cells, B cells, natural killer (NK) cells, and platelets. Activation of T lymphocytes results in the induced expression of CD69 at the cell surface. In addition, cross-linking of CD69 by specific antibodies leads to the activation of cells bearing this receptor and to the induction of effector functions. However, the physiological ligand of CD69 is unknown. We report here the X-ray crystal structure of the extracellular C-type lectin-like domain (CTLD) of human CD69 at 2.27 A resolution. Recombinant CD69 was expressed in bacterial inclusion bodies and folded in vitro. The protein, which exists as a disulfide-linked homodimer on the cell surface, crystallizes as a symmetrical dimer, similar to those formed by the related NK cell receptors Ly49A and CD94. The structure reveals conservation of the C-type lectin-like fold, including preservation of the two alpha-helical regions found in Ly49A and mannose-binding protein (MBP). However, only one of the nine residues coordinated to Ca(2+) in MBP is conserved in CD69 and no bound Ca(2+) is evident in the crystal structure. Surprisingly, electron density suggestive of a puckered six-membered ring was discovered at a site structurally analogous to the ligand-binding sites of MBP and Ly49A. This sugar-like density may represent, or mimic, part of the natural ligand recognized by CD69. PMID- 11101294 TI - Characterization of a Rhodobacter capsulatus reaction center mutant that enhances the distinction between spectral forms of the initial electron donor. AB - A large scale mutation of the Rhodobacter capsulatus reaction center M-subunit gene, sym2-1, has been constructed in which amino acid residues M205-M210 have been changed to the corresponding L subunit amino acids. Two interconvertable spectral forms of the initial electron donor are observed in isolated reaction centers from this mutant. Which conformation dominates depends on ionic strength, the nature of the detergent used, and the temperature. Reaction centers from this mutant have a ground-state absorbance spectrum that is very similar to wild-type when measured immediately after purification in the presence of high salt. However, upon subsequent dialysis against a low ionic strength buffer or the addition of positively charged detergents, the near-infrared spectral band of P (the initial electron donor) in sym2-1 reaction centers is shifted by over 30 nm to the blue, from 852 to 820 nm. Systematically varying either the ionic strength or the amount of charged detergent reveals an isobestic point in the absorbance spectrum at 845 nm. The wild-type spectrum also shifts with ionic strength or detergent with an isobestic point at 860 nm. The large spectral separation between the two dominant conformational forms of the sym2-1 reaction center makes detailed measurements of each state possible. Both of the spectral forms of P bleach in the presence of light. Electrochemical measurements of the P/P+ midpoint potential of sym2-1 reaction centers show an increase of about 30 mV upon conversion from the long-wavelength form to the short-wavelength form of the mutant. The rate constant of initial electron transfer in both forms of the mutant reaction centers is essentially the same, suggesting that the spectral characteristics of P are not critical for charge separation. The short-wavelength form of P in this mutant also converts to the long-wavelength form as a function of temperature between room temperature and 130 K, again giving rise to an isobestic point, in this case at 838 nm for the mutant. A similar, though considerably less pronounced spectral change with temperature occurs in wild-type reaction centers, with an isobestic point at about 855 nm, close to that found by titrating with ionic strength or detergent. Fitting the temperature dependence of the sym2-1 reaction center spectrum to a thermodynamic model resulted in a value for the enthalpy of the conformational interconversion between the short- and long-wavelength forms of about -6 kJ/mol and an entropy of interconversion of about -35 J/(K mol). Similar values of enthapy and entropy changes can be used to model the temperature dependence in wild-type. Thus, much of the temperature dependence of the reaction center special pair near-infrared absorbance band can be described as an equilibrium shift between two spectrally distinct conformations of the reaction center. PMID- 11101295 TI - Voltammetric probes of cytochrome electroreactivity: the effect of the protein matrix on outer-sphere reorganization energy and electronic coupling probed through comparisons with the behavior of porphyrin complexes. AB - Using surface-modified electrodes composed of omega-hydroxyalkanethiols, an experimentally based value for the inner-sphere reorganization energy of the bis(imidazole)iron porphyrin system has been obtained by examining the solvent dependence of the reorganization energy of bis(N-methylimidazole)meso-tetraphenyl iron porphyrin. The value obtained (0.41 +/- 0.06 eV) is remarkably similar to values we have recently reported for the reorganization energy of cytochrome b(5) (0.43 +/- 0.02 eV) and cytochrome c (0.58 +/- 0.06 eV). This strongly suggests that the protein matrix mimics the behavior of a low dielectric solvent and effectively shields the heme from the solvent. The effect of the orientation of the heme relative to the electrode was also explored by sytematically varying the steric bulk of the axial ligands. On the basis of a good linear correlation between the electronic coupling and the cosine of the angle between the heme plane and the surface of the electrode, it is suggested that a parallel orientation of the heme yields a maximum in the electronic coupling. Relevance to interheme protein electron transfer is discussed. PMID- 11101296 TI - Direct voltammetric investigation of the electrochemical properties of human hemoglobin: relevance to physiological redox chemistry. AB - Voltammetric measurements on solutions of human hemoglobin using gold electrodes modified with omega-hydroxyalkanethiols have yielded the first direct measure of the reorganization energy of the protein. The value obtained based on extrapolation of the experimentally measured currents, 0.76 eV, is independent of pH (i.e., over the physiologically relevant rage, pH 6.8-7.4) and is remarkably similar to values obtained for myoglobin. This result is perhaps surprising given the marked dependence of the measured reduction potential of hemoglobin on pH (i.e., the redox Bohr effect). Electron transfer rates from the electrode to hemoglobin were also measured. Using similarly measured heterogeneous electron transfer rates for cytochrome b(5), it is possible to predict the magnitude of the homogeneous electron-transfer rate from cytochrome b(5) to methemoglobin using a formalism developed by Marcus. These predicted rates are in reasonable agreement with reported rates of this physiological reaction based on stopped flow kinetics experiments. These results suggest that the intrinsic electroreactivity of these heme proteins is sufficient to account for physiologically observed rates. Residual differences between homogeneous phase kinetics and those predicted by heterogeneous phase reactions are suggested to be due to small reductions in the outer-sphere reorganization energy of both component proteins which arise due to solvent exclusion at the interface between the two proteins in complex. PMID- 11101297 TI - Heme redox potential control in de novo designed four-alpha-helix bundle proteins. AB - The effects of various mechanisms of metalloporphyrin reduction potential modulation were investigated experimentally using a robust, well-characterized heme protein maquette, synthetic protein scaffold H10A24 [?CH(3)()CONH CGGGELWKL.HEELLKK.FEELLKL.AEERLKK. L-CONH(2)()?(2)](2). Removal of the iron porphyrin macrocycle from the high dielectric aqueous environment and sequestration within the hydrophobic core of the H10A24 maquette raises the equilibrium reduction midpoint potential by 36-138 mV depending on the hydrophobicity of the metalloporphyrin structure. By incorporating various natural and synthetic metalloporphyrins into a single protein scaffold, we demonstrate a 300-mV range in reduction potential modulation due to the electron donating/withdrawing character of the peripheral macrocycle substituents. Solution pH is used to modulate the metalloporphyrin reduction potential by 160 mV, regardless of the macrocycle architecture, by controlling the protonation state of the glutamate involved in partial charge compensation of the ferric heme. Attempts to control the reduction potential by inserting charged amino acids into the hydrophobic core at close proximity to the metalloporphyrin lead to varied success, with H10A24-L13E lowering the E(m8.5) by 40 mV, H10A24-E11Q raising it by 50 mV, and H10A24-L13R remaining surprisingly unaltered. Modifying the charge of the adjacent metalloporphyrin, +1 for iron(III) protoporphyrin IX or neutral for zinc(II) protoporphyrin IX resulted in a loss of 70 mV [Fe(III)PPIX](+) - [Fe(III)PPIX](+) interaction observed in maquettes. Using these factors in combination, we illustrate a 435-mV variation of the metalloporphyrin reduction midpoint potential in a simple heme maquette relative to the about 800-mV range observed for natural cytochromes. Comparison between the reduction potentials of the heme maquettes and other de novo designed heme proteins reveals global trends in the E(m) values of synthetic cytochromes. PMID- 11101298 TI - Preferential binding of equine ferricytochrome c to the bacterial photosynthetic reaction center from Rhodobacter sphaeroides. AB - Redox titration of horse heart cytochrome c (cyt c), in the presence of varying concentrations of detergent-solubilized photosynthetic reaction center (RC) from Rhodobacter sphaeroides, revealed an RC concentration-dependent decrease in the measured cyt c midpoint potential that is indicative of a 3.6 +/- 0.2-fold stronger binding affinity of oxidized cytochrome to a single binding site. This effect was correlated with preferential binding in the functional complex by redox titration of the fraction of RCs exhibiting microsecond, first-order, special pair reduction by cytochrome. A binding affinity ratio of 3.1 +/- 0.4 was determined by this second technique, confirming the result. Redox titration of flash-induced intracomplex electron transfer also showed the association in the electron transfer-active complex to be strong, with a dissociation constant of 0.17 +/- 0.03 microM. The tight binding is associated with a slow off-rate which, in the case of the oxidized form, can influence the kinetics of P(+) reduction. The pitfalls of the common use of xenon flashlamps to photoexcite fast electron transfer reactions are discussed with relation to the first electron transfer from primary to secondary RC quinone acceptors. The results shed some light on the diversity of kinetic behavior reported for the cytochrome to RC electron transfer reaction. PMID- 11101299 TI - Mutations in the CD-loop region of the D2 protein in Synechocystis sp. PCC 6803 modify charge recombination pathways in photosystem II in vivo. AB - The lumenal CD-loop region of the D2 protein of photosystem II contains residues that interact with the primary electron donor P680 and the redox active tyrosyl residue Y(D). Photosystem II properties were studied in a number of photoautotrophic mutants of Synechocystis sp. PCC 6803, most of which carried combinatorial mutations in residues 164-170, 179-186, or 187-194 of the D2 protein. To facilitate characterization of photosystem II properties in the mutants, the CD-loop mutations were introduced into a photosystem I-less background. According to variable fluorescence decay measurements in DCMU-treated cells, charge recombination of Q(A)(-) with the donor side was faster in the majority of mutants (t(1/2) = 45-140 ms) than in the control (t(1/2) = 180 ms). However, in one mutant (named C7-3), the decay of Q(A)(-) was 2 times slower than in the control (t(1/2) = 360 ms). The decay half-time of each mutant correlated with the yield of the Q-band of thermoluminescence (TL) emitted due to S(2)Q(A)( ) charge recombination. The C7-3 mutant had the highest TL intensity, whereas no Q-band was detected in the mutants with fast Q(A)(-) decay (t(1/2) = 45-50 ms). The correlated changes in the rate of recombination and in TL yield in these strains suggest the existence of a nonradiative pathway of charge recombination between Q(A)(-) and the donor side. This may involve direct electron transfer from Q(A)(-) to P680(+) in a way not leading to formation of excited chlorophyll. Many mutations in the CD-loop appear to increase the equilibrium P680(+) concentration during the lifetime of the S(2)Q(A)(-) state, for example, by making the midpoint potential of the P680(+)/P680 redox couple more negative. The nonradiative charge recombination pathway involves a low activation energy and is less temperature-dependent than the formation of excited P680 that leads to TL emission. Therefore, during the TL measurements in these mutants, the S(2)Q(A)(-) state can recombine nonradiatively before temperatures are reached at which radiative charge recombination becomes feasible. The results presented here highlight the presence of two charge recombination pathways and the importance of the CD-loop of the D2 protein in determination of the energy gap between the P680(+)S(1) and P680S(2) states. PMID- 11101300 TI - Thermal behavior of proteins: heat-resistant proteins and their heat-induced secondary structural changes. AB - Most proteins are denatured by heat treatment, and the process is usually irreversible. However, some proteins, such as hyperthermophilic proteins are known to be stable even at the boiling temperature of water. We here describe a systematic investigation of thermal behavior of proteins by purifying and characterizing some heat-resistant proteins (HRPs) that are not aggregated upon heat treatment. Although most proteins were precipitated by boiling in a water bath, about 20 and 70 wt % of total proteins appeared to be heat-resistant in Jurkat T-cell lysates and human serum, respectively. We identified major HRPs from Jurkat T-cells and human serum by N-terminal amino acid sequencing and Western blot analysis. HRPs of 20 and 45 kDa (HRP20 and HRP45) were identified as alpha-synuclein and calreticulin, respectively, and HRPs of 60, 27, and 16 kDa (HRP60, HRP27, and HRP16) were identified as human serum fetuin, apolipoprotein A I, and transthyretin, respectively. By a systematic investigation of the effect of heat on the secondary structure of the purified HRPs by circular dichroic spectroscopy, we observed four major types of thermal behavior, suggesting that the proteins could protect themselves through these pathways. Although our analysis is restricted to protein secondary structural changes, our data indicate that heat resistance of protein can be achieved in several different ways depending on the thermodynamic stability of native (N), unfolded (U), denatured (D), and intermediate (I) states. PMID- 11101301 TI - Interaction of a bacterially expressed peptide from the receptor binding domain of Pseudomonas aeruginosa pili strain PAK with a cross-reactive antibody: conformation of the bound peptide. AB - The C-terminal receptor binding region of Pseudomonas aeruginosa pilin protein strain PAK (residues 128-144) has been the target for the design of a vaccine effective against P. aeruginosa infections. We have recently cloned and expressed a (15)N-labeled PAK pilin peptide spanning residues 128-144 of the PAK pilin protein. The peptide exists as a major (trans) and minor (cis) species in solution, arising from isomerization around a central Ile(138)-Pro(139) peptide bond. The trans isomer adopts two well-defined turns in solution, a type I beta turn spanning Asp(134)-Glu-Gln-Phe(137) and a type II beta-turn spanning Pro(139) Lys-Gly-Cys(142). The cis isomer adopts only one well-defined type II beta-turn spanning Pro(139)-Lys-Gly-Cys(142) but displays evidence of a less ordered turn spanning Asp(132)-Gln-Asp-Glu(135). These turns have been implicated in cross reactive antibody recognition. (15)N-edited NMR spectroscopy was used to study the binding of the (15)N-labeled PAK pilin peptide to an Fab fragment of a cross reactive monoclonal antibody, PAK-13, raised against the intact PAK pilus. The results of these studies are as follows: the trans and cis isomers bind with similar affinity to the Fab, despite their different topologies; both isomers maintain the conformational integrity of their beta-turns when bound; binding leads to the preferential stabilization of the first turn over the second turn in each isomer; and binding leads to the perturbation of resonances within regions of the trans and cis backbone that undergo microsecond to millisecond motions. These slow motions may play a role in induced fit binding of the first turn to Fab PAK-13, which would allow the same antibody combining site to accommodate either trans or cis topology. More importantly for vaccine design, these motions may also play a role in the development of a broad-spectrum vaccine capable of generating an antibody therapeutic effective against the multiple strains of P. aeruginosa. PMID- 11101302 TI - Structural and mechanistic investigation of 3-deoxy-D-manno-octulosonate-8 phosphate synthase by solid-state REDOR NMR. AB - 15N?(31)P? REDOR NMR experiments were applied to lyophilized binary complexes of 3-deoxy-D-manno-2-octulosonate-8-phosphate synthase (KDO8PS), with each of its natural substrates, phosphoenolpyruvate (PEP) and arabinose-5-phsophate (A5P), and with a mechanism-based inhibitor (K(i) = 0.4 microM), directly characterizing the active site basic residues involved in the binding of their carboxylate and phosphate moieties. KDO8PS was labeled uniformly with (15)N or [eta-(15)N(2)]Arg, and the ligands were selectively labeled with (13)C and (15)N. The NMR data established that PEP is bound by KDO8PS via a preserved set of structurally rigid and chemically unique Arg and Lys residues, with 5 A (upper limit) between epsilon-(15)N of this Lys and (31)P of PEP. A5P is bound in its cyclic forms to KDO8PS via a different set of Lys and Arg residues. The two sets arise from adjacent subsites that are capable of independent and sufficiently strong binding. The inhibitor is best characterized as an A5P-based substrate analogue inhibitor of KDO8PS. Five mutants in which highly conserved arginines were replaced with alanines were prepared and kinetically characterized. Our solid state NMR observations complement the crystallographic structure of KDO8PS, and in combination with the mutagenesis results enable tentative assignment of the NMR-identified active site residues. Lys-138 and Arg-168 located at the most recessed part of the active site cavity are the chemically distinct and structurally rigid residues that bind PEP phosphate; R168A resulted in 0.1% of wild-type activity. Arg-63, exposed at the opening of the active site barrel, is the flexible residue with a generic chemical shift that binds A5P; R63A resulted in complete deactivation. The mechanistic implications of our results are discussed. PMID- 11101303 TI - Participation of Na,K-ATPase in FGF-2 secretion: rescue of ouabain-inhibitable FGF-2 secretion by ouabain-resistant Na,K-ATPase alpha subunits. AB - We have examined the relationship between Na,K-ATPase and FGF-2 secretion in transfected primate cells. FGF-2 lacks a classic hydrophobic export signal, and the mechanisms mediating its secretion are unknown. To monitor secretion, a FLAG epitope tag was inserted into the carboxyl terminus of the 18 kDa form of human FGF-2, and the construct was transfected into either human HEK 293 or monkey CV-1 cells. Exported FGF-2 was detected in the culture medium using the FLAG-specific monoclonal antibody M2. FGF-2 secretion from HEK 293 or CV-1 cells was linear over time and sensitive to inhibition by the cardiac glycoside ouabain, a specific inhibitor of the Na,K-ATPase. In contrast, the secretion of FGF-8 (an FGF family member that contains a hydrophobic secretory signal) was not inhibited by treatment of HEK 293 or CV-1 cells with ouabain. FGF-2 secretion was also assayed in CV-1 cells expressing the naturally ouabain-resistant rodent Na,K ATPase alpha1 subunit. In cells expressing the rodent alpha1 subunit, FGF-2 secretion was unaffected by high levels of ouabain, indicating that the rodent alpha1 subunit was capable of rescuing ouabain-inhibitable FGF-2 export. Expression of ouabain-resistant mutants of the rodent alpha2 and alpha3 subunits, or the naturally ouabain-resistant rodent alpha4 subunit, also supported FGF-2 secretion in ouabain-treated cells. Taken together, our studies are consistent with the idea that the Na,K-ATPase plays a prominent role in regulating FGF-2 secretion, although none of the alpha subunit isoforms exhibited specificity with regard to FGF-2 export. PMID- 11101305 TI - Deletions in the N-terminal segment of the plasma membrane Ca(2+) pump impair the expression of a correctly folded functional enzyme. AB - Mutant cDNAs encoding h4 plasma membrane Ca(2+) pumps with deletions in the N terminal segment have been constructed and expressed in COS cells. As judged by immunoblotting, each construct was expressed at a high level similar to that of the wild-type enzyme. The removal of the first six amino acids had no effect on the Ca(2+) transport activity, but deletions in the segment 15-75 reduced the activity to undetectable levels. The d(43-56)h4 mutant, lacking amino acids 43 56, was also efficiently expressed in stable form in CHO cells. The Ca(2+) transport activity of d(43-56)h4 in this system was about 40% of that of the wild type. The d(43-56)h4 enzyme exhibited a similar affinity for Ca(2+), a slightly increased apparent affinity for ATP, and a slightly lower sensitivity to inhibition by vanadate than the wild-type enzyme. Analysis of the phosphoenzyme intermediate formed in the presence of lanthanum showed that the phosphorylation reaction was not affected, but the maximum amount of phosphoenzyme was reduced to the same extent as the Ca(2+) transport activity. These results suggest that the expressed d(43-56)h4 was a mixture of fully active and inactive enzyme. The d(43 56)h4 enzyme was more easily degraded by proteases and had a higher sensitivity to heat inactivation than the wild type suggesting that the loss of function was due to the improper folding and instability of the mutant protein. On the basis of these findings, it appears that the N-terminal segment of the plasma membrane Ca(2+) pump is neither essential for synthesis nor for catalytic activity but is critical for the expression of a correctly folded functional enzyme. PMID- 11101304 TI - Interaction of collagen-like peptide models of asymmetric acetylcholinesterase with glycosaminoglycans: spectroscopic studies of conformational changes and stability. AB - The effect of heparin on the conformation and stability of triple-helical peptide models of the collagen tail of asymmetric acetylcholinesterase expands our understanding of heparin interactions with proteins and presents an opportunity for clarifying the nature of binding of ligands to collagen triple-helix domains. Within the collagen tail of AChE, there are two consensus sequences for heparin binding of the form BBXB, surrounded by additional basic residues. Circular dichroism studies were used to determine the effect of the addition of increasing concentrations of heparin on triple-helical peptide models for the heparin binding domains, including peptides in which the basic residues within and surrounding the consensus sequence were replaced by alanine residues. The addition of heparin caused an increased triple-helix content with saturation properties for the peptide modeling the C-terminal site, while precipitation, with no increased helix content resulted from heparin addition to the peptide modeling the N-terminal site. The results suggest that the two binding sites with a similar triple-helical conformation have distinctive ways of interacting with heparin, which must relate to small differences in the consensus sequence (GRKGR vs GKRGK) and in the surrounding basic residues. Addition of heparin increased the thermal stability of all peptides containing the consensus sequence. Heparan sulfate produced conformational and stabilization effects similar to those of heparin, while chondroitin sulfate led to a cloudy solution, loss of circular dichroism signal, and a smaller increase in thermal stability. Thus, specificity in both the sequence of the triple helix and the type of glycosaminoglycan is required for this interaction. PMID- 11101306 TI - Molecular determinants of ligand binding to the human melanocortin-4 receptor. AB - To elucidate the molecular basis for the interaction of ligands with the human melanocortin-4 receptor (hMC4R), agonist structure-activity studies and receptor point mutagenesis were performed. Structure-activity studies of [Nle(4), D Phe(7)]-alpha-melanocyte stimulating hormone (NDP-MSH) identified D-Phe7-Arg8 Trp9 as the minimal NDP-MSH fragment that possesses full agonist efficacy at the hMC4R. In an effort to identify receptor residues that might interact with amino acids in this tripeptide sequence 24 hMC4R transmembrane (TM) residues were mutated (the rationale for choosing specific receptor residues for mutation is outlined in the Results section). Mutation of TM3 residues D122 and D126 and TM6 residues F261 and H264 decreased the binding affinity of NDP-MSH 5-fold or greater, thereby identifying these receptor residues as sites potentially involved in the sought after ligand-receptor interactions. By examination of the binding affinities and potencies of substituted NDP-MSH peptides at receptor mutants, evidence was found that core melanocortin peptide residue Arg8 interacts at a molecular level with hMC4R TM3 residue D122. TM3 mutations were also observed to decrease the binding of hMC4R antagonists. Notably, mutation of TM3 residue D126 to alanine decreased the binding affinity of AGRP (87-132), a C terminal derivative of the endogenous melanocortin antagonist, 8-fold, and simultaneous mutations D122A/D126A completely abolished AGRP (87-132) binding. In addition, mutation of TM3 residue D122 or D126 decreased the binding affinity of hMC4R antagonist SHU 9119. These results provide further insight into the molecular determinants of hMC4R ligand binding. PMID- 11101307 TI - Insulin regulation of beta-cell function involves a feedback loop on SERCA gene expression, Ca(2+) homeostasis, and insulin expression and secretion. AB - The insulin receptor signaling pathway is present in beta-cells and is believed to be important in beta-cell function. We show here that insulin directly regulates beta-cell function in isolated rodent islets. Long-term insulin treatment caused a sustained increase in [Ca(2+)](i) and enhanced glucose stimulated insulin secretion in rat islets, but failed to increase insulin content. Chronic activation of insulin receptor signaling by IRS-1 overexpression in the beta-cell inhibited gene expression of SERCA3, an endoplasmic reticulum Ca(2+)-ATPase. Insulin gene transcription was stimulated by insulin receptor signaling and insulin mimetic compound (L-783 281) in a glucose- and Grb2 dependent manner. Thus, beta-cell SERCA3 is a target for insulin regulation, which implies that beta-cell Ca(2+) homeostasis is regulated in an autocrine feedback loop by insulin. This study identifies a novel regulatory pathway of insulin secretion at the molecular level with two main components: (1) regulation of intracellular Ca(2+) homeostasis via SERCA3 and (2) regulation of insulin gene expression. PMID- 11101308 TI - The n-alcohol site in the nicotinic receptor pore is a hydrophobic patch. AB - Alcohols and volatile anesthetics inhibit peripheral nicotinic acetylcholine receptors noncompetitively, primarily via an open-channel block mechanism. Analysis of hydrophobic mutations near the middle of the pore-forming M2 domains suggested that alcohols interact with the pore in this vicinity. To establish the extent of this inhibitory site, we created a series of hydrophobicity-altering mutations scanning most of the alpha subunit M2 domain. Using both single-channel and rapid patch perfusion electrophysiology, we measured how these mutations affect nAChR sensitivity to ethanol and hexanol. We find a near-contiguous series of amino acids in alpha-M2, extending from alphaL250 (8') to alphaV255 (13'), where mutagenesis strongly influences inhibition by alcohols. These results support the existence of a large inhibitory patch in the nAChR pore lining where interactions with alcohols are primarily due to hydrophobic forces. Ethanol appears to interact with deeper regions of this site than does hexanol. Because alcohols apparently act as open-channel blockers, we infer from our results that most of the residues between alphaL250 and alphaV255 are exposed to the aqueous environment of the pore when the channel is open. The location and extent of this site can explain why small alcohols occupy the nAChR pore at the same time as larger alcohols or charged blockers, while two large alcohols bind in a mutually exclusive manner. PMID- 11101309 TI - Intrinsic fluorescence of the P-glycoprotein multidrug transporter: sensitivity of tryptophan residues to binding of drugs and nucleotides. AB - P-glycoprotein is a member of the ATP binding cassette family of membrane proteins, and acts as an ATP-driven efflux pump for a diverse group of hydrophobic drugs, natural products, and peptides. The side chains of aromatic amino acids have been proposed to play an important role in recognition and binding of substrates by P-glycoprotein. Steady-state and lifetime fluorescence techniques were used to probe the environment of the 11 tryptophan residues within purified functional P-glycoprotein, and their response to binding of nucleotides and substrates. The emission spectrum of P-glycoprotein indicated that these residues are present in a relatively nonpolar environment, and time resolved experiments showed the existence of at least two lifetimes. Quenching studies with acrylamide and iodide indicated that those tryptophan residues predominantly contributing to fluorescence emission are buried within the protein structure. Only small differences in Stern-Volmer quenching constants were noted on binding of nucleotides and drugs, arguing against large changes in tryptophan accessibility following substrate binding. P-glycoprotein fluorescence was highly quenched on binding of fluorescent nucleotides, and moderately quenched by ATP, ADP, and AMP-PNP, suggesting that the site for nucleotide binding is located relatively close to tryptophan residues. Drugs, modulators, hydrophobic peptides, and nucleotides quenched the fluorescence of P-glycoprotein in a saturable fashion, allowing estimation of dissociation constants. Many compounds exhibited biphasic quenching, suggesting the existence of multiple drug binding sites. The quenching observed for many substrates was attributable largely to resonance energy transfer, indicating that these compounds may be located close to tryptophan residues within, or adjacent to, the membrane-bound domains. Thus, the regions of P-glycoprotein involved in nucleotide and drug binding appear to be packed together compactly, which would facilitate coupling of ATP hydrolysis to drug transport. PMID- 11101310 TI - Multisite mutagenesis of interleukin 5 differentiates sites for receptor recognition and receptor activation. AB - Multisite mutagenesis of single-chain and monomeric forms of human interleukin 5 (IL-5) was performed to investigate mechanistic features of receptor activation and the possibility of differentiating sites of activation from those for receptor interaction. The normally dimeric human IL-5 contains two domains, each containing a four-helix bundle. IL-5 has previously been re-engineered into the monomeric, one-domain GM1 form by introducing an eight-residue linker between the third and fourth helices. In this study, we tested a combination of mutations in a single-chain IL-5 (scIL-5) construct, [(89)SLRGG(92),W(110)/(89)AAAAA(92), A(110)]scIL-5. This mutein was found to retain substantial IL-5 receptor alpha chain binding but with selectively suppressed proliferation of the IL-5-dependent cell line TF-1.28. This result confirms recent findings that IL-5 receptor alpha chain recognition can be supported by the (89)SLRGG(92) epitope and that, in contrast, Glu110 is important in receptor activation. On the basis of this result, two mutants of GM1 were constructed with the intent to retain receptor alpha-chain binding while modifying receptor activation epitopes. In the first, [(88)SLRGG(92),W(110)]GM1, the wild-type CD-loop sequence (89)EERRR(92) was converted to the mimotope (89)SLRGG(92), and Glu110 to Trp. In the second, [A(13), A(110)]GM1, wild-type Glu13, and Glu110 were both mutated to Ala. GM1 and mutants were expressed in high yield in Escherichia coli, purified under denaturing conditions from inclusion bodies, and refolded. Monomers were screened for binding to shIL-5Ralpha-Fc using optical biosensor and ELISA and for bioactivity by proliferation of TF-1.28 cells. Both [(88)SLRGG(92),W(110)]GM1 and [A(13),A(110)]GM1 were found to interact with the shIL-5Ralpha-Fc, with affinities of 69-585 nM, 2-15-fold weaker than that of the original GM1. The mutants also were able to compete with IL-5 for binding to shIL-5Ralpha in an ELISA. In contrast, both mutants exhibited a disproportionately decreased capacity to stimulate TF-1. 28 cell proliferation. [A(13),A(110)]GM1 bioactivity was 160-fold lower than that of GM1, while that for the [(88)SLRGG(92),W(110)]GM1 mutant was 2600-fold lower. The largely retained IL-5 receptor alpha-chain binding affinities versus relatively suppressed bioactivities of [A(13),A(110)]GM1 and [(88)SLRGG(92),W(110)]GM1 variants, in particular the latter, point to the existence of separable IL-5 epitopes for receptor binding and activation and establish the potential to design smaller IL-5 mimetic antagonists. PMID- 11101311 TI - Probing the three-dimensional structure of human calreticulin. AB - Calreticulin (CRT) is an abundant soluble protein of the endoplasmic reticulum lumen that functions as a molecular chaperone for nascent glycoproteins. We have probed the three-dimensional structure of human CRT using a series of biochemical and biophysical approaches in an effort to understand the molecular basis of its chaperone function. Sedimentation analysis and chemical cross-linking experiments showed that CRT is monodisperse and monomeric in solution with a molecular mass (MW) of 46 +/- 1 kDa. This MW value together with a sedimentation coefficient, s(o)(20,w), of 2.71 S yielded a frictional ratio, f/f(0), of 1.65. Assuming CRT to be a prolate ellipsoid, we calculated an apparent length of 29.8 nm and diameter of 2.44 nm consistent with an asymmetric elongated molecule. These hydrodynamic dimensions account for the apparent anomalous elution position of CRT on gel filtration columns. Far-UV circular dichroism experiments showed that CRT has a cooperative thermal denaturation transition with a midpoint temperature of 42.5 degrees C suggesting a marginally stable structure. Proteolysis experiments showed that the highly acidic segment at the C-terminus of CRT is most susceptible to digest, consistent with the absence of a well-defined polypeptide backbone structure in this region of the protein. Temperature dependent proteolysis with thermolysin revealed a stable core region within the N and P-domains. A stable fragment encompassing most of the P-domain was also identified in the thermolytic mixture. Collectively, our results suggest that CRT is likely to be a flexible molecule in solution which may be important for its chaperone function. PMID- 11101312 TI - Amino acid propensities for the collagen triple-helix. AB - Determination of the tendencies of amino acids to form alpha-helical and beta sheet structures has been important in clarifying stabilizing interactions, protein design, and the protein folding problem. In this study, we have determined for the first time a complete scale of amino acid propensities for another important protein motif: the collagen triple-helix conformation with its Gly-X-Y repeating sequence. Guest triplets of the form Gly-X-Hyp and Gly-Pro-Y are used to quantitate the conformational propensities of all 20 amino acids for the X and Y positions in the context of a (Gly-Pro-Hyp)(8) host peptide. The rankings for both the X and Y positions show the highly stabilizing nature of imino acids and the destabilizing effects of Gly and aromatic residues. Many residues show differing propensities in the X versus Y position, related to the nonequivalence of these positions in terms of interchain interactions and solvent exposure. The propensity of amino acids to adopt a polyproline II-like conformation plays a role in their triple-helix rankings, as shown by a moderate correlation of triple-helix propensity with frequency of occurrence in polyproline II-like regions. The high propensity of ionizable residues in the X position suggests the importance of interchain hydrogen bonding directly or through water to backbone carbonyls or hydroxyprolines. The low propensity of side chains with branching at the C(delta) in the Y position supports models suggesting these groups block solvent access to backbone C=O groups. These data provide a first step in defining sequence-dependent variations in local triple helix stability and binding, and are important for a general understanding of side chain interactions in all proteins. PMID- 11101313 TI - DNA binding and alkylation by the "left half" of azinomycin B. AB - Azinomycin B (also known as carzinophilin A) contains two electrophilic functional groups-an epoxide and an aziridine residue-that react with nucleophilic sites in duplex DNA to form cross-links at 5'-dGNT and 5'-dGNC sequences. Although the aziridine residue of azinomycin is undoubtedly required for cross-link formation, analogues containing an intact epoxide group but no aziridine residue retain significant biological activity. Azinomycin epoxide analogues (e.g., 5 and 6) are of interest due to their potent biological activity and because there is evidence that azinomycin may decompose in vivo to yield such compounds. To investigate the chemical events underlying the toxicity of azinomycin epoxides, DNA binding and alkylation by synthetic analogues of azinomycin B (6, 8, and 9) that comprise the naphthalene-containing "left half" of the antibiotic have been investigated. The epoxide-containing analogue of azinomycin (6) efficiently alkylates guanosine residues in duplex DNA. DNA alkylation by 6 is facilitated by noncovalent binding of the compound to the double helix. The results of UV-vis absorbance, fluorescence spectroscopy, DNA winding, viscometry, and equilibrium dialysis experiments indicate that the naphthalene group of azinomycin binds to DNA via intercalation. Equilibrium dialysis experiments provide an estimated binding constant of (1.3 +/- 0.3) x 10(3) M(-)(1) for the association of a nonalkylating azinomycin analogue (9) with duplex DNA. The DNA-binding and alkylating properties of the azinomycin epoxide 6 provide a basis for understanding the cytotoxicity of azinomycin analogues which contain an epoxide residue but no aziridine group and may provide insight into the mechanisms by which azinomycin forms interstrand DNA cross-links. PMID- 11101314 TI - Structural studies on some dityrosine-cross-linked globular proteins: stability is weakened, but activity is not abolished. AB - We have carried out conformational and stability studies on three proteins that have previously been shown to undergo dityrosine (DT) cross-linking. They include the monomers and dimers of DT-cross-linked calmodulin and the dimers of bovine pancreatic ribonuclease A and bovine eye lens gammaB-Crystallin. In each of these cases, we find the secondary and tertiary structure of the parent protein to be largely maintained. The DT dimer is, however, weaker than the parent. In this sense, the properties of these DT dimers are somewhat similar to those of glutaraldehyde-cross-linked protein crystals. In contrast, the intramolecularly DT-linked monomeric protein that we studied (DT monomer of calmodulin) is seen to have suffered greater changes in its conformation and stability. These results gain significance in light of the growing identification of DT formation as a marker of oxidative stress, aging, and disease. PMID- 11101315 TI - Removal of hydantoin products of 8-oxoguanine oxidation by the Escherichia coli DNA repair enzyme, FPG. AB - An intriguing feature of 7,8-dihydro-8-oxo-2'-deoxyguanosine (OG) is that it is highly reactive toward further oxidation. Indeed, OG has been shown to be a "hot spot" for oxidative damage and susceptible to oxidation by a variety of cellular oxidants. Recent work has identified two new DNA lesions, guanidinohydantoin (Gh) and spiroiminodihydantoin (Sp), resulting from one-electron oxidation of OG. The presence of Gh and Sp lesions in DNA templates has been shown to result in misinsertion of G and A by DNA polymerases, and therefore, both are potentially mutagenic DNA lesions. The base excision repair (BER) glycosylases Fpg and MutY serve to prevent mutations associated with OG in Escherichia coli, and therefore, we have investigated the ability of these two enzymes to process DNA duplex substrates containing the further oxidized OG lesions, Gh and Sp. The Fpg protein, which removes OG and a variety of other oxidized purine base lesions, was found to remove Gh and Sp efficiently opposite all four of the natural DNA bases. The intrinsic rate of damaged base excision by Fpg was measured under single-turnover conditions and was found to be highly dependent upon the identity of the base opposite the OG, Gh, or Sp lesion; as expected, OG is removed more readily from an OG:C- than an OG:A-containing substrate. However, when adenine is paired with Gh or Sp, the rate of removal of these damaged lesions by Fpg was significantly increased relative to the rate of removal of OG from an OG:A mismatch. The adenine glycosylase MutY, which removes misincorporated A residues from OG:A mismatches, is unable to remove A paired with Gh or Sp. Thus, the activity of Fpg on Gh and Sp lesions may dramatically influence their mutagenic potential. This work suggests that, in addition to OG, oxidative products resulting from further oxidation of OG should be considered when evaluating oxidative DNA damage and its associated effects on DNA mutagenesis. PMID- 11101316 TI - Nitrite inhibition of Vitreoscilla hemoglobin (VHb) in recombinant E. coli: direct evidence that VHb enhances recombinant protein production. AB - Bacteria engineered with the gene (vgb) encoding Vitreoscilla hemoglobin (VHb) typically produce more protein than unengineered cells, and it has generally been assumed that VHb is responsible for this effect. Here, using matched strains of E. coli that bear a recombinant alpha-amylase gene (MK57) or the alpha-amylase gene and vgb (MK79), we provide evidence supporting this assumption. Sodium nitrite (which is known to inhibit heme proteins) was tested over a range of concentrations regarding effects on growth, alpha-amylase production, respiration, and VHb function in MK57 and MK79. Nitrite concentrations were identified at which respiration of cell membranes was inhibited only slightly and to approximately equal degrees in both strains, while whole cell respiration was inhibited to a greater extent and about twice as much in MK79 as MK57. This suggests that these concentrations inhibit VHb while having a much smaller effect on cytochrome oxidase. Direct measurements of VHb showed, in fact, that the same nitrite concentrations greatly decreased the levels of active (ferrous) and, to a somewhat lesser extent, total (ferrous plus ferric) VHb in MK79. Finally, these same nitrite concentrations reversed the advantage regarding alpha-amylase production of MK79 over MK57 seen at 0 mM nitrite, linking the presence of active VHb with the increase in alpha-amylase production. PMID- 11101317 TI - Directed evolution of metabolically engineered Escherichia coli for carotenoid production. AB - We have previously introduced a reconstructed isoprenoid pathway into Escherichia coli that exhibits amplified biosynthetic flux to geranylgeranyl diphosphate (GGPP), a common isoprenoid precursor. It was shown that GGPP synthase is an important rate-controlling enzyme in this reconstructed isoprenoid pathway. In this investigation, we applied directed evolution to GGPP synthase from Archaeoglobus fulgidus to enable the enhanced production of carotenoids in metabolically engineered E. coli. Eight mutants were isolated, and the best one increased lycopene production by 100%. Among the mutants that were isolated, mutation points were clustered in four "hot regions". The "hottest" region is located in the sequence upstream of the coding region, which presumably improves the expression level of the enzyme. The other three are within the coding sequence and are believed to improve the enzyme-specific activity in E coli. These results demonstrate that modulating both enzymatic expression and specific activity are important for optimizing the metabolic flux distribution. PMID- 11101318 TI - Robustness analysis of the Escherichia coli metabolic network. AB - Genomic, biochemical, and strain-specific data can be assembled to define an in silico representation of the metabolic network for a select group of single cellular organisms. Flux-balance analysis and phenotypic phase planes derived therefrom have been developed and applied to analyze the metabolic capabilities and characteristics of Escherichia coli K-12. These analyses have shown the existence of seven essential reactions in the central metabolic pathways (glycolysis, pentose phosphate pathway, tricarboxylic acid cycle) for the growth in glucose minimal media. The corresponding seven gene products can be grouped into three categories: (1) pentose phosphate pathway genes, (2) three-carbon glycolytic genes, and (3) tricarboxylic acid cycle genes. Here we develop a procedure that calculates the sensitivity of optimal cellular growth to altered flux levels of these essential gene products. The results indicate that the E. coli metabolic network is robust with respect to the flux levels of these enzymes. The metabolic flux in the transketolase and the tricarboxylic acid cycle reactions can be reduced to 15% and 19%, respectively, of the optimal value without significantly influencing the optimal growth flux. The metabolic network also exhibited robustness with respect to the ribose-5-phosphate isomerase, and the ribose-5-phosephate isomerase flux was reduced to 28% of the optimal value without significantly effecting the optimal growth flux. The metabolic network exhibited limited robustness to the three-carbon glycolytic fluxes both increased and decreased. The development presented another dimension to the use of FBA to study the capabilities of metabolic networks. PMID- 11101319 TI - Enhanced production of (R)-1,2-propanediol by metabolically engineered Escherichia coli. AB - 1,2-Propanediol (1,2-PD) is a major commodity chemical currently derived from propylene. Previously, we have demonstrated the production of enantiomerically pure (R)-1,2-propanediol from glucose by an engineered E. coli expressing genes for NADH-linked glycerol dehydrogenase and methylglyoxal synthase. In this work, we investigate three methods to improve 1,2-PD in E. coli. First, we investigated improving the host by eliminating production of a byproduct, lactate. To do this, we constructed strains with mutations in two enzymes involved in lactate production, lactate dehydrogenase and glyoxalase I. (Surprisingly, when mutations were made in its ability to produce lactate, one strain of E. coli [MM294], produced a small amount of 1,2-PD without any added genes.) Second, we constructed a complete pathway to 1,2-PD from the glycolytic intermediate, dihydroxyacetone phosphate. Our previous 1, 2-PD producing strains relied on at least one endogenous E. coli activity and only produced 0.7 g/L of 1,2-PD. The complete pathway involved the coexpression of methylglyoxal synthase (mgs), glycerol dehydrogenase (gldA), and either yeast alcohol dehydrogenase (adhI) or E. coli 1,2-propanediol oxidoreductase (fucO). Third, we investigated bioprocessing improvements by carrying out a fed-batch fermentation with the best engineered strain (expressing mgs, gldA, and fucO). A final titer of 4.5 g/L of (R)-1,2-PD was produced, with a final yield of 0.19 g of 1,2-PD per gram of glucose consumed. This work provides a basis for further strain and process improvement. PMID- 11101320 TI - Softwood forest thinnings as a biomass source for ethanol production: a feasibility study for California. AB - A plan has been put forth to strategically thin northern California forests to reduce fire danger and improve forest health. The resulting biomass residue, instead of being open burned, can be converted into ethanol that can be used as a fuel oxygenate or an octane enhancer. Economic potential for a biomass-to-ethanol facility using this softwood biomass was evaluated for two cases: stand-alone and co-located. The co-located case refers to a specific site with an existing biomass power facility near Martell, California. A two-stage dilute acid hydrolysis process is used for the production of ethanol from softwoods, and the residual lignin is used to generate steam and electricity. For a plant processing 800 dry tonnes per day of feedstock, the co-located case is an economically attractive concept. Total estimated capital investment is approximately $70 million for the co-located plant, and the resulting internal rate of return (IRR) is about 24% using 25% equity financing. A sensitivity analysis showed that ethanol selling price and fixed capital investment have a substantial effect on the IRR. It can be concluded that such a biomass-to-ethanol plant seems to be an appealing proposition for California, if ethanol replaces methyl tert-butyl ether, which is slated for a phaseout. PMID- 11101321 TI - Improved performances and control of beer fermentation using encapsulated alpha acetolactate decarboxylase and modeling. AB - The use of the enzyme alpha-acetolactate decarboxylase allows the acceleration of beer fermentation/maturation because it shunts diacetyl formation, whose elimination is the rate-limiting step of the process. To obtain a cost reduction by using this exogenous enzyme, we propose a new process involving recoverable encapsulated alpha-acetolactate decarboxylase. The performance of traditional and new processes was investigated by a modeling approach. A simple model, focused on alpha-acetolactate and diacetyl profiles during beer fermentation, was set up. The simulated profiles are consistent with literature data. This study shows also that encapsulated alpha-acetolactate decarboxylase allows the acceleration of beer fermentation as efficiently as free alpha-acetolactate decarboxylase. The advantage of immobilized alpha-acetolactate decarboxylase versus free enzyme is that it is recoverable and reusable, which means a process cost reduction. PMID- 11101322 TI - Two-liquid-phase slurry bioreactors to enhance the degradation of high-molecular weight polycyclic aromatic hydrocarbons in soil. AB - High-molecular-weight (HMW) polycyclic aromatic hydrocarbons (PAHs) are pollutants that persist in the environment due to their low solubility in water and their sequestration by soil and sediments. The addition of a water immiscible, nonbiodegradable, and biocompatible liquid, silicone oil, to a soil slurry was studied to promote the desorption of PAHs from soil and to increase their bioavailability. First, the transfer into silicone oil of phenanthrene, pyrene, chrysene, and benzo[a]pyrene added to a sterilized soil (sandy soil with 0.65% total volatile solids) was measured for 4 days in three two-liquid-phase (TLP) slurry systems each containing 30% (w/v) soil but different volumes of silicone oil (2.5%, 7.5%, and 15% [v/v]). Except for chrysene, a high percentage of these PAHs was transferred from soil to silicone oil in the TLP slurry system containing 15% silicone oil. Rapid PAH transfer occurred during the first 8 h, probably resulting from the extraction of nonsolubilized and of poorly sorbed PAHs. This was followed by a period in which a slower but constant transfer occurred, suggesting extraction of more tightly bound PAHs. Second, a HMW PAH degrading consortium was enriched in a TLP slurry system with a microbial population isolated from a creosote-contaminated soil. This consortium was then added to three other TLP slurry systems each containing 30% (w/v) sterilized soil that had been artificially contaminated with pyrene, chrysene, and benzo[a]pyrene, but different volumes of silicone oil (10%, 20%, and 30% [v/v]). The resulting TLP slurry bioreactors were much more efficient than the control slurry bioreactor containing the same contaminated soil but no oil phase. In the TLP slurry bioreactor containing 30% silicone oil, the rate of pyrene degradation was 19 mg L(-)(1) day(-)(1) and no pyrene was detected after 4 days. The degradation rates of chrysene and benzo[a]pyrene in the 30% TLP slurry bioreactor were, respectively, 3.5 and 0.94 mg L(-)(1) day(-)(1). Low degradation of pyrene and no significant degradation of chrysene and benzo[a]pyrene occurred in the slurry bioreactor. This is the first report in which a TLP system was combined with a slurry system to improve the biodegradation of PAHs in soil. PMID- 11101323 TI - Kinetic resolution of racemic alpha-methyl-beta-propiothiolactone by lipase catalyzed hydrolysis. AB - Kinetic resolution of racemic alpha-methyl-beta-propiothiolactone (rac-MPTL) using lipases in organic solvent was studied. The lipase from Pseudomonas cepacia (PCL) showed the highest (S)-enantioselectivity (E > 100), and cyclohexane containing 1% (v/v) buffer was identified as the best reaction medium for maintaining high enantioselectivity as well as high reaction rate. While the substrate inhibition was not observed up to 300 mM rac-MPTL, severe product inhibition was observed even at 50 mM racemic 3-mercapto-alpha-methyl propionic acid (rac-MMPA), which made the use of high substrate concentration difficult. To overcome the product inhibition, the products, (R)-MMPA, were neutralized by addition of a dilute basic solution. Although the resolution reaction proceeded further by the base titration, the enantioselectivity of the reaction decreased as a result of nonenantioselective hydrolysis of rac-MPTL in the basic solution. Under these conditions, 200 mM rac-MPTL was successfully resolved to above 95% ee(S) with 53% conversion. PMID- 11101324 TI - Fed-batch bioreactor strategies for microbial decolorization of azo dye using a Pseudomonas luteola strain. AB - A Pseudomonas luteola strain possessing azoreductase activity was utilized to decolorize a reactive azo dye (C. I. Reactive Red 22) with fed-batch processes consisting of an aerobic cell growth stage and an anaerobic fed-batch decolorization stage. The fed-batch decolorization was conducted with different agitation and aeration rates, initial culture volumes, dye loading strategies, and yeast extract to dye (Y/D) ratios, and the effect of those operation parameters on azo dye decolorization was evaluated. Dissolved oxygen strongly inhibited the azo reduction activity; thus aeration should be avoided during decolorization but slight agitation (around 50 rpm) was needed. With the periodical feeding strategy, the specific decolorization rate (v(dye)) and overall decolorization efficiency (eta(dye)) tended to increase with increasing feeding concentrations of dye, whereas substrate inhibition seems to arise when the feeding concentration exceeded 600 mg dye/L. In the continuous feeding mode, higher initial culture volume resulted in better eta(dye) due to higher biomass loading, but lower v(dye) due to lower dye concentration in the bioreactor. With a volumetric flow rate (F) of 25 mL/h, both v(dye) and eta(dye) increased almost linearly with the increase in the loading rate of dye (F(dye)) over the range of 50-200 mg/h, while further increase in F(dye) (400 mg/h) gave rise to a decline in v(dye) and eta(dye). As the F was doubled (50 mL/h), the v(dye) and eta(dye) increased with F(dye) only for F(dye) < 80 mg/h. The best v(dye) (113.7 mg dye g cell(-)(1) h(-)(1)) and eta(dye) (86.3 mg dye L(-)(1) h(-)(1)) were achieved at F(dye) = 200 mg/h and F = 25 mL/h. The yield coefficient representing the relation between dye decolorized and yeast extract consumed was estimated as 0.8 g/g. With F(dye) = 75 mg/h, the Y/D ratio should be higher than 0.5 to ensure sufficient supply of yeast extract for stable fed-batch operations. However, performance of the fed-batch decolorization process was not appreciably improved by raising the Y/D ratio from 0.5 to 1.875 but was more sensitive to the changes in the dye loading rate. PMID- 11101325 TI - Enantioselective synthesis of (S)-ibuprofen ester prodrug in cyclohexane by Candida rugosa lipase immobilized on Accurel MP1000. AB - An enantioselective esterification process was developed for the synthesis of 2-N morpholinoethyl (S)-ibuprofen ester prodrug from racemic ibuprofen by using Candida rugosa lipase immobilized on Accurel MP1000 in cyclohexane. Compared with the performance of Lipase MY, the immobilized lipase possesses a higher enzyme activity and thermal stability, but with a slightly suppressed enantioselectivity. A kinetic model was proposed and confirmed from experiments, for the simulation of time-course conversions of both enantiomers at various combinations of substrate concentrations in a batch reactor. Preliminary results of employing the proposed model and the immobilized lipase in a continuous packed bed reactor were also reported and discussed. PMID- 11101326 TI - Production of cyclodepsipeptides destruxin A and B from Metarhizium anisopliae. AB - Maltose and peptone were the best carbon and nitrogen sources for the production of destruxins from Metarhizium anisopliae. With the addition of 0.1% (w/v) beta alanine to the basal medium, the yields of cyclodepsipeptides DA and DB were 7.2 and 279 mg/L, respectively, which was 2-fold higher than that of control experiment. Response surface methodology (RSM) was applied to optimize the compositions of maltose, peptone, beta-alanine, and glucose used in a shaker flask cultivation of M. anisopliae for the production of DA and DB. Estimated optimal compositions for the DA production were maltose 2.58%, peptone 0.72%, beta-alanine 0.02%, and glucose 0.55%. The predicted DA yield was 18.5 mg/L. On the other hand, the optimal compositions for DB production were maltose 2.51%, peptone 0.75%, beta-alanine 0.02%, and glucose 0.43%. A maximum DB yield of 232 mg/L was predicted. These were confirmed by cultivation experiments conducted at the optimized conditions for maximum destruxins production in a shaker-flask. Furthermore, a modest high level of DA (49 mg/L) and DB (268 mg/L) yields were obtained by employing the response surface methodology optimized DB production medium in a no-baffle, stirred-tank fermentor. PMID- 11101327 TI - Kinetics of heat-shock response and inclusion body formation during temperature induced production of basic fibroblast growth factor in high-cell-density cultures of recombinant Escherichia coli. AB - The kinetics of the heat-shock response and the formation of inclusion bodies in recombinant Escherichia coli TG1 were studied in glucose-limited high-cell density cultures in response to temperature-induced production of human basic fibroblast growth factor (hFGF-2), a protein which partially aggregates into inclusion bodies. The maximum synthesis rates of heat-shock proteins were similar to those in a control cultivation with a strain carrying an expression vector without inducible structural gene. However, the maximum of induction for many heat-shock proteins including DnaK, ClpB, and HtpG was reached at least 30 min later when synthesis of hFGF-2 was simultaneously induced by the temperature upshift. During this first production phase, hFGF-2 was exclusively deposited in the insoluble cell fraction. Thereafter, accumulation of soluble hFGF-2 was observed, too, indicating that the recombinant protein needs heat-shock chaperones for proper folding at elevated temperatures. Strong recombinant protein production prolonged the synthesis of the majority of heat-shock proteins (including GroELS, DnaK, ClpB, and HtpG) even in a wildtype dnaK(+) background. In contrast, the synthesis rates of the small heat-shock proteins IbpA and IbpB declined within 1 h to preinduction values in control and hFGF-2 producing cultures. In the producing cultivation, IbpA and IbpB synthesis ceased to an undetectable level when soluble hFGF-2 started to accumulate, whereas the synthesis rates of the other heat-shock proteins including those belonging to the DnaK and GroEL families remained high throughout the entire production phase. PMID- 11101328 TI - Acetic acid production from lactose by an anaerobic thermophilic coculture immobilized in a fibrous-bed bioreactor. AB - An anaerobic thermophilic coculture consisting of a heterofermentative bacterium (Clostridium thermolacticum) and a homoacetogen (Moorella thermoautotrophica) was developed for acetic acid production from lactose and milk permeate. The fermentation kinetics with free cells in conventional fermentors and immobilized cells in a recycle batch fibrous-bed bioreactor were studied. The optimal conditions for the cocultured fermentation were found to be 58 degrees C and pH 6.4. In the free-cell fermentation, C. thermolacticum converted lactose to acetate, ethanol, lactate, H(2) and CO(2), and the homoacetogen then converted lactate, H(2), and CO(2) to acetate. The overall acetate yield from lactose ranged from 0.46 to 0.65 g/g lactose fermented, depending on the fermentation conditions. In contrast, no ethanol was produced in the immobilized-cell fermentation, and the overall acetate yield from lactose increased to 0.8-0.96 g/g lactose fermented. The fibrous-bed bioreactor also gave a higher final acetate concentration (up to 25. 5 g/L) and reactor productivity (0.18-0.54 g/L/h) as compared to those from the free-cell fermentation (final acetate concentration, 15 g/L; productivity, 0.06-0.08 g/L/h). The superior performance of the fibrous-bed bioreactor was attributed to the high cell density (20 g/L) immobilized in the fibrous-bed and adaptation of C. thermolacticum cells to tolerate a higher acetate concentration. The effects of yeast extract and trypticase as nutrient supplements on the fermentation were also studied. For the free-cell fermentation, nutrient supplementation was necessary for the bacteria to grow in milk permeate. For the immobilized-cell fermentation, plain milk permeate gave a high acetate yield (0.96 g/g), although the reactor productivity was lower than those with nutrient supplementation. Balanced growth and fermentation activities between the two bacteria in the coculture are important to the quantitative conversion of lactose to acetic acid. Lactate and hydrogen produced by C. thermolacticum must be timely converted to acetic acid by the homoacetogen to avoid inhibition by these metabolites. PMID- 11101329 TI - Estimation of biological kinetic parameters from a continuous integrated ozonation-activated sludge system treating domestic wastewater. AB - The feasibility of treating municipal wastewater by a combined ozone-activated sludge continuous flow system was studied. Lab-scale experiments of both single activated sludge and combined ozone-activated sludge processes were carried out to determine the kinetic coefficients of the biological stage. The results obtained indicated a clear improvement in the kinetic parameters of the aerobic oxidation when a pre-ozonation stage was applied. Particularly, COD removal and nitrification rates were highly increased. The biokinetic parameters were also used to simulate and optimize the continuous reaction system. From the model prediction it was concluded that the integrated process (i.e., ozone-ASP) may significantly increase the waste reduction capacity. The results presented here provide a useful basis for further scaling up and efficient operation of ozone ASP units in wastewater treatment processes. PMID- 11101330 TI - Enzymatic treatment of mechanical pulp fibers for improving papermaking properties. AB - Three enzyme preparations (crude cellulase, laccase, and proteinase) were evaluated for their potential to improve the papermaking properties of mechanical pulp. After treating a long fibre-rich fraction of the pulp with enzyme, the fibres were recombined with untreated fines for handsheet making and testing. None of the enzymes altered the retention of fines or the consolidation of the furnish mix during handsheet formation. All three enzymes increased tensile stiffness index, which is a measure of the initial resistance of the handsheets to strain. Only the laccase preparation, an enzyme that modifies pulp lignin, consistently increased fibre bonding to enhance other strength properties of the handsheets. PMID- 11101331 TI - High-pressure shift freezing. Part 1. Amount of ice instantaneously formed in the process. AB - A mathematical model to calculate the amount of ice formed instantaneously after a rapid expansion in high-pressure shift processes (HPSF) was developed. It considers that when water is expanded it does not extend over its melting curve but reaches a metastable state (supercooled water), which also occurs in practice. Theoretical results appear to agree with experimental data. PMID- 11101332 TI - High-presssure shift freezing. Part 2. Modeling of freezing times for a finite cylindrical model. AB - A comprehensive vision of the heat transfer process involved in high-pressure shift freezing (HPSF) is shown in comparison to the process at atmospheric pressure. In addition, a mathematical model to predict the freezing times is presented. This model takes into consideration the dependence of the thermophysical properties relating to temperature and pressure and the supercooling reached by liquid water at atmospheric pressure after adiabatic expansion in the HPSF process. Experimental and theoretical data appear to agree. PMID- 11101333 TI - Pressure and flux profiles in bead-filled ultrafiltration/microfiltration hollow fiber membrane modules. AB - A general mathematical model for the prediction of pressure, flow rate, and flux profiles in an ultrafiltration/microfiltration hollow fiber membrane module whose shell side is filled with beads has been developed. The model was studied for a variety of operational modes in such modules, e.g., ultrafiltration/microfiltration, permeate flow rate control, Starling flow (encountered in hollow fiber bioreactors), and tube-side elution (encountered in filtration-cum-chromatography processes), etc., with or without a bead-filled extended section at the permeate outlet. An algorithm is provided to determine the model parameters from experimental data using the model equations. The solutions developed have been used to study the uniformity of transmembrane pressure profile along the module length using a quantity called the uniformity factor alpha. This factor shows that the model can be a useful tool for achieving the desired module performance in a number of quite different applications. The model predicts successfully the nature of the transmembrane pressure profile and the solvent flux profile in situations that are quite different, namely, conventional ultrafiltration and Starling flow. The approach used in this study can also be adopted to develop a model for description of other operational modes such as backflushing and shell-side elution used in the processes of filtration cum-chromatography. Those applications employing similar device configurations may also use this model to predict the pressure and flux profiles to facilitate the design of the process and the operation conditions. PMID- 11101334 TI - Optimized single-step affinity purification with a self-cleaving intein applied to human acidic fibroblast growth factor. AB - To reduce the number of recovery steps during downstream processing and to overcome the limitations of present fusion-based affinity separations, a controllable self-splicing protein element in the form of a mini-intein was used to optimize the recovery of proteins for both batch and flow purification strategies. The ability to recover purified proteins was demonstrated using a tripartite fusion consisting of a maltose binding domain, a truncated intein as a controllable linker molecule, and a protein of interest. To characterize expression level, solubility, cleavage rates, pH and temperature controllability, and protein activity, recombinant human acidic fibroblast growth factor (aFGF) was used as a model protein. A simple mass transport model, based on cleavage reaction-limited mass transfer and constant dispersion, was successfully used to predict product concentration and peak shape in relation to critical process parameters (with no fitting parameters). Insight into the nature of the cleavage reaction and its regulation was obtained via temperature- and pH-dependent kinetic data. PMID- 11101335 TI - Purification of oligonucleotides by high affinity, low molecular weight displacers. AB - High affinity, low molecular weight anionic displacers were successfully employed for the purification of antisense oligonucleotides. Several important structural characteristics were identified that contribute to the affinity of low molecular weight displacers to a hydrophilized polystyrene divinyl benzene anion exchanger. Sulfonic acid groups were found to possess higher affinity than carboxylic acid and phosphate functionalities, and nonspecific interactions (particularly hydrophobic interactions) were shown to play a major role in the retention process on this stationary phase material. Using this information, two high affinity, low molecular weight displacers were identified. These molecules are relatively inexpensive organic dyes that possess multiple sulfonic acid moieties, as well as aromatic functionalities, which increase nonspecific interactions with the stationary phase. These high affinity displacers, which can be readily detected, were then employed to displace several strongly retained antisense oligonucleotides that could not be displaced by previously established low molecular weight displacers. The displacement process resulted in very high purities of the antisense oligonucleotides. The results presented in this paper are significant in that they demonstrate that low molecular weight displacers for ion-exchange chromatography can possess equal to or greater affinities than their higher molecular weight counterparts, when nonspecific interactions with the stationary phase are exploited. In addition, the results illustrate the high resolutions possible with displacement chromatography and demonstrate an attractive technology for the process scale purification of oligonucleotides. PMID- 11101336 TI - The hydrodynamics of a Graesser ("raining bucket") contactor with a reverse micellar phase. AB - A variety of contactor types have been assessed for the liquid-liquid extraction of proteins using reversed micelles; however, many of these contactors suffer from drawbacks such as emulsion formation and poor mass transfer performance. In this study, a small (1.25 L) Graesser "raining bucket" contactor was assessed for use with this system since it has the potential to ameliorate many of these problems. The aim of the work was to evaluate the hydrodynamics of the contactor in order to use this information for future work on mass transfer performance. Hydrodynamic characteristics such as the axial mixing coefficient were determined by residence time distribution studies using a tracer injection method. The effect of rotor speed and flow rate of each phase on axial mixing was investigated, and as a result of its unusual structure, i.e., falling/rising sheet, the interfacial mass transfer area in the Graesser was determined by image analysis. It was found that rotor speed had more influence on the axial mixing coefficient in the aqueous phase than in the reverse micellar phase. The axial mixing coefficient in each phase increased by increasing the flow rate of the same phase. The images obtained in a dropping cell showed that under the conditions of this study (3 rpm, 22 degrees C), the bucket pours one phase through the other in the form of a curtain or sheet. A new image technique was developed to determine the interfacial area of both phases, and it was found that the specific area was 8.6 m(2)/m(3), which was higher than in a spray column but considerably lower than in a RDC or a Graesser run at high rotational speed (50 rpm) without the addition of a surfactant. PMID- 11101337 TI - Important parameters affecting efficiency of protein refolding by reversed micelles. AB - Refolding of denatured RNase A as a model of inclusion bodies was performed by reversed micelles formulated with sodium di-2-ethylhexyl sulfosuccinate (AOT) in isooctane. In the novel refolding process, a solid-liquid extraction was utilized as an alternative to the ordinary protein extraction by reversed micelles based on a liquid-liquid extraction. First, the effects of operational parameters such as concentration of AOT, W(o) (= [H(2)O]/[AOT]), and pH were examined on the solubilization of solid denatured proteins into a reversed micellar solution. The solubilization was facilitated by a high AOT concentration, a high W(o) value, and a high pH in water pools. These conditions are favorable for the dispersion of the solid protein aggregates in an organic solvent. Second, the renaturation of the denatured RNase A solubilized into the reversed micellar solution was conducted by addition of glutathione as a redox reagent. A complete renaturation of RNase A was accomplished by adjusting the composition of the redox reagent even at a high protein concentration in which protein aggregation would usually occur in aqueous media. In addition, the renaturation rates were improved by optimizing water content (W(o)) and the pH of water pools in reversed micelles. Finally, the recovery of renatured RNase A from the reversed micellar solution was performed by adding a polar organic solvent such as acetone into the reversed micellar solution. This precipitation method was effective for recovering proteins from reversed micellar media without any significant reduction in enzymatic activity. PMID- 11101338 TI - Shear stress affects the kinetics of Staphylococcus aureus adhesion to collagen. AB - Staphylococcus aureus is a major human pathogen that has been shown to bind collagen under static conditions. However, many staphylococcal infections are hematogenously acquired and adhesion events may be influenced by shear stress. In this study, we used a dynamic experimental system consisting of a parallel-plate perfusion chamber and phase-contrast video microscope to study the effects of shear stress on the adhesion kinetics of intact S. aureus to collagen surfaces in vitro. The adhesion of S. aureus Phillips to collagen types I, II, and IV was investigated over a physiologically relevant range of wall shear stresses at 37 degrees C. S. aureus PH100, a collagen adhesin-deficient mutant strain, was used as a control strain for the experiments. We found that S. aureus Phillips could adhere to collagens I, II, and IV at wall shear stresses less than 15 dyn/cm(2) and that the kinetics of the adhesion process were wall shear stress-dependent. Similar studies with PH100 demonstrated that these cells are unable to adhere firmly to collagen surfaces. Transient interactions between PH100 and the collagen surfaces were observed at low levels of shear stress suggesting that S. aureus may also interact with collagen by an alternative mechanism that does not lead to firm adhesion. PMID- 11101339 TI - Chitosan-based coagulating agents for treatment of cheddar cheese whey. AB - Chitosan-Polyanion (Chi-Pol) complexes were used as coagulating agents for treating Cheddar cheese whey. Complexation and coagulation time played a significant role in adsorption, whereas polymer concentration was significant only for chitosan-alginate complexes. Complexes of chitosan with alginate (ALG), pectin (PEC), and carrageenan (CAR) used at 30 mg complex/L whey showed turbidity reductions of 40-43% and 65-72% after 1 and 39 h, respectively. At 10 mg/L, the percent reduction in turbidity after 1 and 39 h were 35-39% and 61-64%, respectively. No significant differences in turbidity reduction (P > 0.05) were observed when using complexes at different Chi-Pol monomeric mixing ratios (MR) except for Chi-Alg at 30 mg/L, wherein reduction at 0.2 was higher than 0.8 MR. Also, UV-vis spectroscopy suggested the preference of this complex for the absorption of specific whey protein fractions. This study successfully demonstrated the effectiveness of Chi-Pol complexes in flocculation of suspended solid wastes in cheese whey with over 70% protein recovery. PMID- 11101340 TI - Critical evaluation of models developed for monitoring an industrial submerged bioprocess for antibiotic production using near-infrared spectroscopy. AB - Near-infrared spectroscopy (NIRS) is known to have potential for cost-effective monitoring of bioprocesses. Although this has been demonstrated in many instances and several models have been reported, information regarding the complexity of models required and their utility over extended periods of time is lacking. In the present study, the complexity of the models required for the NIRS prediction of substrate (oil) and product (tylosin) concentration in an industrial bioprocess that employs a physicochemically heterogeneous medium for antibiotic production was assessed. Measurements made by both the diffuse reflectance and transmittance modes were investigated. SEP values for the prediction of the analytes averaged 5% or less, for the successful models, when evaluated using an external validation set, 2 years after the initial model development exercise. Diffuse reflectance measurements showed poorer results, compared to transmittance measurements, especially for monitoring tylosin. In general, this investigation provides evidence to support the fact that models built for the prediction of analytes in a commercial bioprocess that employs a physicochemically complex production medium can be robust in performance over an extended period of time and that simple models based on fewer terms or latent variables can perform well, even in the context of matrices that are relatively complex. It also indicates that sample presentation is likely to be a critical factor in the successful application of NIRS in bioprocess monitoring, which merits further detailed investigation. PMID- 11101341 TI - Use of green fluorescent protein-conjugated beta-actin as a novel molecular marker for in vitro tumor cell chemotaxis assay. AB - To study the dynamics of actin cytoskeleton rearrangement in living cells, an eukaryotic expression vector expressing a beta-actin-GFP fusion protein was generated. The expression construct when transfected into NIH3T3 fibroblast, A2058 human melanoma and 293T human embryonic kidney carcinoma cell lines expressed beta-actin-GFP fusion protein, which colocalized with endogenous cellular actin as determined by histoimmunofluorescence staining. The beta-actin GFP was also observed to be reorganized in response to treatments with the chemoattractant type IV collagen. Cells extended pseudopodial protrusions and altered the morphology of their cortical structure in response to type IV collagen stimulation. More importantly, beta-actin-GFP accumulated in areas undergoing these dynamic cytoskeleton changes, indicating that beta-actin-GFP could participate in actin polymerization. Although ectopic expression of beta actin-GFP lead to minor side effects on cell proliferation, these studies suggest that this strategy provides an alternative to the invasive techniques currently used to study actin dynamics and permits real-time visualization of actin rearrangements in response to environmental cues. PMID- 11101342 TI - Bone affinity of a bisphosphonate-conjugated protein in vivo. AB - Growth factors capable of stimulating bone formation are potential therapeutic agents for osteoporosis treatment. It is essential, however, that a targeting mechanism is incorporated into the growth factors to deposit them at osseous tissue with minimal distribution to extraskeletal sites. To this end, a strategy has been developed in which a bone-seeking molecule, 1-amino-1,1-diphosphonate methane (aminoBP), was chemically conjugated to a model protein, bovine serum albumin (BSA). This study was carried out to assess the bone affinity of the conjugates in a tibia injection model. Using ovariectomized (OVX) rats, initial (3 h) retention of BSA and aminoBP-BSA were found to be equivalent when injected into the medullary cavity of tibia. After 1 day, an 8- and 12-fold higher tibiae retention of the protein was obtained in normal and OVX rats as a result of aminoBP conjugation. A similar result ( approximately 12-fold difference) was also obtained in OVX rats after 3 days. We concluded that aminoBP conjugation to BSA imparted a high bone affinity and enhanced bone retention of proteins in normal and OVX rats. PMID- 11101343 TI - Compensation of temperature and concanavalin A concentratration effects for glucose determination by the viscometric affinity assay. AB - A viscometer suitable for rapid measurements in small volumes of highly viscous liquids is described. Using this device the viscometric affinity assay for glucose was studied under variable conditions in order to obtain basic information for the design of a viscometric glucose sensor. The viscosity of the dextran/Concanavalin A (ConA) solution is sensitive to glucose in a broad range of the shear stress. However, for measuring the glucose concentration with this sensitive liquid the strong dependence of the absolute viscosity on temperature and ConA concentration has to be taken into account. For the purpose of calibration a parameter more suitable than the absolute viscosity is the relative fluidity (F(r)) that is defined by the actual measured viscosity at a given glucose concentration, the reference viscosity at a standard glucose concentration, and a constant linearization coefficient. F(r) shows a linear dependence on the glucose concentration in the therapeutically interesting range up to 30 mM and is not significantly changed by moderate variations of the ConA concentration or temperature. PMID- 11101344 TI - Glutaraldehyde treatment of proteinaceous gas vesicles from cyanobacterium Anabaena flos-aquae. AB - As potential gas microcarriers, gas vesicles (GVs) were isolated from cultures of the filamentous cyanobacterium Anabaena flos-aquae and treated with glutaraldehyde. The effects of glutaraldehyde treatment on the stability of GVs, against elevated temperatures (40-121 degrees C) and protein-stripping agents such as urea and sodium dodecyl sulfate (SDS), were then examined with the pressure collapse curves generated using pressure nephelometry. The treatment was very beneficial to GVs against the exposure to SDS and urea; however, it did not make the evolution-optimized vesicle structure stronger or more temperature resistant. In the presence of these protein-stripping agents, the treated vesicles had higher median (50%) collapse pressures (by > or =1 atm) than the untreated ones, at both room temperature and 40 degrees C. This increase has been presumably attributed to the cross-linking of the large GvpC protein to the ribbed GvpA shell, thereby resisting the stripping of GvpC that provides the primary mechanical strength to the vesicle wall. The glutaraldehyde treatment also restored the strength of GVs weakened by a 5-week storage in a refrigerator and, therefore, is expected to improve the stability of GVs for long-term storage. GVs could not be autoclaved. If necessary for the intended applications, glutaraldehyde treatment may also serve to chemically sterilize the vesicles, with the glutaraldehyde subsequently removed by dialysis. PMID- 11101345 TI - Enzymatic catalysis of formation of Z-aspartame in ionic liquid - An alternative to enzymatic catalysis in organic solvents. AB - We present the first report of enzymatic catalysis in an ionic liquid. The virtually nonexistent vapor pressure makes ionic liquids an exciting new alternative for enzyme-catalyzed syntheses in environmentally friendly environments. Z-aspartame was synthesized in a thermolysin-catalyzed reaction of carbobenzoxy-L-aspartate and L-phenylalanine methyl ester hydrochloride in 1 butyl-3-methylimidazolium hexafluorophosphate (BP6). Ionic liquids such as BP6 are thermally stable and have a remarkable range of temperatures over which they remain liquid (300 degrees C). With an initial rate of 1.2 +/- 0.1 nmol min(-)(1) mg(-)(1), we observed a competitive rate in comparison to that of enzymatic synthesis in organic solvent. Additionally, the enzyme exhibits outstanding stability, which would normally require immobilization. PMID- 11101346 TI - Bifunctional adsorbents for hydrophobic displacement chromatography of proteins. AB - The separation of pancreatic trypsinogen and alpha-chymochypsinogen A by displacement chromatography was tested on a bifunctional adsorbent containing both butyl and carboxymethyl groups. Methacrylic triblock copolymer was synthesized and used as the displacer. Compared to the displacement results on commercial Butyl-Sepharose, it was found that both the separation and recovery of trypsinogen and alpha-chymochypsinogen A were improved. Adsorption isotherms of proteins were measured on both the commercial Butyl-Sepharose and the synthesized bifunctional adsorbents. It was found that the improvement of protein separation on bifunctional adsorbents was attributed to the alteration of the adsorption of trypsinogen. Charge repulsion between trypsinogen and the negatively charged carboxymethyl groups may account for the alteration. In addition, taking advantage of the effect of charge repulsion, the column regeneration became much easier on the column packed with bifunctional adsorbents. PMID- 11101347 TI - Medium optimization of transformed root cultures of Stizolobium hassjoo producing L-DOPA with response surface methodology. AB - Medium optimization of B5 medium for hairy root cultures producing secondary metabolites was studied through statistical experimental design. Transformed root cultures of Stizolobium hassjoo producing L-DOPA were used as a model system. The serial dilution experiments facilitated logical choice of the upper and lower bounds on executing 2(11)(-)(6) fractional factorial design. Steepest ascent method as well as central composite design were sequentially employed to optimize the media of shake flask cultures. The modified B5 media of GM, PM, and GPM were obtained, indicating the optimum medium compositions for enhancing hairy root dry weight, L-DOPA content in hairy roots, and L-DOPA production, respectively. When cultivating S. hassjoo hairy roots in GM, PM and GPM for 16 days, the dry wt of hairy roots, L-DOPA content, and L-DOPA production obtained were ca. 530 mg per flask (10.6 g/L), 10.8% dry wt, and 806 mg/L, which were 1.8-, 2-, and 2.8-fold of basal B5 medium control runs, respectively. PMID- 11101348 TI - Discovery of chiral N,N-disubstituted trifluoro-3-amino-2-propanols as potent inhibitors of cholesteryl ester transfer protein. PMID- 11101349 TI - Analogues of bacteriocins: antimicrobial specificity and interactions of leucocin A with its enantiomer, carnobacteriocin B2, and truncated derivatives. PMID- 11101350 TI - 1,3-Diaryl-4,5,6,7-tetrahydro-2H-isoindole derivatives: a new series of potent and selective COX-2 inhibitors in which a sulfonyl group is not a structural requisite. AB - Novel tetrahydro-2H-isoindoles have been prepared and evaluated as inhibitors of the COX-2 isoenzyme. A 1,3-diaryl substitution on the central polycyclic ring system and absence of a sulfonyl moiety are the two structural features of this chemical series. A short and easy synthetic pathway produced several derivatives which were shown to be potent and selective COX-2 vs COX-1 inhibitors (IC(50) = 0. 6-100 nM for COX-2, 100->1000 nM for COX-1). Structural modifications established that a bicyclic ring appended to the pyrrole nucleus and 4,4' difluoro substitution on the phenyl rings were optimal for high inhibitory potency. Activity was confirmed in the human whole blood assay and subsequently in the murine air-pouch model in which in vivo PGE2 inhibitory activity was evaluated with respect to gastric tolerance (ED(50) for inhibition of exudate PGE2 of 3 mg/kg and gastric PGE2 of 20 mg/kg). Gastric tolerance was further assessed after administration to mice of high doses (up to 400 mg/kg) of the inhibitors by measurement of gastric damage. This panel of studies allowed selection of a number of tetrahydro-2H-isoindoles which were compared in the adjuvant-induced arthritis model. Compounds 32 and 37 showed the most potent activity with ED(50) values for edema inhibition in the noninjected paw of 0. 35 and 0.15 mg/kg/day, respectively, after oral administration. In addition, this interesting antiinflammatory profile was accompanied by a protective effect against arthritis-induced osteopenia, the decrease being 50% with a dose of 0.25 mg/kg/day. PMID- 11101351 TI - Simulations of the estrogen receptor ligand-binding domain: affinity of natural ligands and xenoestrogens. AB - We have carried out molecular dynamics (MD) simulations and free energy calculations on the alpha-subtype of the human estrogen receptor ligand-binding domain (ERalpha LBD) complexed with a number of known agonists and putative xenoestrogens. Our dynamical simulations of ligand-receptor complexes underscore the highly structured nature of the complex and offer some interesting insights into the structure-activity relationship (SAR) for these ligands. With traditional thermodynamic integration (TI) calculations, we calculate relative binding free energies for three known agonists, in good agreement with experimental values. The sheer number of possible xenoestrogenic compounds makes an approach using traditional free energy calculations unfeasible. Instead, we have made use of a single-step perturbation methodology that allows the calculation of relative free energies for a large number of related polyaromatic hydrocarbons (PAHs) from a single simulation. Our results show good (maximum deviation 3.3 kJ mol(-1)) agreement with experimental data, suggesting the possibility of large-scale xenoestrogen screening in silico to obtain strongly estrogenic compounds for subsequent experimental testing. PMID- 11101352 TI - Pyrido[2,3-d]pyrimidin-7-one inhibitors of cyclin-dependent kinases. AB - The identification of 8-ethyl-2-phenylamino-8H-pyrido[2, 3-d]pyrimidin-7-one (1) as an inhibitor of Cdk4 led to the initiation of a program to evaluate related pyrido[2, 3-d]pyrimidin-7-ones for inhibition of cyclin-dependent kinases (Cdks). Analysis of more than 60 analogues has identified some clear SAR trends that may be exploited in the design of more potent Cdk inhibitors. The most potent Cdk4 inhibitors reported in this study inhibit Cdk4 with IC(50) = 0.004 microM ([ATP] = 25 microM). X-ray crystallographic analysis of representative compounds bound to the related kinase, Cdk2, reveals that they occupy the ATP binding site. Modest selectivity between Cdks is exhibited by some compounds, and Cdk4 selective inhibitors block pRb(+) cells in the G(1)-phase of the cell division cycle. PMID- 11101353 TI - Cation substitution in cationic phosphonolipids: a new concept to improve transfection activity and decrease cellular toxicity. AB - Cationic lipids have been shown to be an interesting alternative to viral vector mediated gene delivery into in vitro and in vivo model applications. Prior studies have demonstrated that even minor structural modifications of the lipid hydrophobic domain or of the lipid polar domain result in significant changes in gene delivery efficiency. Previously, we developed a novel class of cationic lipids called cationic phosphonolipids and described the ability of these vectors to transfer DNA into different cell lines and in vivo. Up until now, in all new cationic lipids, nitrogen atoms have always carried the cationic or polycationic charge. Recently we have developed a new series of cationic phosphonolipids characterized by a cationic charge carried by a phosphorus or arsenic atom. In a second step, we have also examined the effects of the linker length between the cation and the hydrophobic domain as regards transfection activity. Transfection activities of this library of new cationic phosphonolipids were studied in vitro in different cell lines (HeLa, CFT1, K562) and in vivo using a luciferase reporter gene. A luminescent assay was carried out to assess luciferase expression. We demonstrated that cation substitution on the polar domain of cationic phosphonolipids (N --> P or As) results in significant increase in transfection activity for both in vitro and in vivo assays and decrease of cellular toxicity. PMID- 11101354 TI - Chiral derivatives of 2-cyclohexylideneperhydro-4,7-methanoindenes, a novel class of nonsteroidal androgen receptor ligand: synthesis, X-ray analysis, and biological activity. AB - A series of 2-cyclohexylideneperhydro-4,7-methanoindene derivatives was synthesized as novel androgen receptor ligands. Asymmetric hydroboration of key intermediate 2 afforded single enantiomer alcohol derivatives (3aR)-3 and (3aS)-3 which could be further transformed to give 12 variously substituted keto alcohol target compounds. X-ray crystallography of the 4-bromobenzenesulfonyl ester (3aS) 13 was used to establish their absolute configuration. The binding of these compounds to the rat ventral prostate androgen receptor showed moderate affinity with IC(50) values of 1.2 microM and above but with substantial enantiomeric dependencies which varied in accordance to Pfieffer's rule. Surprisingly, the (3aS)-5alpha-alcohols displayed similar affinity to the (3aR)-5beta-alcohols, and molecular modeling suggested an alternative mode of binding for the (3aS) series. The three compounds with the best androgen receptor affinity were assayed in vivo for antiandrogenic and androgenic effects on sex accessory organ growth in castrated immature rats and were found to be ineffective. PMID- 11101355 TI - 7-Deazaadenines bearing polar substituents: structure-activity relationships of new A(1) and A(3) adenosine receptor antagonists. AB - A series of 28 new pyrrolo[2,3-d]pyrimidine-4-amines, pyrimido[4, 5-b]indole-4 amines, and tetrahydropyrimido[4,5-b]indole-4-amines was synthesized and their adenosine receptor affinity determined in radioligand binding assays at rat A(1) and A(2A) adenosine receptors (ARs). Selected compounds were additionally investigated in binding assays at recombinant A(3) ARs. The 2-phenyl residue in (R)-7-(1-methylbenzyl)-2-phenylpyrrolo[2,3-d]pyrimidine-4-amine (ADPEP, 1) and in the corresponding pyrimido[4,5-b]indole (APEPI, 3) could be bioisosterically replaced by heterocyclic rings, such as 2-thienyl and 4-pyridyl. The resulting compounds retained high affinity and selectivity for A(1) ARs. Judging from the investigation of selected compounds, it appears that they are also potent at human A(1) ARs and selective not only versus A(2A) ARs but also highly selective versus A(2B) and A(3) ARs. The p-pyridyl-substituted derivatives 11 and 27 (APPPI) may be interesting pharmacological tools due to their fluorescent properties. Pyrrolo[2,3-d]pyrimidine-4-amine derivatives which were simultaneously substituted at N7 and N(4), combining the substitution pattern of ADPEP (1) and DPEAP (2), showed very low affinity for A(1) ARs. This finding supports our previously published hypothesis of different binding modes for pyrrolopyrimidines, such as ADPEP (1) and DPEAP (2). DPEAP (2), a pyrrolo[2,3 d]pyrimidine-4-amine substituted at the amino group (N(4)), was found to exhibit high affinity for human A(3) ARs (K(i) = 28 nM), whereas N(4)-unsubstituted analogues were inactive. DPEAP (2) and related compounds provide new leads for the development of antagonists for the human A(3) AR. PMID- 11101356 TI - 5-Substituted N(4)-hydroxy-2'-deoxycytidines and their 5'-monophosphates: synthesis, conformation, interaction with tumor thymidylate synthase, and in vitro antitumor activity. AB - Convenient procedures are described for the synthesis of 5-substituted N(4) hydroxy-2'-deoxycytidines 5a,b,d-h via transformation of the respective 5 substituted 3', 5'-di-O-acetyl-2'-deoxyuridines 1a-c,e-h. These procedures involved site-specific triazolation or N-methylimidazolation at position C(4), followed by hydroxylamination and deblocking with MeOH-NH(3). Nucleosides 5a,b,d h were selectively converted to the corresponding 5'-monophosphates 6a,b,d-h with the aid of the wheat shoot phosphotransferase system. Conformation of each nucleoside in D(2)O solution, deduced from (1)H NMR spectra and confirmed by molecular mechanics calculations, showed the pentose ring to exist predominantly in the conformation S (C-2'-endo) and the N(4)-OH group as the cis rotamer. Cell growth inhibition was studied with two L5178Y murine leukemia cell lines, parental and 5-fluoro-2'-deoxyuridine (FdUrd)-resistant, the latter 70-fold less sensitive toward FdUrd than the former. With FdUrd-resistant L5178Y cells, 5 fluoro-N(4)-hydroxy-2'-deoxycytidine (5e) caused almost 3-fold stronger growth inhibition than FdUrd; 5e was only some 3-fold weaker growth inhibitor of the resistant cells than of the parental cells. Thymidylate synthase inhibition was studied with two forms of the enzyme differing in sensitivities toward 5-fluoro 2'-deoxyuridine 5'-monophosphate (FdUMP), isolated from parental and FdUrd resistant L1210 cell lines. All N(4)-hydroxy-dCMP (6a,b,d-h) and dUMP analogues studied were competitive vs dUMP inhibitors of the enzyme. Analogues 6b,d-h and 5 hydroxymethyl-dUMP, similar to N(4)-hydroxy-dCMP (6a) and FdUMP, were also N(5), N(10)-methylenetetrahydrofolate-dependent, hence mechanism-based, slow-binding inhibitors. 5-Chloro-dUMP, 5-bromo-dUMP, and 5-iodo-dUMP, similar to dTMP, did not cause a time-dependent inactivation of the enzyme. Instead, they behaved as classic inhibitors of tritium release from [5-(3)H]dUMP. 5-Bromo-dUMP and 5-iodo dUMP showed substrate activity independent of N(5), N(10) methylenetetrahydrofolate in the thymidylate synthase-catalyzed dehalogenation reaction. The =N-OH substituent of the pyrimidine C(4) prevented the enzyme catalyzed release from the C(5) of Br(-) and I(-) (the same shown previously for H(+)). While FdUMP and 6a showed a higher affinity and greater inactivation power with the parental cell than FdUrd-resistant cell enzyme, an opposite relationship could be seen with 5-hydroxymethyl-dUMP. PMID- 11101357 TI - New tacrine-huperzine A hybrids (huprines): highly potent tight-binding acetylcholinesterase inhibitors of interest for the treatment of Alzheimer's disease. AB - Several new 12-amino-6,7,10,11-tetrahydro-7, 11-methanocycloocta[b]quinoline derivatives (tacrine-huperzine A hybrids, huprines) have been synthesized and tested as acetylcholinesterase (AChE) inhibitors. All of the new compounds contain either a methyl or ethyl group at position 9 and one or two (chloro, fluoro, or methyl) substituents at positions 1, 2, or 3. Among the monosubstituted derivatives, the more active are those substituted at position 3, their activity following the order 3-chloro > 3-fluoro > 3-methyl > 3-hydrogen. For the 1,3-difluoro and 1,3-dimethyl derivatives, the effect of the substituents is roughly additive. No significant differences were observed for the inhibitory activity of 9-methyl vs 9-ethyl derivatives mono- or disubstituted at positions 1 and/or 3. The levorotatory enantiomers of these hybrid compounds are much more active (eutomers) than the dextrorotatory forms (distomers) as AChE inhibitors. Compounds rac-20, (-)-20, rac-26, (-)-26, rac-30, (-)-30, and rac-31 showed human AChE inhibitory activities up to 28.5-fold higher than for the corresponding bovine enzyme. Also, rac-19, (-)-20, (-)-30, and rac-31 were very selective for human AChE vs butyrylcholinesterase (BChE), the AChE inhibitory activities being 438-871-fold higher than for BChE. Several hybrid compounds, specially (-)-20 and (-)-30, exhibited tight-binding character, showing higher activity after incubation of the enzyme with the inhibitor than without incubation, though the reversible nature of the enzyme-inhibitor interaction was demonstrated by dialysis. The results of the ex vivo experiments also supported the tight-binding character of compounds (-)-20 and (-)-30 and showed their ability to cross the blood-brain barrier. Molecular modeling simulations of the AChE-inhibitor complex provided a basis to explain the differences in inhibitory activity of these compounds. PMID- 11101358 TI - 4-Aminoquinolines: novel nociceptin antagonists with analgesic activity. AB - Small-molecule nociceptin antagonists were synthesized to examine their therapeutic potential. After a 4-aminoquinoline derivative was found to bind with the human ORL(1) receptor, a series of 4-aminoquinolines and related compounds were synthesized and their binding was evaluated. Elucidation of structure activity relationships eventually led to the optimum compounds. One of these compounds, N-(4-amino-2-methylquinolin-6-yl)-2-(4-ethylphenoxymethyl)benzamide hydrochloride (11) not only antagonized nociceptin-induced allodynia in mice but also showed analgesic effect in a hot plate test using mice and in a formalin test using rats. Its analgesic effect was not antagonized by the opioid antagonist naloxone. These results indicate that this nociceptin antagonist has the potential to become a novel type of analgesic that differs from mu-opioid agonists. PMID- 11101360 TI - Adenosine deaminase inhibitors: synthesis and biological evaluation of unsaturated, aromatic, and oxo derivatives of (+)-erythro-9-(2'S-hydroxy-3'R nonyl)adenine [(+)-EHNA]. AB - The synthesis and biological evaluation of three classes of chain-modified derivatives of (+)-EHNA are described. Among the 5', 6'-unsaturated derivatives, the Z-isomer was the most potent inhibitor of adenosine deaminase (ADA) but 3 fold less active than (+)-EHNA. Several 9-aralkyladenines (ARADs) have been prepared, and their inhibitory activity was determined. A minimum of two carbon atoms separating the aromatic ring from the adenine-bearing carbon (C-3') was found to be essential for ADA activity equal to or slightly greater than that of (+)-EHNA. Finally, replacement of the C-5' carbon with an oxygen resulted in reduced potency. PMID- 11101359 TI - Conformationally constrained butyrophenones with affinity for dopamine (D(1), D(2), D(4)) and serotonin (5-HT(2A), 5-HT(2B), 5-HT(2C)) receptors: synthesis of aminomethylbenzo[b]furanones and their evaluation as antipsychotics. AB - A series of novel conformationally restricted butyrophenones (6-aminomethyl 4,5,6,7-tetrahydrobenzo[b]furan-4-ones bearing 4-(6 fluorobenzisoxazolyl)piperidine, 4-(p-fluorobenzoyl)piperidine, 4-(o methoxyphenyl)piperazine, 4-(2-pyridyl)piperazine, 4-(2-pyrimidinyl)piperazine, or linear butyro(or valero)phenone fragments) were prepared and evaluated as antipsychotic agents by in vitro assays for affinity for dopamine receptors (D(1), D(2), D(4)) and serotonin receptors (5-HT(2A), 5-HT(2B), 5-HT(2C)), by neurochemical studies, and by in vivo assays for antipsychotic potential and the risk of inducing extrapyramidal side effects. Potency and selectivity depended mainly on the amine fragment connected to the cyclohexanone structure. Compounds 20b, with a benzoylpiperidine moiety, and 20c, with a benzisoxazolyl fragment, were selective for 5-HT(2A) receptors. The in vitro and in vivo pharmacological profiles of N-[(4-oxo-4,5,6, 7-tetrahydrobenzo[b]furan-6-yl)methyl]-4-(p fluorobenzoyl)piperidine (20b, QF1003B) and N-[(4-oxo-4,5,6, 7 tetrahydrobenzo[b]furan-6-yl)methyl]-4-(6-fluorobenzisoxazol-3-yl)p iperidine (20c, QF1004B) suggest that they may be effective as antipsychotic (neuroleptic) drugs. PMID- 11101361 TI - Effect of ring fluorination on the pharmacology of hallucinogenic tryptamines. AB - A series of fluorinated analogues of the hallucinogenic tryptamines N,N diethyltryptamine (DET), 4-hydroxy-N,N-dimethyltryptamine (4-OH-DMT, psilocin), and 5-methoxy-DMT was synthesized to investigate possible explanations for the inactivity of 6-fluoro-DET as a hallucinogen and to determine the effects of fluorination on the molecular recognition and activation of these compounds at serotonin receptor subtypes. The target compounds were evaluated using in vivo behavioral assays for hallucinogen-like and 5-HT(1A) agonist activity and in vitro radioligand competition assays for their affinity at 5-HT(2A), 5-HT(2C), and 5-HT(1A) receptor sites. Functional activity at the 5-HT(2A) receptor was determined for all compounds. In addition, for some compounds functional activity was determined at the 5-HT(1A) receptor. Hallucinogen-like activity, evaluated in the two-lever drug discrimination paradigm using LSD-trained rats, was attenuated or abolished for all of the fluorinated analogues. One of the tryptamines, 4 fluoro-5-methoxy-DMT (6), displayed high 5-HT(1A) agonist activity, with potency greater than that of the 5-HT(1A) agonist 8-OH-DPAT. The ED(50) of 6 in the two lever drug discrimination paradigm using rats trained to discriminate the 5 HT(1A) agonist LY293284 was 0.17 micromol/kg, and the K(i) at [(3)H]8-OH-DPAT labeled 5-HT(1A) receptors was 0.23 nM. The results indicate that fluorination of hallucinogenic tryptamines generally has little effect on 5-HT(2A/2C) receptor affinity or intrinsic activity. Affinity at the 5-HT(1A) receptor was reduced, however, in all but one example, and all of the compounds tested were full agonists but with reduced functional potency at this serotonin receptor subtype. The one notable exception was 4-fluoro-5-methoxy-DMT (6), which had markedly enhanced 5-HT(1A) receptor affinity and functional potency. Although it is generally considered that hallucinogenic activity results from 5-HT(2A) receptor activation, the present results suggest a possible role for involvement of the 5 HT(1A) receptor with tryptamines. PMID- 11101362 TI - Formaldehyde-induced alkylation of a 2'-aminoglucose rebeccamycin derivative to both A.T and G.C base pairs in DNA. AB - Rebeccamycin derivatives represent a promising class of antitumor agents. In this series, two glycosylated indolocarbazoles, NB-506 and NSC-655649, are currently undergoing clinical trials. Their anticancer activities are associated with their capacities to interact with DNA and to inhibit DNA topoisomerases. Previous studies revealed that the planar indolocarbazole chromophore can intercalate into DNA, locating the appended carbohydrate residue in one of the two helical grooves, probably the minor groove as is the case with the anthracyclines and other DNA-binding antibiotics. The sugar residue contributes significantly to the DNA binding free energy of NB-506. However, the exact positioning of the glycosyl residue of rebeccamycin derivatives in the drug-DNA complex remains poorly understood. To better understand how glycosylated indolocarbazoles interact with DNA, we investigated the interaction of a rebeccamycin derivative (85) bearing a 2'-amino group on the sugar residue. We show that the presence of the 2'-amino function permits the formation of covalent drug-DNA complexes in the presence of formaldehyde. Complementary biochemical and spectroscopic measurements attest that 85 reacts covalently with the 2-amino group of guanines exposed in the minor groove of the double helix, as is the case with daunomycin. In contrast to daunomycin, 85 also forms cross-links with an oligonucleotide containing only A.T base pairs. The covalent binding to A.T base pairs was detected using a gel mobility shift assay and was independently confirmed by thermal denaturation studies and by fluorescence measurements using a series of synthetic polynucleotides. The HCHO-mediated alkylation reaction of the drug with A.T base pairs apparently involves the 6-amino group of adenines exposed in the major groove whereas the covalent attachment to G.C base pairs implicates the 2-amino group of guanines situated in the opposite minor groove. Therefore, the results suggest that either the drug is able to switch grooves in response to sequence or it can simultaneously bind to both the minor and major grooves of the double helix. This study will help to guide the rational design of new DNA-binding antitumor indolocarbazole drugs and also provides a general experimental approach for probing minor versus major groove interactions between small molecules and DNA. PMID- 11101363 TI - Natural product biosynthesis: a new interface between enzymology and medicine. PMID- 11101365 TI - Complexation between tert-butyl ketones and beta-cyclodextrin. Structural study by NMR and MD simulations AB - A structural study (NMR and MD) of the complexation between tert-butyl ketones and beta-cyclodextrin has been performed. A priority order for the alkyl and phenyl groups composing the ketones has been determined based on association constants: Ph- > C(6)H(11)- = t-Bu- > Bu-, Pr-, Me-. Geometries for the complexes are proposed based on NOE values and on the MD simulations. Bimodal complexation occurs in all the compounds studied. PMID- 11101364 TI - Studies on the synthesis of trans-dihydrodiols of polycyclic aromatic thiaarenes as potential proximate carcinogenic metabolites: first synthesis of trans-10,11 dihydroxy-10,11-dihydroacenaphtho[1, 2-b]benzo[d]thiophene and 6,7-dihydroxy-6,7 dihydronaphtho[1, 2-b]thiophene. AB - Polyaromatic thiophene compounds are found to occur concomitantly with numerous coal-derived products and shale oils and are suspected mutagens and/or carcinogens. The first synthesis of the two title compounds 9 and 16 has been achieved in five or four steps starting from 8,9-dihydroacenaphtho[1,2 b]benzo[d]thiophene (1) and 7-methoxynaphtho[1,2-b]thiophene (12), respectively. Compound 1 was converted to the cis-diol (11) (via treatment with OsO(4)/pyridine) or to trans-diol (3) [via Prevost reaction (PhCOOAg/I(2)) followed by hydrolysis] in 95-98% yield, respectively. Subsequent dehydration (PTS/benzene) of the diol followed by aromatization of the resulting ketone (5) produced the phenolic compound 6 in 97% yield. Oxidation of the phenol with phenyl iododiacetate followed by hydrolysis of the o-quinone monoketal 7 gave the o-quinone (8) in 86% yield. Stereoselective reduction of 8 with NaBH(4)/EtOH under oxygen afforded trans-10,11-dihydroxy-10,11-dihydroacenaphtho[1,2 b]benzo[d]thi oph ene(9) (orange yellow solid) in 55% yield. Compound 16 was obtained as a colorless solid, through the stereoselective reduction of the o quinone 15 (with NaBH(4)), which in turn was prepared from 12 following the protocol of functional group transformation of methoxy --> phenol --> o-quinone monoketal --> o-quinone, as used in the previous case. The yields for all the steps are very good. The mutagenicity assay of compound 9 and 16 as well as their parent thiaarenes have been performed. The results showed that 9 may not be the proximate carcinogen of acenaphtho[1,2-b]benzo[d]thiophene, while it is likely that compound 16 is one of the possible proximate carcinogens for naphtho[1,2 b]thiophene. PMID- 11101366 TI - General and efficient syntheses of C(18)-4,8-sphingadienines via S(N)2'-type homoallylic coupling reactions mediated by thioether-stabilized copper reagents. AB - The stereoselective syntheses of C(18)-4,8-sphingadienines 3 and 4 as analogues of sphingosine 1 are described. The key step in these syntheses involved a novel S(N)2'-type homoallylic coupling reaction between the corresponding thioether stabilized allylic copper reagents and the allylic mesylate 7. The thioether stabilized allylic copper reagents were easily prepared and retained the configuration of their double bond during the coupling reactions, thus overcoming the problem of isomerization which was normally associated with the use of allylic organometallic reagents in such applications. PMID- 11101367 TI - Copper(I)-assisted mild and convenient synthesis of new Se--N heterocycles: access to a promising class of GPx mimics. AB - Benzisoselenazolines 15 and benzisoselenazines 21, designed as low molecular weight mimics of glutathione peroxidases, were synthesized for the first time. Starting from amines 13 and 14, a smooth introduction of selenium in nonactivated aryl bromides using KSeCN in the presence of CuI was developed. An equimolar quantity of CuI and the presence of Et(3)N as a base are necessary to achieve a complete conversion of the starting material. The reaction is feasible in various solvents such as DMF, acetonitrile, and THF. The desired new Se-N heterocycles 15 and 21 were isolated under optimized conditions in yields of 82 and 68%, respectively. Experiments have been conducted with various copper(I) and copper(II) salts, a chloroamine 17, an aryl bromide 18, and an N-acylated amine 19 to show the scope and the limitations of this method. The previously unknown sulfur analogues 20 and 22 have been synthesized in moderate yields using a slightly modified procedure. Finally, a mechanistic scheme has been proposed to discuss some interesting findings, which were obtained during the optimization process of this new introduction of selenium. PMID- 11101368 TI - Mechanisms of the oxidations of NAD(P)H model Hantzsch 1,4-dihydropyridines by nitric oxide and its donor N-methyl-N-nitrosotoluene-p-sulfonamide. AB - 4-Substituted derivatives of Hantzsch 1,4-dihydropyridine were treated by nitric oxide (NO) or its donor N-methyl-N-nitrosotoluene-p-sulfonamide (MNTS) to give the corresponding pyridine derivatives. When the 4-substituted group was methyl, ethyl, n-propyl, and aryl groups, it was preserved, but when the group was isopropyl or benzyl one, it was lost. 2,3-Dichloro-5, 6-dicyano-1,4-benzoquinone (DDQ) was used in place of NO and MNTS to react with the 4-substituted Hantzsch 1,4-dihydropyridines, no the corresponding 4-dealkyl Hantzsch pyridines were obtained from all the reactions. 1-Benzyl-1,4-dihydronicotinamide (BNAH), a close analogue of Hantzsch 1,4-dihydropyridine (HEH), was used instead of HEH to react with either of NO and MNTS, no reactions were observed for 3 days. Replacement of HEH by N-d-HEH and HEH-4,4-d(2) to react with NO, MNTS and DDQ gave the observed kinetic isotope effects of 3.1 and 1.4 for NO, 1.1 and 1.3 for MNTS, and 1.1 and 2.1 for DDQ, respectively. When p-dinitrobenzene, an electron-transfer inhibitor, was added into the title reaction systems, no remarkable inhibitory effect was observed. These results indicated that the oxidation of HEH by NO was initiated by hydrogen transfer from the N(1)-position to give the corresponding aminyl radical, which then underwent homolytic cleavage to become the final aromatized product (A). But the reaction of HEH with MNTS was initiated by nitrosation to give the corresponding N-nitroso compound, which was subsequently subjected to two steps of homolytic cleavage to afford the aromatized Hantzsch pyridine A. PMID- 11101369 TI - Synthesis and structure of 4-O,6-O-glycosylidene glycosides. AB - Interglycosidic spiro ortho esters (9-20) were efficiently prepared from methyl 2,6-di-O-benzylgluco- or galactopyranoside and various sugar lactones in the presence of methoxytrimethylsilane and a catalytic amount of trimethylsilyl triflate. All of the prepared sugar ortho esters possess perhydrospiro[2H-pyran 2,2'-pyrano[3,2-d][1,3]dioxin] ring systems commonly in their molecules and, remarkably, were afforded as single isomers. The configurations of the spiro centers in their molecules were determined or estimated by X-ray single crystallographic analysis and molecular modeling studies. By comparing the conformations of prepared ortho esters, we revealed that the difference in the stability between two possible isomers was principally caused from that between the spiro ring systems in their molecules in each case. PMID- 11101370 TI - A highly stereoselective construction of beta-glycosyl linkages by reductive cleavage of cyclic sugar ortho esters. AB - The preparation of beta-glycosides by the reductive cleavage of spiro sugar ortho esters is described in this report. This procedure is based on a concept completely different from those of other methods for glycosylation. Twelve sugar ortho esters that commonly possess perhydrospiro[2H-pyran-2,2'-pyrano[3,2 d][1,3]dioxin] ring systems in their molecules were reduced by LiAlH(4)/AlCl(3) or NaCNBH(3)/AlCl(3). Among these ortho esters, those (9a-12a) prepared from the D-sugar lactones (1-4) and 2, 3-di-O-benzyl-alpha-D-glucopyranoside (7) or those (19a, 20a) prepared from the L-sugar lactones (5, 6) and 2, 3-di-O-benzyl-alpha-D galactopyranoside (8) were selectively converted into beta-(1 --> 4)-glycosides (9b-12b or 19b, 20b) in excellent yields by the treatment of LiAlH(4)/AlCl(3). In contrast, the ortho esters (13a-16a or 17a, 18a) that were prepared from combinations of the D-sugar lactones and 8 or those of the L-sugar lactones and 7 were efficiently reduced with NaBH(3)CN/AlCl(3) to afford beta-(1 --> 6) glycosides (13b-16b or 17b, 18b) selectively. It was remarkable that the resulting disaccharides were obtained with extremely high beta-selectivity even in the cases with mannosyl or rhamnosyl glycosides. Moreover, these products would be useful units for the construction of branched saccharides, because the newly formed hydroxy groups could be again glycosylated without further deprotection procedures. The high regio- and stereoselectivity was totally explained by considering the structures and the conformations of these ortho ester molecules and the stereoelectoronic effects of their spiro ring systems. In addition, the preparation of the sugar ortho esters with glucosamine derivatives and the reactivity of these ortho esters are described in this report. N Phthaloyl glucosamine derivatives (21, 22) were efficiently reacted with the benzyl-protected gluconolactone (1) in the presence of TMSOMe and TMSOTf to afford ortho esters (23a-c). After the conversion of the phthalimido functionality to the dibenzyl amino group, glucosylideneglucosamine (25) was reduced with LiAlH(4)/AlCl(3) to afford beta-(1 --> 4)-glycoside (26) selectively. PMID- 11101371 TI - Mechanism of the AB - Stereochemical studies on [2 + 2] photoaddition of cis-/trans-4-propenylanisole (cis-1 and trans-1) and cis-1-(p-methoxyphenyl)ethylene-2-d(1) (cis-3-d(1)) to C(60) exhibit stereospecificity in favor of the trans-2 cycloadduct in the former case and nonstereoselectivity in the latter. The observed stereoselectivity in favor of the cis-6-d(3) [2 + 2] diastereomer by 12% in the case of the photochemical addition of (E)-1-(p-methoxyphenyl)-2-methyl-prop-1-ene-3,3,3-d(3) (trans-5-d(3)) to C(60) is attributed to a steric kinetic isotope effect (k(H)/k(D) = 0.78). The loss of stereochemistry in the cyclobutane ring excludes a concerted addition and is consistent with a stepwise mechanism. Intermolecular secondary kinetic isotope effects of the [2 + 2] photocycloaddition of 3-d(0) vs 3-d(1), and 3-d(6) as well as 5-d(0) vs 5-d(1), and 5-d(6) to C(60) were also measured. The intermolecular competition due to deuterium substitution of both vinylic hydrogens at the beta-carbon of 3 exhibits a substantial inverse alpha secondary isotope effect k(H)/k(D) = 0.83 (per deuterium). Substitution with deuterium at both vinylic methyl groups of 5 yields a small inverse k(H)/k(D) = 0. 94. These results are consistent with the formation of an open intermediate in the rate-determining step. PMID- 11101372 TI - Pyrrole-inverted isomer of 5,10,15,20-tetraaryl-21-selenaporphyrin AB - A novel 5,10,15,20-tetraaryl-21-selenaporphyrin isomer with an inverted pyrrole ring, i.e., 5,10,15, 20-tetraaryl-2-aza-21-carba-22-selenaporphyrin (SeC-TArPH) has been produced by a [3 + 1] condensation of 2, 5 bis(phenylhydroxymethyl)selenophene and 5,10-ditolyltripyrrin. The reaction yielded 5,20-diphenyl-10,15-bis(p-tolyl)-21-selenaporphyrin Se-DPDTPH (19%) and its isomer with an inverted pyrrole ring, i.e., 5,10-diphenyl-15,20-bis(p-tolyl) 2-aza-21-carba-22-selenaporphyrin, SeC-DPDTPH (1%). Mechanistically the synthesis of SeC-DPDTPH requires one beta-condensation at the pyrrole moiety of 5, 10 ditolyltripyrrin instead of the stereotypical alpha-condensation. The identity of inverted selenaporphyrin has been confirmed by high-resolution mass spectrometry and (1)H NMR spectroscopy. A saddle distortion mode for the inverted selenaporphyrin macrocycle SeC-DPDTPH has been determined by X-ray crystallography. NMR spectra are consistent with the existence of tautomeric equilibria that involve three tautomeric species of the neutral form of SeC DPDTPH. The preference for the tautomer with the labile proton located at the peripheral N(2) nitrogen atom has been detected in pyridine-d(5) solution. The density functional theory (DFT) has been applied to determine the molecular and electronic structure of three tautomers of 2-aza-21-carba-22-selenaporphyrin: 2 N, 23-N, 24-NH, 2-N, 23-NH, 24-N, and 2-NH, 23-N, 24-N formally created from SeC DPDTPH by a replacement of phenyl and tolyl groups with hydrogen. The total energies calculated using the B3LYP/6-311G//B3LYP/6-311G approach, demonstrate that relative stability of postulated tautomers decreases in the order 2-N, 23 NH, 24-N > 2-N, 23-N, 24-NH > 2-NH, 23-N, 24-N. The small energy differences between tautomeric species suggests their simultaneous presence in equilibrium. PMID- 11101373 TI - Synthesis and surface and antimicrobial properties of novel cationic surfactants. AB - A series of surfactants with tuned polarity were prepared, including a new class of compounds: gluco-pyridinium surfactants. Pure anomers were obtained by chromatographic separation. The conductivity and surface tension of surfactant solutions in water were measured, and provided interesting information regarding their aggregation behavior. Peculiarities were observed in the premicellar range. Tensidic parameters correlated with antimicrobial activity. A few parameters, mainly the hydrophobicity of the headgroup, may play a role in finding more efficient antimicrobial structures. PMID- 11101374 TI - The synthesis and fluorescent properties of analogues of the zinc(II) specific fluorophore zinquin ester. AB - The synthesis and fluorescent properties in the absence and presence of zinc(II) of a range of 2-substituted derivatives of N-(6-methoxy-2-methyl-8-quinolyl)-4 methylbenzenesulfonamide are described. These analogues formed complexes with zinc(II) as indicated by a bathochromic shift in their UV/vis spectra. Analogues with isobutenyl and isobutyl side chains at the 2-position formed fluorescent complexes whose fluorescence was stronger than that of the 2-methyl-containing parent. These derivatives were converted, via conversion to the phenol with boron tribromide and reaction with ethyl bromoacetate, to systems with ester-containing side chains analogous to zinquin ester, a specific cellular fluorophore for zinc(II). All of these ester derivatives formed complexes with zinc(II) resulting in a bathochomic shift in their UV/vis spectra. Compounds with isobutyl, isobutenyl, and styryl side chains exhibited increased fluorescence compared to that of zinquin ester in the presence of zinc(II). The compound with the 2 isobutyl side chain was more selective in its fluorescence for zinc(II) over cadmium(II) compared to zinquin ester. PMID- 11101375 TI - N-Acylbenzotriazoles: neutral acylating reagents for the preparation of primary, secondary, and tertiary amides AB - Readily available N-acylbenzotriazoles 2a-q efficiently acylate aqueous ammonia and primary and secondary amines to give primary, secondary, and tertiary amides in good to excellent yields. The wide applicability of the procedure is illustrated by the preparation of (i) alpha-hydroxyamides from alpha-hydroxy acids and of (ii) perfluoroalkylated amides. PMID- 11101376 TI - Regioselective enzymatic acylations of polyhydroxylated eudesmanes: semisynthesis, theoretical calculations, and biotransformation of cyclic sulfites. AB - Different lipase enzymes have been tested in order to perform regioselective acetylations on the eudesmane tetrol from vulgarin. High yields (95%) of 1,12 diacetoxy derivative (4) were achieved in 1 h with Candida antarctica lipase (CAL). However, only the 12-acetyl derivative (6) was obtained in similar yield with Mucor miehei (MML) or Candida cylindracea (CCL) lipases. The enzymatic protection at C-1 and C-12 has been used to form eudesmane cyclic-sulfites between C-6 and C-4 atoms. The R/S-sulfur configuration has been assigned by means of the experimental and theoretical (13)C and (1)H NMR chemical shifts. The theoretical shifts were calculated using the GIAO method, with a MM+ geometry optimization followed by a single-point calculation at the B3LYP/6-31G(*) level (B3LYP/6-31G(*)//MM+). Moreover, B3LYP/6-31G(*) geometry optimizations were carried out to test the B3LYP/6-31G(*)//MM+ results, for the deacetylated sulfites (12 and 15). In addition to the delta(C) and delta(H) shifts, the (3)J(HH) coupling constants were also calculated and compared with the experimental values when available. Finally, different reactivities have been checked in both sulfites by biotransformation with Rhizopus nigricans. While the R-sulfite gave 2 alpha- and 11 beta-hydroxylated metabolites, the S-sulfite yielded only regioselective deacetylations. Furthermore, both sulfites showed different reactivities in redox processes. PMID- 11101377 TI - Conversion of bis(trichloromethyl) carbonate to phosgene and reactivity of triphosgene, diphosgene, and phosgene with methanol(1) AB - Triphosgene was decomposed quantitatively to phosgene by chloride ion. The reaction course was monitored by IR spectroscopy (React-IR), showing that diphosgene was an intermediate. The methanolysis of triphosgene in deuterated chloroform, monitored by proton NMR spectroscopy, gave methyl chloroformate and methyl 1,1, 1-trichloromethyl carbonate in about a 1:1 ratio, as primary products. The reaction carried out in the presence of large excess of methanol (0.3 M, 30 equiv) was a pseudo-first-order process with a k(obs) of 1.0 x 10( )(4) s(-)(1). Under the same conditions, values of k(obs) of 0.9 x 10(-)(3) s( )(1) and 1.7 x 10(-)(2) s(-)(1) for the methanolysis of diphosgene and phosgene, respectively, were determined. The experimental data suggest that, under these conditions, the maximum concentration of phosgene during the methanolysis of triphosgene and diphosgene was lower than 1 x 10(-)(5) M. Methyl 1,1,1 trichloromethyl carbonate was synthesized and characterized also by the APCI-MS technique. PMID- 11101378 TI - Synthesis and properties of aminoacylamido-AMP: chemical optimization for the construction of an N-acyl phosphoramidate linkage. AB - This paper describes the design and synthesis of a new type of aminoacyl adenylate analogue (aa-AMPN) having an N-acyl phosphoramidate linkage where the oxygen atom of the mixed anhydride bond of aminoacyl-adenylate (aa-AMP) is replaced by an amino group. This new type of aa-AMP analogue is expected to be useful as material for studies on the recognition mechanism of the aminoacylation of tRNA and other biochemical reactions. The condensation of phosphoramidite derivatives of carboxamides with nucleoside derivatives failed, because the activated phosphoramidite derivatives reacted with not only the hydroxyl groups but also another reactive species. An alternative approach was examined by the reaction of 5'-O-phosphoramidite adenosine derivatives with carboxamide derivatives. The TBTr and TSE groups were chosen for protection of the amino group of amino acid amides and the phosphate group, respectively. Detailed studies revealed that the use of 5-(3,5-dinitrophenyl)-1H-tetrazole as an activating catalyst of phosphoramidites resulted in rapid condensation within 10 min to give fully protected aa-AMPN derivatives. No side reaction occurred. Deprotection of these products via a two-step procedure gave aa-AMPN derivatives in good yields. It also turned out that aa-AMPNs thus obtained are stable under both acidic and basic conditions, such as 0.1 M HCl (pH 1.0) and 0.1 M NaOH (pH 13.0). PMID- 11101379 TI - Highly diastereoselective synthesis of the 1,1'-binaphthol unit on a bile acid template AB - The use of 7-deoxycholic acid as a chiral template in the asymmetric syntheses of 1,1'-binaphthyl-2,2'-diol derivatives is reported. Intramolecular coupling of compounds 7 and 11 have been carried out with Mn(acac)(3) in CH(3)CN to afford coupled binaphthol products 8 and 12 with 65% and >99% diastereoselectivity, respectively. In both cases the predominant formation of the (S) isomers were predicted by computer modeling studies. This was confirmed in the case of compound 12. PMID- 11101380 TI - Multivalency effects in protein--carbohydrate interaction: the binding of the Shiga-like toxin 1 binding subunit to multivalent C-linked glycopeptides. AB - A series of monovalent and bivalent glycopeptides displaying a C-linked analogue of the Pk trisaccharide, the in vivo ligand for the pentavalent Shiga-like toxin binding subunit (SLT-1B), were prepared and evaluated as ligands for SLT-1B by isothermal titration microcalorimetry and competitive enzyme-linked immunosorbent assay (ELISA). Although none of the monovalent ligands showed any enhancement in affinity compared to O-methyl glycoside, two bivalent ligands show significant enhancements in affinity in assays. This observation represents the first calorimetric observation of an enhancement in affinity for this system. In contrast, only one of the two ligands shows an enhancement in the competitive ELISA. Together, these data signal a difference in the means by which the two ligands achieve affinity, apparently triggered by a change in the nature of the linker domain. These results provide a rationalization for apparently contradictory reports from the recent literature and again emphasize the importance of investigating complex binding phenomena by multiple techniques. PMID- 11101381 TI - The first density functional study on the AB - The hetero-Diels-Alder reactions of 1,2-diaza-1,3-butadiene with ethylene, methyl vinyl ether, and methyl acrylate have been investigated theoretically with the aid of density functional theory using the Becke3LYP/6-31G(d) computational level. In the reactions with substituted alkenes, the transition states are concerted but asynchronous; the shortest bond-forming distance involves the nonsubstituted carbon of the alkene. In agreement with the experimental results, the reaction with methyl vinyl ether proceeds with high endo stereoselectivity and with complete regioselectivity to form the 6-methoxy cycloadduct. The conformational study of the transition states shows a sharp s-trans preference. In contrast, the [4 + 2]-cycloaddition of 1,2-diaza-1,3-butadiene with methyl acrylate have been found to occur with low regio- and stereoselectivity but with a s-cis preference in the transition structures. PMID- 11101382 TI - Complexation of fullerenes with 5,5'-Biscalix AB - 5,5'-Biscalix[5]arene, prepared by oxidative coupling of the tetrabenzoyl ester of calix[5]arene (shown by X-ray crystallographic analysis to have a o,u,u,d,u conformation), forms complexes with C(60) (K(assoc) = 43 M(-)(1)) and C(70) (K(assoc) = 233 M(-)(1)). The X-ray crystallographic structure of the C(60) complex reveals its clamshell-shaped architecture, presumably the result of a change in the conformation of biscalix[5]arene from anti (uncomplexed) to syn (complexed). PMID- 11101383 TI - Chemo-enzymatic total synthesis of 3-epiaustraline, australine, and 7-epialexine. AB - Sequential enzymatic aldol reaction and bis-reductive amination leads to the total syntheses of tetrahydroxylated pyrrolizidine alkaloids, 3-epiaustaline (14), australine (1), and 7-epialexine (11). This approach allows for their rapid construction without the need for protecting group manipulation of the hydroxyl functionality. In addition, an improved procedure for the asymmetric epoxidation of divinyl carbinol (3) was described, and the product was used in a concise synthesis of the required triol 7 and ent-7. PMID- 11101384 TI - New crown annelated tetrathiafulvalenes: synthesis, electrochemistry, self assembly of thiol derivatives, and metal cation recognition AB - The synthesis of new S(2)O(4)-crown annelated tetrathiafulvalene (TTF) derivatives substituted with one terminal thiol group is described. Self assembled monolayers (SAMs) of these compounds have been assembled on gold and platinum surfaces, the latter substrate giving improved quality films. SAMs of TTF derivative 16b are the most stable of those we have studied. Electrochemical data for SAMs of 5a, 5b, 8, 16a, and 16b in acetonitrile reveal two reversible one-electron waves, typical of the TTF system; the current increased linearly with scan rate, indicating a surface wave response. SAMs of 8, 16a, and 16b exhibited an electrochemical response in aqueous electrolytes, which was observed between 50 and 100 cycles. Moreover, if the potential scanned was limited to the first TTF oxidation, the cyclic voltammetry response was observed for at least 1000 cycles. Metal complexation by the crown ionophore of the SAMs in acetonitrile has been monitored by a positive shift in the first oxidation potential of the TTF unit (maximum DeltaE(1)(1/2) = 80 mV for Ag(+)). We also report the X-ray crystal structure of TTF-crown derivative 21 bearing two hydroxymethyl substituents, synthesized during the course of this work. The structure is characterized by infinite chains of molecules linked by strong intrachain hydrogen bonds between the terminal hydroxy groups. PMID- 11101385 TI - Acyclic stereoselection in the reaction of nucleophilic reagents with chiral N acyliminium ions generated from N- AB - Optically active N-[1-(phenylsulfonyl)alkyl]imidazolidin-2-ones react at low temperature in the presence of tin tetrachloride to give acyclic N-acyliminium ions. These electrophilic substrates give addition products upon reaction with pi nucleophiles. Allyltrimethylsilane affords the corresponding allylated products in good yields and high diastereoselectivity. The stereochemical outcome of this process can be rationalized by taking into account the preference of the intermediate N-acyliminium ion for an E configuration that favors the attack of the nucleophile from the si-si face. Disappointing results are obtained using silyl ketene acetals; conversely trimethylsilyl enol ether of acetophenone gives the corresponding adducts in high diastereoselectivity. The utilization of trimethylsilyl enol ether of 2-acetylfuran is particularly interesting since the corresponding adducts are obtained with good diastereoselectivity and the furan ring could be amenable of further synthetic transformations. PMID- 11101386 TI - Synthesis of cone, partial-cone, and 1,3-alternate 25, 27-Bis AB - The preparation of 25,27-bis[1-(2-ethyl)hexyl]- and 25, 27-bis[1-(2-tert butoxy)ethyl]calix[4]arene-crown-6 combining one polyether crown-6 and one alkylchain O-attached on each side of a calix[4]arene in the cone, partial-cone, and 1,3-alternate conformations are reported. The control over 25, 27 bisalkylcalix[4]arene-crown-6 conformation via varying specific reaction conditions was studied. The series of calix[4]arenes have been prepared by two routes, which differ in the order in which the alkyl or polyether groups were introduced. Moreover, methods have been developed to selectively prepare the cone and partial-cone conformers by using an appropriate base in the alkylation reactions. The conformations of these new derivatives have been probed by (1)H NMR analysis and X-ray crystallography. The (1)H and (13)C NMR spectra of 25,27 bis[1-(2-ethyl)hexyl]calix[4]arene-crown-6, 1, 3-alternate 1, cone 2, and partial cone 3 are also discussed. PMID- 11101387 TI - Synthesis of vitamin D(3) and calcitriol dimers as potential chemical inducers of vitamin D receptor dimerization. AB - The design and synthesis of vitamin D(3) dimers 2 and 3 and 1 alpha, 25 dihydroxyvitamin D(3) (calcitriol) dimers 4 and 5 are described. The dimers were designed with a view to doubly binding the vitamin D receptor (VDR) and inducing the receptor homodimerization. In the dimers the units are linked through the C 11 position in ring C by an alkyl side chain of six or 10 carbon atoms, far from the hydroxy groups responsible for the VDR binding. The linker is formed by olefin metathesis of an olefinic side chain at the C-11 position introduced by stereoselective cuprate addition. The synthesis, which is both short and convergent, uses the Wittig-Horner approach to construct the vitamin D triene system and allows the preparation of dimers with a linker of modulated length with the purpose of optimizing the vitamin D(3)-VDR interaction. PMID- 11101388 TI - A convergent synthesis of Hexahomotriazacalix AB - A new, convergent synthesis of hexahomotriazacalix[3]arenes 1a-e is described. The key transformation in this synthesis involves the coupling of the triamines 4a-d with 2, 6-bis(chloromethyl)-4-methylphenol 5 and results in the formation of the hexahomotriazacalix[3]arenes 1a-d in 90-95% yield. The triamines 4a-d were constructed by the one-pot reaction of monochloroaldehyde 3 and a primary amine followed by reduction to yield the triamines 4a-d in 50-55% yield. Deallylation of macrocycle 1d was accomplished by palladium catalysis to obtain the N unsubstituted macrocycle 1e, which has the potential to be a precursor to a variety of N-substituted hexahomotriazacalix[3]arenes. PMID- 11101389 TI - Synthesis and complexation properties of poly(ethylene glycol)-linked mono- and bis-dioxocyclams. AB - Both tri- and tetra(ethylene glycol) linked bis-chromium carbene complexes have been synthesized. These carbene complexes were photolyzed with N-protected imidazolines to give protected azapenams. These were transformed into polyether linked basket dioxocyclams 4a,b and bis-dioxocyclams 5a,b. These compounds have cavities for the complexation of both "hard" and "soft" metal ions. The complexes of Ni, Ba, and Gd were synthesized. PMID- 11101390 TI - Development of an efficient, regio- and stereoselective route to libraries based on the beta-D-glucose scaffold. AB - The design and execution of an efficient synthetic route for the sequential functionalization of the five hydroxyl substituents of beta-D-glucose has been achieved. This strategy, based on the stereoselective glycosidation of thioglycosides, provides a new approach for the parallel construction of a wide variety of beta-D-glucose analogues and as such holds promise for solid-support library syntheses. PMID- 11101391 TI - First total synthesis of ent-gelsedine via a novel iodide-promoted allene N acyliminium ion cyclization. AB - The first total synthesis of the oxindole alkaloid gelsedine (1) starting from (S)-malic acid is described. The key step is a novel iodide-promoted intramolecular reaction of an allene with an N-acyliminium ion intermediate which provided in a single step the bicyclic vinyl iodide 11. Other important steps are the highly stereoselective Pd-catalyzed Heck cyclization of N-methylanilide 23a which led to the desired spiro-oxindole 24a, the fully regioselective intramolecular oxymercuration of 25a to the desired cyclic ether, and the remarkable oxindole N-demethylation of 29 via a radical mechanism by using dibenzoyl peroxide. The total synthesis was concluded by the stereoselective introduction of the ethyl group from the bis-Boc compound 41 followed by methoxylation of the oxindole nitrogen. This total synthesis leads to the unnatural (+)-enantiomer of gelsedine in 21 steps and 0.10% overall yield. PMID- 11101393 TI - Development of a protocol for eight- and nine-membered ring synthesis in the annulation of sp(2),sp(3)-hybridized organic dihalides with keto esters. AB - A protocol has been developed in which annulation reactions of various dihalides with keto esters can be carried out to provide entry to eight- and nine-membered carbocycles. In this process wherein one alkenyl- or aryl bromide and a tethered alkyl chloride comprise the organic dihalide, a selective metal-halogen exchange reaction between the sp(2)-hybridized bromide and an organolithium initiates the process. Transmetalation to an organoytterbium reagent generates a species that undergoes selective carbonyl addition to the ketone of the keto ester, creating a lactone intermediate. Subjection of the resulting chloroalkyl lactone to intramolecular reductive coupling with samarium(II) iodide completes the desired annulation. PMID- 11101392 TI - Azidomercurations of alkenes: mercury-promoted Schmidt reactions. AB - Azides bearing a suitably disposed alkene, when treated with either mercuric perchlorate or mercuric trifluoromethanesulfonate, produce bicyclic iminium ions. This new version of the Schmidt reaction proceeds by capture of the mercuronium ion intermediate by the azide to produce an aminodiazonium ion, which suffers a 1,2-shift to give an iminium ion (e.g., 10 --> 16 --> 17 --> 18). Reduction of the iminium ion may then be carried out to produce an amine. Compared to earlier work on the protic acid-promoted intramolecular Schmidt reaction of azido alkenes, the mercury-promoted Schmidt reaction has several advantages. First, the acid-promoted Schmidt reaction of azido-alkenes requires that the intermediate carbocations be tertiary, allylic, benzylic, or propargylic. The mercury-promoted method has no such limitation; thus even 1,2-disubstituted alkenes may be used. Second, the mercury-promoted method is milder, allowing the presence of acid sensitive functionality. The protic version, typically employing trifluoromethanesulfonic acid, is limited in its functional group tolerance. Third, whereas carbocation rearrangement is often observed prior to cyclization/rearrangement in the acid-promoted Schmidt reaction, the mercury promoted method avoids this problem. Fourth, the presence of the mercurio group during the rearrangment may alter the regioselectivity of the 1,2-migration. Finally, the mercury-bearing iminium ions that are the result of the Schmidt reaction were found to be sensitive to protodemercuration, precluding their use in other transformations. PMID- 11101394 TI - 3,3'-Bis(diphenylphosphino)-1,1'-disubstituted-2,2'-biindoles: easily accessible, electron-rich, chiral diphosphine ligands for homogeneous enantioselective hydrogenation of oxoesters AB - Racemic (+/-)-3,3'-bis(diphenylphosphinyl)-1,1'-dimethyl-2, 2'-biindole (1c) (N Me-2-BINPO) and (+/-)-3, 3'-bis(diphenylphosphinyl)-1,1'-bis(methoxymethyl)-2,2' biindole (1d) (N-MOM-2-BINPO) were synthesized in satisfactory yields following a three-step reaction sequence, starting from indole. Resolution of racemic 1c and 1d was achieved through fractional crystallization of their diastereomeric adducts with optically active dibenzoyl tartaric acids, followed by alkaline decomplexation of the diastereomerically pure salts. Their trichlorosilane reduction gave enantiopure phosphines (+)- and (-)-(1a) (N-Me-2-BINP) and (+)- and (-)-(1b) (N-MOM-2-BINP). The electrochemical oxidative potential of 1a and 1b was found to be 0. 52 and 0.60 V, respectively. Both the enantiomers of (1a) were tested as ligands of Ru(II) in asymmetric hydrogenation reactions of alpha- and beta-oxoesters. Reactions were found to be outstandingly fast and enantioselection quite good. Comparative kinetic experiments on the hydrogenation reaction of methyl acetoacetate carried out with 1a, 1c, BINAP, and other biheteroaromatic diphosphines as ligands of Ru(II) demonstrated that all the reactions follow a first-order kinetic. A linear relationship was found between the kinetic constant log and the electrochemical oxidative potential of the diphosphine ligand. PMID- 11101395 TI - Computational study of the electronic structure of substituted phenylcarbene in the gas phase AB - Gas-phase calculations for ortho-, meta-, and para-substituted phenylcarbenes demonstrate that aromatic ring substituents can have a large effect on the singlet-triplet splitting in these molecules. For example, p-aminophenylcarbene is predicted to have a singlet-triplet splitting (DeltaE(ST)) of 0.7 kcal/mol, which is 4.7 kcal/mol smaller than that of the parent phenylcarbene. However, when the mesomeric effect of pi-donation is removed, as in the meta-substituted molecules, this affect is greatly attenuated. m-Amino substitution lowers the DeltaE(ST) to 5.2 kcal/mol, a change of only 0.2 kcal/mol from the parent phenylcarbene. The sensitivity of phenylcarbene to substitution is judged by the slope of the line correlating the singlet-triplet gaps to linear free energy (LFE) parameters. In this study, the best linear correlation for DeltaE(ST) in the para-substituted phenycarbenes is observed when plotted against sigma(p)(+). The resultant slope (rho) is 5.0. The origin of this effect is preferential interaction of aryl substituents with the singlet rather than the triplet species as demonstrated by electron density analyses. PMID- 11101396 TI - Additions of enantioenriched allenylzinc and indium reagents to lactic aldehyde ethers AB - Allenylzinc and indium reagents were generated in situ through Pd(0)-catalyzed metalation of (R)- and (S)-3-butyn-2-ol methanesulfonate with Et(2)Zn and InI. These reagents add to the benzyl and TBS ethers of (S)-lactic aldehyde to afford diastereomeric stereotriads in moderate to high yield. The (S)/(S) combination afforded the anti,anti adducts with 94:6-100:0 diastereoselectivity. The (R)/(S) combination was mismatched, affording a mixture of anti,syn and syn,anti adducts in diastereomeric ratios of ca. 80:20-85:15. Addition of the racemic allenylmetal reagents to the (S)-lactic aldehyde ethers afforded the products of matched and mismatched pairings in equal amount. PMID- 11101397 TI - Synthesis and properties of chiral Tetrahomodiazacalix PMID- 11101398 TI - Use of the ramberg-Backlund rearrangement for the synthesis of medium and large heterocyclic alkenes: stereoselective olefin formation PMID- 11101399 TI - A new synthesis of a key intermediate of beta-lactam antibiotics via diastereoselective alkylation of beta-hydroxy ester. PMID- 11101400 TI - Chemical evidence for thiyl radical addition to the C6-position of a pyrimidine nucleoside and its possible relevance to DNA damage amplification. PMID- 11101401 TI - Alcohol inversion using cesium carboxylates and DMAP in toluene. PMID- 11101402 TI - Efficient lewis acid catalyzed intramolecular cannizzaro reaction PMID- 11101403 TI - Cycloaddition of lewis acid-induced N-methyleneanilines as azadienes to 1,2 bistrimethylsilyloxycyclobutene and oxidative ring expansion to 1,2,4,5 tetrahydro-1-benzazocine-3,6-diones(1) PMID- 11101404 TI - Efficient synthesis of a stereochemically defined carbohydrate scaffold: carboxymethyl 2-acetamido-6-azido-4-O-benzyl-2-deoxy-alpha-D-glucopyranoside. PMID- 11101405 TI - Preparation and reactivities of novel (Diacetoxyiodo)arenes bearing heteroaromatics PMID- 11101406 TI - A novel approach for mannich-type bases having a terminal olefin: zinc triflate and water-promoted Cyclization/C-N bond cleavage process PMID- 11101407 TI - A simple chemoselective method for the deprotection of acetals and ketals using bismuth nitrate pentahydrate PMID- 11101409 TI - Regiocontrolled synthesis of the antitumor antibiotic AT2433-A1 PMID- 11101408 TI - Total synthesis of anti-microtubule diketopiperazine derivatives: phenylahistin and aurantiamine. PMID- 11101410 TI - Computer-assisted structure determination. Structure Of the peptide moroidin from laportea moroides PMID- 11101411 TI - Fluorinated building blocks. The discovery of a stable difluoroallenyl indium and the synthesis of gem-difluoroallenyl and -propargyl synthons in aqueous media PMID- 11101412 TI - The first asymmetric synthesis of alpha-sulfanylphosphonates. AB - [reaction: see text] Diastereoselectivity of up to 88% was achieved for the synthesis of an alpha-mercapto gamma-unsaturated phosphonate using the readily available chiral dimenthylphosphonyl ester group and a carbanionic [2,3] sigmatropic rearrangement. Absolute configuration of the newly formed chiral center of this nonracemic thiol was determined, and the corresponding phosphono thiolane and thiolane S-oxide were also stereoselectively prepared. PMID- 11101413 TI - Photochemical and thermal bergman cyclization of a pyrimidine enediynol and enediynone. AB - [reaction: see text] Novel 10-membered pyrimidine enediynes (3 and 4) were synthesized in seven and eight steps, respectively. These compounds were compared for their abilities to undergo Bergman cyclization both thermally and photochemically. Alcohol 3 readily cyclized both thermally and photochemically in (i)PrOH, while ketone 4 only showed efficient thermal cyclization. Both compounds were also shown to cleave dsDNA under the appropriate conditions. PMID- 11101414 TI - Strong bicyclic guanidine base-promoted wittig and horner-wadsworth-emmons reactions AB - A convenient procedure to effect the Wittig and Horner-Wadsworth-Emmons reactions employs guanidine TBD and MTBD as base-promoters; mild reaction conditions, high efficiency, and facile isolation of the final products make the present methodology, at least in some cases, a practical alternative to known procedures. PMID- 11101415 TI - Annulation of the cyclohexane ring by tandem free radical alkylation of a nonactivated delta-carbon atom-intramolecular carbanion cycloalkylation AB - Annulation of the cyclohexane ring by a combination of free radical and ionic reactions sequences was achieved. Free radical alkylation of the remote nonactivated delta-carbon atom involves addition of delta-carbon radicals, generated by 1,5-hydrogen transfer in alkoxy radical intermediates, to radicophilic olefins, while the polar sequence involves enolate anions as intermediates which undergo a cycloalkylation reaction. Thus, the cyclohexane ring was constructed using diverse acyclic and cyclic structures as precursors of alkoxy radicals. PMID- 11101416 TI - Chiral sulfonamide induced enantioselective protonation of samarium enolate in the reaction of alpha,beta-unsaturated ester with ketone AB - Good enantioselectivity (up to 84% ee) has been achieved in the formation of alpha,gamma-substituted-gamma-butyrolactone by the SmI(2)-mediated reductive coupling of ketones with methyl methacrylate using various chiral proton sources. PMID- 11101417 TI - Synthesis of a triphenylphosphine reagent on non-cross-linked polystyrene support: application to the Staudinger/Aza-wittig reaction AB - A new triphenylphosphine reagent linked to a linear polystyrene was synthesized. The reactivity of this phosphine-bound polymer is superior to that of the phosphine bound to cross-linked polystyrene. The polymer reacted very rapidly with azides to generate iminophosphoranes which could then react with aldehydes to generate imines in good yields and high purities. PMID- 11101418 TI - A new polymer-attached reagent for the oxidation of primary and secondary alcohols AB - A new, polymer-bound reagent system for the efficient oxidation of primary alcohols to aldehydes and secondary alcohols to ketones in the presence of a catalytic amount of 2,2,6, 6-tetramethyl-1-piperidinyloxyl (TEMPO) is described. In most cases, workup of this heavy metal-free oxidation is achieved by simple filtration followed by removal of the solvent. In selected examples this reagent was compared with the known polymer-bound permanganate and chromium(VI) reagents. PMID- 11101419 TI - A new method for the synthesis of imidazolidinone- and benzimidazolone-containing AB - A novel acid-catalyzed double nucleophilic addition of bisthiols to heterocyclic bisacetals gives sulfur-containing macrocycles in good yield. Crossover experiments revealed that the acid-catalyzed N-acyl iminium cyclization for the imidazolidinones is reversible. The sulfur atom in the bridge was extruded photochemically, giving new [2.2]heterophanes containing the imidazolidinone and benzimidazolone ring systems. The determined crystal structures for representative members in the benzimidazolone series are also reported. PMID- 11101420 TI - BtCH(2)TMS-assisted homologation of carboxylic acids: A safe alternative to the arndt-eistert reaction AB - One-carbon homologation of carboxylic acids is achieved by (i) treatment of an acyl chloride with 1-[(trimethylsilyl)methyl]-1H-1,2, 3-benzotriazole (BtCH(2)TMS) (1) to afford N-(acylmethyl)benzotriazoles 3a-f, followed by (ii) conversion of 3a-f with triflic anhydride into RC(OTf)=CHBt 4a-f, and (iii) the subsequent reaction of 4a-c with NaOCH(3) followed by 1N HCl to afford esters RCH(2)CO(2)R' 7a-c in overall yields of 50-70%. For the aliphatic compounds 5d-f, treatment of 5d-f with p-toluenesulfonic acid followed by TBAF/THF afforded acids RCH(2)COOH 7d-f. PMID- 11101421 TI - A remarkably simple chemicoenzymatic approach to structurally complex bicyclo[3.1.0]hexane carbocyclic nucleosides. AB - [reaction: see text] Intramolecular cyclopropanation of a carbene engendered from the corresponding diazo beta-ketoester produced the desired bicyclo[3.1. 0]hexane pseudosugar. Purine nucleosides obtained via Mitsunobu coupling were resolved with adenosine deaminase. The requisite beta-ketoester was assembled in one step from ethyl acetoacetate and acrolein. PMID- 11101422 TI - Highly efficient and practical synthesis of 3,6-branched oligosaccharides. AB - [reaction: see text] A one-pot formation of the 3- and 6-OH differentially protected sugar synthon was described. A mannopyranosyl pentasaccharide and a glucopyranosyl hexasaccharide were prepared employing this new finding. PMID- 11101423 TI - Unexpected facile sequential halolactamization-hydroxylation of 2, 3-allenamides with CuX(2) for the efficient synthesis of 4-halo-5-hydroxypyrrol-2(5H)-ones. AB - [reaction: see text] 4-Halo-5-hydroxypyrrol-2(5H)-ones were synthesized from the efficient sequential halolactamization-hydroxylation reaction of 4 monosubstituted 2,3-allenamides with CuX(2) (X = Br, Cl) in high yields. Halolactamization of fully substituted 2,3-dienamide (1f) afforded 4-halo-pyrrol 2(5H)-ones. PMID- 11101424 TI - Solution and solid-state structure of (1-methoxy-8-naphthyllithium. THF)(2) AB - The structure of (1-methoxy-8-naphthyllithium.THF)(2), (2.THF)(2), determined by single-crystal X-ray diffraction (crystal data: monoclinic, a = 8.1816 (5), b = 21.9649 (14), and c = 8.2345 (3) A, beta = 117.969 (3) degrees; V = 1306.97 (12) A(3); space group P2(1)/n; Z = 4) reveals a centrosymmetrical dimer with a twist angle of about 63 degrees between the naphthyl rings and the Li-C(ipso)-Li plane, representing an intermediate between perpendicular and planar tetracoordinated C(ipso). NMR studies at low temperature indicate that 2 is dimeric in THF-d(8) solution under these conditions. PMID- 11101425 TI - Design, synthesis, and evaluation of a pyrrolinone-based matrix metalloprotease inhibitor. AB - [reaction: see text] A pyrrolinone-based hydroxamate matrix metalloprotease inhibitor, (-)-1, has been designed and synthesized. Enzymatic assay revealed that (-)-1 inhibited three of the ten matrix metalloprotease enzymes examined and as such represents a new, potentially important lead structure. PMID- 11101426 TI - An unusual seven-bond H-H spin coupling AB - A through-space, seven-bond (1)H-(1)H coupling has been observed in tiliacorinine (1) caused by the close spacial proximity of two aromatic protons that are located in different benzene rings which are separated by two sp(3) carbons. Results from DQF-COSY and NOESY NMR experiments, as well as molecular modeling, reveal that the two protons are unusually close to one another in space. PMID- 11101427 TI - Molecular assembly and disassembly: novel photolabile molecular hosts AB - A new approach to the assembly and photochemical disassembly of molecular hosts is developed. It is based on photoinduced fragmentation in hydroxyalkyl dithianes and utilizes a novel spiro-bis-dithiane as a photolabile molecular tether to link two formylated macromolecular blocks, e.g., formyl calixarenes or formyl dibenzocrown ethers. A key feature of this molecular system is that after an assembly-disassembly cycle the starting macromolecular blocks are recovered intact and can be used again. PMID- 11101428 TI - A novel synthesis of 4H-chromen-4-ones via intramolecular wittig reaction AB - The acylphosphoranes formed in a sequential manner from the reaction of the silyl ester of O-acyl(aroyl)salicylic acids and (trimethylsilyl)methylenetriphenylphosphorane undergo intramolecular Wittig cyclization on the ester carbonyl to afford the 4H-chromen-4-ones in good to excellent yields. PMID- 11101429 TI - Thermal cyclization of (2-Ethynylphenyl)triazenes: facile synthesis of substituted cinnolines and isoindazoles AB - High-temperature intramolecular cyclization of N, N-dialkyl-N'-(4-substituted-2 ethynylphenyl)triazenes provides under neutral conditions both 6-substituted cinnolines and 5-substituted isoindazoles in moderate to excellent yields. PMID- 11101430 TI - Meso aryl heptaphyrins: the first 30pi aromatic expanded porphyrins with an inverted structure. AB - [reaction: see text] Synthesis of new meso aryl 30pi heptaphyrins 2, 3, and 4 is achieved. Spectroscopic studies reveal that 2, 3, and 4 are aromatic and possess an inverted structure. PMID- 11101431 TI - Stereoselective synthesis of optically active alpha-hydroxy ketones and anti-1,2 diols via asymmetric transfer hydrogenation of unsymmetrically substituted 1,2 diketones. AB - [reaction: see text] A well-defined chiral Ru catalyst RuCl(N-(p-toluenesulfonyl) 1, 2-diphenylethylenediamine)(eta(6)-arene) effectively promotes asymmetric transfer hydrogenation of 1-aryl-1,2-propanedione with HCOOH/N(C(2)H(5))(3), leading preferentially to optically active 1-aryl-2-hydroxy-1-propanone with up to 99% ee and 89% yield at 10 degrees C. The reaction at 40 degrees C gives anti 1-aryl-1, 2-propanediol with up to 95% ee and 78% yield. This is a highly efficient procedure for the synthesis of optically active anti-diols. PMID- 11101432 TI - Synthesis of 6-acetamido-5-amino- and -5-guanidino-3, 4-dehydro-N-(2 ethylbutyryl)- 3-piperidinecarboxylic acids related to zanamivir and oseltamivir, inhibitors of influenza virus neuraminidases. AB - [reaction: see text] 6-Acetamido-5-amino- and -5-guanidino-3, 4-dehydro-N-(2 ethylbutyryl)-3-piperidinecarboxylic acids (8 and 9) have been synthesized starting from natural siastatin B, a bacterial neuraminidase inhibitor isolated from Streptomyces culture in a stereospecific fashion. These compounds are related to zanamivir and oseltamivir, inhibitors of influenza virus neuraminidases. PMID- 11101433 TI - Formation of beta-glucosamine and beta-mannose linkages using glycosyl phosphates. AB - [reaction: see text] Glycosyl phosphates were examined for their utility in the synthesis of challenging glycosidic linkages. beta-Glucosamine glycosides were formed preferentially and in good yield. beta-Mannosides were constructed in high overall yield with modest anomeric selectivity. Interesting solvent and conformational influences on the stereochemical outcome of the coupling reactions were observed. PMID- 11101434 TI - Reduced-symmetry deep-cavity cavitands AB - The syntheses of reduced-symmetry deep-cavity cavitands by two-stage stereoselective bridging with substituted benzal bromides is reported. Conditions for the optimal formation of the trisbridged derivatives were readily established. However, it was not possible to determine conditions which selectively promoted formation of either one of the two bisbridged species, or the monobridged compound, above the other products. A comparison of yields for A/B bisbridged derivatives verses A/C bisbridged derivatives may indicate that the one-pot formation of deep-cavity cavitands occurs primarily through the former species. PMID- 11101435 TI - Synthesis and structure of a new [6.6]metacyclophane with enediyne bridges. AB - [reaction: see text] Synthesis and structure of a novel [6.6]metacyclophane with enediyne bridges is reported. PMID- 11101436 TI - Palladium(0)-catalyzed cross-benzannulation between conjugated enynes. reactivity controlled synthesis of multifunctionalized benzenes AB - The synthesis of multifunctionalizbenzenes such as polysubstituted alkoxycarbonyl or cyanostyrenes was carried out by the regioselective cross-benzannulation between conjugated enynes in the presence of Pd(PPh(3))(4). PMID- 11101438 TI - Partial reduction of electron-deficient pyridines AB - The partial reduction of electron-deficient pyridines is described herein. Mono- and disubstituted pyridines can be transformed into functionalized dihydropyridines using either Birch reduction conditions or sodium/naphthalene in THF. The compounds formed by these high-yielding reductive alkylation protocols have potential as synthetic intermediates, and we have shown that bicyclo compounds containing 6,5, 6,6, and 6,7 ring systems can be prepared in one step via a base-promoted cylization. PMID- 11101437 TI - Vinylogous amide analogues of diaminopimelic acid (DAP) as inhibitors of enzymes involved in bacterial lysine biosynthesis. AB - [reaction: see text] Vinylogous amides 5 and 6 have been synthesized from L propargyl glycine and tested against diaminopimelate (DAP) enzymes involved in bacterial lysine biosynthesis. Both are reversible inhibitors of DAP D dehydrogenase and DAP epimerase with IC(50) values in the 500 microM range. Compound 5 shows competitive inhibition against the L-dihydrodipicolinate (DHDP) reductase with a K(i) value of 32 microM, which is comparable to the planar dipicolinate 16 (K(i) = 26 microM), the best known inhibitor of the enzyme. PMID- 11101440 TI - Facile lewis acid catalyzed synthesis of C(4) symmetric resorcinarenes AB - The Lewis acid catalyzed condensation of 3-methoxyphenol with octanal produced the C(4) symmetric calix[4]resorcinarene 2, in high yield. Of the numerous stereo and regioisomers possible, the rccc isomer with C(4) symmetry was the only product isolated (as a racemate). The structure has been established by single crystal X-ray structure analysis. PMID- 11101439 TI - Synthesis and biophysical characterization of tRNA(Lys,3) anticodon stem-loop RNAs containing the mcm(5)s(2)U nucleoside. AB - [reaction: see text] Phosphoramidite reagents of the naturally occurring modified nucleosides mcm(5)s(2)U and mcm(5)U were synthesized and along with pseudouridine were incorporated into 17-nucleotide lysine tRNA anticodon stem-loop domains. Standard RNA phosphoramidite coupling chemistry allowed us to systematically investigate the thermodynamic effects of nucleoside modification and to correlate thermodynamic trends with qualitative structure effects seen by NMR spectroscopy. PMID- 11101441 TI - A new one-Pot synthesis of alkynylphosphonates AB - A method for the palladium-catalyzed synthesis of alkynylphosphonates from 1,1 dibromo-1-alkenes has been developed. In general, the best catalyst system for this transformation was found to be Pd(OAc)(2), dppf, H-phosphonate, propylene oxide, DMF, 80 degrees C. The reaction appears tolerant of a range of functional groups in both the 1,1-dibromo-1-alkene and H-phosphonate coupling partners. The synthesis of a backbone-modified thymidine dimer is used to illustrate the application of this methodology in the synthesis of complex target molecules. PMID- 11101442 TI - Palladium-catalyzed additions of silyl-element bonds to bicyclopropylidene-A new access to bicyclopropyl and functionally substituted cyclopropylidenepropane derivatives AB - Palladium-catalyzed additions of disilanes, silylboranes, silylstannanes, and silyl cyanides across the double bond of bicyclopropylidene proceed with remarkable ease by two modes yielding either bicyclopropyl or cyclopropylidenepropane derivatives. PMID- 11101443 TI - One-pot oligosaccharide synthesis exploiting solvent reactivity effects. AB - [reaction: see text] One-pot syntheses of trisaccharides have been accomplished simply by changing the solvent system between the two subsequent glycosylation reactions and utilizing the difference in glycosylation rate between different solvents. By tuning the reactivity of acceptors and donors and performing the first glycosylation in Et(2)O (low glycosylation rate) and the second in CH(2)Cl(2)/Et(2)O (higher glycosylation rate), trisaccharides were synthesized in high yields (76-84%). PMID- 11101444 TI - Memory of chirality in diastereoselective alpha-alkylation of isoleucine and allo isoleucine derivatives. AB - [reaction: see text] alpha-Methylation of 3 gave 5 as a major product whereas 4 gave 6 predominantly, although both 3 and 4 have an (S)-chiral center at C(3). This indicates that chirality at C(2) in 3 and 4 was memorized in the corresponding intermediate enolates and the induced chirality made a major contribution in the stereochemical course of the reaction, while chirality at the adjacent chiral center C(3) had little effect. PMID- 11101445 TI - Intramolecular hydrogen-bond participation in phosphonylammonium salt formation. AB - [reaction: see text] A series of phosphonochloridates and phosphonyl dichlorides were prepared, and their reactivity with triethylamine has been investigated using (31)P NMR spectroscopy. Taken together these studies provide evidence that an intramolecular hydrogen-bond is required for phosphonylammonium salt formation to render the phosphorus more electron-deficient. PMID- 11101446 TI - First rational synthesis of the thiothiono analogue of an unsymmetrically substituted phthalic anhydride AB - Treatment of the dithiolane derivative of an alpha-carboxyethyl benzaldehyde with LDA at -78 degrees C smoothly produced the thiothionophthalic anhydride. The mechanism is proposed to involve loss of ethene and attack of an intermediate dithiocarboxylate onto the ester. Heating the thiothionophthalic anhydride gave the 3, 3'-bithiophthalide. PMID- 11101447 TI - Unique selectivity of iodohydroxylation reaction of allenyl phenyl sulfoxides in aqueous MeCN. A stereodefined synthesis of (E)-2-iodo-3-hydroxy-1-alkenyl sulfoxides AB - Iodohydroxylation reaction of allenyl phenyl sulfoxides with I(2) can smoothly proceed to generate (E)-2-iodo-3-hydroxy-1-alkenyl sulfoxides with excellent regio- and stereoselectivity in high or excellent yields. The configuration of E 2a was determined by the X-ray diffraction study. PMID- 11101448 TI - Syntheses with organoboranes. XI. Allylboration Of vinylic epoxides with allylic dialkylboranes AB - Allylboration of representative vinylic epoxides with allyldiethylborane (1) and (2-cyclohexenyl)dicyclohexylborane (2) affords the corresponding 1,2- and 1,4 addition products. cis-1, 2-Addition is favored in the reaction of 1 with 3, 4 epoxycycloalkenes of six- to eight-membered rings. 3, 4-Epoxycyclopentene (3a) and 5,5-dimethyl-3,4-epoxycyclopentene (3b) undergo five-membered ring opening during allylboration with 1 and 2, producing the corresponding (Z)-trienols (4a and 4b) with high stereoselectivity. 1,4-Addition of 1 and 2 to monoepoxides of 1, 3-butadiene and isoprene is favored, producing predominantly the corresponding (E)-alcohols. PMID- 11101449 TI - Sulfinimine-mediated asymmetric synthesis of 1,3-disubstituted tetrahydroisoquinolines: a stereoselective synthesis of cis- and trans-6,8 dimethoxy-1,3-dimethyl-1,2,3,4-tetrahydroisoquinoline. AB - [reaction: see text] The highly diastereoselective addition of lateral lithiated o-tolunitriles to sulfinimines followed by treatment of the resulting sulfinamide with MeLi, hydrolysis, and reduction represents a concise new methodology for the asymmetric synthesis of 1,3-disubstituted tetrahydroisoquinolines. PMID- 11101450 TI - Polymer-supported Bis(oxazoline)-copper complexes as catalysts in cyclopropanation reactions AB - Bis(oxazolines) are easily immobilized by functionalization of the central methylene bridge with polymerizable groups and subsequent polymerization. Polymers are transformed into copper catalysts active in the cyclopropanation of styrene with ethyl diazoacetate. The results are similar or even better than those obtained with the similar homogeneous systems, and the catalysts can be easily recovered and reused. The substitution in the methylene bridge leads to a slight reduction in the enantioselectivity and an unexpected cis-preference. PMID- 11101451 TI - Enantiocontrolled synthesis of highly functionalized tropanes via [5 + 2] cycloaddition to eta(3)-pyridinylmolybdenum pi-complexes. AB - [reaction: see text] A chiral, nonracemic eta(3)-pyridinyl scaffold participates in [5 + 2] cycloaddition with electron-deficient alkenes, an allene, and an alkyne to give eta(3)-allylmolybdenum bicyclic adducts. The adducts can be demetalated, providing a convergent route to highly functionalized tropanes. High enantiocontrol can be achieved throughout the cycloaddition and demetalation sequence. PMID- 11101452 TI - Studies directed toward the total synthesis of azaspiracid: stereoselective construction of C(1)-C(12), C(13)-C(19), and C(21)-C(25) fragments. AB - [reaction: see text] The efficient entry to the C(1)-C(12), C(13)-C(19), and C(21)-C(25) fragments of azaspiracid is outlined. The C(1)-C(12) portion is constructed using a key asymmetric allenyl borane addition to the corresponding alpha,beta-unsaturated aldehyde. The synthesis of the C(13)-C(19) portion utilizes an Evans asymmetric alkylation followed by Sharpless asymmetric dihydroxylation. In addition, a novel solution to the mismatched effects of a neighboring chiral oxazolidinone during a Sharpless dihydroxylation is detailed. PMID- 11101453 TI - Preparation of a resin-based chromium catalyst for effecting [6pi + 2pi] cycloaddition reactions. AB - [reaction: see text] A resin-based chromium catalyst for performing [6pi + 2pi] cycloaddition reactions has been prepared from chloromethylated polystyrene. The catalyst provides cycloadducts in yields comparable to the photochemical and thermal versions of these transformations, and the process is effective with a wide range of 6pi and 2pi reaction partners. PMID- 11101454 TI - B(C(6)F(5))(3)-Catalyzed hydrosilation of imines via silyliminium intermediates AB - A broad range of benzaldimines and ketimines can be hydrosilated efficiently, employing B(C(6)F(5))(3) as a catalyst in conjunction with PhMe(2)SiH. Spectral evidence supports the intermediacy of a silyliminium cation with a hydridoborate counterion formed via abstraction of a hydride from PhMe(2)SiH by B(C(6)F(5))(3) in the presence of imines. PMID- 11101455 TI - Concise enantiospecific synthesis of a coccinellied alkaloid, (-)-adalinine. AB - [reaction: see text] An enantiospecific synthesis of a coccinellied alkaloid, (-) adalinine, was established starting from (S)-(-)-pyroglutamic acid, where a stereoselective Michael addition and a samarium iodide-promoted regioselective carbon-nitrogen bond cleavage reaction were involved as the key reactions. PMID- 11101456 TI - Synthesis of D-hexos-5-uloses by novel in situ hydrolysis of epoxides derived from 6-deoxyhex-5-enopyranosides. AB - [reaction: see text] Epoxides derived from 2,3, 4-tri-O-protected-6-deoxyhex-5 enopyranosides are hydrolyzed in situ to ultimately give novel protected-D-hexos 5-ulose derivatives (sugar 1,5-dicarbonyls, 5-ketohexoses) in moderate to high yields. The products adopt a bicyclic structure (1,6-anhydropyranos-5-ulose) in solution with the pyranose ring in (4)C(1) conformation. The methodology has been used to prepare D-xylo-hexos-5-ulose (5-ketoglucose), a synthetic precursor to 1 deoxynojirimycin and a possible intermediate in the biosynthesis of inositols. PMID- 11101457 TI - Synthesis of biaryls via intermolecular radical addition of heteroaryl and aryl bromides onto arenes AB - Tris(trimethylsilyl)silane (TTMSS) and azobisisobutironitrile (AIBN) promoted the easy intermolecular arylation of aryl and heteroaryl bromides under thermal conditions via a radical pathway. PMID- 11101458 TI - Intramolecular diels-alder reaction by self-assembly of the components on a lewis acid template AB - Diels-Alder reactions of 2,4-hexadienol or its O-methyl ether with acrylate derivatives at 120 degrees C give mixtures of the four possible adducts with low selectivity. At ambient temperature and in the presence of Mg(II) or Al(III) Lewis acids, reactions of the dienol (but not the ether) are highly selective. Control experiments suggest that the Lewis acid serves both to tether the diene and dienophile and to induce an "intramolecular" reaction of the resulting "self assembled" intermediate. PMID- 11101459 TI - Efficient, diastereoselective chemical synthesis of a beta-mannopyranosyl phosphoisoprenoid. AB - [reaction: see text] Tetrabutylammonium benzyl dihydrophytylphosphate was coupled to S-phenyl 2,3-di-O-benyl-4, 6-O-benzylidene-1-thio-alpha-D-mannopyranoside S oxide on activation with triflic anhydride in toluene at -78 degrees C to give the corresponding beta-mannosyl phosphate in 56% yield with no detectable formation of the alpha-anomer. Treatment with sodium in liquid ammonia then afforded the unprotected beta-mannosyl phosphoisoprenoid. PMID- 11101460 TI - Novel cytotoxic, polyprenylated heptacyclic xanthonoids from Indonesian Garcinia gaudichaudii (Guttiferae). AB - [reaction: see text] The structures of novel gaudichaudiic acids F-I (1-4), isolated from the bark of Indonesian Garcinia gaudichaudii, have been elucidated by detailed spectral analysis. Gaudichaudiic acid I (4) is probably derived from 1 as a result of allylic oxidation at C-24 and C-21, followed by aromatization. PMID- 11101461 TI - Preparation of (S)-N-substituted 4-hydroxy-pyrrolidin-2-ones by regio- and stereoselective hydroxylation with Sphingomonas sp. HXN-200. AB - [reaction: see text] Enantiopure (S)-N-substituted 4-hydroxy-pyrrolidin-2-ones have been prepared for the first time by regio- and stereoselective hydroxylation of the corresponding pyrrolidin-2-ones by use of a biocatalyst. Hydroxylation of 6 and 8 with Sphingomonas sp. HXN-200 afforded 68% of (S)-7 in >99.9% ee and 46% of (S)-9 in 92% ee, respectively. Simple crystallization increased the ee of (S) 9 to 99. 9% in 82% yield. PMID- 11101462 TI - A crystallization-induced stereoselective glycosidation reaction in the synthesis of the anticancer drug etoposide PMID- 11101463 TI - Malnutrition in chronic obstructive pulmonary disease. PMID- 11101464 TI - Fat cat(aract) PMID- 11101465 TI - Defining the steps of the folate one-carbon shuffle and homocysteine metabolism. PMID- 11101466 TI - Cigarettes, cancer, and carotenoids: a continuing, unresolved antioxidant paradox. PMID- 11101467 TI - Bitter taste, phytonutrients, and the consumer: a review. AB - Dietary phytonutrients found in vegetables and fruit appear to lower the risk of cancer and cardiovascular disease. Studies on the mechanisms of chemoprotection have focused on the biological activity of plant-based phenols and polyphenols, flavonoids, isoflavones, terpenes, and glucosinolates. Enhancing the phytonutrient content of plant foods through selective breeding or genetic improvement is a potent dietary option for disease prevention. However, most, if not all, of these bioactive compounds are bitter, acrid, or astringent and therefore aversive to the consumer. Some have long been viewed as plant-based toxins. As a result, the food industry routinely removes these compounds from plant foods through selective breeding and a variety of debittering processes. This poses a dilemma for the designers of functional foods because increasing the content of bitter phytonutrients for health may be wholly incompatible with consumer acceptance. Studies on phytonutrients and health ought to take sensory factors and food preferences into account. PMID- 11101468 TI - Trends in waist-to-hip ratio and its determinants in adults in Finland from 1987 to 1997. AB - BACKGROUND: Although abdominal obesity has been shown to be an important risk factor for cardiovascular disease and a variety of other diseases, secular changes in fat distribution in populations have rarely been documented. OBJECTIVE: Our objective was to assess trends in waist-to-hip ratio (WHR) in the Finnish population during a 10-y period. In addition, we investigated the associations of WHR with body mass index (BMI), age, education, and lifestyle factors. DESIGN: Three independent cross-sectional surveys were carried out at 5 y intervals between 1987 and 1997. Altogether, 15096 randomly selected men and women aged 25-64 y participated in these surveys. RESULTS: The WHR increased in both men and women during the 10-y period (P: < 0.0001). In men, the strongest upward trend took place in the first 5-y period and then seemed to plateau; in women, the WHR continued to increase into the 1990s. In both sexes, the most prominent increase was observed in subjects aged >/=45 y. The WHR increased in all education-level groups, the lowest WHR being among those with the highest education. Age (18% in men, 12% in women) and BMI (33% in men, 25% in women) accounted for most of the variation in WHR, whereas only 3% was explained by education and lifestyle factors. CONCLUSIONS: Abdominal obesity is a growing problem in Finland, especially in persons aged >/=45 y. These adverse changes in body shape continued to take place, particularly in women, in the 1990s. PMID- 11101469 TI - Effects of acute (-)-hydroxycitrate supplementation on substrate metabolism at rest and during exercise in humans. AB - BACKGROUND: (-)-Hydroxycitrate (HCA), a competitive inhibitor of ATP-citrate lyase, should reduce the extramitochondrial acetyl-CoA pool. It has been hypothesized that HCA ingestion can reduce malonyl-CoA concentrations and consequently increase fatty acid oxidation in vivo. OBJECTIVE: This study investigated the acute effects of HCA supplementation on substrate utilization at rest and during exercise in endurance-trained humans. DESIGN: Ten cyclists [x+/- SD) age: 24 +/- 2 y, weight: 73 +/- 2 kg, maximal oxygen uptake: 4.95 +/- 0.11 L/min, maximal work output (W:max): 408 +/- 8 W] were studied at rest and during 2 h of exercise at 50% W:max on 2 occasions. Both 45 and 15 min before exercise and 30 and 60 min after the start of exercise, 3.1 mL/kg body wt of an HCA solution (19 g/L) or placebo was ingested. Total fat and carbohydrate oxidation rates were assessed. Blood samples were collected at 15-min intervals at rest and every 30 min during exercise. RESULTS: Plasma HCA concentrations increased after HCA ingestion up to 0.39 +/- 0.02 mmol/L (82.0 +/- 4.8 mg/L). However, no significant differences in total fat and carbohydrate oxidation rates were observed between trials. Accordingly, plasma glucose, glycerol, and fatty acid concentrations did not differ between trials. Plasma lactate concentrations were significantly lower in the HCA than in the placebo trial after 30 min of exercise but at the end of the exercise period they did not differ between trials. CONCLUSION: HCA, even when provided in large quantities, does not increase total fat oxidation in vivo in endurance-trained humans. PMID- 11101470 TI - Energy expenditure of nonexercise activity. AB - BACKGROUND: We found recently that changes in nonexercise activity thermogenesis (NEAT) mediate resistance to weight gain with overfeeding in sedentary adults. A potentially important, yet seldom investigated, component of NEAT is the energy expenditure of fidgeting-like activities. OBJECTIVE: Our goal was to measure changes in energy expenditure with fidgeting-like activities. DESIGN: Energy expenditure was measured in 24 subjects (17 women and 7 men x+/- SD body weight: 76 +/- 21 kg) while recumbent at rest, sitting motionless, standing motionless, partaking of self-selected fidgeting-like movements while seated and while standing, and walking on a treadmill at 1.6, 3.2, and 4.8 km/h (1, 2, and 3 mph). Measurements were performed by using a high-precision, indirect calorimeter connected to the subject via a transparent, lightweight facemask that enabled almost unrestricted movement. RESULTS: Compared with metabolic rate in the supine position (5.4 +/- 1.5 kJ/min), energy expenditure increased while sitting motionless by 4 +/- 6%, while fidgeting while seated by 54 +/- 29% (P: < 0.0001), while standing motionless by 13 +/- 8% (P: < 0.0001), while fidgeting while standing by 94 +/- 38% (P: < 0.0001), while walking at 1.6 km/h by 154 +/- 38% (P: < 0.0001), while walking at 3.2 km/h by 202 +/- 45% (P: < 0.0001), and while walking at 4.8 km/h by 292 +/- 81% (P: < 0.0001). There was a significant, positive correlation between changes in energy expenditure and body weight for fidgeting-like activities while standing (r = 0.43, P: = 0.02) but not while seated. CONCLUSIONS: There is marked variance between subjects in the energy expenditure associated with self-selected fidgeting-like activities. The thermogenic potential of fidgeting-like and low-grade activities is sufficiently great to substantively contribute to energy balance. PMID- 11101471 TI - Alterations in growth and body composition during puberty. IV. Energy intake estimated by the youth-adolescent food-frequency questionnaire: validation by the doubly labeled water method. AB - BACKGROUND: Estimates of energy intake are required for an understanding of growth and disease; however, few methods of energy intake in children have been validated. OBJECTIVE: Our objective was to validate energy intake estimated by the Youth-Adolescent Food-Frequency Questionnaire (YAQ) against the criterion total energy expenditure (TEE) by doubly labeled water (DLW). DESIGN: Twenty three boys and 27 girls (8.6-16.2 y of age) completed the YAQ and TEE measurements in 1 y. RESULTS: Energy intake by the YAQ (10. 03 +/- 3.12 MJ) and energy expenditure by DLW (9.84 +/- 1.79 MJ) were similar (P: = 0.91) with large lower (-6.30 MJ) and upper (6.67 MJ) +/-2 SD limits of agreement. When within subject CVs of repeated measures of the DLW and YAQ methods were used, 25 of the 50 subjects were deemed to have misreported their energy intake. The discrepancy in energy intake (YAQ - TEE) was related to body weight (r = -0.25, P: = 0.077) and percentage body fat (r = -0.24, P: = 0.09) but not to age (r = -0.07, P: = 0.63) or the time between measures. From logistic regression, fatter boys were more likely to underreport energy intake than were fatter girls. CONCLUSION: The YAQ provides an accurate estimation of mean energy intake for a group but not for an individual. PMID- 11101472 TI - Effect of diets high or low in unavailable and slowly digestible carbohydrates on the pattern of 24-h substrate oxidation and feelings of hunger in humans. AB - BACKGROUND: The pattern of substrate utilization with diets containing a high or a low proportion of unavailable and slowly digestible carbohydrates may constitute an important factor in the control, time course, and onset of hunger in humans. OBJECTIVE: We tested the hypothesis that isoenergetic diets differing only in their content of unavailable carbohydrates would result in different time courses of total, endogenous, and exogenous carbohydrate oxidation rates. DESIGN: Two diets with either a high (H diet) or a low (L diet) content of unavailable carbohydrates were fed to 14 healthy subjects studied during two 24-h periods in a metabolic chamber. Substrate utilization was assessed by whole-body indirect calorimetry. In a subgroup of 8 subjects, endogenous and exogenous carbohydrate oxidation were assessed by prelabeling the body glycogen stores with [(13)C]carbohydrate. Subjective feelings of hunger were estimated with use of visual analogue scales. RESULTS: Total energy expenditure and substrate oxidation did not differ significantly between the 2 diets. However, there was a significant effect of diet (P: = 0.03) on the carbohydrate oxidation pattern: the H diet elicited a lower and delayed rise of postprandial carbohydrate oxidation and was associated with lower hunger feelings than was the L diet. The differences in hunger scores between the 2 diets were significantly associated with the differences in the pattern of carbohydrate oxidation among diets (r = 0.67, P: = 0. 006). Exogenous and endogenous carbohydrate oxidation were not significantly influenced by diet. CONCLUSIONS: The pattern of carbohydrate utilization is involved in the modulation of hunger feelings. The greater suppression of hunger after the H diet than after the L diet may be helpful, at least over the short term, in individuals attempting to better control their food intake. PMID- 11101473 TI - Influence of methylenetetrahydrofolate reductase genotype, age, vitamin B-12, and folate status on plasma homocysteine in children. AB - BACKGROUND: Several studies have examined the association of the methylenetetrahydrofolate reductase (MTHFR) genotype with plasma homocysteine in adults, but few studies have been performed in children. OBJECTIVE: We measured the concentrations of plasma total homocysteine, folate, and vitamin B-12 in a group of healthy fasting children and related these to MTHFR genotype. DESIGN: After the subjects fasted, blood samples were collected into EDTA-containing tubes. Plasma, red blood cells, and the buffy coat were immediately stored at -80 degrees C for biochemical and molecular analyses. Plasma total homocysteine was determined by HPLC. Folate and vitamin B-12 were measured by a double-labeled radioimmunoassay, and the genotypic analysis was performed by polymerase chain reaction amplification of genomic DNA extracted from blood leukocytes. RESULTS: Plasma homocysteine concentrations correlated negatively with folate and vitamin B-12(,) but positively with age (P: < 0. 0001). Whereas folate and vitamin B-12 accounted for 27% and 19% of the variation in homocysteine, respectively, age accounted for 48% of the variation. When the cohort was divided into older (>10 y) and younger (10 y. CONCLUSION: Our data show that in a healthy pediatric population, MTHFR genotype played a significant role in determining homocysteine concentrations in older (>10 y), nutritionally stressed children. PMID- 11101474 TI - Chronic consumption of fresh but not heated yogurt improves breath-hydrogen status and short-chain fatty acid profiles: a controlled study in healthy men with or without lactose maldigestion. AB - BACKGROUND: Ingestion of fermented dairy products induces changes in the equilibrium and metabolism of the intestinal microflora and may thus have beneficial effects on the host. OBJECTIVE: We compared the effects of chronic consumption of yogurt with (fresh) or without (heated) live bacterial cultures (Lactobacillus bulgaricus and Streptococcus thermophilus) on plasma glucose, insulin, triacylglycerols, cholesterol, fatty acids, and short-chain fatty acids. DESIGN: Two groups of 12 healthy men with or without lactose malabsorption were selected with use of a breath-hydrogen test after a 30-g lactose load. Subjects were randomly assigned in a crossover design to 500 g/d of either fresh or heated yogurt for 2 periods of 15 d each, separated by a 15-d washout interval. RESULTS: Chronic consumption of fresh or heated yogurt had no detrimental effects on plasma glucose, insulin, or fatty acid areas under the curve in response to acute ingestion of 500 g yogurt in healthy men with or without lactose malabsorption. There were also no detectable changes in fasting plasma glucose, insulin, fatty acid, triacylglycerol, or cholesterol concentrations. In contrast, plasma butyrate was higher (P: < 0.03) and plasma propionate tended to be higher (P: = 0.059) in subjects without lactose malabsorption after fresh yogurt consumption than after heated yogurt consumption. There were no significant changes in plasma acetate. In subjects with lactose malabsorption, 15 d of fresh yogurt consumption also increased propionate production compared with values at baseline (P: < 0.04). In the same group, the production of breath hydrogen was lower after fresh yogurt consumption than after heated yogurt consumption (P: < 0.01). CONCLUSIONS: In men with lactose malabsorption, chronic consumption of yogurt containing live bacterial cultures ameliorated the malabsorption, as evidenced by lower breath hydrogen excretion, but increased propionate concentrations. In subjects without lactose malabsorption, such yogurt tended to increase propionate and increased butyrate. PMID- 11101475 TI - Factors contributing to alterations in skeletal muscle and plasma amino acid profiles in patients with chronic obstructive pulmonary disease. AB - BACKGROUND: There is increasing evidence of abnormal protein metabolism in patients with chronic obstructive pulmonary disease (COPD), as reflected by lower plasma branched-chain amino acid (BCAA) concentrations and different muscle amino acid (AA) patterns than in age-matched control subjects. OBJECTIVE: We examined whether the low plasma BCAA concentrations in COPD reflect an imbalance between anabolic and catabolic processes as evidenced by a low fat-free mass (FFM) and alterations in the anabolic hormone insulin and whether discrepancies in muscle AA concentrations between studies are related to different patient characteristics. DESIGN: AA profiles in arterial plasma and quadriceps femoris muscle and insulin concentrations in venous plasma were analyzed in 28 postabsorptive COPD patients (14 with and 14 without macroscopic emphysema) and in 28 control subjects. FFM was measured by dual-energy X-ray absorptiometry. RESULTS: The lower sum of plasma BCAAs in the COPD group than in the control subjects was the result of a lower leucine concentration (P: < 0.001); no significant difference in valine and isoleucine was found between the groups. In the COPD group, the lower leucine concentrations were associated with low FFM (P: < 0.01). Compared with the control group, the muscle-to-plasma leucine gradient was higher in the COPD group (P: < 0.001) and was associated with a higher insulin concentration (P: < 0.01). Several muscle AA concentrations were higher or tended to be higher in the group without emphysema than in the control group, whereas nearly all AA concentrations were lower in the group with emphysema. CONCLUSIONS: Leucine metabolism is altered in COPD patients and is associated with low FFM and high insulin concentrations. There were striking differences in the skeletal muscle AA profile between the COPD subtypes. PMID- 11101476 TI - Contribution of dietary protein to sulfide production in the large intestine: an in vitro and a controlled feeding study in humans. AB - BACKGROUND: Hydrogen sulfide is a luminally acting, bacterially derived cell poison that has been implicated in ulcerative colitis. Sulfide generation in the colon is probably driven by dietary components such as sulfur-containing amino acids (SAAs) and inorganic sulfur (eg, sulfite). OBJECTIVE: We assessed the contribution of SAAs from meat to sulfide production by intestinal bacteria with use of both a model culture system in vitro and an in vivo human feeding study. DESIGN: Five healthy men were housed in a metabolic suite and fed a sequence of 5 diets for 10 d each. Meat intake ranged from 0 g/d with a vegetarian diet to 600 g/d with a high-meat diet. Fecal sulfide and urinary sulfate were measured in samples collected on days 9 and 10 of each diet period. Additionally, 5 or 10 g bovine serum albumin or casein/L was added to batch cultures inoculated with feces from 4 healthy volunteers. Concentrations of sulfide, ammonia, and Lowry reactive substances were measured over 48 h. RESULTS: Mean (+/-SEM) fecal sulfide concentrations ranged from 0.22 +/- 0.02 mmol/kg with the 0-g/d diet to 3.38 +/- 0.31 mmol/kg with the 600-g/d diet and were significantly related to meat intake (P: < 0.001). Sulfide formation in fecal batch cultures supplemented with both bovine serum albumin and casein correlated with protein digestion, as measured by the disappearance of Lowry-reactive substances and the appearance of ammonia. CONCLUSION: Dietary protein from meat is an important substrate for sulfide generation by bacteria in the human large intestine. PMID- 11101477 TI - Relations of body fat distribution and height with cataract in men. AB - BACKGROUND: Cataract is the leading cause of blindness worldwide. Body mass index (BMI; in kg/m(2)) is a risk factor for cataract, but other anthropometric measurements may also be important. OBJECTIVE: We tested relations of alternative measures of body size, including height and waist-to-hip ratio (WHR), as well as BMI, with cataract. DESIGN: This was a prospective follow-up study. We analyzed data from 20271 participants in the Physicians' Health Study who did not have cataract at baseline and for whom there was complete information on weight, height, and other risk factors. For analyses concerning WHR, we excluded 3121 additional men for whom we did not have these measurements, assessed at the ninth year of follow-up. The main outcome measures were incident cataract and cataract surgery. RESULTS: Among the 17150 men for whom there were complete data, we confirmed an incident cataract in 1727 during an average of 14 y of follow-up. In proportional hazards regression models that adjusted for many known or suspected risk factors, higher BMI [rate ratio (RR) = 1.25 for >/=27.8 compared with <22, P: for trend = 0. 03], height (RR = 1.23 for >/=184 cm compared with /=65 y), received daily phylloquinone (1000 microg) or placebo for 2 wk. Serum undercarboxylated osteocalcin (ucOC) and total osteocalcin, N:-telopeptides of type I collagen (NTx), bone-specific alkaline phosphatase (BSAP), and phylloquinone concentrations were measured at baseline and after weeks 1 and 2. RESULTS: At baseline, the mean serum phylloquinone concentration was lower in the young than in the old group; there was no effect of sex. Concomitantly, baseline %ucOC was highest in the young and lowest in the old men (P: < 0.0001) but did not differ significantly by age in women. After supplementation, serum phylloquinone concentration increased approximately 10 fold (P: < 0.0001) at week 1 (from 0.93 +/- 0.08 to 8.86 +/- 0.70 nmol/L, x+/- SEM); this was sustained through week 2. Among all supplemented groups, mean %ucOC decreased from 7.6% to 3. 4% without significant differences by age or sex; 102 of 112 subjects had a >1% decrease. Phylloquinone supplementation reduced serum osteocalcin but did not alter NTx or BSAP concentration. CONCLUSIONS: Usual dietary practices in this population did not provide adequate vitamin K for maximal osteocalcin carboxylation. Phylloquinone supplementation reduced serum osteocalcin concentration but did not alter other markers of serum bone turnover. PMID- 11101482 TI - Vitamin D deficiency among older women with and without disability. AB - BACKGROUND: Vitamin D deficiency is associated with bone loss and bone fractures, and the identification of vulnerable populations is important to clinical practice and public health. OBJECTIVE: The objectives of this study were to determine the prevalence of vitamin D deficiency and to examine associated risk factors for vitamin D deficiency in older women. DESIGN: We measured serum concentrations of 25-hydroxyvitamin D [25(OH)D], 1,25-dihydroxyvitamin D [1, 25(OH)(2)D], intact parathyroid hormone (PTH), osteocalcin, and ionized calcium in women aged >/=65 y who were participating in the Women's Health and Aging Study I, an observational study of women representing the approximately one-third most disabled women living in the community, and women aged 70-80 y who were participating in the Women's Health and Aging Study II, an observational study of women among the two-thirds least disabled women living in the community in Baltimore. RESULTS: The women were classified into 4 domains of physical disability. Among 371 women with 0 or 1 domain of disability and 682 women with >/=2 domains of disability, 6.2% and 12.6%, respectively, had vitamin D deficiency [serum concentrations of 25(OH)D < 25 nmol/L]. In univariate analyses, risk factors for vitamin D deficiency included increasing age, black race, low educational level, high body mass index, high triceps skinfold thickness, increasing level of disability, winter season, and elevated creatinine concentration. In multivariate models, black race had a strong association with vitamin D deficiency when other risk factors were adjusted for. CONCLUSIONS: Vitamin D deficiency, a preventable disorder, is a common and important public health problem for older disabled women living in the community; black women are at higher risk than are white women. PMID- 11101483 TI - Primed, constant infusion with [2H3]serine allows in vivo kinetic measurement of serine turnover, homocysteine remethylation, and transsulfuration processes in human one-carbon metabolism. AB - BACKGROUND: One-carbon metabolism involves both mitochondrial and cytosolic forms of folate-dependent enzymes in mammalian cells, but few in vivo data exist to characterize the biochemical processes involved. OBJECTIVE: We conducted a stable isotopic investigation to determine the fates of exogenous serine and serine derived one-carbon units in homocysteine remethylation in hepatic and whole-body metabolism. DESIGN: A healthy man aged 23 y was administered [2,3,3 (2)H(3)]serine and [5,5,5-(2)H(3)]leucine by intravenous primed, constant infusion. Serial plasma samples were analyzed to determine the isotopic enrichment of free glycine, serine, leucine, methionine, and cystathionine. VLDL apolipoprotein B-100 served as an index of liver free amino acid labeling. RESULTS: [(2)H(1)]Methionine and [(2)H(2)]methionine were labeled through homocysteine remethylation. We propose that [(2)H(2)]methionine occurs by remethylation with [(2)H(2)]methyl groups (as 5-methyltetrahydrofolate) formed only from cytosolic processing of [(2)H(3)]serine, whereas [(2)H(1)]methionine is formed with labeled one-carbon units from mitochondrial oxidation of C-3 serine to [(2)H(1)]formate to yield cytosolic [(2)H(1)]methyl groups. The labeling pattern of cystathionine formed from homocysteine and labeled serine suggests that cystathionine is derived mainly from a serine pool different from that used in apolipoprotein B-100 synthesis. CONCLUSIONS: The appearance of both [(2)H(1)]- and [(2)H(2)]methionine forms indicates that both cytosolic and mitochondrial metabolism of exogenous serine generates carbon units in vivo for methyl group production and homocysteine remethylation. This study also showed the utility of serine infusion and indicated functional roles of cytosolic and mitochondrial compartments in one-carbon metabolism. PMID- 11101484 TI - Leptin and maternal growth during adolescent pregnancy. AB - BACKGROUND: Maternal growth on the basis of knee height occurs in nearly 50% of pregnant teenagers and is associated with greater gestational weight gain and accrual of subcutaneous fat in the mother but lower fetal growth compared with nongrowing teenagers and mature pregnant women. OBJECTIVE: The objective of this study was to determine whether leptin is a biomarker for continued maternal growth. DESIGN: Leptin concentrations were measured in 162 growing and nongrowing teenage gravidas (aged 5-fold, fetal growth restriction increased >6-fold, and infant birth weight decreased by approximately 200 g. Gravidas who developed pregnancy-induced hypertension showed a different pattern-higher leptin concentrations at entry and week 28, no difference in the leptin surge, and no postpartum difference in leptin concentration. CONCLUSION: A leptin surge by week 28 appears to mark reduced mobilization of maternal fat stores that is associated with maternal growth on the basis of knee height during adolescent pregnancy. PMID- 11101485 TI - Effects of parenteral cysteine and glutathione feeding in a baboon model of severe prematurity. AB - BACKGROUND: The availability of cysteine for glutathione synthesis is low in premature infants with respiratory distress. OBJECTIVE: The effects of gestational age, oxygen delivery, and cysteine infusion or glutathione infusion, or both, on plasma total cysteine and other methionine metabolites were studied in a baboon model of severe premature birth with respiratory distress. DESIGN: Premature baboons were studied as part of the multiinvestigator National Institutes of Health Collaborative Project on Bronchopulmonary Dysplasia. Premature baboons, 125 d (69% of term) or 140 d (78% of term) of gestational age, were maintained in neonatal intensive care units for or = 3 vaginal deliveries (OR = 5.2, 95% CI : 2.4 11.1) were associated with CIN II/III. History of infertility was also associated with CIN II/III (OR = 2.1, 95% CI : 1.0-4.2). CONCLUSIONS: The data suggest that history of infertility, IUD use and vaginal deliveries were associated with CIN among American Indian women. PMID- 11101539 TI - Nasopharyngeal carcinoma in Malaysian Chinese: occupational exposures to particles, formaldehyde and heat. AB - BACKGROUND: During 1990-1992, 282 Chinese residents of Selangor and the Federal Territory, Malaysia with histologically confirmed nasopharyngeal carcinoma (NPC) were interviewed about occupational history, diet, alcohol consumption, and tobacco use, as were an equal number of Malaysian Chinese population controls, pair-matched to cases by age and sex. METHODS: Exposures to 20 kinds of workplace substances, solar and industrial heat, and cigarette smoke, were analysed by univariate and multivariate methods. RESULTS: Nasopharyngeal carcinoma was associated with occupational exposures to construction, metal and wood dusts; motor fuel and oil; paints and varnishes; certain other chemicals; industrial heat; solar heat from outdoor occupations; certain smokes; cigarette smoking; and childhood exposure to parental smoking. After adjustment for risk from diet and cigarette smoke, only wood dust (OR = 2.36; 95% CI : 1.33- 4.19), and industrial heat (OR = 2.21; 95% CI : 1.12-4.33) remained clearly associated. Wood dust remained statistically significant after further adjustment for social class. No significant crude or adjusted association was found between NPC and formaldehyde (adjusted OR = 0.71; 95% CI : 0.34-1.43). CONCLUSIONS: This study supports previous findings that some occupational inhalants are risk factors for NPC. The statistical effect of wood dust remained substantial after adjustment for diet, cigarette smoke, and social class. Intense industrial heat emerged as a previously unreported risk factor, statistically significant even after adjustment for diet and cigarette smoke. No association was found between NPC and formaldehyde. PMID- 11101540 TI - Multiple primaries in pancreatic cancer patients: indicator of a genetic predisposition? AB - BACKGROUND: The genetic basis of several familial cancers including breast and colon cancers has been identified recently. The occurrence of multiple cancers in one individual is also suggestive of a genetic predisposition. To evaluate inherited predisposition in pancreatic cancer we compared the clinical data of pancreatic cancer patients with and without multiple primaries as well as the frequency of malignancies among their relatives. METHODS: Detailed data on 69 pancreatic cancer patients included survival time and TNM-classification. Index case data were separated into two groups. The first group (group 1) developed only pancreatic cancer during their lifetime, whereas the second group (group 2) developed additional primary tumours. A systematic family history was taken from 59 of these pancreatic cancer patients using a standardized questionnaire. The pancreatic cancers and the multiple primaries of the 59 patients were histologically proven. RESULTS: Of the 69 pancreatic cancer patients, 13 (18.8%) had multiple primaries. Neither the clinical data nor the survival data of the index cases revealed differences between the two groups (all nominal P-values >0.05). In the family history study blood relatives developed a malignancy in 51% (24 of 47) of the families in group 1 compared to 75% (9 of 12) in group 2. The risk of relatives in group 2 of developing a malignant tumour was significantly higher (P = 0.034) than in group 1 after adjustments for family size and age of disease onset of the index case. The cancer spectrum of the 59 families mainly included tumours of the digestive tract and the reproductive organs. CONCLUSIONS: A multiple primary cancer history is a common condition among pancreatic cancer patients. Relatives of these patients seem to have an increased risk for the development of distinct malignant solid tumours, which might be caused by an inherited predisposition. Clinical and genetic investigation of pancreatic cancer patients with multiple primaries and their families might lead to the identification of predisposing gene defects providing a new goal for the understanding of a shared genetic basis of different solid tumours. PMID- 11101541 TI - Occupation and pancreatic cancer in Spain: a case-control study based on job titles. PANKRAS II Study Group. AB - BACKGROUND: Occupational exposures may increase the risk of exocrine pancreatic cancer. This study aimed to identify occupations that in Spain may be associated with such risk. METHODS: Incident cases of pancreatic cancer and hospital controls were prospectively identified and interviewed during their hospital stay. Occupational history was obtained by direct interview with the patient and was available for 164 (89%) of 185 pancreatic cancer cases and for 238 (90%) of 264 controls. Occupations were coded according to the Spanish version of the International Standard Classification of Occupations 1988. RESULTS: A significant increased odds ratio (OR) was observed in men for 'physical, chemistry and engineering science technicians'. Elevated risks were also found for 'metal moulders, sheet-metal workers, structural metal workers, welders and related workers', 'painters and varnishers' and 'machinery mechanics and fitters'. 'Agricultural workers' did not present an increased risk for pancreas cancer in men. In women, however, high OR were observed for 'agricultural workers' and for 'textile and garment workers'. Most associations remained unchanged after considering long duration of the exposure and the period 5-15 years before diagnosis. CONCLUSIONS: Few occupations were at increased risk for pancreatic cancer, and the associations observed are in accordance with previous studies. The increases in risk observed for women in agricultural and textile jobs, and for men in the manufacture of dyes and pigments may deserve further attention. PMID- 11101542 TI - Occupational risk factors for renal cell carcinoma: agent-specific results from a case-control study in Germany. MURC Study Group. Multicenter urothelial and renal cancer study. AB - BACKGROUND: This case-control study was conducted to estimate the renal cell cancer (RCC) risk for exposure to occupation-related agents, besides other suspected risk factors. METHODS: In a population-based multicentre study, 935 incident RCC cases and 4298 controls matched for region, sex, and age were interviewed between 1991 and 1995 for their occupational history and lifestyle habits. Agent-specific exposure was expert-rated with two job-exposure matrices and a job task-exposure matrix. Conditional logistic regression was used to calculate smoking adjusted odds ratios (OR). RESULTS: Very long exposures in the chemical, rubber, and printing industries were associated with risk for RCC. Males considered as 'substantially exposed to organic solvents' showed a significant excess risk (OR = 1.6, 95% CI : 1.1-2.3). In females substantial exposure to solvents was also a significant risk factor (OR = 2.1, 95% CI : 1.0 4.4). Excess risks were shown for high exposure to cadmium (OR = 1.4, 95% CI : 1.1-1.8, in men, OR = 2.5, 95% CI : 1.2-5.3 in women), for substantial exposure to lead (OR = 1.5, 95% CI : 1.0-2.3, in men, OR = 2.6, 95% CI : 1.2-5.5, in women) and to solder fumes (OR = 1.5, 95% CI : 1.0-2.4, in men). In females, an excess risk for the task 'soldering, welding, milling' was found (OR = 3.0, 95% CI : 1.1-7.8). Exposure to paints, mineral oils, cutting fluids, benzene, polycyclic aromatic hydrocarbons, and asbestos showed an association with RCC development. CONCLUSIONS: Our results indicate that substantial exposure to metals and solvents may be nephrocarcinogenic. There is evidence for a gender specific susceptibility of the kidneys. PMID- 11101543 TI - Do larger people have more naevi? Naevus frequency versus naevus density. AB - BACKGROUND: It is unclear which of the number or the density of naevi on the skin is the more appropriate measure of risk of melanoma. Furthermore, the relationship between the number of naevi and their density in an individual has not been explored. Thus, for example, it is unknown if larger people tend to have more naevi by virtue of having a larger skin area, or if the density of naevi is similar in people of different body sizes. In this study, we explored the relationship between the number and the density of naevi in a sample of adolescents. SUBJECTS AND METHODS: A sample survey of naevi in 472 grade 9 secondary school students (aged 14-15 years) was conducted in Tasmania, Australia during 1992, and a subset of these individuals was followed up in 1997. Counts of naevi of various sizes were taken on the arm, leg, and back. Naevus density was estimated by using an algorithm to estimate body surface area from the height and weight of an individual. More general relationships of the naevus counts to height and weight were also explored. Finally, we considered whether the relationship between naevus density and the anthropometric variables could be confounded by exposure to ultraviolet radiation. RESULTS: The mean number of naevi was very similar in the two samples. Naevus density was slightly lower in the 1997 sample, mainly because of increasing body size in the cohort. The numbers of naevi were only weakly related to height and weight in males, and there was essentially no relationship in females. Regression analysis showed significant relationships of weight to the back naevus counts in males in 1992 and 1997, and to the arm naevus count in males in 1997; otherwise, none of the regression coefficients for height and weight were statistically significant. This picture did not change following adjustment for potentially confounding variables indicating time spent outdoors or in the sun. Furthermore, there was no evidence that time spent in the sun was related to the body mass index. CONCLUSIONS: It appears that the number and density of naevi in an individual are unrelated. Accordingly, with the present state of knowledge concerning the risk of melanoma, both the number and density of naevi should be considered as equally valid in future studies as markers of the risk of melanoma, and in studies on the natural history of naevi. If the disease mechanism is systemic, and not related to particular naevi, naevus density might form the better marker of risk. However, if the disease mechanism is related to effects on particular naevi, then the risk would vary in proportion to the number of naevi. PMID- 11101544 TI - Determinants of infant and early childhood mortality levels and their decline in the Netherlands in the late nineteenth century. AB - OBJECTIVE: To study the relative importance of various determinants of total and cause-specific infant and early childhood mortality rates and their decline in The Netherlands in the period 1875-1879 to 1895-1899. DATA AND METHODS: Mortality and population data were derived from Statistics Netherlands for 16 towns and 11 rural areas. Mortality levels and their decline were estimated with a Poisson regression model. The associations of the estimated levels and declines, and determinants of infant and early childhood mortality were analysed using multivariate linear regression analysis. The causes of death studied were major contributors to infant mortality (convulsions, acute digestive disease, acute respiratory disease) and early childhood mortality (encephalitis/meningitis, acute respiratory disease, measles). RESULTS: Infant mortality rates were high in the south-western part of The Netherlands in 1875-1879. Due to a rapid decline in the western regions, this pattern changed to a north-south gradient in 1895-1899. Early childhood mortality showed an urban-rural gradient in 1875-1879 with mortality high in towns. This gradient had largely disappeared by 1895-1899, due to a rapid decline in mortality in towns. Roman Catholicism was significantly associated with infant mortality (particularly from diarrhoeal disease) in 1875 1879 and 1895-1899. The association with Roman Catholicism was stronger in 1895 1899 because mortality declines were less rapid in Roman Catholic areas in 1875 1879 to 1895-1899. Urbanization was significantly associated with early childhood mortality (particularly from respiratory disease) in 1875-1879 and 1895-1899. This association weakened over time, due to the rapid decline in mortality in towns. CONCLUSIONS: Different determinants of mortality (decline) were important in infant and early childhood mortality and they acted on different causes of death. Therefore, infant and childhood mortality should be studied separately. International comparison of the results showed that findings with respect to determinants of mortality (decline) for one country do not necessarily apply to other countries. The results for The Netherlands with respect to infant mortality differed from England and Wales. PMID- 11101545 TI - How exclusive is exclusive breastfeeding? A comparison of data since birth with current status data. AB - BACKGROUND: There is no accepted and widely used indicator for exclusive breastfeeding since birth. Indeed, the difference between 'current status' data on exclusive breastfeeding and data on 'exclusive breastfeeding since birth' is rarely recognized. We used data from a longitudinal study to examine this issue. METHODS: A descriptive longitudinal, prospective study design was used in which 506 mother-infant pairs were included. The mothers completed daily recordings on infant feeding during the first nine months after birth. A research assistant conducted fortnightly home visits with structured interviews. The resulting data on breastfeeding patterns are presented in two different ways: analysis of 'current status' data based on a single 24-hour recording of infant feeding at 2, 4 and 6 months of age, and analysis of data 'since birth', i.e. data on infant feeding for every day, starting from birth until the ages of 2, 4 and 6 months. RESULTS: A wide discrepancy between the results obtained from the two analyses was found. The difference in the exclusive breastfeeding rate was over 40 percentage points at both 2 and 4 months of age (92% versus 51% at 2 months and 73% versus 30% at 4 months) and 9 percentage points at 6 months (11% versus 1.8%). CONCLUSIONS: Current status indicators based on a 24-hour period may be inadequate and even misleading for many purposes. We propose that in many studies an indicator called 'exclusive breastfeeding since birth' could be added. PMID- 11101546 TI - Arsenic in drinking water and the prevalence of respiratory effects in West Bengal, India. AB - BACKGROUND: A large population in West Bengal, India has been exposed to naturally occurring inorganic arsenic through their drinking water. A cross sectional survey involving 7683 participants of all ages was conducted in an arsenic-affected region between April 1995 and March 1996. The main focus of the study was skin keratoses and pigmentation alterations, two characteristic signs of ingested inorganic arsenic. Strong exposure-response gradients were found for these skin lesions. The study also collected limited information concerning respiratory system signs and symptoms, which we report here because increasing evidence suggests that arsenic ingestion also causes pulmonary effects. METHODS: Participants were clinically examined and interviewed, and the arsenic content in their current primary drinking water source was measured. There were few smokers and analyses were confined to non-smokers (N = 6864 participants). RESULTS: Among both males and females, the prevalence of cough, shortness of breath, and chest sounds (crepitations and/or rhonchi) in the lungs rose with increasing arsenic concentrations in drinking water. These respiratory effects were most pronounced in individuals with high arsenic water concentrations who also had skin lesions. Prevalence odds ratio (POR) estimates were markedly increased for participants with arsenic-induced skin lesions who also had high levels of arsenic in their current drinking water source (> or = 500 microg/l) compared with individuals who had normal skin and were exposed to low levels of arsenic (<50 microg/l). In participants with skin lesions, the age-adjusted POR estimates for cough were 7.8 for females (95% CI : 3.1-19.5) and 5.0 for males (95% CI : 2.6-9.9); for chest sounds POR for females was 9.6 (95% CI : 4.0-22.9) and for males 6.9 (95% CI : 3.1-15.0). The POR for shortness of breath in females was 23.2 (95% CI : 5.8 92.8) and in males 3.7 (95% CI : 1.3-10.6). CONCLUSION: These results add to evidence that long-term ingestion of inorganic arsenic can cause respiratory effects. PMID- 11101547 TI - Estimating deaths from industrial injury by capture-recapture: a cautionary tale. AB - BACKGROUND: Capture-recapture methods are increasingly being used to improve surveillance for a number of diseases. However, concerns persist regarding the validity of estimates obtained. METHOD: Capture-recapture methods were applied to estimate the ability of four separate data sources on occupational fatalities to predict the 237 deaths ('gold standard') we determined from a special in-depth study of medical examiner records. RESULTS AND CONCLUSION: Capture-recapture results based upon the four sources vary according to different models. However, both separately and in aggregate of industry type and cause of death, most models seriously underestimate the gold standard, and give a misleading impression of precision of the estimate of hidden individuals. It is commonly believed that reliable estimates from such methods require lists with high coverage and parsimonious models. Here, to obtain an estimate consistent with the gold standard, the list with almost complete coverage must be discarded and a complex model fitted. It is argued that this conclusion is of widespread application. PMID- 11101548 TI - Comparing odds ratios for nested subsets of dietary components. AB - BACKGROUND: In nutritional epidemiology, it is often of interest to disentangle the risk of disease associated with related foods or nutrients, where the food items are in a nested arrangement within a larger group. To compare odds ratios (OR) derived from a standard quantile-based analysis can be misleading since the amounts consumed may differ substantially for different dietary components. METHODS: The authors applied different logistic regression models on a case control study concerning the risk of colorectal adenomas due to meat and its different subsets such as white meat, red meat and well-done red meat. RESULTS: By calculating OR for a fixed amount of intake, the authors suggest a method for partitioning the risk of one dietary item into that associated with increasingly detailed sub-components. A graph is presented for illustrating such partitions in terms of both addition and substitution effects. CONCLUSIONS: Odds ratios based on upper versus lower quantiles or percentiles are useful as they compare the risk between the upper and lower ends of the consumption range. A complimentary set of OR are those based on fixed amounts of consumption. These allow for direct comparisons between nested subgroups of dietary components, in order to disentangle the risk linked to specific groups of foods or nutrients. PMID- 11101549 TI - Assessing research outcomes by postal questionnaire with telephone follow-up. TOTAL Study Group. Trial of Occupational Therapy and Leisure. AB - BACKGROUND: Face-to-face assessment of research outcomes is expensive and may introduce bias. Postal questionnaires offer a cheaper alternative which avoids observer bias, but non-response and incomplete response reduce the effective sample size and may be equally serious sources of bias. This study examines the extent and potential effects of missing data in the postal collection of outcomes for a large rehabilitation trial. METHODS: Questionnaires containing a number of established scales were posted to participants in a trial of occupational therapy after stroke. Response was maximized by telephone and postal reminders, and incomplete questionnaires were followed up by telephone. Scale scores obtained by imputing values to questionnaire items missing on return were compared with those achieved by telephone follow-up. FINDINGS: Response to the initial posting was 60%, rising to 85% after reminders. Participants receiving the experimental treatment were more likely to respond without a reminder. There were no significant differences on any known factors between eventual responders and non responders. Of the questionnaires, 43% were incomplete on return: partial responders were significantly different to complete responders on baseline disability and home circumstances. Of the incomplete questionnaires, 71% were resolved by telephone follow-up. In these, the scale scores achieved by telephone were generally higher than those derived by conventional imputation. CONCLUSION: Postal outcome assessment achieved a good response rate, but considerable effort was needed to minimize non-response and incomplete response, both of which could have been serious sources of bias. PMID- 11101550 TI - The effect of collapsing multinomial data when assessing agreement. AB - BACKGROUND: In epidemiological studies researchers often depend on proxies to obtain information when primary subjects are unavailable. However, relatively few studies have performed formal statistical inference to assess agreement among proxy informants and primary study subjects. In this paper, we consider inference procedures for studies of interobserver agreement characterized by two raters and three or more outcome categories. Of particular interest is the consequence of dichotomizing such data on the expected confidence interval width for the kappa coefficient. The effect of dichotomization on sample size requirements for testing hypotheses concerning kappa is also evaluated. METHODS: Simulation studies were used to compare coverage levels and widths for constructing confidence intervals. Sample size requirements were compared for multinomial and dichotomous data. We illustrate our results using a published data set on drinking habits that assesses agreement among primary and proxy respondents. RESULTS: Our results show that when multinomial data are treated as dichotomous, not only do the expected confidence interval widths become greater, but the penalty in terms of larger sample size requirements for hypothesis testing can be severe. CONCLUSION: We conclude that there are clear advantages in preserving multinomial data on the original scale rather than collapsing the data into a binary trait. PMID- 11101551 TI - Human T-cell lymphotropic virus testing of blood donors in Norway: a cost-effect model. AB - BACKGROUND: Human T-cell lymphotropic virus type I and II (HTLV-I and II) are human retroviruses that can be transmitted by transfusion of whole blood. An HTLV I infection is associated with adult T-cell leukaemia (ATL) and with tropical spastic paraparesis (TSP). Antibody tests from 5.5 million European blood donors have shown that the HTLV prevalence is low, ranging from 0 to 0.02%. This paper examines costs and effects associated with the intervention of testing all new blood donors for HTLV. METHODS: A mathematical model was used to calculate the number of cases prevented by the intervention. For a given prevalence of HTLV in the blood donor population, the model calculates the number of recipients infected by transfusion, and the number of partners and offspring that will in turn be infected. The model then calculates the number of subjects with disease due to HTLV-I infection and the number of deaths from disease. From these numbers the measures of cost and effect are calculated. RESULTS: Testing all new blood donors for HTLV is calculated to cost US$ 9.2 million per life saved, or US$ 420,000 per quality adjusted life year gained by the intervention, when the HTLV prevalence among donors is 1 per 100,000. When the prevalence among donors is 10 per 100,000 the intervention will cost US$ 0.9 million per life saved, or US$ 41,000 per quality adjusted life year gained. The same analysis shows that testing blood donors for human immunodeficiency virus (HIV) saves money when the HIV prevalence among donors is above 0.7 per 100,000. CONCLUSION: For Norway, studies suggest a willingness to pay to save a statistical life of approximately US$ 1.2 million. The costs fall under this value when the number of infected persons is > or = 8 per 100,000 donors. The results are uncertain because of the uncertainty in HTLV infection and disease parameters. PMID- 11101552 TI - BCG vaccine effectiveness in preventing tuberculosis and its interaction with human immunodeficiency virus infection. AB - BACKGROUND: To explore Bacillus Calmette-Guerin vaccine (BCG) as a protective factor against tuberculosis (TB) and how human immunodeficiency virus (HIV) infection modifies the effect of BCG on TB. METHODS: Two matched case-control studies were conducted. One study compared TB cases and controls who were HIV positive. The second compared TB cases and controls who were HIV negative. The study population consisted of 88 TB cases and 88 controls among HIV-positive individuals and 314 TB cases and 310 controls among HIV-negative individuals. Cases were new TB diagnoses, confirmed by either bacteriology, pathology, radiology or clinical response to treatment; controls were selected from people without TB symptoms and who sought medical attention in the same institution where a case was enrolled. BCG was assessed by the presence of a typical scar. RESULTS: The level of protection against all clinical forms of TB was 22% among HIV positive individuals (odds ratio [OR] = 0.78, 95% CI : 0.48-1.26) and 26% among HIV negatives (OR = 0.74, 95% CI : 0.52-1.05). There was a significant difference (P = 0.002) in the level of protection against extrapulmonary TB (ETB) between HIV-negative (OR = 0.54, 95% CI : 0.32-0.93) and HIV-positive individuals (OR = 1.36, 95% CI : 0.72-2.57). CONCLUSION: BCG has a modest protective effect against all forms of TB independent of HIV status, and BCG confers protection against extrapulmonary TB among HIV-negative individuals. However, HIV infection seems to abrogate the protective effect of BCG against extrapulmonary TB. Our data support the public health importance of BCG vaccine in the prevention of extrapulmonary TB among immunocompetent individuals. PMID- 11101554 TI - An introduction To instrumental variables for epidemiologists PMID- 11101553 TI - Outbreak of Legionnaires' disease associated with a display whirlpool spa. AB - BACKGROUND: Recognized outbreaks of Legionnaires' disease (LD) are rare; when they occur, they provide opportunities to understand the epidemiology of the illness and improve prevention strategies. We investigated a population-based outbreak. METHODS: After the confirmation of LD in October 1996 in five people in neighbouring towns in southwest Virginia, active surveillance for additional cases was undertaken. A case-control study was conducted to identify exposures associated with illness, followed by a cohort study among employees of the facility at which the source of the outbreak was located in order to assess unrecognized exposure and illness. Samples of likely sources of LD in the facility were cultured for LEGIONELLA: RESULTS: In all, 23 laboratory-confirmed cases of LD were eventually identified. Of the 15 cases in the case-control study, 14 (93%) reported visiting a home-improvement store, compared with 12 (27%) of 45 controls (matched odds ratio [MOR] = 23.3; 95% CI : 3-182). Among home-improvement centre patrons, 10 (77%) of 13 cases questioned recalled either visiting or walking by a display whirlpool spa, compared with 3 (25%) of 12 controls (MOR = 5.5; 95% CI : 0.7-256.0). Two cases' sputum isolates were an exact match, by monoclonal antibody subtyping and arbitrarily primed polymerase chain reaction, to a whirlpool spa filter isolate from the store. Employees reporting more exposure to the display spas were more likely to report symptoms of LD or to have an elevated titre. CONCLUSIONS: This investigation shows that LD can be transmitted from a whirlpool spa used for display only, and highlights the need for minimizing the risk of transmission of LD from all water-filled spas. Key messages This paper describes an investigation of a population-based outbreak of Legionnaires' disease (LD). A case-control study first identified a home improvement store as the likely source of the outbreak. An environmental investigation later confirmed that finding, as two cases' sputum isolates were an exact match, by monoclonal antibody subtyping and arbitrarily primed polymerase chain reaction, to a whirlpool spa filter isolate from the store. The spa was intended and used for display only. PMID- 11101555 TI - Identification and detection of Stenotrophomonas maltophilia by rRNA-directed PCR. AB - Stenotrophomonas maltophilia has recently emerged as an important nosocomial pathogen in immunocompromised patients, in transplant recipients, and in persons with cystic fibrosis (CF). While this organism is nonpathogenic in healthy individuals, it is increasingly associated with morbidity and mortality in susceptible populations. Recent studies have indicated that for approximately 10% of CF patients with moderate lung disease, S. maltophilia can be cultured from respiratory tract secretions. Identification of S. maltophilia can be problematic, and analysis of isolates from the Burkholderia cepacia Research Laboratory and Repository showed that several isolates presumptively identified as B. cepacia by clinical microbiology laboratories were in fact S. maltophilia. To overcome the problems associated with definitive identification, we developed species-specific PCR (SS-PCR) primers, designated SM1 and SM4, directed to the 23S rRNA gene, and tested their utility to accurately identify S. maltophilia directly from sputum. The SS-PCR was developed and tested against a panel of 112 S. maltophilia isolates collected from diverse geographic locations. To test for specificity, 43 isolates from 17 different species were analyzed. PCR with the SM1-SM4 primer pair and isolated genomic DNA as a template resulted in amplification of a band from all S. maltophilia isolates and was uniformly negative for all other species tested, yielding a sensitivity and a specificity of 100% for the SS-PCR. The utility of the SS-PCR to directly identify S. maltophilia in sputum was examined. Thirteen expectorated sputum samples from CF patients were analyzed by SS-PCR. Three samples were PCR positive, in complete concordance with the conventional laboratory culture. Thus, we have developed an SS-PCR protocol that can rapidly and accurately identify S. maltophilia isolates and which can be used for the direct detection of this organism in CF patient sputum. PMID- 11101556 TI - Application of representational difference analysis to genomic fragments of Marek's disease virus. AB - A rapid and simple method for isolation of DNA fragments of Marek's disease virus (MDV) based on representational difference analysis (RDA) was developed. Multiple viral DNA fragments, the sizes of which were restricted to 0.3 to 3.5 kbp, were simultaneously amplified after subtraction of chicken DNA from BamHI-, BglII-, EcoRI-, HindIII-, or XhoI-digested DNA fragments of MDV-infected cells. Nucleotide sequence of two RDA-derived fragments coincided with the sequence determined from direct sequencing of the MDV genome. We detected an interstrain difference in the size of restriction enzyme-digested fragments on agarose gel. This method was used on a single feather pulp to generate sufficient MDV DNA for cloning. PMID- 11101557 TI - Rapid genotyping of varicella-zoster virus vaccine and wild-type strains with fluorophore-labeled hybridization probes. AB - We developed a single-tube rapid method for the detection and differentiation of varicella-zoster virus (VZV) vaccine and wild-type strains that combines rapid cycle PCR with wild-type-specific fluorescent probe melting profiles for product genotyping. A region including the polymorphic site in VZV open reading frame (ORF) 62 was amplified in the presence of two fluorescence-labeled hybridization probes. During the annealing step of the thermal cycling, both probes bound to their complementary sequences in the amplicon, resulting in resonance energy transfer, thus providing real-time fluorescence monitoring of PCR. Continuous acquisition of fluorescence data during a melting curve analysis at the completion of PCR revealed that loss of fluorescence occurred in a strain specific manner as the detection probe, which was fully complementary to the wild type VZV ORF 62 region, melted off the template. Use of this method allowed genotyping of samples within minutes after the completion of PCR, eliminating the need for post-PCR sample manipulation. In addition to reducing the time required to produce a result, this method substantially reduces the risk of contamination of the final product as well as the risk of sample tracking errors. The genotypes of 79 VZV-positive samples determined by this fluorescent resonance energy transfer (FRET) method were identical to the genotypes obtained by conventional PCR and restriction fragment length polymorphism analysis. The genotyping of VZV strains by the FRET method is a rapid and reliable method that is suitable for typing and that is also practical for use for the processing of large numbers of specimens. PMID- 11101558 TI - Dissemination of CTX-M-3 and CMY-2 beta-lactamases among clinical isolates of Escherichia coli in southern Taiwan. AB - A total of 1,210 clinical isolates of Escherichia coli collected from a university hospital in southern Taiwan were screened for production of extended spectrum beta-lactamases (ESBLs). Expression of classical ESBLs (resistant to extended-spectrum beta-lactam agents and susceptible to beta-lactam inhibitors) was inferred in 18 isolates by the phenotypic confirmatory test. These included 10 isolates producing CTX-M-3, 2 strains carrying SHV-12, 1 strain harboring SHV 5, 1 strain expressing TEM-10, and 4 strains producing unidentifiable ESBLs with a pI of 8.05, 8.0, or 7.4. Eighteen isolates that showed decreased susceptibilities to ceftazidime and/or cefotaxime, negative results for the confirmatory test, and high-level resistance to cefoxitin (MICs of >/=128 microg/ml) were also investigated. Five isolates were found to produce CMY-2 AmpC enzymes, one isolate carried both CTX-M-3 and CMY-2, and the remaining three and nine isolates expressed putative AmpC beta-lactamases with pIs of >9.0 and 8.9, respectively. Thus, together with the isolate producing CTX-M-3 and CMY-2, 19 (1.6%) isolates produced classical ESBLs. Pulsed-field gel electrophoresis revealed that all isolates carrying CTX-M-3 and/or CMY-2 were genetically unrelated, indicating that dissemination of resistance plasmids was responsible for the spread of these two enzymes among E. coli in this area. Among the 16 isolates expressing CTX-M-3 and/or CMY-2, 5 might have colonized outside the hospital environment. Our data indicate that CTX-M-3 and CMY-2, two beta lactamases initially identified in Europe, have been disseminated to and are prevalent in Taiwan. PMID- 11101559 TI - Use of whole blood specimens for routine clinical quantitation of hepatitis C virus RNA does not increase assay sensitivity. AB - The measurement of hepatitis C virus (HCV) RNA levels in the blood has, in the last few years, become a critical component in the therapy of patients with HCV infections. Initially, extraction methods for serum and plasma were used, but a newer method that uses Catrimox-14 as the extraction agent for whole blood has been reported. Because the whole blood extraction method may yield higher virus levels if significant levels of virus are present in the white blood cells (WBC), the method was evaluated for use in our clinical diagnostic laboratory despite its higher reagent costs and more time-consuming methodology. RNA was simultaneously extracted from 39 clinical samples by four different methods: Catrimox-14-Trizol extraction from whole blood, Trizol extraction from whole blood, Trizol extraction from serum, and a commercial serum extraction method, the EZNA total RNA kit. In addition, in an effort to quantitate the amount of HCV RNA virus in the WBC, Trizol extraction from isolated WBC was also performed. Quantitative results for samples from which RNA was extracted by all four methods were essentially the same; the Catrimox-14-Trizol method did not yield increased virus levels. Insignificant levels of virus were found in the WBC. The results did not demonstrate a clinical usefulness for the Catrimox-14-Trizol method. PMID- 11101560 TI - Randomly amplified polymorphic DNA analysis of Erysipelothrix spp. AB - The usefulness of randomly amplified polymorphic DNA method (RAPD) to identify each species of genus Erysipelothrix and for epidemiological analysis of this genus was studied. Eighty-one strains and 18 random primers were tested. Among the tested primers, the primers NK51 (GGTGGTGGTATC) and NK6 (CCCGCGCCCC) produced noticeable results. The primer NK51 revealed four species-specific RAPD patterns. Of the 66 strains of E. rhusiopathiae, 64 had the same unique band of 884 bp. Of the 12 strains of E. tonsillarum, 11 produced a 1,265-bp band. In addition, two strains, previously thought to be E. rhusiopathiae, produced the 1,265-bp band, suggesting that they had been misclassified. One strain of E. tonsillarum produced the 884-bp band, suggesting that it too was E. rhusiopathiae. The E. rhusiopathiae strain of serovar 13 produced a 650-bp band, and the strain of serovar 18 produced a clear 420-bp band as well as three weak bands of 1,265, 918, and 444 bp. The primer NK6 revealed 14 RAPD patterns that were not serovar specific. However, different patterns were produced among strains of the same serovar showing that the RAPD method is able to identify the genetic variations of strains of this genus and can rapidly and easily differentiate strains of the same serovar. Based on these results, we concluded that the RAPD method with primers NK51 and NK6 is a rapid and reliable method to identify the species of this genus; we also concluded that this method might be a useful tool for the epidemiological analysis of the Erysipelothrix species. PMID- 11101561 TI - Improved genotyping vaccine and wild-type poliovirus strains by restriction fragment length polymorphism analysis: clinical diagnostic implications. AB - The combination of preventive vaccination and diagnostic typing of viral isolates from patients with clinical poliomyelitis constitutes our main protective shield against polioviruses. The restriction fragment length polymorphism (RFLP) adaptation of the reverse transcriptase (RT)-PCR methodology has advanced diagnostic genotyping of polioviruses, although further improvements are definitely needed. We report here on an improved RFLP procedure for the genotyping of polioviruses. A highly conserved segment within the 5' noncoding region of polioviruses was selected for RT-PCR amplification by the UC(53)-UG(52) primer pair with the hope that it would be most resistant to the inescapable genetic alteration-drift experienced by the other segments of the viral genome. Complete inter- and intratypic genotyping of polioviruses by the present RFLP method was accomplished with a minimum set of four restriction endonucleases (HaeIII, DdeI, NcoI, and AvaI). To compensate for potential genetic drift within the recognition sites of HaeIII, DdeI, or NcoI in atypical clinical samples, the RFLP patterns generated with HpaII and StyI as replacements were analyzed. The specificity of the method was also successfully assessed by RFLP analysis of 55 reference nonpoliovirus enterovirus controls. The concerted implementation of these conditional protocols for diagnostic inter- and intratypic genotyping of polioviruses was evaluated with 21 clinical samples with absolute success. PMID- 11101562 TI - Citrobacter rodentium, the causative agent of transmissible murine colonic hyperplasia, exhibits clonality: synonymy of C. rodentium and mouse-pathogenic Escherichia coli. AB - Citrobacter rodentium (formerly Citrobacter freundii biotype 4280 and Citrobacter genomospecies 9) was described on the basis of biochemical characterization and DNA-DNA hybridization data and is the only Citrobacter species known to possess virulence factors homologous to those of the human pathogens enteropathogenic Escherichia coli and enterohemorrhagic E. coli. These virulence factors are encoded on the locus of enterocyte effacement (LEE), a pathogenicity island required for the characteristic attaching and effacing (AE) pathology seen in infection with these three pathogens. C. rodentium, which apparently infects only mice, provides a useful animal model for studying the molecular basis of AE pathology. No work has been done to assess differences in pathogenicity between C. rodentium isolates from diverse sources. Here, we report the examination of 15 C. rodentium isolates using a battery of genetic and biochemical approaches. No differences were observed between the isolates by repetitive-element sequence based PCR analysis, biochemical analysis, and possession of LEE-specific virulence factors. These data suggest that members of the species are clonal. We further characterized an atypical E. coli strain from Japan called mouse pathogenic E. coli (MPEC) that, in our hands, caused the same disease as C. rodentium. Applying the same battery of tests, we found that MPEC possesses LEE encoded virulence factors and is indistinguishable from the previously characterized C. rodentium isolate DBS100. These results demonstrate that MPEC is a misclassified C. rodentium isolate and that members of this species are clonal and represent the only known attaching and effacing bacterial pathogen of mice. PMID- 11101563 TI - Glyceraldehyde-3-phosphate dehydrogenase-encoding gene as a useful taxonomic tool for Staphylococcus spp. AB - The gap gene of Staphylococcus aureus, encoding glyceraldehyde-3-phosphate dehydrogenase, was used as a target to amplify a 933-bp DNA fragment by PCR with a pair of primers 26 and 25 nucleotides in length. PCR products, detected by agarose gel electrophoresis, were also amplified from 12 Staphylococcus spp. analyzed previously. Hybridization with an internal 279-bp DNA fragment probe was positive in all PCR-positive samples. No PCR products were amplified when other gram-positive and gram-negative bacterial genera were analyzed using the same pair of primers. AluI digestion of PCR-generated products gave 12 different restriction fragment length polymorphism (RFLP) patterns, one for each species analyzed. However, we could detect two intraspecies RFLP patterns in Staphylococcus epidermidis, Staphylococcus hominis, and Staphylococcus simulans which were different from the other species. An identical RFLP pattern was observed for 112 S. aureus isolates from humans, cows, and sheep. The sensitivity of the PCR assays was very high, with a detection limit for S. aureus cells of 20 CFU when cells were suspended in saline. PCR amplification of the gap gene has the potential for rapid identification of at least 12 species belonging to the genus Staphylococcus, as it is highly specific. PMID- 11101564 TI - Quantitative analysis of cytomegalovirus (CMV) viremia using the pp65 antigenemia assay and the COBAS AMPLICOR CMV MONITOR PCR test after blood and marrow allogeneic transplantation. AB - The performance of a commercially available qualitative PCR test for plasma (AMPLICOR CMV Test; Roche Diagnostics) and a quantitative PCR test for plasma and leukocytes (COBAS AMPLICOR CMV MONITOR Test; Roche Diagnostics) was evaluated with samples from 50 blood or marrow allogeneic transplant recipients who received short courses of sequential ganciclovir therapy (2 weeks intravenously followed by 2 weeks orally) based on a positive cytomegalovirus (CMV) pp65 antigenemia (AG) assay. The number of persons with a positive CMV test was significantly higher for leukocyte-based assays (AG, 67.5%; PCR, 62.5%) compared to both quantitative and qualitative PCR tests of plasma (42.5 and 35%, respectively). One person developed CMV disease during the study despite a negative AG assay; in this particular case, all PCR assays were found to be positive 10 days before his death. There was a trend for earlier positivity after transplantation and more rapid negativity after initiation of ganciclovir for the tests performed on leukocytes. The mean number of CMV copies as assessed by PCR was significantly higher in leukocytes than in plasma (P = 0.02). Overall, excellent agreement (kappa coefficient, >0.75) was found only between the two PCR assays (qualitative and quantitative) based on plasma. These results suggest that either the pp65 AG assay or the COBAS AMPLICOR CMV MONITOR Test using leukocytes could be used to safely monitor CMV viremia in related allogeneic blood or marrow transplant recipients. Such a strategy will result in preemptive treatment for about two-thirds of the persons with a relatively low rate (<33%) of secondary viremic episodes following short courses of ganciclovir therapy. PMID- 11101565 TI - Molecular epidemiology of penicillin-nonsusceptible Streptococcus pneumoniae among children in Greece. AB - A total of 145 penicillin-nonsusceptible Streptococcus pneumoniae strains were isolated from young carriers in Greece and analyzed by antibiotic susceptibility testing, serotyping, restriction fragment end labeling (RFEL), and penicillin binding protein (PBP) genotyping. The serotypes 23A and 23F (54%), 19A and 19F (25%), 9V (5%), 15A, 15B, and 15C (4%), 6A and 6B (4%), and 21 (4%) were most prevalent in this collection. Fifty-three distinct RFEL types were identified. Sixteen different RFEL clusters, harboring 2 to 32 strains each, accounted for 82% of all strains. Eight of these genetic clusters representing 60% of the strains were previously identified in other countries. A predominant lineage of 66 strains (46%) harboring five RFEL types and the serotypes 19F and 23F was closely related to the pandemic clone Spain(23F)-1 (genetic relatedness of > or =85%). Another lineage, representing 11 strains, showed close genetic relatedness to the pandemic clone France(9V)-3. Another lineage of 8 serotype 21 strains was Greece specific since the RFEL types were not observed in an international collection of 193 genotypes from 16 different countries. Characterization of the PBP genes pbp1a, pbp2b, and pbp2x revealed 20 distinct PBP genotypes of which PBP type 1-1-1, initially observed in the pandemic clones 23F and 9V, was predominantly present in 11 RFEL types in this Greek collection of penicillin nonsusceptible strains (55%). Sixteen PBP types covering 52 strains (36%) were Greece specific. This study underlines the strong contribution of penicillin resistant international clones to the prevalence and spread of penicillin nonsusceptible pneumococci among young children in Greece. PMID- 11101566 TI - Clonal groups of penicillin-nonsusceptible Streptococcus pneumoniae in Baltimore, Maryland: a population-based, molecular epidemiologic study. AB - Few data are available on the molecular subtypes of all penicillin-nonsusceptible Streptococcus pneumoniae (PNSP) from a defined population base. Pulsed-field gel electrophoresis (PFGE), serotyping, and antibiotic susceptibility testing were performed for all available invasive PNSP isolates for which the penicillin (MIC) was > or =0.1 microg/ml from Baltimore, Md., during 1995-1996 (n = 143). The dendrogram analysis of PFGE patterns included 32 distinct clonal groups. Six major clonal groups included two-thirds of the PNSP strains. Major clonal groups 2, 3, 4, and 6 strains were genetically related to four previously described international clones and were all multidrug resistant. Major clonal group 3 was genetically related to the Tennessee(23F)-4 clone and contained all four strains for which the penicillin MIC was 8 microg/ml. Most of the clonal group 1 and 5 strains had intermediate susceptibility to penicillin and were rarely multidrug resistant. The latter clonal groups represent two previously undescribed penicillin-intermediate pneumococcal clones. Clonal group homogeneity was greater for serotype 9V, 19A, and 23F strains than for serotype 6A, 6B, 14, and 19F strains. The classification of PNSP strains into clonal groups is essential for future population-based epidemiologic studies of PNSP. PMID- 11101567 TI - Results of multiple diagnostic tests for Mycobacterium avium subsp. paratuberculosis in patients with inflammatory bowel disease and in controls. AB - Mycobacterium avium subsp. paratuberculosis has been incriminated as a cause of Crohn's disease (CD); however, studies to date have been relatively small and generally only used a single diagnostic assay. The objective of the study was to reexamine the association of M. avium subsp. paratuberculosis and CD using multiple diagnostic tests. Five methods were used to detect M. avium subsp. paratuberculosis infections in 439 inflammatory bowel disease (IBD) patients and 324 control subjects in the United States and Denmark. Most assays were adaptations of diagnostic tests for this infection performed routinely on animals. PCR for IS900, a genetic element unique to M. avium subsp. paratuberculosis, was positive significantly more often on resected bowel and lymph node tissues from CD patients (19.0%) and ulcerative colitis (UC) patients (26.2%) than from controls (6. 3%) (P < 0.05). Positive IS900 PCR results occurred more often in U. S. than in Danish IBD patients, 32.0 versus 13.3% (P = 0.025). The majority of Danish patients were bacillus Calmette-Guerin (Mycobacterium bovis BCG) vaccinated (CD, 77.5%; UC, 86.6%; controls, 83.0%) whereas none of the U.S. patients with IBD and only 2% of U. S. controls were vaccinated. Among Danish IBD patients, positive PCR findings were four times more common among subjects who were not BCG vaccinated (33.3%) than among BCG vaccinates (8.8%, P = 0.02). Culture of the same tissues tested by PCR using modified BACTEC 12B medium failed to grow M. avium subsp. paratuberculosis from patients or controls. U.S. CD patients had the highest serological evidence (enzyme-linked immunosorbent assay [ELISA] for serum antibodies) of M. avium subsp. paratuberculosis infection (20.7% of patients positive) which was higher than for all UC patients studied (6.1%) or healthy controls (3.8%, P < 0.005). Among Danish patients alone, however, no significant differences in rates of ELISA-positive results among CD, UC, or control patients were found. For 181 study subjects, both IS900 PCR and ELISA were performed. Although 11 were ELISA positive and 36 were PCR positive, in no instance was a patient positive by both tests, suggesting that these states are mutually exclusive. Evaluation of cytokine-mediated immune responses of IBD patients was complicated by the influence of immunosuppressive therapy given most IBD patients. Gamma interferon (IFN-gamma) release by peripheral blood leukocytes after M. avium purified protein derivative PPD antigen stimulation showed significantly lower responses in CD patients than in UC patients or controls in both U.S. (by ex vivo assay) and Danish (by in vitro assay) populations (P < 0.05). Interleukin-5 responses were not different among CD, UC, or control groups. Collectively, the PCR, ELISA, and IFN-gamma tests for M. avium subsp. paratuberculosis together with the unexpected observation that BCG vaccination influenced M. avium subsp. paratuberculosis detection, lead us to conclude that M. avium subsp. paratuberculosis, or some similarly fastidious mycobacterial species, infects at least a subset of IBD patients. Whether the infection is primary (causal) or secondary, it may contribute to the etiopathogenesis of IBD. PMID- 11101568 TI - Evaluation of nucleic acid amplification tests as reference tests for Chlamydia trachomatis infections in asymptomatic men. AB - Urine ligase chain reaction (LCR) and PCR tests and urethral swab culture were compared for their abilities to detect Chlamydia trachomatis infection in 3,639 asymptomatic men by using one-, two-, and three-test reference standards. Frozen urine at four of five participating centers was also tested by a transcription mediated amplification assay which was used as a reference test. LCR increased the yield of positive results by 27% and PCR increased the yield of positive results by 26% over the yield of positive results by culture (n = 295). LCR and PCR sensitivities were similar, ranging from 80.4 to 93.5%, depending on the reference standard. Culture sensitivity was substantially less. A multiple-test standard yielded LCR, PCR, and culture specificities of 99.6%, with or without discrepant analysis. Test performance varied among centers partly due to different interpretations of the testing protocols. The study confirms that urine LCR and PCR for the detection of C. trachomatis have substantially improved sensitivities over that of urethral swab culture for testing of asymptomatic men, enabling screening of this important target group. These tests, perhaps in combination, are also candidate reference tests for the conduct of test evaluation studies. PMID- 11101569 TI - Restriction endonuclease analysis discriminates Bordetella bronchiseptica isolates. AB - One hundred ninety-five Bordetella bronchiseptica isolates from 12 different host species worldwide were characterized by restriction enzyme analysis (REA). These isolates had previously been categorized into 19 PvuII ribotypes. Twenty restriction endonucleases were evaluated for use in REA. Digestion of chromosomal DNA with HinfI, followed by submarine electrophoresis in agarose gels and staining with ethidium bromide, produced DNA fragments in the 4.0- to 10-kb range, which readily discriminated B. bronchiseptica isolates, resulting in 48 fingerprint patterns. Moreover, AluI digestion of chromosomal DNA produced 39 distinct fingerprint profiles with DNA fragments ranging from 6.0 to 20.0 kb. While REA frequently provided more discriminatory power than ribotyping, there were examples where the use of ribotyping was more discriminatory than REA. Passage of selected isolates up to passage 25 did not change the REA profile. Moreover, the Bvg phase did not alter the fingerprint profile of chromosomal DNA from B. bronchiseptica strains digested with HinfI or AluI. Based on the results presented herein, the combination of REA and ribotyping should provide valuable information in understanding the molecular epidemiology of B. bronchiseptica infections. PMID- 11101570 TI - Molecular epidemiology of human group A rotavirus infections in the United Kingdom between 1995 and 1998. AB - The G and P types of 2,912 rotavirus-positive fecal specimens collected from eight geographical areas of the United Kingdom between 1995 and 1998 were determined by reverse transcription-PCR. Although 15 different G-P combinations were identified, G1P[8], G2P[4], G3P[8], and G4P[8] strains constituted 95% of all the rotaviruses typed. Other genotypes included G9P[6] and G9P[8], which were first identified in the United Kingdom in 1995, or other uncommon G and/or P types of strains that may have had an animal origin. Unusual combinations of G1 or G4 with P[4] and G2 with P[8] which may have arisen by reassortment between human strains were also identified. G1P[8] was the genotype most frequently found (57 to 87%) in each season, followed by G2P[4] in the 1995-1996 (18%) and 1997 1998 (16%) seasons, although the incidence of infection with this virus decreased significantly to 2% during the 1996-1997 season. Significant differences were seen in the distributions of G1P[8], G2P[4], and G9P[8] strains between children and adults, in the temporal distributions of G4P[8] and G9P[8] strains within a season, and in the geographical distributions of each of the four most common genotypes from one season to the next. PMID- 11101571 TI - Evaluation of three newly developed enzyme-linked immunosorbent assays and two agglutination tests for detecting Salmonella enterica subsp. enterica serovar dublin infections in dairy cattle. AB - In this study test characteristics of three newly developed enzyme-linked immunosorbent assays (ELISAs) for Salmonella enterica subsp. enterica serovar Dublin were evaluated and compared with two agglutination tests. The ELISAs involved were an indirect ELISA with serovar Dublin lipopolysaccharide (LPS ELISA), an indirect ELISA with serovar Dublin flagellar antigen (GP ELISA), and a double-antibody sandwich blocking ELISA that uses monoclonal antibodies against S. enterica subsp. enterica serovar Enteritidis flagellin (GM-DAS ELISA). The agglutination tests involved were two routine serum agglutination tests with either somatic (O) or flagellar (H) antigen. Diagnostic specificity of the three ELISAs was determined using 840 serum samples from seven dairy herds without any history of serovar Dublin infection. Cutoff values at a titer of 100, 100, and 10, respectively, for the LPS ELISA, GP ELISA, and GM-DAS blocking ELISA resulted in a specificity of 99.3, 100, and 100%, respectively. Using these cutoff values the LPS ELISA, GP ELISA, and GM-DAS ELISA were able to detect, respectively, 30, 46, and 38% of 50 fecal culture-positive animals from 13 herds with a recent serovar Dublin infection. With the same cutoff values, active carriers (n = 18) were detected for 94.4% with the LPS ELISA and for 100% with the GP and GM-DAS ELISAs. Kappa values determined on the results of all tests from 8 of the 13 serovar Dublin-infected herds and the 7 control herds demonstrated a good correlation between the results of all ELISAs and the H-agglutination test. The results of the O-agglutination test failed to correlate with those of the other tests. Using a set of sera from 170 aborting cows (with 25 abortions due to serovar Dublin), test results of the ELISAs and the H-agglutination test were comparable. The H-agglutination test may be used successfully for single sample testing, especially to diagnose abortion due to serovar Dublin. It is concluded that the ELISAs are useful diagnostic tools in serovar Dublin control programs and that they are preferred to agglutination tests for reasons of automation and costs. PMID- 11101572 TI - Detection of Chlamydia pneumoniae and Helicobacter pylori DNA in human atherosclerotic plaques by PCR. AB - Chlamydia pneumoniae and Helicobacter pylori can cause persistent infections of the respiratory and gastrointestinal tract, respectively. It has been suggested that persistent infection of arteries with these bacteria can contribute to the development of atherosclerosis. The aims of this study were to determine the presence of C. pneumoniae and H. pylori DNA in atherosclerotic plaque samples by PCR and to evaluate the correlation between clinical status and DNA positivity of these bacteria. Eighty-five consecutive patients (mean age, 59 +/- 10; 75 male, 10 female) undergoing coronary artery bypass grafting, carotid endarterectomy, and surgery of the abdominal aorta for atherosclerotic obstructive lesions were included in the study. Forty-six endarterectomy specimens from the atherosclerotic lesions and 39 specimens from healthy regions of the ascending aorta, which were accepted as the control group, were excised. The presence of microorganism DNA in endarterectomy specimens was assessed by PCR. C. pneumoniae DNA was found in 12 (26%) of 46 endarterectomy specimens and none of the healthy vascular-wall specimens (P < 0.001), while H. pylori DNA was found in 17 (37%) of 46 endarterectomy specimens and none of the controls (P < 0.001). Either C. pneumoniae or H. pylori DNA was positive in 23 (50%) of 46 patients and none of the controls (P < 0. 001). Six of the atherosclerotic lesions showed coexistence of both of the microorganism DNAs. The presence of C. pneumoniae and H. pylori DNA in a considerable number of atherosclerotic plaques but their absence in healthy vascular wall supports the idea that they may have a role in the development of atherosclerosis, especially in countries where infection is prevalent and where conventional risk factors fail to explain the high prevalence of atherosclerotic vascular disease. PMID- 11101573 TI - Antigenic diversity of Haemophilus somnus lipooligosaccharide: phase-variable accessibility of the phosphorylcholine epitope. AB - The lipooligosaccharide (LOS) of Haemophilus somnus undergoes antigenic phase variation, which may facilitate evasion from the bovine host immune response and/or colonization and dissemination. However, LOS antigenic diversity in H. somnus has not been adequately investigated. In this study, monoclonal antibodies (MAbs) specific to various LOS epitopes were used to investigate antigenic variation and stability in LOS from H. somnus strains and phase variants. Clinical isolates of H. somnus exhibited intrastrain, as well as interstrain, antigenic heterogeneity in LOS when probed with MAbs to outer core oligosaccharide epitopes in an enzyme-linked immunosorbent assay (ELISA). However, epitopes reactive with MAbs directed predominately to the inner core heptose region were highly conserved. At least one epitope, which was expressed in few strains, was identified. One LOS component affected by phase variation was identified as phosphorylcholine (PCho), which is linked to the primary glucose residue. Inhibition ELISA, immunoblotting, and electrospray-mass spectrometry were used to confirm that MAb 5F5.9 recognized PCho. LOS reactivity with MAb 5F5.9 was associated with loss of most of the outer core oligosaccharide, indicating that reactivity with PCho was affected by phase variation of the glucose residues in this region. Our results indicate that outer core epitopes of H. somnus LOS exhibit a high degree of random, phase-variable antigenic heterogeneity and that such heterogeneity must be considered in the design of vaccines and diagnostic tests. PMID- 11101574 TI - Bacterial colonization of disposable soft contact lenses is greater during corneal infiltrative events than during asymptomatic extended lens wear. AB - Microorganisms, especially gram-negative bacteria, are considered to play a role in the etiology of certain corneal infiltrative events (CIEs) observed during soft contact lens wear. This study explored the possibility of microbial colonization of soft contact lenses as a risk factor leading to CIEs. In a clinical trial conducted from March 1993 to January 1996, 330 subjects wore disposable soft contact lenses on a 6-night extended-wear and disposal schedule. During this period, 4,321 lenses (118 during CIEs; 4,203 during asymptomatic lens wear) were recovered aseptically and analyzed for microbial colonization. A greater percentage of lenses were free from microbial colonization during asymptomatic wear than during CIEs (42 versus 23%; P < 0.0001). The incidence of gram-positive bacteria, gram-negative bacteria and fungi was greater during CIEs than during asymptomatic lens wear (P < 0.05). During asymptomatic lens wear, gram-positive bacteria were isolated most frequently and were usually normal external ocular microbiota. Of the gram-positive bacteria, the incidence of Streptococcus pneumoniae was greater during CIE than during asymptomatic wear (7.6 versus 0.6%; P < 0. 0001). While gram-negative bacteria were seen in few cases during asymptomatic wear, their incidence during CIE in comparison to asymptomatic wear was substantial and significant (23.7 versus 3.8%; P < 0.0001). Also, the level of colonization was high. Of CIEs, events of microbial keratitis, contact lens acute red eye, and asymptomatic infiltrative keratitis were associated with lens colonization with gram-negative bacteria or S. pneumoniae. Colonization of soft contact lenses with pathogenic bacteria, especially gram negative bacteria and S. pneumoniae, appears to be a significant risk factor leading to CIE. PMID- 11101575 TI - Prevalence of diarrheagenic Escherichia coli in finns with or without diarrhea during a round-the-world trip. AB - The incidence of diarrhea and the prevalence of bacterial enteropathogens, viruses, and parasites in feces of subjects with and without diarrhea were evaluated in 204 Finns traveling round the world (from Finland to China, Malaysia, Australia, Fiji, Chile, and Brazil and back to Finland). Special emphasis was placed on the finding of diarrheagenic Escherichia coli (enterotoxigenic, enteropathogenic, Shiga toxin-producing, and enteroaggregative strains) by PCR from growth on primary culture plates. From the PCR-positive samples, corresponding strains were isolated, confirmed as E. coli, and O serotyped. Of all the subjects, 37% experienced a total of 90 episodes of diarrhea. No adenoviruses or rotaviruses were detected, and findings of parasites were insignificant. In contrast, enteropathogenic bacteria were present in 62% of the 65 diarrheal and in 33% of the 127 nondiarrheal samples (P < 0.001); diarrheagenic E. coli strains were found in 35 and 26% of these, respectively (not statistically significant). As a single pathogen, E. coli was found in 20 and 24% of samples (not significant). Of all diarrheagenic E. coli strains, enteropathogenic strains were the most commonly found independently of the clinical picture of the subjects, whereas Salmonella enterica as a single pathogen was the most common non-E. coli organism found in diarrheal samples. Multiple bacterial pathogens were found 10 times more commonly in diarrheal than in nondiarrheal samples (20 versus 2%; P < 0.001). PMID- 11101577 TI - Molecular characterization and diagnostic value of Taenia solium low-molecular weight antigen genes. AB - Neurocysticercosis (NCC) caused by infection with the larvae of Taenia solium is an important cause of neurological disease worldwide. In order to establish an enzyme-linked immunosorbent assay (ELISA) for this infection using recombinant proteins, we carried out molecular cloning and identified four candidates as diagnostic antigens (designated Ag1, Ag1V1, Ag2, and Ag2V1). Except for Ag2V1, these clones could encode a 7-kDa polypeptide, and Ag2V1 could encode a 10-kDa polypeptide. All of the clones were very similar. Except for Ag2V1, recombinant proteins were successfully expressed using an Escherichia coli expression system. Immunoblot analysis of NCC patient sera detected recombinant proteins, but because reactivity to recombinant Ag1 was too weak, Ag1 was not suitable as an immunodiagnostic antigen. So, Ag1V1 and Ag2 were chosen as ELISA antigens, and the Ag1V1/Ag2 chimeric protein was expressed. Of 49 serum samples from NCC patients confirmed to be seropositive by immunoblot analysis, 44 (89.7%) were positive by ELISA. No assays of serum samples from patients with other parasitic infections recognized the Ag1V1/Ag2 chimeric protein. The Ag1V1/Ag2 chimeric protein obtained in this study had a high value for differential immunodiagnosis. PMID- 11101576 TI - Comparison of sequencing of the por gene and typing of the opa gene for discrimination of Neisseria gonorrhoeae strains from sexual contacts. AB - Typing of gonococcal strains is a valuable tool for the biological confirmation of sexual contacts. We have developed a typing method based on DNA sequencing of two overlapping por gene fragments generated by a heminested PCR. We compared sequencing of the por gene (POR sequencing) and typing of the opa gene (OPA typing) for the characterization of strains from 17 sexual partnerships. Both methods were highly discriminatory. A different genotype was detected in 15 of the 17 epidemiologically unconnected couples by POR sequencing and in 16 of the 17 couples by OPA typing with restriction enzyme HpaII. Within partnerships, identical genotypes were obtained from 16 of the 17 known sex contacts by POR sequencing and from 15 of the 17 by OPA typing. Compared to OPA typing, which relies on interpretation of bands in a gel, DNA sequence data offer the advantage of being objective and portable. As costs for sequencing decline, the method should become affordable for most laboratory personnel who wish to type gonococcal strains. PMID- 11101578 TI - Simple and inexpensive but highly discriminating method for computer-assisted DNA fingerprinting of Pseudomonas aeruginosa. AB - We describe here a method for computer-assisted fingerprinting of Pseudomonas aeruginosa. In this method, DNA is digested with SalI, and bands with molecular sizes of >/=9.7 kb are visually scored after electrophoresis on agarose gels. Pattern scores are entered into a Microsoft Excel database. In scoring, the number of bands within each of a set of molecular size ranges is scored, rather than the absolute molecular size of each band, substantially enhancing the speed and reproducibility of the method, while eliminating the need for using expensive gel scanning equipment and software. Pattern scores are used to generate matrices of genetic distance values, which can be visualized in neighbor-joining trees. The method reliably distinguishes two epidemiologically unrelated isolates in 99.3% of all comparisons. The genetic relationships between isolates observed with the method were consistent with those obtained by analysis of two P. aeruginosa genes, indicating that it provides valid estimates of genetic divergence between isolates. Using the method, respiratory tract isolates from cystic fibrosis patients in Green Lane Hospital in Auckland, New Zealand, were shown to be genetically less diverse than epidemiologically unrelated isolates from other patients. This finding was not due to the existence of clusters of related strains specialized toward colonization of the respiratory tract and thus was indicative of transmission between patients. Analysis of multiple isolates from individual cystic fibrosis patients suggested that up to five separate clusters of genetically related strains may simultaneously be present in a patient. The method described should significantly enhance our ability to investigate the epidemiology of P. aeruginosa. PMID- 11101579 TI - Distinct differences in repertoires of low-molecular-mass secreted antigens of Mycobacterium avium complex and Mycobacterium tuberculosis. AB - Antigens in a 4-week-old culture filtrate (CF) of Mycobacterium avium subsp. avium were separated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by Western blotting. The culture had minimal lysis of bacilli, giving a CF preparation consisting mainly of secreted proteins. Comparison with a similar CF of Mycobacterium tuberculosis with almost no contamination with intracellular proteins showed the presence of cross-reactive antigens homologous to the four components of the antigen 85 complex, as well as MPT32. These were major constituents of the M. avium subsp. avium CF. In addition, there were several low-molecular-mass bands (<15 kDa) in both species that did not cross react with polyclonal and polyvalent rabbit antibodies in Western blotting. Furthermore, these bands were not detected in corresponding sonicate preparations, indicating high localization indexes, which is typical of soluble secreted proteins. A 14-kDa protein was selected for purification and more detailed characterization. The N-terminal amino acid sequence was determined, and a matching gene was found within the genomic sequence of M. avium subsp. avium which was highly homologous to Rv0455c of M. tuberculosis. The gene encoded a signal peptide typical of secreted mycobacterial proteins. A rabbit antiserum was raised against the purified protein, and the antigen was demonstrated by Western blotting in CFs of M. avium subsp. avium, Mycobacterium avium subsp. paratuberculosis, Mycobacterium intracellulare, and Mycobacterium scrofulaceum but was not detected in M. tuberculosis. This is a new example of a highly homologous gene being differentially expressed by different mycobacterial species. PMID- 11101580 TI - Diarrhea in children under 5 years of age from Ifakara, Tanzania: a case-control study. AB - A matched case-control study was conducted in the Maternal and Child Health Clinic (MCH) in Ifakara, Tanzania, during the rainy season in order to elucidate the risk factors for and etiology of diarrheal diseases in children under 5 years of age. Cases (103) and controls (206) were matched for sex and age group. Precoded questionnaires with demographic details, clinical history, and physical signs were completed. Stools samples were collected for bacterial, parasitological, and viral studies. A high number of siblings (odds ratio [OR], 0.86; P = 0.027), the number of siblings surviving (OR, 0.82; P = 0.007), the birth order (OR, 0.85; P = 0.018) and the distance from the house to the water source (OR, 0.33; P = 0.011) were associated with the risk of diarrhea. There were high rates of enteropathogen isolates in stool samples from children without diarrhea (52.23%). Shigella species were the only enteropathogen statistically related with diarrhea (OR, 2.90; P < 0.029). Enterotoxigenic, enteropathogenic, and enteroaggregative strains of Escherichia coli were not related with diarrhea, and neither were Giardia lamblia or Salmonella species. PMID- 11101581 TI - Effects of amplification facilitators on diagnostic PCR in the presence of blood, feces, and meat. AB - The full potential of diagnostic PCR is limited, in part, by the presence of inhibitors in complex biological samples that reduce the amplification efficiency. Therefore, different pre-PCR treatments are being used to reduce the effects of PCR inhibitors. The aim of the present study was to investigate the effects of 16 amplification facilitators to enhance DNA amplification in the presence of blood, feces, or meat. Different concentrations of amplification facilitators and inhibitory samples were added to PCR mixtures containing rTth or Taq DNA polymerase. The addition of 0.6% (wt/vol) bovine serum albumin to reaction mixtures containing Taq DNA polymerase reduced the inhibitory effect of blood and allowed DNA amplification in the presence of 2% instead of 0.2% (vol/vol) blood. Furthermore, the addition of bovine serum albumin (BSA) to reaction mixtures containing feces or meat enhanced the amplification capacities of both polymerases. Taq DNA polymerase was able to amplify DNA in the presence of 4% instead of 0.4% (vol/vol) feces and 4% instead of 0.2% (vol/vol) meat, and rTth was able to amplify DNA in the presence of 4% instead of 0.4% (vol/vol) feces and 20% instead of 2% (vol/vol) meat. The single-stranded DNA binding T4 gene 32 protein (gp32) had a relieving effect similar to that of BSA, except when it was added to PCR mixtures of rTth containing meat and of Taq DNA polymerase containing feces. The relieving effects of betaine and a cocktail of proteinase inhibitors were more sample specific. The addition of 11.7% (wt/vol) betaine allowed Taq DNA polymerase to amplify DNA in the presence of 2% (vol/vol) blood, while the addition of proteinase inhibitors allowed DNA amplification by both polymerases in the presence of 4% (vol/vol) feces. When various combinations of betaine, BSA, gp32, and proteinase inhibitors were tested, no synergistic or additive effects were observed. The effects of facilitators on real-time DNA synthesis instead of conventional PCR were also studied. PMID- 11101582 TI - NSP4 gene analysis of rotaviruses recovered from infected children with and without diarrhea. AB - The transmembrane glycoprotein NSP4 functions as a viral enterotoxin capable of inducing diarrhea in young mice. It has been suggested that NSP4 may be a key determinant of rotavirus pathogenicity and a target for vaccine development. Twenty two G1P[6] rotaviruses from babies with and without diarrhea were comparatively analyzed along with reference strains and another 22 Taiwanese human rotaviruses of G and P combination types different from the G1P[6] type. The sequence variations in the NSP4 genes were studied by direct sequencing analysis of the amplicons of reverse transcription-PCR. Two genetic groups could be identified in this analysis. While the majority of these strains were closely related to the Wa strain, the G2 viruses were closely related to the S2 strain. Furthermore, phylogenetic analysis of the NSP4 gene among the G2 rotaviruses revealed three distinct lineages associated with DS-1, S2, and E210, respectively, as has been reported previously for the VP7 gene. However, we found no apparent correlation in the deduced amino acid sequences corresponding to the proposed enterotoxic peptide region between the rotaviruses recovered from individuals with and without diarrhea. The NSP4 gene product being a pathogenic determinant may not be a generalized phenomenon. PMID- 11101583 TI - Genetic heterogeneity in Mycobacterium tuberculosis isolates reflected in IS6110 restriction fragment length polymorphism patterns as low-intensity bands. AB - Mycobacterium tuberculosis isolates with identical IS6110 restriction fragment length polymorphism (RFLP) patterns are considered to originate from the same ancestral strain and thus to reflect ongoing transmission. In this study, we investigated 1,277 IS6110 RFLP patterns for the presence of multiple low intensity bands (LIBs), which may indicate infections with multiple M. tuberculosis strains. We did not find any multiple LIBs, suggesting that multiple infections are rare in the Netherlands. However, we did observe a few LIBs in 94 patterns (7.4%) and examined the nature of this phenomenon. With single-colony cultures it was found that LIBs mostly represent mixed bacterial populations with slightly different RFLP patterns. Mixtures were expressed in RFLP patterns as LIBs when 10 to 30% of the DNA analyzed originated from a bacterial population with another RFLP pattern. Presumably, a part of the LIBs did not represent mixed bacterial populations, as in some clusters all strains exhibited LIBs in their RFLP patterns. The occurrence of LIBs was associated with increased age in patients. This may reflect either a gradual change of the bacterial population in the human body over time or IS6110-mediated genetic adaptation of M. tuberculosis to changes in the environmental conditions during the dormant state or reactivation thereafter. PMID- 11101584 TI - Diagnostic implications of human cytomegalovirus immediate early-1 and pp67 mRNA detection in whole-blood samples from liver transplant patients using nucleic acid sequence-based amplification. AB - Nucleic acid sequence-based amplification (NASBA) was used for detection of the human cytomegalovirus (CMV) immediate early-1 (IE) and the late pp67 mRNA in 353 blood samples collected from 34 liver transplant patients. The diagnostic value of these assays was compared to that of the pp65 antigenemia assay. Overall, 95 and 42% of the antigenemia-positive samples were IE NASBA and pp67 NASBA positive, respectively. Although the results from pp67 NASBA and the antigenemia assay appeared to correspond poorly, a clear correlation was seen between pp67 NASBA-negative results and low numbers of pp65 antigen-positive cells. Twenty patients (59%) were treated with ganciclovir after the diagnosis of symptomatic CMV infection. Before initiation of the antiviral therapy, the antigenemia assay detected the onset of symptomatic infection in all patients, whereas 95 and 60% of these patients were IE NASBA and pp67 NASBA positive, respectively. Although the sensitivity of IE NASBA was very high, the positive predictive value (PPV) of this assay for the onset of a symptomatic infection was only 63%. The PPV of the antigenemia assay as well as pp67 NASBA was considerably higher (80 and 86%, respectively). Thus, the detection of IE mRNA using NASBA appears to be particularly useful as a marker for early initiation of antiviral therapy in patients at high risk for the development of a symptomatic infection. Also, IE NASBA was found to be more sensitive than the antigenemia assay for monitoring CMV infection during antiviral therapy. On the contrary, pp67 NASBA did not appear to have additional diagnostic value compared to the antigenemia assay. PMID- 11101585 TI - Carried meningococci in the Czech Republic: a diverse recombining population. AB - Population and evolutionary analyses of pathogenic bacteria are frequently hindered by sampling strategies that concentrate on isolates from patients with invasive disease. This is especially so for the gram-negative diplococcus Neisseria meningitidis, a cause of septicemia and meningitis worldwide. Meningococcal isolate collections almost exclusively comprise organisms originating from patients with invasive meningococcal disease, although this bacterium is a commensal inhabitant of the human nasopharynx and very rarely causes pathological effects. In the present study, molecular biology-based techniques were used to establish the genetic relationships of 156 meningococci isolated from healthy young adults in the Czech Republic during 1993. None of the individuals sampled had known links to patients with invasive disease. Multilocus sequence typing (MLST) showed that the bacterial population was highly diverse, comprising 71 different sequence types (STs) which were assigned to 34 distinct complexes or lineages. Three previously identified hyperinvasive lineages were present: 26 isolates (17%) belonged to the ST-41 complex (lineage 3); 4 (2.6%) belonged to the ST-11 (electrophoretic type [ET-37]) complex, and 1 (0.6%) belonged to the ST-32 (ET-5) complex. The data were consistent with the view that most nucleotide sequence diversity resulted from the reassortment of alleles by horizontal genetic exchange. PMID- 11101586 TI - Rapid-cycle PCR method to detect Bordetella pertussis that fulfills all consensus recommendations for use of PCR in diagnosis of pertussis. AB - No standardized PCR method is available for the laboratory diagnosis of the pertussis syndrome. Consensus recommendations for the use of PCR in the diagnosis of Bordetella pertussis infections have been proposed, and the aim of this study was to develop a method that fulfills all of these criteria. A rapid-cycle shared primer PCR method with a microwell format and probe hybridization detection step (POR) was developed using novel oligonucleotides targeted to the outer membrane porin gene (Bordetella spp.). In specimens positive for Bordetella spp., B. pertussis was differentiated from Bordetella parapertussis and Bordetella bronchiseptica by hybridization with organism-specific oligonucleotide probes. An internal control was developed using overlap extension PCR and mouse beta-actin DNA. The analytical specificity was 100%. The analytical sensitivity was comparable to that of nested IS481 and IS1001 PCR ( approximately 1 organism per reaction). The clinical sensitivity and specificity were ascertained using 705 specimens (from 705 patients). The results were compared to those of a nested-PCR method targeting the insertion sequences IS481 and IS1001. Fifty-one specimens were positive for B. pertussis by POR and IS481 PCR. Two specimens which fulfilled a clinical definition of pertussis were positive by POR and negative by IS481 PCR. A total of 652 specimens were negative by both methods. B. parapertussis was not detected in any specimens. PCR inhibition was detected in 21 out of 705 specimens (2.98%). Thus, a rapid (4 h, including specimen preparation) PCR method which fulfills all of the consensus recommendations was developed and validated for the detection of B. pertussis. PMID- 11101587 TI - Identification and phylogenetic relationship of the most common pathogenic Candida species inferred from mitochondrial cytochrome b gene sequences. AB - We sequenced a 396-bp region of the mitochondrial cytochrome b gene of the most common clinically important Candida species: Candida albicans, C. glabrata, C. parapsilosis, C. tropicalis, C. krusei, and C. lusitaniae. The recently described species of Candida, C. dubliniensis, associated with mucosal candidiasis in human immunodeficiency virus-infected individuals, was also included. Two to five strains of each species were examined. Some species represented intraspecies variation, which was not more than 1.8% (DNA). However, interspecies variations were more than 10 and 7%, respectively, for DNA and amino acid sequences. Multiple alignments of nucleotide and deduced amino acid sequences revealed species-specific nucleotides and amino acids. Nucleotide- and amino acid-based phylogenetic trees were constructed and are discussed. Using the database, it is possible to identify presumptive Candida species within a working day. PMID- 11101588 TI - Transfusion-acquired, autochthonous human babesiosis in Japan: isolation of Babesia microti-like parasites with hu-RBC-SCID mice. AB - We have isolated piroplasms from a patient who developed the first case of human babesiosis in Japan by using NOD/shi-scid mice whose circulating erythrocytes (RBCs) had been replaced with human RBCs (hu-RBC-SCID mice). Following inoculation of the patient's blood specimen into hu-RBC-SCID mice, parasites proliferated within the human RBCs in the mice, resulting in a high level of parasitemia. Parasite DNA was prepared from blood samples of the patient and the mice, and the nuclear small-subunit rRNA gene (rDNA) was amplified and sequenced. Both DNA samples gave rise to identical sequences which showed the highest degree of homology (99.2%) with the Babesia microti rDNA. Because the patient had received a blood transfusion before the onset of babesiosis, we investigated the eight donors who were involved. Their archived blood samples were analyzed for specific antibody and parasite DNA; only a single donor was found to be positive by both tests, and the parasite rDNA sequence from the donor coincided with that derived from the patient. The donor's serum exhibited a high antibody titer against the isolate from the patient, whereas it exhibited only a weak cross reaction against B. microti strains isolated in the United States. We conclude that the first Japanese babesiosis case occurred due to a blood transfusion and that the etiological agent is an indigenous Japanese parasite which may be a geographical variant of B. microti. Our results also demonstrated the usefulness of hu-RBC-SCID mice for isolation of parasites from humans and for maintenance of the parasite infectivity for human RBCs. PMID- 11101589 TI - Conservation of type-specific B-cell epitopes of glycoprotein G in clinical herpes simplex virus type 2 isolates. AB - Glycoprotein G (gG-2) of herpes simplex virus type 2 (HSV-2) is cleaved to a secreted amino-terminal portion and to a cell-associated, heavily O-glycosylated carboxy-terminal portion that constitutes the mature gG-2 (mgG-2). The mgG-2 protein is commonly used as a type-specific antigen in the serodiagnosis of HSV-2 infection. As the amino acid sequence variability of mgG-2 in clinical isolates may affect the performance of such assays, the gG-2 gene was sequenced from 15 clinical HSV-2 isolates. Few mutations were identified, and these were mostly localized outside the epitope regions described earlier. Five isolates were identical to different laboratory strains, indicating that the gG-2 gene is highly conserved over time. In the search for HSV-2 isolates harboring mutations within the immunodominant region of mgG-2, a pool of 2,400 clinical HSV-2 isolates was tested for reactivity with two anti-mgG-2 monoclonal antibodies (MAbs). Ten MAb escape HSV-2 mutants, which all harbored structurally restricted single- or dual-point mutations within the respective epitopes explaining the loss of binding, were identified. Sera from corresponding patients were reactive to mgG-2, as well as to a peptide representing the immunodominant region, suggesting that the point mutations detected did not diminish seroreactivity to mgG-2. The conservation of the gG-2 gene reported here further supports the use of mgG-2 as a type-specific antigen in the diagnosis of HSV-2 infections. PMID- 11101590 TI - Characterization of a coronavirus isolated from a diarrheic foal. AB - A coronavirus was isolated from feces of a diarrheic foal and serially propagated in human rectal adenocarcinoma (HRT-18) cells. Antigenic and genomic characterizations of the virus (isolate NC99) were based on serological comparison with other avian and mammalian coronaviruses and sequence analysis of the nucleocapsid (N) protein gene. Indirect fluorescent-antibody assay procedures and virus neutralization assays demonstrated a close antigenic relationship with bovine coronavirus (BCV) and porcine hemagglutinating encephalomyelitis virus (mammalian group 2 coronaviruses). Using previously described BCV primers, the N protein gene of isolate NC99 was amplified by a reverse transcriptase PCR (RT PCR) procedure. The RT-PCR product was cloned into pUC19 and sequenced; the complete N protein of NC99 (446 amino acids) was then compared with published N protein sequences of other avian and mammalian coronaviruses. A high degree of identity (89.0 to 90.1%) was observed between the N protein sequence of NC99 and published sequences of BCV (Mebus and F15 strains) and human coronavirus (strain OC43); only limited identity (<25%) was observed with group 1 and group 3 coronaviruses. Based on these findings, the virus has been tentatively identified as equine coronavirus (ECV). ECV NC99 was determined to have close antigenic and/or genetic relationships with mammalian group 2 coronaviruses, thus identifying it as a member of this coronavirus antigenic group. PMID- 11101591 TI - Strain identification of Trichophyton rubrum by specific amplification of subrepeat elements in the ribosomal DNA nontranscribed spacer. AB - Trichophyton rubrum is the commonest cause of dermatophytosis of skin and nail tissue. Molecular characterization of the T. rubrum ribosomal DNA nontranscribed spacer region revealed two novel tandemly repetitive subelements (TRSs): TRS-1, containing a 27-bp palindromic sequence, and TRS-2. Specific amplification of TRS 1 produced strain-characteristic banding patterns (PCR types), with 21 TRS-1 PCR types recognized from 101 clinical isolates. Four simple patterns representing 1 to 4 copies of TRS-1 accounted for 75 (75%) of all 101 strains, whereas more complex patterns were observed for 21 (20%) of the 101 isolates. The copy number of TRS-2 was 0 to 3 repeats per cistron, with a majority of isolates having two copies of this element. Eleven isolates were polymorphic for TRS-2, and in combination, 23 separate PCR types were recognized by amplification of both TRS-1 and TRS-2. The PCR patterns from both elements were stable and reproducible. Elements with homology to TRS-1 were present in three phylogenetically related species, Trichophyton violaceum, Trichophyton gourvilii, and Trichophyton soudanense, but these elements were not identified in other dermatophyte taxa. There was no clear correlation of PCR type with specimen (skin or nail tissue), but certain PCR types appeared to show a bias in geographic distribution. This new method of typing T. rubrum will enable important questions about pathogenesis and epidemiology of this fungus to be addressed. PMID- 11101592 TI - Variable numbers of TTC repeats in Mycobacterium leprae DNA from leprosy patients and use in strain differentiation. AB - Strain differentiation of Mycobacterium leprae would be of great value for epidemiological investigation to identify the infectious sources of leprosy, to understand transmission patterns, and to distinguish between relapse and reinfection. From the M. leprae genome sequence database, TTC DNA repeats were identified. Primer sets designed to amplify the region flanking TTC repeats revealed PCR products of different sizes, indicating that the number of repeats at each locus may be variable among M. leprae strains. The TTC repeats were not found in Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium marinum, or human tissues, which indicated their specificity to M. leprae. Sequence analysis of the TTC repeat region in each of the M. leprae strains showed a variation of 10 to 37 repeats. In the M. leprae strains of 34 multibacillary patients at Cebu, Philippines, M. leprae with 24 and 25 TTC repeats was most common, and this was followed by strains with 14, 15, 20, 21, and 28 repeats. This study thus indicates that there are variable numbers of TTC repeats in a noncoding region of M. leprae strains and that the TTC region may be useful for strain differentiation for epidemiological investigations of leprosy. PMID- 11101593 TI - Duodenal microflora in very-low-birth-weight neonates and relation to necrotizing enterocolitis. AB - Necrotizing enterocolitis (NEC) is the most common gastrointestinal emergency in the neonatal period. Small-bowel overgrowth with aerobic gram-negative bacteria has previously been implicated in the development of NEC. This prospective study performed quantitative bacteriology on 422 duodenal aspirates collected from 122 very-low-birth-weight (<1,500-g) newborns, at the time of routine changing of nasogastric tubes. Isolates of Enterobacteriaceae were typed by repetitive extragenic, palindromic PCR and pulsed-field gel electrophoresis. One or more samples from 50% of these infants yielded gram-negative bacteria, predominantly Escherichia coli, Klebsiella spp., and Enterobacter spp., with counts up to 10(8) CFU/g. The proportion of samples with gram-negative bacteria increased with postnatal age, while the percentage of sterile samples declined. Molecular typing revealed marked temporal clustering of indistinguishable strains. All infants had been fed prior to isolation of gram-negative organisms. Antibiotic use had no obvious effect on colonization with Enterobacteriaceae. There were 15 episodes of suspected NEC (stage I) and 8 confirmed cases of NEC (2 stage II and 6 stage III) during the study period. Duodenal aspirates were collected prior to clinical onset in 13 episodes of NEC. Seven of these yielded Enterobacteriaceae, of which five strains were also isolated from infants without NEC. Very-low-birth-weight infants have high levels of duodenal colonization with Enterobacteriaceae, with evidence of considerable cross-colonization with indistinguishable strains. There was no association between duodenal colonization with particular strains of Enterobacteriaceae and development of NEC. PMID- 11101594 TI - Genetic relatedness within serotypes of penicillin-susceptible Streptococcus pneumoniae isolates. AB - The molecular epidemiological characteristics of all Streptococcus pneumoniae strains isolated in a nationwide manner from patients with meningitis in The Netherlands in 1994 were investigated. Restriction fragment end labeling analysis demonstrated 52% genetic clustering among these penicillin-susceptible strains, a value substantially lower than the percentage of clustering among Dutch penicillin-nonsusceptible strains. Different serotypes were found within 8 of the 28 genetic clusters, suggesting that horizontal transfer of capsular genes is common among penicillin-susceptible strains. The degree of genetic clustering was much higher among serotype 3, 7F, 9V, and 14 isolates than among isolates of other serotypes, i.e., 6A, 6B, 18C, 19F, and 23F. We further studied the molecular epidemiological characteristics of pneumococci of serotype 3, which is considered the most virulent serotype and which is commonly associated with invasive disease in adults. Fifty epidemiologically unrelated penicillin susceptible serotype 3 invasive isolates originating from the United States (n = 27), Thailand (n = 9), The Netherlands (n = 8), and Denmark (n = 6) were analyzed. The vast majority of the serotype 3 isolates (74%) belonged to two genetically distinct clades that were observed in the United States, Denmark, and The Netherlands. These data indicate that two serotype 3 clones have been independently disseminated in an international manner. Seven serotype 3 isolates were less than 85% genetically related to the other serotype 3 isolates. Our observations suggest that the latter isolates originated from horizontal transfer of the capsular type 3 gene locus to other pneumococcal genotypes. In conclusion, epidemiologically unrelated serotype 3 isolates were genetically more related than those of other serotypes. This observation suggests that serotype 3 has evolved only recently or has remained unchanged over long periods. PMID- 11101595 TI - Comparative genotyping of Candida albicans bloodstream and nonbloodstream isolates at a polymorphic microsatellite locus. AB - Molecular typing studies have shown that the predominant form of reproduction of Candida albicans is clonal and that, in a majority of situations, persistent or recurrent infections are due to a unique strain. Characterization of distinct subpopulations and correlation with clinical features may thus be important to understanding the pathogenesis of candidiasis. In a clonal model, a unique polymorphic marker may identify populations with different biological properties. We therefore compared 48 bloodstream isolates and 48 nonbloodstream matched strains of C. albicans at the elongation factor 3-encoding gene (CEF3) polymorphic microsatellite locus of C. albicans. Sizing of the alleles was performed by automated capillary electrophoresis. A new, 137-bp allele was characterized, and seven nondescribed combinations were observed, resulting in 15 and 11 distinct CEF3 profiles in bloodstream and control strains, respectively. Genotypes 126-135, 130-136, and 131-131 accounted for 60.4% of both bloodstream and control strains. Four bloodstream isolates but no control strains displayed the 135-135 combination. None of the other genotypes was present at an increased frequency in bloodstream isolates. Bloodstream and nonbloodstream strains of C. albicans thus have a heterogeneous structure at the CEF3 locus, with three major and multiple minor allelic combinations. PMID- 11101596 TI - Genomic and phylogenetic analysis of hepatitis C virus isolates from argentine patients: a six-year retrospective study. AB - Typing of hepatitis C virus (HCV) isolates from Argentine patients was performed by using different methodologies in a population of 243 patients. HCV subtype was assigned based upon restriction fragment length polymorphism (RFLP). HCV RNA genomes obtained from serum samples were classified as belonging to clade 1 (53.5%), 2 (23. 0%), or 3 (8.6%); 14.8% of samples showed HCV mixed infections, more frequently implying different subtypes within the same clade. In addition to RFLP typing, phylogenetic relatedness among sequences from both 5' untranslated region (n = 50) and nonstructural 5B coding region (n = 15) was established. PMID- 11101597 TI - Acrophialophora fusispora brain abscess in a child with acute lymphoblastic leukemia: review of cases and taxonomy. AB - A 12-year-old girl with acute lymphoblastic leukemia was referred to King Faisal Specialist Hospital and Research Center. The diagnosis without central nervous system (CNS) involvement was confirmed on admission, and chemotherapy was initiated according to the Children Cancer Group (CCG) 1882 protocol for high risk-group leukemia. During neutropenia amphotericin B (AMB) (1 mg/kg of body weight/day) was initiated for presumed fungal infection when a computed tomography (CT) scan of the chest revealed multiple nodular densities. After 3 weeks of AMB therapy, a follow-up chest CT revealed progression of the pulmonary nodules. The patient subsequently suffered a seizure, and a CT scan of the brain was consistent with infarction or hemorrhage. Because of progression of pulmonary lesions while receiving AMB, antifungal therapy was changed to liposomal AMB (LAMB) (6 mg/kg/day). Despite 26 days of LAMB, the patient continued to have intermittent fever, and CT and magnetic resonance imaging of the brain demonstrated findings consistent with a brain abscess. Aspiration of brain abscess was performed and the Gomori methenamine silver stain was positive for hyphal elements. Culture of this material grew Acrophialophora fusispora. Lung biopsy showed necrotizing fungal pneumonia with negative culture. The dosage of LAMB was increased, and itraconazole (ITRA) was added; subsequently LAMB was discontinued and therapy was continued with ITRA alone. The patient demonstrated clinical and radiological improvement. In vitro, the isolate was susceptible to low concentrations of AMB and ITRA. A. fusispora is a thermotolerant, fast growing fungus with neurotropic potential. We report the first case of human infection involving the CNS. Acrophialophora resembles Paecilomyces but differs in having colonies that become dark and in the development of phialides along the sides or at the tips of echinulate brown conidiophores. Conidia are borne in long chains and are smooth or ornamented with fine-to-coarse echinulations, sometimes in spiral bands. The taxonomy of the genus Acrophialophora is reviewed, and Acrophialophora nainiana and Acrophialophora levis are considered as synonyms of A. fusispora. PMID- 11101598 TI - Infection of hickman catheter by Pseudomonas (formerly flavimonas) oryzihabitans traced to a synthetic bath sponge. AB - Pseudomonas (formerly Flavimonas) oryzihabitans is an uncommon pathogen that may cause catheter-associated infections. Although it has occasionally been isolated from the environment, the source of human infection has not previously been documented. We describe an AIDS patient who developed Pseudomonas oryzihabitans bacteremia due to colonization of a Hickman catheter. The patient reported having strictly followed the recommendations for catheter hygiene. The only flaw detected was the use of a synthetic bath sponge in the shower. The sponge was cultured and yielded P. oryzihabitans among other nonfermentative, gram-negative bacilli. To determine the prevalence of P. oryzihabitans in sponges, we cultured 15 samples from unrelated households. The microorganism was isolated from 3 of the 15 samples. Molecular typing by arbitrarily primed PCR (AP-PCR) was performed with the environmental and clinical isolates. Three different profiles were obtained for the six isolates analyzed from the patient's sponge. The strain from the AIDS patient was identical to one of those from his sponge and was different from all the remaining strains. The AP-PCR typing results were subsequently confirmed by pulsed-field gel electrophoresis. It can be concluded that sponges are occasionally colonized by P. oryzihabitans. For the first time a probable source of an indwelling catheter contamination with this bacterium has been found. Patients carrying these devices should avoid using sponge-like materials, as these are suitable environments for nonfermentative, gram-negative bacilli. PMID- 11101599 TI - Fluorescent amplified-fragment length polymorphism genotyping of Neisseria meningitidis identifies clones associated with invasive disease. AB - Fluorescent amplified-fragment length polymorphism (FAFLP), a genotyping technique with phylogenetic significance, was applied to 123 isolates of Neisseria meningitidis. Nine of these were from an outbreak in a British university; 9 were from a recent outbreak in Pontypridd, Glamorgan; 15 were from sporadic cases of meningococcal disease; 26 were from the National Collection of Type Cultures; 58 were carrier isolates from Ironville, Derbyshire; 1 was a disease isolate from Ironville; and five were representatives of invasive clones of N. meningitidis. FAFLP analysis results were compared with previously published multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) results. FAFLP was able to identify hypervirulent, hyperendemic lineages (invasive clones) of N. meningitidis as well as did MLST. PFGE did not discriminate between two strains from the outbreak that were classified as similar but distinct by FAFLP. The results suggest that high resolution of N. meningitidis for outbreak and other epidemiological analyses is more cost efficient by FAFLP than by sequencing procedures. PMID- 11101600 TI - Cost-effective and rapid presumptive identification of gram-negative bacilli in routine urine, pus, and stool cultures: evaluation of the use of CHROMagar orientation medium in conjunction with simple biochemical tests. AB - The algorithm for a new identification system was designed on the basis of colony color and morphology on CHROMagar Orientation medium in conjunction with simple biochemical tests such as indole (IND), lysine decarboxylase (LDC), and ornithine decarboxylase (ODC) utilization tests with gram-negative bacilli isolated from urine samples as well as pus, stool, and other clinical specimens by the following colony characteristics, biochemical reactions, and serological results: pinkish to red, IND positive (IND(+)), Escherichia coli; metallic blue, IND(+), LDC(+), and ODC negative (ODC(-)), Klebsiella oxytoca; IND(+), LDC(-), and ODC(+), Citrobacter diversus; IND(+) or IND(-), LDC(-), and ODC(-), Citrobacter freundii; IND(-), LDC(+), and ODC(+), Enterobacter aerogenes; IND(-), LDC(-), and ODC(+), Enterobacter cloacae; IND(-), LDC(+), and ODC(-), Klebsiella pneumoniae; diffuse brown and IND(+), Morganella morganii; IND(-), Proteus mirabilis; aqua blue, Serratia marcescens; bluish green and IND(+), Proteus vulgaris; transparent yellow-green, serology positive, Pseudomonas aeruginosa; clear and serology positive, Salmonella sp.; other colors and reactions, the organism was identified by the full identification methods. The accuracy and cost-effectiveness of this new system were prospectively evaluated. During an 8-month period, a total of 345 specimens yielded one or more gram-negative bacilli. A total of 472 gram-negative bacillus isolates were detected on CHROMagar Orientation medium. For 466 of the isolates (98.7%), no discrepancies in the results were obtained on the basis of the identification algorithm. The cost of identification of gram-negative bacilli during this period was reduced by about 70%. The results of this trial for the differentiation of the most commonly encountered gram-negative pathogens in clinical specimens with the new algorithm were favourable in that it permitted reliable detection and presumptive identification. In addition, this rapid identification system not only significantly reduced costs but it also improved the daily work flow within the clinical microbiology laboratory. PMID- 11101601 TI - Characterization of chloramphenicol and florfenicol resistance in Escherichia coli associated with bovine diarrhea. AB - Florfenicol, a veterinary fluorinated analog of thiamphenicol, is approved for treatment of bovine respiratory pathogens in the United States. However, florfenicol resistance has recently emerged among veterinary Escherichia coli isolates incriminated in bovine diarrhea. The flo gene, which confers resistance to florfenicol and chloramphenicol, has previously been identified in Photobacterium piscicida and Salmonella enterica serovar Typhimurium DT104. The flo gene product is closely related to the CmlA protein identified in Pseudomonas aeruginosa. The cmlA gene confers nonenzymatic chloramphenicol resistance via an efflux mechanism. Forty-eight E. coli isolates recovered from calves with diarrhea, including 41 that were both chloramphenicol and florfenicol resistant, were assayed for the presence of both flo and cmlA genes. Forty-two of the 44 isolates for which florfenicol MICs were > or =16 microg/ml were positive via PCR for the flo gene. All E. coli isolates for which florfenicol MICs were < or =8 microg/ml were negative for the flo gene (n = 4) Twelve E. coli isolates were positive for cmlA, and chloramphenicol MICs for all 12 were > or =32 microg/ml. Additionally, eight isolates were positive for both flo and cmlA, and both florfenicol and chloramphenicol MICs for these isolates were > or =64 microg/ml. DNA sequence analysis of the E. coli flo gene demonstrated 98% identity to the published GenBank sequences of both serovar Typhimurium flo(St) and P. piscicida pp-flo. The flo gene was identified on high-molecular-weight plasmids of approximately 225 kb among the majority of florfenicol-resistant E. coli isolates. However, not all of the florfenicol-resistant E. coli isolates tested contained the large flo-positive plasmids. This suggests that several of the E. coli isolates may possess a chromosomal flo gene. The E. coli flo gene specifies nonenzymatic cross-resistance to both florfenicol and chloramphenicol, and its presence among bovine E. coli isolates of diverse genetic backgrounds indicates a distribution much wider than previously thought. PMID- 11101602 TI - Evaluation of Etest for susceptibility testing of multidrug-resistant isolates of Mycobacterium tuberculosis. AB - To prescribe effective treatment schemes for patients with tuberculosis, more efficient susceptibility testing techniques for Mycobacterium tuberculosis are needed, especially in regions with multidrug resistance. Etest (AB BIODISK, Solna, Sweden) is a simple technique that provides quantitative drug susceptibility results for M. tuberculosis in 5 to 10 days from a culture grown at low cost. The performance of Etest was compared to that of the reference proportion method, using 95 M. tuberculosis clinical isolates of which 42.1% (40 of 95) were resistant to at least one antibiotic by the reference method. Overall agreement between Etest and the reference method was 98.9% (94 of 95) for detection of multidrug resistance; for resistance to individual drugs, agreement was 97.9% (93 of 95) for rifampin, 96.0% (92 of 95) for ethambutol, 94.7% (90 of 95) for isoniazid, and 85.3% (81 of 95) for streptomycin. This study supports the utility of Etest for timely detection of drug resistance in M. tuberculosis and for use in tuberculosis control programs. PMID- 11101604 TI - Evaluation of random amplified polymorphic DNA typing of Pseudomonas aeruginosa. AB - A method for distinguishing among Pseudomonas aeruginosa strains using random amplified polymorphic DNA (RAPD) typing was evaluated for reproducibility and discriminatory power. A total of 200 isolates, blinded in triplicate, were evaluated by RAPD. All 600 samples were typeable; 197 of 200 isolates gave identical results on all three occasions, and 131 distinct RAPD types were identified. PMID- 11101603 TI - Development of reverse transcription-PCR (oligonucleotide probing) enzyme-linked immunosorbent assays for diagnosis and preliminary typing of foot-and-mouth disease: a new system using simple and aqueous-phase hybridization. AB - A reverse transcription-PCR (RT-PCR)-enzyme-linked immunosorbent assay system that detects a relatively conserved region within the RNA genome of all seven serotypes of foot-and-mouth disease virus (FMDV) has been developed. The high specificity of the assay is achieved by including a rapid hybridization step with a biotin-labeled internal oligonucleotide. The assay is highly sensitive, fast, and easy to perform. A similar assay, based on a highly variable region of the FMDV genome and employing a single asymmetric RT-PCR and multiple hybridization oligonucleotides, was developed to demonstrate the method's ability to type FMDV. Based on our theoretical and practical knowledge of the methodology, we predict that similar assays are applicable to diagnosis and strain differentiation in any system amenable to PCR amplification. PMID- 11101605 TI - Genotyping of verocytotoxin-producing Escherichia coli O157: comparison of isolates of a prevalent phage type by fluorescent amplified-fragment length polymorphism and pulsed-field gel electrophoresis analyses. AB - We applied the high-resolution genotyping technique fluorescent amplified fragment length polymorphism (FAFLP) analysis to 71 isolates of a single phage type (PT8) of pulsed-field gel electrophoresis (PFGE)-characterized verocytotoxin producing Escherichia coli O157. Twenty-seven similar, but not identical, groupings were defined by both FAFLP analysis and the PFGE profiles. Given the FAFLP analysis conditions described here, these two methods exhibited equivalent discriminatory powers. PMID- 11101606 TI - Vibrio parahaemolyticus serovar O3:K6 as cause of unusually high incidence of food-borne disease outbreaks in Taiwan from 1996 to 1999. AB - The occurrence of food-borne disease outbreaks in Taiwan increased dramatically in 1996, and the incidence has since remained elevated. This increase in outbreaks is correlated with a high rate of isolation of Vibrio parahaemolyticus, which caused between 61 and 71% of the total outbreaks for the period 1996 to 1999. By serotyping, 40 serovars were identified from 3743 V. parahaemolyticus isolates, of which O3:K6 was the most frequently detected. The O3:K6 serovar could have emerged in Taiwan as early as October 1995 and at that time accounted for only 0.6% of the V. parahaemolyticus infections. This level increased suddenly to 50.1% in 1996 and reached a peak (83.8%) in 1997. Comparison of the outbreak profiles for the etiology groups indicates that the high incidence of food-borne disease outbreaks during 1996 to 1999 can be attributed to the extraordinarily high O3:K6 infections. In 1999, the O3:K6 serovar was still prevalent, and accounted for 61.3% of all V. parahaemolyticus infections. Due to its extraordinarily high infection frequency and its capability to spread globally, this organism needs to be intensively monitored internationally. PMID- 11101607 TI - Tween 80 opacity test responses of various Candida species. AB - A total of 111 Candida isolates representing 11 species were examined for their respective responses to a Tween 80 opacity test. The strains of Candida albicans and C. tropicalis that were examined produced an opacity response around their colonies at 2 to 3 days postinoculation. A second group of Candida species yielded a halo around their colonies at 8 to 10 days postinoculation. The remaining Candida species did not produce a positive test response through 10 days postinoculation. The strains of C. dubliniensis were easily differentiated from strains of C. albicans by this test. The Tween 80 opacity test is simple and economical to prepare and is easy to interpret. PMID- 11101608 TI - PCR detection and evidence of shedding of porcine circovirus type 2 in boar semen. AB - An experimental study was conducted to evaluate the potential presence of porcine circovirus type 2 (PCV2) in the semen of infected boars. Four mature boars were inoculated intranasally with PCV2 isolate LHVA-V53 propagated on PK15 cells. Two boars inoculated with the supernatant of noninfected PK15 cells were kept as controls. Serum samples were collected from all boars at 4, 7, 11, 13, 18, 21, 25, 28, 35, and 55 days postinoculation (dpi) and from the four PCV2-infected boars at 90 dpi. Samples were tested for the presence of antibodies to PCV2 by an indirect immunofluorescence assay and for the presence of PCV2 DNA by PCR and nested PCR. Semen samples were collected from all six boars at 5, 8, 11, 13, 18, 21, 25, 28, 33, and 47 dpi and tested for the presence of PCV2 DNA by a nested PCR assay. Antibodies to PCV2 could be detected as early as 11 dpi in one boar, and all four infected boars were found positive for PCV2 antibodies by 18 dpi. Thereafter all infected boars remained positive for antibodies to PCV2 until 90 dpi. Analysis of serum samples by nested PCR demonstrated the presence of PCV2 DNA as early as 4 dpi in three of four infected boars. Serum samples from all infected boars were positive for PCV2 DNA from 11 dpi until 35 dpi but were negative at 90 dpi. PCV2 DNA was detected as soon as 5 dpi in the semen of two infected boars and intermittently thereafter in the semen of all four infected boars. The semen of two infected boars was positive for PCV2 DNA at 47 dpi. Following infection, PCV2 DNA can be detected in semen concurrently with the presence of PCV2 DNA and antibodies in the serum. The present study suggests that PCV2 may be shed intermittently in the semen of infected boars. PMID- 11101609 TI - Antimicrobial resistance of Salmonella isolates from swine. AB - We examined the antimicrobial resistance of 1,257 isolates of 30 serovars of Salmonella enterica subsp. enterica isolated from swine. Serovars Typhimurium and Typhimurium var. Copenhagen were widespread and were frequently multidrug resistant, with distinct resistance to ampicillin, kanamycin, streptomycin, sulfamethoxazole, and tetracycline and to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline, respectively. PMID- 11101610 TI - Comparison of human immunodeficiency virus type 1 RNA sequence heterogeneity in cerebrospinal fluid and plasma. AB - The source of human immunodeficiency virus type 1 (HIV-1) RNA in cerebrospinal fluid (CSF) during HIV-1 infection is uncertain. The sequence heterogeneity of HIV-1 RNA in simultaneous CSF and plasma samples was characterized for five patients at the baseline and during the first week of antiretroviral therapy by two commercial genotyping methodologies. In individual subjects, the sequences in CSF samples differed significantly from those in plasma. In contrast, the viral sequences in CSF at the baseline did not differ from the sequences in CSF during treatment. Similarly, viral sequences in plasma did not vary over this interval. This study provides evidence that HIV-1 RNA in CSF and plasma arise from distinct compartments. PMID- 11101611 TI - Apparent culture-negative prosthetic valve endocarditis caused by Peptostreptococcus magnus. AB - In two patients with prosthetic valve endocarditis due to Peptostreptococcus magnus, blood cultures in the BacT/Alert and BACTEC 9240 systems were signal negative. The capability of the BacT/Alert system to detect various Peptostreptococcus species was assessed. P. magnus and P. anaerobius could not be detected, and subcultures remained negative. The growth in conventional media of these two species and other Peptostreptococcus species was similar. PMID- 11101612 TI - PCR-restriction enzyme analysis of a bone marrow isolate from a human immunodeficiency virus-positive patient discloses polyclonal infection with two Mycobacterium avium strains. AB - Polyclonal infection by Mycobacterium avium was detected by hsp65 PCR-restriction enzyme analysis (PRA) in a bone marrow isolate from an AIDS patient. Two M. avium strains, differing in colony morphology, PRA HaeIII digestion pattern, insertion element (IS) 1245 amplification, and restriction fragment length polymorphism fingerprints with IS1245 and IS1311 probes, were isolated. PMID- 11101613 TI - Comparison of the Rodac imprint method to selective enrichment broth for recovery of vancomycin-resistant enterococci and drug-resistant Enterobacteriaceae from environmental surfaces. AB - We compared the Rodac imprint technique to selective enrichment broth for detecting vancomycin-resistant enterococci (VRE) and multidrug-resistant Enterobacteriaceae (MDRE) on surfaces. Rodac plates contained tryptic soy agar with 5% sheep blood, vancomycin (6 microg/ml), ceftazidime (2 microg/ml), amphotericin B (2 microg/ml), and clindamycin (1 microg/ml). Two types of broth were used: brain heart infusion (BHI) and BHI plus vancomycin (6 microg/ml) and ceftazidime (2 microg/ml) (BHIVC). Of the 46 surfaces cultured for VRE, 12 (26%) were positive. Of the 12 VRE-positive surfaces, 11 (92%) grew from Rodac, 8 (67%) grew from BHIVC, and 7 (58%) grew from BHI. A larger study is needed for MDRE, as only 4 of 43 surfaces were MDRE positive. The Rodac imprint technique successfully recovered VRE from environmental surfaces. PMID- 11101614 TI - Characterization of non-type B Haemophilus influenzae strains isolated from patients with invasive disease. The HI Study Group. AB - Forty-one non-type b Haemophilus influenzae isolates from cases of invasive disease were characterized. By PCR capsular genotyping, 33 nonencapsulated strains, 4 type f isolates, and 4 b(-) strains were identified. By pulsed-field gel electrophoresis, the nonencapsulated isolates exhibited great genetic heterogenicity, whereas the type f and the b(-) strains seemed to have a clonal spread. Occurrence of the hifA gene was found by PCR in 18% of the nonencapsulated, 50% of the b(-), and all of the type f strains. Hemagglutinating fimbriae were generally expressed by nonencapsulated isolates when fimbrial gene hifA was present. Two nonencapsulated isolates not susceptible to ampicillin were detected; no strains were positive for beta-lactamase production. PMID- 11101615 TI - Cryptic genetic diversity in Dientamoeba fragilis. AB - Uncertainty surrounding the role of Dientamoeba fragilis in human disease could be due in part to the existence of pathogenic and nonpathogenic variants. Evidence for two genetically distinct forms was obtained using PCR-restriction fragment length polymorphism analysis of ribosomal genes. Future studies in humans will need to take D. fragilis diversity into account. PMID- 11101616 TI - Detection of Ehrlichia chaffeensis DNA in Amblyomma americanum ticks in Connecticut and Rhode Island. AB - Ehrlichia chaffeensis, the causative agent of human monocytic ehrlichiosis, is transmitted by Amblyomma americanum ticks, which are most abundant in the southern United States. Because serologic evidence suggests that residents of Connecticut are exposed to E. chaffeensis, A. americanum ticks were collected in Connecticut and Rhode Island for PCR analysis to detect E. chaffeensis DNA. Eight of 106 (7.6%) A. americanum ticks from Connecticut and 6 of 52 (11.5%) from Rhode Island contained E. chaffeensis DNA. Thus, E. chaffeensis is present in ticks in southern New England and transmission of E. chaffeensis may occur there. PMID- 11101617 TI - Identification of Corynebacterium glucuronolyticum strains from the urogenital tract of humans and pigs. AB - Bacterial strains isolated from the genital tracts of humans (predominantly males), semen of boars, and uterine and vaginal secretions of sows were identified as Corynebacterium glucuronolyticum and were compared with the type strains of the recently proposed species Corynebacterium glucuronolyticum and Corynebacterium seminale. The two type strains as well as the clinical strains were shown by DNA-DNA hybridization and sequencing of the 16S rRNA gene to be related at the species level. All strains were classified as C. glucuronolyticum, because this name has nomenclatural priority over C. seminale. PMID- 11101618 TI - Evaluation of R-Mix FreshCells in shell vials for detection of respiratory viruses. AB - The performance of a mixture of mink lung and A549 cell lines in shell vials (MSVs) for the detection of respiratory viruses in 159 specimens was evaluated. MSVs, conventional culture, and direct immunofluorescence assay identified 96, 85, and 67% of the influenza A virus-positive specimens, respectively. MSVs provided both a high degree of sensitivity and rapid turnaround times for the detection of influenza A virus. PMID- 11101619 TI - Comparison of susceptibility testing of Mycobacterium tuberculosis using the ESP culture system II with that using the BACTEC method. AB - The ESP Culture System II was evaluated for its capacity to test the susceptibility of 389 cultures of Mycobacterium tuberculosis to streptomycin, rifampin, ethambutol, and isoniazid. Good agreement with results with the BACTEC TB 460 was found. ESP II is a reliable, rapid, and automated method for performing susceptibility testing. PMID- 11101620 TI - Comparison of iodophor and alcohol pledgets with the Medi-Flex blood culture prep kit II for preventing contamination of blood cultures. AB - Iodophor and alcohol pledgets were compared with the Medi-Flex Prep Kit II for skin disinfection before venipuncture. Of 12,367 blood cultures collected, 6,362 were done with conventional pledgets and 6, 005 were done with Medi-Flex kits. Contamination occurred in 351 of 6,362 blood cultures (5.5%; range, 3.7 to 8.1%) with conventional pledgets versus 328 of 6,005 (5.5%; range, 3.5 to 7.5%) with Medi-Flex kits. PMID- 11101621 TI - Detection of bovine herpesvirus 4 (BoHV-4) DNA in the cell fraction of milk of dairy cattle with history of BoHV-4 infection. AB - We have demonstrated, by PCR and restriction enzyme analysis of the PCR product, the presence of bovine herpesvirus 4 (BoHV-4) DNA in the cell fraction of milk from dairy cattle with a history of BoHV-4 infection. We next evaluated the infectious nature of BoHV-4 DNA in those cells. Cocultivation of a BoHV-4 sensitive cell line with BoHV-4 DNA-positive milk cell samples produced cytopathic effects. The same result was obtained from frozen and thawed milk cell fraction coming from the cell milk fraction PCR-positive cows, ensuring that cells were killed and only infectious virus could be recovered after cocultivation with sensitive cells. This report shows that infectious BoHV-4 can be present in milk cells and that therefore nursing may be one of the transmission routes of BoHV-4. PMID- 11101622 TI - High-pathogenicity island of Yersinia spp. in Escherichia coli strains isolated from diarrhea patients in China. AB - The high-pathogenicity island (HPI) of Yersinia has been observed in 93% of 60 enteroadhesive Escherichia coli strains and 80% of E. coli strains isolated from blood samples. In the present study we investigated 671 fecal samples from patients with diarrhea in Shandong Province, China, and isolated HPI-harboring E. coli from 6. 26% of the samples. The isolation rates for patients with diarrhea in three age groups, 10 to 20, 30 to 40, and 50 to 60 years, were 6. 70, 12.35, and 10.81%, respectively. Therefore, HPI-harboring E. coli is the third most frequently isolated enteric pathogen from patients with diarrhea. Vomiting and abdominal pain were recorded for 33.33 and 66.67% of the patients, respectively. Stools with blood were observed for 9.52% of the patients. Twenty-four of 42 (57%) patients experienced a temperature over 37.4 degrees C. These observations indicate that HPI-harboring E. coli is one of the major causes of diarrheal disease in China and that the clinical symptoms caused by HPI-harboring E. coli differ from those caused by enteroadhesive E. coli. PMID- 11101623 TI - Salmonella enterica serovar virchow with CTX-M-like beta-lactamase in Spain. AB - Four Salmonella enterica serovar Virchow strains resistant to broad-spectrum cephalosporins were isolated from patients with gastroenteritis in 1997 and 1998 in Murcia and Barcelona, Spain. The isolates expressed a beta-lactamase with a pI of about 8 and a positive PCR when specific primers for CTX-M-9 were used. These results suggest the presence of a CTX-M-9 beta-lactamase in these strains. PMID- 11101624 TI - Acute echinococcosis: a case report. AB - We report the case of a 69-year-old man with acute pulmonary echinococcosis. A computed tomographic scan of the thorax revealed the presence of multiple nodules in both lungs, and laboratory tests showed eosinophilia and the presence of antibodies against Echinococcus granulosus. Therapy with albendazole led to resolution of the pulmonary nodules and a normalization of the white cell count. To our knowledge this is the first described case of acute echinococcosis, as the diagnosis of this disease is usually delayed to chronic phases. Therefore, finding unexplained eosinophilia, especially in association with pulmonary nodules, should lead one to suspect acute hydatid disease. PMID- 11101625 TI - Molecular identification of Pasteurella dagmatis peritonitis in a patient undergoing peritoneal dialysis. AB - Pasteurella dagmatis was identified as the etiologic agent of peritonitis in a continuous ambulatory peritoneal dialysis patient by utilizing a molecular kit in our hospital's clinical laboratory. This method would appear a useful approach to identify a species of Pasteurella not included in the existing database of commercial identification kits when discrepancies exist between phenotypic tests. PMID- 11101626 TI - Sensitive and quantitative co-detection of two mRNA species by double radioactive in situ hybridization. AB - A better understanding of biological phenomena involving modulations of gene expression requires quantitative analysis of the expression of several genes in the same structure. For this purpose, we have developed a novel in situ hybridization method to quantify two different mRNA species in the same tissue section simultaneously. Two probes labeled with radioelements of significantly different energies ((3)H and (33)P or (35)S) were used to detect the mRNA species. Radioactive images corresponding to the detected mRNA species were acquired with a Micro Imager, a real-time, high-resolution digital autoradiography system. An algorithm was used to process the data such that the initial radioactive image acquired was filtered into two subimages, each representative of the hybridization result specific to one probe. This novel method allows local discrimination and quantification of the respective contributions of each label to each pixel and can therefore be used for quantitative analysis of two mRNAs with a resolution of 15-20 microm. PMID- 11101627 TI - Fluorescence in situ hybridization of scarce leptin receptor mRNA using the enzyme-labeled fluorescent substrate method and tyramide signal amplification. AB - To increase the sensitivity of fluorescence in situ hybridization (FISH) for detection of low-abundance mRNAs, we performed FISH on cryostat sections of rat hypothalamus with biotin-labeled riboprobes to leptin receptor (ObRb) and amplified the signal by combining tyramide signal amplification (TSA) and Enzyme Labeled Fluorescent alkaline phosphatase substrate (ELF) methods. First, TSA amplification was done with biotinylated tyramide. Second, streptavidin-alkaline phosphatase was followed by the ELF substrate, producing a bright green fluorescent reaction product. FISH signal for ObRb was undetectable when TSA or ELF methods were used alone, but intense ELF FISH signal was visible in hypothalamic neurons when the ELF protocol was preceded by TSA. The TSA-ELF was combined with FISH for pro-opiomelanocortin (POMC) and neuropeptide Y (NPY) mRNAs by hybridizing brain sections in a cocktail containing digoxigenin-labeled riboprobes to NPY or POMC mRNA and biotin-labeled riboprobes to ObRb mRNA. Dioxigenin-labeled NPY or POMC mRNA hybrids were subsequently detected first with IgG-Cy3. Then biotin-labeled leptin receptor hybrids were detected with the TSA ELF method. Combining the ELF and TSA amplification techniques enabled FISH detection of scarce leptin receptor mRNAs and permitted the identification of leptin receptor mRNA in cells that also express NPY and POMC gene products. PMID- 11101628 TI - Expression of the membrane-associated carbonic anhydrase isozyme XII in the human kidney and renal tumors. AB - Carbonic anhydrase isozyme XII (CA XII) is a novel membrane-associated protein with a potential role in von Hippel-Lindau carcinogenesis. Although Northern blotting has revealed positive signal for CA XII in normal human kidney, this is the first study to demonstrate its cellular and subcellular localization along the human nephron and collecting duct. Immunohistochemistry with a polyclonal antibody (PAb) raised against truncated CA XII revealed distinct staining in the basolateral plasma membrane of the epithelial cells in the thick ascending limb of Henle and distal convoluted tubules, and in the principal cells of the collecting ducts. A weak basolateral signal was also detected in the epithelium of the proximal convoluted tubules. In addition to the normal kidney specimens, this immunohistochemical study included 31 renal tumors. CA XII showed moderate or strong plasma membrane-associated expression in most oncocytomas and clear cell carcinomas. The segmental, cellular, and subcellular distribution of CA XII along the human nephron and collecting duct suggests that it may be one of the key enzymes involved in normal renal physiology, particularly in the regulation of water homeostasis. High expression of CA XII in some renal carcinomas may contribute to its role in von Hippel-Lindau carcinogenesis. PMID- 11101629 TI - A new monoclonal anti-CD3epsilon antibody reactive on paraffin sections. AB - We generated a new monoclonal antibody (MAb), F7.2.38, by immunizing mice with CD3varepsilongammadelta/CD3omega complexes purified from human T-cells by OKT3 MAb-Sepharose affinity chromatography. Immunoprecipitation experiments and Western blotting analysis showed that MAb F7.2.38 recognized the CD3varepsilon chain in CD3varepsilon cDNA-transfected FOX B-cells and in various T-cell lines. Using flow cytometry on permeabilized or intact cells, the epitope was found to be located in the cytoplasmic tail of the CD3varepsilon chain. Immunohistochemical staining on paraffin-embedded sections showed that the reactivity of MAb F7.2.38 was comparable to that of the commercially available anti-CD3varepsilon polyclonal antibody. Of the 52 well-characterized T-cell lymphomas, 41 were positive for F7. 2.38 (79%), whereas all 37 B-cell lymphomas and 69 non-lymphoid tumors were unreactive. This new anti-CD3varepsilon antibody would be particularly useful for phenotyping T-cell lymphomas on routinely processed paraffin-embedded tissue sections. PMID- 11101630 TI - Localization of xenin-immunoreactive cells in the duodenal mucosa of humans and various mammals. AB - Xenin is a 25-amino-acid peptide extractable from mammalian tissue. This peptide is biologically active. It stimulates exocrine pancreatic secretion and intestinal motility and inhibits gastric secretion of acid and food intake. Xenin circulates in the human plasma after meals. In this study, the cellular origin of xenin in the gastro-entero-pancreatic system of humans, Rhesus monkeys, and dogs was investigated by immunohistochemistry and immunoelectron microscopy. Sequence specific antibodies against xenin detected specific endocrine cells in the duodenal and jejunal mucosa of all three species. These xenin-immunoreactive cells were distinct from enterochromaffin, somatostatin, motilin, cholecystokinin, neurotensin, and secretin cells, and comprised 8.8% of the chromogranin A-positive cells in the dog duodenum and 4.6% of the chromogranin A positive cells in human duodenum. In all three species, co-localization of xenin was found with a subpopulation of gastric inhibitory polypeptide (GIP) immunoreactive cells. Immunoelectron microscopy in the canine duodenal mucosa demonstrated accumulation of gold particles in round, homogeneous, and osmiophilic secretory granules with a closely adhering membrane of 187 +/- 19 nm diameter (mean +/- SEM). This cell type was found to be identical to the previously described canine GIP cell. Immunocytochemical expression of the peptide xenin in a subpopulation of chromogranin A-positive cells as well as the localization of xenin immunoreactivity in ultrastructurally characterized secretory granules permitted the identification of a novel endocrine cell type as the cellular source of circulating xenin. PMID- 11101631 TI - Novel method to quantify neuropil threads in brains from elders with or without cognitive impairment. AB - Pathological alterations in dendrites and axons (i.e., neuritic pathologies) occur in the normal aging brain as well as in brains from elders with mild cognitive impairment and neurodegenerative dementia. These alterations may correlate with clinical measures of cognitive abilities, but the contribution of neuropil threads (NTs), which constitute 85-90% of cortical tau pathology, has not been clear because of the lack of quantitative methodologies. We combined quantitative fractionation and image analysis to devise a strategy for measuring the burden of tau-rich NTs in the entorhinal and perirhinal cortex of brains from elders with and without cognitive impairment, including dementia due to Alzheimer's disease (AD). On the basis of data presented here using this novel strategy, we conclude that this quantitative imaging technique will facilitate efforts to determine the behavioral correlations of neuritic lesions in AD and other brain disorders. PMID- 11101632 TI - Gender differences and the effect of different endocrine situations on the NOS expression pattern in the anterior pituitary gland. AB - The presence of neuronal nitric oxide synthase (nNOS) in two populations of pituitary cells, gonadotrophs (LH) and folliculostellate (FS) cells, suggests that pituitary nitric oxide (NO) is involved in the control of hormone secretion. We have used single and double immunostaining and quantitative procedures to investigate possible gender-related differences in the nNOS expression pattern in the anterior pituitary lobe and its possible alterations in different endocrine situations. Our results reveal a sexual dimorphism in the pattern of nNOS expression. In males, nNOS is mainly found in FS cells, whereas only a few LH cells express nNOS. Conversely, in females, nNOS is mainly found in LH cells. After gonadectomy, paralleling an increase in LH cell size and serum luteinizing hormone (LH) levels, there is nNOS upregulation in LH cells and nNOS downregulation in FS cells. After testoterone replacement, LH cells become nNOS immunonegative again. In lactating rats, LH cells overexpress nNOS, but LH cell size and serum LH levels are low. This suggests that, depending on its cellular source, pituitary NO can exert either an inhibitory or a stimulatory effect on hormone secretion. When released from FS cells, NO exerts a paracrine inhibitory effect, and when released from gonadotrophs it exerts an autocrine or paracrine stimulatory effect on LH or prolactin secretion, respectively. PMID- 11101633 TI - Distribution of H type 1-4 chains of the ABO(H) system in different cell types of human respiratory epithelium. AB - We used three anti-H monoclonal antibodies (MAbs) specific for H Type 1, H Type 2, and H Type 3/4 antigens to investigate the distribution of H Type 1-H Type 4 chains of the ABO(H) histo-blood group in the human respiratory system. Strong staining of H Type 1 chain and weak staining of H Type 2 chain were observed in mucous cells of submucosal glands of bronchial epithelium, which were dependent on the secretor status. No H Type 3/4 chains were detected in mucous cells. Serous cells of submucosal glands of respiratory system showed no staining by three anti-H antibodies. H Type 1 and H Type 3/4 antigens were detected heterogeneously in apical surfaces of bronchial epithelium from secretors but not from nonsecretors. In contrast, basal cells of bronchial epithelium expressed H Type 2 irrespective of the secretor status, probably regulated by the H gene. Some alveolar Type II cells contained only H Types 3/4, which were dependent on the secretor status, whereas alveolar Type I cells had no H antigens. Our results indicated that different cell types in respiratory epithelium expressed different types of carbohydrate chains of histo-blood group antigens under the control of the H or the Se gene. PMID- 11101634 TI - Developmental mucin gene expression in the gastroduodenal tract and accessory digestive glands. I. Stomach. A relationship to gastric carcinoma. AB - Studies were undertaken to provide information regarding cell-specific expression of mucin genes in stomach and their relation to developmental and neoplastic patterns of epithelial cytodifferentiation. In situ hybridization was used to study mRNA expression of eight mucin genes (MUC1-4, MUC5AC, MUC5B, MUC6, MUC7) in stomach of 13 human embryos and fetuses (8-27 weeks' gestation), comparing these with normal, metaplastic, and neoplastic adult tissues. These investigations have demonstrated that MUC1, MUC4, MUC5AC, MUC5B, and MUC6 are already expressed in the embryonic stomach at 8 weeks of gestation. MUC3 mRNA expression can be observed from 10.5 weeks of gestation. MUC2 is expressed at later stages, concomitant with mucous gland cytodifferentiation. Normal adult stomach is characterized by strong expression of MUC1, MUC5AC, and MUC6, less prominent MUC2, and sporadic MUC3 and MUC4, without MUC5B and MUC7. Intestinal metaplasia is characterized by an intestinal-type pattern with MUC2 and MUC3 mRNA expression. Gastric carcinomas exhibit altered mucin gene expression patterns with disappearance of MUC5AC and MUC6 mRNAs in some tumor glands, abnormal expression of MUC2, and reappearance of MUC5B mRNAs. In conclusion, we have observed that patterns of mucin gene expression in embryonic and fetal stomach could show similarities with some gastric carcinomas in adults. Differences in mucin gene expression in developmental, metaplastic, and neoplastic stomach compared to normal adult stomach suggest a possible regulatory role for their products in gastric epithelial cell proliferation and differentiation. PMID- 11101635 TI - Developmental mucin gene expression in the gastroduodenal tract and accessory digestive glands. II. Duodenum and liver, gallbladder, and pancreas. AB - Studies were undertaken to provide information regarding cell-specific expression of mucin genes and their relation to developmental and neoplastic patterns of epithelial cytodifferentiation. In situ hybridization was used to study mRNA expression of mucin genes in duodenum and accessory digestive glands (liver, gallbladder, pancreas) of 13 human embryos and fetuses (6. 5-27 weeks' gestation), comparing these with normal and neoplastic adult tissues. These investigations demonstrated that the pattern of mucin gene expression in fetal duodenum reiterated the patterns we observed during gastric and intestinal ontogenesis, with MUC2 and MUC3 expression in the surface epithelium and MUC6 expression associated with the development of Brunner's glands. In embryonic liver, MUC3 was already expressed at 6.5 weeks of gestation in hepatoblasts. As in adults, MUC1, MUC2, MUC3, MUC5AC, MUC5B, and MUC6 were expressed in fetal gallbladder, whereas MUC4 was not. In contrast, MUC4 was strongly expressed in gallbladder adenocarcinomas. MUC5B and MUC6 were expressed in fetal pancreas, from 12 weeks and 26 weeks of gestation, respectively. Surprisingly, MUC3 which is strongly expressed in adult pancreas, was not detected in developmental pancreas. Taken together, these data show complex spatio-temporal regulation of the mucin genes and suggest a possible regulatory role for mucin gene products in gastroduodenal epithelial cell differentiation. PMID- 11101636 TI - Immunohistological and flow cytometric analysis of glycosphingolipid expression in mouse lymphoid tissues. AB - Expression of neutral glycosphingolipids (GSLs) and gangliosides in normal lymphoid tissues and cells has been studied mostly by biochemical and immunochemical analysis of lipid extracts separated by thin-layer chromatography. GSLs and gangliosides involved in the GM1b biosynthetic pathway were assigned to T-lymphocytes, whereas B-cell gangliosides and GSLs have been poorly characterized in former publications. We used specific polyclonal antibodies in immunohistochemistry and flow cytometry to analyze the distribution of globotriaosylceramide (Gb(3)Cer), globoside (Gb(4)Cer), gangliotriaosylceramide (Gg(3)Cer), gangliotetraosylceramide (Gg(4)Cer), and gangliosides GM3 and GalNAc GM1b in the mouse thymus, spleen, and lymph node. Immature thymocytes expressed epitopes recognized by all antibodies, except for anti-Gb(4)Cer. Mature thymocytes bound only antibodies to GalNAc-GM1b, Gg(4)Cer, and Gb(4)Cer. In secondary lymphoid organs, antibodies to globo-series GSLs bound to vascular spaces of secondary lymphoid organs, whereas the ganglio-series GSL antibodies recognized lymphocyte-containing regions. In a Western blotting analysis, only GalNAc-GM1b antibody recognized a specific protein band in all three organs. Flow cytometric analysis of spleen and lymph node cells revealed that B-cells carried epitopes recognized by all antibodies, whereas the T-cell GSL repertoire was mostly oriented to ganglio-series-neutral GSLs and GM1b-type gangliosides. The results of immunohistochemistry and flow cytometry were not always identical, possibly because of crossreactivity to glycoprotein-linked oligosaccharides and/or differences between cell surface carbohydrate profiles of isolated cells and cells in a tissue environment. PMID- 11101637 TI - Secretion of renin-angiotensin system (RAS) components by normal and tumoral lactotropes. A comparative study using reverse hemolytic plaque assay (RHPA) and immunoelectron microscopy. AB - Immunodetection of renin-angiotensin system (RAS) components indicates that there is a local RAS in anterior pituitary cells, particularly in lactotropes. We have attempted to determine if RAS molecules are secreted by lactotropes and the secretory pathways and intracellular sites of maturation. We investigated the secretory activity of individual lactotropes, using the reverse hemolytic plaque assay (RHPA), with GH3B6 tumor cells and normal male rat pituitary cells. We also determined the subcellular distributions of RAS components in these cells. Both tumor and normal cells secreted angiotensinogen, prorenin, renin, angiotensin I, angiotensin-converting enzyme, and angiotensin II, although at different levels. The percentage of secretory cells was generally higher in tumor lactotropes than in normal cells. The subcellular distribution of RAS components obtained by immunoperoxidase was very similar in both cell types, although the intensities of immunoreactivity differed. Cleaved and uncleaved components were found in rough endoplasmic reticulum (RER), Golgi saccules, and secretory granules, all compartments of the secretory pathway. The cleaved components in the RER suggest the existence of early maturation, whereas the presence of uncleaved products in the secretory granules of normal lactotropes might indicate late maturation sites. PMID- 11101638 TI - Distinct aggregation of beta- and gamma-chains of the high-affinity IgE receptor on cross-linking. AB - The high-affinity IgE receptor (FcepsilonRI) on mast cells and basophils consists of a ligand-binding alpha-chain and two kinds of signaling chains, a beta-chain and disulfide-linked homodimeric gamma-chains. Crosslinking by multivalent antigen results in the aggregation of the bound IgE/alpha-chain complexes at the cell surface, triggering cell activation, and subsequent internalization through coated pits. However, the precise topographical alterations of the signaling beta and gamma-chains during stimulation remain unclarified despite their importance in ligand binding/signaling coupling. Here we describe the dynamics of FcepsilonRI subunit distribution in rat basophilic leukemia cells during stimulation as revealed by immunofluorescence and immunogold electron microscopy. Immunolocalization of beta- and gamma-chains was homogeneously distributed on the cell surfaces before stimulation, while crosslinking with multivalent antigen, which elicited optimal degranulation, caused a distinct aggregation of these signaling chains on the cell membrane. Moreover, only gamma- but not beta-chains were aggregated during the stimulation that evoked suboptimal secretion. These findings suggest that high-affinity IgE receptor beta- and gamma-chains do not co aggregate but for the most part form homogenous aggregates of beta-chains or gamma-chains after crosslinking. PMID- 11101639 TI - A window into the role of inhibitory and excitatory mechanisms of perception? AB - The last few years have seen important advances in the better understanding of human cortical physiology. Functional neuroimaging and transcranial magnetic stimulation (TMS) have allowed us to address novel questions and shed different and complementary information on human brain function. In particular, TMS, a non invasive and virtually painless tool for delivering currents in the brain, has enhanced our understanding of perceptual and motor functions in intact humans. At the same time it has also yielded data on phenomenology, mechanisms, and strategies to modulate these plastic changes in health and disease. Studies using a paired-pulse protocol (Kujirai et al. 1993) allowed detailed investigations of intracortical excitatory and inhibitory interactions in the human motor cortex. This protocol studied the effects of a TMS pulse, delivered at a subthreshold intensity to elicit a motor evoked potential (MEP), on the MEP response to an upcoming suprathreshold TMS pulse. Depending on the interval, the effect of this conditioning pulse may be inhibitory (intracortical inhibition) or excitatory (intracortical facilitation). Abnormalities in these interactions have been described in Parkinson's disease and dystonia (Ridding et al. 1995); differences with control measures have also been described as a function of motor training or learning (Liepert et al. 1998). This protocol has allowed investigators to study mechanisms of both disease and cortical reorganization, for example after amputations (Chen et al. 1998) or motor training processes. PMID- 11101640 TI - Characterisation of human monocarboxylate transporter 4 substantiates its role in lactic acid efflux from skeletal muscle. AB - Monocarboxylate transporter (MCT) 4 is the major monocarboxylate transporter isoform present in white skeletal muscle and is responsible for the efflux of lactic acid produced by glycolysis. Here we report the characterisation of MCT4 expressed in Xenopus oocytes. The protein was correctly targeted to the plasma membrane and rates of substrate transport were determined from the rate of intracellular acidification monitored with the pH-sensitive dye 2', 7'-bis (carboxyethyl)-5(6)-carboxyfluorescein (BCECF). In order to validate the technique, the kinetics of monocarboxylate transport were measured in oocytes expressing MCT1. Km values determined for L-lactate, D-lactate and pyruvate of 4.4, > 60 and 2.1 mM, respectively, were similar to those determined previously in tumour cells. Comparison of the time course of [14C]lactate accumulation with the rate of intracellular acidification monitored with BCECF suggests that the latter reflects pH changes close to the plasma membrane associated with transport, whilst the former may include diffusion-limited movement of lactate into the bulk cytosol. Km values of MCT4 for these substrates were found to be 28, 519 and 153 mM, respectively, and for a range of other monocarboxylates values were at least an order of magnitude higher than for MCT1. Vmax values appeared to be similar for all substrates. K0.5 values of MCT4 (determined at 30 mM L-lactate) for inhibition by alpha-cyano-4-hydroxycinnamate (991 microM), phloretin (41 microM), 5-nitro-2-(3-phenylpropylamino)benzoate (240 microM), p chloromercuribenzene sulphonate (21 microM) and 3-isobutyl-1-methylxanthine (970 microM, partial inhibition) were also substantially higher than for MCT1. No inhibition of MCT4 by 2 mM 4,4'-diisothiocyanostilbene-2,2'-disulphonate was observed. The properties of MCT4 are consistent with published data on giant sarcolemmal vesicles in which MCT4 is the dominant MCT isoform, and are appropriate for the proposed role of MCT4 in mediating the efflux from the cell of glycolytically derived lactic acid but not pyruvate. PMID- 11101641 TI - Voltage-dependent conductance changes in the store-operated Ca2+ current ICRAC in rat basophilic leukaemia cells. AB - Tight-seal whole-cell patch-clamp experiments were carried out in order to investigate the effects of different holding potentials on the rate of development and amplitude of the Ca2+ release-activated Ca2+ current ICRAC in rat basophilic leukaemia (RBL-1) cells. ICRAC was monitored at -80 mV from fast voltage ramps, spanning 200 mV in 50 ms. At hyperpolarised potentials, the macroscopic CRAC conductance was lower than that seen at depolarised potentials. The conductance increased almost 5-fold over the voltage range -60 to +40 mV and was seen when the stores were depleted either by the combination of IP3 and thapsigargin in high Ca2+ buffer, or passively with 10 mM EGTA or BAPTA. The voltage-dependent conductance of the CRAC channels could not be fully accounted for by Ca2+-dependent fast inactivation, nor by other slower inhibitory mechanisms. It also did not seem to involve intracellular Mg2+ or the polycations spermine and spermidine. Voltage step relaxation experiments revealed that the voltage-dependent conductance changes developed and reversed slowly, with a time constant of several seconds at -60 mV. In the presence of physiological levels of intracellular Ca2+ buffers, ICRAC was barely detectable when cells were clamped at -60 mV and dialysed with IP3 and thapsigargin, but at 0 mV the current in low Ca2+ buffer was as large as that seen in high Ca2+ buffer. Our results suggest that CRAC channels exhibit slow voltage-dependent conductance changes which can triple the current amplitude over the physiological range of voltages normally encountered by these cells. The role of this conductance change and possible underlying mechanisms are discussed. PMID- 11101642 TI - Functional IP3- and ryanodine-sensitive calcium stores in presynaptic varicosities of NG108-15 (rodent neuroblastoma x glioma hybrid) cells. AB - Presynaptic varicosities of the model neuronal cell line NG108-15, a cholinergic neuroblastoma cell x glioma cell hybrid capable of innervating striated myotubes, were examined for the presence of inositol 1,4,5-trisphosphate (IP3)-sensitive and Ca2+-activated (ryanodine-sensitive) Ca2+ stores using confocal microscopic imaging of Ca2+-sensitive fluorescent dye loaded into the cells. Initial demonstration of the presence of IP3 receptors and ryanodine receptors in the NG108-15 varicosities was obtained using immunocytochemistry. Treatment of NG108 15 cells with bradykinin (0.1 microM), whose receptor is linked to IP3 generation, and separately, caffeine (10 mM), an activator of endoplasmic reticulum ryanodine receptors, resulted in substantial increases in [Ca2+]i in the varicosities. K+-evoked changes in [Ca2+]i in the varicosities were reduced (52 %) after emptying the ryanodine-sensitive Ca2+ store using caffeine (10 mM), but were not affected by prior depletion of the IP3-sensitive Ca2+ store using thapsigargin (1 microM). Bradykinin-induced changes in [Ca2+]i were abolished following depletion of the IP3-sensitive Ca2+ store using thapsigargin (1 microM) and were reduced (72 %) by prior emptying of the ryanodine-sensitive Ca2+ store with caffeine (10 mM). The same results were obtained when the varicosities of the NG108-15 cells had formed synaptic junctions with co-cultured rat hindlimb myotubes. Taken together, the results suggest that, in the varicosities, activation of the IP3 pathway evoked the release of Ca2+ from the IP3-sensitive store, which, in turn, secondarily induced the release of Ca2+ from the ryanodine sensitive store via Ca2+-induced Ca2+ release, and that depolarization-induced Ca2+ entry evoked Ca2+-induced Ca2+ release only from the ryanodine-sensitive store. Thus, functional internal Ca2+ stores are inherent components of presynaptic varicosities in this neural cell line. PMID- 11101643 TI - The role of Ca2+ stores in the muscarinic inhibition of the K+ current IK(SO) in neonatal rat cerebellar granule cells. AB - Cerebellar granule neurons (CGNs) possess a standing outward potassium current (IK(SO)) which shares many similarities with current through the two-pore domain potassium channel TASK-1 and which is inhibited following activation of muscarinic acetylcholine receptors. The action of muscarine on IK(SO) was unaffected by the M2 receptor antagonist methoctramine (100 nM) but was blocked by the M3 antagonist zamifenacin, which, at a concentration of 100 nM, shifted the muscarine concentration-response curve to the right by around 50-fold. Surprisingly, M3 receptor activation rarely produced a detectable increase in [Ca2+]i unless preceded by depolarization of the cells with 25 mM K+. Experiments with thapsigargin and ionomycin suggested that the endoplasmic reticulum Ca2+ stores in CGNs were depleted at rest. In contrast, cerebellar glial cells in the same fields of cells possessed substantial endoplasmic reticulum Ca2+ stores at rest. Pretreatment of the cells with BAPTA AM, thapsigargin or the phospholipase C (PLC) inhibitor U-73122 all blocked the muscarine-induced Ca2+ signal but had little or no effect on muscarinic inhibition of IK(SO). Raising [Ca2+]i directly with ionomycin caused a small but significant inhibition of IK(SO). It is concluded that muscarine acts on M3 muscarinic acetylcholine receptors both to inhibit IK(SO) and to mobilize Ca2+ from intracellular stores in CGNs. While the mobilization of Ca2+ occurs through activation of PLC, this does not seem to be the primary mechanism underlying muscarinic inhibition of IK(SO). PMID- 11101644 TI - Modulation of the synaptic Ca2+ current in salamander photoreceptors by polyunsaturated fatty acids and retinoids. AB - Synaptic transmission between retinal photoreceptors and second-order neurones is controlled by an L-type Ca2+ conductance (gCa) in the photoreceptor inner segment. Modulation of this conductance therefore influences the flow of visual information to higher centres. Possible modulation of gCa by retinal factors was investigated using patch clamp and Ca2+ imaging. No significant modulation of gCa by retinal neurotransmitters nor by intracellular signalling pathways was found. gCa was inhibited by retinoids (all-trans retinal) and by polyunsaturated fatty acids (PUFAs) such as arachidonic acid and docosahexaenoic acid, which are known to be released in the retina by exposure to light. Some PUFAs tested are physiological substrates for the cyclo-oxygenase, lipoxygenase and epoxygenase pathways, but specific inhibitors of these pathways had no effect on the inhibition of gCa. Treatments designed to activate or inhibit G-protein-coupled pathways or protein kinases A and C similarly had no effect on the inhibition by PUFAs nor on gCa itself. Inhibitors of phosphatases 1 and 2A were also largely ineffective. The inhibition by PUFAs is, however, dependent on membrane potential, suggesting that it arises from a direct interaction of fatty acids with the Ca2+ channel. The effect was not use or frequency dependent, suggesting that the effect does not depend on channel gating state. Control by retinoids and by PUFAs may be an important mechanism by which the Ca2+ conductance, and consequently the transmission of the visual signal, is modulated at the first retinal synapse. PMID- 11101645 TI - Molecular diversity of the repolarizing voltage-gated K+ currents in mouse atrial cells. AB - Voltage-clamp studies on atrial myocytes isolated from adult and postnatal day 15 (P15) C57BL6 mice demonstrate the presence of three kinetically distinct Ca2+ independent, depolarization-activated outward K+ currents: a fast, transient outward current (Ito,f), a rapidly activating, slowly inactivating current (IK,s) and a non-inactivating, steady-state current (Iss). The time- and voltage dependent properties of to,f, IK,s and Iss in adult and P15 atrial cells are indistinguishable. Pharmacological experiments reveal the presence of two components of IK,s: one that is blocked selectively by 50 microM 4-aminopyridine (4-AP), and a 4-AP-insensitive component that is blocked by 25 mM TEA; Iss is also partially attenuated by 25 mM TEA. There are also two components of IK,s recovery from steady-state inactivation. To explore the molecular correlates of mouse atrial IK,s and Iss, whole-cell voltage-clamp recordings were obtained from P15 and adult atrial cells isolated from transgenic mice expressing a mutant Kv2.1 alpha subunit (Kv2.1N216Flag) that functions as a dominant negative, and from P15 atrial myocytes exposed to (1 microM) antisense oligodeoxynucleotides (AsODNs) targeted against Kv1.5 or Kv2.1. Peak outward K+ current densities are attenuated significantly in atrial myocytes isolated from P15 and adult Kv2.1N216Flag-expressing animals and in P15 cells exposed to AsODNs targeted against either Kv1.5 or Kv2.1. Analysis of the decay phases of the outward currents evoked during long (5 s) depolarizing voltage steps revealed that IK, s is selectively attenuated in cells exposed to the Kv1.5 AsODN, whereas both IK,s and Iss are attenuated in the presence of the Kv2. 1 AsODN. In P15 and adult Kv2.1N216Flag-expressing atrial cells, mean +/- s.e.m. IK,s and Iss densities are also significantly lower than in non-transgenic atrial cells. In addition, pharmacological experiments reveal that the TEA-sensitive component IK,s is selectively eliminated in P15 and adult Kv2.1N216Flag-expressing atrial cells. Taken together, the results presented here reveal that both Kv1.5 and Kv2.1 contribute to mouse atrial IK,s, consistent with the presence of two molecularly distinct components of IK,s. In addition, Kv2.1 contributes to mouse atrial Iss. PMID- 11101646 TI - A subthreshold persistent sodium current mediates bursting in rat subfornical organ neurones. AB - It is widely accepted that while release of amino acid neurotransmitters occurs with relatively high fidelity, peptidergic synapses require clustered bursts of action potentials for optimal transmitter release. Here we describe for the first time the occurrence and mechanisms of bursting by neurones in the subfornical organ (SFO), cells that utilize the peptide angiotensin II (ANG) in neurotransmission in autonomic pathways. In current clamp recording of isolated SFO neurones in vitro, 53 % (n = 74) showed either spontaneous or evoked burst like discharge patterns. Bursts typically appeared as shifts in bistable membrane potential, with action potentials superimposed on a depolarizing afterpotential (DAP). Similarly, in vivo single unit recordings of identified SFO neurones showed that 9 of 15 neurones fired in bursts. The pattern of bursting, as well as duration of evoked DAPs was strongly dependent upon membrane potential, suggesting that the DAP contributes to burst generation. Based on our previous observation of calcium-sensing receptor (CaR)-activated bursts, we investigated the effects of NPS R-467, an allosteric agonist of the CaR, on evoked DAPs. NPS R 467 (1 microM) potentiated DAP duration throughout the voltage range tested. We observed a dependence of evoked DAPs upon Na+ channels, as shown by sensitivity to tetrodotoxin (0.5 microM) or reduction of external [Na+] from 140 to 40 mM. The duration of DAPs suggested that a persistent Na+ current mediates these events. Voltage-clamp analysis revealed the presence of a subthreshold sodium current, INaP. Pharmacological blockade of INaP with 100 microM lidocaine reduced the duration of evoked DAPs, and inhibited bursting in SFO neurones. Facilitation of INaP with 10 nM anemone toxin (ATX) increased DAP duration and led to marked excitation of bursting cells. These data indicate that INaP is the main current underlying bursting in SFO neurones. Our observations of receptor-mediated facilitation of bursting by SFO neurones represents an intriguing mechanism through which the release of the peptide neurotransmitter ANG may be regulated. PMID- 11101647 TI - Presynaptic 5-HT3 receptor-mediated modulation of synaptic GABA release in the mechanically dissociated rat amygdala neurons. AB - Nystatin-perforated patch recordings were made from mechanically dissociated basolateral amygdala neurons with preserved intact native presynaptic nerve terminals to study the mechanism of 5-HT3 receptor-mediated serotonergic modulation of GABAergic inhibition. The specific 5-HT3 agonist mCPBG (1 microM) rapidly facilitated the frequency of GABAergic miniature inhibitory postsynaptic currents (mIPSCs) and this facilitation desensitized within 1 min. Tropisetron (30 nM), a specific 5-HT3 antagonist, blocked the mCPBG effect. mCPBG augmented mIPSC amplitude. However, no direct postsynaptic serotonergic currents were evoked by mCPBG. Neither GABA-evoked current amplitude nor the kinetics of individual GABAergic mIPSCs were affected by mCPBG. Therefore, the augmentation is unlikely to be due to postsynaptic effects evoked by mCPBG. At higher concentrations mCPBG produced shorter-duration facilitation of miniature events. While mCPBG increased the mIPSC frequency in calcium-containing solution with Cd2+, this increase was absent in Ca2+-free external solution. It appears that the Ca2+ influx through voltage-dependent calcium channels was not as crucial as that through 5-HT3 receptors for synaptic GABA release. When two pulses of mCPBG (each 1 microM, 1 min) were given, the response to the second pulse elicited full recovery when the interval between pulses was at least 9 min. Protein kinase A (PKA) activation by 8-Br-cAMP (300 microM) shortened and PKA inhibition by Rp cAMP (100 microM) prolonged the recovery time. PKA activity did not affect the time course of fast desensitization. Our results suggest that a 5-HT3-specific agonist acts on presynaptic nerve terminals facilitating synaptic GABA release without postsynaptic effects. The facilitation requires calcium influx through presynaptic 5-HT3 receptors. PKA modulates the recovery process from desensitization of presynaptic 5-HT3 receptor-mediated regulation of synaptic GABA release. PMID- 11101648 TI - Differential expression of volume-regulated anion channels during cell cycle progression of human cervical cancer cells. AB - This study investigated the volume-regulated anion channel (VRAC) of human cervical cancer SiHa cells under various culture conditions, testing the hypothesis that the progression of the cell cycle is accompanied by differential expression of VRAC activity. Exponentially growing SiHa cells expressed VRACs, as indicated by the presence of large outwardly rectifying currents activated by hypotonic stress with the anion permeability sequence I- > Br- > Cl-. VRACs were potently inhibited by tamoxifen with an IC50 of 4.6 [mu]M. Fluorescence-activated cell sorting (FACS) experiments showed that 59 +/- 0.5, 5 +/- 0.5 and 36 +/- 1.1% of unsynchronized, exponentially growing cervical cancer SiHa cells were in G0/G1, S and G2/M stage, respectively. Treatment with aphidicolin (5 [mu]M) arrested 88 +/- 1.4% of cells at the G0/G1 stage. Arrest of cell growth in the G0/G1 phase was accompanied by a significant decrease of VRAC activity. The normalized hypotonicity-induced current decreased from 48 +/- 5.2 pA pF-1 at +100 mV in unsynchronized cells to 15 +/- 2.6 pA pF-1 at +100 mV in aphidicolin treated cells. After removal of aphidicolin, culturing in medium containing 10% fetal calf serum triggered a rapid re-entry into the cell cycle and a concomitant recovery of VRAC density. Pharmacological blockade of VRACs by tamoxifen or NPPB caused proliferating cervical cancer cells to arrest in the G0/G1 stage, suggesting that activity of this channel is critical for G1/S checkpoint progression. This study provides new information on the functional significance of VRACs in the cell cycle clock of human cervical cancer cells. PMID- 11101649 TI - Regulation of basal intracellular calcium concentration by the sarcoplasmic reticulum in myocytes from the rat gastric antrum. AB - The intracellular calcium concentration ([Ca2+]i) was monitored in fura-2-loaded myocytes isolated from the rat gastric antrum and voltage clamped at -60 1r1rqmV1qusing the perforated patch clamp technique. The rate of quench of fura-2 fluorescence by Mn2+ was used as a measure of capacitative Ca2+ entry. Cyclopiazonic acid (5 microM) did not affect the holding current but produced a sustained elevation in steady-state [Ca2+]i that was dependent on the presence of external calcium. Cyclopiazonic acid increased Mn2+ influx with physiological external [Ca2+], but not in Ca2+-free conditions. Cyclopiazonic acid increased the rate of [Ca2+]i rise following a rapid switch from Ca2+-free to physiological [Ca2+] solution. Sustained application of carbachol (10 microM) produced an elevation in steady-state [Ca2+]i that was associated with an increased rate of Mn2+ influx. Application of cyclopiazonic acid in the presence of carbachol further elevated steady-state [Ca2+]i without changing Mn2+ influx. Ryanodine (10 microM) elevated steady-state [Ca2+]i either on its own or following a brief application of caffeine (10 9i1s1sqmMc1q). Cyclopiazonic acid had no further effect when added to cells pre-treated with ryanodine. Neither caffeine nor ryanodine increased the rate of Mn2+ influx. When brief applications of ionomycin (25 microM) in Ca2+-free solution were used to release stored Ca2+, ryanodine reduced the amplitude of the resulting [Ca2+]i transients by approximately 30 %, indicating that intracellular stores were partially depleted. These findings suggest that continual uptake of Ca2+ by the sarcoplasmic reticulum Ca2+-ATPase into a ryanodine-sensitive store limits the bulk cytoplasmic [Ca2+]i under resting conditions. This pathway can be short circuited by 10 microM ryanodine, presumably by opening Ca2+ channels in the sarcoplasmic reticulum. Depletion of stores with cyclopiazonic acid or carbachol also activates capacitative Ca2+ entry. PMID- 11101650 TI - Cytochalasin D reduces Ca2+ sensitivity and maximum tension via interactions with myofilaments in skinned rat cardiac myocytes. AB - The F-actin disrupter cytochalasin D depresses cardiac contractility, an effect previously ascribed to the interaction of cytochalasin D with cytoskeletal actin. We have investigated the possibility that this negative inotropic effect is due to the interaction of cytochalasin D with sarcomeric actin of the thin filament. Confocal images of Triton X-100-skinned myocytes incubated with a fluorescent conjugate of cytochalasin D revealed a longitudinally striated pattern of binding, consistent with a myofibrillar rather than cytoskeletal structure.Tension-pCa relationships were determined at sarcomere lengths (SLs) of 2.0 and 2.3 [mu]m following 2 min incubation with 1 [mu]M cytochalasin D. Cytochalasin D significantly reduced the pCa for half-maximal activation (pCa50) at both SLs. The shift in pCa50 was significantly greater at a SL of 2.3 [mu]m compared with that at a SL of 2.0 [mu]m. Cytochalasin D had no effect on the Hill co-efficient at either SL. Cytochalasin D significantly reduced the maximum tension at both SLs. We suggest that the length-dependent decrease in myofilament Ca2+ sensitivity in response to cytochalasin D is due to a decrease in the affinity of troponin C for Ca2+. Cytochalasin D has been used for many years as the agent of choice for disruption of cytoskeletal actin. However, we have demonstrated for the first time an interaction of cytochalasin D with sarcomeric actin of the thin filament, which can account for the effects of cytochalasin D on cardiac contractility. PMID- 11101651 TI - A brainstem area mediating cerebrovascular and EEG responses to hypoxic excitation of rostral ventrolateral medulla in rat. AB - We sought to identify the medullary relay area mediating the elevations of regional cerebral blood flow (rCBF) and synchronization of the electroencephalogram (EEG) in the rat cerebral cortex elicited by hypoxic excitation of reticulospinal sympathoexcitatory neurons of the rostral ventrolateral medulla (RVLM ). In anaesthetized spinalized rats electrical stimulation of RVLM elevated rCBF (laser-Doppler flowmetry) by 31 +/- 6 %, reduced cerebrovascular resistance (CVR) by 26 +/- 8 %, and synchronized the EEG, increasing the power of the 5-6 Hz band by 98 +/- 25 %. Stimulation of a contiguous caudal region, the medullary cerebral vasodilator area (MCVA), had comparable effects which, like responses of RVLM, were replicated by microinjection of L-glutamate (5 nmol, 20 nl). Microinjection of NaCN (300 pmol in 20 nl) elevated rCBF (17 +/- 5 %) and synchronized the EEG from RVLM, but not MCVA, while nicotine (1.2 nmol in 40 nl) increased rCBF by 13 +/- 5 % and synchronized the EEG from MCVA. In intact rats nicotine lowered arterial pressure only from MCVA (101 +/- 3 to 52 +/- 9 mmHg). Bilateral electrolytic lesions of MCVA significantly reduced, by over 59 %, elevations in rCBF and, by 78 %, changes in EEG evoked from RVLM. Bilateral electrolytic lesions of RVLM did not affect responses from MCVA. Anterograde tracing with BDA demonstrated that RVLM and MCVA are interconnected. The MCVA is a nicotine-sensitive region of the medulla that relays signals elicited by excitation of oxygen-sensitive reticulospinal neurons in RVLM to reflexively elevate rCBF and slow the EEG as part of the oxygen-conserving (diving) reflex initiated in these neurons by hypoxia or ischaemia. PMID- 11101652 TI - Responses of aortic depressor nerve-evoked neurones in rat nucleus of the solitary tract to changes in blood pressure. AB - Using electrophysiological techniques, the discharge of neurones in the nucleus of the solitary tract (NTS) receiving aortic depressor nerve (ADN) inputs was examined during blood pressure changes induced by I.V. phenylephrine or nitroprusside in anaesthetized, paralysed and artificially ventilated rats. Various changes in discharge rate were observed during phenylephrine-induced blood pressure elevations: an increase (n = 38), a decrease (n = 5), an increase followed by a decrease (n = 4) and no response (n = 11). In cells receiving a monosynaptic ADN input (MSNs), the peak discharge frequency response was correlated to the rate of increase in mean arterial pressure (P < 0.01) but was not correlated to the absolute increase in blood pressure. The peak discharge frequency response of cells receiving a polysynaptic ADN input (PSNs) was correlated to neither the absolute increase in blood pressure nor the rate of increase in mean arterial pressure. Diverse changes in discharge rate were observed during nitroprusside-induced reductions in blood pressure: an increase (n = 3), a decrease (n = 10), an increase followed by a decrease (n = 3) and no response (n = 6). Reductions in pressure of 64 +/- 2 mmHg produced weak reductions in spontaneous discharge of 1.3 +/- 0.9 Hz and only totally abolished spontaneous discharge in one neurone. These response patterns of NTS neurones during changes in arterial pressure suggest that baroreceptor inputs are integrated differently in MSNs compared to PSNs. The sensitivity of MSNs to the rate of change of pressure provides a mechanism for the rapid regulation of cardiovascular function. The lack of sensitivity to the mean level of a pressure increase in both MSNs and PSNs suggests that steady-state changes in pressure are encoded by the number of active neurones and not graded changes in the discharge of individual neurones. Both MSNs and PSNs receive tonic excitatory inputs from the arterial baroreceptors; however, these tonic inputs appear to be insufficient to totally account for their spontaneous discharge. PMID- 11101653 TI - Modulation of synaptic transmission from segmental afferents by spontaneous activity of dorsal horn spinal neurones in the cat. AB - We examined, in the anaesthetised cat, the influence of the neuronal ensembles producing spontaneous negative cord dorsum potentials (nCDPs) on segmental pathways mediating primary afferent depolarisation (PAD) of cutaneous and group I muscle afferents and on Ia monosynaptic activation of spinal motoneurones. The intraspinal distribution of the field potentials associated with the spontaneous nCDPs indicated that the neuronal ensembles involved in the generation of these potentials were located in the dorsal horn of lumbar segments, in the same region of termination of low-threshold cutaneous afferents. During the occurrence of spontaneous nCDPs, transmission from low-threshold cutaneous afferents to second order neurones in laminae III-VI, as well as transmission along pathways mediating PAD of cutaneous and Ib afferents, was facilitated. PAD of Ia afferents was instead inhibited. Monosynaptic reflexes of flexors and extensors were facilitated during the spontaneous nCDPs. The magnitude of the facilitation was proportional to the amplitude of the 'conditioning' spontaneous nCDPs. This led to a high positive correlation between amplitude fluctuations of spontaneous nCDPs and fluctuations of monosynaptic reflexes. Stimulation of low-threshold cutaneous afferents transiently reduced the probability of occurrence of spontaneous nCDPs as well as the fluctuations of monosynaptic reflexes. It is concluded that the spontaneous nCDPs were produced by the activation of a population of dorsal horn neurones that shared the same functional pathways and involved the same set of neurones as those responding monosynaptically to stimulation of large cutaneous afferents. The spontaneous activity of these neurones was probably the main cause of the fluctuations of the monosynaptic reflexes observed under anaesthesia and could provide a dynamic linkage between segmental sensory and motor pathways. PMID- 11101654 TI - Paired transcranial magnetic stimulation protocols reveal a pattern of inhibition and facilitation in the human parietal cortex. AB - Intracortical inhibition (ICI) and facilitation (ICF) of the human motor cortex can be induced by paired transcranial magnetic stimulation (TMS). Although demonstrated in experimental animals, the existence of intracortical inhibitory and excitatory circuits in parietal sensory cortex has not been documented in humans. The aim of this study was to investigate the effects of paired TMS of the parietal cortex on contralateral tactile perception. Fifteen healthy subjects were involved in a task of discrimination of electrical stimuli delivered at near threshold intensity of sensory perception over the left thumb. Paired TMS was delivered with a focal coil on the right posterior parietal lobe after various delays from the presentation of finger stimuli. The effects of different interstimulus intervals (ISI: 1, 3, 5, 7, 10 and 15 1 1 Bms1B) between the conditioning and the test TMS stimulus on tactile perception were studied. The conditioning stimulus intensity was set at 70 % of motor threshold, while test TMS intensity was 130 % of motor threshold. Single pulse suprathreshold TMS interfered with the perception of finger stimuli, while subthreshold stimuli such as the 'conditioning' stimuli had no effect on sensory perception. Paired TMS differentially influenced the performance depending on the ISI. At an ISI of 1 1 1 Bms1B, paired TMS stimuli induced a significant worsening of the performance compared with single pulse TMS; at an ISI of 5 1 1 Bms1B, paired TMS stimuli induced a significant facilitation of the performance compared with single pulse TMS, restoring baseline performance levels. These results suggest that paired TMS can reveal a selective pattern of ICI and ICF in the human parietal cortex. PMID- 11101655 TI - Human cone photoreceptor responses measured by the electroretinogram [correction of electoretinogram] a-wave during and after exposure to intense illumination. AB - We recorded the a-wave of the electroretinogram from human subjects with normal vision, using a corneal fibre electrode and ganzfeld stimulation under photopic conditions, so as to extract the parameters of cone phototransduction. The amplitude of bright flash responses provided a measure of the massed circulating current of the cones, while the amplitude of dim flash responses provided a measure of the product of the fraction of cone photopigment present, and the amplification constant of transduction within the cones. In the presence of steady background illumination, the cone circulating current declined to half at 3000 photopic trolands, and to a quarter at 20 000 photopic trolands. At very early times after the delivery of a near-total bleach, we could not determine the level of circulating current as our bright flashes did not appear to saturate the a-wave (presumably because so little pigment was present). However, by 20-30 s after a total bleach, the cone circulating current had returned to its dark adapted level. Following smaller bleaches (when ca 50 % of the pigment remained present) the bright flashes were able to saturate the a-wave even at very early times. Within 3 s of extinction of the illumination, the cone circulating current had returned to its dark-adapted level. This is at least a factor of 300 times faster than the period of ca 15 min required for full recovery of rods exposed to the same level of bleach, and indicates a major difference between rods and cones in the way that they cope with the photoproducts of bleaching. Despite the very rapid recovery of circulating current after bleaches, the recovery of dim-flash sensitivity was much slower, with a time constant of ca 1.5 min after a near total bleach. This time course is very similar to previous measurements of the regeneration of cone photopigment, and it seems highly probable that the reduction in dim-flash sensitivity results from pigment depletion. PMID- 11101656 TI - Accommodation to depolarizing and hyperpolarizing currents in cutaneous afferents of the human median and sural nerves. AB - To determine whether accommodation to depolarizing and hyperpolarizing stimuli differs for cutaneous afferents in the median and sural nerves, studies were performed in normal human subjects using threshold electrotonus. The changes in threshold for compound sensory action potentials of 50 % of maximum were recorded when the nerves were subjected to long-lasting depolarizing and hyperpolarizing DC. The premise was that the threshold changes largely mirror the underlying electrotonic changes in membrane potential. The maximal threshold changes produced by depolarizing and hyperpolarizing currents were greater for median afferents, suggesting that the DC produced greater changes in membrane potential in these afferents. Median afferents underwent greater accommodation to depolarizing currents than sural afferents and a greater threshold undershoot at the end of the currents, suggesting greater activity of a slow K+ conductance. Median afferents also underwent greater accommodation to hyperpolarizing currents, suggesting greater inward rectification. These conductances are voltage dependent, and the differences in accommodation could be due to greater changes in membrane potential for the median nerve. The changes in threshold produced by long-lasting depolarizing and hyperpolarizing currents of graded intensity were therefore measured. When the threshold changes were matched for the two nerves, median afferents underwent 22.4 % more accommodation to depolarizing currents and 28.7 % more accommodation to hyperpolarizing currents. We conclude that there is greater expression of two internodally located conductances responsible for accommodation on median afferents. The biophysical differences identified in this study might contribute to the finding that sural afferents have a greater tendency to dysfunction than median afferents. PMID- 11101657 TI - Fatiguing inspiratory muscle work causes reflex sympathetic activation in humans. AB - We tested the hypothesis that reflexes arising from working respiratory muscle can elicit increases in sympathetic vasoconstrictor outflow to limb skeletal muscle, in seven healthy human subjects at rest. We measured muscle sympathetic nerve activity (MSNA) with intraneural electrodes in the peroneal nerve while the subject inspired (primarily with the diaphragm) against resistance, with mouth pressure (PM) equal to 60 % of maximal, a prolonged duty cycle (TI/TTot) of 0.70, breathing frequency (fb) of 15 breaths min-1 and tidal volume (VT) equivalent to twice eupnoea. This protocol was known to reduce diaphragm blood flow and cause fatigue. MSNA was unchanged during the first 1-2 min but then increased over time, to 77 +/- 51 % (s.d.) greater than control at exhaustion (mean time, 7 +/- 3 min). Mean arterial blood pressure (+12 mmHg) and heart rate (+27 beats min-1) also increased. When the VT, fb and TI/TTot of these trials were mimicked with no added resistance, neither MSNA nor arterial blood pressure increased. MSNA and arterial blood pressure also did not change in response to two types of increased central respiratory motor output that did not produce fatigue: (a) high inspiratory flow rate and fb without added resistance; or (b) high inspiratory effort against resistance with PM of 95 % maximal, TI/TTot of 0.35 and fb of 12 breaths min-1. The heart rate increased by 5-16 beats min-1 during these trials. Thus, in the absence of any effect of increased central respiratory motor output per se on limb MSNA, we attributed the time-dependent increase in MSNA during high resistance, prolonged duty cycle breathing to a reflex arising from a diaphragm that was accumulating metabolic end products in the face of high force output plus compromised blood flow. PMID- 11101658 TI - Sensory integration in the perception of movements at the human metacarpophalangeal joint. AB - These experiments were designed to investigate illusions of movements of the fingers produced by combined feedback from muscle spindle receptors and receptors located in different regions of the skin of the hand. Vibration (100 Hz) applied in cyclic bursts (4 s 'on', 4 s 'off') over the tendons of the finger extensors of the right wrist produced illusions of flexion-extension of the fingers. Cutaneous receptors were activated by local skin stretch and electrical stimulation. Illusory movements at the metacarpophalangeal (MCP) joints were measured from voluntary matching movements made with the left hand. Localised stretch of the dorsal skin over specific MCP joints altered vibration-induced illusions in 8/10 subjects. For the group, this combined stimulation produced movement illusions at MCP joints under, adjacent to, and two joints away from the stretched region of skin that were 176 +/- 33, 122 +/- 9 and 67 +/- 11 % of the size of those from vibration alone, respectively. Innocuous electrical stimulation over the same skin regions, but not at the digit tips, also 'focused' the sensation of movement to the stimulated digit. Stretch of the dorsal skin and compression of the ventral skin around one MCP joint altered the vibration induced illusions in all subjects. The illusions became more focused, being 295 +/- 57, 116 +/- 18 and 65 +/- 7 % of the corresponding vibration-induced illusions at MCP joints that were under, adjacent to, and two joints away from the stimulated regions of skin, respectively. These results show that feedback from cutaneous and muscle spindle receptors is continuously integrated for the perception of finger movements. The contribution from the skin was not simply a general facilitation of sensations produced by muscle receptors but, when the appropriate regions of skin were stimulated, movement illusions were focused to the joint under the stimulated skin. One role for cutaneous feedback from the hand may be to help identify which finger joint is moving. PMID- 11101659 TI - Identification of potential sigma(N)-dependent promoters in bacterial genomes. PMID- 11101660 TI - Fimbrial surface display systems in bacteria: from vaccines to random libraries. PMID- 11101661 TI - Cooperative, synergistic and antagonistic haemolytic interactions between haemolysin BL, phosphatidylcholine phospholipase C and sphingomyelinase from Bacillus cereus. AB - Haemolysis of erythrocytes from different species (sheep, bovine, swine and human), caused by various combinations of phosphatidylcholine (PC)-preferring phospholipase C (PC-PLC), sphingomyelinase (SMase) and the three-component, pore forming toxin haemolysin BL (HBL) from Bacillus cereus was analysed. The lytic potency of HBL did not correlate with phospholipid (PL) content, but lysis by the individual or combined enzymes did. SMase alone lysed ruminant erythrocytes, which contain 46-53% sphingomyelin (SM). The cooperative action of PC-PLC and SMase was needed to lyse swine and human erythrocytes (22-31% PC and 28-25% SM). SMase synergistically enhanced haemolysis caused by HBL for all erythrocytes tested, which all contained >25% SM. PC-PLC enhanced HBL haemolysis only in cells containing significant amounts of PC (swine, 22% PC; human, 31% PC). Unexpectedly, PC-PLC inhibited HBL lysis of sheep erythrocytes (<2% PC) and enhanced the discontinuous haemolysis pattern that is characteristic of HBL in sheep blood agar. Inhibition and pattern enhancement was abolished by washing PC PLC-treated erythrocytes or by adding EDTA, suggesting that enzymic alteration of the membrane is not involved, but that zinc in the active site is required, perhaps to facilitate binding. These observations highlight the potential for cooperative and synergistic interactions among virulence factors in B. cereus infections and dependence of these effects on tissue composition. PMID- 11101662 TI - Mutations in the ss subunit of the Bacillus subtilis RNA polymerase that confer both rifampicin resistance and hypersensitivity to NusG. AB - Mutations conferring resistance to the antibiotic rifampicin (Rif(r)) occur at specific sites within the ss subunit of the prokaryotic RNA polymerase. Rif(r) mutants of Escherichia coli are frequently altered in the elongation and termination of transcription. Rif(r) rpoB mutations were isolated in Bacillus subtilis and their effects on transcription elongation factor NusG and Rho dependent termination were investigated. RNase protection assay, Northern analysis and the expression of nusG-lacZ fusions in cells with an inducible NusG suggested the B. subtilis nusG gene was autoregulated at the level of transcription. Rif(r) mutations that changed residue Q469 to a basic residue (Q469K and Q469R) enhanced autoregulation of nusG. A mutant expressing a truncated form of NusG, due to a nonsense mutation within the nusG gene, was isolated on the basis of the loss of autoregulation. The mechanism of autoregulation was found to be independent both of transcription termination factor Rho and of the promoter transcribing nusG. Autoregulation required sequences within the 5' coding sequence of the nusG gene or immediately upstream. This is the first evidence from any bacterium that Rif(r) RNA polymerases can display altered transcription regulation by NusG. PMID- 11101663 TI - The aprE leader is a determinant of extreme mRNA stability in Bacillus subtilis. AB - The Bacillus subtilis aprE gene encodes subtilisin, an extracellular proteolytic enzyme produced in stationary phase. The authors examined the stability of aprE mRNA and aprE leader-lacZ fusion mRNA. Both mRNAs were found to be unusually stable, with half-lives longer than 25 min, demonstrating that the aprE leader contains a determinant for extreme mRNA stability. The half-lives were the same in growing and stationary-phase cells. This contrasts with the findings of O. Resnekov et al. (1990) [Proc Natl Acad Sci USA 87, 8355-8359], which suggested a growth-phase-dependent mechanism for decay of aprE mRNA. The discrepancy is explained by the techniques used. Substitution of two bases or deletion of 25 nucleotides in the aprE leader led to a major difference in its predicted secondary structure and resulted in a fivefold reduction of the half-life of aprE mRNA. The authors also determined the half-life of amyE mRNA, which encodes alpha amylase, another stationary-phase, excreted enzyme and found it to be around 5 min. This shows that extreme stability is not a general property of stationary phase mRNAs encoding excreted enzymes. PMID- 11101664 TI - Bacillus subtilis guanine deaminase is encoded by the yknA gene and is induced during growth with purines as the nitrogen source. AB - Bacillus subtilis can utilize the purine bases adenine, hypoxanthine and xanthine as nitrogen sources. The utilization of guanine as a nitrogen source is reported here. The first step is the deamination of guanine to xanthine catalysed by guanine deaminase (GDEase). To isolate mutants defective in GDEase activity, a collection of mutant strains was screened for strains unable to use guanine as a nitrogen source. The strain BFA1819 (yknA) showed the expected phenotype and no GDEase activity could be detected in this strain. A new name for yknA, namely gde, is proposed. The gde gene encodes a 156 amino acid polypeptide and was preceded by a promoter sequence that is recognized by the sigma(A) form of RNA polymerase. High levels of GDEase were found in cells grown with purines and intermediary compounds of the purine catabolic pathway as nitrogen sources. Allantoic acid, most likely, is a low molecular mass inducer molecule. The level of GDEase was found to be subjected to global nitrogen control exerted by the GlnA/TnrA-dependent signalling pathway. The two regulatory proteins of this pathway, TnrA and GlnR, indirectly and positively affected gde expression. This is the first instance of a gene whose expression is positively regulated by GlnR. The GDEase amino acid sequence shows no homology with the mammalian enzyme. In agreement with this are the different physiological roles for the two enzymes. PMID- 11101665 TI - Gene transfer to Clostridium cellulolyticum ATCC 35319. AB - Although much is known about the bacterial cellulosome and its various protein components, their contributions to bacterial growth on cellulose and the process of cellulolysis in vivo cannot currently be assessed. To remedy this, the authors have developed gene transfer techniques for Clostridium cellulolyticum ATCC 35319. Firstly, transfer of Tn1545 has been obtained using an Enterococcus faecalis donor. Secondly, IncP-mediated conjugative mobilization of plasmids from Escherichia coli donors has also been achieved. The yield of transconjugants in both cases was low and was probably limited by the suboptimal growth conditions that must of necessity be employed for the co-culture of oligotrophic C. cellulolyticum with copiotrophic donors. A restriction endonuclease was detected in crude extracts of C. cellulolyticum. This enzyme, named CCE:I, is an isoschizomer of MSP:I (HPA:II). Electro-transformation was employed to establish plasmids containing the replication functions of pAMss1 (En. faecalis), pIM13 (Bacillus subtilis), pCB102 (Clostridium butyricum), pIP404 (Clostridium perfringens) and pWV01 (Lactococcus lactis subsp. cremoris) in C. cellulolyticum. Transformants were only obtained if the DNA was appropriately methylated on the external C of the sequence 5'-CCGG-3' using either BSU:FI methylase in vivo or MSP:I methylase in vitro. Plasmids based on the pAMss1 and pIM13 replicons were more stably maintained than one based on the pCB102 replicon. Selection of transformants on solid medium led to low apparent transformation efficiencies (approx. 10(2) transformants per microg DNA) which might, in part, reflect the low plating efficiency of the organism. Selection of transformants in liquid medium led to a higher apparent yield of transformants (between 10(5) and 10(7) transformants per microg DNA). The methods developed here will pave the way for functional analysis of the various cellulosome components in vivo. PMID- 11101666 TI - The potential for intraspecific horizontal gene exchange by natural genetic transformation: sexual isolation among genomovars of Pseudomonas stutzeri. AB - The potential for natural genetic transformation among the seven genomovars (gvs) of Pseudomonas stutzeri was investigated. Of the 12 strains originating from a variety of environments, six strains (50%) from five gvs were competent for DNA uptake (Rif(R) marker). The transformation frequencies varied over more than three orders of magnitude. With three highly transformable strains (ATCC 17587, ATCC 17641, JM300) from two gvs and all other strains as DNA donors, sexual isolation from other pseudomonad species (Pseudomonas alcaligenes, Pseudomonas mendocina) and also from other P. stutzeri gvs was observed (i.e. heterogamic transformation was reduced). For ATCC 17587 (gv 2) and ATCC 17641 (gv 8), heterogamic transformation was up to two and three orders of magnitude lower with other P. stutzeri gv and the other species employed, respectively, than in homogamic transformations. Interestingly, whereas with ATCC 17587 and ATCC 17641 heterogamic transformation with donors of the same gv was as high as homogamic transformation, JM300 (gv 8) was sexually isolated from its nearest relative (ATCC 17641). Also, sexual isolation of JM300 from other P. stutzeri gvs was most pronounced among the recipients tested, in some cases reaching the highest levels found with the other species as DNA donors (reduction of heterogamic transformation by 4000-fold). Results obtained here from nucleotide sequence analysis of part (422 nt) of the gene for the RNA polymerase ss subunit (rpoB) from various strains indicated that sexual isolation of ATCC 17641 increased with nucleotide sequence divergence. Implications of the observed great heterogeneity in transformability, competence levels and sexual isolation among strains are discussed with regard to the evolution of P. stutzeri. PMID- 11101667 TI - Molecular characterization of the mycobacterial SenX3-RegX3 two-component system: evidence for autoregulation. AB - Environmental regulation of bacterial gene expression is often mediated by two component signal transduction systems, which are themselves tightly regulated. The response regulator RegX3 and the cytoplasmic portion of the histidine kinase SenX3 from Mycobacterium bovis BCG were overproduced in Escherichia coli and purified as N-terminally (His)(6)-tagged proteins. Phosphorylation assays demonstrated autophosphorylation of the cytoplasmic portion of SenX3 and a phosphotransfer from SenX3 to RegX3, involving conserved histidine and aspartate residues, respectively. In addition, as shown by electrophoretic mobility shift assays, (His)(6)RegX3 was able to specifically bind to the promoter region of the senX3-regX3 operon. Furthermore, operon fusion analyses indicated that the overexpression of the senX3-regX3 operon increases the activity of the senX3 promoter in Mycobacterium smegmatis. Together, these results indicate that the mycobacterial SenX3-RegX3 two-component system is positively autoregulated. PMID- 11101668 TI - Prochlorococcus marinus strain PCC 9511, a picoplanktonic cyanobacterium, synthesizes the smallest urease. AB - The urease from the picoplanktonic oceanic Prochlorococcus marinus sp. strain PCC 9511 was purified 900-fold to a specific activity of 94.6 micromol urea min(-1) (mg protein)(-1) by heat treatment and liquid chromatography methods. The enzyme, with a molecular mass of 168 kDa as determined by gel filtration, is the smallest urease known to date. Three different subunits with apparent molecular masses of 11 kDa (gamma or UreA; predicted molecular mass 11 kDa), 13 kDa (ss or UreB; predicted molecular mass 12 kDa) and 63 kDa (alpha or UreC; predicted molecular mass 62 kDa) were detected in the native enzyme, suggesting a quaternary structure of (alphassgamma)(2). The K:(m) of the purified enzyme was determined as being 0.23 mM urea. The urease activity was inhibited by HgCl(2), acetohydroxamic acid and EDTA but neither by boric acid nor by L-methionine-DL sulfoximine. Degenerate primers were designed to amplify a conserved region of the ureC gene. The amplification product was then used as a probe to clone a 5.7 kbp fragment of the P. marinus sp. strain PCC 9511 genome. The nucleotide sequence of this DNA fragment revealed two divergently orientated gene clusters, ureDABC and ureEFG, encoding the urease subunits, UreA, UreB and UreC, and the urease accessory molecules UreD, UreE, UreF and UreG. A putative NtcA-binding site was found upstream from ureEFG, indicating that this gene cluster might be under nitrogen control. PMID- 11101669 TI - The microaerophilic flagellate Giardia intestinalis: oxygen and its reaction products collapse membrane potential and cause cytotoxicity. AB - Trophozoites of the microaerophilic flagellate parasitic protozoon Giardia intestinalis have only a limited capacity to detoxify O(2). Thus, when exposed to controlled concentrations of dissolved O(2) >8 microM, they gradually lose their ability to scavenge O(2). In a washed cell suspension stirred under 10% air in N(2) (equivalent to 25 microM O(2)), inactivation of the O(2)-consuming system was complete after 3.5 h; during this period accumulation of H(2)O(2) (3 micromol per 10(6) organisms) and oxidation of cellular thiols to 16% of their initial level occurred. Under 20% air (50 microM O(2)), respiratory inactivation was complete after 1.5 h, and under air (258 microM O(2)), after 50 min. Loss of O(2) consuming capacity was accompanied by loss of motility. Use of the fluorogen 2, 7 dichlorodihydrofluorescein acetate indicated that intracellular H(2)O(2) is produced at extranuclear sites. Flow cytometric estimation of the plasma membrane electrochemical potentials using bis(1,3-dibutylbarbituric acid) trimethine oxonol, DiBAC(4)(3), showed that values declined from -134 mV to -20 mV after 4.5 h aeration. Incubation of organisms with 60 microM H(2)O(2) for 10 min gave partial collapse of plasma membrane potential and complete loss of O(2) uptake capacity; motility and viability as assessed by DiBAC(4)(3) exclusion were completely lost after 1 h. Inactivation of the O(2)-consuming system and loss of viability were also observed on exposure to singlet oxygen photochemically generated from rose bengal or toluidine blue. PMID- 11101670 TI - The microaerophilic flagellate Giardia intestinalis: Allium sativum (garlic) is an effective antigiardial. AB - Whole garlic (Allium sativum L.) extract and some of its components were assayed for antigiardial activity. Whole garlic extract gave an IC(50) at 24 h of 0.3 mg ml(-1). Most of the components assayed were inhibitory to the organism, especially allyl alcohol and allyl mercaptan, with IC(50) values of 7 microg ml( 1) and 37 microg ml(-1) respectively. Studies with calcofluor white indicated that whole garlic and allyl alcohol collapse the transmembrane electrochemical membrane potential (Deltapsi) of the organism, as indicated by uptake of the fluorochrome. Electron microscopy allowed the morphological changes that occur with garlic inhibition to be recorded. Both the surface topography and internal architecture of the organism changed during incubation with the biocides. Both whole garlic and allyl alcohol resulted in fragmentation of the disc and an overexpression of disc microribbons, internalization of flagella, vacuole formation and an increase in distended vesicles. Allyl mercaptan, however, only gave an increase in distended vesicles, suggesting that this biocide has a different mode of action. PMID- 11101671 TI - Antibacterial activity of synthetic analogues based on the disaccharide structure of moenomycin, an inhibitor of bacterial transglycosylase. AB - Moenomycin is a natural product glycolipid that inhibits the growth of a broad spectrum of Gram-positive bacteria. In Escherichia coli, moenomycin inhibits peptidoglycan synthesis at the transglycosylation stage, causes accumulation of cell-wall intermediates, and leads to lysis and cell death. However, unlike Esc. coli, where 5-6 log units of killing are observed, 0-2 log units of killing occurred when Gram-positive bacteria were treated with similar multiples of the MIC. In addition, bulk peptidoglycan synthesis in intact Gram-positive cells was resistant to the effects of moenomycin. In contrast, synthetic disaccharides based on the moenomycin disaccharide core structure were identified that were bactericidal to Gram-positive bacteria, inhibited cell-wall synthesis in intact cells, and were active on both sensitive and vancomycin-resistant enterococci. These disaccharide analogues do not inhibit the formation of N:-acetylglucosamine ss-1, 4-MurNAc-pentapeptide-pyrophosphoryl-undecaprenol (lipid II), but do inhibit the polymerization of lipid II into peptidoglycan in Esc. coli. In addition, cell growth was required for bactericidal activity. The data indicate that synthetic disaccharide analogues of moenomycin inhibit cell-wall synthesis at the transglycosylation stage, and that their activity on Gram-positive bacteria differs from moenomycin due to differential targeting of the transglycosylation process. Inhibition of the transglycosylation process represents a promising approach to the design of new antibacterial agents active on drug-resistant bacteria. PMID- 11101672 TI - A new broad-spectrum protease inhibitor from the entomopathogenic bacterium Photorhabdus luminescens. AB - A new protease inhibitor was purified to apparent homogeneity from a culture medium of Photorhabdus luminescens by ammonium sulfate precipitation and preparative isoelectric focusing followed by affinity chromatography. Ph. luminescens, a bacterium symbiotically associated with the insect-parasitic nematode Heterorhabditis bacteriophora, exists in two morphologically distinguishable phases (primary and secondary). It appears that only the secondary-phase bacterium produces this protease inhibitor. The protease inhibitor has an M:(r) of approximately 12000 as determined by SDS-PAGE. Its activity is stable over a pH range of 3.5-11 and at temperatures below 50 degrees C. The N-terminal 16 amino acids of the protease inhibitor were determined as STGIVTFKND(X)GEDIV and have a very high sequence homology with the N-terminal region of an endogenous inhibitor (IA-1) from the fruiting bodies of an edible mushroom, Pleurotus ostreatus. The purified protease inhibitor inactivated the homologous protease with an almost 1:1 stoichiometry. It also inhibited proteases from a related insect-nematode-symbiotic bacterium, Xenorhabdus nematophila. Interestingly, when present at a molar ratio of 5 to 1, this new protease inhibitor completely inactivated the activity of both trypsin and elastase. The activity of proteinase A and cathepsin G was partially inhibited by this bacterial protease inhibitor, but it had no effect on chymotrypsin, subtilisin, thermolysin and cathepsins B and D. The newly isolated protease inhibitor from the secondary-phase bacteria and its specific inhibition of its own protease provides an explanation as to why previous investigators failed to detect the presence of protease activity in the secondary-phase bacteria. The functional implications of the protease inhibitor are also discussed in relation to the physiology of nematode-symbiotic bacteria. PMID- 11101673 TI - Temperature regulation of protease in Pseudomonas fluorescens LS107d2 by an ECF sigma factor and a transmembrane activator. AB - The production of extracellular enzymes by Pseudomonas fluorescens is important with respect to phytopathogenesis and, in the case of psychrotrophic strains, food spoilage. The production of extracellular protease has been previously reported to be dependent on temperature in psychrotrophic strains of P. fluorescens; production is decreased above the optimum growth temperature with a relatively small change in growth rate. In this work, a transposon mutant of P. fluorescens LS107d2 has been isolated which, in contrast to the wild-type strain, is completely protease deficient at 29 degrees C, above the optimum growth temperature of 25 degrees C, but which produces protease at 23 degrees C. Further analysis revealed that this mutation is in a gene (prtR) which is part of a dicistronic operon, prtIR, in which the two genes are translationally coupled. Evidence is presented that prtI encodes a sigma factor related to others involved in extracytoplasmic functions (ECF sigma factors) and that prtR encodes a novel transmembrane activator of PrtI. PrtI, like PrtR, is also required for protease production at 29 degrees C but not at 23 degrees C. Analysis of the amino acid sequence of PrtR indicates that it is functionally related to a group of membrane associated anti-sigma factors and a few transmembrane regulators, but is not significantly sequence related. Complementation analysis indicates that PrtR may also interact with sigma factors other than PrtI. The promoter region of the protease-encoding gene (aprX) in LS107d2 has been identified and has sequence features which could indicate interaction with either an ECF sigma factor or a primary sigma factor. PMID- 11101674 TI - YeiL, the third member of the CRP-FNR family in Escherichia coli. AB - The yeiL open reading frame located at 48.5 min (2254 kb) in the nfo-fruA region of the Escherichia coli chromosome was predicted to encode a CRP and FNR paralogue capable of forming inter- or intra-molecular disulphide bonds and incorporating one iron-sulphur centre per 25 kDa subunit. Purified MBP-YeiL (a maltose-binding-protein-YeiL fusion protein) was a high-molecular-mass oligomer or aggregate which released unstable monomers (68 kDa) under reducing conditions. The MBP-YeiL protein contained substoichiometric amounts of iron and acid-labile sulphide, and an average of one disulphide bond per monomer. The iron and sulphide contents increased consistent with the acquisition of one [4Fe-4S] cluster per monomer after anaerobic NifS-catalysed reconstitution. By analogy with FNR and FLP (the FNR-like protein of Lactobacillus casei) it was suggested that the transcription-regulatory activity of YeiL might be modulated by a sensory iron-sulphur cluster and/or by reversible disulphide bond formation. A yeiL-lacZ transcriptional fusion showed that aerobic yeiL expression increases at least sixfold during stationary phase, requires RpoS, and is positively autoregulated by YeiL, positively activated by Lrp (and IHF in the absence of FNR) and negatively regulated by FNR. A regulatory link between the synthesis of YeiK (a potential nucleoside hydrolase) and YeiL was inferred by showing that the yeiK and yeiL genes are divergently transcribed from overlapping promoters. A 10 15% deficiency in aerobic growth yield and an enhanced loss of viability under nitrogen starvation conditions were detected with a yeiL::kan(R) mutant, suggesting that YeiL might function as a post-exponential-phase nitrogen starvation regulator. PMID- 11101675 TI - A method for direct cloning of fur-regulated genes: identification of seven new fur-regulated loci in Escherichia coli. AB - A strain that allows the cloning of Fur-regulated loci was constructed. The strain, named FUR-SEL1, contains a chromosomal fhuA'-'cat transcriptional fusion that is expressed from the Fur-regulated promoter, fhuA(p). Therefore, Fur boxes introduced on a multicopy plasmid can cause derepression of the fusion by titrating the Fur repressor and thereby confer chloramphenicol resistance, which can be used as a selectable phenotype for cloning Fur-regulated loci. However, a number of additional mutations had to be introduced before FUR-SEL1 could be used for cloning Fur-regulated genes. The principal approach consisted of introducing a leaky fur mutation that ensures a more than 10(6)-fold increase in chloramphenicol resistance for FUR-SEL1 transformants carrying FUR-box-containing plasmids. To verify that the cloning procedure selects Fur-regulated genes, 10 recombinant plasmids chosen at random among the ones selected with FUR-SEL1 were analysed by FURTA (Fur-titration assay), a method for identification of Fur regulated genes. In addition, the nucleotide sequences of their chromosomal inserts were determined. Besides known Fur-regulated genes, seven Escherichia coli loci which have not previously been shown to be Fur regulated were found, including the pgmA and nrdHIEF genes, predicted ORF yhhY and four promoters identified first in this study. Three of the promoters preceded the nohA gene, and ORFs ygaC and yhhX. The fourth was located upstream of orf78 predicted in this work. The regulation of the promoter activities by iron and the involvement of Fur in this regulation were shown. Employing FUR-SEL1 for cloning Fur regulated loci from other enterobacteria is discussed. PMID- 11101676 TI - Genomic survey of cAMP and cGMP signalling components in the cyanobacterium Synechocystis PCC 6803. AB - Cyanobacteria modulate intracellular levels of cAMP and cGMP in response to environmental conditions (light, nutrients and pH). In an attempt to identify components of the cAMP and cGMP signalling pathways in Synechocystis PCC 6803, the authors screened its complete genome sequence by using bioinformatic tools and data from sequence-function studies performed on both eukaryotic and prokaryotic cAMP/cGMP-dependent proteins. Sll1624 and Slr2100 were tentatively assigned as being two putative cyclic nucleotide phosphodiesterases. Five proteins were identified as having all the determinants required to be cyclic nucleotide receptors, two of them being probably more specific for cGMP (an element of two-component regulatory systems - Slr2104 - and a putative cyclic nucleotide-gated cation channel - Slr1575), the three others being probably more specific for cAMP: (i) a protein of unidentified function (Slr0842); (ii) a putative cyclic-nucleotide-modulated permease (Slr0593), previously annotated as a kinase A regulatory subunit; and (iii) a putative transcription factor (CRP SYN: =Sll1371), which possesses cAMP- and DNA-binding determinants homologous to those of the cAMP receptor protein of Escherichia coli (CRP-EC:). This homology, together with the presence in Synechocystis of CRP-EC:-like binding sites upstream of crp, cya1, slr1575, and several genes encoding enzymes involved in transport and metabolism, strongly suggests that CRP-SYN: is a global regulator. PMID- 11101677 TI - The Oenococcus oeni genome: physical and genetic mapping of strain GM and comparison with the genome of a 'divergent' strain, PSU-1. AB - The physical and genetic maps of the Oenococcus oeni strains GM and PSU-1, which represent two genomic divergent groups on the basis of macrorestriction and ribotyping analysis, were compared. To achieve this comparison, the GM maps were constructed and the PSU-1 maps, already established, were improved. All the recognition sites of the restriction enzymes ASC:I, I-CEU:I, FSE:I, NOT:I and SFI:I were located in both chromosomes and the position of 26 genetic markers, including two rrn operons and 14 new putative oenococcal genes, were allocated to the restriction fragments generated by the five enzymes. The comparative analysis of O. oeni GM and PSU-1 genomes revealed extensive conservation of loci order. As for the differences encountered in the locations of restriction sites, they seem to be a reflection of the differences in restriction fragment sizes, explainable by insertion/deletion events and point mutations. No evidence for major genomic rearrangements was found. The genomic conservation between the two strains is in agreement and suggests homogeneity within the species, which was not unexpected in view of the restricted ecological niche of O. oeni. Further comparisons of physical maps, both of O. oeni strains and related species, will certainly help to assess whether O. oeni is really an homogeneous species and physical mapping is suitable for taxonomic purposes, both at the supra- and intraspecific levels. PMID- 11101678 TI - Comparison of the proteome of Mycobacterium tuberculosis strain H37Rv with clinical isolate CDC 1551. AB - The genome sequences of two virulent strains of Mycobacterium tuberculosis (H37Rv and CDC 1551) are now available. CDC 1551 is a recent clinical isolate and H37Rv is a commonly used lab strain which has been subject to in vitro passage. The two strains have been shown to display differing phenotypes both in vivo and in vitro. The proteome of the two strains grown in liquid culture were examined over time to determine whether there are any major differences between them at the protein level and the differences were compared to the genome data. Total cell lysates of the two strains were analysed by two-dimensional electrophoresis. Approximately 1750 protein spots were visualized by silver staining and the protein profiles of the two strains were found to be highly similar. Out of a total of 17 protein spot differences, seven were unique to CDC 1551 and three to H37Rv. Two further spots showed increased intensity in H37Rv, one spot showed differing vertical mobility between the strains and four showed differing spot intensities with time. Twelve of the spot differences were identified using mass spectrometry; however, no obvious association with phenotype could be deduced. When genome differences were analysed and related to the proteome differences, a mobility shift identified in the MoxR protein could be explained by a point mutation at the gene level. This proteome analysis reveals that, despite having been maintained under vastly different conditions, namely in vitro passage and in vivo transmission, these two strains have remained highly similar. PMID- 11101679 TI - Differential cytokine expression in avian cells in response to invasion by Salmonella typhimurium, Salmonella enteritidis and Salmonella gallinarum. AB - Salmonella enterica is a facultative intracellular pathogen that is capable of causing disease in a range of hosts. Although human salmonellosis is frequently associated with consumption of contaminated poultry and eggs, and the serotypes Salmonella gallinarum and Salmonella pullorum are important world-wide pathogens of poultry, little is understood of the mechanisms of pathogenesis of Salmonella in the chicken. Type III secretion systems play a key role in host cell invasiveness and trigger the production of pro-inflammatory cytokines during invasion of mammalian hosts. This results in a polymorphonuclear cell influx that contributes to the resulting enteritis. In this study, a chicken primary cell culture model was used to investigate the cytokine responses to entry by the broad host range serotypes S. enteritidis and S. typhimurium, and the host specific serotype S. gallinarum, which rarely causes disease outside its main host, the chicken. The cytokines interleukin (IL)-1ss, IL-2, IL-6 and interferon (IFN)-gamma were measured by quantitative RT-PCR, and production of IL-6 and IFN gamma was also determined through bioassays. All serotypes were invasive and had little effect on the production of IFN-gamma compared with non-infected cells; S. enteritidis invasion caused a slight down-regulation of IL-2 production. For IL 1ss production, infection with S. typhimurium had little effect, whilst infection with S. gallinarum or S. enteritidis caused a reduction in IL-1ss mRNA levels. Invasion of S. typhimurium and S. enteritidis caused an eight- to tenfold increase in production of the pro-inflammatory cytokine IL-6, whilst invasion by S. gallinarum caused no increase. These findings correlate with the pathogenesis of Salmonella in poultry. S. typhimurium and S. enteritidis invasion produces a strong inflammatory response, that may limit the spread of Salmonella largely to the gut, whilst S. gallinarum does not induce an inflammatory response and may not be limited by the immune system, leading to the severe systemic disease fowl typhoid. PMID- 11101680 TI - Invasiveness in chickens, stress resistance and RpoS status of wild-type Salmonella enterica subsp. enterica serovar typhimurium definitive type 104 and serovar enteritidis phage type 4 strains. AB - The heat and acid resistance and the ability to survive airdrying on commonly used kitchen surfaces were assessed for clinical and environmental strains of Salmonella enterica subsp. enterica serovar Typhimurium, definitive type (DT) 104. Three out of thirty-eight strains of DT 104 were found to be more sensitive in stationary phase to the stresses examined than the other strains. This compares to a previous study by the authors which showed that seven out of forty serovar Enteritidis phage type (PT) 4 strains were more sensitive. RpoS activity was examined indirectly in selected strains of DT 104 and PT 4. In those with normal stress resistance a 100-fold induction of an RpoS-dependent spvR/A:'::luxCDABE fusion was observed upon entry into stationary phase. The sensitive strains examined showed either no induction or a reduced level of spvR/A:'::luxCDABE expression. The rpoS gene was sequenced from these strains and three were found to harbour mutations including one deletion, one base-pair substitution resulting in a nonsense codon, and one insertion causing a frameshift resulting in an early stop codon. Strains with negligible or reduced spvR/A:'::luxCDABE expression had low stress resistance. All strains of DT 104 could be recovered from liver and spleen tissues of infected hens 14 d post infection, but one with no induction of spvR/A:'::luxCDABE expression was significantly less likely to be recovered from chicken reproductive tissues, liver or spleen than the majority of other strains, including one with reduced spvR/A:'::luxCDABE expression. This work has demonstrated that clinical and environmental strains of DT 104 and PT 4 not infrequently harbour mutations in the rpoS allele. It is possible that the rpoS mutations may have occurred during the initial isolation of the strains. The ability of a strain to cause infection, however, also depends on factors such as host susceptibility and dose. PMID- 11101681 TI - How Delisea pulchra furanones affect quorum sensing and swarming motility in Serratia liquefaciens MG1. AB - Halogenated furanones produced by the benthic marine macroalga Delisea pulchra inhibit swarming motility of Serratia liquefaciens MG1. This study demonstrates that exogenously added furanones control transcription of the quorum sensing regulated gene swrA in competition with the cognate signal molecule N:-butanoyl-L homoserine lactone. This in turn results in reduced production of the surface active compound serrawettin W2, which is crucial for surface translocation of the differentiated swarm cells. It is demonstrated that furanones interfere with interspecies communication during swarming of mixed cultures and that the mode of interference in quorum-sensing control and interspecies communication is not through inhibition of autoinducer synthesis. PMID- 11101682 TI - Continuous monitoring of the cytoplasmic pH in Methanobacterium thermoautotrophicum using the intracellular factor F(420) as indicator. AB - The absorption spectrum of factor F(420) changes depending on the pH and the redox state of the cytoplasm. Specific wavelengths were used to calibrate absorption changes to allow the measurement of changes in the cytoplasmic pH in Methanobacterium thermoautotrophicum. Upon a hydrogen pulse, a rapid efflux of protons was observed. Under these energized conditions, the DeltapH amounts to 0.2-0.4 pH units at pH 6.6, and 0.6-0.8 pH units at pH 6.0. It decays within 10 20 s. In parallel, a sodium gradient is formed which has a slightly longer lifetime. Both DeltapH and DeltaPsi contribute to the proton-motive force present during methanogenesis. The energy-conversion rate, as indicated by the decay of the energized state of the cell, is fastest under growth conditions, i.e. at pH 6.9 and at a temperature of 58 degrees C. PMID- 11101684 TI - PkwA, a WD-repeat protein, is expressed in spore-derived mycelium of Thermomonospora curvata and phosphorylation of its WD domain could act as a molecular switch. AB - WD-repeat proteins are found in all eukaryotes and are implicated in a variety of regulatory functions as a result of protein-protein interactions. PkwA from Thermomonospora curvata CCM3352 is a first potential example of a WD-repeat protein in a prokaryotic actinomycete. A mAb (3G2) was generated against the carboxy terminus of PkwA and was used to analyse the expression of PkwA in T. curvata. PkwA was detected in exponential growth phase following inoculation with spores, but could not be found at any stage of growth following inoculation with vegetative mycelium. PkwA and its WD domain were expressed in Escherichia coli as His-tag derivatives and purified on a Talon metal affinity matrix. The WD domain was phosphorylated by Pkg2, a membrane-spanning protein Ser/Thr kinase from 'Streptomyces granaticolor'. A membrane fraction from an exponential, spore derived culture of T. curvata was found to phosphorylate the WD domain specifically in the presence of Mn(2+). These data confirm that PkwA is expressed in spore-derived exponential growth phase of T. curvata and could play a role as a molecular switch in a signalling pathway. PMID- 11101683 TI - A functional water channel protein in the pathogenic bacterium Brucella abortus. AB - The gene for a new bacterial aquaporin, AqpX, was cloned from the pathogenic Gram negative bacterium Brucella abortus. The gene was mapped on the large chromosome of B. abortus. It is flanked by one upstream and two downstream copies of the Brucella repeated sequence Bru-RS. Prediction from the nucleotide sequence indicated that the protein is a member of the MIP family, which comprises channels for water and/or solute transport. Expression in Xenopus oocytes and cryoelectron microscopy of Escherichia coli cells transformed with the aqpX gene confirmed that the protein is an efficient water channel. Glycerol uptake experiments in E. coli also showed that the protein is not able to transport glycerol. PMID- 11101685 TI - The importance of the five phosphoribosyl-pyrophosphate synthetase (Prs) gene products of Saccharomyces cerevisiae in the maintenance of cell integrity and the subcellular localization of Prs1p. AB - Phosphoribosyl-pyrophosphate synthetase (Prs) catalyses the synthesis of phosphoribosyl pyrophosphate (PRPP), an intermediate in nucleotide metabolism and the biosynthesis of the amino acids histidine and tryptophan. The Saccharomyces cerevisiae genome contains a family of five PRS genes, PRS1-PRS5. Using anti peptide antisera directed against two different epitopes of Prs1p it was shown that Prs1p localizes to granular cytoplasmic structures. This localization was confirmed by living cell microscopy of strains expressing a functional green fluorescent protein (GFP)-tagged Prs1p. Analysis of Prs1p distribution in conditional secretory-deficient (sec) mutants suggested that the observed distribution of Prs1p is independent of the secretory pathway. Electron microscopy revealed that plasma membrane invaginations and accumulation of cytoplasmic vesicles were more frequent in strains which lack some of the PRS genes than in the wild-type. The fact that Deltaprs1 and Deltaprs3 are hypersensitive to caffeine and unable to recover from exposure to it as judged by the release of alkaline phosphatase points to a possible link between Prs and the maintenance of cell integrity. PMID- 11101686 TI - Landmarks in the early duplication cycles of Aspergillus fumigatus and Aspergillus nidulans: polarity, germ tube emergence and septation. AB - When the spores of filamentous fungi break dormancy, nuclear division is accompanied by a series of ordered morphological events including the switch from isotropic to polar growth, the emergence of a second germ tube from the conidium and septation. Correlation of these morphological events with nuclear number allows them to serve as duplication cycle landmarks. Early duplication cycle landmarks have been characterized in Aspergillus nidulans, but not in other filamentous fungi. To learn more about duplication cycle control in filamentous fungi, a study was undertaken to compare the timing of landmarks in Aspergillus fumigatus and A. nidulans. Nuclear duplication took approximately 45 min in A. fumigatus, with mitosis occupying roughly 5% of this period. Under the same conditions, nuclear duplication in A. nidulans took approximately 60 min, with mitosis occupying roughly 4% of this period. In A. fumigatus the isotropic to polar switch preceded the first mitosis in 22% of cells, while in A. nidulans the isotropic to polar switch did not occur until after the first mitosis. In both A. fumigatus and A. nidulans the earliest emergence of a second germ tube from the conidium occurred after the third mitotic division. However, by the fifth mitosis only 19% of A. fumigatus conidia had a second germ tube, compared to 98% of A. nidulans conidia. In both A. fumigatus and A. nidulans, formation of the first septum occurred after the fourth mitotic division. In all experiments a few cells lagged behind the others in nuclear number. In this delayed group, it was common to see landmark events at an earlier mitotic division. Differences in nuclear number when identical landmarks occur in A. fumigatus versus A. nidulans, and uncoupling of mitotic division and landmarks in delayed cells suggest that nuclear division and morphogenesis lie in parallel pathways, perhaps coordinated by checkpoints. PMID- 11101687 TI - Characterization of IS900 loci in mycobacterium avium subsp. paratuberculosis and development of multiplex PCR typing PMID- 11101688 TI - Molecular screening of the CFTR gene in men with anomalies of the vas deferens: identification of three novel mutations. AB - Many studies have shown that congenital absence of the vas deferens (CAVD) is a genital cystic fibrosis transmembrane conductance regulator (CFTR)-mediated phenotype, with a broad spectrum of abnormalities causing male infertility. The genotype of these patients includes mutations in the CFTR gene, e.g. DeltaDeltaF508, R117H and the T5 allele; all of which are commonly found in CAVD. In this study we have screened the entirety of CFTR gene in 47 males with anomalies of the vas deferens: 37 cases of congenital bilateral absence of the vas deferens, three cases of congenital unilateral absence of the vas deferens and seven cases of obstructive azoospermia with hypoplastic vas deferens. Among the 94 chromosomes studied, 65 mutations, of which three are novel (2789+2insA, L1227S, 4428insGA), were identified. The majority of patients (63.8%) had two detectable CFTR gene mutations. Furthermore, high frequencies of the DeltaDeltaF508 mutation (44.7%), the T5 allele (36.2%) and R117H mutation (19.1%) were observed. PMID- 11101689 TI - Expression and characterization of the human YWK-II gene, encoding a sperm membrane protein related to the alzheimer betaA4-amyloid precursorprotein. AB - The YWK-II cDNA, RSD-2, encoding a sperm membrane protein was isolated from a rat testis cDNA expression library. Using the RSD-2 insert in combination with rapid amplification of cDNA ends (RACE), the corresponding human gene was isolated from a human testis cDNA expression library. The human testis cDNA, HSD-2, is 3654 bp in length and contains an open reading frame of 763 codons. Hydropathicity analysis showed that the deduced polypeptide is a single strand transmembrane protein. The deduced polypeptide has partial homology with the amyloid precursor protein (APP) and high homology with the amyloid precursor homologue, APLP2/APPH. The YWK-II gene was mapped and assigned to human chromosome locus: 11q24-25. Northern blotting of various human tissue RNAs using the HSD-2 cDNA as a probe showed that the gene is transcribed ubiquitously. The cytoplasmic domain of HSD-2 was expressed in Escherichia coli. In-vitro studies showed that the recombinant polypeptide bound to a GTP-binding protein (G(o)) and was phosphorylated by protein kinase C and cdc2 kinase. In mammalian F11 cells, the recombinant polypeptide was found to be coupled to G(o). Thus, the YWK-II component has the characteristics of a G(o)-coupled receptor and may be involved in G(o)-mediated signal transduction pathway. Protein kinase C and cdc2 kinase may regulate this pathway in spermatozoa by phosphorylating the cytoplasmic domain of the YWK-II component. PMID- 11101690 TI - High concentrations of inhibin A and inhibin B in ovarian serous cystadenoma: relationship with oestradiol and nitric oxide metabolites. AB - Inhibin production has been demonstrated in malignant epithelial ovarian tumours, but secretion of inhibins by benign cystadenoma has not yet been reported. The present study evaluated the concentrations of inhibin A and inhibin B and the relationship with oestradiol and nitric oxide metabolites in fluid collected from benign ovarian serous cystadenomas (n = 15). In addition, follicular fluid samples (n = 14) from women with regular ovulatory cycles undergoing ovarian stimulation for IVF were studied as a reference group. High concentrations of inhibin A (median = 89.3 ng/ml) and inhibin B (median = 116.1 ng/ml) were found in the cystic fluid of ovarian serous cystadenomas. These inhibin concentrations were even higher than in follicular fluid of stimulated follicles (inhibins A and B = 41.2 and 46.8 ng/ml respectively; P: < 0.001), whereas oestradiol was approximately 18-fold lower in cystic fluid than in follicular fluid (median = 34 versus 622 pg/ml, P: < 0.001). In ovarian cysts, the concentrations of inhibin A and oestradiol were inversely correlated (r = -0.678, P: = 0.008). Cystic fluid samples containing the highest concentrations of NO(2)(-)/NO(3)(-) (45-60 micromol/l) had lower inhibin A and higher oestradiol concentrations than those samples containing lower concentrations (10-25 micromol/l) of NO(2)(-)/NO(3)(-). It is concluded that high amounts of dimeric inhibins are present in ovarian serous cystadenoma. The source of inhibins and the determinants of the inverse association of inhibin A with oestradiol and nitric oxide remain to be determined. PMID- 11101691 TI - Eosinophils in the human corpus luteum: the role of RANTES and eotaxin in eosinophil attraction into periovulatory structures. AB - We evaluated the presence and number of eosinophils at varying stages in the human corpus luteum from 27 ovaries of women at reproductive age. Eosinophils preferentially accumulated in dilated microvessels of the thecal layer transforming into septa of the corpus luteum. The granulosa layer under luteinization, the thecal layer, and haemorrhages in the former antrum each contained low, moderate and high numbers of extravasated eosinophils respectively. Eosinophils decreased rapidly during the stages of secretion and regression. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT PCR) and enzyme-linked immunosorbent assay (ELISA) systems were used to investigate the expression and regulation of the eosinophil-attracting chemokines RANTES (regulated on activation, normal T cell expressed and secreted) and eotaxin in granulosa cells obtained from follicular aspirates from women undergoing IVF. Contaminating leukocytes were determined by CD18 mRNA quantification. Granulosa cells expressed RANTES (n = 3; 43 +/- 14 pg/ml, mean +/ SEM). 4ss-phorbol-12-myristate-13-acetate (PMA; 211 +/- 53) and tumour necrosis factor alpha (TNFalpha) (238 +/- 59), but not interleukin (IL)-1 up-regulated RANTES at significant levels. In general, higher basal and stimulated RANTES mRNA and protein were found in cultures with higher CD18 mRNA levels than in those with lower levels. We found only traces of eotaxin mRNA and no eotaxin secretion, even in stimulated granulosa cell cultures, independently of leukocyte levels. Taken together, this is the first study demonstrating the selective presence of eosinophils in human periovulatory structures. RANTES, but not eotaxin, may play an active process in the accumulation of these cells. PMID- 11101692 TI - Localization of connective tissue growth factor in human uterine tissues. AB - Connective tissue growth factor (CTGF) is a recently described heparin-binding mitogen for fibroblasts and smooth muscle cells. The aim of this study was to investigate the production of CTGF by human uterine tissues using immunohistochemical and Northern blotting analyses. For immunohistochemistry, formalin-fixed human proliferative (n = 5), early secretory (n = 5; days 15-19), mid-secretory (n = 5; days 20-23), late secretory (n = 5; days 24-28) endometrial, and decidual (n = 5) tissues were stained using a highly specific affinity-purified polyclonal antibody raised against residues 81-94 of human CTGF. Myometrial (n = 5) and leiomyoma (n = 5) tissues were also used for CTGF immunochemistry. In proliferative endometrium, epithelial and vascular endothelial cells showed strong CTGF immunoreactivity, whereas stromal cells were negative or only weakly positive for the CTGF protein. Throughout the entire secretory stage, CTGF was detected in epithelial and endothelial cells of endometrium. Stromal cells showed strong immunoreactivity to CTGF only in oedematous areas for early and mid-secretory endometrium, and in decidualized regions of late secretory endometrium. During pregnancy, the decidual, epithelial and endothelial cells of the endometrium were all immunoreactive to CTGF. In myometrial and leiomyoma samples, CTGF immunoreactivity was found only in the endothelial cells. Northern blotting of mRNA from normal uterus (n = 2) or leiomyoma (n = 6) using a 320 bp human CTGF cDNA probe revealed a single 2.4 kb transcript. This study is the first to demonstrate CTGF gene expression and localization of its encoded protein in human uterine tissues. The cell- and cycle specific localization of CTGF support a role for this molecule in regulating aspects of uterine cell growth, migration, and/or matrix production during the menstrual cycle and pregnancy. PMID- 11101693 TI - In-vivo transfection of the female reproductive tract epithelium. AB - Mouse female genital tract was transfected in vivo using the ss-galactosidase reporter gene. To transfect the female tract, DNA/liposome complexes were injected through the infundibulum of the oviducts of adult, immature, and pseudopregnant females. Females which were in different stages of the ovarian cycle were also employed. Transfection was analysed using histochemical, immunological and molecular (Southern blotting, polymerase chain reaction and gene sequencing) procedures. The lower region of the uterine glands and the oviduct epithelium in the isthmus and juncture regions were the most conspicuous transfected areas. The greatest numbers of transfected cells were 6% in the oviduct and 9% in the uterus, meanwhile the duration of expression reached a maximum of 7 days in the oviduct and 14 days in the uterus. The hormonal stage of the genital tract epithelium directly affected transfection, as the highest number of successful transfections occurred during the meta-oestrus and pseudopregnancy stages. PMID- 11101694 TI - Inhibin and activin subunits are differentially expressed in endometrial cells and leukocytes during the menstrual cycle, in early pregnancy and in women using progestin-only contraception. AB - Inhibins and activins are dimeric hormones which share common subunits and which have diverse endocrine and paracrine roles in regulating reproductive function. Endometrial expression of inhibin alpha, ssA and ssB subunits was examined by immunohistochemistry and in-situ hybridization, across the menstrual cycle and in early pregnancy. All three subunits were found to be expressed in endometrium, primarily by glandular epithelium in the early stages of the cycle. Following the onset of decidualization, expression of alpha, ssA and ssB subunits was up regulated in decidualized stromal cells. A marked down-regulation of alpha subunit was detected in glandular epithelium, whilst expression of ssA and ssB subunits was maintained. This pattern was consistent in decidua from early pregnancy and additionally in endometrium from women using progestin-only contraceptives, either subdermal implants (Norplant((R))) or levonorgestrel releasing intrauterine systems (Lng-IUS). Immunostaining was also observed for both ssA and ssB subunits in subpopulations of endometrial leukocytes, identified to be distinct subsets of macrophages, neutrophils and mast cells. Potential paracrine roles for activins may be envisaged in facilitating tissue remodelling during decidualization, in tissue repair following menstruation, and additionally in modulating premenstrual inflammatory events. PMID- 11101695 TI - Calcium influx in human uterine epithelial RL95-2 cells triggers adhesiveness for trophoblast-like cells. Model studies on signalling events during embryo implantation. AB - RL95-2 is a human uterine epithelial cell line that exhibits adhesion competence on its apical surface for trophoblast-like JAR cells. Using confocal microscopy and an adhesion assay we have found that changes in intracellular free calcium ([Ca(2+)](i)) in RL95-2 cells are involved in binding of JAR spheroids. Impact of spheroids upon, and movement of spheroids across, monolayers of RL95-2 cells produced a transient increase in [Ca(2+)](i). Pretreatment of RL95-2 cells with the Ca(2+) channel inhibitor, diltiazem, reduced the [Ca(2+)](i) increase. Interestingly, resting of JAR spheroids on RL95-2 cells caused no detectable alterations in [Ca(2+)](i) although cell-cell bonds were formed during prolonged contact. However, separation of established bonds did produce an increase in [Ca(2+)](i) which could be reduced by the Ca(2+) channel blocker, SKF-96365, but not by diltiazem. SKF-96365 also reduced adhesion of JAR spheroids to RL95-2 cells. In all experiments, the increase in [Ca(2+)](i) was due to influx from the external medium, as it could be blocked both by removing extracellular Ca(2+) and by nickel. These results suggest that the plasma membrane of uterine RL95-2 cells contains two types of Ca(2+) channels that are involved in trophoblast adhesion, i.e. diltiazem-sensitive channels contributing to initiation of JAR cell binding and SKF-96365-sensitive channels participating in a feedback loop that controls the balance of bonds. PMID- 11101696 TI - Uterine expression of alternatively spliced mRNAs of mouse splicing factor SC35 during early pregnancy. AB - RNA differential display was applied to identify genes critical for the establishment of pregnancy in the mouse. One of the gene fragments identified was homologous to human SC35 splicing factor; the mouse counterpart had not then been cloned. To obtain the full cDNA sequence of the mouse gene, a cDNA library was screened and four positive clones were fully analysed. Sequencing analysis indicated that we had cloned alternatively spliced mRNA species of mouse SC35 splicing factor. A map of splicing structure for this gene's pre-mRNA was then proposed and region-specific mRNA species were tested on Northern blots. This analysis indicated that the overall expression level of SC35 mRNA was much higher in implantation sites than in inter-implantation sites in the mouse uterus during early pregnancy. The expression of alternatively spliced mRNAs for SC35 was differently regulated both during early pregnancy and by steroid hormones. Embryo derived factors were also implicated in the up-regulation of SC35 mRNA at implantation sites. These results demonstrate, for the first time, that an essential splicing factor is regulated in a complex manner during implantation in the mouse uterus. Hence, its correct regulation could be important for the success of pregnancy. PMID- 11101697 TI - Application of a functional genomics approach to identify differentially expressed genes in human myometrium during pregnancy and labour. AB - The molecular mechanisms regulating uterine relaxation and contraction during pregnancy are poorly understood. In the present study, we used for the first time a functional genomics approach applying gene array technology to identify novel candidate genes involved in the regulation of uterine quiescence and contractility during pregnancy. The purpose of this approach was to obtain a molecular snapshot of the expression profile of gene transcripts as a function of the time dependent process regulating myometrial quiescence. Using this approach, we found several genes whose expression in human myometrium was altered with the onset of labour. For example, the expression of insulin-like growth factor (IGF) II, calgranulin A and B, and G-protein coupled receptor were decreased while the expression of IGF-binding proteins, Ca(2+)/CaM binding protein kinase C substrate, and angiotensin converting enzyme were increased in the labouring, compared with non-labouring, pregnant myometrium. The differentially-expressed genes include several genes whose roles in myometrial quiescence are yet to be understood, although they have been reported to regulate vascular smooth muscle tone. Our findings illustrate the advantage of a functional genomics approach over a single gene analysis in identifying a large number of novel and potentially important genes mediating uterine smooth muscle contractile activity. PMID- 11101698 TI - The expression of glutaredoxin is increased in the human cervix in term pregnancy and immediately post-partum, particularly after prostaglandin-induced delivery. AB - Glutaredoxins are glutathione disulphide oxidoreductases catalysing disulphide reductions via a redox active disulphide. We have examined the presence of glutaredoxin in the human cervix, and its differential expression during cervical remodelling in term pregnancy and immediately post-partum as compared to the non pregnant state. Cervical biopsies were obtained from 24 term-pregnant and 24 post partal women, of which 10 were taken after spontaneous delivery, 10 after prostaglandin-induced delivery and four after mifepristone-induced delivery, all obtained within 15 min after delivery. Six non-pregnant women served as controls. The tissues were analysed for the glutaredoxin mRNA levels using a solution hybridization method. Glutaredoxin mRNA was expressed in the human cervix, the level increased > or =2-fold at term pregnancy and immediately post-partum. The level of cervical glutaredoxin mRNA from prostaglandin E(2)-treated women was 3 fold higher than after spontaneous ripening and delivery. Localization of glutaredoxin was visualized with immunohistochemistry in cervices from two post partal women, and was compared to that of thioredoxin. We conclude that glutaredoxin may be involved in the regulation of cervical ripening in humans, particularly in the inflammatory reaction seen during this process. Glutaredoxin mRNA levels are up-regulated after prostaglandin treatment, which is effective and the most commonly used substance for cervical priming and induction of labour. PMID- 11101699 TI - Real-time PCR using molecular beacons for accurate detection of the Y chromosome in single human blastomeres. AB - We describe a highly accurate method for determining the sex of human embryos via real-time polymerase chain reaction (PCR) amplification of highly-conserved, moderately-repeated sequences within the TSPY genes on the Y chromosome and the U2 genes on chromosome 17. Individual male lymphocytes, female lymphocytes, and blastomeres from donated cleavage-stage embryos were lysed prior to PCR using an optimized buffer containing proteinase K. Molecular beacons, a new type of fluorescent probe, were used to detect and quantify accumulating amplicons during each cycle of PCR carried out in closed tubes. The present work is part of an ongoing study to construct and implement a new, convenient and reliable system of preimplantation genetic diagnosis (PGD). PMID- 11101700 TI - Preimplantation genetic diagnosis for medium-chain acyl-CoA dehydrogenase (MCAD) deficiency. AB - Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most common defect in fatty acid oxidation. The disease is inherited in an autosomal recessive fashion (carrier frequency around 1 in 70) and probably affects as many as 1 in 10000 new-borns. Affected children usually present within the two first years of life with recurrent episodes of hypoketotic hypoglycaemia and lethargy leading to death in approximately 25% of the cases. One mutation (c985A-->G) accounts for approximately 90% of the carrier chromosomes. We developed a preimplantation genetic diagnosis (PGD) strategy for MCAD for a couple who had already lost two affected children. When tested on heterozygous lymphoblasts, the amplification efficiency was 67 out of 71 (94%) and the allele drop-out rate was 0 out of 67. The patient became pregnant after one PGD cycle during which two embryos were replaced. The twin pregnancy was checked by chorionic villus sampling (CVS) and was shown to be unaffected. The twins have been born and are healthy. PMID- 11101701 TI - Modern measurement for a modern health service. PMID- 11101702 TI - Evidence-based patient empowerment. PMID- 11101703 TI - Performance management at the crossroads in the NHS: don't go into the red. PMID- 11101704 TI - Voluntary reporting system in anaesthesia: is there a link between undesirable and critical events? AB - BACKGROUND: Reporting systems in anaesthesia have generally focused on critical events (including death) to trigger investigations of latent and active errors. The decrease in the rate of these critical events calls for a broader definition of significant anaesthetic events, such as hypotension and bradycardia, to monitor anaesthetic care. The association between merely undesirable events and critical events has not been established and needs to be investigated by voluntary reporting systems. OBJECTIVES: To establish whether undesirable anaesthetic events are correlated with critical events in anaesthetic voluntary reporting systems. METHODS: As part of a quality improvement project, a systematic reporting system was implemented for monitoring 32 events during elective surgery in our hospital in 1996. The events were classified according to severity (critical/undesirable) and nature (process/outcome) and control charts and logistic regression were used to analyse the data. RESULTS: During a period of 30 months 22% of the 6439 procedures were associated with anaesthetic events, 15% of which were critical and 31% process related. A strong association was found between critical outcome events and critical process events (OR 11.5 (95% confidence interval (CI) 4.4 to 27.8)), undesirable outcome events (OR 4.8 (95% CI 2.0 to 11.8)), and undesirable process events (OR 4.8 (95% CI 1.3 to 13.4)). For other classes of events, risk factors were related to the course of anaesthesia (duration, occurrence of other events) and included factors determined during the pre-anaesthetic visit (risk of haemorrhage, difficult intubation or allergic reaction). CONCLUSION: Undesirable events are associated with more severe events and with pre-anaesthetic risk factors. The way in which information on significant events can be used is discussed, including better use of preoperative information, reduction in the collection of redundant information, and more structured reporting. PMID- 11101705 TI - Validation of a questionnaire measuring patient satisfaction with general practitioner services. AB - BACKGROUND: In order that patient satisfaction may be assessed in a meaningful way, measures that are valid and reliable are required. This study was undertaken to assess the construct validity and internal reliability of the previously developed Patient Satisfaction Questionnaire (PSQ). METHOD: A total of 1390 patients from five practices in the North of England, the Midlands, and Scotland completed the questionnaire. Responses were checked for construct validity (including confirmatory factor analysis to check the factor structure of the scale) and internal reliability. RESULTS: Confirmatory factor analysis showed that items loaded on the appropriate factors in a five factor model (doctors, nurses, access, appointments, and facilities). Scores on the specific subscales showed highly significant positive correlations with general satisfaction subscale scores suggesting construct validity. Also, the prediction (derived from past research) that older people would be more satisfied with the service was borne out by the results (F (4, 1312) = 57.10; p < 0.0001), providing further construct validation. The five specific subscales (doctors, nurses, access, appointments, and facilities), the general satisfaction subscale, and the questionnaire as a whole were found to have high internal reliability (Cronbach's alpha = 0.74-0.95). CONCLUSION: The results suggest that the PSQ is a valid and internally reliable tool for assessing patient satisfaction with general practitioner services. PMID- 11101706 TI - Errors in health care management: what do they cost? AB - BACKGROUND: Iatrogenic injuries are relatively common and a potentially avoidable source of morbidity. The economic evaluation of this area has been limited by the lack of good quality national data to provide an estimate of incidence, associated disability, and preventability of iatrogenic injuries. Two recent surveys, the Quality in Australian Health Care Study (QAHCS) and the Utah Colorado Study (UTCOS), have now made this feasible. AIMS: To determine the direct costs associated with iatrogenic injuries occurring in a hospital setting. METHODS: The QAHCS was used as a representative national source of information on the incidence, disability, and preventability of iatrogenic injuries. Costs were calculated using information from Australian disease related groups (AN-DRGs) relative to the injury categories. RESULTS: The cost of just 12 preventable iatrogenic injuries is significant (0.25 million US dollars) and accounts for 2 3% of the annual budget of a typical Australian community based hospital of 120 beds. Costing data provide additional useful information for policy and decision makers. CONCLUSION: Costing iatrogenic injuries is an important component of the impact of these events. An ongoing national database of iatrogenic injuries is necessary to assist in identifying the incidence of these injuries, monitoring trends, and providing data for cost estimates and economic evaluations. PMID- 11101707 TI - New Zealand and United Kingdom experiences with the RAND modified Delphi approach to producing angina and heart failure criteria for quality assessment in general practice. AB - OBJECTIVES: (1) To describe the development of minimum review criteria for the general practice management in New Zealand (NZ) of two chronic diseases: stable angina and systolic heart failure, and (2) to compare the NZ angina criteria with a set produced in Manchester to assess the extent to which use of the same approach to criteria development yields similar criteria. METHODS: A modified Delphi approach, based on the RAND consensus panel method, was used to produce minimum criteria for reviewing the recorded management of heart failure and angina in NZ general practice. The criteria for angina were compared with those produced in the UK, including assessment of the extent to which each set describes actions that the other panel agrees are necessary to record. RESULTS: For each condition we report minimum criteria describing actions rated as (a) necessary to record and (b) inappropriate to take but, if taken, necessary to record. Although strong scientific evidence underpins approximately one quarter and one third, respectively, of the final sets of NZ and UK angina criteria for actions necessary to record, the NZ criteria agree strongly with the UK criteria (33 of 39 criteria, 85%) but there is less UK agreement with the NZ angina criteria (28 of 40 criteria, 70%). CONCLUSION: Despite the lack of scientific evidence for up to three quarters of angina care in general practice, the RAND based approach to criteria development was used in NZ to reproduce most of the UK angina criteria for actions rated as necessary to record in general practice. It is important to make explicit whether ratings of necessity and appropriateness apply to the recording of actions or to the actions themselves. PMID- 11101708 TI - What is a prescribing error? AB - OBJECTIVE: To develop a practitioner led definition of a prescribing error for use in quantitative studies of their incidence. DESIGN: Two stage Delphi technique. SUBJECTS: A panel of 34 UK judges, which included physicians, surgeons, pharmacists, nurses and risk managers. MAIN OUTCOME MEASURES: The extent to which judges agreed with a general definition of a prescribing error, and the extent to which they agreed that each of 42 scenarios represented a prescribing error. RESULTS: Responses were obtained from 30 (88%) of 34 judges in the first Delphi round, and from 26 (87%) of 30 in the second round. The general definition of a prescribing error was accepted. The panel reached consensus that 24 of the 42 scenarios should be included as prescribing errors and that five should be excluded. In general, transcription errors, failure to communicate essential information, and the use of drugs or doses inappropriate for the individual patient were considered prescribing errors; deviations from policies or guidelines were not. CONCLUSIONS: Health care professionals are in broad agreement about the types of events that should be included and excluded as prescribing errors. A general definition of a prescribing error has been developed, together with more detailed guidance regarding the types of events that should be included. This definition allows the comparison of prescribing error rates among different prescribing systems and different hospitals, and is suitable for use in both research and clinical governance initiatives. PMID- 11101709 TI - The potential use of decision analysis to support shared decision making in the face of uncertainty: the example of atrial fibrillation and warfarin anticoagulation. AB - The quality of patient care is dependent upon the quality of the multitude of decisions that are made daily in clinical practice. Increasingly, modern health care is seeking to pursue better decisions (including an emphasis on evidence based practice) and to engage patients more in decisions on their care. However, many treatment decisions are made in the face of clinical uncertainty and may be critically dependent upon patient preferences. This has led to attempts to develop decision support tools that enable patients and clinicians to make better decisions. One approach that may be of value is decision analysis, which seeks to create a rational framework for evaluating complex medical decisions and to provide a systematic way of integrating potential outcomes with probabilistic information such as that generated by randomised controlled trials of interventions. This paper describes decision analysis and discusses the potential of this approach with reference to the clinical decision as to whether to treat patients in atrial fibrillation with warfarin to reduce their risk of stroke. PMID- 11101710 TI - Psychosocial interventions for schizophrenia. PMID- 11101715 TI - Measuring Patient Outcomes.: Marie T Nolan and Victoria Mock (Pp 248; pound23.00). California: Sage Publications, 2000. 0 7619 1505 2. PMID- 11101714 TI - The Evidence-Based Primary Care Handbook.: Mark Gabbay, editor. (Pp 314; pound19.50). London: Royal Society of Medicine, 1999. 1 85315 415 6Quality in Health Care 2000;9:264-265. PMID- 11101711 TI - Complications of diabetes: renal disease and promotion of self-management. PMID- 11101716 TI - 35th anniversary of the Journal of Pediatric Surgery (1966-2001): a look into the future. PMID- 11101717 TI - Surfactant--a review for pediatric surgeons. PMID- 11101718 TI - A model of bacterial translocation in neuroblastoma-bearing mice. AB - PURPOSE: The aim of this study was to establish a model of bacterial translocation (BT) in neuroblastoma-bearing mice. METHODS: A suspension of 1 x 10(6) cells of the murine neuroblastoma cell line C1300 was injected subcutaneously into the thighs of 8-week-old female A/J mice, which were then killed after 7, 14, and 21 days. Some of the mice were given 1-microm or 2-microm fluorescein-labeled latex beads in their drinking water for 7 days before being killed. Mesenteric lymph nodes (MLNs) were aseptically removed and cultured for 72 hours at 37 degrees C. Segments of distal ileum were obtained for histologic examination. Samples of venous blood were obtained for laboratory tests. RESULTS: Tumors were found at the injection sites on days 14 and 21 after C1300 injection. Although tumors were not found in 7 days, significantly high number of 1-microm latex beads were detected in MLNs compared with the control, and the number increased with tumor growth. The number of 2-microm latex beads was significantly higher on days 14 and 21. The percentage of mice with MLN cultures positive were significantly higher on day 14, and the percentage increased along with tumor growth. On day 21 after C1300 injection, body weight loss and anemia were observed, and histologic findings of the terminal ileum showed mucosal edema and villous thinning. Serum levels of interleukin (IL)-6 were significantly higher in mice killed 14 and 21 days after injection. CONCLUSIONS: The results suggest that BT from the gut to MLNs may occur in neuroblastoma C1300-bearing mice, and it increases along with tumor growth. Even in the early stage of malignancy, particles as small as 1 microm may translocate from the gut to MLNs. PMID- 11101719 TI - Neonatal jaundice: the role of laparoscopy. AB - BACKGROUND: When managing neonatal jaundice, despite continual improvement of diagnostic tests and increasing knowledge regarding its pathogenesis, there is no single test or imaging modality that can reliably define biliary atresia. Early diagnosis is essential for a better surgical outcome. In many situations, mini laparotomy and operative cholangiography may be needed to settle the definitive diagnosis, with the risk of having negative exploration in those high-risk patients with medical etiology. The use of laparoscopy may help in avoiding unnecessary exploration for such group of patients. METHODS: Thirty-three patients aged between 1 and 4 months with conjugated hyperbilirubinemia were the subject for this study. All had a HIDA scan result suggestive of biliary atresia. They underwent a diagnostic laparoscopy before surgical exploration. When the gallbladder was not visualized we proceeded to laparotomy. In patients with a good size gallbladder visualized at laparoscopy, a laparoscopic-guided cholangiogram was then performed, and laparoscopic liver biopsy done for those who had patent biliary tree. RESULTS: Two groups of patients were identified: the first group (21 patients) showed small atretic gallbladder; 18 patients had biliary atresia with complete intra- and extrahepatic atresia; these patients underwent a Kasai hepatic-portoenterostomy. Two patients showed a patent gallbladder and common bile duct with atresia of the common hepatic and intrahepatic ducts, and they underwent a portocholecystostomy. The last patient showed left-sided gallbladder arising from the left lobe of the liver that was missed during laparoscopy, and operative cholangiogram showed hypoplastic biliary ducts. The second group included 12 patients with good-sized gallbladder, and laparoscopic-guided percutaneous cholangiogram showed normal communicating patent biliary system, hypoplastic in 2, and they underwent laparoscopic liver biopsy. No mortality related to the laparoscopic procedure was encountered in this series, and one patient with hypoplastic gallbladder had adhesive intestinal obstruction on the fifth day after laparoscopy necessitating exploration. CONCLUSION: Laparoscopy with laparoscopic-guided cholangiography may be a very useful tool used in accurately diagnosing infants with conjugated hyperbilirubinemia, and in avoiding unnecessary laparotomies performed on these critical babies. PMID- 11101720 TI - Surgical treatment of pulmonary hydatid disease in children: report of 122 cases. AB - BACKGROUND/PURPOSE: The aim of this study was to review the authors' surgical experience in pediatric pulmonary hydatid disease focusing on clinical presentation, parenchyma saving operations, and long-term results. METHODS: One hundred twenty-two children with pulmonary hydatid cyst were treated surgically over the last 2 decades and were reviewed retrospectively. There were 66 boys and 56 girls with a mean age of 9 years. RESULTS: Pulmonary hydatid cyst was seen in 111 (91%) patients and pulmonary and hepatic cysts in 11 (9%). Lateral thoracotomy was performed in 106 (87%) patients, thoracotomy and laparotomy in 6 (5%), median sternotomy in 5 (4%), lateral thoracotomy with phrenotomy in 4 (3%), and median sternotomy with phrenotomy in 1 (0.8%). Parenchyma-saving procedures were performed in 114 patients (93%) and lung resection in 8 (7%). There was no mortality. Postoperative complication was seen in 5 patients (4%). CONCLUSIONS: Parenchyma-saving procedures without capitonnage are preferable. In patients with right or bilateral lung and coexisting cysts in the upper part of the liver, thoracotomy or median sternotomy and transdiaphragmatic approach allows the surgeon to remove the lung and liver cysts in a single operation. Median sternotomy is an alternative method for the bilateral lung hydatidosis compared with sequential thoracotomy. PMID- 11101721 TI - Ultrasonographic diagnosis criteria using scoring for hypertrophic pyloric stenosis. AB - PURPOSE: For diagnosis of infantile hypertrophic pyloric stenosis (HPS), ultrasonography (US) is a useful and objective diagnostic method. In the current study, pyloric diameter, muscular thickness, and pyloric length were measured in normal infants (n = 26) and infants which is an adequate (n = 57). Each score was assigned to relevant measurements, and diagnostic criteria obtained with a scoring system were prepared using statistical skills by the probit analysis. METHODS: For scoring, points were given to relevant measurements in conformity with the probit analysis. Zero points were given to patients with no possibility of HPS, 1 point to those with less than 25% probability, 2 points to those with 25% or more but less than 50% probability, and 3 points to patients with 50% or more probability. Points were totaled, and analysis was performed. RESULTS: The composite score was evaluated by probit analysis, and cases with a composite score of 2 or less were all included in the normal group, whereas those with a composite score of 3 or more were all in the HPS group. Both groups could thereby be 100% identified. CONCLUSION: US was able to diagnose cases with overall score of 2 or less as normal and those with overall score of 3 or higher as having HPS. In addition, after the current diagnostic criteria were prepared, preoperative diagnoses were performed prospectively using them for vomiting neonates and infants, and all cases were correctly discriminated and diagnosed. These findings indicate our ultrasonographic diagnosis criteria are useful for diagnosing HPS. PMID- 11101722 TI - The contemporary outcome of gastroschisis. AB - BACKGROUND: The aim of this study was to evaluate the contemporary outcome in the management of gastroschisis. METHODS: A retrospective analysis was conducted of 91 babies admitted over a 7-year period to a single neonatal surgical unit with a diagnosis of gastroschisis. RESULTS: An antenatal diagnosis was made in 89 (98%) cases. Surgical intervention occurred in 90 babies, at a mean of 5 hours (range, 0.5 to 17) postdelivery. In 72 (80%) cases, primary closure of the abdominal defect was achieved, with a silo fashioned in the remaining 18 (20%). One child died before abdominal closure. The median time to full oral feeding was 30 days (range, 5 to 160 days), and to discharge was 42 days (range, 11 to 183 days). Those children who required a silo, took longer to feed (P =.008) and stayed longer in the hospital (P =.021). The 8 (8.8%) children with an intestinal atresia, required significantly more operative procedures (P =.0001) and took significantly longer to achieve full oral feeding (P =.04), but the presence of an atresia was not an independent risk factor for mortality. There were 7 deaths (7.7%), 3 within the first 7 days. Of the deaths, 5 (71%) were caused by overwhelming sepsis. CONCLUSIONS: The contemporary mortality rate from gastroschisis is less than 8%, and minimizing septic complications would contribute significantly to reducing this. Strategies designed to improve morbidity must focus on optimizing management of those factors associated with a prolonged recovery, namely intestinal atresia, prematurity, and the use of a silo. PMID- 11101723 TI - Rectal mucosal biopsy compared with laparoscopic seromuscular biopsy in the diagnosis of intestinal neuronal dysplasia in children with slow-transit constipation. AB - BACKGROUND/PURPOSE: Intestinal neuronal dysplasia (IND) as a cause for severe chronic constipation remains controversial. The aim of this study is to examine the correlation between a deficiency of substance P (SP) immunoreactive nerve fibers in the colon and enzyme histochemistry of rectal biopsies in children with slow-transit constipation. METHODS: Fifty children with intractable constipation have been assessed by rectal biopsies examined with histochemical staining for lactate dehydrogenase, and 32 children among those 50 have been studied by laparoscopic seromuscular biopsy of the colon labelled with antibodies to SP using immunofluorescence methods. RESULTS: Four children have evidence of IND. Fifteen children, including all 4 IND cases, showed a deficiency of SP immunoreactivity. There is a significant correlation between giant ganglia and SP deficiency (P <.01). CONCLUSION: This study is attempting to propose that a deficiency of SP immunoreactivity in colonic circular muscle nerves may be used as a histologic marker for slow-transit constipation and that IND may be a small subset of patients with SP deficiency. PMID- 11101724 TI - Allogeneic fetal small bowel graft in pigs treated with cyclosporin A. AB - BACKGROUND: The functional integrity of transplanted fetal intestine was proven in rodents. The authors examined the morphology and development of intraperitoneally transplanted fetal intestine under cyclosporin A (CsA) monotherapy in a large mammal. METHODS: Allogeneic fetal intestinal grafts were transplanted intraperitoneally in pigs. The graft was wrapped in omentum. Thirteen recipients received grafts harvested at 60 days of gestation and 5 at 105 days of gestation. All recipients received 25 mg/kg/d CsA. CsA blood levels were measured at the end of the study. The development of the grafts was assessed by inspection and histology studying revascularization, maturation, and immune rejection. RESULTS: All grafts developed neovascularization. The intestinal wall in the 105-day-old group was thick enough to lead to complete mucosal destruction, whereas the 60-day-old group showed viable mucosa. All grafts induced an immune rejection. This immune response was correlated with the CsA blood level. The graft was destroyed within 15 days when CsA trough level was below 70 ng/mL, had a subacute rejection with villi atrophy when CsA trough level ranged from 70 to 150 ng/mL, and had a good appearance in spite of mild blunting of villi when CsA trough level was over 150 ng/mL. CONCLUSION: Allogeneic fetal intestinal transplantation from 60-day-old embryos in pig achieved successful graft. PMID- 11101725 TI - A prospective study of botulinum toxin for internal anal sphincter hypertonicity in children with Hirschsprung's disease. AB - BACKGROUND: Internal anal sphincter hypertonicity with nonrelaxation can cause persistent constipation and obstructive symptoms in children after surgery for Hirschsprung's disease. Intractable symptoms traditionally have been treated with anal myectomy, which may be ineffective or complicated by long-term incontinence. The authors evaluated prospectively the use of intrasphincteric botulinum toxin for these patients. METHODS: Eighteen children were studied (age 1 to 13; median, 4 years). Botulinum toxin was injected (total dose 15 to 60 U) into 4 quadrants of the sphincter. Resting sphincter pressure was measured in 14 patients before and after injection. Ten have had 1 to 5 additional injections (total dose, 30 to 60 U per injection). RESULTS: Four patients had no improvement in bowel function, 2 had improvement for less than 1 month, 7 had improvement for 1 to 6 months, and 5 had improvement more than 6 months. Nine of those with symptomatic improvement longer than 1 month had pressures measured, with a documented decrease in 8. Five with no significant clinical improvement had pressure measurements, with a decrease in 3. There were no adverse effects associated with botulinum toxin injection. Four children had new encopresis postinjection, which was mild and resolved in each case. CONCLUSIONS: Intrasphincteric botulinum toxin is a safe and less-invasive alternative to myectomy for symptomatic internal sphincter hypertonicity. Persistent symptoms, despite a fall in sphincter pressure, suggest a nonsphincteric etiology. Repeat injections often are necessary for recurrent symptoms. PMID- 11101726 TI - Improved survival for patients with advanced neuroblastoma after high-dose combined chemotherapy based in part on N-myc amplification. AB - BACKGROUND/PURPOSE: In spite of many different kinds of chemotherapy for neuroblastoma, the prognosis for advanced neuroblastoma remains unsatisfactory. In particular, the outcome of advanced neuroblastoma with high copies of the N myc gene tend to be poor. Therefore, the new high-dosage combined chemotherapy regimens for advanced neuroblastoma based in part on the N-myc amplification status has been utilized in the Kyushu area of Japan since 1991. This study aims to investigate whether these new regimens based in part on N-myc amplification have improved the survival rate of stage III and stage IV patients in comparison with the old regimens. METHODS: Between 1983 and 1995, 77 patients over 1 year of age and with stage III or IV neuroblastoma were registered in the Kyushu Area. Between 1983 and 1990, 49 patients received 1 of 2 combined chemotherapy regimens consisting of cyclophosphamide, cisplatin plus VM-26, and Adriamycin plus DTIC. Since 1991, two new regimens (New A1 and A3) have been administered based on the N-myc amplification status in a total of 28 patients. The New A1 regimen, which consists of cyclophosphamide, cisplatin, Adriamycin, and VP-16 has been administered in cases of less than 10 copies of N-myc, whereas the A3 regimen, consisting of a higher dose of cyclophosphamide, cisplatin, Adriamycin, and VP 16, has been administered in cases of more than 10 copies of N-myc. The survival rate was then compared between the old regimens and the new regimens. RESULTS: The 3-year survival rate (61.5%) for patients treated by the new regimens was significantly higher than that (32.7%) for patients treated by the old regimens (P <.01). Regarding the 24 cases of more than 10 copies of N-myc, the 3-year survival rate (35.9%) of the 13 patients treated by the A3 regimen was higher than that (0%) of the 11 patients treated by the old regimens (P <.05). However, in the 19 stage IV patients treated by the new regimens, the 3-year survival rate (11.1%) of the 9 cases of more than 10 copies was significantly lower than that (77.8%) of the 10 cases of less than 10 copies of N-myc (P <.01). CONCLUSIONS: These results suggest that high-dose combined chemotherapy based in part on the N myc amplification status significantly improved the prognosis of patients with advanced neuroblastoma. However, stage IV patients with N-myc amplification still require a more effective treatment modality. PMID- 11101727 TI - Hydrostatic balloon dilation of congenital esophageal stenoses associated with esophageal atresia. AB - It has been stated that congenital cartilage rings in the esophagus do not respond to dilation and should be resected. The authors report on 3 infants with congenital esophageal stenoses who were treated successfully with hydrostatic balloon dilation. Based on the appearance during dilation the authors believe that these stenoses were cartilage rings. The technique is described in detail. Balloon dilation is the treatment of choice for these patients. Resection should be reserved for those who do not respond to this form of therapy. PMID- 11101728 TI - Risk of contralateral manifestation in children with unilateral inguinal hernia: should hernia in children be treated contralaterally? AB - PURPOSE: This study was done to identify risk factors for metachronous manifestation of contralateral inguinal hernia in patients with unilateral inguinal hernia. METHODS: Characteristics of 156 patients with metachronous contralateral hernia were compared with those of 156 patients with unilateral hernia who were ascertained not to have presented with contralateral hernia. RESULTS: There was a tendency for the hernia to be more often on the left side in 88 of 156 patients (56.4%) with contralateral manifestation compared with 70 of 156 patients (44.9%) in the control group (P =.054). The age at hernia repair of the patients with contralateral manifestation, 1 to 120 months (median, 14 months), was significantly younger than the 1 to 149 months (median, 20 months) of the control patients (P =.016). More patients with contralateral manifestation had a family history of inguinal hernia, and the percentage, 24.4%, was significantly higher than the 14.7% in the control group (P =.046). A univariate analysis with the Cox regression models found that hernia on the left side and a positive family history were significantly associated with the metachronous manifestation of contralateral hernia (hazard ratio [HR], 1.40; P =. 037 and HR, 1.59; P =.013, respectively). CONCLUSION: The risk of metachronous manifestation of contralateral hernia is high in patients with left-side hernia and in those with a family history, and the incidence of contralateral hernia is at most 10% in these patients. The authors think that the incidence is still too low to justify routine exploration and surgery for a patent processus vaginalis. Contralateral exploration should therefore be reserved for high-risk patients in whom second anesthesia and surgery have to be avoided. PMID- 11101729 TI - Coexpression of hypoxia-inducible factor 1-alpha and vascular endothelial growth factor in Wilms' tumor. AB - BACKGROUND/PURPOSE: Hypoxia-inducible factor 1 alpha (HIF-1alpha) is an important transcriptional factor responsible for regulating expression of the angiogenic cytokine, vascular endothelial growth factor (VEGF). Little information is available regarding factors involved in the hypoxic cascade, such as HIF or VEGF in Wilms' tumor. We concomitantly evaluate the expression of HIF-1alpha and VEGF in ex vivo human Wilms' tumor specimens. METHODS: Immunohistochemical analysis (IHC) utilizing a monoclonal human anti-HIF-1alpha or a polyclonal anti-VEGF antibody was performed on ex vivo specimens of Wilms' tumor (n = 18). Predominant tumor histologic subtype was divided equally between epithelial (n = 6), blastemal (n = 6), and mixed (n = 6). Specimens were scored on a predetermined scale for distribution (percent positive cells) and intensity of HIF-1alpha/VEGF expression within areas of tumor. RESULTS: IHC analysis found that HIF-1alpha and VEGF were expressed in all Wilms' tumor specimens. Strong nuclear staining for HIF-1alpha was seen in all samples evaluated, (n = 18), mean score 2.7 (>50% cells exhibiting nuclear HIF-1alpha expression). Cytoplasmic staining for HIF 1alpha also was seen in 15 of 18 samples (83%). Distribution of VEGF was equivalent between blastemal and epthelial components, mean score 2.23 versus 2.35. CONCLUSIONS: HIF-1alpha and one of its regulatory end-products, the angiogenic cytokine VEGF, are simultaneously expressed in human Wilms' tumor. In Wilms' tumor, intratumoral hypoxia may stimulate tumor conversion to the angiogenic phenotype and incite production of VEGF. Strategies targeting the hypoxic cascade ultimately may prove efficacious against Wilms' tumor. PMID- 11101730 TI - Presentation and management of epigastric hernias in children. AB - BACKGROUND/PURPOSE: Although epigastric hernias are common, there are no reports that describe the presentation and treatment of these defects in children. The authors reviewed their experience with these hernias to develop recommendations for their management in this age group. METHODS: Medical records were reviewed for all children younger than 18 years who presented for evaluation of an epigastric hernia at our institution over 14 years. Data on presentation, operative findings, and postoperative results were obtained. RESULTS: Forty children were evaluated for an epigastric hernia, representing 4% of all pediatric patients seen for treatment of a hernia. An epigastric hernia was first observed at birth in 12 patients (30%). All children presented with a mass in the epigastrium. The hernia was observed to be either symptomatic (abdominal wall pain or tenderness) or enlarging in 22 patients (55%). Thirty-eight children underwent repair, and 2 were lost to follow-up. There was no recurrence or morbidity associated with surgical repair of these defects. CONCLUSIONS: Epigastric hernias are common in children and frequently present in infancy. Because most are either symptomatic or enlarging, the authors recommend repair of these defects at the time of presentation. PMID- 11101731 TI - Prenatal dexamethasone rescues heart hypoplasia in fetal rats with congenital diaphragmatic hernia. AB - BACKGROUND/PURPOSE: Patients and rats with congenital diaphragmatic hernia (CDH) have lung and heart hypoplasia. Prenatal steroids improve lung hypoplasia in CDH rats. The current study tests the hypothesis that prenatal dexamethasone could rescue heart hypoplasia in rats with CDH. METHODS: Timed pregnant rats received intragastrically either 100 mg nitrofen or oil on day 9.5, and other animals had the same treatment with, in addition, either 0.25 mg/kg dexamethasone intraperitoneally or no treatment on days 19 and 20. Fetuses were recovered on day 21, and heart weight to body weight ratios, heart DNA, protein, and glycogen were measured in fresh specimens. Left-to-right ventricular diameter and aortic to-pulmonary diameter ratios were measured after formalin fixation. RESULTS: Wet heart weight to body weight, left-to-right ventricular diameter, and aortic-to pulmonary root diameter ratios, which were lower in fetuses exposed only to nitrofen than in their oil controls, were similar in those exposed to nitrofen plus dexamethasone than in their corresponding oil plus dexamethasone controls. Total heart DNA, which was decreased in fetuses exposed to nitrofen with CDH in comparison with their controls, was increased in those receiving nitrofen and dexamethasone in comparison with theirs. Protein to DNA ratio was decreased in all rats with CDH irrespective of their exposure or not to dexamethasone. Glycogen to DNA ratio was higher in all dexamethasone-treated fetuses than in those without this treatment. No gross histologic differences were seen among groups. CONCLUSIONS: Heart hypoplasia in rats with CDH is in part rescued by prenatal dexamethasone treatment as expressed by increased number of smaller myocytes with higher glycogen content. Prenatal steroids could modify heart involvement in human fetuses with CDH as well. PMID- 11101732 TI - Intrahepatic mast cell population correlates with clinical outcome in biliary atresia. AB - PURPOSE: To determine if mast cells influence the clinical outcome in biliary atresia (BA), the authors examined the intrahepatic mast cell population in BA. METHODS: Mast cells were identified histochemically using Toludin Blue and immunohistochemically using antimast cell tryptase antibody in formalin-fixed paraffin-embedded sections from 21 cases of BA. Patients were divided into 3 groups; group I (n = 8) with good liver function, group II (n = 8) with moderate liver dysfunction, and group III (n = 5) with severe liver dysfunction. Liver biopsies from patients with choledochal cysts (CDC, n = 5), and normal liver (NL, n = 4) served as controls. The results were compared among the groups. RESULTS: Both histochemical and immunohistochemical methods showed similar data. Mast cells were seen mostly in the portal tracts. Mast cell numbers per medium power field (20 x magnification) were higher in BA than in the controls (15. 03 +/- 2.25 v 3.85 +/-.65, [mean +/- SEM], P <.05, BA v CDC; 15.03 +/- 2.25 v 1.73 +/ .06, [mean +/- SEM], P <.05, BA v NL, immunohistochemical data). Clinical correlation showed an association between higher mast cell number and liver dysfunction (32.62 +/-.80 v 8.52 +/-.87 [mean +/- SEM], group III v group I; P <. 05, immunohistochemical data). CONCLUSION: Increased mast cell population in BA adversely affects liver function and raises the possibility that type I allergic reaction may play role in the pathology of BA. PMID- 11101733 TI - Horse-related injuries in pediatric patients. AB - PURPOSE: The aim of this study was to describe the characteristics, nature, severity and outcome of injuries from horse-related trauma in pediatric patients, aged of 19 years or younger. METHODS: Retrospective analysis was conducted of 315 patients recorded in the National Pediatric Trauma Registry from February 1995 to August 1999. RESULTS: A total of 62% of the 315 patients were girls. The median age of injury was 10 years. Sixty-five percent of the patients were injured while mounted on a horse, and the most common mechanism of injury was falling off the horse. The most frequent reason for hospital admission was skeletal fractures followed by head injuries. The head, neck, and face area was the most commonly injured anatomic site, followed by the upper extremity, the abdomen, and then the lower extremity. The median length of stay in the hospital was 2 days. Forty percent of the patients needed treatment in the intensive care unit with a median length of stay of 2 days. Thirty-nine percent of patients underwent surgical procedures. The Injury Severity Score ranged from moderate to critical in 31.5% of the children. There were 8 deaths, 2.5% of the injured children. The most common cause of mortality was head injuries. Of the 307 survivors, 3% were discharged to a rehabilitation center, and 2% of the children had 1 or more functional impairments lasting longer than 7 months after discharge. CONCLUSIONS: Horse-related trauma is frequent in children and can cause severe injuries resulting in death and long-term disability. Awareness of the nature of injuries is important to avoid underestimation of their severity. PMID- 11101734 TI - Biliary atresia with extrahepatic biliary cysts--cholangiographic patterns influencing the prognosis. AB - PURPOSE: Biliary atresia (BA) with extrahepatic biliary cysts (EHBC) has been recognized generally as "correctable" BA, which indicates a good prognosis. The variants of BA with EHBC according to cholangiographic findings and their outcomes were reviewed. METHODS: An EHBC was observed in 8 (20%) of 40 patients with BA who underwent operation at our institute. Intraoperative cholangiographic patterns included visualization of the intrahepatic bile ducts (type I BA with EHBC) in 6 patients and no visualization (type III BA with EHBC) in 2. Intrahepatic biliary cysts (IHBC) and EHBC were observed simultaneously in 2 patients diagnosed at older age. The follow-up periods ranged between 4 months and 20 years. RESULTS: Good bile drainage after a hepaticoenterostomy or portoenterostomy was obtained in all 6 patients with type I BA with EHBC. Two who showed IHBC on intraoperative cholangiography had complications caused by postoperative recurrent cholangitis, which led to a liver transplantation in 1. Revision after the portoenterostomy was required in 2 patients with type III BA with EHBC. One became jaundice free after revision, whereas the other died of hepatic failure without bile drainage. CONCLUSION: Intraoperative cholangiographic findings showing IHBC and type III BA are poor prognostic factors in patients with BA with EHBC. PMID- 11101735 TI - Polyvinylchloride infusion lines expose infants to large amounts of toxic plasticizers. AB - PURPOSE: The purpose of this study was to evaluate whether infusion lines are able to leach plasticizers in substantial amounts and thus be a candidate substance for hepatotoxic effects during long-term total parenteral nutrition (TPN). METHODS: TPN solutions, blood products, and selected drugs typical for preterm infants concerning amount, content, and infusion time were perfused through common polyvinylchloride (PVC) infusion lines. Concentration of diethylhexyl-phthalate (DEHP) before and after perfusion was determined by gas chromatography/mass spectrometry. RESULTS: Daily quantities of DEHP by 24-hour infusions were Lipid emulsion 20%: 10185.6 microg; aminoacid/glucose-solution: 116.2 microg; midazolaminfusion for sedation: 26.4 microg; fentanyl for sedation: 132.5 microg; propofol for sedation: 6561.0 microg. The amount of DEHP by single doses of blood products (20 mL) were packed red blood cells: 144-608 microg; platelet rich plasma: 928 microg; and fresh frozen plasma: 552-8108 microg. The dose of DEHP for a typical preterm neonate requiring TPN and additional therapy like sedation or blood products is at minimum 10 mg and can easily reach 20 mg/d. CONCLUSION: This large amount of DEHP is especially disturbing, because it effects the most vulnerable patients (neonates). Whether there is a relation to TPN-induced hepatobiliary dysfunction remains to be elucidated and is under investigation. With respect to recent literature, a biological effect of these doses must be assumed. PMID- 11101736 TI - Lordosis of lumbar vertebrae in omphalocele: an important factor in regulating abdominal cavity capacity. AB - PURPOSE: Although it is well known that the tiny abdominal cavity associated with larger omphalocele enlarges rapidly after surgery, the responsible mechanism remains obscure. The authors have studied the curve of lumbar vertebrae and established a hypothesis in which lordosis of lumbar vertebrae plays a major role in changing the capacity of the abdominal cavity. METHODS: Fifty-three patients with intact omphalocele and 10 normal newborns as the control were studied. The size of omphalocele was determined from the mean value of the length and breadth of the sac. The curve of lumbar vertebrae (lordotic angle) was measured using Cobb's technique on the lateral radiograph serially taken in the supine position. RESULTS: The mean lordotic angle of 53 patients was 22.2 degrees (range, 0 to 65), which was significantly greater than 11.0 degrees of the control (P <.012). The individual lordotic angle before surgery was significantly correlated with the size of omphalocele (r = 0.668, P <.0001). In serial studies carried out in 15 patients with larger omphalocele, the mean lordotic angle was decreased significantly from 30.5 degrees to 20.0 degrees (P <.009) on the first postoperative day and returned to the control level within a few weeks. CONCLUSIONS: These results suggest that severe lumbar lordosis may well become an alternative factor to produce visceroabdominal disproportion observed before surgery, and its correction after surgery may well induce the abdominal cavity to rapidly enlarge. It may be speculated that the dehiscence of abdominal rectus muscles associated with omphalocele causes the tension of ventral muscles to decline, and, then, imbalance in tension between ventral (abdominal) and dorsal (back) muscles causes the lumbar lordosis to increase. Accordingly, at the staged procedure, the prosthetic sheet should be attached under moderate tension, and its plication should be carried out at regular intervals to maintain the tension of abdominal rectus muscles. PMID- 11101737 TI - High incidence of adriamycin cardiotoxicity in children even at low cumulative doses: role of radionuclide cardiac angiography. AB - BACKGROUND/PURPOSE: Adriamycin (doxorubicin), a chemotherapeutic agent commonly used in the treatment of pediatric solid tumors, is known to have a dose-related cardiotoxicity, which is reported to be more common in children. The clinical manifestation of this is congestive cardiac failure (CHF), and this is fatal in 50% of the cases. Various strategies, including prospective multiple gated acquisition (MUGA) scan for early detection of the onset of cardiac damage has been recommended to decrease this fatality caused by cardiotoxicity. METHODS: All children receiving Adriamycin for solid tumors, registered at our pediatric solid tumor clinic from January 1998 through June 1999, were included in the study. Cardiotoxicity was evaluated by using electrocardiogram (ECG)-gated radionuclide cardiac angiography (MUGA scan). RESULTS: MUGA scans were performed on 55 patients in the above-mentioned period of 18 months. Twenty-three patients had only 1 baseline MUGA scan done, 12 of these are awaiting further studies at appropriate time and 11 are either dead or lost to follow-up. Of the remaining 32 patients, 13 have shown evidence of cardiotoxicity on MUGA scan done at a cumulative dose of 180 to 200 mg/m(2), in the form of decrease in left ventricular ejection fraction (LVEF) or abnormality in myocardial movements. Three of these 13 patients had clinical evidence of CHF. In 10 patients, timely discontinuation of Adriamycin, based on the MUGA report, probably has helped avoid the development of CHF. CONCLUSION: Routine monitoring of all children receiving Adriamycin is required to avoid the mortality and morbidity of Adriamycin-related cardiotoxicity, which may develop at relatively low cumulative doses also. PMID- 11101738 TI - Currarino triad--diagnostic dilemma and a combined surgical approach. AB - PURPOSE: The authors present 2 families with 3 cases of Currarino triad, diagnostic difficulties, their familial occurrence, and genetic mapping, with emphasis on a combined pediatric surgical and pediatric neurosurgical approach in managing these children. RESULTS: The main presentation was intractable constipation. In the first family there was a 4-generation pedigree with recurrence of Currarino triad. The mother and the child have the condition. Family 2 screening showed a 3-generation pedigree with presence of Currarino triad in 3 members. Patients 2 and 3 are cousins whose fathers are affected by spina bifida occulta and Currarino triad, respectively. In patient 1, the diagnosis was made after inadvertent rupture of an anterior meningocele during posterior myectomy. In patient 2, the presacral mass was found on examination under anesthesia, and the planned anorectal myectomy for intractable constipation was abandoned. Patient 3 was a cousin of patient 2, and the diagnosis was considered when she presented with intractable constipation at the age of 7 months. Magnetic resonance scan was useful in showing the presence of presacral mass, spinal abnormalities, and tethered cord. A combined pediatric and neurosurgical approach optimized the extirpation of the presacral mass with minimal complications. Surgical treatment was individualized according to the estimation of the operative risk factors. All patients have a normal bladder function. Patient 1 has required laxatives and enemas for intermittent constipation. She has associated learning difficulties but is otherwise well. Patient 2 and 3, aged 10 and 2 years, respectively, are awaiting closure of colostomy. They are thriving and well. CONCLUSIONS: The authors recommend a combined pediatric and neurosurgical assessment and management for all cases of Currarino triad. Family screening is obligatory. The authors suggest the use of a magnetic resonance scan or computerized axial tomography myelogram to define the presence of anosacral and spinal cord anomalies in patients with intractable constipation. PMID- 11101739 TI - Removal of adherent ventricular catheters by a modified sheath introducer system: technical note. AB - BACKGROUND/PURPOSE: A series of technical notes has been dedicated to the removal of retained intracranial shunt catheters, among which the intraluminal cautery proved to be the most accepted technique. However, several reports showed that these techniques still harbor potentially serious complications. METHODS: In this technique, a modified plastic sheath introducer system is passed over the retained ventricular catheter. While advancing the tube along the longitudinal axis of the catheter, circular movements of the tube around the longitudinal axis of the catheter are performed, allowing the tube to act as a spherical knife cutting the ingrown choroid plexus or ependymal adhesions. RESULTS: There were no procedure-related complications in any of the 9 patients treated by the technique described. The procedure proved to be easy and effective in all cases. In addition, in case a new ventricular catheter was needed at the same site, it could be placed via the same tube. CONCLUSIONS: The technique described seems to be an easy, safe, and effective alternative to other techniques for removal of retained ventricular catheters. However, considering the limited number of patients treated with the technique described and the great number of patients treated by the widely accepted intraluminal cautery, one cannot claim the one technique as superior to the other at this stage. PMID- 11101740 TI - Colon injuries in children. AB - BACKGROUND/PURPOSE: Colonic injuries are rare in childhood, but when they do occur, they are mostly associated with penetrating abdominal injuries. The primary repair of colon injuries without stoma is still controversial within surgical experience, and the potential risk factors affecting morbidity and mortality is not sufficiently known. METHODS: Between 1985 and 1997, 34 children presenting with traumatic colonic perforations were reevaluated by analyzing the relationship between the overall morbidity and mortality and the potential risk factors. RESULTS: Of the 34 children in the case study, 27 boys and 7 girls, there were 7 (21%) isolated colonic injuries. The remaining 27 (79%) patients showed colonic injuries most frequently associated with the small bowel, the liver, and the bladder. Localization of injury was distributed thus: 21% in the right colon, 29% in the transverse colon, and 50% in the left colon. Primary repair, with or without intestinal resection, was performed in 27 (79%) of the patients. In total, postoperative complications occurred in 10 (29%) of the patients. Risk factors such as age, abdominal contamination, and associated abdominal organ injuries were found significant in these complications, however, the mechanism of injury, shock, blood transfusion, and localization of injury were not correlated significantly to postoperative complications. "'Flint's Colon Grading System" was used to ascertain the sensitivity of trauma scoring systems for postoperative complications. CONCLUSION: Colonic wounds can be repaired primarily without the need of colostomy in the majority of cases in children when the required selections are established. PMID- 11101741 TI - Hirschsprung's disease: problems with transition-zone pull-through. AB - BACKGROUND/PURPOSE: It is generally accepted that if surgery for Hirschsprung's disease is to be successful, ganglionic bowel must be anastomosed to the lower rectum or anal canal. Above the aganglionic distal bowel lies a transition zone (TZ) where more subtle abnormalities of innervation are apparent. The significance of this transition zone in respect to the functional outcome of surgery has received little attention. The aim of this study was to identify the incidence of transition zone pull-through (TZPT) in a cohort of children who underwent surgery for Hirschsprung's disease, to identify the reasons why TZPTs occurred, and to identify the functional consequences. The authors report the long-term outcome of these children with emphasis on bowel function and the results of subsequent surgery. METHODS: A Retrospective study was conducted of children treated at a single institution from 1979 through 1994. TZPT patients were subject to detailed review of surgical records and histopathologic material. RESULTS: Thirteen children were identified with a TZPT. In 12 cases, histopathologic errors contributed to the TZPT: in 5 cases this was caused by single point biopsies missing an asymmetrical TZ, whereas in 7 cases the histopathologic features of the TZ were not recognized. In 1 case the TZPT was caused by surgical error. As a consequence of the TZPT 7 children underwent repeat pull-through. One child is fully continent, one has daytime fecal continence, and 2 others are incontinent. Two children have permanent stomas. One child is clean with antegrade colonic washouts. Repeat pull-throughs were not attempted in 6 children. Two children have achieved full continence, 2 have permanent stomas, 1 is clean with antegrade colonic washouts, and 1 child receives regular suppositories. CONCLUSIONS: Transition zone pull-throughs occurred because of a combination of surgical and histopathologic errors. The transition zone may follow an asymmetric course around the circumference of the bowel and may be missed if single-point extramucosal biopsy specimens are taken. Recognition of the subtle histologic features of the transition zone requires an experienced pathologist. The functional consequences of a TZPT are severe, with symptoms of constipation, diarrhea, and incontinence. The results of revisional pull-through were disappointing. Serious consideration should be given to alternative procedures such as the antegrade continence enema operation. PMID- 11101742 TI - Clarification of the process of separation of the cloaca into rectum and urogenital sinus in the rat embryo. AB - BACKGROUND/PURPOSE: The normal process of division of the cloaca into a rectum and urogenital tract is still not fully understood. The main controversies relate to how the urorectal septum (URS) divides the cloaca and whether the URS fuses with the cloacal membrane. This study used a 3-dimensional reconstruction technique, combined with histologic correlation, to observe the developmental and septational processes of the cloaca of the normal rat embryo from gestational days 11 to 16. METHODS: Normal rat embryos from gestational days 11 to 16 were sectioned serially both transversely and sagittally and stained with H&E. 3 dimensional reconstructions were performed on embryos younger than day 13.5. The relevant structures were examined in a temporo-spatial sequence. RESULTS: The tailgut started to regress by apoptosis on day 12 in a cranio-caudal direction. The URS, first evident in day-12.5 embryos, extended and fused with the cloacal membrane on day 15 of gestation, completing the separation of the cloaca into rectum and bladder. Regression of the tailgut and ventral protrusion of the urogenital sinus markedly changed the configuration of the cloaca. The cloacal membrane did not break down until after it had fused with the URS. CONCLUSIONS: These findings clarify the relative contributions made by active septation of the cloaca by the URS and configurational changes of the cloaca to produce a rectum and bladder. The URS fuses with the cloacal membrane before the anal and urogenital membranes break down. PMID- 11101743 TI - Association of bronchopulmonary sequestration with expression of the homeobox protein Hoxb-5. AB - Bronchopulmonary sequestration (BPS) is caused by the abnormal development of an accessory lung diverticulum from the foregut very early in embryogenesis. The developmental abnormalities seen with BPS suggest that this anomaly is caused by abnormal expression of homeobox genes, which control axial identity and organ specific patterning during embryogenesis. The authors previously have shown that the homeobox gene Hoxb-5 is necessary for normal airway branching during lung development. The authors now report that BPS is associated with aberrant developmental expression of Hoxb-5 protein, suggesting that this Hox gene is involved in the development of BPS. PMID- 11101744 TI - Surgical intervention for emphysematous pulmonary regions in a postoperative infant with congenital diaphragmatic hernia. AB - A postoperative infant with congenital diaphragmatic hernia (CDH) developed extrinsic obstruction of the trachea by the innominate artery that ensued from unequal expansion of the lungs followed by left mediastinal shift. Septation of the anterior mediastinum prevented unequal expansion of the lungs, and elongation of the innominate artery improved proximal airway obstruction. Prolonged artificial ventilation, however, resulted in the emphysematous bullae in the left lung. Lung volume reduction surgery (LVRS), at 3 years of age, ameliorated the respiratory distress and resulted in good weight gain. Surgical intervention, including LVRS, should be considered to improve respiratory disturbance caused by difference in compliance of the lungs in children. PMID- 11101745 TI - Salvage of the ruptured spleen in an infant with very low birth weight. AB - Hemorrhage from the traumatized spleen needs surgical intervention if the patient's cardiovascular status is unstable and not stabilizing with medical support. It is well recognized that salvage of the injured spleen is preferable to splenectomy because of risks of overwhelming postsplenectomy infection (OPSI). The authors report the case of a 746-g newborn with splenic hemorrhage that required surgical intervention to control bleeding. A 2-component fibrin tissue adhesive and a cellulose fabric were used. PMID- 11101746 TI - Bilateral eventration of the diaphragm with perforated gastric volvulus in an adolescent. AB - Bilateral congenital eventration of the diaphragm almost uniformly presents in infancy with respiratory compromise and is associated with a high mortality rate. Delayed presentation of diaphragmatic eventration in older children and adults may be associated with acute gastric volvulus. Thus, any patient with abdominal pain, vomiting, or nonspecific gastrointestinal symptoms in association with abnormal diaphragmatic findings on chest x-ray should undergo further diagnostic workup with upper gastrointestinal series or computed tomography (CT) scan. Treatment of gastric volvulus requires immediate surgical repair to prevent subsequent necrosis and perforation. The authors describe a case report of bilateral congenital diaphragmatic eventration complicated by a perforated gastric volvulus in a 13-year-old boy. Emergent reduction of the volvulus, closure of the perforated stomach, plication of the diaphragm, and placement of gastrostomy was performed successfully. PMID- 11101747 TI - Lateral esophagostomy: an alternative in the initial management of long gap esophageal atresia without fistula. AB - The authors report an alternative method of cervical esophagostomy that was used in a child with type A esophageal atresia. This method involved performing a lateral esophagostomy in the proximal pouch, preserving its distal end, allowing the child to swallow normally, without choking, while stimulating the spontaneous growth of the proximal esophagus. As a result, the infant could be discharged home on G-tube feedings while waiting for spontaneous growth of the proximal pouch to occur. There were no episodes of aspiration during this period, and definitive reconstruction through end-to-end esophageal anastomosis was accomplished successfully at the age of 18 months. The authors consider that this alternative might increase the possibility of a definitive correction through delayed primary anastomosis of the infant's own esophagus in children with this type of malformation. PMID- 11101748 TI - Early primary repair of long gap esophageal atresia: the VATER operation. AB - Despite the numerous approaches described for the management of neonates with "long gap" esophageal atresia, controversy still exists as to the preferred method. Delayed primary anastomosis is probably the most frequently adopted practice, but often the native esophagus is abandoned, and the long gap is bridged by some form of esophageal replacement. A case is reported of a 1.38-kg newborn with pure esophageal atresia and a long gap undergoing early primary repair. The technique used in this patient and the various options available for management of long-gap esophageal atresia are discussed. PMID- 11101749 TI - Tailgut cyst in a neonate. AB - Tailgut cyst is a rare lesion of developmental origin located in the retrorectal space, which usually presents as a multilocular cystic mass. It is usually found in adults, and neonatal cases are extremely rare. The authors report a tailgut cyst in a neonate that was found by prenatal ultrasonogram, which was like a teratoma in gross appearance. PMID- 11101750 TI - Vascular control for resection of suprahepatic intracaval Wilms' tumor: technical considerations. AB - The surgical resection of Wilms' tumor can be complicated by tumor thrombus extension into the inferior vena cava. In cases of suprahepatic Wilms' tumor thrombus that may extend into the right atrium, a median sternotomy and cardiopulmonary bypass (CPB) are used to facilitate tumor resection. However, if the tumor can be localized and controlled below the atrium, resection without the use of cardiopulmonary bypass may limit morbidity. The authors describe a novel approach to tumor thrombectomy for a Wilms' tumor extending to the suprahepatic vena cava without the use of CPB. The authors used transesophageal echocardiography to localize the tumor thrombus and detect any tumor or air embolization and a minimal lower sternotomy to obtain intrapericardial control of the inferior vena cava. This technique may be useful in selected cases of Wilms' tumor as an alternative to median sternotomy and use of cardiopulmonary bypass. PMID- 11101751 TI - Squamous cell carcinoma of the bladder in a pediatric patient. AB - A 16-year-old North American girl with a prior history of a bladder calculus removed by open cystolithotomy 8 years earlier, subsequently had squamous cell carcinoma of the bladder along the site of the previous incision. The authors believe that this child is the youngest documented patient with squamous cell carcinoma of the bladder not associated with schistosomiasis. PMID- 11101752 TI - Smith-Lemli-Opitz syndrome: case report and literature review. AB - The authors report the case of a 6-week-old boy with the Smith-Lemli-Opitz syndrome (SLOS) and review the literature on the subject. Intersexuality was suspected and a laparoscopy performed. Abnormalities of the gastrointestinal tract, the lower extremities, and the face prompted DNA analysis, which found a defect of cholesterol biosynthesis in the form of the Smith-Lemli-Opitz-syndrome, a rare congenital defect. The clinical course of this case is compared with similar cases in the literature. PMID- 11101754 TI - Differences between amnioinfusion and amniotic fluid exchange. PMID- 11101753 TI - A case of pediatric Henoch-Schonlein purpura and thrombosis of spermatic veins. AB - The authors report a case of thrombosis of the spermatic veins associated with Henoch-Schonlein purpura mimicking an acute scrotum, which responded to a low molecular-weight heparin treatment. PMID- 11101756 TI - Interval appendectomy after conservative treatment of periappendicular abscess. PMID- 11101757 TI - Knee-jerk response. PMID- 11101758 TI - Therapeutic space invaders. PMID- 11101759 TI - Avoiding frankendrugs. PMID- 11101760 TI - Cloned pig litter update. PMID- 11101761 TI - Breaching principles. PMID- 11101773 TI - Stem cells hunt tumors PMID- 11101762 TI - Direct action. PMID- 11101775 TI - Defensin stimulant PMID- 11101774 TI - Artificial blood PMID- 11101777 TI - Chromosomal checkup PMID- 11101778 TI - Microarrays-a closer look? PMID- 11101779 TI - Evolving antibodies in a test tube PMID- 11101780 TI - Modulating protein stability PMID- 11101781 TI - Ramping up antigen presentation PMID- 11101782 TI - Gene regulation in flux PMID- 11101783 TI - A map of the protein world PMID- 11101784 TI - Evolving sturdier gene therapy vectors PMID- 11101785 TI - Technical reports PMID- 11101787 TI - Brighter aptamers for biosensing PMID- 11101786 TI - Review PMID- 11101789 TI - Gene amplification for plants PMID- 11101788 TI - Crop fungal protection PMID- 11101790 TI - Company, academics argue over data. PMID- 11101791 TI - First genetic trust banks on genes. PMID- 11101792 TI - Sweeping bioethical reform proposals contemplated for trials. PMID- 11101793 TI - Cantab failure shows need for wide portfolio. PMID- 11101794 TI - HGS targets patent-expiring drugs. PMID- 11101795 TI - Chiral drugs viable, despite failure. PMID- 11101796 TI - Dutch bill unlikely to revive industry. PMID- 11101797 TI - New tools for an old workhorse. PMID- 11101798 TI - Protein networks-built by association. PMID- 11101799 TI - Viral vectors do the DNA shuffle. PMID- 11101800 TI - Triplex technology takes off. PMID- 11101801 TI - The changing marketplace of bioinformatics. PMID- 11101802 TI - Applications of display technology in protein analysis. AB - Display technology refers to a collection of methods for creating libraries of modularly coded biomolecules that can be screened for desired properties. It has become a routine tool for enriching molecular diversity and producing novel types of proteins. The combination of an ever-increasing variety of libraries of modularly coded protein complexxes with the development of innovative approaches to select a wide array of desired properties has facilitated large-scale analyses of protein-protein/protein-substrate interactions, rapid isolation of antibodies (or antibody mimetics) without immunization, and function-based protein analysis. Several practical and theoretical challenges remain to be addressed before display technology can be readily applied to proteomic studies. PMID- 11101803 TI - A network of protein-protein interactions in yeast. AB - A global analysis of 2,709 published interactions between proteins of the yeast Saccharomyces cerevisiae has been performed, enabling the establishment of a single large network of 2,358 interactions among 1,548 proteins. Proteins of known function and cellular location tend to cluster together, with 63% of the interactions occurring between proteins with a common functional assignment and 76% occurring between proteins found in the same subcellular compartment. Possible functions can be assigned to a protein based on the known functions of its interacting partners. This approach correctly predicts a functional category for 72% of the 1,393 characterized proteins with at least one partner of known function, and has been applied to predict functions for 364 previously uncharacterized proteins. PMID- 11101804 TI - RNA expression analysis using a 30 base pair resolution Escherichia coli genome array. AB - We have developed a high-resolution "genome array" for the study of gene expression and regulation in Escherichia coli. This array contains on average one 25-mer oligonucleotide probe per 30 base pairs over the entire genome, with one every 6 bases for the intergenic regions and every 60 bases for the 4,290 open reading frames (ORFs). Twofold concentration differences can be detected at levels as low as 0.2 messenger RNA (mRNA) copies per cell, and differences can be seen over a dynamic range of three orders of magnitude. In rich medium we detected transcripts for 97% and 87% of the ORFs in stationary and log phases, respectively. We found that 1, 529 transcripts were differentially expressed under these conditions. As expected, genes involved in translation were expressed at higher levels in log phase, whereas many genes known to be involved in the starvation response were expressed at higher levels in stationary phase. Many previously unrecognized growth phase-regulated genes were identified, such as a putative receptor (b0836) and a 30S ribosomal protein subunit (S22), both of which are highly upregulated in stationary phase. Transcription of between 3,000 and 4,000 predicted ORFs was observed from the antisense strand, indicating that most of the genome is transcribed at a detectable level. Examples are also presented for high-resolution array analysis of transcript start and stop sites and RNA secondary structure. PMID- 11101805 TI - Enhanced major histocompatibility complex class I-dependent presentation of antigens modified with cationic and fusogenic peptides. AB - Soluble extracellular protein antigens are notoriously poor stimulators of CD8+ cytotoxic T-lymphocyte (CTL) responses, largely because these antigens have inefficient access to an endogenous cytosolic pathway of the major histocompatibility complex (MHC) class I-dependent antigen presentation. Here, we present a strategy that facilitates antigen penetration into the cytosol of antigen-presenting cells (APC) by addition to the antigen of charge-modifying peptide sequences. As a result of this intervention, the charge modification enhances antigen uptake into APC by counteracting the repulsive cell surface charge, and then endosomal membranes are disrupted with a subsequent release of antigen into the cytosol. This technology significantly improves MHC class I dependent antigen presentation to CTL, enabling a more efficient generation of specific CTL immunity in vivo. The strategy described here has potential for use in developing efficient vaccines for antigen-specific immunotherapy of human malignancies. PMID- 11101806 TI - Efficient nonviral transfection of dendritic cells and their use for in vivo immunization. AB - Immunization with dendritic cells (DCs) transfected with genes encoding tumor associated antigens (TAAs) is a highly promising approach to cancer immunotherapy. We have developed a system, using complexes of plasmid DNA expression constructs with the cationic peptide CL22, that transfects human monocyte-derived DCs much more efficiently than alternative nonviral agents. After CL22 transfection, DCs expressing antigens stimulated autologous T cells in vitro and elicited primary immune responses in syngeneic mice, in an antigen specific manner. Injection of CL22-transfected DCs expressing a TAA, but not DCs pulsed with a TAA-derived peptide, protected mice from lethal challenge with tumor cells in an aggressive model of melanoma. The CL22 system is a fast and efficient alternative to viral vectors for engineering DCs for use in immunotherapy and research. PMID- 11101807 TI - Breeding of retroviruses by DNA shuffling for improved stability and processing yields. AB - Manufacturing of retroviral vectors for gene therapy is complicated by the sensitivity of these viruses to stress forces during purification and concentration. To isolate viruses that are resistant to these manufacturing processes, we performed breeding of six ecotropic murine leukemia virus (MLV) strains by DNA shuffling. The envelope regions were shuffled to generate a recombinant library of 5 x 106 replication-competent retroviruses. This library was subjected to the concentration process three consecutive times, with amplification of the surviving viruses after each cycle. Several viral clones with greatly improved stabilities were isolated, with the best clone exhibiting no loss in titer under conditions that reduced the titers of the parental viruses by 30- to 100-fold. The envelopes of these resistant viruses differed in DNA and protein sequence, and all were complex chimeras derived from multiple parents. These studies demonstrate the utility of DNA shuffling in breeding viral strains with improved characteristics for gene therapy. PMID- 11101808 TI - Increasing galactose consumption by Saccharomyces cerevisiae through metabolic engineering of the GAL gene regulatory network. AB - Increasing the flux through central carbon metabolism is difficult because of rigidity in regulatory structures, at both the genetic and the enzymatic levels. Here we describe metabolic engineering of a regulatory network to obtain a balanced increase in the activity of all the enzymes in the pathway, and ultimately, increasing metabolic flux through the pathway of interest. By manipulating the GAL gene regulatory network of Saccharomyces cerevisiae, which is a tightly regulated system, we produced prototroph mutant strains, which increased the flux through the galactose utilization pathway by eliminating three known negative regulators of the GAL system: Gal6, Gal80, and Mig1. This led to a 41% increase in flux through the galactose utilization pathway compared with the wild-type strain. This is of significant interest within the field of biotechnology since galactose is present in many industrial media. The improved galactose consumption of the gal mutants did not favor biomass formation, but rather caused excessive respiro-fermentative metabolism, with the ethanol production rate increasing linearly with glycolytic flux. PMID- 11101809 TI - Picomolar affinity antibodies from a fully synthetic naive library selected and evolved by ribosome display. AB - Here we applied ribosome display to in vitro selection and evolution of single chain antibody fragments (scFvs) from a large synthetic library (Human Combinatorial Antibody Library; HuCAL) against bovine insulin. In three independent ribosome display experiments different clusters of closely related scFvs were selected, all of which bound the antigen with high affinity and specificity. All selected scFvs had affinity-matured up to 40-fold compared to their HuCAL progenitors, by accumulating point mutations during the ribosome display cycles. The dissociation constants of the isolated scFvs were as low as 82 pM, which validates the design of the naive library and the power of this evolutionary method. We have thus mimicked the process of antibody generation and affinity maturation with a synthetic library in a cell-free system in just a few days, obtaining molecules with higher affinities than most natural antibodies. PMID- 11101810 TI - In vitro selection of signaling aptamers. AB - Reagentless biosensors that can directly transduce molecular recognition to optical signals should potentiate the development of sensor arrays for a wide variety of analytes. Nucleic acid aptamers that bind ligands tightly and specifically can be readily selected, but may prove difficult to adapt to biosensor applications. We have therefore attempted to develop selection methods that couple the broad molecular recognition properties of aptamers with signal transduction. Anti-adenosine aptamers were selected from a pool that was skewed to contain very few fluoresceinated uridines. The primary family of aptamers showed a doubling of relative fluorescence intensity at saturating concentrations of a cognate analyte, ATP, and could sense ATP concentrations as low as 25 microM. A single uridine was present in the best signaling aptamer. Surprisingly, other dyes could substitute for fluorescein and still specifically signal the presence of ATP, indicating that the single uridine functioned as a general "switch" for transducing molecular recognition to optical signals. PMID- 11101811 TI - A ubiquitin-based tagging system for controlled modulation of protein stability. AB - Many biotechnology applications depend on the expression of exogenous proteins in a predictable and controllable manner. A key determinant of the intracellular concentration of a given protein is its stability or "half-life." We have developed a versatile and reliable system for producing short half-life forms of proteins expressed in mammalian cells. The system consists of a series of destabilization domains composed of varying numbers of a mutant form of ubiquitin (UbG76V) that cannot be cleaved by ubiquitin hydrolases. We show that increasing the number of UbG76V moieties within the destabilization domain results in a graded decrease in protein half-life and steady-state levels when fused to heterologous reporter proteins as well as cellular proteins. Cells expressing a destabilized beta-lactamase reporter act as a robust, high-throughput screening (HTS)-compatible assay for proteasome activity within cells. PMID- 11101812 TI - Tobacco ribosomal DNA spacer element stimulates amplification and expression of heterologous genes. AB - Here we show that the cis-acting genetic element aps (amplification-promoting sequence), isolated from the nontranscribed spacer region of tobacco ribosomal DNA (rDNA), increases the level of expression of recombinant proteins. Transgenic tobacco plants, transformed with expression cassettes containing the herbicide resistant acetolactate synthase (hr-ALS) gene or the green fluorescent protein (GFP) gene fused to the aps sequence, had greater levels of corresponding messenger RNAs (mRNAs) and proteins compared to transformants lacking aps. Analysis of transgenic plants showed that aps increased the copy number and transcription of the adjacent heterologous genes and, in the case of hr-ALS, enhanced the herbicide resistance phenotype. Both the increased transgene copy number and enhanced expression were stably inherited. These data provide the first evidence that the aps sequence can be used for gene amplification in transgenic plants and possibly other multicellular organisms. PMID- 11101813 TI - Fungal pathogen protection in potato by expression of a plant defensin peptide. AB - Defensins are small cysteine-rich peptides with antimicrobial activity. We demonstrate that the alfalfa antifungal peptide (alfAFP) defensin isolated from seeds of Medicago sativa displays strong activity against the agronomically important fungal pathogen Verticillium dahliae. Expression of the alfAFP peptide in transgenic potato plants provides robust resistance in the greenhouse. Importantly, this resistance is maintained under field conditions. There have been no previous demonstrations of a single transgene imparting a disease resistance phenotype that is at least equivalent to those achieved through current practices using fumigants. PMID- 11101814 TI - Stable correction of a genetic deficiency in human cells by an episome carrying a 115 kb genomic transgene. AB - Persistent expression of a transgene at therapeutic levels is required for successful gene therapy, but many small vectors with heterologous promoters are prone to vector loss and transcriptional silencing. The delivery of genomic DNA would enable genes to be transferred as complete loci, including regulatory sequences, introns, and native promoter elements. These elements may be critical to ensure prolonged, regulated, and tissue-specific transgene expression. Many studies point to considerable advantages to be gained by using complete genomic loci in gene expression. Large-insert vectors incorporating elements of the bacterial artificial chromosome (BAC) cloning system, and the episomal maintenance mechanisms of Epstein-Barr virus (EBV), can shuttle between bacteria and mammalian cells, allowing large genomic loci to be manipulated conveniently. We now demonstrate the potential utility of such vectors by stably correcting a human genetic deficiency in vitro. When the complete hypoxanthine phosphoribosyltransferase (HPRT) locus of 115 kilobases (kb) was introduced into deficient human cells, the transgene was both maintained as an episome and expressed stably for six months in rapidly dividing cell cultures. The results demonstrate for the first time that gene expression from an episomal genomic transgene can correct a cell culture disease phenotype for a prolonged period. PMID- 11101815 TI - DNA cloning by homologous recombination in Escherichia coli. AB - The cloning of foreign DNA in Escherichia coli episomes is a cornerstone of molecular biology. The pioneering work in the early 1970s, using DNA ligases to paste DNA into episomal vectors, is still the most widely used approach. Here we describe a different principle, using ET recombination, for directed cloning and subcloning, which offers a variety of advantages. Most prominently, a chosen DNA region can be cloned from a complex mixture without prior isolation. Hence cloning by ET recombination resembles PCR in that both involve the amplification of a DNA region between two chosen points. We apply the strategy to subclone chosen DNA regions from several target molecules resident in E. coli hosts, and to clone chosen DNA regions from genomic DNA preparations. Here we analyze basic aspects of the approach and present several examples that illustrate its simplicity, flexibility, and remarkable efficiency. PMID- 11101816 TI - Research corrigendum PMID- 11101817 TI - Research errata PMID- 11101818 TI - Moving the gene patent debate forward. PMID- 11101820 TI - Promotions, advancements PMID- 11101819 TI - Recent patents in phage display. PMID- 11101822 TI - 2000 subject index - volume 18 PMID- 11101824 TI - Who owns plant genetics? PMID- 11101825 TI - Uncoupling expectations. PMID- 11101826 TI - Recombination: a means to an end in human cells. PMID- 11101827 TI - Breast cancer--not just whether but when? PMID- 11101829 TI - TOUCHINGbase PMID- 11101828 TI - p53 and p73: seeing double? PMID- 11101830 TI - Genetic homogeneity of Icelanders. PMID- 11101831 TI - Analysis of complex traits: mutagenesis versus QTLs. PMID- 11101832 TI - Amegakaryocytic thrombocytopenia and radio-ulnar synostosis are associated with HOXA11 mutation. PMID- 11101833 TI - A region on chromosome 3 is linked to dizygotic twinning. PMID- 11101834 TI - Selfish gene is offside PMID- 11101835 TI - The transcriptome of Arabidopsis thaliana during systemic acquired resistance. AB - Infected plants undergo transcriptional reprogramming during initiation of both local defence and systemic acquired resistance (SAR). We monitored gene expression changes in Arabidopsis thaliana under 14 different SAR-inducing or SAR repressing conditions using a DNA microarray representing approximately 25-30% of all A. thaliana genes. We derived groups of genes with common regulation patterns, or regulons. The regulon containing PR-1, a reliable marker gene for SAR in A. thaliana, contains known PR genes and novel genes likely to function during SAR and disease resistance. We identified a common promoter element in genes of this regulon that binds members of a plant-specific transcription factor family. Our results extend expression profiling to definition of regulatory networks and gene discovery in plants. PMID- 11101836 TI - High constant incidence in twins and other relatives of women with breast cancer. AB - The incidence of breast cancer rises steeply between ages 25 and 50, and more slowly thereafter. In contrast, the incidence in the unaffected (contralateral) breast of women who have had breast cancer remains constant at about 0.7% per year for at least the next 20 years after diagnosis, irrespective of age at first diagnosis. The incidence in relatives of the patients seems to show a similar pattern. The incidence in a prospective study of monozygotic twins of patients was approximately constant at 1.3% per year (77 cases), again about 0.7% per breast. At ages older than a patient's age at diagnosis, her mother and sisters have an incidence of 0.3-0.4% per year. Above the index patient's age at diagnosis, the rate in relatives shows no temporal trend and is independent of the patient's age at diagnosis. A statistically simple explanation is that incidence in susceptible women increases to a high constant level by a predetermined age that varies between families, but this seems inconsistent with conventional models of carcinogenesis and susceptibility. The very high incidence in monozygotic twins of patients indicates that a high proportion, and perhaps the majority, of breast cancers arise in a susceptible minority of women. PMID- 11101837 TI - The core meiotic transcriptome in budding yeasts. AB - We used high-density oligonucleotide microarrays to analyse the genomes and meiotic expression patterns of two yeast strains, SK1 and W303, that display distinct kinetics and efficiencies of sporulation. Hybridization of genomic DNA to arrays revealed numerous gene deletions and polymorphisms in both backgrounds. The expression analysis yielded approximately 1,600 meiotically regulated genes in each strain, with a core set of approximately 60% displaying similar patterns in both strains. Most of these (95%) are MATa/MATalpha-dependent and are not similarly expressed in near-isogenic meiosis-deficient controls. The transcript profiles correlate with the distribution of defined meiotic promoter elements and with the time of known gene function. PMID- 11101838 TI - Cancer predisposition caused by elevated mitotic recombination in Bloom mice. AB - Bloom syndrome is a disorder associated with genomic instability that causes affected people to be prone to cancer. Bloom cell lines show increased sister chromatid exchange, yet are proficient in the repair of various DNA lesions. The underlying cause of this disease are mutations in a gene encoding a RECQ DNA helicase. Using embryonic stem cell technology, we have generated viable Bloom mice that are prone to a wide variety of cancers. Cell lines from these mice show elevations in the rates of mitotic recombination. We demonstrate that the increased rate of loss of heterozygosity (LOH) resulting from mitotic recombination in vivo constitutes the underlying mechanism causing tumour susceptibility in these mice. PMID- 11101839 TI - Dominant modifier DFNM1 suppresses recessive deafness DFNB26. AB - More than 50% of severe childhood deafness is genetically determined, approximately 70% of which occurs without other abnormalities and is thus termed nonsyndromic. So far, 30 nonsyndromic recessive deafness loci have been mapped and the defective genes at 6 loci, DFNB1, DFNB2, DFNB3, DFNB4, DFNB9 and DNFB21, have been identified, encoding connexin-26 (ref. 3), myosin VIIA (ref. 4), myosin XV (ref. 5), pendrin, otoferlin and alpha-tectorin, respectively. Here we map a new recessive nonsyndromic deafness locus, DFNB26, to a 1.5-cM interval of chromosome 4q31 in a consanguineous Pakistani family. A maximum lod score of 8.10 at theta=0 was obtained with D4S1610 when only the 8 affected individuals in this family were included in the calculation. There are seven unaffected family members who are also homozygous for the DFNB26-linked haplotype and thus are non penetrant. A dominant modifier, DFNM1, that suppresses deafness in the 7 nonpenetrant individuals was mapped to a 5.6-cM region on chromosome 1q24 with a lod score of 4.31 at theta=0 for D1S2815. PMID- 11101840 TI - Disruption of the uncoupling protein-2 gene in mice reveals a role in immunity and reactive oxygen species production. AB - The gene Ucp2 is a member of a family of genes found in animals and plants, encoding a protein homologous to the brown fat uncoupling protein Ucp1 (refs 1 3). As Ucp2 is widely expressed in mammalian tissues, uncouples respiration and resides within a region of genetic linkage to obesity, a role in energy dissipation has been proposed. We demonstrate here, however, that mice lacking Ucp2 following targeted gene disruption are not obese and have a normal response to cold exposure or high-fat diet. Expression of Ucp2 is robust in spleen, lung and isolated macrophages, suggesting a role for Ucp2 in immunity or inflammatory responsiveness. We investigated the response to infection with Toxoplasma gondii in Ucp2-/- mice, and found that they are completely resistant to infection, in contrast with the lethality observed in wild-type littermates. Parasitic cysts and inflammation sites in brain were significantly reduced in Ucp2-/- mice (63% decrease, P<0.04). Macrophages from Ucp2-/- mice generated more reactive oxygen species than wild-type mice (80% increase, P<0.001) in response to T. gondii, and had a fivefold greater toxoplasmacidal activity in vitro compared with wild-type mice (P<0.001 ), which was absent in the presence of a quencher of reactive oxygen species (ROS). Our results indicate a role for Ucp2 in the limitation of ROS and macrophage-mediated immunity. PMID- 11101841 TI - The imprinting box of the Prader-Willi/Angelman syndrome domain. AB - A subset of mammalian genes is monoallelically expressed in a parent-of-origin manner. These genes are subject to an imprinting process that epigenetically marks alleles according to their parental origin during gametogenesis. Imprinted genes can be organized in clusters as exemplified by the 2-Mb domain on human chromosome 15q11-q13 and its mouse orthologue on chromosome 7c (ref. 1). Loss of this 2-Mb domain on the paternal or maternal allele results in two neurogenetic disorders, Prader-Willi syndrome (PWS) or Angelman syndrome (AS), respectively. Microdeletions on the paternal allele share a 4.3-kb short region of overlap (SRO), which includes the SNRPN promoter/exon1, cause PWS and silence paternally expressed genes. Microdeletions on the maternal allele share a 0.88-kb SRO located 35 kb upstream to the SNRPN promoter, cause AS and alleviate repression of genes on the maternal allele. Individuals carrying both AS and PWS deletions on the paternal allele show a PWS phenotype and genotype. These observations suggest that cis elements within the AS-SRO and PWS-SRO constitute an imprinting box that regulates the entire domain on both chromosomes. Here we show that a minitransgene composed of a 200-bp Snrpn promoter/exon1 and a 1-kb sequence located approximately 35 kb upstream to the SNRPN promoter confer imprinting as judged by differential methylation, parent-of-origin-specific transcription and asynchronous replication. PMID- 11101842 TI - The insulin gene VNTR is associated with fasting insulin levels and development of juvenile obesity. AB - In millions of people, obesity leads to type 2 diabetes (T2D; also known as non insulin-dependent diabetes mellitus). During the early stages of juvenile obesity, the increase of insulin secretion in proportion to accumulated fat balances insulin resistance and protects patients from hyperglycaemia. After several decades, however,beta-cell function deteriorates and T2D develops in approximately 20% of obese patients. In modern societies, obesity has thus become the leading risk factor for T2D (ref. 5). The factors that predispose obese patients to alteration of insulin secretion upon gaining weight remain unknown. To determine which genetic factors predispose obese patients to beta-cell dysfunction, and possibly T2D, we studied single-nucleotide polymorphisms (SNPs) in the region of the insulin gene (INS) among 615 obese children. We found that, in the early phase of obesity, alleles of the INS variable number of tandem repeat (VNTR) locus are associated with different effects of body fatness on insulin secretion. Young obese patients homozygous for class I VNTR alleles secrete more insulin than those with other genotypes. PMID- 11101843 TI - Telomere maintenance by recombination in human cells. AB - Telomeres of eukaryotic chromosomes contain many tandem repeats of a G-rich sequence (for example, TTAGGG in vertebrates). In most normal human cells, telomeres shorten with each cell division, and it is proposed that this limits the number of times these cells can replicate. Telomeres may be maintained in germline cells, and in many immortalized cells and cancers, by the telomerase holoenzyme (first discovered in the ciliate Tetrahymena), which uses an RNA subunit as template for synthesis of telomeric DNA by the reverse transcriptase catalytic subunit. Some immortalized human cell lines and some tumours maintain their telomeres in the absence of any detectable telomerase activity by a mechanism referred to as alternative lengthening of telomeres (ALT). Here we show that DNA sequences are copied from telomere to telomere in an immortalized human ALT cell line, indicating that ALT occurs by means of homologous recombination and copy switching. PMID- 11101844 TI - Targeted inversion of a polar silencer within the HoxD complex re-allocates domains of enhancer sharing. AB - Mammalian Hox genes are clustered at four genomic loci. During development, neighbouring genes are coordinately regulated by global enhancer sequences, which control multiple genes at once, as exemplified by the expression of series of contiguous Hoxd genes in either limbs or gut. The link between vertebrate Hox gene transcription and their clustered distribution is poorly understood. Experimental and comparative approaches have revealed that various mechanisms, such as gene clustering or global enhancer sequences, might have constrained this genomic organization and stabilized it throughout evolution. To understand what restricts the effect of a particular enhancer to a precise set of genes, we generated a loxP/Cre-mediated targeted inversion within the HoxD cluster. Mice carrying the inversion showed a reciprocal re-assignment of the limb versus gut regulatory specificities, suggesting the presence of a silencer element with a unidirectional property. This polar silencer appears to limit the number of genes that respond to one type of regulation and thus indicates how separate regulatory domains may be implemented within intricate gene clusters. PMID- 11101845 TI - Fgf8 is required for outgrowth and patterning of the limbs. AB - The expression pattern and activity of fibroblast growth factor-8 (FGF8) in experimental assays indicate that it has important roles in limb development, but early embryonic lethality resulting from mutation of Fgf8 in the germ line of mice has prevented direct assessment of these roles. Here we report that conditional disruption of Fgf8 in the forelimb of developing mice bypasses embryonic lethality and reveals a requirement for Fgf8 in the formation of the stylopod, anterior zeugopod and autopod. Lack of Fgf8 in the apical ectodermal ridge (AER) alters expression of other Fgf genes, Shh and Bmp2. PMID- 11101846 TI - Fgf8 signalling from the AER is essential for normal limb development. AB - Vertebrate limb development depends on signals from the apical ectodermal ridge (AER), which rims the distal tip of the limb bud. Removal of the AER in chick results in limbs lacking distal skeletal elements. Fibroblast growth factor (FGF) proteins can substitute for the AER (refs 4-7), suggesting that FGF signalling mediates AER activity. Of the four mouse Fgf genes (Fgf4 , Fgf8, Fgf9, Fgf17) known to display AER-specific expression domains within the limb bud (AER-Fgfs), only Fgf8 is expressed throughout the AER. Moreover, Fgf8 expression precedes that of other AER-Fgfs (refs 8-13), suggesting that Fgf8 may perform unique functions early in limb development. In mice, loss of function of Fgf4 (refs 13,14), Fgf9 (D. Ornitz, pers. comm.) or Fgf17 (ref. 15) has no effect on limb formation. We report here that inactivating Fgf8 in early limb ectoderm causes a substantial reduction in limb-bud size, a delay in Shh expression, misregulation of Fgf4 expression, and hypoplasia or aplasia of specific skeletal elements. Our data identify Fgf8 as the only known AER-Fgf individually necessary for normal limb development, and provide insight into the function of Fgf signalling from the AER in the normal outgrowth and patterning of the limb. PMID- 11101847 TI - Role of the p53-homologue p73 in E2F1-induced apoptosis. AB - Most human cancers harbour aberrations of cell-cycle control, which result in deregulated activity of the E2F transcription factors with concomitant enhanced cell-cycle progression. Oncogenic signalling by E2F1 has recently been linked to stabilization and activation of the tumour suppressor p53 (refs 1,3,4). The p73 protein shares substantial sequence homology and functional similarity with p53 (refs 5-7 ). Hence, several previously considered p53-independent cellular activities may be attributable to p73. Here we provide evidence that E2F1 directly activates transcription of TP73, leading to activation of p53-responsive target genes and apoptosis. Disruption of p73 function by a tumour-derived p53 mutant reduced E2F1-mediated apoptosis. Thus, p73 activation by deregulated E2F1 activity might constitute a p53-independent, anti-tumorigenic safeguard mechanism. PMID- 11101848 TI - A major susceptibility locus for atopic dermatitis maps to chromosome 3q21. AB - Atopic dermatitis (eczema) is a chronic inflammatory skin disease with onset mainly in early childhood It is commonly the initial clinical manifestation of allergic disease, often preceding the onset of respiratory allergies. Along with asthma and allergic rhinitis, atopic dermatitis is an important manifestation of atopy that is characterized by the formation of allergy antibodies (IgE) to environmental allergens. In the developed countries, the prevalence of atopic dermatitis is approximately 15%, with a steady increase over the past decades. Genetic and environmental factors interact to determine disease susceptibility and expression, and twin studies indicate that the genetic contribution is substantial. To identify susceptibility loci for atopic dermatitis, we ascertained 199 families with at least two affected siblings based on established diagnostic criteria. A genome-wide linkage study revealed highly significant evidence for linkage on chromosome 3q21 (Zall=4.31, P= 8.42 10(-6)). Moreover, this locus provided significant evidence for linkage of allergic sensitization under the assumption of paternal imprinting (hlod=3.71,alpha=44%), further supporting the presence of an atopy gene in this region. Our findings indicate that distinct genetic factors contribute to susceptibility to atopic dermatitis and that the study of this disease opens new avenues to dissect the genetics of atopy. PMID- 11101849 TI - Absence of perilipin results in leanness and reverses obesity in Lepr(db/db) mice. AB - Obesity is a disorder of energy balance. Hormone-sensitive lipase (HSL) mediates the hydrolysis of triacylglycerol, the major form of stored energy in the body. Perilipin (encoded by the gene Plin), an adipocyte protein, has been postulated to modulate HSL activity. We show here that targeted disruption of Plin results in healthy mice that have constitutively activated fat-cell HSL. Plin -/- mice consume more food than control mice, but have normal body weight. They are much leaner and more muscular than controls, have 62% smaller white adipocytes, show elevated basal lipolysis that is resistant to beta-adrenergic agonist stimulation, and are cold-sensitive except when fed. They are also resistant to diet-induced obesity. Breeding the Plin -/- alleles into Leprdb/db mice reverses the obesity by ncreasing the metabolic rate of the mice. Our results demonstrate a role for perilipin in reining in basal HSL activity and regulating lipolysis and energy balance; thus, agents that inactivate perilipin may prove useful as anti-obesity medications. PMID- 11101850 TI - Perlecan, the major proteoglycan of basement membranes, is altered in patients with Schwartz-Jampel syndrome (chondrodystrophic myotonia). AB - Schwartz-Jampel syndrome (SJS1) is a rare autosomal recessive disorder characterized by permanent myotonia (prolonged failure of muscle relaxation) and skeletal dysplasia, resulting in reduced stature, kyphoscoliosis, bowing of the diaphyses and irregular epiphyses. Electromyographic investigations reveal repetitive muscle discharges, which may originate from both neurogenic and myogenic alterations. We previously localized the SJS1 locus to chromosome 1p34 p36.1 and found no evidence of genetic heterogeneity. Here we describe mutations, including missense and splicing mutations, of the gene encoding perlecan (HSPG2) in three SJS1 families. In so doing, we have identified the first human mutations in HSPG2, which underscore the importance of perlecan not only in maintaining cartilage integrity but also in regulating muscle excitability. PMID- 11101851 TI - MAML1, a human homologue of Drosophila mastermind, is a transcriptional co activator for NOTCH receptors. AB - Notch receptors are involved in cell-fate determination in organisms as diverse as flies, frogs and humans. In Drosophila melanogaster , loss-of-function mutations of Notch produce a 'neurogenic' phenotype in which cells destined to become epidermis switch fate and differentiate to neural cells. Upon ligand activation, the intracellular domain of Notch (ICN) translocates to the nucleus, and interacts directly with the DNA-binding protein Suppressor of hairless (Su(H)) in flies, or recombination signal binding protein Jkappa (RBP-Jkappa) in mammals, to activate gene transcription. But the precise mechanisms of Notch induced gene expression are not completely understood. The gene mastermind has been identified in multiple genetic screens for modifiers of Notch mutations in Drosophila. Here we clone MAML1, a human homologue of the Drosophila gene Mastermind, and show that it encodes a protein of 130 kD localizing to nuclear bodies. MAML1 binds to the ankyrin repeat domain of all four mammalian NOTCH receptors, forms a DNA-binding complex with ICN and RBP-Jkappa, and amplifies NOTCH-induced transcription of HES1. These studies provide a molecular mechanism to explain the genetic links between mastermind and Notch in Drosophila and indicate that MAML1 functions as a transcriptional co-activator for NOTCH signalling. PMID- 11101852 TI - A transgenic insertion upstream of sox9 is associated with dominant XX sex reversal in the mouse. AB - In most mammals, male development is triggered by the transient expression of the Y-chromosome gene, Sry, which initiates a cascade of gene interactions ultimately leading to the formation of a testis from the indifferent fetal gonad. Several genes, in particular Sox9, have a crucial role in this pathway. Despite this, the direct downstream targets of Sry and the nature of the pathway itself remain to be clearly established. We report here a new dominant insertional mutation, Odsex (Ods), in which XX mice carrying a 150-kb deletion (approximately 1 Mb upstream of Sox9) develop as sterile XX males lacking Sry. During embryogenesis, wild-type XX fetal gonads downregulate Sox9 expression, whereas XY and XX Ods/+ fetal gonads upregulate and maintain its expression. We propose that Ods has removed a long-range, gonad-specific regulatory element that mediates the repression of Sox9 expression in XX fetal gonads. This repression would normally be antagonized by Sry protein in XY embryos. Our data are consistent with Sox9 being a direct downstream target of Sry and provide genetic evidence to support a general repressor model of sex determination in mammals. PMID- 11101853 TI - A point mutation in PTPRC is associated with the development of multiple sclerosis. AB - Multiple sclerosis (MS) is the most common demyelinating disease of the central nervous system. It is widely accepted that a dysregulated immune response against brain resident antigens is central to its yet unknown pathogenesis. Although there is evidence that the development of MS has a genetic component, specific genetic factors are largely unknown. Here we investigated the role of a point mutation in the gene (PTPRC) encoding protein-tyrosine phosphatase, receptor-type C (also known as CD45) in the heterozygous state in the development of MS. The nucleotide transition in exon 4 of the gene locus interferes with mRNA splicing and results in altered expression of CD45 isoforms on immune cells. In three of four independent case-control studies, we demonstrated an association of the mutation with MS. We found the PTPRC mutation to be linked to and associated with the disease in three MS nuclear families. In one additional family, we found the same variant CD45 phenotype, with an as-yet-unknown origin, among the members affected with MS. Our findings suggest an association of the mutation in PTPRC with the development of MS in some families. PMID- 11101855 TI - Erratum PMID- 11101854 TI - Corrections PMID- 11101857 TI - 2000 subject index - volumes 24 - 26 PMID- 11101858 TI - Collaborative conflicts. PMID- 11101859 TI - Benefits of memory. PMID- 11101860 TI - Clemens Freiherr von Pirquet: explaining immune complex disease in 1906. PMID- 11101861 TI - Flying doctors. PMID- 11101862 TI - NKT cells and tumor immunity--a double-edged sword. PMID- 11101863 TI - Murder by proxy. PMID- 11101864 TI - Somatic evolution of antibody specificity: setting the clock forward. PMID- 11101865 TI - Complexity or coherence? Cytokine secretion by B cells. PMID- 11101866 TI - Immunology highlights from the recent literature. PMID- 11101867 TI - The multifaceted role of Fas signaling in immune cell homeostasis and autoimmunity. AB - Originally identified as a cell surface receptor that triggered the death of lymphocytes and tumor cells, it is now recognized that Fas (also known as CD95 or Apo-I) has distinct functions in the life and death of different cell types in the immune system. Fas signaling may also be involved in T cell costimulation and proliferation. Although Fas deficiency in humans and mice predisposes them towards systemic autoimmunity, Fas-FasL interactions can also facilitate organ specific immunopathology. Proximal signaling by Fas and related receptors depends on subunit preassembly, which accounts for the dominant-negative effect of pathogenic receptor mutants and natural splice variants. PMID- 11101868 TI - Reciprocal regulation of polarized cytokine production by effector B and T cells. AB - Although B cells produce cytokines it is not known whether B cells can differentiate into effector subsets that secrete polarized arrays of cytokines. We have identified two populations of "effector" B cells (Be1 and Be2) that produce distinct patterns of cytokines depending on the cytokine environment in which the cells were stimulated during their primary encounter with antigen and T cells. These effector B cell subsets subsequently regulate the differentiation of naive CD4+ T cells to TH1 and TH2 cells through production of polarizing cytokines such as interleukin 4 and interferon gamma. In addition, Be1 and Be2 cells could be identified in animals that were infected with pathogens that preferentially induce a Type 1 and Type 2 immune response. Together these results suggest that, in addition to their well defined role in antibody production, B cells may regulate immune responses to infectious pathogens through their production of cytokines. PMID- 11101869 TI - Control of target cell survival in thyroid autoimmunity by T helper cytokines via regulation of apoptotic proteins. AB - After autoimmune inflammation, interactions between CD95 and its ligand (CD95L) mediate thyrocyte destruction in Hashimoto's thyroiditis (HT). Conversely, thyroid autoimmune processes that lead to Graves' disease (GD) result in autoantibody-mediated thyrotropin receptor stimulation without thyrocyte depletion. We found that GD thyrocytes expressed CD95 and CD95L in a similar manner to HT thyrocytes, but did not undergo CD95-induced apoptosis either in vivo or in vitro. This pattern was due to the differential production of TH1 and TH2 cytokines. Interferon gamma promoted caspase up-regulation and CD95-induced apoptosis in HT thyrocytes, whereas interleukin 4 and interleukin 10 protected GD thyrocytes by potent up-regulation of cFLIP and Bcl-xL, which prevented CD95 induced apoptosis in sensitized thyrocytes. Thus, modulation of apoptosis-related proteins by TH1 and TH2 cytokines controls thyrocyte survival in thyroid autoimmunity. PMID- 11101870 TI - Fas triggers an alternative, caspase-8-independent cell death pathway using the kinase RIP as effector molecule. AB - Cell death is achieved by two fundamentally different mechanisms: apoptosis and necrosis. Apoptosis is dependent on caspase activation, whereas the caspase independent necrotic signaling pathway remains largely uncharacterized. We show here that Fas kills activated primary T cells efficiently in the absence of active caspases, which results in necrotic morphological changes and late mitochondrial damage but no cytochrome c release. This Fas ligand-induced caspase independent death is absent in T cells that are deficient in either Fas associated death domain (FADD) or receptor-interacting protein (RIP). RIP is also required for necrotic death induced by tumor necrosis factor (TNF) and TNF related apoptosis-inducing ligand (TRAIL). In contrast to its role in nuclear factor kappa B activation, RIP requires its own kinase activity for death signaling. Thus, Fas, TRAIL and TNF receptors can initiate cell death by two alternative pathways, one relying on caspase-8 and the other dependent on the kinase RIP. PMID- 11101871 TI - Caspase inhibitors improve survival in sepsis: a critical role of the lymphocyte. AB - Sepsis induces lymphocyte apoptosis and prevention of lymphocyte death may improve the chances of surviving this disorder. We compared the efficacy of a selective caspase-3 inhibitor to a polycaspase inhibitor and to caspase-3-/- mice. Both inhibitors prevented lymphocyte apoptosis and improved survival. Caspase-3-/- mice shared a decreased, but not total, block of apoptosis. The polycaspase inhibitor caused a very substantial decrease in bacteremia. Caspase inhibitors did not benefit RAG-1-/- mice, which had a > tenfold increase in bacteremia compared to controls. Adoptive transfer of T cells that overexpressed the anti-apoptotic protein Bcl-2 increased survival. T cells stimulated with anti CD3 and anti-CD28 produced increased interleukin 2 and interferon gamma by 6 h. Thus, caspase inhibitors enhance immunity by preventing lymphocyte apoptosis and lymphocytes act rapidly, within 24 h, to control infection. PMID- 11101872 TI - OmpA targets dendritic cells, induces their maturation and delivers antigen into the MHC class I presentation pathway. AB - We analyzed the interaction between a bacterial cell wall protein and dendritic cells (DCs). Outer membrane protein A from Klebsiella pneumoniae (kpOmpA) specifically bound to professional antigen presenting cells and was endocytosed by immature DCs via a receptor-dependent mechanism. kpOmpA signaled through Toll like receptor 2, induced DCs to produce interleukin 12 and induced maturation of DCs. Whole antigen that was coupled to kpOmpA and injected into mice was taken up by DCs and delivered to the conventional cytosolic MHC class I presentation pathway. kpOmpA also primed antigen-specific CD8+ CTLs in the absence of CD4+ T cell help or adjuvant and elicited therapeutic immunity to antigen-expressing tumors. Thus, OmpA belongs to a class of proteins that are able to elicit CTL responses to exogenous antigen. PMID- 11101873 TI - IL-6 switches the differentiation of monocytes from dendritic cells to macrophages. AB - Monocytes can give rise to either antigen presenting dendritic cells (DCs) or scavenging macrophages. This differentiation is initiated when monocytes cross the endothelium. But the regulation of DC and macrophage differentiation in tissues remains elusive. When stimulated with granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin-4 (IL-4), monocytes yield DCs. However, we show here that the addition of fibroblasts switches differentiation to macrophages. On contact with monocytes, fibroblasts release IL-6, which up regulates the expression of functional M-CSF receptors on monocytes. This allows the monocytes to consume their autocrine M-CSF. Thus, the interplay between IL-6 and M-CSF switches monocyte differentiation to macrophages rather than DCs, and IL-6 is an essential factor in the molecular control of antigen presenting cell development. PMID- 11101874 TI - NKT cell-mediated repression of tumor immunosurveillance by IL-13 and the IL-4R STAT6 pathway. AB - Using a mouse model in which tumors show a growth-regression-recurrence pattern, we investigated the mechanisms for down-regulation of cytotoxic T lymphocyte mediated tumor immunosurveillance. We found that interleukin 4 receptor (IL-4R) knockout and downstream signal transducer and activator of transcription 6 (STAT6) knockout, but not IL-4 knockout, mice resisted tumor recurrence, which implicated IL-13, the only other cytokine that uses the IL-4R-STAT6 pathway. We confirmed this by IL-13 inhibitor (sIL-13R alpha 2-Fc) treatment. Loss of natural killer T cells (NKT cells) in CD1 knockout mice resulted in decreased IL-13 production and resistance to recurrence. Thus, NKT cells and IL-13, possibly produced by NKT cells and signaling through the IL-4R-STAT6 pathway, are necessary for down-regulation of tumor immunosurveillance. IL-13 inhibitors may prove to be a useful tool in cancer immunotherapy. PMID- 11101875 TI - Immune suppression and skin cancer development: regulation by NKT cells. AB - Ultraviolet (UV) radiation is carcinogenic and immunosuppressive. UV-induced immune suppression is mediated by antigen-specific T cells, which can transfer suppression to normal recipients. These cells are essential for controlling skin cancer development in the UV-irradiated host and in suppressing other immune responses, such as delayed-type hypersensitivity. Despite their importance in skin cancer development, their exact identity has remained elusive. We show here that natural killer T cells from UV-irradiated donor mice function as suppressor T cells and play a critical role in regulating the growth of UV-induced skin cancers and suppressing adaptive immune responses in vivo. PMID- 11101876 TI - BLIMP-I mediates extinction of major histocompatibility class II transactivator expression in plasma cells. AB - Class II transactivator (CIITA), a coactivator required for class II major histocompatibility complex (MHC) transcription, is expressed in B cells but extinguished in plasma cells. This report identifies B lymphocyte-induced maturation protein I (BLIMP-I), a transcriptional repressor that is capable of triggering plasma cell differentiation, as a developmentally regulated repressor of CIITA transcription. BLIMP-I represses the B cell-specific promoter of the human gene that encodes CIITA (MHC2TA) in a binding site-dependent manner. Decreased CIITA correlates with increased BLIMP-I during plasma cell differentiation in cultured cells. Ectopic expression of BLIMP-I represses endogenous mRNA for CIITA and the CIITA targets, class II MHC, invariant chain and H2-DM (the murine equivalent of HLA-DM) in primary splenic B cells as well as 18-81 pre-B cells. Thus, the BLIMP-I program of B cell differentiation includes loss of antigen presentation via extinction of CIITA expression. PMID- 11101878 TI - Erratum. PMID- 11101877 TI - Toll-like receptor 2-mediated NF-kappa B activation requires a Rac1-dependent pathway. AB - Mammalian Toll-like receptors (TLRs) are expressed on innate immune cells and respond to the membrane components of Gram-positive or Gram-negative bacteria. When activated, they convey signals to transcription factors that orchestrate the inflammatory response. However, the intracellular signaling events following TLR activation are largely unknown. Here we show that TLR2 stimulation by Staphylococcus aureus induces a fast and transient activation of the Rho GTPases Rac1 and Cdc42 in the human monocytic cell line THP-1 and in 293 cells expressing TLR2. Dominant-negative Rac1N17, but not dominant-negative Cdc42N17, block nuclear factor-kappa B (NF-kappa B) transactivation. S. aureus stimulation causes the recruitment of active Rac1 and phosphatidylinositol-3 kinase (PI3K) to the TLR2 cytosolic domain. Tyrosine phosphorylation of TLR2 is required for assembly of a multiprotein complex that is necessary for subsequent NF-kappa B transcriptional activity. A signaling cascade composed of Rac1, PI3K and Akt targets nuclear p65 transactivation independently of I kappa B alpha degradation. Thus Rac1 controls a second, I kappa B-independent, pathway to NF-kappa B activation and is essential in innate immune cell signaling via TLR2. PMID- 11101879 TI - Peering into the review process. PMID- 11101880 TI - The promiscuous and specific sides of SecB. PMID- 11101881 TI - Specificity through flexibility. PMID- 11101882 TI - Fraternal twins: AQP1 and GlpF. PMID- 11101883 TI - Phosphothreonine recognition comes into focus. PMID- 11101884 TI - Some assembly required. PMID- 11101885 TI - Going green. PMID- 11101886 TI - Picture story. Milking it for all it's worth. PMID- 11101887 TI - Peroxisomal targeting signal-1 recognition by the TPR domains of human PEX5. AB - Many proteins contain targeting signals within their sequences that specify their delivery to particular organelles. The peroxisomal targeting signal-1 (PTS1) is a C-terminal tripeptide that is sufficient to direct proteins into peroxisomes. The PTS1 sequence closely approximates Ser-Lys-Leu-COO-. PEX5, the receptor for PTS1, interacts with the signal via a series of tetratricopeptide repeats (TPRs) within its C-terminal half. Here we report the crystal structure of a fragment of human PEX5 that includes all seven predicted TPR motifs in complex with a pentapeptide containing a PTS1 sequence. Two clusters of three TPRs almost completely surround the peptide, while a hinge region, previously identified as TPR4, forms a distinct structure that enables the two sets of TPRs to form a single binding site. This structure reveals the molecular basis for PTS1 recognition and demonstrates a novel mode of TPR-peptide interaction. PMID- 11101888 TI - A de novo designed helix-turn-helix peptide forms nontoxic amyloid fibrils. AB - We report here that a monomeric de novo designed alpha-helix-turn-alpha-helix peptide, alpha t alpha, when incubated at 37 degrees C in an aqueous buffer at neutral pH, forms nonbranching, protease resistant fibrils that are 6-10 nm in diameter. These fibrils are rich in beta-sheet and bind the amyloidophilic dye Congo red. alpha t alpha fibrils thus display the morphologic, structural, and tinctorial properties of authentic amyloid fibrils. Surprisingly, unlike fibrils formed by peptides such as the amyloid beta-protein or the islet amyloid polypeptide, alpha t alpha fibrils were not toxic to cultured rat primary cortical neurons or PC12 cells. These results suggest that the potential to form fibrils under physiologic conditions is not limited to those proteins associated with amyloidoses and that fibril formation alone is not predictive of cytotoxic activity. PMID- 11101889 TI - The Mad1-Sin3B interaction involves a novel helical fold. AB - Sin3A or Sin3B are components of a corepressor complex that mediates repression by transcription factors such as the helix-loop-helix proteins Mad and Mxi. Members of the Mad/Mxi family of repressors play important roles in the transition between proliferation and differentiation by down-regulating the expression of genes that are activated by the proto-oncogene product Myc. Here, we report the solution structure of the second paired amphipathic helix (PAH) domain (PAH2) of Sin3B in complex with a peptide comprising the N-terminal region of Mad1. This complex exhibits a novel interaction fold for which we propose the name 'wedged helical bundle'. Four alpha-helices of PAH2 form a hydrophobic cleft that accommodates an amphipathic Mad1 alpha-helix. Our data further show that, upon binding Mad1, secondary structure elements of PAH2 are stabilized. The PAH2 Mad1 structure provides the basis for determining the principles of protein interaction and selectivity involving PAH domains. PMID- 11101890 TI - Structures of two RNA domains essential for hepatitis C virus internal ribosome entry site function. AB - Translation of the hepatitis C virus (HCV) polyprotein is initiated at an internal ribosome entry site (IRES) element in the 5' untranslated region of HCV RNA. The HCV IRES element interacts directly with the 40S subunit, and biochemical experiments have implicated RNA elements near the AUG start codon as required for IRES-40S subunit complex formation. The data we present here show that two RNA stem loops, domains IIId and IIIe, are involved in IRES-40S subunit interaction. The structures of the two RNA domains were solved by NMR spectroscopy and reveal structural features that may explain their role in IRES function. PMID- 11101891 TI - Metal ion binding sites in a group II intron core. AB - Group II introns are catalytic RNA molecules that require divalent metal ions for folding, substrate binding, and chemical catalysis. Metal ion binding sites in the group II core have now been elucidated by monitoring the site-specific RNA hydrolysis patterns of bound ions such as Tb(3+) and Mg(2+). Major sites are localized near active site elements such as domain 5 and its surrounding tertiary interaction partners. Numerous sites are also observed at intron substructures that are involved in binding and potentially activating the splice sites. These results highlight the locations of specific metal ions that are likely to play a role in ribozyme catalysis. PMID- 11101892 TI - Point mutations alter the mechanical stability of immunoglobulin modules. AB - Immunoglobulin-like modules are common components of proteins that play mechanical roles in cells such as muscle elasticity and cell adhesion. Mutations in these proteins may affect their mechanical stability and thus may compromise their function. Using single molecule atomic force microscopy (AFM) and protein engineering, we demonstrate that point mutations in two beta-strands of an immunoglobulin module in human cardiac titin alter the mechanical stability of the protein, resulting in mechanical phenotypes. Our results demonstrate a previously unrecognized class of phenotypes that may be common in cell adhesion and muscle proteins. PMID- 11101893 TI - Crystal structure of a nucleosome core particle containing the variant histone H2A.Z. AB - Activation of transcription within chromatin has been correlated with the incorporation of the essential histone variant H2A.Z into nucleosomes. H2A.Z and other histone variants may establish structurally distinct chromosomal domains; however, the molecular mechanism by which they function is largely unknown. Here we report the 2.6 A crystal structure of a nucleosome core particle containing the histone variant H2A.Z. The overall structure is similar to that of the previously reported 2.8 A nucleosome structure containing major histone proteins. However, distinct localized changes result in the subtle destabilization of the interaction between the (H2A.Z-H2B) dimer and the (H3-H4)(2) tetramer. Moreover, H2A.Z nucleosomes have an altered surface that includes a metal ion. This altered surface may lead to changes in higher order structure, and/or could result in the association of specific nuclear proteins with H2A.Z. Finally, incorporation of H2A.Z and H2A within the same nucleosome is unlikely, due to significant changes in the interface between the two H2A.Z-H2B dimers. PMID- 11101894 TI - Structure of Escherichia coli exonuclease I suggests how processivity is achieved. AB - Exonuclease I (ExoI) from Escherichia coli is a monomeric enzyme that processively degrades single stranded DNA in the 3' to 5' direction and has been implicated in DNA recombination and repair. Determination of the structure of ExoI to 2.4 A resolution using X-ray crystallography verifies the expected correspondence between a region of ExoI and the exonuclease (or proofreading) domains of the DNA polymerases. The overall fold of ExoI also includes two other regions, one of which extends the exonuclease domain and another that can be described as an elaborated SH3 domain. These three regions combine to form a molecule that is shaped like the letter C, although it also contains a segment that effectively converts the C into an O-like shape. The structure of ExoI thus provides additional support for the idea that DNA metabolizing enzymes achieve processivity by completely enclosing the DNA. PMID- 11101895 TI - An evolutionary bridge to a new protein fold. AB - Arc repressor bearing the N11L substitution (Arc-N11L) is an evolutionary intermediate between the wild type protein, in which the region surrounding position 11 forms a beta-sheet, and a double mutant 'switch Arc', in which this region is helical. Here, Arc-N11L is shown to be able to adopt either the wild type or mutant conformations. Exchange between these structures occurs on the millisecond time scale in a dynamic equilibrium in which the relative populations of each fold depend on temperature, solvent conditions and ligand binding. The N11L mutation serves as an evolutionary bridge from the beta-sheet to the helical fold because in the mutant, Leu is an integral part of the hydrophobic core of the new structure but can also occupy a surface position in the wild type structure. Conversely, the polar Asn 11 side chain serves as a negative design element in wild type Arc because it cannot be incorporated into the core of the mutant fold. PMID- 11101896 TI - The structural basis for red fluorescence in the tetrameric GFP homolog DsRed. AB - Green fluorescent protein (GFP) has rapidly become a standard tool for investigating a variety of cellular activities, and has served as a model system for understanding spectral tuning in chromophoric proteins. Distant homologs of GFP in reef coral and anemone display two new properties of the fluorescent protein family: dramatically red-shifted spectra, and oligomerization to form tetramers. We now report the 1.9 A crystal structure of DsRed, a red fluorescent protein from Discosoma coral. DsRed monomers show similar topology to GFP, but additional chemical modification to the chromophore extends the conjugated pi system and likely accounts for the red-shifted spectra. Oligomerization of DsRed occurs at two chemically distinct protein interfaces to assemble the tetramer. The DsRed structure reveals the chemical basis for the functional properties of red fluorescent proteins and provides the basis for rational engineering of this subfamily of GFP homologs. PMID- 11101897 TI - Novel DNA binding domain and genetic regulation model of Bacillus subtilis transition state regulator abrB. AB - We have determined the high resolution NMR solution structure of the novel DNA binding domain of the Bacillus subtilis transition state regulator AbrB. Comparisons of the AbrB DNA binding domain with DNA binding proteins of known structure show that it is a member of a completely novel class of DNA recognition folds that employs a dimeric topology for cellular function. This new DNA binding conformation is referred to as the looped-hinge helix fold. Sequence homology investigations show that this DNA binding topology is found in other disparately related microbes. Structural analysis of the AbrB DNA binding domain together with bioanalytical and mutagenic data of full length AbrB allows us to construct a general model that describes the genetic regulation properties of AbrB. PMID- 11101898 TI - Evidence for cleft closure in actomyosin upon ADP release. AB - Structural insights into the interaction of smooth muscle myosin with actin have been provided by computer-based fitting of crystal structures into three dimensional reconstructions obtained by electron cryomicroscopy, and by mapping of structural and dynamic changes in the actomyosin complex. The actomyosin structures determined in the presence and absence of MgADP differ significantly from each other, and from all crystallographic structures of unbound myosin. Coupled to a complex movement ( approximately 34 A) of the light chain binding domain upon MgADP release, we observed a approximately 9 degrees rotation of the myosin motor domain relative to the actin filament, and a closure of the cleft that divides the actin binding region of the myosin head. Cleft closure is achieved by a movement of the upper 50 kDa region, while parts of the lower 50 kDa region are stabilized through strong interactions with actin. This model supports a mechanism in which binding of MgATP at the active site opens the cleft and disrupts the interface, thereby releasing myosin from actin. PMID- 11101899 TI - Crystal structures of ribosome anti-association factor IF6. AB - Ribosome anti-association factor eIF6 (originally named according to translation initiation terminology as eukaryotic initiation factor 6) binds to the large ribosomal subunit, thereby preventing inappropriate interactions with the small subunit during initiation of protein synthesis. We have determined the X-ray structures of two IF6 homologs, Methanococcus jannaschii archaeal aIF6 and Sacchromyces cerevisiae eIF6, revealing a phylogenetically conserved 25 kDa protein consisting of five quasi identical alpha/beta subdomains arrayed about a five-fold axis of pseudosymmetry. Yeast eIF6 prevents ribosomal subunit association. Comparative protein structure modeling with other known archaeal and eukaryotic homologs demonstrated the presence of two conserved surface regions, one or both of which may bind the large ribosomal subunit. PMID- 11101900 TI - The structure and oligomerization of the yeast arginine methyltransferase, Hmt1. AB - Protein methylation at arginines is ubiquitous in eukaryotes and affects signal transduction, gene expression and protein sorting. Hmt1/Rmt1, the major arginine methyltransferase in yeast, catalyzes methylation of arginine residues in several mRNA-binding proteins and facilitates their export from the nucleus. We now report the crystal structure of Hmt1 at 2.9 A resolution. Hmt1 forms a hexamer with approximate 32 symmetry. The surface of the oligomer is dominated by large acidic cavities at the dimer interfaces. Mutation of dimer contact sites eliminates activity of Hmt1 both in vivo and in vitro. Mutating residues in the acidic cavity significantly reduces binding and methylation of the substrate Npl3. PMID- 11101901 TI - Crystal structure of the bacterial protein export chaperone secB. AB - SecB is a bacterial molecular chaperone involved in mediating translocation of newly synthesized polypeptides across the cytoplasmic membrane of bacteria. The crystal structure of SecB from Haemophilus influenzae shows that the molecule is a tetramer organized as a dimer of dimers. Two long channels run along the side of the molecule. These are bounded by flexible loops and lined with conserved hydrophobic amino acids, which define a suitable environment for binding non native polypeptides. The structure also reveals an acidic region on the top surface of the molecule, several residues of which have been implicated in binding to SecA, its downstream target. PMID- 11101904 TI - 2000 subject index - volume 7 PMID- 11101905 TI - Zidovudine (ZDV) and Lamivudine (3TC) Combination Therapy for HIV Infection - A Review. AB - The current state of knowledge regarding the treatment of HIV infection supports the use of antiretroviral agents in such a way as to maximize the likelihood of viral suppression for as long as possible. This goal is more likely to be achieved with the combined use of two nucleoside reverse transcriptase inhibitors (NRTIs) and a potent protease inhibitor (PI). Theoretically, there are 10 different ways of combining 2 NRTIs using the 5 NRTIs currently available on the market. However, some combinations are not recommended, e.g. zidovudine (ZDV) plus stavudine (d4T) or didanosine (ddI) plus zalcitabine (ddC), due to metabolism and toxicity considerations. The ingredients of an ideal combination should be highly potent as isolated drugs, should have different metabolic pathways, different target cells (activated versus resting cells), no overlapping toxicity and resistance profiles, and, of increasing importance, they should be easy to take with infrequent daily dosing in order to optimize patient compliance. The combination of ZDV and lamivudine (3TC) is probably the best documented association of 2 NRTIs. The combination has been through a full development plan and it has displayed many of the required characteristics. The combination of ZDV/ 3TC is thus considered a reliable component of any potent antiviral regimen aiming to maximize viral suppression. PMID- 11101902 TI - Structural basis of the recognition of the dishevelled DEP domain in the Wnt signaling pathway. AB - The DEP domain of Dishevelled (Dvl) proteins transduces signals to effector proteins downstream of Dvl in the Wnt pathway. Here we report that DEP-containing mutants inhibit Wnt-induced, but not Dvl-induced, activation of the transcription factor Lef-1. This inhibitory effect is weakened by a K434M mutation. Nuclear magnetic resonance spectroscopy revealed that the DEP domain of mouse Dvl1 comprises a three-helix bundle, a beta-hairpin 'arm' and two short beta-strands at the C-terminal region. Lys 434 is located at the tip of the beta-hairpin 'arm'. Based on our findings, we conclude that DEP interacts with regulators upstream of Dvl via a strong electric dipole on the molecule's surface created by Lys 434, Asp 445 and Asp 448; the electric dipole and the putative membrane binding site are at two different locations. PMID- 11101906 TI - Stavudine: A Review. AB - Stavudine was the fourth nucleoside analog to be approved in the United States for the management of HIV-1 infections. It is a dideoxy analog of thymidine and is structurally similar to zidovudine. In adults, the usual dose is 40mg twice daily and probably does not have to be altered based on body weight unless the patient shows signs of toxicity from the drug, or has renal insufficiency. Clinical studies indicate that is very well tolerated, and, hematological toxicity is infrequent. Its dose limiting adverse effect is peripheral neuropathy, which is reversible in most cases with dose reduction or discontinuation. It has been shown to have synergistic activity against HIV-1 in vitro when combined with didanosine, lamivudine, nevirapine, and saquinavir. During clinical trials it has been shown that stavudine in combination with other antiviral drugs can reduce the viral load to 2 log(10) copies/ml. Recently it has been observed that Stavudine may have an increased side-effect profile and reduced efficacy when combined with zidovudine, and, therefore, is best used as an alternative to zidovudine in combination drug regimens. PMID- 11101907 TI - Evaluation of the Antimicrobial Activity of Sparfloxacin, Relative to Other Oral Antibiotics Against 1,125 Bacterial Isolates from 10 Medical Centers in Brazil. AB - A multicenter study was carried out in order to compare the in vitro activity of sparfloxacin to ciprofloxacin, amoxicillin/clavulanic acid, cephalexin, cefuroxine and azithromycin, against 1,125 microorganisms recently isolated from clinical specimens, most of them representative of respiratory tract infections. Sparfloxacin demonstrated potent action and was more active than the beta-lactam agents and azithromycin against most of the bacterial strains tested. Sparfloxacin was more potent (96% and 95% sensitivity, with MIC(90) of 0.19ug/mL and 0.5ug/mL, respectively)than the other antimicrobial agents tested against the Enterobacteriaceae family (Escherichia coli and Elebsiella pneumoniae). It was found to be equivalent in activity to ciprofloxacin (96% and 91% sensitivity and MIC(90) of 0.25 and 0.75ug/mL, respectively). Sparfloxacin was also found to be very active against the most fastidious microorganisms commonly associated to respiratory tract infections such as the penicillin-susceptible and resistant Streptococcus pneumoniae (MIC(90) 0.5ug/mL and 0.38ug/mL, respectively), ampicillin-susceptible and -resistant Haemophilus influenzae (MIC(90) 0.016ug/mL and 0.38ug/mL, respectively), beta-lactamase producing Moraxella catarrhalis (MIC(90) 0.032ug/mL) and non beta-lactamase producing M.catarrhalis (MIC(90) 0.5ug/mL). PMID- 11101908 TI - Evaluation of the in vitro Activity of Cefprozil Relative to Other Oral Antimicrobial Agents Against Bacteria in Clinical Specimens from Patients in Sao Paulo With Respiratory Tract Infections. AB - Cefprozil is a new oral second generation cephalosporin. Its in vitro antimicrobial activity was evaluated against 371 recent clinical isolates from patients with respiratory infections. We tested the susceptibility of 244 streptococci (96 Streptococcus pneumoniae, 105 group viridans streptococci, 32 Streptococcus agalactiae, and 11 group A beta-hemolitic streptococci) 107 Haemophilus influenzae, and 20 oxacillin-susceptible S.aureus (OSSA). The isolates were susceptibility tested against cefprozil, cefaclor and amoxicillin/clavulanic acid by the E-test method; and against cefadroxil, cefuroxime, cefetamet, erythromycin, and azythromycin by disk diffusion. The methods and the susceptibility categorization followed the National Committee for Clinical Laboratory Standards (NCCLS) procedures. Amoxicillin/clavulanic acid was slightly more active againstH.influenzae (MICs(90) 0.5ug/mL) than cefprozil or cefaclor (MICs(90) 4 and 2ug/mL respectively). Cefprozil demonstrated potent activity against streptococci. Against S.pneumoniae, cefprozil was 2-4 fold more active than cefaclor (MICs(90)0.125 and 0.38ug/mL, respectively). S. pneumoniae susceptibility was 84% to penicillin, 95% to erythromycin and 97% to azithromycin by disk diffusion. Viridans streptococci showed higher MICs for cefprozil and cefaclor (MICs(90) 4ug/mL and 8ug/mL, respectively) and only 50% susceptibility to the macrolides. Cefprozil was four times more active than cefaclor and as active as amoxieillin/clavulanic acid against group A beta-hemolytic streptococci and S.agalactiae. These three compounds showed similar activity against OSSA. In conclusion, cefprozil demonstrated excellent in vitro activity against bacterial species responsible for respiratory infections in Sao Paulo. PMID- 11101910 TI - Incentives and Penalties in Health Care: Getting the Best from Pharmaceutical Development. PMID- 11101909 TI - Nosocomial Primary Bacteremia: Clinical and Laboratory Characteristics in Adults and Children. AB - A prospective study was carried out to evaluate the main clinical and laboratory manifestations of nosocomial primary bacteremia among adults and children at the Heart Institute, University of Sao Paulo School of Medicine, Brazil during 1993. Forty occurrences of bacteremia were analyzed; 27 in adults and 13 in children. Among adults, although fever, rigors and alterations in leukocyte count were frequently recorded, rigors and fever were absent in 30% of the cases and the frequency of any associated clinical manifestations was only 55%. Among children, fever occurred most frequently, but rigors were not recorded. It is concluded that the diagnosis of nosocomial bacteremia is imprecise due to frequent absence of the main clinical correlates of bacteremia. Blood cultures should be obtained in patients who are in a setting associated with bacteremia, even if typical clinical signs and symptoms are not presented, and that frequent updating of guidelines for empiric treatment is needed. We also suggest that separate guidelines be established for children and adults because of the differences in the clinical findings in these two groups. PMID- 11101911 TI - Microbial Pathogens Associated With Cystic Fibrosis: Special Focus on Pseudomonas aeruginosa. AB - Cystic fibrosis (CF) is the most common inherited lethal disease among Caucasians. The disease is characterized by a high sodium sweat concentration, malabsorption, malnutrition, and chronic bronchopulmonary infection. Although CF is a multisystem disorder it is the deterioration of lung function, due mainly to bacterial colonization, that is the major determinant contributing to the high morbidity and mortality of affected patients. A relatively large variety of microbial species can be recovered from the CF sputum but it is widely accepted that Staphylococcus aureus, Haemophilus influenzae, Pseudomonas aeruginosa, and Burkholderia cepacia are the most important organisms causing infection in CF patients. Among these, P.aruginosa is the most prevalent pathogen responsible for much of the severe chronic lung infection. Antibiotic therapy aimed at clearing or reducing, the bacterial load in the CF lung, even though temporary, increases the longevity of patients and is one of the mainstays in treatment the disease. PMID- 11101912 TI - Statistical Inference (part 1): Basic Concepts. AB - In the most common research situations, the investigator cannot directly assess the whole population of interest. For this reason, a sample is often studied to infer the actual population measures or parameters. Statistical inference comprises the application of methods to analyze the sample data in order to estimate the population parameters. The basic assumption in statistical inference is that each individual within the population of interest has the same probability of being included in a specific sample. When the sample is not randomly selected. the study findings can still be generalized if the sample can be considered representative of the whole population of interest. A set of statistical methods used to infer the population parameters is performed under the assumption that the sample estimates follow a bell-shaped distribution, called normal distribution. This article presents with the help of examples, the logic used in the sampling distribution theory. The concept of normal (also called gaussian) sampling distribution has an important role in statistical inference, even when the population values are not normally distributed. In fact, in the statistical inference process, the form of the distribution of the sample estimates is more important than the distribution of the individual values. PMID- 11101913 TI - Co-Infection by HTLV-I/II is Associated With Increased Viral Load in PBMC of HIV 1 Infected Patients in Bahia, Brazil. AB - To evaluate the viral load in peripheral blood mononucler cells (PBMC) of patients infected by HIV-1 we performed 96 quantitative cultures in 74 patients infected by HIV-1, using the co-culture method. The viral load was expressed in tissue culture infectious doses (TCID), and the results were analyzed according to gender, age and clinical stage of patients, duration of previous antiretroviral therapy, detectable p24 antigenemia, CD4(+)/CD8(+) cell counts, co infection by HTLV-I/II and viral subtype. We detected a statistically significant association between co-infection by HTLV-I/II and viral load higher than >50 TCID (p=0.003). We also found a significant association between co-infection by HTLV I/II and p24 antigenemia (p=0.028). CD4(+) cell counts were significantly higher for patients presenting negative cultures, but there was no detectable association between lower CD4(+) cell counts and higher TCID. The majority of patients were infected by subtype B virus. The observation in this study that co infection with HTLV-I/II was significantly associated with higher viral load raises the possibility that these agents act as co-factors of AIDS progression, in doubly-infected patients. PMID- 11101914 TI - Staphylococcus aureus Nasopharyngeal Carriage Rates and Antimicrobial Susceptibility Patterns Among Health Care Workers and Their Household Contacts. AB - Dissemination of Staphylococcus aureus within hospitals by nasopharyngeal carriage of the organism by health care workers (HCW) has been well characterized for over 40 years, but physicians and nurses must be reminded of the extent of the problem. To determine the level of colonization among HCW in one hospital in Brazil, and to examine the potential spread to household contacts and the surrounding community, nasal swabs for S. aureus were done on 200 HCW, 87 household contacts, and 77 members of the community. The frequency of positive cultures in each group was recorded, and the organims were then tested for susceptibility to a panel of antibiotics. The average level of antibiotic resistance was calculated for each organism using a scoring technique termed rate of bacterial resistance (RBR). Phage typing was also done. The frequency of colonization was 63/200 (31.5%) among HCW, 27/87 (31%) among their household contacts, and 14/77 (18.1%) in members of the community (p>0.05). The level of antibiotic resistance (RBR) was significantly higher among HCW than among household contacts or the community. Phage typing revealed that 40.7% of isolates had a common phage pattern between HCW and household contacts. Among household contacts, the level of antimicrobial resistance was the same for the shared phage types as for the unique types. We conclude that nasopharyngeal carriage among HCW remains a problem, that the carriage rate is also seen among household contacts, but not in the community. Increased levels of antimicrobial resistance in the strains carried by HCW indicate that the spread of resistant organisms occurs by this mechanism. Careful control of S. aureus among HCW is an important hospital practice. PMID- 11101915 TI - Percutaneous Injuries With Sharp Instruments and the Behavior of Anesthesiologists and Obstetricians in Regard to the Associated Risk of Occupational Infectious Diseases: A Survey in a Town in Brazil. AB - Health care workers are at risk of acquiring several infectious diseases, including human immunodeficiency virus (HIV) infection, from percutaneous injuries with contaminated needles and sharp instruments. A survey among anesthesiologists and obstetricians in a Brazilian town assessed their risk of these injuries and their behavior toward prophylaxis. Both anesthesiologists and obstetricians tended to be more adherent to universal precautions when caring for patients with known HIV positive status, although the difference was statistically significant only for the use of goggles. Of those surveyed, 80% of the obstetricians and 65% of anesthesiologists had at least 1 episode of injury with a needle or sharp instrument in the last 12 months. Injuries were mostly from syringe needles (88.9%) among the anesthesiologists, and from suture needles (56.9%) among the obstetricians. The annual risk of injury was much higher among obstetricians (2 per 100 procedures) than among anesthesiologists (11 per 10,000 anesthesias and 17 per 10,000 venous punctures/intravenous catheterizations). The majority of the anesthesiologists and obstetricians referred for vaccination against hepatitis B received the vaccine as post-exposure prophylaxis. Promoting recognition of percutaneous injuries with sharp instruments among health care workers as an important occupational health problem, distribution of information on methods to prevent such injuries, and when to immunize are objectives yet to be achieved to prevent the transmission of infection by HIV and other microbes. Proper assessment of a number of new devices, techniques, and protective equipment may result in more effective preventive measures in the future. PMID- 11101916 TI - Characteristics of Isolates Streptococcus pneumoniae from Middle-Aged and Elderly Adults in Brazil: Capsular Serotypes and Antimicrobial Sensitivity to Invasive Infections. AB - Pneumococcal infection cause frequent, serious problems among middle aged and elderly populations worldwide. Efforts to prevent mortality caused by pneumococci are based mainly on rapid diagnosis of the infection and appropriate antimicrobial therapy. Vaccination is considered to be the best approach to prevent the disease in at risk populations and the current 23-valent pneumococcal polysaccharide vaccine is recommended for people >e;65 years of age. To evaluate the present antibiotic sensitivity patterns and the potential for vaccine use to prevent this disease in Brazil, 94 isolates of pneumococci from normally sterile body sites from patients >e;50 years of age were analyzed for capsular serotypes and antimicrobial resistance. Among the total isolates, 7.4% (n = 7) of the isolates showed intermediate level resistance to penicillin (IR). Among the IR isolates, 6 were also resistant to trimethoprim-sulfamethoxazole. All isolates were susceptible to cefotaxime, chloramphenicol, erythromycin, and vancomycin. Twenty-eight serotypes were identified and the serotypes included in the 23-valent vaccine accounted for 81.7%. By adding the cross-reacting serotypes, the percentage of vaccine preventable infections was 90.3%. Each of intermediate level resistant strains were among the 23 serotypes included in pneumococcal vaccine. This preliminary data on the distribution of serotype of S.pneumoniae among those >e;50 years of age in Brazil, and the potential for increased antimicrobial resistance to penicillin and trimethoprim sulfamethoxazole, support the use of pneumococcal vaccine in this population. PMID- 11101917 TI - Scabies Norwegian Associated With High IgE and Low IgG1 Levels Presenting as Systemic Lupus Erythematosus. AB - Norwegian type scabies is a severe skin infestation which has been described in both immunosuppressed and immunocompetent people. We report a case of Systemic Lupus Erythematosus (SLE) presenting as Norwegian scabies in a malnourished patient who had exceedingly high levels of serum IgE and low IgG 1. The CD(4) count was normal but the ratio CD(4)/CD(8) was decreased as a result of an increased CD(8) count. The extensive skin disease which led to the diagnosis of Systemic Lupus Erythematosus was unresponsive to treatment until the SLE was properly controlled by therapy. The possible relationship between high IgE and low IgG1 levels and Norwegian scabies is discussed in the context of systemic lupus erythematosus. PMID- 11101918 TI - Will Immunotherapy and Vaccination Replace Antimicrobials? The Future May Be Closer Than We Think! PMID- 11101919 TI - Phase transition in a traffic model with passing AB - We investigate a traffic model in which cars either move freely with quenched intrinsic velocities or belong to clusters formed behind slower cars. In each cluster, the next-to-leading car is allowed to pass and resume free motion. The model undergoes a phase transition from a disordered phase for the high passing rate to a jammed phase for the low rate. In the disordered phase, the cluster size distribution decays exponentially in the large size limit. In the jammed phase, the distribution of finite clusters is independent on the passing rate, but it accounts only for a fraction of all cars; the "excessive" cars form an infinite cluster moving with the smallest velocity. Mean-field equations, describing the model in the framework of Maxwell approximation, correctly predict the existence of phase transition and adequately describe the disordered phase; properties of the jammed phase are studied numerically. PMID- 11101920 TI - Reaction-diffusion models describing a two-lane traffic flow AB - A unidirectional two-lane road is approximated by a set of two parallel closed one-dimensional chains. Two types of cars, i.e., slow and fast ones are considered in the system. Based on the Nagel-Schreckenberg model of traffic flow [K. Nagel and M. Schreckenberg, J. Phys. 2, 2221 (1992)], a set of reaction diffusion processes is introduced to simulate the behavior of the cars. Fast cars can pass the slow ones using the passing lane. We write and solve the mean-field rate equations for the density of slow and fast cars, respectively. We also investigate the properties of the model through computer simulations and obtain the fundamental diagrams. A comparison between our results and the v(max)=2 version of the Nagel-Schreckenberg model is made. PMID- 11101921 TI - Percolation parameter and percolation-threshold estimates for three-dimensional random ellipses with widely scattered distributions of eccentricity and size AB - In fractured materials of very low matrix permeability, fracture connectivity is the first-order determinant of the occurrence of flow. For systems having a narrow distribution of object sizes (short-range percolation), a first-order percolation criterion is given by the total excluded volume which is almost constant at threshold. In the case of fractured media, recent observations have demonstrated that the fracture-length distribution is extremely large. Because of this widely scattered fracture-length distribution, the classical expression of the total excluded volume is no longer scale invariant at the percolation threshold and has no finite limit for infinitely large systems. Thus, the classical estimation method of the percolation threshold established in short range percolation becomes useless for the connectivity determination of fractured media. In this study, we derive an expression for the total excluded volume that remains scale invariant at the percolation threshold and that can thus be used as the proper control parameter, called the parameter of percolation in percolation theory. We show that the scale-invariant expression of the total excluded volume is the geometrical union normalized by the system volume rather than the summation of the mutual excluded volumes normalized by the system volume. The summation of the mutual excluded volume (classical expression) remains linked to the number of intersections between fractures, whereas the normalized geometrical union of the mutual excluded volume (our expression) can be essentially identified with the percolation parameter. Moreover, fluctuations of this percolation parameter at threshold with length and eccentricity distributions remain limited within a range of less than one order of magnitude, giving in turn a rough percolation criterion. We finally show that the scale dependence of the percolation parameter causes the connectivity of fractured media to increase with scale, meaning especially that the hydraulic properties of fractured media can dramatically change with scale. PMID- 11101922 TI - Quantum kinetic equation in the closed-time-path formalism AB - A systematic derivation of the quantum kinetic equation is presented in the framework of a closed-time-path formalism. Introducing a probe, the expectation value of the number operator is calculated as a functional of the probing source. Then, solving for the source by inverting the relation, the removal of the source leads to the quantum kinetic equation as the equation of motion for the number, which gives a generalization of the Boltzmann equation including memory. The inversion formula is used in the course of the derivation. The calculation is presented up to third order in interaction, and the effect of initial correlations is also considered. PMID- 11101923 TI - Spinodal decomposition and the tomita sum rule AB - The scaling properties of a phase-ordering system with a conserved order parameter are studied. The theory developed leads to scaling functions satisfying certain general properties including the Tomita sum rule. The theory also gives good agreement with numerical results for the order parameter scaling function in three dimensions. The values of the associated nonequilibrium decay exponents are given by the known lower bounds. PMID- 11101924 TI - Dynamics of the frustrated ising lattice gas AB - The dynamical properties of a three-dimensional model glass, the frustrated Ising lattice gas (FILG), are studied by Monte Carlo simulations. We present results of compression experiments, where the chemical potential is either slowly or abruptly changed, as well as simulations at constant density. One time quantities like density and two time quantities like correlations, responses, and mean square displacements are measured, and the departure from equilibrium clearly characterized. The aging scenario, particularly in the case of density autocorrelations, is reminiscent of spin glass phenomenology with violations of the fluctuation-dissipation theorem, typical of systems with one replica symmetry breaking. The FILG, as a valid on-lattice model of structural glasses, can be described with tools developed in spin glass theory and, being a finite dimensional model, can open the way for a systematic study of activated processes in glasses. PMID- 11101925 TI - Feedback-controlled forcing of meandering spiral waves in an open gel reactor AB - The control of spiral wave dynamics has attracted much interest recently. Besides external forcing at a fixed frequency, feedback-controlled forcing has been studied. One of the simplest feedback schemes is to measure the activity level at a particular point of the medium continuously and to apply a spatially uniform pulsatory modulation if a certain threshold is reached at the detector. We have realized this feedback loop using an open gel reactor for the light-sensitive Belousov-Zhabotinskii medium. This allows us to maintain stationary nonequilibrium conditions over several hundred rotation periods of the unperturbed spiral wave. By varying the distance between the detector and the spiral tip, we find different stable branches of the resonance attractor for the same medium. We confirm the constant spacing between neighboring stable branches as predicted by Zykov and Karma and measure the dependence of the resonance drift on the forcing amplitude. PMID- 11101926 TI - Temperature of nonequilibrium steady-state systems AB - We determined the operational temperatures of nonequilibrium-molecular-dynamics (NEMD) systems by the recently developed thermometer [A. Baranyai, Phys. Rev. E 61, R3306 (2000)] and compared these values to the dynamic temperatures [H. H. Rough, Phys. Rev. Lett. 78, 772 (1997)] of the same systems. NEMD models use a synthetic thermostat, a numerical feedback procedure to remove the dissipative heat instantaneously. A consequence of this feedback is a splitting of the dynamic temperature. The kinetic part is different from the configurational part because the energy is removed through the momentum subspace of the system. In addition to this, these temperature values also vary with respect to the direction of the external perturbation. In the case of planar Couette flow and color flow, the isotropic operational temperatures of dense liquids are always closer to the configurational than to the kinetic temperatures. We show that the observed split and the pronounced directional dependence of the dynamic temperature is an artifact caused by the instantaneous feedback of NEMD models. Since relaxation of a preset difference between the kinetic and the configurational temperature is an order of magnitude faster than the relaxation of the heat flux vector, for models with realistic thermostas such a split must be very small. We argue that in real systems, even far from equilibrium, the operational temperature and both terms of the dynamic temperature must be practically identical and isotropic. PMID- 11101927 TI - Self-replicating pulses and sierpinski gaskets in excitable media AB - In our previous papers, we have shown by computer simulations that a Sierpinski gasket pattern appears in a Bonhoeffer-van der Pol type reaction-diffusion system. In this paper, we show another class of regular self-similar structure which is found in four different excitable reaction-diffusion systems. This result strongly implies that the existence of the self-similar spatiotemporal evolution is universal in excitable reaction-diffusion media. PMID- 11101928 TI - Cluster diversity and entropy on the percolation model: the lattice animal identification algorithm AB - We present an algorithm to identify and count different lattice animals (LA's) in the site-percolation model. This algorithm allows a definition of clusters based on the distinction of cluster shapes, in contrast with the well-known Hoshen Kopelman algorithm, in which the clusters are differentiated by their sizes. It consists in coding each unit cell of a cluster according to the nearest neighbors (NN) and ordering the codes in a proper sequence. In this manner, a LA is represented by a specific code sequence. In addition, with some modification the algorithm is capable of differentiating between fixed and free LA's. The enhanced Hoshen-Kopelman algorithm [J. Hoshen, M. W. Berry, and K. S. Minser, Phys. Rev. E 56, 1455 (1997)] is used to compose the set of NN code sequences of each cluster. Using Monte Carlo simulations on planar square lattices up to 2000x2000, we apply this algorithm to the percolation model. We calculate the cluster diversity and cluster entropy of the system, which leads to the determination of probabilities associated with the maximum of these functions. We show that these critical probabilities are associated with the percolation transition and with the complexity of the system. PMID- 11101929 TI - First-order phase transition in a nonequilibrium growth process AB - We introduce a simple continuous model for nonequilibrium surface growth. The dynamics of the system is defined by the Kardar-Parisi-Zhang equation with a Morse-like potential representing a short range interaction between the surface and the substrate. The mean field solution displays a nontrivial phase diagram with a first-order transition between a growing and a bound surface, associated with a region of coexisting phases, and a tricritical point where the transition becomes second order. Numerical simulations in three dimensions show quantitative agreement with mean field results, and the features of the phase space are preserved even in two dimensions. PMID- 11101930 TI - Unattainability of carnot efficiency in the brownian heat engine AB - We discuss the reversibility of the Brownian heat engine. We perform an asymptotic analysis of the Kramers equation on a Buttiker-Landauer system and show quantitatively that Carnot efficiency is unattainable even in the fully overdamped limit. The unattainability is attributed to inevitable irreversible heat flow over the temperature boundary. PMID- 11101931 TI - Nontrivial extension of the two-dimensional ising model: the d-dimensional "molecular" model AB - A recently proposed molecular model is discussed as a nontrivial extension of the Ising model. For d=2 the two models are shown to be equivalent, while for d>2 the molecular model describes a peculiar second order transition from an isotropic high-temperature phase to a low-dimensional anisotropic low-temperature state. The general mean-field analysis is compared with the results achieved by a variational Migdal-Kadanoff real space renormalization group method and by standard Monte Carlo sampling for d=3. By finite size scaling the critical exponent has been found to be nu=0.44+/-0.02, thus establishing that the molecular model does not belong to the universality class of the Ising model for d>2. PMID- 11101932 TI - Self-organized localization of macrostates by additive noise in a fast-slow dynamical system: effect of the slow nonreactive mode on the barrier crossing rate of the fast, bistable mode AB - We have studied the stochastic dynamics of a two-dimensional gradient system composed of a fast, bistable mode and a slow, monotonically decaying mode. The coupling is bidirectional and cooperative. Additive white noise acts on the fast mode only. We find that the noise intensity controls the location of macrostates (shape of the probability density function), the appearance of bimodality in the slow-mode probability distribution and, together with the coupling strength, the rate of fast-mode barrier crossing. These features arise from the interplay of noise, widely separated time scales, and bidirectional, excitatory coupling. They are believed to be generic. PMID- 11101933 TI - Local minimal energy landscapes in river networks AB - The existence and stability of a universality class associated with local minimal energy landscapes is investigated. Using extensive numerical simulations, we first study the dependence on a parameter gamma of a partial differential equation, which was proposed to describe the evolution of a rugged landscape toward a local minimum of the dissipated energy. We then compare the results with those obtained by an evolution scheme based on a variational principle (the optimal channel networks). It is found that both models yield qualitatively similar river patterns and similar dependences on gamma. However, the aggregation mechanism is strongly dependent on the value of gamma. A careful analysis suggests that scaling behaviors may depend weakly on both gamma and on initial conditions, but in all cases are within observational data predictions. Consequences of our results are finally discussed, and the most plausible scenario is presented. PMID- 11101934 TI - Strong-property-fluctuation theory for homogenization of bianisotropic composites: formulation AB - The strong-property-fluctuation theory is developed for the homogenization of the linear dielectric, magnetic, and magnetoelectric properties of a two-constituent bianisotropic composite. The notion of a bianisotropic comparison medium (BCM) is introduced to serve as a springboard for the Dyson equation satisfied by the ensemble-averaged electromagnetic field. With the constitutive properties of the BCM serving as the zeroth-order solution of the Dyson equation, the first-order correction, known as the bilocal approximation, is obtained. Wave propagation in the composite can be described in this manner by a nonlocal effective medium containing information about the spatial correlations of the constitutive properties. For scales larger than the correlation length, the nonlocality vanishes and a local effective medium emerges. Analytical results for the local effective constitutive properties are presented after assuming a spherical particulate topology for the constituent mediums. Illustrative numerical results are provided. PMID- 11101935 TI - Mean first passage time for anomalous diffusion AB - When the random force acting on a particle diffusing in an interval [0,L] and subjected to a constant external force is a Gaussian white noise, the "Brownian" mean-squared displacement is described by the seminal relation =2Dt(gamma) with gamma=1. However, for more complicated random forces the diffusion may be slower (gamma<1, "subdiffusion") or faster (gamma>1, "superdiffusion") than the "normal" diffusion. For both these cases we calculated the mean free passage time (MFPT)-the time needed to reach one of the traps at boundaries. The simple formulas for the different diffusive regimes are compared quantitatively for the simplest case of the absence of an external field and for an initial position in the middle of the interval. It turns out that the MFPT's for anomalous diffusion can be both larger or smaller than that for normal diffusion depending on the values of the length of the interval and the diffusion coefficient. Moreover, the MFPT can show nonmonotonic changes with the degree of departure from normal diffusion. PMID- 11101936 TI - Reaction-controlled diffusion AB - The dynamics of a coupled two-component nonequilibrium system is examined by means of continuum field theory representing the corresponding master equation. Particles of species A may perform hopping processes only when particles of different type B are present in their environment. Species B is subject to diffusion-limited reactions. If the density of B particles attains a finite asymptotic value (active state), the A species displays normal diffusion. On the other hand, if the B density decays algebraically approximately t(-alpha) at long times (inactive state), the effective attractive A-B interaction is weakened. The combination of B decay and activated A hopping processes gives rise to anomalous diffusion, with mean-square displacement (A)(t)(2)> approximately t(1-alpha) for alpha<1. Such algebraic subdiffusive behavior ensues for nth-order B annihilation reactions (nB-->) with n>/=3, and n=2 for d<2. The mean-square displacement of the A particles grows only logarithmically with time in the case of B pair annihilation (n=2) and d>/=2 dimensions. For radioactive B decay (n=1), the A particles remain localized. If the A particles may hop spontaneously as well, or if additional random forces are present, the A-B coupling becomes irrelevant, and conventional diffusion is recovered in the long-time limit. PMID- 11101937 TI - Phase space geometry and stochasticity thresholds in hamiltonian dynamics AB - Results of numerical computations of the largest Lyapunov exponent lambda(1)(varepsilon,N) as a function of the energy density varepsilon and the number of particles N are here reported for a Fermi-Pasta-Ulam alpha+beta model. These results show the coexistence at large N of two thresholds: a stochasticity threshold, found before for the alpha model alone, and a strong stochasticity threshold (SST), found before for the beta model alone. Although this coexistence may seem at first sight plausible, it is not obvious a priori that the alpha+beta model superimposes properties of the alpha and beta models independently. The main point of this paper, however, is a geometric characterization of the SST via the mean curvature of the constant energy hypersurfaces in the phase space of the model and the characteristic decay time of its time autocorrelation function tau(c)(varepsilon,N), which correlates with that of lambda(1)(varepsilon,N) for fixed N. This appears to provide important information on the very complicated geometry of the phase space of this simple solidlike model. PMID- 11101938 TI - Order-parameter model for unstable multilane traffic flow AB - We discuss a phenomenological approach to the description of unstable vehicle motion on multilane highways that explains in a simple way the observed sequence of the "free flow <--> synchronized mode <--> jam" phase transitions as well as the hysteresis in these transitions. We introduce a variable called an order parameter that accounts for possible correlations in the vehicle motion at different lanes. So, it is principally due to the "many-body" effects in the car interaction in contrast to such variables as the mean car density and velocity being actually the zeroth and first moments of the "one-particle" distribution function. Therefore, we regard the order parameter as an additional independent state variable of traffic flow. We assume that these correlations are due to a small group of "fast" drivers and by taking into account the general properties of the driver behavior we formulate a governing equation for the order parameter. In this context we analyze the instability of homogeneous traffic flow that manifested itself in the above-mentioned phase transitions and gave rise to the hysteresis in both of them. Besides, the jam is characterized by the vehicle flows at different lanes which are independent of one another. We specify a certain simplified model in order to study the general features of the car cluster self-formation under the "free flow <--> synchronized motion" phase transition. In particular, we show that the main local parameters of the developed cluster are determined by the state characteristics of vehicle motion only. PMID- 11101939 TI - Elasticity of gaussian and nearly gaussian phantom networks AB - We study the elastic properties of phantom networks of Gaussian and nearly Gaussian springs. We show that the stress tensor of a Gaussian network coincides with the conductivity tensor of an equivalent resistor network, while its elastic constants vanish. We use a perturbation theory to analyze the elastic behavior of networks of slightly non-Gaussian springs. We show that the elastic constants of phantom percolation networks of nearly Gaussian springs have a power-law dependence on the distance of the system from the percolation threshold, and we derive bounds on the exponents. PMID- 11101940 TI - Establishing the relation between detrended fluctuation analysis and power spectral density analysis for stochastic processes AB - Stochastic fractal signals can be characterized by the Hurst coefficient H, which is related to the exponents of various power-law statistics characteristic of these processes. Two techniques widely used to estimate H are spectral analysis and detrended fluctuation analysis (DFA). This paper examines the analytical link between these two measures and shows that they are related through an integral transform. Numerical simulations confirm this relationship for ideal synthesized fractal signals. Their performance as estimators of H is compared based on a mean square error criterion and found to be similar. DFA measures are derived for physiological signals of heartbeat R-R intervals through the integral transform of a spectral density estimate. These agree with directly calculated DFA estimates, indicating that the relationship holds for signals with nonideal fractal properties. It is concluded that DFA and spectral measures provide equivalent characterizations of stochastic signals with long-term correlation. PMID- 11101941 TI - 1/f(alpha) noise from self-organized critical models with uniform driving AB - Using the well-known Olami-Feder-Christensen model as our paradigm, we show how to modify uniform driven self-organized critical models to generate 1/f(alpha) noise. This model can reproduce all the main features of 1/f(alpha) noise: (1) alpha is close to one and does not depend on the dimension of the system. (2) The 1/f(alpha) behavior is found for very low frequencies. (3) The spatial correlations do not obey a power law. That proves that spatially extended systems based on activation-deactivation processes do not have to be point-driven to produce 1/f(alpha) noise. The essential ingredient is a local memory of the activation-deactivation process. PMID- 11101942 TI - Kinetics of ordering in fluctuation-driven first-order transitions: simulation and theory AB - Many systems involving competing interactions or interactions that compete with constraints are well described by a model first introduced by Brazovskii [Zh. Eksp. Teor. Fiz. 68, 175 (1975) [Sov. Phys. JETP 41, 85 (1975)]]. The hallmark of this model is that the fluctuation spectrum is isotropic and has a maximum at a nonzero wave vector represented by the surface of a d-dimensional hypersphere. It was shown by Brazovskii that the fluctuations change the free energy structure from a straight phi(4) to a straight phi(6) form with the disordered state metastable for all quench depths. The transition from the disordered phase to the periodic lamellar structure changes from second order to first order and suggests that the dynamics is governed by nucleation. Using numerical simulations we have confirmed that the equilibrium free energy function is indeed of a straight phi(6) form. A study of the dynamics, however, shows that, following a deep quench, the dynamics is described by unstable growth rather than nucleation. A dynamical calculation, based on a generalization of the Brazovskii calculations, shows that the disordered state can remain unstable for a long time following the quench. PMID- 11101943 TI - Variety and volatility in financial markets AB - We study the price dynamics of stocks traded in a financial market by considering the statistical properties of both a single time series and an ensemble of stocks traded simultaneously. We use the n stocks traded on the New York Stock Exchange to form a statistical ensemble of daily stock returns. For each trading day of our database, we study the ensemble return distribution. We find that a typical ensemble return distribution exists in most of the trading days with the exception of crash and rally days and of the days following these extreme events. We analyze each ensemble return distribution by extracting its first two central moments. We observe that these moments fluctuate in time and are stochastic processes, themselves. We characterize the statistical properties of ensemble return distribution central moments by investigating their probability density functions and temporal correlation properties. In general, time-averaged and portfolio-averaged price returns have different statistical properties. We infer from these differences information about the relative strength of correlation between stocks and between different trading days. Last, we compare our empirical results with those predicted by the single-index model and we conclude that this simple model cannot explain the statistical properties of the second moment of the ensemble return distribution. PMID- 11101944 TI - Nonlinear relaxation field in charged systems under high electric fields AB - The influence of an external electric field on the current in charged systems is investigated. The results beyond linear response from the classical hierarchy of density matrices are compared with the results from quantum kinetic theory. It is found that even an infinitesimal friction with the background changes the results in a noncontinuous way. The kinetic theory yields a systematic treatment of the nonlinear current beyond linear response. To this end the dynamically screened and field-dependent Lenard-Balescu equation is integrated analytically and the nonlinear relaxation field is calculated. The classical linear response result known as the Debye-Onsager relaxation effect is obtained only if asymmetric screening is assumed. When considering the kinetic equation of one species, the other species have to be screened dynamically while the screening with the same species itself has to be performed statically. Other different approximations are discussed and compared. PMID- 11101945 TI - Crossover phenomenon in self-organized critical sandpile models AB - We consider a stochastic sandpile where the sand grains of unstable sites are randomly distributed to the nearest neighbors. Increasing the value of the threshold condition the stochastic character of the distribution is lost and a crossover to the scaling behavior of a different sandpile model takes place where the sand grains are equally transferred to the nearest neighbors. The crossover behavior is analyzed numerically in detail; especially we consider the exponents which determine the scaling behavior. PMID- 11101946 TI - Nonlinear stochastic resonance: the saga of anomalous output-input gain AB - We reconsider stochastic resonance (SR) for an overdamped bistable dynamics driven by a harmonic force and Gaussian noise from the viewpoint of the gain behavior, i.e., the signal-to-noise ratio (SNR) at the output divided by that at the input. The primary issue addressed in this work is whether a gain exceeding unity can occur for this archetypal SR model, for subthreshold signals that are beyond the regime of validity of linear response theory: in contrast to nondynamical threshold systems, we find that the nonlinear gain in this conventional SR system exceeds unity only for suprathreshold signals, where SR for the spectral amplification and/or the SNR no longer occurs. Moreover, the gain assumes, at weak to moderate noise strengths, rather small (minimal) values for near-threshold signal amplitudes. The SNR gain generically exhibits a distinctive nonmonotonic behavior versus both the signal amplitude at fixed noise intensity and the noise intensity at fixed signal amplitude. We also test the validity of linear response theory; this approximation is strongly violated for weak noise. At strong noise, however, its validity regime extends well into the large driving regime above threshold. The prominent role of physically realistic noise color is studied for exponentially correlated Gaussian noise of constant intensity scaling and also for constant variance scaling; the latter produces a characteristic, resonancelike gain behavior. The gain for this typical SR setup is further contrasted with the gain behavior for a "soft" potential model. PMID- 11101947 TI - Thermally activated breakdown in the fiber-bundle model AB - Guarino et al., (cond-Mat/9908329) have recently introduced a fiber bundle model where fiber fracture can be thermally activated. Under a fixed (subcritical) loading, the mean failure time of the bundle is studied. An analytical expression for the latter is obtained as a function of the load. The effect of a (narrow) quenched disorder in the fracture stress of the fibers with a Gaussian distribution is shown to lead to an effective temperature simply translated with respect to the actual one. Finally, some "critical" properties of fracture precursors which have been proposed are investigated within the present model. PMID- 11101948 TI - Random networks created by biological evolution. AB - We investigate a model of an evolving random network, introduced by us previously [Phys. Rev. Lett. 83, 5587 (1999)]. The model is a generalization of the Bak Sneppen model of biological evolution, with the modification that the underlying network can evolve by adding and removing sites. The behavior and the averaged properties of the network depend on the parameter p, the probability to establish a link to the newly introduced site. For p=1 the system is self-organized critical, with two distinct power-law regimes with forward-avalanche exponents tau=1.98+/-0.04 and tau(')=1.65+/-0.05. The average size of the network diverges as a powerlaw when p-->1. We study various geometrical properties of the network: the probability distribution of sizes and connectivities, size and number of disconnected clusters, and the dependence of the mean distance between two sites on the cluster size. The connection with models of growing networks with a preferential attachment is discussed. PMID- 11101949 TI - Stochastic dynamics of time correlation in complex systems with discrete time AB - In this paper we present the concept of description of random processes in complex systems with discrete time. It involves the description of kinetics of discrete processes by means of the chain of finite-difference non-Markov equations for time correlation functions (TCFs). We have introduced the dynamic (time dependent) information Shannon entropy S(i)(t) where i=0,1,2,3,ellipsis, as an information measure of stochastic dynamics of time correlation (i=0) and time memory (i=1,2,3,ellipsis). The set of functions S(i)(t) constitute the quantitative measure of time correlation disorder (i=0) and time memory disorder (i=1,2,3,ellipsis) in complex system. The theory developed started from the careful analysis of time correlation involving dynamics of vectors set of various chaotic states. We examine two stochastic processes involving the creation and annihilation of time correlation (or time memory) in details. We carry out the analysis of vectors' dynamics employing finite-difference equations for random variables and the evolution operator describing their natural motion. The existence of TCF results in the construction of the set of projection operators by the usage of scalar product operation. Harnessing the infinite set of orthogonal dynamic random variables on a basis of Gram-Shmidt orthogonalization procedure tends to creation of infinite chain of finite-difference non-Markov kinetic equations for discrete TCFs and memory functions (MFs). The solution of the equations above thereof brings to the recurrence relations between the TCF and MF of senior and junior orders. This offers new opportunities for detecting the frequency spectra of power of entropy function S(i)(t) for time correlation (i=0) and time memory (i=1,2,3,ellipsis). The results obtained offer considerable scope for attack on stochastic dynamics of discrete random processes in a complex systems. Application of this technique on the analysis of stochastic dynamics of RR intervals from human ECG's shows convincing evidence for a non-Markovian phenomemena associated with a peculiarities in short- and long-range scaling. This method may be of use in distinguishing healthy from pathologic data sets based in differences in these non-Markovian properties. PMID- 11101950 TI - Critical behavior and conservation in directed sandpiles AB - We perform large-scale simulations of directed sandpile models with both deterministic and stochastic toppling rules. Our results show the existence of two distinct universality classes. We also provide numerical simulations of directed models in the presence of bulk dissipation. The numerical results indicate that the way in which dissipation is implemented is irrelevant for the determination of the critical behavior. The analysis of the self-affine properties of avalanches shows the existence of a subset of superuniversal exponents, whose value is independent of the universality class. This feature is accounted for by means of a phenomenological description of the energy balance condition in these models. PMID- 11101951 TI - Mesoscopic dynamics of microcracks AB - The mesoscopic concept is applied to the description of microcracks. The balance equations of the cracked continuum result in mesoscopic directional balances of mass, momentum, angular momentum, and energy. Averaging over the length of the cracks gives the corresponding orientational balances. A further averaging process leads to the macroscopic balance equations of the microcracked continua. Dynamic equations for the fabric tensors of different order are derived using a multipole moment expansion of the orientational crack distribution function. The simple example of Griffith cracks is treated. The role of physical assumptions in the microcrack representations and the different macroscopic internal variable representations of microcracks are discussed. PMID- 11101953 TI - Similarity solutions and collapse in the attractive gross-pitaevskii equation AB - We analyze a generalized Gross-Pitaevskii (GP) equation involving a paraboloidal trap potential in D space dimensions and generalized to a nonlinearity of order 2n+1. For attractive coupling constants collapse of the particle density occurs for Dn>/=2 and typically to a delta function centered at the origin of the trap. By introducing a special variable for the spherically symmetric solutions, we show that all such solutions are self-similar close to the center of the trap. Exact self-similar solutions occur if, and only if, Dn=2, and for this case of Dn=2 we exhibit an exact but rather special D=1 analytical self-similar solution collapsing to a delta function which, however, recovers and collapses periodically, while the ordinary GP equation in two space dimensions also has a special solution with periodic delta function collapses and revivals of the density. The relevance of these various results to attractive Bose-Einstein condensation in spherically symmetric traps is discussed. PMID- 11101952 TI - Ziff-gulari-barshad model with random distribution of inert sites AB - A random distribution of inert sites is introduced in the Ziff-Gulari-Barshad model to study the phase transitions between active and poisoned states. The adsorption of CO and O2 molecules is not possible at the position of the inert sites. This model is investigated in the site and pair approximations, as well as through Monte Carlo simulations. We determine the mean coverages of the elements as a function of the dilution and show that the continuous transition between the active and O-poisoned states is slightly affected by moderate values of dilution in the pair approximation and in the simulations. On the other hand, from the analysis of the hysteresis curves, the transition between the active and CO poisoned states changes from first order to continuous as one increases the concentration of inactive sites. The observed transition in the site and pair approximations is always of first-order nature. We also found the lines of transition and spinodal points as a function of the concentration of inert sites. Finally, the production rate of CO2 is calculated as a function of the dilution of sites. PMID- 11101954 TI - Asymptotic step profiles from a nonlinear growth equation for vicinal surfaces AB - We study a recently proposed nonlinear evolution equation describing the collective step meander on a vicinal surface subject to the Bales-Zangwill growth instability [O. Pierre-Louis et al., Phys. Rev. Lett. 80, 4221 (1998)]. A careful numerical analysis shows that the dynamically selected step profile consists of sloped segments, given by an inverse error function and steepening as sqrt[t], which are matched to pieces of a stationary (time-independent) solution describing the maxima and minima. The effect of smoothening by step-edge diffusion is included heuristically, and a one-parameter family of evolution equations is introduced that contains relaxation by step-edge diffusion and by attachment-detachment as special cases. The question of the persistence of an initially imposed meander wavelength is investigated in relation to recent experiments. PMID- 11101955 TI - Generalized chapman-kolmogorov equation: A unifying approach to the description of anomalous transport in external fields AB - The generalized Chapman-Kolmogorov equation [V. M. Kenkre, E. W. Montroll, and M. F. Shlesinger, J. Stat. Phys. 9, 45 (1973)] is discussed. It is demonstrated that this equation unifies recently proposed kinetic equations of fractional order that describe anomalous transport in external fields, as well as continuous time random walks. The conditions under which the individual models can be established are discussed. PMID- 11101956 TI - Extremal-point densities of interface fluctuations in a quenched random medium AB - We give a number of exact, analytical results for the stochastic dynamics of the density of local extrema (minima and maxima) of linear Langevin equations and solid-on-solid lattice growth models driven by spatially quenched random noise. Such models can describe nonequilibrium surface fluctuations in a spatially quenched random medium, diffusion in a random catalytic environment, and polymers in a random medium. In spite of the nonuniversal character for the quantities studied, their behavior against the variation of the microscopic length scale can present generic features, characteristic of the macroscopic observables of the system. PMID- 11101957 TI - Short-time dynamics of an ising system on fractal structures AB - The short-time critical relaxation of an Ising model on a Sierpinski carpet is investigated using Monte Carlo simulation. We find that when the system is quenched from high temperature to the critical temperature, the evolution of the order parameter and its persistence probability, the susceptibility, and the autocorrelation function all show power-law scaling behavior at the short-time regime. The results suggest that the spatial heterogeneity and the fractal nature of the underlying structure do not influence the scaling behavior of the short time critical dynamics. The critical temperature, dynamic exponent z, and other equilibrium critical exponents beta and nu of the fractal spin system are determined accurately using conventional Monte Carlo simulation algorithms. The mechanism for short-time dynamic scaling is discussed. PMID- 11101958 TI - Scaling behaviors of branched polymers AB - We study the thermodynamic behavior of branched polymers. We first study random walks in order to clarify the thermodynamic relation between the canonical ensemble and the grand canonical ensemble. We then show that correlation functions for branched polymers are given by those for straight phi(3) theory with a single mass insertion, not those for the straight phi(3) theory themselves. In particular, the two-point function behaves as 1/p(4), not as 1/p(2), in the scaling region. This behavior is consistent with the fact that the Hausdorff dimension of the branched polymer is 4. In the appendixes we derive the exact two-point correlation function at finite (but large) system size N, which is consistent with the analysis in the body of the paper. PMID- 11101959 TI - Small-world networks: links with long-tailed distributions AB - Small-world networks (SWN), obtained by randomly adding to a regular structure additional links (AL), are of current interest. In this paper we explore (based on physical models) a new variant of SWN, in which the probability of realizing an AL depends on the chemical distance between the connected sites. We assume a power-law probability distribution and study random walkers on the network, focusing especially on their probability of being at the origin. We connect the results to Levy flights, which follow from a mean-field variant of our model. PMID- 11101960 TI - Finite-precision stationary states at and away from equilibrium AB - We study the precision dependence of equilibrium and nonequilibrium phase-space distribution functions for time-reversible dynamical systems simulated with finite, computational precision. The conservative and dissipative cases show different behavior, with substantially reduced period lengths in the dissipative case. The main effect of finite precision is to change the phase-space fraction occupied by the distributions. The convergence of thermodynamic averages is only slightly affected. We introduce and discuss a simple stochastic model which is nicely consistent with all of our numerical results. This model links the length of periodic orbits to the strange attractor's correlation dimension. PMID- 11101961 TI - Surmounting oscillating barriers: path-integral approach for weak noise AB - We consider the thermally activated escape of an overdamped Brownian particle over a potential barrier in the presence of periodic driving. A time-dependent path-integral formalism is developed which allows us to derive asymptotically exact weak-noise expressions for both the instantaneous and the time-averaged escape rate. Our results comprise a conceptually different, systematic treatment of the rate prefactor multiplying the exponentially leading Arrhenius factor. Moreover, an estimate for the deviations at finite noise strengths is provided and a supersymmetry-type property of the time-averaged escape rate is verified. For piecewise parabolic potentials, the rate expression can be evaluated in closed analytical form, while in more general cases, as exemplified by a cubic potential, an action-integral remains to be minimized numerically. Our comparison with very accurate numerical results demonstrates an excellent agreement with the theoretical predictions over a wide range of driving strengths and driving frequencies. PMID- 11101962 TI - Characteristic relations of type-I intermittency in the presence of noise AB - Near the point of tangent bifurcation, the scaling properties of the laminar length of type-I intermittency are investigated in the presence of noise. Based on analytic and numerical studies, we show that the scaling relation of the laminar length is dramatically deformed from 1/sqrt[epsilon] for epsilon>0 to exp(1/D)|epsilon|(3/2) for epsilon<0 as epsilon passes the bifurcation point (epsilon=0). The results explain why two coupled Rossler oscillators exhibit deformation of the scaling relation of the synchronous length in the nearly synchronous regime. PMID- 11101963 TI - Quantum recurrences: probe to study quantum chaos AB - We study the phase space of periodically modulated gravitational cavity by means of quantum recurrence phenomena. We report that the quantum recurrences serve as a tool to connect phase space of the driven system with a spectrum in the quantum domain. With the help of quantum recurrences we investigate the quasienergy spectrum of the system for a certain fixed modulation strength. In addition, we study transition of spectrum from discrete to continuum as a function of modulation strength. PMID- 11101964 TI - Symmetry breaking and dynamical independence in a multimode laser AB - Multimode lasers display various behaviors caused by the asymmetry between the modes belonging to orthogonal polarizations. We discuss dynamical independence, clustering, and grouping in a solid state laser with intracavity second harmonic generation, and show that these effects result from unstable cycles lying within their invariant planes. These invariant planes are dynamically independent. The sequential or random itinerancy of limit unstable cycles lying within the invariant planes explains most of the effects caused by asymmetry. PMID- 11101965 TI - Characteristics of the wave function of coupled oscillators in semiquantum chaos AB - Using the method of adiabatic invariants and the Born-Oppenheimer approximation, we have calculated the ground-state wave function of a pair of coupled oscillators in so-called semiquantum chaos. Some interesting characteristics, e.g., the similarities and differences between the wave functions in the regular and chaotic states have been found. Time-correlation functions of the wave functions and their Fourier spectra in two states have also been investigated. The sensitivity of the wave function in the chaotic state to the initial conditions has been identified. PMID- 11101966 TI - Modeling maps by using rational functions AB - Rational functions are not very useful for obtaining global differential models because they involve poles that may eject the trajectory to infinity. In contrast, it is here shown that they allow one to significantly improve the quality of models for maps. In such a case, the presence of poles does not involve any numerical difficulty when the models are iterated. The models then take advantage of the ability of rational functions to capture complicated structures that may be generated by maps. The method is applied to experimental data from copper electrodissolution. PMID- 11101967 TI - Hierarchy and stability of partially synchronous oscillations of diffusively coupled dynamical systems AB - The paper presents a qualitative analysis of an array of diffusively coupled identical continuous time dynamical systems. The effects of full, partial, antiphase, and in-phase-antiphase chaotic synchronizations are investigated via the linear invariant manifolds of the corresponding differential equations. The existence of various invariant manifolds, a self-similar behavior, and a hierarchy and embedding of the manifolds of the coupled system are discovered. Sufficient conditions for the stability of the invariant manifolds are obtained via the method of Lyapunov functions. Conditions under which full global synchronization cannot be achieved even for the largest coupling constant are defined. The general rigorous results are illustrated through examples of coupled Lorenz-like and Rossler systems. PMID- 11101968 TI - Integral behavior for localized synchronization in nonidentical extended systems AB - We report the synchronization of two nonidentical spatially extended fields, ruled by one-dimensional complex Ginzburg-Landau equations. The two fields are prepared in different dynamical regimes, and interact via an imperfect coupling consisting of a given number of local controllers N(c). The strength of the coupling is ruled by the parameter varepsilon. We show that, in the limit of three controllers per correlation length, the synchronization behavior is not affected if the product varepsilonN(c)/N is kept constant, providing a sort of integral behavior for localized synchronization. PMID- 11101969 TI - Phase-space structure of a thermoreceptor. AB - We analyze the phase-space structure of a model for thermoreceptors in fish and mammals. As a function of the temperature we identify a period doubling scenario at low temperatures, a regime where an unstable stationary fixed point collides with the attractor and blocks the thermoreceptor, and a transition from period n+1 to period n as the temperature is further increased. The period reduction phenomenon is due to an autoresonance between fast and slow ion channels and shows the features typical for mode locking. PMID- 11101970 TI - Convergence to the critical attractor of dissipative maps: log-periodic oscillations, fractality, and nonextensivity AB - For a family of logisticlike maps, we investigate the rate of convergence to the critical attractor when an ensemble of initial conditions is uniformly spread over the entire phase space. We found that the phase-space volume occupied by the ensemble W(t) depicts a power-law decay with log-periodic oscillations reflecting the multifractal character of the critical attractor. We explore the parametric dependence of the power-law exponent and the amplitude of the log-periodic oscillations with the attractor's fractal dimension governed by the inflection of the map near its extremal point. Further, we investigate the temporal evolution of W(t) for the circle map whose critical attractor is dense. In this case, we found W(t) to exhibit a rich pattern with a slow logarithmic decay of the lower bounds. These results are discussed in the context of nonextensive Tsallis entropies. PMID- 11101971 TI - Emergence of quantum chaos in the quantum computer core and how to manage it AB - We study the standard generic quantum computer model, which describes a realistic isolated quantum computer with fluctuations in individual qubit energies and residual short-range interqubit couplings. It is shown that in the limit where the fluctuations and couplings are small compared to the one-qubit energy spacing, the spectrum has a band structure, and a renormalized Hamiltonian is obtained which describes the eigenstate properties inside one band. Studies are concentrated on the central band of the computer ("core") with the highest density of states. We show that above a critical interqubit coupling strength, quantum chaos sets in, leading to a quantum ergodicity of the computer eigenstates. In this regime the ideal qubit structure disappears, the eigenstates become complex, and the operability of the computer is quickly destroyed. We confirm that the quantum chaos border decreases only linearly with the number of qubits n, although the spacing between multiqubit states drops exponentially with n. The investigation of time evolution in the quantum computer shows that in the quantum chaos regime, an ideal (noninteracting) state quickly disappears, and exponentially many states become mixed after a short chaotic time scale for which the dependence on system parameters is determined. Below the quantum chaos border an ideal state can survive for long times, and an be used for computation. The results show that a broad parameter region does exist where the efficient operation of a quantum computer is possible. PMID- 11101972 TI - Asymptotically stable phase synchronization revealed by autoregressive circle maps AB - A specially designed of nonlinear time series analysis is introduced based on phases, which are defined as polar angles in spaces spanned by a finite number of delayed coordinates. A canonical choice of the polar axis and a related implicit estimation scheme for the potentially underlying autoregressive circle map (next phase map) guarantee the invertibility of reconstructed phase space trajectories to the original coordinates. The resulting Fourier approximated, invertibility enforcing phase space map allows us to detect conditional asymptotic stability of coupled phases. This comparatively general synchronization criterion unites two existing generalizations of the old concept and can successfully be applied, e.g., to phases obtained from electrocardiogram and airflow recordings characterizing cardiorespiratory interaction. PMID- 11101973 TI - Spatiotemporal stability of one-way open coupled nonlinear systems AB - The present paper investigates the spatiotemporal stability of homogeneous solutions in one-way open coupled nonlinear systems. We show that the H(infinity) norm concept, which has been used as an important index in the field of robust control theory, allows us to grasp the mechanism of spatial instability in coupled systems. Spatial instability occurs only when the H(infinity) norm of the transfer function of each site is greater than 1. It is shown that numerical simulations for one-way open coupled double scroll circuits are in good agreement with our theoretical results. PMID- 11101974 TI - Atoms in parallel fields: analysis with diffractive periodic orbits AB - We show that fluctuations in the density of states of nonhydrogenic atoms in parallel fields are strongly influenced by diffractive periodic orbits. Unlike typical systems with a diffractive point scatterer, the atomic core of small atoms like lithium and helium is best understood as a combined geometric and diffractive scatterer. Each Gutzwiller (geometric) periodic orbit is paired with a diffractive orbit of the same action. We investigate, particularly, amplitudes for contributions from repetitions, and multiple scattering orbits. We find that periodic orbit repetitions are described by "hybrid" orbits, combining both diffractive and geometric core scatters, and that by including all possible permutations we can obtain excellent agreement between the semiclassical model and accurate fully quantal calculations. For high repetitions, we find even one scatter diffractive contributions become of the same order as those of the geometric periodic orbit for repetition numbers n approximately Planck's over 2pi(-1/2). Although the contribution of individual diffractive orbits is suppressed by O(Planck's over 2pi(1/2)) relative to the geometric periodic orbits, the proliferation of diffractive orbits with increasing period means that the diffractive effect for the atom can persist in the Planck's over 2pi-->0 limit. PMID- 11101975 TI - Failure of chaos control AB - We study the control of chaos in an experiment on a parametrically excited pendulum whose excitation mechanism is not perfect. This imperfection leads to a weakly excited degree of freedom with an associated small eigenvalue. Although the state of the pendulum could be characterized well and although the perturbation is weak, we fail to control chaos. From a numerical model we learn that the small eigenvalue cannot be ignored when attempting control. However, the estimate of this eigenvalue from an (experimental) time series is elusive. The reason is that points in an experimental time series are distributed according to the natural measure. It is this extremely uneven distribution of points that thwarts attempts to measure eigenvalues that are very different. Another consequence of the phase-space distribution of points for control is the occurrence of logarithmic-oscillations in the waiting time before control can be attempted. We come to the conclusion that chaos needs to be destroyed before the information needed for its control can be obtained. PMID- 11101976 TI - Synchronous chaos in coupled map lattices with small-world interactions AB - In certain physical situations, extensive interactions arise naturally in systems. We consider one such situation, namely, small-world couplings. We show that, for a fixed fraction of nonlocal couplings, synchronous chaos is always a stable attractor in the thermodynamic limit. We point out that randomness helps synchronization. We also show that there is a size dependent bifurcation in the collective behavior in such systems. PMID- 11101977 TI - Oscillatory clusters in a model of the photosensitive belousov-zhabotinsky reaction system with global feedback AB - Oscillatory cluster patterns are studied numerically in a reaction-diffusion model of the photosensitive Belousov-Zhabotinsky reaction supplemented with a global negative feedback. In one- and two-dimensional systems, families of cluster patterns arise for intermediate values of the feedback strength. These patterns consist of spatial domains of phase-shifted oscillations. The phase of the oscillations is nearly constant for all points within a domain. Two-phase clusters display antiphase oscillations; three-phase clusters contain three sets of domains with a phase shift equal to one-third of the period of the local oscillation. Border (nodal) lines between domains for two-phase clusters become stationary after a transient period, while borders drift in the case of three phase clusters. We study the evolving border movement of the clusters, which, in most cases, leads to phase balance, i.e., equal areas of the different phase domains. Border curling of three-phase clusters results in formation of spiral clusters-a combination of a fast oscillating cluster with a slow spiraling movement of the domain border. At higher feedback coefficient, irregular cluster patterns arise, consisting of domains that change their shape and position in an irregular manner. Localized irregular and regular clusters arise for parameters close to the boundary between the oscillatory region and the reduced steady state region of the phase space. PMID- 11101978 TI - Topology of high-dimensional chaotic scattering AB - We investigate Hamiltonian chaotic scattering in physically realistic three dimensional potentials. We find that the basin topology of the scattering dynamics can undergo a metamorphosis from being totally disconnected to being connected as a system parameter, such as the particle energy, is varied through a critical value. The dynamical origin of the metamorphosis is investigated, and the topological change in the scattering basin is explained in terms of the change in the structure of the invariant set of nonescaping orbits. A dynamical consequence of this metamorphosis is that the fractal dimension of the chaotic set responsible for the chaotic scattering changes its behavior characteristically at the metamorphosis. This topological metamorphosis has no correspondence in two-degree-of-freedom open Hamiltonian systems. PMID- 11101979 TI - Quasidiagonal approach to the estimation of lyapunov spectra for spatiotemporal systems from multivariate time series AB - We describe methods of estimating the entire Lyapunov spectrum of a spatially extended system from multivariate time-series observations. Provided that the coupling in the system is short range, the Jacobian has a banded structure and can be estimated using spatially localized reconstructions in low embedding dimensions. This circumvents the "curse of dimensionality" that prevents the accurate reconstruction of high-dimensional dynamics from observed time series. The technique is illustrated using coupled map lattices as prototype models for spatiotemporal chaos and is found to work even when the coupling is not strictly local but only exponentially decaying. PMID- 11101980 TI - Disorder can eliminate oscillator death AB - We use an array of diffusively coupled limit-cycle oscillators with a regular monotonic trend of natural frequencies to demonstrate that the disorder introduced in the form of random deviations from a linear trend of frequencies can weaken considerably desynchronization-induced oscillator death and, as a result, increase oscillation intensity substantially. There exist definite optimal levels of magnitude of spatial disorder at which maximal oscillatory energy is attained in the array. PMID- 11101981 TI - Parametric resonance energy exchange and induction phenomenon in a one dimensional nonlinear oscillator chain AB - We study analytically the induction phenomenon in the Fermi-Pasta-Ulam beta oscillator chain under initial conditions consisting of single mode excitation. Our study is based on the analytical computation of the largest characteristic exponent of an approximate version of the variational equation. The main results can be summarized as follows: (1) the energy density epsilon scaling of the induction time T is given by T approximately epsilon(-1), and T becomes smaller for higher-frequency mode excitation; (2) there is a threshold energy density epsilon(c) such that the induction time diverges when epsiloninfinity; (3) the threshold epsilon(c) vanishes as epsilon(c) approximately N-2 in the limit N-->infinity; (4) the threshold epsilon(c) does not depend on the mode number k that is excited in the initial condition; (5) the two modes k+/-m have the largest exponential growth rate, and m increases with increasing epsilon as m/N=sqrt[3betaepsilon]/pi. The above analytical results are thoroughly verified in numerical experiments. Moreover, we discuss the energy exchange process after the induction period in some energy density regimes, based on the numerical results. PMID- 11101982 TI - Phase synchronization and noise-induced resonance in systems of coupled oscillators AB - We study synchronization and noise-induced resonance phenomena in systems of globally coupled oscillators, each possessing finite inertia. The behavior of the order parameter, which measures the collective synchronization of the system, is investigated as the noise level and the coupling strength are varied, and hysteretic behavior is manifested. The power spectrum of the phase velocity is also examined and the quality factor as well as the response function is obtained to reveal noise-induced resonance behavior. PMID- 11101983 TI - Stochastic resonance on a circle without excitation: physical investigation and peak frequency formula AB - In this article the existence of stochastic resonance (SR) without external force in a simplified circular system for different values of the control parameter b is considered. The average power spectra are calculated as well as the signal-to noise ratio as a measure for stochastic resonance. It is shown that in the monostable and semistable (b<1 and b=1) cases coherent oscillations occur and SR exists. For the case b>1, the system is oscillatory and noise plays only a destructive role; therefore no SR occurs. The rotation number of the system is calculated and compared to the peak frequency of the power spectrum. Although the coincidence in the noisy case is not as good as that in the deterministic case, we can derive an empirical formula between the peak frequency of the power spectrum and the rotation number of the system, which is in good agreement with results of numerical simulations. PMID- 11101984 TI - Classical statistical mechanics of a few-body interacting spin model AB - We study the emergence of Boltzmann's law for the "single-particle energy distribution" in a closed system of interacting classical spins. It is shown that for a large number of particles Boltzmann's law may occur, even if the interaction is very strong. Specific attention is paid to classical analogs of the average shape of quantum eigenstates and "local density of states," which are very important in quantum chaology. Analytical predictions are then compared with numerical data. PMID- 11101985 TI - Comparison of the bifurcation scenarios predicted by the single-mode and multimode semiconductor laser rate equations AB - We present a detailed comparison of the bifurcation scenarios predicted by single mode and multimode semiconductor laser rate equation models under large amplitude injection current modulation. The influence of the gain model on the predicted dynamics is investigated. Calculations of the dependence of the time averaged longitudinal mode intensities on modulation frequency are compared with experiments performed on an Al(x)Ga(1-x)As Fabry-Perot semiconductor laser. PMID- 11101986 TI - Estimation of the entropy of the solar wind flow AB - We analyze a time series of velocities of the low-speed stream measured by the Helios spacecraft in the inner heliosphere, which is the region of space dominated by the solar wind flow. We use a nonlinear filter to give a faithful representation of the solar wind nonlinear behavior. We have demonstrated that the influence of noise in the data can be much more efficiently reduced by a nonlinear filter than with the conventional (linear) filters, and this allows a more realistic calculation of the solar wind entropy. The resulting Kolmogorov entropy is positive, and possibly the largest Lyapunov exponent is also positive locally, which would exhibit exponential sensitivity to initial conditions. Thus, these results show that the solar wind in the inner heliosphere is most likely a deterministic chaotic system. PMID- 11101987 TI - Largest lyapunov-exponent estimation and selective prediction by means of simplex forecast algorithms AB - Limited predictability is one of the remarkable features of deterministic chaos and this feature may be quantized in terms of Lyapunov exponents. Accordingly, Lyapunov-exponent estimates may be expected to follow in a natural way from forecast algorithms. Exploring this idea, we propose a method estimating the largest Lyapunov exponent from a time series which uses the behavior of so-called simplex forecasts. The method considers the estimation of properties of the distribution of local simplex expansion coefficients. These are also used for the definition of error bars for the Lyapunov-exponent estimates and allows for selective forecasts with improved prediction accuracy. We demonstrate these concepts on standard test examples and three realistic applications to time series concerning largest Lyapunov-exponent estimation of an experimentally obtained hyperchaotic NMR signal, brain state differentiation, and stock-market prediction. PMID- 11101988 TI - Correlation between the kolmogorov-sinai entropy and the self-diffusion coefficient in simple liquids AB - Molecular dynamics simulations were performed for soft- and hard-sphere systems, for number densities ranging from 0.5 to 1.0, and the Kolmogorov-Sinai entropy (KS entropy) and self-diffusion coefficients were calculated. It is found that the KS entropy, when expressed in terms of average collision frequency, is uniquely related to the self-diffusion coefficient by a simple scaling law. The dependence of the KS entropy on average collision frequency and number density was also explored. Numerical results show that the scaling laws proposed by Dzugutov, and by Beijeren, Dorfman, Posch, and Dellago, can be applied to both soft- and hard-sphere systems by changing to more generalized forms. PMID- 11101989 TI - Internal waves excited by the marangoni effect AB - Traveling periodic internal wave trains are generated in liquid layers during the absorption process of a miscible surface-active substance out of the vapor phase. In our nonstationary experimental runs, internal waves are excited by surface waves, which had been previously generated by a surface-tension-gradient-driven instability. The internal wave trains adjust their wave number by an Eckhaus instability. Close to the instability threshold narrow and extended pulses are observed. Furthermore, the wave trains can alter their traveling direction, i.e., one wave train traveling in one direction yields to another train, in general of different wave number, traveling in the opposite direction. PMID- 11101990 TI - Multiple-scattering effects on smooth neutron-scattering spectra AB - Elastic and inelastic incoherent neutron-scattering experiments are simulated for simple models: a rigid solid (as used for normalization), a glass (with a smooth distribution of harmonic vibrations), and a viscous liquid (described by schematic mode-coupling equations). In cases where the input scattering law factorizes into a wave-number-dependent amplitude and a frequency-dependent spectral distribution, the latter is only weakly affected by multiple scattering, whereas the former is severely distorted. PMID- 11101991 TI - Planform selection in two-layer Benard-marangoni convection AB - Benard-Marangoni convection in a system of two superimposed liquids is investigated theoretically. Extending previous studies, the complete hydrodynamics of both layers is treated and buoyancy is consistently taken into account. The planform selection problem between rolls, squares, and hexagons is investigated by explicitly calculating the coefficients of an appropriate amplitude equation from the parameters of the fluids. The results are compared with recent experiments on two-layer systems in which squares at onset have been reported. PMID- 11101992 TI - Toy model for the mean-field theory of hard-sphere liquids AB - We investigate a toy model of liquid, based on simplified hypernetted chain equations in very large spatial dimension D. The model does not exhibit a phase transition, but several regimes of behavior when D-->infinity can be observed in different intervals of the density. PMID- 11101993 TI - Hydrodynamic fluctuations in the kolmogorov flow: nonlinear regime AB - In a previous paper [I. Bena, M. Malek Mansour, and F. Baras, Phys. Rev. E 59, 5503 (1999)] the statistical properties of linearized Kolmogorov flow were studied, using the formalism of fluctuating hydrodynamics. In this paper the nonlinear regime is considered, with emphasis on the statistical properties of the flow near the first instability. The normal form amplitude equation is derived for the case of an incompressible fluid and the velocity field is constructed explicitly above (but close to) the instability. The relative simplicity of this flow allows one to analyze the compressible case as well. Using a perturbative technique, it is shown that close to the instability threshold the stochastic dynamics of the system is governed by two coupled nonlinear Langevin equations in Fourier space. The solution of these equations can be cast into the exponential of a Landau-Ginzburg functional, which proves to be identical to the one obtained for the case of an incompressible fluid. The theoretical predictions are confirmed by numerical simulations of the nonlinear fluctuating hydrodynamic equations. PMID- 11101994 TI - Density functional theory of long-range critical wetting AB - The wetting properties of a fluid adsorbed at a solid substrate are studied by means of density functional theory. Explicit calculations of the substrate-liquid and substrate-gas density profiles are carried out and used to evaluate the asymptotic expansion for the interface potential of a system with long-range interactions. The range of validity of the asymptotic expansion is checked by comparing it with the interface potential obtained numerically through the constrained minimization of the density functional free energy. Depending on the parameters of the fluid-fluid and substrate-fluid interactions we find first order or critical wetting transitions. In a limited range of parameters, the critical wetting transition is preceded by a first-order transition between a microscopic and a mesoscopic film, thus corroborating previous calculations and experiments for alkanes on brine. We find that the behavior of the alkanes on brine is not universal, since it requires fine-tuning of the fluid-fluid and substrate-fluid interactions. Finally, we investigate the influence of the short- and long-range forces on the location of the first-order transition. We find that for the models studied, the long-range forces cannot be treated perturbatively. Thus for this type of model it is not possible to separate the effects of short- and long-range forces as done in Landau theories, where the long-range forces are treated perturbatively. PMID- 11101995 TI - Density fluctuations and entropy AB - A functional for the entropy that is asymptotically correct both in the high- and low-density limits is proposed. The new form is S=S((id))+S((ln))+S((r))+S((c)), where the term S((c)) depends on the p-body density fluctuations alpha(p) and has the form S((c))/k=ln 2-1+ summation operator(infinity)(p=2) (ln 2)(p)/p! alpha(p)-[exp(alpha(2)-1)-alpha(2)]+Sinsertion mark. Sinsertion mark renormalizes the ring approximation S((r)). This result is obtained by analyzing the functional dependence of the most general expression of the entropy. Two main results for S((c)) are proved: (i) In the thermodynamic limit it is only a functional of the one-body distribution function and (ii) by summing to infinite order the leading contributions in the density a numerical expression for the entropy [Eq. (33)] with a renormalized ring approximation is obtained. The relation of these results to the incompressible approximation for the entropy is discussed and preliminary numerical results on hard spheres are presented. PMID- 11101996 TI - Optical emissions in a sonoluminescing bubble AB - We study how the mechanism of spontaneous decay of atoms (or molecules) in a sonoluminescing bubble (SLB) can be affected by the high density and high temperature environment resulting from the rapid collapse of the gas bubble immediately prior to light emission. We present a detailed study of the density of states of photons in multiple-layered spheres, which mimic various stages of a SLB. In particular, we found that the spontaneous decay rate could be strongly enhanced in the presence of a thin plasma shell inside the bubble, which was predicted recently in numerical hydrodynamic simulations of a SLB. PMID- 11101997 TI - Nonlinear viscosity and velocity distribution function in a simple longitudinal flow AB - A compressible flow characterized by a velocity field u(x)(x, t)=ax/(1+at) is analyzed by means of the Boltzmann equation and the Bhatnagar-Gross-Krook kinetic model. The sign of the control parameter (the longitudinal deformation rate a) distinguishes between an expansion (a>0) and a condensation (a<0) phenomenon. The temperature is a decreasing function of time in the former case, while it is an increasing function in the latter. The non-Newtonian behavior of the gas is described by a dimensionless nonlinear viscosity eta(*)(a(*)), that depends on the dimensionless longitudinal rate a(*). The Chapman-Enskog expansion of eta(*) in powers of a(*) is seen to be only asymptotic (except in the case of Maxwell molecules). The velocity distribution function is also studied. At any value of a(*), it exhibits an algebraic high-velocity tail that is responsible for the divergence of velocity moments. For sufficiently negative a(*), moments of degree 4 and higher may diverge, while for positive a(*) the divergence occurs in moments of degree equal to or larger than 8. PMID- 11101998 TI - Molecular-dynamics simulation of two-dimensional thermophoresis AB - A numerical technique is presented for the thermal force exerted on a solid particle by a gaseous medium between two flat plates at different temperatures, in the free molecular or transition flow. This is a two-dimensional molecular dynamics simulation of hard disks in a inhomogeneous thermal environment. All steady-state features exhibited by the compressible hard-disk gas are shown to be consistent with the expected behaviors. Moreover the thermal force experienced by a large solid disk is investigated, and compared to the analytical case of cylinders moving perpendicularly to the constant temperature gradient for an infinite Knudsen number and in an infinite medium. We show precise examples of how this technique can be used simply to investigate more difficult practical problems, in particluar the influence of nonlinear gradients for large applied differences of temperature, of proximity of the walls, and of smaller Knudsen numbers. PMID- 11101999 TI - Correlation of stress and structure in a simple fluid confined to a pore with furrowed walls AB - A Lennard-Jones (12,6) film confined between two furrowed walls was simulated by the grand canonical ensemble Monte Carlo method. The walls are constructed by gouging triangular grooves in planar substrates that are structureless on the molecular scale. The furrows are infinitely long in one transverse direction (y) and of nanoscopic width (s(x)) and depth (D). The furrows in the two walls are maintained parallel and in register. The diagonal components of the stress tensor (T(alphaalpha), alpha=x,y,z) are computed as functions of D and the separation between the substrates (s(z)) at fixed temperature, chemical potential, and s(x). The T(alphaalpha) for the film between the furrowed walls are strongly shifted from their counterparts for the film between flat (i.e., planar) walls. The shifts are rationalized in terms of the structure of the film, which becomes more ordered as the furrows deepen and the packing of film molecules becomes more restricted in the two dimensions normal to the y direction. The results demonstrate the profound impact of the coupling between molecular and nanoscopic scales on the properties of geometrically constrained fluids. PMID- 11102000 TI - Effect of natural convection in a horizontally oriented cylinder on NMR imaging of the distribution of diffusivity AB - This paper describes the influence of natural convection on NMR measurement of a self-diffusion constant of fluid in the earth's magnetic field. To get an estimation of the effect, the Lorenz model of natural convection in a horizontally oriented cylinder, heated from below, is derived. Since the Lorenz model of natural convection is derived for the free boundary condition, its validity is of a limited value for the natural no-slip boundary condition. We point out that even a slight temperature gradient can cause significant misinterpretation of measurements. The chaotic nature of convection enhances the apparent self-diffusion constant of the liquid. PMID- 11102001 TI - Anomalous burning rates of flamelets induced by self-similar multiple scale (fractal and spiral) initial fields AB - In contrast to the classical problem of a single idealized flamelet (which is described by a nonlinear reaction-diffusion equation of motion) which propagates at a constant burning rate, self-similar multiple scale fields, whether fractal or nonfractal, induce anomalous rates of burning determined by the space-filling properties of the initial field. We compare the regimes induced by (line-cuts through) three specific geometries with distinct space-filling characteristics: (1) an algebraic spiral which has capacity (box-counting dimension) D(k)>0, and fractal dimension H=0; (2) an exponential spiral which has D(k)=0 and H=0, and geometric ratio R>1; (3) a fractal Cantor dust which has D(k)=H>0. The (nondimensional) burning rate U(B) induced by all three geometries takes the general form U(B) approximately F(tau(-zeta)), where F is a function whose form depends on the specific geometry, zeta is an exponent that contains the space filling characteristic of the geometry, and tau is a nondimensional time. (1) For the algebraic spiral, F(x)=1(x), and zeta=D(k); F is continuous. (2) For the exponential spiral, F(x)=ln(x), and zeta=1/(R-1); F is continuous. (3) For the fractal Cantor dust, F(u)(x)=1(x), and zeta=H (for the envelope); F itself is a step-like discontinuous function. Thus, as D(k)-->0, or as H-->0, or as R- >infinity, then zeta-->0 and U(B)-->const; and as D(k)-->1, or as H-->1, (space filling) then zeta-->1; and as R-->1 (space filling) then zeta-->infinity. Two numerical methods, a fundamental (Eulerian) solution to the equation of motion and a Lagrangian model for flamelet propagation, confirm these theoretical predictions. The Lagrangian model is based on the idealized flamelet as a "point" with finite flame thickness Delta(L), (which is determined by the two-flamelet collision process), propagating with a given flame speed U(L). The Lagrangian model allows simulations in parameter ranges not easily accessible by the fundamental method (such as the case for the fractal Cantor dust). Interestingly, the linear regime of scalar diffusion in an algebraic spiral field displays the same dependence on D(k) as in the present reaction-diffusion case. The nonlinear regime of advection-diffusion (Burger turbulence) shows a different dependence on D(k). PMID- 11102002 TI - Shock wave profiles in the burnett approximation AB - This paper is devoted to a discussion of the profiles of shock waves using the full nonlinear Burnett equations of hydrodynamics as they appear from the Chapman Enskog solution to the Boltzmann equation. The system considered is a dilute gas composed of rigid spheres. The numerical analysis is carried out by transforming the hydrodynamic equations into a set of four first-order equations in four dimensions. We compare the numerical solutions of the Burnett equations, obtained using Adam's method, with the well known direct simulation Monte Carlo method for different Mach numbers. An exhaustive mathematical analysis of the results offered here has been done mainly in connection with the existence of heteroclinic trajectories between the two stationary points located upflow and downflow. The main result of this study is that such a trajectory exists for the Burnett equations for Mach numbers greater than 1. Our numerical calculations suggest that heteroclinic trajectories exist up to a critical Mach number ( approximately 2.69) where local mathematical analysis and numerical computations reveal a saddle-node-Hopf bifurcation. This upper limit for the existence of heteroclinic trajectories deserves further clarification. PMID- 11102003 TI - Fluctuations of elastic interfaces in fluids: theory, lattice-boltzmann model, and simulation AB - We study the dynamics of elastic interfaces-membranes-immersed in thermally excited fluids. The work contains three components: the development of a numerical method, a purely theoretical approach, and numerical simulation. In developing a numerical method, we first discuss the dynamical coupling between the interface and the surrounding fluids. An argument is then presented that generalizes the single-relaxation-time lattice-Boltzmann method for the simulation of hydrodynamic interfaces to include the elastic properties of the boundary. The implementation of this method is outlined and it is tested by simulating the static behavior of spherical bubbles and the dynamics of bending waves. By means of the fluctuation-dissipation theorem we recover analytically the equilibrium frequency power spectrum of thermally fluctuating membranes and the correlation function of the excitations. Also, the nonequilibrium scaling properties of the membrane roughening are deduced, leading us to formulate a scaling law describing the interface growth, W2(L,t)=L(3) g(t/L(5/2)), where W, L, and t are the width of the interface, the linear size of the system, and the time, respectively, and g is a scaling function. Finally, the phenomenology of thermally fluctuating membranes is simulated and the frequency power spectrum is recovered, confirming the decay of the correlation function of the fluctuations. As a further numerical study of fluctuating elastic interfaces, the nonequilibrium regime is reproduced by initializing the system as an interface immersed in thermally preexcited fluids. PMID- 11102004 TI - Field theory of absorbing phase transitions with a nondiffusive conserved field AB - We investigate the critical behavior of a reaction-diffusion system exhibiting a continuous absorbing-state phase transition. The reaction-diffusion system strictly conserves the total density of particles, represented as a nondiffusive conserved field, and allows an infinite number of absorbing configurations. Numerical results show that it belongs to a wide universality class that also includes stochastic sandpile models. We derive microscopically the field theory representing this universality class. PMID- 11102005 TI - Scaling behavior in a nonlocal and nonlinear diffusion equation AB - We present the results of analytical studies of a one-dimensional nonlocal and nonlinear diffusion equation describing nonequilibrium processes ranging from aggregation phenomena to the cooperation of individuals. On tuning the initial conditions, a dynamical transition with a universal scaling behavior is observed between two different asymptotic (in time) solutions. The scaling behavior at the transition is also obtained in a self-organized manner, independent of the initial conditions, on temporally evolving the diffusion equation subjected to a mirror symmetry transformation. PMID- 11102006 TI - Nonregular languages in the kicked rotor AB - A symbolic dynamics of the Markov chain model for trajectories of the kicked rotor (standard map) in sticky regions near resonant orbits is developed. Two sets of trajectories are shown to correspond to a context-free and a context sensitive language, respectively. PMID- 11102007 TI - Interface dynamics in hele-shaw flows with centrifugal forces: preventing cusp singularities with rotation AB - A class of exact solutions of Hele-Shaw flows without surface tension in a rotating cell is reported. We show that the interplay between injection and rotation modifies the scenario of formation of finite-time cusp singularities. For a subclass of solutions, we show that, for any given initial condition, there exists a critical rotation rate above which cusp formation is suppressed. We also find an exact sufficient condition to avoid cusps simultaneously for all initial conditions within the above subclass. PMID- 11102008 TI - Manifestation of anisotropy persistence in the hierarchies of magnetohydrodynamical scaling exponents AB - An example of a turbulent system where the failure of the hypothesis of small scale isotropy restoration is detectable both in the "flattening" of the inertial range scaling exponent hierarchy and in the behavior of odd-order dimensionless ratios, e.g., skewness and hyperskewness, is presented. Specifically, within the kinematic approximation in magnetohydrodynamical turbulence, we show that for compressible flows, the isotropic contribution to the scaling of magnetic correlation functions and the first anisotropic ones may become practically indistinguishable. Moreover, the skewness factor now diverges as the Peclet number goes to infinity, a further indication of small-scale anisotropy. PMID- 11102009 TI - Air-water interface-induced smectic bilayer AB - We show, using surface pressure versus molecular area isotherms measurements and x-ray reflectivity, that the long diblock semifluorinated n-hexaeicosane molecules, F(CF2)(8)-(CH2)18H, form a stable smectic bilayer phase, noted M1, with a total thickness of 3. 3 nm, at an apparent molecular area about 0.3 nm(2), though in the bulk the used molecules do not form smectic phases at any temperature. We discuss different molecular packing models according to our experimental data and deduce that molecules are antiparallel with fluorinated chains outwards and interleaved hydrocarbon chains inwards. PMID- 11102010 TI - Formation of string defects at thinning transitions in smectic-C* free-standing films AB - The layer thinning transitions in freely suspended smectic-C* films have been investigated. The defect structure formed by stringlike lines was observed just before the thinning transitions. The string defects disappear after the thinning transition and appear again near the temperature of the next thinning transition. These results clearly indicate that thin free-standing films at the thinning transitions are slightly below the melting temperature of the interior layers. PMID- 11102011 TI - Observation of the soft mode in the Sm-C(*)(alpha) phase AB - We have performed the electro-optical measurements in an antiferroelectric liquid crystal 4-(1-methylheptyloxycarbonyl)phenyl 4(')-octylcarbonyloxybiphenyl-4 carboxylate (MHPOCBC) and observed the soft mode in the chiral Sm-C(*)(alpha) phase. This is a direct consequence that the Sm-C(*)(alpha) phase is induced by the condensation of a tilting mode. A Landau-type theory was developed to analyze frequency dispersions in the second-order response and it was found that the Sm-A Sm-C(*)(alpha) phase transition is close to a tricritical point. PMID- 11102012 TI - Nonlinear dielectric relaxation spectroscopy of ferroelectric liquid crystals AB - The nonlinear dielectric relaxation spectra of ferroelectric liquid crystals (FLCs) have been studied in the chiral smectic-C phase. The linear and third order nonlinear dielectric spectra show the relaxation corresponding to the fluctuation in the azimuthal angle of directors called the Goldstone mode. We calculated the nonlinear dielectric spectra of the Goldstone mode theoretically by the torque balance equation which describes the dynamics of FLCs under the electric field. The calculated spectra make good agreement with the measured ones. We also evaluated the material constants of FLCs from the best-fitted values of the linear and nonlinear dielectric increment and relaxation time. PMID- 11102013 TI - Electric field-induced acoustic-optic mode coupling in an anticlinic liquid crystal AB - A dc electric field was applied perpendicular to the tilt plane of a pitch compensated (unwound helix) anticlinic liquid crystal. By means of quasielastic light scattering, the field was found to couple the acoustic and optic Goldstone modes, resulting in an increase of the relaxation time tau(beta) of the acousticlike eigenmode. Elastic constants were estimated from the relaxation time data. PMID- 11102014 TI - Slow dynamics of equilibrium density fluctuations in suspensions of colloidal hard spheres near the glass transition AB - A mean-field theory for the dynamics of equilibrium suspensions of colloidal hard spheres near the glass transition is presented based on the standpoint recently proposed by the present author. It is shown that although the relative magnitude of the density fluctuations to the mean equilibrium density is small even near the glass transition, they are described by a nonlinear stochastic equation which originates from the long-range hydrodynamic interactions between particles. A nonlinear mean-field equation for the particle mean-square displacement is then derived. This equation is used to analyze the recent experimental data for equilibrium colloidal suspensions. Analyses show that no divergence of the alpha- and beta-relaxation times is found in the experimental data, although the dynamic properties of the colloidal liquid exhibit a drastic slowing down in the so called supercooled region. PMID- 11102015 TI - Convergent calculation of the asymptotic dimension of diffusion limited aggregates: scaling and renormalization of small clusters AB - Diffusion limited aggregation (DLA) is a model of fractal growth that had attained a paradigmatic status due to its simplicity and its underlying role for a variety of pattern forming processes. We present a convergent calculation of the fractal dimension D of DLA based on a renormalization scheme for the first Laurent coefficient of the conformal map from the unit circle to the expanding boundary of the fractal cluster. The theory is applicable from very small (2-3 particles) to asymptotically large (n-->infinity) clusters. The computed dimension is D=1.713+/-0.003. PMID- 11102016 TI - Approach to Monte Carlo calculation of the buckling of supercoiled DNA loops. AB - The short supercoiled circular DNA molecules are shown to be glassy systems and canonical Metropolis Monte Carlo simulations of the systems tend to get stuck in local metastable energy basins. A Monte Carlo algorithm is developed to alleviate the problem of "ergodicity breaking" of the glassy systems, in which the Markov process is driven by an explicitly analytic weight factor with enhanced probability in both low- and high-energy regions. To characterize the degree of puckering of the supercoiled DNA loops, a different quantity of aplanarity is introduced as the shortest principal axis of configurational ellipsoid of DNA. With the suggested Monte Carlo method, the quantitative correlation between supercoiling degree and buckling of DNA is attained. With supercoiling stress increasing, the conformational transition from a circle to mono-, diplo-, or triple interwound superhelical structure will take place in a successive but decreasingly abrupt mode. PMID- 11102017 TI - Experimental evidence of electric inhibition in fast electron penetration and of electric-field-limited fast electron transport in dense matter AB - Fast electron generation and propagation were studied in the interaction of a green laser with solids. The experiment, carried out with the LULI TW laser (350 fs, 15 J), used K(alpha) emission from buried fluorescent layers to measure electron transport. Results for conductors (Al) and insulators (plastic) are compared with simulations: in plastic, inhibition in the propagation of fast electrons is observed, due to electric fields which become the dominant factor in electron transport. PMID- 11102018 TI - X-ray and extreme ultraviolet emission induced by variable pulse-width irradiation of Ar and Kr clusters and droplets AB - Measurements are presented of x-ray (>1.5 keV) and extreme ultraviolet (EUV, lambda=2-44 nm) emission from argon and krypton supersonic gas jets at room (T=300 K) and cryogenic (T=173 K) temperatures irradiated with constant energy (50 mJ), variable width laser pulses ranging from 100 fs to 10 ns. Two regimes of jet operation are explored: cluster formation (radius<100 nm) and droplet formation (radius>1 &mgr;m). The results for both clusters and droplets can be understood in terms of two time scales: a short time scale for optimal resonant absorption at the critical density layer in the expanding plasma, and a longer time scale for the plasma to drop below critical density. PMID- 11102019 TI - Time-resolved experiments on light diffusion in anisotropic random media AB - Multiple light scattering in isotropic and anisotropic media is studied experimentally with an optical gating technique, as commonly used in fluorescence spectroscopy. The experimental setup permits an accurate analysis of the propagation of a short light pulse through disordered or partially ordered media. The diffusion constant of some isotropic systems is reported, and the anisotropy in the diffusion constant for light diffusion through liquid crystals is observed. For the time-resolved data, good agreement with diffusion theory is found in all cases, including the liquid crystal in the nematic phase. PMID- 11102020 TI - Liquid crystal director fluctuations and surface anchoring by molecular simulation AB - We propose a simple and reliable method to measure the liquid crystal surface anchoring strength by molecular simulation. The method is based on the measurement of the long-range fluctuation modes of the director in confined geometry. As an example, molecular simulations of a liquid crystal in slab geometry between parallel walls with homeotropic anchoring have been carried out using the Monte Carlo technique. By studying different slab thicknesses, we are able to calculate separately the position of the elastic boundary condition and the extrapolation length. PMID- 11102021 TI - Fine structure of defects in radial nematic droplets AB - We investigate the structure of defects in nematic liquid crystals confined in spherical droplets and subject to radial strong anchoring. Equilibrium configurations of the order-parameter tensor field in a Landau-de Gennes free energy are numerically modeled using a finite-element package. Within the class of axially symmetric fields, we find three distinct solutions: the familiar radial hedgehog, the small ring (or loop) disclination predicted by Penzenstadler and Trebin, and a solution that consists of a short disclination line segment along the rotational symmetry axis terminating in isotropic end points. Phase and bifurcation diagrams are constructed to illustrate how the three competing configurations are related. They confirm that the transition from the hedgehog to the ring structure is first order. The third configuration is metastable (in our symmetry class) and forms an alternate solution branch bifurcating off the radial hedgehog branch at the temperature below which the hedgehog ceases to be metastable. Dependence on temperature, droplet size, and elastic constants is investigated, and comparisons with other studies are made. PMID- 11102022 TI - Isotropic-nematic interface of soft spherocylinders AB - The isotropic-nematic interface of a simple model of liquid-crystal molecules has been investigated using computer simulation, and by numerical minimization of the Onsager free-energy functional. The molecules are represented by long spherocylindrical particles interacting via the Kihara potential. The agreement between simulation and theory is excellent, apart from the bulk coexistence densities which are over estimated by the theory. Planar alignment of the molecules at the interface is preferred in all cases. The number density profile is found to vary monotonically, both in simulation and in theory. Biaxiality of the molecular orientational distribution near the interface is demonstrated. PMID- 11102023 TI - Thermal renormalization of the anchoring energy of nematic liquid crystals AB - The temperature dependence of the anchoring energy of a nematic liquid crystal on thermal fluctuations is studied. We consider the weak anchoring case, where the interaction of a nematic molecule on the surface with the substrate is small with respect to the mean field energy due to the other nematic molecules. The analysis is performed by means of a perturbation method in which the expansion parameter is the surface interaction. The presented model is valid for any value of the scalar order parameter. We show that the renormalization of the anchoring coefficients due to thermal fluctuations is proportional to the generalized scalar order parameters. We show also that, at the lowest order in the scalar order parameter, Landau-like theories agree with our mean field approach. An expression for the thermal renormalization of the anchoring coefficients valid in the low temperature region, where the fluctuations are small, is derived. The agreement between our theoretical predictions and the experimental data obtained by other groups is fairly good over a large temperature range. PMID- 11102024 TI - Stimulated orientational and thermal scatterings and self-starting optical phase conjugation with nematic liquid crystals AB - A quantitative theory and experimental results on self-starting optical phase conjugation, using stimulated orientational and thermal scattering in nematic liquid crystal films, are presented. The coupled wave-material equations for the laser-induced refractive index changes, grating formation, and coherent wave mixing effects are developed. Analytical solutions are obtained for the case of negligible pump depletion, and numerical solutions for various input and generated signals, taking losses into account, are obtained. Experimentally, we demonstrate the feasibility of realizing these stimulated scattering and phase conjugation processes in thin (200 &mgr;m) nematic liquid crystal with a milliwatt-power cw laser. Theoretical estimates for various gain constants and threshold intensities, and their dependence on various physical parameters, are found to be in good agreement with experimental observations. PMID- 11102025 TI - Surface extrapolation length and director structures in confined nematics AB - We report the results of Monte Carlo simulations of the Lebwohl-Lasher model of nematic liquid crystals confined to cylindrical cavities with homeotropic anchoring. We show that the ratio of the bulk to surface couplings is not in general equal to the corresponding parameter K/W used in elastic theory (where K is the Frank elastic constant in the one-constant approximation and W is the surface anchoring strength). By measuring the temperature dependence of K/W (which is equivalent to the surface extrapolation length) we are able to reconcile the results of our simulations as well as others with the predictions of elastic theory. We find that the rate at which we cool the system from the isotropic to nematic phase plays a crucial role in the development of the final director structure, because of a large free energy barrier separating different director structures as well as the temperature dependence of K/W. With a suitably fast cooling rate we are able to keep the system out of a metastable planar state and form an escaped radial structure for large enough systems. Copyright 2000 The American Physical Society. PMID- 11102026 TI - Tiling the plane with noncongruent toric focal conic domains AB - This paper deals with regular arrays of macroscopic defects (focal conic domains) observed when a slab of lamellar phase is sandwiched between two substrates imposing different orientational anchorings. We report, in particular, detailed observations of the texture of a lyotropic lamellar phase in contact with a glass substrate and a lyotropic sponge phase. We consider several models for the defects depending on the material and the substrate parameters. Their energy has a common form, and the main features of the textures are explained in the framework of a simple model where disks of different sizes tile a plane in order to minimize a particular interface energy. PMID- 11102027 TI - Evolution of interfaces and expansion in width AB - Interfaces in a model with a single, real nonconserved order parameter and purely dissipative evolution equation are considered. We show that a systematic perturbative approach, called the expansion in width and developed for curved domain walls, can be generalized to the interfaces. A procedure for calculating curvature corrections is described. We also derive formulas for local velocity and local surface tension of the interface. As an example, evolution of spherical interfaces is discussed, including an estimate of the critical size of small droplets. PMID- 11102028 TI - Structural study of the smectic-I to smectic-F transition in freely suspended films AB - The smectic-I (S(I)) to smectic-F (S(F)) phase transition in terephthal-bis-(4n) decylaniline (TB10A) has been examined for a possible continuous transition via the intermediate smectic-L (S(L)) phase. X-ray diffraction measurements of thick, single-domain, freely suspended films are used to classify the phases and to determine the hexatic order parameter C6 and its harmonics. Instead of the continuous transition suggested in the literature we find a first-order S(I)- >S(F) transition with a discontinuous change in the direction of the bond orientational order relative to the molecular tilt. The tilt of the molecular form factor in the hexatic phases is inconsistent with the tilt estimated from published layer spacing measurements, suggesting that the hexatic phases of TB10A must have the molecular cores oriented at an angle relative to the tails. This result taken together with published results on the S(C) phase suggests that the S(C)-->S(I)-->S(F) transitions are driven by changes in the conformation of the hydrocarbon tails. Examination of the harmonic scaling relation between the hexatic order parameters shows mean-field behavior in the S(I) phase. This result will make binary mixtures of TB10A with other materials a practical system for the study of the crossover from mean field to the XY behavior seen in other hexatic systems. PMID- 11102029 TI - Orientational ordering in fluids with partially constrained molecule orientations AB - Molecular orientations in anisotropic fluids can be partially constrained as a result of electric or magnetic fields or interface influences. A statistical approach for the investigation of the orientational ordering in such systems is proposed. The long-range correlations are taken into account consistently. The method is illustrated for the well-known thermotropic nematic model in an infinite disorienting field W, when the molecules are constrained to orient perpendicularly to the field direction. For this problem the analytical solution of the anisotropic Ornstein-Zernike equation is obtained, and the asymptotic expression for the long-range correlations on large distances is given. The phase diagram and elastic constants are calculated for W-->infinity and are compared with the usual case of a uniaxial nematic ordering at W=0. In the case W- >infinity when the temperature decreases the orientational phase transition of the second order becomes the one of the first order at a tricritical point. The disorienting field W increases much the region of an ordered fluid. It is shown that at a given pressure the orientational ordering temperature for W-->infinity is higher about 1.2-1.5 times than the one at W=0. The orientational ordering pressure is less about 4-5 times than the pressure of the uniaxial nematic ordering (W=0) at the same temperature. The disorienting field increases elastic properties of the model under consideration. PMID- 11102030 TI - Direct observation of a twist mode in electroconvection AB - I report on the direct observation of a uniform twist mode of the director field in electroconvection in the nematic liquid crystal I52. Recent theoretical work suggests that such a uniform twist mode of the director field is responsible for a number of secondary bifurcations in both electroconvection and thermal convection in nematics. I show here evidence that the proposed mechanisms are consistent with being the source of the previously reported stationary oblique roll pattern identified as the SO2 state of electroconvection in the liquid crystal I52. The same mechanisms also contribute to a tertiary Hopf bifurcation that I observe in electroconvection in the liquid crystal I52. There are quantitative differences between the experiment and calculations that only include the twist mode. These differences suggest that a complete description must include effects described by the weak-electrolyte model of electroconvection. PMID- 11102031 TI - Theoretical investigation into the effects of polar anchoring in antiferroelectric liquid crystal cells AB - We present a theoretical investigation into the effects of polar anchoring, which induces ferroelectric ordering close to the cell surfaces, in a liquid crystal cell containing an antiferroelectric liquid crystalline material. Our model includes effects due to finite polar and nonpolar anchoring, quadrupolar ordering and polarization self interaction. By minimizing the free energy of the system, we find parameter domains in which multiple zero-voltage solutions are stable. We find that these solutions may undergo thresholdless or hysteretic switching depending on the parameter values. In two instances, the presence of quadrupolar ordering or weak anchoring means that the cell must first be primed into the thresholdless state through a discontinuous transition from an initial antiferroelectric state. PMID- 11102032 TI - Determination of the interaction force between two adsorptive surfaces delimiting a critical binary polymer blend AB - We consider a mixture of two incompatible polymers A and B, confined between two parallel surfaces of the same chemical nature, separated by a distance L. It is assumed that both surfaces strongly adsorb one of the species (A) at high temperature. It is also assumed that a demixing transition occurs at a critical temperature T(c) below the adsorption temperature T(a). The strong adsorption implies that the composition of species A on surfaces is quenched even when the temperature is lowered. The presence of strong density fluctuations near the critical point induces an interaction between the surfaces. We reexamine this attractive force and determine its dependence with the thickness L, when the latter is smaller than the thermal correlation length. We find that, in the vicinity of the critical point, this force decreases with distance as L-4. We show that the corresponding amplitude is a universal number, independent of the value of the composition on surfaces, and we give its exact expression. Finally, we note that the present system may be considered as a typical model enabling one to understand qualitatively and quantitatively the flocculation of colloids embedded in critical binary polymer blends. PMID- 11102033 TI - Effects of electric charges on hydrophobic forces. II AB - We study by molecular-dynamics simulations the effect of electric charges of either sign on hydrophobic interactions and on the dynamics of hydration water, using explicit water and very simplified solutes. Results show that the presence of a charged solute can disrupt the "hydrophobic contact bond" between two apolar solutes nearby, by forcing them towards a different configuration. As a consequence of different structural changes of the solvent caused by charges of opposite sign, the effect is markedly charge-sign-dependent. Analogous weaker effects appear to be induced by the presence of one additional apolar element. The dynamics of hydration water around each solute is also seen to be strongly influenced by the presence of other (charged or uncharged) nearby solutes. Comparison between our results on hydration water dynamics around charged solutes and available experimental data allows sorting out the effects of solute charge sign and size. Our results also offer a plain interpretation of the equivalence of the effects on water structure due to solute ions and to high pressures. These results reflect at a basic paradigmatic level the immensely more complex cases of well-known phenomena such as salting-in and salting-out, and of protein conformational changes caused, e.g., by the arrival of a charged or of an apolar group (phosphorilation or methylation). As it will be discussed, they help in the direction of Delbruck's desirable "progress towards a radical physical explanation" for this class of phenomena. PMID- 11102034 TI - Shear response of a frictional interface to a normal load modulation AB - We study the shear response of a sliding multicontact interface submitted to a harmonically modulated normal load, without loss of contact. We measure, at low velocities (V<100 &mgr;m s(-1)), the average value &Fmacr; of the friction force and the amplitude of its first and second harmonic components. The excitation frequency (f=120 Hz) is chosen much larger than the natural one, associated with the dynamical aging of the interface. We show the following: (i) In agreement with the engineering thumb rule, even a modest modulation induces a substantial decrease of &Fmacr;. (ii) The Rice-Ruina state and rate model, though appropriate to describe the slow frictional dynamics, must be extended when dealing with our "high" frequency regime. That is, the rheology which controls the shear strength must explicitly account not only for the plastic response of the adhesive junctions between load-bearing asperities, but also for the elastic contribution of the asperities bodies. This "elastoplastic" friction model leads to predictions in excellent quantitative agreement with all our experimental data. PMID- 11102035 TI - Influence of the free-energy functional form on simulated morphology of spinodally decomposing blends AB - The spinodal decomposition of a binary mixture has been studied within several mesoscopic models. It has been found that the form of the equilibrium free energy has a crucial effect on the morphological development in asymmetric blends. We have shown that the principal quantity that determines the topology of the interface (and type of morphology) is the equilibrium minority phase volume fraction, while the transition from bicontinuous to droplet morphology can be treated as a percolation. The concentration dependence of the square gradient coefficient attributed for the Flory-Huggins-de Gennes free energy has no significant influences on the average domain growth, but can be distinguished experimentally from its constant-coefficient alternative by measuring the maximum wave vector of the scattering intensity as a function of the minority phase volume fraction for spinodally decomposing asymmetric blends. The concentration dependence of the Onsager coefficient has the weak, systematic effect of slowing down the morphological development. The local shape of the interface is not affected considerably by the concentration dependence of the square gradient and Onsager coefficient. PMID- 11102036 TI - Spatially patterned static roughness superimposed on thermal roughness in a condensed phospholipid monolayer AB - Imaging of diffuse light scattering in reflection from a phospholipid monolayer at the air/water interface has revealed a previously undetected separation of the monolayer into two regions distinguishable by the intensity of their scattering. In monolayers of L-dipalmitoyl phosphatidylcholine in the condensed phase, chiral shaped domains are surrounded by a brighter region that covers approximately half the monolayer. While the scattered intensity from both regions increases with surface pressure in a manner consistent with scattering from thermally induced capillary waves, the additional scattering from the brighter region indicates a static surface roughness superimposed on the thermal roughness. PMID- 11102037 TI - Ordering process in quenched block copolymers at low temperatures AB - We have studied domain growth of symmetric diblock copolymers undergoing microphase separation at low temperatures. We introduce a phenomenological nonlinear diffusion model with order-parameter-dependent mobility. Performing two dimensional simulations, we find that the time-dependent scattering function exhibits dynamical scaling with a logarithmic growth law in the strong segregation limit where surface diffusion is the relevant mechanism for coarsening. PMID- 11102038 TI - Dynamic electrorheological effects and interparticle force between a pair of rotating spheres AB - We consider a two-particle system in which a particle is held fixed, and the other one rotates around the axis perpendicular to the line joining the particles' centers. The rotating particle leads to a displacement of its polarization charge on the surface. Our results show that the rotational motion of the particles generally reduces the force between the particles. The dependence of interparticle force on the angular velocity of rotation will be discussed. PMID- 11102039 TI - Avalanches in fine, cohesive powders AB - We have investigated the onset of avalanches in fine, cohesive granular materials. In our experiments shear stress is generated by tilting an initialized bed of powder and increasing the angle of tilt until the powder avalanches. We find that the angle alpha of the avalanche decreases with increasing bed width. The avalanche depth increases with the bed width and, in all cases, is of the order of several millimeters, which is much greater than the particle size. We carry out a macroscopic analysis of the avalanche process based on Coulomb's method of wedges. This analysis shows the fundamental role played by powder cohesion and boundary conditions on avalanches in fine cohesive powders. This behavior contrasts with the behavior of noncohesive grains, such as dry sand, where avalanches consist of superficial layers of about ten grains. The reason behind this is that for our experimental powders (particle diameter approximately 10 &mgr;m) the van der Waals interparticle adhesive force exceeds several orders of magnitude particle weight. Adhesive forces oppose gravity, and as a result fine cohesive powders settle in very open structures as compared to noncohesive granular materials. Because of the dominance of adhesive forces over particle weight, our materials behave more like wet sand. PMID- 11102040 TI - Spreading dynamics of water droplets AB - The spreading dynamics of water droplets on flat silicon surfaces is investigated. It is shown that, for situations close to complete wetting, the radius evolution with time can be described using a power law with a nonstandard exponent of 1/7. This dynamics is interpreted using a hydrodynamic model with an invariant dissipation profile. Such a description is also consistent with the slow dynamics observed for larger contact angles. PMID- 11102041 TI - Fluctuations of lamellar structure prior to a lamellar-->gyroid transition in a nonionic surfactant system AB - Fluctuations of lamellar structure prior to a lamellar-->gyroid transition in a nonionic surfactant-water system has been investigated by means of small-angle x ray scattering (SAXS) and differential scanning calorimeter measurements. For large DeltaT (DeltaT=T-T(LG), where T is the temperature and T(LG) the lamellar- >gyroid transition temperature) in the lamellar phase, the SAXS profiles can be described by a Caille correlation function for undulating lamellar structure. Approaching the temperature to the T(LG), an excess diffuse scattering grows at the lower Q (Q is the magnitude of scattering vector) side of the first lamellar peak. Highly oriented lamellar samples revealed that the excess diffuse scattering arises from in-plane density fluctuations. We attribute this diffuse scattering to the perforation fluctuation layer (PFL) structure and we show that the PFL is an equilibrium structure. At T(LG), the PFL transformed to the gyroid phase through a transient ordered structure having a rhombohedral symmetry. PMID- 11102042 TI - Magnetization of ferrofluids with dipolar interactions: A born-mayer expansion AB - For ferrofluids that are described by a system of hard spheres interacting via dipolar forces we evaluate the magnetization as a function of the internal magnetic field with a Born-Mayer technique and an expansion in the dipolar coupling strength. Two different approximations are presented for the magnetization, considering different contributions to a series expansion in terms of the volume fraction of the particles and the dipolar coupling strength. PMID- 11102043 TI - Interlayer interactions in enantiomeric anticlinic liquid crystalline mixtures AB - The interlayer interaction coefficient U in the anticlinic phase of enantiomeric binary mixtures was determined by measuring the threshold electric field for the onset of solitary waves. U was found to increase with decreasing temperature in the anticlinic phase for a given enantiomer excess X. The ratio U(X,straight theta)/U(X=1,straight theta), where straight theta is the polar angle, was found to be significantly smaller for mixtures with low enantiomer excess than for the optically pure material. The observed behavior is analyzed using a mean-field model for dipole-dipole interactions in adjacent layers. PMID- 11102044 TI - Kinetic pathways of multiphase surfactant systems AB - Relaxation following a temperature quench of two- (L(alpha) and L3) and three phase (L(alpha), L3, and L1) samples has been studied in a sodium dodecyl sulfate octanol-brine system. In the three-phase case we have observed samples that are initially mainly of the sponge phase, with lamellar and micellar phases on the top and bottom of the sample, respectively. Upon decreasing the temperature, most of the volume of the sponge phase is replaced by the lamellar phase. During equilibration we have observed three regimes of behavior within the sponge phase: (i) first there is disruption in the L3 texture, then (ii) after the sponge phase homogenizes there is a L(alpha) nucleation regime; finally (iii), a bizarre plume connects the L(alpha) phase with the L1 phase. The relaxation of the two-phase sample proceeds instead in two stages. First L(alpha) drops nucleate in L3, forming an onion "gel" structure. Over time the L(alpha) structure compacts while equilibrating into a two-phase L(alpha)-L3 sample. We offer possible explanations for some of these observations in the context of a general theory for phase kinetics in systems with one fast composition variable and one slow. PMID- 11102045 TI - Permeation properties of three-dimensional self-affine reconstructions of porous materials AB - A binary medium generation method is presented, which is capable of producing three-dimensional reconstructions of porous solids. The method is based on the midpoint displacement and successive random addition technique, which is, essentially, a graphical reproduction technique for the generation of self-affine media that follow fractional Brownian motion (FBM) statistics. Thresholding of the site values at the desired porosity value leads to three-dimensional porous constructions, the correlation degree of which is defined by the preselected value of the Hurst exponent. The correlation length of such single-cell media is comparable to the size of the working cell and, therefore, this approach would be of local use only. To remedy this problem, a method for producing multicell FBM media is presented, which is capable of generating media with size considerably larger than the correlation length and, as such, they can be employed to simulate a variety of actual porous solids. The percolation properties of these reconstructions are investigated and the specific surface area is calculated as a function of the porosity, the number of interwoven cells, and the Hurst exponent value. Furthermore, the flow equations are solved numerically within the void space of the three-dimensional FBM media and the effects of structure porosity and correlation degree on the permeability are studied. Application of the methodology to a sandstone sample as a case study showed a very good agreement of the numerical predictions for the permeability with actual permeability measurements and with the permeability estimate using a serial sectioning technique. PMID- 11102046 TI - Dynamics and lateral interactions of dipolar chains AB - The dynamics and lateral interactions of dipolar chains in magnetorheological suspensions determine the long-time microscopic structure and resulting rheological response. In this paper we characterize proposed lateral interaction mechanisms and their implications for long-time coarsening of structure and compare them to direct measurements of the lateral interaction of dipolar chains using optical trap micromanipulation. We observe a long-range far-field attraction between flexible chains, while the near-field interaction can be repulsive or attractive. At high field strengths, we observe the short-range attraction of rigid chains. Chain dynamics measured with videomicroscopy and diffusing wave spectroscopy are described by a local-mode model and are consistent with fluctuation-mediated interaction theories. The subdiffusive behavior at intermediate and long times scales as t(0.75), identical to semiflexible molecules. Finally, we show examples of how defects in chains can create lateral attractions or repulsions. PMID- 11102047 TI - Binary hard-sphere fluids near a hard wall AB - By using the Rosenfeld density functional, we determine the number density profiles of both components of binary hard-sphere fluids close to a planar hard wall as well as the corresponding excess coverage and surface tension. The comparison with published simulation data demonstrates that the Rosenfeld functional, both its original version and sophistications thereof, is superior to previous approaches, and exhibits the same excellent accuracy as known from studies of the corresponding one-component system. PMID- 11102048 TI - Simulating formation of voids in charged colloids by brownian dynamics AB - Using Brownian dynamics simulations and Sogami's effective pair potential with a long-range attractive term for the colloidal particles, we report that the mechanism of making voids in charged colloids is the long-range attractive interaction between particles. The present results are in good agreement with the experimental observations of stable voids in equilibrium with ordered structures, which may not be explained by repulsive potentials without a long-range attractive interaction such as the Derjaguin-Landau-Vervey-Overbeck potential. Our results are also in a good agreement with Monte Carlo simulation results. The voids have been reported with ordered structure in charged colloids by Brownian dynamics simulations. PMID- 11102049 TI - Nonequilibrium molecular dynamics simulations of transport and separation of gas mixtures in nanoporous materials AB - The nonequilibrium molecular dynamics simulations of transport and separation of a binary gas mixture through a porous membrane with interconnected pores of distributed sizes are reported. The membrane is modeled by a three-dimensional disordered molecular network of interconnected pores consisting of tens of thousands of atoms, based on a Voronoi tessellation of space. Results are presented for transport and adsorption of the gases, including the existence of an optimal pore structure for maximum separation of the gases. PMID- 11102050 TI - Polydisperse star polymer solutions AB - We analyze the effect of polydispersity in the arm number on the effective interactions, structural correlations, and phase behavior of star polymers in a good solvent. The effective interaction potential between two star polymers with different arm numbers is derived using scaling theory. The resulting expression is tested against monomer-resolved molecular dynamics simulations. We find that the theoretical pair potential is in agreement with the simulation data in a much wider polydispersity range than other proposed potentials. We then use this pair potential as an input in a many-body theory to investigate polydispersity effects on the structural correlations and the phase diagram of dense star polymer solutions. In particular, we find that a polydispersity of 10%, which is typical in experimental samples, does not significantly alter previous findings for the phase diagram of monodisperse solutions. PMID- 11102051 TI - Phase behavior and material properties of hollow nanoparticles AB - Effective pair potentials for hollow nanoparticles such as those made from carbon (fullerenes) or metal dichalcogenides (inorganic fullerenes) consist of a hard core repulsion and a deep, but short-ranged, van der Waals attraction. We investigate them for single-walled and multiwalled nanoparticles and show that in both cases, in the limit of large radii the interaction range scales inversely with the radius R, while the well depth scales linearly with R. We predict the values of the radius R and the wall thickness h at which the gas-liquid coexistence disappears from the phase diagram. We also discuss unusual material properties of the solid, which include a large heat of sublimation and a small surface energy. PMID- 11102052 TI - General view of a liquid-liquid phase transition AB - We present a general view of a liquid-liquid phase transition, based on a simple physical picture that there is "cooperative medium-range bond ordering" for any liquids. Contrary to the common belief, we argue that liquid is not homogeneous and in any liquid there exist locally favored structures, which are frustrated with normal-liquid structures. The cooperative excitation of locally favored structures leads to a gas-liquid-like critical point of bond ordering. This picture naturally leads to the conclusion that liquid-liquid transition is not specific to special materials, but can in principle exist in any liquids. Our model suggests a new possibility that (i) even an ordinary molecular liquid can have a hidden liquid-liquid phase transition and (ii) it may be the origin of a second amorphous phase (e.g., "glacial phase") and critical-like, large-scale fluctuations ("Fischer clusters") observed in supercooled molecular liquids. PMID- 11102053 TI - Hard-core yukawa model for charge-stabilized colloids AB - The hypernetted chain approximation is used to study the phase diagram of a simple hardcore Yukawa model of a charge-stabilized colloids. We calculate the static structure factor, the pair distribution function, and the collective mode energies over a wide range of parameters, and the results are used for studying the freezing transition of the system. The resulting phase diagram is in good agreement with the known estimates and the Monte Carlo simulations. PMID- 11102054 TI - Surface stability of granular systems under horizontal and vertical vibration: the applicability of a coefficient of friction AB - We investigate the conditions under which the surface of a granular pile becomes unstable to vibrations. Three stability boundaries are identified, which depend upon the relative phase of the driving forces and the angle of the prepared slope. The experimental findings can be interpreted within the context of a Coulomb friction model and used to define an effective coefficient of friction. For up-hill motion we find that the coefficient of friction depends strongly on the slope angle and that, in general, it requires less vibration to transport grains uphill than would be otherwise expected. PMID- 11102055 TI - Improvement of the Davydov theory of bioenergy transport in protein molecular systems. AB - The Hamiltonian and the wave function in the Davydov theory have simultaneously been improved and extended, based on some physical and biological grounds and on results from other models. The equations of motion for the improved Davydov model with a quasicoherent two-quanta state and a new interaction term in the Hamiltonian describe bioenergy transport along the molecular chains in protein molecules by a soliton mechanism. Some elementary properties of the soliton, including the nonlinear coupling energy and greatly increased binding energy of the soliton, are also given. The results obtained suggest that the model could be a candidate for a bioenergy transport mechanism in protein molecules. PMID- 11102056 TI - Variational studies and replica symmetry breaking in the generalization problem of the binary perceptron AB - We analyze the average performance of a general class of learning algorithms for the nondeterministic polynomial time complete problem of rule extraction by a binary perceptron. The examples are generated by a rule implemented by a teacher network of similar architecture. A variational approach is used in trying to identify the potential energy that leads to the largest generalization in the thermodynamic limit. We restrict our search to algorithms that always satisfy the binary constraints. A replica symmetric ansatz leads to a learning algorithm which presents a phase transition in violation of an information theoretical bound. Stability analysis shows that this is due to a failure of the replica symmetric ansatz and the first step of replica symmetry breaking (RSB) is studied. The variational method does not determine a unique potential but it allows construction of a class with a unique minimum within each first order valley. Members of this class improve on the performance of Gibbs algorithm but fail to reach the Bayesian limit in the low generalization phase. They even fail to reach the performance of the best binary, an optimal clipping of the barycenter of version space. We find a trade-off between a good low performance and early onset of perfect generalization. Although the RSB may be locally stable we discuss the possibility that it fails to be the correct saddle point globally. PMID- 11102057 TI - Renormalization group analysis of a quivering string model of posture control. AB - Scaling concepts and renormalization group methods are applied to a simple linear model of human posture control consisting of a trembling or quivering string subject to damping and restoring forces. The string is driven by uncorrelated white Gaussian noise, intended to model the corrections of the physiological control system. We find that adding a weak quadratic nonlinearity to the posture control model opens up a rich and complicated phase space (representing the dynamics) with various nontrivial fixed points and basins of attraction. The transition from diffusive to saturated regimes of the linear model is understood as a crossover phenomenon, and the robustness of the linear model with respect to weak nonlinearities is confirmed. Correlations in posture fluctuations are obtained in both time and space domains. There is an attractive fixed point identified with falling. The scaling of the correlations in the front-back displacement, which can be measured in the laboratory, is predicted for both large-separation (along the string) and long-time regimes of posture control. PMID- 11102058 TI - Cellular automata model for citrus variegated chlorosis. AB - A cellular automata model is proposed to analyze the progress of citrus variegated chlorosis epidemics in Sao Paulo orange plantations. In this model epidemiological and environmental features, such as motility of sharpshooter vectors that perform Levy flights, level of plant hydric and nutritional stress, and seasonal climatic effects, are included. The observed epidemic data were quantitatively reproduced by the proposed model on varying the parameters controlling vector motility, plant stress, and initial population of diseased plants. PMID- 11102059 TI - Molecular chirality and domain shapes in lipid monolayers on aqueous surfaces. AB - The shapes of domain boundaries in the mesoscopic phase separation of phospholipids in aqueous surface monolayers are analyzed with particular attention to the influence of molecular chirality. We have calculated equilibrium shapes of such boundaries, and show that the concept of spontaneous curvature derived from an effective pair potential between the chiral molecules-yields an adequate description of the contribution of chirality to the total energy of the system. For enantiomeric dipalmitoylphosphatidylcholine in pure monolayers, and in mixtures with impurities that adsorb preferentially at the (one-dimensional) boundary line between the isotropic and anisotropic fluid phases, such as cyanobiphenyl (5CB), a total energy term that includes line tension, electrostatic dipole-dipole interaction, and spontaneous curvature is sufficient to describe the shapes of well-separated domain boundaries in full detail. As soon as interdomain distances fall below the domain sizes upon compression of a monolayer, fluctuations take over in determining its detailed structural morphology. Using Minkowski measures for the well-studied dimyristoyl phosphatidic acid (DMPA)/cholesterol system, we show that calculations accounting for line tension, electrostatic repulsion, and molecular chirality yield boundary shapes that are of the same topology as the experimentally observed structures. At a fixed molecular area in the phase coexistence region, the DMPA/cholesterol system undergoes an exponential decay of the line tension lambda with decreasing subphase temperature T. PMID- 11102060 TI - NaDNA-bipyridyl-(ethylenediamine)platinum (II) complex: structure in oriented wet spun films and fibers. AB - Complexes of NaDNA with bipyridyl-(ethylenediamine)platinum(II) (abbreviated [(bipy)Pt(en)](2+)) in solid, oriented films, prepared with a wet-spinning method, have been studied using x-ray diffraction, elastic neutron scattering, two-dimensional magic-angle-spinning nuclear magnetic resonance (NMR), infrared (IR) linear dichroism, and IR absorption. All of these experiments indicate that the DNA in this complex is in the B conformation. The neutron diffraction experiments reveal that the rise per residue is 3.31 A, indicating that the [(bipy)Pt(en)](2+) molecular ion causes a small distortion of the B conformation. The neutron data in the direction perpendicular to the helical axis are consistent with a centered orthorhombic unit cell with a=22.65 A and b=32.2 A. The NMR and IR experiments show that the orientation of phosphate groups in the DNA small middle dot[(bipy)Pt(en)](2+) complex is the same as that observed for pure DNA in the B conformation. The IR experiments also show that the [(bipy)Pt(en)](2+) molecular ion stabilizes the B conformation of DNA down to 59% relative humidity, a low water activity. Mechanochemical experiments on wet-spun NaDNA fibers in 68% ethanol with and without [(bipy)Pt(en)](2+) reveal a 9% elongation of the DNA fibers as the complex is formed. PMID- 11102061 TI - Exact solution of site and bond percolation on small-world networks. AB - We study percolation on small-world networks, which has been proposed as a simple model of the propagation of disease. The occupation probabilities of sites and bonds correspond to the susceptibility of individuals to the disease, and the transmissibility of the disease respectively. We give an exact solution of the model for both site and bond percolation, including the position of the percolation transition at which epidemic behavior sets in, the values of the critical exponents governing this transition, the mean and variance of the distribution of cluster sizes (disease outbreaks) below the transition, and the size of the giant component (epidemic) above the transition. PMID- 11102062 TI - Symmetry breaking and coarsening in spatially distributed evolutionary processes including sexual reproduction and disruptive selection. AB - Sexual reproduction presents significant challenges to formal treatment of evolutionary processes. A starting point for systematic treatments of ecological and evolutionary phenomena has been provided by the gene-centered view of evolution which assigns effective fitness to each allele instead of each organism. The gene-centered view can be formalized as a dynamic mean-field approximation applied to genes in reproduction and selection dynamics. We show that the gene-centered view breaks down for symmetry breaking and pattern formation within a population and show that spatial distributions of organisms with local mating neighborhoods in the presence of disruptive selection give rise to such symmetry breaking and pattern formation in the genetic composition of local populations. Global dynamics follows conventional coarsening of systems with nonconserved order parameters. The results have significant implications for the ecology of genetic diversity and species formation. PMID- 11102063 TI - Phase transitions and steady-state microstructures in a two-temperature lattice gas model with mobile active impurities AB - The nonequilibrium, steady-state phase transitions and the structure of the different phases of a two-dimensional system with two thermodynamic temperatures are studied via a simple lattice-gas model with mobile active impurities ("hot/cold spots") whose activity is controlled by an external drive. The properties of the model are calculated by Monte Carlo computer-simulation techniques. The two temperatures and the external drive on the system lead to a rich phase diagram including regions of microstructured phases in addition to macroscopically ordered (phase-separated) and disordered phases. Depending on the temperatures, microstructured phases of both lamellar and droplet symmetry arise, described by a length scale that is determined by the characteristic temperature controlling the diffusive motion of the active impurities. PMID- 11102064 TI - Calculating the local solvent chemical potential in crystal hydrates. AB - Determining solvation patterns in biological systems is crucial in investigating the functional role water may play in structural stabilization and molecular recognition. Determining whether a particular position would be occupied by a solvent molecule requires the local thermodynamics to be known. In this work we introduce a simple and inexpensive approach based on grand canonical molecular simulations to determine the occupancy factors of the cavities. The method is applied to the test case of the sodium salt of hyaluronic acid. The results agree very well with experimental results and demonstrate the success of the method. PMID- 11102065 TI - Force-velocity relation for growing biopolymers. AB - The process of force generation by the growth of biopolymers is simulated via a Langevin-dynamics approach. The monomer-monomer interaction forces are taken to have simple forms that favor the growth of straight fibers from solution, and they are taken to grow against a flat obstacle. The force-velocity relation is obtained from the simulations for two versions of the monomer-monomer force field. We evaluate corrections to the simplest analytic theory based on thermal motion of the obstacle, which yields an exponential velocity decay with applied force. For most orientations of the growing fiber tip, the corrections are small. However, for orientations in which the surface of the growing fiber is parallel to the obstacle, large corrections are obtained in the direction of reduced fiber velocity. These results are explained on the basis of the diffusion properties of monomers near the fiber tip. It is also found that the mobility of the obstacle has little effect on the growth rate over a broad range of possible values. PMID- 11102066 TI - Generalization properties of finite-size polynomial support vector machines AB - The learning properties of finite-size polynomial support vector machines are analyzed in the case of realizable classification tasks. The normalization of the high-order features acts as a squeezing factor, introducing a strong anisotropy in the patterns distribution in feature space. As a function of the training set size, the corresponding generalization error presents a crossover, more or less abrupt depending on the distribution's anisotropy and on the task to be learned, between a fast-decreasing and a slowly decreasing regime. This behavior corresponds to the stepwise decrease found by Dietrich et al. [Phys. Rev. Lett. 82, 2975 (1999)] in the thermodynamic limit. The theoretical results are in excellent agreement with the numerical simulations. PMID- 11102067 TI - Proteins with selected sequences: a heteropolymeric study. AB - Protein sequences are expected not to be random but selected in order to form a stable native structure that is kinetically accessible. Therefore our model contains a selective temperature in sequence space (see [S. Ramanathan and E. Shakhnovich, Phys. Rev. E 50, 1303 (1994)] ) to optimize the sequence for the target conformation statistically. Replica calculations, which go beyond quadratic approximations in the field-theoretical Hamiltonian, are presented. A phase diagram indicating the temperatures and selective temperatures at which transitions to a frozen globule, i.e., the native state, occur is obtained. It is shown that going beyond the quadratic approximation in the field Hamiltonian is very important, since it results in a significant change of the phase diagram. Moreover, we suggest that a one-step replica permutation symmetry scheme is sufficient to solve the model. In addition to this we present a result for the sequence correlation function along the chain in the case of a short-ranged potential between the monomers. A correlation function between monomers that form a contact in the native state is given depending on the temperature and the interaction parameter. PMID- 11102068 TI - Information transmission and recovery in neural communications channels. AB - Biological neural communications channels transport environmental information from sensors through chains of active dynamical neurons to neural centers for decisions and actions to achieve required functions. These kinds of communications channels are able to create information and to transfer information from one time scale to the other because of the intrinsic nonlinear dynamics of the component neurons. We discuss a very simple neural information channel composed of sensory input in the form of a spike train that arrives at a model neuron, then moves through a realistic synapse to a second neuron where the information in the initial sensory signal is read. Our model neurons are four dimensional generalizations of the Hindmarsh-Rose neuron, and we use a model of chemical synapse derived from first-order kinetics. The four-dimensional model neuron has a rich variety of dynamical behaviors, including periodic bursting, chaotic bursting, continuous spiking, and multistability. We show that, for many of these regimes, the parameters of the chemical synapse can be tuned so that information about the stimulus that is unreadable at the first neuron in the channel can be recovered by the dynamical activity of the synapse and the second neuron. Information creation by nonlinear dynamical systems that allow chaotic oscillations is familiar in their autonomous oscillations. It is associated with the instabilities that lead to positive Lyapunov exponents in their dynamical behavior. Our results indicate how nonlinear neurons acting as input/output systems along a communications channel can recover information apparently "lost" in earlier junctions on the channel. Our measure of information transmission is the average mutual information between elements, and because the channel is active and nonlinear, the average mutual information between the sensory source and the final neuron may be greater than the average mutual information at an earlier neuron in the channel. This behavior is strikingly different than the passive role communications channels usually play, and the "data processing theorem" of conventional communications theory is violated by these neural channels. Our calculations indicate that neurons can reinforce reliable transmission along a chain even when the synapses and the neurons are not completely reliable components. This phenomenon is generic in parameter space, robust in the presence of noise, and independent of the discretization process. Our results suggest a framework in which one might understand the apparent design complexity of neural information transduction networks. If networks with many dynamical neurons can recover information not apparent at various waystations in the communications channel, such networks may be more robust to noisy signals, may be more capable of communicating many types of encoded sensory neural information, and may be the appropriate design for components, neurons and synapses, which can be individually imprecise, inaccurate "devices." PMID- 11102069 TI - Single and multiple topologically driven structural transitions in DNA. AB - We derive some exact general results concerning the behavior of topological absorbers (i.e., sequences undergoing topologically driven structural transitions) in closed circular DNA molecules. Starting from the formal physical framework that covers all known structural transitions, like those from standard B-DNA to nonstandard conformers Z-DNA, H-DNA, cruciform-DNA, melt-DNA or others, we develop a reduced state space description that leads to an analytically simplified "black box" view of absorbers. The latter contains only a single state variable-the total sequence unwinding u describing the topological state of the absorber. We show that the statistical mechanics of u is determined by the (one dimensional) absorption free energy function G(abs) and find explicit expressions for G(abs) and for moments in terms of the standard experimental observable-the absorption function alpha:=. The reduced state space method is then applied to systems consisting of several interacting topologically coupled absorbers and a formula predicting their collective behavior (superposition) in terms of their individual absorptions is derived. Using these results we formulate and discuss solution methods for two basic types of inverse problems that turn out to be fundamental for future absorber construction. PMID- 11102070 TI - Fluctuating filaments: statistical mechanics of helices AB - We examine the effects of thermal fluctuations on thin elastic filaments with noncircular cross section and arbitrary spontaneous curvature and torsion. Analytical expressions for orientational correlation functions and for the persistence length of helices are derived, and it is found that this length varies nonmonotonically with the strength of thermal fluctuations. In the weak fluctuation regime, the local helical structure is preserved and the statistical properties are dominated by long-wavelength bending and torsion modes. As the amplitude of fluctuations is increased, the helix "melts" and all memory of intrinsic helical structure is lost. Spontaneous twist of the cross section leads to resonant dependence of the persistence length on the twist rate. PMID- 11102071 TI - Aggregation models at high packing fraction AB - Dense phases of micellar aggregates have strong molecular correlation at two different levels: that of the molecules forming a micelle and that between micelles, leading to a possible phase transition from a micellar fluid to a micellar crystal. The global phase diagram may also include lamellar and other dense phases, which do not have a micellar structure. We present here a generic approach to deal with these systems through a two-level density-functional description, to first describe an isolated micellar aggregate and then the dense micellar system, obtaining the free energy in a self-consistent way from the molecular interactions. Nonmicellar dense phases are included with the same density-functional approach applied at the first level. The results are shown to be very accurate for a one-dimensional model with exact solution, and the method is then applied to a three-dimensional amphiphile model that had been successfully used to describe the properties of diluted amphiphile solutions. PMID- 11102072 TI - Nonlocal electron kinetics in a planar inductive helium discharge AB - A measurement of the electron energy distribution function (EEDF) using the ac superposition method is done over a helium pressure range of 10-100 mTorr in a planar inductive plasma, and the electron energy diffusion coefficient which describes the electron heating is calculated based on the same discharge conditions using a two-dimensional simulation. It is found that the measured EEDF shows a bi-Maxwellian distribution with a low-energy electron group at low pressures below 20 mTorr even in the inductive discharge using helium of the non Ramsauer gas. The major factors which can affect the EEDF formation are investigated. In particular, the concept of the total electron bounce frequency, i.e., the electron residence time, is introduced as an indicator of how the electron-electron collision affects the EEDF shape. As a result, it is shown that the observed bi-Maxwellian distribution at low pressures is attributed to the combined effects of the formation of low-energy electrons through the cooling mechanism of energetic electrons enhanced by the capacitive field, the low heating rate of the low-energy electrons, the confinement of low-energy electrons by the ambipolar space potential, and the low electron-electron collision frequency which can be estimated from the total electron bounce frequency presented in this paper. PMID- 11102073 TI - Basic physics of laser propagation in hollow waveguides AB - The basic theory of laser propagation in hollow waveguides is considered in the context of laser-plasma physics. The physical model of waves reflecting between the guide walls is used to show that there is a discrete series of modes, and to give the mode dispersion relation and losses in terms of a given reflectivity. The mathematical connection between this model and the solution of Maxwell's equations for lossless propagation in a cylinder is given. Thus the solutions for low loss propagation for any given reflectivity can be obtained, provided it is close to 1. Results are given using Fresnel reflectivity for perfect dielectric and finite conductivity waveguides. The relationship of the breakdown intensity in dielectric waveguides to known breakdown intensities is also derived. The practical implications for the guiding of intense laser pulses and the limitations of the model are discussed. The theory is shown to explain, at least qualitatively, a number of previous experimental results. PMID- 11102074 TI - Ball lightning as a force-free magnetic knot AB - The stability of fireballs in a recent model of ball lightning is studied. It is shown that the balls shine while relaxing in an almost quiescent expansion, and that three effects contribute to their stability: (i) the formation in each one during a process of Taylor relaxation of a force-free magnetic field, a concept introduced in 1954 in order to explain the existence of large magnetic fields and currents in stable configurations of astrophysical plasmas; (ii) the so called Alfven conditions in magnetohydrodynamics; and (iii) the approximate conservation of the helicity integral. The force-free fields that appear are termed "knots" because their magnetic lines are closed and linked. PMID- 11102075 TI - Transport coefficients for dense metal plasmas AB - Thermoelectric transport coefficients of metal plasmas are calculated within the linear response theory applied previously to determine the electrical conductivity of Al and Cu plasmas [R. Redmer, Phys. Rev. E 59, 1073 (1999)]. We consider temperatures of 1-3 eV and densities of 0.001-1 g/cm(3) as relevant in rapid wire evaporation experiments. The plasma composition is calculated considering higher ionization stages of atoms up to 5+, and solving the respective system of coupled mass action laws. Interactions between charged particles are treated on T matrix level. Results for the electrical conductivity of various metal plasmas are in reasonable agreement with experimental data. Thermal conductivity and thermopower are also given. In addition, we compare with experimental data for temperatures up to 25 eV and liquidlike densities. PMID- 11102076 TI - Measurement method for electric fields based on stark spectroscopy of argon atoms AB - We report the development of a method for the measurement of electric fields in glow discharge plasmas, based on Stark spectroscopy of argon atoms. The method is based on laser excitation of transitions in atomic argon. The key feature of the method is that the electric field is determined by matching experimentally obtained absorption spectra to theoretically calculated spectra. The dependence of the positions of energy levels of argon atoms on the strength of the electric field was calculated by solving the Schrodinger equation for the argon atom. Measurements of Stark spectra were made in the sheath region of a glow discharge using laser optogalvanic spectroscopy. The wavelength of the laser radiation was tuned to the transitions 4s-->nf (n=7,8,ellipsis,14) of the argon atom. For n=11, the lower limit for electric field measurements was estimated to be 14 V/mm. PMID- 11102077 TI - Dynamics of a microcapillary discharge plasma using a soft x-ray laser backlighter AB - We have used the new technique of soft x-ray laser shadowgraphy in combination with traditional plasma emission spectroscopy and theoretical modeling to study the dynamics of a plasma column created by a discharge through a 380 &mgr;m diameter evacuated microcapillary. The transient microcapillary plasma was imaged with high-spatial and temporal resolution using a tabletop discharge pumped 46.9 nm laser backlighter. Model computations show that the sharp boundary observed between the absorbent and transparent regions of the shadowgrams is defined by the spatial distribution of weakly ionized ions that are strongly photoionized by the probe laser. The plasma was observed to rapidly evolve from an initially nonuniform distribution into a column with good azimuthal symmetry and minimum density on axis [computed electron density on axis n(e)=(1-3)x10(19) cm(-3)]. This concave electron density profile constitutes a plasma waveguide for laser radiation. Heated solely by Joule dissipation from relatively small excitation currents (1.5 kA), this dense plasma reaches substantial electron temperatures of T(e)=15-20 eV as a result of the absence of significant hydrodynamic losses and reduced radiation losses caused by large spectral line opacities. The results illustrate the potential of tabletop soft x-ray lasers as a new plasma diagnostic tool. PMID- 11102078 TI - Internally driven spatiotemporal irregularity in a dc glow discharge AB - Spatiotemporal dynamics of an undriven dc glow discharge at intermediate pressures (p(0)r(0)=6.2 Torr cm, i<50 mA) is investigated experimentally. Spatiotemporal irregularity and windows of regular nonlinear waves occur and are found to depend on the discharge current. Above a threshold current column head oscillations arise and inject high-frequency ionization waves into the positive column that decay towards the anode through nonlinear wave coupling with a discrete eigenmode of the positive column. Regularity was found to be a result of commensuration of both waves and obeys a devil's staircase. Since column head oscillations occur in the transition region from cathode fall to positive column as result of discharge formation, the irregularities were internally driven. Spatiotemporal analysis by means of biorthogonal decomposition gives insights into the mechanism of irregularity and can be employed for characterization of spatiotemporal complexity. PMID- 11102079 TI - Potential formalism for an axial energy cumulation process AB - A central filament is assumed to be axially positioned inside a collapsing cylindrical thin shell. A cumulation process is induced by generating a high magnetic field between the shell and the filament. Assuming a dissipationless approach, it is shown that this problem is equivalent to two point particles moving in a potential. PMID- 11102080 TI - Prepulse effects on the interaction of intense femtosecond laser pulses with high Z solids AB - Kalpha emission of high-Z solid targets irradiated by an intense, short (<100 fs) laser pulse in the 10 keV region is shown to be sensitive to the electron energy cutoff, which is strongly dependent on the density gradient of the plasma corona formed by a long prepulse. The absorption rate of short laser pulses, the hot electron distribution, and x-ray emission from a Cu slab target are studied via a hybrid model, which combines the hydrodynamics, collisional particle-in-cell, and Monte Carlo simulation techniques, and via a direct spectroscopic measurement. An absorption mechanism originating from the interaction of the laser pulse with plasma waves is found to increase the absorption rate by over 30% even for a very short, s-polarized laser pulse. Calculated and measured x-ray spectra are in good agreement, confirming the electron energy cutoff. PMID- 11102081 TI - On-off intermittency at the onset of the ion-acoustic instability in a laboratory plasma AB - The ion-ion beam instability is experimentally studied just above threshold in a laboratory double-plasma device. Intermittent bursts of unstable waves are recorded in the target plasma and the distribution of the recurrence times of the bursts is estimated. At the onset of instability, the measurements are in agreement with the expected evolution deduced from a theoretical model that combines the normal form of a supercritical Hopf-Andronov bifurcation and the parametric noisy deviations from threshold typical of on-off intermittency. In both the experiment and the model, the distribution of the recurrence times of the bursts decays as an inverse power law and the evolution of the mean laminar length when the control parameter is increased beyond threshold exhibits a power law of exponent -1. PMID- 11102082 TI - Observation of spatial asymmetry of THz oscillating electron plasma wave in a laser wakefield AB - The asymmetric spatial distribution of electron density perturbation is observed by using a frequency-domain interferometry technique. The wake amplitude of the outside bump is enhanced by the elliptical distribution of the pump laser pulse. This asymmetry can be explained with a two-dimensional analytical model expanded from cylindrically symmetric linear theory. PMID- 11102083 TI - Detailed-term-accounting-approximation simulation of x-ray transmission through laser-produced Al plasmas AB - An extensive configuration interaction (CI) scheme and the R-matrix method are combined to calculate the x-ray transmission spectrum for high-power laser produced Al plasmas in local thermodynamic equilibrium by using the detailed-term accounting (DTA) approximation. All atomic parameters such as state levels and photoabsorption cross sections for different ionization stages are obtained by using the CI and R-matrix method. Special attention is given to the effects of autoionizing resonance broadening on the transmission. A large difference exists between the convergence of the results with and without taking account of autoionizing resonance broadening when the autoionization resonance broadening is the major broadening mechanism. This shows that autoionizing resonance widths of the K-shell excited states have large effects and should be considered to interpret the spectral-resolved transmission. PMID- 11102084 TI - Generation of one-cycle laser pulses by use of high-amplitude plasma waves AB - The dynamics of a short laser pulse located in the density trough of a background plasma wave is investigated and a scheme is proposed to compress the pulse duration by use of a high-amplitude plasma wave. The threshold amplitude of the plasma wave, at which the compressing effect just balances the dispersive spreading of the laser pulse, is estimated for certain pulse profiles. Numerical simulations are conducted with particle-in-cell codes, where a pump pulse is used to generate a high-amplitude plasma wave and a signal pulse copropagates behind. It is shown that the signal pulse can be compressed by the plasma wave from ten laser cycles to about one cycle within a millimeter in tenuous plasma only a few percent of the critical density. PMID- 11102085 TI - Diffusive alignment of the magnetic field in active regions of plasmas AB - Regions of high magnetic field within plasmas tend to keep this field aligned in a dominant direction. This occurs both in observed phenomena and in simulations of kinematic and nonlinear dynamos. Although most of this effect is due to the particular dynamics of each case, magnetic diffusion also plays an important role. It is shown here that the unitary magnetic field vector satisfies a certain estimate that bounds its possible variations. The dependence of the bound on the plasma parameters is analyzed. PMID- 11102086 TI - High density collimated beams of relativistic ions produced by petawatt laser pulses in plasmas AB - Under optimal interaction conditions ions can be accelerated up to relativistic energies by a petawatt laser pulse in both underdense and overdense plasmas. Two dimensional particle in cell simulations show that the laser pulse drills a channel through an underdense plasma slab due to relativistic self-focusing. Both ions and electrons are accelerated in the head region of the channel. However, ion acceleration is more effective at the end of the slab. Here electrons from the channel expand in vacuum and are followed by the ions dragged by the Coulomb force arising from charge separation. A similar mechanism of ion acceleration occurs when a superintense laser pulse interacts with a thin slab of overdense plasma and the pulse ponderomotive pressure moves all the electrons away from a finite-diameter spot. PMID- 11102087 TI - Cyclotron resonance maser experiments in a bifilar helical waveguide AB - Oscillator and amplifier cyclotron-resonance-maser (CRM) experiments in a spiral bifilar waveguide are presented in this paper. The slow-wave CRM device employs a low-energy low-current electron beam (2-12 keV, approximately 0.5 A). The pitch angle of the helical waveguide is relatively small; hence, the phase velocity in this waveguide, V(ph) congruent with0.8c (where c is the speed of light), is much faster than the axial velocity of the electrons, V(ez)2%). The wide tunable range of this CRM device due to the nondispersive bifilar helix is discussed. PMID- 11102088 TI - Simulation of beam-beam effects in a circular e(+)e(-) collider AB - We have carried out a strong-strong simulation to study the beam-beam effect in a circular electron-positron collider. In the simulation, both the positron and electron beams are represented by macroparticles, and the interaction between the two beams is obtained by solving the Poisson equation for the charge distribution of the macroparticles. Using the simulation, we investigate the beam-beam limit and the coherent beam-beam tune shift, two characteristic phenomena of the beam beam effect. We also study collision with a finite crossing angle, and verify its feasibility for the KEKB factory. Our results for KEKB yield an estimated luminosity of 8.5x10(33) cm(-2) s(-1), only slightly lower than the design value of 1x10(34) cm(-2) s(-1). PMID- 11102089 TI - Three-dimensional analysis of harmonic generation in high-gain free-electron lasers AB - In a high-gain free-electron laser (FEL) employing a planar undulator, strong bunching at the fundamental wavelength can drive substantial bunching and power levels at the harmonic frequencies. In this paper we investigate the three dimensional evolution of harmonic radiation based on the coupled Maxwell Klimontovich equations that take into account nonlinear harmonic interactions. Each harmonic field is a sum of a linear amplification term and a term driven by nonlinear harmonic interactions. After a certain stage of exponential growth, the dominant nonlinear term is determined by interactions of the lower nonlinear harmonics and the fundamental radiation. As a result, the gain length, transverse profile, and temporal structure of the first few harmonics are eventually governed by those of the fundamental. Transversely coherent third-harmonic radiation power is found to approach 1% of the fundamental power level for current high-gain FEL projects. PMID- 11102090 TI - Lorentz torque on a charged sphere rotating in a dielectric fluid in the presence of a uniform magnetic field AB - The Lorentz torque exerted by a uniform magnetic field on a charged sphere rotating steadily in a dielectric fluid is calculated to first order in the charge. For a strongly polar fluid and stick boundary conditions the torque is enhanced significantly with respect to its vacuum value. The modification from the vacuum value depends only on the static dielectric constant of the fluid and on the slip parameter. It is independent of the dielectric response of the sphere and of the shape of the radial charge distribution. There is a nonvanishing Lorentz torque, even when the charge is concentrated in the center of the sphere. PMID- 11102091 TI - Supersonic and multiple topological excitations in the driven frenkel-kontorova model with exponential interaction AB - The criteria for the existence of supersonic and multiple topological excitations (kinks) in the driven Frenkel-Kontorova model (a chain of atoms placed into an external periodic potential) with anharmonic (exponential) interatomic interactions are studied. PMID- 11102092 TI - Ansatz-independent solution of a soliton in a strong dispersion-management system AB - We introduce a theoretical approach to the study of propagation in systems with periodic strong-management dispersion. Our approach does not assume any ansatz about the form of the solution nor does it make use of any average procedure. We find an explicit solution for the pulse evolution in the fast dynamics regime (distances smaller than the dispersion period). We also establish the equation of motion governing the slow dynamics of an arbitrary pulse and prove that the pulse evolution is nonlinear and Hamiltonian. We solve this equation and find that a nonlinear solitonlike solution occurs self-consistently in the form of an asymptotic stationary eigenfunction of the Hamiltonian. PMID- 11102093 TI - Energy-flux pattern in the goos-Hanchen effect AB - The (two-dimensional) wave beam fields and the associated energy flux in the Goos Hanchen effect are studied analytically and numerically. In particular, the time averaged Poynting vector and its flux lines around the interface are calculated for both states of wave polarization, irrespective of a positive shift (the usual one) or a negative shift of the reflected beam. For a given transverse field profile of the incident beam, the flux lines associated with the evanescent waves for the two cases of shift are shown to have the same shape yet to take opposite directions; they are parabolas if the profile is Gaussian. The flux lines in the first medium are shown to connect to those in the second medium on the two sides of the interface. In the case of positive shift, the whole flux pattern expectedly shows the supply of energy from the incident beam to the evanescent wave on one side and the return of energy on the other side to the reflected beam. In the case of negative shift, on the other hand, the flux lines nearby the interface form loops, in addition to the expected incoming-outgoing flux pattern in the remaining region. PMID- 11102094 TI - Three-dimensional walking spatiotemporal solitons in quadratic media AB - Two-parameter families of chirped stationary three-dimensional spatiotemporal solitons in dispersive quadratically nonlinear optical media featuring type-I second-harmonic generation are constructed in the presence of temporal walk-off. Basic features of these walking spatiotemporal solitons, including their dynamical stability, are investigated in the general case of unequal group velocity dispersions at the fundamental and second-harmonic frequencies. In the cases when the solitons are unstable, the growth rate of a dominant perturbation eigenmode is found as a function of the soliton wave number shift. The findings are in full agreement with the stability predictions made on the basis of a marginal linear-stability curve. It is found that the walking three-dimensional spatiotemporal solitons are dynamically stable in most cases; hence in principle they may be experimentally generated in quadratically nonlinear media. PMID- 11102095 TI - Field and intensity correlation in random media AB - We have obtained the spectral and spatial field correlation functions, C(E)(Deltaomega) and C(E)(Deltax), respectively, from measurement of the microwave field spectrum at a series of points along a line on the output of a random dielectric medium. C(E)(Deltaomega) and C(E)(Deltax) are shown to be the Fourier transforms, respectively, of the time of flight distribution, obtained from pulsed measurements, and of the specific intensity. Unlike C(E)(Deltaomega), the imaginary part of C(E)(Deltax) is shown to vanish as a result of the isotropy of the correlation function in the output plane. The complex square of the field correlation function gives the short-range or C1 contribution to the intensity correlation function C. Longer-range contributions to the intensity correlation function are obtained directly by subtracting C1 from C and are in good agreement with theory. PMID- 11102096 TI - Spontaneous pattern formation in driven nonlinear lattices AB - We demonstrate the spontaneous formation of spatial patterns in a damped, ac driven cubic Klein-Gordon lattice. These patterns are composed of arrays of intrinsic localized modes characteristic for nonlinear lattices. We analyze the modulation instability leading to this spontaneous pattern formation. Our calculation of the modulational instability is applicable in one- and two dimensional lattices; however, in the analyses of the emerging patterns we concentrate particularly on the two-dimensional case. PMID- 11102097 TI - Ground states of dispersion-managed nonlinear Schrodinger equation AB - An exact pulse for the parametrically forced nonlinear Schrodinger equation (NLS) is isolated. The equation governs wave envelope propagation in dispersion-managed fiber lines with positive residual dispersion. The pulse is obtained as a ground state of an averaged variational principle associated with the equation governing pulse dynamics. The solutions of the averaged and original equations are shown to stay close for a sufficiently long time. A properly adjusted pulse will therefore exhibit nearly periodic behavior in the time interval of validity of the averaging procedure. Furthermore, we show that periodic variation of dispersion can stabilize spatial solitons in a Kerr medium and one-dimensional solitons in the NLS with quintic nonlinearity. The results are confirmed by numerical simulations. PMID- 11102098 TI - Diffusion of waves in a layer with a rough interface AB - Sound trapped between the rough boundaries of a statistically homogeneous layer can exhibit diffusive behavior which influences the reverberation of a pulsed signal. If the interfaces of the layer can transmit sound, then eventually sound within the layer will be lost and there will be no diffusion. Nevertheless, if the transmission to the exterior of the layer is weak, there will be a remnant of diffusion. This paper examines the description of this kind of quasidiffusive behavior for sound which impinges on the rough interface of such a layer from outside the layer. As in the case of diffuse light scattering by particles in suspension, the diffusion constant is renormalized according to the delay required to build up resonant energy in the layer. In addition, when there is a density contrast between the interior and exterior of the layer, or when there is dispersion, the diffusion constant has another correction associated with energy flux within the layer. PMID- 11102099 TI - Optical addressing at the subwavelength scale AB - The Green dyadic formalism is applied to the study of the optical properties of dielectric subwavelength structures integrated in coplanar geometry. We first consider homogeneous wires with high refractive index featuring subwavelength cross sections. We show that such wires may have guiding properties and that they may be coupled with a local illumination produced by a focused Gaussian beam totally reflected at the substrate interface. When excited by the focused beam, these subwavelength optical waveguides (SOW's) provide a confined source of light that could be used to excite a single nanoscopic object. Well designed heteregeneous wires resulting from the alignment of dielectric particles separated from each other by a subwavelength distance are also found to propagate a Gaussian beam excitation over several micrometers. This propagation occurs with reasonable damping for incident beams in the visible frequency range. The computed transmission spectra of these heterowires may exhibit narrow gaps. Finally, we discuss the relation between the optical properties of the SOW and the calculated electromagnetic local density of states. PMID- 11102100 TI - Scattering-theory analysis of waveguide-resonator coupling AB - Using a formalism similar to the quantum scattering theory, we analyze the problem of coupling between optical waveguides and high Q resonators. We give the optical transmission and reflection coefficients as functions of the waveguide resonator coupling, cavity loss (gain), and cavity resonant frequency. Based on these results, the recently proposed concept of "critical coupling" is discussed. Using a matrix formalism based on the scattering analysis, we find the dispersion relation of indirectly coupled resonator optical waveguides. The coupling between waveguides and multiple cavities is investigated and the reflection and transmission coefficients are derived. PMID- 11102101 TI - General electromagnetic density of modes for a one-dimensional photonic crystal AB - In this paper, we present more general, exact, and concise expressions for calculating the electromagnetic density of modes (EDOM) in one dimension photonic crystal (superlattice) for E and H polarizations. The expression is used for numerical computation of the EDOM in the lower-index (dielectric constant) layer. We discuss the difference between the EDOM in high- and low-index layers as due to the presence of waveguiding modes and evanescent-excited Bloch modes in the higher-index layer. Two methods of computation are presented to compute the EDOM in the lower-index layer. We suggest the possibility of using the EDOM to establish population inversion, which may be useful for higher-frequency lasers (e.g., x rays) and control any radiative processes. We also elaborate on the limitations of the results of Alvarado Rodriguez et al. as due to the approximation used in the evaluation of partial differentialomega/ partial differentialk(y,z) for nabla(k)omega and comment on the limitations of the one dimensional EDOM expression of Bendickson et al. PMID- 11102102 TI - Stabilization of dark solitons in the cubic ginzburg-landau equation AB - The existence and stability of exact continuous-wave and dark-soliton solutions to a system consisting of the cubic complex Ginzburg-Landau (CGL) equation linearly coupled with a linear dissipative equation is studied. We demonstrate the existence of vast regions in the system's parameter space associated with stable dark-soliton solutions, having the form of the Nozaki-Bekki envelope holes, in contrast to the case of the conventional CGL equation, where they are unstable. In the case when the dark soliton is unstable, two different types of instability are identified. The proposed stabilized model may be realized in terms of a dual-core nonlinear optical fiber, with one core active and one passive. PMID- 11102103 TI - Superluminal tunneling of an electromagnetic X wave through a planar slab AB - A study is provided of an X wave undergoing frustrated total internal reflection on the upper surface of a planar slab separating two dielectric media. It is shown that the peak of the field transferred through the slab appears to be transmitted at an ultrafast speed. A general analytic solution is derived and is supported by specific numerical examples that indicate that the transport speed of the peak of the X wave can be much larger than the speed of light. PMID- 11102104 TI - Broad histogram method: extension and efficiency test AB - Compared to standard histogram techniques, the broad histogram method allows us to increase the efficiency of Monte Carlo simulations by a tremendous amount. This gain of efficiency is achieved by measuring simulation averages of particular system observables (different from those used in standard histogram techniques), while the algorithm of the simulation can be left unchanged. In this paper, the broad histogram method is reformulated in a more mathematical and precise way. Furthermore, the method is extended to estimate the density of states as a function of more than one parameter. A quantitative investigation of the gain of efficiency of Monte Carlo simulations is performed. For the broad histogram method, we find a gain of efficiency which amounts to orders of magnitude in comparison to the standard histogram method. PMID- 11102105 TI - Second-order stochastic leapfrog algorithm for multiplicative noise brownian motion AB - A stochastic leapfrog algorithm for the numerical integration of Brownian motion stochastic differential equations with multiplicative noise is proposed and tested. The algorithm has a second-order convergence of moments in a finite time interval and requires the sampling of only one uniformly distributed random variable per time step. The noise may be white or colored. We apply the algorithm to a study of the approach towards equilibrium of an oscillator coupled nonlinearly to a heat bath and investigate the effect of the multiplicative noise (arising from the nonlinear coupling) on the relaxation time. This allows us to test the regime of validity of the energy-envelope approximation method. PMID- 11102106 TI - Ground state of trapped interacting bose-einstein condensates by an explicit imaginary-time algorithm AB - We show that an explicit time-marching method previously developed for the numerical study of the dynamics of Bose-Einstein condensates can be profitably adapted to the numerical determination of their ground state. After reduction to a one-dimensional model, we first reproduce and test known results on condensates in harmonic traps and then determine the ground state of a condensate in a harmonically bound optical lattice in the range of parameters which are relevant to existing experiments. PMID- 11102107 TI - Approach to ergodicity in monte carlo simulations AB - The approach to the ergodic limit in Monte Carlo simulations is studied using both analytic and numerical methods. With the help of a stochastic model, a metric is defined that enables the examination of a simulation in both the ergodic and nonergodic regimes. In the nonergodic regime, the model implies how the simulation is expected to approach ergodic behavior analytically, and the analytically inferred decay law of the metric allows the monitoring of the onset of ergodic behavior. The metric is related to previously defined measures developed for molecular dynamics simulations, and the metric enables the comparison of the relative efficiencies of different Monte Carlo schemes. Applications to Lennard-Jones 13-particle clusters are shown to match the model for Metropolis, J-walking, and parallel tempering based approaches. The relative efficiencies of these three Monte Carlo approaches are compared, and the decay law is shown to be useful in determining needed high temperature parameters in parallel tempering and J-walking studies of atomic clusters. PMID- 11102108 TI - Naudts-like duality and the extreme fisher information principle AB - We show that using Frieden and Soffer's extreme information principle [Phys. Rev. E 52, 2274 (1995)] with a Fisher measure constructed with escort probabilities [C. Beck and F. Schlogel, Thermodynamics of Chaotic Systems (Cambridge University Press, Cambridge, England, 1993)], the concomitant solutions obey a type of Naudts's duality (e-print cond-mat/990470) for nonextensive ensembles [C. Tsallis, in Nonextensive Statistical Mechanics and its Applications, Lecture Notes in Physics, edited by S. Abe and Y. Okamoto (Springer-Verlag, Berlin, in press)]. PMID- 11102109 TI - Phase transitions in the kinetic ising model with competing dynamics AB - We study the nonequilibrium phase diagram and critical properties of a two dimensional kinetic Ising model with competing Glauber and Kawasaki dynamics suggested by Tome and de Oliveira [Phys. Rev. A 40, 6643 (1989)]. The role of the Kawasaki dynamics, chosen with probability 1-p, is to simulate a permanent energy flux into the system. The theoretical prediction for the phase diagram is improved significantly by using four- and six-point dynamical mean-field approximations. Monte Carlo simulations support that the ferromagnetic paramagnetic phase transition changes from second to first order for sufficiently small p. The antiferromagnetic phase is found to be stable for a nonzero value of p even at T=0. PMID- 11102110 TI - Studying avalanches in the ground state of the two-dimensional random-field ising model driven by an external field AB - We study the exact ground state of the two-dimensional random-field Ising model as a function of both the external applied field B and the standard deviation sigma of the Gaussian random-field distribution. The equilibrium evolution of the magnetization consists in a sequence of discrete jumps. These are very similar to the avalanche behavior found in the out-of-equilibrium version of the same model with local relaxation dynamics. We compare the statistical distributions of magnetization jumps and find that both exhibit power-law behavior for the same value of sigma. The corresponding exponents are compared. PMID- 11102111 TI - Branching annihilating random walk on random regular graphs AB - The branching annihilating random walk is studied on a random graph whose sites have a uniform number of neighbors (z). The Monte Carlo simulations in agreement with the generalized mean-field analysis indicate that the concentration decreases linearly with the branching rate for z>/=4, while the coefficient of the linear term becomes zero if z=3. These properties are described by a modified mean-field theory taking explicitly into consideration the probability of mutual annihilation of the parent and its offspring particles using the returning features of a single walker on the same graph. PMID- 11102112 TI - Fluctuation-induced transport in a spatially symmetric periodic potential AB - We present an analytical investigation of the fluctuation-induced transport of Brownian particles in a deterministic spatial symmetrical periodic potential subject to Gaussian noises. We found that directed motion of the Brownian particles can be induced by the correlation between a multiplicative white noise and an additive white noise. The direction of current is determined by the sign of the noise correlation. PMID- 11102113 TI - Dynamic approach to weak first-order phase transitions AB - A short-time dynamic approach to weak first-order phase transitions is proposed. Taking the two-dimensional Potts models as examples, from short-time behavior of nonequilibrium relaxational processes starting from high temperature and zero temperature states, pseudo-critical-points K* and K** are determined. A clear difference of the values for K* and K** distinguishes a weak first-order transition from a second-order one. PMID- 11102114 TI - Analytical density functional theory of homogeneous vapor condensation AB - Starting from an exact gradient transcription of the perturbative density functional theory of homogeneous vapor condensation, we propose an analytical approximation that reproduces the density profile and free energy of critical fluctuations to high accuracy. For a broad variety of substances, including nonpolar, weakly polar, and metallic liquids, the method predicts nucleation rates that are orders of magnitude closer to experiment than those from the classical approach. The present treatment incorporates detailed molecular theory into macroscopic modeling. PMID- 11102115 TI - Rigorous solution for the elasticity of diluted gaussian spring networks AB - We present a rigorous solution of the elasticity of the diluted Gaussian spring networks (DGSNs) at zero temperature. We show that the deformation of a diluted DGSN is homogeneous provided that the displacements of the particles on the boundary are homogeneous. It follows that at zero temperature the nonvanishing elastic stiffness coefficients are proportional to the hydrostatic pressure in both two and three dimensions. Follows a rigorous proof of the equivalence of the elasticity of the DGSN and the conductance of the random resistor network at zero temperature. PMID- 11102116 TI - Tsallis maximum entropy principle and the law of large numbers AB - Tsallis has suggested a nonextensive generalization of the Boltzmann-Gibbs entropy, the maximization of which gives a generalized canonical distribution under special constraints. In this Brief Report, we show that the generalized canonical distribution so obtained may differ from that predicted by the law of large numbers when empirical samples are held to the same constraint. This conclusion is based on a result regarding the large deviation property of conditional measures and is confirmed by numerical evidence. PMID- 11102117 TI - Characterization of intermittent lag synchronization AB - Intermittent lag synchronization of two nonidentical symmetrically coupled Rossler systems is investigated. This phenomenon can be seen as a process wherein the intermittent bursts away from the lag synchronization regime correspond to jumps of the system toward other lag configurations. During these jumps, the chaotic trajectory visits closely a periodic orbit. The identification of the different lag configurations and the measure of the fraction of time passed by the system in each one of them provide information on the global scenario of transitions undergone by the system before reaching perfect lag synchronization. PMID- 11102118 TI - Enhancement of phase synchronization through asymmetric couplings AB - Phase synchronization in lattices of coupled chaotic oscillators is studied. It is found that phase synchronization can be greatly improved by asymmetric biased coupling. The mechanism responsible for this effect is the transition from a localized wave to synchronized flow and nonlocal phase synchronization. PMID- 11102119 TI - Experimental verification of the synchronization condition for chaotic external cavity diode lasers AB - The synchronization condition obtained numerically by Ahlers, Parlitz, and Lauterborn [Phys. Rev. E 58, 7208 (1998)] is verified experimentally. PMID- 11102120 TI - Measuring statistical dependence and coupling of subsystems AB - We investigate recently proposed measures for the statistical dependence of systems with complex dynamical behavior. We consider appropriate model systems, to ensure that influences of individual properties of the systems are excluded. We demonstrate that it is indeed possible to obtain nontrivial directional information, but we also argue that the interpretation of this information is difficult. PMID- 11102121 TI - Intermittent chaos in electron scattering AB - The motion of an electron in a uniform magnetic field and positive point charge is not integrable. Phase space is often divided between regular regions farther from the positive charge and chaotic regions nearby. As the electron transits the chaotic region, intermittent chaotic behavior ensues. An analytic method to estimate the location of the transit parameters is also developed. PMID- 11102122 TI - Influence of neighboring bubbles on the primary bjerknes force acting on a small cavitation bubble in a strong acoustic field AB - The primary Bjerknes force on a small bubble in a strong acoustic field in the presence of another bubble is calculated. It is shown that the influence of the bubbles on each other's primary Bjerknes forces is very substantial even if the separation distance between them is large compared with their size. As a result, the peculiarities of the primary forces in strong fields, such as the change of sign with increasing driving pressure amplitude, manifest themselves earlier and more vigorously. The results obtained are of immediate interest for understanding and modeling collective bubble phenomena in strong fields. PMID- 11102123 TI - Equilibrium and nonequilibrium states in thermostated gledzer-ohkitani-yamada shell models AB - We study Gledzer-Ohkitani-Yamada shell models with heat reservoirs. Time reversible Nose-Hoover dynamics are used for the heat reservoirs. Both equilibrium and nonequilibrium cascade states are obtained by changing the heat reservoirs. PMID- 11102124 TI - Excess-entropy and freezing-temperature scalings for transport coefficients: self diffusion in yukawa systems AB - A semiempirical "universal" corresponding states relationship, for the dimensionless transport coefficients of dense fluids as functions of the reduced configurational entropy, was proposed more than 20 years ago and established by many simulations. Recent density functional analysis predicts a universal freezing-temperature scaling for the excess entropy. Combining these properties we derive an approximate corresponding states relationship for the dimensionless transport coefficients of dense fluids as functions of the temperature scaled by the freezing temperature. The temperature scaling observed in recent computer simulation results for self-diffusion in Yukawa systems is just one more case of our general result. PMID- 11102125 TI - Switching behavior and electro-optical properties of liquid crystals in nematic gels AB - Anisotropic nematic gels are prepared via in situ polymerization of diacrylate monomers in an orientated nematic liquid crystal (LC) matrix. The switching behavior of the LC molecules under electric field is probed in polarized Raman spectroscopy and straight theta-2straight theta elastic light scattering experiments. The electro-optical characteristics of the gels are directly related to the electric field dependence of the fraction of switched molecules. The electro-optical contrast relates to the coexistence of switched LC domains and LC domains anchored to the polymer network. PMID- 11102126 TI - Fast quantum search algorithms in protein sequence comparisons: quantum bioinformatics. AB - Quantum search algorithms are considered in the context of protein sequence comparison in bioinformatics. Given a sample protein sequence of length m (i.e., m residues), the problem considered is to find an optimal match in a large database containing N residues. Initially, Grover's quantum search algorithm is applied to a simple illustrative case-namely, where the database forms a complete set of states over the 2(m) basis states of a m qubit register, and thus is known to contain the exact sequence of interest. This example demonstrates explicitly the typical O(square root of [N]) speedup on the classical O(N) requirements. An algorithm is then presented for the (more realistic) case where the database may contain repeat sequences, and may not necessarily contain an exact match to the sample sequence. In terms of minimizing the Hamming distance between the sample sequence and the database subsequences the algorithm finds an optimal alignment, in O(square root of [N]) steps, by employing an extension of Grover's algorithm, due to Boyer et al. for the case when the number of matches is not a priori known. PMID- 11102127 TI - Adsorption-assisted translocation of a chain molecule through a pore AB - We analyze the free energy for translocation of a polymer from the outside of a spherical vesicle to the inside. The process is assumed to be driven by the adsorption of the polymer on the inner surface of the vesicle. We argue that in the case where the polymer is adsorbed on the outer surface too, the entropic barrier for translocation is absent. We analyze the adsorption process and find the free energy profile for the translocation. We argue that the motion corresponds to a polymer crossing a region with a change in free energy per segment. Based upon our earlier analysis of the behavior of kinks in such a problem, we conclude that the translocation can occur with a crossing time t(trans) approximately N. PMID- 11102128 TI - Compactlike breathers: bridging the continuous with the anticontinuous limit AB - We consider discrete nonlinear lattices characterized by on-site nonlinear potentials and nonlinear dispersive interactions that, in the continuous limit, support exact compacton solutions. We show that the compact support feature of the solutions in the continuous limit persists all the way to the anticontinuous limit. While in the large coupling regime the compact discrete breather solution retains the essential simple cosinelike compacton shape, in the close vicinity of the anticontinuous limit it acquires a spatial shape characterized by a fast stretched exponential decay, preserving thus its essentially compact nature. The discrete compact breathers in the anticontinuous limit are generated through a numerically exact procedure and are shown to be generally stable. PMID- 11102130 TI - Reply to "Comment on 'Experimental proof of standard electrodynamics by measuring the self-force on a part of a current loop' " AB - Our paper [Phys. Rev. E 58, 2505 (1998)] described experimental results that brought into agreement standard theory and experiment, in contrast to two previous experiments that claimed disagreement. The shape of our circuit was designed to improve knowledge of the electric flux lines. Such a shape required numerical calculations to predict the relevant force on the mobile part of the circuit. PMID- 11102129 TI - Comment on "Experimental proof of standard electrodynamics by measuring the self force on a part of a current loop" AB - We discuss the paper of Cavalleri et al. [Phys. Rev. E 58, 2505 (1998)] on the measurement of a force on part of a closed circuit carrying a constant current. PMID- 11102131 TI - Quantum-mechanical nonperturbative response of driven chaotic mesoscopic systems AB - Consider a time-dependent Hamiltonian H(Q,P;x(t)) with periodic driving x(t) = Asin(Omegat). It is assumed that the classical dynamics is chaotic, and that its power spectrum extends over some frequency range |omega|A(prt), where A(prt) approximately Planck's over 2pi, the system may have a relatively strong response for Omega>omega(cl) due to QM nonperturbative effect. PMID- 11102132 TI - Second order theory of excitations in trapped bose condensates at finite temperatures AB - We present a finite temperature field theory for collective excitations of trapped Bose condensates which includes the dynamics of the thermal cloud. In spherical traps we show that excitations couple strongly to a small number of modes, giving resonance structure in their frequency spectra. Where possible, we derive energy shifts and lifetimes of excitations. For the l = 0 mode we show that the simple picture of a decay rate fails, which should be observable in suitable experiments. It also suggests a possible explanation for the anomalous behavior of the m = 0 mode observed in anisotropic traps. PMID- 11102133 TI - Discretized diffusion processes AB - We study the properties of the "rigid Laplacian" operator; that is we consider solutions of the Laplacian equation in the presence of fixed truncation errors. The dynamics of convergence to the correct analytical solution displays the presence of a metastable set of numerical solutions, whose presence can be related to granularity. We provide some scaling analysis in order to determine the value of the exponents characterizing the process. We believe that this prototype model is also suitable to provide an explanation of the widespread presence of power law in a social and economic system where information and decision diffuse, with errors and delay from agent to agent. PMID- 11102134 TI - Continuous quantum measurement and the emergence of classical chaos AB - We formulate the conditions under which the dynamics of a continuously measured quantum system becomes indistinguishable from that of the corresponding classical system. In particular, we demonstrate that even in a classically chaotic system the quantum state vector conditioned by the measurement remains localized and, under these conditions, follows a trajectory characterized by the classical Lyapunov exponent. PMID- 11102135 TI - Measurement of direct photon emission in K+-->pi(+)pi(0)gamma decay AB - We have performed a measurement of the K+-->pi(+)pi(0)gamma decay and have observed 2x10(4) events. The best fit to the decay spectrum gives a branching ratio for direct photon emission of (4.7+/-0.8+/-0. 3)x10(-6) in the pi(+) kinetic energy region of 55 to 90 MeV and requires no component due to interference with inner bremsstrahlung. PMID- 11102136 TI - Search for direct CP violation in nonleptonic decays of charged Xi and lambda hyperons AB - A search for direct CP violation in the nonleptonic decays of hyperons has been performed. In comparing the product of the decay parameters, alpha(Xi)alpha(Lambda), in terms of an asymmetry parameter, A(XiLambda), between hyperons and antihyperons in the charged Xi-->Lambdapi and Lambda-->ppi decay sequence, we found no evidence of direct CP violation. The parameter A(XiLambda) was measured to be 0.012+/-0.014. PMID- 11102137 TI - Transverse momentum distributions and their forward-backward correlations in the percolating color string approach AB - The forward-backward correlations in the p(T) distributions at midrapidity, which present a clear signature of nonlinear effects in particle production, are studied in the model of percolating color strings. Quantitative predictions are given for these correlations at SPS, RHIC, and LHC energies. Interaction of strings also naturally explains the flattening of p(T) distributions and increase of with energy and atomic number for nuclear collisions. PMID- 11102138 TI - Semi-inclusive lambda and K(S) production in p-Au collisions at 17.5 GeV/c AB - The first detailed measurements of the centrality dependence of strangeness production in p-A collisions are presented. Lambda and K(S) dn/dy distributions from 17.5 GeV/ c p-Au collisions are shown as a function of "grey" track multiplicity and the estimated number of collisions, nu, made by the proton. The nu dependence of the Lambda yield deviates from a scaling of p-p data by the number of participants, increasing faster than this scaling for nu/=130 fs) at peak intensities of nl(') in atomic hydrogen induced by collisions with charged particles at ultralow energies. A novel classical expression for the transition probability P(l(')l) is presented. The exact classical results for P(l(')l)(alpha) as a function of l,l(') and the Stark parameter alpha agree exceptionally well with (exact) quantal results. They complement the quantal results by revealing essential characteristics which remain obscured in the quantal treatment. PMID- 11102142 TI - Ripple formation through an interface instability from moving growth and erosion sources AB - The propagation of material interfaces is investigated under the action of a localized moving source which deposits or removes material. Among others the latter process applies to beam cutting techniques. We develop a Kuramoto Sivashinsky-type model and find a new type of ripple forming mechanism. This theory offers a new explanation for the occurrence of striation patterns which often degrade the quality of cutting edges. PMID- 11102143 TI - Eliminating the transverse instabilities of kerr solitons AB - We show analytically, numerically, and experimentally that a transversely stable one-dimensional [(1+1)D] bright Kerr soliton can exist in a 3D bulk medium. The transverse instability of the soliton is completely eliminated if it is made sufficiently incoherent along the transverse dimension. We derive a criterion for the threshold of transverse instability that links the nonlinearity to the largest transverse correlation distance for which the 1D soliton is stable. PMID- 11102144 TI - Nondiffusive transport in tokamaks: three-dimensional structure of bursts and the role of zonal flows AB - Large scale transport events are studied in simulations of resistive ballooning turbulence in a tokamak plasma. The spatial structure of the turbulent flux is analyzed, indicating radially elongated structures (streamers) at the low field side which are distorted by magnetic shear at different toroidal positions. The interplay between self-generated zonal flows and transport events is investigated, resulting in significant modifications of the frequency and the amplitude of bursts. The propagation of bursts is studied in the presence of a transport barrier generated by a strong shear flow. PMID- 11102145 TI - Ba-IV-type incommensurate crystal structure in group-V metals AB - The long-unknown crystal structure of Bi-III has been solved. It comprises a body centered-tetragonal (bct) "host" and a bct "guest" component made up of chains that lie in channels in the host; the guest is incommensurate with the host along the tetragonal c axis. Diffraction data for Sb-II reveal that it too can be fitted with the same composite structure. The structures of these two high pressure phases of Bi and Sb are similar to those reported recently in the alkaline-earth metals Ba and Sr. PMID- 11102146 TI - Out-diffusion and precipitation of copper in silicon: An electrostatic model AB - Concentrations of mobile interstitial copper and precipitated copper in silicon were studied after a high temperature intentional contamination and quench to room temperature. It was found that below a critical contamination the copper predominantly diffuses out to the surface, while for higher initial copper concentrations it mainly precipitates in the bulk. The critical copper contamination equals the acceptor concentration plus 10(16) cm (-3). This behavior can be explained by the electrostatic interaction between the positively charged interstitial copper and the forming copper precipitates. PMID- 11102147 TI - Instabilities in diamond under high shear stress AB - We investigate, through first-principles calculations, lattice instabilities induced in diamond by the application of high shear stresses. For shear stresses as low as 95 GPa a lattice instability will occur, leading to graphitelike layered structures. This effect is highly anisotropic. The reversal of the direction of the applied shear forces may cause a change of 80 GPa in the shear stress value at which the instability develops. The same reversal also causes different bonds to be broken, resulting in a drastic change in the orientation of the resulting graphitelike structures. We also find that an additional compressive stress of 50 GPa along the (111) direction does not eliminate the shear-induced instability. PMID- 11102148 TI - Frost heave in argon AB - The freezing of argon in silica powder is observed to generate bands of pure solid argon in the same manner as in the phenomenon of ice lens formation in the freezing of moist ground. A first principles dynamical theory describes the mechanism of lens formation by the thermomolecular pressure-driven flow of interfacially melted films at the lens-solid boundary. PMID- 11102149 TI - Glass transition and relaxation following PhotoPerturbation in thin polymeric films AB - In this letter we employ null ellipsometry on Langmuir Blodgett multilayers of a glass forming photosensitive side chain polymer to investigate both the structural changes and the related time relaxation as a function of temperature and sample thickness following isothermal optical perturbation. Below a thickness of a few layers the glassy multilayer system collapses to a smecticlike crystal. This effect is discussed in the context of the glass transition in restricted dimensionality. PMID- 11102150 TI - Low-temperature structure of indium quantum chains on silicon AB - The array of quasi-one-dimensional indium chains in the Si(111)- (4x1)-In surface reconstruction exhibits a phase transition to a low-temperature (8x2) phase. It has been suggested that this phase transition is related to a charge density wave (CDW) formation. The x-ray diffraction results presented here demonstrate that at 20 K the CDW has not yet condensed into a superstructure even though good transverse coupling was established. This indicates that CDW formation cannot be the driving force for the phase transition. Furthermore we elucidate the subtle highly anisotropic interchain correlations and reveal the detailed atomic structure of the low-temperature (8x2) phase. PMID- 11102151 TI - Bloch electron in a magnetic field and the ising model AB - The spectral determinant det(H-varepsilonI) of the Azbel-Hofstadter Hamiltonian H is related to Onsager's partition function of the 2D Ising model for any value of magnetic flux Phi = 2piP/Q through an elementary cell, where P and Q are coprime integers. The band edges of H correspond to the critical temperature of the Ising model; the spectral determinant at these (and other points defined in a certain similar way) is independent of P. A connection of the mean of Lyapunov exponents to the asymptotic (large Q) bandwidth is indicated. PMID- 11102152 TI - New classes of quasicrystals and marginal critical states AB - One-dimensional quasilattices, namely, the geometrical objects that represent quasicrystals, are classified into mutual local-derivability (MLD) classes. Besides the familiar class, there exist an infinite number of new MLD classes, and different MLD classes are distinguished by the inflation rules of their representatives. It has been found that electronic properties of a new MLD class are characterized by the presence of marginal critical states, which are considered to be nearly localized states. PMID- 11102153 TI - Coulomb drag between one-dimensional conductors AB - We have analyzed Coulomb drag between currents of interacting electrons in two parallel one-dimensional conductors of finite length L attached to external reservoirs. For strong coupling, the relative fluctuations of electron density in the conductors acquire energy gap M. At energies larger than gamma = constxv( )exp(-LM/v(-))/L+gamma(+), where gamma(+) is the impurity scattering rate, and, for L>v(-)/M, where v(-) is the fluctuation velocity, the gap leads to an "ideal" drag with almost equal currents in the conductors. At low energies the drag is suppressed by coherent instanton tunneling, and the zero-temperature transconductance vanishes, indicating the Fermi-liquid behavior. PMID- 11102154 TI - Low-frequency crossover of the fractional power-Law conductivity in SrRuO3 AB - We combine the results of terahertz time-domain spectroscopy with far-infrared transmission and reflectivity to obtain the conductivity of SrRuO3 over an unprecedented continuous range in frequency, allowing us to characterize the approach to zero frequency as a function of temperature. We show that the conductivity follows a simple phenomenological form, with an analytic structure fundamentally different from that predicted by the standard theory of metals. PMID- 11102155 TI - Controlled modification of individual adsorbate electronic structure AB - Modification of the electronic structure of a single Mn adsorbate placed within a geometrical array of adatoms on Ag(111) is observed using local spectroscopy with the scanning tunneling microscope. The changes result from coupling between the adsorbate level and surface electronic states of the substrate. These surface states are scattered coherently within the adatom array, mediating the presence and shape of the array to the adsorbate within. The dimension and geometry of the adatom array thus provide a degree of control over the induced changes. PMID- 11102156 TI - Quantum phase transitions in d-wave superconductors AB - Motivated by the strong, low temperature damping of nodal quasiparticles observed in some cuprate superconductors, we study quantum phase transitions in d(x(2) y(2)) superconductors with a spin-singlet, zero momentum, fermion bilinear order parameter. We present a complete, group-theoretic classification of such transitions into seven distinct cases (including cases with nematic order) and analyze fluctuations by the renormalization group. We find that only two, the transitions to d(x(2)-y(2))+is and d(x(2)-y(2))+id(xy) pairing, possess stable fixed points with universal damping of nodal quasiparticles; the latter leaves the gapped quasiparticles along (1,0), (0,1) essentially undamped. PMID- 11102157 TI - Quasiparticle localization in disordered d-wave superconductors AB - An extensive numerical study is reported on the disorder effect in two dimensional d-wave superconductors with random impurities in the unitary limit. It is found that a sharp resonant peak shows up in the density of states at zero energy and correspondingly the finite-size spin conductance is strongly enhanced which results in a nonuniversal feature in one-parameter scaling. However, all quasiparticle states remain localized, indicating that the resonant density peak alone is not sufficient to induce delocalization. In the weak disorder limit, the localization length is so long that the spin conductance at small sample size is close to the universal value predicted by Lee [Phys. Rev. Lett. 71, 1887 (1993)]. PMID- 11102158 TI - Theory of plastic vortex creep AB - We develop a theory for plastic vortex creep in a topologically disordered (dislocated) vortex solid phase in type-II superconductors in terms of driven thermally activated dislocation dynamics. Plastic barriers for dislocations show a power-law divergence at small driving currents j, U(pl)( j) approximately j( &mgr;), with &mgr; = 1 for a single dislocation and &mgr; = 2/5 for creep of dislocation bundles. This implies a suppression of the creep rate at the transition from the ordered vortex phase ( &mgr; = 2/11) to the dislocated glass and can manifest itself as an observed increase of the apparent critical current (second peak). Our approach applies to general dynamics of disordered elastic media on a random substrate. PMID- 11102159 TI - Ultrahigh-resolution photoemission spectroscopy of Ni borocarbides: direct observation of the superconducting gap and a change in gap anisotropy by impurity AB - We have performed ultrahigh-resolution photoemission spectroscopy of Y(Ni1 xPtx)2B2C ( x = 0.0 and 0.2) in order to study the changes in the density of states across the superconducting transition. Because of a drastic increase in energy resolution, we clearly observe the opening of superconducting gaps across T(c) in both compounds. Furthermore, we find a small but significant difference in the superconducting-state spectral shape. This can be explained in terms of reduction in gap anisotropy by introducing impurities and provides spectroscopic evidence for an anisotropic s-wave gap in YNi2B2C. PMID- 11102160 TI - Exact and numerical results for a dimerized coupled spin- 1/2 chain AB - We establish exact results for coupled spin-1/2 chains for special values of the four-spin interaction V and dimerization parameter delta. The first exact result is at delta = 1/2 and V = -2. Because we find a very small but finite gap in this dimerized chain, this can serve as a very strong test case for numerical and approximate analytical techniques. The second result is for the homogeneous chain with V = -4 and gives evidence that the system has a spontaneously dimerized ground state. Numerical diagonalization and bosonization techniques indicate that the interplay between dimerization and interaction could result in gapless phases in the regime 0/=2. We show that the main effect of increasing the order of the interactions among the species is to make the system less competitive, in the sense that the fraction of extinct species is greatly reduced. In addition, we find that for p>2 there is a threshold value which gives a lower bound to the concentration of the surviving species, preventing then the existence of rare species and, consequently, increasing the robustness of the ecosystem to external perturbations. PMID- 11102169 TI - Absence of dc-conductivity in lambda-DNA. AB - The electrical conductivity of biomaterials on a molecular scale is of fundamental interest in the life sciences. We perform first principles electronic structure calculations, which clearly indicate that lambda-DNA chains should present large resistance values. We also present two direct procedures to measure electrical currents through DNA molecules adsorbed on mica. The lower limit for the resistivity is 10(6) Omega . cm, in agreement with our calculations. We also show that low energy electron bombardment induces a rapid contamination and dramatically affects the measured conductivity, thus providing an explanation to recent reports of high DNA conductivity. PMID- 11102168 TI - Why is the DNA denaturation transition first order? AB - We study a model for the denaturation transition of DNA in which the molecules are considered as being composed of a sequence of alternating bound segments and denaturated loops. We take into account the excluded-volume interactions between denaturated loops and the rest of the chain by exploiting recent results on scaling properties of polymer networks of arbitrary topology. The phase transition is found to be first order in d = 2 dimensions and above, in agreement with experiments and at variance with previous theoretical results, in which only excluded-volume interactions within denaturated loops were taken into account. Our results agree with recent numerical simulations. PMID- 11102170 TI - Instabilities due to charge-density-curvature coupling in charged membranes. AB - Instabilities are caused by the reduction in the electrostatic energy when the membrane is curved with the higher charge density on the bilayer which is stretched by the curvature. In a bilayer where the charges can flip from one lipid layer to the other, there is a thermodynamic instability to a spontaneously curved state with different charge densities on the two sides. If the charges are not permitted to flip, there is a dynamical instability due to the correlated modulation of the charge density and curvature fields. Numerical estimates show that these effects are present in parameter regimes relevant to biological systems. PMID- 11102171 TI - Frustrations in polymer conformation in gels and their minimization through molecular imprinting. AB - We report an experimental realization of a gel system in which frustrations exist and can be minimized, thus meeting two crucial criteria predicted to enable memory of conformations in polymers. The gels consist of a thermosensitive major monomer component and two minor components. One minor component is positively charged and will form complexes around negatively charged target molecules placed in solution. The complexes can be imprinted into the gel by then cross-linking the second minor component, which will form cross-links additional to those in the major polymer matrix. The complexes are destroyed and reformed upon swelling and reshrinking of the gels, showing that memorization has been achieved. PMID- 11102172 TI - Low cloud properties influenced by cosmic rays AB - The influence of solar variability on climate is currently uncertain. Recent observations have indicated a possible mechanism via the influence of solar modulated cosmic rays on global cloud cover. Surprisingly the influence of solar variability is strongest in low clouds (nu(&mgr;) and nu;(&mgr;)-->nu;(&mgr;) survival probabilities at a baseline L = 732 km can test for CPT-odd contributions at orders of magnitude better sensitivity than present neutrino sector limits. Interference between the CPT-violating interaction and CPT-even mass terms in the Lagrangian can lead to a resonant enhancement of the oscillation amplitude. For oscillations in matter, a simultaneous enhancement of both neutrino and antineutrino oscillation amplitudes is possible. PMID- 11102186 TI - First direct measurement of the parity-violating coupling of the Z0 to the s quark AB - We present the first direct measurement of A(s), the parity-violating coupling of the Z0 boson to the strange quark, using approximately 550 000 e(+)e(-)-->Z0- >hadrons events recorded by the SLC Large Detector with a polarized e(-) beam. We tagged Z0-->s&smacr; events by the absence of B or D hadrons and the presence in each hemisphere of a high momentum K+/- or K(0)(s). Fitting the polar angle distributions of the strangeness-signed thrust axis gave A(s) = 0.895+/ 0.066(stat)+/-0.062(syst). The analyzing power and uu+d&dmacr; background were constrained using the data, greatly reducing any model dependence. PMID- 11102187 TI - Enhanced electric dipole moment of the muon in the presence of large neutrino mixing AB - The electric dipole moment (edm) of the muon ( d(e)(&mgr;)) is evaluated in supersymmetric models with nonzero neutrino masses and large neutrino mixing arising from the seesaw mechanism. It is found that if the seesaw mechanism is embedded in the framework of a left-right symmetric gauge structure, the interactions responsible for the right-handed neutrino Majorana masses lead to an enhancement in d(e)(&mgr;) to values as large as 5x10(-23)e cm, with a correlated value of (g-2)(&mgr;) approximately 13x10(-10). This should provide a strong motivation for improving the edm of the muon to the level of 10(-24)e cm as has recently been proposed. PMID- 11102188 TI - Cross section for b-Jet production in &pmacr;p collisions at radicals = 1.8 TeV AB - Bottom-quark production in &pmacr;p collisions at sqrt[s] = 1.8 TeV is studied with 5 pb(-1) of data collected in 1995 by the D0 detector at the Fermilab Tevatron Collider. The differential production cross section for b jets in the central rapidity region ( | y(b)|<1) as a function of jet transverse energy is extracted from a muon-tagged jet sample. Within experimental and theoretical uncertainties, D0 results are found to be higher than, but compatible with, next to-leading-order QCD predictions. PMID- 11102189 TI - Sudden interchannel interaction in the Tl 6p ionization above the 5d threshold AB - The linear magnetic dichroism in the angular distribution of Tl 5d and 6p photoelectrons and their dynamical spin polarization have been measured between hnu = 30 and 50 eV. In contrast to the Xe 5p photoionization at the 4d threshold, our results show that above the Tl 5d threshold strong interchannel coupling effects induce a sudden increase in the asymptotic phase difference of the s and d waves for the Tl 6p ionization. This shows that the valence excitation is different for resonant (Xe 4d) and nonresonant (Tl 5d) excitation from subvalence shells. PMID- 11102190 TI - Observation of inner electron ionization from radial rydberg wave packets in two electron atoms AB - We have observed multiphoton ionization of the 5s core electron from a 5snd radial Rydberg wave packet of Sr atoms using a short optical pulse. When the outer nd electron is at its outer turning point the inner 5s electron is removed from the atom, and the outer electron is left in a Sr+ Rydberg state, but when the outer electron is at the inner turning point this does not occur. Analysis of the final Sr+ Rydberg states shows that the two electrons interact as the inner electron leaves, so that the outer electron is not simply projected onto the Sr+ Rydberg states. PMID- 11102191 TI - High harmonic generation beyond the electric dipole approximation AB - A generalization of the analytical theory of high harmonic generation in the long wavelength limit and in the single active electron approximation is developed taking into account the magnetic dipole and electric quadrupole interaction. Quantum mechanical and classical theories are found to be in excellent agreement, which allows one to explain the influence of multipole effects in terms of an intuitive picture. For Ti:S lasers ( 0.8 &mgr;m) multipole contributions are found to be small below an intensity of about 10(17) W/cm(2), at which harmonic radiation with photon energies of several keV is generated. This promises the extension of high harmonic generation well into the sub-nm wavelength regime. PMID- 11102192 TI - Boundary-free propagation with the time-dependent Schrodinger equation AB - We present two methods that allow for the efficient numerical propagation of continuum wave packets to large times. Time-dependent solutions of the Schrodinger equation that include continuum components are numerically challenging to solve because the wave packet travels, spreads, and acquires a spatial phase gradient. The methods we propose account for these kinematic effects analytically in general and numerically tractable schemes. PMID- 11102193 TI - X-Ray emission following low-energy charge exchange collisions of highly charged ions AB - K-shell x-ray emission following low-energy charge exchange collisions ( W(D)/(1+W(D)) leads to a stationary state, while in the opposite case, the broken resistor fraction reaches the percolation threshold p(c). We study the resistance noise of this system under stationary conditions by Monte Carlo simulations. The variance of resistance fluctuations is found to follow a scaling law |p-p(c)|( kappa(0)) with kappa(0) = 5.5. The proposed model relates quantitatively the defectiveness of a disordered media with its electrical and excess-noise characteristics. PMID- 11102231 TI - Multiple-sequence information provides protection against mis-specified potential energy functions in the lattice model of proteins AB - The neutral nets in the lattice models of proteins are composed of a group of similar sequences that encode for the same native structure. The maximal neutral nets in an HP and an AB lattice model are investigated by exhaustive enumeration in the conformation space. The result demonstrates that the performance of mis specified potential functions can be improved significantly by averaging over sequences in the neutral nets. PMID- 11102232 TI - Elasticity of the rod-shaped gram-negative eubacteria. AB - We report a theoretical calculation of the elasticity of the peptidoglycan network, the only stress-bearing part of rod-shaped Gram-negative eubacteria. The peptidoglycan network consists of elastic peptides and inextensible glycan strands, and it has been proposed that the latter form zigzag filaments along the circumference of the cylindrical bacterial shell. The zigzag geometry of the glycan strands gives rise to nonlinear elastic behavior. The four elastic moduli of the peptidoglycan network depend on its stressed state. For a bacterium under physiological conditions the elasticity is proportional to the bacterial turgor pressure. Our results are in good agreement with recent measurements. PMID- 11102233 TI - Do thiols merely passivate gold nanoclusters? PMID- 11102234 TI - Comment on "Relativistic effects of light in moving media with extremely low group velocity" PMID- 11102235 TI - Leonhardt and piwnicki reply: PMID- 11102236 TI - Comment on "Canonical and microcanonical calculations for fermi systems" PMID- 11102237 TI - Comment on "Potts model with long-range interactions in one dimension" PMID- 11102238 TI - Bayong et al. reply: PMID- 11102239 TI - Andreev reflection at point contacts with heavy-fermion UBe13? PMID- 11102240 TI - Walti et al. reply PMID- 11102242 TI - Bellessa et al. reply: PMID- 11102241 TI - Spin tunneling in Mn12 cannot Be assisted by phonons at millikelvin temperatures PMID- 11102243 TI - Hypertrophic cardiomyopathy and sudden death: new perspectives on risk stratification and prevention with the implantable cardioverter-defibrillator. PMID- 11102244 TI - Hotline sessions at the 22nd European Congress of Cardiology. PMID- 11102245 TI - Non-Q wave myocardial infarction management strategies. PMID- 11102246 TI - How hot is inflammation in acute coronary syndrome? PMID- 11102247 TI - A cold start for oral glycoprotein IIb/IIIa antagonists. PMID- 11102248 TI - Brachytherapy in the Journal: European cardiologists have their own forum and should use it! PMID- 11102249 TI - In TIME-II: more than a study on lanoteplase. PMID- 11102250 TI - The utilization of the implantable defibrillator--a European enigma. PMID- 11102251 TI - Intravenous NPA for the treatment of infarcting myocardium early; InTIME-II, a double-blind comparison of single-bolus lanoteplase vs accelerated alteplase for the treatment of patients with acute myocardial infarction. AB - AIMS: To compare the efficacy and safety of lanoteplase, a single-bolus thrombolytic drug derived from alteplase tissue plasminogen activator, with the established accelerated alteplase regimen in patients presenting within 6 h of onset of ST elevation acute myocardial infarction. METHODS AND RESULTS: 15,078 patients were recruited from 855 hospitals worldwide and randomized in a 2:1 ratio to receive either lanoteplase 120 KU. kg(-1)as a single intravenous bolus, or up to 100 mg accelerated alteplase given over 90 min. The primary end-point was all-cause mortality at 30 days and the hypothesis was that the two treatments would be equivalent. By 30 days, 6.61% of alteplase-treated patients and 6.75% lanoteplase-treated patients had died (relative risk 1.02). Total stroke occurred in 1.53% alteplase- and 1.87% lanoteplase-treated patients (ns); haemorrhagic stroke rates were 0.64% alteplase and 1.12% lanoteplase (P=0.004). The net clinical deficit of 30-day death or non-fatal disabling stroke was 7.0% and 7.2%, respectively. By 6 months, 8.8% of alteplase-treated patients and 8.7% of lanoteplase-treated patients had died. CONCLUSION: Single-bolus weight-adjusted lanoteplase is an effective thrombolytic agent, equivalent to alteplase in terms of its impact on survival and with a comparable risk-benefit profile. The single bolus regimen should shorten symptoms to treatment times and be especially convenient for emergency department or out-of-hospital administration. PMID- 11102252 TI - Early invasive versus ischaemia-guided strategies in the management of non-Q wave myocardial infarction patients with and without prior myocardial infarction; results of Veterans Affairs Non-Q Wave Infarction Strategies in Hospital (VANQWISH) trial. AB - AIMS: To compare the role of early invasive vs conservative management strategies in treating patients with non-Q wave myocardial infarction with or without a prior myocardial infarction. BACKGROUND: In patients recovering from non-Q wave myocardial infarction, the prognosis among patients with a first non-Q wave myocardial infarction is significantly better than in patients with a prior myocardial infarction, yet physicians often adopt an early invasive strategy to treat patients with a first non-Q wave myocardial infarction. METHODS: Non-Q wave myocardial infarction patients enrolled in the VANQWISH trial with a history of prior myocardial infarction were compared to those with a first non-Q wave myocardial infarction, for the trial primary end-point of death or myocardial infarction at 1 and 12 months, as well as for the initial randomized treatment strategy. RESULTS: Of the 920 non-Q wave myocardial infarction patients, 396 had a history of prior myocardial infarction and 524 did not. Patients with a history of prior myocardial infarction were older and had a higher incidence of multiple high-risk baseline characteristics than those with a first non-Q wave myocardial infarction. Compared to the group with a first myocardial infarction, the prior myocardial infarction group suffered more events at both 1 month (11% vs 6%, P=0.007) and at 12 months (29% vs 16%, P<0.001). This difference in outcome remained significant even after adjusting for confounding variables (P<0.0001 at 12 months). Among the non-Q wave myocardial infarction patients with a prior myocardial infarction, the frequency of death or recurrent myocardial infarction was similar in both invasive and conservative groups during the first year of follow-up. Among the first non-Q wave myocardial infarction group, those assigned to the conservative strategy had significantly fewer events (3% vs 9%, P=0.009 at 1 month; 12% vs 20%, P=0.016 at 12 months) and mortality (1% vs 5%, P=0.012 at one month; 5% vs 11%, P=0.009 at 12 months) than those assigned to early invasive strategy. CONCLUSION: A history of prior myocardial infarction identifies a moderately high-risk subset of non-Q wave myocardial infarction patients who display similar long-term outcomes regardless of the strategy assignment; however, patients with a first non-Q wave myocardial infarction may fare better with a conservative or ischaemia-guided approach during the first post infarction year. PMID- 11102253 TI - A randomized placebo-controlled trial to assess the efficacy of antiinflammatory therapy with methylprednisolone in unstable angina (MUNA trial). AB - AIMS: The purpose of this study was to assess the efficacy of antiinflammatory therapy with methylprednisolone during the acute phase of unstable angina. METHODS: This is a randomized 'prospective' double-blind, placebo-controlled trial. Patients with the diagnosis of unstable angina were randomized to a 48-h course of methylprednisolone (n=81) or placebo (n=85). Patient care and therapy were otherwise decided by their attending cardiologist. The primary end-point was a composite of in-hospital recurrence of angina, silent ischaemia on Holter recording, emergency coronary revascularization, readmission with unstable angina, and myocardial infarction or death during the 30-day follow-up. RESULTS: The two groups were well balanced and had similar clinical characteristics at baseline. Forty-eight hours after randomization, mean C-reactive protein levels decreased by 2.6 mg. l(-1)in the methylprednisolone group, but increased by 1.6 mg. l(-1)in the placebo group (P=0.03). The primary end-point occurred in 44% of the methylprednisolone patients and in 33% of the placebo patients (P=0.12). Coronary revascularization rates were equal between the two groups (38% and 40%). When adjustment was made for the difference in revascularization times, a trend towards better event-free survival was seen in the control group (67% vs 57%;P=0.09). CONCLUSION: A 48 h course of antiinflammatory therapy with methylprednisolone given at the doses of this study did not improve the short term outcome of patients with unstable angina. PMID- 11102254 TI - The Clopidogrel in Unstable angina to prevent Recurrent Events (CURE) trial programme; rationale, design and baseline characteristics including a meta analysis of the effects of thienopyridines in vascular disease. AB - BACKGROUND: Other than aspirin, there are few oral antithrombotic treatments with proven efficacy in patients with acute coronary syndrome. In this report, we present the rationale, design and baseline characteristics of the Clopidogrel in Unstable angina to prevent Recurrent ischaemic Events (CURE) trial, which includes a meta-analysis of the effects of thienopyridines in patients with vascular disease. METHODS AND RESULTS: Combined data from randomized trials of thienopyrindines in patients with atherosclerotic disease demonstrated a 29% reduction in vascular events when compared with placebo/control (n=2392) (OR 0.71, 95% CI 0.58-0.86, P=0.0006) and a 10% reduction in vascular events when compared with aspirin (n=22 254) (OR 0.91, 95% Cl 0.84-0.99, P=0.039). Similarly, randomized trials of aspirin plus thienopyridines in patients undergoing intracoronary stenting, demonstrated a marked benefit of aspirin plus ticlopidine in reducing death or myocardial infarction compared with aspirin alone (OR 0.23, 95% CI 0.11-0.49, P=0.0001) or aspirin plus warfarin (OR 0.51, 95% CI 0.33-0.78, P=0.002). Whether these benefits extend to the much larger population of patients with acute coronary syndrome is unknown. CURE is an international, randomized, double-blind trial, in which patients with acute coronary syndrome will be randomized to receive either a bolus dose of clopidogrel (300 mg) followed by 75 mg per day for 3-12 months, or matching placebo. Both groups will receive aspirin. The co-primary efficacy end-points of CURE are: (1) the composite of cardiovascular death, myocardial infarction or stroke; and (2) the composite of cardiovascular death, myocardial infarction, stroke or refractory ischaemia. CURE will recruit approximately 12 500 patients with acute coronary syndrome (from 28 countries) and its power to detect moderate treatment benefits will be in the region of 80-90%, while maintaining an overall type I error (alpha) of 0.05. The baseline characteristics of the study population are consistent with at least a moderate risk group of patients with acute coronary syndrome. CONCLUSIONS: Randomized trials of thienopyridines in patients with vascular disease demonstrate that thienopyridines are effective in reducing vascular events when compared with placebo/control or aspirin, as well as when used in combination with aspirin in patients undergoing intracoronary stent implantation. The CURE trial is a large international study to determine if acute and long-term treatment with the combination of clopidogrel and aspirin is superior to aspirin alone in patients with acute coronary syndrome. PMID- 11102255 TI - Safety and preliminary efficacy of one month glycoprotein IIb/IIIa inhibition with lefradafiban in patients with acute coronary syndromes without ST-elevation; a phase II study. AB - AIMS: Oral glycoprotein IIb/IIIa inhibitors might enhance the early benefit of an intravenous agent and prevent subsequent cardiac events in patients with acute coronary syndromes. We assessed the safety and preliminary efficacy of 1 month treatment with three dose levels of the oral GP IIb/IIIa blocker lefradafiban in patients with unstable angina or myocardial infarction without persistent ST elevation. METHODS: The Fibrinogen Receptor Occupancy STudy (FROST) was designed as a dose-escalation trial with 20, 30 and 45 mg lefradafiban t.i.d. or placebo. Five hundred and thirty-one patients were randomized in a 3:1 ratio to lefradafiban or placebo in a double-blind manner. Efficacy was assessed by the incidence of death, myocardial infarction, coronary revascularization and recurrent angina. Safety was evaluated by the occurrence of bleeding classified according to the TIMI criteria and by measuring clinical laboratory parameters. RESULTS: There was a trend towards a reduction in cardiac events with lefradafiban 30 mg when compared with placebo and lefradafiban 20 mg. The benefit was particularly apparent in patients with a positive (> or = O.1 ng. ml(-1)) troponin I test at baseline and less so in those with a negative test result. In patients receiving lefradafiban, the cardiac event rate decreased with increasing minimal levels of fibrinogen receptor occupancy. There was a dose-dependent increase in the incidence of bleeding: the composite of major or minor bleeding occurred in 1% of placebo patients, 5% of patients receiving lefradafiban 20 mg and in 7% of patients receiving 30 mg, with an excessive risk (15%) in the 45 mg group which resulted in early discontinuation of this dose level. Gingival and arterial or venous puncture site bleedings were most common and accounted for more than 60% of all haemorrhagic events. There was an increased incidence of neutropenia (neutrophils <1. 5 x 10(9)/l) in the lefradafiban groups (5.2% vs 1.5% in the placebo group), which did not result from bone marrow depression but rather from a reversible redistribution of neutrophils by margination or clustering. CONCLUSION: One month's treatment with the oral glycoprotein IIb/IIIa inhibitor lefradafiban in patients with unstable angina and myocardial infarction without persistent ST elevation resulted in a decrease in cardiac events with lefradafiban 30 mg and a dose-dependent increase in haemorrhagic events. The observed favourable trend towards a reduction in cardiac events in patients with elevated troponin levels requires confirmation in a large clinical trial. PMID- 11102256 TI - A novel balloon angioplasty catheter impregnated with beta-particle emitting radioisotopes for vascular brachytherapy to prevent restenosis; first in vivo results. AB - BACKGROUND: According to early clinical trials, vascular brachytherapy performed prior to or shortly after angioplasty is very effective in reducing restenosis rates. The purpose of this study was to investigate the effects of a novel radioactive catheter that allows simultaneous balloon angioplasty and beta particle irradiation in the prevention of restenosis. MATERIAL AND METHODS: The balloon surface of an angioplasty catheter was impregnated with the radioisotope(32)P. Dosimetry calculations using a Monte Carlo method were performed at a radial distance of 0.2 mm from the balloon surface. Rabbit iliac arteries were dilated and simultaneously irradiated with a dose of 20 Gy delivered to the adventitia. Control arteries were only dilated and not irradiated. Neointimal areas, cell numbers and the perimeter of the arteries were measured by histomorphometry after 6 weeks. RESULTS: Neointima formation was reduced after balloon dilatation and simultaneous beta-particle irradiation using the(32)P impregnated angioplasty catheter as compared to balloon dilatation alone with a non-impregnated catheter (0.09+/-0.06 vs 0.27+/-0.09 mm(2)neointimal area and 168+/-45 vs 360+/-133 cells/0.05 mm(2)neointima, P<0.001 vs control, respectively). In addition, balloon dilatation with the(32)P impregnated angioplasty catheter increased the vessel perimeter as compared to balloon dilatation with a non-impregnated catheter (4. 7+/-0.2 vs 3.9+/-0.3 mm, P<0.001 vs control). CONCLUSIONS: Simultaneous balloon dilatation and vascular brachytherapy with a novel(32)P impregnated angioplasty catheter markedly reduces restenosis in vivo by preventing neointimal hyperplasia and constrictive vascular remodelling. PMID- 11102257 TI - Relationship between tensile stress and plaque growth after balloon angioplasty treated with and without intracoronary beta-brachytherapy. AB - AIMS: We investigated the influence of tensile stress on plaque growth after balloon angioplasty with and without beta-radiation therapy. METHODS AND RESULTS: Thirty-one consecutive patients successfully treated with balloon angioplasty were analysed qualitatively and quantitatively by means of an ECG-gated three dimensional intravascular ultrasound post-procedure and at follow-up. Eighteen patients were irradiated with catheter-based beta-radiation ((90)Sr/(90)Y source) and 13 were not (control). Studied segments were divided into 2 mm subsegments. Thus 184 irradiated and 111 non-irradiated subsegments were included. Tensile stress was calculated according to Laplace's law. The radiation dose was calculated by means of dose-volume histograms. Plaque growth was positively correlated to tensile stress in both the radiation and control groups (r=0.374, P=0.0001 and r=0.305, P=0.001). Low-dose subsegments (<6 Gy) had a significant correlation (r=0.410, P=0.0001) whereas no correlation was observed in the effective-dose subsegments (> or = 6 Gy). Multivariate analysis identified tensile stress as the only independent predictor of plaque increase in non irradiated subsegments, whereas actual dose and plaque morphology were stronger predictors in irradiated subsegments. CONCLUSION: The results of this study suggest that plaque growth is related to tensile stress after balloon angioplasty. Intracoronary brachytherapy may alter the biophysical process on plaque growth when the prescribed dose is effectively delivered. PMID- 11102258 TI - Meta-analysis of the implantable cardioverter defibrillator secondary prevention trials. AVID, CASH and CIDS studies. Antiarrhythmics vs Implantable Defibrillator study. Cardiac Arrest Study Hamburg . Canadian Implantable Defibrillator Study. AB - AIMS: Three randomized trials of implantable cardioverter defibrillator (ICD) therapy vs medical treatment for the prevention of death in survivors of ventricular fibrillation or sustained ventricular tachycardia have been reported with what might appear to be different results. The present analysis was performed to obtain the most precise estimate of the efficacy of the ICD, compared to amiodarone, for prolonging survival in patients with malignant ventricular arrhythmia. METHODS AND RESULTS: Individual patient data from the Antiarrhythmics vs Implantable Defibrillator (AVID) study, the Cardiac Arrest Study Hamburg (CASH) and the Canadian Implantable Defibrillator Study (CIDS) were merged into a master database according to a pre-specified protocol. Proportional hazard modelling of individual patient data was used to estimate hazard ratios and to investigate subgroup interactions. Fixed effect meta-analysis techniques were also used to evaluate treatment effects and to assess heterogeneity across studies. The classic fixed effects meta-analysis showed that the estimates of ICD benefit from the three studies were consistent with each other (P heterogeneity=0.306). It also showed a significant reduction in death from any cause with the ICD; with a summary hazard ratio (ICD:amiodarone) of 0.72 (95% confidence interval 0.60, 0.87;P=0.0006). For the outcome of arrhythmic death, the hazard ratio was 0.50 (95% confidence interval 0.37, 0.67;P<0.0001). Survival was extended by a mean of 4.4 months by the ICD over a follow-up period of 6 years. Patients with left ventricular ejection fraction < or = 35% derived significantly more benefit from ICD therapy than those with better preserved left ventricular function. Patients treated before the availability of non-thoracotomy ICD implants derived significantly less benefit from ICD therapy than those treated in the non-thoracotomy era. CONCLUSION: Results from the three trials of the ICD vs amiodarone are consistent with each other. There is a 28% reduction in the relative risk of death with the ICD that is due almost entirely to a 50% reduction in arrhythmic death. PMID- 11102259 TI - Clinical outcome after coronary events in patients treated with HIV-protease inhibitors. PMID- 11102260 TI - Clinical trials for conversion of recent onset atrial fibrillation must consider the role of digoxin. PMID- 11102261 TI - Cavotricuspid isthmus ablation and atrial fibrillation. PMID- 11102264 TI - Volume 21 Contents and Index. PMID- 11102262 TI - Decreased heart rate variability as a predictor for sudden death was known in China in the third century A.D. PMID- 11102266 TI - The revolution that wasn't: a new interpretation of the origin of modern human behavior. AB - Proponents of the model known as the "human revolution" claim that modern human behaviors arose suddenly, and nearly simultaneously, throughout the Old World ca. 40-50 ka. This fundamental behavioral shift is purported to signal a cognitive advance, a possible reorganization of the brain, and the origin of language. Because the earliest modern human fossils, Homo sapiens sensu stricto, are found in Africa and the adjacent region of the Levant at >100 ka, the "human revolution" model creates a time lag between the appearance of anatomical modernity and perceived behavioral modernity, and creates the impression that the earliest modern Africans were behaviorally primitive. This view of events stems from a profound Eurocentric bias and a failure to appreciate the depth and breadth of the African archaeological record. In fact, many of the components of the "human revolution" claimed to appear at 40-50 ka are found in the African Middle Stone Age tens of thousands of years earlier. These features include blade and microlithic technology, bone tools, increased geographic range, specialized hunting, the use of aquatic resources, long distance trade, systematic processing and use of pigment, and art and decoration. These items do not occur suddenly together as predicted by the "human revolution" model, but at sites that are widely separated in space and time. This suggests a gradual assembling of the package of modern human behaviors in Africa, and its later export to other regions of the Old World. The African Middle and early Late Pleistocene hominid fossil record is fairly continuous and in it can be recognized a number of probably distinct species that provide plausible ancestors for H. sapiens. The appearance of Middle Stone Age technology and the first signs of modern behavior coincide with the appearance of fossils that have been attributed to H. helmei, suggesting the behavior of H. helmei is distinct from that of earlier hominid species and quite similar to that of modern people. If on anatomical and behavioral grounds H. helmei is sunk into H. sapiens, the origin of our species is linked with the appearance of Middle Stone Age technology at 250-300 ka. PMID- 11102267 TI - The hunters and the hunted revisited. AB - The dietary niches of extinct animals, including hominids and predators, may be constrained using stable carbon isotope ratios in fossil tooth enamel.(13)C/(12)C ratios of many of the primates abundant in the faunal assemblages of Members 1 and 2 at Swartkrans, including cercopithecoids and Australopithecus (Paranthropus) robustus, and a range of other possible prey species, have been reported previously. Resulting suggestions of a mixed, or omnivorous, diet for A. robustus raise questions about niche overlap with coeval, larger brained Homo. Here we present(13)C/(12)C data from Homo and several large predators including Panthera pardus, Dinofelis sp., Megantereon cultridens and Chasmoporthetes nitidula in Member 1, and P. pardus and P. leo in Member 2, in order to compare the two hominid species and to determine likely predators of the various primates and other macrovertebrates. Results for three Homo cf. ergaster individuals are indistinguishable from those of A. robustus, showing that proportions of C(3)- and C(4)-based foods in their diets did not differ. P. pardus, Megantereon and Crocuta are shown to be likely predators of the hominids and Papio baboons in Member 1, while the Dinofelis individual concentrated on prey which consumed C(4)grasses. The hunting hyaenid C. nitidula preyed on either mixed feeders or on a range of animals across the spectrum of C(3)and C(4)variation. The data from Members 1 and 2 confirm a shift in leopard diets towards animals that consumed C(4)grasses. PMID- 11102268 TI - Geochronology of Oreopithecus -bearing succession at Baccinello (Italy) and the extinction pattern of European Miocene hominoids. PMID- 11102269 TI - The shoulders follow the head: postcranial constraints on human childbirth. PMID- 11102270 TI - Sex and significance of Lake Mungo 3: reply to Brown "Australian pleistocene variation and the sex of Lake Mungo 3". PMID- 11102272 TI - Contents and index of volume 39 PMID- 11102271 TI - Tabun revisited: revised ESR chronology and new ESR and U-series analyses of dental material from Tabun C1. PMID- 11102273 TI - Blood groups and their function. AB - The function(s) assigned to red blood cell membrane components is based on an observed effect in the red cells that lack the component, comparison of the protein sequence (predicted from the nucleotide sequence of the gene) to proteins of known function, and extrapolation of function of the component in other cells. The functions are varied and include membrane structure, transport, receptor, adhesion, enzyme activity, complement components, complement regulation and glycocalyx formation. Several components have more than one function. PMID- 11102274 TI - The HLA system in blood transfusion. AB - The human leukocyte antigen (HLA) system, originally discovered as the result of a transfusion reaction, is now known to play a crucial role in many areas of clinical medicine. The main function of the HLA molecules is to present antigenic peptides to the immune system and in this way regulate the induction of immune responses. This is a highly regulated process which requires a close interaction between the HLA molecules, the antigenic peptide and the T cell receptor.HLA molecules are also known to be associated with a variety of autoimmune, non autoimmune and infectious diseases and to restrict the antibody response to certain antigens and to vaccines. It is likely that the mechanism responsible for this restriction is the preferential presentation of antigen-derived peptides to T cells. Furthermore, HLA antigens, in contrast to most polymorphic molecules, have the ability to activate the immune system using two different pathways of T cell activation, the direct and indirect pathways. As a result of these features, HLA antigens and antibodies are responsible for some of the serious clinical complications of blood transfusion, and have an important influence on the outcome of solid organ and haemopoietic stem cell transplantation. PMID- 11102275 TI - Autologous transfusion and other approaches to reduce allogeneic blood exposure. AB - When used as the sole source of transfused blood, the principal advantage of autologous blood transfusion is the avoidance of transmission of infectious agents and the avoidance of the purported adverse immunomodulatory effects of allogeneic transfusion. In the 1990s, however, the risks of transfusion transmitted diseases have been greatly reduced, and estimates of the cost effectiveness of pre-operative autologous blood donations now vary between 2470 dollars and 3,400,000 dollars per quality-adjusted year of life saved, depending on assumptions about the existence and magnitude of any adverse immunomodulatory effects of allogeneic transfusion. There is a paucity of randomized controlled trials evaluating the clinical outcomes and the cost-effectiveness of autologous transfusion procedures, and this situation is unlikely to change in the near future because of the difficulties in conducting such trials. This chapter reviews the available evidence on the efficacy, safety and cost-effectiveness of the three common autologous transfusion procedures, that is, pre-operative autologous blood donation, acute normovolaemic haemodilution, and intra-operative and post-operative blood recovery. PMID- 11102276 TI - New technologies for the inactivation of infectious pathogens in cellular blood components and the development of platelet substitutes. AB - Despite the increased safety of blood components, achieved through improved donor selection and testing, transfusion recipients remain at risk of transfusion associated diseases. Transfusion of cellular blood components has been implicated in transmission of viral, bacterial and protozoan diseases. While it is commonly recognized that hepatitis B virus (HBV), hepatitis C virus (HCV), cytomegalovirus (CMV), and retroviruses, such as human immunodeficiency virus (HIV) and the human lymphotrophic viruses (HTLV), can be transmitted through cellular components, other pathogens are emerging as potentially significant transfusion-associated infectious agents. For example, transmission of protozoan infections due to trypanosomes and Babesia have been reported. In addition to viral and protozoan infectious agents, cases of bacterial contamination of platelet and red cell concentrates continue to be reported and may be an under-reported transfusion complication. More importantly, new infectious agents continue to enter the donor population, and there is an inherent time delay before the new pathogens are definitively identified and new tests implemented in order to maintain consistent safety of the blood supply. The paradigm for this problem is the HIV pandemic. During the past decade a number of methods for inactivating infectious pathogens in platelet concentrates have been investigated as a strategy to improve the safety of platelet transfusion therapy. One method of treating platelet concentrates to inactive pathogens has now reached the advanced clinical trial phase in the United States and Europe. Similar efforts with a new class of compounds are underway for red cell concentrates, and two of these are in early phase trials. In addition to studies with allogeneic platelets and red cells, a number of laboratories have described methods for developing platelet substitutes or modified platelets to avoid the use of traditional platelet concentrates as a means to improve safety. PMID- 11102277 TI - Placental blood for bone marrow replacement: the New York Blood Center's program and clinical results. AB - BACKGROUND AND OBJECTIVES: Placental/umbilical cord blood (PCB) has been used for allogeneic bone marrow replacement since 1988. The Placental Blood Program of the New York Blood Center has developed techniques for collecting, testing, freezing and searching units on behalf of unrelated patients in need of hematopoietic stem cell replacement since 1993 and provided analysis of the outcomes of these transplants identified variables associated with clinical outcomes. In this review, after considering practical and conceptual aspects of the technology, we update information on the clinical outcomes of these transplants. MATERIALS AND METHODS: All 861 patients transplanted through 1999 with PCB from our Program are included in this report. Two thirds were diagnosed with leukaemia or lymphoma, 25% with inherited conditions and 7% with acquired diseases. Outcome data were provided by the respective Transplant Center and analyses included both univariate and multivariate regression tests and actuarial (Kaplan-Meier) techniques. RESULTS: Engraftment was achieved by over 90% of recipients (Kaplan Meier estimate). Multivariate analysis confirmed the influence of cell dose, HLA matching, disease diagnosis and transplant center location (US vs. foreign). Patient age and HLA match grade independently affected the frequency and severity of acute graft vs. host disease. Leukaemic relapse was associated with the stage of disease at transplantation and the prior existence of acute graft vs. host disease. The probability of transplant-related events was independently associated with disease diagnosis, cell dose, number of HLA mismatches and transplant center, while the cell dose failed to associate significantly with the relative risk of reaching this endpoint in the subset of patients who achieved engraftment. Overall, event-free survival rates at one year post-transplant were 49 and 30%, respectively for genetic disease and haematologic malignancies and 35% for patients with acquired diseases, respectively. CONCLUSIONS: These results confirm and extend earlier data, particularly establishing the significant association of transplant success with histocompatibility matching grades, and indicatng the urgency of improving the transplant match levels. PMID- 11102278 TI - From leukocyte reduction to leukocyte transfusion: the immunological effects of transfused leukocytes. AB - In transfusion medicine, mononuclear leukocytes have been studied more often as contaminants of red blood cells or platelets responsible for adverse transfusion outcomes than as therapeutic cells; leukocyte transfusion has been effective in augmenting recipient immunity only in limited clinical situations. Studies in leukocyte reduction and leukocyte transfusion have progressed separately as if the leukocytes' adverse and therapeutic effects result from different immunological mechanisms. With growing clinical experience, however, it is increasingly clear that some adverse immune effects may be exploited for therapeutic benefit. Advances in clinical immunology, understanding of the variety of cells and functions in the leukocyte fraction of blood, and blood component preparation technology may lead to new ways of deriving immunological benefit from transfused blood leukocytes while minimizing their adverse effects. This chapter reviews the current uses of leukocyte reduction and mononuclear leukocyte transfusion, with an emphasis on the relationship between transfusion associated graft-versus-host disease and donor lymphocyte infusion in controlling relapsed leukaemias. PMID- 11102279 TI - Which agents threaten blood safety in the future? AB - The safety of the blood supply is critical to many parts of modern medicine. In a time when prescriber's and the public's expectations are increasing, it is essential that transfusion services globally ensure the safety of the blood supply. There are, however, many threats to this safety, one being the appearance of new infectious agents. Such agents may be truly 'novel', or may be existing agents, known but not routinely screened for, posing a new or increased threat. However, before an agent is considered to be a true threat to blood safety it must be well characterized, and evidence must be presented that (i) transfusion transmission is a significant route of spread, and (ii) the agent causes significant clinical disease. If either of these criteria are not met, the question has to be asked as to whether the agent is truly a threat to blood safety. PMID- 11102280 TI - Will genome detection replace serology in blood screening for microbial agents? AB - The residual risk of transfusion-transmitted viral infection in developed countries is considered minimal or negligible. However, zero risk remains a strong political objective. Genomic screening for HCV, HIV and HBV represents a major advance, eliminating infectious blood donations collected during the pre seroconversion window period, rare cases of immunosilent infections and, possibly, a large spectrum of viral variants. In Western countries, HCV RNA genomic screening started on pools of 16-400 plasma samples from individual donations. Pooling may produce false-positive and false-negative results. Individual donation testing is more suitable to blood screening but requires multiplexing, automation, and affordable cost. Because donations from individuals who are HBV DNA-negative/serologically positive, or those apparently recovered from HCV infection, may remain infectious, it is unlikely that HBsAg, anti-HCV, and anti-HIV will be discontinued when genomic screening is extended to all three viruses. HIV-1 p24 antigen may prove redundant with HIV RNA screening. Anti-HTLV I and HTLV-II will remain more effective than genomic testing. PMID- 11102281 TI - The risks of transfusion-transmitted infection: direct estimation and mathematical modelling. AB - Direct measurement of the risk of transfusion-transmitted infection (TTI) is practical and accurate only if the level of risk is high. Historically, studies that established frozen repositories of transfusion recipient and/or blood donor samples were important in establishing the risk of many TTI agents, including the human immunodeficiency virus (HIV), hepatitis B virus (HBV) and hepatitis C virus (HCV). However, given the current very low risk of TTI, mathematical modelling is necessary to estimate the magnitude of such a risk. For agents for which routine blood donor screening is performed, most of this risk comes from transfusion of units collected in the window period between donor infection and a positive blood screening assay. The incidence/window period model has been used to estimate the magnitude of such risks (of the order of 1:100 000 to 1:1 000 000) and for predicting the extent of risk reduction that can be expected with implementation of new tests. Direct estimation and mathematical modelling approaches are both important tools for future assessment of potential, new or emerging TTI agents. PMID- 11102282 TI - Red blood cell substitutes. AB - Soluble polymerized haemoglobin (polyhaemoglobin) is now in a phase III clinical trials. Patients have received up to 20 units (10 litres) in trauma surgery and other surgery. Polyhaemoglobin can be stored for more than 1 year. Haemoglobin solutions have no blood group antigen and can be used as a 'universal donor' oxygen carrier. They can also be sterilized. With a circulation half-life of 24 hours they are undergoing trials for peri-operative use. For conditions with potential for ischaemia-reperfusion injuries, a new polyhaemoglobin-superoxide dismutase-catalase, which can reduce oxygen radicals, is being developed. Recombinant human haemoglobin has been tested in clinical trials, and a new type of recombinant human haemoglobin that has low affinity for nitric oxide is being developed for clinical trials. To increase the circulation time, artificial red blood cells have been prepared with a bilayer lipid membrane (haemoglobin liposomes) or with a biodegradable polymer membrane-like polylactide (haemoglobin nanocapsules). Synthetic chemicals such as perfluorochemicals are also being developed and tested in clinical trials as red blood cell substitutes. PMID- 11102284 TI - Index PMID- 11102283 TI - Biotechnology: alternatives to human plasma-derived therapeutic proteins. AB - Proteins derived from human plasma have become critically important therapeutic products since their introduction in the 1940s. In the last 20 years, the tools of molecular biology have provided alternatives to the administration of the natural products. Recombinant analogues of Factor VIII and Factor IX are commercially available, and recombinant forms of other plasma proteins are under development. Genetic engineering also provides the opportunity to modify a natural protein to improve the efficiency with which it can be produced in vitro, or to change its therapeutic profile. More efficient production systems, such as transgenic plants or animals, may yield less costly therapies and a wider availability of products that are now in limited supply. Finally, gene therapy offers the prospect of permanently correcting conditions arising from deficiencies in any one of several plasma proteins, freeing individuals from the need to undergo periodic treatments with exogenous proteins. PMID- 11102285 TI - Preface PMID- 11102286 TI - Use of closely related affected individuals for the genetic study of complex diseases in founder populations. AB - We propose a method, the maximum identity length contrast (MILC) statistic, to locate genetic risk factors for complex diseases in founder populations. The MILC approach compares the identity length of parental haplotypes that are transmitted to affected offspring with the identity length of those that are not transmitted to affected offspring. Initially, the statistical properties of the method were assessed using randomly selected affected individuals with unknown relationship. Because both nuclear families with multiple affected sibs and large pedigrees are often available in founder populations, we performed simulations to investigate the properties of the MILC statistic in the presence of closely related affected individuals. The simulation showed that the use of closely related affected individuals greatly enhances the power of the statistic. For a given sample size and type I error, the use of affected sib pairs, instead of affected individuals randomly selected from the population, could increase the power by a factor of two. This increase was related to an increase of kinship-coefficient contrast between haplotype groups when closely related individuals were considered. The MILC approach allows the simultaneous use of affected individuals from a founder population and affected individuals with any kind of relationship, close or remote. We used the MILC approach to analyze the role of HLA in celiac disease and showed that the effect of HLA may be detected with the MILC approach by typing only 11 affected individuals, who were part of a single large Finnish pedigree. PMID- 11102287 TI - Genetics of event-related brain potentials in response to a semantic priming paradigm in families with a history of alcoholism. AB - Event-related brain potentials (ERPs) are altered in patients with a variety of psychiatric disorders and may represent quantitative correlates of disease liability that are more amenable to genetic analysis than disease status itself. Results of a genomewide linkage screen are presented for amplitude of the N4 and P3 components of the ERP, measured at 19 scalp locations in response to a semantic priming task for 604 individuals in 100 pedigrees ascertained as part of the Collaborative Study on the Genetics of Alcoholism. N4 and P3 amplitudes in response to three stimuli (nonwords, primed words [i.e., antonyms], and unprimed words) all showed significant heritabilities, the highest being.54. Both N4 and P3 showed significant genetic correlations across stimulus type at a given lead and across leads within a stimulus, indicating shared genetic influences among the traits. There were also substantial genetic correlations between the N4 and P3 amplitudes for a given lead, even across stimulus type. N4 amplitudes showed suggestive evidence of linkage in several chromosomal regions, and P3 amplitudes showed significant evidence of linkage to chromosome 5 and suggestive evidence of linkage to chromosome 4. PMID- 11102288 TI - Ecological risk assessment of endocrine disruptors. AB - The European Centre for Ecotoxicology and Toxicology of Chemicals proposes a tiered approach for the ecological risk assessment of endocrine disruptors, integrating exposure and hazard (effects) characterization. Exposure assessment for endocrine disruptors should direct specific tests for wildlife species, placing hazard data into a risk assessment context. Supplementing the suite of mammalian screens now under Organization for Economic Cooperation and Development (OECD) validation, high priority should be given to developing a fish screening assay for detecting endocrine activity in oviparous species. Taking into account both exposure characterization and alerts from endocrine screening, higher tier tests are also a priority for defining adverse effects. We propose that in vivo mammalian and fish assays provide a comprehensive screening battery for diverse hormonal functions (including androgen, estrogen, and thyroid hormone), whereas Amphibia should be considered at higher tiers if there are exposure concerns. Higher tier endocrine-disruptor testing should include fish development and fish reproduction tests, whereas a full life-cycle test could be subsequently used to refine aquatic risk assessments when necessary. For avian risk assessment, the new OECD Japanese quail reproduction test guideline provides a valuable basis for developing a test to detecting endocrine-mediated reproductive effects; this species could be used, where necessary, for an avian life-cycle test. For aquatic and terrestrial invertebrates, data from existing developmental and reproductive tests remain of high value for ecological risk assessment. High priority should be given to research into comparative endocrine physiology of invertebrates to support data extrapolation to this diverse fauna. PMID- 11102289 TI - Determination of monomethylarsonous acid, a key arsenic methylation intermediate, in human urine. AB - In this study we report on the finding of monomethylarsonous acid [MMA(III)] in human urine. This newly identified arsenic species is a key intermediate in the metabolic pathway of arsenic biomethylation, which involves stepwise reduction of pentavalent to trivalent arsenic species followed by oxidative addition of a methyl group. Arsenic speciation was carried out using ion-pair chromatographic separation of arsenic compounds with hydride generation atomic fluorescence spectrometry detection. Speciation of the inorganic arsenite [As(III)], inorganic arsenate [As(V)], monomethylarsonic acid [MMA(V)], dimethylarsinic acid [DMA(V)], and MMA(III) in a urine sample was complete in 5 min. Urine samples collected from humans before and after a single oral administration of 300 mg sodium 2,3 dimercapto-1-propane sulfonate (DMPS) were analyzed for arsenic species. MMA(III) was found in 51 out of 123 urine samples collected from 41 people in inner Mongolia 0-6 hr after the administration of DMPS. MMA(III )in urine samples did not arise from the reduction of MMA(V) by DMPS. DMPS probably assisted the release of MMA(III) that was formed in the body. Along with the presence of MMA(III), there was an increase in the relative concentration of MMA(V) and a decrease in DMA(V) in the urine samples collected after the DMPS ingestion. PMID- 11102290 TI - Prospective study of exposure to environmental tobacco smoke and dysmenorrhea. AB - Dysmenorrhea is a common gynecologic disorder in women of reproductive age. Previous studies have found an association between current cigarette smoking and prevalence of dysmenorrhea. This study investigated the association between exposure to environmental tobacco smoke (ETS) and the occurrence of dysmenorrhea among women without a history of this disorder. The study population consisted of 165 newly wed, nonsmoking Chinese women (in Shenyang, China), who intended to get pregnant and who had no past history of dysmenorrhea at the time of enrollment. These women completed a baseline questionnaire interview upon enrollment and were prospectively followed by daily diary. Dysmenorrhea was defined as a diary recording of abdominal pain or low back pain for at least 2 days during a menstrual period. A subject's ETS exposure was defined as the mean number of cigarettes smoked per day at home by household members over an entire menstrual cycle before the menstrual period. A logistic regression model was used to assess the effect of ETS on the risk of dysmenorrhea, with adjustment for age, body mass index, education, season, area of residence, occupation, shift work, perceived stress, passive smoking at work, and occupational exposure to chemical hazards, dust, and noise. Generalized estimating equations were used to account for autocorrelations as a result of multiple cycles per subject. This report is based on 625 prospectively followed menstrual cycles with complete baseline and diary data. ETS exposure was reported in 77% of cycles, within which average daily exposures throughout the cycle ranged from 0.02 to 10. 3 cigarettes. The incidence of dysmenorrhea was 9.7% and 13.3% among nonexposed and exposed cycles, respectively. Among ETS-exposed cycles, there was a positive dose-response relationship between the numbers of cigarettes smoked and the relative risk of dysmenorrhea. The adjusted odds ratios of dysmenorrhea associated with "low," "middle," and "high" tertiles of ETS exposure versus no exposure were 1.1 [95% confidence interval (CI), 0.5-2.6], 2.5 (CI, 0.9-6.7), and 3.1 (CI, 1.2-8.3), respectively. The findings were consistent with those of analyses limited to the first follow-up menstrual cycle from each woman. These data suggest a significant dose-response relationship between exposure to ETS and an increased incidence of dysmenorrhea in this cohort of young women. PMID- 11102292 TI - Data quality in predictive toxicology: identification of chemical structures and calculation of chemical properties. AB - Every technique for toxicity prediction and for the detection of structure activity relationships relies on the accurate estimation and representation of chemical and toxicologic properties. In this paper we discuss the potential sources of errors associated with the identification of compounds, the representation of their structures, and the calculation of chemical descriptors. It is based on a case study where machine learning techniques were applied to data from noncongeneric compounds and a complex toxicologic end point (carcinogenicity). We propose methods applicable to the routine quality control of large chemical datasets, but our main intention is to raise awareness about this topic and to open a discussion about quality assurance in predictive toxicology. The accuracy and reproducibility of toxicity data will be reported in another paper. PMID- 11102291 TI - Longitudinal study of dust and airborne endotoxin in the home. AB - To characterize the seasonal variability of endotoxin levels, we measured endotoxin in dust from the bed, bedroom floor, and kitchen floor in 20 homes, and in air from the bedroom in 15 of the homes. All homes were located in the greater Boston, Massachusetts, area and were sampled each month from April 1995 to June 1996. Outdoor air was collected at two locations. We found greater within-home than between-home variance for bedroom floor, kitchen floor, and airborne endotoxin. However, the reverse was true for bed dust endotoxin. Thus, studies using single measurements of dust endotoxin are most likely to reliably distinguish between homes if bed dust is sampled. Dust endotoxin levels were not significantly associated with airborne endotoxin. Airborne endotoxin was significantly (p = 0. 04) and positively associated with absolute humidity in a mixed-effect model adjusting for a random home effect and fixed effect of sampling month and home characteristics. This finding implies that indoor humidity may be an important factor controlling endotoxin exposure. We found a significant (p < 0.05) seasonal effect in kitchen floor dust (spring > fall) and bedroom airborne endotoxin (spring > winter), but not in the other indoor samples. We found significant seasonal pattern in outdoor airborne endotoxin (summer > winter). PMID- 11102293 TI - Influence of the consumption of fatty Baltic Sea fish on plasma levels of halogenated environmental contaminants in Latvian and Swedish men. AB - We examined the influence of widely varied consumption of fatty fish from the Baltic Sea and of age on plasma concentrations of polychlorinated biphenyls (PCBs), polychlorobiphenylols (OH-PCBs), 2, 2-bis(4-chlorophenyl)-1,1,1 trichloroethane (4,4'-DDT), 2, 2-bis(4-chlorophenyl)-1,1-dichloroethane (4,4' DDE), 2,2',4, 4'-tetrabromodiphenyl ether (BDE-47), hexachlorobenzene (HCB), and pentachlorophenol (PCP) in Latvian and Swedish men. Both age and fish consumption were significantly correlated with the concentrations of [sigman]PCB, [sigman]OH PCB, 4,4'-DDE, 4,4'-DDT, and HCB. In the case of BDE-47, no significant relationship with age was observed, and fish consumption had the largest relative effect on plasma concentrations of this contaminant. This relationship may be a result of exposure to BDE-47 having been more recent than that of PCBs and DDE, or because the half-life of BDE-47 may be shorter than that of PCB and DDE. Latvian men demonstrated higher plasma levels of DDE and DDT but lower levels of [sigman]PCB and PCP than did Swedish men. The corresponding levels of HCB and BDE 47 were similar in both countries. The Spearman's rank correlation coefficient obtained by comparing the level of the metabolite 4-hydroxy-2,3,3',4',5 pentachlorobiphenyl (4-OH-CB107) to the combined levels of its parent compounds, 2,3,3',4, 4'-pentachlorobiphenyl (CB-105) and 2,3',4,4',5-pentachlorobiphenyl (CB 118), was higher than the median correlation coefficient obtained upon comparing the level of this metabolite to all other possible combinations of two PCB levels. No other increased correlation between metabolite and parent PCB concentration was observed. PMID- 11102294 TI - Selenium concentration in the milk of breast-feeding mothers and its geographic distribution. AB - A total of 905 human milk samples collected in all provinces of Poland, between 12 and 75 days of lactation, were analyzed for selenium concentration. The distribution of Se levels in milk between the provinces was narrow and varied from 8.81 to 11.58 ng/mL, with the mean value (+/- SD) of 10.24 +/- 2.82 ng/mL. The regions with lower levels of Se were in the central and eastern part of Poland; the areas with higher values were in the northern, western, and southern parts of Poland. No significant correlations were found between Se levels in milk and the age of lactating mothers or between Se levels and the postpartum period. The calculated daily Se intakes by breast-fed infants varied from 6.46 to 8.50 microg/day, with the mean value of 7.52 microg/day. This amount does not meet the recommended dietary allowances for infants between 0 and 6 months of age. Based on Se levels in human milk, we present a selenium map of Poland. PMID- 11102295 TI - Environmental malignant mesothelioma in southern Anatolia: a study of fifty cases. AB - Malignant mesothelioma is a highly aggressive tumor of the serous membranes, which in humans results from exposure to asbestos and asbestiform fibers. Although occupational malignant mesothelioma is still the most common form of this lesion, naturally contaminated soil can play an important role in the development of environmental malignant mesothelioma in some parts of the world. Fifty cases of malignant mesothelioma (MM) from southern Turkey with no occupational history of asbestos exposure were reviewed regarding pathologic and clinical features. A case of hyaline fibrous plaque of the pleura was also included in this series. Histologically the cases were classified as epithelial (36 cases); sarcomatous (7 cases); and biphasic (7 cases). One of the sarcomatous cases was desmoplastic. Ultrastructural examination of the tumor tissue in three cases revealed long-surface microvilli in epithelial cells. Interstitial cells of the lung in one case showed electron-dense asbestos fibers in the cytoplasm. Mineralogical analyses of the lung tissue in three cases of MM and the case of pleural plaque showed high amounts of asbestos fibers most consistent with tremolite and actinolite. The clinical and pathologic features of our cases support that the environmental inhalation of asbestos is still a major health problem in some parts of Turkey. PMID- 11102296 TI - Particle concentrations in urban microenvironments. AB - Although ambient particulate matter has been associated with a range of health outcomes, the health risks for individuals depend in part on their daily activities. Information about particle mass concentrations and size distributions in indoor and outdoor microenvironments can help identify high-risk individuals and the significant contributors to personal exposure. To address these issues in an urban setting, we measured particle count concentrations in four size ranges and particulate matter (3/4) 10 microm (PM(10)) concentrations outdoors and in seven indoor microenvironments in Boston, Massachusetts. Particle counts and PM(10) concentrations were continuously measured with two light-scattering devices. Because of the autocorrelation between sequential measurements, we used linear mixed effects models with an AR-1 autoregressive correlation structure to evaluate whether differences between microenvironments were statistically significant. In general, larger particles were elevated in the vicinity of significant human activity, and smaller particles were elevated in the vicinity of combustion sources, with indoor PM(10) concentrations significantly higher than the outdoors on buses and trolleys. Statistical models demonstrated significant variability among some indoor microenvironments, with greater variability for smaller particles. These findings imply that personal exposures can depend on activity patterns and that microenvironmental concentration information can improve the accuracy of personal exposure estimation. PMID- 11102297 TI - Prevalence of headache among handheld cellular telephone users in Singapore: a community study. AB - We carried out a cross-sectional community study in Singapore to determine the prevalence of specific central nervous system (CNS) symptoms among hand-held cellular telephone (HP) users compared to nonusers and to study the association of risk factors and CNS symptoms among HP users. A total of 808 men and women between 12 and 70 years of age, who lived in one community, were selected using one-stage cluster random sampling and responses to a structured questionnaire. The prevalence of HP users was 44.8%. Headache was the most prevalent symptom among HP users compared to non-HP users, with an adjusted prevalence rate ratio of 1.31 [95% confidence interval, 1.00-1.70]. There is a significant increase in the prevalence of headache with increasing duration of usage (in minutes per day). Prevalence of headache was reduced by more than 20% among those who used hand-free equipment for their cellular telephones as compared to those who never use the equipment. The use of HPs is not associated with a significant increase of CNS symptoms other than headache. PMID- 11102298 TI - Distribution of particulate matter and tissue remodeling in the human lung. AB - We examined the relationship between intrapulmonary particle distribution of carbonaceous and mineral dusts and remodeling of the airways along anatomically distinct airway paths in the lungs of Hispanic males from the central valley of California. Lung autopsy specimens from the Fresno County Coroner's Office were prepared by intratracheal instillation of 2% glutaraldehyde at 30 cm H(2)O pressure. Two distinct airway paths into the apico-posterior and apico-anterior portions of the left upper lung lobe were followed. Tissue samples for histologic analysis were generally taken from the intrapulmonary second, fourth, sixth, and ninth airway generations. Parenchymal tissues beyond the 12th airway generation of each airway path were also analyzed. There was little evidence of visible particle accumulation in the larger conducting airways (generations 2-6), except in bronchial-associated lymphoid tissues and within peribronchial connective tissue. In contrast, terminal and respiratory bronchioles arising from each pathway revealed varying degrees of wall thickening and remodeling. Walls with marked thickening contained moderate to heavy amounts of carbonaceous and mineral dusts. Wall thickening was associated with increases in collagen and interstitial inflammatory cells, including dust-laden macrophages. These changes were significantly greater in first-generation respiratory bronchioles compared to second- and third-generation respiratory bronchioles. These findings suggest that accumulation of carbonaceous and mineral dust in the lungs is significantly affected by lung anatomy with the greatest retention in centers of lung acini. Furthermore, there is significant remodeling of this transitional zone in humans exposed to ambient particulate matter. PMID- 11102299 TI - Airborne particles are a risk factor for hospital admissions for heart and lung disease. AB - We examined the association between particulate matter [less than/equal to] 10 microm; (PM(10)) and hospital admission for heart and lung disease in ten U.S. cities. Our three goals were to determine whether there was an association, to estimate how the association was distributed across various lags between exposure and response, and to examine socioeconomic factors and copollutants as effect modifiers and confounders. We fit a Poisson regression model in each city to allow for city-specific differences and then combined the city-specific results. We examined potential confounding by a meta-regression of the city-specific results. Using a model that considered simultaneously the effects of PM(10) up to lags of 5 days, we found a 2.5% [95% confidence interval (CI), 1.8-3. 3] increase in chronic obstructive pulmonary disease, a 1.95% (CI, 1. 5-2.4) increase in pneumonia, and a 1.27% increase (CI, 1-1.5) in CVD for a 10 microg/m(3) increase in PM(10). We found similar effect estimates using the mean of PM(10) on the same and previous day, but lower estimates using only PM(10) for a single day. When using only days with PM(10) < 50 mg/m(3), the effect size increased by [greater/equal to] 20% for all three outcomes. These effects are not modified by poverty rates or minority status. The results were stable when controlling for confounding by sulfur dioxide, ozone, and carbon monoxide. These results are consistent with previous epidemiology and recent mechanistic studies in animals and humans. PMID- 11102300 TI - Cord serum cotinine as a biomarker of fetal exposure to cigarette smoke at the end of pregnancy. AB - This study investigated the association between biomarkers of fetal exposure to cigarette smoke at the end of pregnancy, cotinine in cord serum and in maternal and newborn urine samples, and quantitative measurement of smoking intake and exposure evaluated by maternal self-reported questionnaire. Study subjects were 429 mothers and their newborns from a hospital in Barcelona, Spain. A questionnaire including smoking habits was completed in the third trimester of pregnancy and on the day of delivery. Cotinine concentration in cord serum was associated with daily exposure to nicotine in nonsmokers and with daily nicotine intake in smokers. The geometric mean of cotinine concentration in cord serum statistically discriminated between newborns from nonexposed and exposed nonsmoking mothers, and between these two classes and smokers, and furthermore was able to differentiate levels of exposure to tobacco smoke and levels of intake stratified in tertiles. Urinary cotinine levels in newborns from nonsmoking mothers exposed to more than 4 mg nicotine daily were statistically different from levels in two other categories of exposure. Cotinine concentration in urine from newborns and from mothers did not differentiate between exposure and nonexposure to environmental tobacco smoke (ETS) in nonsmoking mothers. Cord serum cotinine appeared to be the most adequate biomarker of fetal exposure to smoking at the end of pregnancy, distinguishing not only active smoking from passive smoking, but also exposure to ETS from nonexposure. PMID- 11102301 TI - Limb malformations and abnormal sex hormone concentrations in frogs. AB - Declines in amphibian populations, and amphibians with gross malformations, have prompted concern regarding the biological status of many anuran species. A survey of bullfrogs, Rana catesbeiana, and green frogs, Rana clamitans, conducted in central and southern New Hampshire showed malformed frogs at 81% of the sites sampled (13 of 16 sites). Brain gonadotropin-releasing hormone (GnRH) and the synthesis of androgens and estradiol, hormones essential to reproductive processes, were measured from limb-malformed and normal (no limb malformation) frogs. Normal frogs had significantly higher concentrations (nearly 3-fold) of in vitro produced androgens and of brain GnRH than malformed frogs. Because most malformations are thought to occur during development, we propose that environmental factors or endocrine-disrupting chemicals that may cause developmental abnormalities also act during early development to ultimately cause abnormally reduced GnRH and androgen production in adult frogs. The consequences of reduced GnRH and androgens on anuran reproductive behavior and population dynamics are unknown but certainly may be profound and warrant further research. PMID- 11102302 TI - Lead isotopes as a supplementary tool in the routine evaluation of household lead hazards. AB - The advent of magnetic sector inductively coupled plasma-mass spectrometry (ICP MS) allows rapid, accurate, and precise measurement of lead isotopes in environmental and biological samples at a lower cost than traditional methods. This may increase the feasibility of including lead isotope measurements as a routine tool to identify household sources of lead exposure to children. Here, we present three household case studies to illustrate how lead hazard evaluations by an environmental specialist could be supplemented with routine lead isotope analyses of potential lead sources and blood. Sampling for lead isotopes was undertaken following the U.S. Department of Housing and Urban Development regulatory guidelines for the evaluation of lead hazards in housing, and with the consideration of minimizing the additional costs associated with lead isotope measurements. The range of isotopic ratios within a single residence was large enough to allow the characterization of different lead sources, particularly when both major (e.g., (207)Pb/(206)Pb) and minor (e.g., (206)Pb/(204)Pb) isotope ratios were considered. These cases illustrate the utility of the lead isotope method to identify main source(s) of lead exposure to the child; discard unlikely sources of exposure to the child; point to sources of lead to dust; and substantiate or refine the environmental assessment based exclusively on lead concentrations and loadings. Thus, a more effective evaluation of household lead hazards would likely benefit from considering a) lead concentrations and loadings in and around the household environment; b) all isotopic ratios of potential lead sources within that environment; and c) information about behavioral habits, as well as an evaluation of viable pathways of exposure to the child. PMID- 11102303 TI - The association between caries and childhood lead exposure. AB - Epidemiologic studies suggest an association between lead exposure and caries. Our objective was to establish whether children with a higher lead exposure as toddlers had more caries at school age than children with a lower lead exposure. We used a retrospective cohort design. A sample of children who attended second and fifth grades in the Rochester, New York, public schools during the 1995-1996 and 1996-1997 school years were examined for caries through a dental screening program. For each child we assessed the number of decayed, missing, or filled surfaces on permanent teeth (DMFS), and the number of decayed or filled surfaces on deciduous teeth (dfs); the number of surfaces at risk (SAR) was also recorded. Lead exposure was defined as the mean of all blood lead levels collected between 18 and 37 months of age by fingerstick [provided the blood lead level was [less than/equal to] 10 microg/dL)] or venipuncture. A total of 248 children (197 second graders and 51 fifth graders) were examined for caries and had a record of blood lead levels to define lead exposure. The mean dfs was 3.4 (range 0-29); the mean DMFS was 0.5 (range 0-8). Logistic regression was used to examine the association between the proportion of children with DMFS [Greater/equal to] 1, and the proportion with dfs [Greater/equal to] 1, and lead exposure [< 0.48 micromol/L vs. [Greater/equal to] 0.48 micromol/L (< 10 microg/dL vs. [Greater/equal to] 10 microg/dL)] while controlling for SAR, age at examination, and grade in school. For DMFS, the adjusted odds ratio was 0.95 [95% confidence interval (CI), 0.43-2.09; p = 0.89); for dfs, the odds ratio was 1.77 (95% CI, 0.97-3.24; p = 0.07). This study did not demonstrate that lead exposure > 10 microg/dL as a toddler was a strong predictor of caries among school-age children. However, the results should be interpreted cautiously because of limitations in the assessment of lead exposure and limited statistical power. PMID- 11102305 TI - A challenge for the new millennium: eliminating health disparities and achieving educational and workforce diversity. PMID- 11102304 TI - An unusual case of organophosphate intoxication of a worker in a plastic bottle recycling plant: an important reminder. AB - A young man was sent to our emergency unit because he had suffered from vomiting and cold sweating for 2 days. At the time he was admitted, he had no acute abdominal pains or gastrointestinal symptoms, and a physical examination revealed nothing but a faster heart rate and moist, flushing skin. The patient had worked for 6 years at a plastic bottle-recycling factory, but none of his co-workers had the same symptoms. Nevertheless, because the plant also recycled pesticide bottles, we suspected organophosphate pesticide intoxication. The patient's plasma acetylcholinesterase level was checked, revealing 1498.6 microU/L (normal range: 2,000-5, 000) on the first day and 1,379 microU/L on the second day. Upon questioning, the patient recalled that one of his shoe soles had been damaged and that his foot had been wet from walking all day in rain collected on the factory floor on the day that his symptoms first occurred. We conducted a study in the change of preshift and postshift acetylcholinesterase levels among six of his co workers on a rainy day. We used the Wilcoxon signed rank test to compare the preshift and postshift plasma acetylcholinesterase levels; no significant difference was revealed (p = 0.600), leaving contamination via the damaged shoe sole suspect. We reviewed the literature on organophosphate intoxication; pesticide bottle-recycling factories were reported to be at a low risk of organophosphate toxicity in the working environment. However, because the potential risk of intoxication is still present, protective equipment such as clothing, gloves, and water-proof shoes should be worn, and employees should be educated on the potential risks. PMID- 11102306 TI - Brown's response: difficulties with "the balkans" AB - Respon to Csaba Varga' comments. Environ Health Perspect 108:494-494 (2000). PMID- 11102307 TI - Comments on "The worst of both worlds: poverty and politics in the Balkans. PMID- 11102308 TI - Arsenic in urine and drinking water. PMID- 11102309 TI - Questioning coal ash deregulation. PMID- 11102310 TI - Panama left with an explosive issue. PMID- 11102311 TI - Estuarine nutrient loads still a problem PMID- 11102313 TI - Logging gets a break PMID- 11102312 TI - New grasspeas lessen risk of paralysis. PMID- 11102314 TI - Let's work together: connecting research and the community. PMID- 11102315 TI - Caveolins in cholesterol trafficking and signal transduction: implications for human disease. AB - Caveolins are a family of proteins that coat the cytoplasmic face of caveolae, vesicular invaginations of the plasma membrane. These proteins are central to the organization of the proteins and lipids that reside in caveolae. Caveolins transport cholesterol to and from caveolae, and they regulate the activity of signaling proteins that reside in caveolae. Through studying the genes encoding the caveolae coat proteins, we have learned much about how they perform these multiple functions. PMID- 11102316 TI - Bruton tyrosine kinase (BTK) in X-linked agammaglobulinemia (XLA). AB - X-linked agammaglobulinemia (XLA) is a heritable immunodeficiency disorder that is caused by a differentiation block leading to almost complete absence of B lymphocytes and plasma cells. The affected protein is a cytoplasmic protein tyrosine kinase, Bruton's agammaglobulinemia tyrosine kinase (Btk). Btk along with Tec, Itk, Bmx and Txk belong to a distinct family of protein kinases. These proteins contain five regions; PH, TH, SH3, SH2 and kinase domains. Mutations causing XLA may affect any of these domains. About 380 unique mutations have been identified and are collected in a mutation database, BTKbase. Here, we describe the structure, function, and interactions of the affected signaling molecules in atomic detail. PMID- 11102317 TI - Cell-cycle dysregulation in breast cancer: breast cancer therapies targeting the cell cycle. AB - Breast cancer is the most commonly diagnosed cancer in American women. The underlying mechanisms that cause aberrant cell proliferation and tumor growth involve conserved pathways, which include components of the cell cycle machinery. Proto-oncogenes, growth factors, and steroids have been implicated in the pathogenesis of breast cancer. Surgery, local irradiation, and chemotherapy have been the mainstay of treatment for early and advanced stage disease. Potential targets for selective breast cancer therapy are herein reviewed. Improved understanding of the biology of breast cancer has led to more specific "targeted therapies" directed at biological processes that are selectively deregulated in the cancerous cells. Examples include tamoxifen for estrogen receptor positive tumors and imunoneutralizing antibodies such as trastuzumab for Her2/neu overexpressing tumors. Other novel anticancer agents such as paclitaxel, a microtubule binding molecule, and flavopiridol, a cyclin dependent kinase inhibitor, exert their anticancer effects by inhibiting cell cycle progression. PMID- 11102318 TI - Yeast perspectives on HIV-1 VPR. AB - Increasing evidence suggests that HIV-1 viral protein R (Vpr) plays an important role in viral pathogenesis, as its functions are being linked to viral activation, suppression of human immune functions and depletion of human CD4 lymphocytes, which are the major clinical manifestation of AIDS. In vitro, Vpr shows multiple activities both in mammalian and yeast cells, which include nuclear transport, induction of cell cycle G2 arrest, morphological changes and cell death. The occurrence of these activities in yeast indicates that Vpr interacts with highly conserved cellular processes to cause these effects and allows Vpr activities to be studied in these genetically well characterized organisms. Studies of Vpr in fission yeast (Schizosaccharomyces pombe) and budding yeast (Saccharomyces cerevisiae) have helped to establish these major conclusions. 1) Vpr induces G2 arrest through inhibitory phosphorylation of the cyclin-dependent kinase by a pathway in which protein phosphatase 2A plays an important role. 2) Vpr fulfills its essential role in the nuclear transport of the viral pre-integration complex by binding to a novel site on importin ?. 3) Vpr induces apoptosis by directly permeabilizing the mitochondrial membrane. 4) Vpr also appears to kill cells by mitochondrial-independent mechanisms. 5) G2 arrest and cell death induced by Vpr are two independent functions, and 6) amino acid residues of Vpr at position 29, 33 and 71 are important sites for maintaining the overall structure of Vpr. Future studies of Vpr in yeast are expected to make additional contributions to understanding the mechanisms of Vpr activities and may also help address the importance of these activities during the course of a HIV-1 infection. PMID- 11102319 TI - Evolving model of depression as an expression of multiple interacting risk factors. PMID- 11102320 TI - Sex and suicide. Gender differences in suicidal behaviour. PMID- 11102321 TI - Gender differences in depression. Critical review. AB - BACKGROUND: With few exceptions, the prevalence, incidence and morbidity risk of depressive disorders are higher in females than in males, beginning at mid puberty and persisting through adult life. AIMS: To review putative risk factors leading to gender differences in depressive disorders. METHOD: A critical review of the literature, dealing separately with artefactual and genuine determinants of gender differences in depressive disorders. RESULTS: Although artefactual determinants may enhance a female preponderance to some extent, gender differences in depressive disorders are genuine. At present, adverse experiences in childhood, depression and anxiety disorders in childhood and adolescence, sociocultural roles with related adverse experiences, and psychological attributes related to vulnerability to life events and coping skills are likely to be involved. Genetic and biological factors and poor social support, however, have few or no effects in the emergence of gender differences. CONCLUSIONS: Determinants of gender differences in depressive disorders are far from being established and their combination into integrated aetiological models continues to be lacking. PMID- 11102322 TI - Needs assessment for mentally disordered offenders: measurement of 'ability to benefit' and outcome. AB - BACKGROUND: The Department of Health defines needs as "the ability to benefit from healthcare interventions". Outcome measurement is an integral component of needs assessment because it underpins 'ability to benefit'. AIMS: To propose a framework for addressing the measurement of outcome in relation to mentally disordered offenders (MDOs). METHOD: Based on a literature search, the paper reviews the definition and measurement of outcome in general mental health care and specifically in relation to MDOs. It analyses the problems of conducting outcome research in relation to MDOs. RESULTS: A framework for outcome measurement in relation to MDOs is presented. Outcome is placed within a broader framework that relates to service evaluation. CONCLUSIONS: Current measurement of outcome in relation to MDOs is inadequate. A comprehensive framework that acknowledges the multi-dimensional nature of outcome is essential. Researchers must be able to justify the dimensions they prioritise. PMID- 11102323 TI - Recent life events, cortisol, dehydroepiandrosterone and the onset of major depression in high-risk adolescents. AB - BACKGROUND: It is not clear whether cortisol or dehydroepiandrosterone (DHEA) hypersecretion increases the risk for major depression in the presence of undesirable life events. AIMS: To determine whether there is a specific pattern of psychoendocrine factors that predicts the onset of major depressive disorder. METHOD: 180 adolescents (73 boys, 107 girls) at high risk for psychopathology were assessed for cortisol, DHEA, depressive symptoms, life events and psychiatric disorder at entry and 12 months later. RESULTS: Major depression was predicted for both genders by the additive effects of: higher depressive symptoms; personal disappointments and losses only in the month before onset; one or more daily levels of cortisol at 08.00 h or DHEA at 20.00 h greater than the 80th percentile of the daily mean. CONCLUSIONS: A subgroup of adolescents may carry a physiological risk for major depression which may be either of genetic and/or earlier psychosocial origin. PMID- 11102324 TI - Morning cortisol as a risk factor for subsequent major depressive disorder in adult women. AB - BACKGROUND: Whether individual differences in cortisol contribute to subsequent major depressive disorder (MDD) is unknown. AIMS: To determine whether premorbid levels of salivary cortisol and dehydroepiandrosterone (DHEA) were associated with subsequent MDD and how these related to psychosocial factors known to increase the risk for MDD. METHOD: Adult women (n=116) were recruited from general practices. None was currently depressed; 83 were 'psychosocially vulnerable' to MDD, 33 were not. Salivary steroids (cortisol and DHEA at 08.00 h and 20.00 h), recent life events, current mood and social support were assessed at entry. Onset of MDD was recorded during 13 months' follow-up. RESULTS: There were no associations between salivary cortisol or DHEA and recent life events or vulnerability. Twenty-eight onsets of MDD occurred during the follow-up period. This was associated with: severe adverse life events and difficulties during the follow-up period; mean morning cortisol levels at entry; and the presence of any of three vulnerability factors. CONCLUSIONS: Individual differences in morning salivary cortisol levels may represent an independent risk factor for subsequent MDD. The origin of these differences in cortisol is not yet understood. PMID- 11102325 TI - Causes and consequences of duration of untreated psychosis in schizophrenia. AB - BACKGROUND: It is unclear what determines duration of untreated psychosis (DUP) in schizophrenia and why long DUP predicts poor outcome. AIMS: First, to test the hypothesis that specific patterns of symptoms and social functioning acting before treatment prolong DUP. Second, to clarify the mechanisms linking DUP with recovery after treatment. METHOD: Two hundred and forty-eight consecutive first admissions with schizophrenia were interviewed to assess DUP, symptoms and social functioning at admission, and symptoms were re-assessed after 6-12 weeks. RESULTS: Median DUP was 12 weeks. Long DUP was predicted by poor insight, social isolation and preserved coping skills, but not by demographic factors. Even allowing for all these variables, long DUP predicted poor outcome. CONCLUSIONS: Longer DUP results partly from a pattern of symptoms and social functioning which reduces concern by the sufferer and relevant others. DUP's relationship to outcome is strongest in the initial months of psychosis. This has implications for targeting early intervention. PMID- 11102326 TI - Cognitive approach to depression and suicidal thinking in psychosis. 1. Ontogeny of post-psychotic depression. AB - BACKGROUND: Depression in schizophrenia is a rather neglected field of study, perhaps because of its confused nosological status. Three course patterns of depression in schizophrenia, including post-psychotic depression (PPD), are proposed. AIMS: We chart the ontogeny of depression and psychotic symptoms from the acute psychotic episode over a 12-month period and test the validity of the proposed course patterns. METHOD: One hundred and five patients with ICD-10 schizophrenia were followed up on five occasions over 12 months following the acute episode, taking measures of depression, positive symptoms, negative symptoms, neuroleptic exposure and side-effects. RESULTS: Depression accompanied acute psychosis in 70% of cases and remitted in line with the psychosis; 36% developed PPD without a concomitant increase in psychotic symptoms. CONCLUSIONS: The results provided support for the validity of two of the three course patterns of depression in schizophrenia, including PPD. Post-psychotic depression occurs de novo without concomitant change in positive or negative symptoms. PMID- 11102327 TI - Cognitive approach to depression and suicidal thinking in psychosis. 2. Testing the validity of a social ranking model. AB - BACKGROUND: In paper I we reported that depression in the acute stage remitted in line with the psychosis and that 36% of patients developed post-psychotic depression (PPD). AIMS: We apply our cognitive framework to PPD and chart the appraisal of self and psychosis and their link with the later emergence of PPD. METHOD: Patients with ICD-10 schizophrenia (n=105) were followed up over 12 months following the acute episode, taking measures of depression, working self concept, cognitive vulnerability, insight and appraisals of psychosis. RESULTS: Before developing PPD, these patients felt greater loss, humiliation and entrapment by their illness than those who relapsed or did not become depressed, and were more likely to see their future selves in 'lower status' roles. Upon becoming depressed, participants developed greater insight, lower self-esteem and a worsening of their appraisals of psychosis. CONCLUSIONS: Depression in psychosis arises from the individual's appraisal of psychosis and its implications for his/her perceived social identity, position and 'group fit'. Patients developing PPD feel forced to accept a subordinate role without opportunity for escape. Implications for treatment are discussed. PMID- 11102328 TI - Experience of caregiving: relatives of people experiencing a first episode of psychosis. AB - BACKGROUND: There has been relatively little research on caregivers of people experiencing their first episode of psychosis. AIMS: To investigate dimensions of caregiving and morbidity in caregivers of people with first-episode psychosis. METHOD: Caregivers of 40 people with first-episode psychosis were interviewed at home about their experience of caregiving, coping strategies and distress. RESULTS: Caregivers used emotional and practical strategies to cope with participants' negative symptoms and difficult behaviours and experienced more worry about these problems. They increased supervision when the participants displayed difficult behaviours. Twelve per cent of caregivers were suffering from psychiatric morbidity as defined by the General Health Questionnaire. Those living with the participant had more frequent visits to their general practitioner. CONCLUSIONS: At first-episode psychosis, caregivers are already having to cope with a wide range of problems and are developing coping strategies. Caregivers worried most about difficult behaviours and negative symptoms in participants. PMID- 11102329 TI - Using the Strengths and Difficulties Questionnaire (SDQ) to screen for child psychiatric disorders in a community sample. AB - BACKGROUND: Child psychiatric disorders are common and treatable, but often go undetected and therefore remain untreated. AIMS: To assess the Strengths and Difficulties Questionnaire (SDQ) as a potential means for improving the detection of child psychiatric disorders in the community. METHOD: SDQ predictions and independent psychiatric diagnoses were compared in a community sample of 7984 5- to 15-year-olds from the 1999 British Child Mental Health Survey. RESULTS: Multi informant (parents, teachers, older children) SDQs identified individuals with a psychiatric diagnosis with a specificity of 94.6% (95% Cl 94.1-95.1%) and a sensitivity of 63.3% (59.7-66.9%). The questionnaires identified over 70% of individuals with conduct, hyperactivity, depressive and some anxiety disorders, but under 50% of individuals with specific phobias, separation anxiety and eating disorders. Sensitivity was substantially poorer with single-informant rather than multi-informant SDQs. CONCLUSIONS: Community screening programmes based on multi informant SDQs could potentially increase the detection of child psychiatric disorders, thereby improving access to effective treatments. PMID- 11102330 TI - Unconsciousness, amnesia and psychiatric symptoms following road traffic accident injury. AB - BACKGROUND: Although road traffic accident injury is the most common cause of traumatic brain injury, little is known of the prevalence of psychiatric complications or the significance of unconsciousness and amnesia. AIMS: To describe amnesia and unconsciousness following a road traffic accident and to determine whether they are associated with later psychological symptoms. METHOD: Information was obtained from medical and ambulance records for 1441 consecutive attenders at an emergency department aged 17-69 who had been involved in a road traffic accident. A total of 1148 (80%) subjects completed a self-report questionnaire at baseline and were followed up at 3 months and 1 year. RESULTS: Altogether, 1.5% suffered major head (and traumatic brain) injury and 21% suffered minor head injury. Post-traumatic stress disorder (PTSD) and anxiety and depression were more common at 3 months in those who had definitely been unconscious than in those who had not, but there were no differences at 1 year. CONCLUSIONS: PTSD and other psychiatric complications are as common in those who were briefly unconscious as in those who were not. PMID- 11102331 TI - Deliberate self-harm and antidepressant drugs. Investigation of a possible link. AB - BACKGROUND: It is not clear if the frequency of deliberate self-harm (DSH) is the same in patients taking different pharmacological classes of antidepressant drugs. AIMS: To compare the frequency of DSH in patients who had been prescribed a tricyclic antidepressant (TCA) or a selective serotonin reuptake inhibitor (SSRI) prior to the DSH event. METHOD: This was a prospective study in 2776 consecutive DSH cases attending an accident and emergency department. The incidence of DSH in TCA-treated cases and SSRI-treated cases is expressed as number of DSH events per 10 000 prescriptions of each antidepressant. RESULTS: Significantly more DSH events occurred following the prescription of an SSRI than that of a TCA (P<0.001). The occurrence of DSH was highest with fluoxetine and lowest with amitriptyline. CONCLUSIONS: Merely prescribing safer-in-overdose antidepressants is unlikely to reduce the overall morbidity from DSH. PMID- 11102332 TI - Stress and psychiatric disorder in urban Rawalpindi. Community survey. AB - BACKGROUND: Recent studies in rural areas of Pakistan have yielded high prevalence rates of common mental disorders, especially among women. AIMS: To investigate emotional distress and common mental disorders in a poor urban district using the same survey method. METHOD: First-stage screening of a slum district of Rawalpindi used the Bradford Somatic Inventory. Psychiatric interviews were conducted with stratified samples using the ICD-10 research diagnostic criteria. RESULTS: On a conservative estimate, 25% of women and 10% of men suffered from anxiety and depressive disorders. Levels of emotional distress increased with age in both men and women. Women living in joint households reported more distress than those living in unitary families. Higher levels of education were associated with lower risk of common mental disorders, especially in younger women. Emotional distress was negatively correlated with socio economic variables among women. CONCLUSIONS: This study found levels of emotional distress and psychiatric morbidity in a poor district of Rawalpindi to be less than half those in a nearby rural village in the Punjab, although rates in women were still double those in men. Possible explanations are that more healthy people migrate to the cities or that urban living is more conducive to good mental health in Pakistan. PMID- 11102333 TI - Concerns over reform of the Mental Health Act. PMID- 11102334 TI - Concerns over reform of the Mental Health Act. PMID- 11102335 TI - Relevant training for case managers in severe mental illness. PMID- 11102336 TI - Neurocognitive deficits in infants of mothers with schizophrenia. PMID- 11102337 TI - Assertive outreach could be cost-effective. PMID- 11102338 TI - Need for neuropathological studies in pre-senile dementia. PMID- 11102339 TI - Disclosing the diagnosis of dementia. PMID- 11102340 TI - Ethnic differences in forensic hospitalisation. PMID- 11102341 TI - Stigmatisation: classifying drug and alcohol misuse as mental illness. PMID- 11102342 TI - Psychotherapy and developmental disability. PMID- 11102343 TI - Olanzapine and pancreatitis. PMID- 11102344 TI - Reboxetine-induced spontaneous ejaculation. PMID- 11102346 TI - A virtual cry for help? PMID- 11102348 TI - Origins of the coalescent. 1974-1982. PMID- 11102349 TI - Cost of host radiation in an RNA virus. AB - Although host radiation allows a parasite to expand its ecological niche, traits governing the infection of multiple host types can decrease fitness in the original or alternate host environments. Reasons for this reduction in fitness include slower replication due to added genetic material or modifications, fitness trade-offs across host environments, and weaker selection resulting from simultaneous adaptation to multiple habitats. We examined the consequences of host radiation using vesicular stomatitis virus (VSV) and mammalian host cells in tissue culture. Replicate populations of VSV were allowed to evolve for 100 generations on the original host (BHK cells), on either of two novel hosts (HeLa and MDCK cells), or in environments where the availability of novel hosts fluctuated in a predictable or random way. As expected, each experimental population showed a substantial fitness gain in its own environment, but those evolved on new hosts (constant or fluctuating) suffered reduced competitiveness on the original host. However, whereas evolution on one novel host negatively correlated with performance on the unselected novel host, adaptation in fluctuating environments led to fitness improvements in both novel habitats. PMID- 11102350 TI - Compensatory evolution in rifampin-resistant Escherichia coli. AB - This study examines the intrinsic fitness burden associated with RNA polymerase (rpoB) mutations conferring rifampin resistance in Escherichia coli K12 (MG1655) and explores the nature of adaptation to the costs of resistance. Among 28 independent Rif(r) mutants, the per-generation fitness burden (in the absence of rifampin) ranged from 0 to 28%, with a median of 6.4%. We detected no relationship between the magnitude of the cost and the level of resistance. Adaptation to the costs of rif resistance was studied by following serial transfer cultures for several Rif(r) mutants both in the presence of rifampin and in the absence. For cultures evolved in the absence of rifampin, single clones isolated after 200 generations were more fit than their ancestor; we saw no association between increased fitness and changes in the level of rifampin resistance; and in all cases, increased fitness was due to compensatory mutations, rather than to reversion to drug sensitivity. However, in the parallel evolution experiments in the presence of rifampin, overall levels of resistance increased as did relative fitness-for all strains save one that had an initially high level of resistance. Among the evolved clones tested, five (of seven) demonstrated increased transcription efficiency (assessed using a semiquantitative RT-PCR protocol). The implications of these results for our understanding of adaptive molecular evolution and the increasing clinical problem of antibiotic resistance are discussed. PMID- 11102351 TI - Selected amplification of the cell division genes ftsQ-ftsA-ftsZ in Escherichia coli. AB - Rapidly growing Escherichia coli is unable to divide in the presence of the antibiotic mecillinam, whose direct target is penicillin-binding protein 2 (PBP2), responsible for the elongation of the cylindrical portion of the cell wall. Division can be restored in the absence of PBP2 activity by increasing the concentration of the cell division proteins FtsQ, FtsA, and FtsZ. We tried to identify regulators of the ftsQ-ftsA-ftsZ operon among mecillinam-resistant mutants, which include strains overexpressing these genes. By insertional mutagenesis with mini-Tn10 elements, we selected for insertions that conferred mecillinam resistance. Among 15 such mutants, 7 suppressed the thermosensitivity of the ftsZ84(Ts) mutant, strongly suggesting that they had increased FtsZ activity. In all 7 cases, however, the mutants resulted from a duplication of the ftsQAZ region. These duplications seemed to result from multiple events, suggesting that no simple insertional inactivation can result in a mutant with sufficiently amplified ftsQAZ expression to confer mecillinam resistance. The structure of the duplications suggests a general method for constructing directed duplications of precise sequences. PMID- 11102352 TI - Experimental analysis of molecular events during mutational periodic selections in bacterial evolution. AB - A fundamental feature of bacterial evolution is a succession of adaptive mutational sweeps when fitter mutants take over a population. To understand the processes involved in mutational successions, Escherichia coli continuous cultures were analyzed for changes at two loci where mutations provide strong transport advantages to fitness under steady-state glucose limitation. Three separate sweeps, observed as classic periodic selection events causing a change in the frequency of neutral mutations (in fhuA causing phage T5 resistance), were identified with changes at particular loci. Two of the sweeps were associated with a reduction in the frequency of neutral mutations and the concurrent appearance of at least 13 alleles at the mgl or mlc loci, respectively. These mgl and mlc polymorphisms were of many mutational types, so were not the result of a mutator or directed mutation event. The third sweep observed was altogether distinct and involved hitchhiking between T5 resistance and advantageous mgl mutations. Moreover, the hitchhiking event coincided with an increase in mutation rates, due to the transient appearance of a strong mutator in the population. The spectrum of mgl mutations among mutator isolates was distinct and due to mutS. The mutator-associated periodic selection also resulted in mgl and fhuA polymorphism in the sweeping population. These examples of periodic selections maintained significant genotypic diversity even in a rapidly evolving culture, with no individual "winner clone" or genotype purging the population. PMID- 11102353 TI - Genetic and physical interactions between factors involved in both cell cycle progression and pre-mRNA splicing in Saccharomyces cerevisiae. AB - The PRP17/CDC40 gene of Saccharomyces cerevisiae functions in two different cellular processes: pre-mRNA splicing and cell cycle progression. The Prp17/Cdc40 protein participates in the second step of the splicing reaction and, in addition, prp17/cdc40 mutant cells held at the restrictive temperature arrest in the G2 phase of the cell cycle. Here we describe the identification of nine genes that, when mutated, show synthetic lethality with the prp17/cdc40Delta allele. Six of these encode known splicing factors: Prp8p, Slu7p, Prp16p, Prp22p, Slt11p, and U2 snRNA. The other three, SYF1, SYF2, and SYF3, represent genes also involved in cell cycle progression and in pre-mRNA splicing. Syf1p and Syf3p are highly conserved proteins containing several copies of a repeated motif, which we term RTPR. This newly defined motif is shared by proteins involved in RNA processing and represents a subfamily of the known TPR (tetratricopeptide repeat) motif. Using two-hybrid interaction screens and biochemical analysis, we show that the SYF gene products interact with each other and with four other proteins: Isy1p, Cef1p, Prp22p, and Ntc20p. We discuss the role played by these proteins in splicing and cell cycle progression. PMID- 11102354 TI - Accumulation of phosphorylated sphingoid long chain bases results in cell growth inhibition in Saccharomyces cerevisiae. AB - Sphingolipid metabolites in mammals can function as signaling molecules with cell specific functions. In Saccharomyces cerevisiae, phosphorylated long chain bases, such as dihydrosphingosine 1-phosphate and phytosphingosine 1-phosphate, have also been implicated in stress responses. To further explore the biological roles of these molecules, we created disruption mutants for LCB4, LCB5, DPL1, YSR2, YSR3, and SUR2. LCB4 and LCB5 encode kinases that phosphorylate long chain bases. DPL1 and YSR2/YSR3 are involved in degradation of the phosphorylated long chain bases. SUR2 catalyzes conversion of dihydrosphingosine to phytosphingosine. We adapted an HPLC method to measure intracellular concentrations of the phosphorylated long chain bases. Double mutants of dpl1 and ysr2 were inviable, whereas dpl1 ysr2 lcb4 triple mutants were viable. Further, growth inhibition associated with accumulated phosphorylated long chain bases was observed in the triple mutant dpl1 ysr2 lcb4 overexpressing LCB4 or LCB5. These results indicate that phosphorylated long chain bases can inhibit cell growth. Mutants defective in both YSR2 and SUR2, which accumulated dihydrosphingosine 1-phosphate only, grew poorly. The phenotypes of the ysr2 sur2 mutants were suppressed by overexpression of DPL1. Our results clearly show that elevated levels of phosphorylated long chain bases have an antiproliferative effect in yeast. PMID- 11102355 TI - Spontaneous loss of heterozygosity in diploid Saccharomyces cerevisiae cells. AB - To obtain a broad perspective of the events leading to spontaneous loss of heterozygosity (LOH), we have characterized the genetic alterations that functionally inactivated the URA3 marker hemizygously or heterozygously situated either on chromosome III or chromosome V in diploid Saccharomyces cerevisiae cells. Analysis of chromosome structure in a large number of LOH clones by pulsed field gel electrophoresis and PCR showed that chromosome loss, allelic recombination, and chromosome aberration were the major classes of genetic alterations leading to LOH. The frequencies of chromosome loss and chromosome aberration were significantly affected when the marker was located in different chromosomes, suggesting that chromosome-specific elements may affect the processes that led to these alterations. Aberrant-sized chromosomes were detected readily in approximately 8% of LOH events when the URA3 marker was placed in chromosome III. Molecular mechanisms underlying the chromosome aberrations were further investigated by studying the fate of two other genetic markers on chromosome III. Chromosome aberration caused by intrachromosomal rearrangements was predominantly due to a deletion between the MAT and HMR loci that occurred at a frequency of 3.1 x 10(-6). Another type of chromosome aberration, which occurred at a frequency slightly higher than that of the intrachromosomal deletion, appeared to be caused by interchromosomal rearrangement, including unequal crossing over between homologous chromatids and translocation with another chromosome. PMID- 11102356 TI - Decreased meiotic intergenic recombination and increased meiosis I nondisjunction in exo1 mutants of Saccharomyces cerevisiae. AB - Exonuclease I was originally identified as a 5' --> 3' deoxyribonuclease present in fractionated extracts of Schizosaccharomyces pombe and Saccharomyces cerevisiae. Genetic analysis of exo1 mutants of both yeasts revealed no major defect in meiosis, suggesting that exonuclease I is unlikely to be the primary activity that processes meiosis-specific double-strand breaks (DSBs). We report here that exo1 mutants of S. cerevisiae exhibit subtle but complex defects in meiosis. Diploids containing a homozygous deletion of EXO1 show decreased spore viability associated with an increase in meiosis I nondisjunction, while intergenic recombination is reduced about twofold. Exo1p functions in the same pathway as Msh5p for intergenic recombination. The length of heteroduplex tracts within the HIS4 gene is unaffected by the exo1 mutation. These results suggest that Exo1p is unlikely to play a major role in processing DSBs to form single stranded tails at HIS4, but instead appears to promote crossing over to ensure disjunction of homologous chromosomes. In addition, our data indicate that exonuclease I may have a minor role in the correction of large DNA mismatches that occur in heteroduplex DNA during meiotic recombination at the HIS4 locus. PMID- 11102358 TI - Extragenic suppressors of the nimX2(cdc2) mutation of Aspergillus nidulans affect nuclear division, septation and conidiation. AB - The Aspergillus nidulans NIMX(CDC2) protein kinase has been shown to be required for both the G(2)/M and G(1)/S transitions, and recent evidence has implicated a role for NIMX(CDC2) in septation and conidiation. While much is understood of its G(2)/M function, little is known about the functions of NIMX(CDC2) during G(1)/S, septation, and conidiophore development. In an attempt to better understand how NIMX(CDC2) is involved in these processes, we have isolated four extragenic suppressors of the A. nidulans nimX2(cdc2) temperature-sensitive mutation. Mutation of these suppressor genes, designated snxA-snxD for suppressor of nimX, affects nuclear division, septation, and conidiation. The cold-sensitive snxA1 mutation leads to arrest of nuclear division during G(1) or early S. snxB1 causes hyperseptation in the hyphae and sensitivity to hydroxyurea, while snxC1 causes septation in the conidiophore stalk and aberrant conidiophore structure. snxD1 leads to slight septation defects and hydroxyurea sensitivity. The additional phenotypes that result from the suppressor mutations provide genetic evidence that NIMX(CDC2) affects septation and conidiation in addition to nuclear division, and cloning and biochemical analysis of these will allow a better understanding of the role of NIMX(CDC2) in these processes. PMID- 11102357 TI - The Aspergillus nidulans xprF gene encodes a hexokinase-like protein involved in the regulation of extracellular proteases. AB - The extracellular proteases of Aspergillus nidulans are produced in response to limitation of carbon, nitrogen, or sulfur, even in the absence of exogenous protein. Mutations in the A. nidulans xprF and xprG genes have been shown to result in elevated levels of extracellular protease in response to carbon limitation. The xprF gene was isolated and sequence analysis indicates that it encodes a 615-amino-acid protein, which represents a new type of fungal hexokinase or hexokinase-like protein. In addition to their catalytic role, hexokinases are thought to be involved in triggering carbon catabolite repression. Sequence analysis of the xprF1 and xprF2 alleles showed that both alleles contain nonsense mutations. No loss of glucose or fructose phosphorylating activity was detected in xprF1 or xprF2 mutants. There are two possible explanations for this observation: (1) the xprF gene may encode a minor hexokinase or (2) the xprF gene may encode a protein with no hexose phosphorylating activity. Genetic evidence suggests that the xprF and xprG genes are involved in the same regulatory pathway. Support for this hypothesis was provided by the identification of a new class of xprG(-) mutation that suppresses the xprF1 mutation and results in a protease-deficient phenotype. PMID- 11102359 TI - Scooter, a new active transposon in Schizophyllum commune, has disrupted two genes regulating signal transduction. AB - Two copies of scooter, a DNA-mediated transposon in the basidiomycetous fungus Schizophyllum commune, were characterized. Scooter is the first transposon isolated from S. commune. Scooter creates 8-bp target site duplications, comparable to members of the hAT superfamily, and has 32-bp terminal inverted repeats. Both copies of scooter are nonautonomous elements capable of movement. Southern blot hybridizations show that scooter-related sequences are present in all S. commune strains tested. Scooter-1 was identified initially as an insertion in the Bbeta2 pheromone receptor gene, bbr2, leading to a partial defect in mating. Scooter-2 spontaneously disrupted a gene to produce the frequently occurring morphological mutant phenotype known as thin. The scooter-2 insert permitted cloning of the disrupted gene, thn1, which encodes a putative regulator of G protein signaling (RGS) protein. Spontaneous insertion of scooter into genes with identifiable mutant phenotypes constitutes the first evidence of active transposition of a DNA-mediated transposon in a basidiomycete. PMID- 11102360 TI - Null mutations in the lin-31 gene indicate two functions during Caenorhabditis elegans vulval development. AB - The lin-31 gene is required for the proper specification of vulval cell fates in the nematode Caenorhabditis elegans and encodes a member of the winged-helix family of transcription factors. Members of this important family have been identified in many organisms and are known to bind specific DNA targets involved in a variety of developmental processes. DNA sequencing of 13 lin-31 alleles revealed six nonsense mutations and two missense mutations within the DNA-binding domain, plus three deletions, one transposon insertion, and one frameshift mutation that all cause large-scale disruptions in the gene. The missense mutations are amino acid substitutions in the DNA-binding domain and probably disrupt interactions of the LIN-31 transcription factor with its DNA target. In addition, detailed phenotypic analysis of all 19 alleles showed similar penetrance for several characteristics examined. From our analysis we conclude: (1) the null phenotype of lin-31 is the phenotype displayed by almost all of the existing alleles, (2) the DNA-binding domain plays a critical role in LIN-31 function, and (3) direct screens for multivulva and vulvaless mutants will probably yield only null (or strong) alleles of lin-31. PMID- 11102361 TI - The Caenorhabditis elegans dosage compensation machinery is recruited to X chromosome DNA attached to an autosome. AB - The dosage compensation machinery of Caenorhabditis elegans is targeted specifically to the X chromosomes of hermaphrodites (XX) to reduce gene expression by half. Many of the trans-acting factors that direct the dosage compensation machinery to X have been identified, but none of the proposed cis acting X chromosome-recognition elements needed to recruit dosage compensation components have been found. To study X chromosome recognition, we explored whether portions of an X chromosome attached to an autosome are competent to bind the C. elegans dosage compensation complex (DCC). To do so, we devised a three dimensional in situ approach that allowed us to compare the volume, position, and number of chromosomal and subchromosomal bodies bound by the dosage compensation machinery in wild-type XX nuclei and XX nuclei carrying an X duplication. The dosage compensation complex was found to associate with a duplication of the right 30% of X, but the complex did not spread onto adjacent autosomal sequences. This result indicates that all the information required to specify X chromosome identity resides on the duplication and that the dosage compensation machinery can localize to a site distinct from the full-length hermaphrodite X chromosome. In contrast, smaller duplications of other regions of X appeared to not support localization of the DCC. In a separate effort to identify cis-acting X recognition elements, we used a computational approach to analyze genomic DNA sequences for the presence of short motifs that were abundant and overrepresented on X relative to autosomes. Fourteen families of X-enriched motifs were discovered and mapped onto the X chromosome. PMID- 11102363 TI - A test for epistasis among induced mutations in Caenorhabditis elegans. AB - Synergistic epistasis, in which deleterious mutations tend to magnify each other's effects, is a necessary component of the mutational deterministic hypothesis for the maintenance of sexual production. We tested for epistasis for life-history traits in the soil nematode Caenorhabditis elegans by inducing mutations in two genetic backgrounds: a wild-type strain and a set of genetically loaded lines that contain large numbers of independent mildly detrimental mutations. There was no significant difference between the effect of new mutations on the wild-type background and the genetically loaded background for four out of five fitness correlates. In these four cases, the maximum level of epistasis compatible with the data was very low. The fifth trait, late productivity, is not likely to be an important component of fitness. This suggests either that specific environmental conditions are required to cause epistasis or that synergistic epistasis is not a general phenomenon. We also suggest a new mechanism by which deleterious mutations may provide an advantage to sexual reproduction under low selection coefficients. PMID- 11102362 TI - spe-29 encodes a small predicted membrane protein required for the initiation of sperm activation in Caenorhabditis elegans. AB - Caenorhabditis elegans spermatids complete a dramatic morphogenesis to crawling spermatozoa in the absence of an actin- or tubulin-based cytoskeleton and without synthesizing new gene products. Mutations in three genes (spe-8, spe-12, and spe 27) prevent the initiation of this morphogenesis, termed activation. Males with mutations in any of these genes are fertile. By contrast, mutant hermaphrodites are self-sterile when unmated due to a failure in spermatid activation. Intriguingly, mutant hermaphrodites form functional spermatozoa and become self fertile upon mating, suggesting that spermatids can be activated by male seminal fluid. Here we describe a mutation in a fourth gene, spe-29, which mimics the phenotype of spe-8, spe-12, and spe-27 mutants. spe-29 sperm are defective in the initiation of hermaphrodite sperm activation, yet they maintain the ability to complete the morphogenetic rearrangements that follow. Mutant alleles of spe-12, spe-27, and spe-29 exhibit genetic interactions that suggest that the wild-type products of these genes function in a common signaling pathway to initiate sperm activation. We have identified the spe-29 gene, which is expressed specifically in the sperm-producing germ line and is predicted to encode a small, novel transmembrane protein. PMID- 11102364 TI - A role for RIC-8 (Synembryn) and GOA-1 (G(o)alpha) in regulating a subset of centrosome movements during early embryogenesis in Caenorhabditis elegans. AB - RIC-8 (synembryn) and GOA-1 (G(o)alpha) are key components of a signaling network that regulates neurotransmitter secretion in Caenorhabditis elegans. Here we show that ric-8 and goa-1 reduction of function mutants exhibit partial embryonic lethality. Through Nomarski analysis we show that goa-1 and ric-8 mutant embryos exhibit defects in multiple events that involve centrosomes, including one-cell posterior centrosome rocking, P(1) centrosome flattening, mitotic spindle alignment, and nuclear migration. In ric-8 reduction of function backgrounds, the embryonic lethality, spindle misalignments and delayed nuclear migration are strongly enhanced by a 50% reduction in maternal goa-1 gene dosage. Several other microfilament- and microtubule-mediated events, as well as overall embryonic polarity, appear unperturbed in the mutants. In addition, our results suggest that RIC-8 and GOA-1 do not have roles in centrosome replication, in the diametric movements of daughter centrosomes along the nuclear membrane, or in the extension of microtubules from centrosomes. Through immunostaining we show that GOA-1 (G(o)alpha) localizes to cell cortices as well as near centrosomes. Our results demonstrate that two components of a neuronal signal transduction pathway also play a role in centrosome movements during early embryogenesis. PMID- 11102365 TI - Transposons but not retrotransposons are located preferentially in regions of high recombination rate in Caenorhabditis elegans. AB - We analyzed the distribution of transposable elements (TEs: transposons, LTR retrotransposons, and non-LTR retrotransposons) in the chromosomes of the nematode Caenorhabditis elegans. The density of transposons (DNA-based elements) along the chromosomes was found to be positively correlated with recombination rate, but this relationship was not observed for LTR or non-LTR retrotransposons (RNA-based elements). Gene (coding region) density is higher in regions of low recombination rate. However, the lower TE density in these regions is not due to the counterselection of TE insertions within exons since the same positive correlation between TE density and recombination rate was found in noncoding regions (both in introns and intergenic DNA). These data are not compatible with a global model of selection acting against TE insertions, for which an accumulation of elements in regions of reduced recombination is expected. We also found no evidence for a stronger selection against TE insertions on the X chromosome compared to the autosomes. The difference in distribution of the DNA and RNA-based elements along the chromosomes in relation to recombination rate can be explained by differences in the transposition processes. PMID- 11102366 TI - Embryonic morphogenesis in Caenorhabditis elegans integrates the activity of LET 502 Rho-binding kinase, MEL-11 myosin phosphatase, DAF-2 insulin receptor and FEM 2 PP2c phosphatase. AB - let-502 rho-binding kinase and mel-11 myosin phosphatase regulate Caenorhabditis elegans embryonic morphogenesis. Genetic analysis presented here establishes the following modes of let-502 action: (i) loss of only maternal let-502 results in abnormal early cleavages, (ii) loss of both zygotic and maternal let-502 causes elongation defects, and (iii) loss of only zygotic let-502 results in sterility. The morphogenetic function of let-502 and mel-11 is apparently redundant with another pathway since elimination of these two genes resulted in progeny that underwent near-normal elongation. Triple mutant analysis indicated that unc-73 (Rho/Rac guanine exchange factor) and mlc-4 (myosin light chain) act in parallel to or downstream of let-502/mel-11. In contrast mig-2 (Rho/Rac), daf-2 (insulin receptor), and age-1 (PI3 kinase) act within the let-502/mel-11 pathway. Mutations in the sex-determination gene fem-2, which encodes a PP2c phosphatase (unrelated to the MEL-11 phosphatase), enhanced mutations of let-502 and suppressed those of mel-11. fem-2's elongation function appears to be independent of its role in sexual identity since the sex-determination genes fem-1, fem-3, tra-1, and tra-3 had no effect on mel-11 or let-502. By itself, fem-2 affects morphogenesis with low penetrance. fem-2 blocked the near-normal elongation of let-502; mel-11 indicating that fem-2 acts in a parallel elongation pathway. The action of two redundant pathways likely ensures accurate elongation of the C. elegans embryo. PMID- 11102367 TI - Mutations affecting the development of the peripheral nervous system in Drosophila: a molecular screen for novel proteins. AB - In our quest for novel genes required for the development of the embryonic peripheral nervous system (PNS), we have performed three genetic screens using MAb 22C10 as a marker of terminally differentiated neurons. A total of 66 essential genes required for normal PNS development were identified, including 49 novel genes. To obtain information about the molecular nature of these genes, we decided to complement our genetic screens with a molecular screen. From transposon-tagged mutations identified on the basis of their phenotype in the PNS we selected 31 P-element strains representing 26 complementation groups on the second and third chromosomes to clone and sequence the corresponding genes. We used plasmid rescue to isolate and sequence 51 genomic fragments flanking the sites of these P-element insertions. Database searches using sequences derived from the ends of plasmid rescues allowed us to assign genes to one of four classes: (1) previously characterized genes (11), (2) first mutations in cloned genes (1), (3) P-element insertions in genes that were identified, but not characterized molecularly (1), and (4) novel genes (13). Here, we report the cloning, sequence, Northern analysis, and the embryonic expression pattern of candidate cDNAs for 10 genes: astray, chrowded, dalmatian, gluon, hoi-polloi, melted, pebble, skittles, sticky ch1, and vegetable. This study allows us to draw conclusions about the identity of proteins required for the development of the nervous system in Drosophila and provides an example of a molecular approach to characterize en masse transposon-tagged mutations identified in genetic screens. PMID- 11102368 TI - Specific genetic interference with behavioral rhythms in Drosophila by expression of inverted repeats. AB - We describe a new experimental technique that allows for a tissue-specific reduction of gene activity in the Drosophila nervous system. On the basis of the observation that certain gene functions can be ubiquitously blocked by injecting double-stranded RNA into Drosophila embryos, we employed a method to interfere with an individual gene function permanently in a predetermined cell type. This was achieved by the formation of an inverted-repeat RNA sequence in the tissue of interest under control of the GAL4/UAS binary expression system. As an example, we show that inverted-repeat-mediated interference with the period gene produces a hypomorphic period phenotype. A selective decrease of period RNA appears to be a component of the cellular response. PMID- 11102369 TI - The oxen gene of Drosophila encodes a homolog of subunit 9 of yeast ubiquinol cytochrome c oxidoreductase complex: evidence for modulation of gene expression in response to mitochondrial activity. AB - A P-element insertion in the oxen gene, ox(1), has been isolated in a search for modifiers of white gene expression. The mutation preferentially exerts a negative dosage effect upon the expression of three genes encoding ABC transporters involved in pigment precursor transport, white, brown, and scarlet. A precise excision of the P element reverts the mutant phenotype. Five different transcription units were identified around the insertion site. To distinguish a transcript responsible for the mutant phenotype, a set of deletions within the oxen region was generated. Analysis of gene expression within the oxen region in the case of deletions as well as generation of transgenic flies allowed us to identify the transcript responsible for oxen function. It encodes a 6.6-kD homolog of mitochondrial ubiquinol cytochrome c oxidoreductase (QCR9), subunit 9 of the bc(1) complex in yeast. In addition to white, brown, and scarlet, oxen regulates the expression of three of seven tested genes. Thus, our data provide additional evidence for a cellular response to changes in mitochondrial function. The oxen mutation provides a model for the genetic analysis in multicellular organisms of the effect of mitochondrial activity on nuclear gene expression. PMID- 11102371 TI - A selective sweep associated with a recent gene transposition in Drosophila miranda. AB - In Drosophila miranda, a chromosome fusion between the Y chromosome and the autosome corresponding to Muller's element C has created a new sex chromosome system. The chromosome attached to the ancestral Y chromosome is transmitted paternally and hence is not exposed to crossing over. This chromosome, conventionally called the neo-Y, and the homologous neo-X chromosome display many properties of evolving sex chromosomes. We report here the transposition of the exuperantia1 (exu1) locus from a neo-sex chromosome to the ancestral X chromosome of D. miranda. Exu1 is known to have several critical developmental functions, including a male-specific role in spermatogenesis. The ancestral location of exu1 is conserved in the sibling species of D. miranda, as well as in a more distantly related species. The transposition of exu1 can be interpreted as an adaptive fixation, driven by a selective advantage conferred by its effect on dosage compensation. This explanation is supported by the pattern of within-species sequence variation at exu1 and the nearby exu2 locus. The implications of this phenomenon for genome evolution are discussed. PMID- 11102370 TI - Extensive amino acid polymorphism at the pgm locus is consistent with adaptive protein evolution in Drosophila melanogaster. AB - PGM plays a central role in the glycolytic pathway at the branch point leading to glycogen metabolism and is highly polymorphic in allozyme studies of many species. We have characterized the nucleotide diversity across the Pgm gene in Drosophila melanogaster and D. simulans to investigate the role that protein polymorphism plays at this crucial metabolic branch point shared with several other enzymes. Although D. melanogaster and D. simulans share common allozyme mobility alleles, we find these allozymes are the result of many different amino acid changes at the nucleotide level. In addition, specific allozyme classes within species contain several amino acid changes, which may explain the absence of latitudinal clines for PGM allozyme alleles, the lack of association of PGM allozymes with the cosmopolitan In(3L)P inversion, and the failure to detect differences between PGM allozymes in functional studies. We find a significant excess of amino acid polymorphisms within D. melanogaster when compared to the complete absence of fixed replacements with D. simulans. There is also strong linkage disequilibrium across the 2354 bp of the Pgm locus, which may be explained by a specific amino acid haplotype that is high in frequency yet contains an excess of singleton polymorphisms. Like G6pd, Pgm shows strong evidence for a branch point enzyme that exhibits adaptive protein evolution. PMID- 11102372 TI - An enhancer trap screen for ecdysone-inducible genes required for Drosophila adult leg morphogenesis. AB - Although extensive studies of Drosophila imaginal disc development have focused on proliferation and patterning, relatively little is known about how the patterned imaginal discs are transformed into adult structures during metamorphosis. Studies focused primarily on leg development have shown that this remarkable transformation is coordinated by pulses of the steroid hormone ecdysone and requires the function of ecdysone-inducible transcription factors as well as proteases and components of the contractile cytoskeleton and adherens junctions. Here, we describe a genetic screen aimed at expanding our understanding of the hormonal regulation of Drosophila adult leg morphogenesis. We screened 1300 lethal P-element enhancer trap insertions on the second chromosome for a series of sequential parameters including pupal lethality, defects in leg morphogenesis, and ecdysone-induced lacZ reporter gene expression. From this screen we identified four mutations, one of which corresponds to bancal, which encodes the Drosophila homolog of hnRNP K. We also identified vulcan, which encodes a protein that shares sequence similarity with a family of rat SAPAP proteins. Both bancal and vulcan are inducible by ecdysone, thus linking the hormone signal with leg morphogenesis. This screen provides new directions for understanding the hormonal regulation of leg development during Drosophila metamorphosis. PMID- 11102373 TI - A directed mutagenesis screen in Drosophila melanogaster reveals new mutants that influence hedgehog signaling. AB - The Hedgehog signaling pathway has been recognized as essential for patterning processes in development of metazoan animal species. The signaling pathway is, however, not entirely understood. To start to address this problem, we set out to isolate new mutations that influence Hedgehog signaling. We performed a mutagenesis screen for mutations that dominantly suppress Hedgehog overexpression phenotypes in the Drosophila melanogaster wing. We isolated four mutations that influence Hedgehog signaling. These were analyzed in the amenable wing system using genetic and molecular techniques. One of these four mutations affects the stability of the Hedgehog expression domain boundary, also known as the organizer in the developing wing. Another mutation affects a possible Hedgehog autoregulation mechanism, which stabilizes the same boundary. PMID- 11102374 TI - Genetic analysis of the Drosophila DNAprim gene. The function of the 60-kd primase subunit of DNA polymerase opposes the fat facets signaling pathway in the developing eye. AB - The Drosophila DNAprim gene encodes the large subunit (60 kD) of DNA primase, the part of DNA polymerase alpha that synthesizes RNA primers during DNA replication. The precise function of the 60-kD subunit is unknown. In a mutagenesis screen for suppressors of the fat facets (faf) mutant eye phenotype, we identified mutations in DNAprim. The faf gene encodes a deubiquitinating enzyme required specifically for patterning the compound eye. The DNA sequences of four DNAprim alleles were determined and these define essential protein domains. We show that while flies lacking DNAprim activity are lethal, flies with reduced DNAprim activity display morphological defects in their eyes, and unlike faf mutants, cell cycle abnormalities in larval eye discs. Mechanisms by which DNA primase levels might influence the faf-dependent cell communication pathway are discussed. PMID- 11102375 TI - Toward a physical map of Drosophila buzzatii. Use of randomly amplified polymorphic dna polymorphisms and sequence-tagged site landmarks. AB - We present a physical map based on RAPD polymorphic fragments and sequence-tagged sites (STSs) for the repleta group species Drosophila buzzatii. One hundred forty four RAPD markers have been used as probes for in situ hybridization to the polytene chromosomes, and positive results allowing the precise localization of 108 RAPDs were obtained. Of these, 73 behave as effectively unique markers for physical map construction, and in 9 additional cases the probes gave two hybridization signals, each on a different chromosome. Most markers (68%) are located on chromosomes 2 and 4, which partially agree with previous estimates on the distribution of genetic variation over chromosomes. One RAPD maps close to the proximal breakpoint of inversion 2z(3) but is not included within the inverted fragment. However, it was possible to conclude from this RAPD that the distal breakpoint of 2z(3) had previously been wrongly assigned. A total of 39 cytologically mapped RAPDs were converted to STSs and yielded an aggregate sequence of 28,431 bp. Thirty-six RAPDs (25%) did not produce any detectable hybridization signal, and we obtained the DNA sequence from three of them. Further prospects toward obtaining a more developed genetic map than the one currently available for D. buzzatii are discussed. PMID- 11102376 TI - The tricornered gene, which is required for the integrity of epidermal cell extensions, encodes the Drosophila nuclear DBF2-related kinase. AB - During their differentiation epidermal cells of Drosophila form a rich variety of polarized structures. These include the epidermal hairs that decorate much of the adult cuticular surface, the shafts of the bristle sense organs, the lateral extensions of the arista, and the larval denticles. These cuticular structures are produced by cytoskeletal-mediated outgrowths of epidermal cells. Mutations in the tricornered gene result in the splitting or branching of all of these structures. Thus, tricornered function appears to be important for maintaining the integrity of the outgrowths. tricornered mutations however do not have major effects on the growth or shape of these cellular extensions. Inhibiting actin polymerization in differentiating cells by cytochalasin D or latrunculin A treatment also induces the splitting of hairs and bristles, suggesting that the actin cytoskeleton might be a target of tricornered. However, the drugs also result in short, fat, and occasionally malformed hairs and bristles. The data suggest that the function of the actin cytoskeleton is important for maintaining the integrity of cellular extensions as well as their growth and shape. Thus, if tricornered causes the splitting of cellular extensions by interacting with the actin cytoskeleton it likely does so in a subtle way. Consistent with this possibility we found that a weak tricornered mutant is hypersensitive to cytochalasin D. We have cloned the tricornered gene and found that it encodes the Drosophila NDR kinase. This is a conserved ser/thr protein kinase found in Caenorhabditis elegans and humans that is related to a number of kinases that have been found to be important in controlling cell structure and proliferation. PMID- 11102377 TI - In vivo Structure/Function analysis of the Drosophila fat facets deubiquitinating enzyme gene. AB - The Drosophila Fat facets protein is a deubiquitinating enzyme required for patterning the developing compound eye. Ubiquitin, a 76-amino-acid polypeptide, serves as a tag to direct proteins to the proteasome, a protein degradation complex. Deubiquitinating enzymes are a large group of proteins that cleave ubiquitin-protein bonds. Fat facets belongs to a class of deubiquitinating enzymes called Ubps that share a conserved catalytic domain. Fat facets is unique among them in its large size and also because Fat facets is thought to deubiquitinate a specific substrate thereby preventing its proteolysis. Here we asked which portions of the Fat facets protein are essential for its function. P element constructs that express partial Fat facets proteins were tested for function. In addition, the DNA sequences of 12 mutant fat facets alleles were determined. Finally, regions of amino acid sequence similarity in 18 Drosophila Ubps revealed by the Genome Project were identified. The results indicate functions for specific conserved amino acids in the catalytic region of Fat facets and also indicate that regions of the protein both N- and C-terminal to the catalytic region are required for Fat facets function. PMID- 11102378 TI - Linkage disequilibria and the site frequency spectra in the su(s) and su(w(a)) regions of the Drosophila melanogaster X chromosome. AB - Over the last decade, surveys of DNA sequence variation in natural populations of several Drosophila species and other taxa have established that polymorphism is reduced in genomic regions characterized by low rates of crossing over per physical length. Parallel studies have also established that divergence between species is not reduced in these same genomic regions, thus eliminating explanations that rely on a correlation between the rates of mutation and crossing over. Several theoretical models (directional hitchhiking, background selection, and random environment) have been proposed as population genetic explanations. In this study samples from an African population (n = 50) and a European population (n = 51) were surveyed at the su(s) (1955 bp) and su(w(a)) (3213 bp) loci for DNA sequence polymorphism, utilizing a stratified SSCP/DNA sequencing protocol. These loci are located near the telomere of the X chromosome, in a region of reduced crossing over per physical length, and exhibit a significant reduction in DNA sequence polymorphism. Unlike most previously surveyed, these loci reveal substantial skews toward rare site frequencies, consistent with the predictions of directional hitchhiking and random environment models and inconsistent with the general predictions of the background selection model (or neutral theory). No evidence for excess geographic differentiation at these loci is observed. Although linkage disequilibrium is observed between closely linked sites within these loci, many recombination events in the genealogy of the sampled alleles can be inferred and the genomic scale of linkage disequilibrium, measured in base pairs between sites, is the same as that observed for loci in regions of normal crossing over. We conclude that gene conversion must be high in these regions of low crossing over. PMID- 11102379 TI - Evolution of dosage compensation in Diptera: the gene maleless implements dosage compensation in Drosophila (Brachycera suborder) but its homolog in Sciara (Nematocera suborder) appears to play no role in dosage compensation. AB - In Drosophila melanogaster and in Sciara ocellaris dosage compensation occurs by hypertranscription of the single male X chromosome. This article reports the cloning and characterization in S. ocellaris of the gene homologous to maleless (mle) of D. melanogaster, which implements dosage compensation. The Sciara mle gene produces a single transcript, encoding a helicase, which is present in both male and female larvae and adults and in testes and ovaries. Both Sciara and Drosophila MLE proteins are highly conserved. The affinity-purified antibody to D. melanogaster MLE recognizes the S. ocellaris MLE protein. In contrast to Drosophila polytene chromosomes, where MLE is preferentially associated with the male X chromosome, in Sciara MLE is found associated with all chromosomes. Anti MLE staining of Drosophila postblastoderm male embryos revealed a single nuclear dot, whereas Sciara male and female embryos present multiple intranuclear staining spots. This expression pattern in Sciara is also observed before blastoderm stage, when dosage compensation is not yet set up. The affinity purified antibodies against D. melanogaster MSL1, MSL3, and MOF proteins involved in dosage compensation also revealed no differences in the staining pattern between the X chromosome and the autosomes in both Sciara males and females. These results lead us to propose that different proteins in Drosophila and Sciara would implement dosage compensation. PMID- 11102380 TI - Artificial and epigenetic regulation of the I factor, a nonviral retrotransposon of Drosophila melanogaster. AB - The I factor (IF) is a LINE-like transposable element from Drosophila melanogaster. IF is silenced in most strains, but under special circumstances its transposition can be induced and correlates with the appearance of a syndrome of female sterility called hybrid dysgenesis. To elucidate the relationship between IF expression and female sterility, different transgenic antisense and/or sense RNAs homologous to the IF ORF1 have been expressed. Increasing the transgene copy number decreases both the expression of an IF-lacZ fusion and the intensity of the female sterile phenotype, demonstrating that IF expression is correlated with sterility. Some transgenes, however, exert their repressive abilities not only through a copy number-dependent zygotic effect, but also through additional maternal and paternal effects that may be induced at the DNA and/or RNA level. Properties of the maternal effect have been detailed: (1) it represses hybrid dysgenesis more efficiently than does the paternal effect; (2) its efficacy increases with both the transgene copy number and the aging of sterile females; (3) it accumulates slowly over generations after the transgene has been established; and (4) it is maintained for at least two generations after transgene removal. Conversely, the paternal effect increases only with female aging. The last two properties of the maternal effect and the genuine existence of a paternal effect argue for the occurrence, in the IF regulation pathway, of a cellular memory transmitted through mitosis, as well as through male and female meiosis, and akin to epigenetic phenomena. PMID- 11102381 TI - Molecular population genetics of male accessory gland proteins in Drosophila. AB - Drosophila seminal proteins have an unusually high rate of molecular sequence evolution, suggesting either a high rate of neutral substitution or rapid adaptive evolution. To further quantify patterns of polymorphism and divergence in genes encoding seminal proteins, also called accessory gland proteins (Acp's), we conducted a sequencing survey of 10 Acp genes in samples of Drosophila melanogaster and D. simulans (Acp29AB, Acp32CD, Acp33A, Acp36DE, Acp53Ea, Acp62F, Acp63F, Acp76A, Acp95EF, and Acp98AB). Mean heterozygosity at replacement sites in D. simulans was 0.0074 for Acp genes and 0.0013 for a set of 19 non-Acp genes, and mean melanogaster-simulans divergence at replacement sites was 0.0497 for Acp genes and 0.0107 at non-Acp genes. The elevated divergence of Acp genes is thus accompanied by elevated within-species polymorphism. In addition to the already reported departures of Acp26A, Acp29AB, and Acp70A from neutrality, our data reject neutrality at Acp29AB and Acp36DE in the direction of excess replacements in interspecific comparisons. PMID- 11102382 TI - The Ketel gene encodes a Drosophila homologue of importin-beta. AB - The Drosophila melanogaster Ketel gene was identified via the Ketel(D) dominant female sterile mutations and their ketel(r) revertant alleles that are recessive zygotic lethals. The maternally acting Ketel(D) mutations inhibit cleavage nuclei formation. We cloned the Ketel gene on the basis of a common breakpoint in 38E1. 2-3 in four ketel(r) alleles. The Ketel(+) transgenes rescue ketel(r)-associated zygotic lethality and slightly reduce Ketel(D)-associated dominant female sterility. Ketel is a single copy gene. It is transcribed to a single 3.6-kb mRNA, predicted to encode the 97-kD Ketel protein. The 884-amino-acid sequence of Ketel is 60% identical and 78% similar to that of human importin-beta, the nuclear import receptor for proteins with a classical NLS. Indeed, Ketel supports import of appropriately designed substrates into nuclei of digitonin permeabilized HeLa cells. As shown by a polyclonal anti-Ketel antibody, nurse cells synthesize and transfer Ketel protein into the oocyte cytoplasm from stage 11 of oogenesis. In cleavage embryos the Ketel protein is cytoplasmic. The Ketel gene appears to be ubiquitously expressed in embryonic cells. Western blot analysis revealed that the Ketel gene is not expressed in several larval cell types of late third instar larvae. PMID- 11102383 TI - The Ketel(D) dominant-negative mutations identify maternal function of the Drosophila importin-beta gene required for cleavage nuclei formation. AB - The Ketel(D) dominant female-sterile mutations and their ketel(r) revertant alleles identify the Ketel gene, which encodes the importin-beta (karyopherin beta) homologue of Drosophila melanogaster. Embryogenesis does not commence in the Ketel(D) eggs deposited by the Ketel(D)/+ females due to failure of cleavage nuclei formation. When injected into wild-type cleavage embryos, cytoplasm of the Ketel(D) eggs does not inhibit nuclear protein import but prevents cleavage nuclei formation following mitosis. The Ketel(+) transgenes slightly reduce effects of the Ketel(D) mutations. The paternally derived Ketel(D) alleles act as recessive zygotic lethal mutations: the Ketel(D)/- hemizygotes, like the ketel(r)/ketel(r) and the ketel(r)/- zygotes, perish during second larval instar. The Ketel maternal dowry supports their short life. The Ketel(D)-related defects originate most likely following association of the Ketel(D)-encoded mutant molecules with a maternally provided partner. As in the Ketel(D) eggs, embryogenesis does not commence in eggs of germline chimeras with ketel(r)/- germline cells and normal soma, underlining the dominant-negative nature of the Ketel(D) mutations. The ketel(r) homozygous clones are fully viable in the follicle epithelium in wings and tergites. The Ketel gene is not expressed in most larval tissues, as revealed by the expression pattern of a Ketel promoter lacZ reporter gene. PMID- 11102385 TI - Heteroplasmy in the mtDNA control region of sturgeon (Acipenser, Huso and Scaphirhynchus). AB - Data from 1238 fishes from 19 sturgeon species and 1 paddlefish were used to analyze heteroplasmy in sturgeon. Lengths of central repeat units ranged from 74 to 83 bp among sturgeon species. No repeat sequence was found in the paddlefish, Polyodon spathula. A general feature of the repeat units was the presence of termination associated sequence (TAS) motifs. About 50% of 138 interspecific mutations observed among the D-loop sequences are located 10 bp down- and upstream from these TAS motifs. Interestingly, most homoplasmic species showed deletions upstream to the TAS motifs, whereas deletions downstream to the TAS motifs observed in two species do not seem to preclude heteroplasmy. Calculations of secondary structures and thermal stabilities of repeat units showed DeltaG values for all heteroplasmic species to be <-8 and for most homoplasmic species DeltaG value to be >-8. Most heteroplasmic fishes had two and/or three repeat units. No homoplasmic sturgeon with >2 repeat units were observed. Molecular phylogeny based on the entire cytochrome b showed that heteroplasmy probably resulted from a single evolutionary event. Our data demonstrate that heteroplasmy is present in most sturgeon species and suggest that the thermal stability of the secondary structure of the repeat unit in combination with mutations downstream of the TAS sequences influences heteroplasmy. PMID- 11102384 TI - The population genetics of the origin and divergence of the Drosophila simulans complex species. AB - The origins and divergence of Drosophila simulans and close relatives D. mauritiana and D. sechellia were examined using the patterns of DNA sequence variation found within and between species at 14 different genes. D. sechellia consistently revealed low levels of polymorphism, and genes from D. sechellia have accumulated mutations at a rate that is approximately 50% higher than the same genes from D. simulans. At synonymous sites, D. sechellia has experienced a significant excess of unpreferred codon substitutions. Together these observations suggest that D. sechellia has had a reduced effective population size for some time, and that it is accumulating slightly deleterious mutations as a result. D. simulans and D. mauritiana are both highly polymorphic and the two species share many polymorphisms, probably since the time of common ancestry. A simple isolation speciation model, with zero gene flow following incipient species separation, was fitted to both the simulans/mauritiana divergence and the simulans/sechellia divergence. In both cases the model fit the data quite well, and the analyses revealed little evidence of gene flow between the species. The exception is one gene copy at one locus in D. sechellia, which closely resembled other D. simulans sequences. The overall picture is of two allopatric speciation events that occurred quite near one another in time. PMID- 11102386 TI - Sequence variation at two eosinophil-associated ribonuclease loci in humans. AB - Host defense against invading pathogens is of great importance to the survival of higher organisms. We have been studying the evolution of mammalian eosinophil associated ribonucleases (EARs), which are members of the ribonuclease A superfamily with known antipathogen activities. Earlier studies showed that positive selection promoted rapid diversification of paralogous EAR genes in both primates and rodents. Intraspecifically, however, it is unknown whether these genes also have divergent alleles. The recent discovery that the gene repertoire of the EAR family is much larger in rodents than in primates has led us to consider the possibility that primates maintain a large number of polymorphic alleles to compensate for a smaller gene repertoire. Here we present sequences of 2417 nucleotides at the two EAR loci, the eosinophil-derived neurotoxin (EDN, RNase 2) and eosinophil cationic protein (ECP, RNase 3), from >50 human individuals. Our data demonstrate that the nucleotide diversities (0.06-0.11%) at these loci are typical for human nuclear genes, thus permitting us to reject this polymorphism hypothesis. No significant departure from neutrality is noted and no signs of overdominant selection are observed. Similar patterns were observed in a preliminary study of chimpanzees. In summary, our results suggest that the antipathogen functions of the primate EARs are conserved after they are established and that these proteins are not currently undergoing rapid diversification in response to challenge from invading microorganisms. PMID- 11102387 TI - Genetic structure and evolution of RAC-GTPases in Arabidopsis thaliana. AB - Rho GTPases regulate a number of important cellular functions in eukaryotes, such as organization of the cytoskeleton, stress-induced signal transduction, cell death, cell growth, and differentiation. We have conducted an extensive screening, characterization, and analysis of genes belonging to the Ras superfamily of GTPases in land plants (embryophyta) and found that the Rho family is composed mainly of proteins with homology to RAC-like proteins in terrestrial plants. Here we present the genomic and cDNA sequences of the RAC gene family from the plant Arabidopsis thaliana. On the basis of amino acid alignments and genomic structure comparison of the corresponding genes, the 11 encoded AtRAC proteins can be divided into two distinct groups of which one group apparently has evolved only in vascular plants. Our phylogenetic analysis suggests that the plant RAC genes underwent a rapid evolution and diversification prior to the emergence of the embryophyta, creating a group that is distinct from rac/cdc42 genes in other eukaryotes. In embryophyta, RAC genes have later undergone an expansion through numerous large gene duplications. Five of these RAC duplications in Arabidopsis thaliana are reported here. We also present an hypothesis suggesting that the characteristic RAC proteins in higher plants have evolved to compensate the loss of RAS proteins. PMID- 11102388 TI - Inferring recent outcrossing rates using multilocus individual heterozygosity: application to evolving wheat populations. AB - Using multilocus individual heterozygosity, a method is developed to estimate the outcrossing rates of a population over a few previous generations. Considering that individuals originate either from outcrossing or from n successive selfing generations from an outbred ancestor, a maximum-likelihood (ML) estimator is described that gives estimates of past outcrossing rates in terms of proportions of individuals with different n values. Heterozygosities at several unlinked codominant loci are used to assign n values to each individual. This method also allows a test of whether populations are in inbreeding equilibrium. The estimator's reliability was checked using simulations for different mating histories. We show that this ML estimator can provide estimates of outcrossing rates for the final generation outcrossing rate (t(0)) and a mean of the preceding rates (t(p)) and can detect major temporal variation in the mating system. The method is most efficient for low to intermediate outcrossing levels. Applied to nine populations of wheat, this method gave estimates of t(0) and t(p). These estimates confirmed the absence of outcrossing t(0) = 0 in the two populations subjected to manual selfing. For free-mating wheat populations, it detected lower final generation outcrossing rates t(0) = 0-0.06 than those expected from global heterozygosity t = 0.02-0.09. This estimator appears to be a new and efficient way to describe the multilocus heterozygosity of a population, complementary to Fis and progeny analysis approaches. PMID- 11102389 TI - FARE, a new family of foldback transposons in Arabidopsis. AB - A new family of transposons, FARE, has been identified in Arabidopsis. The structure of these elements is typical of foldback transposons, a distinct subset of mobile DNA elements found in both plants and animals. The ends of FARE elements are long, conserved inverted repeat sequences typically 550 bp in length. These inverted repeats are modular in organization and are predicted to confer extensive secondary structure to the elements. FARE elements are present in high copy number, are heterogeneous in size, and can be divided into two subgroups. FARE1's average 1.1 kb in length and are composed entirely of the long inverted repeats. FARE2's are larger, up to 16.7 kb in length, and contain a large internal region in addition to the inverted repeat ends. The internal region is predicted to encode three proteins, one of which bears homology to a known transposase. FARE1.1 was isolated as an insertion polymorphism between the ecotypes Columbia and Nossen. This, coupled with the presence of 9-bp target-site duplications, strongly suggests that FARE elements have transposed recently. The termini of FARE elements and other foldback transposons are imperfect palindromic sequences, a unique organization that further distinguishes these elements from other mobile DNAs. PMID- 11102390 TI - A simple sequence repeat-based linkage map of barley. AB - A total of 568 new simple sequence repeat (SSR)-based markers for barley have been developed from a combination of database sequences and small insert genomic libraries enriched for a range of short simple sequence repeats. Analysis of the SSRs on 16 barley cultivars revealed variable levels of informativeness but no obvious correlation was found with SSR repeat length, motif type, or map position. Of the 568 SSRs developed, 242 were genetically mapped, 216 with 37 previously published SSRs in a single doubled-haploid population derived from the F(1) of an interspecific cross between the cultivar Lina and Hordeum spontaneum Canada Park and 26 SSRs in two other mapping populations. A total of 27 SSRs amplified multiple loci. Centromeric clustering of markers was observed in the main mapping population; however, the clustering severity was reduced in intraspecific crosses, supporting the notion that the observed marker distribution was largely a genetical effect. The mapped SSRs provide a framework for rapidly assigning chromosomal designations and polarity in future mapping programs in barley and a convenient alternative to RFLP for aligning information derived from different populations. A list of the 242 primer pairs that amplify mapped SSRs from total barley genomic DNA is presented. PMID- 11102391 TI - Ac insertion site affects the frequency of transposon-induced homologous recombination at the maize p1 locus. AB - The maize p1 gene regulates the production of a red pigment in the kernel pericarp, cob, and other maize floral tissues. Insertions of the transposable element Ac can induce recombination between two highly homologous 5.2-kb direct repeat sequences that flank the p1 gene-coding region. Here, we tested the effects of the Ac insertion site and orientation on the induction of recombination at the p1 locus. A collection of unique p1 gene alleles was used, which carry Ac insertions at different sites in and near the p1 locus, outside of the direct repeats, within the direct repeat sequences, and between the direct repeats, in both orientations. Recombination was scored by the numbers of colorless pericarp sectors (somatic frequency) and heritable mutations (germinal frequency). In both the somatic and germinal tests, the frequency of homologous recombination is significantly higher when Ac is inserted between the direct repeats than when Ac is inserted either within or outside the repeats. In contrast, Ac orientation had no significant effect on recombination frequency. We discuss these results in terms of the possible mechanisms of transposon-induced recombination. PMID- 11102393 TI - Flow sorting of mitotic chromosomes in common wheat (Triticum aestivum L.). AB - The aim of this study was to develop an improved procedure for preparation of chromosome suspensions, and to evaluate the potential of flow cytometry for chromosome sorting in wheat. Suspensions of intact chromosomes were prepared by mechanical homogenization of synchronized root tips after mild fixation with formaldehyde. Histograms of relative fluorescence intensity (flow karyotypes) obtained after the analysis of DAPI-stained chromosomes were characterized and the chromosome content of all peaks on wheat flow karyotype was determined for the first time. Only chromosome 3B could be discriminated on flow karyotypes of wheat lines with standard karyotype. Remaining chromosomes formed three composite peaks and could be sorted only as groups. Chromosome 3B could be sorted at purity >95% as determined by microscopic evaluation of sorted fractions that were labeled using C-PRINS with primers for GAA microsatellites and for Afa repeats, respectively. Chromosome 5BL/7BL could be sorted in two wheat cultivars at similar purity, indicating a potential of various wheat stocks for sorting of other chromosome types. PCR with chromosome-specific primers confirmed the identity of sorted fractions and suitability of flow-sorted chromosomes for physical mapping and for construction of small-insert DNA libraries. Sorted chromosomes were also found suitable for the preparation of high-molecular-weight DNA. On the basis of these results, it seems realistic to propose construction of large-insert chromosome-specific DNA libraries in wheat. The availability of such libraries would greatly simplify the analysis of the complex wheat genome. PMID- 11102392 TI - Mutator-like elements in Arabidopsis thaliana. Structure, diversity and evolution. AB - While genome-wide surveys of abundance and diversity of mobile elements have been conducted for some class I transposable element families, little is known about the nature of class II transposable elements on this scale. In this report, we present the results from analysis of the sequence and structural diversity of Mutator-like elements (MULEs) in the genome of Arabidopsis thaliana (Columbia). Sequence similarity searches and subsequent characterization suggest that MULEs exhibit extreme structure, sequence, and size heterogeneity. Multiple alignments at the nucleotide and amino acid levels reveal conserved, potentially transposition-related sequence motifs. While many MULEs share common structural features to Mu elements in maize, some groups lack characteristic long terminal inverted repeats. High sequence similarity and phylogenetic analyses based on nucleotide sequence alignments indicate that many of these elements with diverse structural features may remain transpositionally competent and that multiple MULE lineages may have been evolving independently over long time scales. Finally, there is evidence that MULEs are capable of the acquisition of host DNA segments, which may have implications for adaptive evolution, both at the element and host levels. PMID- 11102394 TI - A mixed-model approach to mapping quantitative trait loci in barley on the basis of multiple environment data. AB - In this article, I propose a mixed-model method to detect QTL with significant mean effect across environments and to characterize the stability of effects across multiple environments. I demonstrate the method using the barley dataset by the North American Barley Genome Mapping Project. The analysis raises the need for mixed modeling in two different ways. First, it is reasonable to regard environments as a random sample from a population of target environments. Thus, environmental main effects and QTL-by-environment interaction effects are regarded as random. Second, I expect a genetic correlation among pairs of environments caused by undetected QTL. I show how random QTL-by-environment effects as well as genetic correlations are straightforwardly handled in a mixed model framework. The main advantage of this method is the ability to assess the stability of QTL effects. Moreover, the method allows valid statistical inferences regarding average QTL effects. PMID- 11102395 TI - A note on algorithms for genotype and allele elimination in complex pedigrees with incomplete genotype data. AB - Elimination of genotypes or alleles for each individual or meiosis, which are inconsistent with observed genotypes, is a component of various genetic analyses of complex pedigrees. Computational efficiency of the elimination algorithm is critical in some applications such as genotype sampling via descent graph Markov chains. We present an allele elimination algorithm and two genotype elimination algorithms for complex pedigrees with incomplete genotype data. We modify all three algorithms to incorporate inheritance restrictions imposed by a complete or incomplete descent graph such that every inconsistent complete descent graph is detected in any pedigree, and every inconsistent incomplete descent graph is detected in any pedigree without loops with the genotype elimination algorithms. Allele elimination requires less CPU time and memory, but does not always eliminate all inconsistent alleles, even in pedigrees without loops. The first genotype algorithm produces genotype lists for each individual, which are identical to those obtained from the Lange-Goradia algorithm, but exploits the half-sib structure of some populations and reduces CPU time. The second genotype elimination algorithm deletes more inconsistent genotypes in pedigrees with loops and detects more illegal, incomplete descent graphs in such pedigrees. PMID- 11102396 TI - Empirical Bayes procedure for estimating genetic distance between populations and effective population size. AB - We developed an empirical Bayes procedure to estimate genetic distances between populations using allele frequencies. This procedure makes it possible to describe the skewness of the genetic distance while taking full account of the uncertainty of the sample allele frequencies. Dirichlet priors of the allele frequencies are specified, and the posterior distributions of the various composite parameters are obtained by Monte Carlo simulation. To avoid overdependence on subjective priors, we adopt a hierarchical model and estimate hyperparameters by maximizing the joint marginal-likelihood function. Taking advantage of the empirical Bayesian procedure, we extend the method to estimate the effective population size using temporal changes in allele frequencies. The method is applied to data sets on red sea bream, herring, northern pike, and ayu broodstock. It is shown that overdispersion overestimates the genetic distance and underestimates the effective population size, if it is not taken into account during the analysis. The joint marginal-likelihood function also estimates the rate of gene flow into island populations. PMID- 11102397 TI - Mapping quantitative trait loci in complex pedigrees: a two-step variance component approach. AB - There is a growing need for the development of statistical techniques capable of mapping quantitative trait loci (QTL) in general outbred animal populations. Presently used variance component methods, which correctly account for the complex relationships that may exist between individuals, are challenged by the difficulties incurred through unknown marker genotypes, inbred individuals, partially or unknown marker phases, and multigenerational data. In this article, a two-step variance component approach that enables practitioners to routinely map QTL in populations with the aforementioned difficulties is explored. The performance of the QTL mapping methodology is assessed via its application to simulated data. The capacity of the technique to accurately estimate parameters is examined for a range of scenarios. PMID- 11102398 TI - Detecting the undetected: estimating the total number of loci underlying a quantitative trait. AB - Recent studies have begun to reveal the genes underlying quantitative trait differences between closely related populations. Not all quantitative trait loci (QTL) are, however, equally likely to be detected. QTL studies involve a limited number of crosses, individuals, and genetic markers and, as a result, often have little power to detect genetic factors of small to moderate effects. In this article, we develop an estimator for the total number of fixed genetic differences between two parental lines. Like the Castle-Wright estimator, which is based on the observed segregation variance in classical crossbreeding experiments, our QTL-based estimator requires that a distribution be specified for the expected effect sizes of the underlying loci. We use this expected distribution and the observed mean and minimum effect size of the detected QTL in a likelihood model to estimate the total number of loci underlying the trait difference. We then test the QTL-based estimator and the Castle-Wright estimator in Monte Carlo simulations. When the assumptions of the simulations match those of the model, both estimators perform well on average. The 95% confidence limits of the Castle-Wright estimator, however, often excluded the true number of underlying loci, while the confidence limits for the QTL-based estimator typically included the true value approximately 95% of the time. Furthermore, we found that the QTL-based estimator was less sensitive to dominance and to allelic effects of opposite sign than the Castle-Wright estimator. We therefore suggest that the QTL-based estimator be used to assess how many loci may have been missed in QTL studies. PMID- 11102399 TI - Monte Carlo evaluation of the likelihood for N(e) from temporally spaced samples. AB - A population's effective size is an important quantity for conservation and management. The effective size may be estimated from the change of allele frequencies observed in temporally spaced genetic samples taken from the population. Though moment-based estimators exist, recently Williamson and Slatkin demonstrated the advantages of a maximum-likelihood approach that they applied to data on diallelic genetic markers. Their computational methods, however, do not extend to data on multiallelic markers, because in such cases exact evaluation of the likelihood is impossible, requiring an intractable sum over latent variables. We present a Monte Carlo approach to compute the likelihood with data on multiallelic markers. So as to be computationally efficient, our approach relies on an importance-sampling distribution constructed by a forward-backward method. We describe the Monte Carlo formulation and the importance-sampling function and then demonstrate their use on both simulated and real datasets. PMID- 11102400 TI - A method for estimating the intensity of overdominant selection from the distribution of allele frequencies. AB - A method is proposed for estimating the intensity of overdominant selection scaled by the effective population size, S = 2Ns, from allele frequencies. The method is based on the assumption that, with strong overdominant selection, allele frequencies are nearly at their deterministic equilibrium values and that, to a first approximation, deviations depend only on S. Simulations verify that reasonably accurate estimates of S can be obtained for realistic sample sizes. The method is applied to data from several loci in the major histocompatibility complex (Mhc) in numerous human populations. For alleles distinguished by both serological typing and the sequence of the peptide-binding region, our estimates of S are comparable to those obtained by analysis of DNA sequences in showing that selection is strongest on HLA-B and weaker on HLA-A, HLA-DRB1, and HLA-DQA1. The intensity of selection on HLA-B varied considerably among populations. Two populations, Native American and Inuit, showed an excess rather than a deficiency in homozygosity. Comparable estimates of S were obtained for alleles at Mhc class II loci distinguished by serological reactions (serotyping) and by differences in the amino acid sequences of the peptide-binding region (molecular typing). A comparison of two types of data for DQA1 and DRB1 showed that serotyping led to generally lower estimates of S. PMID- 11102401 TI - Testing for concordant equilibria between population samples. AB - A substantial body of theory has been developed to assess the effect of evolutionary forces on the distribution of genotypes, both single and multilocus, within populations. One area where the potential for application of this theory has not been fully appreciated concerns the extent to which population samples differ. Within populations, the divergence of genotype or haplotype frequencies from that expected under Hardy-Weinberg (HW) or linkage equilibrium can be measured as disequilibria coefficients. To assess population samples for concordant equilibria, an analytical framework for comparing disequilibria coefficients between populations is necessary. Here we present log-linear models to evaluate such hypotheses. These models have broad utility ranging from conventional population genetics to genetic epidemiology. We demonstrate the use of these log-linear models (1) as a test for genetic association with disease and (2) as a test for different levels of linkage disequilibria between human populations. PMID- 11102402 TI - New quinolones--a fresh answer to the pneumococcus. PMID- 11102403 TI - Infection in the twenty-first century: predictions and postulates. AB - The late Paul Garrod, in whose honour this lecture is named, was 'the right man at the right time'. He seized the opportunities offered by the dawning of the chemotherapeutic era with vigour and enthusiasm and was a formidable link between the traditional laboratory-based bacteriologist and the more clinically orientated 'modern' medical microbiologist. Professor Garrod was a founder member of the British Society for Antimicrobial Chemotherapy and I had the privilege of meeting him on many occasions. He would have relished the many challenges facing today's microbiologists, infectious disease physicians and public health experts. These will have major implications for antimicrobial chemotherapy in the twenty first century. The emergence and prevalence of infectious diseases, and the necessity for discovering therapies to treat them, are influenced by many factors. In this lecture I will discuss four which could have a major influence on infectious diseases in the twenty-first century-global warming, biological warfare/terrorism, the dissemination of infections, including those caused by resistant pathogens, by travellers and certain untreatable zoonotic diseases. PMID- 11102404 TI - A TEM-2beta-lactamase encoded on an active Tn1-like transposon in the genome of a clinical isolate of Stenotrophomonas maltophilia. AB - A constitutively expressed beta-lactamase gene from a clinical isolate of Stenotrophomonas maltophilia, J675Ia, has been cloned. Its DNA sequence is almost identical to that of bla(TEM2) (one nucleotide change) and the expressed enzyme is a Bush type 2a penicillinase with an amino acid sequence identical to that of TEM-2. The bla(TEM) gene was present within a novel Tn1/Tn3-type transposon in the genome of isolate J675Ia and the transposon was able to mobilize bla(TEM) on to the broad host-range conjugative plasmid, R388. When transferred to an Escherichia coli recipient, R388::Tn conferred high-level ampicillin resistance. This represents the first identification of a TEM beta-lactamase in S. maltophilia and the first evidence that this important clinical pathogen is able to act as a reservoir for mobile beta-lactamase genes in the hospital environment. PMID- 11102405 TI - Influence of the MexA-MexB-oprM multidrug efflux system on expression of the MexC MexD-oprJ and MexE-MexF-oprN multidrug efflux systems in Pseudomonas aeruginosa. AB - Of the Pseudomonas aeruginosa multidrug efflux systems, MexAB-OprM is expressed in wild-type cells, while MexCD-OprJ is not, and MexEF-OprN shows variable, strain-specific expression. In defined mutant strains, MexCD-OprJ expression increased with decreases in MexAB-OprM and was generally inversely related to MexAB-OprM expression. In so-called wild-type strains expressing MexEF-OprN, MexAB-OprM hyperexpression correlated with a decline in MexEF-OprN expression, while loss of MexAB-OprM was associated with increased expression of MexEF-OprN, also indicative of an inverse correlation between MexAB-OprM and MexEF-OprN expression. Still, the increases in MexCD-OprJ and MexEF-OprN failed to compensate for the loss of MexAB-OprM with respect to antibiotic resistance. Nonetheless, these data suggest that the overall complement of these MDR efflux systems is monitored and that alterations in the level of one efflux system may effect compensatory changes in the levels of the others. PMID- 11102406 TI - Emergence of imipenem resistance in Klebsiella pneumoniae owing to combination of plasmid-mediated CMY-4 and permeability alteration. AB - Klebsiella pneumoniae BM2974 isolated from an abdominal abcess was resistant to high concentrations of all available beta-lactams, including recently developed third-generation cephalosporins and carbapenems. Isoelectric focusing of beta lactamases and amplification, cloning and sequencing of the corresponding genes, together with conjugation and transformation experiments, indicated that, in addition to the chromosomally encoded beta-lactamase, the strain produced three plasmid-mediated beta-lactamases with pIs of 5.4, 8.2 and 9.0, which corresponded to TEM-1, SHV-5 and AmpC-type CMY-4, respectively. Strain BM2974 also lacked a major outer membrane protein of c. 40 kDa which was present in the spontaneous imipenem-susceptible revertant BM2974-1. We suggest that imipenem resistance in strain BM2974 is attributable to production of CMY-4 beta-lactamase combined with permeability alteration. PMID- 11102407 TI - Combined effects of meropenem and aminoglycosides on Pseudomonas aeruginosa in vitro. AB - To investigate combinations of antibiotics against Pseudomonas aeruginosa, the in vitro effects of combinations of meropenem with each of three aminoglycosides, arbekacin, amikacin and netilmicin, were evaluated using an agar dilution chequerboard technique. The combinations of meropenem and aminoglycosides were effective against almost all P. aeruginosa strains tested, which included meropenem-resistant strains. Increased synergic effects were observed in combinations that included arbekacin or amikacin. None of the combinations had an antagonistic effect. Most of the synergic and additive effects were achieved at clinically relevant concentrations. PMID- 11102408 TI - In vitro activity of ketolides telithromycin and HMR 3004 against italian isolates of Streptococcus pyogenes and Streptococcus pneumoniae with different erythromycin susceptibility. AB - Two ketolides, telithromycin and HMR 3004, were evaluated for their in vitro activity against erythromycin-susceptible and -resistant strains of Streptococcus pyogenes and Streptococcus pneumoniae. On the basis of their resistance to macrolide, lincosamide and streptogramin (MLS) antibiotics, erythromycin resistant test strains were assigned to the constitutive resistance (cMLS) phenotype, the inducible resistance (iMLS) phenotype or the M phenotype. iMLS S. pyogenes strains were further subdivided into the three recently described subtypes iMLS-A, -B and -C. Telithromycin and HMR 3004 were uniformly and highly active against pneumococci (regardless of their susceptibility or resistance to erythromycin and/or penicillin), erythromycin-susceptible S. pyogenes and erythromycin-resistant S. pyogenes strains of the M phenotype (in which resistance is mediated by an efflux system) or iMLS-B or -C phenotype (in which resistance is mediated by a methylase encoded by the ermTR gene). Both ketolides were less active against erythromycin-resistant S. pyogenes strains with the cMLS phenotype or the iMLS-A subtype (where resistance is mediated by a methylase encoded by the ermAM gene), these strains ranging in phenotype from the upper limits of susceptibility to low-level resistant. PMID- 11102409 TI - In vitro development of resistance to ceftriaxone, cefprozil and azithromycin in Streptococcus pneumoniae. AB - Approval of ceftriaxone for the treatment of otitis media has led to fear of selection of resistant mutants owing to widespread use. To test this, we examined the ability of sequential subcultures in sub-MICs of ceftriaxone, cefprozil and azithromycin to select resistant mutants in 12 pneumococci. Daily subculturing was performed 50 times or until mutants with raised ceftriaxone, cefprozil or azithromycin MICs were selected. Of eight ceftriaxone-susceptible parents, ceftriaxone did not select for any resistant mutants, while cefprozil selected for four mutants (MICs 2-4 mg/L after 21-50 subcultures). Among four ceftriaxone resistant parents, subculturing in ceftriaxone selected for one stable mutant with raised ceftriaxone MIC (>16 mg/L after 21 subcultures) and subculturing in cefprozil selected for one mutant with raised cefprozil MIC (64 mg/L after 44 subcultures). Mutations were observed in pbp2x and pbp1a. Among six azithromycin susceptible parents, subculturing in azithromycin selected for five resistant mutants (MIC 0.5-32 mg/L after 10-42 passages) and among six azithromycin resistant strains, subculturing selected for mutants with raised azithromycin MICs in all six strains (MIC 16-32 mg/L after 4-18 passages). All azithromycin resistant mutants derived from azithromycinsusceptible parents had mutations in domain V of 23S rRNA while all azithromycin-resistant parents and derived mutants had mefE. Single-step mutation rates among the 12 strains at the MIC ranged from 1.5 x 10(-6) to <6.2 x 10(-10) for ceftriaxone, >1.3 x 10(-5) to 8.9 x 10(-8) for cefprozil and >1.1 x 10(-6) to 6.7 x 10(-10) for azithromycin. Multi-step and single-step testing showed that ceftriaxone selected for resistant mutants less often than cefprozil and azithromycin. PMID- 11102410 TI - Bactericidal activity of nitrofurans against growing and dormant Mycobacterium bovis BCG. AB - Depletion of oxygen triggers the shift-down of Mycobacterium bovis BCG to a state of dormancy. Bacilli in their dormant state are resistant to standard anti mycobacterials. The nitroimidazole metronidazole was the first compound identified to show bactericidal activity against dormant tubercle bacilli. In contrast to metronidazole's selective toxicity for dormant bacilli, we report here that the nitrofurans nitrofurantoin, furaltadone and nitrofurazone showed bactericidal activity against dormant and growing bacteria. Importantly, the bactericidal effect of nitrofurans on dormant bacilli was 35- to 250-fold higher compared with metronidazole. PMID- 11102411 TI - The effect of three broad-spectrum antimicrobials on mononuclear cell responses to encapsulated bacteria: evidence for down-regulation of cytokine mRNA transcription by trovafloxacin. AB - The effect of trovafloxacin, ciprofloxacin and ceftriaxone on cytokine production of human peripheral blood mononuclear cells (PBMCs) was examined. PBMC responses were measured after stimulation with lipopolysaccharide (LPS), lipoteichoic acid (LTA) or killed or viable Streptococcus pneumoniae and Haemophilus influenzae. Trovafloxacin inhibited the production of tumour necrosis factor alpha (TNF alpha), interleukin-1beta (IL-1beta), IL-6 and IL-8 by PBMCs after stimulation with either LPS or LTA by 83%. Similar inhibition occurred in PBMCs incubated with killed or live bacteria and trovafloxacin, but not with ciprofloxacin or ceftriaxone. The relevance of this in vitro observation was explored by examining TNF-alpha and IL-6 responses in trovafloxacin-treated mice. Serum concentrations of both cytokines 1 h after LPS challenge were 95% less than serum concentrations in mice that were not given trovafloxacin. Reverse transcription- polymerase chain reaction studies of the mechanisms determining cytokine down-regulation demonstrated that trovafloxacin reduced TNF-alpha, IL-1beta and IL-6 mRNA to levels similar to those of unstimulated cells. These observations indicate that trovafloxacin can consistently and significantly reduce production of cytokines that play an important role in sepsis. In vitro, this effect can occur in the presence of bacteriolysis and is associated with inhibition of transcription of cytokine genes. PMID- 11102412 TI - Susceptibility of gram-positive cocci from 25 UK hospitals to antimicrobial agents including linezolid. The Linezolid Study Group. AB - The prevalence of antibiotic resistance amongst Gram-positive cocci from 25 UK hospitals was studied over an 8 month period in 1999. A total of 3770 isolates were tested by the sentinel laboratories using the Etest; these bacteria comprised 1000 pneumococci, 1005 Staphylococcus aureus, 769 coagulase-negative staphylococci (CNS) and 996 enterococci. To ensure quality, 10% of the isolates were retested centrally, as were any found to express unusual resistance patterns. The prevalence of penicillin-resistant Streptococcus pneumoniae, vancomycin-resistant enterococci and methicillin-resistant S. aureus (MRSA) varied widely amongst the sentinel laboratories. The resistance rates to methicillin among S. aureus and CNS were 19.2 and 38.9%, respectively, with MRSA rates in individual sentinel sites ranging from 0 to 43%. No glycopeptide resistance was seen in S. aureus, but 6.5% of CNS isolates were teicoplanin resistant and 0.5% were vancomycin resistant. Vancomycin resistance was much more frequent among Enterococcus faecium (24.1%) than E. faecalis (0.5%) (P<0.05), with most resistant isolates carrying vanA. The rate of penicillin resistance in pneumococci was 8.9%, and this resistance was predominantly intermediate (7.9%), with only six hospitals reporting isolates with high level resistance. The prevalence of erythromycin resistance among pneumococci was 12.3%, with the majority of resistant isolates having the macrolide efflux mechanism mediated by mefE. All the organisms tested were susceptible to linezolid with MICs in the range 0.12-4 mg/L. The modal MICs of linezolid were 1 mg/L for CNS and pneumococci, and 2 mg/L for S. aureus and enterococci. Linezolid was the most potent agent tested against Gram-positive cocci, including multiresistant strains, and as such may prove a valuable therapeutic option for the management of Gram-positive infections in hospitals. PMID- 11102413 TI - Patterns of phenotypic resistance to the macrolide-lincosamide-ketolide streptogramin group of antibiotics in staphylococci. AB - Phenotypes of resistance to the macrolide-lincosamide-ketolide-streptogramin (MLKS) group of antibiotics have been determined in 540 clinical isolates of staphylococci (210 Staphylococcus aureus and 330 coagulase-negative species). Results of disc diffusion tests using erythromycin A, oleandomycin, rokitamycin, clindamycin, telithromycin, quinupristin and dalfopristin delineated four main groups corresponding to those defined classically using erythromycin and clindamycin only, but with sub-divisions. Resistance to erythromycin was more common in coagulase-negative strains (56%) than in S. aureus (16%); telithromycin, clindamycin, quinupristin-dalfopristin and rokitamycin were active against >97% of S. aureus strains and >88% of the coagulase-negative strains. The commonest resistance phenotype was 'inducible MLS(B)' (12% in S. aureus, 31% in coagulase-negative strains); this group could be divided in terms of the different inducing abilities of erythromycin and oleandomycin. 'Constitutive MLS(B)' and 'MS' phenotypes were more often found in coagulase-negative strains (11 and 13%, respectively) than in S. aureus (2 and 1%). Novel phenotypes were found during the isolation of constitutively resistant mutants from inducible strains, and of resistant mutants from 'MS' strains. This extended phenotyping scheme has revealed further complexities and evolutionary possibilities in patterns of resistance to this group of antibiotics. PMID- 11102414 TI - The beta-lactamases and beta-lactam antibiotic susceptibility of Yersinia enterocolitica. AB - One hundred and forty-five isolates of Yersinia enterocolitica of different serotypes and biotypes, including atypical biotypes, collected from various parts of the world, were examined for their susceptibility to beta-lactam antibiotics and expression of intracellular beta-lactamases. The reasons for the specificity of patterns of susceptibility to beta-lactams for each biotype or subtype of Y. enterocolitica were elucidated by examining their ss-lactamase activity. Whilst the biotypes and subtypes were uniformly susceptible to the newer beta-lactam antibiotics, the susceptibility pattern observed with other beta-lactams was specific to each biotype or subtype, because of the characteristics of beta lactamase expression by strains within these groups. The susceptibility to these beta-lactam agents depended entirely on the extent of elaboration or the absence of one of the two beta-lactamases, enzyme A and enzyme B, found in the species. Detection of enzyme B by a disc diffusion test yielded inconsistent results, but detection of enzyme A by disc diffusion was highly reliable. This test clearly distinguished strains of biotype 2, serotype O:5,27 from those of biotype 2, serotype O:9 and biotype 3, serotypes O:1, 2a-3, O:3 and O:5. PMID- 11102415 TI - Streptococcus pyogenes resistance to erythromycin in relation to macrolide consumption in Spain (1986-1997). AB - The relationship between Streptococcus pyogenes resistance to erythromycin and macrolide consumption in Spain was studied. Erythromycin resistance was highly correlated with the consumption of total macrolides (r = 0.88, P<0.01). When macrolides were grouped into posological subgroups according to their pharmacokinetic and pharmacodynamic properties and analysed separately, erythromycin resistance appeared to be related mainly to those macrolides taken twice daily (bd) (r = 0.86, P<0.01) and those taken once daily (od) (r = 0.87, P<0.01), but not to those taken four (qds) or three times a day (tds) (r = -0.04, P: = 0.90). A progressive increase in the erythromycin resistance curve was seen after the consecutive introduction of both bd and od macrolides, which contributed to the increase in the total macrolide consumption, replacing tds macrolide prescription. Although this ecological analysis cannot establish an unequivocal causal relationship between antibiotic consumption and S. pyogenes resistance, the data are consistent with the hypothesis that widespread use of macrolides, mainly of bd and od macrolides, resulted in an increased prevalence of S. pyogenes resistant to erythromycin in Spain. PMID- 11102416 TI - Antibiotic resistance in salmonellae isolated from humans and animals in France: comparative data from 1994 and 1997. AB - Among 25526 recorded isolates of salmonellae, 5086 isolated from humans and 20440 from animals in 1994 and 1997 in France, the antibiotic resistance phenotype was determined for all human and 5336 animal isolates. In Salmonella enterica serovar typhimurium, one of the two most frequently isolated serovars from humans as well as animals, resistance to ampicillin was observed in 61% of both human and animal isolates in 1994 and in 73% of human and 53% of animal isolates in 1997. During these periods, resistance to co-amoxiclav was between 45% and 66% for both types of isolate. Resistance to ampicillin was associated with resistance to streptomycin, spectinomycin, sulphonamide, tetracycline and chloramphenicol in over 70% of isolates. Resistance to ampicillin as well as co-amoxiclav never exceeded 7% in Salmonella enteritidis. While Salmonella hadar was practically absent among the human isolates in 1994, this serovar was the third most frequent in 1997, and at that time 92% were resistant to nalidixic acid. Among the animal S. hadar isolates, the prevalence of resistance to nalidixic acid increased from 3% in 1994 to 72% in 1997. None of these isolates manifested high-level resistance to ofloxacin. The levels of resistance to aminoglycosides (< or =3%) and trimethoprim-suphamethoxazole (< or =14%) remained practically unchanged in all three serovars. The resistance markers of 463 ampicillin-resistant S. typhimurium isolated in 1997 were determined. Among the 24 phenotypes observed, six multiresistance phenotypes, representing 82% of these isolates (as compared with 80% in 1994), were associated with the PSE-1 gene typically found in the lysotype DT104 of this serovar. PMID- 11102417 TI - Antagonism between penicillin and erythromycin against Streptococcus pneumoniae in vitro and in vivo. AB - The combination of beta-lactam antibiotics and macrolides is often recommended for the initial empirical treatment of acute pneumonia in order to obtain activity against the most important pathogens. Theoretically, this combination may be inexpedient, as the bacteriostatic agent may antagonize the effect of the bactericidal agent. In this study, the possible interaction between penicillin and erythromycin was investigated in vitro and in vivo against four clinical isolates of Streptococcus pneumoniae with MICs of penicillin ranging from 0.016 to 0.5 mg/L and of erythromycin from 0. 25 to >128 mg/L. In vitro time-kill curves were generated with clinically relevant concentrations of penicillin (10 mg/L) and erythromycin (1 mg/L), either individually or in combination. Antagonism between penicillin and erythromycin was observed for the four isolates. In vivo interaction was investigated in the mouse peritonitis model. After intraperitoneal inoculation, penicillin and erythromycin were given either individually or in combination. For two of the four isolates, mortality was significantly higher in the groups treated with the combination of penicillin and erythromycin than in the groups treated with penicillin alone [32/36 (86%) vs. 3/12 (25%), P<0.05; and 24/36 (67%) vs. 3/12 (25%), P<0.05, respectively]. Using the mouse peritonitis model, in vivo time-kill curves showed that there was antagonism between erythromycin and penicillin for the examined isolate. The antagonism demonstrated in vitro and in vivo between penicillin and erythromycin suggests that ss-lactam antibiotics and macrolides should not be administered together unless pneumococcal infection is ruled out. PMID- 11102418 TI - Linezolid against penicillin-sensitive and -resistant pneumococci in the rabbit meningitis model. AB - Linezolid, a new oxazolidinone antibiotic, showed good penetration (38+/-4%) into the meninges of rabbits with levels in the CSF ranging from 9.5 to 1.8 mg/L after two i.v. injections (20 mg/kg). Linezolid was clearly less effective than ceftriaxone against a penicillin-sensitive pneumococcal strain. Against a penicillin-resistant strain, linezolid had slightly inferior killing rates compared with the standard regimen (ceftriaxone combined with vancomycin). In vitro, linezolid was marginally bactericidal at concentrations above the MIC (5 x and 10 x MIC). PMID- 11102419 TI - Short-term infection with Helicobacter pylori and 1 week exposure to metronidazole does not enhance gastric mutation frequency in transgenic mice. AB - The aim of this study was to determine whether exposure of Helicobacter pylori infected mice to metronidazole resulted in the delivery of mutagenic compounds to the gastric epithelium via the oxygen-insensitive NADPH nitroreductase (RdxA) of H. pylori. C57BL/6 transgenic mice containing the lambda/lacI transgene were inoculated with peptone trypsin broth, H. pylori SS1 or SS1-rdxA(-), an SS1 derived mutant in rdxA. Twelve weeks after inoculation, the mice were treated for 7 days with a control solution or with the mouse equivalent of a human dose of metronidazole 1 g od. Three weeks after completion of treatment, the animals were killed and mutations in the target lacI gene assessed by a transgenic mutagenesis assay system. There was no increase in lacI mutations in cells harvested from mice infected with H. pylori and/or exposed to metronidazole. These data suggest that short-term infection with H. pylori and exposure to metronidazole does not enhance the mutation frequency in the gastric cells of mice. Whether chronic infection and/or repeated exposure to metronidazole or other nitroaromatic compounds causes genetic damage to gastric epithelial cells remains to be determined. PMID- 11102420 TI - Pharmacokinetics of enoxacin and its oxometabolite after multiple oral dosing and penetration into prostatic tissue. AB - The objective of this study was to determine the concentrations of enoxacin and its oxo-metabolite in human prostatic tissue after multiple oral doses (400 mg bd) in 13 patients. On the first day of treatment, elimination half-lives were 6.8 h for enoxacin and 7.1 h for its metabolite; they were increased on day 4 (10.3 and 13.2 h, respectively). The ratios of drug concentration in prostatic tissue and plasma averaged 2.2 for enoxacin and 1.4 for its metabolite. In conclusion, concentrations of enoxacin achieved within the prostatic tissue were higher than plasma concentrations suggesting that there was an active transport mechanism. PMID- 11102421 TI - Comparison of the Etest and microdilution method for antifungal susceptibility testing of Cryptococcus neoformans to four antifungal agents. AB - We performed a prospective study to compare the Etest and the microdilution method (NCCLS guidelines) for determining the MICs of fluconazole, itraconazole, flucytosine and amphotericin B for 35 strains of Cryptococcus neoformans. For the microdilution method (MDM) RPMI 1640 medium with 2% glucose was used for fluconazole, itraconazole and flucytosine, and Antibiotic Medium 3 for amphotericin B. For the Etest, RPMI 1640 medium with 2% glucose and solidified with 1.5% agar was used for the four antifungal agents. Amphotericin B was also tested on Antibiotic Medium 3 solidified with 1.5% agar. Fluconazole and flucytosine MICs by the Etest showed good correlation with the broth MDM (81.1 and 89.2% agreement within two dilutions, respectively). With the tested population of itraconazole- and amphotericin B-susceptible isolates, the MIC agreement for itraconazole was 54%; amphotericin B showed the lowest agreement (8.1% on Antibiotic Medium 3 and 13.5% on RPMI). PMID- 11102422 TI - Impact of fluconazole prophylaxis on fungal colonization and infection rates in neutropenic patients. The Canadian Fluconazole Study. AB - Fungal colonization profiles from four different anatomical sites were evaluated in 266 neutropenic cancer patients receiving intensive cytotoxic therapy for acute leukaemia or for autologous marrow transplantation. At the beginning of chemotherapy patients were allocated randomly to receive oral fluconazole 400 mg daily or an identical placebo until prophylaxis failure or marrow recovery. Candida albicans colonization was reduced from 30 to 10% in the fluconazole recipients while it increased from 32 to 57% in the placebo patients (P<0.001). By the end of prophylaxis, colonization with non-albicans Candida species increased from 7 to 21% and 8 to 18% in the fluconazole and placebo patients, respectively (P = 0.396). Although Candida glabrata was isolated more frequently at the end of the prophylactic period in the fluconazole patients than in the placebo patients (16 versus 7%), only one definite invasive C. glabrata infection was noted. Overall, definite invasive fungal infections were documented in 26 patients [four fluconazole versus 22 placebo patients (P< or =0.001)]. In 23 (92%) patients the infections were caused by persistently colonizing or newly acquired organisms. While probable invasive fungal infections were noted in five fluconazole patients, 10 placebo patients were also affected (P = 0.19). An end of-prophylaxis colonization index >0.25 was 76% sensitive but only 69% specific for invasive fungal infection. However, a colonization index < or =0.25 at baseline had a negative predictive value of 88% for development of invasive fungal infection. Fluconazole prophylaxis decreased colonization by fungi and subsequent invasive fungal infections in neutropenic cancer patients. PMID- 11102423 TI - Expression of the carbapenemase gene (cfiA) in Bacteroides fragilis. AB - Bacteroides fragilis strains were studied to examine carbapenemase gene (cfiA) expression and insertion sequence (IS) element promoters. High-level resistance was associated with high meropenemase activity and IS elements upstream of cfiA. Sequencing revealed two element types; IS1187 and elements related to IS942 and IS1170. The latter was implicated in the conversion to carbapenem resistance in a cfiA-positive isolate during imipenem therapy. Two strains showing low-level resistance, and strains susceptible to meropenem, did not possess IS elements upstream of cfiA. The prevalence in Nottingham of clinical isolates of B. fragilis with cfiA and efficient IS element promoters was low (0.6%). PMID- 11102424 TI - Pheromone responses and high-level aminoglycoside resistance of conjugative plasmids of Enterococcus faecalis from Greece. AB - Fifteen of 22 clinical isolates of Enterococcus faecalis with high-level aminoglycoside resistance isolated in Greece, which were tested for mating ability, co-transferred pheromone response genes together with aminoglycoside resistance determinants to a sensitive recipient strain. Nine of them belonged to the same pulsotype, while the remaining six isolates were genetically unrelated. The prgB gene, which encodes aggregation substance, was detected in all the clinical isolates and transconjugants by both PCR and DNA hybridization but prgA, which encodes the surface exclusion protein, was only detected in two isolates, whereas it is present in most pheromone response plasmids from other sources. PMID- 11102425 TI - The in vitro activity of ABT773, a new ketolide antimicrobial agent. AB - The in vitro activity of ABT773, a ketolide antimicrobial agent, was investigated and compared with those of seven other antibiotics. Type strains and 733 Gram positive, Gram-negative and anaerobic isolates of clinical origin and four CHLAMYDIA: isolates were used. The activity of ABT773 was very similar to that of telithromycin, the other ketolide tested. The MIC(90) was < or = 0.5 mg/L for all bacteria examined except methicillin-resistant Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Haemophilus influenzae and BACTEROIDES: spp. The antichlamydial activity of ABT773 was greater than that of telithromycin, erythromycin and ciprofloxacin. Neither an increase in the size of the inoculm nor the addition of human serum had any marked affect on the in vitro activity of ABT773. PMID- 11102426 TI - In vitro activity of gemifloxacin (SB-265805) compared with 14 other antimicrobials against intestinal pathogens. AB - We studied the in vitro activity of gemifloxacin (SB-265805) and 14 comparator antimicrobials against 288 recent isolates of enteropathogenic bacteria (106 Salmonella: spp., 32 Hafnia alvei, 22 Yersinia enterocolitica, 21 Shigella: spp., 16 Aeromonas: spp. and 91 Campylobacter jejuni). Gemifloxacin, the other fluoroquinolones and cefotaxime were very active against all microorganisms tested except for C. jejuni. Seventy-seven per cent of isolates of C. jejuni were inhibited by erythromycin < or =0.5 mg/L. Only one strain of C. jejuni was highly resistant to this antimicrobial agent. Of the compounds tested, gentamicin was the most active in vitro. The in vitro activity of the other antibiotics tested was variable. A quinolone could be a good choice for treating gastrointestinal infections when antimicrobial therapy is indicated. For C. jejuni, another antibiotic such as erythromycin should be considered. PMID- 11102427 TI - Features and trends in Helicobacter pylori antibiotic resistance in Lisbon area, Portugal (1990-1999). AB - The features of Helicobacter pylori antibiotic resistance in Lisbon from 1990 to 1999 were studied. Overall resistance rates to amoxycillin, tetracycline, metronidazole, clarithromycin and ciprofloxacin were 0, 0, 30.6, 19.0 and 9.6%, respectively. The incidence of resistance to clarithromycin was much higher in isolates from children (44.8%) than adults (14.6%). For metronidazole, the contrary was observed (children: 19.0%, adults: 32.3%). Ciprofloxacin-resistant isolates were all from adult patients. Concerning the adult population, the resistance rate to metronidazole showed a slight increase during the decade, while for clarithromycin and ciprofloxacin a significant increase was observed (4.6 to 22.0% and 0 to 20.9%, respectively). PMID- 11102428 TI - An analysis of antibiotic prescriptions from general dental practitioners in England. AB - The aim of this study was to determine the antibiotics prescribed by general dental practitioners (GDPs). Adult antibiotic prescriptions issued by GDPs from 10 Health Authorities (HAs) in England were analysed. The type of antibiotic prescribed, dose, frequency and duration were investigated. Most of the 17007 prescriptions were for generic antibiotics; nine different antibiotics were prescribed. Many practitioners prescribed antibiotics inappropriately with inconsistent frequency and dose, and for prolonged periods. PMID- 11102429 TI - In vitro activity of gemifloxacin and five other fluoroquinolones against defined isogenic mutants of Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus. PMID- 11102430 TI - The first clinical isolates of Enterococcus faecium with the VanA phenotype in a tertiary Greek hospital. PMID- 11102431 TI - Emergence of teicoplanin-resistant Staphylococcus haemolyticus clinical isolates in Greece. PMID- 11102432 TI - Carbapenem-hydrolysing VIM-2 metallo- beta-lactamase in Pseudomonas aeruginosa from Greece. PMID- 11102433 TI - Laboratory implications of incorrectly taken blood samples for antibiotic assay. PMID- 11102434 TI - delta-aminolaevulinic acid modulates the resistance to fluconazole in a hem1 mutant of Saccharomyces cerevisiae. PMID- 11102435 TI - Antiviral susceptibilities of herpes simplex virus from immunocompetent subjects with recurrent herpes labialis: a UK-based survey. PMID- 11102436 TI - An assessment of the impact of antibiotic resistance in different bacterial species and of the contribution of animal sources to resistance in human infections. PMID- 11102437 TI - A new member of the HCO3(-) transporter superfamily is an apical anion exchanger of beta-intercalated cells in the kidney. AB - The kidneys play pivotal roles in acid-base homeostasis, and the acid-secreting (alpha-type) and bicarbonate-secreting (beta-type) intercalated cells in the collecting ducts are major sites for the final modulation of urinary acid secretion. Since the H(+)-ATPase and anion exchanger activities in these two types of intercalated cells exhibit opposite polarities, it has been suggested that the alpha- and beta-intercalated cells are interchangeable via a cell polarity change. Immunohistological studies, however, have failed to confirm that the apical anion exchanger of beta-intercalated cells is the band 3 protein localized to the basolateral membrane of alpha-intercalated cells. In the present study, we show the evidence that a novel member of the anion exchanger and sodium bicarbonate cotransporter superfamily is an apical anion exchanger of beta intercalated cells. Cloned cDNA from the beta-intercalated cells shows about 30% homology with anion exchanger types 1-3, and functional expression of this protein in COS-7 cells and Xenopus oocytes showed sodium-independent and 4,4' diisothiocyanostilbene-2,2'-disulfonic acid-insensitive anion exchanger activity. Furthermore, immunohistological studies revealed that this novel anion exchanger is present on the apical membrane of beta-intercalated cells, although some beta intercalated cells were negative for AE4 staining. We conclude that our newly cloned transporter is an apical anion exchanger of the beta-intercalated cells, whereas our data do not exclude the possibility that there may be another form of anion exchanger in these cells. PMID- 11102438 TI - The Src kinase p56(lck) up-regulates VLA-4 integrin affinity. Implications for rapid spontaneous and chemokine-triggered T cell adhesion to VCAM-1 and fibronectin. AB - In circulating lymphocytes, the VLA-4 integrin preexists in multiple affinity states that mediate spontaneous tethering, rolling, and arrest on its endothelial ligand, vascular cell adhesion molecule-1 (VCAM-1). The regulation and function of VLA-4 affinity in lymphocytes has never been elucidated. We show here that p56(lck), the major Src kinase in T cells, is a key regulator of high affinity VLA-4. This high affinity is essential for the rapid development of firm adhesion of resting T cells to VCAM-1 and to their extracellular matrix ligand, fibronectin. Lck-regulated VLA-4 function does not require intact TCR nor several key components of the TCR signaling pathway, including ZAP-70 and SLP-76. Furthermore, stimulation of p56(lck) by the phosphatase inhibitor, pervanadate, triggers firm VLA-4-dependent adhesion to VCAM-1. Although Lck is not required for chemokine receptor signaling to mitogen-activated protein kinase, the presence of Lck-regulated high affinity VLA-4 also facilitates firm adhesion triggered by the chemokine, SDF-1, at short-lived contacts. Surprisingly, bond formation rates, ability to tether cells to VLA-4 ligand, and VLA-4 tether bond stability under shear flow are not affected by VLA-4 affinity or Lck activity. Thus, the ability of high affinity VLA-4 to arrest cells on VCAM-1 under flow arises from instantaneous post-ligand strengthening rather than from increased kinetic stability of individual VLA-4 bonds. These results suggest that p56(lck) maintains high affinity VLA-4 on circulating lymphocytes, which determines their ability to strengthen VLA-4 adhesion and rapidly respond to proadhesive chemokine signals at endothelial sites. PMID- 11102439 TI - Regulation of the aldehyde dehydrogenase gene (aldA) and its role in the control of the coinducer level necessary for induction of the ethanol utilization pathway in Aspergillus nidulans. AB - Expression of the structural genes for alcohol and aldehyde dehydrogenase, alcA and aldA, respectively, enables the fungus Aspergillus nidulans to grow on ethanol. The pathway-specific transcriptional activator AlcR mediates the induction of ethanol catabolism in the presence of a coinducing compound. Ethanol catabolism is further subject to negative control mediated by the general carbon catabolite repressor CreA. Here we show that, in contrast to alcA and alcR, the aldA gene is not directly subject to CreA repression. A single cis-acting element mediates AlcR activation of aldA. Furthermore, we show that the induction of the alc gene system is linked to in situ aldehyde dehydrogenase activity. In aldA loss-of-function mutants, the alc genes are induced under normally noninducing conditions. This pseudo-constitutive expression correlates with the nature of the mutations, suggesting that this feature is caused by an intracellular accumulation of a coinducing compound. Conversely, constitutive overexpression of aldA results in suppression of induction in the presence of ethanol. This shows unambiguously that acetaldehyde is the sole physiological inducer of ethanol catabolism. We hypothesize that the intracellular acetaldehyde concentration is the critical factor governing the induction of the alc gene system. PMID- 11102440 TI - Interleukin-3 withdrawal induces an early increase in mitochondrial membrane potential unrelated to the Bcl-2 family. Roles of intracellular pH, ADP transport, and F(0)F(1)-ATPase. AB - Cytokines such as interleukin-3 (IL-3) promote the survival of hematopoietic cells through mechanisms that are not well characterized. Withdrawal of IL-3 from an IL-3-dependent pro-B cell line induced early stress-related events that preceded cell death by more than 40 h. Intracellular pH rose above pH 7.8, peaking 2-3 h post-IL-3 withdrawal, and induced a transient increase in mitochondrial membrane potential (Delta Psi(m)) detected using several different dyes. Similar events were observed following IL-7 withdrawal from a different dependent cell line. Bcl-2, Bax, and caspases were unrelated to these early events. Intracellular alkaline pH inhibited the mitochondrial import of ADP, which would limit ATP synthesis. Total cellular ATP sharply declined during this early period, presumably as a consequence of suppressed ADP import. This was followed by an increase in reduced pyridine nucleotides. The transient increase in Delta Psi(m) was blocked by oligomycin, an inhibitor of F(0)F(1-)ATPase that may have undergone reversal caused by the abnormal ADP-ATP balance within mitochondria. These findings suggest a novel sequence of early events following trophic factor withdrawal in which alkaline pH inhibits ADP import into mitochondria, reversing the F(0)F(1-)ATPase, which in turn consumes ATP and pumps out protons, raising Delta Psi(m). PMID- 11102441 TI - Pro-caspase-8 is predominantly localized in mitochondria and released into cytoplasm upon apoptotic stimulation. AB - The recruitment and cleavage of pro-caspase-8 to produce the active form of caspase-8 is a critical biochemical event in death receptor-mediated apoptosis. However, the source of pro-caspase-8 available for activation by apoptotic triggers is unknown. In human fibroblasts and mouse clonal striatal cells, confocal microscopy revealed that pro-caspase-8 immunofluorescence was colocalized with cytochrome c in mitochondria and was also distributed diffusely in some nuclei. Biochemical analysis of subcellular fractions indicated that pro caspase-8 was enriched in mitochondria and in nuclei. Pro-caspase-8 was found in the intermembrane space, inner membrane, and matrix of mitochondria after limited digestion of mitochondrial fractions, and this distribution was confirmed by immunogold electron microscopy. Pro-caspase-8 and cytochrome c were released from isolated mitochondria that were treated with an inhibitor of the ADP/ATP carrier atractyloside, which opens the mitochondria permeability transition pore. Release was blocked by the mitochondria permeability transition pore inhibitor cyclosporin A (CsA). After clonal striatal cells were exposed for 6 h to an apoptotic inducer tumor necrosis factor-alpha (TNF-alpha), mitochondria immunoreactive for cytochrome c and pro-caspase-8 became clustered at perinuclear sites. Pro-caspase-8 and cytochrome c levels decreased in mitochondrial fractions and increased, along with pro-caspase-8 cleavage products, in the cytoplasm of the TNF-alpha-treated striatal cells. CsA blocked the TNF-alpha-induced release of pro-caspase 8 but not cytochrome c. Internucleosomal DNA fragmentation started at 6 h and peaked 12 h after TNF-alpha treatment. These results suggest that pro caspase-8 is predominantly localized in mitochondria and is released upon apoptotic stimulation through a CsA-sensitive mechanism. PMID- 11102442 TI - Calpain mutants with increased Ca2+ sensitivity and implications for the role of the C(2)-like domain. AB - The ubiquitous calpain isoforms (mu- and m-calpain) are Ca(2+)-dependent cysteine proteases that require surprisingly high Ca(2+) concentrations for activation in vitro ( approximately 50 and approximately 300 microm, respectively). The molecular basis of such a high requirement for Ca(2+) in vitro is not known. In this study, we substantially reduced the concentration of Ca(2+) required for the activation of m-calpain in vitro through the specific disruption of interdomain interactions by structure-guided site-directed mutagenesis. Several interdomain electrostatic interactions involving lysine residues in domain II and acidic residues in the C(2)-like domain III were disrupted, and the effects of these mutations on activity and Ca(2+) sensitivity were analyzed. The mutation to serine of Glu-504, a residue that is conserved in both mu- and m-calpain and interacts most notably with Lys-234, reduced the in vitro Ca(2+) requirement for activity by almost 50%. The mutation of Lys-234 to serine or glutamic acid resulted in a similar reduction. These are the first reported cases in which point mutations have been able to reduce the Ca(2+) requirement of calpain. The structures of the mutants in the absence of Ca(2+) were shown by x-ray crystallography to be unchanged from the wild type, demonstrating that the increase in Ca(2+) sensitivity was not attributable to conformational change prior to activation. The conservation of sequence between mu-calpain, m-calpain, and calpain 3 in this region suggests that the results can be extended to all of these isoforms. Whereas the primary Ca(2+) binding is assumed to occur at EF hands in domains IV and VI, these results show that domain II-domain III salt bridges are important in the process of the Ca(2+)-induced activation of calpain and that they influence the overall Ca(2+) requirement of the enzyme. PMID- 11102443 TI - Stable histone deacetylase complexes distinguished by the presence of SANT domain proteins CoREST/kiaa0071 and Mta-L1. AB - Human histone deacetylases I (HDAC1) and II (HDAC2) are homologous proteins (84% identity) that catalyze release of acetyl groups from modified N-terminal lysines of core histones. Histone deacetylation is correlated with both transient and persistent states of transcriptional inactivity (i.e. silencing) in many eukaryotes. In this study, we analyzed complexes containing HDAC1 and HDAC2 to identify the proteins most stably associated with these deacetylases. Complex cI (9.5 S) contained transcriptional corepressor CoREST/kiaa0071 and a protein homologous to FAD-dependent oxidoreductases, kiaa0601. Complex cII (15 S) contained >/=15 proteins, including CHD3/4 (Mi-2), Mta-L1, RbAp48/46, and MBD3, characteristic of vertebrate nucleosome-remodeling complexes. Under native conditions, cI and cII may contain HDAC1, HDAC2 or both; these can be dissociated to cI and cII core complexes containing only HDAC1 or HDAC2. The (m)CpG-binding protein MBD2 was associated only with the HDAC1 cII core complex. A model is proposed in which HDAC1 core complexes can be targeted to methylated DNA via MBD2 with recruitment of HDAC2 occurring through formation of HDAC1/2 cII dimers. We note that the cI component CoREST/kiaa0071 and the cII component Mta-L1 share a region of homology that includes a SANT domain; this domain may play a role in complex assembly. PMID- 11102444 TI - Paradoxical block of parathormone secretion is mediated by increased activity of G alpha subunits. AB - The paradox of blunted parathormone (PTH) secretion in patients with severe hypomagnesemia has been known for more than 20 years, but the underlying mechanism is not deciphered. We determined the effect of low magnesium on in vitro PTH release and on the signals triggered by activation of the calcium sensing receptor (CaSR). Analogous to the in vivo situation, PTH release from dispersed parathyroid cells was suppressed under low magnesium. In parallel, the two major signaling pathways responsible for CaSR-triggered block of PTH secretion, the generation of inositol phosphates, and the inhibition of cAMP were enhanced. Desensitization or pertussis toxin-mediated inhibition of CaSR stimulated signaling suppressed the effect of low magnesium, further confirming that magnesium acts within the axis CaSR-G-protein. However, the magnesium binding site responsible for inhibition of PTH secretion is not identical with the extracellular ion binding site of the CaSR, because the magnesium deficiency dependent signal enhancement was not altered on CaSR receptor mutants with increased or decreased affinity for calcium and magnesium. By contrast, when the magnesium affinity of the G alpha subunit was decreased, CaSR activation was no longer affected by magnesium. Thus, the paradoxical block of PTH release under magnesium deficiency seems to be mediated through a novel mechanism involving an increase in the activity of G alpha subunits of heterotrimeric G-proteins. PMID- 11102445 TI - Glutathione stimulates sulfated estrogen transport by multidrug resistance protein 1. AB - Multidrug resistance protein 1 (MRP1) is an ATP-binding cassette (ABC) transporter that transports a range of hydrophobic xenobiotics, as well as relatively hydrophilic organic anion conjugates. The protein is present at high levels in testicular Leydig and Sertoli cells. Studies with knockout mice suggest that MRP1 may protect germ cells from exposure to some cytotoxic xenobiotics, but potential endobiotic substrates in this organ have not been identified. Previously, we have shown certain D-ring, but not A-ring, estrogen glucuronides can act as competitive inhibitors of MRP1 mediated transport, suggesting that they are potential substrates for the protein. In the case of 17 beta-estradiol 17 beta-d-glucuronide, this has been confirmed by direct transport studies. The Leydig cell is the major site of estrogen conjugation in the testis. However, the principal products of conjugation are A-ring estrogen sulfates, which are then effluxed from the cell by an unknown transporter. To determine whether MRP1/mrp1 could fulfill this function, we used membrane vesicles from MRP1-transfected HeLa cells to assess this possibility. We found that estradiol and estrone 3-sulfate alone were poor competitors of MRP1-mediated transport of the cysteinyl leukotriene, leukotriene C(4). However, in the presence of reduced glutathione (GSH), their inhibitory potency was markedly increased. Direct transport studies using [(3)H]estrone 3-sulfate confirmed that the conjugated estrogen could be efficiently transported (K(m) = 0.73 microm, V(max) = 440 pmol mg(-)1 protein min(-)1), but only in the presence of either GSH or the nonreducing alkyl derivative, S-methyl GSH. In contrast to previous studies using vincristine as a substrate, we detected no reciprocal increase in MRP1-mediated GSH transport. These results provide the first example of GSH-stimulated, MRP1-mediated transport of a potential endogenous substrate and expand the range of MRP1 substrates whose transport is stimulated by GSH to include certain hydrophilic conjugated endobiotics, in addition to previously identified hydrophobic xenobiotics. PMID- 11102446 TI - Caveolin-1 regulates transforming growth factor (TGF)-beta/SMAD signaling through an interaction with the TGF-beta type I receptor. AB - Transforming growth factor-beta (TGF-beta) signaling proceeds from the cell membrane to the nucleus through the cooperation of the type I and II serine/threonine kinase receptors and their downstream SMAD effectors. Although various regulatory proteins affecting TGF-beta-mediated events have been described, relatively little is known about receptor interactions at the level of the plasma membrane. Caveolae are cholesterol-rich membrane microdomains that, along with their marker protein caveolin-1 (Cav-1), have been implicated in the compartmentalization and regulation of certain signaling events. Here, we demonstrate that specific components of the TGF-beta cascade are associated with caveolin-1 in caveolae and that Cav-1 interacts with the Type I TGF-beta receptor. Additionally, Cav-1 is able to suppress TGF-beta-mediated phosphorylation of Smad-2 and subsequent downstream events. We localize the Type I TGF-beta receptor interaction to the scaffolding domain of Cav-1 and show that it occurs in a physiologically relevant time frame, acting to rapidly dampen signaling initiated by the TGF-beta receptor complex. PMID- 11102448 TI - Intracellular Ca(2+) mobilization and kinase activity during acylated homoserine lactone-dependent quorum sensing in Serratia liquefaciens. AB - Quorum sensing in Gram-negative bacteria involves acylated homoserine lactones (AHLs) and a transcription factor, activated by the AHLs. In this study, a possible involvement of intracellular Ca(2+) as second messenger and/or protein kinase activity during signal transduction is analyzed. When N-hexanoyl-l homoserine lactone was added to a suspension of Fura-2-loaded Serratia liquefaciens, there was a decline in [Ca(2+)](i), measured as a decrease in the Fura-2 fluorescence ratio. As controls, the addition of the signal molecule N-3 oxohexanoyl-l-homoserine lactone, which is not produced by S. liquefaciens, did not induce changes in [Ca(2+)](i). Using a protein kinase activity assay on AHL stimulated cells, an increase in kinase activity after N-butanoyl-l-homoserine lactone stimulation of S. liquefaciens cells was detected, whereas the kinase activity induced by N-3-oxohexanoyl-l-homoserine lactone was not statistically significant. The conclusion from this study is that changes in [Ca(2+)](i) are involved in quorum sensing signal transduction in the Gram-negative bacteria S. liquefaciens. We also conclude that kinase activity is induced in S. liquefaciens upon AHL stimulation. We suggest that the transient intracellular [Ca(2+)] changes and kinase activity, activated by the AHL signal, are critical for the quorum-sensing signal transduction. PMID- 11102447 TI - Inhibition of osteoclast function by adenovirus expressing antisense protein tyrosine kinase 2. AB - Osteoclast activation is initiated by adhesion to bone, cytoskeletal rearrangement, formation of the sealing zone, and formation of the polarized ruffled membrane. Previous findings suggest that protein-tyrosine kinase 2 (PYK2), a cytoplasmic kinase related to focal adhesion kinase, participates in these events. This study examines the role of PYK2 in adhesion-mediated signaling and osteoclast function, using PYK2 antisense. We produced a recombinant adenovirus containing a 300-base pair reversed 5'-coding region of PYK2 and used full-length PYK2 as a control. Murine osteoclast-like cells or their mononuclear precursors were generated in a co-culture of bone marrow and osteoblasts. Infection with antisense adenovirus significantly reduced the expression of endogenous PYK2 protein relative to uninfected cells or to cells infected with sense PYK2 and caused: 1) a reduction in osteoclast formation in vitro; 2) inhibition of cell spreading and of actin ring formation in osteoclasts plated on glass or bone and of attachment and spreading of osteoclast precursors plated on vitronectin; 3) inhibition of bone resorption in vitro; 4) marked reduction in p130(Cas) tyrosine phosphorylation; and 5) no change in alpha(v)beta(3) integrin expression or c-Src tyrosine phosphorylation. Taken together, these findings support the hypothesis that PYK2 plays a central role in the adhesion-dependent cytoskeletal organization and sealing zone formation required for osteoclastic bone resorption. PMID- 11102449 TI - The human origin recognition complex protein 1 dissociates from chromatin during S phase in HeLa cells. AB - We investigated the association of human origin recognition complex (ORC) proteins hOrc1p and hOrc2p with chromatin in HeLa cells. Independent procedures including limited nuclease digestion and differential salt extraction of isolated nuclei showed that a complex containing hOrc1p and hOrc2p occurs in a nuclease resistant compartment of chromatin and can be eluted with moderate high salt concentrations. A second fraction of hOrc2p that dissociates in vitro at low salt conditions was found to occur in a chromatin compartment characterized by its high accessibility to micrococcal nuclease. Functional differences between these two sites become apparent in HeLa cells that synchronously enter the S phase after a release from a double-thymidine block. The hOrc1p/hOrc2p-containing complexes dissociate from their chromatin sites during S phase and reassociate at the end of mitosis. In contrast, the fraction of hOrc2p in nuclease-accessible, more open chromatin remains bound during all phases of the cell cycle. We propose that the hOrc1p/hOrc2p-containing complexes are components of the human origin recognition complex. Thus, the observed cell cycle-dependent release of the hOrc1p/hOrc2p-containing complexes is in line with previous studies with Xenopus and Drosophila systems, which indicated that a change in ORC stability occurs after prereplication complex formation. This could be a powerful mechanism that prevents the rereplication of already replicated chromatin in the metazoan cell cycle. PMID- 11102450 TI - Molecular mechanism for functional interaction between DnaA protein and acidic phospholipids: identification of important amino acids. AB - DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli, seems to be reactivated from the ADP-bound form to its ATP-bound form through stimulation of ADP release by acidic phospholipids such as cardiolipin. We previously reported that two potential amphipathic helices (Lys-327 to Ile-344 and Asp-357 to Val-374) of DnaA protein are involved in the functional interaction between DnaA and cardiolipin. In relation to one of these helices (Asp-357 to Val-374), we demonstrated that basic amino acids in the helix, especially Lys-372, are vital for this interaction. In this study, we have identified an amino acid in the second potential amphipathic helix (Lys-327 to Ile-344), which would also appear to be involved in the interaction. We constructed three mutant dnaA genes with a single mutation (dnaAR328E, dnaAR334E, and dnaAR342E) and examined the function of the mutant proteins. DnaAR328E, but not DnaAR334E and DnaAR342E, was found to be more resistant to inhibition of its ATP binding activity by cardiolipin than the wild-type protein. The stimulation of ADP release from DnaAR328E by cardiolipin was also weaker than that observed with the other mutants and the wild-type protein. These results suggest that Arg 328 of DnaA protein is involved in the functional interaction of this protein with acidic phospholipids. We propose that acidic phospholipids bind to two basic amino acid residues (Arg-328 and Lys-372) of DnaA protein and change the higher order structure of its ATP-binding pocket, which in turn stimulates the release of ADP from the protein. PMID- 11102451 TI - Modulation of circulating leptin levels by its soluble receptor. AB - Leptin is an adipocyte-derived hormone with potent weight reducing effects. Genetically obese rodents with mutations of leptin or the leptin receptor are defective in leptin signaling and develop morbid obesity and diabetes. Interestingly, the levels of both leptin mRNA and protein are increased by up to 20-fold in these animals, suggesting the existence of a feedback mechanism controlling the amount of leptin in circulation. In this report, we attempted to determine whether the up-regulation of circulating leptin in Zucker Diabetic Fatty rats, which are nonresponsive to leptin due to a receptor point mutation, is entirely due to increased expression of leptin. We demonstrate that the high level of circulating leptin in these rats is attributable to at least two factors: increased leptin expression by the adipose tissue and delayed clearance of leptin from circulation due to binding to its soluble receptor. The latter conclusion was supported by three lines of evidence: 1) The soluble leptin receptor is up-regulated by about 20-fold in Zucker Diabetic Fatty rats; 2) Adenovirus-mediated overexpression of the soluble leptin receptor results in a similar -fold increase of circulating leptin; 3) In ob/ob mice, which have no endogenous leptin, exogenously administered leptin reaches a higher level when the soluble leptin receptor is overexpressed. The weight-reducing effect of leptin is enhanced in C57Bl/6 ob/ob mice with overexpression of the soluble leptin receptor. Soluble leptin receptor may be a significant factor determining the amount of total leptin in circulation. PMID- 11102452 TI - Familial hypertrophic cardiomyopathy mutations in the regulatory light chains of myosin affect their structure, Ca2+ binding, and phosphorylation. AB - The effect of the familial hypertrophic cardiomyopathy mutations, A13T, F18L, E22K, R58Q, and P95A, found in the regulatory light chains of human cardiac myosin has been investigated. The results demonstrate that E22K and R58Q, located in the immediate extension of the helices flanking the regulatory light chain Ca(2+) binding site, had dramatically altered Ca(2+) binding properties. The K(Ca) value for E22K was decreased by approximately 17-fold compared with the wild-type light chain, and the R58Q mutant did not bind Ca(2+). Interestingly, Ca(2+) binding to the R58Q mutant was restored upon phosphorylation, whereas the E22K mutant could not be phosphorylated. In addition, the alpha-helical content of phosphorylated R58Q greatly increased with Ca(2+) binding. The A13T mutation, located near the phosphorylation site (Ser-15) of the human cardiac regulatory light chain, had 3-fold lower K(Ca) than wild-type light chain, whereas phosphorylation of this mutant increased the Ca(2+) affinity 6-fold. Whereas phosphorylation of wild-type light chain decreased its Ca(2+) affinity, the opposite was true for A13T. The alpha-helical content of the A13T mutant returned to the level of wild-type light chain upon phosphorylation. The phosphorylation and Ca(2+) binding properties of the regulatory light chain of human cardiac myosin are important for physiological function, and alteration any of these could contribute to the development of hypertrophic cardiomyopathy. PMID- 11102453 TI - Arresting pore formation of a cholesterol-dependent cytolysin by disulfide trapping synchronizes the insertion of the transmembrane beta-sheet from a prepore intermediate. AB - Perfringolysin O (PFO), a member of the cholesterol-dependent cytolysin family of pore-forming toxins, forms large oligomeric complexes comprising up to 50 monomers. In the present study, a disulfide bridge was introduced between cysteine-substituted serine 190 of transmembrane hairpin 1 (TMH1) and cysteine substituted glycine 57 of domain 2 of PFO. The resulting disulfide-trapped mutant (PFO(C190-C57)) was devoid of hemolytic activity and could not insert either of its transmembrane beta-hairpins (TMHs) into the membrane unless the disulfide was reduced. Both the size of the oligomer formed on the membrane and its rate of formation were unaffected by the oxidation state of the Cys(190)-Cys(57) disulfide bond; thus, the disulfide-trapped PFO was assembled into a prepore complex on the membrane. The conversion of this prepore to the pore complex was achieved by reducing the C190-C57 disulfide bond. PFO(C190-C57) that was allowed to form the prepore prior to the reduction of the disulfide exhibited a dramatic increase in the rate of PFO-dependent hemolysis and the membrane insertion of its TMHs when compared with toxin that had the disulfide reduced prior mixing the toxin with membranes. Therefore, the rate-limiting step in pore formation is prepore assembly, not TMH insertion. These data demonstrate that the prepore is a legitimate intermediate during the insertion of the large transmembrane beta sheet of the PFO oligomer. Finally, the PFO TMHs do not appear to insert independently, but instead their insertion is coupled. PMID- 11102454 TI - Ligand- and coactivator-mediated transactivation function (AF2) of the androgen receptor ligand-binding domain is inhibited by the cognate hinge region. AB - Transactivation functions (AF2) in the ligand-binding domains (LBD) of many steroid receptors are well characterized, but there is little evidence to support such a function for the LBD of the androgen receptor (AR). We report a mutant AR, with residues 628-646 in the hinge region deleted, which exhibited transactivation activity that was more than double that of the wild type (WT) AR. Although no androgen-dependent AF2 activity could be observed for the WT ARLBD fused to a heterologous DNA-binding domain, the mutant ARLBD(Delta628-646) was 30 40 times more active than the WT ARLBD. In the presence of the p160 coactivator TIF2, AR(Delta628-646) was significantly more active than similarly treated WT AR. Deletion of residues 628-646 also enhanced TIF2-ARLBD activity 8-fold, an effect not present when the LBD-interacting LXXLL motifs of TIF2 were mutated, suggesting that the negative modulatory activity of residues 628-646 were exerted via coactivator pathways. Although the AP-1 (c-Jun/c-Fos) system and NcoR have been reported to interact with and repress the activity of some steroid receptors, c-Jun, c-Fos, c-Jun/c-Fos, nor NcoR function was consistently affected by the absence or presence of residues 628-646, implying that the AR hinge region exerts its silencing effects in a manner independent of these corepressors. Our data provide evidence for the novel finding that strong androgen-dependent AF2 exists in the ARLBD and is the first report of a negative regulatory domain in the AR. Because mutations in this region are commonly associated with prostate cancer, it is important to characterize the mechanisms by which the hinge region exerts its repressor effect on ligand-activated and coactivator-mediated AF2 activity of the ARLBD. PMID- 11102455 TI - Polymerization of plasminogen activator inhibitor-1. AB - The activity of the serine proteinase inhibitor (serpin) plasminogen activator inhibitor-1 (PAI-1) is controlled by the intramolecular incorporation of the reactive loop into beta-sheet A with the generation of an inactive latent species. Other members of the serpin superfamily can be pathologically inactivated by intermolecular linkage between the reactive loop of one molecule and beta-sheet A of a second to form chains of polymers associated with diverse diseases. It has long been believed that PAI-1 is unique among active serpins in that it does not form polymers. We show here that recombinant native and latent PAI-1 spontaneously form polymers in vitro at low pH although with distinctly different electrophoretic patterns of polymerization. The polymers of both the native and latent species differ from the typical loop-A-sheet polymers of other serpins in that they readily dissociate back to their original monomeric form. The findings with PAI-1 are compatible with different mechanisms of linkage, each involving beta-strand addition of the reactive loop to s7A in native PAI-1 and to s1C in latent PAI-1. Glycosylated native and latent PAI-1 can also form polymers under similar conditions, which may be of in vivo importance in the low pH environment of the platelet. PMID- 11102456 TI - Functional roles of the conserved threonine 250 in the target recognition domain of HhaI DNA methyltransferase. AB - DNA cytosine-5-methyltransferase HhaI recognizes the GCGC sequence and flips the inner cytosine out of DNA helix and into the catalytic site for methylation. The 5'-phosphate of the flipped out cytosine is in contact with the conserved Thr-250 from the target recognition domain. We have produced 12 mutants of Thr-250 and examined their methylation potential in vivo. Six active mutants were subjected to detailed biochemical and structural studies. Mutants with similar or smaller side chains (Ser, Cys, and Gly) are very similar to wild-type enzyme in terms of steady-state kinetic parameters k(cat), K(m)(DNA), K(m)(AdoMet). In contrast, the mutants with bulkier side chains (Asn, Asp, and His) show increased K(m) values for both substrates. Fluorescence titrations and stopped-flow kinetic analysis of interactions with duplex oligonucleotides containing 2-aminopurine at the target base position indicate that the T250G mutation leads to a more polar but less solvent-accessible position of the flipped out target base. The x-ray structure of the ternary M. HhaI(T250G).DNA.AdoHcy complex shows that the target cytosine is locked in the catalytic center of enzyme. The space created by the mutation is filled by water molecules and the adjacent DNA backbone atoms dislocate slightly toward the missing side chain. In aggregate, our results suggest that the side chain of Thr-250 is involved in constraining the conformation the DNA backbone and the target base during its rotation into the catalytic site of enzyme. PMID- 11102458 TI - Neurobeachin: A protein kinase A-anchoring, beige/Chediak-higashi protein homolog implicated in neuronal membrane traffic. AB - We describe the identification and initial characterization of neurobeachin, a neuron-specific multidomain protein of 327 kDa with a high-affinity binding site (K(d), 10 nm) for the type II regulatory subunit of protein kinase A (PKA RII). Neurobeachin is peripherally associated with pleomorphic tubulovesicular endomembranes near the trans sides of Golgi stacks and throughout the cell body and cell processes. It is also found in a subpopulation of synapses, where it is concentrated at the postsynaptic plasma membrane. In live cells, perinuclear neurobeachin is dispersed by brefeldin A (BFA) within 1 min, and in permeabilized cells a recruitment of neurobeachin from cytosol to Golgi-near membranes is stimulated by GTPgammaS and prevented by brefeldin A. Spots of neurobeachin recruitment are close to but distinct from recruitment sites of COP-I, AP-1, and AP-3 coat proteins involved in vesicle budding. These observations indicate that neurobeachin binding to membranes close to the trans-Golgi requires an ADP ribosylation factor-like GTPase, possibly in association with a novel type of protein coat. A neurobeachin isoform that does not bind RII, beige-like protein (BGL), is expressed in many tissues. Neurobeachin, BGL, and approximately 10 other mammalian gene products share a characteristic C-terminal BEACH-WD40 sequence module, which is also present in gene products of invertebrates, plants, protozoans, and yeasts, thus defining a new protein family. The prototype member of this family of BEACH domain proteins, lysosomal trafficking regulator (LYST), is deficient in genetic defects of protein sorting in lysosome biogenesis (the beige mouse and Chediak-Higashi syndrome). Neurobeachin's subcellular localization, its coat protein-like membrane recruitment, and its sequence similarity to LYST suggest an involvement in neuronal post-Golgi membrane traffic, one of its functions being to recruit protein kinase A to the membranes with which it associates. PMID- 11102459 TI - The status of voltage-dependent calcium channels in alpha 1E knock-out mice. AB - It has been hypothesized that R-type Ca currents result from the expression of the alpha(1E) gene. To test this hypothesis we examined the properties of voltage dependent Ca channels in mice in which the alpha(1E) Ca channel subunit had been deleted. Application of omega-conotoxin GVIA, omega-agatoxin IVA, and nimodipine to cultured cerebellar granule neurons from wild-type mice inhibited components of the whole-cell Ba current, leaving a "residual" R current with an amplitude of approximately 30% of the total Ba current. A minor portion of this R current was inhibited by the alpha(1E)-selective toxin SNX-482, indicating that it resulted from the expression of alpha(1E). However, the majority of the R current was not inhibited by SNX-482. The SNX-482-sensitive portion of the granule cell R current was absent from alpha(1E) knock-out mice. We also identified a subpopulation of dorsal root ganglion (DRG) neurons from wild-type mice that expressed an SNX-482 sensitive component of the R current. However as with granule cells, most of the DRG R current was not blocked by SNX-482. We conclude that there exists a component of the R current that results from the expression of the alpha(1E) Ca channel subunit but that the majority of R currents must result from the expression of other Ca channel alpha subunits. PMID- 11102460 TI - Phase shifting the retinal circadian clock: xPer2 mRNA induction by light and dopamine. AB - A circadian clock is located in the retinal photoreceptors of the African clawed frog Xenopus laevis. These photoreceptor clocks are thought to govern a wide variety of output rhythms, including melatonin release and gene expression. Both light and dopamine phase shift the retinal clock in a phase-dependent manner. Two homologs of the Drosophila period gene have been cloned in Xenopus, and one of these (xPer2) is acutely regulated by light. Light and dopamine induce xPer2 mRNA in a similar manner. In addition, the increase of xPer2 mRNA in response to light and dopamine is the same at all times of day tested. In contrast, xPer1 mRNA exhibits circadian oscillations but is relatively insensitive to phase-shifting treatments of light or dopamine. Our data suggest that xPer2 functions as the molecular link between the light/dark cycle and the circadian clock. PMID- 11102461 TI - Postsynaptic signaling via the [mu]-opioid receptor: responses of dorsal horn neurons to exogenous opioids and noxious stimulation. AB - Although both pre- and postsynaptic mechanisms have been implicated in the analgesia produced by mu-opioids at the spinal cord, it is not known under what conditions these different controls come into play. Because the mu-opioid receptor (MOR) can be visualized in individual lamina II excitatory interneurons and internalizes into endosomes on ligand binding, we tested whether MOR internalization could be monitored and used to measure postsynaptic MOR signaling. To test whether endogenous opioids modulate these lamina II interneurons during noxious stimulation, we next assessed the magnitude of postsynaptic MOR internalization under a variety of nociceptive conditions. As observed in other systems, we show that MOR internalization in dorsal horn interneurons is demonstrated readily in response to opioid ligands. The MOR internalization is dose-dependent, with a similar dose-response to that observed for opioid-induced increases in potassium conductance. We demonstrate that MOR internalization in lamina II neurons correlates precisely with the extent of analgesia produced by intrathecal DAMGO. These results suggest that MOR internalization provides a good marker of MOR signaling in the spinal cord and that postsynaptic MORs on lamina II interneurons likely participate in the analgesia that is produced by exogenous opioids. We found, however, that noxious stimuli, under normal or inflammatory conditions, did not induce MOR internalization. Thus, endogenous enkephalins and endomorphins, thought to be released during noxious peripheral stimuli, do not modulate nociceptive messages via postsynaptic MORs on lamina II interneurons. We suggest that any endogenous opioids that are released by noxious stimuli target presynaptic MORs or delta opioid receptors. PMID- 11102462 TI - Independence of and interactions between GABA-, glutamate-, and acetylcholine activated Cl conductances in Aplysia neurons. AB - In certain Aplysia neurons, glutamate, GABA, and acetylcholine (ACh) all elicit desensitizing Cl-dependent responses. This fact and the finding that the glutamate and GABA responses "cross-desensitize" led to the suggestion (Swann and Carpenter, 1975; King and Carpenter, 1987) that the responses to these transmitters were mediated by the same receptor-channel complex. This hypothesis is incompatible with the demonstration given here that the GABA- and glutamate gated channels are clearly distinct; the GABA channel, but not the glutamate channel, shows outward rectification (Matsumoto, 1982; King and Carpenter, 1987, 1989) and is selectively blocked by intracellular sulfate. Exploiting these distinctive characteristics and the independent expression of the receptors in some cells, we have been able to reevaluate the so-called cross-desensitization by analyzing the ability of GABA, glutamate, and other agonists to interact with each of the receptor molecules. The cross-desensitization was found to be exclusively attributable to the ability of GABA to interact with the glutamate receptor (Oyama et al., 1990). The GABA receptor is unaffected by glutamate. Nevertheless, in cells expressing both receptors, glutamate can reduce the GABA response by auto-desensitizing the part of the response that is mediated by the glutamate receptor. No interactions were observed between ACh-induced responses and either of the responses elicited by the amino acids. The invertebrate glutamate-gated Cl channels that have been cloned resemble the vertebrate glycine receptor (Vassilatis et al., 1997). Our pharmacological evaluation of the molluscan glutamate receptor points in the same direction. PMID- 11102463 TI - Growth/differentiation factor-15/macrophage inhibitory cytokine-1 is a novel trophic factor for midbrain dopaminergic neurons in vivo. AB - Transforming growth factor-betas (TGF-betas) constitute an expanding family of multifunctional cytokines with prominent roles in development, cell proliferation, differentiation, and repair. We have cloned, expressed, and raised antibodies against a distant member of the TGF-betas, growth/differentiation factor-15 (GDF-15). GDF-15 is identical to macrophage inhibitory cytokine-1 (MIC 1). GDF-15/MIC-1 mRNA and protein are widely distributed in the developing and adult CNS and peripheral nervous systems, including choroid plexus and CSF. GDF 15/MIC-1 is a potent survival promoting and protective factor for cultured and iron-intoxicated dopaminergic (DAergic) neurons cultured from the embryonic rat midbrain floor. The trophic effect of GDF-15/MIC-1 was not accompanied by an increase in cell proliferation and astroglial maturation, suggesting that GDF 15/MIC-1 probably acts directly on neurons. GDF-15/MIC-1 also protects 6 hydroxydopamine (6-OHDA)-lesioned nigrostriatal DAergic neurons in vivo. Unilateral injections of GDF-15/MIC-1 into the medial forebrain bundle just above the substantia nigra (SN) and into the left ventricle (20 microgram each) immediately before a 6-OHDA injection (8 microgram) prevented 6-OHDA-induced rotational behavior and significantly reduced losses of DAergic neurons in the SN. This protection was evident for at least 1 month. Administration of 5 microgram of GDF-15/MIC-1 in the same paradigm also provided significant neuroprotection. GDF-15/MIC-1 also promoted the serotonergic phenotype of cultured raphe neurons but did not support survival of rat motoneurons. Thus, GDF 15/MIC-1 is a novel neurotrophic factor with prominent effects on DAergic and serotonergic neurons. GDF-15/MIC-1 may therefore have a potential for the treatment of Parkinson's disease and disorders of the serotonergic system. PMID- 11102464 TI - Estrogen is essential for maintaining nigrostriatal dopamine neurons in primates: implications for Parkinson's disease and memory. AB - There are sexual differences in several parameters of the nigrostriatal dopamine neurons, as well as in the progression of diseases associated with this system, e.g., Parkinson's disease and dementia. These differences, as well as direct experimental data in rodents, suggest that gonadal hormones play a role in modulating this system. To determine whether circulating estrogen might have long term effects by altering the number of dopamine neurons, the density of dopamine neurons was calculated in the compact zone of the substantia nigra of male and intact female short- (10 d) and longer-term (30 d) ovariectomized and short- and longer-term ovariectomized but estrogen-replaced nonhuman primates (African green monkeys). Furthermore, the number of tyrosine hydroxylase-expressing neurons, the total number of all types of neurons, and the volume of the compact zone of the substantia nigra were calculated in 30 d ovariectomized and in 30 d ovariectomized and estrogen-replaced monkeys. Unbiased stereological analyses demonstrated that a 30 d estrogen deprivation results in an apparently permanent loss of >30% of the total number of substantia nigra dopamine cells. Furthermore, the density calculations showed that brief estrogen replacement restores the density of tyrosine hydroxylase-immunoreactive cells after a 10 d, but not after a 30 d, ovariectomy. Moreover, the density of dopamine cells is higher in females than in males. These observations show the essential role of estrogen in maintaining the integrity of the nigral dopamine system, suggest a new treatment strategy for patients with Parkinson's disease and with certain forms of memory impairing disorders, and provide another rationale for estrogen replacement therapy for postmenopausal women. PMID- 11102465 TI - The human skeletal muscle Na channel mutation R669H associated with hypokalemic periodic paralysis enhances slow inactivation. AB - Missense mutations of the human skeletal muscle voltage-gated Na channel (hSkM1) underlie a variety of diseases, including hyperkalemic periodic paralysis (HyperPP), paramyotonia congenita, and potassium-aggravated myotonia. Another disorder of sarcolemmal excitability, hypokalemic periodic paralysis (HypoPP), which is usually caused by missense mutations of the S4 voltage sensors of the L type Ca channel, was associated recently in one family with a mutation in the outermost arginine of the IIS4 voltage sensor (R669H) of hSkM1 (Bulman et al., 1999). Intriguingly, an arginine-to-histidine mutation at the homologous position in the L-type Ca(2+) channel (R528H) is a common cause of HypoPP. We have studied the gating properties of the hSkM1-R669H mutant Na channel experimentally in human embryonic kidney cells and found that it has no significant effects on activation or fast inactivation but does cause an enhancement of slow inactivation. R669H channels exhibit an approximately 10 mV hyperpolarized shift in the voltage dependence of slow inactivation and a twofold to fivefold prolongation of recovery after prolonged depolarization. In contrast, slow inactivation is often disrupted in HyperPP-associated Na channel mutants. These results demonstrate that, in R669H-associated HypoPP, enhanced slow inactivation does not preclude, and may contribute to, prolonged attacks of weakness and add support to previous evidence implicating the IIS4 voltage sensor in slow inactivation gating. PMID- 11102466 TI - Zinc inhibits miniature GABAergic currents by allosteric modulation of GABAA receptor gating. AB - Zinc is abundantly present in the CNS, and after nerve stimulation is thought to be released in sufficient quantity to modulate the synaptic transmission. Although it is known that this divalent cation inhibits the GABAergic synaptic currents, the underlying mechanisms were not fully elucidated. Here we report that zinc reduced the amplitude, slowed the rise time, and accelerated the decay of mIPSCs in cultured hippocampal neurons. The analysis of current responses to rapid GABA applications and model simulations indicated that these effects on mIPSCs are caused by zinc modulation of GABA(A) receptor gating. In particular, zinc slowed the onset of GABA-evoked currents by decreasing both the binding (k(on)) and the transition rate from closed to open state (beta(2)). Moreover, slower onset and recovery from desensitization as well as an increased unbinding rate (k(off)) were shown to underlie the accelerated deactivation kinetics in the presence of zinc. The nonequilibrium conditions of GABA(A) receptor activation were found to strongly affect zinc modulation of this receptor. In particular, an extremely fast clearance of synaptic GABA is implicated to be responsible for a stronger zinc effect on mIPSCs than on current responses to exogenous GABA. Finally, the analysis of currents evoked by GABA coapplied with zinc indicated that the interaction between zinc and GABA(A) receptors was too slow to explain zinc effects in terms of competitive antagonism. In conclusion, our results provide evidence that inhibition of mIPSCs by zinc is attributable to the allosteric modulation of GABA(A) receptor gating. PMID- 11102467 TI - Role of Ca2+ stores in metabotropic L-glutamate receptor-mediated supralinear Ca2+ signaling in rat hippocampal neurons. AB - The role of metabotropic l-glutamate (mGlu) receptors in supralinear Ca(2+) signaling was investigated in cultured hippocampal cells using Ca(2+) imaging techniques and whole-cell voltage-clamp recording. In neurons, but not glia, global supralinear Ca(2+) release from intracellular stores was observed when the mGlu receptor agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) was combined with elevated extracellular K(+) levels (10.8 mm), moderate depolarization (15-30 mV), or NMDA (3 micrometer). There was a delay (2-8 min) before the stores were fully charged, and the enhancement persisted for a short period (up to 10 min) after removal of the store-loading stimulus. Studies with the mGlu receptor antagonist 2-methyl-6-(phenylethynyl)-pyridine demonstrated that these effects were mediated by activation of the mGlu(5) receptor subtype. The L-type voltage-gated Ca(2+) channel antagonist nifedipine (10 micrometer) substantially reduced responses to DHPG obtained in the presence of elevated extracellular K(+) but not NMDA. This suggests that the Ca(2+) that is required to load the stores can enter either through L-type voltage-gated Ca(2+) channels or directly through NMDA receptors. The findings that both depolarization and NMDA receptor activation can facilitate mGlu receptor Ca(2+) signaling adds considerable flexibility to the processes that underlie activity-dependent changes in synaptic strength. In particular, a temporal separation between the store-loading stimulus and the activation of mGlu receptors could be used as a recency detector in neurons. PMID- 11102468 TI - Induction of interleukin-6 by depolarization of neurons. AB - Interleukin-6 (IL-6) has neuromodulatory and neuroprotective effects in vivo. It is expressed in glial cells and neurons both under physiological conditions and in various neurological diseases. Although the expression of IL-6 in glia has been intensely investigated, little is known about the regulation of IL-6 production by neurons. Therefore, we investigated the regulation of IL-6 expression in neurons. Membrane depolarization raised IL-6 mRNA accumulation in primary cortical cells and the PC-12 cell line. In vivo, IL-6 mRNA in the brain increased significantly after epileptic seizures. To investigate IL-6 gene transcription, PC-12 cells were transfected with reporter gene constructs containing the human IL-6 promoter. Membrane depolarization raised IL-6 transcription twofold to fourfold. This increase could be blocked by lowering extracellular Ca(2+) levels or by inhibiting L-type Ca(2+) channels or Ca(2+)/calmodulin-dependent protein kinases. Internal mutations in various elements of the IL-6 promoter revealed the glucocorticoid response element (GRE) 2 to be a depolarization-responsive element. Although the GRE2 bound the glucocorticoid receptor (GR) and was stimulated by dexamethasone, the GR was not responsible for the effect of membrane depolarization because a consensus GRE did not mediate stimulation by membrane depolarization. Instead, another yet undefined factor that binds to the IL-6 GRE2 may mediate the response to membrane depolarization. These data demonstrate that the expression of IL-6 in neurons is regulated by membrane depolarization and suggest a novel Ca(2+)-responsive promoter element. Through this mechanism, IL-6 may function as a neuromodulator induced by neuronal activity. PMID- 11102469 TI - GABAC receptor sensitivity is modulated by interaction with MAP1B. AB - GABA(C) receptors contain rho subunits and mediate feedback inhibition from retinal amacrine cells to bipolar cells. We previously identified the cytoskeletal protein MAP1B as a rho1 subunit anchoring protein. Here, we analyze the structural basis and functional significance of the MAP1B-rho1 interaction. Twelve amino acids at the C terminus of the large intracellular loop of rho1 (and also rho2) are sufficient for interaction with MAP1B. Disruption of the MAP1B-rho interaction in bipolar cells in retinal slices decreased the EC(50) of their GABA(C) receptors, doubling the receptors' current at low GABA concentrations without affecting their maximum current at high concentrations. Thus, anchoring to the cytoskeleton lowers the sensitivity of GABA(C) receptors and provides a likely site for functional modulation of GABA(C) receptor-mediated inhibition. PMID- 11102470 TI - GABA spillover from single inhibitory axons suppresses low-frequency excitatory transmission at the cerebellar glomerulus. AB - GABA type B receptors (GABA(B)-Rs) are present on excitatory terminals throughout the CNS, but surprisingly little is known about their role in modulating neurotransmission under physiological conditions. We have investigated activation of GABA(B)-Rs on excitatory terminals within the cerebellar glomerulus, a structure where glutamatergic excitatory and GABAergic inhibitory terminals are in close apposition and make axodendritic synapses onto granule cells. Application of the GABA(B)-R agonist baclofen depressed evoked mossy fiber EPSCs by 54% at 1 Hz. The amplitude of miniature EPSCs recorded in tetrodotoxin was unchanged in the presence of baclofen, but the frequency was significantly reduced, indicating a purely presynaptic action of baclofen under our recording conditions. At physiological temperature (37 degrees C) presynaptic GABA(B)-Rs were not tonically activated by spontaneous GABA release from Golgi cells, which fire at approximately 8 Hz in slices at this temperature. However, tonic activation could be induced by blocking GABA uptake or by lowering temperature. GABA(B)-Rs were activated at physiological temperature when Golgi cell firing was increased above the basal level by stimulating a single inhibitory Golgi cell input at 50 Hz, suppressing the mossy fiber-evoked EPSC by 24% at 1 Hz. Furthermore, glutamate release was selectively inhibited at low-frequency mossy fiber inputs (<10 Hz) during Golgi cell stimulation. Our findings suggest that GABA spillover in the glomerulus modulates sensory input to the cerebellar cortex. PMID- 11102471 TI - Decreases in endogenous opioid peptides in the rat medullo-coerulear pathway after chronic morphine treatment. AB - Several biochemical changes have been described in noradrenergic neurons of the locus coeruleus (LC) after chronic morphine treatment. Changes in neurochemical expression in opioid afferent projections to the LC may be equally important in modulating noradrenergic neurons during chronic opiate exposure. To test the hypothesis that opioid peptides in LC afferents are altered after chronic opiate administration, we exposed adult male rats to either morphine or placebo pellets for 5 d. Tissue sections through the LC were processed for peroxidase or gold silver labeling of methionine(5)-enkephalin (met-ENK) and analyzed using light or electron microscopy, respectively. Light level densitometry and ultrastructural analysis showed that there was a significant decrease in immunolabeling for ENK in LC-afferent terminals of morphine-treated rats. Western immunoblot analysis confirmed that protein levels for both leucine(5)- and methionine(5)-ENK were significantly decreased in tissue samples containing the LC after chronic morphine treatment. To test whether decreases in ENK protein expression were mirrored by decreases in gene expression, Northern blot analysis of preproenkephalin (PPE) mRNA was conducted in tissue samples obtained through the medulla, a brainstem area that contains the major opioid afferents to the LC. PPE mRNA was reduced in samples obtained from morphine-treated rats. Finally, in situ hybridization experiments confirmed significant decreases in PPE mRNA expression in the nucleus paragigantocellularis, a region known to provide a robust opioid input to the LC. These data suggest that there is a decrease in the synthesis of the opioid peptide mRNA and protein in the medullo-coerulear pathway after chronic exposure to morphine. Such alterations in opioid peptide levels during opiate dependence may contribute to the observed hyperactivity of LC neurons during opiate withdrawal. PMID- 11102472 TI - A conserved clathrin assembly motif essential for synaptic vesicle endocytosis. AB - Although clathrin assembly by adaptor proteins (APs) plays a major role in the recycling of synaptic vesicles, the molecular mechanism that allows APs to assemble clathrin is poorly understood. Here we demonstrate that AP180, like AP-2 and AP-3, binds to the N-terminal domain of clathrin. Sequence analysis reveals a motif, containing the sequence DLL, that exists in multiple copies in many clathrin APs. Progressive deletion of these motifs caused a gradual reduction in the ability of AP180 to assemble clathrin in vitro. Peptides from AP180 or AP-2 containing this motif also competitively inhibited clathrin assembly by either protein. Microinjection of these peptides into squid giant presynaptic terminals reversibly blocked synaptic transmission and inhibited synaptic vesicle endocytosis by preventing coated pit formation at the plasma membrane. These results indicate that the DLL motif confers clathrin assembly properties to AP180 and AP-2 and, perhaps, to other APs. We propose that APs promote clathrin assembly by cross-linking clathrin triskelia via multivalent interactions between repeated DLL motifs in the APs and complementary binding sites on the N-terminal domain of clathrin. These results reveal the structural basis for clathrin assembly and provide novel insights into the molecular mechanism of clathrin mediated synaptic vesicle endocytosis. PMID- 11102473 TI - Dopamine D3 receptors expressed by all mesencephalic dopamine neurons. AB - A polyclonal antibody was generated using synthetic peptides designed in a specific sequence of the rat D(3) receptor (D(3)R). Using transfected cells expressing recombinant D(3)R, but not D(2) receptor, this antibody labeled 45-80 kDa species in Western blot analysis, immunoprecipitated a soluble fraction of [(125)I]iodosulpride binding, and generated immunofluorescence, mainly in the cytoplasmic perinuclear region of the cells. In rat brain, the distribution of immunoreactivity matched that of D(3)R binding, revealed using [(125)I]R(+)trans 7-hydroxy-2-[N-propyl-N-(3'-iodo-2'-propenyl)amino] tetralin ([(125)I]7-trans-OH PIPAT), with dense signals in the islands of Calleja and mammillary bodies, and moderate to low signals in the shell of nucleus accumbens (AccSh), frontoparietal cortex, substantia nigra (SN), ventral tegmental area (VTA) and lobules 9 and 10 of the cerebellum. Very low or no signals could be detected in other rat brain regions, including dorsal striatum, or in D(3)R-deficient mouse brain. Labeling of perikarya of AccSh and SN/VTA appeared with a characteristic punctuate distribution, mostly at the plasma membrane where it was not associated with synaptic boutons, as revealed by synaptophysin immunoreactivity. In SN/VTA, D(3)R immunoreactivity was found on afferent terminals, arising from AccSh, in which destruction of intrinsic neurons by kainate infusions produced a loss of D(3)R binding in both AccSh and SN/VTA. D(3)R-immunoreactivity was also found in all tyrosine hydroxylase (TH)-positive neurons observed in SN, VTA and A8 retrorubral fields, where it could represent D(3) autoreceptors controlling dopamine neuron activities, in agreement with the elevated dopamine extracellular levels in projection areas of these neurons found in D(3)R-deficient mice. PMID- 11102474 TI - alpha-Latrotoxin releases calcium in frog motor nerve terminals. AB - alpha-Latrotoxin (alpha-LTX) is a neurotoxin that accelerates spontaneous exocytosis independently of extracellular Ca(2+). Although alpha-LTX increases spontaneous transmitter release at synapses, the mechanism is unknown. We tested the hypothesis that alpha-LTX causes transmitter release by mobilizing intracellular Ca(2+) in frog motor nerve terminals. Transmitter release was measured electrophysiologically and with the vesicle marker FM1-43; presynaptic ion concentration dynamics were measured with fluorescent ion-imaging techniques. We report that alpha-LTX increases transmitter release after release of a physiologically relevant concentration of intracellular Ca(2+). Neither the blockade of Ca(2+) release nor the depletion of Ca(2+) from endoplasmic reticulum affected Ca(2+) signals produced by alpha-LTX. The Ca(2+) source is likely to be mitochondria, because the effects on Ca(2+) mobilization of CCCP (which depletes mitochondrial Ca(2+)) and of alpha-LTX are mutually occlusive. The release of mitochondrial Ca(2+) is partially attributable to an increase in intracellular Na(+), suggesting that the mitochondrial Na(+)/Ca(2+) exchanger is activated. Effects of alpha-LTX were not blocked when Ca(2+) increases were reduced greatly in saline lacking both Na(+) and Ca(2+) and by application of intracellular Ca(2+) chelators. Therefore, although increases in intracellular Ca(2+) may facilitate the effects of alpha-LTX on transmitter release, these increases do not appear to be necessary. The results show that investigations of Ca(2+) independent alpha-LTX mechanisms or uses of alpha-LTX to probe exocytosis mechanisms would be complicated by the release of intracellular Ca(2+), which itself can trigger exocytosis. PMID- 11102475 TI - Increased production of tumor necrosis factor-alpha by glial cells exposed to simulated ischemia or elevated hydrostatic pressure induces apoptosis in cocultured retinal ganglion cells. AB - Although glial cells in the optic nerve head undergo a reactivation process in glaucoma, the role of glial cells during glaucomatous neurodegeneration of retinal ganglion cells is unknown. Using a coculture system in which retinal ganglion cells and glial cells are grown on different layers but share the same culture medium, we studied the influences of glial cells on survival of retinal ganglion cells after exposure to different stress conditions typified by simulated ischemia and elevated hydrostatic pressure. After the exposure to these stressors, we observed that glial cells secreted tumor necrosis factor-alpha (TNF alpha) as well as other noxious agents such as nitric oxide into the coculture media and facilitated the apoptotic death of retinal ganglion cells as assessed by morphology, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling, and caspase activity. The glial origin of these noxious effects was confirmed by passive transfer experiments. Furthermore, retinal ganglion cell apoptosis was attenuated approximately 66% by a neutralizing antibody against TNF alpha and 50% by a selective inhibitor of inducible nitric oxide synthase (1400W). Because elevated intraocular pressure and ischemia are two prominent stress factors identified in the eyes of patients with glaucoma, these findings reveal a novel glia-initiated pathogenic mechanism for retinal ganglion cell death in glaucoma. In addition, these findings suggest that the inhibition of TNF alpha that is released by reactivated glial cells may provide a novel therapeutic target for neuroprotection in the treatment of glaucomatous optic neuropathy. PMID- 11102476 TI - Involvement of the extracellular signal-regulated kinase cascade for cocaine rewarding properties. AB - A central feature of drugs of abuse is to induce gene expression in discrete brain structures that are critically involved in behavioral responses related to addictive processes. Although extracellular signal-regulated kinase (ERK) has been implicated in several neurobiological processes, including neuronal plasticity, its role in drug addiction remains poorly understood. This study was designed to analyze the activation of ERK by cocaine, its involvement in cocaine induced early and long-term behavioral effects, as well as in gene expression. We show, by immunocytochemistry, that acute cocaine administration activates ERK throughout the striatum, rapidly but transiently. This activation was blocked when SCH 23390 [a specific dopamine (DA)-D1 antagonist] but not raclopride (a DA D2 antagonist) was injected before cocaine. Glutamate receptors of NMDA subtypes also participated in ERK activation, as shown after injection of the NMDA receptor antagonist MK 801. The systemic injection of SL327, a selective inhibitor of the ERK kinase MEK, before cocaine, abolished the cocaine-induced ERK activation and decreased cocaine-induced hyperlocomotion, indicating a role of this pathway in events underlying early behavioral responses. Moreover, the rewarding effects of cocaine were abolished by SL327 in the place-conditioning paradigm. Because SL327 antagonized cocaine-induced c-fos expression and Elk-1 hyperphosphorylation, we suggest that the ERK intracellular signaling cascade is also involved in the prime burst of gene expression underlying long-term behavioral changes induced by cocaine. Altogether, these results reveal a new mechanism to explain behavioral responses of cocaine related to its addictive properties. PMID- 11102477 TI - Dendritic and axonal targeting of type 5 metabotropic glutamate receptor is regulated by homer1 proteins and neuronal excitation. AB - The physiological actions of neurotransmitter receptors are intimately linked to their proper neuronal compartment localization. Here we studied the effect of the metabotropic glutamate receptor (mGluR)-interacting proteins, Homer1a, b, and c, in the targeting of mGluR5 in neurons. We found that mGluR5 was exclusively localized in cell bodies when transfected alone in cultured cerebellar granule cells. In contrast, mGluR5 was found also in dendrites when coexpressed with Homer1b or Homer1c, and in both dendrites and axons when cotransfected with Homer1a. In dendrites, cotransfected mGluR5 and Homer1b/c formed clusters that colocalized with the synaptic marker synaptophysin. Interestingly when transfected alone, the Homer proteins were also translocated to neurites but did not form such clusters. Depolarization of the neurons with a mixture of ionotropic glutamate receptor agonists, NMDA and kainate, or potassium channel blockers, tetraethylammonium and 4-aminopyridine, induced transient expression of endogenous Homer1a and persistent neuritic localization of transfected mGluR5 even long after degradation of Homer1a. These results suggest that Homer1a/b/c proteins are involved in the targeting of mGluR5 to dendritic synaptic sites and/or axons and that this effect can be regulated by neuronal activity. Because the activity-dependent effect of endogenous Homer1a was also long-lasting, the axonal targeting of mGluR5 by this protein is likely to play an important role in synaptic plasticity. PMID- 11102478 TI - Presenilin-1 P264L knock-in mutation: differential effects on abeta production, amyloid deposition, and neuronal vulnerability. AB - The pathogenic mechanism linking presenilin-1 (PS-1) gene mutations to familial Alzheimer's disease (FAD) is uncertain, but has been proposed to include increased neuronal sensitivity to degeneration and enhanced amyloidogenic processing of the beta-amyloid precursor protein (APP). We investigated this issue by using gene targeting with the Cre-lox system to introduce an FAD-linked P264L mutation into the endogenous mouse PS-1 gene, an approach that maintains normal regulatory controls over expression. Primary cortical neurons derived from PS-1 homozygous mutant knock-in mice exhibit basal neurodegeneration similar to their PS-1 wild-type counterparts. Staurosporine and Abeta1-42 induce apoptosis, and neither the dose dependence nor maximal extent of cell death is altered by the PS-1 knock-in mutation. Similarly, glutamate-induced neuronal necrosis is unaffected by the PS-1P264L mutation. The lack of effect of the PS-1P264L mutation is confirmed by measures of basal- and toxin-induced caspase and calpain activation, biochemical indices of apoptotic and necrotic signaling, respectively. To analyze the influence of the PS-1P264L knock-in mutation on APP processing and the development of AD-type neuropathology, we created mouse lines carrying mutations in both PS-1 and APP. In contrast to the lack of effect on neuronal vulnerability, cortical neurons cultured from PS-1P264L homozygous mutant mice secrete Abeta42 at an increased rate, whereas secretion of Abeta40 is reduced. Moreover, the PS-1 knock-in mutation selectively increases Abeta42 levels in the mouse brain and accelerates the onset of amyloid deposition and its attendant reactive gliosis, even as a single mutant allele. We conclude that expression of an FAD-linked mutant PS-1 at normal levels does not generally increase cortical neuronal sensitivity to degeneration. Instead, enhanced amyloidogenic processing of APP likely is critical to the pathogenesis of PS-1 linked FAD. PMID- 11102479 TI - Adult spinal cord stem cells generate neurons after transplantation in the adult dentate gyrus. AB - The adult rat spinal cord contains cells that can proliferate and differentiate into astrocytes and oligodendroglia in situ. Using clonal and subclonal analyses we demonstrate that, in contrast to progenitors isolated from the adult mouse spinal cord with a combination of growth factors, progenitors isolated from the adult rat spinal cord using basic fibroblast growth factor alone display stem cell properties as defined by their multipotentiality and self-renewal. Clonal cultures derived from single founder cells generate neurons, astrocytes, and oligodendrocytes, confirming the multipotent nature of the parent cell. Subcloning analysis showed that after serial passaging, recloning, and expansion, these cells retained multipotentiality, indicating that they are self-renewing. Transplantation of an in vitro-expanded clonal population of cells into the adult rat spinal cord resulted in their differentiation into glial cells only. However, after heterotopic transplantation into the hippocampus, transplanted cells that integrated in the granular cell layer differentiated into cells characteristic of this region, whereas engraftment into other hippocampal regions resulted in the differentiation of cells with astroglial and oligodendroglial phenotypes. The data indicate that clonally expanded, multipotent adult progenitor cells from a non-neurogenic region are not lineage-restricted to their developmental origin but can generate region-specific neurons in vivo when exposed to the appropriate environmental cues. PMID- 11102480 TI - Localization and enhanced current density of the Kv4.2 potassium channel by interaction with the actin-binding protein filamin. AB - Kv4.2 potassium channels play a critical role in postsynaptic excitability. Immunocytochemical studies reveal a somatodendritic Kv4.2 expression pattern, with the channels concentrated mainly at dendritic spines. The molecular mechanism that underlies the localization of Kv4.2 to this subcellular region is unknown. We used the yeast two-hybrid system to identify the Kv4.2-associated proteins that are involved in channel localization. Here we demonstrate a direct interaction between Kv4.2 and the actin-binding protein, filamin. We show that Kv4.2 and filamin can be coimmunoprecipitated both in vitro and in brain and that Kv4.2 and filamin share an overlapping expression pattern in the cerebellum and cultured hippocampal neurons. To examine the functional consequences of this interaction, we expressed Kv4.2 in filamin(+) and filamin(-) cells and performed immunocytochemical and electrophysiological analyses. Our results indicate that Kv4.2 colocalizes with filamin at filopodial roots in filamin(+) cells but shows a nonspecific expression pattern in filamin(-) cells, with no localization to filopodial roots. Furthermore, the magnitude of whole-cell Kv4.2 current density is approximately 2.7-fold larger in filamin(+) cells as compared with these currents in filamin(-) cells. We propose that filamin may function as a scaffold protein in the postsynaptic density, mediating a direct link between Kv4.2 and the actin cytoskeleton, and that this interaction is essential for the generation of appropriate Kv4.2 current densities. PMID- 11102481 TI - Insulysin hydrolyzes amyloid beta peptides to products that are neither neurotoxic nor deposit on amyloid plaques. AB - Insulysin (EC. 3.4.22.11) has been implicated in the clearance of beta amyloid peptides through hydrolytic cleavage. To further study the action of insulysin on Abeta peptides recombinant rat insulysin was used. Cleavage of both Abeta(1-40) and Abeta(1-42) by the recombinant enzyme was shown to initially occur at the His(13)-His(14), His(14)-Gln(15), and Phe(19)-Phe(20) bonds. This was followed by a slower cleavage at the Lys(28)-Gly(29), Val(18)-Phe(19), and Phe(20)-Ala(21) positions. None of the products appeared to be further metabolized by insulysin. Using a rat cortical cell system, the action of insulysin on Abeta(1-40) and Abeta(1-42) was shown to eliminate the neurotoxic effects of these peptides. Insulysin was further shown to prevent the deposition of Abeta(1-40) onto a synthetic amyloid. Taken together these results suggest that the use of insulysin to hydrolyze Abeta peptides represents an alternative gene therapeutic approach to the treatment of Alzheimer's disease. PMID- 11102482 TI - Exacerbation of noise-induced hearing loss in mice lacking the glutamate transporter GLAST. AB - Acoustic overstimulation is one of the major causes of hearing loss. Glutamate is the most likely candidate neurotransmitter for afferent synapses in the peripheral auditory system, so it was proposed that glutamate excitotoxicity may be involved in noise trauma. However, there has been no direct evidence that noise trauma is caused by excessive release of glutamate from the inner hair cells (IHCs) during sound exposure because studies have been hampered by powerful glutamate uptake systems in the cochlea. GLAST is a glutamate transporter highly expressed in the cochlea. Here we show that after acoustic overstimulation, GLAST deficient mice show increased accumulation of glutamate in perilymphs, resulting in exacerbation of hearing loss. These results suggest that GLAST plays an important role in keeping the concentration of glutamate in the perilymph at a nontoxic level during acoustic overstimulation. These findings also provide further support for the hypothesis that IHCs use glutamate as a neurotransmitter. PMID- 11102483 TI - Glial-derived neurotrophic factor upregulates expression of functional SNS and NaN sodium channels and their currents in axotomized dorsal root ganglion neurons. AB - Dorsal root ganglion (DRG) neurons produce multiple sodium currents, including several different TTX-sensitive (TTX-S) currents and TTX-resistant (TTX-R) currents, which are produced by distinct sodium channels. We previously demonstrated that, after sciatic nerve transection, the levels of SNS and NaN sodium channel alpha-subunit transcripts and protein in small (18-30 micrometer diameter) DRG neurons are reduced, as are the amplitudes and densities of the slowly inactivating and persistent TTX-R currents produced by these two channels. In this study, we asked whether glial-derived neurotrophic factor (GDNF), which has been shown to prevent some axotomy-induced changes such as the loss of somatostatin expression in DRG neurons, can ameliorate the axotomy-induced downregulation of SNS and NaN TTX-R sodium channels. We show here that exposure to GDNF can significantly increase both slowly inactivating and persistent TTX-R sodium currents, which are paralleled by increases in SNS and NaN mRNA and protein levels, in axotomized DRG neurons in vitro. We also show that intrathecally administered GDNF increases the amplitudes of the slowly inactivating and persistent TTX-R currents, and SNS and NaN protein levels, in peripherally axotomized DRG neurons in vivo. Finally, we demonstrate that GDNF upregulates the persistent TTX-R current in SNS-null mice, thus demonstrating that the upregulated persistent sodium current is not produced by SNS. Because TTX-R sodium channels have been shown to be important in nociception, the effects of GDNF on axotomized DRG neurons may have important implications for the regulation of nociceptive signaling by these cells. PMID- 11102484 TI - The Agrin/MuSK signaling pathway is spatially segregated from the neuregulin/ErbB receptor signaling pathway at the neuromuscular junction. AB - The neuregulin/erbB receptor and agrin/MuSK pathways are critical for communication between the nerve, muscle, and Schwann cell that establishes the precise topological arrangement at the vertebrate neuromuscular junction (NMJ). ErbB2, erbB3, and erbB4 as well as neuregulin, agrin, and MuSK are known to be concentrated at the NMJ. Here we have examined NMJs from gastrocnemius muscle of adult rat using immunofluorescence confocal microscopy to characterize in detail the distribution of these proteins relative to the distribution of acetylcholine receptors (AChRs). We have determined that erbB2 and erbB4 are enriched in the depths of the secondary junctional folds on the postsynaptic muscle membrane. In contrast, erbB3 at the NMJ was concentrated at presynaptic terminal Schwann cells. This distribution strongly argues that erbB2/erbB4 heterodimers are the functional postsynaptic neuregulin receptors of the NMJ. Neuregulin was localized to the axon terminal, secondary folds, and terminal Schwann cells, where it was in a position to signal through erbB receptors. MuSK was concentrated in the postsynaptic primary gutter region where it was codistributed with AChRs. Agrin was present at the axon terminal and in the basal lamina associated with the primary gutter region, but not in the secondary junctional folds. The differential distributions of the neuregulin and agrin signaling pathways argue against neuregulin and erbB receptors being localized to the NMJ via direct interactions with either agrin or MuSK. PMID- 11102485 TI - Disparity in neurotransmitter release probability among competing inputs during neuromuscular synapse elimination. AB - Competition among the several motor axons transiently innervating neonatal muscle fibers results in an increasing disparity in the quantal content and synaptic territory of each competitor, culminating in the permanent loss of all but one axon from neuromuscular junctions. We asked whether differences in the probability of neurotransmitter release also contribute to the increasing disparity in quantal content among competing inputs, and when in the process of competition changes in release probability become apparent. To address these questions, intracellular recordings were made from dually innervated neonatal mouse soleus muscle fibers, and quantal content and paired-pulse facilitation were evaluated for each input. At short interpulse intervals, paired-pulse facilitation was significantly higher for the weaker input with the smaller quantal content than the stronger input with the larger quantal content. Because neurotransmitter release probability across all release sites is inversely related to the extent of facilitation observed after paired-pulse stimulation, this result suggests that release probability is lower for weak compared with strong inputs innervating the same junction. A disparity in the probability of neurotransmitter release thus contributes to the disparity in quantal content that occurs during synaptic competition. Together, this work suggests that an input incapable of sustaining a high release probability may be at a competitive disadvantage for synaptic maintenance. PMID- 11102486 TI - Differential postnatal development of catecholamine and serotonin inputs to identified neurons in prefrontal cortex of rhesus monkey. AB - The monoaminergic innervation of cerebral cortex has long been implicated in its development. Methods now exist to examine catecholamine and serotonin inputs to identified neurons in the cerebral cortex. We have quantified such inputs on pyramidal and nonpyramidal cells in prefrontal cortex of rhesus monkeys ranging in age from 2 weeks to 10 years. Individual layer III neurons were filled with Lucifer yellow and double-immunostained with axons containing either tyrosine hydroxylase (TH) or 5-hydroxytryptamine (5-HT). The filled cells were reconstructed, and putative appositions between the axons and dendritic spines and shafts were quantified at high magnification using light microscopy. The density of catecholamine appositions on pyramidal neurons matures slowly, reaching only half the adult level by 6 months of age and thereafter rising gradually to adult levels by 2 years of age. By contrast, the density of serotonin appositions on pyramidal cells reaches the adult level before the second week after birth. The average adult pyramidal neuron in layer III of area 9m receives three times stronger input from catecholaminergic than from serotoninergic axons. The overall density of both inputs to interneurons does not appear to change during postnatal development. Selective changes in the TH innervation of pyramidal cells against a backdrop of constant TH innervation of interneurons suggest that the balance between excitation and inhibition may change developmentally in the prefrontal cortex. By contrast, 5-HT innervation of both types of neurons remains relatively constant over the age range studied. PMID- 11102487 TI - Fetal hippocampal grafts containing CA3 cells restore host hippocampal glutamate decarboxylase-positive interneuron numbers in a rat model of temporal lobe epilepsy. AB - Degeneration of CA3-pyramidal neurons in hippocampus after intracerebroventricular kainic acid (KA) administration, a model of temporal lobe epilepsy, results in hyperexcitability within both dentate gyrus and the CA1 subfield. It also leads to persistent reductions in hippocampal glutamate decarboxylase (GAD) interneuron numbers without diminution in Nissl-stained interneuron numbers, indicating loss of GAD expression in a majority of interneurons. We hypothesize that enduring loss of GAD expression in hippocampal interneurons after intracerebroventricular KA is attributable to degeneration of their CA3 afferent input; therefore, fetal CA3 grafts can restore GAD interneuron numbers through graft axon reinnervation of the host. We analyzed GAD interneuron density in the adult rat hippocampus at 6 months after KA administration after grafting of fetal mixed hippocampal, CA3 or CA1 cells into the CA3 region at 45 d after lesion, in comparison with "lesion-only" and intact hippocampus. In dentate and CA1 regions of the lesioned hippocampus receiving grafts of either mixed hippocampal or CA3 cells, GAD interneuron density was both significantly greater than lesion-only hippocampus and comparable with the intact hippocampus. In the CA3 region, GAD interneuron density was significantly greater than lesion-only hippocampus but less than the intact hippocampus. Collectively, the overall GAD interneuron density in the lesioned hippocampus receiving either mixed hippocampal or CA3 grafts was restored to that in the intact hippocampus. In contrast, GADinterneuron density in the lesioned hippocampus receiving CA1 grafts remained comparable with lesion-only hippocampus. Thus, grafts containing CA3 cells restore CA3 lesion-induced depletions in hippocampal GAD interneurons, likely by reinnervation of GAD-deficient interneurons. This specific graft mediated effect is beneficial because reactivation of interneurons could ameliorate both loss of functional inhibition and hyperexcitability in CA3 lesioned hippocampus. PMID- 11102488 TI - Long-term nicotine adaptation in Caenorhabditis elegans involves PKC-dependent changes in nicotinic receptor abundance. AB - Chronic exposure to nicotine leads to long-term changes in both the abundance and activity of nicotinic acetylcholine receptors, processes thought to contribute to nicotine addiction. We have found that in Caenorhabditis elegans, prolonged nicotine treatment results in a long-lasting decrease in the abundance of nicotinic receptors that control egg-laying. In naive animals, acute exposure to cholinergic agonists led to the efficient stimulation of egg-laying, a response mediated by a nicotinic receptor functionally expressed in the vulval muscle cells. Overnight exposure to nicotine led to a specific and long-lasting change in egg-laying behavior, which rendered the nicotine-adapted animals insensitive to simulation of egg-laying by the nicotinic agonist and was accompanied by a promoter-independent reduction in receptor protein levels. Mutants defective in the gene tpa-1, which encodes a homolog of protein kinase C (PKC), failed to undergo adaptation to nicotine; after chronic nicotine exposure they remained sensitive to cholinergic agonists and retained high levels of receptor protein in the vulval muscles. These results suggest that PKC-dependent signaling pathways may promote nicotine adaptation via regulation of nicotinic receptor synthesis or degradation. PMID- 11102489 TI - Stable Hebbian learning from spike timing-dependent plasticity. AB - We explore a synaptic plasticity model that incorporates recent findings that potentiation and depression can be induced by precisely timed pairs of synaptic events and postsynaptic spikes. In addition we include the observation that strong synapses undergo relatively less potentiation than weak synapses, whereas depression is independent of synaptic strength. After random stimulation, the synaptic weights reach an equilibrium distribution which is stable, unimodal, and has positive skew. This weight distribution compares favorably to the distributions of quantal amplitudes and of receptor number observed experimentally in central neurons and contrasts to the distribution found in plasticity models without size-dependent potentiation. Also in contrast to those models, which show strong competition between the synapses, stable plasticity is achieved with little competition. Instead, competition can be introduced by including a separate mechanism that scales synaptic strengths multiplicatively as a function of postsynaptic activity. In this model, synaptic weights change in proportion to how correlated they are with other inputs onto the same postsynaptic neuron. These results indicate that stable correlation-based plasticity can be achieved without introducing competition, suggesting that plasticity and competition need not coexist in all circuits or at all developmental stages. PMID- 11102490 TI - GABAergic inhibition suppresses paroxysmal network activity in the neonatal rodent hippocampus and neocortex. AB - In the adult cerebral cortex, the neurotransmitter GABA is strongly inhibitory, as it profoundly decreases neuronal excitability and suppresses the network propensity for synchronous activity. When fast, GABA(A) receptor (GABA(A)R) mediated neurotransmission is blocked in the mature cortex, neuronal firing is synchronized via recurrent excitatory (glutamatergic) synaptic connections, generating population discharges manifested extracellularly as spontaneous paroxysmal field potentials (sPFPs). This epileptogenic effect of GABA(A)R antagonists has rarely been observed in the neonatal cortex, and indeed, GABA in the neonate has been proposed to have an excitatory, rather than inhibitory, action. In contrast, we show here that when fast GABAergic neurotransmission was blocked in slices of neonatal mouse and rat hippocampus and neocortex, sPFPs occurred in nearly half the slices from postnatal day 4 (P4) to P7 neocortex and in most slices from P2 to P7 hippocampus. In Mg(2+)-free solution, GABA(A)R antagonists elicited sPFPs in nearly all slices of P2 and older neocortex and P0 and older hippocampus. Mg(2+)-free solution alone induced spontaneous events in the majority of P2 and older slices from both regions; addition of GABA(A)R antagonists caused a dramatic increase in the mean amplitude, but not frequency, of these events in the hippocampus and in their mean frequency, but not amplitude, in the neocortex. In the hippocampus, GABA(A)R agonists suppressed amplitudes, but not frequency, of sPFPs, whereas glutamate antagonists suppressed frequency but not amplitudes. We conclude that neonatal rodent cerebral cortex possesses glutamatergic circuits capable of generating synchronous network activity and that, as in the adult, tonic GABA(A)R-mediated inhibition prevents this activity from becoming paroxysmal. PMID- 11102492 TI - Human cortical muscle coherence is directly related to specific motor parameters. AB - Cortical oscillations have been the target of many recent investigations, because it has been proposed that they could function to solve the "binding" problem. In the motor cortex, oscillatory activity has been reported at a variety of frequencies between approximately 4 and approximately 60 Hz. Previous research has shown that 15-30 Hz oscillatory activity in the primary motor cortex is coherent or phase locked to activity in contralateral hand and forearm muscles during isometric contractions. However, the function of this oscillatory activity remains unclear. Is it simply an epiphenomenon or is it related to specific motor parameters? In this study, we investigated task-dependent modulation in coherence between motor cortex and hand muscles during precision grip tasks. Twelve right handed subjects used index finger and thumb to grip two levers that were under robotic control. Each lever was fitted with a sensitive force gauge. Subjects received visual feedback of lever force levels and were instructed to keep them within target boxes throughout each trial. Surface EMGs were recorded from four hand and forearm muscles, and magnetoencephalography (MEG) was recorded using a 306 channel neuromagnetometer. All subjects showed significant levels of coherence (0.086-0.599) between MEG and muscle in the 15-30 Hz range. Coherence was significantly smaller when the task was performed under an isometric condition (levers fixed) compared with a compliant condition in which subjects moved the levers against a spring-like load. Furthermore, there was a positive, significant relationship between the level of coherence and the degree of lever compliance. These results argue in favor of coherence between cortex and muscle being related to specific parameters of hand motor function. PMID- 11102491 TI - NMDA and glutamate evoke excitotoxicity at distinct cellular locations in rat cortical neurons in vitro. AB - The development of cortical neurons in vivo and in vitro is accompanied by alterations in NMDA receptor subunit expression and concomitant modifications in the pharmacological profile of NMDA-activated ionic currents. For example, we observed that with decreasing NR2B/NR2A subunit expression ratio, the block of NMDA receptor-mediated whole-cell responses by the NR2B-selective antagonist haloperidol was also decreased. In mature cultures (>22 d in vitro), however, NMDA responses obtained from excised nucleated macropatches, which comprised a large portion of the soma, remained strongly antagonized by haloperidol. These results suggest that in more mature neurons NR1/NR2B receptors appear to be preferentially expressed in the cell body. As predicted from the whole-cell recording pharmacological profile, NMDA-induced toxicity was largely unaffected by haloperidol in mature cultures. However, haloperidol effectively blocked glutamate toxicity in the same cultures, suggesting that the neurotoxic actions of this amino acid were mostly due to the activation of somatic NMDA receptors. In experiments in which the potency of glutamate toxicity was increased by the transport inhibitor l-trans-pyrrolidine-2,4-dicarboxylic acid, the neuroprotective effects of haloperidol were significantly diminished. This was likely because of the fact that glutamate, now toxic at much lower concentrations, was able to reach and activate dendritic receptors under these conditions. These results strongly argue that exogenous glutamate and NMDA normally induce excitotoxicity at distinct cellular locations in mature mixed neuronal cultures and that NR1/NR2B receptors remain an important component in the expression of glutamate, but not NMDA-induced excitotoxicity. PMID- 11102493 TI - DOI-Induced activation of the cortex: dependence on 5-HT2A heteroceptors on thalamocortical glutamatergic neurons. AB - Administration of the hallucinogenic 5-HT(2A/2C) agonist 1-[2, 5-dimethoxy-4 iodophenyl]-2-aminopropane (DOI) induces expression of Fos protein in the cerebral cortex. To understand the mechanisms subserving this action of DOI, we examined the consequences of pharmacological and surgical manipulations on DOI elicited Fos expression in the somatosensory cortex of the rat. DOI dose dependently increased cortical Fos expression. Pretreatment with the selective 5 HT(2A) antagonist MDL 100,907 completely blocked DOI-elicited Fos expression, but pretreatment with the 5-HT(2C) antagonist SB 206,553 did not modify DOI-elicited Fos expression. These data suggest that DOI acts through 5-HT(2A) receptors to increase cortical Fos expression. However, we found that DOI did not induce Fos in cortical 5-HT(2A) immunoreactive neurons but did increase expression in a band of neurons spanning superficial layer V to deep III, within the apical dendritic fields of layer V 5-HT(2A)-immunoreactive cells. This band of Fos immunoreactive neurons was in register with anterogradely labeled axons from the ventrobasal thalamus, which have previously been shown to be glutamatergic and express the 5 HT(2A) transcript. The effects of DOI were markedly reduced in animals pretreated with the AMPA/KA antagonist GYKI 52466, and lesions of the ventrobasal thalamus attenuated DOI-elicited Fos expression in the cortex. These data suggest that DOI activates 5-HT(2A) receptors on thalamocortical neurons and thereby increases glutamate release, which in turn drives Fos expression in cortical neurons through an AMPA receptor-dependent mechanism. These data cast new light on the mechanisms of action of hallucinogens. PMID- 11102494 TI - Impaired recognition memory in rats after damage to the hippocampus. AB - Rats with radio-frequency or ibotenic acid lesions of the hippocampus and rats with radio-frequency lesions of the fornix were tested on the visual paired comparison task (VPC), a test of recognition memory. Memory was assessed at five different delay intervals ranging from 10 sec to 24 hr. All operated groups performed normally at the shorter delays (10 sec and 1 min). Across longer delays, the two groups with hippocampal damage were impaired. Rats with fornix lesions performed well on the VPC task but were impaired on a spatial task (spontaneous alternation). The results show that the hippocampus is essential for normal recognition memory. Moreover, fornix lesions need not mimic the effects of direct damage to hippocampal tissue. The findings are discussed in the context of the contribution of the hippocampus to recognition memory. PMID- 11102495 TI - Brain-stimulation reward thresholds raised by an antisense oligonucleotide for the M5 muscarinic receptor infused near dopamine cells. AB - Oligonucleotides targeting M5 muscarinic receptor mRNA were infused for 6 d into the ventral tegmental area of freely behaving rats trained to bar-press for lateral hypothalamic stimulation. The bar-pressing rate was determined at a range of frequencies each day to evaluate the effects of infusions on reward. M5 antisense oligonucleotide (oligo) infusions increased the frequency required for bar pressing by 48% over baseline levels, with the largest increases occurring after 4-6 d of infusion. Two control oligos had only slight effects (means of 5 and 11% for missense and sense oligos, respectively). After the infusion, the required frequency shifted back to baseline levels gradually over 1-5 d. Antisense oligo infusions decreased M5 receptors on the ipsilateral, but not the contralateral, side of the ventral tegmentum, as compared with a missense oligo. Therefore, M5 muscarinic receptors associated with mesolimbic dopamine neurons seem to be important in brain-stimulation reward. PMID- 11102496 TI - Purinergic and adrenergic agonists synergize in stimulating vasopressin and oxytocin release. AB - The A1 catecholamine neurons of the caudal ventrolateral medulla transmit hemodynamic information to the vasopressin (VP) neurons in the hypothalamus. These neurons corelease ATP with norepinephrine. Perifused explants of the hypothalamoneurohypophyseal system were used to investigate the role of these substances on VP release. ATP (100 micrometer) increased VP release 1.5-fold (p = 0.027). The response was rapid but unsustained. It was blocked by the P(2) receptor antagonist pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS). The alpha(1)-adrenergic agonist phenylephrine (PE; 100 micrometer) also increased VP release by 1.5-fold (p = 0.014). Again, the response was rapid and unsustained. However, simultaneous perifusion of explants with ATP (100 micrometer) and PE (100 micrometer) resulted in a threefold to fourfold increase in VP release, which was sustained for as long as 4 hr. There was a similar synergistic effect of ATP and PE on oxytocin release. Interestingly, the synergistic response was delayed approximately 40 min relative to the response to either agent alone. Several experiments were performed to elucidate the cellular mechanisms of this synergism. The effect was blocked by PPADS, a protein kinase C inhibitor (bisindolylmaleimide I HCl), and actinomycin, an inhibitor of gene transcription. These data suggest that P(2X) receptor activation, PKC-mediated phosphorylation, and gene transcription are required for the synergistic response. The marked synergism of these coreleased agents is probably important to achieve sustained increases in plasma VP in response to prolonged hypotension. These observations may also have broad applications to CNS function, because ATP may be coreleased at noradrenergic synapses throughout the CNS. PMID- 11102497 TI - Enhanced vulnerability to cocaine self-administration is associated with elevated impulse activity of midbrain dopamine neurons. AB - Individual differences in responding to a novel environment predict behavioral and neurochemical responses to psychostimulant drugs. Rats with a high locomotor response to a novel environment (HRs) exhibit enhanced self-administration (SA) behavior, sensitization, and basal or drug-induced dopamine release in the nucleus accumbens compared with rats with a low response to the novel context (LRs). In this study, we determined whether such differences in vulnerability to drug addiction might be related to differences in dopamine (DA) neuron activity. Rats were divided into HRs and LRs according to their response to a novel environment and then tested for acquisition of cocaine SA. HRs rapidly acquired cocaine SA (175 microg/kg per infusion), whereas LRs did not. Differences in cocaine SA were not caused by differences in exploratory behavior or sampling because these behaviors did not differ in HRs and LRs self-administering a saline solution. In a separate experiment, we used extracellular single-unit recordings and found that HRs exhibit higher basal firing rates and bursting activity of DA neurons in the ventral tegmental area and, to a lesser extent, in the substantia nigra pars compacta. The greater activity of midbrain DA cells in HRs was accompanied by reduced sensitivity to the inhibitory effects of a DA D2-class receptor agonist, indicating possible subsensitivity of impulse-regulating DA autoreceptors. These results demonstrate that differences in the basal activity of DA neurons may be critically involved in determining individual vulnerability to drugs of abuse. PMID- 11102498 TI - Reliability of a fly motion-sensitive neuron depends on stimulus parameters. AB - The variability of responses of sensory neurons constrains how reliably animals can respond to stimuli in the outside world. We show for a motion-sensitive visual interneuron of the fly that the variability of spike trains depends on the properties of the motion stimulus, although differently for different stimulus parameters. (1) The spike count variances of responses to constant and to dynamic stimuli lie in the same range. (2) With increasing stimulus size, the variance may slightly decrease. (3) Increasing pattern contrast reduces the variance considerably. For all stimulus conditions, the spike count variance is much smaller than the mean spike count and does not depend much on the mean activity apart from very low activities. Using a model of spike generation, we analyzed how the spike count variance depends on the membrane potential noise and the deterministic membrane potential fluctuations at the spike initiation zone of the neuron. In a physiologically plausible range, the variance is affected only weakly by changes in the dynamics or the amplitude of the deterministic membrane potential fluctuations. In contrast, the amplitude and dynamics of the membrane potential noise strongly influence the spike count variance. The membrane potential noise underlying the variability of the spike responses in the motion sensitive neuron is concluded to be affected considerably by the contrast of the stimulus but by neither its dynamics nor its size. PMID- 11102499 TI - Thalamic reticular nucleus activation reflects attentional gating during classical conditioning. AB - All senses, except olfaction, are routed through the thalamus to cerebral cortex. Thus, the thalamus is often referred to as the sensory gateway to cortex. Located between thalamus and cortex is a thin lamina of neurons called the thalamic reticular nucleus, which may function as an attentional gate. The phenomenon of blocking in classical conditioning provides an opportunity to test whether an attended stimulus activates the thalamic reticular nucleus more than an unattended stimulus: when a second stimulus is presented together with a previously conditioned stimulus, conditioned responding to the second stimulus is inhibited. Different groups of rats were given conditioning sessions with a single stimulus, a light or a tone, and then given conditioning sessions with compound (light and tone) stimuli. Blocking was confirmed using probe trials of single stimulus presentations. After a final test session of compound stimulus presentations, the brains were processed for the presence of Fos protein. Here we show that Fos-positive neurons were more numerous in the sector of the thalamic reticular nucleus associated with the attended conditioned stimulus than in the sector associated with the unattended stimulus. Thus, we provide evidence for an involvement of the thalamic reticular nucleus in selective attention. PMID- 11102500 TI - Prenatal cocaine exposure increases sensitivity to the attentional effects of the dopamine D1 agonist SKF81297. AB - Sensitivity to the attentional effects of SKF81297, a selective full agonist at dopamine D(1) receptors, was assessed in adult rats exposed to cocaine prenatally (via intravenous injections) and controls. The task assessed the ability of the subjects to monitor an unpredictable light cue of either 300 or 700 msec duration and to maintain performance when presented with olfactory distractors. SKF81297 decreased nose pokes before cue presentation and increased latencies and response biases (the tendency to respond to the same port used on the previous trial), suggesting an effect of SKF81297 on the dopamine (DA) systems responsible for response initiation and selection. The cocaine-exposed (COC) and control animals did not differ in sensitivity to the effects of SKF81297 on these measures. In contrast, the COC animals were significantly more sensitive than were controls to the impairing effect of SKF81297 on omission errors, a measure of sustained attention. This pattern of results provides evidence that prenatal cocaine exposure produces lasting changes in the DA system(s) subserving sustained attention but does not alter the DA system(s) underlying response selection and initiation. These findings also provide support for the role of D(1) receptor activation in attentional functioning. PMID- 11102501 TI - Putative cortical and thalamic inputs elicit convergent excitation in a population of GABAergic interneurons of the lateral amygdala. AB - Synaptic circuitry in the rat lateral amygdala (AL) was studied in brain slices using electrophysiological recordings. Electrical stimulation of external and internal capsules evoked an EPSC followed by a sequence of GABA(A) and GABA(B) receptor-mediated IPSC in principal neurons. Paired stimulation of either afferents resulted in a significant reduction ( approximately 45%) of the second GABA(A) receptor-mediated IPSC. A priming stimulation, consisting of a priming pulse to one pathway followed by a pulse to the other pathway, resulted in a strong depression of the second IPSC basically identical to that during paired stimulation. Paired- and primed-pulse depressions were largely relieved by 10 micrometer CGP 55845A, indicating regulation through presynaptic GABA(B) receptors. Furthermore, putative interneurons responded with EPSCs of constant latencies to minimal stimulation of both cortical and thalamic fibers, indicating convergent monosynaptic input. At higher stimulation strength, an approximately 15% reduction of EPSCs occurred in interneurons after paired and primed stimulation, which was not sensitive to CGP 55845A. These findings indicate that a rather homogeneous population of interneurons exists in the AL with respect to their afferent connectivity, in that they receive convergent input through putative thalamic and cortical fibers, both directly and indirectly (through principal neurons), and mediate inhibitory control of postsynaptic principal neurons. This symmetrically built GABAergic circuitry can be of functional significance, given the distinctive role of the two afferent input systems for the mediation of different components of fear responses and the importance of GABAergic mechanisms for limitation of excessive neuronal activity. PMID- 11102502 TI - Learning of visuomotor transformations for vectorial planning of reaching trajectories. AB - The planning of visually guided reaches is accomplished by independent specification of extent and direction. We investigated whether this separation of extent and direction planning for well practiced movements could be explained by differences in the adaptation to extent and directional errors during motor learning. We compared the time course and generalization of adaptation with two types of screen cursor transformation that altered the relationship between hand space and screen space. The first was a gain change that induced extent errors and required subjects to learn a new scaling factor. The second was a screen cursor rotation that induced directional errors and required subjects to learn new reference axes. Subjects learned a new scaling factor at the same rate when training with one or multiple target distances, whereas learning new reference axes took longer and was less complete when training with multiple compared with one target direction. After training to a single target, subjects were able to transfer learning of a new scaling factor to previously unvisited distances and directions. In contrast, generalization of rotation adaptation was incomplete; there was transfer across distances and arm configurations but not across directions. Learning a rotated reference frame only occurred after multiple target directions were sampled during training. These results suggest the separate processing of extent and directional errors by the brain and support the idea that reaching movements are planned as a hand-centered vector whose extent and direction are established via learning a scaling factor and reference axes. PMID- 11102503 TI - Reduced inhibition in an animal model of cortical dysplasia. AB - Cortical dysplasia has a strong association with epilepsy in humans, but the underlying mechanisms for this are poorly understood. In utero irradiation of rats produces diffuse cortical dysplasia and neuronal heterotopia in the neocortex and hippocampus. Using in vitro neocortical slices, whole-cell patch clamp recordings were obtained from pyramidal neurons in dysplastic cortex and control neocortex. Spontaneous IPSCs were reduced in amplitude (35%) and frequency (70%) in pyramidal cells from dysplastic cortex. Miniature IPSCs were reduced in frequency (66%) in dysplastic cortex. Two additional measures of cortical inhibition, monosynaptic evoked IPSCs and paired pulse depression of evoked EPSCs, were also impaired in dysplastic cortex. Spontaneous EPSCs were increased in amplitude (42%) and frequency (77%) in dysplastic cortex, but miniature EPSCs were not different between the two groups. These data demonstrate significant physiological impairment in inhibitory synaptic transmission in experimental cortical dysplasia. This supports previous immunohistochemical findings in this model and observations in humans of a reduction in the density of inhibitory interneurons in dysplastic cortex. PMID- 11102504 TI - Cannabinoids reveal the necessity of hippocampal neural encoding for short-term memory in rats. AB - The memory-disruptive effects of Delta(9)-tetrahydrocannabinol (Delta(9)-THC) and the synthetic cannabinoid WIN 55,212-2 (WIN-2) were assessed in rats exposed to varying doses of each drug (Delta(9)-THC, 0.5-2.0 mg/kg; WIN-2, 0.25-0.75 mg/kg) during performance of a delayed nonmatch to sample (DNMS) task. Cannabinoids affected performance in a dose x delay-dependent manner, with WIN-2 showing a potency more than four times that of Delta(9)-THC. These effects on DNMS performance were eliminated if the cannabinoid CB1 receptor antagonist SR141617A (Sanofi Research Inc.) was preadministered, but doses of the antagonist alone had no effect on performance. Simultaneous recording from ensembles of hippocampal neurons revealed that both WIN-2 and Delta(9)-THC produced dose-dependent reductions in the frequency (i.e., "strength") of ensemble firing during the sample phase of the task to the extent that performance was at risk for errors on >70% of trials as a function of delay. This decrease in ensemble firing in the Sample phase resulted from selective interference with the activity of differentiated hippocampal functional cell types, which conjunctively encoded different combinations of task events. A reduction in ensemble firing strength did not occur in the nonmatch phase of the task. The findings indicate that activation of CB1 receptors renders animals at risk for retention of item specific information in much the same manner as hippocampal removal. PMID- 11102505 TI - Projection neurons with shared cotransmitters elicit different motor patterns from the same neural circuit. AB - Specificity in the actions of different modulatory neurons is often attributed to their having distinct cotransmitter complements. We are assessing the validity of this hypothesis with the stomatogastric nervous system of the crab Cancer borealis. In this nervous system, the stomatogastric ganglion (STG) contains a multifunctional network that generates the gastric mill and pyloric rhythms. Two identified projection neurons [modulatory proctolin neuron (MPN) and modulatory commissural neuron 1 (MCN1)] that innervate the STG and modulate these rhythms contain GABA and the pentapeptide proctolin, but only MCN1 contains Cancer borealis tachykinin-related peptide (CabTRP Ia). Selective activation of each projection neuron elicits different rhythms from the STG. MPN elicits only a pyloric rhythm, whereas MCN1 elicits a distinct pyloric rhythm as well as a gastric mill rhythm. We tested the degree to which CabTRP Ia distinguishes the actions of MCN1 and MPN. To this end, we used the tachykinin receptor antagonist Spantide I to eliminate the actions of CabTRP Ia. With Spantide I present, MCN1 no longer elicited the gastric mill rhythm and the resulting pyloric rhythm was changed. Although this rhythm was more similar to the MPN-elicited pyloric rhythm, these rhythms remained different. Thus, CabTRP Ia partially confers the differences in rhythm generation resulting from MPN versus MCN1 activation. This result suggests that different projection neurons may use the same cotransmitters differently to elicit distinct pyloric rhythms. It also supports the hypothesis that different projection neurons use a combination of strategies, including using distinct cotransmitter complements, to elicit different outputs from the same neuronal network. PMID- 11102506 TI - The effect of lesions of the insular cortex on instrumental conditioning: evidence for a role in incentive memory. AB - In three experiments, we assessed the effect of lesions aimed at the gustatory region of the insular cortex on instrumental conditioning in rats. In experiment 1, the lesion had no effect on the acquisition of either lever pressing or chain pulling in food-deprived rats whether these actions earned food pellets or a maltodextrin solution. The lesion did, however, attenuate the impact of outcome devaluation, induced by sensory-specific satiety, on instrumental performance but only when assessed in an extinction test. This effect was not secondary to an impairment in instrumental learning; in experiment 2, no evidence was found to suggest that the lesioned rats differed from shams in their ability to encode the specific action-outcome contingencies to which they were exposed during training. In experiment 3, however, lesioned rats were found to be insensitive to the impact of an incentive learning treatment conducted when they were undeprived; although, again, this deficit was confined to a test conducted in extinction. These results are consistent with the view that, in instrumental conditioning, the gustatory region of the insular cortex is involved in encoding the taste of food outcomes in memory and, hence, in encoding the incentive value assigned to these outcomes on the basis of prevailing motivational conditions. PMID- 11102508 TI - A conserved interaction between Moe1 and Mal3 is important for proper spindle formation in Schizosaccharomyces pombe. AB - Moe1 is a conserved fission yeast protein that negatively affects microtubule stability/assembly. We conducted a two-hybrid screen to search for Moe1-binding proteins and isolated Mal3, a homologue of human EB1. We show that Moe1 and Mal3 expressed in bacteria form a complex and that Moe1 and Mal3 expressed in fission yeast cosediment with microtubules. Deletion of either moe1 or mal3 does not result in lethality; however, deletion of both moe1 and mal3 leads to cell death in the cold. The resulting cells appear to die of chromosome missegregation, which correlates with the presence of abnormal spindles. We investigated the cause for the formation of monopolar spindles and found that only one of the two spindle pole bodies (SPBs) contains gamma-tubulin, although both SPBs appear to be equal in size and properly inserted in the nuclear membrane. Moreover, the moe1 mal3 double null mutant in the cold contains abnormally short and abundant interphase microtubule bundles. These data suggest that Moe1 and Mal3 play a role in maintaining proper microtubule dynamics/integrity and distribution of gamma tubulin to the SPBs during mitosis. Finally, we show that human Moe1 and EB1 can each rescue the phenotype of the moe1 mal3 double null mutant and form a complex, suggesting that these proteins are part of a well-conserved mechanism for regulating spindle functioning. PMID- 11102507 TI - A snc1 endocytosis mutant: phenotypic analysis and suppression by overproduction of dihydrosphingosine phosphate lyase. AB - The v-SNARE proteins Snc1p and Snc2p are required for fusion of secretory vesicles with the plasma membrane in yeast. Mutation of a methionine-based sorting signal in the cytoplasmic domain of either Sncp inhibits Sncp endocytosis and prevents recycling of Sncp to the Golgi after exocytosis. snc1-M43A mutant yeast have reduced growth and secretion rates and accumulate post-Golgi secretory vesicles and fragmented vacuoles. However, cells continue to grow and secrete for several hours after de novo Snc2-M42A synthesis is repressed. DPL1, the structural gene for dihydrosphingosine phosphate lyase, was selected as a high copy number snc1-M43A suppressor. Because DPL1 also partially suppresses the growth and secretion phenotypes of a snc deletion, we propose that enhanced degradation of dihydrosphingosine-1-phosphate allows an alternative protein to replace Sncp as the secretory vesicle v-SNARE. PMID- 11102509 TI - Insulin recruits GLUT4 from specialized VAMP2-carrying vesicles as well as from the dynamic endosomal/trans-Golgi network in rat adipocytes. AB - Insulin treatment of fat cells results in the translocation of the insulin responsive glucose transporter type 4, GLUT4, from intracellular compartments to the plasma membrane. However, the precise nature of these intracellular GLUT4 carrying compartments is debated. To resolve the nature of these compartments, we have performed an extensive morphological analysis of GLUT4-containing compartments, using a novel immunocytochemical technique enabling high labeling efficiency and 3-D resolution of cytoplasmic rims isolated from rat epididymal adipocytes. In basal cells, GLUT4 was localized to three morphologically distinct intracellular structures: small vesicles, tubules, and vacuoles. In response to insulin the increase of GLUT4 at the cell surface was compensated by a decrease in small vesicles, whereas the amount in tubules and vacuoles was unchanged. Under basal conditions, many small GLUT4 positive vesicles also contained IRAP (88%) and the v-SNARE, VAMP2 (57%) but not markers of sorting endosomes (EEA1), late endosomes, or lysosomes (lgp120). A largely distinct population of GLUT4 vesicles (56%) contained the cation-dependent mannose 6-phosphate receptor (CD MPR), a marker protein that shuttles between endosomes and the trans-Golgi network (TGN). In response to insulin, GLUT4 was recruited both from VAMP2 and CD MPR positive vesicles. However, while the concentration of GLUT4 in the remaining VAMP2-positive vesicles was unchanged, the concentration of GLUT4 in CD-MPR positive vesicles decreased. Taken together, we provide morphological evidence indicating that, in response to insulin, GLUT4 is recruited to the plasma membrane by fusion of preexisting VAMP2-carrying vesicles as well as by sorting from the dynamic endosomal-TGN system. PMID- 11102510 TI - Distinct FTDP-17 missense mutations in tau produce tau aggregates and other pathological phenotypes in transfected CHO cells. AB - Multiple tau gene mutations are pathogenic for hereditary frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), with filamentous tau aggregates as the major lesions in the CNS of these patients. Recent studies have shown that bacterially expressed recombinant tau proteins with FTDP-17 missense mutations cause functional impairments, i.e., a reduced ability of mutant tau to bind to or promote the assembly of microtubules. To investigate the biological consequences of FTDP-17 tau mutants and assess their ability to form filamentous aggregates, we engineered Chinese hamster ovary cell lines to stably express tau harboring one or several different FTDP-17 mutations and showed that different tau mutants produced distinct pathological phenotypes. For example, delta K, but not several other single tau mutants (e.g., V337 M, P301L, R406W), developed insoluble amorphous and fibrillar aggregates, whereas a triple tau mutant (VPR) containing V337M, P301L, and R406W substitutions also formed similar aggregates. Furthermore, the aggregates increased in size over time in culture. Significantly, the formation of aggregated delta K and VPR tau protein correlated with reduced affinity of these mutants to bind microtubules. Reduced phosphorylation and altered proteolysis was also observed in R406W and delta K tau mutants. Thus, distinct pathological phenotypes, including the formation of insoluble filamentous tau aggregates, result from the expression of different FTDP-17 tau mutants in transfected Chinese hamster ovary cells and implies that these missense mutations cause diverse neurodegenerative FTDP-17 syndromes by multiple mechanisms. PMID- 11102511 TI - Human orthologs of yeast vacuolar protein sorting proteins Vps26, 29, and 35: assembly into multimeric complexes. AB - Sorting nexin (SNX) 1 and SNX2 are mammalian orthologs of Vps5p, a yeast protein that is a subunit of a large multimeric complex, termed the retromer complex, involved in retrograde transport of proteins from endosomes to the trans-Golgi network. We report the cloning and characterization of human orthologs of three additional components of the complex: Vps26p, Vps29p, and Vps35p. The close structural similarity between the yeast and human proteins suggests a similarity in function. We used both yeast two-hybrid assays and expression in mammalian cells to define the binding interactions among these proteins. The data suggest a model in which hVps35 serves as the core of a multimeric complex by binding directly to hVps26, hVps29, and SNX1. Deletional analyses of hVps35 demonstrate that amino acid residues 1-53 and 307-796 of hVps35 bind to the coiled coil containing domain of SNX1. In contrast, hVps26 binds to amino acid residues 1-172 of hVps35, whereas hVps29 binds to amino acid residues 307-796 of hVps35. Furthermore, hVps35, hVps29, and hVps26 have been found in membrane-associated and cytosolic compartments. Gel filtration chromatography of COS7 cell cytosol showed that both recombinant and endogenous hVps35, hVps29, and hVps26 coelute as a large complex ( approximately 220-440 kDa). In the absence of hVps35, neither hVps26 nor hVps29 is found in the large complex. These data provide the first insights into the binding interactions among subunits of a putative mammalian retromer complex. PMID- 11102512 TI - Mutation of cyclin/cdk phosphorylation sites in HsCdc6 disrupts a late step in initiation of DNA replication in human cells. AB - Cyclin-dependent kinases (Cdk) are essential for promoting the initiation of DNA replication, presumably by phosphorylating key regulatory proteins that are involved in triggering the G1/S transition. Human Cdc6 (HsCdc6), a protein required for initiation of DNA replication, is phosphorylated by Cdk in vitro and in vivo. Here we report that HsCdc6 with mutations at potential Cdk phosphorylation sites was poorly phosphorylated in vitro by Cdk, but retained all other biochemical activities of the wild-type protein tested. Microinjection of mutant HsCdc6 proteins into human cells blocked initiation of DNA replication or slowed S phase progression. The inhibitory effect of mutant HsCdc6 was lost at the G1/S transition, indicating that phosphorylation of HsCdc6 by Cdk is critical for a late step in initiation of DNA replication in human cells. PMID- 11102513 TI - Complex phenotype of mice lacking occludin, a component of tight junction strands. AB - Occludin is an integral membrane protein with four transmembrane domains that is exclusively localized at tight junction (TJ) strands. Here, we describe the generation and analysis of mice carrying a null mutation in the occludin gene. Occludin -/- mice were born with no gross phenotype in the expected Mendelian ratios, but they showed significant postnatal growth retardation. Occludin -/- males produced no litters with wild-type females, whereas occludin -/- females produced litters normally when mated with wild-type males but did not suckle them. In occludin -/- mice, TJs themselves did not appear to be affected morphologically, and the barrier function of intestinal epithelium was normal as far as examined electrophysiologically. However, histological abnormalities were found in several tissues, i.e., chronic inflammation and hyperplasia of the gastric epithelium, calcification in the brain, testicular atrophy, loss of cytoplasmic granules in striated duct cells of the salivary gland, and thinning of the compact bone. These phenotypes suggested that the functions of TJs as well as occludin are more complex than previously supposed. PMID- 11102514 TI - Expression and partial characterization of kinesin-related proteins in differentiating and adult skeletal muscle. AB - Using pan-kinesin antibodies to screen a differentiating C2C12 cell library, we identified the kinesin proteins KIF3A, KIF3B, and conventional kinesin heavy chain to be present in differentiating skeletal muscle. We compared the expression and subcellular localization characteristics of these kinesins in myogenic cells to others previously identified in muscle, neuronal, and mitotic systems (KIF1C, KIF3C, and mitotic-centromere-associated kinesin). Because members of the KIF3 subfamily of kinesin-related proteins showed altered subcellular fractionation characteristics in differentiating cells, we focused our study of kinesins in muscle on the function of kinesin-II. Kinesin-II is a motor complex comprised of dimerized KIF3A and KIF3B proteins and a tail associated protein, KAP. The Xenopus homologue of KIF3B, Xklp3, is predominantly localized to the region of the Golgi apparatus, and overexpression of motorless Xklp3 in Xenopus A6 cells causes mislocalization of Golgi components (). In C2C12 myoblasts and myotubes, KIF3B is diffuse and punctate, and not primarily associated with the Golgi. Overexpression of motorless-KIF3B does not perturb localization of Golgi components in myogenic cells, and myofibrillogenesis is normal. In adult skeletal muscle, KIF3B colocalizes with the excitation contraction-coupling membranes. We propose that these membranes, consisting of the transverse-tubules and sarcoplasmic reticulum, are dynamic structures in which kinesin-II may function to actively assemble and maintain in myogenic cells. PMID- 11102515 TI - Self-association of coilin reveals a common theme in nuclear body localization. AB - We have found that coilin, the marker protein for Cajal bodies (coiled bodies, CBs), is a self-interacting protein, and we have mapped the domain responsible for this activity to the amino-terminus. Together with a nuclear localization signal, the self-interaction domain is necessary and sufficient for localization to CBs. Overexpression of various wild-type and mutant coilin constructs in HeLa cells results in disruption of both CBs and survival motor neurons (SMN) gems. Additionally, we have identified a cryptic nucleolar localization signal (NoLS), within the coilin protein, which may be exposed in specific coilin phospho isoforms. The implications of these findings are discussed in light of the fact that other proteins known to localize within nuclear bodies (e. g., PML, SMN and Sam68) can also self-associate. Thus protein self-interaction appears to be a general feature of nuclear body marker proteins. PMID- 11102516 TI - Identification of ribonucleotide reductase protein R1 as an activator of microtubule nucleation in Xenopus egg mitotic extracts. AB - Microtubule nucleation on the centrosome and the fungal equivalent, the spindle pole body (SPB), is activated at the onset of mitosis. We previously reported that mitotic extracts prepared from Xenopus unfertilized eggs convert the interphase SPB of fission yeast into a competent state for microtubule nucleation. In this study, we have purified an 85-kDa SPB activator from the extracts and identified it as the ribonucleotide reductase large subunit R1. We further confirmed that recombinant mouse R1 protein was also effective for SPB activation. On the other hand, another essential subunit of ribonucleotide reductase, R2 protein, was not required for SPB activation. SPB activation by R1 protein was suppressed in the presence of anti-R1 antibodies or a partial oligopeptide of R1; the oligopeptide also inhibited aster formation on Xenopus sperm centrosomes. In accordance, R1 was detected in animal centrosomes by immunofluorescence and immunoblotting with anti-R1 antibodies. In addition, recombinant mouse R1 protein bound to gamma- and alpha/beta-tubulin in vitro. These results suggest that R1 is a bifunctional protein that acts on both ribonucleotide reduction and centrosome/SPB activation. PMID- 11102517 TI - Mammalian meiotic telomeres: protein composition and redistribution in relation to nuclear pores. AB - Mammalian telomeres consist of TTAGGG repeats, telomeric repeat binding factor (TRF), and other proteins, resulting in a protective structure at chromosome ends. Although structure and function of the somatic telomeric complex has been elucidated in some detail, the protein composition of mammalian meiotic telomeres is undetermined. Here we show, by indirect immunofluorescence (IF), that the meiotic telomere complex is similar to its somatic counterpart and contains significant amounts of TRF1, TRF2, and hRap1, while tankyrase, a poly-(ADP ribose)polymerase at somatic telomeres and nuclear pores, forms small signals at ends of human meiotic chromosome cores. Analysis of rodent spermatocytes reveals Trf1 at mouse, TRF2 at rat, and mammalian Rap1 at meiotic telomeres of both rodents. Moreover, we demonstrate that telomere repositioning during meiotic prophase occurs in sectors of the nuclear envelope that are distinct from nuclear pore-dense areas. The latter form during preleptotene/leptotene and are present during entire prophase I. PMID- 11102518 TI - Endosome to Golgi transport of ricin is regulated by cholesterol. AB - We have here studied the role of cholesterol in transport of ricin from endosomes to the Golgi apparatus. Ricin is endocytosed even when cells are depleted for cholesterol by using methyl-beta-cyclodextrin (m beta CD). However, as here shown, the intracellular transport of ricin from endosomes to the Golgi apparatus, measured by quantifying sulfation of a modified ricin molecule, is strongly inhibited when the cholesterol content of the cell is reduced. On the other hand, increasing the level of cholesterol by treating cells with mbetaCD saturated with cholesterol (m beta CD/chol) reduced the intracellular transport of ricin to the Golgi apparatus even more strongly. The intracellular transport routes affected include both Rab9-independent and Rab9-dependent pathways to the Golgi apparatus, since both sulfation of ricin after induced expression of mutant Rab9 (mRab9) to inhibit late endosome to Golgi transport and sulfation of a modified mannose 6-phosphate receptor (M6PR) were inhibited after removal or addition of cholesterol. Furthermore, the structure of the Golgi apparatus was affected by increased levels of cholesterol, as visualized by pronounced vesiculation and formation of smaller stacks. Thus, our results indicate that transport of ricin from endosomes to the Golgi apparatus is influenced by the cholesterol content of the cell. PMID- 11102519 TI - The carboxy-terminal cysteine of the tetraspanin L6 antigen is required for its interaction with SITAC, a novel PDZ protein. AB - PDZ domains are protein modules that mediate protein-protein interactions. Here, we present the identification and characterization of a protein similar to the recently identified PDZ-containing protein TACIP18, which we have named SITAC (similar to TACIP18). SITAC is preferentially expressed in cells of the digestive tract, associated with intracellular membranes. Despite the high degree of sequence identity between the PDZ domains of TACIP18 and those of SITAC, none of the known ligands of the former shows interaction with the latter, as judged by two-hybrid analysis. SITAC interacts with peptides containing bulky hydrophobic amino acids two positions upstream of the C-terminal residue. Surprisingly, SITAC also shows interaction with peptides ending in C, a previously unacknowledged ability of PDZ domains. The sequence -Y-X-C-COOH, bound in vitro by SITAC, is present in the member of the tetraspanin superfamily, the L6 antigen. Coimmunoprecipitation experiments show that SITAC interacts with L6A, but not with an L6A C-terminal mutant, confirming the capacity of SITAC to interact with proteins ending in C. Confocal analysis shows that the interaction between L6A and SITAC is necessary for the precise colocalization of both molecules in the same subcellular compartment. In summary, the characterization of the protein SITAC has unveiled novel sequences recognized by PDZ domains, and it suggests that L6A is a natural ligand of this PDZ protein. PMID- 11102520 TI - Golgi apparatus immunolocalization of endomannosidase suggests post-endoplasmic reticulum glucose trimming: implications for quality control. AB - Trimming of N-linked oligosaccharides by endoplasmic reticulum (ER) glucosidase II is implicated in quality control of protein folding. An alternate glucosidase II-independent deglucosylation pathway exists, in which endo-alpha-mannosidase cleaves internally the glucose-substituted mannose residue of oligosaccharides. By immunogold labeling, we detected most endomannosidase in cis/medial Golgi cisternae (83.8% of immunogold labeling) and less in the intermediate compartment (15.1%), but none in the trans-Golgi apparatus and ER, including its transitional elements. This dual localization became more pronounced under 15 degrees C conditions indicative of two endomannosidase locations. Under experimental conditions when the intermediate compartment marker p58 was retained in peripheral sites, endomannosidase was redistributed to the Golgi apparatus. Double immunogold labeling established a mutually exclusive distribution of endomannosidase and glucosidase II, whereas calreticulin was observed in endomannosidase-reactive sites (17.3% in intermediate compartment, 5.7% in Golgi apparatus) in addition to the ER (77%). Our results demonstrate that glucose trimming of N-linked oligosaccharides is not limited to the ER and that protein deglucosylation by endomannosidase in the Golgi apparatus and intermediate compartment additionally ensures that processing to mature oligosaccharides can continue. Thus, endomannosidase localization suggests that a quality control of N glycosylation exists in the Golgi apparatus. PMID- 11102521 TI - Genomic expression programs in the response of yeast cells to environmental changes. AB - We explored genomic expression patterns in the yeast Saccharomyces cerevisiae responding to diverse environmental transitions. DNA microarrays were used to measure changes in transcript levels over time for almost every yeast gene, as cells responded to temperature shocks, hydrogen peroxide, the superoxide generating drug menadione, the sulfhydryl-oxidizing agent diamide, the disulfide reducing agent dithiothreitol, hyper- and hypo-osmotic shock, amino acid starvation, nitrogen source depletion, and progression into stationary phase. A large set of genes (approximately 900) showed a similar drastic response to almost all of these environmental changes. Additional features of the genomic responses were specialized for specific conditions. Promoter analysis and subsequent characterization of the responses of mutant strains implicated the transcription factors Yap1p, as well as Msn2p and Msn4p, in mediating specific features of the transcriptional response, while the identification of novel sequence elements provided clues to novel regulators. Physiological themes in the genomic responses to specific environmental stresses provided insights into the effects of those stresses on the cell. PMID- 11102522 TI - Exocyst is involved in cystogenesis and tubulogenesis and acts by modulating synthesis and delivery of basolateral plasma membrane and secretory proteins. AB - Epithelial cyst and tubule formation are critical processes that involve transient, highly choreographed changes in cell polarity. Factors controlling these changes in polarity are largely unknown. One candidate factor is the highly conserved eight-member protein complex called the exocyst. We show that during tubulogenesis in an in vitro model system the exocyst relocalized along growing tubules consistent with changes in cell polarity. In yeast, the exocyst subunit Sec10p is a crucial component linking polarized exocytic vesicles with the rest of the exocyst complex and, ultimately, the plasma membrane. When the exocyst subunit human Sec10 was exogenously expressed in epithelial Madin-Darby canine kidney cells, there was a selective increase in the synthesis and delivery of apical and basolateral secretory proteins and a basolateral plasma membrane protein, but not an apical plasma membrane protein. Overexpression of human Sec10 resulted in more efficient and rapid cyst formation and increased tubule formation upon stimulation with hepatocyte growth factor. We conclude that the exocyst plays a central role in the development of epithelial cysts and tubules. PMID- 11102523 TI - The sodium/proton exchanger Nhx1p is required for endosomal protein trafficking in the yeast Saccharomyces cerevisiae. AB - We show that the vacuolar protein sorting gene VPS44 is identical to NHX1, a gene that encodes a sodium/proton exchanger. The Saccharomyces cerevisiae protein Nhx1p shows high homology to mammalian sodium/proton exchangers of the NHE family. Nhx1p is thought to transport sodium ions into the prevacuole compartment in exchange for protons. Pulse-chase experiments show that approximately 35% of the newly synthesized soluble vacuolar protein carboxypeptidase Y is missorted in nhx1 delta cells, and is secreted from the cell. nhx1 delta cells accumulate late Golgi, prevacuole, and lysosome markers in an aberrant structure next to the vacuole, and late Golgi proteins are proteolytically cleaved more rapidly than in wild-type cells. Our results show that efficient transport out of the prevacuolar compartment requires Nhx1p, and that nhx1 delta cells exhibit phenotypes characteristic of the "class E" group of vps mutants. In addition, we show that Nhx1p is required for protein trafficking even in the absence of the vacuolar ATPase. Our analysis of Nhx1p provides the first evidence that a sodium/proton exchange protein is important for correct protein sorting, and that intraorganellar ion balance may be important for endosomal function in yeast. PMID- 11102524 TI - The latent transforming growth factor-beta-binding protein-1 promotes in vitro differentiation of embryonic stem cells into endothelium. AB - The latent transforming growth factor-beta-binding protein-1 (LTBP-1) belongs to a family of extracellular glycoproteins that includes three additional isoforms (LTBP-2, -3, and -4) and the matrix proteins fibrillin-1 and -2. Originally described as a TGF-beta-masking protein, LTBP-1 is involved both in the sequestration of latent TGF-beta in the extracellular matrix and the regulation of its activation in the extracellular environment. Whereas the expression of LTBP-1 has been analyzed in normal and malignant cells and rodent and human tissues, little is known about LTBP-1 in embryonic development. To address this question, we used murine embryonic stem (ES) cells to analyze the appearance and role of LTBP-1 during ES cell differentiation. In vitro, ES cells aggregate to form embryoid bodies (EBs), which differentiate into multiple cell lineages. We analyzed LTBP-1 gene expression and LTBP-1 fiber appearance with respect to the emergence and distribution of cell types in differentiating EBs. LTBP-1 expression increased during the first 12 d in culture, appeared to remain constant between d 12 and 24, and declined thereafter. By immunostaining, fibrillar LTBP-1 was observed in those regions of the culture containing endothelial, smooth muscle, and epithelial cells. We found that inclusion of a polyclonal antibody to LTBP-1 during EB differentiation suppressed the expression of the endothelial specific genes ICAM-2 and von Willebrand factor and delayed the organization of differentiated endothelial cells into cord-like structures within the growing EBs. The same effect was observed when cultures were treated with either antibodies to TGF-beta or the latency associated peptide, which neutralize TGF-beta. Conversely, the organization of endothelial cells was enhanced by incubation with TGF-beta 1. These results suggest that during differentiation of ES cells LTBP-1 facilitates endothelial cell organization via a TGF-beta-dependent mechanism. PMID- 11102525 TI - New components of a system for phosphate accumulation and polyphosphate metabolism in Saccharomyces cerevisiae revealed by genomic expression analysis. AB - The PHO regulatory pathway is involved in the acquisition of phosphate (P(i)) in the yeast Saccharomyces cerevisiae. When extracellular P(i) concentrations are low, several genes are transcriptionally induced by this pathway, which includes the Pho4 transcriptional activator, the Pho80-Pho85 cyclin-CDK pair, and the Pho81 CDK inhibitor. In an attempt to identify all the components regulated by this system, a whole-genome DNA microarray analysis was employed, and 22 PHO regulated genes were identified. The promoter regions of 21 of these genes contained at least one copy of a sequence that matched the Pho4 recognition site. Eight of these genes, PHM1-PHM8, had no previously defined function in phosphate metabolism. The amino acid sequences of PHM1 (YFL004w), PHM2 (YPL019c), PHM3 (YJL012c), and PHM4 (YER072w) are 32-56% identical. The phm3 and phm4 single mutants and the phm1 phm2 double mutant were each severely deficient in accumulation of inorganic polyphosphate (polyP) and P(i). The phenotype of the phm5 mutant suggests that PHM5 (YDR452w) is essential for normal catabolism of polyP in the yeast vacuole. Taken together, the results reveal important new features of a genetic system that plays a critical role in P(i) acquisition and polyP metabolism in yeast. PMID- 11102526 TI - Head and/or CaaX domain deletions of lamin proteins disrupt preformed lamin A and C but not lamin B structure in mammalian cells. AB - The nuclear lamina is an important determinant of nuclear architecture. Mutations in A-type but not B-type lamins cause a range of human genetic disorders, including muscular dystrophy. Dominant mutations in nuclear lamin proteins have been shown to disrupt a preformed lamina structure in Xenopus egg extracts. Here, a series of deletion mutations in lamins A and B1 were evaluated for their ability to disrupt lamina structure in Chinese hamster ovary cells. Deletions of either the lamin A "head" domain or the C-terminal CaaX domain formed intranuclear aggregates and resulted in the disruption of endogenous lamins A/C but not lamins B1/B2. By contrast, "head-less" lamin B1 localized to the nuclear rim with no detectable effect on endogenous lamins, whereas lamin B1 CaaX domain deletions formed intranuclear aggregates, disrupting endogenous lamins A/C but not lamins B1/B2. Filter binding assays revealed that a head/CaaX domain lamin B1 mutant interacted much more strongly with lamins A/C than with lamins B1/B2. Regulated induction of this mutant in stable cell lines resulted in the rapid elimination of all detectable lamin A protein, whereas lamin C was trapped in a soluble form within the intranuclear aggregates. In contrast to results in Xenopus egg extracts, dominant negative lamin B1 (but not lamin A) mutants trapped replication proteins involved in both the initiation and elongation phases of replication but did not effect cellular growth rates or the assembly of active replication centers. We conclude that elimination of the CaaX domain in lamin B1 and elimination of either the CaaX or head domain in lamin A constitute dominant mutations that can disrupt A-type but not B-type lamins, highlighting important differences in the way that A- and B-type lamins are integrated into the lamina. PMID- 11102527 TI - A decrease in membrane tension precedes successful cell-membrane repair. AB - We hypothesized that the requirement for Ca(2+)-dependent exocytosis in cell membrane repair is to provide an adequate lowering of membrane tension to permit membrane resealing. We used laser tweezers to form membrane tethers and measured the force of those tethers to estimate the membrane tension of Swiss 3T3 fibroblasts after membrane disruption and during resealing. These measurements show that, for fibroblasts wounded in normal Ca(2+) Ringer's solution, the membrane tension decreased dramatically after the wounding and resealing coincided with a decrease of approximately 60% of control tether force values. However, the tension did not decrease if cells were wounded in a low Ca(2+) Ringer's solution that inhibited both membrane resealing and exocytosis. When cells were wounded twice in normal Ca(2+) Ringer's solution, decreases in tension at the second wound were 2.3 times faster than at the first wound, correlating well with twofold faster resealing rates for repeated wounds. The facilitated resealing to a second wound requires a new vesicle pool, which is generated via a protein kinase C (PKC)-dependent and brefeldin A (BFA)-sensitive process. Tension decrease at the second wound was slowed or inhibited by PKC inhibitor or BFA. Lowering membrane tension by cytochalasin D treatment could substitute for exocytosis and could restore membrane resealing in low Ca(2+) Ringer's solution. PMID- 11102528 TI - Apoptosis induced by Rac GTPase correlates with induction of FasL and ceramides production. AB - Rho proteins, members of the Ras superfamily of GTPases, are critical elements in signal transduction pathways governing cell proliferation and cell death. Different members of the family of human Rho GTPases, including RhoA, RhoC, and Rac1, participate in the regulation of apoptosis in response to cytokines and serum deprivation in different cell systems. Here, we have characterized the mechanism of apoptosis induced by Rac1 in NIH 3T3 cells. It requires protein synthesis and caspase-3 activity, but it is independent of the release of cytochrome c from mitochondria. Moreover, an increase in mitochondria membrane potential and the production of reactive oxygen species was observed. Rac1 induced apoptosis was related to the simultaneous increase in ceramide production and synthesis of FasL. Generation of FasL may be mediated by transcriptional regulation involving both c-Jun amino terminal kinase as well as nuclear factor kappa B-dependent signals. None of these signals, ceramides or FasL, was sufficient to induce apoptosis in the parental cell line, NIH 3T3 cells. However, any of them was sufficient to induce apoptosis in the Rac1-expressing cells. Finally, inhibition of FasL signaling drastically reduced apoptosis by Rac1. Thus, Rac1 seems to induce apoptosis by a complex mechanism involving the generation of ceramides and the de novo synthesis of FasL. These results suggest that apoptosis mediated by Rac1 results from a signaling mechanism that involves biochemical and transcriptional events under control of Rac1. PMID- 11102529 TI - Dual requirement for rho and protein kinase C in direct activation of phospholipase D1 through G protein-coupled receptor signaling. AB - G protein-coupled and tyrosine kinase receptor activation of phospholipase D1 (PLD1) play key roles in agonist-stimulated cellular responses such as regulated exocytosis, actin stress fiber formation, and alterations in cell morphology and motility. Protein Kinase C, ADP-ribosylation factor (ARF), and Rho family members activate PLD1 in vitro; however, the actions of the stimulators on PLD1 in vivo have been proposed to take place through indirect pathways. We have used the yeast split-hybrid system to generate PLD1 alleles that fail to bind to or to be activated by RhoA but that retain wild-type responses to ARF and PKC. These alleles then were employed in combination with alleles unresponsive to PKC or to both stimulators to examine the activation of PLD1 by G protein-coupled receptors. Our results demonstrate that direct stimulation of PLD1 in vivo by RhoA (and by PKC) is critical for significant PLD1 activation but that PLD1 subcellular localization and regulated phosphorylation occur independently of these stimulatory pathways. PMID- 11102530 TI - Cytoplasmic and nuclear phospholipase C-beta 1 relocation: role in resumption of meiosis in the mouse oocyte. AB - The location of the phospholipase C beta 1-isoform (PLC-beta 1) in the mouse oocyte and its role in the resumption of meiosis were examined. We used specific monoclonal antibodies to monitor the in vitro dynamics of the subcellular distribution of the enzyme from the release of the oocyte from the follicle until breakdown of the germinal vesicle (GVBD) by Western blotting, electron microscope immunohistochemistry, and confocal microscope immunofluorescence. PLC-beta 1 became relocated to the oocyte cortex and the nucleoplasm during the G2/M transition, mainly in the hour preceding GVBD. The enzyme was a 150-kDa protein, corresponding to PLC-beta 1a. Its synthesis in the cytoplasm increased during this period, and it accumulated in the nucleoplasm. GVBD was dramatically inhibited by the microinjection of anti-PLC-beta1 monoclonal antibody into the germinal vesicle (GV) only when this accumulation was at its maximum. In contrast, PLC-gamma 1 was absent from the GV from the time of release from the follicle until 1 h later, and microinjection of anti-PLC-gamma 1 into the GV did not affect GVBD. Our results demonstrate a relationship between the relocation of PLC-beta 1 and its role in the first step of meiosis. PMID- 11102531 TI - Identification of a novel transcription factor binding element involved in the regulation by differentiation of the human telomerase (hTERT) promoter. AB - Three different cell differentiation experimental model systems (human embryonic stem cells, mouse F9 cells, and human HL-60 promyelocytic cells) were used to determine the relationship between the reduction in telomerase activity after differentiation and the regulation of the promoter for the hTERT gene. Promoter constructs of three different lengths were subcloned into the PGL3-basic luciferase reporter vector. In all three experimental systems, all three promoter constructs drove high levels of reporter activity in the nondifferentiated state, with a marked and time-dependent reduction after the induction of differentiation. In all cases, the smallest core promoter construct (283 nt upstream of the ATG) gave the highest activity. Electrophoretic mobility shift assays revealed transcription factor binding to two E-box domains within the core promoter. There was also a marked time-dependent reduction in this binding with differentiation. In addition, a distinct and novel element was identified within the core promoter, which also underwent time-dependent reduction in transcription factor binding with differentiation. Site-directed mutagenesis of this novel element revealed a correlation between transcription factor binding and promoter activity. Taken together, the results indicate that regulation of overall telomerase activity with differentiation is mediated at least in part at the level of the TERT promoter and provides new information regarding details of the regulatory interactions that are involved in this process. PMID- 11102532 TI - Schizosaccharomyces pombe rho2p GTPase regulates cell wall alpha-glucan biosynthesis through the protein kinase pck2p. AB - Schizosaccharomyces pombe rho1(+) and rho2(+) genes are involved in the control of cell morphogenesis, cell integrity, and polarization of the actin cytoskeleton. Although both GTPases interact with each of the two S. pombe protein kinase C homologues, Pck1p and Pck2p, their functions are distinct from each other. It is known that Rho1p regulates (1,3)beta-D-glucan synthesis both directly and through Pck2p. In this paper, we have investigated Rho2p signaling and show that pck2 delta and rho2 delta strains display similar defects with regard to cell wall integrity, indicating that they might be in the same signaling pathway. We also show that Rho2 GTPase regulates the synthesis of alpha D-glucan, the other main structural polymer of the S. pombe cell wall, primarily through Pck2p. Although overexpression of rho2(+) in wild-type or pck1 delta cells is lethal and causes morphological alterations, actin depolarization, and an increase in alpha-D-glucan biosynthesis, all of these effects are suppressed in a pck2 delta strain. In addition, genetic interactions suggest that Rho2p and Pck2p are important for the regulation of Mok1p, the major (1-3)alpha-D-glucan synthase. Thus, a rho2 delta mutation, like pck2 delta, is synthetically lethal with mok1-664, and the mutant partially fails to localize Mok1p to the growing areas. Moreover, overexpression of mok1(+) in rho2 delta cells causes a lethal phenotype that is completely different from that of mok1(+) overexpression in wild-type cells, and the increase in alpha-glucan is considerably lower. Taken together, all of these results indicate the presence of a signaling pathway regulating alpha-glucan biosynthesis in which the Rho2p GTPase activates Pck2p, and this kinase in turn controls Mok1p. PMID- 11102534 TI - Smoking and sleep position are only pieces of the puzzle resulting in the Sudden Infant Death Syndrome. PMID- 11102535 TI - In some cases, once may be enough. PMID- 11102533 TI - The TRAPP complex is a nucleotide exchanger for Ypt1 and Ypt31/32. AB - In yeast, the Ypt1 GTPase is required for ER-to-cis-Golgi and cis-to-medial-Golgi protein transport, while Ypt31/32 are a functional pair of GTPases essential for exit from the trans-Golgi. We have previously identified a Ypt1 guanine nucleotide exchange factor (GEF) activity and characterized it as a large membrane-associated protein complex that localizes to the Golgi and can be extracted from the membrane by salt, but not by detergent. TRAPP is a large protein complex that is required for ER-to-Golgi transport and that has properties similar to those of Ypt1 GEF. Here we show that TRAPP has Ypt1 GEF activity. GST-tagged Bet3p or Bet5p, two of the TRAPP subunits, were expressed in yeast cells and were precipitated by glutathione-agarose (GA) beads. The resulting precipitates can stimulate both GDP release and GTP uptake by Ypt1p. The majority of the Ypt1 GEF activity associated with the GST-Bet3p precipitate has an apparent molecular weight of > 670 kDa, indicating that the GEF activity resides in the TRAPP complex. Surprisingly, TRAPP can also stimulate nucleotide exchange on the Ypt31/32 GTPases, but not on Sec4p, a Ypt-family GTPase required for the last step of the exocytic pathway. Like the previously characterized Ypt1 GEF, the TRAPP Ypt1-GEF activity can be inhibited by the nucleotide-free Ypt1 D124N mutant protein. This mutant protein also inhibits the Ypt32 GEF activity of TRAPP. Coprecipitation and overexpression studies suggest that TRAPP can act as a GEF for Ypt1 and Ypt31/32 in vivo. These data suggest the exciting possibility that a GEF complex common to Ypt1 and Ypt31/32 might coordinate the function of these GTPases in entry into and exit from the Golgi. PMID- 11102536 TI - Transcriptional regulation of cardiac development: implications for congenital heart disease and DiGeorge syndrome. AB - In recent years, impressive advances have occurred in our understanding of transcriptional regulation of cardiac development. These insights have begun to elucidate the mystery of congenital heart disease at the molecular level. In addition, the molecular pathways emerging from the study of cardiac development are being applied to the understanding of adult cardiac disease. Preliminary results support the contention that a thorough understanding of molecular programs governing cardiac morphogenesis will provide important insights into the pathogenesis of human cardiac diseases. This review will focus on examples of transcription factors that play critical roles at various phases of cardiac development and their relevance to cardiac disease. This is an exciting and burgeoning area of investigation. It is not possible to be all-inclusive, and the reader will note important efforts in the areas of cardiomyocyte determination, left-right asymmetry, cardiac muscular dystrophies, electrophysiology and vascular disease are not covered. For a more complete discussion, the reader is referred to recent reviews including the excellent compilation of observations assembled by Harvey and Rosenthal (1). PMID- 11102537 TI - Mechanisms and disturbances of neuronal migration. AB - Neuronal migration appears as a complex ontogenic step occurring early during embryonic and fetal development. Control of neuronal migration involves different cell populations including Cajal-Retzius neurons, subplate neurons, neuronal precursors or radial glia. The integrity of multiple molecular mechanisms, such as cell cycle control, cell-cell adhesion, interaction with extracellular matrix protein, neurotransmitter release, growth factor availability, platelet activating factor degradation or transduction pathways seems to be critical for normal neuronal migration. The complexity and the multiplicity of these mechanisms probably explain the clinical, radiologic and genetic heterogeneity of human disorders of neuronal migration. The present review will be focused on mechanisms and disturbances of migration of neurons destined to the neocortex. New insights gained from the analysis of animal models as well as from the study of human diseases will be included. PMID- 11102538 TI - Amelioration of intestinal disease severity in cystic fibrosis mice is associated with improved chloride secretory capacity. AB - The variability in intestinal disease severity in patients with cystic fibrosis (CF) has been associated with the expression of secondary modifier genes. The locus containing these modifier genes in CF patients is syntenic with a modifier locus previously associated with survival in CF transmembrane conductance regulator-knockout mice. These previous studies showed that the proportion of CF mice that survive weaning (mildly affected mice) versus those that succumb to obstruction of the small intestine (severely affected) is related to their genetic background and the expression of modifier genes. In the present work, we show that the basal transepithelial chloride transport measured across jejuna obtained from mice of mixed genetic backgrounds segregates into two groups, some mice having low and others having high, near normal chloride transport. Further, we report that the segregation of mice with respect to intestinal chloride transport correlates with their predicted segregation on the basis of genotype at the "modifier locus." These findings support the hypothesis that intestinal disease modification in CF mice correlates with improved chloride transport through non-CF transmembrane conductance regulator chloride channels. PMID- 11102539 TI - Parallel secretion of pancreatic phospholipase A(2), phospholipase A(1), lipase, and colipase in children with exocrine pancreatic dysfunction. AB - The cosecretion of pancreatic lipase and colipase are important in normal fat digestion. As adsorption of phosphatidylcholine to the lipid substrate interferes with lipase activity, hydrolysis to lysophosphatidylcholine with subsequent desorption is also essential for fat digestion. There are some data regarding the secretion of pancreatic phospholipases in normal adults but none in children or patients with pancreatic disease. In the present study, we aimed a) to develop an accurate fast assay method to measure phospholipase A(2) and b) to determine the secretion rate of pancreatic phospholipase A(2) and whether it is cosecreted with lipase and colipase in children with exocrine pancreatic dysfunction. Nine male patients aged 0.5 to 16 y (seven with cystic fibrosis, two with malabsorption) underwent pancreatic stimulation tests. Their colipase and lipase secretion rates were measured by titrimetric methods and phospholipase A(2) and A(1) by phosphorus magnetic resonance spectroscopy ((31)P NMR). It was found that the phospholipases, colipase, and lipase were absent in the two patients with pancreatic insufficiency. In patients with normal absorption, there were marked inter-and intrasubject variations of lipase, colipase, and phospholipase secretion rates that were consistent with the degree of exocrine pancreatic dysfunction. However, in the three 20-min stimulation periods of the pancreatic function test, pancreatic phospholipase is cosecreted with lipase and colipase, and average colipase and phospholipase A(2) secretion rates follow a similar or parallel pattern. These findings are consistent with the important role of pancreatic phospholipases in intestinal phospholipid hydrolysis leading to the desorption of phospholipids from the lipid substrate and enhancing lipid hydrolysis and phospholipid absorption. PMID- 11102540 TI - Long-time persistence of superantigen-producing Staphylococcus aureus strains in the intestinal microflora of healthy infants. AB - Staphylococcus aureus has been isolated at an increasing rate from infants' stools during the last decades, but it is not known whether this species can colonize and persist in the intestinal microflora. To investigate this, 49 Swedish infants were followed prospectively from birth until 12 months of age. S. aureus was identified in a rectal swab obtained 3 d after delivery and in quantitative cultures of fecal samples collected at 1, 2, 4, and 8 weeks and at 6 and 12 months of age. A random amplified polymorphic DNA (RAPD) method was developed to distinguish individual S. aureus strains from one another and the strains were tested for production of enterotoxins A-D and TSST-1. By 3 days of age, 16% of infants had S. aureus in their intestines, which increased to 73% by 2-6 months, whereafter it decreased slightly to 53%. At the same time S. aureus population counts in colonized infants declined from an average 10(6.8) CFU/g feces during the first months of life to 10(4.0) CFU/g feces by 12 months. Colonized infants usually harbored one or two S. aureus strains in their microflora for long periods of time. Few strains were transient passengers and the median time of persistence of S. aureus strains in the microflora was several months. Of the 75 S. aureus strains identified, 43% produced one or more toxins: 13% SEA, 7% SEB, 23% SEC, 4% SED, and 11% TSST-1. Altogether, 47% of the investigated infants were colonized by a toxin-producing S. aureus during their first year of life. Despite this they were apparently healthy and did not have more gastrointestinal problems than noncolonized infants. This report is the first to show that S. aureus may be a resident member of the normal intestinal microflora in infancy. PMID- 11102541 TI - Analysis of exonic mutations leading to exon skipping in patients with pyruvate dehydrogenase E1 alpha deficiency. AB - The pyruvate dehydrogenase (PDH) complex is situated at a key position in energy metabolism and is responsible for the conversion of pyruvate to acetyl CoA. In the literature, two unrelated patients with a PDH complex deficiency and splicing out of exon 6 of the PDH E1 alpha gene have been described, although intronic/exonic boundaries on either side of exon 6 were completely normal. Analysis of exon 6 in genomic DNA of these patients revealed two exonic mutations, a silent and a missense mutation. Although not experimentally demonstrated, the authors in both publications suggested that the exonic mutations were responsible for the exon skipping. In this work, we were able to demonstrate, by performing splicing experiments, that the two exonic mutations described in the PDH E1 alpha gene lead to aberrant splicing. We observed a disruption of the predicted wild-type pre-mRNA secondary structure of exon 6 by the mutated sequences described. However, when we constructed mutations that either reverted or disrupted the wild-type predicted pre-mRNA secondary structure of exon 6, we were unable to establish a correlation between the aberrant splicing and disruption of the predicted structure. The mutagenic experiments described here and the silent mutation found in one of the patients suggest the presence of an exonic splicing enhancer in the middle region of exon 6 of the PDH E1alpha gene. PMID- 11102542 TI - Mutations in the chloride channel gene, CLCNKB, leading to a mixed Bartter Gitelman phenotype. AB - Gitelman syndrome is an inherited renal disorder characterized by impaired NaCl reabsorption in the distal convoluted tubule and secondary hypokalemic alkalosis. In clinical practice, it is distinguished from other hypokalemic tubulopathies by the presence of both hypomagnesemia and normocalcemic hypocalciuria. To date, only mutations in a single gene encoding the thiazide-sensitive NaCl cotransporter have been found as the molecular basis of GS. We describe three unrelated patients presenting with the typical laboratory findings of GS. Mutational analysis in these patients revealed no abnormality in the SLC12A3 gene. Instead, all patients were found to carry previously described mutations in the CLCNKB gene, which encodes the kidney-specific chloride channel ClC-Kb, raising the possibility of genetic heterogeneity. Review of the medical histories revealed manifestation of the disease within the first year of life in all cases. Clinical presentation included episodes of dehydration, weakness, and failure to thrive, much more suggestive of classic Bartter syndrome than of GS. The coexistence of hypomagnesemia and hypocalciuria was not present from the beginning. In the follow-up, however, a drop of both parameters below normal range was a consistent finding reflecting a transition from cBS to GS phenotype. The phenotypic overlap may indicate a physiologic cooperation of the apical thiazide-sensitive NaCl cotransporter and the basolateral chloride channel for salt reabsorption in the distal convoluted tubule. PMID- 11102543 TI - Alternatively spliced nephrin in experimental glomerular disease of the rat. AB - Nephrin is a novel transmembrane protein of kidney glomerular podocytes, which appears crucially important for the maintenance of the glomerular filtration barrier. According to its predicted structure, nephrin has additional roles in cell-cell adhesion and/or signal transduction. We have previously cloned the rat homologue of nephrin and described its alternatively spliced transcripts alpha and beta. In this study we examined the alterations in expression and regulation of particularly the major alternatively spliced nephrin-alpha giving rise to a variant lacking the membrane spanning domain in the puromycin nephrosis of the rat. A down-regulation of up to 78% was observed of the full length mRNA after 10 d of PAN treatment. The expression changes of nephrin-alpha followed closely the expression of the full length mRNA. Interestingly, we also found nephrin protein in urine at the peak proteinuria samples of this model. These results suggest that soluble nephrin variants may be important markers for proteinuric diseases. PMID- 11102544 TI - Right ventricular injury in young swine: effects of catecholamines on right ventricular function and pulmonary vascular mechanics. AB - Acute right ventricular (RV) injury is commonly encountered in infants and children after cardiac surgery. Empiric medical therapy for these patients results from a paucity of data on which to base medical management and the absence of animal models that allow rigorous laboratory testing. Specifically, exogenous catecholamines have unclear effects on the injured right ventricle and pulmonary vasculature in the young. Ten anesthetized piglets (9-12 kg) were instrumented with epicardial transducers, micromanometers, and a pulmonary artery flow probe. RV injury was induced with a cryoablation probe. Dopamine at 10 microg/kg/min, dobutamine at 10 microg/kg/min, and epinephrine (EP) at 0.1 microg/kg/min were infused in a random order. RV contractility was evaluated using preload recruitable stroke work. Diastolic function was described by the end-diastolic pressure-volume relation, peak negative derivative of the pressure waveform, and peak filling rate. In addition to routine hemodynamic measurements, Fourier transformation of the pressure and flow waveforms allowed calculation of input resistance, characteristic impedance, RV total hydraulic power, and transpulmonary vascular efficiency. Cryoablation led to a stable reproducible injury, decreased preload recruitable stroke work, and impaired diastolic function as measured by all three indices. Infusion of each catecholamine improved preload recruitable stroke work and peak negative derivative of the pressure waveform. Dobutamine and EP both decreased indices of pulmonary vascular impedance, whereas EP was the only inotrope that significantly improved transpulmonary vascular efficiency. Although all three inotropes improved systolic and diastolic RV function, only EP decreased input resistance, decreased pulmonary vascular resistance, and increased transpulmonary vascular efficiency. PMID- 11102545 TI - Evidence for autosomal recessive inheritance of infantile dilated cardiomyopathy: studies from the Eastern Province of Saudi Arabia. AB - Familial dilated cardiomyopathy is being increasingly recognized, but affected individuals <10 y are rarely identified. We describe the natural history of dilated cardiomyopathy and evaluate the mode of inheritance among infants of Arab descent from the Eastern Province of Saudi Arabia. We evaluated 55 consecutive cases of dilated cardiomyopathy in patients <10 y of age seen during a 5-y interval. Echocardiography was the primary diagnostic modality. The 55 cases represented 20% of the offspring of 41 families of Arab descent. In 19 families (46%), parents were first cousins; there was no obvious consanguinity in 22 families (54%). Age at presentation was <30 mo (95%) (range, 1 to 100 mo); males (38%) and females (62%) were affected. Patients died (25 patients, 46%), improved (15 patients, 27%), or recovered (15 patients, 27%). The left ventricular shortening fraction at diagnosis ranged from 5 to 28% and did not differ in those who died, improved, or recovered. Complex segregation analysis of the family data using the mixed model of inheritance showed that a model of recessive inheritance best fits the data. Recessively inherited dilated cardiomyopathy has been infrequently reported, perhaps because it may be difficult to recognize in other patient groups in which consanguineous marriage is uncommon and the number of children per family is small. In the setting of consanguineous marriage, homozygosity mapping should lead to identification of the gene(s) causing dilated cardiomyopathy in the families we studied. PMID- 11102546 TI - Recurrent milk aspiration produces changes in airway mechanics, lung eosinophilia, and goblet cell hyperplasia in a murine model. AB - Recurrent aspiration of milk into the respiratory tract has been implicated in the pathogenesis of a variety of inflammatory lung disorders including asthma. However, the lack of animal models of aspiration-induced lung injury has limited our knowledge of the pathophysiological characteristics of this disorder. This study was designed to evaluate the effects of recurrent milk aspiration on airway mechanics and lung cells in a murine model. Under light anesthesia, BALB/c mice received daily intranasal instillations of whole cow's milk (n = 7) or sterile physiologic saline (n = 9) for 10 d. Respiratory system resistance (Rrs) and dynamic elastance (Edyn,rs) were measured in anesthetized, tracheotomized, paralyzed and mechanically ventilated mice 24 h after the last aspiration of milk. Rrs and Edyn,rs were derived from transrespiratory and plethysmographic pressure signals. In addition, airway responses to increasing concentrations of i.v. methacholine (Mch) were determined. Airway responses were measured in terms of PD(100) (dose of Mch causing 100% increase from baseline Rrs) and Rrs,max (% increase from baseline at the maximal plateau response) and expressed as % control (mean +/- SE). We found recurrent milk aspiration did not affect Edyn and baseline Rrs values. However, airway responses to Mch were increased after milk aspiration when compared with control mice. These changes in airway mechanics were associated with an increased percentage of lymphocytes and eosinophils in the bronchoalveolar lavage, mucus production, and lung inflammation. Our findings suggest that recurrent milk aspiration leads to alterations in airway function, lung eosinophilia, and goblet cell hyperplasia in a murine model. PMID- 11102547 TI - Antenatal endotoxin and glucocorticoid effects on lung morphometry in preterm lambs. AB - In utero inflammation may accelerate fetal lung maturation but may also play a role in the pathogenesis of chronic lung disease. We examined the impact of endotoxin, a potent proinflammatory stimulus, on structural and functional maturation of preterm sheep lungs. Date bred ewes received 20 mg Escherichia coli endotoxin or saline by ultrasound guided intra-amniotic injection at 119 d gestation. A comparison group of animals received 0.5 mg/kg betamethasone, a known maturational agent, at 118 d gestation. Lambs were delivered by cesarean section at 125 d (term = 150 d) and ventilated for 40 min. Lung function data are reported elsewhere. Total and differential white cell counts were performed on amniotic fluid and fetal lung fluid samples. Morphometric analyses were performed on inflation fixed right upper lobes. Total cell count increased slightly but not significantly in both amniotic fluid and fetal lung fluid. Both endotoxin and betamethasone had similar effects on alveolarization: average alveolar volume increased by approximately 20% and total alveolar number decreased by almost 30%. Both treatments led to thinning of alveolar walls, although this was statistically significant in the betamethasone-treated group only. Although antenatal endotoxin leads to striking improvements in postnatal lung function, this may be at the expense of normal alveolar development. PMID- 11102548 TI - Antenatal dexamethasone administration increases fetal lung DNA synthesis and RNA and protein content in nitrofen-induced congenital diaphragmatic hernia in rats. AB - Antenatal glucocorticoids treatment has been shown to correct pulmonary immaturity. The thymidine analog bromodeoxyuridine (BrdU) is incorporated into S phase cells and used as a marker of DNA synthesis. In this study, we investigated the effect of antenatal glucocorticoid administration on DNA synthesis and RNA and protein content in nitrofen-induced congenital diaphragmatic hernia (CDH) in rats to better understand the effect of antenatal glucocorticoids on CDH lung. The CDH model was induced in pregnant rats using nitrofen. Dexamethasone (0.25 mg/kg) was given on d 18.5 and 19.5 of gestation (term = 22 d). BrdU was administered 1 h before fetuses were killed on d 21, and detected by immunohistochemistry. DNA synthesis was evaluated by percentage of BrdU incorporated nuclei (BrdU labeling index). Total RNA and soluble protein were extracted from another set of left lungs to measure RNA and protein content. BrdU labeling index and total RNA content were significantly decreased in CDH lung compared with control rats. Antenatal dexamethasone treatment significantly increased BrdU labeling index and RNA and protein content in the left CDH lung. Our findings of decreased DNA synthesis and decreased RNA and protein content in CDH lung suggest that lung growth and development are suppressed in hypoplastic CDH lung. Increased DNA synthesis and increased RNA and protein content in dexamethasone-treated CDH lung suggest that antenatal glucocorticoids may accelerate fetal lung growth and development in CDH. PMID- 11102549 TI - EGF regulates early embryonic mouse gut development in chemically defined organ culture. AB - The profound intestinal epithelial defects in the newborn epidermal growth factor receptor (EGFR) knockout mouse suggests that EGFR signaling plays important roles in embryonic gut development. Herein, we further elucidated the function of EGFR signaling on early embryonic gut development by comparing the effects of 1-10 ng/mL of exogenous epidermal growth factor (EGF) or 10-25 microM of the tyrphostin 3,4,5 trihydroxybenzene malononitrile, a specific inhibitor of EGFR tyrosine kinase, on intact E12 Swiss-Webster mouse midgut grown in chemically defined organ culture using Fitton-Jackson BGJb medium for 4 or 6 d. Intestinal development during culture was assayed by morphometry, histology, reverse transcription/competitive PCR for villin and intestinal fatty acid binding protein mRNA, and immunohistochemistry for epithelial proliferative markers. During organ culture, control specimens grew in length, developed smooth muscle, simple columnar epithelial and goblet cell phenotypes, showed early villus formation in the proximal intestine, and increased expression of villin and intestinal fatty acid binding protein mRNA. EGF failed to significantly alter small intestinal lengthening, whereas EGF 10 ng/mL inhibited colonic length growth. Tyrphostin 25 microM resulted in regional losses of stromal and smooth muscle cells in the small intestine and absent colonic goblet cells. In controls, cellular proliferation initially occurred throughout the small intestinal epithelium but became increasingly localized to the intervillus crypt regions. This sequestration of epithelial proliferation into crypts was much more apparent in EGF-treated versus tyrphostin-treated specimens. EGFR activation, therefore, appears to accelerate the maturation rate of goblet cells and the differential crypt/villus proliferation pattern in early embryonic mouse gut. PMID- 11102550 TI - The effects of intra-uterine growth retardation and postnatal undernutrition on onset of puberty in male and female rats. AB - The nutritional status, prenatally and early postnatally, plays a critical role in postnatal growth and development. Early malnutrition may change the original programming of organs, especially those in developmental phases, which can result in long-term changes in metabolism. The association between a low birth weight and the increased risk on type 2 diabetes, hypertension and cardiovascular disease is well known. In the present study we investigated whether intrauterine malnutrition or direct postnatal food restriction affects the onset of puberty in male and female rats. Intrauterine growth retardation (IUGR) was induced by uterine artery ligation on day 17 of gestation and postnatal food restriction (FR) by litter-enlargement to 20 pups per mother from day 2 after birth until weaning (24 d). Both models of malnutrition resulted in a persistent growth failure postnatally. The parameter of the onset of puberty was balano-preputial separation (BPS) in the male rat and vaginal opening (VO) in the female rat. In both male IUGR (n = 26) and FR (n = 20) rats, the age at BPS was significantly delayed, with 48.1 +/- 1.9 d (p < 0.0001) and 50.4 +/- 2.9 d (p < 0. 0001), respectively, compared with controls (n = 30) with 45.8 +/- 1.4 d. In female IUGR rats (n = 37) the age at VO was significantly delayed, with 37.4 +/- 2.7 d (p < 0.04) compared with 36.1 +/- 1.5 d in controls (n = 23), but not in female FR rats (n = 18) with 36.5 +/- 2.2 d. Weight at onset of puberty did not differ between male IUGR and control rats, 194.5 +/- 20.0 g and 201.7 +/- 16.8 g, respectively, but was significantly lower in male FR rats with a weight of 175.6 +/- 17.5 g (p < 0.0001). In female IUGR as well as in female FR rats, weight at onset of puberty was significantly lower compared with controls: weight in IUGR 106.1 +/- 13.1 g (p < 0.001), weight in FR 85.3 +/- 7.6 g (p < 0.0001) and weight in controls 116.9 +/- 9.3 g. We conclude that early malnutrition, during late gestation or direct postnatally, results in a delayed onset of puberty in IUGR and FR male rats and in IUGR female rats, but not in FR female rats. The onset of puberty in these growth retarded rats as well as in controls does not depend on the achievement of a certain, crucial weight. The perinatal period appears to be a "critical time period" for the maturational process of pubertal development. PMID- 11102551 TI - Placental insufficiency and fetal growth restriction lead to postnatal hypotension and altered postnatal growth in sheep. AB - Low birth weight has been associated with elevated arterial pressure in later life but mechanisms are unknown. Our aim was to determine the effects of low birth weight resulting from intrauterine growth restriction (IUGR) on fetal and postnatal arterial pressures and the potential roles of circulating cortisol and renin. We induced IUGR by umbilico-placental embolization (UPE) in fetal sheep from 120 d of gestation until birth (approximately 147 d); postnatal lambs (8 IUGR, 8 controls) were studied for 8 wk. Fetal and postnatal arterial pressures were measured and blood samples taken for measurement of gas tensions, cortisol concentrations and renin activity. In IUGR fetuses, mean arterial pressure (MAP) initially increased with UPE, but near term was not different to values in controls. IUGR lambs weighed 33% less than controls at birth and remained lighter than controls during the 8 postnatal weeks; their growth pattern was different to that of controls. IUGR lambs had lower MAP than controls, and this relative hypotension (-4 mm Hg) persisted throughout the 8 postnatal weeks. Covariate analysis showed that the relative hypotension of IUGR lambs could have resulted from their smaller size. Plasma cortisol concentrations were not different between IUGR and control animals before or after birth. Plasma renin activity was not different in postnatal IUGR lambs compared with controls. Thus, postnatal cortisol and renin levels were not consistent with the development of hypotension or hypertension. We conclude that late gestational IUGR in sheep leads to relative hypotension in the early postnatal period, probably a result of reduced body size. PMID- 11102552 TI - Effects of intrauterine and early postnatal growth restriction on hypothalamic somatostatin gene expression in the rat. AB - In the human, intrauterine growth retardation (IUGR) can result in persistent postnatal growth failure, which may be attributable, in part, to abnormal GH secretion. Whether putative alterations in GH secretion are the result of abnormalities intrinsic to the pituitary or reflect changes in the production of GH-releasing hormone or somatostatin (SS) is unknown. We tested the hypothesis that growth failure associated with IUGR or early postnatal food restriction (FR) is caused by a central defect in hypothalamic SS gene expression. Both models displayed persistent growth failure postnatally without any catch-up growth. We measured levels of SS mRNA levels in rats experimentally subjected to IUGR or FR. SS mRNA levels were measured by semiquantitative in situ hybridization throughout development. Levels of SS mRNA in the periventricular nucleus were significantly higher in both male and female IUGR rats in the juvenile and adult stages compared with matched controls (p < or = 0.05). FR was associated with higher SS mRNA levels only in neonatal female rats (p < or = 0.05). These results suggest that intrauterine malnutrition induces a persistent increase in the expression of SS mRNA in the periventricular nucleus, whereas early postnatal FR results in only a transient increase in SS gene expression. Because IGF-I levels were normal in juvenile IUGR and FR rats, central dysregulation of SS neurons does not appear to be the cause of early postnatal growth failure in either model. However, these observations are consistent with the hypothesis that nutritional stress at critical times during development can have persistent and potentially irreversible effects on organ function. PMID- 11102553 TI - Discordant amino acid profiles in monochorionic twins with twin-twin transfusion syndrome. AB - To test the hypothesis that discordant growth in monochorionic (MC) twins occurs at least in part due to disparity in placental amino acid transporter function, we measured plasma amino acid levels by HPLC in maternal and fetal blood samples collected at birth from gestational age matched twins with (n = 12) and without (n = 12) twin-twin transfusion syndrome (TTTS). In the donor twin, fetal plasma concentrations and feto-maternal ratios of five essential amino acids-valine (p < 0.001), leucine (p < 0.001), iso-leucine (p < 0.05), histidine (p < 0.001) and L arginine (p < 0. 001)-were lower than the recipient and non-TTTS twin pairs. Fetal concentrations of the nonessential amino acids taurine (p < 0.001), serine (p < 0.01), glycine (p < 0.001) and tyrosine (p < 0.05) were also markedly lower in the donor than the recipient and non-TTTS twin pairs. In contrast, the fetal alanine level in the donor twin was higher than the recipient (664 +/- 64 versus 396 +/- 23 microM; p < 0.001) and the non-TTTS twin pairs (p < 0.01). No such differences between amino acid profiles in non-TTTS MC twin pairs were found. Maternal plasma amino acid levels between TTTS and non-TTTS groups were comparable. This study provides the first evidence that certain amino acids in the donor twin of chronic TTTS differ significantly from those of the co-twin while others were comparable between twin pairs. These data, therefore, argue against inter-twin transfusion as the sole cause of growth restriction of the donor twin and suggests instead that impaired placental transport of amino acids may be a likely mechanism. PMID- 11102554 TI - A model to study antioxidant regulation of endotoxemia-modulated neonatal granulopoiesis and granulocyte apoptosis. AB - Neonates with septicemia tend to develop granulocytopenia, which may, in part, be due to septic mediators such as oxygen free radicals and tumor necrosis factor alpha (TNF-alpha). Granulocytopenia may be caused by a decrease in granulocyte growth and/or an increase in granulocyte destruction. In the present study, we investigated antioxidant regulation of endotoxin-modulated neonatal granulopoiesis and granulocyte apoptosis. Using human umbilical cord blood (HUCB), we found that simulating endotoxemia in vitro elicited significant superoxide production within a few minutes. Endotoxin exposure suppressed colony forming unit-granulocyte and monocyte formation in a dose-dependent fashion. Addition of antioxidants such as N-acetyl-cysteine could reverse the endotoxin suppression of colony-forming unit-granulocyte and monocyte formation (13 +/- 5 versus 75 +/- 5 colony-forming units/mL). Spontaneous in vitro granulocyte apoptosis in 6 h, as reflected by phosphatidylserine expression on the cell surface, was higher in granulocytes from HUCB than in those from adult blood (10.8 +/- 1.0% versus 5.6 +/- 1.2%). The addition of endotoxin or IL-8 to the cells in the in vitro model did not promote granulocyte apoptosis, but TNF-alpha, a major mediator of the effects of endotoxin, significantly induced granulocyte apoptosis in HUCB (control versus TNF-alpha: 8.9 +/- 1.2% versus 35.9 +/- 2.9%). Addition of the antioxidant N-acetyl-cysteine effectively blocked TNF-alpha induced granulocyte apoptosis as demonstrated by DNA fragmentation. Results from these studies indicate that oxygen radicals are directly involved in endotoxin suppression of granulopoiesis, and indirectly promote granulocyte apoptosis, presumably through TNF-alpha-mediated action. Thus, under certain conditions, modulation of oxygen radical production in the blood may benefit neonates with granulocytopenia. PMID- 11102555 TI - The use of low-EPA fish oil for long-chain polyunsaturated fatty acid supplementation of preterm infants. AB - Because docosahexaenoic acid (DHA) may be an essential nutrient for the visual and early cognitive development of preterm infants, DHA enrichment of preterm formulas has been recommended. This randomized trial was designed to study the n 6 and n-3 fatty acid status of healthy preterm infants fed a formula enriched with a low eicosapentaenoic-fish oil until 4 mo corrected age compared with that of infants fed a standard formula. A reference group of breast-fed infants was studied concurrently. The fatty acid content of red blood cell (RBC) phospholipid was assessed at enrollment, hospital discharge, expected term, and 3 and 6 mo postterm. The DHA content of RBC phospholipid was higher in infants fed the enriched versus the standard formula at hospital discharge, expected term, and 3 and 6 mo postterm. However, compared with infants fed the standard formula, infants fed the enriched formula had also higher RBC phospholipid eicosapentaenoic content (0.69 +/- 0.15% versus 0.25 +/- 0.12%, p < 0.001), and lower RBC phospholipid arachidonic acid content (15.1 +/- 0.93% versus 18.8 +/- 0.89%; p < 0.001). We conclude that supplementing preterm infants with low eicosapentaenoic fish oil is effective in improving DHA status, but results in worsening of n-6 fatty acid status. We speculate that preterm infants may require a dietary supply of arachidonic acid as well as DHA if the same fatty acid status as that of breast-fed infants is to be achieved. PMID- 11102556 TI - Expression of wild-type and mutant human ornithine transcarbamylase genes in Chinese hamster ovary cells and lack of dominant negative effect of R141Q and R40H mutants. AB - Chinese hamster ovary cultured cells were transformed to continuously express wild-type and two mutant ornithine transcarbamylase genes, R141Q and R40H. In addition, these cells were transfected to transiently express the same genes. The R141Q mutation abolishes the enzymatic activity, and the amount of "mature" protein present in transfected cells is equivalent to the wild type. The R40H mutation causes a reduction of enzymatic activity to approximately 26 to 35% of wild type concomitant with a significant reduction in the amount of protein present. Transfection with wild-type and mutant genes together in various proportions did not reveal dominant negative effects of the two mutations studied. This expression system can be used to examine the deleterious effect of private mutations or lack thereof in families with ornithine transcarbamylase deficiency as well as evaluate the potential dominant negative effects of gene delivery for treatment of ornithine transcarbamylase deficiency. PMID- 11102557 TI - Iodothyronine sulfotransferase activity in rat uterus during gestation. AB - In developing mammals, we and others demonstrated that sulfation is an important pathway in the metabolism of thyroid hormone, and there is significant fetal maternal transfer of sulfated iodothyronine. In the present study, we characterized a novel iodothyronine sulfotransferase (IST) in pregnant rat uterus. (125)I-labeled 3,3'-diiodothyronine (T(2)), T(3), rT(3), and T(4) were used as substrates with unlabeled 3'-phosphoadenosine-5'-phosphosulfate (PAPS) as the sulfate donor. Sulfated iodothyronine products were separated by Sephadex LH 20 column and further identified on reverse phase HPLC. We measured IST activity in pregnant rat uterus by incubating 1 microM substrate, 50 microM PAPS, and 50 microg cytosol protein, pH 7.2, 30 min at 37 degrees C. The results show that the substrate preference of the uterine IST activity is: T(2 )> rT(3 )> T(3)> T(4); the pH optimum is 6.0 for T(2). The K(m) and V:(max) (for gestational day 21 uterus) for T(2) are 0.62 microM and 3466 pmol/mg protein/h, respectively; for PAPS the values are 2.6 microM and 1523 pmol/mg protein/h, respectively. During pregnancy, the total activities exhibit a U-shaped curve with minimum activity at day 13 of gestation; while a thermostable activity increases significantly near term. In summary, there is significant uterine IST that varies during pregnancy. The role of this uterine sulfotransferase activities in regulating the bioavailability of thyroid hormone in the developing fetus remains to be elucidated. PMID- 11102558 TI - Progressive infantile neurodegeneration caused by 2-methyl-3-hydroxybutyryl-CoA dehydrogenase deficiency: a novel inborn error of branched-chain fatty acid and isoleucine metabolism. AB - We report a novel inborn error of metabolism identified in a child with an unusual neurodegenerative disease. The male patient was born at term and recovered well from a postnatal episode of metabolic decompensation and lactic acidosis. Psychomotor development in the first year of life was only moderately delayed. After 14 mo of age, there was progressive loss of mental and motor skills; at 2 years of age, he was severely retarded with marked restlessness, choreoathetoid movements, absence of directed hand movements, marked hypotonia and little reaction to external stimuli. Notable laboratory findings included marked elevations of urinary 2-methyl-3-hydroxybutyrate and tiglylglycine without elevation of 2-methylacetoacetate, mild elevations of lactate in CSF and blood, and a slightly abnormal acylcarnitine profile. These abnormalities became more apparent after isoleucine challenge. Enzyme studies showed absent activity of 2 methyl-3-hydroxybutyryl-CoA dehydrogenase (MHBD) in the mitochondrial oxidation of 2-methyl branched-chain fatty acids and isoleucine. Under dietary isoleucine restriction, neurologic symptoms stabilized over the next 7 months. PMID- 11102559 TI - Current problems with non-specific COX inhibitors. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used and effective treatments for pain and inflammation. They have a substantial toxicity profile with side effects mainly affecting the gastrointestinal tract, heart and kidneys. Although they comprise a chemically diverse group of drugs, NSAIDs are unified by a common mode of action the ability to inhibit the enzyme cyclo-oxygenase (COX). This also accounts for much of their toxicity. The enzyme exists in at least 2 isoforms. COX-1 generates prostaglandins with physiological functions, COX-2 is induced by inflammation and its physiologic functions are unclear at present. Conventional NSAIDs, like diclofenac, ibuprofen, and naproxen, are non-selective COX inhibitors, blocking the production of both physiologic and inflammatory prostaglandins. In this chapter, we describe the main predictable gastrointestinal, cardiac and renal toxicities that can be explained by such blockade and review the supporting clinical and epidemiological evidence. In the gastrointestinal tract, the side effects associated with conventional NSAIDs are both local and systemic, and include ulceration, bleeding, perforation, and obstruction. The upper gastrointestinal tract is more commonly affected than the lower. The cardiac and renal side effects are most likely to occur in patients with existing heart or kidney disease, where prostaglandins play an essential role in maintaining the vasoconstrictor/dilator balance necessary for homeostasis. The patients at highest risk of toxicity are the elderly, those with a prior history of ulceration or bleeding, and those with a history of cardiac disease. Among such patients, the decision to prescribe NSAIDs requires careful consideration of the potential benefits and harms. PMID- 11102560 TI - Clinical experience with specific COX-2 inhibitors in arthritis. AB - The common mechanism of action of aspirin and the chemically unrelated non steroidal anti-inflammatory drugs (NSAIDs) is the inhibition of prostaglandin (PG) production due to interference with the enzymatic activity of cyclooxygenase (COX). These agents have long been used as effective treatments for arthritis. The recognition that the inducible isoform COX-2 was associated with inflammation and arthritis led to the hypothesis that PGs produced by a COX-2-dependent pathway were responsible for the inflammation, pain, and tissue destruction. Since the constitutive COX-1 enzyme was identified as responsible for gastroprotection and inhibition of platelet function, the potential for compounds that were both effective and safer than NSAIDs led to rapid development of agents that specifically inhibit COX-2. These agents have now been tested and approved for use by the US Food and Drug Administration for patients with osteoarthritis and rheumatoid arthritis. They have been shown equally effective to comparitor NSAIDs. More importantly, there is a 3.5-fold reduction in the incidence of endoscopic gastroduodenal ulcerations and early data suggesting a similar reduction in clinically significant perforations, symptomatic ulcers, and bleeds. In patients with arthritis at risk for gastrointestinal complications of NSAIDs, specific inhibitors of COX-2 provide an effective and apparently safer form of anti-inflammatory agent. PMID- 11102561 TI - COX-2 and the kidneys. AB - The kidney is the second most frequent target of serious adverse effects of non steroidal antiinflammatory drugs (NSAIDs). The renal side effects of NSAIDs related to inhibition of cyclooxygenase (COX) comprise reduction in renal blood flow (RBF) and glomerular filtration rate (GFR), sodium/water retention, water intoxication and hyperkalemia. The discovery of two COX-isoenzymes, a constitutive COX-1, serving homeostatic prostanoid synthesis, and an inducible COX-2, responsible for proinflammatory prosta noid production, led to the development of new NSAIDs: Preferential and specific COX-2 inhibitors, promising minimal NSAID-typical toxicity with equivalent efficacy. However, we learned that there is no clear distinction in "physiologic" constitutive COX-1 and "inflammatory" inducible COX-2. This is particular true for the kidney of humans and other mammalians, where COX-2 was found constitutively in meaningful amounts. Animal experiments and clinical trials with preferential and specific COX-2 inhibitors revealed that COX-2 is the critical enzyme for sodium excretion, renin release and likely antagonism of antidiuretic hormone. Additionally, a significant role of COX-2 for nephro genesis is suggested. For renal hemodynamics the given evidence point to COX-1 as the predominant enzyme, but further investigations are required. In summary, the gain of renal safety by use of preferential or specific COX-2 inhibitors is small or negligible with respect to sodium retention, hyperkalemia and probably water intoxication. These drugs may be advantageous regarding renal perfusion, but presently the same precautions as for conventional NSAIDs must be used. PMID- 11102562 TI - Structure-based design of compounds inhibiting Grb2-SH2 mediated protein-protein interactions in signal transduction pathways. AB - Receptor protein tyrosine kinases are usually activated upon binding their growth factors, or other suitable ligands, to their extracellular domains. These activated receptors initiate cytoplasmic signalling cascades which, when aberrant, can result in different disease states, such as oncogenic transformation. Many receptor protein tyrosine kinases use Src homology 2 domains (SH2) to couple growth factor activation with intracellular signalling pathways to mediate cell control and other biological events. The characterization of the components involved in these signal transduction pathways has resulted in the identification of new attractive targets for therapeutic intervention. Such is the case for the protein-protein interactions involving the SH2 domain of growth factor receptor bound protein 2 (Grb2). Agents that specifically disrupt Grb2-SH2 binding interactions involved in aberrant signalling could potentially shut down these oncogenic pathways and thus block human malignancies. This paper reviews the structural characteristics of the Grb2-SH2 domain and the approaches which have been used to identify antagonists of the Grb2-SH2 domain. Examples have been selected from our own research to illustrate how the unique structural features of the ligand-bound Grb2-SH2 have been exploited to design potent and selective Grb2-SH2 antagonists. PMID- 11102563 TI - From a classic approach in cancer chemotherapy towards differentiation therapy: acyclic and cyclic seven-membered 5-fluorouracil O,N-acetals. AB - Novel derivatives of 5-fluorouracil (5-FU) possessing a broader spectrum of antitumor activity and fewer toxic side effects than 5-FU have been sought. Herein, we report three different types of 5-FU O,N-acetals: a) a novel class of 5-fluorouracil-containing acyclonucleosides. The antitumor activities of such compounds were assessed against HEp human cells showing that (RS)-1-?[3-(2 hydroxyethoxy)-1-cyclopentoxy]propyl?-5-fluorouracil 3c is 4-fold more active than 5-FU. (RS)-1-?[3-(2-hydroxyethoxy)-1-isopropoxy]propyl?-5-fluorouracil 3b has important potential advantages over 5-FU because of its lower toxicity and its ability to induce myogenic differentiation in rhabdomyosarcoma cells. Our results suggest that this drug may be useful for differentiation therapy in this type of tumor; b) within the cyclic prodrugs of 5-FU, a series of new ring expanded isosteres (1,4-oxaheteroepanes) of Ftorafur were synthesized. The level of diastereoselectivity in the preparation of cis and trans 1-(3-chloromethyl) 1,4-dioxepan-5-yl)-5-fluorouracil, although modest, suggests a potentially general approach for controlling the stereochemistry of this unexplored class of reactions involving the preparation of 5-FU seven-membered O,N-acetals; c) new 5 FU acyclic analogs containing two 5-FU moieties at both ends of the molecules with a linker having two amide bonds have been designed and synthesized. These bis(5-FU-O,N-acetals) show interesting antineoplastic activities against the HT 29 cell line. PMID- 11102564 TI - Antitumor properties of podophyllotoxin and related compounds. AB - The lignan family of natural products includes compounds with important antineoplastic and antiviral properties such as podophyllotoxin and two of their semisynthetic derivatives, etoposide and teniposide. The latter are included in a wide variety of cancer chemotherapy protocols. Due to these biological activities, lignans, and especially cyclolignans, have been the objective of numerous studies focused to prepare better and safer anticancer drugs. The mechanism by which podophyllotoxin blocks cell division is related to its inhibition of microtubule assembly in the mitotic apparatus. However, etoposide and teniposide were shown not to be inhibitors of microtubule assembly which suggested that their antitumor properties were due to another mechanism of action, via their interaction with DNA and inhibition of DNA topoisomerase II. Other podophyllotoxin derivatives has also been reported which retained or even improved the cytotoxic activity, but these were weak inhibitors of topoisomerase II in vitro; the data revealed that such analogs exhibit a different, as yet unknown, mechanism of action. The main deficiency of these compounds is their cytotoxicity for normal cells and hence side effects derived from their lack of selectivity against tumoral cells. In this regard it is necessary to investigate and prepare new more potent and less toxic analogs, that is, with better therapeutic indices. It is well accepted from structure-activity studies in this field that the trans-lactones are more potent as antineoplastics than the cis lactones. Not only the configuration of the D ring is an important factor for high cytotoxic activity, but also a quasi-axial arrangement of the E ring is necessary. On this basis, studies on lignans have been addressed to modify the lactone moiety and prepare analogs with heteroatoms at different positions of the cyclolignan skeleton. Our group has been working during the last few years on chemical transformations of podophyllotoxin and analogs and we have prepared a large number of cyclolignan derivatives some of which display potent antiviral, immunosuppressive and cytotoxic activities. We have reported several new cytotoxic agents with nitrogen atoms at C-7 or C-9 or at both C-7 and C-9: imine derivatives, oxime derivatives, pyrazoline-, pyrazo- and isoxazoline-fused cyclolignans. At present, we are preparing mainly new compounds by modifications of the A and E cyclolignan-rings. They are being tested on cultures of different tumoral cell lines (P-388 murine leukemia, A-549 human lung carcinoma, HT-29 human colon carcinoma and MEL-28 human melanoma) and some of them have shown an interesting and selective cytotoxicity. PMID- 11102565 TI - Understanding and exploiting nature's chemical arsenal: the past, present and future of calicheamicin research. AB - The enediyne antitumor antibiotics are appreciated for their novel molecular architecture, their remarkable biological activity and their fascinating mode of action and many have spawned considerable interest as anticancer agents in the pharmaceutical industry. Of equal importance to these astonishing properties, the enediynes also offer a distinct opportunity to study the unparalleled biosyntheses of their unique molecular scaffolds and what promises to be unprecedented modes of self-resistance to highly reactive natural products. Elucidation of these aspects should unveil novel mechanistic enzymology, and may provide access to the rational biosynthetic modification of enediyne structure for new drug leads, the construction of enediyne overproducing strains and eventually lead to an enediyne combinatorial biosynthesis program. This article strives to compile and present the critical research discoveries relevant to the clinically most promising enediyne, calicheamicin, from a historical perspective. Recent progress, particularly in the areas of biosynthesis, self-resistance, bio engineering analogs and clinical studies are also highlighted. PMID- 11102566 TI - Mucins in the diagnosis and therapy of pancreatic cancer. AB - Mucins are large glycoproteins that form a protective layer along the lumens of the organs of the gastrointestinal and reproductive tracts. Frequently in tumors of the pancreas there are changes in the structure of mucin carbohydrates and/or levels of apomucin types. Originally mucins were of interest clinically because diagnostic tests could be based on their levels in circulation. More recently mucin directed monoclonal antibodies have been used to target tumors with cytotoxic agents. There is now a considerable literature on the development of mucin-based vaccines. Both monoclonal antibodies and vaccines could be powerful tools to specifically target tumor cells in distant metastases. Gene therapy based upon the MUC1 gene promoter is being investigated to target therapeutic genes to MUC1 expressing cells. The carbohydrates of mucins on the surface of tumor cells have been reported to inhibit cells of the immune system. These carbohydrates also act as ligands during the process of tumor cell metastasis. Another approach to therapy is to block interactions between the ligands and their receptors. PMID- 11102567 TI - Fibroblast growth factors and their inhibitors. AB - Fibroblast growth factors (FGFs) are members of a family of polypeptides synthesized by a variety of cell types during the processes of embryonic development and in adult tissues. FGFs have been detected in normal and malignant cells and show a biological profile that includes mitogenic and angiogenic activity with a consequent crucial role in cell differentiation and development. To activate signal transduction pathways, FGFs use a dual receptor system based on tyrosine kinases and heparan sulfate (HS) proteoglycans. Based on these considerations, a variety of inhibitors able to block the interactions between FGFs and their receptors have been designed and investigated for their biological properties related to antiangiogenesis and antitumor activity. In this paper, in addition to an extensive description of the FGF family members, we report several compounds acting as FGF inhibitors by direct interaction with the growth factors. Suramin and other diverse polyanionic polysulfated and polysulfonated compounds are described, with a particular focus on suradistas. For this class of molecules, by means of molecular modeling procedures, a binding model to FGF-2 has been proposed and the structure-activity relationships of suradistas have been analyzed on the basis of the computational model described. PMID- 11102568 TI - delta-, but not mu- and kappa-, opioid receptor activation protects neocortical neurons from glutamate-induced excitotoxic injury. AB - Recent observations from our laboratory have led us to hypothesize that delta opioid receptors may play a role in neuronal protection against hypoxic/ischemic or glutamate excitotocity. To test our hypothesis in this work, we used two independent methods, i.e., "same field quantification" of morphologic criteria and a biochemical assay of lactate dehydrogenase (LDH) release (an index of cellular injury). We used neuronal cultures from rat neocortex and studied whether (1) glutamate induces neuronal injury as a function of age and (2) activation of opioid receptors (delta, mu and kappa subtypes) protects neurons from glutamate-induced injury. Our results show that glutamate induced neuronal injury and cell death and this was dependent on glutamate concentration, exposure period and days in culture. At 4 days, glutamate (up to 10 mM, 4 h-exposure) did not cause apparent injury. After 8-10 days in culture, neurons exposed to a much lower dose of glutamate (100 microM, 4 h) showed substantial neuronal injury as assessed by morphologic criteria (>65%, n=23, P<0.01) and LDH release (n=16, P<0. 001). Activation of delta-opioid receptors with 10 microM DADLE reduced glutamate induced injury by almost half as assessed by the same criteria (morphologic criteria, n=21, P<0.01; LDH release, n=16, P<0.01). Naltrindole (10 microM), a delta-opioid receptor antagonist, completely blocked the DADLE protective effect. Administration of mu- and kappa-opioid receptor agonists (DAMGO and U50488H respectively, 5-10 microM) did not induce appreciable neuroprotection. Also, mu- or kappa-opioid receptor antagonists had no appreciable effect on the glutamate induced injury. This study demonstrates that activation of neuronal delta-opioid receptors, but not mu- and kappa-opioid receptors, protect neocortical neurons from glutamate excitotoxicity. PMID- 11102569 TI - Temporary inactivation of the retrorubral fields decreases the rewarding effect of medial forebrain bundle stimulation. AB - Prior studies indicate that lesioning the retrorubral fields (RRF) decreases the rewarding effect of medial forebrain bundle (MFB) stimulation, although these studies did not make the RRF their primary target. This study directly investigates the role of the RRF in MFB self-stimulation using transient lidocaine-induced inactivation of target tissue rather than permanent lesioning. In 18 rats with MFB stimulation electrodes, inactivation of the RRF via 0. 5 and 1.0 microl of 4% lidocaine produced immediate, substantial upward shifts in the frequency required to maintain half-maximal self-stimulation response rates whereas injecting comparable volumes of saline did not. Bilateral inactivation was particularly effective, especially at medium and high stimulation currents, although unilateral inactivation ipsilateral to the stimulation site was also effective. Contralateral inactivation alone did not substantially change the stimulation's reward value, although contralateral inactivation appeared to contribute to the effectiveness of bilateral inactivation. The frequency required to maintain half-maximal responding returned to baseline levels by 15-20 min after lidocaine infusion. In seven rats whose infusion sites were not in the RRF, lidocaine inactivation did not consistently degrade the stimulation's reward value. These results indicate that some neural elements located in the RRF contribute to the rewarding effect of MFB stimulation. Possible roles for these elements in the anatomical substrate for MFB self-stimulation are discussed. PMID- 11102570 TI - Neuronal degeneration in the limbic system of weanling rats exposed to saline, hyperthermia or d-amphetamine. AB - Neuronal degeneration was detected in the tenia tecta and other regions of the anterior limbic system of male weanling rats 3 days after four doses of 5 mg/kg d amphetamine (4 x 5 mg/kg AMPH) when seizures occurred during AMPH exposure. Neurodegeneration in the parietal cortex, loss of tyrosine hydroxylase immunoreactivity in the caudate-putamen (CPu) and decreases in CPu tissue dopamine levels in weanlings was much less than those previously observed in adults. The neurotoxicity seen in the parietal cortex and CPu of the weanlings was much less than previously seen in adults even though severe hyperthermia and the behavior of retrograde propulsion occurred during AMPH exposure. Neurodegeneration was not detected in any of the previously mentioned brain regions in controls and weanlings made hyperthermic by a warm environment. However, signs of spontaneous neurodegeneration were seen in the posterior piriform cortex (Pir), posteriolateral cortical amygdaloid nucleus (PLCo), and the amygdalopiriform transition area (APir) of control weanlings. The doses of AMPH and the degree of hyperthermia necessary to induce seizures were substantially lower in weanlings compared to those previously observed in adult rats. Further studies will be necessary to determine if the susceptibility of weanlings to AMPH-induced seizures is related to or dependent on the same processes involved in producing degeneration in the posterior limbic system of saline controls. PMID- 11102571 TI - Expression of interleukin-6 and its receptor in the sciatic nerve and cultured Schwann cells: relation to 18-kD fibroblast growth factor-2. AB - Expression of interleukin-6 (IL-6) and fibroblast growth factor-2 (FGF-2) in Schwann cells is modulated by external stimuli. To study possible interactions of both factors we have analyzed mutual effects of exogenous IL-6 and FGF-2 on the expression of each other and the corresponding receptor (R) molecules IL-6R and FGFR1 after peripheral nerve lesion in vivo and in vitro using cultured Schwann cells. Using rat Schwann cells we found that IL-6 did not exert any effects on the expression of FGF-2 and FGF receptor type 1 (R1) whereas exogenously applied 18-kD FGF-2 strongly increased the expression of the mRNAs of IL-6 and its receptor. In addition, immortalized Schwann cells over-expressing the 18-kD FGF-2 isoform showed elevated levels of IL-6 and IL-6R whereas immortalized Schwann cells over-expressing the high-molecular-weight isoforms (21 kD and 23 kD) displayed unaltered IL-6 and IL-6R expression levels. According to in situ hybridization studies of intact and crushed sciatic nerves in vivo, Schwann cells seems to be the main source of IL-6 and IL-6R. Following sciatic nerve crush, the FGF-2 and the IL-6 system are upregulated after the first hours. Furthermore, we showed that the early increase of the FGF-2 protein is mainly confined to the 18 kD isoform. These results are consistent with the idea of a functional coupling of FGF-2 and the IL-6 system in the early reaction of Schwann cells to nerve injury. PMID- 11102572 TI - Corticosterone inhibits generation of long-term potentiation in rat hippocampal slice: involvement of brain-derived neurotrophic factor. AB - In the present study, the effect of corticosterone (CORT) on the generation of long-term potentiation (LTP) and its underlying mechanism involving neurotrophin gene expression in CA1 synapses of rat hippocampal slice were examined. Our experimental results showed incubation of hippocampal slice with CORT for 3 h had no effect on either the slope or amplitude of excitatory postsynaptic potentials (EPSP) evoked in hippocampal CA1 pyramidal dentrites, indicating no marked change in basal synaptic transmission. However, when tetanic stimulation (100 pulses, 100 Hz) was delivered to the Schaffer collateral pathway, CORT application significantly attenuated the tetanus-induced increases of both EPSP slope and amplitude, demonstrating an inhibitory effect of CORT on LTP generation. In addition, CORT treatment significantly reduced both slope and amplitude ratios of the second evoked EPSP to the first one when paired-pulse facilitation (PPF) was established at different interpulse intervals from 20 to 40 ms, suggesting that a presynaptic mechanism may be involved in CORT-induced hippocampal synaptic plasticity. Reverse-transcription polymerase chain reaction (RT-PCR) analysis showed that CORT-treated hippocampal CA1 cells underwent a significant decrease in the expression of mRNA for nerve growth factor-beta (NGF-beta) and brain derived neurotrophic factor (BDNF), but not for neurotrophin-3 (NT-3) compared with those in control. Moreover, BDNF co-applied with CORT significantly antagonized CORT-induced deficit in PPF. Taken together, the present results suggest that CORT-induced inhibition of LTP may be, at least to some extent, mediated by a presynaptic mechanism and decrease in the BDNF expression in rat hippocampal CA1 cells induced by CORT may partially account for this presynaptic mechanism. PMID- 11102573 TI - Sodium valproate alters GnRH-GABA interactions during development in seizure prone mice. AB - During reproductive maturation, characteristic changes occur in the morphology of the gonadotropin releasing hormone (GnRH) cell population within the hypothalamus. In the early stages of development, GnRH neurons are bipolar cells; however, just before pubertal onset, the majority of these neurons transform into unipolar cells. Our laboratory has reported that valproic acid (VPA), an antiepileptic medication that has previously been shown to slow the velocity of pubertal development in both humans and seizure-prone mice, is capable of delaying the normal process of GnRH morphological differentiation. As VPA is primarily believed to act via a GABAergic mechanism, the present study investigated potential influences of VPA on GnRH-GABA interactions within the medial preoptic area (mPOA) across pubertal development (experiment 1), as well as in adult animals (experiment 2). The results from experiment 1 revealed the expected drug effects on GnRH cell morphology. For VPA animals, there was a greater percentage of bipolar neurons at every time period except for the 24-day sample. Additionally, VPA animals had greater numbers of bipolar and unipolar GnRH neurons with GABA associations across all ages. However, experiment 2 showed a lack of drug effects on GnRH-GABA interactions in adulthood. These results suggest that VPA may delay GnRH cell morphological maturation by altering the density of GABAergic inputs to GnRH neurons. These inputs may normally play a role in timing the activation of the GnRH pulse generator. However, any neuroendocrine effects of VPA in adulthood are most likely due to the actions of VPA at another level of the hypothalamic-pituitary-gonadal axis. PMID- 11102574 TI - Degree of immediate early gene induction in striatum by eticlopride determines sensitivity to N-methyl-D-aspartate receptor blockade. AB - Cortical afferents excite striatal efferent neurons through activation of N methyl-D-aspartate (NMDA) receptors, which can be modulated by D2 dopamine receptors. It is suggested that activation of PKA by D2 receptor blockade leads to NMDA receptor phosphorylation in the dendrites or phosphorylation of transcription factors in the nucleus. Thus, the levels and cellular localization of activated PKA may determine if D2 antagonist-mediated gene expression is dependent on NMDA receptor activation. We have previously demonstrated that NMDA receptor antagonists block gene expression induced by a high dose of eticlopride in medial and central but not lateral striatum. Here, we examined the effects of NMDA receptor antagonists on striatal gene expression after administration of a low dose of eticlopride. The results showed that NMDA receptor antagonists blocked gene induction by eticlopride throughout striatum. Less PKA activation by the low dose of eticlopride might explain why the expression was more sensitive in the lateral striatum to NMDA receptor blockade than in our previous study. To increase levels of PKA activation to the extent that NMDA receptor blockade would have less effect on eticlopride-mediated gene induction in all regions of striatum, we administered the phosphodiesterase inhibitor IBMX to animals treated with eticlopride. The combined administration of IBMX and eticlopride induced gene expression that was only partially attenuated (c-fos) or unaffected (zif268) by NMDA receptor blockade. These data support the suggestion that the degree of second messenger activation by D2 receptor blockade determines whether D2 dopamine receptor antagonist-mediated gene expression is dependent on NMDA receptor activation. PMID- 11102575 TI - Diffusion- and perfusion-weighted magnetic resonance imaging of focal cerebral ischemia and cortical spreading depression under conditions of mild hypothermia. AB - In a model of experimental stroke, we characterize the effects of mild hypothermia, an effective neuroprotectant, on fluid shifts, cerebral perfusion and spreading depression (SD) using diffusion- (DWI) and perfusion-weighted MRI (PWI). Twenty-two rats underwent 2 h of middle cerebral artery (MCA) occlusion and were either kept normothermic or rendered mildly hypothermic shortly after MCA occlusion for 2 h. DWI images were obtained 0.5, 2 and 24 h after MCA occlusion, and maps of the apparent diffusion coefficient (ADC) were generated. SD-like transient ADC decreases were also detected using DWI in animals subjected to topical KCl application (n=4) and ischemia (n=6). Mild hypothermia significantly inhibited DWI lesion growth early after the onset of ischemia as well as 24 h later, and improved recovery of striatal ADC by 24 h. Mild hypothermia prolonged SD-like ADC transients and further decreased the ADC following KCl application and immediately after MCA occlusion. Cerebral perfusion, however, was not affected by temperature changes. We conclude that mild hypothermia is neuroprotective and suppresses infarct growth early after the onset of ischemia, with better ADC recovery. The ADC decrease during SD was greater during mild hypothermia, and suggests that the source of the ADC is more complex than previously believed. PMID- 11102576 TI - Manganese induces neurite outgrowth in PC12 cells via upregulation of alpha(v) integrins. AB - Previous studies have demonstrated that the divalent cation manganese (Mn) causes PC12 cells to form neurites in the absence of NGF. Since divalent cations modulate the binding affinity and specificity of integrins, and integrin function affects neurite outgrowth, we tested the hypothesis that Mn induces neurite outgrowth through an integrin-dependent signaling pathway. Our studies support this hypothesis. Function-blocking antisera specific for beta(1) integrins block the neurite-promoting activity of Mn by 90-95%. Bioassays and biochemical studies with antisera specific for the alpha(v), alpha(5), or alpha(8) integrin subunit suggest that the alpha(v)beta(1) heterodimer is one of the principal beta(1) integrins mediating the response of PC12 cells to Mn. This is corroborated by studies in which Mn failed to induce neurite outgrowth in a clone of PC12 cells that does not express alpha(v) at levels detectable by immunoprecipitation or immunocytochemistry. SDS-PAGE analysis of biotinylated surface proteins immunoprecipitated from Mn-responsive PC12 cells, as well as confocal laser microscopy of PC12 immunostained for surface alpha(v) indicate that Mn increases the surface expression of alpha(v) integrins. This increase appears to be due in part to synthesis of alpha(v) since specific inhibitors of RNA and protein synthesis block the neurite-promoting activity of Mn. These data indicate that Mn induces neurite outgrowth in PC12 cells by upregulating alpha(v) integrins, suggesting that Mn potentially represents an additional mechanism for regulating the rate and direction of neurite outgrowth during development and regeneration. PMID- 11102577 TI - Activation of purinergic receptors by ATP inhibits secretion in bovine adrenal chromaffin cells. AB - Autoinhibition is a common mechanism observed in neurons to regulate neurotransmission. Released neurotransmitter interacts with presynaptic autoreceptors to inhibit subsequent release. The requisite elements for autoinhibition are present in chromaffin cells: secretory granules contain millimolar levels of ATP which is coreleased with catecholamines upon stimulation and the cells express purinergic receptors. We were interested to determine whether autoinhibition produced by ATP binding to purinergic receptors plays an important role in catecholamine release from chromaffin cells. In these studies, short depolarizations were used to elicit transmitter release measured by membrane capacitance. We find that stimulation of chromaffin cells results in the release of endogenous ATP which may suppress Ca(2+) channel currents and secretion. In the presence of a maximal concentration of ATP, both the amount of secretion and the maximal rate of release are about half that observed in the absence of ATP. ATP-mediated inhibition of secretion was blocked by Reactive Blue 2 suggesting the involvement of P(2Y) purinergic receptors. Prepulses to positive potentials that relieve the Ca(2+) channel block largely relieve the inhibition of secretion. Furthermore, when secretion is plotted as a function of Ca(2+) influx there is no apparent change in the relationship between control cells and those stimulated in the presence of ATP and prepulses. These results suggest that ATP diminishes secretion by inhibiting Ca(2+) influx into the cells. Our results indicate that feedback inhibition by ATP, mediated primarily by Ca(2+) channels, may be an important regulator of catecholamine release in chromaffin cells. PMID- 11102578 TI - Distinct signaling pathways involved in multiple effects of basic fibroblast growth factor on cultured rat hippocampal neurons. AB - We investigated possible involvement of voltage-dependent Ca(2+) channels (VDCCs) and several intracellular signaling mechanisms in multiple actions of basic fibroblast growth factor (bFGF), such as survival promotion, induction of calbindin D(28k) expression as well as acceleration of neuritic branch formation of cultured rat hippocampal neurons. Immunocytochemical staining with anti-gamma aminobutyric acid (GABA) antibody showed that the promotion of neuron survival by bFGF in high cell-density cultures were exerted exclusively on GABA-negative neurons. Nicardipine (5 microM) attenuated the effect of bFGF on neuronal survival and formation of neurite branches, suggesting that the activity of L type VDCCs is required for these effects. In contrast, stimulation of calbindin expression by bFGF was not attenuated by nicardipine. A phospholipase C inhibitor U73122 (1 microM) prevented the effect of bFGF on neurite branch formation, but not on neuronal survival or calbindin expression. On the other hand, chronic application of phorbol-12-myristate-13-acetate (1 microM) inhibited the effect of bFGF on neuronal survival, without inhibiting the other bFGF actions. Forskolin (100 microM) attenuated the effect of bFGF on neuronal survival and neurite branch formation, indicating that cyclic AMP plays negative regulatory roles in these actions of bFGF. Taken together, these results suggest that multiple biological actions of bFGF on hippocampal neurons are exerted through, and modulated by, distinct signaling pathways. PMID- 11102579 TI - Structural alterations of tight junctions are associated with loss of polarity in stroke-prone spontaneously hypertensive rat blood-brain barrier endothelial cells. AB - The mechanisms leading to stroke in stroke-prone spontaneously hypertensive rats (SHRSP) are not well understood. We tested the hypothesis that the endothelial tight junctions of the blood-brain barrier are altered in SHRSP prior to stroke. We investigated tight junctions in 13-week-old SHRSP, spontaneously hypertensive stroke-resistant rats (SHR) and age-matched Wistar-Kyoto rats (WKY) by electron microscopy and immunocytochemistry. Ultrathin sections showed no difference in junction structure of cerebral capillaries from SHRSP, SHR and WKY, respectively. However, using freeze-fracturing, we observed that the blood-brain barrier specific distribution of tight junction particles between P- and E-face in WKY (58.7+/-3.6%, P-face; 41.2+/-5.59%, E-face) and SHR (53.2+/-19. 3%, P-face; 55.6+/-13.25%, E-face) was changed to an 89.4+/-9.9% predominant E-face association in cerebral capillaries from SHRSP. However, the expression of the tight junction molecules ZO-1, occludin, claudin-1 and claudin-5 was not changed in capillaries of SHRSP. Permeability of brain capillaries from SHRSP was not different compared to SHR and WKY using lanthanum nitrate as a tracer. In contrast, analysis of endothelial cell polarity by distribution of the glucose-1 transporter (Glut-1) revealed that its abluminal:luminal ratio was reduced from 4:1 in SHR and WKY to 1:1 in endothelial cells of cerebral capillaries of SHRSP. In summary, we demonstrate that early changes exist in cerebral capillaries from a genetic model of hypertension-associated stroke. We suggest that a disturbed fence function of the tight junctions in SHRSP blood-brain barrier endothelial cells may lead to subtle changes in polarity. These changes may contribute to the pathogenesis of stroke. PMID- 11102580 TI - Changes in electrophysiological properties of cat hypoglossal motoneurons during carbachol-induced motor inhibition. AB - The control of hypoglossal motoneurons during sleep is important from a basic science perspective as well as to understand the bases for pharyngeal occlusion which results in the obstructive sleep apnea syndrome. In the present work, we used intracellular recording techniques to determine changes in membrane properties in adult cats in which atonia was produced by the injection of carbachol into the pontine tegmentum (AS-carbachol). During AS-carbachol, 86% of the recorded hypoglossal motoneurons were found to be postsynaptically inhibited on the basis of analyses of their electrical properties; the electrical properties of the remaining 14% were similar to motoneurons recorded during control conditions. Those cells that exhibited changes in their electrical properties during AS-carbachol also displayed large-amplitude inhibitory synaptic potentials. Following sciatic nerve stimulation, hypoglossal motoneurons which responded with a depolarizing potential during control conditions exhibited a hyperpolarizing potential during AS-carbachol. Both spontaneous and evoked inhibitory potentials recorded during AS-carbachol were comparable to those that have been previously observed in trigeminal and spinal cord motoneurons under similar experimental conditions as well as during naturally occurring active sleep. Calculations based on modeling the changes that we found in input resistance and membrane time constant with a three-compartment neuron model suggest that shunts are present in all three compartments of the hypoglossal motoneuron model. Taken together, these data indicate that postsynaptic inhibitory drives are widely distributed on the soma-dendritic tree of hypoglossal motoneurons during AS-carbachol. These postsynaptic inhibitory actions are likely to be involved in the pathophysiology of obstructive sleep apnea. PMID- 11102581 TI - Rescue of ischemic brain injury by adenoviral gene transfer of glial cell line derived neurotrophic factor after transient global ischemia in gerbils. AB - Glial cell line-derived neurotrophic factor (GDNF), a member of the transforming growth factor (TGF)-beta superfamily, is one of the most potent neurotrophic factors and promotes survival of many populations of cells. We examined neuroprotective effect of an adenoviral vector encoding glial cell line-derived neurotrophic factor (AxCAhGDNF) on the transient global ischemia. Gerbils received administration of AxCAhGDNF or an adenoviral vector encoding bacterial beta-galactosidase gene (AxCALacZ) through the lateral ventricle. Two days later, occluding bilateral common carotid arteries for 5 min using aneurysm clips produced the transient global forebrain ischemia. Animals showed intense immunolabeling for GDNF in ependymal cells on 2, 4 and 7 days after the operation. The exogenous gene transducted by adenovirus in the same cells was detected by in situ hybridization. The treatment with AxCAhGDNF significantly prevented the loss of hippocampal CA1 pyramidal neurons 2 to 7 days after the operation, as compared to AxCALacZ treatment. Also terminal deoxynucleotidyl transferase-mediated dUTP-biotin in situ nick end labeling (TUNEL) staining was markedly reduced in the case with AxCAhGDNF treatment at 7 days after the operation. These results indicated that the adenovirus-mediated gene transfer of GDNF might prevent the delayed neuronal death of stroke and other disorders of the cerebral vasculature. PMID- 11102582 TI - Increased striatal dopamine turnover following acute administration of rotenone to mice. AB - Because of the potential role of mitochondrial dysfunction in nigrostriatal degeneration in Parkinson's disease, the effects of rotenone (an inhibitor of mitochondrial NADH dehydrogenase and a naturally occurring toxicant) on the levels of striatal dopamine (DA) and DA metabolites were evaluated after acute and subchronic administration to mice. Systemic acute treatment with relatively high doses of rotenone did not affect DA concentration, but caused a significant increase in both DA metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). DOPAC and HVA changes were measured at 1 day and were reversed within 1 week, paralleling the time course of rotenone-induced increase in striatal lactate levels. Subchronic administration with a relatively mild dose of rotenone did not significantly alter the striatal levels of DA and DOPAC, while it slightly reduced HVA concentration. No neurochemical signs of dopaminergic damage were seen when mice were co-exposed to rotenone and diethyldithiocarbamate, a compound known to enhance nigrostriatal injury caused by the neurotoxicant 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP). Also, rotenone did not cause additional injury to animals previously lesioned by MPTP. Taken together, data indicate that rotenone is not capable of causing overt dopaminergic toxicity under the testing paradigms used in this study. Rather, an increase in DA turnover, as indicated by a higher (DOPAC+HVA)/DA ratio, seems to be associated to rotenone-induced striatal energy impairment. PMID- 11102583 TI - Action of carbamazepine on epileptiform activity of the verartidine model in CA1 neurons. AB - The veratridine epileptiform model was utilized to assess the antiepileptic effect of Carbamazepine (CBZ) in rat hippocampal CA1 pyramidal neurons using conventional intracellular recording techniques. In the veratridine model, where brain slices are treated with veratridine (0.3 microM), a single intracellular stimulus evokes epileptiform bursting. Additionally, spontaneous epileptiform activity commonly appears on prolonged exposure to veratridine in this model. In this model, therapeutic (7-15 microM) and high (50 microM) concentrations of CBZ inhibited the evoked and spontaneous epileptiform bursting in a concentration- and voltage-dependent manner. At all concentrations tested, CBZ produced inhibition of epileptiform activity without affecting the membrane resting potential or input resistance. However, at 50 microM, the drug increased the firing threshold of neurons. These results confirm the suitability of this model for testing sodium channel-dependent antiepileptic agents. PMID- 11102584 TI - 7-nitroindazole reduces cerebral blood flow following chronic nitric oxide synthase inhibition. AB - Blood flow and glucose utilization were measured in rat brain after chronic L NAME treatment followed by acute 7-nitroindazole. Following chronic L-NAME, blood flow was not significantly different from control. Treatment with acute 7 nitroindazole reduced blood flow to the same extent in both chronic saline and L NAME groups. Glucose utilization was unaffected. These results suggest that residual NOS activity in brain is sufficient to provide tonic, NO-dependent cerebrovascular dilator tone. PMID- 11102585 TI - The pineal gland is not essential for circadian expression of rat period homologue (rper2) mRNA in the suprachiasmatic nucleus and peripheral tissues. AB - To investigate the functional involvement of the pineal gland in circadian expression of the rat period homolog gene (rPer2) in the suprachiasmatic nucleus (SCN) and peripheral tissues, we performed Northern blot analysis in tissues from pinealectomized rats. The ectomy did not have any significant effects on rPer2 mRNA expression patterns both in a daily light-dark condition and in a constant darkness. These results suggest that the rhythmic secretion of pineal melatonin is not essential for the circadian expression of clock genes in the SCN and other peripheral tissues of rats. PMID- 11102586 TI - Gene expression in the brain across the sleep-waking cycle. AB - Sleep and waking differ significantly in terms of behavior, metabolism, and neuronal activity. Recent evidence indicates that sleep and waking also differ with respect to the expression of certain genes. To systematically investigate such changes, we used mRNA differential display and cDNA microarrays to screen approximately 10000 transcripts expressed in the cerebral cortex of rats after 8 h of sleep, spontaneous waking, or sleep deprivation. We found that 44 genes had higher mRNA levels after waking and/or sleep deprivation relative to sleep, while 10 were upregulated after sleep. Known genes that were upregulated in waking and sleep deprivation can be grouped into the following categories: immediate early genes/transcription factors (Arc, CHOP, IER5, NGFI-A, NGFI-B, N-Ras, Stat3), genes related to energy metabolism (glucose type I transporter Glut1, Vgf), growth factors/adhesion molecules (BDNF, TrkB, F3 adhesion molecule), chaperones/heat shock proteins (BiP, ERP72, GRP75, HSP60, HSP70), vesicle- and synapse-related genes (chromogranin C, synaptotagmin IV), neurotransmitter/hormone receptors (adrenergic receptor alpha(1A) and beta(2), GABA(A) receptor beta(3), glutamate NMDA receptor 2A, glutamate AMPA receptor GluR2 and GluR3, nicotinic acetylcholine receptor beta(2), thyroid hormone receptor TRbeta), neurotransmitter transporters (glutamate/aspartate transporter GLAST, Na(+)/Cl(-) transporter NTT4/Rxt1), enzymes (aryl sulfotransferase, c-jun N-terminal kinase 1, serum/glucocorticoid-induced serine/threonine kinase), and a miscellaneous group (calmodulin, cyclin D2, LMO-4, metallothionein 3). Several other genes that were upregulated in waking and all the genes upregulated in sleep, with the exception of the one coding for membrane protein E25, did not match any known sequence. Thus, significant changes in gene expression occur across behavioral states, which are likely to affect basic cellular functions such as RNA and protein synthesis, neural plasticity, neurotransmission, and metabolism. PMID- 11102587 TI - Depo-provera associated with weight gain in Navajo women. AB - Depo-medroxyprogesterone acetate (DMPA) is an increasingly popular contraceptive choice among Navajo women. Weight gain is cited as a common side effect and major reason for discontinuation of DMPA. No controlled trials have evaluated the association between weight gain and DMPA in Navajo women. We aimed to clarify whether DMPA is associated with weight gain in Navajo women and to quantify the magnitude of weight gain. A cohort of 172 Navajo women who had used DMPA continuously for one or 2 years comprised the study group. A cohort of 134 Navajo women who used a non-progestin method or no method over 1 or 2 years comprised the comparison group. Initial weight, one-year weight and 2-year weights were recorded for all patients. Study subjects gained a mean of 6 pounds over one year and 11 pounds over 2 years relative to the comparison group (p < 0.001) after controlling for possible confounding variables including age, parity and initial weight. Use of DMPA is associated with significant weight gain in Navajo women. This weight gain is greater than that reported in previous uncontrolled studies in non-Navajo populations. This information should be utilized in counseling Navajo women about the side effects of DMPA. PMID- 11102588 TI - Acceptability and performance of the levonorgestrel-releasing intrauterine system (Mirena) in Campinas, Brazil. AB - The objectives of this study were to evaluate the acceptability of the LNG-IUS Mirena(R), when offered as an additional option in a free choice context, and to evaluate the possibility of using this method in women with increased bleeding wanting an IUD and in copper IUD users requesting removal of the device for bleeding problems. A total of 256 women chose Mirena and were accepted into the study during the enrollment period. This represents 23.3% of all new acceptors of contraceptive methods in the clinic during the same period. Discontinuations were fairly evenly distributed among expulsion, bleeding changes, pain, and personal reasons. Bleeding changes were decreased bleeding, oligomenorrhea or amenorrhea. Comparing the performance in the group of women who chose the LNG-IUS as a first option with those having heavy bleeding, the only difference found was a higher expulsion rate in the group with bleeding problems. No pregnancies occurred and continuation rate was slightly over 75% in the total sample and in both groups. The characteristics of the LNG-IUS (Mirena) allow predicting that this method can effectively contribute to the increase in contraceptive options when introduced to family planning programs in Brazil. PMID- 11102589 TI - Structural integrity of the female condom after a single use, washing, and disinfection. AB - The Reality(R) female condom is approved for use during a single act of intercourse, but is expensive relative to other barrier methods. Re-use is a potential strategy to reduce its per-use cost. We tested the structural integrity of female condoms (n = 318) after a single act of vaginal intercourse. We also measured the impact of laboratory washing (1, 5, or 10 times) with and without disinfection on the structural integrity of unused condoms. Structural integrity was measured via 5 tests: seam tensile strength, water leakage, air-burst, tear propagation, and device dimensions. No degradation in device structural integrity occurred after a single use when compared to control for seam tensile (16.0 vs.15.7 mPa; p = 0.558); water leakage (1.9% vs. 0.9%; p = 0.618); air burst (3.9 vs. 3.6 kPa; p <0.001); or tear propagation (344.6 vs. 336.8 psi; p = 0.313). Mean length was slightly increased [single use vs. control (177.9 vs. 172.5 mm; p <0.001)]. No consistent pattern of structural degradation emerged across all wash/disinfection groups. Our data suggest the structural integrity of the female condom remains intact after a single use and cleaning. PMID- 11102590 TI - Efficacy and acceptability of testosterone implants, alone or in combination with a 5alpha-reductase inhibitor, for male hormonal contraception. AB - Testosterone (T) treatment suppresses serum gonadotropins and reduces sperm output sufficiently for contraceptive efficacy in approximately 70% of Caucasian men. In the remaining 30% of men, an increase in 5alpha-reductase activity may maintain testicular androgen activity, thus accounting for the failure of sperm suppression. The form of T therapy is a major consideration in the safety and acceptability of T-based contraception. As compared to T ester injections, T implants provide a more physiological serum T profile and fewer side effects, but have not yet been used in contraceptive efficacy studies. We have used T implants (800-1200 mg every 3 months) in 29 normal men for 3-16 months. T implants produced long-term suppression of sperm densities below 1 million/mL in approximately 70% of men without significant androgenic side effects. No pregnancies occurred in 214 months of exposure. In 16 men failing to suppress within 3 months of T 800 mg, no evidence of enhanced spermatogenic suppression was seen with the co-administration of the type 2 5alpha-reductase inhibitor, finasteride, for 3 months when compared to placebo. We conclude that: 1) T implants provide adequate spermatogenic suppression in approximately 70% of Caucasian men, a rate comparable to intramuscular T injections but with minimal side effects; and, 2) the inclusion of a type 2 5alpha-reductase inhibitor does not enhance spermatogenic suppression. PMID- 11102591 TI - Demographic and clinical profile of human immunodeficiency virus-infected postpartum women who undergo sterilization. AB - The decision of human immunodeficiency virus (HIV)-infected women to accept a contraceptive method has implications related to the prevention of HIV infection to their children. A case-control study was performed in 57 HIV seropositive pregnant women with prenatal care and delivery at the National Institute of Perinatology, Mexico City. Thirty-five cases were women who accepted postpartum sterilization and twenty-two controls were women who refused this method. The acceptance of tubal occlusion was statistically more frequent in multiparous women, and in those with previous children infected with HIV. The antecedent of at least one previous pregnancy had an association with the acceptance of tubal occlusion with an OR of 11.2 (CI 95% 2.9 to 42.9); having at least one previous child HIV-infected had an OR of 4.6 (CI 95% 1.3 to 23.1). The stratified analysis did not show modification of the association strength between previous pregnancy and the precedent of previous children HIV-infected with the acceptance of sterilization. PMID- 11102592 TI - A randomized comparison of the effects on vaginal and cervical epithelium of a placebo vaginal ring with non-use of a ring. AB - Little is known about the effects of contraceptive vaginal rings on the vaginal surface epithelium, although most studies have not demonstrated any significant deleterious effect. However, one study found that some medium-to-long-term levonorgestrel-releasing ring users developed chronic erythematous and ulcerative lesions in the posterior vaginal fornix. Subsequently, this ring was completely redesigned (IVR-2) with different dimensions and much greater flexibility. The first version of IVR-2 was designed as a placebo ring to explore effects on the vagina and cervix without addition of a progestogen. One-hundred-sixty-six healthy sexually active women volunteers were recruited in four centers and randomly assigned for 6 months to either placebo ring use or control (non-use) using a predetermined randomization code generated by WHO in a 2:1 ratio. Careful inspections of the vaginal and cervical epithelium were performed with a colposcope at admission and at 2-month intervals. No clinically significant lesions were detected in any center either among ring users or controls. However, a number of minor changes in appearance of the vaginal and cervical epithelium (erythema, petechiae, ecchymosis, and minor aceto-white changes) were described from the Sydney Center, some of which were present on admission and some of which were found on subsequent examination. Ten of eleven "red" changes on the cervix and vagina were noted in IVR-2 users, and only one in the controls, suggesting a contribution by the IVR-2 to minor epithelial surface changes. Five of ten resolved completely with continued ring use. There was no correlation in this study between epithelial changes and cigarette smoking or frequency of intercourse in the 14 days prior to colposcopic examination but a significant relationship between tampon use in the last 7 days and all epithelial changes (p = 0.05) and especially red changes (p = 0. 027) was noted. Red changes were significantly less likely to be found among condom users (p = 0.007). The IVR-2 placebo ring did not produce clinically significant changes in the vaginal epithelium and cervical mucosa and a carefully controlled and randomized study should be considered to compare the epithelial appearances in women using a placebo IVR-2 and one releasing 20 microg levonorgestrel. PMID- 11102593 TI - A new vaginal antimicrobial contraceptive formulation: phase I clinical pilot studies. AB - Pilot clinical trials were performed with a new vaginal suppository called "Long Acting, Sustained Release of Spermicide" ("LASRS"). No visual or colposcopic lesions or patient complaints occurred as a result of using LASRS with increasing doses of nonoxynol-9 (up to 20%) for 5 days or of applying the highest dose of nonoxynol-9 (20%; total 400 mg) for 8 h. Colposcopic or visual lesions were also not induced when LASRS with 20% nonoxynol-9 was used for 7 consecutive days by the study participants except for those who developed symptomatic monilia vaginitis. Symptoms were reported although these were mostly minor. A long lasting, bioadhesive, translucent layer (film) of formulation formed over the vaginal and cervical surfaces. Postcoital spermicidal studies showed LASRS to be highly effective for prolonged periods of time. Although intercourse was delayed for 5 to 8.5 h after insertion of the formulation, an average of only 0. 2 motile sperm/HPF could be found in cervical mucus. These studies suggest LASRS to possess advantages over presently marketed formulations by having long-term efficacy and by forming a bioadhesive, presumably protective layer over the genital tract epithelium. The results also suggest the formulation to decrease the vaginal irritation caused by nonoxynol-9 as noted by colposcopy. These pilot data support a more extensive study with the LASRS suppository. PMID- 11102594 TI - Cyclodextrin enhances spermicidal effects of magainin-2-amide. AB - Magainins are antimicrobial peptides with known spermicidal activity. Their activity is inhibited by cholesterol present in eukaryotic cell membranes. Pretreatment of spermatozoa with methyl-beta-cyclodextrin, which extracts cholesterol from cell membranes and induces capacitation, sensitizes them to magainin-2-amide as shown by a decrease in human sperm motility determined by computer-assisted sperm analysis and a concomitant decrease in sperm viability, as measured by MitoTracker(R) Red CMXRos labeling. Magainin-2-amide affects mainly the fast progressive spermatozoa inducing them directly into an immotile state, without an increase in motile non-progressive and slow progressive spermatozoa. We conclude that methyl-beta-cyclodextrin highly potentiates the deleterious effect of magainin-2-amide on human spermatozoa. Most probably, this effect can be explained by cholesterol extraction from sperm cell membranes. PMID- 11102595 TI - Hypersensitivity reactions following laminaria placement. AB - Laminaria tents are commonly placed intracervically prior to elective termination of pregnancy. Three women, each of whom had undergone at least one previous abortion in which a laminaria was utilized, developed hypersensitivity reactions following laminaria placement. The reactions included urticaria, angioedema and respiratory distress. All responded to removal of the laminaria and administration of either diphenhydramine, prednisone, inhaled bronchodilators or subcutaneous epinephrine. One of the women subsequently underwent skin testing and was positive to laminaria. Patients undergoing laminaria placement may manifest a Type I reaction, IgE-mediated sensitivity. Providers should counsel patients with histories of multiple previous laminaria insertions about this possibility, have alternative methods of cervical dilation available, and be able to promptly recognize and treat reactions when they occur. PMID- 11102596 TI - Oral contraceptives and cardiovascular risk-an end to the debate? PMID- 11102597 TI - Oral contraceptives and cardiovascular outcomes: cause or bias? AB - The 1995-1996 "pill" scare, which suggested that third-generation oral contraceptives (OCs) were associated with a greater risk of venous thromboembolism (VTE) than second-generation OCs, had serious social and public health consequences, as women discontinued OCs, resulting in unwanted pregnancies and unnecessary abortions. This article uses the Bradford Hill criteria, for diagnosing causality from an observed association, to interpret evidence from recent studies as to whether there is any difference in the risk of VTE between third- and second-generation OCs. Bias and the influence of confounders have also been examined in relation to the difference in the risk of VTE between third- and second-generation OCs reported in the 1995-1996 studies. It is clear from the results of this analysis that none of the Bradford Hill criteria are fulfilled. Thus, a causal relationship cannot be inferred from the alleged association of third-generation OCs with VTE. Indeed, it would appear that the unavoidable bias in observational research is a more likely explanation for the apparent difference in the risk of VTE between third- and second-generation OCs in the 1995-1996 studies. Recent studies, which employed more appropriate controls for these biases showed no difference in the risk of VTE between third- and second generation OCs. A Danish study (1994-1996) demonstrated a lower risk of thrombotic morbidity and mortality with third-generation OCs compared with second generation OCs. In addition, the Transnational study has shown that third generation OCs have a significantly lower relative risk (0.3 [0.1-0.9]) for acute myocardial infarction (MI) compared with second-generation products. In conclusion, there is no convincing evidence for a difference in the risk of stroke or VTE between third- and second-generation OCs. Moreover, third generation OCs may be associated with a lower risk of MI than second-generation OCs. PMID- 11102598 TI - Hemostatic effects of third- and second-generation oral contraceptives: absence of a causal mechanism for a difference in risk of venous thromboembolism. AB - Some observational studies have found a difference in the risk of nonfatal venous thromboembolism (VTE) with low-dose, oral contraceptives (OCs) containing desogestrel (DSG) or gestodene (GSD) and those containing levonorgestrel (LNG). However, this does not agree with current pathophysiological concepts. This review compares all 17 comparative studies on the hemostatic effects of DSG/GSD- and LNG- or norgestimate (NGM)-containing OCs, and comments on two recent cross sectional studies on the effects of third- and second-generation OCs on activated protein C (APC) sensitivity. In the comparative studies, the only difference in hemostatic parameters between DSG/GSD- and LNG- or NGM-containing OC users was a tendency towards higher factor VII (FVII) levels with DSG/GSD OCs. Differential effects on APC sensitivity were observed with the endogenous thrombin generation potential (ETP) assay, but not with the classical APC resistance test. FVII is not a risk marker for VTE, but is affected by dietary fat, estrogens and androgens and may interfere with the ETP assay. As no differences in established VTE risk markers were observed, there is no plausible reason for a differential risk of VTE with DSG/GSD- and LNG-containing OCs. In fact, the lack of differences with regard to established risk markers of VTE gives further support to the findings of the most recent epidemiological studies, which have not found any difference in the risk of VTE between third- and second-generation OCs. PMID- 11102599 TI - Venous thromboembolism and combined oral contraceptives: does the type of progestogen make a difference? AB - Venous thromboembolism (VTE) is rare in young women but is associated with the use of combined oral contraceptives (OCs). In 1995 and 1996, three studies showed a difference in the risk of VTE with third-generation OCs containing the progestogens, desogestrel or gestodene, compared with earlier formulations. However, the subsequent MediPlus study did not show any difference in the risk of VTE between users of third- and second-generation OCs. To re-examine the risks of VTE with various OCs, a nested case-control study was undertaken using the General Practice Research Database (GPRD). This study identified 293 cases and selected up to four controls matched for year of birth, practice, and event date. Adjustment for confounding variables included: body mass index, smoking, asthma, diastolic blood pressure, and a proxy for recent illness. The new analysis of the GPRD showed that there was no statistically significant difference in the risk of VTE among users of third-generation OCs compared with second-generation OCs containing levonorgestrel 150 microg plus ethinylestradiol 30 microg. Important associations with idiopathic VTE included: age, obesity, smoking, recent concurrent illness, and asthma. Thus, any difference previously noted between OCs containing desogestrel or gestodene and levonorgestrel are likely to be due to the healthy-user effect, prescribing bias and inadequate control of known confounding variables, such as age and obesity. PMID- 11102600 TI - Emerging evidence on oral contraceptives and arterial disease. AB - A number of case-control studies published in recent years have shown an apparent increase in the risk of venous thromboembolism (VTE) associated with the use of third-generation oral contraceptives (OCs) compared with second-generation OCs. However, it is thought that these studies were subject to a number of biases that would have increased the risk estimates for third-generation OCs while lowering those for second-generation preparations. Data on the risk of ischemic stroke and myocardial infarction (MI) show no such difference between generations of OCs, with a statistically significant reduction in the risk of acute MI from first- to third-generation in one major study. Available results indicate that there is no significant increase in the risk of ischemic stroke or acute MI associated with the use of low-dose estrogen OCs in young women who are properly screened before use, and who have no pre-existing cardiovascular risk factors, such as smoking and hypertension, for these conditions. These findings should be taken into account when interpreting the results of studies on the risk of VTE in women taking combined OCs. PMID- 11102601 TI - Brief communications: why they are important PMID- 11102602 TI - The impact of free antenatal care on perinatal mortality. PMID- 11102603 TI - Factors affecting perinatal mortality in India. PMID- 11102604 TI - The challenge of reducing maternal mortality in Nigeria. PMID- 11102605 TI - Pregnancy and term delivery after neovaginoplasty in a patient with vaginal agenesis. PMID- 11102606 TI - Potential efficacy of nitric oxide for cervical ripening in pregnancy at term. PMID- 11102607 TI - Detection of Gardnerella vaginalis in the vagina and amniotic fluid using the polymerase chain reaction. PMID- 11102608 TI - Neurological complications associated with the pre-eclampsia/eclampsia syndrome. PMID- 11102609 TI - Incidence and risk factors for third degree perineal tears. PMID- 11102610 TI - Fourth-degree lacerations and epidural anesthesia. PMID- 11102611 TI - The value of a clinical fetal biophysical profile in high risk pregnancies. PMID- 11102612 TI - Loop electrosurgical excision procedure (LEEP) during first trimester of pregnancy. AB - Loop electrosurgical excision procedure (LEEP) during pregnancy can be performed safely as in non-pregnant women and can replace traditional cone biopsy when performed in the first trimester. PMID- 11102613 TI - Laparoscopic removal of translocated retroperitoneal IUD. PMID- 11102614 TI - Regression of liver metastases after high-dose chemotherapy and peripheral-blood progenitor-cell support in Stage IV ovarian cancer. PMID- 11102615 TI - Estrogen-progestagen pre-treatment before HMG induction in hypogonadotropic patients. PMID- 11102616 TI - Role of transvaginal ultrasonography and diagnostic hysteroscopy in the evaluation of patients with abnormal uterine bleeding. PMID- 11102617 TI - Is peritoneal closure necessary after abdominal hysterectomy? PMID- 11102618 TI - Sexual assault--a neglected public health problem in the developing world. PMID- 11102619 TI - DNA-ploidy as an independent prognostic factor in patients with serous ovarian carcinoma. PMID- 11102620 TI - Ultrasonography and ectopic pregnancy--a review. PMID- 11102621 TI - Abortion among rural women in north Ethiopia. PMID- 11102622 TI - Association of Acanthosis nigricans with risk of diabetes mellitus and hormonal disturbances in females. AB - OBJECTIVE: To determine the association of Acanthosis nigricans, hyperinsulinemia, and hormonal levels in female subjects from the United Arab Emirates (UAE). DESIGN: Prospective study. SETTING: Tawam Teaching Hospital of Faculty of Medicine & Health Sciences. SUBJECTS: Ninety-two females (age range 16 65 years) were recruited. METHODS: Height, weight, and sitting blood pressure were recorded on 92 female subjects with A. nigricans. Fasting blood samples were obtained for measurement of uric acid, glucose, cholesterol, high-density lipoprotein- (HDL) cholesterol, and triglyceride levels. Serum levels of thyroid stimulating hormone (TSH), testosterone, luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin are obtained by radio-immunoassay. RESULTS: Ninety-two females with A. nigricans were enrolled in the study. Of the 92 females, 36 subjects were considered to have diabetes mellitus (DM) and 56 euglycemic subjects. The analysis showed that in cases of family history of DM, HDL-cholesterol (mmol/l) and uric acid (mmol/l) levels were higher. Overall, DM subjects had significantly higher values for hormone levels of TSH, FSH, LH, progesterone, testosterone, cortisol, prolactin, (growth hormone) GH, and ferritin. CONCLUSION: Patients with A. nigricans have a high prevalence of DM and insulin resistance in UAE. Since A. nigricans is rather prevalent in the UAE, identifying this skin lesion can help detect those subjects with a higher risk of DM and hormonal disturbances. PMID- 11102623 TI - Experience with the intrauterine device (TCU-380A) in Enugu, eastern Nigeria. PMID- 11102624 TI - Successful treatment of pelvic recurrent vulvar melanoma. PMID- 11102625 TI - Expression of p53 in epithelial ovarian cancer. PMID- 11102627 TI - Nuchal translucency: association of size and persistence with fetal abnormalities. PMID- 11102626 TI - Short anovaginal distance: a risk factor for recurrent vaginitis. PMID- 11102628 TI - Hysteroscopic surgery for endometrial polyps using a bipolar microelectrode. PMID- 11102629 TI - Catheterization following abdominal hysterectomy. PMID- 11102630 TI - Intra-mammary lymph node metastases in an early stage ovarian cancer of low malignant potential. PMID- 11102631 TI - Serum CA 125 level as a reflection of proliferative activity of serous borderline ovarian tumor. PMID- 11102632 TI - Ethical guidelines in the prevention of iatrogenic multiple pregnancy. PMID- 11102633 TI - Transcranial magnetic stimulation applications and potential use in chronic pain: studies in waiting. AB - Transcranial magnetic stimulation (TMS) is a new technology which uses electromagnetic principles to produce small electrical currents in the cortex. Evidence indicates that TMS can produce plastic changes in the CNS which are observable at both the cellular and physiological levels. It is proposed that studies are justified to determine whether TMS can provide short-term or long term relief in chronic pain. PMID- 11102634 TI - Early pregnancy factor suppresses experimental autoimmune encephalomyelitis induced in Lewis rats with myelin basic protein and in SJL/J mice with myelin proteolipid protein peptide 139-151. AB - Early pregnancy factor (EPF) is a secreted protein with immunosuppressive and growth factor properties. During pregnancy, it appears in maternal serum within 6 24 h of fertilization, is present for at least the first two-thirds of pregnancy in all species studied and is essential for embryonic survival. It is a homologue of chaperonin 10, a heat shock protein, but, unlike other members of this family, EPF has an extracellular role. As it has the ability to modulate CD4+ T cell dependent immune responses, its role in treatment of experimental autoimmune encephalomyelitis (EAE) was investigated. EAE is a CD4+ T cell-mediated disease, the best available animal model of multiple sclerosis (MS). Two models of EAE were investigated, acute EAE induced in Lewis rats by inoculation with myelin basic protein (MBP-EAE) and chronic relapsing EAE induced in SJL/J mice by inoculation with myelin proteolipid protein peptide (residues 139-151) (PLP-EAE). EPF, delivered intraperitoneally or orally to rats or intraperitoneally to mice, suppressed clinical signs of disease. Mice with PLP-EAE were also treated with interferon-beta, with and without EPF. Both EPF and IFN-beta suppressed clinical signs of EAE and, when administered together, gave greater suppression than when given separately. These findings suggest that EPF may be a potential candidate for use in treatment of MS and may be of use in combined therapy with IFN-beta. PMID- 11102635 TI - Definition of the anterior choroidal artery territory in rats using intraluminal occluding technique. AB - This manuscript delineates the territory of the anterior choroidal artery (AChA) in rats, as defined by the induction of an AChA infarction. By advancing a 0.24 mm surgical suture up the internal carotid artery (ICA) to a point 0.5-2 mm proximal to the middle cerebral artery (MCA) origin, the AChA could be occluded and a reliable AChA distribution infarction was produced in 62% (23/37) of animals. The infarct volume, as defined by TTC staining, was 55+/-7 mm(3). Maps of the infarction, generated by measuring the entire area of overlapping coronal slices, demonstrated that the internal capsule was always damaged. Other areas that might be affected included the hippocampus, thalamus, amygdaloid complex, piriform cortex, dorsal caudatoputamen, and lateral ventricular wall. Positioning the coated suture proximal to the AChA produced a much smaller infarct involving the medial and lateral hypothalamus, preoptic region, optic chiasm, and marginal region of the internal capsule near to the lateral hypothalamus exempt from AChA territory damage. A causative relationship between AChA occlusion and a deep cerebral infarct centered on the internal capsule was further established by: (1) identifying the AChA on the non-ischemic side with colored silicone perfusion, and subsequent similar delineation on the ischemic side, and (2) delineating infarction in the silicone perfused AChA region using hematoxylin and eosin staining and the TUNEL method. The AChA usually originated from the ICA (91% of cases), 1.75+/-0.12 mm proximal to the MCA bifurcation. Approximately 27% of the AChAs had periamygdaloid branch(es) on its initial segment. PMID- 11102636 TI - Cortical-basal ganglionic degeneration: a clinical, functional and cognitive evaluation (1-year follow-up). AB - We decided to evaluate a patient who was diagnosed with cortical-basal ganglionic degeneration from a clinical, instrumental and neuropsychological perspective. Our aim was to employ a new instrumental tool, functional magnetic resonance, in order to evaluate his cortical damage. We then followed the pathological course for 1 year and tested the patient again: we discuss the results of our evaluation, having an overview of the literature on the topic. In particular, we focused our attention on his apraxia, trying to suggest a dynamic and anatomical model to guarantee a possible explanation of his behavior. PMID- 11102637 TI - Adult bacterial meningitis in Southern Taiwan: epidemiologic trend and prognostic factors. AB - In two investigative phases over a 13.5-year study period (January 1986-June 1999), 202 adult patients with culture-proven bacterial meningitis were enrolled in this study. In order to determine the epidemiologic trend, prognostic factors and therapeutic results for this disease. Klebsiella pneumoniae (K. pneumoniae), Pseudomonas aeruginosa, and Streptococcus pneumoniae were the three most commonly revealed pathogens, accounting for about 48% of the episodes. Although there was a change in relative frequency for the pathogens, K. pneumoniae remained the most prevalent during the two periods studied (January 1986-December 1992 and January 1993-June 1999). Multiantibiotic resistant strains have been in evidence since their appearance in 1994, with most of our patients acquiring their infection nosocomially. The overall mortality rates during the two periods were 40% and 34%, respectively. In stepwise logistic regression analysis, only initial conscious level, appropriate antibiotic therapy and septic shock were independently associated with mortality, after adjustment for other potentially confounding factors. Initial empirical antibiotics with both third-generation cephalosporin and penicillin G, should be considered for the majority of meningitis cases resulting from infection with Gram-negative bacilli and streptococcal species. Besides the evolution of newer pathogens, there has been increasing incidence for nosocomially acquired bacterial meningitis for patients postneurosurgery, with the emergence of resistant strains presenting a therapeutic challenge in recent years. Vancomycin and imipenem/cilastatin should be considered as the initial empirical antibiotics of choice for the treatment of this special group of patients. PMID- 11102638 TI - The involvement of the neuronal Golgi apparatus and trans-Golgi network in the human olivary hypertrophy. AB - We studied the Golgi apparatus (GA) and trans-Golgi network (TGN) in the human olivary hypertrophy by immunohistological methods with organelle specific antibodies against the medial cisternae of the organelle (MG160) and the trans Golgi network (TGN46). The GA and TGN of enlarged neurons in the inferior olivary nuclei in the early stages after central tract lesions lost the normal network like configuration, and they were reduced to numerous small disconnected granules (fragmentation). In chronic stages after lesions, the GA and TGN of vacuolated or enlarged neurons showed a variety of morphological profiles, such as normal looking patterns, fragmentation, reduction in number, and aggregation around nuclei or at a distance in the cytoplasm. In patients with multiple system atrophy, the GA and TGN of the neurons in the inferior olivary nuclei showed almost similar findings to those seen in the chronic stages after brainstem lesions. These results suggest that the GA and TGN are affected in degenerating neurons by anterograde transneuronal mechanisms. PMID- 11102639 TI - Crossed linguo-buccal reflex in post-stroke patients. AB - A pathological crossed orofacial reflex, called crossed linguo-buccal reflex in the present study, was observed in approximately 1/3 of post-stroke patients with central facial palsy. Stroking with pressure two or three times with a split wooden tongue-blade to the tongue or palate contralateral to the central facial palsy elicited a reflex movement consisting of retraction of the angle of mouth and medio-posterior withdrawal of the buccal mucosa on the paretic side. Seventy seven patients with central hemifacial palsy caused by a unilateral cerebral lesion were examined clinically, electromyographically and by computed tomography (CT) and magnetic resonance imaging (MRI). In addition, three men with bilateral cerebral lesions and bilateral crossed linguo-buccal reflexes were electromyographically examined. Twenty-two patients with unilateral cerebral lesions had this reflex. It was found that this reflex was most frequently observed in patients with a capsulo-caudate lesion involving the head of the caudate nucleus, the anterior limb and genu of the internal capsule. The electromyogram of the reflex showed increased activity in the orbicularis oris, depressor anguli oris, risorius, zygomaticus major and buccinator muscles on the paretic side with a long latency (254-856 ms), and a prolonged after-discharge after the stimulation. Reciprocal inhibition was observed in patients with bilateral positive reflexes. These findings suggest that liberation of the polysynaptic brainstem reflex in the medulla oblongata and pons from the indirect corticobulbar inhibition may underlie the occurrence of the crossed linguo-buccal reflex in post-stroke patients. PMID- 11102640 TI - Defective electron transfer in complexes I and IV in patients with aceruloplasminemia. AB - Aceruloplasminemia is a disorder of iron metabolism caused by mutations in the ceruloplasmin gene. It is characterized by progressive neurodegeneration of the retina, basal ganglia, dentate nucleus and cerebral cortex in association with iron accumulation in these tissues. Enzyme activities in the mitochondrial respiratory chain of the cerebral cortices of two patients were reduced to 62% and 71% for complexes I and IV. Malondialdehyde, a marker of lipid peroxidation, was three times higher than the control value and was accompanied by increased expression of superoxide dismutase 2 (Mn SOD). These findings suggest that iron mediated free radicals contribute to the impairment of mitochondrial energy metabolism in aceruloplasminemia PMID- 11102641 TI - Recovery of brain dysfunction after methylmercury exposure in rats. AB - We studied the time course of central nervous system (CNS) involvement after the termination of methylmercury exposure to rats, in order to investigate whether or not the involvement still progresses even after the termination of exposure. Methylmercury chloride (MMC), at a dose of 2 mg/kg/day, was subcutaneously injected for 25 consecutive days in 12 adult male Sprague-Dawley rats. Six of them were sacrificed on the final day of exposure (group A) after completing the observations of behavioral changes and determining the local cerebral glucose utilization (LCGU) as an indicator of cerebral neuronal activities. Histological examinations of the brain and the sciatic nerve were done. The other six rats were further followed up for 90 days after the termination of exposure (group B). In addition, six rats that received physiological saline served as a control. Group A showed a significant reduction of LCGU without any accompanying cerebral histological alterations and a moderate loss of myelinated fibers in the sciatic nerve. Group B showed normal LCGU rates while severe axonal degeneration of the sciatic nerve was found on the final day of the 90-day follow-up period. The present results demonstrate that a transient involvement of the CNS can occur after MMC exposure. In addition, a complete recovery may occur when the process is mild enough not to cause histological alterations. In contrast, the involvement of the peripheral nerve is much more severe than that of the CNS and it was observed to progress even after the cessation of MMC exposure. Therefore, it seems unlikely, at least in rats, that a steadily progressive course occurs in the CNS but not in the peripheral nerves over a long period of time after MMC exposure. PMID- 11102642 TI - Chronic effects of methylmercury on the cerebral function in rats. AB - We studied the effects of the long-term and small-dose administration of methylmercury chloride (MMC) on the cerebral function in rats. MMC, at a dose of 0.7 mg/kg/day, was subcutaneously injected for 85 consecutive days in nine adult male Sprague-Dawley rats. They were then sacrificed on the final day of exposure (MMC group) after both completing observations on behavioral changes and also determining the local cerebral glucose utilization (LCGU) as an indicator of the cerebral neuronal activities. Histological examinations of the brain and the sciatic nerve were also performed. In addition, seven rats who received physiological saline also served as a control. LCGU significantly decreased in the visual cortex, lateral geniculate nucleus and medial geniculate nucleus without any accompanying histological alterations. Severe axonal degeneration of the sciatic nerve was also observed, which corresponded to the previously described crossed leg phenomenon. The present results suggest that the damage to the peripheral nerve was much more severe than that to the brain, which caused behavioral changes. Although no cerebral morphological changes were observed, brain dysfunction showed a selective involvement of the visual and auditory systems. This finding suggests that LCGU is a sensitive method for detecting the subclinical cerebral dysfunction caused by long-term and small-dose MMC intoxication in the rat brain. PMID- 11102643 TI - Asymptomatic CTG expansion at the SCA8 locus is associated with cerebellar atrophy on MRI. AB - Spinocerebellar ataxia type 8 (SCA8) is the first example of dominantly inherited ataxia reported to be caused by a dynamic mutation of the untranslated CTG trinucleotide repeat. We performed genetic and clinical analyses of a family with an isolated case with young onset cerebellar ataxia carrying an expanded 95 CTA/CTG repeats, and revealed that the asymptomatic father was also carrying a much greater expansion of 136 repeats. This paternal transmission developed a large contraction of -41 CTG repeats. The ataxia patient showed almost pure cerebellar symptoms, and a cerebral MRI of the patient demonstrated significant atrophy of the cerebellar vermis and hemispheres with preservation of brainstem and cerebrum. Although the father did not show any neurological abnormalities, his MRI demonstrated mild atrophy of the cerebellar hemispheres. The genetic phenomenon on this family has not been observed in other types of SCAs, and this reduced penetrance may cause reproduction of sporadic SCA8 frequently. Therefore, we must perform careful interviews regarding family history, and suggest the genetic and neuroradiological investigations on family members when we encounter a sporadic patient with the CTG expansion at the SCA8 locus. PMID- 11102644 TI - Neuropeptides 2000. Proceedings of the 10th Annual Meeting of the European Neuropeptide Club. Innsbruck, Austria, May 10-13, 2000. PMID- 11102645 TI - Comparison of currents activated by noxious heat in rat and chicken primary sensory neurons. AB - The vanilloid receptor 1 (VR1) gene is responsible for both capsaicin-, and low threshold (LT) noxious heat-sensitivity in mammalian primary sensory neurons. Although, birds lack capsaicin-sensitivity they express LT noxious heat sensitivity. Here, we compared LT noxious heat-activated whole-cell currents produced by rat and chicken cultured dorsal root ganglion neurons in order to find out the similarities and differences in the LT noxious heat transduction mechanisms between the two species. No significant differences between rat and chicken neurons were found in the mean cell diameter of the LT noxious heat sensitive cells (20.4+/-0.8 microm, n=19 and 20.6+/-0.6 microm, n=11, respectively) and the average threshold (45.7+/-0.5 degrees C, n=19 and 46.1+/ 0.7 degrees C, n=11, respectively) and peak amplitude (-2.9+/-0.6 nA, n=19 and 2.1+/-0.6 nA, n=11, respectively) of the heat-evoked responses. The current voltage curves of the responses both in rat and chicken cells reversed at the same range (-19.5+/-3.8 mV, n=4 and -15.5+/-1. 2 mV, n=3, respectively) and showed strong outward rectification at negative membrane potentials. While all LT noxious heat-sensitive rat cells responded to capsaicin, none of the chicken neurons produced detectable response to it. These findings suggest that a VR1 homologue which lacks to sequence for capsaicin-sensitivity is possibly the LT noxious heat transducer in chicken. PMID- 11102646 TI - NKP608: a selective NK-1 receptor antagonist with anxiolytic-like effects in the social interaction and social exploration test in rats. AB - NKP608 is a non-peptidic derivative of 4-aminopiperidine which acts as a selective, specific and potent antagonist at the neurokinin-1 (NK-1) receptor both in vitro and in vivo. In vitro, the binding of NKP608 to bovine retina was characterized by an IC50 of 2.6+/-0.4 nM, whereas the compound's affinity to other receptor binding sites, including NK-2 and NK-3, was much lower. Species differences in IC(50) values with NKP608 were less pronounced than with previously described NK-1 receptor antagonists, being 13+/-2 and 27+/-2 nM in gerbil midbrain and rat striatum, respectively. In vivo, using the hind foot thumping model in gerbils, NKP608 exhibited a potent NK-1 antagonistic activity following oral administration (ID(50)=0.23 mg/kg; 2 h pretreatment), supporting a central activity of NKP608. The compound had a long duration of action with an ID(50) value of 0. 15 mg/kg p.o. and 0.38 mg/kg p.o. following a pretreatment of 5 and 24 h, respectively. Following a subchronic administration for 7 consecutive days (once daily) there was no evidence for the development of tolerance or accumulation. In the social interaction test performed in a highly illuminated, unfamiliar test arena, NKP608 specifically increased the time the two rats spent in social contact, and there was no concomitant increase in parameters reflecting general activity, i.e. ambulation (number of square entries) or the number of rearings. Active social time was maximally increased at a dose range of 0.01-1 mg/kg p.o. NKP608, the effect being weaker or absent at both lower (0.001 mg/kg p.o.) and higher (10 mg/kg p.o.) doses. A comparable bell-shaped dose-response relation was seen in the social exploration test in rats. In this modified resident/intruder paradigm, maximal increase in social contact of the intruder rat directed towards the resident rat was seen at a similar dose range (0.03-3 mg/kg p.o.) The effects observed following an acute oral administration of NKP608 were comparable to those seen following a treatment with the well-known benzodiazepine, chlordiazepoxide, in both these tests. These findings indicate that NKP608 exhibits an anxiolytic-like effect and that this effect, as concluded from the observed antagonism of the hind foot thumping induced by i.c.v. administration of the NK-1 receptor agonist SPOMe, is centrally mediated. This makes this compound a potentially promising candidate for treating anxiety related disorders in humans. PMID- 11102647 TI - Differential role of neurokinin receptors in human lymphocyte and monocyte chemotaxis. AB - The precise nature of neurokin receptor involvement in human immune cell chemotaxis is unclear. This study therefore sought to directly compare the chemotactic effects of neurokinins on human T lymphocytes and monocytes. Substance P was found to have a similar dose-dependent chemotactic action on T lymphocyte and monocyte populations. In contrast, T lymphocytes were found to be more responsive than monocytes both to the highly selective NK-1 agonist, [Sar(9)Met O(2)(11)]-substance P, and also to the NK-2 selective agonist, beta alanine neurokinin A((4-10)). Consistent with these findings, substance P-induced chemotaxis of both T lymphocyte and monocytes was attenuated by the selective NK 1 antagonist LY303870. However, the selective NK-2 antagonist MEN 10,376 was only effective in inhibiting the T lymphocyte response. The study confirms that neurokinins have chemotactic actions on immune cells and indicates important functional differences between human T lymphocyte and monocyte responses. This provides a potential mechanism by which the nervous system can selectively influence cellular recruitment in inflammatory disease. PMID- 11102648 TI - Vasopressinergic regulation of the hypothalamic-pituitary-adrenal axis: implications for stress adaptation. AB - In addition to its role on water conservation, vasopressin (VP) regulates pituitary ACTH secretion by potentiating the stimulatory effects of corticotropin releasing hormone (CRH). The pituitary actions of VP are mediated by plasma membrane receptors of the V1b subtype, coupled to calcium-phospholipid signaling systems. VP is critical for adaptation of the hypothalamic-pituitary-adrenal (HPA) axis to stress as indicated by preferential expression of VP over CRH in parvocellular neurons of the hypothalamic paraventricular nucleus, and the upregulation of pituitary VP receptors during stress paradigms associated with corticotroph hyperresponsiveness. V1b receptor mRNA levels and coupling of the receptor to phospolipase C are stimulated by glucocorticoids, effects which may contribute to the refractoriness of VP-stimulated ACTH secretion to glucocorticoid feedback. The data suggest that vasopressinergic regulation of the HPA axis is critical for sustaining corticotroph responsiveness in the presence of high circulating glucocorticoid levels during chronic stress. PMID- 11102649 TI - Brain oxytocin inhibits the (re)activity of the hypothalamo-pituitary-adrenal axis in male rats: involvement of hypothalamic and limbic brain regions. AB - In response to various stressors, oxytocin is released not only into blood, but also within hypothalamic and extrahypothalamic limbic brain regions. Here, we describe the involvement of intracerebrally released oxytocin in the regulation of the activity of the hypothalamo-pituitary-adrenal (HPA) axis by infusion of the oxytocin receptor antagonist (des Gly-NH(2) d(CH(2))(5) [Tyr(Me)(2), Thr(4)] OVT; pH 7.4; Dr. M. Manning, Toledo, OH, USA) either into the lateral cerebral ventricle (icv[0.75 microg/5 microl,]) or via retrodialysis (10 microg/ml, 3.3 microl/min, 15 min) into the hypothalamic paraventricular nuclei (PVN), the medio lateral septum or the amygdala. Male Wistar rats fitted with a chronic jugular vein catheter and an icv guide cannula or a microdialysis probe targeting the respective brain region 4 days prior to the experiment were blood sampled under basal as well as stressful conditions. Rats were exposed to the elevated platform (emotional stressor) and/or to forced swimming (combined physical and emotional stressor). Blockade of the receptor-mediated action of endogenous oxytocin within the PVN resulted in an enhanced basal secretion of ACTH whereas, in response to forced swimming, ACTH secretion was rather reduced, indicating a tonic inhibitory effect of OXT on basal HPA axis activity, but a potentiating action under conditions of stress. Within the medio-lateral septum, antagonist treatment did not alter basal ACTH secretion, but significantly disinhibited ACTH secretion in response to the elevated platform, but not to forced swimming. Within the amygdala, no significant effects either on basal or stress-induced HPA axis activity could be found. The results indicate a differential involvement of brain oxytocin in the regulation of the HPA axis activity which depends both on the site of intracerebral oxytocin release and the stressor the animals are exposed to. PMID- 11102650 TI - A novel peptide prevents death in enriched neuronal cultures. AB - We have recently cloned a novel protein (activity-dependent neuroprotective protein, ADNP) containing an 8-amino-acid, femtomolar-acting peptide, NAPVSIPQ (NAP). Here we show, for the first time, that NAP exerted a protective effect on glia-depleted neurons in culture. The number of surviving neurons was assessed in cerebral cortical cultures derived from newborn rats. In these cultures, a 24-h treatment with the beta-amyloid peptide (the Alzheimer's disease associated toxin) induced a 30-40% reduction in neuronal survival that was prevented by NAP (10(-13)-10(-11) M). Maximal survival was achieved at NAP concentrations of 10( 12) M. In a second set of experiments, a 5-day incubation period, with NAP added once (at the beginning of the incubation period) exhibited maximal protection at 10(-10) M NAP. In a third set of experiments, a 10-min period of glucose deprivation resulted in a 30-40% neuronal death that was prevented by a 24-h incubation with NAP. Glucose deprivation coupled with beta-amyloid treatment did not increase neuronal death, suggesting a common pathway. We thus conclude, that NAP can prevent neurotoxicity associated with direct action of the beta-amyloid peptide on neurons, perhaps through protection against impaired glucose metabolism. PMID- 11102651 TI - A selective orexin-1 receptor antagonist reduces food consumption in male and female rats. AB - A variety of evidence implicates the orexins, especially orexin-A, in the regulation of food intake, but it has not been established whether this effect is mediated by the orexin-1 or orexin-2 receptor. In the present study, a selective orexin-1 receptor antagonist, 1-(2-methylbenzoxazol-6-yl)-3-[1,5]naphthyridin-4 yl urea hydrochloride (SB-334867-A), was administered intraperitoneally to rats under various conditions, and food consumption was subsequently measured over 24 h. In male rats, a single dose of SB-334867-A (30 mg/kg, i.p.) given during the light phase reduced both orexin-A-induced food intake (7 nmol, i.c.v.) and feeding stimulated by an overnight fast for 4 h. When given at the start of the dark phase, food consumption was reduced in both male and female rats over 24 h. Daily injections at the start of the dark phase for 3 days reduced natural feeding in male rats over 24 h on days one and three. These findings demonstrate direct inhibition of orexin-A induced food intake with a selective orexin-1 receptor antagonist. Furthermore, the suppression of nocturnal feeding and food intake stimulated by an overnight fast supports other evidence that orexin-A is involved in the regulation of natural feeding and suggests that orexin-1 receptor antagonists could be useful in the treatment of obesity. PMID- 11102652 TI - Alterations within the endogenous opioid system in mice with targeted deletion of the neutral endopeptidase ('enkephalinase') gene. AB - The biological inactivation of enkephalins by neutral endopeptidase (enkephalinase, NEP, EC3.4.24.11) represents a major mechanism for the termination of enkephalinergic signalling in brain. A pharmacological blockade of NEP-activity enhances extracellular enkephalin concentrations and induces opioid dependent analgesia. Recently, knockout mice lacking the enzyme NEP have been developed [Lu et al., J. Exp. Med. 1995;181:2271-2275]. The present study investigates the functional consequences and biochemical compensatory strategies of a systemic elimination of NEP activity in these knockout mice. Using biochemical and behavioural tests we found that the lack of NEP activity in brain is not compensated by enhanced activities of alternative enkephalin-degrading enzymes. Also no change in enkephalin biosynthesis was detectable by in situ methods quantifying striatal proenkephalin-mRNA levels in NEP-deficient mice compared with wildtype. Only a 21% reduction of mu receptor density in crude brain homogenates of NEP knockout mice was observed, while delta- and kappa opioid receptor densities were unchanged. This receptor downregulation was also confirmed functionally in the hot-plate paradigm. NEP knockouts developed normally, but showed enhanced aggressive behaviour in the resident-intruder paradigm, and altered locomotor activity as assessed in the photobeam system. Thus, although NEP plays a substantial role in enkephalinergic neurotransmission, the biochemical adaptations within the opioid system of NEP-deficient mice are of only modest nature. PMID- 11102653 TI - Retinoic acid treatment enhances the acetylcholine contents in the human teratocarcinoma cell line NTera-2. AB - Human NTera-2/clone D1 teratocarcinoma cells are induced by retinoic acid (RA) to differentiate into postmitotic cells with morphological and biochemical characteristics of embryonic human neurones. Currently only limited information concerning peptide-contents and neurotransmitter pools of these cells is available. Zeller and Strauss [Int. J. Dev. Neurosci. 1995;13(5):437] described an increase in choline acetyltransferase (ChAT) activity in RA-treated, but not in untreated NTera-2 cells, suggesting the induction of a cholinergic phenotype during treatment with RA. In the present study we investigated the effect of RA differentiation on the amount of the neurotransmitters acetylcholine (ACh), and dopamine in NTera-2 in order to specify the transmitter phenotype induced by RA differentiation. We found that a 4-week treatment of NTera-2 cells with 10 microM RA markedly increased the ACh-content of these cells, while dopamine levels were unchanged. Depolarisation with potassium (60 mM) enhanced ACh-outflow in the differentiated cells in a Ca(++) dependent way. Also neuropeptides like substance P and NPY were detectable in the undifferentiated NTera-2 cells, while vasointestinal peptide (VIP) could not be found in either precursor or RA differentiated cells. Differentiation was accompanied by a marked reduction of neutral endopeptidase enzyme activity and aminopeptidase activity. From these observations it was concluded that RA induces a cholinergic neurochemical differentiation of this human teratocarcinoma cell line, and that these cells might provide a model system to investigate cholinergic properties of human origin. PMID- 11102654 TI - Orexins: a role in medullary sympathetic outflow. AB - Orexin A and B, also known as hypocretin 1 and 2, are two recently isolated hypothalamic peptides. As orexin-containing neurons are strategically located in the lateral hypothalamus, which has long been suspected to play an important role in feeding behaviors, initial studies were focused on the involvement of orexins in positive food intake and energy metabolism. Recent studies implicate a more diverse biological role of orexins, which can be manifested at different level of the neuraxis. For example, canine narcolepsy, a disorder with close phenotypic similarity to human narcolepsy, is caused by a mutation of hypocretin receptor 2 gene. Results from our immunohistochemical and functional studies, which will be summarized here, suggest that the peptide acting on neurons in the rostral ventrolateral medulla augment sympathoexcitatory outflow to the spinal cord. This finding is discussed in the context of increased sympathetic activity frequently associated with obesity. PMID- 11102655 TI - Dose-response effects of orexin-A on food intake and the behavioural satiety sequence in rats. AB - Although intracerebroventricular (i.c.v.) administration of orexin-A has been reported to stimulate food intake and/or feeding behaviour in rats, mice and goldfish, little attention has thus far been paid to its effects on normal patterns of feeding. In the present study, a continuous monitoring technique was used to characterise the effects of this novel neuropeptide on the microstructure of rat behaviour during a 1-h test with palatable wet mash. Particular attention was devoted to the behavioural satiety sequence, in which feeding is followed by grooming and resting. Although results confirmed the hyperphagic effects of orexin-A (3.33-30.0 microg i.c. v.), gross behavioural analysis failed to reveal any reliable effects of peptide treatment on eating, drinking, sniffing, grooming, resting, locomotion or rearing. However, microstructural analysis revealed behavioural effects of orexin-A that are both dose- and time-dependent. At lower doses (3.33-10.0 microg), orexin-A primarily delayed behavioural satiety, i.e. the normal transition from eating to resting. In contrast, the 30 microg dose initially induced a sedative-like effect, significantly suppressing eating and other active behaviours for the first 15-20 min of the test period. This sedative-like effect resulted in a phase-shifting of the entire behavioural sequence with higher than control levels of eating, grooming, locomotion, rearing and sniffing observed over the second half of the test session. Present findings illustrate the advantages of microstructural behavioural analysis and suggest that the hyperphagic response to low doses of orexin-A results largely from a delay in behavioural satiety while that seen in response to high doses may occur in rebound to initial behavioural suppression. Further studies will be required to confirm the identity of the specific orexin receptors (i.e. OX(1) or OX(2)) involved in mediating the dose-dependent behavioural effects reported. PMID- 11102656 TI - Boning up on tissue engineering. PMID- 11102657 TI - The Achilles' heel of proteomics. PMID- 11102658 TI - An analytical approach to the implementation of genetically modified crops. AB - Public scepticism towards genetically modified (GM) crops is increasing. To address this, the risks and benefits of GM crops must be examined across scientific disciplines, and be discussed with the authorities, the agricultural industry and the consumers. In a feasibility study we have systematically analysed the challenges of the development and marketing of GM crops in Europe. A life-cycle inventory was used together with established technology foresight techniques in an interdisciplinary and empirical framework. The approach taken in this study established a dialogue between stakeholders and provided a framework for discussions about the future direction of GM crops. PMID- 11102659 TI - Interactive visualization and exploration of relationships between biological objects. AB - Genome sequencing and microarray technology produce ever-increasing amounts of complex data that need analysis. Visualization is an effective analytical technique that exploits the ability of the human brain to process large amounts of data. Here, we review traditional visualization methods based on clustering and tree representation, and also describe an alternative approach that involves projecting objects onto a Euclidean space in a way that reflects their structural or functional distances. Data are visualized without preclustering and can be dynamically explored by the user using 'virtual-reality'. We illustrate this approach with two case studies from protein topology and gene expression. PMID- 11102660 TI - Searching for process information in the aroma of cell cultures. AB - Aroma emissions from living cells can provide valuable information about the metabolic and physiological condition of those cells. Electronic noses are chemical gas-sensor arrays that use artificial neural network models to evaluate aromas. They can interpret the complex aroma information emitted from cultures of bacteria, yeast cells and animal cells. Potential applications for electronic noses range from medical diagnosis to industrial bioprocessing. PMID- 11102661 TI - Industrial wastewater bioreactors: sources of novel microorganisms for biotechnology. AB - Microorganisms exist in nature as members of complex, mixed communities. The microbial communities in industrial wastewater bioreactors can be used as model systems to study the evolution of new metabolic pathways in natural ecosystems. The evolution of microbial metabolic capability in these bioreactors is presumably analogous to phenomena that occur in natural ecosystems. The microorganisms in these bioreactors compete for different carbon sources and constantly have to evolve new metabolic capabilities for survival. Thus, industrial bioreactors should be a rich source of novel biocatalysts. PMID- 11102662 TI - Microalgae: a green source of renewable H(2). AB - This article summarizes recent advances in the field of algal hydrogen production. Two fundamental approaches are being developed. One involves the temporal separation of the usually incompatible reactions of O(2) and H(2) production in green algae, and the second involves the use of classical genetics to increase the O(2) tolerance of the reversible hydrogenase enzyme. The economic and environmental impact of a renewable source of H(2) are also discussed. PMID- 11102663 TI - Oscillatory gamma activity in humans: a possible role for object representation. AB - The coherent representation of an object has been suggested to be established by the synchronization in the gamma range (20-100 Hz) of a distributed neural network. So-called '40-Hz' activity in humans could reflect such a mechanism. We have presented here experimental evidence supporting this hypothesis, both in the visual and auditory modalities. However, different types of gamma activity should be distinguished, mainly the evoked 40-Hz response and the induced gamma activities. Only induced gamma activities seem to be related to coherent object representations. In addition, their topography depends on sensory modality and task, which is in line with the idea that they reflect the oscillatory synchronization of task-dependent networks. They can also be functionally and topographically distinguished from the classical evoked potentials and from the alpha rhythm. It was also proposed that the functional role of gamma oscillations is not restricted to object representation established through bottom-up mechanisms of feature binding, but also extends to the cases of internally driven representations and to the maintenance of information in memory. PMID- 11102664 TI - The loci of oscillatory visual-object priming: a combined electroencephalographic and reaction-time study. AB - The detection of reaction-times (RTs) to a target Kanizsa-type square (an illusory square defined by the colinear arrangement of 90 degrees corner junctions) within a matrix of distractor junctions are expedited when the target display is preceded by a 40-Hz flickering display of premask crosses presented prior to, and at the locations subsequently occupied by the junctions of the target display. Priming effects were obtained when four crosses (which together matched the Gestalt arrangement of the target) were presented at the display locations subsequently occupied by the junctions forming the target Kanizsa square (Elliott and Muller, 1998, 2000). The present study was conducted with the aim of replicating the 40-Hz RT priming effects, while simultaneously recording the observers EEG in order to establish the presence and location of Gestalt priming in the brain. The statistical pattern obtained in the RT data corresponded well with previous studies and was matched by the pattern of target P300 latencies across bilateral central and posterior electrodes. Planned analyses focused upon the evoked 40-Hz activity that co-occurs with the P300, revealing a more specific pattern of 40-Hz priming over the visual cortex. A subsequent series of cross-correlational analyses examined the cortical distribution and timing of Gestalt-prime generation during and subsequent to premask-display presentation. Correlations were revealed between stimulus related 40-Hz activity over a range of cortical loci, including the right temporal lobe, which is considered important for figure coding. Taken together, these findings not only support the role of a distributed 40-Hz mechanism during Gestalt-figure priming, but also suggest that patterns of oscillatory brain activity may be directly influenced by, and interpretable in terms of equivalent temporal patterns of stimulus activity. PMID- 11102665 TI - Reversal-rate dependent differences in the EEG gamma-band during multistable visual perception. AB - It is an often reported observation in the literature on multistable perception that the reversal rate within a given observation time is subject to a high interindividual variability. Recently, we reported frontal gamma-band enhancement during multistable visual perception. The aim of the present study was to investigate whether changes in the gamma-band correspond to the variability of the reversal rate. Therefore, a total of 25 observers were divided into two subgroups according to their reversal rate during a 400-s observation period of a reversible pattern based on apparent motion. Subjects with more than 40 reversals within the 400-s were defined as high-rate switchers (HRS). Subjects with a reversal rate below 40 switches were defined as low-rate switchers (LRS). EEG was recorded from frontal, central, parietal, and occipital locations of both hemispheres. The results showed significantly higher gamma activity for the HRS in both phase-locked and non-phase-locked oscillations. Both subgroups showed the highest gamma amplitudes at frontal locations. The results support the involvement of attentional top-down processing in figure reversal. It is concluded that the higher gamma activity for the HRS reflects states of higher arousal, alertness and/or attention according to their fast reversal rate. PMID- 11102666 TI - Alternative perceptual states 'apparent motion' and 'perceived simultaneity' lead to differences of induced EEG rhythms. AB - An investigation of the cortical response (EEG) to periodically presented stimuli producing an ambiguity between long-range apparent motion and flicker is reported. ERPs to stimulus onsets differed slightly between the two percepts, in accordance with the results of Manning et al. (1988), Selmes et al. (1997). Induced rhythms exhibited a strong increase in induced beta and gamma powers at electrode positions T7 and T8 during the perception of apparent motion in two out of 10 participants. In addition, a small overall increase in alpha power at 12-13 Hz and a decrease in delta power below 3.5 Hz during perceived motion were found. The results indicate that a variety of different neural rhythms are involved in the perception of long-range apparent motion. PMID- 11102667 TI - Magnetoencephalographic responses to illusory figures: early evoked gamma is affected by processing of stimulus features. AB - We examined evoked and induced responses in event-related fields and gamma activity in the magnetoencephalogram (MEG) during a visual classification task. The objective was to investigate the effects of target classification and the different levels of discrimination between certain stimulus features. We performed two experiments, which differed only in the subjects' task while the stimuli were identical. In Experiment 1, subjects responded by a button-press to rare Kanizsa squares (targets) among Kanizsa triangles and non-Kanizsa figures (standards). This task requires the processing of both stimulus features (colinearity and number of inducer disks). In Experiment 2, the four stimuli of Experiment 1 were used as standards and the occurrence of an additional stimulus without any feature overlap with the Kanizsa stimuli (a rare and highly salient red fixation cross) had to be detected. Discrimination of colinearity and number of inducer disks was not necessarily required for task performance. We applied a wavelet-based time-frequency analysis to the data and calculated topographical maps of the 40 Hz activity. The early evoked gamma activity (100-200 ms) in Experiment 1 was higher for targets as compared to standards. In Experiment 2, no significant differences were found in the gamma responses to the Kanizsa figures and non-Kanizsa figures. This pattern of results suggests that early evoked gamma activity in response to visual stimuli is affected by the targetness of a stimulus and the need to discriminate between the features of a stimulus. PMID- 11102668 TI - Modulation of induced gamma band activity in the human EEG by attention and visual information processing. AB - Here we present a series of four studies aimed to investigate the link between induced gamma band activity in the human EEG and visual information processing. We demonstrated and validated the modulation of spectral gamma band power by spatial selective visual attention. When subjects attended to a certain stimulus, spectral power was increased as compared to when the same stimulus was ignored. In addition, we showed a shift in spectral gamma band power increase to the contralateral hemisphere when subjects shifted their attention to one visual hemifield. The following study investigated induced gamma band activity and the perception of a Gestalt. Ambiguous rotating figures were used to operationalize the law of good figure (gute Gestalt). We found increased gamma band power at posterior electrode sites when subjects perceived an object. In the last experiment we demonstrated a differential hemispheric gamma band activation when subjects were confronted with emotional pictures. Results of the present experiments in combination with other studies presented in this volume are supportive for the notion that induced gamma band activity in the human EEG is closely related to visual information processing and attentional perceptual mechanisms. PMID- 11102669 TI - Different frequencies for different scales of cortical integration: from local gamma to long range alpha/theta synchronization. AB - Cortical activity and perception are not driven by the external stimulus alone; rather sensory information has to be integrated with various other internal constraints such as expectations, recent memories, planned actions, etc. The question is how large scale integration over many remote and size-varying processes might be performed by the brain. We have conducted a series of EEG recordings during processes thought to involve neuronal assemblies of varying complexity. While local synchronization during visual processing evolved in the gamma frequency range, synchronization between neighboring temporal and parietal cortex during multimodal semantic processing evolved in a lower, the beta1 (12-18 Hz) frequency range, and long range fronto-parietal interactions during working memory retention and mental imagery evolved in the theta and alpha (4-8 Hz, 8-12 Hz) frequency range. Thus, a relationship seems to exist between the extent of functional integration and the synchronization-frequency. In particular, long range interactions in the alpha and theta ranges seem specifically involved in processing of internal mental context, i.e. for top-down processing. We propose that large scale integration is performed by synchronization among neurons and neuronal assemblies evolving in different frequency ranges. PMID- 11102670 TI - Inhibition-based rhythms: experimental and mathematical observations on network dynamics. AB - An increasingly large body of data exists which demonstrates that oscillations of frequency 12-80 Hz are a consequence of, or are inextricably linked to, the behaviour of inhibitory interneurons in the central nervous system. This frequency range covers the EEG bands beta 1 (12-20 Hz), beta 2 (20-30 Hz) and gamma (30-80 Hz). The pharmacological profile of both spontaneous and sensory evoked EEG potentials reveals a very strong influence on these rhythms by drugs which have direct effects on GABA(A) receptor-mediated synaptic transmission (general anaesthetics, sedative/hypnotics) or indirect effects on inhibitory neuronal function (opiates, ketamine). In addition, a number of experimental models of, in particular, gamma-frequency oscillations, have revealed both common denominators for oscillation generation and function, and subtle differences in network dynamics between the different frequency ranges. Powerful computer and mathematical modelling techniques based around both clinical and experimental observations have recently provided invaluable insight into the behaviour of large networks of interconnected neurons. In particular, the mechanistic profile of oscillations generated as an emergent property of such networks, and the mathematical derivation of this complex phenomenon have much to contribute to our understanding of how and why neurons oscillate. This review will provide the reader with a brief outline of the basic properties of inhibition-based oscillations in the CNS by combining research from laboratory models, large-scale neuronal network simulations, and mathematical analysis. PMID- 11102671 TI - Introduction. PMID- 11102672 TI - Gamma activity as a functional correlate of cognition. special issue. PMID- 11102673 TI - Enhanced delivery of AZT to macrophages via acetylated LDL. AB - It is known that infected macrophages play an important role in HIV pathogenesis acting as a reservoir for dissemination of the virus to various organs. Enhanced and/or specific delivery of anti-HIV agents to infected cells is expected to improve their therapeutic index by increasing efficacy and reducing toxicity. Acetylated low density lipoproteins (AcLDL) are known to be taken up by macrophages via scavenger receptors and appear to be good carriers for targeting drug molecules to macrophages. This study investigated the delivery of 3'-azido 3'-deoxythymidine (AZT), an anti-HIV agent, to macrophages using AcLDL. Since the incorporation of AZT into AcLDL was found to be low, several derivatives of AZT including 5'-O-13-oxamyristate-AZT (5'-O-oxaMyr-AZT) have been synthesized as prodrugs. The prodrugs were incorporated into AcLDL using two different methods, namely the contact method and the microemulsion method. Our results demonstrated that the microemulsion method was more effective. The physicochemical properties of the AcLDL/prodrug complex were evaluated by electrophoresis and electron microscopy (EM). Incubation of the complex with plasma resulted in little distribution of the incorporated drug molecules from AcLDL to other components of the plasma, suggesting that the complex was quite stable. Cellular uptake studies using J774.A and U937 demonstrated that AcLDL/prodrug was taken up about 10 times more than AZT. The presence of excess AcLDL was found to inhibit the cellular uptake of AcLDL/5'-O-oxaMyr-AZT by macrophages while excess high density lipoprotein (HDL) or low density lipoprotein (LDL) was found to have little effect, suggesting that the AcLDL/prodrug complex is taken up into macrophages via the scavenger receptor. PMID- 11102674 TI - Two methods for quantifying DNA extracted from poly(lactide-co-glycolide) microspheres. AB - DNA can be formulated with synthetic polymers such as poly(lactide-co-glycolide) (PLG) to generate microparticles. Researchers have used either UV spectroscopy or fluorometry with PicoGreen((R)) dye to quantify PLG-encapsulated DNA. While the sensitivity of DNA detection and quantification by PicoGreen is higher ( approximately 12 pg/ml) compared to UV ( approximately 0.5 microg/ml), each method as an analytical tool has limitations. The premise of this work addresses the usefulness and limitations of each method to determine encapsulation efficiencies in PLG microspheres post-process, and to quantify release of DNA from microspheres during in vitro release experiments. In addition, assay conditions for accurate and reproducible extraction of DNA from PLG microspheres using a biphasic (aqueous/organic) solvent system are described. It was also determined that residual poly(vinyl alcohol) and DNA isoforms (linear, nicked, supercoiled) affected PicoGreen/DNA fluorescence values. PMID- 11102675 TI - Percolation thresholds in ultrasound compacted tablets. AB - Twenty matrix systems with different KCl content (as drug model, from 10 to 90% w/w) and Eudragit RS-PM (as inert excipient) were prepared using an ultrasound assisted press and a traditional eccentric machine. The release behavior from both types of matrices was examined; the kinetic parameters for the release (intrinsic dissolution) and the technological properties of the final tablets (total porosity) were used to estimate the percolation threshold for the drug model and the excipient in both systems. For the systems compacted by ultrasound (US) the estimated value for the excipient percolation threshold ranges from 13.4 to 20.2% v/v (lower than that found for traditional tablets), that agrees with a continuum percolation model suggesting the presence of a continuum phase inside the tablet. This depends on a thermoplastic deformation of Eudragit RS-PM under ultrasound, that destroyed the particulate system of the excipient and transform it into a continuum medium. The percolation threshold for KCl ranged from 58.6 to 61.0% v/v for US and from 26.7 to 42.2% v/v for the traditional tablets. The higher value for ultrasound compacted tablets can be explained by the difficulty of KCl to outcome from a matrix containing insoluble phase that surrounds KCl crystals. PMID- 11102676 TI - Validation protocol of an automated in-line flow-through diffusion equipment for in vitro permeation studies. AB - Transdermal drug delivery experiments are often tedious and time consuming in terms of sampling, labor, etc. In this way, the automation of such experiments has increased in the last few years. A protocol suitable to validate an automated diffusion equipment with seven in-line flow-through cells is described. The proposed protocol was divided into two parts. First, validation of each component which makes up the whole equipment, including the study of the statistical variability of the internal volumes between the cells, the temperature into the different chambers, the time and sample volumes, etc. In the second part, a series of permeation studies were carried out comparing the performance of the system against a classical Franz-type diffusion cell. Ketoprofen was used as a model drug. It was proved the low variability of the replicates obtained with the automated flow-through diffusion cells. The best work conditions as flow rate into the receptor chamber, temperature, etc., as well as the best mathematical approach for the diffusion data, were determined. The advantages in terms of time saved and easiness of validation of the flow-through cell design in comparison to the Franz-type cell were evidenced. PMID- 11102677 TI - Controlled biodistribution of galactosylated liposomes and incorporated probucol in hepatocyte-selective drug targeting. AB - Two types of galactosylated liposomes containing cholesten-5-yloxy-N-(4-((1-imino 2-beta-D-thiogalactosyle thyl)amino)b utyl)formamide (Gal-C4-Chol) as a homing device were prepared to study the biodistribution of liposomal carriers and the incorporated drug. Distearoylphosphatidylcholine (DSPC)/cholesterol (Chol)/Gal-C4 Chol (60:35:5) (Gal DSPC), DSPC/Chol (60:40) (DSPC), egg yolk phosphatidylcholine (eggPC)/Chol/Gal-C4-Chol (60:35:5) (Gal eggPC), and eggPC/Chol (60:40) (eggPC) liposomes labeled with [(3)H]cholesteryl hexadecyl ether (CHE) were tested and [(14)C]probucol, with a partition coefficient between octanol and water (PC(oct)) of 10(10.8), was selected as a model drug with lipophilicity suitable for liposomal incorporation. After intravenous injection of the combination of [(14)C]probucol and [(3)H]liposomes, the liver uptake of [(3)H]CHE was the highest in Gal DSPC liposomes, followed by Gal egg PC liposomes, egg PC liposomes, and DSPC liposomes in that order. [(14)C]Probucol incorporated in Gal DSPC liposomes exhibited lower liver uptake than [(3)H]CHE, suggesting that substantial release from liposomes had taken place. In contrast, [(14)C]probucol incorporated in Gal eggPC liposomes was more stably incorporated under in vivo conditions. Co-administration with galactosylated bovine serum albumin significantly inhibited the liver uptake of [(14)C]probucol in both types of galactosylated liposomes, suggesting that the hepatic uptake of liposomes should be mediated by asialoglycoprotein receptors being [(14)C]probucol incorporated in them. PMID- 11102678 TI - Carbopol/pluronic phase change solutions for ophthalmic drug delivery. AB - The major purpose of this study is to develop and characterize a series of carbopol- and pluronic-based solutions as the in situ gelling vehicles for ophthalmic drug delivery. The rheological properties, in vitro release as well as in vivo pharmacological response of various polymer solutions, including carbopol, pluronic and carbopol/pluronic solution, were evaluated. It was found that the optimum concentration of carbopol solution for the in situ gel forming delivery systems was 0.3% (w/w), and that for pluronic solution was 14% (w/w). The mixture of 0.3% carbopol and 14% pluronic solutions showed a significant enhancement in gel strength in the physiological condition; this gel mixture was also found to be free flowing at pH 4.0 and 25 degrees C. The rheological behaviors of carbopol/pluronic solution were not affected by the incorporation of pilocarpine hydrochloride. Both the in vitro release and in vivo pharmacological studies indicated that the carbopol/pluronic solution had the better ability to retain drug than the carbopol or pluronic solutions alone. The results demonstrated that the carbopol/pluronic mixture can be used as an in situ gelling vehicle to enhance the ocular bioavailability. PMID- 11102679 TI - Macrophage-mediated activation of camptothecin analogue T-2513-carboxymethyl dextran conjugate (T-0128): possible cellular mechanism for antitumor activity. AB - Camptothecin (CPT) analogue T-2513-carboxymethyl (CM) dextran conjugate (T-0128) suppressed human tumor xenografts that were refractory to CPTs. This improvement was explained by its altered pharmacokinetics, but the cellular mechanism of action is still not clear. For this reason, in the present study we examined the determinants of T-0128 action at the cellular level. In vitro tests showed that T 0128 was inactive, indicating that the requirement for its activity lies in the release of linked T-2513, accompanied by the cellular uptake of the conjugate. The accumulation varied between cell lines: tumor cells, including Walker-256 carcinoma and B16 melanoma, showed only a marginal uptake and an undetectable drug release in the medium. In contrast, macrophage-like cells, such as J774.1, internalized T-0128 very efficiently, and liberated T-2513. With regard to the mode of accumulation, fluid-phase pinocytosis seems to be a key factor based on the followings: a similar cell-specificity existed in the uptake of FITC dextran, a marker of fluid-phase pinocytosis. Also, the macrophage uptake of T-0128 increased almost linearly with its medium concentration and was insensitive to dextran sulfate, a ligand for macrophage scavenger receptor. Comparative efficacy studies of T-0128 in the presence and absence of macrophages demonstrated that macrophages increased the efficacy of T-0128. The enhancement could be explained in terms of increases in the amount of released T-2513. Overall, these results lead us to the conclusion that T-0128 acts like a Trojan horse with the help of macrophages: T-0128 is taken up by macrophages in tumor tissues, and the liberated T-2513 kills tumor cells. PMID- 11102680 TI - Determinants for the drug release from T-0128, camptothecin analogue carboxymethyl dextran conjugate. AB - To improve pharmacological profiles of camptothecins (CPTs), a new macromolecular prodrug, denoted T-0128, was synthesized. This prodrug comprises a novel CPT analog (T-2513: 7-ethyl-10-aminopropyloxy-CPT) bound to carboxymethyl (CM) dextran through a Gly-Gly-Gly linker, with a molecular weight of 130 kDa. The present study was designed to elucidate the mechanisms that promote the release of linked T-2513. First, we compared the abilities of a rat liver homogenate, a cocktail of its lysosomal enzymes, and different types of pure enzymes, to liberate T-2513 from the conjugate. The releasing rate in the homogenate was very slow, but was accelerated with the lysosomes. Lysosomal cysteine proteinases, such as cathepsin B, were responsible, coupled with the results of in vitro and in vivo inhibition studies using proteinase inhibitors. The pH optimum for the cathepsin B-mediated drug release was approximately 4. This corresponds to the pH in lysosomes, suggesting lysosomotropic release. Second, to assess the effect of the length and composition of the peptidyl linker, we synthesized the conjugates with a different linker and compared the drug-releasing rates. We found that the insertion of Phe into Gly-Gly-Gly allowed various kinds of enzymes to produce a rapid cleavage, and the Gly-chain lengthening enhanced the lysosome-mediated drug release. The released T-2513 levels in the liver and tumor of the tumor-bearing rats dosed with each conjugate increased with the length of Gly linker, suggesting a good in vitro to in vivo relationship. Comparative efficacy studies of the conjugates with a different linker demonstrated that T-0128 showed the maximum efficacy against MX-1 human mammary xenograft tumors. Thus the Gly-Gly Gly linker exploits lysosomal cathepsin B to liberate T-2513 slowly and steadily, resulting in improved therapeutic efficacy. PMID- 11102681 TI - Drug release from w/o/w emulsions prepared with different chitosan salts and concomitant creaming up. AB - Water-in-oil-in-water (w/o/w) emulsions containing chitosan and tryptophan in the inner aqueous phase were prepared. The acids used to dissolve chitosan were formic, acetic, butyric and lactic acids. The effects of these organic acids and pH of the inner aqueous phase on the release of tryptophan and on the separation of the aqueous phase due mainly to creaming up during storage were studied. When the inner aqueous phase was an acidic chitosan solution, the release rate was almost the same irrespective of the acids mentioned above. When acetic and butyric acids were neutralized with sodium hydroxide, the release of tryptophan was prolonged. However, it was enhanced again by an excess amount of sodium hydroxide. On the other hand, the release was enhanced after neutralization of formic and lactic acids. Separation of the aqueous phase from the w/o/w emulsion during storage was remarkably delayed after neutralizing the chitosan solution containing acetic or butyric acids, while not so much in the cases of formic and lactic acids. It is concluded that these w/o/w emulsions were stabilized in the presence of neutralized chitosan gel suspensions containing acetate or butyrate ions in the inner aqueous phase. PMID- 11102682 TI - NMR characterisation and transdermal drug delivery potential of microemulsion systems. AB - The purpose of this study was to investigate the influence of structure and composition of microemulsions (Labrasol/Plurol Isostearique/isostearylic isostearate/water) on their transdermal delivery potential of a lipophilic (lidocaine) and a hydrophilic model drug (prilocaine hydrochloride), and to compare the drug delivery potential of microemulsions to conventional vehicles. Self-diffusion coefficients determined by pulsed-gradient spin-echo NMR spectroscopy and T(1) relaxation times were used to characterise the microemulsions. Transdermal flux of lidocaine and prilocaine hydrochloride through rat skin was determined in vitro using Franz-type diffusion cells. The formulation constituents enabled a broad variety of microemulsion compositions, which ranged from water-continuous to oil-continuous aggregates over possible bicontinuous structures, with excellent solubility properties for both lipophilic and hydrophilic compounds. The microemulsions increased transdermal flux of lidocaine up to four times compared to a conventional oil-in-water emulsion, and that of prilocaine hydrochloride almost 10 times compared to a hydrogel. A correlation between self-diffusion of the drugs in the vehicles and transdermal flux was indicated. The increased transdermal drug delivery from microemulsion formulations was found to be due mainly to the increased solubility of drugs and appeared to be dependent on the drug mobility in the individual vehicle. The microemulsions did not perturb the skin barrier, indicating a low skin irritancy. PMID- 11102683 TI - Protein encapsulation into biodegradable microspheres by a novel S/O/W emulsion method using poly(ethylene glycol) as a protein micronization adjuvant. AB - A new method for preparing protein-loaded biodegradable microspheres by a process involving solid-in-oil-in-water (S/O/W) emulsion was established using poly(ethylene glycol) (PEG). In the first step, a protein solution was lyophilized with PEG, which resulted in the formation of spherical protein microparticles, less than 5 microm in diameter, dispersed in a continuous PEG phase. This process was well explained by the aqueous phase separation phenomenon induced by freezing-condensation. Since this lyophilizate could be directly dispersed in an organic phase containing biodegradable polymer by dissolving PEG with methylene chloride, a conventional in-water drying method could be adopted in the second step. Through this S/O/W emulsion process, horseradish peroxidase was effectively entrapped into monolithic-type microspheres of poly(DL-lactic-co glycolic acid) (PLGA), without significant loss of activity. Bovine superoxide dismutase (bSOD), as another model protein, could be encapsulated into reservoir type microspheres by the 'polymer-alloys method' using both poly(DL-lactic acid) (PLA) and PLGA. The initial release of bSOD from this reservoir-type microsphere was efficiently reduced. Further, the bSOD release kinetics could be suitably modified by adjusting the loading amounts of PEG or polymer composition. In this study, the multi-functional nature of PEG was successfully utilized in the preparation and designing of protein-loaded microspheres. PMID- 11102684 TI - Biodegradable microparticles as a two-drug controlled release formulation: a potential treatment of inflammatory bowel disease. AB - A multiple unit dosage form for oral delivery based on the microencapsulation of anti-inflammatory drugs using different biodegradable polymers, poly(epsilon caprolactone), polylactic acid and poly(lactic-co-glycolic acid), prepared either by the water-in-oil-in-water (w/o/w) or the solid-in-oil-in-water (s/o/w) solvent evaporation method was developed. Microparticles were characterized for their size, morphology, encapsulation efficiency and drug release. The physical state of drugs and polymers was determined by differential scanning calorimetry (DSC), imaging of the particles was performed by scanning electron microscopy and confocal laser scanning microscopy. Sulfasalazine and betamethasone used for the treatment of inflammatory bowel disease, were chosen as model drugs. The microparticles were spherical with diameters in the range of 91 to 258 microm by the w/o/w-method, and in the range of 102 to 277 microm by the s/o/w-method. The encapsulation efficiency (EE) varied between 11 and 16% for sulfasalazine and 50 and 67% for betamethasone with the w/o/w-method, and between 73 and 79% for sulfasalazine and 60 and 70% for betamethasone with the s/o/w-method. DSC showed no interaction between polymers and drugs, while the drugs were dispersed in the polymer. In vitro release studies showed a controlled release of sulfasalazine and betamethasone from microparticles prepared by the s/o/w-method; a pronounced burst release of sulfasalazine was observed from microparticles prepared by the w/o/w-method. PMID- 11102685 TI - Bimodal drug release achieved with multi-layer matrix tablets: transport mechanisms and device design. AB - The aim of this study was to develop new multi-layer matrix tablets to achieve bimodal drug release profiles (fast release/slow release/fast release). Hydroxypropyl methylcellulose acetate succinate (HPMCAS, type MF) was chosen as a matrix former, because it is water-insoluble at low, and water-soluble at high pH values. Studies focused on the elucidation of the drug release mechanisms from HPMCAS-MF:drug tablets. In 0.1 N HCl the resulting release kinetics can be described using Fick's second law of diffusion, taking into account axial and radial mass transfer in cylindrical geometry. As the diffusion coefficients are found to be constant and the boundary conditions to be stationary, these systems are purely drug diffusion-controlled. In contrast, the dominating mass transport phenomena in phosphate buffer pH 7.4 are more complex. Due to polymer dissolution the resulting matrix structure is time-variant, leading to increasing drug diffusion coefficients and decreasing tablet dimensions, and thus moving boundary conditions. Drug release is affected by water imbibition, drug diffusion and polymer dissolution and is faster compared to 0.1 N HCl. With knowledge of these underlying release mechanisms, multi-layer matrix tablets were developed to achieve bimodal drug release. HPMCAS-MF:drug mixtures were used as tablet cores. As expected, changing the release medium from 0.1 N HCl to phosphate buffer pH 7. 4 after 2 h, lead to a significant increase in drug release. The abruptness of this rate change could be enhanced by adding two drug-free HPMCAS-MF barrier layers (one on each side) to the system. The addition of a fourth, drug containing and fast disintegrating initial dose layer yielded the desired bimodal drug release patterns. The process and formulation parameters affecting the resulting release rates were investigated using theophylline and acetaminophen as model drugs. PMID- 11102686 TI - A bioluminescent derivative of Pseudomonas putida KT2440 for deliberate release into the environment. AB - Recombinant derivatives of Pseudomonas putida strain KT2440 are of potential interest as microbial inoculants to be deliberately released for agricultural applications. To facilitate tracking of this strain and its derivatives after introduction into the environment, a mini-Tn5-'luxAB transposon was introduced into the chromosome of P. putida KT2440, yielding strain P. putida S1B1. Sequencing of the DNA region located upstream of the 'luxAB genes and similarity search with the P. putida KT2440 genome sequence, localized the transposon within a 3021-bp open reading frame (ORF), whose translated sequence showed significant similarity with the hypothetical YdiJ proteins from Escherichia coli and Haemophilus influenzae. A second ORF adjacent to and divergent from the ydiJ sequence was also found and showed significant homology with various LysR-type transcriptional activator proteins from several bacteria. Disruption of the ydiJ locus in P. putida S1B1 did not affect the survival of the strain in unvegetated or vegetated soils. Bioluminescent detection of P. putida S1B1 cells enriched in selective media directly from soil allowed detection of culturable cells in soil samples over a period of at least 8 months. The addition of the luxAB biomarker facilitates tracking in the root system of several plant species grown under sterile and non-sterile conditions. The correlation of the bioluminescent phenotype with the growth activity of P. putida S1B1 cells colonizing the root system of barley and corn plants was estimated by monitoring ribosomal contents using quantitative hybridization with fluorescence-labeled ribosomal RNA probes. A correlation between inoculum density, light output, and ribosomal contents was found for P. putida cells colonizing the root system of barley seedlings grown under sterile conditions. Although ribosomal contents, and therefore growth activity, of P. putida S1B1 cells extracted from the rhizosphere of corn plants grown in non-sterile soil were similar to those found in starved cells, the luminescent system permitted non-destructive in situ detection of the strain in the upper root system. PMID- 11102687 TI - Identification and characterization of humic substances-degrading bacterial isolates from an estuarine environment. AB - Bacterial isolates were obtained from enrichment cultures containing humic substances extracted from estuarine water using an XAD-8 resin. Eighteen isolates were chosen for phylogenetic and physiological characterization based on numerical importance in serial dilutions of the enrichment culture and unique colony morphology. Partial sequences of the 16S rRNA genes indicated that six of the isolates were associated with the alpha subclass of Proteobacteria, three with the gamma-Proteobacteria, and nine with the Gram-positive bacteria. Ten isolates degraded at least one (and up to six) selected aromatic single-ring compounds. Six isolates showed ability to degrade [(14)C]humic substances derived from the dominant salt marsh grass in the estuary from which they were isolated (Spartina alterniflora), mineralizing 0.4-1.1% of the humic substances over 4 weeks. A mixture of all 18 isolates did not degrade humic substances significantly faster than any of the individual strains, however, and no isolate degraded humic substances to the same extent as the natural marine bacterial community (3.0%). Similar studies with a radiolabeled synthetic lignin ([beta (14)C]dehydropolymerisate) showed measurable levels of degradation by all 18 bacteria (3.0-8.8% in 4 weeks), but mineralization levels were again lower than that observed for the natural marine bacterial community (28.2%). Metabolic capabilities of the 18 isolates were highly variable and generally did not map to phylogenetic affiliation. PMID- 11102688 TI - 5S rRNA fingerprints of marine bacteria, halophilic archaea and natural prokaryotic assemblages along a salinity gradient. AB - Natural prokaryotic assemblages from two multi-pond solar salterns and pure cultures of both marine bacteria and halophilic archaea were analyzed and compared by electrophoretic analysis of 5S rRNAs. A salinity gradient from seawater (3.7%) to NaCl precipitation (37%) was studied. The culture-independent, PCR-free, fingerprinting analysis covered two objectives: (i) to compare natural assemblages among them and with results previously obtained through a PCR dependent approach and (ii) to estimate the in situ relevance of those prokaryotic groups obtained with classical culture methodologies. Natural assemblages were analyzed through cluster analysis of quantitative 5S rRNA band patterns. The resulting groups were in accordance with environmental parameters (i.e., NaCl concentration) and with the clustering obtained after a PCR-dependent approach, showing the formation of three salinity-based groups of samples (<10%, 10-25% and >25% salinity). Similarities between the laboratory strains tested and dominant community members were studied by comparing 5S rRNA band patterns. The lack of match obtained after cluster analysis indicated that the prokaryotic populations relevant in the ponds below 25% salinity were neither Flavobacteria nor haloarchaeal strains belonging to the genera Halococcus, Haloarcula and Halobacterium. Members of Proteobacteria and Gram-positive bacteria were found to match bands in these samples. The 5S rRNA fingerprint from the dominant community members in the ponds above 30% salinity did not fit any of the cultured halophilic archaea studied, in agreement with earlier PCR results. This is consistent with a greater bias introduced by culture-dependent methods than by those based on PCR, especially for archaeal populations. PMID- 11102689 TI - 16S rDNA analysis for characterization of denitrifying bacteria isolated from three agricultural soils. AB - Bacteria capable of denitrification are spread among phylogenetically diverse groups. In the present investigation, molecular methods (amplified ribosomal DNA restriction analysis (ARDRA) and partial 16S rDNA gene sequencing) were used to determine the genetic diversity of culturable denitrifying soil bacteria. The purpose of this work was to study the microbial density and diversity of denitrifying communities isolated from two luvisols and a rendosol. The denitrifying bacterial density was significantly higher in the two luvisols (3x10(6) and 4x10(6) bacteria g(-1) dry soil) than in the rendosol (4x10(5) bacteria g(-1) dry soil). Denitrifying isolates from soils were grouped according to the similarity of their restriction patterns into 26 ARDRA types. Interestingly ARDRA analysis suggests that some denitrifying isolates are specific to a soil type while others seem to be geographically widespread. The number of individual isolates found in each ARDRA type appeared to be highly variable between the two sampling dates but some denitrifying types were capable of persisting in soil. The tree obtained from the partial sequences revealed five major branches exhibiting highest identity to the following genera: (i) Burkholderia-Ralstonia, (ii) Pseudomonas, (iii) Xanthomonas-Frateuria, (iv) Bacillus and (v) Streptomyces. Our 16S rDNA-based analysis clearly reveals broad diversity exceeding that previously described in the literature. PMID- 11102690 TI - Isolation of new fastidious alpha Proteobacteria and Afipia felis from hospital water supplies by direct plating and amoebal co-culture procedures. AB - As water is a source of nosocomial infections in hospitals, the presence of fastidious Gram-negative bacteria in water samples taken in a university hospital was investigated. Water samples were inoculated onto agar plates and into amoebal microplates for co-culture. Sixty-eight alpha proteobacteria isolates were obtained and characterized using phenotypic methods and 16S rRNA gene sequence comparison. The latter approach divided the strains into seven clusters. Of these, one corresponded to previously recognized Afipia felis and it is likely that six were closely related new species. As these bacteria are fastidious and can not be cultivated on standard microbiological media, their possible role in hospital-acquired human infections should be investigated. PMID- 11102691 TI - Cloning and characterization of two repeated sequences in the symbiotic fungus Tuber melanosporum Vitt. AB - Two repeated DNA sequences of European strains of the symbiotic fungus Tuber melanosporum were isolated and characterized. One of these, SS14, representing about 0.05% of the fungal genome, was shown to be a T. melanosporum-specific sequence by Southern and dot-blot hybridization. The second one, named SS15, is about 0.0025% of the entire genome, and it is specific not only to T. melanosporum but also to the Asian black truffle Tuber indicum. Neither of these two fragments hybridizes with any of the other European truffle species tested. By sequence analysis of these two fragments, PCR primers were designed and used to selectively amplify DNA from T. melanosporum ascocarps and ectomycorrhizae by simple and multiplex PCR. No amplification products were obtained with DNA from either mycorrhizal roots or fruit bodies of other ectosymbiotic fungi. The two identified genomic traits also provided useful information for a better understanding of the phylogenetic relationships among black truffle species and for testing T. melanosporum intraspecific variability. PMID- 11102692 TI - Screening of sulfate-reducing bacteria in colonoscopy samples from healthy and colitic human gut mucosa. AB - A PCR-based approach combined with microbiological cultivation methods was employed to determine the occurrence of sulfate-reducing bacteria (SRB) in colon biopsy samples from ulcerative colitis patients and from non-colitic controls. The detection of mucosa-associated SRB was carried out by digoxigenin-dUTP labelled PCR amplification, in liquid Postgate medium B and in a new liquid medium, termed VM medium I. Using Postgate medium B, the growth of SRB was confirmed in 92% of the colitic specimens and in 52% of non-colitic samples. However, PCR analysis and incubation in VM medium I detected SRB in 100% of biopsy material indicating ubiquitous presence of SRB in human colon mucosa. PMID- 11102693 TI - Carbon monoxide production is not enhanced by nitrogenase activity. AB - A diverse group of nitrogen-fixing bacteria and two heme degraders were grown with and without fixed nitrogen sources under oxic and suboxic conditions, with and without addition of heme-containing compounds. Several of the strains produced carbon monoxide (CO) under one or more of these conditions, but nitrogenase activity did not stimulate rates of production relative to controls. Although nitrogenase can reduce CO(2) to CO in vitro in the absence of N(2), this process likely contributes minimally to CO production in soils under in situ conditions. In contrast, myoglobin or hematin addition under oxic conditions significantly stimulated CO production by the heme degraders. However, estimates of CO production from microbial heme turnover suggest that this too is likely to be only a small source of CO in soils in situ. PMID- 11102694 TI - Competition between methanogenesis and quinone respiration for ecologically important substrates in anaerobic consortia. AB - Anaerobic consortia obtained from a wide variety of environments were tested for oxidizing several ecologically significant substrates with the humic model compound, anthraquinone-2,6-disulfonate (AQDS), as terminal electron acceptor. All the substrates, including hydrogen, acetate, propionate, methanol and lactate, were completely or partially converted to methane when bicarbonate was the only electron acceptor available. Addition of AQDS (20 mM) to the cultures prevented methanogenesis in most cases and AQDS reduction became the preferred pathway. AQDS was shown to be toxic for methanogenesis and this effect played an important role in enabling quinone-respiring bacteria to outcompete methanogens. Furthermore, AQDS respiration is thermodynamically more favorable than methanogenesis. All the consortia evaluated were capable of oxidizing hydrogen linked to the reduction of AQDS. Most inocula tested were also able to oxidize acetate and lactate in the same way. When methanol was provided as an electron donor competition between methanogenesis and acetogenesis occurred. Acetate accumulated from the latter process was responsible for quinone respiration. These results suggest that quinone-respiring bacteria are ubiquitous and that quinones in humus may significantly contribute to carbon cycling process by serving as a terminal electron acceptor for the anaerobic microbial oxidation of a wide variety of ecologically important substrates. PMID- 11102695 TI - Genetic diversity of carbofuran-degrading soil bacteria. AB - The genetic diversity of 128 carbofuran-degrading bacteria was determined by ARDRA (amplified ribosomal DNA restriction analysis) of 16S rDNA and restriction fragment length polymorphism analysis of the 16S-23S rDNA spacer region (IGS) using five endonucleases. The isolates were distributed in 26 distinct ARDRA groups and 45 IGS types revealing a high level of microbial diversity confirmed by ARDRA clustering and sequencing of 16S rDNA. The occurrence of a methylcarbamate-degrading gene (mcd) was monitored by polymerase chain reaction amplification using specific primers. The mcd gene was detected only in 58 bacteria and there was no clear relationship between the presence of this gene and the phylogenetic position of the strain. PMID- 11102696 TI - A new study challenges the current belief of a high human male:female mutation ratio. AB - A recent comparison of a DNA region that was transposed from the X to the Y chromosome 3-4 million years ago, with the same region on the X chromosome showed only a slight excess of mutant changes on the Y chromosome. This translates to an estimate of 1.7 for the ratio of the male to female mutation rate, much less than the average 5.1 of previous studies. The authors argue that this throws doubt not only on higher male mutation rates in human ancestry, but also on the standard assumption of a high male:female ratio in contemporary human populations. Clearly, more studies are needed to clear up this discrepancy in the ancestral rates, but I believe that the high contemporary male:female ratio for base substitutions is too well established to be overthrown by even a very good evolutionary study. PMID- 11102697 TI - Detecting the form of selection from DNA sequence data. AB - Clues to our evolutionary history lie hidden within DNA sequence data. One of the great challenges facing population geneticists is to identify and accurately interpret these clues. This task is made especially difficult by the fact that many different evolutionary processes can lead to similar observations. For example, low levels of polymorphism within a region can be explained by a low local mutation rate, by selection having eliminated deleterious mutations, or by the recent spread to fixation of a beneficial allele. Theoretical advances improve our ability to distinguish signals left by different evolutionary processes. In particular, a new test might better detect the footprint of selection having favored the spread of a beneficial allele. PMID- 11102698 TI - The evolutionary history of ribosomal protein RpS14: horizontal gene transfer at the heart of the ribosome. PMID- 11102699 TI - Mining archaeal proteomes for eukaryotic proteins with novel functions: the PACE case. PMID- 11102700 TI - Author correction. Adele De arcangelis and elisabeth georges-labouesse (2000) integrin and ECM functions: roles in vertebrate development. Trends genet. 16, 389-395 PMID- 11102701 TI - Crossing over a boundary PMID- 11102702 TI - Prion-mediated diversity in yeast PMID- 11102703 TI - The link between DNA synthesis and asymmetric division PMID- 11102704 TI - Form of the worm: genetics of epidermal morphogenesis in C. elegans. AB - The development of the epidermis of the nematode worm Caenorhabditis elegans illustrates many common processes of epithelial morphogenesis. In the worm, these morphogenetic movements have been described with single-cell resolution, and the roles of individual cells have been probed in laser killing experiments. Genetic dissection is yielding insights into the molecular mechanisms of these complex morphogenetic processes. PMID- 11102705 TI - Microsatellite mutations in the germline: implications for evolutionary inference. AB - Microsatellite DNA sequences mutate at rates several orders of magnitude higher than that of the bulk of DNA. Such high rates mean that spontaneous mutations that form new-length variants can realistically be seen in pedigree analysis. Data on observed mutation events from various organisms are now accumulating, allowing inferences on DNA sequence evolution to be made through an unusually direct approach. Here I discuss and integrate microsatellite mutation data in an evolutionary context. A striking feature of the mutation process is that it seems highly heterogeneous, with distinct differences between species, repeat types, loci and alleles. Age and sex also affect the mutation rate. Within genomes at equilibrium, the microsatellite-length distribution is a delicate balance between biased mutation processes and point mutations acting towards the decay of repetitive DNA. Indeed, simple repeats do not evolve simply. PMID- 11102706 TI - Regulation of the L-arabinose operon of Escherichia coli. AB - Over forty years of research on the L-arabinose operon of Escherichia coli have provided insights into the mechanism of positive regulation of gene activity. This research also discovered DNA looping and the mechanism by which the regulatory protein changes its DNA-binding properties in response to the presence of arabinose. As is frequently seen in focused research on biological subjects, the initial studies were primarily genetic. Subsequently, the genetic approaches were augmented by physiological and then biochemical studies. Now biophysical studies are being conducted at the atomic level, but genetics still has a crucial role in the study of this system. PMID- 11102707 TI - Towards an understanding of the genetics of human male infertility: lessons from flies. AB - It has been argued that about 4-5% of male adults suffer from infertility due to a genetic causation. From studies in the fruitfly Drosophila, there is evidence that up to 1500 recessive genes contribute to male fertility in that species. Here we suggest that the control of human male fertility is of at least comparable genetic complexity. However, because of small family size, conventional positional cloning methods for identifying human genes will have little impact on the dissection of male infertility. A critical selection of well defined infertility phenotypes in model organisms, combined with identification of the genes involved and their orthologues in man, might reveal the genes that contribute to human male infertility. PMID- 11102708 TI - Natural selection and the function of genome imprinting: beyond the silenced minority. AB - Most hypotheses of the evolutionary origin of genome imprinting assume that the biochemical character on which natural selection has operated is the expression of the allele from only one parent at an affected locus. We propose an alternative - that natural selection has operated on differences in the chromatin structure of maternal and paternal chromosomes to facilitate pairing during meiosis and to maintain the distinction between homologues during DNA repair and recombination in both meiotic and mitotic cells. Maintenance of differences in chromatin structure in somatic cells can sometimes result in the transcription of only one allele at a locus. This pattern of transcription might be selected, in some instances, for reasons that are unrelated to the original establishment of the imprint. Differences in the chromatin structure of homologous chromosomes might facilitate pairing and recombination during meiosis, but some such differences could also result in non-random segregation of chromosomes, leading to parental-origin-dependent transmission ratio distortion. This hypothesis unites two broad classes of parental origin effects under a single selective force and identifies a single substrate through which Mendel's first and second laws might be violated. PMID- 11102724 TI - Neurological bases of behavioral development in infancy. AB - This article presents selected psychological competences that emerge in children during the first 2 years, together with correlated structural, biochemical and physiological changes in the brain. Major behavioral events of the 1st year are the disappearance of the neonatal reflexes, the improvement of recognition and working memory and the appearance of the universal fears of strangers and of separation from the caretaker. These behaviors are correlated in time with changes in the brain that allow the increased ability of the cortex to inhibit brainstem reflexes, with processes in the prefrontal cortex and hippocampus that facilitate the formation, storage and retrieval of memories, and with strengthened connections between the cortex and limbic system. Behavioral events of the 2nd year include the explosion of language and the emergence of self awareness, both of which depend on the capacity for inference. The emergence of these capabilities is correlated with the intensified connectivity of the two hemispheres, the maturation of the prefrontal cortex and cortical-subcortical network. PMID- 11102725 TI - Assessment of brainstem function in Chiari II malformation utilizing brainstem auditory evoked potentials (BAEP), blink reflex and masseter reflex. AB - Brainstem dysfunction was evaluated in 67 patients with myelomeningocele and Chiari II malformation using brainstem auditory evoked potentials (BAEP), blink reflex (BR) and masseter reflex (MR). Signs and symptoms related to Chiari II malformation were observed in 18 patients while 49 patients had normal brainstem findings. BAEP and BR showed a higher sensitivity of brainstem involvement than MR (BAEP=1.0, BR=0.83, MR=0.50). BR, and in particular, MR were of higher accuracy (BR=0.52, MR=0.72) than BAEP (0.39) in separating patients with brainstem signs and symptoms related to Chiari II malformation. We feel that this is due to anatomic and physiologic peculiarities of the brainstem structures mediating BR and MR. Our results suggest that brainstem reflexes can support the decision of further treatment. PMID- 11102726 TI - Cognitive and neurophysiological evaluation of Japanese dyslexia. AB - Seven Japanese dyslexic boys were evaluated as to their pedagogic performance on the pupil rating scale (PRS), and psychological and neurophysiological characteristics. One of them suffered from severe English dyslexia despite that his Japanese dyslexia was feeble. PRS did not successfully reveal their reading difficulties. Psychological examination (WISC-R and K-ABC) revealed their cognitive dysfunction, but the results were heterogeneous. The Token test was most useful for detecting their poor reading comprehension. Electroencephalogram (EEG) coherence analysis showed high inter- and intra-hemispheric values. These findings may imply hyperconnectivity of the cerebral white matter in dyslexia. We assumed that the Token test demonstrates the discrepancy between reading and hearing comprehension best of all among these psychological tests and that connectivity between non-functional cortical lesions remains in dyslexic children. PMID- 11102727 TI - Cerebrospinal fluid purine metabolite and neuron-specific enolase concentrations after febrile seizures. AB - If febrile seizures cause significant compromise of neuronal metabolism (whether permanent or reversible), this should be reflected in an increase in the cerebrospinal fluid concentrations of neuron-specific enolase (NSE) and/or adenosine triphosphate (ATP) breakdown products. In the present study, AMP, IMP, inosine, adenosine, guanosine, adenine, guanine, hypoxanthine, xanthine, uric acid and NSE concentrations were determined in the cerebrospinal fluid of 90 children 1 h after febrile seizure (73 simple febrile seizures (SFS); 17 complex febrile seizures (CFS)), and in a control group of 160 children. There was no statistically significant difference between the SFS group and the control group for any of the substances determined, suggesting that SFS neither significantly depletes neuronal ATP concentration, nor significantly increases NSE concentration; thus, SFS do not appear to constitute a threat to neuronal integrity. However, patients with CFS showed significantly lower IMP concentrations and significantly higher adenine concentrations than controls, and significantly higher AMP concentrations than SFS patients; these results suggest that CFS may affect energy metabolism in the brain. However, NSE concentrations were normal in the cerebrospinal fluid of both SFS and CFS patients, suggesting that neither type of seizure causes significant neuronal damage, at least early after the seizure. PMID- 11102728 TI - Co-segregation of benign infantile convulsions and paroxysmal kinesigenic choreoathetosis. AB - We report seven families and two sporadic cases in which benign infantile convulsions and paroxysmal kinesigenic choreoathetosis were co-segregated. Clinical investigations included physical and neurological examinations, blood electrolyte values, interictal and ictal electroencephalograms, and computed tomography or magnetic resonance imaging of the brain. The family pedigree was confirmed and the clinical history of the relatives was obtained. Seventeen individuals developed infantile convulsions followed by paroxysmal dyskinesias during childhood or adolescence. Six had only infantile convulsions, and two had only paroxysmal dyskinesias. The seizures never persisted into childhood or recurred in adulthood. The seizure type was a complex partial seizure, with or without secondary generalization, in nine of 14 patients. Paroxysmal dyskinesias, a subgroup of paroxysmal kinesigenic choreoathetosis, occurred for less than 5 min. The attacks of dyskinesias began at age 5-12 years in most patients, and tended to remit in adulthood. The mode of inheritance was apparently autosomal dominant in four of the families (17 affected individuals), who were diagnosed with ICCA syndrome (infantile convulsions and paroxysmal choreoathetosis). However, the condition occurred only among siblings in three families (six patients), and sporadically in two patients, suggesting genetic heterogeneity in this distinct co-segregation. PMID- 11102729 TI - Developmental sequence of postural control in prone position in children with spastic diplegia. AB - The aim of this study was to assess the development of postural control in the prone position in children with spastic diplegia and triplegia, and determine the influence of clinical characteristics, visual acuity and cognitive performance on that development. We also analysed the relation between these early motor achievements in the prone position and the subsequent acquisition of motor competence in the sitting position. We followed 24 diplegic and triplegic children from before age 2 years (mean age 12 months) to mean age 41 months, videorecording motor behaviour every six months and abstracting acquisitions in alignment and balance using a standardised procedure. We confirm a developmental sequence of all the acquired movements in the prone position. 83.3% of the children completed the uprighting sequence in the sagittal plane, acquired good balance, and ability to rotate the head and trunk. 70.8% of the children (all but one of the diplegic children and none among triplegic children) acquired symmetric posture in the frontal plane and 83.3% reduced leg hyperextension. Development was not uniform, and at 12-18 months two groups began to emerge: diplegic children who rapidly achieved all or most of the steps in the sequence and had a favourable prognosis for subsequent motor development; and triplegic children who achieved these steps at a much slower rate or in some cases not at all and had a less favourable prognosis for future development. Diplegic children with normal visual acuity, and general quotient GQ>70 did better than triplegic children with compromised visual acuity and GQ<70. Acquisition of the full uprighting sequence in the prone position before the age of two related to the later acquisition of autonomous sitting. PMID- 11102730 TI - Complex regional pain syndrome in childhood: report of three cases. AB - We describe three patients with the limb pain of complex regional pain syndrome (CRPS) in childhood with autonomic nervous system function involvement. Their autonomic nerve abnormality was non-invasively examined by means of laser doppler flowmetry (LDF) and a sympathetic skin response (SSR) test. In one it was resolved with physiotherapy, but the others needed epidural anesthesia for pain control. Though CRPS used to be recognized as a refractory disorder in adults, childhood cases have been found in recent years, generally having a better prognosis than adult ones. However, even in the children, the prognosis or responses to the same therapy vary, and there are progressive and refractory cases. CRPS should be considered as a differential diagnosis of unexplained persistent limb pain even in childhood for early and appropriate management. PMID- 11102731 TI - Infantile convulsions and paroxysmal kinesigenic choreoathetosis in a patient with idiopathic hypoparathyroidism. AB - We reported a 15-year-old boy with idiopathic hypoparathyroidism who presented with paroxysmal kinesigenic choreoathetosis at age 10. Calcium levels were low and intact parathyroid hormones were undetectable in serum. Computed tomography showed calcifications in the basal ganglia, thalamus, and cerebral white matter. He had a history of infantile convulsions with a benign outcome. The convulsions occurred in clusters at age 2.5 months, but they never recurred. This patient's clinical features were phenotypically indistinguishable from those of infantile convulsions and choreoathetosis (ICCA) syndrome PMID- 11102732 TI - Diagnostic usefulness of diffusion-weighted magnetic resonance imaging in influenza-associated acute encephalopathy or encephalitis. AB - A magnetic resonance imaging (MRI) study was performed for a 20-month-old girl with an influenza type A infection who presented acute encephalopathy. Conventional MRI performed 8 days after the onset of encephalopathy, including T1 weighted, T2-weighted, and fluid-attenuated inversion recovery imaging, revealed only vague lesions in the right frontal, temporal, and parietal lobes. In contrast, diffusion-weighted imaging (DWI) then demonstrated the lesions much more intensively. On the 26th day, the lesions previously observed on DWI had become less discernible. The hyperintensity observed on DWI might reflect cytotoxic edema. Thus, DWI may be useful for evaluation of acute influenzal encephalopathy/encephalitis. PMID- 11102733 TI - A case of Walker-Warburg syndrome. AB - Walker-Warburg syndrome (WWS) is an autosomal recessive disorder characterized by type II lissencephaly, cerebellar and retinal anomalies, and congenital muscular dystrophy. We report a female diagnosed with WWS based on clinical criteria. This patient was found to have fetal hydrocephalus on ultrasonography at 29 weeks of gestation, and exhibited severe hypotonia, ocular malformations, and hydrocephalus at birth. MRI revealed type II lissencephaly, hydrocephalus, and other severe brain malformations. Genetic analysis was performed to distinguish WWS from severe Fukuyama-type congenital muscular dystrophy (FCMD), which has numerous findings in common. This revealed no expression of the founder haplotype or single-stranded conformation polymorphism (SSCP) abnormalities. Since the life expectancy of patients with FCMD is longer, differential diagnosis should be performed precisely. PMID- 11102734 TI - Preface. PMID- 11102735 TI - In vitro and in vivo investigations on fluoroquinolones; effects of the P glycoprotein efflux transporter on brain distribution of sparfloxacin. AB - The role of mdr1a-encoded P-glycoprotein on transport of several fluoroquinolones across the blood-brain barrier was investigated. In vitro, P-glycoprotein substrates were selected by using a confluent monolayer of MDR1-LLC-PK1 cells. The inhibition of fluoroquinolones (100 microM) on transport of rhodamine-123 (1 microM) was compared with P-glycoprotein inhibitors verapamil (20 microM) and SDZ PSC 833 (2 microM). Subsequently, transport polarity of fluoroquinolones was studied. Sparfloxacin showed the strongest inhibition (26%) and a large polarity in transport, by P-glycoprotein activity. In vivo, using mdr1a (-/-) and wild type mice, brain distribution of pefloxacin, norfloxacin, ciprofloxacin, fleroxacin and sparfloxacin was determined at 2, 4, and 6 h following intra arterial infusion (50 nmol/min). Brain distribution of sparfloxacin was clearly higher in mdr1a (-/-) mice compared with wild-type mice. Sparfloxacin was infused (50 nmol/min) for 1, 2, 3 and 4 h in which intracerebral microdialysis was performed. At 4 h, in vivo recovery (dynamic-no-net-flux method) was 6.5+/-2.2 and 1.5+/-0.5%; brain(ECF) concentrations were 5.1+/-0.2 and 26+/-21 microM; and total brain concentrations were 7.2+/-0.3 and 23+/-0.3 microM in wild-type and mdr1a (-/-) mice, respectively. Plasma concentrations were similar (18.4+/-0.7 and 17.9+/-0.5 microM, respectively). In conclusion, sparfloxacin enters the brain poorly mainly because of P-glycoprotein activity at the blood-brain barrier. PMID- 11102736 TI - Relationship between permeability status of the blood-brain barrier and in vitro permeability coefficient of a drug. AB - OBJECTIVE: The aim was to test the hypothesis that the assessment of basal and drug-induced changes in permeability of the blood-brain barrier (BBB) during in vitro drug transport assays is essential for an accurate estimation of the permeability coefficient of a drug. METHODS: An in vitro BBB model was used, comprising of brain capillary endothelial cells (BCEC) and astrocytes co-cultured on semi-permeable filter inserts. Experiments were performed under control and challenged experimental circumstances, induced to simulate drug effects. The apparent BBB permeability coefficient for two markers for paracellular drug transport, sodium fluorescein (P(app,FLU), M(w) 376 Da) and FITC-labeled dextran (P(app,FD4), M(w) 4 kDa), was determined. Transendothelial electrical resistance (TEER) was used to quantify basal and (simulated) drug-induced changes in permeability of the in vitro BBB. The relationship between P(app) and TEER was determined. Drug effects were simulated by exposure to physiologically active endogenous and exogenous substances (i.e., histamine, deferroxamine mesylate, adrenaline, noradrenaline, bradykinin, vinblastine, sodium nitroprusside and lipopolysaccharide). RESULTS: P(app,FLU) and P(app,FD4) in control experiments varied from 1.6 up to 17.6 (10(-6)cm/s) and 0.3 up to 7. 3 (10(-6)cm/s), respectively; while for individual filters P(app, FLU) was 4 times higher than P(app,FD4) (R(2)=0.97). As long as TEER remained above 131.Omega cm(2) for FLU or 122.Omega cm(2) for FD4 during the transport assay, P(app) remained independent from the basal permeability of the in vitro BBB. Below these TEER values, P(app) increased exponentially. This nonlinear relationship between basal BBB permeability and P(app) was described by a one-phase exponential decay model. From this model the BBB permeability status independent permeability coefficients for FLU and FD4 (P(FLU) and P(FD4)) were estimated to be 2.2+/-0.1 and 0.48+/ 0.03 (10(-6)cm/s), respectively. In the experimentally challenged experiments, a reliable indication for P(FLU) and P(FD4) could be estimated only after the (simulated) drug-induced change in BBB permeability was taken into account. CONCLUSIONS: The assessment of basal BBB permeability status during drug transport assays was essential for an accurate estimation of the in vitro permeability coefficient of a drug. To accurately extrapolate the in vitro permeability coefficient of a drug to the in vivo situation, it is essential that drug-induced changes in the in vitro BBB permeability during the drug transport assay are determined. PMID- 11102737 TI - Modulation of oral bioavailability of anticancer drugs: from mouse to man. AB - Oral bioavailability of many anticancer drugs is poor and highly variable. This is a major impediment to the development of new generation drugs in oncology, particularly those requiring a chronic treatment schedule, a.o. the farnesyltransferase inhibitors. Limited bioavailability is mainly due to: (1) cytochrome P450 (CYP) activity in gut wall and liver, and (2) drug transporters, such as P-gp in gut wall and liver. Shared substrate drugs are affected by the combined activity of these systems. Available preclinical in vitro and in vivo models are in many cases only poorly predictive for oral drug uptake in patients because of a.o. interspecies differences in CYP drug metabolism and intestinal drug-transporting systems. Clearly, novel systems that allow reliable translation of preclinical results to the clinic are strongly needed. Our previous work, also using P-gp knockout (KO) mice, already showed that P-gp has a major effect on the oral bioavailability of several drugs and that blockers of P-gp can drastically improve oral bioavailability of paclitaxel and other drugs in mice and humans (Schinkel et al., Cell 77 (1994) 491; Sparreboom et al., Proc. Natl. Acad, Sci. USA 94 (1997) 2031; Meerum Terwogt et al. Lancet 352 (1998) 285). This work revealed, however, that apart from P-gp other drug-transporting systems and CYP effects also determine overall oral drug uptake. The taxanes paclitaxel and docetaxel are considered excellent substrate drugs to test the concept that by inhibition of P-gp in the gut wall and CYP activity in gut wall and/or liver low oral bioavailability can be increased substantially. In current studies we focus on the development of chronic oral treatment schedules with these drugs and on other drug transport systems that may play a significant role in regulation of oral bioavailability of other classes of (anti-cancer) drugs. The current review paper describes the background and summarizes our recent results of modulation of oral bioavailability of poorly available drugs, focused on drug transport systems and CYP in gut wall and liver. PMID- 11102738 TI - Clinical pharmacokinetics of phenobarbital in neonates. AB - Demographic and clinical pharmacokinetic data collected from term and preterm neonates who were treated with intravenous phenobarbital have been analysed to evaluate the role of patient characteristics in pharmacokinetic parameters. Significant relationships between total body weight (TBW) or body surface area (BSA) and volume of distribution (Vd) and total body clearance (CL) were found. Coefficients of determination were: 0.55 and 0.59 for Vd, and 0.76 and 0.72 for CL against TBW and BSA, respectively. In addition, significant relationships between height of the infants and volume of distribution of phenobarbital and total body clearance were observed. Coefficients of determination were 0.58 for Vd and 0.56 for CL. A weaker but significant correlation existed between gestational age and Vd or CL of phenobarbital. Coefficients of determination were 0.43 and 0.64, respectively. There was no correlation between volume of distribution per kg body weight or total body clearance per kg body weight and any patient parameter investigated. However, these latter pharmacokinetic parameters tended to decrease with increasing gestational age and height of the neonates. The results obtained were used to develop new loading and maintenance doses for phenobarbital in neonates based on total body weight and body surface area and based on height and gestational age for cases that weight is not available. PMID- 11102739 TI - Prediction of interindividual variation in drug plasma levels in vivo from individual enzyme kinetic data and physiologically based pharmacokinetic modeling. AB - A strategy is presented to predict interindividual variation in drug plasma levels in vivo by the use of physiologically based pharmacokinetic modeling and human in vitro metabolic parameters, obtained through the combined use of microsomes containing single cytochrome P450 enzymes and a human liver microsome bank. The strategy, applied to the pharmaceutical compound (N-[2-(7-methoxy-1 naphtyl)-ethyl]acetamide), consists of the following steps: (1) estimation of enzyme kinetic parameters K(m) and V(max) for the key cytochrome P450 enzymes using microsomes containing individual P450 enzymes; (2) scaling-up of the V(max) values for each individual cytochrome P450 involved using the ratio between marker substrate activities obtained from the same microsomes containing single P450 enzymes and a human liver microsome bank; (3) incorporation into a physiologically based pharmacokinetic model. For validation, predicted blood plasma levels and pharmacokinetic parameters were compared to those found in human volunteers: both the absolute plasma levels as well as the range in plasma levels were well predicted. Therefore, the presented strategy appears to be promising with respect to the integration of interindividual differences in metabolism and prediction of the possible impact on plasma and tissue concentrations of drugs in humans. PMID- 11102740 TI - Diffusion of n-butyl-p-aminobenzoate (BAB), lidocaine and bupivacaine through the human dura-arachnoid mater in vitro. AB - INTRODUCTION: Epidural administration of a suspension of n-butyl-p-aminobenzoate (BAB) to humans resulted in a selective, ultra-long lasting sensory block without motor function impairment. The absence of motor block was attributed to 'spatial' confinement of active concentrations of dissolved BAB within the epidural space. This study was designed to investigate the diffusion of BAB through the human dura-arachnoid membrane in vitro relative to lidocaine and bupivacaine and to quantify the influence of the composition of the suspension formulation on this flux. MATERIALS AND METHODS: Human dura-arachnoid specimens were mounted between the donor and the receiver compartment of a diffusion cell. Five concentrations of BAB, lidocaine and bupivacaine in phosphate-buffered saline, pH 7.4, were added to the donor compartment and the increase of the concentration of the agent in time in the receiver compartment was measured by automated UV-spectrometry. Fluxes and permeabilities were calculated. The influence of pH, polysorbate 80 (PS 80) and polyethylene glycol 3350 (PEG 3350) on the flux of BAB in solution and in suspension formulations were analyzed. RESULTS: The flux of both lidocaine and bupivacaine at pH 4 was considerably smaller than at pH 7.4. Permeabilities decreased in the order bupivacaine>lidocaine&z.Gt;BAB and at the level T12>T1. PS 80 at concentrations exceeding 0.025 mg/ml and PEG 3350 decreased the flux of BAB from BAB-solutions. Used in the preparation of the suspension, PS 80 and PEG 3350 did significantly reduce the permeability. DISCUSSION: The results of this study are consistent with the hypothesis that the selective action of epidurally applied BAB suspension can be attributed to the spatial confinement of active BAB concentrations within the epidural space. Additives used in the preparation of the aqueous suspension formulation may substantially influence the local pharmacokinetics and by that the pharmacodynamic effects. PMID- 11102741 TI - Sorbitol as a marker for drug-induced decreases of variable duration in liver blood flow in healthy volunteers. AB - OBJECTIVE: Sorbitol has been suggested as a suitable marker to assess liver blood flow (LBF), after it was shown to adequately reflect prolonged changes in LBF but changes of a shorter duration have not been investigated. We therefore used sorbitol to evaluate drug-induced decreases in LBF of variable duration with i.v. infusions of somatostatin and its synthetic analogue octreotide. METHODS: In a double-blind, placebo controlled, randomised study, six healthy males received sorbitol for 170 min. At sorbitol steady state, which was at 45 min after the start of the infusion (t=0), somatostatin or octreotide was infused for 30 min. Sampling for sorbitol assay and echo-Doppler hepatic portal vein flow measurements were done regularly and treatments were compared using ANOVA. RESULTS: The sorbitol AUC over the 30-min intervention period was 15% (95% C.I.: +4, +22%) and 13% (+5, +24%) higher compared to placebo after somatostatin and octreotide respectively. The decline of sorbitol levels after termination of the intervention was faster for somatostatin compared to octreotide, demonstrated by the difference in the AUC (0-2 h) with placebo which was 8% (-3, +19%) lower after somatostatin, and 15% (+5, +26%) after octreotide. Portal venous blood flow decreased during the 30-min interventions; after somatostatin 27% (-14, -40%) and after octreotide 29% (-17, -42%). Portal flow was lower than placebo during the entire experiment after octreotide 30% (-10, -50%), but not after somatostatin 13% (-33, +7%). Changes in sorbitol levels and portal venous blood flow occurred simultaneously and were well correlated for each individual, making it likely that the interventions did not interfere with metabolism. CONCLUSION: Sorbitol can be used to adequately assess decreases in LBF of variable duration in healthy volunteers. PMID- 11102742 TI - Pharmacokinetic-pharmacodynamic modelling of tiagabine CNS effects upon chronic treatment in rats: lack of change in concentration-EEG effect relationship. AB - The pharmacodynamics of the gamma-aminobutyric acid (GABA) uptake inhibitor (R)-N (4,4-di-(methylthien-2-yl)but-3-enyl) nipecotic acid (tiagabine) was quantified in rats following chronic (14 days) administration by an integrated pharmacokinetic-pharmacodynamic (PK/PD) modelling approach. The increase in beta activity (11.5-30 Hz) of the EEG as derived by fast Fourier transformation analysis was used as pharmacodynamic endpoint. Two groups of male Wistar rats were treated for 14 days with either tiagabine at a steady-state concentration of 198+/-10 ng ml(-1) or placebo. Chronic treatment with tiagabine resulted in an increase of the EEG effect parameter by 38+/-2 microV. In the PK/PD experiment the time course of the EEG effect was determined in conjunction with the decline of drug concentrations after an i.v. bolus administration of 10 mg kg(-1). The pharmacokinetics of tiagabine was most adequately described by a bi-exponential function. No influence of chronic treatment on the pharmacokinetics was observed. Hysteresis between plasma concentration and EEG effect was accounted for by incorporation of an 'effect-compartment' in the model. The observed relationship between tiagabine concentrations and EEG effect was non-linear and described on the basis of the Hill equation. Between the treatment groups no differences in the pharmacodynamic parameters were observed. The population means for the different pharmacodynamic parameters were: maximum EEG effect 82 microV, EC(50) 486 ng ml(-1), Hill factor 2.0 and k(e0) 0.060 min(-1). In the in vitro [(3)H]GABA uptake assay no changes in affinity or functionality for the GABA uptake transporter were observed, consistent with the absence of adaptation. It is concluded that chronic treatment with tiagabine in an effective dose range for 14 days does not produce functional adaptation to tiagabine-induced GABA-ergic inhibition in vivo. PMID- 11102743 TI - Evaluation of a novel high-throughput assay for cytochrome P450 2D6 using 7 methoxy-4-(aminomethyl)-coumarin. AB - We recently reported on the design, synthesis and characterisation of a novel and selective substrate of human cytochrome P450 2D6 (CYP2D6), 7-methoxy-4 (aminomethyl)-coumarin (MAMC). Here, we describe a high-throughput microplate reader assay, which makes use of MAMC as a fluorescent probe for determining the inhibition and activity of CYP2D6 in heterologously expressed systems and human liver microsomes. The high-throughput screening (HTS) assay can be used both in an end-point and real-time configuration, and is easy to use, rapid and sensitive. In addition, end-point measurements by means of flow injection analysis have also successfully been performed. The HTS-assay was validated by performing inhibition experiments for several low- and high-affinity ligands (n=6) of CYP2D6, and comparing the findings to those obtained with the standard O demethylation assay of dextromethorphan. The results indicate that all compounds tested display competitive inhibition in both the MAMC and dextromethorphan assay, and the K(i) values reveal a very good correlation (R(2)=0.984) between the two assays. To further demonstrate the usefulness of the HTS-assay, IC(50) values of a series of five N-substituted analogs of 3, 4 methylenedioxyamphetamine for CYP2D6 have been determined. The results obtained demonstrate that the current HTS-assay represents a significant improvement over previous assays, with a higher turnover of MAMC and a higher selectivity for CYP2D6. PMID- 11102744 TI - Investigation of a correlation between monoamine oxidase B and catechol-O methyltransferase activity in human blood cells. AB - Monoamine oxidase B (MAO-B) and catechol-O-methyltransferase (COMT) are pivotal enzymes in the catabolism of several neurotransmitters. MAO-B and COMT activity can be reliably measured in human platelets and erythrocytes, respectively. This study investigated whether a correlation exists between the activity of the two enzymes in a panel of 47 elderly subjects (age range 55-80 years). No correlation was apparent between the two activities (r(2)<0.01), which suggests that, genetically, they are determined independently. COMT activity in a panel of 163 subjects showed a bimodal distribution with a nadir at approximately 38 pmol/h/mg Hb. PMID- 11102745 TI - Matricellular proteins: an overview. PMID- 11102746 TI - Thrombospondin 2, a matricellular protein with diverse functions. AB - Thrombospondin (TSP) 2 is a close relative of TSP1 but differs in its temporal and spatial distribution in the mouse. This difference in expression undoubtedly reflects the marked disparity in the DNA sequences of the promoters in the genes encoding the two proteins. The synthesis of TSP2 occurs primarily in connective tissues of the developing and growing mouse. In the adult animal the protein is again produced in response to tissue injury and in association with the growth of tumors. Despite the abnormalities in collagen fibrillogenesis, fragility of skin, and laxity of tendons and ligaments observed in the TSP2-null mouse, TSP2 does not appear to contribute directly to the structural integrity of connective tissue elements. Instead, emerging evidence supports a mode of action of TSP2 'at a distance', i.e. by modulating the activity and bioavailability of proteases and growth factors in the pericellular environment and, very likely, by interaction with cell-surface receptors. Thus, TSP2 qualifies as a matricellular protein, as defined in the introduction to this minireview series. The phenotype of TSP2-null mice has been very helpful in providing clues to the functions of TSP2. In addition to histological and functional abnormalities in connective tissues, these mice display an increased vascularity of the dermis and subdermal tissues, increased endosteal bone growth, a bleeding defect, and a marked adhesive defect of dermal fibroblasts. Our laboratory has established that TSP2 binds matrix metalloproteinase 2 (MMP2) and that the adhesive defect in TSP2-null fibroblasts results from increased MMP2 activity. The investigation of the basis for the other defects in the TSP2-null mouse is likely to yield equally interesting results. PMID- 11102747 TI - SPARC, a matricellular protein: at the crossroads of cell-matrix. AB - SPARC is a multifunctional glycoprotein that belongs to the matricellular group of proteins. It modulates cellular interaction with the extracellular matrix (ECM) by its binding to structural matrix proteins, such as collagen and vitronectin, and by its abrogation of focal adhesions, features contributing to a counteradhesive effect on cells. SPARC inhibits cellular proliferation by an arrest of cells in the G1 phase of the cell cycle. It also regulates the activity of growth factors, such as platelet-derived growth factor (PDGF), fibroblast growth factor (FGF)-2, and vascular endothelial growth factor (VEGF). The expression of SPARC in adult animals is limited largely to remodeling tissue, such as bone, gut mucosa, and healing wounds, and it is prominent in tumors and in disorders associated with fibrosis. The crystal structure of two of the three domains of the protein has revealed a novel follistatin-like module and an extracellular calcium-binding (EC) module containing two EF-hand motifs. The follistatin-like module and the EC module are shared by at least four other proteins that comprise a family of SPARC-related genes. Targeted disruption of the SPARC locus in mice has shown that SPARC is important for lens transparency, as SPARC-null mice develop cataracts shortly after birth. SPARC is a prototypical matricellular protein that functions to regulate cell-matrix interactions and thereby influences many important physiological and pathological processes. PMID- 11102748 TI - Tenascin-C in development and disease: gene regulation and cell function. AB - Tenascin-C (TN-C) is a modular and multifunctional extracellular matrix (ECM) glycoprotein that is exquisitely regulated during embryonic development and in adult tissue remodeling. TN-C gene transcription is controlled by intracellular signals that are generated by multiple soluble factors, integrins and mechanical forces. These external cues are interpreted by particular DNA control elements that interact with different classes of transcription factors to activate or repress TN-C expression in a cell type- and differentiation-dependent fashion. Among the transcriptional regulators of the TN-C gene that have been identified, the homeobox family of proteins has emerged as a major player. Downstream from TN C, intracellular signals that are relayed via specific cell surface receptors often impart contrary cellular functions, even within the same cell type. A key to understanding this behavior may lie in the dual ability of TN-C-enriched extracellular matrices to generate intracellular signals, and to define unique cellular morphologies that modulate these signal transduction pathways. Thus, despite the contention that TN-C null mice appear to develop and act normally, TN C biology continues to provide a wealth of information regarding the complex nature of the ECM in development and disease. PMID- 11102749 TI - The cell biology of thrombospondin-1. AB - Thrombospondin-1 (TSP-1) is a matricellular protein that regulates cellular phenotype during tissue genesis and repair. It acts as a molecular facilitator by bringing together cytokines, growth factors, matrix components, membrane receptors and extracellular proteases. TSP-1 binds to a wide variety of integrin and non-integrin cell surface receptors. The binding sites for these receptors on TSP-1 are dispersed throughout the molecule, with most domains binding multiple receptors. In some cases, TSP-1 binds to multiple receptors concurrently, and recent data indicate that there is cross-talk between the receptor systems. Thus, TSP-1 may function to direct the clustering of receptors to specialized domains for adhesion and signal transduction. PMID- 11102750 TI - Osteopontin: a versatile regulator of inflammation and biomineralization. AB - Osteopontin is a secreted glycoprotein with a multidomain structure and functions characteristic of a matricellular protein. Osteopontin interacts with cell surface receptors via arginine-glycine-aspartate (RGD)- and non-RGD containing adhesive domains, in addition to binding to components of the structural extracellular matrix. While normally expressed in bone and kidney, osteopontin levels are elevated during wound healing and inflammation in most tissues studied to date. Since 1986, over one thousand studies have been published on osteopontin, including recent experiments in osteopontin-deficient mice. These studies reveal osteopontin as a cell adhesive, signaling, migratory, and survival stimulus for various mesenchymal, epithelial, and inflammatory cells, in addition to being a potent regulator of osseous and ectopic calcification. Based on these reports, a general picture of osteopontin as an important regulator of inflammation and biomineralization is emerging. A common denominator in osteopontin function in these situations is its ability to regulate the function of macrophage and macrophage-derived cells (i.e. osteoclasts). While we have learned much about osteopontin and the processes it appears to regulate over the past decade, many questions regarding this important multifunctional protein remain unanswered and provide important directions for future studies. PMID- 11102751 TI - Polymorphism in matrix metalloproteinase gene promoters: implication in regulation of gene expression and susceptibility of various diseases. AB - The matrix metalloproteinases (MMPs) can degrade a range of extracellular matrix proteins and have been implicated in connective tissue destruction and remodelling associated with cancer invasion and metastasis, cartilage destruction in arthritis, atherosclerotic plaque rupture, and the development of aneurysms. Recently, naturally occurring sequence variation has been detected in the promoter of a number of MMP genes. These genetic polymorphisms have been shown to have allele-specific effects on the transcriptional activities of MMP gene promoters, and to be associated with susceptibility to coronary heart disease, aneurysms and cancers. These findings indicate that variation in the MMP genes may contribute to inter-individual differences in susceptibility to these common, complex diseases, likely through effects on the balance between the synthesis and degradation of extracellular matrix proteins. PMID- 11102752 TI - Influence of decorin expression on transforming growth factor-beta-mediated collagen gel retraction and biglycan induction. AB - Complex formation of transforming growth factor-beta (TGF-beta) with the small proteoglycan decorin has been considered to inactivate the cytokine. However, neither the TGF-beta-mediated stimulation of the retraction of collagen lattices in culture nor the enhanced transcription of biglycan were influenced by an excess of native decorin in the culture medium. In contrast, when MG-63 osteosarcoma cells were transfected with sense- or antisense-decorin-cDNA, which led to an over- or under-expression of the proteoglycan, they responded to TGF beta differently. An inverse correlation between decorin expression and the TGF beta-mediated stimulation of collagen gel retraction and biglycan induction, respectively, was found. These results are best explained by assuming that decorin is not inactivating but sequestering TGF-beta in the extracellular matrix. PMID- 11102753 TI - Differential expression of laminin alpha chains during proliferative and differentiation stages in a model for skin morphogenesis. AB - The purpose of this study was to determine the mRNA and protein expression of laminin alpha chains at various stages of in vitro skin morphogenesis. Fibroblasts in mono-cultures express low levels of the mRNA of laminin alpha1,alpha2, alpha3 and alpha4 chains. When co-cultured with keratinocytes for 28 days, they expressed the mRNA for all these chains. Keratinocytes in monolayer expressed the laminin alpha3 chain mRNA and very low levels of the mRNA of the alpha1 and alpha2 chains, although, when recombined with fibroblasts they also expressed laminin alpha1and alpha2 mRNA, but not the laminin alpha4 mRNA. Immunocytochemistry of cells in co-culture showed that laminin alpha1, alpha3 and alpha5 chains were expressed in the epidermis, while the laminin alpha2, beta1, and gamma1 chains were noted in the dermis and at the epidermo-dermal interface. The laminin alpha1chain was first expressed during the proliferative stage (14-21 days) and the laminin alpha2 and alpha5 chains appeared later, during the differentiation stage (28-42 days). The above results suggest that epithelial mesenchymal interactions are involved in the expression of laminin alpha chain mRNA during in vitro skin morphogenesis. In addition, there is distinct temporal and spatial expression of these chains during proliferative and differentiation stages, possibly reflecting different functions. PMID- 11102754 TI - Expression of matrilin-1, -2 and -3 in developing mouse limbs and heart. AB - The expression of matrilin-1, -2 and -3 was studied in the heart and limb during mouse development. Matrilin-1 is transiently expressed in the heart between days 9.5 and 14.5 p.c. Matrilin-2 expression was detected in the heart from day 10.5 p.c. onwards. In the developing limb bud, both matrilin-1 and -3 were observed first at day 12.5 p.c. Throughout development matrilin-3 expression was strictly limited to cartilage, while matrilin-1 was also found in some other forms of connective tissue. Matrilin-2, albeit present around hypertrophic chondrocytes in the growth plate, was mainly expressed in non-skeletal structures. The complementary, but in part overlapping, expression of matrilins indicates the possibility for both redundant and unique functions among the members of this novel family of extracellular matrix proteins. PMID- 11102755 TI - Coordinate induction of collagenase-1, stromelysin-1 and urokinase plasminogen activator (uPA) by the 120-kDa cell-binding fibronectin fragment in fibrocartilaginous cells: uPA contributes to activation of procollagenase-1. AB - Specific fibronectin (Fn) fragments found in synovial fluid of arthritic joints potentially contribute to the loss of cartilage proteoglycans by inducing matrix metalloproteinase (MMP) expression. However, whether or not the Fn fragment modulated changes in expression of MMPs result in a net increase in matrix degradative activity through alterations in the balance between MMP activation and inhibition has not been established. To understand the mechanisms by which proteolytic Fn fragments may contribute to joint degeneration, conditioned medium from fibrocartilaginous cells exposed to Fn, its 30-kDa fragment containing the collagen/gelatin-binding domain, its 120-kDa fragment containing the central cell binding domain, and the RGD peptide were assayed for MMPs, and MMP activators and inhibitors. We found that the 120-kDa fragment of Fn (but not intact Fn), the 30 kDa fragment, and the RGD peptide, dose-dependently induced procollagenase-1 and prostromelysin-1 and decreased levels of the tissue inhibitor of metalloproteinases (TIMPs) -1 and -2. The alpha5beta1 integrin was implicated in the induction of collagenase by the 120-kDa Fn fragment, since collagenase induction was abrogated in the presence of blocking antibody to this integrin. Conditioned medium from cells exposed to the 120-kDa Fn fragment also demonstrated increased levels of the activated collagenase-1, which resulted in significantly elevated collagen degradative activity. That the urokinase plasminogen activator (uPA) was involved in the activation of procollagenase-1 was suggested by findings that the 120-kDa Fn fragment induced uPA coordinately with procollagenase-1, and the activation of procollagenase-1 was dose dependently inhibited in the presence of plasminogen activator inhibitor-1. These data demonstrate that the 120-kDa cell-binding fragment of Fn induces a net increase in matrix-degradative activity in fibrocartilaginous cells by concomitantly inducing MMPs and their activator, uPA, while decreasing TIMPs. PMID- 11102756 TI - Revised genomic structure of the human MAGP1 gene and identification of alternate transcripts in human and mouse tissues. AB - The human MAGP1 (or MFAP2) and mouse Magp1 genes code for the microfibril associated glycoprotein-1 (MAGP-1), an extracellular matrix protein of microfibrillar structures. We report a revised 5' genomic structure including the use of a single transcription start site that gives rise to a 32-bp 5' exon spanning a segment of the previously described exon B. No evidence of heterogeneous 5' ends from the use of alternative promoters was found in human tissues and cell lines. We located the genetic marker D1S170 to a position 3 kb downstream of the polyadenylation site. Large-scale comparison of the human and mouse genes revealed conservation of sequence outside the coding exons. Although the 5' flanking regions were found to be divergent certain cis-elements for transcription factors are conserved, including Sp1, AP-2, AP-4, NF-kappaB, and c ETS motifs. We identified a total of five splice variants in addition to the canonical MAGP1A/Magp1A form. These transcripts are species-specific and are generated by different processing mechanisms. The alternate forms MAGP1A', MAGP1B, and MAGP1C are expressed in human tissues; and the two variants Magp1A" and Magp1D were found only in mouse. The alternatively spliced forms show restricted patterns of expression relative to the canonical isoform. PMID- 11102757 TI - Monoclonal antibodies to mineralized matrix molecules of the avian eggshell. AB - The extracellular matrix of the mineralizing eggshell contains molecules hypothesized to be regulators of biomineralization. To study eggshell matrix molecules, a bank of monoclonal antibodies was generated that bound demineralized eggshell matrix or localized to oviduct epithelium. Immunofluorescence staining revealed several staining patterns for antibodies that recognized secretory cells: staining for a majority of columnar lining cells, staining for a minor sub set of columnar lining cells, intensified staining within epithelial crypts, and staining of the entire tubular gland. Western blotting with the antibody Epi2 on eggshell matrix showed binding to molecules with the apparent molecular weight of eggshell matrix dermatan sulfate proteoglycan (eggshell DSPG). Immunoblots of cyanogen bromide-cleaved eggshell DSPG revealed broad band of reactivity that shifted to 25 kDa after chondroitinase digestion; indicating that the Epi2 binding site is located on a fragment which contains dermatan sulfate side chains. Immunogold labeling showed that Epi2 binds to secretory vesicles within the non-ciliated cells of the columnar epithelium, while the antibodies Tg1 and Tg2 bind to secretory vesicles of tubular gland cells. Immunogold labeling of demineralized shell matrix showed binding of Epi2, Tg1, and Tg2 to the matrix of the palisade layer, and showed little reactivity to other regions of the shell matrix. Quantification of the immunogold particles within the eggshell matrix revealed that antibodies Epi2 and Tg1 bind all calcified regions equally while antibody Tg2 has a greater affinity for the baseplate region of the calcium reserve assembly. PMID- 11102758 TI - Molecular cloning and relative tissue expression of keratocan and mimecan in embryonic quail cornea. AB - We have cloned and sequenced the cDNAs for quail cornea keratan sulfate proteoglycan core proteins, keratocan and mimecan. The deduced quail keratocan protein contains a single conservative amino acid difference from the chick sequence, whereas quail mimecan protein contains a 58 amino acid-long avian unique sequence that shares no homology with mammalian mimecan. Ribonuclease protection assay of Day 16 embryonic quail tissues reveals that keratocan and lumican are expressed at highest levels in cornea, whereas mimecan mRNA is expressed at a much lower level. Keratocan is expressed only in quail cornea, whereas mimecan is expressed in many different tissues as four transcripts of different sizes. Both lumican and mimecan are expressed at lowest levels in brain, liver and sternum. PMID- 11102759 TI - Molecular cloning and relative tissue expression of decorin and lumican in embryonic quail cornea. AB - We have cloned and sequenced the cDNAs for quail cornea proteoglycan core proteins, decorin and lumican. Comparison of deduced amino acid sequences shows that two of five amino acid differences in the mature protein between quail and chick decorin, and two of three for lumican, are non-conservative. Ribonuclease protection assay of Day 16 embryonic quail tissues reveals that decorin and lumican are most highly expressed in cornea, and that both are also highly expressed at approximately equal levels in most other tissues. Decorin is highly expressed in sclera and sternum, whereas lumican is expressed in these tissues, as well as in liver, at very low levels. Both decorin and lumican are expressed at lowest levels in brain. PMID- 11102760 TI - Immunology PMID- 11102761 TI - Web alert. Atopic allergy and other hypersensitivities. Autoimmunity. PMID- 11102762 TI - Allergy, a disease of the internal and external environments PMID- 11102763 TI - Regulation of the IgE isotype switch: new insights on cytokine signals and the functions of epsilon germline transcripts. AB - In allergic responses, B cells are driven to undergo an immunoglobulin isotype switch, shifting from IgM to IgE synthesis. This process involves the rearrangement of germline DNA in the immunoglobulin heavy-chain locus and is stimulated by cytokines (IL-4 and IL-13) and CD40 activation. It is now evident that cytokine-induced 'germline' epsilon-RNA transcripts associate with DNA in the genomic switch region (S epsilon) to form DNA-RNA hybrid structures, which target nucleases in for deletional switch recombination. Alterations in cytokine production and signaling affect the efficiency of this process and are associated with inherited predisposition to allergy. PMID- 11102764 TI - Roles of mast cells and basophils in innate and acquired immunity. AB - There have been several recent advances in knowledge about mast cells and basophils in immune responses, of which some are particularly important: a role has been found for heparin in the storage of certain proteases and other mediators in mast cell cytoplasmic granules; an important role for mast cells in the development of several chronic aspects of an asthma model in mice has been discovered; and a new approach has been developed, based on the generation of mast cells from embryonic stem cells in vitro, to investigate mast cell function in vitro or in vivo. PMID- 11102765 TI - The influence of infections on the development and severity of allergic disorders. AB - The frequency and severity of atopic disorders are steadily increasing, particularly in developing countries. The reason for this observation is not clear. Recent studies indicate that infections with viruses and especially with bacteria early in life may help to inhibit allergic Th2 responses by skewing the immune system towards Th1 responses. However, infections can also lead to the exacerbation of atopic disorders. PMID- 11102766 TI - Immune regulation in atopic dermatitis. AB - Atopic dermatitis is a chronic inflammatory skin disease with a pathogenesis of complex immune dysregulation and interplay of genetic, environmental and psychological factors. Activation and skin-selective homing of peripheral-blood T cells, and effector functions in the skin, represent sequential immunological events in the pathogenesis of atopic dermatitis. Both CD4(+) and CD8(+) T cells bearing the cutaneous-lymphocyte-associated antigen represent activated memory/effector T cell subsets and induce IgE, mainly via IL-13, and prolong eosinophil lifespan, mainly via IL-5. Dysregulated apoptosis in skin-homing T cells and keratinocytes contributes to the elicitation and progress of atopic dermatitis. T cell survival is enhanced in the skin by cytokines and extracellular-matrix proteins. These activated T cells induce keratinocyte apoptosis, leading to eczema formation. PMID- 11102767 TI - Mechanisms of food allergy. AB - The prevalence of food allergy continues to rise, particularly in 'westernized' societies; it has been linked to the 'hygiene hypothesis' and the increased diversity of food consumption worldwide. The pathogenic mechanisms and Th1/Th2 paradigm are being closely examined with respect to the occurrence of inflammatory and injury/repair responses at different mucosal sites. Genetically modified plants as potential food sources and allergenicity are current topics of controversy. PMID- 11102769 TI - Editorial overview PMID- 11102768 TI - T cell differentiation: a mechanistic view. AB - It has recently become clear that cytokine expression by T cells involves epigenetic changes in chromatin structure, locus accessibility and DNA methylation that occur during differentiation of naive T cells into mature effector T cells. These changes require the coordinate actions of antigen- and cytokine-induced transcription factors, chromatin remodeling complexes, histone modifying enzymes and subset-specific transcription factors. PMID- 11102770 TI - Helper T cells in antibody-mediated, organ-specific autoimmunity. AB - The production of pathogenic autoantibodies in organ-specific autoimmune diseases is largely T cell dependent. For many of these diseases, the precise specificities and cytokine profiles of the T cells that respond to the corresponding autoantigens have now been identified. This knowledge has been exploited to treat some models of antibody-mediated autoimmunity using peptides corresponding to the dominant helper epitopes, giving impetus to the development of a similar approach in the equivalent human diseases. PMID- 11102771 TI - Present difficulties and future promise of MHC multimers in autoimmune exploration. AB - Class I tetramers have been used to track cytotoxic T cells during bacterial and viral infections. During the past year, the use of such molecules has revealed important information about the role of CD8(+) T cells in autoimmune diabetes. Furthermore, class II multimers have been produced and successfully used to stain autoreactive CD4(+) T cells from patients with rheumatoid arthritis or Borrelia burgdorferi-induced Lyme arthritis. PMID- 11102772 TI - The role of different subsets of T regulatory cells in controlling autoimmunity. AB - T regulatory cells-in addition to clonal deletion and anergy-are essential for the downregulation of T cell responses to both foreign and self antigens, and for the prevention of autoimmunity. Recent progress has been made in characterising the different subsets of T regulatory cells, the factors that drive their differentiation, and their mode of action. The resolution of these mechanisms will make it possible to use T regulatory cells therapeutically in human autoimmune diseases. PMID- 11102773 TI - Animal models of autoimmunity and their relevance to human diseases. AB - T cells mediate various autoimmune diseases. Pathologic autoimmunity can be induced by manipulating thymic or peripheral control of self-reactive T cells. There is, for example, accumulating evidence that elimination or dysfunction of regulatory T cells can elicit T cell mediated, destructive autoimmune disease in otherwise normal animals and enhance autoimmunity in spontaneous models. PMID- 11102774 TI - Mapping and identification of autoimmunity genes. AB - Over the past several years, intense effort has been made to map the chromosomal locations of genes involved in the susceptibility to autoimmune diseases. The first phase of this mapping effort-performed in most cases by using microsatellite markers to scan the genome for loci that are linked with disease has generated first-draft maps for numerous autoimmune diseases in humans, mice and rats, pointing to as many as 20 different loci in some diseases. The second phase is now beginning, with efforts focused on narrowing the loci sufficiently to allow the positional cloning of disease-associated alleles. From these mapping data it is clear that some of these loci overlap between various autoimmune diseases and preliminary results suggest that indeed there is a sharing of 'autoimmunity genes' between various autoimmune diseases. PMID- 11102775 TI - Human autoimmunity genes in mice. AB - In the post-genomic era, the expression and investigation of human (auto)immunity genes seems more relevant than ever. The generation of humanized animal models of human diseases will be useful to study the interplay between genetic and non genetic factors in disease development and may form a basis for the development of new drugs that act more specifically than the ones currently in use. Transgenic mice have been generated that express various human proteins- candidate autoantigens, disease-associated MHC class II molecules, TCRs and/or CD4--in order to study diseases such as rheumatoid arthritis, multiple sclerosis and diabetes. PMID- 11102776 TI - Antigen-specific therapy for autoimmune disease. AB - The application of self-antigens as therapeutic tools is validated in inbred animal models of autoimmune disease. Mechanisms of antigen-induced tolerance (apoptosis, anergy, regulatory T cells and immune deviation) are being clarified in relation to the properties of antigens and the modes and routes of their delivery. Mucosa-mediated tolerance remains the predominant mode of antigen specific therapy but awaits demonstration of clinical efficacy in human autoimmune disease. PMID- 11102777 TI - Novel, non-antigen-specific therapeutic approaches to autoimmune/inflammatory diseases. AB - Anti-TNF therapy has made a major impact on the treatment of inflammatory arthritides and Crohn's disease. It leads to prompt and prolonged clinical response, even in patients refractory to conventional therapy. Moreover, the progression of joint damage noted in patients with rheumatoid arthritis treated with methotrexate was prevented by an anti-TNF-alpha antibody, suggesting a genuine disease-modifying potential of TNF-alpha blockade. PMID- 11102778 TI - Apoptosis genes and autoimmunity. AB - To try to understand autoimmunity, attention has often fallen on the process of cell death. After all, apoptosis is used during selection of immunocytes, cells in the target organs end up dying and mutations to cell death genes have been found in some autoimmune diseases. Furthermore, some autoimmune-prone mice fail to develop disease when certain cell death genes are deleted, and transgenic mice expressing other cell death genes develop autoimmunity. However, only a tiny proportion of human autoimmune disease is associated with mutations to individual genes and even in these rare cases the genetic background has a major influence on the severity of disease. An understanding of the pathophysiology of common autoimmune diseases will require elucidation of many different systems that interact in complex ways, of which the process of apoptosis is just one. PMID- 11102779 TI - The role of autoantigens in autoimmune disease. AB - Many autoantigens have been identified in human patients and in rodent models. In numerous experimental settings, these autoantigens or related autoreactive lymphocytes can transfer autoimmunity. Although autoreactivity spreads to new epitopes during the course of disease, single-epitope-specific therapies show considerable efficacy in multi-epitope-induced models of autoimmunity. These observations may indicate that epitope-specific therapies operate at the level of regulating mechanisms of immune tolerance rather than exerting a direct effect on autoreactive T lymphocytes. PMID- 11102781 TI - The immunogenicity of human and murine cytomegaloviruses PMID- 11102782 TI - Chemical biotechnology pharmaceutical biotechnology. Web alert. AB - A selection of World Wide Web sites relevant to papers published in this issue of Current Opinion in Biotechnology. PMID- 11102780 TI - Tolerance to the foeto-placental 'graft': ten ways to support a child for nine months. AB - Tolerance to the foetal 'allograft' has been extensively studied in the past few years, providing interesting new insights. In addition to a potential role for HLA-G, which has been widely discussed, there are hypotheses suggesting roles for several other molecules or cells: leukemia inhibitory factor and its receptor; indoleamine 2. 3-dioxygenase; the Th1/Th2 balance; suppressor macrophages; hormones such as progesterone or the placental growth hormone; CD95 and its ligand; and, as recently proposed, annexin II. Tolerance of the foetal allograft is probably the consequence of a wide panel of mechanisms that may or may not be pregnancy-specific, that are of major or secondary importance and that may be interconnected. PMID- 11102783 TI - Chemical biotechnology. A happy marriage between chemistry and biotechnology: asymmetric synthesis via green chemistry. PMID- 11102784 TI - Enzymatic asymmetric synthesis by decarboxylases. AB - Decarboxylation reactions using microbial cells or enzymes are increasingly being used for the synthesis of enantiomerically pure compounds because of their high degree of regio- and stereo-specificity. Pyruvate decarboxylase, benzoylformate decarboxylase and phenylpyruvate decarboxylase enzymes are capable of acyloin type condensation reactions leading to formation of chiral alpha-hydroxy ketones, which are versatile building blocks in the pharmaceutical and chemical industries. Availability of three-dimensional structures of some decarboxylases in recent years has facilitated understanding of reaction mechanisms and the creation of mutants with enhanced activity and stability. PMID- 11102785 TI - Applications of transketolases in organic synthesis. AB - The enzyme transketolase has been employed as a catalyst for asymmetric carbon carbon bond formation in the synthesis of biologically important molecules. A number of important parameters have been addressed including substrate specificity, over-expression of the protein in suitable host systems, scale-up of the reaction and use of transketolase in multi-enzyme experiments. X-ray structural studies have been used to probe the origin of the asymmetry of the carbon-carbon bond-forming process. PMID- 11102786 TI - Hydroxynitrile lyases in stereoselective catalysis. AB - (R)- as well as (S)-cyanohydrins are now easily available as a result of the excellent accessibility, the relatively high stability and the easy handling of hydroxynitrile lyases (HNLs). The optimization of reaction conditions (solvent, temperature, and using site-directed mutagenesis, etc.) has enabled HNL-catalyzed preparations of optically active cyanohydrins on a technical scale. The enantioselectivity of chiral metal-complex-catalyzed additions of trimethylsilyl cyanide to aldehydes has been improved, but is, by far, not yet competitive with the HNL-catalyzed reactions. PMID- 11102787 TI - Biodesulfurization and the upgrading of petroleum distillates. AB - Biotechnology offers an alternative way to process fossil fuels. There have been several important advances in the elucidation of the mechanisms of biodesulfurization and the development of a biocatalytic desulfurization process. These include a detailed analysis of the rate and extent of desulfurization of real target molecules in a diesel matrix, the directed evolution of rate- and extent-limiting enzymes for better performance and the expression of the genes in alternative hosts. Process innovations include new reactor designs, separations and recovery strategies and the production of value-added byproducts during desulfurization. PMID- 11102788 TI - Enzymatic hydroxylation reactions. AB - During the past 18 months, considerable progress has been made in the understanding of the key enzyme-substrate interactions that control the regioselectivity and stereoselectivity of the hydroxylation reaction performed by cytochrome-P450-dependent enzymes of mammalian origin. The manipulation of microbial hydroxylating enzymes, in both whole-cell and cell-free environments, has also been examined in the context of controlling the regioselectivity and stereoselectivity of the hydroxylation reaction. Several new applications for hydroxylating enzymes have been reported, and the construction of chimeric hydroxylating enzymes has been used both for mechanistic studies and for the production of enzymes with high hydroxylating activity for a defined substrate. PMID- 11102789 TI - Selective oxygen transfer catalysed by heme peroxidases: synthetic and mechanistic aspects. AB - The synthetic and mechanistic aspects of the use of heme peroxidases as functional mimics of the cytochrome P450 monooxygenases in oxygen-transfer reactions have been described. The chloroperoxidase from Caldariomyces fumago (CPO) is the catalyst of choice in sulfoxidation, hydroxylation and epoxidation on account of its high activity and enantioselectivity. Other heme peroxidases were less active by orders of magnitude; protein engineering has resulted in impressive improvements but even the most active mutant was still at least an order of magnitude less active than CPO. The 'oxygen-rebound' mechanisms of oxygen transfer mediated by heme enzymes - as originally conceived - have proved to be untenable. Dual pathway mechanisms, via oxoferryl species that insert oxygen as well as iron hydroperoxide species that insert OH(+), have been proposed that accommodate all of the known experimental data. PMID- 11102790 TI - Dynamic resolution and stereoinversion of secondary alcohols by chemo-enzymatic processes. AB - To overcome the maximum 50% yield limitation of classical resolution methods, deracemization processes involving a racemization step (dynamic resolution) or a prochiral intermediate (stereoinversion) have been developed. The use of transition metal complexes as racemizing agents, in combination with an enzymatic reaction, has been successfully extended to the deracemization of a number of simple or functionalized sec-alcohols. A two-enzyme process has been also investigated for their sequential or simultaneous deracemization. Other prominent results arise from an (apparently general) oxidoreduction process catalyzed by a single whole-cell microorganism. PMID- 11102791 TI - Online NMR for monitoring biocatalysed reactions. AB - Monitoring biocatalysed reactions and metabolic pathways using NMR spectroscopy is of growing interest. As a non-invasive analytical method providing simultaneous information about intracellular and extracellular constituents, it is superior to other analytical techniques and has a wide range of applications: kinetics and stoichiometrics of metabolic events, metabolic fluxes and enzyme activities can be detected in situ or after taking a sample from the biotransformation mixture. New NMR pulse sequences provide even more valuable experiments in these fields. Research topics range from the monitoring of polymer formation to fermentations producing beverages or antibiotics. Routine monitoring of industrial fermentations by NMR seems to be imminent. PMID- 11102792 TI - Pharmaceutical biotechnology. The genomes are just the beginning PMID- 11102793 TI - Whole genomes: the foundation of new biology and medicine. AB - Our genomic DNA sequence provides a unique glimpse of the provenance and evolution of our species, the migration of peoples, and the causation of disease. Understanding the genome may help resolve previously unanswerable questions, including perhaps which human characteristics are innate or acquired. Such an understanding will make it possible to study how genomic DNA sequence varies among populations and among individuals, including the role of such variation in the pathogenesis of important illnesses and responses to pharmaceuticals. The study of the genome and the associated proteomics of free-living organisms will eventually make it possible to localize and annotate every human gene, as well as the regulatory elements that control the timing, organ-site specificity, extent of gene expression, protein levels, and post-translational modifications. For any given physiological process, we will have a new paradigm for addressing its evolution, development, function, and mechanism. PMID- 11102794 TI - Drug discovery opportunities from apoptosis research. AB - Cell suicide is a normal process that participates in a wide variety of physiological processes, including tissue homeostasis, immune regulation, and fertility. Physiological cell death typically occurs by apoptosis, as opposed to necrosis. Defects in apoptotic cell-death regulation contribute to many diseases, including disorders associated with cell accumulation (e.g. cancer, autoimmunity, inflammation and restenosis) or where cell loss occurs (e.g. stroke, heart failure, neurodegeneration, AIDS and osteoporosis). At the center of the apoptosis machinery is a family of intracellular proteases, known as 'caspases', that are responsible directly or indirectly for the morphological and biochemical events that characterize apoptosis. Multiple positive and negative regulators of these cell-death proteases have been discovered in the genomes of mammals, amphibians, insects, nematodes, and other animal species, as well as a variety of animal viruses. Inputs from signal-transduction pathways into the core of the cell-death machinery have also been identified, demonstrating ways of linking environmental stimuli to cell-death responses or cell-survival maintenance. Knowledge of the molecular mechanisms of apoptosis has provided important insights into the causes of multiple diseases where aberrant cell-death regulation occurs and has revealed new approaches for identifying small-molecule drugs for more effectively treating these illnesses. PMID- 11102795 TI - Small-molecule inhibitors of cell signaling. AB - Small-molecule inhibitors of several intracellular signaling proteins, mostly protein kinases, show tremendous selectivity and potency. The complexity and redundancy of signaling pathways presents opportunities for therapeutic selectivity and some clinical results are remarkable. New strategies are being developed to interfere with previously intractable targets, such as protein protein interactions. PMID- 11102796 TI - Gene expression profiling of cardiovascular disease models. AB - Recent development of gene expression profiling technologies has enabled the large-scale analysis of gene expression changes during disease progression. Frequently, cardiovascular diseases involve complex interactions of multiple cell types over prolonged periods of time. A better understanding of the pathology of cardiovascular diseases and the potential identification of underlying genetic defects are currently being explored by using profiling methodologies in a number of animal and tissue-culture models. PMID- 11102797 TI - Applied genomics: integration of the technology within pharmaceutical research and development. AB - Multiple novel technologies have recently been developed to improve the analysis of genetic sequences, to rapidly assess RNA or protein levels in relevant tissues, and to validate function of potential new drug targets. The challenge facing pharmaceutical research is one of effective integration of these new technologies in ways that can maximally affect the discovery and development pipeline. Although database mining and transcriptional profiling clearly have increased the number of putative targets, the current focus is to assign function to new gene targets in a high-throughput manner. This requires a restructuring of the classical linear progression from gene identification, functional elucidation, target validation and screen development. New approaches are called for that can make this process non-linear and high-throughput. PMID- 11102798 TI - Phage display in pharmaceutical biotechnology. AB - Over the past year, methods for the construction of M13 phage-display libraries have been significantly improved and new display formats have been developed. Phage-displayed peptide libraries have been used to isolate specific ligands for numerous protein targets. New phage antibody libraries have further expanded the practical applications of the technology and phage cDNA libraries have proven useful in defining natural binding interactions. In addition, phage-display methods have been developed for the rapid determination of binding energetics at protein-protein interfaces. PMID- 11102799 TI - VEGF: an update on biological and therapeutic aspects. AB - Vascular endothelial growth factor (VEGF) is an endothelial cell-specific mitogen and an angiogenic inducer as well as a mediator of vascular permeability. VEGF is essential for developmental angiogenesis and is also required for female reproductive functions and endochondral bone formation. Substantial evidence also implicates VEGF in tumors and intraocular neovascular syndromes. Currently, several clinical trials are ongoing to test the hypothesis that the inhibition of VEGF activity may be beneficial for these conditions. PMID- 11102800 TI - Bacterial virulence as a target for antimicrobial chemotherapy. AB - As bacterial resistance to currently used antibiotics increases, so too must efforts to identify novel agents and strategies for the prevention and treatment of bacterial infection. In the past, antimicrobial drug discovery efforts have focused on eradicating infection by either cidal or static agents, resulting in clearance of the bacterium from the infected host. To this end, drug discovery targets have been those proteins or processes essential for bacterial cell viability. However, inhibition of the interaction between the bacterium and its host may also be a target. During establishment of an infection, pathogenic bacteria use carefully regulated pathways of conditional gene expression to transition from a free-living form to one that must adapt to the host milieu. This transition requires the regulated production of both extracellular and cell surface molecules, often termed virulence factors. As the biological imperatives of the invading organism change during the course of an infection, the expression of these factors is altered in response to environmental cues. These may be changes in the host environment, for example, pH, metabolites, metal ions, osmolarity, and temperature. Alternatively, effector molecules produced by the bacterium to sense changing cell density can also lead to changes in virulence gene expression. Although the mechanisms of pathogenesis among different bacteria vary, the principles of virulence are generally conserved. Bacterial virulence may therefore offer unique opportunities to inhibit the establishment of infection or alter its course as a method of antimicrobial chemotherapy. PMID- 11102801 TI - DAPP1 undergoes a PI 3-kinase-dependent cycle of plasma-membrane recruitment and endocytosis upon cell stimulation. AB - BACKGROUND: Phosphoinositide (PI) 3-kinase and its second messenger products, phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) and phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P(2)), play important roles in signalling processes crucial for cell movement, differentiation and survival. Previously, we isolated a 32kDa PtdIns(3,4,5)P(3)-binding protein from porcine leukocytes. This protein contains an amino-terminal Src homology 2 (SH2) domain and a carboxy-terminal pleckstrin homology (PH) domain, and is identical to the recently described DAPP1 (also known as PHISH or Bam32) protein. Here, we characterised the subcellular distribution of DAPP1 in response to cell stimulation. RESULTS: When expressed transiently in porcine aortic endothelial (PAE) cells, DAPP1 translocated from the cytosol to the plasma membrane in response to platelet-derived growth factor (PDGF). This translocation was dependent on both PI 3-kinase activity and an intact DAPP1 PH domain. Following recruitment to the plasma membrane, DAPP1 entered the cell in vesicles. Similar responses were seen in DT40 chicken B cells following antibody treatment, and Rat 1 fibroblasts following epidermal growth factor (EGF) or PDGF treatment. Colocalisation studies in PAE cells suggested entry of DAPP1 by endocytosis in a population of early endosomes containing internalised PDGF-beta receptors. DAPP1 also underwent PI 3-kinase-dependent phosphorylation on Tyr139 in response to PDGF stimulation, and this event was involved in the vesicular response. CONCLUSIONS: This is the first report of plasma-membrane recruitment and endocytosis of a PI 3-kinase effector protein in response to cell stimulation. The results suggest a novel role for DAPP1 in endosomal trafficking or sorting. PMID- 11102802 TI - Reciprocal inheritance of centrosomes in the parthenogenetic hymenopteran Nasonia vitripennis. AB - BACKGROUND: In the majority of animals, the centrosome-the microtubule-organizing center of the cell-is assembled from components of both the sperm and the egg. How the males of the insect order Hymenoptera acquire centrosomes is a mystery, as they originate from virgin birth. RESULTS: To address this issue, we observed centrosome, spindle and nuclear behavior in real time during early development in the parthenogenetic hymenopteran Nasonia vitripennis. Female meiosis was identical in unfertilized eggs. Centrosomes were assembled before the first mitotic division but were inherited differently in unfertilized and fertilized eggs. In both, large numbers of asters appeared at the cortex of the egg after completion of meiosis. In unfertilized eggs, the asters migrated inwards and two of them became stably associated with the female pronucleus and the remaining cytoplasmic asters rapidly disappeared. In fertilized eggs, the Nasonia sperm brought in paternally derived centrosomes, similar to Drosophila melanogaster. At pronuclear fusion, the diploid zygotic nucleus was associated only with paternally derived centrosomes. None of the cytoplasmic asters associated with the zygotic nucleus and, as in unfertilized eggs, they rapidly degenerated. CONCLUSIONS: Selection and migration of the female pronucleus is independent of the sperm and its aster. Unfertilized male eggs inherit maternal centrosomes whereas fertilized female eggs inherit paternal centrosomes. This is the first system described in which centrosomes are reciprocally inherited. The results suggest the existence of a previously undescribed mechanism for regulating centrosome number in the early embryo. PMID- 11102803 TI - Dynamic actin-based epithelial adhesion and cell matching during Drosophila dorsal closure. AB - BACKGROUND: The adhesion of two epithelial sheets is a fundamental process that occurs throughout embryogenesis and during wound repair. Sealing of the dorsal epidermis along the midline of the Drosophila embryo provides a genetically tractable model to analyse the closure of such holes. Several studies indicate that the actin cytoskeleton plays a critical role in dorsal closure. Although many components of the signalling cascade directing this process have been identified, the precise cell-biological events upon which these signals act remain poorly described. RESULTS: By confocal imaging of living fly embryos expressing green fluorescent protein (GFP)-tagged actin, we found that dorsal closure relies on the activity of dynamic filopodia and lamellipodia that extend from front-row cells to actively zipper the epithelial sheets together. As these epithelial fronts approach one another, we observed long, thin filopodia, apparently 'sampling' cells on the opposing face. When the assembly of these actin-based protrusions was blocked (by interfering with the activities of Cdc42 and Jun N-terminal kinase signalling), the adhesion and fusion of opposing epithelial cells was prevented and their ability to 'sense' correct partners was also blocked, leading to segment misalignment along the midline seam. CONCLUSIONS: Dynamic, actin-based protrusions (filopodia and lamellae) are critical, both in the mechanics of epithelial adhesion during dorsal closure and in the correct 'matching' of opposing cells along the fusion seam. PMID- 11102804 TI - A novel Dictyostelium RasGEF is required for normal endocytosis, cell motility and multicellular development. AB - BACKGROUND: Dictyostelium possesses a surprisingly large number of Ras proteins and little is known about their activators, the guanine nucleotide exchange factors (GEFs). It is also unclear, in Dictyostelium or in higher eukaryotes, whether Ras pathways are linear, with each Ras controlled by its own GEF, or networked, with multiple GEFs acting on multiple Ras proteins. RESULTS: We have identified the Dictyostelium gene that encodes RasGEFB, a protein with homology to known RasGEFs such as the Son-of-sevenless (Sos) protein. Dictyostelium cells in which the gene for RasGEFB was disrupted moved unusually rapidly, but lost the ability to perform macropinocytosis and therefore to grow in liquid medium. Crowns, the sites of macropinocytosis, were replaced by polarised lamellipodia. Mutant cells were also profoundly defective in early development, although they eventually formed tiny but normally proportioned fruiting bodies. This defect correlated with loss of discoidin Igamma mRNA, a starvation-induced gene, although other genes required for development were expressed normally or even precociously. RasGEFB was able to rescue a Saccharomyces CDC25 mutant, indicating that it is a genuine GEF for Ras proteins. CONCLUSIONS: RasGEFB appears to be the principal activator of the RasS protein, which regulates macropinocytosis and cell speed, but it also appears to regulate one or more other Ras proteins. PMID- 11102805 TI - Inhibition of PDK-1 activity causes a reduction in cell proliferation and survival. AB - 3-Phosphoinositide-dependent kinase-1 (PDK-1) was identified by its ability to phosphorylate and activate protein kinase B (PKB) in vitro [1,2] and can phosphorylate and activate additional protein kinases in the AGC family in vitro [3-6]. Its role in vivo has, however, only begun to be addressed. We used antisense oligonucleotides directed against PDK-1 expression to explore the role of PDK-1 in human glioblastoma cells (U87-MG), which express a mutant PTEN allele. Reduction in PDK-1 levels resulted in inhibition of PKB activity, and a reduction in phosphorylation on Thr308 and Ser473 of PKB. p70 S6 kinase (p70(S6K)) activity was also reduced. Cell proliferation was dramatically inhibited following treatment with PDK-1 antisense oligonucleotides, due to a combination of decreased cell doubling and an increase in apoptosis. This is in contrast to direct inhibition of phosphoinositide 3-OH kinase (PI 3-kinase), which results in G1 arrest with no effect on apoptosis. This study confirms both PKB and p70(S6K) as in vivo substrates for PDK-1. The effect of acute PDK-1 loss on cell proliferation and survival suggests the involvement of PI 3-kinase dependent and independent signaling events, and implicates PDK-1 as a potential therapeutic target for human neoplasms. PMID- 11102806 TI - Chaperones that cure yeast artificial [PSI+] and their prion-specific effects. AB - The [PSI(+)] nonsense-suppressor determinant of Saccharomyces cerevisiae results from the ability of Sup35 (eRF3) translation termination factor to undergo prion like aggregation [1]. Although this process is autocatalytic, in vivo it depends on the chaperone Hsp104, whose lack or overexpression can cure [PSI(+)] [2]. Overproduction of the chaperone protein Ssb1 increased the [PSI(+)] curing by excess Hsp104, although it had no effect on its own, and excess chaperone protein Ssa1 protected [PSI(+)] against Hsp104 [3,4]. We used an artificial [PSI(+)(PS)] based on the Sup35 prion-forming domain from yeast Pichia methanolica [5] to find other prion-curing factors. Both [PSI(+)(PS)] and [PSI(+)] have prion 'strains', differing in their suppressor efficiency and mitotic stability. We show that [PSI(+)(PS)] and a 'weak' strain of [PSI(+)] can be cured by overexpression of chaperones Ssa1, Ssb1 and Ydj1. The ability of different chaperones to cure [PSI(+)(PS)] showed significant prion strain specificity, which could be related to variation in Sup35 prion structure. Our results imply that homologs of these chaperones may be active against mammalian prion and amyloid diseases. PMID- 11102807 TI - Asymmetric stem-cell divisions define the architecture of human oesophageal epithelium. AB - In spite of its clinical importance, little is known about the stem-cell compartment of the human oesophageal epithelium [1,2]. The epithelial basal layer consists of two distinct zones, one overlying the papillae of the supporting connective tissue (PBL) and the other covering the interpapillary zone (IBL) [3]. In examining the oesophageal basal layer, we found that proliferating cells were rare in the IBL and a high proportion of mitoses were asymmetrical, giving rise to one basal daughter and one suprabasal, differentiating daughter. In the PBL, mitoses were more frequent and predominantly symmetrical. The IBL was characterised by low expression of ?1 integrins and high expression of the beta2 laminin chain. By combining fluorescence-activated cell sorting (FACS) with in vitro clonal analysis, we obtained evidence that the IBL is enriched for stem cells. A normal oesophageal epithelium with asymmetric divisions was reconstituted on denuded oesophageal connective tissue. In contrast, asymmetric divisions were not sustained on skin connective tissue, and the epithelium formed resembled epidermis. We propose that stem cells located in the IBL give rise to differentiating daughters through asymmetric divisions in response to cues from the underlying basement membrane. Until now, stem-cell fate in stratified squamous epithelia was believed to be achieved largely through populational asymmetry [4-6]. PMID- 11102808 TI - The SM19 gene, required for duplication of basal bodies in Paramecium, encodes a novel tubulin, eta-tubulin. AB - The discovery of delta-tubulin, the fourth member of the tubulin superfamily, in Chlamydomonas [1] has led to the identification in the genomes of vertebrates and protozoa of putative delta homologues and of additional tubulins, epsilon and zeta [2-4]. These discoveries raise questions concerning the functions of these novel tubulins, their interactions with microtubule arrays and microtubule organising centres, and their evolutionary status. The sm19-1 mutation of Paramecium specifically inhibits basal body duplication [5] and causes delocalisation of gamma-tubulin, which is also required for basal body duplication [6]. We have cloned the SM19 gene by functional complementation and found that it encodes another new member of the tubulin superfamily. SM19p, provisionally called eta-tubulin (eta-tubulin), shows low sequence identity with the tubulins previously identified in Paramecium, namely, alpha [7], beta [8], gamma [6], delta (this work) and epsilon (P. Dupuis-Williams, personal communication). Phylogenetic analysis indicated that SM19p is not consistently grouped with any phylogenetic entity. PMID- 11102809 TI - An fMRI study of the selective activation of human extrastriate form vision areas by radial and concentric gratings. AB - The ventral form vision pathway of the primate brain comprises a sequence of areas that include V1, V2, V4 and the inferior temporal cortex (IT) [1]. Although contour extraction in the V1 area and responses to complex images, such as faces, in the IT have been studied extensively, much less is known about shape extraction at intermediate cortical levels such as V4. Here, we used functional magnetic resonance imaging (fMRI) to demonstrate that the human V4 is more strongly activated by concentric and radial patterns than by conventional sinusoidal gratings. This is consistent with global pooling of local V1 orientations to extract concentric and radial shape information in V4. Furthermore, concentric patterns were found to be effective in activating the fusiform face area. These findings support recent psychophysical [2,3] and physiological [4,5] data indicating that analysis of concentric and radial structure represents an important aspect of processing at intermediate levels of form vision. PMID- 11102810 TI - The telomerase reverse transcriptase is limiting and necessary for telomerase function in vivo. AB - Mammalian telomerase is essential for the maintenance of telomere length [1-5]. Its catalytic core comprises a reverse transcriptase component (TERT) and an RNA component. While the biochemical role of mammalian TERT is well established [6 11], it is unknown whether it is sufficient for telomere-length maintenance, chromosome stability or other cellular processes. Cells from mice in which the mTert gene had been disrupted showed progressive loss of telomere DNA, a phenotype similar to cells in which the gene encoding the telomerase RNA component (mTR) has been disrupted [1,12]. On prolonged growth, mTert-deficient embryonic stem (ES) cells exhibited genomic instability, aneuploidy and telomeric fusions. ES cells heterozygous for the mTert disruption also showed telomere attrition, a phenotype that differs from heterozygous mTR cells [12]. Thus, telomere maintenance in mammals is carried out by a single, limiting TERT. PMID- 11102811 TI - The Drosophila RAD21 cohesin persists at the centromere region in mitosis. AB - 'Cohesin' is a highly conserved multiprotein complex thought to be the primary effector of sister-chromatid cohesion in all eukaryotes. Cohesin complexes in budding yeast hold sister chromatids together from S phase until anaphase, but in metazoans, cohesin proteins dissociate from chromosomes and redistribute into the whole cell volume during prophase, well before sister chromatids separate (reviewed in [1,2]). Here we address this apparent anomaly by investigating the cell-cycle dynamics of DRAD21, the Drosophila orthologue of the Xenopus XRAD21 and Saccharomyces cerevisiae Scc1p/Mcd1p cohesins [3]. Analysis of DRAD21 in S2 Drosophila tissue culture cells and live embryos expressing a DRAD21-green fluorescent protein (GFP) fusion revealed the presence of four distinct subcellular pools of DRAD21: a cytoplasmic pool; a chromosome-associated pool which dissociates from chromatin as chromosomes condense in prophase; a short lived centrosome-associated pool present during metaphase-anaphase; and a centromere-proximal pool which remains bound to condensed chromosomes, is found along the junction of sister chromatids between kinetochores, and persists until the metaphase-anaphase transition. We conclude that in Drosophila, and possibly all metazoans, a minor pool of cohesin remains bound to centromere-proximal chromatin after prophase and maintains sister-chromatid cohesion until the metaphase-anaphase transition. PMID- 11102813 TI - Depletion of a Cks homolog in C. elegans embryos uncovers a post-metaphase role in both meiosis and mitosis. AB - In eukaryotic cells, the key regulators of cell-cycle transitions are the cyclin dependent kinases (CDKs). The best studied CDK is a component of the M-phase promoting factor (MPF), which promotes entry into and progression through meiosis and mitosis. One of the enduring mysteries of the MPF complex has been the role of Cks/Suc1, a highly conserved member of the cell-cycle machinery in eukaryotes [1,2]. Cks has been proposed to be involved in activation of MPF [3], general interactions of MPF with its mitotic substrates [4] and/or inactivation of MPF [5,6]. We identified two Cks homologs in the genome of Caenorhabditis elegans and used RNA-mediated interference (RNAi) to investigate their roles in development. Whereas cks-2(RNAi) embryos display no apparent defects, cks-1(RNAi) embryos display defects in both meiosis and mitosis. Specifically, cks-1(RNAi) embryos fail to exit M phase properly. We propose that CKS-1 has an essential role in the inactivation of MPF during early C. elegans embryogenesis. PMID- 11102812 TI - Differentiated neurons retain the capacity to generate axons from dendrites. AB - Cutting the axon of a morphologically polarized neuron (stage 3) close to the cell body causes another neurite to grow as an axon [1-3]. Stage 3 neurons still lack molecular segregation of axonal and dendritic proteins, however. Axonal and dendritic compartments acquire their distinct composition at stage 4 (4-5days in culture), when proteins such as the microtubule-associated protein 2 (MAP-2) and the glutamate receptor subunit GluR1 localize to the dendrites and disappear from the axon [4,5]. We investigated whether cultured hippocampal neurons retained axon/dendrite plasticity after axons and dendrites have created their distinct cytoskeletal architecture and acquired their specific membrane composition. We found that axotomy of stage 4 neurons transformed a dendrite into an axon. Using axonal and dendritic markers, we tested whether cytoskeletal changes could cause similar transformations, and found that actin depolymerization induced multiple axons in unpolarized neurons. Moreover, depletion of actin filaments from both morphologically and molecularly polarized cells also resulted in the growth of multiple axons from pre-existing dendrites. These results imply that dendrites retain the potential to become axons even after molecular segregation has occurred and that the dendritic fate depends on the integrity of the actin cytoskeleton. PMID- 11102814 TI - Direct evidence from parietal extinction of enhancement of visual attention near a visible hand. AB - Brain areas exist that appear to be specialized for the coding of visual space surrounding the body (peripersonal space). In marked contrast to neurons in earlier visual areas, cells have been reported in parietal and frontal lobes that effectively respond only when visual stimuli are located in spatial proximity to a particular body part (for example, face, arm or hand) [1-4]. Despite several single-cell studies, the representation of near visual space has scarcely been investigated in humans. Here we focus on the neuropsychological phenomenon of visual extinction following unilateral brain damage. Patients with this disorder may respond well to a single stimulus in either visual field; however, when two stimuli are presented concurrently, the contralesional stimulus is disregarded or poorly identified. Extinction is commonly thought to reflect a pathological bias in selective vision favoring the ipsilesional side under competitive conditions, as a result of the unilateral brain lesion [5-7]. We examined a parietally damaged patient (D.P.) to determine whether visual extinction is modulated by the position of the hands in peripersonal space. We measured the severity of visual extinction in a task which held constant visual and spatial information about stimuli, while varying the distance between hands and stimuli. We found that selection in the affected visual field was remarkably more efficient when visual events were presented in the space near the contralesional finger than far from it. However, the amelioration of extinction dissolved when hands were covered from view, implying that the effect of hand position was not mediated purely through proprioception. These findings illustrate the importance of the spatial relationship between hand position and object location for the internal construction of visual peripersonal space in humans. PMID- 11102816 TI - First DNA malaria vaccine on trial in Africa. PMID- 11102818 TI - Gaur power. PMID- 11102819 TI - Root development. PMID- 11102820 TI - Sister chromatid separation: falling apart at the seams. AB - Cohesion between sister chromatids must be dissolved at the time of chromosome segregation. Recent studies reveal that the principles of cohesion dissolution in mitosis and meiosis are the same, but that there are important differences that stem from the distinct natures of these two processes. PMID- 11102821 TI - Embryonic polarity: a role for microtubules. AB - Researchers have suspected that initial polarization of the Caenorhabditis elegans embryo might be directed by microtubules, but demonstrating this has faced obstacles. A new study has cleverly bypassed these obstacles. PMID- 11102822 TI - Microbial pathogenesis: lipid rafts as pathogen portals. AB - The route of initial entry influences how host cells respond to intracellular pathogens. Recent studies have demonstrated that a wide variety of pathogens target lipid microdomains in host cell membranes, known as lipid rafts, to enter host cells as an infectious strategy. PMID- 11102823 TI - Plant stem cells: the only constant thing is change. AB - Recent studies in Arabidopsis have uncovered a negative feedback loop that couples the antagonistic functions of the WUSCHEL and CLAVATA loci to control stem cell fate in the shoot apical meristem. Abundance of the CLAVATA3 protein limits signaling through this pathway. PMID- 11102824 TI - Insect metamorphosis: out with the old, in with the new. AB - During insect metamorphosis, the steroid hormone ecdysone activates programmed cell death of larval tissues and the further development of adult tissues. Recent studies suggest that the E93 gene is both necessary and sufficient to target tissues for ecdysone-induced apoptosis. PMID- 11102825 TI - Evolutionary genetics: what is driving male mutation? AB - In mammals, most new mutations occur in males. But a study of the evolution of a human X to Y chromosomal translocation has revealed a sex bias much lower than previous estimates. Patterns of substitution suggest that differential methylation between male and female germ lines is a key determinant of the mutation rate. PMID- 11102826 TI - Visual perception: monkeys see things our way. AB - Neural mechanisms of visual perception can be studied in detail only in non-human animals. But recent work in humans has revealed a striking functional homology between the human and monkey visual systems, confirming the relevance of animal data and establishing a paradigm for cross-species studies of brain function. PMID- 11102827 TI - Antigen presentation: peptides and proteins scramble for the exit. AB - The fate of peptides that fail to bind to major histocompatibility complex class I molecules in the endoplasmic reticulum (ER)has remained unclear. A recent study has revealed that these peptides exit the ER via the Sec61 channel and compete for this pathway with misfolded proteins. PMID- 11102828 TI - Vertebrate development: the fast track to nodal signalling. AB - The transcription factors Fast-1 and Mixer are important terminal components of Smad2-mediated TGF-beta signal transduction. Recent studies demonstrate that Fast 1 and Mixer play critical roles in the formation of endoderm and dorsal mesoderm in zebrafish. PMID- 11102829 TI - Purification and characterisation of a non-plant myrosinase from the cabbage aphid Brevicoryne brassicae (L.). AB - Plant myrosinases and glucosinolates constitute a defence system in cruciferous plants towards pests and diseases. We have purified for the first time a non plant myrosinase from the cabbage aphid Brevicoryne brassicae (L.) to homogeneity. The protein was N-terminally blocked and protease (trypsin and lys c) degradation gave peptides of which five were sequenced. The protein is a dimer with subunits of mass 54 kDa+/-500 Da. Western blot analysis with an anti-aphid myrosinase antibody showed a strong cross reaction with a protein extract from the Brassica specialist, B. brassicae. The anti-aphid myrosinase antibody does not cross react with plant myrosinase neither does an anti-plant myrosinase antibody cross react with aphid myrosinase. PMID- 11102831 TI - The V410M mutation associated with pyrethroid resistance in Heliothis virescens reduces the pyrethroid sensitivity of house fly sodium channels expressed in Xenopus oocytes. AB - Some strains of Heliothis virescens carry a novel sodium channel mutation, corresponding to the replacement of Val410 by Met (designated V410M) in the house fly Vssc1 sodium channel, that is genetically and physiologically associated with pyrethroid resistance. To test the functional significance of this mutation, we created a house fly Vssc1 sodium channel containing the V410M mutation by site directed mutagenesis, expressed wildtype and specifically mutated sodium channels in Xenopus laevis oocytes, and evaluated the effects of the V410M mutation on the functional and pharmacological properties of the expressed channels by two electrode voltage clamp. The V410M mutation caused depolarizing shifts of approximately 9mV and approximately 5mV in the voltage dependence of activation and steady-state inactivation, respectively, of Vssc1 sodium channels. The V410M mutation also reduced the sensitivity of Vssc1 sodium channels to the pyrethroid cismethrin at least 10-fold and accelerated the decay of cismethrin-induced sodium tail currents. The degree of resistance conferred by the V410M mutation in the present study is sufficient to account for the degree of pyrethroid resistance in H. virescens that is associated with this mutation. Although Val410 is located in a sodium channel segment identified as part of the binding site for batrachotoxin, the V410M mutation did not alter the sensitivity of house fly sodium channels to batrachotoxin. The effects of the V410M mutation on the voltage dependence and cismethrin sensitivity of Vssc1 sodium channels were indistinguishable from those caused by another sodium channel point mutation, replacement of Leu1014 by Phe (L1014F), that is the cause of knockdown resistance to pyrethroids in the house fly. The positions of the V410M and L1014F mutations in models of the tertiary structure of sodium channels suggest that the pyrethroid binding site on the sodium channel alpha subunit is located at the interface between sodium channel domains I and II. PMID- 11102830 TI - Lipid storage and mobilization in insects: current status and future directions. AB - In this paper we review the current status of research on fatty acid absorption and conversion to diacylglycerol in the midgut. We further discuss how diacylglycerol may leave the midgut and associate with lipophorin in hemolymph. We review the present understanding of the role of the lipid transfer particle and lipophorin receptors in lipid delivery between lipophorin and tissues. Finally, we discuss recent studies on the mobilization of diacylglycerol from the fat body in response to adipokinetic hormone. Several suggestions for exciting areas of future research are described. PMID- 11102832 TI - Sulfated glycosaminoglycans from ovary of Rhodnius prolixus. AB - We have characterized sulfated glycosaminoglycans from ovaries of the blood sucking insect Rhodnius prolixus, and determined parameters of their synthesis and distribution within this organ by biochemical and histochemical procedures. The major sulfated glycosaminoglycan is heparan sulfate while chondroitin 4 sulfate is a minor component. These glycosaminoglycans are concentrated in the ovarian tissue and are not found inside the oocytes. Besides this, we detected the presence of a sulfated compound distinguished from sulfated glycosaminoglycans and possibly derived from sulfated proteins. Conversely to the compartmental location of sulfated glycosaminoglycans, the unidentified sulfated compound is located in the ovarian tissue as well as inside the oocytes. Based on these and other findings, the possible roles of ovarian sulfated glycosaminoglycans on the process of oogenesis in these insects are discussed. PMID- 11102833 TI - Digestion of legume starch granules by larvae of Zabrotes subfasciatus (Coleoptera: bruchidae) and the induction of alpha-amylases in response to different diets. AB - Zabrotes subfasciatus larvae possess three alpha-amylase isoforms as determined by in gel assays following SDS-PAGE. The two minor isoforms present lower electrophoretic mobility than the major form, and seem to occur as a heterodimer. When developed inside Vigna unguiculata (cowpea) seeds, fourth instar larvae have minor quantities of the slow-migrating forms, but when reared on seeds of Phaseolus vulgaris (common bean) or Phaseolus lunatus, the two slow-migrating forms are expressed in higher amounts, while activity of the major form was independent of the host seed. Larvae developing inside cowpea seeds at the beginning of the fourth instar were fed on flour from cotyledons of cowpea or common bean. Larvae fed on the common bean flour started to express the dimer in higher amounts when compared with the control larvae fed on cowpea flour. In an attempt to correlate differences between starch granules and the induction of alpha-amylases, a detailed study on the digestive process of the granules was conducted. Incorporation of purified starch granules into artificial diets did not induce the two minor alpha-amylases. The in vitro hydrolysis rates of purified granules and the pattern of dextrins liberated by the different alpha amylases were similar for the two legume species. The starch granules enter the midgut extensively damaged, which may facilitate the access to the more susceptible parts of the granules to enzymatic attack. PMID- 11102835 TI - A new form of arthropod phenoloxidase is abundant in venom of the parasitoid wasp Pimpla hypochondriaca. AB - We have recently identified phenoloxidase (PO) activity among several biologically active factors in venom from the parasitoid wasp Pimpla hypochondriaca. We have now isolated three genes, designated POI, POII and POIII, from a cDNA library made from venom-producing glands and found that their products are related to pro-phenoloxidases (PPOs), which are expressed as proenzymes in haemocytes and which mediate immune processes in arthropods. This is the first report of PO as a venom constituent. Amino acid sequence comparisons between the three Pimpla POs and PPOs revealed several notable differences, including the absence of sequences which specify the site of proteolytic activation in insect PPOs and the unprecedented occurrence of signal peptide sequences. NH(2)-terminal amino acid analysis of PO purified from venom yielded a peptide sequence matching the predicted mature NH(2) termini of POI and POII, confirming the authenticity of the signal peptide and indicating that proteolytic processing, other than to remove the signal peptide, does not occur in the wasp. Expression of POI, analysed by Northern hybridization, was approximately uniform from the time of adult emergence to day 6 post-emergence, after which it declined. A novel means of host immune suppression, mediated by the unregulated activity of venom PO in the haemocoel, is proposed. PMID- 11102834 TI - Ligand blot analysis of juvenile hormone esterase binding proteins in Manduca sexta L. AB - Biotinylated recombinant juvenile hormone esterase (JHE) was used for ligand blotting of proteins from fat body tissue and pericardial athrocytes of Manduca sexta. Proteins were separated by SDS-polyacrylamide gel electrophoresis or by two-dimensional electrophoresis. Eight putative JHE binding proteins were detected in fat body tissue and in pericardial athrocytes of both M. sexta and Heliothis virescens. The predominant bands were 29, 72, 75, 125 and 240kDa, with minor bands at 50, 80 and 205kDa. All putative JHE binding proteins were present from the second through to the fifth instar larvae of M. sexta. On wide-range isoelectric focusing, the 29kDa JHE binding protein separated into three species with isoelectric points of 6.5, 6.6 and 6.8. Biotinylated-JHE did not bind recombinant M. sexta-derived juvenile hormone binding protein. The mutant JHE with mutations K29R and K524R binds weakly to the JHE binding protein P29, relative to binding of wild-type JHE [Shanmugavelu et al., J. Biol. Chem., 275 (2000) 1802-1806]. A similar reduction in binding was not seen for the 29kDa binding protein identified here in pericardial athrocytes by ligand blot. This result is discussed. PMID- 11102836 TI - Single amino acid changes outside the active site significantly affect activity of glutathione S-transferases. AB - Glutathione S-transferases (GSTs: E.C. 2.5.1.18) are a multigene family of multifunctional dimeric proteins that play a central role in detoxication. Four allelic forms of the mosquito Anopheles dirus GST, adGST1-1, were cloned, expressed and characterized. The one or two amino acid changes in each allelic form was shown to confer different kinetic properties. Based on an available crystal structure, several of the residue changes were not in the putative substrate-binding pocket. Modeling showed that these insect Delta class GSTs also possess a hydrophobic surface pocket reported for Alpha, Mu and Pi class GSTs. The atom movement after replacement and minimization showed an average atom movement of about 0.1 A for the 0 to 25 A distance from the alpha carbon of the single replaced residue. This does not appear to be a significant movement in a static modeled protein structure. However, 200-500 atoms were involved with movements greater than 0.2 A. Dynamics simulations were performed to study the effects this phenomenon would exert on the accessible conformations. The data show that residues affecting nearby responsive regions of tertiary structure can modulate enzyme specificities, possibly through regulating attainable configurations of the protein. PMID- 11102837 TI - Genomic organization and putative promoters of highly conserved glutathione S transferases originating by alternative splicing in Anopheles dirus. AB - The genomic DNA of a GST class I alternative splicing gene has been characterized from Anopheles dirus, a Thai malaria vector. This gene organization is highly conserved in An. dirus and Anopheles gambiae (aggst1alpha), with >80% nucleotide identity in the coding region. Their gene organization contains six exons for four mature GST transcripts, which share exon 1 and exon 2 but vary between four different exon 3 sequences (exon 3A-3D). The deduced amino acid sequence of the GST transcripts from these two genes also shows very high conservation, with 85 93% identity for each orthologous gene. Two putative promoters and possible regulatory elements were predicted by a combination of the TSSW and MatInspector programs. The Ad214 promoter is proposed to be involved in developmental stage regulation. The Ad2112 promoter would appear to respond to intra- or extracellular stimuli. These two Anopheline species appear to have diverged in the distant past based on gene neighbors and phylogenetic data, yet these GST genes are still conserved. Therefore it is highly probable that this GST gene organization has one or more important roles. PMID- 11102838 TI - In vitro degradation of the Neb-Trypsin modulating oostatic factor (Neb-TMOF) in gut luminal content and hemolymph of the grey fleshfly, Neobellieria bullata. AB - The unblocked hexapeptidic Trypsin Modulating Oostatic Factor of the fleshfly, an inhibitor of both trypsin and ecdysone biosynthesis, resists very well proteolytic breakdown by enzymes present in the lumen of the gut of previtellogenic fleshflies. However, when incubated in hemolymph of adult flies, females and males, its half-life time is a mere 0.5 min. In hemolymph of last instar larvae, this value increases to about 1.5 min. Whereas PMSF, a potent inhibitor of serine proteases has no effect, captopril and lisinopril, both known to be specific inhibitors of mammalian angiotensin I converting enzyme (ACE), effectively inhibit TMOF breakdown in fly hemolymph. Digestion of Neb-TMOF by recombinant Drosophila AnCE on itself results in identical degradation products as with total hemolymph. In both cases ESI-Qq-oa-Tof mass spectrometry demonstrated the appearance of peptide fragments with the sequences NPTN, LH and NP. These observations not only confirm the reported presence of circulating ACE like activity in flies but also strongly suggest that in flies this hemolymph ACE like activity might be involved in the regulation of the oostatic activity as exerted by Neb-TMOF. PMID- 11102840 TI - Special editor introduction. Surface electromyography. . PMID- 11102839 TI - Isolation and expression of the ecdysteroid-inducible angiotensin-converting enzyme-related gene in wing discs of Bombyx mori. AB - We isolated a clone encoding a putative angiotensin-converting enzyme-related gene from the wing disc cDNA library of the silkworm, Bombyx mori (refer to as BmAcer). The predicted open reading frame encoded 648 amino acids with about 50% identities with the Drosophila melanogaster angiotensin-converting enzyme Ance and Acer. Northern analysis identified a 2.2-kilobase mRNA which was abundant in wing discs two days after the beginning of wandering. An accumulation of the transcript was observed approximately 2 h after 20-hydroxyecdysone (20E) exposure in vitro and was blocked slightly by a protein synthetic inhibitor. These data suggest that the transcription of the BmAcer gene is directly 20E-inducible. PMID- 11102841 TI - Use of surface electromyography to estimate neck muscle activity. AB - This paper reviews the literature concerning the use of surface electromyography (sEMG) for the study of the neck musculature in response to work and workplace design during light work and semi-static tasks. The paper also draws upon basic research and biomechanical modeling in order to provide methodological recommendations for the use of surface electromyography in this region of the body and to identify areas which require further investigation. The paper includes review and discussion of electrode site location, methods of normalization, data reliability, and factors that can affect sEMG signals from this region, including noise, physiologic artifact, stress, visual deficiencies, and pain. General guidance for maximum exertions with the neck musculature, for sEMG normalization or other purposes, is also included. PMID- 11102842 TI - Differential control during maximal concentric and eccentric loading revealed by characteristics of the electromyogram. AB - Maximal eccentric loading has been associated with higher levels of spindle afferent activity but lower levels of integrated EMG as compared to maximal concentric loading. Elbow flexor EMG was recorded from 17 subjects during concentric (CONC) and eccentric (ECC) elbow flexion at 70 degrees s(-1) using a Kin-Com dynamometer. We hypothesized that peak EMG amplitude would be more sensitive to fluctuations in facilitation by the spindle primary afferents via the segmental stretch reflex pathway, and that the mean EMG would be more reflective of the ongoing level of muscle activation. A ratio of peak to mean EMG (P/M EMG ratio) was predicted to be larger during maximal eccentric loading than maximal concentric loading. The peak EMG (P<0.013) and the P/M EMG ratio (P<0.001) were significantly greater during the ECC condition than the CONC condition. In a subgroup of three subjects who underwent 3 weeks of eccentrically biased weight training, EMG, peak torque and torque variability were assessed before and after training. P/M EMG ratio decreased, while peak torque and torque variability increased following the training. Differences in the P/M EMG ratio appear to reflect differences in the way eccentric and concentric muscle actions are controlled and do not simply represent less control during the eccentric task. PMID- 11102843 TI - Coactivation during gait as an adaptive behavior after stroke. AB - The aims of the present study were to quantify the impairment in ankle coactivation on the paretic and non-paretic sides of subjects with hemiparesis and to examine the relationship of ankle coactivation with postural instability, motor deficit of the paretic lower extremity and locomotor performance. Electromyography of the medial gastrocnemius (MG) and tibialis anterior (TA) muscles were recorded bilaterally during gait in 30 subjects (62.1+/-9.9 years) who had suffered a recent stroke (<6 months) as well as on one side of 17 healthy controls (59.3+/-9.1 years) walking at very slow speed. Ankle muscle coactivation was calculated by dividing the time of overlap between MG and TA signals (threshold of 20 microV) by the duration of the gait phases of interest: stance, swing, first and second double support sub-phases and single support sub-phase. The time spent in single support and the peak plantarflexor moment of force on the paretic side were used to measure, respectively, postural stability and dynamic strength of the paretic plantarflexors. The subjects with hemiparesis demonstrated less coactivation on the paretic side during the single support sub phase (p<0.01) and more coactivation during first and second double support sub phases on the non-paretic side (p<0.001) compared to control values. The patients with coactivation patterns that differed the most from controls were the patients with the more severe impairments and disabilities. While the reduced coactivation on the paretic side may contribute to poor postural stability and poor locomotor performance, the presence of excessive coactivation on the non-paretic side when both limbs were in ground contact may be an adaptation to help maintain postural stability during gait. PMID- 11102844 TI - Neuromuscular responses to explosive and heavy resistance loading. AB - The EMG power spectrum may shift towards higher frequencies with higher movement velocities. Fatigue, on the other hand, can cause a decrease in the frequency components. The purpose of this study was to examine acute effects of explosive (EE) and heavy resistance (HRE) concentric leg press exercise on muscle force, EMG and blood lactate. The EE included five sets of ten repetitions with 40+/-6% of the isometric maximum at a 100 degrees knee angle performed as explosively as possible. The same number of repetitions was performed in HRE but with a heavier load (67+/-7% of the isometric maximum at a 100 degrees knee angle). Maximal isometric and single concentric actions of different loads, and an isometric fatigue test were measured before and after both exercises. Surface EMG was recorded from the vastus medialis muscles for analyses of average EMG (aEMG) and EMG power spectrum. Muscle fiber composition of the vastus lateralis was determined and blood lactate measured throughout the exercises. Mean power frequency and median frequency were higher during EE than during HRE (P<0.05). They increased during EE (P<0.05) as the exercise progressed, whereas during HRE no change or even slight decreases were observed. Signs of fatigue after pure concentric work were not observed after EE, and even after HRE, possibly due to the relatively small range of motion and short duration of action time, the fatigue was not that extensive. The relative number of fast twitch fibers was correlated (r=0.87, P<0.05) with the change in blood lactate in HRE. It was concluded that there may be a greater use of fast twitch motor units in explosive movements and that instead of fatigue, the present number of concentric actions in explosive exercise seems to have facilitated the neuromuscular system. PMID- 11102845 TI - Different neuromuscular recruitment patterns during eccentric, concentric and isometric contractions. AB - Aim. The purpose of this study was to determine the neuromuscular fatigue profiles during 100 s isometric (ISO), concentric (CON), and eccentric (ECC) activity.Methods. Twelve subjects (age 25.1+/-3.7 years, mass 70.1+/-8.2 kg, mean+/-SD) performed ISO, CON and ECC maximal voluntary contractions and 100 s endurance trials on an isokinetic dynamometer. Raw EMG data were recorded throughout each trial from the rectus femoris of the right limb. Corresponding data for integrated electromyography (IEMG), percentile frequency shifts (MPFS) and peak torque output were divided into five 5 s epochs and subsequently normalised with the first epoch being the reference point, in order to assess changes over time.Results. There were no significant differences between ECC, CON and ISO peak torque output (211+/-63 vs 169+/-41 vs 177+/-61 Nm; ECC, CON, ISO) and IEMG activity (280+/-143 vs 305+/-146 vs 287+/-143 mV; ECC, CON, ISO) during maximal contractions. Serial reductions in torque output were greatest in ISO in which torque output during the final epoch was 31+/-13% of initial values, similar to the final torque values in CON (58+/-15%), but significantly less than ECC (108.6+/-38.6%; P<0. 001) values. In CON and ECC, IEMG was maintained (95+/ 27% and 93+/-21%; CON and ECC), whereas IEMG for ISO decreased to 38+/-13% of initial values. The greatest reduction in MPFS occurred in CON (69+/-10%) compared to ISO (78+/-9%; P<0.05) and ECC (93+/-6%; P<0.001).Conclusion. These data demonstrate distinct neuromuscular fatigue profiles for the different types of muscle contraction. Whereas eccentric activity was largely fatigue resistant, isometric and concentric contractions displayed different neuromuscular fatigue profiles. PMID- 11102847 TI - The science of chemical discovery: probing the unknown with new technologies. PMID- 11102846 TI - Intensity analysis in time-frequency space of surface myoelectric signals by wavelets of specified resolution. AB - Surface myoelectric signals often appear to carry more information than what is resolved in root mean square analysis of the progress curves or in its power spectrum. Time-frequency analysis of myoelectric signals has not yet led to satisfactory results in respect of separating simultaneous events in time and frequency. In this study a time-frequency analysis of the intensities in time series was developed. This intensity analysis uses a filter bank of non-linearly scaled wavelets with specified time-resolution to extract time-frequency aspects of the signal. Special procedures were developed to calculate intensity in such a way as to approximate the power of the signal in time. Applied to an EMG signal the intensity analysis was called a functional EMG analysis. The method resolves events within the EMG signal. The time when the events occur and their intensity and frequency distribution are well resolved in the intensity patterns extracted from the EMG signal. Averaging intensity patterns from multiple experiments resolve repeatable functional aspects of muscle activation. Various properties of the functional EMG analysis were shown and discussed using model EMG data and real EMG data. PMID- 11102849 TI - Key strategies in functional genomics for drug discovery. PMID- 11102848 TI - Improved prospects for anti-cancer gene therapy. PMID- 11102850 TI - Pseudocomplementary strategy strengthens PNA therapeutic potential. PMID- 11102851 TI - Ion chelators for stroke. PMID- 11102852 TI - Dietary supplement to treat asthma. PMID- 11102854 TI - Quo vadis, biotech? (Part 1). AB - The recent increase in the capitalization value of the biotechnology industry appears to be sustainable. The phenomenon is interpreted as an acknowledgement by the markets that this industry has become the main source of innovation for the pharmaceutical industry. In addition, biotechnology is beginning to impact on other industries. However, from a methodological and strategic point of view, the biotechnology industry is still too fragmented. Consolidation along the lines of value generation in drug research and development, however, will occur. Over the next 5 to 10 years, the biotechnology industry will remain the fastest growing industry in the health care arena. PMID- 11102855 TI - A patent strategy for genomic and research tool patents: are there any differences between the USA, Europe and Japan? AB - The patenting of genomics and research tools, and its effects on the development of therapeutics are attracting considerable attention. Regardless of whether one is in favor of patents for this technology, it is not specifically excluded from patenting in most countries. Accordingly, it is imperative that a suitable global patent strategy be developed and followed to maximize both intellectual property rights and returns on R&D investment. PMID- 11102856 TI - Differential gene expression technologies for identifying surrogate markers of drug efficacy and toxicity. AB - Advances in the rapidly evolving discipline of pharmacogenomics have forced the biotechnology and pharmaceutical industries to integrate differential gene expression profiling into their drug discovery and development strategies. Here we highlight the use of differential gene expression technologies for the elucidation of both drug efficacy and toxicity as well as novel candidate genes for pharmacogenetic analyses to assess individual variability to drug response. This will include an overview of the different technologies created to facilitate pharmacogenomic analyses and to highlight advantages and disadvantages of these emerging methodologies. Two high-throughput differential gene expression technologies, microarrays and GeneCalling((R)), will be presented in detail. PMID- 11102857 TI - Monitor Editorship. PMID- 11102858 TI - Farewell. PMID- 11102859 TI - Monitor: molecules and profiles. AB - Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues. PMID- 11102860 TI - Combinatorial chemistry. PMID- 11102861 TI - Web alert. Biopolymers model systems. PMID- 11102862 TI - Biopolymers. Chemical and biological approaches for understanding form and function editorial overview PMID- 11102863 TI - Synthetically modified DNAs as substrates for polymerases. AB - DNA polymerase enzymes process their natural substrates with very high specificity. Yet recent experiments have shown that these enzymes can also process DNA in which the backbone or bases are modified to a surprising degree. Such experiments have important implications in understanding the mechanisms of DNA replication, and suggest important biotechnological uses as well. PMID- 11102864 TI - Self-assembled nanostructures based on DNA: towards the development of nanobiotechnology. AB - DNA is a promising construction material for the supramolecular 'bottom-up' engineering of artificial nanostructured devices. The use of DNA for the selective positioning of macromolecular components, the fabrication of nanostructured DNA scaffolds, as well as the DNA-templated synthesis of nanometer sized and mesoscopic complexes, consisting of inorganic and bioorganic compounds, are exciting areas of current research. PMID- 11102865 TI - Chemical and enzymatic synthesis of glycopolymers. AB - The development of efficient, fast, flexible and general synthetic routes to glycopolymers is an ongoing challenge and much progress has been made in recent years. Chemical coupling methods have become increasingly sophisticated to fine tune reactivity of reagents by fortuitous choices of anomeric activating group and protecting groups. As a result, oligosaccharide synthesis has become more predictable and reliable even to the extent that first examples of saccharide library syntheses in solution and on the solid phase have been published. In biology, the repertoire of biocatalysts that can be used for glycoside synthesis is ever-increasing, and enzyme-catalysed glycosylation steps have been successfully incorporated into synthetic strategies. PMID- 11102866 TI - Heparin and heparan sulfate: biosynthesis, structure and function. AB - Heparin and heparan sulfate glycosaminoglycans are acidic complex polysaccharides found on the cell surface and in the extracellular matrix. Recent progress has uncovered a virtual explosion of important roles of these biopolymers in fundamental biological processes. Advances in the understanding of biosynthesis and structure and the development of novel analytical methods for composition and sequence analysis have provided remarkable insights into structure/function relationships of these complex and once elusive polysaccharides. PMID- 11102867 TI - Dissecting and manipulating the pathway for glycosylphos-phatidylinositol-anchor biosynthesis. AB - The pathway for glycosylphosphatidylinositol-anchor biosynthesis consists of at least 10 reaction steps. Many of the genes encoding the enzymes and regulators involved in this pathway have been recently cloned and their products characterised. These studies have revealed the common and also different characteristics of glycosylphosphatidyl-inositol biosynthesis enzymes in different organisms, leading to the development of species-specific inhibitors of the pathway. PMID- 11102868 TI - Understanding peptide interactions with the lipid bilayer: a guide to membrane protein engineering. AB - In recent years, studies of the interactions of designed hydrophobic and amphipathic polypeptides with biological membranes have progressed considerably. In-plane and transmembrane helical domains have been engineered as well as sequences that exhibit dynamic distributions of different topologies. These sequences not only help our understanding of the thermodynamic interaction contributions that determine peptide orientation, but also exhibit interesting biological functions as autonomous units. In addition, helices are considered to be folding intermediates of multi-spanning membrane proteins. The specificity and thermodynamic stability of transmembrane helix-helix interactions or the assembly of beta sheets at the membrane surface have, therefore, become a focus of ongoing research. PMID- 11102869 TI - Unnatural amino acids as probes of protein structure and function. AB - Nonsense suppression methodology, for incorporating unnatural amino acids into proteins, has enabled a wide range of studies into protein structure and function using both in vitro and in vivo translation systems. Although methodological challenges remain, scores of unnatural amino acids have been employed that include both subtle and dramatic variants of the natural set. A number of insights that would not have been possible using conventional site-directed mutagenesis have been gained. PMID- 11102871 TI - Chemistry at the crossroads PMID- 11102870 TI - Bacterial adhesins: structural studies reveal chaperone function and pilus biogenesis. AB - During the past year, remarkable progress has been made in understanding how periplasmic chaperones fold and protect protein modules that are destined for assembly into adhesive pili in Gram-negative bacteria. The first two three dimensional structures of complexes of periplasmic chaperones with substrate pilus subunits have revealed much about the structural basis for chaperone mediated folding and aggregation prevention, and have provided insight into the structure of adhesive pili. PMID- 11102873 TI - Allosterically controllable ribozymes with biosensor functions. AB - The concept of allosteric regulation has already been exploited in the creation of artificial ribozymes and the activities of certain ribozymes can be controlled allosterically by specific effectors. Ribozymes with such properties are in the spotlight as biosensors. Such artificial allosterically regulated ribozymes have potential utility as nucleic-acid-based biosensors. PMID- 11102872 TI - Chemical strategies for the global analysis of protein function. AB - As the human genome project nears completion, biological research is entering a new era in which experimental focus will shift from identifying novel genes to determining the function of gene products. Rising to this challenge, several technologies have emerged that aim to characterise genes and/or proteins collectively rather than individually. Of particular interest is a new breed of strategies that employs synthetic chemistry to enrich our understanding of protein function on a global scale. PMID- 11102874 TI - RNA as a target for small molecules. AB - Proteins are folded to form a small binding site for catalysis or ligand recognition and this small binding site is traditionally the target for drug discovery. An alternative target for potential drug candidates is the translational process, which requires a precise reading of the entire mRNA sequence and, therefore, can be interrupted with small molecules that bind to mRNA sequence-specifically. RNA thus presents itself as a new upstream target for drug discovery because of the critical role it plays in the life of pathogens and in the progression of diseases. In this post-genomic era, RNA is becoming increasingly amenable to small-molecule therapy as greater structural and functional information accumulates with regard to important RNA functional domains. The study of aminoglycoside antibiotics and their binding to 16S ribosomal RNA has been a paradigm for our understanding of the ways in which small molecules can be developed to affect the function of RNA. PMID- 11102875 TI - Nitric oxide synthase: models and mechanisms. AB - The overproduction or underproduction of nitric oxide has been implicated in pathological symptoms such as endotoxic shock, diabetes, allograft rejection, and myocardial ischimia/reperfusion injury. A thorough understanding of the biosynthesis of nitric oxide is necessary to probe and manipulate these signaling events. There is also considerable pharmacological interest in developing selective inhibitors of the several isoforms of nitric oxide synthase. The recently determined crystal structures of complexes between nitric oxide synthase and substrate, the mechanisms of the enzymatic reaction that generate nitric oxide and chemical precedents and models for these reactions are now coming into focus, but there are still numerous fascinating and unanswered questions regarding nitric oxide biosynthesis. PMID- 11102876 TI - Synthetic multivalent ligands in the exploration of cell-surface interactions. AB - Processes such as cell-cell recognition and the initiation of signal transduction often depend on the formation of multiple receptor-ligand complexes at the cell surface. Synthetic multivalent ligands are unique probes of these complex cell surface-binding events. Multivalent ligands can be used as inhibitors of receptor ligand interactions or as activators of signal transduction pathways. Emerging from these complementary applications is insight into how cells exploit multivalent interactions to bind with increased avidity and specificity and how cell-surface receptor organization influences signaling and the cellular responses that result. PMID- 11102877 TI - Molecular transport and organization in supported lipid membranes. AB - The mechanism by which vesicles spontaneously form supported lipid bilayer membranes on glass surfaces is becoming better understood and this knowledge is the basis of a technology of patterning membrane arrays and controlling composition. Controlled interactions between supported membranes and cells, particularly from the immune system, provide direct insight into cell-cell surface interactions. PMID- 11102878 TI - Imprinted polymers with transition metal catalysts. AB - The microenvironment of immobilized transition metal catalysts can be manipulated using catalyst-substrate conjugates in combination with the technology of molecular imprinting. Recent results have shown that catalysts with a significantly enhanced activity combined with an improved substrate-, regio- and enantioselectivity can be prepared in this way. These artificial systems resemble metalloenzymes because the catalytic transformation is effectively controlled by a well-defined second coordination sphere. PMID- 11102879 TI - Molecular photonic and electronic circuitry for ultra-sensitive chemical sensors. AB - Molecular wires have progressed from an intellectual curiosity to become the basis for chemical sensors with unprecedented sensitivity. Particularly exciting opportunities are those that make use of biological superstructures to effect conduction through assemblies of molecular wires. PMID- 11102880 TI - A view on the science: physical anthropology at the millennium. PMID- 11102881 TI - Biomechanical analysis of vertical climbing in the spider monkey and the Japanese macaque. AB - Climbing is one of the most important components of primate locomotor modes. We previously reported that the kinesiological characteristics of vertical climbing by the spider monkey and Japanese macaque are clearly different, based on their kinetics and kinematics. In this study, a more detailed analysis using inverse dynamics was conducted to estimate the biomechanical characteristics of vertical climbing in the spider monkey and Japanese macaque. One of the main findings was the difference in forelimb use by the two species. The results of a joint moment analysis and estimates of muscular force indicate that the spider monkey uses its forelimbs to keep the body close to the substrate, rather than to generate propulsion. The forelimb of the Japanese macaque, on the other hand, likely contributes more to propulsion. This supports the idea that "forelimb-hindlimb differentiation" is promoted in the spider monkey. The estimated muscular force also suggests that the spider monkey type of climbing could develop the hindlimb extensor muscles, which are important in bipedal posture and walking. As a result, we conclude that the spider monkey type of climbing could be functionally preadaptive for human bipedalism. This type of climbing would develop the hip and knee extensor muscles, and result in more extended lower limb joints, a more erect trunk posture, and more functionally differentiated fore- and hindlimbs, all of which are important characteristics of human bipedalism. PMID- 11102882 TI - Fourier analysis of acetabular shape in native American Arikara populations before and after acquisition of horses. AB - The goal of this study was to identify changes in acetabular morphology associated with the use of horses by Native Americans. Previous studies reported "elongate" acetabula in horseback-riding members of the Omaha and Ponca populations. Such a difference in acetabular shape is a potentially useful osteological marker of habitual horseback riding. This report compares acetabula of adult males from two Native American Arikara populations known to have differed substantially in their use of horses. Population samples were from separate sites in South Dakota: Larson (nonriding) and Leavenworth (riding). Outlines of acetabular rims were digitized and analyzed, using a simplified 12 point Fourier analysis. A Fourier series with six terms accurately described acetabular shape. Significant differences (P<0.10) between riding and nonriding populations were observed in two Fourier coefficients. Acetabula of riding Arikara were found to have smaller B(4) coefficients (P = 0. 061) and more positive B(2) coefficients (P = 0.080), indicating expanded anterior-superior borders relative to acetabula of non-riding Arikara. PMID- 11102883 TI - Subadult health and disease in prehistoric Tonga, Polynesia. AB - This article focuses on the differential diagnosis of pathologic lesions recorded on the limbs and crania of 17 subadults from two pre-European burial mounds in Tonga, western Polynesia. All affected subadults were between the ages of 6 months and 3 years at death. The lesions described consist primarily of subperiosteal new bone deposition on the limbs and endocranial surface. However, the presence of cribra orbitalia in a number of individuals indicates concurrent iron-deficiency anaemia. A differential diagnosis of haematogenous osteomyelitis, congenital syphilis, yaws, scurvy, hypervitaminosis A, trauma, Caffey's disease, and iron-deficiency anaemia is discussed. It was concluded that the most likely cause for the lesions observed is a synergistic relation between infection (weanling diarrhoea, yaws) and metabolic disease (scurvy and possibly hypervitaminosis A). Trauma is not ruled out as contributing to the development of some pathologic lesions. It is concluded that, in the Pacific Islands at least, multiple causes for skeletal pathology in subadults should be considered rather than a single aetiology. PMID- 11102884 TI - Body mass prediction from skeletal frame size in elite athletes. AB - Body mass can be estimated from measures of skeletal frame size (stature and bi iliac (maximum pelvic) breadth) fairly accurately in modern human populations. However, it is not clear whether such a technique will lead to systematic biases in body mass estimation when applied to earlier hominins. Here the stature/bi iliac method is tested, using data available for modern Olympic and Olympic caliber athletes, with the rationale that these individuals may be more representative of the general physique and degree of physical conditioning characteristic of earlier populations. The average percent prediction error of body mass among both male and female athletes is less than 3%, with males slightly underestimated and females slightly overestimated. Among males, the ratio of shoulder to hip (biacromial/bi-iliac) breadth is correlated with prediction error, while lower limb/trunk length has only a weak inconsistent effect. In both sexes, athletes in "weight" events (e.g. , shot put, weight lifting), which emphasize strength, are underestimated, while those in more endurance-related events (e.g., long distance running) are overestimated. It is likely that the environmental pressures facing earlier hominins would have favored more generalized physiques adapted for a combination of strength, speed, agility, and endurance. The events most closely approximating these requirements in Olympic athletes are the decathlon, pentathlon, and wrestling, all of which have average percent prediction errors of body mass of 5% or less. Thus, "morphometric" estimation of body mass from skeletal frame size appears to work reasonably well in both "normal" and highly athletic modern humans, increasing confidence that the technique will also be applicable to earlier hominins. PMID- 11102885 TI - Proceedings of the sixty-ninth meeting of the american association of physical anthropologists PMID- 11102886 TI - American association of physical anthropologists constitution and By-laws PMID- 11102887 TI - American Association of Physical Anthropologists. Membership list. PMID- 11102888 TI - AJPA reviewers during 1999 PMID- 11102889 TI - K-ras oncogene subtype mutations are associated with survival but not expression of p53, p16(INK4A), p21(WAF-1), cyclin D1, erbB-2 and erbB-3 in resected pancreatic ductal adenocarcinoma. AB - Previous studies of molecular prognostic markers following resection for exocrine pancreatic cancer have produced conflicting results. Our aim was to undertake a comprehensive analysis of potentially useful molecular markers in a large, multicentre patient population and to compare these markers with standard pathological prognostic variables. Formalin-fixed, paraffin-embedded specimens of pancreatic ductal adenocarcinoma were analysed from 157 patients [100 men and 57 women with a median (range) age of 60 (33-77) years] who had undergone pancreatectomy. Immunohistochemistry was used to detect expression of p16(INK4), p53, p21(WAF1), cyclin D1, erbB-2 and erbB-3. Mutations in codons 12 and 13 of the K-ras oncogene were detected by SSCP and sequencing following DNA extraction and amplification by PCR. The median (range) survival post-resection was 12.5 (3 83) months. Abnormalities of p16(INK4), p53, p21(WAF1), cyclin D1, erbB-2 and erbB-3 expression were found in 87%, 41%, 75%, 72%, 33% and 57% of cases, respectively. There was no significant correlation between expression of any of these markers and patient survival. K-ras mutations were found in 73 (75%) of 97 cases with amplifiable DNA. The presence of K-ras mutation alone did not correlate with survival, but there were significant differences in survival according to the type of K-ras mutation (p = 0.0007). Reduced survival was found in patients with GaT, cGT and GcT K-ras mutations compared to GtT, aGT and GaC mutations. In conclusion, survival was associated with type of K-ras mutation but not expression of p16(INK4), p53, p21(WAF1), cyclin D1, erbB-2 and erbB-3. PMID- 11102890 TI - Correlation of total VEGF mRNA and protein expression with histologic type, tumor angiogenesis, patient survival and timing of relapse in non-small-cell lung cancer. AB - We have quantified the expression of all 4 isoforms of vascular endothelial growth factor (VEGF) mRNA in non-small-cell lung cancer (NSCLC) using a new kinetic quantitative PCR method, real-time quantitative (RTQ) RT-PCR, and investigated the association between VEGF expression at the mRNA and protein levels and the clinicopathologic variables, tumor angiogenesis, patient survival and timing of relapse. Surgical tumor specimens from 72 NCSLC patients (37 squamous-cell carcinomas, 35 adenocarcinomas) were examined. Twenty-eight patients had stage I, 10 stage II and 34 stage IIIA or IIIB disease. Total VEGF mRNA (all 4 isoforms) was quantified by RTQ RT-PCR, while VEGF protein expression and microvessel number in tumors were assessed immunohistochemically. VEGF mRNA was detected in all 72 tumor samples at significantly higher levels than in adjacent normal tissue. Tumoral VEGF mRNA levels correlated strongly with the VEGF protein staining score and microvessel count. Adenocarcinomas showed significantly higher VEGF mRNA expression and a higher protein staining score than squamous-cell carcinomas. High tumoral VEGF mRNA expression was associated with advanced (IIIA or IIIB) tumor stage, lymph node metastasis, high tumoral microvessel counts, short patient survival (<24 months) and early relapse (<12 months), while a high VEGF protein staining score was associated with high tumoral microvessel counts, short patient survival and early relapse. Patients with high tumoral levels of both VEGF mRNA and protein had significantly shorter survival and earlier relapse. In multivariate analysis, the VEGF protein staining score and nodal status were the most important independent predictors of survival and recurrence. We conclude that RTQ RT-PCR is a sensitive method for detecting and quantifying VEGF mRNA expression in NSCLC and that the expression levels of total VEGF mRNA and protein in NSCLC are strongly associated with histologic type, tumor angiogenesis, survival and timing of relapse. High VEGF expression in adenocarcinomas may contribute to their greater metastatic potential. PMID- 11102891 TI - Co-expression of epidermal growth factor receptor and transforming growth factor alpha predicts worse prognosis in breast-cancer patients. AB - Epidermal growth factor receptor (EGF-R) and its ligand, transforming growth factor-alpha (TGF-alpha), play an important role through the autocrine growth regulation system in several human cancers, including breast cancer. However, the clinical significance of co-expression of EGF-R and TGF-alpha has not been elucidated. One hundred seventy-three female patients diagnosed as invasive ductal carcinoma who had undergone a mastectomy (159 patients) or breast conserving surgery (14 patients) were followed up for 81 to 119 months (median 94 months) post-operatively. Immunoreactivity for EGF-R, TGF-alpha, p53 and c-erbB-2 with paraffin-embedded carcinoma tissue was investigated using labeled streptavidin-biotin methods. Positive rates of carcinoma cells were 27%, 33%, 32% and 26% for EGF-R, TGF-alpha, p53 and c-erbB-2, respectively. Expression of EGF-R only was observed in 16% (28/173), of TGF-alpha only in 22% (38/173), of both EGF R and TGF-alpha in 11% (19/173) and of neither in 51% (88/173). By univariate analysis, significant differences in overall survival and disease-free survival were noted according to the co-expression of EGF-R and TGF-alpha (p< 0.0001, p<0.0001), co-expression of EGF-R and c-erbB-2 (p = 0.0029, p = 0.0028), nodal status (p = 0.0028, p = 0.0001), tumor size (p = 0.0001, p<0.0001) and c-erbB-2 expression (p = 0.0034, p = 0.018), respectively. The status of p53 expression (p = 0.01), estrogen receptor (p = 0.042) and progesterone receptor (p = 0.046) showed significant differences in overall survival. According to Cox's multivariate analysis, co-expression of EGF-R and TGF-alpha had the most significant effect on disease-free survival (p<0.0001) and overall survival (p<0.0001), followed by nodal status. Co-expression of EGF-R and TGF-alpha by immunohistochemical detection is an independent prognostic indicator, and it may be helpful for determining the group of breast-cancer patients with an aggressive phenotype. PMID- 11102892 TI - Expression and prognostic significance in lung cancer of human tumor-associated antigen RCAS1. AB - A new monoclonal antibody (MAb), 22-1-1, acts against a novel tumor-associated antigen (Ag) strongly expressed in human uterine cervical adenocarcinomas. A cDNA encoding the Ag recognized by the 22-1-1 MAb has been isolated and called RCAS1 (receptor-binding cancer antigen expressed on SiSo cells). RCAS1 can induce growth arrest and apoptosis in RCAS1 receptor-positive cells including T cells and natural killer cells in vitro. These results suggest that RCAS1 is involved in tumor escape from the immune system. Immunohistochemical analysis revealed the relationship between RCAS1 expression and clinicopathological variables (age, sex, smoking, histology, differentiation grade, pathological T factor, N factor and stage) and the prognostic significance of RCAS1 in 66 lung-cancer patients who underwent curative operations: 33 adenocarcinomas, 24 squamous-cell carcinomas, 3 large-cell carcinomas, 4 adenosquamous carcinomas and 2 small-cell carcinomas. Median follow-up period of 64 non-small-cell carcinomas (NSCLCs) was 67.4 months. RCAS1 was expressed in 74.2% of lung cancers. RCAS1 in NSCLC cases with advanced T factor or pathological stage or in poorly differentiated adenocarcinomas was highly expressed. Furthermore, RCAS1 expression inducing apoptosis of tumor-infiltrating lymphocytes was a significant prognostic factor in NSCLC (p<0.03). PMID- 11102893 TI - Nuclear morphometry in male breast carcinoma: association with cell proliferative activity, oncogene expression, DNA content and prognosis. AB - To investigate the prognostic value of nuclear morphometry in male breast carcinoma (MBC), histological samples from 50 patients (mean age 62.2 years) were retrospectively analyzed by computerized nuclear morphometry. All patients received surgery; 35 had multiple combinations of adjuvant therapies. Mean follow up was 67 months (range 1-230). In each case, 100 tumor cells were measured, and the mean nuclear area (MNA), standard deviation of the nuclear area (SDNA), mean nuclear perimeter (MNP), standard deviation of the nuclear perimeter (SDNP) and shape factor (SHF) were calculated. Morphometric features were compared with tumor histological grade, size, nodal status, DNA ploidy evaluated by flow cytometry and cell proliferative activity assessed by the quantity of argyrophilic nucleolar organizer region-associated proteins (AgNORs), monoclonal antibody (MAb) PC10 against proliferating cell nuclear antigen and MAb MIB-1. Comparison was also made with the immunohistochemical detection of p53, bcl-2, c erbB-2 and c-myc proteins. Significant association was found between nuclear morphometric parameters and tumor grade, DNA content and cell proliferation indices. SDNA was greater in p53-positive and bcl-2-negative cases; SDNP was greater in p53-positive cases; SHF was lower in p53- and c-myc-positive cases. Overall survival was shorter in carcinomas with high MNA, SDNA, MNP and SDNP and low SHF. In multivariate analysis, performed by testing nuclear morphometric parameters, histological grade, tumor size, nodal status and p53 immunostaining in the Cox model, p53 over-expression and histological grade retained independent prognostic significance. When p53 was excluded, only SDNP appeared as an independent prognostic variable. Our results indicate that nuclear morphometric parameters can identify an aggressive tumor phenotype and provide additional prognostic information for patients with MBC. PMID- 11102894 TI - Altered expression of HLA class I antigens in breast cancer: association with prognosis. AB - Loss of surface expression of class I major histocompatibility antigens is commonly observed in malignant tumors and has been considered one of the mechanisms for escape from cytotoxic T cells. However, natural killer cells kill cells lacking HLA class I antigens. In the present study, we characterized by immunohistochemistry the HLA class I expression of breast carcinomas from 187 patients with TNM stages I and II, diagnosed 1981-1984, using beta(2) microglobulin as a marker and evaluated the effect on survival with a follow-up of up to 14 years. The largest group (48%) consisted of HLA class I-negative tumors (< or =10% of cells stained), mixed expression (>10% and <80% of cells stained) was seen in 36% and only 15% were classified as HLA class I-positive (> or=80% cells stained). No associations could be established with various clinicopathological parameters, such as tumor size, presence of lymph node metastases, histological grade, expression of hormone receptors, S phase and p53 mutations. There was no effect on recurrence-free survival in the whole group; but among node-negative patients (n = 86), those who had tumors with mixed HLA class I expression had a significantly higher probability of disease recurrence (OR = 3.42, p = 0.014) than patients with either HLA class I-positive or negative tumors, particularly after more than 5 years. In node-positive patients who received adjuvant therapy, this phenotype was not associated with disease recurrence. PMID- 11102895 TI - Down-regulation of insulin-like growth factor-I receptor and insulin receptor substrate-1 expression in advanced human breast cancer. AB - The ligands, receptors and related signaling proteins of the insulin-like growth factor family are involved in the regulation of breast-cancer cell growth. We investigated the expression pattern of insulin-like growth factor-I receptor (IGF IR), insulin receptor (IR) and insulin receptor substrate-1 (IRS-1), a core downstream signaling protein, in 69 primary breast-cancer specimens of different grades and in 21 control tissues by immunohistochemistry. In addition, cell proliferation (percentage of Ki67(+) nuclei) and estrogen receptor (ER) expression were determined. IGF-IR, IRS-1 and IR were expressed mainly in epithelial cells. IRS-1 and IGF-IR were expressed at high levels in control tissues and in well and moderately differentiated carcinomas but at low levels in poorly differentiated breast cancers. IR expression did not show a significant correlation with the differentiation grade of the tissues investigated. Statistical analysis (ROC analysis for tumor grade) demonstrated that down regulation of IGF-IR and IRS-1 correlated better with tumor progression than reduction of ER expression or increase in cell proliferation, IGF-IR showing the best correlation, followed by IRS-1 and, less significant, ER and Ki67. Our findings clearly show that progression of breast cancer is accompanied by a reduction of IGF-IR/IRS-1 expression and that IGF-IR/IRS-1 expression inversely correlates with high proliferation rate in dedifferentiated breast cancers. The strong correlation of IGF-IR and IRS-1 down-regulation with tumor progression suggests the use of IGF-IR and IRS-1 as a novel set of marker proteins for tumor grading. PMID- 11102896 TI - Papillary urothelial hyperplasia is a clonal precursor to papillary transitional cell bladder cancer. AB - Papilloma and papillary hyperplasia (PH) have been proposed to be the putative precursor lesions of papillary transitional-cell carcinoma of the urinary bladder. We examined 15 PH lesions and 4 papillomas for loss of heterozygosity (LOH) at 17 microsatellite markers on 9 chromosomal arms. Eight of 15 (53%) PHs were clonal, demonstrating LOH of at least 1 microsatellite marker. In contrast, none of the papillomas showed any genetic changes among the markers tested. In PH, chromosomal arm 9q was the most frequently lost (4/15), followed by 9p and 18q (n = 2) and, less frequently, 8p, 10q, 11p and 17p (n = 1). Furthermore, 2 hyperplastic lesions demonstrated LOH at 9q only, confirming the notion that allelic loss on chromosomal arm 9q is among the earliest events in bladder-cancer progression. In 1 patient, identical LOH patterns were observed between PH and a recurrent transitional-cell carcinoma. Our molecular data demonstrate that at least a proportion of PHs represent pre-cancerous lesions of the bladder that subsequently progress to papillary bladder cancer. Moreover, chromosomal arm 9q may harbor a tumor-suppressor gene(s) inactivated in the earliest stages of human bladder tumorigenesis. PMID- 11102897 TI - Predictive values of serum tumour markers tetranectin, OVX1, CASA and CA125 in patients with a pelvic mass. AB - Our objective was to compare the predictive value of the well-established tumour marker CA125 with the newer tumour markers tetranectin (TN), OVX1 and CASA in distinguishing benign and malignant pelvic masses in women. Participants included 185 women, 19 years or older, with a pelvic mass planned for surgical exploration. Significantly different CA125 levels were found between benign tumours and localised ovarian cancer (OC), advanced OC and other non-OCs. Significantly different TN levels were found between benign tumours and advanced OC (stage III/IV), between benign tumours and other cancers and between all OCs and other cancers. For CASA, significant differences were found between benign tumours and all OCs as well as advanced OC. No significant differences could be demonstrated for OVX1. Significant correlations for the 44 OC patients were found between CA125, TN and CASA. No significant correlations were found for OVX1, possibly because of the method used for collection and handling of serum samples. None of the new markers had any additional predictive value compared to CA125. TN and CASA levels correlated with FIGO stage and could be used to discriminate between benign and advanced OC. However, in comparison to the performance of CA125, the additional discriminative value of TN and CASA was minor. PMID- 11102898 TI - Circulating antibodies to p40(AIS) in the sera of respiratory tract cancer patients. AB - Studies of immune recognition in cancer have defined several tumor antigens using autologous cytotoxic T lymphocytes and by detection of serum antibodies to tumor associated products such as p53 and HER-2/neu. The AIS gene is a p53 homologue with multiple protein products (p40, p51, p63, p73L) on chromosomal arm 3q, frequently amplified and over-expressed in squamous-cell carcinoma of the respiratory tract. We analyzed the humoral response to p40(AIS) (a core domain of AIS products without the transactivation domain) by Western blot and ELISA using bacterially synthesized p40(AIS) protein. Antibodies were detected in the sera of 17/94 (18%) HNSCCs and 13/76 (17%) lung cancers, including 5/18 (26%) squamous cell carcinomas. Anti-p40(AIS) antibodies were not associated with factors such as sex, age, histopathological grading, extent or size of primary tumor, lymph node involvement and staging. Our results indicate that amplification and over expression of p40(AIS) may lead to antigen recognition by an autologous host with cancer. AIS may thus represent a new group of developmentally regulated genes that are recognized as tumor antigens. PMID- 11102899 TI - Screening for high-grade cervical intra-epithelial neoplasia and cancer by testing for high-risk HPV, routine cytology or colposcopy. AB - The validity of testing for high-risk types of human papillomavirus (HPV) in cervical cancer prevention programs is undetermined. We compared the performance on primary screening of HPV DNA testing, cytology and colposcopy in detecting cervical intra-epithelial neoplasia (CIN) grade 2 or 3 or cancer. A cohort of 4,761 women, median age 35 years, was screened by routine cytology, routine colposcopy and testing for high-risk HPV by a PCR-based method. Within an 8-month period, women with abnormal findings on cytology or screening colposcopy or in whom high-risk HPV types were detected were referred for colposcopy and biopsy. Women negative on all initial screening tests were followed by a second screening examination. To correct for work-up bias, the true prevalence of CIN 2 or 3 or cancer was estimated by projection from histologically verified subgroups. Cervical biopsies were taken in 364 women (7.6%), of whom 114 (2.4%) showed CIN 2 (n = 34) or CIN 3 (n = 71) or cancer (n = 9). High-risk HPV testing achieved bias corrected performance measures of 89.4% sensitivity, 93.9% specificity, 35.8% positive predictive value and 99.6% negative predictive value. Bias-corrected rates of true- and false-positives by high-risk HPV testing compared to cytology (colposcopy) were about 4.5 (6.7) and 19.1 (7.4) times higher, respectively. The quality of routine cytology was controlled by computer-assisted review, and the observed number of true-positives more than doubled after adding automated review results. In middle-aged women, testing for high-risk HPV types, particularly when negative, may be used to increase the screening interval in programs for secondary prevention of cervical cancer. PMID- 11102900 TI - PLK (polo-like kinase), a new prognostic marker for oropharyngeal carcinomas. PMID- 11102901 TI - Critical illness myopathy. PMID- 11102902 TI - Clinical and genetic heterogeneity in myotonic dystrophies. AB - This review of myotonic dystrophies primarily concentrates on the clinical and genetic findings that can distinguish a novel form of myotonic dystrophy, myotonic dystrophy type 2 (DM2); proximal myotonic myopathy (PROMM); and proximal myotonic dystrophy (PDM) from myotonic dystrophy type 1 (DM1). The multisystemic nature of these disorders leads to a spectrum of symptoms and signs. Careful clinical evaluation of patients with DM2/PROMM shows that the similarities among the multisystemic myotonic disorders outweigh the differences. An important point in the comparison of the phenotypes of DM1 and DM2/PROMM is that no severe congenital type of DM2/PROMM has yet been described. Genetic linkage analyses show that myotonic dystrophies can be divided into three types: the conventional Steinert type linked to chromosome 19q13.3 (DM1); DM2/PROMM and PDM linked to chromosome 3q21.3; and families not linked to either chromosomal site. Although the diagnosis may be clinically suspected, it depends on DNA analysis. PMID- 11102903 TI - Paraneoplastic syndromes of the spinal cord, nerve, and muscle. AB - Cancer can affect the peripheral nervous system by non-metastatic, sometimes immune-mediated mechanisms. Recognition of these paraneoplastic syndromes is important because it can lead to the detection of the tumor, and also helps to avoid unnecessary studies to determine the cause of the neurologic symptoms in patients with cancer. Many paraneoplastic syndromes of the peripheral nervous system are not associated with serum antibodies that serve as markers of paraneoplasia. For this group of disorders the diagnosis depends on the clinician's index of suspicion and conventional electrophysiologic and laboratory tests. Treatment of the tumor, immunotherapy, or both may improve some of these syndromes. This review focuses on paraneoplastic syndromes of the spinal cord, peripheral nerve, and muscle. PMID- 11102905 TI - Mechanomyographic amplitude and mean power output during maximal, concentric, isokinetic muscle actions. AB - The purpose of this study was to determine the velocity-related patterns for mechanomyographic (MMG) amplitude, electromyographic (EMG) amplitude, mean power output (MP), and peak torque (PT) of the superficial muscles of the quadriceps femoris (vastus lateralis [VL], rectus femoris [RF], and vastus medialis [VM]) during maximal, concentric, isokinetic leg extensions. Twelve adult women (mean +/- SD: 22 +/- 3 years of age) performed such leg extensions at velocities of 60 degrees, 120 degrees, 180 degrees, 240 degrees, and 300 degrees /s on a Cybex 6000 dynamometer. PT decreased (P < 0.05) across velocity to 240 degrees /s. MP and MMG amplitude for each muscle (VL, RF, and VM) increased (P < 0.05) with velocity to 240 degrees /s and then plateaued. EMG amplitude increased (P < 0.05) to 240°/s for the VL, remained unchanged across velocity (P > 0.05) for the RF, and increased (P < 0.05) to 300 degrees /s for the VM. The results indicated close similarities between the velocity-related patterns for MMG amplitude and MP, but dissociations among EMG amplitude, MMG amplitude, and PT. These findings support the recent hypothesis that MMG amplitude is more closely related to MP than PT during maximal, concentric, isokinetic muscle actions and, therefore, may be useful for monitoring training-induced changes in muscle power. PMID- 11102904 TI - Recovery from 6 weeks of repeated strain injury to rat soleus muscles. AB - Recovery from chronic strain injury (50 strains daily, five times weekly for 6 weeks to hyperactive soleus muscles) was followed for 3 months in female rats after cessation of chronic hyperactivity induced by pretreatment of the plantar flexor muscles with tetanus toxin. After 6 weeks of repeated strains, muscle mass decreased by 62%, myofiber areas were reduced by 87%, and noncontractile tissue expanded dramatically by 222%. Collagen content increased by almost ninefold (control 40 +/- 3 microg/mg, chronic injury 392 +/- 53 microg/mg), whereas the molar ratio of collagen (pyridinoline) crosslinks to collagen remained the same (control 0.20 +/- 0.01, chronic injury 0.16 +/- 0.01). After 3 months of ambulation, muscle mass returned to normal but myofiber areas remained smaller by 21%, noncontractile tissue was still markedly elevated by 18% with increased collagen content (107 +/- 15 microg/mg), and the molar ratio of crosslinks to collagen increased by 75% during recovery. Thus, rat soleus muscles recovered very slowly and incompletely from chronic strain injuries that produced muscle fibrosis, highlighting the necessity of devising preventative strategies for repeated strain injuries. PMID- 11102906 TI - Neutralizing capacity of commercial bothropic antivenom against Bothrops jararacussu venom and bothropstoxin-I. AB - Bothrops jararacussu venom and its major toxin, bothropstoxin-I (BthTX-I), possess myotoxic and neurotoxic activities. The ability of commercial equine antivenom to neutralize these activities was studied in mouse isolated phrenic nerve-diaphragm (PND) and extensor digitorum longus (EDL) preparations by indirect stimulation (0.1 HZ, 0.2 ms). The time required to produce 50% neuromuscular blockade in the PND and EDL preparations was, respectively, 70 +/- 11.5 min and 58 +/- 8 min for B. jararacussu venom (50 microg/mL), and 31 +/- 6 min and 30 +/- 3 min for BthTX-I (20 microg/mL). After a 120-min incubation, the creatine kinase (CK) concentrations in the EDL preparations were 3464 +/- 346 U/L and 3422 +/- 135 U/L following exposure to venom (50 microg/mL) and BthTX-I (20 microg/mL), respectively. Antivenom neutralized the neuromuscular blockade induced by the venom and toxin in PND preparations in a dose-dependent fashion, but only partially neutralized this effect in EDL. Antivenom also effectively prevented the venom- and toxin-induced release of CK from EDL. In contrast, histological analysis showed that the morphological damage caused by B. jararacussu venom and BthTX-I in the EDL was only partially prevented by the anti venom. These results indicate that commercial equine antiserum fully protects against the neurotoxic action of B. jararacussu and BthTX-I in PND preparations, but only partially protects against the neurotoxic and myotoxic actions of the venom and its toxin in EDL preparations. Care must therefore be exercized in extrapolating results from different preparations even when similar pharmacological or physiological responses are involved. PMID- 11102907 TI - Changes in corticomotor excitability after fatiguing muscle contractions. AB - To investigate whether the type and duration of activity influences corticomotor excitability following fatiguing exercise, we compared motor evoked potential (MEP) responses of the biceps brachii to transcranial magnetic stimulation (TMS) during recovery from two different exercise regimens. Responses were recorded in both the resting state and during a weak contraction. Ten subjects performed a 60 s maximal voluntary contraction (MVC) and, on a subsequent occasion, a sustained 20% MVC to the point of exhaustion. Resting MEP amplitude declined following maximal and submaximal protocols, reaching 34% and 31% of pre-exercise means, respectively (P < 0.001 for both). In contrast, mean facilitated MEP amplitude showed a smaller and more transient decrement following the sustained submaximal effort (64%; P < 0.05), but not the 60-s MVC. Abolition of the postexercise depression in resting MEP amplitude by a weak tonic contraction indicates that decreases in excitability at the spinal level contribute to the reduced corticomotor excitability observed after fatiguing exercise. PMID- 11102908 TI - Prior heat stress improves survival of ischemic-reperfused skeletal muscle in vivo. AB - The ability of heat stress to improve the survival of ischemic-reperfused skeletal muscle in vivo was investigated. Ischemia-reperfusion was applied using the rat hindlimb tourniquet model. The viability of ischemic-reperfused muscle (11 +/- 1%) was increased by prior mild heat stress (86 +/- 2%). To investigate whether heat shock protein 70 (Hsp 70) expression in the muscle of the heated limb was responsible for this protection, the survival of Hsp 70-expressing transduced myoblasts and myocytes was measured after exposure to mediators of ischemia-reperfusion injury. Survival was improved in Hsp 70-positive myoblasts but not in myocytes, suggesting that the mechanism of protection conferred by heat stress in vivo cannot be explained by the expression of Hsp 70 in myocytes and may involve a more complex mechanism. In conclusion, prior heat stress is effective in protecting mature skeletal muscle in vivo against necrosis after ischemia-reperfusion and has potential for use in microsurgical procedures requiring tourniquet applications. PMID- 11102909 TI - Electromyographic evidence of subclinical myopathy in hypertrophic cardiomyopathy. AB - Hypertrophic cardiomyopathy (HCM) is due to a number of mutations of contractile protein genes such as beta-cardiac myosin, myosin binding protein-C, and troponin T. Unlike troponin-T, beta-myosin is a constituent of slow skeletal muscle and its mutations generally have a better prognosis. In order to investigate the usefulness of electromyography in detecting skeletal muscle involvement in HCM, 46 patients were examined using both conventional electromyography (EMG) and quantitative electromyography (QEMG) methods. The QEMG involved motor unit potential (MUP) analysis, turns/amplitude (TAA) analysis, and power spectrum analysis of the interference pattern. Using conventional EMG, myopathic findings were demonstrated in 13 patients (28%). Receiver operating characteristic (ROC) analysis of the results of a discriminant function extracted using QEMG values, identified correctly 10 out of 11 normal controls and all 9 myopathic control patients, and displayed a 15% presence of myopathy (7 patients) among the cardiomyopathy group. The duration of MUPs was the most sensitive among the quantitative parameters in differentiating normal from myopathic subjects. Since skeletal muscle involvement may be due to distinct gene mutations, normal and myopathic EMG findings may reflect HCM subpopulations with a different genetic substrate. PMID- 11102910 TI - Bcl-2 and bax immunohistochemistry in denervation-reinnervation and necrosis regeneration of rat skeletal muscles. AB - Bcl-2 and Bax immunohistochemistry was examined in the skeletal muscle of rats after cutting the sciatic nerve, as a model of denervation and reinnervation, and in the anterior tibialis muscle of rats after an intramuscular injection of metoclopramide, as a model of necrosis and regeneration of muscle fibers, to better understand the role of these proteins in muscle disorders. An increase in Bax immunoreactivity was seen in long-standing denervated and reinnervated muscle fibers. Bcl-2 and Bax immunoreactivity was limited to macrophages in necrotic muscle fibers at 24 h after the intramuscular injection of metoclopramide. However, increased Bax immunoreactivity was observed in regenerating muscle fibers by the fourth day after the injection. Muscle fiber nuclei with the morphological features of apoptosis were not observed in rat muscles after the intramuscular injection of metoclopramide or after the severing of the sciatic nerve. Furthermore, using the Tunel method, no stained nuclei were observed in the two groups of animals. Our observations in the experimental models of skeletal muscle denervation-reinnervation and necrosis-regeneration here described suggest that modification in the intensity of the Bax protein is not related to the process of cell death but rather that increased Bax expression is associated with muscle fiber regeneration. PMID- 11102911 TI - Prognostic significance of preoperative electrophysiologic investigation for facial nerve outcome in acoustic neuroma surgery. AB - The prognostic significance of transcranial magnetic stimulation and nasal muscle F-wave recording to predict postoperative facial nerve function was assessed in 24 patients with unilateral acoustic neuromas (mean diameter, 31 mm) and clinically intact facial nerve function. Latency of F waves and response to cortical magnetic stimulation, as well as F ratios, central motor conduction time, and the ratio of response latency to cortical and cisternal magnetic stimulation were significantly increased. Outcome analysis revealed no significant correlation between preoperative electrophysiologic changes and postoperative facial nerve function. However, a significant correlation with tumor diameter was detected. Thus, acoustic neuroma size seems to be the best predictor of facial nerve function after surgery. PMID- 11102912 TI - Complex fasciculations and their origin in amyotrophic lateral sclerosis and Kennedy's disease. AB - Complex fasciculations are common in patients with amyotrophic lateral sclerosis (ALS). Combined fasciculations, defined as a complex fasciculation consisting of two or more components that occur independently but also in combination with another component, also occur in ALS. To test the hypothesis that combined fasciculations originate at the supraspinal level, we analyzed 2681 fasciculation potentials from 17 patients with definite or probable ALS by the El Escorial criteria. Results were compared with 304 fasciculation potentials recorded from 6 patients with Kennedy's disease, in which the upper motoneurons are spared. The mean firing frequency of the fasciculations was 24.4 +/- 25.6 per minute in ALS, significantly higher than the 2.9 +/- 3.4 per minute found in Kennedy's disease (P < 0.0001). In ALS, the mean combination ratio (the number of times that a combined fasciculation occurred divided by the total number of fasciculations) was 4.6 +/- 8.3% (range 0-33). Fourteen of 17 ALS patients had combined fasciculations, but only one combined fasciculation was found in a patient with Kennedy's disease. Combined fasciculations are distinctive in ALS, and we hypothesize that they are triggered by a supraspinal mechanism reflecting dysfunction of descending motor pathways. PMID- 11102913 TI - An autosomal dominant early adult-onset distal muscular dystrophy. AB - In this study we describe an autosomal dominant distal muscular dystrophy in a small Austrian family. The myopathy started in early adulthood with a slowly progressive weakness of the muscles of the anterior tibial compartment, followed by the long finger extensors and sternocleidomastoids in some family members. Other muscles were spared. Histopathology showed fiber size variation and autophagic vacuoles. This disease pattern is similar to Laing distal myopathy, which has been described previously in only one other family. PMID- 11102915 TI - Tibial nerve entrapment in the popliteal fossa. AB - Details are presented of nine cases of tibial nerve entrapment by the tendinous arch of origin of the soleus muscle. The diagnosis was confirmed by surgical exploration of the popliteal fossa in six patients, who recovered fully after division of the soleus arch, whereas the other three improved spontaneously. This condition can be distinguished clinically from tibial nerve compression at the ankle, and from S1 radiculopathy, by the presence of severe pain and tenderness and a positive Tinel sign in the popliteal fossa, and by electrodiagnostic studies. PMID- 11102914 TI - Optimizing the number of tests for carpal tunnel syndrome. AB - The combined sensory index (CSI), the sum of three latency differences, median ulnar across the palm (palmdiff), median-ulnar to the ring finger (ringdiff), and median-radial to the thumb (thumbdiff), has higher sensitivity and reliability for carpal tunnel syndrome than individual tests. The objective in this study was to develop an approach that minimizes testing but maximizes accuracy. We retrospectively studied 300 hands. There were endpoints for individual tests that confidently predicted the CSI; for ranges between these endpoints, further testing was required. These ranges were: palmdiff 0-0.3 ms; ringdiff 0.1-0.4 ms; and thumbdiff 0.2-0.7 ms. One may use a strategy in which more tests are performed when results are in these ranges. This approach can allow accurate diagnosis with fewer tests when values are extreme, yet uses the greater diagnostic power of more tests when values are midrange. PMID- 11102916 TI - Multifocal motor neuropathy presenting with respiratory failure. AB - Multifocal motor neuropathy is a disorder characterized by slowly progressive asymmetrical limb weakness and multiple motor conduction blocks. We report a 56 year-old woman with this disorder who presented unusually with respiratory failure and who initially had absent responses to phrenic nerve stimulation bilaterally. The mechanism of the patient's respiratory failure may have been chronic conduction blocks in the phrenic nerves leading to diaphragmatic weakness. PMID- 11102917 TI - Hypoglycemic sensorimotor polyneuropathy associated with insulinoma. AB - Hypoglycemia-induced peripheral neuropathy due to insulinoma is unusual and, as far as we know, has previously been reported in only 34 patients. In this case report, we describe the clinical features, electrophysiological features, and pathological findings in a 37-year-old patient with polyneuropathy from repeated hypoglycemic episodes over a 9-year period that related to an insulinoma. The literature is discussed. The reported case is of special interest because the peripheral neuropathy led to the correct diagnosis. PMID- 11102918 TI - Is the sural nerve a pure sensory nerve? PMID- 11102920 TI - AAEM news and comments PMID- 11102921 TI - Gerhard koch 1913-1999 PMID- 11102922 TI - Oto-palato-digital syndrome, type II: report of three cases with further delineation of the chondro-osseous morphology. AB - Oto-palato-digital syndrome type II (OPD II) is a lethal X-linked skeletal dysplasia with pleiotropic manifestations. The basic defect is not known. There has been only one detailed report of the chondro-osseous abnormalities in this condition describing abnormal periosteal ossification in a single case [1990: Am J Med Genet 36:226-231]. We report on three cases of OPD II emphasizing the chondro-osseous morphology. Although endochondral ossification was normal, periosteal ossification was defective with islands of cortical bone aplasia and hyperplasia of the periosteum. The trabecular bone was also extremely poorly formed and markedly hypercellular. Both membranous ossification and bone remodeling appear to be defective in OPD II and should account for part of the observed phenotype. The biglycan gene maps to Xq28 and is involved in bone formation, but was excluded as a candidate by direct sequencing of cDNA in one case. PMID- 11102923 TI - Char syndrome: an additional family with polythelia, a new finding. AB - This report describes a father and daughter with Char syndrome, a rare autosomal dominant disorder. Both affected individuals had typical face, strabismus, and foot anomalies. The girl also had a patent ductus arteriosus. In addition, both patients had polythelia (supernumerary nipples), a finding not described before in the Char syndrome. PMID- 11102924 TI - Variable presentation of Rothmund-Thomson syndrome. AB - The recent finding that a subset of patients with Rothmund-Thomson syndrome (RTS) have mutations of a helicase gene has prompted reexamination of the phenotypes of individuals diagnosed with this disorder. We report on two patients with variable presentations of RTS. Initial presenting symptoms included growth deficiency and absent thumbs in one patient and osteogenic sarcoma and poikiloderma in the second patient. The growth-deficient patient was diagnosed with growth hormone deficiency and had a subnormal response to growth hormone supplementation. Neither malformations nor growth deficiency were present in the patient with osteogenic sarcoma, and her only other manifestation of RTS was poikiloderma. The diagnosis of RTS should be considered in all patients with osteogenic sarcoma, particularly if associated with skin changes. PMID- 11102925 TI - Phenotypic overlap of McKusick-Kaufman syndrome with bardet-biedl syndrome: a literature review. AB - Hydrometrocolpos (HMC) and post-axial polydactyly (PAP) are common to both McKusick-Kaufman syndrome (MKS) and Bardet-Biedl syndrome (BBS). We review reported cases of MKS and BBS presenting with HMC and PAP early in life to determine if there are clinical features that allow discrimination between the two syndromes as the primary features of retinitis pigmentosa, obesity, learning disability in BBS are age-dependent. We did not find any phenotypic features that allowed reliable differentiation between the two syndromes in the neonatal period. However, uterine, ovarian, and fallopian tube anomalies are more common in BBS patients, and it may be that these clinical features prove to be useful discriminating features. We conclude that sporadic female infants with HMC and PAP cannot be diagnosed with MKS until at least age 5 years and that monitoring for the complications of BBS should be performed in these patients. PMID- 11102926 TI - Folate pathway gene alterations in patients with neural tube defects. AB - Periconceptional folate supplementation reduces the recurrence and occurrence risk of neural tube defects (NTD) by as much as 70%, yet the protective mechanism remains unknown. Inborn errors of folate and homocysteine metabolism may be involved in the aetiology of NTDs. Previous studies have demonstrated that both homozygosity for the C677T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene, and combined heterozygosity for the C677T and for another mutation in the same gene, the A1298C polymorphism, represent genetic risk factors for NTDs. In an attempt to identify additional folate related genes that contribute to NTD pathogenesis, we performed molecular genetic analysis of folate receptors (FRs). We identified 4 unrelated patients out of 50 with de novo insertions of pseudogene (PS)-specific mutations in exon 7 and 3'UTR of the FRalpha gene, arising by microconversion events. All of the substitutions affect the carboxy terminal amino acid membrane tail, or the GPI anchor region of the nascent protein. Furthermore, among 150 control individuals, we also identified one infant with a gene conversion event within the FRalpha coding region. This study, though preliminary, provides the first genetic association between molecular variations of the FRalpha gene and NTDs and suggests that this gene can act as a risk factor for human NTD. PMID- 11102927 TI - Plasma levels of amyloid beta 40 and 42 are independent from ApoE genotype and mental retardation in Down syndrome. AB - In Down syndrome (DS) brain an early, selective accumulation of amyloid beta (Abeta) peptides ending at residue 42 (Abeta42) occurs. Whether this event depends on an altered processing of amyloid beta precursor protein (APP) or on defective clearance is uncertain. To investigate this issue, we measured Abeta species 40 and 42 in plasma from 61 patients with DS, 77 age-matched normal controls, and 55 mentally retarded subjects without chromosomal abnormalities. The Abeta 40 and 42 plasma levels were then correlated with apolipoprotein E (apoE) genotypes in all groups of cases, and with I. Q. and Mini Mental Status Examination values in DS subjects. Both Abeta species were significantly elevated in DS compared to control groups, and the extent of their increase reflects that expected from APP gene overexpression. Plasma levels of Abeta 40 and 42 did not correlate with apoE genotypes in DS and control cases, and with the extent of mental retardation in DS subjects. The results indicate that accumulation and clearance of plasma and cerebral Abeta are regulated by different and independent factors. PMID- 11102928 TI - Phenotypic features in a boy with monosomy 18 mosaicism. AB - We report on a patient with mosaicism for monosomy 18, a chromosomal abnormality that has been reported only once previously. The patient had cleft lip and palate and mild behavioral and academic problems. His phenotype was milder in comparison with the previously reported patient by Khalifa et al. [1996: Clin Genet 49:318 320]. Further case reports of this chromosomal anomaly will be needed to determine a consistent phenotypic pattern to assist in management and genetic counseling. PMID- 11102929 TI - Clinical variability and genetic homogeneity of the camptodactyly-arthropathy coxa vara-pericarditis syndrome. AB - The camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP) is an autosomal recessive condition characterized by the association of congenital or early onset camptodactyly and noninflammatory arthropathy with synovial hyperplasia. Progressive coxa vara deformity and/or noninflammatory pericardial or pleural effusions have been observed in some patients. Recently, the disease gene has been assigned to human chromosome region 1q25-q31, and truncating mutations have been identified in the megakaryocyte stimulating factor gene. Studying 12 patients from 8 unrelated families, we emphasized hip and spine involvement, particularly in the course of the disease as shown in a 58-year-old patient. Despite clinical variability, linkage studies support genetic homogeneity of the disease. PMID- 11102930 TI - Familial choanal atresia with maxillary hypoplasia, prognathism, and hypodontia. AB - We report on two sibs and a cousin with bilateral choanal atresia. At 2 months, one sib died of complications following surgical correction of her defects. We evaluated her brother and cousin at age 7 and 9 years, respectively. Both had a tall forehead, maxillary hypoplasia, prognathism, and absence of certain deciduous and permanent teeth. Psychomotor development was appropriate for age. Roentgenocephalometric analyses of several relatives showed that one grandfather of these children and two of the five uncles and aunts also had maxillary hypoplasia and/or prognathism. To our knowledge, this condition has not been described previously and may represent a newly recognized autosomal dominant condition with incomplete penetrance and variable expressivity caused by a defect of neural crest development. PMID- 11102931 TI - Syndrome of gingival hypertrophy, hirsutism, mental retardation and brachymetacarpia in two sisters: specific entity or variant of a described condition? AB - Two sisters born to consanguineous Lebanese parents had mental retardation and epilepsy, brachymetacarpalia, hirsutism, bulbous soft nose, thick floppy ears with abnormal configuration and gingival hypertrophy. One girl presented additionally with tetralogy of Fallot and the other with congenital hypothyroidism and bilateral ureteral stenosis. These manifestations resemble the syndrome of hypertrichosis-gingival fibromatosis-mental retardation and seizures of Anavi et al. [1989: Dev Med Child Neurol 31:538-542] but our two girls additionally have brachymetacarpia. The inheritance seems to be autosomal recessive. These two sisters may represent a hitherto undescribed syndrome. We discuss the findings in our patients in relation to the literature. PMID- 11102932 TI - Uruguay facio-cardio-musculo-skeletal syndrome: a novel X-linked recessive disorder. AB - We report on three male patients from a single family with a brachyturricephaly, "pugilistic" facial appearance, a muffled voice, cardiomyopathy, muscular hypertrophy, broad hands, wide feet with progressive pes cavus deformities, dislocation of toes, variable congenital hip dislocation, and scoliosis. Three other males in the family, now deceased from cardiac disease, appear to have had the same disorder. The mother of the propositus has milder signs of the syndrome. All affected males are related through the maternal line. These cases represent an apparently previously undescribed X-linked recessive syndrome. PMID- 11102933 TI - Clinical heterogeneity in mitochondrial DNA deletion disorders: a diagnostic challenge of Pearson syndrome. AB - The clinical presentation of mitochondrial DNA (mtDNA) disorders is quite diverse. Very often, the initial symptoms do not fit a specific disease, and diagnosis is difficult to make. We describe a patient who presented with macrocytic anemia. Extensive biochemical and clinical work-up failed to provide an etiology for the macrocytic anemia. The patient over the course of 6 years developed gait problems, exercise intolerance, episodic vomiting, short stature, dermatological problems, and recurrent infection. At age 8 years she had encephalopathy with ataxia and dysphagia. The presence of elevated lactate, bilateral basal ganglia calcification, and ragged red fibers led to mtDNA mutational analysis. A novel 4.4-kb deletion from nucleotide position 10,560 to nucleotide position 14, 980 was identified in muscle biopsy. The same heteroplasmic mtDNA deletion was present in blood, buccal cells, and hair follicles, but not in mother's blood, consistent with sporadic mutation in the patient. This case emphasizes the importance of considering mtDNA disorder in patients with multisystemic symptoms that cannot be explained by a specific diagnosis. PMID- 11102934 TI - Transmission of the dysgnathia complex from mother to daughter. AB - We report the first observation of parent-to-child transmission of dysgnathia, a rare disorder characterized by severe mandibular hypoplasia or agenesis, ear anomalies, microstomia, and microglossia. Patient 1 was noted prenatally by ultrasound to have severe micrognathia and, after birth, abnormal ears with canal stenosis and non-contiguous lobules located dorsally to the rest of the pinnae, normal zygomata, severe jaw immobility and microstomia with an opening of only 4 to 5 mm, hypoplastic tongue, and cleft palate. The 21-year-old mother of patient 1 was born with severe micrognathia requiring tracheostomy, microglossia, cleft palate with filiform alveolar bands, abnormal pinnae, and decreased conductive hearing. Dysgnathia is thought to result from a defect in the development of the first branchial arch. A similar phenotype has been seen in Otx2 haplo insufficiency and endothelin-1 homozygous null mice, suggesting that these genes contribute to branchial arch development. Our report of a long-surviving mother and her daughter with non-syndromal dysgnathia may lead to identification of the molecular basis of these findings and provide insight into the genetics of first branchial arch formation. The survival of patient 1 and patient 2 beyond the neonatal period has implications for improvements in prenatal diagnosis and counseling and for neonatal treatment of this condition. PMID- 11102935 TI - Unbalanced 4;6 translocation and progressive renal disease. AB - Two sibs are described with an unbalanced 4;6 translocation resulting in partial trisomy 6p and monosomy for distal 4p. Growth retardation, psychomotor retardation, and characteristic facial appearance are present. The facial anomalies include high prominent forehead, blepharoptosis, blepharophimosis, high nasal bridge, bulbous nose, long philtrum, small mouth with thin lips, and low set ears. Both children have small kidneys and have had proteinuria since early childhood. The older boy developed progressive renal disease including hypertension and renal failure necessitating renal transplantation at age 18 years. Renal biopsy of the younger girl also indicates significant renal involvement. Progressive renal disease is likely an important part of the trisomy 6p phenotype. PMID- 11102936 TI - Blaschkolinear skin pigmentary variation due to trisomy 7 mosaicism. AB - Mosaic trisomy 7 is a rare condition that can be seen in individuals with Blaschkolinear skin pigmentary variation, somatic asymmetry, and variable other clinical anomalies. In any patient presenting with Blaschkolinear skin pigmentary variation, varying degrees of asymmetrical growth disturbance, developmental delay, and a normal lymphocytic karyotype, chromosomal mosaicism may be present. To rule out tissue-specific or occult chromosomal mosaicism, it is recommended to culture and karyotype skin fibroblasts, since lymphocyte cell lines may not demonstrate the abnormal cell line. Early diagnosis is of paramount importance, since early physical, occupational, and speech/language therapy can greatly improve the developmental outcome of these patients. We report on a fourth patient with trisomy 7 mosaicism in whom early diagnosis and developmental therapy contributed to an improved developmental outcome when compared with patients in previous reports. Early intervention can greatly benefit patients with this diagnosis, especially in minimizing the aggressive behavior associated with communication difficulties. Our patient has milder manifestations than the previously reported patients with no seizure activity or asymmetry and fewer cells with trisomy 7. PMID- 11102937 TI - Nonsyndromic total anomalous venous return associated with a de novo translocation inolving chromosomes 10 and 21 t(10;21)(q23.1;q11.2). PMID- 11102938 TI - Case of congenital hypotransferrinemia suggests that tissue hypoxia during fetal development may cause hypospadias. PMID- 11102939 TI - Response to letter to the editor by goldwurm and biondi-"Case of congenital hypotransferrinemia suggests that tissue hypoxia during fetal development may cause hypospadias" PMID- 11102940 TI - Response to the letter to the editor by fung PMID- 11102941 TI - Malpuech syndrome: a possible relationship with the Wolf-Hirschhorn/Pitt-Roger Danks phenotype. PMID- 11102942 TI - Lack of sex-ratio distortion in neurofibromatosis 2. PMID- 11102943 TI - Premature adrenal cortical dysfunction in mandibuloacral dysplasia: a progeroid like syndrome. PMID- 11102944 TI - Partial deletion of chromosome 12q is not usually associated with CFC syndrome. PMID- 11102945 TI - New model for the avian magnetic compass. AB - It is proposed that the avian magnetic compass depends on the angle between the horizontal component B(h) of the geomagnetic field (GMF) and E(r), the radial electric field distribution generated by gamma-oscillations within the optic tectum (TeO). We hypothesize that the orientation of the brain relative to B(h) is perceived as a set of electric field ion cyclotron resonance (ICR) frequencies that are distributed in spatially recognizeable regions within the TeO. For typical GMF intensities, the expected ICR frequencies fall within the 20-50 Hz range of gamma-oscillation frequencies observed during visual stimulation. The model builds on the fact that the superficial lamina of the TeO receive signals from the retina that spatially map the visual field. The ICR frequencies are recruited from the local wide-band gamma-oscillations and are superposed on the tectum for interpretation along with other sensory data. As a first approximation, our analysis is restricted to the medial horizontal plane of the TeO. For the bird to fly in a preferred, previously mapped direction relative to B(h), it hunts for that orientation that positions the frequency maxima at appropriate locations on the TeO. This condition can be maintained even as B(h) varies with geomagnetic latitude during the course of long-distance flights. The magnetovisual coordinate system (straight phi, omega) overlaying the two halves of the tectal surface in a nonsymmetric way may imply an additional orienting function for the TeO over and above that of a simple compass (e.g., homing navigation as distinct from migrational navigation). PMID- 11102946 TI - Prenatal exposure to 900 MHz, cell-phone electromagnetic fields had no effect on operant-behavior performances of adult rats. AB - To clarify potential health risks of radio-frequency electromagnetic fields (EMFs) used in cellular telephone technology to the developing brain, Wistar rats were continuously exposed during pregnancy to a low-level (0.1 mW/cm(2)) 900 MHz, 217 Hz pulse modulated EMF that approximated the highest legal exposure of normal populations to the radiation of base antennas of the GSM digital cell-phone technology. Whole body average specific absorption rate (SAR) values for the freely roaming, pregnant animals were measured in models; they ranged between 17.5 and 75 mW/kg. The offspring of exposed and of sham-exposed dams were coded and tested later as adults in a battery of ten simultaneously operated test chambers (Skinner boxes) during night time. Eight groups of ten coded animals in each group were tested for learning deficits in a sequence of nine, computer controlled, 15 h sessions of the food-reinforced contingency Differential Reinforcement of Rate with increasing performance requirements. Two different sets of events were recorded: The food-reinforced lever-pressing activity of the animals and the inter-response intervals (IRIs) between consecutive lever presses. IRI-occurence patterns discriminated consistently between "learners" and "non-learners". Analyses of performance scores and of IRI-patterns both showed that exposure in-utero to the GSM field did not induce any measurable cognitive deficits. PMID- 11102947 TI - In vivo enhancement of chemotherapy with static electric or magnetic fields. AB - We evaluated the ability of both static electric and static magnetic fields to enhance the in vivo action of a chemotherapeutic agent, adriamycin, against transplanted mammary tumors in mice. Female B6C3F1 mice with transplanted mammary adenocarcinoma were divided into four randomized groups and injected with 10 mg/kg adriamycin on day 7 of the study. Three of the groups were then exposed to nonuniform static electric or static magnetic fields. The resulting tumor regression in each group was measured four times during the remaining 13 days of the 20 day study. Two-sided statistical tests revealed all of the static field exposed groups achieved significantly greater (P G (L244V), 907C>G (R303G), and 370C>T (P124S), in the coding region of one FUT6 allele. Another individual, expressing weak plasma alpha3-fucosyltransferase activity, had the 907C>G together with the 370C>T mutation, but did not have the 730C>G mutation. PCR-RFLP analyses of complete families confirmed the segregation of these alleles and illustrated the existence and inheritance of the [370C>T; 907C>G] mutated allele in three additional families. Altogether, this allele was found heterozygously in nine Indonesian and two Swedish individuals, all with detectable plasma alpha3-fucosyltransferase activities. The FUT6 allele with the three mutations (370C>T; 730C>G; 907C>G) was identified heterozygously in only two Indonesian individuals, both having the inactivating 739G>A mutation in the other allele and both lacking plasma alpha3-fucosyltransferase activity. Enzyme studies made on transiently transfected COS-7 cells demonstrated that the combination of the 370C>T, 730C>G and 907C>G mutations decreased the V(max) by more than 80%, but caused no obvious change of the apparent K(m) values for GDP fucose and Gal-N-acetyllactosamine. In comparison, chimeric constructs with the isolated 730C>G or 907C>G mutations decreased the V(max) values by about two thirds and one third, respectively. PMID- 11102977 TI - High frequency of recurrent mutations in BRCA1 and BRCA2 genes in Polish families with breast and ovarian cancer. AB - Germ-line mutations in BRCA1 and BRCA2 genes result in a significantly increased risk of breast and ovarian cancer. Other genes involved in an increased predisposition to breast cancer include the TP53 gene, mutated in Li-Fraumeni syndrome. To estimate the frequency of germ-line mutations in these three genes in Upper Silesia, we have analyzed 47 breast/ovarian cancer families from that region. We found five different disease predisposing mutations in 17 (36%) families. Twelve families (25.5%) carried known BRCA1 mutations (5382insC and C61G), four families (8.5%) carried novel BRCA2 mutations (9631delC and 6886delGAAAA), and one family (2%) harbored novel mutation 1095del8 in the TP53 gene, which is the largest germline deletion in coding sequence of this gene identified thus far. The 5382insC mutation in BRCA1 was found in 11 families and the 9631delC mutation in BRCA2 occurred in three families. These two mutations taken together contribute to 82% of all mutations found in this study, and 30% of the families investigated harbor one of these mutations. The very high frequency of common mutations observed in these families can only be compared to that reported for Ashkenazi Jewish, Icelandic, and Russian high-risk families. This frequency, however, may not be representative for the entire Polish population. The observed distribution of mutations will favor routine pre-screening of predisposed families using a simple and cost-effective test. PMID- 11102978 TI - Mutational analyses of BRCA1 and BRCA2 in Ashkenazi and non-Ashkenazi Jewish women with familial breast and ovarian cancer. AB - In Ashkenazi (East European) Jews, three predominant mutations in BRCA1 (185delAG and 5382insC) and BRCA2 (6174delT) account for the majority of germline mutations in high-risk breast and/or ovarian cancer families. Among non-Ashkenazi Jews, the 185delAG, Tyr978Ter, and a handful of "private" mutations have been reported anecdotally within both genes. In this study we attempted to determine the spectrum of BRCA1 and BRCA2 mutations in high-risk Jewish individuals, non carriers of any of the predominant Jewish mutations. We employed multiplex PCR and denaturing gradient gel electrophoresis (DGGE) analysis for BRCA2, and combined denaturing high performance liquid chromatography (DHPLC) and protein truncation test (PTT) for BRCA1, complemented by DNA sequencing. We screened 47 high-risk Jewish individuals, 26 Ashkenazis, and 21 non-Ashkenazis. Overall, 13 sequence alterations in BRCA1 and eight in BRCA2 were detected: nine neutral polymorphisms and 12 missense mutations, including five novel ones. The novel missense mutations did not co-segregate with disease in BRCA1 and were detected at rates of 6.25% to 52.5% in the general population for BRCA2. Our findings suggest that except for the predominant mutations in BRCA1 and BRCA2 in Jewish individuals, there are only a handful of pathogenic mutations within these genes. It may imply novel genes may underlie inherited susceptibility to breast/ovarian cancer in Jewish individuals. PMID- 11102979 TI - Genetic testing for hereditary hearing loss: connexin 26 (GJB2) allele variants and two novel deafness-causing mutations (R32C and 645-648delTAGA). AB - Mutations in GJB2 are the most common cause of hereditary congenital hearing loss in many countries and are found in about half of persons with severe-to-profound congenital autosomal recessive non-syndromic hearing loss (ARNSHL). We report the results of GJB2 mutation screening in 209 consecutive persons with congenital deafness of indeterminate etiology using an allele-specific polymerase chain reaction assay, single-strand conformational polymorphism analysis, and direct sequencing. GJB2 allele variants were detected in 74 of 209 deaf individuals (35%). Over one-fourth of screened individuals were either homozygous (n=31) or heterozygous (n=24) for the 35delG mutation. Of those with the 35delG mutation, 51 (92.7%) were diagnosed with GJB2-related deafness. Nineteen persons were identified with other GJB2 allele variants - two novel deafness-causing mutations (R32C, 645-648delTAGA), one mutation of unknown significance (E47K), and one benign polymorphism (I128I). While these data enable health care professionals to provide parents and patients with improved genetic counseling data, difficulty still exists is determining whether some missense mutations compromise auditory function and are deafness-causing. PMID- 11102980 TI - Genetic variation in ICF syndrome: evidence for genetic heterogeneity. AB - ICF syndrome is a rare autosomal recessive immunoglobulin deficiency, sometimes combined with defective cellular immunity. Other features that are frequently observed in ICF syndrome patients include facial dysmorphism, developmental delay, and recurrent infections. The most diagnostic feature of ICF syndrome is the branching of chromosomes 1, 9, and 16 due to pericentromeric instability. Positional candidate cloning recently discovered the de novo DNA methyltransferase 3B (DNMT3B) as the responsible gene by identifying seven different mutations in nine ICF patients. DNMT3B specifically methylates repeat sequences adjacent to the centromeres of chromosome 1, 9, and 16. Our panel of 14 ICF patients was subjected to mutation analysis in the DNMT3B gene. Mutations in DNMT3B were discovered in only nine of our 14 ICF patients. Moreover, two ICF patients from consanguineous families who did not show autozygosity (i.e. homozygosity by descent) for the DNMT3B locus did not reveal DNMT3B mutations, suggesting genetic heterogeneity for this disease. Mutation analysis revealed 11 different mutations, including seven novel ones: eight different missense mutations, two different nonsense mutations, and a splice-site mutation leading to the insertion of three aa's. The missense mutations occurred in or near the catalytic domain of DNMT3B protein, indicating a possible interference with the normal functioning of the enzyme. However, none of the ICF patients was homozygous for a nonsense allele, suggesting that absence of this enzyme is not compatible with life. Compound heterozygosity for a missense and a nonsense mutation did not seem to correlate with a more severe phenotype. PMID- 11102981 TI - Evaluation of DHPLC analysis in mutational scanning of Notch3, a gene with a high G-C content. AB - Notch3 mutations cause CADASIL, an increasingly recognized cause of subcortical ischemic stroke and vascular dementia in human adults. In the absence of any specific diagnostic criteria, CADASIL diagnosis is based on mutational scanning of Notch3, which is a large gene composed of 33 exons with a high G-C content. In this study we examined the sensitivity of denaturing high performance liquid chromatography (DHPLC). First we established the theoretical optimal parameters, then we examined a large collection of amplicons in which we had previously identified distinct pathogenic mutations or polymorphisms. We further performed Notch3 mutational scanning in five patients suspected of CADASIL diagnosis in which previous scanning, including SSCP and heteroduplexes analysis, failed to detect any pathogenic mutation. DHPLC resolved 97% of mutations previously detected by sequencing and allowed identification of two novel pathogenic mutations: R607C and F984C. These data indicate that DHPLC is a sensitive screening method particularly suitable for epidemio-genetic screening of CADASIL. PMID- 11102982 TI - Mutational analysis of GLUT1 (SLC2A1) in glut-1 deficiency syndrome; dong wang; pamela kranz-eble; darryl C. De vivo; (Article was originally published in human mutation 16:224-231, 2000) AB - An error was made in the reproduction of Figure 2. Therefore, the corrected version is being reprinted here. PMID- 11102983 TI - Novel thymidylate synthase enhancer region alleles in African populations. AB - Thymidylate synthase (TS) regulates the production of DNA synthesis precursors and is an important target of cancer chemotherapy. A polymorphic tandem repeat sequence in the enhancer region of the TS promoter was previously described, where the triple repeat gives higher in vitro gene expression than a double repeat. We recently identified ethnic differences in allele frequencies between Caucasian and Asian populations. We now describe assessment of genotype and allele frequencies of the TS polymorphism in 640 African (African American, Ghanaian and Kenyan) and Caucasian (UK, USA) subjects. The double and triple repeat were the predominant alleles in all populations studied. The frequency of the triple repeat allele was similar between Kenyan (49%), Ghanaian (56%), African American (52%), American Caucasian (54%) and British Caucasian (54%) subjects. However, two novel alleles contained 4 and 9 copies of the tandem repeat. These novel alleles were found at a higher allele frequency in African populations (Kenyan 7%, Ghanaian 3%, African American 2%) than Caucasians (UK 1%, USA 0%). The novel alleles identified in this study decrease in frequency with Western migration, while the common alleles are relatively stable. This is a unique example suggesting the influence of multiple selection pressures within individual populations. Hum Mutat 16:528, 2000. PMID- 11102984 TI - BTK mutations in patients with X-linked agammaglobulinemia: lack of correlation between presence of peripheral B lymphocytes and specific mutations. AB - X-linked agammaglobulinemia (XLA) is a human antibody deficiency that results from mutation of the tyrosine kinase btk. We tested the hypothesis that XLA patients who varied from the classic phenotype of XLA by presence of normal or near normal number of peripheral B lymphocytes would have a set of mutations of BTK that is different from the mutations found in patients without peripheral B lymphocytes. The mutations of BTK we found in two patients with normal numbers of peripheral B lymphocytes have been previously identified in patients without peripheral B lymphocytes. A third patient, without peripheral B cells, was found to express normal levels of wild type btk. Exmination of the mutations of the BTK gene in patients in the BTKbase who were identified as having peripheral B lymphocytes found that these same mutations, or mutations of the same protein domains, were also present in patients identified as lacking peripheral B lymphocytes. Analysis of mutations in BTK has previously led to the conclusion that severity of disease in XLA cannot be predicted from the specific mutation of BTK. The results of this study suggest that whether an XLA patient will develop peripheral B lymphocytes cannot be predicted from the specific mutation of BTK. PMID- 11102985 TI - Low frequency of ankyrin mutations in hereditary spherocytosis: identification of three novel mutations. AB - Hereditary spherocytosis (HS) is a common hemolytic anemia caused by defects in the erythrocyte membrane proteins. The screening of mutations in the ankyrin-1 (ANK1) gene of 28 Brazilian HS patients showed two new missense mutations (His276Arg and Ile1054Thr) and one novel promoter mutation (-153 G-->A). The His276Arg mutation affected the invariable TPLH sequence on repeat 9. The -153 mutation was linked in cis to the known -108 T-->C mutation. In contrast to other populations, we were able to detect mutations in the ankyrin-1 gene in only 10% of our patients. It is also interesting to point out that, from 15 informative subjects for the 3' Acn repeats, only one presented a loss of heterozigosity at the cDNA level. Taken together, these results suggest that mutations in the ankyrin-1 gene might not be as common in Brazil as described for other populations. PMID- 11102986 TI - Ten novel BRCA1 and BRCA2 mutations in breast and/or ovarian cancer families from northern Germany. AB - Germline mutations in the BRCA1 and BRCA2 gene account for the majority of high risk breast/ovarian cancer families. We have screened such families from Northern Germany by using DHPLC analysis and subsequent direct sequencing techniques. In ten families we identified six novel BRCA1 and 4 novel BRCA2 mutations comprising four frame shift mutations, one nonsense and one splice site mutation in the BRCA1 gene as well as three frameshift mutations and one nonsense mutation in the BRCA2 gene. Our analysis contributes to the further characterisation of the mutational spectrum of BRCA1 and BRCA2. PMID- 11102987 TI - Polymorphisms in a pseudogene highly homologous to PMS2. AB - PMS2 is one of a complex of genes encoding DNA repair proteins that includes MSH2, MLH1, MSH6 and MSH3. Mutation of any of these DNA mismatch repair genes leads to impairment of DNA repair and can lead to tumorigenesis. Germline mutation of PMS2 has been reported as a rare cause of hereditary nonpolyposis colorectal cancer (HNPCC) and Turcot's syndrome. The PMS2 gene is located on chromosome 7p22 and consists of 15 exons. Within exon 11 of PMS2 is a coding repeat of eight adenosines. This study reports on the finding of a nonexpressed pseudogene that is highly homologous to the PMS2 gene in this region. The pseudogene is polymorphic for two alterations in the repeat region: a 3 bp delAAA at a site corresponding to nucleotide 1231 in PMS2; and an AA-->GG change at nucleotide 1238. Due to the high homology in both intronic and exonic sequences, polymorphisms in this pseudogene could be mistaken for mutations in the PMS2 gene and erroneously thought to be a cause of HNPCC and/or Turcot's syndrome. PMID- 11102988 TI - Frameshift mutations with severe and moderate clinical phenotypes in Thai hemophilia A patients. AB - Six frameshift mutations in exon 14 of the factor VIII gene were identified in Thai hemophilia A patients. Although all these mutations created premature stop codons and expected to cause severe disease, the molecular defects and clinical severity were in discrepancy in some patients. Four mutations (delT3490, delACAC3618-21, delGA4429-30, and delA4658) were found in the patients with the severe clinical phenotype while two (delA3629-37 and insA4372-9) were observed in the patients who had moderate severity, with FVIII:C of 4.2 and 2.8%. The frameshift mutations in these two patients were due to deletion and insertion of an 'A' nucleotide in the stretches of 9As and 8As in codons 1191-4 and 1439-41, respectively. This indicates that deletion or insertion in the stretches of poly A nucleotides in exon 14 of the factor VIII gene is a likely cause of the moderate clinical severity in some cases of Thai hemophilia A patients. PMID- 11102989 TI - Molecular analysis of the TFR2 gene: report of a novel polymorphism (1878C>T). PMID- 11102990 TI - Two new mutations (H106P and L178V) in the protoporphyrinogen oxidase gene in Argentinean patients with variegate porphyria. PMID- 11102991 TI - Identification of a novel T398A mutation in the ND5 subunit of the mitochondrial complex I and of three novel mtDNA polymorphisms in two patients presenting ocular symptoms. PMID- 11102992 TI - A novel stop mutation in exon 18 (W1145X) of the CFTR (ABCC7) gene in an adult CF patient. PMID- 11102993 TI - TIGR/MYOC proximal promoter GT-repeat polymorphism is not associated with myopia. PMID- 11102994 TI - A novel mutation E179K of the MEN1 gene predisposes for multiple endocrine neoplasia-type 1 (MEN1). PMID- 11102995 TI - Novel silent variant (c1722G>A) in the coding region of the insulin receptor substrate 1 (IRS1) gene. PMID- 11102996 TI - Novel variants in 3 kb of 5'UTR of the beta 1-adrenergic receptor gene (-93C>T, 210C>T, and -2146T>C): -2146C homozygotes present in patients with idiopathic dilated cardiomyopathy and coronary heart disease. PMID- 11102998 TI - Antibiotic Selection in the Treatment of Febrile Neutropenia: Current Approach and New Directions. AB - The combination of neutropenia and fever, with or without documented infection is a common complication in cancer patients receiving chemotherapy. The prompt institution of empirical broad-spectrum antibiotic therapy for febrile neutropenic patients has been shown to reduce the rate of early death and morbidity in cancer populations undergoing chemotherapy. Three different regimens for consideration as initial empiric therapy are presented. Alternatives such as outpatient regimens to low risk patients and monotherapy are discussed. If confirmed prospectively, the results presented here may enable clinicians to identify low risk groups and treat neutropenia and fever with good results and better economic and psychological cost. PMID- 11102997 TI - A de novo adrenoleukodystrophy gene (ABCD1) mutation S636I without detectable ABCD1 protein and a R104C mutation with normal amounts of protein from an Austrian patient collective. PMID- 11102999 TI - Management of Opportunistic Infections in HIV(+) Patients: Contrasts Between Europe and South America. AB - The author discusses the management of some opportunistic diseases more commonly observed in South American AIDS patients than in European ones. Characteristics of coinfection with HIV and leprosy, paracoccidioidomycosis, Chagas' disease, mucocutaneous leishmaniasis, malaria, disseminated BCG and strongyloidiasis are reviewed, with special emphasis on preferred therapeutic schedules for these conditions. PMID- 11103000 TI - Frequency, Type and Associated Diseases of Bacteria and Virus in The Oropharynx of Children Born to Human Immunodeficiency Virus-Infected Mothers. AB - HIV-infected children are more likely than other children to develop pneumonia, which in these children is often recurrent or persistent. The main reservoir of the major pathogens is the nasopharynx, but to date no data has been published on the frequency and biologic characteristics of S.pneumoniae, H.influenzae and respiratory viruses found in the upper respiratory tract of children born to human immunodeficiency virus-infected mothers. To document these aspects, 105 children was monitored by pharyngeal swab (PS) and nasopahryngeal aspirates (NPA) who attended an outpatient clinic for HIV-infection evaluation. Bacterial identification was performed by standard procedures. Serotype, biotype and beta lactamase production was investigated in H.influenzae isolates. S.pneumoniae serotypes were recognized by "quellung" and the susceptibility to 4 antibiotics was assessed. Respiratory syncytial viruses, parainfluenza, influenza A and B, and adenoviruses were diagnosed by indirect immunofluorescence and/or viral isolation in cell cultures. Twenty-nine children were identified as infected by HIV as a result of maternal-child transmission. Seventy children born to HIV positive mothers but who were not HIV-infected served as controls. Of 269 PS, 110 110 S. pneumoniae and 92 H.influenzae were identified. Also 31 viruses were detected in 188 NPA. After stratifying by age no differences were observed in the frequency of bacterial colonization or in the presence of viruses in the upper respiratory tract of the two groups. Some biologic characteristics of the agents were noteworthy such as the frequency of colonization by S.pneumoniae serotype 14, the predominance of H.influenzae biotype I and the high frequency of viruses in NPA of asymptomatic children. Of note, although colonization frequencies were similar, children presenting with acute respiratory illness (ARI) were more likely to have bacteria isolated if they also had HIV-infection than if they were HIV-negative. It is concluded that HIV-infection in infants as a result of maternal virus transmission have a similar frequency of bacteria and virus colonization of their respiratory tract, but a higher frequency of ARI and perhaps a higher frequency of types of bacteria with special characteristics. PMID- 11103001 TI - Risk Factors for HTLV-I Mother to Child Transmission: Influence of Genetic Markers. AB - This study was designed to evaluate the influence of genetic markers on the seropositivity of offspring of HTLV-I positive mothers in Tumaco, Colombia, an endemic area for HTLV-I infection and a site where there exists a racially mixed population of Black and Caucasian ancestors. 33 HTLV-I seropositive women with at least one offspring were studied. A total of 111 offspring were tested using hemaglutination-inhibition for testing sera for the allotypic markers G1m (1, 2, 3, 17) and G3m (5, 6, 13, 21). Potential risk factors such as mother s age at child's birth, mother's age at the time of the study, breastfeeding months, TSP vs. asymptomatic HTLV-I carrier, sibship's size, children's age and sex, were not found to be associated with mother to child transmission. Mother's Negroid genetic marker genotype (1, 17, 5, 13/1, 17, 5, +/- 13) was marginally associated with mother to child transmission of HTLV-I (P=0.0057; OR=11.97; CI=0.92-155.96) PMID- 11103002 TI - Human Immunodeficiency Virus Infection in Nigeria. AB - Study of the increasing epidemic of human immunodeficiency virus (HIV) infection in Nigeria provides insight into the magnitude of its spread, and allows identification of particular population groups which must be the target for preventive measures and increased public awareness campaigns. We have reviewed records documenting the disease in Nigeria since the first case was reported in 1986. Prevalence surveys have allowed identification of the rate of increase in the infection in various regions of Nigeria, and among various population groups: blood donors, those attending ante-natal care clinics (ANC), those attending sexually transmitted diseases clinics (STD), commercial sex workers (CSW), and patients with tuberculosis. We also reviewed the success of campaigns to increase awareness of AIDS in Nigeria. There were over 2 million cases of HIV infection in Nigeria in 1996, based on a national prevalence of 3.8%. The highest prevalence of infection was in the Middle Belt region of the country with 9.6% positivity of those tested in 1992, and 33.6% in 1994. Among blood donors, the percentage of those infected rose from 0% in 1987, to 4.4% in 1992, in one medical center; and in 1995, in Ante Natal Care clinics (ANC) the prevalence ranged from 0.2% to 12.9% of mothers (median 5.0%). High rates of infection were recorded among CSW ( 71.3% in one region), those attending STD clinics (27.4% in one region), long haul truck drivers and policemen. Patients with tuberculosis also had a high prevalence of infection. Public awareness and adequate surveillance data remain inadequate in view of the magnitude of the problem. Increased funding and judicious expenditures are essential to obtain and distribute accurate information in order to begin to resolve this epidemic. PMID- 11103003 TI - Vancomycin-Resistant Enterococcus faecium: First Case in Brazil. AB - We report a fatal case of septicemia due to a vancomycin-resistant Enterococcus faecium in a 9 year old girl with aplastic anemia. The isolate was also resistant to ampicillin, teicoplanin, gentamicin (high level), and streptomycin (high level). We believe that this is the first case of vancomycin-resistant Enterococcus (VRE) reported from a clinical specimen in Brazil. PMID- 11103004 TI - To Honor Hideyo Noguchi: 1876-1928. PMID- 11103005 TI - Statistical Inference (part II): The Normal and Related Distributions. AB - The normal (or gaussian) is the most important probability distribution in the statistical inference process. By transforming the normal distribution to a standard distribution (z-distribution) it is possible to determine the probabilities where observations or values fall in certain intervals for variables with different means and standard deviations. Areas under the normal distribution may be represented in a table where the z is the number of standard deviations away from the mean. For example, the area under the normal distribution delimited by a z value of 1.96 to the left and 1.96 to the right of the mean corresponds to 95% of the total area of the normal distribution. Sampling distribution of estimates of population parameters may also be described by the normal distribution. It is important to note, however, that the estimation of a population mean based on the normal distribution is conditional to the assumption that the population standard deviation is a known parameter. The t distribution is used to infer about a population mean when the population standard deviation is estimated by the sample data. In statistical inference about proportions, the normal approximation of the binomial distribution may be used provided the data fit certain assumptions. Statistical methods that do not depend on the form of the distribution (distribution-free or nonparametric methods) and those based on the actual probability distribution, called exact methods, are often used in situations where the data do not comply with the assumption that the distribution of the estimate is approximately normal. PMID- 11103006 TI - The Dynamics of the AIDS Epidemic in Brazil: A Space-Time Analysis in the Period 1987-1995. AB - The dynamics of the spread of the AIDS epidemic in Brazil have been studied by an analysis of cases reported between 1987 and 1995. The analysis evaluates characteristics of the epidemic as it changed over time, geographic region and special sub-populations affected. Data collected by the National Coordination of sexually transmitted diseases (STD's) and AIDS of the Health Ministry were reviewed and incidence rates calculated. Three periods of time were identified (1987-89, 1990-92, and 1993-95) and the distribution of cases by age group, sex, risk factor sub-population, and major geographic region recorded. Several observations can be made from this information: 1)The disease began among homosexual / bisexual males in large metropolitan areas of the southeast of Brazil. 2) It spread to other risk groups such as intravenous drug users (IDU) and recipients of blood products. 3) The diseases increased rapidly between 1987 89 and 1990-92, but then showed a plateau, particularly in the homosexual / bisexual group. 4) Increase in cases persisted in the IDU group. 5) The incidence among heterosexuals and in smaller municipalities has continued to increase indicative of a shift in the dynamics of the epidemic. The shift in the characteristics of the epidemic away from special groups and towards the socially vulnerable, poorly educated, heterosexual, smaller municipalities require new preventive strategies adaptable to regional patterns of each social structure, economy and culture. PMID- 11103007 TI - Effect of Antifungal Agents on Candida spp. and Pichia anomala Isolated from Oropharyngeal Candidiasis of AIDS Patients in a University Hospital in Brazil. AB - In order to determine the clinical significance of oropharyngeal candidiasis in AIDS patients, 44 isolates collected from individual clinical episodes were evaluated. The isolates were identified by microbiologic standard methods and in vitro antifungal susceptibility testing was evaluated for amphotericin B, fluconazole, flucytosine, miconazole, itraconazole, and ketoconazole according to the National Committee for Clinical Laboratory Standards. Candida albicans ATCC90028 was used as control for the MICs. The microbiologic isolation revealed 2 strains of Pichia anomala, an uncommon pathogen in AIDS patients. C.albicans and Candida tropicalis comprised 35 and 6 isolates, respectively. The antifungal susceptibility testing for C.albicans isolates (35 isolates) revealed low sensitivity and 19 strains (54%) were resistant to at least 1 antifungal agent. 5 strains (14%) showed multi-drug resistance, including to fluconazole. The resistant profiles of these Candida spp. are of major concern since at present, fluconazole is not commonly prescribed for the treatment of oral candidiasis in our AIDS patients. All C.tropicalis isolates were resistant to fluconazole and miconazole. Both P.anomala strains were resistant to fluconazole. The local diversity of species and the variability of MICs and IC80s of antifungal agents seen in this study indicate that continuing evaluation of clinical and laboratory aspects of oral candidiasis in our HIV1-patients is needed, and that a new approach and therapy will be necessary to manage the increasing problem. PMID- 11103008 TI - HIV Seroprevalence and Risk Factors in a Brazilian Prison. AB - The burden of the AIDS epidemic in Brazil is characterized by high prevalence rates in specific groups, including prison populations. Reports o HIV seroprevalence rates from several, prisons in different countries have varied widely. Those rates have usually reflected the HIV seroprevalence in the community served by the correctional facility studied and the risk factors of inmates for HIV infection. The present study was designed: 1) to determine the HIV seroprevalence among inmates of Casa de Detencao de Sao Paulo; 2) to identify independent risk factors for HIV acquisition; and 3) to determine the relevance of transmission of HIV infection within the prison. From December 20, 1993, through January 5, 1994, 780 inmates were interviewed using a standardized questionnaire and had their blood drawn for HIV testing by ELISA with confirmatory Western Blot. Out of 766 inmates tested, 637 (83.1%) were negative for HIV, 105 (13.7%) were positive, and 24 (3.1%) had indeterminate test results. Multivariate logistic regression analysis identified the following variables as independent risk factors for HIV seropositivity: 1) age less than 29 years-old; 2) previous incarceration in Casa de Detenca; 3) more than one sexual partner in the last year in Casa de Detenca; and 4) intravenous drug use before admission to Casa de Detenca. We conclude from this study that HIV infection among prisoners is high (13.7%) and that several risk factors are responsible. Of these, intravenous drug use before imprisonment is the most likely factor, but HIV transmission can also occur during incarceration. PMID- 11103009 TI - Aggregation of Rotavirus Particles and Improvement of the Detection of Rotavirus RNA by Polycrylamide Gel Electrophoresis After Treatment With Tween-80. AB - Treatment of feces rotavirus-positive with Tween-80 at 0.25% enhances by a factor of 4-fold the viral detection of rotavirus RNA by polyacrylamide gel electrophoresis (PAGE) after silver impregnation. By use of electron microscopy 35% of the rotavirus particles in untreated feces are aggregated, but after Tween 80 treatment, the percentage aggregated is decreased significantly to approximately 6%. Disaggregation of virus particles by Tween-80 can amplify the range of rotavirus RNA detection in feces. PMID- 11103011 TI - New Roles for Infectious Diseases Physicians in the Hospital; Leadership, Organization, and Teaching by Example. PMID- 11103010 TI - Successful Treatment of Mucormycosis and Aspergillus sp. Rhinosinusitis in an Immunocompromised Patient. AB - Fungal rhinosinusitis is a serious infection related to immunosuppressive conditions. It is caused by common opportunistic mycoses, such as Mucor, Aspergillus sp. and Candida sp. Treatment is difficult and most of the patients with an invasive form, die. We report a case of a 13 year old boy who developed Mucor and Aspergillus sp rhinosinusitis while receiving chemotherapy for acute lymphoblastic leukemia. Even though the patient was intensely immunossupressed, early diagnoses associated with surgical debridement and effective doses of amphotericin B allowed a favorable outcome and hospital discharge. PMID- 11103012 TI - Statistical Inference (Part 3): Statistical Hypothesis Testing and Confidence Interval Estimation. AB - An association between an independent and a dependent variable found in a study may have several explanations, including chance (i.e., random error). This article presents two approaches to assess the role played by chance in an association: confidence interval estimation and statistical hypothesis testing. Statistical hypothesis testing estimates the probability (i.e., the P value) of getting a difference as large or larger than the one observed in a specific study assuming the absence of association. Confidence intervals are estimates of the range of values that are expected to include the actual parameter with a certain probability, or confidence level (often 0.95 or 95%). PMID- 11103014 TI - Response of HIV/AIDS Patients to Hepatitis B Recombinant Vaccine. AB - Studies have demonstrated that HIV infection negatively affects the immune response to hepatitis B vaccine. The present study evaluated the seroconversion to the recombinant vaccine against hepatitis B applied in HIV patients. Twenty two patients were included in the study group all with confirmed HIV infection and with negative serum markers to hepatitis B. The control group was composed of 18 healthy individuals with negative markers for hepatitis B. All subjects were vaccinated with 20ug of ENGERIX B((R)) at 0, 1 and 6 months (3 doses). The antibody response was quantitatively assessed 1 month after the third dose of recombinant vaccine. CD4 T lymphocyte counts were also performed in those beginning vaccination. Of 22 patients in the study group, only 10 (45.5%) responded to vaccination with protective levels (over 10mIU/ml). In the control group, all of the subjects responded (p=0.005). Seventeen patients in the study group had their CD4 lymphocytes measured. The results suggested a direct relationship between the level of CD4 lymphocyte counts and response to the vaccine. The rate of response to hepatitis B recombinant vaccine with 3 doses of 20ug of HBsAg in patients infected by the human immunodeficiency virus was significantly lower than in the control group. Patients with low CD4 T lymphocyte counts are likely to have an inadequate response to the current method of hepatitis B vaccination. PMID- 11103013 TI - Comparison of Ritonavir in a First Choice Three Drug Regimen, After Two Nucleoside Analogues and in Saquinavir Experienced Patients. AB - Ritonavir is a potent, orally bioavailable inhibitor of HIV-1 protease. Our investigators undertook a retrospective study to compare the effectiveness of ritonavir (600mg twice daily) associated with 2 reverse transcriptase inhibitors (RTIs) in 38 patients in 3 situations. Group I patients previously treated with 2 RTIs, Group II treatment-naive patients, and Group III patients previously treated with 2 RTIs and saquinavir. Routine hematological and biochemical studies, HIV-1 viremia, and CD4+ lymphocyte counts were performed before and after ritonavir. In Group I, the median of HIV-1 RNA plasma levels decreased from 4.8 to 3.4 log(10) copies/mL, in Group II from 5.9 to 2.9 log(10) copies/mL, and in Group III from 5.2 to 4.1 log(10) copies/mL. (p=0.003, p=0.014, p=0.002, respectively, Wilcoxon signed rank test). The median increases of CD4(+) cells occurred as follows: in Group I from 173 to 282 cells/mm(3), in Group II from 92 to 254 cell/mm(3), and in Group III from 68 to 133 cell/mm(3) (p=0.002, p=0.008, p<0.001, respectively, Wilcoxon signed rank test). In Group II the mean weight increased from 55.2 +/-14.3 kg to 59.4+/-15.7 kg and, in Group III, from 62.2+/ 10.5 kg to 67.5+/-12 kg (p = 0.026, p = 0.002, respectively, paired T test). Patients in Group I presented no weight gain. Mild reversible hypertriglyceridemia occurred in 6 of 38 patients. The results of this study showed that ritonavir is a good choice for treatment naive patients and as a sequential option, not only after 2 RTIs, but also after a 3 drug regimen with saquinavir. PMID- 11103015 TI - Piperacillin/Tazobactam: Evaluation of Its In vitro Activity against Bacteria Isolated in Two Brazilian Hospitals and an Overview of Its Antibacterial Activity, Pharmacokinetic Properties and Therapeutic Potential. AB - Piperacillin/tazobactam is a highly active antibiotic against most clinically important species of Gram-negative and positive bacteria, including anaerobes. It has never been used or tested against bacteria isolated in Brazil. In this article we report the in vitro activity of piperacillin/tazobactam against clinical isolates from two tertiary hospitals in Sao Paulo and Rio de Janeiro and review the evolving clinical literature. The study was performed before its commercialization in Brazil. Its activity was compared to that of several broad spectrum antimicrobial agents commercially available in Brazil. Piperacillin/tazobactam was active against 83% of the isolates tested, while imipenem was active against 91%, cefepime 77%, and ciprofloxacin 73%. Against Enterobacteriaceae (n=398), cefepime was more active than piperacillin/tazobactam (92% versus 88%). Klebsiella pneumoniae strains (n=95) presented low susceptibility to piperacillin/tazobactam (79%), ceftazidime (67%), and to cefepime (82%) indicating a high percentage of ESBL-producing strains. The most active compounds against this species were imipenem (100%) and ciprofloxacin (93%). Piperacillin/tazobactam was the most active compound against Gram-positive cocci (n=238; percentage of susceptibility rank order: piperacillin/tazobactam = imipenem > ciprofloxacin > cefepime) and the second most active against nonenteric Gram-negative bacilli (n=250, rank order: imipenem [72%] > piperacillin/tazobactam [60%] > cefepime [56%] > ceftazidime [52%] > gentamicin [45%] > ciprofloxacin = aztreonam [42%]). Cefepime was the most active compound against P. aeruginosa (n=128, only 67%), followed by ceftazidime (64%), piperacillin/tazobactam (63%) and imipenem (59%). Only imipenem (91%) was active against more than 50% of the A. baumannii isolates (n=79) tested. Piperacillin/tazobactam was the second most active compound against A. baumannii (49%) and the most active against B. cepacia (91%). Our results demonstrated a high level of antimicrobial resistance in the hospitals evaluated, especially among nonenteric Gram-negative bacilli; and clonal dissemination of multiresistant strains. Piperacillin/tazobactam may contribute to the treatment of nosocomial infections in Brazil, however, some degree of resistance was detected in some species in the instance of frequent multiresistant bacteria in the tertiary level hospitals where the drug was evaluated. PMID- 11103017 TI - Statistics in Medicine; A Look at the Problems, and a Challenge for the Future. PMID- 11103016 TI - Semilunar Valvar Stenosis Associated With Congenital Rubella Syndrome. AB - We studied four children who were born with congenital rubella syndrome, in whom cardiac malformations including semilunar valvar stenosis were present. These children attended the Pediatric Cardiology Unit of Hospital Fundacao Santa Casa de Misericordia do Para. The diagnosis was made clinically and by laboratory results. Specific anti-rubella antibodies IgM and IgG were measured using a capture-antibody enzyme immunoassay. Three patients presented with aortic stenosis associated with patient ductus arteriosus, and 1 patient had isolated pulmonary valvar stenosis. Rubella-specific IgM antibodies were detected in 3 of the 4 children and IgG antibodies in 1. We review the role of rubella virus as a teratogenic agent implicated in a pathogenesis of aortic and pulmonary valvar stenosis. These pathologic lesion are part of the diffuse ateriopathy that affects the fetus with rubella virus infection. PMID- 11103018 TI - Epidemiology of the Ebola Virus: Facts and Hypotheses. AB - Marburg and Ebola viruses are emerging pathogens recognized since 1967, and in 1976, when they were first identified. These viruses are the only members of the Filoviridae family. They cause severe, frequently fatal, hemorrhagic fever. Each genus includes some serotypes with the distinctive characteristics to cause high mortality rate during outbreaks. The Ebola-Zaire subtype is the most lethal variant. The epidemiology of human pathogenic filovirus is reviewed in this paper considering the most relevant facts. Primary human cases arise probably through close contact with infected primates. This point may be the key to preventing the introduction of these viruses in human populations. Once introduced in humans, the infection may spread through close contact with infected individuals or their body fluids, particularly in hospital environments. A main feature of filovirus outbreaks is the occurrence of cycles of secondary infection. PMID- 11103019 TI - Estimated Prevalence of Viral Hepatitis in the General Population of the Municipality of Sao Paulo, Measured by a Serologic Survey of a Stratified, Randomized and Residence-Based Population. AB - The present study was done to estimate the prevalence of Hepatitis A (HAV), B (HBV), C (HCV), and E (HEV) infection in the general population residing in the municipality of Sao Paulo, and to evaluate the level of knowledge related to the various modes of infection transmission by and protection against the different viruses. Blood samples and health questionnaires were collected from 1,059 individuals. The study design used an inductive method of predictive statistical inferences through randomized sampling stratified by Sex, age and residence region. The estimated prevalence rated found were: Hepatitis A = 66.59% (63.75% 69.44% CI); Hepatitis B = 5.94% (4.50%-7.35%); Hepatitis C = 1.42% (0,70%-2.12%); Hepatitis E = 1.68% (0.91%-2.46%). The frequency of hepatitis was similar in males and females. HAV showed an estimated prevalence of 56.16% in the population up to 17 years old, increasing to 65.30% in individuals between 18 and 29 years. The infection reached its peak of 90% in individuals 40 years of age or older. The study showed a greater tendency of dissemination of HBV among the population between 15 and 17 years. This specific age group showed an estimated prevalence of active infection of 1.04% (0.43%-1.65% CI), and also demonstrated an ascending level of acquired immunity with an estimated prevalence of 4.90% (3.60%-6.20% CI). HCV demonstrated an estimated prevalence of 1.42% (0.70%-2.12% CI). This specific infection occurred more frequently among adults 30 years of age or older, with the prevalence reaching a peak of 3.80% among the group aged 50 to 59 years. HEV showed zero prevalence among the age group between 2 and 9 years. This was followed by a slightly ascending rate starting from age 10, with an estimated prevalence of 1.05% (0.94%-3.04% CI) among those 10 to 14 years of age. This infection reached its peak of 3.00% (0.55%-6.74% CI) at the age of 60 years or older. Individuals with lower educational levels had a higher tendency of acquiring HAV and HCV, while there was no statistically significant difference for this parameter related to HBV and HEV. HBV occurred more frequently among inhabitants of the northern region of the city. All other hepatitis forms occurred at similar frequencies among the five regions of the city. Among the population, 1.90% (1.08%-2.72% CI) demonstrated an elevated hepatic enzyme with no serologic evidence indicating the cause was the viruses studied. This observation suggests the presence of other hepatic diseases, possibly including other viral diseases. It was also estimated that 75.12% of the city's population did not know the modes of transmission of hepatitis viruses and 76.70% did not know how to prevent them. This clearly suggests the need for a full-scale education program combined with public health measures regarding prevention of all forms of vial hepatitis. PMID- 11103020 TI - Morphofunctional Study of Blood Polymorphonuclear Leucocytes in HIV-Seropositive Individuals. AB - The impairment of leukocytic functions in AIDS infected individuals, where opportunistic infections are manifested, has been under study since 1985. However, controversy remains concerning leukocyte function during initial stages of HIV infection. In the context of the precarious immunologic and phagocytic defense of persons with AIDS, and the resulting difficulty they have to control microorganism invasion and opportunistic infections, examination of the host defense functions played by leukocytes in seropositive HIV persons is particularly important. To that end, our study sought to assess, during the initial stage of HIV infection, the laboratory parameters associated myeloid cells which are known to be altered during disease stage. Seventy-five (75) persons seropositive to HIV-1 (by the ELISA test, confirmed by immunofluorescence), and twenty-six (26) controls were tested. These individuals were screened by infectologists and their disease severity classified according to the Walter Reed Army Institute System. We observed that myeloperoxidase enzyme activity, superoxide anion production, and fungicidal action in homologous serum were all diminished in patients classified as WR-1, and progressively decreased in WR-2-4. The percent phagocytizing neutrophils and the number of C. albicans phagocytized were normal in WR-1 patients, but diminished in WR-2-4. We conclude that neutrophil function is diminished in HIV-infected persons at the beginning of infection, and that the defects increase as the HIV disease progresses. PMID- 11103021 TI - Drug Resistance in Bacteria Isolated From a Brazilian Hospital. AB - Bacteria commonly associated with cases of hospital infection were isolated from samples of food, from food handless, and from objects and surfaces from different places of a hospital in Piracicaba, Sao Paulo, Brazil, and the resistance patterns to antibiotic of these strains of bacteria were evaluated. The resistance patterns of these bacteria showed a large variation, and a high frequency of resistance to ampicillin (60.9%), cephalothin (58.7%) and carbenicillin (52.2%) was observed. The frequency of resistance to cephalosporins of 3rd and 4th-generations was 26.1% and 17.4% of the samples, respectively. Resistance to more than two drugs was observed in 27 samples (56.5%), and in four strains multiple resistance to 17 or more tested drugs was recorded. Five bacteria which were multi-resistant to antibiotics (Enterobacter aerogenes, Escherichia coli, Proteus sp, Pseudomonas sp and Staphylococcus aureus) were studied to determine the chromosomal or plasmidial genetic basis of the resistance, using plasmid curing and agarose gel electrophoresis of plasmidial DNA. It was possible to verify that for the antibiotics chloramphenicol and kanamycin, the resistance seems to be of plasmidial origin. PMID- 11103022 TI - Baccilary Angiomatosis: Negative Results Using Normal Balb/c and Balb/c Nude Mice. AB - Baccilary angiomatosis has recently been described as a disease that can spread systematically and that is potentially fatal. It is caused by Bartonella henselae and B. quintana, and presents as especially pronounced signs and symptoms in patients suffering from acquired immunodeficiency syndrome (AIDS). To clarify the pathogenesis of the disease and to try to define the relationships among baccilary angiomatosis, cat scratch disease and Carrion's bartonellosis, the authors of this study have attempted to develop an experimental model using mice that were immunocompetent as well as those that had their cellular immunity genetically compromised. A known concentration of B. henselae was inoculated intradermally in Balb/c an isogenic mice or an athymic group of the same lineage. Blood samples were taken on days 0, 3, 7, 10, 14, 28, and 60 after inoculation for indirect immunofluorescence antibody testing. On the 21st and 60th day, one animal from each group was sacrificed and a post mortem carried out including histological evaluation of the liver, spleen, lymph nodes, skin and other organs. Hemocultures of the sacrificed animals were collected. All results of serologic response, cultures and histologic examination were negative. The authors discuss the methodology, especially the use of isogenic animals of the same lineage in B. henselae infection, with and without immunodeficiency, and the resources for the negative results of histopathology, serology and cultures. PMID- 11103024 TI - Careful Evaluation of Clinical and Pre-Clinical Data is Needed -- Not More Clinical Trials! PMID- 11103023 TI - Facial Cutaneous Anthrax in a Pregnant Woman: a Case Report. AB - Anthrax remains an uncommon, but worldwide problem, particularly in countries in which domestic animals and processing of animal by-products are an important part of the economy. The disease has received attention recently because of its potential for use in biologic warfare. In Poland during the last 10 years, several human cases of cutaneous anthrax occurred. We report here a case of a pregnant woman with this disease. The lesion was atypical and in a potentially dangerous location since it was on the upper part of the face; a site which could be associated wth either respiratory or central nervous system complications. The patient recovered without complication after antibiotic treatment and local surgery. The fetus and the subsequent labor and delivery were not affected. The case is presented as a reminder of the continued presence of this disease, of the need for attention to its special clinical signs and symptoms, of the need to supervise the agriculture and textile industries, such as by use of vaccines when appropriate, and of the increased concern about this disease in regard to biologic warfare. PMID- 11103025 TI - Early coronary angioplasty for acute myocardial infarction complicated by cardiogenic shock: have novel therapies led to better results? AB - Patients with acute myocardial infarction (MI) and cardiogenic shock constitute a very high risk subset despite an aggressive management. The objective of this study was to evaluate if the results of early coronary angioplasty in patients with acute myocardial infarction and cardiogenic shock have changed over the last years, and to address which role the recent adjuvant therapies have played in this evolution. From 1991 to April 1999, 94 patients with acute MI and cardiogenic shock were treated with coronary angioplasty within the first 12 hours from the onset of symptoms. Temporal changes of the utilization of adjuvant therapies and operators experience were studied over these years, as well as their impact on the angiographic results and in-hospital outcome. Over the years, a progressive and significant increase on the use of coronary stents and c7E3Fab was observed, as well as an increased number of primary angioplasties performed per month. The proportion of patients treated with intraaortic balloon pump did not changed significantly over the years. An angiographic successful result (< 50% residual stenosis and TIMI flow 2 or 3) and a final TIMI grade 3 flow were obtained in 76 (80.9%) and 61 (64.9%) patients, respectively. The angiographic success rate progressively increased over the years, from 72.3% in patients treated before 1994 to 94.1% in those admitted in 1998Eth 1999 (p for trend 0.0409). The proportion of patients with a final TIMI grade 3 flow also grew progressively over the years: from 36.4% before 1994 to 76.5% after 1997 (p for trend 0. 0209). The overall in-hospital mortality rate was 63.8% (60 patients), and there was no significant change in mortality rate over the years. Therefore, apart from the growing operators experience, we have observed an incremental change in the use of coronary stents and c7E3 Fab (abciximab) in patients with acute myocardial infarction and cardiogenic shock treated with early coronary angioplasty. All these factors have led to an improvement in the angiographic results, although this change has not meant a significant reduction of mortality. PMID- 11103026 TI - 8 french transradial coronary interventions: clinical outcome and late effects on the radial artery and hand function. AB - BACKGROUND: Limited information is available on the effects of 8 French (Fr) transradial procedures on radial patency. In addition, the effects of radial procedures and radial occlusion on hand function are unknown. METHODS: Two groups were recruited: twenty-four patients who had undergone 26 transradial 8 Fr interventions and 16 patients who had undergone 16 transradial 6 Fr procedures. At 1 year, radial patency, hand strength and hand endurance were measured. RESULTS: No major adverse cardiac events or vascular complications were noted in either group. Late radial occlusion was noted in 2/18 (11%) 8 Fr patients and 3/16 (19%) 6 Fr patients (p = ns). There were no differences in the 8 Fr group between the catheterized and uncatheterized radial arteries for diameter (3.2 +/- 1.1 mm versus 3.3 +/- 0.7 mm, respectively; p = NS) or volumetric flow (55 +/- 51 ml/minute versus 57 +/- 45 ml/minute, respectively; p = NS). No differences in hand strength or hand endurance were seen between the catheterized and uncatheterized arms in the 8 Fr group, between the 8 Fr and 6 Fr groups, or between occluded and non-occluded patients. CONCLUSION: Transradial use of 8 Fr guiding catheters appears to be feasible and safe in highly selected patients, albeit associated with a low incidence of silent radial occlusion. Additionally, neither the use of 8 Fr sheaths nor the presence of radial artery occlusion appear to adversely affect hand strength or endurance. PMID- 11103027 TI - Radial interventions. PMID- 11103028 TI - De novo stenting of descending thoracic aorta in Takayasu arteritis: intermediate term follow-up results. AB - We report our intermediate-term follow-up results of de novo stenting of descending thoracic aorta in Takayasu arteritis. Six patients (5 males and 1 female) underwent aortoplasty and stenting (8 Wallstents were deployed in 6 patients). Aortoplasty was performed with conventional balloons in 5 patients and with an Inoue balloon in 1 patient. The mean diameter of the aorta increased from 5.36 +/- 0.62 mm to 13.91 +/- 1.8 mm after stenting while the peak systolic gradient was totally abolished in all cases. These results were significantly better than aortoplasty alone (p < 0.0001). Lower limb claudication improved in all patients. Hypertension was cured in 3 patients and improved in 3 patients. All patients were asymptomatic over 6Eth 30 months (mean, 22.8 +/- 4.0 months) follow-up. All patients underwent angiographic follow-up after 6 months. They continued to have an absence of gradient with excellent flow across the stents. A minimal intimal reaction was observed at the stent margins. On follow-up angiogram at 6 months, one of the patients was noted to have developed a small asymptomatic pseudoaneurysm at the lower margin of the stent, which gradually increased in size over the next year and was treated by percutaneous endovascular deployment of a Wallstent graft. Our series demonstrates the safety and efficacy of stent deployment in stenotic lesions of the aorta in Takayasu arteritis and proves that the results are superior to plain balloon aortoplasty alone. This is also the only study that demonstrates intermediate-term success by angiographic follow-up at 6 months. PMID- 11103029 TI - Efficacy of a new hemostatic device, Adapty , after transradial coronary angiography and intervention. AB - A novel hemostatic device, Adapty (Medikit, Tokyo, Japan), was developed to achieve effective and comfortable hemostasis following transradial procedures. The device consists of a pad fixed to a transparent plastic plate and a self adhesive strap. The catheter sheath is removed from the radial artery after the pad has been positioned precisely over the puncture site, with the strap attached to the plate. Compression pressure then is adjusted with the self-adhesive strap, as is required with occlusive clamps. Patients do not need to maintain hyperextension of the wrist after the procedure. The wrist can be mobilized immediately after application. The efficacy of Adapty was evaluated in prospective observations of 200 patients. The device was successfully applied in all patients immediately after sheath removal. No patient required interruption of compression because of pain, congestion or ischemia. Complete hemostasis was obtained in 199 patients (99.5%), and the device caused no vascular complications. This study demonstrates that Adapty is highly effective for achieving hemostasis after transradial procedures. PMID- 11103030 TI - Radial hemostasis devices: alternatives for a simple pile of gauze and elastic plasters? PMID- 11103031 TI - Determining appropriate small vessels for stenting by intravascular ultrasound. AB - To re-evaluate outcomes of stenting in small vessels, we studied 176 patients successfully treated with several types of stent by way of intravascular ultrasound (IVUS) and quantitative coronary angiography (QCA). These lesions were divided into 3 subgroups according to reference diameter (RD) by QCA, and vessel diameter (VD) by IVUS (group I: pre-RD 2.5 mm; group II: pre-RD < 2.5 mm and pre VD 4.0 mm; group III: pre-RD < 2.5 mm and pre-VD < 4.0 mm). Post-procedure percent diameter stenosis (4 +/- 3%), post-procedure percent plaque area (42 +/- 3%), and loss index (39 +/- 11%) in group II were not significantly different from those in group I (7 +/- 2%, 37 +/- 2%, 45 +/- 5%, respectively). The rates of restenosis and target lesion revascularization in group II (24.0% and 16.0%, respectively) were not different from those in group I (25.9% and 21. 2%, respectively), and were significantly favorable compared to group III (66.7% and 39.4%, respectively; p < 0.05). By the use of IVUS, we not only identified those vessels which would normally go unstented when only using QCA, but also documented excellent long-term outcome in these patients. PMID- 11103032 TI - Are small vessels "truly" small? PMID- 11103033 TI - "Skirt" technique for coronary artery bifurcation stenting. AB - Stent implantation in the treatment of coronary artery bifurcation lesions frequently impairs blood flow and gives the coronary tree a new, metallic configuration. The new technique we describe uses a single short stent in a "skirt" shape which produces no "jailing" effects and can be used in the treatment of true coronary Y-shaped bifurcation lesions. PMID- 11103034 TI - A novel strategy to overcome resistance in stent placement at lesion site after adequate predilatation. AB - Resistance was encountered in passing a 3 x 18 mm stent across a lesion in the proximal left anterior descending coronary artery. Successive changes in stent with repeated balloon dilatations did not succeed. Finally, a 9 mm stent was passed across the lesion and deployed at the site of maximal resistance. The 18 mm stent was then placed through this stent. A novel strategy to overcome resistance in the stent passage through the lesion after an adequate balloon predilatation is reported. PMID- 11103035 TI - Percutaneous stenting of anomalous coronary arteries. AB - We present a case series of stenting de novo lesions in anomalous coronary arteries. Our experience suggests that appropriate guiding catheter selection would allow diseased anomalous coronary arteries to be amenable to rotational atherectomy, excimer laser coronary angioplasty or brachytherapy. PMID- 11103036 TI - IVUS based dosimetry and treatment planning. PMID- 11103037 TI - Comparison of Clinical Features of Acute HIV-1 Infection in Patients Infected Sexually or Through Injection Drug Use. AB - Acute HIV-1 infection (AHI) may present with a clinical picture that represents a diagnostic challenge. We tested the hypothesis that two different routes of infection, that is, sexual versus parenteral, might be associated with a difference in the clinical features of AHI. A prospective cohort of seroconvertors was established in Montreal in private medical clinics and hospitals from February 1996 to May 1999. The prevalence of the symptomatic presentation was almost overlapping within the two groups of newly infected individuals 69% (42 of 61) for men having sex with men (MSM) and 69% (18 of 26) for injection drug users (IDUs; p =.98). Comparison of all types of symptoms and signs as well as their duration was also similar in both groups. Of particular interest, the site of lymph node enlargement was not different despite the estimated sites of intravenous inoculation. Oral and anal ulcers were more frequently observed in MSM than in IDUs (6 versus 0 and 4 versus 1, respectively). Neither the mean CD4+ count (514.8 and 414.7 cells/mm3; p =.14) nor the mean viral load (4.45 and 4.70 log copies/ml; p =.40) were different between the two groups at the time of the first study visit. Our study results clearly indicate that health care workers can expect similar clinical presentation of AHI in MSM and in IDUs despite the different routes of infection. PMID- 11103038 TI - Strong in vitro synergy between the fusion inhibitor T-20 and the CXCR4 blocker AMD-3100. AB - Attachment and entry of HIV-1 into CD4 cells involve a series of events in which different viral envelope proteins interact with specific cell receptors, culminating in fusion of viral and cell membranes. AMD-3100 is a small molecule inhibitor of HIV-1 attachment to the CXCR4 chemokine receptor, and T-20 is a synthetic peptide corresponding to a region of HIV-1 gp41 that blocks fusion to cell membranes. To evaluate the interaction between agents acting at two different steps of the entry process, we conducted in vitro studies of the combination of T-20 and AMD-3100 against an X4 HIV-1 isolate. Single drugs or multiply diluted fixed ratio combinations of drugs were added to peripheral blood mononuclear cells infected with a clinical isolate, 14aPre. Drug interactions were evaluated using the median-effect principle and the combination index technique. The 50% inhibitory concentration (IC50) for T-20 was 0.10 microg/ml and for AMD-3100 was 0.19 microg/ml. Synergy was observed between T-20 and AMD 3100 and this increased with higher inhibitory concentrations, with combination indices ranging from 0.62 at IC50 to 0.02 at IC95. Whether these synergistic interactions translate into clinical benefit will need to be addressed in the context of clinical trials. PMID- 11103039 TI - No association of HIV-1 envelope (C2-V3-C3) sequence pattern with long-term nonprogression. AB - We previously identified a group of 10 long-term nonprogressors (LTNP) with HIV-1 infection. In this study, we have sequenced the envelope gene (C2-V3-C3) from the 10 LTNPs and from a control group of 9 people with rapidly progressing infection (RPI). The 19 individuals' CCR5 genotype and virus phenotype (i.e., syncytium inducing/non-syncytium-inducing [SI/NSI]) were obtained from a previous study. A phylogenetic tree was constructed containing the 19 envelope sequences together with 42 local control env sequences obtained from other studies. Analysis of the phylogenetic tree did not reveal any relation between the envelope gene (C2-V3 C3) from LTNPs versus RPIs. When data from the CCR5 genotype and the virus phenotype were assembled in the phylogenetic tree, no significant clustering was observed. From alignment of the protein sequences, we found a possible N-glycan in position aa294 in env that was conserved in only 1 of 10 LTNPs; however, it was conserved in 6 of 9 RPIs. Our study could not demonstrate any association between LTNPs and the sequenced envelope gene segment (C2-V3-C3). This lack of association could be due to the relatively small sample size of the data set. Nor did we find any relation between the CCR5 genotype or the SI/NSI phenotype with the sequenced envelope genes from the 19 participants. The possible N-glycan position we have described is an interesting observation that may require further investigation. PMID- 11103040 TI - Effect of cryopreservation on measurement of cytomegalovirus-specific cellular immune responses in HIV-infected patients. AB - To determine the feasibility of cytomegalovirus (CMV)-specific cell-mediated immunity (CMI) studies using cryopreserved cells, we compared lymphocyte proliferation assays (LPA), responder cell frequency (RCF), interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) production using fresh and cryopreserved peripheral blood mononuclear cells (PBMCs) from 53 HIV-infected patients and 15 uninfected controls. Qualitative CMV LPA results were concordant in >/=84% of the specimens from either HIV-infected patients or controls. Proliferation-based RCF, IL-2, and IFN-gamma comparisons showed that cryopreservation reduces the number of CMV-specific responders and decreases cytokine secretion, without changing the rank order of the results (p <.01). In contrast, the number of flow cytometry enumerated IFN-gamma-producing CD4+ cells was not significantly changed by cryopreservation. In HIV-infected patients, the differences between fresh and frozen cell assays were not influenced by CD4 cell numbers or HIV viral load. These data indicate that cryopreserved cells are suitable for longitudinal studies of the CMV-specific immune response in HIV-infected patients and uninfected controls. PMID- 11103041 TI - Prevalence and predictors of highly active antiretroviral therapy use in patients with HIV infection in the united states. HCSUS Consortium. HIV Cost and Services Utilization. AB - BACKGROUND: Highly active antiretroviral therapy (HAART) became standard for HIV in 1996. Studies at that time showed that most people infected with HIV had initiated HAART, but that members of minority groups and poor people had lower HAART use. It is not known whether high levels of HAART use have been sustained or whether socioeconomic and racial disparities have diminished over time. OBJECTIVES: To determine the proportion of patients who had received and were receiving HAART by January 1998, and to evaluate predictors of HAART receipt. DESIGN AND PARTICIPANTS: Prospective cohort study of a national probability sample of 2267 adults receiving HIV care who completed baseline, first follow-up, and second follow-up interviews from January 1996 to January 1998. MAIN OUTCOME VARIABLES: Proportion currently using HAART at second follow-up (August 1997 to January 1998), contrasted with the cumulative proportions using HAART at any time before January 1998 and before December 1996. ANALYSES: Bivariate and multiple logistic regression analysis of population characteristics predicting current use of HAART at the time of the second follow-up interview. RESULTS: The proportion of patients ever having received HAART increased from 37% in December 1996 to 71% by January 1998, but only 53% of people were receiving HAART at the time of the second follow-up interview. Differences between sociodemographic groups in ever using HAART narrowed after 1996. In bivariate analysis, several groups remained significantly less likely to be using HAART at the time of the second follow-up interview: blacks, male and female drug users, female heterosexuals, people with less education, those uninsured and insured by Medicaid, those in the Northeast, and those with CD4 counts of >/=500 cells/microl (all p <.05). Using multiple logistic regression analysis, low CD4 count (for CD4 <50 cells/microl: odds ratio [OR], 3.20; p <.001) remained a significant predictor of current HAART use at the time of the second follow-up interview, but lack of insurance (OR, 0.71; p <.05) predicted not receiving HAART. CONCLUSIONS: The proportion of persons under HIV care in the United States who had ever received HAART increased to over 70% of the affected population by January 1998 and the disparities in use between groups narrowed but did not disappear. However, nearly half of those eligible for HAART according to the U.S. Department of Health and Human Services guidelines were not actually receiving it nearly 2 years after these medications were first introduced. Strategies to promote the initiation and continuation of HAART are needed for those without contraindications and those who can tolerate it. PMID- 11103042 TI - Eradication of cryptosporidia and microsporidia following successful antiretroviral therapy. AB - OBJECTIVES: Incidence of opportunistic protozoal infections causing diarrheal illnesses in patients with HIV has decreased since the introduction of highly active antiretroviral therapy (HAART). The objective of this study was to determine whether the parasites, cryptosporidia, and microsporidia were effectively eradicated or only suppressed following treatment. DESIGN: Six HIV positive patients with diarrheal symptoms caused by cryptosporidia or microsporidia were prospectively followed up with stool samples and duodenal biopsies. Samples were taken before HAART, between 1 to 3 months, and 6 months post-HAART. METHODS: Duodenal samples were analyzed using routine histology and transmission electron microscopy. Stool samples were analyzed by both light microscopy and polymerase chain reaction (PCR) techniques. RESULTS: Patients who responded successfully to HAART eradicated both cryptosporidial and microsporidial organisms. Symptoms improved within 1 month of therapy but complete eradication of the organisms was only observed after 6 months of treatment. CONCLUSIONS: AIDs-related cryptosporidiosis and microsporidiosis can be cured following successful antiretroviral therapy. PMID- 11103043 TI - Fat redistribution in indinavir-treated patients with HIV infection: A review of postmarketing cases. AB - CONTEXT: Fat redistribution (FR) occurring alone or in association with hyperlipidemia has been associated with protease inhibitors (PI) and nucleoside reverse transcriptase inhibitors (NRTIs); however, the relationship between FR features, relationship of FR to hyperlipidemia, and pathogenesis of FR is unknown. OBJECTIVE: To characterize the spectrum of FR, assess relationships among FR features, determine trends in occurrence of FR, and determine relationship of FR to hyperlipidemia. DESIGN: Review of postmarketing indinavir reports of FR in Merck & Co. Inc.'s database. SETTING AND PARTICIPANTS: 282 reports of FR among HIV-positive patients taking indinavir submitted through the passive postmarketing system to Merck through February 23, 1998. RESULTS: 282 FR reports were compared across 3 groups: fat accumulation (FA) only, FA with peripheral wasting (FA with PW), and peripheral wasting only (PWO). Of 282 reports, 56% (159 of 282) had FA only, 22% (63 of 282) had FA with PW, and 21% (60 of 282) had PWO. The proportions of reports of PWO was higher in men, whereas the proportion of reports of FA was higher in women. Blood lipids were provided in 93 of 282 reports; were elevated in 69 of 93, and were normal in 24 of 93 reports. Proportions of hyperlipidemia and hypertriglyceridemia reports were significantly higher in the PWO group versus FA only group (p =.024 and.003, respectively) and versus FA with/without PW groups (p =.038 and.005, respectively). Weight gain was more frequently reported in those with FA (100%) or FA with PW (68%), whereas weight loss was usually reported in those with PWO (83%). In all, 98% of patients reporting FR on indinavir for whom a concomitant drug history was available were also taking lamivudine, stavudine, or both. A higher proportion of patients reporting PWO (34 of 60; 56.7%) versus FA (42 of 159; 26.4%) only took both lamivudine and stavudine. CONCLUSIONS: Differences observed from analysis of cases in clinical features, gender, weight change, concomitant medications, and presence of hyperlipidemia among the three groups of FR cases reported to Merck suggests that PWO may be a distinct entity from other features of FR. The data suggest that certain antiretroviral combinations predispose HIV persons to development of FR. PMID- 11103044 TI - Suppression of maternal virus load with zidovudine, didanosine, and indinavir combination therapy prevents mother-to-fetus HIV transmission in macaques. AB - Recently, we developed a maternal-fetal macaque model using a highly pathogenic HIV-2 strain, HIV-2287, to study the time course of HIV transmission in utero. Most pregnant macaques (Macaca nemestrina) infected with HIV-2287 (10-103 infective doses) transmitted HIV to their fetuses, as verified by positive identification of virus-infected mononuclear cells and free viral RNA in fetal blood. To determine whether an antiretroviral drug combination therapy composed of two dideoxynucleosides, azidothymidine (15 mg/kg) and dideoxyinosine (15 mg/kg), and a protease inhibitor, indinavir (25 mg/kg), could completely inhibit mother-to-fetus HIV transmission, we administered these drugs orally through gastric catheters to five pregnant macaques infected with 10 infective doses of HIV-2287. Beginning 30 minutes after HIV inoculation, the dams were given the combination antiviral therapy three times daily until delivery by cesarean section. Drug treatment reduced the maternal virus load to a minimally detectable level but did not prevent primary HIV-2287 infection. All fetal and infant blood samples were virus negative by internally controlled RNA polymerase chain reaction (QC-RNA-PCR) and virus coculture assays. Fetal and infant CD4+ T-cell levels remained normal throughout the experiment. These findings strongly suggest that combination chemotherapy with azidothymidine, dideoxyinosine, and indinavir can suppress maternal viral load enough to prevent mother-to-fetus transmission of HIV. PMID- 11103045 TI - Normalization of immune activation in lymphoid tissue following highly active antiretroviral therapy. AB - Although significant progress has been made in understanding immune reconstitution in peripheral blood following highly active antiretroviral therapy (HAART), less is known about immune changes in lymphoid tissue. Here, the expression of cytokine proteins (interferon gamma [IFN-gamma], interleukin [IL] 2, IL-4, IL-10, IL-1alpha, and IL-1beta) and surface antigens (CD4, CD8, CD1a, CD68) as well as cellular proviral HIV-1 DNA were determined in sequential tonsil biopsies before and at 4, 12, and 48 to 56 weeks posttherapy by quantitative in situ image analysis and fluorescent in situ 5;-nuclease assay (FISNA). Despite plasma virus suppression, a fraction of tonsil cells harbored pro-viral DNA for up to 1 year. A fourfold to eightfold increase in CD8+ T cells in tissue compared with seronegative controls and an increased frequency of CD1a+ dendritic cells prior to HAART reached control levels at week 56. The frequency of IFN-gamma expressing cells was 10-to 15-fold higher than controls before therapy and was comparable with findings in seronegative controls by week 56. Elevated baseline expression of IL-1alpha and IL-1beta was reduced by week 4 but IL-1alpha levels remained elevated in 1 of 3 patients at week 56. These findings suggest that with effective viral suppression the immune system in tissue may return to a more resting state. PMID- 11103046 TI - Prevalences of HTLV-1 infection and associated risk determinants in an urban population in Guinea-Bissau, West Africa. AB - OBJECTIVE: To assess the prevalence and modes of transmission of HTLV-1 infection in an adult population in Bissau, and to evaluate possible interactions between the pattern of spread of HTLV-1 and HIV-1/HIV-2. DESIGN AND METHODS: Univariate and multivariate analyses were used to evaluate gender-and age-specific HTLV-1 prevalences as well as associated risk determinants in an adult population based on a serosurvey comprising 2127 individuals from 304 randomly selected houses in Bissau. RESULTS: Using stringent Western blot criteria, the overall seroprevalence of HTLV-1 was 3.6%, 2.2% among men and 4.7% among women, respectively. One individual was seropositive to HTLV-2. The prevalence of HTLV 1, which increased with age in both genders, however more markedly among women, was >4 times higher (9.4%) among older individuals (>44 years of age) than among younger individuals (2.4%). Blood transfusion and HIV-2 seropositivity were independently associated with HTLV-1 seropositivity in men. Among women, both HIV 2 seropositivity and HIV-1 seropositivity were significant risk determinants. Having had sexual partners was associated with a fivefold increased risk among women but did not reach significance. CONCLUSION: The adult population of Guinea Bissau has a higher prevalence of HTLV-1 than reported from most other countries in West Africa. The gender-and age-specific pattern of spread of HTLV-1 closely resembles that observed for HIV-2, another retrovirus prevalent to the region. The close correlation between HTLV-1 and HIV-2 most likely reflects the shared risk factors related to sexual behavior. The implication of the high percentage of double infections in this population needs to be determined. PMID- 11103047 TI - Economic evaluation of HIV risk reduction intervention in African-American male adolescents. AB - PURPOSE: To evaluate the cost-effectiveness of a cognitive-behavioral HIV risk reduction intervention for African-American male adolescents that has previously been shown to be effective at reducing sexual risk taking. METHODS: Standard techniques of cost-utility analysis were employed. A societal perspective and a 3% discount rate were used in the main analysis. Program costs were ascertained retrospectively. A mathematical model of HIV transmission was used to translate observed changes in sexual behavior into an estimate of the number of HIV infections the intervention averted. Intervention effects were assumed to last for 1 year. For each infection averted, the corresponding savings in future HIV related medical care costs and quality-adjusted life years (QALYs) were estimated. The overall net cost per QALY saved (cost-utility ratio) was then calculated. Sensitivity analyses were performed to assess the robustness of the main results. RESULTS: The cost-utility ratio was approximately $57,000 U.S. per QALY saved when training costs were included, and $41,000 U.S. per QALY saved when they were excluded. The intervention appeared substantially more cost effective when the analysis was restricted to the subgroup of participants who reported being sexually active at baseline. Assumptions about the prevalence of HIV infection and the duration of intervention effectiveness also greatly affected the cost-utility ratio. CONCLUSIONS: The HIV prevention intervention was moderately cost-effective in comparison with other health care programs. Selectively implementing the intervention in high-HIV prevalence communities and with sexually active youth can enhance cost-effectiveness. PMID- 11103048 TI - Partner concordance in reports of joint risk behaviors. AB - OBJECTIVE: To evaluate the accuracy of self reports on sexual and drug use behaviors. METHODS: Data from a network study of HIV transmission among a sample of drug users and nonusers are used to compare reports of sexual and drug use behaviors by partners who engaged in those behaviors. Partner concordance (self report agreement between two people) was used as an estimate of validity. RESULTS: Results showed that persons are able to recall and report about 85% of their recent partners (15%-20% less for recent drug use partners). For relationships that were reported by both partners, a high degree of concordance existed about recent behaviors (83%-96%) and variable agreement about frequency (0.48 /=3 days (27% versus 7%, respectively; p <.001), and weight loss of >/=2 kg (21% versus 9%, respectively; p =.05). STDs were more common in seroconverters (gonorrhea: 9% versus 1%, respectively; p <.01 and condyloma: 9% versus 3%, respectively; p =. 08). The first case definition was ID for >3 days, fever, pharyngitis, and myalgia (seroconverters, 3 of 32, versus nonseroconverters, 2 of 640). The second case definition was was ID for >3 days, anti-core hepatitis b-positive, and age <21 years (seroconverters: 6 of 32 versus nonseroconverters 4 of 640). The sensitivity and specificity for the first and second case definitions were: 9.4%, 99.4%, and 18.8%, 99.8%, respectively. CONCLUSIONS: Among HIV seroconverters, symptoms consistent with ARS were common. We were unable to identify a sensitive case definition that could be used as a screening tool. Although the clinical case definition was not validated, the specificity of our case definitions was high, suggesting that subjects within this HIV risk group who fulfill the case definition should be tested for HIV. PMID- 11103051 TI - Association between crack cocaine use and high-risk sexual behaviors after HIV diagnosis. AB - OBJECTIVE: To describe the prevalence of crack cocaine use in an HIV-infected population and to examine the association between crack use after HIV diagnosis and high-risk sexual behaviors for heterosexual men, heterosexual women, and men who have sex with men (MSM). METHODS: Analysis of cross-sectional interviews conducted from January 1995 through December 1998 with HIV infected adults in 12 states. RESULTS: Of 10,415 persons with HIV or AIDS, 66.6% never used crack, 10.7% used crack before HIV diagnosis but not after, and 22.7% used crack after diagnosis. High-risk sexual behaviors were more prevalent among those who had ever used crack and were most prevalent among those who used crack after diagnosis. In multivariable analyses, crack use after diagnosis was associated with having multiple sex partners and trading sex for drugs/money in all three groups: heterosexual men, heterosexual women, and MSM. For heterosexual women and MSM, crack use after diagnosis was associated with unprotected sex with a main partner, and among heterosexual men and MSM, with unprotected sex with casual partners. CONCLUSIONS: Crack use after HIV diagnosis was associated with high risk sexual behaviors. Treatment programs to assist people in quitting crack are needed to help reduce the risk of HIV transmission from this population. PMID- 11103052 TI - Increase of atherogenic plasma profile in HIV-infected patients treated with protease inhibitor-containing regimens. PMID- 11103053 TI - Directly observed therapy in HIV therapy: A realistic perspective? PMID- 11103054 TI - Erythropoietin with iron supplementation to prevent allogeneic blood transfusion in total hip joint arthroplasty. A randomized, controlled trial. AB - BACKGROUND: The optimum regimen of epoetin alfa for prevention of allogeneic blood transfusion is unknown. OBJECTIVE: To determine whether a modified regimen of epoetin alfa reduces allogeneic blood transfusion in patients undergoing hip arthroplasty. DESIGN: Randomized, double-blind, multicenter trial comparing two modified dose regimens of epoetin alfa with placebo. SETTING: 13 teaching hospitals and 4 community hospitals in Canada. PATIENTS: 201 patients undergoing primary hip arthroplasty who had a hemoglobin concentration of 98 to 137 g/L and did not predonate blood. INTERVENTION: Patients were assigned in a 3:5:5 ratio to receive four weekly doses of epoetin alfa, 40 000 U (high-dose; n = 44) or 20 000 U (low-dose; n = 79), or placebo (n = 78), starting 4 weeks before surgery. All patients received oral iron supplementation, 450 mg/d, for 42 or more days before surgery. MEASUREMENTS: The primary end point was allogeneic transfusion. Secondary end points were thromboembolic events and change in reticulocyte count and hemoglobin concentration. RESULTS: Both modified epoetin alfa regimens significantly reduced the need for allogeneic transfusion: Five (11.4%) patients in the high-dose group (P = 0.001) and 18 (22. 8%) patients in the low-dose group (P = 0.003) had transfusion, compared with 35 (44.9%) patients in the placebo group. The hematologic response was substantial in patients who received epoetin alfa. In the high-dose group, low-dose group, and placebo group, the preoperative increase in reticulocyte count was 58.8, 37. 0 and 1.8 x 10(9) cells/L (P < 0.001), respectively, and the increase in hemoglobin concentration was 19.5, 17.2, and 1.2 g/L (P < 0.001). The incidence of thromboembolic events did not differ among groups. CONCLUSIONS: Both modified epoetin alfa regimens were effective compared with placebo in reducing allogeneic transfusion in patients undergoing hip arthroplasty. Patients who received high-dose epoetin alfa had the lowest transfusion rate. PMID- 11103055 TI - Performance of screening mammography among women with and without a first-degree relative with breast cancer. AB - BACKGROUND: Although it is recommended that women with a family history of breast cancer begin screening mammography at a younger age than average-risk women, few studies have evaluated the performance of mammography in this group. OBJECTIVE: To compare the performance of screening mammography in women with a first-degree family history of breast cancer and women of similar age without such history. DESIGN: Cross-sectional. SETTING: Mammography registries in California (n = 1), New Hampshire (n = 1), New Mexico (n = 1), Vermont (n = 1), Washington State n = 2), and Colorado (n = 1). PARTICIPANTS: 389 533 women 30 to 69 years of age who were referred for screening mammography from April 1985 to November 1997. MEASUREMENTS: Risk factors for breast cancer; results of first screening examination captured for a woman by a registry; and any invasive cancer or ductal carcinoma in situ identified by linkage to a pathology database, the Surveillance, Epidemiology, and End Results program, or a state tumor registry. RESULTS: The number of cancer cases per 1000 examinations increased with age and was higher in women with a family history of breast cancer than in those without (3.2 vs. 1.6 for ages 30 to 39 years, 4.7 vs. 2.7 for ages 40 to 49 years, 6.6 vs. 4.6 for ages 50 to 59 years, and 9.3 vs. 6.9 for ages 60 to 69 years). The sensitivity of mammography increased significantly with age (P = 0.001 [chi square test for trend]) in women with a family history and in those without (63.2% [95% CI, 41. 5% to 84.8%] vs. 69.5% [CI, 57.7% to 81.2%] for ages 30 to 39 years, 70.2% [CI, 61.0% to 79.5%] vs. 77.5% [CI, 73.3% to 81.8%] for ages 40 to 49 years, 81.3% [CI, 73.3% to 89.3%] vs. 80.2% [CI, 76.5% to 83.9%] for ages 50 to 59 years, and 83.8% [CI, 76.8% to 90.9%] vs. 87.7% [CI, 84.8% to 90.7%] for ages 60 to 69 years). Sensitivity was similar for each decade of age regardless of family history. The positive predictive value of mammography was higher in women with a family history than in those without (3.7% vs. 2.9%; P = 0.001). CONCLUSIONS: Cancer detection rates in women who had a first-degree relative with a history of breast cancer were similar to those in women a decade older without such a history. The sensitivity of screening mammography was influenced primarily by age. PMID- 11103056 TI - Cost-effectiveness of radiofrequency ablation for supraventricular tachycardia. AB - BACKGROUND: Radiofrequency ablation is an established but expensive treatment option for many forms of supraventricular tachycardia. Most cases of supraventricular tachycardia are not life-threatening; the goal of therapy is therefore to improve the patient's quality of life. OBJECTIVE: To compare the cost-effectiveness of radiofrequency ablation with that of medical management of supraventricular tachycardia. DESIGN: Markov model. DATA SOURCES: Costs were estimated from a major academic hospital and the literature, and treatment efficacy was estimated from reports from clinical studies at major medical centers. Probabilities of clinical outcomes were estimated from the literature. To account for the effect of radiofrequency ablation on quality of life, assessments by patients who had undergone the procedure were used. TARGET POPULATION: Cohort of symptomatic patients who experienced 4.6 unscheduled visits per year to an emergency department or a physician's office while receiving long term drug therapy for supraventricular tachycardia. TIME HORIZON: Patient lifetime. PERSPECTIVE: Societal. INTERVENTIONS: Initial radiofrequency ablation, long-term antiarrhythmic drug therapy, and treatment of acute episodes of arrhythmia with antiarrhythmic drugs. OUTCOME MEASURES: Costs, quality-adjusted life-years, life-years, and marginal cost-effectiveness ratios. RESULTS OF BASE CASE ANALYSIS: Among patients who have monthly episodes of supraventricular tachycardia, radiofrequency ablation was the most effective and least expensive therapy and therefore dominated the drug therapy options. Radiofrequency ablation improved quality-adjusted life expectancy by 3.10 quality-adjusted life-years and reduced lifetime medical expenditures by $27 900 compared with long-term drug therapy. Long-term drug therapy was more effective and had lower costs than episodic drug therapy. RESULTS OF SENSITIVITY ANALYSIS: The findings were highly robust over substantial variations in assumptions about the efficacy and complication rate of radiofrequency ablation, including analyses in which the complication rate was tripled and efficacy was decreased substantially. CONCLUSIONS: Radiofrequency ablation substantially improves quality of life and reduces costs when it is used to treat highly symptomatic patients. Although the benefit of radiofrequency ablation has not been studied in less symptomatic patients, a small improvement in quality of life is sufficient to give preference to radiofrequency ablation over drug therapy. PMID- 11103057 TI - Fatal hepatorenal failure associated with hydrazine sulfate. AB - BACKGROUND: The Internet has revolutionized the manner in which patients obtain information about health care. This technology has also allowed patients to obtain directly both prescription and nonprescription therapies. OBJECTIVE: To report a case of fulminant hepatorenal failure associated with the use of hydrazine sulfate, an unregulated alternative remedy for cancer marketed on the Internet. DESIGN: Case report. SETTING: Academic medical center. PATIENT: A 55 year-old man with maxillary sinus cancer. INTERVENTION: Self-medication with hydrazine sulfate. MEASUREMENTS: Serum liver and renal function tests; histologic evaluation of liver and kidney tissue. RESULTS: The patient developed hepatic encephalopathy, renal failure, and profound coagulopathy. He died after severe gastrointestinal hemorrhage developed. Autopsy revealed autolysis of the kidneys and submassive bridging necrosis of the liver. CONCLUSION: Fatal hepatorenal failure may occur after the use of hydrazine sulfate. This fatal complication must be considered in anyone taking or contemplating the use of hydrazine sulfate. PMID- 11103058 TI - Prolongation of the QT interval and ventricular tachycardia in patients treated with arsenic trioxide for acute promyelocytic leukemia. AB - BACKGROUND: Recently, arsenic trioxide has increasingly been used for relapsed acute promyelocytic leukemia. However, it is known to have several adverse effects, including acute cardiac toxicities. OBJECTIVE: To determine cardiac toxicities resulting from arsenic trioxide therapy in patients with relapsed or refractory acute promyelocytic leukemia. DESIGN: Phase II clinical prospective cohort study. SETTING: A university hospital in Hamamatsu, Japan. PATIENTS: 8 patients with relapsed acute promyelocytic leukemia. INTERVENTION: Arsenic trioxide, 0.15 mg/kg of body weight, administered daily by 2-hour infusion for a maximum of 60 days. MEASUREMENTS: Continuous monitoring with ambulatory electrocardiography. RESULTS: Five patients (63%) achieved complete remission. During induction therapy with arsenic trioxide, prolonged QT intervals were observed in all patients. Ventricular premature contractions were noticed during 8 of 12 courses of therapy. Four patients developed nonsustained ventricular tachycardia and required treatment with antiarrhythmic agents. CONCLUSIONS: Cardiac toxicity occurs during arsenic trioxide therapy in patients with acute promyelocytic leukemia. Such patients should be monitored for prolonged QT intervals and ventricular arrhythmia. PMID- 11103059 TI - Dying patients in the intensive care unit: forgoing treatment, maintaining care. AB - End-of-life care of patients in the intensive care unit (ICU) often requires dramatic shifts in attitudes and interventions, from traditional intensive rescue care to intensive palliative care. The care of patients dying in ICUs raises both clinical and ethical difficulties. Because fewer ICU patients are able to make decisions about withdrawing treatment, careful attention must be paid to previously expressed preferences and surrogate input. Cultural and spiritual values of patients and families may differ markedly from those of clinicians. Although prognostic models are increasingly able to predict mortality rates for groups of ICU patients, their usefulness in guiding specific decisions to forego treatment has not been established. When a decision to forego treatment is made, the focus should be on specifying the patient's goals of care and assessing all treatments in light of these goals; interventions that do not contribute to the patient's goals should be discontinued. Symptoms accompanying withdrawal of life support can almost always be controlled with appropriate palliative measures. After ICU interventions are foregone, patient comfort must be the paramount objective. Whether in the ICU or elsewhere, hospitals have an ethical obligation to provide settings that offer dignified, compassionate, and skilled care. PMID- 11103060 TI - Update in geriatrics. PMID- 11103061 TI - The role of the implantable cardioverter-defibrillator for prevention of sudden cardiac death. AB - Sudden cardiac death, which accounts for approximately 350,000 deaths each year, is a major health care problem. Antiarrhythmic drugs have not been reliable in preventing sudden cardiac death. Although beta-blockers, angiotensin-converting enzyme inhibitors, and revascularization play a role in prevention of sudden cardiac death, the development and subsequent refinement of the implantable cardioverter-defibrillator has made the most important contribution to its management. Several randomized, controlled trials have demonstrated improved survival in patients resuscitated from cardiac arrest. Two recent trials also suggest a role for primary prevention in selected patients with coronary artery disease, ventricular dysfunction, and nonsustained ventricular tachycardia in whom sustained ventricular tachycardia is induced. Further technological refinements and development of new, more sensitive risk stratifiers with a higher positive predictive value for sudden cardiac death will expand the indications for this life-saving therapy. PMID- 11103062 TI - Hydrazine, cancer, the Internet, isoniazid, and the liver. PMID- 11103064 TI - Knees. PMID- 11103063 TI - Anatomy Lesson. PMID- 11103065 TI - The rebirth of Isabelle. PMID- 11103066 TI - Measuring the quality of breast cancer care. PMID- 11103067 TI - Medicine and spirituality. PMID- 11103068 TI - Cholesterol and lipoprotein levels as predictors of response to interferon for hepatitis C. PMID- 11103069 TI - Mycophenolate-associated onycholysis. PMID- 11103070 TI - Recovery from theophylline toxicity by continuous hemodialysis with filtration. PMID- 11103071 TI - Hepatitis C virus genotype 4 and response to combination therapy with interferon alpha2b plus ribavirin. PMID- 11103072 TI - Cocaine-related vasculitis causing upper-limb peripheral vascular disease. PMID- 11103073 TI - The deletion allele of angiotensin-converting enzyme gene polymorphism as a cardiovascular risk factor in patients undergoing long-term hemodialysis. PMID- 11103074 TI - Organism responsible for nodular cutaneous microsporidiosis in a patient with AIDS. PMID- 11103075 TI - Acute leukemia in a patient with sickle-cell anemia treated with hydroxyurea. PMID- 11103076 TI - Internuclear ophthalmoplegia after insecticide exposure. PMID- 11103077 TI - "Show your Wound" medicine and the work of Joseph Beuys. PMID- 11103080 TI - Improvement of Schwann cell attachment and proliferation on modified hyaluronic acid strands by polylysine. AB - Hyaluronic acid (HyA) has the intrinsic ability to promote cell proliferation and reduce scar formation. However, the clinical use of HyA has so far been limited because of its water solubility and nonadhesive characteristics. Increasing interest in HyA as a clinically useful biomaterial has prompted our study of altering HyA's physical properties to render it a potential component of nerve grafts. In this study, strands of HyA were cross-linked by glutaraldehyde (Glut), coated with polylysine, and then inoculated with Schwann cells (SCs). Results in vivo and in vitro demonstrated that cross-linked HyA strands were water insoluble and thus less biodegradable. Poly-D-lysine-resurfaced strands showed significant SC attachment of 350-400 cells/mm(2), compared to uncoated controls (0-10 cells/mm(2), p < 0.01). Fibroblast control groups showed an attachment of 40-100 cells/mm(2) on coated strands. Immunostaining for proliferating cells showed SCs as and fibroblasts as +. Cells neither adhered to nor proliferated on the modified HyA strands that were not resurfaced. The results suggest that polylysine promotes SC attachment and proliferation to glutaraldehyde-cross linked HyA strands, the product being a three-dimensional composite with low solubility that may have potential application in nerve grafts. PMID- 11103081 TI - Formation of new tissue from an arteriovenous loop in the absence of added extracellular matrix. AB - A major requirement for the microsurgical repair of contour defects of the skin, for example, following removal of a skin cancer on the face, is a mass of vascularised subcutaneous tissue. Such tissue can be generated in vivo using basic tissue engineering principles. In previous studies in our laboratory, we have used a model comprising an arteriovenous (AV) shunt loop sandwiched in artificial dermis, placed in a cylindrical plastic growth chamber, and inserted subcutaneously to grow new connective tissue progressively up to 4 weeks. To learn more about the basic growth characteristics with this model, the same AV shunt loop within a chamber without added extracellular matrix was inserted subcutaneously into the groins of rats for 2, 4, or 12 weeks (n = 5 per group). There was a progressive increase in the mass and volume of tissue such that the chamber was two-thirds full after 12 weeks. Histological examination showed that at 2 weeks there was evidence of fibroblast and vascular outgrowth from the AV shunt, with the formation of granulation tissue, surrounded by a mass of coagulated exudate. At 4 weeks the connective tissue deposition was more extensive, with a mass of more mature granulation tissue containing considerable collagen. By 12 weeks there was an extensive, well vascularized mass of mature fibrous tissue. The blood vessels and residual adventitia of the AV shunt were the likely source of growth factors and of the cells which populated the chamber with new maturing connective tissue. A patent AV shunt in an isolated chamber appears to be the minimal requirement for the generation of new vascularized tissue that is potentially suitable for microsurgical transplantation. PMID- 11103082 TI - Engineered bone development from a pre-osteoblast cell line on three-dimensional scaffolds. AB - Bone regeneration is based on the hypothesis that healthy progenitor cells, either recruited or delivered to an injured site, can ultimately regenerate lost or damaged tissue. Three-dimensional porous polymer scaffolds may enhance bone regeneration by creating and maintaining a space that facilitates progenitor cell migration, proliferation, and differentiation. As an initial step to test this possibility, osteogenic cells were cultured on scaffolds fabricated from biodegradable polymers, and bone development on these scaffolds was evaluated. Porous polymer scaffolds were fabricated from biodegradable polymers of lactide and glycolide. MC3T3-E1 cells were statically seeded onto the polymer scaffolds and cultured in vitro in the presence of ascorbic acid and beta-glycerol phosphate. The cells proliferated during the first 4 weeks in culture and formed a space-filling tissue. Collagen messenger RNA levels remained high in these cells throughout the time in culture, which is consistent with an observed increase in collagen deposition on the polymer scaffold. Mineralization of the deposited collagen was initially observed at 4 weeks and subsequently increased. The onset of mineralization corresponded to increased mRNA levels for two osteoblast-specific genes: osteocalcin and bone sialoprotein. Culture of cell/polymer constructs for 12 weeks led to formation of a three-dimensional tissue with architecture similar to that of native bone. These studies demonstrate that osteoblasts within a three-dimensional engineered tissue follow the classic differentiation pathway described for two-dimensional culture. Polymer scaffolds such as these may ultimately be used clinically to enhance bone regeneration by delivering or recruiting progenitor cells to the wound site. PMID- 11103083 TI - Priming of hepatocytes for cell culture by partial hepatectomy prior to cell isolation. AB - The combination of ex vivo gene transfer and a sufficient transplant model for hepatocytes may permit treatment of single enzyme-based metabolic liver diseases. Induction of replicative potential (priming) in hepatocyte cultures may enhance the efficiency of gene transfer under stable in vitro conditions. It is known that hepatocyte replication is increased in vivo after partial hepatectomy. We investigated the effect of partial hepatectomy prior to cell isolation on hepatocytes in vitro. Male Lewis rats served as donors. Hepatocytes were isolated by collagenase digestion from either intact livers or from livers 48 h after 70% hepatectomy (PH). Cells were seeded on collagen-coated culture dishes with hormone-supplemented culture media. Hepatocyte morphology, number, albumin secretion rate, and mono-ethyl-glycin-xylidid (MEGX)-biotransformation capacity were assessed on days 1, 3, and 5 in culture. PH significantly increased hepatocyte number and albumin secretion of cultured hepatocytes over the whole observation period. In contrast, MEGX-biotransformation capacity was significantly decreased. Morphology of cultured hepatocytes was not affected by PH prior to hepatocyte isolation. These results suggest a prolonged and complex response of hepatocytes to PH in vitro. Hepatocyte priming by PH is a promising approach toward stable cultures of proliferating hepatocytes and may provide a model for in vitro studies of hepatic regeneration mechanisms. Further research on hepatocyte priming toward an application in ex vivo gene transfer and hepatic tissue engineering seems justified. PMID- 11103084 TI - Transmission electron microscopic study of hepatocytes in bioartificial liver. AB - A bioartificial liver (BAL) was prepared by simple inoculation of hepatocytes into the inner space of hollow fibers of a hemodialyzer and it was maintained in a closed circuit for in vitro culture. Morphology of hepatocytes in the hollow fibers was studied in detail using transmission electron microscopy (TEM). The hepatocytes formed three-dimensional, rod-shaped aggregates of 200 microm in diameter throughout the whole dimension of the hollow fibers after 1 day of culture. Approximately five hepatocyte layers existed from the surface to the center of the aggregate. The hepatocytes in the aggregate displayed mostly polygonal shapes and were surrounded by five to six cells. Abundant bile canaliculi were formed between the hepatocytes and were sealed by tight junctions. The distance between the adjacent hepatocytes except the bile canaliculus domain was approximately 20 nm, and interdigitation was observed between some hepatocytes. These observations indicate that the hepatocytes formed functionally associated aggregates, that is, organoids. Although the cells facing the inner surface of the hollow fiber lost their polygonal shape and became flattened during the following several-day culture, no drastic change was observed in the morphology of the hepatocytes located inside the aggregate. After 14 days of culture, the number of living cells decreased and most of these had a deformed nucleus, few numbers of organelles, and intermittent lipid droplets. PMID- 11103085 TI - Concept for organ engineering: a reconstruction method of rat liver for in vitro culture. AB - In the past decade, there have been remarkable advances in tissue engineering technology toward the goal of creating organoids in vitro from cells and cellular scaffolding. Indeed, tissue-engineered organoids such as skin and cartilage, each with comparatively simple architectures, are presently at the clinical stage. However, conventional tissue engineering techniques have not allowed for the reconstruction of an organoid that mimics an organ of complex architecture of abundant vascular networks. We established a method for organ engineering that can remodel a rat liver into a reconstructed organoid without separating the majority of liver cells by a continuous three-step perfusion. The liver was perfused through its vascular system with a buffered balanced salt solution to cleanse blood from the organ, with a collagenase/dispase medium to deconstruct cellular scaffolds, and with a culture medium containing collagen type I to reorganize the multicellular architecture. The reconstructed organoid was then prepared by excising the perfused liver from the rat and culturing it at 37 degrees C for 2 h. Histologically healthy parenchymal hepatocytes expressing albumin were observed in the excised organoid even after culture for 3 weeks. Furthermore, a fibroblast-implanted organoid was prepared by using a culture medium containing suspended fibroblasts in the third step of the perfusion procedure, demonstrating the efficacy of heterogeneous cells for the reconstruction of an organoid. This method may be applicable to the formation of organoids from other organs, such as kidney and spleen, each of which have abundant capillaries, and therefore the method provides a novel concept for the development of lab-grown organs, i. e., organ engineering. PMID- 11103087 TI - Poster presentations PMID- 11103086 TI - Oral presentations PMID- 11103089 TI - Bulletin board PMID- 11103088 TI - Biological sex analysis in clinical research. PMID- 11103090 TI - Hormones, chromosomes, and the future of sex-based biology. PMID- 11103091 TI - The role of the environmental health laboratory in women's health. PMID- 11103092 TI - Toward optimal health: the experts discuss colon cancer. PMID- 11103093 TI - Helping women find everyday solutions. PMID- 11103094 TI - The first annual conference on sex and gene expression. PMID- 11103095 TI - Health issues for women with epilepsy: a descriptive survey to assess knowledge and awareness among healthcare providers. AB - The American Academy of Neurology and the American College of Obstetricians and Gynecologists recently issued practice parameters for women with epilepsy. These parameters suggest optimal care practices. To assess knowledge of the issues covered in the parameters and to facilitate educational efforts to promote best care, the Epilepsy Foundation conducted a survey of healthcare professionals likely to provide care to women with epilepsy. The survey sampled 3535 healthcare professionals across a wide range of specialties. Most respondents did not know the specific effects of estrogen and progesterone on the seizure threshold, were not aware of menstrual-associated seizure patterns, and could not identify which antiepileptic drugs interfere with oral contraceptives. The majority of respondents did not know that women with epilepsy have higher rates of infertility, reproductive endocrine disorders, and sexual dysfunction. Most respondents did not know the frequency of birth defects in children born to women with epilepsy. Providers seeing the largest number of persons with epilepsy were more likely to have correct answers. By specialty, neurologists provided the highest number of correct responses, followed (in descending order) by endocrinologists, obstetricians/gynecologists, internal medicine physicians, family practice physicians, and pediatricians. These results suggest that women with epilepsy are not receiving adequate counseling and that care practices may not conform to those recommended. PMID- 11103096 TI - Social and contextual etiology of coronary heart disease in women. AB - We explored the social and contextual etiology of coronary heart disease (CHD) prevention and management in women. Social and contextual influences on CHD risk include such factors as socioeconomic status, access to healthcare, cultural mores, working conditions including work overload, multiple role responsibilities, and social isolation. Women, particularly economically disadvantaged women, occupy lower levels on the social status hierarchy and, therefore, experience more stressful life experiences, less favorable living conditions, and less opportunity to affect positive health behavior and outcomes. Women are often discriminated against economically, politically, and socially, and this discrimination may adversely affect their efforts at CHD health promotion and treatment. Multiple role responsibilities within the family and psychosocial factors, including chronic life stress, are critical to an understanding of the health status of women, particularly poor and minority women. Although community-based interventions appear to be ideal for addressing the contextual risks related to CHD in women, a number of issues need to be considered, for example, the limited acknowledgment of secular trends in economic development that influence lifestyle decisions and health promotion efforts. Directions for research and interventions include recognition of the full spectrum of CHD risk in women, recognition of culturally competent interventions, and recognition of the need for empowerment of women. PMID- 11103097 TI - Implementing a new model of integrated women's health in academic health centers: lessons learned from the National Centers of Excellence in Women's Health. AB - The National Centers of Excellence in Women's Health Program (CoE) represents a new model for women's health in academic health centers that unites women's health research, teaching, clinical care, public education and outreach, and career advancement for women in the health sciences. Lessons learned from the first 3 years of implementation indicate that this type of model requires a transformation from the traditionally fragmented set of activities in academic health centers to an integrated system united around the goal of advancing women's health. The transformation requires institutional commitment, dedicated players, and an ability to build on existing resources and bring added value to the institution. Challenges and strategies to link women's health activities and increase collaboration are also discussed. PMID- 11103098 TI - Carotid endarterectomy for women and men. AB - Carotid endarterectomy is the standard of care for people with severe symptomatic carotid stenosis. We analyzed population administrative data and clinical trial data to determine whether sex differences exist in the use and outcomes of this surgical procedure. We studied patients in Ontario who underwent carotid endarterectomy between 1982 and 1994 (n = 12,949) and patients with severe carotid stenosis who were enrolled in two randomized trials of endarterectomy (n = 1646). We compared the proportion of men and women who underwent carotid endarterectomy in each group, over time, and after adjustment for demographic factors. Men were twice as likely as women to receive carotid endarterectomy in the administrative analysis (65% versus 35%, p < 0.001) and in the clinical trial analysis (70% versus 30%, p < 0.001). The relatively lower use in women was consistent in every age group and in every year studied. Men in the administrative database were somewhat less likely than women to die or be institutionalized after surgery (5% versus 6%, p = 0.007). Men in the clinical trial database were also less likely than women to experience perioperative stroke or death, although the results were not statistically significant (6% versus 7%, p = 0.32). Patients who were assigned to surgical therapy, compared with those assigned to medical therapy, had a significant decrease in the risk of adverse events at 1 year, and the net benefit appeared similar in women and men. Carotid endarterectomy is performed relatively infrequently on women despite their similar lifetime burden of disease and similar short-term perioperative risks compared with men. PMID- 11103099 TI - The effect of an ipriflavone-containing supplement on urinary N-linked telopeptide levels in postmenopausal women. AB - Osteoporosis is a significant health concern to our aging population. We report here the results of a pilot placebo-controlled trial of a dietary supplement containing ipriflavone, calcium, and vitamin D on a urinary marker of bone breakdown in postmenopausal women. Seven postmenopausal women not currently receiving hormone replacement therapy received either an ipriflavone-containing supplement or placebo for 3 months. Urinary N-linked telopeptides, a marker of bone breakdown, declined by 29% in those receiving the supplement, whereas an increase in this marker was observed in the group receiving the placebo. No changes were observed in salivary hormone measurements. Although our sample size was small, to the best of our knowledge, this is the first report that demonstrates changes in N-linked telopeptide levels as a result of consuming an ipriflavone-containing product. Our findings confirm those of other researchers that demonstrate the usefulness of ipriflavone at slowing the progression of bone loss and suggest that measuring N-linked telopeptides may be a useful tool to assess therapeutic efficacy. PMID- 11103100 TI - How does breast cancer mortality compare with that of other cancers and selected cardiovascular diseases at different ages in U.S. women? AB - Women tend to fear breast cancer and thus overestimate their risk of developing it, have less concern about developing heart disease, and do not know that lung cancer is the major cause of cancer death. Death certificate data, consolidated into a national database by the National Center for Health Statistics, were used to compare age-specific mortality due to selected cardiovascular diseases and cancers among women who died in 1997 in the United States. The outcomes examined included underlying cause of death categorized as all circulatory system disease, cerebrovascular disease, and heart disease, including coronary and noncoronary disease, and as all cancers combined plus cancer of the lung, breast, and colon/rectum. In 1997, 500,703 women in the United States died from diseases of the circulatory system, including 370,357 deaths from heart disease. Most deaths from heart disease were due to coronary heart disease, which exceeded mortality from cerebrovascular disease at all ages except under age 40. In 1997, 258,463 women in the United States died from cancer, and before age 55, breast cancer death rates exceeded lung and colorectal cancer death rates. Mortality due to total heart disease exceeded breast and lung cancer mortality among women at all ages, but before age 55, when absolute death rates are low, breast cancer death rates exceeded those for coronary heart disease. In conclusion, aside from mortality due to all cancers combined and circulatory system disease, only accidents, which were not included in this study, and total heart disease caused more deaths than breast cancer before age 55. PMID- 11103101 TI - A profile of early versus late onset of obesity in postmenopausal women. AB - Obesity is a serious health problem among women across the life span. Although people can become obese at any age, there is a large proportion of older women who have been obese since childhood. The purpose of this study was to determine whether postmenopausal women with an early versus late onset of obesity manifested differences in body habitus, eating behaviors, and mood. One hundred thirty-five postmenopausal women with obesity responded to self-report questionnaires on weight history, weight loss and maintenance expectancy, eating behaviors, and mood. Women with an early onset of obesity had a significantly higher body mass index (BMI), waist circumference, and highest attained adult body weight than women with a late onset of obesity. They had attempted a significantly larger number of diets and had lost more weight on any single diet. The groups also differed significantly on binge eating and overeating in response to negative affect. There was a tendency for women with an early onset to have more depressive and anxious symptoms. Postmenopausal women with an early onset of obesity differed physiologically and psychologically from those with a late onset. Tailoring dietary and behavioral interventions to profiles of postmenopausal women based on onset of obesity may improve the overall efficacy of weight loss programs. PMID- 11103102 TI - Intimate partner violence and cervical neoplasia. AB - Intimate partner violence (IPV) is associated with a range of adverse physical health outcomes, including chronic and infectious diseases. An emerging literature suggests that partner violence and specifically sexual violence may be associated with an increased risk of cervical neoplasia. To assess the risk of preinvasive and invasive cervical cancer in a cross-sectional study of women screened for IPV by type, frequency and duration, 1152 women ages 18-65 were recruited from family practice clinics in 1997-1998. They were screened for IPV during a brief in-clinic interview, and health history and current status were assessed in a follow-up interview. Of 1152 women surveyed, 14 (1.2%) reported cervical cancer, and 20. 3% (n = 234) reported treatment for cervical neoplasia. Ever experiencing IPV was associated with an increased risk of invasive cervical cancer (adjusted relative risk [aRR] = 4.28; 95% CI 1.94, 18.39) and with preinvasive cervical neoplasia (aRR = 1.47; 95% CI 1. 16, 1.82). This association was stronger for women experiencing physical or sexual IPV than for women experiencing psychological IPV. Women with cervical cancer reported being in violent relationships longer and experiencing more frequent physical and sexual assaults and more IPV-associated injuries than did controls. This exploratory study suggests that IPV may increase a woman's risk of cervical neoplasia. The mechanism by which IPV effects cervical neoplasia may be indirect through psychosocial stress or negative coping behaviors or direct through sexual assaults and transmission of human papillomavirus (HPV). PMID- 11103103 TI - Differences in breast cancer screening rates: an issue of ethnicity or socioeconomics? AB - Previous reports suggest that use of preventive measures, such as screening mammography (SM), differs by ethnicity. It is unclear, however, if this is determined directly by ethnicity or indirectly by related socioeconomic factors. We studied self-reported data from 18,245 women aged 40-49 who participated in the Behavioral Risk Factor Surveillance System telephone survey in 1992 and 1993. Of these, 11,509 (63%) reported having obtained mammography within the preceding 2 years for screening purposes only. Using reports of other preventive healthcare behaviors, education level, socioeconomic status, and healthcare access problems as independent variables, bivariate associations were assessed, and a logistic regression model was developed. Models for each ethnic group were developed, with consistent results. Women who engaged in other preventive health measures, such as Pap smear (odds ratio [OR] 8.99, 95% confidence interval [CI] = 7.6-10.7), cholesterol measurement (OR 2.64, 95% CI = 2.3-3.0), and seatbelt use, were more likely to obtain SM. Women with healthcare access or insurance problems (OR 0. 59, 95% CI = 0.5-0.7) and current smokers (OR 0.71, CI = 0.6-0.8) had a lower likelihood of obtaining SM. Ethnicity, alcohol use, marital status, and education level were not significantly associated with women's reports of SM. Although ethnicity apparently does not influence a woman's likelihood of obtaining SM, access to healthcare and insurance and engaging in other healthy behaviors do. Health policy planners should consider the importance of these related factors when developing preventive health programs for women. PMID- 11103104 TI - Webwatch-women's health and gender-based medicine. Sexually transmitted diseases. PMID- 11103106 TI - Correction PMID- 11103105 TI - Women's health literaturewatch. PMID- 11103107 TI - [Liver cell culture in bioreactors for in vitro drug studies as an alternative to animal testing]. AB - An important consideration for the utilisation of in vitro culture models for studies on drug metabolism as an alternative to animal testing is the maintenance of a defined degree of cell differentiation. Thus, in vitro conditions reflecting as near as possible the in vivo situation of the cells within the whole organ are required. A bioreactor was developed for the cultivation of liver cells which allows the reorganisation of hepatocytes and non-parenchymal cells of the liver in coculture to form three-dimensional, tissue-like structures including extracellular matrix components produced by the cells. In this study, the vitality and metabolic activity of isolated rat hepatocytes was investigated over a two week culture period in bioreactors. The results show that after a reorganisation phase, the cells preserve specific functions, such as protein and urea synthesis capacity and specific cytochrome P450 activities during the culture period, with maximal values during the first week. Possible applications of the model in pharmaceutical industry are studies on metabolite patterns, enzyme induction, drug-drug-interactions, first pass effects and long-term toxicity of drugs. PMID- 11103108 TI - [In vitro model to investigate the effect of estrogens on neointima development after endothelial injury in the rabbit aorta]. AB - Animal experiments are widely accepted in arterosclerosis research. Estrogens have lipid lowering properties and beneficial effects on the vasomotion. They act antiproliferative on those cells of the vascular wall which play a major role in lumen narrowing after vascular injury and in atherogenesis. The aim of the present study was to establish an organ culture model (rabbit aorta) to investigate post injury estrogen effects in the vessel wall. We chose the rabbit abdominal aorta which is the target organ in various animal experiments on this matter. The endothelial mono-layer was manipulated in a way that caused a measurable and reproducible post injury reaction (neointima formation). Then the effect of different estrogens (17beta-Estradiol, the phytoestrogens Genistein and Daidzein) on neointima development was investigated in male and female rabbit aortae. In equivalent dosages of 50 microg/ml all three estrogens inhibited the neointima formation significantly in male and female vessels. By the use of this organ culture model it was possible to show post injury effects of different estrogens in the vasculature while the consumption of animals was significantly reduced. Because 10 aortic segments could be taken from one aortic vessel, the number of animals that would have been necessary for an in vivo experiment could be reduced by the factor 10. PMID- 11103109 TI - [A comprehensive form for the standardized characterization of transgenic rodents: genotype, phenotype, welfare assessment, recommendations for refinement]. AB - When "creating" transgenic mouse strains it is not possible to predict their phenotype with certainty, particularly not with respect to welfare. The generation of models for (human) diseases deliberately implies a compromised health ranging from minor (clinically inapparent) to lethal. To ensure animal welfare requirements and apply the criteria of the 3R, a careful phenotype characterisation and welfare assessment has to be done routinely for each newly produced strain, at individual and strain level, starting by the standardised monitoring of founders and their consequent generations. A comprehensive form has been developed for a standardised characterisation of transgenic mouse strains. It is subdivided into basic and detail information. It can be kept up to date continuously in the form of a computerised database, incorporating growing knowledge and experience of the strain. Basic information mainly serves the requirements of housing and breeding facilities as well as the authorities in view of animal welfare measures, detail information mainly serves the interests of research and efficiency. PMID- 11103110 TI - [Comments--counter culture or cultural struggle?]. PMID- 11103111 TI - [Hypocritical "positive attitude to animal welfare" of the new German government]. PMID- 11103112 TI - [Intraoperative transesophageal echocardiography: why? how? for whom?]. PMID- 11103113 TI - [0.2% ropivacaine vs. 0.1% ropivacaine plus fentanyl in obstetric epidural analgesia]. AB - OBJECTIVE: To compare the analgesic efficacy of epidural administration of 0.2% ropivacaine alone to that of 0.1% ropivacaine plus 0.0002% fentanyl during childbirth. PATIENTS AND METHODS: We performed a prospective, randomized single blind study of 84 women in labor (aged 16 to 40 y, ASA I-II, weight over 110 kg, height over 150 cm, gestational age 37 to 42 weeks). The women were randomly assigned to two groups: group I consisted of 42 patients who received an initial bolus of 10 ml of ropivacaine 0.2% followed by continuous perfusion of ropivacaine 0.2% at a rate of 6 to 10 ml/h; group II was composed of 42 women who received an initial bolus of ropivacaine 0.2% with 50 micrograms of fentanyl followed by continuous infusion of ropivacaine 0.1% and fentanyl 2 micrograms/ml at a rate of 6 to 10 ml/h. Data recorded were parity and type of delivery, blood pressure, heart rate (HR), time to onset of pain relief, motor blockade on a modified Bromage scale, pain on a visual analog scale (VAS) and fetal HR, Apgar score and arterial and venous pH of umbilical blood. RESULTS: We found no significant differences in demographic or hemodynamic data in mothers or fetuses, in type of delivery or motor block, although the latter tended to be slightly lower in group II. In group II, the total anesthetic dose used was significantly lower (p = 0.003); time until onset of pain relief was significantly shorter (p = 0.044); and VAS scores were significantly lower at 15 min (p = 0.005), 30 min (p = 0.029), 60 min (p = 0.017) and 90 min (p = 0.002). The number of top-up boluses needed for deliveries involving instruments was significantly greater in group II (p = 0.37). CONCLUSION: The protocol of ropivacaine 0.1% with 2 micrograms/ml of fentanyl provides satisfactory analgesia throughout labor, allowing lower doses of local anesthetic to be used, with shorter onset of pain relief and reduced motor blockade; however the analgesia provided is insufficient for deliveries assisted by instruments. PMID- 11103115 TI - [A new toy: upswing in curare use in Spanish anesthesia]. AB - The introduction of curare for general anesthesia by Harold Griffith in 1942 was one of the most important moments in the development of anesthesiology. However, several years passed before curare came to be used in Spain. We review the early application of curare and the role played by Robert Macintosh, Professor of Anaesthesia at Oxford, in introducing the drug to Spain. PMID- 11103114 TI - [Efficacy of a single dose of urapidil for preventing arterial hypertension during the pre-bypass period in coronary surgery]. AB - OBJECTIVES: To assess the effect of a single prophylactic dose of urapidil for arterial hypertension during the period before start of extracorporeal circulation. PATIENTS AND METHODS: Forty-four patients with good ventricular function (ejection fraction < 40%) scheduled for coronary surgery were enrolled for prospective study. The patients were randomly assigned to receive 0.5 mg/kg of urapidil (group U, n = 22) or nothing (group N, n = 22) 3 min before skin incision. If hypertension developed sodium nitroprusside was administered, starting with a dose of 0.5 microgram/kg/min. Monitoring of arterial pressure, heart rate and ST segment (DII and V5) was continuous. The study ended with cannulation of the aorta. RESULTS: The demographic features, cardiovascular history, medication and duration of surgery were comparable in the two groups. Six patients in group U (27%) and 19 in group N (86%) developed arterial hypertension (p < 0.001), the duration of which was 2.23 +/- 4.49 min in group U and 9.64 +/- 9.7 min in group N (p < 0.05). Arterial hypotension was observed in 13 group U patients and 7 group N patients (NS). No significant differences in duration of tachycardia, bradycardia or myocardial ischemia were found. CONCLUSIONS: The administration of a single dose of urapidil prevents arterial hypertension during the phase before extracorporeal circulation for coronary surgery and reduces the need for nitroprusside. No clinically relevant side effects are evident. PMID- 11103116 TI - [Laryngeal mask for intubation (Fastrach)]. AB - The laryngeal mask for intubation (MLI), or "Fastrach", is a new device designed by Brain for airway management. The MLI, a modified version of the conventional laryngeal mask, allows for blind intubation through the device using endotracheal tubes up to 8 mm in diameter. Insertion with the head in a neutral position makes this system useful for managing the airway when neck injury is present. The device has been used successfully in patients assessed as having difficult-to manage airways and its use in emergencies inside or outside the hospital is promising. The MLI has been used with high rates of success in combination with other techniques such as fiberoptic bronchoscopy (success rate 99 to 100%) and transillumination (95 to 100% success rate) in patients whose airways have been considered difficult to manage. Given such high rates of success for MLI placement (95 to 100%) and for blind orotracheal intubation (81 to 100%), the Fastrach may offer an alternative to the conventional laryngeal mask in algorithms for airway management. PMID- 11103117 TI - [Intraoperative transesophageal echocardiography: basic aspects and recommendations for the training of the anesthesiologist. Joint Working Group of the Section on Echocardiography and Other Imaging Techniques of the Spanish Cardiology Society and the Cardiac Anesthesia Section of the Spanish Society for Anesthesiology, Resuscitation and Pain Management]. AB - Transesophageal echocardiography has demonstrated to be a very useful technique in the operating room. Although cardiologists are responsible for the performance and interpretation of conventional echocardiographic studies, anesthesiologists should have an active role in the intraoperative ones, because of their active role in the operating room and their responsibility for the evaluation and treatment of perioperative complications. The increasing practice of intraoperative echocardiography in Spain, mainly due to the interest of the anesthesiologists and cardiac surgeons, and the lack of recommendations for an adequate training in transesophageal echocardiography oriented to the anesthesiologist, has prompted the creation of a Joint Task Force including members of the Sections of Echocardiography and Cardiothoracic Surgery of the Spanish Societies of Cardiology and Anesthesia, respectively. The Task Force has draw up a set of guidelines focused on the training of anesthesiologists in intraoperative transesophageal echocardiography, with the ultimate goal of stimulating the cooperation between anesthesiologists, cardiologists and surgeons in the surgical area. PMID- 11103118 TI - [Neurophysiological monitoring during scoliosis surgery using controlled hypotension]. AB - Controlled arterial hypotension understood to be a mean arterial pressure (MAP) between 55 and 60 mmHg is often used as a complementary technique in anesthesia even though it is not without complications and associated mortality even in young patients. During surgery to reduce scoliosis in a young boy, MAP fell to 60 mmHg accompanied by bilateral loss of sensory and motor evoked potentials (SEP and MEP). Detecting the absence of SEP and MEP allowed us to prevent medullar injury due to ischemia secondary to hypotension, once possible surgical or technical causes had been ruled out. We believe that monitoring SEP and MEP is useful not only to the surgeon but also to the anesthesiologist. PMID- 11103119 TI - [Intraoperative bursting of a laryngeal mask]. PMID- 11103120 TI - [Acute transitory parotiditis secondary to general anesthesia using a laryngeal mask]. PMID- 11103121 TI - [Discovery of an intra-arterial hydatid cyst during a diagnostic thoracotomy]. PMID- 11103122 TI - [Anaphylactic shock after anesthesia induction]. PMID- 11103123 TI - [To what is apoptosis related?]. PMID- 11103124 TI - [The beginnings of drug therapy of infectious diseases in France]. PMID- 11103125 TI - [Clinical elements of the diagnostic guidelines for low back pain]. AB - If the majority of low back pain spontaneously recover in a few weeks, the main problem is to eliminate the possibility of a specific low back pain due to a serious underlying medical condition with radically different therapeutics and consequences. Medical history and physical examination will make it possible to detect these conditions (tumor, infection, spinal fracture, spondylarthropathy) and to specify useful complementary explorations. The second objective, for which we have few reliable clinical criteria, will consist in specifying the cause of low back problem. Lastly, it will be needed to detect the factors of risk of chronic course in order to improve the medical care which must be early. PMID- 11103126 TI - [Imaging strategies in common low back pain]. AB - Imaging strategy varies according to the background of acute or persistent low back pain. In case of acute low back pain with associated symptoms of a cauda equina syndrome or any atypical clinical signs PA and lateral radiographs of the lumbar spine are required. An additional MRI examination is often necessary, even with negative radiographs. In the other cases, radiographs are only obtained when the low back pain persists longer than 7 weeks. Secondary, MRI may be performed in case of worsening and persistence of the clinical symptoms or if a specific low back pain is suspected. In case of chronic low back pain, with a severe socio professional impact or a planed invasive treatment, plain films of the lumbar spine must be obtained, eventually with additional MRI examination. PMID- 11103127 TI - [Factors in the chronic progression of common low back pain]. AB - The main risk factors for developing chronic low back pain are related to comorbidity, the social circle of the patient and family status, the patient's concept of his health status, the patient's profession and the interval before medical treatment. For patients undergoing surgery, the same risk factors for chronic low back pain are found, with a predominance of professional status and age of the patient. On the contrary, the initial handicap and the type of management are of little importance in patient outcome. It is important to detect early the factors that risk to favour the transition to chronicity of low back pain. The effectiveness of both medical and socio-professional approaches to prevention remains to be evaluated and defined in prospective studies. PMID- 11103128 TI - [Local and general rest in the treatment of common low back pain]. AB - The routine prescription of rest for patients with common low back pain is now widely questioned. Local rest using lumbar corsets and back supports is useful only in case of acute or extreme low back pain, in particular if trauma is involved. In chronic low back pain, such support should be used only to facilitate pursuit of an at-risk professional activity. In acute low back pain, general rest should be indicated by the intensity of pain and not prescribed as a treatment itself. General prolonged rest in acute low back pain has now been demonstrated to entail nefarious long-term effects. In the patient with chronic low back pain, general rest will favour loss of physical condition and exacerbate the difficulties socio-professional rehabilitation. PMID- 11103129 TI - [Which rehabilitation for which low back pain?]. AB - Many rehabilitation technics for low back pain are available. Their aims are short time pain decrease, muscular strengthening in flexion or extension, increased hip and lumbar spine mobility, improved lumbar and pelvic proprioceptive sensibility, improved general fitness. During the past ten years, studies meeting widely accepted validity and applicability for therapeutic trials have addressed the clinical efficacy of rehabilitation in low back pain patients. Most studies assessing the back school approach have found no benefit. Spinal extension and flexion programs have yielded short-time improvements, without difference between the two methods. There is now strong evidence that functional restoration programs provide long-term benefits including better social and occupational outcomes. PMID- 11103130 TI - [Spinal manipulation in the treatment of common low back pain]. AB - Although the best designed studies have shown promising results, the efficacy of spinal manipulations has not as yet been proved. The mode of action may involve mechanical changes in the disk and facet joints, but neurological mechanisms probably play the key role. Complications of cervical spinal manipulations are rare. To protect patients, French legislation requires that spinal manipulations should only be performed by licensed physicians, who are capable of establishing the accurate diagnosis before undertaking manipulations. PMID- 11103131 TI - [Local and general anti-inflammatory treatment of common low back pain]. AB - Non-steroidal anti-inflammatory drugs are often prescribed with analgesics. They are proposed at the acute phase of low back pain or sciatica, or during flares of chronic back pain. Efficacy of NSAIDs have been demonstrated in acute back pain with randomised clinical trials. Efficacy is less clearly demonstrated in chronic back pain and sciatica. Local injections of steroids are required after failure of analgesics and NSAIDs, in patients with sciatica and chronic back pain. Different routes of injection are possible, depending of the symptoms and the mechanism of compression. PMID- 11103132 TI - [Indications and limitations of surgery of common low back pain]. AB - Surgical treatment of the low-back pain remains controversial in term of efficacy. Surgical treatment is advocated only when conservative management fails, a clearly identifiable cause of lumbar pain is identified, worker's compensations are detected, psychological disorders are treated and disability and pain are still present. In addition, we used a preoperative external diagnostic immobilisation with hip spica cast to select the patients. Posterior instrumented fusion is more often used, however several studies demonstrated the interest of interbody fusion when an inflammatory signal of the disc was detected on the MRI. Spinal fusion provide 50-93% of clinical success in the literature, spondylolisthesis remains the better indication. Only 30% returning to work were obtained. PMID- 11103133 TI - [Recertification: expectation or reality?]. PMID- 11103134 TI - [Criteria of imputability in accidents of drug-induced origin]. PMID- 11103135 TI - [Nephrotic syndrome. Physiopathology, diagnosis, evaluation, treatment of lipoid nephrosis]. PMID- 11103136 TI - [The post-partum period. Clinical follow-up, breast-feeding and its complications]. PMID- 11103137 TI - [Certificates. Death certificates, accident and injury certificates; drafting and issuance. The requirements]. PMID- 11103138 TI - [Alzheimer disease. Diagnosis, progression]. PMID- 11103139 TI - [Acute disseminated lupus erythematosus. Diagnosis, progression, principles of treatment]. PMID- 11103140 TI - [Vertigo. Diagnostic guidelines]. PMID- 11103141 TI - [Statistical methods necessary for the analyses of information from clinical tests--with special reference to problems in testing significance levels and multivariate analyses]. PMID- 11103142 TI - [Evaluation of diagnostic possibility of clinical tests]. PMID- 11103143 TI - [Estimation of normal ranges]. PMID- 11103144 TI - [Methods for quality control]. PMID- 11103145 TI - [Methods for database management]. PMID- 11103146 TI - [Standards and standardization]. PMID- 11103147 TI - [Reporting of laboratory test results]. PMID- 11103148 TI - [Data interpretation]. PMID- 11103149 TI - [Evidence-based medicine and laboratory diagnosis]. PMID- 11103150 TI - Leprosy elimination campaigns. PMID- 11103151 TI - Hunterian Lecture. The ontogeny of the peptide innervation of the human pylorus with special reference to understanding the aetiology and pathogenesis of infantile hypertrophic pyloric stenosis. AB - Infantile hypertrophic pyloric stenosis is the most common cause for urgent abdominal surgery in infancy. The aetiology of the condition is unknown. The ontogeny of the innervation and structure of the normal infant pylorus is unknown. A variety of differing histological features have been attributed to this condition and a number of animal models have been described. The histological changes in the human condition and those in the animal models have not been quantified and statistically verified. Thus, precise comparisons cannot be made. Immunohistochemistry was the principal technique employed in this study. Using this technique, the ontogeny and structure of the normal infant pylorus have been documented. The morphological and immunohistochemical changes underlying infantile hypertrophic pyloric stenosis have been quantified for the first time and compared with the quantified changes in natural and experimental animal models of this condition. PMID- 11103152 TI - Gastric stromal tumours: a practical approach. AB - Recent findings on the pathological diversity of gastric stromal tumours and their unpredictable behaviour prompted us to review our series of 16 patients who had undergone surgery for these tumours from 1991 to 1998. There were 13 benign and 3 malignant lesions. The majority of patients presented with either upper gastrointestinal bleeding or anaemia alone (12 of 16). Endoscopy was an extremely useful diagnostic tool, revealing the lesion as an intraluminal protuberant tumour with or without ulcer in 10 cases and as an ulcer alone in 4 cases, and in 1 case features suggesting an extrinsic mass. All the patients in the series underwent surgery. We used staplers (AutosutureR TA 55) to excise the tumours in 7 cases, all of which on histological examination were benign with clear resection margins. Gastric resections were performed in 5 cases for either large tumours or those situated at the fundus or antrum and local excision of the remaining 4. The mean follow-up of these patients was 24 months. Two patients with malignant lesions died of irresectable recurrences, one 2 months and one 18 months after surgery. There have been no recurrences in the tumours diagnosed as benign on histology. Tumour size, position and the ability to apply the stapler leaving adequate margin below the tumour should be the determinants of extent and type of excision. Reliable determinants of behaviour are tumour size, grade and mitotic index. PMID- 11103153 TI - Management of pancreatic pseudocysts. AB - BACKGROUND: This review analyses the outcome for patients with acute and chronic pancreatic pseudocysts managed in two major referral centres. PATIENTS AND METHODS: From 1987 to 1997, 33 patients were treated with either acute (n = 19) or chronic (n = 14) pseudocysts. Procedures performed included cystgastrostomy (64%), cystduodenostomy (6%), cystjejunostomy (3%), distal pancreatectomy with resection of pseudocyst (12%), laparotomy with external drainage (9%), endoscopic transpapillary stenting (3%) and endoscopic pancreatic duct sphincterotomy with percutaneous drainage of the pseudocyst (3%). RESULTS: All patients had resolution of their pseudocyst and no patient developed recurrence. There were no deaths in this series. There was a 9% incidence of major complications and a 21% incidence of minor complications. Outcome was excellent in 63% and good in 27% of patients. Two patients (6%) had persistent chronic pain and one patient (3%) had evidence of exocrine pancreatic insufficiency with malabsorption. CONCLUSIONS: Surgical internal drainage of pancreatic pseudocysts can be performed safely with low morbidity and mortality provided patients are carefully selected and their medical management is optimized. Although minimally invasive techniques now offer a variety of treatment options, open surgical drainage is still indicated for a significant number of cases. PMID- 11103154 TI - Laparoscopic appendicectomy: safe and useful for training. AB - Debate exists about the benefits of laparoscopic appendicectomy when compared to a conventional open procedure. The majority of appendices are removed by the open route in the UK. We report a series of 132 cases of suspected appendicitis managed laparoscopically: 112 (85%) of the patients had acute appendicitis, the remaining 20 (15%) had non-appendiceal pathology. The median operative time was 30 min and there were no conversions to an open operative procedure. The median postoperative stay was two days. Complications were seen in two patients. The published evidence comparing laparoscopic and open appendicectomy is contradictory. Our series shows that laparoscopic appendicectomy is a safe procedure with low morbidity; it is also an excellent training tool in laparoscopic technique and, with sufficient experience, takes no longer than an open procedure. Negative appendicocecotomies are most common in women of fertile age and can be associated with significant morbidity; therefore, laparoscopy should be used to make the diagnosis and, if appendicitis is the cause, the appendix could safely be removed laparoscopically. However, the choice between open and laparoscopic procedure is a subjective decision for the patient and their surgeon. Laparoscopic appendicectomy cannot be regarded as the gold standard. PMID- 11103155 TI - Emergency surgical admissions in patients aged more than 80 years: a study over four decades. AB - BACKGROUND: The proportion of older patients in the community is rising. The aim of this study was to determine the trend in emergency surgical admissions in patients over 80 years of age in 1997 compared with the previous three decades. PATIENTS AND METHODS: Data were obtained on all patients over 80 years of age admitted as general surgical emergencies in 1997 to the Royal Berkshire and Battle Hospitals, Reading, UK. Reasons for admission, management, mortality and duration of hospital stay were recorded and compared with results from 1966, 1976 and 1989. RESULTS: During 1997, 4807 patients over the age of 80 years were admitted as emergencies to all specialities. Of these, 447 (9.3%) were surgical. This compares with 122 in 1966, 248 in 1976 and 339 in 1989. Emergency surgical workload in patients over 80 years of age had increased from 6.2% in 1966 to 12% in 1997. A random sample of 261 patients was analysed. In-patient mortality was 13.8% in 1997 compared with 21.8% for 1976 and 22.4% for 1989. Median length of stay was 8 days (range, 0-41 days) for 1997 and 1989 compared with 14 days in 1976. Twenty-four patients either needed admission to other specialities or need not have been admitted as emergencies at all and were classified as inappropriate admissions to the general surgical ward. CONCLUSIONS: The trend of increased number of patients over the age of 80 years being admitted as emergencies to general surgery continues through four decades. There has been a decrease in mortality and length of stay since 1966, but no decrease in length of stay in 1997 compared with 1989. Avoiding inappropriate admissions would result in a significant improvement in bed utilisation for elective surgery and help to reduce waiting lists. PMID- 11103156 TI - Early results of inguinal hernia repair by the 'mesh plug' technique--first 200 cases. AB - INTRODUCTION: Inguinal hernia repair is the most common surgical procedure performed in the UK. Evidence from several earlier studies suggests that primary inguinal hernia repair has a high recurrence rate of 10-15%. The Royal College of Surgeons of England guidelines suggested the use of layered suture (Shouldice) or prosthetic (Lichtenstein) repair. Per-fix plugs have been used in the US for more than a decade with excellent results. This study was a series of 200 consecutive cases. The aim was to evaluate the mesh plug technique in the repair of all types of inguinal hernias and its results in one consultant practice within a district general hospital. PATIENTS AND METHODS: In a 15-month period between 1997 and 1998, all patients with inguinal hernias presenting to the general surgical clinic of one consultant were recruited to the study. All had mesh plug repair under local (n = 40), regional (n = 50) or general (n = 110) anaesthesia either by the consultant, associate specialist or specialist registrar (following initial training), using the same standard technique. The majority 80% (n = 160) were done as day cases. The results were evaluated by questionnaire and personal outpatient review initially at 3 weeks, then at 1 year (9-13 months). RESULTS: 200 consecutive patients with inguinal hernias underwent mesh plug repair; mean age was 54 years (95% CI, 46-61). The majority of patients had primary (n = 180) and others had recurrent (n = 20) hernia. All types of hernia (Gilbert's I-VII) were included. Median follow-up was 1 year (9-15 months). Groin pain, which was the leading symptom at presentation, was relieved in 96% of the patients; 79% returned to previous jobs within 4 weeks (95% CI, 0.71-0.87). All retired patients resumed normal life activities within 2 days. Postoperative pain was minimal; 28 patients did not require any postoperative analgesia. There were very few minor (n = 6) and no major complications. During the follow-up, one recurrence occurred. CONCLUSIONS: Mesh plug repair is associated with minimal postoperative pain, quick recovery and return to work. It is an ideal technique for day-case surgery. Although longer follow-up will be required to assess true recurrence rate, so far the recurrence rate at 0.5% is acceptable, particularly in the light of other published series. PMID- 11103157 TI - Performance data and the mortuary register. AB - OBJECTIVE: To compare departmental records of deaths after cardiac surgery with the hospital's information system. DESIGN: Matched pairs comparisons: (i) historic record compared with current record from another source; (ii) contemporary records from different sources; and (iii) timed records from different sources. SETTING: Regional cardiothoracic units at St George's and St. Thomas's Hospitals. SUBJECTS: 2664 cardiac surgical operations at St George's between January 1992 and June 1994, 215 deaths in the cardiac surgery database at St Thomas's between April 1993 and March 1997, 120 in-hospital deaths received by the mortuary at St George's during June 1999. MAIN OUTCOME MEASURES: The difference in the number of in-hospital deaths from departmental, hospital, and mortuary sources. RESULTS: Four of 2664 operations (0.15%) had been incorrectly coded as leaving hospital alive. Fewer than 80% of the actual number of deaths after cardiac surgery at St Thomas's had been recorded on either the departmental database or the hospital administration system. For 9% of deaths received in the mortuary, it took more than 6 working days for the hospital record to be updated, and at the time of reporting 1 case had not been updated after 14 working days: the date of death was inaccurate in 4/113 (3.5%) of cases. CONCLUSIONS: The mortuary staff can contribute to improving the accuracy of body counts. Death rates and performance data should not be published without statistical peer review. PMID- 11103158 TI - The passage of bacteria through surgical drapes. AB - The passage of bacteria through surgical drapes is a potential cause of wound infection. Previous studies have shown that liquids and human albumin penetrate certain types of drapes. We studied the passage of bacteria through seven different types of surgical drape and an operating tray. Bacteria easily penetrated all the woven re-usable fabrics within 30 min. The disposable non woven drapes proved to be impermeable, as did the operating tray. We recommend the use of non-woven disposable drapes or woven drapes with an impermeable operating tray in all surgical cases. PMID- 11103159 TI - A prospective, randomised trial of prophylactic antibiotics versus bag extraction in the prophylaxis of wound infection in laparoscopic cholecystectomy. AB - Septic complications are rare following laparoscopic cholecystectomy if prophylactic antibiotics are given, as demonstrated in previous studies. Antibiotic treatment may be unnecessary and, therefore, undesirable, so we compared two forms of prophylaxis: a cephalosporin antibiotic and bag extraction of the dissected gallbladder. A total of 76 patients undergoing laparoscopic cholecystectomy were randomised to either receive an antibiotic or to have their gallbladder removed from the abdomen in a plastic bag. Complicated cases were excluded. There was a total of 6 wound infections (7.9%), 3 in each of the study groups. All these were associated with skin commensals. There were no other septic complications. Bacteriological studies grouped the organisms isolated from the bile and the wound as potential pathogens and likely commensals. A total of 10 potential pathogens were isolated, 9 of which were found in the group receiving antibiotics. We conclude that septic sequelae of uncomplicated laparoscopic cholecystectomy are uncommon, but clearly not entirely prevented by antibiotic or mechanical prophylaxis. Prophylactic antibiotics may not be required in uncomplicated laparoscopic cholecystectomy. Further study is warranted. PMID- 11103160 TI - Epidermoid cysts of the testis: the case for conservative surgery. AB - The series comprises 6 patients (mean age, 21 years) who presented with an epidermoid cyst of the testis between 1991 and 1998. Pre-operative ultrasonography suggested the presence of a testicular cancer in 3 patients who underwent a radical orchidectomy. The ultrasound successfully predicted the true diagnosis in 3 patients who had a wedge excision of the cyst together with a cuff of normal surrounding tissue. All patients are free of disease with a mean follow up of 3 years. With increasing awareness of the condition coupled with accurate pre-operative radiological imaging, local excision of an epidermoid cyst with preservation of the remainder of the testis is now a feasible and rational alternative to more radical surgery. PMID- 11103161 TI - Open access carotid duplex scanning: throughput and resultant surgical workload. AB - It is now generally accepted that high-quality duplex scanning obviates the need for pre-operative angiography in the assessment of most cases of carotid disease. An information leaflet was circulated to all general practitioners and hospital doctors in early 1996. Indications, results of treatment and details of the open access service were described. This paper outlines the workload that followed, for both imaging and surgical departments. PMID- 11103162 TI - Effect of left renal vein division during aortic surgery on renal function. AB - A total of 398 consecutive patients underwent surgery for an aneurysm or occlusive disease of the aorta at Norfolk and Norwich Hospital between December 1994 and October 1998. It was necessary to divide the left renal vein in 58 (14.6%) cases. We examined the effect of this division on the mortality rate and renal function. Renal function was assessed by measuring serum creatinine pre operatively, peri-operatively and long-term postoperatively. There was no significant difference in the mortality rate between patients who had the left renal vein divided (LRVD) and in whom the left renal vein remained intact (LRVI)- 31% versus 32%, P = 0.83. There was no significant difference in the pre operative serum creatinine level between both groups (107 +/- 21 mumol/l in LRVD versus 103 +/- 29 mumol/l in LRVI, P = 0.14). There was an insignificant rise in the mean serum creatinine 7 days postoperatively (111 +/- 21 mumol/l in LRVD versus 107 +/- 31 mumol/l in LRVI, P = 0.05). The mean serum creatinine returned back to the pre-operative level at 30 days postoperatively (106 +/- 16 mumol/l in LRVD and 105 +/- 29 mumol/l, P = 0.20). After 1 month, there was no significant difference in the number of patients who had a sustained elevation of serum creatinine level (7.5% in LRVD versus 2.7% in LRVI, P = 0.11). We feel that division of the left renal vein is a safe and helpful procedure during juxtarenal aortic surgery. PMID- 11103163 TI - Rectal strictures following abdominal aortic aneurysm surgery. AB - Rectal stricture formation is a rare complication of aortic aneurysm repair. Two case are described here. A combination of hypotension, a compromised internal iliac circulation and poor collateral supply following inferior mesenteric artery ligation can result in acute ischaemic proctitis--an infrequently described clinical entity. Ulceration and necrosis are the sequelae of prolonged ischaemia and fibrous stricture formation may result. One patient responded to dilatation and posterior mid-rectal myotomy; the other failed to respond to conservative measures and eventually had an end colostomy fashioned following intractable symptoms. PMID- 11103164 TI - Smoking--do vascular surgeons practise what they preach? AB - Smoking is a major health problem in Great Britain and cigarette consumption is rising. Although there are studies concerning the smoking habits of hospital physicians, nurses and oral and maxillofacial surgeons, little is known about the smoking habits of vascular surgeons and the advice given by them to their patients. A questionnaire survey was conducted involving 422 members of the Vascular Surgical Society of Great Britain and Ireland. The response rate was 74%. The median age of responders was 51 years (range, 32-69 years) of whom 98% were men. Of responders, 98% routinely advise patients to stop smoking, 10% advise nicotine gum/patch, 39% provide antismoking information sheets, 11% are involved in an antismoking clinic/group and 74% check to see whether patients continue to smoke. The majority of responders would be prepared to offer revascularisation in patients who continue to smoke. Only 8 surgeons (3%) would not advise revascularisation in this group of patients. Only 10% of respondents were current smokers, 37% were ex-smokers and 53% had never smoked. Vascular surgeons, therefore, seem to practise what they preach. PMID- 11103165 TI - Delivery of low molecular weight heparin for prophylaxis against deep vein thrombosis using a novel, needle-less injection device (J-Tip). AB - Given daily, low molecular weight (LMW) heparins are established for prophylaxis against deep vein thrombosis (DVT). We describe delivery by a novel, needle-less device that is virtually painless in action. Its use could provide benefits for patients in terms of comfort both psychologically and physically, and for healthcare workers in terms of safety from needle-stick injury. Patients undergoing elective surgery received LMW heparin delivered subcutaneously by either a standard needle and syringe or by the needle-less injection device, J Tip. Pain was scored at the time of injection and plasma anti-factor Xa levels compared between the two methods of drug delivery 4 h later: 29 patients received LMW heparin delivered by the J-Tip and 31 patients by standard needle and syringe. The J-Tip was significantly more comfortable for the patient as the method of drug delivery (P < 0.001). When delivered by the J-Tip, LMW heparin was equally as efficacious, as plasma anti-factor Xa levels were similar for both methods of delivery (P < 0.42). In summary, delivery of LMW heparin by the J-Tip device was both comfortable and effective. These findings, taken in conjunction with its ease of use and complete freedom from risk of needle-stick injury might encourage further examination and use of this type of product. PMID- 11103166 TI - A 'safe' surgical technique for stabilisation of the sternoclavicular joint: a cadaveric and clinical study. AB - In symptomatic patients with recurrent anterior sternoclavicular dislocation, surgery may be required to stabilise the joint. Posterior sternoclavicular dislocations may also require open reduction and stabilisation due to the complications that may arise. We present a new, 'safe' technique of surgical stabilisation of the sternoclavicular joint that is not technically demanding and does not require exposure of the first rib, as is often the case in other methods described. The repair was tested in cadavers before being employed in three patients and was found to be effective under both static and dynamic loading. The early clinical results prove encouraging. PMID- 11103167 TI - Audit of open tibial diaphyseal fracture management at a district accident centre. AB - Preston Acute Hospital is a designated district accident centre with a 24 h flying squad and on-site plastic and orthopaedic units. We performed a retrospective 5-year survey of open tibial shaft fracture management at our unit and compared our treatment to the guidelines of the British Orthopaedic Association (BOA) and British Association of Plastic Surgeons (BAPS). Deficiencies were highlighted and changes in practice made. We then re-audited our figures over an 18-month period to see if clinical improvements had been made. The audit demonstrated an incidence of open tibial shaft fractures of 15 per 50,000 new patients per year in accident and emergency. Gustilo grading, and thus full appreciation of soft tissues injury, was being underestimated, with 8% of the injuries undergraded at the time of surgery: 17% of Gustilo IIIA and 85% Gustilo IIIB required flap cover. Seventy-four percent of patients received their first orthopaedic procedure within the recommended 6 h of admission, but despite the on-site plastics unit, only 50% of cases in the initial survey had their soft tissue defect covered by the recommended 5 days. After changes to practice, 80% patients received their first orthopaedic procedure within the recommended 6 h of admission, and all had their soft tissue defect covered within 5 days; 5% of cases required fasciotomy to relieve compartment syndrome. We highlight features to alert the high energy (Gustilo III) status and recommend immediate involvement of plastic surgical colleagues with these injuries. We also highlight a high incidence of compartment syndrome in the young male patient with the lower energy Gustilo I injury. PMID- 11103168 TI - Technical and medico-legal suggestions for Liechtenstein hernia repair. PMID- 11103169 TI - The use of two face masks to prevent clouding of spectacles and surgical visors. PMID- 11103170 TI - Ethics in publishing; are we practising to the highest possible standards? PMID- 11103171 TI - Ropivacaine in children. PMID- 11103172 TI - Pharmacokinetics and clinical efficacy of long-term epidural ropivacaine infusion in children. AB - The clinical efficacy and pharmacokinetics of long-term epidural ropivacaine infusion were investigated in 18 postoperative children aged between 0.3 and 7.3 yr. A lumbar or thoracic epidural catheter was inserted after the anaesthetic induction. Sixty minutes following a bolus dose of ropivacaine 1 mg kg-1, 0.2% ropivacaine was infused at a fixed rate of 0.4 mg kg-1 h-1 for a mean of 61.3 h (range 36-96 h). Clinical evaluation comprised hourly recording of pain, sedation, motor block, nausea/vomiting, pruritus-scores, SpO2, pulse and respiratory rates, and recording of non-invasive arterial pressure every 4 h. Total and free plasma concentrations were measured by high-performance liquid chromatography at 0, 1, 6, 12, 24, 36, 48, 72 and 96 h. Analgesia was of high quality and side effects were minor. No clinical signs of local anaesthetic toxicity were seen. Total (100-3189 micrograms litre-1) and free (10-56 micrograms litre-1) ropivacaine concentrations were within the range reported to be 'safe' in previous studies in adults. Mean (95% CI) volume of distribution was 3.1 litre kg-1 (2.1-4.2 litre kg-1), total clearance was 8.5 ml kg-1 min-1 (5.8 11.1 ml kg-1 min-1), free clearance was 220 ml kg-1 min-1 (170-270 ml kg-1 min-1) and elimination half-life was 4.9 h (3.0-6.7 h). PMID- 11103173 TI - Is obstructive sleep apnoea a rapid eye movement-predominant phenomenon? AB - Obstructive sleep apnoea (OSA) is thought to be worse during rapid eye movement (REM) sleep. REM rebound in the late postoperative period can follow the REM suppression shown to occur after some types of surgery. This is thought to worsen nocturnal episodic hypoxaemia, leading to greater cardio-respiratory risk. We set out to determine if OSA was a REM-predominant phenomenon. We reviewed the sleep clinic records of 64 consecutive patients with a diagnosis of OSA on full overnight polysomnography and sufficient data to determine the presence of a sleep stage predominance. OSA was diagnosed if the number of apnoeas/hypopnoeas per hour of sleep, the respiratory disturbance index (RDI), was greater than 10. The variables recorded for the purposes of this study were the RDI and the minimum blood oxygen saturation using pulse oximetry (SpO2min) for both REM and non-rapid eye movement (NREM) sleep. All values are presented as mean (SD). The Wilcoxon signed rank test was used for statistical analysis. The means for NREM and REM RDI were, respectively, 36 (26) and 38 (27) per hour (P = 0.96). In 32 of the 64 patients (50%) the RDI in NREM was greater than in REM. Thirty-one (48%) had a larger number during REM. One patient had identical RDIs for both REM and NREM. Sixty-two patients had satisfactory pulse oximetry recordings for both NREM and REM, and the mean SpO2min values were, respectively, 84 (7) and 82 (13)% (P = 0.15). Twenty-nine patients (47%) had a lower SpO2min in REM (seven by more than 10% and two by more than 40%), while 24 (39%) were lower in NREM (two by more than 10%). Nine patients (14%) had identical values in REM and NREM. In contrast to suggestions that OSA is a REM-predominant phenomenon, this study suggests that respiratory disturbance is not greatly affected by sleep stage, in most patients. While a small number clearly desaturate much more during REM, the majority do not. Thus, postoperative REM rebound may worsen OSA in some patients, but in many it may do otherwise. The implications of postoperative sleep disturbance are therefore likely to be more complex than previously suggested. PMID- 11103174 TI - Bispectral analysis of the electroencephalogram does not predict responsiveness to verbal command in patients emerging from xenon anaesthesia. AB - The bispectral index (BIS) is derived empirically from the electroencephalogram database of patients receiving common anaesthetics, but it may not be valid for uncommon agents. Therefore, we investigated how xenon affects the BIS. Nine and 11 patients were anaesthetized with 0.8 of the minimal alveolar concentration (MAC) of isoflurane (0.92%) and xenon (56%), respectively. After the end of operation, these concentrations were decreased in decrements of 0.1 MAC (isoflurane 0.12% or xenon 7%) and each new concentration was maintained for 15 min. This was repeated until the patient first opened her eyes or squeezed the investigator's hand on command. Isoflurane and xenon at 0.8 MAC reduced the BIS to a median of 40 (range 36-53) and 36 (30-61), respectively. With decreasing concentrations of isoflurane, the BIS increased progressively and it reached a median of 96 (90-98) when the patients awoke. In contrast, four patients receiving xenon responded to verbal command while the BIS was below 50 [median 45 (range 41-49)]. The remaining seven patients in the xenon group awoke when their BIS was greater than 80 [median 96 (range 82-98)], but in four of them the BIS was no greater than 50 when the xenon concentration was only 0.1 MAC (7%) higher than that associated with awakening. We conclude that low BIS values (< 50) do not guarantee adequate hypnosis during xenon anaesthesia. PMID- 11103176 TI - Nodal rhythm and bradycardia during inhalation induction with sevoflurane in infants: a comparison of incremental and high-concentration techniques. AB - We studied heart rate and rhythm changes during sevoflurane inhalation induction in 60 healthy, unpremedicated infants. Patients were allocated randomly to receive an incremental (2% sevoflurane, increased every four to six breaths by 2% increments, to 8%) or high-concentration induction technique (8% sevoflurane from the outset). The ECG was recorded for 330 s (30 s pre- and 300 s postinduction) using a mini-Holter device (Recollect Dual Channel, Hertford Medical) and later analysed by an independent observer. Twelve patients developed nodal rhythm (six in each group), but no other dysrhythmias were recorded. The onset of nodal rhythm was associated with bradycardia (< 80 bpm) in seven out of 12 cases, and occurred significantly earlier in the high-concentration group (median 123 (range 99-139) s versus 164 (127-138) s). Its duration was similar in both groups (62 (2 84) s versus 90 (20-167) s). These findings highlight the importance of using continuous ECG analysis when studying volatile anaesthetic agents in young children. PMID- 11103175 TI - Intravenous opioids reduce airway irritation during induction of anaesthesia with desflurane in adults. AB - Desflurane is not used for the induction of anaesthesia despite its favourable pharmacokinetic characteristics because it causes airway irritation. We investigated whether pretreatment with i.v. narcotics reduced unwanted effects. One hundred and eighty adults were randomized to three groups (60 per group) to receive i.v. saline, fentanyl 1 microgram kg-1 and morphine 0.1 mg kg-1, respectively, before inhalational induction with desflurane in nitrous oxide and oxygen. Mean time to loss of response to commands was 4.0 min, without significant differences between groups. The incidence of coughing was greater (25%) in the control group than in the fentanyl (5.0%) and morphine groups (8.3%). The incidence of apnoea was 20.0% in the control group versus 13.3 and 5.0% in the fentanyl and morphine groups, respectively. Laryngospasm developed in 11.7% of controls compared with 3.3 and 1.7% in the fentanyl and morphine groups, respectively. More patients in the control group had excitatory movements (46.7%) than in the fentanyl (16.7%) and morphine (8.3%) groups. These results demonstrate that i.v. opioids reduce airway irritability significantly during inhalational induction with desflurane in adults. PMID- 11103177 TI - Estimation of pulmonary blood flow from sinusoidal gas exchange during anaesthesia: a theoretical study. AB - We simulated the use of simultaneous sinusoidal changes of inspired O2 and N2O (Williams et al., J Appl Physiol, 1994; 76: 2130-9) at fractional concentrations up to 0.3 and 0.7, respectively, to estimate FRC and pulmonary blood flow (PBF) during anaesthesia, using O2 as an insoluble indicator. Hahn's approximate equations, which neglect the effect of pulmonary uptake and excretion on expiratory flow, estimate dead space and alveolar volume (VA) with systematic errors less than 10%, but yield systematic errors in PBF which are approximately proportional to FIN2O in magnitude. A correction factor (1 - P)-1 for Hahn's equations for PBF (where P is the mean partial pressure of the soluble indicator) reduces the dependence of PBF estimates of FIN2O, and the solution of equations describing the simultaneous mass balance of both indicators yields accurate results for a wide range of mean FIN2O. However, PBF estimates are sensitive to measurement errors and a third gas must be present to ensure that the indicator gases behave independently. PMID- 11103178 TI - Left ventricular pressure-area relations as assessed by transoesophageal echocardiographic automated border detection: comparison with conductance catheter technique in cardiac surgical patients. AB - The aim of this study was to validate measurements of intraoperative left ventricular (LV) area by transoesophageal echocardiography against simultaneous measurements of LV volume by conductance catheter (CC) in cardiac surgical patients with normal systolic LV function. Echo area was compared with CC volume during steady state and during acute changes of pre- and afterload by partial clamping of the inferior vena cava and the ascending aorta in eight patients scheduled for coronary artery bypass grafting. At steady state, Bland-Altman analysis of 32 recordings revealed a bias (SD) of 0.6% (2.5%) between echo area and CC volume, related to the initial values of end-diastolic area (100% area) and volume (100% volume), respectively. During loading interventions, bias between the two methods, as assessed by 112 measurement sequences, was 0.5% (3.7%) during aortic occlusion and -3.9% (4.4%) during cava occlusion at end systole (P < 0.0001); at end-diastole, this bias was 1.3% (4%) during aortic occlusion and 0.2% (5.7%) during cava occlusion (P < 0.0001). Intraoperative area measurements with transoesophageal echocardiography in cardiac surgical patients with normal systolic LV function show good correlation with CC volume measurements under steady-state conditions. During acute unloading by vena cava occlusion, the resulting small end-systolic echo area measurements differ significantly more from CC volume measurements than during acute increase in afterload by aortic occlusion. PMID- 11103179 TI - Effects of amrinone on ischaemia-reperfusion injury in cirrhotic patients undergoing hepatectomy: a comparative study with prostaglandin E1. AB - The effects of amrinone, a selective phosphodiesterase III inhibitor, on liver ischaemia reperfusion injury have not yet been clarified. Forty-five patients with hepatocellular carcinoma who underwent partial liver resection using Pringle's manoeuvre were studied. Patients were divided into three groups: those given amrinone, those given prostaglandin E1 (PGE1) and those not treated (controls). An indocyanine green (ICG) clearance test was performed before the operation and three times during surgery: just before induction of liver ischaemia, just after liver resection and 60 min after reperfusion. Blood lactate and base excess were measured at the same times. Systolic and diastolic arterial pressure, heart rate, cardiac index and oesophageal temperature were monitored. Aminotransferase levels were recorded the day before surgery, 1 h after operation and on the first and third postoperative days. These data were compared between groups. The ICG elimination rate, lactate and base excess in the amrinone group differed significantly from those in controls during the observation period (P = 0.03, P = 0.04 and P = 0.03, respectively). The differences between the PGE1 and control groups were not significant. There were no significant differences between the groups in perioperative vital signs, cardiac index or postoperative aminotransferase. Amrinone enhanced intraoperative ICG elimination in cirrhotic patients who underwent liver resection. PMID- 11103180 TI - Blood propofol concentration and psychomotor effects on driving skills. AB - We studied psychomotor performance in 10 healthy volunteers during recovery after a target-controlled infusion of propofol. Choice reaction time, dual task tracking with secondary reaction time and a within-list recognition task were assessed at target blood propofol concentrations of 0.8, 0.4 and 0.2 microgram ml 1. Performance was impaired most at the highest blood propofol concentration (choice reaction time increased by a mean of 247 ms and secondary reaction time by a mean of 178 ms). Choice reaction time and dual task tracking with secondary reaction time were the most sensitive and reliable methods of assessment (significant difference from baseline (P < 0.05) at a propofol concentration of 0.2 microgram ml-1 with choice and secondary reaction time testing). Within-list recognition assessment of memory was not sufficiently sensitive at very low propofol concentrations. The impairment in choice and secondary reaction time with a blood propofol concentration of 0.2 microgram ml-1 was less than that observed with a blood alcohol concentration of 50 mg 100 ml-1 and no greater than that observed with a blood alcohol concentration of 20 mg 100 ml-1 in a previous study involving healthy volunteers. PMID- 11103181 TI - Blood alcohol concentration and psychomotor effects. AB - This study assessed the effect of intravenous alcohol infusions on psychomotor impairment and compared it with that of alcohol administered orally. Comparisons were made between three European drink-driving limits of blood alcohol concentration (BAC) (20, 50 and 80 mg 100 ml-1) and an oral dose of alcohol 0.75 mg kg-1. Twelve volunteers, aged 22-34 yr, were recruited. At targets of 20, 50 and and 80 mg 100 ml-1, the mean (SD) BAC was 22.1 (3.7), 51.5 (3.3) and 80.5 (4.2) mg 100 ml-1, respectively. The peak BAC following an oral dose of alcohol 0.75 mg kg-1 ranged from 19 to 68 mg 100 ml-1. In psychomotor testing, choice reaction time deteriorated with increasing BAC and showed significant differences between baseline and the 50 (P < 0.05) and 80 mg 100 ml-1 (P < 0.01) conditions. Dual-task secondary reaction time deteriorated with increasing BAC and showed a statistically significant difference between all groups and baseline (oral and 20 mg groups, P < 0.05; 50 and 80 mg groups, P < 0.01). Dual-task tracking in the 50 and 80 mg groups was significantly different from baseline (P < 0.05 and P < 0.01, respectively). Oral dosing resulted in widely variable BACs, making it difficult to assess psychomotor impairment reliably. An intravenous infusion enables the BAC to be maintained within a narrow range. This allows precision when investigating the effects of alcohol on psychomotor performance. PMID- 11103182 TI - Continuous propofol anaesthesia for patients with myotonic dystrophy. AB - Myotonic dystrophy, a rare genetic disorder, may pose a serious problem to the anaesthesiologist due to muscular and extramuscular involvement. Thirteen patients, median age 21 yr were anaesthetized by continuous propofol infusion, fentanyl, atracurium and N2O to evaluate this combination in myotonic dystrophy. Intraoperatively, neither exaggerated reactions nor haemodynamic instability was observed. Recovery was smooth and quick. Although there was a significant decrease in mean postoperative vital capacity (965 (349) ml) from the preoperative value (1664 (566) ml, P = 0.0028), there was no change in mean postoperative SpO2 and there were no perioperative respiratory complications. Only two patients complained of nausea and vomiting. Similarly, muscular hypertonia and shivering were not observed. We conclude that the combination of continuous propofol infusion and fentanyl was a successful anaesthetic technique in these young myotonic dystrophy patients undergoing peripheral surgery. PMID- 11103183 TI - Resistance of laryngeal mask airway and tracheal tube in mechanically ventilated patients. AB - We compared the airflow resistance of 7.5 and 8.5 mm internal diameter (i.d.) endotracheal tubes (ETTs) with that of a size 4 laryngeal mask airway (LMA). We thought that any difference in the resistance of the devices alone might be offset by the resistance of the larynx. Sixteen adult ASA physical status I and II patients (14 males, two females) undergoing general anaesthesia were anaesthetized and paralysed with intravenous propofol, ketamine and vecuronium. After insertion of the LMA, controlled ventilation (tidal volume 10 ml kg-1, frequency 12 min-1) was established with three different settings for inspiratory flow (5.5, 7.5 and 12.5 ml kg-1 s-1). Ventilation with the same settings was used after orotracheal intubation with an ETT of i.d. 7.5 mm (females) or 8.5 mm (males). The position of the LMA mask and the tip of the ETT were checked through a fibrescope. The resistance of the devices and, in case of the LMA, of the larynx, was derived by relating proximal and distal pressures (measured via catheters) to inspiratory flow. Four patients--young, tall men--had to be excluded from further study because of a leak around the LMA. In the remaining 10 males and two females, resistance of the LMA (mean (SD) at high flow, 1.19 (0.22) mbar.s litre-1 in males) was less than that of the 8.5 mm i.d. ETT (3.34 (0.52) mbar.s litre-1) (P < 0.01). However, the structures between the LMA and the trachea added another, highly variable, resistance component, so that the mean resistance of the LMA and larynx together was similar (in males: 3.20 (2.71) mbar.s litre-1) to that of the 8.5 mm ETT. In eight patients the epiglottis projected on to one-tenth to two-thirds of the distal opening of the LMA; this was in no case associated with greater resistance. Greater resistance occurred in two patients with a central LMA position and unobstructed view of the glottis and in one patient with marked lateral deviation. In conclusion, there is no clinically relevant difference between the resistance of a size 4 LMA plus that of the larynx and that of an 8.5 mm i.d. ETT. PMID- 11103184 TI - Influence of different colloids on molecular markers of haemostasis and platelet function in patients undergoing major abdominal surgery. AB - Synthetic colloids have been reported to cause haemorrhagic complications. The effects of perioperative volume replacement with 4% gelatin (n = 20), 6% low molecular weight (LMW) hydroxyethyl starch (HES) (Mw: 70,000 dalton; HES 70/0.5; n = 20) and 6% medium-molecular weight (MMW) HES (Mw: 200,000 dalton; HES 200/0.5; n = 20) on haemostasis were assessed in patients undergoing major abdominal surgery. Volume was administered to keep central venous pressure (CVP) between 10 and 14 mm Hg. Conventional global coagulation tests, molecular markers of coagulation, and platelet function (using a platelet function analyser (PFA 100) with ADP as inductor) were monitored prior to surgery (T0), at the end of surgery (T1), 4 h after the end of surgery (T2), and on the morning of the first postoperative day (T3). Significantly more gelatin (2900 (SD 320) ml) than HES 200 (2150 (312) ml) was given during the study period. Bleeding and the use of allogeneic blood-blood products were similar in all groups. Markers of thrombin generation (F1 + 2), of thrombin neutralization (TAT III complex), and of fibrin formation and its degradation (D-dimer) increased significantly during and after surgery without showing significant group differences. Factor VIII and von Willebrand factor (vWF) also increased in all groups beyond the normal range, showing the significantly highest increase in the gelatin-treated group (VIII: from 173 (36) to 266 (33) U dl-1; vWF: from 164(33) to 238 (31) U dl-1). Platelet function remained within the normal range and without group differences throughout the study period. We can conclude that all three solutions can be used safely in patients undergoing major abdominal surgery with regard to the haemostatic process. PMID- 11103185 TI - Anaesthesia with propofol decreases FMLP-induced neutrophil respiratory burst but not phagocytosis compared with isoflurane. AB - Propofol has been reported to produce a dose-dependent inhibition of phagocytosis and superoxide anion production during the respiratory burst (RB) of polymorphonuclear cells (PMNs) in vitro. In this randomized, blinded study, these two parameters were compared during propofol or isoflurane anaesthesia in patients undergoing elective interventional embolization of cerebral arterio venous malformations. Anaesthesia was performed with continuous intravenous propofol 6-8 mg kg-1 h-1 (n = 15) or isoflurane 0.8-1.0% end tidal (n = 15). Heparinized blood was drawn before, and 2 and 4 h after induction of anaesthesia. The RB in isolated leucocytes was measured with the fluorescent dye rhodamine after ex vivo induction by Escherichia coli or tumour necrosis factor alpha/N formyl-methionyl-leucylphenylalanine (TNF-alpha/FMLP). Phagocytosis was carried out in whole blood after incubation with fluorescein isothiocyanate (FITC) labelled, opsonized E. coli and also measured with a flow cytometer. The two groups were similar in terms of biometric data and haemodynamic responsiveness. After 4 h of propofol or isoflurane anesthesia, the mean (SD) phagocytosis of E. coli was 93.2% (7.0%) and 94.3% (9.2%), respectively, of that before anaesthesia. The percentage of PMN with RB activity following TNF-alpha/FMLP stimulation was significantly reduced after 2 h (80.9% (24.2%); P < 0.05) and 4 h (53.7% (27.3); P < 0.05) of anaesthesia with propofol compared with the values before induction. This effect of propofol anaesthesia was significantly different from the effect of isoflurane anaesthesia. In contrast to published in vitro results, 4 h of anaesthesia with propofol did not reduce the phagocytotic capacity of human blood PMN more than isoflurane anaesthesia. PMID- 11103186 TI - Somatosensory evoked potentials for closed-loop control of anaesthetic depth using propofol in the urethane-anaesthetized rat. AB - Primary somatosensory cortical mass responses have been shown to exhibit dose dependent changes in latency when general anaesthetics are administered. Here we describe a system in which the latency of evoked responses was measured automatically in real time in five animals. Latency changes were used to operate a closed-loop control of propofol delivery by intravenous infusion. The system attempted to induce and maintain a 1 ms increase in evoked response latency; this was reversed when infusion was discontinued. Allowing for the rapid and large biological fluctuations in the evoked response, this was achieved successfully. The system maintained a mean increase in latency of 1.27 (SD 0.42) ms. The mean statistical dispersion index of data obtained during the controlled period was 1.23 (0.3); in an ideal controllable system it approximates to 1. Such a system may provide a means for the automatic delivery of anaesthetics. PMID- 11103187 TI - Effects of sevoflurane on hypoxic pulmonary vasoconstriction in anaesthetized piglets. AB - In vitro, halogenated agents reduce the pulmonary vasoconstrictor response to alveolar hypoxia in isolated perfused lungs. However, studies in intact animals have been less convincing. The aim of the present study was to assess the effect of sevoflurane on hypoxic pulmonary vasoconstriction (HPV) in anaesthetized piglets using the pressure/cardiac index relationship (P/Q). Ten large white piglets were anaesthetized and mechanically ventilated, alternately in hyperoxia (FIO2 = 0.4) and hypoxia (FIO2 = 0.12). Multipoint plots of pulmonary arterial pressure (PAP) or differences between PAP and left atrial pressure (LAP) against Q were generated by gradual inflation of a balloon introduced into the inferior vena cava. P/Q relationships were established in hyperoxia and hypoxia at baseline, and then with sevoflurane. In hypoxia, pressure gradients (PAP-LAP) increased at every level of Q, thus demonstrating active pulmonary vasoconstriction. Sevoflurane at 1 MAC did not affect these P/Q relationships in hyperoxia or hypoxia as compared with baseline. Sevoflurane at a clinically relevant concentration (1 MAC) has no significant effect on HPV in anaesthetized piglets. PMID- 11103188 TI - Peri-operative management of patients with coagulation disorders. PMID- 11103189 TI - Within-breath arterial PO2 oscillations in an experimental model of acute respiratory distress syndrome. AB - Tidal ventilation causes within-breath oscillations in alveolar oxygen concentration, with an amplitude which depends on the prevailing ventilator settings. These alveolar oxygen oscillations are transmitted to arterial oxygen tension, PaO2, but with an amplitude which now depends upon the magnitude of venous admixture or true shunt, QS/QT. We investigated the effect of positive end expiratory pressure (PEEP) on the amplitude of the PaO2 oscillations, using an atelectasis model of shunt. Blood PaO2 was measured on-line with an intravascular PaO2 sensor, which had a 2-4 s response time (10-90%). The magnitude of the time varying PaO2 oscillation was titrated against applied PEEP while tidal volume, respiratory rate and inspired oxygen concentration were kept constant. The amplitude of the PaO2 oscillation, delta PaO2, and the mean PaO2 value varied with the level of PEEP applied. At zero PEEP, both the amplitude and the mean were at their lowest values. As PEEP was increased to 1.5 kPa, both delta PaO2 and the mean PaO2 increased to a maximum. Thereafter, the mean PaO2 increased but delta PaO2 decreased. Clear oscillations of PaO2 were seen even at the lowest mean PaO2, 9.5 kPa. Conventional respiratory models of venous admixture predict that these PaO2 oscillations will be reduced by the steep part of the oxyhaemoglobin dissociation curve if a constant pulmonary shunt exists throughout the whole respiratory cycle. The facts that the PaO2 oscillations occurred at all mean PaO2 values and that their amplitude increased with increasing PEEP suggest that QS/QT, in the atelectasis model, varies between end-expiration and end inspiration, having a much lower value during inspiration than during expiration. PMID- 11103190 TI - Spasmolytic effects of prostaglandin E1 on serotonin-induced bronchoconstriction and pulmonary hypertension in dogs. AB - In this study, we simultaneously evaluated the spasmolytic effects of prostaglandin E1 (PGE1) on serotonin-induced bronchoconstriction and pulmonary hypertension. Eleven mongrel dogs (8-12 kg) anaesthetized with pentobarbital were assigned to two groups: saline (n = 4) and PGE1 (n = 7). Bronchoconstriction and pulmonary hypertension were elicited with serotonin 10 micrograms kg-1 + 1 mg kg 1 h-1 and assessed as the percentage change in bronchial cross-sectional area (BCA) measured by bronchoscopy and pulmonary vascular resistance (PVR), respectively. Thirty minutes after starting the serotonin infusion, saline or PGE1 0 (saline), 0.01, 0.1, 1.0 or 10 micrograms kg-1 i.v. was given. %BCA and %PVR (basal = 100%) were assessed before and 30 min after serotonin, and 30 and 60 min after saline (saline group) or 5 min after each dose of PGE1 (PGE1 group). In the saline group, pulmonary hypertension and bronchoconstriction were stable. In the PGE1 group, PGE1 at > or = 0.1 microgram kg-1 significantly decreased %BCA and 10 micrograms kg-1 almost fully reversed the constriction (from mean (SEM) 56.2% (4.9%) to 94.4% (3.7%)). %PVR was significantly decreased at 10 micrograms kg-1 (from 230% (24%) to 176% (11%)) only. We suggest that PGE1 may produce bronchodilation rather than pulmonary vasodilation. PMID- 11103191 TI - Influence of airway-occluding instruments on airway pressure during jet ventilation for rigid bronchoscopy. AB - We measured changes in airway pressure (Paw) caused by microsurgical instruments introduced into a rigid bronchoscope during high frequency jet ventilation (HFJV). With approval of the institutional Ethics Committee, 10 adults undergoing elective tracheobronchial endoscopy and endosonography during general anaesthesia were investigated. Inflation of an endosonography probe balloon in the left main stem bronchus caused airway obstruction. Pressure measurements proximal and distal to the obstruction were compared after three degrees of obstruction (0%, 50% and 90%) and with two different driving pressure settings. Airway obstruction increased the mean (SD) peak inspiratory pressure (PIP) from 7.5 (2.6) to 9.5 (3.5) mm Hg for 2 atm (P = 0.0008) and from 9.7 (3.7) to 13.0 (5.1) mm Hg for 3 atm (P = 0.0001). Airway obstruction did not alter peripheral PIP (7.2 (4.1) to 7.1 (3.7) mm Hg for 2 atm and 8.8 (4.3) to 9.4 (5.2) mm for 3 atm), but resulted in an end-expiratory pressure (EEP) beyond the narrowing being significantly greater than in the unobstructed airway (2.5 (3.4) to 5.5 (3.7) mm Hg for 2 atm; P = 0.0005) and 3.2 (3.6) to 8.0 (4.3) mm for 3 atm; P < 0.0001). Severe airway narrowing increases inspiratory pressure proximal and expiratory pressure distal to the obstruction in relation to the applied driving pressure. Since the distal EEP never exceeded PIP, even near-total airway obstruction should not cause severe lung distension or barotrauma in subjects with normal lungs. PMID- 11103192 TI - Transcranial magnetic-evoked potentials under total intravenous anaesthesia and nitrous oxide. AB - Magnetic stimulation of the cortex and recording of the motor-evoked potentials (MEPs) by electromyography (EMG) is a well proven method to assess the descending pathways of the spinal cord and detect neurological impairment. We have assessed, in 33 adult patients undergoing spinal surgery, the influence of four total i.v. anaesthesia regimens (TIVA) on this recording technique. In 20 patients, the effect of 50% nitrous oxide was also studied. MEP amplitudes, latencies and success rates of stimulation were obtained in the steady-state after induction of anaesthesia. Combinations of midazolam and ketamine, and alfentanil and etomidate had the least effect on MEPs. Propofol (in combination with alfentanil or ketamine) showed marked depression of the MEP amplitude and the lowest success rates of stimulation. The latencies did not change at all. The addition of nitrous oxide significantly depressed the registered MEPs and lowered the success rates. PMID- 11103193 TI - Motor block during patient-controlled epidural analgesia with ropivacaine or ropivacaine/fentanyl after intrathecal bupivacaine for caesarean section. AB - We compared patient-controlled epidural analgesia (PCEA) with ropivacaine alone or combined with fentanyl in terms of analgesic efficacy, motor weakness and side effects in patients who had received spinal anaesthesia for elective Caesarean section. ASA I patients received combined spinal-epidural anaesthesia and were randomly assigned, in a double-blind study, into two groups after operation: group R (n = 23) received PCEA ropivacaine 0.1%, bolus 5 mg, lock-out 15 min, 3 mg h-1 background infusion, and group RF (n = 24) received PCEA 0.1% ropivacaine/fentanyl 2 micrograms ml-1 at identical settings. Pain and satisfaction on a 100 mm visual analogue scale (VAS) and side-effects were noted. Incidence of motor weakness (Bromage grade 1 or higher) was 48% (11/23) at 8 h in group R compared with 13% (3/24) in group RF (P = 0.025). Pain scores on movement were lower in group RF at 8 and 12 h and at rest at 6 and 8 h (P < 0.05 for each comparison). Analgesic consumption was less in RF (P = 0.041), but there was no difference in time to first request for supplementary analgesia. Patient satisfaction with postoperative analgesia (mean (SD)) was higher in RF (79 (23) vs 57 (29) mm, P = 0.045). Caution should be exercised using ropivacaine PCEA after spinal bupivacaine for Caesarean section, because its reputed motor-sparing property may be unreliable. PMID- 11103194 TI - Repetitive synchronized cyclical oscillations of multisystem parameters subsequent to high-dose thiopental therapy for status epilepticus secondary to herpes encephalitis. AB - We report a case of status epilepticus secondary to herpes encephalitis, treated with thiopental infusion and mechanical ventilation. The computerized storage and analysis of physiological data led to the detection of repetitive synchronized cyclical oscillations of arterial pressure, heart rate, EEG parameters, peripheral temperature and core temperature. Arterial pressure oscillations have been described in patients who are severely systemically unwell; cardiovascular and brain electrical activity may also oscillate in the presence of raised intracranial pressure. In contrast, this patient had no features of severe systemic illness or of raised intracranial pressure. Our hypothesis is that high dose thiopental may have been a cause of our findings by producing autonomic dysfunction. PMID- 11103195 TI - Loss of consciousness following spinal anaesthesia for caesarean section. AB - A healthy parturient under spinal anaesthesia for Caesarean section lost consciousness for an hour, 20 min after the intrathecal injection of 2 ml of 0.5% heavy bupivacaine. The patient was haemodynamically stable before losing consciousness. The differential diagnosis is discussed. PMID- 11103196 TI - Traumatic bilateral internal carotid artery dissection following airbag deployment in a patient with fibromuscular dysplasia. AB - This case describes a 39-yr-old male, presenting with left hemiplegia after a road traffic accident involving frontal deceleration and airbag deployment. Brain computerized tomography (CT) scan revealed a right parietal lobe infarct. Contrast angiography demonstrated bilateral internal carotid artery dissection and fibromuscular dysplasia. The patient was treated with systemic heparinization. Neurological improvement, evidenced by full return of touch sensation, proprioception and nociception began 10 days after the injury. To our knowledge, this is the first case report of carotid artery dissection associated with airbag deployment. Forced neck extension in such settings may result in carotid artery dissection because of shear force injury at the junction of the extracranial and intrapetrous segments of the vessel. Clinicians should consider carotid artery injury when deterioration in neurological status occurs after airbag deployment. We propose that the risk of carotid artery dissection was increased by the presence of fibromuscular dysplasia. PMID- 11103197 TI - Anaesthetic management of a pregnant patient in a persistent vegetative state. AB - Pregnancy in a patient in a persistent vegetative state presents challenging therapeutic questions about the level of supportive management required, the assessment of fetal well-being, the timing and mode of delivery and the anaesthetic management of labour and delivery. We report the case of a 29-yr-old woman who had a favourable fetal outcome despite suffering hypoxic brain damage after a suicide attempt by a drug overdose. She was managed until the onset of labour on an intensive care unit and had a spontaneous vaginal delivery assisted by epidural anaesthesia. PMID- 11103198 TI - The Reflector: a new method for saving anaesthetic vapours. AB - Anaesthesia systems that minimize the use of volatile anaesthetics to reduce cost and pollution are of interest. Closed circuit anaesthesia is the ideal solution, but requires continuous adjustment of fresh gas flow and composition and thus is demanding in routine practice. We describe an alternative system, the Reflector system, which is open in regard to oxygen, nitrogen and N2O, and semiclosed in regard to volatile anaesthetics. The Reflector system is a circle system with a carbon dioxide absorber and an automatic vapour delivery device placed in the inspiratory limb of the circle. A zeolite filter, the Reflector, is placed between the ventilator and the circle. The Reflector functions as a molecular sieve, preventing the volatile anaesthetic from leaving the circle. Isoflurane consumption using the Reflector system in bench tests and an animal study was compared with that of an open system. In bench tests consumption was reduced by 79% and 82%, at a respiratory frequency of 10 and 20 min-1, respectively. The corresponding mean figures from the animal experiment were 65% and 77%. PMID- 11103199 TI - Management of massive blood loss: a template guideline. AB - The management of acute massive blood loss is considered and a template guideline is formulated, supported by a review of the key literature and current evidence. It is emphasized that, if avoidable deaths are to be prevented, surgeons, anaesthetists, haematologists and blood-bank staff need to communicate closely in order to achieve the goals of secure haemostasis, restoration of circulating volume, and effective management of blood component replacement. PMID- 11103200 TI - General versus regional anaesthesia for hip fracture surgery. PMID- 11103201 TI - Off-pump revascularization and the brain. PMID- 11103202 TI - Off-pump revascularization and the brain. PMID- 11103203 TI - Intubating laryngeal mask airway (ILMA) seems to be an ideal device for blind intubation in case of immobile spine. PMID- 11103204 TI - Preparation for regional anaesthesia induces changes in thromboelastography. PMID- 11103205 TI - Epidural top-ups solutions for emergency caesarean section. PMID- 11103206 TI - Postoperative orphenadrine withdrawal. PMID- 11103207 TI - Protecting anaesthetic tubing from occlusion: an inbuilt solution. PMID- 11103208 TI - Headaches in the prenatal period. PMID- 11103209 TI - Dietary fatty acids, cholesterol, and the lipoprotein profile. PMID- 11103210 TI - Orexins, feeding and the big picture. PMID- 11103211 TI - The use of visual analogue scales to assess motivation to eat in human subjects: a review of their reliability and validity with an evaluation of new hand-held computerized systems for temporal tracking of appetite ratings. AB - This present paper reviews the reliability and validity of visual analogue scales (VAS) in terms of (1) their ability to predict feeding behaviour, (2) their sensitivity to experimental manipulations, and (3) their reproducibility. VAS correlate with, but do not reliably predict, energy intake to the extent that they could be used as a proxy of energy intake. They do predict meal initiation in subjects eating their normal diets in their normal environment. Under laboratory conditions, subjectively rated motivation to eat using VAS is sensitive to experimental manipulations and has been found to be reproducible in relation to those experimental regimens. Other work has found them not to be reproducible in relation to repeated protocols. On balance, it would appear, in as much as it is possible to quantify, that VAS exhibit a good degree of within subject reliability and validity in that they predict with reasonable certainty, meal initiation and amount eaten, and are sensitive to experimental manipulations. This reliability and validity appears more pronounced under the controlled (but more artificial) conditions of the laboratory where the signal:noise ratio in experiments appears to be elevated relative to real life. It appears that VAS are best used in within-subject, repeated-measures designs where the effect of different treatments can be compared under similar circumstances. They are best used in conjunction with other measures (e.g. feeding behaviour, changes in plasma metabolites) rather than as proxies for these variables. New hand-held electronic appetite rating systems (EARS) have been developed to increase reliability of data capture and decrease investigator workload. Recent studies have compared these with traditional pen and paper (P&P) VAS. The EARS have been found to be sensitive to experimental manipulations and reproducible relative to P&P. However, subjects appear to exhibit a significantly more constrained use of the scale when using the EARS relative to the P&P. For this reason it is recommended that the two techniques are not used interchangeably. PMID- 11103212 TI - The role of high-fat diets and physical activity in the regulation of body weight. AB - The prevalence of obesity is increasing in westernized societies. In the USA the age-adjusted prevalence of BMI > 30 kg/m2 increased between 1960 and 1994 from 13% to 23% for people over 20 years of age. This increase in the prevalence of obesity has been attributed to an increased fat intake and a decreased physical activity. However, the role of the impact of the level of dietary fat intake on human obesity has been challenged. High-fat diets, due to their high energy density, stimulate voluntary energy intake. An increased fat intake does not stimulate its own oxidation but the fat is stored in the human body. When diet composition is isoenergetically switched from low to high fat, fat oxidation only slowly increases, resulting in positive fat balances on the short term. Together with a diminished fat oxidation capacity in pre-obese subjects, high-fat diets can therefore be considered to be fattening. Another environmental factor which could explain the increasing prevalence of obesity is a decrease in physical activity. The percentage of body fat is negatively associated with physical activity and exercise has pronounced effects on energy expenditure and substrate oxidation. High-intensity exercise, due to a lowering of glycogen stores, can lead to a rapid increase in fat oxidation, which could compensate for the consumption of high-fat diets in westernized societies. Although the consumption of high-fat diets and low physical activity will easily lead to the development of obesity, there is still considerable inter-individual variability in body composition in individuals on similar diets. This can be attributed to the genetic background, and some candidate genes have been discovered recently. Both leptin and uncoupling protein have been suggested to play a role in the prevention of diet-induced obesity. Indeed, leptin levels are increased on a high fat diet but this effect can be attributed to the increased fat mass observed on the high-fat diet. No effect of a high-fat diet per se on leptin levels is observed. Uncoupling proteins are increased by high-fat diets in rats but no data are available in human subjects yet. In conclusion, the increased intake of dietary fat and a decreasing physical activity level are the most important environmental factors explaining the increased prevalence of obesity in westernized societies. PMID- 11103213 TI - Intestinal metabolism of rye lignans in pigs. AB - To study the intestinal metabolism of lignans, the concentrations of plant and mammalian lignans in intestinal digesta sampled along the intestinal tract of pigs were determined by isotope dilution GC-MS. The pigs were fed rye-bread diets made from either whole rye-grains or rye-grain milling fractions enriched in pericarp-testa, aleurone or endosperm cells. The content and characteristics of dietary fibre varied between diets and had been shown to induce different colon fermentation patterns. As the metabolism of lignans depends on the action of the intestinal flora, we tested whether the rye-bread diets influence the metabolism of lignans. In the ileum, the lignans were mainly present as conjugated plant lignans, which were determined only when the analytical procedure included a hydrolysis step. High recovery of dietary lignans in the ileum may indicate that the lignans enter the enterohepatic circulation. In addition, two to three times the intake of lignans were recovered in the faeces when the diets had a high content of dietary fibre suggesting underestimation of plant lignans in the diet. Most of the plant lignans disappeared from the intestinal tract between the terminal ileum and the caecum. The intestinal concentrations and the disappearance of lignans correlated with the content of lignans in the diet, being highest on the pericarp-testa diet and lowest on the endosperm diet. No effect of fermentation pattern on the intestinal metabolism of lignans was observed. The lignans were liberated from the pericarp-testa diet although the plant cell walls remained largely undegraded. PMID- 11103214 TI - Interactive effects of dietary cholesterol and different saturated fatty acids on lipoprotein metabolism in the hamster. AB - The present study examines the interactive effects of three fatty acids: myristic, palmitic and stearic acids, with dietary cholesterol, on lipoprotein metabolism in the hamster. Each saturated fatty acid was fed at a concentration of 100 g pure synthetic triacylglycerol/kg in the presence of 100 g triolein/kg and was fed in the presence of 0.05, 1.2 or 2.4 g dietary cholesterol/kg. Dietary cholesterol increased the concentration of cholesterol in each of the major plasma lipoprotein fractions. The largest effects on VLDL and LDL were seen in the presence of tripalmitin where the increase between the lowest and highest dietary cholesterol groups were 129% and 38% respectively. In contrast, HDL showed the greatest change in the tristearin group when the equivalent increase was 59%. No interactive effects of dietary cholesterol and fat were seen on hepatic mRNA concentrations for the LDL receptor, hydroxymethylglutaryl-CoA reductase or the microsomal triacylglycerol transfer protein. As the amount of cholesterol in the diet increased, large differences were seen in the storage of hepatic cholesterol ester. At the highest dietary cholesterol intake the amount of hepatic cholesterol ester was 1.7-fold higher in the animals fed trimyristin compared with those fed tripalmitin. These results suggest that, as the amount of cholesterol in the diet is increased, palmitic acid becomes more hypercholesterolaemic. This is associated with a reduced ability to store cholesterol ester in the liver. PMID- 11103215 TI - Quantification of the absorption of nutrients derived from carbohydrate assimilation: model experiment with catheterised pigs fed on wheat- or oat-based rolls. AB - The main purpose of this study was to quantify the absorption of nutrients derived from carbohydrate assimilation in a model experiment with catheterised pigs. A low-fibre (LF) diet based on wheat flour and two high-fibre diets with added insoluble fibre from wheat bran (HFWB) or soluble fibre from oat bran (HFOB) were used. The diets were offered as baked rolls to three catheterised pigs in a 3 x 3 Latin square design. The pigs were surgically fitted with catheters placed in the portal vein and mesenteric artery and with an ultrasonic flow probe attached to the portal vein to monitor the blood-flow rate. The pigs were fed the diets three times daily and portal and arterial blood samples collected twice weekly up to 8 h after the morning feeding. Glucose, insulin, lactic acid (LA) and short-chain fatty acids (SCFA) were determined on the samples. The baseline level of glucose in the portal vein was about 6 mmol/l increasing to 10-11 mmol/l 20-30 min post-feeding with no difference among the different diets. Portal and arterial insulin mirrored portal glucose concentration and was also unaffected by the dietary composition. The net absorption of glucose (per 24 h) was: diet LF 4190 mmol; diet HFWB 3050 mmol and diet HFOB 3190 mmol corresponding to a recovery of 0.76-0.92 of ingested starch. The levels of LA and SCFA in the portal vein were relatively constant in the postprandial period. The net absorption of LA and SCFA was in the same order (749 and 720 mmol/d respectively) with diet LF, while LA was lower (384 and 582 mmol/d) and SCFA higher (738 to 891 mmol/d) when feeding the two high-fibre diets. There was a higher molar proportion of butyrate in the portal vein after feeding the high-fibre diet supplemented with oat bran as compared with the wheat based diets. PMID- 11103216 TI - Effects of diet quality on urea fates in sheep as assessed by refined, non invasive [15N15N]urea kinetics. AB - The effect of diet quality on urea production, entry into the gastrointestinal tract (GIT) and subsequent diversion to anabolic or catabolic fates was examined in four sheep (mean live weight 49.5 kg). The animals received, in a crossover design, each of two rations, hay-grass pellets (1:1 HG) and a mixed concentrate forage (CF). Measurements were made of N balance and urea kinetics based on a 4 d continuous intravascular infusion of [15N15N]urea. Enrichments of [15N15N]- and [14N15N]urea in the urine, and faecal 15N content were determined each day. After 24 h of infusion, urinary [15N15N]urea enrichments reached constant enrichment but a further 24 h was required before [14N15N]urea enrichment was at plateau. The latter is derived from hydrolysis of urea to 15NH3 in the digestive tract with subsequent absorption and reconversion to urea. The diets were not isonitrogenous (14.3 v. 17.1 g N supplied daily for HG and CF respectively) but showed no difference in N balance. Urea-N production was much greater (16.3 v. 11.1 g/d; P = 0.011) for CF compared with HG and more urea-N entered the GIT (9.9 v. 7.7; P = 0.07). A larger proportion of GIT entry was returned to ureagenesis (51 v. 42%; P = 0.047) for the CF diet but a smaller fraction was lost in the faeces (3.3% v. 7.1%; P = 0.013). In consequence, most of the additional urea-N which entered the GIT on the CF diet was returned to the ornithine cycle (probably as NH3) and the absolute amount available for anabolic purposes was similar between the rations (3.9 v. 4.5 g N/d). PMID- 11103217 TI - Soluble polysaccharide and biomass of red microalga Porphyridium sp. alter intestinal morphology and reduce serum cholesterol in rats. AB - The present study investigated the effects of the red microalga Porphyridium sp. on gastrointestinal physiology and lipid metabolism in male Sprague-Dawley rats. Diets containing dietary fibre from pelleted red microalgal cells (biomass) or their sulfated polysaccharide, pectin or cellulose (control) were fed to rats for a period of 30 d. All three fibre-supplemented diets increased the length of both the small intestine and colon, with a significantly greater effect in rats fed the algal polysaccharide. The polysaccharide also increased mucosa and muscularis cross-sectional area of the jejunum, and caused hypertrophy in the muscularis layer. The algal biomass significantly lowered gastrointestinal transit time by 44% in comparison with the control rats. Serum and mucosal cholecystokinin levels were lower in rats on the pectin and polysaccharide diets, while cholecystokinin levels in rats fed algal biomass were not different from those in the control animals. In comparison with the control diet, all the experimental diets significantly lowered serum cholesterol levels (22-29%). Feeding of non fermentable algal polysaccharide or biomass significantly increased faecal weight and bile acid excretion compared with pectin-fed or control rats. The algal polysaccharide and biomass were thus shown to be potent hypocholesterolaemic agents active at low concentrations in the diet. Both metabolic and morphological changes were observed following consumption of algae, suggesting several possible mechanisms by which the alga affects lipid metabolism. The results presented in the present study encourage the use of red microalga as a functional food. PMID- 11103218 TI - Portal recovery of short-chain fatty acids infused into the temporarily-isolated and washed reticulo-rumen of sheep. AB - The present study was undertaken to study the metabolism of short-chain fatty acids (SCFA) by the reticulo-ruminal epithelium and the portal-drained viscera (PDV) under in vivo conditions with no interference from the metabolism of the rumen microbes. The technique of temporary isolation of the reticulo-rumen was applied to wethers implanted with catheters in a mesenteric artery, the hepatic portal vein and the right ruminal vein. Portal blood flow was measured by downstream dilution of p-aminohippuric acid; the PDV uptake of arterial acetate, as well as the whole-body irreversible loss rate (ILR) of acetate, was estimated by [2-(13)C]acetate infusion into the right ruminal vein. The sheep were maintained with a bicarbonate-buffered solution of SCFA in the reticulo-rumen along with continuous intraruminal infusion of SCFA for 4 h. The portal appearance of SCFA of non-reticulo-ruminal origin was estimated before and after the infusion protocol. Of the acetate absorbed by the sheep, 89 (SE 5), 109 (SE 7) and 101 (SE 7)% was recovered as portal net appearance of acetate, portal net appearance of acetate corrected for PDV uptake of arterial acetate and increase in the ILR of acetate respectively. Of the propionate, isobutyrate, butyrate, isovalerate and valerate absorbed by the sheep, 95 (SE 7), 102 (SE 9), 23 (SE 3), 48 (SE 5) and 32 (SE 4)% respectively was recovered as portal net appearance. In contrast to current concepts, the present study showed that the reticulo-ruminal epithelium metabolizes none (or only a small proportion) of the acetate and propionate absorbed from the rumen. This observation could lead to the more efficient use of results obtained with multi-catheterized animals to quantify the net metabolite output of the rumen microbes. PMID- 11103219 TI - Portal net appearance of amino acids in growing pigs fed a barley-based diet with inclusion of three different forage meals. AB - The net absorption of amino acids (AA) in young pigs fed a barley-based control diet (C) and diets where barley was replaced by 200 g/kg fresh weight of dried lucerne (Medicago sativa; L20), white clover (Trifolium repens; W20) or perennial ryegrass (Lolium perenne; PR20) meal was studied. Castrated male pigs were fitted with permanent catheters in the hepatic portal vein and mesenteric artery, and the hepatic portal net absorption of AA was estimated from the porto-arterial plasma concentration differences and the hepatic portal-vein blood flow. In general, the essential AA (EAA) concentrations in the hepatic portal vein reached peak levels 90 min after feeding and thereafter exhibited a transient decline. Maximum porto-arterial differences were reached between 1 and 3 h postprandially for most of the AA. The cumulative net absorption of non-essential AA (NEAA) and EAA did not differ significantly between the barley-based diet and diets W20 and PR20. Due to a lower intake of AA on diet L20, the cumulative net absorption of NEAA and EAA was significantly (P < 0.05) lower than diet C. With the exceptions of the EAA arginine, cystine and valine, and the NEAA glutamic acid + glutamine and glycine, there were no significant differences in the absorption coefficients for the EAA and NEAA between the diets. In addition, the pattern of the total EAA in the mixture absorbed postprandially did not differ significantly between the diets. The present study gives support to the contention that the replacement of barley AA with forage meal AA in a barley-based diet for growing pigs should be expected to result in minor differences in the net portal flux of AA. PMID- 11103221 TI - The effect of grape-skin extract on oxidative status. AB - Epidemiological studies indicate that moderate alcohol consumption, particularly wine, reduce the risk of CHD. The present study was designed to investigate the effect of grape-skin extract on markers of oxidative status. The study was designed as a randomised crossover. A diet with a low content of flavonoids was served with strict control of intake in two consecutive 1-week intervention periods to fifteen subjects (nine women, six men) divided randomly into two groups. During one of the weeks the subjects from either group consumed 200 ml grape-skin extract in water (1 mg extract/ml) at each of three daily meals (31.3 mg total phenolics, including 9.0 mg catechin). An increased activity of glutathione reductase and a borderline increase of glutathione peroxidase activity in erythrocytes were observed after grape-skin intervention, while the intervention had no significant effect on superoxide dismutase or catalase. Likewise, no effect was found on 2-aminoadipic semialdehyde (AAS) residues, a plasma protein oxidation product, or on malondialdehyde in plasma or in LDL, which are markers of lipoprotein oxidation. A marginal effect of grape-skin intervention was observed on plasma ascorbate levels. Intake of the experimental diet significantly reduced plasma vitamin C and plasma AAS in both groups. This effect was most pronounced in the particular week with no grape-skin extract addition. We speculate that grape-skin extract may have a sparing effect on vitamin C. The effects of the experimental diet may be partly ascribed to a low content of several fruit- and vegetable-related antioxidants like flavonoids and vitamin C and a relatively high content of carrot-derived antioxidants, such as carotenes. PMID- 11103220 TI - Dietary calcium and phosphate restriction in guinea-pigs during pregnancy: fetal mineralization induces maternal hypocalcaemia despite increased 1 alpha,25 dihydroxycholecalciferol concentrations. AB - Guinea-pig fetuses at term are mineralized to a degree comparable with human fetuses, which makes the guinea-pig an attractive animal model to study maternal fetal interactions with regard to Ca and phosphate (P) homeostasis. We studied non-pregnant and pregnant (day 57) vitamin D-replete guinea-pigs, fed either a normal guinea-pig chow with 9.6 g Ca/kg and 4.9 g P/kg or a study diet with 2 g Ca/kg and 1 g P/kg (low-Ca-P diet) for 7-8 weeks. Both pregnancy and the low-Ca-P diet decreased plasma concentrations of 25-hydroxycholecalciferol (25(OH)D3), but increased total and free 1 alpha,25-dihydroxycholecalciferol (1,25(OH)2D3), strongly suggesting an additive stimulation of 1 alpha-hydroxylase activity. Maternal and fetal 25(OH)D3 and 1,25(OH)2D3 levels were highly correlated (r 0.82 and 0.92 respectively, P < 0.001). Dual-energy absorption X-ray absorptiometry (DXA) showed that both pregnancy and the low-Ca-P diet decreased bone mineral density (BMD) of the maternal femur, particularly at the distal metaphysis. Despite higher 1,25(OH)2D3 concentrations and lower BMD, pregnant animals on the low-Ca-P diet were hypocalcaemic; blood Ca2+ levels were inversely correlated with the number of fetuses in this group (r -0.93, P < 0.001). Fetal growth as well as mineralization (assessed by whole-body and femoral DXA, bone histomorphometry and plasma-bone osteocalcin measurements) were unaltered in the low-Ca-P group. In conclusion, fetal mineralization proceeds normally but induces maternal hypocalcaemia in guinea-pigs with dietary restriction of Ca and P. PMID- 11103222 TI - Resting metabolic rate, fat-free mass and catecholamine excretion during weight loss in female obese patients. AB - The reduction in resting metabolic rate (RMR) during weight loss exceeds that accounted for by changes in body composition by 15%, suggesting that factors other than fat-free mass (FFM) explain the metabolic adaptation during food restriction in obesity. Our study aimed to establish if changes in the sympathoadrenal system activity, as inferred from an integrated measure such as 24 h urinary excretion of catecholamines, may play a role in the RMR adaptation observed during dietary restriction in obese patients. Ninety-three obese female subjects consumed a low-energy diet (LED) (2930 kJ/d (700 kcal/d)) for a 3-week period. At the beginning and at the end of the study, 24 h urinary excretion of catecholamines, FFM and RMR were measured. The LED induced a significant reduction in body weight (-3.3 (SEM 0.4) kg; P < 0.01), FFM (-1.9 (SEM 0.7) kg; P < 0.01) and in the fat mass (-1.2 (SEM 0.5) kg; P < 0.01). Noradrenalin excretion (24 h) decreased during the LED from 264 (SEM 26) during a weight-maintenance period to 171 (SEM 19) nmol/24 h after consumption of the LED for 3 weeks (P < 0.001); mean 24 h adrenalin excretion did not change during the LED (22 (SEM 3) during the weight-maintenance period v. 21 (SEM 3) nmol/24 h after consumption of the LED for 3 weeks; NS). The LED induced a significant decrease in RMR (7300 (SEM 218) v. 6831 (SEM 138) kJ/24 h; P < 0.001). The only independent variable that significantly explained variations in RMR both before and after consumption of the LED for 3 weeks, was FFM (r2 0.79 and r2 0.80 respectively). Urinary noradrenalin excretion explained a further 4% of the variability in RMR, but only before the diet, so that a role of sympathoadrenal system on RMR seems to be present in obese patients in basal conditions but not at the end of the LED. PMID- 11103223 TI - Comparison of high-fat and high-carbohydrate foods in a meal or snack on short term fat and energy intakes in obese women. AB - The present study aimed to compare the action of high-fat and high-carbohydrate (CHO) foods on meal size (satiation) and post-meal satiety in obese women. A within-subjects design was used; each participant received all four nutritional challenges. Fifteen healthy obese women (age 21-56 years, BMI 35-48 kg/m2) participated; thirteen completed all four test days. On two test days, participants were exposed to a nutritional challenge comprising an ad libitum high-fat or high-CHO lunch. On the other two test days they were exposed to a challenge comprising an ad libitum sweet high-fat or high-CHO mid-afternoon snack. Energy and macronutrient intakes were measured at each eating episode. Visual analogue rating scales were completed periodically to record subjective feelings of appetite. When offered a high-CHO selection of foods at lunch and mid afternoon participants consumed less energy than when offered a high-fat selection. However, post-meal satiety was similar. Total test-day energy intake was significantly higher when high-fat foods were consumed at lunch, but not as a snack. Consumption of high-fat foods at a lunch and snack increased the amount of fat consumed over the whole test day. In conclusion, energy intake of an eating episode was influenced by nutrient composition in this group of obese women. Consumption of high-fat foods at lunch or as a snack led to overconsumption relative to high-CHO foods. However, high-fat foods at meals may have greater potential to influence daily intake than at snacks, probably because meals are larger eating episodes and therefore give greater opportunity to overconsume. PMID- 11103225 TI - Tracking of nutrient intakes in adolescence: the experiences of the Young Hearts Project, Northern Ireland. AB - This study evaluated the tracking of energy and nutrient intakes, assessed by diet history, in a random sample of adolescents (boys n 225, girls n 230) at baseline (age 12 years), and subsequently at age 15 years. Median energy (MJ/d) and macronutrient (g/d) intakes increased significantly (all P < 0.001) with increasing age in the boys. The girls' reported energy intake (MJ/d) remained stable over time, despite significant increases in BMI, weight and % body fat. Age-related changes in the girls' macronutrient intakes were inconsistent. When expressed in terms of nutrient density, the diets of both sexes became significantly richer, over time, in total folate (both sexes, P < 0.01), but poorer in Ca (boys P < 0.01, girls P < 0.001) and riboflavin (both sexes P < 0.001). Vitamin B6 (P < 0.001) and Fe (P < 0.05) densities increased in the boys, while the thiamin density of the girls' diets decreased (P < 0.001). Tracking, defined as maintenance of rank over time, was summarised using weighted kappa statistics (kappa). There were some significant changes in intakes at the group level; however, tracking of energy and nutrients in both sexes was only poor to fair (kappa < 0.40), indicating substantial drift of individuals between classes of intake over time. Particularly poor tracking was evident for % energy from sugars (kappa 0.09) and total fat (kappa 0.09) in the girls' diets. In conclusion, the poor to fair tracking observed in this cohort suggests that individual dietary patterns exhibited at 12 years of age are unlikely to be predictive of energy and nutrient intake at age 15 years. PMID- 11103224 TI - Components and variations in daily energy expenditure of athletic and non athletic adolescents in free-living conditions. AB - The objectives of the study were to determine: (1) daily energy expenditure (EE) of athletic and non-athletic adolescents of both sexes in free-living conditions; (2) day-to-day variations in daily EE during 1 week; (3) energy costs of the main activities; and (4) the effect of usual activity on EE during sleep, seated and miscellaneous activities. Fifty adolescents (four groups of eleven to fifteen boys or girls aged 16-19 years) participated in the study. Body composition was measured by the skinfold-thickness method, and VO2max and external mechanical power (EMP) by a direct method (respiratory gas exchanges) on a cycloergometer. Daily EE and partial EE in free-living conditions were computed from heart-rate (HR) recordings during seven consecutive days using individual prediction equations established from the data obtained during a 24 h period spent in whole body calorimeters with similar activities. Fat-free mass (FFM), VO2max, EMP, daily EE and EE during sleep were significantly higher in athletic than in non athletic subjects. After adjustment for FFM, VO2max, EMP, daily EE and EE during exercise were still higher in athletic than in non-athletic adolescents (P < 0.001). However, adjusted sleeping EE was not significantly different between athletic and non-athletic adolescents. Increases in exercise EE were partly compensated for by significant reductions in EE during schoolwork and miscellaneous activities. Thus, the differences in daily EE between athletic and non-athletic subjects resulted mainly from increases in FFM and EE during exercise (duration and energy cost). PMID- 11103226 TI - Fruit and vegetable availability among ten European countries: how does it compare with the 'five-a-day' recommendation? DAFNE I and II projects of the European Commission. AB - Recasting the role of fruit and vegetables (F&V) in the diet, and planning national and international campaigns to enhance their consumption are major public health service objectives. The present study seeks to describe F&V availability patterns in ten European countries and examine compliance with current recommendations. The mean and median F&V availability (g/person per d) was estimated based on household budget survey data retrieved from the Data Food Networking (DAFNE) databank. Low F&V consumers were identified based on WHO international recommendations (minimum combined F&V intake of about 400 g/person per d) and current conservative guidelines of a minimum daily intake of three portions of vegetables and two portions of fruit. Considerable disparities in F&V availability were found among the surveyed European populations. Only in Mediterranean countries did the mean daily population intake clearly exceed combined F&V recommendations. Dietary patterns were positively skewed in all populations studied, on account of the presence of exceptionally high values among segments of the populations. Moreover, the correlation was unexpectedly weak between the proportion of low fruit and low vegetable consumers (Spearman's correlation coefficient +0.18). More than 50% of the households in the surveyed populations are likely to consume less than the recommended daily vegetable intake of three portions, and this applies even to the two Mediterranean populations. The efficiency of F&V promoting strategies may be enhanced if F&V are addressed separately; furthermore, interventions that would specifically focus on vegetables are probably needed. PMID- 11103227 TI - Effects of replacing meat with soyabean in the diet on sex hormone concentrations in healthy adult males. AB - A randomised crossover dietary intervention study was performed to evaluate the effects of replacing meat protein in the diet with a soyabean product, tofu, on blood concentrations of testosterone, dihydrotestosterone, androstanediol glucuronide, oestradiol, sex hormone-binding globulin (SHBG), and the free androgen index (total testosterone concentration/SHBG concentration x 100; FAI). Forty-two healthy adult males aged 35-62 years were studied. Diets were isoenergetic, with either 150 g lean meat or 290 g tofu daily providing an equivalent amount of macronutrients, with only the source of protein differing between the two diets. Each diet lasted for 4 weeks, with a 2-week interval between interventions. Fasting blood samples were taken between 07.00 and 09.30 hours. Urinary excretion of genistein and daidzein was significantly higher after the tofu diet (P < 0.001). Blood concentrations of sex hormones did not differ after the two diets, but the mean testosterone:oestradiol value was 10% higher (P = 0.06) after the meat diet. SHBG was 3% higher (P = 0.07), whereas the FAI was 7% lower (P = 0.06), after the tofu diet compared with the meat diet. There was a significant correlation between the difference in SHBG and testosterone:oestradiol and weight change. Adjusting for weight change revealed SHBG to be 8.8% higher on the tofu diet (mean difference 3 (95% CI 0.7, 5.2) nmol/l; P = 0.01) and testosterone:oestradiol to be significantly lower, P = 0.049). Thus, replacement of meat protein with soyabean protein, as tofu, may have a minor effect on biologically-active sex hormones, which could influence prostate cancer risk. However, other factors or mechanisms may also be responsible for the different incidence rates in men on different diets. PMID- 11103228 TI - Properties of maltodextrins and glucose syrups in experiments in vitro and in the diets of laboratory animals, relating to dental health. AB - The objective of the study was to examine the cariogenic potentials of maltodextrins and glucose syrups (two glucose polymers derived from starch) using a range of techniques in vitro and in laboratory animals. The experimental methods used were: (1) measurement of acid production from glucose syrups and maltodextrins by human dental plaque micro-organisms; (2) evaluation of the role salivary alpha-amylase in degrading oligosaccharides (degree of polymerisation > 3) in the glucose polymers, estimating the products by HPLC; (3) assessment of the fermentability of trioses relative to maltose; (4) measurement of dental caries levels in three large-scale studies in laboratory rats fed on diets containing the glucose polymers. It was found that acid production from the glucose polymers increased as their higher saccharide content fell. Salivary alpha-amylase rapidly degraded the oligosaccharides (degree of polymerisation > 3), mainly to maltose and maltotriose. In the presence of oral micro-organisms, maltotriose took longer to ferment than maltose, but by the end of a 2 h period the total amount of acid produced was the same from both. Incorporated into the diets in solid form, the glucose syrups and maltodextrins were associated with unexpectedly high levels of dental caries. In conclusion, the findings were unforeseen in the light of earlier data that a glucose syrup was less cariogenic than sucrose. PMID- 11103229 TI - High-fructose feeding of streptozotocin-diabetic rats is associated with increased cataract formation and increased oxidative stress in the kidney. AB - We examined the effects of high-fructose (FR) feeding on the development of diabetic complications in the lens and the kidney of streptozotocin (STZ) diabetic rats. Male Wistar Furth rats were treated with one of two doses of STZ (HIGH STZ, 55 mg/kg body weight; MOD STZ, 35 mg/kg body weight) or vehicle alone (SHAM) and were then assigned to a control (CNTL) or 400 g FR/kg diet for 12 weeks. At the end of the study, body weight, plasma glucose and insulin concentrations differed among STZ groups (HIGH v. MOD v. SHAM, P < 0.001) but did not differ due to diet. Plasma FR concentrations were significantly higher in FR fed v. CNTL-fed groups (P < 0.0001) and in HIGH-STZ groups v. MOD-STZ and SHAM groups (P < 0.0004 and P < 0.0001 respectively). Focal length variability of the lens, a quantitative measure of cataract formation, was increased in the HIGH STZ, FR group compared with the HIGH STZ, CNTL group (P < 0.01). The concentration of H2O2 in kidney microsomes was significantly higher in HIGH STZ, FR rats v. HIGH STZ, CNTL rats (P < 0.01). Micro-albuminuria was not observed in any of the groups examined, and there was no evidence of extensive histological damage in the kidney from any rats. Under conditions of severe hyperglycaemia, high FR intake promotes the development of cataracts in the lens of the eye, and results in increased concentrations of substances indicative of oxidative stress in the kidney. Although FR has been suggested as a carbohydrate source for diabetics, a high FR diet coupled with hyperglycaemia produces effects that may promote some of the complications associated with diabetes. PMID- 11103230 TI - [Selection of multi-modality treatment of hepatocellular carcinoma]. AB - Multi-modalities of the treatment of hepatocellular carcinoma were utilized in clinical, however, we should consider not only tumor factors but also background of the patients and hepatic function reserve as well as multicentric carcinogenesis by hepatitis viruses. Most important message for clinician is how to select the suitable treatment for the patient based on survival rate and recurrence rate in each modalities. In this chapter, the author summarized the result of each treatment and indicated a selection of each treatment of hepatocellular carcinoma. PMID- 11103231 TI - [New trends in neoadjuvant chemoradiotherapy for locally-advanced esophageal cancer: esophagectomy--is it necessary?]. AB - In responders to neoadjuvant chemoradiotherapy for locally-advanced esophageal cancer, there was no significant difference in the long-term outcome between patients who underwent esophagectomy and those who did not. Esophagectomy might be unnecessary for patients who achieve a complete response with chemoradiotherapy for an esophageal cancer, in cases when salvage surgery is considered in order to treat any future recurrence. PMID- 11103232 TI - [Postoperative adjuvant therapies for esophageal carcinoma]. AB - Recent studies have focused on biochemical modulation with the combined use of CDDP and 5-FU for esophageal cancer. The combination of chemo- and radiotherapy is one of the most effective regimens for esophageal carcinoma. This combination therapy led to complete clinical remission in up to 30% of the cases in a prospective randomized study. We introduce many trials about postoperative adjuvant therapy after surgical treatment for esophageal carcinoma. The development and investigation of more effective chemotherapy regimens, as well as precise assessments of the response to preoperative treatment, is the challenge for randomized trials in the future. PMID- 11103233 TI - [Significance of neoadjuvant chemotherapy for gastric cancer]. AB - Neoadjuvant chemotherapy for high-risk patients with advanced gastric cancer is important to increase the chance for curative resection and make unresectable gastric cancer tumors resectable by down-staging of the tumor. Tumors with H0, P0, T3, T4, or N3 are the best candidates for this therapy. Randomized controlled phase III studies are needed in conjunction with accurate staging of the disease by laparoscopy. The results of histopathologic evaluation of resected materials following preoperative chemotherapy using oral fluoropyrimidine are thought to be useful as an indicator of chemosensitivity for postoperative adjuvant setting. PMID- 11103234 TI - [Current status and problem of adjuvant chemotherapy for curatively resected gastric cancer]. AB - Randomized controlled trials (RCT) on adjuvant chemotherapy for gastric cancer published in the West and Japan were reviewed. Although several small trials showed positive data, adjuvant chemotherapy for curatively resected gastric cancer has been thought to be ineffective in western countries. Results of Japanese RCTs also have not become evidence of its benefit. Despite this, suggestive data by non-predefined subset analyses of old RCTs have been misread as definitive evidence of benefit because of less understanding of clinical statistics in Japan. As a result most Japanese patients have received postoperative adjuvant chemoimmunotherapy. Recently understanding of clinical trial has spread gradually and well designed RCTs with sufficient sample size have been reported. First of all we have to determine the efficacy of adjuvant chemotherapy by carefully designed RCT using surgery alone arm as control. PMID- 11103235 TI - [Recent advance in gastric cancer chemotherapy]. AB - Because of the low chemosensitivity of gastric cancer to conventionally available agents, several approaches were investigated to design "order made" treatments using chemosensitivity tests, including the histoculture drug response assay (HDRA) which was useful in evaluating the appropriate cancer chemotherapy for the patients with Stage III/IV gastric cancer. A recent investigation using a molecular biological method was introduced to predict the sensitivity of gastric cancer specimens to 5-fluorouracil (5-FU) by dihydropyrimidine dehydrogenase (DPD) activity and its mRNA. The low activity of DPD and DPD mRNA resulted in the low sensitivity to 5-FU, although thymidylate synthetase activity was not related to the sensitivity to 5-FU. This method is promising, since a small amount of material obtained through gastrofiberscopy will be adequate to assess DPD mRNA to predict the sensitivity to 5-FU. On the other hand, some randomized control trials with a huge cohort have indicated the usefulness of a "docetaxel + cisplatin + 5-FU" regimen for advanced and recurrent gastric cancer, and "5-FU + LV + radiation" as an adjuvant therapy for advanced gastric cancer. Furthermore, the efficacy of adjuvant chemotherapy after curative resection for gastric cancer was warranted by a meta-analysis of 19 published randomized trials. The "order made" and "standard" therapies will be complementary in the further development of chemotherapy against gastric cancer. PMID- 11103236 TI - [Latest progress on chemotherapy for advanced gastric cancer]. AB - Although recent phase II studies have demonstrated high antitumor activity in the treatment of advanced gastric cancer, no significant survival benefit has been clearly demonstrated yet, when compared with 5-FU alone. More recently, a number of new agents including irinotecan and S-1 have demonstrated significant activity against gastric cancer as single agent or in combination with other chemotherapeutic agents. A phase III trial of 5-FU alone versus irinotecan plus cisplatin versus S-1 alone in advanced gastric cancer patients will be initiated in Japan Clinical Oncology Group (JCOG) within a few months. These new regimens have a potential becoming a new standard chemotherapy for the treatment of gastric cancer. The patients with peritoneal dissemination has usually not yet evaluated and explored from clinical study because of risk of toxicity and having no measurable disease. A next randomized phase III trial comparing 5-FU alone with sequential methotrexate and 5-fluorouracil in patients with peritoneal metastasis will be initiated in JCOG next year. The development of molecular biology has demonstrated the molecular mechanisms of chemoresistance or chemosensitivity, as well as a number of molecular targets against cancer cells. To date, many molecular targeted agents are being evaluated in various stages of clinical testing. These advances may provide a possibility of tailor made treatment. PMID- 11103237 TI - [Analysis of cases of complete response (CR) in esophageal and gastric cancer]. AB - We investigated 117 patients with advanced esophageal cancer who had undergone radio-chemotherapy from 1990 to 2000 in our department. Concurrent radiotherapy with chemotherapy improved the response rate, and adjuvant surgery improved the prognosis. Future problems are the establishment of a method to estimate the sensitivity of tumors to chemotherapy or radiotherapy, the improvement of diagnostic methods for evaluation of the effect and the development of new therapies and regimens for non-responders in the present radio-chemotherapy group. The CR cases of inoperable, noncurative and recurrent patients with gastric cancer in the past decade were examined. A CR of the lymph nodes was obtained in 5 cases, and that of the hepatic metastasis and peritoneal recurrence was observed in one case each. However, a CR of the primary lesion has not been attained. Though re-swelling of the lymph nodes was not observed in 4 out of 5 CR cases, a maintenance treatment after CR should be established. Topical treatment is promising for the hepatic metastasis and peritoneal seeding. Since it is difficult to attain a CR of the primary lesion, surgical resection is required to prolong the survival time. PMID- 11103238 TI - World summit against cancer for the new millennium: 4th February 2000 charter of Paris against cancer. PMID- 11103239 TI - [Pharmacokinetics of cisplatin and methotrexate in a patient suffering from advanced ureteral tumor accompanied by chronic renal failure, undergoing combined hemodialysis and systemic M-VAC chemotherapy]. AB - The pharmacokinetics of cisplatin and methotrexate were determined in a patient suffering from advanced ureteral tumor accompanied by chronic renal failure undergoing 4 consecutive cycles of M-VAC chemotherapy and hemodialysis. No significant difference was observed in t1/2, AUC or CLtot of total platinum between the patient with the chronic renal failure and patients with normal renal function. The AUC and CLtot of free platinum in the patient with the chronic renal failure were higher and lower, respectively, than in the patients with normal renal function. The free cisplatin rebounded remarkably after the end of dialysis, which may be partly attributed to an increase in the AUC and decrease in CLtot. However, the dialysis index was about 75 and 85% in the 3rd and 4th cycles, respectively. The t1/2 and CLtot of methotrexate in the patient with the chronic renal failure tended to be longer and smaller than those in patients with normal renal function, respectively. Seventy-two hours after administration, the methotrexate level was 0.02 microM, which was not at the high-risk level of high dose therapy. After four cycles of M-VAC therapy, the rest of the right ureteral tumor was extirpated and the clinical response was CR. In conclusion, it is considered that cisplatin and methotrexate can be given to a patient with chronic renal failure. However, the cisplatin and methotrexate serum levels must be monitored, even after very low doses. PMID- 11103240 TI - [Evaluation of adverse effects including neurotoxicity of combination chemotherapy with paclitaxel and carboplatin]. AB - The adverse effects of combination chemotherapy with paclitaxel and carboplatin, including neurotoxicity, arthralgia and muscle pain, were evaluated in 21 patients (30 courses) using questionnaires of the Gynecologic Oncology Chemotherapy Joint Research Group. The scores of pain and numbness peaked from the third to fourth day of treatment. Although the pain score improved subsequently, the numbness score persisted at a high level. Compared to the first and second courses, the peak pain score was higher and persisted for a longer duration in the fifth and sixth courses. Using the questionnaires, we were able to recognize a high incidence of numbness and pain in patients on combination chemotherapy with paclitaxel and carboplatin, and identify the degree and temporal changes of the adverse effects. Our results suggest that the questionnaires used in this study are clinically useful for evaluating the degree and clinical course of pain and numbness in anticancer chemotherapy. PMID- 11103241 TI - [The evaluation of pamidronate therapy for bone metastases from breast cancer]. AB - We devised a method to evaluate comprehensively the therapy to alleviate the pain of bone metastases from breast cancer according to the three items of bone pain and effects of analgesia and radiology. In 12 patients, we evaluated the therapeutic effect of pamidronate as an alleviative treatment for the pain of bone metastases from breast cancer. Bone pain was evaluated on a 6-grade scale, as was use of analgesics. Improvement in bone pain, in addition to improvement in use of analgesia, was evaluated as markedly improved, improved, unchanged, aggravated, no pain or undeterminable. Radiological improvement in bone lesions was evaluated as markedly improved, improved, unchanged, aggravated or undeterminable. An overall evaluation was made by combining the above two. In this evaluation method, pamidronate therapy resulted in an evaluation of markedly improved in 2 patients, improved in 5, unchanged in 4 and aggravated in 1, demonstrating that the therapy was very useful as an alleviative treatment for the pain of bone metastases from breast cancer. The evaluation method, in which pain, a subjective complaint, is combined with use of analgesics, an objective factor, and to which radiological evaluation is added for further objectively, may in the future to be applicable for evaluation of various alleviative treatments of pain of bone metastases. PMID- 11103242 TI - [Improvement in quality of life with vinorelbine as a single agent in two patients with recurrent lung cancer]. AB - Two patients with lung cancer that recurred after surgery and chemotherapy were administered vinorelbine (20 mg/m2) as a single agent on days 1 and 8 every 4 weeks. Patient 1 had recurrences in both lung lobes and metastases in the bone and liver after surgery, while patient 2 had a recurrence in the left lung lobe and metastases in the brain and liver after prior chemotherapy. In both patients, vinorelbine treatment improved several clinical indicators of cancer symptomatology including serum LDH, levels of other clinical blood tests, as well as a subjective assessment of the quality of life (QOL). Both patients were discharged from the hospital and subsequently treated on an outpatient basis. Because recurrent lung cancer patients generally have a poor performance status, chemotherapy for these patients is thought to worsen the QOL and increase hospital stays. The present report suggests that vinorelbine, as a single agent, might improve the QOL and make it possible to treat recurrent lung cancer patients on an outpatient basis. PMID- 11103243 TI - [Effectiveness of docetaxel plus cisplatin in large cell lung cancer showing little response to prior chemotherapy with MVP]. AB - The present patient was a 53-year-old male who had large cell lung cancer of c T4N1M0. We administered multi-drug regimen including mitomycin C, vindesine and cisplatin (CDDP) because of cancer invasion into the great vessels seen on a chest CT. After 3 courses, the cancer showed no change in size. Therefore, we adopted chemotherapy of docetaxel (Taxotere: TXT) and CDDP. After 4 courses, the size of the mass had decreased (partial response). The only major toxic defect was grade 3 neutropenia. A good response to TXT and CDDP could lead to complete resection of lung cancer. It is suggested that TXT is effective in the treatment of large cell lung cancer. PMID- 11103244 TI - [A case of advanced breast cancer that responded remarkably to chemotherapy]. AB - The case of a 49-year-old woman with axillary lymph nodes, multiple bone and multiple lung metastases from advanced breast cancer is reported. The patient responded remarkably to combination chemo- and endocrine-therapy. This patient was discharged after receiving 2 cycles of cyclophosphamide, pirarubicin and 5' DFUR, while continuing to receive daily oral doses of 5'-DFUR and MPA. The patient experienced few adverse effects during chemotherapy. The patient enjoys an improved quality of life. This combined regimen has been confirmed to be an effective treatment for patients in advanced stages of breast cancer. PMID- 11103245 TI - [A patient with recurrent uterine cervical cancer successfully treated by combined modality therapy including local injection of anticancer agents based on CDDP]. AB - The patient was a 77-year-old woman who had undergone total abdominal hysterectomy and bilateral salpingo-oophorectomy following the diagnosis of uterine cervical cancer at the age of 57. A recurrent tumor was detected on the anterior surface of the right iliopsoas muscle and fine needle aspiration of the tumor revealed squamous cell carcinoma. A needle was inserted into the tumor under transabdominal ultrasound guidance, and anticancer agents, chiefly cis diamminedichloroplatinum [II] (CDDP), were injected. The patient also received systemic chemotherapy (CAP therapy) and radiotherapy. Subsequently, computed tomography showed a 73% reduction in the size of the tumor. A mass 7 cm in diameter was still present on the anterior surface of the iliopsoas after treatment, but tumor markers fell to normal levels. The patient is presently receiving UFT (600 mg/day) as maintenance therapy. At 48 months after recurrence, there has been no increase in the tumor size or tumor marker levels, and the performance status is grade 0. In conclusion, a good outcome was achieved by local injection of anticancer agents combined with systemic chemotherapy and radiotherapy, and there were few side effects. PMID- 11103246 TI - [Three cases of metastatic breast cancer, resistant to pre-chemo and/or endocrine therapy, markedly improved with UFT and cyclophosphamide oral combination therapy]. AB - The cases of 3 female patients with metastatic breast cancer treated with oral UFT and cyclophosphamide (CPA) are reported. Patient 1 had lymph node and bone metastases. Patient 2 had bone metastasis. Patient 3 had skin, lymph node, and peritoneal metastases. All had a history of mastectomy and chemo- and/or endocrine therapy for metastatic lesions. Patients 2 and 3 had also undergone CAF combination chemotherapy. However, the lesions did not change. UFT 400 mg and CPA 100 mg, everyday, were administered to patient 1. UFT 400 mg and CPA 100 mg, 2 weeks, and UFT 400 mg, 2 weeks, were given every 4 weeks to patient 2. UFT 300 mg and CPA 150 mg, 6 weeks per 8 weeks were given to patient 3. Improvements in the metastatic lesions were seen 4 weeks after the beginning of UFT and CPA therapy. Therapy is now continuing, and no patients had a progression of the disease. All had leukopenia 2 or 4 weeks after the beginning of this therapy, and two temporarily stopped the therapy. No other side-effect was observed. Oral UFT and CPA combination therapy was considered useful for metastatic breast cancer. To prevent leukopenia and prolong the term of treatment efficacy, a treatment regimen will need to be established. PMID- 11103247 TI - [A case of stage IV gastric cancer responding to combined chemotherapy by intravenous, intraarterial and intraperitoneal injection]. AB - We treated a case of unresectable gastric cancer in which peritoneal lavage cytology and the primary tumor responded to combined chemotherapy by intravenous, intraarterial and intraperitoneal injection. The patient was a 61-year-old male with loss of appetite. He underwent laparotomy for gastric cancer in November, 1997. An intraperitoneal catheter was set without gastrectomy because of the unresectability due to extensive peritoneal dissemination and local invasion to pancreas. The patient was given combined chemotherapy using 5-FU 6,000 mg i.v., CDDP 360 mg i.p. and 5-FU 10,500 mg ia by the catheters in the supra vena cava, right gastroepiploic artery and peritoneal cavity. One month after laparotomy, peritoneal lavage cytology had changed to class I from class V, and the primary tumor had been reduced to 10 mm in size from a diffusely infiltrating tumor. His quality of life in appetite and activity was improved for 13 months. PMID- 11103248 TI - [Combined 5-FU and CDDP in a gastric cancer patient undergoing hemodialysis- pharmacokinetics of 5-FU and CDDP]. AB - The pharmacokinetics of 5-FU and CDDP was examined in a gastric cancer patient receiving regular hemodialysis (HD) for renal failure. The patient received combination chemotherapy of 5-FU and CDDP, then on the day of HD we measured the plasma concentration of 5-FU, total platinum, and non-protein-bound platinum of the patient. In the present case, the 5-FU concentration was kept at an almost even level during HD. Non-protein-bound platinum disappeared after being maintained in blood for a certain time when HD was started 30 minutes after the end of CDDP administration. From these findings, we conclude that combined 5-FU and low-dose CDDP therapy should be done by decreasing the dose of 5-FU, administrating CDDP only on the day the patient undergoes HD, and starting HD 30 minutes after the end of CDDP administration. PMID- 11103249 TI - [A complete response persisting for seven months with the use of TS-1 in a patient with paraaortic lymph node metastasis of gastric cancer]. AB - TS-1, a novel oral anticancer agent, was administered to a 68-year-old male patient with a paraaortic lymph node metastasis, 42 x 28 mm in size, of gastric cancer. The patient received four courses of treatment. Each treatment course consisted of a four-week administration followed by two drug-free weeks. The daily dose was 120 mg for the first two courses and 100 mg for the last two. A partial response was obtained after completion of the first course and a complete response was observed after the second. As of February, 2000, eleven months after the first administration, no regrowth of the tumor was observed either radiographically or by tumor marker levels. Adverse effects included stomatitis (grade 1), neutropenia (grade 3) and anemia (grade 1), all of which were temporary and needed no treatment. The patient's QOL improved gradually after the treatment. He is now in a good health and undergoes regular check-ups. PMID- 11103250 TI - [A case report of TS-1 in a patient with advanced gastric cancer]. AB - TS-1 is an oral anticancer drug that produces biochemical modulation. TS-1 is composed of FT (tegafur), CDHP (gimestat, which inhibits 5-FU degradation enzyme), and Oxo (otastat potassium, which reduces 5-FU gastrointestinal toxicities), in a molar ratio of 1:0.4:1. We administered TS-1 to a 68-year-old female gastric cancer patient, after distal gastrectomy (Stage IV, cur C). As a result of abdominal CT, the diameter of metastatic lymph node increased before and after surgery, and before TS-1 (45 x 35 mm), but it was reduced after 1 course of TS-1 (37 x 25 mm), 2 courses of TS-1 (35 x 20 mm), 3 courses of TS-1 (30 x 20 mm), 4 courses of TS-1 (30 x 20 mm), and 6 months after 4 courses of TS 1 (20 x 20 mm). The reduction rate is 74.6%. The value of CA125 was reduced 74.4 to 8.6 after TS-1. Anorexia and back pain, which occurred after operation, disappeared after TS-1. There was no side effect over grade 3. PMID- 11103251 TI - [The strategy of combination phase I/II study]. AB - Combination chemotherapy plays an important role in anticancer chemotherapy. Many new anticancer drugs, including target-based drugs, have been introduced clinically. The opportunity to evaluate combination chemotherapy including new drugs has increased recently. Combination phase I/II studies must be designed scientifically and ethically in order to evaluate the efficacy and safety of combination chemotherapy including new drugs. It is important to choose appropriate drugs and to determine an effective method of administration, because drug-drug interaction can be a problem in combination chemotherapy. It is important to clarify the causes of drug-drug interactions through pharmacokinetics analysis because these interactions influence toxicity and efficacy. To conduct high quality clinical trials, we need to improve the infrastructure of such trials. PMID- 11103252 TI - [The heartache of muscular dystrophy]. AB - Duchenne and Becker muscular dystrophy are caused by a mutation in the dystrophin gene, located on the short arm of the X chromosome. Three so called dystrophinopathy patients, a women aged 54 and two men aged 23 and 21 years, suffered from a severe dilated cardiomyopathy. Such a cardiomyopathy can develop in both carriers and patients. In addition, it is often more important for prognosis than muscle weakness. For these two reasons it is important to screen both groups for (early) cardiological abnormalities. If these are present, regular follow-up is necessary to start timely therapy. When cardiological investigations yield normal results, it is advised to screen carriers with a five year interval. Dystrophinopathy patients should be checked every year, because the cardiomyopathy sometimes develops and deteriorates over a short period of time. Patients with dilated cardiomyopathy and with a positive family history for dilated cardiomyopathy, muscle weakness or high serum creatine kinase activity should be screened for a mutation in the dystrophin gene. PMID- 11103253 TI - [Nobel prize in physiology of medicine for year 2000 for research of signal transduction in the nervous system]. AB - The three Nobel laureates Arvid Carlsson, Paul Greengard and Eric Kandel have made pioneering discoveries concerning slow synaptic transmission between neurons. As common theme, for which the Nobel Prize in Physiology or Medicine for 2000 is given, the Nobel Assembly chose 'signal transduction in the nervous system'. The work of Carlsson led to the discovery of dopamine as transmitter in the brain and opened the way for the development of the levodopa therapy of patients suffering from Parkinson's disease. His later work concentrated on the dopamine hypothesis of schizophrenia and the rationale for the mechanism of action of antipsychotics. Greengard pioneered the field of receptor-mediated phosphorylation and dephosphorylation of brain proteins. He was the first to describe the cyclic-AMP-dependent protein kinase in the brain and the activation of this kinase following dopamine receptor activation. A substrate enriched in cells that bear dopamine receptors is 'dopamine- and cyclic-AMP-regulated phosphoprotein' (DARPP-32). Phosphorylation by the cyclic-AMP-dependent kinase influences its protein phosphatase inhibiting capacity and, as such, DARPP-32 is an important 'feed-forward activator' in the dopamine signal transduction cascade. Kandel received the prize for his contributions to our understanding of the neural substrate of learning and memory. Most of his work was carried out in the sea slug Aplysia in which he was able to relate three psychologically defined forms of learning--habituation, sensitisation, and classical conditioning--to subcellular processes and intercellular signalling. Kandel is known all over the world for his eminent textbook Principles of Neural Science which inspired generations of young neuroscientists. It seems that it is not so much the signal transduction that joins these laureates but their outstanding conceptual approach to, in fact, three different themes of the neurosciences during the second part of the last century. PMID- 11103254 TI - [Intra-articular injection of hyaluronic acid as an alternative option to corticosteroid injections for arthrosis]. AB - Intra-articular hyaluronic acid injections in the case of osteoarthritis of the knee may restore the viscous properties of the synovial fluid and protect the joint's cartilage. In theory the hyaluronic acid has an analgesic and anti inflammatory effect as well as a favourable influence on chondrocyte metabolism. In six published studies which compared the effectiveness of hyaluronic acid with injection of a placebo of a non-steroidal anti-inflammatory medication, the hyaluronic acid injections had a relatively long-term favourable effect (compared with corticosteroid injections) on pain, function, serous gonitis and concomitant analgesic use. Just as for corticosteroid injections, the placebo effect was large. Intra-articular hyaluronic acid injections have few side effects. However, in general three to five injections, with weekly intervals, are needed for the effect to be achieved. Furthermore, the medication is relatively expensive. Future investigations will be necessary to provide insights into both the cost benefit aspects and the long-term effects of the medication. PMID- 11103255 TI - [Immunology in medical practice. XXXIV. Screening for suspected immunodeficiency: Introduction]. AB - A multistage laboratory protocol for the diagnosis of immunodeficiency is useful for the efficient identification of immunodeficient patients. The protocol presented in this article starts with the patient's clinical presentation. In the initial stages a low threshold for the performance of simple screening is applied, thus allowing early exclusion of potential immunodeficiencies, as well as identification of patients before serious infections have compromised their general condition. In the later stages, more elaborate tests leading to diagnosis and definitive classification are reserved for those few patients in whom the presence of an immunodeficiency is more probable. This definitive classification is important for the identification of carriers and for the genetic counselling of the family. The protocol described has been developed in cooperation with the Dutch national working parties of clinical immunologists (paediatric as well as internal medicine) and laboratory immunologists who are involved in diagnosing or treating patients with immunodeficiency. PMID- 11103256 TI - [Immunology in medical practice. XXXV. Screening of suspected immunodeficiency: diagnostic protocols for patients with opportunistic or recurrent severe infections, wasting and failure to thrive]. AB - With a multistage laboratory protocol immunodeficiencies can be efficiently identified. The article presents a diagnostic protocol that consists of three schemes. Scheme 1 describes the diagnostic protocol for the large group of patients with recurrent pulmonary and ENT-infections, where an antibody deficiency can occasionally be found. Scheme 2 presents the diagnostic protocol for the much smaller group of patients with opportunistic infections, wasting or failure to thrive. Several of these patients suffer from a severe T-lymphocyte disorder. Early diagnosis and treatment is important for the prognosis in these patients. Scheme 3 shows the diagnostic protocol for patients with recurrent infections of surface areas and deeper organs; these patients may suffer from a phagocyte disorder. PMID- 11103257 TI - [Diagnostic image (11). (Urine sample with fat droplets)]. AB - In a urine sample of a 44-year-old female patient suppository base was found due to a wrong insertion of a diclofenac suppository. PMID- 11103258 TI - [From gene to disease; ion channel proteins and the long QT syndrome]. AB - The long QT syndrome is characterised by QT prolongation on the ECG, repeated syncope and sudden cardiac death. QT prolongation is the result of delayed repolarisation at the cellular level, secondary to dysfunctioning ion channels. Ventricular arrhythmias underlie syncope and death. At least six genes, all encoding (parts of) ion channels, are causally involved. A molecular diagnosis is often feasible and can be reached reasonably straightforwardly, based on the clinical (family) history and the ECG pattern. PMID- 11103259 TI - [Multidisplinary approach to chronic back pain: postrehabilitation resumption of work the same 3-4 years later as after 6 months]. AB - OBJECTIVE: To determine the results of a multidisciplinary programme for people with chronic back pain 3 to 4 years post-programme. DESIGN: Prospective cohort study. METHOD: In 1996, 143 patients with chronic back pain had participated in an outpatients programme, of 4 weeks duration, at a Back AdviceCentre. A follow up study took place 6 months post-programme. This involved 99 men and 44 women, who at the start of the programme had a mean age of 41.6 (SD: 8.5; range: 23-58) and had experienced back pain for a mean period of 46.3 months (SD: 36.3). In January/February 2000 the patients were contacted by telephone and completed questionnaires concerning their current work status, pain intensity, perceived disability and complaints. The same questionnaires had been completed at the start of the programme and 6 months post-programme. Differences across the three scores were tested in a multivariate test for repeated measurements. RESULTS: Data were obtained from 130 patients; 103 (79%) persons were completely returned to work. The response was 92% (n = 130). As a result of back pain, 14 patients (11%) could not work or worked reduced hours compared with the premorbid situation. (This percentage was 13% at 6 months post-programme.) A further 9 patients (7%) could not work or worked reduced hours due to other health complaints. The average level of pain, disability and complaints was still lower (p < 0.001) than the pre-programme situation. CONCLUSION: The percentage of people who permanently returned to work following the multidisciplinary approach to chronic back pain, remained stable over a period of 3 to 4 years. The results were comparable to those obtained at 6 months post-programme. PMID- 11103261 TI - [Acute gastric dilatation in Duchenne's muscular dystrophy]. AB - A 15 year old boy with Duchenne muscular dystrophy had severe pain in the lower abdomen and complained of nausea and bilious vomiting. A physical examination and an abdominal X-ray indicated an acute gastric dilation. With a treatment policy of administering nothing orally, a downward-hanging stomach tube and the intravenous administration of fluid the symptoms subsided. In Duchenne muscular dystrophy there may also be atrophy of the smooth muscle layers, in addition to the known progressive atrophy of striated skeletal and cardiac muscle. This may cause clinical dysfunctioning of the gastro-intestinal tract in the second decade of life. PMID- 11103260 TI - [Indirect traumatization of spouses of Dutch war victims]. AB - OBJECTIVE: To determine the specific problems experienced by spouses of Dutch war victims. DESIGN: Cross-sectional written questionnaire investigation. METHOD: A written questionnaire was sent to 382 supporters of a foundation for spouses of war victims (Stichting Partners van Oorlogsgetroffenen (SPO)). This organization facilitates contact between fellow sufferers. The questionnaire incorporated elements from the 'Symptom checklist'-90, the 'Impact of event scale', and the 'Maudsley marital questionnaire'. The data obtained were compared to those from previous investigations. The first of these was among 346 spouses of veterans, members of a veteran organization (Bond van Nederlandse Militaire Oorlogs- en Dienstslachtoffers), who were subdivided into two groups, married to veterans with post-traumatic stress disorder (PTSS) (n = 76) and married to veterans without PTSS (n = 264). The second was with 555 women who had participated in a study, carried out in 1992, concerning the long-term effects of severe war experiences among elderly Dutch people. RESULTS: The response rate was 161/382 (42%). The spouses experienced a high burden of care. All of their symptoms were significantly more severe than in the two comparison groups, but equalled those in the partners of veterans with PTSS, with the exception of the quality of the partner relation. This was significantly less than in the partners of veterans with PTSS. In particular, the spouses reported having problems with the war victim being uncommunicative, emotionally numb, sad, and irritable. Spouses themselves also reported posttraumatic reactions such as re-experiencing and avoidance that could not be explained by their own exposure to war and violence. CONCLUSION: The results confirm the theory that by their close contact with the war victim spouses are indirectly traumatized. It is recommended that the physician who has traumatized patients in his or her practice actively inquiries as to whether the spouse can cope with the situation and to what extent. PMID- 11103262 TI - [Recognition of case histories by a third party]. AB - A case report of psittacosis during pregnancy resulting in death of the child was published in this journal. The item was taken up by a newspaper, whereupon the owners of the birds that had been implicated realised that they were being described. They now felt responsible for the tragedy and complained. Anonymising patient information will never prevent recognition by the patient himself or herself. When publishing a patient history, the authors must be aware that third parties may be involved as well. PMID- 11103263 TI - [Attention deficit-hyperactivity disorder (ADHD); etiology, diagnosis and treatment]. PMID- 11103264 TI - [Roaming through methodology. XXIII. The need for randomization and blinding in therapeutic investigations]. PMID- 11103265 TI - Simultaneous formation and detection of the reaction product of solid-state aspartame sweetener by FT-IR/DSC microscopic system. AB - The solid-state stability of aspartame hemihydrate (APM) sweetener during thermal treatment is important information for the food industry. The present study uses the novel technique of Fourier transform infrared microspectroscopy equipped with differential scanning calorimetry (FT-IR/DSC microscopic system) to accelerate and determine simultaneously the thermal-dependent impurity formation of solid state APM. The results indicate a dramatic change in IR spectra from 50, 110 or 153 degrees C, which was respectively attributed to the onset temperature of water evaporation, dehydration and cyclization processes. It is suggested that the processes of dehydration and intramolecular cyclization occurred in the solid state APM during the heating process. As an impurity, 3-carboxymethyl-6-benzyl 2,5-diketopiperazine (DKP) degraded from solid state APM via intramolecular cyclization and liberation of methanol. This was evidenced by this novel FT IR/DSC microscopic system in a one-step procedure. PMID- 11103266 TI - Nicarbazin contamination in feeds as a cause of residues in eggs. AB - A survey was carried out to investigate the prevalence of nicarbazin residues in eggs in Northern Ireland. Nicarbazin, in the form of 4,4'-dinitrocarbanilide (DNC), was detected in 39 of the 190 eggs analysed. An experiment was designed to establish the relationship between nicarbazin-contaminated feed and nicarbazin residues in eggs. The concentrations of both the DNC and 4,6-dimethyl-2 hydroxypyrimidine (DHP) components of the drug in eggs were proportional to feed levels. The maximum feed nicarbazin concentration of 12.1 mg/kg (8.6 mg/kg DNC and 3.5 mg/kg DHP) gave rise to mean maximum whole egg concentrations of 631 micrograms/kg DNC and 51.8 micrograms/kg DHP. After withdrawal of the experimental diet, DNC was undetectable in eggs after 12 days and DHP after 3 days. Feed contaminated with nicarbazin at concentrations greater than about 2 mg/kg gave rise to egg DNC residues at concentrations greater than the Differential Action Limit (DAL) set by the UK (100 micrograms/kg). DNC was contained almost entirely in the yolk of the egg, whereas DHP was distributed between albumen and yolk in a ratio of approximately 3:1. PMID- 11103267 TI - Evaluation of commercial immunoassays for cross-reactivity to clenbuterol stereoisomers and bovine metabolites. AB - Several commercially available immunoassay kits have been developed to detect the beta-adrenergic agonist clenbuterol HCl. Technical materials supplied with the kits do not generally report cross-reactivity with clenbuterol metabolites. Use of such kits to quantitate clenbuterol might lead to an overestimation of parent drug if metabolites were present. The objective of this study was to measure the cross-reactivity of clenbuterol metabolites with several commercially available clenbuterol immunoassays. Three clenbuterol-glucuronide conjugates, clenbuterol sulphamate, 4-amino-3,5-dichloro-hippuric acid (clenbuterol-hippurate), and purified clenbuterol-stereoisomers were tested for cross-reactivity. The clenbuterol-sulphamate metabolite showed significant cross-reactivity (42-77%), but clenbuterol-hippurate showed very little competition (< 0.2%) towards clenbuterol. Clenbuterol-glucuronides had little (0.1-1.6%) cross-reactivity. In addition, (R)-, (S)-, and racemic clenbuterol were used to determine the stereospecificity of the kits. Both (R)- and (S)-clenbuterol competed for binding in two of the kits, however, in one kit the (S)-clenbuterol stereoisomer had an affinity 100 times greater than the (R)-stereoisomer. The presence of significant quantities of the sulphamate metabolite of clenbuterol in a biological matrix would cause an overestimation of the amount of parent clenbuterol. This study illustrates the inherent problems of using unvalidated immunoassays for quantitation purposes. PMID- 11103268 TI - Pesticide residues in the Canadian Market Basket Survey--1992 to 1996. AB - Market basket food samples from six Canadian cities collected from 1992 to 1996 were analysed for pesticide residues. One hundred and thirty-six composites were prepared for each city, representing 99% of the Canadian diet. Residues were found most frequently in peanut butter and butter. DDE, malathion and captan occurred most frequently, while the fungicides chlorothalonil, dicloran and captan were present in the highest concentrations. Processed commodities contained fewer residues and at lower concentrations than the raw products. No residues were detected in either milk or soy-based infant formula. Of the infant foods sampled, fruit contained both the greatest number and highest concentrations of pesticides. PMID- 11103269 TI - Acephate and buprofezin residues in olives and olive oil. AB - Field trials were carried out to study the persistence of acephate and buprofezin on olives. Two cultivars, pizz'e carroga and pendolino, with very large and small fruits respectively were used. After treatment, no difference was found between the two pesticide deposits on the olives. The disappearance rates, calculated as pseudo first order kinetics, were similar for both pesticides (on average 12 days). Methamidophos, the acephate metabolite, was always present on all olives, and in some pendolino samples it showed higher residues than the maximum residue limit (MRL). During washing, the first step of olive processing, the residue level of both pesticides on the olives did not decrease. After processing of the olives into oil, no residues of acephate or methamidophos were found in the olive oil, while the residues of buprofezin were on average four times higher than on olives. PMID- 11103270 TI - Tetrahydro-beta-carboline-3-carboxylic acid compounds in fish and meat: possible precursors of co-mutagenic beta-carbolines norharman and harman in cooked foods. AB - The presence of tetrahydro-beta-carbolines and beta-carbolines was studied in raw, cooked and smoked fish and meat. 1,2,3,4-Tetrahydro-beta-carboline-3 carboxylic acid (THCA) usually was the major beta-carboline found, whereas 1 methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (MTCA) appeared in smoked and 'well done' cooked samples. THCA was detected in raw fish (nd-2.52 micrograms/g), cooked fish (nd-6.43 micrograms/g), cooked meats (nd-0.036 microgram/g), smoked fish (0.19-0.67 microgram/g) and smoked meats (0.02-1.1 micrograms/g). Smoked and cooked samples contained higher amounts of THCA and MTCA than raw products. Deep cooking of fish and meat increased both THCA and MTCA, and this was accompanied by the formation of more beta-carbolines, norharman and harman. The tetrahydro-beta-carbolines THCA and MTCA were chemical precursors of the co-mutagens norharman and harman during cooking. These and previous results confirm that foods are an important source of beta-carbolines in humans. PMID- 11103271 TI - Ochratoxin A in Brazilian roasted and instant coffees. AB - Thirty-four samples of roast and ground coffee, 14 samples of instant coffee and two samples of decaffeinated instant coffee were collected in markets and supermarkets in the city of Campinas, Brazil, and analysed for ochratoxin A using immunoaffinity columns for clean-up and HPLC with fluorescence detection for quantification. The limit of detection was 0.2 ng/g ochratoxin A. Twenty-three samples of ground and roast coffee were found to be contaminated with the toxin at levels ranging between 0.3 and 6.5 ng/g. The average concentration in all 34 samples was 0.9 ng/g. All samples of instant coffee contained ochratoxin A at levels ranging from 0.5 to 5.1 ng/g, with an average figure of 2.2 ng/g. Roast and ground coffee is the type of coffee most used by Brazilians for the preparation of the beverage. Considering that an average Brazilian adult takes five cups of coffee per day, which corresponds to 30 g of roast and ground coffee, the probable daily intake of ochratoxin A by a 70 kg adult would be 0.4 ng/kg bw, which is far below the current Provisional Tolerable Daily Intake of 14 ng/kg bw for ochratoxin A as set by the Codex Alimentarius. To study the transfer of ochratoxin A into coffee brew, the beverage was prepared by two methods: (a) the drip method and (b) the Brazilian country style method. No significant difference was observed between the two methods in terms of extraction of the toxin using five contaminated samples containing between 0.8 and 6.5 ng/g ochratoxin A. The drip method extracted 86 +/- 15% and the Brazilian country style 74 +/- 20% of the ochratoxin A initially present in the roast and ground coffee. PMID- 11103272 TI - Aflatoxin M1 in raw and ultra high temperature-treated milk commercialized in Portugal. AB - During June to September 1999, 31 samples of raw milk were obtained from individual farms and 70 samples of ultrahigh-temperature-treated (UHT) milk (18 samples of whole milk, 22 of semi-skimmed and 30 samples of skimmed milk) were collected in supermarkets in Lisbon, Portugal. The total number of samples analysed was 101. The incidence of aflatoxin M1 (AFM1) contamination in the milk samples analysed was very high (83.2%). AFM1 was detected in 80.6% of raw milk and 84.2% of UHT milk. Concerning raw milk, 17 samples (54.8%) contained low levels (0.005-0.010 microgram/l), two samples (6.5%) had levels ranging from 0.011 to 0.020 microgram/l and six samples (19.3%) had levels between 0.021 and 0.050 microgram/l. Of 70 samples of UHT milk analysed, 10 samples had levels below 0.005 microgram/l (one whole milk, two semi-skimmed milk and seven skimmed milk). Nine samples--two whole milk (11.1%) and seven skimmed milk (23.3%)--were contaminated with levels between 0.005 and 0.010 microgram/l. Twenty-five samples (eight whole milk, 44.4%; one semi-skimmed milk, 4.5%; and 16 skimmed milk, 53.4%) had AFM1 contamination levels of 0.011-0.020 microgram/l. Six samples of whole milk (33.3%) and 18 samples of semi-skimmed milk (81.9%) had levels of 0.021-0.050 microgram/l. Only two samples were contaminated at levels above the legal limit for AFM1 (one whole milk and one semi-skimmed milk, 0.059 and 0.061 microgram/l, respectively). At the moment, the contamination of milk with AFM1 does not appear to be a risk to public health, although the presence of this toxin was detected in 83.2% of the samples analysed. PMID- 11103273 TI - Fumonisins B1 and B2 in Brazilian corn-based food products. AB - Eighty-one samples of corn products were acquired from markets and supermarkets in the city of Campinas, SP, Brazil, and were analysed for fumonsins B1 and B2 (FB1 and FB2). Forty samples (49%) were positive for FB1 (0.03-4.93 (micrograms/g) and 44 samples (54%) for FB2 (0.02-1.38 (micrograms/g). The samples, in order of decreasing contamination, were, corn meal (all contaminated, 0.56-4.93 (micrograms/g FB1), followed by degerminated corn (8/11 samples, nd 4.52 (micrograms/g FB1), corn flour (9/11 samples, nd-1.46 (micrograms/g FB1), precooked corn flour (4/6 samples, nd-1.79 (micrograms/g FB1), corn grits (2/2 samples, 0.17-1.23 (micrograms/g FB1), and popcorn (4/9 samples, nd-1.72 (micrograms/g FB1). Relatively lower incidences and levels of contamination were found in corn flakes (1/4 samples, nd-0.66 (microgram/g FB1) and corn flour baby cereal (1/2 samples, nd and 0.44 (microgram/g FB1). The samples of corn on the cob (common corn in the milky stage, 7 samples) and of the typical foods 'curau' (2 samples) and 'pamonha' (7 samples), both prepared with corn in the milky stage, did not show any detectable contamination. Canned sweet corn, also harvested in the milky stage, exhibited a very low incidence of and level of contamination (2/11 samples, nd-0.08 (microgram/g FB1). The intake of corn products is low in urban areas and in most rural areas in Brazil. In certain rural areas, however, corn products play a greater role in daily meals and the calculated intake of FB1 is higher than a proposed Tolerable Daily Intake of 800 ng/kg bw/day. This is the first report on fumonisins in Brazilian corn-based food products. PMID- 11103274 TI - Occurrence of bisphenol-F-diglycidyl ether (BFDGE) in fish canned in oil. AB - The levels of bisphenol-F-diglycidyl ether (BFDGE) were quantified as part of a European survey on the migration of residues of epoxy resins into oil from canned fish. The contents of BFDGE in cans, lids and fish collected from all 15 Member States of the European Union and Switzerland were analysed in 382 samples. Cans and lids were separately extracted with acetonitrile. The extraction from fish was carried out with hexane followed by re-extraction with acetonitrile. The analysis was performed by reverse phase HPLC with fluorescence detection. BFDGE could be detected in 12% of the fish, 24% of the cans and 18% of the lids. Only 3% of the fish contained BFDGE in concentrations considerably above 1 mg/kg. In addition to the presented data, a comparison was made with the levels of BADGE (bisphenol-A-diglycidyl ether) analysed in the same products in the context of a previous study. PMID- 11103275 TI - An Internet compendium of analytical methods and spectroscopic information for monomers and additives used in food packaging plastics. AB - An internet website (http:?cpf.jrc.it/smt/) has been produced as a means of dissemination of methods of analysis and supporting spectroscopic information on monomers and additives used for food contact materials (principally packaging). The site which is aimed primarily at assisting food control laboratories in the European Union contains analytical information on monomers, starting substances and additives used in the manufacture of plastics materials. A searchable index is provided giving PM and CAS numbers for each of 255 substances. For each substance a data sheet gives regulatory information, chemical structures, physico chemical information and background information on the use of the substance in particular plastics, and the food packaging applications. For monomers and starting substances (155 compounds) the infra-red and mass spectra are provided, and for additives (100 compounds); additionally proton NMR are available for about 50% of the entries. Where analytical methods have been developed for determining these substances as residual amounts in plastics or as trace amounts in food simulants these methods are also on the website. All information is provided in portable document file (PDF) format which means that high quality copies can be readily printed, using freely available Adobe Acrobat Reader software. The website will in future be maintained and up-dated by the European Commission's Joint Research Centre (JRC) as new substances are authorized for use by the European Commission (DG-ENTR formerly DGIII). Where analytical laboratories (food control or other) require reference substances these can be obtained free-of-charge from a reference collection housed at the JRC and maintained in conjunction with this website compendium. PMID- 11103277 TI - Medical publishing: how does it affect you? PMID- 11103276 TI - The acute abdomen and its management. PMID- 11103278 TI - Controversies within colorectal surgery. PMID- 11103279 TI - Investigation of colonic disease. AB - Bowel cancer awareness among the general public has heightened in recent years. The promotion of prompt referral and the pressure on early diagnosis will alter our previous strategies on colonic evaluation. This article gives an overview of the colonic investigations currently available. PMID- 11103280 TI - Ulcerative colitis. AB - Ulcerative colitis is a relatively common cause of altered bowel habit and rectal bleeding. Coordinated management of the investigation and care of patients with this condition is vital to optimize treatment. This article reviews current management options. PMID- 11103281 TI - Improving outcomes in colonic cancer. AB - Many of the symptoms of colon cancer do not start until the tumour has spread outside the bowel, and treatment at this stage has reduced chances of cure. Early detection and the optimum combination of surgery and adjuvant treatment can make a significant impact on outcome. PMID- 11103282 TI - Improving outcomes in rectal cancer. AB - Concerted efforts are being made to improve the poor surgical outcomes in rectal carcinomas and this includes the use of total mesorectal excision by specially trained colorectal surgeons, in addition to the use of pre- or postoperative radiotherapy and chemotherapy. The role of each modality should be carefully evaluated, and benefits weighed against toxicity and added costs. PMID- 11103283 TI - Management of oesophageal varices. AB - Beta-blockers are the treatment of choice to prevent the first episode of variceal bleeding and further rebleeding episodes. In acute bleeding all patients should receive pharmacological treatment with vasoconstrictors and endoscopic treatment. Failure of therapy should lead to consideration of transjugular intrahepatic portosystemic shunting. PMID- 11103284 TI - Treatment of paediatric cerebral palsy with Dysport. AB - Dysport (Clostridium botulinum type A toxin-haemagglutin complex) has had its licence extended to include treatment of children aged 2 years and over with dynamic equinus foot deformity, caused by spasticity associated with cerebral palsy. Dysport reduces muscle tone, thus improving function, relieving pain, and facilitating physiotherapy, application and tolerability of splints. PMID- 11103285 TI - The risks of clinical practice. AB - The hospital workplace is changing dramatically, moulded in part by the need to protect medical staff from the increasing physical and mental risks of medical practice today. This article examines the risks and look to the future. PMID- 11103286 TI - Acute heart failure: a practical guide to management. AB - Acute heart failure is a common cause of referral and admission to hospital. It requires prompt recognition and treatment. This article will address the practical issues regarding diagnosis, investigation, treatment and prevention of recurrence. PMID- 11103287 TI - Sample size determination in clinical research: 1. AB - In this two-part series, the main designs in clinical research are considered and, using examples, sample size calculation is explained. The information required in order to calculate the sample size for clinical research is highlighted to enable researchers to spend their time most efficiently with statisticians. PMID- 11103288 TI - Issues in measurement. AB - Measurement lies at the heart of all quantitative methods in clinical, epidemiological and health services research. If the measures employed lack the essential features of validity and reliability then the conclusions drawn from empirical findings may mislead. This short article explains and explores the desirable features of measurement instruments used in health-care research. PMID- 11103289 TI - Finding health information on the Internet: health professionals. AB - This article discusses some of the key problems that clinicians encounter when searching the Internet for health information. Guidance is provided on how best to search the Internet to find high-quality information and resources. The Internet is a dynamic and valuable source of information for health professionals, but information accessed through the Internet should be critically evaluated before being applied to practice. PMID- 11103290 TI - Spontaneous rectus sheath haematoma mimicking an enlarged urinary bladder. PMID- 11103291 TI - Causes of the acute abdomen: add thrombophilia to your list. PMID- 11103292 TI - Retained gall-stone as a differential diagnosis of acute appendicitis. PMID- 11103293 TI - Orthopaedic surgery in the elderly. PMID- 11103294 TI - Multiple myeloma with hyperparathyroidism. PMID- 11103295 TI - Severe postoperative nausea and vomiting in a parkinsonian patient. PMID- 11103296 TI - Consumption of black tea elicits an increase in plasma antioxidant potential in humans. AB - Epidemiological studies suggest that the consumption of tea flavonoids may be associated with reduced risk of coronary heart disease, stroke and cancer-related deaths. The flavonoids are polyphenols which in vitro exhibit antioxidant properties. Tea flavonoids are known to be rapidly absorbed into the circulation following oral ingestion. To date few studies have demonstrated that these bioavailable flavonoids retain antioxidant properties in vivo. Nine healthy subjects aged between 26 and 59 (one male and eight females) took part in 3 study days. On 1 day subjects consumed no tea and on the other 2 days subjects drank either black tea with milk, or black tea alone at hourly intervals between 9.00 a.m. and 14.00 p.m. Blood was sampled at 9.00 a.m. and at 12.00 p.m. and 15.00 p.m. The antioxidant potential of plasma was determined using the ferric reducing antioxidant power (FRAP) assay. Subjects consuming no tea exhibited no significant change in FRAP across the 6 hours of the study day. Similarly consumption of milky tea produced no change in FRAP between 9.00 a.m. and 12.00 p.m. and the 50% increase in FRAP noted between 12.00 p.m. and 15.00 p.m. was not statistically significant. When the subjects consumed black tea without milk FRAP increased by 65% between 9.00 a.m. and 12.00 p.m. (P = 0.02) and at 15.00 p.m. was 76% higher than at 9.00 a.m. (P = 0.002). Heavy consumption of black tea thus appears to elevate circulating antioxidant potentials in vivo. This is an effect which appears to be totally negated by the drinking of tea with milk. Although tea may thus represent an important source of dietary antioxidants in many societies, the role of tea in reducing risk of major disease states needs to be investigated in more detail. PMID- 11103297 TI - Sources of dietary fluoride intake in 4-year-old children residing in low, medium and high fluoride areas in Iran. AB - Accurate estimation of fluoride dietary intake is desirable for optimising caries prevention. Little is known about the dietary fluoride intake of children aged 4 years, an age when many permanent teeth are forming. This study was undertaken in Fars Province, Iran, in 1995-1996, where there are big differences in temperature between winter and summer. The aims were to determine: (a) the relative contributions of different components of the diet to fluoride intake, (b) the effect of variation in fluoride concentration in drinking water, and (c) the effect of climate temperature. Temperature varied between +40 degrees C in summer to -5 degrees C in winter. The mean fluoride concentrations in drinking water in the three areas were 0.3, 0.6 and 4.0 mgF/L. Dietary information was obtained by two 3-day diet diaries with interview, validated with reference to international standards. The fluoride content of foods was measured using the silicon facilitated diffusion method. One hundred and three 4-year-old children completed the study. The mean (and 95% confidence interval) dietary fluoride intakes in each of the three areas, respectively, were 413 (+/- 21), 698 (+/- 89) and 3472 (+/- 557) micrograms/day. Drinks provided 72 to 87% of dietary fluoride--this proportion increased with increasing water fluoride concentration and increasing climate temperature. Tea (infusion) was an important source of dietary fluoride, providing 31 to 38% of total dietary intake. Tap water was a more important source of fluoride than soft drinks. Cooked rice and bread were the most important food source of fluoride and the amount of fluoride they contributed increased as water fluoride concentration increased. The results of this first such survey in the Middle East showed (a) that water (as a drink) and tea were by far the most important contributors to dietary fluoride intake, (b) substantial increases in fluoride intake with increasing water fluoride concentrations, and (c) substantially higher fluoride intakes in summer than in the winter. PMID- 11103298 TI - Preservation of alpha-tocopherol in sunflower oil by herbs and spices. AB - The ability of some commercially available herb and spice extracts to preserve alpha-tocopherol in sunflower oil during heating at 85-105 degrees C was assessed using sunflower oil as a model system. The Rancimat was evaluated for the heating stage and was used throughout as it was shown to be viable: alpha-tocopherol did not evaporate under the test conditions. The delay in the onset of rancidity was found to be directly related to the initial alpha-tocopherol concentration (P < 0.01). Rosemary, thyme, turmeric, sage, oregano and cumin extracts (2000 mg.kg-1) delayed rancidity (P < 0.01) and preserved alpha-tocopherol (P < 0.01). Some preservation was observed with clove extract but coriander and cardamom extracts were pro-oxidants. With thyme extract, the log of the induction time (as an indicator of the delay in rancidity) was directly proportional to the temperature (85-100 degrees C). The ethyl acetate, hexane and methanol extracts of fresh sage were effective for preserving alpha-tocopherol (P < 0.01). With thyme, rosemary and sage extracts, the increase in the preservation of alpha-tocopherol was directly related to the concentration of the herb extract (P < 0.01) and was quite effective even at 100 mg.kg-1. The increased delay in the onset of rancidity was due directly to the improved preservation of alpha-tocopherol (P < 0.01). In further experiments, the preservative effect of turmeric was shown not to be due to its reported major antioxidant, curcumin, even though it delayed rancidity. When herb/spice extracts were examined mixed with thyme, bay and turmeric showed synergism (P < 0.01) whereas bay alone was slightly inhibitory. The mode of action appeared to be due to free radical activity rather than through singlet oxygen generation. PMID- 11103299 TI - Use of a single method in the extraction of the seed storage globulins from several legume species. Application to analyse structural comparisons within the major classes of globulins. AB - In this study, a single, improved methodology was used to extract, fractionate and purify the 11S (legumin-type or related to the alpha-conglutin from Lupinus albus L.), 7S (vicilin-type or related to the beta-conglutin from L. albus) and 2S (related to the gamma-conglutin from L. albus) families of proteins from eight legume species: L. albus, Glycine max (L.) Merr., Pisum sativum L., Vicia faba L., Cicer arietinum L., Phaseolus vulgaris L., Lens culinaris Med. and Arachis hypogaea L. The sedimentation coefficients obtained varied from 1.9 to 8.1 for the gamma-conglutin-related proteins, from 5.1 to 10.5 for the beta-conglutin related proteins and from 12.0 to 14.9 for the alpha-conglutin-related globulins. The gamma-conglutin-related proteins is the most heterogeneous group. Antibodies produced against each type of gamma-conglutin polypeptide chain recognize the other polypeptide chain as well as other polypeptides in the corresponding globulins from all species examined. The 7S globulins are typically composed of a large number of polypeptides, covering a wide range of molecular masses (10 to 70 kD). The presence of disulphide bonds is apparently absent and the occurrence of glycopolypeptides is not widespread. Finally, the 11S globulins are characteristically formed by a limited number of polypeptides that may be divided into a lighter group (20-25 kD) and a heavier group (35-50 kD). The presence of disulphide bonds is apparently widespread but the occurrence of glycopolypeptides seems to be relatively rare. Both the 7S family and the 11S globulins studied by immunoblotting exhibit a low level of structural similarity. PMID- 11103300 TI - Particle size of solid food after human mastication and in vitro simulation of oral breakdown. AB - Mastication, the first step in food digestion, results in the breakdown of solid food and its lubrication with saliva. Although the rate and extent of starch digestion are closely dependent on the way food is chewed, this factor has not been adequately considered in the preparation of food for in vitro digestion experiments. The purpose of this study was to determine the size distribution of starchy food particles before swallowing and to use an in vitro mincing procedure to simulate how food is divided up during chewing. Foods differing in texture and size (bread, spaghetti and tortiglioni) were chewed by 12 healthy subjects and spat out before swallowing. Chewing time and saliva impregnation were measured for each mouthful. The particle sizes resulting from experiments with chewed and minced bread and pasta were analysed respectively by light laser diffraction and image analysis. Chewing time was longer for bread than pasta, resulting in higher saliva impregnation. Chewed bread showed a bimodal distribution of particle size (30 microns, 500 microns), whereas both kinds of pasta produced particles of similar size (0.5 to 30 mm2) after mastication. Mincing reproduced the division of bread and pasta as achieved by chewing in an acceptable way. From our results it seems that the size of particles resulting from mastication depends on food texture. We succeeded by wetting and mincing food to prepare food in a similar bolus-like form before swallowing. Mincing provides a simple means of simulating the reduction of food into particles for in vitro digestion studies. PMID- 11103301 TI - Amino acid profile of milk-based infant formulas. AB - The protein content and amino acid profile of three milk-based infant formulas, two of which were powdered (adapted and follow-on) and the third liquid, were determined to check their compliance with the EU directive and to evaluate whether or not they fulfil an infant's nutritional needs. To obtain the amino acid profile proteins were subjected to acid hydrolysis, prior to which the sulfur-containing amino acids were oxidized with performic acid. The amino acids were derivatized with phenylisothiocyanate (PITC) and then determined by ion-pair reverse phase high performance liquid chromatography (HPLC) In the case of tryptophan a basic hydrolysis was applied and there was no need of derivatization. The protein contents of the analysed formulas were in the ranges established by the EU directive for these products and the amino acid contents were in the ranges reported by other authors for these types of formulas. In all cases the tryptophan content determined the value of the chemical score, which was always lower than 80% of the reference protein but in the ranges reported by other authors. The analysed adapted infant formula provides amino acids in amounts higher than the established nutritional requirements. PMID- 11103302 TI - Concentrations of iron, copper and zinc in human milk and powdered infant formula. AB - Concentrations of iron, copper and zinc were determined in 56 samples of mature human milk from Canarian women and 5 samples of powdered infant formula. According to the literature our data fall within the normal limits in each kind of milk. The mean concentration of Fe, Cu and Zn of powdered infant formula was significantly higher than those concentrations found in the human milks. Significant differences among the concentrations of the studied metals for the milks of considered mothers were observed. The Fe, Cu and Zn intakes of infants fed with human milk are lower than the requirements recommended by the Food and Nutrition Board (1989). However, the infants fed with powdered infant formula had consumed an adequate intake of Fe and Cu. A progressive decrease of the metal concentrations with the lactation stage was observed. The human milk obtained in spring presented Fe and Zn concentrations lower than in autumn, which could be due to changes in nutritional habits of the mothers. Age of mother and number of previous children seem to influence the Zn and Cu concentrations of human milk. PMID- 11103303 TI - Influence of legume blends on fried papad quality. AB - Legumes and their blends are widely used for the production of papads. Papads with low fat content would be a boon to populations looking for low-calorie foods with retention of organoleptic profile. Judicious blending of legumes such as black gram, green gram, bengal gram, red gram and cowpea revealed that low-fat fried papads could be prepared from a blend of 40:36:24 blend of bengal gram:black gram:green gram flours. The blend had 15.6% lower fat content as compared to the control prepared from black gram flour alone. Other quality parameters such as expansion ratio, texture in terms of crispness, colour and overall organoleptic quality were also evaluated. PMID- 11103304 TI - Effects of moisture content, hybrid variety, kernel size, and microwave wattage on the expansion volume of microwave popcorn. AB - This study investigated several independent variables affecting the expansion volume of microwave popcorn; (a) different hybrids were used, (b) hybrids were popped at different moisture contents, (c) microwave ovens with various wattages were employed, and (d) kernels were separated by size. The greatest expansion volume was achieved with Hunt-Wesson 214 at 11% moisture. Overall, the large capacity bag and the 1000-watt microwave oven produced the greatest expansion volume. Medium-sized kernels produced the greatest expansion volume. PMID- 11103305 TI - Aspects of manufacture, composition and safety of orubisi: a traditional alcoholic beverage in the north-western region of Tanzania. AB - The processing technology and characteristics of orubisi/amarwa, an opaque beer commonly consumed in Kagera region in the north-western part of Tanzania is described in detail. The protein content of orubisi increased from 2.0 to 2.7% after 120 hours of fermentation. The maximum alcohol content of orubisi as determined by specific gravity method was 2.5%. The alcohol profile of orubisi analysed by gas liquid chromatography (GLC) was found to contain ethanol and iso butanol. The test for methanol was negative. Orubisi was characterised as product with relatively high acidity ranging from 0.35-0.89 g/100 ml and a final pH of 3.7. The levels of fermentable sugars--sucrose, maltose, glucose and fructose- were 0.5, 0.7, 1.8 and 0.6 g/100 ml after 120 hours of fermentation, respectively. High microbial counts were encountered in orubisi. The viable counts included yeasts: 2.0 x 10(7) cfu/ml, moulds 7.4 x 10(6) cfu/ml, coliforms 1.18 x 10(2) cfu/ml, lactic acid bacteria 6.5 x 10(7) cfu/ml and total aerobic count 2.95 x 10(7) cfu/ml. Based on these results, orubisi poses a serious danger to public health due to the presence of high numbers of total count and coliforms. In order to improve the safety of orubisi the pasteurisation step is recommended in the process of preparing orubisi. Hygienic handling of the final product is necessary in order to avoid contamination before consumption. The presence of trace amount of iso-butanol can lead to irritation of mucous membranes and depression of central nervous system. PMID- 11103306 TI - Nutritive value of traditional sweets consumed in the Arab Gulf countries. AB - Proximate, mineral, fatty acid and cholesterol composition of eight traditional sweets commonly consumed in the Arab Gulf countries were determined. Four sweets were obtained from Bahrain, whereas the other sweets were obtained from Oman. Protein level ranged from 0.2 to 9.0%, while the fat content ranged from 7.9 to 18.0%. In general, the sweets were found to be high in energy content but poor in most minerals. Iron and zinc contents were low (less than 2 and less than 1 mg/100 g for iron and zinc, respectively). Cholesterol was high in four sweets (range from 10.6 to 20.4 mg/100 g), mainly because of the use of animal fat in preparation of these sweets. The fatty acids profiles showed that palmitic and oleic acids were predominant. One sweet (eggbaith) was found to be very high in linoleic (42%) and low in palmitic (9.6%) acids. The study showed that some traditional sweets are nutritious, while others should be consumed with moderation. PMID- 11103307 TI - Development of food products based on millets, legumes and fenugreek seeds and their suitability in the diabetic diet. AB - A multitude of investigations have demonstrated the beneficial hypoglycemic effect of millets, fenugreek seeds and legumes in diabetic subjects. However, the bitter taste of fenugreek seeds and coarse nature of millets have been limitations in using them in daily dietaries. Moreover, as of today, the availability of special foods for diabetics in the Indian market is negligible. The millets, fenugreek seeds and legumes in judicious combination, after suitable processing, were used to formulate three nutritious food products--dhokla (leavened steamed cake), uppuma (kedgeree) and laddu (sweet balls), which are popular traditional snack foods in India. Evaluation of these food products for glycemic response in five normal and five diabetic subjects showed hypoglycemic effects in terms of glycemic-index (GI). The highest GI was observed for dhokla (34.96) followed by laddu (23.52) and uppuma (17.60) in normal subjects. All three food products differed significantly from each other in GI. Comparison of GI of all three food products in normal subjects with diabetes did not show significant differences (P approximately 0.05). The food products were well tolerated and acceptable to the subjects. These food products may have an important role in dietary management for diabetic people and may cater for their needs on a large scale if commercialized. PMID- 11103308 TI - Distribution of key ions of chloroquine biotransformation and cholesterol uptake in rat liver. AB - Oral administration to rats of chloroquine (5 mg/kg) three times per week for 8 weeks was investigated in two nutritionally compounded diets. It was observed that ferrous ion was preferentially retained in the presence of high (25%) protein diet even though all other ions, Ca2+, Mg2+ and K+ had varying uptake. The study shows no effect on the utilization of glucose and acid phosphatase. The difference in the level of lactate dehydrogenase (LDH) in both diets was not statistically significant. However, the level of cholesterol was significantly lower in both high and low protein diets when compared to control, suggesting that chloroquine biotransformation may interact with cholesterol metabolism in an unknown manner. The decrease in total cholesterol content corresponds to a similar decrease in malondialdehyde formation (lipid peroxidation). This result suggests that the biologic effects of chloroquine including its antimalarial action may be directly related to its lysosomotrophism. PMID- 11103309 TI - Severe falciparum malaria. World Health Organization, Communicable Diseases Cluster. PMID- 11103311 TI - Alpha- and beta-adrenergic blood pressure response of the rabbit hind limb vessels during cold adaptation. PMID- 11103310 TI - Lambertian acid and its amino derivatives: a new group of perspective neurotropic agents. PMID- 11103312 TI - Organ specificity of the Gus gene expression under the effect of testosterone as related to the activity of lysosomal beta-glucuronidase (EC 3.2.1.31) and of androgen-dependent pheromones in laboratory mice. PMID- 11103313 TI - Multilevel effects of transthoracic microelectrostimulation in acute myocardial infarction. PMID- 11103314 TI - Analysis of the beta-adrenergic reaction of the chick amnion. PMID- 11103315 TI - A new experimental model of depression: WAG/Rij rats genetically predisposed to absence epilepsy. PMID- 11103316 TI - Effect of the Ala-Glu-Asp-Gly peptide on lifespan in Drosophila melanogaster. PMID- 11103317 TI - Effects of neurotransmitters on the chloride sensitivity of microsomal Mg(2+) ATPase of fish (Abramis brama L.) brain. PMID- 11103318 TI - Interaction between roach Rutilus rutilus embryos in a clutch and the postembryonal consequences of this interaction. PMID- 11103319 TI - The role of the left hemisphere in estimation of emotional significance of words: examination of patients with affective disorders treated by unilateral electroconvulsions and psychotropic drugs. PMID- 11103320 TI - The oldest placental mammal. PMID- 11103321 TI - Effect of methyl jasmonate on growth processes in pea (Pisum sativum L.). PMID- 11103322 TI - Changes in background levels of heavy metals in a marine environment. PMID- 11103323 TI - Low-dose ionizing radiation inhibits the aging-related rise in accumulation of spontaneous cytogenetic abnormalities: the genome-stabilizing effect. PMID- 11103324 TI - The current state of the Atlantic salmon Salmo salar populations in small rivers of East Murman (Kola Peninsula). PMID- 11103325 TI - Odor attractiveness of males in laboratory mice in the case of humoral immune response to nonreplicable antigens. PMID- 11103326 TI - Independent inheritance and expression of apomeiosis and parthenogenesis in maize gama grass hybrids. PMID- 11103328 TI - Goldfish Carassius auratus gibelio (Bloch) occur in the Russian area of the Black Sea. PMID- 11103327 TI - Fixation of heterosis in the course of successive generations without hybridization. PMID- 11103329 TI - Histochemical indices of cutaneous glands as criteria of taxonimic evaluation of two vole species (Microtus socialis and Microtus guentheri). PMID- 11103330 TI - Burrows of mammals as acoustic devices: a study of the bobac burrow as an example. PMID- 11103331 TI - An aquatic ecosystem: a large-scale diversified bioreactor with a water self purification function. PMID- 11103332 TI - Determination of plant families using fuzzy set theory. PMID- 11103333 TI - Effect of environmental conditions on hormone concentrations in roach (Rutilus rutilus) fry from two phenotypic groups. PMID- 11103334 TI - Phoronis ovalis (Phoronida, Lophophorata) in the White Sea: the first discovery of phoronids in the Arctic Basin. PMID- 11103335 TI - Fish response to hydrostatic pressure changes in water currents varying in turbulence intensity. PMID- 11103336 TI - Nephridium structure in sipunculans (Sipuncula). PMID- 11103337 TI - Flight distances of the reindeer. PMID- 11103338 TI - Effects of Semax on Ca2+ responses of human neutrophils. PMID- 11103339 TI - Cytotoxicity of polyhydroxyalkanoates in animal cell cultures. PMID- 11103340 TI - Mercury may serve as a growth factor in microorganisms subjected to stress. PMID- 11103341 TI - Na(+)- and H(+)-dependent ATP synthesis in extremely alkaliphilic anaerobes. PMID- 11103342 TI - [Air bags and their effect on the ear]. PMID- 11103343 TI - [Telemedicine in otorhinolaryngology exemplified by a Tubingen-Leipzig video conference]. AB - "Telemedicine" is a major new development with great potential for improving health care delivery. It therefore affects each department in medicine. There is a great deal of telemedicine activity around the world. However, the term telemedicine is not clear. It describes all forms of medical information, transferred from a relevant distance by an electronic transfer media. An essential condition for communication is the intelligibility between transmitter and receiver. Because of different transmitting technologies and networks in distinct countries, towns, or even academic institutions, satisfactory contact is not possible. In the last decade, the demand for worldwide audiovisual data transmission has led to the standardization of telecommunication media. Therefore it is no longer necessary to transport medical data (or even patients) by conventional manners, e.g., post, car, or aircraft. Telemedicine for diagnosis and management can be bidirectional in real-time, long-distance videoconferencing, in which the patient consults a specialist located at the remote site, or it can be the transmission of either real-time or pre-recorded images and data to a remote expert, as in teleradiology or telepathology. Another application is the use of videoconference systems in the course of meetings. The remote specialist has the opportunity to take part in the session, e.g., with a lecture. Furthermore, the remote specialist can demonstrate special operative techniques for teleteaching purposes, some of which may be specialities of the particular medical unit, e.g., operation in open NMR, telemanipulation, or telerobotic procedures. In this paper, we describe the use and benefit of a videoconference between the departments of otolaryngology and head and neck surgery of the Universities of Tubingen and Leipzig by means of an "ISDN-based videoconference system". During the meeting, the "operating course for reconstructive surgery in the head and neck", the practicability, reliability, costs and quality were determined and compared with other technologies for audiovisual data transfer. PMID- 11103344 TI - [Functional and esthetic results of osteoplastic frontal sinus operations and profile reconstruction with bitemporal coronal incision]. AB - BACKGROUND AND OBJECTIVE: The majority of inflammatory frontal sinus diseases requiring operative intervention can now be managed successfully by endonasal surgery. In problematic cases, however, optimal exposure of the entire frontal sinus and possibly complete mucous membrane removal combined with sinus obliteration must be achieved. In these cases, external frontal sinus surgery is indicated. PATIENTS/METHODS: The operative approach and findings of 35 osteoplastic sinusotomy operations and 5 forehead reconstruction operations with porous polyethylene are reviewed. Median follow-up was 3.7 years, ranging from 1 to 7 years. Indications for surgery were trauma (12), infections (8), tumors (11), muco- or pyoceles (4), and frontal deformities (5). RESULTS: The overall functional and esthetic result was excellent. No recurrent infection was noted. None of the patients have required revision surgery so far, and no serious complications occurred. Abnormal forehead sensation was reported by 12 patients (30%). Temporary double vision was described in 3 cases (7.5%). CONCLUSIONS: Due to optimal sinus exposure and advantageous incision, osteoplastic frontal sinusotomy leads to excellent functional and esthetic results. Porous polyethylene demonstrates superior material properties and seems to be, in comparison with other implants, the current material of choice in frontal reconstruction. PMID- 11103345 TI - [Healing process of cartilage attached to a polydioxanone implant]. AB - BACKGROUND AND OBJECTIVE: To correct severe septal deformities, it is often necessary to perform an extracorporal septoplasty with temporary removal of the deviated septal cartilage, followed by straightening and reimplantation into the nose as free graft. The utilization of a compound graft, which sutures the cartilage fragments to a resorbable polydioxanon (PDS) foil, facilitates this technique immediately while simultaneously ensuring support of the nasal dorsum. PATIENTS/METHODS: On account of the good clinical experiences (during the past 3 years 71 patients have been treated using this method), we investigated the biological properties of the foil and its degradation products in connection with cartilage in an animal model from a histological point of view. In five rabbits, a 0.15-mm-thick PDS foil was implanted in combination with the cartilage into the outer ear. The follow-up period was between 2 and 25 weeks. RESULTS: The results showed the following: (1) Up to the tenth week, the foil maintains the original structure; (2) The foil will be completely resorbed after 25 weeks; (3) The cartilage underneath the foil is protected from necrosis; (4) The degradation products of the polydioxanon do not interfere with the healing process; (5) Concerning the regeneration of cartilage, the foil acts as a guarding splint; (6) After resorption, almost no scar tissue remains. CONCLUSIONS: Due to the fact that the PDS foil endonasal implant means no additional risk to the patient, we can recommend the compound graft for simplification of extracorporal septoplasty. PMID- 11103346 TI - [Morphological and functional changes in nasal mucosa after nCPAP therapy]. AB - BACKGROUND AND OBJECTIVE: The treatment success of nCPAP therapy (nasal continuous positive airway pressure) depends partly on the relief of symptoms and partly on long-term patient acceptance and the related avoidance of complications. Nasal complaints constitute the most frequently reported side effects and, together with problems of mask application, are among the primary factors causing an nCPAP-therapy to be prematurely discontinued. PATIENTS/METHODS: To assess the morphological changes of the nasal mucosa during nCPAP-therapy, we excised specimens of nasal mucosa tissue in twelve patients with obstructive sleep apnea syndrome (OSAS) both before and 3-10 months after establishing nCPAP-mask acceptance. The specimens were examined by electron microscopy. RESULTS: In all these patients, acceptance of the CPAP mask marked the initial part of therapy. In addition, mucociliary clearance was assessed by the saccharin test before and after therapy. In all patients, the nasal epithelium underwent fundamental changes upon CPAP therapy, which became manifest as modifications in the shape of epithelial cells, conglutination and clumping of the microvilli, and the appearance of immunocompetent cells. Once patients were nCPAP mask compliant, mucociliary clearance was distinctly prolonged in all cases. CONCLUSIONS: A successful therapeutic concept should provide normalization of room temperature and air humidity once nCPAP mask compliance has been achieved, and include regular assessment of the condition of the mucosa in the upper respiratory tract. Only by these measures can nasal complications be countered or therapy be applied at an early stage. PMID- 11103347 TI - [Reconstruction of traumatic skull base defects with alloplastic, resorbable fleece]. AB - BACKGROUND AND OBJECTIVE: Defects of the frontal skull base can be reconstructed with different types of material. Homologous material possibly involves the risk of infection and thus complicated wound healing. Therefore, alloplastic material seems to be an interesting alternative. PATIENTS/METHODS: In the present paper, we report on a series of 45 patients who underwent reconstruction of the anterior skull base. The defects were localized in different regions and had a maximum diameter of 10 cm2. Of the defects, 39 were closed via a sandwich technique and 12 via an onlay. The rhinosurgical approaches were aimed at closing the defects with Ethisorb, a resorbable collagen texture, and supporting the surgical field with a balloon catheter from below. Postoperative controls were performed after 6 weeks (endoscopy, determination of beta 2-transferrin) and 6 months (CT scans). RESULTS: All defect reconstructions were stable and tight, i.e., neither a CSF leakage nor a meningoencephalocele was found. The healing was completely normal and the endoscopic controls showed a regular mucosal lining over the defects. CONCLUSIONS: We conclude that Ethisorb is a useful alternative material in the treatment of frontal skull base defects. PMID- 11103348 TI - [Acceptance of wearing hearing aids by children: a longitudinal analysis]. AB - There exist no systematic longitudinal studies concerning the acceptance of hearing aids in Germany. This study examines the acceptance of a hearing aid (defined by its daily/weekly use) in the management of children with persistent sensorineural hearing loss over a period of years. 35 children with monaural or binaural hearing loss were treated with a hearing aid. All children had at least a 25-dB averaged mid-frequency pure-tone hearing loss (500, 1000, 2000, 4000 Hz). The data consist of standardized parent ratings at 4 points in time over a period of nearly 30 months. Unilateral-impaired children wore their hearing aids less often than bilateral-impaired children. This effect was not significant at the beginning (P = 0.85) but increased over time. By the end the difference was significant (P = 0.004). Mild to moderate monaural hearing-impaired children accepted their hearing aids, whereas children with severe to profound hearing loss refused to wear them. Bilateral hearing-impaired children demonstrated, a priori, a better wearing acceptance that even improved with time. There was never a significant difference between boys and girls in their average wearing time. A significant correlation of age and wearing acceptance was also not observed at any time. Hearing aids are an effective treatment with high acceptance and compliance, especially by children with bilateral sensorineural hearing loss. The quality of acceptance of monaural hearing-impaired children needs to be studied further. PMID- 11103349 TI - [Bilateral acoustic trauma after air bag inflation]. AB - Airbag inflation is associated with a loud noise. This very short noise has a peak amplitude of 170 dB sound pressure level and can make otologic injuries. We report a case of acoustic trauma after airbag inflation. The 24-year-old man has hearing loss both side in the 3000- to 6000-Hz range and in the deep-frequency. The possibility of acoustic trauma from airbag noise are probably much more common and need more attention in the clinic. PMID- 11103350 TI - [Ear dysfunction due to air bag detonation?]. AB - Air bags are among the latest developments in extensive automobile safety systems. They successfully have saved the lives of car occupants in road accidents. Many additional injuries caused by air bags from minor to severe have been reported. With the help of two acceleration sensors, the electronic tuner amplifier records the vehicle's deceleration. This is the adequate trigger for air bag deployment, which creates an intense noise of up to 170 dB sound pressure level. This noise level can cause cochlear damage. We present two patients with otologic symptoms after spontaneous air bag deployment. PMID- 11103351 TI - [Personnel management in German hospitals. Leadership deficits hinder organization and staff engagement]. PMID- 11103352 TI - [Andy Warhol's plastic nose revision reflected in his work]. AB - Andy Warhol underwent dermabrasion in 1957 because of a nasal skin lesion, which is best diagnosed as a rhinophyma on the basis of the available biographical literature. Throughout his creative life, the artist worked on different aspects of plastic surgery especially in the nasal area. As one of the founders of Pop Art, Warhol left essential marks on the modern society and influenced our standards of aesthetic shapes. PMID- 11103353 TI - [Slowly growing cheek tumor. Capillary hemangioma of the masseter muscle]. PMID- 11103354 TI - [Unusual submandibular tumor transformation. Myoepithelial carcinoma in a pleomorphic adenoma of the submandibular gland]. PMID- 11103355 TI - [Manifestations of Langerhans-cell histiocytosis in the ENT area]. PMID- 11103356 TI - Recent developments in community acquired pneumonia. PMID- 11103357 TI - National data sources on the care and outcomes of patients with community acquired pneumonia. AB - Despite improved understanding and treatment of community acquired pneumonia (CAP), variations in clinical practice and patient outcomes still exists, resulting in excess healthcare dollars spent and decreased patient satisfaction. The use of treatment and outcomes research data can help providers improve their methods and standardize techniques to control costs and provide the best care for their patients. To better understand the utility and capabilities of this research, this article will compare several administrative and clinical databases. Two, data sources in particular, the EPI-Q Inc. CAP-Compare database and the University HealthSystem Consortium's (UHC) CAP Benchmarking Program contain clinical and utilization data specific to CAP. In addition there are several government and non-government sponsored data sources that include administrative data on many diagnosis including CAP. These include: Healthcare Benchmarking Systems International's EXPLORE database, the Center for Healthcare Industry Performance Studies database, the National Center for Health Statistics' -National Hospital Discharge Survey, and the Medicare Provider Analysis and Review data files. PMID- 11103358 TI - Management strategies for community acquired pneumonia. AB - The past decade has witnessed dramatic changes in the etiology, diagnosis, and management of community acquired pneumonia (CAP). Due to the wide variation in practice patterns, physicians and professional societies have taken the lead in developing practice guidelines. To better understand the range of these multiple protocols, various stages of the disease and treatment are described and compared. Experts agree that CAP management should include: defining a correct diagnosis, identifying high risk populations in order to determine the severity of the disease, recommending appropriate treatment therapies, and obtaining positive and cost beneficial outcomes. PMID- 11103359 TI - Evaluation of community acquired pneumonia guidelines. AB - Community acquired pneumonia (CAP) is a serious illness, in which the etiology of disease cannot be determined in 50% of the patients. The American Thoracic Society (ATS) and the Infectious Disease Society of America (IDSA) have independently developed treatment guidelines for CAP. These guidelines, however, differ in their approaches. The differences can be attributed to their unique approaches. ATS favors the application of empiric methodology verse IDSA, which recommends a definitive clinical approach. Presently, a widely accepted, standardized guideline for the diagnosis and treatment of CAP is unavailable. This contributes to the large variation in admission rates, use of institutional resources, and quality of care given to CAP patients. PMID- 11103360 TI - Ongoing issues in pneumonia care: when to admit, how to treat and the role of oral therapy. AB - In recent decades the understanding of the diagnosis and treatment of community acquired pneumonia (CAP) has increased within the medical scientific community. The challenge remains how best to disseminate new information to the practicing clinician to facilitate improved outcomes and cost-effective care. To accomplish this end, professional organizations such as the Infectious Disease Society of America (IDSA) and the American Thoracic Society (ATS) have developed and published consensus guidelines reflecting the proposed standard of medical care. Three ongoing areas of ambiguity and divergence between published guidelines and clinical practice include treatment selection, the admission decision, and the utility and role of oral therapy. PMID- 11103361 TI - [Conversion from typical to atypical neuroleptics. Guidelines for ambulatory and inpatient treatment]. AB - The introduction of atypical antipsychotics has presented new options for the pharmacological treatment of schizophrenic patients. It is assumed that in the near future, neuroleptic medication will shift from conventional to atypical antipsychotics for an increasing number of patients. The present article addresses guidelines for the conversion from conventional to atypical neuroleptics in inpatient and outpatient settings. PMID- 11103362 TI - [Diagnosis of body dysmorphic disorder. A pilot study]. AB - The assessment of body dysmorphic disorder (BDD) is complicated by poorly defined diagnostic criteria and a lack of valid instruments. The present pilot study examines the possible benefit of specific instruments in the discrimination of other body image disorders. Thirteen patients with BDD, 13 with disfiguring defects, and 21 with no significant impairment of appearance participated in the study. The participants were recruited from dermatological outpatients on the basis of semistructured diagnostic interviews and clinical ratings of disfigurement. Furthermore, structured interviews were conducted to determine comorbidity with depressive disorders and social phobia and to obtain a clinical rating of obsessive-compulsive symptoms using the modified version of the YBOCS for body dysmorphic disorders. In addition, the Beck depression inventory, the Social Phobia Scale, and the Social Interaction Anxiety Scale were completed. Patients with BDD differed significantly from disfigured and unimpaired patients in obsessive-compulsive symptoms, in particular obsessions, and the discrepancy between personal and clinical rating of disfigurement. However, depression and social phobia scores were only higher in comparison with unimpaired patients. Comorbidity with depressive disorders was significantly increased. Our results support the utility of the specific diagnostic instruments and indicate the particular importance of obsessive-compulsive symptoms in BDD. PMID- 11103363 TI - [The Eppendorf Schizophrenia Inventory (ESI). Development and evaluation of a questionnaire for assessment of characteristic self perception of cognitive dysfunctions by schizophrenic patients]. AB - This study explored characteristic subjective experiences of schizophrenia. A questionnaire for self-assessment of disturbances in several cognitive and perceptual areas (the Eppendorf Schizophrenia Inventory, or ESI) was constructed and administered to first-episode schizophrenics (SCHe, n = 45), negative syndrome schizophrenics (SCHn, n = 45), remitted schizophrenics (SCHr, n = 24), depressives (DEP, n = 43), alcoholics (ALK, n = 48), obsessive-compulsive patients (ZWA, n = 46), and healthy controls (KON, n = 57). Comparisons between the SCHe, SCHn, DEP, ALK, and ZWA groups and a subsequent factor analysis revealed four schizophrenia-specific dimensions: attention and speech impairment (AS), ideas of reference (IR), auditory uncertainty (AU), and deviant perception (DP). Further analyses suggested that the AS syndrome may represent a mediating vulnerability factor while IR, AU, and DP probably are reversible episode indicators. The results may contribute to refinements in the measurement of specific prepsychotic signs, thus facilitating the development of early intervention approaches. PMID- 11103364 TI - [Identity measurement with the Frankfurt self concept scales and their relationship to parental bonding]. AB - Psychiatric inpatients and nonpatients were compared using the Frankfurt Self Concept Scales (FSKN). Firstly, we tried to validate FSKN as a measure of identity using the DSM-III-R identity rating as an external criterion. Then the FSKN ratings of patients and nonpatients were compared. Lastly, we correlated individual FSKN ratings with certain demographic variables, clinical diagnoses, and the two dimensions of the Parental Bonding Instrument (PBI). All FSKN scores of patients with identity disorders differentiated significantly from those without identity disorder, thus confirming FSKN validity. The most significant differences were found in practically all FSKN results comparing patients with nonpatients of both sexes, always in favor of nonpatients. Male patients presented better self-concepts than female patients, whereas practically no significant relationships were found between FSKN scales and sex in nonpatients or between FSKN, vocational category, and diagnosis. Also, the relationship between the scales and age appears to be slight, showing positive in patients and negative in nonpatients. Many significant correlations were found between FSKN scales and PBI dimensions which were negative between the scales and PBI "control" and positive between the scales and PBI "care", both especially in men with regard to their relationship with the father. PMID- 11103365 TI - [Differences between psychopathological evaluation in German and Turkish language of Turkish immigrants]. AB - The elevated rate of schizophrenia among migrants has been explained, among others, with misdiagnosis due to symptoms' being misinterpreted as psychotic. Previous studies have shown both higher and lower rates of psychotic symptoms of these patients when interviewed in their mother tongues, compared to being interviewed in the second language. In this study, 91 patients of Turkish origin and 50 of German origin with a paranoid-hallucinatory syndrome at admission were examined by an one interviewer each of Turkish and German origins using a standardized psychopathological instrument. In spite of comparative samples, correlation of psychopathological evaluation as well as diagnostic agreement between the two interviewers was significantly higher in the German patient group. Within the Turkish patient group, correlation was higher for those with good German language knowledge than for those with poor knowledge, yet only on a few items and without an effect on diagnostic agreement. The greatest difficulties lie in the evaluation of delusions. In spite of higher disagreement on psychopathology, the potential misdiagnoses cannot sufficiently explain the higher rate of schizophrenia among migrants. PMID- 11103366 TI - [Schizophrenic psychoses and suicide in the clinic. Risk factors, psychopharmacologic treatment]. AB - For all 5,352 patients treated for schizophrenia at the Psychiatric Hospital of the University of Munich in 1981 and 1992, detailed routine and data processing assisted documentations were made of the psychopharmacological therapies. Nineteen of the patients committed suicide while undergoing inpatient treatment; the control group consisted of all other patients (n = 5,333). More than 77 sociodemographic and anamnestic variables as well as 195 items from the admission summaries were taken into account while comparing the groups. Furthermore, the pharmacological data were classified according to drug groups and comparison was based on the mean frequency of prescription of each group. We analyzed the mean number of prescriptions for neuroleptics, tranquilizers, and antidepressants, which were further differentiated into sedating and nonsedating types. For frequently administered drugs, mean daily doses were also compared. Bivariate analysis of the data suggests that the suicide cases presented depressive signs, symptoms, and tendencies already present on admission more frequently than with controls; the same applies to previously attempted suicides. Discriminating analysis showed that the variables "feeling of loss of feelings," thought insertion," "visible depression," "free-floating anxiety," "suicidal tendencies," and "previously attempted suicide" have the greatest predictive value with respect to suicide, in descending order. No differences in psychopharmacological treatment between suicides and controls were found, apart from a significantly higher percentage of antidepressive treatments and a higher mean number of antidepressant prescriptions for the suicides. PMID- 11103367 TI - [The internet as a delusional topic in paranoid schizophrenia]. AB - Two patients with paranoid schizophrenia had delusions involving the internet. Additionally, one of them experienced computer databases as being distributed to other people in a phenomenologically similar way to that encountered in thought broadcasting. The presented cases illustrate the historical association of the contents of delusions in schizophrenia. PMID- 11103368 TI - [Community psychiatric management of severely ill schizophrenic patients. An exemplary case study]. AB - Psychiatric care for chronic schizophrenic patients has improved in recent years, but for severely ill patients, often with multiple comorbid psychiatric conditions and in great need of care, the situation remains unsatisfactory. Community-based psychiatric care is limited and psychiatric hospitals are now cutting back on inpatient care. The quality of care that chronically mentally ill patients receive at independent psychiatric nursing homes removed from population centers is undetermined, and many patients end up in asylums or homeless shelters unable to provide for their psychiatric needs. Neither a reliable way to assess the needs of these complicated patients nor a scientific basis for therapeutic intervention exists yet. The case study presented here illustrates an interdisciplinary approach to the care and treatment of a chronic schizophrenic patient with several comorbid conditions. Based on the individual needs of the patient and developed over the last 4 years, this approach had positive effects on the course and outcome of treatment and resulted in an impressive reduction of costs. The average annual number of days of hospitalization over a 3-year period was reduced from 206 to two and the total cost of care was reduced from US$70,000 annually to $8000. The results from this single case study are promising. Further investigation of the development and evaluation of highly individualized treatment plans for severely disturbed schizophrenic patients is warranted. PMID- 11103369 TI - [Acute tongue and pharyngeal spasms during changeover from clozapine to amisulpride. A case example]. AB - With the advent of atypical antipsychotics, quality of life for patients with schizophrenia has improved significantly. The positive effects are based not only on the compliance-enhancing reduction of extrapyramidal side effects but also due to improved cognitive function and social integration, shorter duration, and overall reduction of hospital treatment. Numerous controlled studies have addressed the issue of switching patients from typical to atypical antipsychotics. However, published data on substituting one atypical antipsychotic for another are preliminary and very limited. This case report describes acute side effects which occurred when switching from clozapine to amisulpride and discusses mechanisms on the receptor level. Regarding these two agents, the clinical relevance of the knowledge of receptor profiles is outlined. PMID- 11103370 TI - Readers disagree with erythropoietin alpha described as nontraditional treatment. PMID- 11103371 TI - Puget sound oncology nursing education cooperative. AB - The purpose of the Leadership & Professional Development feature is to provide readers with information, ideas, and exemplars of leadership competencies and professional roles in oncology nursing. Manuscripts submitted to the Leadership & Professional Development feature should be prepared according to the Information for Authors published in the Oncology Nursing Forum (ONF) but limited to six to eight double-spaced typed pages. Submit two copies of the manuscript using IBM compatible software along with a computer disk copy, or submit a copy of the manuscript as an e-mail attachment to Joan Such Lockhart, PhD, RN, CORLN, ONF Associate Editor, 1365 Simona Drive, Pittsburgh, PA 15201; lockhart /duq.edu (e mail). Manuscripts should be referenced and include tables, figures, or illustrations as appropriate. Ideas for possible manuscripts are welcome. PMID- 11103373 TI - Prophylactic mastectomy and genetic testing: an update. AB - PURPOSE/OBJECTIVES: To examine and discuss the possible benefits and difficulties with recommending prophylactic mastectomy to BRCA1- and BRCA2-positive women. DATA SOURCES: Published research articles, professional review articles, textbooks. DATA SYNTHESIS: Women with BRCA1 and BRCA2 mutations face a much higher risk of developing breast cancer than the general population, with limited options available for prevention. Prophylactic mastectomy has been shown to have a survival advantage in young women who carry BRCA1 and BRCA2 mutations. Challenges exist, however, in the use of prophylactic mastectomy and genetic testing. CONCLUSIONS: Methods of preventing breast cancer in BRCA1- and BRCA2 positive women currently are limited to watch-and-wait surveillance, prophylactic mastectomy, and, perhaps, chemoprevention. Genetic testing and prophylactic mastectomy each present unique challenges while offering certain benefits as well. Recent studies have shown survival advantages to BRCA1- and BRCA2-positive women who undergo prophylactic mastectomy. IMPLICATIONS FOR NURSING PRACTICE: Nurses need to be aware of the complex issues surrounding testing for BRCA1 and BRCA2 mutations and prophylactic mastectomy to be able to provide current information to patients and assist in decision making. PMID- 11103372 TI - Tobacco-control attitudes, advocacy, and smoking behaviors of oncology nurses. AB - PURPOSE/OBJECTIVES: To describe oncology nurses' attitudes, smoking behaviors, and involvement in tobacco-control policy and legislation. DESIGN: Descriptive, cross-sectional survey. SAMPLE: Responses from 1,508 (38% return) of 4,000 randomly selected members of the Oncology Nursing Society (ONS). The typical respondent was female, age 44, a staff nurse, had practiced as an oncology nurse for 12 years, and was certified as an OCN. METHODS: A mailed survey with specific questions about smoking status and the importance of nursing involvement in healthcare policy and legislation for tobacco control. MAIN RESEARCH CONCEPTS: Attitudes about tobacco-control policies and legislation; sociodemographic, professional, and institutional variables; and tobacco use. FINDINGS: The majority (85%) of members stated that nursing involvement in tobacco-control healthcare policy and legislation was important. More than 90% of respondents supported prevention of youth access to tobacco and providing information about health effects of tobacco and cessation. Seven percent (n = 106) were current smokers. Significantly fewer smokers valued involvement in tobacco-control activities. Respondents with personal experience of tobacco-related illnesses were more likely to value involvement in advocacy activities. Educational programs to prevent tobacco use among youth and to help patients stop smoking received the most support (80%). IMPLICATIONS FOR NURSING PRACTICE: This sample of ONS members strongly supported involvement in tobacco-control policies and legislation. Smoking was associated with more negative attitudes about the importance of actively engaging in tobacco control. These oncology nurses recognized the need for additional educational programs to prevent tobacco initiation by youth. PMID- 11103374 TI - Quality of life, survivorship, and psychosocial adjustment of young women with breast cancer after breast-conserving surgery and radiation therapy. AB - PURPOSE/OBJECTIVES: To examine changes in quality of life (QOL), psychosocial adjustment, and survivorship issues over time of women younger than 45 years who underwent breast-conserving surgery and radiation therapy (RT) for breast cancer. DESIGN: Repeated measures, longitudinal design. METHODS: Data were collected at four time points: start of RT, midpoint of RT, end of RT, and six months after RT. Three instruments were used to collect data: Quality-of-Life Index, Psychosocial Adjustment to Illness Scale, and the newly developed Adaptation to Survivorship Experience. Subjects also participated in an indepth interview at the start of RT. SETTING: A large radiation oncology department located in an urban teaching hospital in the Northeast United States. SAMPLE: 23 women with newly diagnosed stage I or II breast cancer who were starting RT following breast conserving surgery, with a mean age of 37.8 years (range = 25-45 years). MAIN RESEARCH VARIABLES: QOL, psychosocial adjustment, and adaptation to survivorship experience. FINDINGS: Although subjects adjusted their lives to accommodate RT, QOL declined from the start of RT to midpoint, with gradual improvement reported six months later. Social and sexual adjustment declined from start of RT to six months later. Negative perceptions of the survivorship experience and worry about cancer increased from the start of RT to six months later. CONCLUSIONS: Young women with breast cancer experience changes in QOL, psychosocial adjustment, and adaptation to survivorship issues during RT. Changes may not reflect what is observed in clinical practice. IMPLICATIONS FOR NURSING PRACTICE: Nurses need to be aware of changes in QOL, psychosocial adjustment, and survivorship to better understand and support young women during RT. PMID- 11103375 TI - The effects of standard care counseling or telephone/in-person counseling on beliefs, knowledge, and behavior related to mammography screening. AB - PURPOSE/OBJECTIVES: To determine the most effective methods of increasing mammography adherence while also considering ease of intervention delivery in evolving healthcare systems. DESIGN: Experimental. SETTING: Women from a health maintenance organization and a large general medicine practice. SAMPLE: Women 50 85 years of age who had not had breast cancer and did not have a mammogram within the last 15 months. METHODS: Once consent and baseline information were obtained, women were randomized to receive in-person, telephone, or no mammography counseling. MAIN RESEARCH VARIABLES: Mammography adherence, perception of susceptibility to breast cancer, and benefits, barriers to, and knowledge of mammography. FINDINGS: Compared to standard care, telephone counseling was more than twice as effective at increasing mammography adherence, whereas in-person counseling resulted in almost three times the mammography adherence postintervention. Both telephone and in-person counseling are successful in changing perceived susceptibility, knowledge, barriers, and benefits. CONCLUSION: Both telephone and in-person counseling interventions were successful in changing beliefs, which, in turn, increased mammography adherence. IMPLICATIONS FOR NURSING PRACTICE: Interventions based on altering beliefs are effective for increasing mammography adherence. PMID- 11103376 TI - Nutritional status of Korean Americans: implications for cancer risk. AB - PURPOSE/OBJECTIVES: To examine nutrient intake of Korean Americans, especially those foods and supplements implicated in cancer. DESIGN: Cross-sectional survey and descriptive analysis. SETTING: Chicago, IL. SAMPLE: 103 Korean Americans who were between 40 and 69 years of age. METHODS: An Instrument, culturally and linguistically adapted from the Health Habits and History Questionnaire, was administered to assess nutrient intake from food and vitamin and mineral supplements. Bilingual interviewers collected data at respondents' homes. FINDINGS: Relative to their diet in Korea, more than one-third of the respondents reported an increase in the consumption of beef, dairy products, coffee, soda, and bread, as well as a decrease in the intake of fish and rice and other grains. Compared to the general U.S. population included in the National Health Interview Survey (NHIS), Korean Americans had a greater intake of carbohydrates and vitamins A and C and lower intake of total fat, cholesterol, and saturated fat. Moreover, the percentages of calories were higher from carbohydrates and lower from fat, sweets, and alcohol for Korean Americans than those reported by NHIS respondents. Gender, education, and marital status were significantly associated with nutrient intake. The use of daily vitamin and calcium supplements was similar between respondents and those from NHIS. CONCLUSIONS: At their stage of cultural adaptation, the incorporation of a larger quantity of Western food items did not make for a less healthy dietary pattern among respondents. Data showed that Korean Americans continued to consume diets more consistent with Korean than with American food patterns, in as much as greater than 60% of their calories came from carbohydrates and about 16% of calories from fat. As a group, respondents met the recommended dietary guidelines for most nutrients, except for dietary fiber and calcium. IMPLICATIONS FOR NURSING PRACTICE: Variation in dietary intake by age, culture, gender, and years in the United States is well accepted. Effective cancer prevention and initiatives for dietary reform call for the incorporation of available research findings and considerable attention to data gaps regarding Korean Americans and other Asian Americans and Pacific Islander populations. Culturally competent, community-based programs should include the reinforcement of positive traditional dietary habits, encourage the adaptation of healthy Western food items, as well as assist minority populations in developing strategies that will effectively correct likely deficiencies in diet. PMID- 11103377 TI - Breast cancer screening knowledge, attitudes, and practices among Korean American women. AB - PURPOSE/OBJECTIVES: To describe the knowledge and beliefs about breast cancer and breast cancer screening and practices of clinical breast examination (CBE) and mammography of Korean American women. DESIGN: Cross-sectional survey. SETTING: Two Korean churches in a mid-sized Southeastern U.S. city. SAMPLE: A convenience sample of 107 Korean women ages 40 and older. METHODS: Data were collected using Champion's Health Belief Model instrument (susceptibility, seriousness, benefits, and barriers) and the Breast Cancer Knowledge test through mailed questionnaires. MAIN RESEARCH VARIABLES: Knowledge and beliefs about breast cancer screening and practices of CBE and mammography. FINDINGS: The percentages of Korean American women who ever had a CBE and mammography were 67 and 58, respectively. Among the Health Belief Model variables, women who never had a CBE had significantly lower knowledge scores and higher perceived barriers to CBE than those who had. Women who never had a mammogram reported significantly higher perceived barriers to mammography. Logistic regression analyses demonstrated that husband's nationality, regular checkups, and encouragement from family members and physicians were significant predictors of CBE and mammography use. CONCLUSIONS: The frequency of breast cancer screening practices among Korean American women is below national objectives. IMPLICATIONS FOR NURSING PRACTICE: As healthcare professionals in a culturally diverse nation, nurses need to increase their awareness of cultural variations and provide culturally and linguistically appropriate breast health education. Additional studies with women from a variety of settings are needed to validate present study findings. PMID- 11103378 TI - Hospice and hospital oncology unit nurses: a comparative survey of knowledge and attitudes about cancer pain. AB - PURPOSE/OBJECTIVES: To identify knowledge strengths and weaknesses and misperceptions about cancer pain management between two groups of registered nurses in different settings. DESIGN: Descriptive, comparative survey. SETTING: 11 community-based hospices and 7 inpatient hospital oncology units within an urban county. SAMPLE: A convenience sample of 30 hospice and 34 hospital oncology unit nurses. Sample criteria included registered nurses who had worked for at least the preceding six months exclusively in either a hospice or hospital oncology unit. METHODS: The North Carolina Cancer Pain Initiative survey and a demographic survey were distributed to the work mailboxes of nurses in the participating facilities who met the inclusion criteria. MAIN RESEARCH VARIABLES: Hospice and hospital oncology unit nurses' knowledge and attitudes about basic pharmacologic cancer pain management. FINDINGS: Hospice nurses scored significantly higher than hospital oncology unit nurses regarding overall pain management knowledge, opioids, scheduling, and liberalness. Hospice nurses also reported more pain education and a higher frequency of pain guideline review requirements than hospital oncology unit nurses. CONCLUSIONS: The most prevalent knowledge deficits concerned opioids. Practice setting and pain education may influence knowledge, as well as attitudes, about pain. IMPLICATIONS FOR NURSING PRACTICE: Further research is needed regarding nurses' pain management behavior and outcomes of pain management education in various settings. PMID- 11103379 TI - Experience of palliative home care according to caregivers' and patients' ages in Hong Kong Chinese people. AB - PURPOSE/OBJECTIVES: To identify the relationship between family caregivers' reported difficulty in managing caregiver tasks and ages of caregivers and patients. DESIGN: Cross-sectional, descriptive survey. SETTING: A hospice homecare program in Hong Kong. SAMPLE: Twenty-nine Chinese family caregivers who had experienced at least weekly caregiving responsibility for more than two months and were able to read and understand Chinese. METHODS: Respondents completed a caregiver task inventory. Four homecare nurses assisted in the distribution and collection of questionnaires. MAIN RESEARCH VARIABLES: Caregiver tasks and age and patient age. FINDINGS: Caregivers' age was negatively correlated with reported difficulty in overall tasks and in interpersonal ties. The patients' age was negatively correlated with reported difficulty in direct care to patients, intrapersonal tasks, and overall tasks. CONCLUSIONS: The younger the caregiver, the more difficulty he or she experienced in the caregiving role, particularly in the maintenance of social and family ties. Caregivers of younger patients experienced more difficulty in most aspects of caregiving tasks. More research with a larger sample size is required to fully investigate the effect of age on the family caregiving experience and the validity of the caregiver task inventory. IMPLICATIONS FOR NURSING PRACTICE: Nursing support and preparation to younger caregivers and caregivers of younger patients are suggested in the practice of palliative home care. PMID- 11103380 TI - Development of a method for clinical medication review by a pharmacist in general practice. AB - Medication review of patients on long-term treatment in general practice in the UK has been reported to be inadequate. Proposals followed suggesting that pharmacists could use their expertise to lead such a medication review in conjunction with the general practitioner. This paper describes the concept of clinical medication review by a pharmacist based in general practice. We describe the development of a method for a structured and systematic process for undertaking such a review in clinics conducted by a pharmacist. The method was developed for a nationally funded study in the UK. We provide a definition of clinical medication review and suggests a structure for the process through data gathering, evaluation and implementation. PMID- 11103381 TI - Pharmacokinetics of intraventricularly administered teicoplanin in Staphylococci ventriculitis. AB - Following craniotomy for a medulloblastoma in the posterior cranial fossa, a 6 year old girl developed a ventriculitis with coagulase negative staphylococci associated with the use of a ventriculostomy. Treatment with intravenous (i.v.) and intraventricular (ivt) vancomycin resulted in negative cultures of the cerebrospinal fluid, but had to be stopped because of a severe allergic skin reaction. Teicoplanin was administered i.v. (240 mg once daily) and ivt (10 mg once daily), resulting in high teicoplanin CSF levels that were used to model the pharmacokinetics of ivt teicoplanin in this patient. No signs of recurrent infection or adverse events occurred. It is concluded that a pharmacokinetic model can be derived from this case that can be used as prior to guide teicoplanin intraventricular therapy in other patients. PMID- 11103382 TI - Community pharmacists' views and beliefs about the treatment of symptoms suggestive of vaginal thrush in community pharmacies. AB - OBJECTIVE: To investigate the views and beliefs of community pharmacists about the benefits and disadvantages to the customer, pharmacy and pharmacist of treating women with symptoms suggestive of vaginal thrush. DESIGN: Semi structured interviews. SETTING: Community pharmacists from within Grampian Primary Care NHS Trust. OUTCOME MEASURES: Pharmacists' views and beliefs analysed using content analysis. RESULTS: Of the 26 pharmacists contacted, 19 (73%) pharmacists from 16 pharmacies completed interviews. The pharmacists were generally positive towards the treatment of women with vaginal symptoms and perceived few disadvantages. Immediate access to treatment and rapid symptom relief were perceived to be the greatest advantages to the customer. The main problems were customer embarrassment, cost and the risk of masking a serious condition. Customer embarrassment was perceived to be influenced by lack of privacy and the gender of the member of staff involved in the consultation. Five pharmacists perceived vaginal thrush to be an infection that could be spread by sexual transmission. DISCUSSION: There is a need to make pharmacists aware of the current evidence regarding the treatment of vaginal thrush, particularly the sexual partners of women with acute, uncomplicated thrush do not require treatment with an antifungal. The main difficulties that community pharmacists reported with the treatment of this condition were obtaining an accurate history and this was influenced by customer embarrassment. The gender of pharmacy staff and lack of private consultation facilities were suggested as factors that are associated with customer embarrassment and hence, the ability to obtain an accurate history. PMID- 11103383 TI - Television advertising of pharmacy medicines in the United Kingdom. AB - In the UK pharmacy is often promoted as the first port of call for minor ailments and the pharmacist's armamentarium of products is increasing as further products are reclassified from prescription only control. Non-prescription medicines in the UK can be advertised directly to the public but the advertisements must comply with the law. Some medicines, namely pharmacy can only be sold from pharmacies by, or under, the supervision of a pharmacist. This report describes the major findings of a postal survey of community pharmacists on the subject of advertising of pharmacy medicines and put them into context by outlining the legal and quasi-legal requirements. Further other studies concerning the advertising of non-prescription medicines are discussed in the context of requests for, and sales of, medicines available without a prescription. PMID- 11103384 TI - Sex differences in the medication choice for hypertension in general practice. A study with written case simulations. AB - The objective of this study was to explore explanations for the preference of physicians to prescribe beta-blockers to hypertensive men and diuretics to hypertensive women. A qualitative study among 12 family physicians was conducted with a combination of written case simulations, semi-structured interviews and statements on attitudes of physicians towards antihypertensive drug choice. Among the male hypertensive cases the most frequently prescribed drugs were beta blockers, whereas among the female hypertensive cases diuretics were more often prescribed. Physician characteristics associated with a preferred prescribing of beta-blockers to hypertensive men and diuretics to hypertensive women were: older age (no residency in family medicine), the believe that beta-blockers are more effective in men with regard to lowering blood pressure and that diuretics are more effective in women, a non-evidence based attitude and a sex-related attitude towards the choice of beta-blockers and diuretics in general, and in particular towards the prescribing of beta-blockers to hypertensive men because men have a higher absolute risk of coronary heart disease than women. An additional explanation for these findings may be the higher prevalence of ankle oedema among women. Patient characteristics associated with more prescribing of beta-blockers to hypertensive men and diuretics to hypertensive women were: current employment and a "high-risk" profile in terms of blood pressure level and additional cardiovascular risk factors. Although, most considerations underlying a preferred prescribing of beta-blockers to hypertensive men and diuretics to hypertensive women were not evidence-based, the actual choice of antihypertensive drug (diuretic or beta-blocker) was evidence-based. These considerations may also play a role in the sex difference in the choice of calcium antagonists and angiotensin converting enzyme inhibitors and require further investigation. PMID- 11103385 TI - Stabilization of oral anticoagulant therapy in hospitalized patients and characteristics associated with lack of stabilization. AB - The initiation and stabilization of oral anticoagulant therapy in hospitalized patients in a setting without specialized medical or pharmaceutical advice, was studied. In addition, potential risk factors for lack of stabilization were studied. All patients from three wards (orthopaedic surgery, general surgery and internal medicine) in two Dutch hospitals, who were started on oral anticoagulant therapy and who gave informed consent, were included in this three months prospective follow-up study. When a patient had two consecutive INR's within the range 2-3 during hospitalization (on day 6 or later), he was defined as stable. Stable and unstable patients were compared with respect to age, gender, quetelet index, length of hospital stay, indication for oral anticoagulant therapy, induction dosing schedule of oral anticoagulant therapy, prescribing physician, type of hospital (teaching or non-teaching), concurrently used drugs, concurrently used drugs known to potentially interact with oral anticoagulant therapy (drug-drug interactions that influence INR) and (co)morbidity. A total of 125 patients, who all used acenocoumarol as oral anticoagulant, were recruited in the study. The study population mainly comprised orthopaedic discharges on prophylactic oral anticoagulants. The mean length of hospital stay was 14.5 days (median 11.0, standard deviation (SD) 10.2) for the patients included in the study (patients with a short length of stay < 6 days were excluded from the study, because of the definition of stability). 43 patients (34%) became stable during hospitalization. The second INR within the range was reached after on average 11.1 days (median 10.0, SD 4.5). 18 different induction dosing schedules were used. Differences in apparent risk of INR instability were statistically associated with length of hospital stay (odds ratio (OR) 0.85, 95% confidence interval (CI) 0.78-0.92), concurrent use of musculoskeletal drugs, mainly NSAIDs, (OR 1.68, 95% CI 1.04-2.72) and two individual prescribing physicians (OR 6.61, 95% CI 1.47-29.82 for one physician and OR 0.23, 95% CI 0.06-0.99 for the other physician). This population has a high percentage of instability and reaches stability relatively late. The instability was associated with length of hospital stay, the concurrent use of musculoskeletal drugs (mostly NSAID's) and physician. Most of the unstable patients had INR's below therapeutic range, suggesting a conservative dosing habit. Part of the instability may also be due to the many different physicians who dose their own patients. Interventions to improve dosing may aid in better stabilization in hospitalized patients and thus in reduced length of hospital stay. PMID- 11103386 TI - Rational pharmacotherapy and clinical practice guidelines. Theories and perspectives on implementing pharmacotherapeutic treatment guidelines. AB - Several theories behind implementing clinical guidelines have been described within the literature. At first sight, these may seem different. However, there are similarities and eventually they are rather complementary than mutually exclusive. This article integrates several theoretical views on implementation of pharmacotherapeutic treatment guidelines and subsequently addresses some empirical considerations. Furthermore, specific limitations and potential harms of implementing guidelines are addressed. Several checklists are provided in order to make pharmacists, pharmacologists and clinicians aware of perspectives on dissemination, implementation, and subsequent application of pharmacotherapeutic treatment guidelines. PMID- 11103387 TI - Desire for information about drugs. A multi-method study in general medical inpatients. AB - The purpose of this paper is to investigate patients' drug information preferences using a combination of quantitative and qualitative methods. Patient interviews (n = 299) were conducted on general medical wards in three London teaching hospitals. The purpose was to refine and validate a quantitative 12-item scale, the Intrinsic Desire for Information (IDI), by interfacing quantitative and qualitative data, and to explore the relationship between this scale score and patient demographics. The IDI-scale was subjected to factor analysis. Two secondary factors were found in the IDI scale; a 5-item factor describing the extent of information desired and a weaker 3-item factor describing an inhibited desire for knowledge about illness/drugs. Reliability analysis and multiple regression analysis were undertaken. Responses to open answer questions during the qualitative interviews were transcribed at the bedside and imported into QSR NUD*IST software program for coding and analysis. The methodology employed in this study involved importing quantitative, summative data into a qualitative data base and re-analysing both the quantitative and qualitative data to validate the scale. Age was a predominant factor associated with patient desire for information, although the data suggest that educational and socio-economic status are also influential. Factor 1, the extent of information desired, may have value in targeting receptive patients, or in identifying those who may be refractory to drug information. The refined tool could help health services to effectively target information provision based on evidence, rather than supposition. PMID- 11103388 TI - Rectal absorption of oxcarbazepine. AB - Physicians regularly challenge the hospital pharmacy departments to find alternative routes for the administration of drugs, which can't be withhold, e.g. anti-epileptic drugs. In our hospital we were confronted with the question whether it was possible to administer oxcarbazepine rectally. In the present report data on the absorption of rectally administered oxcarbazepine is presented. No therapeutic bloodlevels were attained after rectal administration. Administration via the oral route, however, gave within the same period of time a therapeutic bloodlevel. It is concluded that the absorption after rectal administration of oxcarbazepine at least in this dose and frequency used is too low to justify application in practice. PMID- 11103389 TI - [Stigmatization of the mentally ill: is it our problem?]. PMID- 11103390 TI - [Stigma, stigma management, destigmatization]. AB - WHO and WPA recently inaugurated worldwide "anti-stigma-campaigns". On this background the exploration discrimination of against the mentally ill of the term stigma and its meaning is useful. Stigma dates from ancient Greece and was meant to be a visible sign of disgrace. Today stigma and stigma management are sociological terms described by the American sociologist Ervin Goffman (1963) in is essay "Stigma--notes on the management to spoiled identity". Changes and hazards of attempts of destigmatization are explored. PMID- 11103391 TI - [Colloquial terminology in German depicting mentally ill and odd people. Etymologic-sociolinguistic review]. AB - OBJECTIVE: This paper investigates the picture of the mentally ill in German colloquial language. METHOD: The empirically collected single words and phrases were subject to systematic semantical, linguo-historical as well as etymological investigations, the results of which were then contrasted with selected psychiatric concepts of the 18th/19th century. RESULTS: Most of the words found aim at the head or brain which are often compared with a mechanism and regarded as the actual starting point of the disease. Other terms refer to blows or other mechanical impressions as the cause of mental illnesses. There is another group of words depicting weakness or even total loss of mind. Some of the ideas expressed by the words or structures analysed coincide with concepts psychiatrists had in the past. Many of the words even had been terms in psychiatric theory before they became part of the everyday language after having been loaded with negative connotative meanings making them inappropriate for being a scientific term. Some lexemes have been taken over from other languages, above all from English. CONCLUSION: Language mirrors stigmatisation of mentally ill people by society in an extraordinarily drastic way. Thus the colloquial names for them share the same fate as the words for other--ethnic, sexual, whatsoever--minorities: by pure means of language those people are regarded as somewhat strange, not belonging, something negative one does not wish to have contact with. On the other hand however, there have always been attempts to counter this by replacing negatively connotated words by other, neutral or even positive ones. Thus was the introduction of the new word nervenkrank ("ill in the nerves", cf English brainsick) in the 19th century, making mentally ill people even to bodily ill ones, what meant an enormous enhancement in value for both the patients and the psychiatrists. Today's name Betroffene ("affect-ed") is another example for those attempts. As can be seen from the historical retrospective this had been the case in former times as well. PMID- 11103392 TI - [Public image of psychiatry. Results of a representative poll in the new federal states of Germany]. AB - RESEARCH PROBLEM: The readiness to use psychiatric services is, among other things, determined by the image of psychiatry held by the public. METHOD: A representative survey of the adult population in the new German Lander was carried out, part of which was the assessment of what people associated with the term "psychiatry" and of their idea of a psychiatric hospital. RESULTS: Psychiatric hospitals, in particular the large-scale mental institutions, shape the dominant image of psychiatry among the public. In this, the custodial or repressive character of psychiatry is the central constituent of the public's representation. DISCUSSION: The chances that a positive change of the public's ideas toward more informed images and attitudes might be achieved appear to be slim as long large scale institutions, though with a new appearance, continue to exist. PMID- 11103393 TI - [Depression and stigma]. AB - AIM: Until now, the interest of stigma research in psychiatry has been predominantly focused on schizophrenia. The fact that individuals suffering from depression may also be exposed to stigmatization, on the other hand, has received little attention. METHOD: Using the scales for the assessment of perceived stigma and stigma coping developed by Link et al., 39 patients who had received in patient treatment for a depressive episode were questioned 4-7 months after their discharge from hospital. RESULTS: 45% of the respondents reported concrete instances of stigmatization. Respondents expected to find discrimination and devaluation on the part of their environment especially with regard to their changes on the labour market. The vast majority of those questioned was in favour of keeping the fact that one had experienced a mental illness or had received psychiatric treatment to themselves. Further, our results show a tendency to avoid situations which might imply an increased risk of stigmatization. While respondents considered it important to facilitate a better understanding of mental illness on the part of their closest friends and relatives, the idea of being actively involved in educating the lay public received comparatively little support. CONCLUSION: Although individuals suffering from depression may be confronted with less rejection or resentment than schizophrenic patients or patients with substance use disorder, this does not imply that they do not have to struggle with the problem of stigmatization. PMID- 11103394 TI - [Mentally ill and dangerous? Attitudes of female journalists and medical students]. AB - OBJECTIVE: The realisation of community mental health services may be adversely affected by society's prejudices about mentally disordered people. In this regard, the frequently replicated stereotype that people with mental illness are dangerous is of paramount relevance. METHODS: Journalists and medical students were asked by means, of an interview whether they consider certain crimes (murder, rape, arson, larceny, disorderly conduct) to be more frequently committed by mentally disordered persons or by the general population or whether no difference is supposed. RESULTS: Journalists and medical students agreed about the frequency of commitment concerning offences such as murder (no difference between the mentally disordered and the general population), arson (more frequently committed by mentally disordered persons) and larceny (more frequently committed by the general population). While almost half the journalists supposed that there is no difference between mentally disordered persons and the general population regarding rape and disorderly conduct, just as much medical students charged mentally disordered persons with these offences. Personal contact to people with mental disorders slightly influenced these attitudes to the credit of mentally disordered persons. CONCLUSIONS: Compared with the results of a representative survey among the general population of the old Lander of Germany in 1990, journalists asked in our investigation markedly less often charged mentally disordered people with murder, rape and disorderly conduct. Medical students, in contrast, shared the general population's attitude except for murder. PMID- 11103395 TI - ["...not dangerous, but nevertheless frightening". A program against stigmatization of schizophrenia in schools]. AB - OBJECTIVE: Evaluation of a brief school programme providing information about schizophrenia to high school students. It was investigated whether the programme changed the students' attitude towards this target group and whether information given by a patient affects the results. METHODS: 114 high school students (6 school classes) took part in the programme: 57 students were informed by a psychiatrist and an afflicted person (group A), the other 57 were informed by a psychiatrist and a social worker (group B). The students' attitude towards schizophrenics was assessed using a vignette of a fictitious class-mate suffering from schizophrenia (according to DSM criteria) and by questionnaires assessing the students' emotional and cognitive reactions and their social distance towards the person described by the vignette. RESULTS: Only in group A (psychiatrist and afflicted) there was a significant improvement regarding the emotional reaction (reduction of fear, increase in positive emotions) and a significant decrease in social distance. Moreover, students no longer associated psychiatric illness with being "crazy". In group B (psychiatrist and social worker) these changes could not be observed. There was even a significant increase in describing patients with schizophrenia as dangerous. CONCLUSION: As high school students themselves consider their level of knowledge about psychiatric illness as low, information about this topic should be given more attention than is currently done (also from a preventive point of view). For such information to be effective and to affect students' attitudes positively, it appears necessary that students have the opportunity to get in contact with a person affected by psychiatric illness. For changing attitudes it is also essential how information is presented. PMID- 11103396 TI - [Volunteering in psychiatry: determining factors of attitude and actual commitment]. AB - OBJECTIVE: To assess public attitude, actual working commitment and the respective influence of demographic, psychological and sociological variables on voluntary help in psychiatry. METHODS: Multiple logistic regression analysis of the results of a representative population survey in Switzerland. RESULTS: Public attitude is mostly positive, but the respective working commitment is small. Attitude depends on gender, psychological factors (social distance, stereotypes), and on attitude to community psychiatry. For the working commitment, clearly distinct predictors are found: age, emotions, participation, and perceived discrimination to the mentally ill. For both attitude and commitment, having a social profession and interest in mass media are predictors. CONCLUSIONS: Internationally compared, Switzerland has a positive attitude and a big commitment in lay helping in psychiatry. But attitude is different from actual commitment. Lay helpers' work must be limited to realizable tasks and they need professional recruitment, instruction, and supervision otherwise they tend to be over-burden. The unused potential of voluntary helpers has to be opened specifically, e.g. by involving mass media and opinion-makers. PMID- 11103397 TI - [Psychological education of inpatients]. PMID- 11103398 TI - [Psychiatry in print media. Information acquired through reading of the daily papers]. PMID- 11103399 TI - [Current German laws for foreigners as a contributory factor in the treatment of a first-time acute schizophrenic episode]. AB - In a case report we describe an 18 year old Croat refugee with a first episode schizophrenia to discuss the current German laws for foreigners as a possible factor for the maintenance of the disease. Short residence permits, the release of the professional discretion to the authorities, deficient information about the laws and the unclear situation in the native country of the patients can lead to a worst course and prognosis of the disease. PMID- 11103400 TI - [Fools, simpletons, crazies. History of contradictory societal attitudes towards the mentally ill]. PMID- 11103401 TI - Observations on some aspects of current psychoanalytic theories. AB - The competing theories in the psychoanalytic marketplace today should be judged on their merits, not on the basis of the authority of whoever first proposed them. What is valid in each theory should be included in any formulation of a psychoanalytic theory of mental development and functioning. Since psychoanalysis, as part of psychology, is a branch of natural science, pluralism in theory is to be avoided in psychoanalysis as in every branch of science. The psychoanalytic method is a valid one of studying a particular aspect of brain functioning. The method and the theories based upon it are as "organic" as is the case with any of the other neurosciences. Any valid psychoanalytic theory of mental functioning and development should include the following conclusions: (1) Unconscious mental processes are omnipresent and of great importance in mental functioning; (2) Thoughts are as causally related to one another as are other events in the universe; (3) Mental functioning is a developmental phenomenon with describable, sequential features; and (4) A major role in mental functioning and development is played by conflicts over the sexual and aggressive wishes that characterize mental life during the period from three to six years of age, and by the compromise formations that result from those conflicts. The last of these conclusions, though disputed by many, is abundantly supported by evidence that is not dependent on the use of the psychoanalytic method, as well as by evidence furnished by the use of the psychoanalytic method. There is also much evidence to support the assertion that any psychoanalytic theory that attributes language dependent thoughts to a child whose brain is not yet mature enough to be capable of language is to be considered invalid, as are any observations made by the psychoanalytic method (= clinical observations) that are influenced by such an invalid theory. In psychoanalysis, as in every other branch of science, an observer--no matter how astute and how experienced--who subscribes to an invalid theory will be led astray by that theory, sooner or later, in one way or another. PMID- 11103402 TI - The empty mother: women's fear of their destructive envy. AB - This paper explains the importance of understanding the little girl's envy of her mother and how the resolution of this envy (and her fear of other women's envy) is crucial to a woman's development. I postulate that envy is a universal part of female development (with more or less destructive effects on a woman's personality, depending on the libidinal/sexual components of her attachment to both parents). I hope to show that by interpreting a woman's fear of her destructive envy, one can free her not only to enjoy her own sexuality and to find appropriate ways to express her aggression, but also to be more creative. I believe that guilt about these envious feelings often leads to profound inhibitions and masochistic behavior. Two clinical examples illustrate how envy manifests itself in treatment with a woman analyst, and how the working through of intense envious feelings leads to a greater ability to enjoy one's own capacities without constant fear of retribution. PMID- 11103403 TI - On lying and the lie of a toddler. AB - For most of its history, the psychoanalytic literature on lying dealt exclusively with the dynamic, genetic meanings of lying: the problems for treatment presented by a patient who lies, and the technique used in dealing analytically with lies. In recent decades, issues relating to the moral and general development of children in relation to lying have been considered. In this paper, a lie told by a 21-month-old child is used to raise and explore questions about lying and its relation to intrapsychic structure and development. It is suggested that cognitive abilities and the psychic apparatus have to develop to the point that self can be distinguished from object, and a superego prototype must be present, before the means and motivations for lying are in place. This would date the beginning of the capacity to lie to sixteen to twenty-four months of age. PMID- 11103404 TI - Subjective reality, objective reality, modes of relatedness, and therapeutic action. AB - This paper describes a dialectic believed to be at the heart of therapeutic interaction within a relational model. The dialectic consists of the interrelationship of two modes: the dyadic and the triadic. In the dyadic mode, the analyst responds with aspects of his or her self that singularly reflect the patient's subjectivity. This mode of attunement is uniquely suited to bringing the patient's experience into a place where it can then be seen and known. In the triadic mode, realities are recognized that are important to, but still outside of, the subjectivity of the patient. The analyst invites the patient to see him- or herself not only from inside his or her own space, but also from a point outside, through the perspective of others. The analyst is charged with asymmetric but not exclusive responsibility for negotiating and sustaining a fluid and flexible relationship between these modes. Optimally, this occurs through spontaneous and authentic engagement informed by intuition, empathy, and clinical judgment. However, when this dialectic loses its robust and kinetic quality (as frequently occurs in approaches ranging from the classical to the postmodern), an impermeable dyad is formed by extruding potentially triangulating aspects of reality (and subjectivity). This can result in curiosity and the openness of uncertainty being replaced by closed-mindedness and proclamation. PMID- 11103405 TI - At one with death: destructive narcissism. AB - In this paper, narcissism is considered to be the relation of self with an idealized internal object, and Narcissus's romance with his reflection is taken to be a two-party affair. Destructiveness, an inborn capability, is distinguished from destructive narcissism, a two-party situation between the self and a sadistic internal figure built on the idealization of power. Too often, only half the narcissistic pair is analyzed. The internal object becomes the persecutor of self, while the sadism of self, projected onto the persecutor, goes unanalyzed. This paper takes up a clinical solution: how the analysis can seize the destructive internal object and resolve it down to its nucleus, the self. PMID- 11103406 TI - The roots of violence: converging psychoanalytic explanatory models for power struggles and violence in schools. AB - This paper demonstrates that several psychoanalytic models taken together converge to collectively explain school violence and power struggles better than each does alone. Using my own experience in doing psychoanalytically informed community intervention, I approach the problem of school violence from a combination of Adlerian, Stollerian, dialectical social systems, and Klein-Bion perspectives. This integrated model is then applied to the Columbine High School massacre in Littleton, Colorado. PMID- 11103407 TI - [Functional magnetic resonance tomography in neuroradiology]. AB - The assessment of cerebral functions has long been the domain of positron emission tomography and single photon emission computed tomography. The use of rapid imaging sequences and contrast agents enables physiological and pathophysiological cerebral processes to be assessed and monitored by magnetic resonance imaging. Both T1- and T2-weighted contrast-enhanced fast imaging sequences can be used to assess tissue perfusion, vascularity, and microcirculation by applying models developed in nuclear medicine. The diffusion of water molecules and hemodynamic aspects of the macrovasculature can also be monitored. Functional magnetic resonance (MR) imaging enables the visualization of neuronal function and activity, and MR spectroscopy makes possible the metabolic mapping of lesions and surrounding tissue. The advantages of MR techniques includes their low invasiveness, multiplanar imaging ability, and lack of radiation. This contribution discusses the clinical use of functional MR imaging methods and their role in neuroradiological diseases. Measuring perfusion and diffusion allows detailed insight into the pathophysiology of cerebral ischemia and is already being used routinely in acute ischemic stroke. Dynamic MR angiography enables the hemodynamic assessment of vascular malformations. In CNS neoplasms these imaging techniques can improve lesion characterization and the selecting, planning, and monitoring of therapy. Functional MR imaging techniques have also revolutionized the study of psychiatric illness; however, their clinical utility here is still limited. Initial results in patients with dementia and schizophrenia have provided insight into the pathophysiological changes of these diseases. PMID- 11103408 TI - [Nuclear medicine diagnosis in diseases of the central nervous system]. AB - Positron-emission tomography (PET) and single photon emission computed tomography (SPECT) can be used to visualize and quantify cerebral perfusion, glucose consumption, neurotransmission, and amino acid uptake. These techniques are clearly superior to conventional structural imaging techniques for several indications. This contribution describes the clinical role of PET and SPECT in clinical neurology. PMID- 11103409 TI - [Multimodal SPECT and MRT imaging data analysis for an improvement in the diagnosis of idiopathic Parkinson's syndrome]. AB - Parkinson's disease (PD) is characterized by a degeneration of nigrostriatal dopaminergic neurons, which can be imaged with 123I-labeled 2 beta-carbomethoxy-3 beta-(4-iodophenyl) tropane ([123I]beta-CIT) and single-photon emission computed tomography (SPECT). However, the quality of the region of interest (ROI) technique used for quantitative analysis of SPECT data is compromised by limited anatomical information in the images. We investigated whether the diagnosis of PD can be improved by combining the use of SPECT images with morphological image data from magnetic resonance imaging (MRI)/computed tomography (CT). We examined 27 patients (8 men, 19 women; aged 55 +/- 13 years) with PD (Hoehn and Yahr stage 2.1 +/- 0.8) by high-resolution [123I]beta-CIT SPECT (185-200 MBq, Ceraspect camera). SPECT images were analyzed both by a unimodal technique (ROIs defined directly within the SPECT studies) and a multimodal technique (ROIs defined within individual MRI/CT studies and transferred to the corresponding interactively coregistered SPECT studies). [123I]beta-CIT binding ratios (cerebellum as reference), which were obtained for heads of caudate nuclei (CA), putamina (PU), and global striatal structures were compared with clinical parameters. Differences between contra- and ipsilateral (related to symptom dominance) striatal [123I]beta-CIT binding ratios proved to be larger in the multimodal ROI technique than in the unimodal approach (e.g., for PU: 1.2 vs. 0.7). Binding ratios obtained by the unimodal ROI technique were significantly correlated with those of the multimodal technique (e.g., for CA: y = 0.97x + 2.8; r = 0.70; P < 0.001). Concerning the correlations between SPECT data and clinical parameters, the significance levels in the multimodal ROI technique, for example, for the correlation between CA and the UPDRScom subscore (r = -0.49 vs. -0.32). These results show that the impact of [123I]beta-CIT SPECT for diagnosing PD is affected by the method used to analyze the SPECT images. The described multimodal approach, which is based on coregistration of SPECT and morphological imaging data, leads to improved determination of the degree of this dopaminergic disorder. PMID- 11103410 TI - [Radiodiagnosis of the lung]. AB - Radiological cross-sectional imaging modalities, particularly computed tomography (CT) have become the mainstays for diagnosing lung disease in recent years. These enable morphological visualization of pathological processes with the greatest possible spatial resolution. Modern technical developments and complementary strategies have led to new applications and new functional assessments which need to be reviewed together with state-of-the-art techniques in nuclear imaging. The diagnosis of pulmonary embolism using spiral CT angiography and magnetic resonance (MR) angiography certainly belongs in this category. CT has become the an alternative modality of first choice, and it is also challenging pulmonary angiography as the gold standard. Direct visualization of patent pulmonary arteries and thromboembolic material is complemented by that of effects on the pulmonary parenchyma and right heart function; it also provides perfusion studies and MR-based flow measurement to assess hemodynamic compromise. Ventilation studies have long been a domain of nuclear imaging, and new techniques for the direct visualization of ventilation are emerging from recent developments in the field of MR imaging, for example, using hyperpolarized inert gases. New functional parameters of ventilation can be derived from these studies. For the diagnosis of metabolically active disease, such as tumor and pneumonia, CT offers very high sensitivity, for example, in screening for intrapulmonary nodules using low-dose CT and in the early detection of pulmonary infiltrates in high-risk patients. Especially for characterizing pulmonary nodules there is a need to combine nuclear medicine techniques, such as in positron-emission tomography. PMID- 11103411 TI - [Nuclear medicine diagnosis of the lung]. AB - Scintigraphic recording of regional ventilation and perfusion with 99mTc-Aerosol and 99mTc-MAA remain in the foreground of nuclear medicine pulmonary diagnostics. The most important indication for ventilation scintigraphy is the prediction of postoperative pulmonary function, which is still performed in many hospitals with perfusion scintigraphy, and with which, in turn, intrapulmonary right-left shunts can be simply and also semiquantitatively recorded. Combined ventilation/perfusion scintigraphy offers a very high degree of sensitivity in the proof of acute pulmonary embolism, is therefore exceptionally well suited for exclusion diagnostics, while specificity compared to pulmonary angiography and spiral CT still needs some clarification. The self-cleaning mechanism of the lung can be quantitatively examined using mucociliary and resorptive clearance. The clinical areas of application are limited for methodical reasons. Primary diagnostics of bronchial carcinoma and dignity differentiation of solitary pulmonary nodules, preferably with 18F-FDG PET are gaining steadily in importance. PMID- 11103412 TI - [MR tomography of the heart]. AB - The introduction of magnetic resonance (MR) tomography has fundamentally changed radiological diagnosis for many diseases. Invasive digital subtraction angiography has already been widely replaced by noninvasive MR angiography for most of the vascular diseases. The rapid technical development of MR imaging in recent years has opened new functional imaging techniques. MR imaging of the heart allows simultaneous measurement of morphological and functional parameters in a single noninvasive examination without any radiation exposure. Because of the high spatial resolution and the reproducibility cine MR imaging is now the gold standard for functional analysis. With the improvement of myocardial perfusion and viability studies many diseases of the heart can be diagnosed in a single examination. MR spectroscopy is the only method which allows a view of the metabolism of the heart. New examinations for vascular imaging and flow quantification complete the goal of "one-stop-shop" imaging of the heart. MR imaging is the only diagnostic modality which allows a complete evaluation of many diseases of the heart with one technique, basic examination as well as follow-up studies. The very rapid improvement in MRI will overcome most of the limitations in the near future, especially concerning MR coronary angiography. PMID- 11103413 TI - [Nuclear medicine studies of the heart]. AB - Nuclear imaging methods provide noninvasive indexes of myocardial function, perfusion, and metabolism and are well accepted in clinical cardiology. Advances in prevention and treatment of cardiac disease have resulted in decreasing cardiovascular mortality in industrialized nations. The improvement in therapeutic options has increased the demand for diagnostic tests that might guide clinical decision making. Information beyond the pure anatomic characterization of coronary stenoses is required. Nuclear imaging can be used for early detection and monitoring of the severity and extent of disease. The prognostic potential of such functional testing is being increasingly appreciated and used to guide therapy, thereby resulting in improvement of the quality and cost-effectiveness of the workup of patients with cardiovascular disease. Extensive clinical validation has resulted in growing acceptance of these techniques. Furthermore, ongoing improvement of imaging techniques and development of new radiopharmaceuticals will pave the way for disease-specific, molecular-targeted cardiac imaging in the future. PMID- 11103414 TI - [Radiological diagnosis of the liver]. AB - Highly specific methods are required for the diagnostic workup of focal hepatic lesions, since benign circumscribed liver changes are very common. Although cross sectional imaging techniques have a high diagnostic accuracy, radionuclide imaging techniques such as colloid, red blood cell, or hepatobiliary scan are commonly performed when a benign lesion is assumed since these permit a definite diagnosis with high specificity. The diagnosis of a primary or secondary malignant liver tumor, however, usually relies on radiological imaging techniques alone, supported by needle biopsy. Whether positron emission tomography as a primary or supplementary diagnostic tool will have a role in the routine staging of malignant tumors remains to be determined. PMID- 11103415 TI - [Nuclear medicine diagnosis of the liver]. AB - Four types of radionuclide investigations are described here: 99mTc-labeled red blood cell scintigraphy, colloid liver scintigraphy, hepatobiliary scintigraphy, and positron emission tomography with [18F]fluorodeoxyglucose. The role of nuclear imaging techniques in the diagnosis of liver diseases has changed in recent years and now compliments morphological imaging modalities by offering the unique ability to visualize function and metabolism. The studies described here are therefore rarely performed now by themselves for the delineation of secondary liver tumors. These radionuclide investigations are used principally to narrow the differential diagnosis of focal liver disease. PMID- 11103416 TI - [Quantitative recording of renal function with magnetic resonance tomography]. AB - AIM: To show the potential of various methods in magnetic resonance imaging for the evaluation of renal function. MATERIAL AND METHODS: A combined assessment of renal morphology, renal hemodynamics and function is proposed. Various techniques are explained, including multiphasic 3D gadolinium MR angiography, MR phase contrast flow measurements, quantitative perfusion measurements with intravascular contrast agents, and MR renography and MR urography. The use of these techniques is demonstrated for renovascular diseases. RESULTS: The combined use of these techniques allows renal artery stenosis to be accurately detected and evaluation of renal blood flow, perfusion, glomerular filtration rate, and renal excretion. Based on true quantitative parameters, the hemodynamic and functional significance of the stenosis can be assessed. Renovascular diseases can be differentiated from renoparenchymal disease. CONCLUSION: For the assessment of renal function, functional magnetic resonance imaging techniques are an important alternative to nuclear medicine. The predictive value regarding the effect of revascularization is currently under investigation. PMID- 11103417 TI - [Nuclear medicine diagnosis of the kidneys]. AB - BACKGROUND: Renal studies have a long tradition in nuclear medicine and are available for routine use for more than 30 years. Their high clinical acceptance is mainly based upon the fact that they allow for quantitative evaluation of different functional parameters such as glomerular filtration, effective renal plasma flow, or postrenal transport. The used methods are validated by experimental as well as numerous clinical studies and are performed throughout the world in a highly standardized way. INDICATIONS: Indications are verification/exclusion of a disturbed renal function, detection or evaluation of renal artery stenosis, and differential diagnosis of urinary tract obstruction. Further diagnostic improvement might be achieved by use of positron emission tomography which has the potential for absolute quantification of physiological parameters such as renal blood flow in ml.min-1.100 g-1 tissue. PMID- 11103418 TI - [Radiological diagnosis of the skeletal system]. AB - Diagnostic strategies, radiological signs on conventional techniques and MRI as well as problems in skeletal radiology are discussed. Emphasis is given on inflammatory conditions, bone metastasis and traumatology. Particularly common aspects of diagnostic radiology and nuclear medicine are considered. PMID- 11103419 TI - [Nuclear medicine study potentials in diseases of the skeletal system]. AB - Even though new diagnostic methods, as CT/MRI, are widely available, the bone scintigraphy still does remain an important tool for imaging bone pathology. An increase in diagnostic accuracy is possible by using different imaging modalities: wholebody scan, three phase bone scintigraphy and SPECT. For this reason, the bone scintigraphy can be used for diagnosis of tumors/infections and therapy monitoring. Applying these methods, important informations can be gained for differential diagnosis. The bone scintigraphy is easy to perform, allows good whole-body overview by low radiation burden to the patient. PMID- 11103420 TI - [Computed tomography and magnetic resonance tomography in tuberculous spondylitis. A review of the literature and the authors' own observations]. AB - PURPOSE: The findings of tuberculous spondylitis in MRT have been described extensively. Nevertheless the diagnostic value of both methods in the diagnosis of this severe manifestation of the tuberculous disease was not yet defined definitely. MATERIALS: We performed a review of the recent literature and a retrospective analysis of the findings in ten patients with proven tuberculous spondylitis. Here we evaluated 10 CT and 6 MRT. RESULTS: Major findings in computed tomography (n = 10) were osseous sequestration (8/10), subperiosteal bone apposition (6/10), abscess of the surrounding tissue. (8/10) and calcification of the masses (3/10). In all cases which were examined by MRT (n = 6) marrow edema was seen. Affection of the soft tissue was described by means of MRT in 5/6 patients. All patients showed rim enhancement. CONCLUSIONS: MRT shows signs of infection (bone marrow edema) which is an early but rather unspecific finding. The commonness of osseous lesions in advanced tuberculous spondylitis suggests a benefit of computed tomography in the later stages. Both methods are complementary in the differential diagnosis of tuberculous and non-specific spondylitis. PMID- 11103421 TI - [Vomiting and watery diarrhea after a trip to Egypt]. PMID- 11103422 TI - [The x-ray findings in lung diseases]. PMID- 11103423 TI - [IgE allergy due to formaldehyde paste during endodontic treatment. Apropos of 4 cases: 2 with anaphylactic shock and 2 with generalized urticaria]. AB - We report 4 cases of allergic reaction to formaldehyde-containing root canal sealant after endodontic care: 2 anaphylactic shocks and 2 local reactions with generalized urticaria. Allergic IgE mediated mechanisms were suggested by the clinical presentation, skin tests and high levels of anti-formaldehyde IgE. These infrequent but potentially severe reactions after canal treatment led us to examine the involved mechanisms, the diagnostic procedure and the possibility of prevention in odontostomatology. PMID- 11103424 TI - [Allergy to local anesthetics]. AB - Suspected allergic reaction to local anesthetics is a frequently encountered problem. Although some reactions are proven to result from an allergic mechanism, many remain unexplained, raising various levels of risk depending on the type of hypersensitivity involved. Good clinical management of allergic reactions is now well standardized, allowing for proper identification of the causal agent in proven cases. PMID- 11103425 TI - [Treatment of low-pressure vascular malformations by injection of Ethibloc. Study of 19 cases and analysis of complications]. AB - We retrospectively studied the cases of 19 patients suffering from low flow vascular malformations who were treated with Ethibloc sclerotherapy. Out of 10 venous angiomas, 5 entirely disappeared, 3 considerably decreased and 2 remained unaltered. Out of 7 cystic lymphangiomas, 7 were completely cured and in the 2 other cases, outcome was quite good. Post-treatment complications consisted in a local inflammatory reaction. This reaction occurred immediately and disappeared within a few days without after-effects. It materialized as an Ethibloc exteriorization among 5 patients suffering from venous angiomas and 3 patients suffering from lymphangiomas and as an inflammatory lump for 2 venous angiomas carriers and for 4 lymphangiomas carriers. This lump was located at the puncture point of the product and it was due to some Ethibloc residue. Fourteen of these local reactions spontaneously decreased; 5 of them required a surgical operation but in all cases, after-effects were minor. These satisfactory results (volume decreasing in 90% of the malformations) as well as the mildness of the side effects encourage to use Ethibloc in the treatment of low flow vascular malformations. In our experience, Ethibloc is particularly appropriate for the treatment of large lymphangiomas. PMID- 11103426 TI - [Treatment of recurrent luxation of the temporomandibular joint with botulinum toxin]. AB - We report the case of a 70-year-old man who suffered recurrent dislocations of the temporomandibular joint secondary to severe Parkinson syndrome. The patient was given repeated injections of botulinum toxin. After 3 injections over a 9 month period, no further dislocation occurred. Botulinum toxin may be an alternative to surgery. PMID- 11103427 TI - [Case report: radiographic anomaly of the parotid space]. PMID- 11103428 TI - [Gorlin-Goltz syndrome. Apropos of a maxillary cyst]. AB - We report a case of Gorlin-Goltz syndrome and recall the principle features of this uncommon disease. The syndrome associates multiple skeletal malformations, skin tumors and epidermoid maxillary cyst. We discuss the differential diagnosis of maxillary cysts. PMID- 11103429 TI - [Writing a scientific medical article: the original article]. PMID- 11103431 TI - [Use of variously shaped miniplates in osteosynthesis of sagittal osteotomy of the mandible]. PMID- 11103430 TI - [Functional management of fractures of the mandibular condyle]. PMID- 11103432 TI - [Recognizing the situation (1): a plea for an open debate about rationing]. AB - The rise in health costs is due to the tremendous advances in medical science and other causes, such as a system offering a unique financial stimulus for health professionals to provide more services, non-functioning competition, patients' demand for every possible form of treatment as rapidly as possible, a compliant political community and also new players such as consultants, the ethics industry, lawyers and nursing academics. On the other hand, the public is not willing to pay more in taxes and health insurance premiums. The resultant dilemma sooner or later forces restrictions on health care which can be called either optimisation, rationalisation or rationing. PMID- 11103433 TI - [New physiopathological findings in paraproteinemias]. AB - Apart from their antigen-recognising function, immunoglobulins have other physicochemical properties. The best described is cryoprecipitation of monoclonal and/or polyclonal immunoglobulins. The cryoprecipitation phenomenon is unique for each cryoglobulin, and depends on the intrinsic properties of the monoclonal immunoglobulin, the protein environment, temperature and the surrounding ionic concentrations. The complement activating properties of monoclonal immunoglobulins are also unique. We have described a series of monoclonal immunoglobulins which are able to activate the classical pathway of complement without the formation of antigen/antibody complexes. Analysis of the properties of these monoclonal immunoglobulins suggests new therapeutic strategies in oncology. Monoclonal chimeric antibodies, combining antibody specific against tumour antigens and strong complement activation, may be more damaging to tumor cells than currently used antibodies. PMID- 11103434 TI - Paraproteinaemias: pathophysiology. AB - Paraproteinaemias may be associated with benign or malignant proliferations of lymphocytes or plasma cells, including multiple myeloma, monoclonal gammopathy of undetermined significance (MGUS) and Waldenstrom's macroglobulinaemia. Primary amyloidosis may be associated with multiple myeloma and rarely with lymphoid malignancies, but most cases can be considered as a particular form of monoclonal gammopathy of undetermined significance, where the paraprotein causes damage by virtue of its amyloidogenic properties. This article discusses recent advances in understanding of the biology of multiple myeloma. Multiple myeloma is now known to arise from a post-germinal centre B cell in the lymph node which homes to the bone marrow. Interactions with stromal cells in the marrow facilitate homing and growth of the myeloma cells. The stromal cells produce IL-6, which is an important growth factor for myeloma cells, while the myeloma cells produce factors such as TNF-alpha and IL-1 beta that activate osteoclasts, resulting in myeloma bone disease. Myeloma cells also produce vascular endothelial growth factor which results in increased microvessel formation in the marrow, promoting tumour growth. There has been interest in the possible role of the Kaposi's sarcoma associated herpes virus (HHV8) in multiple myeloma, following the demonstration of viral gene sequences in multiple myeloma marrow. However, results of further studies have been conflicting and at present there is no clear evidence for an aetiological role of HHV8 in multiple myeloma. Cytogenetic studies using modern techniques have demonstrated that almost all multiple myeloma cases are cytogenetically abnormal, the predominant abnormalities being various translocations involving chromosome 14q and deletions of chromosome 13. 14q translocations are equally common in monoclonal gammopathy of undetermined significance, but deletions of chromosome 13 seem to be associated with progression to multiple myeloma, and also have powerful prognostic significance for survival in multiple myeloma patients. PMID- 11103435 TI - [Treatment of multiple myeloma]. AB - The chief advances observed over the last 15 years in the treatment of multiple myeloma arise from intensive procedures and in particular autologous bone marrow transplantation. However, even if autologous bone marrow transplantation increases the rate of total remissions, no plateau is yet observable in the survival curve and transplantation is probably not a curative therapy. Hopes for the future seem to rest on intensive chemotherapy combined with innovative therapeutic approaches such as diphosphonates, thalidomide or immunotherapy. PMID- 11103436 TI - Fas-mediated cell death in toxic epidermal necrolysis and graft-versus-host disease: potential for therapeutic inhibition. AB - Death receptors are a growing family of transmembrane proteins which can detect the presence of specific extracellular death signals and rapidly trigger cellular destruction by apoptosis. The best studied to date is Fas (CD95). Expression and signalling by Fas and its ligand (FasL, CD95L) is a tightly regulated process essential for key physiological functions in a variety of organs, including the maintenance of immune homoeostasis. Recently, strong evidence has shown that dysregulation of Fas expression and/or signalling contributes to the pathogenesis of toxic epidermal necrolysis and acute graft-versus-host disease. With these new developments, strategies for modulating the function of Fas signalling have emerged and opened up novel therapeutic possibilities. Specific blockade of Fas, for example with intravenous immunoglobulin preparations containing specific anti Fas antibodies, has shown great promise in the treatment of toxic epidermal necrolysis and may also be useful in the treatment of acute graft-versus-host disease. Further developments in this field may have important clinical implications for the treatment of such diseases. PMID- 11103437 TI - [Ethics and genetic diagnosis]. AB - The implications of current ethical concerns in the field of genetic research and applications are discussed. Genetic diagnosis is perceived as ethically ambivalent both in general and in its individual applications. Genetic testing, by inviting participation, reflects this ambivalence and can be taken as the basis for both a "right to know" and a "right not to know". The connection between prenatal diagnosis and abortion is also complex and cannot be regarded as ethically clearcut. In these circumstances the legislator must exercise caution in regulating genetic diagnosis in its present state of development. PMID- 11103438 TI - [Violence against the elderly--by the elderly: does it exist?--Medical aspects] . AB - Thus far there has been very little research on violence in relation to age. This is due to various problems: the definition and delimitation of the term "violence" is unclear, and violence is considered an interactive event in which everyone involved is both victim and perpetrator. Violence is a taboo topic which people prefer to conceal, and those involved do not talk about the violence they experience. The different forms of violence are physical violence, psychic violence, neglect, restraint, financial exploitation and structural violence. Views on the incidence of violence vary: one certainty is that it occurs in care relationships and that psychic violence and neglect are commoner than physical violence. There is no doubt about the correlatives of violence: limited cognitive abilities, disruptive behaviour, troubled relationships, overtaxing, exhaustion and inappropriate structures. Medical care requires awareness of the problem, facing up to one's own violent behaviour, and knowledge of the risks, causes and consequences of violence. Only in this way is effective prevention and intervention possible. Empirical research offers an approach to future prevention and rehabilitation. People required to deal with violence are under a heavy burden, and decisions must often be made in conflict situations. The law provides a framework, which nevertheless allows scope for action. Ethical guidelines may be of great assistance in this context. PMID- 11103439 TI - [Care aspects on the theme "Violence against the elderly--by the elderly: does it exist?"]. AB - Violence against and by the elderly is a day-to-day concern when working with old and very old people. However, the symptoms are undifferentiated and only discernible if carers come to terms with the (violent) history of those involved. The author's aim is not to provide answers to a complex problem but to contribute to the greater mutual understanding which would prevent the emergence of new traumas. PMID- 11103440 TI - [Recurrent Clostridium difficile enterocolitis]. AB - Pseudomembranous enterocolitis generally occurs after antibiotic treatment. The standard treatment is oral metronidazol or vancomycin. Nevertheless, relapses of Clostridium difficile enterocolitis are observed in 10-25% of cases. Factors associated with recurrences include endogenous reinfection by spore formation, selective IgG1 or IgA deficiency or infection with mutated strains of Clostridium difficile. Recurrent Clostridium difficile enterocolitis may be treated with repeat oral vancomycin combined with Sacchoromyces boulardii, with intravenous immunoglobulin for severe colitis. PMID- 11103441 TI - [Reactivation of herpes virus infections by vaccination: evidence or coincidence?]. AB - Varicella zoster and herpes simplex viruses cause latent infections by persisting in human cells. Reactivation has been associated with increasing age, immunosuppression, cancer, stress, fever, exposure to ultraviolet light, and tissue damage. Based on three cases reported to the Swiss Drug Monitoring Centre SANZ, we postulated previously that vaccinations may trigger reactivation of herpes virus infections due to vaccine-induced immunomodulation. In the meantime, 10 new cases of reactivated herpes virus infections soon after vaccinations have been reported. They involved 5 women and 5 men with an age range between 16 and 60. In only one case had a trauma preceded, otherwise healthy subjects with no known relevant comorbidity were vaccinated. The clustering of reports after publication points to a previous underreporting of similar cases. This may be explained by the fact that both vaccinations and reactivations of herpes virus infections are frequent, and a causal link is not suspected. However, these new cases do not prove causality, and extensive epidemiological or experimental studies are needed to elucidate the possible link between vaccination and reactivation of herpes virus infections. PMID- 11103442 TI - Hypokalaemic periodic paralysis associated with controlled thyrotoxicosis. AB - Familial hypokalaemic periodic paralysis is an autosomal dominant muscle disease which has been linked to point mutations in the skeletal muscle L-type calcium channel alpha 1 subunit (alpha 1 s). It consists of muscular weakness episodes due to hypokalaemia caused by intracellular shifting of potassium. We describe the case of a young man of Kurdish origin, with a history of Graves' disease, who was admitted to the emergency room with hypotonic tetraplegia associated with severe hypokalaemia. PMID- 11103443 TI - [A case of pseudophlebitis of the great saphenous vein: focal nodular myositis of the gracilis muscle]. AB - A case of focal myositis in a healthy 68-year-old woman is described. The patient was admitted for evaluation of a painful soft-tissue mass localised on the medial side of the left thigh, initially misdiagnosed as thrombophlebitis of the v. saphena magna. Laboratory data were normal, in particular sedimentation rate and muscle enzyme levels. After exclusion of venous thrombosis, the mass localised in the left m. gracilis was surgically removed. Histologic examination of the biopsy specimen showed muscle cell necrosis and severe inflammation, with lymphocytic infiltration leading to the diagnosis of focal myositis. This is a rare benign inflammatory pseudotumour of skeletal muscle. The aetiology and pathogenesis of the disease remain unclear. It is most commonly seen in the lower extremities and may mimic thrombophlebitis or soft-tissue neoplasm. Ultrasound and magnetic resonance scans are helpful, but definitive diagnosis is obtained only by histology. Because recurrent lesions in other skeletal muscles are possible, and a third of patients develop polymyositis, a follow-up of several years is recommended. PMID- 11103444 TI - [Conservative management of spontaneous splenic rupture as a complication of infectious mononucleosis: two case reports and literature review]. AB - We report on conservative management of 2 patients with spontaneous splenic rupture associated with infectious mononucleosis. Both patients had an unremarkable hospital course and were discharged within 7 days of admission. Resolution of the haematoma was followed by ultrasound monitoring during the hospital stay. A literature review to 1999 shows that approximately 45 patients with serologically proven infectious mononucleosis have suffered spontaneous rupture of the spleen. Spontaneous splenic rupture is a rare but potentially fatal complication of infectious mononucleosis. Although splenectomy has been advocated in the past as the definitive therapy, we recommend that non-surgical management be considered in haemodynamically stable patients, to avoid the complications of splenectomy (e.g. post-splenectomy sepsis). PMID- 11103445 TI - [Percutaneous endoscopic gastrostomy (PEG) for palliative decompression drainage in inoperable ileus]. AB - Bowel obstruction, causing repetitive vomiting and reduced quality of life, is a common complication in patients with intraabdominal malignancies. Conservative treatment with nasogastric tubes is limited by patient discomfort. Antisecretory drug treatment with octreotide may be insufficient. We describe the application of percutaneous endoscopic gastrostomy (PEG) in 3 terminally ill cancer patients as simple and effective method for decompression in the upper gastrointestinal tract. PMID- 11103447 TI - [Hepatic hydrothorax without ascites]. PMID- 11103446 TI - [Rational diagnostic strategy for tuberculous lymphadenitis] . AB - OBJECTIVES: To examine the clinical, radiographic and laboratory findings in patients with tuberculous lymphadenitis and to analyse the investigational strategies which lead to the diagnosis of tuberculous lymphadenitis. METHODS: Retrospective study including 16 HIV-negative patients at the Cantonal Hospital, Winterthur with tuberculous lymphadenitis diagnosed between 1994 and 1999. RESULTS: The majority of patients presented with local symptoms and without signs of severe systemic disease. All the PPD skin tests performed were positive. Cultures for M. tuberculosis were more often positive using fine-needle aspiration than surgical biopsy. We found a lack of systematic diagnostic strategy. CONCLUSIONS: We suggest a standardised investigation procedure. When tuberculous lymphadenitis is suspected, the first diagnostic step consists of a PPD skin test and fine-needle aspiration for acid fast smear, mycobacterial culture and cytology. Surgical biopsy should be done if the cytological and mycobacteriological results of fine-needle aspiration are not diagnostic. PMID- 11103448 TI - Pennywise bioplastics? PMID- 11103450 TI - Gravity, revised. PMID- 11103449 TI - From ague to West Nile. PMID- 11103451 TI - The second abortion pill. PMID- 11103452 TI - Paleolithic pit stop. PMID- 11103453 TI - Red team versus the agents PMID- 11103454 TI - The amazing acenes PMID- 11103455 TI - A trace of the corona PMID- 11103456 TI - Completing the circuit. PMID- 11103457 TI - In the waiting room. PMID- 11103458 TI - Bits of radio. PMID- 11103459 TI - Rulers of the Jurassic seas. PMID- 11103460 TI - Nanotubes for electronics. PMID- 11103461 TI - The secrets of stardust. PMID- 11103462 TI - Piecing together Alzheimer's. PMID- 11103464 TI - A new theory of urbanism. PMID- 11103463 TI - The science of smart growth. PMID- 11103465 TI - The coolest gas in the universe PMID- 11103466 TI - Diapers--disposable. Superabsorbers. PMID- 11103468 TI - Jumping champions PMID- 11103467 TI - Calibrating with cold. PMID- 11103470 TI - Not Nelson's Obelisk PMID- 11103469 TI - Gleaning nuggets. PMID- 11103471 TI - [Where can pre-implantation diagnosis lead?]. PMID- 11103472 TI - [Contingent negative variation (CNV) in children with hyperkinetic syndrome--an experimental study using the Continuous Performance Test (CPT)]. AB - OBJECTIVES: Evaluation of event-related potentials for selective attention in attention deficit/hyperactivity disordered children. METHODS: Selective attention processes were examined in a group of 18 boys aged 6 to 12 years with attention deficit/hyperactivity disorder and compared with the data of a control group of 21 age and sex matched boys. Parallel thereto the event-related potentials (ERP) were derived during the test at the electrode positions Fz, Cz, Pz, and Oz with reference to the linked ears. RESULTS: Two components of the contingent negative variation (CNV) with different topography were identified in the ERP following a preparatory stimulus (CNV-1: 600 to 1100 ms and CNV-2: 1000 to 1500 ms after the stimulus). There were no group differences at the behavioural level (number of correct detected targets, number of errors). Significant group differences resulted with regard to the topography of the two CNV components. Children with ADD showed an attenuated frontal CNV-1 amplitude and a trend towards increased CNV-1 and CNV-2 occipital amplitudes. CONCLUSIONS: The results support the hypothesis of impaired frontal inhibitory processes in children with ADD. PMID- 11103473 TI - [Promoting reading and spelling in children with computer programs]. AB - OBJECTIVES AND METHODS: Sixteen students trained their reading and spelling skills with the computer programmes Budenberg 1 and 2 and the Comles Package for 1000 minutes over a one-month period. RESULTS: Following one month of computer training, reading test scores had improved for seven of the 16, and spelling test scores for three children whose basic performance had been poor. Three and a half months of school instruction later, the reading test scores had improved for nine children, while there was no effect upon the spelling scores for most of the students. PMID- 11103474 TI - [Therapy motivation of young prisoners]. AB - OBJECTIVES: Therapy adherence was examined within a sample of 145 imprisoned male juveniles with regard to factors of influence, structure and predictability. METHODS: Predictors included biographic data, expectations with regard to therapy, personality traits (measured with the FPI-R) and psychological impairment (investigated by means of the Symptom Checklist). Therapy adherence as a criterion can be divided into the following dimensions: affliction, dissatisfaction, requests for change and help, and expectations of success. RESULTS: Within the sample a clear impairment of biographical and psychological data, as well as of personality traits was found. Values were average for expectations regarding therapy and therapy adherence, while two thirds of the sample were willing to undergo treatment during imprisonment. CONCLUSIONS: Therapy adherence proved to be a one-dimensional construct, best predicted by emotional instability, followed by the symptom score, expectations regarding therapy and inhibition. Dividing the entire sample into subgroups, significant differences were found mainly for psychological test data and less so for the construct of therapy adherence. PMID- 11103475 TI - [Anxiety and depression in students. A study in Dresden middle schools and high schools]. AB - OBJECTIVES: Recent investigations by the WHO have shown once again the high prevalence of mental disorders in European countries. Mainly in the field of mental disorders considerable deficits exist particularly with regard to primary prevention. METHODS: Although there is no doubt as to the importance of primary prevention within the scope of Public Health, there are deficits in this branch in Germany. This is unfortunate, considering that the proof of successful interventions is evidence for causal risk models. The current investigation is part of a study focusing upon the development, implementation and evaluation of a prevention program for anxiety and depression in adolescents and young adults. RESULTS: Presented here are the prevalence rates of anxiety disorders and depression ascertained in the first cross-sectional study carried out in secondary schools and high schools. A total of 627 high school students and 485 secondary school students in grades nine and ten in Dresden were examined. PMID- 11103476 TI - [Legal binding character of guidelines]. AB - Guidelines as recommendations by the medical associations concerning the diagnosis and treatment of certain diseases do not have the character of judicial norms. However, they may have an impact on decision-making in court. Within social security insurance law, and especially in medical malpractice litigation, the medical standard has become a legal term. As far as statements in guidelines render the actual standard as affirmed, the more binding is their character with respect to judicial questions. This article deals with this interplay between medical quality assurance and the legal demands on medical action. As a basis for discussion of this issue the relevant jurisdiction will be represented. Its impact on the guidelines for child and adolescent psychiatry will be pointed out as a concrete example. PMID- 11103477 TI - [Feeding disorders and failure to thrive in small and/or handicapped children]. AB - Feeding disorders and failure-to-thrive (prevalence 2% to 4%) rarely have an isolated cause, but most often a number of inappropriate conditions are leading up to the development and, especially, the maintenance of the disorder. These can include organic causes like chronic diseases, peculiarities of the person, strange behavior of the child or the care person or of the interaction-problems. An obligatory classification of feeding disorders does not exist. Feeding disorders and failure-to-thrive can ask for a long-term additional or full tube feeding or the child rejects age-appropriate food texture, has a very selective eating behavior or there are massive interaction problems during feeding. Feeding disorders and failure-to-thrive can not only have direct physical effects but also long-term unfavourable influences on behavioral aspects as well as on mental abilities. The diagnosis of feeding disorders and failure-to-thrive comprises next to the clarification of a basic organic disease, the clarification of swallowing and oral-motor capabilities as well as the exclusion of a gastroesophageal reflux. A differentiated feeding protocol must include the oral feeding as well as the tube feeding. A behavior observation comprises the feeding situation and, if necessary, further situations of interaction. Besides the treatment of the basic disease, a direct guidance in the feeding situation for the care person is necessary. Furthermore, a therapy of the oral motorics as well as one of the care person and guidelines for interaction during different situations can be important. PMID- 11103478 TI - Report of the BSE inquiry published. PMID- 11103479 TI - Major milestone reached in global polio eradication. PMID- 11103480 TI - AIDS and HIV infection in the United Kingdom: monthly report. PMID- 11103481 TI - [Pelvic osteotomies in children and adolescents]. AB - The main types of pelvic osteotomies in children and adolescents are reviewed. Osteotomies in the first group aim at reorienting the acetabulum: Salter's innominate osteotomy is widely used; its technique, possible drawbacks and indications are analyzed; double and triple osteotomies are then reviewed (Sutherland, Le Coeur, Steel, Tonnis and Trousseau) with their prerequisites, drawbacks and specific indications. A second group of osteotomies do not involve complete transsection of the hemipelvis; they are acetabuloplasties following the techniques described by Dega and by Pemberton, the indications of which are also presented together with their prerequisites. Last comes Chiari's osteotomy: it appears as a palliative operation, with limited indications in children and adolescents. Finally, the indications for pelvic osteotomies are reviewed, according to patient's age, anatomical status of the hip and underlying pathology. Unstable and dysplastic DDH hips may be treated by Salter's osteotomy, Pemberton's acetabuloplasty of triple pelvic osteotomy if the hip is mobile, well centered and congruous. The more simple Salter and Pemberton operations are to be preferred to triple osteotomy as long as they are indicated, i.e. until the age of 5 to 8 years. Established congenital dislocations may be treated using Chiari's osteotomy in cases where a reorientation osteotomy or acetabuloplasty is no longer indicated, provided the hip remains mobile. The indications for pelvic osteotomy in Perthes disease are analyzed, and the arguments for a pelvic rather than femoral osteotomy in some cases are presented. Pelvic osteotomies with the numerous techniques developed over the years, have been a major advance in the treatment of hip anomalies in children. In older adolescents, their indication must be balanced against those of hip reconstruction; they must anyway never make subsequent arthroplasty in adult age difficult or impossible. PMID- 11103482 TI - [Rheumatoid arthritis of the wrist with adult onset]. AB - The author reviews the consequences of rheumatoid synovitis of tendons and joints at the wrist, consequences which are different on the volar and dorsal aspects of the wrist. He refers to a modified Larsen classification to describe the consequences of instability in the radiocarpal (RC), midcarpal (MC) and radioulnar (RU) joints, both in the coronal and sagittal planes. A. On the volar aspect, tenosynovitis of the flexor tendons is frequent but may be difficult to diagnose. Synovitis in the carpal tunnel, although frequent, rarely results in compression of the median nerve; persistence of synovitis despite medical treatment is an indication to synovectomy. The latter may have to be extended into the palm and over the proximal phalanges, using the appropriate approach in the individual cases. Flexor tendon ruptures may occur, mostly of the flexor pollicis longus (FPL) and the flexor tendons to the index finger. Rupture of the FPL may be treated by a tendon graft or by arthrodesis of the i.p. joint. Rupture of the deep flexor tendon to the index may be treated by anastomosis to that of the medius; rupture of the superficial flexor tendon to the index may be treated similarly; rupture of both flexor tendons requires a tendon graft. B. On the dorsal aspect, the indications vary according to the stage of the disease. In Larsen's stage IV or V (destruction of one or more of the radiocarpal and intracarpal joints with navicular dislocation), arthrodesis or arthroplasty is indicated; the latter is ruled out however if extensor tendons are ruptured or the bone stock is insufficient. An original or modified Mannerfelt technique is used for arthrodesis, with the wrist in neutral or slightly extended position. Several wrist prostheses are available. Swanson's silastic implant has been discontinued; the Meuli, CFV, Biax, Trispherical, ATW, and GUEPAR prostheses have all been used with varying degrees of success. The choice between arthrodesis and arthroplasty is based on the severity of articular and tendon pathology, on uni- or bilateral involvement and on the condition of other joints, particularly in the upper limb. In less advanced stages, the author advocates using a combined operation with synovectomy of the extensor tendons and of the RC, MC and RU joints, relaxation by tendon transfers and Sauve-Kapandji's technique; he stresses important technical points. The specific indications for radiolunate arthrodesis are discussed. PMID- 11103483 TI - Destination of debris during intramedullary reaming. An experimental study on sheep femurs. AB - This study was designed to further explain the better fracture healing in fractures treated with a reamed nail. It investigates the location and quantity of the reaming debris in an ex vivo animal model to test the autograft theory. In 10 cadaveric sheep femurs, a 5-mm semicircular gap was created at the midshaft. The medullary cavity was opened and the reaming debris that dropped from the gap during reaming and the debris from the proximal opening were collected and weighed separately. The mean harvest of reaming debris at the gap was 0.99 g +/- 0.12 g (24%) and from the proximal opening at the medullary cavity 3.08 g +/- 0.31 g (76%) (total 4.07 +/- 0.34 g). This study proves that a significant amount of reaming debris collects at an artificial fracture gap during reaming of the medullary cavity. This finding supports the theory of bone autografting. PMID- 11103484 TI - Short- and long-term effects of regional application of morphine and bupivacaine on the iliac crest donor site. AB - We investigated the analgesic effect of regional application of bupivacaine and a morphine-bupivacaine combination on iliac crest donor-site pain in a randomized, double-blind controlled study of 45 patients. Patients were divided into three groups: group I (control group), group II (bupivacaine) and group III (morphine bupivacaine combination). Pain in the acute stage was evaluated by visual analogue scale scoring and analgesic consumption. Chronic pain and dysesthesia were evaluated at 12 weeks after operation at a follow-up visit. It was found that local bupivacaine administration with or without morphine provided satisfactory analgesia in the acute stage following iliac crest bone harvesting. The amount of analgesic consumption was found to be significantly less with the addition of morphine to bupivacaine, when compared to bupivacaine alone. Effective pain control in the acute stage had a favorable effect on long-term pain and dysesthesia, which are the main complaints after iliac crest bone harvesting. This effect was augmented significantly by addition of morphine to the local anesthetic solution. PMID- 11103485 TI - [Surgical treatment of metastases from thyroid cancer in the axial skeleton. A retrospective study of 18 cases]. AB - Between 1990 and 1997, 18 patients with a mean age of 55.5 years (11 females, 7 males) underwent surgical treatment for a metastasis from thyroid cancer involving the axial skeleton. At the time of surgery all patients had a poor prognosis: 7 metastases revealed the thyroid cancer, all 18 patients had a neurological or mechanical complication, 9 had multiple metastases, all were over 40 years of age. After arteriography with embolization, the surgical procedure consisted of curettage of the tumor and reconstruction, followed by treatment with iodine 131. The survival rate 3 years after surgery was 50%. At the last review, the functional outcome was good and 17 patients had total neurological recovery. Four complications occurred: 1 operative hemorrhage, 3 postoperative infections. Four patients had local recurrence of the metastasis with a one-year survival rate of 20%. When the thyroid cancer was revealed by the axial metastasis, the 3-year-survival rate was 42%. In cases with huge metastases, the 3-year-survival rate was 71%. It appears from these data that surgical treatment of metastases from thyroid cancer in the axial skeleton still achieves a good functional outcome even in cases where neurological or mechanical complications had occurred before surgery. PMID- 11103486 TI - The floating radial head prosthesis for comminuted radial head fractures: a multicentric study. AB - We report our experience with the floating radial head prosthesis of Judet for comminuted fractures of the radial head. We present the results in 15 patients with a mean follow-up of 25.2 months. Thirteen prostheses were inserted for acute Mason III fractures of the radial head, and 2 were inserted for chronic problems after radial head fracture. According to the Mayo Elbow Performance Index there were 7 excellent, 3 good, 1 fair and 2 poor results in the group with acute injuries. In this group, one prosthesis was removed after 8 months for severely decreased elbow function. In the group with chronic problems, there were 2 fair results. There were no dislocations or prosthesis fractures. None of the prostheses showed signs of loosening. Three patients in the acute group developed wrist pain, and in one patient in the chronic group, preexisting wrist pain disappeared after insertion of the radial head prosthesis. Our short-term results suggest that the floating radial head prosthesis is a suitable solution for early or delayed treatment of Mason type III fractures, either isolated or associated with more complex injuries. PMID- 11103487 TI - Radial head dislocation with plastic deformation of the ulna in children. A rare and frequently missed condition. AB - Although often reported in the literature, 'isolated' traumatic radial head dislocation in children is a rare condition which has not been studied extensively. There is very often a delay in diagnosis and treatment. Lincoln and Mubarak described the 'ulnar bow sign' in 1994. They accurately described the plastic deformity of the ulna and explained how to make a correct diagnosis from the radiographs. Probably most 'isolated' radial head dislocations in children are associated with plastic deformation of the ulna as stated in 1984 by Dubuc et al. (2). The recognition of these "plastic" Monteggia fractures is mandatory, as action on the ulna plays a central role in the treatment. On the other hand, the 'ulnar bow sign' may be used to detect subtle changes in the position of the radial head. We present 4 cases of chronic radial head dislocation associated with plastic deformation of the ulna. Open reduction of the radial head and reconstruction of the annular ligament was performed. An osteotomy of the ulna was deemed necessary in 3 cases. PMID- 11103488 TI - Prognosis of wrist ganglion operations. AB - A retrospective study was conducted to evaluate the results of treatment of 40 wrist ganglia operated under local anesthesia over four years. The mean follow-up period was 27 months (range 6-48 months). There were 24 dorsal and 16 volar ganglia. The mean complication rate was 56% for volar ganglia, 12.5% for dorsal ganglia, and the difference was significant (p < 0.05). The recurrence rates were 31.2% and 8.3%, respectively (mean 17.5%). There was evidence of nerve damage to the superficial branch of the radial nerve in one patient (dorsal cyst) and to the palmar cutaneous branch of the median nerve in two patients (volar cysts). The mean nerve injury rate was 7.5%. In two patients with volar ganglia, the palmar superficial branch of the radial artery was lace-rated and had to be ligated. The significantly higher complication rate after excision of volar ganglia in contrast to dorsal ones might indicate that the former should be approached more carefully in contrast to dorsal ones and preferably by a senior surgeon. PMID- 11103489 TI - [Fibrocartilaginous dysplasia and tibia vara in young children: presentation of 3 new cases with spontaneous resolution and review of the literature]. AB - Focal fibrocartilaginous dysplasia is a rare and benign condition associated with unilateral tibia vara in toddlers. Three additional cases are reported in children aged 17, 18 and 26 months with spontaneous resolution. The authors discuss, based on a meta-analysis, the natural history of the disorder and review the diagnostic and pathophysiological problems. Since spontaneous remodeling of varus angulation and healing of the bony defect can be expected, biopsy and surgical intervention should be avoided. PMID- 11103490 TI - Long-term clinical results of cemented revision of primary cemented total hip arthroplasties. AB - Seventy-six patients who had undergone revision of a cemented total hip replacement were reviewed with an average follow-up of almost ten years. The average age at primary total hip replacement (PTHR) was 63.3 years. The average time between primary total hip replacement and revision was 62.5 months. Revision surgery was performed without using special techniques such as acetabulum reconstruction or femoral bone grafting. We evaluated patients pre- and postoperatively using the Merle d'Aubigne-Postel(M d'A) hip score. Clinically we observed an improvement of the hip score after total hip revision, particularly regarding pain. Thirty hips required a second, and six a third revision. If re revision is used as an end-point, our results are unsatisfactory, as we had a cumulative failure rate of 54% after 12 years. This is mainly due to not using special techniques adapted to revision situations. PMID- 11103491 TI - [Malleolar fractures. Predictive factors for secondary osteoarthritis. Retrospective study of 32 cases]. AB - The authors report a series of 32 ankle fractures treated by internal fixation and reviewed with a follow-up of more than 15 years. The series includes 12 fibular, 14 bimalleolar and 6 trimalleolar fractures. Following Weber's classification, there were 4 type A, 18 type B and 10 type C fractures. The postoperative x-ray showed 28 anatomy reductions; shortening of the fibula from 3 to 5 mm was noted in 4 cases. Clinical results were evaluated according to Kitaoka's criteria, and radiological results according to Magnusson's criteria. Statistical analysis was made with a Chi-square test. The retrospective review at an average follow-up of 15 years showed 19 painfree ankles, normal mobility in 22 cases, absence of edema in 18. The shoe-wear was normal in 30 cases. Walking had returned to normal in 23 cases but radiography showed narrowing of the tibiotalar joint line in 12 cases and lengthening of the medial malleolus in 16. Narrowing of the tibiotalar joint space was associated with lengthening of the medial malleolus in 10 cases. The objective results were rated as follows: 23 good, 8 fair, and 1 poor. With a follow-up of 15 years, we noted degenerative changes in the ankle in 37% of cases in spite of an anatomic reconstruction which had been perfect in 28. Shortening of the fibula, observed in 4 cases, was associated with subsequent ossification below the medial malleolus corresponding to avulsion of the non sutured medial collateral ligament. Nevertheless, degenerative changes of the ankle were clinically well-tolerated. The long term result of internal fixation of malleolar fractures was good. This was achieved only through perfect restoration of the joint anatomy. Contrary to other series, non-operative repair of the medial collateral ligament was associated with long team degenerative changes and reduced mobility of the joint. We therefore now advocate surgical repair of the medial collateral ligament. PMID- 11103492 TI - A retroperitoneal tumor as a late complication of the use of bone wax. AB - Ordinary bone wax was used to stop bleeding from the iliac crest after procurement of autogenous bone graft harvesting. This gave rise to a large, symptomatic retroperitoneal tumor, which had to be removed operatively 19 years later. Microscopically, a bone wax granuloma was diagnosed. As far as the authors know this is the first case reported with such late and severe clinical complications after the use of bone wax. PMID- 11103493 TI - Atlanto axial instability due to neurofibromatosis: case report. AB - Neurofibromatosis is an autosomal dominant genetic disease, characterized by cafe au lait spots, neurofibromas and several bony anomalies. Deformities of the spine are the most frequent alterations. Involvement of the cervical spine has been studied less frequently. The case of a 16-year-old male patient affected by neurofibromatosis, with cervical pain without neurological symptoms is presented. X-rays, CT-scan and MRI demonstrated the presence of cervical kyphosis, occipitoaxial instability and atlantoaxial instability with subluxation. Posterior occipito-C2 fusion was performed with prior placement of a halo-vest. The outcome at four years was good with solid occipito axial fusion, moderate loss of cervical spine flexion and moderate-to-severe limitation of cervical spine rotation. The incidence and variety of alterations of the cervical spine in patients affected with neurofibromatosis is discussed, as well as the results obtained by the treatment. PMID- 11103494 TI - Reconstructive surgery for a defect in the shaft of the ulna due to osteomyelitis. Long-term result of a case. AB - An eight and a half-year-old boy suffered from chronic osteomyelitis of the left ulna with sinuses, destruction of the middle three-quarters with the presence of necrotic bone and posterolateral dislocation of the radial head. The operative treatment included sequestrectomy and gradual reduction of the radial head after application of an Anderson apparatus. In a second procedure a corticocancellous tibial bone graft was used to bridge the ulnar gap, and later the redislocated radial head was excised. At the latest follow-up, 45 years postoperatively, the limb is fit with normal muscle strength and very satisfactory motion of the elbow and wrist joints, and the patient works as a hard manual laborer. PMID- 11103495 TI - Extraskeletal chondroma, a rare soft tissue tumor. Case report. AB - A rare case of a chondroma of soft tissue located in the first metatarsophalangeal joint of the right foot in a 65-year-old male patient is reported. Defining elements of the tumor under study are: 1) the asymptomatic and harmless clinical course 2) the lack of connection between the tumor and the underlying bone 3) the slow tumor development 4) the absence of age and sex predominance and 5) the characteristic tumor histological picture. PMID- 11103496 TI - Bilateral total hip replacement in pseudoachondroplasia. AB - Pseudoachondroplasia is an inherited skeletal dysplasia with short-limbed dwarfism and early onset of osteoarthritis. A 29-year-old pseudoachondroplastic woman presented with progressively painful hips secondary to severe osteoarthritis of both joints, so that total joint replacements were necessary to restore her mobility and quality of life. The implants inserted had to be specifically manufactured in accordance with the individual geometry and reduced bone size. In addition, the implants mechanical resistance to dynamic loading conditions had to be tested prior to total hip replacement surgery. PMID- 11103497 TI - Shortening of the calcaneus treated with Ilizarov's method. AB - The author reports the case of a sixteen year old young lady with heel shortening, due to a growth arrest of the calcaneal epiphysis, secondary to its septic involvement by a heel puncture. At the age of sixteen a lengthening according to Ilizarov's method was performed, using progressive distraction of an osteotomy of the calcaneal tuberosity. The result was satisfying with a cosmetical improvement, a painless function and a more comfortable shoe fitting. PMID- 11103498 TI - [Molecular biology in bladder cancer]. AB - Progress of molecular biology with regard to etiopathogeny of tumours in general, and cancer of the bladder in particular, is taking place at such a vertiginous pace that practising urologists find themselves overwhelmed in terms of their ability to learn and keep updated in so complex a subject. The understanding of certain molecular factors with critical influence on the formation, growth and progression of a tumour of the bladder, is forcing us to make unbiased assessments on the role they will play in the evolution and survival of this neoplasia. It is anticipate they will be much more reliable than traditionally established morphological factors such as grade and stage. We also include a literature review with an analysis and elucidation of the role played by oncogenes, tumor suppressor genes, vascular density markers, telomerase etc., in the formation and growth of cancer of the bladder and their likely relationships with already established clinico-pathological factors. PMID- 11103499 TI - [Intracranial metastasis in prostatic cancer]. AB - The intracranial metastasis due to prostatic adenocarcinoma are quite rare, inside them, the ones placed in the parasellar region on the cranial base are exceptional. There are only 3 clinical cases found in the literature consulted, now we report here two more cases and we review the etiopathogenia, clinic presentation, diagnosis and treatment for this type of lesions. Usually there are very undifferentiated neoplasms, developed stages and with multiple metastasis at others levels. A patient with prostatic carcinoma known and neurological signs we should suspect the presence of intracranial metastasis. The diagnosis is made with image tests (basically with CT and MRI), being necessary in some cases the histological confirmation with a biopsy. Although the prognostic of these patients (less than 6 months in our cases) depends more of the evolutive stage of the illness than the type of treatment that the patients will be someated, we should establish it rapidly, on this way we can revert the neurological status and we will improve the quality of life of these patients. PMID- 11103500 TI - [Bilateral germ tumors of the testis. Report of 5 cases and review of the literature]. AB - OBJECTIVE: To know the prevalence of the bilateral germ cell tumours of testis diagnosed in our Department and to review the literature. MATERIAL AND METHODS: 64 patients diagnosed of a germ cell tumour of the testis were followed during an average period of 51.4 months (1-168 months). RESULTS: 5 (7.8%) patients developed a second germ cell testicular tumour. In one patient the tumours were synchronous while in the remaining four were metachronous, occurring in an average interval of 59 months. One patient with a metachronous tumour died as consequence of the second tumour. In two of the five patients risk factors were identified, one presented testicular atrophy and the second referred history of undescended testis. DISCUSSION: The probability of developing a germinal testis tumour between the patients with history of a previous germ cell tumour of the testis is sensibly greater than between the general population. The prevalence of the bilateral tumours of the testis oscillates between 1-5% and approximately 75% will be metachronous. The principal factor that can predict the appearance of a second testicular tumour is the presence of the carcinoma in situ (Cis) in the contralateral testicular biopsy. Except in cases of testicular atrophy or previous history of undescended testis we do not recommend routine biopsy of the other testis. PMID- 11103501 TI - [Differential traits of prostatic cancer detected through an early screening program versus the one detected during urologic consultation]. AB - OBJECTIVE: To compare some features of prostate cancers (PCa) detected in a screening program, versus cancers diagnosed in an outpatient clinic. MATERIAL AND METHODS: Retrospective study of 393 patients with biopsy evidence of PCa: 93 (23.7%) from a screening campaign, and 300 (76.3%) detected in an outpatient Urology clinic. Features studied at the moment of diagnosis were age, PSA, digital rectal examination (DRE), transrectal ultrasound (TRUS) characteristics and volume stimation, PSA density (PSAD), clinical stage and Gleason score. A comparison was established between the two groups of patients taking into account the mentioned parameters. RESULTS: A higher age, PSA and DPSA values were found among cancers detected out of the screening program. A greater probability of abnormal DRE and a more advanced clinical stage was also noted. In the screening group, 78.5% of the cancers were localized and 8.6% metastatic. In the outpatient clinic group, the proportions were 50.7% and 26%, respectively. No differences were found with respect to TRUS findings, prostate volume, or Gleason score. CONCLUSIONS: Cancers detected in screening programs are found in earlier stages. Nevertheless, results from long term randomized studies are necessary to verify if these data really mean that a disease-specific mortality reduction can be achieved. PMID- 11103502 TI - [Repetition of ultrasonography-guided prostatic biopsy for the detection of cancer. Study of a series of 192 re-biopsied patients]. AB - PURPOSE: We reviewed the result of transrectal ultrasound (TRUS)-guided needle biopsies to find the re-biopsy criteria, emphasizing on the Focal Glandular Atypia (FGA) histological changes. MATERIAL AND METHOD: 192 cases were selected, from a total of 1957 patients older than 50, re-biopsied because of high PSA levels and/or abnormal DRE, or because of the histological findings on initial biopsies (high grade PIN and/or FGA). The results are related to the serum PSA levels and DRE characteristics. RESULTS: A 38.83% global positivity for cancer was obtained and 27.08% for re-biopsy. When the first biopsy was negative, the positivity of the re-biopsies was 19.37%; if it was negative for cancer but had high grade PIN and/or FGA changes, the positivity was 65.62%, being higher in FGA changes than in the PIN cases (68.00% vs. 57.14%). The abnormal DRE raised the positivity rate from 17.82% to, 35.75%. CONCLUSIONS: The positivity was especially related to abnormal DRE and/or PSA > or = 10 ng/ml. The tumor rate detected at second and third or successive biopsies was similar (19.28% vs 21.74%). The FGA changes (3.47% globally) had a cancer predictive value of 65.62%. We recommend re-biopsy in all patients with FGA changes. PMID- 11103503 TI - [Malignant neoplasms of the penis]. AB - OBJECTIVE: To assess the behaviour and management of these uncommon neoplasias. MATERIAL AND METHODS: Between March 1975 and July 1999, a total of 95 malignant neoplasias of the penis were diagnosed and treated by our unit. Patients mean age was 62 years (28 to 87 years). A retrospective analysis of any associated lesions, biological behaviour of the various neoplasias, as well as therapies used is carried out. RESULTS: The squamous carcinoma of the penis (SCP) is the most frequent pathohistological entity entailing 78 cases (82%), followed by verrucous carcinoma (VC) 13 cases (13.5%), basal cells carcinoma 1 case, and melanoma, lymphoma and penile metastasis 1 case each. There is a significant presence of associated lesions with marked predominance of phimosis. The most frequent reason for the call is an injury of the penis (74 cases; 78%). Treatment was mainly partial penectomy (51 patients; 53.7%), followed by conservative treatment in 28 cases (30%). Inguinal lymphadenectomy was performed in 13 patients (14%), due to either a positive nodular biopsy or a persistent adenopathy following antibiotic therapy. CONCLUSIONS: Neoplasias showing superior biological behaviour are basal cell carcinoma of the penis, and verrucous carcinoma. Prognosis in SCP is based on pathological status and node involvement. Patients with pT1 tumours showed no metastatic adenopathies after follow-up regardless of cytological grading, and are therefore candidates to watchful waiting with regular monitoring. Melanoma of the penis is a highly aggressive tumour due to its high metastatic capacity with a poor prognosis. PMID- 11103504 TI - [Structural changes in the ileal mucosa of urinary conduits]. AB - OBJECTIVES: To study the changes found in the terminal ileum mucosa in contact with urine in patients with skin ureteroileostomy following cystectomy due to infiltrant carcinoma of the bladder. MATERIAL AND METHODS: 21 biopsies of gut mucosa were performed in as many patients. Measurements included height of intestinal villi (average values -AV-: 350-300 mu), height of crypts (AV: 70-100 mu), crypt/villus ratio (AV: 0.2), goblet cells/enterocytes ratio, presence of lymphangiectasis and inflammatory infiltrate. Also the existence of lab changes were measured with ionogram and venous gasometry. RESULTS: Mean age of patients was 65.2 years +/- 7.4 SD. Males 66.7%. Time of evolution with urinary by-pass was 59.5 months +/- 53.2 SD. Mean height of villi 178.2 mu +/- 70.2 SD, mean height of crypts 290.9 mu +/- 114.4 SD and mean crypt/villus ratio 4.2 +/- 9.2 SD. Submucous inflammatory infiltrate was mild in 57.1%, moderate in 23.8%, and severe in 19.0%. Only 2 cases had lymphangiectasis images. Goblet cells/enterocytes ratio was 3.3 +/- 1.3 SD. No correlation was seen between time of evolution of urinary by-pass with the various changes in gut mucosa or between degree of mucosal atrophy and existence of metabolic disorders. CONCLUSIONS: Changes in the terminal ileum mucosa in patients with skin ureteroileostomy-like urinary by-pass are characterised by a marked atrophy of intestinal villi with increased crypt length, increased crypt/villus ratio and presence of mild-to moderate inflammatory infiltrate. There is also an increase of goblet cells in detriment of enterocytes. All these changes are independent from the time of evolution of patients with urinary by-pass. PMID- 11103505 TI - [Tumor thrombosis of the left renal vein and inferior vena cava secondary to renal cell carcinoma. Findings with ultrasonography, Echo-Doppler, and computerized tomography]. AB - Renal cell carcinoma represents a 2.5-3% of all neoplastic processes, usually seen un patients older than 50 years. 60-75% are resectable at diagnosis, representing local or metastatic advanced disease the rest of them. This tumor tends to spread intravascularly, leading to tumoral thrombosis within the inferior caval vein (ICV) and renal vein 4-10% and 21-35% of cases, respectively. As the only effective treatment is surgical resection, preoperative determining of the thrombus extension is crucial. Thus, an accurate radiological study including ultrasound, doppler sonography, computed tomography and/or Magnetic Resonance, is key for these patients. We present a 49 year-old patient with renal cell carcinoma and associated tumoral thrombosis in inferior caval vein and left renal vein; we provide the most significant figures, explaining its most characteristic radiological findings. PMID- 11103506 TI - [Paratesticular lipoma]. AB - Among the neoplasias affecting the scrotum and other elements contained within, testicular tumours are the most common; paratesticular neoplasias account for about 2%. 70% of them are benign tumours, lipoma being one the most frequent types. We contribute a new case of paratesticular lipoma in a 45-year old patient, who presented with scrotal complaints. Physical examination found fibroelastic scrotal tumoration; diagnosis being aided with ultrasound and CAT that showed fat tissue radiographic patterns. Complete removal of tumoration and subsequent pathoanatomical study confirmed the diagnosis of paratesticular lipoma. A literature review on the subject is included with reflections on the clinical, diagnostic and therapeutical aspects. PMID- 11103507 TI - [Congenital bladder diverticulum and Ehlers-Danlos syndrome: an unusual association]. AB - The existence of a vesical diverticulum in the context of a congenital connective tissue disorder such as Ehlers-Danlos syndrome led us to consider the possibility of a relationship. Four types of diverticula can be found in the literature: congenital, acquired, iatrogenic and syndrome-associated. Within the later, Ehlers-Danlos syndromes type IV and IX, even type V, are associated to the existence of vesical diverticula. The potential spontaneous rupture of the diverticulum is a typical feature, as well as post-surgery relapse. The attitude towards such diverticula should be one of watchful waiting, and simple, plasty free diverticulectomy on the bladder's neck is indicated when performing a surgical procedure. PMID- 11103508 TI - [Self-administration of an oil solution in the penis]. AB - We report the case of a patient who has autoinjected himself the penis and the scrotum with a solution constituted by petroleum, vaseline, oxygenated water and ketoconazol gel. Treatment consists in the excision of the injected deposit of lipid material and antibiotic therapy with good result at 4 months. We review the existing literature related with this exceptional pathology. PMID- 11103509 TI - [Solitary renal metastasis of primary esophageal cancer]. AB - We present an unusual case of single kidney metastasis from an primary esophageal neoplasm. Its main clinical and diagnostic topics are described. They made us consider him as a good candidate for surgery. The pathological study of the nephrectomy specimen was surprising. The postoperative period was unfortunate, however. PMID- 11103510 TI - [Granulomatous orchitis]. AB - Granulomatous orchitis is an inflammatory change of the testis. This is a rare lesion of unknown etiology. Usually are unilateral. The clinical appearance varies, and it is difficult to differentiate from testicular cancer. The diagnosis usually being made on histological examination after orchiectomy. Our paper report one case of granulomatous orchitis in a 27 year old patient. A review of the literature is made on the diagnosis and pathogenesis. PMID- 11103511 TI - [Retroperitoneal benign schwannoma. Report of a new case]. AB - The schwannoma is a tumor resulting from the Schwann cells of neural shwath, being its retroperitoneal localization quite unusual. This tumor is clinically unspecified and in most of the cases it originates symptoms coming from the compression of the close structures when its localization is retroperitoneal. Its diagnosis is quite often fortuitous being confirmed by anatomopathological study afterwards. The treatment is surgical radical exeresis with subsequent followup. We report a new case of this uncommon retroperitoneal pathology in a female patient showing a nonspecific clinic. Two years after the surgery she remains asymptomatic without any radiological evidence of recidive. PMID- 11103512 TI - [Rupture of penile dorsal vein]. PMID- 11103513 TI - Angiogenesis and vascular regression in the ovary. AB - Angiogenesis is prominent during development and downregulated in the adult. Strictly controlled angiogenesis in the healthy adult occurs cyclically in the ovary and corpus luteum, which therefore make an excellent model with which to study vascular growth. Dysfunctional or uncontrolled angiogenesis is involved in a number of diseases and is responsible for growth and dissemination of tumours. This review focuses on the following aspects of the ovary: the gross and microscopical anatomy of the blood vessels, described mainly--but not exclusively -in the bovine; vascularization of the follicle before and after ovulation; angiogenesis in the developing and the mature corpus luteum as well as in the corpus luteum of pregnancy. The potential mechanisms of vascular regression during luteolysis and the potential role of vascular growth in dominance and atresia of follicles will be described. Furthermore, recent research on ovarian angiogenic and potential anti-angiogenic factors including fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), angiopoietin and metalloproteinase inhibitor will be presented. Finally, the role of hormones including FSH, LH, sexual steroids, prostaglandins, prolactin, oxytocin and activin/inhibin in ovarian angiogenesis will be summarized. Future research is likely to yield valuable information that can contribute to the development of novel therapeutic strategies for the treatment of diseases characterized by disregulated angiogenesis and vascular regression. PMID- 11103514 TI - Immunohistochemical localization of inhibin and steroidogenic enzymes in the ovary of common tree shrew (Tupaia glis) and northern smooth-tailed tree shrew (Dendrogale murina). AB - To study the ovarian function of the Order Scandentia, the localization of inhibin and steroidogenic enzymes (3 beta-hydroxysteroid dehydrogenase/isomerase and aromatase) in the ovaries of common tree shrew (Tupaia glis) and northern smooth-tailed tree shrew (Dendrogale murina) was immunohistochemically analysed. As in the results reported for other mammals, inhibin alpha-chain was localized in the follicular epithelium of secondary or Graafian follicles in the two species. The localization of aromatase in the ovary of these two species, however, was different. In the common tree shrew, the aromatase was localized in the thecal cells, whilst in other mammals it is localized in the granulosa cells. These results indicate that in the ovary of the common tree shrew, the oestradiol may be synthesized in the thecal cells. PMID- 11103515 TI - Arteries of the hindfoot of the llama (Lama glama). AB - The objective of this study was to describe the arterial distribution of the hindfoot of the llama (Lama glama). Ten adult llamas, preserved in 6% formalin solution at 0 degree C, were dissected. The arterial system was perfused with a solution of 14% coloured plaster; the venous system was perfused with a solution of 17% coloured industrial gelatin. Angiographies were also obtained. In the llama, the arterial distribution is of the saphenous type and this simple sort of irrigation could be used as a didactic model. The caudal branch of the saphenous artery divides into the small lateral plantar artery and the larger medial plantar artery, which continues as the plantar common digital artery III, and it is the main blood supply of the hindfoot. The dorsal pedal artery is underdeveloped and the perforating tarsal artery does not exist in this species. The plantar common digital artery III divides into the plantar proper digital II, III and IV. Branches from the plantar proper digital artery III supply the digits. We compared the arterial distribution of the hindfoot of the llama with that of other domestic animals including the one-humped camel (Camelus dromedarius). PMID- 11103516 TI - Comparison of localization of the neurokinin 1 receptor with AMPA-type glutamate receptors in the rat spinal cord. AB - With the cloning of the alpha-amino-3-hydroxy-5-methyl-4-isaxole propionic acid (AMPA)-type receptor subunits, it is now possible to localize these receptor subunits in the spinal cord. Comparison of the neurokinin 1 receptor distribution with that of non-N-methyl-D-aspartate glutamate receptor subunits (GluR1-4), considered to be AMPA-type, was investigated in rat spinal cord by immunocytochemical methods. Different patterns of immunolabelling were observed with the antibodies to the GluR1, GluR2/3 and GluR4 subunits in the lumbar spinal cord. Immunolabelling with antibodies to both GluR1 and GluR2/3 revealed intensive staining in the dorsal horn, while staining for GluR2/3 and GluR4 was dense in the motor neurons of the ventral horn. These results suggest that in the rat spinal cord AMPA-type receptors vary their composition according to the region where they are expressed. Neurokinin-1-receptor-expressing neurons in the dorsal horn do not appear to express the GluR4 subunit, however, whether they express the GluR1, GluR2/3 receptors subunits could not be determined. PMID- 11103517 TI - Morphological, histochemical and morphometric study of the myotomal muscle tissue of the pacu (Piaractus mesopotamicus Holmberg 1887: Serrasalminae, Characidae, Teleostei). AB - Histochemical, ultrastructural and morphometric methods were used to study growth patterns of red, pink and white muscle fibres and their relation to body weight and total length in the fast-growing freshwater fish Piaractus mesopotamicus Holmberg. The correlations amongst body weight, body length and diameter of red, pink and white fibres were low. From 10-15 to 40-50 cm, body weight increased 102.7 times, while the diameter of each type of fibre increased by factors of 0.94, 0.74 and 0.70, respectively. Muscle fibres revealed different morphological and histochemical stages of maturation. The frequencies of < 20 microns fibres of red, pink and white muscle tissue in the youngest and oldest classes were 64.5 and 11.0, 38.2 and 7.7 and 24.0 and 1.4%, respectively. In 30-40 cm fish, the frequency of < 20 microns fibres in the red and pink tissue was 24.5 and 25.5%, while in the white tissue it was 11.5%. During sexual maturity (40-50 cm), the recruitment of < 20 microns fibres in white muscle was 1.4%. Muscle fibres of this species showed continuous growth by both hyperplastic and hypertrophic mechanisms, and hyperplasia was particularly active in the juvenile phase. PMID- 11103518 TI - Arterial vascularization of the uropygial glands (Gl. uropygialis) in geese (Anser anser) and ducks (Anas platyrhynches). AB - In the present study, arterial vascularization of the uropygial glands (Gl. uropygialis) of 10 adult geese (Anser anser) and 10 adult ducks (Anas platyrhynches) were studied. Takilon was injected into the median coccygeal arteries of six specimens from each species, and Latex (a natural rubber with ammonia) into those of four specimens. Takilon-injected specimens were corrosion casted, and arteries nourishing the gland were revealed via dissection. Vascularization of the uropygial glands of both the goose and the duck was observed to be the right (a. gl. uropygii dextra), left (a. gl. uropygi sinistra) and ventral (a. gl. uropygi ventralis) glandular uropygial arteries, arising from the median coccygeal (a. coccygea media) artery. Both the right and left glandular uropygial arteries were observed, divided into four branches as follows; muscular ramus (ramus muscularis), medial ramus (ramus medialis), ventral ramus (ramus ventralis) and lateral ramus (ramus lateralis). Of these, as the lateral, medial and ventral branches feed the gland, the muscular branch provides blood for the lateral coccygeal (m. coccygealis lateralis) and levator coccygeal (m. levator coccygealis) muscles, and the skin. Among the arteries mentioned above, anastomosis between the first and the second branches of the right ventral uropygial arteries in the five geese and five ducks was found. PMID- 11103519 TI - Computed tomography of normal cranioencephalic structures in two horses. AB - The purpose of this investigation was to define the anatomy of the cranioencephalic structures in horses using computed tomography (CT). Transverse images of two isolated equine cadaver heads were obtained using a Toshiba 600 HQ (third-generation equipment TCT). CT images were compared to corresponding frozen cross-sections of the cadaver head. Relevant anatomical structures were identified and labelled at each level. The resulting images provided excellent anatomic detail of the structures of the central nervous system and associated formations. Annotated CT images from this study are intended as a reference for clinical CT imaging studies of the equine head. PMID- 11103520 TI - The vascular bed in the rabbit ear: microangiography and scanning electron microscopy of vascular corrosion casts. AB - The vascular bed of the rabbit ear has been studied with light microscopy, microangiography and scanning electron microscopy of resin casts in a series of 20 ears. The major arteries supplying the ear were the central and rostral auricular branches of the caudal auricular artery. The caudal auricular branch was not observed, except as a small vessel supplying the rostral auricular base. The central auricular branch supplied blood to most of the auricular integument and was surrounded by capillaries extended from those supplying the skin. The periarterial capillaries formed a fine, compact network in the tunica media and were closely related to the central auricular branch. Evidence is presented suggesting that this vascular mechanism has a counter-current heat-exchange function for regulating body temperature. The artery had the well-developed internal elastic lamina and intimal cushions that regulate blood flow at the branching sites. A number of arteriovenous anastomoses were also observed between arterioles and venules, particularly in marginal regions of the ear. The intimal cushions and arteriovenous anastomoses might play an additional role in thermoregulation by regulating local blood flow in the ear. PMID- 11103521 TI - The ultrastructure of the subcommissural organ ependyma of the goat. AB - The cells of pseudostratified columnar ciliated ependyma of the subcommissural organ in the goat were classified into two types on the basis of the distribution of chromatin material and nuclear clefts. Amongst the cell organelles the endoplasmic reticulum was highly developed and formed three types of Nebenkerne systems. The type-I Nebenkerne had spiral concentric lamellae and was associated with round lipid droplets. The type-II Nebenkerne, with widely spaced coils, was expanded towards its central and peripheral parts. The type-III Nebenkerne, composed of a meshwork of lamellae, was modified into a vacuolated form. The concentration of mitochondria was greatly increased towards the basal processes of the ependymal cells. The inclusion bodies included small electron-dense bodies, osmiophilic asteroid droplets, large round to spherical bodies and large round osmiophilic bodies with inner structures. PMID- 11103522 TI - Morphological study of metaphase-specific apoptosis in MRL mouse testis. AB - Apoptosis of male germ cells is a complex phenomenon in many animal species. Understanding its mechanisms could be useful in the diagnosis and therapy of male infertility. To examine the differences of distribution of apoptosis among mouse strains, the terminal transferase-mediated nick end labelling (TUNEL) method was employed. In the testes of MRL mice, many TUNEL-positive cells were identified at the metaphases of meiotic spermatocytes. Morphometrical analysis revealed that metaphase-specific apoptosis occurred at the region between secondary spermatocytes and step 1 spermatids in stage XII seminiferous tubules. In the investigation of the developing first-wave of seminiferous tubules, there were some metaphases showing apoptotic morphology prior to becoming secondary spermatocytes. Details of the apoptotic structure revealed by electron microscopy showed that cellular arrest occurred after the beginning of the M phase of the cell cycle. These results suggested that metaphase-specific apoptosis in the testis of the MRL mouse strain took place at least at the first meiotic division, perhaps showing the spindle assembly checkpoint of the cell cycle. PMID- 11103523 TI - Sulphates, nitrates, and chlorides in particle fractions of different size. AB - This paper describes a pilot study of chloride, nitrate, and sulphate content in thoracic and high-risk respirable fractions of airborne particles. Samples were collected at one measuring site in Zagreb in autumn 1998 and spring 1999. The results showed that almost total chloride, nitrate, and sulphate content was present in the respirable particle fraction. The average mass contribution of these pollutants to the particle mass amounted to 25%. Although chloride mass concentrations were quite low, the findings indicated that all pollutants originated from the same source. PMID- 11103524 TI - Distribution of airborne bacteria in swine housing facilities and their immediate environment. AB - This paper describes a bacteriological analysis of air samples taken from swine housing facilities and the immediate environment. The air volume of the samples was pre-programmed by a standard air sampler (MAS-100, Merck) and was directly impacted onto the bacteriologic agar surface (Petri dishes, standard diameter of 90 mm). The bacterial contamination in forty-eight samples was 2.59 x 10(5) CFU/m3 (ranging from 8.46 x 10(4) to 5.30 x 10(5) CFU/m3). Potentially pathogenic bacterial agents predominated in all samples (100%), while primarily pathogenic bacteria were isolated in a minor proportion of samples (33%-66%). Airborne bacterial contamination in samples (N = 16) obtained from emptied facilities ranged from 1.8 x 10(3) CFU/m3 (that is, after coarse mechanical washing) to 0.8 x 10(2) CFU/m3 (upon completion of disinfection). Control measurements at different locations and distance from the farm (N = 32) pointed to the presence of non-pathogenic airborne bacteria, ranging from 1.55 x 10(2) to 3.70 x 10(2) CFU/m3. The results of this preliminary study showed that the emission of potentially pathogenic bacteria from animal housing facilities to the immediate farm environment via aerosol was very low. PMID- 11103525 TI - A method for monitoring deposited matter in forest ecosystems. AB - Systematic and elaborate multidisciplinary research of pedunculate oak and common hornbeam began in the Forest Research Institute in Jastrebarsko in 1991. That led to a development of a method for monitoring deposition in forest ecosystems which is now closely related to the International Co-operative Programme on Assessment and Monitoring of Air Pollution Effects on Forest. Several forest communities on the territory of Croatia were monitored for Cl-, SO4(2-)-S, NH4(+)-N, NO3(-)-N, Na+, K+, Ca2+, and Mg2+ using that method. Samples were collected by funnels (bulks). Plastic lysimeters were placed in the soil at the depth of 10 cm or beneath the humus layer. They served to collect liquid that seeped into the soil (seepage). Sampling was carried out once a month or once in three months. The results show that the monitored forest ecosystems absorbed more deposited particles (wet and dry sedimentation) than control areas. PMID- 11103527 TI - Levels of nitrogen dioxide in the Zagreb air. AB - Surveillance of NO2 in Zagreb started in 1994 at five measuring sites. This paper presents the levels and trends of NO2 annual mean values for the period between 1994 and 1998. The obtained data are compared with the recommended and limit values according to the Law on Air Quality Protection in Croatia and the Ordinance on Recommended and Limit Air Quality Values. For the past five years, ever since the measurement commenced, the concentration levels of NO2 in Zagreb have kept between the national recommended and limit values at all measuring sites, indicating that the air has been moderately polluted. This paper also shows the variations of monthly mean values through 1998 at all sites. The levels of NO2 in the northern part of Zagreb were measured at two distances from traffic. Daily variations in the levels were measured near the road and presented as hourly averages for workdays and weekends in winter and in summer reflecting the activity pattern of emission sources. PMID- 11103526 TI - The meaning of air quality and flue gas emission standards for public acceptance of new thermal power plants. AB - For the time being only 30-40% of the electric energy supply in Croatia comes from burning fossil fuel. New capacities of 800-1400 MW for the next decade will have to rely on the exclusive use of fossil fuels in thermal power plants (TPP). Public opinion will probably have a decisive influence on the issuing of construction permissions. The potential adverse effects on air seem to be the main argument against construction of TPPs. The priority is therefore to unambiguously state what air quality is warranted in the influenced area for the whole operation period of a TPP. It is important that the public should understand the real meaning of current air quality standards and emission limits. The only known way to do it today is through comparison with the corresponding standards and limits accepted worldwide. This paper discusses some important aspects of such comparison. PMID- 11103528 TI - Lead, manganese, and cadmium content in PM10 and PM2.5 particle fractions--a pilot study. AB - This paper describes a pilot study of lead, manganese, and cadmium content in thoracic particle fraction and high-risk respirable fraction of airborne particles. Samples were collected at one measuring site in Zagreb during autumn 1998 and spring 1999. The results show that the total heavy metal contents is found in the high-risk respirable particle fraction, and point to two main pollution sources, the first for lead and manganese and the other for cadmium. PMID- 11103529 TI - Airborne metal concentrations in shipyard environment. AB - Protection against corrosion in the shipyard is a source of airborne particles. From October 1996 to September 1997 samples of suspended particles (1 site) and dustfall (6 sites) were collected in the vicinity of a repairs shipyard situated in the martinscica Cove, east of the city of Rijeka, Croatia. Collected samples were analysed for lead, cadmium, iron, copper, and zinc content. Though annual mean concentrations of suspended particles, lead, and cadmium kept below the guideline values, the metal contents were generally higher than values measured in the city centre. The correlation between the quantity of abrasives used at the shipyard and monthly mean concentrations of all parameters except cadmium suggests that the shipyard was the main source of those pollutants. The annual mean, as well as maximum monthly amount of dustfall at the site next to the shipyard zone exceeded the national limit values, indicating considerable pollution of this area with coarse particles. The annual mean quantity of lead in dustfall exceeded the guideline values at the same site. The content of metals occasionally observed in dustfall at particular sites surrounding the shipyard depended on the location of corrosion protection activities and meterological conditions within the Martinscica Cove. PMID- 11103530 TI - Ambient air quality programmes for health impact assessment in the WHO European region. AB - An important aim of air quality assessment is to provide information about population exposure and health impact assessment. Numerous epidemiological studies have already shown that exposure to excessive levels of ambient air pollutants are associated with either acute or chronic health effects. Until recently, the adequacy of monitoring population exposure in relation to quantitative assessment of health effects of air pollution was rarely considered in ambient air monitoring strategies. This made the formulation of health-related recommendations to risk management difficult and weakens preventive and other measures to reduce adverse health effects of air pollution. To improve local and national capacities for health impact assessment, the European Centre for Environment and Health of the World Health Organization has prepared methodology guidelines concerning selected aspects of air monitoring. The WHO Collaborating Centre for Air Quality Management and Air Pollution Control support efforts in line with international programmes on quality assurance and control for Europe. PMID- 11103531 TI - Fractionation schedules and radiation therapy waiting lists. PMID- 11103532 TI - Quality assurance: Hodgkin's disease and beyond. PMID- 11103533 TI - Synovial, chondropathic, depositional: the radiological categories of arthritis. A review. AB - The accepted optimum logic frame for complex diagnostic problems is sequential hypothesis testing. The X-ray diagnosis of arthritis has now become sufficiently complex to make this the procedure of choice, but standard classifications of arthritis lack the discriminatory power needed for its effective deployment. This obstacle can be overcome if these standard classifications are replaced by a more discriminant classification based on radiologically identifiable discriminators. This classification divides arthritis into three groups: synovial, chondropathic and depositional. The initial categorization is usually made fairly simply from two or three markers, of which the most important is the site of any erosions present. Once categorized, the subsequent analysis can proceed in an approximately binary manner using the discriminators appropriate for that category. The use of this radiological classification simplifies the diagnostic approach, reduces the workload, and provides the algorithm needed for sequential hypothesis testing. PMID- 11103534 TI - Magnetic resonance findings in knee dislocation: pictorial essay. AB - Magnetic resonance images depicting the range of soft-tissue and bony injuries associated with knee dislocation are presented. Those injuries that are less well described or more difficult to diagnose are emphasized. PMID- 11103535 TI - Analysis of intracranial mass lesions using proton magnetic resonance spectroscopy at 1.0 T: pictorial essay. AB - Proton MR spectroscopy has been used to characterize the spectroscopic profiles of a wide variety of neurological disease processes. In particular, the spectroscopic characteristics of a range of focal intracranial mass lesions have been described. The use of proton MR spectroscopy has until recently been restricted to major research centres with high-field MR scanners. The spectroscopic findings in a number of illustrative cases imaged at Royal Perth Hospital using a 1-T scanner are described here. The cases illustrated in the present paper confirm the clinical utility of proton MR spectroscopy as a supplement to conventional MR in routine radiological practice. PMID- 11103536 TI - Value of pharyngography in patients without suprasternal symptoms. AB - The purpose of the present paper was to determine if pharyngeal or cervical oesophageal lesions may present with distal symptoms. All patients presenting for barium swallow underwent examination of the pharynx and oesophagus. The pharyngeal examination included spot films of the pharynx as well as views of the pharyngo-oesophageal segment filmed at three frames per second. During the 18 month period of the present study interrogations were carried out to identify patients without symptoms in the cervical or suprasternal region. One hundred and twelve patients were identified; 58 were male and 54 were female. The age range was 18-84 years. Examinations revealed abnormalities within the pharynx in 42 patients (38%); of this group of 42, 34 also had an oesophageal abnormality. The majority of the pharyngeal findings were minor. There were, however, three patients who each had a pharyngeal abnormality (pharyngeal carcinoma, obstructive cricopharyngeal narrowing, pharyngo-oesophageal junction stricture) as well as an oesophageal lesion (hiatal hernia, achalasia, reflux oesophagitis), either of which may have been the source of the symptoms. The remaining eight patients (7%) of this group of 42 with detected pharyngeal abnormality had normal oesophageal examinations. Most of these were again minor changes and were unlikely to be significant. There was, however, one patient in whom the only abnormality was an infiltrative cancer of the posterior wall of the pharyngo-oesophageal junction. In conclusion, the identification of patients in the present study with pharyngeal lesions and without distal abnormal findings indicates that a proximal lesion may present with downstream symptoms. Furthermore, there were also patients in whom the examination found abnormalities in multiple segments of the pharynx and oesophagus. We suggest that examination of the pharynx is warranted as part of the barium swallow in patients without cervical or suprasternal symptoms. PMID- 11103537 TI - Modified computed tomography peritoneography: clinical utility in continuous ambulatory peritoneal dialysis patients. AB - The purpose of the present paper is to review the indications, results, clinical correlation and implications for management of a modified CT peritoneography (CTP) technique in the evaluation of continuous ambulatory peritoneal dialysis (CAPD) patients. Forty CTP in 33 patients were reviewed by two observers blinded to clinical history or outcome. The CTP technique included 100 mL non-ionic intraperitoneal contrast, 1 h of ambulation and prone positioning during the CT. No precontrast or delayed examinations were performed. A CAPD-related complication was diagnosed in 33 of 40 (82%) CTP. Twenty-nine CTP were performed for evaluation of suspected dialysate leaks. In this group there were 18 leaks, a total of seven umbilical hernias (five as isolated findings) and five normal studies. Nine of 18 (50%) leaks resolved with conservative management and six of nine patients (67%) of the remainder continued CAPD after appropriate surgery. Seven patients were evaluated for scrotal swelling (one bilateral), and there were five communicating hydroceles (all with inguinal fat herniation and all surgically confirmed) and three non-communicating hydroceles (none of which progressed). Umbilical hernias were present in nine of 33 patients (27%). No false positive or false negative studies were identified. The CTP technique described provides accurate diagnostic information in the management of common CAPD-related complications, particularly dialysate leak, genital swelling, abdominal wall hernias and peritoneal adhesions. Computed tomography peritoneography directs appropriate conservative or surgical management. PMID- 11103538 TI - Acute disseminated encephalomyelitis and multiple sclerosis: magnetic resonance imaging differentiation. AB - The study was undertaken to compare the MR imaging features of acute disseminated encephalomyelitis (ADEM) and multiple sclerosis (MS) in a country with a high prevalence of ADEM. Magnetic resonance scans from 33 patients diagnosed clinically with MS (14 patients) or ADEM (19 patients) were reviewed concurrently by two radiologists blinded to the clinical diagnosis. The size, site, morphology and pattern of brain and spinal cord involvement were recorded and the MR imaging diagnosis was compared with the clinical diagnosis. The MR imaging findings matched with the clinical diagnosis in 11 of 14 patients with MS (sensitivity = 78.6%), and with the clinical diagnosis in 15 of 18 patients with ADEM (sensitivity = 78.9%). Three patients had non-specific findings and in a further three patients discordant imaging features were present. One patient with imaging features typical of Balo's concentric sclerosis was diagnosed clinically as suffering from ADEM. In a country with a high prevalence of ADEM, the majority of patients with ADEM and MS can be differentiated on MR imaging. PMID- 11103539 TI - Magnetic resonance imaging of lipoma and liposarcoma: potential of short tau inversion recovery as a technique of fat suppression. AB - The present limited retrospective study was performed to assess MR imaging of lipomatous tumours of the musculoskeletal system and to evaluate the potential of the T2 short tau inversion-recovery (STIR) technique for differentiating lipomas from liposarcomas. Magnetic resonance imaging of 12 patients with lipomatous tumours of the musculoskeletal system (eight benign lipomas, three well differentiated liposarcomas and one myxoid liposarcoma) were reviewed. Benign lipomas were usually superficial and showed homogeneity on T1- and T2-weighted spin echo sequences. Full suppression at T2 STIR was readily demonstrated. In contrast, the liposarcomas in the present series were all deep-seated. Two well differentiated liposaromas showed homogeneity at long and short relaxation time (TR) but failed to show complete suppression at T2 STIR. One case of well differentiated liposarcoma (dedifferentiated liposarcoma) and one of myxoid liposarcoma showed mild and moderate heterogeneity at T1 and T2, respectively, and posed no difficulty in being diagnosed correctly. In conclusion, short and long TR in combination with T2 STIR show promise in differentiating benign from malignant lipomatous tumours of the musculoskeletal system, when taken in combination with the position of the tumour. PMID- 11103540 TI - Computed tomographic cephalometric analysis of Chinese patients with obstructive sleep apnoea. AB - Variations of craniofacial and upper airway structures are recognized to contribute to the phenomenon of obstructive sleep apnoea (OSA). Most previous cephalometric studies were performed using erect lateral radiographs in Caucasian patients. The present project aims to determine cephalometric measurements, utilizing CT, in normal Chinese subjects and in Chinese patients with OSA. Computed tomography of 25 patients with OSA (proven using overnight polysomnography), and of 25 age-, sex-, height-, bodyweight- and body mass index (BMI)-matched control subjects were prospectively performed. Thirteen standard bony and soft-tissue measurements were obtained from the CT lateral scout view of the head and neck, taken with each subject in the neutral supine position. The cross-sectional area was calculated at two axial levels (velopharynx and hypopharynx). The measurements in the two groups, OSA and control subjects, were compared. The measurements for hyoid position (P = 0.00), nasal cavity length (P = 0.01), mandibular prognathism (P = 0.05), tongue size (P = 0.02), oropharyngeal airway space (P = 0.02), posterior tongue airway space (P = 0.04) and cross sectional areas at the level of the velopharynx (P = 0.00) and hypopharynx (P = 0.01) differed significantly between the two groups. In conclusion, CT cephalometric measurements show that certain anatomical variations in the head and neck are likely to contribute to the pathogenesis of OSA in Chinese patients. PMID- 11103541 TI - Subsequent investigation and management of patients with intermediate-category and -probability ventilation-perfusion scintigraphy. AB - The authors wished to determine the proportion of patients with intermediate category and intermediate-probability ventilation-perfusion scintigraphy (IVQS) who proceed to further imaging for investigation of thromboembolism, to identify the defining clinical parameters and to determine the proportion of patients who have a definite imaging diagnosis of thromboembolism prior to discharge from hospital on anticoagulation therapy. One hundred and twelve VQS studies performed at the Flinders Medical Centre over a 9-month period were reported as having intermediate category and probability for pulmonary embolism. Medical case notes were available for review in 99 of these patients and from these the pretest clinical probability, subsequent patient progress and treatment were recorded. Eight cases were excluded because they were already receiving anticoagulation therapy. In the remaining 91 patients the pretest clinical probability was considered to be low in 25; intermediate in 30; and high in 36 cases. In total, 51.6% (n = 47) of these patients (8% (n = 2) with low, 66% (n = 20) with intermediate, and 69.4% (n = 25) with high pretest probability) proceeded to CT pulmonary angiography (CTPA) and/or lower limb duplex Doppler ultrasound (DUS) evaluation. Of the patients with IVQS results, 30.7% (n = 28) were evaluated with CTPA. No patient with a low, all patients with a high and 46% of patients with an intermediate pretest probability initially received anticoagulation therapy. This was discontinued in three patients with high and in 12 patients with intermediate clinical probability prior to discharge from hospital. Overall, 40% of patients discharged on anticoagulation therapy (including 39% of those with a high pretest probability) had a positive imaging diagnosis of thromboembolism The results suggest that, although the majority of patients with intermediate-to-high pretest probability and IVQS proceed to further imaging investigation, CTPA is relatively underused in this group. Most patients with a high pretest clinical probability receive anticoagulation therapy irrespective of imaging findings, and less than half of all patients discharged from hospital on anticoagulation therapy have a positive imaging diagnosis of thromboembolism. PMID- 11103542 TI - How do waiting times affect radiation dose fractionation schedules? AB - The purpose of the present paper was to evaluate the changing patterns of dose prescription at the Queensland Radium Institute from 1995 to 1998 inclusive. Data were analysed from the treatment files collected on each patient and these were compared with data on delay time. There has been an increased use of shorter fractionation schedules in the period studied. Paradoxically, radical treatments have become longer. The average number of fractions for all patients was 17.4 and for palliative treatments it was 7.4. The monthly delay varied from 0 to 22 working days and the average was 7 working days. When fraction number was compared to treatment delay, there was a negative linear correlation (R = -0.25). The correlation was stronger (R = -0.467) when palliative treatments were compared, indicating that clinicians were more prepared to alter palliative treatments in the presence of a treatment delay than curative ones. PMID- 11103543 TI - Mantle planning: report of the Australasian Radiation Oncology Lymphoma Group film survey and consensus guidelines. AB - The purpose of the present paper was to measure the variation in mantle planning in Australia and New Zealand. A chest X-ray (CXR) of a patient in the supine position with a neck node marked by wire was sent to every radiation oncologist in Australia and New Zealand. They were to mark on the CXR the lung blocks that they would use to treat this patient, assuming that the patient had stage IA Hodgkin's disease. These marks were compared with a small sample of radiologists who were asked to define the mediastinum on the same CXR. Radiation oncologists were also asked to complete a short questionnaire about other modifications to their treatment fields and their experience with this technique. One hundred and six films were sent out and 44 radiation oncologists replied. There was a maximum variation in the placement of their lung blocks of 6 cm. Half of the lung blocks were within a 2-cm range. One respondent said they would not use a mantle field to treat this patient. Mediastinal coverage was inadequate in at least 50% of cases. There was a very large variation in mantle field planning practices within Australia and New Zealand. For this reason Australasian Radiation Oncology Lymphoma Group has produced consensus guidelines for mantle block design. These are appended to the present paper. PMID- 11103544 TI - Use of digitally reconstructed radiographs in radiotherapy treatment planning and verification. AB - The authors present 3 years of experience in using digitally reconstructed radiographs (DRR) for radiotherapy planning and verification. Comparison is made with simulation film (SF), to illustrate the advantages of DRR over SF. Emphasis is placed on using the appropriate equipment and applying the correct technique. A brief discourse on the principle of CT imaging is presented to illustrate the operation of CT software and optimization of image display for axial slices and DRR. Emphasis is placed on the application of clinical knowledge to enhance the usefulness, as well as the technical quality, of the DRR. Illustrative examples are given. PMID- 11103545 TI - Introduction and implementation of the basic treatment equivalent in a Varian based department. AB - The increasing popularity of 3-D planning leads to procedural alterations as both workload and resource utilization increase. Although the complexity of the techniques has increased (as well as the set-up and treatment times), the workload statistics must still include the number of fields treated. It is commonly known that machine statistics of fields treated per day do not accurately represent workload because there are major differences between techniques. A mantle treatment technique and an opposed spine technique both have (statistically) two fields, although the set-up requirements and treatment times are very different. A basic treatment equivalent (BTE) formula was reported in early 1999 by Delaney et al. and incorporates a large number of variables inherent in patient treatment. The formula considers different factors that affect overall treatment time, and aims to represent a more accurate treatment time indicator. The aim of introducing the BTE into the Department of Radiation Oncology at Sir Charles Gairdner Hospital was to create a more accurate scheduling system and even out workloads on all treatment units. Therefore the BTE formula was used to assess accuracy of treatment times in order to determine if the values could be relied upon as accurate time indicators. Patients undergoing a variety of treatment techniques were timed for the duration of their treatment procedure, and their treatment times compared to the time estimated using the BTE formula. A few minor alterations were made to the equation for treatment units with multi-leaf collimation (MLC). A trial conducted at Sir Charles Gairdner Hospital found that, using the BTE formula (with a few modifications required for the MLC treatment units), of 60 patients timed for the duration of their set-up and treatment, 85% of values were in the range of +/- 3 min, and 95% were in the range of +/- 5 min of the estimated times. Through the routine use of the BTE equation a more sensitive indication of treatment machine workload can be found. Advantages such as: (i) a more accurate measure of treatment workload (for comparison with other departments) and (ii) increased scheduling accuracy will succeed over the currently accepted system of fields per hour. PMID- 11103546 TI - Diastrophic dysplasia with severe primary kyphosis and 'monkey wrench' appearance of the femora. AB - The cases of two sisters with severe diastrophic dysplasia who showed some unusual radiographic features (kyphosis secondary to hypoplasia/dysplasia of the lumbar spine and a 'monkey wrench' appearance of the proximal femur) are reported here. Absent patellae were another feature that has not previously been reported in diastrophic dysplasia. PMID- 11103547 TI - Nuclear medicine imaging in early vertebral osteomyelitis: still of clinical utility. AB - A case is presented of vertebral osteomyelitis in an elderly confused patient with poorly localizing signs. The lesion was not diagnosed on the initial MRI study of the spine due to poor targeting. The abnormality was detected on a bone scan the following day. This was confirmed with a gallium scan 3 days later, and also a repeat MRI study 11 days after the first MRI, using an optimized protocol over the region of interest established by the bone scan. PMID- 11103548 TI - Pigmented villonodular synovitis: magnetic resonance features of an unusual case of bilateral hip joint involvement. AB - Pigmented villonodular synovitis (PVNS) is usually a mono-articular disorder. Bilateral symmetrical affection of this disease process is exceptional. A case is reported of a patient with PVNS with bilateral, symmetrical hip joint involvement. PMID- 11103549 TI - Meningioangiomatosis. AB - The imaging findings of a case of meningioangiomatosis with pathologic correlation are presented. PMID- 11103550 TI - Computed tomography features of small bowel volvulus. AB - A case of small bowel volvulus in an adult is presented. The established CT signs of small bowel volvulus are discussed. These include an abnormal orientation of the superior mesenteric artery to the superior mesenteric vein; the whirl sign, the triangle sign and the beak sign. PMID- 11103551 TI - Hereditary haemorrhagic telangiectasia simulating biliary tree obstruction secondary to malignant porta hepatis lymphadenopathy. AB - Hereditary telangiectasis is an uncommon condition and is thus not usually considered in the differential diagnosis of porta hepatis lymphadenopathy causing biliary tree obstruction. The present case illustrates a patient in whom tortuous vessels in the porta hepatis simulated masses on delayed contrast-enhanced CT with subsequent referral for biopsy. PMID- 11103552 TI - Pancreatic head melanoma producing obstructive jaundice: magnetic resonance characteristics aiding differentiation from adenocarcinoma. AB - A case is reported of malignant melanoma presenting as a mass in the head of the pancreas producing obstructive jaundice. On MR imaging the mass displayed signal characteristics not typical of adenocarcinoma but instead exhibited a low signal on T2-weighted images and early phase dynamic enhancement, the combination of which should suggest the possibility of melanoma. PMID- 11103553 TI - Acute arthritis as an unusual complication of malignancy. AB - A 59-year-old man presented with an inability to weight-bear due to an acute, red, hot, swollen knee joint which was subsequently found to be due to a malignant joint effusion consequent to colonic carcinoma. Treatment with radiation therapy relieved his symptoms. A review of the literature reveals that malignant joint effusion is rare and this is only the third reported case secondary to colonic carcinoma. PMID- 11103554 TI - Malignant retroperitoneal paraganglioma: case report and review of treatment options. AB - A case of retroperitoneal paraganglioma metastasizing to bone is presented. This is followed by a literature review of treatment options, including external beam radiotherapy, chemotherapy and 131I-metaiodobenzylguanidine. PMID- 11103555 TI - Diffuse lymphangiomatosis of bone: comment. PMID- 11103556 TI - Radiotherapy utilization in New South Wales from 1996 to 1998: comment. PMID- 11103557 TI - [Information and support needs of the family of psychiatric patients]. AB - AIM: This study aimed at the systematic assessment of the educational and service needs among relatives of psychiatric patients. METHOD: A postal survey was carried out on a sample drawn from the members of the associations of relatives of mentally ill people in Germany and Austria. Data collected from parents (n = 509) and spouses (n = 54) of patients with schizophrenia or affective disorders were analysed. RESULTS: More than half of the participants expressed the need for more support and complained of not having enough opportunities to relieve the burden imposed on them. Further, families have a strong desire for more information about the disorder as well as practical advice concerning how to cope with the patient. CONCLUSIONS: The results document the great need for services among families and the need for enhancement of quality of care and negotiated partnership between families and professionals. PMID- 11103558 TI - [Need for and availability of services by nursing insurance for patients with rheumatoid arthritis]. AB - Although rheumatoid arthritis is amongst those functional disabling diseases requiring massive help and care, there is as yet no study on how RA patients master their everyday lives, nor are there reports on how many of those patients are in need of and/or do in fact receive benefits from the German Nursing Care Insurance. METHODS: In a representative sample of considerably disabled RA patients (functional capacity < 67%) in rheumatological care it was investigated how many patients received nursing care insurance benefits and how many more would have been entitled to receive them. Standardized interviews exploring functional capacity, amount of help and care needed and help-seeking behaviour were conducted to determine which patient-related and resource-related characteristics were associated with unmet need regarding the patients entitlement to benefits of nursing care insurance. Using the relation between justified need for nursing care insurance benefits and the functional status score, a coefficient was computed by logistic regression to project the expected proportion of RA patients entitled to care insurance benefits. RESULTS: A projected 5.4% of all RA patients needed benefits from the nursing care insurance, but only 63% of those did in fact receive benefits. It was found that unmet need was almost exclusively due to the fact that patients actually eligible for insurance benefits did not apply for it, whereas unjustified rejection of applications by insurance experts made up for only a marginal proportion. CONCLUSION: Applying for nursing care insurance benefits is still not a matter of course. It must be considered that about one third of all obviously care dependent RA patients either claim care insurance benefits too late or never do. To avoid unmet need, experts should encourage particularly those patients who are unaware of their right to ask for help. PMID- 11103559 TI - [How many intensive care beds are necessary? A quantitative study with the Therapeutic Intervention Scoring System (TISS) in 5 Hessian hospitals)]. AB - In five major Hessian hospitals all patients of critical care units have been grouped according to the Therapeutic Intervention Scoring System (TISS) over a time span of four weeks. The objective was to establish the critical care patient capacity needed in accordance with the Hessian Guidelines for Critical Care. Sample surveys showed that the ascertained data are highly reliable. 10,756 TISS classifications have been evaluated in total. 9.4% of the classifications have been assigned to general ward, 27.5% to intensive monitoring and 63.1% to intensive treatment. Assuming a standard rate of use of 85% over the year this revealed an average requirement of critical care patient capacity of 6.1% of the total number of beds. The results of the investigation were readily accepted by health insurances and hospitals involved. PMID- 11103560 TI - [Psychosomatic health promotion in adolescents. An intervention study in 2 high schools]. AB - AIMS: The effects of health-promotion lesson modules on teaching climate, school stress and physical and psychological well-being were investigated in a controlled intervention study in two high schools. The lesson plans were developed jointly by teachers and students and integrated into normal classroom teaching. METHODS: Three questionnaire assessments were carried out with students in the intervention and control schools. Health promotion lessons were implemented in the intervention school between T1 and T2 (an interval of 12 months) and evaluated by participants after completion. RESULTS: Indicated that the modules were positively appraised and that participation in a longitudinal study led to a more critical evaluation of the teaching climate and school stress. A decrease in physical and psychological symptomatology ran in parallel with increases in critical appraisal of the school environment. This pattern of results was found in both schools, i.e., no intervention effect could be demonstrated. CONCLUSIONS: It is likely that participation made the adolescents more sensitized to individual and interactional problems as well as led them to mobilize their own resources. The health-promotion activities supported a participatory/interactive relationship between teachers and students which in turn resulted in systemic structural changes. It can be assumed that these factors will have long-term positive effects on the well-being of the participants. PMID- 11103561 TI - [Goppingen 1999 vaccination program in the 5th and 6th grade]. AB - During a campaign carried out by the Ministry for Social Affairs in Baden Wuerttemberg to promote vaccination against measles, mumps and rubella (German measles) (MMR), the Juvenile Medical Service at the Public Health Office in Goeppingen carried out a broadly offered vaccination (advisory) programme in the Goeppingen area for 5,800 schoolchildren in their first and second years of secondary education (10-12-year olds) from September to December 1999. Our first aim was to detect in this target group vaccination gaps which could be closed by practising physicians, but our medical service also offered the mentioned vaccinations in those cases where no family doctor had been requested to vaccinate. Furthermore, the vaccination rates of standard immunizations were registered and a regional health report was published. Vaccination gaps were found especially at schools for children requiring special care and attention and schools for the less academically inclined, and showed an increased low-threshold need for immunization offers. PMID- 11103562 TI - [Electronic data processing assisted quality management in general practice networks]. AB - Possibilities of an EDP-based quality management for medical networks are presented and some organizational requirements are derived. To show the relevance of the approach, three networks are sketched. PMID- 11103563 TI - [Drug advertising--users want information. Report of telephone survey conducted by North-Rhine Westphalia Public Health Service on the topic of drug advertising and drug information for users]. AB - In Germany, drug advertising of non-prescription drugs is a controversial subject. On the one hand, consumer organisations plead for placing a ban on advertising or at least to offer a detailed description of medical risks in respect of protection. On the other hand, the pharmaceutical industry is keen on liberalizing the specific advertising law for drugs. A representative telephone survey among the population of North Rhine-Westphalia was conducted in April and May 1999. It showed consumer interest in advertising, the value of information on risks, the institution with maximum credibility in drug information for consumers, the importance of the now obligatory sentence after every advertisement: Regarding risks and side effects read the leaflet in the package and ask your physician or pharmacist, and to what extend the consumer would take advice from independent experts over the telephone about drugs. It was found that, in particular women, about 30% are occasionally interested in advertising, younger people are more open-minded about advertising than older people; and that doctors and pharmacists have the most credibility and are consulted for further information. It was also found that more than 80% of the population demanded precise information on the side effects of drugs. One-third of the consumers declared that the obligatory sentence (see above) led to greater demand for information from doctors or to read attentively the instruction leaflet. Nevertheless, there is a need for more information from more than half of the consumers, who would take advantage of an independent advice centre if this should exist. PMID- 11103564 TI - [Routine documentation based on disability data of the legal health insurance]. AB - Data on sickness absence of employees are routinely used for health reporting and the analysis of work-related morbidity by institutions of the German health insurance. Since the insurance system comprises several different branches, these health reports differ in respect of objectives, data selection, methods of analysis, and presentation of results. A further lack of comparability is caused by the heterogeneous populations, since membership in a certain health insurance depends on social status and job requirements. Aim of this paper was to review the methods and characteristics of health reports as they are routinely published in Germany. By evaluating these reports recommendations should be derived to improve comparability. The review showed that at least four different kinds of health reports should be differentiated: company-based health reports, health reports for business branches, morbidity statistics, and reports from research projects. Such reports have different objectives and therefore require different methods of data analysis. However, group-specific and common standards could be set up and it is suggested that health insurance institutions should work out report guidelines based on the recommendations given in this review. PMID- 11103565 TI - ["Deficiencies in the mandatory health insurance system"--possibilities and limits of medical expert assessment]. AB - In Germany, the Federal Committee of Physicians and Health Insurance Agencies is responsible for assessing which medical procedures are covered by the insurance agencies (therapeutic/economic value). The failure to evaluate certain medical procedures which promise therapeutic benefit may be considered a failure in the system of statutory health insurance as decreed by the German Federal Social Court. In cases of system failure the assessment of medical procedures is taken over by social courts. Evidence of therapeutic value will continue to be the decisive criterion of evaluation. In special cases, however, it may be replaced by the particular procedure's rate of incidence in everyday medical practice, as reflected by a widespread resonance in medical discussion and its use by a considerable number of physicians. Medical experts may face various difficulties in handling these alternative criteria. They usually lack reliable data on the frequency of the procedure, i.e. its distribution. Even if such data and data on secondary factors--such as disease incidence etc.--were available to them, they would still not be able to come up with a definite conclusion on the degree (widespread, considerable) of dissemination, as defined by the German Federal Social Court. Nevertheless, on request of statutory health insurances, social medical experts may investigate facts in order to establish basic knowledge for later decisions that are to be done by others. Factual investigation includes clarification of etiology, incidence, importance and natural history of the particular disease processes. Potential deficiencies in the medical services rendered for specific diseases as well as the characteristics of the procedure under scrutiny, and special requirements for evaluating the outcome have to be investigated as well. Additional attention needs to be paid to the quality of related publications (analysis of statistical data), possible recommendations for therapy (e.g. guidelines and their scientific basis), as well as to economic aspects. The result of such an investigation has to be stated without use of court criteria (widespread/considerable). PMID- 11103566 TI - [Quality assurance in treatment of stroke: basic module of the German Stroke Registry Study Group]. AB - BACKGROUND AND PURPOSE: Comparable, standardised data on the quality and efficiency of stroke care in Germany are lacking. The Arbeitsgemeinschaft Deutscher Schlaganfall-Register (ADSR--German Stroke Registries Study Group) has defined a "Minimum DataSet" for the evaluation of quality indicators of stroke treatment in Germany. METHODS: The ADSR is a voluntary network of current regional stroke registries aiming at a standardisation in the use of stroke terminology and methods of data collection for German stroke databases. Currently six regional stroke registries are cooperating in the ADSR, combining data from about 18,000 stroke patients in 110 hospitals annually. RESULTS: In the design of the ADSR DataSet a modular approach was chosen. The ADSR "Minimum DataSet" was adapted for a wide use in different health care facilities. In addition to the "Minimum DataSet" an "Advanced DataSet" will be developed to document additional items of stroke care in specialised stroke centres. The ADSR DataSet collection will be completed by special "Extended DataSets", designed for answering centre specific and research questions. CONCLUSION: The ADSR "Minimum DataSet" allows a standardised assessment of stroke care in Germany. It is the first questionnaire that provides valid and reliable comparisons between different clinical settings as well as regional stroke databases. The ADSR "Minimum DataSet" defines core items for a future National German Health Report on stroke care. PMID- 11103567 TI - [German League of the Bone and Joint Decade 2000-2010--for prevention and therapy of diseases and injuries of the skeletal and locomotor system]. PMID- 11103568 TI - Leveling the playing field. PMID- 11103569 TI - Tooth sensitivity. PMID- 11103570 TI - Class II composite restorations. PMID- 11103571 TI - Sleep disordered breathing. PMID- 11103572 TI - Clinical research: learning right from wrong. PMID- 11103573 TI - Occlusal vs. non-occlusal etiology. PMID- 11103574 TI - Nanodentistry. AB - BACKGROUND: Nanodentistry will make possible the maintenance of comprehensive oral health by involving the use of nanomaterials, biotechnology (including tissue engineering) and, ultimately, dental nanorobotics (nanomedicine). RESULTS: When the first micrometer-sized dental nanorobots can be constructed within 10 to 20 years, these devices will allow precisely controlled oral analgesia, dentition replacement therapy using biologically autologous whole replacement teeth manufactured during a single office visit, and rapid nanometer-scale precision restorative dentistry. CLINICAL IMPLICATIONS: New treatment opportunities may include dentition renaturalization, permanent hypersensitivity cure, complete orthodontic realignments during a single office visit, covalently bonded diamondized enamel and continuous oral health maintenance through the use of mechanical dentifrobots. PMID- 11103575 TI - Nanodentistry. Fact or fiction? PMID- 11103576 TI - Dental caries prevalence and dental care utilization among the very old. AB - BACKGROUND: This article reports on coronal and root caries prevalence and dental care utilization patterns of elderly Iowans aged 79 years or older. METHODS: The sample for this study was 449 people who were surviving members of the Iowa 65+ Rural Health Study cohort originally recruited in 1981. The authors focused their analyses on the 342 of these who were dentate. Examinations were conducted in subjects' homes by trained and calibrated examiners, using a halogen headlight, a mouth mirror, a color-coded periodontal probe and a no. 23 explorer. RESULTS: The mean age of subjects was 85.1 years (range 79-101 years), and they had a mean of 19.4 remaining teeth. Nearly all subjects (96 percent) had coronal decay experience, while 23 percent of the subjects had untreated coronal decay, about one-fourth of which was recurrent. Nearly two-thirds (64 percent) of the sample had root caries experience, with 23 percent having untreated root caries. Utilization of dental services was high among the dentate elderly, with nearly three-quarters reporting having visited a dentist within the past year. Nearly all reported that they paid for dental care themselves with no third-party coverage. CONCLUSIONS: The findings from this study of the very old suggest that coronal and root caries remain prevalent, with high levels of dental care utilization among those who have retained natural teeth. CLINICAL IMPLICATIONS: As the U.S. population ages, and more teeth are retained, demand for dental services in the population of the oldest elderly people is likely to increase. PMID- 11103577 TI - Nonsurgical periodontal therapy in 2000: a literature review. AB - BACKGROUND: This article addresses the advantages and limitations of nonsurgical periodontal therapies to treat patients with mild-to-moderate chronic periodontitis. TYPES OF STUDIES REVIEWED: Controlled clinical trials were selected that assessed the efficacy of the following treatment methods: mechanical instrumentation, ultrasonic debridement, supragingival irrigation, subgingival irrigation, local drug delivery, administration of systemic antibiotics and host-response modulation. Evidently, data with regard to alterations of probing depth, clinical attachment levels and inflammatory status were evaluated. RESULTS: Comparison of the data from test and control groups revealed the following results. Manual and ultrasonic debridement can be used to treat most patients with mild-to-moderate chronic periodontitis. Patients who do not practice optimal plaque control can enhance their personal hygiene procedures by using supragingival irrigation. Subgingival irrigation usually does not provide any benefit beyond that achieved with root planing. Systemic and locally delivered antimicrobial agents appear to be most beneficial among patients who do not respond to conventional treatment. Host modulation may enhance root planing modestly. CLINICAL IMPLICATIONS: The data indicate that most patients with mild to-moderate periodontitis can be treated with nonsurgical therapies. However, clinicians need to be aware of the limitations of each technique with regard to the magnitude of improvement that it can induce at specific sites. PMID- 11103578 TI - Unrecognized carotid artery stenosis discovered by calcifications on a panoramic radiograph. AB - BACKGROUND: Approximately 730,000 strokes occur each year in the United States, costing an estimated $40 billion annually. One-half of all strokes are the result of atherosclerotic plaques found in the carotid artery. Such plaques frequently are heavily calcified and can be identified on a panoramic radiograph by the incidental finding of calcifications overlying the carotid bifurcation. CASE DESCRIPTION: The authors found that a 67-year-old asymptomatic woman had calcium deposits overlying both carotid bifurcation regions on a panoramic radiograph. Subsequent duplex ultrasonic examination indicated bilateral, high-grade carotid arterial stenoses. The patient underwent uneventful bilateral carotid endarterectomy. CLINICAL IMPLICATIONS: The patient had critical carotid arterial stenoses associated with significant risk of stroke that had not been identified otherwise. The findings on the panoramic radiograph led to appropriate and potentially life-saving treatment. While the positive predictive value of this finding has yet to be defined, the authors believe that calcifications overlying the carotid system region seen on panoramic radiography in an asymptomatic patient should be followed by formal evaluation of the carotid bifurcation. PMID- 11103579 TI - Pink teeth resulting from Russian endodontic therapy. PMID- 11103580 TI - A survey of antibiotic use in dentistry. AB - BACKGROUND: Antibiotics are important in the management and prophylaxis of infection in patients at risk of experiencing microbial disease. As a result of the increase in antimicrobial resistance, the authors conducted a survey to assess current antibiotic use in dental practice. METHODS: The authors mailed a two-page, pretested survey to all licensed dental practitioners in British Columbia, Canada. A total of 2,542 surveys were mailed; 19.9 percent were returned by fax or mail. The authors examined an association between factors analyzed using a chi 2 test. RESULTS: Respondents were demographically consistent with all registered dentists in British Columbia. They reported writing an average of 4.45 prescriptions per week. Antibiotics prescribed after treatment primarily were penicillin and its derivatives. Recommended adult doses of penicillin were prescribed by 59.2 percent of respondents; recommended daily doses of amoxicillin were prescribed by 72.2 percent of respondents. The average prescription duration was 6.92 days. Respondents prescribed prophylactic antibiotics an average of 1.15 times per week for prophylaxis of bacterial endocarditis; 17.5 percent reported postoperative dosing for prophylaxis, ranging from a one- to seven-day prescription with an average of 6.91 postoperative doses. Preoperative antibiotics were prescribed for patients with a history of rheumatic fever or any heart murmur or prosthetic hip. Antibiotics were prescribed more frequently for surgical procedures and patients with acquired immunodeficiency syndrome than for other circumstances. CONCLUSIONS: More than 80 percent of respondents reported that they followed current American Heart Association prophylaxis guidelines. The authors, however, noted discrepancies in prophylactic use of antibiotics for bacterial endocarditis and for patients with large joint prostheses, as well as in prescribing antibiotics in the presence of clinical infection. In therapeutic use, approximately 85 percent of respondents followed appropriate prescription guidelines for dosing and duration of therapy. CLINICAL IMPLICATIONS: Appropriate and correct use of antibiotics is essential to ensure that effective and safe treatment is available and that practices that may enhance microbial resistance are avoided. To improve standards of care, dentists need up-to-date pharmacology in dental education, as well as continuing education, further outcome studies and continuous assessment of dental practices. PMID- 11103582 TI - Characteristics of dentists participating in capitation and Preferred Provider Organization dental plans. AB - BACKGROUND: In 1998, the American Dental Association Survey Center conducted a telephone and mail survey of U.S. dentists in private practice in an effort to determine the extent of dentists' participation in capitation and preferred provider organization, or PPO, dental plans and the characteristics of dentists who participate in those plans. METHODS: An initial telephone screening survey of a random sample of 11,550 dentists in private practice was conducted to identify dentists who participated in PPO or capitation dental plans. Dentists who participated in either of these plan types then were asked to complete a mail survey on their plan participation. RESULTS: The majority of dentists participating in either type of dental plan reported having never left a dental plan. Dentists who belonged to more than one PPO or capitation plan reported that a larger percentage of their patients were enrolled in these plans and that more of their practice's gross income came from the plans. Participation in PPO and capitation plans has had a positive impact on the practices of many of the responding dentists, particularly with regard to expanding their patient base. CONCLUSIONS: The authors found that the majority of dentists participating in PPO dental plans found it to be a positive experience overall. Dentists participating in capitation plans were less satisfied; more than 50 percent of capitation plan participants reported some level of dissatisfaction with the plans. The majority of dentists participating in a PPO plan expected to renew participation when their current contract expired; a much smaller percentage (though still a majority) of responding capitation-plan participants indicated the same. PRACTICE IMPLICATIONS: Responding dentists' overall indication of satisfaction with their current PPO plan participation probably indicates further growth for these dental plans. On the other hand, capitation plan participants seem much less satisfied with their plans. PPO plans, therefore, seem much more likely to be the type of plan that dentists will choose in the future. PMID- 11103581 TI - The physical properties of packable and conventional posterior resin-based composites: a comparison. AB - BACKGROUND: The authors compared the physical properties of three packable hybrid resin-based composites with those of a conventional hybrid and a microfill composite material advocated for use as posterior restorative materials. They evaluated diametral tensile strength, or DTS; compressive strength, or CS; flexural strength, or FS; and depth of cure, or DC. METHODS: The authors studied the following resin-based restorative materials: three packable composites, Alert Condensable Composite (Jeneric Pentron), SureFil High Density Posterior Restorative (Dentsply Caulk) and Solitaire (Heraeus Kulzer); one conventional hybrid composite, TPH Spectrum (Dentsply Caulk); and one microfill, Heliomolar Radiopaque (Ivoclar-Vivadent). The authors evaluated DTS, CS, FS and DC, according to American National Standards Institute criteria. They made scanning electron micrographs of the packable resin-based composites. RESULTS: Results demonstrated that the conventional hybrid, TPH Spectrum, had significantly greater DTS and FS than other resin-based composites. Alert and SureFil had comparable DTS and FS, which were significantly greater than Heliomolar's DTS and FS. Solitaire had significantly lower DTS and FS than all other resin-based composites. SureFil had the highest CS, followed by TPH Spectrum, Solitaire and Alert, which were comparable and had significantly greater CS than Heliomolar. TPH Spectrum and Alert had significantly greater DC than all other resin-based composites, followed in decreasing order by SureFil, Solitaire and Heliomolar. CONCLUSION: While the packable composites tested in this study had physical properties superior to those of the microfill composite, they were no better suited for use as a posterior restorative material than was the conventional hybrid resin-based composite. CLINICAL IMPLICATIONS: Packable composites may be easier for clinicians to handle than conventional resin-based composites; however, their physical properties were not superior to those of the conventional small-particle hybrid resin-based composite. In addition, these materials may have the clinical drawback of increased wear and surface roughness that was seen with early, large-particle composite restorative materials. PMID- 11103583 TI - When and why should I have my practice appraised? PMID- 11103584 TI - Improving the quality of fixed prosthodontic services. AB - I have discussed only the three problems most frequently mentioned by laboratory technicians in the Lab Management Today survey. In my opinion, there are many more areas in fixed prosthodontics that need attention. To achieve this, we must improve the quality of the procedures we use in creating fixed prostheses. Laboratory technicians observe the quality of impressions, tooth preparations and interocclusal records that are sent to them by the dentists with whom they work. Their statements about the need for improvement on several of the aspects of the clinical procedure are of concern. It is dentists' professional responsibility to upgrade their clinical techniques so that they produce high-quality impressions of adequate tooth preparations and transfer information about the proper interocclusal relationship to the dental technician. PMID- 11103585 TI - Vindication in court. A review of the 9th Circuit's ruling in California Dental Association vs. FTC. PMID- 11103587 TI - [Trastuzumab. Monoclonal antibody for the treatment of breast cancer]. PMID- 11103586 TI - [Beta-lactams--complete or able to be improved?]. PMID- 11103588 TI - [Depressions. Picture of the disease and current therapy]. PMID- 11103589 TI - [Importance and various risks of anesthesia procedures]. PMID- 11103590 TI - [Use of ear drops]. PMID- 11103591 TI - [An antibiotic for tartar removal?]. PMID- 11103593 TI - Relationship between neonatal birthweight variation in respect to 50 degrees percentile and maternal education level in a high selected and homogeneous population. AB - BACKGROUND: To evaluate the influence of maternal education on exceeding and missing weight of newborns in respect to 50th percentile. METHODS: DESIGN: observational study. Length: five years. SETTING: obstetric and Gynecology Division Care. Neonatology Division Care. SUBJECTS AND MEASUREMENTS: 66 male and 57 female newborns having birth weight of around the 50th percentile; missing or exceeding weight was evaluated in respect of the 50th percentile of the relative week of birth. Parents were selected on the basis of the following criteria: mother: age 30-35, BMI before pregnancy 20-25, no habit for smoking or alcohol, first pregnancy, urban living area, no hypertension, normal glucose tolerance, normal thyroid function, regular pregnancy course, Caucasian race, Italian nationality, regularly married; father: BMI 20-25, no habit for smoking or alcohol, urban living area, steady job, just higher, equal or lower education level than wife's. Mothers underwent OGTT, measurement of thyroid function and recording of blood pressure and educational level. RESULTS: No relationship was observed between mother's education and exceeding weight both in male and in female newborns, while negative relationship was found between mother's education and missing weight both in male and in female in respect of the 50th percentile (Linear regression). CONCLUSIONS: Our data show that low maternal education exerts an influence in reducing but not increasing birth weight. PMID- 11103592 TI - Effectiveness of forced rehydration and early re-feeding in the treatment of acute diarrhoea in a tropical area. AB - BACKGROUND: The administration of oral re-hydration solution (ORS) via continuous infusion through a nasogastric (NG) tube and early refeeding facilitates delivery in hospitalised children and the return back home. METHODS: DESIGN: the observation was made during a one-year stage in the Camillian Medical Centre (CMC) of Ouagadougou in Burkina Faso. 4,131 infants and children under 5 years old, affected by acute diarrhoea and severe dehydration (loss of weight > 10%) were studied. Those children having difficulties for oral re-hydration were hydrated by continuous infusion through naso-gastric (NG) tube; the NG tube was put in by the nurses and connected to a 500 ml bottle, in which a mixture of glucose and electrolytes was dissolved according to the formula (glucose 20 g + NaCl 3.5 g + NaHCO3 2.5 g KCl 1.5 g in 1 litre of water). The infusion rate was 20-30 drops/minute. No sedative or anti-emetic drug was given, unless in the presence of uncontrolled vomiting. At the end of infusion, flour of millet (60%), soy bean (20%), peanut butter (10%), sugar (10%) and salt (1%) was administered and continued at home or in the nearby areas available for the night. RESULTS: After 4-5 hrs of infusion 3,717 children (90%) showed a significant gain of weight, although the weight prior to the acute event preceding hospitalisation was never reached during their stay at the CMC. Only 413 children (10%) required a longer period of forced infusion: at the end of the day, however, they were fed with this flour. CONCLUSIONS: A simple strategy, based on a NG infusion plus an oral administration of flour has proven safe and effective in encompassing those difficulties encountered in the treatment and prevention of dehydration in developing countries where the therapy, in children affected by diarrhoea, still represents a major daily occupation. PMID- 11103594 TI - Childhood glomerulonephritis associated with varicella and streptococcal infection. AB - Varicella is a usually benign disease of childhood and its complications are uncommon in immunocompetent children. In recent years we have witnessed the increasing virulence of group A beta-haemolytic streptococci (GABHS). In particular, in 1993, 50% of new cases of invasive GABHS disease were associated with varicella infection and all were suppurative complications. Because also a non suppurative complication of varicella as glomerulonephritis associated with GABHS infection, has been published in only one case, we feel that it could be of interest to describe this condition in two other cases we have observed. PMID- 11103595 TI - Ectrodactyly. A case report. AB - A case of ectrodactyly characterized by simple absence of the third finger of the right hand is reported. Clinical and genetic aspects are considered. PMID- 11103597 TI - Raymond V. Gilmartin. PMID- 11103596 TI - [Acute infectious diseases]. PMID- 11103598 TI - Merck: committed to access to medicines for patients throughout the world. PMID- 11103599 TI - Patients not profiting from for-profit care. PMID- 11103600 TI - World AIDS Day: men make a difference. PMID- 11103601 TI - Pride of the nation tomorrow. Where UMDNJ is headed. PMID- 11103602 TI - Medicine and mediation: healing the conflicts that divide us. PMID- 11103603 TI - Influenza: prevention and treatment. PMID- 11103604 TI - Osteoporosis. Recommended guidelines and New Jersey legislation. AB - More than 28 million Americans aged 50 and over have osteoporosis or low bone density. While 80% of those affected are women, one in eight men also suffer from the disease. This number is expected to increase as men live longer. Commonly called the "silent disease," osteoporosis exhibits no symptoms or pain until a fracture occurs. The most common fractures involve the wrist, vertebra, and hip. Half of all women and 20% of men will have at least one osteoporosis fracture in their lifetime. This article describes statewide efforts to prevent and manage osteoporosis and the recommended practice guidelines for the diagnosis and treatment of the disease. PMID- 11103605 TI - Lyme disease. Pitfalls in diagnosis. PMID- 11103606 TI - How bone density testing influenced osteoporosis treatment in a community hospital. PMID- 11103607 TI - Recent changes in vaccination guidelines. An update on vaccines and target population initiatives. PMID- 11103608 TI - Preventing childhood poisoning. An intervention in a family medicine residency program. PMID- 11103610 TI - Public health in North Carolina. The new deal comes to Raleigh. PMID- 11103609 TI - A woman with abdominal pain and bilious vomiting. A very late aftermath of Billroth II gastrectomy. AB - Patients with a history of Billroth II gastrojejunostomy who present with a symptom complex of postprandial nausea, fullness, and bilious vomiting leading to relief should be suspected of having an afferent loop syndrome. Diagnosis depends on barium radiography and upper intestinal endoscopy. Surgical correction is the treatment. The current age of medical therapy has dramatically decreased the frequency and necessity of surgery for peptic ulcer disease. However, we should not forget the lessons of the past and fail to diagnose a patient who has a chronic complication of a previously common operation. PMID- 11103612 TI - Caroliance. North Carolina's health insurance cooperative for small businesses needs a doctor. PMID- 11103611 TI - The beginning of emergency medicine. A personal view from North Carolina. PMID- 11103613 TI - [Use of bone markers in veterinary medicine]. AB - Bone metabolism can be monitored in humans and several animal species in vivo by measuring enzymes and other protein products released by osteoblasts and osteoclasts, respectively. The biochemical markers of bone formation currently in use include bone isoenzyme of alkaline phosphatase, osteocalcin and propeptides derived from the N or C terminal ends of the typ I procollagen molecule. The most useful markers of bone resorption are breakdown products of type I collagen. The oldest established method is the measurement of hydroxyproline in urine, which is not specific for bone, because it can be found in all collagen types and is also derived from diets. The measurement of collagen crosslinks, deoxypyridinoline and pyridinoline, is comparatively more specific to monitor bone resorption. Deoxypyridinoline and pyridinoline are used in human medicine for diagnosis and evaluation of bone diseases and in predicting the occurrence of fractures and rates of bone loss. The carboxyterminal telopeptide of type I collagen, which has been used in several animal species, is also a promising bone marker. This article reviews the use of different bone markers in veterinary medicine and the possibilities for diagnosing and preventing bone diseases. PMID- 11103614 TI - [Main virulence factors in Escherichia coli isolates from swine over two weeks old with edema disease and/or E. coli diarrhea]. AB - The OK antigens and the fimbriae F4 of E. coli with haemolysis isolated from 113 cases of oedema disease and/or diarrhoea were identified serologically. The genes for F18 and for enterotoxins LT, STIa and STII as well as Shigatoxin Stx2e were determined by PCR. Fimbrial variants F18ab and F18ac were distinguished by means of indirect immunofluorescence on smears prepared from the intestinal mucosa and from cultures grown under appropriate conditions. Adhesive fimbriae were detected with every case or isolate, respectively, by means of at least one out of the techniques mentioned above. The serogroup O149:K91 with fimbriae F4ac (K88ac) and genes for the enterotoxins LT and STII was most prevalent. Serogroup O139:K12 with fimbriae F18ab and the gene for Stx2e was second, whereas serogroups O141ab and O141ac with fimbriae F18ac and genes for Stx2e, STII and often LT were much less prevalent. The serogroup O147:K89 with fimbriae F18ac, and genes for STIa and STII was detected for the first time in Switzerland. PMID- 11103615 TI - [Radiation therapy in two cats with pituitary tumors]. AB - Two cats with large pituitary neoplasms (adenoma and adenocarcinoma) were treated with fractionated radiation therapy. Total doses of 40 Gy, respectively 36 Gy, were applied in 10 fractions of 4 Gy, and 3.6 Gy respectively. Side effects were minimal and transient. Anesthesia was well tolerated. Improvement of clinical signs could be observed during radiation therapy in both cats. One cat had a complete, the other a partial tumor response. One cat (suspicion of adenoma) was euthanized 1 3/4 years after therapy due to unrelated disease. No tumor was found on histopathology, however a small focal necrosis of brain tissue in the irradiated field was observed. The second animal with a pituitary adenocarcinoma was euthanized because of tumor recurrence 1 1/2 years after therapy. Radiation therapy was effective, despite the low total doses of radiation applied. PMID- 11103616 TI - [Demodicosis in a Toggenburg goat]. AB - This report describes the findings in a four-year-old Toggenburg goat with demodicosis. The skin had multifocal nodules, which were approximately 5 mm in diameter and contained thick yellow exsudate. Microscopic examination of the exsudate revealed numerous Demodex caprae. The goat was clipped and treated topically every five to seven days for a total of 12 treatments with a 1:100 dilution of amitraz (Ectodex, Hoechst Roussel Vet). The treatment resulted in a marked decrease in the number of skin nodules. However, new nodules appeared after treatment was discontinued and complete clinical cure was not achieved. PMID- 11103617 TI - [Cell transplantation in surgery--reality and prospects for tissue engineering]. AB - Traditionally surgical repair of tissue defects and loss or failure of function has relied on mechanical means, medical (drug) treatment, autologous and allogenic transplantation, and alloplastic/synthetic devices. Tissue engineering represents a new interdisciplinary field of applied research combining engineering and biosciences together with clinical application (mainly in surgical specialities) to develop living substitutes for tissues and organs. The understanding of cell-cell interactions and chemical signalling (growth factors) and the selection of appropriate matrices (cell-matrix interaction) is the key for success. Gene therapy represents the logical combination with tissue engineering on the molecular biology level. Application of cultivated skin and cartilage has already become reality, engineering of vascularized, more complex organs remains a challenge for this century. PMID- 11103618 TI - [Familial Mediterranean fever--from gene test to clinical aspects]. AB - Familial Mediterranean Fever (FMF) is a genetically defined disease affecting mostly families of jewish, turkish or armenian origin whose ancestors originate from the mediterranean basin. The first officially acknowledged description was given by SIEGAL in 1945 but previous cases were reported since 1908. The main clinical signs which are very varying in intensity and appearance are periodic attacks of fever with peritonitis, pleurisy and arthritis. The classical but not always found complication is amyloidosis with renal failure which is preventable by lifelong colchicine therapy. By using a novel genetest it is now possible to definitely diagnose FMF instead of relying on a diagnosis made merely by exclusion. This will emphasize the use of colchicine and should bring us nearer to the pathophysiology of this interesting disease. PMID- 11103619 TI - [DNA vaccination]. AB - DNA vaccination is based on the observation that plasmid DNA could directly transfect muscle cells, dendritic cells, and other cell types in vivo. Intramuscular, intraepidermal, or oral application of expression plasmids can induce humoral and cellular immune responses against the encoded proteins. This strategy has now been used to elicit protective antibody and cellular immune responses in a wide variety of preclinical animal models for viral, bacterial, parasitic, and malignant diseases. DNA vaccination is particularly useful for the induction of cytotoxic T cells and may, therefore, have therapeutic potential as well. This was recently demonstrated in an animal model of tuberculosis. Early phase human trials are in progress. The results from these trials may be eagerly awaited. PMID- 11103620 TI - [Cell transplantation in hepatology]. AB - Preclinical studies have shown that hepatocytes transplanted into the liver function normally on a long term basis. The procedure is well tolerated and offers a wide range of potential clinical applications. Clinical case studies have been performed in the setting of acute and chronic liver failure as well as metabolic diseases. In particular, the case of a patient with Crigler-Najjar Syndrome Type I clearly demonstrated the long term viability of transplanted hepatocytes with stable metabolic function. Further studies are warranted to assess the full therapeutic potential of hepatocyte transplantation. PMID- 11103621 TI - [Non-alcoholic fatty liver]. PMID- 11103622 TI - [Herpes zoster]. PMID- 11103623 TI - Trust and the healing encounter: an examination of an unorthodox healing performance. AB - Just why a patient should trust a particular healer is a question that has not been adequately explored in the literature on healing. This ethnographic case report examines the healing performance of a chiropractor and proposes that it contains four intrinsic claims to trustworthiness: he claims to be a qualified and sincere healer who is in possession of knowledge and techniques that derive their power from their truth content and which empower him to make beneficial changes in the patient. Taking each claim in turn I described the nature of the claim, how it might be adequately validated, ways in which his healing performance might validate it and how he might be assisted by the patient, and how their actual validation may be distorted by the healer and patient. It is suggested that while unusual in many regards, this unorthodox healing performance may be a foil by which to examine other more orthodox healing performances. PMID- 11103624 TI - A pathological view of disease. AB - This paper is a response to Christopher Boorse's recent defense of his Biostatistical Theory (BST) of health and disease. Boorse maintains that his concept of theoretical health and disease reflects the "considered usage of pathologists." I argue that pathologists do not use "disease" in the purely theoretical way that is required by the BST. Pathology does not draw a sharp distinction between theoretical and practical aspects of medicine. Pathology does not even need a theoretical concept of disease. Its focus is not theoretical, but practical; pathology's goal is to contribute to the healing of patients. Pathology, even experimental pathology, is not value-free. Not only "disease" but also such terms as "nerve" and "organ" are laden with conceptual values. PMID- 11103625 TI - Priest attended death in medieval and early modern period: translation and commentary on an old Croatian text circa 1600. PMID- 11103626 TI - Health of organisms and health of persons: an embedded instrumentalist approach. AB - In a time when we as a society are in the process of deciding what our basic rights to health care are, it is critically important for us to have a full and complete understanding of what constitutes health. We argue for an analysis of health according to which certain states are healthy not in themselves but because they allow an individual to reach actual goals. Recognizing that the goals of an individual considered from the point of view of biology and the goals of the same individual considered as an agent in the world might be different, we introduce a distinction between the health of an individual qua organism and the health of an individual qua person. We then argue that this distinction characterizes the evaluations made by patients and healthcare providers better than the widely discussed distinction between disease and illness. PMID- 11103627 TI - Losing perspective with Nurse Betty. The real causes of the nursing shortage. PMID- 11103628 TI - Do you smoke? If so, stop. Please, stop. PMID- 11103629 TI - Nursing organizations unite. Opposition to the AMA's petition is strong. PMID- 11103630 TI - A spoonful of medicine. Or is it more? PMID- 11103631 TI - Lurking nurses. The perils of post-op. PMID- 11103632 TI - A hidden menace: hemolytic uremic syndrome. PMID- 11103633 TI - Sickle cell pain & hydroxyurea. PMID- 11103634 TI - Rheumatoid arthritis. PMID- 11103635 TI - Mightier than the syringe. PMID- 11103636 TI - Elder abuse and neglect. It takes many forms--if you're not looking, you may miss it. PMID- 11103637 TI - Oxandrolone restores appetite. An increase in weight helps heal wounds. PMID- 11103638 TI - Proving nursing negligence. PMID- 11103639 TI - Kegel exercises. Strengthening the weak pelvic floor muscles that cause urinary incontinence. PMID- 11103640 TI - State regulation of RN-to-patient ratios. PMID- 11103641 TI - Pocket full of miracles. A professional portfolio can be a powerful force in your career advancement. PMID- 11103642 TI - Violence in the health care workplace. PMID- 11103643 TI - The world wide nursing Web. PMID- 11103644 TI - When domestic violence leaves home. It can and does invade the workplace. PMID- 11103645 TI - Writing basics: elements of the case study. PMID- 11103646 TI - Status report on nutrition in the news. PMID- 11103647 TI - A view on high-protein, low-carb diets. PMID- 11103648 TI - Lactose intolerance: a new perspective. PMID- 11103649 TI - What should we do about excess weight? PMID- 11103650 TI - Biotechnology: mobilizing dietitians to be a resource. PMID- 11103651 TI - Solving global nutrition challenges requires more than new biotechnologies. PMID- 11103652 TI - Food biotechnology in the new millennium: promises, realities, and challenges. PMID- 11103653 TI - Impact of declines in nutritional status on outcomes in adult patients hospitalized for more than 7 days. AB - OBJECTIVE: To assess the association between changes in nutritional status in hospitalized patients and the occurrence of infections, complications, length of stay in hospital, and hospital charges. DESIGN: A prospective observational study with a retrospective component was conducted over a 7-month interval at a university hospital. SUBJECTS: A total of 404 adults (> or = 18 years old) admitted to the inpatient service for more than 7 days who were not pregnant or lactating and not a psychiatric patient were included. MAIN OUTCOME MEASURES: Major outcome variables included changes in nutritional status as assessed by subjective global assessment (SGA) at hospital admission and discharge, length of stay, hospital charges, complications, and infections. STATISTICAL ANALYSIS PERFORMED: Analysis of variance with a Tukey adjustment for multiple comparisons was used to examine the impact of changes in nutritional status between nutrition change categories for continuous variables (charges and length of stay). Discrete variables were assessed using chi 2 analysis. Logistic regression was used to calculate odds ratios with 95% confidence intervals for the development of complications and infections when compared with the reference group. RESULTS: Compared with the reference group (normally nourished at admission and discharge), patients who declined nutritionally, regardless of nutritional status at admission, had significantly higher hospital charges ($28,631 +/- 1,835 vs $45,762 +/- 4,021). Odds of complications were significantly greater for patients who declined nutritionally, regardless of nutritional status at admission, compared with the reference group. APPLICATIONS/CONCLUSIONS: Declines in patients' nutritional status while they are hospitalized, regardless of their nutritional status at admission, were associated with significantly higher hospital charges and a higher likelihood of complications. Practicing clinicians should make reducing declines in patients' nutritional status a priority regardless of patients' nutritional status at admission. PMID- 11103654 TI - Both food preferences and food frequency scores predict fat intakes of women with breast cancer. AB - OBJECTIVE: To determine whether self-reported frequencies of food use were linked to self-reported preferences for the same foods. The hypothesis was that both food frequencies and food preferences can predict nutrient intakes. RESPONDENTS: Participants were adult women patients (n = 339), recruited through the University of Michigan Breast Care Center. The sample included both persons with breast cancer and persons who were cancer-free. DESIGN: All women completed a 98 item food frequency questionnaire and rated preferences for many of the same foods using a 9-point category scale. Percent energy from fat and saturated fat, and intakes of dietary fiber and vitamin C were estimated from analyses of 4-day food records. STATISTICAL ANALYSES: Pearson correlations coefficients were used for data analysis. RESULTS: Dislike was a strong predictor of nonuse. In contrast, the more preferred foods were also reported as more frequently consumed. Significant correlations between preference and frequency scores were obtained for virtually all item pairs. Median Pearson correlation coefficient was 0.30 (range 0.04 to 0.56). Correlations improved when foods were aggregated into the chief dietary sources of fat, saturated fat, and vitamin C. Food frequencies and food preferences showed the same strength of association with percent energy from fat and saturated fat (r = 0.20 to 0.25). Food frequencies showed a stronger association with vitamin C intakes than did preferences for vegetables and fruit. APPLICATIONS: Food preferences may provide a potential alternative to the food frequency approach. PMID- 11103655 TI - Pregnant adolescent and adult women have similarly low intakes of selected nutrients. AB - OBJECTIVE: To examine the dietary intake of pregnant adolescents during the second and third trimester of pregnancy, and to compare their nutrient intake with that of pregnant adults. DESIGN: Two 7-day food records (14 days) from subjects participating in a larger randomized clinical calcium trial: the first at 19 to 21 weeks and the second between 29 and 31 weeks gestation. Intake of energy and selected nutrients were calculated and compared with dietary standards. SUBJECTS/SETTING: Fifty-nine pregnant adolescents and 97 pregnant adults recruited from prenatal clinics at a metropolitan university hospital. STATISTICAL ANALYSES: Two sample t tests, equality of variances, and repeated measures (analysis of variance). RESULTS: There was no difference in mean nutrient intakes between the second and third trimesters. Using two 7-day food records, we found mean intakes for energy, iron, zinc, calcium, magnesium, folate, and vitamins D and E to be below recommended standards in both groups. Other nutrients examined met or exceeded reference values. Total daily intakes for energy and 11 nutrients were significantly higher in the adolescent compared to the adult diets (P < .05). These differences were not evident when nutrient values were corrected for energy, indicating that increased energy intake in the teen-aged population was contributed by nutrient-dense foods. APPLICATIONS: This study indicates the need for continued dietary monitoring of pregnant adolescents and pregnant adults, including nutrition guidance that stresses food sources of calcium, magnesium, zinc, iron, fiber, folate, and vitamins D and E, the nutrients found deficient in their diets. PMID- 11103656 TI - Parents' restrictive feeding practices are associated with young girls' negative self-evaluation of eating. AB - OBJECTIVE: This study was conducted to determine whether parents' restriction of young girls' access to palatable foods promotes the consumption of those foods while evoking negative self-evaluation. DESIGN: Girls' intake of 10 snack foods was measured immediately following a standard lunch, in a setting with free access to palatable snack foods. Girls' self-evaluation about their eating was assessed following the free access snack session. In addition, reports of parental restriction were obtained from mothers, fathers, and girls. PARTICIPANTS: Participants were 197 girls aged 4.6 to 6.4 years and their parents. STATISTICAL ANALYSIS: Structural equation modeling was used to test models describing relationships between parents' restriction and girls' eating. RESULTS: Following the standard lunch, girls' snack food intake during the 10 minute free access session ranged from 0 to 436 kcal, with a mean of 123 +/- 7 kcal. Approximately half of the girls reported negative self-evaluation about eating 1 or more of the 10 foods provided. The revised path model indicated that parents' restriction predicted both girls' snack food intake and girls' negative self-evaluation of eating. Girls' negative self-evaluation of eating was not associated with the amount of food that they consumed when not hungry, but was linked to their perceptions of being restricted from those foods. APPLICATIONS/CONCLUSIONS: These findings indicate that restricting young girls' access to palatable foods may promote the intake of restricted foods and may also generate negative feelings about eating restricted foods. PMID- 11103658 TI - Absence of nutritional or clinical consequences of decentralized bulk food portioning in elderly nursing home residents with dementia in Montreal. AB - OBJECTIVES: To evaluate the nutritional and clinical consequences of changing from a centralized food delivery system to decentralized bulk food portioning; a system in which meal portioning occurs on residents' floors of a nursing home. DESIGN: A pilot study with a pre-post design SUBJECTS/SETTING: The study took place on one floor of a home for elderly persons with dementia. Of the 34 residents, 22 (1 man) participated in this study. Average age was 82 years (range = 55 to 94 years). Nutritional status was verified before introduction of the bulk food portioning system by 3 nonconsecutive days of observed food intakes, anthropometric measurements (height, weight, triceps skinfold thickness, mid upper-arm circumference), and biochemical parameters (albumin, lymphocytes, glucose, sodium, potassium, transferrin, vitamin B-12, folate, hemoglobin). Trained dietitians collected the dietary and anthropometric data and validated the food intake estimates and anthropometric measurements. Data were also collected 10 weeks after implementation of the new food distribution system. STATISTICAL ANALYSES PERFORMED: Paired t tests adjusted by a Bonferroni correction assessed differences between values measured before and after introduction of the new food distribution system. RESULTS: Average food consumption increased substantially and significantly after introduction of the bulk food portioning system. Mean energy intakes rose from 1,555 to 1,924 kcal/day and most other nutrients also increased, many significantly, but there were no changes in anthropometric values or biochemical parameters, except for albumin level which decreased to the lower normal limit. APPLICATIONS: Portioning of food in the residents' dining room simulates a homelike atmosphere thereby encouraging increased food consumption. With well-trained and enthusiastic staff, this system could contribute to improved nutritional status in the very elderly, even those who have dementia. Dietitians have a key role to play in overseeing residents' nutritional needs and in training, supervising, and motivating foodservice personnel. PMID- 11103657 TI - Exercise mitigates the association of abdominal obesity with high-density lipoprotein cholesterol in premenopausal women: results from the third National Health and Nutrition Examination Survey. AB - OBJECTIVE: To examine the relationship between abdominal obesity, as measured by waist-to-hip ratio (WHR) and high-density lipoprotein cholesterol (HDL-C) level within the context of age, body fatness, exercise, saturated fat intake, and other plasma lipids. DESIGN/SUBJECTS: Subjects were premenopausal, white, non Hispanic women from the third National Health and Nutrition Examination Survey. Smokers, heavy drinkers, and women who took lipid-altering drugs were excluded. Of 1,188 subjects who met the inclusion criteria, complete data were available for 435 women. STATISTICAL ANALYSES: Means were calculated using all subjects for each variable, then F-protected t tests and linear contrasts were performed to test differences in means between subgroups. A P < .05 was considered significant. RESULTS: Age was not significantly associated with HDL-C level. Comparisons of HDL-C by WHR, percentage body fat (%BF), and exercise level revealed that HDL-C level was significantly lower at the higher levels of WHR and %BF and higher at the highest levels of exercise. Higher levels of HDL-C were generally accompanied by lower levels of triacylglycerol. When HDL-C was compared by exercise level within each WHR tertile and %BF tertile, the association of exercise with HDL-C diminished. Saturated fat intake was not associated with HDL C. CONCLUSIONS/APPLICATIONS: Increased exercise is associated with a lower WHR and subsequently a higher HDL-C level. This association between WHR and HDL-C appears to be mediated through %BF. Women exercisers with the highest WHR had consistently more favorable plasma lipid profiles and lower mean body mass index and %BF than nonexercisers. Thus, for women who exhibit abdominal obesity, exercise mitigates the association of WHR with HDL-C level. Vigorous exercise in the premenopausal years may promote a more favorable lipid profile, even in the presence of increased body fat and abdominal girth. PMID- 11103659 TI - Development of a standardized methodology for double-blind, placebo-controlled food challenge in patients with brittle asthma and perceived food intolerance. AB - OBJECTIVE: To develop a standardized, double-blind, placebo-controlled, food challenge (DBPCFC) methodology for identifying food intolerance in patients with brittle asthma. SUBJECTS/SETTING: Patients with brittle asthma and perceived food intolerance were studied in hospital. DESIGN: Each of 3 protocols began with 5 days of dietary exclusion. Protocol 1 consisted of open food challenges in 29 patients, protocol 2 consisted of 2 daily DBPCFCs in 22 patients, and protocol 3 involved 1 daily DBPCFC in 18 patients. Total immunoglobulin E level was measured and food-specific radioallergosorbent tests and skin prick tests were conducted. A standard panel of hyperallergenic foods were masked in a soup (developed specially for this study) for every food challenge. Peak expiratory flow, forced expiratory volume, and symptoms were assessed as objective measures of response. Open food challenges at home followed each protocol. Each protocol took approximately 14 days in the hospital and 4 to 6 months at home. RESULTS: For protocols 1, 2 and 3, positive reactions were experienced by 52%, 55%, and 66% of patients, respectively. Radioallergosorbent tests and skin prick tests were shown to have 40% and 71% sensitivity, respectively, and 74% and 77% specificity for predicting a positive food challenge. APPLICATIONS/CONCLUSIONS: The high prevalence of food intolerance in patients with brittle asthma was confirmed, as was the poor positive predictive value of skin prick tests and radioallergosorbent tests. The food challenge method developed enables standardized identification of food intolerances in patients with brittle asthma and may be useful in other groups. PMID- 11103660 TI - Potential mechanisms of diet therapy for fibrocystic breast conditions show inadequate evidence of effectiveness. AB - Fibrocystic breast conditions, formerly referred to as fibrocystic breast disease, affect about half of all women and typically present as any combination of breast nodularity, swelling, and pain. We reviewed the literature to evaluate evidence supporting nutrition interventions commonly recommended for fibrocystic breast conditions by health care providers. Randomized, controlled studies of the effectiveness of caffeine restriction fail to support any benefit in fibrocystic breast conditions. Similarly, evidence supporting evening primrose oil, vitamin E, or pyridoxine as treatments for the discomforts of fibrocystic breast conditions is insufficient to draw conclusions about effectiveness. Dietary alterations that influence the intermediate markers for fibrocystic breast conditions include low-fat (15% to 20% energy), high-fiber (30 g/day), and soy isoflavone regimens. However, our findings provide no solid evidence for secondary prevention or treatment of fibrocystic breast conditions through a dietary approach. Health care providers should limit recommendations to proven diet therapies supported by randomized, placebo-controlled trials, given the instability inherent in fibrocystic breast conditions and the near 20% placebo effect associated with intervention. Because excessive estrogen or altered sensitivity to estrogen is the dominant theory of etiology, interventions that may modulate endogenous steroid hormones warrant further investigation as potential treatments for symptomatic fibrocystic breast conditions. PMID- 11103661 TI - The latex and food allergy connection. AB - Natural rubber latex is used in the manufacture of many products in the United States. As natural rubber latex allergy becomes of increasing concern, dietitians need to have an understanding of this allergy and how it relates to workplace safety, employee health, and patient feeding and counseling. Natural rubber latex contains more than 35 proteins that may be related to Type I, Ig-E-mediated allergy in numerous segments of the population, including health care workers and patients. Many foods, especially chestnut, banana, and avocado, have the potential to cross-react with natural rubber latex. Chitinase enzymes, related to plant defense, are believed to be involved in this cross-reaction. A strong connection between food allergy and natural rubber latex allergy is recognized and described in this review. PMID- 11103662 TI - Nutrient adequacy of diets of adults with hypercholesterolemia after a cholesterol-lowering intervention: long-term assessment. PMID- 11103663 TI - Folate, iron, and dietary fiber contents of the gluten-free diet. PMID- 11103664 TI - Comparison of food groups and health claims appearing in food advertisements in 3 popular magazine categories. PMID- 11103665 TI - Fruit and vegetable consumption among Mexican-American college students. PMID- 11103666 TI - [Intensive treatment of blood pressure in patients with kidney disease and proteinuria]. AB - Blood pressure and proteinuria are important determinants of progressive renal failure in patients with renal diseases. In a 53-year-old man with hypertension and nephrotic-range proteinuria, lowering the blood pressure to a value of 125/75 mmHg resulted in a disappearance of the proteinuria. Recent literature data indicate that the treatment of blood pressure in patients with proteinuria, with emphasis on the benefits of reaching a blood pressure target of 125/75 mmHg and the use of angiotensin-converting enzyme inhibitors, may lead to a serious improvement in their prognosis. PMID- 11103667 TI - [Stereotaxic irradiation of vestibular schwannoma (acoustic neuroma)]. AB - A vestibular schwannoma (acoustic neurinoma) is a benign tumour localized in the cerebellopontine angle; it can give rise to cranial nerve symptoms. In recent years stereotactic irradiation has become an alternative to radical surgery. Stereotactic irradiation is administered with a gamma knife unit or with an adapted linear accelerator, as a single fraction (radiosurgery) or fractionated (stereotactic radiation therapy). Stereotactic irradiation gives local control rates of over 90%. Post treatment hearing preservation rate is over 60% and treatment related toxicity is low. Comparable treatment results are also found in the Netherlands at the VU-Ziekenhuis in Amsterdam. PMID- 11103668 TI - ['Advanced trauma life support' in Netherlands]. AB - Introduction of the principles of advanced trauma life support (ATLS) in the management of accident victims has been in progress in the Netherlands since 1995. The main ATLS principles are that the aid giver treats the most dangerous disorder first and does no further damage. After assessment and, if necessary, treatment of the airways, the respiration, the circulation and any craniocerebral injury, an exploratory examination is carried out. Physicians receive theoretical and practical instructions in this form of management during an intensive two-day course, counselled by a coordinating organization in the USA. Most of those attending are interns in general surgery, traumatology and orthopaedics, gatekeeper doctors of emergency rooms and army medical officers. The standardized way of thinking improves the communication and understanding between the various disciplines involved in trauma care, in part because there exist comparable programmes for ambulance care and emergency care. Other measures improving the quality of trauma care are regionalization of the trauma care, medical helicopter teams and evaluation of the effects of ATLS as an operating procedure. PMID- 11103669 TI - [Roaming through methodology. XXVI. The ecological fallacy and its less well known counterpart, the atomistic fallacy]. AB - Usually, individuals form the unit of observation and analysis in epidemiological studies, but such is not the case in the ecological study design in which determinants and diseases are related at the group level (school classes, companies, suburbs, countries). Ecological studies are subject to the ecological fallacy when they are used to make inferences on relationships between determinants and diseases at the individual level: the relationship on the group level may not reflect the relationship on the individual level. On the other hand, individual-level studies are subject to the atomistic fallacy, when they are used to make inferences on relationships between determinants and diseases at the group level: a relationship between a determinant and disease on group level may not be exclusively based on the relationship on the individual level. PMID- 11103670 TI - [From gene to disease; from p16 to melanoma]. AB - Approximately 10% of human cutaneous melanoma cases occur in families with the familial atypical multiple mole melanoma (FAMMM) syndrome, which is characterised by the familial occurrence of melanomas and atypical precursor naevi. A melanoma associated gene has been mapped to 9p2l, encoding for the tumour suppressor gene CDKN2A. Worldwide, germline mutations in melanoma kindreds implicate this cell cycle regulator (p16) as a susceptibility gene for malignant melanoma. Most FAMMM families registered at the Leiden Pigmented Lesions Clinic share the same CDKN2A inactivating deletion (P16-Leiden). Presymptomatic DNA diagnosis will now be available for P16-Leiden positive FAMMM family members at the Leiden University Medical Centre. PMID- 11103671 TI - [Diagnostic image (9). Chronic mucocutaneous candidiasis]. AB - An 18-year-old man had lifelong mucocutaneous lesions from which Candida albicans was persistently cultured. Upon fluconazole treatment the lesions diminished notably. PMID- 11103672 TI - [Experiences of women who decided to continue the pregnancy after diagnosis of Down's syndrome]. AB - OBJECTIVE: To gain insight into the motives and experiences of women who had decided to continue with the pregnancy after Down's syndrome had been diagnosed in the foetus. DESIGN: In-depth interviews. METHOD: In ten women who had decided to continue her pregnancy after Down's syndrome had been diagnosed in the foetus, qualitative in-depth interviews were held. Four women were pregnant at the time of the interview, the other six were parent of a Down's syndrome child already. One of the women was in her first pregnancy, the other nine had been pregnant once or several times. Four women had problems in their history (subfertility, miscarriage, in-vitro fertilisation). RESULTS: Many pregnant women were confronted with an increased risk as the result of maternal serum testing or nuchal translucency. They hoped to reduce the uncertainty which had arisen by submitting to an amniocentesis or a chorionic villus sampling. The result of this diagnostic test put those concerned in the position of having to make a difficult decision. They had to make the choice between having to bring up a child with intellectual limitations or allowing the termination of an already well-advanced pregnancy. For the ten respondents, the latter proved to be unacceptable. Initially, little understanding was shown for the parent's decision by some social and medical workers; however, sufficient help and support were usually given. The respondents received a lot of support from members of their family, friends and acquaintances, but there were also negative and disapproving reactions. Only one woman regretted the examination. CONCLUSION: As the technological possibilities for determining individual risks during pregnancy increase, it will occur more often that women hesitate to have their pregnancy terminated after diagnostic testing has identified Down's syndrome. Whatever decision is made, those involved should be treated with understanding. PMID- 11103673 TI - [Routine laboratory tests unnecessary for children referred for recurrent wheezing and/or asthma]. AB - OBJECTIVE: To investigate the usefulness of laboratory testing and thorax radiography in children, referred to the paediatrician for evaluation of recurrent wheezing. DESIGN: Retrospective. METHODS: In this study, 158 children referred for recurrent wheezing to a specialized child outpatient clinic of the Medisch Centrum Leeuwarden, the Netherlands, in the period 1 January 1994-31 December 1996, were evaluated according to a routine protocol including haemoglobin, ESR, leucocytes, immunoglobulins, sweat chloride levels and allergy testing and chest roentgenograms. It was determined whether these investigations had yielded abnormal results and whether these test results aided in confirming/rejecting the diagnosis of asthma or were helpful in clinical management. RESULTS: In 144 of the 158 (91%) children the diagnosis 'asthma' or 'recurrent wheezing' was made. Although numerous test results were abnormal they were not helpful in establishing the diagnosis. In only one child an abnormal chest radiograph was helpful (the radiograph showed infiltrative abnormalities). Tests for aero-allergy were rarely positive in children younger than 2 years; in children older than 6 years aero-allergy was found frequently, notably to dust mite (41/144). CONCLUSION: The results of this study suggest that--except for allergy testing--routine laboratory testing and chest roentgenograms are not indicated in children referred for evaluation of wheezing disorders. Aero-allergy testing may help to decide on preventive measures. PMID- 11103674 TI - [Distraction osteogenesis of the mandible in 2 children with obstruction of the upper respiratory tract due to micrognathia]. AB - A girl approximately 2 years old with Pierre Robin sequence had periods of nocturnal respiratory insufficiency as a consequence of micrognathia and a boy nearly 4 years old with Nager syndrome and tracheostomy was retarded in his speech development, had problems swallowing and often had respiratory tract infections. The obstruction of the upper respiratory tract was resolved by performing a distraction osteogenesis of the lower jaw. A tracheostomy was avoided or else removed. The girl became more active and there was an improvement in the boy's speech development. Distraction osteogenesis is a good alternative to the current practice of mandibular reconstruction in micrognathic patients, which involves extensive bone grafts. PMID- 11103675 TI - [Acquired medical condition and driving ability]. AB - Under Dutch law a driver's licence requires a written statement of physical and mental health from the applicant. However, after the licence has been issued, no further statements are required, not even when the person involved develops a medical condition that poses a threat to safe driving. If an accident occurs and it is established that the driver suffered from an intercurrently acquired condition, the insurance companies will not pay. Physicians have a moral obligation to advise patients in their care whom this may affect to report to the physician of the licence administration. PMID- 11103676 TI - [Increased proportion of elementary school children with asthmatic symptoms in the Netherlands 1984/85-1994/95; a literature review]. PMID- 11103677 TI - [Increased proportion of elementary school children with asthmatic symptoms in the Netherlands, 1984/85-1994/95; a literature review]. PMID- 11103678 TI - [Favorable results of surgical treatment of mucoid cysts of the fingers and thumb in 20 patients, Leyenburg Hospital, Den Haag, 1992-99]. PMID- 11103679 TI - [The importance of the endogenous cannabinoid system in various neuropsychiatric disorders]. AB - The endogenous cannabinoid system was first described in 1988. There are two specific receptors, the CB2-receptor, located in the lymphatic system (spleen, lymphocytes), and the CB1-receptor occurring predominantly in the central nervous system. The CB1-receptor shows a distinct distribution in the CNS with a very high density in the cerebellum, the basal ganglia and in the hippocampus. In 1992 endogenous ligands of the cannabinoid system were discovered for the first time (e.g. anandamide and 2-arachidonylglycerol). The physiological role of these arachidonic acid derivates is still unclear. Implications of these recent discoveries for the Gilles de la Tourette syndrome, ischaemic brain lesions, schizophrenic psychoses and opiate drug dependence are described. A dysregulation in the endogenous cannabinoid/anandamide system could possibly play an import role in the etiology of Gilles de la Tourette syndrome and schizophrenic psychoses; administration of cannabinoids affects the symptoms of the Gilles de la Tourette syndrome positively, whereas cannabinoids probably have rather negative effects in schizophrenic psychoses. In ischaemic brain lesions cannabinoids seem to have a neuroprotective effect; they appear to minimize the extent of a lesion by reduction of glutamate release. Additionally the meaning of the endogenous cannabinoid system for the development of opioid drug dependency is discussed and interactions between the endogenous opioid system and the endogenous cannabainoid system are pointed out. This is of interest since it could be shown in animal experiments that the absence of CB1 receptors reduces the positive reinforcement of opiate administration. PMID- 11103681 TI - [Clinical aspects of frontotemporal dementia]. AB - Frontotemporal neurodegeneration can cause three typical clinical syndromes: frontotemporal dementia (FTD), primary progressive aphasia (PPA) and semantic dementia (SD). In the present paper we review these syndromes, highlighting FTD. Four case examples are presented. At the early stage of FTD changes of personality and social conduct are prominent, whereas cognitive functions are relatively well preserved. Since the usual dementia tests are not sufficiently sensitive to disclose non-cognitive symptoms, clinical diagnosis as well as differentiation from non-organic psychiatric disorders can be difficult. Detailed history, thorough clinical examination, and neuropsychological testing are required to establish the diagnosis. EEG and functional brain imaging may be helpful. The choice of therapeutic options for FTD is extremely limited. Medications may be used to treat neuropsychiatric symptoms. There is little experience with non-pharmacologic behaviour modification and milieu treatment approaches. The problems that FTD imposes on caregivers are dissimilar to those arising from Alzheimer's disease. Families receive little or no support so that early nursing home admission of patients is common. PMID- 11103680 TI - [Cerebrospinal fluid protein tau levels in the differential diagnosis of Alzheimer's disease]. AB - Tau protein concentration in cerebrospinal fluid was determined in 55 patients with Alzheimer's disease (AD), 18 patients with vascular dementia (VD), 19 patients with dementia caused by other disorders and 14 patients with major depression. Significantly (p < 0.05) elevated protein tau concentrations were found in AD patients (564.5 +/- 275.5 pg/ml) compared to all other patient groups (VD: 406.5 +/- 263.9 pg/ml; other dementia: 275.0 +/- 135.4 pg/ml; depression: 212.9 +/- 115.6 pg/ml). However, tau levels in AD patients covered a broad range (163.2 pg/ml-1200 pg/ml). AD patients with tau levels below the 25%-percentile of the distribution (among them a high percentage of patients with presenile onset) showed tau levels similar to those of the patients with late life depression. No significant correlations between tau levels and clinical variables such as severity of dementia, age, age of onset, duration of illness, and cerebral changes as assessed by volumetric magnetic resonance imaging could be demonstrated. Similarly, we could not find an influence of either APO-E genotype or psychotropic medication on the tau levels in AD patients. In accordance with other studies our results confirm elevated tau levels in AD compared to elderly not demented control subjects. Comparing groups, this finding applies as well with respect to VD and other dementing disorders. However, elevated tau levels cannot be detected in a subgroup of AD patients. This finding needs to be further investigated in future studies. PMID- 11103682 TI - [Posttraumatic stress disorder in patients with neurogenic amnesia for the traumatic event]. AB - The development of symptoms of posttraumatic stress disorder (PTSD) in patients with neurogenic amnesia for the traumatic event is recorded in 2 own patients and in 19 cases from the clinical literature. With a single exception, all patients were accident victims with closed head injuries. Only about three quarters of the patients completely fulfilled DSM-III-R criteria of PTSD. Nineteen patients displayed involuntary conscious memories of aspects of the traumatic event (presenting as recurrent intrusive thoughts, images or dreams) co-existent with a complete or partial lack of voluntary conscious memories of the trauma, suggesting that different memory systems and distinct brain mechanisms subserve these phenomena. The said clinical observations are discussed against the background of current neuropsychological models of multiple memory systems. The recorded cases demonstrate that declarative episodic memory is not necessary for symptoms of PTSD to emerge, whereas preserved functions of non-declarative memory systems represent a sufficient condition for the development of PTSD symptoms. PMID- 11103683 TI - [A transcultural study of eye perception. Comparison between social phobia and tai-jin-kyofu]. AB - Recently Western psychiatry has been paying more attention to the sense of shame, as is evident from the introduction of the concept of sociophobia which may be regarded as a typical example of "shame-disease", to DSM and ICD. Sociophobia is without doubt similar to Tai-jin-kyofu(= TK), which was often considered as a specifically Japanese culture syndrome. To explore the relation between sociophobia and TK syndrome, the eye perception of the sociophobic patient is compared with that of the TK Patient. The former, to which until now--different from the latter--little attention has been paid, is presented by three cases. This approach shows up a cultural difference between the basic orientation toward the individual in the West and that toward Ma ("Between") in Japan. PMID- 11103684 TI - [The Susac syndrome--a retinocochleocerebral angiopathy]. AB - INTRODUCTION: There has been a series of case reports of otherwise healthy patients suffering from microangiopathy of the brain, retina and cochlea. Most patients were young women presenting clinically with a subacute encephalopathy, branch retina artery occlusions, and hearing loss. In 1994 the name "Susac syndrome" has been proposed for this disease entity. METHOD: Case report and review of 64 published cases, identified through MEDLINE are given. CASE REPORT: We describe a 32-year-old otherwise healthy woman presenting with a subacute encephalopathy, multiple branch retinal artery occlusions and bilateral hearing loss. MRI of the brain revealed multiple small white and grey matter lesions without contrast enhancement. CSF protein was elevated, oligoclonal bands were negative. Immunological laboratory parameters, microbiology, virology, coagulation studies, SEP, AEP, VEP and cerebral DSA were normal. REVIEW OF THE LITERATURE: Of 64 identified patients 58 were women. The mean age of the patients was 30 years. 60 patients (94%) had arterial occlusions, which were bilateral in 39%. 48 patients reported hearing loss, 37 patients (58%) had a global encephalopathy, but other neurologic manifestations were common. CONCLUSION: This rare syndrome has a strong young female preponderance. MRI of the brain often shows lesions suggestive of multiple sclerosis. Fluorescein angiography may show arteriolar wall hyperfluorescence. Patients can be identified at an early stage with a careful history and physical examination. Early treatment with corticosteroids is often associated with a good prognosis. Cyclophosphamide and antiplatelet drugs may be added in complicated cases. PMID- 11103686 TI - [The musician's hand: aspects of music physiology and performing arts medicine]. AB - Musicians are particularly demanding hand patients. Increased expectancy towards the hand surgeon as well as principle aversion to any kind of operative treatment of the musician's hand impede the relationship between hand surgeon and patient. Knowledge and experience of the specific demands towards the musician's hand- including the background of basics in physiology of music making and instrumental techniques--facilitate history taking, examinations, diagnosing as well as the selection of therapy. Besides tendinitis and tendovaginitis, pathologies of the musician's hand are not more frequent compared to the entire population. Still several pathologies reach a significantly pronounced importance through the specific professional requirements. This is true especially for peripheral nerve compression syndromes, arthritis, ganglia, contractures, joint laxity and focal dystonias. The treatment of these disorders and hand trauma require, besides an utmost of hand surgical care, creativity and the implementation of the professional musical surroundings in therapy and rehabilitation, including also the musical instruments. The indication for surgery might be restrained in certain cases, although there are, without doubt, situations, when a professional musician should be operated on at an early stage. This should enhance a rapid return to play the musical instrument and to exercise delicate and demanding skilled movement patterns of his hand. PMID- 11103687 TI - [The musician's hand: 2 case reports from the hand surgery clinic]. AB - Report of the hand surgical treatment in two musicians, which enabled them to continue with their occupation. A young cellist was no longer able to play his instrument due to a painful scar at the pulp of his left index finger. The surgical treatment, performed only after a period of hesitation, consisted in scar excision and reconstruction of a normal pulp by a palmar advancement flap. Following a fracture of the distal radius, healed in 35 degrees dorsal angulation, a violinist was not able to flex his wrist maximally as required for playing in high octaves. This was again possible after a corrective osteotomy with interposition of a corticocancellous bone graft from the iliac crest and an intensive program of postoperative excercises. The important role of the cooperation of the patient, particularly in the rehabilitation, is emphasized. PMID- 11103688 TI - [Reduction of breasts ... Hans Schaller and the first mammaplasty in 1561. Contribution to history of medicine]. AB - In comparison with other surgical procedures concerning the breast, the history of reduction mammaplasty is relatively short. Some authors have mistaken Paulos of Aegina for the pioneer in this field, although he occupied himself exclusively with gynaecomastia. Since some decades Hanns Schaller, the so-called "barber" of Augsburg, is considered to be the first surgeon to have performed a reduction mammaplasty by breast amputation in 1561. However, exact references have not been available so far. We found the original text containing the description of the procedure written by a contemporary in a rather unexpected place as well as some details about the surgeon. We conclude that Hanns Schaller was the first surgeon to undertake a reduction mammaplasty in an otherwise healthy woman in order to relieve her physical symptoms. Undoubtedly, his intentions were purely functional without any further aesthetic considerations. PMID- 11103689 TI - [Ulnar lengthening in osteochondroma (multiple cartilagenous exostoses) of the forearm]. AB - The osteochondroma is the most frequent bone tumor to occur during the period of growth. The multiple hereditary form often involves the forearms. Depending on localisation and size of the tumor, shortening of the bones in the forearm, angular malalignments and functional impairment of the wrist and elbow joints may result. Early diagnosis and surgery in the growing child can prevent these complications. 13 children were operated on altogether 15 forearms. In nine cases lengthening of the ulna was necessary to correct ulnar instability of the wrist as well as improving the support of the carpus and preventing dislocation of the radial head. PMID- 11103690 TI - [Commentary on the article by B. Bader and F. Grill: Ulnar lengthening in osteochondroma (multiple cartilagenous exostoses) of the forearm]. PMID- 11103691 TI - [Results after minimally invasive therapy of acute scapholunate dissociation]. AB - In 27 cases with acute scapholunate ligament injury, minimal invasive treatment was performed. After closed reduction scapholunate transfixation was achieved by Kirschner wires. Postoperatively, a short-arm cast was applied for six weeks. All patients have been reexamined on average 26 months after operation. Follow-up results have been good concerning clinical examination and patients' satisfaction. Radiological examination has shown recurrence of scapholunate instability in 15 cases. In fact, 11% of the patients had dynamic and 44% static instability at follow-up. Recurrence has been most frequent in cases involving primary static instability. Consequently, these cases should be treated by open reduction. Minimal invasive treatment of acute scapholunate dissociation is not recommended. The therapeutic procedure must be adapted to the different stages of scapholunate ligament injury. PMID- 11103692 TI - [Radiocarpal fracture dislocation]. AB - Radiocarpal fracture dislocation is a very rare, often complex injury and the result of high energy trauma. We present a retrospective review of four patients. The injury in our series was characterized by a complete dorsal dislocation of the radiocarpal joint with fracture of the radial styloid, associated with avulsion of the palmar and dorsal cortical margin of the distal radius (radiotriquetral ligament included). One case presented a die punch fracture, another case a partial lesion of the scapholunate ligament. All injuries were immediately treated by closed reduction in local anaesthesia. All patients underwent operative treatment with a dorsal approach within seven days. The radial styloid was fixed with screws or Kirschner-wire, the impaction of the dorsoulnar aspect of the radius with elevation and fixation with a 2.7 mm plate. The tear of the scapholunate ligament was sutured and stabilized with Kirschner wire between scaphoid and lunatum. All wrists were immobilized with a forearm cast for four weeks. The mean follow-up was 25 months. At the time of follow-up all patients showed a very good functional outcome, although the radiographic analysis revealed some osteoarthrotic changes. PMID- 11103693 TI - [Chronic paronychia and synovialitis of extensor tendons due to Mycobacterium marinum. Is diagnosis or treatment the problem?]. AB - Most infections of the upper extremity are caused by staphylococcus or streptococcus and respond well to beta-lactam antibiotics. Hand surgeons should be aware of the possible diagnosis of Mycobacterium marinum infection: 90% of the lesions are found in the upper extremity. We present a case of a chronic, cutaneous lesion of the right middle finger with synovialitis of the extensor tendons observed in a 35-year-old woman. Routine cultures from tissue of the infected finger led to the diagnosis of paronychia due to staphylococcus aureus. Despite surgical and antibacterial treatment, the lesion persisted and the patient developed multiple raised, non-tender satellite lesions to the right hand and elbow. Based on the clinical aspect and a detailed history (she kept fish and had suffered a chicken bone stab to her middle finger 12 weeks earlier), we suspected a Mycobacterium marinum infection. Tissue was obtained mainly by synovialectomy. Culture of the biopsy tissue for Mycobacterium marinum confirmed the diagnosis. The patient responded to a triple therapy (rifabutin, ethambutol and clarithromycin) and had an uncomplicated recovery. The importance of a high index of suspicion, adequate examination and a complete patient's history for a correct diagnosis is stressed. Culture for Mycobacterium marinum is not routinely performed and ought to be initiated once an infection is suspected. We also discuss the best timing for the onset of medical treatment. PMID- 11103694 TI - [DIGITOS-prosthesis for the proximal interphalangeal joint. A 2-year follow-up]. AB - The treatment of osteoarthrosis of the proximal interphalangeal joint by prosthesis bears several specific problems. With the DIGITOS-prosthesis a new implant is now available. It is a cemented modular hinged prosthesis. We report our experiences with this implant from a prospective clinical trial. Seven patients with osteoarthrosis of the proximal interphalangeal joint had ten prosthetic middle joint replacements performed with DIGITOS-prostheses. Over an observation period of two years, the functional results were found to be good. The mean preoperative range of motion in the affected joints was 51.5 degrees. Three months after surgery, this had improved to 60.5 degrees, but then decreased to 53.0 degrees after one year and to 49.5 degrees, i.e. the original range of motion, by the end of the second postoperative year. No implant became loose. Nearly all patients were free from pain. The only serious complication was an iatrogenic lesion of the middle slip of the extensor aponeurosis resulting in a buttonhole deformity. In spite of these encouraging two-year results, the further follow-up has to be observed carefully. PMID- 11103695 TI - Communication in the evolving world of case management. PMID- 11103696 TI - Expanding the boundaries of primary care for elderly people. AB - This article reports the results of a qualitative evaluation of the Generalist Physician Initiative, designed to enhance the care of older people provided by primary care physicians in nine demonstration projects around the country. A theme entitled "Pushing the Comfort Zone" examines activities in which physicians engage before collaboration: selecting elderly patients, "opening cans of worms," recognizing patient and family expectations, and going outside the comfort zone. A second theme called "Linking with Collaborators" reveals activities in which physicians engage as they collaborate: teaming, using intervention agents as eyes and ears, communicating, and tracking patients. Findings indicate that social workers are logical collaborators with primary care physicians as contemporary practice is expanding to be more holistic. PMID- 11103697 TI - Examining the HIV/AIDS case management process. AB - Eight themes in the HIV/AIDS case management process emerged from a 1998 study of 14 Ryan White Title I-funded case management programs in the New York City tri county region. For individuals who were struggling with multiple environmental stressors, the diagnosis of HIV or AIDS was merely one of the many pressures that brought them to case management programs. Most came when they were in crisis. Using both chart reviews and focus groups with case managers and supervisors, this article reports that the activities that characterize this region's case management introduce alternative ways of thinking about the HIV/AIDS case management process. PMID- 11103698 TI - The Caregiver Well-Being Scale revisited. AB - The Caregiver Well-Being Scale measures caregiver well-being from a strengths based perspective by assessing caregivers' basic human needs and satisfaction with activities of daily living. This article revisits the scale to examine further the scale's psychometric properties using a caregiver-only sample. Reliability is determined through internal consistency. Construct validity is supported through factorial validity with factor analysis. Criterion-related validity is established by examining the concurrent validity of the Well-Being Scale with a measure of depression. Using a sample of family caregivers, results suggest that the Well-Being Scale is a valid and reliable measure. PMID- 11103699 TI - Utilization review: a powerful social work role in health care settings. PMID- 11103700 TI - Inequality and health: implications for social work. PMID- 11103701 TI - Suggestions to social workers for surviving in managed care. PMID- 11103702 TI - Long-term care for people with developmental disabilities. PMID- 11103703 TI - Biomedical engineering: yesterday, today, and tomorrow. PMID- 11103704 TI - Glucose monitoring using implanted fluorescent microspheres. PMID- 11103705 TI - Finite element modeling of the neuron-electrode interface. PMID- 11103706 TI - Effects of shoulder stability on endpoint stiffness. PMID- 11103707 TI - Engineering a more accessible world. PMID- 11103708 TI - Temperature-compensated bioimpedance system for estimating body composition. PMID- 11103709 TI - Noninvasive early detection of focal cerebral ischemia. PMID- 11103710 TI - Constructing head models by computation. PMID- 11103711 TI - Antenatal fetal risk assessment by blood-flow velocity waveforms. PMID- 11103712 TI - Syntactic pattern recognition for X-ray diagnosis of pancreatic cancer. PMID- 11103713 TI - Pulse type classification by varying contact pressure. AB - We introduced a new technique of quantification of pulse characteristics in connection with contact pressure. We provided data only for illustrative purposes, deferring statistical analysis of samples of a larger population. This quantification can be useful for diagnostic purposes; for example, by detecting changes of the three quantities according to the health condition or particular disease of a patient, or by analyzing correlations among these quantities and other physiological variables. PMID- 11103714 TI - Topics in membrane electrophysiology. Aspects of nerve conduction. PMID- 11103715 TI - Interpretation of genome expression patterns: computational challenges and opportunities. PMID- 11103716 TI - When does the Freedom of Information Act apply to privately held data produced under a federal grant? (Part II). PMID- 11103717 TI - Variability in centred house-of-cards mutation models. AB - Convergence of variability in phenotypic models with balance between selection and mutation is analysed. The mutation assumed occurs with weak probability and brings down the evolutionary process built up by selection around the mean in the population. Gaussian approximations are used. PMID- 11103718 TI - Multiple dose vaccination against childhood diseases: high coverage with the first dose remains crucial for eradication. AB - The high vaccination coverage required to eradicate communicable diseases like measles, mumps and rubella, with a single dose of vaccine, has prompted many countries to introduce a second dose. In this paper we investigate the conditions to eradicate childhood diseases with multiple doses of vaccine by obtaining explicit analytical solutions to the classical compartment model that assumes an age-independent force of infection and conceptualizes the host population as divided into maternally protected (P), susceptibles (S), latents (E), infectious (I), and removed (R). The solutions allow a quantitative discussion of the long term impact of vaccination schedules with an arbitrary number of doses of vaccine. It becomes possible to determine the effect of the number of doses, ages at vaccination, and coverage rates of vaccines against childhood diseases. In an example with a two-dose vaccination schedule against measles, we show that, in spite of a second dose, a high (> 90%) immunization coverage in the first dose is still crucial to achieve eradication. With a high first-dose coverage, however, eradication is relatively insensitive to the age of the second dose and requires only moderate coverage rates in the latter. PMID- 11103719 TI - Behaviour change and treatment of core groups: its effect on the spread of HIV/AIDS. AB - A general model is considered for treatment and behaviour change of the Human Immunodeficiency Virus (HIV) infected in a highly sexually active core group of female commercial sex workers (CSWs) and a 'bridge population' of young unpartnered males. In this model, the spread of HIV/AIDS in the community is carried out mainly through the sexual interaction between the core group and the bridge population which acts as a bridge for the spread of disease to the general population. We will consider the effect of treatment of the infected and/or the subsequent behaviour change when targeted toward the core group and the bridge population. Analytical results will be given for a strategy which targets treatment and behaviour change at either the core group or the bridge population. Numerical examples are also provided to illustrate the biological significance of the treatment/behaviour change and its effect on the threshold parameter values. The results show that if the contact rates and transmission probabilities of the treated individuals are sufficiently reduced, the treatment/behaviour change can eradicate the disease provided that the level of treatment in the infected population is sufficiently high. However, an ill-planned treatment program which fails to meet the required reductions in contact rate or transmission probability could have a detrimental effect on the spread of the epidemic. PMID- 11103720 TI - Higher-dimensional separation principle for the analysis of relaxation oscillations in nonlinear systems: application to a model of HIV infection. AB - In this paper, geometric and singular perturbation arguments are utilized to develop a separation condition for the identification of limit cycles in higher dimensional (n > or = 4) dynamical systems characterized by highly diversified time responses, in which there exists an (n - 3)-dimensional subsystem which quickly reaches a quasi-steady state. The condition, which has been used up to now to analyze relaxation oscillation in slow-fast systems, is extended to accommodate dynamical systems in which more state variables are involved in a special manner which still allows for the use of singular perturbation techniques. Application is then made to a model of human immunodeficiency virus infection in T helper (TH) cell clones with limiting resting TH cell supply. PMID- 11103721 TI - Stochastic models for systems of interacting ion channels. AB - We consider a variety of Markov based models for systems of ion channels exhibiting dependence between channels. It is shown how many useful properties which may be calculated for an aggregated single-channel model, or a system of independent channels, can be extended to various types of interacting channel systems. Key structure and results from the theory of aggregated Markov processes are summarized in a convenient matrix form. These are then applied to the superposition of independent and dependent channels, including a patch of channels in a random environment, and a system of channels with spatial interactions. Calculations based on the resultant matrix expressions and intensity arguments can be implemented straightforwardly in a matrix-oriented package such as Matlab. The role of reversibility is also studied. A number of examples illustrate the strengths of the methods and enable numerical comparisons between the different types of systems. PMID- 11103722 TI - Increasing paramedics' comfort and knowledge about children with special health care needs. AB - This study evaluated a continuing education program for paramedics about children with special health care needs (CSHCN). Pretraining, posttraining, and follow-up surveys containing two scales (comfort with CSHCN management skills and comfort with Pediatric Advanced Life Support [PALS] skills) were administered. Objective measures of knowledge were obtained from pre- and posttraining tests. Differences in average scores were assessed using t-tests. Response rates for paramedics completing the program ranged from 94% for the posttraining survey, 81% for the initial comfort survey, 56% for the knowledge pretest, and 56% for the follow-up survey. PALS comfort scores were significantly higher than CSHCN comfort scores both before and after training, both P < .01. Posttraining surveys showed an increase in CSHCN comfort, P < .01. The follow-up surveys showed a significant decline in CSHCN comfort, P = .05. Scores on the tests showed a similar pattern, with a significant increase in knowledge from pre- to posttraining (P = .02) and a significant decrease in knowledge from posttraining to follow-up (P < .01). Comfort was significantly higher for standard pediatric skills than for specialized management skills. Completion of the self-study program was associated with an increase in comfort and knowledge, but there was some decay over time. PMID- 11103723 TI - Prospective randomized study of analgesic use for ED patients with right lower quadrant abdominal pain. AB - Giving an analgesic to patients with right lower quadrant (RLQ) pain causes greater alteration of abdominal signs predictive of appendicitis than placebo. A randomized double-blinded controlled trial of 68 patients who received either tramadol or placebo. Absence or presence of seven abdominal signs (tenderness on light and deep palpation, tenderness in the RLQ and elsewhere, rebound, cough, and percussion tenderness) and pain (100 mm Visual Analog Scale [VAS]) at 0 and 30 minutes were recorded. The predictive value of each physical finding (PF) was measured using an 11-point PF score weighted by likelihood ratios. There was significant reduction in mean VAS of 14.2 mm (95% CI 5.6 to 22.8) in analgesic group versus 6.5 mm (95% CI 1.6 to 11.4) in placebo group. The analgesic group had less normalization of signs as measured by the PF score in all patients [32 of 154 (20.8%) versus 40 of 121 (33.1 %) (P = .031)] and in those with proven appendicitis [4 of 33 (12.1%) versus 10/22 (45.5%) (P = .014)]. Parenteral use of tramadol in emergency department patients with RLQ pain resulted in significant levels of pain reduction without concurrent normalisation of abdominal examination findings indicative of acute appendicitis. PMID- 11103724 TI - Etomidate versus succinylcholine for intubation in an air medical setting. AB - The objective was to compare rates of successful endotracheal intubation (ETI) and requirement for multiple ETI attempts in patients receiving etomidate (ETOM) versus succinylcholine (SUX). This retrospective study analyzed adults in whom oral ETI was attempted by a helicopter EMS (HEMS) service between July 1997 to July 1999. Data were from records of the HEMS service, which uses a RN/EMTP crew; analysis was with chi-square and logistic regression (P = .05). ETI was successful in 269 (97.8%) of 275 patients, with multiple attempts occurring in 54 (20.1%) of 269. Success rates for SUX (209 of 213, 98.1%) and ETOM (60 of 62, 96.8%) were similar (P = .62). However, of 60 ETOM patients successfully intubated, 7 (11.7%) required rescue succinylcholine. When these patients are tallied as ETOM failures and SUX successes, resultant success rates for ETOM (86.9%) and SUX (98.2%) are different (P = .001). ETOM patients were more likely (P = .004) than SUX patients to require multiple attempts (33.3% versus 16.3%). ETI success rates were high in patients receiving SUX or ETOM as primary adjuncts for airway control, but initial success was more likely with SUX, and ETOM patients were more likely to require multiple attempts. PMID- 11103725 TI - Medical communication: do our patients understand? AB - The objective of this study was to determine emergency department (ED) patient's understanding of common medical terms used by health care providers (HCP). Consecutive patients over 18 years of age having nonurgent conditions were recruited from the EDs of an urban and a suburban hospital between the hours of 7 a.m. and 11 p.m. Patients were asked whether six pairs of terms had the same or different meaning and scored on the number of correct answers (maximum score 6). Multiple linear regression analysis was used to assess possible relationships between test scores and age, sex, hospital site, highest education level, and predicted household income (determined from zip code). Two hundred forty-nine patients (130 men/119 women) ranging in age from 18 to 87 years old (mean = 39.4, SD = 14.9) were enrolled on the study. The mean number of correct responses was 2.8 (SD = 1.2). The percentage of patients that did not recognize analogous terms was 79% for bleeding versus hemorrhage, 78% for broken versus fractured bone, 74% for heart attack versus myocardial infarction, and 38% for stitches versus sutures. The percentage that did not recognize nonanalogous terms was 37% for diarrhea versus loose stools, and 10% for cast versus splint. Regression analysis (R2 = .13) revealed a significant positive independent relationship between test score and age (P < .024), education (P < .001), and suburban hospital site (P < .004). Predicted income had a significant relationship with test score (P < .001); however, this was no longer significant when controlled for the confounding influence of age, education and hospital site. Medical terminology is often poorly understood, especially by young, urban, poorly educated patients. Emergency health care providers should remember that even commonly used medical terminology should be carefully explained to their patients. PMID- 11103726 TI - Hospital factors associated with emergency center patients leaving without being seen. AB - We developed a statistical model that would identify and quantify the relative contributions of different factors hypothesized to impact the frequency of emergency center (EC) patients who leave without being seen (LWBS). We performed an analysis of the daily counts of patients that registered in our EC during a 21 month period who then LWBS. Candidate predictor variables included the number of patients seen, and the number admitted to the hospital, for each area of our EC, as well as the hours of faculty double coverage, and the day of the week. Univariate analyses were performed using standard methods. Multivariate analysis was performed using the general linear model. A backward selection procedure was used to eliminate statistically insignificant variables until all remaining independent variables had P-values < or = .05. External validation and analysis of the stability of the estimated regression coefficients of the model were evaluated using bootstrap methods. Two-tailed tests and a type I error of 0.05 were used. During the period studied, 133,666 patients were registered in the EC and 9,894 (7.4%) left. Multivariate analysis identified six variables that were significantly associated with LWBS. The fitted model containing all six variables explained 52.8% of the variability observed in LWBS frequency. The most powerful predictor of LWBS was total number of patients cared for in the main ED. This accounted for 46.4% of the observed variation in LWBS. The total number of trauma and resuscitation patients, and the total number of observation unit admissions to the hospital were also associated with increased LWBS. More pediatric cases seen in the main ED, weekends, and additional faculty coverage were associated with fewer patients leaving. Efforts to decrease the LWBS rate will be most successful if they address the issue of main ED volume. PMID- 11103727 TI - Which chest pain patients potentially benefit from continuous 12-lead ST-segment monitoring with automated serial ECG? AB - A prospective observational study was performed in 678 chest pain patients with suspected acute coronary ischemic syndrome (ACS) and absence of clinical and ECG criteria for emergent reperfusion therapy on presentation to determine how often continuous 12-lead ST-segment monitoring with automated serial ECG (SECG) results in a significant change in therapy during the initial emergency department (ED) evaluation in typical high- and low-risk chest pain patients. After initial history, physical, and ECG were obtained, patients were grouped into high and low risk subgroups based on ED physician's assessment of likelihood of ACS. Significant change in therapy was defined as thrombolytic drug administration, emergent percutaneous coronary angioplasty (PTCA), and intensive anti-ischemic therapy with intravenous heparin and/or intravenous nitroglycerin. SECG monitoring was continued until either the patient was taken for emergent PTCA or until 2-hour serum markers measurements were obtained. A total of 26 patients therapy was changed secondary to SECG monitoring which represented 14.6% of high risk patients and 1.1% of low-risk patients. New injury (21 patients) and new ischemia (4 patients) were the only SECG findings that led to a change in therapy. SECG monitoring had a 15.2 times increased odds of changing therapy in the high risk patients as compared with the low risk patients (P < .0001; 95% CI 6.1 to 38.2). Chest pain evaluation protocols that exclude these high risk ED patients from SECG monitoring should be reevaluated. Our data also suggests that researchers designing randomized studies to show utility of SECG monitoring should focus on the high-risk patients. PMID- 11103728 TI - Outcome analysis of chest pain patients discharged from the ED--a pilot study. AB - The objective of this pilot study was to determine clinical predictors of adverse outcome, defined as myocardial infarction, angioplasy or stent placement, coronary artery bypass graft, or death, within 60 days for patients discharged from the emergency department with a presenting complaint of chest pain. All patients presenting to the emergency department with a chief complaint of chest pain were eligible for the study. A chest pain risk analysis sheet was completed as part of the patient evaluation. Patients discharged from the emergency department, in whom a risk analysis sheet was completed, were contacted to determine their clinical course within 60 days of their discharge from the emergency department. During the 6-month study period, 129 eligible patients were enrolled. Of these 129 patients, four had an adverse outcome within 60 days of their discharge. All four patients had either a balloon angioplasty procedure, coronary artery bypass graft, or both. None of the study patients had a myocardial infarction or died. Statistically significant predictors of adverse outcome in our study population were an abnormal electrocardiogram (ECG), a history of myocardial infarction, and a history of hypertension. In conclusion, patients discharged from the emergency department with a presenting complaint of chest pain were at a low risk for having a myocardial infarction or dying within 60 days of their discharge. Several patients, however, did have significant coronary artery disease requiring angioplasty or bypass. These patients were more likely to have an abnormal ECG, a history of myocardial infarction, or have a history of hypertension. A prospective study with larger numbers of patients is needed to validate these findings. PMID- 11103729 TI - The frequency of blood pressure measurements in children in four EDs. AB - The study's objective was to assess the frequency of triage blood pressure measurements in pediatric patients and the recognition of an elevated blood pressure. The design was retrospective and included chart review. The setting consisted of four emergency departments associated with one medical school, including one level I academic center, two level II Community departments, and a regional children's hospital. A convenience sample of 437 patients, aged 1 month to 18 years, was selected. The frequency of triage blood pressure measurements was recorded. The number of patients whose blood pressure was higher than the 90th percentile for age and sex as established by the Second Task Force on Blood Pressure Control in Children was also recorded. The frequency of a second blood pressure measurement in patients with an elevated initial blood pressure was recorded. All frequency data were stratified by hospital and by age group. The results showed 294/437 (66%) of children had blood pressures measured at triage. Of these measurements, 153/294 (52%) reflected blood pressures greater than the 90th percentile for age and sex, but only 58/153 (38%) of patients with such blood pressures had a second blood pressure measured. Hospitals varied in their frequency of blood pressure measurement. Adolescents had their blood pressure measured more frequently, 981105 (93%) than two to 12-year-olds, 144/185 (78%) or 1-month to 2-year-olds, 52/147 (35%). Frequency of triage blood pressure measurements in children varied by institution and increased in frequency with age. PMID- 11103730 TI - Pretest probability assessment for selective rest sestamibi scans in stable chest pain patients. AB - The objective of this study was to determine whether pretest probability assessments permit more selective testing of chest pain patients with technetium 99m sestamibi scanning. Pretest probabilities of cardiac ischemia were measured both objectively (Acute Cardiac Ischemia Time-Insensitive Predictive Instrument [ACI-TIPI]) and subjectively (physician's estimate of the probability of unstable angina). Two groups were defined: patients whose postsestamibi scan led to a "downgrade" of the intensity of monitoring and those that resulted in no change in monitoring intensity. Sixty-five patients met study criteria; 25 had a disposition downgrade and 40 had no change. Pretest ACI-TIPI scores were similar in the two groups (29% +/- 18% versus 27% +/- 11%, mean +/- standard deviation; P = .95) as were the physician's assessment of unstable angina (39% +/- 22% versus 40% +/- 24%; P = .75). Objective or subjective pretest probabilities are not significantly different in patients who are likely to have their disposition altered by sestamibi scanning. PMID- 11103731 TI - Provocative testing for chest pain. AB - Since the first Chest Pain Center (CPC) was set up in 1981 to speed up the evaluation and treatment of patients with acute myocardial infarction, the original concept has been expanded to include rapid evaluation of chest pain patients with the appropriate streamlining of care and incorporation of the latest in technology. It has also been established that among patients presenting with acute chest pain, a very low-risk group with less than 5% probability of a coronary event can be identified. The recognition of this group could prevent unnecessary admissions, affording more appropriate patient care and improved cost effectiveness. The efficient management of these chest pain patients requires that there be reductions in: (1) delays in therapy, (2) "soft" admissions, (3) inappropriate dispositions, and (4) cost. With time, provocative testing (PT) for chest pain patients has been brought forward to the frontline. PT methods are now being studied in hundreds of emergency department (ED) patients, followed up over several months to ascertain the predictive value of both positive and negative test results. More and more CPCs are now using PT as part of their management protocol, in terms of decision-making pertaining to prognostification, treatment and disposition. This could be in the form of the ECG graded exercise test (GXT), stress echocardiography (SE) and stress single-photon emission computed tomography (SPECT) radionuclide perfusion imaging. The GXT is fairly widely used currently, SE is gaining popularity and stress radionuclide perfusion imaging will perhaps gain more acceptance as the experience with its use as well as the number of randomized controlled studies increase. As we move into the new millennium, the emergency physicians must familiarize themselves with the latest in the state-of-the-art concepts and technology to render improved, up-to-date and more cost-effective patient care. PMID- 11103732 TI - Pseudo myocardial infarction and pseudo ventricular hypertrophy ECG patterns in Wolff-Parkinson-White syndrome. AB - In Wolff-Parkinson-White (WPW) syndrome, the ventricles are pre-excited through an accessory conduction pathway, bundle of Kent, which directly connects atria with ventricles bypassing the atrioventricular node. The altered sequence of ventricular activation secondary to presence of the bundle of Kent may cause pseudo myocardial infarction and pseudo ventricular hypertrophy patterns on electrocardiogram. The morphology of these pseudo electrocardiographic patterns depends on the anatomical location of the bundle of Kent around the circumference of the atrioventricular ring. Electrocardiograms of the WPW syndrome displaying morphology of different pseudo patterns are presented and the mechanisms causing pseudo patterns are reviewed. PMID- 11103733 TI - Pseudo ventricular hypertrophy and pseudo myocardial infarction in Wolff Parkinson-White syndrome. AB - In Wolff-Parkinson-White syndrome, the sequence of ventricular activation is altered and depending on the anatomic site of the accessory conduction pathway may result in pseudo ventricular hypertrophy and pseudo myocardial infarction patterns on electrocardiogram. The right-sided accessory pathway may direct the depolarization vector towards left amplifying R-wave amplitude in left-sided limb leads simulating left ventricular hypertrophy. The left-sided accessory pathways may give rise to prominent R-waves in right precordial leads simulating right ventricular hypertrophy. The right lateral accessory pathways may simulate anterior infarction because of prominent Q-waves in right precordial leads. The left lateral accessory pathways directing depolarization vector towards right may cause Q-waves in lateral limb-leads simulating high lateral myocardial infarction. In posteroseptal accessory pathway, the ventricular depolarization vector is directed superiorily giving rise to prominent Q-waves in inferior limb leads simulating inferior myocardial infarction. Therefore, ventricular hypertrophy and myocardial infarction should not be diagnosed from the electrocardiograms of Wolff-Parkinson-White syndrome. PMID- 11103734 TI - CT scans essential after posttraumatic loss of consciousness. AB - The frequency of "talk and deteriorate" in the emergency department (ED), subsequent deterioration of patients with seemingly "mild" head injury at the time of presentation, is summarized. Among the 1,073 patients with minor head injury treated in the last 5 years, five patients (0.5%) deteriorated in the ED. All of the five patients had experienced transient loss of consciousness (LOC) before presentation. Deterioration had occurred during treatment of trivial associated injuries in four-fifths of the cases. Computed tomography (CT) scans revealed four acute epidural hematomas and one cerebellar contusion. Retrospectively, immediate brain CT shortly after their arrival may have revealed the presence of traumatic intracranial hematomas before deterioration. Although routine use of CT scans in patients with mild head injury has been controversial, the authors conclude that CT scans should be taken if patients have experienced transient LOC to prevent or reduce the occurrence of deterioration in ED. PMID- 11103735 TI - Mallory-Weiss syndrome with severe bleeding: treatment by endoscopic ligation. AB - There is no consensus as to the best treatment for Mallory-Weiss tears with severe bleeding. Endoscopic ligation is an inexpensive, readily available, and easily learned technique, in contrast to conventional thermocoagulation or hemoclipping. To evaluate the utility of endoscopic ligation, we performed this technique during emergency endoscopy to treat severe bleeding from Mallory-Weiss tears in four patients in our hospital with continuous active bleeding from Mallory-Weiss tears. The patients were all male with an average age of 40.5 years. Symptoms associated with increased intra-abdominal pressure, including retching and vomiting were reported by all patients. The bleeding points were aspirated and controlled by endoscopic ligation, and complete hemostasis was achieved in all cases. We conclude that endoscopic ligation is easy to perform and may provide an alternative treatment for severe bleeding from Mallory-Weiss tears. PMID- 11103736 TI - Elastic adhesive dressing treatment of bleeding wounds in trauma victims. AB - Conventional methods for hemorrhage control in the trauma patient fall short of providing a full solution for the life-threatening bleeding injury. The tourniquet is limited specifically to injuries of the distal limbs. Local pressure or tight bandaging with military bandages is cumbersome and often insufficient. Therefore, we sought a superior method to stop bleeding in emergency situations. Our objective is report and description of our experience with this method. Since 1992 our trauma team repeatedly encountered multiple trauma victims presenting with bleeding wounds. We achieved hemorrhage control by means of an adhesive elastic bandage applied directly over a collection of 4 x 4 gauze pads placed on the wound surface. The roll is then wrapped around the body surface, over the bleeding site, until sufficient pressure is reached to terminate ongoing hemorrhage. Three typical cases are described in detail. Adhesive elastic dressing compression was successful in fully controlling bleeding without compromise of distal blood flow. Our method corresponded to the demand for an immediate, effective and lasting form of hemorrhage control without complications. Furthermore, this technique proved successful even over body surfaces normally recognized as difficult to compress. We experienced equal favorable success while working during transit by either ambulance or helicopter transportation. We find our preliminary experience using elastic adhesive dressing for bleeding control encouraging and suggest that this may substitute existing practices as the selected treatment when indicated. This method is presently underrecognized for this purpose. Development of a single unit bandage may further enhance success in the future. PMID- 11103737 TI - High-pressure injection injuries to the hand. AB - High-pressure injection injury hides the true extent of the lesions behind an apparent small and harmless puncture of the finger or the hand. Through clinical description, we wish to point out the need for prompt treatment to avoid mutilating and function-threatening complications. We wish to outline the role of the emergency physician who must be aware of the incidence of high-pressure injection injury and become accustomed to early referral to a surgeon, experienced in extensive surgical exploration, removal of foreign bodies, and rehabilitation. The open-wound technique gives the best results. We also point out that failure to refer may become an increasing focus of negligence claims. PMID- 11103738 TI - Chest injuries transferred to trauma centers after the 1999 Taiwan earthquake. AB - To better understand the effects of delayed medical care and long transportation times when emergency medical services (EMS) failed after the 1999 Chi-Chi, Taiwan earthquake, we analyzed the patterns and outcomes of patients with chest injuries who were transferred to an unaffected back-up hospital. The medical records of 164 trauma patients who were transferred to Taichung Veterans General Hospital from September 21 to September 24, 1999 were reviewed. Of the 164 patients, 26 (15.9%) had chest injuries. Chest injuries were caused by blunt trauma in all cases. Minor chest injury was noted in 16 patients (61.5%). Mortality developed in two patients, who were transferred after first aid in the field hospital and were in shock status on arrival to emergency department of the back-up hospital. Inadequate resuscitation attributable to insufficient manpower in field hospitals and long transportation times to back-up hospitals are the major problems to be solved in developing disaster plans. For evacuation of overwhelming casualties and for support of medical resources, transportation by helicopter is suggested in aftermath of a large earthquake. PMID- 11103739 TI - Emergency medicine in Ukraine: challenges in the post-Soviet era. AB - The practice of emergency medicine in Ukraine is markedly different from the practice in North America. The emergency physician counterpart in Ukraine attends 6 years of medical school then 18 months of prehospital physician training at an EMS base station. Once trained, prehospital physicians work 160 hours/month in 24 hour shifts at the base station as part of a physician-nurse team which answers ambulance requests. Most patients are seen and treated on site of the ambulance call. Patients are transported to the hospital only 20% of the time. Prehospital physicians can expect to earn $35 to $65 per month. Nearly all prehospital physicians are government employees. Since becoming an independent democratic republic, Ukraine's turbulent economy has negatively affected health care. Deaths from infectious diseases, including vaccine-preventable illnesses are 10-fold to that of Western countries. A 90% income tax discourages the private practice of medicine. Medical care is provided free of charge, however, if a patient wants a higher standard of care, they may have to pay an attending physician up to $200. Most medications used to treat emergencies are free, but if thrombolytics are required, the patient will have to pay for them before they are administered. Budgetary constraints limit equipment and technology. The disparity between urban and rural resources is striking as even the most basic equipment is antiquated and in need of repair. Despite the economic challenges facing Ukranian physicians, they are enthusiastic about the care and services they provide. EMS is well organized and offers services not seen in the United States. Prehospital physicians in Ukraine are viewed as an integral part of the health care system by their hospital-based colleagues. PMID- 11103740 TI - Parkinsonian syndrome after acute carbon monoxide poisoning. PMID- 11103741 TI - Pelvic mass after a sexual assault. PMID- 11103742 TI - Cardiac tamponade secondary to cardiopulmonary resuscitation in a patient receiving antiplatelet therapy. PMID- 11103743 TI - Spontaneous spinal epidural hematoma. PMID- 11103744 TI - Cardiopulmonary arrest secondary to haloperidol. PMID- 11103745 TI - Pediatric respiratory arrest due to a pedunculated laryngeal polyp. PMID- 11103746 TI - Emphysema of the face, neck, and mediastinum from compressed gas barotrauma to the ear. PMID- 11103747 TI - Penetrating abdominal injury caused by nonlethal ammunition. PMID- 11103748 TI - Do caffeine-containing analgesics promote dependence? A review and evaluation. AB - OBJECTIVE: Debates about the suspected association between kidney disease and use of analgesics have led to concern about whether caffeine could stimulate an undesirable overuse of phenacetin-free combined analgesics. A committee was asked to critically review the pertinent literature and to suggest guides for clinical practice and for consideration of international regulatory authorities. PARTICIPANTS: A group of international scientists, jointly selected by the regulatory authorities of Germany, Switzerland, and Austria and the pharmaceutical industry. EVIDENCE: All invited experts evaluated relevant literature and reports and added further information and comments. CONCLUSIONS: Caffeine has a synergistic effectiveness with analgesics. Although caffeine has a dependence potential, the potential is low. Experimental data regarding dependence potential for caffeine alone may not correspond to the conditions in patients with pain. Withdrawal is not likely to cause stimulation or sustainment of analgesic intake. For drug-induced headache, no single or combined analgesic was consistently identified as causative, and no evidence exists for a special role of caffeine. Strong dependence behavior was observed only in patients using phenacetin-containing preparations, coformulated with antipyretics/analgesics and caffeine. This finding may have led to the impression that caffeine stimulates overuse of analgesics. SUMMARY: Although more experimental and long-term data would be desirable to show possible mechanisms of dependence and to offer unequivocal proof of safety, the committee concluded that the available evidence does not support the claim that analgesics coformulated with caffeine, in the absence of phenacetin, stimulate or sustain overuse. PMID- 11103749 TI - Grapefruit-felodipine interaction: effect of unprocessed fruit and probable active ingredients. AB - OBJECTIVES: To determine whether unprocessed grapefruit can cause a drug interaction, whether the active ingredients are naturally occurring, and whether specific furanocoumarins or flavonoids are involved. METHODS: The oral pharmacokinetics of felodipine and its dehydrofelodipine metabolite were determined after administration of felodipine 10 mg extended-release tablet with 250 mL commercial grapefruit juice, homogenized grapefruit segments, or extract of segment-free parts equivalent to one unprocessed fruit or water in a randomized four-way crossover study. Inhibition of recombinant CYP3A4 by furanocoumarins (bergamottin, 6',7'-epoxybergamottin, 6',7'-dihydroxybergamottin) and flavonoids (naringenin optical isomers) was determined. Furanocoumarin and naringenin precursor (naringin) concentrations were measured in each grapefruit treatment. RESULTS: Felodipine AUC with commercial grapefruit juice, grapefruit segments, or grapefruit extract was on average 3-fold higher than that with water. Felodipine peak concentration was higher, but the half-life was unchanged. The dehydrofelodipine/felodipine AUC ratio was reduced. The furanocoumarins produced mechanism-based and competitive inhibition of CYP3A4. Bergamottin was the most potent mechanism-based inhibitor. Naringenin isomers produced only competitive inhibition. Bergamottin, 6',7'-dihydroxybergamottin, and naringin concentrations varied among grapefruit treatments but were sufficient to inhibit markedly in vitro CYP3A4 activity. CONCLUSIONS: Unprocessed grapefruit can cause a drug interaction with felodipine. The active ingredients are naturally occurring in the grapefruit. Bergamottin is likely important in drug interactions with commercial grapefruit juice. 6',7'-Dihydroxybergamottin and naringin may be more important in grapefruit segments because they are present in higher concentrations. Any therapeutic concern for a drug interaction with commercial grapefruit juice should now be extended to include whole fruit and possibly confectioneries made from grapefruit peel. PMID- 11103750 TI - Gender-dependent racial difference in disposition of cyclosporine among healthy African American and white volunteers. AB - Pharmacokinetic studies of intravenous and oral cyclosporine (cyclosporin) were performed in 22 healthy African American (n = 11) and white (n = 11) volunteers. Blood cyclosporine concentrations were measured by high performance liquid chromatography. Concentration versus time data were analyzed by noncompartmental models, and statistical analyses were performed by ANOVA. The clearance of intravenous and oral cyclosporine was 4.3 +/- 0.9 mL/min/kg and 13.5 +/- 4.5 mL/min/kg, respectively, in African Americans and 3.7 +/- 0.5 mL/min/kg and 9.6 mL/min/kg, respectively, in the white volunteers (P = .0001). There was a significant race and gender interaction (P = .038). Bioavailability was lower in African Americans (32.8 +/- 6.6%) compared with white volunteers (39.3 +/- 7.1%; P = .049), with a significant race and gender interaction (P = .048). The dose adjusted area under the curve (AUC) of intravenous and oral cyclosporine was 54.3 +/- 10.6 ng x hr/mL per milligram and 18.1 +/- 4.1 ng x hr/mL per milligram, respectively, in African Americans and 61.9 +/- 6.8 ng x hr/mL per milligram and 24.2 +/- 4.6 ng x hr/mL per milligram, respectively, in white volunteers (P = .023). These findings suggest that disposition of cyclosporine is dependent both on race and on gender. PMID- 11103751 TI - The cytochrome P450 3A4 inhibitor itraconazole markedly increases the plasma concentrations of dexamethasone and enhances its adrenal-suppressant effect. AB - OBJECTIVE: To examine the possible interaction of itraconazole with orally and intravenously administered dexamethasone. METHODS: In a randomized, double-blind, placebo-controlled crossover study with four phases, eight healthy subjects took either 200 mg itraconazole (in two phases) or placebo (in two phases) orally once daily for 4 days. On day 4 each subject received an oral dose of 4.5 mg dexamethasone or an intravenous dose of 5.0 mg dexamethasone sodium phosphate during both itraconazole and placebo phases. Plasma dexamethasone and cortisol concentrations were determined by HPLC up to 71 hours, itraconazole and hydroxyitraconazole up to 23 hours. RESULTS: Itraconazole decreased the systemic clearance of intravenously administered dexamethasone by 68% (P < .001), increased the total area under the plasma dexamethasone concentration-time curve [AUC(0-infinity)] 3.3-fold (P < .001), and prolonged the elimination half-life of dexamethasone 3.2-fold (P < .001). The AUC(0-infinity) of oral dexamethasone was increased 3.7-fold (P < .001), the peak plasma concentration 1.7-fold (P < .001), and the elimination half-life 2.8-fold (P < .001) by itraconazole. The morning plasma cortisol concentrations measured 47 and 71 hours after administration of dexamethasone were substantially lower after exposure to itraconazole than to placebo (P < .001). Accordingly, the adrenal-suppressant effect of dexamethasone was greatly enhanced during the itraconazole phases. CONCLUSIONS: Itraconazole markedly increases the systemic exposure to and effects of dexamethasone. A careful follow-up is recommended when itraconazole or other potent inhibitors of the cytochrome P450 3A4 are added to the drug regimen of patients receiving dexamethasone. PMID- 11103752 TI - Rifampin decreases the plasma concentrations and effects of repaglinide. AB - OBJECTIVE: To study the effects of rifampin (INN, rifampicin) on the pharmacokinetics and pharmacodynamics of repaglinide, a new short-acting antidiabetic drug. METHODS: In a randomized, two-phase crossover study, nine healthy volunteers were given a 5-day pretreatment with 600 mg rifampin or matched placebo once daily. On day 6 a single 0.5-mg dose of repaglinide was administered. Plasma repaglinide and blood glucose concentrations were measured up to 7 hours. RESULTS: Rifampin decreased the total area under the concentration time curve of repaglinide by 57% (P < .001) and the peak plasma repaglinide concentration by 41% (P = .001). The elimination half-life of repaglinide was shortened from 1.5 to 1.1 hours (P < .01). The blood glucose decremental area under the concentration-time curve from 0 to 3 hours was reduced from 0.94 to 0.23 mmol/L x h (P < .05), and the maximum decrease in blood glucose concentration from 1.6 to 1.0 mmol/L (P < .05) by rifampin. CONCLUSIONS: Rifampin considerably decreases the plasma concentrations of repaglinide and also reduces its effects. This interaction is probably caused by induction of the CYP3A4 mediated metabolism of repaglinide. It is probable that the effects of repaglinide are decreased during treatment with rifampin or other potent inducers of CYP3A4, such as carbamazepine, phenytoin, or St John's wort. PMID- 11103753 TI - Effect of angiotensin II infusion with and without angiotensin II type 1 receptor blockade on nitric oxide metabolism and endothelin in human beings: a placebo controlled study in healthy volunteers. AB - BACKGROUND: Angiotensin II has been shown to induce the synthesis of endothelium derived relaxing factor nitric oxide (NO) and endothelin in vitro. In human beings, to our knowledge, no data on NO release in response to angiotensin II and on the influence of angiotensin II type 1 receptor blockade have been published. METHODS: In a placebo-controlled study in nine healthy volunteers, angiotensin II was administered intravenously for 6 hours with and without pretreatment with valsartan, a specific angiotensin II type 1 receptor antagonist. NO (NO2 + NO3) and endothelin plasma concentrations, clearance values for inulin and paraaminohippuric acid and NO (NO2 + NO3) excretion in urine were determined. RESULTS: During angiotensin II infusion NO plasma concentrations remained unaltered compared with placebo after 3 hours: 6.66 +/- 5.49 versus 5.56 +/- 3.09 micromol/L (P = ns) but increased after 6 hours: 18.36 +/- 20.02 versus 7.13 +/- 3.87 micromol/L (P < .04). The same was noted after pretreatment with valsartan: 7.61 +/- 5.69 versus 5.56 +/- 3.09 micromol/L (P= ns) after 3 hours, and 21.70 +/ 11.51 versus 7.13 +/- 3.87 micromol/L (P = .02) after 6 hours. In urine fractional NO excretion decreased after angiotensin II infusion: 0.87 +/- 0.72 versus 0.95 +/- 0.71 (P = .5) during the first 3 hours, and 0.44 +/- 0.39 versus 0.78 +/- 0.43 (P = .01) during the following 3 hours. After valsartan pretreatment the decrease in fractional urinary NO excretion began earlier: 0.40 +/- 0.15 versus 0.95 +/- 0.71 (P = .04) during the first 3 hours, and 0.17 +/- 0.11 versus 0.78 +/- 0.43 (P = .01) during the following 3 hours. Endothelin plasma concentrations showed no difference after angiotensin II infusion with or without valsartan. CONCLUSIONS: Our observations demonstrate for the first time that angiotensin II increases NO plasma concentrations in human beings and that this response is not mediated by angiotensin II type 1 receptor. In spite of increased NO plasma levels, urinary NO excretion decreased. Endothelin plasma levels remained unchanged during angiotensin II infusion. PMID- 11103754 TI - Pharmacokinetic analysis of bioequivalence trials: implications for sex-related issues in clinical pharmacology and biopharmaceutics. AB - OBJECTIVES: To address the questions of whether women should be included in bioequivalence trials and whether dosage adjustment may be needed in women relative to men. METHODS: Sex-related analysis was conducted for 26 bioequivalence studies involving both sexes. A total of 94 data sets [47 each for the areas under the plasma concentration-time curve (AUC) and maximum concentration (Cmax)] were used. ANOVA was performed. Three statistical models were used to estimate population means and intrasubject variability between sexes, as well as sex-by-formulation interactions. Comparisons were made by use of confidence intervals, magnitude of observed differences, and statistical significance (alpha = .05). RESULTS: With some exceptions, intrasubject variabilities were similar for men and women. In about 10% of the data sets (AUC or Cmax), women had significantly higher variability. Although fewer, there were examples with higher variability in men. With a 20 percentage point difference used in the test-over-reference mean ratios between sexes as a signal of sex-by formulation interaction, the frequency of this interaction (AUC or Cmax) is approximately 13% and approximately 35%, counting by data sets and studies, respectively. Mean sex-related differences of > or = 20% in the pharmacokinetic parameters for the reference product were observed in 39% of the data sets (AUC or Cmax). In approximately 28% of the data sets, these differences were statistically significant. The frequency was approximately 15% after body weight correction. CONCLUSIONS: In general, men and women have similar intrasubject variability. Where variability differs between sexes, there is a suggestion that higher variability in women may be more frequent. The data also suggest that a sex-based subject-by-formulation interaction can occur, although the frequency may be low. Sex-related differences in pharmacokinetics are apparent in many drugs studied. Dosage adjustment with body weight may be warranted for drugs that exhibit a steep dose-response curve. Although exploratory, the results of this study support recommendations of the 1993 Food and Drug Administration gender guideline that women not be excluded from bioequivalence trials. PMID- 11103755 TI - Myocardial efficiency during levosimendan infusion in congestive heart failure. AB - AIMS: Levosimendan, a novel calcium-dependent calcium sensitizer of the myocardial contractile proteins, also enhances diastolic relaxation and induces peripheral vasodilation by opening potassium channels. To assess the combined energetical effects of levosimendan infusion in vivo, we performed positron emission tomography in patients with decompensated chronic heart failure. METHODS AND RESULTS: Eight hospitalized patients with New York Heart Association functional class III or IV heart failure received levosimendan or placebo intravenously in a randomized double-blind cross-over study. During steady-state, dynamic positron emission tomography with [11C]acetate was used to assess myocardial oxygen consumption and [15O]H2O to measure myocardial blood flow. Cardiac performance and dimensions were assessed by pulmonary artery catheterization and echocardiography. Compared with healthy volunteers, myocardial oxygen consumption during placebo was elevated in the right ventricle but comparable in the left ventricle. During administration of levosimendan, cardiac output increased by 32% (P = .002) mainly because of higher stroke volume. Coronary, pulmonary, and systemic vascular resistance values were significantly reduced. Mean myocardial blood flow increased from 0.76 to 1.02 mL/min/g (P = .033). Levosimendan was neutral on myocardial oxygen consumption and left ventricular efficiency, but it improved right ventricular mechanical efficiency by 24% (P = .012). CONCLUSIONS: Levosimendan has an energetically favorable short-term profile in the treatment of congestive heart failure. It enhances cardiac output without oxygen wasting, particularly by improving efficiency in the right ventricle. PMID- 11103756 TI - Clinical impact of cyclosporine cellular pharmacodynamics in minimal change nephrotic syndrome. AB - BACKGROUND: Cellular pharmacodynamics of cyclosporine (INN, cyclosporin) is considered to be closely implicated in clinical efficacy of the drug in kidney transplantation and other immunologic disorders. We applied this strategy to patients with minimal change nephrotic syndrome to predict individual clinical efficacy of cyclosporine. METHODS: Drug sensitivity tests were carried out with peripheral blood mononuclear cells from 31 patients with minimal change nephrotic syndrome. The 50% lymphocyte-mitosis inhibition of cyclosporine on in vitro blastogenesis of peripheral blood mononuclear cells stimulated with concanavalin A were estimated, and interpatient variations of 50% lymphocyte-mitosis inhibition were evaluated. The relationship between cyclosporine-50% lymphocyte mitosis inhibition and clinical outcomes indicated a decrease of urinary protein and the period required for complete remission under cyclosporine therapy was examined in 14 patients. We also evaluated the correlation between cyclosporine 50% lymphocyte-mitosis inhibition and interleukin-2 production and percentages of interleukin 2 receptor-positive peripheral blood mononuclear cells in vitro. RESULTS: Cyclosporine 50% lymphocyte-mitosis inhibition on peripheral blood mononuclear cell blastogenesis deviated largely between patients from 0.2 to 86.0 ng/mL. We found a statistically significant negative correlation between cyclosporine-50% lymphocyte-mitosis inhibition in vitro and decreasing rates of urinary protein at 1 week after onset of cyclosporine administration (r = -0.655, P < .02). When we arbitrarily divide the 14 patients who received cyclosporine therapy according to their median 50% lymphocyte-mitosis inhibition of cyclosporine into two groups, that is, a high-sensitivity group (50% lymphocyte mitosis inhibition < 18.1 ng/mL, n = 6) and a low-sensitivity group (50% lymphocytemitosis inhibition > 18.1 ng/mL, n = 8), the period required for complete remission was significantly shorter in the high-sensitivity group (P < .03). The 50% lymphocyte-mitosis inhibition of cyclosporine on interleukin-2 production in culture medium was correlated with 50% lymphocyte-mitosis inhibition of the drug on peripheral blood mononuclear cell blastogenesis (r = 0.806, P < .02). Decreasing rates of interleukin-2R-positive cells by cyclosporine treatment in vitro were negatively correlated with peripheral blood mononuclear cells blastogenesis in the presence of the drug (r = -0.694, P < .02). CONCLUSIONS: Peripheral blood mononuclear cell response to cyclosporine in vitro is closely related to clinical efficacy of the drug in minimal change nephrotic syndrome. Peripheral blood mononuclear cell resistance to cyclosporine was correlated with ability of the cells to express interleukin 2 and interleukin 2R. PMID- 11103757 TI - Developmental changes in pharmacokinetics and pharmacodynamics of warfarin enantiomers in Japanese children. AB - OBJECTIVE: To clarify developmental changes in the pharmacokinetics and dynamics of warfarin enantiomers to establish rational pediatric dosage. METHODS: Plasma concentrations of unbound warfarin enantiomers, vitamin K1 and vitamin K dependent proteins (that is, prothrombin fragments 1+2, protein C, and the protein-induced by vitamin K absence) and international normalized ratio were measured in 38 prepubertal (1 to 11 years), 15 pubertal (12 to 18 years), and 81 adult (37 to 76 years) patients given long-term warfarin therapy. Unbound oral clearance values for warfarin enantiomers and its body weight-, body surface area , and liver weight-normalized values, as well as the pharmacodynamic parameters, were compared among the groups. RESULTS: The prepubertal, pubertal, and adult patients exhibited comparable mean plasma concentrations of unbound warfarin enantiomers for pharmacologically more active (S)-warfarin. Although the unbound oral clearance of (S)-warfarin for the prepubertal patients was significantly (P < .01) less than that for the adult group (346 versus 637 mL/min), the body weight-normalized unbound oral clearance for the prepubertal patients was significantly (P < .01) greater than that for the adults and showed a negative correlation (P < .05) with age. In contrast, no differences were observed in the liver weight-normalized unbound oral clearance for (S)-warfarin between the prepubertal and adult groups. The prepubertal patients showed significantly (P < .01 or .05) lower plasma concentrations of protein C and prothrombin fragments 1+2 and greater international normalized ratio and international normalized ratio/dose than the adults. In contrast, the pubertal patients showed largely similar pharmacokinetic and pharmacodynamic properties to adults. CONCLUSION: Liver weight may be a better parameter than body weight for estimating the warfarin doses for prepubertal patients on the basis of the corresponding adult values. Augmented responses to warfarin in children should also be taken into account for estimating warfarin doses for children. PMID- 11103758 TI - Pegylated interferon-alpha2b: pharmacokinetics, pharmacodynamics, safety, and preliminary efficacy data. Hepatitis C Intervention Therapy Group. AB - AIMS: The objectives of this study were to assess the safety, pharmacokinetic and pharmacodynamic profiles, and antiviral efficacy of pegylated interferon-alpha2b monotherapy in patients with chronic hepatitis C. METHODS: Fifty-eight patients (38 men, 20 women; age range, 25 to 65 years) with compensated chronic hepatitis C were enrolled in this open-label, randomized, active controlled study. Patients received 0.035 to 2.0 microg/kg pegylated interferon-alpha2b subcutaneously weekly or the active control, interferon-alpha2b 3 million IU subcutaneously three times/week, for 24 weeks. Safety and antiviral efficacy assessments were performed during treatment and in a subsequent 4-week follow-up period. Detailed pharmacokinetic assessments were performed at weeks 1 and 4. RESULTS: Pegylated interferon-alpha2b produced dose-related reductions in white blood cells, neutrophils, and platelets, and dose-related increases in oral temperature, serum neopterin, and serum 2'5'-oligoadenylate synthetase activity, which were qualitatively similar to those produced by nonpegylated interferon-alpha2b. Reported adverse events (flu-like symptoms, asthenia) were qualitatively similar in pegylated interferon-alpha2b- and nonpegylated interferon-alpha2b-treated groups. Dose-related antiviral activity, as measured by loss of detectable serum hepatitis C virus RNA (<100 copies/mL), was noted at the end of treatment and after 4 weeks of follow-up. Both pegylated and nonpegylated interferon-alpha2b were rapidly absorbed, with maximal concentrations occurring approximately 8 to 12 hours after dose administration. Pegylated interferon-alpha2b had sustained maximal serum concentrations for 48 to 72 hours after dose administration, whereas nonpegylated interferon-alpha2b concentrations declined rapidly. Volume of distribution for both compounds was similar (approximately 1 L/kg). Pegylated interferon-alpha2b elimination half-life was approximately 10-fold greater, and mean apparent clearance was one tenth that of nonpegylated interferon-alpha2b. CONCLUSIONS: Pegylated and nonpegylated interferon-alpha2b safety and pharmacodynamic profiles were comparable. Pegylated interferon-alpha2b demonstrated delayed clearance compared with nonpegylated interferon-alpha2b, consistent with once-weekly administration. PMID- 11103759 TI - Prediction of the outcome of a phase 3 clinical trial of an antischizophrenic agent (quetiapine fumarate) by simulation with a population pharmacokinetic and pharmacodynamic model. AB - A completed phase 3 trial result was simulated 100 times on the basis of a simulation model of quetiapine fumarate (Seroquel), an antischizophrenic agent. The simulation was executed by analysts who were completely blinded from results of the actual trial until after the simulations were submitted to the holder of the trial results. Data from two clinical investigations of quetiapine in patients with schizophrenia were analyzed by use of nonlinear mixed effects modeling to derive a population pharmacokinetic- and pharmacodynamic-based simulation model. The time course of quetiapine concentrations was described by use of a one-compartment open linear pharmacokinetic model with first-order absorption and elimination. The combination of an inhibitory maximum effect pharmacodynamic model for the active treatment effect and a linear function of time for the placebo effect characterized the observed time course of change in the Brief Psychiatric Rating Scale. Simulation results were compared with those in the actual trial to evaluate how well the simulations predicted the outcome. The actual trial results for all doses except the placebo group fell within the predicted Brief Psychiatric Rating Scale scores +/- 1 SE. Unlike the phase 2 trial, from which the pharmacokinetic/pharmacodynamic model was developed, the placebo group in the actual phase 3 trial showed deterioration of Brief Psychiatric Rating Scale scores with time. We conclude that variable placebo responses observed in short-term studies of schizophrenia provide an inadequate basis for the modeling and simulation of placebo subjects in clinical trials. Knowledge of the range of placebo response observed in other studies may have provided an improved basis for the placebo effect model. The model for active drug produced adequate predictions of the actual trial outcomes. PMID- 11103760 TI - Calcaneal fractures in the industrial patient. AB - A retrospective review was performed on industrial patients who sustained calcaneal fractures within the State of Idaho during the years 1992 to 1994, and these patients were insured by the Idaho State Insurance Fund. Of 48 calcaneal fractures that occurred during this period, 18 were non-displaced extra-articular fractures and 30 were displaced intra-articular fractures. An independent evaluator contacted each patient and performed chart reviews regarding the work history, period of time off work, and cost incurred with the injury. A total of 24 primary surgical procedures were performed on patients who sustained a displaced intra-articular calcaneal fracture and 31 secondary procedures were performed including wound debridement, hardware removal, skin grafting, and secondary subtalar fusion. For patients whose calcaneal fractures could be treated with non-operative care, the average time from injury until return to work was 18 weeks, and the average total cost of injury was $14,230. For patients whose calcaneal fractures required open reduction and internal fixation, the average time loss from work was 35 weeks, and the average total cost of injury was $31,004. Seven patients whose calcaneal fractures were initially treated with an open reduction, internal fixation later underwent a hindfoot arthrodesis. The average time off work for these patients was 69 weeks and the average total cost of injury was $65,384. Fractures were rated on postoperative radiographs according to the quality of their operative reduction. Fractures that were non anatomically reduced had an increased tendency to require a subtalar fusion. Nine patients sustained other injuries associated with their calcaneal fracture and three patients sustained an open fracture. Both concurrent injuries and open fractures were associated with increased total cost and increased time off work. The total cost of injury was doubled as was time off work when an open reduction and internal fixation was followed later by a secondary subtalar arthrodesis. PMID- 11103761 TI - Outcomes in hallux rigidus patients treated nonoperatively: a long-term follow-up study. AB - The purpose of this study was to analyze radiographic outcome and patient satisfaction in non-operative care of hallux rigidus. Twenty-two patients representing 24 feet were surveyed and radiographed. Average follow-up was 14.4 years (range, 12-19 years). In 75% (18/24) of the feet, the patients would "still chose not to have surgery" if they had to make the decision again. The pain remained about the same in 22 feet, improved with time in one, and became worse in one. The most common reason given for not having surgery was that the pain was not severe enough. The most common type of self-care was a shoe with an "ample toe box." More patients benefited from a stiff sole than a soft sole, but the majority of patients did not cite the sole of the shoe as being important. There was measurable loss of cartilage space radiographically over time in 16 of 24 feet, and in eight of the 16 feet, the loss of cartilage space was dramatic. The majority of hallux rigidus patients rated their pain as staying the same over a twelve-year period, despite significant deterioration of joint space noted radiographically. PMID- 11103762 TI - Treatment of sesamoid disorders with a rocker sole shoe modification. PMID- 11103763 TI - Current practice patterns in the treatment of Charcot foot. AB - Treatment of Charcot foot osteoarthropathy has emerged as a major component of the American Orthopaedic Foot and Ankle Society (AOFAS) Diabetes 2000 Initiative. A two-part survey described treatment patterns and current footwear use of patients with Charcot osteoarthropathy of the foot and ankle. In the first part, 94 consecutive patients with a history of Charcot foot and ankle presenting for care were questioned on their foot-specific treatment and current footwear use. A history of diabetic foot ulcer was given by 39 (41%) patients, and an infection had been present in a foot of 20 (21%) patients. The initial treatment of the Charcot foot and ankle had been a total contact cast in 46 (49%) patients, and a pre-fabricated walking boot in 19 (20%). Charcot related surgery had consisted of 76 procedures in 46 (49%) patients. Sixty-three (67%) patients were currently using accommodative footwear (depth-inlay shoes in 46 [49%], custom shoes in 10 [11%], and CROW in 7 [7%] patients), and 72 (77%) were currently using custom accommodative foot orthoses. The second part of this study consisted of a questionnaire completed by 37 orthopaedic surgeons (members of AOFAS) interested in forming a Charcot Study Group. They treated an average of 11.8 patients having Charcot foot or ankle per month. Thirty (81%) used the Semmes-Weinstein 5.07 monofilament as a screening tool for peripheral neuropathy. For treatment of Eichenholtz Stage I, 29 (78%) used a total contact cast and 15 (41%) allowed weightbearing; for Stage II, 30 (81%) physicians used a total contact cast and 18 (49%) allowed weightbearing. Although the literature contains uniform recommendations for immobilization and non-weightbearing as treatment for the initial phases of Charcot arthropathy, the results of this benchmarking study reveal that currenl treatment is varied. PMID- 11103764 TI - Hindfoot motion after isolated and combined arthrodeses: measurements in anatomic specimens. AB - Passive motions at the subtalar joint, talonavicular joint and calcaneocuboid joint were measured in eight ankle specimens, using an ultrasonic motion analysis system. Arthrodeses of the three joints were performed in all feasible combinations and the resulting motion change at the unfused joints was determined. Motion at the subtalar joint was not significantly affected by fusion of the calcaneocuboid joint, reduced to one quarter by fusion of the talonavicular and calcaneocuboid joints (double arthrodesis) and almost completely eliminated with all other fusions. Motion at the talonavicular joint was not significantly affected by calcaneocuboid fusion and reduced to approximately one third with the subtalar and the double arthrodesis. Motion at the calcaneocuboid joint was not significantly reduced by subtalar fusion but almost completely eliminated in all fusions involving the talonavicular joint. It is concluded that the talonavicular joint is the key articulation for hindfoot motion. Double arthrodesis preserved significant motion at the subtalar joint. Fusion of the calcaneocuboid joint had no significant influence on remaining hindfoot motion. PMID- 11103765 TI - Quantitative assessment of simultaneous capacitive and resistive plantar pressure measurements during walking. AB - Plantar pressure data were collected simultaneously, during walking, from capacitive and resistive in-shoe pressure measurement systems. Overall mean peak pressure recordings from the resistive system were 32%, 20% and 14% greater than recordings from the capacitive system, at the heel, central metatarsal heads, and great toe, respectively. Placement of one system's insoles above or below the other's somewhat affected peak pressure measurements from both systems, while calibration via air bladder or single limb standing techniques somewhat affected resistive measurements as well. Capacitive measurement variability was 60%, 20% and 22% lower than resistive measurement variability, at the heel, central metatarsal heads, and great toe, respectively. Both systems tended to exhibit greater variability when capacitive insoles were placed above resistive insoles; however, the effects on variability of the experimental insole arrangements were well overshadowed by the overall variability differences between systems. PMID- 11103766 TI - Dynamic pedobarography (DPB) in operative management of cavovarus foot deformity. AB - Dynamic pedobarography (DPB) was performed in 21 patients, 9 male and 12 female with cavovarus foot deformity mostly of Charcot-Marie-Tooth origin. Age ranged from 14 to 52 years (mean 30 y). Twenty-six feet were examined pre- and postoperatively clinically, radiologically and by dynamic pedobarography with a follow-up time from 9 to 49 months (mean 22.5 mo). The EMED SF system was used for data collection during walking. Gait line, contact areas (CA), peak pressures (PP) and pressure time integral (PTI) were determined. According to the contact pattern the examined feet could be divided into three groups with antegrade, retrograde and inversion contact pattern. Data analysis showed postoperatively considerable increase in CA and decrease in PP and PTI. Clinical results such as plantar callosities and "roll over avoidance gait" did not always correlate with pedobarographic data. DPB adds a dynamic component in the diagnosis and management of cavovarus feet but certain limitations exist. PMID- 11103767 TI - Accessory soleus muscle as a cause of resistance to correction in congenital club foot: a case report. AB - A 14-month-old female with bilateral clubfeet was initially treated by serial casting and percutaneous tenotomy of the Achilles tendon, bilaterally. Both clubfeet subsequently underwent surgical treatment with a posteromedial release through a Cincinnati incision. At surgery on one clubfoot, an accessory Soleus muscle was found anterior to the Achilles tendon with a distinct insertion on the upper surface of calcaneus, anterior and medial to the insertion of Achilles tendon. This accessory Soleus muscle may have been the cause of resistance to correction in this congenital clubfoot. PMID- 11103768 TI - Simultaneous arthrodesis of the metatarsophalangeal and interphalangeal joints of the hallux. PMID- 11103769 TI - The diabetic plantar hallux ulcer: a curative soft tissue procedure. PMID- 11103770 TI - Highlights of the Sixteenth Annual Summer Meeting of the American Orthopaedic Foot and Ankle Society, Vail, Colorado, July 13-15, 2000. PMID- 11103771 TI - Psychogenic equinovarus. PMID- 11103772 TI - Role of metatarsus primus elevatus in the pathogenesis of hallux rigidus. PMID- 11103773 TI - Duplication of the mutant RET allele in trisomy 10 or loss of the wild-type allele in multiple endocrine neoplasia type 2-associated pheochromocytomas. AB - Inherited mutations of the RET proto-oncogene are tumorigenic in patients with multiple endocrine neoplasia type 2 (MEN 2). However, it is not understood why only few of the affected cells in the target organs develop into tumors. Genetic analysis of nine pheochromocytomas from five unrelated patients with MEN 2 showed either duplication of the mutant RET allele in trisomy 10 or loss of the wild type RET allele. Our results suggest a "second hit" causing a dominant effect of the mutant RET allele, through either duplication of the mutant allele or loss of the wild-type allele, as a possible mechanism for pheochromocytoma tumorigenesis in patients with MEN 2. PMID- 11103774 TI - t(9;11)(p22;p15) in acute myeloid leukemia results in a fusion between NUP98 and the gene encoding transcriptional coactivators p52 and p75-lens epithelium derived growth factor (LEDGF). AB - A t(9;11)(p22;p15) chromosomal translocation was identified in an adult patient with de novo acute myelogenous leukemia. Fluorescence in situ hybridization and Southern blot analysis mapped the 11p15 break-point to the NUP98 gene. Using 3' rapid amplification of cDNA ends, we have identified a chimeric mRNA that fused the NUP98 FXFG repeats in frame to the COOH-terminal portion of the gene encoding the transcriptional coactivators p52 and p75, also known as lens epithelium derived growth factor (LEDGF). As expected, both NUP98-p52 and NUP98-p75 (LEDGF) chimeric mRNAs were detected by reverse transcription-PCR; however, the reciprocal p52/p75 (LEDGF)-NUP98 fusion mRNA was not detected. Our results demonstrate that this is the most 5' NUP98 fusion reported and reveal a previously unrecognized genetic target, the gene encoding p52/p75 (LEDGF). PMID- 11103775 TI - Frequent amplification of the telomerase reverse transcriptase gene in human tumors. AB - Activation of telomerase is a crucial step during cellular immortalization and malignant transformation of human cells and requires the induction of the catalytic component, human telomerase reverse transcriptase (hTERT), encoded by the hTERT gene. It is poorly understood how the hTERT gene is activated in human cancer cells. In the present study, we examined the hTERT gene copy number in human cancer cell lines and in primary tumor tissues. Amplification of the hTERT gene was observed in 8 of 26 (31%) tumor cell lines and 17 of 58 (30%) primary tumors examined (8 of 21 lung tumors, 3 of 10 cervical tumors, 5 of 19 breast carcinomas, and 1 of 8 neuroblastomas). In addition, 13 of 26 (50%) cell lines and 13 of 58 (22%) primary tumors displayed gain of hTERT gene copies with 3-4 copies/cell. The present findings imply that the hTERT locus may be a frequent target for amplification during tumorigenesis and that this genetic event probably contributes to the dysregulation of telomerase activity occurring in human tumors. PMID- 11103776 TI - TMS1, a novel proapoptotic caspase recruitment domain protein, is a target of methylation-induced gene silencing in human breast cancers. AB - Gene silencing associated with aberrant methylation of promoter region CpG islands is an acquired epigenetic alteration that serves as an alternative to genetic defects in the inactivation of tumor suppressor and other genes in human cancers. The hypothesis that aberrant methylation plays a direct causal role in carcinogenesis hinges on the question of whether aberrant methylation is sufficient to drive gene silencing. To identify downstream targets of methylation induced gene silencing, we used a human cell model in which aberrant CpG island methylation is induced by ectopic expression of DNA methyltransferase. Here we report the isolation and characterization of TMS1 (target of methylation-induced silencing), a novel CpG island-associated gene that becomes hypermethylated and silenced in cells overexpressing DNA cytosine-5-methyltransferase-1. We also show that TMS1 is aberrantly methylated and silenced in human breast cancer cells. Forty percent (11 of 27) of primary breast tumors exhibited aberrant methylation of TMS1. TMS1 is localized to chromosome 16p11.2-12.1 and encodes a 22-kDa predicted protein containing a COOH-terminal caspase recruitment domain, a recently described protein interaction motif found in apoptotic signaling molecules. Ectopic expression of TMS1 induced apoptosis in 293 cells and inhibited the survival of human breast cancer cells. The data suggest that methylation-mediated silencing of TMS1 confers a survival advantage by allowing cells to escape from apoptosis, supporting a new role for aberrant methylation in breast tumorigenesis. PMID- 11103777 TI - Activation of a caspase-9-mediated apoptotic pathway by subcellular redistribution of the novel caspase recruitment domain protein TMS1. AB - Genetic and epigenetic alterations affecting proteins involved in apoptosis can contribute to the establishment and progression of cancer. Recently, our laboratory has isolated a novel gene, TMS1, that is aberrantly methylated and silenced in a significant proportion of human breast cancers. TMS1 contains a caspase recruitment domain (CARD), suggesting a role in caspase-mediated cell death. In the present study, we characterize the participation of TMS1 in apoptosis and examine the subcellular localization of the protein. Inducible expression of TMS1 inhibited cellular proliferation and induced DNA fragmentation in a time-dependent manner. These apoptotic events were blocked by the general caspase inhibitor, Z-VAD-fmk. The ability of TMS1 to trigger apoptosis was also suppressed by a dominant negative form of caspase-9 but not by a dominant negative form of caspase-8, indicating that TMS1 functions through activation of caspase-9. Unlike a number of other CARD-containing proteins, TMS1 did not activate nuclear factor kappaB-dependent transcription, consistent with a proapoptotic role for TMS1 in death signaling pathways. Timed localization studies revealed that TMS1-induced apoptosis was accompanied by the redistribution of TMS1 from the cytoplasm to perinuclear spherical structures. Whereas the apoptotic activity of TMS1 was blocked by caspase inhibition, the formation of TMS1-containing subcellular structures was not, suggesting that the redistribution of TMS1 precedes caspase activation. Both the proapoptotic activity of TMS1 and aggregate formation were dependent on the CARD. In summary, the data indicate that TMS1-induced apoptosis proceeds through a CARD-dependent aggregation step followed by activation of a caspase-9-mediated pathway. PMID- 11103778 TI - Vascular endothelial growth factor (VEGF) modulation by targeting hypoxia inducible factor-1alpha--> hypoxia response element--> VEGF cascade differentially regulates vascular response and growth rate in tumors. AB - Although tumors can activate vascular endothelial growth factor (VEGF) promoter in host stromal cells, the relative contribution to VEGF production of host versus tumor cells and the resulting vascular response have not been quantitated to date. To this end, we implanted VEGF-/- and wild-type (WT) embryonic stem (ES) cells in transparent dorsal skin windows in severe combined immunodeficient mice. VEGF-/- ES cell-derived tumors produced approximately 50% of VEGF compared with the WT tumors, suggesting significant contribution of host stromal cells. To discern the hypoxia-induced hypoxia inducible factor (HIF)-1alpha --> hypoxia response element (HRE) --> VEGF signaling cascade, we also examined tumors derived from HIF-1alpha-/- and HRE-/- ES cells. As expected, the VEGF protein level in HIF-1alpha-/- ES tumors was intermediate between VEGF-/- and WT ES cell tumors. Surprisingly, HRE-/- ES tumors produced the same level of VEGF as the VEGF-/- ES tumors, suggesting a critical role of HRE in tumor cell VEGF production. Angiogenesis in these tumors was proportional to their VEGF levels (VEGF-/- approximate to HRE-/- < HIF-1alpha-/- < WT). In contrast, vascular permeability, leukocyte-endothelial adhesion, and tumor growth were reduced in VEGF-/- and HRE-/- tumors but were comparable in HIF-1a-/- and WT tumors. This discrepancy suggests that different intracellular signaling pathways may be involved in each of these functions of VEGF. More importantly, these data suggest that host cells are active players in tumor angiogenesis and growth and need to be taken into account in the design of any therapeutic strategy. PMID- 11103779 TI - Complete inhibition of rhabdomyosarcoma xenograft growth and neovascularization requires blockade of both tumor and host vascular endothelial growth factor. AB - Growth of the human rhabdomyosarcoma A673 cell line in nude mice is substantially reduced but not completely suppressed after systemic administration of the antihuman vascular endothelial growth factor (VEGF) monoclonal antibody (Mab) A.4.6.1. Potentially, such escape might be attributable to incomplete local penetration of the antibody because of a diffusion barrier associated with tumor growth. Alternatively, it might reflect a compensatory up-regulation of murine VEGF, produced by the stroma of the host, or of other angiogenic factor genes. To test these potential mechanisms, systemic administration of Mab A.4.6.1, was performed in conjunction with intratumoral administration of an irrelevant antibody, an antihuman VEGF Fab or mFlt(1-3)-IgG that neutralizes both human and murine VEGF. Tumor growth in the systemic-plus-intratumoral anti-VEGF group was not different from that in the systemic anti-VEGF-plus-intratumoral-control antibody group, arguing against the possibility that bioavailability is the factor that limits the antitumor efficacy of Mab A.4.6.1. However, intratumoral mFlt(l-3)-IgG administration dramatically enhanced the activity of systemic anti VEGF Mab and resulted in complete suppression of tumor growth, which indicated that host VEGF significantly contributes to tumor growth. Systemic administration of mFlt(1-3)-IgG alone replicated these findings. Histological analysis of residual tumor tissues revealed an almost complete absence of host-derived vasculature and massive tumor-cell necrosis in the mFlt(1-3)-IgG groups. Such extensive necrotic areas were not present in the other groups. Real-time reverse transcription-PCR analysis of total RNA derived from tumor tissues indicated strong up-regulation of both human and murine VEGF as well as other genes regulated by hypoxia. Our findings emphasize the need to completely block VEGF for maximal inhibition of tumor growth. PMID- 11103780 TI - The caspase 9 inhibitor Z-LEHD-FMK protects human liver cells while permitting death of cancer cells exposed to tumor necrosis factor-related apoptosis-inducing ligand. AB - Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a potent inducer of apoptosis of transformed and cancer cells but not of most normal cells. Recent studies have revealed an unforeseen toxicity of TRAIL toward normal human hepatocytes, thereby bringing into question the safety of systemic administration of TRAIL in humans with cancer. We found that SW480 colon adenocarcinoma, or H460 non-small cell lung cancer cell lines, which are sensitive to TRAIL, were not protected by the caspase 9 inhibitor Z-LEHD-FMK from TRAIL-induced apoptosis. However, a human colon cancer cell line HCT116 and a human embryonic kidney cell line 293, which are sensitive to TRAIL, were protected by Z-LEHD-FMK from TRAIL-mediated death. Both HCT116 and SW480 cells were protected from TRAIL by the caspase 8 inhibitor Z-IETD-FMK, dominant negative FADD and cellular FLIP-s and interestingly both cell lines displayed caspase 9 cleavage to a similar extent after TRAIL exposure. We confirmed that normal human liver cells are sensitive to TRAIL. Moreover, we found that normal human liver cells could be protected from TRAIL-induced apoptosis by simultaneous exposure to Z-LEHD-FMK. A similar brief exposure to TRAIL plus Z-LEHD-FMK inhibited colony growth of SW480 but not HCT116 cells. Because some cancer cell lines are not protected from TRAIL-mediated killing by Z-LEHD-FMK, we believe that a brief period of caspase 9 inhibition during TRAIL administration may widen the therapeutic window and allow cancer cell killing while protecting normal liver cells. This strategy could be further developed in the effort to advance TRAIL into clinical trials. PMID- 11103781 TI - Altered expression of estrogen receptor coregulators during human breast tumorigenesis. AB - The hypothesis that altered expression of specific coactivators/repressors of the estrogen receptor occurs during human breast tumorigenesis in vivo is examined in this study. Using in situ hybridization and reverse transcription-PCR assays, the expression of two coactivators (SRA and AIB1) and one repressor (REA) of the estrogen receptor was compared between matched breast tumors and adjacent normal human breast tissue. The levels of SRA and AIB1 mRNA were increased in tumors compared with normal tissues (n = 19; Wilcoxon matched pairs test; P < 0.01). In contrast, the expression of REA mRNA was not different between tumors and normal tissues (n = 19; Wilcoxon; P = 0.110). The ratios of AIB1:REA and SRA:REA were higher (Wilcoxon; P < 0.05) in tumors compared with normal tissues. Furthermore, SRA:AIB1 was higher (Wilcoxon; P = 0.0058) in tumors compared with normal tissues. Although our study is small, these data are consistent with the above hypothesis and suggest that such alterations may have a role in the altered estrogen action occurring during breast tumorigenesis. PMID- 11103782 TI - A MAGE-A3 peptide presented by HLA-DP4 is recognized on tumor cells by CD4+ cytolytic T lymphocytes. AB - Antigens encoded by MAGE-A3 and recognized by T cells are interesting targets for tumor immunotherapy because they are strictly tumor specific and shared by many tumors of various histological types. A number of MAGE-A3 antigenic peptides presented by HLA class I molecules have been used in clinical trials, and regressions of melanoma metastasis have been observed. We report here the identification of a MAGE-A3 epitope, TQHFVQENYLEY, presented to CD4+ T lymphocytes by HLA-DP4 molecules, which are expressed in approximately 76% of Caucasians. This new epitope may be useful both for therapeutic vaccination and for the evaluation of the immune response in cancer patients. Interest ingly, the CD4+ T cells lysed HLA-DP4 tumor cells expressing MAGE-A3, indicating that this epitope, in contrast to other class-II MAGE-A3 epitopes, is presented at the surface of tumor cells. The study of this disparity in the presentation of two epitopes from the same protein may lead to a better understanding of the endogenous class II presentation pathway. PMID- 11103783 TI - Activation of the transcription factor Oct-1 in response to DNA damage. AB - Mammalian cells exhibit complex cellular responses to genotoxic stress, including cell cycle checkpoint, DNA repair, and apoptosis. Inactivation of these important biological events will result in genomic instability and cell transformation. It has been demonstrated that gene activation is a critical initial step during the cellular response to DNA damage. A number of investigations have shown that transcription factors are involved in the regulation of stress-inducible genes. These transcription factors include p53, c-Myc, and AP-1 (c-fos and c-jun). However, the role for the octamer-binding transcription factor Oct-1 in the DNA damage-activated response is unknown. In this report, we have presented the novel observation that the transcription factor Oct-1 is induced after cells are exposed to multiple DNA-damaging agents and therapeutic agents, including UV radiation, methylmethane sulfonate, ionizing radiation, etoposide, cisplatin, and camptothecin. The induction of the Oct-1 protein is mediated through a posttranscriptional mechanism and does not require the normal cellular function of the tumor suppressor p53, indicating that the Oct-1 protein, as a transcription factor, may play a role in p53-independent gene activation. In addition to increased protein level, the activity of Oct-1 DNA binding to its specific consensus sequence is also enhanced by DNA damage. Therefore, these results have implicated that the transcription factor Oct-1 might participate in cellular response to DNA damage, particularly in p53-independent gene activation. PMID- 11103784 TI - Large-scale serial analysis of gene expression reveals genes differentially expressed in ovarian cancer. AB - Difficulties in the detection, diagnosis, and treatment of ovarian cancer result in an overall low survival rate of women with this disease. A better understanding of the pathways involved in ovarian tumorigenesis will likely provide new targets for early and effective intervention. Here, we have used serial analysis of gene expression (SAGE) to generate global gene expression profiles from various ovarian cell lines and tissues, including primary cancers, ovarian surface epithelia cells, and cystadenoma cells. The profiles were used to compare overall patterns of gene expression and to identify differentially expressed genes. We have sequenced a total of 385,000 tags, yielding >56,000 genes expressed in 10 different libraries derived from ovarian tissues. In general, ovarian cancer cell lines showed relatively high levels of similarity to libraries from other cancer cell lines, regardless of the tissue of origin (ovarian or colon), indicating that these lines had lost many of their tissue specific expression patterns. In contrast, immortalized ovarian surface epithelia and ovarian cystadenoma cells showed much higher similarity to primary ovarian carcinomas than to primary colon carcinomas. Primary tissue specimens therefore appeared to be a better model for gene expression analyses. Using the expression profiles described above and stringent selection criteria, we have identified a number of genes highly differentially expressed between nontransformed ovarian epithelia and ovarian carcinomas. Some of the genes identified are already known to be overexpressed in ovarian cancer, but several represent novel candidates. Many of the genes up-regulated in ovarian cancer represent surface or secreted proteins such as claudin-3 and -4, HE4, mucin-1, epithelial cellular adhesion molecule, and mesothelin. Interestingly, both apolipoprotein E (ApoE) and ApoJ, two proteins involved in lipid homeostasis, are among the genes highly up regulated in ovarian cancer. Selected serial analysis of gene expression results were further validated through immunohistochemical analysis of ApoJ, claudin-3, claudin-4, and epithelial cellular adhesion molecule in archival material. These experiments provided additional evidence of the relevance of our findings in vivo. The publicly available expression data reported here should stimulate and aid further research in the field of ovarian cancer. PMID- 11103785 TI - Frequent association of beta-catenin and WT1 mutations in Wilms tumors. AB - The etiology of Wilms tumor, an embryonic kidney tumor, is genetically heterogeneous. One Wilms tumor gene, WT1, which encodes a zinc finger transcription factor, is mutated in 10-20% of Wilms tumors, but it is still not clear what critical cellular pathway(s) is affected by these mutations. Recently beta-catenin mutations have been reported in 6 of 40 (15%) of Wilms tumors. Beta catenin is the central effector in the Wnt signal transduction pathway, and deregulation of beta-catenin signaling is critical in the development of a number of malignancies. The observation of beta-catenin mutations in Wilms tumors suggests that abrogation of the Wnt signaling pathway also plays a role in some Wilms tumors. To assess the relationship of WT1 mutations vis-a-vis beta-catenin mutations in Wilms tumor, we analyzed 153 primary tumors, and 21 of 153 (14%) carried beta-catenin mutations. Surprisingly, we observed a highly significant (P = 3.6 x 10(-13)) association between WT1 and beta-catenin mutations; 19 of 20 beta-catenin-mutant tumors had also sustained WT1 mutations. By analogy to the patterns of concordant and discordant gene mutations observed in other tumors, our data suggest that mutation of WT1 and beta-catenin affects two different cellular pathways, both of which are critically altered in at least a subset of Wilms tumors. PMID- 11103786 TI - Loss of annexin 1 correlates with early onset of tumorigenesis in esophageal and prostate carcinoma. AB - Annexin I protein expression was evaluated in patient-matched longitudinal study sets of laser capture microdissected normal, premalignant, and invasive epithelium from human esophageal squamous cell cancer and prostatic adenocarcinoma. In 25 esophageal cases (20 by Western blot and 5 by immunohistochemistry) and 17 prostate cases (3 by Western blot and 14 by immunohistochemistry), both tumor types showed either complete loss or a dramatic reduction in the level of annexin I protein expression compared with patient matched normal epithelium (P < or = 0.05). Moreover, by using Western blot analysis of laser capture microdissected, patient-matched longitudinal study sets of both tumor types, the loss of protein expression occurred in premalignant lesions. Concordance of this result with immunohistochemical analysis suggests that annexin I may be an essential component for maintenance of the normal epithelial phenotype. Additional studies investigating the mechanism(s) and functional consequences of annexin I protein loss in tumor cells are warranted. PMID- 11103787 TI - Thymidine phosphorylase induces carcinoma cell oxidative stress and promotes secretion of angiogenic factors. AB - Thymidine phosphorylase (TP) (E.C. 2.4.2.4), also known as platelet-derived endothelial cell growth factor, is a potent angiogenic factor. The expression of TP correlates with poor prognosis in a range of tumor types. 2-Deoxy-D-ribose-1 phosphate, a product of thymidine catabolism by TP, is a strongly reducing sugar that generates oxygen radical species during the early stages of protein glycation. We show that thymidine induces oxidative stress in TP-overexpressing carcinoma cells, promoting secretion of the stress-induced angiogenic factors vascular endothelial growth factor and interleukin-8, and inducing matrix metalloproteinase-1. Our findings outline a putative mechanism for TP-induced angiogenesis and identify novel targets for intervention. PMID- 11103788 TI - The oncogenic potential of the high mobility group box protein Sox3. AB - Sox proteins belong to the superfamily of high mobility group (HMG) proteins. Sox3 is expressed predominantly in the immature neuroepithelium. Ectopic expression of Sox3 causes oncogenic transformation of chicken embryo fibroblasts (CEFs). The oncogenicity of Sox3 is correlated with nuclear localization and transcriptional regulatory activity; mutants containing deletions in the HMG box or the transactivation domain fail to induce foci of transformation. These observations suggest that Sox proteins can induce aberrant cell growth and strengthen the link of HMG proteins to oncogenesis. PMID- 11103789 TI - O6-benzylguanine potentiates the antitumor effect of locally delivered carmustine against an intracranial rat glioma. AB - Local delivery of carmustine (BCNU) via biodegradable polymers prolongs survival against experimental brain tumors and in human clinical trials. O6-benzylguanine (O6-BG), a potent inhibitor of the DNA repair protein, O6-alkylguanine-DNA alkyltransferase (AGT), has been shown to reduce nitrosourea resistance and, thus, enhance the efficacy of systemic BCNU therapy in a variety of tumor models. In this report, we demonstrate that O6-BG can potentiate the activity of BCNU delivered intracranially via polymers in rats challenged with a lethal brain tumor. Fischer 344 rats received a lethal intracranial challenge of 100,000 F98 glioma cells (F98 cells have significant AGT activity, 328 fmol/mg protein). Five days later, animals receiving an i.p. injection of O6-BG (50 mg/kg) 2 h prior to BCNU polymer (3.8% BCNU by weight) implantation had significantly improved survival (n = 7; median survival, 34 days) over animals receiving either O6-BG alone (n = 7; median survival, 22 days; P = 0.0002) or BCNU polymer alone (n = 8; median survival, 25 days; P = 0.0001). Median survival for the control group (n = 8) was 23.5 days. Moreover, there was no physical, behavioral, or pathological evidence of treatment-related toxicity. These findings suggest that O6-BG can potentiate the effects of interstitially delivered BCNU and, for tumors expressing significant AGT, may be necessary for the BCNU to provide a meaningful therapeutic benefit. Given the clinical use of BCNU polymers against malignant gliomas, concurrent treatment with O6-BG may provide an important addition to our therapeutic armamentarium. PMID- 11103790 TI - Somatic mutations of LKB1 and beta-catenin genes in gastrointestinal polyps from patients with Peutz-Jeghers syndrome. AB - Peutz-Jeghers syndrome (PJS) is characterized by multiple gastrointestinal hamartomatous polyps, mucocutaneous melanin deposition, and increased risk of cancer, mainly in the gastrointestinal tract. We examined mutations of the LKB1, beta-catenin, APC, K-ras, and p53 genes in 27 gastrointestinal hamartomatous polyps from 10 patients in nine PJS families. Of these hamartomatous polyps, one intestinal polyp had an adenomatous lesion, and one gastric polyp contained adenomatous and carcinomatous lesions. Germ-line mutations of the LKB1 gene were detected in six PJS families. Somatic mutations of the LKB1 gene were found in 5 polyps, whereas loss of heterozygosity (LOH) at the LKB1 locus at 19p was seen in 14 other polyps. In adenomatous lesions microdissected from hamartomatous polyps, both beta-catenin mutation and 19p LOH were detected. Furthermore, a carcinomatous lesion in a gastric hamartomatous polyp was found to contain a mutation of the p53 gene and LOH at the p53 locus in addition to LOH at the LKB1 locus and a beta-catenin mutation. K-ras mutations were detected in a few polyps, whereas no APC mutation or 5q LOH was detected in hamartomatous polyps. These results suggest that gastrointestinal hamartomatous polyps in PJS patients develop through inactivation of the LKB1 gene by germ-line mutation plus somatic mutation or LOH of the unaffected LKB1 allele, and that additional mutations of the beta-catenin gene and p53 gene convert hamartomatous polyps into adenomatous and carcinomatous lesions. PMID- 11103791 TI - Functional analysis of human ornithine decarboxylase alleles. AB - It has been known for > 10 years that there are two alleles of the human ornithine decarboxylase (ODC) gene, defined by a polymorphic PstI RFLP in intron 1. We have sequenced a large portion of each of the two alleles, including some of the 5' promoter region, exon 1, intron 1, and exon 2, and determined that a single nucleotide polymorphism at base +317 (relative to transcription start site) is responsible for the presence or absence of the PstI restriction site. We have developed two genotyping assays, a PCR-RFLP assay and a high-throughput TaqMan-based method, and determined the ODC genotype distribution in >900 North American DNA samples. On the basis of its location between two closely spaced Myc/Max binding sites (E-boxes), we speculated that the single nucleotide polymorphism at base +317 could have functional significance. Results of transfection assays with allele-specific reporter constructs support this hypothesis. The promoter/regulatory region derived from the minor ODC allele (A allele) was more effective in driving luciferase expression in these assays than the identical region from the major allele (G allele). Our results suggest that individuals homozygous for the A allele may be capable of greater ODC expression after environmental exposures, especially those that up-regulate c-MYC expression. PMID- 11103792 TI - Bax is a transcriptional target and mediator of c-myc-induced apoptosis. AB - The c-Myc oncoprotein is a transcription factor involved in cellular transformation as well as apoptotic cell death. We show here that over-expression of c-Myc delivered by an adenovirus vector up-regulates endogenous proapoptotic bax mRNA and protein expression in human cells. In contrast, the cytotoxic tumor necrosis factor-related apoptosis-inducing ligand induces cell death without up regulating bax expression. c-Myc/Max heterodimers bind to canonical E-box elements located in the bax promoter region as demonstrated by electrophoretic mobility shift analysis and DNaseI foot-printing assays. Analysis of bax regulatory region mutants suggests a model involving myc-dependent activation as well as relief of repression through distinct E-box elements. c-Myc-null cells are deficient in bax-promoter activation as compared with wild-type c-Myc expressing cells. Overexpression of c-Myc in serum-starved human or mouse embryonic cells leads to apoptosis which is significantly reduced in the presence of growth factor-containing serum. c-Myc-induced apoptosis appears to be deficient in bax-null as compared with bax-wild-type mouse embryonic fibroblasts. The results suggest that the cell death-promoting gene bax is directly downstream of c-Myc in a pathway leading to apoptosis. PMID- 11103793 TI - Nitric oxide synthase is induced in tumor promoter-sensitive, but not tumor promoter-resistant, JB6 mouse epidermal cells cocultured with interferon-gamma stimulated RAW 264.7 cells: the role of tumor necrosis factor-alpha. AB - Expression of inducible nitric oxide synthase (iNOS) has been reported to be involved in certain organs of potential tumorigenesis, including the stomach and colon. The mechanisms for iNOS expression in epithelial cells, however, are not fully understood. In the present study, we investigated the role of macrophages in epithelial iNOS expression by coculturing a stimulated murine macrophage-like cell line, RAW 264.7, with either tumor promoter-sensitive (P+) or promoter resistant (P-) JB6 murine epidermal cells. After monoculture, treatment of RAW 264.7 cells with IFN-gamma for 24 h generated a large amount of nitrite (NO2-), as reported previously, whereas no increase in NO2- concentration was observed in the IFN-gamma-treated P+ or P-subclones. Interestingly, when IFN-gamma-treated RAW 264.7 cells were cocultured with P+ but not P- cells, we observed a marked increase in NO2- concentration (30.8+/-3.6 microM), which significantly exceeded (P < 0.01) the sum of the concentrations (20.0+/-2.3 microM) added from each cell line monoculture. Western blotting analysis revealed that, after coculture, iNOS protein was up-regulated 55-fold more than the control in JB6 P+ but not in P- cells. IFN-gamma-treated RAW 264.7 cells secreted proinflammatory cytokines, including tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta. The addition of IFN-gamma-treated RAW 264.7 cell-conditioned media to P+ subclones led to a significant enhancement of NO2- formation that was diminished by the TNF alpha-specific but not IL-1beta-specific antibody. When combined with IFN-gamma, the recombinant TNF-alpha (1-100 ng/ml) enhanced NO2- formation in JB6 P+ cells, whereas IL-1beta (1-100 ng/ml) did not. These results led us to conclude that IFN gamma-treated RAW 264.7 cells release TNF-alpha to induce iNOS expression in promoter-sensitive JB6 cells. Thus, we propose the hypothesis that macrophages stimulate neoplastic cells with TNF-alpha via a paracrine loop to induce epithelial iNOS protein expression. PMID- 11103794 TI - Activator protein 1 transcription factors are fundamental to v-rasHa-induced changes in gene expression in neoplastic keratinocytes. AB - The induction of mouse skin papillomas by initiation-promotion protocols is associated with aberrant expression of epithelial markers in the tumor mass. Similarly, initiation of mouse keratinocytes with a retrovirus encoding the v rasHa gene (v-rasHa keratinocytes) causes characteristic alterations of epidermal gene expression (A. A. Dlugosz et at, Cancer Res., 54: 6413-6420, 1994). Because activator protein 1 (AP-1) proteins are likely targets of Ras activation, we have examined the role of AP-1 factors in v-rasHa keratinocytes. Introduction of v rasHa into keratinocytes up-regulates c-Fos, deltaFos B, and Fra-1 transcripts and protein levels in nuclear extracts. The expression of Jun proteins is not significantly altered in v-rasHa keratinocytes. Transduction of cells with v rasHa results in increased AP-1-dependent transcriptional activity, which is also simulated by transfection of keratinocytes with either c-Fos or deltaFos B but not Fra-1, suggesting that the up-regulation of c-Fos and deltaFos B contributes to this effect. To explore the role of AP-1 proteins in regulating keratinocyte markers in v-rasHa keratinocytes, we blocked the binding of AP-1 proteins to DNA by infecting keratinocytes with an adenovirus encoding a dominant-negative Fos mutant (A-FOS). A-FOS replaces endogenous Fos proteins in the formation of heterodimers with Jun family members and thus prevents the AP-1 transcription factor from binding to DNA. In v-rasHa keratinocytes, the A-FOS virus reversed the suppression of keratins 1 and 10 transcripts and protein, which is characteristically seen in tumors and v-rasHa keratinocytes. A-FOS also increased protein levels but reduced transcripts for the late marker, loricrin, a component of the cornified envelope. These findings indicate that AP-1 proteins are involved in the changes in gene expression that define the v-rasHa phenotype in mouse keratinocytes. PMID- 11103795 TI - The RB1 gene is the target of chromosome 13 deletions in malignant fibrous histiocytoma. AB - Forty-four malignant fibrous histiocytomas (MFHs) were studied by comparative genomic hybridization. Among the observed imbalances, losses of the 13q14-q21 region were observed in almost all tumors (78%), suggesting that a gene localized in this region could act as a tumor suppressor gene and that its inactivation could be relevant for MFH oncogenesis and/or progression. We determined by CA repeat analyses a consensus region of deletion focusing on the RB1 region. The RB1 gene was then analyzed by protein truncation test, direct sequencing, fluorescence in situ hybridization, Southern blotting, and immunohistochemistry. RB1 mutations and/or homozygous deletions were found in 7 of the 34 tumors analyzed (20%). Among the 35 tumors with comparative genomic hybridization imbalances analyzed by immunohistochemistry, 30 (86%) did not exhibit significant nuclear labeling. The high correlation between chromosome 13 losses and absence of RB1 protein expression and the mutations detected strongly suggest that RB1 gene inactivation is a pivotal event in MFH oncogenesis. Moreover, the observation of a high incidence of MFH in patients previously treated for hereditary retinoblastoma fits well this hypothesis. PMID- 11103796 TI - Arsenite induces p53 accumulation through an ATM-dependent pathway in human fibroblasts. AB - Arsenic compounds are potent human carcinogens. Accumulated evidence has shown that arsenite-induced cytogenetic alterations are associated with the carcinogenicity of arsenic. Because p53 plays a guarding role in maintaining genome integrity and accuracy of chromosome segregation, the mechanistic effects of arsenite on p53 activation were analyzed. In the present study, arsenite induced DNA strand breaks were confirmed by alkaline single-cell gel electrophoresis (comet assay) in human fibroblast (HFW) cells. Accompanying the appearance of DNA strand breaks was a significant accumulation of p53 in arsenite treated HFW cells, as demonstrated by immunoblotting and immunofluorescence techniques. p53 downstream proteins, such as p21 and the human homologue of murine double minute-2, were also significantly induced by arsenite treatment. Cell cycle retardation and G2-M arrest were observed in 5-bromo-2'-deoxyuridine pulse-labeled HFW cells by flow cytometry. Wortmannin, an inhibitor of phosphatidylinositol 3-kinases, inhibited arsenite- or X-ray irradiation-induced p53 accumulation but did not alter UV irradiation- or N-acetyl-Leu-Leu norleucinal-induced p53 accumulation. p53 phosphorylation on serine 15 was also confirmed by immunoblotting technique in arsenite- and X-ray-treated HFW cells but was not observed in UV- or N-acetyl-Leu-Leu-norleucinal-treated HFW cells. These results suggest the involvement of a phosphatidylinositol 3-kinase-related protein kinase in arsenite-induced p53 accumulation. For confirmation, we demonstrated that arsenite treatment, similar to X-ray irradiation, did not induce p53 accumulation in GM3395 fibroblasts derived from a patient with ataxia telangiectasia. In contrast, UV irradiation did cause p53 accumulation in these cells. Together, these findings infer that arsenite-induced DNA strand breaks may lead to p53 phosphorylation and accumulation through an ataxia telangiectasia mutated-dependent pathway in HFW cells. PMID- 11103797 TI - Oleate activates phosphatidylinositol 3-kinase and promotes proliferation and reduces apoptosis of MDA-MB-231 breast cancer cells, whereas palmitate has opposite effects. AB - Epidemiological studies and experiments using animal models and cultured breast cancer cells have suggested that a high intake of dietary fat could increase breast cancer risk. Little is known about the biochemical pathways by which various free fatty acids (FFAs) influence breast cancer cell proliferation and apoptosis. The present study was designed to investigate the effects of the two most abundant circulating FFAs, oleate and palmitate, on established human breast cancer cell lines after a short period of serum starvation. The unsaturated FFA oleate (C:18:1) stimulated cell proliferation, whereas the saturated FFA palmitate (C:16) dose dependently inhibited it. The half maximal effective concentrations of oleate and palmitate in the presence of albumin were 5 and 25 microM, respectively. The growth-inhibitory effect of palmitate in MDA-MB-231 cells was related to the induction of apoptosis as indicated by morphological and biochemical criteria. Moreover, oleate protected cells against the proapoptotic action of palmitate. Oleate and palmitate increased and decreased phophatidylinositol 3-kinase (PI3-K) activity, respectively, and the actions of the two FFAs on the enzyme were antagonistic. The PI3-K inhibitors wortmannin and LY294002 completely blocked the proliferative action of oleate. 2-Bromopalmitate, a nonmetabolizable analogue, did not affect MDA-MB-231 cell proliferation, suggesting that palmitate must be metabolized to exert its effect. Thus, various types of fatty acids are not equivalent with respect to their actions on breast cancer cell proliferation and apoptosis. The results support the concept that PI3 K is implicated in the control of breast cancer cell growth by FFAs and that PI3 K may provide a link between fat and cancer. The data are also consistent with the view that the type of FFA and their ratios in the diet in addition to the total amount of fat influence mammary carcinogenesis. PMID- 11103798 TI - Selective replication and oncolysis in p53 mutant tumors with ONYX-015, an E1B 55kD gene-deleted adenovirus, in patients with advanced head and neck cancer: a phase II trial. AB - ONYX-015 is an E1B-55kDa gene-deleted adenovirus engineered to selectively replicate in and lyse p53-deficient cancer cells. To evaluate the selectivity of ONYX-015 replication and cytopathic effects for the first time in humans, we carried out a Phase II clinical testing of intratumoral and peritumoral ONYX-015 injection in 37 patients with recurrent head and neck carcinoma. Patients received ONYX-015 at a daily dose of 1 x 10(10) plaque-forming units (pfu) via intratumoral injection for 5 days during week 1 of each 3-week cycle (n = 30; cohort A), or 1 x 10(10) pfu twice a day for 10 days during weeks 1 and 2 of each 3-week cycle. Posttreatment biopsies documented selective ONYX-015 presence and/or replication in the tumor tissue of 7 of 11 patients biopsied on days 5-14, but not in immediately adjacent normal tissue (0 of 11 patients; P = 0.01). Tissue destruction was also highly selective; significant tumor regression (>50%) occurred in 21% of evaluable patients, whereas no toxicity to injected normal peritumoral tissues was demonstrated. p53 mutant tumors were significantly more likely to undergo ONYX-015-induced necrosis (7 of 12) than were p53 wild-type tumors (0 of 7; P = 0.017). High neutralizing antibody titers did not prevent infection and/or replication within tumors. ONYX-015 is the first genetically engineered replication-competent virus to demonstrate selective intratumoral replication and necrosis in patients. This agent demonstrates the promise of replication-selective viruses as a novel therapeutic platform against cancer. PMID- 11103799 TI - PDZK1 and GREB1 are estrogen-regulated genes expressed in hormone-responsive breast cancer. AB - The function of estrogen in breast cancer proliferation and progression is likely to be due to the expression of a repertoire of genes regulated by estrogen receptor (ER). Using suppression subtractive hybridization, we have isolated a set of 14 estrogen-responsive genes that was differentially expressed in MCF7 cells stimulated by beta-estradiol as compared with unstimulated cells. Tamoxifen repressed the expression of all 14 estrogen-responsive genes. Thirteen of the genes were induced within 6 h of estrogen treatment, indicating that these were early response genes in the ER-regulated pathway. PDZK1 and a new gene, GREB1, demonstrated a significant correlation with ER phenotype in a panel of breast cancer cell lines. Treatment with cycloheximide indicated that ER directly controls GREB1 expression. Three cDNAs (GREB1a, GREB1b, and GREB1c) were isolated by screening a MCF7 cDNA library. These three cDNAs of GREB1 shared extensive sequences through the open reading frame but had divergent 5' untranslated regions, indicating the possibility of multiple promoters regulated by beta estradiol. Studies in primary breast cancers showed that PDZK1 and GREB1 were overexpressed in ER-positive breast cancers as compared with ER-negative breast cancers by 19-fold and 3.5-fold, respectively. GREB1 was also induced by beta estradiol in the ER-positive endometrial cell line ECC-1. The pattern of expression suggests a critical role for these two genes in the response of tissues and tumors to beta-estradiol. PMID- 11103800 TI - Risk for gastric cancer after antibiotic prophylaxis in patients undergoing hip replacement. AB - Despite strong evidence of an association between Helicobacter pylori and gastric cancer, the benefit of eradicating H. pylori infection is unknown. Our aim was to test the hypothesis that exposure to high doses of antibiotics reduces risk for gastric cancer via possible eradication of H. pylori We conducted a nationwide case-control study nested in a cohort of 39,154 patients who underwent hip replacement surgery between 1965 and 1983. Such patients frequently receive prophylactic antibiotic treatment. During follow-up through 1989, we identified 189 incident cases of gastric cancer. For each case, three controls were selected from the cohort. Exposure data were abstracted from hospital records. Blood samples from a separate cohort undergoing hip replacement surgery were analyzed for anti-H. pylori IgG before and after surgery. Both long-term antibiotic treatment before surgery [odds ratio (OR), 0.3; 95% confidence interval (CI), 0.1 0.7] and prophylactic antibiotic treatment (OR, 0.7; 95% CI, 0.5-1.1) conferred a reduction in gastric cancer risk. The reduction appeared stronger after 5 years (OR, 0.6; 95% CI, 0.3-1.2) than during shorter follow-up after hip replacement (OR, 0.8; 95% CI, 0.4-1.7). There was an apparent decrease in risk with increasing body weight-adjusted doses of antibiotics (P = 0.13). However, the rate of H. pylori antibody disappearance was not strikingly higher in the cohort of patients undergoing hip replacement than in a control cohort. Our findings provide indirect support for the hypothesis that treatment with antibiotics at a relatively advanced age reduces the risk of gastric cancer. PMID- 11103801 TI - Glutathione peroxidase codon 198 polymorphism variant increases lung cancer risk. AB - Human cellular glutathione peroxidase 1 (hGPX1) is a selenium-dependent enzyme that participates in the detoxification of hydrogen peroxide and a wide range of organic peroxides. We conducted a case-control study nested within the alpha Tocopherol, beta-Carotene Cancer Prevention Study cohort to evaluate the association between the proline to leucine polymorphism at codon 198 of hGPX1 and lung cancer risk. Cases (n = 315) were matched to controls on age (+/-5 years), intervention group, and study clinic using incidence density sampling in a 1:1 ratio. The prevalence of the hGPX1 Pro198Leu variant allele was 58% for controls and 71% for cases (P < 0.001). Using conditional logistic regression, we found a significant association between hGPX1 genotype and lung cancer risk. The odds ratio for heterozygotes was 1.8 (95% confidence interval, 1.2-2.8) and 2.3 (95% confidence interval, 1.3-3.8) for homozygous variants compared to wild-type individuals. Due to its high prevalence, the hGPX1 variant may contribute significantly to lung cancer risk among Caucasians but not among ethnic Chinese who do not exhibit this polymorphism. PMID- 11103802 TI - Predicting tumor responses to mitomycin C on the basis of DT-diaphorase activity or drug metabolism by tumor homogenates: implications for enzyme-directed bioreductive drug development. AB - Mitomycin C (MMC) is a clinically used anticancer drug that is reduced to cytotoxic metabolites by cellular reductases via a process known as bioreductive drug activation. The identification of key enzymes responsible for drug activation has been investigated extensively with the ultimate aim of tailoring drug administration to patients whose tumors possess the biochemical machinery required for drug activation. In the case of MMC, considerable interest has been centered upon the enzyme DT-diaphorase (DTD) although conflicting reports of good and poor correlations between enzyme activity and response in vitro and in vivo have been published. The principle aim of this study was to provide a definitive answer to the question of whether tumor response to MMC could be predicted on the basis of DTD activity in a large panel of human tumor xenografts. DTD levels were measured in 45 human tumor xenografts that had been characterized previously in terms of their sensitivity to MMC in vitro and in vivo (the in vivo response profile to MMC was taken from work published previously). A poor correlation between DTD activity and antitumor activity in vitro as well as in vivo was obtained. This study also assessed the predictive value of an alternative approach based upon the ability of tumor homogenates to metabolize MMC. This approach is based on the premise that the overall rate of MMC metabolism may provide a better indicator of response than single enzyme measurements. MMC metabolism was evaluated in tumor homogenates (clarified by centrifugation at 1000 x g for 1 min) by measuring the disappearance of the parent compound by HPLC. In responsive [T/C <10% (T/C defined as the relative size of treated and control tumors)] and resistant (T/C >50%) tumors, the mean half life of MMC was 75+/-48.3 and 280+/-129.6 min, respectively. The difference between the two groups was statistically significant (P < 0.005). In conclusion, these results unequivocally demonstrate that response to MMC in vivo cannot be predicted on the basis of DTD activity. Measurement of MMC metabolism by tumor homogenates on the other hand may provide a better indicator of tumor response, and further studies are required to determine whether this approach has real clinical potential in terms of individualizing patient chemotherapy. PMID- 11103803 TI - Tumor regression induced by intratumor administration of adenovirus vector expressing CD40 ligand and naive dendritic cells. AB - We have previously shown that in vivo genetic modification of tumors with an adenovirus (Ad) vector engineered to express CD40 ligand (AdmCD40L) induces tumor specific CTLs and suppresses tumor growth in vivo. In the present study, we investigate the hypothesis that this treatment can be made more efficient with 10(2)-fold less Ad vector by also administering bone marrow-generated dendritic cells (DCs) to the tumor. Using AdmCD40L and the number of DCs that alone had no effect on tumor growth, the data show that the growth of CT26 (colon adenocarcinoma; H-2d) and B16 (melanoma; H-2b) murine s.c. tumors is significantly suppressed by direct administration of DCs into s.c. established tumors that had been pretreated with AdmCD40L 2 days previously. The antitumor effect produced by the combination therapy AdmCD40L + DCs correlated with in vivo priming of tumor-specific CTLs. The antitumor cell-mediated immunity was transferable to naive mice by spleen cells from AdmCD40L + DC-treated animals. The interactions between CD40L and CD40 within treated tumors were critical because tumor suppression was abrogated by coadministration to the tumors of neutralizing monoclonal antibody against CD40L along with the DCs. Finally, in vivo depletion and knockout mice experiments demonstrated that tumor regression produced by this combination therapy depends on CD8+ T cells, but not on CD4+ T cells. These findings should be useful in designing strategies for use of DCs and AdmCD40L in cancer immunotherapy. PMID- 11103804 TI - Viral fusogenic membrane glycoprotein expression causes syncytia formation with bioenergetic cell death: implications for gene therapy. AB - Viral fusogenic membrane glycoproteins (FMGs) are candidates for gene therapy of solid tumors because they cause cell fusion, leading to formation of lethal multinucleated syncytia. However, the cellular mechanisms mediating cell death after FMG-induced cell fusion remain unclear. The present study was designed to examine the mechanisms by which FMG expression in hepatocellular carcinoma cells lead to cell death. Transfection of Hep3B cells with the Gibbon Ape leukemia virus hyperfusogenic envelope protein (GALV-FMG) resulted in the formation of multinucleated syncytia that reached a maximum 5 days after transfection (100 nuclei/syncytia). The syncytia were viable for a period of 2 days and then rapidly lost viability by day 5. Mitochondrial dysfunction occurred in GALV-FMG induced syncytia prior to loss of viability with loss of the mitochondrial membrane potential, cellular ATP depletion, and release of mitochondrial cytochrome c-GFP into the cytosol. The pan-caspase inhibitor, Z-VAD-fmk, did not prevent cell death. However, glycolytic generation of ATP with fructose effectively increased cellular ATP and preserved syncytial viability. These data suggest that expression of FMG in hepatoma cells results in the formation of multinucleated syncytia, causing mitochondrial failure with ATP depletion, a bioenergetic form of cell death with necrosis. This form of cell death should be effective in vivo and enhance the bystander effect, suggesting that FMG-based gene therapy deserves further study for the treatment of hepatocellular and other cancers. PMID- 11103805 TI - Modulation of mitogen-activated protein kinases and phosphorylation of Bcl-2 by vinblastine represent persistent forms of normal fluctuations at G2-M1. AB - Microtubule inhibitors, widely used in cancer chemotherapy, induce G2-M arrest and apoptosis and have in common the ability to stimulate Raf-1/Bcl-2 phosphorylation and activate c-Jun NH2-terminal protein kinase (JNK). These signal transduction pathways are thought to be activated in response to microtubule damage to promote apoptosis. However, Bcl-2 phosphorylation has been reported to occur at G2-M in nonapoptotic cells, raising the possibility that this and perhaps other signaling pathways altered by microtubule inhibitors reflect perturbations of normal mitotic events. In this study, we sought to test this hypothesis. We show that Bcl-2 phosphorylation and JNK activation, as well as extracellular response kinase and p38 inactivation, occur not only in response to vinblastine but also as discrete transient events at G2-M phase in untreated synchronized KB-3 cells. Thus, modulation of these pathways is not a response to microtubule damage; rather they occur normally at G2-M, and it is the extent, duration, and/or irreversible nature of the signals that distinguish a preapoptotic cell from one destined to divide. These findings provide novel insight into the relationship between mitotic and apoptotic signaling and the mechanism of action of antimitotic drugs. PMID- 11103806 TI - The role of c-Jun kinase in the apoptotic response to nucleoside analogue-induced DNA damage. AB - Activation of the c-Jun NH2-terminal kinase type 1 (JNK1) signaling pathway is often associated with apoptosis. In this report, we elucidated the role of this kinase in the programmed cell death induced by the nucleoside analogue 9-beta-D arabinosyl-2-fluoroadenine (F-ara-A). Treatment of ML-1 cells with 3 or 10 microM F-ara-A specifically killed cells in the S-phase of the population. Incorporation of F-ara-ATP, the nucleoside triphosphate of F-ara-A, into DNA resulted in the activation of JNK1 in a time- and dose-dependent fashion. Activation of JNK1 temporally preceded DNA fragmentation. When incorporation of F-ara-A into DNA was blocked by pretreatment of the cells with aphidicolin to inhibit DNA synthesis, neither JNK1 signaling nor apoptosis was evident. Furthermore, inhibition of JNK1 by treatment of the cells with forskolin or by pretreatment with an antisense oligonucleotide directed against JNK1 mRNA resulted in a decrease in F-ara-A induced apoptosis. Finally, the JNK1 signaling pathway appeared to be upstream to that of the effector caspases in nucleoside analogue-induced apoptosis. Thus, our data strongly suggest that JNK1 is involved in transduction of F-ara-A-induced distress signals into an apoptotic response. PMID- 11103807 TI - Antitumor activity of tumor necrosis factor-alpha conjugated with polyvinylpyrrolidone on solid tumors in mice. AB - We attempted the development of a novel polymer conjugation to further improve the therapeutic potency of antitumor cytokines compared with PEGylation for clinical application. Compared with native tumor necrosis factor (TNF)-alpha in vitro, specific bioactivities of polyvinyl-pyrrolidone (PVP)-modified TNF-alphas (PVP-TNF-alphas) were decreased by increasing the degree of PVP attachment. PVP TNF-alpha fraction 3, Mr 101,000, had the most effective antitumor activity of the various PVP-TNF-alphas in vivo. PVP-TNF-alpha fraction 3 had >200-fold higher antitumor effect than native TNF-alpha, and the antitumor activity of PVP-TNF alpha fraction 3 was >2-fold higher than that of MPEG-TNF-alpha (Mr 108,000), which had the highest antitumor activity among the polyethylene glycol (PEG) conjugated TNF-alphas. Additionally, a high dose of native TNF-alpha induced toxic side effects such as body weight reduction, piloerection. and tissue inflammation, whereas no side effects were observed after i.v. administration of PVP-TNF-alpha fraction 3. The plasma half-life of PVP-TNF-alpha fraction 3 (360 min) was about 80- and 3-fold longer than those of native TNF-alpha (4.6 mm) and MPEG-TNF-alpha (122 min), respectively. The mechanism of increased antitumor effect in vivo caused the prolongation of plasma half-life and increase in stability. These results suggested that PVP is a useful polymeric modifier for bioconjugation of TNF-alpha to increase its antitumor potency, and multifunctionally bioconjugated TNF-alpha may be a potentiated antitumor agent for clinical use. PMID- 11103808 TI - Cystathionine-beta-synthase cDNA transfection alters the sensitivity and metabolism of 1-beta-D-arabinofuranosylcytosine in CCRF-CEM leukemia cells in vitro and in vivo: a model of leukemia in Down syndrome. AB - The significantly higher event-free survival rates of Down syndrome (DS) children with acute myeloid leukemia compared with non-DS children is linked to increased sensitivity of DS myeloblasts to 1-beta-D-arabinofuranosylcytosine (ara-C) and the enhanced metabolism of ara-C to ara-C triphosphate (J. W. Taub et al., Blood, 87: 3395-3403, 1996). The cystathionine-beta-synthase (CBS) gene (localized to chromosome 21q22.3) may have downstream effects on reduced folate and S adenosylmethionine pathways; ara-C metabolism and folate pools are linked by the known synergistic effect of sequential methotrexate and ara-C therapy. We have shown that relative CBS transcripts were significantly higher in DS compared with non-DS myeloblasts, and CBS transcript levels correlated with in vitro ara-C sensitivity (J. W. Taub et al., Blood, 94: 1393-1400, 1999). A leukemia cell line model to study the relationship of the CBS gene and ara-C metabolism/sensitivity was developed by transfecting CBS-null CCRF-CEM cells with the CBS cDNA. CBS transfected cells were a median 15-fold more sensitive in vitro to ara-C compared with wild-type cells and generated 8.5-fold higher [3H]ara-C triphosphate levels after in vitro incubation with [3H]ara-C. Severe combined immunodeficient mice implanted with CBS-transfected CEM cells demonstrated greater responsiveness to therapy, reflected in significantly prolonged survivals after ara-C administration compared with mice implanted with wild-type cells and treated with the same dosage schedule. The transfected cells also demonstrated increased in vitro and in vivo sensitivity to gemcitabine. Deoxycytidine kinase (dCK) activity was approximately 22-fold higher in transfected CEM cells compared with wild-type cells. However, levels of dCK transcripts on Northern blots and protein levels on Western blots were nearly identical between CBS-transfected and wild-type cells. Collectively, these results suggest a posttranscriptional regulation of dCK in CBS-overexpressing cells that contributes to increased ara-C phosphorylation and drug activity. Further elucidating the mechanisms of increased sensitivity of DS cells to ara-C related to the CBS gene may lead to the application of these novel approaches to acute myeloid leukemia therapy for non-DS patients. PMID- 11103809 TI - Superior tumor protection induced by a cellular vaccine carrying a tumor-specific T helper epitope by genetic exchange of the class II-associated invariant chain peptide. AB - Efficient loading of MHC class II molecules with a T helper epitope of choice can be achieved through genetic exchange of the MHC class II-associated invariant chain peptide (CLIP) sequence with a sequence encoding the helper peptide. We have now used this method to engineer a cellular vaccine that continuously expresses a tumor-specific helper epitope in a defined costimulatory context. We provide evidence (a) that this cellular vaccine induces peptide-specific helper T cells in vivo that are functional in protecting mice from challenge with a highly aggressive tumor, (b) that this vaccine can directly prime tumor-specific helper T cells in vivo, and (c) that this cellular vaccine is superior compared with similar cells loaded with synthetic T helper peptide in inducing tumor protection. In conclusion, cellular vaccines for activation of antigen-specific helper T cells can be greatly improved by the introduction of invariant chain constructs containing a T helper epitope by class II-associated invariant chain peptide exchange. PMID- 11103810 TI - Targeting of bivalent anti-ErbB2 diabody antibody fragments to tumor cells is independent of the intrinsic antibody affinity. AB - In immunodeficient mice antitumor single-chain Fv (scFv) molecules penetrate tumors rapidly and have rapid serum clearance, leading to excellent tumor:normal organ ratios. However, the absolute quantity of scFv retained in the tumor is low due to rapid serum clearance and monovalent scFv binding. We previously demonstrated that the presence of an additional binding site prolongs in vitro and in vivo association of scFv-based molecules with tumor cells expressing relevant antigen. The contribution of the intrinsic affinity of each component scFv to the association between a dimeric scFv and its target antigen is largely unknown. Here, we have constructed bivalent diabody molecules from three affinity mutants of the human anti-ErbB2 (HER2/neu) scFv molecule C6.5 by shortening the peptide linker between the heavy (VH) and light (VL) chains variable domains from 15 to 5 amino acids. The shorter linker prevents intramolecular pairing of VH and VL, resulting in intermolecular pairing and creation of a dimeric Mr 50,000 molecule with two antigen-binding sites. The scFv used to create the diabodies span a 133-fold range of affinity for the same epitope of ErbB2 [133 nM (C6G98A), 25 nM (C6.5), and 1 nM (C6ML3-9)] and differ by only one to three amino acids. Diabody binding kinetics were determined by surface plasmon resonance on the immobilized ErbB2 extracellular domain. The association rate constants obtained for each diabody molecule were similar to that of the parental (component) scFv. However, the dissociation rate constants obtained for the bivalent diabodies were up to 15-fold slower. The magnitude of the decrease in the bivalent dissociation rate constant was inversely proportional to the monovalent interaction, ranging from only 3-fold for that of the C6ML3-9 diabody to 15-fold for the C6G98A diabody. This resulted in only a 22-fold difference in bivalent affinity, compared with a 133-fold difference in affinity for the respective scFv. Equilibrium-binding constants obtained by surface plasmon resonance correlated well with the equilibrium-binding constants determined in vitro on ErbB2 overexpressing cells. Biodistribution studies were performed in scid mice bearing established SKOV3 tumors. At 24 h, 3-37-fold more diabody was retained in tumor compared with the parental scFv monomers. This likely results from a higher apparent affinity, because of bivalent binding, and a slower serum clearance. Surprisingly, the differences in affinity between diabodies did not result in differences in quantitative tumor retention or tumor to blood ratios. In fact, the diabody constructed from the lowest affinity scFv exhibited the best tumor targeting properties. We conclude that, above a threshold affinity, other factors regulate quantitative tumor retention. In addition, straightforward dimerization of a low-affinity scFv leads to significantly greater tumor localization than does exhaustive scFv affinity maturation. PMID- 11103811 TI - Dead or alive: immunogenicity of human melanoma cells when presented by dendritic cells. AB - Uveal melanoma is an aggressive malignancy with a poor prognosis despite current therapeutic intervention. These tumors have been shown to be antigenic because they express a number of melanoma-associated antigens and are therefore attractive targets for immunotherapy. Here, we investigated the immunogenicity of uveal melanoma cells that have undergone apoptosis and compared this with their necrotic or live counter-parts. The fate of the tumor antigens in these cells largely depends on their ability to be processed and phagocytosed by dendritic cells (DCs). Flow cytometric analysis shows that human DCs form conjugates more efficiently with dead uveal melanoma cells, and consequently these are effective stimuli of lymphocyte proliferation. However, only DCs pulsed with apoptotic cells were able to induce proliferation of CD8+ cytotoxic T cells and stimulate antigen-specific T cells. This study demonstrates for the first time that DCs derived from melanoma patients process and present antigens derived from both HLA matched or HLA-mismatched human apoptotic tumor cells stimulating both CD4+ and CD8+ T cells. This approach may be important to the development of DC-based immunotherapies for melanoma. PMID- 11103812 TI - A novel melanoma gene (MG50) encoding the interleukin 1 receptor antagonist and six epitopes recognized by human cytolytic T lymphocytes. AB - We identified a novel 8.1-kb human melanoma gene, MG50, derived from subtractive hybridization with a squamous lung carcinoma cell line, LU-1. 6.8 kb containing an open reading frame were sequenced, and the structure of the encoded 1496 amino acid protein was deduced. With HLA-A2.1-transduced Drosophila cells as antigen presenting cells, we identified six epitopes restricted by HLA-A2.1 that elicited CTLs in vitro. Reactivity of the CTLs to melanoma cells containing MG50 indicated that the epitopes were displayed naturally. Significant cross-reactivity of CTLs immunized against a melanoma cell line that lacked HLA-A2.1 indicated that at least four of the epitopes were also recognized in a different HLA class I context, most likely HLA-A*6802. By quantitative reverse transcription, MG50 message was found in one of two skin melanoma cell lines, an ocular melanoma cell line, two of four metastatic skin melanomas, two of three mammary carcinomas, one of two colon carcinomas, and an ovarian carcinoma. Of six normal tissues, MG50 was found only in a specimen of normal skin and was absent from a congenital nevus. It is likely that MG50 is the gene for the interleukin 1 receptor antagonist because a reported sequence of cDNA from the latter had a sequence of 528 bases in the 3' region, a long contiguous base sequence, and 176 encoded amino acids identical with those of MG50. MG50 is one of the few melanoma associated antigens that is not a differentiation antigen or a mutated protein. Because of its nature, it may prove to be important in the pathogenesis of the tumors in which it is found, as well as an immunogen and target for immunotherapy. PMID- 11103813 TI - Adenovirus-interleukin-12-mediated tumor regression in a murine hepatocellular carcinoma model is not dependent on CD1-restricted natural killer T cells. AB - The cytokine interleukin-12 (IL-12) has shown potent antitumor activity in several tumor models. Recently, natural killer (NK) T cells have been proposed to mediate the antitumor effects of IL-12. In this study, the antitumor response of IL-12 was investigated in a gene therapeutic model against s.c. growing mouse hepatocellular carcinomas using an adenoviral vector expressing murine IL-12 (AdVmIL-12). An adenoviral-based system was chosen because of the ability of adenoviruses to transduce dividing and nondividing cells and because of their high transduction efficiencies. Our goals were to examine the efficacy of AdVmIL 12 in a hepatocellular carcinoma model and to investigate the mechanism of the AdVmIL-12-mediated antitumor response with specific interest in the role of NK T cells. Our studies demonstrate that intratumoral AdVmIL-12-mediated regression of s.c. hepatocellular tumors is associated with rapid antitumor responses. AdVmIL 12 treatment was associated with an immune cellular infiltrate consisting of CD4 and CD8 T lymphocytes, macrophages, NK cells, and NK T cells. Antibody ablation of CD4 and CD8 T cells and use of NK cell-defective beige mice failed to abrogate the response to AdVmIL-12. Studies in T-cell- and B-cell-deficient severe combined immunodeficient and recombinase activating gene-2-deficient mice and T cell-, B-cell-, and NK cell-defective severe combined immunodeficient/beige mice also failed to abrogate this response. AdVmIL-12 retained potent antitumor activity in mice with specific genetic defects in immune cellular cytotoxicity (perforin knockout mice) and costimulation (CD28 knockout mice). Use of mice with specific NK T cell deficiencies, Valpha14 T-cell receptor and CD1 knockout mice, also failed to abrogate the response to AdVmIL-12. Histological and immunohistochemical studies of AdVmIL-12-treated tumors showed extensive inhibition of neovascularization and a marked decrease in factor VIII-stained endothelial cells. Our studies indicate that the antitumor response of AdVmIL-12 is independent of direct cytotoxic cellular immunity (specifically, the function of NK T cells) and suggest that the initial mechanisms of AdVmIL-12-mediated tumor regression involve inhibition of angiogenesis. PMID- 11103814 TI - Differential effects of theaflavin monogallates on cell growth, apoptosis, and Cox-2 gene expression in cancerous versus normal cells. AB - Theaflavin (TF-1), theaflavin-3-monogallate and theaflavin-3'-monogallate mixture (TF-2), and theaflavin-3,3'-digallate (TF-3) are the major black tea polyphenols. Here we compared the effects of these polyphenols on cell growth, apoptosis, and gene expression in normal and cancerous cells. We showed that TF-2 (10-50 microM) inhibited the growth of SV40 transformed WI38 human cells (WI38VA) and Caco-2 colon cancer cells but had little effect on the growth of their normal counterparts. The IC50s of TF-2 for the growth inhibition of WI38 and WI38VA cells were, respectively, 300 and 3 microM. The other two black tea polyphenols, TF-1 and TF-3, did not exhibit such differential growth-inhibitory effect. TF-2, but not TF-1 or TF-3, induced apoptosis in transformed WI38VA cells but not in normal WI38 cells, suggesting that apoptosis was responsible, at least in part, for the differential growth-inhibitory effect of TF-2. Cox-2 has been implicated in intestinal carcinogenesis. Among the tea polyphenols tested, TF-2 and, to a lesser degree, (-)-epigallocatechin gallate inhibited cyclooxygenase (Cox)-2 gene expression. TF-2 at 50 microM completely blocked the serum-induced Cox-2 gene expression at both mRNA and protein level. Other genes, including c-fos, c-myc, thymidine kinase, proliferating cell nuclear antigen, BRCA1, BRCA2, and Cox-1, were not significantly affected by TF-2. These findings suggest that TF-2 may be responsible, at least in part, for the chemopreventive activity in black tea extracts. PMID- 11103816 TI - Two percent of Finnish prostate cancer patients have a germ-line mutation in the hormone-binding domain of the androgen receptor gene. AB - Mutations of the androgen receptor (AR) gene have been reported in prostate cancer, usually from tumor tissue specimens from late-stage, androgen-independent cancer. Occasionally, germ-line mutations have been found, but a link between AR mutations and predisposition to human prostate cancer has not been firmly established. Recently, two independent studies reported the same germ-line mutation at codon 726 in exon E (CGC to CTC) in two apparently unrelated Finnish prostate cancer patients. This arginine to leucine substitution was reported to alter the transactivational specificity of the AR protein. In the present study, the R726L mutation was analyzed by allele-specific oligohybridization in DNA specimens from 418 consecutive prostate cancer patients who reported a negative family history (sporadic group) and from 106 patients with a positive family history (hereditary group). The population frequency of the R726L mutation in blood donors was 3 of 900 (0.33%). In contrast, eight (1.91%) mutations (odds ratio = 5.8; P = 0.006) were found in the sporadic group, and two (1.89%) mutations were found in the hereditary group (odds ratio = 5.8; P = 0.09). Suggestive evidence of the segregation of the mutation with prostate cancer was seen in these two families. The present study indicates that the R726L substitution in the AR may confer an up to 6-fold increased risk of prostate cancer and may contribute to cancer development in up to 2% of Finnish prostate cancer patients. These results warrant additional large-scale studies of the significance of rare mutations and polymorphisms in candidate genes along the androgen signaling pathway as risk factors for prostate cancer. PMID- 11103815 TI - Prognostic value of genetically diagnosed lymph node micrometastasis in non-small cell lung carcinoma cases. AB - The predictive value of lymph node micrometastasis, detected by immunohistochemical or genetic methods, is well appreciated in terms of prognosis. However, a major problem is high false-positive rates, because most methods focus on cytokeratin, which is a component not only of carcinoma but also normal epithelial and nonepithelial cells. Mutant allele-specific amplification (MASA) can detect DNAs derived from cancer cells itself, reportedly with high sensitivity. It was, therefore, used with nested-PCR using p53 or K-ras mutation for analysis of lymph node micrometastasis in non-small cell lung carcinoma (NSCLC) patients in the present study, in comparison with the immunohistochemical method using an anti-cytokeratin reagent for the same samples. Lymph nodes from 31 NSCLC patients with p53 and K-ras mutated tumors (30 and 1, respectively) staged as pathological (p)-T1-4 N0-1 and M0 were examined. Genetic and immunohistochemical methods demonstrated positive reactions in 34 (15%) and 61 (27%) of 229 lymph nodes, respectively (9 cases, 29%, and 24 cases, 77%). The concordance with the two methods was 77%, but 13 (39%) of 34 genetically positive lymph nodes could not be detected by immunohistochemistry (IHC). Of 22 cases with p-N0 disease, 6 (27%) were genetically positive in hilar and/or mediastinal lymph nodes, and 4 (67%) of them died after cancer relapse. In contrast, none of the patients without micrometastasis died of cancer (P < 0.001, log rank analysis). Of the same p-N0 patients, 17 (77%) were positive by IHC, and 4 (24%) of them died of cancer, whereas 5 negative patients did not suffer cancer relapse. Survival did not significantly differ between cases positive and negative (P = 0.246) by IHC. According to the g-N (N factor restaged by a genetic method), patients with g-N1 and g-N2 disease had a shorter survival than those with g-N0 disease (P = 0.042 and P < 0.001, respectively). However, no significant difference was observed with grading by IHC. Thus, detection of micrometastasis in regional lymph nodes with the MASA method, in other words with a carcinoma specific marker, is of greater prognostic significance for early stage NSCLC patients than immunohistochemical results. This approach should facilitate selection of patients for whom postoperative adjuvant chemotherapy should be performed. PMID- 11103817 TI - Induction of mammary differentiation by mammary-derived growth inhibitor-related gene that interacts with an omega-3 fatty acid on growth inhibition of breast cancer cells. AB - We previously identified and characterized a novel tumor growth inhibitor and a fatty acid-binding protein in human mammary gland and named it the mammary derived growth inhibitor-related gene (MRG). Here, the effects of MRG on mammary gland differentiation and its interaction with omega-3 polyunsaturated fatty acids (omega-3 PUFAs) on growth inhibition were investigated. MRG protein expression was associated with human mammary gland differentiation, with the highest expression observed in the differentiated alveolar mammary epithelial cells from the lactating gland. Overexpression of MRG in human breast cancer cells induced differentiation with changes in cellular morphology and a significant increase in the production of lipid droplets. Treatment of mouse mammary gland in organ culture with MRG protein resulted in a differentiated morphology and stimulation of beta-casein expression. Treatment of human breast cancer cells with the omega-3 PUFA docosahexaenoic acid resulted in a differential growth inhibition proportional to their MRG expression. MRG transfected cells or MRG protein treated cells were much more sensitive to docosahexaenoic acid-induced growth inhibition than MRG-negative or untreated control cells. Our results suggest that MRG is a candidate mediator of the differentiating effect of pregnancy on breast epithelial cells and may play a major role in omega-3 PUFA-mediated tumor suppression. PMID- 11103818 TI - Increase of GKLF messenger RNA and protein expression during progression of breast cancer. AB - Genetic alterations found in carcinomas can alter specific regulatory pathways and provide a selective growth advantage by activation of transforming oncogenes. A subset of these genes, including wild-type alleles of GLI or c-MYC, and activated alleles of RAS or beta-catenin, exhibit transforming activity when expressed in diploid epithelial RK3E cells in vitro. By in vitro transformation of these cells, the zinc finger protein GKLF/KLF-4 was recently identified as a novel oncogene. Although GKLF is normally expressed in superficial, differentiating epithelial cells of the skin, oral mucosa, and gut, expression is consistently up-regulated in dysplastic epithelium and in squamous cell carcinoma of the oral cavity. In the current study, we used in situ hybridization, Northern blot analysis, and immunohistochemistry to detect GKLF at various stages of tumor progression in the breast, prostate, and colon. Overall, expression of GKLF mRNA was detected by in situ hybridization in 21 of 31 cases (68%) of carcinoma of the breast. Low-level expression of GKLF mRNA was observed in morphologically normal (uninvolved) breast epithelium adjacent to tumor cells. Increased expression was observed in neoplastic cells compared with adjacent uninvolved epithelium for 14 of 19 cases examined (74%). Ductal carcinoma in situ exhibited similar expression as invasive carcinoma, suggesting that GKLF is activated prior to invasion through the basement membrane. Expression as determined by Northern blot was increased in most breast tumor cell lines and in immortalized human mammary epithelial cells when these were compared with finite-life span human mammary epithelial cells. Alteration of GKLF expression was confirmed by the use of a novel monoclonal antibody that detected the protein in normal and neoplastic tissues in a distribution consistent with localization of the mRNA. In contrast to most breast tumors, expression of GKLF in tumor cells of colorectal or prostatic carcinomas was reduced or unaltered compared with normal epithelium. The results demonstrate that GKLF expression in epithelial compartments is altered in a tissue-type specific fashion during tumor progression, and suggest that increased expression of GKLF mRNA and protein may contribute to the malignant phenotype of breast tumors. PMID- 11103819 TI - Visualization of the timing of gene amplification during multistep head and neck tumorigenesis. AB - Head and neck tumorigenesis is thought to represent a multistep process whereby carcinogen exposure leads to genetic instability in the tissue and the accumulation of specific genetic events, which result in dysregulation of proliferation, differentiation, and cell loss and the acquisition of invasive capacity. Chromosome 11q13 amplification is frequently observed in head and neck squamous cell carcinoma (HNSCC), and the amplified gene products are assumed to play important functional roles in the tumor phenotype. However, it is not well understood whether gene amplification precedes carcinoma development or results from the unstable nature of intact tumors. To determine the timing of gene amplification during tumorigenesis, tissue sections from amplified HNSCC specimens (containing a contiguous transition from normal epithelium to hyperplasia to dysplasia to carcinoma) were probed for INT2 gene copy number by chromosome in situ hybridization. In addition, representative epithelia were microdissected from the tissue sections, and the DNA was isolated and assessed for INT2 gene copy number by semiquantitative PCR. In those cases containing amplified INT2 in the carcinoma, gene amplification appeared to precede HNSCC development. In one case, INT2 gene amplification appeared in the hyperplasia to dysplasia transition, whereas in two other cases, gene amplification was apparent at dysplasia. These results suggest that gene amplification can occur early during head and neck tumorigenesis and that genetic instability is an important driving force in the tumorigenesis process. PMID- 11103820 TI - Construction of evolutionary tree models for renal cell carcinoma from comparative genomic hybridization data. AB - Renal cell carcinoma is characterized by an accumulation of complex chromosomal alterations during tumor progression. Chromosome 3p deletions are known to occur early in the carcinogenesis, but the nature of subsequent events, their interrelationships, and their sequence is poorly understood, as one usually only obtains a single "view" of the dynamic process of tumor development in a particular cancer patient. To address this limitation, we used comparative genomic hybridization analysis in combination with a distance-based and a branching-tree method to search for tree models of the oncogenesis process of 116 conventional (clear cell) renal carcinomas. This provides a means to analyze and model cancer development processes based on a more dynamic model, including the presence of multiple pathways, as compared with the fixed linear model first proposed by Vogelstein et al. (N. Engl. J. Med., 319: 525-532, 1988) for colorectal cancer. The most common DNA losses involved 3p (61%), 4q (50%), 6q (40%), 9p (35%), 13q (37%), and Xq (21%). The most common gains were seen at chromosome 17p and 17q (20%). The tree model derived from the distance-based method is consistent with the established theory that -3p is an important early event in conventional (clear cell) renal cancer and supports the prediction made from the branching tree that -4q is another important early event. Both tree models suggest that there may be two groups of clear cell renal cancers: one characterized by -6q, +17q, and + 17p, and another by -9p, -13q, and -18q. Putative prognostic parameters were -9p and -13q. The distance-based tree clarifies that -8p (present in 12% of tumors) is a late event, largely independent of other events. In summary, tree modeling of comparative genomic hybridization data provided new information on the interrelationships of genetic changes in renal cancer and their possible order, as well as a clustering of these events. Using tree analysis, one can derive a more in-depth understanding of the renal cancer development process than is possible by simply focusing on the frequencies of genetic events in a given cancer type. PMID- 11103821 TI - Clonal selection versus genetic instability as the driving force in neoplastic transformation. AB - Recent clonal studies of spontaneous neoplastic transformation in cell culture indicate that it develops at confluence in a small minority of individual clonal populations before it does in the uncloned parental culture. Either preferential selection of spontaneous variants or genetic destabilization in clones can be inferred to explain the result. In the present experiments, using a subline of NIH 3T3 cells that is relatively refractory to transformation, we demonstrate unequivocally that transformed foci appear under selective conditions in some clones long before there is any sign of neoplastic change in the polyclonal culture from which they were derived. Because the transformed cells that appear in the susceptible clones are not inhibited in the size or number of foci formed on a confluent background of the uncloned parental population, the genetic events underlying transformation must occur much less frequently in the latter. This disparity can be accounted for by the much larger number of selectable cells in the susceptible clones at confluence than in the parental culture, where such cells are a minority. The preferential transformation exhibited by experimental isolation and expansion of susceptible clones accords with evidence from various sources that neoplastic transformation in culture is a multistep process dependent primarily on selection of spontaneously occurring genetic variants. There is no necessity to posit a significant role for genetic destabilization in neoplastic transformation. These considerations bolster computer models of human cancer that implicate selective expansion of rogue clones rather than genetic instability as the driving force in the origin of most tumors. Both the genetics of the selected clone and the epigenetics of the selective environment would then contribute to tumor development. PMID- 11103822 TI - Involvement of the Ets-1 gene in overexpression of matrilysin in human hepatocellular carcinoma. AB - Although matrix metalloproteinases (MMPs) are thought to be involved in the invasion and metastasis of a variety of malignant tumors, including human hepatocellular carcinoma (HCC), the mechanisms for the expression of MMPs in HCC are not known. To understand the mechanism(s) of MMP expression, the expression of matrilysin (MMP-7) and several genes of the Ets transcription factor family was investigated in human HCC and hepatoma-derived cell lines. The role of Ets-1 gene expression in HCC was also studied. Analysis by semiquantitative reverse transcription-PCR revealed that MMP-7 and Ets-1 are overexpressed and closely associated in HCC. To clarify the role of Ets-1, hepatoma cells were transduced with human Ets-1 or targeted with the Ets-1-specific antisense oligonucleotides. Cells stably transduced with the Ets-1 gene showed increased MMP-7 expression compared to parental and mock-transfected cells. Cells targeted with Ets-1 specific antisense oligonucleotides showed reduced expression of MMP-7. Cotransfection of cells with a MMP-7 promoter-reporter gene plasmid and an Ets-1 expression vector yielded an increase in MMP-7 promoter activity in an Ets-1 responsive element-dependent manner. Taken together, these data suggested that the Ets-1 oncogene is up-regulated and involved in the overexpression of MMP-7 in human HCC and may contribute to the progression of HCC. PMID- 11103823 TI - Treatment of prostate cancer by radioiodine therapy after tissue-specific expression of the sodium iodide symporter. AB - Causing prostate cancer cells to express functionally active sodium iodide symporter (NIS) by targeted NIS gene transfer might offer the possibility of radioiodine therapy of prostate cancer. Therefore, we investigated radioiodine accumulation and therapeutic effectiveness of 131I in NIS-transfected prostate cancer cells in vitro and in vivo. The human prostatic adenocarcinoma cell line LNCaP was stably transfected with NIS cDNA under the control of the prostate specific antigen promoter. The stably transfected LNCaP cell line NP-1 showed perchlorate-sensitive, androgen-dependent iodide uptake in vitro that resulted in selective killing of these cells by 131I in an in vitro clonogenic assay. Xenografts were established in athymic nude mice and imaged using a gamma camera after i.p. injection of 500 microCi of 123I. In contrast to the NIS-negative control tumors (P-1) which showed no in vivo uptake of 123I, NP-1 tumors accumulated 25-30% of the total 123I administered with a biological half-life of 45 h. In addition, NIS protein expression in LNCaP cell xenografts was confirmed by Western blot analysis and immunohistochemistry. After a single i.p. application of a therapeutic 131I dose (3 mCi), significant tumor reduction was achieved in NP-1 tumors in the therapy group compared with P-1 tumors and tumors in the control group. In conclusion, a therapeutic effect of 131I has been demonstrated in prostate cancer cells after induction of tissue-specific iodide uptake activity by prostate-specific antigen promoter-directed NIS expression in vitro and in vivo. This study demonstrates the potential of NIS as a novel therapeutic gene for nonthyroidal cancers, in particular prostate cancer. PMID- 11103825 TI - Induction of Fas expression and augmentation of Fas/Fas ligand-mediated apoptosis by the synthetic retinoid CD437 in human lung cancer cells. AB - The synthetic retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) induces apoptosis in a variety of cancer cells. Recently, we demonstrated that CD437 induces apoptosis in human non-small cell lung cancer (NSCLC) cells expressing wild-type p53 by increasing the level of the death domain-containing cell surface receptor Killer/DR5. In the present study, we investigated whether CD437 induced the expression of Fas (CD95/APO-1), a cell surface protein belonging to the tumor necrosis factor receptor superfamily, which induces apoptosis upon interaction with Fas ligand (FasL) or agonistic antibodies. We found that CD437 increased the level of Fas mRNA in a time- and concentration-dependent manner in NSCLC H460 cells. The increased Fas expression was also identified at the protein level. CD437 induced Fas expression in three NSCLC cell lines with wild-type p53 but not in six NSCLC cell lines containing mutant p53. Moreover, enhanced degradation of wild-type p53 protein in NSCLC cells expressing human papillomavirus-16 E6 oncoprotein blocked CD437-induced Fas expression. These results implicate the involvement of wild-type p53 in CD437 induced Fas expression in human NSCLC cells. CD437 did not change Fas mRNA stability, and actinomycin D abolished CD437-induced expression of Fas mRNA, suggesting that CD437 induces Fas expression at the transcriptional level. The combination of CD437 and FasL or CD437 and agonistic anti-Fas antibody caused synergistic induction of apoptosis. Furthermore, CD437 augmented Fas/ FasL induced apoptosis in cell lines with wild-type p53 but not in cell lines having mutant p53, indicating that a p53-dependent mechanism is also involved in this effect. Taken together, these results demonstrate that increased Fas expression may play an important role in CD437-induced, p53-dependent apoptosis in human NSCLC cells. PMID- 11103824 TI - Sp1 decoy transfected to carcinoma cells suppresses the expression of vascular endothelial growth factor, transforming growth factor beta1, and tissue factor and also cell growth and invasion activities. AB - Vasculature development is thought to be an important aspect in the growth and metastasis of solid tumors. Among the angiogenic factors produced by tumor cells, vascular endothelial growth factor is considered to be the most potent and pathologically important. The synthesis of this growth factor has been shown to be modulated through Sp1 function following stimulation by tumor necrosis factor alpha (TNF-alpha). Oligodeoxynucleotides (ODNs) were synthesized with either the consensus sequence for Sp1 binding (Sp1 decoy ODNs) or a mutated form of this sequence (mt-Sp1 decoy ODNs). Using the hemagglutinating virus of Japan (HVJ) liposome method, we transferred these ODNs into cultured cancer cells (A549 and U251 cells). The TNF-alpha-mediated expression of both VEGF and transforming growth factor beta1 and tissue factor (TF) by the cancer cells could be simultaneously suppressed to less than 30% by transfection of Sp1 decoy ODNs but not by mt-Sp1 decoy ODNs. In addition, in vitro invasiveness, synthesis of mRNA for urokinase-type plasminogen activator, and cell proliferation of both cell lines were also inhibited to 40% by the transfection of only Sp1 decoy ODNs. These results suggested that the Sp1 decoy strategy would be effective for regulating tumor growth by simultaneously reducing cancer cell (a) angiogenic growth factor expression, (b) proliferation, and (c) invasiveness. PMID- 11103826 TI - Psychosocial variables and coronary heart disease. PMID- 11103827 TI - On the electrocardiographic diagnosis of biventricular infarctions. AB - To diagnose posterior and anterior biventricular infarctions it is necessary to record from right and left thoracic and high abdominal unipolar leads. These supplementary leads are dependable, can be repeated as many times as needed and show the evolution from signs of myocardial injury to those of dead tissue (Q waves of 0.04 sec or more). This electrocardiographic evolution increases the diagnostic value of the electrical exploration, since the injury current can be observed also in other conditions. The diagnosis of right ventricular infarction can be established even in the presence of RBBB. Signs of a dead zone in the free right ventricular wall are more frequently observed in posterior biventricular infarctions than in anterior ones. In these cases, the signs of subepicardial injury are more accentuated in the right thoracic unipolar leads than in V3, indicating anterior right ventricular involvement. These signs are also observed in experimental studies made in animals. This electrocardiographic exploration opens a wide field for the diagnosis of myocardial infarction, particularly in biventricular involvement, including old myocardial scars, and in discarding signs of pericarditis manifested only by the upward displacement of the ST segment. A review of the medical literature concerning diagnosis of biventricular infarctions is presented. PMID- 11103828 TI - Comparison of low-dose dobutamine echocardiography and thallium-201 reinjection for the determination of myocardial viability in the early post myocardial infarction period. AB - OBJECTIVE: Determination of viability in the infarction zone in the early post Ml period is an important parameter in clinical decision making. METHODS: In an attempt to compare the places of low-dose dobutamine echocardiography (LDDE) and thallium-201 reinjection SPECT (TI-SPECT) in the determination of viability in dyssynergic myocardial segments, 17 patients (mean age: 54.6 +/- 12.8 years, 16 male, 1 female) with a recent myocardial infarction and an uneventful early clinical course underwent both tests within 5-13 days of infarction. The 16 segment model was utilised to evaluate the left ventricular wall motion and each segment was graded as 1) normokinetic, 2) hypokinetic, 3) akinetic and 4) dyskinetic or aneurysmal on a 4-scale basis. A dyssynergic segment of myocardium was considered to be viable by LDDE if it showed an improvement in wall motion of at least one grade with low-dose dobutamine infusion (10 microg/kg/min). On the other hand, mild to moderate (< 50%) fixed perfusion defects and reversible (at least a 10% improvement in perfusion on either redistribution or reinjection images) severe (50% or more) perfusion defects were considered positive for viability by TI-SPECT. RESULTS: Of the 76 segments with resting dyssynergy (10 dyskinetic/aneurysmal, 33 akinetic, 33 hypokinetic), 51 (67%) were shown to be viable by LDDE and 61 (80%) by TI-SPECT. There was an agreement of 76% (p = 0.03, K = 0.63) between the two methods. CONCLUSION: This study disclosed a moderate degree of agreement between LDDE and TI-SPECT for the determination of viability in dyssynergic myocardial segments in the early post-myocardial infarction period. PMID- 11103829 TI - Chlamydia and atherosclerotic coronary arterial disease in Turkey. AB - OBJECTIVE: Chlamydia pneumoniae, which is a Gram(-) intracellular bacteria, besides being a respiratory pathogen, is thought to play an active role in the progress of acute myocardial infarction and chronic coronary artery disease. In this study we aim to determine the frequency of C. pneumoniae in coronary artery lesions of Turkish people. METHODS AND RESULTS: The atherosclerotic material taken from 8 cases by directional atherectomy and from 23 cases by surgical endarterectomy and examined by indirect immunofluorescence (IIFA) test and polymerase chain reaction (PCR). C. pneumoniae positivity was 32.3% (10/31) by IIFA and 29.0% (9/31) by PCR while the evaluation of the methods together yield a positivity of 35.5% (11/31). CONCLUSIONS: A statistically significant difference could not be established between C. pneumoniae positive and negative groups according to age and the classical atherosclerotic risk factors such as diabetes mellitus, smoking, hypercholesterolaemia, hypertension, family history; besides, a statistically significant difference could not be found between the presence of C. pneumoniae and the severity and clinical picture of coronary artery disease. PMID- 11103830 TI - Effects of cigarette smoking on the circadian rhythm of heart rate variability. AB - BACKGROUND: The effects of cigarette smoking on the circadian rhythm of heart rate variability (HRV) are not known. METHODS: We studied the effects of cigarette smoking on the circadian rhythm of HRV in 24 smoking and 21 non-smoking healthy subjects. Twenty-four hour ambulatory electrocardiograms were recorded and time domain parameters of HRV (SDNN [standard deviation of all R-R intervals], SDANN [standard deviation of the averages of R-R intervals in all 5 minute segments of the entire recording], RMSSD [the square root of the mean of the sum of the squares of differences between adjacent R-R intervals]) were determined for the entire 24-hour period and for each 3-hour period. RESULTS: In total, SDNN and SDANN were significantly lower in smokers than non-smokers (116 +/- 26 vs 136 +/- 27, p < 0.05 for SDNN, 109 +/- 25 vs 121 +/- 24, p < 0.05 for SDANN). However, there were no statistical differences between smokers and non smokers in heart rate (81 +/- 9 vs 76 +/- 10, p > 0.05) and RMSSD (32 +/- 12 vs 37 +/- 18, p > 0.05). These HRV parameters showed a circadian variation: they increased at night and decreased during the day in both groups. The parameters were lower in smokers than non-smokers during daytime (especially, between 8-14 hours). However, no differences were detected during night-time. CONCLUSIONS: Time domain parameters of HRV (SDNN, SDANN and RMSSD) in both smoking and non smoking healthy subjects have a circadian rhythm. SDNN and SDANN were lower in smokers than non-smokers during daytime. PMID- 11103831 TI - A case report of a patient with a large aneurysmatic coronary artery fistula. AB - A coronary artery fistula is a rare congenital malformation, which can become symptomatic in adulthood. This report describes a 65-year-old patient with a large aneurysmatic fistula who presented with signs of heart failure. Angiographically a large aneurysmatic fistula was found running from the left coronary artery to the junction of the superior vena cava and the right atrium. PMID- 11103832 TI - Long-term left ventricular pacing. AB - A 57-year-old female was implanted with a Biotronik Pikos VVI pulse generator. During her follow-up period right bundle-branch block was observed. Therefore further posteroanterior and lateral chest X-ray and echocardiography were performed. Only the transoesophageal echocardiography showed exactly the abnormal pathway of the electrode through the foramen ovale apertum to the left side of the heart. Abnormal electrode position can create serious complications, however, our patient remained free of symptoms throughout her 5 year follow-up. PMID- 11103833 TI - Coronary aneurysm five months after intracoronary beta-irradiation. AB - Intracoronary beta-irradiation is believed to be useful in preventing restenosis after coronary angioplasty or as adjunct therapy of an in-stent restenosis. Intracoronary aneurysms after gamma-irradiation were reported by Condado et al. in 1995, especially after doses higher than 25 Gy and without a centering device. In repeated small trials using intracoronary beta-irradiation no aneurysms were reported at 6 months follow-up. We report the development of a coronary aneurysm at 5 months after intracoronary beta-irradiation and stenting. Intracoronary brachytherapy is a new promising technique although one should be cautious about its possible unknown long-term complications. PMID- 11103834 TI - Occupation and lung disease. PMID- 11103835 TI - Biopsychosocial rehabilitation for repetitive-strain injuries among working-age adults. AB - The objective of this study was to determine the effectiveness of biopsychosocial rehabilitation for upper-limb repetitive-strain injuries among working-age adults. Studies were identified from electronic bibliographic databases, reference checks, and consultations with experts in rehabilitation. Four blinded reviewers selected randomized controlled and controlled trials. Two experts evaluated the clinical relevance of the findings. Two other reviewers extracted the data and assessed the main results and the methodological quality of the studies. Finally, a qualitative analysis was performed. Only 2 studies satisfied the criteria. They were both considered to be low-quality trials. The clinical relevance of the included studies was also unsatisfactory. The level of scientific evidence was limited, showing that hypnosis as a supplement to comprehensive treatment can decrease the pain intensity of acute repetitive strain injury in short follow-ups. There appears to be little scientific evidence for the effectiveness of biopsychosocial rehabilitation with respect to repetitive-strain injuries. PMID- 11103836 TI - Impact of occupation on respiratory disease. AB - OBJECTIVES: This study identified occupations with a marked impact on sick leaves due to respiratory disease. METHODS: A national sick-leave register containing information on all sick leaves exceeding 14 days, physicians' diagnoses, and the occupational status of all manual and service employees in the private sector in Sweden was studied. Sick leaves during 1992-1994 (N=210,755) were analyzed with special attention to respiratory disease and occupation. RESULTS: Respiratory disease accounted for 4.4% of the total number of sick leaves. The incidence of long-term (> or = 90 days) sick leaves due to respiratory disease was 3 times higher in occupations with a high incidence than in those with a low incidence. There was a high correlation (r=0.80) between the incidence of long-term sick leave due to respiratory disease and sick leave due to all other conditions; this finding suggests that market and selection factors may play an important role in determining the overall risk for sick leave in various occupations. The proportion of sick leaves due to long-term respiratory disease out of all long term disease was compared between occupations. Agricultural workers had a 46% higher proportion of long-term respiratory disease than metal workers. Industrial workers, food industry workers, and painters were also occupations with an increased risk. These findings could not be explained by differences in age or smoking habits. CONCLUSIONS: Major differences were found among manual and service occupations regarding long-term sick leave due to respiratory disease. Several occupations, in which exposure to respiratory sensitizers and irritants are known to occur, were among those in which workers had an increased risk for long-term respiratory disease. PMID- 11103837 TI - Differences between work methods and gender in computer mouse use. AB - OBJECTIVES: The aim of this study was to investigate whether gender or different methods of operating a computer mouse have an effect on performance and musculoskeletal load in the use of a computer mouse. METHODS: Thirty experienced computer mouse users, 15 men and 15 women, participated in the study. Electromyography (right first dorsal interossei, right extensor digitorum and right and left trapezius), a force-sensing mouse, and subjective ratings were used to register muscular load. An electrogoniometer was used to register the wrist movements. The subjects worked with 3 different methods, their own, a wrist based method and an arm-based method. Gender comparisons were made when the subjects used their own method. RESULTS: The women worked with greater extension and range of motion and tended to work with a greater ulnar deviation of the wrist. They also applied higher forces to the mouse when expressed as a percentage of a maximum voluntary contraction and had higher muscular activity in the right extensor digitorum. When using the arm-based method, the subjects worked with greater wrist extension, had higher muscular activity in the right and left trapezius muscles, and had the highest ratings of perceived exertion in the neck and shoulder. The wrist-based method resulted in higher forces being applied to the sides of the mouse and the highest ratings of perceived exertion in the wrist and hand-fingers. CONCLUSIONS: Gender differences were found for musculoskeletal load, and for most of the measured variables the women worked with higher loads than the men. The work method affected performance and musculoskeletal load. Finally, subjective measures appeared to have some utility in characterizing muscular load. PMID- 11103838 TI - Measuring and characterizing force exposures during computer mouse use. AB - OBJECTIVES: The purpose of this study was to develop and validate a sampling strategy for characterizing the finger force exposures associated with computer mouse use. METHODS: Mouse forces were measured from 16 subjects (8 men, 8 women), on 3 separate days, at their actual workstations while they performed (i) their regular work, (ii) a battery of standardized tasks, and (iii) simulated mouse use. RESULTS: The forces applied to the mouse did not vary between hours or days. During regular work, the mouse was used 78.0 (SD 40.7) times per hour, accounting for 23.7 (SD 9.5)% of the worktime. The mean forces applied to the sides and button of the mouse were low, averaging 0.6 % (0.35 N) and 0.8 % (0.43 N) of the maximal voluntary contraction, respectively. The forces applied to the mouse during the standardized tasks differed from the regular work forces; however, there were moderate-to-strong correlations between the 2 measures. CONCLUSIONS: With respect to performing exposure assessment studies, the 3 major findings were (i) mouse force measurements should be made while subjects perform their actual work in order to characterize the absolute applied force accurately, (ii) the forces applied to the mouse during the performance of a short battery of standardized tasks can be used to characterize relative exposure and identify computer operators or work situations for which higher forces are applied to the mouse, and (iii) subjects cannot accurately simulate mouse forces. PMID- 11103839 TI - Misclassification of physical work exposures as a design issue for musculoskeletal intervention studies. AB - OBJECTIVES: This study determined the impact of misclassification due to using job titles as surrogate variables for physical work exposures to assess confounding in a study of the preventive effect of back belts on back injury. The authors present retail merchandise data that quantify misclassification from residual confounding by physical work exposures on injury rate ratios when available administrative job titles are used. METHODS: Job title and direct observation data on 134 workers were used to calculate the percentage to which the job-title-adjusted rate ratio for back injury accounts for confounding by the true physical work exposures, awkward postures, and heavy weight handling. Workers' compensation data, an estimate of the effect of back belts from the literature, and the percentage of adjustment of the rate ratio due to the job title variable were used to calculate the magnitude of bias from the rate ratio adjusted for job title. RESULTS: The job title variable was found to have sensitivities of 97% and 85% and specificities of 68% and 58% for awkward postures and heavy weight handling, respectively. The magnitude of confounding bias remaining for the back-injury rate ratio when the job title surrogate was used was 24% for postures and 45% for heavy weight handling. CONCLUSIONS: The administrative job title performed poorly in this setting; residual confounding was sufficient to bias the rate ratio from 2.0 to 1.3. The effect of additional sources of misclassification and the need for better exposure measures than job title are discussed. PMID- 11103840 TI - Standing at work and varicose veins. AB - OBJECTIVES: This study attempts to determine whether or not prolonged standing at work involves an excess risk for the occurrence of varicose veins. METHODS: A cohort of 1.6 million 20-to-59-year-old Danes gainfully employed in 1991 were followed for 3 years according to first hospitalization due to varicose veins of the lower extremities. The exposure data came from a representative sample of the baseline population. Altogether 5940 people were interviewed about occupational exposure and confounding factors. RESULTS: For men working mostly in a standing position, the risk ratio for varicose veins was 1.85 [95% confidence interval (95% CI) 1.33-2.36] in a comparison with all other men. The corresponding risk ratio for women was 2.63 (95% CI 2.25-3.02). The results were adjusted for age, social group, and smoking. CONCLUSIONS: Working in a standing position is associated with subsequent hospitalization due to varicose veins for both men and women. PMID- 11103841 TI - Effects of shift work on 24-hour ambulatory blood pressure and its variability among Japanese workers. AB - OBJECTIVES: This study examined the effects of rotating shift work on blood pressure in a comparison of ambulatory blood pressure and long-term changes in blood pressure between shift and day workers. METHODS: Ambulatory blood pressure was measured for 24-hour periods at an interval of 30 minutes for 27 shift workers and 26 day workers when they worked during the day. Blood pressure was compared between these 2 groups of workers for 4 time categories (awake, sleep, nonwork awake, and work periods). Their long-term blood pressures, recorded in annual surveys, were reviewed for long-term changes. These comparisons were adjusted for the effects of body mass index, alcohol intake, anger expression, and physical activity. RESULTS: On the average, sleep time was shorter and the anger-in (ie, anger suppressed) score was higher for the shift workers than for the day workers, but body mass index and alcohol intake did not differ between the 2 groups. Even after adjustment for these co-variables, the mean systolic blood pressure during the 24-hour, awake, and work periods were higher among the shift workers than among the day workers. The 24-hour standard deviations of the systolic blood pressures were also higher for the shift workers than for the day workers. Among the shift workers, but not among the day workers, a significant long-term increase was observed in systolic blood pressure measured in the annual surveys. CONCLUSIONS: These results suggest that shift work may increase systolic blood pressure levels among Japanese men. PMID- 11103842 TI - Renal and immunologic markers for chloralkali workers with low exposure to mercury vapor. AB - OBJECTIVES: The aim of this study was to investigate renal function and immunologic markers among chloralkali workers with long-term low exposure to mercury vapor. METHODS: Forty-seven currently exposed workers were compared with reference workers matched for age in a cross-sectional design. RESULTS: The mean urinary mercury concentration was 5.9 (range 1.1-16.8) nmol/mmol creatinine (Cr) for the exposed workers and 1.3 (range 0.2-5.0) nmol/mmol Cr for the referents. The chloralkali workers had been exposed for an average of 13.3 (range 2.8-34.5) years. The activity of N-acetyl-beta-D-glucosaminidase in urine (U-NAG) was higher in the exposed workers (mean 0.18 U/mmol Cr versus 0.14 U/mmol Cr, P=0.02). Associations between current urinary mercury, cumulative urinary mercury, and cumulative urinary mercury per year (intensity) and U-NAG, autoantibodies to myeloperoxidase (anti-MPO) and proteinase 3 in serum, respectively, were observed. The activity of U-NAG and anti-MPO was increased in the workers with the highest exposure, as assessed by their mean intensity of exposure. The highest activity of U-NAG was observed in the exposed workers with the lower concentrations of selenium in whole blood. CONCLUSIONS: The study indicates an effect of exposure on the kidney proximale tubule cells, possibly modified by individual selenium status, and an effect mediated by neutrophil granulocytes. PMID- 11103843 TI - Chromosome aberrations in pesticide-exposed greenhouse workers. AB - OBJECTIVES: The aim of this study was to investigate the possibility of subtoxic exposure to pesticides causing chromosome aberrations in greenhouse workers. METHODS: In a cross-sectional and prospective study design chromosome aberration frequencies in cultured lymphocytes were examined for 116 greenhouse workers exposed to a complex mixture of almost 50 insecticides, fungicides, and growth regulators and also for 29 nonsmoking, nonpesticide-exposed referents. RESULTS: The preseason frequencies of chromosome aberrations were slightly but not statistically significantly elevated for the greenhouse workers when they were compared with the referents. After a summer season of pesticide spraying in the greenhouses, the total frequencies of cells with chromosome aberrations were significantly higher than in the preseason samples (P=0.02) and also higher than for the referents (P=0.05). This finding was especially due to an increased number of cells with chromatid gaps between the first and second samples (P=0.001). The results may reflect an additive genotoxic effect of the spraying season, for which the use of insecticides and growth regulators (but not fungicides) culminates. The highest elevation in the risk of chromatid gaps was observed for persons who did not use gloves during re-entry activities such as nipping, cutting, pricking, and potting (risk ratio 2.88, 95% confidence interval 1.63-5.11). CONCLUSIONS: The present results suggest a genotoxic effect from a complex subtoxic occupational pesticide exposure. In general, the findings indicate the importance of personal protection, during high-exposure re-entry activities, in preventing pesticide uptake and genetic damage. PMID- 11103844 TI - Evidence-based primary prevention? PMID- 11103845 TI - International expert meeting on new advances in the radiology and screening of asbestos-related diseases. PMID- 11103846 TI - Physiological issues surrounding the performance of adolescent athletes. AB - More than ever, many young athletes are being encouraged to train intensely for sporting competitions from an early age. Compared with studies in adults, less is known about the physiological trainability of adolescents. The velocity of physical growth during the adolescent years makes research with a group of young athletes particularly difficult. The purpose of this review is to discuss a number of physiological issues that surround the performances of the adolescent athlete. Research has highlighted the role of growth hormone (GH) in the abrupt acceleration of linear growth that occurs during adolescence. In addition, GH has been shown to be sensitive to exercise following short term intervention studies. The reduced anaerobic power of the adolescent athlete compared with that of an adult athlete has been attributed to the intrinsic properties of the muscle that are yet to be fully understood. Resistance training studies in male adolescents, and to a lesser extent female adolescents, highlight the substantial relative strength gains that can be obtained. Aerobic trainability in young boys appears to improve markedly during the adolescent years. One of the most plausible explanations for this observation is the 'trigger hypothesis' which links increased aerobic improvements in adolescence with hormonal changes and substantial growth of the cardiorespiratory and musculoskeletal systems. Studies of aerobic trainability in adolescent girls are too scarce to be conclusive. An understanding of the impact of long term intensive training on adolescent athletes is difficult to ascertain because physical stresses vary both between and within sports. There is, however, limited evidence to suggest that 'intense' training does not impair normal growth, development or maturation. Adolescent athletes who experience rapid growth as well as large increases in training volumes may be vulnerable to overuse injuries. PMID- 11103848 TI - Adaptation to a fat-rich diet: effects on endurance performance in humans. AB - The focus of this review is on studies where dietary fat content was manipulated to investigate the potential ergogenic effect of fat loading on endurance exercise performance. Adaptation to a fat-rich diet is influenced by several factors, of which the duration of the adaptation period, the exercise intensity of the performance test and the content of fat and carbohydrate in the experimental diet are the most important. Evidence is presented that short term adaptation, < 6 days, to a fat-rich diet is detrimental to exercise performance. When adaptation to a fat-rich diet was performed over longer periods, studies where performance was tested at moderate intensity, 60 to 80% of maximal oxygen uptake, demonstrate either no difference or an attenuated performance after consumption of a fat-rich compared with a carbohydrate-rich diet. When performance was measured at high intensity after a longer period of adaptation, it was at best maintained, but in most cases attenuated, compared with consuming a carbohydrate-rich diet. Furthermore, evidence is presented that adaptation to a fat-rich diet leads to an increased capacity of the fat oxidative system and an enhancement of the fat supply and subsequently the amount of fat oxidised during exercise. However, in most cases muscle glycogen storage is compromised, and although muscle glycogen breakdown is diminished to a certain extent, this is probably part of the explanation for the lack of performance enhancement after adaptation to a fat-rich diet. PMID- 11103849 TI - Injury prevention in women's gymnastics. AB - The most serious problem faced by contemporary gymnasts is injury. Given that prevention is superior to treatment, can the gymnastics community and the scientific and medical community do a better job at injury prevention? Most research in gymnastics has been descriptive in nature. Injury prevention ultimately requires that one can predict the outcome of certain activities and their injurious nature. Making such predictions requires a knowledge of the scientific and medical aspects of injury, but more than that, one must have an intimate knowledge of the sport. Injury prevention efforts must be firmly grounded in science and medicine while making pragmatic linkages to gymnastics as it exists and is practiced. This article attempts to bridge the gap between the scientific and medical community and what actually happens in gymnastics. PMID- 11103850 TI - Typical error versus limits of agreement. AB - We have shown that the SEM (typical error) and LOA are very similar when defined at the same level of abstraction. The calculation of these sample statistics does not depend on sample size, but the precision of their estimate for the population parameter does. Only the latter concept involves the t-statistic. They do differ in the type of measurement error that is described (true score error versus test retest error) and the coverage probability of the reference interval (0.6 versus 0.95). Bland and Altman and Atdinson and Nevill have promoted the citation of either the SEM or the LOA to help researchers in their discussion of the impact of error to real uses of the measurement tol. What is vital in this discussion is the researcher having a thorough understanding of the underlying theory behind the measurement error statistic(s) that is/are employed, especially the definition of error and the coverage probability that is selected. Such issues have been built into the title of the 95% LOA statistic, and are also an inherent part of SEM. Whilst the concept of typical error appears to be easy to understand and teach, this is only because the underlying theory and definition of what the statistic actually represents is not communicated. Only if all researchers adopt one single statistic (e.g. typical error) for measurement error is it remotely possible to push underlying theory into the background, since there would be a baseline of comparison for all. Because this scenario is highly unlikely, it is important that any measurement error statistic is well defined and understood by researchers. PMID- 11103847 TI - Insulin resistance with aging: effects of diet and exercise. AB - Insulin resistance, a reduction in the rate of glucose disposal elicited by a given insulin concentration, is present in individuals who are obese, and those with diabetes mellitus, and may develop with aging. Methods which are utilised to measure insulin sensitivity include the hyperinsulinaemic-euglycaemic and hyperglycaemic clamps and the intravenous glucose tolerance tests. Several hormones and regulatory factors affect insulin action and may contribute to the insulin resistance observed in obesity. In addition, abnormal free fatty acid metabolism plays an important role in insulin resistance and the abnormal carbohydrate metabolism seen in individuals who are obese or diabetic. Thus, the mechanisms underlying the development of insulin resistance are multifactorial, and also involve alterations of the insulin signalling pathway. Aging is associated with an increase in bodyweight and fat mass. Not only is abdominal fat associated with hyperinsulinaemia but visceral adiposity is correlated with insulin resistance as well. Modifications of the changes in body composition with aging by diet and exercise training could delay the onset of insulin resistance. Weight loss and aerobic and resistive exercise training result in losses of total body fat and abdominal fat. Several studies report that bodyweight loss increases insulin sensitivity and improves glucose tolerance. In addition, the insulin resistance observed in aged persons can be modified by physical training. Longitudinal studies indicate significant improvements in glucose metabolism with aerobic exercise training in middle-aged and older men and women. Moreover, the improvements in insulin sensitivity with resistive training are similar in magnitude to those achieved with aerobic exercise. The improvements in glucose metabolism after bodyweight loss and exercise training may in some cases be partially attributed to changes in body composition, including reductions in total and central body fat. Yet, additional changes in skeletal muscle, blood flow and other mechanisms likely interact to modify insulin resistance with exercise training. Lifestyle modifications including bodyweight loss and physical activity provide health benefits and functional gains and should be promoted to increase insulin sensitivity and prevent glucose intolerance and type 2 diabetes mellitus in older adults. PMID- 11103851 TI - The rainbow trout (Salmo gairdneri Richardson) granular leukocytes and mast-cells system versus the human one. Cytomorphometrical and cytochemical characters in these two species philogenetically poles apart. AB - In man, circulating granular leukocytes constitute a cellular system and are able to migrate in the tissues to take part in the immune reactions. This study was to characterize the granular leukocytes and the eventual existence of mast-cells of a low vertebrate, rainbow trout (Salmo gairdneri R.) in order to compare it with man. The blood smears and tissues sections have been tested whit panoptic methods, cytochemistry and toluidine blue. White blood cell count, leukocytes formula and cytomorphometric characterization was performed using an image analyser. Scanning of tissues sections in order to identify mast-cells has been also performed. In this model the granular leukocytes are all neutrophilic like; no eosinophilic, no basophilic no tissue mast-cells, basically existing in allergic and anaphylactic reactions, were found. PMID- 11103852 TI - The ameloblast movement in rat incisor. L.M., S.E.M. and C.L.S.M. study. AB - The internal epithelium of enamel organ and the below enamel surface during growth of the lower incisor, were examinated in ten Wistar rat 12-27 weeks old and weighing between 150/200 gr, by means of immuno histochemical, light and scanning electron microscopy techniques. Our specimens indicate that during the outer enamel secretion the anti-actin positivity goes from distal terminal web to infra nuclear region of cell body. The results of the present study do not support the active movement hypothesis, conversely they support the Warshawsky (1992) hypothesis, i.e. the distal terminal web permits the maintenance and the assembling of ameloblasts during enamel growth. Hence we do agree with Osborn (1970) who reported that, during secretion, ameloblasts move passively in response to secretory forces. PMID- 11103853 TI - Light microscopic analysis of cellular networks in the pineal gland of the golden hamster as revealed by methylene blue labeling. AB - Pineal glands of golden hamsters were stained transcardially with methylene blue and studied in paraffin sections by light microscopy. In the superficial portion of the pineal complexes, a selective labeling of a subpopulation of polymorphic cells with processes of different length was achieved. It could not be clarified, whether these cells form a syncytium or are only in contact with each other. Although functional interpretations of the morphological results have to be drawn with great care, the investigation might be of special interest for a better understanding of the well-known inhibiting effects of the dye to soluble guanylyl cyclase and nitric oxide synthase within this neuroendocrine gland. PMID- 11103854 TI - Extra cellular matrix features in human meninges. AB - We collected human fetal and adult normal meninges to relate the age of the tissue with the presence of collagenous and non-collagenous components of Extra Cellular Matrix (ECM). Immunohistochemistry led us to observe some differences in the amount and in the distribution of these proteins between the two sets of specimens. In particular, laminin and tenascin seem to be expressed more intensely in fetal meninges when compared to adult ones. In order to investigate whether the morphofunctional characteristics of fetal meninges may be represented in pathological conditions we also studied meningeal specimens from human meningiomas. Our attention was particularly focused on the expression of those non-collagenous proteins involved in nervous cell migration and neuronal morphogenesis as laminin and tenascin, which were present in lesser amount in normal adult specimens. Microscopical evidences led us to hipothesize that these proteins which are synthesized in a good amount during the fetal development of meninges can be newly produced in tumors. On the contrary, the role of tenascin and laminin in adult meninges is probably only interesting for their biophysical characteristics. PMID- 11103855 TI - Localization, morphology and ultrastructure of taste buds in the domestic duck (Cairina moschata domestica L.) oral cavity. AB - The Authors report on localization of the taste buds in epithelium of the palate (70%), the floor of the oral cavity (28%) and the tongue (2%) in Domestic Duck. Each taste bud is oval-shaped, measuring roughly 130 x 60 microm, and communicates with the oral cavity by a short duct. There is topographical correspondence between the buds and the parietal salivary glands. Ultrastructural examination showed the following 4 cell types: 1) light cells--characterized by cytoplasm containing sparse perinuclear filaments, numerous light vesicles, a uniformly granular nucleus. 2) dark cells--with electron-dense cytoplasm, numerous perinuclear filaments, occasional light vesicles, an irregular and densely granular nucleus. 3) intermediate cells--with characteristics common to the two previous cell types. 4) basal cells--located at the ventral part of the button. PMID- 11103856 TI - Anatomical variations of the ulnar and median nerves in the upper limb. AB - The aim of our study was the evaluation of the anatomy of ulnar and median nerves in the upper limb in order to ameliorate knowledge on the clinical anatomy of these nerves. In fact, further information on this topic may be useful owing to its possible clinical relevance when planning surgical anatomy and reconstructive surgery in tumor affected and injured patients. The relationships between ulnar and median nerve and neighbouring anatomical structures have been examined, together with the course and ramification of the ulnar and median nerves in six fresh cadavers. Moreover, we have performed a review of the literature. Four specific aspects were evaluated during dissection: 1) division modality of the ulnar nerve at the wrist; 2) anatomical details of the medial humeral epicondyle; 3) anatomical relationships between median nerve and retinaculum flexorum; 4) median-ulnar nerves anastomosis. Our results show that: the medial humeral epicondyle shows specific anatomical details in relation to the ulnar nerve; the relationships between the median nerve and the transverse carpal ligament may be characterized by one or two nerve trunks (two cases of bifid median nerve in our experience); median-ulnar nerve anastomosis may be also found at various levels. Comparing our results with those of the available literature we can conclude that anatomical variations of ulnar and median nerve in the upper limb are not an infrequent finding and their clinical, diagnostic and surgical relevance should be considered. PMID- 11103857 TI - Stroke: pathogenesis, investigations, and prognosis--Part II of III. AB - The aim of this review is to present the current knowledge regarding stroke. It will appear in three parts (in part I the epidemiology, clinical picture, and risk factors were discussed, while part III will consist of the management and rehabilitation). In the present part (II) the pathogenetic and pathophysiologic aspects of stroke are described. Regarding the investigations apart from the history and clinical examination and general investigations, the following specialized investigations and their role are discussed in detail: Computed tomography (CT), magnetic resonance imaging (MRI), xenon-blood-flow, positron emission tomography (PET), cerebral angiography, magnetic resonance angiography (MRA), ultrasonography, transcranial Doppler (TCD), echocardiography, Holter monitoring, and biopsies. In addition, taking into account the information from the above-cited modalities a prognosis for the final outcome is presented. PMID- 11103858 TI - Evaluation of electron beam tomographic coronary arteriography with three dimensional reconstruction in healthy subjects. AB - In this study, the authors evaluated the performance characteristics of contrast enhanced electron-beam tomography (EBT) with three-dimensional reconstruction in defining the coronary artery lumen in healthy subjects. Thirty patients with normal coronary angiograms by selective coronary arteriography (SCA) underwent contrast-enhanced EBT examination. Measured parameters included degree of luminal enhancement, intravascular contrast-to-noise ratio (CNR), and diameter and length of visualized lumen. Ventricular cavity, aortic blood pool, and coronary artery attenuation were found to be significantly different before and after intravenous injection of contrast material (p < 0.001). CNR decreased from proximal to distal segments within each vessel (p < 0.001), with a peak of 11.2 +/- 2.3 occurring in the proximal left anterior descending coronary artery (LAD) to a low of 4.8 +/- 2.0 in the distal left circumflex (LCX). Luminal diameters visualized by EBT had no significant difference with that of SCA (p > 0.05). Therefore, EBT angiography with three-dimensional reconstruction allows for noninvasive coronary arteriography revealing long segments of the major coronary arteries in normal subjects. PMID- 11103859 TI - Differences in beat-to-beat variability of the QT interval between day and night. AB - The objective of this study was to evaluate the beat-to-beat variability of the QT interval during the day and night. A new algorithm was used to detect the onset of the QRS, the apex of the T wave, and end of the T in ambulatory electrocardiographic recordings. Beat-to-beat variability of QT, QaT, and QTc during the day and night was studied in the time, frequency, and chaotic domains. Participants were adults without clinical evidence of heart disease. Although the QT duration was higher (p = 0.0001) at night, the beat-to-beat variability of this interval was lower: in the time domain (decreased standard deviation, p = 0.0005), in the frequency domain (decreased low-frequency power of the spectra, p = 0.004), and the chaotic domain (tighter clustering of the points in the Poincare plots). The high-frequency to low-frequency ratio of the power spectra of the QT (and the RR) was higher (p = 0.03) at night. Beat-to-beat QT variability in the time, frequency, and chaotic domains is decreased at night with shift of the QT modulation to higher frequencies corresponding to respiration and representing vagal preponderance. The techniques presented here and the findings in normal subjects may be useful in evaluating the risk for arrhythmic events in patients with heart disease. PMID- 11103860 TI - Short- and long-term results of abciximab versus aspirin in conjunction with thrombolysis for patients with peripheral occlusive arterial disease and arterial thrombosis. AB - Acute peripheral occlusive arterial disease is an important cause of morbidity and mortality, particularly among older persons. Catheter-directed thrombolytic therapy is the treatment of choice but has limitations: long lytic times, occlusions refractory to thrombolysis, and a high rate of restenosis. We conducted a pilot study to evaluate the use of the platelet GP IIb/IIIa receptor antagonist abciximab versus aspirin in conjunction with thrombolysis in patients with acute peripheral occlusive arterial disease associated with arterial thrombosis. A total of 84 patients were randomized into two equal groups to receive 5 mg recombinant tissue plasminogen activator intravenously and 500 IU heparin/hour along with either 500 mg acetylsalicylic acid or a bolus of 0.25 mg/kg abciximab followed by 10 microg/min abciximab over 12 hours (heparin reduced to 250 IU/hour). Primary efficacy criteria included the number of rehospitalizations, reinterventions, and amputations during the following 6 months. Secondary endpoints were the changes in the Fontaine stage, Bollinger index (vessel occlusion), ankle-to-brachial ratios, distance to claudication after 6 months, and the duration of the initial local lysis treatment. Adjunctive use of abciximab reduced the rates of rehospitalization, reinterventions, and amputations versus results with the use of aspirin (10 vs 14 occurrences, respectively; 9 vs 11; 3 vs 5; when summed, intergroup difference p < 0.05). Secondary peripheral occlusive arterial disease variables became highly significant versus aspirin (p < 0.001 or greater) at 3 and 6 months after treatment. The duration of lysis was markedly shorter upon addition of abciximab versus aspirin (75 vs 110 min; p < 0.001). No major bleeding complications or embolisms occurred. These preliminary results indicate that abciximab may have a useful role when used adjunctively with a thrombolytic agent in older persons with acute peripheral occlusive arterial disease and arterial thrombosis. PMID- 11103861 TI - Evaluation of dysrhythmia in children with muscular dystrophy. AB - Myocardial involvement and dysrhythmia are common findings with muscular dystrophy and are among the leading causes of death. The authors evaluated rhythm and conduction abnormalities in children with muscular dystrophy by electrocardiography, signal-averaged electrocardiography, and Holter monitoring. Twenty-nine patients (mean age, 8 years) and 29 healthy control subjects were included in the study. Sixty-two percent of patients had electrocardiographic abnormalities defined as deep Q waves in V6, tall R waves in V1, and QRS axis deviation. The cardiomyopathy index was significantly greater in the patient group whereas QT and QTc dispersion values showed no significant difference. Holter monitoring revealed premature atrial and ventricular contractions more frequently than normal. However all were classified as Lown I and II. Mean heart rate was significantly higher in the patient group. The electrocardiograms of 41% of the patients showed late potentials. No relationship with these changes and cardiac function was observed. During the study, one patient died whose cardiomyopathy index was longer and had late potentials detected with signal averaged electrocardiography. In conclusion, standard electrocardiography, cardiomyopathy index, signal-averaged electrocardiography, and Holter monitoring are valuable and reliable monitoring methods in children with muscular dystrophy. PMID- 11103862 TI - Aortic distensibility is closely related to the progression of left ventricular hypertrophy in patients receiving hemodialysis. AB - Aortic stiffening and left ventricular hypertrophy are believed to be major determinants for the prognosis of patients with end-stage renal disease. However, the relationship between left ventricular hypertrophy and aortic stiffness remains to be determined. Echocardiographically determined parameters and aortic distensibility determined with cine magnetic resonance were evaluated in 21 patients undergoing chronic hemodialysis. Hemodynamic variables measured at the beginning of the study were compared with those measured after 28 months. Aortic distensibility determined at the descending aorta was markedly lower in patients undergoing hemodialysis than in healthy control subjects. During the follow-up period, blood pressure and hemodynamic variables, including left ventricular mass index, remained unchanged. However, multiple regression analysis indicated that aortic distensibility independently contributed to the left ventricular mass index and to the change in left ventricular mass index between baseline and after 28 months. Baseline left ventricular mass index negatively correlated to aortic distensibility (r = -0.74, p < 0.0001), and the changes in left ventricular mass index positively correlated to aortic distensibility (r = 0.52, p < 0.05). Our study demonstrates that aortic distensibility at the descending aorta is a predictable marker for the development or regression of left ventricular hypertrophy. Therefore, patients with end-stage renal disease must be treated with appropriate drugs to improve aortic distensibility. PMID- 11103863 TI - Effects of perindopril on left ventricular remodeling and aortic regurgitation in rats assessed by echocardiography. AB - This study investigated the effect of the angiotensin-converting enzyme (ACE) inhibitor perindopril on left ventricular (LV) remodeling and cardiac function in rats with aortic regurgitation (AR). Twenty male Sprague-Dawley rats in which AR was produced by closed-chest aortic valve puncture were divided into untreated and perindopril-treated (5 mg/kg/day) rats. Ten control rats were sham-operated. Blood pressure, body weight, and echocardiographic recordings were followed every 2 weeks for a period of 12 weeks. LV dimension (LVD) and fractional shortening (FS) were calculated. The heart was finally excised for weight, hydroxyproline measurements, and histopathology. At 12 weeks, end-diastolic LVD increased in untreated rats (10.8 +/- 0.2 mm) but did not dilate in treated rats (9.6 +/- 0.3 mm, p < 0.01 vs untreated rats). FS decreased from 6 weeks in untreated rats (27.3 +/- 0.9% at 12 weeks), but did not change in treated rats (33.8 +/- 0.5%, p < 0.001). The ratio of LV weight to body weight in untreated rats (2.62 +/- 0.11 mg/g, p < 0.05) was higher than in sham-operated rats (1.52 +/- 0.02 mg/g) and than in treated rats (1.95 +/- 0.07 mg/g). Interstitial collagen accumulation histopathologically increased in untreated rats and was inhibited in treated rats. LV collagen of untreated rats (1.46 +/- 0.08 mg/100 mg, p = 0.03) was higher than those of treated (1.08 +/- 0.09 mg/100 mg) and sham-operated rats (1.06 +/- 0.14 mg/100 mg). Perindopril inhibited LV remodeling induced by volume overload and preserved LV function in AR rats. PMID- 11103864 TI - Temporal arteritis and fever: report of a case and a clinical reanalysis of 360 cases. AB - The purposes of this article are to report a case with temporal arteritis (TA) and to summarize and reanalyze the cases of temporal arteritis associated with fever in published articles for understanding better the clinical features of TA. A case with biopsy-proven TA is reported. The publications with TA and fever were searched by using MEDLINE in English from 1966 to 1999. Three hundred sixty cases of temporal arteritis associated with fever were reanalyzed. The results showed that a case of biopsy-proven TA with typically clinical manifestation was initially misdiagnosed and that the reanalysis of 360 cases revealed that the common clinical findings at presentation were abnormal temporal arteries, headache, low fever, loss of weight, polymyalgia rheumatica, jaw claudication, vision disorder, arthralgis or myalyias, and ear pain and that the uncommon clinical findings at presentation were high fever, malaise, anorexia, breast pain, transient ischemic attack/stroke, cough, mental disorder, diarrhea, and uterine prolapse, etc. Laboratory findings were the range of erythrocyte sedimentation rate (ESR) 14 to 149 with a mean of 97.0 mm/hr, white blood cells being normal or increased in the range of 10.9 to 22.9 x 10(9)/L, hemoglobin level 7 to 16 g/dL, the platelets count increased to 785 x 10(9)/L, and microscopic hematuria. The diagnosis was made by a combination of clinical features, an increased ESR, a response to steroids, and, most specifically, temporal artery biopsy. The initial diagnosis was misdiagnosed in 38.2% of patients. In conclusion, the features of TA associated with fever have not been widely appreciated yet. TA is a common cause of fever of unknown origin (FUO) in the elderly. TA should be considered when patients complain of common and uncommon manifestations. An elevated ESR will aid in the diagnosis of TA, and temporal artery biopsy will provide certainty. PMID- 11103865 TI - Embolic stroke secondary to an aortic arch tumor--a case report. AB - An acute stroke from an aortic arch tumor is reported. These tumors are rare and have to be differentiated from atheromas. Aortic atheromas commonly present with embolic phenomena and occasionally as masses. Aortic tumors are more likely to produce obstructive phenomena, presenting as a coarctation or dissection. Magnetic resonance imaging with gadolinium can facilitate the diagnosis. A literature review of aortic masses and their diagnosis and treatment are presented. PMID- 11103866 TI - Superior vena cava syndrome due to permanent transvenous pacemaker electrodes: successful treatment with combined thrombolysis and angioplasty--a case report. AB - Superior vena cava syndrome is a rare complication of permanent transvenous pacing electrodes. Multiple treatment options are available, namely thrombolytics, venoplasty, stenting, surgery, and combinations of the above, yet initially the optimal approach is uncertain. Whether plain balloon angioplasty provides durable and satisfactory long-term results is equally uncertain. The authors report a patient treated with a combination of local thrombolytic therapy and balloon venoplasty with good long-term outcome at two years of follow-up. PMID- 11103867 TI - Transition from classic aortic dissection to aortic intramural hemorrhage--a case report. AB - A 64-year-old man was hospitalized with chief complaints of chest and back pain. A diagnosis of Stanford type A aortic dissection with a false lumen extending from the ascending to the descending aorta was made based on the results of computed tomography (CT). A CT obtained the following day showed resolution of the false lumen and increased brightness of the aortic wall, typical of aortic dissection with intramural hemorrhage. Although previous studies have described a gradual transition from aortic intramural hemorrhage to aortic dissection with a false lumen, there are no reports of the transition from an aortic dissection with a false lumen to the intramural hemorrhage type of aortic dissection. This patient is of interest when considering the pathogenesis of aortic dissection with intramural hemorrhage and the relationship between the intramural hemorrhage and false-lumen types of aortic dissection. PMID- 11103868 TI - Development and applications of a model for cellular response to multiple chemotactic cues. AB - The chemotactic response of a cell population to a single chemical species has been characterized experimentally for many cell types and has been extensively studied from a theoretical standpoint. However, cells frequently have multiple receptor types and can detect and respond chemotactically to more than one chemical. How these signals are integrated within the cell is not known. and we therefore adopt a macroscopic phenomenological approach to this problem. In this paper we derive and analyze chemotactic models based on partial differential (chemotaxis) equations for cell movement in response to multiple chemotactic cues. Our derivation generalizes the approach of Othmer and Stevens [29], who have recently developed a modeling framework for studying different chemotactic responses to a single chemical species. The importance of such a generalization is illustrated by the effect of multiple chemical cues on the chemotactic sensitivity and the spatial pattern of cell densities in several examples. We demonstrate that the model can generate the complex patterns observed on the skin of certain animal species and we indicate how the chemotactic response can be viewed as a form of positional indicator. PMID- 11103869 TI - A self-consistent cell flux expression for simultaneous chemotaxis and contact guidance in tissues. AB - During wound healing, both chemotaxis and contact guidance can contribute to the migration of blood and tissue cells to the wound. In order to understand the wound healing process, we must thus understand how cells respond to both these simultaneous directional cues, which are not necessarily coaligned. Although chemotaxis and contact guidance have been studied individually, the interaction between them has not been addressed. We extend a stochastic cell movement model, developed by Dickinson and Tranquillo (1995) [6] for individual cues, for simultaneous chemotaxis and contact guidance by a two-parameter perturbation analysis in terms of the two associated cues, a chemotactic factor gradient and aligned tissue fibers. We present results from analysis of the first-order perturbation, which includes the cell flux expression heuristically proposed by others, but reveals paradoxical results for other indices of cell movement, such as the mean-squared displacement. We then present second-order perturbation results that resolve these paradoxical results. Finally, we relate these results to a continuum mechanical model developed by Barocas and Tranquillo (1997) [3] that predicts fiber alignment due to cell traction induced tissue contraction. PMID- 11103870 TI - A mathematical analysis for the Brownian dynamics of a DNA tether. AB - In the single-particle tracking experiment, the internal motion of a single DNA or polymer molecule whose one end is attached to a microsphere (optical marker) and the other end is anchored to a substratum is studied (Finzi and Gelles, 1995). The stochastic Brownian dynamics of the sphere reflect the spontaneous fluctuations, thus the physical characteristics, of the DNA or polymer molecule (Qian and Elson, 1999, Qian, 2000). In this paper, two continuous models of polymer molecules, a flexible elastic string and a weakly bendable elastic rod, are analyzed. Both models are cast mathematically in terms of linear stochastic differential equations. Based on Fourier analyses, we calculate the mean square displacement (MSD) of the particle motion, the key observable in the experiment. We obtain for both models the short-time asymptomatics for the MSD, as well as the long-time behavior in terms of the smallest non-zero eigenvalues. It is shown that: (i) the long-time dynamics of continuous elastic string model quantitatively agree with that of the discrete bead-spring model. (ii) The short time MSD of both models are controlled by the tethered particle, with linear dependence on t. (iii) The two models show characteristic difference for long time behavior: The longest relaxation time is proportional to L2 for long elastic string and to L for short elastic string, but is proportional to L4 for both long and short weakly bendable rod. PMID- 11103871 TI - Drug-resistant parasites and aggregated infection--early-season dynamics. AB - We investigate the effect of spatial aggregation in the infection dynamics of nematode parasites in ruminants. We show that a high degree of spatial aggregation is likely to lead to a dramatically enhanced rate of invasion by drug resistant strains. PMID- 11103872 TI - A dynamic energy budget model based on partitioning of net production. AB - We formulate a Dynamic Energy Budget (DEB) model for the growth and reproduction of individual organisms based on partitioning of net production (i.e. energy acquisition rate minus maintenance rate) between growth and energy reserves. Reproduction uses energy from reserves. The model describes both feeding and non feeding stages, and hence is applicable to embryos (which neither feed nor reproduce), juveniles (which feed but do not reproduce), and adults (which commonly both feed and reproduce). Embryonic growth can have two forms depending on the assumptions for acquisition of energy from yolk. By default, when the energy acquisition rate exceeds the maintenance rate, a fixed proportion of the resulting net production is spent on growth (increase in structural biomass), and the remaining portion is channelled to the reserves. Feeding organisms, however, modulate their allocation of net production energy in response to their total energy content (energy in the reserves plus energy bounded to structural biomass). In variable food environment an organism alternates between periods of growth, no-growth, and balanced-growth. In the latter case the organism adopts an allocation strategy that keeps its total energy constant. Under constant environmental conditions. the growth of a juvenile is always of von Bertalanffy type. Depending on the values of model parameters there are two long-time possibilities for adults: (a) von Bertalanffy growth accompanied by reproduction at a rate that approaches zero as the organism approaches asymptotic size, or (b) abrupt cessation of growth at some finite time, following which, the rate of reproduction is constant. We illustrate the model's applicability in life history theory by studying the optimum values of the energy allocation parameters for constant environment and for each of the dynamic regimes described above. PMID- 11103873 TI - Animal models of deficient sensorimotor gating: what we know, what we think we know, and what we hope to know soon. AB - Sensorimotor gating of the startle reflex can be studied in humans and laboratory animals using measures of prepulse inhibition (PPI) of the startle reflex. PPI is reduced in patients with specific neuropsychiatric disorders and in rats after manipulation of the limbic cortex, striatum, pallidum or pontine tegmentum. Studies are rapidly identifying the neurochemical and neuroanatomical substrates regulating PPI in laboratory animals; this detailed circuit information has been used as a 'blueprint' to identify possible candidate substrates responsible for PPI deficits in psychiatrically disordered humans. In parallel, studies have also begun to assess the homology of pharmacological effects on PPI across species, as an initial step towards translating detailed neural circuit information from rats to humans. Despite this rapid progress, there is an increasing danger of overlooking important methodological and interpretative issues that could impact either positively or negatively on the ultimate utility of models based on measures of PPI. Some of these issues--ranging from the cross-species methods for quantifying specific variables to the relevance of genetic drift to animal and human studies of PPI--and their implications for future studies are the focus of this review. PMID- 11103874 TI - Can animal models help to understand human diseases? Commentary on Swerdlow et al., 'Animal models of deficient sensorimotor gating: what we know, what we think we know, and what we hope to know soon'. PMID- 11103875 TI - A human perspective: commentary on Swerdlow et al., 'Animal models of deficient sensorimotor gating: what we know, what we think we know, and what we hope to know soon'. PMID- 11103876 TI - Context determines the type of sensitized behaviour: a brief review and a hypothesis on the role of environment in behavioural sensitization. AB - Behavioural sensitization to psychostimulants may develop context-dependency in certain circumstances. Animals given a stimulant repeatedly in a test cage but not in other environments may show enhanced drug-induced behaviour in the test cage. Conditioning mechanisms have been claimed to be responsible for these phenomena. However, several recent findings are not properly accounted for by conditioning. In addition, growing evidence supports the hypothesis that behavioural sensitization reflects neural changes induced by repeated exposure to psychostimulants (the pharmacological hypothesis). However, the pharmacological hypothesis itself fails to account for environmental influences. In this paper, we propose a hypothesis on the role of environment that is complementary to the pharmacological hypothesis. According to our hypothesis, environment does not have a causal role in the development of sensitization, but it modifies the mode of expression of the sensitized behaviour. Sensitization primarily reflects a neuroadaptive change induced by repeated exposure of the neural system to psychostimulants. However, psychostimulants are known to induce different behaviours in different environments. Therefore, repeated administration of a psychostimulant in different environments would result in augmentation of different behaviours. Our hypothesis potentially accommodates various previous observations. We briefly review the literature and present our hypothesis. PMID- 11103877 TI - Animal models for the negative symptoms of schizophrenia. AB - Negative or defect symptoms refer to a reduction in normal functioning. In schizophrenia, negative symptoms encompass, among others, anhedonia, flat affect, avolition and social withdrawal. These symptoms have been found to be particularly prominent in the more chronic phase of the illness and seem to be virtually insensitive to current antipsychotic treatment. This review focuses on the possibilities and limitations of animal models for the negative symptoms of schizophrenia. Following a review of the negative symptoms in schizophrenia, attention is focused on the two symptoms most often modelled in animals - anhedonia and social withdrawal. We then look at the important question of how to model schizophrenic pathology in animals. Since the exact pathology is still far from clear, most efforts have in the past concentrated on using psychotomimetic drugs such as amphetamine or phencyclidine. The recently accumulated knowledge that schizophrenia probably results from disturbances in the normal development of the brain has led to a surge of new animal models in which the long-term consequences of early manipulations are investigated. However, so far these models have predominantly concentrated on the positive rather than the negative symptoms of schizophrenia. The last part of this review is dedicated to the question of validation of animal models for anhedonia and social withdrawal. The general conclusion is that very few models have so far been adequately tested. The lack of currently effective treatment further hampers the study of such validation. PMID- 11103878 TI - Studies in animal models and humans suggesting a role of nerve growth factor in schizophrenia-like disorders. AB - Neurotrophic factors, such as nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), are known to play a crucial role in growth, differentiation and function in a variety of brain neurons during development and in adult life. We have recently shown that environmental changes, aggressive behavior and anxiety-like responses alter both circulating and brain basal NGF levels. In the present review, we present data obtained using animal models which suggest that neurotrophic factors, particularly NGF and BDNF, might be implicated in mechanism(s) leading to a condition associated with schizophrenic-like behaviors. The hypothesis that neurotrophins of the NGF family can be implicated in some maldevelopmental aspects of schizophrenia is supported by findings indicating that the constitutive levels of NGF and BDNF are affected in schizophrenic patients. PMID- 11103879 TI - Pharmacological and molecular targets in the search for novel antipsychotics. AB - The recent enthusiasm among clinicians for the so-called 'atypical antipsychotics' has both improved treatment for schizophrenic patients and provided a welcome stimulus for basic research on antipsychotic mechanisms. Even the newer drugs have shortcomings, and research is underway aimed at identifying novel agents with greater efficacy and safety. Much of this effort is directed towards compounds which, in addition to blocking dopamine receptors, also act on other neurotransmitter receptors such as 5-HT2, 5-HT1A and alpha2-adrenergic receptors. However, there is also a large amount of scientific activity seeking to discover and develop selective dopamine receptor subtype antagonists (including compounds which specifically block D3 or D4 receptors) or drugs that specifically target the dopamine autoreceptor. Finally, a number of drug development programmes are searching for non-dopaminergic antipsychotics. Drugs that do not have affinity for dopamine receptors but act through neurotensin, sigma or cannabinoid CB1 receptors or glutamatergic mechanisms are currently being evaluated. If any of these agents prove to have clinical efficacy this may lead to a third generation of antipsychotics. It is likely, however, that the mechanisms of action of such drugs will nevertheless imply the intimate involvement of dopaminergic pathways. PMID- 11103880 TI - Spatial and associative learning deficits induced by neonatal excitotoxic hippocampal damage in rats: further evaluation of an animal model of schizophrenia. AB - Neonatal ventral hippocampal lesions in the rat result in post-pubertal onset of behavioural abnormalities, modelling some aspects of schizophrenia. We further assessed the behavioural effects of neonatal lesions in rats in a variety of cognitive tasks and in the prepulse inhibition (PPI) of startle response paradigm. Prepubescent, lesioned rats exhibited startle responses and PPI similar to controls whereas, at adulthood, they showed a deficit in PPI. Lesioned rats acquired both passive and active avoidance responses. However, compared to controls, they showed a deficit in passive avoidance retention and in acquisition of active avoidance responses. In a cued Morris water-maze task, lesioned rats demonstrated adequate sensorimotor functions and appropriate motivation to escape from water. However, they were impaired in place learning and in remembering the location of a submerged platform. In conclusion, neonatal ventral hippocampal lesions result in the post-pubertal emergence of long-lasting deficits in sensorimotor gating and in the capacity to acquire and retain information in tests of spatial and avoidance learning. Therefore, this neurodevelopmental model of schizophrenia seems to exhibit an interesting degree of validity in possibly simulating some cognitive impairments and sensorimotor gating deficits frequently observed in psychotic patients. PMID- 11103881 TI - Exaggerated MK-801-induced motor hyperactivity in rats with the neonatal lesion of the ventral hippocampus. AB - Neonatal lesions of the ventral hippocampus in rats produce changes in spontaneous and pharmacologically induced dopamine-dependent behaviors that emerge in early adulthood. Neural mechanisms underlying these changes may have implications for understanding the neurobiology of schizophrenia, putatively a neurodevelopmental disorder. In this study, we evaluated the effects of MK-801 (dizocilpine), on automated measures of distance traveled and stereotypies in adult rats with neonatal (postnatal day 7) lesions, and tested the effects of haloperidol, clozapine and an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic (AMPA) antagonist LY293558 on the MK-801-induced behaviors. The lesioned rats showed significantly greater increases in motor activity after 0.05 and O.1 mg/kg of MK-801 than did controls. Both haloperidol (0.1 and 0.4 mg/kg) and clozapine (4 and 10 mg/kg) reduced hyperlocomotion elicited by 0.2 mg/kg MK-801 in the ventral hippocampus (VH)-lesioned and sham rats. Haloperidol was more potent than clozapine in decreasing MK-801-induced stereotypy, especially in the lesioned rats. Moreover, an AMPA antagonist normalized exaggerated MK-801-induced hyperolocomotion in the lesioned rats at doses that had no effect in controls. These results demonstrate that the lesioned rats are more sensitive to MK-801 during adulthood than control rats, and that antidopaminergic drugs as well as AMPA antagonists antagonize the MK-801-induced behaviors. The neonatal lesion rat model may be useful to further our understanding of the interactions between dopamine and glutamate and their role in the pathophysiology of schizophrenia. PMID- 11103883 TI - In-vivo assessment of 5-HT2A and 5-HT2C antagonistic properties of newer antipsychotics. AB - The effects of serotonin (5-HT) receptor ligands on the MK 212 (6-chloro-2[1 piperazinyl]pyrazine) discriminative stimulus and quipazine-induced head twitches were studied in rats. 5-HT1A (8-OH-DPAT) and preferential 5-HT2A (DOI) receptor agonists did not generalize to the discriminative stimulus. The 5-HT2B/2C receptor antagonist, SB 206553 (5-methyl-1-(3-pyridylcarbamoyl)-1,2,3,5 tetrahydropyrrolo[2 ,3-f]indole), and the 5-HT2A/2C-receptor antagonist, ritanserin, acted as potent antagonists, whereas the 5-HT2A-receptor antagonist, MDL 100.151 ([(+/-)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4- piperidine-methanol), produced minor and inconsistent inhibition. SB 206553 was a weak antagonist against quipazine-induced head twitches, whereas MDL 100.151 and ritanserin were potent antagonists. This suggests that the MK 212 discriminative stimulus is mediated by 5-HT2C receptors, while quipazine-induced head twitches are mediated primarily by 5-HT2A receptors. The effects on quipazine-induced head twitches were comparable to previously published effects on the DOI discriminative stimulus. 5-HT2A- and 5-HT2C-receptor antagonistic potencies of clozapine, olanzapine, risperidone, sertindole and ziprasidone were compared in the same models. Clozapine showed similar potencies in both models, while sertindole, olanzapine and risperidone inhibited quipazine-induced effects more potently than the MK 212 discriminative stimulus. Ziprasidone exerted a minor preference for 5-HT2A- compared to 5-HT2C-receptor-mediated effects. The ratio between in vivo inhibitory potencies at 5-HT2A and 5-HT2C receptors did not correlate with corresponding ratios from in-vitro affinity and ex-vivo occupancy studies in the literature. PMID- 11103882 TI - Behavioural disturbances associated with hyperdopaminergia in dopamine transporter knockout mice. AB - Mice lacking the dopamine transporter (DAT-/-) are characterized by high extracellular dopamine levels and spontaneous hyperlocomotion. We performed a detailed analysis of the behavioural phenotype of DAT-/- mice in order to identify other behavioural impairments associated with the hyperdopaminergic tone of these mutant mice. In particular, we investigated locomotor activity, exploration, and social and maternal behaviours, which are known to be regulated by dopamine. DAT-/- mice were easily aroused by novelty and always responded with hyperlocomotion, which interfered with habituation to the testing environment, exploratory behaviour in an open field and the coping response to forced swimming stress. Social behaviours such as interaction with an unknown congener or aggressiveness were not modified in DAT-/- mice compared with DAT+/- and DAT+/+ mice, although the maternal behaviour of mutant females was severely disturbed. Haloperidol and clozapine reversed the hyperactivity in DAT-/- mice, with a rightward shift of the dose-response curve compared with control animals, suggesting a dopamine-mediated effect. These results emphasize the role of dopamine regulation in locomotion, exploration and maternal behaviours and suggest that mice with a genetic deletion of DAT may represent a useful model to elucidate the altered behavioural processes accompanying pathological conditions associated with hyperdopaminergic function. PMID- 11103884 TI - Repeated treatment with 8-OH-DPAT induces tolerance to its ability to produce the 5-HT1A behavioural syndrome, but not to its ability to attenuate haloperidol induced catalepsy. AB - When administered acutely, 5-hydroxytryptamine1A (5-HT1A) agonists attenuate the cataleptic side effects of antipsychotics. We investigated whether tolerance occurs to these effects after repeated administration of the 5-HT1A agonist 8 hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT). For comparison, we also assessed the ability of 8-OH-DPAT to produce elements of the 5-HT1A behavioural syndrome (i.e. forepaw treading, flat body posture and lower lip retraction), some of which readily demonstrate tolerance. Catalepsy was measured in rats using both the cross-legged position test and the bar test. Repeated treatment with 8 OH-DPAT (0.63-2.5 mg/kg subcutaneously), once daily for 4 days, did not significantly alter the ability of acute treatment with 8-OH-DPAT (0.01-2.5 mg/kg) to inhibit catalepsy induced by haloperidol (2.5 mg/kg) in either test. In contrast, the ability of 8-OH-DPAT to produce the 5-HT1A behavioural syndrome was significantly attenuated by the repeated treatment. The present data, showing an absence of tolerance to the anti-cataleptic effects of a 5-HT1A agonist, indicate that mixed dopamine antagonist/5-HT1A agonist compounds may continue to have a low propensity to induce extrapyramidal side effects during chronic treatment. PMID- 11103885 TI - Low-dose clozapine pretreatment partially prevents haloperidol-induced deficits in conditioned active avoidance. AB - The effectiveness of neuroleptics in disrupting conditioned active avoidance has led to the widespread use of this test as an index of antipsychotic efficacy, whereas the tendency for these drugs to induce catalepsy is believed to reflect their propensity to cause extrapyramidal motor side-effects. Although the typical neuroleptic haloperidol produces catalepsy as well as profound deficits in conditioned active avoidance, the atypical neuroleptic clozapine does not induce catalepsy and is less effective than haloperidol in disrupting active avoidance. Furthermore, clozapine pretreatment prevents haloperidol-induced catalepsy. We investigated whether clozapine pretreatment might also reduce the disruptive effects of haloperidol on two-way active avoidance. We assessed the avoidance acquisition of the following drug treatment groups in which all animals received two injections prior to testing: vehicle + vehicle, vehicle + haloperidol (0.1 mg/kg, i.p.), clozapine (2.5, 5.0 or 10 mg/kg, i.p.) + haloperidol (0.1 mg/kg, i.p.), or clozapine (2.5, 5.0 or 10 mg/kg, i.p.) + vehicle. Haloperidol pretreated animals showed markedly impaired active avoidance, deficits which were improved by 2.5 and 5 mg/kg but not by 10 mg/kg clozapine pretreatment. These data suggest that the disruptive effects of haloperidol on conditioned active avoidance partially mirror its capacity to induce catalepsy and extrapyramidal motor symptoms. Furthermore, this study indicates that clozapine may be effective in reducing motor side-effects caused by typical neuroleptics. PMID- 11103886 TI - Neuropsychological and conditioned blocking performance in patients with schizophrenia: assessment of the contribution of neuroleptic dose, serum levels and dopamine D2-receptor occupancy. AB - Patients with schizophrenia show impairments of attention and neuropsychological performance, but the extent to which this is attributable to antipsychotic medication remains largely unexplored. We describe here the putative influence of the dose of antipsychotic medication (chlorpromazine equivalents, CPZ), the antipsychotic serum concentration of dopamine (DA) D2-blocking activity and the approximated central dopamine D2-receptor occupancy (DA D2-occupancy), on conditioned blocking (CB) measures of attention and performance on a neuropsychological battery, in 108 patients with schizophrenia (compared with 62 healthy controls). Antipsychotic serum concentration and D2-occupancy were higher in patients with a paranoid versus non-paranoid diagnosis, and in female versus male patients (independent of symptom severity). Controlling for D2-occupancy removed the difference between high CB in paranoid and impaired low CB in non paranoid patients. Similar partial correlations for antipsychotic drug dose and serum levels of DA D2-blocking activity with performance of the trail-making and picture completion tests (negative) and the block-design task (positive) showed the functional importance of DA-related activity. High estimates of central DA D2 occupancy were related to impaired verbal fluency but were associated with improved recall of stories, especially in paranoid patients. This, the first study of its kind, tentatively imputes a role for DA D2-related activity in left frontal (e.g. CB, verbal fluency) and temporal lobe functions (verbal recall) as well as in some non-verbal abilities mediated more in the right hemisphere in patients with schizophrenia. PMID- 11103887 TI - A comparison of the behavioural effects of 5-HT2A and 5-HT2C receptor agonists in the pigeon. AB - Activity at the 5-HT2A receptor versus that of the 5-HT2C receptor was studied in three behavioural paradigms. In pigeons trained to discriminate 0.32 mg/kg of 1 (2,5-diemethoxy-4-iodophenyl)-2-aminopropane (DOI) (a mixed 5-HT2A/C receptor agonist) from vehicle, quipazine (0.1-1 mg/kg) and m-chlorophenylpiperazine (mCPP) (1-3 mg/kg) substituted for DOI in a dose-related manner, and this generalization was blocked by MDL100907 (0.0001-0.01 mg/kg), a selective 5-HT2A receptor antagonist. RO60-0175 (a relatively selective 5-HT2C agonist) induced partial substitution at 3 mg/kg that was antagonized by both MDL100907 and by 3 mg/kg of SB242084, a relatively selective 5-HT2C antagonist. MK212 (a mixed 5 HT2C/A agonist) induced partial substitution that was antagonized by SB242084, but not by MDL100907. On a progressive ratio 5 operant schedule (PR5) for food reinforcement, DOI, quipazine, mCPP, MK212 and R060-0175 decreased the break point; mCPP, DOI, MK212 and quipazine also induced vomiting. Although MDL100907 antagonized both the reductions of break point and vomiting, SB242084 only partially attenuated the decrease in break point observed with MK212 and DOI, and was unable to eliminate vomiting. Thus pharmacological activity at the 5-HT2A receptor can be behaviourally distinguished from pharmacological activity at the 5-HT2C receptor in the pigeon. Furthermore, the decrease in the break point of a PR5 schedule induced by 5-HT2C receptor agonists may be related to decreased appetite, whereas that induced by 5-HT2A receptor agonists may be due to unrelated factors, such as emesis. PMID- 11103888 TI - Conditioned locomotion in rats following amphetamine infusion into the nucleus accumbens: blockade by coincident inhibition of protein kinase A. AB - Recent studies demonstrate a role for cyclic adenosine monophosphate (cAMP) dependent protein kinase (PKA) in the nucleus accumbens (NAc) in reward-related learning. To clarify this role, we assessed the effect of PKA inhibition on the unconditioned and conditioned locomotor activating properties of intra-NAc amphetamine. Rats underwent three 60 min conditioning sessions, pairing a test environment with bilateral co-infusions of amphetamine (25 microg/side) and the PKA inhibitor Rp-adenosine 3',5'-cyclic monophosphothioate triethylamine (Rp cAMPS) (0, 2.5, 250, 500 ng, 1, 10 or 20 microg/side). Two additional groups - receiving amphetamine explicitly unpaired with the environment or saline/environment pairings - served as controls. In a subsequent drug-free 60 min session, animals that received amphetamine/environment pairings demonstrated conditioned locomotion relative to controls. Rp-cAMPS co-treatment during pairing sessions differentially affected conditioned and unconditioned locomotor activation. Amphetamine-induced unconditioned activity was significantly enhanced by 500 ng and 1 microg Rp-cAMPS, locomotor sensitization was enhanced by 250 ng-1 microg Rp-cAMPS, and conditioned activity was attenuated by 1 microg Rp-cAMPS and blocked by 10 and 20 microg Rp-cAMPS. Thus, unconditioned activity and locomotor sensitization were enhanced at doses (250 ng-1 microg) that did not affect or attenuated conditioned activity, while conditioned activity was reduced or blocked at doses (1-20 microg) that enhanced or did not affect overall unconditioned activity. These results demonstrate that the activation of PKA plays a critical role in the process by which properties of drugs become associated with environmental stimuli. PMID- 11103889 TI - Effects of the cannabinoid ligand SR 141716A alone or in combination with delta9 tetrahydrocannabinol or scopolamine on learning in squirrel monkeys. AB - To investigate the effects of the cannabinoids on learning and on scopolamine induced disruptions in learning, delta9-tetrahydrocannabinol (delta9-THC), SR 141716A (an antagonist at CB1 receptors) and scopolamine were administered to squirrel monkeys responding in a repeated-acquisition task. In this task, monkeys acquired a different three-response sequence each session and responding was maintained by food presentation under a second-order fixed-ratio 5 schedule. When either delta9-THC (0.1-0.56 mg/kg, i.m.) or SR 141716A (1-10 mg/kg, i.m.) was administered alone, 60 and 75 min before the session, respectively, both cannabinoid ligands dose-dependently decreased the overall rate of responding and increased the overall percentage of errors. However, at a dose that had little or no effect alone (i.e. 1 mg/kg), SR 141716A antagonized the disruptive effects of delta9-THC (0.18-1.8 mg/kg) on acquisition, shifting the dose-effect curves for rate of responding and percentage of errors at least 1/2 log unit to the right. Finally, when either delta9-THC (0.001-1 mg/kg) or SR 141716A (0.32-10 mg/kg) was administered with scopolamine (0.01 or 0.032 mg/kg, 15 min before the session), greater rate-decreasing and error-increasing effects were obtained than with scopolamine alone. These results suggest that while low doses of SR 141716A can antagonize the effects of delta9-THC in squirrel monkeys, high doses can also disrupt acquisition when administered alone and potentiate the disruptive effects of scopolamine on acquisition. PMID- 11103890 TI - Effects of cocaine-induced sensitization on ethanol drinking: sex and strain differences. AB - Sensitization induced by repeated drug exposure has been proposed to increase 'wanting' the drug and to facilitate the transition from moderate to excessive drug intake. The present study examined the effects of cocaine-induced sensitization on ethanol-drinking behavior in male and female rats from different strains. In experiment 1, rats were pretreated with six injections of saline or cocaine (10 mg/kg, i.p.), spaced by 3-day intervals, and were subsequently allowed access to ethanol intake in an unrestricted free-choice procedure. In experiment 2, rats had acquired ethanol-drinking behavior and were exposed to the sensitizing treatment described previously or were left undisturbed. Subsequently, all animals again had access to ethanol. Whatever the sex and strain concerned, sensitized and control animals did not differ in either the acquisition or the maintenance of ethanol-drinking behavior, suggesting that cocaine-induced behavioral sensitization does not modify ethanol intake. The present results also confirm the sex- and strain-dependent character of alcohol intake and of the 'alcohol deprivation effect'. PMID- 11103891 TI - Conditioned compensatory response to ethanol as indicated by locomotor activity in rats. AB - Spontaneous motor activity (SMA) was used to investigate conditioned tolerance to the depressant effect of ethanol, and conditioned responses to stimuli predicting ethanol injection. Rats were injected with saline or ethanol at 800 or 1600 mg/kg, on alternate days, in two distinctly different locations, over a period of 20 conditioning days. The two ethanol doses were administered to separate groups of rats, but conditioned effects were determined by within-subject comparisons. Conditioned SMA responses (rearing and ambulatory activity) were measured after injection of saline in the location previously paired with ethanol treatment, and conditioned tolerance was determined by observing ethanol effects in rats tested in the environment previously paired with saline treatment. Ethanol-paired stimuli increased SMA (both activity measures, both dose-groups) during the conditioned response test. Absence of these conditioned stimuli during the tolerance test resulted in greater behavioral depression with the 800 mg/kg ethanol dose for both the rearing and ambulation measures; however, this effect was seen with the 1600 mg/kg dose for the rearing measure only. These results provide further evidence that Pavlovian conditioning is involved in tolerance to the depressant action of ethanol on overt behavior, and demonstrate the presence of such conditioned compensatory responses in the absence of ethanol treatment. PMID- 11103892 TI - Housing conditions and the anxiolytic efficacy of buspirone: the relationship between main and side effects. AB - Serotonergic anxiolytics yield contradictory results both in the laboratory and clinically. In an attempt to investigate the cause of discrepancies, the anxiolytic effect of buspirone (0, 3 or 10 mg/kg, single treatment) was tested 1 h and 4 h after injection in rats in different housing conditions. At 1 h after drug administration, buspirone increased corticosterone production and decreased locomotor behaviour in both the elevated plus-maze and the social interaction tests. No anxiolytic-like effect was produced in either test. At 4 h after drug injection, no corticosterone or locomotor effects of buspirone were observed. In contrast, anxiolytic effects emerged in this phase. Open arm exploration and social investigation were increased in the plus-maze and social interaction test, respectively. In the plus-maze, the anxiolytic effect was significant in isolated animals only. In the social interaction test, the anxiolytic effect was stronger in isolated than in group-housed animals. When corticosterone secretion was inhibited by adrenalectomy, a full anxiolytic effect of buspirone was observed 1 h after drug administration. It appears that the side effects of buspirone have a shorter duration than the main anxiolytic effect. The buspirone-induced increase in corticosterone may have abolished the anxiolytic effects of the drug shortly after injection. Individual housing enhanced the anxiolytic efficacy of buspirone 4 h after administration. PMID- 11103893 TI - Concurrent variable-interval drug self-administration and the generalized matching law: a drug-class comparison. AB - It has previously been shown that self-administration of cocaine under concurrent variable-interval schedules is well described by the generalized matching law. That is, choice between two cocaine-maintained options was apportioned in accordance with relative frequency of reinforcement. However, the generality of this conclusion to drugs of other pharmacological classes has not been determined. In the present study, four male rhesus monkeys (Macaca mulatta) lever pressed under various pairs of concurrent variable-interval schedules with drug injection as the maintaining event. An opioid (alfentanil, 0.001 or 0.004 mg/kg/injection), a barbiturate (methohexital, 0.25 or 0.5 mg/kg/injection), and a psychomotor stimulant (cocaine, 0.05 mg/kg/injection) were selected as representatives of major classes of abused drugs and because of their relatively short duration of action. As has been found for cocaine, choice was well accounted for by the generalized matching law. There were no systematic differences in matching-law parameters across drugs and/or doses. As in earlier studies with drug and nondrug reinforcers, undermatching was a consistent finding. Therefore, the conclusion that relative reinforcement frequency is a crucial determinant of choice, as proposed by the generalized matching law, can be extended to behavior maintained by drugs from a variety of pharmacological classes. PMID- 11103895 TI - The development and expression of locomotor sensitization to nicotine in the presence of ibogaine. AB - Ibogaine is a naturally occurring psychoactive alkaloid with claimed efficacy in the treatment of certain drug addictions, including nicotine. It has been reported to be a non-competitive blocker of nicotinic receptors, with a potent inhibitory action on nicotinic acetylcholine receptor-mediated catecholamine release. We have investigated the effect of different doses of ibogaine on the development and expression of sensitization to the locomotor stimulant effect of nicotine in rats, a facilitatory process in which a history of exposure to nicotine results in enhanced locomotor activity when the same dose of nicotine is administered repeatedly. The effects were determined of co-administering ibogaine (0.0, 5.0 or 10 mg/kg i.p.) with nicotine (0.0 or 0.4 mg/kg s.c.) daily for 21 days. Dose-response curves for nicotine (0.04-0.8 mg/kg s.c.) were then determined in groups of 10 rats. There was clear sensitization of the locomotor activity produced by nicotine in photocell activity cages but co-administration of ibogaine with nicotine had no effect on the degree of sensitization. Ibogaine (5-20 mg/kg) itself did not influence locomotor activity and was also without effect on the expression of the sensitized response to 0.4 mg/kg of nicotine (n = 10). Thus, there was no evidence that ibogaine may retard or suppress sensitization to nicotine. PMID- 11103894 TI - Effects of nitric oxide synthase inhibitors on timing and short-term memory in rats. AB - Nitric oxide synthase (NOS) inhibitors have been shown to affect the development of long-term potentiation and the acquisition of new learning. In the present study, we investigated the effects of NOS inhibitors in two animal models in which aspects of cognition are measured in well-learned operant tasks - a delayed non-match-to-position (DNMTP) task and a multiple signalled-unsignalled differential reinforcement of low rates (DRL) 15 s schedule - models of short term memory and behavioral inhibition/timing, respectively. Since an overlap in the behavioral effects of NOS inhibitors and phencyclidine (PCP)-like N-methyl-D aspartate (NMDA) antagonists has been observed previously, we compared our results with NOS inhibitors to those obtained with PCP. Whereas PCP produced a delay-independent decrease in the DNMTP task and increased burst responding (consecutive responses with inter-response intervals of < 3 s) in both the signalled and unsignalled components of the DRL procedure, 7-nitroindazole did not affect accuracy in the DNMTP task nor did it alter the pattern of responding in either component of the DRL schedule. Similarly, NG-nitro-L-arginine (L-NOARG) and NG-nitro-L-arginine-methyl-ester (L-NAME) did not affect accuracy in the DNMTP task. These results suggest that NOS inhibitors do not produce PCP-like disruption of behavioral inhibition or timing, nor do they decrease accuracy in a conditional discrimination task, as has been observed with PCP. The present results lend further support to the hypothesis that nitric oxide modulation does not affect retention of well-learned tasks, although it may affect acquisition of novel behavior. PMID- 11103896 TI - Electromigration methods for amino acids, biogenic amines and aromatic amines. AB - Methods of electromigration in laboratory apparatus of small-bore size have recently undergone development at a remarkably rapid pace, leading to a variety of new analytical techniques. One such technique is called "capillary electrophoresis" (CE), which is further classified on the basis of electromigration mode, viz., "capillary zone electrophoresis" (CZE), which, in turn, has several variations. This review aims to give a short overview of the various electromigration methods for amino compounds by using CE. Firstly, this review briefly summarizes the detection methods employed for detection of monoamines and polyamines by CE for both native and derivative forms. Next, current CE methods are described, and their applications to detection of amino acids, biogenic amines, aromatic amines, including heteroaromatic amines and their enantiomers, are introduced from representative papers. Finally, new methods for single-cell analysis and microchip CE techniques are focused on. PMID- 11103897 TI - Analysis of biogenic amines in microdialysates of the brain. AB - Microdialysis is a method of sampling a liquid compartment by means of a hollow fibre dialysis membrane. The method was developed in the 1980s as a technique for sampling the extracellular fluid of the brain of conscious animals. When used in combination with sensitive analytical chemical tools, microdialysis can be used to study the regulation of neurotransmission in the living brain. Here we describe the application of microdialysis for sampling and detection of biogenic amines (dopamine, noradrenaline and serotonin) in brain tissue. A short overview of the microdialysis technique and its applications are given. In addition, the analytical chemical methods that are currently used to assay biogenic amines in dialysates are briefly discussed. PMID- 11103898 TI - Clinical chemistry of serotonin and metabolites. AB - Analyses of serotonin and other 5-hydroxyindoles, such as its precursor 5 hydroxytryptophan and major metabolite 5-hydroxyindoleacetic acid (5-HIAA), are indispensable for the elucidation of their (patho)physiological roles. In clinical chemistry attention is mainly focused on the diagnosis and follow-up of carcinoid tumours. For this most laboratories routinely measure urinary 5-HIAA. More recently, measurements of serotonin in platelets and urine have been advocated. Platelet serotonin may be the most sensitive indole marker for the detection of carcinoid tumours that secrete only small amounts of serotonin and/or its precursor 5-hydroxytryptophan. Although several chromatographic techniques have emerged for the analysis of tryptophan-related indoles, HPLC with either electrochemical or fluorometric detection have become the methods of choice for their quantification. HPLC-based methods combine selectivity, sensitivity and high precision, and enable the simultaneous investigation of several metabolically related indoles. This review aims to place the analysis of indoles in biological matrices in a biochemical, physiological and clinical perspective and highlights several important steps in their chromatographic analysis and quantification. PMID- 11103899 TI - Analysis of amines in plant materials. AB - Biogenic amines are conveniently divided into aliphatic monoamines, aliphatic di- and polyamines and aromatic amines. These compounds are shown to fulfill an array of roles in cellular metabolism. Thus, amines are needed for growth and development and their metabolism appears to be coordinated with the cell cycle. Di- and polyamines, among which are putrescine, spermidine and spermine, are ubiquitous polycationic molecules that occur in all living cells. However, plants accumulate a number of specific related compounds under free or conjugated forms. In plant tissues, the molecular diversity combined with the fact that amine contents are highly responsive to developmental and environmental signals encouraged analysts to develop specific procedures for their isolation and characterization. The main goals were to develop high performance routine procedures in terms of selectivity, repeatability and detectability with minimum running costs. Domains of application concern not only fundamental aspects of amine biochemistry and physiology in plants but also increasing needs in the control of food and beverage quality from plant origin. The present review reports the most recent advances in extraction, identification and quantitation of amines in plant tissues with special interest in the analysis of original and uncommon metabolites. Emphasis is directed towards chromatographic and electrophoretic separation methodologies and new detection technologies of both derivatized and underivatized compounds including photometry, fluorometry, amperometry and mass spectrometry. PMID- 11103900 TI - Current methodologies for the analysis of aminoglycosides. AB - The aminoglycosides are a large and diverse class of antibiotics that characteristically contain two or more aminosugars linked by glycosidic bonds to an aminocyclitol component. Structures are presented for over 30 of the most important members of this family of compounds. The use of aminoglycosides in clinical and veterinary medicine and in agriculture is described. Qualitative methods for aminoglycoside analysis include X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS). The major part of this article comprises a comprehensive review of quantitative methods for the determination of aminoglycosides. These are microbiological assay, radiochemical assay, radioimmunoassay, enzyme immunoassay, fluoroimmunoassay and other immunoassays, spectrophotometric and other non-separative methods, gas chromatography (GC), thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), and capillary electrophoresis (CE). Simple spectrophotometric methods may be adequate for the assay of bulk pharmaceuticals and their formulations. Microbiological assays make useful semi-quantitative screening tests for the analysis of veterinary drug residues in food, but rapid enzyme immunoassays are more suitable for accurate measurements of aminoglycosides in complex matrices. Automated immunoassays are the most appropriate methods for serum aminoglycoside determinations during therapeutic drug monitoring. HPLC techniques provide the specificity and sensitivity required for pharmacokinetic and other research studies, while HPLC-MS is employed for the confirmation of veterinary drug residues. The potential for further development of chromatographic and CE methods for the analysis of biological samples is outlined. PMID- 11103901 TI - Separation and assay methods for melatonin and its precursors. AB - Melatonin is an indoleamine hormone that is synthesized from tryptophan via 5 hydroxytryptophan, serotonin and N-acetylserotonin in the vertebrate pineal gland. Many chromatographic and non-chromatographic techniques have been developed and improved for the determination and measurement of melatonin and its related indoleamines. At present, gas chromatography with mass spectrometry and reversed-phase high-performance liquid chromatography with fluorescence or electrochemical detection are widely used for indoleamine determinations in the pineal gland. This review will deal with methods for the separation and determination of the melatonin and its related indoleamines. PMID- 11103902 TI - Separation methods used in the determination of choline and acetylcholine. AB - Cholinergic neurotransmission has been the subject of intensive investigations in recent years due to increasing recognition of the importance of its roles in physiology, pathology and pharmacology. The fact that the disposition of a neurotransmitter may reflect its functional status has made the measurement of acetylcholine and/or its precursors and metabolites in biological fluids an integral part of cholinergic research. With evolving complexity in experimental approaches and designs, and correspondingly increasing demand on sensitivity, specificity and accuracy matching advancements in sophistication in analytical methods have been made. The present review attempts to survey the array of analytical techniques that have been adopted for the measurement of acetylcholine or its main precursor/metabolite choline ranging from simple bioassays, radioenzymatic assays, gas chromatography (GC) with flame ionization detection, GC with mass spectrometry (GC-MS) detection, high-performance liquid chromatography (HPLC) with electrochemical detection (ED), HPLC with MS (HPLC-MS) to the sophisticated combination of micro-immobilized enzymatic reactor, microbore HPLC and modified electrode technology for the detection of ultra-low levels with particular emphasis on the state of the art techniques. PMID- 11103903 TI - Chromatography of guanidino compounds. AB - Guanidino compounds involved in the urea and guanidine cycles have been found in serum of nephritic patients, and some guanidino compounds have been suspected to be uremic toxins. The simultaneous analysis of naturally occurring metabolites is important for diagnosis of diseases. In this review, liquid chromatographic analysis of natural metabolites of guanidino compounds are described. the information about arginine as a precursor of nitric oxide are included. The reports of pharmaceutical compounds having a guanidino group, peptides containing arginine and aminoglycosides are summarized in Table 1. PMID- 11103905 TI - Separation of heteroaromatic amines in food products. AB - In recent years, many studies have dealt with the role of certain heteroaromatic amines (HAs) as mutagenic compounds, and their occurrence in foodstuffs. Here we examine the determination of HAs, focusing on the analytical strategies for their extraction and preconcentration from several matrices. We summarise the properties of heteroaromatic amines and the main drawbacks involved in their analysis, and then concentrate on the separation procedures, sorbents and solvents used in the sample treatment. We discuss the requirements of the analytical techniques and the strategies most frequently followed to achieve accurate results. PMID- 11103904 TI - Chromatographic and related techniques for the determination of aromatic heterocyclic amines in foods. AB - Some 20 years ago, Japanese scientists discovered a new group of highly toxic compounds, classified as heterocyclic aromatic amines, from broiled and grilled meat and fish products. Numerous studies have shown that most HAs are mutagenic and carcinogenic, and the safety of HA-containing foods has become a concern for the public. To date, more than 20 different mutagenic and/or carcinogenic heterocyclic amines have been identified in foods. This paper reviews the analysis of foods for HAs with 145 references. We survey some of the numerous methods available for the chromatographic analysis of heterocyclic amines and highlight the recent advances. We discuss chromatographic and related techniques, including capillary electrophoresis, and their coupling to mass spectrometry for the determination of these contaminants in foods. In addition, the review summarises data on the content of HAs in various cooked foods. PMID- 11103906 TI - Chromatographic separation of 2,3-benzodiazepines. AB - A review of chromatographic methods for the determination of 2,3-benzodiazepines (2,3-BZs) is presented. The determinations are performed to investigate the presence of potential impurities in drug substances and to study their pharmacokinetic profile in biological samples, either in animals or in humans. Several methods dealt with a pretreatment of samples, i.e., liquid-liquid extraction by using a variety of solvents, solid-phase extraction, direct injection of specimens into the chromatographic apparatus. Different chromatographic techniques have been used. High-performance liquid chromatography allows optimal sensitivity and specificity by using ultraviolet or diode array detection methods. Gas chromatography-mass spectrometry and gas chromatography with nitrogen-phosphorous or electron-capture detectors have been also reported. Suitable methods for the separation of enantiomers of 2,3-BZs have been described. Thin-layer chromatography has been shown to be capable to isolate analytes from biological samples as urine or faeces. The reported chromatographic techniques are currently applied to define the metabolic pathways of 2,3-BZs in experimental and clinical studies. PMID- 11103907 TI - Determination of uremic toxins in biofluids: creatinine, creatine, uric acid and xanthines. AB - Rapid and accurate determination of small molecule metabolic end-products is vital for clinical diagnosis and study of many metabolic disorders and medical abnormalities. Chromatographic and electrophoretic techniques are attractive for clinical analyses because of the inherent ability to analyze multiple component biofluids and determine the analytes of interest with minimal interference from other species. This manuscript reviews recent (1990-present) developments in chromatography and electrophoresis methodology for the determination of creatinine, creatine, uric acid and xanthines in biofluids. PMID- 11103908 TI - Chromatographic and capillary electrophoretic methods for the analysis of nicotinic acid and its metabolites. AB - Methods for the assay of nicotinic acid (NiAc) and its metabolites in biological fluids using high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE) are reviewed. Most of the references cited in this review concern HPLC methods. A few CE methods that have been recently reported are also included. As these compounds are relatively polar and have a wide range of physico-chemical properties, the sample pre-treatment or clean-up process prior to analysis is included. Most HPLC methods using an isocratic elution system allow determination of a single or few metabolites, but gradient HPLC methods enable simultaneous determination of five to eight compounds. Simultaneous determination of NiAc including many metabolites in a single run can be achieved by CE. We also discuss the pharmacokinetics of NiAc and some of its metabolites. PMID- 11103909 TI - Separation methods for amino group-possessing pesticides in biological samples. AB - The separation methods for pesticides include liquid-liquid extraction, solid phase extraction and solid-phase microextraction, gas chromatography (GC), GC mass spectrometry (MS), GC-MS-MS, high-performance liquid chromatography (LC), LC MS and LC-MS-MS. This review deals with each technique commonly used for extraction, chromatographic separation and detection of amino group possessing pesticides, such as diazines, triazines, carbamates, dinitroanilines and chloroacetanilides in biological samples. The methods presented for analysis of the pesticides in complicated biological matrices seem to be easily applicable to surface or groundwater in environmental chemistry. PMID- 11103911 TI - The effects of amphetamine and raclopride on food transport: possible relation to defensive behavior in rats. AB - Recent work has shown that transport of food items from open, exposed food sources to a covered shelter is reduced by drugs thought to have anxiolytic properties in rodents and humans. We studied the effects of amphetamine and the dopamine D2/3-receptor antagonist, raclopride, in this test of food transport that pits immediate food consumption against exposure in an open space. Rats traveled from a home cage along an elevated beam to obtain single food items of varying sizes located at one of 12 distances from the home cage. Large food items and items located close to the home cage were carried back and consumed inside the cage. Small items and items located farther from the cage were eaten immediately at the food source while sitting on the beam. Amphetamine sulfate (0.001-2.0 mg/kg, i.p.) decreased eating on the beam and increased carrying of food items to the home cage. Raclopride (0.005-0.2 mg/kg, i.p.) tended to reduce carrying of food to the home cage, but 0.05 mg/kg raclopride did not block the increase in food carrying seen with amphetamine treatment (2 mg/kg). The increased food carrying seen with amphetamine is opposite to the effect produced by anxiolytic drugs, raising the possibility that amphetamine promotes carrying by increasing defense or 'anxiety'. Consistent with this hypothesis, amphetamine (2 mg/kg; the maximally effective dose in the food-carrying experiment) decreased open-arm exploration in the elevated plus-maze, considered to be an anxiogenic effect. These results indicate that stimulation of monoaminergic neurotransmission increases food transport from exposed food sources to a shelter; D2/3-receptor blockade tends to reduce it. The food-carrying test provides a rich, ethologically valid paradigm to assess the effects of psychoactive drugs on species-specific, defensive behaviors in rodents. PMID- 11103910 TI - Prenatally protein-malnourished rats are less sensitive to the amnestic effects of medial septal infusions of chlordiazepoxide. AB - Evidence is mounting that prenatal protein malnutrition affects the physiological properties of the GABAergic neurotransmitter system in rats. To investigate the functional behavioral consequences of these changes, chlordiazepoxide (CDP, a positive modulator of the GABA(A) receptor) was applied directly to the medial septum and the amnestic response appraised. In adulthood, male offspring of rats provided with a protein-deficient diet (6% casein) for 5 weeks prior to mating and throughout pregnancy underwent stereotaxic surgery to implant steel cannulae aimed at the medial septum. After recovery, spatial learning performance in the submerged platform version of the Morris water maze task was assessed immediately following a 1 microl infusion of either artificial cerebrospinal fluid (aCSF), or one of three doses of CDP (15, 30 and 60 nmol). Well-nourished control rats demonstrated a robust amnestic response to intraseptal CDP. During task acquisition, well-nourished rats administered each of the doses exhibited significantly longer escape latencies than those given aCSF. On the probe trial (platform removed) a lower proportion of time was spent in the target quadrant (all three doses) at a greater average distance from the former platform location (30 and 60 nmol doses). In contrast, prenatally malnourished rats exhibited a muted sensitivity to CDP, most notable at the 30 nmol dose. These findings provide further support for functional changes within the GABAergic system consequent to malnutrition. PMID- 11103912 TI - Does locomotor response to novelty in rats predict susceptibility to develop sensitization to cocaine and PHNO? AB - It has been suggested that the locomotor response of rats to novelty is positively correlated with motor stimulant effects of acute injections with psychomotor stimulants, and liability to self-administer these drugs. In addition, response to novelty appears to be inversely correlated with an individual's susceptibility to develop behavioural sensitization (an increase in the behavioural response to a given dose of stimulant after repeated treatments). To test some of these putative relationships, 96 rats were allocated to one of two subgroups based on a median split of locomotor responses to novelty. Animals then received 10 successive injections of either vehicle, cocaine (10 mg/kg), or the direct D2 agonist, (+)-4-propyl-9-hydroxynaphthoxazine (PHNO: 15 microg/kg), and locomotor activity was monitored. Conditioning tests and additional sensitization and cross-sensitization tests were conducted. Results showed that locomotor responses to novelty are not significantly correlated with locomotor effects of either acute injection with cocaine or PHNO, or rate of development of behavioural sensitization to these drugs. However, locomotor responses to novelty did predict level of locomotor and stereotypy responses to cocaine, and to a lessor extent to PHNO. Cocaine-treated, but not PHNO-treated, rats exhibited drug conditioned-like effects. Cross-sensitization between cocaine and PHNO was not observed, indicating independent mechanisms for sensitization. It is concluded that the locomotor response to novelty can predict level of locomotion and stereotypy produced by cocaine and PHNO, but does not predict the degree or rate of behavioural sensitization to either of these drugs. PMID- 11103913 TI - Previous experience of diazepam withdrawal prevents the formation of a withdrawal conditioned taste aversion: test of a blocking hypothesis. AB - Prior experience of withdrawal from chronic diazepam treatment reduces the aversiveness of withdrawal when precipitated withdrawal is made the unconditioned stimulus in a conditioned taste aversion (CTA) paradigm. Accounts of the mechanism by which unconditioned stimulus pre-exposure reduces its effectiveness in CTA postulate that unconditioned stimulus pre-exposure leads to the formation of associations with the environment, resulting in blocking of taste conditioning. We tested whether a blocking explanation accounted for the reduced effectiveness of withdrawal as a unconditioned stimulus in a CTA following prior exposure. Mice received chronic diazepam (15mg/kg/day, s.c. in sesame oil), or sesame oil vehicle, for three periods of 7 days, interspersed with 3-day withdrawal periods. The first two withdrawals occurred either in the home cage, or in one compartment of a place-conditioning apparatus (PCA). Animals which experienced withdrawal in the home cage were given equivalent experience of the PCA outside the withdrawal period. The third withdrawal was precipitated by i.p. administration of flumazenil (20mg/kg). Thirty minutes before injection, all animals were placed individually in the compartment of the PCA to which they had been previously exposed, allowed to drink a novel 10% sucrose solution, injected with flumazenil, and replaced in the PCA for 2 h, before being returned to the home cage. When sucrose consumption was measured 24 h later, only that group which had experienced all three withdrawals in the PCA showed evidence of a CTA. These animals (but not those that had experienced withdrawal in the home cage, or vehicle-treated mice) also showed strong avoidance of the chamber in which they had experienced withdrawal. Thus, no evidence was adduced that prior conditioning of an environment-conditioned stimulus to a withdrawal unconditioned stimulus blocked the formation of a CTA. When the CTA conditioning was repeated in the home cage, again only the mice that had experienced withdrawal in the place conditioning apparatus showed evidence of conditioning. These observations are discussed in the context of blocking explanations of unconditioned stimulus pre exposure. PMID- 11103914 TI - Discriminative stimulus effects of putative D3 dopamine receptor agonists in rats. AB - Three separate groups of rats were trained to discriminate the putative D3 dopamine receptor agonists (+/-)-7-hydroxydipropylaminotetralin (7-OH-DPAT) (0.03 mg/kg), PD 128,907 (1.0 mg/kg) and quinpirole (0.03 mg/kg) from saline. Food was presented after each 10 (7-OH-DPAT and PD 128,907) or 20 (quinpirole) consecutive responses on one lever after administration of the training drug, and the other lever after the administration of saline. Once stable performances were obtained, the effects of various doses of several dopaminergic agonists were assessed during test sessions in which responses on either lever were reinforced. The substitution tests were conducted to determine if differences in potencies would be obtained, which would be suggestive of differences in the mechanisms underlying the discriminative effects of the training drugs. Non-selective agonists with activity at both D2 and D3 dopamine receptors (D2-like agonists) substituted for each of the three training drugs. In addition, the selective D2 dopamine receptor agonist U91356A also generalized to both 7-OH-DPAT and PD 128,907. The potencies of the D2-like agonists in substituting for each training drug were highly correlated with potencies in substituting for the others. SKF 82958 and SKF 81297, agonists with selectivity for D1 and D5 dopamine receptors (D1-like agonists), partially substituted for 7-OH-DPAT but not PD 128,907. The D1-like partial agonist SKF 38393 did not substitute for any of the training drugs for which it was tested. Cocaine produced intermediate substitution in 7-OH DPAT- and PD 128,907-trained subjects and did not substitute at all in quinpirole trained subjects. The dopamine D1-like antagonist SCH 39166 (0.001-0.03 mg/kg) did not alter the discriminative stimulus effects of PD 128,907, whereas the D2 like dopamine antagonist spiperone (0.001-0.1 mg/kg) produced at the highest dose an insurmountable antagonism of the discriminative effects of PD 128,907. In contrast, there was no appreciable antagonism of the effects of PD 128,907 on response rates. The data collected are consistent with a distinction between the effects of each of these training drugs and the indirectly acting agonist cocaine. Further, these data indicate that there are differences in the mechanisms underlying the discriminative effects of PD 128,907 and its effects on response rates. Moreover, these data indicate that each of the training drugs is distinct from drugs acting through D1 dopaminergic mechanisms. However, there were no data that clearly distinguished these training drugs from each other or from drugs acting through D2 dopaminergic mechanisms. PMID- 11103915 TI - Contextual fear conditioning and baseline startle responses in the rat fear potentiated startle test: a comparison of benzodiazepine/gamma-aminobutyric acid A receptor agonists. AB - In the rat, fear-potentiated startle (FPS) test animals are first trained to associate brief light presentations with a mild electric footshock and then tested for startle responses to acoustic stimuli, delivered either in darkness (i.e. baseline startle) or after the conditioning stimulus. Following light presentation the magnitude of the startle response is markedly increased, and the test is commonly used to distinguish anxiolytic drug effects (i.e. a reduction in FPS) from non-specific effects such as sedation/muscle relaxation. However, recent studies suggest that the environment in which the animal is trained may also contribute towards the acquisition of a conditioned fear response (i.e. contextual fear conditioning) and that this may elevate startle responses recorded in the dark. In the present study, therefore, we have compared the benzodiazepine/gamma-aminobutyric acid-A receptor agonist chlordiazepoxide with the partial agonists FG 8205 and bretazenil, which are known to have a reduced propensity to produce sedation/myorelaxation, using two different FPS procedures: (i) conditioning and testing in stabilimeter chambers, and (ii) conditioning and testing in different environments. The results show that FPS can be demonstrated in both procedures and that treatment with chlordiazepoxide, FG 8205 or bretazenil dose-dependently attenuates the response. However, animals conditioned and tested in stabilimeter chambers also showed a significant increase in dark startle amplitudes compared with non-shocked rats, suggesting that this response was elevated by contextual fear conditioning. Furthermore, despite clear differences in side-effect liabilities, FG 8205 and bretazenil significantly reduced dark-startle responses, suggesting that this measure is also sensitive to the anxiolytic effects of benzodiazepines. In contrast, when animals were conditioned and tested in different environments, dark-startle responses were not significantly different from those recorded in non-shocked rats and treatment with FG 8205 or bretazenil had no effect. Thus, conditioning and testing animals in different environments may provide a more effective means of distinguishing anxiolytic from non-specific drug effects in the rat FPS test. PMID- 11103916 TI - Nitric oxide synthetase inhibition hinders facilitation of active avoidance learning by nicotine in rats. AB - Nicotine produces dose-dependent enhancement of performance in an active avoidance test, and also increases the levels of NO2- and NO3-, which are stable metabolites of nitric oxide (NO), in various brain regions of rats. On the basis of these two observations, we hypothesized that the beneficial effect of nicotine on learning could result from increased NO in relevant brain regions. We therefore tested active avoidance performance in rats given L-Nomega nitroarginine (L-NA) to inhibit NO synthetase (NOS) prior to nicotine administration. Male Sprague-Dawley rats received L-NA (30 or 50 mg/kg), nicotine (0.4 mg/kg), saline or combinations of these treatments before learning trials. Rats were also tested on the inclined plane, to assess the possible effects due to impairment of motor function by drug treatments on active avoidance learning. L-NA treatment impaired the acquisition of active avoidance learning, and this defect was partially overcome by the co-administration of nicotine. Nicotine facilitated learning and significantly increased the number of correct responses. The threshold for the effect of NOS inhibition on performance exceeded 30 mg/kg L NA, whereas 50 mg/kg impaired learning and also eliminated the nicotine-induced enhancement of learning. On the fifth day of learning trials, no facilitation of learning by nicotine was observed in rats receiving either dose of L-NA. Our results suggest that NO is involved in the facilitation of active avoidance learning by nicotine. PMID- 11103917 TI - Discrimination of intranasal cocaine. AB - In the development of medications for the treatment of cocaine abuse, the drug discrimination paradigm can be used to identify medications that can attenuate the discriminative stimulus effects of cocaine. To ascertain that participants are basing the discrimination on the drug's central effects, this paradigm requires that the drug and placebo administrations do not produce any peripheral effects on which the discrimination can be based. This study examined whether intranasal cocaine (50 mg) can be discriminated from placebo (46 mg lactose + 4 mg cocaine), how quickly this discrimination can be made, and whether pretreatment with intranasal benzocaine can affect this discrimination. Results showed that subjects were generally able to discriminate the drug conditions correctly 15 s after administration, and this was unaffected by benzocaine. These results suggest that subjects base the discrimination on peripheral drug effects (e.g. taste) that are not affected by anaesthesia of the nasal passage, and that the intranasal route of cocaine administration is unlikely to be feasible with a drug discrimination paradigm. PMID- 11103918 TI - Towards multidisciplinary research into health inequalities. PMID- 11103919 TI - Working together for equity. PMID- 11103920 TI - Health inequalities: bringing the hidden assumptions into the open. PMID- 11103921 TI - Equity in the allocation of health care resources. AB - This paper examines some of the issues that arise when seeking to tackle health inequalities in a context of limited health care resources. Increasingly, central agencies are using devolved budgets for health care providers as a central instrument of expenditure control. Equity objectives play an important role in the resource allocation methods used to determine such budgets. Yet, unless integrated into a proper system of risk management, the use of budgets can lead to serious inequity. The paper discusses the potential contributions of different disciplines to promoting equity within a health care budgetary regime. PMID- 11103922 TI - Rationing for health equity: is it necessary? PMID- 11103923 TI - The need for an interdisciplinary perspective on the social determinants of health. PMID- 11103924 TI - Equity in health: a challenge for researchers and policy makers. PMID- 11103925 TI - Interpreting the rational addiction model. AB - The rational addiction (RA) model of Becker and Murphy (Becker GS, Murphy KM. A theory of rational addiction. J Pol Econ 1988; 96(4): 675-700) has rapidly become one of the standard models in the literature on addictive behaviour. This paper reviews some theoretical issues surrounding its use, and indicates areas in which caution should be used in applying this model. PMID- 11103926 TI - A Bayesian approach to economic analyses of clinical trials: the case of stenting versus balloon angioplasty. AB - New results about the costs and effects of a new therapy may be weighted with prior information. As such, classical confidence intervals surrounding the costs and effects of a therapy may not reflect the real uncertainties. Bayesian techniques may improve this by formalizing the way that prior information is taken into account in assessing the new evidence. Costs and effects can be analysed separately, but also, when considering the balance between costs and effects, they can be analysed simultaneously. Here, an example is given using data from two trials that compared costs and effectiveness of stent implantation with balloon angioplasty. The Bayesian results make it clear that different prior distributions may lead to different decisions, and it is concluded that even Bayesian analysis may not always reflect the process of capturing the remaining uncertainties. PMID- 11103927 TI - The social value of health programmes: is age a relevant factor? AB - In cost-effectiveness analysis (CEA) it is usually assumed that a quality adjusted life-year (QALY) is of equal value to everybody, irrespective of the patient's age. However, it is possible that society assigns different social values to a QALY, according to who gets it. In this paper, we discuss the possibility of weighting health benefits for age in CEA. We also examine the possibility that age-related preferences depend on the size of the health gain. An experiment was performed to test these hypotheses. The assessment of results suggests that the patient's age is a relevant factor when assessing health gains. PMID- 11103928 TI - Definition, interpretation and calculation of cost-effectiveness acceptability curves. AB - This paper discusses the definition, interpretation and computation of cost effectiveness (CE) acceptability curves. A formal definition of the CE acceptability curve based on the net benefit approach is provided. The curve can be computed using parametric or non-parametric techniques and for both computational approaches we establish a formal relation between the CE acceptability curve and statistical inference based on confidence intervals and P values in CE analysis. PMID- 11103929 TI - Institutional considerations in priority setting: transactions cost perspective on PBMA. AB - Programme budgeting and marginal analysis (PBMA) is increasingly being used as a method of priority setting in the health care sector. Despite this, PBMA has, on occasions, been subject to problems in its application which can be seen as being 'institutional' in nature. This paper examines the extent to which the institutional setting of PBMA affects the way in which it can be conducted. In particular, a transactions costs perspective is taken to analyse the extent to which variation in such costs can alter the incentives of the individual participants. A number of recommendations for improving the sustainability of such projects is then provided. Following this, the implications which this 'institutional' approach has for the evaluation of PBMA are set out. PMID- 11103930 TI - The price of placements in residential and nursing home care: the effects of contracts and competition. AB - A variety of contract types are used in the placement of elderly people in residential and nursing care homes in the UK. Contracts vary according to how and when providers are paid. Among other things, prices can be made contingent on the total quantity of service to be purchased and on production cost characteristics. They can be determined at the time of placement or in advance. The primary objective of this paper is to assess the impact of contract choices on the price of placements. Regression analysis was conducted on a final sample of 1780 publicly funded placements made in 18 local authorities in the UK over a 6-month period ending in early 1996. Controlling factors included in the price analysis were production cost indicators and those measuring market competitiveness. Choices of both quantity and cost contingent contracts were found to be significantly associated with placement prices. The findings support the hypothesis that contract payment arrangements have different risk, insurance and information properties, and so have implications for the performance of residential care providers. PMID- 11103931 TI - IC261, a specific inhibitor of the protein kinases casein kinase 1-delta and epsilon, triggers the mitotic checkpoint and induces p53-dependent postmitotic effects. AB - The p53-targeted kinases casein kinase 1delta (CK1delta) and casein kinase 1epsilon (CK1epsilon) have been proposed to be involved in regulating DNA repair and chromosomal segregation. Recently, we showed that CK1delta localizes to the spindle apparatus and the centrosomes in cells with mitotic failure caused by DNA damage prior to mitotic entry. We provide here evidence that 3-[(2,4,6 trimethoxyphenyl)methylidenyl]-indolin-2-one (IC261), a novel inhibitor of CK1delta and CK1epsilon, triggers the mitotic checkpoint control. At low micromolar concentrations IC261 inhibits cytokinesis causing a transient mitotic arrest. Cells containing active p53 arrest in the postmitotic G1 phase by blockage of entry into the S phase. Cells with non-functional p53 undergo postmitotic replication developing an 8N DNA content. The increase of DNA content is accompanied by a high amount of micronucleated and apoptotic cells. Immunfluorescence images show that at low concentrations IC261 leads to centrosome amplification causing multipolar mitosis. Our data are consistent with a role for CK1delta and CK1epsilon isoforms in regulating key aspects of cell division, possibly through the regulation of centrosome or spindle function during mitosis. PMID- 11103933 TI - A cancer modifier role for parathyroid hormone-related protein. AB - The parathyroid hormone-related protein (PTHrP) gene (Pthlh) maps in the distal region of mouse chromosome 6 that contains a quantitative trait locus associated with genetic predisposition to skin tumorigenesis. Here, we report a genetic polymorphism located in the osteostatin encoding region of the Pthlh gene and that produces Thr/ Pro PTHrP variants. PthlhThr and PthlhPro alleles were significantly linked with resistance and susceptibility to skin carcinogenesis in phenotypically selected Car-R and Car-S outbred mice. Transfection of human NCI H520 squamous cell carcinoma cells with the PthlhPro allele resulted in cells growing in clusters, tending to pile up, and growing at a significantly faster rate in nude mice than non-transfected and PthlhThr-transfected cells. These results point to the role of the Pthlh gene as a cancer modifier gene in skin tumorigenesis. PMID- 11103932 TI - Reduction of Cdc25A contributes to cyclin E1-Cdk2 inhibition at senescence in human mammary epithelial cells. AB - Replicative senescence may be an important tumor suppressive mechanism for human cells. We investigated the mechanism of cell cycle arrest at senescence in human mammary epithelial cells (HMECs) that have undergone a period of 'self selection', and as a consequence exhibit diminished p16INK4A levels. As HMECs approached senescence, the proportion of cells with a 2N DNA content increased and that in S phase decreased progressively. Cyclin D1-cdk4, cyclin E-cdk2 and cyclin A-cdk2 activities were not abruptly inhibited, but rather diminished steadily with increasing population age. In contrast to observations in fibroblast, p21Cip1 was not increased at senescence in HMECs. There was no increase in p27Kip1 levels nor in KIP association with targets cdks. While p15INK4B and its binding to both cdk4 and cdk6 increased with increasing passage, some cyclin D1-bound cdk4 and cdk6 persisted in senescent cells, whose inhibition could not be attributed to p15INK4B. The inhibition of cyclin E-cdk2 in senescent HMECs was accompanied by increased inhibitory phosphorylation of cdk2, in association with a progressive loss of Cdc25A. Recombinant Cdc25A strongly reactivated cyclin E-cdk2 from senescent HMECs suggesting that reduction of Cdc25A contributes to cyclin E-cdk2 inhibition and G1 arrest at senescence. Although ectopic expression of Cdc25A failed to extend the lifespan of HMECs, the exogenous Cdc25A appeared to lack activity in these cells, since it neither shortened the G1-to-S phase interval nor activated cyclin E-cdk2. In contrast, in the breast cancer-derived MCF-7 line, Cdc25A overexpression increased both cyclin E-cdk2 activity and the S phase fraction. Thus, mechanisms leading to HMEC immortalization may involve not only the re-induction of Cdc25A expression, but also activation of this phosphatase. PMID- 11103934 TI - Reduced latency but no increased brain tumor penetrance in mice with astrocyte specific expression of a human p53 mutant. AB - p53-germline mutations located in the core DNA-binding domain have been associated with a more dominant tumor penetrance especially for breast cancer and brain tumors. We previously reported an unusual accumulation of CNS tumors associated with a unique p53 germline mutation, Y236delta (deletion of codon 236). To test whether this tissue-specific tumor predisposition reflects a gain of-function activity of Y236delta, we generated transgenic mice expressing Y236delta in astrocytes using the regulatory elements of the glial fibrillary acidic protein (GFAP) gene. After transplacental exposure to N-ethyl-N nitrosourea (25 mg/kg BW) brain tumors developed in 18% (7/39) of GFAP-Y236delta transgenic p53-/- mice, while in p53+/- mice the incidence was 28% (11/40) (P>0.3). However, the mean tumor latency for GFAP-Y236delta/p53+/- mice was significantly shorter than for p53+/- mice, with 19.9 weeks vs 31.6 weeks (P=0.039), respectively. Taken together, cell specific expression of Y236delta results in an acceleration of tumor progression but does not confer a higher tumor penetrance. Conceivably, the transdominant effect of Y236delta provided a growth advantage early in the progression of neoplastic cells, since the endogenous p53 wild-type allele was lost in all brain tumors independent of the genotype. This reflects well observations from human astrocytic neoplasms with p53 mutations. PMID- 11103936 TI - Regulation of a multigenic invasion programme by the transcription factor, AP-1: re-expression of a down-regulated gene, TSC-36, inhibits invasion. AB - The transcription factor AP-1 (activator protein-1) is required for transformation by many oncogenes, which function upstream of it in the growth factor-ras signal transduction pathway. Previously, we proposed that one role of AP-1 in transformation is to regulate the expression of a multigenic invasion programme. As a test of this proposal we sought to identify AP-1 regulated genes based upon their differential expression in 208F rat fibroblasts transformed by FBR-v-fos (FBR), and to determine if they functioned in the invasion programme. Subtracted cDNA libraries specific for up- or down-regulated genes in FBRs compared to 208Fs were constructed and analysed. Northern analysis revealed that the cDNAs in both libraries represented differentially expressed genes. Nucleic acid sequence analysis of randomly selected cDNA clones from each library coupled with searches of nucleic acid and amino acid sequence databases determined that many of the cDNAs represented proteins that function in various aspects of the invasion process. Functional analysis of one the down-regulated genes, TSC 36/follistatin-related protein (TSC-36/Frp), which has not previously been associated with invasion, demonstrated that its expression in FBRs inhibited in vitro invasion. These results support the proposal that AP-1 in transformed cells regulates a multigenic invasion programme. PMID- 11103935 TI - Loss of p21WAF1/CIP1 accelerates Ras oncogenesis in a transgenic/knockout mammary cancer model. AB - Upregulation of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 and subsequent cell growth arrest or senescence is one mechanism by which normal cells are believed to respond to stress induced by the constitutively activated GTPase Ras. We hypothesize that in the absence of p21, the onset of Ras-dependent oncogenesis is accelerated. To test this hypothesis, we crossed MMTV/v-Ha-ras transgenic mice into a p21-deficient background. By 63 days of age, all 8 ras/p21-/- mice developed either malignant (mammary and/or salivary adenocarcinomas) or benign (Harderian hyperplasia) tumors. In contrast, by the same age, only one out of nine of the ras/p21+/+ mice developed a tumor. Furthermore, by 94 days of age, half of the ras/p21-/- mice, but none of the ras/p21+/+ mice, developed mammary tumors. p21-deficiency also accelerated the development of salivary (T50=66 days for ras/p21-/- vs T50=136 days for ras/p21+/+) and Harderian (T50=52 days for ras/p21-/- vs T50>221 days for ras/p21+/+) tumors. Furthermore, two out of the eight ras/p21-/- mice had metastatic lesions, one in its lungs, the other in its abdomen. None of the nine ras/p21+/+ mice had metastatic lesions. By 4 months of age, the mammary tumor multiplicity was 10-fold greater in ras/p21-/- (average 3.40 tumors/mouse) than in ras/p21+/+ (average 0.33 tumor/mouse) mice. However, once the tumors appeared, their growth rate, apoptosis level, and mitotic index were not affected by the loss of p21, suggesting that loss of p21 is critical in early but not late events of Ras oncogenesis. Altogether, the results show that tumor onset in MMTV/v-Ha-ras mice is p21-dependent with loss of p21 associated with earlier tumor appearance and increased tumor multiplicity and aggressiveness. PMID- 11103937 TI - Adenovirus-mediated p16INK4a gene expression radiosensitizes non-small cell lung cancer cells in a p53-dependent manner. AB - We examined the influence of adenovirus-mediated wild-type p16INK4a (Ad/p16) expression on the radiation sensitivity of NSCLC cell lines, all of which lacked constitutive p16INK4a but each of which varied in p53 status: A549 (-p16INK4a/ +pRb/wt-p53), H322 (-p16INK4a/ +pRb/mt-p53), and H1299 (-p16INK4a/ +pRb/deleted p53). The in vitro clonogenic survival results indicate that Ad/p16 enhanced the radiosensitivity of A549 but not H322 or H1299. Further analysis indicated that the apoptosis induced by combination therapy using Ad/p16 plus irradiation was dependent on the endogenous p53 status of the cancer cells. We performed Western blotting to analyse the p53 protein expression of A549 cells treated with either Ad/p16 or Ad/Luc. Endogenous p53 protein levels were higher in A549 cells transfected with Ad/p16 than in those transfected with Ad/Luc. Furthermore, when wt-p53 protein expression was restored in H1299 using Ad/ p53, Ad/p16 stabilized p53 protein expression and radiosensitized the cells. These results suggest that Ad/ p16-induced stabilization of p53 protein may play an important role in Ad/p16 mediated radiosensitization by enhancing or restoring apoptosis properties. Thus, Ad/ p16 plus radiation in combination may be a useful gene therapy strategy for tumors that have wt-p53 but nonfunctional p16INK4a. PMID- 11103938 TI - Oncogenic insertional mutations in the P-loop of Ras are overactive in MAP kinase signaling. AB - Mutations of Ras with three extra amino acids inserted into the phosphate-binding (P) loop have been investigated both in vitro and in vivo. Such mutants have originally been detected as oncogenes both in the ras and the TC21 genes. Biochemical experiments reveal the molecular basis of their oncogenic potential: the mutants show a strongly attenuated binding affinity for nucleotides, most notably for GDP, leading to a preference for GTP binding. Furthermore, both the intrinsic as well as the GAP-stimulated GTP hydrolysis are drastically diminished. The binding interaction with GAP is reduced, whereas binding to the Ras-binding domain of the downstream effector c-Raf1 is not altered appreciably. Microinjection into PC12 cells shows the mutants to be as potent to induce neurite outgrowth as conventional oncogenic Ras mutants. Unexpectedly, their ability to stimulate the MAP kinase pathway as measured by a reporter gene assay in RK13 cells is much higher than that of the normal oncogenic mutant G12V. This characteristic was attributed to an increased stimulation of c-Raf1 kinase activity by the insertional Ras mutants. PMID- 11103939 TI - Synchronous and regulated expression of two AU-binding proteins, AUF1 and HuR, throughout murine development. AB - The AUF1 (hnRNPD) and HuR (ELAV-like) proteins, potential trans-acting factors for regulated mRNA decay, bind in vitro to A+U-rich elements (AREs) found in the 3' untranslated region (3' UTR) of many labile transcripts. In an effort to determine whether these trans-acting factors are likely to play a role in embryogenesis, we have analysed their expression during mouse development both at the mRNA and protein levels. We show that AUF1 and HuR are expressed at all the developmental stages analysed from day 8.5 of embryonic development to adulthood. Expression levels are dynamic, varying between tissues and developmental stages. However, a strong positive correlation between AUF1 and HuR protein levels was observed in all examined tissues. Finally, we compared AUF1 and HuR expression with accumulation of one common target mRNA, c-myc. The similar spatio-temporal distribution of these proteins and of c-myc mRNA is in agreement with a potential concerted role in ARE-mediated control of mRNA stability. PMID- 11103940 TI - Differential requirements of the MAP kinase and PI3 kinase signaling pathways in Src- versus insulin and IGF-1 receptors-induced growth and transformation of rat intestinal epithelial cells. AB - There have been few studies on the specific signaling pathways involved in the transformation of epithelial cells by oncogenic protein tyrosine kinases. Here we investigate the requirement of MAP (MAPK) and phosphatidylinositol 3- (PI3K) kinases in the transformation of rat intestinal epithelial (RIE) cells by oncogenic forms of insulin receptor (gag-IR), insulin-like growth factor-1 receptor (gag-IGFR), and v-Src. MAPK is not significantly activated in cells transformed by gag-IR and gag-IGFR but is activated in v-Src transformed cells. Treatment with PD98059, a MEK inhibitor, at concentrations where MAPK activity was reduced below the basal level showed that MAPK is partially required for the monolayer growth of parental and transformed RIE cells. However, MAPK is not essential for the focus forming ability of the three oncogene-transformed cells. It is also not necessary for the colony forming ability of gag-IR- and gag-IGFR-, but is partially required for v-Src-transformed cells. PI3K is significantly activated in all three oncogene transformed RIE cells. LY294002, a PI3K inhibitor, potently inhibited monolayer growth of all three oncogene-transformed cells. However, at concentrations of LY294002 where activated forms of Akt, a downstream component of the PI3K pathway, were undetectable, colony and focus forming abilities of the v-Src-RIE cells were only slightly affected whereas those of gag-IR/IGFR-RIE cells were greatly inhibited. These results were confirmed using a different pharmacological inhibitor, wortmannin, and a dominant negative form of PI3K, Ap85. Similarly, rapamycin, known to inhibit p70S6 kinase, a downstream component of the PI3K-Akt pathway, also inhibited gag-IR/IGFR induced, but not v-Src-induced, focus and colony formation. We conclude that the MAPK and PI3K signaling pathways are differentially required for transformation of RIE cells by oncogenic IR and IGFR versus Src and the pattern of requirements is different from that of fibroblast transformation. PMID- 11103941 TI - Chimeric VEGFRs are activated by a small-molecule dimerizer and mediate downstream signalling cascades in endothelial cells. AB - Despite much interest in vascular endothelial growth factor (VEGF) and its receptors (VEGFRs -1 and -2), VEGF-induced signalling cascades remain incompletely defined. Attempts to assign individual responses to a particular receptor have used either transfected cell lines, receptor-specific growth factors or antisense oligonucleotides. Such studies have attributed the majority of VEGF-induced responses to activation of VEGFR-2. As a consequence of poor growth factor-induced VEGFR-1 autophosphorylation however, observations from these studies may instead reflect the relative activation of the two receptors. We have generated novel chimeric VEGF receptors in which the dimerization domain of the B subunit of DNA gyrase is fused to the cytoplasmic domain of VEGFRs -1 and -2. When expressed in porcine aortic endothelial cells, both chimeric VEGFR-1 and -2 autophosphorylate in response to addition of the small-molecule dimerizing agent, coumermycin. Once activated, both receptors induce downstream signalling cascades, exemplified here by the activation of MAPK, PLCgamma and PKB/Akt. Furthermore, we demonstrate that the Y1175 residue of VEGFR-2 is essential for the activation of PLCgamma mediated by this chimeric receptor. In contrast to previous reports which show a limited ability of VEGFR-1 to mediate signalling cascades, we show that once sufficiently activated, VEGFR-1 signals in a similar manner to VEGFR-2 in endothelial cells. PMID- 11103942 TI - MMAC1/PTEN inhibits cell growth and induces chemosensitivity to doxorubicin in human bladder cancer cells. AB - The development and progression of bladder cancer is associated with multiple alterations in the genome, including loss of chromosome 10. Recently, MMAC1/PTEN, a phosphatidylinositol phosphatase, has been mapped to chromosome 10q23. We previously demonstrated that MMAC1/PTEN has tumor suppressive properties in glioblastoma and prostate cancer. To investigate the efficacy of gene therapy with MMAC1/PTEN, we examined whether the exogenous introduction of MMAC1/PTEN via an adenoviral vector (Ad-MMAC) can inhibit tumor growth and reverse drug resistance to doxorubicin in human bladder cancer cells. Human bladder cancer cell lines UM-UC-3 and T24 were infected with Ad-MMAC to induce exogenous expression of MMAC1/PTEN. The cells were then analysed for cell growth and expression of phosphorylated protein kinase B (Akt/PKB) and MMAC1/PTEN. UM-UC 6dox, a doxorubicin resistant subline, was infected with Ad-MMAC to evaluate its role in reversing drug resistance to doxorubicin. We found that MMAC1/PTEN suppressed tumor growth in UM-UC-3 and T24 cells with arrest in the G1 phase of the cell cycle. We also showed that gene therapy with MMAC1/PTEN abrogated phosphorylated Akt/PKB expression in UM-UC-3, T24 and UMUC-6dox cells, and restored doxorubicin sensitivity in UM-UC-6dox. These data demonstrate that MMAC1/PTEN can induce growth suppression and increase sensitivity to doxorubicin in bladder cancer cells and suggest that the MMAC1/PTEN gene and its pathways can be therapeutic targets for bladder cancer. PMID- 11103943 TI - Loss of heterozygosity, allele silencing and decreased expression of p73 gene in breast cancers: prevalence of alterations in inflammatory breast cancers. AB - The p73 gene is a p53 homologue located at 1p36-33, a region submitted to deletions in breast cancer (BC) and putatively imprinted. To study whether p73 was associated with breast carcinogenesis, loss of heterozygosity (LOH), allele expression and transcript levels were assessed in 59 BC, including 39 BC presenting no inflammatory symptoms (NBC) and 20 inflammatory BC (IBC). IBC is a rare but aggressive form of cancer with a very poor prognosis. Normal breast epithelium (BE) and lymphocytes from patients were used as controls. StyI polymorphism generating GC and/or AT alleles was used to select 22 heterozygous patients. p73 LOH was significantly higher in IBC than in NBC [five of eight cases (62%) versus two of 14 cases (14%); Fisher's exact test, P=0.05]. p73 was biallelically expressed in all BE. In contrast, 12 of 16 (75%) BC were monoallelically expressed, showing that allele silencing was significantly associated with breast carcinogenesis (P=0.012), AT being the preferential silent allele (10 out of 12 tumours). p73 mRNA levels in NBC and IBC were two- and threefold lower than in BE, respectively, suggesting that decreased expression could be related to tumour aggressiveness. In conclusion, LOH, allele silencing and decreased expression of the p73 gene may play a role in breast carcinogenesis. PMID- 11103945 TI - Pattern formation in the cerebellar cortex. AB - The cerebellar cortex is subdivided rostrocaudally and mediolaterally into a reproducible array of zones and stripes. This makes the cerebellum a valuable model for studying pattern formation in the vertebrate central nervous system. The structure of the adult mouse cerebellar cortex and the series of embryological events that generate the topography are reviewed. PMID- 11103944 TI - New thoughts on the role of the beta-gamma subunit in G-protein signal transduction. AB - Heterotrimeric G proteins are involved in numerous biological processes, where they mediate signal transduction from agonist-bound G-protein-coupled receptors to a variety of intracellular effector molecules and ion channels. G proteins consist of two signaling moieties: a GTP-bound alpha subunit and a beta-gamma heterodimer. The beta-gamma dimer, recently credited as a significant modulator of G-protein-mediated cellular responses, is postulated to be a major determinant of signaling fidelity between G-protein-coupled receptors and downstream effectors. In this review we have focused on the role of beta-gamma signaling and have included examples to demonstrate the heterogeneity in the heterodimer composition and its implications in signaling fidelity. We also present an overview of some of the effectors regulated by beta-gamma and draw attention to the fact that, although G proteins and their associated receptors play an instrumental role in development, there is rather limited information on beta gamma signaling in embryogenesis. PMID- 11103946 TI - Attraction vs. repulsion: the growth cone decides. AB - Axons are guided through their environment in response to signals provided by extracellular cues. These cues are transduced into motile responses by the tip of the growing axon, the growth cone, and can be either repulsive or attractive in nature. Recent studies have suggested that how an axon responds to any given signal depends on the internal state of the growth cone. This review discusses these studies and their importance for understanding how nerve connections are made in the developing embryo. PMID- 11103947 TI - Cellular mechanisms of netrin function: long-range and short-range actions. AB - Netrins are secreted proteins that direct axon extension and cell migration during neural development. They are bifunctional cues that act as an attractant for some cell types and as a repellent for others. Several lines of evidence suggest that two classes of receptors, the deleted in colorectal cancer (DCC) family and the UNC-5 family, mediate the attractant and repellent response to netrin. Although netrins were first identified as diffusible long-range cues for developing axons, recent findings provide evidence that they also function as short-range cues close to the surface of the cells that produce them. This short range function of netrin contributes to guiding neurite outgrowth and mediating cell-cell interactions during development and perhaps also in adults. PMID- 11103948 TI - Regulation and function of FGF8 in patterning of midbrain and anterior hindbrain. AB - In this article, an adjunct to a platform presentation at the Winternational 2000 Symposium, we summarize the recent findings of this group concerning the regulation and functions of FGF8 expressed at the isthmus of the developing brain. We show that several different FGF8 isoforms, ectopically expressed in midbrain or posterior forebrain, are able to mimic the proliferative and patterning functions previously attributed to the isthmus in tissue grafting studies. Moreover, we also show that FGF8 protein is sufficient to induce an ectopic isthmic organiser (Fgf-8+, Gbx2+) in anterior midbrain. We also provide evidence that isthmic FGF8 patterns anterior hindbrain, repressing Hox-a2 expression and setting aside a territory of the brain that includes the cerebellar anlage. We show that these effects of FGF8 are likely to be mediated via FGFR1 and be modulated by the putative FGF antagonist, Sprouty2, identified using a differential display screen. Finally, we provide evidence that the onset of Fgf8 expression is regulated by En1 and that its expression at the isthmus is subsequently maintained by a specific and direct interaction between rhombomere 1 and midbrain. PMID- 11103949 TI - Hedgehog signaling and the axial patterning of Drosophila wings. AB - Growth and cell fate in the anterior-posterior (A/P) axis of the developing wing of Drosophila melanogaster are controlled by a stripe of cells bisecting the axis called the A/P organizer. Hedgehog (Hh) signaling from posterior to anterior cells induces the organizer. Several Hh-responsive genes expressed by cells of the organizer mediate its patterning activity. The Hh-signaling pathway controls the post-translational modification of the transcription factor Cubitus interruptus (Ci) and the resulting local activation of Ci is required for the correct location of the A/P organizer. PMID- 11103950 TI - Distal-less-related homeobox genes of vertebrates: evolution, function, and regulation. AB - Homeobox genes of the Distal-less family have been identified in virtually all metazoan groups where they play roles in the ontogeny of these animals. The vertebrate Distal-less related genes (Dlx genes) are thought to have arisen as a result of a tandem gene duplication event followed by a number of larger genomic scale duplications and thus represent an interesting model with which to study the evolution of clustered gene families. Dlx genes are involved in the development of the forebrain, branchial arches, sensory organs, and limbs. Here we describe the current state of knowledge of the Dlx genes in terms of their developmental expression, how this expression is regulated and how the products of these genes function, once expressed. We highlight a number of recent studies that have shed light on the transcriptional regulation of this gene family. These findings have not only contributed to our understanding of the selective pressures involved in the maintenance of familial gene clustering in genomes, but also to our understanding of how genes may diverge in function during the course of evolution as a result of divergence of regulatory mechanisms. PMID- 11103951 TI - Semaphorin function in the developing invertebrate peripheral nervous system. AB - Different members of the semaphorin family of secreted and transmembrane guidance molecules play important and diverse roles during neuronal development. Within the developing grasshopper limb bud, two semaphorins are expressed in relatively non-overlapping and distinct expression patterns. The establishment of the tibial sensory projection within the limb bud relies on the combinatorial action of both semaphorins. In this review, we describe the function of the two semaphorins in axonal guidance and propose that a hierarchy of cues guide sensory neurons in the developing peripheral nervous system. PMID- 11103952 TI - Neurons from stem cells: implications for understanding nervous system development and repair. AB - Neurodegenerative diseases cost the economies of the developed world billions of dollars per annum. Given ageing population profiles and the increasing extent of this problem, there has been a surge of interest in neural stem cells and in neural differentiation protocols that yield neural cells for therapeutic transplantation. Due to the oncogenic potential of stem cells a better characterisation of neural differentiation, including the identification of new neurotrophic factors, is required. Stem cell cultures undergoing synchronous in vitro neural differentiation provide a valuable resource for gene discovery. Novel tools such as microarrays promise to yield information regarding gene expression in stem cells. With the completion of the yeast, C. elegans, Drosophila, human, and mouse genome projects, the functional characterisation of genes using genetic and bioinformatic tools will aid in the identification of important regulators of neural differentiation. PMID- 11103953 TI - Genetic and biochemical diversity in the Pax gene family. AB - The mammalian Pax gene family comprises nine members that are characterized by a conserved DNA-binding motif, the paired domain, which was originally described in the Drosophila protein paired. Both loss- and gain-of-function studies reveal that Pax genes carry out essential roles during embryogenesis, and in some instances, may function as master regulatory genes. This review focuses on both genetic and biochemical aspects of the Pax family, and emphasizes important differences in the activity of individual Pax genes and their protein products. PMID- 11103954 TI - More on cortical strut grafting for thigh pain. PMID- 11103955 TI - MRI and arthroscopic comparison of chondral lesions. PMID- 11103956 TI - The demise of value: reasons against Medicare E&M guidelines. PMID- 11103957 TI - Radiologic case study. Subscapularis tendon tear. PMID- 11103958 TI - The figure-of-four view to evaluate ACL injury. AB - Standard arthroscopic assessment of the anterior cruciate ligament (ACL) injury through an anterior view can be sub-optimal for evaluation of the femoral origin, particularly the posterior component. The figure-of-four view provides increased exposure to the posterolateral aspect of the intercondylar notch, thereby facilitating diagnosis of proximal ACL injury and avulsions of the ACL origin. PMID- 11103959 TI - The significance of stem-cement loosening of grit-blasted femoral components. AB - This study analyzed 15 patients who underwent revision for loosening at the stem cement interface. The femoral components were from the same manufacturer and had grit-blast roughened surfaces. An apparent radiographic deficiency in the cement mantle was present in at least one zone in 1 3 patients. In 9 of 12 patients with localized osteolysis, the osteolysis developed in a zone with an apparent radiographic cement mantle defect. Loosening occurred due to tension failure of the stem-cement interface followed by axial subsidence and movement into relative retroversion. Motion between the stem and the cement mantle fueled an abrasive wear mechanism between the roughened metal surface and the cement mantle, generating excessive metal and cement particles that gained access to endosteal bone via defects in the cement mantle and resulting in localized osteolysis. Although the roughened surface played a central role in these failures, it is unlikely the layer of polymethylmethacrylate (precoat) played a role in the mechanism of failure. In some cases, debonding occurred as a result of tension failure of the metal-precoat interface. In others, tension failure occurred within the cement mantle, leaving the precoat and some cement from the mantle on the stems. There was no difference in the mechanism of failure of stems with precoat proximally compared to stems with precoat proximally and distally. One stem had no precoat; findings in this patient were indistinguishable from the others. The significance of debonding depends on the surface roughness of the stem. Debonding carries a poorer prognosis with a rougher stem surface because of abrasive wear with the generation of numerous metal and cement particulates, which can lead to rapid osteolysis if there are cement mantle defects. Stems with a higher metal-cement bond strength may require a higher quality cement mantle for long-term success. PMID- 11103960 TI - Medium- and long-term results of open reduction and internal fixation for unstable pelvic ring fractures. AB - Over a 10-year period, 74 patients with unstable pelvic injuries were treated with open reduction and internal fixation. Radiographic and clinical follow-up averaged 71 months (range: 38-141 months). Satisfactory (ie, good and very good) radiographic results were obtained in 90% of patients. Clinical results were superior in patients without associated injuries (P=.05-.001). Most of the complications in this series were due to associated injuries. Sepsis was mostly due to open pelvic injuries and malunion to either lack of patient cooperation or inadequate open reduction and internal fixation. Careful preoperative analysis of the nature of the pelvic injury and selection of the appropriate operative technique for open reduction and internal fixation result in a satisfactory outcome for the majority of operative patients. PMID- 11103961 TI - Expression of nm23 protein in human osteosarcoma in relationship with early metastasis. AB - Nm23 protein expression was analyzed by immunohistochemical staining using formalin-fixed, paraffin-embedded sections from 39 cases with osteosarcomas and compared with the histologic findings and early metastasis for the purpose of detecting nm23 expression in osteosarcoma and elucidating the clinical significance of its expression. Immunoreactivity of nm23 protein was detected in 48.7% of the total cases. There was no statistical difference between nm23 expression and early metastasis, but there was a trend for cases with nm23 expression to progress to early metastasis within 1 year after operation. The role of nm23 as a tumor metastasis suppressor in osteosarcomas appeared less prominent. PMID- 11103962 TI - Musculoskeletal complications of Crohn's disease: the role of computed tomography in diagnosis and patient management. AB - The delayed diagnosis of musculoskeletal complications of Crohn's disease may produce major morbidity in patients. This study compared abdominal and pelvic computed tomography (CT) with conventional radiography in the diagnosis of musculoskeletal complications in 23 of 552 patients with Crohn's disease examined by CT over a 7-year period. Surgical confirmation was available in 15 of 21 patients. The clinical features of psoas/gluteal abscesses, abdominal wall fistulae, and sacral osteomyelitis are described. Because the clinical manifestations of these musculoskeletal complications are often nonspecific, CT is often useful in diagnosing and directing therapeutic interventions. PMID- 11103963 TI - Endoscopic harvesting of the sural nerve graft: a cadaveric investigation. AB - This study investigated an endoscopic technique of harvesting the sural nerve graft. Using endoscopic instrumentation, the sural nerve was harvested from six cadaveric legs. A 2-cm longitudinal incision was made immediately posterior to the lateral malleolus, and a 5-mm endoscope was introduced. The path of the nerve was followed to the popliteal space, and nerve dissection was performed from proximal to distal. Air inflation of a balloon was used to enlarge the endoscopic cavity. The cavity created around the nerve was insufflated with carbon dioxide gas, allowing complete nerve isolation. Using a 0.5-cm transverse incision, the nerve was cut and removed. This endoscopic sural nerve grafting approach offers potential advantages such as less injury to soft tissues, decreased pain, nerve integrity preservation, and good aesthetic results. PMID- 11103964 TI - Anatomical variation of the posterior interosseous nerve: a cadaver dissection study. AB - An anatomical variation of the posterior interosseous nerve was found in a cadaver. The posterior interosseous nerve entered the supinator muscle 3 cm distal to the radiohumeral joint, but exited from two sites. Fifty percent of the nerve exited under the distal edge of the supinator muscle. The other 50% of the nerve pierced through the supinator muscle, 4.2 cm distal to the articular surface of the radial head and then joined the remaining posterior interosseous nerve as it emerged from the supinator muscle distally. Variations were not found concerning the order and the manner of branches to the muscles. This variation in the posterior interosseous nerve could be an additional compression site for this nerve and therefore responsible for some of the atypical presentations of symptoms and for partial recovery after surgical decompression. Careful surgical dissection is recommended to avoid injury to this branch. PMID- 11103965 TI - Salter-Harris type II distal tibia fracture. PMID- 11103966 TI - Subacromial impingement due to the locking bolt of a humeral nail. PMID- 11103967 TI - Salmonella osteomyelitis transmitted from an iguana. PMID- 11103968 TI - Secondary aneurysmal bone cyst simulating malignant transformation in fibrous dysplasia. PMID- 11103969 TI - Iliotibial band friction syndrome. AB - Overuse knee injuries are common, but ITBFS is often overlooked as a cause of lateral knee pain in an active population. Iliotibial band friction syndrome is an overuse injury usually seen in long distance runners, cyclists, and military personnel. The exact incidence of the syndrome has been estimated to range from 1.6%-52% depending on the population studied. The diagnosis is often made from a thorough history and clinical examination with an infrequent need for additional studies. Treatment is mostly conservative consisting of rest and anti inflammatory agents, with only the refractory cases requiring surgical resection of the impinging portion of the ITB. PMID- 11103970 TI - Response to nursing newspaper headlines. PMID- 11103971 TI - Critical thinking: beyond nursing process. PMID- 11103972 TI - Congruency in defining critical thinking by nurse educators and non-nurse scholars. AB - This study investigated nurse educators' definition of the concept critical thinking. A sample of 201 baccalaureate nurse educators in midwest nursing programs completed a questionnaire identifying their perception of critical thinking skills and characteristics, and their agreement with non-nurse critical thinking experts on items often considered to be critical thinking. This study found that nurse educators agreed with non-nurse critical-thinking experts on the skills and dispositions; however, significant differences were found between nurse educators and non-nurse experts regarding concepts related to critical thinking. Nurse educators were more likely to identify researching, problem solving, decision-making, and planning as critical thinking. Despite their assertion otherwise, it is apparent from this study that nurse educators have a different perception of critical thinking than scholars in other disciplines. This study suggests that practice disciplines such as nursing may perceive critical thinking differently than educators in nonpractice disciplines. PMID- 11103973 TI - A consensus statement on critical thinking in nursing. AB - The purpose of this study was to define critical thinking in nursing. A Delphi technique with 5 rounds of input was used to achieve this purpose. An international panel of expert nurses from nine countries: Brazil, Canada, England, Iceland, Japan, Korea, Netherlands, Thailand, and 23 states in the U.S. participated in this study between 1995 and 1998. A consensus definition (statement) of critical thinking in nursing was achieved. The panel also identified and defined 10 habits of the mind (affective components) and 7 skills (cognitive components) of critical thinking in nursing. The habits of the mind of critical thinking in nursing included: confidence, contextual perspective, creativity, flexibility, inquisitiveness, intellectual integrity, intuition, open mindedness, perseverance, and reflection. Skills of critical thinking in nursing included: analyzing, applying standards, discriminating, information seeking, logical reasoning, predicting and transforming knowledge. These findings can be used by practitioners, educators and researchers to advance understanding of the essential role of critical thinking in nursing. PMID- 11103974 TI - The impact of the use of inquiry-based learning as a teaching methodology on the development of critical thinking. AB - Problem-based learning (PBL) uses patients' problems to develop students' problem solving and clinical skills. Inquiry-based learning (IBL) was developed as a similar methodology that was more holistic and flexible. This study sought to determine if inquiry-based learning (IBL) enhances critical-thinking ability as measured by the Watson Glaser Critical Thinking Appraisal (WGCTA). The WGCTA was administered to 228 nursing students in the first semester and 257 students in the final semester of their program. When the scores were stratified into groups, the students in the low group showed a significant increase in mean score, no change in the medium group, and a significant drop for the high group. PMID- 11103975 TI - Language as a constitutive: critical thinking for multicultural education and practice in the 21st century. AB - Postmodern understandings of language can function as revolutionary critical thinking tools and enable multicultural education in a way yet to be resolutely embraced by the discipline. This thesis is illustrated with critical thinking examples relevant to topics in nursing education, such as maternal infant attachment, HIV prevention education, standardized instruments measuring quality of life and self-esteem, domain of person, and adolescent male identity formation. Working through postmodern positions on language produces important questions. It offers nursing provocative ways of thinking about education and provides radically different approaches to critical thinking and cultural competence. Capitalizing on postmodern sensibilities about language to create multicultural education and practice will take persistent self-reflective educational practices that question the ground that nursing stands on, as well as good intentions regarding a deep and broad embrace of complexly understood cultural competence. PMID- 11103976 TI - The relationship of critical-thinking skills and the clinical-judgment skills of baccalaureate nursing students. PMID- 11103977 TI - Fostering bachelor of nursing students' research skills using public health education. PMID- 11103978 TI - Sparking students' interest in the clinical relevance of qualitative research. PMID- 11103979 TI - [Natural history of antibiotic resistance of community bacteria]. PMID- 11103980 TI - [Environmental factors (antibiotics in animals, day care centers, etc...(]. PMID- 11103981 TI - [Antibiotic therapy policy at the national level in different countries]. PMID- 11103982 TI - [Collective bases: conclusions, synthesis and perspectives]. PMID- 11103983 TI - [Doses, length of treatment]. PMID- 11103984 TI - [Antibiotic therapy indications]. PMID- 11103985 TI - [Individual approaches: conclusions, synthesis and perspectives]. PMID- 11103986 TI - [Is Streptococcus pneumoniae and Haemophilus influenzae resistance reversible?]. PMID- 11103987 TI - [Is the penicillin and pneumococcal model transferable to the new quinolones?]. PMID- 11103988 TI - [Practical perspectives: conclusions, synthesis and perspectives]. PMID- 11103989 TI - New roles for structure in biology and drug discovery. PMID- 11103990 TI - Structural genomics programs at the US National Institute of General Medical Sciences. PMID- 11103991 TI - An overview of structural genomics. AB - With access to sequences of entire human genomes plus those of various model organisms and many important microbial pathogens, structural biology is on the verge of a dramatic transformation. Our newfound wealth of sequence information will serve as the foundation for an important initiative in structural genomics. We are poised to embark on a systematic program of high-throughput X-ray crystallography and NMR spectroscopy aimed at developing a comprehensive view of the protein structure universe. Structural genomics will yield a large number of experimental protein structures (tens of thousands) and an even larger number of calculated comparative protein structure models (millions). This enormous body of structural data will be freely available, and promises to accelerate scientific discovery in all areas of biological science, including biodiversity and evolution in natural ecosystems, agricultural plant genetics, breeding of farm and domestic animals, and human health and disease. PMID- 11103992 TI - Structural genomics in North America. AB - Structural genomics in North America has moved remarkably quickly from ideas to pilot projects. Just three years ago, the field was only a concept, independently being discussed by its many inventors. Now it is already a well-organized, increasingly-funded, consortium-based effort to determine protein structures on a large scale. PMID- 11103993 TI - Structural genomics in Europe: slow start, strong finish? AB - Structural genomics in Europe is slowly coming off the ground. So far, European Commission funding is restricted to methods development projects, whereas structural genomics programs are funded from national sources. The research outlook is more toward techniques for high-throughput structure analysis than toward a systematic coverage of protein fold space. At present, there is little European coordination of the structural genomics effort. PMID- 11103994 TI - Structural genomics projects in Japan. AB - Two major structural genomics projects exist in Japan. The oldest, the RIKEN Structural Genomics Initiative, has two major goals: to determine bacterial, mammalian, and plant protein structures by X-ray crystallography and NMR spectroscopy and to perform functional analyses with the target proteins. The newest, the structural genomics project at the Biological Information Research Center, focuses on human membrane proteins. PMID- 11103995 TI - Structural genomics in the biotechnology sector. AB - Commercial efforts in structural genomics focus on providing to pharmaceutical customers information that relates to the suitability of specific proteins as drug targets and the informed selection and refinement of lead compounds generated by high-throughput screening and rational approaches. These efforts follow a variety of business models and are impacted by activities in the public domain, recent technological advances, and the changing intellectual property landscape. PMID- 11103996 TI - Patent protection for protein structures and databases. AB - Patent protection is available for certain inventions in the field of structural genomics. A review of the patent application procedure is provided, and patentable aspects of protein structural information under US law are discussed. Strategic and international factors to consider when seeking patent protection for an invention also are presented. PMID- 11103997 TI - Creating a structural genomics consortium. AB - A group of multinational companies, together with the Wellcome Trust, is attempting to form a charitable organization, the Structural Genomics Consortium. The goal will be to obtain X-ray structures for a broad representation across families of human proteins and to place the structural coordinates in the public database. PMID- 11103998 TI - Structural genomics of RNA. AB - A detailed understanding of the functions and interactions of biological macromolecules requires knowledge of their molecular structures. Structural genomics, the systematic determination of all macromolecular structures represented in a genome, is focused at present exclusively on proteins. It is clear, however, that RNA molecules play a variety of significant roles in cells, including protein synthesis and targeting, many forms of RNA processing and splicing, RNA editing and modification, and chromosome end maintenance. To comprehensively understand the biology of a cell, it will ultimately be necessary to know the identity of all encoded RNAs, the molecules with which they interact and the molecular structures of these complexes. This report focuses on the feasibility of structural genomics of RNA, approaches to determining RNA structures and the potential usefulness of an RNA structural database for both predicting folds and deciphering biological functions of RNA molecules. PMID- 11103999 TI - The Protein Data Bank and the challenge of structural genomics. AB - The PDB has created systems for the processing, exchange, query, and distribution of data that will enable many aspects of high throughput structural genomics. PMID- 11104000 TI - Integrative database analysis in structural genomics. AB - An important aspect of structural genomics is connecting coordinate data with whole-genome information related to phylogenetic occurrence, protein function, gene expression, and protein-protein interactions. Integrative database analysis allows one to survey the 'finite parts list' of protein folds from many perspectives, highlighting certain folds and structural features that stand out in particular ways. PMID- 11104001 TI - Structural genomics for science and society. AB - The field of robotics is affecting structural biology, enabling the era of structural genomics. The potential impact on protein fold prediction, biology, protein engineering and medicine is immense. Unraveling mysteries in the protein structure universe will require a dedicated effort for decades to come with computational toxicology as possibly a century long challenge. PMID- 11104002 TI - Target selection for structural genomics. AB - Structural genomics aims to use high-throughput structure determination and computational analysis to provide three-dimensional models of every tractable protein. The process of choosing proteins for experimental structure characterization is known as target selection. In this nomenclature, the targets are regions of proteins to be studied by crystallography or NMR. Selection of the targets is principally a computational process of restricting candidate proteins to those that are tractable and of unknown structure, and prioritizing according to expected interest and accessibility. PMID- 11104003 TI - Protein production: feeding the crystallographers and NMR spectroscopists. AB - Protein purification efforts for structural genomics will focus on automation for the readily-expressed proteins, and process development for the more difficult ones, such as membrane proteins. Thousands of proteins are expected to be produced in the next few years. The purified proteins will be valuable reagents for the entire research community. PMID- 11104004 TI - Automation of X-ray crystallography. AB - Structure-based biological discovery is entering a new era with the development of industrialized macromolecular structure determination pipelines. Intense, highly focused X-rays from integrated synchrotron radiation beam lines combined with significant advances in protein expression, purification, and micro crystallization automation allow for the full streamlining of the traditionally tedious and time consuming process of determining the three dimensional structures of macromolecules. PMID- 11104005 TI - Current state of automated crystallographic data analysis. AB - A goal of structural biology--and of structural genomics in particular--is to improve the underlying methodology for high-throughput determination of three dimensional structures of biological macromolecules. Here we address issues related to the development, automation and streamlining of the process of macromolecular X-ray crystal structure solution. PMID- 11104006 TI - Protein NMR spectroscopy in structural genomics. AB - Protein NMR spectroscopy provides an important complement to X-ray crystallography for structural genomics, both for determining three-dimensional protein structures and in characterizing their biochemical and biophysical functions. PMID- 11104007 TI - Protein structure modeling for structural genomics. AB - The shapes of most protein sequences will be modeled based on their similarity to experimentally determined protein structures. The current role, limitations, challenges and prospects for protein structure modeling (using information about genes and genomes) are discussed in the context of structural genomics. PMID- 11104008 TI - From structure to function: approaches and limitations. AB - This review presents a summary of current approaches to extract functional information from structural data on proteins and their complexes. While structural homologs may reveal possible biochemical functions (which may be hidden at the sequence level), elucidating the exact biological role of a protein in vivo will only be possible by including other results, such as data on expression and localization. PMID- 11104009 TI - Biotransformation of linoleic acid by Clavibacter sp. ALA2: heterocyclic and heterobicyclic fatty acids. AB - Clavibacter sp. ALA2 transformed linoleic acid into a variety of oxylipins. In previous work, three novel fatty acids were identified, (9Z)-12, 13, 17 trihydroxy-9-octadecenoic acid and two tetrahydrofuran-(di)hydroxy fatty acids. In this report, we confirm the structures of the tetrahydrofuran-(di)hydroxy fatty acids by nuclear magnetic resonance as (9Z)-12-hydroxy-13,16-epoxy-9 octadecenoic acid and (9Z)-7,12-dihydroxy-13,16-epoxy-9-octadecenoic acid. Three other products of the biotransformation were identified as novel heterobicyclic fatty acids, (9Z)-12,17;13, 17-diepoxy-9-octadecenoic acid, (9Z)-7-hydroxy 12,17;13,17-diepoxy-9-octadecenoic acid, and (9Z)-12,17;13,17-diepoxy-16-hydroxy 9-octadecenoic acid. Thus, Clavibacter ALA2 effectively oxidized linoleic acid at C-7, -12, -13, -16, and/or -17. PMID- 11104010 TI - Production of eicosapentaenoic acid by a recombinant marine cyanobacterium, Synechococcus sp. AB - The eicosapentaenoic acid (EPA) synthesis gene cluster from an EPA-producing bacterium, Shewanella sp. SCRC-2738, was cloned into a broad-host range vector, pJRD215, and then introduced into a marine cyanobacterium, Synechococcus sp. NKBG15041c, by conjugation. The transconjugant cyanobacteria produced 3.7 +/- 0.2% (2.24 +/- 0.13 mg/L) EPA (n-3) and 2.5 +/- 0.2% (1.49 +/- 0.06 mg/L) eicosatetraenoic acid (n-3) of the total fatty acids when the cells were cultured at 23 degrees C at a light intensity of 1,000-1,500 Lux. The EPA and eico satetraenoic acid contents of the cells were increased to 4.6 +/- 0.6% (3.86 +/- 1.11 mg/L) and 4.7 +/- 0.3% (3.86 +/- 0.82 mg/L), and 7.5 +/- 0.3% (1.76 +/- 0.10 mg/L) and 5.1 +/- 0.2% (1.19 +/- 0.06 mg/L) when they were cultured at low temperature (18 degrees C) and at lower light intensity (40 Lux), respectively. PMID- 11104011 TI - Dietary conjugated linoleic acid did not alter immune status in young healthy women. AB - The purpose of this study was to examine whether conjugated linoleic acid (CLA) supplementation in human diets would enhance indices of immune status as reported by others for animal models. Seventeen women, 20-41 yr, participated in a 93-d study conducted in two cohorts of 9 and 8 women at the Metabolic Research Unit of Western Human Nutrition Research Center. Seven subjects were fed the basal diet (19, 30, and 51% energy from protein, fat, and carbohydrate, respectively) throughout the study. The remaining 10 subjects were fed the basal diet for the first 30 d, followed by 3.9 g CLA (Tonalin)/d for the next 63 d. CLA made up 65% of the fatty acids in the Tonalin capsules, with the following isomeric composition: t10, c12, 22.6%; c11, t13, 23.6%; c9, t11, 17.6%; t8, c10, 16.6%; and other isomers 19.6%. Most indices of immune response were tested at weekly intervals, three times at the end of each period (stabilization/intervention); delayed-type hypersensitivity (DTH) to a panel of six recall antigens was tested on study day 30 and 90; all subjects were immunized on study day 65 with an influenza vaccine, and antibody titers were examined in the sera collected on day 65 and 92. None of the indices of immune status tested (number of circulating white blood cells, granulocytes, monocytes, lymphocytes, and their subsets, lymphocytes proliferation in response to phytohemagglutinin, and influenza vaccine, serum influenza antibody titers, and DTH response) were altered during the study in either dietary group. Thus, in contrast to the reports with animal models, CLA feeding to young healthy women did not alter any of the indices of immune status tested. These data suggest that short-term CLA supplementation in healthy volunteers is safe, but it does not have any added benefit to their immune status. PMID- 11104012 TI - Absorption by rats of tocopherols present in edible vegetable oils. AB - The absorption of tocopherols (alpha, gamma, and delta) and fatty acids from rapeseed (RO), soybean (SOO), and sunflower (SUO) oil, both from the natural oils and from the oils following moderate heating (180 degrees C for 15 min), was measured in lymph-cannulated rats. Oils were administered as emulsions through a gastrostomy tube, and lymph samples were collected for 24 h. The composition of tocopherols in oils and lymph fractions was measured by high-performance liquid chromatography, and fatty acids were measured by gas-liquid chromatography. The highest accumulated transport of alpha-tocopherol was observed after SUO administration, the lowest after SOO, with RO in between, corresponding to their relative contents (41.6 +/- 8.8, 32.7 +/- 5.0, and 24.9 +/- 4.3 microg at 24 h after administration of SUO, RO, and SOO, respectively). The calculated recoveries (in %) 24 h after oil administration were 21.4 +/- 4.5, 45.7 +/- 7.0, and 78.8 +/- 13.5 for SUO, RO, and SOO, respectively, suggesting that the absorption efficiency decreased when the alpha-tocopherol concentration increased. The recovery of alpha-tocopherol was higher than the recoveries of gamma- and delta-tocopherol, indicating that the different tocopherols were not absorbed to the same extent or with similar rates. No differences between unheated and heated oils were observed in the absorption of tocopherols, whereas heating led to lower absorption of fatty acids, thus showing no direct association between absorption of tocopherols and fatty acids. PMID- 11104013 TI - Escherichia coli sepsis increases hepatic apolipoprotein B secretion by inhibiting degradation. AB - Sepsis leads to hypertriglyceridemia in both humans and animals. Previously, we reported that plasma very low density lipoprotein apolipoprotein (apo) B and hepatic production of apoB increased during Escherichia coli sepsis. The present experiments were undertaken to determine whether the altered hepatic secretion of apoB was associated with an increase in synthesis or a decrease in degradation rate. Sepsis was induced in male, Lewis rats (225-275 g) by intravenous injection of 3.8 x 10(8) live E. coli colonies/100 g body. Twenty-four hours later rats were sacrificed, and primary hepatocytes were prepared and incubated overnight with 35S-methionine. Hepatocytes from E. coli-treated rats secreted twice as much apoB-48 and total apoB than the hepatocytes from control rats. Escherichia coil sepsis increased cellular triglyceride mass by 86%, which was due to a stimulation in triglyceride synthesis from newly synthesized fatty acids, measured by 3H2O incorporation into triglycerides. The apoB synthesis rate, apoB mRNA levels, and apoB mRNA editing were not altered during E. coil sepsis. The pulse-chase experiments showed that the rate of apoB degradation decreased in E. coli-treated rats. These findings demonstrate that the secretion of apoB is regulated posttranslationally during E. coli sepsis by decreasing the degradation of newly synthesized apoB, which contributes to the development of hypertriglyceridemia. PMID- 11104014 TI - Cu2+ -induced low density lipoprotein peroxidation is dependent on the initial O2 concentration: an O2 consumption study. AB - Atherosclerotic plaques form in the arterial intima, where low density lipoprotein (LDL) is thought to be oxidatively modified at sites which may contain catalytic amounts of copper in the presence of low O2 tension. We have investigated O2 consumption during LDL peroxidation induced by Cu2+ ions in vitro and found two phases: a lag phase followed by a phase of rapid O2 consumption. The length of the lag phase was dependent on Cu2+ and on initial O2 concentrations; increasing either decreased the lag time; however, LDL. concentration had no effect. LDL-induced Cu2+ reduction, however, was not affected by low initial O2 concentrations, suggesting that O2 is not required for LDL-mediated reduction of Cu2+. Following the lag phase, O2 consumption was dependent upon LDL or initial O2 concentrations; Cu2+ concentrations had little effect, suggesting that the propagation phase is more dependent on the presence of LDL lipids and O2 as substrates for the reaction. In summary, LDL peroxidation takes place in the presence of Cu2+ at low O2 tension; however, the reaction is dependent upon initial O2 concentrations; increases shorten the lag phase and accelerate O2 consumption. PMID- 11104015 TI - Adaptation of composition and biophysical properties of phospholipids to temperature by the Crustacean, Gammarus spp. AB - The compositions of lipid classes as well as the molecular species composition of subclasses (diacyl, alkylacyl, and alkenylacyl forms) of choline and ethanolamine phosphoglycerides in marine amphipod crustaceans, Gammarus spp., collected in the Baltic Sea at 8 and 15 degrees C, were studied in relation to environmental temperature. The structural order of phospholipid multibilayers was also determined. Environmental temperature had little effect on fatty acid composition. The level of some polyunsaturated fatty acids, such as 20:4, even increased in choline and ethanolamine phosphoglycerides at 15 degrees C. Ethanolamine phosphoglycerides were rich in alkenylacyl forms, especially in crustaceans collected at 15 degrees C. The accumulation of sn-1 monoenic, sn-2 polyenic diacyl, alkyl, and alkenylacyl phosphatidylethanolamines and diacyl phosphatidylcholines was observed at 8 degrees C. The phospholipid vesicles of crustaceans collected at 8 degrees C were more disordered than expected compared to those obtained from animals collected at 15 degrees C. It was concluded that, in addition to variations in the levels of sn-1 monoenic and sn-2 polyenic phospholipid molecular species with temperature, ethanolamine plasmalogens may play a role in controlling membrane biophysical properties in marine amphipod crustaceans. PMID- 11104016 TI - Cold acclimation or grapeseed oil feeding affects phospholipid composition and mitochondrial function in duckling skeletal muscle. AB - The phospholipid fatty acid (FA) composition and functional properties of skeletal muscle and liver mitochondria were examined in cold-acclimated (CA, 4 degrees C) ducklings. Phospholipid FA of isolated muscle mitochondria from CA birds were longer and more unsaturated than those from thermoneutral (TN, 25 degrees C) reared ducklings. The rise in long-chain and polyunsaturated FA (PUFA, mainly 20:4n-6) was associated with a higher State 4 respiration rate and a lower respiratory control ratio (RCR). Hepatic mitochondria, by contrast, were much less affected by cold acclimation. The cold-induced changes in phospholipid FA profile and functional properties of muscle mitochondria were reproduced by giving TN ducklings a diet enriched in grapeseed oil (GO, rich in n-6 FA), suggesting a causal relationship between the membrane structure and mitochondrial functional parameters. However, hepatic mitochondria from ducklings fed the GO diet also showed an enrichment in long-chain PUFA but opposite changes in their biochemical characteristics (lower State 4, higher RCR). It is suggested that the differential modulation of mitochondrial functional properties by membrane lipid composition between skeletal muscle and liver may depend on muscle-specific factors possibly interacting with long-chain PUFA and affecting the proton leakiness of mitochondrial membranes. PMID- 11104017 TI - Effect of n-3 fatty acid deficiency on fatty acid composition and metabolism of aminophospholipids in rat brain synaptosomes. AB - Docosahexaenoic acid (DHA, 22:6n-3) is one of the major polyunsaturated fatty acids esterified predominantly in aminophospholipids such as ethanolamine glycerophospholipid (EtnGpl) and serine glycerophospholipid (SerGpl) in the brain. Synaptosomes prepared from rats fed an n-3 fatty acid-deficient safflower oil (Saf) diet had significantly decreased 22:6n-3 content with a compensatory increased 22:5n-6 content when compared with rats fed an n-3 fatty acid sufficient perilla oil (Per) diet. When the Saf group was shifted to a diet supplemented with safflower oil plus 22:6n-3 (Saf + DHA) after weaning, 22:6n-3 content was found to be restored to the level of the Per group. The uptake of [3H]ethanolamine and its conversion to [3H]EtnGpl did not differ significantly among the three dietary groups, whereas the formation of [3H]lysoEtnGpl from [3H]ethanolamine was significantly lower in the Saf group than in the other groups. The uptake of [3H]serine, its incorporation into [3H]SerGpl, and the conversion into [3H]EtnGpl by decarboxylation of [3H]SerGpl did not differ among the three dietary groups. The observed decrease in lysoEtnGpl formation associated with a reduction of 22:6n-3 content in rat brain synaptosomes by n-3 fatty acid deprivation may provide a clue to reveal biochemical bases for the dietary fatty acids-behavior link. PMID- 11104018 TI - Further characterization of rat dihydroceramide desaturase: tissue distribution, subcellular localization, and substrate specificity. AB - The introduction of the double bond in the sphingoid backbone of sphingolipids occurs at the level of dihydroceramide via an NADPH-dependent desaturase, as discovered in permeabilized rat hepatocytes. In the rat, the enzyme activity, which has now been further characterized, appeared to be mostly enriched in liver and Harderian gland. By means of subcellular fractionation of rat liver homogenates and density gradient separation of microsomal fractions, the desaturase was localized to the endoplasmic reticulum. Various detergents were inhibitory to the enzyme, and maximal activities were obtained in the presence of NADPH and when the substrate was complexed to albumin. In the presence of albumin, the chain length of the fatty acid of the truncated dihydroceramides hardly affected the activity. Finally, in view of a likely evolutionary relationship between desaturases and hydroxylases, the formation of hydroxylated intermediates was analyzed. No evidence for their presence was found under our assay conditions. PMID- 11104020 TI - Novel lipidic enaminones from a C18 keto-allenic ester. AB - Primary amines (ammonia, methyl, propyl, octyl, octadecyl, phenyl, benzyl, phenethyl) including methyl esters of amino acids (glycine, DL-alanine, L-valine, L-leucine, L-tyrosine, and L-methionine), and secondary amines (dimethyl, diethyl, dipropyl, diisopropyl, dioctyl, and diphenyl) attack regiospecifically the central carbon atom of the allene system of methyl 12-keto-9,10 octadecadienoate (1) to give the corresponding lipidic enaminone derivatives (2 21) with an average yield of 77%. The E- and Z-configuration of the enaminone system of these novel lipid derivatives was confirmed by infrared and nuclear magnetic resonance spectroscopic techniques. Primary amines furnished Z enaminones, while secondary amines gave E-enaminones. PMID- 11104019 TI - Identification of the pathway of alpha-oxidation of cerebronic acid in peroxisomes. AB - Cerebronic acid (2-hydroxytetracosanoic acid), an alpha-hydroxy very long-chain fatty acid (VLCFA) and a component of cerebrosides and sulfatides, is unique to nervous tissues. Studies were carried out to identify the pathway and the subcellular site involved in the oxidation of cerebronic acid. The results from these studies revealed that cerebronic acid was catabolized by alpha-oxidation to CO2 and tricosanoic acid (23:0). Studies with subcellular fractions indicated that cerebronic acid was alpha-oxidized in fractions having particulate bound catalase and enzyme systems for the beta-oxidation of VLCFA (e.g., lignoceric acid), suggesting peroxisomes as the subcellular organelle responsible for alpha oxidation of cerebronic acid. Etomoxir, an inhibitor of mitochondrial fatty acid oxidation, had no effect on cerebronic acid alpha-oxidation. Further, cerebronic acid oxidation was found to be dependent on the presence of NAD+ but not FAD, NADPH, ATP, Mg2+, or CoASH. Intraorganellar localization studies indicated that the enzyme system for the alpha-oxidation of cerebronic acid was associated with the peroxisomal limiting membranes. Studies on cultured fibroblasts from normal subjects and patients with peroxisomal disorders indicated an impairment of alpha oxidation of cerebronic acid in cell lines that lack peroxisomes [e.g., Zellweger syndrome (ZS)]. On the other hand, alpha-oxidation of cerebronic acid was found to be normal in cell lines from X-linked adrenoleukodystrophy, adult Refsum disease, and rhizomelic chondrodysplasia punctata. Our results clearly demonstrate that alpha-oxidation of alpha-hydroxy VLCFA (cerebronic acid) is a peroxisomal function and that this oxidation is impaired in ZS. Furthermore, this alpha-oxidation enzyme system is distinct from the one for the alpha-oxidation of beta-carbon branched-chain fatty acids (e.g., phytanic acid). PMID- 11104021 TI - Comparison of silver-ion high-performance liquid chromatographic quantification of free and methylated conjugated linoleic acids. AB - Silver-ion high-performance liquid chromatography was used to fractionate a mixture of conjugated linoleic acid (CLA) isomers (as the free fatty acids, CLAFFA) in commercial CLA mixtures and biological samples. Due to the unchanged retention mechanism, it was assumed that the elution order of the isomers remained the same as that of methyl esters separated on the same column. The most abundant isomers, cis/trans 10, 12-18:2 and cis/trans 9,11-18:2, were separated better as free acids on a single column than in the methyl ester form. Quantification of the CLA standard was used as the reference profile to evaluate different methylation methods commonly used to prepare CLA methyl esters for quantitation. Acid-and base-catalyzed derivatization methods resulted in CLA intraisomerization and losses in total conjugated dienes content. Acid (HCl and BF3) methylations significantly elevated the level of trans,trans isomers and significantly reduced the cis/trans isomers. Base methylation, tetramethylguanidine/methanol, resulted in loss of trans,trans isomers, and a substantial loss of total underivatized conjugated dienes. Other catalysts such as the trimethylsilyldiazomethane produced additional peaks of unidentified artifacts. The analysis of CLAFFA appears to provide more accurate quantification of CLA isomers in commercial and biological samples. PMID- 11104022 TI - Characterization of caldarchaetidylglycerol analogs, dialkyl-type and trialkyl type, from Thermoplasma acidophilum. AB - The structures of three kinds of phospholipids (PL-X, PL-Y, and PL-T) isolated from Thermoplasma acidophilum have been characterized. The core lipid of PL-Y was caldarchaeol, and that of PL-X was archaeol. The composition of the hydrocarbon chains of the PL-T core lipid was C20 phytane and C40 isoprenoid in a molar ratio of 2 to 1. The major molecular species of the C40 isoprenoid was acyclic without the cyclopentane ring. These three kinds of intact phospholipids commonly had glycerophosphate residues as polar head groups. The structure of PL-T was characterized as trialkyl-type caldarchaetidylglycerol, PL-Y as caldarchaetidylglycerol, and PL-X as archaetidylglycerol. PMID- 11104023 TI - The effect of maternal diets on the mean melting points of human milk fatty acid. PMID- 11104024 TI - E-cadherin mutation-based genetic counseling and hereditary diffuse gastric carcinoma. AB - The E-cadherin mutation has been identified in a subset of families with multiple cases of diffuse gastric carcinoma. However, the true penetrance of this mutation and its association with other carcinomas in such families remains elusive. We aim to show the importance of DNA-based genetic counseling in hereditary diffuse gastric carcinoma. The proband was self-referred after three of his siblings died of diffuse gastric carcinoma. Medical and pathology records confirmed diagnoses. The family was educated about diffuse gastric carcinoma. Analysis for the 70G-->T mutation was performed by sequencing genomic DNA from lymphocyte pellets. DNA results and genetic counseling were provided individually to those tested. Twenty four family members were tested for the E-cadherin mutation. Nine were found to be positive and 15 were negative. Three who tested positive and were affected are now deceased. None of the 19 patients counseled wanted results sent to their physicians once they recognized the potential for insurance discrimination. None had undergone endoscopic ultrasound. Three who were positive for the E-cadherin mutation expressed strong interest in prophylactic gastrectomy. The E-cadherin mutation strongly predicts susceptibility to diffuse gastric carcinoma. Emotional stress in at-risk patients, the limited knowledge of the mutation's penetrance, and limitations of available screening mandate patient-centered genetic counseling. PMID- 11104025 TI - Comparison of comparative genomic hybridization and interphase fluorescence in situ hybridization in ovarian carcinomas: possibilities and limitations of both techniques. AB - Comparative genomic hybridization (CGH) is a valuable technique for cytogenetic analysis of solid tumors. To evaluate the reliability of CGH, we examined DNA of 10 ovarian carcinomas after CGH analysis with single- and double-locus fluorescence in situ hybridization (FISH). The FISH experiments, involving 5 chromosomes (chromosomes 3, 6, 8, 12, and 18) with different FISH probes, confirmed the CGH results in 66.2% of cases (92 of 139 investigated loci). In 4 patients, inconsistent results (41 loci) were related to polyploidy, because CGH cannot detect polyploid karyotypes. The remaining 6 discordant loci can be referred to limitations in both techniques. Re-evaluation of FISH and CGH results by one other is therefore recommended to overcome these technical artifacts. Nevertheless, CGH is of potential value in characterizing chromosomal alterations and might help in generating tumor-specific sets of FISH probes to obtain genetic information of prognostic value within a few days. PMID- 11104026 TI - Identification of chromosomes 3, 6, and 8 aberrations in uveal melanoma by microsatellite analysis in comparison to comparative genomic hybridization. AB - In uveal melanoma, monosomy 3 is strongly associated with metastic disease and poor prognosis. Cytogenetic analysis and comparative genomic hybridization (CGH) have been used to identify chromosomal aberrations in uveal melanoma. As these methods are costly and time consuming in routine diagnostic settings, we evaluated whether tumors with monosomy 3 can be reliably identified by microsatellite analysis (MSA). In addition, we also tested if aberrations of chromosomes 6 and 8, which have also been associated with the course of the disease, can be detected by MSA. We established a protocol for MSA of 23 markers, 3-4 on each arm of chromosomes 3, 6, and 8. Twenty tumors were analyzed by CGH and MSA, and 10 tumors were analyzed by MSA only. For chromosome 3, the results of CGH and MSA were concordant, thus indicating that the dosage of this chromosome can reliably be determined by MSA. However, MSA failed to detect copy number gains at 6p in some tumors. Moreover, despite quantitative evaluation of allele ratios, it was not possible to discern 8p losses and gains reliably. We thus conclude that while MSA can be used to determine monosomy 3 in uveal melanoma, careful interpretation of results for chromosomes 6 and 8 is recommended. PMID- 11104027 TI - Comparative genomic hybridization detects losses of chromosomes 22 and 16 as the most common recurrent genetic alterations in primary ependymomas. AB - In this study, we used comparative genomic hybridization to provide an overview of chromosomal imbalances in a series of 20 adult and 8 childhood ependymomas. All tumors displayed multiple genomic imbalances. Loss of genetic material was observed in chromosomes 22q (71%), 16 (57%), 17 (46%), 6 (39%), 19q (32%), 20q (32%), and 1p (29%), with the overlapped deletion regions determined at 16p13.1 13.3, 16q22-q24, 19q13.1-13.4, 20q13.1-13.2 and 1p36.1-36.3. Gain of DNA was commonly detected on chromosomes 5q (46%), 12q (39%), 7q (36%), 9q (36%), and 4q (32%), with overlapped regions of gain mapped to 5q21-22, 12q15-24.1, 7q11.2 31.2, 9q12-32, and 4q23-28, respectively. These findings suggest a greater degree of genomic imbalance in ependymomas than has been recognized previously and highlight chromosomal loci likely to contain oncogenes or tumor suppressor genes that may contribute to the molecular pathogenesis of this tumor. Our study also confirmed previous findings on frequent losses of 17 and 22q in ependymomas and further identified chromosome 16 loss as a common recurrent genetic aberration in ependymomas. PMID- 11104028 TI - Acute myeloid leukemia associated with hemophagocytic syndrome and t(4;7)(q21;q36). AB - Hemophagocytic syndrome (HS) is a histiocytic reactive process often associated with infections and/or malignancies. Clonal karyotypic abnormalities have been the hallmark of several hematological malignancies and have been shown to be of clinical significance in terms of both diagnosis and prognosis. While there are limited reports of both clonal and nonclonal abnormalities in HS, their clinical significance has not been established. Detection of such clonal abnormalities, as seen in some cases of HS, may indicate the presence of an occult malignant process, even when there is no microscopic evidence of a hematological malignancy. We report a case of HS in a child with clonal t(4;7)(q21;q36) which later progressed to acute myeloid leukemia (AML) with further clonal evolution. Our case strengthens the argument that cytogenetic studies in HS may be important in identifying the underlying occult malignant process. PMID- 11104030 TI - Deletion of Xq23 is a recurrent karyotypic abnormality in acute myeloid leukemia. AB - Deletion of chromosome Xq23 has been reported in a number of solid tumors, including soft tissue sarcoma, malignant melanoma, astrocytoma, and adenocarcinoma. The deleted Xq often occurs in a setting of very complex karyotypic changes. A similar abnormality has also been described in rare cases of acute myeloid leukemia (AML) but in no other hematologic malignancies. In this study, we report the occurrence of del(X)(q23) in two cases of AML. PMID- 11104029 TI - Recurrent chromosome aberrations in fibrous dysplasia of the bone: a report of the CHAMP study group. CHromosomes And MorPhology. AB - The nosologic status of fibrous dysplasia (FD), a well-known and relatively common bone lesion, is controversial. Information collected by the CHromosomes And MorPhology (CHAMP) study group on published and unpublished cases of fibrous dysplasia shows the presence of clonal chromosome changes in at least a proportion of these lesions. The chromosome aberrations found in FD lesions have been quite variable and have included both structural and numerical changes. Two of the three cases investigated at the study group had trisomy 2 as the sole acquired anomaly. Combined with previously published data, +2 and rearrangements involving chromosome band 12p13 have each been detected in 3 of 8 cases with abnormal karyotype of 11 in which chromosomal analysis has been performed, suggesting that FD is a neoplastic lesion rather than a "dysplastic" process, as has been generally believed and as implied by its very name. PMID- 11104031 TI - HSNF5/INI1 gene mutations in lymphoid malignancy. AB - hSNF5/INI1 is one of the components of the SWI/SNF multiprotein complex that is necessary for the transcriptional activation of several genes and functions by altering chromatin structure. This gene has been thought to be one of the tumor suppressor genes (TSGs) because deletions or mutations were reported in malignant rhabdoid tumors and atypical teratoid and rhabdoid tumors. To evaluate the hSNF5/INI1 gene as a TSG in lymphoid malignancies, we performed a mutational analysis in 23 patients with non-Hodgkin lymphoma (NHL), 24 with acute lymphoblastic leukemia (ALL), 24 with multiple myeloma (MM), 24 with adult T-cell lymphoma/leukemia (ATLL), and 19 with lymphoid cell lines, by polymerase chain reaction-single-strand conformational polymorphism (PCR-SSCP) analysis. Nonsense and missense mutations were found in 1 NHL case and 2 cell lines. Mutations from this NHL case proved to be somatic in origin. These data indicated that alterations in the hSNF5/INI1 gene might be involved in the pathogenesis of lymphoid malignancies. PMID- 11104033 TI - Loss of heterozygosity at chromosomes 3, 6, 8, 11, 16, and 17 in ovarian cancer: correlation to clinicopathological variables. AB - Tumor specimens from 78 epithelial ovarian cancer patients were examined for loss of heterozygosity (LOH) at 11 microsatellite markers at chromosomes 3p14.2, 6q27, 8p12, 11p15.5, 11q23.1-q24, 16q24.3, and 17p13.1, to evaluate the involvement, possible clustering, and prognostic significance of these lesions in the progression of the disease. The LOH analysis was performed on polymerase chain reaction (PCR)-amplified DNA from sections of paraffin-embedded tumor and normal tissue pairs. In addition to primary tumors, specimens of metastatic tissues were studied from 19 patients. In the combined results from primary and metastatic tumors, LOH frequencies varied between 31% (6q27) and 69% (17p13.1). Only LOH at chromosomal regions 3p14.2 (D3S1300), 11p15.5 (D11S1318), 11q23.3-q24 (D11S1340 and D11S912), 16q24.3 (D16S476 and D16S3028), and 17p13.1 (D17S938) was associated with an adverse disease course. Our results indicate that LOH at 17p13.1 occurs independently from the other chromosomal sites studied, and is an early event in ovarian tumorigenesis. The LOH at 16q24.3, 11q23.3/q24, and 11p15.5 seems to occur later. The LOH at 11p15.5 and 11q23.3 was associated with reduced cancer-specific survival time; therefore, the studied markers could be located close to genes with influence on patient survival. Of the studied chromosomal regions, the most important tumor suppressor genes involved in the evolution of ovarian cancer appear to be located on chromosomes 11, 16, and 17. The genetic heterogeneity observed in primary and metastatic specimens demonstrates that there are multiple pathways involved in the progression of ovarian cancer. PMID- 11104032 TI - Microdissection, DOP-PCR, and comparative genomic hybridization of paraffin embedded familial prostate cancers. AB - There is a clear genetic component to prostate cancer susceptibility. Regions reported to be linked to prostate cancer include 1q24-25 (HPC-1), 1q42.2-43, and Xq27-28. There is limited genetic information on familial prostate tumors. We used the Utah Population Database to identify familial prostate cancer cases and selected 35 cases from high-risk families. Tissue blocks containing discernable tumor were available from 19 cases; 13 of these yielded adequate specimens for analysis. Six cases came from families with linkage to HPC-1, 3 were known to have linkage to Xq27-28, and 4 had no linkage to a known locus; 7 cases were analyzed from patients who showed no known linkage (sporadic tumors) as controls. These paraffin-embedded tumors were laser microdissected, degenerate oligonucleotide (DOP)-amplified, and labeled for fluorescence detection by comparative genomic hybridization (CGH). Loss of 7q, 10q, and 16q and gain of 8q were common abnormalities present in both familial and sporadic tumors. Distinctive abnormalities included loss of 3p12-3p22 in 3 of 6 HPC-7-linked cases and in 2 of 3 X-linked cases and gain of 6q11-6q21 in 2 each of HPC-1 and X linked tumors. In conclusion, laser microdissection, DOP-PCR, and CGH is a feasible method for analysis of paraffin-embedded prostate tumors. This study provides preliminary data suggesting that familial prostate cancer harbors some unique genetic changes when compared with sporadic prostate tumors. PMID- 11104034 TI - DNA replication error is frequent in ovarian granulosa cell tumors. AB - DNA replication errors (RER) have been detected in epithelial ovarian cancers, as well as in other human tumor types. These observations suggest that this genetic defect is present in ovarian granulosa cell tumors, and that a DNA mismatch repair deficiency may be involved in their development and/or progression. We therefore assayed tissue samples from 29 patients with granulosa cell tumors for RER, using polymerase chain reaction (PCR) and 5 microsatellite markers. The RER were observed at greater than or equal to 1 loci in 15 (58%) of 26 informative cases. The incidence of RER was unrelated to the patient's age or the histologic subtype or clinical stage of the tumors. The RER, however, were observed in 57% (8/14) of the informative patients with stage IA disease. These findings suggest that a DNA mismatch repair deficiency may contribute to the pathogenesis of ovarian granulosa cell tumors, and that this deficiency may be an early event in their development and/or progression. PMID- 11104035 TI - Acute megakaryoblastic leukemia in an infant with a novel t(1;9)(p32;q34). AB - We report a case of a 14-month-old girl with acute megakaryoblastic leukemia (AMKL). May-Giemsa staining of the bone marrow cells revealed the proliferation of two distinct types of blasts. One type of blasts had cytoplasmic blebs, and the other showed a lymphoblastic morphology without blebs. Both types of blasts were negative for peroxidase and esterase reactions. Electron microscopic platelet peroxidase (PPO) reaction also revealed the presence of two types of blasts. One had irregular-shaped nuclei and positive PPO reaction in the nuclear envelope and rough endoplasmic reticulum but not in the Golgi apparatus. These types of blasts were considered to be megakaryoblasts. The other had an immature phenotype with round nuclei and positive PPO reaction in the nuclear envelope, rough endoplasmic reticulum, and the Golgi apparatus. The origin of this type of blasts could not be defined by their morphology. Surface marker analysis indicated that most of the leukemic cells expressed platelet markers, gpIIb, gpIIb/IIIa, gpIX, and gpIbalpha. Karyotypic analysis of the bone marrow cells of this unique subset of AMKL demonstrated a novel translocation, t(1;9)(p32;q34). PMID- 11104037 TI - Treating the unowned animal: who should pay? PMID- 11104036 TI - Cytogenetic analysis in three cerebral subependymomas: further evidence for a hamartomatous nature? PMID- 11104038 TI - Illthrift and scouring frequently reported in lambs in September. PMID- 11104039 TI - Investigations towards an efficacious and safe strangles vaccine: submucosal vaccination with a live attenuated Streptococcus equi. AB - As part of a search for a safe and efficacious strangles vaccine, several different vaccines and different vaccination routes were tested in foals. The degree of protection was evaluated after an intranasal challenge with virulent Streptococcus equi by clinical, postmortem and bacteriological examinations. Inactivated vaccines containing either native purified M-protein (500 microg per dose) or whole S equi cells (10(10) cells per dose) administered at least twice intramuscularly at intervals of four weeks, did not protect against challenge. Different live attenuated S equi mutants administered at least twice at intervals of four weeks by the intranasal route were either safe but not protective or caused strangles. In contrast, a live attenuated deletion mutant administered intramuscularly, induced complete protection but also induced unacceptable local reactions at the site of vaccination. Submucosal vaccination in the inner side of the upper lip with the live attenuated mutant at > or =10(8) colony-forming units per dose, appeared to be safe and efficacious in foals as young as four months of age. The submucosal vaccinations caused small transient swellings that resolved completely within two weeks, and postmortem no vaccine remnants or other abnormalities were found at the site of vaccination. PMID- 11104040 TI - Efficacy of non-acaricidal containing otic preparations in the treatment of otoacariasis in dogs and cats. AB - Eighty-nine cats and 38 dogs naturally infested with the ear mite Otodectes cynotis were randomly allocated into two treatment groups. One group was treated with a product containing miconazole nitrate, polymyxin B sulphate and prednisolone acetate, the other with a combination of diethanolamine fusidate, framycetin sulphate, nystatin and prednisolone. The treatment (five drops in each ear) was applied twice daily for 14 days, and its efficacy was evaluated on days 7, 14 and 21 on the basis of an otoscopic examination of the external ear canal, a microscopical examination of scrapings for the presence of ear mites and clinical signs of pruritus, pain, erythema and/or exudate. Both treatments were highly effective, and there were no significant differences between the two products, either in efficacy or in the clinical improvements observed. Apart from an allergic reaction in one cat treated with the second product, no adverse effects were observed. PMID- 11104041 TI - Use of moxidectin treatment in the investigation of abomasal nematodiasis in wild reindeer (Rangifer tarandus platyrhynchus). AB - An experiment was conducted to evaluate moxidectin as a tool for understanding the impact of parasitism on wild Svalbard reindeer (Rangifer tarandus platyrhynchus). Adult females were injected subcutaneously with moxidectin at a dose rate of 0-4 mg/kg bodyweight, and groups of animals were culled within its expected period of efficacy (around 14 days) or around 12 or 24 weeks after treatment. Moxidectin was effective in eliminating the reindeers' abomasal worm burdens, and although they became reinfected, worm burdens were significantly lower in the treated animals compared to the untreated controls for up to 24 weeks after treatment. Nematode eggs did not reappear in faeces until five weeks after treatment, a similar period to that claimed by the manufacturer for sheep and cattle. Animals culled 12 and 24 weeks after treatment had been reinfected and harboured a wide range of abomasal worm burdens which contributed to the understanding of the seasonal variation in the relationship between faecal egg count and worm burden. PMID- 11104042 TI - Repeatability of a lameness scoring system for finishing pigs. PMID- 11104043 TI - Quinolone resistance in Escherichia coli strains isolated from diarrhoeic lambs in Spain. PMID- 11104045 TI - Effect of a corpus luteum on the recovery and developmental potential of buffalo oocytes. PMID- 11104044 TI - Intramammary Aspergillus fumigatus infection in dairy ewes associated with antibiotic dry therapy. PMID- 11104046 TI - Data protection. PMID- 11104047 TI - Farmers' views on veterinary services. PMID- 11104048 TI - Leptospirosis control in UK pigs. PMID- 11104049 TI - Early neutering of dogs. PMID- 11104050 TI - The role of muscle activity and mental load in the development of pain and degenerative processes at the muscle cell level during computer work. PMID- 11104051 TI - Changes in muscle afferents, motoneurons and motor drive during muscle fatigue. AB - Fatigue is a reduction of maximal muscle force or power that occurs with exercise. It is accompanied by changes at multiple levels in the motor pathway and also by changes in the discharge patterns of muscle afferents. Changes in afferent firing can lead to altered perceptions and can also act on the efferent pathway. Changes in the motor pathway include slowing of motor unit firing rates during sustained maximal voluntary contractions (MVCs). Muscle responses to stimulation at different levels of the motor pathway also change. Transcranial magnetic stimulation of the motor cortex and stimulation of descending tracts in the spinal cord in human subjects show an increase in the response of the cortex and a decrease in response of the motoneuron pool during sustained MVCs. In addition, the silent period following magnetic stimulation is prolonged. During relaxation after fatiguing exercise, muscle responses to stimulation of the motor cortex are initially facilitated and are then depressed for many minutes, whereas responses to descending tract stimulation are initially depressed but recover over about 2 min. Although some of the loss of force of fatigue does occur through inadequate drive to the muscle, it is not clear which, if any, of the changes described in the cortex or the motoneurons are responsible for loss of maximal voluntary force and thus contribute to fatigue. Changes may be associated with muscle fatigue without causing it. PMID- 11104052 TI - Control of the wrist joint in humans. AB - As one considers changes in motor activity from lower mammals to higher primates, one of the major changes one observes lies in the cortical control of forelimb muscles. There has been a shift from disynaptic control of spinal motoneurons in, for example, the cat, to a greater and greater percentage of monosynaptic control of hand and forelimb motoneurons in the primate. In spite of the species and evolutionary changes in the synaptic connections of the corticospinal tract, it appears that the interneurons identified in the cat are retained in the monkey and human. These interneurons, under the influence of descending pathways, modulate the output of motoneuron pools. Perhaps the control of these interneurons has also changed towards finer control of movement, as has been suggested by recent studies in the monkey. Whether in cat or human, the recruitment pattern for motor units is the same; the change from disynaptic to monosynaptic connections has not changed the recruitment pattern of muscles. Differences in the recruitment patterns of muscles may lie in the finer control of inputs to motoneurons in the primate. This review seeks to integrate the current knowledge of the mechanisms involved in the motor control of the wrist joint and especially in the recruitment patterns of the muscles. These motor control mechanisms include the biomechanics of the wrist joint, recruitment patterns of wrist muscles, interneurons and spinal cord circuits in the cervical regions mediating the output of spinal motoneurons, and the supraspinal control of these muscles. PMID- 11104053 TI - Adaptations in motor unit discharge activity with force control training in young and older human adults. AB - Six young (aged 18-22 years) and six older (aged 66-76 years) healthy humans participated in a visually guided isometric force modulation training program designed to improve accurate control of force during ankle dorsiflexion. Isometric force and the discharge activity of motor units (MU) supplying the tibialis anterior muscle were sampled concurrently at the beginning of the study, following 2 weeks of force modulation training and again after a 4 week retention period which followed immediately. The initial maximal voluntary force (MVC) and MU discharge rates were similar between young and older adults at 10-60% MVC while MU discharge rates during maximal effort were significantly reduced in older adults. Following the 2 weeks of force modulation training, both young and older adults demonstrated significant improvements in force accuracy (44% young, 48% older) and significantly reduced MU discharge rates at 30%, 40%, and 60% MVC. Young adults also demonstrated increased MVC force (11%), while older adults demonstrated significantly increased (30%) maximal MU discharge rate. Thus, following 2 weeks of force modulation training, young and older individuals demonstrated similar MU discharge rates at all force levels. The MU discharge rate adaptations were retained after the 4 week retention period. In young adults, improved force accuracy and increased MVC force were accompanied by significantly reduced MU recruitment thresholds. In the older subjects, improved force accuracy was accompanied by an increase in the difference between the recruitment-derecruitment force threshold and significantly reduced antagonist co contraction. Age-related alterations in force regulation and MU discharge activity cannot be explained solely on the basis of contractile changes in senescent muscle. Rather, reliance on compensatory neuromuscular changes including antagonist muscle co-contraction is suggested. PMID- 11104054 TI - Non-invasive approach of motor unit recording during muscle contractions in humans. AB - Information about the structural and functional characteristics of the motor unit (MU) is highly relevant for the diagnosis of neuromuscular disorders. Electromyography (EMG) is a suitable method for obtaining the information needed. The problem is the separation of the activity of one MU from others which are simultaneously active. Such investigations of single MU activity have commonly used invasive methods, e.g. employing a needle or a wire. Conventional surface EMG methods have limited resolution and detect, at high contraction levels, multiple MU superimposed one on the other. The separation of the activity of a single MU can be achieved in a non-invasive way when highly specialised acquisition techniques are used. One approach, called high spatial resolution EMG (HSR-EMG), is based on the use of multi-electrode arrays in combination with a two-dimensional Laplace filter. The HSR-EMG permits the completely non-invasive detection of single MU activity even during maximal voluntary contractions. First applications have shown that the method provides a deeper insight into the functional and structural characteristics of the MU. In this paper the application of HSR-EMG to the diagnosis of neuromuscular disorders will be presented, and the latest results will be given of its application in the evaluation of treatment of patients with plexus lesion. PMID- 11104055 TI - Human muscle activity related to non-biomechanical factors in the workplace. AB - This paper presents current knowledge on low-level, long-lasting work-related muscle activity, focusing on the shoulder and the upper part of the trapezius muscle, and on mental, rather than biomechanical reasons for the muscle activation. The paper identifies three sources of vocational muscle activity: the biomechanical need for force production in order to perform movements or maintain postures against the force of gravity, the biomechanical need to stabilise body parts as a reference for performing movements and securing a stable visual field, and finally, muscle activity without obvious biomechanical purposes. This last category has been labelled non-biomechanical muscle activity in this review. Non biomechanical muscle activity is related to the mental load, the emotional load and the individual characteristics of the subject, and is identified as having a low-level and a low second-to-second variability, resembling a static muscle contraction. Recent research has indicated that the size principle for motor unit recruitment order puts a strain on a limited number of low-threshold motor units which might be heavily taxed despite the overall low level of this muscle activity. However, the paper also cites a recent report showing that motor unit substitution may occur in prolonged low-level muscle activation (longer than a few minutes). Evaluations of muscle load at work usually omit the possibility of extra muscle activation due to nonbiomechanical factors, and thus may often give estimates of the muscle load that are too low, or misinterpret nonbiomechanical muscle activity as biomechanical muscle load. PMID- 11104056 TI - Human muscle fibre abnormalities related to occupational load. AB - Ten biopsy studies addressing structural and histochemical muscle fibre abnormalities related to occupational work and myalgia have been reviewed. Eight of these were focused on the upper trapezius muscle while the two others were made on the first dorsal interosseous muscle and the extensor carpi radialis brevis muscle. Standard histological methods have mainly been used with some variations. All except two studies were on women. To achieve desirable contrasts in effect, subject groups usually were individuals with myalgia exposed to static and/or highly repetitive work and healthy non-exposed subjects. In three studies work-exposed subjects without myalgia were included, in two others two groups with myalgia were formed based on the degree of disorder. Major findings in the subjects with myalgia or exposed to work load were increased fibre cross sectional areas, various indications of mitochondrial disturbances in type I fibres and reduced capillarisation per normal fibre cross-sectional area, the last only in women. The relationship between these findings and pain perception are unclear and frequently similar findings are made also in work-exposed pain free subjects. Some signs of mitochondrial disturbances in the trapezius muscle were also found in reference groups. However, there are some indications of a relationship between degree of abnormality and degree of pain. There are also indications of differences between the type of finding in the trapezius muscle and in the hand/forearm muscles. PMID- 11104057 TI - Functional significance of Ca2+ in long-lasting fatigue of skeletal muscle. AB - Repeated activation of skeletal muscle causes fatigue, which involves a reduced ability to produce force and slowed contraction regarding both the speed of shortening and relaxation. One important component in skeletal muscle fatigue is a reduced sarcoplasmic reticulum (SR) Ca2+ release. In the present review we will describe different types of fatigue-induced inhibition of SR Ca2+ release. We will focus on a type of long-lasting failure of SR Ca2+ release which is called low-frequency fatigue, because this type of fatigue may be involved in the muscle dysfunction and chronic pain experienced by computer workers. Paradoxically it appears that the Ca2+ released from the SR, which is required for contraction, may actually be responsible for the failure of SR Ca2+ release during low frequency fatigue. We will also discuss the relationship between gross morphological changes in muscle fibres and long-lasting failure of SR Ca2+ release. Finally, a model linking muscle cell dysfunction and muscle pain is proposed. PMID- 11104058 TI - Ca2+ accumulation and cell damage in skeletal muscle during low frequency stimulation. AB - Electrical stimulation has been shown to produce a marked increase in Ca2+ influx and Ca2+ content in rat skeletal muscle. Long-term low-frequency stimulation (1 Hz, 240 min) increased 45Ca uptake by 30% and 154% in soleus and extensor digitorum longus muscles, respectively. Studies using Ca2+-fluorescent dyes have shown that intracellular concentrations of free Ca2+ are increased up to threefold during long-term low-frequency stimulation, suggesting that muscle cells have difficulties in handling the Ca2+ taken up during stimulation. Furthermore, long-term low-frequency stimulation induces leakage of the intracellular enzyme lactate dehydrogenase from the muscles. This leakage may reflect degradation of membrane proteins by the Ca2+-activated neutral protease calpain. This, in turn, leads to further influx of Ca2+ and further acceleration of protein breakdown. Membrane leakages are likely to result in sensations of pain in the damaged muscle. It is suggested that Ca2+ plays a central role in the development of muscle fibre injury during prolonged muscle activity of workers using a computer mouse. PMID- 11104060 TI - Motor unit recruitment and rate coding in response to fatiguing shoulder abductions and subsequent recovery. AB - The purpose of the present study was to investigate motor unit (MU) recruitment and firing rate, and the MU action potential (MUAP) characteristics of the human supraspinatus muscle during prolonged static contraction and subsequent recovery. Eight female subjects sustained a 30 degrees shoulder abduction, requiring 11-12% of maximal voluntary contraction (MVC), for 30 min. At 10 and 30 min into the recovery period, the shoulder abduction was repeated for 1 min. The rating of perceived exertion for the shoulder region increased to "close to exhaustion" during the prolonged contraction, and the surface electromyography (EMG) recorded from the deltoid and trapezius muscles showed signs of local muscle fatigue. From the supraspinatus muscle, a total of 23,830 MU firings from 265 MUs were identified using needle electrodes. Of the identified MUs, 95% were continuously active during the 8-s recordings, indicating a low degree of MU rotation. The mean (range) MU firing rate was 11.2 (5.7-14.5) Hz, indicating the relative force contribution of individual MUs to be larger than the overall mean shoulder muscle load. The average MU firing rate remained stable throughout the prolonged abduction, although firing rate variability increased in response to fatigue. The average concentric MUAP amplitude increased by 38% from the beginning (0-6 min) to the end (24-29 min) of the contraction period, indicating recruitment of larger MUs in response to fatigue. In contrast, after 10 min of recovery the average MU amplitude was smaller than seen initially in the prolonged contraction, but not different after 30 min, while the MU firing rate was higher during both tests. In conclusion, MU recruitment plays a significant role during fatigue, whereas rate coding has a major priority during recovery. Furthermore, a low degree of MU rotation in combination with a high relative load at the MU level may imply a risk of overloading certain MUs during prolonged contractions. PMID- 11104059 TI - Motor-unit activity in the trapezius muscle during rest, while inputting data, and during fast finger tapping. AB - The Cinderella hypothesis postulates the continuous activity of specific motor units during low-level muscle contraction and contradicts the concept of motor unit substitution. Constant trapezius muscle activity has been reported in typical visual-display-unit-related tasks. If it can be shown that constant muscle activity can be caused by the continuous firing of single motor units, this could explain the frequent complaints of muscular neck pain reported by computer users. The present study was undertaken to investigate motor-unit activity in the trapezius muscle during resting with closed eyes, while inputting three-digit numbers with auditory presentation at a rate of 0.5 Hz, and while tapping on a key with the right index finger at a rate of 5 Hz. Electrodes with four fine wires were inserted into the right upper trapezius muscle of six healthy subjects, and three-channel intramuscular electromyography was recorded. The decomposition programme MAPQuest, developed to analyse short-term one-channel signals, was complemented with MAPView, a programme that merges the short-term results of 10 s to a 3-min analysis. The results showed that activity in the trapezius muscle was induced in one subject while resting, in two subjects while inputting data, and in five subjects while finger tapping. Long-lasting single motor-unit firing was observed in two subjects while inputting data and in one subject while finger tapping. Whilst our findings may support the Cinderella hypothesis, the measurement periods are too short to confirm it fully, and for further discussion it is necessary to record and analyse for longer periods. PMID- 11104061 TI - The influence of experimental muscle pain on motor unit activity during low-level contraction. AB - In the present study we compared motor unit (MU) activity in a painful extensor carpi ulnaris (ECU) muscle to that of a pain-free control. According to the pain adaptation model the activity of the painful ECU muscle may be inhibited and its antagonist activity increased during wrist extension performed as a pre-defined low-force ramp. The pre-defined low force may then be maintained by increased activity in the pain-free synergist muscles such as the extensor carpi radialis (ECR) muscle. Nine females (31-47 years old) participated in the study. Maximal voluntary contraction (MVC) of the wrist extensors was performed. A catheter was inserted into the ECU muscle to allow the injection of hypertonic saline to evoke muscle pain, and a concentric needle was inserted for the recording of MU activity. Surface electromyograms were recorded from a synergist and an antagonist (ECR and flexor carpi radialis) to the painful ECU muscle. A force ramp of isometric wrist extensions up to 10% MVC, with a force increase of 1% MVC x s(-1), were performed followed by 60 s of sustained contraction at 10% MVC. The number of MUs recruited was almost identical for baseline and with pain, and no effect of experimental muscle pain was found on the properties of the MUs (amplitude, area) or their firing characteristics (mean firing rate, firing variability) during low-force ramp contraction. During the sustained 10% MVC, no effect of pain was found for concentric or surface EMG of the forearm muscles. At low force levels no pain-induced modulations were found in MU activity, when the mechanical condition was similar to that of a control situation. PMID- 11104062 TI - Co-activity of the trapezius and upper arm muscles with finger tapping at different rates and trunk postures. AB - In the context of finding a model that describes the pathophysiological mechanisms leading to muscle pain at low-intensity repetitive work, in this study we investigated whether a simplified finger motor task that requires little mental demand can cause increased muscle activity in the upper arms and neck, and examined the impact of the variation of two parameters, finger tapping rate and body posture. Using the 5th and 95th percentiles from the surface electromyogram of six muscles of the fingers, upper arm and neck, we determined the static and dynamic components of the muscle activity. Correlation methods were used to find a component in the muscle activity that originated from the rhythm of the finger tapping. Further investigations included tapping steadiness and finger force. It was found that in many, but not all subjects, low or even high activity was constantly present in the upper arm and trapezius muscles, sometimes even during relaxation. Fast tapping and a forward-leaning body posture caused considerable increases, while a slightly reclined posture helped to reduce co-activity. However, motor control patterns varied strongly between individuals. Since certain subjects showed no co-activity at all we can assume that trapezius and upper-arm activation is not necessarily required for the completion of a task similar to ours. This may explain why some VDU users develop work-related musculoskeletal disorders while others remain healthy. PMID- 11104063 TI - Surface EMG and psychophysiological stress reactions in women during repetitive work. AB - In order to understand the high prevalence of musculoskeletal disorders associated with stressful work, it is important to explore the relationship between muscle activity and psychophysiological stress responses. The present real-life study examines surface trapezius electromyographic (sEMG) activity, heart rate, blood pressure, and levels of urinary catecholamines and salivary cortisol among 31 female employees working at supermarkets, where the prevalence of neck and shoulder disorders is high (60-70%). As expected, the results show that psychophysiological arousal was high during work. Significant correlations were found between self-reports indicating negative stress (stressed, exhausted, tense) and sEMG activity during work. No significant correlations were found between self-reports of positive reactions (stimulated, concentrated, happy) and sEMG activity. No associations were found between sEMG activity and pain or between negative stress ratings and pain. Objectively measured workload and physiological stress responses did not correlate significantly with sEMG activity. Thus, our data indicate that perceived negative stress may have a specific influence on muscle activity, which may be of importance for musculoskeletal disorders in jobs with low-to-moderate physical load and negative psychosocial factors. PMID- 11104064 TI - Intramuscular pressure of the infra- and supraspinatus muscles in relation to hand load and arm posture. AB - In work engaging the upper extremities, the musculoskeletal system of the shoulder is sometimes exposed to prolonged excessive load, leading to musculoskeletal disorders of the shoulder. One way of reducing work-related shoulder disorders is to establish guidelines for working postures. The purpose of this study was to identify harmful working positions, by performing a comprehensive survey of the intramuscular pressure (IMP) in the infra- and supraspinatus muscles in relation to different arm positions and external loads. Ten healthy males participated, and the IMP in the infra- and supraspinatus muscles was studied in a total of 112 combinations of arm positions and hand loads at levels that occur frequently in industrial work. High-precision spatial recordings were accomplished with a three-dimensional motion-analysis system, and the IMP was measured using the microcapillary infusion technique. The mean IMP of the infraspinatus muscle as well as that of the supraspinatus muscle increased continuously from a resting pressure at 0 degrees of upper arm elevation to a maximal pressure at 90 degrees of upper arm elevation, for all elevation planes. The mean IMP of the supraspinatus muscle appeared to be more dependent upon the elevation plane and less dependent upon the hand load, compared to the infraspinatus muscle. Even during only moderate arm elevation, the mean IMP of the infra- and supraspinatus muscles, presented here in polar diagrams, had already exceeded the levels of reduced recovery from local muscle fatigue and blood flow impairment. The elevation angle and the hand load primarily influence the development of IMP in the infra- and supraspinatus muscles. PMID- 11104065 TI - Consistency of motor-unit identification during force-varying static contractions. AB - Due to inter-operator variability, two operators were used to assess the consistency of motor unit (MU) identification during ramp contractions, by the comparison of semi-automatic decompositions of the same recordings. Static shoulder abduction was performed against a force transducer in a position with the upper arms vertical and elbows flexed to 90 degrees. The subjects followed an 8-s force trajectory: 30% maximum voluntary contraction (MVC, 2 s), a reduction in force from 30% to 0% MVC (2 s), 0% MVC (1 s), an increase in force from 0 to 30% MVC (2 s), and 30% MVC (1 s). Muscle activity was recorded from the supraspinatus muscle with a quadripolar needle. From six recordings of 8 s duration, a total of 2527 MU firings were identified by both operators, and 93% of these were identified identically into 31 MUs. Both operators identified 8 of these MUs as continuously firing, 5 as only being active either before or after the 1 s at 0% MVC, and 18 as being de-recruited during force decreases and recruited during force increases. Both operators agreed that 16 of these 18 MUs were de-recruited at a higher force level than that at which they were recruited, which may be due to the electromechanical delay. The coefficient of variation for double determination of the results obtained by operators A and B was 8.5% for the number of MU firings, 4.5% for the MU mean firing rate, and 8.4% for the MU action potential (MUAP) amplitude. Therefore, the operator interactive decomposition method was considered to be valid for studying recruitment and de recruitment as well as firing rate and MUAP amplitude during static, force varying ramp contractions. PMID- 11104066 TI - Trapezius muscle activity, neck and shoulder pain, and subjective experiences during monotonous work in women. AB - The electromyographic (EMG) activity patterns of 18 female supermarket employees reporting neck and shoulder pain were compared with those of 6 of their female colleagues reporting no pain when doing cash-register work. It was found that the EMG activity of the trapezius muscle tended to show a lack of low and high levels among pain subjects, and that the time the trapezius muscle was at rest was longer in the group reporting no pain. In the non-dominant side, the muscle rest time was significantly longer (P < 0.05) in the group reporting no pain, and this group also showed a larger EMG activity difference between the dominant and non dominant sides, indicating a less static bilateral muscle activation. Self reports of negative experiences (stressed, exhausted and tense) were somewhat higher in the group reporting pain, while positive experiences during work (concentrated, stimulated and happy) appeared to be similar in the two groups of supermarket employees. PMID- 11104067 TI - Responses of cloned rainbow trout Oncorhynchus mykiss to an attenuated strain of infectious hematopoietic necrosis virus. AB - The objective of this work was to examine the response of homozygous clones of rainbow trout to vaccination by an attenuated strain (Nan Scott Lake; NSL) of infectious hematopoietic necrosis virus (IHNV). Adult rainbow trout of the Hot Creek Strain (YY males maintained in a recirculating system at 12 degrees C) were injected 3 times with 10(5) to 10(7) plaque forming units (pfu) of NSL. Intraperitoneal injections were given at Day 0 and at 2 and 4 mo post-infection. All fish were nonlethally bled at monthly intervals for 18 mo. Serum from each fish was analyzed by the complement-dependent neutralization assay and by western blot against purified NSL virus. The highest virus neutralization titers were detected 4 mo after the first injection, and peaked at 1280. When sera were analyzed by western blot, the predominating responses of the serum from immunized fish on the reduced western blot were against M1, a matrix protein of the virus and to a 90 kDa stress protein. The 90 kDa protein was identified by a monoclonal antibody as a stress protein derived from the CHSE-214 cells in which the purified IHN virus was grown and which associates with the virus during purification. PMID- 11104068 TI - Concomitant herpes-like virus infections in hatchery-reared larvae and nursery cultured spat Crassostrea gigas and Ostrea edulis. AB - Concomitant sporadic high mortalities were reported in France in May 1994 among batches of hatchery-reared larval Pacific oysters Crassostrea gigas and European flat oysters Ostrea edulis in 2 hatcheries, and in June and July 1994 among batches of cultured spat of both species in a shellfish nursery. Histological observation showed the presence of cellular abnormalities in moribund animals. Transmission electron microscopy revealed the presence of herpes-like virus particles in infected larvae and spat of both oyster species. This is the first description of a herpes-like virus infection in larval O. edulis. Viruses observed in diseased larvae and spat of both species are similar with respect to ultrastructure and morphogenesis. They were detected simultaneously in C. gigas and O. edulis larvae and spat, indicating possible interspecific transmission. Moreover, these viruses are associated with high mortality rates in both oyster species. An electron microscopic examination revealed hemocytes with condensed chromatin and extensive perinuclear fragmentation of chromatin. These data suggest that herpes-like viruses infecting oysters may induce apoptosis in oyster hemocytes. PMID- 11104069 TI - Protistan parasite QPX of hard-shell clam Mercenaria mercenaria is a member of Labyrinthulomycota. AB - Biomass of the protistan parasite QPX (quahaug parasite X) of hard-shell clam Mercenaria mercenaria was enriched from in vitro culture. The nuclear gene encoding the 18S RNA of the small-subunit ribosomal (ssu-rDNA) was recovered using the polymerase chain reaction (PCR) and sequenced. Phylogenetic analysis clearly showed that QPX is a member of phylum Labyrinthulomycota, within which it appears as a specific relative of Thraustochytrium pachydermum. These results confirm the provisional assignment of QPX to the Labyrinthulomycota made previously on the basis of morphological and ultrastructural characters found in some, but not all, geographic isolates. PMID- 11104070 TI - Standardization of experimental infection with Flavobacterium psychrophilum, the agent of rainbow trout Oncorhynchus mykiss fry syndrome. AB - Rainbow trout fry syndrome (RTFS) is a septicaemic infection of young rainbow trout Oncorhynchus mykiss occurring at low temperatures and responsible for severe economic losses in European fish farming. The causative agent, Flavobacterium psychrophilum, is a gliding bacterium, and difficulties in culturing it have long been an impediment to investigations on pathogenesis and immunity. Successful attempts at experimentally inducing the disease have been reported, but no experimental model resulting in well-controlled and quantitatively reproducible effects has been described. Recent improvements in F. psychrophilum cultivation made it possible to produce bacterial suspensions with nearly constant viability and to complete challenge injections in rainbow trout fingerlings, using accurately adjusted infective doses. Parenteral injection resulted in significant mortality, which was higher when administered intramuscularly (IM) than intraperitoneally (IP). Lethal doses 50 % lower than 10(3) colony forming units were consistently obtained in trout weighing 3 to 5 g, and the regular shape of the cumulative mortality curves appeared to lend itself to quantitative analyses. Bath experiments produced milder effects, although mortality ranging between 45 and 60 % was obtained in 6 g trout when skin lesions or stressors were induced along with bacterial exposure. Temperature, salinity and the process of preserving isolates (at least over short periods of time) did not seem to be associated with the severity of infection. Nevertheless, infection trials performed at 2 different locations differing both in water quality and in the system of fish maintenance resulted in different mortalities. These findings notwithstanding, the proposed IM model appears easy to apply under standardized experimental conditions and should contribute to effective advances in the study of the disease. PMID- 11104071 TI - Development of a PCR assay for detection of the oyster pathogen Bonamia ostreae and support for its inclusion in the Haplosporidia. AB - The development of diagnostic assays more sensitive and specific than traditional histological techniques is important for the management of bonamiasis in flat oysters Ostrea edulis. A specific polymerase chain reaction (PCR) protocol was developed for the detection of very small amounts of Bonamia ostreae (Pichot et al. 1980) ribosomal DNA (rDNA) in bulk DNA from oyster gill and hemolymph. The presence of a 760 bp PCR amplification product corresponded with B. ostreae infections determined cytologically in 185 oysters from Ireland, Spain, and the USA. All (100%) 'heavily' and 'moderately' infected oysters, 86.7 % of the 'lightly' infected oysters, and 66.7 % of the 'scarcely' infected oysters were confirmed to be infected using the PCR. In addition, 37.9% of the oysters in which B. ostreae was not detected using cytology were positive using the PCR. Sampling error and the subjectivity of cytological diagnoses are the likely sources of disagreement between diagnostic methods in oysters with very light infections. The PCR assay developed here is more sensitive and less ambiguous than standard histological and cytological techniques. Phylogenetic analysis of DNA sequence data confirmed B. ostreae to be a member of the Haplosporidia. PMID- 11104072 TI - Haplosporidiosis in the Pacific oyster Crassostrea gigas from the French Atlantic coast. AB - Two cases of haplosporidian infection occurred during 1993 in Pacific oysters Crassostrea gigas from the French Atlantic coast. The localization and ultrastructure of the plasmodia are described. In situ hybridization of infected tissue sections was conducted with DNA probes for oyster-infecting haplosporidians. The Haplosporidium nelsoni-specific DNA probe MSX1347 hybridized with the C. gigas parasite, and the H. costale-specific probe SSO1318 did not hybridize. Total genomic DNA was extracted from the infected tissue sections for polymerase chain reaction (PCR) amplification of the haplosporidian. PCR amplifications with H. nelsoni-specific primers and with 'universal' actin primers did not yield the expected products of 573 and 700 bp, respectively. A series of primers was designed to amplify short regions of small subunit ribosomal DNA (SSU rDNA) from most haplosporidians. The primers encompass a highly variable region of the SSU rDNA and did not amplify oyster DNA. PCR amplification of the infected C. gigas genomic DNA with these primers yielded the expected-sized product from the primer pair targeting the shortest region (94 bp). This PCR product was sequenced and it was identical to the corresponding SSU rDNA region of H. nelsoni. PMID- 11104073 TI - Prolonged in vitro cultivation of Ichthyophthirius multifiliis using an EPC cell line as substrate. AB - The ciliate Ichthyophthirius multifiliis, which normally requires a fish host to develop from the theront stage to the trophont stage, was cultivated in vitro for part of its life cycle. Experiments were conducted using a laboratory strain of the parasite originally isolated from rainbow trout Oncorhynchus mykiss in a Danish trout farm. Theronts escaping from tomontocysts were kept in water, cell culture media (E-MEM or L-15), or cultures of EPC (Epithelioma Papulosum Cyprini) cells in plastic tissue culture dishes (Nunc multidish plates). In addition, a 2 compartment system, with water separated from tissue culture media by a monolayer of EPC cells on an Anopore Tissue Culture Insert (mimicking the fish epidermis) was tested as an experimental habitat for the parasite. Theronts transformed into trophonts in all treatments except in water alone. However, development was accelerated in wells containing EPC cells, and survival and growth of trophonts were significantly increased compared to water or tissue culture media alone. Further, the 2-compartment system allowed superior performance of the parasites (attachment of parasites to cells and growth from 36 to 46 microm). In all experiments it was found that the presence of host factors (mucus and serum) stimulated parasite development. PMID- 11104074 TI - Differences in the susceptibility of some penaeid prawn species to gill associated virus (GAV) infection. AB - Four species of penaeid prawn cultured in Australia (Penaeus monodon, Penaeus esculentus, Marsupenaeus japonicus and Fenneropenaeus merguiensis) were injected with a virulent preparation of gill-associated virus (GAV). P. monodon (average weight = 8.9, 13.9 and 19.2 g), P. esculentus (average weight = 19.5 g), F. merguiensis (average weight = 10.5 g), and small (average weight = 5.8 g) M. japonicus displayed overt signs of disease and mortalities which reached 82 to 100% within 23 d post-injection. Cumulative mortalities in P. esculentus and F. merguiensis were significantly lower than for P. monodon of the same size class. Medium (average weight = 13.0 g) M. japonicus also developed overt signs of disease but cumulative mortalities were not significantly higher than uninfected controls. Large (average weight = 20.3 g) M. japoncius did not display symptoms of disease and there were no significant mortalities up to 23 d post-injection. PMID- 11104075 TI - Infection with Edwardsiella tarda causes hypertrophy of liver cells in the Japanese flounder Paralichthys olivaceus. AB - To study the direct cause of liver enlargement in the Japanese flounder Paralichthys olivaceus infected with Edwardsiella tarda, the fish were challenged with E. tarda and reared without feeding. The liver of fish exposed to the bacteria was markedly enlarged compared to that of the controls while no severe histopathological change appeared in the organ during the experiments. No notable difference was observed in the crude fat, glycogen, and water content of the liver between challenged and control fish. The size of liver cells and nuclei of the challenged fish was apparently larger than that of the controls. Analysis of crude DNA in the liver suggested that the number of liver cells of starved control fish significantly decreased during the experiment while that of the challenged fish was maintained at a level of the initial control. RNA/DNA ratio of the liver of challenged fish clearly increased while it decreased in the control fish during the experiment. These observations suggest that liver enlargement of flounder infected with E. tarda, at least in the early stage of infection, is not a result of any readily observable histopathological changes and that E. tarda infection causes hypertrophy of the cells, as well as preventing decrease in liver cell number. PMID- 11104076 TI - Lipid peroxidation induced by Clinostomum detruncatum in muscle of the freshwater fish Rhamdia quelen. AB - The effect of Clinostomum detruncatum metacercaria infection on the activities of the antioxidant enzymes superoxide dismutase and catalase in muscle of the freshwater fish Rhamdia quelen was analyzed. Tert-butyl hydroperoxide-initiated chemiluminescence, a measure of lipid peroxidation, was also investigated. Enzyme activities were similar in infected and uninfected fishes. However, the chemiluminescence was almost 2-fold higher in muscle of infected fishes than in muscle of uninfected ones. These results indicate that parasite infection induces oxidative stress and a higher level of membrane damage in the fish muscle due to an imbalance between pro-oxidants and non-enzymatic antioxidants. Our results suggest that fish response to parasite infection could involve, as in other vertebrates, reactive oxygen intermediates. PMID- 11104077 TI - A preventive approach. PMID- 11104078 TI - 'Dr' for dentists. PMID- 11104079 TI - Nitrous oxide. PMID- 11104080 TI - Shared world. PMID- 11104082 TI - Piercing difficulties PMID- 11104081 TI - NHS discrimination. PMID- 11104083 TI - Special care dentistry. PMID- 11104084 TI - Antibiotic prescribing. PMID- 11104085 TI - Frank Ashley PMID- 11104086 TI - Oral temazepam. PMID- 11104087 TI - Paediatric dentistry. PMID- 11104088 TI - Kaiser Bill PMID- 11104089 TI - Mobile telephones. PMID- 11104090 TI - Orthodontics on TV. PMID- 11104091 TI - Sinusitis prescribing. PMID- 11104092 TI - Oral cancer screening. PMID- 11104093 TI - Oral cancer screening. PMID- 11104094 TI - Special needs dentistry. PMID- 11104095 TI - Copycat papers. PMID- 11104096 TI - Specialist lists. PMID- 11104098 TI - Cycling capers PMID- 11104097 TI - Recalled attendance. PMID- 11104099 TI - Communication between the dentist and the dental technician. AB - Factors contributing to good RPD design are described, including the respective inputs of the dentist and dental technician. Poor communication in current practice is reported and an appropriate format for a work authorization presented. PMID- 11104100 TI - Root extrusion, a practical solution in complicated crown-root incisor fractures. AB - Implants and fixed and removable prostheses are very successful in replacing missing units but their cost can be inhibitory to a number of patients. In addition fixed and removable prostheses can be destructive to sound abutment teeth and can result in damage to dental and soft tissue. This report describes the restoration of a tooth with a complicated incisor crown-root fracture that extended below both the gingival cuff and the alveolar crest, by using remaining tooth tissue. The restoration was completed after root extrusion with a cast post, diaphragm and core, and porcelain crown. PMID- 11104101 TI - Disease activity and need for dental care in a capitation plan based on risk assessment. AB - This article describes a capitation model of care which would stimulate both dentists and patients to apply existing preventive knowledge. PMID- 11104102 TI - A geodemographic analysis of the Denplan patient population in the North West Region. AB - OBJECTIVE: To provide a preliminary descriptive investigation of Denplan patients in the North West Region, by plotting the age/gender and payment banding distribution, and to identify the area types where Denplan patients live and the areas in the North West Region where Denplan practices are most likely to thrive. SETTING: North West Region of England SUBJECTS AND MATERIALS: The study included Denplan patients resident in the North West Region. Age/gender and payment banding frequency distributions were constructed. A market penetration ranking report using the Target Market level of the Super Profiles geodemographic classification was produced by a spreadsheet analysis in Microsoft Excel. A Lorenz curve was plotted to graphically represent the output of the market penetration analysis. Following the market penetration analysis the enumeration districts (EDs) of the six top ranked Target Markets in the North West Region were identified and mapped out across the Region. Finally, the number and percentage of EDs in the six top ranked Target Markets were identified for each health authority in the Region. RESULTS: 47,106 patients were registered with Denplan. In all but one 5-year age band (16-20-year-olds) female patients were in the majority. Patients were concentrated (40.5%) into the 40-55 age group. Nearly 50% (22,329) of patients were allocated to the second lowest payment banding. Under 0.5% of patients (N = 199) were categorised into the highest payment band. The Target Markets at the top of the penetration ranking were more affluent in nature, with a strong rural element and an older demographic profile as part of their descriptive titles. At the bottom of the ranking deprived area types with young demographic profiles predominated. About one half (49.9%) of Denplan patients were present in just over a quarter (25.7%) of the total population of the North West Region. The Lorenz curve demonstrated that Super Profiles at Target Market level had an effectiveness of 37.9% in segmenting the population of the North West Region according to Denplan registration status. CONCLUSIONS: The population using this service in the North West Region tend to be from more mature, rural and affluent populations. The Super Profiles classification was moderately successful in segmenting the population of the North West Region according to their Denplan registration status. PMID- 11104103 TI - The provision of primary care dental general anaesthesia and sedation in the north west region of England, 1996-1999. AB - AIM: To investigate trends in the provision of primary care dental general anaesthesia (DGA) and sedation following the new guidance from the General Dental Council. DESIGN: Cross-sectional analysis of data about the provision of DGA in the General Dental Service and Community Dental Service from 1996/97 to 1998/99. SETTING: The North West Health Region of England. METHOD: The numbers of DGAs in the General Dental Service (GDS) and Community Dental Service (CDS) for the three financial years, 1996/97, 1997/98 and 1998/99 were examined. In addition General Dental Service quarterly information about the numbers of general anaesthetics, sedations requiring another dentist or doctor, and claims for conscious sedation or inhalation sedation from April 1997 to December 1999 was requested from the Dental Practice Board. RESULTS: The numbers of DGAs declined by 24 per cent between 1996/97 and 1998/99. Those in the GDS fell from 14,550 in the first quarter of 1997/98 to 3,527 in the first quarter of 1999/2000. The number of claims for sedation, which required another dentist or doctor, increased from 712 in the first quarter of 1997/98 to 2,989 in the same quarter of 1999/2000, while the number of claims for conscious sedation and inhalation sedation increased slightly from 2,847 to 2,963 over the same period. CONCLUSIONS: The revised General Dental Council guidance has reduced the numbers of DGAs being carried out in both GDS and CDS. The number of sedations involving another dentist or doctor has increased considerably but the new guidance seems to have had little effect on the numbers of patients receiving operated administered conscious sedation and inhalation sedation. PMID- 11104105 TI - Risking it. PMID- 11104104 TI - Perceived inability to cope and care-seeking in patients with toothache: a qualitative study. AB - AIMS: To explore the subjective experience of a sample of patients attending a dental teaching hospital emergency clinic with toothache. MATERIALS AND METHODS: Subjects 21 female and 14 male dental patients, of different ages, marital status, employment status and levels of education, presenting with toothache at a dental teaching hospital emergency clinic. Data collection Unstructured in-depth interviews, following a topic guide. Analysis Transcribing, sifting, indexing and charting data according to key issues and themes. FINDINGS: A dimension of toothache pain that emerged was the perceived inability to cope. Patients reported a dependency on a dentist or other person to alleviate their pain, suggesting connotations of helplessness, disempowerment and incapacitation. The perceived inability to cope was also expressed in terms of loss of control, despair and isolation. A number of care-seeking patterns for toothache was identified: repeated visits to the same dentist for emergency care, repeated visits to different dentists, attendance at the dental hospital emergency clinic and consulting non-dental health workers such as doctors and pharmacists. CONCLUSIONS: The perceived inability to cope and care-seeking patterns are two unexplored dimensions of the toothache pain experience. Both dimensions may be associated with pain intensity, the clinical conditions that manifest as toothache, quality of treatment provided and management of demand for emergency dental care. A conceptual framework is proposed for future research to investigate these relationships. PMID- 11104106 TI - Different sensitivity to ethanol in alcohol-preferring sP and -nonpreferring sNP rats. AB - BACKGROUND AND OBJECTIVES: Clinical research has proposed that initial sensitivity to ethanol may be negatively correlated with levels of subsequent ethanol intake; consistently, alcohol-preferring P rats were found to be less sensitive to the ataxic and sedative/hypnotic effects of ethanol than nonpreferring NP rats. The present study investigated the initial sensitivity to the ataxic and sedative/hypnotic effects of ethanol and to the sedative/hypnotic effects of pentobarbital and diazepam in selectively bred Sardinian alcohol preferring sP and -nonpreferring sNP rats. METHODS: In experiment 1, time to lose (onset) and regain (sleep time) the righting reflex after the acute intraperitoneal (ip) administration of 3.0 and 3.5 g/kg ethanol were measured in sP and sNP rats. In experiment 2, sP and sNP rats were required to perform a motor coordination task on a Rota-Rod after the acute intragastric administration of 2.0, 2.5, and 3.0 g/kg ethanol. Experiment 3 assessed onset and sleep time in sP and sNP rats after the acute injection of pentobarbital (40 mg/kg; ip) and diazepam (15 and 20 mg/kg; ip). RESULTS: In experiment 1, sP rats took shorter times to lose the righting reflex and regained this reflex over longer periods of time and at lower blood ethanol levels than sNP rats. In experiment 2, ethanol affected motor coordination to a greater extent in sP than sNP rats. In contrast, results from experiment 3 showed that sP and sNP rats were not differentially sensitive to the sedative/hypnotic effects of pentobarbital and diazepam. CONCLUSIONS: The results of experiments 1 and 2 suggest that sP rats possess a genetically determined, greater sensitivity to the motor impairing and sedative/hypnotic effects of ethanol than sNP rats. Although caution should be adopted before hypothesizing any comparison to humans, these results may feature sP rats as an experimental model of those subsets of human alcoholics with initial high sensitivity to ethanol challenges. Finally, the results of experiment 3 suggest a minimal involvement of the benzodiazepine and barbiturate recognition sites in the differential sensitivity to ethanol of sP and sNP rats. PMID- 11104107 TI - Serotonin, impulsivity, and alcohol use disorders in the older adolescent: a psychobiological study. AB - BACKGROUND: Alcohol use disorders (AUDs) among adolescents are associated with a high prevalence of conduct disorder (CD), much as type II alcoholism in adults is associated with impulsive-aggressive behavior and antisocial personality traits. Adults with impulsive personality disorders and AUD demonstrate diminished central serotonergic responsiveness to serotonergic agonists. Dysregulation of central serotonergic function may contribute to a vulnerability to impulsive aggressive behavior, CD, and AUD. We studied older adolescents, both male and female, to examine the relationships between sex, dispositional impulsivity, aggressivity, CD, and responsiveness to serotonergic challenge with d,l fenfluramine (FEN) early in the development of AUD. METHODS: Thirty-six adolescents between the ages of 16 and 21 years were assessed for DSM-IV AUD and other Axis I disorders by using the Psychoactive Substance Use Disorders section of the Structured Clinical Interview for DSM III-R, the Schedule for Affective Disorders and Schizophrenia for School-Age Children-Present and Lifetime Version, and CD interviews. Impulsivity and aggressivity were assessed by the Barratt Impulsiveness Scale, Lifetime History of Aggression, Buss-Durkee Hostility Inventory, Eysenck Impulsiveness Questionnaire, Youth Self Report, and Multidimensional Personality Questionnaires. FEN was administered as 0.8 mg/kg to a maximum of 60 mg, and blood was sampled at fixed intervals for prolactin, cortisol, fenfluramine, and norfenfluramine levels. RESULTS: Eighteen adolescents (12 male, 6 female) with AUD scored significantly higher on all measures of impulsivity and aggressivity compared with 18 healthy controls (12 male, 6 female). There were no significant differences between groups in peak prolactin or cortisol responses (minus baseline), or area-under-the-curve determinations (AUC); however, 9 subjects with AUD and comorbid CD had significantly elevated cortisol AUC levels compared with subjects with AUD and no CD or with normal controls. In the total sample, cortisol AUC was associated positively with measures of aggression. CONCLUSIONS: Adolescents with early-onset AUD are characterized by impulsivity and aggressivity compared with healthy peers but do not demonstrate the diminished prolactin or cortisol responses to FEN characteristic of adult alcoholics with impulsive-aggression. PMID- 11104108 TI - Thiamine status in liver and brain of rats genetically selected for different sensitivity to hypnotic effect of alcohol. AB - BACKGROUND: The mechanisms of the different sensitivity or resistance of animals and humans to alcohol are still not completely understood. For further biochemical characterization of animals genetically selected for high-alcohol sensitivity (HAS) and low-alcohol sensitivity (LAS) with the hypnotic effect of alcohol, the thiamine status and thiamine metabolizing enzymes in these animals have been studied. METHODS: We investigated thiamine diphosphate and thiamine triphosphate levels as well as the activity of thiamine-dependent enzyme, transketolase, and thiamine-metabolizing enzymes, thiamine kinase, and thiamine triphosphatase in the liver and brain of HAS, LAS, and CAS (control) rats by standard biochemical techniques. RESULTS: It was found that the activity of transketolase, and the level of the coenzyme form of thiamine, thiamine diphosphate (TDP), were significantly lower in HAS versus LAS rats. The activation of transketolase by the exogenous TDP (TDP-effect) was significantly higher in the liver and brain regions of HAS rats compared with LAS rats. The level of TDP in the liver and cerebellum of HAS rats was significantly lower compared with LAS rats. These results indicate a severe deficiency of TDP in HAS rats. HAS rats have a significantly lower activity of thiamine triphosphatase, the additional source of TDP. Accordingly, HAS rats have much higher thiamine triphosphate levels in the liver and brain, compared with LAS rats. There were no significant differences between groups with respect to the thiamine diphosphatase and thiamine kinase activity. Most of the above parameters had the intermediate values in CAS rats, compared with LAS and HAS rats. These data indicate the possible role of the thiamine phosphate esters and related enzymes in the mechanisms that bring about the differential sensitivity to the hypnotic effect of alcohol. CONCLUSIONS: HAS rats have the genetically mediated thiamine diphosphate deficiency and increased thiamine triphosphate levels, probably due to reduced activity of thiamine triphosphatase in the liver and brain, compared with LAS rats. It can be related with the higher initial sensitivity of HAS rats to hypnotic effect of ethanol. PMID- 11104109 TI - No sex and age influence on the expression pattern and activities of human gastric alcohol and aldehyde dehydrogenases. AB - BACKGROUND: Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are the principal enzymes responsible for ethanol metabolism in humans. The stomach is involved in the metabolism of alcohol during absorption. Conflicting reports exist with regard to the influence of sex and age on the activity of ADH in the human gastric mucosa. The purpose of the present study was to determine the effects of age and sex on the expression pattern and activities of stomach ADH and ALDH. METHODS: A total of 115 endoscopic gastric biopsy specimens were investigated from Han Chinese men (n = 70) and women (n = 45) aged 20-79 years with approximately even distribution among 10-year age intervals. The expression patterns of ADH and ALDH were identified by isoelectric focusing, and the activities were assayed spectrophotometrically. RESULTS: The expression patterns of gastric ADH and ALDH remained unchanged with respect to sex and age. At 33 mM or 500 mM ethanol, pH 7.5, the ADH activities did not differ significantly among the various age groups or between men and women. At 200 microM or 20 mM acetaldehyde, the ALDH activities did not differ significantly in relation to sex and age. No correlations were found between the ADH or ALDH activities at both the high and low substrate concentrations and the ages in men and women. CONCLUSIONS: The results indicate that there is no significant effect of either sex or age on the expression pattern and activity of ADH and ALDH in the human gastric mucosa. The stomach ADH seems unlikely to account for possible variations in the first-pass metabolism of alcohol with regard to sex and age. PMID- 11104110 TI - Nicotine is more reinforcing in smokers with a past history of alcoholism than in smokers without this history. AB - BACKGROUND: Whether smokers with a past history (PH) but not current history of alcohol dependence are more nicotine dependent than smokers with no such history (NH) is unclear. The present study was an experimental test of this hypothesis. METHOD: Twenty PH and 10 NH smokers abstained from smoking for 16 hr on each of 4 days. On each of 3 days, participants received three doses per day of 0, 2, or 4 mg nicotine gum in a within-subjects, randomized, double-blind, crossover design. To examine subjective effects, participants completed the Profile of Mood States, Addiction Research Inventory, and other ratings before and after each dose. To examine nicotine reinforcement, participants reported preference among the gums, reported on money versus gum choices, and, on the 4th day, underwent a double blind self-administration test. RESULTS: Across the 21 subjective measures, with one exception, PH and NH smokers did not differ in subjective response to nicotine. However, across all three reinforcement measures, nicotine was a more potent reinforcer in PH than NH smokers. CONCLUSIONS: These results provide a behavioral mechanism to explain prior findings that PH smokers are more nicotine dependent than NH smokers. PMID- 11104111 TI - Validation of multidimensional scaling-based modeling of alcohol expectancies in memory: age and drinking-related differences in expectancies of children assessed as first associates. AB - BACKGROUND: As evidence has accumulated that alcohol expectancies mediate the effects of other drinking antecedents, attempts to understand the mechanism by which expectancies influence behavior have focused on modeling memory processes. Previous expectancy work, however, has used relatively indirect approaches to retrieve and model information stored in memory. By using the method most recommended by memory researchers for directly obtaining uncontaminated memory contents, we assessed children's expectancies and related findings to empirically modeled organization and activation of expectancies in memory based on scaled instruments. METHODS: Individual interviews were conducted with 462 children in 2nd through 5th grades, and surveys were completed by 1,003 children in 3rd, 6th, 9th, and 12th grades. Interviews and surveys consisted of a measure designed to retrieve participants' first expectancy associate to an alcohol prompt and several drinking quantity/frequency questions. RESULTS: Older and higher drinking children were more likely to report positive expectancies as their first associate to an alcohol prompt. Age and drinking-based findings were consistent with organizational structure, dimension emphasis shift, and paths of association identified by prior multidimensional scaling techniques. CONCLUSIONS: Consumption of alcohol among children corresponded to accessibility of positive expectancies in memory. In addition, the use of multidimensional scaling to study the organization and activation of alcohol expectancies in memory was validated. PMID- 11104112 TI - Do risk factors for re-arrest differ for female and male drunk-driving offenders? AB - BACKGROUND: The present study investigated gender differences in factors affecting recidivism among 628 female and 659 male drunk-driving offenders. The study population included residents from New Mexico who completed a screening program for offenders and who were still residents when contacted 5 years later. METHOD: Risk factors for re-arrest in the 5-year period after screening referral were examined using multiple logistic regression models. Predictor variables included gender, age, ethnicity, education, marital status, blood alcohol concentration at arrest, parental alcohol problems, spousal alcohol problems, lifetime use of cannabis, cocaine, or amphetamines, abusive behavior toward spouse, and scores on two standardized assessments. RESULTS: Risk factors for re arrest were similar for males and females except that young age predicted higher recidivism among males but not females. The overall 5-year re-arrest rate was 26% 20% for women, 38% for males age 30 and under, and 24% for males age 31 and older. CONCLUSIONS: Young age predicts re-arrest for males but not for females. Neither the type of risk factors nor the number of risk factors fully explained female offenders' disproportionately lower recidivism rates, compared with young males. PMID- 11104113 TI - Effects of alcohol on the response to hyperventilation of participants high and low in anxiety sensitivity. AB - BACKGROUND: Previous research suggests that high levels of anxiety sensitivity (AS; fear of anxiety symptoms) may constitute a risk factor for alcohol abuse. The present study evaluated the hypothesis that high AS levels may increase risk for alcohol abuse by promoting a heightened sober reactivity to theoretically relevant stressors and heightened sensitivity to alcohol's emotional reactivity dampening effects, which would negatively reinforce drinking in this population. METHODS: One hundred and two undergraduate participants (51 high AS, 51 low AS) with no history of panic disorder were assigned to either a placebo, low-dose alcohol, or high-dose alcohol beverage condition (17 high AS, 17 low AS per beverage condition). After beverage consumption and absorption, participants underwent a 3 min voluntary hyperventilation challenge. RESULTS AND CONCLUSIONS: High-AS/placebo participants displayed greater affective and cognitive reactivity to the challenge than low-AS/placebo participants, which indicated increased fear and negative thoughts (e.g., "losing control") during hyperventilation among sober high AS individuals. Dose-dependent alcohol dampening of affective and cognitive reactivity to hyperventilation was observed only among high-AS participants, which suggested that high-AS individuals may be particularly sensitive to alcohol-induced reductions in their degree of fear and negative thinking in response to the experience of physical arousal sensations. In contrast, dose-dependent alcohol dampening of self-reported somatic reactivity was observed among both high- and low-AS participants. We discuss implications of these results for understanding risk for alcohol abuse in high-AS individuals, as well as directions for future research. PMID- 11104114 TI - Alcohol use in the year before marriage: alcohol expectancies and peer drinking as proximal influences on husband and wife alcohol involvement. AB - BACKGROUND: Models of adolescent alcohol involvement that include individual difference, family, and peer risk factors indicate a significant association between the drinking of adolescents and that of their peers. Peer drinking influences, however, have not been investigated extensively in integrative models of adult drinking. The purpose of this study was to test a model of adult drinking that incorporated the potentially important risk factor of partner drinking and in which proximal risk factors (peer drinking, alcohol expectancies) were hypothesized to be strongly associated with adult alcohol use and to mediate relationships between more distal risk factors and drinking. METHODS: Couples (n = 389) were assessed at the time of their first marriage. Separate, self administered questionnaires were completed at home by both husbands and wives. Distal risk factors included family history of alcoholism, antisocial behavior, and depressive symptomatology. Substantive relationships were tested in a model that included spousal associations with respect to distal risk factors, proximal risk factors, and drinking. RESULTS: Findings demonstrate the unique association of alcohol expectancies and peer drinking with adult alcohol use. Of particular relevance is the significance of the social network as a correlate of adult drinking. A peer network characterized by a higher level of alcohol involvement was strongly associated with heavier drinking among both men and women. This relationship was independent of sociodemographic and individual difference factors, alcohol expectancies, and partner's drinking. Results also demonstrate the similarity between husband and wife drinking, an association that cannot be attributed to assorting with respect to the other risk factors. CONCLUSIONS: The social network continues to significantly impact drinking behavior in adulthood. The relevancy of peer and partner drinking influences to adult alcohol involvement suggests that the immediate social environment may have a prominent role in the continuity/discontinuity of heavy or problem drinking during the transition to marriage. PMID- 11104115 TI - Brief intervention for female heavy drinkers in routine general practice: a 3 year randomized, controlled study. AB - BACKGROUND: Today, heavy drinking is a common health hazard among women. The evidence in favor of providing some kind of brief intervention to reduce drinking is quite convincing. However, we do not know if intervention works in a natural environment of routine health care. The purpose of this study was to evaluate the effectiveness of long-lasting, brief alcohol intervention counseling for women in a routine general practice setting. METHODS: In five primary care outpatient clinics in a Finnish town, 118 female early-phase heavy drinkers who consulted their general practitioners for various reasons were given brief alcohol intervention counseling. Intervention groups A (n = 40) and B (n = 38) were offered seven and three brief intervention sessions, respectively, over a 3-yr period. The control group C (n = 40) was advised to reduce drinking at baseline. Main outcome measures were self-reported weekly alcohol consumption, carbohydrate deficient transferrin, mean corpuscular volume (MCV), aspartate aminotransferase, alanine aminotransferase, and gamma-glutamyltransferase. RESULTS: Depending on the outcome measure and the study group, clinically meaningful reduction of drinking was found in 27% to 75% of the heavy drinkers. Within all the groups, MCV significantly decreased. However, there were no statistically significant differences between study groups A, B, and C in the mean changes between the beginning and endpoint in the main outcome measures. CONCLUSIONS: The present study indicated that minimal advice, as offered to group C, was associated with reduced drinking as much as the brief intervention, as offered to groups A and B, given over a 3-yr period. Furthermore, in the routine setting of the general practice office, the effectiveness of the brief intervention may not be as good as in special research conditions. The factors possibly reducing the effectiveness in a routine setting are unknown. Thus, different methods of implementing brief intervention need to be evaluated to find better ways to support general practice personnel in their efforts to help heavy-drinking female patients to reduce their drinking. PMID- 11104116 TI - Application of a quality of life measure, the life situation survey (LSS), to alcohol-dependent subjects in relapse and remission. AB - BACKGROUND: Recent studies have shown that quality of life (QOL) is improved significantly when subjects do not relapse to heavy drinking, and QOL deteriorates significantly on prolonged relapse. This article further investigates these relationships using a QOL index, the Life Situation Survey (LSS). METHODS: Eighty-two DSM-IV alcohol-dependent subjects admitted for alcohol detoxification were studied at baseline and 12 week follow-up. Sociodemographic data were collected, and severity of alcohol dependence (SADQ) and General Health Questionnaire (GHQ-12) were baseline indices only. The main outcome measure, the LSS, was administered at both time points. RESULTS: Two subjects were lost to follow-up and one died during the study period. Thus, the relapse/ nonrelapse analysis related to 79 subjects. Fifty subjects (63%) had relapsed to heavy drinking at 3 months follow-up. There was a significant correlation between LSS and GHQ-12 scores. Significant changes occurred in total LSS scores as a result of relapse and nonrelapse. The improvement in LSS scores associated with nonrelapse was larger than the deterioration that accompanied relapse. In those subjects who did not relapse to heavy drinking, the mean follow-up score remained in the poor/borderline LSS range. Remission from heavy drinking was accompanied by significant improvements in appetite, sleep, and self-esteem. Relapse to heavy drinking coincided with a significant deterioration in mood/affect, public support, and work/life role scores. CONCLUSION: QOL as assessed by the LSS in recently detoxified alcoholics is impaired significantly. In the nonrelapse group, there was a significant improvement in LSS scores after 3 months. Relapse was accompanied by a smaller deterioration in LSS scores. The LSS can play an important role in monitoring the clinical care and progress of alcohol-dependent subjects. PMID- 11104117 TI - Skeletal response to alcohol. AB - This review briefly assesses the well-established effects of alcohol consumption on bone and mineral metabolism and addresses areas of controversy that need additional research. Alcohol consumption is a risk factor for osteoporosis based on the frequent finding of a low bone mass, decreased bone formation rate, and increased fracture incidence in alcoholics. Alcohol also has been shown to reduce bone formation in healthy humans and animals and to decrease proliferation of cultured osteoblastic cells. On the other hand, it has been difficult to demonstrate alcohol-induced bone loss and increased fracture rate in population based studies. Indeed, most population-based studies have shown a positive association between alcohol and bone mass and no change or a decrease in fracture risk. Overall, the evidence generally supports a detrimental effect of chronic alcohol abuse on the skeleton of men and a neutral or generally beneficial effect of light to moderate alcohol consumption, especially in older women. This latter putative beneficial effect may be due to a reduction in the age-related increase in bone remodeling associated with postmenopausal bone loss. Specific areas of research are recommended to clarify the dose and sex effects of alcohol consumption and to determine cellular and molecular mechanisms of action. The goals of this proposed research emphasis are to determine the degree of risk for the range of alcohol consumption, to set guidelines of consumption compatible with maintaining bone health, and to develop appropriate countermeasures to prevent or reverse the detrimental skeletal effects of alcohol abuse. PMID- 11104118 TI - Effects of moderate and high doses of alcohol on attention, impulsivity, discriminability, and response bias in immediate and delayed memory task performance. AB - BACKGROUND: Prior studies that examined the effects of alcohol on Continuous Performance Test (CPT) performance have resulted in inconsistent outcomes. Most studies that examined the effects of alcohol on concentrated attention tasks (like the CPT) found little effect of alcohol on performance measures, even when doses that exceeded 0.8 g/kg were used. One likely reason for these inconsistencies is the varying difficulty (and sensitivity) of the task used, and as a result, comparisons between studies are difficult. This study is one in a series that examines the effects of alcohol on attention by using a difficult version of the CPT (Immediate and Delayed Memory Task--IMT/DMT). Our purpose for these studies has been two-fold, examining the effects of alcohol (1) on concentrated attention (i.e., correct detections) and (2) on errors (i.e., commission errors) previously correlated with impulsive behaviors. The first is important because previous studies have shown little effect of alcohol on attention, and the second is important because commission errors have been related to impulsive behaviors. METHODS: In the IMT/DMT, participants respond to a briefly displayed number when it is identical to the one displayed before it. The procedure includes immediate and delayed conditions where successive stimuli to be matched are delayed by 0.5 sec or by 3.5 sec. The three stimulus types included target (identical match), catch (four of five digits match), and filler (no match) stimuli. Twenty subjects completed this task after consuming either a placebo drink or a drink that contained 0.5 g/kg or 1.0 g/kg of alcohol on different days. RESULTS: The main findings were that (1) alcohol decreased the percentage of correct identifications of target stimuli; (2) alcohol increased the percentage of commission errors in relation to the number of correct target responses; and (3) alcohol decreased discriminability whereas response bias became more conservative. CONCLUSIONS: These results clearly demonstrated a time course effect of the 1.0 g/kg alcohol dose on attention, impulsivity, discrimination, and response criteria when a variety of dependent measures are used. PMID- 11104119 TI - Binge ethanol consumption causes differential brain damage in young adolescent rats compared with adult rats. AB - BACKGROUND: Adolescents respond differently to alcohol than adults. Furthermore, binge drinking in young adolescents is becoming increasingly common. METHODS: To determine if the effects of binge drinking on brain damage are different in juveniles compared with adults, the effects of a 4 day binge ethanol treatment (e.g., 4 days of 4 times per day 15% ethanol intragastrically, approximately 9-10 g/kg/day ethanol) were investigated in adolescent-juvenile rats (JVN) 35 days old and compared with adult (ADT) rats 80 to 90 days old. Brain damage was measured by using the amino cupric silver stain of de Olmos et al. (1994). RESULTS: Significant brain damage was found in both groups. The olfactory bulbs were equally damaged in both groups; however, the associated frontal cortical olfactory regions were damaged only in JVN. The anterior portions of the piriform and perirhinal cortices also were damaged only in JVN rats. Quantitation of silver-stained frontal areas in binge ethanol-treated JVN rats ranged from 400% to 1,260% of control values. For example, in anterior perirhinal cortex, silver stain increased from 48 +/- 14 to 444 +/- 114 (mm2 x 10(3) argyrophilic area; p < 0.01) in JVN control and binge ethanol-treated animals, respectively. In contrast, posterior perirhinal cortex showed greater damage in adults, being 236 +/- 76 vs. 875 +/- 135 (mm2 x 10(3) argyrophilic area; p < 0.005) in JVN and ADT, respectively. CONCLUSIONS: The young-adolescent brain shows differential sensitivity to alcohol-induced brain damage compared with adults. PMID- 11104120 TI - Event-related potentials and cue-reactivity in alcoholism. AB - BACKGROUND: Relapse is a major problem in the treatment of addictive behaviors. Conditioning models of alcohol addiction suggest that stimuli associated with previous drug use (cues) may initiate relapse in a definite group of alcoholics. Event-related potentials (ERPs) might be useful to reveal the brain functional substrates of cue-reactivity. METHODS: In a preliminary investigation, 11 alcohol dependent patients who did not take part in the electrophysiological study completed a structured interview to rate 80 words as to the degree of relatedness to alcohol. Based on these results, cue-reactivity for 15 alcohol-related and 15 unrelated word cues, each repeated eight times, was investigated in 19 alcohol dependent men (44.2 +/- 8.5 years) and 19 healthy control men (42.5 +/- 12.5 years). RESULTS: A cue-reactivity that consisted of significantly higher amplitudes in the ERPs after alcohol-related words compared with unrelated words was found in alcohol-dependent patients, but not in controls, at the electrode location Pz [F(1,36) = 5.2,p < 0.05]. CONCLUSIONS: Consistent with the hypothesis, only alcohol-dependent patients were characterized by signs of increased cerebral activity associated with alcohol-related compared with unrelated cues. Therefore, the results support the concept of cue-reactivity in alcoholism based on a neurobiological measurement. Future investigations will show whether this cue-reactivity can be applied to assess the risk of relapse in individual alcohol-dependent patients. PMID- 11104121 TI - Serotonergic modulation of ethanol-induced electrophysiological depression in young and aged rats. AB - BACKGROUND: Ethanol (EtOH)-induced electrophysiological depressions in cerebellar Purkinje neurons have been shown to be potentiated by exogenously applied serotonin (5HT). In this study, we determined whether this modulatory action can be activated by endogenous release from presynaptic serotonergic terminals, and whether such a response is altered by age or rat strain. METHODS: Extracellular 5HT levels in cerebellar cortex were measured in real time by in vivo chronoamperometry, by using Nafion-coated carbon fiber electrodes, in anesthetized young (3-5 months old) or aged (18-24 months old) Sprague Dawley and Fischer 344 rats. Some animals were prelesioned with 5,7 dihydroxytryptamine (5,7 DHT). Single unit electrophysiological activity was recorded from cerebellar Purkinje neurons. Serotonin or its presynaptic antagonist methiothepin was applied directly to cerebellar neurons through multibarrel pipettes. RESULTS: Local application of methiothepin dose-dependently induced 5HT overflow in young Sprague Dawley and Fisher 344 rats. Methiothepin-induced 5HT release was decreased significantly in aged or 5,7 DHT-lesioned rats. Local application of methiothepin or 5HT potentiated EtOH-induced electrophysiological depression of Purkinje neurons in young animals of both strains. Methiothepin-potentiated, EtOH elicited neuronal inhibition was reduced greatly in aged or 5,7 DHT-lesioned rats. Serotonin-facilitated EtOH responses were reduced in the aged Sprague Dawley rats. CONCLUSIONS: EtOH-induced electrophysiological responses in cerebellum can be facilitated by endogenous 5HT release by using a 5HT autoreceptor antagonist. Such actions are attenuated in aged rats perhaps through a presynaptic serotonergic mechanism. PMID- 11104122 TI - Depression of neuronal firing rates in somatosensory and posterior parietal cortex during object acquisition in a prehension task. AB - Prehension is an object-oriented behavior consisting of four components: reach, grasp, manipulation, and release. To determine how such actions are represented in primary somatosensory (S-I) and posterior parietal cortex (PPC), we used digital video to synchronize spike trains of neurons recorded in Brodmann's areas 3b, 1, 2, 5, and 7 with the hand kinematics as monkeys performed a prehension task. Statistical analyses indicated that one-third of task-modulated neurons showed significantly depressed firing rates during object acquisition and/or manipulation. This population was dominated by neurons innervated by deep receptors that sensed extension movements of the fingers, or by tactile receptors in hairy skin sensing stretch. Grasp-inhibited responses were the most common type. Tonic firing rates of these cells dropped significantly during approach as the hand was preshaped for grasping, or at contact when grasp was initiated, and persisted until hand motion ceased or as the grip relaxed. Maximum suppression of firing occurred at grasp completion. Their lack of specificity for particular hand behaviors formed the inhibitory counterpart of broadly tuned cells that fired prolonged bursts during grasp and manipulatory stages of prehension. The remainder of the task-inhibited population showed biphasic responses. Firing rates were significantly depressed during grasping and manipulation when the hand interacted directly with the object, but were enhanced prior to contact, when the hand was preshaped (approach-tuned), or upon relaxation of grasp and release of the object from the hand (loweror relax-tuned). Grasp-inhibited responses occurred primarily in S-I, whereas biphasic inhibitory activity was recorded mainly in PPC. Suppression of activity within these populations may thereby increase the saliency of excitatory responses to acquisition and manipulation of objects. Reduction of firing during prehension might also signal the flexed postures used to retain objects in the hand, rather than a generalized gating of sensory information. The similarity of responses to active and passive extension movements suggests that the inhibitory responses may provide important postural and motor information about the hand kinematics when performing skilled tasks. PMID- 11104123 TI - Visual and non-visual cues in the perception of linear self-motion. AB - Surprisingly little is known of the perceptual consequences of visual or vestibular stimulation in updating our perceived position in space as we move around. We assessed the roles of visual and vestibular cues in determining the perceived distance of passive, linear self motion. Subjects were given cues to constant-acceleration motion: either optic flow presented in a virtual reality display, physical motion in the dark or combinations of visual and physical motions. Subjects indicated when they perceived they had traversed a distance that had been previously given to them either visually or physically. The perceived distance of motion evoked by optic flow was accurate relative to a previously presented visual target but was perceptually equivalent to about half the physical motion. The perceived distance of physical motion in the dark was accurate relative to a previously presented physical motion but was perceptually equivalent to a much longer visually presented distance. The perceived distance of self motion when both visual and physical cues were present was more closely perceptually equivalent to the physical motion experienced rather than the simultaneous visual motion, even when the target was presented visually. We discuss this dominance of the physical cues in determining the perceived distance of self motion in terms of capture by non-visual cues. These findings are related to emerging studies that show the importance of vestibular input to neural mechanisms that process self motion. PMID- 11104124 TI - Activation of the cerebellum in co-ordinated eye and hand tracking movements: an fMRI study. AB - Dysfunction of the cerebellum leads to significant deterioration of movements performed under visual guidance and of co-ordinated eye and hand movement. Visually guided tracking tasks combine both of these control features, as the eyes and hand together track a visual target. To better understand the involvement of the cerebellum in tracking tasks, we used functional magnetic resonance imaging to study the activation of cerebellar structures in visually guided tracking movements of the eye and hand. Subjects were tested performing ocular tracking, manual tracking without eye movement or combined eye and hand tracking of a smoothly moving visual target. Three areas were activated in the cerebellum: a bilateral region in the ansiform lobule of the lateral hemisphere, a region in the ipsilateral paramedian lobule and a region in the oculomotor vermis. The ansiform and paramedian areas were most strongly activated by hand movement, although the vermal site was also active. The reverse was found for ocular tracking, with predominantly vermal activation. Activation of these cerebellar cortical areas related to movement of eyes or hand alone was significantly enhanced when the subjects performed co-ordinated eye and hand tracking of a visual target. These results provide the first direct evidence from a functional-imaging study for cerebellar activation in eye and hand co ordination. PMID- 11104125 TI - Enhanced responsiveness of human extravisual areas to photic stimulation in patients with severely reduced vision. AB - Lesions in the primary visual cortex induce severe loss of visual perception. Depending on the size of the lesion, the visual field might be affected by small scotomas, hemianopia, or complete loss of vision (cortical blindness). In many cases, the whole visual field of the patient is affected by the lesion, but diffuse light-dark discrimination remains (residual rudimentary vision, RRV). In other cases, a sparing of a few degrees can be found (severely reduced vision, SRV). In a follow-up study, we mapped visually induced cerebral activation of three subjects with SRV using functional magnetic resonance imaging. We were especially interested in the visual areas that would be activated if subjects could perceive the stimulus consciously although information flow from V1 to higher visual areas was strongly reduced or virtually absent. Because subjects were only able to discriminate strong light from darkness, we used goggles flashing intense red light at a frequency of 3 Hz for full visual field stimulation. Besides reduced activation in V1, we found activation in the parietal cortex, the frontal eye fields (FEF), and the supplementary eye fields (SEF). In all patients, FEF activation was pronounced in the right hemisphere. These patterns were never seen in healthy volunteers. In a patient who recovered completely, we observed that extrastriate activation disappeared in parallel with the visual field restitution. This result suggests that damage to the primary visual cortex changes the responsiveness of parietal and extravisual frontal areas in patients with SRV. This unexpected result might be explained by increased stimulus-related activation of attention-related networks. PMID- 11104126 TI - Hydrocephalus induced by immunological blockage of the subcommissural organ Reissner's fiber (RF) complex by maternal transfer of anti-RF antibodies. AB - Stenosis of the cerebral aqueduct seems to be a key event for the development of congenital hydrocephalus. The causes of such a stenosis are not well known. Overholser et al. in 1954 (Anat Rec 120:917-933) proposed the hypothesis that a dysfunction of the subcommissural organ (SCO) leads to aqueductal stenosis and congenital hydrocephalus. The SCO is a brain gland, located at the entrance of the cerebral aqueduct, that secretes glycoproteins into the cerebrospinal fluid that, upon release, assemble into a fibrous structure known as Reissner's fiber (RF). By the permanent addition of new molecules to its rostral end, RF grows and extends along the aqueduct, fourth ventricle, and central canal of the spinal cord. The immunological blockage of the SCO-RF complex has been used to test Overholser's hypothesis. The following was the sequence of events occurring in pregnant rats that had been immunized with RF glycoproteins: the mother produced anti-RF antibodies and transferred them to the fetus through the placenta and to the pup through the milk, and the antibodies reached the brain of the fetus and pup and blocked the SCO-RF complex. This resulted in a permanent absence of RF that was followed by stenosis of the cerebral aqueduct, and then by the appearance of hydrocephalus. The latter was patent until the end of the 6-month observation period. The chronic hydrocephalic state appeared, in turn, to induce new alterations of the SCO. It is concluded that a selective immunological knock out of the SCO-RF complex leads to hydrocephalus. PMID- 11104127 TI - Changes in motor representation related to facial nerve damage and regeneration in adult rats. AB - The present study examined how facial nerve regeneration shapes movement representation patterns in previously disconnected motor cortices. Electrical microstimulation was used to bilaterally map the motor cortices of adult rats subjected to unilateral facial nerve lesion and reanastomosis stamps at the stylomastoid foramen level. The motor cortex output patterns of two groups of experimental hemispheres (contralateral to lesioned side) were compared before and after facial nerve reinnervation. The motor cortex output patterns in the control hemispheres (ipsilateral to the lesioned side) were used as reference. Before facial nerve reinnervation, the motor cortex forelimb and eye output area extended into the vibrissa area; such enlargement did not occupy the medial part of the former vibrissa area where ipsilateral vibrissa or neck movements were represented. After facial nerve reinnervation, the contralateral vibrissa movement reorganized into a shrunken cortical area corresponding to the medial portion of the former vibrissa representation, i.e., where the ipsilateral vibrissa and neck movements were mainly represented prior to facial nerve reinnervation. The enlargement of the forelimb and eye representation remained unchanged, even after the vibrissa motor innervation was reactivated. Before facial nerve reinnervation into expanded forelimb and eye representations, a minimal current was required to evoke these movements, which did not vary from the normal range. A higher current was necessary to evoke the ipsilateral vibrissa and neck movements in the medial part of the vibrissa representation than the threshold needed to elicit the vibrissa movement normally represented in this cortical region. After facial nerve reinnervation, the overall current required to evoke movement remained the same as that which evoked movement before the vibrissa motor innervation was reactivated. PMID- 11104128 TI - Cerebral activation during bicycle movements in man. AB - The cerebral activation during bicycle movements was investigated by oxygen-15 labelled H2O positron emission tomography (PET) in seven healthy human subjects. Compared to rest active bicycling significantly activated sites bilaterally in the primary sensory cortex, primary motor cortex (M1) and supplementary motor cortex (SMA) as well as the anterior part of cerebellum. Comparing passive bicycling movements with rest, an almost equal activation was observed. Subtracting passive from active bicycle movements, significant activation was only observed in the leg area of the primary motor cortex and the precuneus, but not in the primary sensory cortex (S1). The M1 activation was positively correlated (alpha=0.75-0.85, t=6.4, P<10(-5)) with the rate of the active bicycle movements. Imagination of bicycle movements compared to rest activated bilaterally sites in the SMA. It is suggested that the higher motor centres, including the primary and supplementary motor cortices as well as the cerebellum, take an active part in the generation and control of rhythmic motor tasks such as bicycling. PMID- 11104129 TI - Control of eye movement reflexes. AB - We investigated the effect of strategic suppression of reflexive eye movements on external control over fixation using a fixation offset paradigm. A visual signal at fixation facilitates the fixation reflex and inhibits eye movements. Certain preparatory states render the fixation reflex less reactive to visual stimulation at fixation, as evidenced by a reduction in the fixation offset effect (FOE). For example, past studies have suggested that the reduced FOE during anti-saccade tasks results from the requirement to inhibit reflexive eye movements. We tested whether suppressing reflexive saccades reduces external control over ocular fixation using a go-nogo saccade paradigm. During each trial, one of two targets appeared in the periphery. Participants were instructed to saccade to one target (go), but when the other target appeared they either had to maintain fixation (nogo) or move their eyes in the direction opposite the target (anti). When nogo trials were admixed with target-directed saccades a large FOE was observed compared to when target-directed saccades occurred alone (experiment 1); however, when anti-saccades were mixed with target-directed saccades, a small FOE was observed for both types of eye movements (experiment 2). We conclude that suppressing reflexive eye movements does not reduce external control over fixation. Further research is necessary to elucidate which other component of preparing to make an anti-saccade diminishes the FOE. PMID- 11104130 TI - The effects of instability and additional hand support on anticipatory postural adjustments in leg, trunk, and arm muscles during standing. AB - We investigated the role of additional perceptual information (finger touch) and additional mechanical support (hand grasp) on the anticipatory postural adjustments (APAs) associated with fast arm movements performed by standing subjects. The subjects performed fast, unilateral shoulder ante-flexion movements while standing on a stable force platform or on an unstable board with instability in a sagittal or in a frontal plane. Changes in the background activity of leg, trunk, and arm muscles and displacements of the center of pressure were quantified within time intervals typical for APAs. Leg and trunk muscles showed a significant drop in APAs with added finger touch and no further changes when the touch was substituted with hand grasp. Arm muscles showed no changes or a small drop in APAs with touch and a significant increase in APAs with grasp. These changes were seen during both stable and unstable standing. We conclude that APAs can show changes associated not only with mechanical aspects of a task, but also with its perceptual aspects. Based on the equilibrium-point hypothesis of motor control, an additional analysis of APAs was performed. With additional support, we observed a significant modulation of an index related to the co-activation of agonist-antagonist muscle pairs, while no changes in reciprocal activation were found. A similar analysis performed across all the leg and trunk muscles with respect to control of the center of mass lead to similar results. We conclude that the central nervous system seems to simplify adjustments of control patterns to changes in task parameters by modulation of only one of the two major central commands. PMID- 11104131 TI - Development of spontaneous leg movements in infants with and without periventricular leukomalacia. AB - The main question asked in the present study was whether support could be found for the notion that supraspinal influences on the generation of spontaneous kicking movements become increasingly apparent in the first half-year after birth. In comparing groups of infants with and without damage in tracts connected with the cortex surrounding the central sulcus, such support would consist of the finding that similar patterns of spontaneous kicking are observed early in development, whereas differences between groups should occur with increasing age. Using 3-D registrations, the spontaneous kicking movements of 19 infants with differing degrees of periventricular, lobar, and subcortical leukomalacia based on white matter (WM) abnormalities on ultrasound were compared to those of 10 healthy control infants at 6, 12, 18, and 26 weeks of corrected age. Magnetic resonance imaging recordings were used to identify the location and severity of the brain lesions. Infants with extensive lesions in the periventricular and lobar WM with or without diffuse lesions in the subcortical WM showed a decreased variability on some spatial and temporal parameters of kicks. More importantly, these infants showed a different developmental course for intralimb couplings when compared to the other infants; they were unable to dissociate tight intralimb couplings at 18 and 26 weeks. As all of these infants had substantial damage of the corticospinal tracts, these findings suggest that these tracts are involved in the regulation of intralimb joint dissociations between 4 and 6 months of age. However, caution is needed as areas outside those in which the corticospinal tracts are located could be damaged as well and most of the infants with moderate to severe lesions in the corticospinal tract had additional psychomotor problems. For interlimb couplings and most of the spatial and temporal parameters of kicks, no differences were found between groups. This strengthens the claim that inter- and intralimb couplings are organized in fundamentally different ways. PMID- 11104132 TI - Movement-related potentials are task or end-effector dependent: evidence from a multifinger experiment. AB - In a number of recent studies, the specific sensitivity of movement-related EEG potentials toward experimental manipulations of motor tasks using the index finger as a primary end-effector is well documented. The major question in this study was whether different movement-related EEG components are primarily end effector or task dependent. Accordingly, the experimental task (i.e., the rate of force development - a ratio of peak force to time-to-peak force) was systematically manipulated and the effects of this manipulation on movement related potentials were examined while subjects used either the index, middle, ring or little finger. Significant effects observed in this study were due mainly to the sensitivity of movement-related potentials preceding movement onset (Bereit shafts potential and motor potential) toward the specific finger performing the task and the sensitivity of components accompanying the task (movement-monitoring potential) toward the rate of force development. In addition, both movement-related potentials preceding and accompanying movement significantly changed as a function of the finger performing the slow task (lower rate of force development) with maximum values observed for the ring finger and minimal values observed for the index finger. Behaviorally subjects were less accurate during slow tasks regardless of the finger performing the task. In contrast, the amplitude of neither early nor late components of movement-related potentials changed as a function of the finger performing the fast task (higher rate of force development). Overall, our results are consistent with the notion that the whole complex of movement-related EEG potentials reflect a combination of factors including the selection of corresponding general motor programs as reflected in the amplitude of potentials preceding movement and specific elements of the task including rate of force development as reflected in the amplitude of potentials accompanying movement execution. PMID- 11104133 TI - Relative stability improves with experience in a dynamic standing task. AB - This study tested the hypothesis that subjects improve their relative stability as they learn a dynamic pulling task. Healthy adult subjects practiced making brief horizontal pulls (<300 ms) on a handle to a range of target forces ranging from 20 to 80% of their estimated maximum for 5 days. They were instructed to always keep their feet flat and begin and end their motion in an upright posture. In order to do this, subjects had to develop the appropriate body momentum prior to the pull and then recover their balance following the pull. We analyzed relative stability during balance recovery, using two measures: spatial safety margin (minimum distance of the center of pressure, COP, to the edges of the feet) and temporal safety margin (minimum extrapolated time for the COP to reach the edges of the feet). We hypothesized that: (1) spatial and temporal safety margins would be uncorrelated; (2) safety-margin means would increase with practice; and (3) safety-margin standard deviations would decrease with practice. Two experiments were conducted: one where subjects practiced three force targets and positioned their initial COP in a small window, and one where subjects practiced two force targets with no initial COP constraint. Results showed that spatial and temporal safety margins were correlated but shared less than 6% variance, indicating that they reflected different aspects of control. Safety margin averages increased with practice and standard deviations decreased with practice, indicating that the stability of balance control in the execution of this task became more robust. We suggest that the nervous system could use safety margins in both feedback and feedforward control of balance. PMID- 11104134 TI - Interjoint coordination during handwriting-like movements. AB - The present study investigates intrinsic preferences and tendencies in coordination of the wrist and finger movements during handwriting-like tasks. Movement of the inkless pen tip in nine right-handed subjects was registered with a digitizer. One circle-drawing task and four line-drawing tasks were included in the experiment. The line-drawing task included: (1) drawing with the wrist only, (2) drawing with the fingers only, (3) an equivalent pattern consisting of the simultaneous flexion/extension of the wrist and fingers, and (4) a nonequivalent pattern in which wrist flexion was accompanied by finger extension and wrist extension was accompanied by finger flexion. Both the line and circle drawing were performed repetitively at four speed levels, ranging from slow to "as fast as possible" movements. The analysis of the line drawing revealed differential variability and temporal characteristics across the four movement patterns. While the equivalent pattern had characteristics of performance similar to those observed in the wrist-only and fingers-only pattern, the nonequivalent pattern was more variable and was executed slower when as fast as possible movement was required, compared to the other three patterns. The circle-drawing task also revealed intrinsic tendencies in coordination of the wrist and fingers. These tendencies were manifested by a spontaneous transition of the circular path of the pen tip to a tilted oval with increases in movement speed. The transition to the oval shape was accompanied by decreases in relative phase between the wrist and finger movements, whereas amplitudes of these movements were not affected by movement speed manipulations. The results suggest that subjects did not display a tendency to decrease the number of joints involved when executing the patterns that required simultaneous wrist and finger movements. Instead, there were preferences during these patterns to integrate wrist and finger movements with low relative phase. The findings are interpreted in terms of biomechanical constraints imposed on the wrist-finger linkage. This interpretation was further examined by testing two left-handed subjects. The data obtained showed symmetrical preferences in joint coordination. Collectively, the findings support a supposition that the shape of cursive letters may have been adjusted to the biomechanical structure of the hand to facilitate the motor act of handwriting. PMID- 11104135 TI - Paid attendant carers hold important and unexpected roles which contribute to the lives of people with brain injury. AB - OBJECTIVE: Paid attendant carers spend many hours assisting people with a brain injury. Despite this considerable responsibility, most carers receive little support or training and their roles are often ill-defined. This exploratory study set out to define the key roles of paid carers. METHOD: Ten semi-structured interviews were conducted. Perspectives were sought from 10 participants: five people with a traumatic brain injury and five paid carers. A computer software package, NUD*IST was used during analysis to help identifvy and categorize commonly recurring themes. RESULTS: Five major roles were identified: Attendant, Protector, Friend, Coach and Negotiator. Friendship was the most important aspect of the care relationship for three of the people with a brain injury, most of whom had lost their pre-injury friends and associates. Carers were required to negotiate frequently with clients and their families, and with other service providers. Sound communication skills were required. CONCLUSION: In addition to further research, industry guidelines are required which take account of the wider suite of roles fulfilled by paid carers, address training and support needs, and occupational health and safety issues. PMID- 11104136 TI - Behavioural differences between psychiatric patients with confirmed versus non confirmed traumatic brain injuries. AB - The relationship between history of Traumatic Brain Injury (TBI) and BDI-2 depression scores at admission and discharge from the hospital was assessed in acutely hospitalized psychiatric patients. The participants were assigned to three groups: (1) no reported history of TBI (n = 18), (2) reported but not confirmed TBI history (n = 13), and (3) reported and confirmed TBI history (n = 15). It was found that confirmed history of TBI was associated with elevated BDI 2 depression scores. In contrast, the non-confirmed TBI group was characterized by over-reporting of psychological distress, as measured by MMPI-2 validity indices, 100% prevalence of alcohol use history, and depression scores that were intermediate between the control and confirmed TBI groups. PMID- 11104137 TI - TBI knowledge and pragmatic assessment among Connecticut school speech-language pathologists. AB - School-based speech-language pathologists from Connecticut responded to a random survey which had a twofold purpose, (1) to replicate a previous conclusion that clinicians' specific experience with traumatic brain injury (TBI) influences their knowledge of this subject, and (2) to explore the topic of pragmatic assessment and whether it is also influenced by specific TBI experience. Results indicate that Connecticut school clinicians favourably regard both their own knowledge of TBI and the contemporary issue of pragmatic assessment. Connecticut clinicians also report a relatively low degree of prior TBI training and clinical experience. Clinicians' degree of TBI training and clinical experience did not appear directly related to their reported competence in TBI knowledge and pragmatic assessment. PMID- 11104138 TI - Cognitive and functional recovery at 6 and 12 months post-TBI. AB - Outcome studies examining recovery from traumatic brain injury (TBI) often fail to provide a clear understanding of the time course of cognitive, emotional, and behavioural recovery. The present study represents an effort to prospectively study individuals with TBI at fixed intervals, specifically 6 and 12 months post injury with a window of +/- 1 month. Seventy-two individuals with new-onset TBI underwent neuropsychological evaluation and clinical interview at 6 and 12 months post-injury. Results revealed significant improvements in cognitive abilities, including memory, processing speed, language abilities, and constructional skills. There were significant gains in community integration and involvement in productive activities, but limitations in driving activities remained. Although individuals with mild-moderate TBI performed better than individuals with severe TBI, both groups demonstrated equivalent rates of recovery across domains. The results of this study provide important information regarding the time course of TBI recovery. PMID- 11104139 TI - Stimulating consciousness and cognition following severe brain injury: a new potential clinical use for lamotrigine. AB - No medications clearly enhance consciousness or cognition following severe brain injury. This series (n = 13) suggests that lamotrigine may stimulate improvement of patients with impairment equivalent to level I-III on the Rancho Los Amigos Cognitive Scale. After a serendiptious clinical result, severely brain injured patients who were taking an anticonvulsant had an opportunity to start lamotrigine. This cohort had been transferred to this rehabilitation unit 14-304 (mean 73.9) days and started lamotrigine 20-310 (mean 87.5) days after acute brain injury. Compared to this unit's experience with patients with similar severe brain injuries, more patients (n = 10) were discharged to the conmmunity and fewer to skilled nursing facilities (n = 3) than were expected. This preliminary and provocative case series corresponds to basic science results, and further investigation of lamotrigine is warranted. PMID- 11104140 TI - Slow-to-recover severe traumatic brain injury: a review of outcomes and rehabilitation effectiveness. AB - Severe traumatic brain injury may result in very severe disability with prolonged recovery. Because of this slow recovery, survivors of severe traumatic brain injury may not be considered as good candidates for typical brain injury rehabilitation programmes and, thus, there is relatively little published information concerning the nature of this group. The recent literature regarding functional outcomes and the effectiveness of rehabilitation for this sub population of brain-injury survivors is reviewed and suggestions for further research are discussed. The existing evidence suggests that this emerging but important group of brain-injury survivors is capable of significant functional recovery over a period of months-to-years after injury, and that rehabilitation may serve to further ameliorate disability and reduce longterm costs of care. It is suggested that further research focus on delineating the nature of recovery in the slow-to-recover brain injury population, exploring the current prevalence of slow-to-recover brain injury survivors, and assessing the effectiveness of currently existing programmes specializing in rehabilitation of this type. PMID- 11104141 TI - Speech recognition training for enhancing written language generation by a traumatic brain injury survivor. AB - Impairments in motor functioning, language processing, and cognitive status may impact the written language performance of traumatic brain injury (TBI) survivors. One strategy to minimize the impact of these impairments is to use a speech recognition system. The purpose of this study was to explore the effect of mild dysarthria and mild cognitive-communication deficits secondary to TBI on a 19-year-old survivor's mastery and use of such a system-specifically, Dragon Naturally Speaking. Data included the % of the participant's words accurately perceived by the system over time, the participant's accuracy over time in using commands for navigation and error correction, and quantitative and qualitative changes in the participant's written texts generated with and without the use of the speech recognition system. Results showed that Dragon NaturallySpeaking was approximately 80% accurate in perceiving words spoken by the participant, and the participant quickly and easily mastered all navigation and error correction commands presented. Quantitatively, the participant produced a greater amount of text using traditional word processing and a standard keyboard than using the speech recognition system. Minimal qualitative differences appeared between writing samples. Discussion of factors that may have contributed to the obtained results and that may affect the generalization of the findings to other TBI survivors is provided. PMID- 11104142 TI - Histopathology of Kabatana arthuri (Microspora) infection in sutchi catfish, Panagasius sutchi. AB - The microsporidian Kabatana arthuri (Lom, Dykova et Shaharom, 1990) induced severe regressive changes in trunk muscles of Pangasius sutchi (Fowler) from Thailand. Necrotic changes developed in muscle fibres around the developmental stages and on the periphery of giant aggregates of spores. The main feature of the host defence reaction was the phagocytic activity of macrophages. Inflammatory reaction was only exceptionally observed. Spore-laden macrophages were found in various tissues and organs; their infiltration in epidermis including its outermost layers may effectively enhance the spread of infection while the hosts still live. PMID- 11104143 TI - Myxobolus intrachondrealis sp. n. (Myxosporea: Myxobolidae), a parasite of the gill cartilage of the common carp, Cyprinus carpio. AB - A species not identifiable with any of the about 23 Myxobolus species recorded from the common carp so far, was detected in the gills of one- and two-summer-old specimens of the common carp ((Cyprinus carpio L.) cultured in pond farms in Hungary. The strictly tissue-specific plasmodia of the parasite were located, surrounded by hyaline cartilage cells, in the chondrous substance of the terminal parts of the gill arches and in the cartilage structure ventrally connecting the gill arches. The spores of the parasite described as Myxobolus intrachondrealis sp. n. developed in globular or ellipsoidal plasmodia measuring 300-600 microm. By their elongated ellipsoidal shape and similarly elongated polar capsules the spores were well distinguishable from the hitherto described Myxobolus species parasitic in the common carp and also from the cartilage-parasitic Myxobolus species of other fishes. PMID- 11104144 TI - Dynamics of the IgG3 responses following immunisation of BALB/c mice with somatic and excretory/secretory antigens from various Trichinella species. AB - Comparison of the dynamics and antigen recognition profiles of IgG3 following immunisation with larval crude extracts (LCE) and excretory-secretory (ES) products from muscle larvae of different species of Trichinella (T. spiralis, T. nativa, T. britovi, T. nelsoni and genotype T6) was made in BALB/c mice. High levels of gG3 response were obtained in ELISA following immunisation with LCE from all species with maximum levels achieved between days 59 and 64 post immunisation (p.i.) and maintained up to the limit of the observation (day 164). Antigen recognition profiles as measured by western-blot showed dense and numerous bands in the range 45-64 kDa that were consistent from week 5th with variation in epitope recognition among the different species. Following immunisation with ES antigens a significant decrease in IgG3 response was observed for all species especially for T. nativa in comparison to LCE. Antigen recognition on ES antigens showed three main bands in the range of 45-60 kDa for all species excepting T. nelsoni and T. britovi where an additional band was also present. These results clearly show that IgG3 epitopes are more abundant in somatic (LCE) than in ES products of Trichinella muscle larvae and that quantitative as well as qualitative variations exist among different species. PMID- 11104145 TI - Morphometrics and seasonal occurrence of metacestodes of Neogryporhynchus cheilancristrotus (Cyclophyllidea: Dilepididae) in the blue bream (Abramis ballerus) from the Oder River (Germany/Poland). AB - From May 1993 to April 1995, the seasonal occurrence of metacestodes of Neogryporhynchus cheilancristrotus (Wedl, 1855) (Cyclophyllidea: Dilepididae) in its second intermediate host, the blue bream Abramis ballerus (L.) was studied monthly in the Oder River on the borders of Germany and Poland. Based on the parasite specimens found, detailed data on their morphometrics are presented. The metacestodes occurred in the blue bream intestine throughout the year (overall prevalence 27% and intensity 1-56 (mean 4.8) metacestodes per infected fish). Increased prevalences and mean intensities of infection were noted from March to June and November to December indicating that spring, late autumn and early winter are the main periods of new infections. PMID- 11104146 TI - Three new helminth species from two endemic plethodontid salamanders, Typhlomolge rathbuni and Eurycea nana, in central Texas. AB - Helminthological examination of two rare, endemic species of plethodontid salamanders, the Texas blind salamander (Typhlomolge rathbuni Stejneger) and the San Marcos dwarf salamander (Eurycea nana Bishop), from the subterranean waters and springs in San Marcos, Hays County, central Texas, USA revealed the presence of three new, previously undescribed species of intestinal helminths: Brachycoelium longleyi sp. n. (Trematoda) from T. rathbuni (type host) and E. nana, Dendronucleata americana sp. n. (Acanthocephala) from T. rathbuni, and Amphibiocapillaria texensis sp. n. (Nematoda) from T. rathbuni; nematode larvae probably belonging to the last named species were recorded from E. nana. Brachycoelium longleyi can be distinguished from all congeners primarily by its conspicuously small eggs among other features, whereas A. texensis differs from its closest congeneric species A. tritonispiunctati mainly in the structure of mature eggs and a markedly shorter spicule. Dendronucleata americana is the first species of the family Dendronucleatidae from the New World, differing from its Asian congeners mainly in the number and arrangement of proboscis hooks, number of giant hypodermic nuclei and in the position of testes. PMID- 11104147 TI - Digeneans and cestodes parasitic in the white-faced ibis Plegadis chihi (Aves: Threskiornithidae) from Argentina. AB - Some digeneans and cestodes parasitic in a population of the white-faced ibis Plegadis chihi (Vieillot) from Buenos Aires province, Argentina, are presented. The digeneans Dietziella egregia (Dietz, 1909), Patagifer bilobus (Rudolphi, 1819), Ascocotyle (Leighia) hadra Ostrowski de Nunez, 1992 and Posthodiplostomum nanum Dubois, 1937 from the intestine; Prosthogonimus ovatus (Rudolphi, 1803) from the cloaca; Athesmia heterolecithodes (Braun, 1899) from the bile ducts and the cestode Hymenolepis megalops (Nitzsch in Creplin, 1829) from the cloaca, were recorded. The discovery of D. egregia, P. ovatus, A. heterolecithodes and P. nanum constitute new host and/or new geographical records. Adults of A. (L.) hadra, previously described in experimental definitive hosts, are first reported from a naturally infected bird. Hymenolepis megalops, a cestode of Anseriformes is first reported from Ciconiiformes. PMID- 11104148 TI - Wardium paucispinosum sp. n. (Eucestoda: Hymenolepididae), parasite of Larus maculipennis (Aves: Laridae) in Mar del Plata, Argentina; with comments on Wardium semiductilis (Szidat, 1964) comb. n. AB - A new species Wardium paucispinosum (Eucestoda: Hymenolepididae) parasite from the intestine of Larus maculipennis (Lichtenstein) from Mar del Plata, Argentina is described. The distinctive features of the new species are: strobilar length 52.8 mm; 10 aploparaksoid rostellar hooks, 14 (12-17) microm long; ratio between cirrus pouch length and mature proglottid width (CPL/MPW) 0.38 (0.27-0.50); regular cylindrical evaginated cirrus, 90 x 10 microm, with distal end without spines and proximal and medium thirds covered with spines 7 microm long; simple tubular membranous vagina, 110 x 10 microm, without sclerotised portions and sphincters; eggs fusiform, 77 x 44 microm. Besides, Hymenolepis semidutctilis Szidat, 1964, from the intestine of Larus dominicanus and L. maculipennis from Santa Fe, Argentina is transferred to the genus Wardium Mayhew, 1925, based on the presence and shape of the rostellar hooks. PMID- 11104149 TI - Rhabdochona mexicana sp. n. (Nematoda: Rhabdochonidae) from the intestine of characid fishes in Mexico. AB - A new nematode, Rhabdochona mexicana sp. n., is described based on specimens recovered from the intestine of two species of fishes, Astyanax mexicanus (De Filippi) (type host) and Astyanax fasciatus (Cuvier) (Characidae: Characiformes) in central Mexico. This species is characterised by the following characters: 10 anteriorly directed teeth in the prostom, a larger (left) spicule which is slender in form with a small bifurcation at its distal tip covered by a cuticular membrane, a smaller (right) spicule without a barb at its distal tip, eggs bearing an irregular flock-like coating, and a conical tail without a cuticular spike (in both sexes). PMID- 11104150 TI - Development of Procamallanus saccobranchi (Nematoda: Camallanidae), a parasite of a freshwater fish in India. AB - The development of the nematode Procamallanus saccobranchi Karve, 1952, a parasite in the stomach of the fish Heteropneustes fossilis (Bloch), was studied in Mesocyclops crassus (Fischer) and Mesocyclops leuckarti (Claus). After being ingested by the copepods the nematode first-stage larvae penetrated into the haemocoel of the intermediate host; there they moulted twice (on days 3 and 5 p.i. at 28-30 degrees C) attaining the third, infective stage. The definitive host H. fossilis acquired infection by feeding on copepods harbouring infective stage larvae; in the stomach of this definitive host, the larvae were observed to undergo two more moults. The third moult occurred on day 13 p.i. and the fourth moult on day 38 p.i. and day 66 p.i. in "male" and "female" larvae, respectively. The larval stages, including the moulting forms are described and illustrated. PMID- 11104151 TI - Anuretes grandis sp. n., a caligid copepod (Siphonostomatoida) parasitic on Diagramma pictum (Pisces) in Taiwan, with discussion of Anuretes Heller, 1865. AB - A new species of caligid copepod (Siphonostomatoida), Anuretes grandis sp. n., parasitic on the painted sweetlips [Diagramma pictum (Thunberg)] in Taiwan is described. The new species is distinguished from its congeners by having: (1) free margin of cephalothorax not covering fourth pediger, (2) large genital complex longer than 2/3 of the cephalic shield, (3) no maxillary whip, (4) leg 3 with 9 setae on the terminal segment of exopod and 8 plumose setae on the terminal segment of endopod, and (5) armature of I,III on leg 4 exopod. Genus Anuretes Heller, 1865 is reviewed and redefined. Based on the new diagnosis three species (A. chelatus Prabha et Pillai, A. fedderni Price and A. parvulus Wilson) were transferred to Pseudanuretes, and two species (A. furcatus Capart and A. renalis Heegaard) were transferred to Lepeophtheirus. In addition, the following three species of caligids were transferred to Anuretes: Lepeophtheirus fallolunulus Lewis, Heniochophilus indicus Pillai, and Lepeophtheirus rotundigenitalis Prabha et Pillai. The latter is renamed Anuretes occultus nom. n. due to the homonym encountered through this transfer. "Anuretes plectorhynchi Yamaguti" reported by Prabha and Pillai (1986) is renamed Anuretes similis sp. n. and Anuretes yamagutii Prabha et Pillai is relegated to the synonym of Anuretes anomalus Pillai. A key to the 18 species of Anuretes is provided. PMID- 11104152 TI - A new species, Gnathia nkulu sp. n. (Crustacea: Isopoda: Gnathiidae) from southern Africa. AB - A new species, Gnathia nkulu sp. n. is described from material collected off the South African coast at 80-200m depth. It differs from the intertidal species Gnathia africana Barnard, 1914 in that the mediofrontal process is not deeply divided into two lobes, article 2 of the pylopod is rounded and small wart-like tubercles and long simple setae are present on both the cephalosome and pereon. PMID- 11104153 TI - Policy lessons from children's allowances for children's savings accounts. AB - This article examines the history and current structure of children's allowances around the world as well as the history of such allowances in the United States in an effort to provide the United States with a policy framework for children's savings accounts. The authors also provide policy direction for children's savings accounts. PMID- 11104154 TI - Outcome assessment: suggestions for agency practice. AB - Managed care proponents emphasize outcome research to determine the cost effectiveness of interventions, suggesting the use of multimeasurement procedures that will assess both specific and general effects of interventions. Although most agencies can perform outcome assessment and outcome management, they have difficulty conducting meaningful outcome research. This articles provides an introduction to outcome research; discusses the concepts of statistical significance, effect size, and the meaningfulness of effects; and emphasizes the need for client value-based judgments regarding the effectiveness of interventions. To encourage agencies to use client-completed outcome assessment procedures, the steps needs to effectively do so are outlined. PMID- 11104155 TI - Linking child maltreatment retrospectively to birth and home visit records: an initial examination. AB - The study reported here tested the feasibility of linking administrative datasets for evaluation of home visiting as a strategy to reduce the incidence of child abuse and neglect. It also examined associations between maternal and child attributes coded in the birth record, and subsequent child maltreatment. The results show that home visiting efforts in Vermont were, in general, targeted to the populations most at-risk for child maltreatment. Mother's educational attainment, in particular, was identified as a potent correlate of child maltreatment, a finding with implications for high school dropout prevention and recovery efforts. PMID- 11104156 TI - Concept mapping the needs of foster parents. AB - This study describes the needs of foster parents as perceived by the foster parents themselves. Forty-nine parents from 30 foster families were asked to describe their needs in response to the question: "What do you need to be a good foster parent?" Five themes were apparent in their answers: (1) good working relationships; (2) cultural sensitivity; (3) harmonious and stable family relationships; (4) adequate payment for services; and (5) a range of personality characteristics and parenting skills. These themes are consistent with the literature, with the notable exception of respite, a need identified in the literature but not by the sampled foster parents. The study results lend credibility to the existing literature on the needs of foster parents. PMID- 11104157 TI - The role of podocyte injury in the pathogenesis of focal segmental glomerulosclerosis. AB - The podocyte has diverse functions including glomerular filtration, biosynthesis and maintenance of the glomerular capillary architecture. It discharges these functions by virtue of a unique morphology, an intimate relationship with the capillary wall, and diverse synthetic and membrane specializations. Despite the complex role that it plays in glomerular function, the clinical manifestations of podocyte dysfunction are limited to proteinuria and renal insufficiency. Recurrent focal segmental glomerulosclerosis (FSGS) in renal transplants provides a unique opportunity to study the pathogenesis of FSGS in human beings, because the patients are monitored carefully to identify the onset of disease, the recurrence is presumed to have the same etiology as the primary disease, renal biopsy is a tool to study the pathogenesis of the lesion, and therapeutic intervention provides a mechanism to test pathogenesis. Pathologic studies have identified a proliferative lesion of the podocytes as the first sign of recurrent disease. The glomerular lesions evolve to form segmental glomerular scars with time. These findings and studies in experimental models of FSGS implicate podocyte injury in the pathogenesis of the recurrent disease. The cellular lesion (similar to the proliferative lesion of podocytes in recurrent FSGS), seen early in the course of primary FSGS suggests that the pathogenic sequence in recurrent FSGS also applies to primary FSGS. A soluble circulating factor that increases glomerular permeability and correlates with recurrence of FSGS has been identified in the pretransplant serum of patients with end-stage FSGS, but the mechanism of podocyte injury by this factor remains speculative. In any case, podocytes in the cellular lesion undergo morphologic changes and lose specialized functions seen in the normal mature cell, and these structural and functional abnormalities cause the permeability changes associated with proteinuria and destruction of glomerular filtration surface by scarring associated with loss of glomerular function. PMID- 11104158 TI - The treatment of primary focal segmental glomerulosclerosis. AB - Nephrotic patients with primary focal segmental glomerulosclerosis (FSGS) have a poor prognosis with 50% progressing to end stage renal disease (ESRD) over 3 to 8 years. The achievement of a remission in proteinuria has been associated with a significantly improved renal survival as compared to those patients not attaining a remission. Unfortunately, spontaneous remissions are rare in FSGS, and the response to therapy has historically been poor. Recent experience with more aggressive immunosuppressive therapy has lead to an increase in the remission rate for FSGS patients and given rise to optimism in the treatment of this glomerulopathy. PMID- 11104160 TI - Water: normal balance, hyponatremia, and hypernatremia. PMID- 11104159 TI - Treatment of idiopathic membranous nephropathy (IMN). AB - The best treatment of idiopathic membranous nephropathy remains an area of clinical controversy. At the moment only patients with nephrotic syndrome and/or declining renal function should be treated. Despite the negative trials, prolonged oral administration of corticosteroids alone may be a safe and an effective first-line treatment in nephrotic patients. If corticosteroids are ineffective, prolonged use of cyclosporine in low-doses can be recommended as an alternative treatment, that diminishes rapidly proteinuria in the majority of patients. Both treatments (intravenous high doses of corticosteroids and cyclosporine) may also be effective in patients with declining renal function. Because of their toxicity, the routine use of alkylating agents for patients with nephrotic syndrome is not justified. They may be retained for patients, in whom other treatment modalities have failed. Chlorambucil may be preferred over cyclophosphamide since it carries less toxicity. A lower dose of chlorambucil, than that usually suggested, for a short period of time seems to be prudent in an effort to avoid serious side-effects. PMID- 11104161 TI - Alport syndrome: abnormalities of type IV collagen genes and proteins. PMID- 11104162 TI - Renal pathology and ultrastructural findings in Alport's syndrome. AB - Morphological study of the kidney is generally the first step in the diagnosis of Alport's syndrome. Light microscopy study allows to suggest the diagnosis with the association of focal and segmental glomerulosclerosis, GBM anomalies when studied with silver staining, interstitial foam cells, and negative standard immunofluorescence study. GBM anomalies observed by electron microscopy are nearly specific with thickening splitting and fragmenting of the lamina densa. GBM anomalies are the consequence of a collagen IV disease. Thus, immunohistochemical results obtained with 6 different alpha(IV) are essential and allow to evaluate the mode of inheritance. Schematically, in the X dominant AS form, GBM, distal tubular BM and collecting duct BM do not express alpha3/alpha4, alpha5(IV). In the autosomic recessive AS form, collecting duct BM alone express alpha5(IV) without expression of alpha3(IV) and alpha5(IV) chains along the GBM and distal TBM. PMID- 11104163 TI - Contemporary diagnostic approach in Alport's syndrome. AB - Diagnosis of Alport's syndrome rests on clinical, pathological and genetic criteria. The clinical criteria include positive family history, persistent microhematuria and extrarenal abnormalities involving eyes and ears. Besides kidney biopsy, skin biopsy has emerged recently as an interesting diagnostic tool: the absence of staining for the alpha5 chain of type IV collagen in the epidermal basement membrane is highly specific of x-linked Alport's syndrome but its sensitivity is approximately 75%. Genetic diagnosis is based on direct identification of the mutation involved (positive in 50% of the families, after long and tedious search) or on linkage analysis. An integrated approach to diagnosis of Alport's syndrome (which encompasses various diseases characterized by different molecular defects) has been greatly facilitated in the last 10 years. PMID- 11104164 TI - Alport syndrome: renal transplantation and donor selection. PMID- 11104165 TI - Re-assessment of the renal hydrosaline dysfunction in rats bearing the Walker-256 tumor. AB - Sodium retention is a frequent effect of cancer in humans and animals, but the mechanism involved is not yet understood. In the Walker-256 tumor, sodium retention has been considered to be a late effect, secondary to retention in the tumor mass, and/or to adrenal hypertrophy. Normally, (in rats receiving single tumor implants), the development of different tumor systemic effects (TSE) such as anorexia, sodium and fluid retention, anemia and immune depression in rats is synchronous within each individual but random among individuals of a given group in which they appear 6-47 days, or more, after inoculation. In present study, multifocal simultaneous inoculations of tumor cells resulted in a rapid and synchronous initiation of TSE (in 3-4 days) in all rats when no local effects of metastases could mask the results. Sodium retention is a special tumor effect on Na+ balance and a very sensitive indicator of TSE initiation. The results from multifocally inoculated rats were averaged in each (sub-clinical (SubC), moderate (mCP) and grave (gCP)) clinical phase and compared to food-restricted (FR) rats. There was a significant, early decrease in urinary Na+ excretion during mCP when compared to SubC and FR. The renal sites involved were studied in awake, unrestrained animals by measuring of sodium, creatinine and lithium clearances. There was an initial increase in the absolute proximal (mCP: 21.4 +/- 1.7 vs FR: 16.0 +/- 1.1 mmol/min/100 g b.w., p < 0.05) and post-proximal (mCP: 11.1 +/- 0.4 vs FR: 6.6 +/- 0.4 mmol/min/100 g b.w., p < 0.001) Na+ reabsorption, which were partially compensated for by a rise in glomerular filtration rate (mCP: 213 +/- 11.4 vs FR: 162 +/- 10.2 microL/min/100 g b.w., p < 0.01) and by a fall in fractional proximal Na+ reabsorption (mCP: 62.8 +/- 2.2% vs FR: 70.1 +/- 1.7%, p < 0.05), despite significant Na+ and fluid retention. The terminal phase of illness (gCP) culminated with a marked decrease in creatinine clearance, suggesting a significant fall in renal function. The multifocal model proved useful for studying the initial TSE, since the sites of action would, in principle, be easy to identify. These observations may be of physiological interest since TSE may result from the abnormal production of physiological modulators. PMID- 11104166 TI - Effects of ethanol on plasma chloroquine, arginine vasopressin (AVP) concentrations and renal hydro-electrolyte handling in the rat. AB - Current evidence in literature suggests that acute effects of either chloroquine or ethanol on kidney function partly depend on influencing plasma concentrations of arginine vasopressin (AVP). Therefore, the goal of the current study was to explore the effects of chloroquine and/or various doses of ethanol on plasma AVP levels and associated effects on renal hydro-electrolyte handling. Separate groups of male anaesthetized Sprague-Dawley rats were placed on a continuous jugular infusion of 0.077 M NaCl at 150 microL/min(-1). After 3 h equilibration period, consecutive 20 min urine collections were made over the subsequent 4 h of 1 h control, 1 h 20 min treatment and 1 h 40 min postequilibration periods for measurements of urine flow and Na+ and K+ excretion rates. Chloroquine (0.06 microg/min(-1)) and/or ethanol at either 2.4, 6, 18 or 24 microg/min(-1) were added to the infusate during the treatment period. Trunk blood was collected after the treatment period from parallel groups for AVP, ethanol and chloroquine measurements. Vehicle infused animals acted as control animals. Infusion of ethanol at low rate of 2.4 microg/min(-1) increased Na+ excretion rates, but high rates (6-24 microg/min(-1)) did not elicit such effects. Plasma ethanol concentrations were undetectable following administration of ethanol alone at 2.4 or 6 microg/min(-1). However, ethanols were measurable following co-infusion of chloroquine and ethanol at 6 microg/min(-1) (6+/-1 mg/dL(-1)). Concurrent chloroquine and ethanol (24 microg/min(-1)) administration elevated plasma ethanol concentrations by 26% by comparison with that of ethanol alone at the same dose. Chloroquine and ethanol infusion at all doses significantly (p < 0.01) increased plasma chloroquine concentrations. Intravenous infusion of ethanol increased plasma AVP concentrations in a dose-dependent manner. The observations of this study suggest that acute ethanol increases plasma AVP levels in a dose dependent manner to affect hydro-electrolyte balance. PMID- 11104167 TI - Mildly elevated serum creatinine concentration correlates with the extent of coronary atherosclerosis. AB - Mildly elevated serum creatinine concentration was proposed to be a marker for increased risk of cardiovascular disease mortality. The aim of our prospective study was to evaluate a possible association between serum creatinine concentration and extent of coronary atherosclerosis together with conventional risk factors for atherosclerosis. Serum creatinine concentration was measured in 40 male patients without overt renal or ischemic renal disease (mean age 53 +/- 7 years) with stable or unstable angina undergoing routine coronary arteriography. The extent of coronary atherosclerosis was assessed by Gensini score. In univariate linear regression analysis Gensini score significantly correlated with serum concentrations of apolipoprotein AII (r=-0.3242, P<0.05) and creatinine (r=+0.3194, P<0.05), but not with serum concentrations of lipids (total, low- and high-density lipoprotein cholesterol, triglycerides), other apolipoproteins (apo B, apo AI), lipoprotein(a), autoantibodies to oxidatively modified low-density lipoprotein or age, weight and status of smoking, diabetes or hypertension. Multivariate linear regression analysis revealed that elevated serum creatinine was associated with the extent of coronary atherosclerosis independently of conventional risk factors for atherosclerosis. Mildly elevated serum creatinine was probably the marker of generalised vascular disease denoting early nephrovasculopathy in correlation with established atherosclerotic risk factors. PMID- 11104168 TI - Cyanate as a hemolytic factor. AB - During advanced renal failure, and particularly in patients with end-stage renal disease, proteins are carbamylated as a result of a reaction with cyanate. If the carbamylation of proteins adversely alters their biologic activities and structures, then urea must be viewed as an uremic toxin, rather than a surrogate. Therefore, we studied in this paper the role of cyanate as a hemolytic factor of erythrocytes to explain anemia observed in patients with high blood urea levels due to inadequate dialysis. Cyanate was added to make the final concentration 150, 300 and 600 nmol to each test tube containing the final concentration of 140 x 10(6) with human erythrocytes per mL of phosphate buffered saline solution. And they were incubated at 37 degrees C for 24, 48 and 72 hours. The extent of hemolysis and carbamylation was monitored. The levels of hemolysis and carbamylated erythrocytes increased as the time of exposure to cyanate increased from 24 hours to 72 hours. Furthermore, those increased as cyanate concentration in the incubation media rose from 150 nmol to 600 nmol. Cyanate can induce hemolysis by carbamylation of erythrocytes. Urea, through cyanate, may contribute to hemolysis. If one extrapolates these results to patients with end-stage renal disease, it may help explain one of the reasons for the anemia in patients with high levels of BUN due to inadequate dialysis. PMID- 11104169 TI - End-stage renal failure after irbesartan prescription in a diabetic patient with previously stable chronic renal insufficiency. AB - We report the case of a 78-year-old hypertensive diabetic patient without evidence of renal artery stenosis who had moderate chronic renal insufficiency, which had been stable for several years under low-dose captopril therapy, and who rapidly developed acute renal failure when irbesartan was prescribed. Unfortunately the medication was not stopped promptly and the patient never recovered his basal renal function and had to undergo chronic hemodialysis. This observation emphasizes the importance of a careful monitoring of renal function in patients receiving angiotensin II receptor antagonists. PMID- 11104170 TI - Continuous arteriovenous hemodiafiltration in the acute treatment of hyperammonaemia due to ornithine transcarbamylase deficiency. AB - BACKGROUND: Acute hyperammonemia caused by urea cycle disorder is a medical emergency for which immediate managements should be taken to minimize permanent brain damage. Among different enzyme defects, ornithine transcarbamylase deficiency (OTC) is one of the most common enzyme defect in urea cycle disorders. We utilized continuous renal replacement therapy techniques in the acute treatment of hyperammonemia due to ornithine transcarbamylase deficiency. PATIENTS AND METHODS: Three male neonates with elevated serum ammonia levels were shown, based on urine organic acid analysis and serum amino acid studies, to have OTC deficiency. Administration of sodium benzoate and sodium phenylacetate for activating alternative nitrogen waste pathway were used associated with protein restriction. Other modalities, including blood exchange transfusion, peritoneal dialysis, continuous renal replacement therapy were utilized in an attempt to lower serum ammonia concentration. RESULTS: We report the successful use of continuous arteriovenous hemofiltration (CAVH), continuous arteriovenous hemodialysis (CAVHD), continuous arteriovenous hemodiafiltration (CAVHDF) in the acute management of hyperammonemia due to OTC deficiency. We also compared the ammonia clearance between peritoneal dialysis, exchange transfusion, CAVH, CAVHD and CAVHDF. It demonstrated the evidence that CAVHDF provides the best ammonia clearance. CONCLUSION: Continuous renal replacement therapy including CAVH, CAVHD, and CAVHDF may be the alternative techniques for acute management of hyperammonemia in inborn error of metabolism when dialysis machine is not available. Our data suggests CAVHDF provides the best ammonia clearance. PMID- 11104171 TI - Scrotal pathology in pediatrics with sonographic imaging. AB - Scrotal pathology in pediatrics ranges from the more benign hydrocele and varicocele to acute testicular torsion requiring emergent surgery. Malignant testicular tumors can be insidious in onset or may present acutely when trauma brings a swollen scrotum to the patient's or physician's attention. Three common conditions can present as an acute scrotum, all of which can suggest testicular torsion clinically. Epididymitis often has a less acute onset than testicular torsion, although it does not always present with a straightforward diagnosis. Although it is generally an inflammatory process affecting males from 9 to 14 years of age, it can be seen in younger males with Henoch-Schonlein purpura and Kawasaki disease. Torsion of the appendix of the testis and epididymis can present acutely and mimic acute testicular torsion and generally occurs from 6 to 12 years of age. Testicular torsion itself usually occurs from 12 to 18 years of age and usually results from the anatomical "bell-and-clapper" deformity. Infarction of the testis can occur within as early as 4 to 6 hours after torsion, depending on the duration of symptoms and degree of twist of the spermatic cord. Advances in ultrasound technology in recent years have made ultrasound the examination of choice for imaging scrotal pathology, whether acute or chronic in nature. Doppler technology has tremendously increased the radiologist's ability to assess flow within the prepubertal testicle, which allows assessment of viability in the undescended testis as well as in neonatal torsion. The ability of ultrasound to diagnose the pathogenesis of the acute scrotum is unsurpassed by any other imaging modality. Ultrasound is a readily available, noninvasive examination without radiation that provides excellent anatomic detail and serves as an important and tremendously helpful imaging modality in all types of pediatric scrotal pathology. PMID- 11104172 TI - Calcium pyrophosphate dihydrate crystal deposition disease: imaging perspectives. AB - Calcium pyrophosphate dihydrate (CPPD) crystal deposition disease is widespread in elderly persons and has various clinical presentations that can be confounding to clinicians. It is characterized by acute, subacute, or chronic joint inflammation and deposition of CPPD crystals in hyaline cartilage, fibrocartilage, and other soft tissue structures. We have learned a great deal about imaging findings of CPPD crystal deposition disease. New facts about the disorder and clues to radiologic diagnosis continue to be revealed. This article will provide a review of imaging characteristics of this disease with emphasis on some recent findings. The nomenclature, epidemiology, classification, and pathophysiology will be explained. A discussion of the clinical manifestations and treatment will be followed by a review of the characteristic imaging features. PMID- 11104173 TI - Micro-affordance: the potentiation of components of action by seen objects. AB - It is suggested that seen objects potentiate a range of actions associated with them, irrespective of the intentions of the viewer. Evidence for this possibility is provided by the data from two experiments, both of which required a participant to make a binary motor response to an auditory stimulus. In the first experiment the response was a power or precision grip, which was performed whilst simultaneously viewing a real object which would normally be grasped using either a power or precision grip. A significant interaction of response and grip compatibility of the object was observed. Similar results were obtained in the second experiment when a wrist rotation of a given direction was used as a response, whilst viewing objects which would require wrist rotations if they were to be grasped. The effects of the seen objects on components of action are described as microaffordances which are said to be dispositional states of the viewer's nervous system. PMID- 11104174 TI - Iconic hand gestures and the predictability of words in context in spontaneous speech. AB - This study presents a series of empirical investigations to test a theory of speech production proposed by Butterworth and Hadar (1989; revised in Hadar & Butterworth, 1997) that iconic gestures have a functional role in lexical retrieval in spontaneous speech. Analysis 1 demonstrated that words which were totally unpredictable (as measured by the Shannon guessing technique) were more likely to occur after pauses than after fluent speech, in line with earlier findings. Analysis 2 demonstrated that iconic gestures were associated with words of lower transitional probability than words not associated with gesture, even when grammatical category was controlled. This therefore provided new supporting evidence for Butterworth and Hadar's claims that gestures' lexical affiliates are indeed unpredictable lexical items. However, Analysis 3 found that iconic gestures were not occasioned by lexical accessing difficulties because although gestures tended to occur with words of significantly lower transitional probability, these lower transitional probability words tended to be uttered quite fluently. Overall, therefore, this study provided little evidence for Butterworth and Hadar's theoretical claim that the main function of the iconic hand gestures that accompany spontaneous speech is to assist in the process of lexical access. Instead, such gestures are reconceptualized in terms of communicative function. PMID- 11104175 TI - When further learning fails: stability and change following repeated presentation of text. AB - Kay (1955) presented a text passage to participants on a weekly basis and found that most errors and omissions in recall persisted despite repeated re presentation of the text. Experiment I replicated and extended Kay's original research, demonstrating that after a first recall attempt there was very little evidence of further learning, whether measured in terms of further acquisition or error correction, over three more presentations of the text passages. Varying the schedule of presentations and tests had little effect, although performance was better when intermediate trials included both presentation and test than when only presentations or tests occurred. Experiment 2 explored whether this 'failure of further learning' effect could be overcome by (a) warning participants against basing their recall on their previous recall efforts and specifically directing them to base their recall upon the passages, (b) making each presentation more distinctive, or (c) drawing participants' attention to areas that would benefit from further learning by requiring them to tally their omissions and errors. The effect persisted in all cases. The findings have serious implications for the learning of text material. PMID- 11104176 TI - Running shared mental models as a distributed cognitive process. AB - Shared mental models theory normally takes the individual as its unit of analysis. This paper proposes a theoretical framework for studying shared mental models in which the model is considered to be distributed amongst the team. From this framework a cognitive process is predicted which describes how shared mental models are run. A team reasoning task requiring planning was used to illustrate this framework and test predictions derived from it. Two aspects of sharing mental models were studied; the degree of overlap between team members' mental models and the organization of the division of the model between team members. Experimental results showed that the cognitive processes used were distributed amongst the team and support was found for most, but not all, aspects of the proposed process of running a shared mental model. The organization of sharing was found to influence this process. PMID- 11104177 TI - The effect of explicit negatives and of different contrast classes on conditional syllogisms. AB - One experiment tested the effects of systematically negating the constituents of four fundamental inferences based on conditionals: Modus Ponens (i.e. inferences of the form: if p then q; p therefore q), Modus Tollens (if p then q; not-q therefore not-p); Affirmation of the Consequent (if p then q; q therefore p), and Denial of the Antecedent (if p then q; not-p therefore not-q). The latter two inferences are valid only for bi-conditionals (if, and only if, p then q). The participants drew their own conclusions from premises about letters and numbers on cards. We observed a significant effect of explicit negatives on Modus Tollens and Denial of the Antecedent problems: The inferences were drawn more often for conditionals that yield a negative conclusion (e.g. if p then not q; q therefore not p) than for conditionals that yield an affirmative conclusion (e.g. if not p then q; not q therefore p). Additionally, we observed a similar, but smaller effect on Affirmation of the Consequent problems. Furthermore, we observed a significant effect of the categorical premise (affirmative or negative), especially on Affirmation of the Consequent problems. Finally, we observed an effect of the magnitude of the contrast class. If the contrast is larger (a set of three, five or nine values), then the making of a double negation or the production of an affirmative conclusion is more difficult for Denial of the Antecedent inferences. We discussed the results in relation to a negative categorical premise bias, an affirmative premise bias, a negative conclusion bias and a double negation effect. PMID- 11104178 TI - Some shortcomings of long-term working memory. AB - Within the framework of their long term working memory theory, Ericsson and Kintsch (1995) propose that experts rapidly store information in long-term memory through two mechanisms: elaboration of long-term memory patterns and schemas and use of retrieval structures. They use chess players' memory as one of their most compelling sources of empirical evidence. In this paper, I show that evidence from chess memory, far from supporting their theory, limits its generality. Evidence from other domains reviewed by Ericsson and Kintsch, such as medical expertise, is not as strong as claimed, and sometimes contradicts the theory outright. I argue that Ericsson and Kintsch's concept of retrieval structure conflates three different types of memory structures that possess quite different properties. One of these types of structures--generic, general purpose retrieval structures--has a narrower use than proposed by Ericsson and Kintsch: it applies only in domains where there is a conscious, deliberate intent by individuals to improve their memory. Other mechanisms, including specific retrieval structures, exist that permit a rapid encoding into long-term memory under other circumstances. PMID- 11104179 TI - Shortcomings of generic retrieval structures with slots of the type that Gobet (1993) proposed and modelled. PMID- 11104180 TI - Causing harm and allowing harm: a study of beliefs in obsessional problems. AB - This study investigates two factors hypothesised as relevant to obsessional problems because of the way in which they influence decisions whether or not to act to prevent harm. These are (i) the way in which intrusive thoughts increase the internal awareness of harm, and confront the person with the possibility of taking action to prevent such harm and (ii) the extent to which there is some obvious external factor which increases awareness of the possibility of preventing harm. Obsessional patients, anxious and non-clinical controls completed a scale which systematically measured these factors across a wide range of situations. Results across all situations evaluated confirmed previous findings that both obsessionals and nonobsessionals were more likely to report acting to prevent harm when awareness of it is prompted by an intrusion than when it is not. It was also found that participants in all groups acted more 'obsessionally' when a scenario is described in ways which suggest that harm may be by 'commission' than when it is described in terms of an 'omission'. When scenarios about which each individual is most disturbed were analysed, anxious and non-clinical controls continued to differentially rate omission and commission situations; as predicted, this differential was not present for obsessional patients. It is concluded that obsessionals are more sensitive to omission than are nonobsessionals when considering scenarios about which they are concerned, and that this sensitivity is one factor influencing the decision whether to act to prevent harm. PMID- 11104181 TI - Effects of distraction and guided threat reappraisal on fear reduction during exposure-based treatments for specific fears. AB - To test predictions derived from the emotional processing theory of fear reduction, claustrophobics (N = 58) were randomized to one of four exposure conditions: (a) exposure with guided threat reappraisal, (b) exposure with a cognitive load distracter task, (c) exposure with both guided threat reappraisal and cognitive load distracter task and (d) exposure without guided threat reappraisal or cognitive load distracter task. We hypothesized that self-guided in vivo exposure would lead to less fear reduction if performed simultaneously with a cognitive load distracter task that severely taxes information processing resources. In contrast, we hypothesized that focusing on core threats during exposure would enhance fear reduction. The main findings were largely consistent with predictions. The cognitive load task (regardless of focus of available attention) had a detrimental effect on fear reduction, while guided threat reappraisal (regardless of cognitive load) had a facilitative effect. The greatest level of fear reduction and the lowest level of return of fear were observed in the exposure condition involving guided threat reappraisal without cognitive load. Clinical implications and directions for future research are discussed. PMID- 11104182 TI - Social anxiety and self-impression: cognitive preparation enhances the beneficial effects of video feedback following a stressful social task. AB - Negative and distorted images of the observable self are important in the development and maintenance of social phobia. Previous research has shown that video feedback has potential to correct the distorted self-perception [Rapee, R. M. & Hayman, K. (1996). The effects of video feedback on the self-evaluation of performance in socially anxious subjects. Behaviour Research and Therapy, 34, 315 322]. The present experiment investigated whether the construction of a self image prior to viewing the video may enhance the therapeutic effects of video feedback. High and low socially anxious individuals gave a speech and then viewed the video of their performance. Half of the sample were given cognitive preparation prior to viewing the video. Cognitive preparation involved asking participants to (1) predict in detail what they will see in the video, (2) form an image of themselves giving the speech and (3) watch the video as though they were watching a stranger. Participants who received cognitive preparation prior to the video feedback made higher ratings of their overall performance and of specific aspects of their performance compared to those who were not given cognitive preparation and compared to the same ratings made prior to the video feedback. These results suggest that the therapeutic effects of video feedback can be enhanced by careful cognitive preparation which maximises the perceived discrepancy between self and video images. PMID- 11104183 TI - Paradoxical breathlessness in asthma. AB - This study tested the hypothesis that breathlessness in asthma relates linearly to airway obstruction when situational, attentional and emotional influences are held constant via random presentation of different intensities of externally applied airflow obstruction. Adolescents with stable asthma and normal controls (n = 25 + 25) with lung functions of approximately 3.5 1 forced expiratory volume in 1 s (FEV1) breathed through a device which obstructed airflow with five stimulus intensities, analogous to a mean reduction in FEV1 of 8-66%. A session consisted of 10 blocks, each with presentation of five stimulus intensities plus the baseline resistance of the apparatus. Breathlessness was continuously reported by moving a lever along a 10-point scale. The mean breathlessness was computed per stimulus intensity. Lung function and anxiety were measured before and after the test. Participants with asthma, not controls, manifested a paradoxical response: they reported significantly more breathlessness, but undifferentially. One patient against 12 controls' reported consistently more breathlessness from baseline to severe obstruction. The hypothesis was only supported for controls. Breathlessness did not correlate with severity of asthma, lung function, duration of asthma, number of exacerbations over the last six months, age, sex or anxiety. It was concluded that the meaning of airflow obstruction in patients with asthma has changed and underlies their paradoxical responses, even when situational, attentional and emotional factors are controlled. PMID- 11104184 TI - Thought control strategies in schizophrenia: a comparison with non-patients. AB - This study tested the hypothesis that patients with a diagnosis of schizophrenia would report the use of different thought control strategies in comparison with non-patients. The Thought Control Questionnaire [TCQ; Wells, A. & Davies, M. (1994). The thought control questionnaire: a measure of individual differences in the control of unwanted thoughts. Behaviour Research and Therapy, 32, 871-878.] was administered to 22 patients who met DSM-IV criteria for schizophrenia and 22 non-patients. The results showed that schizophrenic patients used different thought control strategies (more worry and punishment-based strategies, less distraction-based strategies) in compairison with non-patients. The theoretical and clinical implications of these findings are discussed. PMID- 11104185 TI - A comparison of a brief and long version of the Situational Confidence Questionnaire. AB - Assessing confidence to resist drinking in high risk situations is an important part of behavioral treatments for alcohol problems. The present study assessed the reliability and validity of the original 100-item Situational Confidence Questionnaire (SCQ) and of an 8-item brief version (BSCQ). Using a visual analog scale, the BSCQ asked respondents to report their confidence to resist urges to drink heavily using the original eight SCQ subscales (e.g., pleasant times with others, social pressure). Data were collected from 120 adult problem drinkers who voluntarily entered an outpatient alcohol treatment program. The comparability of the BSCQ and the SCQ-100 was evaluated through intraclass correlations between the eight subscales and comparison of both instruments' underlying factor structures. Correlation coefficients for the subscales ranged from 0.56 to 0.80. Both instruments showed similar, but not identical factor structures. The present results indicate that the BSCQ provides comparable information to the SCQ-100. Limitations, as well as the clinical advantages, of using the BSCQ over longer versions are discussed. PMID- 11104186 TI - Current issues in Tourette syndrome. PMID- 11104187 TI - 5-HT2C receptor binding is increased in the substantia nigra pars reticulata in Parkinson's disease. AB - The involvement of abnormalities in nondopaminergic transmitter systems in Parkinson's disease is noteworthy because of the complications, such as dyskinesia, associated with long-term dopamine replacement therapy. The output regions of the basal ganglia, the substantia nigra pars reticulata, and the medial segment of the globus pallidus are overactive in Parkinson's disease but underactive in dyskinesia. 5-HT2C receptors are localized in these regions and are excitatory. A 5-HT2C receptor binding assay using [3H]-mesulergine and SB 200646A to define nonspecific binding was applied to postmortem tissue from patients with Parkinson's disease and from age-matched control patients. [3H] mesulergine binding was increased in the substantia nigra pars reticulata by 108% in Parkinson's disease tissue as compared with control tissue. These data suggest abnormalities of 5-HT2C transmission in the basal ganglia of patients with Parkinson's disease. PMID- 11104188 TI - Allelic association between the DRD2 TaqI A polymorphism and Parkinson's disease. AB - Genes encoding proteins involved in dopaminergic transmission have been of special interest during the evaluation of candidate genes for Parkinson's disease (PD). The dopamine D2 receptor gene (DRD2) is located on chromosome 11 q22-q23, and several polymorphisms of the gene have been described. The DRD2 gene has a TaqI A restriction fragment length polymorphism that is located in the untranslated region, approximately 10 kilobases from the 3' end of the gene. This polymorphism creates the two alleles A1 (variant) and A2. In this study, we investigated the hypothesis that a TaqI repeat fragment length polymorphism in the dopamine D2 receptor gene may be associated with PD. DNA from 72 patients with PD, classified as definite, probable, or atypical PD, and from 81 controls was genotyped by polymerase chain reaction and gel electrophoresis for the presence of the TaqI A1 polymorphism. The controls were matched for age, race, and geographic origin. There were significant differences in allelic distribution between the overall PD group and control groups (chi2 = 5.009, p = 0.025). When only patients with definite PD were considered an even more significant association was found (chi2 = 8.2121, p = 0.004). Among the overall PD group, the odds ratio for having the variant allele A1 was found to be 2.2 (95% confidence interval, [1.1; 4.4]), whereas it was calculated to be 3.0 (95% confidence interval, [1.4; 6.4]) when only patients with definite PD were considered. The current study showed that there is a statistically significant association between the DRD2 variant allele A1 and PD. This association is most pronounced in patients with definite PD and becomes nonsignificant when the clinical picture is classified as atypical PD. PMID- 11104189 TI - Linkage exclusion in French families with probable Parkinson' s disease. AB - We analyzed the segregation of genetic markers spanning chromosomal regions 2p13, 4p14-15, 4q21-23, 6q25-27, and 17q21 in nine French families affected by autosomal-dominant probable Parkinson's disease. These regions have been linked or associated with familial Parkinson's disease. Multipoint linkage and haplotype analyses excluded 2p13 and 4p14-15 loci in five of nine families. For three families, which were equivocal for two-point linkage at D4S405, the ubiquitin carboxy-terminal hydrolase gene (UCH-L1) was sequenced. In one family, a novel UCH-L1 M124L mutation that did not segregate with early-onset disease was identified. This suggests that rare variants in this gene may not be pathogenic. In seven of nine families, it could be inferred that affected individuals did not share 4q21-23 (alpha-synuclein) haplotypes. All families were unequivocally excluded by haplotype analysis from the parkin locus on 6q25-27. Finally, the 17q21 region was excluded in four of nine families, and no mutation in the tau gene was identified in the five remaining families. Findings from this study confirm genetic heterogeneity within familial parkinsonism. PMID- 11104190 TI - Kinetic tremor in a reach-to-grasp movement in Parkinson's disease. AB - The aim of this study was to quantify the tremor of the hand during a natural movement (kinetic tremor) in tremor-dominant parkinsonian patients (n = 13). We used a three-dimensional camera system to kinematically analyze rest and kinetic tremors in an unrestrained reach-to-grasp movement, and additional tremor recordings were performed under standard postural and rest conditions using electromyography and accelerometry. The standard analysis showed a highly synchronized tremor with similar frequencies at rest and in sustained postural tasks, with and without loading. A kinematic recording was used to compare rest and action conditions. A strong inhibition of the resting tremor was present at the onset of the movement and reached its peak during deceleration. A kinetic tremor of low amplitude was present in most of the parkinsonian patients, but its occurrence was confined mainly to the terminal periods of the movement. The frequency of kinetic tremor was significantly higher than that at rest, before the onset of the movement in Parkinson's disease, as determined by the kinematic analysis (mean, 5.5 Hz vs 6.5 Hz; p <0.01). Our results confirm similarities between the tremor at rest and the oscillations during a sustained postural task in classic parkinsonian tremor. In contrast to this stable tremor, which seems to be generated by basal ganglia oscillators, a different pathophysiology of oscillations during motion must be considered. The kinetic tremor is most likely related to an enhancement of the physiologic tremor in the terminal phase of the reach-to-grasp movement. PMID- 11104191 TI - Visuomotor skill learning on serial reaction time task in patients with early Parkinson's disease. AB - This study tested the role of basal ganglia in visuomotor skill learning. Thirty nine patients early in the course of Parkinson's disease (PD) and 30 patients after operation for an aneurysm of the anterior communicating artery (ACoA) were compared with 31 matched control subjects on a Serial Reaction Time test (SRTt). The patients with PD showed impaired visuomotor skill learning across the repeating blocks, in the presence of preserved declarative knowledge of embedded sequences, in contrast to the ACoA group in whom the reverse pattern was observed. The significant correlation in patients with PD between the standard neuropsychological and motor measures and the performance observed in the skill acquisition test, in the ACoA group and control subjects was not observed. The suggestion that this learning impairment could not be attributed to a motor deficit per se was also confirmed more directly for patients with PD. Accuracy of performance after the initial learning phase on the SRTt in patients with PD was associated predominantly with visual span capacity measures. Declarative knowledge of the embedded sequence of the SRTt was correlated to general cognitive and verbal span abilities in the PD group. The impairment observed in the PD group was not the result of a general decline in cognitive functioning, mood disturbances, or the severity of the motor symptoms. PMID- 11104192 TI - Detection and assessment of the severity of levodopa-induced dyskinesia in patients with Parkinson's disease by neural networks. AB - Levodopa-induced dyskinesias (LID) in Parkinson's disease (PD) have remained a clinical challenge. We evaluated the feasibility of neural networks to detect LID and to quantify their severity in 16 patients with PD at rest and during various activities of daily living. The movements of the patients were measured using four pairs of accelerometers mounted on the wrist, upper arm, trunk, and leg on the most affected side. Using parameters obtained from the accelerometer signals, neural networks were trained to detect and to classify LID corresponding to the modified Abnormal Involuntary Movement Scale. Important parameters for classification appeared to be the mean segment velocity and the cross-correlation between accelerometers on the arm, trunk, and leg. Neural networks were able to distinguish voluntary movements from LID and to assess the severity of LID in various activities. Based on the results in this study, we conclude that neural networks are a valid and reliable method to detect and to assess the severity of LID corresponding to the modified Abnormal Involuntary Movement Scale. PMID- 11104193 TI - How does Parkinson's disease affect quality of life? A comparison with quality of life in the general population. AB - BACKGROUND: Adequate provision of appropriate healthcare resources for patients with chronic neurologic disorders such as Parkinson's disease (PD) requires knowledge of the impact of the illness on their life. Quality of life (QoL) instruments measure the impact of the disease on general well-being that cannot be fully appreciated by clinical rating scales and allow comparisons with other patient groups and the general population. OBJECTIVES: To assess QoL in a population-based sample of patients with PD in different disease stages in comparison with the general population. METHODS: All 124 patients with PD seen in a population-based study on the prevalence of parkinsonism in the London area were asked to complete a QoL battery including the EuroQoL 5D (EQ-5D), the Medical Outcome Study Short Form (SF 36), and the 39-item Parkinson's Disease questionnaire (PDQ-39). An interview and complete neurologic examination, including the Hoehn and Yahr scale, were performed on the same day. The patients' QoL scores were compared with published QoL norms from the United Kingdom population. RESULTS: Quality of life, as measured by the PDQ-39, the EQ-5D, and the physical summary of the SF 36, deteriorated significantly with increasing disease severity. The greatest impairment was seen in the areas related to physical and social functioning, whereas reports of pain and poor emotional adjustment had similar prevalence in patients with PD and the general population. The impairment of QoL was seen in all age groups and was similar for men and women, but the differences between patients with PD and the general population were most marked in the younger patient groups. CONCLUSIONS: Parkinson's disease interferes with various aspects of QoL, particularly those related to physical and social functioning. This information should be taken into account in the clinical management and planning and allocation of healthcare resources to this population. PMID- 11104194 TI - Estrogen use among nursing home residents with a diagnosis of Parkinson's disease. AB - BACKGROUND: The role of estrogen in motor, cognitive, and behavioral functions in Parkinson's disease (PD) remains unclear. OBJECTIVES/METHODS: To determine differences in functional, cognitive, and behavioral patterns between estrogen users and non-users, we performed an observational study on 10,145 elderly women with PD using the Systematic Assessment in Geriatric drug use via Epidemiology (SAGE) database. The SAGE database consists of the Minimum Data Set (MDS), data collected on a cross-section of over 400,000 nursing home (NH) residents in five US states. Using a cross-sectional study design, we evaluated the demographics, physical and cognitive function, and mortality rates of women with PD who received estrogen (n = 195) versus those who did not receive estrogen (n = 9950). RESULTS: Independent of age, estrogen users were less cognitively impaired and more independent in their activities of daily living. Surprisingly, more estrogen users were depressed and more likely to be on an antidepressant than non-users. One-year death rates were comparable between estrogen users and non-users. CONCLUSION: This study supports the growing number of recent data suggesting estrogen's potential beneficial effects on PD motor and cognitive functions. PMID- 11104195 TI - Volume perception in parkinsonian speech. AB - This study contrasted the volume level of speech production with perceived volume. Fifteen idiopathic patients with Parkinson's disease who have hypophonic dysarthria and 15 healthy age- and sex-matched control subjects participated in this study. Testing took place in a sound-proof room. Ability to regulate volume was tested at three instructional levels of loudness: participants were given no instructions regarding volume (to elicit normal default volume) or were asked to read loudly or quietly. Two types of volume-perception judgments were made. First, an estimate of one's own volume, immediately after speaking (that is, immediate perception), and secondly, an estimation of reading volume after hearing one's own voice played back (that is, playback perception). These perceptual ratings were compared with actual speech volume produced in reading and conversation tasks. It was found that there was less of a difference between patients' production and perception of speech volume compared with that of the control subjects. While patients spoke more quietly than control subjects, they nevertheless perceived (immediate and playback perception) their own speech to be louder than did the control subjects. Patients overestimated the volume of their speech during both reading and conversation. The findings raise the question as to whether impaired speech production is driven by a basic perceptual fault or whether perception is abnormal as a consequence of impaired mechanisms involved in the generation of quiet speech. PMID- 11104196 TI - An investigation of the effects of subthalamic nucleus stimulation on acoustic measures of voice. AB - Seven patients with Parkinson's disease were implanted with deep brain stimulators to provide chronic electrical stimulation to the subthalamic nucleus bilaterally. Acoustic recordings and neurologic assessments were undertaken before surgery in the medication-off and medication-on conditions and after surgery with and without electrical stimulation in the medication-off and medication-on conditions. The data showed significant improvements in limb motor performance in response to medication before surgery and when the subthalamic nucleus was stimulated after surgery. Six months after surgery, there were small but statistically significant increases in sound pressure level and fundamental frequency variability in response to stimulation in the medication-on condition. No other statistically significant speech changes were measured. These findings are consistent with several other studies that have reported disparity between limb and speech improvements after neurosurgical intervention for Parkinson's disease. PMID- 11104197 TI - Dilated perivascular spaces in the putamen and pallidum in patients with Parkinson's disease scheduled for pallidotomy: a comparison between MRI findings and clinical symptoms and signs. AB - Forty patients with Parkinson's disease without mental deterioration who were scheduled for ventroposterolateral (VPL) pallidotomy were randomly selected for retrospective stereotactic magnetic resonance image (MRI) analysis. The preoperative MRI study was performed on a 1.0-T MRI machine with a three dimensional gradient-echo sequence. The MRI analysis was focused on five consecutive 2 mm thick axial slices without gap and parallel to the intercommissural line, starting from the level of the foramen of Monro and continuing in a ventral direction. Lacunar cysts of varying sizes (4-424 mm3) were seen at least in one hemisphere of all patients. The cysts had a clear dominance in posteroventral regions of the lateral-most pallidal regions (GP) and posteroventral regions of the putamen (PUT). No statistical correlation was found between the number or volume of the cysts and the sex, age, or duration of illness of the patients. Patients with predominantly left-sided clinical symptoms had a concentration of the cysts in the left GP, whereas those with predominantly right-sided symptoms had cysts significantly larger and more frequent in the right than the left GP. The cysts did not seem to affect the clinical outcome of pallidotomy. The authors think striatopallidal cysts develop from dilated perivascular spaces of the lenticulostriate vessels in the posteroventral regions of the GP and PUT. They are not pathognomonic for PD, but they may play some role in lateralization of the clinical symptoms in this classically asymmetric condition. PMID- 11104198 TI - Sociocultural differences in gait. AB - Transcultural differences in routine motor behavior and movement disorders have rarely been assessed. In the present study gait was studied in 47 healthy inhabitants of Tyrol living in rural or semi-urban (Innsbruck, Austria) settings and 43 healthy subjects residing in Berlin, Germany. In addition, gait was assessed in 23 patients in early stages of idiopathic Parkinson's disease (11 in Berlin, 12 in Innsbruck). Healthy subjects in Berlin showed faster gait velocity than their counterparts in Tyrol, and patients with Parkinson's disease were slightly slower than their respective healthy peers in both environments. Surprisingly, patients with Parkinson's disease from Berlin had significantly faster walking speeds than both patients and healthy control subjects from Tyrol. High gait tempo in parkinsonian patients from Berlin was characterized by fast step-rates and short strides. Differences in normal gait in different sociocultural settings are thus reflected in parkinsonian slowing of gait. PMID- 11104199 TI - The value of external anal sphincter electromyography for the diagnosis of multiple system atrophy. AB - OBJECTIVE: To assess the value of external anal sphincter electromyography (ASEMG) for the diagnosis of multiple system atrophy (MSA) among various causes of parkinsonism. ASEMG denervation profiles have previously been proposed as a diagnosis test for MSA, but their specificity is disputed. METHODS: ASEMG variables of 52 parkinsonian patients were analyzed according to the clinical diagnosis: MSA (n = 31) or no MSA (n = 21). Mean motor unit potential duration, percentage of polyphasicity, and the electromyographer's interpretation were analyzed according to clinical diagnosis, disease duration, genitourinary symptoms, gender, parity, and history of pelvic surgery. RESULTS: All patients with MSA showed ASEMG denervation. Mean motor unit potential duration was the most discriminant variable. No patient with MSA had a mean duration less than 12 ms and no patient without MSA had one greater than 16 ms. ASEMG discriminates between patients with MSA and Parkinson's disease. Using a threshold of 13 ms, the sensitivity was 80% and specificity was almost 70% (positive predictive value, 80%) for the diagnosis of MSA. Age, history of pelvic surgery, and to a lesser extent, female gender, parity, disease duration, and presence of urinary symptoms increased the likelihood of abnormal ASEMG. CONCLUSION: ASEMG was highly sensitive and rather specific for the diagnosis of MSA. PMID- 11104200 TI - [123I]beta-CIT SPECT in multiple system atrophy, progressive supranuclear palsy, and corticobasal degeneration. AB - Differentiation between Parkinson's disease (PD) and other neurodegenerative disorders with parkinsonian features, such as multiple system atrophy (MSA), progressive supranuclear palsy (PSP), and corticobasal degeneration (CBD), is difficult on clinical grounds. We studied the pattern of dopaminergic degeneration in 18 patients with probable MSA, 8 patients with PSP, 4 patients with CBD, 48 patients with PD and a similar degree of disability, and 14 control subjects performing single photon emission computed tomography (SPECT) 20 hours after injection of [123I]beta-CIT. Overall striatal binding was significantly reduced in MSA (-51% of normal mean), PSP (-60%), CBD (-35%), and PD (-58%), without overlap with control values. Asymmetry of striatal beta-CIT binding was significantly increased in patients with CBD and PD, as compared with control subjects. Although asymmetry seemed to be less pronounced in MSA and PSP than in PD, this was not statistically significant. Putamen-caudate nucleus ratios in patients with PD, MSA, and PSP, but not with CBD, were significantly reduced, as compared with control subjects. In conclusion, [123I]beta-CIT SPECT reliably enables the visualization of the presynaptic dopaminergic lesion in patients with MSA, PSP, and CBD. In most patients, however, it does not seem to be possible to differentiate these disorders from PD with this method. PMID- 11104201 TI - Markedly severe dystonia in Japanese encephalitis. AB - Encephalitis has been reported to be a rare cause of severe dystonia. We describe five patients with markedly severe dystonia from Japanese encephalitis. These patients with markedly severe dystonia were seen during the past 8 years as a subgroup of 50 patients with Japanese encephalitis. The diagnosis of markedly severe dystonia was based on increasingly frequent episodes of generalized dystonia with bulbar, respiratory, or metabolic derangement or leading to exhaustion or pain. The diagnosis of JE was based on clinicoradiologic features and a fourfold increase of hemagglutination-inhibiting antibody titers in paired serum. The outcome of the patients was defined as a good, partial, or poor recovery on the basis of 1-year clinical status. All the patients were males, and their ages ranged from 6 to 19 years. Movement disorders appeared 1 to 3 weeks after the illness as the level of consciousness started improving. During the next 1 to 4 weeks, patients began to experience markedly severe dystonia. It was associated with marked axial dystonia resulting in opisthotonus and retrocollis in five patients, jaw-opening dystonia in two patients, teeth clenching in one patient, and oculogyric crisis and neck deviation in another patient. The attacks of markedly severe dystonia lasted for 2 to 30 minutes and occurred as many as 20 to 30 times daily. Other developments included fixed limb dystonia in one patient, severe spasticity and rigidity in five patients, and focal muscle wasting in one patient. These patients had only a modest improvement after treatment. Markedly severe dystonia abated by 2 to 6 months in all the patients who were followed up. Cranial magnetic resonance imaging showed bilateral thalamic involvement in all patients, brainstem involvement in three patients, and basal ganglia involvement in two patients. At the 3-month follow-up, all patients had a poor outcome. At 1 year, one patient had a complete recovery; one had a partial recovery; and two were bedridden. It can be concluded that markedly severe dystonia is an important and serious sequela of Japanese encephalitis and may occur as the result of thalamus, midbrain, or basal ganglia involvement in various combinations. PMID- 11104202 TI - Implication of sensorimotor integration in the generation of periodic dystonic myoclonus in subacute sclerosing panencephalitis (SSPE). AB - To clarify the mechanism of periodic dystonic myoclonus in subacute sclerosing panencephalitis (SSPE), a 22-year-old patient with a clinical diagnosis of SSPE was electrophysiologically investigated. Involuntary movements consisted of generalized dystonic posturing which occurred quasiperiodically once every 4 to 8 seconds. Effects of sensory stimuli and voluntary movements were studied by means of polygraphic recording of surface electromyogram (EMG), scalp electroencephalogram (EEG), and magnetoencephalogram (MEG). EEG showed quasi periodic, generalized, transient complexes synchronous to each dystonic myoclonus, which were preceded by a slow negative EEG shift at the parietal region by approximately 5 seconds. Neither external stimuli nor self-paced movements alone influenced the periodicity of dystonic myoclonus or EEG complexes. In the reaction time task, however, the external stimuli given as an imperative cue to execute a motor task elicited dystonic myoclonus and generalized EEG complexes only if they were presented in the latter segment of the interval between the two successive EEG complexes while the slow negative EEG shift appeared. These findings suggest that EEG complexes and periodic movements spontaneously occur when cortical excitability reaches a certain critical level, but both phenomena are elicited even before if the sensory stimuli as an imperative signal requiring motor execution are presented. This finding most likely implies involvement of the sensorimotor integration mechanism in these periodic phenomena. PMID- 11104203 TI - Movement sequencing in children with Tourette's syndrome and attention deficit hyperactivity disorder. AB - Little research has been conducted to examine sequential motor functioning of children with Tourette's syndrome (TS) and attention deficit hyperactivity disorder (ADHD). Movement sequencing performance for a group of 12 children with TS and 24 children with ADHD children (12 taking and 12 not taking stimulant medication) and matched control subjects was examined using a serial choice reaction time button-pressing procedure. Aspects of movement preparation and execution were measured for 10 sequential two-way choice points along a response board that extinguished the illuminated target buttons at certain specific times contingent on the previous button press or release. The level of advance information was systematically reduced to provide three levels of reduction of advance information, including no reduction, moderate reduction, and high reduction. Children with TS and ADHD (unmedicated) showed larger increases in down time, reflecting aspects of movement preparation, for the highest level of reduction of advance information than did their respective control groups. These deficits are suggestive of underlying frontostriatal dysfunction. Furthermore, the normalization of performance for children with ADHD taking stimulant medication assists in the confirmation of the validity of such a clinical diagnosis and seems to add to the clinical efficacy of this form of treatment, which has previously been associated with improvements for predominantly attentional and inhibitory symptoms of ADHD. PMID- 11104204 TI - Response to levodopa challenge in Tourette syndrome. AB - A dopaminergic excess has been commonly postulated in the pathophysiology of tics, and an early report described acute worsening of tics with levodopa. However, dopamine agonists sometimes improve tics. We undertook this pilot study to determine whether people with tics could tolerate an acute dose of levodopa. Six adults with Tourette syndrome (TS) who had never been treated with neuroleptics took 150 mg levodopa by mouth under single-blind conditions after carbidopa pretreatment. All six subjects reported a decrease in self-rated tic severity (mean -40%, p <0.05), and three spontaneously asked if they could be prescribed levodopa for chronic treatment. Blinded videotape ratings of motor tic severity improved by 37% (p <0.02). A large, placebo-controlled trial will be required to confirm these findings, which raise important questions concerning the relationship of tic expression to dopaminergic activity. PMID- 11104205 TI - Niemann-Pick disease type C: two cases and an update. AB - We describe two patients with juvenile-onset Niemann-Pick disease type C (NPC) to illustrate the variable neurologic features of this condition. One presented with hypersplenism at age 10 and was misdiagnosed with Gaucher disease. He developed complex partial seizures in his teens but remained otherwise neurologically asymptomatic until his mid 30s. At age 45, he had mild dementia and dysarthria, vertical supranuclear ophthalmoplegia, axonal sensorimotor polyneuropathy, and cerebellar ataxia. The second patient presented with rapidly progressive dystonia at age 8, and mild hepatosplenomegaly, vertical supranuclear ophthalmoplegia, severe behavioral disorder, and dementia by age 14. The diagnosis of NPC was based on deficient cholesterol esterification and excessive lysosomal filipin staining in cultured skin fibroblasts. Current notions about diagnosis and pathogenesis of NPC are reviewed. PMID- 11104206 TI - Clinical report of three patients with hereditary hemochromatosis and movement disorders. AB - Neurologic manifestations are rarely described in hereditary hemochromatosis (HH). We describe three patients with HH and movement disorders. Patient 1, a 69 year-old man, had a 13-year history of disabling cerebellar syndrome, action tremor and myoclonus, and secondary dementia. Patient 2 was a 40-year-old man with a 9-year history of cerebellar syndrome, head and arm tremor, and cervical dystonia. Patient 3, a 75-year-old woman, had a 5-year history of rapidly disabling parkinsonian syndrome unresponsive to levodopa. The diagnosis of HH was established in the three patients by iron tests, evidence of a C282Y mutation, and, in two patients, by liver biopsy. High-field T2-weighted magnetic resonance imaging showed hyperintense signals in hemispheric white matter in patient 1, cerebellar atrophy in patient 2, and cerebellar and cerebral atrophy in patient 3 and no significant hypointense signals in the three patients. Phlebotomies and symptomatic treatments did not change the course of the disease. Our cases are compared with the five previously reported observations of HH with movement disorders. This rare association is one cause of the chronic acquired non Wilsonian hepatocerebral degeneration syndromes and represents a separate entity from aceruloplasminemia. The pathophysiologic mechanism of movement disorders in HH is unresolved. No hepatic insufficiency and portosystemic encephalopathy is evidenced in our cases, whereas the putative role of abnormal iron load remains to be ascertained. HH should be investigated more systematically in patients with movement disorders. PMID- 11104207 TI - Abnormalities of motor cortical excitability are not correlated with clinical features in atypical parkinsonism. AB - OBJECTIVE: To evaluate the specificity of motor cortical excitability changes in parkinsonian syndromes and their relevance to the pathophysiology of cardinal parkinsonian features. METHODS: Paired transcranial magnetic stimulation (TMS) was used to assess cortico-cortical inhibition (CCI) and facilitation (CCF) in the opponens pollicis muscle of patients with atypical, non-L-dopa- (LD) responsive parkinsonism. RESULTS: Compared with age-matched normal control subjects, CCI (interstimulus interval [ISI], 3 ms) was significantly reduced in 10 patients with predominantly parkinsonian multiple system atrophy (MSA-P) and in seven with vascular parkinsonism (VP), but not in four with predominantly cerebellar MSA. No significant change of CCF (ISI, 12 ms) was observed. No correlation was found between the amount of CCI and clinical status as evaluated with the Unified Parkinson's Disease Rating Scale (UPDRS). In 10 patients (5 MSA P, 5 VP), CCI was significantly increased by LD acute administration without concurrent clinical changes. CONCLUSIONS: Abnormalities of CCI are not peculiar to idiopathic Parkinson's disease and seem unlikely to underlie any specific parkinsonian feature, but rather possibly reflect a nonspecific imbalance of inhibitory and facilitatory motor cortical circuits. PMID- 11104208 TI - Sympathetic skin response and cardiovascular autonomic function tests in Parkinson's disease and multiple system atrophy with autonomic failure. AB - The relationship between sympathetic skin response (SSR) and cardiovascular autonomic function tests (CVTs) was investigated in 15 patients with idiopathic Parkinson's disease (PD), 15 patients with clinical evidence of multiple system atrophy (MSA) with autonomic failure, and in 15 healthy control subjects. SSR was elicited by electrical stimulation of the right and left median nerves and simultaneously recorded on the palms of both hands. CVTs included the following sympathetic and parasympathetic tests: orthostatism, head-up tilt, cold pressor test, deep breathing, Valsalva maneuver, and hyperventilation. The SSR was normal in all patients with PD and control subjects but was abnormal or absent in all patients with MSA. For patients with MSA, SSR latency was significantly longer and amplitude was significantly smaller than that of patients with PD and control subjects. For patients with PD, SSR did not differ from that of control subjects. In these patients, SSR latency was significantly longer and SSR amplitude was smaller when the side with more marked motor symptoms was stimulated, both ipsilaterally and contralaterally to the side of stimulation. A statistically significant difference in SSR latencies and amplitudes was found between patients with PD and control subjects only when motor asymmetries were considered. CVTs showed severe sympathetic and parasympathetic hypofunction in patients with MSA, but not in patients with PD or control subjects. No correlation was found between SSR and CVTs that assess sympathetic function in patients and control subjects. SSR is indicated as an additional test for the evaluation of sympathetic degeneration in patients with MSA. PMID- 11104209 TI - The validity of the Beck Depression Inventory as a screening and diagnostic instrument for depression in patients with Parkinson's disease. AB - PURPOSE: To evaluate the validity of the Beck Depression Inventory (BDI) as a screening and diagnostic scale for depression in Parkinson's disease (PD). PATIENTS AND METHODS: Fifty-three nondemented patients with PD were diagnosed according to a standardized protocol consisting of the depression module of the Structured Clinical Interview for DSM axis I disorders (SCID) and the BDI. A "receiver operating characteristics" (ROC) curve was obtained and the sensitivity, specificity, positive and negative predictive values (PPV and NPV, respectively) were calculated for different cut-off points of the BDI. RESULTS: Maximum discrimination was obtained with a cut-off score of 13/14. High sensitivity and NPV were obtained with cut-off scores of 8/9 or lower; a high specificity and PPV were obtained with cut-off scores of 16/17 or higher. The area under the ROC curve was 85.67%. CONCLUSION: A single cut-off score on the BDI to distinguish nondepressed from depressed patients with PD is not feasible. If one accepts the low specificity, then the BDI can be used as a valid screening instrument for depression in PD with a cut-off of 8/9. With a cut-off score of 16/17, it can be used as a diagnostic scale, at the cost of a low sensitivity. The use of diagnostic criteria for depression remains necessary. PMID- 11104210 TI - S-Adenosyl-Methionine improves depression in patients with Parkinson's disease in an open-label clinical trial. AB - We report a pilot study of S-adenosyl-methionine (SAM) in 13 depressed patients with Parkinson's disease. All patients had been previously treated with other antidepressant agents and had no significant benefit or had intolerable side effects. SAM was administered in doses of 800 to 3600 mg per day for a period of 10 weeks. Eleven patients completed the study, and 10 had at least a 50% improvement on the 17-point Hamilton Depression Scale (HDS). One patient did not improve. Two patients prematurely terminated participation in the study because of increased anxiety. One patient experienced mild nausea, and another two patients developed mild diarrhea, which resolved spontaneously. The mean HDS score before treatment was 27.09 +/- 6.04 (mean +/- standard deviation) and was 9.55 +/- 7.29 after SAM treatment (p < 0.0001). Although uncontrolled and preliminary, this study suggests that SAM is well tolerated and may be a safe and effective alternative to the antidepressant agents currently used in patients with Parkinson's disease. PMID- 11104211 TI - Risperidone in the treatment of dopamine-induced psychosis in Parkinson's disease: an open pilot trial. AB - PURPOSE: To evaluate the safety and efficacy of risperidone in patients with Parkinson's disease (PD) who are experiencing significant dopamine-induced psychosis. PATIENTS AND METHODS: Seventeen patients (median age, 72 yrs) participated in this 12-week, open pilot study receiving 0.5 to 3 mg oral risperidone per day. Maintenance antiparkinsonian medication was continued throughout, although psychotropic medication was discontinued. EFFICACY RESULTS: Risperidone produced a substantial improvement in psychotic symptoms, shown on the mean total positive subscale score on the Positive and Negative Syndrome Scale (PANSS) by a 30% improvement (-3.1 decrease) after 1 week and a 66% improvement (-6.8 decrease) at end point. This improvement was most evident in the items delusions, hallucinatory behavior, and suspiciousness/persecution. Risperidone also achieved significant improvement from baseline in Clinical Global Impression (CGI)-severity and CGI-improvement (p < 0.001, Page test). Risperidone treatment did not adversely affect symptoms specific to Parkinson's disease, as assessed by the Unified Parkinson's Disease Rating Scale (UPDRS). SAFETY RESULTS: Sixteen patients reported at least one adverse event, but only two patients withdrew as a result of adverse events. No significant changes or clinically relevant abnormalities were observed in laboratory parameters or vital signs. CONCLUSION: Short-term use of risperidone (mean dosage, 1.1 mg per day) improves the psychopathology of patients with PD who have dopamine-induced psychosis without adversely affecting the symptoms of PD. Higher doses and long term use were not addressed in this study and may be precluded by extrapyramidal side effects. PMID- 11104212 TI - The DYT1 GAG deletion is infrequent in sporadic and familial writer' s cramp. AB - A 3-base pair (GAG) deletion in the DYT1 gene has recently been found to be responsible for most cases of early-onset primary generalized dystonia. In some cases, this mutation has been associated with writer's cramp. To determine the frequency of this mutation in a larger series of patients, we examined 44 index patients with sporadic or familial (seven patients) writer's cramp for the presence of the DYT1 GAG deletion, including eight patients with segmental dystonia involving at least one upper limb. We found the mutation in none of these index patients, which confirms that isolated writer's cramp is only in rare cases a phenotypic manifestation of this mutation, even if a positive family history of writer's cramp is present. PMID- 11104214 TI - Internal globus pallidotomy in dystonia secondary to Huntington's disease. AB - INTRODUCTION AND METHOD: The prototypic motor feature of Huntington's disease (HD) is chorea, but parkinsonism and involuntary movements such as dystonia and myoclonus can also be present. Pallidotomy has been shown to be an effective treatment for medically refractory Parkinson's disease (PD). We performed bilateral microelectrode guided-stereotactic pallidotomies targeted at globus pallidum internus (GPi) to treat a 13-year-old patient diagnosed with Westphal variant of HD with intractable generalized dystonia and parkinsonism. RESULTS: Intraoperative microelectrode recordings of GPi cells showed a relatively low firing rate, 29 +/- 14 Hz, with most neurons showing pauses. Acutely, after surgery, limb dystonia mildly improved but trunk dystonia persisted. Postoperative follow up 3 months later showed minimal clinical improvement in dystonic features with marked worsening of spasticity. CONCLUSION: In our case, bilateral pallidotomy produced modest palliative functional improvement in dystonic features. Cellular firing patterns were markedly different than in PD and were similar to those found in dystonia. PMID- 11104213 TI - Intrathecal baclofen for dystonia: benefits and complications during six years of experience. AB - Fourteen patients with primary or secondary dystonia received intrathecal baclofen (ITB) through an implanted pump following a trial dose. Patients were selected for ITB trial if they had clinically unsatisfactory responses to oral antidystonic medications, including oral baclofen. Patients were rated using the Burke-Fahn-Marsden rating scale by a blinded rater after the dose of ITB was optimized. Five patients experienced improvement in symptoms as determined by a change in rating scale scores, although only two had a clear clinical benefit. Etiology of dystonia did not determine the efficacy of ITB therapy, as benefit or failure was seen in both primary and secondary dystonia. PMID- 11104215 TI - Patterns of response to acute naloxone infusion in Tourette's syndrome. AB - The purpose of this study was to replicate findings from an earlier pilot study in which we found a dose-related effect of the opioid antagonist naloxone on tic behavior in patients with Tourette's syndrome (TS). Fifteen subjects with TS were challenged with randomized doses (30 and 300 microg/kg) of naloxone at 3-day intervals. Videotaped recordings of tic behavior were counted in a "blind" fashion. We found that naloxone had opposite effects on tics at different dosages. The low dose caused a significant decrease in tics, whereas the high dose caused a significant increase in tics. Therefore, activity at opioid receptors appears to influence the expression of TS, and the difference in response to naloxone in TS subjects may be based on a dose-response effect. PMID- 11104216 TI - Accuracy of clinical diagnostic criteria for Friedreich's ataxia. AB - The accuracy of the diagnostic criteria for Friedreich's ataxia proposed by Harding and by the Quebec Cooperative Study on Friedreich's Ataxia was studied in 142 patients with progressive unremitting ataxia of autosomal recessive inheritance or sporadic occurrence. Eighty-eight patients received the molecular diagnosis of Friedreich's ataxia. Traditional diagnostic criteria are characterized by high specificity, but they yield a high number of false-negative diagnoses. We suggest three levels of diagnostic certainty: (1) possible Friedreich's ataxia, defined as sporadic or recessive progressive ataxia with (a) lower limb areflexia and dysarthria, Babinski sign, or electrocardiographic repolarization abnormalities, or (b) with lower limb retained reflexes and electrocardiographic repolarization abnormalities (95% sensitivity and 88% positive predictive value); (2) probable Friedreich's ataxia as defined by Harding's criteria (63% sensitivity and 96% positive predictive value) or by Quebec Cooperative Study on Friedreich's Ataxia criteria (63% sensitivity and 98% positive predictive value); (3) definite diagnosis, molecularly confirmed. PMID- 11104217 TI - Bilateral painful hand-moving fingers: electrophysiological assessment of the central nervous system oscillator. AB - We describe a 35-year-old woman who presented with the syndrome of painful hand moving fingers on the right side. Eight months later, she developed similar finger movements and hand discomfort on the left side. She had a history of hand trauma and recurrent shoulder dislocation on the right side. Kinesiologic electromyography suggested a common central oscillator for finger movements in both hands. Electrophysiological assessment of spinal alpha motor neuron excitability, reciprocal inhibition, and Renshaw cell inhibition failed to show any abnormalities. Somatosensory evoked potential test showed marked attenuation of N20 potential recorded from the left somatosensory cortex; paired transcortical magnetic stimulation of the left motor cortex suggested failure of cortical facilitation. The data suggest that the central oscillator responsible for finger movements is located above the spinal cord level in this patient. PMID- 11104218 TI - Nocturnal frontal lobe epilepsy: a wide spectrum of seizures. PMID- 11104219 TI - No association between paraoxonase 1 (PON1) gene polymorphisms and susceptibility to Parkinson's disease in a Chinese population. PMID- 11104220 TI - Lack of association between cytochrome P450 2E1 gene polymorphisms and Parkinson's disease in a Chinese population. PMID- 11104221 TI - Modafinil treatment of pramipexole-associated somnolence. PMID- 11104222 TI - Neuroacanthocytosis presenting as parkinsonism. PMID- 11104223 TI - Dopa-resistant parkinsonism, oculomotor disturbances, chorea, mirror movements, dyspraxia, and dementia: the expanding clinical spectrum of hypoparathyroidism. A case report. PMID- 11104224 TI - Nicotine-sensitive writer's cramp. PMID- 11104225 TI - Can intravenous immunoglobulin improve antibody-mediated botulinum toxin therapy failure? PMID- 11104226 TI - Treatment of persistent hemiballism with botulinum toxin type A. PMID- 11104227 TI - McLeod syndrome and neuroacanthocytosis with a novel mutation in the XK gene. PMID- 11104228 TI - Chorea in new variant Creutzfeldt-Jacob disease. PMID- 11104229 TI - Primary anticholinergic-responsive Pisa syndrome. PMID- 11104230 TI - Radiation-induced 'Meige syndrome'. PMID- 11104231 TI - Pseudoathetosis in a patient with cervical myelitis: neurophysiologic and functional MRI studies. PMID- 11104232 TI - Hyperekplexia in the first year of life. PMID- 11104233 TI - Persistence of rhythmic movement disorder beyond childhood: a videotape demonstration. PMID- 11104234 TI - Friedreich ataxia associated with dystonic head tremor provoked by prolonged exercise. PMID- 11104235 TI - Atypical antipsychotics: clozapine-related cardiac complications. PMID- 11104236 TI - Acute medicine: the physician's role. A working party report of the Federation of Royal Colleges of Physicians of the United Kingdom. PMID- 11104237 TI - A national census of ambulance response times to emergency calls in Ireland. AB - BACKGROUND: Equity of access to appropriate pre-hospital emergency care is a core principle underlying an effective ambulance service. Care must be provided within a timeframe in which it is likely to be effective. A national census of response times to emergency and urgent calls in statutory ambulance services in Ireland was undertaken to assess current service provision. METHODS: A prospective census of response times to all emergency and urgent calls was carried out in the nine ambulance services in the country over a period of one week. The times for call receipt, activation, arrival at and departure from scene and arrival at hospital were analysed. Crew type, location of call and distance from ambulance base were detailed. The type of incident leading to the call was recorded but no further clinical information was gathered. Results-2426 emergency calls were received by the services during the week. Fourteen per cent took five minutes or longer to activate (range 5-33%). Thirty eight per cent of emergencies received a response within nine minutes (range 10-47%). Only 4.5% of emergency calls originating greater than five miles from an ambulance station were responded to within nine minutes (range 0-10%). Median patient care times for "on call" crews were three times longer than "on duty" crews. CONCLUSION: Without prioritized use of available resources, inappropriately delayed responses to critical incidents will continue. Recommendations are made to improve the effectiveness of emergency medical service utilisation. PMID- 11104238 TI - The laws of violence. AB - Working in an accident and emergency (A&E) department inevitably involves dealing with the consequences of violence, and a knowledge of the laws of violence is a useful adjunct to the clinical practice of A&E medicine. The police and the Crown Prosecution Service decide whether or not to charge a suspect, and which charge is appropriate. All criminal offences are initially considered in the magistrates' court but the more serious offences may be committed to crown court. Specific offences include common assault, actual bodily harm, grievous bodily harm, and grievous bodily harm with intent. If the defendant is found guilty, an appropriate sentence is imposed. PMID- 11104239 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. Vomiting and serious head injury in children. PMID- 11104240 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. Low molecular weight heparin or unfractionated heparin in the treatment of patients with uncomplicated deep vein thrombosis. PMID- 11104241 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. Outpatient treatment for patients with uncomplicated above knee deep vein thrombosis. PMID- 11104242 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. SimpliRed D-dimer assay in suspected pulmonary embolus. PMID- 11104243 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. Elastic compression stockings and the risk of post-thrombotic syndrome in patients with symptomatic proximal vein thrombosis. PMID- 11104244 TI - Towards evidence based emergency medicine: best BETs from the Manchester Royal Infirmary. Prior injection of local anaesthetic and the pain and success of intravenous cannulation. PMID- 11104245 TI - Article 4. An introduction to estimation--1. Starting from Z. PMID- 11104246 TI - Phrenic nerve injury following blunt trauma. AB - Phrenic nerve trauma in the absence of direct injury is unusual and may present diagnostic difficulty. Diaphragmatic paralysis resulting from phrenic nerve injury may closely mimic diaphragmatic rupture. This case highlights the value of magnetic resonance imaging in establishing diaphragmatic integrity and of ultrasonographic assessment during respiratory excursion in confirming diaphragmatic paralysis. In cases of non-contact injury involving torsional injury to the neck, an index of clinical awareness may help to establish the diagnosis of phrenic nerve trauma. PMID- 11104247 TI - Non-penetrating chest blows and sudden death in the young. AB - Sudden death in the young after low energy anterior chest wall impact is an under recognised phenomenon in this country. Review of the literature yields several American references to commotio cordis, mainly in the context of sporting events. Two cases are reported of sudden death in young men as a result of blunt impact anterior chest wall trauma. It is suggested that these cases draw attention to a lethal condition of which many practitioners are unaware. PMID- 11104248 TI - Extracorporeal rewarming in a severely hypothermic patient using venovenous haemofiltration in the accident and emergency department. AB - Severe hypothermia is a medical emergency and requires active and occasionally rapid core rewarming to prevent cardiac arrhythmias and death. In the accident and emergency department rewarming is often limited to warmed intravenous fluids, heated blankets, gastric and bladder lavage. Extracorporeal methods, which rewarm core blood directly, for example haemodialysis and cardiopulmonary bypass, require expertise and equipment not always found in a district general hospital. Venovenous haemofiltration is now commonly found in district general hospitals around the country and can be used safely for core rewarming. A case is reported of a severely hypothermic elderly patient successfully rewarmed using venovenous haemofiltration, in an accident and emergency department, when other conventional methods had failed. PMID- 11104249 TI - Snap without crackle or pop: a rude awakening. A case history of penile fracture. AB - Penile fracture is a rare but worrying condition. The presentation to accident and emergency or primary care should not present difficulty in diagnosis but may cause concern with regard to initial treatment and definitive management. Emergency admission to a urologist is mandatory. PMID- 11104250 TI - Brugada syndrome--the missed epidemic. AB - About 10-20% of patients dying suddenly or resuscitated from ventricular fibrillation do not have demonstrable heart disease. These people are often young and tragically in some cases sudden death is the first and only clinical event. One of the three main electrophysiological diagnoses to be considered in these situations is the Brugada syndrome. A case of Brugada syndrome is described, together with an example of the classic electrocardiographic manifestations and a discussion of the possible aetiology, diagnosis and management of this condition. PMID- 11104251 TI - Posterior dislocation of hip in adolescents attributable to casual rugby. PMID- 11104252 TI - Retropharyngeal haematoma after blunt trauma. PMID- 11104253 TI - Radiology case report: a nasty orbital abscess. PMID- 11104254 TI - Punchbag machine injuries in a nightclub. PMID- 11104255 TI - Does intraosseous have to mean intramedullary? PMID- 11104256 TI - Modeling of pH and acidity for industrial cheese production. AB - A three-layer feedforward neural network was successfully used to model and predict the pH of cheese curd at various stages during the cheese-making process. An extended database, containing more than 1800 vats over 3 yr of production of Cheddar cheese with eight different starters, from a large cheese plant was used for model development and parameter estimation. Neural network models were developed with inputs selected among 33 quantitative and qualitative process variables for final pH of cheese, pH at cutting, and acidity at whey drawing-off and at pressing. In all cases, very high correlation coefficients, ranging from 0.853 to 0.926, were obtained with the validation data. A sensitivity analysis of neural network models allowed the relative importance of each input process variable to be identified. The sensitivity analysis in conjunction with a priori knowledge permitted a significant reduction in the size of the model input vector. A neural network model using only nine input process variables was able to predict the final pH of cheese with the same accuracy as for the complete model with 33 original input variables. This significant decrease in the size of neural networks is important for applications of process control in cheese manufacturing. PMID- 11104257 TI - Viscoelastic properties of oil-water interfaces covered by bovine beta-casein tryptic peptides. AB - A combination of proteolysis and dilational rheology has been used to study the behavior of films of beta-casein (beta-CN) and of peptides spread at the oil water interface. Identification of the peptides produced by trypsin hydrolysis of beta-CN in emulsion at 37 degrees C provided information on the structure of beta CN adsorbed at the oil-water interface. Good interface properties were observed for beta-CN or its peptides, probably because of the amphipathic nature of beta CN or a synergistic effect between hydrophilic and hydrophobic peptides. Remarkable surface activity was found for the amphipathic peptide beta-CN (f114 169). Rheological studies had shown that interface films made with peptide fractions or with beta-CN were elastic rather than viscous. Film made with the purified peptide beta-CN (f114-169) was merely elastic at the triolein-water interface. A decrease of the viscoelastic modulus was observed for aging beta-CN film but not for aging peptide films; The beta-CN decrease was related to the flexibility of its structure. When the interface is increased by the dilation of an aqueous droplet plunged into oil, beta-CN may expose new polypeptide trains to cover the increased interface, unlike peptides with simpler structures. PMID- 11104258 TI - Technical note: production of butter with enhanced conjugated linoleic acid for use in biomedical studies with animal models. AB - Cancer models utilize massive doses of carcinogen so that investigations of anticarcinogenic effects require equally large doses. Conjugated linoleic acids (CLA), predominately consumed in dairy products, are thought to be anticarcinogenic. Our objective was to naturally produce a CLA-enhanced butter for use in biomedical studies with animal models. To do this, we fed cows a low forage diet supplemented with sun-flower oil. This resulted in increases in content of CLA of milk fat, but the markedly elevated concentrations were transient and declined over a 3-wk period. By collecting milk fat over the first few days on the diet (d 7 to 10) and selecting cows with the greatest CLA concentrations, we were able to produce a butter in which CLA content was enhanced sevenfold over control butter (41 vs. 5 mg/g of fatty acids) and the cis 9, trans-11 isomer predominated (91%). Thus, butter produced by this method can be used to investigate the preventive role of CLA in natural foods with biomedical models of different types of cancer. Furthermore, the butter allows examination of the other beneficial health effects of CLA reported with animal models. PMID- 11104259 TI - Influence of high-oryzanol rice bran oil on the oxidative stability of whole milk powder. AB - The effect of high oryzanol rice bran oil (RBO) on the oxidative stability of low heat and high-heat whole milk powder (WMP) was investigated. Milk (3.6% fat) was fortified with RBO at 0.1 and 0.2% (wt/wt) and was concentrated and dried. Control WMP was made without RBO addition. Thiobarbituric acid reactive substances (TBARS) were used to monitor oxidation during storage at 45 degrees C for 40 d. The oxidation of low-heat WMP was significantly reduced by addition of 0.1% RBO, but there was no significant effect on the oxidation of high-heat WMP. An increase of RBO to 0.2% did not significantly improve the oxidative stability when compared with 0.1% RBO. The TBARS in RBO-fortified, low-heat WMP increased with storage time up to 30 d but decreased with further storage to 40 d. The TBARS in all high-heat WMP and low-heat control WMP increased up to 20 d storage and then decreased with further storage. The most likely reason for this increase was due to the reaction of TBARS with milk proteins. Addition of RBO reduced the L (lightness) value and increased the b (yellowness) value but had no effect on the a (redness) value. When compared with the control milk powder, consumers could not detect any effect on the flavor of the reconstituted 0.1% RBO WMP but could detect a flavor difference in the 0.2% RBO WMP. PMID- 11104260 TI - Milk identification of different species: 13C-NMR spectroscopy of triacylglycerols from cows and buffaloes' milks. AB - Triacylglycerols from cows and buffaloes' milk fat were investigated by 13C nuclear magnetic resonance (NMR) spectroscopy. By the addition of pure triacylglycerols standards, we identified the resonances of both milk fats, and the peaks were used for qualitative and quantitative analysis of acyl groups. Multivariate analysis treatment of triacylglycerols distribution and composition parameters enabled us to identify milk. This study shows that NMR can safely be used to quantitate milk fatty acid content, providing unique information for milk identification of different animal species. PMID- 11104261 TI - Short communication: associations between blood calcium status at calving and milk yield in dairy cows. AB - The purpose of the present study was to estimate the effect of total blood plasma calcium (TBPCC) concentration at calving on milk yield in dairy cows. Data originated from 153 dairy cows in 27 herds from a single veterinary practice. For each cow, data included calcium concentration in a blood sample taken within 12 h postpartum, monthly test-day milk yield until 300 d in milk, calving date, parity, breed, and herd. The TBPCC ranged from 0.69 to 2.73 mmol/L, with a mean value of 1.80 mmol/L. The statistical analysis adjusted for the fixed effects of parity and lactation stage, random effects of herd and cow, and the correlation between repeated measures of test-day milk yield. The results showed that TBPCC at calving was not significantly related to fat- and protein-corrected milk yield at any lactation period. The present study indicates that hypocalcemia (low TBPCC) at calving is not an important risk factor for decreased milk yield. PMID- 11104262 TI - A cross-sectional study on the prevalence of Listeria monocytogenes and Salmonella in New York dairy herds. AB - As part of our long-term objective of assessing risk for Listeria monocytogenes and Salmonella spp. in dairy herds, we carried out a cross-sectional study to determine the prevalence of the two organisms. The study population consisted of a sample of dairy herds enrolled in the Quality Milk Promotion Services at Cornell during the period of April 1998 to March 1999. The sample was stratified by geographical region to assure representation. Four hundred and four dairy farms were enrolled in the study. In-line milk filters were collected from each farm for bacteriological examination of L. monocytogenes and Salmonella spp. Four hypothesized risk factors were evaluated for their association with the likelihood of the presence of each of the two organisms using logistic regression analysis. Listeria monocytogenes was isolated from 51 (12.6%) of the milk filters. We found region-specific differences in the rate of farms with positive milk filters for this pathogen. Salmonella spp. were isolated from 6 (1.5%) milk filters. One isolate was confirmed as Salmonella enterica Serotype Typhimurium DT 104. There was no significant association between any of the hypothetical risk factors and the likelihood of Salmonella spp. isolation. Our study demonstrated that both L. monocytogenes and Salmonella spp. were prevalent in milk filters in New York dairy herds and that Salmonella was isolated at a significantly lower rate then L. monocytogenes. PMID- 11104263 TI - A biological approach to lactation curve analysis for milk yield. AB - Lactation records of milk yield are commonly analyzed with empirical mathematical models. A family of new models is described based on the known biology of the mammary gland during pregnancy and lactation. The new models fit two logistic curves representing secretory cell differentiation and cell death (apoptosis) throughout lactation. A further function describes secretion rate changes. Both additive and multiplicative forms of the new models are described. Additional terms in the model can account for factors that alter the course of milk yield during the lactation, either by new cell differentiation, a change in the rate of milk secretion loss, or both. The new models were compared with three widely used empirical functions and another biologically based function using weekly records of milk yield taken from 40 dairy cows. The new models fitted the data with a lower residual mean square than the other functions. In addition, the parameters of the new models have a biological interpretation and can be used to discuss key characteristics of lactation. The application of the new models to monthly lactation records is possible, although a reduction in the number of parameters is likely to be required. Some parameters in the cell differentiation function of the models will be poorly estimated from monthly records and can be replaced with average or standard values. PMID- 11104264 TI - Induced lactation in prepubertal Holstein heifers. AB - Lactation was hormonally induced in six prepuberal Holstein heifers by seven daily injections of estrogen and progesterone and three injections of dexamethasone on d 18, 19, and 20, followed by twice daily hand milking beginning on d 21. Heifers were about 6 mo old and weighed 162 kg at the beginning of the experiment. Secretions were obtained from five of six of heifers, and twice daily milking continued for 75 d in three of five heifers. The volume of milk obtained on d 7 ranged from 32 to 500 ml and averaged 4.7, 4.1, and 3.7% lactose, protein, and fat, respectively. In the first natural lactation, milk yield and composition were nearly identical for controls and induced heifers. Serum alpha-lactalbumin was increased in induced heifers after treatment with dexamethasone and was highest on d 10 after onset of milking. Our data suggest that sufficient secretions for extensive biochemical testing can be obtained following hormonal induction of lactation in a majority of prepubertal heifers. Moreover, hormonal induction of lactation had no apparent effect on reproduction or first natural lactation. While it is unlikely that hormonal induction of lactation in prepubertal heifers is practical from a dairy production viewpoint, the advent of biotechnology for production of therapeutic recombinant proteins in the mammary gland of transgenic livestock has made early detection of these transgenic proteins very desirable. We conclude that induction of lactation in prepubertal heifers is a viable technique for testing the expression of mammary-linked gene constructs in transgenic cattle. PMID- 11104265 TI - Risk factors associated with clinical mastitis in low somatic cell count British dairy herds. AB - A cross-sectional survey of dairy farms with low bulk milk somatic cell counts was carried out to assess the level of clinical mastitis and to quantify risk factors associated with the incidence rate of clinical mastitis. Questionnaires were sent to 3009 milk operations with an annual mean bulk milk somatic cell count of less than 100,000 cells/ml during 1997. A response rate was 61%. The mean incidence of clinical mastitis reported was 22.8 cases per 100 cows/yr. Negative binomial regression models were used to assess statistically significant risk factors associated with the incidence of clinical mastitis. The incidence increased when farmers reported that they had straw yard housing for milking cows (compared with cubicle housing), mucked out the calving area less frequently than once per month, kept cows standing in a yard after milking, always practiced postmilking teat disinfection, had greater than 50% replacement rate, had some cows that leaked milk on entry to the parlor, had some cows that leaked milk at other times, and foremilked before cluster attachment. The incidence of clinical mastitis was lower on farms when the gathering yard used before milking was scraped at least twice a day, cows were offered feed after both milkings, rubber gloves were not worn during milking, teat liners were changed after 6000 milkings, and the average dry period was less than 40 d. The study has identified areas of the environment in which efforts to improve hygiene should be focused. PMID- 11104266 TI - Can relative spermatozoal galactosyltransferase activity be predictive of dairy bull fertility? AB - The best and poorest bovine semen samples used commercially for artificial insemination in dairy cattle typically differ in pregnancy rates by 20 to 25% but are within a range that pregnancy rates cannot be predicted consistently by commonly used laboratory assays. Sperm motility and morphology are the characteristics most often evaluated. Laboratory assays that measure other functional traits of sperm may be useful as supplemental assays to increase the reliability of predicting fertility. One such functional trait is the ability of sperm to bind to the zona pellucida, a process mediated by complementary receptors on each gamete. On mouse sperm, beta1,4-galactosyltransferase acts as a receptor for the zona pellucida. Beta1,4-galactosyltransferase is expressed on sperm from many mammals, including bovine sperm, and is a candidate for a zona pellucida receptor. The ability of sperm to bind to the zona pellucida may be related to the amount of beta1,4-galactosyltransferase present on sperm. The aim of this work was to determine if bull sperm beta1,4-galactosyltransferase activity was related to fertility. Beta1,4-galactosyltransferase enzyme assays were performed on sperm from 24 bulls whose fertility was estimated by nonreturn rate and on sperm from a second group of seven bulls whose fertility was ranked by in vivo competitive fertilization. Beta1,4-galactosyltransferase activity varied between individual bulls but was not correlated to fertility as estimated by nonreturn rate or by competitive fertilization. These results demonstrate that beta1,4-galactosyltransferase activity on sperm varies between animals, but that beta1,4-galactosyltransferase activity alone is not an accurate indicator of fertility in dairy bulls. PMID- 11104267 TI - Postprandial metabolism and endocrine status in veal calves fed at different frequencies. AB - Veal calves fed by bucket often develop postprandial insulin resistance, hyperglycemia, and glucosuria during fattening. Automatic feeding systems allow feed intake for 24 h, and small ingested portions are expected to decrease postprandial glucose loads. We have studied metabolic and endocrine traits in calves that were either 1) fed identical daily amounts of whole milk plus milk replacer by a computer-programmed automatic feeder (> or =6 portions from 0800 to 2400 h) (GrA) or 2) fed by bucket at 0800 and 1630 h (GrB). Calves started at a body weight of 118 kg, and the experiment lasted for 3 wk. During wk 3, lactose was supplemented to stress postabsorptive glucose homeostasis. Feed intake and average daily gains in GrA and GrB were similar. Plasma concentrations during an 8-h period of glucose (in part), lactate, urea, and somatostatin (in wk 3), and of glucagon and insulin (wk 2 and 3) were smaller in GrA than in GrB, whereas growth hormone, insulin-like growth factor I, insulin-like growth factor binding protein-1 (wk 2), and prolactin concentrations (wk 2 and 3) were higher. Lactose supplementation in wk 3 enhanced transient postprandial hyperglycemia and hyperinsulinemia. Thus, there were marked metabolic and endocrine differences when calves sucked their feed in six or more portions during a 16-h period from an automatic feeder compared with twice daily drinking from a bucket. Ingestion of small portions by calves avoided marked hyperglycemia and lactate increments, and lower plasma urea concentrations mirrored enhanced nitrogen utilization, possibly mediated by the altered growth hormone, IGF-I and insulin status. PMID- 11104268 TI - Effects of dietary inclusion of chromium propionate and calcium propionate on glucose disposal and gastrointestinal development in dairy calves. AB - In experiment 1, 21 male Holstein calves (43.9 kg) were fed only milk replacer at 1.4% of their body weight as dry matter for 6 wk. Dietary treatments included a commercial milk replacer (22% protein, 15% fat) containing (dry basis) either 6.4% Ca propionate or 6.4% dextrose (control) and either 0 or 0.5 mg/kg of supplemental Cr as Cr propionate. Neither Cr nor Ca propionate affected body weight gain; however, Ca propionate tended to increase the growth of the entire foregut measured after slaughter at 6 wk of age. A Minimal Model glucose tolerance test indicated that insulin sensitivity was not affected by treatment. However, calves fed Cr had higher glucose disappearance indexes than controls when propionate was not fed (0.013 vs. 0.019 units) but similar clearance when propionate was included (0.018 vs. 0.018 units, Cr x P interaction). The area under the glucose response curves after propionate-loading tests was much greater for calves fed the Cr versus control replacer when propionate was not present; however, when propionate was included, the response was less dramatic. In experiment 2, 25 Holstein calves were used to study performance and metabolic responses when milk replacer, and then postweaning starter, were supplemented with 0.5 mg/kg of Cr as Cr propionate. The metabolic responses of these calves were not affected by treatment. Overall, combined data suggested that supplemental Cr may improve glucose effectiveness; however, these responses seemed to be attenuated by supplemental propionate. PMID- 11104269 TI - Effect of maturity on degradation kinetics of sod-seeded cereal grain forage grown in northern Arkansas. AB - Wheat (Triticum aestivum L.), oat (Avena sativa L.), and rye (Secale cereale L.) were overseeded into a dormant bermudagrass (Cynodon dactylon (L.) Pers.) sod and harvested at 3-wk intervals throughout March, April, May, and early June. Plant growth stage was documented for each forage on each harvest date, and harvested forages were evaluated for forage quality characteristics. Degradation kinetics of DM and NDF for these forages were evaluated by the in situ method. Fractional degradation rates for DM and NDF in all three species were relatively rapid for vegetative forage (> or =0.086 h(-1)) but declined rapidly by the heading stage of development and stabilized thereafter. Forage quality declined and forages were more resistant to ruminal degradation as plants entered the reproductive stages of growth. Based on these findings, growth stage is an effective predictor of most characteristics of in situ DM and NDF disappearance. The relationships between these degradation parameters and growth stage were typically explained with quadratic or cubic models. Clearly, forage quality characteristics of overseeded rye deteriorated more rapidly with phenological development and growth stage than quality characteristics of overseeded wheat and oat grown in the same environment. For rye, this problem is further complicated by its accelerated phenological development. These factors combine to permit a very narrow harvest window in early spring, relative to the other cereal grains evaluated. Acceptable forage quality may persist for an extended period in wheat and oat; this suggests that producers wishing to utilize these forages may lengthen the harvest window by planting more than one species, either as a mixture or preferably in independent stands. PMID- 11104270 TI - A comparison of methods of adding fibrolytic enzymes to lactating cow diets. AB - Holstein cows (n = 43) in early lactation were used to investigate effects of method of adding fibrolytic enzymes to diets on feed intake, milk production, and digestibility. Cows were blocked according to parity and calving date and randomly assigned to three treatments: control, enzymes applied to the total mixed ration (E-TMR), or enzymes added to the barley-based concentrate (E-Conc). The enzyme product used was a proprietary blend that contained relatively high xylanase and low cellulase activities (Biovance Technol. Inc., Omaha, NE). An enzyme solution (50 mg of enzyme powder dissolved into 20 ml of water) was sprayed onto each kilogram of total mixed ration (dry matter basis) before feeding. Alternatively, 73 g of enzyme powder, dissolved in 20 L of water, was added per ton of concentrate (50 mg of enzyme/kg of diet dry matter). The total mixed rations contained 24% corn silage, 14% alfalfa hay, and 62% concentrate (dry matter basis) and were offered ad libitum. In vitro gas production assays and two experiments using sheep were conducted to measure the effects of enzyme treatment on digestibility. Dry matter intake (mean: 19.8 kg/d) was not affected by enzyme supplementation. Milk yield (kg/d) was higher for cows fed E-Conc (37.4) than for cows fed control (35.3) or E-TMR (35.2) with no effects on milk composition. Total tract digestibility (%) of dry matter was higher for E-Conc (66.6%) than for the control diet (63.9%) and intermediate for E-TMR (65.7%) when measured in dairy cows. However, the digestibility of the diets was substantially higher when measured in sheep than in dairy cows, with no effects of enzyme supplementation. The results indicate that fibrolytic enzymes have the potential to increase digestibility and milk production in dairy cows because digestion is low relative to potential digestibility. When digestion is higher, as was observed in lambs or in vitro, no improvement in digestibility occurs. Method of enzyme delivery must also be considered to maximize the benefits of using fibrolytic enzymes in dairy cow diets. PMID- 11104271 TI - Influence of mechanical processing on utilization of corn silage by lactating dairy cows. AB - We conducted three experiments to determine the influence of mechanical processing on corn silage utilization by lactating dairy cows. Total mixed rations contained either unprocessed or processed corn silage harvested between 1/4 and 3/4 milk line. In trial 1, 12 multiparous Holstein cows were used in a replicated double switchback design with 21-d periods. Intake of dry matter (DM) was increased 1.2 kg/d by processing, but milk yield was unaffected. Processing did not affect apparent total-tract DM digestibility, but processing tended to lower starch and corn excretion in feces and reduced concentration of sieved corn kernel particles in feces. In trial 2, 42 Holstein cows were used in an 18-wk randomized complete-block design. Intake of DM and milk yield were unaffected by processing, but milk fat percent was increased 0.35 percentage units by processing. Processing tended to increase total-tract digestibility of starch, but reduced organic matter, crude protein, and neutral detergent fiber digestibilities. In trial 3, 30 Holstein cows were used in a 15-wk randomized complete block design. There was no influence of mechanical processing on intake or lactation performance in this trial. Despite indications of increased starch digestion in two trials and increased DM intake in one trial, effects of processing corn silage on lactation performance were minimal with corn silage at the maturity and moisture contents used in these trials. PMID- 11104272 TI - Effects of corn processing and supplemental hay on rumen environment and lactation performance of dairy cows grazing grass-legume pasture. AB - The effect of corn processing (9 kg of dry matter/d of ground dry shelled or 9 kg of dry matter/d of steam rolled) and supplemental hay (0 or 3.2 kg of dry matter/d of alfalfa hay) on milk yield and composition, rumen environment, and starch utilization by lactating cows grazing grass-legume pasture was studied. Twelve rumen cannulated, multiparous Holstein cows in early lactation (95 d in milk), were assigned to a 4 x 4 Latin square design replicated three times. Treatments were ground shelled corn-based concentrate, ground shelled corn-based concentrate plus alfalfa hay, steam-rolled, corn-based concentrate, or steam rolled, corn-based concentrate plus alfalfa hay. Supplements were fed in equal proportions twice daily. Cows fed steam-rolled corn tended to have higher percentage of milk protein and lower milk urea nitrogen concentrations than cows fed shelled corn. Milk yield was not affected by corn processing or hay supplementation. Intake of pasture forage but not total dry matter intake was reduced by hay supplementation. Starch plus free glucose digestibility in the total tract was not affected by grain processing; however, starch plus free glucose digestibility tended to increase with hay supplementation. Supplemental hay increased starch plus free glucose digestibility through changes in rumen digestion kinetics. Hay supplementation reduced the liquid rate of passage, and tended to reduce particulate turnover. Rumen degradability of pasture forage organic matter tended to be higher for cows fed supplemental hay. Supplemental hay in these diets had a greater impact on starch utilization than corn processing. PMID- 11104273 TI - Effect of whole and expanded-expelled cottonseed on milk yield and blood gossypol. AB - Thirty-two primiparous and 12 multiparous Holstein cows were randomly assigned at calving to treatments to determine the effects of type and amount of cottonseed product on plasma gossypol, milk production, and composition, and conjugated linoleic acid concentration in milk fat. Rations consisted of corn silage, corn grain, soybean meal, and cottonseed hulls, and contained on average 16.8% crude protein and 25.3% acid detergent fiber on a dry matter basis. On a dry matter basis, diets contained one of the following: 1) 14% whole cottonseed; 2) 14% expanded-expelled cottonseed; 3) 21% expanded-expelled cottonseed; or 4) 28% expanded-expelled cottonseed. Cows remained on treatment from 30 through 120 d in milk. Dry matter intakes were not significantly different, but intakes of crude protein, acid detergent fiber, and fat were higher for multiparous cows fed whole cottonseed. Multiparous cows fed whole cottonseed had higher yields of milk, fat corrected milk, crude protein, fat and solids-not-fat than those fed any level of expanded-expelled cottonseed. Concentrations of milk fat, protein, and SNF were not affected by treatment. Although there were treatment differences in fat intake, there were no production differences in primiparous cows. Milk production efficiency (fat-corrected milk/dry matter intake) was not affected by treatment for either multiparous or primiparous cows. Cows fed 14% whole or 14% expanded expelled cottonseed had similar levels of total plasma gossypol and plasma levels of the negative isomer of gossypol. Increasing the level of expanded-expelled cottonseed in the diet increased both total plasma gossypol and the negative isomer. In this experiment, multiparous but not primiparous cows fed whole cottonseed produced more milk than those fed expanded-expelled cottonseed at 14 to 28% of the diet dry matter, however, feed efficiencies were similar for all treatments. PMID- 11104274 TI - Potential of fermentation byproducts as nitrogen supplements for lactating dairy cows. AB - Two feeding trials evaluated several byproducts from commercial amino acid fermentations as N supplements for lactating cows. Trial 1 was a replicated 5 x 5 Latin square that used 2-wk periods and 25 Holstein cows (five with ruminal cannulae) fed diets containing [dry matter (DM) basis] 28% alfalfa silage, 31% corn silage, 28% high moisture ear corn plus 4 percentage units of crude protein (CP) from: soybean meal, urea, commercial fermentation byproduct 1 or 2, or a blend of fermentation byproducts plus wheat middlings. Diets averaged 15.1% CP and 32% neutral detergent fiber. Intake of DM, body weight (BW) gain, and yield of milk and milk components were greatest for cows fed soybean meal; animal performance was similar with urea, byproduct 1 and the byproduct blend. Intake, BW change, and yield of milk and protein when cows were fed byproduct 2 were lower than when fed urea. Urine output (estimated with creatinine in spot urine samples) was greater on fermentation byproduct 1 and the byproduct blend. There were no differences due to N source in microbial synthesis (based on estimated purine derivative excretion), in situ digestion of alfalfa hay DM, or molar proportions of ruminal volatile fatty acids. Trial 2 was a replicated 5 x 5 Latin square using 2-wk periods and 10 Holstein cows fed diets containing (DM basis) 37% alfalfa silage, 28% corn silage, 29% high moisture ear corn plus 2 percentage units of CP from urea, fermentation byproduct 1, or one of three blends of fermentation byproducts plus wheat middlings. Except for greater DM intake in cows fed the byproduct blends, performance and urinary metabolite excretion did not differ because of N supplement. Relative to other fermentation byproducts and urea, byproduct 1 resulted in reduced milk urea N in both trials. Under the conditions of these trials, fermentation byproducts were less effective than soybean meal, and no more effective than urea, as N supplements. PMID- 11104275 TI - The effect of roasting nonlinted whole cottonseed on milk production by dairy cows. AB - This study was conducted to examine the effect of roasting nonlinted whole cottonseed on ruminal crude protein (CP) degradability and performance in high yielding dairy cows. Multiparous Israeli Holstein-Friesian cows (parity average 2.5+/-1.5; n = 132) with 571+/-65 kg of body weight (BW), 107+/-48 d in milk (DIM), and 37+/-5.8 kg of milk yield/d were used in the study. Cows were divided into two dietary treatment groups according to their BW, DIM, and milk production. The two diets were similar in CP, net energy for lactation, and neutral detergent fiber content [17%, 1.74 Mcal/kg, and 30% on a dry matter (DM) basis] and included either 15% (on a DM basis) whole cottonseed or roasted whole cottonseed. Ruminal effective degradability of CP, organic matter (OM), and ether extract (EE) decreased 14, 11, and 10%, respectively, compared to whole cottonseed. Total tract digestibilities of CP and EE were similar for both treatments and averaged 57 and 59%, respectively. However, DM and OM digestibilities were 6 and 5% higher in cows offered roasted whole cottonseed relative to those fed whole cottonseed diet. The inclusion of roasted whole cottonseed in the ration decreased ruminal ammonia and blood urea N concentration by 12% compared with diet with the raw whole cottonseed. Milk production, milk fat content, and production, and milk protein yield increased when roasted, nonlinted whole cottonseed was included in the diet. Milk protein content was similar for both treatments, averaging 2.92%. PMID- 11104276 TI - Effects of amount and source of fat on the rates of lipolysis and biohydrogenation of fatty acids in ruminal contents. AB - Because the percentage loss of unsaturated fatty acids across the rumen has varied considerably in previous in vivo studies, we conducted five experiments to identify potential factors that might affect the in vitro rates of lipid lipolysis and biohydrogenation in ruminal contents. The factors examined included the amount of fat added to the substrate, the source of added fat, the diet fed to the donor fistulated cow, and the time of collection of inoculum from the donor cow. Lipolysis and biohydrogenation were expressed as the rates of disappearance of neutral lipid and unsaturated fatty acids, respectively, from the culture contents over time using a first-order model. The rate of lipolysis of soybean oil declined from 44%/h to less than 30%/h as the percentage of soybean oil in the culture substrate increased from 2 to 10%. The overall rate of biohydrogenation of C18:2 was 14.3%/h, but declined 1.2%/h for each percentage unit increase in C18:2 added to the substrate. Compared with C18:2, the rates of biohydrogenation of C18:1 were generally lower (averaged 3.6 %/h) for all fat sources. The rate of biohydrogenation of C18:2 in soybean oil was not affected by the amount of grain or fat fed to the donor cow, or the time after feeding that ruminal inoculum was collected. Based on these findings, high linoleic acid concentrations in the diet would possibly reduce biohydrogenation and increase the postruminal flow of this unsaturated fatty acid. Also, lipolysis may vary considerably due to amount and source of lipid added to the diet, but this has little influence on the initial disappearance rates of linoleic or oleic acids from ruminal contents. PMID- 11104277 TI - Effect of sugars and malate on ruminal microorganisms. AB - The objective of this study was to examine the effects of a commercial feed supplement that contains sugars and malate on lactate fermentation by Selenomonas ruminantium grown in batch culture. Experiments also were conducted to examine the effects of this feed supplement on the mixed ruminal microorganism fermentation of ground corn and soluble starch in the presence and absence of 5 mg/kg of monensin. When S. ruminantium strains HD4 and H18 were incubated in basal medium that contained DL-lactate, some DL-lactate was utilized by both strains after 24 h. In the presence of 1 g/L of sugars plus malate commercial feed supplement, both strains used most of the carbohydrate associated with the feed supplement between 6 and 8 h, and lactate was the main end product. In ground corn fermentations by mixed ruminal microorganisms, 2.25 and 3.25 g/L of sugars plus malate commercial feed supplement increased concentrations of acetate, propionate, and total volatile fatty acids, while 3.25 g/L increased lactate and decreased final pH and butyrate. Fermentation of soluble starch in the presence of both concentrations of sugars plus malate commercial feed supplement increased concentrations of acetate, propionate, and total volatile fatty acids and decreased the acetate:propionate ratio. In the presence of 5 mg/kg of monensin, sugars plus malate treatment increased concentrations of propionate and total volatile fatty acids in ground corn and soluble starch fermentations. Collectively, these results suggest that the sugars plus malate commercial feed supplement stimulates the ruminal fermentation. PMID- 11104278 TI - Mixed ruminal microbes of cattle produce isopropanol in the presence of acetone but not 3-D-hydroxybutyrate. AB - The objective was to evaluate the ability of mixed rumen microbes to synthesize isopropanol from acetone or 3-D-hydroxybutyrate. Rumen fluid from seven mature, nonpregnant, dry Holstein cows was incubated with starch or cellulose and additions of acetone, 3-D-hydroxybutyrate, or saline. Rumen fluid was analyzed for isopropanol after 0, 3, 6, and 9 h. No isopropanol was present in any sample at 0 h, and none was present in incubations containing saline or 3-D hydroxybutyrate at any subsequent time. Incubations that included acetone produced small amounts of isopropanol from 0 to 3 and 3 to 6 h and significantly larger amounts from 6 to 9 h. With starch as the energy substrate, production from 6 to 9 h was 3.8 micromol/min per liter of rumen fluid and 3.7 micromol/min per liter with cellulose as the energy substrate; however, these values did not differ significantly. Mixed rumen microbes could synthesize isopropanol from acetone but not from 3-D-hydroxybutyrate, and rumen microbial metabolism of acetone was the likely source of plasma isopropanol seen in ketotic ruminants. PMID- 11104279 TI - Nitrogen metabolism of early lactation cows fed diets with two different levels of protein and different amino acid profiles. AB - Four multiparous Holstein cows (569+/-122 kg) surgically prepared with indwelling catheters in the mesenteric, portal, and hepatic veins and carotid artery were allocated in a 4 x 4 Latin square to determine the effects of dietary crude protein (CP) level and amino acid (AA) profile on N metabolism during early lactation (from 25 to 65 d in milk). Cows received their diets in two equal meals and were milked twice daily. The dietary treatments were: 18% CP with a high (18H) or a low (18L) quality AA profile, and 15% CP with a high (15H) or a low (15L) quality AA profile. The four diets were similar in net energy for lactation (1.75 NEL Mcal/kg) and contained the same amount of RUP (34% of CP). The quality of the AA profile pertained only to the essential AA (EAA), and was assessed by comparison with the EAA profile of casein and considered the potential contribution of EAA from ruminal bacteria. The 18H and 15H diets were supplemented with 50 and 25 g/d of ruminally protected Met, respectively. After 10 d on treatment, a blood flow marker (p-amino-hippurate) was infused into a mesenteric vein, and arterial, portal, hepatic, and mammary blood samples were obtained at 3, 6, and 12 h after feeding. Dry matter intake was similar across treatments (23.4+/-0.5 kg/d). Amino acid oxidation, and consequent urea production, in the liver were numerically greater with the 18% CP rations, and, as a result, arterial urea concentrations were greatest (P < 0.01) with these rations. The amount of total AA extracted by the mammary gland tended to be greater with the H than with the L diets (21.4 vs. 18.2 mmol/ h, respectively). Milk yield tended to be greater (P = 0.16) with the 18H and 15H diets (47.7 and 46.3 kg/d, respectively) compared with the 18L and 15L diets (45.9 and 44.6 kg/d, respectively). Also, milk CP and casein contents were greatest (P = 0.09) with the H diets compared with the L diets. Milk and plasma urea N were greatest (P < 0.01) with the 18% CP diets. The efficiency of N utilization for milk protein synthesis was greatest (P < 0.09) with the 15% CP diets. It is concluded that milk protein production during early lactation is less susceptible to variations in dietary CP contents than variations in the AA profile of the dietary protein. PMID- 11104280 TI - Responses to graded postruminal doses of histidine in dairy cows fed grass silage diets. AB - Five Finnish Ayrshire cows were used in a 4 x 4 Latin square experiment designed to study the effects of graded doses of postruminal His infusion on milk production, arterial concentrations, and mammary uptakes of plasma amino acids (AA) as well as utilization of added His. Grass silage (16.9% CP) was given ad libitum with 8 kg of cereal-based concentrate per day (11.3% CP). Treatments were abomasal or duodenal infusions of 250 g of glucose/d in combination with 0, 2, 4, or 6 g of His/d. Infusions did not affect dry matter intake (mean 18 kg/d). Infusion of His increased milk yield linearly from 27.0 to 28.8 kg/d, protein yield from 861 to 919 g/d and lactose yield from 1345 to 1457 g/d. Milk fat yield and content changed in a cubic manner (1240, 1167, 1296, and 1177 g/d and 4.60, 4.16, 4.60, and 4.09). Infusion of His had no influence on milk protein or lactose concentrations. Arterial Lys and His concentrations increased linearly, but other AA concentrations were unaffected as well as calculated arteriovenous differences and mammary AA uptakes. The extraction of His decreased linearly with an increasing amount of His. The utilization of added His (28%) was not affected by the level of infusion, and mammary AA uptake seemed to be regulated by an inverse relationship between arteriovenous difference of essential AA and calculated mammary plasma flow. This experiment confirmed that His is the first limiting AA on grass silage-cereal based diets. PMID- 11104281 TI - Milk composition and apparent digestibilities of dietary fatty acids in lactating dairy cows abomasally infused with Cis or Trans fatty acids. AB - Fat supplementation of diets for dairy cows produces changes in nutrient supply and milk composition. The effect of abomasal infusion of either cis-C18:1 or trans-C18:1 fatty acid isomers on the digestibility of fatty acids and milk composition was determined in lactating dairy cows. Six multiparous midlactation Holstein cows were used and fed a control diet containing 50% forage and 50% concentrate. Treatments were (per day): no infusion, infusion of a 630-g fat mixture high in cis-C18:1 isomers, and infusion of a 623-g fat mixture high in trans-C18:1 isomers using two 3 x 3 Latin squares with 4-wk experimental periods. Fat infusion did not affect total dry matter intake and increased apparent digestibilities of total fatty acids. Apparent digestibilities of C18 fatty acids were directly related to the number of double bonds within isomers, and cis-C18:1 isomers were slightly more digestible than trans-C18:1 isomers. The lower yield of C12:0, C14:0, and C16:0 fatty acids in milk fat and higher milk citrate observed when cows were infused with trans-C18:1 suggests a depressed de novo milk fatty acid synthesis. Effects of trans infusion on milk fat were independent of ruminal fermentation, fatty acid apparent absorption, and fatty acid plasma concentrations. Lower milk protein yield in cows infused with fat may have been caused by a decrease in milk protein synthesis. PMID- 11104282 TI - Influence of dietary fish oil on conjugated linoleic acid and other fatty acids in milk fat from lactating dairy cows. AB - Lactating cows were fed menhaden fish oil to elevate concentrations of conjugated linoleic acid, transvaccenic acid, and n-3 fatty acids in milk. Twelve multiparous Holstein cows at 48+/-11 DIM were assigned randomly to a replicated 4 x 4 Latin square. Each treatment period was 35 d in length, with data collected d 15 to 35 of each period. On a dry matter (DM) basis, diets contained 25% corn silage, 25% alfalfa hay, and 50% of the respective concentrate mix. Fish oil was supplemented at 0, 1, 2, and 3% of ration DM. Linear decreases were observed for DM intake (28.8, 28.5, 23.4, and 20.4 kg/d) and milk fat (2.99, 2.79, 2.37, and 2.30%) for 0 to 3% dietary fish oil, respectively. Milk yield (31.7, 34.2, 32.3, and 27.4 kg/d) increased as dietary fish oil increased from 0 to 1% but decreased linearly from 1 to 3% dietary fish oil. Milk protein percentages (3.17, 3.19, 3.21, and 3.17) were similar for all treatments. When the 2% fish oil diet was fed, concentrations of conjugated linoleic acid and transvaccenic acid in milk fat increased to 356% (to 2.2 g/ 100 g of total fatty acids) and 502% (to 6.1 g/100 g), respectively, of amounts when 0% fish oil was fed. There were no additional increases in these fatty acids when cows were fed 3% fish oil. The n-3 fatty acids increased from a trace to over 1 g/100 g of milk fatty acids, when the 3% fish oil diet was fed. Fish oil supplementation to diets of dairy cows increased the conjugated linoleic acid, transvaccenic acid, and n-3 fatty acids in milk. PMID- 11104283 TI - Genetic evaluation for longevity of Dutch dairy bulls. AB - Parameters needed for survival analysis of longevity records of cows to predict breeding values of their sires were estimated with data on Dutch Black and White and Red and White cows. The heritabilities of functional productive life were 0.041 and 0.036 on the log scale for Black and White and Red and White cows, respectively. Although the heritabilities and other parameters differed between both breeds, the resulting breeding values were hardly affected: the correlation between breeding values of Red and White bulls using either Red and White parameters or Black and White parameters was 0.992. Genetic correlations between the direct breeding value for functional longevity (based solely on longevity of sires' daughters) and breeding values for conformation, health, and fertility traits were calculated. Several alternative selection indices were investigated using these correlations. Based on the resulting reliabilities, it was concluded that the Dutch breeding value for functional longevity should be based on longevity, rump angle, teat placement, udder depth, feet and legs, and somatic cell count. The index was expressed on a scale with average of 100 and a standard deviation of 4 points (at 80% reliability). The economic value was Dfl. 65 per genetic standard deviation, which was 0.46 times the economic value of INET (Net Milk Revenue Index). For the breeding value for functional longevity that was first published in August 1999, slight modifications in the model were made. PMID- 11104284 TI - Genetic parameters of legendre polynomials for first parity lactation curves. AB - Variance components of the covariance function coefficients in a random regression test-day model were estimated by Legendre polynomials up to a fifth order for first-parity records of Dutch dairy cows using Gibbs sampling. Two Legendre polynomials of equal order were used to model the random part of the lactation curve, one for the genetic component and one for permanent environment. Test-day records from cows registered between 1990 to 1996 and collected by regular milk recording were available. For the data set, 23,700 complete lactations were selected from 475 herds sired by 262 sires. Because the application of a random regression model is limited by computing capacity, we investigated the minimum order needed to fit the variance structure in the data sufficiently. Predictions of genetic and permanent environmental variance structures were compared with bivariate estimates on 30-d intervals. A third order or higher polynomial modeled the shape of variance curves over DIM with sufficient accuracy for the genetic and permanent environment part. Also, the genetic correlation structure was fitted with sufficient accuracy by a third order polynomial, but, for the permanent environmental component, a fourth order was needed. Because equal orders are suggested in the literature, a fourth-order Legendre polynomial is recommended in this study. However, a rank of three for the genetic covariance matrix and of four for permanent environment allows a simpler covariance function with a reduced number of parameters based on the eigenvalues and eigenvectors. PMID- 11104285 TI - Bias in genetic evaluations by records of cows treated with bovine somatotropin. AB - Records from Dairy Records Management Systems in Raleigh were used to estimate effects of bovine somatotropin (bST) treatment and to predict breeding values for milk production traits. The data comprised 5245 test-day records of bST-treated cows and 126,223 test-day records of untreated cows in first lactation for milk, fat, and protein yields. Fixed effects of bST treatment were estimated from test day animal models with herd-test-date as another fixed factor. Percentage increases due to bST treatment ranged from 7 to 8% for test-day milk, fat, and protein yields. Random regression coefficients for additive genetic and permanent environmental effects were included in the model. To assess the potential for bias in genetic evaluations when some and not all cows are treated with bST, breeding values predicted by the test-day model with and without effects of bST treatment were compared for cows and sires. Correlations between breeding values predicted from models with and without effects of bST treatment were 0.99. However, relatively large bias was found for individual animals. This result suggests that bias in genetic evaluation caused by ignoring bST treatment may be significant. PMID- 11104286 TI - Interactions among factors affecting stillbirths in Holstein cattle in the United States. AB - Each year about 7% of the Holstein calves born in the United States die within 48 h of birth. The exact cause of death is unknown. The purpose of this article is to examine the complex interactions among factors (e.g., parity, season of birth, dystocia, year) contributing to stillbirth rates. A modified chi-squared automated interaction detection algorithm was used to develop classification trees explaining the most likely sequence of factors that result in a stillborn calf. The data were 666,341 births from the MidStates Dairy Records Processing Center and the National Association of Animal Breeders. Primiparous and multiparous cows clearly differ in the rate of stillbirths, 11.0 and 5.7%, respectively. Dystocia followed parity as the next most important factor within both primiparous and multiparous cows. In primiparous cows, season, year of birth, or gestation length ranked third as an important predictor for dystocia equal to 1, 2, or 3+, respectively. Gestation length ranked third in importance among the factors that affect stillbirth rates for all levels of dystocia in multiparous cows. Among multiparous cows needing assistance (dystocia 3+), stillbirth rates were greatest for shorter gestations less than the average of 280 d, 55.3% for -15 to -12 d, 45.5% for -11 to -9 d, 33.7% for -8 to -5 d, 23.8% for -4 to 13 d, and 35.4% for 14 to 15 d. Gestation length pinpointed the time when stillbirths occurred, as indicated by the increase from 23.8% stillbirth rate among calves born at or above the mean gestation length to 55.3% for those calves born -15 to -12 d below the mean gestation. Further investigation of the relationship between stillbirth rates and gestation length is needed to develop a more complete understanding of the biological processes resulting in the loss of calves at birth. PMID- 11104287 TI - The genetic relationship between calving interval, body condition score and linear type and management traits in registered Holsteins. AB - The trend to poorer fertility in dairy cattle with rising genetic merit for production over the last decade suggests that breeding goals need to be broadened to include fertility. This requires reliable estimates of genetic (co)variances for fertility and other traits of economic importance. In the United Kingdom at present, reliable information on calving dates and hence calving intervals are available for most dairy cows. Data in this study consisted of 44,672 records from first lactation heifers on condition score, linear type score, and management traits in addition to 19,042 calving interval records. Animal model REML was used to estimate (co)variance components. Genetic correlations of body condition score (BCS) and angularity with calving interval were -0.40 and 0.47, respectively, thus cows that are thinner and more angular have longer calving intervals. Genetic correlations between calving interval and milk, fat, and protein yields were between 0.56 and 0.61. Records of phenotypic calving interval were regressed on sire breeding values for BCS estimated from records taken at different months of lactation and breeding values for BCS change. Genetic correlations inferred from these regressions showed that BCS recorded 1 mo after calving had the largest genetic correlation with calving interval in first lactation cows. It may be possible to combine information on calving interval, BCS, and angularity into an index to predict genetic merit for fertility. PMID- 11104288 TI - Approaches to estimating daily yield from single milk testing schemes and use of a.m.-p.m. records in test-day model genetic evaluation in dairy cattle. AB - Statistical models were presented to estimate daily yields from either morning or evening test results. The 64,451 test-day records from 10,392 lactations of 8800 cows were available for analysis from experiments that were designed to investigate the accuracy of an alternate morning and evening four-weekly milk testing scheme. The experiments were conducted in 152 herds from six German states and covered a span from 1994 to 1998. Milk yield, fat, and protein percentage were recorded for all of the morning and evening milkings. Seven statistical models were fitted to the data to derive formulas for estimating daily yields from morning or evening yields. In general, use of evening milkings less accurately estimated yields than did use of morning milkings. Among the three yield traits the lowest accuracy of estimation of daily yield was found for fat yield. Although the models do not differ much in the correlation between estimated and true daily yields, systematic under- and overestimation of daily yield at the beginning and end of lactation were observed in all models with the exception of model 6, which accounted for heterogeneous variances by parity class, milking interval class, and lactation stage by fitting separate regression formulas within each combination of the three factors. A study to validate the models showed that model 6 is also robust for the analyzed populations. Smoothing model 6 regression formulas across lactation stages caused a systematic pattern of estimation error, although loss in accuracy was minimal by fitting far fewer parameters in the regression formulas. Differences in the accuracy of alternate milking schemes to predict daily yields were found between traits, between morning and evening milkings, and between parity classes. Compared with true daily yields from different lactation stages, variances and correlations of the estimated yields were reduced, which must be accounted for in genetic evaluation. The use of estimated daily yields from morning or evening milkings has a smaller impact on estimated breeding values of bulls than cows. As a result of lower heritability and repeatability of estimated daily yields than true daily yields, the weight on own test-day records for estimating cows' breeding values is lower when cows are in a.m.-p.m. than conventional monthly testing schemes. However, the difference in the weights between estimated and true daily yields decreases as lactation progresses. Use of estimated daily yields is less reliable for estimating breeding value than use of true daily yields. PMID- 11104289 TI - Relationships between energy balance and health traits of dairy cattle in early lactation. AB - The objective of the study was to calculate phenotypic relationships between energy balance in early lactation and health and reproduction in that lactation. Data were 26,701 daily records of dry matter intake and milk production, periodic measures of milk composition and body weight, and all health and reproductive information from 140 multiparous Holstein cows. Daily energy balance was calculated by multiplying feed intake by the concentration of energy of the ration and subtracting the amount of energy required for maintenance (based on parity and body weight) and for milk production (based on yield and concentrations of fat, protein, and lactose). Six measures of energy balance were defined: mean daily energy balance during the first 20, 50, and 100 d of lactation; minimum daily energy balance; days in negative energy balance; and total energy deficit. Measures of health were the numbers of occurrences of each of the following during lactation: all udder problems, mastitis, all locomotive problems, laminitis, digestive problems, and reproductive problems. Reproductive traits were the number of days to first observed estrus and number of inseminations. Several significant relationships between energy balance and health were observed. Increased digestive and locomotive problems were associated with longer and more extreme periods of negative energy balance. PMID- 11104290 TI - Bayesian analysis of lactation curves of Holstein-Friesian cattle using a nonlinear model. AB - A Bayesian procedure was developed for fitting Wood's incomplete Gamma function to test-day milk records of Spanish Holstein Friesian cattle. Each parameter of Wood's function was considered as a dependent variable in a submodel that accounted for systematic effects and genetic relationships among animals. Marginal posterior distributions of model parameters were obtained using Gibbs sampling. Variables of economic interest, such as 305-d yield, persistency, peak yield, and days in milk at peak day were predicted as functions of Wood's function curve parameters. Heritability estimates were 0.26, 0.32, and 0.19 for parameters of Wood's function and 0.26, 0.14, 0.26, and 0.05 for 305-d yield, persistency, peak yield, and days in milk at peak yield. These estimates indicate that it is possible to modify the shape of the lactation curve through genetic selection. Genetic correlations between parameters of Wood's curve and the aforementioned functions of these parameters suggest that selection for 305-d milk yield would result in higher and later peak yield, but only a slight improvement in persistency is expected. PMID- 11104291 TI - Genetic variation of susceptibility to Mycobacterium avium subsp. paratuberculosis infection in dairy cattle. AB - Paratuberculosis is an infectious disease that is not easily amenable to classical control methods such as treatment and vaccination. Experimental animal models suggest that there could be genetic factors responsible for susceptibility or resistance to infection with the causative agent, Mycobacterium avium subsp. paratuberculosis. The aim of this study was to estimate genetic variation in susceptibility to paratuberculosis in Dutch dairy cattle. Data collected during a vaccination trial, conducted from 1984 to 1994, was used. A total of 3020 cows, with complete pedigree records and infection status at slaughter, were available for analysis. A standard polygenic statistical probit model was used to estimate heritabilities. The estimated heritability of susceptibility to M. avium. subsp. paratuberculosis infection was 0.06 for the overall population. In the subpopulation of vaccinated animals the estimated heritability was 0.09. Other calculations based on the model used in this study argue against a prominent role for vertical transmission. Because the establishment of genetic variation is one of the first steps towards the exploration of the possible use of selection for genetic improvement, the present study provides evidence for the presence of genetic variation in the susceptibility of cattle to paratuberculosis. Because the economic impact of the disease is substantial, the development and application of genetic tools, along with other control methods, could be instrumental in the eradication of paratuberculosis. PMID- 11104292 TI - Comparison of models for describing the lactation curve of latxa sheep and an analysis of factors affecting milk yield. AB - The objectives of this work were 1) to compare the goodness-of-fit of empirical models of the lactation curve and 2) to analyze the factors affecting the shapes of the lactation curves, the parameters describing them, and the overall milk yield of Latxa dairy sheep. A total of 14,699 records from 2711 ewes, collected during three consecutive years (1995 to 1997) by the milk recording program of the Latxa ewe of the Basque Country (Spain), were used. Six mathematical models and three fitting procedures were compared. The estimation of model parameters by nonlinear fitting procedures was superior to that by linear regression methods. A nonlinear variable decay model fitted the data better than the other models, as judged by lower mean square prediction error, residual sums of squares, and a lack of first-order positive autocorrelation as assessed by the Durbin Watson coefficient. The effects of the flock, flock-year interaction, month of lambing, length of lactation nested within month of lambing, parity, and number of live lambs born had significant effects on the parameters of the model and the total milk yield (P < 0.01). The prediction of milk yield from the selected model was similar to the estimates obtained with the Fleischmann method currently used by the national breeding program for the Latxa breed. PMID- 11104293 TI - "Airway management defines the specialty of emergency medicine". PMID- 11104294 TI - Migraine and female hormones--what factors increase the risk of ischemic stroke? PMID- 11104295 TI - Evaluation of treatment efficacy of Raynaud phenomenon by digital blood pressure response to cooling. Raynaud's Treatment Study Investigators. AB - Our previous studies have suggested that digital blood pressure response to cooling could provide a measure of the efficacy of treatments that are administered to patients with Raynaud phenomenon (RP). This method was used on 158 primary RP patients participating in a multicenter, randomized clinical trial that compared the efficacy of sustained-release nifedipine with temperature biofeedback in the treatment of RP. A pill placebo and electromyography served as controls. The response to local finger cooling was measured at 30 degrees, 20 degrees, 15 degrees and 10 degrees C in a temperature-controlled room under standardized conditions. The results showed that, at the 15 degrees C and 10 degrees C local cooling temperatures, the patients in the nifedipine group had a higher mean digital systolic blood pressure, a higher relative digital systolic blood pressure (RDSP), a smaller proportion of subjects with RDSP < 70% and a smaller proportion of subjects with a zero reopening pressure than the patients in the three other treatment groups. These results were statistically significant at 10 degrees C, the nifedipine group being significantly different from all others (p < 0.05); no significant difference was found between the three other treatment groups. PMID- 11104296 TI - Spurious systolic hypertension in youth. AB - Six young men diagnosed with systolic hypertension had normal carotid pressure wave contours, normal synthesized aortic pressure wave contours and normal diastolic and mean pressures in upper limb arteries. Elevated brachial systolic pressure was caused by a high narrow systolic peak of the pressure wave. This was attributed to amplification of the pressure wave between the ascending aorta and upper limb (radial and brachial) arteries that is associated with attainment of full body length and very distensible arteries. These young men were not truly hypertensive. Exaggeration of the upper limb systolic peak represented an extreme of the normal pressure wave pattern in youth, where amplification is greater than in childhood or in older subjects. This phenomenon accounts for the rapid increase in systolic pressure between the ages of 5 and 20 years, and the relative plateau in systolic pressure between the ages of 20 and 45 years that is seen in population studies. PMID- 11104297 TI - Differential expression of nitric oxide by dermal microvascular endothelial cells from patients with scleroderma. AB - Vascular abnormalities in scleroderma are fundamental to the pathogenesis of this disease. The objective of this study was to characterize dermal microvascular endothelial cells (DMEC) isolated from scleroderma patients with respect to growth and expression of the constitutive form of endothelial nitric oxide synthase (eNOS). DMEC from patients with both systemic sclerosis (SSc) and localized scleroderma (Loc Scl) contained small intact microvascular structures in contrast to single cell isolations obtained from control skin. Immunoaffinity selection on anti-PECAM-1 beads yielded pure populations of DMEC expressing normal markers. While the morphology and initial growth of SSc DMEC closely paralleled control cells, the growth of SSc DMEC decreased with time in culture (doubling time of 3 days vs. 5 days). Expression of ecNOS mRNA was reduced in both Loc Scl and SSc as shown by semi-quantitative RT-PCR (p < 0.001). Western blots showed variable but generally lower ecNOS protein levels and decreased levels of nitrogen oxides in media were found from both SSc and Loc Scl relative to control cells. The results indicate an intrinsic defect in the mechanism of nitric oxide production in DMEC isolated from scleroderma patients and suggest its possible involvement in the pathophysiology of scleroderma. PMID- 11104298 TI - Successful anticoagulation with hirudin in a patient with mesenteric venous thrombosis and multiple coagulation abnormalities. AB - A case of multiple thrombotic diatheses discovered in the setting of mesenteric venous infarction is discussed. The patient had deficiencies of protein C, protein S, antithrombin III; was heterozygous for factor V Leiden; and had polycythemia vera. Adequate anticoagulation could not be established with heparin administration and hirudin was used. The diagnosis of mesenteric venous infarction, thrombotic tendency of multiple coagulation diatheses, and use of hirudin are discussed. PMID- 11104299 TI - Predicting plaque rupture: enhancing diagnosis and clinical decision-making in coronary artery disease. AB - Atherosclerosis is the process underlying coronary artery disease, myocardial infarction and cerebrovascular disease and is a leading cause of morbidity and mortality in industrialized countries. The atherosclerotic plaque is often indolent and progressive and may destabilize without warning. Components of the atherosclerotic plaque, including structural, cellular and molecular characteristics, determine its vulnerability to rupture. The imaging techniques currently available utilize invasive and non-invasive methods to characterize coronary artery stenoses. Detection, however, usually occurs late in the course of disease after symptoms have presented. Much effort has recently been directed at early detection and in defining markers of atherosclerotic disease. Our challenge for the future is to find non-invasive imaging modalities that can predict plaque vulnerability before irreversible damage has occurred. Through early detection and a targeted treatment strategy we hope to reduce the burden of ischemic heart disease. PMID- 11104300 TI - Protein kinase C activation and its pharmacological inhibition in vascular disease. AB - Vascular complications in diabetes mellitus are known to be associated with the activation of the protein kinase C (PKC) pathway through the de novo synthesis of diacylglycerol (DAG) from glycolytic intermediates. Specific PKC isoforms, mainly the beta- and delta-isoforms, have been shown to be persistently activated in diabetic mellitus. Multiple studies have reported that the activation of PKC leads to increased production of extracellular matrix and cytokines, enhances contractility, permeability and vascular cell proliferation, induces the activation of cytosolic phospholipase A2 and inhibits the activity of Na+-K+ ATPase. These events are not only frequently observed in diabetes mellitus but are also involved in the actions of vasoactive agents or oxidative stress. Inhibition of PKC by two different kinds of PKC inhibitors - LY333531, a selective PKC-beta-isoform inhibitor, and vitamin E, d-alpha-tocopheron - were able to prevent or reverse the various vascular dysfunctions in vitro and in vivo. Clinical studies using these compounds are now ongoing to evaluate the significance of DAG-PKC pathway activation in the development of vascular complications in diabetic patients. PMID- 11104301 TI - Surgical management of abdominal aortic aneurysm. AB - Abdominal aortic aneurysms (AAA) are increasingly common in the aging population. While the etiology of abdominal aortic aneurysms is unknown, there is growing evidence that suggests an immune response. The majority of AAA are asymptomatic and when treated are standard open surgical procedures. The overall mortality rate is 5% or less. The current recommendations for the treatment of aneurysms are based on diameter: diameters exceeding 5 cm in good-risk younger patients should be treated. Aortic aneurysms tend to enlarge over time with a growth-rate between 0.2 and 0.4 mm per year. Once rupture occurs mortality is estimated to exceed 75%, with half of the patients dying prior to arriving at the hospital and the remaining one-half following surgical correction. Recently, minimally invasive techniques have been developed to treat AAA in high-risk patients. These techniques involve the use of covered stented grafts. Current clinical investigations are underway both in this country and in Europe, which have yielded promising results. However, long-term complications are unknown. Currently, aortic aneurysms are best treated with open surgical management. PMID- 11104302 TI - Temporal patterns of injury during a rugby season. AB - The aim of this study was to describe temporal patterns in the frequency, nature and circumstances of injuries occurring among a cohort of 356 rugby players during a club rugby season in New Zealand. It was found that the rate of injury in games decreased significantly over time in both males and females. The reduction in injury rate over the season was more pronounced in some grades, but no differences were found when examined by gender. playing position, age, ethnicity or by health and fitness types. Trends in injury rate were consistent over the rugby season and did not appear to be the result of a bias involving under-reporting of end-of-season injuries. The types and severity of injury remained relatively constant, but the proportion of injuries occurring in back play fell significantly over the season and injuries were more likely to occur in the trunk body region as the season progressed. This study supported the hypothesis that higher rates of injury occur at the start of the rugby season and decrease over the course of the season. This reduction is consistent over time and across player types, and is not attributable either to decreasing injury severity or to increasing player fitness. PMID- 11104303 TI - The relationship between coaching behaviours and sport anxiety in athletes. AB - Previous research has identified the relationship between athlete sport anxiety and various sport outcomes (e.g., performance and dropout). For the majority of athletes involved in sport, the coach is an influential element of the competitive experience. Two hundred and twenty-eight athletes from 15 sports, completed the Sport Anxiety Scale (SAS) and the Coaching Behavior Scale for Sport (CBS-S). The predictive ability of athletes' perceived frequency of seven coaching behaviours (physical training, mental preparation, goal setting, technical skills, competition strategies, personal rapport and negative personal rapport) on four forms of sport anxiety (total anxiety, somatic anxiety, concentration disruption and worry) was examined. Results indicate that negative personal rapport was a significant predictor of all measured forms of sport anxiety while competition strategies was a significant predictor for total anxiety, concentration disruption, and worry. Other behaviours were not significant. The findings suggest that negative rapport between coach and athlete is an important contributor to athlete anxiety. In addition, behaviours that the coach demonstrates relative to competition can be influential in reducing athlete anxiety. PMID- 11104304 TI - Risk reduction in diving spinal cord injury: teaching safe diving skills. AB - Thirty-four recreational swimmers underwent an intervention program to improve diving skills. Participants with low diving skills completed seven 10-minute sessions which emphasised locking thumbs and holding arms extended beyond the head, and steering and gliding skills. Various dive entries were video-recorded and maximum depth reached was used as the criterion measure. A one-way repeated measures analysis of variance was conducted for each dive condition. Maximum depth decreased for all dives. Velocity at maximum depth was greater for the Treadwater, Deck and Block conditions. Improved streamlining and increased 'spring' were evident in more confident participants. Hands separated in 71% of pre-intervention dives but only in 3% of post-intervention dives. Preintervention, arms were pulled backward before. or at, maximum depth in 30% of participants but none did this post-intervention. Diving skills were improved following participation in the intervention program. PMID- 11104305 TI - Time and motion analysis of the AFL field umpire. Australian Football League. AB - The purpose of the present study was to quantify the movement patterns and work intensities of field umpires while officiating in the three-umpire system of the Australian Football League (AFL). Five umpires were randomly selected and videoed throughout five AFL-matches played at the Brisbane Cricket Ground. Each video was analysed manually for time spent in each of four movement modalities: forward, backward, sideways and stationary which were further analysed into the categories of forward sprinting, forward cruising/jogging and forward walking; backward fast/moderate and backward slow; sideways movement and stationary. The following calculations were made: a) the total time spent performing each movement modality; b) the relative contribution (%) of time spent in each activity; and c) the work to rest ratio. The relative time contribution of each movement modality was: Sprinting (1.9+/-0.2%), Cruising/Jogging (26.1+/-3.2%), Walking (21.9+/ 3.1%). Fast/Moderate Backward (14.6+/-1.2%), Slow Backward (13.6+/-1.0%), Sideways (2.2+/-0.3%) and Stationary (19.7+/-2.7%). The average time of effort for each movement modality were found to be: Sprinting (2.2+/-0.4 secs), Cruising/Jogging (6.09+/-1.3 secs), Walking (9.9+/-1.1 secs), Backward Fast/Moderate (4.4+/-0.3 secs), Backward Slow (5.2+/-0.8 secs), Sideways (1.7+/ 0.1 secs) and Stationary (7.4+/-1.4 secs). The average work to rest ratio was approximately between 1:4-1:5. The current findings provide a detailed description of the movement patterns and work intensities of AFL field umpires which may be used in the development of training programs specific to the three umpire system. PMID- 11104306 TI - Personal risk factors associated with injury among female recreational ice hockey players. AB - INTRODUCTION: Women's ice hockey is a rapidly growing sport, however little is known about the injuries sustained by this group of athletes. PURPOSE: The objective of this research was to identify risk factors associated with injury among female recreational ice hockey players. METHODS: This prospective study followed players from two women's ice hockey leagues in Edmonton, Canada during the 1997-98 hockey season. The occurrence of injuries was monitored during the season through standardized telephone follow-up. Risk factors were determined using multiple logistic regression. RESULTS: The initial study sample consisted of 314 players, however as the season progressed 19 (6%) were lost to follow-up. The results of the study are based on 295 (94%) participants. A total of 125 injuries were reported; the injury rate was 7.5 injuries/1,000 player-exposures. Risk factors found to be significantly related to the occurrence of injury were: injury in the past year (OR= 1.57), more than 5 years of hockey experience (OR=1.49), and high exposure level (OR=1.41). CONCLUSION: This research is the first to quantify personal risk factors associated with injury among female recreational ice hockey players. A sports injury in the previous 12 months appears to be highly associated with injury and further research is required to more fully understand this relationship. The importance of controlling for level of exposure when investigating risk factors for sports injury was demonstrated. PMID- 11104307 TI - Does childhood and adolescence provide a unique opportunity for exercise to strengthen the skeleton? AB - Osteoporosis is a major, and increasing, public health problem. In this review we examine the evidence that childhood physical activity is an important determinant of bone mineral in adult years, and as such, may help to prevent osteoporosis. Animal studies provide incontrovertible evidence that growing bone has a greater capacity to add new bone to the skeleton than does adult bone. Observational studies in children undertaking routine physical activity and cross-sectional athlete studies in young sportspeople both reveal that activity is positively associated with bone mineral density (BMD). Longitudinal studies in pre- and peripubertal gymnasts reveal BMD gains far in excess of those that can be achieved in adulthood. However, such studies permit only limited conclusions as they contain the potential for selection bias and can be confounded by other determinants of bone mineral (e.g. dietary and lifestyle factors). Thus, research comparing inter-individual playing-to-nonplaying arm differences in bone mineral (e.g., in racquet sports) have proven to be extremely useful. These studies suggest that the BMD differences are clearly greater when bone is subjected to mechanical loading prior to the end of puberty and longitudinal growth of the body (in women, before menarche) rather than after it. Tanner stage II and III appears to be the maturational stage when the association between exercise and BMD becomes manifest in most adolescents. Do conclusions drawn from athlete studies apply to the general population? Randomised intervention studies of physical activity and bone mineral accrual in normal children confirm that childhood activity is strongly associated with bone mineral accrual. Furthermore, some retired athlete studies and a detraining study suggest that adolescent bone gain may, at least partly, persist despite reduced adult physical activity. Mechanisms that may underlie the association between childhood physical activity and bone mineral accrual are outlined. Thus, it appears that physical activity during the most active period of maturity (with respect to longitudinal growth of the body) plays a vital role in optimising peak bone mass and that benefits may extend into adulthood. PMID- 11104308 TI - The effects of breathing supplemental oxygen during altitude training on cycling performance. AB - To compare the training effects of doing high intensity intervals at 1,840 m in a normoxic vs. hyperoxic environment, eight cyclists (NORM) performed intervals on ergometers 3d/wk while breathing normoxic gas (P1O2 = 128 Torr), and seven subjects (HYPER) performed identical intervals at the same relative workload while breathing hyperoxic gas (P1O2 = 156 Torr). HYPER subjects were able to train at a higher percentage of their altitude lactate inflection point than were NORM subjects (HYPER = 126+/-2%, NORM = 109+/-3% p<0.05). Improvements in power output at maximal steady state (NORM = 8 W, HYPER = 20 W,) and improvement in time to complete a 120 kJ cycling performance test (NORM = 2 s, HYPER = 15 s) were significant in the HYPER group pre- vs. post-training (p<0.05) while the NORM group exhibited no significant changes. No significant changes in power output at lactate inflection point were seen in either group (NORM = -12 W, HYPER = +11 W). The results demonstrate that while training at moderate altitude, breathing hyperoxic gas vs. ambient air allows for higher training intensities and this higher intensity training results in significant improvements in maximal steady state power output and time to complete a 120 kJ performance test. PMID- 11104309 TI - Correlations between physiological parameters and performance in elite ten-pin bowlers. AB - The increasing acceptance of ten-pin bowling as a sport, as well as the keen competition amongst bowlers, necessitates the identification of performance indicators to aid training. The aim of this cross-sectional study was to determine if age, height, weight, aerobic power index, bowling grip strength, 10 RM leg press performance, and the sit-and-reach distance correlated with bowling performance in 42 elite bowlers (26 males and 16 females). At the same time, the physiological profiles of bowlers classified as heavy ball strokers, heavy ball crankers, and light ball spinners were compared. The results showed that for the male bowlers, none of the parameters correlated with performance, while for the female bowlers, the only parameter that correlated with performance was the aerobic power index. Bowlers using the three different releases had similar anthropometric and physiological profiles. The implications from this study are that bowlers of diverse age and build can be equally competitive in the sport; that aerobic capacity (as reflected by the aerobic power index) may, to a certain extent, contribute to bowling performance; and that strength and flexibility measures do not seem to be useful performance indicators amongst elite bowlers. PMID- 11104310 TI - Effects of heat stress on physiological responses and exercise performance in elite cyclists. AB - This study examined the effect of heat stress on physiological responses and exercise performance in elite road cyclists. Eleven members of the Australian National Road Cycling Squad completed two 30 min cycling time-trials in an environmental chamber set at either 32 degrees C, (HT) or 23 degrees C (NT) with a relative humidity of 60% in each circumstance. The trials were separated by two days, with six subjects performing HT first. Power output was 6.5% lower (P<0.05) during HT compared with NT. Mean skin temperature and sweat rate were higher (P<0.05) in HT compared with NT. In contrast, rectal temperature was remarkably similar throughout each trial. During the first 10 min of exercise in HT when power output was not different between trials, blood lactate was higher (P<0.05), and blood pH lower (P<0.05). In contrast, during the last 10 min of exercise when power output was reduced (P<0.05), blood lactate was lower (P<0.05), and pH higher (P<0.05), in HT. These data indicate that heat stress is associated with a reduced power output during self-paced exercise in highly trained men. This decrease in performance appears to be associated with factors associated with body temperature rather than metabolic capacity. PMID- 11104311 TI - Masking effects of social desirability response set on relations between psychosocial factors and sport injuries: a methodological note. AB - Social desirability has long been viewed as a potential source of error variance in self-report measures. We suggest that social desirability (whether in the form of impression management or self-deception) has the capacity to mask relations between psychosocial variables and sport-related outcome or criterion measures that are not measured by selfreport. To illustrate what can occur, we present data from a longitudinal study in which life stress and psychological coping skills were studied as predictors of behaviorally-defined athletic injuries. When data from the entire sample of 352 athletes were analyzed, virtually no injury variance was accounted for by life stress, psychological coping skills, or their interaction. In contrast, deletion from the sample of athletes with high social desirability response set scores resulted in significant predictive relations involving both life stress and coping skills, as well as a significant moderator effect for coping skills. We propose that social desirability masking effects can significantly increase the likelihood of Type II errors in sports medicine research that involves self-report measures, and that social desirability responding needs to be controlled or minimized. PMID- 11104312 TI - The 'price' of Olympic Gold. AB - In 1981 the Commonwealth Government established the Australian Institute of Sport (AIS). The Australian Sports Commission (ASC) which administers the AIS has 2 objectives: (1) excellence in sports performances; and (2) increased participation in sports and sports activities. State-based institutes of sport have also been established with the same or very similar objectives. Federal policy directs the bulk of the ASC budget to elite athlete programs. A smaller proportion goes towards community participation. The official reason is based on the notion of the 'trickle-down' or 'demonstration' effect. That is, a flow-on of benefits to the broader community in the form of increased participation as a direct result of elite sports success. The aims of this study were to determine the (1) spending pattern to elite sports programs for the 5 Olympics 1976/77 to 1995/96, (2) evidence for the two ASC objectives having been met, and (3) expected medal tally at the 2000 Olympic Games. Results show funding (in 1998 dollars), has accelerated from about $1.2 million (1976/77) to $106 million in (1997/98), particularly since the Games were awarded to Sydney. The total amount spent on elite athletes was $0.918 billion. In the period 1980-96 Australia won 25 gold and 115 total Olympic medals. This equates to approximately $37 million per gold and $8 million per medal in general. There was a significant linear relationship between money spent and total medals won. This was also found when all medal types were analysed independently. The predicted medal tally in 2000 (based on the cost per medal and the expenditure towards Sydney) indicates the medal count will be about 14+/-1 gold, 15+/-2 silver and 33+/-4 bronze. Based on our nation's record of international sporting achievement, there is little doubt we have fulfilled the ASC's first objective. Current data on physical activity patterns of Australians suggest the second objective has not been met. Focusing attention on and achieving the first objective does not appear to have any bearing on the second objective. It is time to revisit the notion that elite sporting success leads to greater mass participation as a result of the so-called 'trickle-down' effect. PMID- 11104313 TI - Psychosocial treatment strategies in the MTA study: rationale, methods, and critical issues in design and implementation. AB - The Collaborative Multimodal Treatment Study of Children with Attention Deficit Hyperactivity Disorder (ADHD), the MTA, is the first multisite, cooperative agreement treatment study of children, and the largest psychiatric/psychological treatment trial ever conducted by the National Institute of Mental Health. It examines the effectiveness of Medication vs. Psychosocial treatment vs. their combination for treatment of ADHD and compares these experimental arms to each other and to routine community care. In a parallel group design, 579 (male and female) ADHD children, aged 7-9 years, 11 months, were randomly assigned to one of the four experimental arms, and then received 14 months of prescribed treatment (or community care) with periodic reassessments. After delineating the theoretical and empirical rationales for Psychosocial treatment of ADHD, we describe the MTA's Psychosocial Treatment strategy applied to all children in two of the four experimental arms (Psychosocial treatment alone; Combined treatment). Psychosocial treatment consisted of three major components: a Parent Training component, a two-part School Intervention component, and a child treatment component anchored in an intensive Summer Treatment Program. Components were selected based on evidence of treatment efficacy and because they address comprehensive symptom targets, settings, comorbidities, and functional domains. We delineate key conceptual and logistical issues faced by clinical researchers in design and implementation of Psychosocial research with examples of how these issues were addressed in the MTA study. PMID- 11104314 TI - Behavioral versus behavioral and pharmacological treatment in ADHD children attending a summer treatment program. AB - As part of the behavioral treatment in the Multimodal Treatment Study of Children with ADHD (MTA9), children participated in an intensive summer treatment program (STP). This study examined the differences between 57 children in the combined treatment (Comb) group, who were medicated, and 60 children in the behavioral treatment (Beh) group, who were unmedicated throughout the STP. Comb children were significantly better than Beh on 5 measures: rule following, good sportsmanship, peer negative nominations, and STP teacher posttreatment ratings of inattention/overactivity. Groups did not differ on any of the other 30 measures, and responded similarly to the STP over time. Comparisons to normative data revealed that Comb children were more likely to fall within the normative range on 6 measures. The differences between these results and the main MTA results, in which Comb was always superior to Beh, are discussed in terms of the relative intensity of combined treatments. The implications for future studies of pharmacological and behavioral treatment for ADHD are discussed. PMID- 11104315 TI - Anxiety as a predictor and outcome variable in the multimodal treatment study of children with ADHD (MTA). AB - Initial moderator analyses in the Multimodal Treatment Study of Children with ADHD (MTA) suggested that child anxiety ascertained by parent report on the Diagnostic Interview Schedule for Children 2.3 (DISC Anxiety) differentially moderated the outcome of treatment. Left unanswered were questions regarding the nature of DISC Anxiety, the impact of comorbid conduct problems on the moderating effect of DISC Anxiety, and the clinical significance of DISC Anxiety as a moderator of treatment outcome. Thirty-three percent of MTA subjects met DSM-III R criteria for an anxiety disorder excluding simple phobias. Of these, two-thirds also met DSM-III-R criteria for comorbid oppositional-defiant or conduct disorder whereas one-third did not, yielding an odds ratio of approximately two for DISC Anxiety, given conduct problems. In this context, exploratory analyses of baseline data suggest that DISC Anxiety may reflect parental attributions regarding child negative affectivity and associated behavior problems (unlike fearfulness), particularly in the area of social interactions, another core component of anxiety that is more typically associated with phobic symptoms. Analyses using hierarchical linear modeling (HLM) indicate that the moderating effect of DISC Anxiety continues to favor the inclusion of psychosocial treatment for anxious ADHD children irrespective of the presence or absence of comorbid conduct problems. This effect, which is clinically meaningful, is confined primarily to parent-reported outcomes involving disruptive behavior, internalizing symptoms, and inattention; and is generally stronger for combined than unimodal treatment. Contravening earlier studies, no adverse effect of anxiety on medication response for core ADHD or other outcomes in anxious or nonanxious ADHD children was demonstrated. When treating ADHD, it is important to search for comorbid anxiety and negative affectivity and to adjust treatment strategies accordingly. PMID- 11104317 TI - Family processes and treatment outcome in the MTA: negative/ineffective parenting practices in relation to multimodal treatment. AB - To elucidate processes underlying therapeutic change in a large-scale randomized clinical trial, we examined whether alterations in self-reported parenting practices were associated with the effects of behavioral, medication, or combination treatments on teacher-reported outcomes (disruptive behavior, social skills, internalizing symptoms) in children with attention-deficit hyperactivity disorder (ADHD). Participants were 579 children with Combined-type ADHD, aged 7 9.9 years, in the Multimodal Treatment Study of Children with ADHD (MTA). We uncovered 2 second-order factors of parenting practices, entitled Positive Involvement and Negative/Ineffective Discipline. Although Positive Involvement was not associated with amelioration of the school-based outcome measures, reductions in Negative/Ineffective Discipline mediated improvement in children's social skills at school. For families showing the greatest reductions in Negative/Ineffective Discipline, effects of combined medication plus behavioral treatment were pronounced in relation to regular community care. Furthermore, only in combination treatment (and not in behavioral treatment alone) was decreased Negative/Ineffective Discipline associated with reduction in children's disruptive behavior at school. Here, children in families receiving combination treatment who showed the greatest reductions in Negative/Ineffective Discipline had teacher-reported disruptive behavior that was essentially normalized. Overall, the success of combination treatment for important school-related outcomes appears related to reductions in negative and ineffective parenting practices at home; we discuss problems in interpreting the temporal sequencing of such process-outcome linkages and the means by which multimodal treatment may be mediated by psychosocial processes related to parenting. PMID- 11104316 TI - Parenting and family stress treatment outcomes in attention deficit hyperactivity disorder (ADHD): an empirical analysis in the MTA study. AB - Parenting and family stress treatment outcomes in the MTA study were examined. Male and female (579), 7-9-year-old children with combined type Attention Deficit Hyperactivity Disorder (ADHD), were recruited at six sites around the United States and Canada, and randomly assigned to one of four groups: intensive, multi faceted behavior therapy program alone (Beh); carefully titrated and monitored medication management strategy alone (MedMgt); a well-integrated combination of the two (Comb); or a community comparison group (CC). Treatment occurred over 14 months, and assessments were taken at baseline, 3, 9, and 14 months. Parenting behavior and family stress were assessed using parent-report and child-report inventories. Results showed that Beh alone, MedMgt alone, and Comb produced significantly greater decreases in a parent-rated measure of negative parenting, Negative/Ineffective Discipline, than did standard community treatment. The three MTA treatments did not differ significantly from each other on this domain. No differences were noted among the four groups on positive parenting or on family stress variables. Results are discussed in terms of the theoretical and empirically documented importance of negative parenting in the symptoms, comorbidities and long-term outcomes of ADHD. PMID- 11104318 TI - Parent cognitions as predictors of child treatment response in attention deficit/hyperactivity disorder. AB - Using a subsample of 105 children and their parents (100 mothers, 57 fathers) from the Multimodal Treatment Study of Children with ADHD (the MTA), the value of parents' baseline cognitions as predictors of children's treatment outcome at 14 months was examined. Measures of parents' cognitions about themselves, their ADHD children, and their parenting, as well as a self-report measure of dysfunctional discipline were included. Both mothers' and fathers' self-reported use of dysfunctional discipline predicted worse child treatment outcome. Low self-esteem in mothers, low parenting efficacy in fathers, and fathers' attributions of noncompliance to their ADHD child's insufficient effort and bad mood also were associated with worse child treatment outcome. All of these predictive relations were obtained even after MTA treatment effects had been taken into account. Secondary analyses indicated that mothers had a more external locus of control, lower self-esteem, lower parenting efficacy, and a greater tendency to attribute noncompliance to their ADHD child's bad mood than did fathers. PMID- 11104320 TI - Commentary on the multimodal treatment study of children with ADHD. AB - The multimodal treatment study of attention deficit hyperactivity disorder (MTA Study) constitutes a landmark in the history of treatment research in child psychopathology, being the largest single study of its kind ever undertaken. Important findings have emerged from this project, as the papers in the present volume will attest. This commentary focuses on several concerns about the assumptions that appear to have guided the design of the MTA study, particularly its psychosocial treatment component, as well as the manner in which treatment results have been presented to date. In particular, no explicit theory of ADHD appears to have guided the construction of the treatment components, relying instead on implicit theories associated with those treatments, such as the notion that the symptoms of ADHD arise through faulty learning and defective contingencies of reinforcement. Future articles from this study will need to address these and other concerns if the results of the study are to be properly interpreted and the scientific and clinical yield is to be maximized. PMID- 11104319 TI - Familial aggregation of ADHD characteristics. AB - Patterns of familial aggregation of ADHD symptoms in parents of ADHD and non-ADHD children were examined. Within the ADHD sample, symptom aggregation was examined as a function of biological relationship, parent and child gender, and children's comorbid diagnoses. Participants consisted of parents of 579 children with ADHD, Combined Type participating in the multimodal treatment study of children with ADHD and parents of 288 normal control participants. Adult symptoms of ADHD were measured by both self-report and report of a significant other. Results indicated that the parents of children with ADHD had higher ratings of inattention/cognitive problems, hyperactivity/restlessness, impulsivity/emotional lability, and lower self-concept than parents of children without ADHD on both self-report and other-report ratings. Within the ADHD sample of children, other report ratings of inattention/cognitive problems and impulsivity/emotional lability were higher for biological parents compared to nonbiological parents whereas self-ratings were not related to biological status. These findings support previous research documenting familial aggregation of ADHD and appear to strengthen the hypothesis that there is a genetic contribution to ADHD. PMID- 11104321 TI - A nursing critique of US welfare system reform. AB - In 1996, the Aid to Families with Dependent Children program was repealed, and the welfare system in the United States was changed. This article critiques, from a nursing perspective, US welfare system reform. It interrogates dominant ideologies about poverty, welfare, and waged labor; examines federal welfare reform legislation of the 1990s and its programmatic implementation at the state level; discusses global health and safety implications of welfare replacement initiatives; and challenges nurses to political and scholarly action. PMID- 11104322 TI - Nursing participation in health care reform efforts of 1993 to 1994: advocating for the national community. AB - This report of a postmodern feminist oral history tells a contemporary story of the success of nursing in overcoming the impediments of tradition, organizing and acting as an identifiable group, and speaking out with clarity as advocates for the health of American society. This was an important historical, transitional, and celebratory time for nursing. Continuing advocacy for health care for all Americans requires developing expertise in both traditional and feminist leadership, understanding how political theories and history affect policy development, and active participation in American democracy. Future actions require incorporation of lessons from the recent past. PMID- 11104323 TI - Watson's philosophy, science, and theory of human caring as a conceptual framework for guiding community health nursing practice. AB - Criticisms of existing nursing theories in relation to community health nursing practice are that they focus on individuals and have been developed primarily for practice within the context of infirmity and disease, making them inadequate to guide community health nursing practice. Despite being developed for individuals, Watson's theory is proposed as a nursing framework that is philosophically congruent with contemporary global approaches to community health and health promotion. An overview of her theory identifies the centrality of caring, holism, and ecology in the theory as it has evolved over the past 20 years. Concepts developed for individual-nurse interactions are extrapolated to the community in a discussion of the suitability of the theory to guide community health nursing practice. A community assessment tool based on Watson's theory is provided. PMID- 11104324 TI - From theory to practice: community health nursing in a public health neighborhood team. AB - An interdisciplinary team in a local public health district tested its ability to implement the core public health functions of assessment, policy development, and assurance by changing its practice to a community-driven model of building partnerships for health with groups and communities in a designated locale. Evaluation of this innovation revealed that the public health nurse members of the team enacted their community health nursing knowledge to strengthen agency to cocreate health. Interdisciplinary collaboration was essential to the team's community mobilization efforts. Additional findings suggested that this organizational innovation was associated with developing a more participatory organizational climate, increasing system effectiveness, and building community capacity. PMID- 11104325 TI - Nursing in a technological world: searching for healing communities. AB - A research dialectic between philosophy of technology and nurses' work in acute care surfaces parallel technological practices that threaten the healing nature of two modern projects: health care and ecological restoration. A metaphor of ecological restoration is used to explore the consequences of denatured health care work for the welfare of patients, families, practitioners, and healing communities. It is argued that in health care systems where the mismatch between treatment options and resources for care steadily grows, the nursing discipline must develop ecological literacy for a technological world. PMID- 11104326 TI - Caregiving and care receiving among a technologically dependent heart failure population. AB - This descriptive study investigated health-related quality of life (HRQOL) and caregiving/care receiving among 20 end stage heart failure patients receiving community-based inotropic infusions and among their 18 family caregivers. The analysis revealed that care recipients perceived considerable impairment from their disease process and poor HRQOL despite the use of inotropic infusions. Perceived powerlessness was identified as a predictor of the recipients' mental health status, while caregiver esteem adversely affected recipient HRQOL. Although insufficient preparation to care and caregiving tasks significantly contributed to the negative aspects of care provision. the esteem and mental health of the caregiver significantly enhanced HRQOL among caregivers. PMID- 11104327 TI - Should patients receiving long-term gastric acid inhibition therapy be evaluated for vitamin B12 deficiency? PMID- 11104328 TI - A 65-year-old factory worker with dyspnea on exertion and a normal chest x-ray. AB - Chronic beryllium disease is an occupationally acquired granulomatous lung disease similar to sarcoidosis. It is caused by exposure to beryllium in genetically susceptible persons. It should be suspected in patients with beryllium exposure who present with pulmonary symptoms or have a positive screening blood beryllium-specific lymphocyte proliferation test. The diagnosis is confirmed by the finding of granulomas on transbronchial biopsy in the appropriate clinical and epidemiologic setting. Although there is no cure, treatment with corticosteroids is usually beneficial. In view of the potential side effects, treatment is reserved for patients with symptoms or a decline in pulmonary function. PMID- 11104329 TI - Heart failure is a fever: the cytokine connection. AB - Inflammation probably contributes to the development and progress of heart failure. Proteins called cytokines, which are produced by damaged tissues and leukocytes as part of the inflammatory response, affect the heart both directly and indirectly. They exacerbate hemodynamic imbalances, act as negative inotropes, stimulate left ventricular hypertrophy, and promote the production of still more cytokines, which continue the cycle. Several medications counter these effects in vitro, and some are in clinical testing. PMID- 11104330 TI - In diabetes, treat hidden heart disease. AB - Both diabetic and prediabetic patients have abnormal vascular reactivity and should be considered to have occult cardiovascular disease. Angiotensin converting-enzyme (ACE) inhibitors are particularly beneficial in diabetes because they reduce the incidence of both cardiovascular events and diabetes related complications. In prediabetic patients, ACE inhibitors also reduce the risk of a new diagnosis of type 2 diabetes. Managing hypertension is even more beneficial for diabetic patients than for nondiabetic patients. To further reduce the risk of heart disease in patients with diabetes or prediabetes, dyslipidemia should also be treated aggressively. PMID- 11104331 TI - A 70-year-old farmer with a skin lesion. PMID- 11104332 TI - Pharmacologic aids to smoking cessation. AB - Nicotine replacement and bupropion have shown significant benefits for those seeking pharmacologic assistance with smoking cessation. This discussion reviews the efficacy literature for both treatments, including potential side effects. PMID- 11104334 TI - Treatment of ovarian cancers not of epithelial origin. AB - The ovary can harbor neoplasms other than epithelial ovarian cancer; these include less-common primary cancers of the ovary and cancers that metastasize to the ovary from other parts of the body. Depending on the specific histology, site of origin of a metastatic lesion, extent to which the cancer has spread, and comorbid medical conditions, women with these less-common neoplasms may be candidates for aggressive surgical intervention, systemic chemotherapy, or therapy focused on symptom management and comfort. PMID- 11104333 TI - Demystifying two common genetic predispositions to venous thrombosis. AB - Two recently discovered genetic abnormalities substantially increase the risk of venous thromboembolism. Yet we do not advocate screening for these abnormalities except in cases in which the information gained would affect the course of action. PMID- 11104335 TI - Using troponin T to diagnose acute coronary syndromes. AB - Elevated troponin T is a useful marker for acute myocardial infarction: it is more specific than is elevated creatine kinase MB isoenzyme, and it remains elevated for many days after creatine kinase levels have returned to normal, providing a useful indicator for late presentations. Nevertheless, creatine kinase MB still has many important roles, including providing estimates of infarct size and diagnosing acute myocardial infarction in patients with renal failure. Often, measuring both markers provides additional information. This article provides a diagnostic algorithm for using both markers. PMID- 11104336 TI - Carotid stenosis: current strategies for choosing between medical and surgical management. AB - The effectiveness of carotid revascularization depends on appropriate patient selection and balancing the expected benefits with the risks of treatment. Exceeding a rate of serious complications (strokes and deaths) of 5% for asymptomatic and 9% for symptomatic patients negates any benefit for carotid endarterectomy. Endovascular techniques such as stent-supported angioplasty will likely change the management approach for some patients with carotid occlusive disease. This paper contains the author's recommendations for choosing between medical and surgical management of carotid stenosis. PMID- 11104337 TI - Significance of lipoxygenase-derived monohydroxy fatty acids in cutaneous biology. AB - The skin displays a highly active metabolism of polyunsaturated fatty acids (PUFA). Dietary deficiency of linoleic acid (LA), an 18-carbon (n-6) PUFA, results in characteristic scaly skin disorder and excessive epidermal water loss. Although arachidonic acid (AA), a 20-carbon (n-6) PUFA, is metabolized via cyclooxygenase pathway into predominantly prostaglandin E2 (PGE2) and PGF2alpha. The 15-lipoygenase is very active in this tissue and catalyzes the transformation of 20-carbon AA into predominantly 15-hydroxyeicosatetraenoic acid (15-HETE). Similarly, the epidermal 15-lipoxygenase also catalyzes the transformation of 18 carbon LA and 20-carbon dihomo-gamma-linolenic acid (DGLA) to 13 hydroxyoctadecadienoic acid (13-HODE) and 15-hydroxyeicosatrienoic acid (15 HETrE), respectively. The monohydroxy fatty acids are incorporated in phospholipids which undergo catalysis to yield substituted-diacylglycerols (13 HODE-DAG) and 15-HETrE-DAG) which exert anti-inflammatory/antiproliferative effects on the skin. PMID- 11104338 TI - Cyclooxygenase isoforms in human skin. PMID- 11104339 TI - Arachidonic acid metabolism in skin health and disease. PMID- 11104340 TI - Eicosanoids in inflammatory skin diseases. PMID- 11104341 TI - Of flies, mice, and men. PMID- 11104342 TI - Therapeutic effects of functional electrical stimulation of the upper limb of eight children with cerebral palsy. AB - Functional electrical stimulation (FES) of the upper limb has been used for patients with a variety of neurological conditions, although few studies have been conducted on its use on the upper limb of children with cerebral palsy (CP). The aim of this study was to investigate the effect of cyclic FES on the wrist extensor muscles of a group of eight children (five boys, three girls) with hemiplegic CP (mean age 10 years). The study design involved a baseline (3 weeks), treatment (6 weeks), and follow-up (6 weeks). FES was applied for 30 minutes daily during the treatment period of the study. Improvements in hand function (p < or = 0.039) and active wrist extension (p = 0.031) were observed at the end of the treatment period. These improvements were largely maintained until the end of the follow-up period. No significant change was observed in the measurements of wrist extension moment during the treatment period (p = 0.274). Hand function in this group of children improved after they were exposed to FES of wrist extensor muscles. This suggests that FES could become a useful adjunct therapy to complement existing management strategies available for this patient population. PMID- 11104343 TI - Quantitative assessment of mirror movements in children and adolescents with hemiplegic cerebral palsy. AB - Mirror movements in individuals with hemiplegic cerebral palsy (CP) may result from a reorganization of the central sensorimotor system. Motor performances of both hands were measured to characterize mirror activity (or mirroring) and hand functions in 22 participants (6 to 18 years) with hemiplegic CP and in 17 control participants. During a unimanual repetitive squeezing task, contractions of the active hand and fingertip forces of the opposite hand were recorded simultaneously. In the control group, slight mirror activity (or mirroring) was found that decreased with age. In participants with CP, mirror activity was 15 times stronger than in the control group, and was found at all age levels. Mirroring was more prominent in the unaffected hand of the CP group. The amount of mirror activity was not related to the degree of hemiplegia, which was assessed with measures of spasticity, strength, and dexterity. Mirror movements disturbed functional bimanual skills, although to some extent they could be suppressed by voluntary effort. PMID- 11104344 TI - Macrocephaly in autism and other pervasive developmental disorders. AB - To assess the prevalence of macrocephaly (head circumference > or = 1.88 standard deviations above normative data for age and sex or > 97th centile) in autism and other pervasive developmental disorders, 41 children with autism, and a comparison group of 21 children with tuberous sclerosis complex (TSC) or an unspecified seizure disorder were studied. Familiality of head circumference was also assessed from measurements of 133 first-degree relatives. Significantly higher rates of macrocephaly were found in probands with autism (12.2%) and their first-degree relatives (15.5%) when compared against a published normative sample. The incidence of macrocephaly in the comparison group of probands with TSC and seizure disorder (9.5%) and their first-degree relatives (8.3%) was higher than normative data as well, although the relation between macrocephaly and autism was more pronounced. Head circumference and extreme scores reflecting macrocephaly were moderately heritable in the present sample (H2 = 0.47). The increased prevalence of macrocephaly in relatives of children with autism compared with control children suggests that this characteristic may be a familial risk factor in the pathogenesis of autism. PMID- 11104345 TI - Patterns of neuropsychological deficits in children with medulloblastoma according to craniospatial irradiation doses. AB - This study aimed to analyse the relationship between supratentorial irradiation dose and the intellectual outcome in 36 children (aged between 5 and 15 years) treated for medulloblastoma. The supratentorial radiation dose was reduced to 25 Gy in 23 children and given at the standard dose, 35 Gy, in 13 other children. Neuropsychological evaluation was performed at a mean of 4.3 years (SD 4.7 years) after radiotherapy. The supratentorial radiation dose was the principal risk factor associated with impaired intellectual outcome. Verbal fluency, immediate word list recall, block design, and fine motricity of the dominant hand were significantly lower in children irradiated at the standard doses than in those irradiated at reduced doses. These findings suggest that the dose of radiotherapy applied to the brain strongly influences later verbal and non-verbal skills in children with medulloblastoma. This should be taken into account in treatment planning and in rehabilitation programs. PMID- 11104346 TI - Serum NGF levels in children and adolescents with either Williams syndrome or Down syndrome. AB - The neurotrophin nerve growth factor (NGF) is a major regulator of peripheral and central nervous system development. Serum NGF was measured in normally developing control children (n=26) and in individuals affected by congenital syndromes associated with learning disability: either Williams syndrome (WS; n=12) or Down syndrome (DS; n=21). Participants were assessed at three distinct developmental stages: early childhood (2 to 6 years), childhood (8 to 12 years), and adolescence (14 to 20 years). A sample was taken only once from each individual. Serum NGF levels were markedly higher in participants with WS, than DS and control participants. In addition, different developmental profiles emerged in the three groups: while in normally developing individuals NGF levels were higher in early childhood than later on, children with WS showed constantly elevated NGF levels. When compared to control participants, those with DS showed lower NGF levels only during early childhood. Neuropsychological assessment confirmed previously reported differences among the three groups in the development of linguistic/cognitive abilities. Some features of individuals with WS, such as hyperacusis and hypertension, could be related to high-circulating NGF levels. PMID- 11104347 TI - Can sodium valproate improve learning in children with epileptiform bursts but without clinical seizures? AB - This study aimed to determine whether sodium valproate (VPA) improves cognitive performance and behaviour in children with learning and behavioural problems associated with electrographic epileptiform discharges but without clinical seizures. A randomized, double-blind, single-crossover trial was carried out with VPA or placebo on eight participants with different learning and behaviour problems. Participants also underwent neuropsychological testing under video EEG and the parent and teacher Behaviour Check List (CBCL; Achenbach 1991a, b) during each treatment phase. Clinically none of the children improved on VPA. On formal testing children were more distractable, had increased delay in response time, and showed lower memory scores while on VPA. In addition, parents reported higher internalizing scores on the CBCL while children were on VPA. Our data do not support the use of VPA in similar patients. PMID- 11104348 TI - Outcome of infants with unilateral Sturge-Weber syndrome and early onset seizures. AB - Patients with Sturge-Weber syndrome often present with seizures during the first year of life. Currently, only patients with clinically significant seizures who do not respond to medical treatment are candidates for early epileptic surgery. However, a delay of surgical treatment may result in cognitive deterioration. We studied the correlation between parameters and outcome of seizures to re-examine the criteria for early epilepsy surgery. We performed a retrospective chart review combined with telephone interviews of parents of all Israeli infants with unilateral Sturge-Weber syndrome and early onset seizures, and we examined whether age of seizure onset and seizure intensity were correlated with cognitive level and the degree of hemiparesis at follow-up. We recruited a total of 15 patients with unilateral Sturge-Weber syndrome and early onset seizures, five of whom underwent epilepsy surgery. The mean follow-up period of all the patients was 15 years: six patients had normal intelligence, four had borderline cognitive level, three had mild mental retardation and two had moderate mental retardation. Eight of the ten non-operated patients still experience seizures at follow-up. Cognitive delay was significantly correlated with seizure intensity in the early period, but not with the age of seizures onset, the degree of hemiparesis, or the presence of ongoing seizures. We conclude that high seizure intensity in young patients with Sturge-Weber syndrome is a prognostic marker for mental deterioration. PMID- 11104349 TI - Association of epilepsy with different groups of microcephaly. AB - Sixty-six participants (33 males, 33 females) with microcephaly (MC), age range from 2 to 19 years old, were evaluated. MC was classified pathogenetically into isolated MC (IMC) and multiple MC (MMC) and classified etiologically into primary MC (PMC) and secondary MC (SMC). Both IMC and MMC were further classified. Overall prevalence of epilepsy was 40.9%. Furthermore, there was a significantly higher prevalence of epilepsy in males. Main seizure type was generalized tonic clonic seizures. Generally, learning disability (LD) was diagnosed in 93.9% and profound LD was evident in 43.9% of participants. There was an inverse correlation between severity of epilepsy and IQ but a positive correlation between severity of epilepsy and degree of LD. Differences in the success rate between monotherapy and polytherapy or response to antiepileptic drugs were not observed. Results suggest that epilepsy may be associated with the lower cognitive ability of the participants with microcephaly. The pathogenetic classification proposed is of value in delineating the prevalence of epilepsy and LD in the different varieties of MC as compared with the etiological classification. PMID- 11104350 TI - Monozygotic boys with fragile X syndrome. AB - Monozygotic twin boys with fragile X syndrome underwent thorough genetic, psychiatric, neurological, and language evaluations at 10 years of age. They both demonstrated physical features, speech and language difficulties, social problems, and attentional deficits that characterize the behavioural phenotype of fragile X syndrome. Despite identical genetic constitutions, there were important developmental and behavioural heterogeneities. Twin A showed less social interaction and symbolic play and more speech and language dysfunction than twin B. Twin A also had significantly larger caudate volumes. It is suggested that the Xq27.3 anomaly may not be sufficient to account for all the behavioural phenotypic and neuroanatomical features of fragile X syndrome. PMID- 11104351 TI - Cerebellar ataxia, anterior horn cell disease, learning difficulties, and dystonia: a new syndrome. AB - The following case reports describe a new condition of cerebellar ataxia, anterior horn cell disease, dystonia, and learning difficulties. Four cases are described. The condition appears to be of autosomal recessive inheritance as the group is made up of two pairs of sisters. All cases were evident by 3 years of age. Anterior horn cell disease was of a type not previously described at this age in association with cerebellar ataxia. Further genetic studies suggest the condition is not allelic with spinal muscular atrophy having no evidence of deletion of the survival motor neurone gene. PMID- 11104352 TI - Genetics and the muscular dystrophies. PMID- 11104353 TI - Use of rectal diazepam in the community. PMID- 11104354 TI - Transient cytochrome oxidase deficiency with Ohtahara syndrome. PMID- 11104355 TI - Pseudohypertrophy of the temporalis muscle in Xp21 muscular dystrophy. PMID- 11104356 TI - A high dose of long term treatment with deprenyl loses its effect on antioxidant enzyme activities as well as on survivals of Fischer-344 rats. AB - The survival rate of male Fischer-344/Du rats treated chronically with high doses of deprenyl was investigated. Eighteen month old rats were treated with 1 mg/kg s.c. deprenyl 3 times per week for 13 months. At the age of 31 months, treated rats showed a greater mortality rate with three of 12 rats surviving, while in saline-treated control animals seven of 12 animals survived. No significant differences in superoxide dismutase (SOD) or catalase (CAT) activities in brain regions of control and treated animals were seen at 31 months of age. In contrast, when 27 month old rats were treated in the same manner for one month, significant increases in SOD (both Cu,Zn- and Mn-) and CAT activities were found in substantia nigra, striatum and cerebral cortex, but not in hippocampus. This effect was produced with a wide range of deprenyl doses (0.25-2 mg/kg, but not 4 mg/kg). Although a causal relationship between the two different effects of the drug, i.e. 1) increases in antioxidant enzyme activities and 2) the prolongation of survival of animals, has not been directly demonstrated, the loss of both effects with the high dose of the drug in the present experiment may be taken as circumstantial evidence for their causal relationship. PMID- 11104357 TI - Regulation of inhibin/activin subunits and follistatin mRNA expression in the rat pituitary at early estrus. AB - In the rat pituitary, activin stimulates whereas inhibin prevents FSH synthesis and secretion. Besides, the activin binding protein follistatin neutralizes the action of activin. The control of the FSH secondary surge at early estrus is not completely understood. To investigate the regulation of the inhibin/activin alpha , betaA- and betaB-subunits and follistatin mRNA expression in the pituitary during the time of the FSH secondary surge, cyclic rats treated with LHRH antagonist (ANT) and ovine LH (oLH), progesterone (P), the anti-steroid RU486, adrenalectomy (ADX) or ADX plus corticosterone (B), were killed at early estrus. The serum concentrations of FSH were measured and the mRNA levels of the above mentioned transcripts were analysed and quantitated by using RNase protection assays. ANT abolished the FSH secondary surge and increased mRNA for alpha- and betaA-subunits and follistatin, but reduced that for betaB-subunit. Both oLH and P reversed these effects. RU486 blocked the effect of oLH on FSH levels and prevented the reduction in the mRNA for follistatin. ADX in ANT+oLH-treated rats reduced the serum FSH concentrations, enhanced mRNA for betaA- and betaB-subunits and, similar to RU486, blocked the drop in follistatin mRNA. Finally, replacement of B in ADX animals reversed these effects. These results demonstrate that, in the cyclic rat, the preovulatory secretion of LH and the surges of P and B on proestrus regulate the synthesis of inhibin/activin subunits and follistatin mRNA in the rat pituitary at early estrus, probably by reducing inhibin and follistatin and increasing activin. Moreover, these effects of LH, P and B at the pituitary level, together with the decrease in the amount of inhibin coming from the ovary, might be responsible for the occurrence of the FSH secondary surge. PMID- 11104358 TI - Dietary polyenylphosphatidylcholine decreases cholesterolemia in hypercholesterolemic rabbits: role of the hepato-biliary axis. AB - The aim of this work was to study the cholesterol-lowering mechanisms induced by dietary soybean lecithin in hypercholesterolemic rabbits. Male New Zealand white rabbits (n = 6 in each group) were fed for 10 weeks either a low-fat control C diet, containing 27 g fat/kg, or high-fat diets enriched with 2 g cholesterol/kg and 77 g fat/kg. The high-fat diets contained 50 g lard (L), 50 g soybean triacylglycerol (SO), or 50 g pure soybean phosphatidylcholine (PLE). PLE diet decreased by 30% beta-VLDL-cholesterol, compared with SO diet. HDL2-, HDL3- and LDL-lipid contents were unchanged in the L, SO and PLE groups. In gallbladder bile, amounts of phospholipids, bile salts and cholesterol were significantly increased in PLE group by respectively 45%, 11% and 44%, in comparison with SO group. Intestinal and hepatic Hydroxy Methyl Glutaryl Coenzyme A reductase activities were not increased by PLE diet. Triacylglycerol hepatic content was lower in PLE group than in L or SO groups. Compared with triacylglycerol enriched diet, phosphatidylcholine enriched diet developed significant higher cholesterol- and triacylglycerol-lowering effects by a two-step mechanism: i) by reducing the beta-VLDLs, ii) by enhancing the secretion of bile cholesterol. Such results constitute promising effects of soybean phosphatidylcholine at the hepato-biliary level, in the treatment or prevention of hyperlipidemia and related atherosclerosis. PMID- 11104359 TI - Pain threshold, learning and formation of brain edema in mice lacking the angiotensin II type 2 receptor. AB - The main biological role of angiotensin II type 2 receptor (AT2) has not been established. We made use of targeted disruption of the mouse AT2 gene to examine the functional role of the AT2 receptor in the central nervous system (CNS). We have previously shown that AT2-deficient mice displayed anxiety-like behavior in comparisons with wild-type mice. In the present study, we analyzed the pain threshold, learning behavior and brain edema formation using the tail-flick test, the tail-pinch test, the passive avoidance task and cold injury, respectively. In the passive avoidance task and cold injury, no differences were found between wild-type mice and AT2-deficient mice. In contrast, the pain threshold was significantly lower in AT2-deficient mice, compared with findings in wild-type mice. The immunohistochemical distribution of beta-endorphin in the brain was analyzed quantitatively in AT2-deficient mice and wild-type mice, using microphotometry. The fluorescence intensity of beta-endorphin in the arcuate nucleus of the medial basal hypothalamus (ARC) was significantly lower in AT2 deficient mice, compared with findings in wild-type mice. We found that the AT2 receptor does not influence learning behavior and brain edema formation. As AT2 deficient mice have increased sensitivity to pain and decreased levels of brain beta-endorphin, AT2 receptors may perhaps mediate regulation of the pain threshold. PMID- 11104360 TI - Mechanism of wortmannin-induced inhibition of secretory responses in rat adrenal medullary cells. AB - The effect of wortmannin (WT), an inhibitor of myosin light chain kinase (MLCK) as well as PI3-kinase, on catecholamine (CA) secretion in the perfused rat adrenal medulla was studied. After a 35-min application of 10 microM WT, secretory response to repetitive stimulation with 30 mM extracellular K+ was reduced to 30% of that obtained with intact medullae, and the response to 200 nM bradykinin (BK) was almost completely abolished. Aiming to identify the target for the WT effect, the WT derivative KT7692, which retains the same potency to inhibit MLCK as that of WT but its potency to PI3-kinase is one-hundredth that of WT, was used. Unlike WT, KT7692 at 10 microM did not affect the high-K+-evoked secretion and slightly potentiated the BK-evoked secretion. These results oppose the notion that WT inhibits the secretory responses through inhibition of MLCK. However, the alternative idea, that PI3-kinase is a target for WT, is also difficult to accept since WT concentrations required for the inhibition of the secretions are much higher than those needed to inhibit PI-3-kinase by WT. PMID- 11104361 TI - Differential effects of hypoxia on untreated and NGF treated PC12 cells. AB - Perinatal hypoxia is known to induce long-lasting changes in the central dopaminergic system. In order to understand the cellular mechanism of these changes, we studied the effects of hypoxia on the levels of dopamine (DA) and tyrosine hydroxylase (TH) mRNA in untreated and NGF treated PC12 cells. On the second day after plating (DAP), cells were exposed to a hypoxic episode (pO2 = 10 20 mm Hg, 24 h), and the levels of DA and TH mRNA were examined on DAP 4 and DAP 8. In untreated cells, hypoxia induced a two fold increase both in DA and TH mRNA levels on DAP 4 which normalized up to DAP 8. This increase correlated with an activation of the hypoxia inducible factor (HIF-1alpha), measured with a reporter gene. In contrast, NGF treated cells responded to hypoxia with an increase of DA level on DAP 8. In these cells neither an increase of the HIF-1alpha activity measured immediately after hypoxia nor a significant increase of the TH mRNA level on DAP 8 were found. The findings indicate that NGF shifts the hypoxia induced changes of DA levels from a short-term to a long-term mode. The long-term increase of dopamine levels is the most likely result of changes connected with cell growth and differentiation and not the result of a long-term TH mRNA level increase. PMID- 11104362 TI - Stretch-induced morphological changes of human endothelial cells depend on the intracellular level of Ca2+ rather than of cAMP. AB - When exposed to a uni-axial cyclic stretch, cultured human umbilical vein endothelial cells (HUVECs) align and elongate perpendicular to the stretch axis. Previous studies showed that forskolin inhibited stretch-induced orientation of endothelial cells, suggesting that adenosine 3:5-cyclic monophosphate (cAMP) plays an important role in the shape change. However, we have recently shown that stretch-induced shape changes in cultured HUVECs are due to increased [Ca2+]i. In the present study, we examined the possible role of cAMP in stretch-induced shape changes in cultured HUVECs. Application of uni-axial cyclic stretch induced a gradual rise in cAMP reaching a peak level at 60 min after the onset of stretch. The adenylate cyclase activator, forskolin, increased the basal level of cAMP but inhibited the rise in [Ca2+]i resulting in no cell shape changes. In contrast, N 6,2-dibutyryladenosine 3:5-cyclic monophosphate (dbcAMP) enhanced the stretch induced increase in cAMP and [Ca2+]i and resulted in cell shape changes. On the other hand, 2'5'-dideoxyadenosine (DDA), an adenylate cyclase inhibitor, inhibited stretch-induced increases in cAMP and [Ca2+]i resulting in no cell shape changes. In summary, our data showed that cell shape changes were consistently dependent on [Ca2+]i rather than cAMP levels. We conclude that the primary second messenger in the stretch-induced shape changes in HUVECs is intracellular Ca2+ rather than cAMP. PMID- 11104363 TI - The reduction of myocardial damage and leukocyte polymorphonuclear accumulation following coronary artery occlusion by the tyrosine kinase inhibitor tyrphostin AG 556. AB - We investigated the effects of tyrophostin AG 556, a tyrosine kinase inhibitor, on the phenomenon of leukocyte accumulation during ischaemia and reperfusion of the myocardium. Male anaesthetized rats were subjected to total occlusion (45 min) of the left main coronary artery followed by 5 h reperfusion (MI/R). Sham myocardial ischaemia-reperfusion rats (Sham MI/R) were used as controls. Myocardial necrosis, myocardial myeloperoxidase activity (MPO), serum creatinine phosphokinase activity (CPK) serum Tumor Necrosis Factor (TNF-alpha) and Interleukin 6 (IL-6), cardiac intercellular adhesion molecule-1 (ICAM-1) and TNF alpha expression and myocardial contractility (left ventricle dP/dt(max)) were evaluated. Myocardial ischaemia plus reperfusion in untreated rats produced marked myocardial necrosis, increased serum CPK activity (196.5 +/- 19 U/100 ml, at the end of reperfusion) and myeloperoxidase activity (MPO, a marker of leukocyte accumulation) both in the area-at-risk (4.5 +/- 0.5 U/g/tissue) and in necrotic area (8.2 +/- 1.2 U/g/tissue), reduced myocardial contractility (1,706 +/- 52 mmHg/s, at the end of reperfusion) and induced a marked increase in the serum levels of TNF-alpha (1,950 +/- 97 pg/ml, at 1 h of reperfusion) and IL-6 (998 +/- 16 U/ml, at the end of reperfusion). Finally, myocardial ischaemia reperfusion injury also increased cardiac mRNA for TNF-alpha and ICAM-1 in the myocardium-at risk. Tyrphostin AG 556 (0.5, 1 and 2 mg/kg subcutaneously 5 min after the onset of reperfusion) lowered myocardial necrosis and myeloperoxidase activity in the area-at-risk (1.5 +/- 0.2 U/g/tissue, following the highest dose) and in necrotic area (2.9 +/- 0.3 U/g/tissue following the highest dose), decreased serum CPK activity (96 +/- 9 U/100 ml, at the end of reperfusion), lowered serum TNF-alpha and IL-6, increased myocardial contractility (2,096 +/- 88 mmHg s, at the end of reperfusion) and reduced cardiac mRNA levels for TNF alpha and ICAM-1. The present data suggest that tyrosine kinase inhibitors protect against myocardial ischaemia-reperfusion injury by reducing leukocyte accumulation to the ischaemic myocardium. PMID- 11104364 TI - Lagenin, a novel ribosome-inactivating protein with ribonucleolytic activity from bottle gourd (Lagenaria siceraria) seeds. AB - The seeds of Lagenaria siceraria (Family Cucurbitaceae) were extracted with water and the extract was lyophilized. The lyophilized extract was chromatographed on a DEAE-cellulose column in 10 mM Tris-HCl buffer (pH 7.2). The unadsorbed fraction was applied to an Affi-gel Blue gel column previously equilibrated with the same buffer. After removal of unadsorbed materials, the adsorbed proteins were eluted with 1.5 M NaCl in the Tris-HCl buffer. After dialysis the adsorbed fraction was loaded on a CM-Sepharose CL-6B column which had been equilibrated with and was eluted with the same buffer. After elution of unadsorbed proteins, the column was eluted with a gradient of 0-1 M NaCl in 10 mM Tris-HCl buffer (pH 7.2). The fraction eluting at about 0.55 M NaCl, which represented pure ribosome inactivating protein (RIP), inhibited cell-free translation in a rabbit reticulocyte system with an IC50 of 0.21 nM and exerted ribonuclease activity on yeast tRNA with an activity of 45 U/mg. The RIP was designated lagenin. It possessed a molecular weight of 20 kDa, smaller than the range of 26-32 kDa reported for other RIPs. The N-terminal sequence of lagenin exhibited a lesser extent of similarity to those of other Cucurbitaceae RIPs, characterized by a deletion of the first three amino acid residues and a replacement of the 4th (Phe), 17th (Phe), 18th (Ile) and 22nd (Arg) residues which are invariant in other RIPs. PMID- 11104365 TI - Effects of nitric oxide synthase inhibitors on the febrile response to lipopolysaccharide and muramyl dipeptide in guinea pigs. AB - We investigated the effect of N-nitro-L-arginine methyl ester (L-NAME), an unspecific nitric oxide synthase (NOS) inhibitor, and aminoguanidine, a relatively selective inhibitor of the inducible NOS enzyme, on both gram-negative lipopolysaccharide (LPS) and gram-positive muramyl dipeptide (MDP) fever in guinea pigs. Intraperitoneal injection of either 10 mg/kg L-NAME or 25 mg/kg aminoguanidine inhibited the febrile response to an intramuscular injection of 50 microg/kg MDP. However, LPS fever (20 microg/kg) was inhibited only by L-NAME. The development of LPS fever may therefore occur independently of the synthesis of nitric oxide by the inducible NOS enzyme, while MDP fever may involve synthesis of nitric oxide by both the inducible and the constitutively expressed NOS enzymes. PMID- 11104366 TI - Effects of Centella asiatica on ethanol induced gastric mucosal lesions in rats. AB - Centella asiatica is a herbal medicine widely used in China and India for wound healing. The aim of this study was to examine its effects on the prevention of ethanol induced gastric lesions in rats. Gastric transmucosal potential difference (PD) was reduced by the application of 50% ethanol in the gastric ex vivo chamber model and Centella extract (CE) accelerated its recovery. Oral administration of CE (0.05 g/kg, 0.25 g/kg and 0.50 g/kg) before ethanol administration significantly inhibited gastric lesions formation (58% to 82% reduction) and decreased mucosal myeloperoxidase (MPO) activity in a dose dependent manner. These results suggested that CE prevented ethanol induced gastric mucosal lesions by strengthening the mucosal barrier and reducing the damaging effects of free radicals. PMID- 11104367 TI - Nifedipine inhibits activation of transcription factor NF-kappaB. AB - This study was performed to examine the effects of the calcium channel blockers, nifedipine, amlodipine, diltiazem, and verapamil on the activation of the transcription factor NF-kappaB. A549 cells, a human epithelium-like lung carcinoma cell line, were transfected with the NF-kappaB reporter plasmid, which contains the luciferase gene driven by promoters containing a TATA element and 5 copies of the kappaB cis-acting element, and co-transfected with 0.2 microg of pSV2neo vector using LipofectAMINE. Nifedipine significantly decreased the expression of luciferase protein stimulated with IL-1beta (1 ng/mL) compared with controls: 80+/-4% at 3 micromol/L, 47+/-2% at 10 micromol/L and 30+/-2% at 30 micromol/L (each, n=3, p<0.0001). The inhibitory effect of nifedipine on promoter activity was concentration-dependent, with a maximal effect obtained at 30 micromol/L. In contrast, high concentrations (30 micromol/L) of amlodipine, diltiazem or verapamil decreased promoter activity to only 89+/-3%, 90+/-3% or 87+/-2% of control, respectively. A comparable inhibitory effect of nifedipine was observed when cells were stimulated with tumor necrosis factor (TNF)-alpha (50 ng/mL), or phorbol 12-myristate 13-acetate (PMA, 100 ng/mL). Electrophoretic mobility shift assay by lipopolysaccharide stimulation, using the RAW 264.7 macrophage cell line, also showed inhibition of NF-kappaB activation by nifedipine in concentrations of 30 and 50 micromol/L. Nifedipine possesses the unique property of inhibiting NF-kappaB, which may be independent of its calcium channel blocking activity, and may, in part, explain its immunosuppressive effect. PMID- 11104368 TI - The content and accessibility of sex education information on the Internet. AB - The objective of this study was to describe Web sites with sex education material and assess the accessibility of specific information on the Internet. First, the authors conducted a review of Web sites using specific sex education keywords. Second, 27 undergraduate students were asked to locate information on proper condom use and sexually transmitted disease (STD) symptoms. The time, number of search attempts, and number of clicks needed to identify each piece of information were recorded. The authors identified 41 sites with sex education material from almost 6 million pages yielded by the keywords. Sixty-three percent of the 1,556 most compatible pages were categorized as pornography. The students found the information on condom use and STD symptoms in an average of 4 minutes, using fewer then six clicks and two searches. The authors concluded that general information on sex education is difficult to locate on the Internet and often lacks essential elements, but accurate and useful information on specific topics can be more easily obtained. PMID- 11104369 TI - Research, the Internet, and the way things are. PMID- 11104370 TI - Understanding individual and collective capacity to enhance quality of life. PMID- 11104371 TI - Readiness to change: newspaper coverage of tobacco farming and diversification. AB - Diversification, like tobacco use prevention and cessation, is an important public health concern. The multilevel patterns of tobacco dependency suggest the need for public health approaches to the "tobacco problem." To understand how newspaper and wire service journalists cover issues involving diversification among tobacco farmers, the authors performed a content analysis of a subset of 100 articles on diversification and tobacco farming. Prochaska and DiClemente's stages of change model was applied to the "problem behavior" of tobacco farming. Among news accounts relating to tobacco farmers or tobacco farming, print media accounts gave relatively little attention to the issue of diversification. Farmers in the sample of news accounts were generally cognizant of pressures to diversify away from reliance on tobacco cultivation but were frustrated due to obstacles to diversification such as limited diversification options and relative absence of infrastructure supports. Community leaders and policy-relevant sources generally supported diversification. PMID- 11104372 TI - The social and cultural context of risk and prevention: food and physical activity in an urban Aboriginal community. AB - One of the key public health challenges facing indigenous and other minority communities is how to develop and implement effective, acceptable, and sustainable strategies for the prevention of non-insulin-dependent diabetes mellitus (NIDDM). In this article, the authors describe how an ethnographic approach was used to contextualize the behavioral risk factors for NIDDM and applied to the development of a more meaningful and appropriate epidemiological risk factor survey instrument for an urban Aboriginal population in Australia. The overall research design comprised a mixture of qualitative and quantitative methods. The ethnographic study showed that the complex web of meanings that tie people to their family and community can and should be taken into account in any social epidemiology of health and illness if the findings are to have any effective and long-term potential to contribute to successful public health interventions targeting these behavioral risk factors. PMID- 11104373 TI - A theory-based motivational approach for reducing alcohol/drug problems in college. AB - The Campuswide Alcohol and Drug Abuse Prevention Program (CADAPP) was implemented and evaluated over a 1.5-year span at the University of New Mexico (UNM). Drawing on self-regulation theory as a basis for understanding motivation for change, the program was designed to increase risk perceptions and thereby reduce the use of alcohol and other drugs among university students. The program was evaluated from 1988 to 1989 through repeated anonymous random sample surveys of all enrolled students on the UNM campus and on a similar control campus not implementing new prevention efforts during the same period. As predicted, relative to the control campus, students on the CADAPP campus after the program showed significantly higher perceived risks from substance use and significantly reduced levels of alcohol and marijuana use. These findings provide encouraging evidence for this theory-based approach to primary and secondary prevention. PMID- 11104374 TI - Assessing community change at multiple levels: the genesis of an evaluation framework for the California Healthy Cities Project. AB - More than 40 cities have participated in the California Healthy Cities Project since its inception in 1988. Because Healthy Cities efforts are community driven, these cities address diverse health and social issues using a wide variety of strategies. This complexity, in addition to the usual difficulties associated with evaluating community interventions, creates many challenges for evaluation. Given the community building and process orientation of Healthy Cities, it may be most appropriate to measure intermediate community changes that have been linked to health outcomes in previous research or, at a minimum, theoretically. The California Healthy Cities evaluation framework conceptualizes change at five levels: individual, civic participation, organizational, interorganizational, and community. The framework, developed collaboratively with Healthy Cities participants, attempts to synthesize current thinking and practice on evaluation of community projects by applying concepts from community capacity/competence, social ecology, and urban planning. PMID- 11104375 TI - Guarding against threatening HIV prevention messages: an information-processing model. AB - Previous research has suggested that fear-provoking HIV prevention messages can lead to defensive coping strategies among sexually active students who encounter such messages. An information-processing model of defensive responses is proposed that identifies and operationally defines four mediating processes--attention avoidance, blunting, suppression, and counterargumentation--that may lead to the rejection or denial of threatening health messages. Attention avoidance occurs when people indiscriminately avoid all messages; blunting is the use of distraction to avoid only threatening information in the message. Suppression occurs when people try to stop thinking about the information and avoid forming inferences about its self-relevance; counterargumentation is the biased assessment that follows comprehension and arises along with self-relevant elaboration. Situational influences on people's choice of defensive coping strategy are considered, and implications for researchers and practitioners are discussed. PMID- 11104376 TI - Failure of voluntary activation of the quadriceps femoris muscle after patellar contusion. AB - STUDY DESIGN: Descriptive study of phenomenon. OBJECTIVES: To determine the extent of failure of voluntary activation of the quadriceps femoris muscle in patients early after patellar contusion. BACKGROUND: Pain and effusion are related to the presence of quadriceps inhibition. We hypothesized that patients with patellar contusions would be unable to fully recruit their quadriceps muscles and that the activation deficit would be associated with self-report measures of function. METHODS AND MEASURES: Sixteen patients who had sustained a unilateral patellar contusion fewer than 4 months prior to testing participated in the study (7 men, 9 women; mean age = 30.0 +/- 11.6). Subjects completed a self-report questionnaire to assess knee function and performed an isometric burst superimposition test on the involved and uninvolved quadriceps at 60 degrees of knee flexion. The subjects were assigned to 2 groups according to the presence (n = 5) or absence (n = 11) of quadriceps inhibition. RESULTS: Sixty nine percent of the subjects tested were able to fully activate their quadriceps. Both groups had a decreased knee extensor force on the involved side compared to the uninvolved, but the group with inhibition had a lower side-to-side percentage of knee extensor force (mean = 65.5% +/- 18.9) than those without inhibition (mean = 85.5% +/- 16.4). CONCLUSION: Early after patellar contusion, approximately one-third of the patients demonstrated quadriceps inhibition. According to our working hypothesis, the majority of the patients tested should have demonstrated inhibition. Quadriceps inhibition was not associated with the activities of daily living, sports activity, or global rating scales in this study. Decreased volitional quadriceps force production (the hallmark of inhibition) was the only variable that discriminated patients with patellar contusion who had inhibition from those who did not. PMID- 11104377 TI - Effect of foot orthotics on calcaneal eversion during standing and treadmill walking for subjects with abnormal pronation. AB - STUDY DESIGN: Repeated measures analysis of intervention. OBJECTIVES: To determine the effects of foot orthotics and shoewear on calcaneal eversion for standing and treadmill walking. BACKGROUND: Foot orthotics are commonly used as an intervention for treating lower extremity musculoskeletal pathology. Qualitative research regarding the benefit of foot orthotics tends to be favorable, while the results of quantitative studies often conflict. METHODS AND MEASURES: Eight men (mean age = 35.8 +/- 12.7 years) and 5 women (mean age = 30.4 +/- 10.6 years), who demonstrated abnormal pronation, walked quickly (average velocity = 1.9 m/s) on a treadmill with and without foot orthotics. Subjects were filmed using a 2-dimensional video system and plastic molds designed to indicate calcaneal position inside the shoe during static standing and treadmill walking. RESULTS: Paired t tests indicated that foot orthotics significantly reduced the mean maximum calcaneal eversion angle by 2.2 degrees and the mean calcaneal eversion angle at heel rise by 2.1 degrees during fast walking. Orthotic and nonorthotic conditions did not differ significantly for the remaining kinematic variables. A one-way ANOVA indicated that calcaneal eversion in standing was significantly greater for barefoot standing compared with standing in shoes with or without orthotics. ANOVA also indicated that the plastic molds provided reliable measures of calcaneal position. CONCLUSIONS: Foot orthotics have a significant effect on calcaneal eversion and shoes also should be considered in conjunction with foot orthotic prescription. PMID- 11104378 TI - Durkan gauge and carpal compression test: accuracy and diagnostic test properties. AB - STUDY DESIGN: A prospective, criterion-based validity study. OBJECTIVES: To assess the diagnostic properties of the carpal compression test (CCT) when performed with the Durkan carpal tunnel syndrome (CTS) gauge, and to determine the measurement validity of the gauge. BACKGROUND: The CCT has been reported to be highly sensitive (.87-.89) and specific (.93-1.0) in the diagnosis of CTS when it is done with thumb pressure. The accuracy of measurements with the Durkan CTS gauge, however, has not been established and the diagnostic sensitivity and specificity of the CCT when the gauge is used has not been independently confirmed. METHODS AND MEASURES: The study sample included 33 women and 19 men, aged 18 to 85 years (45.7 +/- 13.5 years). The accuracy of the gauge was determined with a force dynamometer and holding frame. Standard nerve conduction studies (NCS) and the CCT were performed on the symptomatic extremity of all subjects. A compatible history and the NCS results were used to confirm CTS. RESULTS: The Durkan gauge registered pressures of 11.94 psi and 15.25 psi at the 12 and 15 psi gauge marks, respectively. Test sensitivity and specificity were .36 (95% CI = .17-.54) and .57 (95% CI = .39-.74), respectively. CONCLUSIONS: Pressure measurements obtained with the Durkan CTS gauge were accurate. The CCT when performed with the Durkan gauge, however, was neither sensitive or specific for the diagnosis of CTS. PMID- 11104379 TI - Self-reported oral contraceptive use and peripheral joint laxity. AB - STUDY DESIGN: A masked, single-factor, posttest-only control group design. OBJECTIVE: To explore the relationship between reported oral contraceptive use and peripheral joint laxity. BACKGROUND: Studies have found an association between increased ligamentous laxity and changes in serum levels of hormones such as estrogen, progesterone, and relaxin. Two of these hormones, estrogen and progesterone, are present in most oral contraceptives. Oral contraceptive users, therefore, provide a population for studying the effects of these hormones on the degree of ligamentous laxity. METHODS AND MEASURES: Fifty-five women between the ages of 20 and 25 years participated in this study. Thirty users of oral contraceptives were a test group and 25 nonusers of oral contraceptives were controls. The KT-1000 Arthrometer was used to measure passive anterior translation of the tibia in relation to the femur in both knees. Passive abduction and adduction of the proximal interphalangeal (PIP) joint of the second digit of the nondominant hand and distal interphalangeal (DIP) joint hyperextension of the fifth digit of the nondominant hand were measured using a goniometer. A subjective measurement of passive second PIP joint motion was also assessed and a value of minimum, moderate, or maximum laxity was assigned. Independent sample t tests were performed to compare the measurements of the oral contraceptive user and nonuser groups for each joint. A chi-square test compared the subjective PIP joint data between the 2 groups. RESULTS: No significant differences in laxity measurements at the knee or hand were found between the 2 groups. Average knee laxity varied between 5.7-7.9 mm of anterior displacement for both groups. Average PIP abduction and adduction varied between 6.5-6.7 degrees for both groups and DIP hyperextension was 28.6-29.9 degrees. CONCLUSIONS: Results of this study indicate that self-reported oral contraceptive use was not associated with peripheral joint laxity. PMID- 11104380 TI - Prediction of knee extensor and flexor isokinetic strength in young male soccer players. AB - STUDY DESIGN: Single group, cross-sectional study. OBJECTIVES: To measure various anthropometric and demographic variables in young male soccer players and to use these measurements to develop equations with which to predict the isokinetic, concentric, and eccentric moment of force. BACKGROUND: The development of equations that can predict isokinetic muscle strength from commonly measured subject characteristics can assist in the effective design of training and rehabilitation programs for athletic children. METHODS AND MEASURES: One hundred thirteen male soccer players (13.50 +/- 2.21 years of age) performed eccentric and concentric maximum efforts of the knee extensors and flexors at 60, 120, and 180 degrees/second. Stepwise regression tests were used to develop predictive equations using combinations of age, height, body mass, sexual maturation (Tanner) stage, percentage of body fat, and hours spent training per week. RESULTS: The results indicated a significant relationship of concentric and eccentric isokinetic strength for both knee extensors and flexors with 73-93% of the variance explained by using combinations of age, body mass, percentage of body fat, and hours training per week. Body mass and age were the main predictor variables under concentric conditions, whereas chronological age was not included in the models under eccentric conditions. CONCLUSIONS: The relation between isokinetic moment and multiple anthropometric and demographic variables depends on the type of muscle action examined. The results suggest that the equations we developed can be used to predict the isokinetic moment in trained, young soccer players. A cross-validation analysis is required to confirm the accuracy and the suitability of the equations developed in our study. PMID- 11104381 TI - Mobilization of surgically stabilized pelvis dangerous. PMID- 11104382 TI - Was the patient informed of risk? PMID- 11104383 TI - More expertise on trial. PMID- 11104384 TI - Pathoanatomical model and diagnosis. PMID- 11104385 TI - Trigger points and limited motion. PMID- 11104386 TI - Stratified tests, stratified slopes, and random effects models for clinical trials with missing data. AB - Because missing observations may affect the size and power of statistical tests of equality, various analytical techniques explicitly or implicitly condition the analysis on the amount of information available per person. We illustrate the difference between stratifying a slope estimate and stratifying a test statistic based on slopes. We compare a nonparametric version of the latter approach with the parametric tests available from SAS Proc Mixed. Power and size of these two approaches are considered under different parametric settings, distributions, and missing data mechanisms. PMID- 11104387 TI - On testing for drug/chemical interactions: definitions and inference. AB - The notion of zero interaction in the statistical literature is not always equivalent to what is found in the toxicology literature. A discussion about when they are the same is provided here. Design issues are of paramount importance in the analysis of drug combinations (mixtures of chemicals) when the number of constituents in the combination is larger than, say, three as the usual factorial designs are not feasible. An economical design necessary and sufficient to support the estimation of an additivity model is single drug (chemical) dose response data. Once estimated, the additivity surface can be used to make comparisons to the observed data at combination points of interest. Examples are provided to demonstrate the methods. PMID- 11104388 TI - Foundation for nonlinear models with thresholds for longitudinal data. AB - Threshold models first appeared in the literature nearly half a century ago. Threshold segments have been added to many commonly used forms of models from linear models and generalized linear models through mixed models for the analysis of cross-sectional data. Nonlinear models with thresholds for cross-sectional data are less prevalent in the literature. Nonlinear models with thresholds for longitudinal data are new. The historical developments leading to this point are reviewed as a means of introducing terms necessary for discussing features of these newer models. Nonlinear models for longitudinal data with thresholds are presented and discussed. PMID- 11104389 TI - Progress report on the guidance for industry for statistical aspects of the design, analysis, and interpretation of chronic rodent carcinogenicity studies of pharmaceuticals. AB - The U.S. Food and Drug Administration (FDA) is in the process of preparing a draft Guidance for Industry document on the statistical aspects of carcinogenicity studies of pharmaceuticals for public comment. The purpose of the document is to provide statistical guidance for the design of carcinogenicity experiments, methods of statistical analysis of study data, interpretation of study results, presentation of data and results in reports, and submission of electronic study data. This article covers the genesis of the guidance document and some statistical methods in study design, data analysis, and interpretation of results included in the draft FDA guidance document. PMID- 11104390 TI - Dropouts in longitudinal studies: definitions and models. AB - The widely used distinction of Little and Rubin (1) about types of randomness for missing data presents difficulties in its application to dropouts in longitudinal repeated measurement studies. In its place, a new typology of randomness for dropouts is proposed that relies on using a survival model for the dropout process. In terms of a stochastic process, dropping out is a change of state. Then, the longitudinal measures and dropout processes can be modeled simultaneously, each conditional on the complete previous history of both repeated measures and states. In this context, Poisson regression is used to fit various proportional hazards models, some of which are new, to the dropout process using the longitudinal measurements responses as time-varying covariates. As examples of longitudinal measurement studies displaying nonrandom dropout processes, a dental study of testosterone production in rats and clinical trials for treatment of gallstones and of depression are analyzed. PMID- 11104391 TI - A modified large sample approach in the assessment of population bioequivalence. AB - The U.S. Food and Drug Administration (FDA) requires pharmaceutical companies to show bioequivalence between different formulations or generic companies to show bioequivalence between generic drugs and brand drugs before approval. In a recent FDA guidance on bioequivalence, new criteria were proposed for assessment of population and individual bioequivalence. In this article, computer simulation is used to compare a modified large sample (MLS) upper bound for the population bioequivalence ratio with the bootstrap upper bound recommended by the FDA. The comparison criteria are the ability to maintain the stated confidence level and the estimated power of tests based on these bounds. PMID- 11104392 TI - Qualifying ELISA data: combining information. AB - With immunoassay or bioassay data, the assay standards often exhibit considerable inter-assay variability. However, the assay controls, which are used to monitor the assay performance and set acceptance criteria, should have no or less interassay variability. In this paper, we develop a mixed-effect calibration model for the assay controls to set new acceptance criteria and qualify the enzyme-linked immunosorbent assay (ELISA) data, which incorporates the interassay variation of assay standards and the nature of the assay controls, and overcomes the problems caused by traditional fixed-effect calibration model. PMID- 11104393 TI - Statistical perspective on adjusting for center effect in a 2 x 2 crossover design with application to clinical trials. AB - Patients for large-scale clinical trials are typically recruited from multiple centers to ensure adequate patient population and timely enrollment. In this note, carryover, treatment, and period effects for continuous data in multicenter studies with a 2 x 2 crossover design are examined using two types of covariate analysis adjusted for center effect proposed by Castellana and Patel (1). The correspondence among F-tests for various sources of variation in the two approaches is derived in detail. An example from a clinical study with complete and incomplete data is given to illustrate the use of the two types of covariate analysis. Also, the impact of complete and incomplete data on the covariate analyses is discussed along with the weighted squares of means analysis used in SAS procedure PROC GLM and residual maximum likelihood (REML) estimation employed in SAS procedure PROC MIXED. PMID- 11104394 TI - A remark on small-sample properties of logistic regression in three-point designs. AB - Nottingham and Birch (1) recently alleged that the maximum likelihood (ML) estimator beta of the steepness parameter in a logistic regression model could be seriously underestimated. They based their conclusion on a simulation study, investigating in particular a small-sample three-point design with a relatively large spacing between the doses. In the present work we study such situations in more detail and use complete enumeration to find the exact properties of the ML estimators. The result presented here show that the allegation by Nottingham and Birch was misleading. There is a substantial probability for an infinite outcome of beta, which appears to have been neglected by Nottingham and Birch. In fact, it will be demonstrated that the asymptotic normal approximation for beta fits quite well even with small samples, except in the upper tail where outcomes are infinite instead of large finite. The consequences for coverage probabilities of confidence intervals for both of the regression parameters are elucidated. PMID- 11104395 TI - Gender differences in risk factors for suicide in Denmark. AB - BACKGROUND: Gender is one of the most frequently replicated predictors for suicide. AIMS: To identify risk factors for suicide among males and females and to investigate whether risk factors for suicide differ by gender. METHOD: A time matched nested case-control design was performed using Danish longitudinal register databases to obtain 811 suicide cases and 79 871 controls. Data were analysed using conditional logistic regression. RESULTS: A history of hospitalised mental illness was the most marked risk factor for suicide for both genders. Unemployment, retirement, being single and sickness absence were significant risk factors for men, whereas having a child <2 years old was significantly protective for women. The relative risks for suicide differed significantly between genders according to psychiatric admission status and being the parent of a child <2 years. However, adjustment for these factors did not eliminate the gender difference in suicide risk. CONCLUSIONS: Risk factors for suicide differed by gender and gender differences could not be explained by differential exposure to known risk factors. PMID- 11104397 TI - Perspectives on modern orthopaedics: use of Adcon-L for epidural scar prevention. AB - Adcon-L is a biodegradable gel matrix that was recently approved by the Food and Drug Administration for use during single-level posterior lumbar laminectomy or laminotomy procedures. Experimental and clinical studies have suggested that the use of this product will decrease postoperative epidural scarring. However, the relationship between epidural fibrosis and patient outcome remains unclear. If the treating surgeon is of the opinion that there is a need to reduce epidural scar, the use of this product appears worthwhile. However, further studies are needed to evaluate clinical outcome and justify the use of this product on a routine basis. PMID- 11104398 TI - High-risk stress fractures: evaluation and treatment. AB - Stress fractures are common overuse injuries seen in athletes and military recruits. The pathogenesis is multifactorial and usually involves repetitive submaximal stresses. Intrinsic factors, such as hormonal imbalances, may also contribute to the onset of stress fractures, especially in women. The classic presentation is a patient who experiences the insidious onset of pain after an abrupt increase in the duration or intensity of exercise. The diagnosis is primarily clinical, but imaging modalities such as plain radiography, scintigraphy, computed tomography, and magnetic resonance imaging may provide confirmation. Most stress fractures are uncomplicated and can be managed by rest and restriction from the precipitating activity. A subset of stress fractures can present a high risk for progression to complete fracture, delayed union, or nonunion. Specific sites for this type of stress fracture are the femoral neck (tension side), the patella, the anterior cortex of the tibia, the medial malleolus, the talus, the tarsal navicular, the fifth metatarsal, and the great toe sesamoids. Tensile forces and the relative avascularity at the site of a stress-induced fracture often lead to poor healing. Therefore, high-risk stress fractures require aggressive treatment. PMID- 11104399 TI - Reconstruction of the failed femoral component and proximal femoral bone loss in revision hip surgery. AB - Advances in implant technology and surgical techniques have greatly improved the results of femoral stem revision in total hip arthroplasty. The 10-year results obtained with extensively coated noncemented revision stems parallel those obtained with cemented stems revised by using contemporary techniques. Proximal femoral bone loss is an important consideration when planning and performing revision arthroplasty. Proximal femoral bone defects can be managed with either metal or bone. Insignificant defects can be reconstructed by using primary hip arthroplasty techniques. Proximal femoral replacement prostheses are best restricted to sedentary elderly patients. Cortical strut grafts can be used reliably to reconstruct noncircumferential segmental defects. Calcar allografts are associated with unacceptably high rates of resorption. Proximal femoral allografts with either noncemented or cemented long-stem prostheses have the potential advantage of biologic soft-tissue attachment and restoration of bone stock. Impaction allografting with cement is indicated for cavitary defects and may also restore bone stock. PMID- 11104400 TI - Healing and repair of ligament injuries in the knee. AB - Although methods of treating ligamentous injuries have continually improved, many questions remain about enhancing the rate, quality, and completeness of ligament healing. It is known that the ability of a torn ligament to heal depends on a variety of factors, including anatomic location, presence of associated injuries, and selected treatment modality. A grade III injury of the medial collateral ligament (MCL) of the knee usually heals spontaneously. Surgical repair followed by immobilization of an isolated MCL tear does not enhance the healing process. In contrast, tears of the anterior cruciate ligament (ACL) and the posterior cruciate ligament often require surgical reconstruction. The MCL component of a combined ACL-MCL injury has a worse prognosis than an isolated MCL injury. The results of animal studies suggest that nonoperative treatment of an MCL injury is effective if combined with operative reconstruction of the ACL. Experimentation using animal models has helped to define the effects of ligament location, associated injuries, intrinsic factors, surgical repair, reconstruction, and exercise on ligament healing. New techniques utilizing growth factors and cell and gene therapies may offer the potential to enhance the rate and quality of healing of ligaments of the knee, as well as other ligaments in the body. PMID- 11104401 TI - Displaced three- and four-part proximal humerus fractures: evaluation and management. AB - Three- and four-part fractures are the most severe injuries in the spectrum of fractures of the proximal humerus. Despite the shortcomings of the currently available imaging techniques, fracture displacement remains an important principle in guiding management. As a result, increasing emphasis has been placed on the use of Neer's criteria in intraoperative decision making. Patients with four-part fractures with valgus impaction of the head fragment should be treated with limited open reduction and minimal internal fixation, as the blood supply to the humeral head is better preserved than with other fracture patterns and the potential for osteonecrosis is less. In the case of displaced three- and four part fractures, the physiologic age and bone quality also help guide treatment selection. In young patients with good bone quality, attempts to preserve the humeral head by meticulous handling of soft tissues and the use of low-profile implants to secure fracture fragments is recommended. Vertical fixation alone with Rush rods in patients with poor bone quality and in those with four-part fractures is no longer considered adequate and should not be used. For selected patients with three-part fractures and satisfactory bone quality, fixation with Ender rods and tension-band wiring may be appropriate. Elderly patients and those with poor bone quality have a greater risk of loss of reduction after open reduction and internal fixation, and the current consensus is that early hemiarthroplasty is the appropriate treatment. Late reconstruction necessitated by malunion and soft-tissue contracture is technically difficult, and the outcome is less favorable. The outcome of treatment of three- and four-part fractures is dependent on the surgeon's ability to analyze the fracture pattern and execute appropriate techniques to restore anatomy and function. The use of cement for prosthetic fixation and rigorous attention to tuberosity stabilization and anatomic reduction are two factors that will optimize outcome. Adequate pain relief after hemiarthroplasty has been consistently demonstrated, but return of motion and function is less predictable. PMID- 11104402 TI - Fractures of the proximal interphalangeal joint. AB - Fractures of the proximal interphalangeal joint constitute a broad spectrum of injuries. An understanding of the anatomy, the potential for joint instability, and the treatment options is essential to management of these fractures. Commonly observed fracture patterns involve one or both condyles of the proximal phalanx or the base of the middle phalanx. Fractures of the middle phalanx may involve the palmar lip or the dorsal lip or may be a "pilon" type of injury involving both the palmar and the dorsal lip with extensive intra-articular comminution. Intra-articular injuries may lead to joint subluxation or dislocation and must be identified in a timely manner to limit loss of motion, degenerative changes, and impaired function. These injuries range from those requiring minimal intervention to obtain an excellent outcome to those that are challenging to the most experienced surgeon. The treatment options include extension-block splinting, percutaneous pinning, traction, external fixation, open reduction and internal fixation, and volar-plate arthroplasty. Prompt recognition of the complexity of the injury and appropriate management are essential for an optimal functional outcome. PMID- 11104403 TI - Mis-medication and Under-utilization of Aspirin in the Prevention and Treatment of Cardiovascular Disease. AB - CONTEXT: Aspirin has been clearly established as an important therapy in the secondary prevention of cardiovascular disease (CVD), and also plays a role in primary prevention. OBJECTIVE: To determine the prevalence of use of aspirin or other over-the-counter analgesics to prevent or treat CVD. DESIGN: Representative national survey conducted in 1996. SETTING: National Family Opinion (NFO) Mail Panel. PARTICIPANTS: 23,158 persons aged 40 or older with no prior CVD, and 3818 who reported prior CVD. MAIN OUTCOME MEASURES: Use of aspirin and other over-the counter (OTC) analgesics to prevent or treat cardiovascular problems. RESULTS: Overall, 10% of survey respondents reported regular use of any analgesic for primary prevention, and 8% specifically reported using aspirin. The prevalence of aspirin use was associated with age and cardiovascular risk factors, including overweight, family history of heart problems, high cholesterol, and high stress. Of those reporting analgesic use, 11% used only non-aspirin analgesics to prevent CVD, and an additional 10% used these with aspirin. Such mis-medication was greater in women than in men, but was not related to age. Among those reporting prior CVD, only 43% used aspirin for secondary prevention. Of those using analgesics, 11% used only non-aspirin products and an additional 14% used these with aspirin. We project that 1.3 million people nationally may be erroneously taking non-aspirin products for CVD prevention, with another 1.4 million taking these along with aspirin. CONCLUSION: Efforts to increase public awareness and advocate the appropriate use of aspirin are needed, and could have an important impact on public health. PMID- 11104404 TI - A Double-Blind, Placebo-Controlled Trial of Secretin for the Treatment of Autistic Disorder. AB - OBJECTIVE: This study examines the efficacy of intravenous porcine secretin for the treatment of autism. METHODS: Using a randomized, double-blind, placebo controlled crossover design, 20 subjects with autistic disorder received either a secretin or placebo infusion at baseline and the other substance at week 4. Subjects were given the Autism Diagnostic Interview-Revised, the Autism Diagnostic Observation Schedule-Generic (ADOS-G), and other pertinent developmental measures at baseline and at weeks 4 and 8 to assess drug effects. RESULTS: For the primary efficacy analysis, change of ADOS-G social-communication total score from week 0 to week 4, no statistically significant difference was obtained between placebo (-1.0 +/- 2.4) and secretin groups (-0.7 +/- 1.4; t 0.34, df 18, P less than.74). No significant differences were obtained for the other measures, including when all 20 subjects were compared by paired t-test from baseline to 4 weeks after secretin infusion. CONCLUSION: There was no evidence for efficacy of secretin in this preliminary randomized controlled trial. These data were collected as part of a multicenter study with the University of California-Irvine and the University of Utah. PMID- 11104405 TI - Neurologic Manifestations of AIDS in Children and Adolescents: A Review of Cases in Santos, Brazil. AB - OBJECTIVE: To review the neurologic manifestations of AIDS in all children and adolescents who were seen by specialist doctors at 2 centers in Santos, Brazil, over the past 7 years. MATERIALS AND METHODS: Files of all patients aged 17 and under who were infected by HIV and admitted to 2 specialized AIDS centers between 1990 and 1997 were reviewed. RESULTS: Of the 239 children and adolescents admitted to AIDS centers, 20 presented with a variety of neurologic complications, including focal motor signs, altered tonus, retarded neurodevelopment, cognitive disturbances, intractable headache, seizures, and coma. Opportunistic infections were the exception, an important difference from the adult population of the same area. CONCLUSION: Neurologic complications of AIDS in children and adolescents in the city of Santos, Brazil, were relatively unusual, found in less than 10% of this population. The neurologic involvement did not increase the mortality of these children and adolescents. These finding may be attributable to the quality of diagnoses, treatment, and follow-up of children infected with HIV by specialized professionals in adequate institutions. PMID- 11104406 TI - How Fast Can the Racial Gap in Life Expectancy Between Whites and Blacks Be Eliminated? AB - BACKGROUND: The racial gap in life expectancy between whites and blacks fluctuated from 7.6 to 5.7 years from 1970-1996, but the causes of this gap and the years required to eliminate it remain unclear. This paper analyzes the leading causes of death and how they contribute to the racial gap in life expectancy, and estimates the number of years required to eliminate this gap. METHODS: Standard abridged life table methods and life table partitioning techniques were used to estimate the total and the cause-specific racial gap in life expectancy. Cause-specific mortality rates by age, sex, and race in the United States from 1970-1996 were obtained from the Centers for Disease Control and Prevention. The predictions of years needed to eliminate the racial gap in life expectancy are based on international and domestic trends in life expectancy improvement. RESULTS: The racial gap in life expectancy declined before 1982, increased from 1982 to 1989, and slowly declined after 1989. In 1996, about 54% and 62% of the racial gap was attributable to cancer, heart disease, homicide, and HIV for females and males, respectively. If blacks could experience substantial improvement in life expectancy, the current racial gap in life expectancy could be eliminated in about 40 years. CONCLUSIONS: The goal of eliminating the racial gap in life expectancy is a critical national priority. Differences in life expectancy are the result of multiple health and socioeconomic determinants, which will require multiple intervention strategies. The time it will take to reduce the overall gap will depend on the speed of reduction of the leading cause-specific mortality differences, which will require intensified efforts in both prevention and treatment. PMID- 11104407 TI - Indications for Cardiac Pacemaker Implantation in Myotonic Dystrophy. AB - OBJECTIVES: To determine risk factors for cardiac complications in patients with myotonic dystrophy, and to determine whether permanent cardiac pacemakers may be beneficial in the treatment of myotonic dystrophy heart disease. BACKGROUND: Myotonic dystrophy affects the cardiac conduction system. Cardiac pacemakers are easily implanted and can be life-saving in patients with severe or complete heart block. METHODS: A total of 94 patients with myotonic dystrophy were examined; 46 were followed for a mean of 6.4 (+/- 3.5 SD) years to determine predictors of the end point events of sudden death, Stokes-Adams attacks, or onset of atrial fibrillation. The end points were chosen to determine when cardiac pacemakers should be implanted (Fig. 1). RESULTS: Four out of 5 patients (and all 4 patients >60 years of age) with PR intervals longer than 275 msec had sudden death (n=1), Stokes-Adams attacks (n=1), or onset of atrial fibrillation (n=2). Conversely, only 1 of 89 patients with a PR interval shorter than 275 msec had end point events; 1 patient, aged 63 years, developed left bundle branch block and palpitations accompanied by dyspnea, which responded to a pacemaker. CONCLUSIONS: We recommend that patients with myotonic dystrophy and any indication or a family history of myotonic heart disease have at least a yearly electrocardiogram, particularly if they have a prolonged baseline or progressively increasing PR interval, or symptoms suggestive of heart block. PMID- 11104408 TI - Delay in Treatment of Pulmonary Tuberculosis: An Analysis of Symptom Duration Among Ethiopian Patients. AB - Despite the heavy burden of tuberculosis in Ethiopia, little is known about the length of time taken by the patient to seek medical care. We therefore assessed the duration of symptoms before treatment starts in patients with pulmonary tuberculosis. We studied 198 patients (134 men and 66 women) from Yirga Alem, Ethiopia, who were consecutively treated for newly diagnosed pulmonary tuberculosis. Tuberculosis was considered proven when a Ziehl-Neelsen stain of sputum showed acid-fast bacilli. The mean duration was 5.9 months, with a median (range) duration of illness for all patients of 4 months (0.5-36 months). Seventy five percent of the patients had a duration of illness of more than 2 months, and in 25% of the patients, the illness lasted more than 8 months. Patients with severe disease had a longer duration. Patients with a long duration of symptoms had a greater number of bacilli on direct microscopy of their sputum, suggesting a higher degree of infectivity. Married patients, persons with no formal education, and people living in rural areas had long illness duration. Also, patients with occupations such as farmers, housewives, soldiers, and houseworkers had increased risk compared with students. In south Ethiopia, patients with pulmonary tuberculosis present late to treatment. For some patients, the long pretreatment duration may have had consequences for the severity of the disease and for poor treatment results. Interventions that aim at earlier case detection may therefore be appropriate. PMID- 11104410 TI - The House of Managed Care. PMID- 11104409 TI - A Guide to the Assessment and Care of the Patient Whose Medical Decision-Making Capacity is in Question. AB - In general terms, medical-decision-making "capacity" is the ability to give informed consent to accept a medical intervention or to make an informed refusal of the intervention. This article presents a method to guide physicians who assess and care for patients whose medical decision-making capacities are in question. The guide relies on legal and ethical principles pertaining to informed consent. The guide recommends a specific definition of capacity. Relying on that definition, the guide employs the best empirically validated bedside tests of the neuropsychiatric elements of capacity: attention, language, memory, and frontal lobe functions (awareness and judgment). This article accomplishes 5 things that previously published capacity-assessment instruments do not do: (1) guides the physician in assessing attention, language, memory and general frontal-lobe functions; (2) guides the physician in assessing certain elements of capacity that are legally required in certain cases in some jurisdictions; (3) defines and recommends a specific standard of judgment; (4) guides the physician in caring for the patient beyond the point of simply rendering a decision about capacity; and (5) guides the physician in dealing with the complex legal issues surrounding the assessment of capacity. PMID- 11104411 TI - Internet Health Monitors for Outcomes of Chronic Illness. AB - Chronic illness is a dominant feature of healthcare. Its dominance will increase as the historically large baby boomer population reaches retirement age in 10 years. There is no current infrastructure to provide care at this scale. We propose a model for a scalable infrastructure that will continuously monitor health status. The model is based upon outcomes measures, which are dynamic versions of standardized questionnaires. To handle chronic conditions, customized interactions are administered via computer directly in patients' homes. The standard protocols of the Internet make it possible, for the first time, to reach large populations with daily interactions that record health status. Future technology will enable databases to be effectively searched across interactions from many different patients. The offloading of routine patient interactions to Internet health monitors has significant potential to increase quality and decrease costs. PMID- 11104412 TI - Aflatoxin, Tobacco, Ammonia and the p53 Tumor-Suppressor Gene: Cancer's Missing Link? AB - Aflatoxin, the fungal carcinogen first identified in 1960, is now recognized as the prototypical laboratory carcinogen. It causes mutations in the p53 tumor suppressor gene as well as ras mutations, which are involved in the majority of human cancers. Aflatoxin has been shown to contaminate tobacco products. Tobacco related cancers, including those associated with ETS, often show the same p53 mutations associated with aflatoxin exposure. The role of ammonia in neutralizing aflatoxin contamination is examined, as well as the potential role of the FDA in regulating aflatoxin contamination of tobacco products. PMID- 11104413 TI - Fractures of the Os Calcis. AB - Calcaneal fractures remain among the most challenging fractures encountered by orthopaedic traumatologists, but the technology exists today to treat any isolated calcaneal fracture. Critical to patient outcome is the restoration of the morphologic structure of the entire calcaneus, including the relationship between the articular surfaces and the overall length, width, and height. Open reduction and internal fixation via the extensile lateral approach is the preferred treatment for displaced calcaneal injuries, because it allows superior access to the anterior process, posterior facet, and tuberosity. Despite recent claims to the contrary, excellent results can be obtained with minimal risk to the patient. PMID- 11104414 TI - Academic Health Centers for the Millennium: Threats, Problems, and Proposed Strategies and Solutions. PMID- 11104415 TI - The Unbearable Confusion of Being a Consumer of Managed Care. PMID- 11104416 TI - A Scientific Evaluation of Health Effects of Two Plasticizers Used in Medical Devices and Toys: A Report from the American Council on Science and Health. PMID- 11104417 TI - Lessons Learned From the Terminally, Critically Ill Patient Who Demands to Live as Long as Possible. AB - A 67-year-old man with metastatic pancreatic cancer was admitted to the hospital for terminal care. The patient requested intensive medical support in order "to live as long as possible." Management goals included preserving end-of-life autonomy; therefore, life-extending treatments were delivered as he had adamantly requested. Simultaneously, there was agreement among physicians that futile treatments were unwarranted. Discussions with the power of attorney did not alter medical management. Elements of a "Fair Process Approach to Futility," published in March 1999 by the American Medical Association (AMA) Council on Ethical and Judicial Affairs, were utilized in an attempt to achieve conflict resolution. Medical subspecialist and ethics committee consultations had contributory roles in resolving conflict between the patient and patient surrogate and the patient's physicians. This case offers the following lessons: 1.Potential conflicts in end of-life care management philosophy should be addressed early in the patient physician relationship. 2.Patients and surrogates should be made aware that physicians are under no obligation to provide futile care. 3.Futile or medically inappropriate care should not be offered "theoretically" with the expectation that it will be refused. 4.The Advance Directive can guide end-of-life care but does not substitute for physician judgements. 5.A "fair process" end-of-life care management algorithm can provide limited structure to the process of patient physician deliberation and conflict resolution. PMID- 11104419 TI - Profiles in Primary Care: Introduction. PMID- 11104418 TI - Primary Psoas Abscess Due to Salmonella typhi. AB - Typhoid fever is a febrile illness caused by Salmonella typhi. The usual symptoms include fever, malaise, and compromise of the gut such as diarrhea and abdominal pain. Occasionally, an extraintestinal involvement can be seen and it may be accompanied by severe complications like bowel perforation or massive hemorrhage. The general manifestations are always present in the extraintestinal compromise, and can be preponderant. However, nonintestinal involvement without the general symptoms that this case reports is so rare that no related records can be found in the MEDLINE database. PMID- 11104420 TI - Profiles in Primary Care: His Father's Son. PMID- 11104421 TI - The Empire Strikes Back: Medical Unions -- Have No Fear! PMID- 11104422 TI - Genes, Neurochemicals, and Persons. PMID- 11104423 TI - Brainology. PMID- 11104424 TI - The Effect of the Information Revolution on American Medical Schools. PMID- 11104425 TI - Peer Review vs Timeliness: A Delicate Balancing Act. PMID- 11104426 TI - Critical Choices Face Healthcare in How to Use Information Technology. PMID- 11104427 TI - The Ethics of the Medical Internet: Medscape's Advertising Policy. PMID- 11104428 TI - Who Shall Lead Us? PMID- 11104429 TI - Editorial Board Announcement II. PMID- 11104430 TI - Introducing the Editorial Board of Medscape. PMID- 11104431 TI - Launch of a New Journal and Invitation to Authors and Readers. PMID- 11104432 TI - Prophylactic Antidepressant Treatment in Hospitalized Patients. PMID- 11104433 TI - From Matthew Cushing, MD. PMID- 11104434 TI - From Ware G. Kuschner, MD. PMID- 11104435 TI - Dysgeusia by Losartan in a Patient Intolerant of Captopril. PMID- 11104436 TI - From H. Rex Greene, MD. PMID- 11104438 TI - From Joseph Goldman, MPhil, MA. PMID- 11104437 TI - From Ware G. Kuschner, MD. PMID- 11104439 TI - From Kent Bottles, MD, FACPE, FCAP. PMID- 11104440 TI - From Michael Arnold Glueck, MD. PMID- 11104441 TI - Internet Empowers Patients: Response to Comments. PMID- 11104442 TI - From Kent Bottles, MD, FACPE, FCAP. PMID- 11104443 TI - From Sherwood Vine, MD. PMID- 11104444 TI - Internet Empowers Patients. PMID- 11104445 TI - From Alexander B. Niculescu III. PMID- 11104446 TI - The Good Medical Teacher. PMID- 11104447 TI - When criminals are shot: A survey of Washington, DC, jail detainees. AB - INTRODUCTION: Criminals are at high risk of being victims of violence, but little is known about their victimization. METHODS: A screen of Washington, DC, detainees found that 1 in 4 had been wounded in events that appear unrelated to their incarceration. Extensive interviews were conducted with 79 men entering the city jail from March through June 1997; the men reported 93 prior events in which they had been shot within the past 5 years. RESULTS: Eighty-three percent had personally witnessed someone being shot, and 46% had a family member killed with a gun. In the incidents in which they were shot, most were victims of robberies, assaults, and crossfires. The shootings were serious -- 35% were hit by more than 1 bullet, more than 90% went to the hospital, and 40% still had some disability from the wounds. These detainees report being shot by other criminals rather than by law-abiding citizens. Ninety percent would prefer to live in a world without easy access to firearms. CONCLUSION: These young men live in a violent world of gunplay. The overwhelming majority would prefer that firearms were not so readily available. PMID- 11104448 TI - Case-control study on radiology work, medical x-ray investigations, and use of cellular telephones as risk factors for brain tumors. AB - CONTEXT: Ionizing radiation is a well-established risk factor for brain tumors. During recent years, microwave exposure from the use of cellular telephones has been discussed as a potential risk factor. OBJECTIVE: To determine risk factors for brain tumors. DESIGN: A case-control study, with exposure assessed by questionnaires. PARTICIPANTS: A total of 233 currently living men and women, aged 20 to 80 years, were included. The case patients had histopathologically verified brain tumors and lived in the Uppsala-Orebro region (1994-1996) or the Stockholm region (1995-1996). Two matched controls to each case were selected from the Swedish Population Register. MAIN OUTCOME MEASURES: Ionizing radiation and use of cellular telephones as risk factors for brain tumors. RESULTS: A total of 209 cases (90%) and 425 controls (91%) answered the questionnaire. Work as a physician yielded an odds ratio (OR) of 6.00, with a 95% confidence interval (CI) of 0.62 to 57.7. All three case patients had worked with fluoroscopy. Radiotherapy of the head and neck region yielded an OR of 3.61 (95% CI, 0.65 19.9). Medical diagnostic x-ray examination of the same area yielded an OR of 2.10 (95% CI, 1.25-3.53), with a tumor induction period of 5 years or more. Chemical industry work yielded an OR of 4.10 (95% CI, 1.25-13.4), and laboratory work yielded an OR of 3.21 (95% CI, 1.16-8.85). Ipsilateral use of cellular telephones increased the risk for tumors in the temporal, temporoparietal, and occipital lobes (OR, 2.42; 95% CI, 0.97-6.05), ie, the anatomic areas with highest exposure to microwaves from a mobile telephone. The result was further strengthened (OR, 2.62; 95% CI, 1.02-6.71) in a multivariate analysis that included laboratory work and medical diagnostic x-ray investigations of the head and neck. CONCLUSION: Exposure to ionizing radiation, work in laboratories, and work in the chemical industry increased the risk of brain tumors. Use of a cellular telephone was associated with an increased risk in the anatomic area with highest exposure. PMID- 11104449 TI - Adverse drug reactions in hospitalized patients: A critique of a meta-analysis. AB - OBJECTIVE: To replicate and to critique a recently published meta-analysis[1] of the incidence of nonpreventable serious and fatal adverse drug reactions (ADRs) in hospitalized patients, to better understand its results and conclusions. METHODS: The published methods described in the meta-analysis of Lazarou and colleagues were followed.[1] This meta-analysis reviewed 30 original publications describing 39 prospective studies. In each study, the numbers of patients with nonpreventable ADRs, probably or definitely related to drugs, were sought to allow calculation of the incidence of "all-severities," serious and fatal, ADRs. In the original meta-analysis, these ADR incidences were then pooled to provide estimates of the incidence in all hospitalized patients. In our analysis, the original studies were examined by 2 investigators for consistency with the study search and inclusion criteria of the meta-analysis by Lazarou and colleagues, as well as accuracy and appropriateness of data extraction, meta-analysis, and conclusions. RESULTS: Multiple sources of heterogeneity among studies and data were found and include important differences in populations and hospital wards monitored, surveillance techniques, ADR definitions, determination of preventability of ADRs, distinguishing relationship to drugs, and in formats of reporting ADRs (by numbers of events or by patients). Imputations of event numerators made by the authors of the original meta-analysis were questionable and may overestimate the results of any individual study. With regard to fatal ADRs, the problem of small numbers of events is likely to introduce large errors in incidence estimates. Simple pooling of fatal event frequencies from only those studies specifically reporting the number of fatal ADRs, as was done in the meta analysis of Lazarou and colleagues, is likely to dramatically overestimate the death rate. CONCLUSION: Meta-analysis was invalid because of heterogeneity of the studies. Most of these studies did not report the data needed for incidence calculations. The methodology used was seriously flawed, and no conclusions regarding ADR incidence rates in the hospitalized population in the United States should be made on the basis of the original meta-analysis. PMID- 11104450 TI - Postcompetition elevation of muscle enzyme levels in professional football players. AB - Strenuous exercise causes a rise in circulating levels of creatine kinase (CK), and well-trained athletes liberate smaller amounts than do untrained individuals. Plasma CK, aspartate aminotransferase, and lactate dehydrogenase were measured in a group of 23 professional football players 48 hours after competition. All players were asymptomatic for myalgias or chest discomfort despite elevations of CK levels. Physicians should be aware of these elevations in muscle enzymes postexertion and interpret each in conjunction with the athlete's symptoms. PMID- 11104451 TI - Rates of hospitalization for asthma by insurance status. AB - OBJECTIVE: To compare utilization of emergency department (ED) and inpatient resources for asthmatics across insurance types. METHODS: Retrospective cohort analysis consisting of patients over 18 years of age admitted to the ED of 27 hospitals located throughout the United States for asthma between October 1, 1996 and September 30, 1997. RESULTS: 2738 patients were identified who met all inclusion/exclusion criteria. Approximately 25% of the sample had traditional indemnity insurance, 22% were managed care enrollees, 35% were enrolled in Medicaid or Medicare, and 18% were self-pay. Cost of treatment varied by insurance type in the ED and for inpatient asthma care: Medicare patients tended to have higher ED costs than all other insurance types, while indemnity patients had higher costs than Medicaid and Medicare patients. No significant differences were observed between managed care or indemnity patients for ED or inpatient costs; however, indemnity patients were less likely to be hospitalized for asthma subsequent to visiting the ED than managed care patients (OR: 0.40, 95% CI: 0.31 0.52). CONCLUSION: Significant differences in types and costs of care were observed across differing insurance types, which may suggest an "insurance effect" on asthma-related treatment in the ED and/or hospital. PMID- 11104452 TI - Linking evidence-based medicine therapeutic summary measures to clinical decision analysis. AB - OBJECTIVE: Evidence-based medicine (EBM) seeks to improve clinical practice by evaluating the quality of clinical evidence and ensuring that only the "best" evidence from clinical research is used in the management of individual patients. EBM has contributed to our understanding of the meaning of the benefit and harm of treatment as reported in the literature, and it is often promoted as an aid to clinical decision making. However, EBM therapeutic summary measures reflect only a single dimension of clinical decision making. The purpose of this work is to show how EBM therapeutic summary measures can be effectively incorporated into medical decision making. DESIGN: The effective application of the therapeutic summary measures advocated by EBM requires their integration into the framework of clinical decision analysis. Clinical decision analysis involves not only the identification and specification of the probabilities of clinical events but also the assessment of their relative values or utilities. We present here several analytic models for the integration of EBM therapeutic summary measures within the framework of clinical decision analysis. MAIN RESULTS: As expected, our analysis demonstrated that treatment should never be administered if its harm is greater than its efficacy, which is generally expressed as relative risk reduction. Likewise, a diagnostic test should never be ordered if the therapeutic harm is greater than the therapeutic efficacy. Intervention is always favored if the number needed to treat to avoid one adverse outcome (NNT) is smaller than the number needed to treat to harm one individual (NNH). When faced with a choice between two therapeutic options, the action threshold above which an intervention is favored can be expressed in terms of the harm inflicted (H) as H x NNT or NNT/NNH. If a patient's preferences are taken into account as relative value judgments (RV) of adverse events relative to that of therapeutic events, the action threshold is defined as NNT x (RV/NNH). CONCLUSIONS: In the setting of clinical decision making, EBM summary measures derived from population studies can be effectively used to define diagnostic and therapeutic action thresholds that may help in the management of individual patients. PMID- 11104453 TI - Physicians' Internet activities and their perceived coping with the medical information. AB - OBJECTIVE: To describe and analyze physicians' Internet activities and how this relates to their coping with medical information. METHODS: Postal survey among 1276 Norwegian physicians (response rate 78%). RESULTS: Seventy-two percent of all physicians had access to the Internet in 1999, up from 38% in 1997. One out of two physicians use the Internet for professional purposes. Web-based search is the dominant activity and Internet use is closely related to other ways of information-seeking (reading and attending professional meetings). A total of 70% of the respondents reported ability to obtain sufficient information for keeping updated in their daily work. "Internet-active"-physicians reported a higher rate of such ability than physicians without Internet access (74% vs 65%). CONCLUSION: The Internet plays an increasingly important role in physicians' professional updating. The impact of new information technology on the medical community should be carefully monitored in the future. PMID- 11104454 TI - Medical error and outcomes measures: where have all the autopsies gone? PMID- 11104455 TI - Is mammography indicated for women with defective BRCA genes? Implications of recent scientific advances for the diagnosis, treatment, and prevention of hereditary breast cancer. AB - About 5% of breast cancer patients have inherited their disease because of a mutation in genes encoding either the BRCA-1 or BRCA-2 proteins. Inheriting one of these mutations confers a 50% to 87% risk of breast cancer. Many physicians faced with such a patient would, at a minimum, suggest increased and earlier screening for breast cancer by routine mammography.[1] Normally, regular mammographic screening combined with appropriate and prompt treatment can reduce mortality from breast cancer by 30% in women aged 50-59 years and by about 14% 18% in women aged 40-49. There are no controlled clinical trials for screening young women who have multiple first-degree relatives developing breast cancer before age 45, or those known to carry BRCA-1 or BRCA-2 mutations. In fact, recent advances point out that BRCA-1 and BRCA-2 gene products are needed to repair radiation damage to DNA.[4,5] Based on this finding, I propose that women with defective BRCA genes are likely to have an inordinate sensitivity to radiation, and this raises a question about the advisability of routinely screening these women by frequent mammography. PMID- 11104456 TI - The healthcare CEO as leader: practical advice for improving patient care, clinical performance, and employee productivity. PMID- 11104457 TI - Stachybotrys chartarum: current knowledge of its role in disease. AB - Stachybotrys chartarum is one of several species of filamentous fungi capable of producing mycotoxins under certain environmental conditions. In some observational studies, the growth of this toxigenic mold in the indoor environment has been implicated as a cause of building-related illness. Following reports of a cluster of cases of pulmonary hemosiderosis and hemorrhage associated with exposure to Stachybotrys, public health measures have been recommended which have far-reaching implications. Although the hazards associated with exposure to some mycotoxins have been well studied, the health risks from environmental exposure to Stachybotrys remain poorly defined. The purpose of this review is to critically evaluate the current body of epidemiologic knowledge regarding Stachybotrys and to increase physician awareness regarding this emerging environmental health issue. PMID- 11104458 TI - The use of information technology in improving medical performance. Part III. Patient-support tools. AB - Despite the proliferation of computer-based resources for patients, usefulness has been limited to date. Already, 17,000 biomedical Internet sites exist, and patients are increasingly finding support and knowledge on the Internet, but the accuracy of the information found is highly variable and difficult for patients to assess. Patients have also found value in electronic communication with physicians, although relatively few physicians routinely use email to communicate with patients on a regular basis. Nonetheless, patient-focused information technologies potentially will have profound effects on medical care. With advancing sophistication of technology, patients will increasingly be able to compare and choose doctors using the Internet and to access information that allows them to monitor and regulate the quality of their own care. Further, technologies will likely be developed to allow patients to increasingly manage their own care -- whether they are patients with chronic illnesses such as diabetes or congestive heart failure who use customized software to adjust drug dosages and other treatments or patients with such common illnesses as headache or gastrointestinal infection who access self-management programs that may even write prescriptions for them. Thoughtful analysis and policy development will be critical for ensuring that the benefits are maximized and potential harm minimized. Specific areas include assessing the effects on outcomes and the characteristics of patients and technologies that succeed with self-management, and developing policies regarding liability for Web-based medical transactions and the privacy of information provided to physicians by email and via interactive Web sites. PMID- 11104459 TI - The use of information technology in improving medical performance. Part II. Physician-support tools. AB - Increasing data from a few sites demonstrate that information technologies can improve physician decision making and clinical effectiveness. For example, computer-based physician order entry systems, automated laboratory alert systems, and artificial neural networks have demonstrated significant reductions in medical errors. In addition, Internet services to disseminate new knowledge and safety alerts to physicians more rationally and effectively are rapidly developing, and telemedicine to improve rural access to specialty services is undergoing substantial growth. However, even technologies demonstrated to yield beneficial effects have not yet achieved widespread adoption, though the pace of change appears to be increasing as the Internet takes hold. Scientific evaluation of many technologies is also lacking, and the dangers of some of these technologies may be underappreciated. Research on the effects of specific technologies should be a priority. Policies should be developed to press information technology companies, such as pharmaceutical and medical device manufacturers, to recognize the importance of clinical evaluation. Research could also analyze the characteristics of effective technologies and of physicians and organizations who implement these technologies effectively. PMID- 11104460 TI - The use of information technology in improving medical performance. Part I. Information systems for medical transactions. AB - Investment in medical information technologies reached $15 billion in 1996. However, these technologies have not had the wide impact predicted in streamlining bureaucracy, improving communications, and raising the effectiveness of care. In this series, we identify how such technologies are being used to improve quality and performance, the future directions for advancement, and the policy and research developments required to maximize public benefit from these technologies. Each of these articles focuses on a different type of information technology: (1) information systems to manage medical transactions; (2) physician support technologies to improve medical practice; and (3) patient-focused technologies designed to change how people manage their own care. This first article of a 3-part series examines the successes of and opportunities for using advanced information systems that track and manage medical transactions for large populations to improve performance. Examples of such systems include: HEDIS, which gathers standardized data from health plans on quality of care; the USQA Health Services Research Program, which tracks treatment patterns and outcomes for 14 million insurance members; Ford's program to collect medical data for over 600,000 employees; and Harvard Pilgrim Health Care's system of computerized laboratory, pharmacy, ambulatory, and hospital admission records for its 1.5 million members. Data from these systems have led to modest improvements in knowledge and practice patterns for some diseases. Significant barriers are slowing efforts to add outcomes data to these databases and broaden the databases to cover larger populations. Nonetheless, existing data in currently evolving systems could be used to greater benefit in tracking public health and in identifying more effective treatments and causes of diseases. PMID- 11104461 TI - Medscape and the National Committee on Quality Assurance (NCQA). PMID- 11104462 TI - Understanding adoption of new technologies by physicians. PMID- 11104463 TI - Open invitation from the International Poverty and Health Network to all health professionals. PMID- 11104464 TI - An e-challenge for print journals. PMID- 11104465 TI - Letter in response to ACSH report on plasticizers. PMID- 11104466 TI - Response letter from ACSH phthalate panel. PMID- 11104467 TI - From Allen Markowicz, MD. PMID- 11104468 TI - The use of information technology in improving medical performance. PMID- 11104470 TI - Victor cohn, dean of science writers, dies at 80 PMID- 11104469 TI - Does effective secondary prevention of ischemic events start at hospitalization? PMID- 11104471 TI - Masking or blinding? An unscientific survey of mostly medical journal editors on the great debate. AB - I intended to end the great debate about whether "masking" is preferred to "blinding" when describing clinical trials in which group assignment is concealed from patients, data collectors, and sometimes statisticians. I conducted a highly unscientific survey, mostly of medical journal editors from the World Association of Medical Editors, but also of several strangers and distant relatives. Respondents were asked whether they preferred masking, blinding, or had no preference or considered the distinction to be unimportant. More than 80 people responded, representing 6 continents and several languages. Masking was preferred to blinding 3 to 1. However, the number of people reporting that either term is acceptable or that the issue is unimportant was about the same as the number who voted for masking. I conclude that both terms are acceptable in the context of reporting clinical research and that either may be used without confusion. I pronounce the great debate dead. PMID- 11104472 TI - The good physician PMID- 11104473 TI - Advocacy and community: the social roles of physicians in the last 1000 years. Part III. AB - The 19th and 20th centuries were to witness dramatic developments in Western medicine. The Industrial Revolution was to transform the means by which societies generated wealth. Populations grew exponentially throughout Europe and America as epidemics receded into the pages of history, and clinical medicine -- grandchild of the Enlightenment project -- was beginning to produce long-promised therapeutic benefits for individual patients. As these factors merged, healthcare would be transformed simultaneously into a commodity -- to be bought and sold on the market -- as well as a public good, and even a right, expected by citizens from their governments. Physicians would be called upon to mediate this tension, which would come to define the context of medical practice through the end of the 20th century. PMID- 11104474 TI - Advocacy and community: the social roles of physicians in the last 1000 years. Part II. AB - Medicine in the 16th through the 18th centuries underwent profound changes -- from its understanding of the human body to its growing significance in the body politic. Key to advances in medicine as well as to the growth of nations was a conceptual shift in the perception of the natural world, which would ultimately provide a methodology for therapeutic advances as well as for the building of wealthy, strong nations. PMID- 11104475 TI - Advocacy and community: the social roles of physicians in the last 1000 years. Part I. AB - Over the last 1000 years, the practice of medicine in the Western world has been shaped by extraordinary transformations -- in the organizational structures of healthcare delivery, the changing concepts of disease and illness, and the ethical and social issues posed to a growing and diversified profession. Some critical aspects that characterize contemporary Western medicine -- as professionally defined, highly organized and regulated, and scientifically and technologically based -- have emerged only within the last 200 years. For most of its history, medicine was practiced without these distinctions -- but precursors to many current tensions can be traced back to Hippocratic times. In the last millennium, medicine developed in tandem with emerging political ideologies and social structures, and the roles of physicians evolved to respond to the needs of individual patients, the profession, and society at large. As medicine became increasingly effective, it was harnessed into the political objectives of promoting social cohesion and productivity. Professional regulation and social mechanisms for the equitable distribution of healthcare became significant considerations for the profession and society. In this brief 3-part history, we will trace the major organizational, conceptual, and political changes that, together, by the year 2000, created a profession with responsibilities of advocacy for individual patients in concert with attention to the needs and demands of all the individuals in the larger community. PMID- 11104476 TI - The Millennium in infectious diseases: focus on the last Century 1900-2000. AB - Among the myriad remarkable changes in the last thousand years has been the evolution of our understanding and management of infectious diseases. To encapsulate a millennium of change is a daunting task, because the progress that has been made has been monumental in scope and breadth, with individual achievments of immense proportions. Hence, I have chosen to look at a snapshot of the last century with a focus on the years 1900 and 2000, with a brief analysis of the changes that have accounted for the obvious differences in the two eras. I have chosen this period because of the remarkable changes that have occurred and because that period provided both greater and more significant advances than in the preceding 900 years. PMID- 11104477 TI - Millennial landmarks in obstetrics and gynecology. PMID- 11104478 TI - Medscape millennium presentation. PMID- 11104479 TI - Vice president Al Gore's health care agenda and the utilization of medical services: An empirical analysis. AB - OBJECTIVE: To estimate the impact of Vice President Al Gore's healthcare agenda on the utilization of physician and hospital services among 4 uninsured target populations: parents of publicly insured children; near elderly adults, ages 55 64; employed adults with disabilities; and adults employed in small firms or self employed. METHODS: From the 1993 National Health Interview Survey, we select 4 representative samples of uninsured adults, ages 18-64, corresponding to the target groups described in Gore's healthcare agenda. For each adult in these samples, we estimate the change in medical service utilization caused by becoming insured using results from Craig and Ko.[1] The weighted average of these estimates represents the expected change in medical service utilization attributable to insurance. RESULTS: The increase in the utilization caused by insurance depends on the target group and the service in question. The increase in utilization of physician visits is 16% among parents of publicly insured children, 37% among the near elderly, 8% among the employed, disabled adults, and 21% among the self-employed. This effect is small compared with the increase in surgical procedures (31%, 110%, 316%, and 101%, respectively). However, given the size of the US healthcare system, this amounts to about a 0.5% increase in the production of medical services. Even if a universal coverage plan were instated in place of Gore's incremental coverage plan, production would increase by about 2%. CONCLUSIONS: Points 2 through 5 of Vice President Al Gore's healthcare agenda have an impact on the utilization of medical care by the 4 target populations. However, this impact varies by service and population, and its system-wide impact on the production of care is minor. PMID- 11104480 TI - Cost associated with the treatment of influenza in a managed care setting. AB - OBJECTIVE: The purpose of this study was to assess the costs and treatments associated with influenza patients with and without secondary viral or bacterial infections in a managed care setting. METHODS: Patients with influenza diagnoses (ICD-9 = 487) were identified in the PharMetrics database between January 1, 1997 and June 30, 1998. Patients were placed into 3 cohorts: influenza only (INF), influenza plus a secondary bacterial respiratory infection (BRI), and influenza plus a secondary viral respiratory infection (VRI). The index date was defined as the date of the first occurrence of an influenza diagnosis during the study period. Medical claims were assessed from the index date to the end of the influenza episode, which was defined as the date of the last claim for influenza followed by a 90-day "clean period" during which no influenza-related charges occurred. RESULTS: A total of 18,000 patients met the inclusion criteria. The mean age was 29 years, and 54% were female. Approximately 93% of patients were placed in the INF cohort, and 3% each in the BRI and VRI groups. The BRI cohort had the highest mean total cost ($5593* SD = 10,939), compared with the VRI cohort ($847 SD = 1782) and INF cohort ($602 SD = 2813) (P less than.0001 vs INF; P less than.0001 vs VRI). This total cost disparity was primarily driven by differences in inpatient costs: BRI ($3509, SD = 9474); VRI ($208, SD = 1327); INF ($138, SD = 2145). Patients in the BRI cohort averaged 0.5 hospitalizations per patient vs 0.06 in the VRI cohort and 0.03 in the INF cohort. CONCLUSIONS: Subjects in the BRI cohort were significantly more costly and had an increased risk of hospitalization as compared with subjects in the VRI or INF cohorts. Early intervention with antiviral agents and/or antibiotics, where appropriate, could result in significant cost savings for managed care organizations. PMID- 11104481 TI - Proposed frameworks to improve the quality of health web sites: review. PMID- 11104482 TI - Short-term outcomes of chronic back pain patients on an airbed vs innerspring mattresses. AB - OBJECTIVE: To compare SF-36, pain Visual Analog Scale (VAS), and sleep VAS outcomes of an adjustable airbed with innerspring mattresses in a population of chronic back pain sufferers. STUDY DESIGN: A-B-A trial, in 3 phases: the patients on their own bed for 1 night, on an adjustable airbed for 28 nights, and on their bed for 14 nights. SETTING: Outpatient pain rehabilitation, physical therapy, and alternative medicine clinics. PATIENTS: Three centers recruited 30 patients each with severe chronic back pain and without sleep apnea or other sleep disorders. MAIN OUTCOME MEASURES: SF-36 health status survey and VAS pain and sleep quality scales. RESULTS: On VAS scales, 95% showed pain improvement, and 88% reported better sleep. The average improvements were a 32% pain decrease and a 73% increase in sleep quality, significant at P less than.001 (two-tail t test). Eighty percent improved on the SF-36 physical functioning dimension and 88% improved on the bodily pain dimension. The average score on each dimension improved (P less than.001). Eighty-five percent preferred the adjustable airbed. DISCUSSION AND CONCLUSION: SF-36 and VAS outcomes measures showed a highly significant benefit for the airbed design in this short-term comparison. The airbed appears to be a useful sleep aid and an adjunct to medical and physical therapies for chronic back pain sufferers. PMID- 11104483 TI - Inadequate health insurance: costs and consequences. AB - CONTEXT: Changes in the healthcare marketplace have begun to test the nature and adequacy of health insurance. The complex nature of insurance is driving us away from the notion that there are 2 distinct groups - the insured and the uninsured toward an idea that insurance is best represented along a continuum, from the very well insured to the chronically uninsured, with a wide range of quality of coverage in between. OBJECTIVE: The objective of this study was to examine the experiences of insured adults as they try to get needed healthcare and balance the payment for these services against other basic needs. DESIGN: Using data from the Commonwealth Fund 1999 Survey of Workers' Health Insurance, the study analyzes the cost and access problems of insured adults by a number of different variables including income, plan satisfaction, health status, and insurance stability. RESULTS: Bivariate results indicate that insured adults with low incomes and those reporting fair or poor health are more likely to experience problems getting and paying for healthcare. These groups are also more likely to have problems paying for basic living expenses. CONCLUSIONS: The most essential notion of insurance is that it will provide protection against financial risk and assurance that we can get healthcare services when we are sick. Yet, we find substantial proportions of low- and modest-income, insured adults who struggle to afford insurance premiums; we also find that their insurance plans do not provide them with either access to care when needed or financial protection from the cost of that care. PMID- 11104484 TI - The National Patient Safety Foundation agenda for research and development in patient safety. PMID- 11104485 TI - How the internet can help clinicians improve their clinical skills. PMID- 11104486 TI - Scientific progress - wireless phones and brain cancer: current state of the science. AB - CONTEXT: The current science is not definitive about health risks from wireless phones; however, the legitimate questions about safety that have arisen from recent studies make claims of absolute safety no longer supportable. OBJECTIVE: The objective of this paper is to outline for primary care providers the results of the most current research on the possible impact of wireless phone use on human health. Presented are study results from Wireless Technology Research (WTR) program, the 7-year, $27 million effort funded by the wireless industry in the United States, that represents the world's most comprehensive research effort addressing this issue to date. Science-based recommendations for consumer interventions and future research are presented. DATA SOURCES: Original studies performed under the WTR program as well as other relevant research from around the world. STUDY SELECTION: This article presents a synopsis of the peer-reviewed in vitro and in vivo laboratory research, and the peer-reviewed epidemiology studies supported by the WTR, as well as a summary of other relevant work. DATA EXTRACTION: Only peer-reviewed scientific studies are presented, primarily WTR sponsored research. In addition, results of the WTR literature surveillance program, which identified other relevant toxicology and epidemiology studies on an ongoing basis, are presented. These studies are presented in the context of their usefulness in providing intervention recommendations for consumers. DATA SYNTHESIS: Following a qualitative synthesis of specific relevant non-WTR research and a critical assessment of the WTR results, the following represents the current state of scientific understanding relevant to the public health impact of wireless phones: laboratory studies appear to have confirmed that radio frequency radiation from wireless phone antennas is insufficient to cause DNA breakage; however, this same radiation appears to cause genetic damage in human blood as measured through the formation of micronuclei. An increase in the rate of brain cancer mortality among hand-held cellular phone users as compared to car phone users, though not statistically significant, was observed in the WTR cohort study. A statistically significant increase in the risk of neuro-epithelial brain tumors was observed among cellular phone users in another case-control study. CONCLUSIONS: As new data emerge, our understanding of this complex problem will improve; however, at present there is a critical need for ongoing and open evaluation of the public health impact of new science, and communication of this science and derivative intervention options to those who are potentially affected. PMID- 11104487 TI - Self-surgery: removal of ankle surgical implants--A case report. AB - Self-surgery is rare, but numerous cases of self-mutilation are reported in the literature (eg, castration, enucleation of an eye, and amputation of a limb).[1] We have found no previous reports in the literature of a patient who has performed self-surgery to remove fracture fixation implants. PMID- 11104488 TI - The Institute of Medicine Report on Medical Errors: misunderstanding can do harm. Quality of Health Care in America Committee. PMID- 11104489 TI - Religion and medicine: why faith should not Be mixed with science PMID- 11104490 TI - Religion and medicine - in reply. PMID- 11104491 TI - Religion and medicine - responses. PMID- 11104493 TI - Functional somatic syndromes in rheumatic disorders - A Patient's response PMID- 11104492 TI - Functional somatic syndromes in rheumatic disorders. PMID- 11104494 TI - Functional somatic syndromes in rheumatic disorders. PMID- 11104495 TI - Children and pain - In reply PMID- 11104496 TI - Children and pain. PMID- 11104497 TI - Institutional and economic influences on autopsy performance. PMID- 11104499 TI - In memoriam: john H. Renner, healthcare consumer advocate PMID- 11104498 TI - Low doses of gabapentin may be helpful in the management of chronic daily headache. AB - CONTEXT: The difficult management of chronic daily headache and the lack of clinical trials for this medical condition led the authors to perform this study. Gabapentin has been successfully used for a variety of chronic pain conditions and therefore may be of use in the treatment of chronic headache. OBJECTIVE: To assess the efficacy and safety of low doses of gabapentin in cases of chronic daily headache. DESIGN: Open-label study on a series of cases of chronic daily headache. PATIENTS: Twenty-one consecutive patients with primary headache lasting 4 or more hours a day, at least 15 days per month, were invited to participate in this open trial. They were treated with low doses of gabapentin for sufficient time to lead to a headache-free period of at least 7 consecutive days. A minimum of 1 follow-up visit and 1 phone call were made in the subsequent 6-9 months. MEASUREMENTS: A simple "Patient Impression of Change" was used to evaluate the results. Patients were also invited to compare this treatment to previous ones, whenever possible. RESULTS: The efficacy of the treatment with gabapentin was rated as "excellent" by 19% of the patients, "good" by 47.6%, "fair" by 19%, and "poor" by 14.4%. CONCLUSIONS: Despite the inherent limitations of such a small open trial, the authors concluded that ratings of excellent and good by two thirds of this population of patients with chronic daily headache should encourage the setup of a large double-blind, multicentric, placebo-controlled trial of low doses of gabapentin for chronic daily headache. PMID- 11104500 TI - Insight into the pathophysiology of restless legs syndrome. AB - Restless Legs Syndrome (RLS) is a disorder of sensation with a prevalence of around 2-5% of the population. Relevant to understanding the possible pathophysiological mechanism is the fact that RLS is extremely responsive to dopaminergic agents. A second issue is that iron deficiency states may precipitate RLS in as much as 25-30% of people with iron deficiency. Studies looking at basal ganglia dopaminergic function using PET and SPECT techniques have shown a decrease in binding potential for the dopamine receptor and transporter. Similar phenomena occurs in iron-deficient animals. Using MRI techniques and CSF analysis of iron-related protein, studies have suggested a reduction in brain iron concentration occurs in RLS patients. The relevance of CNS iron metabolism to the pathophysiology of RLS is discussed. PMID- 11104501 TI - Voltage-gated potassium channels in retinal ganglion cells of trout: a combined biophysical, pharmacological, and single-cell RT-PCR approach. AB - Retinal ganglion cells of young mature trout were acutely isolated by tissue printing and analyzed with the whole-cell mode of the patch-clamp technique in combination with single-cell RT-PCR. All cells either exhibited spontaneous spiking activity or could be induced to fire trains of action potentials by current injection. Depolarizing voltage steps elicited a TTX-sensitive sodium inward current and a complex outward current that could be subdivided into a calcium-dependent component that was sensitive to 100 nM iberiotoxin as well as three major types of voltage-sensitive currents: 1) a high-threshold (-20 mV) noninactivating current that was highly sensitive to submicromolar TEA and quinine, resembling recombinant mammalian Kv3.1 channels; 2) a low-threshold DTX sensitive current, matching mammalian Kv1; and 3) a fast-inactivating transient current that was highly sensitive to TEA (3 mM) but resistant to alpha-DTX (1 microM) and quinine (0.1 mM). By multiplex single-cell RT-PCR, coexpression of multiple transcripts encoding Shaker-related channel genes of trout (termed Tsha1 Tsha4) as well as two Shaw-related channels (termed Traw1 and Traw2) could be demonstrated in individual cells. PMID- 11104502 TI - Early expression of GABA(A) receptor delta subunit in the neonatal rat hippocampus. AB - The cDNA library screening strategy was used to identify the genes encoding for GABA(A) receptor subunits in the rat hippocampus during development. With this technique, genes encoding eleven GABA(A) receptor subunits were identified. The alpha5 subunit was by far the most highly expressed, followed by the gamma2, alpha2 and alpha4 subunits respectively. The expression of the beta2, alpha1, gamma1, beta1 and beta3 subunits was moderate, although that of the alpha3 and delta subunits was weak. In situ hybridization experiments, using digoxigenin labeled cRNA probes, confirmed that the delta subunit was expressed in the neonatal as well as in the adult hippocampus, and is likely to form functional receptors in association with other subunits of the GABA(A) receptor. When the more sensitive RT-PCR approach was used, the gamma3 subunit was also detected, suggesting that this subunit is present in the hippocampus during development but at low levels of expression. The insertion of the delta subunit into functional GABA(A) receptors may enhance the efficacy of GABA in the immediate postnatal period when this amino acid is still exerting a depolarizing and excitatory action. PMID- 11104503 TI - Maternal adrenalectomy at the early onset of gestation impairs the postnatal development of the rat hippocampal formation: effects on cell numbers and differentiation, connectivity and calbindin-D28k immunoreactivity. AB - The possible role of the maternal glucocorticoids on the postnatal development of the hippocampus was tested with bilateral adrenalectomy of pregnant rats. Surgery was performed 24 hr after sperm-positiveness was determined. The offspring from adrenalectomized mothers, compared with animals from control sham-operated mothers, showed decreased body weight and increased brain weight. The CA1 field of the hippocampus of these animals showed lower number of both Nissl-stained and Calbindin-immunoreactive cells, whereas the granule cell layer of the dentate gyrus showed higher number of both populations. Both types of cell numbers were statistically similar from postnatal Day 21, however, suggesting some compensatory mechanism. The neuronal populations of adrenalectomized animals appeared with a delay in the development of their dendritic trees, cytoplasmic differentiation, and synaptic connections. In the same way, both septohippocampal and hippocamposeptal projections appeared delayed in the adrenalectomized animals with respect to control ones by several days, mainly with regard to regressive events typical of the first 8 days of age. The ultrastructural study showed that every ADX postnatal group appeared more immature than the corresponding control group. These results suggest that gestational levels of maternal glucocorticoids (that were removed by adrenalectomy) influence the normal postnatal development of the hippocampus as reflected in neuron numbers and cell maturation, as well as in the developmental timing of the pattern of connectivity, and that this effect must be accomplished both in neuroepithelium and post-mitotic cells before the endogenous fetal hormones are secreted and reach concentrations capable to produce a response. PMID- 11104504 TI - Increase in neurotrophin-3 expression followed by Purkinje cell degeneration in the adult rat cerebellum after spinal cord transection. AB - Changes in brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) contents following thoracic spinal cord transection were investigated in the cerebral cortex, hippocampus, and cerebellum of rats. The NT-3 content became significantly elevated at 3 days after transection only in the cerebellum and gradually declined to the control level by 6 days after the injury, remaining unchanged in the cerebral cortex and hippocampus. No significant change in the BDNF content was observed in any of the regions tested. Immunohistochemical analysis showed that the labeling indicating NT-3-like immunoreactivity was intensified in both cerebellar granule and Purkinje cells 3 days after the injury. The number of Purkinje cells with aggregation of chromatin around the nuclear membrane and swelling of the cytoplasm and/or organelles gradually increased with time starting 4 days after the injury, demonstrating morphological changes indicative of necrosis. However, no abnormal morphology was found in cerebellar granule cells at any time examined. We suggest that it is reasonable that increased NT-3 stimulated the death of Purkinje cells, because 1) the degeneration was necrosis, which is known to be accelerated by neurotrophins under certain pathological conditions, and 2) the increase in NT-3 occurred prior to Purkinje cell degeneration. Therefore, our present results may imply that spinal cord injury-induced NT-3 accelerates injury rather than alleviates degeneration of Purkinje cells. PMID- 11104505 TI - Expression of ODC and its regulatory protein antizyme in the adult rat brain. AB - Ornithine decarboxylase and its inhibitor protein, antizyme are key regulators of polyamine biosynthesis. We examined their expression in the adult rat brain using in situ hybridization and immunocytochemistry. Both genes were widely expressed and their expression patterns were mostly overlapping and relatively similar. The levels of antizyme mRNA were always higher than those of ornithine decarboxylase mRNA. The highest expression for both genes was detected in the cerebellar cortex, hippocampus, hypothalamic paraventricular and supraoptic nuclei, locus coeruleus, olfactory bulb, piriform cortex and pontine nuclei. Ornithine decarboxylase and antizyme mRNAs appeared to be localized in the nerve cells. ODC antibody displayed mainly cytoplasmic staining in all brain areas. Antizyme antibody staining was mainly cytoplasmic in the most brain areas, although predominantly nuclear staining was detected in some areas, most notably in the cerebellar cortex, anterior olfactory nucleus and frontal cortex. Our study is the first detailed and comparative analysis of ornithine decarboxylase and antizyme expression in the adult mammalian brain. PMID- 11104506 TI - Collagen IV deposits do not prevent regrowing axons from penetrating the lesion site in spinal cord injury. AB - Scarring is suggested to impede axon regrowth across the lesion site in the injured adult mammalian central nervous system. Collagen Type IV, as a major component of the scar formed after injury, is an impediment for successful axonal regeneration and a decrease in its amount is a prerequisite for regrowing axons to cross the lesion in the postcommissural fornix in the injured adult rat (Stichel et al. [1999] Neurosci. 93:321-333). The aim of the present study was to analyze the relationship between collagen IV deposits and regrowing axons at various times after dorsal hemi-section of the adult rat spinal cord. Immunohistochemical double staining revealed that penetrating neurofilament positive axons and collagen IV deposits were co-localized in the lesion site in the initial stages of axonal sprouting (between 7 and 14 days post-operatively) and were still present 1 and 2 months post-operatively. Interestingly, collagen IV-immunoreactive areas located around cystic cavities formed at the site of injury 1 month post-operatively, were devoid of axons. In conclusion, our observations indicate that collagen IV deposits after spinal cord injury do not prevent neurofilament-positive regrowing axons from penetrating the lesion site. PMID- 11104507 TI - Site of action of suramin and reactive blue 2 in preventing neuronal death induced by dequalinium. AB - Dequalinium (DQ, an anticancer drug) is a potent neurotoxicant in the cultured developing cerebellar granule neurons (CGNs) with an IC(50) of 1.31. M after 24 hr incubation. By utilizing fluorometric technique, we found that DQ initially induced apoptosis and then necrosis associated with a marked decrease in ATP contents. The purinergic P(2) receptor antagonists (suramin, and reactive blue 2) prevented DQ-cytotoxicity, although glutamate ionotropic receptor antagonists (MK 801 and NBQX) could not. Furthermore, we quantitatively determined a reduction of mitochondrial membrane potential and an increase of free radical production induced by DQ. Suramin abolished these detrimental events of DQ. This suggests that neuronal death induced by DQ is mediated, at least in part, through a signaling pathway of free radical production-mitochondrial dysfunction. Further evidence supporting this contention is that CGN progressively became more sensitive to both DQ-induced cytotoxicity and reduced mitochondrial membrane potential. This implies that neuronal mitochondria are apparently one of the target sites for DQ and suramin and directly or indirectly induce neurotoxicity and neuroprotection respectively. The alteration in mitochondrial membrane potential during neuronal maturation may be one of the determinants accounting for the increased susceptibility to neurotoxicants such as DQ. PMID- 11104508 TI - Insulin-like growth factor-I (IGF-I) protects myelination from undernutritional insult: studies of transgenic mice overexpressing IGF-I in brain. AB - Using insulin-like growth factor-I (IGF-I)-overexpressing transgenic (Tg) mice as a model, we have shown that IGF-I promotes myelination by increasing the number of oligodendrocytes and stimulating the expression of myelin-specific protein genes. In the present study, we investigated whether IGF-I protects myelination from undernutritional insult in Tg mice. Mice were undernourished beginning from postnatal (P) day 1, a time coincident with the onset of transgene expression, and sacrificed at P20. Consistently with our previous studies, brain weights of undernourished non-Tg control mice were decreased by approximately 18%. Brain weights of undernourished IGF-I Tg mice, however, were the same as those of well fed control mice and much greater than those of undernourished control mice. The expression of two major myelin proteins [myelin basic protein (MBP) and proteolipid protein (PLP)] in cerebral cortex (CTX) and hippocampus (HIP) was decreased by 73-92% in undernourished control mice, as judged by Northern and Western blot hybridization. The abundances of MBP and PLP mRNAs and proteins, however, were decreased by only 40-70% in undernourished IGF-I Tg mice. To assess the number of oligodendrocytes and their precursors, antibodies specific for carbonic anhydrase II (CAII; an oligodendrocyte marker) and NG2 (a precursor marker) were used. Compared to their well-fed counterparts, undernourished control mice exhibited 17-30% decreases in the number of oligodendrocytes and their precursors in CTX and corpus callosum (CC), whereas well-fed IGF-I Tg mice had 21-35% increases in CTX and CC. Undernourished IGF-I Tg mice exhibited cell numbers similar to those of well-fed control mice. These data indicate that IGF-I protects myelination from undernutrition damage during development. PMID- 11104509 TI - Lipid-free versus lipid-bound P2 protein-induced experimental allergic neuritis: clinicopathological, neurophysiological, and immunological study. AB - The P2 protein of the peripheral nervous system myelin is a neuritogenic protein capable of inducing experimental allergic neuritis (EAN) in the Lewis rat. It has been suggested that the addition of some lipids to the protein isolated in the lipid-free form might enhance its immunogenicity. In this study, we compared lipid-free P2 (the EAN factor) and the corresponding lipid-bound form of the protein regarding their ability to induce EAN. Lipid-bound P2, copurified with all the myelin lipids, shows a conformation different from that of LF-P2. The timing of disease and the clinical scores of lipid-bound P2-induced EAN animals (n = 23) did not differ statistically from those injected with lipid-free P2 (n = 23), with only a tendency to higher clinical severity in the former group. Tail nerve conduction velocities did not differ in the two groups and in both were significantly lower in comparison to Freund adjuvant controls (n = 8). Inflammation and demyelination predominated in the spinal roots and were less evident in the sciatic nerve for both groups of animals. The ELISA determination of antibodies to lipid-free and lipid-bound P2 revealed the development of antibodies recognizing the lipid-free form of the protein in both groups of animals. Our results stand in contrast to results of previous studies performed after addition of exogenous lipids to the P2 purified in the lipid-free form and indicate that lipid-bound P2 is not significantly more immunogenic than lipid depleted P2. PMID- 11104510 TI - P0 protein peptide 180-199 together with pertussis toxin induces experimental autoimmune neuritis in resistant C57BL/6 mice. AB - The C57BL/6 mice strain is known to be reputedly resistant to induction of experimental autoimmune neuritis (EAN), an animal model of Guillain-Barre syndrome in human by bovine peripheral myelin (BPM), and P2 protein or the P2 protein peptide 57-81. The P0 peptide 180-199 is a stronger neuritogenic antigen than the P2 peptide 57-81. We found that this synthetic peptide induced both clinical and pathological characteristics of an acute monophasic EAN in C57BL/6 mice. Only male mice were more sensitive to EAN induction with the P0 peptide 180 199. Intravenously administrated pertussis toxin (PT) had an adjuvant effect that increased the incidence of P0 peptide 180-199-induced EAN as well as the inflammation and demyelination in the peripheral nerves. Spontaneous and P0 peptide 180-199 stimulated proliferation of peripheral T-cells were enhanced by PT-treatment as well. The enhancing effect was lower before onset of the disease (Day 6 post immunization) (p.i.) as compared to the early phase of the disease (Day 22 p.i.). Thus, P0 peptides together with PT are able to break tolerance to myelin in C57BL/6 mice. PMID- 11104511 TI - K(+)-Evoked [(3)H]D-aspartate release in rat spinal cord synaptosomes: modulation by neuropeptide Y and calcium channel antagonists. AB - This study was conducted to investigate mechanisms regulating the release of [(3)H]D-aspartate (or endogenous glutamate) in the rat spinal cord. Presynaptic modulation of glutamate release was studied in superfused synaptosomes depolarized with 20 mM KCl. Calcium-channel antagonists, omega-conotoxin GVIA (omega-CgTx GVIA; N-type), nifedipine (L-type), and omega-conotoxin MVIIC (omega CmTx MVIIC; P/Q type), were used to characterize the voltage-operated Ca(2+) channels (VOCCs) involved in this release. Nifedipine had no significant effect on the K(+)-evoked release of [(3)H]D-aspartate, but the omega-conotoxins GVIA and MVIIC produced dose-dependent inhibitory effects that were additive. The most substantial reduction (54.30% +/- 4.40%) was seen with omega-CgTx GVIA, indicating that N-type channels play a major role in the release of glutamate in this tissue. We investigated the effects of neuropeptide Y (NPY), NPY(13-36), and [Leu(31)][Pro(34)]NPY on Ca(2+)-dependent, K(+)-evoked [(3)H]D-aspartate release. NPY and NPY(13-36) equipotently inhibited the release of glutamate in a concentration-dependent manner. The half-maximal response was observed at about 12 nM; maximal inhibition of 44.22% +/- 4.60% was achieved with 0.3 microM. The selective GABA(B) agonist (-)baclofen inhibited K(+)-evoked [(3)H]D-aspartate release from superfused spinal cord synaptosomes by 50.00% +/- 4.80% at 10 microM. When NPY(13-36) and (-)baclofen were used together at maximal doses, their release-inhibiting effects were not additive. In addition, neither of the agonists was able to enhance the inhibition produced by pretreating the synaptosomes with the selective inhibitor of N-type VOCCs omega-CgTx GVIA. These results are consistent with the hypothesis that presynaptic Y(2)-like and GABA(B) receptors regulate glutamate release by blocking Ca(2+) currents through N-type VOCCs. Characterization of the receptors that can inhibit the release of glutamate may provide useful information for treatment of conditions characterized by excessive glutamatergic transmission in the spinal cord. PMID- 11104512 TI - Quercetin inhibits c-fos, heat shock protein, and glial fibrillary acidic protein expression in injured astrocytes. AB - Quercetin, a bioflavonoid, is found widely in many kinds of fruits and vegetables. It is known to engage in many bioactivities, such as interfering with of the progress of stress responses to injury. In the present study, we investigated the effects of quercetin on some injury responses in primary cultures of astrocytes. These injury responses included the elevation of c-fos protein, heat shock protein (HSP70), and glial fibrillary acidic protein (GFAP). After heat shock insult, the levels of c-fos protein and HSP70 in astrocytes increased. With quercetin treatment, these proteins were significantly reduced. The inhibition of these injury responses by quercetin in astrocytes indicated a dose dependency, with the highest effect at 100 microM. We have previously established a scratch injury model in a primary culture of astrocytes. In that model, astrocytes responded to the scratch injury by an elevation in their GFAP level and formation of hypertrophic cell processes, which extend into the scratch areas. Quercetin treatment reduced the number of hypertrophic cell processes being extended into the scratch areas. With 100 microM of quercetin, there was a complete inhibition of the formation of the hypertrophic cell process. Western blot analysis for GFAP indicated that quercetin significantly reduced the induction of GFAP in the scratch model. At 100 microM, the total GFAP content in the injured cultures was reduced to a level lower than that of the control. This implied that quercetin might possess an antigliotic property. PMID- 11104513 TI - Characteristics of odorant elicited calcium changes in cultured human olfactory neurons. AB - An important step in establishing and utilizing a cell culture system for the in vitro study of olfaction is assessing whether the cultured cells possess physiological properties similar to those of mature olfactory neurons. Various investigators have successfully established proliferating cell lines from olfactory tissue, but few have demonstrated the characteristics of odor sensitivity of these cells. We successfully established cultured cell lines from adult human olfactory tissue obtained using an olfactory biopsy procedure and measured their ability to respond to odor stimulation using calcium imaging techniques. A subset of the human olfactory cells in culture displayed a distinct morphology and specifically expressed immunocytochemical markers characteristic of mature human olfactory neurons such as OMP, G(olf), NCAM and NST. Under defined growth conditions, these cultured cells responded to odorant mixes that have been previously shown to elicit intracellular calcium changes in acutely isolated human olfactory neurons. These odorant-elicited calcium responses displayed characteristics similar to those found in mature human olfactory neurons. First, cultured cells responded with either increases or decreases in intracellular calcium. Second, increases in calcium were abolished by removal of extracellular calcium. Third, inhibitors of the olfactory signal transduction cascades reversibly blocked these odorant-elicited intracellular calcium changes. Our results demonstrate that cultures of adult human olfactory cells established from olfactory biopsies retain some of the in vivo odorant response characteristics of acutely isolated cells from the adult olfactory epithelium. This work has important ramifications for investigation of olfactory function and dysfunction using biopsy procedures and in vitro assays of odor sensitivity. PMID- 11104514 TI - Sex-dependent changes in blood-brain barrier permeability and brain NA(+),K(+) ATPase activity in rats following acute water intoxication. AB - To understand the increased susceptibility of the development of serious complications to hypoosmotic hyponatremia in young females, we examined the resistance of blood brain barrier (BBB) permeability to water along with the synaptosomal Na(+),K(+)ATPase activity in both sexes of rats during acute water intoxication. Four groups of rats were used: Group I and II were normal female and male rats injected with only Evans-blue. Group III and IV were water intoxicated female and male rats respectively. BBB permeability in female rats was found to be increased following acute water intoxication. In contrast, synaptosomal Na(+),K(+)ATPase activities in both water intoxicated male and female rats were found significantly lower than those in control rats. But inhibition in enzyme activity in synaptosomes from water intoxicated female rats was more pronounced than those of corresponding male rats. Our results concluded that female sex steroids may be responsible for the highly significant decrease in synaptosomal Na(+),K(+)ATPase activity and increased BBB permeability in female rats following water intoxication. PMID- 11104515 TI - The grail problem. PMID- 11104516 TI - Whither genomics? AB - The flood of data from genome-wide analysis is transforming biology. We need to develop new, interdisciplinary approaches to convert these data into information about the components and structures of individual biological pathways and to use the resulting information to yield knowledge about general principles that explain the functions and evolution of life. PMID- 11104517 TI - Are plant formins integral membrane proteins? AB - BACKGROUND: The formin family of proteins has been implicated in signaling pathways of cellular morphogenesis in both animals and fungi; in the latter case, at least, they participate in communication between the actin cytoskeleton and the cell surface. Nevertheless, they appear to be cytoplasmic or nuclear proteins, and it is not clear whether they communicate with the plasma membrane, and if so, how. Because nothing is known about formin function in plants, I performed a systematic search for putative Arabidopsis thaliana formin homologs. RESULTS: I found eight putative formin-coding genes in the publicly available part of the Arabidopsis genome sequence and analyzed their predicted protein sequences. Surprisingly, some of them lack parts of the conserved formin-homology 2 (FH2) domain and the majority of them seem to have signal sequences and putative transmembrane segments that are not found in yeast or animals formins. CONCLUSIONS: Plant formins define a distinct subfamily. The presence in most Arabidopsis formins of sequence motifs typical or transmembrane proteins suggests a mechanism of membrane attachment that may be specific to plant formins, and indicates an unexpected evolutionary flexibility of the conserved formin domain. PMID- 11104518 TI - Phylogenetic variation and polymorphism at the toll-like receptor 4 locus (TLR4). AB - BACKGROUND: Differences in responses to bacterial surface lipopolysaccharides (LPSs) are apparent between and within mammalian species. It has been shown in mice that resistance to LPS is caused by defects in the Toll-like receptor 4 gene (Tlr4), the product of which is thought to bind LPS and mediate LPS signal transduction in immune system cells. RESULTS: We have sequenced the Toll-like receptor 4 gene of humans (TLR4; 19.0 kilobases, kb) and mice (Tlr4; 91.7 kb), as well as the coding region and splice junctions of Tlr4 from 35 mouse (Mus musculus) strains, from the chimpanzee and from the baboon. No other discernible genes or regions of interspecies conservation lies close to Tlr4 and, in both humans and mice, flanking sequences and introns are rich in repeats of retroviral origin. Interstrain analyses reveal that Tlr4 is a polymorphic protein and that the extracellular domain is far more variable than the cytoplasmic domain, both among strains and among species. The cytoplasmic domain of the Tlr4 protein is highly variable at the carboxy-terminal end. CONCLUSIONS: We suggest that selective evolutionary pressure exerted by microbes expressing structurally distinguishable LPS molecules has produced the high level of variability in the Tlr4 extracellular domain. The highly variable carboxy-terminal region of the cytoplasmic domain is likely to determine the magnitude of the response to LPS within a species. PMID- 11104520 TI - Molecular mechanisms of spindle function. AB - The key molecules involved in regulating the assembly and function of the mitotic spindle are shared by evolutionarily divergent species. Studies in different model systems are leading to convergent conclusions about the central role of microtubule nucleation and dynamics and of kinesin-related motor proteins in spindle function. PMID- 11104519 TI - An overview of the structures of protein-DNA complexes. AB - On the basis of a structural analysis of 240 protein-DNA complexes contained in the Protein Data Bank (PDB), we have classified the DNA-binding proteins involved into eight different structural/functional groups, which are further classified into 54 structural families. Here we present this classification and review the functions, structures and binding interactions of these protein-DNA complexes. PMID- 11104521 TI - What initiates actin polymerization? AB - For those working on the actin cytoskeleton, a major theme of the 39th annual meeting of the American Society for Cell Biology [http://www.faseb.org/ascb/meetings/am99/+ ++main99mtg.htm] (Washington DC, December 11-15, 1999) was the elucidation of how actin polymerization is initiated. The emphasis was on the regulation and localization of the Arp2/3 complex, which over the last two years has been shown to stimulate actin nucleation, and on the identification of additional proteins that interact with actin and Arp2/3 in a variety of organisms. PMID- 11104522 TI - A cultivated taste for yeast. AB - The availability of complete genomic sequences of Saccharomyces cerevisiae has catalyzed a cultural change in the practice of yeast biology, providing opportunities to develop high throughput techniques to define protein function, to define drug targets, and to discover and characterize drugs. PMID- 11104523 TI - Membrane traffic between genomes. AB - Proteins of the Rab and SNARE families target vesicles to their intracellular destinations. A comparison of these families from the budding yeast, fission yeast, nematode and fruitfly genomes has implications for the organization of membrane traffic in different organisms. PMID- 11104524 TI - Measuring a cell's response to stress: the p53 pathway. AB - The characterization of complex cellular responses to diverse stimuli can be studied by the use of emerging chip-based technologies. PMID- 11104525 TI - Fly immunity: great expectations. AB - Preliminary analysis of the Drosophila genome sequence reveals important similarities and differences between the functioning of mammalian and invertebrate immune systems. PMID- 11104526 TI - The many hues of plant heterochromatin. AB - Recent sequence and cytogenetic analyses of heterochromatin in Arabidopsis, together with other results from Arabidopsis and maize, indicate that plant heterochromatin can have very different origins, compositions and dynamics. Shared features that must determine and/or be a result of its unique biological properties are also revealed. PMID- 11104527 TI - Chaotic actin. AB - Living cells organize their internal contents by coordinating the chaotic thermal motion of protein molecules. A recent study, combining experiment with stochastic simulation, shows how this might be achieved in the case of actin polymerization. PMID- 11104528 TI - Dementia with Lewy bodies: a pure case. AB - A pure case of autopsy-confirmed dementia with Lewy bodies (DLB) is described. The patient presented with distinctive verbal fluency deficits in the context of mild language impairment, intact recognition memory, and impaired paragraph recall. Neuroimaging (CT and SPECT) showed progressive medial temporal lobe atrophy. Neuropathology revealed Lewy bodies, degeneration in the substantia nigra, nucleus basalis of Meynert (Nakano & Hirano, 1984), and locus ceruleus, but no pathology characteristic of Alzheimer's disease. It is in this sense that the case is "pure" DLB. Early neuropsychological diagnosis of DLB is essential (Salmon et al., 1996) given the potentially fatal hazard of neuroleptics (McKeith et al., 1992) and the difficulties associated with clinical neurological diagnoses (Litvan et al., 1998). PMID- 11104529 TI - Extinction-like effects in normals: independence of localization and response selection. AB - An extinction-like effect in normal subjects was previously elicited when a low salience target in the left was simultaneously presented with a highly salient distractor in the right visual hemifield, but not vice versa (Pollmann, 1996). We investigated in four experiments whether this extinction-like effect depends on (a) explicit localization and (b) response competition. It was found that the extinction-like effect could be replicated in the absence of both. In contradistinction to our previous results, low-salience distractors had no effect on pop-out target search. This showed that explicit spatial localization demands lead to low-salience distractor interference on pop-out search. PMID- 11104530 TI - Neuropsychological impairments in the recognition of faces, voices, and personal names. AB - In order to determine the dissociability of face, voice, and personal name recognition, we studied the performance of 36 brain-lesioned patients and 20 control subjects. Participants performed familiarity decisions for portraits, voice samples, and written names of celebrities and unfamiliar people. In those patients who displayed significant impairments in any of these tests, the specificity of these impairments was tested using corresponding object recognition tests (with pictures of objects, environmental sounds, or written common words as stimuli). The results showed that 58% of the patients were significantly impaired in at least one test of person recognition. Moreover, 28% of the patients showed impairments that appeared to be specific for people (i.e., performance was preserved in the corresponding object recognition test). Three patients showed a deficit that appeared to be confined to the recognition of familiar voices, a pattern that was not described previously. Results were generally consistent with the assumption that impairments in face, voice, and name recognition are dissociable from one another. In contrast, there was no clear evidence for a dissociability between deficits in face and voice naming. The results further suggest that (a) impairments in person recognition after brain lesions may be more common than was thought previously and (b) the patterns of impairment that were observed can be interpreted using current cognitive models of person recognition (Bruce & Young, 1986; Burton, Bruce, & Johnston, 1990). PMID- 11104531 TI - Language and cognition-Kurt Goldstein's theory of semantics. AB - Kurt Goldstein is regarded as one of the major proponents of the holistic movement at the beginning of the 20th century. He rejected the strong localization hypothesis in the field of aphasiology and attempted to link language disturbances to an underlying general intellectual impairment. Goldstein's criticism was based on his subtle symptomatology, his organismic biology, and his philosophical reflections. In his concept of abstract attitude Goldstein searched for a general psychological function that might explain a variety of aphasic symptoms. Abstract attitude bridges the gap between cognitive and linguistic structures. According to Goldstein, it is the basis for words to have a meaning, to be employed in a categorical sense. Since amnesic aphasics are confined to a concrete attitude, their words have lost their representational function. Although Goldstein's concept of abstract attitude is no longer used in scientific discourse, it is analyzed for its heuristic value. It led Goldstein to questions about the relation between cognition and language and to fragments of a semantic theory. PMID- 11104532 TI - Nature of the working memory deficit in fragile-X syndrome. AB - Working memory performance in a group of young Fragile X males with FMR-1 full mutation was compared to a learning disabled comparison group comprising Down's syndrome males and two control groups of mainstream schoolchildren. Performance was assessed on a battery of tasks tapping the three components of working memory phonological loop, visual-spatial sketch pad, and the central executive. The results indicated that the Fragile X group displayed a general impairment on working memory tasks that cannot be attributed to a single working memory component per se. Instead, the results suggest that Fragile X males have a working memory deficit that may be attributed to how much attentional resource a specific task requires and their overall available executive capacity, irrespective of the working memory subsystem. PMID- 11104533 TI - A performance measure of the degree of hand preference. AB - The present paper describes a performance method for determining hand preference. The task requires participants to reach into different regions of hemispace to perform various actions (point, pick up, toss, sweep, and position) with a dowel located at each position. In accordance with the participants' hand preference as measured by the Waterloo Handedness Questionnaire, the preferred hand was used more frequently on the various performance tasks. The distribution of hand use in working space indicates that preferred hand use was almost exclusive for actions carried out in ipsilateral hemispace, while it is used only moderately for actions in contralateral hemispace, revealing that this hand is used throughout a wider range of extrapersonal space than the nonpreferred hand. These trends were observed across all of the performance tasks, suggesting that task complexity did not affect the frequency of preferred hand use either overall or, more specifically, in right hemispace, as was predicted. This finding is inconsistent with empirical work on questionnaires indicating that verbal reports of preferred hand use increase for more complex tasks (e.g., Steenhuis & Bryden, 1988). As well, performance on the preferential reaching task correlated significantly with hand preference as measured on the Waterloo Handedness Questionnaire (Bryden, 1977), unlike the other performance measure examined, indicating that the preferential reaching task is sensitive to differences in the degree of hand preference. PMID- 11104534 TI - Development of perceptual asymmetry for free viewing of chimeric stimuli. AB - Free-viewing chimeric stimuli tasks have been used in a number of studies to assess perceptual asymmetries and draw inferences about hemispheric lateralization in children and adults. In order to determine whether perceptual asymmetries for nonverbal information are present in children, a free-viewing chimeric stimuli task was used in 63 normally developing 6- through 16-year-old children. Stimuli included affect (happy faces), gender, quantity, and shape. An overall left hemispace (LHS) advantage was present by 6 years of age. This LHS preference was more prominent by age 10 and then plateaued. No preference for shape was detected at any of the age ranges studied. These results suggest that perceptual asymmetries for visual stimuli develop during childhood and appear to reach a plateau by age 10. The observed specificity for certain types of nonverbal stimuli should be taken into account in future studies of perceptual asymmetry in both normal and neurologically impaired children. PMID- 11104535 TI - Covert processing of faces in prosopagnosia is restricted to facial expressions: evidence from cross-modal bias. AB - We present a single case study of a brain-damaged patient, AD, suffering from visual face and object agnosia, with impaired visual perception and preserved mental imagery. She is severely impaired in all aspects of overt recognition of faces as well as in covert recognition of familiar faces. She shows a complete loss of processing facial expressions in recognition as well as in matching tasks. Nevertheless, when presented with a task where face and voice expressions were presented concurrently, there was a clear impact of face expressions on her ratings of the voice. The cross-modal paradigm used here and validated previously with normal subjects (de Gelder & Vroomen, 1995, 2000), appears as a useful tool in investigating spared covert face processing in a neuropsychological perspective, especially with prosopagnosic patients. These findings are discussed against the background of different models of the covert recognition of face expressions. PMID- 11104536 TI - Influences of handedness and gender on the grooved pegboard test. AB - We studied performance on the Grooved Pegboard Test upon repeated trials and transfer of training between the hands in the first trial. The classification of handedness was based on the writing hand. We employed three trials for each hand and two different protocols for the order in which the hands started the test. For the three trials combined, women were faster than men. From the first to the second trial, there was an improvement in performance for both sexes. Within the first trial, sex differences reached significance and the protocol interacted with handedness. In this trial, only left-handed men were found to benefit from previous opposite-hand performance. It is speculated that a larger corpus callosum in left-handed men allows for the greater transfer of training between the hands. PMID- 11104537 TI - Timing and force components in bilateral transfer of learning. AB - Bilateral transfer of perceptual and motor components in movement control was investigated through two experiments. In Experiment 1 a simple anticipatory timing task was practiced with either the preferred or the nonpreferred hand. After a short resting interval an additional set of trials was performed with the contralateral hand. In Experiment 2, the same experimental design was used to investigate bilateral transfer of fine force control in a wrist-flexion movement. Analysis of the results showed that bilateral transfer of learning took place for both anticipatory timing and force control, with more noticeable transfer of training for the former. Asymmetry in transfer was found for force control, with significant transfer only in the preferred-to-nonpreferred direction. Transfer of anticipatory timing occurred similarly in both directions. These results indicated anticipatory timing as a powerful component for bilateral transfer, while force control showed to be more dependent on practice with the specific muscular system. PMID- 11104538 TI - The variability of practice hypothesis in motor learning: does it apply to Alzheimer's disease? AB - Based on Schmidt's (1975) variability of practice hypothesis, this study examined acquisition and transfer of a gross motor skill, namely tossing, in 58 patients with Alzheimer's disease (AD) and 58 healthy older adults under constant, blocked, and random practice conditions. While healthy older adults were able to learn the tossing task equally well under the three practice conditions, only AD patients receiving constant practice showed significant improvements. Tests of intermediate transfer yielded the expected random practice advantage in healthy controls but not AD patients. None of the practice conditions facilitated intermediate transfer in AD patients; however, constant practice did benefit these impaired individuals on tests of near transfer. These results indicate that the variability of practice hypothesis does not extend to AD patients. As motor learning and transfer were clearly a function of constant practice, future attempts to retrain basic activities of daily living in AD patients should emphasize consistency in training. PMID- 11104539 TI - Defective representation of knowledge in Parkinson's disease: evidence from a script-production task. AB - The deficits seen in frontal-lobe patients and in the elderly show clearly that spontaneous script generation depends on good frontal-lobe function. Shallice, however, has proposed that one aspect of script generation (contention scheduling, CS) which is involved in the activation and maintenance of overlearned or routine scripts may depend more on the basal ganglia. Patients with Parkinson's disease would thus be expected to manifest deficits somewhat different from those observed in frontal-lobe patients when generating scripts. The performances of 16 nondemented and nondepressed patients with idiopathic Parkinson's disease were compared to those of 16 age-matched normal control subjects under two experimental conditions; routine, forward script generation and nonroutine, backward script generation. Parkinsonian patients generated scripts significantly deprived of contextual elements in the forward condition and made significantly more sequencing and perseverative errors in both forward and backward conditions than did normal subjects. They also produced a significantly higher number of irrelevant intrusions, in both conditions, than did controls. These results support, in a general sense, Shallice's notion that the basal ganglia are important in script generation; however, other specific predictions of Shallice's model were not supported by our findings. PMID- 11104540 TI - Selective attention and interhemispheric response competition in the split-brain. AB - The interfering effect of an unattended stimulus on processing of an attended item was studied in a single split-brain participant (LB) and in normal controls. Pairs of letters were presented to the left visual field (LVF), right visual field (RVF), or bilaterally. Participants attended to the rightmost letter while attempting to ignore the leftmost letter. Responses associated with the attended and to-be-ignored letters could be compatible or incompatible. Manual response latencies were generally slower on Response Incompatible compared to Response Compatible trials. Notably, LB displayed this effect on Bilateral trials, where target and distractor were presented to opposite visual fields. LB was unable to perform a same-different matching task with bilateral letter stimuli, but was able to name bilateral letters accurately. Hence, in the bilateral condition, the ability to cross-compare letters was dissociated from attentional interference and from letter naming. Implications of these findings are discussed. PMID- 11104541 TI - Dimensional complexity and power spectral measures of the EEG during functional versus predicative problem solving. AB - Electroencephalograms were recorded in 22 men while solving tasks of visual pattern completion and during mental relaxation. They were primed (by foregoing trials) to solve these tasks either in a predicative or functional mode of thinking. Predicative thinking required that in order to complete the pattern the subject had to get involved with the logic of the static structure of the pattern and therefore had to recognize the recurrence of certain features of the elements (e.g., shape, color, and size). Functional thinking required involvement in a dynamic reading of the logic of the pattern and therefore to search for operations and actions to be performed on the pattern elements (e.g., pushing, mirroring, and rotating). The EEG complexity during predicative thinking decreased in comparison to functional thinking and mental relaxation, with this reduction being most pronounced over the right parietal cortex. A reduction in dimensional complexity during functional thinking as compared to mental relaxation, which was concentrated over the left central cortex, although significant, was less clear. The reduced EEG complexity during predicative thought, dominant over the right hemisphere, could reflect increased competitive inhibition among respective cortical neuron assemblies in association with the visual analysis of static element features, converging upon those predicates relevant for the solution. PMID- 11104542 TI - The effect of familial sinistrality on the relation between schizophrenialike thinking and pseudoneglect. AB - We have recently reported a correlation between schizophrenialike symptoms and the degree of pseudoneglect in healthy right-handers. We aimed to investigate the effect of familial sinistrality (FS) on this relation. Seventy-six healthy right handers were divided into four groups on the basis of gender and FS. A computerized version of Corsi's task was used as the visuospatial task. Subjects filled in the Magical Ideation Scale (MI), which asked for delusionlike beliefs, and performed the Corsi's task using each hand. Performance of both hemispaces was separately evaluated. In all groups, performance of the left hemispace was better than that of the right hemispace and FS+ subjects performed better than FS subjects. When the right hand was used, performance was correlated to MI scores only for FS- groups. Findings suggest that the correlation between right-sided neglect and proneness to schizotypy in normal right-handers is affected by FS. PMID- 11104543 TI - Interhemispheric effects of simulated lesions in a neural model of letter identification. AB - Experimental studies have produced conflicting results about the extent to which the intact, nonlesioned cerebral hemisphere is responsible for recovery from cognitive deficits following focal brain damage such as a stroke. To obtain a better theoretical understanding of interhemispheric interactions during recovery, we examined the effects of simulated lesions to a bihemispheric neural model of letter identification under various assumptions about hemispheric asymmetries, corpus callosum influence, and lesion size. Among other results, the model demonstrates that the intact hemispheric region's participation in the recovery process is a function of preexisting lateralization and lesion size, indicating that interpretation of experimental work should take these factors into account. PMID- 11104544 TI - The Williams syndrome cognitive profile. AB - Williams syndrome is a rare neurodevelopmental disorder caused by a hemizygous deletion of approximately 1.5 megabases on chromosome 7q11.23. In this article, we outline a Williams Syndrome Cognitive Profile (WSCP) that operationalizes the cognitive characteristics of the syndrome using measures of absolute and relative performance on subtests of the Differential Abilities Scales (Elliot, 1990a). Testing confirmed excellent sensitivity and specificity scores for the WSCP. Seventy-four of 84 individuals with Williams syndrome fit the WSCP while only 4 participants in a contrast group met all of the WSCP criteria. It was also found that the WSCP does not vary greatly with chronological age or overall level of cognitive ability for individuals with Williams syndrome. Possible applications for the WSCP include psychoeducational evaluation and empirical research such as the search for genotype/phenotype relations in this genetically based syndrome. PMID- 11104545 TI - Afferent dysgraphia after right cerebral stroke: an autonomous syndrome? AB - Afferent dysgraphia is an acquired writing deficit characterized by deletions and duplications of letters and strokes. The commonly accepted interpretation states that afferent dysgraphia is associated with three main clinical features: production of spatial writing errors; the presence of left unilateral neglect; and no deterioration in performance when writing blindfolded. In order to test whether these symptoms necessarily co-occur with afferent dysgraphia, we studied the writing performances of a series of eight right brain-damaged patients. In sentence copying, spontaneous handwriting, and writing to dictation they showed afferent dysgraphia. However, signs of left neglect and spatial dysgraphia were evident only in some cases. Furthermore, the frequency of afferent errors increased when patients were required to write without vision. The present study demonstrates that afferent dysgraphia is an autonomous clinical entity and that it results from a selective impairment of a mechanism whose function is that of comparing the information about the number of letters and strokes specified at the level of letter motor programs and the actual number of movements already realized. PMID- 11104546 TI - Ratings of emotion in laterally presented faces: sex and handedness effects. AB - Sixteen right-handed participants (8 male and 8 female students) and 16 left handed participants (8 male and 8 female students) were presented with cartoon faces expressing emotions ranging from extremely positive to extremely negative. A forced-choice paradigm was used in which the participants were asked to rate the faces as either positive or negative. Compared to men, women rated faces more positively, especially in response to right visual field presentations. Women rated neutral and mildly positive faces more positively in the right than in the left visual field, whereas men rated these faces consistently across visual fields. Handedness did not affect the ratings of emotion. The data suggest a positive emotional bias of the left hemisphere in women. PMID- 11104548 TI - Theoretical experimental neuropsychology (TENNET XII) PMID- 11104547 TI - Influence of the movement parameter to be controlled on manual RT asymmetries in right-handers. AB - In this experiment we test whether the effects of manual asymmetries on movement preparation depend on the parameter (amplitude or direction) to be programmed. In two experiments, only the amplitude, or the direction, of aiming movements was constrained. Reaction and movement times were measured. Results show that RTs are always shorter for left-hand than for right-hand movements. There is an effect of target extent in the amplitude condition, but not in the direction one. RTs for ipsilateral movements are shorter than RTs for contralateral movements. These results are discussed in the light of the processes involved in setting the amplitude or direction of the movement and with regard to the competency of the two hemispheres regarding these processes. PMID- 11104550 TI - Epigenetic gene deregulation in cancer. AB - A mini-review of the literature concerning epigenetic gene regulation in cancer. PMID- 11104551 TI - AQ4N: a new approach to hypoxia-activated cancer chemotherapy. AB - Preclinical studies demonstrate that in vivo AQ4N enhances the anti-tumour effects of radiation and chemotherapeutic agents with a dose-modifying factor of approximately 2.0. With careful scheduling no, or very little, additional normal tissue toxicity should be observed. AQ4N is a bioreductive prodrug of a potent, stable, reduction product which binds non-covalently to DNA, facilitating antitumour activity in both hypoxic and proximate oxic tumour cells. AQ4N is clearly different in both its mechanism of action and potential bystander effect compared to previously identified bioreductive drugs. In particular AQ4N is the only bioreductive prodrug topoisomerase II inhibitor to enter clinical trials. Targeting this enzyme, which is crucial to cell division, may help sensitize tumours to repeated (fractionated) courses of radiotherapy. This is because in principle, the bioreduction product of AQ4N can inhibit the topoisomerase activity of hypoxic cells as they attempt to re-enter the cell cycle. PMID- 11104552 TI - Phase I and pharmacologic study of CT-2584 HMS, a modulator of phosphatidic acid, in adult patients with solid tumours. AB - CT-2584 HMS, 1-(11-dodecylamino-10-hydroxyundecyl)-3, 7-dimethylxanthine-hydrogen methanesulphonate, is a modulator of intracellular phosphatidic acid. We treated 30 patients as part of a Phase I and pharmacokinetic study to determine the maximum-tolerated dose of CT-2584 HMS, toxicity profiles, pharmacokinetic profile and antitumour effects at escalating dose levels. CT-2584 HMS was given as a continuous infusion for 6 hours for 5 consecutive days every 3 weeks. Plasma samples for pharmacokinetic studies were analysed using a validated high performance liquid chromatographic assay. Mean C(max)and AUC values for each dose group were similar on days 1 and 5 and increases in plasma concentration (C(max)and AUC) appeared proportional to the dose. CT-2584 HMS had a mean elimination half-life of 7.3 hours. Values of V(d)and clearance were independent of dose and duration of treatment. Dose escalation was halted at 585 mg/m(2)because of malaise and lethargy, which was sometimes accompanied by nausea and headache. 26 patients were evaluable for response, one patient with pleural mesothelioma achieved a partial response to treatment confirmed by CT scanning. A dose level of 520 mg/m(2)daily x 5 days would be suitable for Phase II testing. Alternative schedules of CT-2584 HMS to overcome the limiting toxicity of malaise would be worthy of examination. PMID- 11104553 TI - Application of positron emission tomography imaging to cancer screening. AB - Whole-body positron emission tomography (PET) with(18)F-fluorodeoxyglucose (FDG) is a diagnostic modality that can noninvasively survey the entire body and sensitively detect various cancers. In this study, we examined the potential application of whole-body PET for cancer screening in asymptomatic individuals. PET was performed in conjunction with conventional examinations including physical examination, laboratory study, ultrasonography and chest computed tomography. Between September 1994 and March 1999, 3165 asymptomatic individuals participated in 5575 screening sessions (2017 men and 1148 women; mean +/- SD age, 52.2+/-10.4 years). Follow-up periods were no less than 10 months. PET results were compared with the screening outcomes. Within 1 year after screening, malignant tumours were discovered in 67 of the 3165 participants (2.1%). PET findings were true-positive in 36 of the 67 cancers (54%). Most of the 36 patients underwent potentially curative surgery; thus a wide variety of cancers were detected by PET at potentially curable stages. However, PET findings were false-negative in 31 of the 67 patients (46%). 14 of these 31 (45%) were of urological origin. FDG PET imaging has the potential to detect a wide variety of cancers at potentially curable stages. However, PET imaging is not suited to screening test of general population because PET examination involves substantial cost. PMID- 11104554 TI - Evidence for a schedule-dependent deleterious interaction between paclitaxel, vinblastine and cisplatin (PVC) in the treatment of advanced transitional cell carcinoma. AB - A phase II study to evaluate the efficacy and toxicity of the combination of vinblastine, paclitaxel and cisplatin (PVC) in previously untreated patients with advanced transitional cell carcinoma. Chemotherapy naive patients with locally advanced or metastatic transitional cell carcinoma received the intravenous combination of paclitaxel 175 mg/m(2)over three hours followed by cisplatin 70 mg/m(2)over 3 hours on day 1 and vinblastine 3 mg/m(2)as a bolus on days 1 and 8 on a 21-day cycle, to a maximum of 6 cycles. The day 8 vinblastine was omitted if the total neutrophil count was <1.0. 15 patients (13 M, 2 F) of median age 66 (54 75) received a median of 5 cycles of treatment. There were two complete responses (13%; 95% CI 2-40%) and five partial responses (33%; 95% CI 12-62%), for an overall response rate of 46% (95% CI 21-73%). Responses occurred only in those with locally recurrent tumours and/or lymph nodes involved. Neutropenia at Grade 3-4 occurred in 14 of 67 cycles (21%) resulting in 7 episodes of neutropenic sepsis. Grade 3-4 thrombocytopenia was not observed. Other Grade 3 toxicity included alopecia (10 pts), diarrhoea (2 pts), constipation resulting in bowel obstruction (2 pts), nephrotoxicity (1 pt), myalgic pain (1 pt) and peripheral neuropathy (1 pt). Six patients developed Grade 2 paraesthesia. The median time to progression was 6 months and the median survival was 11 months. The regimen PVC was both less effective against transitional cell carcinoma and less toxic than expected. This may reflect an inhibitory interaction between vinblastine and paclitaxel and this schedule cannot be recommended for further investigation. PMID- 11104555 TI - Prognostic study of continuous variables (white blood cell count, peripheral blast cell count, haemoglobin level, platelet count and age) in childhood acute lymphoblastic leukaemia. Analysis Of a population of 1545 children treated by the French Acute Lymphoblastic Leukaemia Group (FRALLE). AB - Many cutpoints have been proposed to categorize continuous variables in childhood acute lymphoblastic leukaemia (white blood cell count, peripheral blast cell count, haemoglobin level, platelet count and age), and have been used to define therapeutic subgroups. This variation in the choice of cutpoints leads to a bias called the 'Will Rogers phenomenon'. The aim of this study was to analyse variations in the relative risk of relapse or death as a function of continuous prognostic variables in childhood ALL and to discuss the choice of cutpoints. We studied a population of 1545 children with ALL enrolled in three consecutive protocols named FRALLE 83, FRALLE 87 and FRALLE 89. We estimated the risk of relapse or death associated with different values of each continuous prognostic variable by dividing the sample into quintiles of the distribution of the variables. As regards age, a category of children under 1 year of age was distinguished and the rest of the population was divided into quintiles. The floated variance method was used to calculate the confidence interval of each relative risk, including the reference category. The relation between the quantitative prognostic factors and the risk was monotonic for each variable, except for age. For the white blood cell count (WBC), the relation is log linear. The risk associated with WBC values in the upper quintile was 1.9 times higher than that in the lower quintile. The peripheral blast cell count correlated strongly with WBC (correlation coefficient: 0.99). The risk increased with the haemoglobin level, and the risk in the upper quintile was 1.3 times higher than that in the lower quintile. The risk decreased as the platelet count increased: the risk in the lower quintile was 1.2 times higher than that in the upper quintile. The risk increased gradually with increasing age above one year. The small subgroup of patients (2.5% of the population) under 1 year of age at diagnosis had a risk 2.6 times higher than the reference category of patients between 3 and 4.3 years of age. When the risk associated with a quantitative prognostic factor varies monotonously, the selection of a cutpoint is arbitrary and represents a loss of information. Despite this loss of information, such arbitrary categorization may be necessary to define therapeutic stratification. In that case, consensus cutpoints must be defined if one wants to avoid the Will Rogers phenomenon. The cutpoints proposed by the Rome workshop and the NCI are arbitrary, but may represent an acceptable convention. PMID- 11104556 TI - A medical research council randomized trial of single agent carboplatin versus etoposide and cisplatin for advanced metastatic seminoma. MRC Testicular Tumour Working Party. AB - The UK Medical Research Council conducted this trial of carboplatin chemotherapy in advanced seminoma to compare single agent carboplatin with a standard combination of etoposide with cisplatin. The use of single agent carboplatin was expected to be associated with reduced toxicity. A total of 130 patients with advanced seminoma were randomly assigned to treatment with either single agent carboplatin (C) at a dose of 400 mg/m(2)to be corrected for glomerular filtration rate outside the range 81-120 ml min(-1)and to be administered on day 1 of a 21 day cycle to a total of 4 cycles or to etoposide + platinum (EP). The trial was designed as an equivalence study aiming to exclude a reduction in the 3-year progression-free survival in patients allocated to carboplatin of between 10 and 15%, requiring initially a target accrual of 250 patients (90% power significance level 5% (one-sided)). The trial closed after 130 patients had been randomized following recommendation by an independent data monitoring committee. At a median follow-up time of 4.5 years, 81% of patients had been followed up for at least 3 years and 19 patients have died. The estimated PFS rate (95% Confidence Intervals (CI)) at 3 years was 71% (60-82%) in patients allocated C and 81% (71-90%) in those allocated EP; the 95% CI for the difference in 3 year PFS was - 6% to +19%. The hazard ratio of 0.64 (95% CI 0.32-1.28) favoured EP but the difference was not statistically significant (log rank chi-squared = 1.59 P = 0.21). The 3-year survival rate was 84% (75-92%) in those allocated C, and 89% (81-96%) in those allocated EP. The hazard ratio for survival was 0.85 with 95% CI, 0.35-2.10, log rank chi-squared = 0.12, P = 0.73. The trial has not demonstrated statistically significant differences in the major survival endpoints comparing single agent carboplatin with a combination of etoposide + cisplatin. This cannot be taken as an indication of equivalence since the limited size of this trial rendered it unable to exclude a 19% lower progression-free survival and survival in those treated with single agent carboplatin which would be important clinically. Standard initial chemotherapy for advanced seminoma should be based on cisplatin combinations and the role of carboplatin awaits the outcome of further studies. PMID- 11104557 TI - Timing within the menstrual cycle, sex, and the use of oral contraceptives determine adrenergic suppression of NK cell activity. AB - Physiological responses that involve adrenergic mechanisms, such as stress induced changes in cardiovascular indices, were reported to fluctuate along the menstrual cycle. Metastatic development following surgery was also reported to vary according to the menstrual phase during which a primary breast tumour was removed. Natural killer (NK) cells are believed to play an important role in controlling metastases. Our recent studies in rats demonstrated that adrenergic suppression of NK activity and of resistance to metastasis is more profound during oestrous phases characterized by high levels of oestradiol. In the current study in humans, we examined the in vitro impact of a beta-adrenergic agonist, metaproterenol (MP), on NK activity, comparing blood drawn from (a) women tested at 3-4 different phases of their menstrual cycle (n = 10), (b) women using oral contraceptives (OC) (n = 10), and (c) men (n = 7). NK activity in each blood sample was assessed in the presence of 5 different concentrations of MP (10(-8)M to 10(-6)M), and in its absence (baseline). The results indicated marked group differences in the magnitude of NK suppression by MP: EC(50)was 2. 6-fold lower in the luteal phase compared to the follicular phase, and 1.8-fold lower in OC users compared to men, who were least susceptible to the effects of MP. No significant group differences or menstrual effects in baseline levels of NK activity were evident. These findings provide the first empirical evidence for menstrual regulation of adrenergic impact on cellular immune competence. Relevance of these findings to the relation between the timing of breast cancer excision within the menstrual cycle and survival rates is discussed. PMID- 11104558 TI - Intra-arterial chemotherapy in locally advanced or recurrent carcinomas of the penis and anal canal: an active treatment modality with curative potential. AB - The prognosis of locally advanced or recurrent carcinomas of the penis (PE) and of the anal canal (AC) after conventional treatment is dismal. We report 16 patients (eight with AC carcinomas and eight with PE cancers) treated by intra arterial (IA) chemotherapy. Fifteen of them were treated for locally advanced or recurrent disease and one in an adjuvant setting. The chemotherapy was administered via a femoral IA catheter with its tip located above the aortic bifurcation, under the inferior mesenteric artery. It consisted of eight push injections, given over a 48-h period, of the following drug combination: cisplatin 8.5 mg m(-2), 5-FU 275 mg m(-2), methotrexate 27.5 mg m(-2), mitomycin C 1.2 mg m(-2), and bleomycin 4 mg m(-2). Leucovorin was given po, 4 x 15 mg day( 1), during the chemotherapy and for 3 days thereafter. A total of 52 cycles of treatment were administered. Of the 15 patients evaluable for response, six obtained a CR (three PE, three AC) and eight a PR. Among the complete responders, four are alive and disease-free 2-15 years after treatment. The other patients enjoyed an objective response lasting 3-25 months (median 7 months). Four patients developed grade III/IV haematological toxicity with three episodes of febrile neutropenia, one of them with a fatal outcome due to patient's failure to obtain medical attention at the onset of his fever, one a grade III mucositis of the glans, and four a grade III/IV cutaneous toxicity, the latter caused by the IA administration of bleomycin. In conclusion, IA chemotherapy is effective and potentially curative in locoregionally advanced or recurrent carcinomas of the penis and of the anus. Its contribution in the primary management of advanced penile or anal carcinoma should be prospectively investigated. PMID- 11104559 TI - Population carrier frequency of hMSH2 and hMLH1 mutations. AB - Knowledge of population carrier frequency for DNA mismatch repair (MMR) gene mutations would contribute to understanding the burden of cancer due to genetic susceptibility, but robust prevalence estimates are lacking. To estimate carrier frequency, we genotyped a cohort of relatives of mutation carriers and determined their colorectal cancer prevalence. Systematic Finnish and US data were combined with Scottish genotype and cancer prevalence data in a Bayesian calculation. The estimated carrier prevalence in the population aged 15-74 years is 1:3139 (95% Cl = 1:1247-1:7626) and these carriers are at high risk of colorectal and other cancers. PMID- 11104560 TI - Most microsatellite unstable sporadic colorectal carcinomas carry MBD4 mutations. AB - The MBD4 gene is involved in the repair of mutation at methyl-CpG dinucleotides. In microsatellite unstable tumours MBD4 can itself be mutated at an exonic polynucleotide tract. By analysing DNA from microdissected tumour samples we have found that both frequency and pattern of mutation are more significant than originally reported. PMID- 11104561 TI - Screening breast cancer patients for Norwegian ATM mutations. AB - 483 Norwegian breast cancer patients were screened for six different ataxia telangiectasia mutated (ATM) mutations previously found to account for 83% of the disease alleles in Norwegian ataxia telangiectasia (AT) patients. Only one carrier was found. These results provide no evidence in favour of an excess risk of breast cancer associated with heterozygosity for classical AT mutations, but remain consistent with a maximum 2.4-fold increased risk. PMID- 11104562 TI - No evidence of linkage to chromosome 1q42.2-43 in 131 prostate cancer families from the ACTANE consortium. Anglo, Canada, Texas, Australia, Norway, EU Biomed. AB - Genetic linkage studies worldwide have proposed various chromosomal localizations for prostate cancer susceptibility genes. A recent study found evidence for linkage to chromosome 1q42.2-43. The aim of our study was to attempt to confirm these findings by performing linkage analysis in 131 families with multiple prostate cancer cases selected from the ACTANE (Anglo, Canada, Texas, Australia, Norway, EU Biomed) Consortium. Parametric and non-parametric linkage (NPL) analyses were performed. Two-point LOD scores failed to show evidence of linkage at any marker (maximum two-point LOD score = 0. 40 at recombination fraction theta = 0.2 with marker D1S2850). Using a multipoint heterogeneity analysis, the estimated proportion of families linked to this putative locus (alpha) was 0% (95% CI = 0. 00-0.33). Non-parametric linkage analysis also found no evidence of linkage (maximum NPL score = -0.12, P = 0.55). This analysis of 131 ACTANE families does not support the presence of a locus for a prostate cancer susceptibility gene at 1q42.2-43. Although we cannot rule out the existence of such a locus, analysis indicates that less than 16% of families could be linked to this region. These findings may be a reflection of the locus heterogeneity involved in this disease indicating that there are still other major susceptibility loci to be identified. PMID- 11104563 TI - Progressive genetic aberrations detected by comparative genomic hybridization in squamous cell cervical cancer. AB - Genetic changes orchestrated by human papillomaviruses are the most important known factors in carcinogenesis of the uterine cervix. However, it is clear that additional genetic events are necessary for tumour progression. We have used comparative genomic hybridization to document non-random chromosomal gains and losses within a subset of 37 cervical carcinomas matched for clinical stage Ib, but with different lymph node status. There were significantly more chromosomal changes in the primary tumours when the lymph nodes were positive for metastases. The most frequent copy number alterations were loss of 3p, 11q, 6q and 10q and gain of 3q. The smallest areas of loss and gain on chromosome 3 were 3p14-22 and 3q24-26. The study identifies progressive DNA copy number changes associated with early-stage invasive cervical cancers with and without lymph node metastases, a factor of potential prognostic and therapeutic value. PMID- 11104564 TI - Prognostic value of genomic alterations in minimal residual cancer cells purified from the blood of breast cancer patients. AB - The prognostic value of disseminated tumour cells derived from 353 breast cancer patients was evaluated. Disseminated tumour cells were purified from blood using a newly established method and nucleic acids were subsequently isolated. We investigated genomic imbalances (GI) such as mutation, amplification and loss of heterozygosity of 13 tumour suppressor genes and 2 proto-oncogenes using DNA from isolated minimal residual cancer cells. Significant correlations were found between genomic alterations of the DCC - and c-erbB-2 genes in disseminated breast cancer cells and actuarial relapse-free survival. Furthermore, increasing numbers of genomic imbalances measured in disseminated tumour cells were significantly associated with worse prognosis of recurrent disease. Logistic regression and Cox multivariate analysis led to the identification of genomic imbalances as an independent prognostic factor. Determination of disseminated tumour cells by genotyping of oncogenes and tumour suppressor genes seems not only to be a useful adjunct in follow up of carcinoma patients but provides also valuable additional individualized prognostic and predictive information in breast cancer patients beyond the TNM system. PMID- 11104565 TI - Elevated expression of thyroid hormone receptor alpha 2 (c-erb A- alpha 2) in nasopharyngeal carcinoma. AB - Differential display was used to identify genes differentially expressed between cultured normal nasal epithelial cells and nasopharyngeal carcinoma (NPC) cell lines. A 130 bp cDNA fragment showing homology with thyroid hormone receptor alpha2 (TR-alpha2 or c-erb A-alpha2) was identified in NPC cell lines. Northern blot analysis using the 130 bp cDNA fragment and a TR-alpha2 specific cDNA containing part of the coding region as probes, we were able to detect a 2.7 kb transcript corresponding to that of TR-alpha2 in NPC cell lines but not in normal nasal epithelial cells. RNA in situ hybridization was used to detect TR-alpha2 expression in clinical biopsies obtained from NPC patients and non-tumour controls. TR-alpha2 mRNA was detected in 1 out of 24 (4.2%) normal nasopharynx epithelium biopsies, in 5 out of 27 (18.5%) primary and 15 out of 24 (62.5%) recurrent tumours. The positive rate of TR-alpha2 expression in recurrent NPC biopsies was significantly higher than that in normal nasopharynx epithelium (P<0.00001). The relevance of the elevated expression of TR-alpha2 in the pathogenesis process of NPC was discussed. PMID- 11104566 TI - Clinicopathologic significance of sialyl Le(x) expression in advanced gastric carcinoma. AB - Sialyl Lewis(x)antigen (SLX) is a carbohydrate antigen that serves as a ligand for selectin, an adhesion molecule expressed on vascular endothelial cells. The expression of SLX in 245 patients with advanced gastric carcinoma was examined immunohistochemically, and its clinicopathologic significance was analysed. We classified the patients with advanced gastric carcinoma into 91 with differentiated type and 154 with undifferentiated type. SLX expressed in 135 of 245 patients (55%), comprising 68 (75%) patients with differentiated carcinoma and 67 (44%) with undifferentiated carcinoma. The positive rate for SLX expression was significantly higher among patients with differentiated carcinoma than among those in undifferentiated carcinoma (P<0.0001). With differentiated carcinoma, the incidence of lymph node metastasis, advanced tumour stage (stage III and IV) and liver recurrence was significantly higher in SLX-positive patients than in SLX-negative ones (P<0.0001, P = 0.0065 and P = 0. 028, respectively). Moreover, the prognoses were better in patients with SLX-negative tumours than in those with SLX-positive tumours (P = 0.019). With undifferentiated carcinoma, there were no significant correlations between SLX expression and any clinicopathological features or prognoses. The clinicopathologic significance of SLX expression in gastric carcinoma patients depends on histologic type. SLX expression may be of great relevance in predicting liver metastases in patients with differentiated carcinoma. PMID- 11104567 TI - C-myc amplification in breast cancer: a meta-analysis of its occurrence and prognostic relevance. AB - Data from basic research suggests that amplification of the proto-oncogene c-myc is important in breast cancer pathogenesis, but its frequency of amplification and prognostic relevance in human studies have been inconsistent. In an effort to clarify the clinical significance of c-myc amplification in breast cancer, we conducted a comprehensive literature search and a meta-analysis in which 29 studies were evaluated. The weighted average frequency of c-myc amplification in breast tumours was 15.7% (95% CI = 12.5-18.8%), although estimates in individual studies exhibited significant heterogeneity, P<0.0001. C-myc amplification exhibited significant but weak associations with tumour grade (RR = 1.61), lymph node metastasis (RR = 1.24), negative progesterone receptor status (RR = 1.27), and postmenopausal status (RR = 0.82). Amplification was significantly associated with risk of relapse and death, with pooled estimates RR = 2.05 (95% CI = 1.51 2.78) and RR = 1.74 (95% CI = 1.27-2.39), respectively. This effect did not appear to be merely a surrogate for other prognostic factors. These results suggest that c-myc amplification is relatively common in breast cancer and may provide independent prognostic information. More rigorous studies with consistent methodology are required to validate this association, and to investigate its potential as a molecular predictor of specific therapy response. PMID- 11104568 TI - The prognostic value of the tumour marker Cyfra 21-1 in carcinoma of head and neck and its role in early detection of recurrent disease. AB - This study examines a new tumour marker, Cyfra 21-1, as a prognostic marker in predicting the survival of H&N cancer patients, and its correlation with clinical outcome during prolonged follow up of these patients. The study included 67 patients with primary detection of carcinoma of H&N. The survival of these patients was evaluated in correlation with the disease stage and Cyfra 21-1 levels at initial diagnosis. 38 patients were followed clinically and with serial assays for at least 12 months, or until recurrence was diagnosed. Cyfra 21-1 levels were determined periodically, using an Elisa kit. Patients with Cyfra 21-1 < 1.5 ng ml(-1)had a higher survival rate compared to patients with Cyfra 21-1 > or = 1.5 ng ml(-1)(63% vs. 20%, respectively). The risk ratio of Ln(Cyfra 21-1) is 1.62 (P = 0.028). In a Cox regression model that included the disease stage and Ln(Cyfra 21-1), Ln(Cyfra 21-1) was preferred as the main parameter for predicting patients survival. In 83% of the 12 patients with recurrent or residual disease, Cyfra 21-1 was elevated before or during clinical detection of the recurrence. Cyfra 21-1 was found to be a prognostic marker for carcinoma of H&N, unrelated to the stage of the disease. Elevated levels of Cyfra 21-1 without clinical evidence of disease can be attributed to the marker's mean lead-time as compared to the clinical appearance of the disease. PMID- 11104569 TI - Expression of Ku70 correlates with survival in carcinoma of the cervix. AB - Cervical carcinoma affects around 3400 women in the UK each year and advanced disease is routinely treated with radiation. As part of a programme to establish rapid and convenient methods of predicting tumour and patient responses to radiotherapy, we have examined the relationship between the pre-treatment expression of the Ku components of the DNA damage recognition complex DNA-PK and patient survival in cervical carcinoma. Using immunohistochemistry of formalin fixed sections of tumour biopsies, antibodies to Ku70 and Ku80 stained identical regions of tumour and there was a high degree of correlation between the mean number of cells stained positive for the two components in 77 tumours (r = 0.82, P<0.001). In 53 tumours there was a borderline significant correlation between measurements of tumour radiosensitivity (surviving fraction at 2 gray: SF2) and Ku70 expression (r = 0.26, P = 0.057) and no correlation for Ku80 (r = 0.18, P = 0.19). However, all tumours with a low number of Ku70 or Ku80 positive cells were radiosensitive. Furthermore, using log-rank analysis there was significantly higher survival in the patients whose tumours had a low Ku70 expression (P = 0.046). This difference was also reflected with Ku80, but did not reach statistical significance (P = 0.087). The study suggests that lack of Ku protein leads to radiosensitivity in some tumours and that other factors are responsible for radiosensitive tumours with high Ku expression. It is likely that the most accurate prediction of treatment outcome will lie in assessing the expression of several proteins involved in the recognition and repair of DNA damage, one of which will be Ku. PMID- 11104570 TI - Definition of the role of chromosome 9p21 in sporadic melanoma through genetic analysis of primary tumours and their metastases. The Melanoma Cooperative Group. AB - Malignant melanoma (MM) is thought to arise by sequential accumulation of genetic alterations in normal melanocytes. Previous cytogenetic and molecular studies indicated the 9p21 as the chromosomal region involved in MM pathogenesis. In addition to the CDKN genes (p16/CDKN2A, p15/CDKN2B and p19(ARF), frequently inactivated in familial MM), widely reported data suggested the presence within this region of other melanoma susceptibility gene(s). To clearly assess the role of the 9p21 region in sporadic melanoma, we evaluated the presence of microsatellite instability (MSI) and loss of heterozygosity (LOH) in primary tumours as well as in synchronous or asynchronous metastases obtained from the same MM patients, using 9 polymorphic markers from a 17-cM region at 9p21. LOH and MSI were found in 27 (41%) and 11 (17%), respectively, out of 66 primary tumours analysed. In corresponding 58 metastases, MSI was found at higher rate (22; 38%), whereas a quite identical pattern of allelic deletions with 27 (47%) LOH+ cases were observed. Although the CDKN locus was mostly affected by LOH, an additional region of common allelic deletion corresponding to marker D9S171 was also identified. No significant statistical correlation between any 9p21 genetic alteration (LOH, MSI or both) and clinicopathological parameters was observed. PMID- 11104571 TI - Intracellular Fas ligand in normal and malignant breast epithelium does not induce apoptosis in Fas-sensitive cells. AB - Fas ligand (FasL) is expressed on some cancers and may play a role in the immune evasion of the tumour. We used immuno-histochemistry to study the expression of Fas and FasL in tissue samples from breast cancer patients, as well as normal breast tissue. Our results show that Fas and FasL are co-expressed both in normal tissue and in breast tumours. Fas and FasL mRNA were expressed in fresh normal and malignant breast tissue, as well as cultured breast epithelium and breast cancer cell lines. Flow cytometry analysis of live cells failed to detect FasL on the surface of normal or malignant breast cells; however, both stained positive for FasL after permeabilization. Fas was detected on the surface of normal breast cells and T47D and MCF-10A cell lines but only intracellularly in other breast cell lines tested. Neither normal breast epithelium nor breast cell lines induced Fas-dependent apoptosis in Jurkat cells. Finally, 20 tumour samples were stained for apoptosis. Few apoptotic cells were detected and there was no increase in apoptotic cells on the borders between tumour cells and lymphocytes. We conclude that FasL is expressed intracellularly in both normal and malignant breast epithelium and unlikely to be important for the immune evasion of breast tumours. PMID- 11104572 TI - HERG potassium channels are more frequently expressed in human endometrial cancer as compared to non-cancerous endometrium. AB - HERG K(+)channels, besides contributing to regulate cardiac and neuronal excitability, are preferentially expressed in tumour cell lines of different histogenesis, where their role in the development and maintenance of the neoplastic phenotype is under study. We show here that both herg gene and HERG protein are expressed with high frequency in primary human endometrial cancers, as compared to normal and hyperplastic endometrium. RT-PCR and immunohistochemistry, using specific anti-HERG antibodies developed in our laboratory, were applied to tissue specimens obtained from 18 endometrial cancers and 11 non-cancerous endometrial tissues. herg RNA and HERG protein are expressed in 67% and 82%, respectively, of cancerous, while in only 18% of non-cancerous tissues. In particular, no expression was found in endometrial hyperplasia. Moreover, electrophysiological experiments confirmed the presence of functioning HERG channels on the plasma membrane of tumour cells. On the whole, these data are the first demonstration of the presence of HERG channels in primary human neoplasias, and could candidate HERG as a potential tool capable of marking cancerous versus hyperplastic endometrial growth. PMID- 11104573 TI - Regulation of endometrial cancer cell growth by luteinizing hormone (LH) and follicle stimulating hormone (FSH). AB - Gonadotrophin releasing hormone analogues (GnRHa) have been used to treat recurrent endometrial cancer. However, the mode of action is uncertain. Our previous studies showed no direct effect of GnRHa on endometrial cancer cell growth in vitro. We have now examined the effect of luteinizing hormone (LH) and follicle stimulating hormone (FSH) on endometrial cancer cell growth. The aim was to determine whether suppression of pituitary LH and FSH by GnRHa could explain the tumour regression seen in up to 44% of patients treated with this drug. We show that recombinant human LH and FSH (rhLH and rhFSH) produce a concentration dependent stimulation of the endometrial cancer cell line HEC-1A, in serum-free medium (maximum increase of 62 and 50% respectively relative to untreated controls). This increase is equivalent to that obtained by addition of 10% newborn calf serum. Growth of the Ishikawa cell line in culture increases in the presence of rhLH (maximum increase of 67%) but not with rhFSH. Using RT-PCR, we show that the Ishikawa cell line intermittently expresses receptor mRNA of LH but not of FSH; there is no expression of either mRNA by HEC-1A. Classically, both LH and FSH act via cAMP linked membrane receptors. However, neither rhLH nor rhFSH elicit cAMP production in either of our endometrial cancer cell lines. Thus, although a growth response to LH and FSH can be shown, and some cells express the LH receptor, stimulation appears to be via a pathway separate from that of the classical gonadotrophin receptor. PMID- 11104574 TI - Glycerol restores p53-dependent radiosensitivity of human head and neck cancer cells bearing mutant p53. AB - Mutation or inactivation of p53 is known to be present in approximately 50% of human cancers. We propose here a novel strategy for overcoming this problem in mutant p53-targeting cancer therapies. We examined the restoration of radiation induced p53-dependent apoptosis by a chemical chaperone (glycerol) in human head and neck cancer cells (SAS cells, showing wild-type p53 phenotype). SAS cells transfected with mutant p53 (SAS/m p53) showed radioresistance compared with SAS cells (SAS/ neo) transfected with neo vector as a control, but became radiosensitive when pre-treated with glycerol before X-ray irradiation. Apoptosis in the SAS/m p53 cells was induced by X-rays with glycerol pre-treatment, but not without glycerol pre-treatment, whereas apoptosis in the SAS/ neo cells was induced in both cases. Gel mobility-shift assays showed that after X-ray irradiation combined with glycerol pre-treatment, mp53 was able to bind to the sequence-specific region upstream of the bax gene regulating apoptosis. These results suggest that glycerol is effective in inducing a conformational change of p53 and restoring normal function to mp53, leading to enhanced radiosensitivity through the induction of apoptosis. This novel tool for enhancement of radiosensitivity in cancer cells bearing mp53 may be useful for p53-targeted radiotherapy. PMID- 11104575 TI - Characterization of DNA-strand breakage induced in V79 cells by F 11782, a catalytic inhibitor of topoisomerases. AB - DNA damage induced in V79 cells by F 11782, or 2",3"-bis pentafluorophenoxyacetyl 4',6'-ethylidene-beta-D glucoside of 4'-phosphate-4'-demethylepipodophyllotoxin 2N-methyl glucamine salt, a novel dual catalytic inhibitor of topoisomerases I and II, was investigated using both alkaline and neutral versions of the comet assay methodology. A comparison was then made of the DNA damage induced by F 11782 with that induced by either etoposide or camptothecin under comparable experimental conditions. The results revealed that F 11782 initially induced less DNA strand breaks that either etoposide or camptothecin and rejoined such breaks more slowly. However, unlike these other drugs, the extent of DNA damage induced by F 11782 increased linearly with time of incubation. F 11782 produced both DNA single- and double-strand breaks without any clear specificity relative to phase of the cell cycle, although proliferating cells were preferentially damaged. The marked time-dependency of induction of DNA strand breaks by F 11782 may serve to explain, at least in part, its major in vivo antitumour activities. PMID- 11104576 TI - Timing within the oestrous cycle modulates adrenergic suppression of NK activity and resistance to metastasis: possible clinical implications. AB - Clinical observations suggest that the rate of metastatic development and long term mortality following surgery in breast cancer patients is influenced by the menstrual phase during which surgery is conducted. The menstrual cycle is known to modulate various physiological responses and medical conditions that involve adrenergic mechanisms (e.g., asthma). Natural killer activity (NKA), an immune function controlling metastasis, is suppressed following surgery, and in vitro by adrenaline. We therefore hypothesize that the clinical observation may be partly attributable to surgery-induced adrenergic suppression of NK-dependent resistance to metastasis, a suppression that depends on menstrual phase during surgery. To test this hypothesis in rats, 140 F344 females at different phases of their oestrous cycle were injected with a beta-adrenergic agonist, metaproterenol (MP) (0.4 or 0.8 mg kg(-1), s.c.), or with vehicle, before i.v. inoculation with MADB106 tumour cells. This syngeneic mammary adenocarcinoma line metastasizes only to the lungs, and is highly sensitive to NKA. In a second experiment, the suppression of NKA by MP was studied in vitro in blood drawn at different phases of the oestrous cycle (n = 36). Finally, the effects of stress on the number and activity of NK cells were assessed along the oestrous cycle (n = 71). The findings indicate that the suppressive effects of MP on resistance to metastasis and on NKA, are significantly greater during the oestrous phase characterized by high oestradiol levels (D3/proestrus/oestrus). Similarly, NKA per cell was suppressed by stress only during this phase. In untreated animals, in which inadvertent stress was minimized, no effects of the oestrous cycle on NKA or on resistance to metastasis were evident. These findings indicate that the oestrous cycle modulates adrenergic suppression of NKA and of resistance to metastasis. The relevance of these findings to the above clinical observation, as well as that of our related findings in women from a parallel study, is discussed. PMID- 11104577 TI - Anti-CD7 antibody and immunotoxin treatment of human CD7(+)T-cell leukaemia is significantly less effective in NOD/LtSz-scid mice than in CB.17 scid mice. AB - Groups of 8 to ten SCID (CB.17 scid/scid) or NOD/SCID (NOD/LtSz- scid/scid) mice were injected i.v. with two million human HSB-2 T-ALL cells on day 1 (SCID-HSB-2 and NOD/SCID-HSB-2 mice) and treated later with 3 i.v. 10 microg doses of the anti-CD7 antibody HB2 on days 7, 9 and 11 or with a single 10 microg dose of HB2 SAPORIN or a 7.4 microg dose of HB2-F(ab)(2)-SAPORIN immunotoxin (IT) on day 7. Treatment of SCID-HSB-2 mice with HB2-SAPORIN led to a significant prolongation in the time to development of signs and symptoms of disease compared with PBS sham-treated controls with 80% of animals surviving disease-free. In contrast treatment with HB2-F(ab)(2)-SAPORIN was significantly less effective in SCID-HSB 2 mice with 80% of animals in this treatment group developing leukaemia over the course of the study. HB2 antibody treatment of SCID-HSB-2 mice also led to a significant prolongation in time to leukaemia development compared with sham treated controls with 37% of animals in this treatment group disease-free at termination of the study. In contrast HB2 antibody treatment of NOD/SCID-HSB-2 mice had no therapeutic effect in these animals and the therapeutic effectiveness of both HB2-SAPORIN and HB2-F(ab)(2)-SAPORIN ITs was similar and significantly reduced compared to the effect observed in SCID-HSB-2 mice. It was initially thought that the lack of therapeutic effect of antibody and IT in NOD-SCID-HSB-2 mice might relate to their putative lack of NK cells but flow cytometric and functional studies with NOD-SCID mouse splenocytes revealed that these animals do have some functional NK cells though fewer in number and possibly lower in functionality than those of SCID mice. We reason that the complete lack of therapeutic effect of HB2 antibody and the reduced effect of HB2-SAPORIN in NOD/SCID mice is due to the reduced cytolytic activity of NOD/SCID NK cells which is probably below a certain critical threshold value in these animals. We conclude from this that immunotherapeutics like HB2-SAPORIN would be more accurately assessed for intrinsic potency in NOD/SCID mice where the effects of NK cell and possibly other non-adaptive immune mechanisms would not have a significant influence. PMID- 11104578 TI - Antitumour activity of novel taxanes that act at the same time as cytotoxic agents and P-glycoprotein inhibitors. AB - Taxanes antitumour agents such as paclitaxel and docetaxel represent a successful family of chemotherapeutic drugs. Unfortunately, acquired and innate resistance represents a clinical problem for these drugs. We investigated, on a panel of 7 human cancer cell lines, the growth inhibition effect of 3 newly developed taxanes (SB-T-1213, SB-T-1250 and SB-T-101187) with modification at the C10 and C3' positions of the taxane framework. These positions have been previously characterized as critical to make taxanes highly active against cells overexpressing the efflux pump P-glycoprotein (P-gp). Paclitaxel and docetaxel were used as reference compounds. Results unambiguously indicate the exceptional activity of the novel taxanes toward P-gp positive cells (up to >400 fold higher potency than that of paclitaxel). SB-T-1213 and SB-T-1250 are also substantially more active than the reference compounds against P-gp negative cells. To better understand the mechanisms underlying the enhanced activity of the newly developed taxanes, we performed cell cycle and apoptosis analysis. This study demonstrates that the striking growth inhibition effect exhibited by the novel taxanes is ascribed to their increased ability in inducing apoptosis and G(2)/M cell cycle block. SB-T-1213 and SB-T-1250 are also more active than reference compounds in inducing intracellular accumulation of the beta-tubulin subunits. Finally, it is revealed that these novel taxanes have ability to inhibit the function of the P gp efflux pump on the basis of the Rhodamine 123 assay. These findings strongly suggest that SB-T-1213, SB-T-1250 and SB-T-101187 represent a new tool to overcome innate or acquired P-gp mediated taxane-resistance. PMID- 11104579 TI - Identification of a ras oncogene peptide that contains both CD4(+) and CD8(+) T cell epitopes in a nested configuration and elicits both T cell subset responses by peptide or DNA immunization. AB - Mutations in ras proto-oncogenes are commonly found in a diversity of malignancies and may encode unique, non-self epitopes for T cell-mediated antitumor activity. In a BALB/c (H-2(d)) murine model, we have identified a single peptide sequence derived from the ras oncogenes that contained both CD8(+) and CD4(+) T cell epitopes in a nested configuration. This peptide reflected ras sequence 4-16, and contained the substitution of Gly to Val at position 12 ?i.e., 4-16(Val12)?. Mice immunized with this 13-mer peptide induced a strong antigen (Ag)-specific CD4(+) proliferative response in vitro. In contrast, mice inoculated with the wild-type ras sequence failed to generate a peptide-specific T cell response. Additionally, mice immunized with the ras 4-16(Val12) peptide concomitantly displayed an Ag-specific CD8(+) cytotoxic T lymphocyte (CTL) response, as determined by lysis of syngeneic tumor target cells incubated with the nominal 9-mer nested epitope peptide ?i.e., 4-12(Val12)?, as well as lysis of tumor target cells expressing the corresponding ras codon 12 mutation. Analysis of the Valpha- and Vbeta-chains of the T cell receptor (TCR) expressed by these CTL revealed usage of the Valpha1 and Vbeta9 subunits, consistent with the TCR phenotype of anti-ras Val12 CTL lines produced by in vivo immunization with the nominal peptide epitope alone. Moreover, immunization with the nested epitope peptide, as compared to immunization with either the 9-mer CTL peptide alone or an admixture of the 9-mer CTL peptide with an overlapping 13-mer CD4(+) T cell helper peptide ?i.e., 5-17(Val12)? lacking the class I N-terminus anchor site, enhanced the production of the CD8(+) T cell response. Finally, immunization with plasmid DNA encoding the ras 4-16(Val12) sequence led to the induction of both Ag specific proliferative and cytotoxic responses. Overall, these results suggested that a single peptide immunogen containing nested mutant ras-specific CD4(+) and CD8(+) T cell epitopes: (1) can be processed in vivo to induce both subset specific T lymphocyte responses; and (2) leads to the generation of a quantitatively enhanced CD8(+) CTL response, likely due to the intimate coexistence of CD4(+) help, which may have implications in peptide- or DNA-based immunotherapies. PMID- 11104580 TI - Expression of delta opioid receptors by splenocytes from SEB-treated mice and effects on phosphorylation of MAP kinase. AB - Delta opioid receptors (DORs) are known to modulate multiple T-cell responses. However, little is known about the expression of these receptors. These studies evaluated the expression of DOR mRNA and protein after a single in vivo exposure to staphylococcal enterotoxin B (SEB). SEB (20 microg, ip) significantly enhanced splenocyte DOR mRNA expression 8 and 24 h after injection. SEB also increased the fractions of the total splenocyte (5 to 20%) and T-cell (8 to 50%) populations expressing DOR protein. In saline-treated animals, DOR relative fluorescence intensity per cell was 11.1 +/- 0.62 units (mean +/- SEM), increasing to 16.1 +/- 1.7 after exposure to SEB. DOR fluorescence intensity significantly increased to 33.5 +/- 2.0 units in a subpopulation of T-cells. Thus, SEB significantly increased DOR expression in vivo, affecting both mRNA and protein levels primarily within the T-cell population. To determine whether T-cell DORs modulate the activity of extracellular-regulated kinases (ERKs), the phosphorylation of ERKs 1 and 2 was studied using splenocytes from SEB-treated mice. At concentrations from 10(-8) to 10(-6) M, [d-Ala(2)-d-Leu(5)]-enkephalin, a selective DOR agonist, significantly inhibited anti-CD3-epsilon-induced phosphorylation of the ERKs. Therefore, the DORs expressed by activated T-cells are capable of attenuating T-cell activation that depends on ERK phosphorylation. PMID- 11104581 TI - Differential production of prostaglandin E(2) in male and female mice subjected to thermal injury contributes to the gender difference in immune function: possible role for 15-hydroxyprostaglandin dehydrogenase. AB - We have previously reported a macrophage-mediated gender difference in postburn immunosuppression, which was dependent upon elevated levels of circulating 17beta estradiol (E(2)) and, in part, interleukin-6. Herein we examined the role of prostaglandin E(2) (PGE(2)), a potent suppressor of cell-mediated immunity. Circulating levels of PGE(2) were significantly elevated in females but not males at 10 days postburn (P < 0.01), and indomethacin treatment fully restored the delayed-type hypersensitivity and splenocyte proliferative responses of thermally injured females. While there was no difference in cyclooxygenase-2 protein expression in the lungs and liver of thermally injured male and female mice, there was a marked decrease in the protein expression of 15-hydroxyprostaglandin dehydrogenase in females. These data demonstrate that PGE(2) is a critical mediator of immunosuppression in thermally injured female mice and that the increase in circulating PGE(2) is derived, in part, from decreased degradation and clearance of PGE(2). PMID- 11104582 TI - Effects of raloxifene, a selective estrogen receptor modulator, on thymus, T cell reactivity, and inflammation in mice. AB - Raloxifene is a selective estrogen receptor modulator approved for prevention of osteoporosis in postmenopausal women. It is selective by virtue of having estrogen agonistic effects in bone, vessels, and blood lipids, while it is antagonistic with mammary and uterine tissue. The aim of the study was to examine whether the raloxifene analogue LY117018 (LY) has estrogenic effects on the thymus, T cell responsiveness, and inflammation. Oophorectomized normal mice were treated with subcutaneous injections of equipotent antiosteoporotic doses of LY (3 mg/kg) and 17beta-estradiol (E2) (0.1 mg/kg) or vehicle as controls. Effects on thymus were studied by analyses of thymus weight, cellularity, and CD4 and CD8 phenotype expression and histology, while inflammation was determined as T-cell mediated delayed-type hypersensitivity (DTH) and granulocyte-mediated footpad swelling. LY lacked the suppressive properties of E2 on DTH and granulocyte mediated inflammation. Furthermore, LY induced only minor thymus atrophy compared with E2 and did not, in contrast to E2, alter the thymic CD4/CD8 phenotypes. These results clearly demonstrate that raloxifene principally lacks the modulatory effects of estrogen on T cell responsiveness and inflammation. Our data are discussed in the context of recent findings in estrogen receptor biology and also with respect to estrogen-mediated alteration of autoimmune rheumatic diseases. PMID- 11104583 TI - Neonatal (cord blood) T cells can competently raise type 1 and 2 immune responses upon polyclonal activation. AB - In the neonate, cellular immunity has generally been hypothesized as being incompetent. Accumulating evidence from several recent studies, together with our present report, contradicts this hypothesis. T-helper cell and T cytotoxic type 1 and 2 (Th1/Th2 and Tc1/Tc2, respectively) cytokine responses to polyclonal T cell receptor (TCR) activation were assessed in medium-term cultures of human cord blood T cells using intracellular cytokine staining, which could measure the frequencies of cytokine-producing cells. In this study, we examined the responses of cord blood CD4(+) and CD8(+) T cells in regard to the production of interferon (IFN)-gamma and interleukin (IL)-4 and compared the responses with those obtained from T cells of healthy adults. We found that the responses in cord blood T cells activated with TCR stimulation were comparable to those of their adult counterparts. Moreover, the Th/Tc cells that developed in cord blood were as competent as adult cells for both IFN-gamma and IL-4 secretion. In addition, IL 12 production, which is critical for both Th1 and Tc1 responses, was equally comparable in the two groups. The production of two major cross-regulatory cytokines, tumor necrosis factor-alpha and IL-10, was similarly comparable and not significantly different between the two groups. Taken together, these results indicate that, though naive, the neonatal T cell is competent to respond to TCR mediated stimulation and to produce both type 1 and type 2 cytokines. PMID- 11104584 TI - Morphine Up-regulates expression of substance P and its receptor in human blood mononuclear phagocytes and lymphocytes. AB - In vitro and in vivo studies have indicated that there is an important relationship between morphine and neuropeptide substance P (SP). We therefore investigated the interaction of morphine and cultured human immune cells on the expression of SP, a neuropeptide which we have recently demonstrated to be produced by human monocytes and lymphocytes. Morphine up-regulated SP production in human mononuclear phagocytes and lymphocytes at both the mRNA and the protein level. In addition, morphine induced SP receptor (NK-1R) expression in human lymphocytes. The specific morphine receptor antagonist (naltrexone) blocked morphine-induced SP expression in human mononuclear phagocytes, supporting the concept of authentic morphine receptor-mediated regulation. Since SP modulates neurogenic inflammation and immunologic events, these data suggest that morphine induced SP expression in cells of the immune system may be of importance in the pathogenesis of immune-mediated diseases, including neuroimmunologic diseases and AIDS. PMID- 11104585 TI - Thrombin induces IL-6 but not TNFalpha secretion by mouse mast cells: threshold level thrombin receptor and very low level FcepsilonRI signaling synergistically enhance IL-6 secretion. AB - Mast cells become activated in multiple diseases wherein thrombin generation is often clinically apparent, but the effect of thrombin on cytokine release by mast cells remains unexplored. Thus, we examined IL-6 and TNFalpha release by thrombin challenged mast cells. Thrombin and the protease-activated receptor (PAR)-1 peptide TRAP(14) induced these cells to secrete IL-6 in a dose-dependent fashion. Mast cells secreted > or =2800 pg IL-6/10(6) cells over 24 h, but only low levels of serotonin and no significant TNFalpha. Furthermore, at near-background levels of allergen, threshold doses of alpha-thrombin synergistically enhanced the IL-6 response (by up to 100-fold), but high-dose costimulation led to a simple additive response. Both the PI(3)- and sphingosine-kinase signaling pathways contributed importantly to the thrombin response. Our data thus clearly demonstrate that low-level thrombin and FcepsilonRI signaling can synergize to augment mast cell IL-6 responses, and that thrombin also differentially induces cytokine secretion by mast cells. PMID- 11104586 TI - Glutamine supplemented nutrition support: saving nitrogen and saving money? PMID- 11104587 TI - Hunger disease. AB - This paper examines three aspects of hunger disease: the effect of initial fat stores on macronutrient fuel selection during total starvation (no energy) and how it influences survival; the effects of different rates of weight loss on tissue and body function; and the importance of appetite sensations, including hunger, during malnutrition and during enteral and parenteral nutritional support. Long-term starvation studies in humans reveal major differences in fat carbohydrate and protein metabolism between lean and obese subjects, including a 2-4-fold lower contribution of protein oxidation to energy expenditure in obese subjects, which ensures that more of the excess body fat is oxidized. The rate of weight loss, determined by recent dietary intake, can have major effects on tissue and body function, including wound healing, the acute phase protein response, muscle fatigue and psychological/behavioural function in both clinical and non-clinical settings. In depleted states uncomplicated by disease, changes in appetite sensations can result in energy intakes as high as 6000 to 10,000 kcal/day ( 25-42 MJ/day). Long-term enteral tube feeding and parenteral nutrition are associated with frequent disturbances in appetite sensations, and in those able to eat normally they tend to add rather than replace oral intake to an extent that appears to depend on the regimen. It is concluded that 1) differences between lean and obese subjects in macronutrient fuel selection during starvation are adaptive because they optimize survival in both groups of subjects; 2) the rate of weight loss in health and disease has a major effect on certain tissue and body functions, independently of the magnitude of weight loss; and 3) clinically relevant disturbances in appetite sensations are common subjects receiving long-term enteral and parenteral nutrition. The clinical modulation of all these variables would be aided by greater knowledge of the mechanisms involved. PMID- 11104588 TI - Pathogenesis of malnutrition in cystic fibrosis, and its treatment. AB - PATHOGENESIS: We have developed a model of the pathogenesis of malnutrition in cystic fibrosis. It consists of the relationship between nutrient balance and nutrient requirement. The validation has been conducted with respect to energy, but the same general principals can be applied to any nutrient. A patient with CF either loses weight or fails to grow normally if their absorbed energy intake is less than their total daily energy expenditure. Multiple factors have the potential to contribute to reduced energy intake including, anorexia, gastroeosophageal (GE) reflux leading to vomiting and hence food loss, as well as maldigestion. Another more recently recognized source of energy loss, is glucosuria as a result of CF related diabetes (CFRD). Conversely, lung inflammation appears to be related to increases in resting metabolic rate (RMR). Acute exacerbations of the chronic lung disease increases RMR which returns to a basal level some weeks after the inflammation is treated. In clinically stable patients with CF, RMR rises in a quadratic fashion as lung function falls. When FEV(1)is >85% predicted RMR is not different from controls, but it rises in a curvilinear fashion as FEV(1)falls. Initially it appears that patients adapt to their increased RMR by reducing their activity so their total daily energy expenditure (TDEE) is often no higher than controls. But this is by no means always the case. Furthermore good lung care requires CF patients to be involved in aerobic activities, hence their TDEE would rise. Although there has been considerable interest as to whether the genetic defect has an energy wasting effect, it appears genetic factors have little or no effect on RMR. TREATMENT: This starts with making an energy diagnosis. First, a 3 day faecal fat balance study is conducted. This provides information with regard to intake as well as to maldigestion. In addition a history of GE reflux is sought, since it can readily be treated with H(2)-blockers. If significant fat malabsorption exists, efforts are made to improve pancreatic enzyme dose and function. The possibility of CFRD also needs to be considered. We measure the RMR of the patient using open circuit indirect calorimetry. Recommendations for diet therapy are based on estimated TDEE, which is determined from RMR taking into account faecal losses. Diet therapy places the emphasis on increasing the fat content of the diet. We have conducted a study to determine whether or not oral supplements help increase TDEE and they did not; they merely replaced food energy. Conversely, nocturnal gastrostomy supplemental feeding, while reducing voluntary food energy intake by about 20%, does result in a significant increase in total daily energy intake. Our target is to achieve a completely normal nutritional status. Long term follow up of these patients has shown significantly better survival in patients who achieve normal nutritional status. The advent of lung transplantation has added another dimension. In our experience, following a successful lung transplant, most patients no longer need their supplemental gastrostomy feeding. SUMMARY: Our clinic policy is to encourage a high fat diet (35-40% total energy) and our patients grow normally in height and weight until their lung disease deteriorates significantly. Patients who develop a negative energy balance seldom if ever respond to diet therapy and hence are candidates for supplemental nocturnal gastrostomy feeds. Gastrostomy fed patients constitute 3 to 5% of our total CF population of approximately 590 patients. PMID- 11104590 TI - Lean body mass changes in cancer patients with weight loss. AB - BACKGROUND AND AIMS: Metabolic measurements (e.g. resting energy expenditure) are adjusted to lean body mass to account for body composition differences. Usually lean body mass is estimated from total body water. However, this may be compromised in weight-losing cancer patients owing to alterations in the degree of hydration of the lean body mass. This study examined the relationship between two independent estimates of lean body mass in healthy subjects and cancer patients with weight loss. METHODS AND RESULTS: Height, weight, total body water and total body potassium were measured in healthy subjects (n=9) and weight losing cancer patients (n=13). They were similar in terms of age and gender. However, the cancer group had a significantly lower percentage ideal body weight (P<0.001). The measured total body water values in both groups were similar to those predicted. In contrast, measured total body potassium values in the cancer group were significantly lower than predicted (P<0.001). There was a correlation between the ratio of measured lean body mass (water/lean bodymass (potassium) and the percentage weight loss (r=0.698, P<0.001). CONCLUSIONS: These results suggest that total body water significantly overestimates metabolically active tissue in weight-losing cancer patients and therefore its use as the basis for metabolic requirements in this group of patients is questionable. PMID- 11104589 TI - Cost containment through L-alanyl-L-glutamine supplemented total parenteral nutrition after major abdominal surgery: a prospective randomized double-blind controlled study. AB - BACKGROUND & AIMS: Glutamine is recognized as a conditionally essential amino acid. Recent studies indicate that glutamine-containing total parenteral nutrition improves nitrogen economy, enhances gastrointestinal and immune functions and shortens hospital stay. METHODS: Thirty-seven patients (19 w and 18 m; age 61. 4+/-10.4 years; BMI 23.7+/-2.8 kg/m(2)) following major abdominal surgery receiving an isonitrogenous isoenergetic TPN with or without alanyl glutamine supplementation (0.5 g/kg BW/day), were evaluated in a double-blind, randomized, controlled trial over a five-day period by measuring nitrogen balance, selected biochemical parameters and length of hospital stay. RESULTS: Supplemental alanyl-glutamine improved the overall mean (-3.5+/-1.6 vs. -5.5+/-1. 4 g N;P<0.05) and cumulative nitrogen balance (-14.1+/-9.1 vs. -21.7+/-11.4 g N;P<0.05) compared with the isonitrogenous, isoenergetic standard regimen. Alanyl glutamine normalized plasma glutamine concentration and reduced the length of hospital stay (12.8+/-2.6 vs. 17.5+/-6.4 days;P<0.05). CONCLUSIONS: The results of the study confirm that supplementation with synthetic alanyl-glutamine dipeptide is associated with cost containment due to shortened hospitalization and improved nitrogen economy. PMID- 11104591 TI - Postoperative enteral immunonutrition in head and neck cancer patients. AB - AIMS: to determine if postoperative feeding of head and neck cancer patients, using an enteral diet supplemented with arginine, improves immunological and nutritional status, and clinical outcome, i.e., reduces postoperative infectious/wound complications and length of stay, when compared with an isocaloric, isonitrogenous control diet. METHODS: at operation 44 patients were randomized into two groups to receive: a) an enriched diet (n=23);b) an isocaloric, isonitrogenous control diet (n=21). Thirteen patients with a history of significant weight loss (> or = 10% over the last 6 months) were considered malnourished. Preoperatively and on postoperative days 1, 4 and 8 the following parameters were evaluated: albumin, prealbumin, transferrin, total number of lymphocytes, lymphocyte subsets (CD3, CD4, CD8 and CD4/CD8 ratio) and immunoglobulins. Postoperative complications and length of stay were recorded. RESULTS: 'visceral' serum proteins and immunological parameters decreased on postoperative day 1 in both groups. However, only the enriched group demonstrated a significant increase (P<0.05) in the total number of lymphocytes, CD4, CD4/CD8 on postoperative day 4, and total number of lymphocytes, CD3, CD4, CD4/CD8 on postoperative day 8. In the malnourished subgroup the administration of the enriched formula significantly reduced both postoperative infectious/wound complications and length of stay compared with the control group (P<0.05). CONCLUSIONS: enteral immunonutrition of head and neck cancer patients improves postoperative immunological response. Significant clinical advantages were observed in malnourished patients. PMID- 11104592 TI - Influence on gallbladder volume of early postoperative gastric supply of nutrients. AB - BACKGROUND & AIM: The gallbladder volume is a predictor of biliary stasis and the formation of biliary sludge. Biliary stasis and sludge have been recently recognized as the precursors of acute acalculous cholecystitis, as well as 'idiopathic' postoperative pancreatitis, rare but very serious complications after surgery. The aim of the study was to establish how early postoperative gastric supply of nutrients affects the gallbladder volume in patients after noncardiac and cardiac surgery. METHODS: In the two prospective, randomized studies 40 patients (study I-noncardiac surgery) treated at surgical ICU after major elective extrahepatobiliary and extragastrointestinal surgeries (7 thoracic, 19 vascular, 14 urological) and 40 patients (study II-cardiac surgery) treated at cardiosurgical ICU after CABG surgery were analyzed. In both studies the patients were divided into two groups: control group C (study I: 20 patients, age 45+/-18 yrs, male 65%; study II: 20 patients age 58+/-7 yrs, male 60%) and group E (group of early postoperative gastric supply of nutrients) (study I: 20 patients, age 52+/-17 yrs, male 50%; study II: 20 patients; age 59+/-8 yrs, male 65%). For the first 24 hours the patients in group C received only crystalloid solutions and the gallbladder volume was verified 24 hours after the surgery. In group E, postoperative gastric supply of nutrients began 18 hours after surgery (Osmolite, Ross; first 3 hours 30 ml/h and second 3 hours 50 ml/h; total 240 ml after 6 hours). In all patients sonographic measurement of gallbladder volume was performed immediately before surgery and 6 hours after the start of feeding (24 hours after surgery). The measurement was done with ultrasonographic scanner Hitachi 405 EUB (convex probe 3.5-5MHz) by the same specialist, and the volume was calculated using the ellipsoid method. RESULTS: The gallbladder volume measured by ultrasonography 24 hours after surgery in study I (noncardiac surgery) in group E amounted to 43+/-25 ml while in control group C it was significantly higher, i.e. 67+/-30 ml (P<0.05). In study II (cardiac surgery) in group E gallbladder volume amounted to 59+/-15 ml while in control group C it was also significantly higher, i.e. 71+/-11 ml (P<0.05). CONCLUSION: An early postoperative gastric supply of nutrients after both noncardiac and cardiac adult surgery diminishes the volume and probably stimulates the motility of the gallbladder, thus preventing biliary stasis and the formation of biliary sludge. PMID- 11104593 TI - Eicosapentaenoic acid ethyl ester supplementation in cachectic cancer patients and healthy subjects: effects on lipolysis and lipid oxidation. AB - BACKGROUND & AIMS: Recent reports suggest that weight loss in cachectic cancer patients may be inhibited by supplementation of the n-3 fatty acid eicosapentaenoic acid (20:5n-3; EPA), presumably due to inhibition of lipolysis. The aim of the present double-blind, randomized trial was to assess whether short term oral EPA ethyl ester (EE) supplementation inhibits lipolysis and lipid oxidation in weight-losing cancer patients and in healthy subjects. METHODS: Seventeen weight-losing, cancer patients of different tumor types, and 16 healthy subjects were randomized to receive EPA-EE (6 g/d) or placebo (oleic acid (OA) EE; 6 g/d) for seven days. At baseline (day 0) and during supplementation (days 2 and 7) whole-body lipolysis and palmitic acid release were measured in the overnight fasting state using [1, 1, 2, 3, 3-(2)H(5)]glycerol and [1 (13)C]palmitic acid. Palmitate oxidation was determined by measuring(13)CO(2)enrichment in expired breath. RESULTS: No significant effects of EPA-EE on whole-body lipolysis, palmitic acid release, or palmitate oxidation were detected in cancer patients nor in healthy subjects in comparison with OA EE. EPA-EE supplementation reduced plasma-free fatty acid and triacylglycerol concentrations significantly in healthy subjects but not in cancer patients. CONCLUSION: We conclude that supplementation of EPA-EE does not significantly inhibit lipolysis or lipid oxidation in weight-losing cancer patients or in healthy subjects during short-term supplementation when using OA-EE as a placebo supplement. PMID- 11104594 TI - Simplification of the method of assessing daily and nightly energy expenditure in children, using heart rate monitoring calibrated against open circuit indirect calorimetry. AB - Total daily energy expenditure (TDEE) can be assessed in children using several methods: the double-labelled water technique (DLW); indirect whole-body calorimetry; activity diary; accelerometry and heart rate (HR) monitoring. This last, low-cost and convenient technique has been validated against the DLW technique that is normally considered the reference method. The main advantages of HR monitoring are its social acceptability, and the examination of the time spent in specific activities or intensive exercise sessions. The aim of this study was to assess a new method for computing energy expenditure (EE) with the HR monitoring technique in children, using an open-circuit ventilated hood indirect calorimetry system. Eleven healthy children participated in this study, with a mean age of age 8.9+/-3 years. Seven of them were studied again 6 months later, so that 17 measurements were available for analysis. The calibration period was used to collect simultaneous data from HR and EE measured by indirect calorimetry (IC). Measurements were made when the children were resting (REE=Resting Energy Expenditure), during 30 min of a post-prandial period, and during two different activity levels on a cyclergometer (two periods of 15 min at 20 and 60 W output). Results of EE during a longer calibration period (2.5 h) were compared with those of a shorter period (1.5 h), for a group of six subjects. Results of nightly EE measurements of five subjects were recorded using IC and compared with the HR predicted method. Results of EE measurements estimated with different regression equations were compared to EE measured by IC. The short calibration period (1.5 h) gave similar results to a longer period (2.5 h), with a mean difference of less than 3%. The polynomial third-order relationship between HR and EE gave the highest correlation coefficient, the smallest mean square error and the best agreement (Bland-Altman method). EE predicted with this model gave the best results during the total calibration period, the post-prandial period and the medium and high activity periods. Comparison between measured and predicted nightly EE showed that REE - REE/10 was the best formula to assess nightly EE, and the mean difference was less than 4%. This study demonstrates the validity of a shortening of the calibration period and the reliability of the REE - REE/10 formula to compute nightly EE. The best model for computing daily EE is a polynomial third-order regression equation. PMID- 11104595 TI - Perioperative enteral nutrition and quality of life of severely malnourished head and neck cancer patients: a randomized clinical trial. AB - BACKGROUND AND AIMS: This study evaluated the use of perioperative nutritional support on Quality of Life (QOL) in malnourished head and neck cancer patients undergoing surgery. METHODS: 49 Malnourished (weight loss >10%) head and neck cancer patients who were included in a nutrition intervention trial were randomized to receive either no preoperative and standard postoperative tube feeding (group I), standard preoperative and postoperative tube-feeding (group II) or arginine-supplemented preoperative and postoperative tube-feeding (group III). Of these patients, 31 completed a full QOL assessment on the first day of preoperative nutritional support, one day before surgery, and 6 months after surgery. Both a disease-specific (EORTC QLQ-C30) and a generic questionnaire (COOP-WONCA) were used. One way analysis of variance (ANOVA) and the Kruskal Wallis test were applied for testing differences in scores between groups. RESULTS: Between baseline and the day before surgery, both preoperatively fed groups revealed a positive change for the dimensions physical and emotional functioning and dyspnea (with significance in group II, P=0.050,0.031,0.045 respectively). Group III showed a negative change in appetite (P=0.049). Between baseline and 6 months after surgery, there were no differences between group I and both pre-fed groups. There were no differences in favour of group III compared to group II. CONCLUSION: Enteral nutrition improves QOL of severely malnourished head and neck cancer patients in the period preceding surgery. No benefit of preoperative enteral feeding on QOL could be demonstrated 6 months after surgery. PMID- 11104596 TI - High food wastage and low nutritional intakes in hospital patients. AB - BACKGROUND AND AIMS: The aim of this study was to investigate the cause of continuing weight-loss in hospitalized patients. We determined 1. whether the hospital menu was able to meet the patients' minimum nutritional requirements, 2. the proportion of food being wasted and 3. the mean nutritional intakes of patients. METHODS: This study was carried out in a University hospital (1200 beds). All the food supplied and wasted was measured over a 28 day period on one ward in each of 4 different specialties. Average food intake per patient was calculated and checked against individual food intake measurements. RESULTS: The hospital menu provided over 2000 kcal/day and could meet patients' nutritional requirements. However, high wastage rates of greater than 40% resulted in energy and protein intakes within all specialties being less than 80% of that recommended. The cost of this waste was 139,655 pounds sterling in these four specialties. CONCLUSIONS: More than 40% of hospital food was wasted. Energy and protein intakes were low and patients did not, therefore, meet their recommended intakes. This helps to explain continuing weight-loss in hospital patients and represents a large waste of resources. Hospital feeding policies therefore need reviewing and made more appropriate to the needs of the sick. PMID- 11104597 TI - A recipe for improving food intakes in elderly hospitalized patients. AB - BACKGROUND & AIMS: The aim of this study was to compare food wastage and intake between the normal hospital menu and one where more energy dense but smaller portions were provided. METHODS: This study was carried out on an Elderly Rehabilitation ward in a University hospital. Patients were randomly allocated to receive either a normal or a reduced portion size fortified menu for a 14 day cycle and then swapped-over at the end of each cycle for the 56 day study. One group received a cooked breakfast and normal menus throughout the study. RESULTS: All the menu combinations could meet the patients recommended intake. The fortified menu provided 14% more energy than the normal menu. Food wastage was highest in the cooked breakfast group (32%) and lowest in the Fortified group (27%). The total weight of wasted food was less than in the previous study. Nutritional intakes were 25% higher on the fortified menu compared with the normal menu. The mean protein intakes were still below that recommended. All patients had higher energy intakes on the Fortified menu compared with their intake on the normal menu despite being served a lower weight of food. CONCLUSIONS: We conclude from our own data and that of others that it is possible for elderly patients to achieve their nutritional targets using a combination of smaller portions of increased energy and protein density and between-meal snacks. The needs of other groups of patients also needs to be assessed in a similar way to make hospital food appropriate to the needs of the sick. PMID- 11104598 TI - Pregnancy in a patient with chronic intestinal pseudo-obstruction on long-term parenteral nutrition. AB - Parenteral nutrition support is provided in most instances for short intervals during pregnancy in conditions where oral/enteral intake is severely compromised. Few reports describe the use of parenteral nutrition from conception to delivery. We report the case of a 30-year-old woman suffering from a severe form of chronic intestinal pseudo-obstruction on long-term parenteral nutrition because of malabsorption and malnutrition. Pregnancy and delivery developed uneventfully. The fetus grew normally throughout pregnancy. Our patient needed only slight modifications in her parenteral nutrition regimen during lactation. There were no metabolic complications during pregnancy. We conclude that female patients even with severe forms of gastrointestinal diseases, such as chronic intestinal pseudo obstruction requiring long-term home parenteral nutrition, can conceive and carry successfully a pregnancy to term. PMID- 11104599 TI - Metabolic responses to tumour disease and progression: tumour-host interaction. AB - The progressive nutritional deterioration frequently found in cancer patients, is often referred to as cancer cachexia. In contrast to starvation, where it is possible to reverse the body composition changes by the provision of extra calories, in cancer cachexia this reversal is not observed, suggesting that anorexia alone is unlikely to be responsible for this wasting syndrome. Over the past decades a number of studies have focused on the possible mediators which may be responsible for metabolic abnormalities observed in cancer patients. Pro inflammatory cytokines have been strongly implicated, but evidence supporting such a direct role is lacking. Recently, exciting work regarding molecules produced by tumour cells, and which may induce lipolysis and proteolysis, has been published. There is also evidence that increased metabolism of host resources may provide substrates which might promote tumour growth. A number of studies have demonstrated that polyunsaturated fatty acids, such as linoleic and arachidonic acid, are able to promote tumour cell growth either by directly stimulating mitosis or by inhibiting apoptosis. Even more interesting is the discovery of antagonists of these catabolic factors such as eicosapentanoic acid for the lipolytic factor, which may play a role in the treatment of these patients in the near future. PMID- 11104601 TI - Environmental Research, Section B. PMID- 11104600 TI - Spontaneous chromosome breakage in pernicious anemia. PMID- 11104602 TI - An opinion in support of second-look surgery in ovarian cancer. PMID- 11104603 TI - Complete salvage surgical cytoreduction improves further survival of patients with late recurrent ovarian cancer. AB - OBJECTIVE: The aim of this study was to assess the clinical benefit of salvage surgical cytoreduction in patients with late recurrent ovarian cancer. METHODS: Thirty patients with recurrent ovarian cancer who underwent salvage surgical cytoreduction were retrospectively reviewed. All had been initially treated by primary surgery and platinum-based chemotherapy and had a period of clinical remission of at least 6 months. Median time to recurrence was 17.5 months (range, 6-76 months). RESULTS: A macroscopically complete salvage cytoreduction was obtained in 17 (56.7%) patients, whereas 8 patients were left with macroscopic residual disease <2 cm and 5 patients with a larger residuum. Logistic regression showed that the probability of achieving a complete cytoreduction was significantly related to the residual disease after initial surgery (<2 cm versus >2 cm, P = 0.0027, odds ratio = 36.000, 95% confidence interval = 3. 473 373.176), but not to FIGO stage, tumor grade, histologic type, patient age at recurrence, and time to recurrence. In the whole series median survival following salvage surgery was 21 months. Survival was significantly longer in patients who were completely cytoreduced compared to those who were not (median: 37 months versus 19 months, P = 0.04). Moreover, survival was significantly related to time to recurrence (>17.5 months versus <17.5 months, median: 25 months versus 15 months, P = 0.039), number of recurrence sites (single versus multiple, median: 40 months versus 19 months, P = 0. 009), and residual disease after initial surgery (<2 cm versus >2 cm, median: 37 months versus 19 months, P = 0.01), but not to patient age, recurrence site with the largest size, FIGO stage, tumor grade, and histologic type. CONCLUSIONS: The present data seem to show that complete salvage surgical cytoreduction significantly improves further survival of ovarian cancer patients who recur at least 6 months after the completion of primary therapy. PMID- 11104604 TI - The abandoned radical hysterectomy: a Gynecologic Oncology Group Study. AB - OBJECTIVE: The aim of this study was to evaluate the frequency with which intended radical hysterectomy for cervical cancer is abandoned and the outcomes for those patients. METHODS: A secondary evaluation of a prospective surgical pathological trial was performed. There were 1127 patients with Stage IB carcinoma of the cervix entered on Gynecologic Oncology Group Protocol No. 49. These patients were to undergo radical hysterectomy and pelvic lymphadenectomy with careful analysis of pathologic findings, complications, and outcomes. RESULTS: Ninety-eight women were found, at operation, to have extrauterine disease and the proposed radical operation was abandoned at the discretion of the operating surgeon. The records of these patients were evaluated. Subgroups of patients with extrapelvic disease (30) and pelvic extension (26), including grossly positive pelvic nodes (12), other pelvic implants (8), and gross serosal extension (2), were identified. Sixty-three (93%) patients subsequently underwent pelvic radiation therapy and one or two intracavitary applications. Para-aortic fields were added for 8 patients who were found to have positive para-aortic nodes. Five patients received radiotherapy and chemotherapy; 4 patients received chemotherapy alone. One patient declined any further therapy. The disease-free survival was shorter for patients whose radical procedure was abandoned than for those patients who underwent radical hysterectomy. Among the abandoned-operation patients, those with extrapelvic disease had the shortest progression-free interval and survival and those with direct pelvic extension the longest. CONCLUSIONS: Retrospective comparisons of radical hysterectomy to radiation therapy are not valid unless the group of patients whose radical operation was abandoned is included. The morbidity of the operation is low even when followed by radiation therapy. However, no recommendations for optimal therapy can be made from this analysis. PMID- 11104605 TI - Comparison of clinical outcome in black and white women treated with radiotherapy for cervical carcinoma. AB - BACKGROUND: The purpose of this investigation was to evaluate the significance of race on the cancer-specific survival outcome of women treated with radiotherapy for advanced-stage cancer of the uterine cervix. METHODS: Data from 922 women with cancer of the uterine cervix treated with radiotherapy were reviewed. Patients were treated at the Mallinckrodt Institute of Radiology from 1959 through 1993. There were 576 women with clinical Stage II cancer and 346 women with clinical Stage III cancer. All women were treated following standard medical care treatment policies according to the stage of their disease. Data were analyzed by race and known treatment-related prognostic factors. Overall and cancer-specific survivals were evaluated. RESULTS: The 5-year cancer-specific survivals for clinical Stage II were 66 and 61% (P = 0.56) for white and black women, respectively. The corresponding 5-year overall survivals were 60 and 51% (P = 0.02). The 5-year cancer-specific survivals for clinical Stage III were 38 and 47% (P = 0.34) for white and black women, respectively. The associated 5-year overall survivals were 32 and 40% (P = 0.37). No differences in treatment-related factors were identified. CONCLUSIONS: In a cancer treatment system where black and white women with clinical Stage II and III cancer of the uterine cervix are all treated with radiotherapy alone, following standard treatment policies, no significant difference in cancer-specific survival outcome is shown. Multivariate analysis revealed that clinical stage and overall treatment time are significant variables affecting the control of tumor by radiotherapy. Overall survivals for the two racial groups are different and may be related to other non-cancer related factors. PMID- 11104607 TI - Tenascin-A marker for the malignant potential of preinvasive breast cancers. AB - OBJECTIVE: Up to now, the mechanisms responsible for progression from noninvasive to invasive breast cancer have remained obscure. Tenascin is an extracellular matrix glycoprotein, present in embryonal and fetal tissues, which is also found in the stroma of various benign and malignant pathologies. We studied the expression and immunohistochemical behavior of tenascin in specimens of invasive and preinvasive breast cancer in order to assess its potential role as a marker for tumor invasion. MATERIALS AND METHODS: Sixty-eight specimens including 29 noninvasive ductal cancers, 12 invasive ductal cancers, 5 adenoses, 7 fibroadenomas, and 15 samples of normal human breast tissue were evaluated. An immunofluorescent microscopic technique was used for analysis of the localization and distribution of tenascin. Paraffin-embedded biopsies were incubated with primary monoclonal anti-tenascin antibodies (1:25, Dako-tenascin, TN2). Subsequently, trimethylrhodamine-isothiocyanate-conjugated secondary antibodies (rabbit anti-mouse immunglobulins (Dakopatts, Denmark) were added to visualize the protein. RESULTS: A significant tenascin expression was observed around the ducts in all samples of patients with preinvasive breast cancers. Intensive staining was also found in the periductal stroma of all specimens of patients with invasive breast cancers. Benign breast lesions showed weaker reactivity. No tenascin expression was detectable in normal human breasts, while tissue samples of in situ cancers presented variable staining intensities positively correlating with the degree of differentiation. CONCLUSION: Tenascin immunofluorescence may prove a suitable and helpful adjunct for diagnosing malignant disease and for predicting the invasive potential of premalignant breast lesions. PMID- 11104606 TI - E-Cadherin complex protein expression and survival in ovarian carcinoma. AB - OBJECTIVE: The aim of this study was to analyze the correlation between expression of E-cadherin complex proteins, epidermal growth factor receptor (EGFR), and c-erbB-2 and disease outcome in advanced-stage ovarian carcinomas. METHODS: Sections from 75 primary ovarian carcinomas (=37) and metastatic lesions (=38) from 45 patients diagnosed with advanced-stage ovarian carcinoma (FIGO stage III-IV) were immunostained and evaluated for staining pattern, extent, and intensity. Patients were divided in two groups based on disease outcome. Long term survivors (21 patients) and short-term survivors (24 patients) were defined using a double cutoff of 36 months for disease-free survival (DFS) and 60 months for overall survival (OS). Mean follow-up period was 70 months. The mean values for DFS and OS were 109 and 125 months for long-term survivors, as compared to 3 and 21 months for short-term survivors, respectively. RESULTS: Comparison of all primary and metastatic lesions showed upregulation of gamma-catenin protein expression in the latter (P = 0.05). When segregated according to disease outcome, the expression of all studied proteins, with the exception of EGFR, was more diffuse in tumors of short-term survivors. The presence of cytoplasmic staining for c-erbB-2 was associated with poor survival in the entire cohort (P = 0.007), as well as in primary tumors alone (P = 0.003), in survival analysis. Similar results were seen in the evaluation of primary tumors for gamma-catenin (P = 0.002). CONCLUSIONS: gamma-Catenin, and possibly c-erbB-2, are valid markers of poor survival in advanced-stage ovarian carcinoma. PMID- 11104608 TI - The Society of Gynecologic Oncologists Outcomes Task Force. Study of endometrical cancer: initial experiences. AB - OBJECTIVE: The aim of this study was to develop an outcomes measure, which incorporates patient reported information, for The Society of Gynecologic Oncologists (SGO) to establish benchmarks in the treatment of endometrial cancer and demonstrate quality to third parties. METHODS: The Outcomes Task Force (OTF) developed an outcomes tool that included preoperative, intraoperative, and 120 day-postoperative assessments. Measures included demographics, patient-reported health status (SF36), comorbid conditions, living status, satisfaction surveys, operative events and disease characteristics. Patients (n = 297) were surveyed at 11 pilot sites from 10/1/97 to 9/1/99. RESULTS: The mean age of patients was 64.4 years and their mean Quetelet index was 33.2 kg/m(2). Forty-eight percent were Medicare beneficiaries and 25% were HMO patients. Mean comorbidity score was 19.1 (maximum possible 100). This represents approximately three comorbidities per average patient. Seventy-four percent were FIGO stage I, 9% stage II, 11% stage III, and 5% stage IV. Forty percent were FIGO grade 1, 35% grade 2, and 24% grade 3. Ninety-two percent of patients were able to live independently preoperatively and 91% were independent postoperatively. Seventy-seven percent of patients underwent total abdominal hysterectomy, 8% radical abdominal hysterectomy, 9% laparoscopic hysterectomy, and 1% vaginal hysterectomy. Mean length of stay was 3. 3 days and mean operative time was 119 min. Ninety-nine percent were staged and 80% underwent lymph node sampling. Two patients required unplanned returns to surgery and 8 required blood transfusion (27 units total). Postoperatively, 20% received radiation therapy and 13% received cytotoxic chemotherapy. Mean satisfaction score (scale 0 to 100) preoperatively was 86 and postoperative was 83. SF36 component summaries were preoperatively and 120 days postoperatively: physical component 43.6 vs 43.1; mental component 49.1 vs 50.6. CONCLUSION: The SGO has developed a tool for assessing outcomes for the treatment of endometrial cancer that can be made available to the membership to assess and objectively demonstrate quality of care to third parties. PMID- 11104609 TI - Survival following extended field irradiation in carcinoma of cervix metastatic to para-aortic lymph nodes. AB - OBJECTIVE: Our goal was to determine survival after extended-field treatment of para-aortic lymph node (PALN) metastasis. METHODS: Thirty-five patients were treated from 1975-1989 for PALN metastasis. The FIGO stages were IB 10, 2A 3, IIB 9, IIIA 1, IIIB 10, 4A 1, and unstaged 1. The diagnosis in 34 patients was by operative staging and in 1 by CT scan and fine-needle aspiration biopsy. Twelve patients had microscopic PALN metastasis (PALN1) and 23 had grossly enlarged lymph nodes (PALN2). Thirty-four patients had extended-field radiotherapy (RT) plus brachytherapy or pelvic boost. Kaplan-Meier estimates were computer calculated for the entire population. Late radiation morbidity was classified by RTOG/EORTC criteria. RESULTS: The 5-year overall survival rate was approximately 29%. Four patients (3 stage IB, 1 stage IIIA) survived without recurrence. All four had extended field RT. The 5-year survival rate was 41.7% for PALN1 cases and 26.1% for PALN2 cases. Three patients (8.6%) had Grade 4 morbidity. CONCLUSIONS: PALN metastasis in stage IB is curable in approximately 30% of cases. The management approach in this series in stage IB was as follows: If PALN metastasis was identified at exploration for radical hysterectomy, the procedure was aborted and extended-field RT administered. In stages IIB through IVA, operative staging or CT scanning with FNA biopsy of suspicious PALN was performed. If PALN metastasis was confirmed, extended-field RT was administered. A 35% 5-year survival rate was observed in the advanced group. The value of chemotherapy for PALN metastasis remains to be defined but results from clinical trials suggest that cisplatin-based chemotherapy may be beneficial. PMID- 11104610 TI - High external parametrial dose can increase the probability of radiation proctitis in patients with uterine cervix cancer. AB - OBJECTIVE: The aim of this study was to evaluate the relationship between external parametrial dose and radiation proctitis after external irradiation and high-dose-rate intracavitary (HDR-IC) brachytherapy among patients with cervical cancer. METHODS: From May 1993 through December 1996, 191 patients with stage IB IVA cervical cancer were managed by curative-intent radiotherapy. External irradiation to the whole pelvis (44-45 Gy/ 22-25 fractions) was delivered to all patients initially. One hundred twenty-seven patients received additional bilateral parametrial and sidewall boost (5.4-14.4 Gy/ 3-8 fractions) with 4-cm midline shielding. HDR-IC brachytherapy, 19.2-24 Gy/ 5 fractions to Point A, was given after external irradiation. Patients receiving an external dose of 44-45, 50-54, and >54 Gy were categorized as no parametrial boost (NPMB), low parametrial boost (LPMB), and high parametrial boost (HPMB) group, respectively. The actuarial proctitis rate was compared among the three groups. RESULTS: Three year overall and Grade 2-4 proctitis rates were 30 and 15%, respectively. Overall proctitis rates were 12, 34, and 51% in the NPMB, LPMB, and HPMB groups (P < 0.0001), respectively. Grade 2-4 proctitis rates were 5, 17, and 27% in the NPMB, LPMB, and HPMB groups (P = 0.0022), respectively. In multivariate analysis of overall and Grade 2-4 radiation proctitis, external parametrial dose was the only independent prognostic factor (P = 0.0002 and 0.0030, respectively). CONCLUSION: Regardless of central shielding after 44-45 Gy whole pelvis irradiation, more patients with high external parametrial dose developed radiation proctitis. Incomplete midline shielding of the upper rectum may be the cause. Diminishing the external beam doses further may decrease rectal complications. PMID- 11104611 TI - Laparoscopic assessment of the sentinel lymph node in early stage cervical cancer. AB - OBJECTIVE: The aim of this study was to describe a minimally invasive technique enabling us to identify the sentinel lymph node in patients affected by early stage cervical cancer and to report the preliminary data. METHOD: Patent Blue Violet was injected around the tumor. Laparoscopy was undertaken and the blue dyed lymph nodes (BDLN) were sought. The evidenced BDLN were removed, and then the systematic dissection was carried out. Material. Thirty-five patients were submitted to surgery. A systematic dissection was performed on 69 pelvic sidewalls (no dissection was performed on the second side of the patient for whom we decided to renounce surgery after assessment of the first side). RESULTS: One or more BDLN was evidenced in 59 of 69 dissections. The rate of failure depends on the quantity of injected blue dye. Failure to identify a BDLN depended on the quantity of injected blue dye: 3 of 6 (50%) for 1.5 ml or less, 3 of 18 (17%) for 2 ml, and only 4 of 45 (10%) after injection of 4 ml (P = 0.05). Among the 63 BDLN (in 4 cases 2 BDLN were identified), 53 were located in contact with the external iliac vein, lateral to the inferior vesical artery, and ventral to the origin of the uterine artery, 7 were located close to the origin of one of the collaterals of the internal iliac artery, and 3 were adjacent to the left common iliac vein. One or more positive pelvic lymph nodes was found in 11 pelvic wall dissections done on 8 patients. The BDLN was the positive node or one of them in all cases. CONCLUSION: If the sensitivity of the assessment of the BDLN is confirmed to be 100%, this laparoscopic approach could transform the management of early cervical cancer. PMID- 11104612 TI - The prognostic value of pretherapeutic tetranectin and CA-125 in patients with relapse of ovarian cancer. AB - OBJECTIVE: The aim of the study was to examine the prognostic values of, respectively, tetranectin (TN) and CA-125 measured in serum from patients presenting with relapse of ovarian cancer (OC). METHODS: TN and CA-125 were measured in serum samples from 75 patients with relapse of OC before the start of second-line chemotherapy. The endpoint used was death of OC. The variables were analyzed by univariate life table analysis and multivariate Cox analysis. RESULTS: A significantly shortened survival was found for patients with low serum TN values compared to patients with serum TN levels above one of the cutoff levels. The survivals are illustrated by life tables. No prognostic function was found for CA-125. TN and relapse 5 mm. Dynamic technique provided only limited additional value in the detection of microinvasive cervical carcinoma. CONCLUSION: Simple T2 MRI is useful in differentiating noninvasive or microinvasive cervical lesions from invasive cervical carcinoma of the cervix >5 mm. PMID- 11104619 TI - Intraoperative radiation therapy in the treatment of pelvic gynecologic malignancies: a review of fifteen cases. AB - OBJECTIVE: The aim of this study was to review the experience with intraoperative radiation therapy (IORT) in the treatment of gynecologic pelvic malignancies at the Massachusetts General Hospital. METHODS: From July 3, 1996, through July 28, 1999, 15 patients were treated with IORT for gynecologic malignancies in a dedicated IORT operating room suite at the Massachusetts General Hospital. Hospital medical records, radiation oncology records, and office charts were reviewed on all patients treated with IORT. IORT was given in the presence of positive surgical margins and where the doses needed for adjuvant postoperative external beam radiotherapy (EBRT) would exceed those tolerated by normal structures. One patient presented with primary disease and 14 with local or regional recurrence. Follow-up time ranged from 3 to 36 months. RESULTS: Treatment in conjunction with IORT included surgery only (7 patients); preoperative EBRT, preoperative brachytherapy, and surgery (1 patient); preoperative chemotherapy and surgery (2 patients); and surgery and postoperative chemotherapy (5 patients). IORT doses ranged from 10 to 22.5 Gy. At the completion of this review, 4 patients (26.6%) have died, 6 (40%) are alive and free of disease, and 5 (33%) are alive with disease persistence or relapse. Of the 10 patients with gross total resection, 5 are alive and free of disease. Of the 5 women with gross residual disease at the time of IORT, only 1 is alive and free of disease. CONCLUSIONS: The volume of residual disease prior to IORT may be an important prognostic indicator for disease relapse. Both local recurrence and distant metastasis were more common among patients with gross residual disease at the time of IORT. Our institutional experience with IORT further supports the importance of optimal surgical resection. PMID- 11104620 TI - Phase II trial of CI-958 in recurrent platinum-refractory ovarian carcinoma. A Gynecologic Oncology Group Study. AB - OBJECTIVES: The objectives of this study were twofold: (1) to estimate the anti tumor efficacy of CI-958 in patients with measurable platinum-resistant ovarian cancer; (2) to determine the nature and degree of toxicity of CI-958 in these patients. METHODS: Patients received CI-958 560 mg/m(2) intravenously every 3 weeks and tumor measurements were performed every one to two cycles. RESULTS: In 25 cases with recurrent platinum-resistant disease, there was one partial response (PR) and six patients had stable disease (SD). CONCLUSIONS: CI-958 has minimal activity in platinum-resistant ovarian cancer at the dose and schedule tested. PMID- 11104621 TI - Long-term follow-up of the Stockholm screening study on ovarian cancer. AB - OBJECTIVES: Seventy percent of ovarian cancer is diagnosed at advanced stages. Having a method for early diagnosis is a very attractive concept. Several attempts have been made, using monoclonal antibody-based immunoassays, ultrasound, or combinations of both, to identify methods that might prove to be sufficiently sensitive and specific as a screening test. Despite promising results, a mortality study of a large population has yet to be completed due in part to the high cost involved. METHODS: One of the first studies aimed at devising a screening strategy for ovarian cancer used the CA 125 immunoassay followed by ultrasound. The study was performed in Stockholm from 1986 through 1988. Ten years now having passed, an analysis has been performed to further evaluate the results of that study. RESULTS: Screening led to the diagnosis of ovarian cancer in six patients, five of whom have since died of the disease. By searching the Cancer Registry, we were able to identify 20 ovarian cancer patients who developed the disease after the screening period. Of these, 12 died of the disease, 2 are alive with disease, and 6 have no evidence of disease following treatment. The median survival for patients diagnosed by screening was 100 months. Median survival for ovarian cancer patients identified subsequent to screening was 20 months. Although there was no difference in survival between these two groups, median survival was better for women diagnosed by screening (borderline significance, P = 0.059). CONCLUSION: These results indicate that a study of a large number of women with a sufficiently long observation time will be required to establish whether or not screening can reduce ovarian cancer mortality. Such a study may also provide insight into the natural history of ovarian cancer. PMID- 11104622 TI - Preclinical studies of a new generation retroviral vector for ovarian cancer BRCA1 gene therapy. AB - OBJECTIVE: The aim of this study was to determine the preclinical stability, toxicity, and efficacy of a second-generation complement-resistant retroviral BRCA1 vector, MFG-BRCA1, for ovarian cancer gene therapy. METHODS: MFG-BRCA1 was packaged in human 293 renal cells and manufactured and tested under cGMP conditions and is allowed for use in humans by the Food and Drug Administration. Vector stability studies were performed in mice and human serum by PCR analysis. Toxicity in the animals was assessed at necropsy, evaluating for histological signs of inflammation and organ damage. Tissue culture efficacy studies were performed on ovarian and breast cancer cells. Animal efficacy studies were conducted in female nu/nu mice. Mice were injected intraperitoneally with SKOV-3 ovarian cancer cells and tumors were allowed to grow for 4 weeks. Mice were treated intraperitoneally with MFG-BRCA1 or control vectors. Survival of animals was compared in the MFG-BRCA1 versus the control groups. RESULTS: MFG-BRCA1 was more stable in human serum than LXSN-BRCA1sv. Toxicity as demonstrated by an inflammatory peritonitis was minimal. Significantly fewer clones were obtained using the MFG-BRCA1 versus the MFG vector alone in both cell lines. Efficacy studies in animals of MFG-BRCA1 demonstrated a near threefold increase in survival over control vector and twofold increase compared to the first generation LXSN-BRCA1sv vector. CONCLUSION: The reengineered complement-resistant MFG-BRCA1 retroviral vector is more effective and more stable than the previous generation LXSN-BRCA1sv vector. PMID- 11104623 TI - Is uterine papillary serous adenocarcinoma a manifestation of the hereditary breast-ovarian cancer syndrome? AB - BACKGROUND: Uterine papillary serous carcinoma (UPSC) shares common pathologic, genetic, and clinical features with other serous cancers of mullerian origin. The most common histologic type of ovarian tumor associated with BRCA mutations is papillary serous. Because of these histologic similarities, we postulated that, in some cases, UPSC may be a manifestation of a field defect in BRCA1 carriers, which also includes ovarian carcinoma, fallopian tube carcinoma, and primary peritoneal carcinoma. METHODS: Fifty-six living patients with UPSC were contacted through their treating physicians and agreed to a family history interview and to provide a blood specimen for BRCA testing. The protein truncation test was used to detect mutations in exons 10 and 11 of BRCA1 and in exon 11 of BRCA2. The presence of four common mutations was assessed by PCR-based specific assays. RESULTS: A high proportion of patients had a past history of breast cancer (11%) or a first-degree relative with breast cancer (29%). Four patients were from families with site-specific hereditary breast cancer. However, there was no clear example of the hereditary breast-ovarian cancer syndrome, and none of the 56 patients was found to carry a BRCA1 or BRCA2 mutation. CONCLUSIONS: BRCA mutations do not appear to predispose to UPSC and this type of cancer does not appear to be a manifestation of the classical hereditary breast-ovarian cancer syndrome. The observed association between UPSC and breast cancer may be due to the presence of mutations in other cancer predisposing genes. PMID- 11104624 TI - Tubal sterilization and risk of cancer of the endometrium. AB - OBJECTIVE: Surgical sterilization is a common method of contraception among U.S. women. Most surgical sterilizations are tubal ligations, but few studies have investigated their potential impact on endometrial cancer risk. METHODS: A case control study included 405 women diagnosed with endometrial cancer at 5 U.S. medical centers between 1987 and 1990 and 297 age-, race-, and location-matched controls who were identified by random-digit-dialing. Questionnaires ascertained information on tubal sterilization, and logistic regression models generated odds ratios (ORs) to estimate relative risk. RESULTS: The OR and 95% confidence interval for tubal sterilization, which was reported by 47 cases and 40 controls, was 0.9 (0.6-1.4) before adjustment and 1. 4 (0.8-2.4) after adjustment for age, parity, and oral contraceptive use. Age at surgery, years since surgery, or calendar years of surgery were not associated with endometrial cancer, and associations did not vary according to parity or stage of disease at diagnosis. CONCLUSIONS: Tubal sterilization is not substantially associated with endometrial cancer. PMID- 11104625 TI - Effect of MRI on therapeutic decisions in invasive cervical carcinoma. Direct comparison with the pelvic examination as a preperative test. AB - OBJECTIVES: Our aim was to compare magnetic resonance imaging (MRI) with the current standard clinical practice (pelvic examination including general anesthesia in selected patients) with regard to treatment planning in invasive cervical carcinoma. It was of particular interest to compare the accuracy of both methods for allocating the patients to the appropriate treatment modality: surgery versus primary radiotherapy. METHODS: One hundred and three consecutive patients with primary invasive cervical carcinoma underwent both MRI at 1.5 T and pelvic examination. The gold standard for comparing treatment decisions was based on the surgico-pathologic data: tumor confined to the cervix (treatment decision for surgery) or extracervical tumor spread (treatment decision for primary radiotherapy). RESULTS: A gold standard was available in 91 patients. The pelvic examination made correct treatment decisions in 89% of patients. However, the sensitivity for extracervical spread was only 44% (8/18 patients). MRI was better at identifying extracervical tumor spread: 67 and 89% for observers 1 and 2, respectively. MRI, however, had more false positive results and correct treatment decisions were made in 69-84% of patients (observer 1, 76/91; observer 2, 63/91). CONCLUSION: Treatment decisions based on the pelvic examination were correct in 89%, with MRI not bringing improvement. MRI, however, is better in diagnosing extracervical spread, but at the cost of more false positives. PMID- 11104626 TI - Local recurrence in high-risk node-negative stage I endometrial carcinoma treated with postoperative vaginal vault brachytherapy. AB - OBJECTIVES: The aim of this study is to examine the patterns of failure after extended surgical staging and postoperative vaginal vault brachytherapy as the only adjuvant treatment in high-risk surgical Stage I patients with endometrial carcinoma. METHODS: The records of all patients with endometrial carcinoma (adenocarcinoma or adenosquamous) receiving vaginal vault brachytherapy as the only adjuvant treatment from January 1989 to December 1997 were examined. A total of 489 patients were found. Of these, 133 had extended surgical staging. The study group consists of 77 surgical Stage I patients with Substages IBG3 and any grade IC. Recurrences were recorded as in the vagina, pelvis, or distant. RESULTS: The mean follow-up interval was 45 months (range 14 to 96 months). Eleven patients had recurrence (14%). Median time to recurrence was 15 months (range 6 to 56 months). Recurrences occurred in the vagina in 7, pelvis in 1, and distantly in 3 patients. Five of 7 vaginal recurrences occurred within 2 years. All patients with distant recurrence died from disease. One patient with pelvic recurrence is alive with disease. Only 1 patient with vaginal recurrence died from disease. Six patients with isolated recurrences in the vagina were successfully treated with radiotherapy with or without local excision. All 6 have no evidence of disease at follow-up (median survival 29 months, range 20 to 71 months). CONCLUSIONS: The vagina remains the most common site of recurrence for high-risk surgical Stage I patients treated with postoperative vaginal vault brachytherapy. Close follow-up in the first 2 years is essential to detect isolated vaginal recurrences. These are amenable to salvage treatment with good disease-free survival. PMID- 11104628 TI - Coexistent choriocarcinoma and malignant mixed mesodermal tumor of the uterus. AB - OBJECTIVE: The purpose of this article is to report a case of coexisting uterine choriocarcinoma and uterine malignant mixed mesodermal tumor (MMMT). The relevant literature is reviewed and possible pathogenesis discussed. METHODS: The clinical course and histopathology of the case were reviewed and a Medline literature search for other cases was performed. RESULTS: The patient's uterine tumor contained syncytiotrophoblastic and cytotrophoblastic cells that stained positively for the beta subunit of human chorionic gonadotrophin consistent with uterine choriocarcinoma. Pathology also revealed a second distinct neoplasm composed of adenocarcinoma admixed with sarcoma, compatible with a uterine MMMT. The patient experienced metastatic choriocarcinoma to her lungs, lymph nodes, and brain. She suffered a complicated clinical course and died 7 months after her initial diagnosis. The literature search revealed that various gynecologic and nongynecologic carcinomas with trophoblastic differentiation have been described, but an association with uterine MMMT has not been previously reported. CONCLUSIONS: Trophoblastic differentiation and choriocarcinoma associated with gynecologic and nongynecologic tumors is rare. We document the presence of uterine MMMT coexisting with uterine choriocarcinoma that followed an aggressive clinical course and review the possible pathogenesis of this lesion. PMID- 11104627 TI - A phase I trial of a 3-day topotecan Q 21 days for recurrent epithelial cancers of the ovary, fallopian tube, and peritoneum. AB - OBJECTIVE: This trial was undertaken to determine the dose limiting toxicity (DLT) and maximum tolerated dose (MTD) of topotecan that can be administered for 3 days q 21 days. A 3-day schedule is more convenient and less expensive than standard 5-day dosing. METHODS: Patients with recurrent epithelial ovary, tubal, or peritoneal carcinoma were treated with escalating doses of topotecan beginning at 2.50 mg/m(2) as an outpatient days 1-3 q 21 days. Colony stimulating factors were not employed prophylactically, but could be added for grade 4 marrow toxicity. RESULTS: Twenty patients with a median age of 61 (range 46-80) and performance status of 0 or 1 were entered. All patients had received at least one prior paclitaxel/platinum regimen; 6 had received two. Ninety-one cycles were delivered (median = 6) and 98.9% were on schedule. Grade 4 neutropenia was seen in 17 of 20 patients (85%) in cycle 1 and in 38 of 91 (41.8%) total cycles. Sixteen of 20 patients (80%) started G-CSF on cycle 2. Two of 91 (2.2%) cycles had grade 4 thrombocytopenia. Four cycles (4.4%) were associated with febrile neutropenia. Two patients experienced grade 4 neurotoxicity (DLT) at 4.25 mg/m(2). Other nonhematologic toxicity was mild. CONCLUSIONS: Topotecan can be safely administered on schedule as an outpatient days 1-3 q 21 days. Neurotoxicity was the DLT when G-CSF was added; the MTD was 3.75 mg/m(2). There was minimal other nonhematologic toxicity. Neutropenia was predictable and easily managed with G-CSF. Febrile neutropenia was uncommon and thrombocytopenia was rare at the doses evaluated. PMID- 11104629 TI - Immunohistochemical detection of parathyroid hormone-related protein in a squamous cell carcinoma arising from mature cystic teratoma causing humoral hypercalcemia of malignancy. AB - BACKGROUND: Humoral hypercalcemia of malignancy is a cancer-related hypercalcemia caused by the production of humoral factors by malignant cells in patients without bone metastases. Squamous cell carcinomas are the tumors most frequently associated with humoral hypercalcemia of malignancy, and parathyroid hormone related protein (PTH-rP) is the main humoral factor implicated. Squamous cell carcinoma arising from mature cystic teratoma is a rare diagnosis itself, much less the description of associated hypercalcemia, despite the fact that the normal keratinocytes produce parathyroid hormone-related protein. CASE: We present a well-documented case of squamous cell carcinoma arising from mature cystic teratoma of the ovary, complicated by hypercalcemia in a patient with high levels of plasma parathyroid hormone-related protein and immunohistochemical evidence of parathyroid hormone-related protein expression by the tumor cells. CONCLUSION: In this case, the carcinoma cells had already produced PTH-rP in the primary tumor although the serum calcium levels had not been significantly high at surgery. It is therefore suggested that hypercalcemia may have occurred after PTH-rP production had overcome the homeostatic level during the terminal stage. PMID- 11104630 TI - Malignant struma ovarii with Graves' disease. AB - BACKGROUND: Malignant struma ovarii is a rare tumor, consisting of a struma ovarii with malignant change. Because of the rarity of the tumor, only a few reports with detailed data of thyroid function of the patient have been published. CASE: Medical and surgical treatments were performed in a case of malignant struma ovarii with Graves' disease and data from thyroid function tests were analyzed. After removal of the tumor, rapid improvement of thyroid function was observed and thyroglobulin level decreased. CONCLUSION: This case demonstrates the possible, if not dominant, contribution from the malignant struma ovarii to the hyperthyroidism of the patient. PMID- 11104631 TI - Female urethral adenocarcinoma arising from urethritis glandularis. AB - INTRODUCTION: Female urethral adenocarcinoma is extremely rare and more than one tissue of origin has been suggested other than the Skene's gland. Immunohistochemistry with cytokeratins (CK) 7 and 20 is used to define the origin of the tumor. CASE REPORT: A 72-year-old woman presented with a 2-cm polypoid tumor at the external urethral meatus and bleeding. Wide local excision and bilateral inguinal lymphadenectomy were performed. Postoperative convalescence was uneventful. It was a poorly differentiated mucinous adenocarcinoma without direct urothelial involvement. There were focal areas of intestinal metaplasia. The tumor cells were positive for CK 7 and 20 and negative for prostate-specific antigen. DISCUSSION: This case provides supportive evidence that mucinous urethral adenocarcinoma may arise from malignant transformation of urethritis glandularis. PMID- 11104632 TI - A case of rapidly growing ovarian squamous cell carcinoma successfully controlled by weekly paclitaxel-carboplatin administration. AB - BACKGROUND: Primary squamous cell carcinoma of the ovary is uncommon and has a poor prognosis. Because of its rarity, the effective postoperative treatment is unknown. We describe a remarkable response of this tumor to weekly paclitaxel carboplatin administration. CASE: A 53-year-old woman had rapidly growing primary squamous cell carcinoma of the ovary that metastasized to the abdominal wall and transverse colon after maximum cytoreductive surgery. The tumor was resistant to primary chemotherapy with cisplatin, vincristine, mitomycin C, and bleomycin. A combination of paclitaxel and carboplatin was used for second-line chemotherapy and was repeated every week. The patient tolerated the chemotherapy well and demonstrated a pathological complete response in the abdominal metastases following the five courses of chemotherapy. CONCLUSION: Weekly paclitaxel carboplatin administration may be a safe and effective treatment for advanced and rapidly growing ovarian squamous cell carcinoma with primary resistance to chemotherapy. PMID- 11104633 TI - Pregnancy in a woman with a Y chromosome after removal of an ovarian dysgerminoma. AB - BACKGROUND: It appears to be a general belief that pregnancy might be impossible in women with the XY karyotype. Therefore, it is recommended that patients with dysgerminoma of the ovary associated with the XY karyotype should undergo a bilateral salpingo-oophorectomy. CASE: We report an extremely rare case of a true hermaphrodite with a 20% 46,XX/80% 46,XY karyotype who became pregnant after removal of an ovarian dysgerminoma. The patient had a completely normal female phenotype. A dysgerminoma with ovotestis was found in the right ovary. Two courses of chemotherapy following a right salpingo-oophorectomy were carried out. Nine months later she became pregnant and delivered a healthy male infant. CONCLUSION: A unilateral salpingo-oophorectomy followed by combination chemotherapy can be the treatment of choice for any woman who wishes to preserve her capacity for conception at the time of operation for dysgerminoma of the ovary. PMID- 11104634 TI - The clinical usefulness of preoperative intratumoral Doppler analysis in predicting lymph node metastasis in patients with endometrial carcinoma. PMID- 11104635 TI - Activity of docetaxel in chemoresistant gestational choriocarcinoma. PMID- 11104636 TI - Comment on: "Complete hydatidiform mole and a coexistent viable fetus". PMID- 11104637 TI - Reply PMID- 11104638 TI - Social signals influence hormones independently of calling behavior in the treefrog (Hyla cinerea). AB - Social signals play an important role in regulating hormone-behavior relationships. In anurans (frogs and toads), acoustic signals are an essential aspect of reproductive behavior; however, the physiological consequences of receiving social signals has remained largely undescribed. Each night for 5, 10, or 20 days, we presented acoustically isolated male treefrogs with a conspecific mating chorus, an array of tones, or no sound. We recorded calling rate of individuals throughout the experiment and collected blood before and after treatment. Days of stimulus exposure had no effect on any dependent measure. Acoustic treatment influenced steroid levels; testosterone, dihydrotestosterone, and corticosterone increased only in the group exposed to the chorus. Chorus exposed males also showed an increase in stimulus-evoked calling. We found no correlation between androgens and calling within each treatment group. In addition, noncallers in the chorus group had higher levels of androgens than males in the tone or no sound groups. Further, chorus-exposed males with zero, low, or high rate of calling had similar levels of androgens. These data indicate that social signals increase circulating androgens independently of calling behavior. Elevated corticosterone associated with chorus reception did not inhibit calling behavior, and corticosterone showed no correlation with androgen levels. PMID- 11104639 TI - Adult plasticity in hormone-sensitive motoneuron morphology: methodological/behavioral confounds. AB - Changes in androgen levels can alter the structure of motoneurons in the spinal nucleus of the bulbocavernosus (SNB), a motor nucleus that innervates perineal muscles involved in copulatory behavior. While sexual activity can alter androgen levels in normal males, it has no effect on SNB motoneuron soma size or dendritic morphology (Beversdorf, Kurz, and Sengelaub, 1990). However, Breedlove (1997) reported reductions in the size of SNB somata, nuclei, and target muscles of copulating versus noncopulating castrated rats maintained on subphysiological testosterone. To reconcile the results obtained using intact versus implant paradigms, we tested the hypothesis that the implant/behavior paradigm could produce differences in hormone levels, potentially confounding sexual behavior effects on the morphology of this androgen-sensitive neuromuscular system. Young adult male rats were castrated and immediately given 5-mm Silastic implants containing crystalline testosterone. One week later, blood samples were drawn and the males were housed with receptive females (copulators) or nonreceptive females (noncopulators) or housed alone (singles). After 27 days, blood samples were drawn again, and SNB target muscles and spinal cords removed. No differences in target muscle weight or SNB somata and nuclei size were observed between copulators, noncopulators, or singles; as expected, all measures were significantly reduced relative to intact males. Radioimmunoassay showed that testosterone declined differentially over the course of the behavioral manipulation across groups, being greatest in copulators and least pronounced in single males. These data indicate that differences in sexual or housing experience can alter testosterone titers under these implant conditions, potentially confounding hormone-sensitive measures of morphology. PMID- 11104640 TI - Testosterone, endurance, and Darwinian fitness: natural and sexual selection on the physiological bases of alternative male behaviors in side-blotched lizards. AB - The mechanistic bases of natural and sexual selection on physiological and behavioral traits were examined in male morphs of three colors of the side blotched lizard, Uta stansburiana. Orange-throated males are aggressive and defend large territories with many females. Blue-throated males defend smaller territories with fewer females; however, blue-throated males assiduously mate guard females on their territory. Yellow-throated males do not defend a territory, but patrol a large home range. They obtain secretive copulations from females on the territories of dominant males. Males with bright orange throats had higher levels of plasma testosterone (T), endurance, activity, and home range size and concomitantly gained greater control over female home ranges than blue- or yellow-throated males. Experimentally elevating plasma T in yellow- and blue throated males increased their endurance, activity, home range size, and control over female territories to levels that were seen in unmanipulated orange-throated males that had naturally high plasma T. However, the enhanced performance of orange-throated males is not without costs. Orange-throated males had low survival compared to the other morphs. Finally, some yellow-throated males transformed to a partial blue morphology late in the season and the endurance of these transforming yellow-throated males increased from early to late in the season. In addition, yellow-throated males that transformed to blue also had significantly higher plasma T late in the season compared to the plasma T earlier in the season. T appears to play an important role in the physiological changes that all three color morphs undergo during the process of maturation. In some yellow males, T plays an additional role in plastic changes in behavior and physiology late in the reproductive season. We discuss natural and sexual selection on physiological and behavioral traits that leads to the evolution of steroid regulation in the context of alternative male strategies. PMID- 11104641 TI - Acute administration of estrogen and progesterone impairs the acquisition of the spatial morris water maze in ovariectomized rats. AB - Although several markers of synaptic efficacy are enhanced during proestrus, spatial water maze performance is impaired. Because levels of both estrogen and progesterone are elevated in proestrus, the nature of their individual and combined effects on spatial learning was examined. Long-Evans hooded rats were ovariectomized postpubertally and pretrained on a water maze with a visible platform (nonspatial). Following pretraining, rats were administered estrogen (5 microg sc) or oil 48 and 24 h prior to testing and progesterone (500 microg sc) or oil 4 h prior to testing. Rats were tested on a water maze in a different room with a submerged platform (spatial) for 16 trials with random start location in a single testing day. Latency and path length to the target platform were significantly greater in estrogen plus progesterone-treated animals than in controls. Neither estrogen nor progesterone alone significantly impaired performance relative to controls on either measure. Swim speed was not significantly affected by any of the hormone treatments. Performance on a nonspatial cue task was not significantly altered by ovarian steroids. Thus, the combination of estrogen and progesterone produces deficits in the acquisition of the Morris water maze that may be specific to spatial reference memory. These deficits could be due to hormonal influences on extrahippocampal structures or to detrimental effects on behavior resulting from the increased synaptic activity intrinsic to the hippocampus proper. PMID- 11104642 TI - Testicular hormones modulate circadian rhythms of the diurnal rodent, Octodon degus. AB - Sex differences have been identified in a variety of circadian rhythms, including free-running rhythms, light-induced phase shifts, sleep patterns, hormonal fluctuations, and rates of reentrainment. In the precocial, diurnal rodent Octodon degus, sex differences have been found in length of free-running rhythm (tau), phase response curves, rates of reentrainment, and in the use of social cues to facilitate reentrainment. Although gonadal hormones primarily organize circadian rhythms during early development, adult gonadal hormones have activational properties on various aspects of circadian rhythms in a number of species examined. Gonadectomy of adult female O. degus did not influence tau, phase angle of entrainment, or activity patterns in previous experiments. The present experiment examined the role of gonadal hormones in adult male degus' circadian wheel-running rhythms. We predicted that male gonadal hormones would have an activational effect on some aspects of circadian rhythms, particularly those in which we see sex differences. Phase angles of entrainment, tau, length of the active period (alpha), maximum and mean activity levels, and activity amplitude were examined for intact and castrated males housed in LD 12:12. Responses to light pulses while housed in constant darkness (DD) were also compared. Castration had no significant effect on tau or light-induced phase shifts. However, castration significantly increased phase angle of entrainment and decreased activity levels. The data indicate that adult gonadal steroids are not responsible for the sex differences in endogenous circadian mechanisms of O. degus (tau, PRC), although they influence activity level and phase angle of entrainment. This is most likely due to masking properties of testosterone, similar to the activity-increasing effects of estrogen during estrus in O. degus females. PMID- 11104644 TI - A beneficial effect of estrogen on working memory in postmenopausal women taking hormone replacement therapy. AB - Recent neurophysiological data suggest that the prefrontal cortex (PFC) may be susceptible to modulation by estrogen. In humans, the PFC mediates a number of cognitive processes that contribute to memory function, particularly working memory. The present study examined whether memory tasks that recruit PFC dependent information processing might exhibit estrogen sensitivity in women. Performance on several memory tasks, including measures of working memory, was evaluated in three groups of postmenopausal women: (1) women who were tested when taking estrogen only (n = 38, M(age) = 55.1 years), (2) women who were tested when taking estrogen and a progestin concurrently (n = 23, M(age) = 55.9 years), and (3) women who were not taking hormone replacement therapy (n = 35, M(age) = 56.0 years). Estrogen users exhibited significantly better performance on a verbal task and on a spatial task, each with a prominent working memory component, but did not differ from nonusers on control tasks involving simple passive recall. These findings are consistent with the hypothesis that estrogen is active within PFC and is capable of influencing functions dependent on this region. The results of this study raise the possibility that estrogen may play a role in maintaining certain frontal lobe functions in women. PMID- 11104645 TI - Insulin tolerance during endotoxic shock in 10-day-old rats. AB - PURPOSE: The purpose was to investigate insulin tolerance during endotoxic shock in 10-day-old rats. MATERIALS AND METHODS: [(14)C]Deoxy-glucose (2DG) with or without insulin (1 unit/kg) was injected to 10-day-old and 6-week-old rats 3 h after an injection of endotoxin (lipopolysaccharide: LPS). Plasma concentrations of glucose and 2DG were serially measured for 45 min. Gluconeogenesis was measured in hepatocytes isolated from control and endotoxic 10-day-old rats to evaluate effects of insulin on gluconeogenesis. RESULTS: In endotoxic 10-day-old rats, plasma glucose concentration at 45 min was 48 +/- 3% (P < 0.05) of value at 0 min, and when insulin was injected with 2DG, it was 29 +/- 4% (P < 0.05) after insulin injection. Plasma 2DG disappearance was enhanced by insulin injection in the control (t(1/2) = 17.9 vs 20.5 min, P < 0.05), but not in the endotoxic rats (t(1/2) = 17.9 vs 18.4 min), indicating the presence of insulin tolerance in septic rats. Insulin decreased gluconeogenesis (P < 0.05) in hepatocytes from both control and endotoxic 10-day-old rats. In endotoxic 6-week-old rats, plasma glucose concentration was decreased to 46 +/- 10% at 45 min and further decreased to 38 +/- 4% (P < 0.05) by insulin injection. Plasma 2DG disappearance was enhanced by insulin injection in the control (t(1/2) = 11.8 vs 17.4 min, P < 0.05) and in the septic rats (t(1/2) = 14.8 vs 12.2 min). However, the enhancement of plasma 2DG disappearance by insulin was less (P < 0.05) in the septic rats than in the control, confirming reports of other investigators which showed insulin tolerance in septic shock. CONCLUSION: Although hepatocytes from endotoxic rats retained insulin sensitivity, insulin tolerance which was evaluated by 2DG disappearance occurred during septic shock in newborn rats. PMID- 11104643 TI - Seasonal changes in courtship song and the medial preoptic area in male European starlings (Sturnus vulgaris). AB - In male starlings (Sturnus vulgaris) courtship song plays a critical role in mate attraction. During the breeding season courtship song occurs prior to copulation and appears to reflect male sexual arousal. Outside the breeding season starlings sing, but song appears unrelated to reproduction. The aromatization of testosterone (T), likely within the medial preoptic nucleus (POM), is critical for the expression of male sexual arousal. The present study was performed to determine whether seasonal changes in the POM might relate to seasonal changes in courtship singing behavior in male starlings. T concentrations, the volume of the POM, and aromatase within the POM were examined both during and outside of the breeding season in male starlings. Song was also recorded at these times both with and without a female present. The POM was largest and contained dense aromatase immunostaining only during the spring breeding season, when T concentrations were highest and males responded to a female with an increase in courtship song. Outside the breeding season the volume of the POM was small, T concentrations were low, and males displayed no changes in song expression in response to female conspecifics. Song bout length was positively related to POM volume, and males sang longer songs in spring. Only males with nestboxes in spring responded to a female, and the POM tended to be larger in these males, suggesting that nestbox possession might influence neuroplasticity within the POM. Overall, the findings suggest that T-dependent plasticity and aromatase activity within the POM might regulate courtship singing in a wild songbird. PMID- 11104646 TI - Factors influencing mortality in the rat elastase-induced-aneurysm model. AB - BACKGROUND: Intraarterial infusion of elastase has been used to create an experimental model of aortic aneurysm in rats. Unfortunately, the utility of this model is limited by a high mortality rate among experimental animals. This study examined the factors influencing mortality to help refine this model. MATERIALS AND METHODS: A total of 126 Wistar rats were divided into six groups. A 1.0-cm segment of infra-renal abdominal aorta was clamped (n = 21), clamped and cannulated via the femoral artery (n = 21), infused with saline (n = 21), or infused with solution containing 25 U of elastase from three different lots (each group, n = 21). Clamping or infusion was performed for 30 (n = 7), 60 (n = 7), and 120 min (n = 7). The mortality rates were calculated at 7 days. RESULTS: There were no deaths in the clamp group, but 29% of the rats that underwent cannulation with clamping or saline infusion for 2 h died. The mortality rate for a 30-min elastase infusion ranged from 0 to 71%. Mortality for a 60-min infusion ranged from 14 to 100%. Mortality for a 2-h infusion ranged from 43 to 100%. The mortality rate was dependent on the treatment time and the elastase lot number. CONCLUSIONS: Aortic cannulation, elastase infusion, and prolonged infusion times all increase the mortality rate in the elastase-induced rat aortic aneurysm model. Mortality is also dependent on the elastase lot number. PMID- 11104647 TI - CD34, CD117, and actin expression in phyllodes tumor of the breast. AB - BACKGROUND: This study investigated the immunophenotypic patterns of CD34, CD117 (a product of the c-kit proto-oncogene), and actin (HHF35) in benign and malignant phyllodes tumors (PTs). We correlated the expression of CD34, CD117, and actin with histopathological grade. MATERIALS AND RESULTS: We analyzed 19 cases (7 benign and 12 malignant cases) of PTs using immunohistochemical analysis. Six of 7 benign PT stromal lesions stained positively for CD34, while only 3 of 12 cases of malignant PT were focally CD34 positive (P = 0.0106). Only 1 of the 7 benign PTs stromal lesions expressed CD117. Nine of the malignant PTs were composed CD117-positive fibroblasts. This result demonstrated that CD117 expression is associated with the malignant potential of PTs (P = 0. 0106). Actin (HHF-35) expression was found in 8 of 12 cases of malignant PTs (P = 0.027), but in only 1 of 7 cases of benign PTs. Actin expression was significantly (P = 0.04) correlated to frequent mitotic activity (>5 mitoses per 10 high-power fields). The immunophenotypic markers were not related to tumor size. Additionally, we sequenced part of the juxtamembrane region of the c-kit proto-oncogene and found point mutations in two malignant PTs. CONCLUSION: Our results demonstrated that expression of CD34 was associated with benign PTs, while CD117 and actin were preferentially expressed in malignant PTs. Our results implied that these immunohistological markers might be used for the histopathological grading of PTs. PMID- 11104648 TI - Evaluation of sepramesh biosurgical composite in a rabbit hernia repair model. AB - BACKGROUND: In cases such as incisional hernia repair, polypropylene mesh (PPM) can be exposed to the underlying viscera and cause adhesions to the mesh. In this study, a composite prosthesis that was designed to be less susceptible to adhesion formation than PPM was evaluated in a rabbit incisional hernia repair model. MATERIALS AND METHODS: A 5 x 7-cm full-thickness defect was created in the abdominal wall of 30 female New Zealand White rabbits. Ten animals each were repaired with PPM, Bard Composix (PP/ePTFE), or Sepramesh biosurgical composite-a polypropylene mesh coated on one side with chemically modified sodium hyaluronate and carboxymethylcellulose (HA/CMC). The animals were sacrificed after 28 days and the overall performance, including adhesion formation and tissue integration by histology and mechanical testing, was evaluated. RESULTS: In the Sepramesh group, there was a significant reduction in the percentage of surface area covered by adhesions and a significant increase in the percentage of animals with no adhesions compared to standard materials. The tissue integration strength and overall cellular response were similar in all groups. A partially remesothelialized peritoneal surface was often apparent overlying the Sepramesh implant. CONCLUSIONS: Sepramesh biosurgical composite effectively repaired abdominal wall defects in rabbits and reduced adhesion development to the mesh compared to the use of a PPM and a PP/ePTFE composite. PMID- 11104649 TI - Quantitative study of changes in intestinal morphology and mucus gel on total parenteral nutrition in rats. AB - Quantification of changes in gastrointestinal morphology and mucus gel has been difficult to study. In the present study, we investigated changes in rat intestine under total parenteral nutrition (TPN) using fluoresceinated lectin staining and image analysis. Wistar rats (n = 34) were divided into two groups: one group received TPN for 2 weeks, and a control group received standard rat chow and water ad libitum for the same period. A 1-cm segment of distal ileum was removed and cut into cross sections. Sections were stained with hematoxylin and eosin, and to stain the mucus, periodic acid-Schiff (PAS), alcian blue (AB), and fluoresceinated lectin, that is, FITC-labeled Ulex europaeus agglutinin I (FITC UEA-I), were used. Light microscope images were stored in a personal computer and analyzed using image analysis. We measured perimeter length, mucosal thickness, villus area, villus surface area index, mucus stain-positive area, mucosal area ratio, and mucosal surface area ratio. Perimeter length, mucosal thickness, villus area, and villus surface area index in the TPN group were significantly less than those in the control group (P < 0.001 for each parameter). In all mucus stainings, the stain-positive area in the TPN group was significantly less than that in the control group. However, there were no significant differences in mucosal area or mucosal surface area ratios between the two groups. The FITC-UEA I-positive area was significantly greater than the PAS- or and AB-positive area. There were significant positive correlations between the FITC-UEA-I-positive area and both the PAS-positive and AB-positive areas. TPN for 2 weeks promoted intestinal atrophy and decreased absolute quantity of mucus gel. We successfully introduced the FITC-UEA-I staining method to evaluate changes in mucus gel. PMID- 11104651 TI - Opening of potassium channels protects mitochondrial function from calcium overload. AB - Ischemic preconditioning (IPC) protects myocardium from ischemia reperfusion injury by activating mitochondrial K(ATP) channels. However, the mechanism underlying the protective effect of K(ATP) channel activation has not been elucidated. It has been suggested that activation of mitochondrial K(ATP) channels may prevent mitochondrial dysfunction associated with Ca(2+) overload during reperfusion. The purpose of this experiment was to study, in an isolated mitochondrial preparation, the effects of mitochondrial K(ATP) channel opening on mitochondrial function and to determine whether it protects mitochondria form Ca(2+) overload. Mitochondria (mito) were isolated from rat hearts by differential centrifugation (n = 5/group). Mito respiratory function was measured by polarography without (CONTROL) or with a potassium channel opener (PINACIDIL, 100 microM). Different Ca(2+) concentrations (0 to 5 x 10(-7) M) were used to simulate the effect of Ca(2+) overload; state 2, mito oxygen consumption with substrate only; state 3, oxygen consumption stimulated by ADP; state 4, oxygen consumption after cessation of ADP phosphorylation; respiratory control index (RCI: ratio of state 3 to state 4); rate of oxidative phosphorylation (ADP/Deltat); and ADP:O ratio were measured. PINACIDIL increased state 2 respiration and decreased RCI compared to CONTROL. Low Ca(2+) concentrations stimulated state 2 and state 4 respiration and decreased RCI and ADP:O ratios. High Ca(2+) concentrations increased state 2 and state 4 respiration and further decreased RCI, state 3, and ADP/Deltat. PINACIDIL improved state 3, ADP/Deltat, and RCI at high Ca(2+) concentrations compared to CONTROL. Pinacidil depolarized inner mitochondrial membrane, as evidenced by decreased RCI and increased state 2 at baseline. Depolarization may decrease Ca(2+) influx into mito, protecting mito from Ca(2+) overload, as evidenced by improved state 3 and RCI at high Ca(2+) concentrations. The myocardial protective effects resulting from activating K(ATP) channels either pharmacologically or by IPC may be the result of protecting mito from Ca(2+) overload. PMID- 11104650 TI - Urinary trypsin inhibitor reduces C-X-C chemokine production in rat liver ischemia/reperfusion. AB - BACKGROUND AND AIM: Protease inhibitors attenuate ischemia/reperfusion injury. However, the underlying mechanisms by which protease inhibitors prevent reperfusion injury remain obscure. Neutrophils play an important role in reperfusion injury. We studied the effects of urinary trypsin inhibitor (UTI) on production of the C-X-C chemokine, cytokine-induced neutrophil chemoattractant (CINC), by Kupffer cells during ischemia/reperfusion of the liver. METHODS: Liver ischemia was induced in rats by occlusion of the portal vein for 30 min. UTI (50,000 U/kg) was injected intravenously 5 min before vascular clamping. Serum CINC concentrations were measured by enzyme-linked immunosorbent assay. Levels of CINC mRNA in the liver were determined by Northern blot analysis. We also examined the inhibitory effects of UTI on in vitro CINC production by peritoneal macrophages in response to neutrophil elastase (NE). RESULTS: Serum CINC concentrations increased and peaked 6 h after reperfusion. However, pretreatment of animals with UTI blunted this increase in CINC and significantly reduced CINC mRNA levels in the liver after ischemia/reperfusion. UTI also decreased neutrophil accumulation in the liver 24 h after reperfusion. In vitro CINC production by Kupffer cells from rats pretreated with UTI 3 h after ischemia/reperfusion was significantly decreased compared to those from untreated animals. UTI reduced NE activity in vitro in a dose-dependent manner, and UTI significantly reduced in vitro CINC production by peritoneal macrophages stimulated with NE. CONCLUSION: UTI reduces the production of CINC by Kupffer cells stimulated with NE, attenuating ischemia/reperfusion injury of the liver. PMID- 11104652 TI - Effects of active and negative mutants of Ras on rat arterial neointima formation. AB - BACKGROUND: Ras protein is a key signal transducer in the cause of cell proliferation. We studied the effects of active and negative mutants of the Ras gene on arterial neointimal formation in rats, with the aim of elucidating the molecular mechanisms regulating restenosis following percutaneous transluminal coronary angioplasty. MATERIALS AND METHODS: AdRasV12 and AdRasN17, the recombinant adenoviruses containing a constitutively active mutant and a dominant negative mutant of Ras, respectively, were used to determine whether Ras is necessary and sufficient to modulate the smooth muscle cell proliferation and neointima formation. Following balloon injury, rat common carotid arteries were treated in their distal half with AdRasV12, AdRasN17, or AdLacZ, with the proximal half used as uninfected control. RESULTS: In rat arteries subjected to balloon injury, either uninfected or treated with AdLacZ, there were pronounced SMC proliferation and neointima formation. These changes were markedly augmented by AdRasV12 and reduced by AdRasN17. CONCLUSION: Ras is necessary and sufficient for SMC proliferation and neointima formation and may play a critical role in restenosis following balloon angioplasty. PMID- 11104653 TI - Dynamic progression of contractile and endothelial dysfunction and infarct extension in the late phase of reperfusion. AB - BACKGROUND: Myocardial injury during early reperfusion (R) has been well documented. However, the extent and time course of myocardial injury during late R are still unclear. The purpose of this study was to determine the extent of regional contractile and endothelial dysfunction and myocardial blood flow (MBF) defect as well as extension of infarction in association with neutrophil (PMN) actions during R. MATERIALS AND METHODS: A total of 29 dogs underwent a protocol of 1 h LAD ischemia followed by 6, 24, 48, and 72 h of R, respectively. Regional contractile function (sonomicrometry), MBF (colored microspheres), infarct size (triphenyltetrazolium chloride staining), and PMN localization (immunohistochemistry) were determined. RESULTS: Percentage segmental shortening at 6, 24, 48, and 72 h of R was significantly blunted (-1.8 +/- 1.2,* - 0.37 +/- 0. 6,* 0.04 +/- 0.2,* and 5.9 +/- 1.2* vs baseline 17.7 +/- 0.8). MBF (ml/min/g) was attenuated at 24 (0.27 +/- 0.03*), 48 (0.46 +/- 0. 07*), and 72 h of R (0.48 +/- 0.06*) vs 6 h of R (0.65 +/- 0.06). Infarct size increased from 6 (27 +/- 2%) to 24 h of R (41 +/- 2%*) with no further increase at 48 and 72 h of R, consistent with a peak of creatine kinase activity. PMN adherence (mm(2) endothelium) to left anterior descending coronary artery (LAD) segments was increased after 6 h of R (63 +/- 3*) vs nonischemic left circumflex coronary artery (LCX) segments (42 +/- 2) with a peak at 48 h of R (111 +/- 5*). Endothelium-dependent vascular relaxation in the LAD was also blunted at 6, 24, and 48 h of R. Immunostaining revealed CD18-positive PMNs were mainly accumulated in intravascular space during 6 h of R with an increase in migration of PMNs seen at 24 h of R, consistent with a peak of myeloperoxidase release. Myeloperoxidase activity in a given area at risk sample was significantly correlated with infarct extension during the first 24 h of R. CONCLUSIONS: These results provide pathologic evidence for myocardial injury during the extended R and a basis for exploration of interventions designed to limit myocardial injury after ischemia. (*P < 0.05 vs Baseline, 6 h of R and LCX segments.) PMID- 11104654 TI - E- and P-selectin expression depends on the resuscitation fluid used in hemorrhaged rats. AB - BACKGROUND: E- and P-selectins are adhesion molecules that effect neutrophil mediated reperfusion injury. Our hypothesis was that the expression of E- and P selectins is dependent on the type of fluid used for resuscitation and that lactated Ringer's (LR) solution would result in an early upregulation of these molecules. METHODS: Male Sprague-Dawley rats (n = 36) were subjected to a 27 ml/kg hemorrhage over 5 min followed by a 1-h shock period and 1-h of resuscitation. The animals were randomized into the following resuscitation groups: (1) sham; (2) hemorrhage, no resuscitation; (3) whole blood (27 ml/kg); (4) 3:1 lactated Ringer's (81 ml/kg); (5) sham hemorrhage, infusion of lactated Ringer's (81 ml/kg); (6) 7. 5% hypertonic saline (9.7 ml/kg). Immediately after resuscitation, the spleen and lung were harvested for measurement of E- and P selectin mRNA expression with reverse transcriptase- polymerase chain reaction (RT-PCR), and protein expression with immunostaining. RESULTS: LR resuscitation and LR infusion without prior hemorrhage significantly increased the E- and P selectin mRNA in the lung and spleen. Immunostaining demonstrated that the adhesion molecule expression was mainly located in perivascular/peribronchial areas in the lung, and the marginal and trabecular areas in the spleen. Pulmonary edema and inflammatory cell infiltration were observed only in the animals that were hemorrhaged and resuscitated with LR. No resuscitation and resuscitation with whole blood caused no significant increase in selectin expression. CONCLUSION: LR resuscitation and LR infusion without hemorrhage are associated with early increased expression of E- and P-selectin molecules in the lung and spleen. PMID- 11104655 TI - Elastase activity enhances the adhesion of neutrophil and cancer cells to vascular endothelial cells. AB - BACKGROUND: Elastase activity in cancer cells has been reported to promote their metastasis. Hence, we analyzed the influence of elastase activity of cancer cells on their responsive adhesion to vascular endothelial cells. MATERIALS AND METHODS: Human pancreatic (S2-007, S2-013, S2-020, S2-028) and colonic (COLO205) cancer cell lines were used. S2-007, S2-013, and S2-020 possess high elastase activity, whereas S2-028 and COLO205 have low elastase activity. Adhesive reactions of these cancer cells and neutrophils to TNFalpha-activated HUVEC were analyzed. Bound cells onto HUVEC were counted after incubation for 10 min. The effects of suppression of elastase activity by ZD8321, a potent elastase inhibitor, and supplementation of human neutrophil elastase (NE) on the adhesive reactions were also analyzed. In addition, E-selectin expression on HUVEC and concentrations of soluble E-selectin in the medium were measured. RESULTS: Adhesion of cells with high intracellular elastase activity to TNFalpha-activated HUVEC was suppressed by ZD8321. On the other hand, adhesion of cells with low elastase activity was enhanced by exogenous NE. Expression of E-selectin, a key molecule in leukocyte-endothelial cell interaction, on HUVEC was increased by NE. Soluble E-selectin concentration in the medium increased after the adhesive reaction between neutrophils and HUVEC. This increase was thought to be due to the shedding of cell surface E-selectin. Such responses were inhibited by ZD8321. CONCLUSION: Elastase activity has a biological function of stimulating both the E selectin expression on HUVEC and the resultant adhesive reaction of cancer cells with them. Inhibition of elastase activity is a potent strategy for controlling cancer metastasis. PMID- 11104656 TI - Lipopolysaccharide-binding protein accelerates and augments Escherichia coli phagocytosis by alveolar macrophages. AB - BACKGROUND: The first step in bacterial clearance by leukocytes is attachment and phagocytosis. Although lipopolysaccharide-binding protein (LBP) is best known for potentiating LPS-induced cytokine production through a CD14-dependent pathway, recent studies suggest that LBP plays a critical role in clearance of gram negative bacteria and is essential for survival after bacterial challenge. We therefore sought to examine LBP's effect on Escherichia coli phagocytosis by alveolar macrophages (AMs) and to determine if this effect is mediated through CD14. MATERIALS AND METHODS: Phosphatidylinositol-specific phospholipase C (PIPLC)-treated and untreated rat AMs were incubated in the presence of increasing doses of recombinant LBP or negative control protein (choramphenicol acetyltransferase) prior to E. coli-FITC (Ec-F) BioParticle challenge. Phagocytosed bacteria were assayed by fluorescence measurement. A time course study was also performed. RESULTS: LBP potentiated phagocytosis of Ec-F BioParticles by AMs in a dose-dependent fashion. Kinetic studies showed that LBP augmented Ec-F phagocytosis by 76% at 30 min. Treatment of AMs with PIPLC to remove CD14 resulted in only a partial decrease in LBP-mediated enhancement of phagocytosis. CONCLUSION: These results clearly demonstrate that LBP plays an important role in enhancing Ec-F binding and phagocytosis in a time- and dose dependent manner. This observed increase may not require the presence of CD14 as significant potentiation of phagocytosis still occurred after PIPLC treatment. We postulate that the LBP-mediated increase in Ec-F phagocytosis can occur in the absence of CD14 through the presence of another receptor. PMID- 11104657 TI - A novel stable reproducible model of hepatic failure in canines. AB - BACKGROUND: Stable and reproducible large animal models of hepatic failure, which allow the assessment of liver-assist devices, are not available. Our objective was to develop a physiologically stable animal model of hepatic failure on which the safety and efficacy of an extracorporeal liver-assist device can be tested. We hypothesized that a surgical model which consists of an end-to-side portocaval shunt combined with common bile duct ligation and transection would create hepatic failure with: (1) elevations in amino transferases, total bilirubin, and ammonia; (2) a decrease in the ratio of branched chain to aromatic amino acids; and (3) histologic evidence of hepatic injury. METHODS: Eleven mongrel dogs underwent common bile duct transection and an end-to-side portocaval shunt. Aminotransferases (AST, ALT), total bilirubin, ammonia, and branched chain and aromatic amino acids were measured prior to operation (baseline) and after 9 days. A necropsy was performed on Postoperative Day 9 and liver biopsies were obtained for histology. RESULTS: By Postoperative Day 9, AST, ALT, total bilirubin, and ammonia values were significantly elevated compared to baseline (P < 0.02). The ratio of branched chain to aromatic amino acids was significantly reduced compared to baseline (P < 0.003). There was histologic evidence of cholestasis and inflammation. CONCLUSION: Portocaval shunt with common bile duct transection produces liver failure with elevations in aminotransferases, total bilirubin, and ammonia, a decreased branched chain to aromatic amino acid ratio, and histologic inflammation. Unlike ischemic or chemically induced models of liver failure, the dogs were hemodynamically and neurologically stable. This model can be used to test the safety and efficacy of liver-assist devices aimed at temporizing the detoxification functions of the failing liver. PMID- 11104658 TI - Naltrexone administration attenuates surgery-induced immune alterations in rats. AB - Surgery is a commonly performed procedure which produces substantial alterations in immune function in both humans and animals. To better understand the mechanism of surgery-induced immunomodulation, the present study investigated the effect of the opioid antagonist naltrexone on surgery-induced immune alterations in rats. Based on previous investigations in our laboratory, rats underwent a 6-cm laparotomy with no internal manipulation and immunological assessments were completed 24 h following the surgical procedure. Naltrexone was administered at the time of surgery and every 4 h thereafter until immune assessment. Results showed that naltrexone attenuated the surgery-induced decrease in natural killer cell cytotoxicity, B-cell proliferation, T-cell proliferation, and production of the cytokine IFN-gamma. These results are among the first to show that pharmacological antagonism of opioid receptors can prevent deleterious immune changes in the postoperative state, suggesting a detrimental role of the endogenous opioids in surgical procedures. PMID- 11104659 TI - Surgical stress induces phospholipid degradation in the intestinal brush border membrane. AB - BACKGROUND: Surgical stress can lead to translocation of bacteria from the intestine into the systemic circulation. The intestinal brush border membrane (BBM) plays an important role in defense against such invasion by luminal bacteria and endotoxin. Our earlier work has shown the development of oxidative stress in the intestine after surgical stress and since the BBM is sensitive to free radical attack, this study examined the effect of surgical stress on the structure and function of intestinal BBM. METHODS: Intestinal BBM were isolated from control and after surgical stress and compared for structural and functional alterations. Surgical stress was also carried out following pretreatment with the xanthine oxidase inhibitor allopurinol or the nitric oxide donor l-arginine, to study the protection offered by these compounds. RESULTS: Surgical stress affected intestinal BBM structure as well as function. A decrease in alkaline phosphatase activity and alpha-tocopherol content, accompanied by an increase in lipid peroxidation, was seen. Surgical stress induced phospholipid degradation with generation of arachidonic acid. Functional impairment with a decrease in glucose transport ability was also seen. These changes are prevented by inhibition of xanthine oxidase by allopurinol pretreatment but not by NO. CONCLUSION: Surgical stress in the small intestine causes structural and functional alterations in the BBM through oxidative stress. This damage could affect gut barrier integrity and generation of arachidonic acid might mediate distal organ dysfunction. PMID- 11104660 TI - Adhesion molecules in liver ischemia and reperfusion. PMID- 11104661 TI - Worldwide genetic analysis of the CFTR region. AB - Mutations at the cystic fibrosis transmembrane conductance regulator gene (CFTR) cause cystic fibrosis, the most prevalent severe genetic disorder in individuals of European descent. We have analyzed normal allele and haplotype variation at four short tandem repeat polymorphisms (STRPs) and two single-nucleotide polymorphisms (SNPs) in CFTR in 18 worldwide population samples, comprising a total of 1,944 chromosomes. The rooted phylogeny of the SNP haplotypes was established by typing ape samples. STRP variation within SNP haplotype backgrounds was highest in most ancestral haplotypes-although, when STRP allele sizes were taken into account, differences among haplotypes became smaller. Haplotype background determines STRP diversity to a greater extent than populations do, which indicates that haplotype backgrounds are older than populations. Heterogeneity among STRPs can be understood as the outcome of differences in mutation rate and pattern. STRP sites had higher heterozygosities in Africans, although, when whole haplotypes were considered, no significant differences remained. Linkage disequilibrium (LD) shows a complex pattern not easily related to physical distance. The analysis of the fraction of possible different haplotypes not found may circumvent some of the methodological difficulties of LD measure. LD analysis showed a positive correlation with locus polymorphism, which could partly explain the unusual pattern of similar LD between Africans and non-Africans. The low values found in non-Africans may imply that the size of the modern human population that emerged "Out of Africa" may be larger than what previous LD studies suggested. PMID- 11104662 TI - CHRNB2 is the second acetylcholine receptor subunit associated with autosomal dominant nocturnal frontal lobe epilepsy. AB - Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is an uncommon, idiopathic partial epilepsy characterized by clusters of motor seizures occurring in sleep. We describe a mutation of the beta2 subunit of the nicotinic acetylcholine receptor, effecting a V287M substitution within the M2 domain. The mutation, in an evolutionary conserved region of CHRNB2, is associated with ADNFLE in a Scottish family. Functional receptors with the V287M mutation are highly expressed in Xenopus oocytes and characterized by an approximately 10-fold increase in acetylcholine sensitivity. CHRNB2 is a new gene for idiopathic epilepsy, the second acetylcholine receptor subunit implicated in ADNFLE. PMID- 11104663 TI - The latent transforming growth factor beta binding protein (LTBP) family. AB - The transforming growth factor beta (TGFbeta) cytokines are a multi-functional family that exert a wide variety of effects on both normal and transformed mammalian cells. The secretion and activation of TGFbetas is regulated by their association with latency-associated proteins and latent TGFbeta binding proteins (LTBPs). Over the past few years, three members of the LTBP family have been identified, in addition to the protoype LTBP1 first sequenced in 1990. Three of the LTBP family are expressed in a variety of isoforms as a consequence of alternative splicing. This review summarizes the differences between the isoforms in terms of the effects on domain structure and hence possible function. The close identity between LTBPs and members of the fibrillin family, mutations in which have been linked directly to Marfan's syndrome, suggests that anomalous expression of LTBPs may be associated with disease. Recent data indicating that differential expression of LTBP1 isoforms occurs during the development of coronary heart disease is considered, together with evidence that modulation of LTBP function, and hence of TGFbeta activity, is associated with a variety of cancers. PMID- 11104664 TI - Association of immunoproteasomes with the endoplasmic reticulum. AB - Proteasomes are complex multisubunit proteases which play a critical role in intracellular proteolysis. Immunoproteasomes, which contain three gamma interferon-inducible subunits, are a subset of proteasomes which have a specialized function in antigen processing for presentation by the MHC class I pathway. Two of the gamma-interferon inducible subunits, LMP2 and LMP7, are encoded within the MHC class II region adjacent to the two TAP (transporter associated with antigen presentation) genes. We have investigated the localization of immunoproteasomes using monoclonal antibodies to LMP2 and LMP7. Immunoproteasomes were strongly enriched around the endoplasmic reticulum as judged by double-immunofluorescence experiments with anti-calreticulin antibodies, but were also present in the nucleus and throughout the cytosol. In contrast, proteasome subunit C2, which is present in all proteasomes, was found to be evenly distributed throughout the cytoplasm and in the nucleus, as was the delta subunit, which is replaced by LMP2 in immunoproteasomes. gamma-Interferon increased the level of immunoproteasomes, but had no effect on their distribution. Our results provide the first direct evidence that immunoproteasomes are strongly enriched at the endoplasmic reticulum, where they may be located close to the TAP transporter to provide efficient transport of peptides into the lumen of the endoplasmic recticulum for association with MHC class I molecules. PMID- 11104665 TI - A role for the actin cytoskeleton in the hormonal and growth-factor-mediated activation of protein kinase B. AB - We show here that cytochalasin D-induced depolymerization of actin filaments markedly reduces the stimulus-dependent activation of protein kinase B (PKB) in four different cell types (HEK-293 cells, L6 myotubes, 3T3-L1 adipocytes and U87MG cells). HEK-293 cells expressing the pleckstrin homology (PH) domains of PKB and general receptor for phosphoinositides-1 (GRP1) fused to green fluorescent protein (GFP) were used to monitor production of 3-phosphoinositides in the plasma membrane. Disassembly of the actin cytoskeleton significantly reduced the insulin-mediated translocation of both PKB-PH-GFP and GRP1-PH-GFP to the plasma membrane, consistent with diminished synthesis of 3-phosphoinositides. Actin depolymerization did not affect the hormonal activation of phosphoinositide 3-kinase (PI 3-kinase), and since cytochalasin D treatment also led to reduced platelet-derived growth factor (PDGF)-induced phosphorylation of PKB in U87MG cells, a PTEN (phosphatase and tensin homologue deleted on chromosome 10) null cell line, lipid phosphatase activity was unlikely to account for any reduction in cellular 3-phosphoinositides. Withdrawal of cytochalasin D from the extracellular medium induced actin filament repolymerization, and reinstated both the recruitment of PH-GFP fusion proteins to the plasma membrane and PKB activation in response to insulin and PDGF. Our findings indicate that an intact actin network is a crucial requirement for PI 3-kinase-mediated production of 3 phosphoinositides and, therefore, for the activation of PKB. PMID- 11104666 TI - Analysis of myo-inositol hexakisphosphate hydrolysis by Bacillus phytase: indication of a novel reaction mechanism. AB - Phytic acid (myo-inositol hexakisphosphate, InsP(6)) hydrolysis by Bacillus phytase (PhyC) was studied. The enzyme hydrolyses only three phosphates from phytic acid. Moreover, the enzyme seems to prefer the hydrolysis of every second phosphate over that of adjacent ones. Furthermore, it is very likely that the enzyme has two alternative pathways for the hydrolysis of phytic acid, resulting in two different myo-inositol trisphosphate end products: Ins(2,4,6)P(3) and Ins(1,3,5)P(3). These results, together with inhibition studies with fluoride, vanadate, substrate and a substrate analogue, indicate a reaction mechanism different from that of other phytases. By combining the data presented in this study with (1) structural information obtained from the crystal structure of Bacillus amyloliquefaciens phytase [Ha, Oh, Shin, Kim, Oh, Kim, Choi and Oh (2000) Nat. Struct. Biol. 7, 147-153], and (2) computer-modelling analyses of enzyme-substrate complexes, a novel mode of phytic acid hydrolysis is proposed. PMID- 11104667 TI - Organization, chromosomal localization and promoter analysis of the gene encoding human acidic fibroblast growth factor intracellular binding protein. AB - Acidic fibroblast growth factor (aFGF) intracellular binding protein (FIBP) is a protein found mainly in the nucleus that might be involved in the intracellular function of aFGF. Here we present a comparative analysis of the deduced amino acid sequences of human, murine and Drosophila FIBP analogues and demonstrate that FIBP is an evolutionarily conserved protein. The human gene spans more than 5 kb, comprising ten exons and nine introns, and maps to chromosome 11q13.1. Two slightly different splice variants found in different tissues were isolated and characterized. Sequence analysis of the region surrounding the translation start revealed a CpG island, a classical feature of widely expressed genes. Functional studies of the promoter region with a luciferase reporter system suggested a strong transcriptional activity residing within 600 bp of the 5' flanking region. PMID- 11104668 TI - Purification, characterization and sequence analysis of Omp50,a new porin isolated from Campylobacter jejuni. AB - A novel pore-forming protein identified in Campylobacter was purified by ion exchange chromatography and named Omp50 according to both its molecular mass and its outer membrane localization. We observed a pore-forming ability of Omp50 after re-incorporation into artificial membranes. The protein induced cation selective channels with major conductance values of 50-60 pS in 1 M NaCl. N terminal sequencing allowed us to identify the predicted coding sequence Cj1170c from the Campylobacter jejuni genome database as the corresponding gene in the NCTC 11168 genome sequence. The gene, designated omp50, consists of a 1425 bp open reading frame encoding a deduced 453-amino acid protein with a calculated pI of 5.81 and a molecular mass of 51169.2 Da. The protein possessed a 20-amino acid leader sequence. No significant similarity was found between Omp50 and porin protein sequences already determined. Moreover, the protein showed only weak sequence identity with the major outer-membrane protein (MOMP) of Campylobacter, correlating with the absence of antigenic cross-reactivity between these two proteins. Omp50 is expressed in C. jejuni and Campylobacter lari but not in Campylobacter coli. The gene, however, was detected in all three species by PCR. According to its conformation and functional properties, the protein would belong to the family of outer-membrane monomeric porins. PMID- 11104669 TI - p67 isoform of mouse disabled 2 protein acts as a transcriptional activator during the differentiation of F9 cells. AB - The mouse disabled 2 (mDab2) gene is a mouse homologue of the Drosophila disabled gene and is alternatively spliced to form two isoforms, p96 and p67. Although p96 has been known to regulate the Ras-Sos G-protein signal transduction pathway by interacting with Grb2, little is known about the biological function of p67. Recent studies have shown that the expression of mDab2 is markedly up-regulated during the retinoic acid (RA)-induced differentiation of F9 cells, suggesting another role for mDab2 in cell differentiation [Cho, Lee and Park (1999) Mol. Cells 9, 179-184). In the present study, we first elucidated the biological function of p67 isoform of mDab2 and identified its binding partner. Unlike p96, p67 largely resides in RA-treated F9 cell nuclei. In this system, p67 interacts with mouse androgen-receptor interacting protein 3, termed the mDab2 interacting protein, which acts as a transcriptional co-regulator. By using a fusion protein with a heterologous DNA-binding domain (GAL4), we showed that p67 had an intrinsic transcriptional activation function. These results suggest that mDab2 p67 may function as a transcriptional co-factor for certain complexes of transcriptional regulatory elements involved in the RA-induced differentiation of F9 cells. PMID- 11104670 TI - The C-terminus of NIPP1 (nuclear inhibitor of protein phosphatase-1) contains a novel binding site for protein phosphatase-1 that is controlled by tyrosine phosphorylation and RNA binding. AB - Nuclear inhibitor of protein phosphatase-1 (NIPP1; 351 residues) is a nuclear RNA binding protein that also contains in its central domain two contiguous sites of interaction with the catalytic subunit of protein phosphatase-1 (PP1(C)). We show here that mutation of these phosphatase-interaction sites did not completely abolish the ability of NIPP1 to bind and inhibit PP1(C). This could be accounted for by an additional inhibitory phosphatase-binding site in the C-terminal region (residues 311-351), with an inhibitory core corresponding to residues 331-337. Following mutation of all three PP1(C)-binding sites in the central and C terminal domains, NIPP1 no longer interacted with PP1(C). Remarkably, while both NIPP1 domains inhibited the phosphorylase phosphatase activity of PP1(C) independently, mutation of either domain completely abolished the ability of NIPP1 to inhibit the dephosphorylation of myelin basic protein. The inhibitory potency of the C-terminal site of NIPP1 was decreased by phosphorylation of Tyr 335 and by the addition of RNA. Tyr-335 could be phosphorylated by tyrosine kinase Lyn, but only in the presence of RNA. In conclusion, NIPP1 contains two phosphatase-binding domains that function co-operatively but which are controlled independently. Our data are in agreement with a shared-site model for the interaction of PP1(C) with its regulatory subunits. PMID- 11104671 TI - Characterization of the unique function of a reduced amide bond in a cytolytic peptide that acts on phospholipid membranes. AB - The incorporation of a reduced amide bond, psi(CH(2)NH), into peptide results in an increase in the net positive charge and the perturbation of alpha-helical structure. By using this characteristic of the reduced amide bond, we designed and synthesized novel pseudopeptides containing reduced amide bonds, which had a great selectivity between bacterial and mammalian cells. A structure-activity relationship study on pseudopeptides indicated that the decrease in alpha helicity and the increase in net positive charge in the backbone, caused by the incorporation of a reduced amide bond into the peptide, both contributed to an improvement in the selectivity between lipid membranes with various surface charges. However, activity results in vitro indicated that a perturbation of alpha-helical structure rather than an increase in net positive charge in the backbone is more important in the selectivity between bacterial and mammalian cells. The present result revealed that the backbone of membrane-active peptides were important not only in maintaining the secondary structure for the interactions with lipid membranes but also in direct interactions with lipid membranes. The present study showed the unique function of a reduced amide bond in cytolytic peptides and a direction for developing novel anti-bacterial agents from cytolytic peptides that act on the lipid membrane of micro-organisms. PMID- 11104672 TI - In vitro-selected RNA cleaving DNA enzymes from a combinatorial library are potent inhibitors of HIV-1 gene expression. AB - Selective inactivation of a target gene by antisense mechanisms is an important biological tool to delineate specific functions of the gene product. Approaches mediated by ribozymes and RNA-cleaving DNA enzymes (DNA enzymes) are more attractive because of their ability to catalytically cleave the target RNA. DNA enzymes have recently gained a lot of importance because they are short DNA molecules with simple structures that are expected to be stable to the nucleases present inside a mammalian cell. We have designed a strategy to identify accessible cleavage sites in HIV-1 gag RNA from a pool of random DNA enzymes, and for isolation of DNA enzymes. A pool of random sequences (all 29 nucleotides long) that contained the earlier-identified 10-23 catalytic motif were tested for their ability to cleave the target RNA. When the pool of random DNA enzymes was targeted to cleave between any A and U nucleotides, DNA enzyme 1836 was identified. Although several DNA enzymes were identified using a pool of DNA enzymes that was completely randomized with respect to its substrate-binding properties, DNA enzyme-1810 was selected for further characterization. Both DNA enzymes showed target-specific cleavage activities in the presence of Mg(2+) only. When introduced into a mammalian cell, they showed interference with HIV-1 specific gene expression. This strategy could be applied for the selection of desired target sites in any target RNA. PMID- 11104673 TI - Steady-state kinetic mechanism of the NADP+- and NAD+-dependent reactions catalysed by betaine aldehyde dehydrogenase from Pseudomonas aeruginosa. AB - Betaine aldehyde dehydrogenase (BADH) catalyses the irreversible oxidation of betaine aldehyde to glycine betaine with the concomitant reduction of NAD(P)(+) to NADP(H). In Pseudomonas aeruginosa this reaction is a compulsory step in the assimilation of carbon and nitrogen when bacteria are growing in choline or choline precursors. The kinetic mechanisms of the NAD(+)- and NADP(+)-dependent reactions were examined by steady-state kinetic methods and by dinucleotide binding experiments. The double-reciprocal patterns obtained for initial velocity with NAD(P)(+) and for product and dead-end inhibition establish that both mechanisms are steady-state random. However, quantitative analysis of the inhibitions, and comparison with binding data, suggest a preferred route of addition of substrates and release of products in which NAD(P)(+) binds first and NAD(P)H leaves last, particularly in the NADP(+)-dependent reaction. Abortive binding of the dinucleotides, or their analogue ADP, in the betaine aldehyde site was inferred from total substrate inhibition by the dinucleotides, and parabolic inhibition by NADH and ADP. A weak partial uncompetitive substrate inhibition by the aldehyde was observed only in the NADP(+)-dependent reaction. The kinetics of P. aeruginosa BADH is very similar to that of glucose-6-phosphate dehydrogenase, suggesting that both enzymes fulfil a similar amphibolic metabolic role when the bacteria grow in choline and when they grow in glucose. PMID- 11104674 TI - Kinetic alteration of a human dihydrodiol/3alpha-hydroxysteroid dehydrogenase isoenzyme, AKR1C4, by replacement of histidine-216 with tyrosine or phenylalanine. AB - Human dihydrodiol dehydrogenase with 3alpha-hydroxysteroid dehydrogenase activity exists in four forms (AKR1C1-1C4) that belong to the aldo-keto reductase (AKR) family. Recent crystallographic studies on the other proteins in this family have indicated a role for a tyrosine residue (corresponding to position 216 in these isoenzymes) in stacking the nicotinamide ring of the coenzyme. This tyrosine residue is conserved in most AKR family members including AKR1C1-1C3, but is replaced with histidine in AKR1C4 and phenylalanine in some AKR members. In the present study we prepared mutant enzymes of AKR1C4 in which His-216 was replaced with tyrosine or phenylalanine. The two mutations decreased 3-fold the K(m) for NADP(+) and differently influenced the K(m) and k(cat) for substrates depending on their structures. The kinetic constants for bile acids with a 12alpha-hydroxy group were decreased 1.5-7-fold and those for the other substrates were increased 1.3-9-fold. The mutation also yielded different changes in sensitivity to competitive inhibitors such as hexoestrol analogues, 17beta-oestradiol, phenolphthalein and flufenamic acid and 3,5,3', 5'-tetraiodothyropropionic acid analogues. Furthermore, the mutation decreased the stimulatory effects of the enzyme activity by sulphobromophthalein, clofibric acid and thyroxine, which increased the K(m) for the coenzyme and substrate of the mutant enzymes more highly than those of the wild-type enzyme. These results indicate the importance of this histidine residue in creating the cavity of the substrate-binding site of AKR1C4 through the orientation of the nicotinamide ring of the coenzyme, as well as its involvement in the conformational change by binding non-essential activators. PMID- 11104675 TI - Elevated levels of protein-bound p-hydroxyphenylacetaldehyde, an amino-acid derived aldehyde generated by myeloperoxidase, are present in human fatty streaks, intermediate lesions and advanced atherosclerotic lesions. AB - Reactive aldehydes might have a pivotal role in the pathogenesis of atherosclerosis by covalently modifying low-density lipoprotein (LDL). However, the identities of the aldehyde adducts that form on LDL in vivo are not yet clearly established. We previously demonstrated that the haem protein myeloperoxidase oxidizes proteins in the human artery wall. We also have shown that p-hydroxyphenylacetaldehyde (pHA), the aldehyde that forms when myeloperoxidase oxidizes L-tyrosine, covalently modifies the N(epsilon)-lysine residues of proteins. The resulting Schiff base can be quantified as N(epsilon) [2-(p-hydroxyphenyl)ethyl]lysine (pHA-lysine) after reduction with NaCNBH(3). Here we demonstrate that pHA-lysine is a marker for LDL that has been modified by myeloperoxidase, and that water-soluble, but not lipid-soluble, antioxidants inhibit the modification of LDL protein. To determine whether myeloperoxidase generated aldehydes might modify LDL in vivo, we used a combination of isotope dilution GC-MS to quantify pHA-lysine in aortic tissues at various stages of lesion evolution. We also analysed LDL isolated from atherosclerotic aortic tissue. Comparison of normal and atherosclerotic aortic tissue demonstrated a significant elevation (more than 10-fold) of the reduced Schiff base adduct in fatty streaks, intermediate lesions and advanced lesions compared with normal aortic tissue. Moreover, the level of pHA-lysine in LDL recovered from atherosclerotic aortic intima was 200-fold that in plasma LDL of healthy donors. These results indicate that pHA-lysine, a specific covalent modification of LDL, is generated in human atherosclerotic vascular tissue. They also raise the possibility that reactive aldehydes generated by myeloperoxidase have a role in converting LDL into an atherogenic lipoprotein. PMID- 11104677 TI - Effects of elevated expression of inositol 1,4,5-trisphosphate 3-kinase B on Ca2+ homoeostasis in HeLa cells. AB - Ins(1,4,5)P(3) 3-kinase (IP3K) phosphorylates the Ca(2+)-mobilizing second messenger Ins(1,4,5)P(3) to yield the putative second messenger Ins(1,3,4,5)P(4). A HeLa cell line was established expressing the rat B isoform of IP3K under the control of an inducible promoter. The IP3KB-transfected cell line possessed 23 fold greater IP3K activity than untransfected cells after induction of IP3KB expression, but only 0.23-fold greater activity when IP3KB expression was not induced. Elevating IP3KB expression significantly reduced levels of Ins(1,4,5)P(3) and increased levels of Ins(1,3,4,5)P(4) after stimulation of cells with histamine, but had no effect on basal levels. Histamine- and ATP evoked cytosolic Ca(2+) responses were dramatically reduced upon elevation of IP3KB expression. On stimulation with a supramaximal dose of histamine, 67% of cells induced to express IP3KB gave no detectable elevation in cytosolic Ca(2+), compared with 3% of uninduced cells. The quantity of Ca(2+) within thapsigargin sensitive and -insensitive stores was unaffected by elevation of IP3KB expression, as was capacitative Ca(2+) entry. These data suggest that IP3KB may play a significant role in the regulation of Ins(1,4,5)P(3) levels, and consequently in Ca(2+) responses following stimulation of cells with Ins(1,4,5)P(3)-elevating agonists. PMID- 11104676 TI - Compartment-specific regulation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinases (MAPKs) by ERK-dependent and non-ERK-dependent inductions of MAPK phosphatase (MKP)-3 and MKP-1 in differentiating P19 cells. AB - Activation of mitogen-activated protein kinases (MAPKs), their upstream activators MAPK kinases (MAPKKs or MEKs) and induction of MKP-1 (CL100/3CH134) and MKP-3 (Pyst1/rVH6) dual-specificity MAPK phosphatases (MKPs) were studied in the mouse embryonic stem cell line P19 during the 7 day induction of neuronal differentiation triggered by aggregation and retinoic acid. ERK (extracellular signal-regulated kinase), but not JNK (c-Jun N-terminal kinase), was found activated with biphasic kinetics: a first transient phase on days 1 and 2, followed by a second activation that was sustained until the appearance of a neuronal phenotype. MEK activation appeared coincident with ERK activation. Cytosolic MKP-3 was induced in parallel to ERK activation, the induction being dependent on ERK activation, as was shown using the MEK-1 inhibitor PD98059. In contrast, nuclear MKP-1 was transiently elevated at 48 h, coincident with ERK inactivation and independently of ERK activity. As shown by cell fractionation, activated ERK is translocated to the nucleus. The complementary induction of ERK specific phosphatases MKP-1 and MKP-3 permits precise and independent control of cytoplasmic and nuclear ERK activity, most probably required to properly induce a complex cellular programme of differentiation. PMID- 11104678 TI - Conversion of Escherichia coli pyruvate oxidase to an 'alpha-ketobutyrate oxidase'. AB - Escherichia coli pyruvate oxidase (PoxB), a lipid-activated homotetrameric enzyme, is active on both pyruvate and 2-oxobutanoate ('alpha-ketobutyrate'), although pyruvate is the favoured substrate. By localized random mutagenesis of residues chosen on the basis of a modelled active site, we obtained several PoxB enzymes that had a markedly decreased activity with the natural substrate, pyruvate, but retained full activity with 2-oxobutanoate. In each of these mutant proteins Val-380 had been replaced with a smaller residue, namely alanine, glycine or serine. One of these, PoxB V380A/L253F, was shown to lack detectable pyruvate oxidase activity in vivo; this protein was purified, studied and found to have a 6-fold increase in K(m) for pyruvate and a 10-fold lower V(max) with this substrate. In contrast, the mutant had essentially normal kinetic constants with 2-oxobutanoate. The altered substrate specificity was reflected in a decreased rate of pyruvate binding to the latent conformer of the mutant protein owing to the V380A mutation. The L253F mutation alone had no effect on PoxB activity, although it increased the activity of proteins carrying substitutions at residue 380, as it did that of the wild-type protein. The properties of the V380A/L253F protein provide new insights into the mode of substrate binding and the unusual activation properties of this enzyme. PMID- 11104679 TI - Unique kinetics of nicotinic acid-adenine dinucleotide phosphate (NAADP) binding enhance the sensitivity of NAADP receptors for their ligand. AB - Nicotinic acid-adenine dinucleotide phosphate (NAADP) is a novel and potent Ca(2+)-mobilizing agent in sea urchin eggs and other cell types. Little is known, however, concerning the properties of the putative intracellular NAADP receptor. In the present study we have characterized NAADP binding sites in sea urchin egg homogenates. [(32)P]NAADP bound to a single class of high-affinity sites that were reversibly inhibited by NaCl but insensitive to pH and Ca(2+). Binding of [(32)P]NAADP was lost in preparations that did not mobilize Ca(2+) in response to NAADP, indicating that [(32)P]NAADP probably binds to a receptor mediating Ca(2+) mobilization. Addition of excess unlabelled NAADP, at various times after initiation of [(32)P]NAADP binding, did not result in displacement of bound [(32)P]NAADP. These data show that NAADP becomes irreversibly bound to its receptor immediately upon association. Accordingly, incubation of homogenates with low concentrations of NAADP resulted in maximal labelling of NAADP binding sites. This unique property renders NAADP receptors exquisitely sensitive to their ligand, thereby allowing detection of minute changes in NAADP levels. PMID- 11104680 TI - Expression and regulation of pyruvate dehydrogenase kinase isoforms in the developing rat heart and in adulthood: role of thyroid hormone status and lipid supply. AB - Activation of the pyruvate dehydrogenase (PDH) complex (PDHC) promotes glucose disposal, whereas inactivation conserves glucose. The PDH kinases (PDHKs) regulate glucose oxidation through inhibitory phosphorylation of PDHC. The adult rat heart contains three PDHK isoforms PDHK1, PDHK2 and PDHK4. Using Western-blot analysis, with specific antibodies raised against individual recombinant PDHK1, PDHK2 and PDHK4, the present study investigated PDHK isoform expression in the developing rat heart and adulthood. We identified clear differences in the patterns of protein expression of each of these PDHK isoforms during the first 3 weeks of post-natal development, with most marked up-regulation of isoforms PDHK1 and PDHK4. Distinctions between the three cardiac PDHK isoforms were also demonstrated with respect to post-neonatal maturational up-regulation; with greatest up-regulation of PDHK1 and least up-regulation of PDHK4 from the post neonatal period until maturity. The study also examined the role of thyroid hormone status and lipid supply on PDHK isoform expression. We observed marked selective increases in the amount of PDHK4 protein present relative to total cardiac protein in both hyperthyroidism and high-fat feeding. Overall, our data identify PDHK isoform PDHK1 as being of more potential regulatory importance for glucose oxidation in the adult compared with the neonatal heart, and cardiac PDHK4 as a PDHK isoform whose expression is specifically responsive to changes in lipid supply, suggesting that its up-regulation during early post-natal life may be the perinatal switch to use fatty acids as the energy source. We also identify regulation of pyruvate sensitivity of cardiac PDHK as a physiological variable, a change in which requires factors in addition to a change in lipid supply. PMID- 11104681 TI - Susceptibility of mitogen-activated protein kinase kinase family members to proteolysis by anthrax lethal factor. AB - The lethal factor (LF) produced by toxigenic strains of Bacillus anthracis is a Zn(2+)-endopeptidase that cleaves the mitogen-activated protein kinase kinases (MAPKKs) MEK1, MEK2 and MKK3. Using genetic and biochemical approaches, we have extended the study of LF proteolytic specificity to all known MAPKK family members and found that LF also cleaves MKK4, MKK6 and MKK7, but not MEK5. The peptide bonds hydrolysed by LF within all MAPKKs were identified. Cleavage invariably occurs within the N-terminal proline-rich region preceding the kinase domain, thus disrupting a sequence involved in directing specific protein-protein interactions necessary for the assembly of signalling complexes. Alignment of the sequences flanking the site of cleavage reveals the occurrence of some consensus motifs: position P2 and P1' are occupied by hydrophobic residues and at least one basic residue is present between P4 and P7. The implications of these findings for the biochemical activity and functional specificity of LF are discussed. PMID- 11104682 TI - Specific induction of RGS16 (regulator of G-protein signalling 16) mRNA by protein kinase C in CEM leukaemia cells is mediated via tumour necrosis factor alpha in a calcium-sensitive manner. AB - The RGS (regulator of G-protein signalling) proteins are GTPase-activating proteins for activated Galpha subunits. We investigated the effects of protein kinase C (PKC) on RGS proteins in various T cell lines by treating them with PMA. mRNA levels of both RGS16 and tumour necrosis factor alpha (TNFalpha) were found to be up-regulated in CEM leukaemia cells in a PKC-dependent manner. Mezerein, a non-phorbol-ester activator of PKC, also elevated RGS16 and TNFalpha mRNA levels, while the specific PKC inhibitor Go6983 abrogated their expression. In view of the slower kinetics of PMA-induced RGS16 expression and the tight correlation between TNFalpha and RGS16 mRNA induction among the cell lines studied, we suggest that activation of PKC up-regulates RGS16 via TNFalpha. Indeed, addition of recombinant TNFalpha to CEM cells rapidly stimulated RGS16 mRNA expression independently of PKC. Furthermore, mobilization of calcium by A23187 and thapsigargin blocked the TNFalpha-mediated induction of RGS16, which was reversed by EGTA and by the immunosuppressants FK506 and cyclosporin A, suggesting that the calcineurin/NF-AT (nuclear factor of activated T cells) pathway may repress the up-regulation process. Our results demonstrate for the first time that activation of PKC induces RGS16 expression via TNFalpha in a calcium-sensitive manner, thereby implicating RGS16 in the regulation of T cell responses to inflammation. PMID- 11104683 TI - Biochemical characterization of a trypanosome enzyme with glutathione-dependent peroxidase activity. AB - In most eukaryotes, glutathione-dependent peroxidases play a key role in the metabolism of peroxides. Numerous studies have reported that trypanosomatids lack this activity. Here we show that this is not the case, at least for the American trypanosome Trypanosoma cruzi. We have isolated a single-copy gene from T. cruzi with the potential to encode an 18 kDa enzyme, the sequence of which has highest similarity with glutathione peroxidases from plants. A recombinant form of the protein was purified following expression in Escherichia coli. The enzyme was shown to have peroxidase activity in the presence of glutathione/glutathione reductase but not in the presence of trypanothione/trypanothione reductase. It could metabolize a wide range of hydroperoxides (linoleic acid hydroperoxide and phosphatidylcholine hydroperoxide>cumene hydroperoxide>t-butyl hydroperoxide), but no activity towards hydrogen peroxide was detected. Enzyme activity could be saturated by glutathione when both fatty acid and short-chain organic hydroperoxides were used as substrate. For linoleic acid hydroperoxide, the rate limiting step of this reaction is the reduction of the peroxidase by glutathione. With lower-affinity substrates such as t-butyl hydroperoxide, the rate-limiting step is the reduction of the oxidant. The data presented here identify a new arm of the T. cruzi oxidative defence system. PMID- 11104684 TI - Nuclear receptors modulate the interaction of Sp1 and GC-rich DNA via ternary complex formation. AB - Binding sites for transcription factor Sp1 have been implicated in the transcriptional regulation of several genes by hormones or vitamins, and here we show that a GC-rich element contributes to the retinoic acid response of the interleukin 1beta promoter. To explain such observations, it has been proposed that nuclear receptors can interact with Sp1 bound to GC-rich DNA. However, evidence supporting this model has remained indirect. So far, nuclear receptors have not been detected in a complex with Sp1 and GC-rich DNA, and the expected ternary complexes in non-denaturing gels were not seen. In search for these missing links we found that nuclear receptors [retinoic acid receptor (RAR), thyroid hormone receptor (TR), vitamin D(3) receptor, peroxisome-proliferator activated receptor and retinoic X receptor] induce an electrophoretic mobility increase of Sp1-GC-rich DNA complexes. Concomitantly, binding of Sp1 to the GC box is enhanced. It is proposed that nuclear receptors may partially replace Sp1 in homo-oligomers at the GC-box. RARs and Sp1 can also combine into a complex with a retinoic acid-response element. The presence of RAR and Sp1 in complexes with either cognate site was revealed in supershift experiments. The C-terminus of Sp1 interacts with nuclear receptors. Both the ligand- and DNA-binding domains of the receptor are important for complex formation with Sp1 and GC-rich DNA. In spite of similar capacity to form ternary complexes, RAR but not TR up-regulated an Sp1-driven reporter in a ligand-dependent way. Thus additional factors limit the transcriptional response mediated by nuclear receptors and Sp1. PMID- 11104685 TI - Autophagosome-associated variant isoforms of cytosolic enzymes. AB - In a search for autophagosome-associated proteins, two-dimensional gel separations of proteins from purified autophagosomes, postnuclear supernatant, cytosol, lysosomes, mitochondria, endosomes and a cytomembrane fraction (mostly endoplasmic reticulum) were compared. Three proteins, with monomeric molecular masses of 43, 35 and 31 kDa, were enriched in total or sedimentable fractions of autophagosomes relative to the corresponding fractions of postnuclear supernatant, suggesting an association with the autophagosomal delimiting membrane. These proteins were also present on lysosomal membranes, but they were absent from mitochondria, and detected only in small amounts in the cytomembrane fraction and in endosomes, indicating that they were not associated with organelles sequestered by autophagy. However, all three proteins were present in the cytosol, suggesting that they were cytosolic proteins binding peripherally to the delimiting membrane of autophagosomes, probably to its innermost surface as indicated by their resistance to treatment of intact autophagosomes with proteinase or protein-stripping agents. Amino acid sequencing identified these proteins as an isoform of argininosuccinate synthase, an N-truncated variant of glyceraldehyde-3-phosphate dehydrogenase, and a sequence variant of short-chain 2 enoyl-CoA hydratase. PMID- 11104686 TI - Inverse regulation of F1-ATPase activity by a mutation at the regulatory region on the gamma subunit of chloroplast ATP synthase. AB - Chloroplast ATP synthase is a thiol-modulated enzyme whose DeltamuH(+)-linked activation is strongly influenced by reduction and the formation of a disulphide bridge between Cys(199) and Cys(205) on the gamma subunit. In solubilized chloroplast coupling factor 1 (CF(1)), reduction of the disulphide bond elicits the latent ATP-hydrolysing activity. To assess the regulatory importance of the amino acid residues around these cysteine residues, we focused on the three negatively charged residues Glu(210)-Asp-Glu(212) close to the two cysteine residues and also on the following region from Leu(213) to Ile(230), and investigated the modulation of ATPase activity by chloroplast thioredoxins. The mutant gamma subunits were reconstituted with the alpha and beta subunits from F(1) of the thermophilic bacterium Bacillus PS3; the active ATPase complexes obtained were purified by gel-filtration chromatography. The complex formed with a mutant gamma subunit in which Glu(210) to Glu(212) had been deleted was inactivated rather than activated by reduction of the disulphide bridge by reduced thioredoxin, indicating inverse regulation. This complex was insensitive to the inhibitory CF(1)-epsilon subunit when the mutant gamma subunit was oxidized. In contrast, the deletion of Glu(212) to Ile(230) converted the complex from a modulated state into a highly active state. PMID- 11104687 TI - Chloride channel activity of ClC-2 is modified by the actin cytoskeleton. AB - The chloride channel ClC-2 has been implicated in essential physiological functions, including cell-volume regulation and fluid secretion by specific epithelial tissues. Although ClC-2 is known to be activated by hyperpolarization and hypo-osmotic shock, the molecular basis for the regulation of this channel remains unclear. Here we show in the Xenopus oocyte expression system that the chloride-channel activity of ClC-2 is enhanced after treatment with the actin disrupting agents cytochalasin and latrunkulin. These findings suggest that the actin cytoskeleton normally exerts an inhibitory effect on ClC-2 activity. An inhibitory domain was previously defined in the N-terminus of ClC-2, so we sought to determine whether this domain might interact directly with actin in binding assays in vitro. We found that a glutathione S-transferase fusion protein containing the inhibitory domain was capable of binding actin in overlay and co sedimentation assays. Further, the binding of actin to this relatively basic peptide (pI 8.4) might be mediated through electrostatic interactions because binding was inhibited at high concentrations of NaCl with a half-maximal decrease in signal at 180 mM NaCl. This work suggests that electrostatic interactions between the N-terminus of ClC-2 and the actin cytoskeleton might have a role in the regulation of this channel. PMID- 11104688 TI - Interaction between DNA-damage protein GADD34 and a new member of the Hsp40 family of heat shock proteins that is induced by a DNA-damaging reagent. AB - GADD34 is one of a subset of proteins induced after DNA damage or cell growth arrest. To examine the function of GADD34, we used the yeast two-hybrid system to clone the protein that interacts with murine GADD34. As bait we used the product of the partial GADD34 cDNA, including the regions rich in proline, glutamic acid, serine and threonine (PEST) and gamma(1)34.5 regions. A cDNA clone, named GAHSP40, which is a mouse DnaJ family protein with a high similarity to human HLJ1 was cloned. The interaction between GADD34 and GAHSP40 in cultured cells was confirmed by a co-immunoprecipitation experiment and in NIH 3T3 cells by two hybrid analysis in vivo. For binding of the two proteins, the gamma(1)34.5 similar region of GADD34 was necessary; however, the PEST region was also involved and the C-terminus of GAHSP40, but not the J-domain, was important. GAHSP40 was detected in all mouse tissues examined, but a different transcript was found in the testis. Both GADD34 mRNA and GAHSP40 mRNA were significantly elevated by treatment with methyl methanesulphonate, although the time courses were different. In addition, both GAHSP40 and GADD34 mRNA were induced by heat shock. PMID- 11104689 TI - Phosphorylation of methylated-DNA-protein-cysteine S-methyltransferase at serine 204 significantly increases its resistance to proteolytic digestion. AB - In a previous paper [Lim, Park, Jee, Lee and Paik (1999) J. Cancer Res. Clin. Oncol. 125, 493-499], we showed two major forms of active DNA-6-O methylguanine:protein-L-cysteine S-methyltransferase (MGMT; EC 2.1.1.63) in the liver with N-nitrosodiethylamine (DEN)-induced carcinogenesis: these were 26 and 24 kDa species. Here we show that a 2 kDa C-terminal fragment was cleaved from the 26 kDa species in vitro by thrombin or microsomal fractions isolated from DEN treated rat livers. When Ser(204) of the 26 kDa protein was replaced with Ala by site-directed mutagenesis, phosphorylation of the protein was completely abolished, indicating Ser(204) to be the site of phosphorylation. We also show that the phosphorylation was performed by Ca(2+)-independent protein kinase isoenzymes, and that the phosphorylated rat MGMT protein was resistant to digestion by protease(s) whose activity was increased during DEN-induced hepatocarcinogenesis and also by digestion with endopeptidase Glu-C (V8 protease). PMID- 11104690 TI - Interaction of sphingosine 1-phosphate with plasma components, including lipoproteins, regulates the lipid receptor-mediated actions. AB - The concentration of sphingosine 1-phosphate (S1P) in plasma or serum is much higher than the half-maximal concentration of the sphingolipid needed to stimulate its receptors. Nevertheless, the inositol phosphate response to plasma or serum mediated by Edg-3, one of the S1P receptors, which was overexpressed in Chinese hamster ovary cells, was much smaller than the response expected from the total amount of S1P in these samples. The inositol phosphate response to exogenous S1P was markedly attenuated in the presence of charcoal-treated low-S1P serum. The inhibitory effect was lost by boiling but not by dialysis of the serum. The inhibitory action of the serum was specific to S1P and was associated with the trapping of exogenous S1P; the inositol phosphate response to P(2) purinergic agonists was somewhat enhanced by the charcoal-treated serum. Among the components of plasma or serum, lipoproteins such as low-density and high density lipoproteins showed a stronger activity for trapping S1P than lipoprotein deficient serum. Consistent with this observation, we detected a 15-100-fold higher amount of S1P per unit amount of protein in lipoproteins than in the lipoprotein-deficient serum. Thus even though the protein content of the lipoprotein fraction contributes to only 4% of the total protein content of plasma or serum, more than 60% of S1P is distributed in this fraction. These results suggest that the tight binding of S1P to the components of serum or plasma, including lipoproteins, may interfere with the S1P binding to its receptors and thereby attenuate the lipid-receptor-mediated actions in the cells. PMID- 11104692 TI - Dynamic palmitoylation of lymphoma proprotein convertase prolongs its half-life, but is not essential for trans-Golgi network localization. AB - Proprotein convertases are responsible for the endoproteolytic activation of proproteins in the secretory pathway. The most recently discovered member of this family, lymphoma proprotein convertase (LPC), is a type-I transmembrane protein. Previously, we have demonstrated that its cytoplasmic tail is palmitoylated. In this study, we have identified the two most proximal cysteine residues in the cytoplasmic tail as palmitoylation sites. Substitution of either cysteine residue by alanine interfered with palmitoylation of the other. Palmitoylation of LPC was found to be sensitive to the protein palmitoyltransferase inhibitor tunicamycin but not cerulenin. It was also insensitive to the drugs brefeldin A, monensin and cycloheximide, indicating that the modification occurs in a late exocytic or endocytic compartment. Turnover of palmitoylated LPC is significantly faster (t(1/2) approximately 50 min) than that of the LPC polypeptide backbone (t(1/2) approximately 3 h), suggesting that palmitoylation is reversible. Abrogation of palmitoylation reduced the half-life of the LPC protein, but did not affect steady-state localization of LPC in the trans-Golgi network. Finally, LPC could not be detected in detergent-resistant membrane rafts. Taken together, these results suggest that dynamic palmitoylation of LPC is important for stability, but does not function as a dominant trafficking signal. PMID- 11104691 TI - Homocysteine stimulates nuclear factor kappaB activity and monocyte chemoattractant protein-1 expression in vascular smooth-muscle cells: a possible role for protein kinase C. AB - Monocyte chemoattractant protein-1 (MCP-1) is a potent chemokine that stimulates the migration of monocytes into the intima of arterial walls. Although many factors that induce MCP-1 expression have been identified, the effect of homocysteine on the expression of MCP-1 in atherogenesis and the underlying mechanisms are not entirely clear. The objective of the present study was to investigate the role of homocysteine in MCP-1 expression in human aorta vascular smooth-muscle cells (VSMCs). After VSMCs were incubated with homocysteine for various time periods, a nuclease protection assay and ELISA were performed. Homocysteine (0.05-0.2 mM) significantly increased the expression of MCP-1 mRNA (up to 2. 7-fold) and protein (up to 3.3-fold) in these cells. The increase in MCP-1 expression was associated with the activation of protein kinase C (PKC) as well as nuclear factor kappaB (NF-kappaB). Further investigation demonstrated that the activation of NF-kappaB was the result of a PKC-mediated reduction in the expression of inhibitory protein (IkappaBalpha) mRNA and protein in homocysteine-treated cells. Oxidative stress might also be involved in the activation of NF-kappaB by homocysteine in VSMCs. In conclusion, the present study has clearly demonstrated that the activation of PKC as well as superoxide production followed by activation of NF-kappaB is responsible for homocysteine induced MCP-1 expression in VSMCs. These results suggest that homocysteine stimulated MCP-1 expression via NF-kappaB activation may play an important role in atherogenesis. PMID- 11104693 TI - Conversion of a synthetic fructosamine into its 3-phospho derivative in human erythrocytes. AB - Intact human erythrocytes catalyse the conversion of fructose into fructose 3 phosphate with an apparent K(m) of 30 mM [Petersen, Kappler, Szwergold and Brown (1992) Biochem. J. 284, 363-366]. The physiological significance of this process is still unknown. In the present study we report that the formation of fructose 3 phosphate from 50 mM fructose in intact erythrocytes is inhibited by 1-deoxy-1 morpholinofructose (DMF), a synthetic fructosamine, with an apparent K(i) of 100 microM. (31)P NMR analysis of cell extracts incubated with DMF indicated the presence of an additional phosphorylated compound, which was partially purified and shown to be DMF 3-phosphate by tandem MS. Radiolabelled DMF was phosphorylated by intact erythrocytes with an apparent K(m) ( approximately 100 microM) approx. 300-fold lower than the value reported for fructose phosphorylation on its third carbon. These results indicate that the physiological function of the enzyme that is able to convert fructose into fructose 3-phosphate in intact erythrocytes is probably to phosphorylate fructosamines. This suggests that fructosamines, which are produced non enzymically from glucose and amino compounds, may be metabolized in human erythrocytes. PMID- 11104694 TI - Size of the ligand complex between the N-terminal domain of the gene III coat protein and the non-infectious phage strongly influences the usefulness of in vitro selective infective phage technology. AB - The selective infective phage (SIP) technology allows a rapid positive selection of interacting pairs of biological molecules that restore to non-infectious phages their ability to infect the bacterial host. After a successful infection, the phage is amplified and the DNA encoding the interacting ligand is isolated from the phage genome and sequenced. In our studies we have evaluated the usefulness of SIP for the identification and cloning of proteins interacting with a biotinylated target binding to a newly designed adapter molecule consisting of streptavidin fused to the C-terminus of the extracellular domain of the phage minor coat protein III. The new adapter was expressed in Escherichia coli and refolded from inclusion bodies. The two different domains joined within the chimaera were found to be biologically functional. We also demonstrated that non covalent interactions between a non-infectious phage displaying a short peptide, which specifically binds the streptavidin, and the adapter molecule restore phage infectivity. To evaluate the potential of SIP as a general and generic tool for the screening of cDNA libraries that encode the ligands displayed at the surface of the phage and binding to biotinylated targets, we have increased both the size of the displayed ligand on the phage and the size of the biotinylated target bound to the streptavidin domain of the adapter molecule. In our model systems we show that the size of either the ligand or the target is a limiting factor for the technology. PMID- 11104695 TI - Human deoxyhypusine synthase: interrelationship between binding of NAD and substrates. AB - Deoxyhypusine synthase catalyses the NAD-dependent transfer of the butylamine moiety from the polyamine, spermidine, to a specific lysine residue of a single cellular protein, eukaryotic translation-initiation factor 5A (eIF5A) precursor. The native enzyme exists as a tetramer of four identical subunits and contains four binding sites for NAD. The binding of spermidine and NAD was studied by a filtration assay. [(3)H]Spermidine binding to the enzyme was not detectable alone or in the presence of the eIF5A precursor, but was detected only in the presence of NAD or NADH, suggesting that a NAD/NADH-induced conformational change is required for the binding of spermidine. A strong NAD-dependent binding was also observed with a spermidine analogue, N(1)-guanyl-1, 7-diamino[(3)H]heptane (GC(7)), but not with [(14)C]putrescine or [(14)C]spermine. Although [(3)H]NAD binding to the enzyme occurred in the absence of spermidine, its affinity for the enzyme was markedly enhanced by spermidine, GC(7) and also by the eIF5A precursor. The maximum binding for NAD and spermidine was estimated to be approximately 4 molecules each/enzyme tetramer. The dependence of spermidine binding on NAD and the modulation of binding of NAD by spermidine and the eIF5A precursor suggest intricate relationships between the binding of cofactor and the substrates, and provide new insights into the reaction mechanism. PMID- 11104696 TI - Thiocyanate binding to the molybdenum centre of the periplasmic nitrate reductase from Paracoccus pantotrophus. AB - The periplasmic nitrate reductase (NAP) from Paracoccus pantotrophus is a soluble two-subunit enzyme (NapAB) that binds two haem groups, a [4Fe-4S] cluster and a bis(molybdopterin guanine dinucleotide) (MGD) cofactor that catalyses the reduction of nitrate to nitrite. In the present study the effect of KSCN (potassium thiocyanate) as an inhibitor and Mo ligand has been investigated. Results are presented that show NAP is sensitive to SCN(-) (thiocyanate) inhibition, with SCN(-) acting as a competitive inhibitor of nitrate (K(i) approximately 4.0 mM). The formation of a novel EPR Mo(V) species with an elevated g(av) value (g(av) approximately 1.994) compared to the Mo(V) High-g (resting) species was observed upon redox cycling in the presence of SCN(-). Mo K edge EXAFS analysis of the dithionite-reduced NAP was best fitted as a mono-oxo Mo(IV) species with three Mo-S ligands at 2.35 A (1 A=0.1 nm) and a Mo-O ligand at 2.14 A. The addition of SCN(-) to the reduced Mo(IV) NAP generated a sample that was best fitted as a mono-oxo (1.70 A) Mo(IV) species with four Mo-S ligands at 2.34 A. Taken together, the competitive nature of SCN(-) inhibition of periplasmic nitrate reductase activity, the elevated Mo(V) EPR g(av) value following redox cycling in the presence of SCN(-) and the increase in sulphur co ordination of Mo(IV) upon SCN(-) binding, provide strong evidence for the direct binding of SCN(-) via a sulphur atom to Mo. PMID- 11104697 TI - Control of pancreatic bile-salt-dependent-lipase secretion by the glucose regulated protein of 94 kDa (Grp94). AB - Bile-salt-dependent lipase (BSDL; EC 3.1.1.13) is an enzyme expressed by the pancreatic acinar cell and secreted as a component of the pancreatic juice. During its route towards secretion, BSDL is associated with intracellular membranes by means of a multiprotein folding complex, which includes the glucose regulated protein of 94 kDa (Grp94). We have postulated that the association of BSDL with membranes is required for the complete O-glycosylation of the protein, which diverts BSDL from a degradation route and consequently allows its secretion. To further characterize the role of Grp94 in BSDL secretion, we have studied the effect of a ribozyme specifically targeted to Grp94 mRNA. This ribozyme has been transfected into AR4-2J cells, and we have shown that a decrease in Grp94 expression leads to a concomitant decrease in BSDL secretion and expression. Geldanamycin (GA), which alters Grp94 functions, also affects the release of BSDL into the culture medium of AR4-2J cells. BSDL expressed in GA treated AR4-2J cells is unstable. Furthermore, under conditions that decrease the level of BSDL secretion, no intracellular accumulation of the enzyme was observed, suggesting that BSDL that cannot associate with (or be structured by) Grp94 could be rapidly degraded. We have further shown that this degradation probably occurs via the ubiquitin-dependent pathway. Altogether, these results indicate that Grp94 has a pivotal role in BSDL folding and in the sorting of this pancreatic enzyme. PMID- 11104698 TI - Purification and characterization of cytosolic pyruvate kinase from banana fruit. AB - Cytosolic pyruvate kinase (PK(c)) from ripened banana (Musa cavendishii L.) fruits has been purified 543-fold to electrophoretic homogeneity and a final specific activity of 59.7 micromol of pyruvate produced/min per mg of protein. SDS/PAGE and gel-filtration FPLC of the final preparation indicated that this enzyme exists as a 240 kDa homotetramer composed of subunits of 57 kDa. Although the enzyme displayed a pH optimum of 6.9, optimal efficiency in substrate utilization [in terms of V(max)/K(m) for phosphoenolpyruvate (PEP) or ADP] was equivalent at pH 6.9 and 7.5. PK(c) activity was absolutely dependent upon the presence of a bivalent and a univalent cation, with Mg(2+) and K(+) respectively fulfilling this requirement. Hyperbolic saturation kinetics were observed for the binding of PEP, ADP, Mg(2+) and K(+) (K(m) values of 0.098, 0.12, 0.27 and 0.91 mM respectively). Although the enzyme utilized UDP, IDP, GDP and CDP as alternative nucleotides, ADP was the preferred substrate. L-Glutamate and MgATP were the most effective inhibitors, whereas L-aspartate functioned as an activator by reversing the inhibition of PK(c) by L-glutamate. The allosteric features of banana PK(c) are compared with those of banana PEP carboxylase [Law and Plaxton (1995) Biochem. J. 307, 807-816]. A model is presented which highlights the roles of cytosolic pH, MgATP, L-glutamate and L-aspartate in the co-ordinate control of the PEP branchpoint in ripening bananas. PMID- 11104699 TI - Metalloprotease-disintegrin ADAM 12 binds to the SH3 domain of Src and activates Src tyrosine kinase in C2C12 cells. AB - ADAM 12, a member of the ADAM (protein containing a disintegrin and metalloprotease) family of metalloprotease-disintegrins, has been implicated in the differentiation and fusion of skeletal myoblasts, and its expression is dramatically up-regulated in many cancer cells. While the extracellular portion of ADAM 12 contains an active metalloprotease and a cell-adhesion domain, the function of the cytoplasmic portion is much less clear. In this paper, we show that the cytoplasmic tail of ADAM 12 mediates interactions with the non-receptor protein tyrosine kinase Src. The interaction is direct, specific, and involves the N-terminal proline-rich region in the cytoplasmic tail of ADAM 12 and the Src homology 3 (SH3) domain of Src. ADAM 12 and Src co-immunoprecipitate from transfected C2C12 cells, suggesting that the two proteins form a complex in vivo. Co-expression of Src and ADAM 12, but not ADAM 9, in C2C12 cells results in activation of the recombinant Src. Moreover, endogenous ADAM 12 associates with and activates endogenous Src in differentiating C2C12 cells. These results indicate that ADAM 12 may mediate adhesion-induced signalling during myoblast differentiation. PMID- 11104700 TI - A carbon-source-responsive element is required for regulation of the hypoxic ADP/ATP carrier (AAC3) isoform in Saccharomyces cerevisiae. AB - The mitochondrial ADP/ATP carrier in Saccharomyces cerevisiae is encoded by three genes that are differentially expressed under different physiological conditions. We investigated the transcriptional control of AAC3, an oxygen-repressed isoform. By deletion analysis, DNA electrophoretic mobility-shift assays, DNase I footprinting and site-directed mutagenesis, we have identified a promoter region (upstream repressing sequence 1, URS(1)) involved in a carbon-source-dependent repression of AAC3. It is different from the previously characterized oxygen dependent ROX1 (regulation by oxygen 1) repressor-binding region (URS(2)). The complex character of URS(1) includes the presence of two different cis-acting sequences: (i) a RAP1 (repressor activator protein 1)-binding site that is capable of binding the RAP1 protein in vitro and (ii) two putative ethanol repression sequences, the modification of which derepresses the AAC3 gene. These findings demonstrate that the hypoxic AAC3 gene is regulated by two upstream repressor sites; one controlled by oxygen and haem, the other by the carbon source. Both sites function to completely switch off the expression of the AAC3 isoform when ATP is made by oxidative phosphorylation, and they modulate AAC3 expression when import of glycolytic ATP into mitochondria is required. PMID- 11104701 TI - Use of alpha-toxin from Staphylococcus aureus to test for channelling of intermediates of glycolysis between glucokinase and aldolase in hepatocytes. AB - We investigated whether hepatocytes permeabilized with alpha-toxin from Staphylococcus aureus are a valid model for studying the channelling of intermediates of glycolysis between glucokinase and triosephosphate isomerase. These cells are permeable to 2-aminoisobutyrate, ATP, glucose 6-phosphate (Glc6P) and fructose 2, 6-bisphosphate [Fru(2,6)P(2)], but maintain cell integrity in the presence of ATP as judged by the retention of cytoplasmic enzymes. During incubation with 25 mM glucose, an ATP-generating system and saturating concentrations of Fru(2,6)P(2), rates of detritiation of [2-(3)H]glucose and [3 (3)H]glucose were similar. Exogenous Glc6P (1 mM) and to a lesser extent fructose 6-phosphate, but not Fru(1, 6)P(2), decreased the rate of detritiation of [3 (3)H]glucose. During incubation with 25 mM glucose and Glc6P (0.2-1 mM), with either [3-(3)H]glucose or [3-(3)H]Glc6P as labelled substrate, there was dilution of metabolism of [3-(3)H]glucose with increasing Glc6P but no overall increase in glycolytic flux from glucose and Glc6P, indicating that glycolysis is apparently saturated with Glc6P despite the permeability of the cells to this metabolite. These findings could be explained by partial channelling of Glc6P between glucokinase and glycolysis in the presence of saturating concentrations of Fru(2,6)P(2). They provide an alternative explanation for the concept that there is more than one Glc6P pool. PMID- 11104702 TI - Coexpression of alpha and beta subunits of prolyl 4-hydroxylase stabilizes the triple helix of recombinant human type X collagen. AB - We have reported previously on the expression of recombinant human type X collagen (hrColX) in HEK 293 and HT 1080 cells by using the eukaryotic expression vector pCMVsis (in which CMV stands for cytomegalovirus). Several stably transfected clones secreted full-length triple-helical hrColX molecules in large amounts, but the secreted collagen was underhydroxylated, with a hydroxyproline to-proline ratio of 0.25 and a melting temperature (T(m)) of 31 degrees C. By comparison, native chicken type X procollagen has a T(m) of 46 degrees C. To stabilize the triple helix of hrColX, an hrColX-expressing clone (A6/16) was co transfected with both alpha and beta subunits of human prolyl 4-hydroxylase. Clones were selected that secreted proalpha1(X) collagen chains with an apparent molecular mass of 75 kDa and an increased hydroxyproline-to-proline ratio of close to 0.5. As a result of enhanced prolyl hydroxylation, the T(m) of the hrColX was increased to 41 degrees C as measured by CD analysis at various temperatures. The CD spectra indicated a minimum ellipticity at 198 nm and a peak at 225 nm at 20 degrees C, confirming the presence of a triple helix. The same T(m) of 41 degrees C was measured for the triple-helical core fragments of hrColX of 60-65 kDa that were retained after brief digestion with chymotrypsin/trypsin at increasing temperatures. This shows that the human cell line HEK-293 is suitable for the simultaneous expression of three genes and the stable production of substantial amounts of recombinant, fully hydroxylated type X collagen over several years. PMID- 11104704 TI - Effects of overexpression of the liver subunit of 6-phosphofructo-1-kinase on the metabolism of a cultured mammalian cell line. AB - Overexpression of the liver subunit of 6-phosphofructo-1-kinase in Chinese hamster ovary K1 cells was shown to increase the steady-state level of the enzyme's product, fructose 1, 6-bisphosphate, and to produce a small but significant decrease in the concentration of fructose 2,6-bisphosphate, which is an allosteric activator of the enzyme. However, overexpression of the enzyme had no effect on glycolytic flux under a variety of different substrate conditions. This latter observation is consistent with similar studies in fungi and in potato tubers which indicate that 6-phosphofructo-1-kinase has very little control over flux in glycolysis. PMID- 11104703 TI - Cross-talk between interleukin 1beta (IL-1beta) and IL-6 signalling pathways: IL 1beta selectively inhibits IL-6-activated signal transducer and activator of transcription factor 1 (STAT1) by a proteasome-dependent mechanism. AB - Interleukin 1beta (IL-1beta) suppresses the IL-6-dependent induction of type II acute-phase response genes, but the underlying mechanism for this suppression remains uncertain. Here we report that treatment of human hepatocullular carcinoma HepG2 cells with IL-1beta inhibited the IL-6-dependent binding of signal transducer and activator of transcription factor (STAT)1, but not that of STAT3, to the high-affinity serum-inducible element ('SIE'). Furthermore, IL 1beta selectively down-regulated the IL-6-induced tyrosine phosphorylation of STAT1 without affecting the level of STAT1 or tyrosine phosphorylation of STAT3. Kinase assays in vitro indicated that the inhibition of STAT1 phosphorylation by IL-1beta was not due to an upstream blockade of Janus kinase (JAK1 or JAK2) activation. However, pretreatment with the proteasome inhibitor MG132 under conditions that prevented the IL-1beta-dependent activation of the nuclear factor NF-kappaB also blocked the inhibitory effect of IL-1beta on IL-6-activated STAT1. In related experiments, the protein tyrosine phosphatase inhibitor Na(3)VO(4) also antagonized the inhibitory effect of IL-1beta on the activation of STAT1 by IL-6. Taken together, these findings indicate that, by using a proteasome dependent mechanism, IL-1beta concomitantly induces NF-kappaB activation and dephosphorylates IL-6-activated STAT1; the latter might partly account for the inhibition by IL-1beta of the IL-6-dependent induction of type II acute-phase genes. PMID- 11104705 TI - Rapid activation and partial inactivation of inositol trisphosphate receptors by adenophostin A. AB - Adenophostin A, the most potent known agonist of inositol 1,4, 5-trisphosphate (InsP(3)) receptors, stimulated (45)Ca(2+) release from the intracellular stores of permeabilized hepatocytes. The concentration of adenophostin A causing the half-maximal effect (EC(50)) was 7.1+/-0.5 nM, whereas the EC(50) for InsP(3) was 177+/-26 nM; both responses were positively co-operative. In rapid superfusion analyses of (45)Ca(2+) release from the intracellular stores of immobilized hepatocytes, maximal concentrations of adenophostin A or InsP(3) evoked indistinguishable patterns of Ca(2+) release. The Ca(2+) release evoked by both agonists peaked at the same maximal rate after about 375 ms and the activity of the receptors then decayed to a stable, partially (60%) inactivated state with a half-time (t(1/2)) of 318+/-29 ms for adenophostin A and 321+/-22 ms for InsP(3). Dissociation rates were measured by recording rates of InsP(3)-receptor channel closure after rapid removal of agonist. The rate of adenophostin A dissociation (t(1/2), 840+/-195 ms) was only 2-fold slower than that of InsP(3) (t(1/2), 436+/ 48 ms). We conclude that slow dissociation of adenophostin A from InsP(3) receptors does not underlie either its high-affinity binding or the reported differences in the Ca(2+) signals evoked by InsP(3) and adenophostin A in intact cells. PMID- 11104706 TI - Learned discrimination of pattern orientation in walking flies. AB - To determine the pattern-orientation discrimination ability of blowflies, Phaenicia sericata, a learning/memory assay was developed in which sucrose served as the reward stimulus and was paired with one of two visual gratings of different orientations. Individual, freely walking flies with clipped wings were trained to discriminate between pairs of visual patterns presented in the vertical plane. During training trials, individual flies learned to search preferentially at the rewarded stimulus. In subsequent testing trials, flies continued to exhibit a learned preference for the previously rewarded stimulus, demonstrating an ability to discriminate between the two visual cues. Flies learned to discriminate between horizontal and vertical gratings, +45 degrees (relative to a 0 degrees vertical) and -45 degrees gratings, and vertical and +5 degrees gratings. Individual patterns of learning and locomotive behavior were observed in the pattern of exploration during training trials. The features of the visual cue critical for discrimination of orientation are discussed. PMID- 11104707 TI - Orientation discrimination independent of retinal matching by blowflies. AB - Blowflies, Phaenicia sericata, can be trained to discriminate in a learning paradigm in which one of the two visual cues is positively rewarded. Retinotopic matching of a learned visual image to the same retinal location from viewing to viewing has been hypothesized to underlie visual pattern learning and memory in insects. To address the theory of retinotopic matching, a detailed analysis was made of the flies' body orientations during learned discriminations between +45 degrees and -45 degrees gratings. Initial approaches to the positive rewarded visual cue did not originate from the same spatial location within the behavioral arena with respect to the visual cues; thus, individual flies approached the positive cue from a different vantage point from trial to trial. During initial approaches to the rewarded visual cue, the distributions of body angles with respect to the cue were different from trial to trial for each individual. These data suggest that Phaenicia sericata can learn a visual pattern with one eye region and later recognize the same pattern with another eye region. Thus, retinotopic matching is not necessary for the recognition of pattern orientation in the experimental paradigm used here. The average amount of head turning in the yaw plane was too small to compensate for the changes in body orientation exhibited by the flies. Flies view the visual patterns with distinct retinal regions from trial to trial during orientation discrimination. PMID- 11104708 TI - Molecular characterization of V-type H(+)-ATPase (B-subunit) in gills of euryhaline crabs and its physiological role in osmoregulatory ion uptake. AB - The vacuolar-type H(+)-ATPase (V-ATPase) has been implicated in osmoregulatory ion uptake across external epithelia of a growing variety of species adapted to life in fresh water. In the present study, we investigated whether the V-ATPase may also function in a euryhaline species that tolerates brackish water (8 salinity) but not fresh water, the shore crab Carcinus maenas. cDNA coding for the regulatory B-subunit of the V-ATPase was amplified and sequenced from C. maenas gills and partially sequenced from four other crab species. Two isoforms differing in the 3'-untranslated region were found in C. maenas. In this species, the abundance of B-subunit mRNA was greater in the respiratory anterior gills than the ion-transporting posterior gills and was not increased by acclimation to dilute salinity. Immunocytochemical analysis showed that the B-subunit protein is not targeted to the apical membrane but is distributed throughout the cytoplasmic compartment. Physiological studies of isolated perfused gills indicated that the V-ATPase inhibitor bafilomycin had no effect on transepithelial potential difference. Thus, in contrast to the freshwater-tolerant Chinese crab Eriocheir sinensis, in which the V-ATPase appears to play an important osmoregulatory role, the V-ATPase in C. maenas probably functions in acidification of intracellular organelles but not in transbranchial NaCl uptake. PMID- 11104709 TI - Acute and chronic influence of temperature on red blood cell anion exchange. AB - Unidirectional (36)Cl(-) efflux via the red blood cell anion exchanger was measured under Cl(-) self-exchange conditions (i.e. no net flow of anions) in rainbow trout Oncorhynchus mykiss and red-eared freshwater turtle Trachemys scripta to examine the effects of acute temperature changes and acclimation temperature on this process. We also evaluated the possible adaptation of anion exchange to different temperature regimes by including our previously published data on other animals. An acute temperature increase caused a significant increase in the rate constant (k) for unidirectional Cl(-) efflux in rainbow trout and freshwater turtle. After 3 weeks of temperature acclimation, 5 degrees C-acclimated rainbow trout showed only marginally higher Cl(-) transport rates than 15 degrees C-acclimated trout when compared at the same temperature. Apparent activation energies for red blood cell Cl(-) exchange in trout and turtle were lower than values reported in endothermic animals. The Q(10) for red blood cell anion exchange was 2.0 in trout and 2.3 in turtle, values close to those for CO(2) excretion, suggesting that, in ectothermic animals, the temperature sensitivity of band-3-mediated anion exchange matches the temperature sensitivity of CO(2) transport (where red blood cell Cl(-)/HCO(3)(-) exchange is a rate-limiting step). In endotherms, such as man and chicken, Q(10) values for red blood cell anion exchange are considerably higher but are no obstacle to CO(2) transport, because body temperature is normally kept constant at values at which anion exchange rates are high. When compared at constant temperature, red blood cell Cl(-) permeability shows large differences among species (trout, carp, eel, cod, turtle, alligator, chicken and man). Cl(-) permeabilities are, however, remarkable similar when compared at preferred body temperatures, suggesting an appropriate evolutionary adaptation of red blood cell anion exchange function to the different thermal niches occupied by animals. PMID- 11104710 TI - Free vertical moments and transverse forces in human walking and their role in relation to arm-swing. AB - We present force plate data on vertical free moments (force couples in the horizontal plane between the foot and the ground) and on transverse force during unloaded walking in different modes and at different speeds (including running) by adults of both sexes and by children, and examine loaded walking by adult males and one boy. Free moments in slow and normal-speed walking are characterised by a lateral peak in the accelerative phase of stance, but the peak during running, and in some cases of fast walking, occurs in the deceleration phase. Free moments are strongly affected by arm fixation in males, but less so in females. The pattern, but not the scale, of free moments is affected by loading position and side, but load magnitude has little effect if the loaded weight is treated as part of the body. Transverse force is more variable than sagittal force. In males, the transverse force curve shows a marked trough at mid stance, whereas in females this trough is rarely seen. The transverse force of males also differs from that of females in response to arm fixation, showing a local medial inflection at three-quarters of the stance phase that is not present in females. Adults differ from children younger than 9 years in the presence of a very short, medially directed peak following heel-strike. Analysis of the effects of arm fixation and the timing of forces suggests strongly that arm-swing and free moments tend to reinforce each other in balancing trunk torques induced by the lower limbs. Both are of reduced importance in slow walking. PMID- 11104711 TI - Contractile properties of the elasmobranch rectal gland. AB - The importance of the rectal gland in elasmobranch osmoregulation is well established. The rate of secretion by the gland is under the control of a variety of secretagogues and inhibitors. Early morphological work suggested that a band of smooth muscle cells surrounds the periphery of the shark rectal gland between the secretory tubules and the connective tissue capsule. To confirm the presence of the muscle ring, we examined histological sections from two species of shark, Squalus acanthias and Carcharodon carcharius, and from the stingray Dasyatis sabina and stained sections from S. acanthias with the actin-specific ligand phalloidin. In all three species, a distinct band of what appeared to be smooth muscle cells was evident, and the putative muscle ring in S. acanthias stained specifically with phalloidin. Moreover, isolated rings of rectal gland tissue from S. acanthias constricted when acetylcholine or endothelin was applied and responded to nitric oxide with an initial dilation, followed by a more substantial constriction. Subsequent addition of porcine C-type natriuretic peptide dilated the rings, but two prostanoids (carbaprostacyclin and prostaglandin E(1)) did not change ring tension significantly. The rings did not respond to the endothelin-B-specific agonist sarafotoxoin S6c, suggesting that the response to endothelin was mediated via endothelin-A-type receptors. Our data confirm the presence of a smooth muscle ring in the periphery of the elasmobranch rectal gland and demonstrate that the gland responds to a suite of smooth muscle agonists, suggesting that changes in the dimensions of the whole rectal gland may play a role in its secretory function. PMID- 11104712 TI - Physical factors affecting the cost and efficiency of sound production in the treefrog Hyla versicolor. AB - The metabolic cost, energy output and efficiency (i.e. the ratio of energy output to metabolic cost) of sound production were compared among male grey treefrogs (Hyla versicolor) as a function of body size and temperature. The effects of call length (in notes per call) and dominant frequency (in kHz) were also considered. Cost, determined from the amount of oxygen consumed, averaged 12.1 mJ per note and was dependent only upon body mass. Acoustic energy per note, determined from oscillograms of recorded calls, averaged 0.34 mJ and was dependent only upon temperature. Conventional theory suggests that the efficiency of sound production should be a function of the ratio of the linear size of the radiating structures to the wavelength of the sound generated (i.e. efficiency is assumed to be a function of the product of mass(0.33) and frequency), but efficiency in H. versicolor was found to be a function of the product of temperature(2.1) and mass(-1.08). Adjusting for temperature and body mass, the efficiency of sound production in H. versicolor (average 2.4 %) is greater than the efficiency of other frog species for which data are available. Temperature may affect acoustic energy output because trunk muscle contraction speed increases with temperature, which increases the velocity of airflow across the vocal cords. PMID- 11104713 TI - The boundary layer of swimming fish. AB - Tangential and normal velocity profiles of the boundary layer surrounding live swimming fish were determined by digital particle tracking velocimetry, DPTV. Two species were examined: the scup Stenotomus chrysops, a carangiform swimmer, and the smooth dogfish Mustelus canis, an anguilliform swimmer. Measurements were taken at several locations over the surfaces of the fish and throughout complete undulatory cycles of their propulsive motions. The Reynolds number based on length, Re, ranged from 3x10(3) to 3x10(5). In general, boundary layer profiles were found to match known laminar and turbulent profiles including those of Blasius, Falkner and Skan and the law of the wall. In still water, boundary layer profile shape always suggested laminar flow. In flowing water, boundary layer profile shape suggested laminar flow at lower Reynolds numbers and turbulent flow at the highest Reynolds numbers. In some cases, oscillation between laminar and turbulent profile shapes with body phase was observed. Local friction coefficients, boundary layer thickness and fluid velocities at the edge of the boundary layer were suggestive of local oscillatory and mean streamwise acceleration of the boundary layer. The behavior of these variables differed significantly in the boundary layer over a rigid fish. Total skin friction was determined. Swimming fish were found to experience greater friction drag than the same fish stretched straight in the flow. Nevertheless, the power necessary to overcome friction drag was determined to be within previous experimentally measured power outputs. No separation of the boundary layer was observed around swimming fish, suggesting negligible form drag. Inflected boundary layers, suggestive of incipient separation, were observed sporadically, but appeared to be stabilized at later phases of the undulatory cycle. These phenomena may be evidence of hydrodynamic sensing and response towards the optimization of swimming performance. PMID- 11104714 TI - Way-finding and landmarks: the multiple-bearings hypothesis. AB - Clark's nutcrackers (Nucifraga columbiana) are capable of very precise searching using the metric relationships between a goal and multiple landmarks to relocate the goal location. They can judge the direction more accurately than the distance to a landmark when the landmark is distant from the goal. On the basis of these findings, we propose that nutcrackers use a set of bearings, each a measure of the direction from the goal to a different landmark, when searching for that goal. The results of a simulation demonstrate that increasing the number of landmarks used results in increasingly precise searching. This multiple-bearings hypothesis makes a series of detailed predictions about how the distribution of searches will vary as a function of the geometry of the locations of the relevant landmarks and the goal. It also suggests an explanation for inconsistencies in the literature on the effects of clock-shifts on searching and on homing. PMID- 11104715 TI - Cardiovascular effects of hypercarbia in rainbow trout (Oncorhynchus mykiss): a role for externally oriented chemoreceptors. AB - In situ and in vivo experiments were performed on rainbow trout (Oncorhynchus mykiss) to examine (i) the direct effect of CO(2) on the systemic vasculature and (ii) the influence of internal versus external hypercapnic acidosis on cardiovascular variables including blood pressure, cardiac output and systemic vascular resistance. Results from in situ saline-perfused trunk preparations indicated that CO(2) (0.6, 1.0 or 2.0% CO(2)) elicited a significant vasodilation, but only in the presence of pre-existing humoral adrenergic tone. In the absence of pre-existing vascular tone, CO(2) was without effect on systemic resistance. In contrast, hypercarbia in vivo triggered a statistically significant increase in systemic resistance (approximately 70 %) that was associated with elevated ventral aortic (approximately 42 %) and dorsal aortic (approximately 43 %) blood pressures and with a significant bradycardia (approximately 12 %); cardiac output was not significantly affected. To determine the potential roles of internal versus external chemoreceptors in mediating the cardiovascular responses to hypercarbia, experiments were performed to elevate the endogenous arterial partial pressure of CO(2) (Pa(CO2)) without an accompanying increase in external P(CO2) (Pw(CO2)). In one series, trout were given a bolus injection of the carbonic anhydrase inhibitor acetazolamide (30 mg kg(-1)) to inhibit CO(2) excretion, and thus raise Pa(CO2), 5-7 h prior to being exposed to an acute increase in Pw(CO2) (maximum Pw(CO2)=6.3+/-0.4 mmHg; 1 mmHg=0.133 kPa). Despite a marked increase in Pa(CO2) (approximately 7 mmHg) after injection of acetazolamide, there was no increase in dorsal aortic blood pressure (P(DA)) or systemic resistance (R(S)). The ensuing exposure to hypercarbia, however, significantly increased P(DA) (by approximately 20 %) and R(S) (by approximately 35 %). A second series of experiments used a 5-7 h period of exposure to hyperoxia (Pw(O2)=643+/-16 mmHg) to establish a new, elevated baseline Pa(CO2) (7.8+/-1.1 mmHg) without any change in Pw(CO2). Despite a steadily increasing Pa(CO2) during the 5-7 h of hyperoxia, there was no associated increase in P(DA) or R(S). Ensuing exposure to hypercarbia, however, significantly increased P(DA) (by approximately 20 %) and R(S) (by approximately 150 %). Plasma adrenaline levels were increased significantly during exposure to hypercarbia and, therefore, probably contributed to the accompanying cardiovascular effects. These findings demonstrate that the cardiovascular effects associated with hypercarbia in rainbow trout are unrelated to any direct constrictory effects of CO(2) on the systemic vasculature and are unlikely to be triggered by activation of internally oriented receptors. Instead, the data suggest that the cardiovascular responses associated with hypercarbia are mediated exclusively by externally oriented chemoreceptors. PMID- 11104716 TI - Intrinsic noise at synapses between a wing hinge stretch receptor and flight motor neurons in the locust. AB - Variability in postsynaptic potential (PSP) amplitude due to intrinsic noise limits the reliability of communication between neurons. I measured PSP variability at synapses between a forewing stretch receptor and wing depressor motor neurons in locusts, a pathway that is important in the control of flying. The intrinsic noise in the stretch receptor output synapse was measured by subtracting the background noise, originating in other synaptic pathways onto the motor neuron, from the variability in the amplitudes of PSPs evoked by the stretch receptor. Intrinsic synaptic noise caused successive PSPs to vary by 4-10 % in basalar and subalar flight motor neurons. Recordings from pairs of these wing depressor motor neurons showed that the amount of transmitter released varied independently between different output sites from the stretch receptor. Histograms of excitatory postsynaptic potential amplitude were normal distributions that lacked separate peaks. I estimate that quantal amplitude is significantly less than 0.1 mV and that several hundred quanta are released for each presynaptic spike. This accords well with a previous estimate of the number of discrete anatomical synapses and would facilitate modulation of output from the stretch receptor. PMID- 11104717 TI - Similarity in flight behaviour between the honeybee Apis mellifera (Hymenoptera: apidae) and its presumed mimic, the dronefly Eristalis tenax (Diptera: syrphidae). AB - It is generally accepted that the dronefly Eristalis tenax is a Batesian mimic of the honeybee Apis mellifera. Previous work has established that the foraging behaviour of droneflies is more similar to that of its model than to that of other more closely related flies, suggesting that behaviour may be important in the mimicry. Locomotor mimicry has been demonstrated in mimetic Heliconius butterflies but not in hoverflies. This study therefore investigated aspects of the flight behaviour of Eristalis tenax, Apis mellifera and two other flies, Syrphus ribesii and a Musca sp. Insects were filmed foraging on Helichrysum bracteum flowers, and flight sequences were analysed to determine flight velocities, flight trajectories and the percentage of time spent hovering. It was found that the flight behaviour of droneflies was more similar to that of honeybees than to that of the other flies. This suggests that the flight behaviour of Eristalis tenax may be mimetic. PMID- 11104718 TI - Identification of a sea urchin Na(+)/K(+)/2Cl(-) cotransporter (NKCC): microfilament-dependent surface expression is mediated by hypotonic shock and cyclic AMP. AB - We report the identification of an invertebrate Na(+)/K(+)/2Cl(-) cotransporter, NKCC. As a model system, we used the immune cells (coelomocytes) of the Mediterranean sea urchin Paracentrotus lividus. These cells are particularly interesting because they can be activated to undergo a rapid and dynamic change in cell shape. We demonstrate that forskolin, a cyclic AMP agonist known to regulate NKCC, induced coelomocyte transformation at doses of 10 micromol l(-)(1) and greater. Using two distinct monoclonal antibodies (T4 and T9) raised against the human intestinal epithelial NKCC, we have identified a high-molecular-mass (195 kDa) protein in coelomocyte extracts. We propose a novel method for the isolation of NKCC in one step by using bumetanide-Sepharose affinity chromatography under low-[Cl(-)] conditions. This method was successful in isolating coelomocyte 195 kDa NKCC. The T4 monoclonal antibody was used in immunocytochemical experiments to localize NKCC in resting and activated coelomocytes. In petalloid coelomocytes, a punctate, cytoplasmic distribution was observed in close proximity to actin filament bundles; in transformed coelomocytes, the immunofluorescence was distributed along the length of the filopodia and uniformly throughout the perinuclear region. The change in subcellular distribution of NKCC between the resting and the activated state was further investigated by using cell surface biotinylation followed by immunoprecipitation. These studies revealed an upregulation of NKCC at the plasma membrane upon activation, a process that was blocked by the F-actin-stabilizing drug phalloidin. These studies identify a novel model system in which to investigate a newly identified invertebrate Na(+)/K(+)/2Cl(-) cotransporter. PMID- 11104719 TI - Oxidative stress, DNA damage and p53 expression in the larvae of atlantic cod (Gadus morhua) exposed to ultraviolet (290-400 nm) radiation. AB - Decreases in stratospheric ozone levels from anthropogenic inputs of chlorinated fluorocarbons have resulted in an increased amount of harmful ultraviolet-B (UVB, 290-320 nm) radiation reaching the sea surface in temperate latitudes (30-50 degrees N). In the Gulf of Maine, present-day irradiances of ultraviolet-A (UVA, 320-400 nm) radiation can penetrate to depths of 23 m and UVB radiation can penetrate to depths of 7-12 m, where the rapidly developing embryos and larvae of the Atlantic cod (Gadus morhua) are known to occur. Laboratory exposures of embryos and larvae of Atlantic cod to ultraviolet radiation (UVR) equivalent to a depth of approximately 10 m in the Gulf of Maine resulted in significant mortality of developing embryos and a decrease in standard length at hatching for yolk-sac larvae. Larvae at the end of the experimental period also had lower concentrations of UVR-absorbing compounds and exhibited significantly greater damage to their DNA, measured as cyclobutane pyrimidine dimer formation, after exposure to UVB radiation. Larvae exposed to UVB radiation also exhibited significantly higher activities and protein concentrations of the antioxidant enzyme superoxide dismutase and significantly higher concentrations of the transcriptional activator p53. p53 is expressed in response to DNA damage and can result in cellular growth arrest in the G1- to S-phase of the cell cycle or to programmed cell death (apoptosis). Cellular death caused by apoptosis is the most likely cause of mortality in embryos and larvae in these laboratory experiments, while the smaller size at hatching in those larvae that survived is caused by permanent cellular growth arrest in response to DNA damage. In addition, the sub lethal energetic costs of repairing DNA damage or responding to oxidative stress may also contribute to poor individual performance in surviving larvae that could also lead to increases in mortality. The irradiances of UVB radiation that elicit these responses in cod larvae can occur in many temperate latitudes, where these ecologically and commercially important fish are known to spawn, and may contribute to the high mortality of cod embryos and larvae in their natural environment. PMID- 11104720 TI - Allometric relationships between embryonic heart rate and fresh egg mass in birds. AB - Previously, we have measured daily changes (developmental patterns) in embryonic heart rate (fh) in altricial and semi-altricial (ASA) birds (range of mean fresh egg mass approximately 1-20 g), semi-precocial seabirds (egg mass approximately 38-288 g) and precocial birds (egg mass approximately 6-1400 g). An allometric relationship between embryonic fh at 80 % of incubation duration (ID) and fresh egg mass (M) has been derived for six species of precocial bird (fh at 80 % ID=429M(-0.118)). In the present study, additional measurements of embryonic fh in three ASA species, the barn owl Tyto alba, the cattle egret Bubulcus ibis and the lanner falcon Falco biarmicus, were made to extend the egg mass range (20-41 g), and the allometric relationships of embryonic fh for these ASA birds and the precocial and semi-precocial (PSP) groups were investigated from published data. The developmental patterns of embryonic fh in three relatively large ASA species did not show a significant increase prior to the pipping period, unlike those in small ASA birds, but tended to be constant, with a subsequent increase during pipping. The allometric relationship derived for ASA birds was fh at 80 % ID=371M(-0.121) (r=-0.846, P<0.001, N=20) and that for PSP birds was fh at 80 % ID=433M(-0.121) (r=-0.963, P<0.001, N=13). The slopes were parallel, but fh of ASA embryos was low compared with that of PSP embryos with the same egg mass. In ASA birds, embyronic fh was maximal during the pipping (perinatal) period, and the maximum fh (fh(max)) was significantly related to fresh egg mass: fh(max)=440M(-0.127) (r=-0.840, P<0.001, N=20). The allometric relationships for fh at 80 % ID in PSP and fh(max) in ASA embryos were statistically identical. Accordingly, embryonic fh at 80 % ID in PSP birds and fh(max) during pipping in ASA birds can be expressed by a single allometric equation: fh=437M(-0.123) (r= 0.948, P<0.001, N=33). PMID- 11104721 TI - Bioinformatics. A user's perspective. AB - This review provides an overview of bioinformatics from the user's point of view. Bioinformatics, defined as the application of computers, databases, and computational methods to the management of biologic information, is essential for almost every aspect of data management in modern biology. The rapid accumulation of genomic sequence information together with the wide availability of new technologies that analyze global gene expression patterns have created an information overload. Molecular biology labs are increasingly dependent on computers, large-capacity databases, search and analysis tools, and high-quality Internet connections. Currently available bioinformatics tools are discussed and a general approach is outlined. Using the resources and approaches in this review, readers should be able to form their own view of bioinformatics and tailor the solutions to the information overload according to their needs. PMID- 11104722 TI - Dislocation of E-cadherin in the airway epithelium during an antigen-induced asthmatic response. AB - The airway epithelium plays a critical role in asthma. E-cadherin, located on the basolateral side of the epithelial cells, forms adherent junctions. To investigate the role of E-cadherin on the regulation of permeability of molecules and fluid in asthmatic responses, we observed the dynamics of E-cadherin after an immunochallenge against guinea pigs. Immunohistochemical studies revealed that E cadherin was expressed on the lateral sides of epithelial cells before the immunochallenge and after immediate airway responses (IAR). However, E-cadherin immunoreactivities decreased from the basolateral region in late airway responses (LAR) 6 h after the challenge. Simultaneously, soluble E-cadherin immunoreactivities were detected in lavage fluid only in LAR, suggesting that E cadherin is partly cleaved and released into the lumen in LAR. Airway permeability, which was examined by penetration of horseradish peroxidase from the airway side into the epithelium, increased in both IAR and LAR. These results suggest that E-cadherin detachment from the lateral side of the epithelial cells increased airway permeability in LAR but not IAR. We conclude that an antigen challenge causes an opening of adherent junctions as well as increases airway permeability in LAR. This mechanism would participate in airflow limitation during attacks and the increase of airway permeability and hyperresponsiveness in asthmatics. PMID- 11104723 TI - Monocyte chemoattractant protein-1 and RANTES are chemotactic for graft infiltrating lymphocytes during acute lung allograft rejection. AB - Graft infiltrating lymphocytes (GILs) are crucial to rejection of lung allografts. However, chemotactic activities, chemokines responsible for GIL recruitment, and cells involved in chemokine production during lung allograft rejection have not been evaluated. This study determined whether chemotactic activity for GILs is upregulated, and whether the chemokines monocyte chemoattractant protein (MCP)-1 and regulated on activation, normal T cells expressed and secreted (RANTES) have roles in GIL chemotaxis during lung allograft rejection. F344 (RT1(lv1)) rat lung allografts were transplanted into WKY (RT1(l)) recipients. Chemotactic activity for GILs and quantities of MCP-1 and RANTES were determined in allograft bronchoalveolar lavage fluid 1 wk after transplantation. Data showed that during rejection, chemotactic activity for GILs is upregulated, MCP-1 and RANTES are produced locally, and both MCP-1 and RANTES are operative in GIL recruitment. Immunohistochemistry showed that alveolar macrophages (AMs) were the major source of MCP-1 and that other lung cells, including AMs, were the source of RANTES. Further, depletion of AMs in the donor lung before transplantation downregulated chemotaxis for GILs and production of MCP-1 during rejection episodes. These data show that chemotaxis for GILs is upregulated locally during lung allograft rejection, and that MCP-1 and RANTES contribute to GIL recruitment during the rejection response. PMID- 11104724 TI - Modulation of alveolar macrophage phagocytosis by leukotrienes is Fc receptor mediated and protein kinase C-dependent. AB - We have previously established an important role for leukotrienes (LTs) in augmenting rat alveolar macrophage (AM) phagocytosis of Klebsiella pneumoniae opsonized with complete immune serum (IS), which contains the two well-known opsonins, immunoglobulin (Ig) G and complement (C). In this report, the specific opsonin requirements for LT modulation of AM phagocytosis and the dependence of this response on protein kinase (PK) C activity were investigated. Phagocytosis of K. pneumoniae opsonized with IS, non-immune serum, or heat-inactivated immune serum and of inert targets (IgG-opsonized fluorescent microspheres or C-opsonized sheep red blood cells) was examined. Inhibition of endogenous LT synthesis or action attenuated, whereas the addition of exogenous LTs augmented, phagocytosis only of targets opsonized with IgG. LTs had no effect on phagocytosis of C opsonized or unopsonized targets. LTs did not affect adherence of IgG-opsonized targets, implying instead an enhancement of internalization. Macrophage internalization of phagocytic targets has previously been shown to require PKC activity. Pretreatment of AMs with the PKC inhibitors staurosporine or calphostin C, or with phorbol 12-myristate 13-acetate to deplete PKC, completely inhibited the ability of LTB(4) and largely inhibited the ability of LTC(4) to augment phagocytosis of IgG-opsonized microspheres. These results demonstrate that LT enhancement is confined to Fc receptor (FcR)-mediated phagocytosis. Moreover, PKC activation represents an important mechanism by which LTs promote FcR-mediated phagocytosis. PMID- 11104725 TI - Characterization of an axonemal dynein heavy chain expressed early in airway epithelial ciliogenesis. AB - The most conspicuous evidence of airway epithelial maturation and vitality is the presence of motile cilia. In an effort to generate genetic and antigenic markers of airway maturation, injury, and repair, we characterized airway epithelial expression of a gene identified by two human expressed sequence tags that encoded peptides with sequence similarity to an invertebrate ciliary dynein heavy chain (DHC). Molecular analyses showed that the gene has a very large RNA transcript that encodes a very high molecular weight polypeptide with biochemical properties that are characteristic of a dynein heavy chain. Expression of the gene transcript correlated with the presence of ciliated cells in tissues, and immunohistochemical localization of the gene product confirmed its presence in the cilia of mature airway epithelium. In epithelium undergoing ciliogenesis ex vivo, expression of the gene transcript preceded ciliation of the epithelium and the gene product was present in the cytoplasm and at the apical border of nonciliated cells. These data suggested that the gene encodes an axonemal DHC that is expressed early during ciliogenesis, before the appearance of cilia. PMID- 11104726 TI - Expression patterns of laminin alpha1 and alpha5 in human lung during development. AB - Laminins are trimeric glycoprotein components of basement membranes. Each laminin has three structurally similar chains, designated alpha, beta, and gamma. Five laminin alpha chains are now known. In previous studies using monoclonal antibody 4C7, laminin alpha1 was thought to be present in basement membranes of human lung throughout development and in the adult, but recent expression studies have demonstrated that 4C7 identifies laminin alpha5 rather than alpha1. To determine the temporal and spatial patterns of laminin alpha1 and laminin alpha5 in developing human lung, we prepared complementary DNA probes specific for laminin alpha1 and alpha5 messenger RNAs (mRNAs). By Northern analysis, laminin alpha1 mRNA was prominent in first-trimester fetal lung, but was not detectable at 23 wk or at later times. In contrast, laminin alpha5 mRNA was readily detected in early fetal lung and remained present thereafter. Immunohistochemical staining demonstrated laminin alpha1 only in early fetal lung, whereas laminin alpha5 was persistent from the early fetal period. In situ hybridization localized laminin alpha1 expression to distal epithelium in the first-trimester lung, and laminin alpha5 to all epithelium and developing pulmonary arteries from the first trimester through the perinatal period. These studies indicate that laminin alpha1 expression is restricted to early human lung morphogenesis, whereas the expression of laminin alpha5 in human lung is continuous from early lung development through adult life. It is evident that laminin alpha1 and laminin alpha5 have different roles in the development of the human lung. PMID- 11104727 TI - Phospholipase D and priming of the respiratory burst by H(2)O(2) in NR8383 alveolar macrophages. AB - Previous investigation showed that preincubation within a range of nontoxic H(2)O(2) concentrations enhanced subsequently stimulated superoxide production by rat alveolar macrophages in response to various stimuli. In the present study, the NR8383 rat alveolar macrophage cell line was used to further investigate the priming effect of H(2)O(2). Using nitroblue tetrazolium, which formed an insoluble formazan when reduced by superoxide, modulation of the respiratory burst was visualized in a cell population exposed to a concentration gradient of H(2)O(2) before stimulation. This model system illustrates how H(2)O(2) may constitute a signaling molecule for a feed-forward regulation of the respiratory burst during inflammation. n-Butanol, which allows consumption of phosphatidic acid by the transphosphatidylation reaction, and propanolol, which inhibits phosphatidic acid phosphohydrolase, were used to investigate the possible involvement of phospholipase D in this phenomenon. These two agents were found to inhibit the basal adenosine diphosphate-stimulated respiratory burst. Inhibition of the H(2)O(2)-enhanced respiratory burst was equally or slightly less effective when expressed as percentage of controls. Furthermore, phospholipase D was not activated by H(2)O(2) concentrations that enhance superoxide production. Thus, phospholipase D does not mediate the enhancement of the respiratory burst by H(2)O(2), although it may be activated by high concentrations of this hydroperoxide. PMID- 11104728 TI - Inhibition of amiloride-sensitive epithelial Na(+) absorption by extracellular nucleotides in human normal and cystic fibrosis airways. AB - Cystic fibrosis (CF) airway epithelia are characterized by enhanced Na(+) absorption probably due to a lack of downregulation of epithelial Na(+) channels by mutant CF transmembrane conductance regulator. Extracellular nucleotides adenosine 5'-triphosphate (ATP) and uridine 5'-triphosphate (UTP) have been shown to activate alternative Ca(2+)-dependent Cl(-) channels in normal and CF respiratory epithelia. Recent studies suggest additional modulation of Na(+) absorption by extracellular nucleotides. In this study we examined the role of mucosal ATP and UTP in regulating Na(+) transport in native human upper airway tissues from patients with 16 patients with CF and 32 non-CF control subjects. To that end, transepithelial voltage and equivalent short-circuit current (I(SC)) were assessed by means of a perfused micro-Ussing chamber. Mucosal ATP and UTP caused an initial increase in lumen-negative I(SC) that was followed by a sustained decrease of I(sc) in both non-CF and CF tissues. The amiloride sensitive portion of I(SC) was inhibited significantly in normal and CF tissues in the presence of either ATP or UTP. Both basal Na(+) transport and nucleotide dependent inhibition of amiloride-sensitive I(SC) were significantly enhanced in CF airways compared with non-CF. Nucleotide-mediated inhibition of Na(+) absorption was attenuated by pretreatment with the Ca(2+)-adenosine triphosphatase inhibitor cyclopiazonic acid but not by inhibition of protein kinase C with bisindolylmaleimide. These data demonstrate sustained inhibition of Na(+) transport in non-CF and CF airways by mucosal ATP and UTP and suggest that this effect is mediated by an increase of intracellular Ca(2+). Because ATP and UTP inhibit Na(+) absorption and stimulate Cl(-) secretion simultaneously, extracellular nucleotides could have a dual therapeutic effect, counteracting the ion transport defect in CF lung disease. PMID- 11104729 TI - Adenovirus-mediated lung vascular endothelial growth factor overexpression protects against hypoxic pulmonary hypertension in rats. AB - Chronic hypoxic pulmonary hypertension (PH) is associated with vasoconstriction and structural remodeling of pulmonary vessels including narrowing of the arterial lumen and loss of distal functional arteries. To test whether lung overexpression of the angiogenic factor vascular endothelial growth factor (VEGF) is beneficial in hypoxic PH, recombinant adenovirus encoding the human VEGF 165 gene under the control of a cytomegalovirus promoter (Ad. VEGF) or control vector containing no gene in the expression cassette (Ad.Null) was administered intratracheally to rats. With Ad. VEGF (10(8) plaque-forming units [pfu]), VEGF protein was present in bronchoalveolar lavage fluid as early as 2 d and until 17 d after gene transfer, but was not detected in serum. Only small patchy areas of mononuclear cells without cell damage, edema, or hemorrhage were observed on lung histology with no significant change in lung permeability. In rats pretreated with Ad.VEGF (10(8) pfu) 2 d before a 2-wk exposure to hypoxia (10% O(2)), lower values versus Ad. Null-pretreated controls were found for pulmonary artery pressure (25 +/- 1 versus 30 +/- 2 mm Hg, P < 0.05), right ventricular over left ventricular-plus-septum weight (0.37 +/- 0.01 versus 0.47 +/- 0. 02, P < 0.001), normalized wall thickness of 50- to 200-microm vessels (P < 0.001), and muscularization of distal vessels (P < 0. 001). Pretreatment with Ad.VEGF (10(8) pfu) increased endothelial nitric oxide synthase activity in lung tissue and partially restored endothelium-dependent vasodilation in isolated lungs from chronically hypoxic rats, as assessed by improvement of ionophore A23187-induced vasodilation and attenuation of endothelin-1 (300 pmol)-induced vasoconstriction, an effect abolished in the presence of nitro-L-arginine methylester. We conclude that adenoviral-mediated VEGF overexpression in the lungs attenuates development of hypoxic PH, in part by protecting endothelium-dependent function. PMID- 11104730 TI - Surfactant protein A differentially regulates IFN-gamma- and LPS-induced nitrite production by rat alveolar macrophages. AB - Although several studies have demonstrated that the pulmonary collectins surfactant protein (SP)-A and SP-D contribute to innate immunity by enhancing pathogen phagocytosis, the role of SP-A and SP-D in regulating production of free radicals and cytokines is controversial. We hypothesized that the state and mechanism of activation of the immune cell influence its response to SP-A. The effects of SP-A and SP-D on production of nitric oxide (NO) and inducible nitric oxide synthase (iNOS) were assessed in isolated rat alveolar macrophages activated with lipopolysaccharide (LPS), interferon gamma (IFN-gamma), or both agonists. SP-A inhibited production of NO and iNOS in macrophages stimulated with smooth LPS, which did not significantly bind SP-A, or rough LPS, which avidly bound SP-A. In contrast, SP-A enhanced production of NO and iNOS in cells stimulated with IFN-gamma or INF-gamma plus LPS. Neither SP-A nor SP-D affected baseline NO production, and SP-D did not significantly affect production of NO in cells stimulated with either LPS or IFN-gamma. These results suggest that SP-A contributes to the lung inflammatory response by exerting differential effects on the responses of immune cells, depending on their state and mechanism of activation. PMID- 11104731 TI - CD14(+) cells are necessary for increased survival of eosinophils in response to lipopolysaccharide. AB - There has been considerable interest in the effect that gram-negative bacterial endotoxin (lipopolysaccharide [LPS]) can have in asthma, given that inhalation of LPS has been shown to cause bronchial hyperresponsiveness. Further, there is evidence that the endotoxin-binding protein CD14 may be a marker for asthma. Inhaled LPS has been shown to cause an influx of eosinophils into the nasal airway and to increase the survival of CD16-negatively selected eosinophils in vitro. In this study, we compared survival of eosinophils isolated via CD16 negative selection with eosinophils that were isolated using both CD16- and CD14 negative selection criteria. Survival of CD16-negatively selected eosinophils was enhanced by LPS in a dose-dependent manner and was inhibited by the endotoxin antagonists polymyxin B or lipid X. In contrast, depletion of CD14(+) cells within the eosinophil preparations (CD14/CD16-negatively selected eosinophils) decreased the effect of LPS on survival. Preincubation of CD16-negatively selected eosinophils with antibody 60bd, which blocks LPS binding to CD14, prevented the survival-enhancing effect of LPS. However, CD14 was not detected on eosinophils by flow cytometry, even after incubation with LPS for up to 24 h. These results suggest that the survival-enhancing effect of LPS on eosinophils requires the presence of CD14(+) cells in the population. It is our hypothesis that enhanced eosinophil survival with LPS involves the contribution of another cell type. PMID- 11104732 TI - Absence of SV40 large T-antigen expression in human mesothelioma cell lines. AB - Simian virus (SV) 40 and SV40-like DNA sequences have recently been detected in several types of human tumors, including malignant mesothelioma. However, the presence of SV40 DNA sequences is not sufficient to account for its possible role in tumor development because the viral proteins must be expressed and ultimately impair the function of relevant cell proteins, such as p53 and pRb. In this study we investigated SV40 large T antigen (SV40 Tag) protein expression in mesothelioma cell lines, established in our laboratory, by Western blotting, immunoprecipitation, and immunocytochemistry using Tag-specific mouse monoclonal antibodies (mAbs) Ab-1 (or Pab 419). By Western blotting of cell extracts, none of the mesothelioma cell lines expressed detectable amounts of SV40 Tag. However, we found that Ab-1 as well as Pab-101, another SV 40 Tag-specific mAb, may generate false-positive signals due to the fact that both antibody preparations are contaminated by a protein of similar size (90 kD) as SV40 Tag and react with the various secondary horseradish peroxidase- conjugated antimouse immunoglobulin Gs tested. The present study suggests that immunodetection of SV40 Tag protein may be puzzling because this contaminating Taglike protein may bind to particular cell structures, thereby generating false-positive signals. PMID- 11104733 TI - Tumor necrosis factor-alpha-induced secretion of RANTES and interleukin-6 from human airway smooth-muscle cells. Modulation by cyclic adenosine monophosphate. AB - Although 3':5' cyclic adenosine monophosphate (cAMP) is known to modulate cytokine production in a number of cell types, little information exists regarding cAMP-mediated effects on this synthetic function of human airway smooth muscle (HASM) cells. We examined the effect of increasing intracellular cAMP concentration ([cAMP](i)) on tumor necrosis factor (TNF)-alpha-induced regulated on activation, normal T cells expressed and secreted (RANTES) and interleukin (IL)-6 secretion from cultured HASM cells. Pretreatment of HASM with prostaglandin (PG) E(2), forskolin, or dibutyryl cAMP inhibited TNF-alpha-induced RANTES secretion but increased TNF-alpha-induced IL-6 secretion. Moreover, stimulation with PGE(2), forskolin, or dibutyryl cAMP alone increased basal IL-6 secretion in a concentration-dependent manner. SB 207499, a specific phosphodiesterase type 4 inhibitor, augmented the inhibitory effects of PGE(2) and forskolin on TNF-alpha-induced RANTES. Collectively, these data demonstrate that increasing [cAMP](i) in HASM effectively increases IL-6 secretion but reduces RANTES secretion promoted by TNF-alpha. Reverse transcriptase/polymerase chain reaction and ribonuclease protection assays suggested that these opposite effects of increased [cAMP](i) on TNF-alpha- induced IL-6 and RANTES secretion may occur at the transcriptional level. Accordingly, we examined the effects of TNF- alpha and cAMP on the regulation of nuclear factor (NF)-kappaB, a transcription factor known to modulate cytokine synthesis in numerous cell types. Stimulation of HASM cells with TNF-alpha increased NF-kappaB DNA-binding activity. However, increased [cAMP](i) in HASM neither activated NF-kappaB nor altered TNF-alpha- induced NF-kappaB DNA-binding activity. These results were confirmed using a NF-kappaB-luciferase reporter assay. Together, our data suggest that TNF-alpha-induced IL-6 and RANTES secretion may be associated with NF-kappaB activation, and that inhibition of TNF-alpha-stimulated RANTES secretion and augmentation of IL-6 secretion by increased [cAMP](i) in HASM cells occurs via an NF-kappaB-independent mechanism. PMID- 11104734 TI - Predicting the long-QT genotype from clinical data: from sense to science. PMID- 11104735 TI - One-year clinical outcome after minimally invasive direct coronary artery bypass. AB - BACKGROUND: Minimally invasive coronary artery bypass (MIDCAB) is a new surgical technique by which the left internal mammary artery is anastomosed under direct visualization to the left anterior descending artery without cardiopulmonary bypass. METHODS AND RESULTS: We followed all 274 patients who underwent MIDCAB from the time it was introduced at a single center. In-hospital and 1-year clinical events were source-documented and adjudicated. The in-hospital major acute cardiac event rate was 2.2%; this included a 1.1% mortality rate. At 1 year, the respective rates were 7.8% and 2. 5%. When compared with the initial 100 procedures, the subsequent 174 procedures had shorter vessel occlusion times (10+/-5 versus 14+/-6 minutes; P:=0.009), times to extubation (6+/-3 versus 14+/ 10 hours; P:<0.001), and lengths of hospital stay (2.1+/-1.9 versus 3. 2+/-3.1 days; P:=0.04). Cumulative 1-year adverse cardiac events were 11% in the initial 100 cases and 6% in the subsequent 174 cases (P:=0.17). CONCLUSIONS: Excellent clinical results can be achieved with the MIDCAB technique. The clinical adverse event rate may decrease with accumulated experience. PMID- 11104736 TI - Simvastatin improves disturbed endothelial barrier function. AB - BACKGROUND: Recent clinical trials have established that inhibitors of the enzyme 3-hydroxy-3-methylglutaryl coenzyme A reductase (statins) reduce the risk of acute coronary events. These effects of statins cannot be fully explained by their lipid-lowering potential. Improved endothelial function may contribute to the positive effects of statin treatment. METHODS AND RESULTS: In the present study, we report that simvastatin reduces endothelial barrier dysfunction, which is associated with the development of atherosclerosis. Treatment of human umbilical vein endothelial cells for 24 hours with 5 micromol/L simvastatin reduced the thrombin-induced endothelial barrier dysfunction in vitro by 55+/-3%, as assessed by the passage of peroxidase through human umbilical vein endothelial cell monolayers. Similar effects were found on the thrombin-induced passage of (125)I-LDL through human aortic endothelial cell monolayers. This reduction in barrier dysfunction by simvastatin was both dose and time dependent and was accompanied by a reduction in the thrombin-induced formation of stress fibers and focal adhesions and membrane association of RhoA. Simvastatin treatment had no effect on intracellular cAMP levels. In Watanabe heritable hyperlipidemic rabbits, treatment for 1 month with 15 mg/kg simvastatin reduced vascular leakage in both the thoracic and abdominal part of the aorta, as evidenced by the Evans blue dye exclusion test. The decreased permeability was not accompanied by a reduction of oil red O-stainable atherosclerotic lesions. CONCLUSIONS: These data show that simvastatin, in a relatively high concentration, improves disturbed endothelial barrier function both in vitro and in vivo. The data also support the beneficial effects of simvastatin in acute coronary events by mechanisms other than its lipid-lowering effect. PMID- 11104737 TI - Nitroglycerin tolerance in human vessels: evidence for impaired nitroglycerin bioconversion. AB - BACKGROUND: The basis for progressive attenuation of the effects of organic nitrates during long-term therapy (nitrate tolerance) remains controversial; proposed mechanisms include impaired nitrate bioconversion resulting in decreased release of nitric oxide (NO) from nitrates and/or increased NO clearance through a reaction with incrementally generated superoxide (O(2)(-)). METHODS AND RESULTS: Patients undergoing elective coronary artery bypass were randomized to receive 24 hours of intravenously infused nitroglycerin (NTG; nitrate group) or no nitrate therapy (control group). Discarded segments of the internal mammary artery and saphenous vein were used to examine (1) vascular responsiveness to NTG, sodium nitroprusside, and the calcium ionophore A23187; (2) bioconversion of NTG to 1,2- and 1,3-glyceryl dinitrate; and (3) the generation of O(2)(-). Responses to NTG were reduced 3- to 5-fold in vessels from the nitrate group compared with control vessels (P:<0. 01 for both types of segments), whereas responses to sodium nitroprusside and A23187 were unchanged. Tissue content of 1, 2-glyceryl dinitrate was lower (P:=0.012) in the saphenous veins from the nitrate group than in those from the control group. O(2)(-) generation was greater (P:<0.01) in internal mammary artery samples from the nitrate group than in those from the control group. However, incremental O(2)(-) generation induced by an inhibitor of superoxide dismutase did not affect NTG responses. CONCLUSIONS: NTG tolerance in patients with coronary artery disease is nitrate-specific and is associated with evidence of impaired NTG bioconversion. Tolerance was associated with incremental O(2)(-) generation, but short-term elevation of O(2)(-) did not affect NTG responsiveness, suggesting increased NO clearance by O(2)(-) has a minimal contribution to tolerance. PMID- 11104738 TI - Comparison of novel hemostatic factors and conventional risk factors for prediction of coronary heart disease. AB - BACKGROUND: This study sought to assess whether novel markers of hemostatic activity are predictive of coronary heart disease (CHD) and improve risk assessment. METHODS AND RESULTS: Conventional CHD risk factors, the activation peptides of factor IX and factor X, factor VII activity and antigen, activated factor XII, prothrombin fragment 1+2, fibrinopeptide A, and fibrinogen were measured in 1153 men aged 50 to 61 years who were free of myocardial infarction at recruitment. Activated factor VII (VIIa) was measured in 829 men. During 7.8 years of follow-up, 104 had a CHD event. Baseline status was related to outcome by logistic regression by using a modified nested case-control design. Screening performance was judged from receiver operating characteristic curves. A high activated factor XII was associated with increased CHD risk, but low levels were not protective. Plasma VIIa and factor X activation peptide were independently and inversely related to risk. Plasma factor IX activation peptide and fibrinogen were positively associated with risk, but the relations were no longer statistically significant after adjustment for other factors, including VIIa and apoA-I. Other hemostatic markers were not associated with CHD risk. CONCLUSIONS: Hemostatic status did not add significant predictive power to that provided by conventional CHD risk factors yet was able to substitute effectively for these factors. PMID- 11104739 TI - Noninvasive coronary angiography by retrospectively ECG-gated multislice spiral CT. AB - BACKGROUND: We investigated the applicability and image quality of contrast enhanced coronary artery visualization by multislice spiral CT using retrospective ECG gating. METHODS AND RESULTS: Twenty-five patients in sinus rhythm (significant coronary artery stenoses ruled out by invasive angiography) were studied with a multislice spiral CT (Siemens SOMATOM Volume Zoom). In inspiration (mean breath-hold, 37 seconds), a volume data set of the heart was acquired (intravenous contrast agent; 4 x 1-mm slice thickness; 500-ms rotation; table feed, 1.5 mm/360 degrees ). Simultaneous recording of the ECG permitted retrospective reconstruction of contiguous cross sections in intervals of 1 mm at any desired interval of the cardiac cycle. The mean duration of the image reconstruction window was 185 ms. Next to 3-dimensional reconstructions of the heart and coronary arteries, multiplanar reconstructions were rendered to determine the visualized length of the coronary arteries, the contrast-to-noise ratio, and the correlation of coronary artery diameters to quantitative coronary angiography. CONCLUSIONS: The coronary arteries could be visualized over long segments (left main, 9+/-4 mm; left anterior descending, 112+/-34 mm; left circumflex, 80+/-29 mm; right coronary artery, 116+/-33 mm). On average, 78+/-16% of these distances were visualized free of motion artifacts. The mean contrast-to noise ratio was 9.3+/-3.3. Coronary artery diameters in multislice spiral CT showed close correlation to quantitative coronary angiography (CT, 3.3+/-1.0 mm; angiography, 3. 2+/-0.9 mm; mean difference, 0.38 mm; r=0.86). Contrast-enhanced multislice spiral CT permits visualization of the coronary artery lumen. Further studies are necessary to determine whether image quality is sufficient to reliably detect coronary artery stenoses. PMID- 11104740 TI - Predictors of disease course in patients with acute myocarditis. AB - BACKGROUND: Clinical manifestations of acute myocarditis, with distinct onset, vary from asymptomatic to fatal. The predictors of the course of the disease in patients with acute myocarditis at initial presentation have not yet been established. In this study, we examined the predictive values of various parameters in the disease course of patients with myocarditis. METHODS AND RESULTS: Twenty-one consecutive patients who had been diagnosed as having acute myocarditis by histological examinations were analyzed. The patients with myocarditis were divided into the survival group (n=13) and the fatal group (n=8). We examined the parameters of the clinical state, hemodynamic variables, required therapies, biochemical laboratory data, and cytokines. The control groups were composed of 23 patients with old myocardial infarction and 20 healthy volunteers. The fatal group had lower blood pressure and higher pulmonary capillary wedge pressure compared with those values in the survival group. Mechanical ventilation support was more frequently required in the fatal group. Serum levels of soluble Fas (sFas) and soluble Fas ligand (sFasL) were significantly higher in the myocarditis group than in the 2 control groups. Furthermore, levels were significantly higher in the fatal group than in the survival group for sFas (13.93+/-4.77 versus 3.77+/-0.52 ng/mL, respectively; P:<0.001) and sFasL (611.4+/-127.7 versus 269.5+/-37.3 pg/mL, respectively; P:<0.05). Other clinical states, hemodynamic variables, required therapies, and biochemical laboratory parameters were not different between the 2 groups. CONCLUSIONS: Elevation of sFas and sFasL levels at initial presentation appear to be a good serological marker to predict the prognosis of acute myocarditis. PMID- 11104741 TI - Evidence for possible involvement of 5-HT(2B) receptors in the cardiac valvulopathy associated with fenfluramine and other serotonergic medications. AB - BACKGROUND: Serotonergic medications with various mechanisms of action are used to treat psychiatric disorders and are being investigated as treatments for drug dependence. The occurrence of fenfluramine-associated valvular heart disease (VHD) has raised concerns that other serotonergic medications might also increase the risk of developing VHD. We hypothesized that fenfluramine or its metabolite norfenfluramine and other medications known to produce VHD have preferentially high affinities for a particular serotonin receptor subtype capable of stimulating mitogenesis. METHODS AND RESULTS: Medications known or suspected to cause VHD (positive controls) and medications not associated with VHD (negative controls) were screened for activity at 11 cloned serotonin receptor subtypes by use of ligand-binding methods and functional assays. The positive control drugs were (+/-)-fenfluramine; (+)-fenfluramine; (-)-fenfluramine; its metabolites (+/ )-norfenfluramine, (+)-norfenfluramine, and (-)-norfenfluramine; ergotamine; and methysergide and its metabolite methylergonovine. The negative control drugs were phentermine, fluoxetine, its metabolite norfluoxetine, and trazodone and its active metabolite m-chlorophenylpiperazine. (+/-)-, (+)-, and (-) Norfenfluramine, ergotamine, and methylergonovine all had preferentially high affinities for the cloned human serotonin 5-HT(2B) receptor and were partial to full agonists at the 5-HT(2B) receptor. CONCLUSIONS: Our data imply that activation of 5-HT(2B) receptors is necessary to produce VHD and that serotonergic medications that do not activate 5-HT(2B) receptors are unlikely to produce VHD. We suggest that all clinically available medications with serotonergic activity and their active metabolites be screened for agonist activity at 5-HT(2B) receptors and that clinicians should consider suspending their use of medications with significant activity at 5-HT(2B) receptors. PMID- 11104742 TI - Risk factors for infective endocarditis: oral hygiene and nondental exposures. AB - BACKGROUND: The risks of infective endocarditis (IE) associated with various conditions and procedures are poorly defined. METHODS AND RESULTS: This was a population-based case-control study conducted in 54 Philadelphia, Pa-area hospitals from 1988 to 1990. Community-acquired IE cases unassociated with intravenous drug use were compared with matched community residents. Subjects were interviewed for risk factors. Diagnoses were confirmed by expert review of medical record abstracts with risk factor data removed. Cases were more likely than controls to suffer from prior severe kidney disease (adjusted OR [95% CI]=16.9 [1.5 to 193], P:=0.02) and diabetes mellitus (adjusted OR [95% CI]=2.7 [1.4 to 5.2], P:=0.004). Cases infected with skin flora had received intravenous fluids more often (adjusted OR [95% CI]=6.7 [1.1 to 41], P:=0.04) and had more often had a previous skin infection (adjusted OR [95% CI]=3.5 [0.7 to 17], P:=0.11). No association was seen with pulmonary, gastrointestinal, cardiac, or genitourinary procedures or with surgery. Edentulous patients had a lower risk of IE from dental flora than patients who had teeth but did not floss. Daily flossing was associated with a borderline decreased IE risk. CONCLUSIONS: Within the limits of the available sample size, the data showed that IE patients differ from people without IE with regard to certain important risk factors but not regarding recent procedures. PMID- 11104743 TI - Spectrum of ST-T-wave patterns and repolarization parameters in congenital long QT syndrome: ECG findings identify genotypes. AB - BACKGROUND: Congenital long-QT syndrome (LQTS) is caused by mutations of genes encoding the slow component of the delayed rectifier current (LQT1, LQT5), the rapid component of the delayed rectifier current (LQT2, LQT6), or the Na(+) current (LQT3), resulting in ST-T-wave abnormalities on the ECG. This study evaluated the spectrum of ST-T-wave patterns and repolarization parameters by genotype and determined whether genotype could be identified by ECG. METHODS AND RESULTS: ECGs of 284 gene carriers were studied to determine ST-T-wave patterns, and repolarization parameters were quantified. Genotypes were identified by individual ECG versus family-grouped ECG analysis in separate studies using ECGs of 146 gene carriers from 29 families and 233 members of 127 families undergoing molecular genotyping, respectively. Ten typical ST-T patterns (4 LQT1, 4 LQT2, and 2 LQT3) were present in 88% of LQT1 and LQT2 carriers and in 65% of LQT3 carriers. Repolarization parameters also differed by genotype. A combination of quantified repolarization parameters identified genotype with sensitivity/specificity of 85%/70% for LQT1, 83%/94% for LQT2, and 47%/63% for LQT3. Typical patterns in family-grouped ECGs best identified the genotype, being correct in 56 of 56 (21 LQT1, 33 LQT2, and 2 LQT3) families with mutation results. CONCLUSIONS: Typical ST-T-wave patterns are present in the majority of genotyped LQTS patients and can be used to identify LQT1, LQT2, and possibly LQT3 genotypes. Family-grouped ECG analysis improves genotype identification accuracy. This approach can simplify genetic screening by targeting the gene for initial study. The multiple ST-T patterns in each genotype raise questions regarding the pathophysiology and regulation of repolarization in LQTS. PMID- 11104744 TI - Cryothermal ablation of the slow pathway for the elimination of atrioventricular nodal reentrant tachycardia. AB - BACKGROUND: We report the first successful slow pathway ablation using a novel catheter-based cryothermal technology for the elimination of atrioventricular nodal reentrant tachycardia (AVNRT). METHODS AND RESULTS: Eighteen patients with typical AVNRT underwent cryoablation. Reversible loss of slow pathway (SP) conduction during cryothermy (ice mapping) was demonstrated in 11 of 12 patients. Because of time constraints, only 2 sites were ice mapped in 1 patient. Seventeen of 18 patients had successful cryoablation of the SP. One patient had successful ice mapping of the SP, but inability to cool beyond -38 degrees C prevented successful cryoablation. A single radiofrequency lesion at this site eliminated SP conduction. No patient has had recurrent AVNRT over 4.9+/-1.7 months of follow up. During cryoablation, accelerated junctional tachycardia was not seen and was therefore not available to guide lesion delivery. Adherence of the catheter tip during cryothermy (cryoadherence) allowed atrial pacing to test for SP conduction. Cryoablation in the anterior septum produced inadvertent transient PR prolongation consistent with loss of fast pathway conduction in 1 patient and transient (6.5 seconds) 2:1 AV block in another. On rewarming, the PR interval returned to normal, and the AV nodal effective refractory period was unchanged in both. Accelerated junctional tachycardia was seen on rewarming in both but not during cryothermy. CONCLUSIONS: Cryothermal ablation of the SP was achieved in patients with this novel technique. Successful ice mapping of both the SP and fast pathway was demonstrated. The ability to test the functionality of specific ablation sites before production of a permanent lesion may eliminate inadvertent AV block. PMID- 11104745 TI - Remodeling of carotid artery is associated with increased expression of matrix metalloproteinases in mouse blood flow cessation model. AB - BACKGROUND: The matrix-degrading activity of matrix metalloproteinases (MMPs), required for cell migration and general tissue reshaping, is thought essential for pathological arterial remodeling in atherosclerosis and restenosis. METHODS AND RESULTS: We triggered remodeling of the carotid artery in C57BL/6 mice by blood flow cessation to study the relationship with gelatinases MMP-9 and MMP-2. Ligated and contralateral carotid arteries from ligated and sham-operated mice were harvested fresh, for biochemical analyses, or were perfusion-fixed, for histological studies, at 0, 1, 3, 7, 14, and 28 days after ligation. An early statistically significant (P:<0.01) 4- to 5-fold increase in MMP-9 expression detected by SDS-PAGE zymography and Western blotting in tissue homogenates of ligated carotid arteries 1 day after flow cessation was maintained through day 7, after which expression gradually fell. Maximal MMP-9 levels were higher than MMP 2 levels, which became significantly increased 7 days after ligation. Proliferating cells, identified by bromodeoxyuridine incorporation, were detectable at day 1 in the adventitia and subsequently throughout the wall. Neointima was visible in 3-day specimens of ligated arteries. Suggested by morphology and predicted by theoretical considerations, maximal MMP-9 expression coincided with cell migration into the neointima, supporting its enabling role. Morphological measurements also demonstrated positive lumen remodeling up to 7 days after ligation. CONCLUSIONS: MMP-9 induction is associated with the formation of intimal hyperplasia and does not require frank mechanical injury. Our data also show that a significant increase in MMP-9 expression preceded the positive geometrical remodeling of arteries, suggesting a potentially beneficial role for this matrix-degrading enzyme. PMID- 11104746 TI - 1alpha,25-dihydroxyvitamin D(3) and its potent synthetic analogs downregulate tissue factor and upregulate thrombomodulin expression in monocytic cells, counteracting the effects of tumor necrosis factor and oxidized LDL. AB - BACKGROUND: We have recently found that a hormonally active form of vitamin D, 1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)], exerts anticoagulant effects by upregulating the expression of an anticoagulant glycoprotein, thrombomodulin (TM), and downregulating the expression of a critical coagulation factor, tissue factor (TF), in monocytic cells including human peripheral monocytes. In this study, we investigated the counteracting effects of 1,25(OH)(2)D(3) and its potent analogs on TF induction and TM downregulation by tumor necrosis factor and oxidized LDL in monocytic cells and the modulatory effects of potent analogs on TF and TM expression. METHODS AND RESULTS: Effects of 1,25(OH)(2)D(3) and its potent synthetic analogs (22R)-22-methyl-20-epi-1,25(OH)(2)D(3) (KY3) and 22 oxacalcitriol on TF and TM antigen levels, cell surface activities, and mRNA levels in monocytic cells were examined. 1, 25(OH)(2)D(3) and its potent analogs showed anticoagulant effects in monocytic cells by downregulating TF and upregulating TM expression, counteracting the effects of tumor necrosis factor and oxidized LDL. KY3 was most potent in its regulatory effect on TF and TM expression. CONCLUSIONS: Because KY3 has the highest affinity for vitamin D receptor, our findings suggest that TF and TM regulation by 1, 25(OH)(2)D(3) analogs is also mediated by vitamin D receptor. The 1, 25(OH)(2)D(3) analogs KY3 and 22-oxacalcitriol may have the potential to serve as an agent for preventing and treating atherosclerotic and other cytokine-mediated thrombotic diseases and as a tool for studying the molecular mechanisms of TF and TM regulation. PMID- 11104747 TI - Insulin prevents cardiomyocytes from oxidative stress-induced apoptosis through activation of PI3 kinase/Akt. AB - BACKGROUND: Loss of cardiomyocytes by apoptosis is proposed to cause heart failure. Reactive oxygen species induce apoptosis in many types of cells including cardiomyocytes. Because insulin has been reported to have protective effects, we examined whether insulin prevents cardiomyocytes from oxidative stress-induced apoptotic death. METHODS AND RESULTS: Cultured cardiomyocytes of neonatal rats were stimulated by hydrogen peroxide (H(2)O(2)). Apoptosis was evaluated by means of the TUNEL method and DNA laddering. Incubation with 100 micromol/L H(2)O(2) for 24 hours increased the number of TUNEL-positive cardiac myocytes (control, approximately 4% versus H(2)O(2), approximately 23%). Pretreatment with 10(-)(6) mol/L insulin significantly decreased the number of H(2)O(2)-induced TUNEL-positive cardiac myocytes (approximately 12%) and DNA fragmentation induced by H(2)O(2). Pretreatment with a specific phosphatidylinositol 3 kinase (PI3K) inhibitor, wortmannin, and overexpression of dominant negative mutant of PI3K abolished the cytoprotective effect of insulin. Insulin strongly activated both PI3K and the putative downstream effector AKT: Moreover, a proapoptotic protein, BAD:, was significantly phosphorylated and inactivated by insulin through PI3K. CONCLUSIONS: These results suggest that insulin protects cardiomyocytes from oxidative stress-induced apoptosis through the PI3K pathway. PMID- 11104748 TI - Aerosol gene transfer with inducible nitric oxide synthase reduces hypoxic pulmonary hypertension and pulmonary vascular remodeling in rats. AB - BACKGROUND: Nitric oxide (NO) is a potent vasodilator with an important role in the regulation of pulmonary vascular tone. The effects of NO synthase (NOS) gene transfer on pulmonary vascular remodeling associated with hypoxic pulmonary hypertension are unknown. METHODS AND RESULTS: We aerosolized 3 x 10(9) pfu of an adenoviral vector containing inducible NOS gene (AdNOS2), constitutive NOS3 gene (AdNOS3), or no transgene (AdRR5) into rat lungs. Exhaled NO levels, monitored with chemiluminescence, were higher in AdNOS2-infected rats than in AdNOS3- and AdRR5-infected rats (at 3 days, 33+/-6 ppb, n=9, versus 17+/-4, n=9, and 6+/-2 ppb, n=3, P:<0.05 for both). Exposure to FIO(2) 0.10 for 7 days increased pulmonary artery pressure from 19+/-4 mm Hg (baseline) to 27+/-1 and 26+/-2 mm Hg in AdNOS3- and AdRR5-infected rats, respectively, but only to 21+/-1 mm Hg in AdNOS2-infected animals (P:<0.05). After 7 days of hypoxia, total pulmonary resistance in AdRR5- and AdNOS3-infected rats was significantly higher than in AdNOS2-infected animals (0.41+/-0.05 and 0.39+/-0.07 versus 0.35+/-0. 03 mm Hg. mL(-)(1). min(-)(1), respectively, P:<0.05). Right ventricular hypertrophy was reduced in AdNOS2-infected rats [right ventricular/(left ventricular+septal) weight, 0.19+/-0.10 versus 0. 28+/-0.10 and 0.32+/-0.10 in AdRR5- and AdNOS3 infected rats, respectively, P:<0.05]. The percentage of muscularized precapillary pulmonary resistance vessels was also significantly decreased (18+/ 4% versus 25+/-8% and 30+/-5% in AdRR5- and AdNOS3-infected rats, P:<0.05). CONCLUSIONS: Aerosol NOS2 gene transfer increases pulmonary NO production and significantly reduces hypoxic pulmonary hypertension and pulmonary vascular remodeling. Aerosol NOS2 gene transfer may be a promising strategy to target pulmonary vascular disorders. PMID- 11104749 TI - Electrophysiological deterioration during long-duration ventricular fibrillation. AB - BACKGROUND: Probability of survival from sudden cardiac arrest caused by ventricular fibrillation (VF) decreases rapidly with fibrillation duration. We hypothesized that cellular ischemia/fibrillation-induced electrophysiological deterioration underlies decreased survival. METHODS AND RESULTS: We determined fibrillation monophasic action potential (MAP) morphology including action potential frequency content, duration, cycle length, developing diastolic intervals, and amplitude as a function of ischemic fibrillation duration in 10 isolated rabbit hearts. We also correlated ECG frequency (used clinically) and MAP amplitude and frequency. Fibrillation cycle length and diastole duration increased, whereas APD(100) shortened significantly with time (P:<0.001). Between 1 and 3 minutes, diastole appeared primarily as the result of APD(100) shortening, with only small changes in cycle length. Between 2 and 5 minutes, diastole increased primarily as the result of increased cycle length. Diastole developed progressively from 5% of VF cycles at 5 seconds to approximately 100% of VF cycles by 120 seconds (P:<0.001). Diastole increased from 1% of cycle length at 5 seconds to 62% at 5 minutes. Its duration increased from 4.7 ms at 5 seconds to 90 ms at 5 minutes (P:<0.001). Both MAP and ECG 1/frequency closely correlated with fibrillation cycle length. CONCLUSIONS: These results show a rapid and progressive electrophysiological deterioration during fibrillation, leading to electrical diastole between fibrillation action potentials. This rapid deterioration may explain the decreased probability of successful resuscitation after prolonged fibrillation. Therefore, a greater understanding of cellular deterioration during fibrillation may lead to improved resuscitation methods, including development of specific defibrillator waveforms for out-of-hospital cardiac arrest. PMID- 11104750 TI - Chronic hypoxia stimulates periarterial sympathetic nerve development in chicken embryo. AB - BACKGROUND: Epidemiological findings suggest an association between low-for-age birth weight and the risk to develop coronary heart diseases in adulthood. During pregnancy, an imbalance between fetal demands and supply may result in permanent alterations of neuroendocrine development in the fetus. We evaluated whether chronic prenatal hypoxia increases arterial sympathetic innervation. METHODS AND RESULTS: Chicken embryos were maintained from 0.3 to 0.9 of the 21-day incubation period under normoxic (21% O(2)) or hypoxic conditions (15% O(2)). At 0.9 incubation, the degree of sympathetic innervation of the embryonic femoral artery was determined by biochemical, histological, and functional (in vitro contractile reactivity) techniques. Chronic hypoxia increased embryonic mortality (32% versus 13%), reduced body weight (21.9+/-0.4 versus 25.4+/-0.6 g), increased femoral artery norepinephrine (NE) content (78.4+/-9.4 versus 57.5+/-5.0 pg/mm vessel length), and increased the density of periarterial sympathetic nerve fibers (14.4+/-0.7 versus 12.5+/-0.6 counts/10(4) microm(2)). Arteries from hypoxic embryos were less sensitive to NE (pD(2), 5.99+/-0.04 versus 6. 21+/-0.10). In the presence of cocaine, however, differences in sensitivity were no longer present. In the embryonic heart, NE content (156.9+/-11.0 versus 108.1+/-14.7 pg/mg wet wt) was also increased after chronic hypoxia. CONCLUSIONS: In the chicken embryo, chronic moderate hypoxia leads to sympathetic hyperinnervation of the arterial system. In humans, an analogous mechanism may increase the risk for cardiovascular disease in adult life. PMID- 11104751 TI - The elusive pathophysiology of neurally mediated syncope. PMID- 11104752 TI - Malignant ventricular arrhythmias due to Aconitum napellus seeds. PMID- 11104753 TI - Controversy over myocardial revascularization, either transmyocardial or percutaneous, continues as experts spar over advantages of each. PMID- 11104754 TI - Exogenous Mg-ATP induces a large inhibition of pyruvate kinase in intact rat hepatocytes. AB - Mg-ATP infusion in vivo has been reported to be beneficial both to organ function and survival rate in various models of shock. Moreover, a large variety of metabolic effects has been shown to occur in several tissues due to purinergic receptor activation. In the present work we studied the effects of exogenous Mg ATP in rat liver cells perifused with dihydroxyacetone to investigate simultaneously gluconeogenetic and glycolytic pathways. We found a significant effect on oxidative phosphorylation as characterized by a decrease in oxygen consumption rate and in the cellular ATP-to-ADP ratio associated with an increase in lactate-to-pyruvate ratio. In addition, exogenous Mg-ATP induced rapid and reversible inhibition of both gluconeogenesis and glycolysis. The main effect on gluconeogenesis was located at the level of the fructose cycle, whereas the decrease in glycolysis was due to a strong inhibition of pyruvate kinase. Although pyruvate kinase inhibition induced by exogenous Mg-ATP was allosteric when assessed in vitro after enzyme extraction, we found a large decrease in the apparent maximal velocity when kinetics were assessed in vivo in intact perifused hepatocytes. This newly described short-term regulation of pyruvate kinase occurs only in the intact cell and may open new potentials for the pharmacological regulation of pyruvate kinase in vivo. PMID- 11104755 TI - Substitution of a glycogen synthase kinase-3beta phosphorylation site in presenilin 1 separates presenilin function from beta-catenin signaling. AB - The majority of cases with early onset familial Alzheimer's disease have been attributed to mutations in the presenilin 1 (PS1) gene. PS1 protein is a component of a high molecular weight membrane-bound complex that also contains beta-catenin. The physiological relevance of the association between PS1 and beta catenin remains controversial. In this study, we report the identification and functional characterization of a highly conserved glycogen synthase kinase-3beta consensus phosphorylation site within the hydrophilic loop domain of PS1. Site directed mutagenesis, together with in vitro and in vivo phosphorylation assays, indicates that PS1 residues Ser(353) and Ser(357) are glycogen synthase kinase 3beta targets. Substitution of one or both of these residues greatly reduces the ability of PS1 to associate with beta-catenin. By disrupting this interaction, we demonstrate that the association between PS1 and beta-catenin has no effect on Abeta peptide production, beta-catenin stability, or cellular susceptibility to apoptosis. Significantly, in the absence of PS1/beta-catenin association, we found no alteration in beta-catenin signaling using induction of this pathway by exogenous expression of Wnt-1 or beta-catenin and a Tcf/Lef transcriptional assay. These results argue against a pathologically relevant role for the association between PS1 and beta-catenin in familial Alzheimer's disease. PMID- 11104756 TI - Elimination of phosphorylation sites of Semliki Forest virus replicase protein nsP3. AB - nsP3 is one of the four RNA replicase subunits encoded by alphaviruses. The specific essential functions of nsP3 remain unknown, but it is known to be phosphorylated on serine and threonine residues. Here we have completed mapping of the individual phosphorylation sites on Semliki Forest virus nsP3 (482 amino acids) by point mutational analysis of threonine residues. This showed that threonines 344 and 345 represented the major threonine phosphorylation sites in nsP3. Experiments with deletion variants suggested that nsP3 itself had no kinase activity; instead, it was likely to be phosphorylated by multiple cellular kinases. Phosphorylation was not necessary for the peripheral membrane association of nsP3, which was mediated by the N-terminal region preceding the phosphorylation sites. Two deletion variants of nsP3 with either reduced or undetectable phosphorylation were studied in the context of virus infection. Cells infected with mutant viruses produced close to wild type levels of infectious virions; however, the rate of viral RNA synthesis was significantly reduced in the mutants. A virus totally defective in nsP3 phosphorylation and exhibiting a decreased rate of RNA synthesis also exhibited greatly reduced pathogenicity in mice. PMID- 11104757 TI - Consequences of omega -6-oleate desaturase deficiency on lipid dynamics and functional properties of mitochondrial membranes of Arabidopsis thaliana. AB - We probed the role of the polyunsaturated fatty acids on the dynamic and functional properties of mitochondrial membranes using the fad2 mutant of Arabidopsis thaliana, deficient in omega-6-oleate desaturase. In mitochondria of this mutant, the oleic acid content exceeded 70% of the total fatty acids, and the lipid/protein ratio was greatly enhanced. As a consequence, local microviscosity, probed by anthroyloxy fatty acid derivatives, was increased by 30%, whereas the lipid lateral diffusion, assayed using 1-pyrenedodecanoic acid, was approximately 4 times increased. Functional parameters such as oxygen consumption rate under phosphorylating and nonphosphorylating conditions and proton permeability of the inner mitochondrial membrane were significantly reduced in fad2 mitochondrial membranes, while the thermal dependence of the respiration was enhanced. Moreover, metabolic control analysis of the respiration clearly showed an enhancement of the control exerted by the membrane proton leaks. Our data suggest that the loss of omega-6-oleate desaturase activity in Arabidopsis cells induced an enhancement of both microviscosity and lipid/protein ratio of mitochondrial membranes, which in turn were responsible for the change in lateral mobility of lipids and for bioenergetic parameter modifications. PMID- 11104758 TI - Possible roles of protein kinase A in cell motility and excystation of the early diverging eukaryote Giardia lamblia. AB - Since little is known of how the primitive protozoan parasite, Giardia lamblia, senses and responds to its changing environment, we characterized a giardial protein kinase A (gPKA) catalytic subunit with unusual subcellular localization. Sequence analysis of the 1080-base pair open reading frame shows 48% amino acid identity with the cyclic AMP-dependent kinase from Euglena gracilis. Northern analysis indicated a 1.28- kilobase pair transcript at relatively constant concentrations during growth and encystation. gPKA is autophosphorylated, although amino acid residues corresponding to Thr-197 and Ser-338 of human protein kinase A (PKA) that are important for autophosphorylation are absent. Kinetic analysis of the recombinant PKA showed that ATP and magnesium are preferred over GTP and manganese. Kinase activity of the native PKA has also been detected in crude extracts using kemptide as a substrate. A myristoylated PKA inhibitor, amide 14-22, inhibited excystation with an IC(50) of 3 microm, suggesting an important role of gPKA during differentiation from the dormant cyst form into the active trophozoite. gPKA localizes independently of cell density to the eight flagellar basal bodies between the two nuclei together with centrin, a basal body/centrosome-specific protein. However, localization of gPKA to marginal plates along the intracellular portions of the anterior and caudal pairs of flagella was evident only at low cell density and higher endogenous cAMP concentrations or after refeeding with fresh medium. These data suggest an important role of PKA in trophozoite motility during vegetative growth and the cellular activation of excystation. PMID- 11104759 TI - Predicted Michaelis-Menten complexes of cocaine-butyrylcholinesterase. Engineering effective butyrylcholinesterase mutants for cocaine detoxication. AB - Butyrylcholinesterase (BChE) is important in cocaine metabolism, but it hydrolyzes (-)-cocaine only one-two thousandth as fast as the unnatural (+) stereoisomer. A starting point in engineering BChE mutants that rapidly clear cocaine from the bloodstream, for overdose treatment, is to elucidate structural factors underlying the stereochemical difference in catalysis. Here, we report two three-dimensional Michaelis-Menten complexes of BChE liganded with natural and unnatural cocaine molecules, respectively, that were derived from molecular modeling and supported by experimental studies. Such complexes revealed that the benzoic ester group of both cocaine stereoisomers must rotate toward the catalytic Ser(198) for hydrolysis. Rotation of (-)-cocaine appears to be hindered by interactions of its phenyl ring with Phe(329) and Trp(430). These interactions do not occur with (+)-cocaine. Because the rate of (-)-cocaine hydrolysis is predicted to be determined mainly by the re-orientation step, it should not be greatly influenced by pH. In fact, measured rates of this reaction were nearly constant over the pH range from 5.5 to 8.5, despite large rate changes in hydrolysis of (+)-cocaine. Our models can explain why BChE hydrolyzes (+)-cocaine faster than (-)-cocaine, and they suggest that mutations of certain residues in the catalytic site could greatly improve catalytic efficiency and the potential for detoxication. PMID- 11104760 TI - Regulation of elongation factor-1alpha expression by growth factors and anti receptor blocking antibodies. AB - The epidermal growth factor (EGF) family and its receptors regulate normal and cancerous epithelial cell proliferation, a process that could be suppressed by anti-receptor blocking antibodies. Polypeptide elongation factor-1alpha (EF 1alpha) is a multifunctional protein whose levels are positively correlated with the proliferative state of cells. To identify genes, whose expression may be modulated by anti-receptor blocking antibodies, we performed a differential display screening and isolated differentially expressed cDNAs. Isolates from one clone were 100% identical to human EF-1alpha. Both EGF and heregulin-beta1 (HRG) induced EF-1alpha promoter activity and mRNA and protein expression. Growth factor-mediated EF-1alpha expression was effectively blocked by pretreatment with humanized anti-EGF receptor antibody C225 or anti-human epidermal growth factor receptor-2 (HER2) antibody herceptin. Mutants and pharmacological inhibitors of p38(MAPK) and MEK, but not phosphatidylinositol 3-kinase, suppressed both constitutive and HRG-induced stimulation of EF-1alpha promoter activity in MCF-7 cells. Deletion analysis of the promoter suggested the requirement of the -393 to -204 region for growth factor-mediated transcription of EF-1alpha. Fine mapping and point mutation studies revealed a role of the SP1 site in the observed HRG mediated regulation of the EF-1alpha promoter. In addition, we also provide new evidence to suggest that HRG stimulation of the EF-1alpha promoter involves increased physical interactions with acetylated histone H3 and histone H4. These results suggest that regulation of EF-1alpha expression by extracellular signals that function through human EGF receptor family members that are widely deregulated in human cancers and that growth factor regulation of EF-1alpha expression involve histone acetylation. PMID- 11104761 TI - Interfacial regulation of acid ceramidase activity. Stimulation of ceramide degradation by lysosomal lipids and sphingolipid activator proteins. AB - The lysosomal degradation of ceramide is catalyzed by acid ceramidase and requires sphingolipid activator proteins (SAP) as cofactors in vivo. The aim of this study was to investigate how ceramide is hydrolyzed by acid ceramidase at the water-membrane interface in the presence of sphingolipid activator proteins in a liposomal assay system. The degradation of membrane-bound ceramide was significantly increased both in the absence and presence of SAP-D when anionic lysosomal phospholipids such as bis(monoacylglycero)phosphate, phosphatidylinositol, and dolichol phosphate were incorporated into substrate bearing liposomes. Higher ceramide degradation rates were observed in vesicles with increased membrane curvature. Dilution assays indicated that acid ceramidase remained bound to the liposomal surface during catalysis. Not only SAP-D, but also SAP-C and SAP-A, were found to be stimulators of ceramide hydrolysis in the presence of anionic phospholipids. This finding was confirmed by cell culture studies, in which SAP-A, -C, and -D reduced the amount of ceramide storage observed in fibroblasts of a patient suffering from prosaposin deficiency. Strong protein-lipid interactions were observed for both SAP-D and acid ceramidase in surface plasmon resonance experiments. Maximum binding of SAP-D and acid ceramidase to lipid bilayers occurred at pH 4.0. Our results demonstrate that anionic, lysosomal lipids are required for efficient hydrolysis of ceramide by acid ceramidase. PMID- 11104762 TI - Phosphorylation of tau is regulated by PKN. AB - For the phosphorylation state of microtubule-associated protein, tau plays a pivotal role in regulating microtubule networks in neurons. Tau promotes the assembly and stabilization of microtubules. The potential for tau to bind to microtubules is down-regulated after local phosphorylation. When we investigated the effects of PKN activation on tau phosphorylation, we found that PKN triggers disruption of the microtubule array both in vitro and in vivo and predominantly phosphorylates tau in microtubule binding domains (MBDs). PKN has a catalytic domain highly homologous to protein kinase C (PKC), a kinase that phosphorylates Ser-313 (= Ser-324, the number used in this study) in MBDs. Thus, we identified the phosphorylation sites of PKN and PKC subtypes (PKC-alpha, -betaI, -betaII, gamma, -delta, -epsilon, -zeta, and -lambda) in MBDs. PKN phosphorylates Ser-258, Ser-320, and Ser-352, although all PKC subtypes phosphorylate Ser-258, Ser-293, Ser-324, and Ser-352. There is a PKN-specific phosphorylation site, Ser-320, in MBDs. HIA3, a novel phosphorylation-dependent antibody recognizing phosphorylated tau at Ser-320, showed immunoreactivity in Chinese hamster ovary cells expressing tau and the active form of PKN, but not in Chinese hamster ovary cells expressing tau and the inactive form of PKN. The immunoreactivity for phosphorylated tau at Ser-320 increased in the presence of a phosphatase inhibitor, FK506 treatment, which means that calcineurin (protein phosphatase 2B) may be involved in dephosphorylating tau at Ser-320 site. We also noted that PKN reduces the phosphorylation recognized by the phosphorylation-dependent antibodies AT8, AT180, and AT270 in vivo. Thus PKN serves as a regulator of microtubules by specific phosphorylation of tau, which leads to disruption of tubulin assembly. PMID- 11104763 TI - Regulation of the TAK1 signaling pathway by protein phosphatase 2C. AB - Protein phosphatase 2C (PP2C) is implicated in the negative regulation of stress activated protein kinase cascades in yeast and mammalian cells. In this study, we determined the role of PP2Cbeta-1, a major isoform of mammalian PP2C, in the TAK1 signaling pathway, a stress-activated protein kinase cascade that is activated by interleukin-1, transforming growth factor-beta, or stress. Ectopic expression of PP2Cbeta-1 inhibited the TAK1-mediated mitogen-activated protein kinase kinase 4 c-Jun amino-terminal kinase and mitogen-activated protein kinase kinase 6-p38 signaling pathways. In vitro, PP2Cbeta-1 dephosphorylated and inactivated TAK1. Coimmunoprecipitation experiments indicated that PP2Cbeta-1 associates with the central region of TAK1. A phosphatase-negative mutant of PP2Cbeta-1, PP2Cbeta-1 (R/G), acted as a dominant negative mutant, inhibiting dephosphorylation of TAK1 by wild-type PP2Cbeta-1 in vitro. In addition, ectopic expression of PP2Cbeta 1(R/G) enhanced interleukin-1-induced activation of an AP-1 reporter gene. Collectively, these results indicate that PP2Cbeta negatively regulates the TAK1 signaling pathway by direct dephosphorylation of TAK1. PMID- 11104764 TI - The C-terminal end of AraC tightly binds to the rest of its domain. AB - Genes were synthesized to express two DNA binding domains of AraC connected by short linkers. The abilities of the resulting proteins to bind to DNA containing AraC half-sites separated by the usual four bases as well as an additional two or three helical turns of the DNA were measured. The inability of some of the protein constructs to bind to widely separated half-sites indicates that the C terminal 14 amino acids of AraC are firmly bound to the rest of the DNA binding domain. PMID- 11104765 TI - The GLFG regions of Nup116p and Nup100p serve as binding sites for both Kap95p and Mex67p at the nuclear pore complex. AB - Our previous studies have focused on a family of Saccharomyces cerevisiae nuclear pore complex (NPC) proteins that contain domains composed of repetitive tetrapeptide glycine-leucine-phenylalanine-glycine (GLFG) motifs. We have previously shown that the GLFG regions of Nup116p and Nup100p directly bind the karyopherin transport factor Kap95p during nuclear protein import. In this report, we have further investigated potential roles for the GLFG region in mRNA export. The subcellular localizations of green fluorescent protein (GFP)-tagged mRNA transport factors were individually examined in yeast cells overexpressing the Nup116-GLFG region. The essential mRNA export factors Mex67-GFP, Mtr2-GFP, and Dbp5-GFP accumulated in the nucleus. In contrast, the localizations of Gle1 GFP and Gle2-GFP remained predominantly associated with the NPC, as in wild type cells. The localization of Kap95p was also not perturbed with GLFG overexpression. Coimmunoprecipitation experiments from yeast cell lysates resulted in the isolation of a Mex67p-Nup116p complex. Soluble binding assays with bacterially expressed recombinant proteins confirmed a direct interaction between Mex67p and the Nup116-GLFG or Nup100-GLFG regions. Mtr2p was not required for in vitro binding of Mex67p to the GLFG region. To map the Nup116-GLFG subregion(s) required for Kap95p and/or Mex67p association, yeast two-hybrid analysis was used. Of the 33 Nup116-GLFG repeats that compose the domain, a central subregion of nine GLFG repeats was sufficient for binding either Kap95p or Mex67p. Interestingly, the first 12 repeats from the full-length region only had a positive interaction with Mex67p, whereas the last 12 were only positive with Kap95p. Thus, the GLFG domain may have the capacity to bind both karyopherins and an mRNA export factor simultaneously. Taken together, our in vivo and in vitro results define an essential role for a direct Mex67p-GLFG interaction during mRNA export. PMID- 11104766 TI - Location of the polyamine binding site in the vestibule of the nicotinic acetylcholine receptor ion channel. AB - To map the structure of a ligand-gated ion channel, we used the photolabile polyamine-containing toxin MR44 as photoaffinity label. MR44 binds with high affinity to the nicotinic acetylcholine receptor in its closed channel conformation. The binding stoichiometry was two molecules of MR44 per receptor monomer. Upon UV irradiation of the receptor-ligand complex, (125)I-MR44 was incorporated into the receptor alpha-subunit. From proteolytic mapping studies, we conclude that the site of (125)I-MR44 cross-linking is contained in the sequence alpha His-186 to alpha Leu-199, which is part of the extracellular domain of the receptor. This sequence partially overlaps in its C-terminal region with one of the three loops that form the agonist-binding site. The agonist carbachol and the competitive antagonist alpha-bungarotoxin had only minor influence on the photocross-linking of (125)I-MR44. The site where the hydrophobic head group of (125)I-MR44 binds must therefore be located outside the zone that is sterically influenced by agonist bound at the nicotinic acetylcholine receptor. In binding and photocross-linking experiments, the luminal noncompetitive inhibitors ethidium and triphenylmethylphosphonium were found to compete with (125)I-MR44. We conclude that the polyamine moiety of (125)I-MR44 interacts with the high affinity noncompetitive inhibitor site deep in the channel of the nicotinic acetylcholine receptor, while the aromatic ring of this compound binds in the upper part of the ion channel (i.e. in the vestibule) to a hydrophobic region on the alpha-subunit that is located in close proximity to the agonist binding site. The region of the alpha-subunit labeled by (125)I-MR44 should therefore be accessible from the luminal side of the vestibule. PMID- 11104767 TI - Pseudo-T-even bacteriophage RB49 encodes CocO, a cochaperonin for GroEL, which can substitute for Escherichia coli's GroES and bacteriophage T4's Gp31. AB - Bacteriophage T4-encoded Gp31 is a functional ortholog of the Escherichia coli GroES cochaperonin protein. Both of these proteins form transient, productive complexes with the GroEL chaperonin, required for protein folding and other related functions in the cell. However, Gp31 is specifically required, in conjunction with GroEL, for the correct folding of Gp23, the major capsid protein of T4. To better understand the interaction between GroEL and its cochaperonin cognates, we determined whether the so-called "pseudo-T-even bacteriophages" are dependent on host GroEL function and whether they also encode their own cochaperonin. Here, we report the isolation of an allele-specific mutation of bacteriophage RB49, called epsilon22, which permits growth on the E. coli groEL44 mutant but not on the isogenic wild type host. RB49 epsilon22 was used in marker rescue experiments to identify the corresponding wild type gene, which we have named cocO (cochaperonin cognate). CocO has extremely limited identity to GroES but is 34% identical and 55% similar at the protein sequence level to T4 Gp31, sharing all of the structural features of Gp31 that distinguish it from GroES. CocO can substitute for Gp31 in T4 growth and also suppresses the temperature sensitive phenotype of the E. coli groES42 mutant. CocO's predicted mobile loop is one residue longer than that of Gp31, with the epsilon22 mutation resulting in a Q36R substitution in this extra residue. Both the CocO wild type and epsilon22 proteins have been purified and shown in vitro to assist GroEL in the refolding of denatured citrate synthase. PMID- 11104768 TI - Cell cycle regulation of the murine cdc25B promoter: essential role for nuclear factor-Y and a proximal repressor element. AB - Expression of the cdc25B gene is up-regulated late during cell cycle progression (S/G(2)). We have cloned the murine cdc25B promoter to identify elements involved in transcriptional regulation. A detailed structure-function analysis led to the identification of several elements that are located upstream of a canonical Inr motif at the site of transcription initiation and are involved in transcriptional activation and regulation. Activation of the promoter is largely mediated by NF-Y and Sp1/3 interacting with one and four proximal binding sites, respectively. In addition, NF-Y plays an essential role in cell cycle regulation in conjunction with a repressor element (cell cycle-regulated repressor) located approximately 30 nucleotides upstream of the putative Inr element and overlapping a consensus TATA motif. The cell cycle-regulated repressor is unrelated to the previously described cell cycle-regulated repressor elements. Taken together, our observations suggest that expression of the cdc25B gene is controlled through a novel mechanism of cell cycle-regulated transcription. PMID- 11104769 TI - The DNMT1 target recognition domain resides in the N terminus. AB - DNA-cytosine-5-methyltransferase 1 (DNMT1) is the enzyme believed to be responsible for maintaining the epigenetic information encoded by DNA methylation patterns. The target recognition domain of DNMT1, the domain responsible for recognizing hemimethylated CGs, is unknown. However, based on homology with bacterial cytosine DNA methyltransferases it has been postulated that the entire catalytic domain, including the target recognition domain, is localized to 500 amino acids at the C terminus of the protein. The N-terminal domain has been postulated to have a regulatory role, and it has been suggested that the mammalian DNMT1 is a fusion of a prokaryotic methyltransferase and a mammalian DNA-binding protein. Using a combination of in vitro translation of different DNMT1 deletion mutant peptides and a solid-state hemimethylated substrate, we show that the target recognition domain of DNMT1 resides in the N terminus (amino acids 122-417) in proximity to the proliferating cell nuclear antigen binding site. Hemimethylated CGs were not recognized specifically by the postulated catalytic domain. We have previously shown that the hemimethylated substrates utilized here act as DNMT1 antagonists and inhibit DNA replication. Our results now indicate that the DNMT1-PCNA interaction can be disrupted by substrate binding to the DNMT1 N terminus. These results point toward new directions in our understanding of the structure-function of DNMT1. PMID- 11104770 TI - Depletion of phosphatidylinositol 4,5-bisphosphate by activation of phospholipase C-coupled receptors causes slow inhibition but not desensitization of G protein gated inward rectifier K+ current in atrial myocytes. AB - G protein-gated inwardly rectifier K+ current in atrial myocytes (I(K(ACh))) upon stimulation with acetylcholine (ACh) shows a fast desensitizing component (t(1/2) approximately 5 s). After washout of ACh, I(K(ACh)) recovers from fast desensitization within < 30 s. A recent hypothesis suggests that fast desensitization is caused by depletion of phosphatidylinositol 4,5-bisphosphate (PtIns(4,5)P(2)), resulting from costimulation of phospholipase C (PLC)-coupled M3 receptors (M3AChR). The effects of stimulating two established PLC-coupled receptors, alpha-adrenergic and endothelin (ET(A)), on I(K(ACh)) were studied in rat atrial myocytes. Stimulation of these receptors caused activation of I(K(ACh)) and inhibition of the M2AChR-activated current. In myocytes loaded with GTPgammaS (guanosine 5'-3-O-(thio)triphosphate), causing stable activation of I(K(ACh)), inhibition via alpha-agonists and ET-1 was studied in isolation. Stimulation of either type of receptor under this condition, via G(q/11), caused a slow inhibition (t(1/2) approximately 50 s) by about 70%. No comparable effect on GTPgammaS-activated I(K(ACh)) was induced by ACh, suggesting that PLC-coupled M3AChRs are not functionally expressed in rat myocytes, which was supported by the finding that M3AChR transcripts were not detected by reverse transcriptase polymerase chain reaction in identified atrial myocytes. Supplementing the pipette solution with PtIns(4,5)P(2) significantly reduced inhibition of I(K(ACh)) but had no effect on fast desensitization. From these data it is concluded that stimulation of PLC-coupled receptors causes slow inhibition of I(K(ACh)) by depletion of PtIns(4,5)P(2), whereas fast desensitization of I(K(ACh)) is not related to PtIns(4,5)P(2) depletion. As muscarinic stimulation by ACh does not exert inhibition of I(K(ACh)) comparable to stimulation of alpha(1)- and ET(A) receptors, expression of functional PLC-coupled muscarinic receptors in rat atrial myocytes is unlikely. PMID- 11104771 TI - Contrasting localizations of MALS/LIN-7 PDZ proteins in brain and molecular compensation in knockout mice. AB - Proteins containing PDZ (postsynaptic density-95, discs large, zonula occludens) domains play a general role in recruiting receptors and enzymes to specific synaptic sites. In Caenorhabditis elegans, a complex of three PDZ proteins, LIN 2/7/10, mediates basolateral targeting of a receptor tyrosine kinase. Homologs of these LIN proteins have also been identified in higher organisms, and here we analyze the MALS/Veli (mammalian LIN-7/vertebrate homolog of LIN-7) proteins in brain. Immunohistochemical staining and in situ hybridization show that MALS occur differentially in discrete populations of neurons throughout the brain. Most neurons express only one MALS protein, although some cells contain two or even all three MALS isoforms. At the subcellular level, MALS proteins are found in both dendritic and axonal locations, suggesting that they may regulate processes at both pre- and postsynaptic sites. Targeted disruption of MALS-1 and MALS-2 does not yield a detectable phenotype, and hippocampal synaptic function and plasticity are intact in the MALS-1/2 double knockouts. Interestingly, MALS-3 protein is dramatically induced in the MALS-1/2 double knockouts, implying that dynamic changes in protein expression may play an important regulatory role for this family of synaptic PDZ proteins. PMID- 11104772 TI - Retinitis pigmentosa GTPase regulator (RPGRr)-interacting protein is stably associated with the photoreceptor ciliary axoneme and anchors RPGR to the connecting cilium. AB - Retinitis pigmentosa (RP) is a blinding retinal disease in which the photoreceptor cells degenerate. Mutations in the gene for retinitis pigmentosa GTPase regulator (RPGR) are a frequent cause of RP. The function of RPGR is not well understood, but it is thought to be a putative guanine nucleotide exchange factor for an unknown G protein. Ablation of the RPGR gene in mice suggested a role in maintaining the polarized distribution of opsin across the cilia. To investigate its function, we used a protein interaction screen to identify candidate proteins that may interact physiologically with RPGR. One such protein, designated RPGR-interacting protein (RPGRIP), is expressed specifically in rod and cone photoreceptors. It consists of an N-terminal region predicted to form coiled coil structures linked to a C-terminal tail that binds RPGR. In vivo, both proteins co-localize in the photoreceptor connecting cilia. RPGRIP is stably associated with the ciliary axoneme independent of RPGR and is resistant to extraction under conditions that partially solubilized other cytoskeletal components. When over-expressed in heterologous cell lines, RPGRIP appears in insoluble punctate and filamentous structures. These data suggest that RPGRIP is a structural component of the ciliary axoneme, and one of its functions is to anchor RPGR within the cilium. RPGRIP is the only protein known to localize specifically in the photoreceptor connecting cilium. As such, it is a candidate gene for human photoreceptor disease. The tissue-specific expression of RPGRIP explains why mutations in the ubiquitously expressed RPGR confer a photoreceptor specific phenotype. PMID- 11104773 TI - The role of dibasic residues in prohormone sorting to the regulated secretory pathway. A study with proneurotensin. AB - The mechanisms by which prohormone precursors are sorted to the regulated secretory pathway in neuroendocrine cells remain poorly understood. Here, we investigated the presence of sorting signal(s) in proneurotensin/neuromedin N. The precursor sequence starts with a long N-terminal domain followed by a Lys-Arg (neuromedin N)-Lys-Arg-(neurotensin)-Lys-Arg- sequence and a short C-terminal tail. An additional Arg-Arg dibasic is contained within the neurotensin sequence. Mutated precursors were expressed in endocrine insulinoma cells and analyzed for their regulated secretion. Deletion mutants revealed that the N-terminal domain and the Lys-Arg-(C-terminal tail) sequence were not critical for precursor sorting to secretory granules. In contrast, the Lys-Arg-(neuromedin N)-Lys-Arg (neurotensin) sequence contained essential sorting information. Point mutation of all three dibasic sites within this sequence abolished regulated secretion. However, keeping intact any one of the three dibasic sequences was sufficient to maintain regulated secretion. Finally, fusing the dibasic-containing C-terminal domain of the precursor to the C terminus of beta-lactamase, a bacterial enzyme that is constitutively secreted when expressed in neuroendocrine cells, resulted in efficient sorting of the fusion protein to secretory granules in insulinoma cells. We conclude that dibasic motifs within the neuropeptide domain of proneurotensin/neuromedin N constitute a necessary and sufficient signal for sorting proteins to the regulated secretory pathway. PMID- 11104774 TI - ADP is the cognate ligand for the orphan G protein-coupled receptor SP1999. AB - P2Y receptors are a class of G protein-coupled receptors activated primarily by ATP, UTP, and UDP. Five mammalian P2Y receptors have been cloned so far including P2Y1, P2Y2, P2Y4, P2Y6, and P2Y11. P2Y1, P2Y2, and P2Y6 couple to the activation of phospholipase C, whereas P2Y4 and P2Y11 couple to the activation of both phospholipase C and the adenylyl cyclase pathways. Additional ADP receptors linked to Galpha(i) have been described but have not yet been cloned. SP1999 is an orphan G protein-coupled receptor, which is highly expressed in brain, spinal cord, and blood platelets. In the present study, we demonstrate that SP1999 is a Galpha(i)-coupled receptor that is potently activated by ADP. In an effort to identify ligands for SP1999, fractionated rat spinal cord extracts were assayed for Ca(2+) mobilization activity against Chinese hamster ovary cells transiently transfected with SP1999 and chimeric Galpha subunits (Galpha(q/i)). A substance that selectively activated SP1999-transfected cells was identified and purified through a series of chromatographic steps. Mass spectral analysis of the purified material definitively identified it as ADP. ADP was subsequently shown to inhibit forskolin-stimulated adenylyl cyclase activity through selective activation of SP1999 with an EC(50) of 60 nM. Other nucleotides were able to activate SP1999 with a rank order of potency 2-MeS-ATP = 2-MeS-ADP > ADP = adenosine 5'-O-2 (thio)diphosphate > 2-Cl-ATP > adenosine 5'-O-(thiotriphosphate). Thus, SP1999 is a novel, Galpha(i)-linked receptor for ADP. PMID- 11104775 TI - Multiple regions of MAP kinase phosphatase 3 are involved in its recognition and activation by ERK2. AB - Mitogen-activated protein kinase phosphatase 3 (MKP3) is a specific regulator of extracellular signal-regulated protein kinase 2 (ERK2). Association of ERK2 with MKP3 results in a powerful increase in MKP3 phosphatase activity. To determine the molecular basis of the specific ERK2 recognition by MKP3 and the ERK2-induced MKP3 activation, we have carried out a systematic mutational and deletion analysis of MKP3. Using activation-based and competition-based assays, we are able to quantitatively evaluate the contributions that residues/regions within MKP3 make to ERK2 binding and ERK2-induced MKP3 activation. Our results show that recognition and activation of MKP3 by ERK2 involves multiple regions of MKP3. Thus, the kinase interaction motif (KIM; residues 61--75) in MKP3 plays a major role (135-fold) for high affinity ERK2 binding. The most important residue in the KIM sequence of MKP3 is Arg(65), which probably interacts with Asp(319) in ERK2. In addition to KIM, a unique sequence conserved in cytosolic MKPs (residues 161- 177 in MKP3) also contributes to ERK2 binding (15-fold). However, these two regions are not essential for ERK2-induced MKP3 activation. A third ERK2 binding site is localized in the C terminus of MKP3 (residues 348--381). Although deletion of this region or mutation of the putative ERK specific docking sequence (364)FTAP(367) in this region reduces MKP3's affinity for ERK2 by less than 10 fold, this region is absolutely required for ERK2-induced MKP3 activation. PMID- 11104776 TI - Protein kinase C activation modulates alpha-calmodulin kinase II binding to NR2A subunit of N-methyl-D-aspartate receptor complex. AB - The N-methyl-d-aspartate (NMDA) receptor subunits NR2 possess extended intracellular C-terminal domains by which they can directly interact with a large number of postsynaptic density (PSD) proteins involved in synaptic clustering and signaling. We have previously shown that PSD-associated alpha-calmodulin kinase II (alphaCaMKII) binds with high affinity to the C-terminal domain of the NR2A subunit. Here, we show that residues 1412-1419 of the cytosolic tail of NR2A are critical for alphaCaMKII binding, and we identify, by site directed mutagenesis, PKC-dependent phosphorylation of NR2A(Ser(1416)) as a key mechanism in inhibiting alphaCaMKII-binding and promoting dissociation of alphaCaMKII.NR2A complex. In addition, we show that stimulation of PKC activity in hippocampal slices either with phorbol esters or with the mGluRs specific agonist trans-1-amino-1,3- cyclopentanedicarboxylic acid (t-ACPD) decreases alphaCaMKII binding to NMDA receptor complex. Thus, our data provide clues on understanding the molecular basis of a direct cross-talk between alphaCaMKII and PKC pathways in the postsynaptic compartment. PMID- 11104777 TI - Structure of a multifunctional protein. Mammalian phosphatidylinositol transfer protein complexed with phosphatidylcholine. AB - Eukaryotic phosphatidylinositol transfer protein is a ubiquitous multifunctional protein that transports phospholipids between membrane surfaces and participates in cellular phospholipid metabolism during signal transduction and vesicular trafficking. The three-dimensional structure of the alpha-isoform of rat phosphatidylinositol transfer protein complexed with one molecule of phosphatidylcholine, one of its physiological ligands, has been determined to 2.2 A resolution by x-ray diffraction techniques. A single beta-sheet and several long alpha-helices define an enclosed internal cavity in which a single molecule of the phospholipid is accommodated with its polar head group in the center of the protein and fatty acyl chains projected toward the surface. Other structural features suggest mechanisms by which cytosolic phosphatidylinositol transfer protein interacts with membranes for lipid exchange and associates with a variety of lipid and protein kinases. PMID- 11104778 TI - Thermal and thermodynamic properties of duplex DNA containing site-specific interstrand cross-link of antitumor cisplatin or its clinically ineffective trans isomer. AB - The effect of the single, site-specific interstrand cross-link formed by cisplatin or transplatin on the thermal stability and energetics of a 20-base pair DNA duplex is reported. The cross-linked or unplatinated 20-base pair duplexes were investigated with the aid of differential scanning calorimetry, temperature-dependent UV absorption, and circular dichroism. The cross-link of both platinum isomers increases the thermal stability of the modified duplexes by changing the molecularity of denaturation. The structural perturbation resulting from the interstrand cross-link of cisplatin increases entropy of the duplex and in this way entropically stabilizes the duplex. This entropic cross-link-induced stabilization of the duplex is partially but not completely compensated by the enthalpic destabilization of the duplex. The net result of these enthalpic and entropic effects is that the structural perturbation resulting from the formation of the interstrand cross-link by cisplatin induces a decrease in duplex thermodynamic stability, with this destabilization being enthalpic in origin. By contrast, the interstrand cross-link of transplatin is enthalpically almost neutral with the cross-link-induced destabilization entirely entropic in origin. These differences are consistent with distinct conformational distortions induced by the interstrand cross-links of the two isomers. Importantly, for the duplex cross-linked by cisplatin relative to that cross-linked by transplatin, the compensating enthalpic and entropic effects almost completely offset the difference in cross-link-induced energetic destabilization. It has been proposed that the results of the present work further support the view that the impact of the interstrand cross-links of cisplatin and transplatin on DNA is different for each and might also be associated with the distinctly different antitumor effects of these platinum compounds. PMID- 11104779 TI - Characterization of phosphatidylserine transport to the locus of phosphatidylserine decarboxylase 2 in permeabilized yeast. AB - In yeast, nascent phosphatidylserine (PtdSer) can be transported to the mitochondria and Golgi/vacuole for decarboxylation to synthesize phosphatidylethanolamine (PtdEtn). In strains with a psd1Delta allele for the mitochondrial PtdSer decarboxylase, the conversion of nascent PtdSer to PtdEtn can serve as an indicator of lipid transport to the locus of PtdSer decarboxylase 2 (Psd2p) in the Golgi/vacuole. We have followed the metabolism of [(3)H]serine into PtdSer and PtdEtn to study lipid transport in permeabilized psd1Delta yeast. The permeabilized cells synthesize (3)H-PtdSer and, after a 20-min lag, decarboxylate it to form [(3)H]PtdEtn. Formation of [(3)H]PtdEtn is linear between 20 and 100 min of incubation and does not require ongoing PtdSer synthesis. PtdSer transport can be resolved into a two-component system using washed, permeabilized psd1Delta cells as donors and membranes isolated by ultracentrifugation as acceptors. With this system, the transport-dependent decarboxylation of nascent PtdSer is dependent upon the concentration of acceptor membranes, requires Mn(2+) but not nucleotides, and is inhibited by EDTA. High speed membranes isolated from a previously identified PtdSer transport mutant, pstB2, contain normal Psd2p activity but fail to reconstitute PtdSer transport and decarboxylation. Reconstitution with permutations of wild type and pstB2Delta donors and acceptors identifies the site of the mutant defect as the acceptor side of the transport reaction. PMID- 11104781 TI - Structural determinants of KvLQT1 control by the KCNE family of proteins. AB - KvLQT1 is a Shaker-like voltage-gated potassium channel that when complexed with minK (KCNE1) produces the slowly activating delayed rectifier I(ks). The emerging family of KCNE1-related peptides includes KCNE1 and KCNE3, both of which complex with KvLQT1 to produce functionally distinct currents. Namely I(ks), the slowly activating delayed rectifier current, is produced by KvLQT1/KCNE1, whereas KvLQT1/KCNE3 yields a more rapidly activating current with a distinct constitutively active component. We exploited these functional differences and the general structural similarities of KCNE1 and KCNE3 to study which physical regions are critical for control of KvLQT1 by making chimerical constructs of KCNE1 and KCNE3. By using this approach, we have found that a three-amino acid stretch within the transmembrane domain is necessary and sufficient to confer specificity of control of activation kinetics by KCNE1 and KCNE3. Moreover, chimera analysis showed that different regions within the transmembrane domain control deactivation rates. Our results help to provide a basis for understanding the mechanism by which KCNE proteins control K(+) channel activity. PMID- 11104782 TI - Unraveling the mechanism of action of thiazolidinediones. PMID- 11104783 TI - Vascular atrophy and VEGFR-2 signaling: old theories of pulmonary emphysema meet new data. PMID- 11104784 TI - Inhibition of VEGF receptors causes lung cell apoptosis and emphysema. AB - Pulmonary emphysema, a significant global health problem, is characterized by a loss of alveolar structures. Because VEGF is a trophic factor required for the survival of endothelial cells and is abundantly expressed in the lung, we hypothesized that chronic blockade of VEGF receptors could induce alveolar cell apoptosis and emphysema. Chronic treatment of rats with the VEGF receptor blocker SU5416 led to enlargement of the air spaces, indicative of emphysema. The VEGF receptor inhibitor SU5416 induced alveolar septal cell apoptosis but did not inhibit lung cell proliferation. Viewed by angiography, SU5416-treated rat lungs showed a pruning of the pulmonary arterial tree, although we observed no lung infiltration by inflammatory cells or fibrosis. SU5416 treatment led to a decrease in lung expression of VEGF receptor 2 (VEGFR-2), phosphorylated VEGFR-2, and Akt-1 in the complex with VEGFR-2. Treatment with the caspase inhibitor Z-Asp CH(2)-DCB prevented SU5416-induced septal cell apoptosis and emphysema development. These findings suggest that VEGF receptor signaling is required for maintenance of the alveolar structures and, further, that alveolar septal cell apoptosis contributes to the pathogenesis of emphysema. PMID- 11104785 TI - Stretch-induced alternative splicing of serum response factor promotes bronchial myogenesis and is defective in lung hypoplasia. AB - Smooth muscle (SM) develops only in organs and sites that sustain mechanical tensions. Therefore, we determined the role of stretch in mouse and human bronchial myogenesis. Sustained stretch induced expression of SM proteins in undifferentiated mesenchymal cells and accelerated the differentiation of cells undergoing myogenesis. Moreover, bronchial myogenesis was entirely controlled in lung organ cultures by the airway intraluminal pressure. Serum response factor (SRF) is a transcription factor critical for the induction of muscle-specific gene expression. Recently, a SRF-truncated isoform produced by alternative splicing of exon 5 has been identified (SRFDelta5). Here we show that undifferentiated mesenchymal cells synthesize both SRF and SRFDelta5 but that SRFDelta5 synthesis is suppressed during bronchial myogenesis in favor of increased SRF production. Stretch induces the same change in SRF alternative splicing, and its myogenic effect is abrogated by overexpressing SRFDelta5. Furthermore, human hypoplastic lungs related to conditions that hinder cell stretching continue to synthesize SRFDelta5 and show a marked decrease in bronchial and interstitial SM cells and their ECM product, tropoelastin. Taken together, our findings indicate that stretch plays a critical role in SM myogenesis and suggest that its decrease precludes normal bronchial muscle development. PMID- 11104786 TI - Induction of the chemokine stromal-derived factor-1 following DNA damage improves human stem cell function. AB - The chemokine stromal-derived factor-1 (SDF-1) controls many aspects of stem cell function. Details of its regulation and sites of production are currently unknown. We report that in the bone marrow, SDF-1 is produced mainly by immature osteoblasts and endothelial cells. Conditioning with DNA-damaging agents (ionizing irradiation, cyclophosphamide, and 5-fluorouracil) caused an increase in SDF-1 expression and in CXCR4-dependent homing and repopulation by human stem cells transplanted into NOD/SCID mice. Our findings suggest that immature osteoblasts and endothelial cells control stem cell homing, retention, and repopulation by secreting SDF-1, which also participates in host defense responses to DNA damage. PMID- 11104787 TI - Bleomycin-induced pulmonary fibrosis in fibrinogen-null mice. AB - Mice deleted for the plasminogen activator inhibitor-1 (PAI-1) gene are relatively protected from developing pulmonary fibrosis induced by bleomycin. We hypothesized that PAI-1 deficiency reduces fibrosis by promoting plasminogen activation and accelerating the clearance of fibrin matrices that accumulate within the damaged lung. In support of this hypothesis, we found that the lungs of PAI-1(-/-) mice accumulated less fibrin after injury than wild-type mice, due in part to enhanced fibrinolytic activity. To further substantiate the importance of fibrin removal as the mechanism by which PAI-1 deficiency limited bleomycin induced fibrosis, bleomycin was administered to mice deficient in the gene for the Aalpha-chain of fibrinogen (fib). Contrary to our expectation, fib(-/-) mice developed pulmonary fibrosis to a degree similar to fib(+/-) littermate controls, which have a plasma fibrinogen level that is 70% of that of wild-type mice. Although elimination of fibrin from the lung was not in itself protective, the beneficial effect of PAI-1 deficiency was still associated with proteolytic activity of the plasminogen activation system. In particular, inhibition of plasmin activation and/or activity by tranexamic acid reversed both the accelerated fibrin clearance and the protective effect of PAI-1 deficiency. We conclude that protection from fibrosis by PAI-1 deficiency is dependent upon increased proteolytic activity of the plasminogen activation system; however, complete removal of fibrin is not sufficient to protect the lung. PMID- 11104788 TI - An abnormal Ca(2+) response in mutant sarcomere protein-mediated familial hypertrophic cardiomyopathy. AB - Dominant-negative sarcomere protein gene mutations cause familial hypertrophic cardiomyopathy (FHC), a disease characterized by left-ventricular hypertrophy, angina, and dyspnea that can result in sudden death. We report here that a murine model of FHC bearing a cardiac myosin heavy-chain gene missense mutation (alphaMHC(403/+)), when treated with calcineurin inhibitors or a K(+)-channel agonist, developed accentuated hypertrophy, worsened histopathology, and was at risk for early death. Despite distinct pharmacologic targets, each agent augmented diastolic Ca(2+) concentrations in wild-type cardiac myocytes; alphaMHC(403/+) myocytes failed to respond. Pretreatment with a Ca(2+)-channel antagonist abrogated diastolic Ca(2+) changes in wild-type myocytes and prevented the exaggerated hypertrophic response of treated alphaMHC(403/+) mice. We conclude that FHC-causing sarcomere protein gene mutations cause abnormal Ca(2+) responses that initiate a hypertrophic response. These data define an important Ca(2+)-dependent step in the pathway by which mutant sarcomere proteins trigger myocyte growth and remodel the heart, provide definitive evidence that environment influences progression of FHC, and suggest a rational therapeutic approach to this prevalent human disease. PMID- 11104789 TI - Generation and phenotype of mice harboring a nonsense mutation in the V2 vasopressin receptor gene. AB - The V2 vasopressin receptor (V2R) plays a key role in the maintenance of a normal body water balance. To generate an in vivo model that allows the physiological and molecular analysis of the role of V2Rs in kidney function, we have created mouse lines that lack functional V2Rs by using targeted mutagenesis in mouse embryonic stem cells. Specifically, we introduced a nonsense mutation known to cause X-linked nephrogenic diabetes insipidus (XNDI) in humans (Glu242stop) into the mouse genome. V2R-deficient hemizygous male pups showed a decrease in basal urine osmolalities and were unable to concentrate their urine. These pups also exhibited an enlargement of renal pelvic space, failed to thrive, and died within the first week after birth due to hypernatremic dehydration. Interestingly, female mice heterozygous for the V2R mutation showed normal growth but displayed an XNDI-like phenotype, characterized by reduced urine concentrating ability of the kidney, polyuria, and polydipsia. Western blot analysis and immunoelectron microscopic studies showed that the loss of functional V2Rs had no significant effect on the basal expression levels of aquaporin-2 and the bumetanide-sensitive Na-K-2Cl cotransporter (BSC-1). The V2R mutant mice described here should serve as highly useful tools for the development of novel therapeutic strategies for the treatment of XNDI. PMID- 11104790 TI - Bromocriptine restores tolerance in estrogen-treated mice. AB - Estrogen can modulate autoimmunity in certain models of systemic lupus erythematosus. Recently, we have shown that it can mediate survival and activation of anti-DNA B cells in a mouse transgenic for the heavy chain of a pathogenic anti-DNA antibody. To identify whether estrogen effects reflect increased prolactin secretion, we characterized B-cell autoreactivity in transgenic mice given both bromocriptine (an inhibitor of prolactin secretion) and estradiol. Treatment of mice with estradiol plus bromocriptine led to reduced titers of anti-DNA antibodies and diminished IgG deposition in kidneys compared with treatment with estradiol alone. However, mice treated with estradiol plus bromocriptine showed an expansion of transgene-expressing B cells and enhanced Bcl-2 expression, similar to those of estradiol-treated mice. We identified anergic high-affinity anti-DNA B cells in mice treated with estradiol plus bromocriptine, and we showed by molecular analysis of anti-DNA hybridomas that their B cells derive from a naive repertoire. Thus, the estradiol-induced breakdown in B-cell tolerance can be abrogated by bromocriptine, which induces anergy in the high-affinity DNA-reactive B cells. These studies demonstrate that some of the effects of estrogen on naive autoreactive B cells require the presence of prolactin and, thus, suggest potential therapeutic interventions in lupus. PMID- 11104791 TI - Mutation of the WI-1 gene yields an attenuated blastomyces dermatitidis strain that induces host resistance. AB - Systemic fungal infections are becoming more common and difficult to treat, and vaccine prevention is not available. Pulmonary infection with the dimorphic fungus Blastomyces dermatitidis often progresses and requires treatment to prevent fatality. We recently created a recombinant strain of the fungus lacking the WI-1 adhesin and pathogenicity. We show here that administration of viable yeast of this attenuated strain vaccinates against lethal pulmonary experimental infection due to isogenic and nonisogenic strains from diverse geographic regions. To our knowledge, this is the first example of a recombinant attenuated vaccine against fungi. The vaccine induces delayed-type hypersensitivity and polarized type 1 cytokine responses, which are linked with resistance. A cell wall/membrane (CW/M) antigen from the vaccine strain also induces polarized and protective immune responses. Lymph node cells and CD4(+) T-cell lines raised with CW/M antigen transfer protective immunity when they release type 1 cytokine IFN gamma, but not when they release IL-4, and neutralization of IFN-gamma confirmed its role in vivo. Thus, by mutating a pathogenetic locus in a dimorphic fungus, we have created an attenuated vaccine strain and have begun to elucidate fungal and host elements requisite for vaccine immunity. PMID- 11104792 TI - Lack of angiotensin II-facilitated erythropoiesis causes anemia in angiotensin converting enzyme-deficient mice. AB - While nephrologists often observe reduced hematocrit associated with inhibitors of angiotensin-converting enzyme (ACE), the basis for this effect is not well understood. We now report that two strains of ACE knockout mice have a normocytic anemia associated with elevated plasma erythropoietin levels. (51)Cr labeling of red cells showed that the knockout mice have a normal total blood volume but a reduced red cell mass. ACE knockout mice, which lack tissue ACE, are anemic despite having normal renal function. These mice have increased plasma levels of the peptide acetyl-SDKP, a possible stem cell suppressor. However, they also show low plasma levels of angiotensin II. Infusion of angiotensin II for 2 weeks increased hematocrit to near normal levels. These data suggest that angiotensin II facilitates erythropoiesis, a conclusion with implications for the management of chronically ill patients on inhibitors of the renin-angiotensin system. PMID- 11104793 TI - Neutral endopeptidase inhibits prostate cancer cell migration by blocking focal adhesion kinase signaling. AB - Neutral endopeptidase 24.11 (NEP, CD10) is a cell-surface enzyme expressed by prostatic epithelial cells that cleaves and inactivates neuropeptides implicated in the growth of androgen-independent prostate cancer (PC). NEP substrates such as bombesin and endothelin-1 induce cell migration. We investigated the mechanisms of NEP regulation of cell migration in PC cells, including regulation of phosphorylation on tyrosine of focal adhesion kinase (FAK). Western analyses and cell migration assays revealed an inverse correlation between NEP expression and the levels of FAK phosphorylation and cell migration in PC cell lines. Constitutively expressed NEP, recombinant NEP, and induced NEP expression using a tetracycline-repressive expression system inhibited bombesin- and endothelin-1 stimulated FAK phosphorylation and cell migration. This results from NEP-induced inhibition of neuropeptide-stimulated association of FAK with cSrc protein. Expression of a mutated catalytically inactive NEP protein also resulted in partial inhibition of FAK phosphorylation and cell migration. Coimmunoprecipitation experiments show that NEP associates with tyrosine phosphorylated Lyn kinase, which then binds the p85 subunit of phosphatidylinositol 3-kinase (PI3-K) resulting in an NEP-Lyn-PI3-K protein complex. This complex competitively blocks FAK-PI3-K interaction, suggesting that NEP protein inhibits cell migration via a protein-protein interaction independent of its catalytic function. These experiments demonstrate that NEP can inhibit FAK phosphorylation on tyrosine and PC cell migration through multiple pathways and suggest that cell migration which contributes to invasion and metastases in PC cells can be regulated by NEP. PMID- 11104794 TI - Immunopathophysiological aspects of an emerging neonatal infectious disease induced by a bacterial superantigen. AB - We recently discovered an emerging neonatal infectious disease, neonatal toxic shock syndrome-like (TSS-like) exanthematous disease (NTED), which is induced by a superantigen, TSS toxin-1 (TSST-1), produced by methicillin-resistant Staphylococcus aureus (MRSA). Here, we analyzed the activation and the response of TSST-1-reactive Vss2(+) T cells in NTED patients during the acute and recovery phases and in asymptomatic infants exposed to MRSA. In the acute phase, Vss2(+) T cells were anergic to stimulation with TSST-1 and underwent marked expansion, but by 2 months after disease onset, their numbers had declined to about 10% of the control level. Although the percentage of Vss2(+) T cells in the ten asymptomatic neonatal MRSA carriers was within the control range, these individuals could be divided into two groups on the basis of Vss2(+) T-cell activation. Vss2(+)CD4(+) T cells from three of these infants (Group 1) highly expressed CD45RO and were anergic to TSST-1, whereas in the other seven asymptomatic neonatal MRSA carriers (Group 2), these cells expressed CD45RO at the control level and were highly responsive to stimulation with TSST-1. The serum anti-TSST-1 IgG Ab titer was negligible in the four NTED patients in the acute phase and the three asymptomatic neonatal MRSA carriers in Group 1, but it was high in the seven asymptomatic carriers in Group 2. We suggest that maternally derived anti-TSST-1 IgGs helps to suppress T-cell activation by TSST-1 and protects infants from developing NTED. PMID- 11104795 TI - Direct regulation of pituitary proopiomelanocortin by STAT3 provides a novel mechanism for immuno-neuroendocrine interfacing. AB - Neuroendocrine ACTH secretion responds to peripheral inflammatory and stress signals. We previously demonstrated that the proinflammatory cytokine, leukemia inhibitory factor (LIF), affects the hypothalamo-pituitary-adrenal axis (HPA) by stimulating in vitro and in vivo pituitary proopiomelanocortin (POMC) gene expression and ACTH secretion and by potentiating the action of hypothalamic corticotropin releasing hormone (CRH). Whereas pathways shown thus far to regulate POMC expression exclusively involve cAMP or calcium, we here describe a direct and indirect STAT3-dependent regulation of POMC transcription by LIF. Using progressive 5'-deletions of POMC promoter, we identified a LIF-responsive 407/-301 region that contains two juxtaposed sequences within -399/-379 related to a STAT3 DNA-binding motif. Each sequence within -399/-379 separately corresponds to a low-affinity and direct binding site for STAT3, but, in combination, these sequences bind STAT3 cooperatively and with high affinity. Moreover, LIF-activated STAT3 indirectly mediates LIF corticotroph action by inducing and potentiating CRH-induced c-fos and JunB expression and binding to the POMC AP-1 element. We therefore conclude that both a direct and indirect route mediate LIF-induced STAT3 activation of POMC transcription. Demonstration of STAT3-dependent regulation of the POMC gene represents a powerful mechanism for immuno-neuroendocrine interfacing and implies a direct stimulation of ACTH secretion by inflammatory and stress-derived STAT3-inducing cytokines. PMID- 11104796 TI - Cross-priming of naive CD8 T cells against melanoma antigens using dendritic cells loaded with killed allogeneic melanoma cells. AB - The goal of tumor immunotherapy is to elicit immune responses against autologous tumors. It would be highly desirable that such responses include multiple T cell clones against multiple tumor antigens. This could be obtained using the antigen presenting capacity of dendritic cells (DCs) and cross-priming. That is, one could load the DC with tumor lines of any human histocompatibility leukocyte antigen (HLA) type to elicit T cell responses against the autologous tumor. In this study, we show that human DCs derived from monocytes and loaded with killed melanoma cells prime naive CD45RA(+)CD27(+)CD8(+) T cells against the four shared melanoma antigens: MAGE-3, gp100, tyrosinase, and MART-1. HLA-A201(+) naive T cells primed by DCs loaded with HLA-A201(-) melanoma cells are able to kill several HLA-A201(+) melanoma targets. Cytotoxic T lymphocyte priming towards melanoma antigens is also obtained with cells from metastatic melanoma patients. This demonstration of cross-priming against shared tumor antigens builds the basis for using allogeneic tumor cell lines to deliver tumor antigens to DCs for vaccination protocols. PMID- 11104797 TI - Follicular B helper T cells express CXC chemokine receptor 5, localize to B cell follicles, and support immunoglobulin production. AB - Chemokines and their receptors have been identified as major regulators controlling the functional organization of secondary lymphoid organs. Here we show that expression of CXC chemokine receptor 5 (CXCR5), a chemokine receptor required for B cell homing to B cell follicles, defines a novel subpopulation of B helper T cells localizing to follicles. In peripheral blood these cells coexpress CD45RO and the T cell homing CC chemokine receptor 7 (CCR7). In secondary lymphoid organs, CD4(+)CXCR5(+) cells lose expression of CCR7, which allows them to localize to B cell follicles and germinal centers where they express high levels of CD40 ligand (CD40L), a costimulatory molecule required for B cell activation and inducible costimulator (ICOS), a recently identified costimulatory molecule of the CD28 family. Thus, when compared with CD4(+)CD45RO(+)CXCR5(-) cells, CD4(+)CD45RO(+)CXCR5(+) tonsillar T cells efficiently support the production of immunoglobulin (Ig)A and IgG. In contrast, analysis of the memory response revealed that long-lasting memory cells are found within the CD4(+)CD45RO(+)CXCR5(-) population, suggesting that CXCR5(+)CD4 cells represent recently activated effector cells. Based on the characteristic localization within secondary lymphoid organs, we suggest to term these cells "follicular B helper T cells" (T(FH)). PMID- 11104798 TI - CXC chemokine receptor 5 expression defines follicular homing T cells with B cell helper function. AB - Leukocyte traffic through secondary lymphoid tissues is finely tuned by chemokines. We have studied the functional properties of a human T cell subset marked by the expression of CXC chemokine receptor 5 (CXCR5). Memory but not naive T cells from tonsils are CXCR5(+) and migrate in response to the B cell attracting chemokine 1 (BCA-1), which is selectively expressed by reticular cells and blood vessels within B cell follicles. Tonsillar CXCR5(+) T cells do not respond to other chemokines present in secondary lymphoid tissues, including secondary lymphoid tissue chemokine (SLC), EBV-induced molecule 1 ligand chemokine (ELC), and stromal cell-derived factor 1 (SDF-1). The involvement of tonsillar CXCR5(+) T cells in humoral immune responses is suggested by their localization in the mantle and light zone germinal centers of B cell follicles and by the concomitant expression of activation and costimulatory markers, including CD69, HLA-DR, and inducible costimulator (ICOS). Peripheral blood CXCR5(+) T cells also belong to the CD4(+) memory T cell subset but, in contrast to tonsillar cells, are in a resting state and migrate weakly to chemokines. CXCR5(+) T cells are very inefficient in the production of cytokines but potently induce antibody production during coculture with B cells. These properties portray CXCR5(+) T cells as a distinct memory T cell subset with B cell helper function, designated here as follicular B helper T cells (T(FH)). PMID- 11104799 TI - Glycosylphosphatidylinositol anchors of Plasmodium falciparum: molecular characterization and naturally elicited antibody response that may provide immunity to malaria pathogenesis. AB - Induction of proinflammatory cytokine responses by glycosylphosphatidylinositols (GPIs) of intraerythrocytic Plasmodium falciparum is believed to contribute to malaria pathogenesis. In this study, we purified the GPIs of P. falciparum to homogeneity and determined their structures by biochemical degradations and mass spectrometry. The parasite GPIs differ from those of the host in that they contain palmitic (major) and myristic (minor) acids at C-2 of inositol, predominantly C18:0 and C18:1 at sn-1 and sn-2, respectively, and do not contain additional phosphoethanolamine substitution in their core glycan structures. The purified parasite GPIs can induce tumor necrosis factor alpha release from macrophages. We also report a new finding that adults who have resistance to clinical malaria contain high levels of persistent anti-GPI antibodies, whereas susceptible children lack or have low levels of short-lived antibody response. Individuals who were not exposed to the malaria parasite completely lack anti-GPI antibodies. Absence of a persistent anti-GPI antibody response correlated with malaria-specific anemia and fever, suggesting that anti-GPI antibodies provide protection against clinical malaria. The antibodies are mainly directed against the acylated phosphoinositol portion of GPIs. These results are likely to be valuable in studies aimed at the evaluation of chemically defined structures for toxicity versus immunogenicity with implications for the development of GPI-based therapies or vaccines. PMID- 11104800 TI - Migration and differentiation of autoreactive B-1 cells induced by activated gamma/delta T cells in antierythrocyte immunoglobulin transgenic mice. AB - Using normal and transgenic (Tg) mice, we have shown that peritoneal B-1 cells are activated by administration of cytokines or lipopolysaccharide and migrate to other lymphoid organs where they differentiate into antibody-secreting cells. However, little is known about the process of B-1 cell migration and differentiation in vivo. We developed a mouse line by crossing the antierythrocyte antibody Tg mice (HL mice) with TCR-gamma/delta Tg mice specific for a self-thymus leukemia (TL) antigen in the recombination activating gene (RAG)2(-/-) background. In the presence of the self-antigen, Tg gamma/delta T cells increased in number and manifested activated phenotypes. Peritoneal B-1 cells in these mice migrated into mesenteric lymph nodes and differentiated into autoantibody-secreting cells, resulting in strong autoimmune hemolytic anemia. Furthermore, transfer of RAG2(-/-) x HL bone marrow or peritoneal cells into the peritoneal cavity of RAG2(-/-) x TCR-gamma/delta Tg mice gave rise to donor derived B-1 cells in mesenteric lymph nodes, and these cells produced the autoantibody. Thus, this study demonstrates that the migration of B-1 cells and differentiation into the antibody-secreting cells can be induced by noncognate T cell help and implies the possibility that gamma/delta T cells may induce B-1 cell differentiation in vivo. PMID- 11104801 TI - Interferon gamma induction of pulmonary emphysema in the adult murine lung. AB - Chronic inflammation containing CD8(+) lymphocytes, neutrophils, and macrophages, and pulmonary emphysema coexist in lungs from patients with chronic obstructive pulmonary disease. Although this inflammatory response is believed to cause the remodeling that is seen in these tissues, the mechanism(s) by which inflammation causes emphysema have not been defined. Here we demonstrate that interferon gamma (IFN-gamma), a prominent product of CD8(+) cells, causes emphysema with alveolar enlargement, enhanced lung volumes, enhanced pulmonary compliance, and macrophage and neutrophil-rich inflammation when inducibly targeted, in a transgenic fashion, to the adult murine lung. Prominent protease and antiprotease alterations were also noted in these mice. They included the induction and activation of matrix metalloproteinase (MMP)-12 and cathepsins B, H, D, S, and L, the elaboration of MMP-9, and the selective inhibition of secretory leukocyte proteinase inhibitor. IFN-gamma causes emphysema and alterations in pulmonary protease/antiprotease balance when expressed in pulmonary tissues. PMID- 11104802 TI - Virulence factors of Helicobacter pylori responsible for gastric diseases in Mongolian gerbil. AB - Helicobacter pylori infection induces various gastroduodenal diseases. We examined the role of two genes, vacA and cagE, in the gastric pathogenesis induced by H. pylori using a long-term (62 wk) animal model. Reportedly, both genes are associated with the virulence of H. pylori: vacA encodes vacuolating cytotoxin, and cagE, with other genes in the cag pathogenicity islands, encodes a type IV secretion system. Mongolian gerbils were challenged in this study by a wild-type TN2 strain and its isogenic mutants of cagE or vacA. The wild-type and vacA mutants induced severe gastritis, whereas cagE mutants induced far milder changes. Gastric ulcer was induced at the highest rate (22/23) by the wild-type TN2, followed by the vacA mutant (19/28). No ulcer was found in the gerbils infected with the cagE mutant (0/27) or in controls (0/27). Intestinal metaplasia was also found in the gerbils infected with the wild-type (14/23) or vacA mutant (15/28). Gastric cancer developed in one gerbil with wild-type infection and in one with vacA mutant infection. In conclusion, the knocking out of the cagE gene deprived wild-type H. pylori of the pathogenicity for gastritis and gastric ulcer, suggesting that the secretion system encoded by cag pathogenicity island genes plays an essential role. PMID- 11104803 TI - Severe B cell deficiency in mice lacking the tec kinase family members Tec and Btk. AB - The cytoplasmic protein tyrosine kinase Tec has been proposed to have important functions in hematopoiesis and lymphocyte signal transduction. Here we show that Tec-deficient mice developed normally and had no major phenotypic alterations of the immune system. To reveal potential compensatory roles of other Tec kinases such as Bruton's tyrosine kinase (Btk), Tec/Btk double-deficient mice were generated. These mice exhibited a block at the B220(+)CD43(+) stage of B cell development and displayed a severe reduction of peripheral B cell numbers, particularly immunoglobulin (Ig)M(lo)IgD(hi) B cells. Although Tec/Btk(null) mice were able to form germinal centers, the response to T cell-dependent antigens was impaired. Thus, Tec and Btk together have an important role both during B cell development and in the generation and/or function of the peripheral B cell pool. The ability of Tec to compensate for Btk may also explain phenotypic differences in X-linked immunodeficiency (xid) mice compared with human X-linked agammaglobulinemia (XLA) patients. PMID- 11104804 TI - Short telomeres result in organismal hypersensitivity to ionizing radiation in mammals. AB - Here we show a correlation between telomere length and organismal sensitivity to ionizing radiation (IR) in mammals. In particular, fifth generation (G5) mouse telomerase RNA (mTR)(-/)- mice, with telomeres 40% shorter than in wild-type mice, are hypersensitive to cumulative doses of gamma rays. 60% of the irradiated G5 mTR(-/)- mice die of acute radiation toxicity in the gastrointestinal tract, lymphoid organs, and kidney. The affected G5 mTR(-/)- mice show higher chromosomal damage and greater apoptosis than similarly irradiated wild-type controls. Furthermore, we show that G5 mTR(-/)- mice show normal frequencies of sister chromatid exchange and normal V(D)J recombination, suggesting that short telomeres do not significantly affect the efficiency of DNA double strand break repair in mammals. The IR-sensitive phenotype of G5 mTR(-/)- mice suggests that telomere function is one of the determinants of radiation sensitivity of whole animals. PMID- 11104805 TI - Melanocyte destruction after antigen-specific immunotherapy of melanoma: direct evidence of t cell-mediated vitiligo. AB - Current strategies for the immunotherapy of melanoma include augmentation of the immune response to tumor antigens represented by melanosomal proteins such as tyrosinase, gp100, and MART-1. The possibility that intentional targeting of tumor antigens representing normal proteins can result in autoimmune toxicity has been postulated but never demonstrated previously in humans. In this study, we describe a patient with metastatic melanoma who developed inflammatory lesions circumscribing pigmented areas of skin after an infusion of MART-1-specific CD8(+) T cell clones. Analysis of the infiltrating lymphocytes in skin and tumor biopsies using T cell-specific peptide-major histocompatibility complex tetramers demonstrated a localized predominance of MART-1-specific CD8(+) T cells (>28% of all CD8 T cells) that was identical to the infused clones (as confirmed by sequencing of the complementarity-determining region 3). In contrast to skin biopsies obtained from the patient before T cell infusion, postinfusion biopsies demonstrated loss of MART-1 expression, evidence of melanocyte damage, and the complete absence of melanocytes in affected regions of the skin. This study provides, for the first time, direct evidence in humans that antigen-specific immunotherapy can target not only antigen-positive tumor cells in vivo but also normal tissues expressing the shared tumor antigen. PMID- 11104806 TI - Resistance of natural killer T cell-deficient mice to systemic Shwartzman reaction. AB - The generalized Shwartzman reaction in mice which had been primed and challenged with lipopolysaccharide (LPS) depends on interleukin (IL)-12-induced interferon (IFN)-gamma production at the priming stage. We examined the involvement in the priming mechanism of the unique population of Valpha14 natural killer T (NKT) cells because they promptly produce IFN-gamma after IL-12 stimulation. We report here that LPS- or IL-12-primed NKT cell genetically deficient mice were found to be resistant to LPS-elicited mortality. This outcome can be attributed to the reduction of IFN-gamma production, because injection of recombinant mouse IFN gamma, but not injection of IL-12, effectively primed the NKT cell-deficient mice. However, priming with high doses of LPS caused mortality of severe combined immunodeficiency, NKT cell-deficient, and CD1-deficient mice, indicating a major contribution of NKT cells to the Shwartzman reaction elicited by low doses of LPS, whereas at higher doses of LPS NK cells play a prominent role. These results suggest that the numerically small NKT cell population of normal mice apparently plays a mandatory role in the priming stage of the generalized Shwartzman reaction. PMID- 11104807 TI - Human malaria in immunocompromised mice: an in vivo model to study defense mechanisms against Plasmodium falciparum. AB - We have recently described that sustained Plasmodium falciparum growth could be obtained in immunodeficient mice. We now report the potential of this new mouse model by assaying the effect of the passive transfer of antibodies (Abs) which in humans have had a well-established effect.Our results show that the total African adult hyperimmune immunoglobulin Gs (HI-IgGs) strongly reduce P. falciparum parasitemia similarly to that reported in humans, but only when mice are concomitantly reconstituted with human monocytes (HuMNs). In contrast, neither HI IgGs nor HuMNs alone had any direct effect upon parasitemia. We assessed the in vivo effect of epitope-specific human Abs affinity-purified on peptides derived either from the ring erythrocyte surface antigen (RESA) or the merozoite surface protein 3 (MSP3). The inoculation of low concentrations of anti-synthetic peptide from MSP3, but not of anti-RESA Abs, consistently suppressed P. falciparum in the presence of HuMNs. Parasitemia decrease was stronger and faster than that observed using HI-IgGs and as fast as that induced by chloroquine. Our observations demonstrate that this mouse model is of great value to evaluate the protective effect of different Abs with distinct specificity in the same animal, a step hardly accessible and therefore never performed before in humans. PMID- 11104808 TI - Fas engagement induces the maturation of dendritic cells (DCs), the release of interleukin (IL)-1beta, and the production of interferon gamma in the absence of IL-12 during DC-T cell cognate interaction: a new role for Fas ligand in inflammatory responses. AB - Ligation of the Fas (CD95) receptor leads to an apoptotic death signal in T cells, B cells, and macrophages. However, human CD34(+)-derived dendritic cells (DCs) and mouse DCs, regardless of their maturation state, are not susceptible to Fas-induced cell death. This resistance correlates with the constitutive expression of the Fas-associated death domain-like IL-1beta-converting enzyme (FLICE)-inhibitory protein (FLIP) ligand. We demonstrate a new role of Fas in DC physiology. Engagement of Fas on immature DCs by Fas ligand (FasL) or by anti-Fas antibodies induces the phenotypical and functional maturation of primary DCs. Fas activated DCs upregulate the expression of the major histocompatibility complex class II, B7, and DC-lysosome-associated membrane protein (DC-LAMP) molecules and secrete proinflammatory cytokines, in particular interleukin (IL)-1beta and tumor necrosis factor alpha. Mature DCs, if exposed to FasL, produce even higher amounts of IL-1beta. Importantly, it is possible to reduce the production of IL 1beta and interferon (IFN)-gamma during DC-T cell interaction by blocking the coupling of Fas-FasL with a Fas competitor. Finally, during cognate DC-T cell recognition, IL-12 (p70) could not be detected at early or late time points, indicating that Fas-induced, IFN-gamma secretion is independent of IL-12. PMID- 11104809 TI - P-Selectin glycoprotein ligand 1 (PSGL-1) is a physiological ligand for E selectin in mediating T helper 1 lymphocyte migration. AB - P-selectin glycoprotein ligand 1 (PSGL-1) is a sialomucin expressed on leukocytes that mediates neutrophil rolling on the vascular endothelium. Here, the role of PSGL-1 in mediating lymphocyte migration was studied using mice lacking PSGL-1. In a contact hypersensitivity model, the infiltration of CD4(+) T lymphocytes into the inflamed skin was reduced in PSGL-1-deficient mice. In vitro-generated T helper (Th)1 cells from PSGL-1-deficient mice did not bind to P-selectin and migrated less efficiently into the inflamed skin than wild-type Th1 cells. To assess the role of PSGL-1 in P- or E-selectin-mediated migration of Th1 cells, the cells were injected into E- or P-selectin-deficient mice. PSGL-1-deficient Th1 cells did not migrate into the inflamed skin of E-selectin-deficient mice, indicating that PSGL-1 on Th1 cells is the sole ligand for P-selectin in vivo. In contrast, PSGL-1-deficient Th1 cells migrated into the inflamed skin of P selectin-deficient mice, although less efficiently than wild-type Th1 cells. This E-selectin-mediated migration of PSGL-1-deficient or wild-type Th1 cells was not altered by injecting a blocking antibody to L-selectin. These data provide evidence that PSGL-1 on Th1 cells functions as one of the E-selectin ligands in vivo. PMID- 11104810 TI - A soluble form of B cell maturation antigen, a receptor for the tumor necrosis factor family member APRIL, inhibits tumor cell growth. AB - A proliferation-inducing ligand (APRIL) is a ligand of the tumor necrosis factor (TNF) family that stimulates tumor cell growth in vitro and in vivo. Expression of APRIL is highly upregulated in many tumors including colon and prostate carcinomas. Here we identify B cell maturation antigen (BCMA) and transmembrane activator and calcium modulator and cyclophilin ligand (CAML) interactor (TACI), two predicted members of the TNF receptor family, as receptors for APRIL. APRIL binds BCMA with higher affinity than TACI. A soluble form of BCMA, which inhibits the proliferative activity of APRIL in vitro, decreases tumor cell proliferation in nude mice. Growth of HT29 colon carcinoma cells is blocked when mice are treated once per week with the soluble receptor. These results suggest an important role for APRIL in tumorigenesis and point towards a novel anticancer strategy. PMID- 11104811 TI - Follicular homing T helper (Th) cells and the Th1/Th2 paradigm. PMID- 11104812 TI - APRIL and BAFF connect autoimmunity and cancer. PMID- 11104813 TI - Probiotic bacteria as biological control agents in aquaculture. AB - There is an urgent need in aquaculture to develop microbial control strategies, since disease outbreaks are recognized as important constraints to aquaculture production and trade and since the development of antibiotic resistance has become a matter of growing concern. One of the alternatives to antimicrobials in disease control could be the use of probiotic bacteria as microbial control agents. This review describes the state of the art of probiotic research in the culture of fish, crustaceans, mollusks, and live food, with an evaluation of the results obtained so far. A new definition of probiotics, also applicable to aquatic environments, is proposed, and a detailed description is given of their possible modes of action, i.e., production of compounds that are inhibitory toward pathogens, competition with harmful microorganisms for nutrients and energy, competition with deleterious species for adhesion sites, enhancement of the immune response of the animal, improvement of water quality, and interaction with phytoplankton. A rationale is proposed for the multistep and multidisciplinary process required for the development of effective and safe probiotics for commercial application in aquaculture. Finally, directions for further research are discussed. PMID- 11104815 TI - Coupling of flagellar gene expression to flagellar assembly in Salmonella enterica serovar typhimurium and Escherichia coli. AB - How do organisms assess the degree of completion of a large structure, especially an extracellular structure such as a flagellum? Bacteria can do this. Mutants that lack key components needed early in assembly fail to express proteins that would normally be added at later assembly stages. In some cases, the regulatory circuitry is able to sense completion of structures beyond the cell surface, such as completion of the external hook structure. In Salmonella and Escherichia coli, regulation occurs at both transcriptional and posttranscriptional levels. One transcriptional regulatory mechanism involves a regulatory protein, FlgM, that escapes from the cell (and thus can no longer act) through a complete flagellum and is held inside when the structure has not reached a later stage of completion. FlgM prevents late flagellar gene transcription by binding the flagellum-specific transcription factor sigma(28). FlgM is itself regulated in response to the assembly of an incomplete flagellum known as the hook-basal body intermediate structure. Upon completion of the hook-basal body structure, FlgM is exported through this structure out of the cell. Inhibition of sigma(28) dependent transcription is relieved, and genes required for the later assembly stages are expressed, allowing completion of the flagellar organelle. Distinct posttranscriptional regulatory mechanisms occur in response to assembly of the flagellar type III secretion apparatus and of ring structures in the peptidoglycan and lipopolysaccharide layers. The entire flagellar regulatory pathway is regulated in response to environmental cues. Cell cycle control and flagellar development are codependent. We discuss how all these levels of regulation ensure efficient assembly of the flagellum in response to environmental stimuli. PMID- 11104814 TI - Molecular properties of bacterial multidrug transporters. AB - One of the mechanisms that bacteria utilize to evade the toxic effects of antibiotics is the active extrusion of structurally unrelated drugs from the cell. Both intrinsic and acquired multidrug transporters play an important role in antibiotic resistance of several pathogens, including Neisseria gonorrhoeae, Mycobacterium tuberculosis, Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa, and Vibrio cholerae. Detailed knowledge of the molecular basis of drug recognition and transport by multidrug transport systems is required for the development of new antibiotics that are not extruded or of inhibitors which block the multidrug transporter and allow traditional antibiotics to be effective. This review gives an extensive overview of the currently known multidrug transporters in bacteria. Based on energetics and structural characteristics, the bacterial multidrug transporters can be classified into five distinct families. Functional reconstitution in liposomes of purified multidrug transport proteins from four families revealed that these proteins are capable of mediating the export of structurally unrelated drugs independent of accessory proteins or cytoplasmic components. On the basis of (i) mutations that affect the activity or the substrate specificity of multidrug transporters and (ii) the three-dimensional structure of the drug-binding domain of the regulatory protein BmrR, the substrate-binding site for cationic drugs is predicted to consist of a hydrophobic pocket with a buried negatively charged residue that interacts electrostatically with the positively charged substrate. The aromatic and hydrophobic amino acid residues which form the drug-binding pocket impose restrictions on the shape and size of the substrates. Kinetic analysis of drug transport by multidrug transporters provided evidence that these proteins may contain multiple substrate-binding sites. PMID- 11104816 TI - Cytopathogenesis and inhibition of host gene expression by RNA viruses. AB - Many viruses interfere with host cell function in ways that are harmful or pathological. This often results in changes in cell morphology referred to as cytopathic effects. However, pathogenesis of virus infections also involves inhibition of host cell gene expression. Thus the term "cytopathogenesis," or pathogenesis at the cellular level, is meant to be broader than the term "cytopathic effects" and includes other cellular changes that contribute to viral pathogenesis in addition to those changes that are visible at the microscopic level. The goal of this review is to place recent work on the inhibition of host gene expression by RNA viruses in the context of the pathogenesis of virus infections. Three different RNA virus families, picornaviruses, influenza viruses, and rhabdoviruses, are used to illustrate common principles involved in cytopathogenesis. These examples were chosen because viral gene products responsible for inhibiting host gene expression have been identified, as have some of the molecular targets of the host. The argument is made that the role of the virus-induced inhibition of host gene expression is to inhibit the host antiviral response, such as the response to double-stranded RNA. Viral cytopathogenesis is presented as a balance between the host antiviral response and the ability of viruses to inhibit that response through the overall inhibition of host gene expression. This balance is a major determinant of viral tissue tropism in infections of intact animals. PMID- 11104817 TI - Positive and negative aspects of the human immunodeficiency virus protease: development of inhibitors versus its role in AIDS pathogenesis. AB - In this review we summarize multiple aspects of the human immunodeficiency virus (HIV) protease from both structural and functional viewpoints. After an introductory overview, we provide an up-to-date status report on protease inhibitors (PI). This proceeds from a discussion of PI structural design, to how PI are optimally utilized in highly active antiretroviral triple therapy (one PI along with two reverse transcriptase inhibitors), the emergence of PI resistance, and the natural role of secretory leukocyte PI. Then we switch to another focus: the interaction of HIV protease with other genes in acute and persistent infection, which in turn may have an effect on AIDS pathogenesis. We conclude with a discussion on future directions in HIV treatment, involving multiple target anti-HIV therapy, vaccine development, and novel reactivation-inhibitory reagents. PMID- 11104819 TI - Origin and evolution of the mitochondrial proteome. AB - The endosymbiotic theory for the origin of mitochondria requires substantial modification. The three identifiable ancestral sources to the proteome of mitochondria are proteins descended from the ancestral alpha-proteobacteria symbiont, proteins with no homology to bacterial orthologs, and diverse proteins with bacterial affinities not derived from alpha-proteobacteria. Random mutations in the form of deletions large and small seem to have eliminated nonessential genes from the endosymbiont-mitochondrial genome lineages. This process, together with the transfer of genes from the endosymbiont-mitochondrial genome to nuclei, has led to a marked reduction in the size of mitochondrial genomes. All proteins of bacterial descent that are encoded by nuclear genes were probably transferred by the same mechanism, involving the disintegration of mitochondria or bacteria by the intracellular membranous vacuoles of cells to release nucleic acid fragments that transform the nuclear genome. This ongoing process has intermittently introduced bacterial genes to nuclear genomes. The genomes of the last common ancestor of all organisms, in particular of mitochondria, encoded cytochrome oxidase homologues. There are no phylogenetic indications either in the mitochondrial proteome or in the nuclear genomes that the initial or subsequent function of the ancestor to the mitochondria was anaerobic. In contrast, there are indications that relatively advanced eukaryotes adapted to anaerobiosis by dismantling their mitochondria and refitting them as hydrogenosomes. Accordingly, a continuous history of aerobic respiration seems to have been the fate of most mitochondrial lineages. The initial phases of this history may have involved aerobic respiration by the symbiont functioning as a scavenger of toxic oxygen. The transition to mitochondria capable of active ATP export to the host cell seems to have required recruitment of eukaryotic ATP transport proteins from the nucleus. The identity of the ancestral host of the alpha-proteobacterial endosymbiont is unclear, but there is no indication that it was an autotroph. There are no indications of a specific alpha-proteobacterial origin to genes for glycolysis. In the absence of data to the contrary, it is assumed that the ancestral host cell was a heterotroph. PMID- 11104818 TI - Signal transduction cascades regulating fungal development and virulence. AB - Cellular differentiation, mating, and filamentous growth are regulated in many fungi by environmental and nutritional signals. For example, in response to nitrogen limitation, diploid cells of the yeast Saccharomyces cerevisiae undergo a dimorphic transition to filamentous growth referred to as pseudohyphal differentiation. Yeast filamentous growth is regulated, in part, by two conserved signal transduction cascades: a mitogen-activated protein kinase cascade and a G protein regulated cyclic AMP signaling pathway. Related signaling cascades play an analogous role in regulating mating and virulence in the plant fungal pathogen Ustilago maydis and the human fungal pathogens Cryptococcus neoformans and Candida albicans. We review here studies on the signaling cascades that regulate development of these and other fungi. This analysis illustrates both how the model yeast S. cerevisiae can serve as a paradigm for signaling in other organisms and also how studies in other fungi provide insights into conserved signaling pathways that operate in many divergent organisms. PMID- 11104822 TI - Effect of the aromatase inhibitor, MEN 11066, on growth of two different MCF-7 sublines. AB - The racemate compound MEN 11066 (1-[(benzofuran-2-yl)(4'-cyanophenyl)methyl]-1H 1,2,4-triazole) and its enantiomers, (+)-MEN 11623 and (-)-MEN 11622, showed potent and selective aromatase activity on human placental microsomes. In addition, to better evaluate their potency as anticancer drugs, the compounds were assayed on testosterone-induced cell proliferation to measure their ability in inhibiting oestrogen-dependent tumour growth. Two different sublines originated from the human breast carcinoma MCF-7 were used. One, named MCF 7(tumour aromatase) (TA), that had maintained its intrinsic aromatase activity, was more sensitive to estradiol or testosterone-induced growth than the second subline named MCF-7(human placental aromatase) (hPA). The latter had been transfected with the human placental aromatase cDNA, after recognizing that the parental cells had aromatase activity reduced to undetectable levels. The MEN compounds completely reverted the testosterone-induced proliferation in both MCF 7(TA) and MCF-7(hPA) cells, while they did not affect the estradiol-triggered proliferation as a proof of their specificity for aromatase enzyme. Interestingly, MCF-7(TA) cells were more susceptible to the effects of aromatase inhibitors than the MCF-7(hPA) cell. These data suggest the efficacy of aromatase inhibitors in breast cancer when the growth dependency from oestrogen is high and a relatively low aromatase activity may be extremely important for tumour development. PMID- 11104823 TI - Cocaine- and amphetamine-regulated transcript peptide-(55-102) and thyrotropin releasing hormone inhibit hypothalamic dopamine release. AB - Cocaine- and amphetamine-regulated transcript (CART) peptide-(55-102) and thyrotropin releasing hormone (TRH) play an anorectic role in the hypothalamus. Catecholamines are also involved in appetite control and we have previously found that leptin, an adipocyte-derived anorectic hormone, inhibits hypothalamic norepinephrine and dopamine release. We have studied the effect of CART peptide (55-102) and TRH on basal and depolarization (K+ 15 mM)-induced norepinephrine and dopamine release from rat hypothalamic neuronal endings (synaptosomes) in vitro. We have found that basal catecholamine release was not modified; both CART peptide-(55-102) and TRH, the former with a higher sensitivity, dose-dependently inhibited depolarization-induced dopamine release, and did not affect the stimulated norepinephrine release. Considering the role played by dopamine in the central mechanisms of reward, these findings suggest that the inhibition of dopamine release could underlie the decreased appetitive behaviour induced by CART peptide-(55-102) and TRH. PMID- 11104820 TI - Proteases for cell suicide: functions and regulation of caspases. AB - Caspases are a large family of evolutionarily conserved proteases found from Caenorhabditis elegans to humans. Although the first caspase was identified as a processing enzyme for interleukin-1beta, genetic and biochemical data have converged to reveal that many caspases are key mediators of apoptosis, the intrinsic cell suicide program essential for development and tissue homeostasis. Each caspase is a cysteine aspartase; it employs a nucleophilic cysteine in its active site to cleave aspartic acid peptide bonds within proteins. Caspases are synthesized as inactive precursors termed procaspases; proteolytic processing of procaspase generates the tetrameric active caspase enzyme, composed of two repeating heterotypic subunits. Based on kinetic data, substrate specificity, and procaspase structure, caspases have been conceptually divided into initiators and effectors. Initiator caspases activate effector caspases in response to specific cell death signals, and effector caspases cleave various cellular proteins to trigger apoptosis. Adapter protein-mediated oligomerization of procaspases is now recognized as a universal mechanism of initiator caspase activation and underlies the control of both cell surface death receptor and mitochondrial cytochrome c Apaf-1 apoptosis pathways. Caspase substrates have bene identified that induce each of the classic features of apoptosis, including membrane blebbing, cell body shrinkage, and DNA fragmentation. Mice deficient for caspase genes have highlighted tissue- and signal-specific pathways for apoptosis and demonstrated an independent function for caspase-1 and -11 in cytokine processing. Dysregulation of caspases features prominently in many human diseases, including cancer, autoimmunity, and neurodegenerative disorders, and increasing evidence shows that altering caspase activity can confer therapeutic benefits. PMID- 11104824 TI - Insulin increased cAMP phosphodiesterase activity antagonizing metabolic actions of glucagon in rat hepatocytes cultured with herbimycin A. AB - The baseline activity of cyclic nucleotide phosphodiesterase 4 was markedly lowered by primary culture of rat hepatocytes with herbimycin A for 4 h [Eur. J. Biochem. 260 (1999) 398-408.]. We now report that insulin added to this preparation of hepatocytes, which had been completely freed of herbimycin, increased the thus lowered phosphodiesterase activity, consequently antagonizing glucagon-induced production of cAMP and activation of glycogen phosphorylase. The insulin receptor beta-subunits and alpha-tubulin were tyrosine-phosphorylated upon the addition of insulin. The phosphorylation of alpha-tubulin afforded conditions unfavorable for microtubule assembly that is responsible for phosphodiesterase inhibition. These effects of insulin observed in herbimycin pretreated hepatocytes were not inhibited by wortmannin that actually abolished insulin-induced activation of phosphatidylinositol 3-kinase (PtdIns 3-kinase) under the same conditions. The physiological significance of the insulin action not mediated by PtdIns 3-kinase in herbimycin-pretreated hepatocytes is discussed. PMID- 11104825 TI - Apoptosis induced by droloxifene and c-myc, bax and bcl-2 mRNA expression in cultured luteal cells of rats. AB - Droloxifene is a tamoxifen derivative whose effects in the therapy of human breast cancer and postmenopausal osteoporosis have been studied widely. We had found that droloxifene could induce apoptosis of luteal cells of rat in vitro, but its mechanisms were unknown. In the present study, the expression of c-myc, bax and bcl-2 mRNA in cultured rat luteal cells during apoptosis induced by droloxifene was investigated and possible associations between these genes and apoptosis were analyzed. Cultured luteal cells of rats were incubated with droloxifene at various concentrations and with treatment durations. Occurrence of apoptosis was detected by terminal deoxyribonucleotidyl tranferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), DNA staining and DNA electrophoresis. Expression of these genes' mRNA was determined by semi quantitative reverse transcription-polymerase chain reaction (RT-PCR). The results showed that the c-myc and bax mRNA levels increased as concentrations or treatment durations of droloxifene increased, while the bcl-2 mRNA level exhibited no changes. A marked increase of c-myc and bax mRNA appeared respectively with 12 and 24 h of treatment, while a clear increase of apoptosis of luteal cells was found at 18 h. These results suggested that droloxifene could induce apoptosis of luteal cells of rat in vitro. The increase of c-myc mRNA expression might be one of the initiating factors and the elevated ratio of bax/bcl-2 mRNA was also probably involved in this effect. PMID- 11104826 TI - Nonpeptide neuromedin B receptor antagonists inhibit the proliferation of C6 cells. AB - The ability of nonpeptide antagonists to interact with neuromedin B receptors on C6 cells was investigated. 2-[3-(2, 6-Diisopropyl-phenyl)-ureido]3-(1H-indol-3 yl)-2-methyl-N-(1-pyridin- 2-yl-cyclohexylmethyl)-proprionate (PD165929), 3-(1H indol-3-yl)-2-methyl-2-[3(4-nitro-phenyl)-ureido]-N-(1-pyridin- 2-yl cyclohexylmethyl)-propionamide (PD168368) and 3-(1H-indol-3-yl)-N-[1-(5-methoxy pyridin-2-yl)-cyclohexylmethyl]- 2-m ethyl-2-[3-(4-nitro-phenyl)-ureido] propionamide (PD176252) inhibited (125I-Tyr0)neuromedin B binding with IC50 values of 2000, 40 and 50 nM, respectively. Because neuromedin B is a G-protein coupled serpentine receptor, the effects of neuromedin B antagonists on second messenger production and proliferation were investigated. PD168368 inhibited the ability of 10 nM neuromedin B to cause elevation of cytosolic Ca2+, whereas it had no effect on basal cytosolic Ca2+. PD168368 inhibited the ability of 100 nM neuromedin B to cause elevation of c-fos mRNA. Also, PD168368 in a dose-dependent manner inhibited the ability of 100 nM neuromedin B to cause phosphorylation of focal adhesion kinase. Using a [3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-2H tetrazolium bromide] assay, the order of antagonist potency to inhibit C6 proliferation was PD168368=PD176252>PD165929. Also, 1 microM PD168368 and PD176252 significantly inhibited colony number using a proliferation assay in vitro. PD168368 significantly inhibited C6 xenograft growth in nude mice in vivo. These results indicate that PD168368 is a C6 cell neuromedin B receptor antagonist, which inhibits proliferation. PMID- 11104821 TI - Microbial biofilms: from ecology to molecular genetics. AB - Biofilms are complex communities of microorganisms attached to surfaces or associated with interfaces. Despite the focus of modern microbiology research on pure culture, planktonic (free-swimming) bacteria, it is now widely recognized that most bacteria found in natural, clinical, and industrial settings persist in association with surfaces. Furthermore, these microbial communities are often composed of multiple species that interact with each other and their environment. The determination of biofilm architecture, particularly the spatial arrangement of microcolonies (clusters of cells) relative to one another, has profound implications for the function of these complex communities. Numerous new experimental approaches and methodologies have been developed in order to explore metabolic interactions, phylogenetic groupings, and competition among members of the biofilm. To complement this broad view of biofilm ecology, individual organisms have been studied using molecular genetics in order to identify the genes required for biofilm development and to dissect the regulatory pathways that control the plankton-to-biofilm transition. These molecular genetic studies have led to the emergence of the concept of biofilm formation as a novel system for the study of bacterial development. The recent explosion in the field of biofilm research has led to exciting progress in the development of new technologies for studying these communities, advanced our understanding of the ecological significance of surface-attached bacteria, and provided new insights into the molecular genetic basis of biofilm development. PMID- 11104827 TI - The CXCR4 agonist ligand stromal derived factor-1 maintains high affinity for receptors in both Galpha(i)-coupled and uncoupled states. AB - The alpha chemokine receptor CXCR4 and its only characterized chemokine ligand, stromal cell-derived factor-1 (SDF-1), are postulated to be important in the development of the B-cell arm of the immune system. In addition, CXCR4 is a critical coreceptor in support of viral entry by T-cell line tropic strains (X4) of the Human Immunodeficiency Virus Type 1 (HIV-1), viral variants which predominate in some infected individuals in end stage disease. SDF-1 can block X4 tropic HIV-1 infection of CD4+ target cells in vitro, and allelic variants of the human gene encoding SDF-1 in vivo correlate with delayed disease progression. Therefore, CXCR4 may be an appropriate target for therapeutic intervention in acquired immunodeficiency syndrome (AIDS), and knowledge of the pharmacology of SDF-1 binding to its cognate receptor will be important in the interpretation of these experiments. We report here a Kd derived using a competition binding assay of 4.5 nM for CXCR4 endogenously expressed on peripheral blood monocytes and T cells. This affinity is similar to that which SDF-1 exhibits when binding to endogenous CXCR4 on an established immortal Jurkat T-cell line as well as recombinant CXCR4 transfected into Chinese Hamster Ovary (CHO) cells. We also demonstrate that the determined affinity of SDF-1 for CXCR4 is reflective of its ability to induce a CXCR4-mediated signal transduction in these different cell types. Furthermore, using Bordetella pertussis toxin, we observe that high affinity binding of SDF-1 to CXCR4 is independent of the G-protein coupled state of the receptor, as uncoupling of G-protein did not lead to the appearance of measurable low affinity SDF-1 binding sites. Moreover, binding affinity and receptor number were unaffected by uncoupling for both recombinant and endogenously expressed CXCR4. Thus, SDF-1 is novel among agonist ligands of G protein-coupled receptors in that it appears to have equal affinity for both the G protein-coupled and uncoupled states of CXCR4. PMID- 11104828 TI - Inhibition of HgCl2-induced mitogen-activated protein kinase activation by LL Z1640-2 in CCRF-CEM cells. AB - Exposure of HgCl2 to CCRF-CEM human lymphoblastoid cells induced phosphorylation of mitogen-activated protein kinases (MAPKs); extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK) and p38. LL-Z1640-2, a macrocyclic nonaketide, inhibited HgCl2-induced JNK phosphorylation at 5-100 ng/ml. It also inhibited phosphorylation of ERK and p38 but only at 100 ng/ml. The same doses of radicicol did not suppress MAPKs activation. LL-Z1640-2 (at 100 ng/ml) inhibited HgCl2-induced JNK phosphorylation in NIH 3T3 fibroblasts but not in LLC-PK(1) renal epithelial cells. Thus, LL-Z1640-2 is a potent inhibitor of HgCl2-induced MAPKs activation, especially that of JNK, in CCRF-CEM cells. PMID- 11104829 TI - Characterisation of ATP-induced facilitation of transmission in rat hippocampus. AB - Superfusion of rat hippocampal slices with ATP induces a form of facilitation that has been poorly characterised. The present study has confirmed that at low concentrations of ATP (10 microM or less), an initial depression of evoked potential size is followed by a rebound facilitation which is not reproduced by alphabeta-methyleneATP, betagamma-methyleneATP, or the dinucleotide P1,P6 diadenosine hexaphosphate. The post-ATP facilitation could be prevented by the adenosine A1 receptor antagonists 8-phenyltheophylline or 1,3-dipropyl-8 cyclopentyltheophylline (50 nM), or superfusion of adenosine deaminase. The adenosine A2A receptor antagonist 8-(chlorostyryl)-caffeine did not affect the inhibition but prevented the post-ATP facilitation. The NMDA receptor antagonist 2-amino-5-phosphonopentanoic acid prevented the establishment of post-ATP facilitation. The post-ATP facilitation was also blocked by suramin at a concentration (50 microM) that does not block glutamate receptors. Suramin prevented the induction but not the maintenance phase of the post-ATP facilitation. The repeated induction of post-ATP facilitation by bursts of electrical stimulation designed to saturate the normal mechanisms of long-term potentiation prevented the induction of post-ATP facilitation. However, repeated applications of ATP to achieve saturation of its receptor did not prevent the subsequent induction of electrically evoked long-term potentiation. It is concluded that ATP can induce a form of synaptic facilitation which resembles only partially that induced by electrical stimulation and which may require the simultaneous activation of P1 and P2 receptors. PMID- 11104830 TI - The antinociceptive effect induced by FR140423 is mediated through spinal 5-HT2A and 5-HT3 receptors. AB - The involvement of 5-HT receptors in the antinociceptive effect of FR140423, 3 (difluoromethyl)-1-(4-methoxyphenyl)-5-[4-(methylsulfinyl)phenyl]py razole, was investigated in mice by means of the tail-pinch test. The antinociceptive effect of FR140423 injected i.t. was completely abolished by co-administration of the non-selective serotonin (5-hydroxytryptamine, 5-HT) receptor antagonist methysergide, the 5-HT2A receptor antagonist ketanserin and the 5-HT3 receptor antagonist MDL-72222 (3-tropanyl-3,5-dichlorobenzoate) but not by the 5-HT2B receptor antagonist SB-204741 (N-(1-methyl-5-indolyl)-N'-(3-methylisothiazol-5 yl)urea) or the 5-HT2C receptor antagonist SB-242084 (6-chloro-5-methyl-N-[6-(2 methylpyridin-3-yloxy)pyridin-3-y l]indolin e-1-carboxamine). The antinociceptive effect of FR140423 administered orally was abolished by i.t., but not by i.c.v., injection of methysergide, ketanserin and MDL-72222. These data indicate that FR140423, unlike morphine, exerts its antinociceptive effect against a mechanical noxious stimulus, such as in the tail-pinch test, by activation of spinal 5-HT2A and 5-HT3 receptors. PMID- 11104831 TI - Estimation of apparent pA2 values for WAY 100635 at 5-HT1A receptors regulating 5 hydroxytryptamine release in anaesthetised rats. AB - 5-HT1A receptor agonists decrease 5-hydroxytryptamine (5-HT) terminal release by activating somatodendritic 5-HT1A autoreceptors. The selective 5-HT1A receptor antagonist, N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohe xanecarboxamide (WAY 100635) inhibits these effects of 5-HT1A receptor agonists. The present study was aimed at estimating apparent pA2 values for WAY 100635 to antagonise 5-HT1A receptor agonist-induced decrease in 5-HT release in rat hippocampus. Extracellular concentrations of 5-HT were measured in microdialysis samples after administration of cumulative doses of 5-HT1A receptor agonists with different intrinsic activity, alone or in the presence of increasing doses of WAY 100635. Administration of cumulative doses of (+/-)-8-hydroxy-2-(di-n propylamino)tetralin (8-OH-DPAT) (0.01-40 mg/kg), 1[2-(4 fluorobenzoylamino)ethyl]-4-(7-methoxynaphtyl)pipe razine (S 14506) (0.00063-2.5 mg/kg), or buspirone (0.16-40 mg/kg), dose-dependently decreased the extracellular concentrations of 5-HT in the ventral hippocampus. Pre-treatment with WAY 100635 (0.01-0.63 mg/kg) shifted the dose-response curve of each agonist to the right in a dose-dependent manner. WAY 100635 antagonised the effects of all three compounds in a competitive manner, with an estimated apparent in vivo pA2 value of 7.95 (95% confidence limits: 7.66-8.24). Taken together, the results are evidence that buspirone, S 14506 and 8-OH-DPAT, administered in cumulative doses, decreased 5-HT release by activating similar 5-HT1A receptors, because a common apparent pA2 value was obtained for WAY 100635. The results also show that orderly microdialysis data can be obtained using cumulative dosing, which enables one to collect dose-response data rapidly, with fewer animals. PMID- 11104832 TI - Effects of lecithinized superoxide dismutase and a neutrophil elastase inhibitor (ONO-5046) on hyperoxic lung injury in rat. AB - Reactive oxygen and neutrophil metabolites have been implicated in the development of hyperoxic lung injury. We determined the protective effects of either a superoxide dismutase or neutrophil elastase inhibitor and the combination of both agents on the development of hyperoxic lung injury in rats. Two drugs (lecithinized superoxide dismutase and ONO-5046) were used in the present study. Lecithinized superoxide dismutase, a lecithin derivative bound to recombinant CuZn superoxide dismutase, has a higher affinity for cells such as polymorphonuclear leukocytes and endothelial cells than recombinant human superoxide dismutase. N-[2-[4-2,2-dimethylpropionyloxy) phenylsulfonylamino] benzoyl]? aminoacetic acid (ONO-5046), a specific neutrophil elastase inhibitor, which was developed as a low-molecular weight inhibitor, showed protective effects against various lung injuries. Rats were exposed to over 90% oxygen for 72 h, and bronchoalveolar lavage was performed to evaluate the permeability and neutrophil accumulation in the lungs. Rats were treated with lecithinized superoxide dismutase (30,000 U/day, intravenously n=7) or ONO-5046 (10 mg/kg, intramuscularly twice a day, n=7) or a combination of both drugs (n=7). Albumin concentration and neutrophil counts in bronchoalveolar lavage fluid were compared between animals with and without drug treatment. Either lecithinized superoxide dismutase or ONO-5046 treatment significantly decreased albumin concentration and neutrophil counts in bronchoalveolar lavage fluid compared to those in the animals of the hyperoxia-alone group (n=9). However, albumin leakage and neutrophil accumulation in the rat lung treated with combined agents were identical to that of either the lecithinized superoxide dismutase or ONO-5046 treatment. These findings suggest that lecithinized superoxide dismutase and ONO 5046 are useful drugs to protect against hyperoxic lung injury in rats. However, there were no additive effects by the combination in preventing hyperoxic lung injury. PMID- 11104833 TI - Peptide and non-peptide bradykinin B2 receptor agonists and antagonists: a reappraisal of their pharmacology in the guinea-pig ileum. AB - We have compared the pharmacology of different antagonists, Icatibant (H-DArg-Arg Pro-Hyp-Gly-Thi-Ser-DTic-Oic-Arg-OH), MEN 11270 (H-DArg-Arg-Pro-Hyp-Gly-Thi-c(Dab DTic-Oic-Arg)c(7 gamma-10 alpha)), and FR173657 ((E)-3-(6-acetamido-3-pyridyl)-N [N-[2, 4-dichloro-3-[(2-methyl-8-quinolinyl)oxymethyl]phenyl]-N-methyl aminocarbonylmethyl]acrylamide) at bradykinin B2 receptors expressed in the guinea-pig ileum by using bradykinin and the non-peptide FR190997 ((8-[2,6 dichloro-3-[N-[(E)-4-(N-methylcarbamoyl)cinnamidoacety l]-N methylamino]benzyloxy]-2-methyl-4-(2-pyridylmethoxy)quinoline) as agonists. In organ bath experiments, Icatibant and FR173657 exerted a non-competitive antagonism (pKB 9.5 and 9.2, respectively) of the contractile response to bradykinin, whereas MEN 11270 showed competitive antagonism (pKB 8.3, slope 0.90). The profile of action and apparent affinities of the three antagonists did not change if contact time was prolonged. The inhibition by the three antagonists of the contractile response to bradykinin was differently reverted by washout (MEN 11270 <30 min, Icatibant <60 min, FR173657 >60 min). The non-peptide ligand FR190997 acted as partial agonist if applied cumulatively to the bath (pD2 8.06, Emax 43% of maximal contractility), but as a full agonist when a maximally effective concentration was added (Emax 83%). FR173657 produced non-competitive antagonism of the response to FR190997 with apparent affinity similar to that measured toward bradykinin. On the contrary, Icatibant and MEN 11270 (300 nM both) competitively antagonized the contractile activity exerted by FR190997 with lower apparent pA2 value (6.9 and 7.2, respectively). In radioligand binding experiments, MEN 11270 and Icatibant displaced the [3H]bradykinin binding with pKi of 10.2 and 10.5 (Hill slope not different from unity), respectively. The non peptide ligands displaced the [3H]bradykinin binding with similar affinity, their pKi being 8.7 and 8.6 for FR173657 and FR190997, respectively (both Hill slopes <1). The present study indicates the difference in the antagonism type (competitive vs. non-competitive) by Icatibant, MEN 11270, and FR173657, as mainly ascribable to their different reversibility from the bradykinin B2 receptor, and affected by the kinetic of the response induced by the different agonists. Results are discussed in view of a different interaction of peptide and non-peptide agonist at the receptor. PMID- 11104834 TI - Pharmacological profile of YM-31636, a novel 5-HT3 receptor agonist, in vitro. AB - We investigated the in vitro pharmacological profile of YM-31636 (2-(1H-imidazol 4-ylmethyl)-8H-indeno[1,2-d]thiazole monofumarate). In cloned human 5-HT3A receptors, YM-31636 had a pKi value of 9.67 vs. ramosetron and pKi values for other 5-HT3 receptor agonists were less than 7. YM-31636 showed very low affinities for other receptors. YM-31636 induced contraction of isolated guinea pig distal colon. The intrinsic activity was approximately 0.90 compared with 5 hydroxytryptamine's (5-HT) 1.0, and the potency was 26 times greater than that of 5-HT. YM-31636 increased short-circuit current (Isc) in the isolated guinea pig distal colon. In this case, the relative intrinsic activity was approximately 0.19. In isolated guinea pig right atrium, YM-31636 induced tachycardia with the relative intrinsic activity of approximately 0.23. All these effects of YM-31636 were antagonized by ramosetron, a selective 5-HT3 receptor antagonist. These results suggest that YM-31636 is a potent and selective 5-HT3 receptor agonist, preferentially acting on the contraction of the colon. PMID- 11104835 TI - The carboxamide feG(NH2) inhibits endotoxin perturbation of intestinal motility. AB - The submandibular gland rat-1 (SMR1) salivary gland prohormone contains several peptides, submandibular gland peptide-T (SGP-T) and the tripeptide, FEG, which possess anti-inflammatory activities. The D-isomeric form of FEG, feG, also is a potent anti-inflammatory peptide. In this study, we compared the inhibitory activity of feG and its carboxamide derivative, feG(NH2), on the perturbations of intestinal motility induced by intravenous lipopolysaccharide. feG(NH2) was 20-30 times more potent than feG in reducing the motility disturbances induced by lipopolysaccharide. feG may undergo square-amidation to yield a hormone that strongly down-regulates intestinal responsiveness to endotoxin. PMID- 11104836 TI - Inhibition of small intestinal secretion by cannabinoids is CB1 receptor-mediated in rats. AB - We tested the hypothesis that cannabinoids, acting via a neuronal mechanism of action decrease small intestinal secretion. In vitro electrical stimulation induced ileal secretion in rats, that was attenuated by a cannabinoid receptor agonist, WIN 55212-2, (mesylate(R)-(+)-[2, 3-dihydro-5-methyl-3-[4 morpholino)methyl]pyrrolo-[1,2,3-de]-1, 4-benzoxazin-6-yl](1-naphthyl)methanone) but not its optical isomer WIN 55212-3. The inhibition of secretion induced by WIN 55212-2 was reversed by SR141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride), a cannabinoid CB1 receptor antagonist. An ileal secretory response stimulated by acetylcholine was unaffected by WIN 55212-2. These findings show that cannabinoids inhibit neurally mediated secretion via cannabinoid CB1 receptors. Thus, cannabinoids may have therapeutic potential for diarrhea unresponsive to available therapies. PMID- 11104837 TI - 3alpha-hydroxy-5alpha-pregnan-20-one exposure reduces GABA(A) receptor alpha4 subunit mRNA levels. AB - To examine the direct effects of neurosteroids on gamma-aminobutyric acid type A (GABA(A)) receptor expression, we exposed developing neuronal cells (P19) in vitro to 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha,5alpha-THP, allopregnanolone). Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed a concentration-dependent decrease in GABA(A) receptor alpha4 subunit mRNA expression that reversed 24 h after steroid withdrawal. These data suggest that variations in neurosteroid levels regulate the pattern of GABA(A) receptor subunit expression and may alter the trophic effects of GABA. PMID- 11104838 TI - Repetitive transcranial magnetic stimulation induces active coping strategies and attenuates the neuroendocrine stress response in rats. AB - The effects of repetitive transcranial magnetic stimulation (rTMS) on various brain functions were investigated in adult male Wistar rats. The stimulation parameters were adjusted according to the results of accurate computer-assisted, magnetic resonance imaging-based reconstructions of the current density distributions induced by rTMS in the rat and human brain, ensuring comparable stimulation patterns in both cases. The animals were subjected to daily rTMS treatment (three trains of 20 Hz; 2.5 s) for 8 weeks from the age of 4 weeks on. In the forced swim test these rats showed a more active stress coping strategy than the control rats. This was accompanied by a significantly attenuated stress induced elevation of plasma ACTH concentrations. Pituitary changes accounting for the attenuation were ruled out by the corticotropin-releasing hormone test. Baseline concentrations of ACTH and corticosterone were indistinguishable in the two groups. No changes were found in the anxiety-related behavior of the rats on the elevated plus-maze or in behavior during the social interaction test. Accordingly, the binding characteristics of the benzodiazepine agonist [(3)H]flunitrazepam at the benzodiazepine/gamma-aminobutyric acid type A receptor complex were similar in the rTMS and control groups. In summary, chronic rTMS treatment of frontal brain regions in rats resulted in a change in coping strategy that was accompanied by an attenuated neuroendocrine response to stress, thus revealing parallels to the effects of antidepressant drug treatment. PMID- 11104839 TI - Spatiotemporal gait patterns during over ground locomotion in major depression compared with healthy controls. AB - Alterations of locomotion are frequent, observable features of patients suffering from depression and have been investigated in these patients by actigraphy, cinematography and ground reaction forces. However, spatiotemporal parameters and neurophysiological mechanisms of gait have not yet been studied in depth in depression. The relationship between spatial and temporal parameters may yield insight into the pathophysiology of altered movements in depression. Therefore, gait patterns were quantitatively assessed and analysed in depressed subjects (n=16) and compared to matched healthy controls. Spatiotemporal gait parameters were measured during over ground walking at self-selected walking speed on a walkway previously validated in healthy subjects and used for orthopaedic and neurological patients. Compared to controls, depressed patients showed significantly lower gait velocity (p<0.001), reduced stride length (p<0.005), double limb support (p<0.005) and cycle duration (p<0.005). There was a significant correlation between cadence and gait velocity in depressed patients (r=0.51, p<0.05), but not in healthy controls (r=0.11, p>0.05). In patients with major depression, reduced gait velocity was associated with stride hypometria and increased cycle duration. Velocity was associated with cadence in depressed patients but not in healthy controls. The results may indicate possible deficiencies in the motor control system in depression. These first results about alterations of spatiotemporal gait patterns in depression warrant further longitudinal and experimental studies. PMID- 11104840 TI - DRD3 and DAT1 genes in schizophrenia: an association study. AB - OBJECTIVE: To investigate the role of the dopamine receptor 3 (DRD3) and transporter 1 (DAT1) genes in schizophrenia or in modulating its phenotype. METHODS: a Ser9Gly polymorphism in codon 9 of the DRD3 and a VNTR polymorphism in the DAT1genes were examined in two groups of schizophrenic patients, one of excellent neuroleptic responders (N=42) and one of nonresponders (N=64). A group of healthy volunteers screened for major psychiatric disorders was also included (N=89). In addition, age at onset of psychotic symptoms, attention performance and family loading for schizophrenia spectrum disorders were compared between patients with different genotypes in the DRD3 and DAT1 genes. RESULTS: No significant differences in the allelic distribution of the DRD3 and DAT1 polymorphisms were detected between schizophrenic patients and controls. A trend toward an excess of DRD3 genotype Gly/Gly was observed in neuroleptic nonresponder schizophrenic patients compared to controls (chi(2)=3. 30, df=1, p=0.07). No significant differences in age at onset of psychotic symptoms, attention task performance or family loading for schizophrenia spectrum disorders were observed between groups with different DRD3 and DAT1 genotypes. CONCLUSION: These results do not support the role of either of these genes in increasing susceptibility to schizophrenia or in modulating its phenotype in the studied population. PMID- 11104841 TI - Independence of sleep EEG responses to GABAergic hypnotics: biological implications. AB - GABAergic hypnotics are known to depress non-rapid eye movement delta and rapid eye movements and to stimulate non-rapid eye movement sigma (spindles) and beta EEG. This study addressed the question of whether the magnitudes of these effects are significantly correlated. Data were from a study in 16 normal subjects whose sleep was recorded for five nights under placebo and for three nights each under zolpidem (10 mg), triazolam (0.25 mg) and temazepam (30 mg). EEG was analyzed with both period-amplitude and power spectral (FFT) analysis. The magnitudes of the EEG and eye movement density responses were not significantly correlated for any of the three drugs. It is therefore unlikely that sleep responses to GABAergic drugs can be explained by the common cellular action (increased chloride conductance) of these drugs. We suggest that the sleep EEG responses are manifestations of complex (but consistent) interactions of excitation and inhibition in large brain systems although certain aspects of these responses (e.g. the different time courses of delta vs sigma and eye movement responses) may reflect molecular adaptations. A separate observation in this study was the strong traitlike characteristics of the sleep variables studied. These variables were highly correlated across nights of baseline sleep; in addition, individual differences in baseline sleep were significantly retained on the third night of temazepam administration. PMID- 11104842 TI - Low cholesterol and violent crime. AB - BACKGROUND: Community cohort studies and meta-analyses of randomized trials have shown a relation between low or lowered cholesterol and death by violence (homicide, suicide, accident); in primates, cholesterol reduction has been linked to increased behavioral acts of aggression (Kaplan J, Manuck S. The effects of fat and cholesterol on aggressive behaviour in monkeys. Psychosom. Med 1990;52:226-7; Kaplan J, Shively C, Fontenot D, Morgan T, Howell S, Manuck S et al. Demonstration of an association among dietary cholesterol, central serotonergic activity, and social behaviour in monkeys. Psychosom. Med 1994;56:479-84.). In this study we test for the first time whether cholesterol level is related to commission of violent crimes against others in a large community cohort. METHODS: We merged one-time cholesterol measurements on 79,777 subjects enrolled in a health screening project in Varmland, Sweden with subsequent police records for arrests for violent crimes in men and women aged 24 70 at enrollment; and with information on covariates. We performed a nested case control comparison of cholesterol in violent criminals - defined as those with two or more crimes of violence against others - to cholesterol in nonoffenders matched on age, enrollment year, sex, education and alcohol, using variable-ratio matching, with a nonparametric sign test. RESULTS: One hundred individuals met criteria for criminal violence. Low cholesterol (below the median) was strongly associated with criminal violence in unadjusted analysis (Men: risk ratio 1.94, P=0.002; all subjects risk ratio 2.32, P<0.001). Age emerged as a strong confounder. Adjusting for covariates using a matching procedure, violent criminals had significantly lower cholesterol than others identical in age, sex, alcohol indices and education, using a nonparametric sign test (P=0.012 all subjects; P=0.035 men). CONCLUSIONS: Adjusting for other factors, low cholesterol is associated with increased subsequent criminal violence. PMID- 11104843 TI - Evidence for anomalous lateralization across domain in ADHD children as well as adults identified with the Wender Utah rating scale. AB - Two studies assessed the relation between ADHD symptomatology and correlates of cerebral dominance. In the first, laterality was examined in school children (N=57), 28 with ADHD. Parental reports of greater attentional symptoms were related to non-righthandedness, but teacher reports were related to anomalous laterality of foot, ear and eye, as well as hand. This suggests that the previously reported association between ADHD and non-righthandedness may not be unique, but instead indicative of a more general condition of anomalous lateralization. This possibility was examined in study two, in which 234 undergraduates were assessed. As expected, the 26 adults identified by the Wender Utah Rating Scale (WURS) as retrospectively reporting more ADHD characteristics were found to be generally male. Also, in a replication of study one, enhanced WURS scores were associated with anomalous lateralization beyond handedness. In this case, ADHD characteristics were associated with a shift away from a right bias for hand, foot, and ear, but not eye. Factor analysis of the extensive Steenhuis and Bryden handedness questionnaire was then undertaken to determine whether all aspects of handedness, or only a subset, are associated with ADHD. The factor analysis indicated that the retrospective reports of ADHD characteristics were associated with only two of the three dimensions. Though limitations such as the gender composition of the groups in study one tempers the conclusions, the results of both studies support previous findings that ADHD is associated with anomalous laterality, but also indicate that non-righthandedness is not an adequate characterization of this relationship. PMID- 11104844 TI - fMRI of neuronal activation with symptom provocation in unmedicated patients with obsessive compulsive disorder. AB - BACKGROUND: Previous studies suggest that a neural circuit involving over activation of cortical, paralimbic, limbic, and striatal structures may underlie OCD symptomatology, but results may have been limited by medication use in those studies. To address this, we examined the effects of symptom induction on fMRI neural activation in medication-free patients with OCD. METHODS: Seven outpatients with OCD were exposed to individually tailored provocative and innocuous stimuli during fMRI scans. Self-ratings of OCD symptoms were performed prior to each scan and after exposure to stimuli. Images were analyzed as composite data sets and individually. RESULTS: Stimulus presentation was associated with significant increases in OCD self-ratings. Significant activation was demonstrated in several regions of the frontal cortex (orbitofrontal, superior frontal, and the dorsolateral prefrontal); the anterior, medial and lateral temporal cortex; and the right anterior cingulate. Right superior frontal activation inversely correlated with baseline compulsion symptomatology and left orbitofrontal cortical activation was inversely associated with changes in OCD self-ratings following provocative stimuli. CONCLUSIONS: These results in unmedicated patients are consistent with those from previous studies with medicated patients and suggest that OCD symptomatology is mediated by multiple brain regions including the anterior cingulate as well as frontal and temporal brain regions. PMID- 11104845 TI - The differential ACTH responses to combined dexamethasone/CRH administration in major depressive and dysthymic disorders. AB - In a preliminary study, we performed the combined dexamethasone/CRH test on patients with major depressive and dysthymic disorders as well as healthy controls. The ACTH response was significantly enhanced in the major depression group compared to the control group and tended to be heightened compared to the dysthymia group. The cortisol response was not significantly different among the three groups. We assume that major depression and dysthymia are neuroendocrinologically distinct disorders, although whether the difference is quantitative or qualitative remains to be examined. PMID- 11104846 TI - Apolipoprotein E epsilon4 and tardive dyskinesia in a Japanese population. AB - The findings that free radicals play a causative role in the occurrence of tardive dyskinesia (TD) and that apolipoprotein E (ApoE) 4 has decreased anti oxidant activity suggest a potential link between TD and ApoE alleles. We, therefore, examined ApoE allelic frequencies in schizophrenic subjects with TD and non-TD. Serum samples were obtained from 333 DSM IV-diagnosed schizophrenic patients and 191 controls in Japan. The presence of TD was evaluated by research diagnostic criteria for TD. ApoE phenotypes of the serum samples were determined by polyacrylamide gel isoelectricfocusing. A total of 62 TD subjects (31 males, 31 females) were identified among all patients examined. No significant differences in ApoE allelic frequency were found between TD and non-TD groups. ApoE epsilon4 allele frequency, however, was significantly lower in the female TD group than in the male TD group. These findings do not clearly demonstrate a certain association between TD and the epsilon4 allele, but may preliminarily reveal a difference in influence of this allele on the development of TD between males and females. PMID- 11104847 TI - Cognitive function in euthymic bipolar patients, stabilized schizophrenic patients, and healthy controls. AB - Studies on cognitive function in bipolar disorder have led to contrasting results and few data are available on affected subjects during the euthymic phase. In the present study we investigated the cognitive function of a cohort of bipolar (n=40) and schizophrenic (n=66) patients compared to healthy controls (n=64). Patients were evaluated in the outpatient setting over at least 3 months using a computerized version of Wisconsin Card Sorting Test. Schizophrenic patients showed the worst performance while that of the bipolar patients was somewhere between schizophrenic and controls. A discriminant analysis was able to classify correctly 60.59% of the subjects (schizophrenics 48.5%, bipolars 40%; healthy controls 85. 9%). The scores of the Wisconsin Card Sorting Test were entered into a principal component analysis, which yielded a 2-factor solution. Even in that analysis bipolar patients showed intermediate features in comparison with the other groups. These data indicate that bipolar patients have subtle neurocognitive deficits even after the resolution of an affective disorder. As well as observing quantitative differences between groups, the results show different dimensions of cognitive performance within groups suggesting that the deficit of euthymic bipolars could be a dishomogeneous entity, probably more heterogeneous than that in schizophrenia. Studies administering a more complete neuropsychological battery could further clarify the nature and meaning of the cognitive deficits in schizophrenia and bipolar disorder. PMID- 11104848 TI - Impaired eye expression recognition in schizophrenia. AB - Schizophrenia has been associated with abnormalities in recognising social emotions, inferring others' mental states and in gaze and visual scanning behaviours. Eye expression is known to convey considerable information in normal circumstances. Our study assessed the ability of individuals with schizophrenia to recognise simple and complex mental states from eye expressions alone. Sixteen individuals with a DSM-IV diagnosis of schizophrenia following SCAN [Schedules for Clinical Assessment in Neuropsychiatry (Wing JK, Babor T, Brugha T, Burke J, Cooper JE, Giel R et al. SCAN: schedules for clinical assessment in neuropsychiatry. Archives of General Psychiatry 1990;47:589-593).] interview and 16 healthy controls acted as participants. Expressions of 10 emotions (e.g. happy, afraid) and 10 complex mental states (e.g. thoughtful, bored), in the form of pictures of whole faces or eyes alone, were presented for recognition using a forced-choice response design. We observed impaired recognition of complex mental states in individuals with schizophrenia, from eye expressions alone (P=0.012). No differences in the recognition of basic emotions were detected. We also observed a negative correlation between illness chronicity and expression recognition performance (r=-0.65, P=0.006). The reduced ability of schizophrenia patients to recognise eye expressions of complex mental states could be interpreted as supporting a lack of "theory of mind". However, more parsimonious explanations based on impairments in basic recognition processes could also apply. An awareness of these processing abnormalities may have implications for future therapeutic strategies and our understanding of the pathophysiology of the disorder. PMID- 11104849 TI - Relationship between sex differences in onset of schizophrenia and puberty. AB - Some neurodevelopmental hypotheses of schizophrenia have postulated that sex differences in onset of illness could be explained by sexual dimorphism in onset of puberty, suggesting that early maturation accounts for the later onset of illness in women. The objective of this study was to analyse the relationship between age of menarche and age of onset of schizophrenia in a sample of Chilean patients. The medical records of 105 schizophrenic women diagnosed according to DSM-III-R criteria were studied. In all cases age of onset (first psychotic symptoms) and age of menarche were obtained. Pearson's correlation and student's t-test were used to analyse the data. The mean age of menarche in the sample of female patients (12. 98 years, S.D.=1.49) was significantly different from that of the general population of Santiago, Chile (12.53 years, S.D.=1.32) (t=2. 38; P<0.05). The mean age of onset of schizophrenia in female patients (19.92 years, S.D.=5.13) was significantly earlier in the Chilean sample than that reported in European and North American samples (P<0.05). No differences were observed when comparing the mean age at menarche. The subtypes with the earliest onset presented the earliest age of menarche and the subtypes with the latest onsets showed the latest ages at menarche. However, no correlation was observed between the age at onset of illness and the age at menarche, both in the total sample and in the analysis by subtype. The results of this study do not support a correlation between puberty and age of onset of illness. PMID- 11104850 TI - Neuromuscular and psychomotor abnormalities in patients with schizophrenia and their first-degree relatives. AB - In previous studies of schizophrenic patients, neuromuscular (histopathological and electrophysiological) and psychomotor (finger tapping) abnormalities were found. The present study was designed to investigate relationships between these abnormalities and a family history of psychosis in 14 schizophrenic patients and 25 unaffected first-degree relatives compared to 14 healthy controls. Muscle biopsies were performed in either m. tibialis anterior or m. lateralis. Macro EMG recordings were made from m. tibialis anterior. A finger tapping test was used to investigate psychomotor performance. Neuromuscular abnormalities (muscle biopsies and/or macro EMG) and/or aberrant psychomotor performance (finger tapping test) were found in 13 (93%) patients, 14 (56%) first-degree relatives and in three (21%) controls. A statistically significant relationship for the psychomotor, but not neuromuscular changes to a family history of psychosis was found using a logistic regression method. The percentage of patients, relatives and healthy controls exhibiting were 36/40/7% in the muscle biopsy, 50/20/0% in the macro EMG, and 71/82/14% in the finger tapping investigations. A higher frequency of neuromuscular and psychomotor abnormalities was found in patients with schizophrenia and their first-degree relatives compared to healthy controls. The relationship between psychomotor findings and a family history of psychosis indicate that central aspects of motor aberrations are associated with a hereditary disposition of psychosis. The neuromuscular as well as psychomotor changes indicate that schizophrenia may be a systemic disease involving the central nervous system as well as peripheral organs. An altered cell membrane is suggested to be an underlying factor based on the type of neuromuscular findings. PMID- 11104851 TI - Contingent negative variation and Dex-CRH test in patients with major depression. PMID- 11104852 TI - The HTR1B 861G>C receptor polymorphism among patients suffering from alcoholism, major depression, anxiety disorders and narcolepsy. AB - The HTR1B receptor gene has been linked to antisocial alcoholism in a Finnish population and an American Indian tribe [Lappalainen et al. , Arch. Gen. Psychiatry, 55 (1998) 989]. Using a candidate gene approach, we genotyped 209 patients with alcoholism, 108 patients with major depression, 32 patients with panic disorder, 50 patients with generalized anxiety disorder, 58 patients with narcolepsy and 74 healthy volunteers for the HTR1B 861G>C polymorphism. There was a higher frequency of the HTR1B 861G alleles among the alcohol-dependent patients as compared to the control subjects (chi(2)=4.02, d.f.=2, P=0.04). However, the association resulted from higher frequencies of the opposite alleles (HTR1B 861G), as originally reported by Lappalainen et al. (1998). Although the association in our study might be due to a type I error, the higher degree of HTR1B allele sharing within both populations could also argue for another alcoholism-relevant gene within the proximity of the HTR1B gene on human chromosome 6. PMID- 11104853 TI - Serotonergic markers and lowered plasma branched-chain-amino acid concentrations in fibromyalgia. AB - The aims of the present study were to examine serotonergic markers, i.e. [3H]paroxetine binding characteristics and the availability of plasma tryptophan, the precursor of serotonin (5-HT), and the plasma concentrations of the branched chain amino acids (BCAAs), valine, leucine and isoleucine, in fibromyalgia. The [3H]paroxetine binding characteristics, B(max) and K(d) values, and tryptophan and the competing amino acids (CAA), known to compete for the same cerebral uptake mechanism (i.e. valine, leucine, isoleucine, phenylalanine and tyrosine), were determined in fibromyalgia patients and normal controls. There were no significant differences in the [3H]paroxetine binding characteristics (B(max) and K(d)) between fibromyalgia and control subjects. There were no significant differences in plasma tryptophan or the tryptophan/CAA ratio between fibromyalgia patients and normal controls. In the fibromyalgia patients, there were no significant correlations between [3H]paroxetine binding characteristics or the availability of tryptophan and myalgic or depressive symptoms. Patients with fibromyalgia had significantly lower plasma concentrations of the three BCAAs (valine, leucine and isoleucine) and phenylalanine than normal controls. It is hypothesized that the relative deficiency in the BCAAs may play a role in the pathophysiology of fibromyalgia, since the BCAAs supply energy to the muscle and regulate protein synthesis in the muscles. A supplemental trial with BCAAs in fibromyalgia appears to be justified. PMID- 11104854 TI - A preliminary study of dehydroepiandrosterone response to low-dose ACTH in chronic fatigue syndrome and in healthy subjects. AB - Abnormalities of the production of dehydroepiandrosterone (DHEA), the adrenal androgen, have been linked with disorders such as obesity and psychological disorders such as major depression. Adrenocorticotropin (ACTH) is the primary stimulant of DHEA, and cortisol, from the adrenal. We chose to examine the DHEA and DHEA/cortisol response to the novel low-dose ACTH test in healthy subjects and a cohort with chronic fatigue syndrome (CFS): this test is useful in assessing subtle irregularities of pituitary-adrenal activity. Nineteen CFS subjects (diagnosed by CDC criteria) and 10 healthy subjects were examined. We demonstrated that 1 microg ACTH significantly elevates DHEA levels, with no difference in output between CFS and healthy subjects. The DHEA/cortisol ratio decreased in response to ACTH stimulation in healthy subjects but not in the CFS cohort. We suggest this divergence of response between the two groups represents an imbalance in the relative synthetic pathways of the CFS group which, if present chronically and if comparable to daily stressors, may manifest itself as an inappropriate response to stress. This difference may be important in either the genesis or propagation of the syndrome. PMID- 11104855 TI - The dimensionality of trauma: a multidimensional scaling comparison of police officers with and without posttraumatic stress disorder. AB - This study assesses the multidimensional structure of traumatic events as perceived by police officers and investigates individual differences in the scaling of such perceptions. Forty-two police officers with posttraumatic stress disorder (PTSD) and 40 officers without PTSD were given descriptions of critical incidents they were likely to encounter at work. They sorted these on the basis of similarity and rated them on 15 descriptive scales. The two groups were comparable in terms of relevant background characteristics. PTSD was diagnosed with the Structured Interview (SI-PTSD). The similarity data were subjected to individual differences multidimensional scaling analysis [Carroll and Chang, Psychometrika 35 (1970) 283]. The objective was, first, to identify the basic cognitive dimensions of psychological trauma that police officers use in discriminating between common critical incidents and, second, to test whether officers with and without PTSD apply such dimensions differently when interpreting critical incidents. The same three-dimensional solution was obtained for both groups: (1) emotional reactivity; (2) vulnerability and physical integrity; and (3) moral responsibility. Significant differences were found between the PTSD and non-PTSD groups in the salience of Dimension 2. Results are discussed with reference to other studies that address the meaning and interpretation of traumatic events. Implications for the conceptualization and assessment of trauma and PTSD are outlined. PMID- 11104857 TI - Emotional discomfort and impairments in verbal memory in schizophrenia. AB - Research has demonstrated that impairments in verbal memory in schizophrenia are linked with psychosocial deficits. Less is known, however, about their relationship to clinical features of illness. This study explores the hypothesis that impairments in verbal memory, particularly forms of memory requiring deeper levels of encoding, are uniquely linked to symptoms of dysphoria or emotional discomfort. Accordingly, we examined the association between concurrent measures of symptoms and verbal memory for 84 subjects with schizophrenia. Measures of positive, negative, cognitive, excitement and emotional discomfort symptoms were derived from factor scores of the Positive and Negative Syndrome Scale. Verbal memory was assessed using two tests requiring relatively superficial levels of encoding: The Hopkins Verbal Memory Test and the Digit Span subtest; and one test requiring deeper levels of encoding: the Logical Memory subtests I and II. As predicted, multiple regressions controlling for age, education and attention revealed that poorer performance on Logical Memory was strongly associated with greater levels of emotional discomfort (R(2)=0.22 and 0.25, respectively) while performance on the Hopkins test was related to cognitive symptoms scores (R(2)=0.08 and 0.09, respectively). Implications for the conceptualization of verbal memory deficits in schizophrenia are discussed. PMID- 11104856 TI - The clinical response to total sleep deprivation and recovery sleep in geriatric depression: potential indicators of antidepressant treatment outcome. AB - The clinical response to antidepressant treatment in late-life depression is often delayed and highly variable. Better indicators of antidepressant efficacy are needed early in the course of treatment, so that augmentation strategies or alternative treatments may be initiated. The goal of this study was to evaluate whether the change in the Hamilton depression rating scale (HDRS) after 36 h of total sleep deprivation (TSD) and recovery sleep predicted clinical outcome after 12 weeks of antidepressant treatment, and whether greater predictive value was observed in certain aspects of depressive symptomology. Fifteen elderly patients diagnosed with major depression underwent combined treatment with an initial 36 hours of TSD and a 12-week trial with the antidepressant paroxetine. Six HDRS subscores were evaluated with respect to how the changes after TSD and after one night of recovery sleep correlated with HDRS scores after 12 weeks of treatment. A significant correlation was obtained between the change in the core depressive symptomology subscale from baseline to recovery sleep and the HDRS score at 12 weeks, but the correlation was not significant when evaluating the change from baseline to TSD. These results indicate that the decrease in symptoms after recovery sleep compared with baseline levels (indicating the persistence of the antidepressant response), rather than the symptom reduction after TSD, has greater predictive value with respect to treatment outcome. PMID- 11104858 TI - A prospective longitudinal 10-year study of schizophrenia's three major factors and depression. AB - This study investigated the nature, independence, and stability of schizophrenia's syndrome factors and depression at 2, 4.5, 7.5 and 10 years post index hospitalization. At the four follow-ups, 71 patients (48 with schizophrenia and 23 with schizoaffective disorder) were assessed for symptoms hypothesized to constitute the reality distortion, disorganized, and negative factors of schizophrenia. At the last three follow-ups, the patients were also assessed for symptoms of depression. Factor analyses of schizophrenia symptoms revealed more than three syndrome factors at each follow-up. Longitudinally, reality distortion was a stable and relatively independent factor. The negative syndrome was independent but was bifurcated into two dimensions, interpreted as social/emotional withdrawal and diminished movement/expressiveness. Although signs of disorganization were not unified or independent early in schizophrenia's course, speech/thought disorder, disorganized affect, and poverty of speech content coalesced to form a disorganization factor by the 7.5-year follow-up. When depressive symptoms were added to the analyses, depression constituted an independent and stable dimension of schizophrenia over time. Each schizophrenia factor demonstrated a unique longitudinal course. Courses included stable symptom consistency (reality distortion), evolving symptom convergence (disorganization), and recurrent bifurcation and symptom instability (the negative syndrome). PMID- 11104860 TI - Chronic stable angina and its treatment; why Cinderella never gets to the ball? PMID- 11104859 TI - Treating problem-solving deficits on an acute care psychiatric inpatient unit. AB - Neuropsychological deficits in problem-solving are commonly found in patients with schizophrenia and severe affective disorders. However, in an acute care setting, treatment efforts do not typically target these deficits, even though they can impede recovery. This study aimed to evaluate the effectiveness of short term problem-solving remediation in acutely ill psychiatric inpatients. Twenty eight psychiatric inpatients identified as having a verbal problem-solving deficit received 6 h of either verbal problem-solving remediation or placebo instruction. Before and after treatment a nurse rated the patient's psychiatric status and the patient completed verbal and nonverbal problem-solving tests, and a self-report rating of symptoms and ability to cope with symptoms. Both groups of patients improved on the measure of verbal problem solving, but those receiving problem-solving remediation improved significantly more. Both groups made symptomatic improvement, but the patients receiving problem-solving remediation made significantly more improvement on the measure of coping ability and the nurses rated them as more improved, both psychiatrically and with regard to coping skills. Verbal problem-solving deficits are responsive to short-term remediation in an acute care setting, and treatment effects may generalize to improve ability to cope with psychiatric symptoms. PMID- 11104861 TI - Shuttle versus six-minute walk test in the prediction of outcome in chronic heart failure. AB - We prospectively analysed the potential usefulness of a symptom-limited walk test, the shuttle walk test (SWT), in the prediction of major cardiac events in 46 consecutive patients with chronic heart failure (NYHA class II-IV, ejection fraction <0.40) and compared it with that of a time-limited walk test, the six minute walk test (6-MT). After a mean follow-up of 17 months (range, 8-28 months), 15 of 46 patients (33%) experienced a major cardiac event, defined as a cardiac death, urgent transplantation, or hospital admission for continuous inotropic or mechanical support. Distance walked in the SWT was shown to be a predictor of outcome at one year of follow-up (P=0.03), but distance ambulated in the 6-MT was not (P=0.07). In multivariate analysis, percentage of age-gender predicted peak oxygen uptake was the best predictor of major cardiac events. When patients were divided into tertiles according to performance in both walk tests, there was an overall difference in event-free survival at 12 months among SWT tertiles (P=0.004), but not among 6-MT tertiles (P=0.09). A low performance in the SWT (distance ambulated <450 m) allowed identification of a subgroup of patients with a high risk for major cardiac events at short-term. We conclude that, in patients with chronic heart failure, distance ambulated in the SWT predicts event-free survival at one year better than that in the 6-MT. PMID- 11104862 TI - Low-cost shuttle walk test for assessing exercise capacity in chronic heart failure. PMID- 11104863 TI - Variable responsiveness of anterograde and retrograde fast pathway conduction to adenosine in patients with typical AV-nodal reentry tachycardia. AB - Adenosine is known as a substance which depresses predominantly the slow pathway of the av-node. However, the effect of adenosine on the anterograde and retrograde fast pathway (FP) has not been studied in a large patient population. Ninety-one patients with inducible typical av-nodal reentrant tachycardias (AVNRT) were included. The clinically used dosage of 12 mg adenosine was administered subsequently as bolus injection during a constant atrial and ventricular pacing (500 ms) in all patients. Electrophysiological av-nodal parameters were determined. A higher responsiveness of the anterograde compared to the retrograde FP was observed: the majority of patients (76%) blocked anterogradely and 55% blocked retrogradely within the FP after the administration of 12 mg adenosine. Thirty-six percent of all patients revealed a differential behaviour to adenosine. Sixteen percent of all patients were completely resistant to adenosine (P=0.012). Electrophysiological parameters did not predict the responsiveness of the FP to adenosine. In patients with typical AVNRT the anterograde FP shows a higher sensitivity than the retrograde FP to adenosine. This might reflect an anatomical and/or functional distinction between anterograde and retrograde FP. The variable response to adenosine could be due to individual anatomical and electrophysiological heterogenity of the perinodal tissue and the av-node. PMID- 11104864 TI - Heterogeneity of the fast pathway in AV nodal re-entrant tachycardia. An additional layer of complexity. PMID- 11104865 TI - Leptin serum levels in cachectic heart failure patients. Relationship with tumor necrosis factor-alpha system. AB - Cachexia is a strong predictor for mortality in patients with congestive heart failure. To investigate the role of leptin and regulators of apoptosis in cardiac cachexia we compared leptin concentrations and their relation to the TNF system, interleukin 1-beta (IL-1b), and soluble Fas in patients with heart failure with and without cachexia. Patients with cardiac cachexia have increased levels of interleukin-1b compared to non-cachectic heart failure patients [mean(S.E.)=1.11(0.62) vs. 0.02(0.02), P=0.01] and decreased concentrations of leptin [10.79(3.93) vs. 23.24 (8.35), P=0.1]. Leptin levels correlate with TNF-RI in cachectic heart failure patients (r=0.58, P=0.018). The TNF-RI levels were also correlated with Fas, both in all the patients taken together (r=0.5, P=0.006) and in those with cachexia (r=0.52, P=0.036). Our data indicate that more prospective studies are needed to clarify the role of leptin in the pathophysiology of heart failure cachexia. PMID- 11104866 TI - The significance of leptin in human--do we know it yet? PMID- 11104868 TI - Differential expression of three types of nitric oxide synthase in both infarcted and non-infarcted left ventricles after myocardial infarction in the rat. AB - In the present report we investigated the differential expression of three types of nitric oxide synthase (NOS) in the left ventricle after myocardial infarction in rats. One, 3, 7, 14, 28 and 56 days (n=6-12 for each group) after ligation of a coronary artery, tissue samples were obtained from infarcted and non-infarcted tissues. The mRNA and protein levels of neuronal (n) NOS, endothelial (e) NOS and inducible (i) NOS were sequentially determined by semi-quantitative reverse transcription-polymerase chain reaction and Western blotting. Progressive left ventricular dilatation and gradual reduction in fractional shortening were confirmed by echocardiography. The expression levels of nNOS were significantly increased 1, 3 and 7 days post-infarct compared to those of sham-operated rats in both the infarcted (P<0.01) and non-infarcted regions (P<0.01). Immunohistochemical analysis showed that nNOS was localized in nerve fibers in the left ventricle and that the number of positive fibers after myocardial infarction had increased compared to that in sham-operated rats. With regard to eNOS, no significant changes in expression levels were detected between infarcted hearts and sham-operated controls. The level of iNOS expression peaked three days post-infarct and then decreased in the infarcted tissue, whereas it increased one day post-infarct, peaked at 14 and 28 days post-infarct and was still elevated in the chronic stage in the ventricular septum. iNOS immunoreactivity was detected in spared cardiomyocytes and macrophages in the infarcted region, and in cardiomyocytes in the ventricular septum. The expressions of three types of NOS were differentially regulated and iNOS produced in the non-infarcted region may contribute to the progression of heart failure after myocardial infarction in rats. PMID- 11104867 TI - Inflammatory cytokines and the possible immunological role for lipoproteins in chronic heart failure. AB - AIMS: We studied the clinical and immunological importance of fasting cholesterol, HDL, LDL and triglycerides in patients with chronic heart failure in relation to plasma concentrations of tumor necrosis factor-alpha (TNFalpha), soluble TNF receptor-1 and -2 (sTNF-R1 and -R2), and a ratio potentially indicating recent endotoxin bioactivity (soluble [s] CD14/total cholesterol). METHODS AND RESULTS: Fifty-eight stable, non-oedematous patients with established heart failure and 19 controls were studied prospectively. Concentrations of sTNF R1 and sCD14 were higher in patients than in controls (1238+/-96 vs. 632+/-72 pg/ml, P=0.005 and 3401+/-120 vs. 2775+/-139 pg/ml, P=0.007, respectively), whereas those of TNFalpha (9.3+/-1.1 vs. 6.7+/-0.6 pg/ml) and sTNF-R2 (2464+/-145 vs. 1920+/-303 pg/ml) were not. Cholesterol (5.6+/-0.1 vs. 5.5+/-0.2 mmol/l) and LDL (3.5+/-0.1 vs. 3.6+/-0.2 mmol/l) were not different (both P>0.75). Patients had lower HDL (1.10+/-0.04 vs. 1.4+/-0.06 mmol/l, P=0.0004) and higher triglycerides (2.1+/-0.1 vs. 1.1+/-0.1 mmol/l, P=0.0006). Aetiology and the presence of cardiac cachexia did not influence the lipid profile. Correlations in patients: cholesterol vs. TNFalpha (r=-0.40, P=0.003), vs. sTNF-R1 (r=-0.24, P=0.08), vs. sTNF-R2 (r=-0.29, P<0.04); sCD14 vs. TNFalpha (r=0.44, P=0.005), vs. sTNF-R1: (r=0.65, P<0.0001), vs. sTNF-R2 (r=0.59, P<0. 0001). The sCD14/cholesterol ratio related powerfully to TNFalpha (r=0.60), sTNF-R1 (r=0.74), and sTNF-R2 (r=0.65, all P<0.0001). This sCD14/cholesterol ratio emerged as the strongest predictor of TNFalpha, sTNF-R1 and -R2 (all P<0.01), independently of renal and hepatic function, and conventional measures of disease severity. A cholesterol level <5.2 mmol/l (n=18) significantly predicted a poor clinical outcome (P<0.04, RR 3.5, 95% CI 1.1-11.0) independently of peak VO(2) (P=0.07), NYHA class (P=0.08), aetiology (P=0.14), and age, body wasting, sodium, LVEF, heart rate, and blood pressure (all P>0.20, follow-up 12 months, event rate 26%). CONCLUSION: Our data supports previous findings that lower, rather than higher cholesterol levels are associated with poor clinical outcome in patients with chronic heart failure. This relationship is unrelated to heart failure aetiology, and suggests that the classic risk profile is not longer relevant in established heart failure. The little-recognised ability of all lipoprotein fractions to bind endotoxin and to serve as natural buffer substances may explain this relationship between lower lipoprotein levels, higher cytokine concentrations and impaired prognosis. PMID- 11104869 TI - Short- and long-term risk factors for sudden death in patients with stable angina. AB - Sudden death is most common and often the first manifestation of coronary heart disease although its risk is difficult to predict. It has been studied mainly in patients with severe ventricular arrhythmia or recent myocardial infarction, but little is known about the different risk factors for short- and long-term risk of sudden death in patients with stable angina. To assess risk factors for sudden death in patients with stable angina and angiographically proven coronary artery disease, 319 consecutive patients were recruited prospectively and followed-up. Patients with clinical heart failure or recent myocardial infarction were excluded. Clinical, angiographic and biological variables were recorded. The association between each variable and the risk of sudden death was assessed in univariate and logistic multivariate analysis. There were 25 sudden deaths during the follow-up period (97+/-29 months). The univariate predictors in the short term (2 years) were: peripheral arterial disease, left ventricular hypertrophy, low density lipoprotein cholesterol and ejection fraction. The independent predictors were: peripheral arterial disease (relative risk: 6.3), ejection fraction (relative risk 1.05) and low density lipoprotein (relative risk: 1.8). In the long-term (8-10 years), body mass index, coronary score, ejection fraction and fibrinogen were univariate predictors. Only body mass index (relative risk: 1. 2), ejection fraction (relative risk: 1.06) and fibrinogen (relative risk: 2) remained independent predictors. The risk factors for sudden death in stable angina were time-dependent, peripheral arterial disease appeared as the best predictor with LDL for short time, and body mass index (obesity: index >27) and fibrinogen for long time. Ejection fraction was the only time-independent predictor. PMID- 11104870 TI - Reasons for adherence with antihypertensive medication. AB - BACKGROUND: Hypertension is often insufficiently controlled in clinical practice, a prominent reason for this being poor patient adherence with therapy. Little is known about the underlying reasons for poor adherence. We set out to investigate hypertensive patients' self-reported reasons for adhering to or ignoring medical advice regarding antihypertensive medication. METHODS: Qualitative analysis of semi-structured interviews with 33 hypertensive patients in a general-practice centre and a specialist hypertension unit in Southern Sweden. Blood-pressure measurements and laboratory measurements of antihypertensive medication were performed. RESULTS: Nineteen out of 33 patients were classified as adherent. Adherence was a function of faith in the physician, fear of complications of hypertension, and a desire to control blood pressure. Non-adherence was an active decision, partly based on misunderstandings of the condition and general disapproval of medication, but mostly taken in order to facilitate daily life or minimize adverse effects. Adherent patients gave less evidence of involvement in care than non-adherent patients. There was no obvious relation between reported adherence, laboratory markers of adherence and blood-pressure levels. CONCLUSIONS: The interview is a powerful tool for ascertaining patients' concepts and behaviour. To optimize treatment of hypertension, it is important to form a therapeutic alliance in which patients' doubts and difficulties with therapy can be detected and addressed. For this, effective patient-physician communication is of vital importance. PMID- 11104871 TI - Detection of mRNA encoding H(1) receptor and iNOS by RT-PCR in autoimmune myocarditis with special reference to changes in heart contractility. AB - Cardiac tissue from autoimmune myocarditis mice was studied to evaluate the expression and biological activity of mRNA encoding H(1) receptor and iNOS. BALB/c inbred mice were immunized with heart protein and sacrificed at 20, 45 and 50 days post immunization. Heart contractility studies and RT-PCR assays were performed. Heart from autoimmune myocarditis mice show mRNA iNOS-related dysfunction with a decrease in heart contractility. This effect was accompanied with an increase production of cyclic GMP and was improved by treating autoimmune mice with an inhibitor of iNOS activity. In addition, autoimmune myocardium expressed an active histamine H(1) receptor mRNA coupled to phospholipase C. The activation of H(1) receptor by ThEA stimulated both phosphoinositide hydrolysis and heart contractility. Normal myocardium did not expressed neither iNOS mRNA nor H(1) receptor mRNA. In conclusions: the development of autoimmune cardiac dysfunction was associated with the expression of iNOS mRNA, cyclic GMP accumulation and the expression of an active histamine H(1) receptor mRNA with increase production of inositol phosphates. These protein emergence during the course of autoimmune myocarditis may be involved a distinct compensatory mechanism operating in this disease. PMID- 11104872 TI - Vectorcardiographic changes during cardioangiography with iodixanol and ioxaglate. AB - AIM: To compare the electrophysiological effects of two contrast media (CM), the non-ionic dimer iodixanol and the ionic dimer ioxaglate using computerised dynamic vectorcardiography (VCG) during coronary angiography. METHODS: The study was designed as a double-blind, three-period crossover, randomised comparison between iodixanol (320 mg I/ml) and ioxaglate (320 mg I/ml). Group 1 (HVV) received ioxaglate (H) in the first injection in the left coronary artery (LCA) and iodixanol (V) in the following injections. Group 2 (VHH) received iodixanol in the first injection in LCA and ioxaglate in the following injections. The first three injections in the LCA were subjected to electrocardiographic analysis. RESULTS: For five out of six VCG variables, there was a significant difference in response between iodixanol and ioxaglate. For these five variables, the deviations from baseline were greater in the ioxaglate than in the iodixanol group (P<0.05). The most pronounced effects from ioxaglate were seen on the ST segment and T-wave. CONCLUSIONS: Iodixanol caused less pronounced electrophysiological changes than ioxaglate, especially during the repolarisation phase. Vectorcardiography is a sensitive and reproducible technique for detecting electrophysiological effects induced by CM. PMID- 11104873 TI - Absent right precordial R waves are associated with reduced left ventricular function and poor prognosis in patients with aortic stenosis. AB - Right precordial Q waves can be present in patients with aortic stenosis as well as in those with anterior myocardial infarction. In order to evaluate the relationship of right precordial Q waves to left ventricular function and prognosis in patients with aortic stenosis, we studied 49 such patients with no history of myocardial infarction, by means of ECG, clinical history and echocardiography. 15 (31%) patients had Q waves in both V1 and V2 and 34 (69%) did not. There were no differences in age (77+/-9.0 years vs. 78+/-9.7), follow up time (15+/-9.0 months vs. 18+/-10), gender (female:male 8:7 vs. 15:19), aortic valve gradient on Doppler (70.0+/-20 mmHg vs. 71+/-20) and left ventricular mass (360+/-118 g vs. 320+/-80) between the two groups (all P=NS). Left ventricular shortening fraction (22+/-9.0% vs. 28+/-8.5, P<0.05), ejection fraction (51+/-15% vs. 62+/-12, P<0.01) and circumferential fibre shortening (0.8+/-0.3 circ/s vs. 1.0+/-0.3, P<0.0s) were all significantly reduced in patients with right precordial Q waves compared to those without. During a mean follow-up of 1.5 years, 9 out of 15 (60%) patients with right precordial Q waves died compared with only 5 out of 34 (15%) patients with a normal QRS pattern died (P<0.01). In summary, a right precordial QS ECG pattern is present in nearly 1/3 patients with aortic stenosis and is associated with impaired left ventricular systolic function and adverse prognosis. PMID- 11104874 TI - The effects of potassium-ATP channel modulation on ventricular fibrillation and defibrillation in the pig heart. AB - BACKGROUND: Drugs acting on the cardiac ATP-sensitive potassium (K-ATP) channels may modulate responses to ischaemia and arrhythmogenesis. We investigated the effects of K-ATP channel modulation on frequency patterns of ventricular fibrillation (VF) and on defibrillation threshold (DFT). METHODS AND RESULTS: Each group of 24 pigs randomly received intravenous levcromakalim (LKM) 40 microgram/kg (K-ATP agonist), glibenclamide (Glib) 20 mg/kg (K-ATP antagonist), saline or vehicle. Firstly, QTc interval was measured before and after drug. VF was then induced by endocardial stimulation and its power spectra and dominant frequencies over 15 min determined by fast Fourier transformation. Secondly, transthoracic DFT was determined (step-up/step-down protocol) before and after each drug. LKM reduced QTc interval (e.g., lead II, 354-321 ms, P<0.05) and increased the dominant VF frequency between 6 and 8 min (9.5+/-0.5 Hz at 6.5 min compared with 7.2+/-0.6 Hz (saline), 7.4+/-0.8 Hz (vehicle), 6.8+/-0.5 Hz (Glib), P=0.03). LKM reduced (to 57.2+/-2.1 mmHg) and Glib increased (to 107.8+/-6.1) mean arterial BP compared with saline (80.3+/-5.6) and vehicle (87. 6+/-7.1; P<0.01). There was no significant difference in defibrillation threshold energy, current or voltage, after any drug. CONCLUSIONS: Activation of K-ATP channels reduced blood pressure and QTc interval. The lack of major effect on VF dominant frequency and DFT of either LKM or Glib suggests that prior administration of similar drugs to patients should not prejudice outcome from VF cardiac arrest. PMID- 11104875 TI - Amiodarone therapy for sustained ventricular tachycardia after myocardial infarction: long-term follow-up, risk assessment and predictive value of programmed ventricular stimulation. AB - We determine the value of the programmed ventricular stimulation (PVS) and of clinical, angiographic and electrophysiologic variables in assessing the long term risk of arrhythmia recurrence in a group of coronary artery diseased patients presenting with a first episode of monomorphic sustained ventricular tachycardia (VT) treated with amiodarone. Mortality and arrhythmia recurrence rates were retrospectively assessed in 55 consecutive patients with previous myocardial infarction presenting with a first VT episode. Results of left heart catheterization, echocardiography and time-domain signal-averaging were collected. Patients underwent PVS after amiodarone oral loading and were classified according to inducibility before being all discharged on amiodarone (200 mg daily). The mean follow-up was 42+/-31 months. Total and cardiac mortality rates were 29% (16 patients) and 23% (13 patients) respectively. Sudden death (SD) occurred in nine patients (16%). VT recurred in 13 patients (23%). Sustained monomorphic VT was inducible in 40 patients (72%) after amiodarone loading. Neither total mortality (10/40 vs. 6/15) nor cardiac mortality (3/40 vs. 1/15) were significantly different between inducible and non-inducible patients. Recurrent VT rate was 27% (11/40 patients) for the inducible group and 13% (2/15 patients) for the non-inducible group (NS). SD occurred in 6/40 inducible patients (15%) and in 2/15 non-inducible patients (13%) (NS). Arrhythmic events occurred in 42% (17/40) inducible patients vs. 26% (4/15) non-inducible patients (P=0.07). Parameters correlated with outcome were ejection fraction (EF) (5 SD/11 patients with EF <0.3 vs. 4/44 with EF >0.3, P=0.003), mitral insufficiency (MI) (4 SD/10 patients with MI vs. 4/44 patients without MI, P=0.004) and age (65+/-9 years for patients with VT recurrence vs. 58+/-9, P=0.02). Although the risk stratification can be improved, reliable and safe long-term prediction of recurrence of malignant ventricular arrhythmia in individual patients cannot be made. Consequently, the systematic implantation of a cardioverter-defibrillator in case of a first episode of sustained VT occurring in coronary artery disease patients should be further debated. PMID- 11104876 TI - Hyperhomocysteinaemia and adverse events complicating coronary catheter interventions. AB - BACKGROUND: Since hyperhomocysteinaemia is an independent risk factor for development of atherosclerosis as well as for arterial and venous thrombosis we investigated whether elevated homocysteine levels are associated with procedural excess risk which complicates coronary interventions including coronary angioplasty (PTCA), stenting, or directional coronary atherectomy (DCA). DESIGN: Consecutive cases receiving coronary catheter interventions. SETTING: Tertiary referral centre in Germany. METHODS: Fasting total plasma homocysteine levels (tHcy) were determined in 648 consecutive coronary artery disease patients who underwent catheter interventions (272 PTCA, 102 DCA, and 274 stenting). Hyperhomocysteinaemia was defined as tHcy >/=15 micromol/l. The patients were investigated for a 30-day composite endpoint, including need for target-vessel revascularization, myocardial infarction, and death. RESULTS: Among the 648 patients, 78 (12%) demonstrated elevated tHcy levels. The composite endpoint occurred in 41 patients (6.3%). For the entire intervention group there was no evidence that hyperhomocysteinaemia was associated with excess procedural risk (odds ratio [OR]: 1.27; 95% confidence interval [CI]=0.52 to -3.13; P=0.62). In further analyses according to device, hyperhomocysteinaemia also failed to predict complications in the device related subgroups. CONCLUSION: The results indicate that hyperhomocysteinaemia is not a major risk factor for 30-day adverse events complicating PTCA, DCA, or stenting. PMID- 11104877 TI - Influence of vessel size, age and body mass index on the flow-mediated dilatation (FMD%) of the brachial artery. AB - BACKGROUND: The non-invasive determination of the endothelial dysfunction (ED) of the brachial artery is a widely used method in clinical research. It remained, however, unclear, whether the test-results are influenced by the anatomical vessel size, the patients age, body mass index (BMI) or gender. METHODS: The flow mediated vasodilatation (FMD%) of the brachial artery was determined in 122 consecutive (88 male, 34 female) patients. FMD% was measured using high resolution ultrasound (13 Mhz) at rest, during reactive hyperaemia and after the sublingual administration of glycerolnitrate (GTN%). RESULTS: Lumen diameters at rest varied from 2.48 mm to 6.33 mm (4.46+/-0.74 mm). The extent of FMD% as well as of GTN% showed an inverse correlation to the resting lumen diameters (r=-0.33, P<0.001/r=-0.51, P<0.001). This correlation was even more distinct in females (females: FMD% r=-0.54, P<0.001; GTN% r=-0.64, P<0.001 vs. males: FMD% -0.23, P<0.001; GTN% -0.59, P<0. 001). No significant influence of age (61+/-9 years, FMD%: r=-0.04, P=0.68, GTN%: r=-0.18, P=0.05) and BMI (27.03+/-3.43 kg/m(2), FMD%: r=0.16, P=0.08, GTN%: r=0.09, P=0.3) on the test results were found. CONCLUSIONS: FMD% was found to be rather independent of age or BMI. The anatomical vessel size had an influence on the test results, which was more obvious in female patients. Our data indicate the necessity of further methodological studies, in larger, community-based populations. In particular, it needs to be clarified, whether vessel size or even gender-specific correction factors are required when using this technique in routine clinical practice. PMID- 11104878 TI - Effectiveness of low dose captopril versus propranolol therapy in infants with severe congestive failure due to left-to-right shunts. AB - To evaluate the therapeutical effects of the angiotensin converting enzyme inhibitor Captopril to the beta-blocker Propranolol in infants with congestive failure due to pulmonary overcirculation, we retrospectively analysed clinical, neurohormonal and hemodynamic data in 22 infants, 11 of whom were treated with Captopril (Group 1), 11 with Propranolol (Group 2). Age, weight, number of palliative operations, plasma renin activities and pulmonary to systemic flow ratios (3.5 vs. 3.5) were not significantly different prior to Captopril or Propranolol therapy. If treatment with digoxin and diuretics did not succeed, the infants were additionally treated with Captopril (1 mg/kg) for a mean of 7.4 months, or with 1.9 mg/kg Propranolol for 9.2 months. RESULTS: 1 mg/kg Captopril did not effectively suppress angiotensin converting enzyme in the steady state at trough level (92+/-52 vs. 87+/-50 nmol/min/ml). In the Propranolol group, the clinical heart failure score (2.6+/-1.5 vs. 7. 4+/-2.5) and plasma renin activities (14+/-10 vs. 101+/-70 ng/ml/h) were significantly lower, compared to the Captopril group. Length of hospital stay (23+/-9 vs. 52+/-24 days) was lower and weight gain (126+/-38 vs. 86+/-84 g/week) was higher within 3 months after starting Propranolol therapy. Significantly lower left atrial pressures (6.2+/ 2.2 vs. 13.4+/-9.2 mmHg) and lower endiastolic ventricular pressures (7.6+/-2.5 vs. 12.6+/-4.0 mmHg) during pre-operative cardiac catheterization indicated a better diastolic ventricular function under chronic Propranolol treatment. CONCLUSION: Although high dose Captopril was not evaluated in this study, when compared to patients on low Captopril dosages, infants who received Propranolol treatment showed improvement in heart failure scores, shorter lengths of hospital stay, lower plasma renin activities and better diastolic ventricular functions. PMID- 11104880 TI - Introduction. A symposium: brachytherapy for localized prostate cancer. PMID- 11104879 TI - Femoral haemostasis after transcatheter therapeutic intervention: a prospective randomised study of the angio-seal device vs. the femostop device. AB - BACKGROUND: A number of haemostatic devices are available to facilitate early haemostasis following transfemoral interventional procedures. METHODS AND RESULTS: We have prospectively compared 150 patients (age: 57+/-12 years, mean+/ S.D.) who were randomly assigned to either external compression using the FemoStop device or direct closure of the arterial puncture using the Angio-Seal device. The Angio-Seal was deployed in the catheter laboratory after the conclusion of the procedure. Patients, randomised to FemoStop, had their sheath removed when the activated clotting time (ACT) was less than 100 s before applying the device. The primary endpoint was the composite of bleeding, haematoma formation, bruise, requirement for blood transfusion, clinical indication for ultrasound examination at 2 h and 24 h following the procedure and crossover to either method at 2 and 24 h after the device deployment. The 95% of the Angio-Seal and 96% of FemoStop patients were discharged on the day following the procedure. An increased number of patients in the Angio-Seal group reached a clinical end-point within the first 2 h (45% vs. 3%, P<0.0001). This difference became insignificant at 24 h (25% vs. 30%, P=0.6). CONCLUSION: Although less comfortable, the overall efficacy of the FemoStop appeared to be higher than that of the Angio-Seal device. PMID- 11104881 TI - The role of active treatment in early prostate cancer. AB - The dramatic increase in the number of patients diagnosed with localized prostate cancer in the last decade presents a difficult challenge for physicians. Because the window of opportunity for cure is short it is vital to begin treatment before the cancer cells invade neighbouring tissues and organs or metastasise to other sites. This pressure of increased patient numbers provided clinicians with the opportunity to investigate other treatment options. New surgical techniques including laparoscopic radical prostatectomy, improving therapeutic radiation by the introduction of conformal radiotherapy, neutron radiation, cryosurgery, high intensity focussed ultrasound (HIF) and the revival of brachytherapy with or without external beam radiation are currently being investigated. The goal of these techniques is to treat localized prostate cancer based on the endpoints of disease specific mortality, no evidence of disease, absent or low levels of prostate-specific antigen (PSA), reduced side-effects, improved quality of life and importantly increased cost-efficacy. It is important to remember however, that watchful waiting and endocrine therapy are still valid therapy options in certain patient groups. The lack of randomized, prospective trials on local treatment of prostate cancer, makes it difficult to compare the efficacy of the different treatments, especially in terms of disease-specific survival. Trials are now in progress but it will be several years before results are available. In the meantime, we need to focus on surrogate endpoints, side effects, quality of life and the cost-efficacy of each treatment. It is also important to ensure that patients are kept informed and up-to-date with any new therapeutic developments. PMID- 11104882 TI - The place of radical prostatectomy in the treatment of early localized prostate cancer. AB - Today a number of treatment options exist for men diagnosed with early localized prostate carcinoma, of which the most important are radical prostatectomy, external beam radiotherapy and brachytherapy. New advances in brachytherapy using the implantation of iodine-125 and palladium-103 seeds have significantly altered its place in the treatment of localized disease and provided an alternative to external beam radiotherapy and potentially radical prostatectomy. Drawing on recently published data and our own experiences of retropubic radical prostatectomy in 100 consecutive men with localized disease, we review the place of radical prostatectomy in the treatment of early prostate cancer today. For many urologists radical prostatectomy remains the treatment of choice for men aged 70 years or less, with localized disease, a life expectancy of over 10 years and no co-morbidity. However, this has to be balanced against recent advances in brachytherapy, which now provides a minimally invasive alternative therapy for some patients with organ-confined disease and for those in whom surgery is contraindicated. PMID- 11104883 TI - Failure free survival following brachytherapy alone for prostate cancer: comparison with external beam radiotherapy. AB - BACKGROUND AND PURPOSE: To compare failure free survival (FFS) for brachytherapy (BT) alone and external beam radiotherapy (EBXRT) alone. MATERIALS AND METHODS: Between 12/88 and 12/95, 1527 and 695 T(1) or T(2) Nx-No Mo prostate cancer patients (from the Arizona Oncology Services database) were treated with either EBXRT or BT, respectively. The median age was 74 years. Median follow-up for EBXRT and BT patients was 41.3 and 51.3 months, respectively. RESULTS: Overall FFS at 5 years for EBXRT and BT were 69 and 71%, respectively (P=0.91). No significant difference in FFS at 5 years was observed between EBXRT and BT for either T(1) (78 vs. 83%, P=0.47) or T(2) (67 vs. 67%, P=0.89) tumours. Superior outcomes for Gleason 8-10 lesions treated with EBXRT vs. BT (5 years FFS 52 vs. 28%, P=0.04) were observed; outcomes for lower grade lesions when analysed by Gleason score alone did not significantly differ according to treatment received. Patients with initial PSA values 10-20 ng/dl had an improved FFS with EBXRT vs. BT (70 vs. 53%, P=0.001); outcomes for patients with initial PSA ranges 0-4 ng/dl, >4-10 ng/dl and >20 ng/dl did not differ significantly with treatment received. CONCLUSIONS: EBXRT and BT appear to be equally efficacious for low-risk patients having T(1)/T(2) disease with Gleason scores <6 and PSA <10 ng/dl. Patients with Gleason scores 8-10 or PSA >10 ng/dl-<20 ng/dl) appear to fare worse with BT alone compared with EBXRT. Neither EBXRT nor BT were particularly effective for patients with a presenting PSA >20 ng/dl. PMID- 11104884 TI - I-125 implantation for localized prostate cancer: the Utrecht University experience. AB - BACKGROUND AND PURPOSE: I-125 seed implantation is one of the treatment modalities for localized prostate carcinoma. It has few side-effects compared with radical prostatectomy and beam irradiation. MATERIALS AND METHODS: At the University Medical Centre, Utrecht, 249 naive patients were treated by perineal implantation between December 1989 and December 1998. Mean age was 69 years (range 45-91 years). Stage and grade were: T(1), 121; T(2), 126; T(3), 2; well differentiated, 136; moderate, 100; undifferentiated, 15; not established, 8. Mean initial PSA level was 16.1 ng/ml (range <1.0-165). Mean prostate volume was 33 cm(3). Sixty-two patients had had previous surgical intervention for voiding problems. Treatment evolved from single seeds to RAPID Strand, and from a probe mounted template to stepping unit and pre-planning. The introduction of RAPID Strand considerably increased the number of seeds (mean 41->65 seeds). Mean follow-up was 32.8 months, median 29.2 months (range 6-94 months). RESULTS: A total of 195 patients had no evidence of disease (18 died of intercurrent causes) and 54 had evidence of disease (13 died with prostate cancer). Toxicity was found in 22 patients. Urinary side-effects occurred in 18 patients, in nine cases after previous TURP. Four patients had intestinal problems, but only one had a rectal ulcer, which healed after hormonal therapy for local recurrence. CONCLUSIONS: Our findings indicate a correlation between the number of seeds implanted per cm(3) prostate volume and the final result. This is also reflected in a better volume coverage from MRI studies. PMID- 11104885 TI - The role of external beam radiotherapy with I-125/Pd-103 brachytherapy for prostate carcinoma. AB - BACKGROUND AND PURPOSE: To compare the biochemical outcomes of patients treated with Pd-103/I-125 brachytherapy alone vs. brachytherapy combined with external beam radiotherapy for early stage prostate carcinoma. METHODS: Brachytherapy monotherapy was used in 403 patients. Brachytherapy was combined with 45 Gy of external beam radiotherapy in 231 patients. Median follow-up was 58 months. To compare the biochemical outcomes of these two treatment approaches, patients were stratified into three relative risk groups: low risk, T(1)-T(2), Gleason 2-6/10, PSA< or =10.0; intermediate risk, T(3), Gleason 7-10/10, PSA>10.0 (one factor); high risk, T(3), Gleason 7-10/10, PSA>10.0 (two factors). RESULTS: The actuarial biochemical progression-free rate (bNED) for the entire 634 patients was 85% at 10 years. The bNED outcomes by risk group for monotherapy vs. combined therapy respectively were: low risk, 94 vs. 87%; intermediate risk, 84 vs. 85%; high risk, 54 vs. 62%. These differences did not reach statistical significance for any risk group. Rectal morbidity was slightly greater in the combined treatment patients. CONCLUSION: Although the addition of external beam irradiation to brachytherapy is conceptually appealing for patients with higher risk prostate carcinoma, we were unable to demonstrate a benefit. Whether this is because of patient selection biases within the risk groupings, an artefact of retrospective review, or because external radiotherapy does not offer additional benefit is uncertain. PMID- 11104886 TI - The combined use of the natural and the cumulative dose-volume histograms in planning and evaluation of permanent prostatic seed implants. AB - BACKGROUND AND PURPOSE: To investigate prostate dose coverage and overdosage in planned and realized permanent iodine seed prostate implants and to explore the use of the natural dose-volume histogram (NDVH) and the cumulative dose-volume histogram (CDVH) as tools to optimize prostate implants. MATERIALS AND METHODS: The optimal prescription dose (PD) or natural prescription dose (NPD) was derived from the NDVH. The mismatch between the NPD and the given PD was called the natural dose ratio (NDR). For an ideal implant the NDR should be 1. The target is overdosed if NDR >1 and underdosed if NDR <1. The NDR and prostate coverage were evaluated in implants of nine patients. Prostate coverage was determined from the CDVH based on pre-implant ultrasound or post-implant MRI for the planned and realized implants, respectively. The use of the NDVH to further optimize the planned prostate implants was also explored. RESULTS: The mean values of the NDRs were 1.30+/-0.34 (range 0.76-1.79), 1.22+/-0.31 (0.76-1.74) and 1.22+/-0.12 (0.98 1.33) for the planned, realized and optimized seed distributions, respectively. The realized prostatic implants showed smaller prostate coverage than the planned implants. The prostate volume fractions receiving 100% of the prescription dose were V(100)=79+/-6% and V(100)=97+/-3% for the realized and the planned implants, respectively. CONCLUSIONS: The NDVH and the CDVH proved to be valuable tools in plan evaluation. The NDVH and its derived parameter NDR quantify the risk of under or overdosage for a given PD. The CDVH is valuable in evaluation of prostate coverage realized prostate. Our strategy to implant just the prostate and not the prostate plus a margin led to NDR values between 1.1 and 1.3 and a prostate coverage of V(100)=79+/-6% in the nine patients. The planned coverage of V(100)=95% was not realized, mainly due to inadequate coverage of the base of the prostate. PMID- 11104887 TI - High dose rate brachytherapy boost treatment in radical radiotherapy for prostate cancer. AB - The natural history of prostate cancer is for early invasion of the prostatic capsule and seminal vesicles. This will be present in the majority of patients presenting with a prostate specific antigen (PSA) >10 or Gleason score >7. In these patients a combination of external beam treatment to provide a regional dose of radiation followed by a high dose rate afterloading brachytherapy boost to enable conformal dose escalation within the prostate gland presents an attractive option in local treatment. Accurate placement of catheters is now possible using transrectal ultrasound to provide high quality implants. A number of centres have now developed this technique as a routine clinical tool. There remains variation in the optimal dose fractionation with a range of BED(10) values from 100 to 77 and BED(3) values from 246.6 to 122.5. This does not, however, take into account geometric variations in dose distribution exploiting the physical advantage of BT in achieving a rapid dose fall off close to critical structures such as the rectum. Early results show PSA response levels of around 90% with grade III toxicity in 5-9% of patients. Critical evaluation of this technique in prospective, randomized trials is required. PMID- 11104888 TI - High dose rate prostate brachytherapy: the California Endocurietherapy (CET) method. AB - BACKGROUND AND PURPOSE: To describe the rationale, protocol and procedure for the treatment of prostate cancer using high dose rate brachytherapy (HDR-BT) and a non-fixed template technique. MATERIALS AND METHODS: Between July 1991 and December 1998, 491 patients with carcinoma of the prostate were treated using HDR BT and a non-fixed template technique. AJC stages T(1C)-T(3B), patients with prior transurethral resections of the prostate (TURP) and gland volumes >60 cm(3), were included. Flexible cystoscopy, fluoroscopy and transrectal ultrasound (TRUS) were used and 17 flexiguides were inserted through the perineum. Dosimetry was carried out using localization films. Treatment volume was defined at 4-6 mm outside the peripheral catheters. BT consisted of two implants, separated by 1 week, with two fractions given per implant for a total of four HDR fractions. Dose prescription to the treatment volume was 6 Gy (HDR) per fraction, with an additional dose of 0.5 to 0.75 Gy given where required. RESULTS: Patients with glands >60 cm(3), narrow pubic arches and TURP defects were treated satisfactorily. Symptoms of urinary irritation occurred with variable intensity and abated rapidly 2 weeks after the procedures. There was no high-grade chronic rectal morbidity and most patients reported no rectal symptoms or treatment related chronic urinary incontinence. CONCLUSIONS: The non-fixed template technique allowed flexibility in flexiguide placement to encompass large glands (>60 cm(3)), extracapsular extension and seminal vesicle involvement without the need for additional flexiguides. Also, small pubic arches and TURP defects posed little problem in positioning the flexiguides. This versatility resulted in complete treatment volume coverage of the prostate. PMID- 11104889 TI - Feasibility of permanent implants for prostate cancer after previous radiotherapy in the true pelvis. AB - BACKGROUND AND PURPOSE: Permanent seed implantation was used in the management of primary and recurrent prostate cancer in patients who had been treated previously by radiotherapy of the true pelvis. MATERIAL AND METHODS: Between 1993 and 1998 a total of 21 patients received an I-125 implant after radiotherapy for bladder cancer (two patients), anal cancer (one patient), seminoma (two patients) and prostate cancer (16 patients). Two seminoma and 10 prostate cancer patients were treated after earlier definitive external beam radiation therapy (EBRT), while the bladder and anal cancer were initially treated with EBRT plus iridium implantation. Six prostate cancer patients were initially treated by brachytherapy alone. The interval between the two treatments was longer in patients with radiotherapy for other malignancies than prostate cancer. RESULTS: After EBRT no serious late toxicity was observed. However, 1/6 patients who had two seed implants experienced serious complications, resulting in a vesico-rectal fistula. CONCLUSIONS: The permanent seed implantation with I-125 is feasible after previous radiotherapy in the prostate area. Also a second implant is possible, but may result in severe complications, depending on the initial dose and interval between the two treatments. PMID- 11104890 TI - Erectile dysfunction following radical therapy for prostate cancer. AB - With the earlier detection of prostate cancer and the increasing demand for treatment of organ-confined dizease, quality of life issues are becoming more important. Development of erectile dysfunction (ED) following radical therapy is a particular concern, and occurs in perhaps a third of patients treated by radiotherapy and 30-70% of patients treated by radical prostatectomy. Although it is assumed that the ED relates to damage to the nerves subserving erection, this view has been questioned recently and in at least a proportion of patients the cause appears to be vascular. Despite the likely cause of their ED, all patients presenting with ED after treatment for prostate cancer should undergo assessment by history and examination to ensure that there are no other correctable risk factors. Patients can then be considered for a number of treatment options, and currently sildenafil (Viagra, Pfizer) is usually used as first-line therapy assuming there are no contraindications, such as severe ischaemic heart disease or nitrate therapy. Sildenafil improves erectile function in 70% of patients with ED post-radiotherapy, but appears less effective in men after radical prostate surgery with a response rate of 40-50%. Other treatment options include self injection or intra-urethral administration of alprostadil, and some patients are happy to use a vacuum erection device. Finally, if all else fails, patients may be suitable for penile implant surgery. PMID- 11104891 TI - Brachytherapy with transperineal (125)Iodine seeds for localized prostate cancer. AB - BACKGROUND AND PURPOSE: To analyze the treatment results of transperineal (125)Iodine seeds in localized prostate cancer. PATIENTS AND METHODS: Between 1985 and 1996, 102 patients with T1-T2 N0 prostate cancer were treated with transperineal (125)Iodine seed implants at the Academic Medical Centre in Amsterdam. Tumours were classified as T1c in four patients, T2a in 73 patients and T2b in 25 patients. The mean pre-treatment PSA was 17 ng/ml. The (125)Iodine seeds were implanted transperineally under transrectal ultrasound guidance. The mean prostate volume was 31 ml (range 15-48 ml). An average of 49 seeds (range 29 74) was implanted. The dose to the periphery of the prostate was 160 Gy. Until 1988, 27 patients had additional external pelvic irradiation to a dose of 40 Gy in 20 daily fractions of 2 Gy. RESULTS: The 5- and 7-year actuarial survival rates were 77 and 63%, respectively (median 102 months). Ten patients (9.5%) died from prostate cancer. The 5- and 7-year clinical progression rates were 12 and 17%, respectively. Biochemical failure rates at 5 and 7 years were 39 and 44%, respectively. Age, alkaline phosphatase, creatinine, differentiation grade, additional treatment, staging procedure, number of seeds, prostate volume, treatment period and PSA were analyzed as prognostic factors. Only pre-treatment PSA was a prognosticator of clinical and biochemical outcome but not of survival. Biochemical control at 6 years varied from 30% for pre-treatment PSA values higher than 20 ng/ml to 95% for values < or =8 ng/ml. Forty-one out of 49 patients who were sexually active before brachytherapy maintained sexual function during the follow-up. Complete urinary incontinence occurred in one patient. No rectal complications were seen in patients receiving brachytherapy alone. CONCLUSIONS: Transperineal (125)Iodine seeds brachytherapy in localized prostate cancer achieves a good clinical control and overall survival with acceptable late toxicity. Biochemical failure was strongly correlated to the pre-treatment PSA value. PMID- 11104892 TI - ESTRO/EAU/EORTC recommendations on permanent seed implantation for localized prostate cancer. PMID- 11104893 TI - Places of emulsions in drug delivery. PMID- 11104894 TI - Preparation and evaluation of w/o/w type emulsions containing vancomycin. AB - The objective of this contribution is to summarize the preparation and application of water-in-oil-in-water type multiple emulsions (w/o/w emulsions) entrapping vancomycin (VCM). Formulations of the emulsions (the composition of an oily phase or the type and concentrations of surfactants) and emulsification methods (a stirring method and a membrane method) or conditions (rotation rates, pore sizes of membrane or operation pressures) were evaluated in order to prepare stable w/o/w emulsions. The pharmaceutical properties of the w/o/w emulsions - particle sizes, viscosity, phase separation and drug entrapment efficiency were measured and evaluated. We prepared stable w/o/w emulsions with a particle size of about 3 micrometer and an entrapment efficiency of VCM of about 70%. When this emulsion was administered intravenously to rats, plasma concentrations of VCM were prolonged compared to the VCM solution alone. The results of this study show the potential of the w/o/w emulsions for several clinical applications as one of the drug delivery systems. PMID- 11104895 TI - Basic study for stabilization of w/o/w emulsion and its application to transcatheter arterial embolization therapy. AB - Stabilization of w/o/w emulsion and its application to transcatheter arterial embolization (TAE) therapy are reviewed. W/o/w emulsion was stabilized by making inner aqueous phase hypertonic, addition of chitosan in inner phase, and techniques of phase-inversion with porous membrane. Lipiodol w/o/w emulsion for TAE therapy was prepared by using a two-step pumping emulsification procedure. The procedure is so easy that the emulsion could be prepared even during the surgical operation. The deposition after hepatic arterial administration of the emulsion was detected by an X-ray CT scanner. The concentration of epirubicin hydrochloride (EPI) in liver was increased and its residence was prolonged by encapsulating it in the w/o/w emulsion. The toxic effects of EPI and lipiodol on the normal hepatic cells were reduced. The w/o/w emulsion prepared by us is a suitable formulation for the TAE therapy. PMID- 11104896 TI - Particle control of emulsion by membrane emulsification and its applications. AB - Particle-size control of emulsion is very important for maintaining stability and giving emulsions new functional roles. Porous glass membrane, prepared by phase separation of a glass composition, is available as an emulsifying element, from which, one can obtain monodispersed emulsion with different particle sizes, and useful water/oil/water (W/O/W) emulsion in very high yield. The authors have called this new technology 'membrane emulsification'. Applications of membrane emulsification technology to drug delivery systems were carried out under cooperative research with Miyazaki Medical College. It was found that the clinical administration of a W/O/W drug emulsion that encapsulated an anticancer drug in its inner droplets was surprisingly effective for both terminal and multiple nodules of hepatocellular carcinoma when the drug was injected to damaged liver through a catheter inserted in the hepatic artery. Other applications have been tried and developed elsewhere. PMID- 11104897 TI - Hepatic arterial injection chemotherapy for hepatocellular carcinoma with epirubicin aqueous solution as numerous vesicles in iodinated poppy-seed oil microdroplets: clinical application of water-in-oil-in-water emulsion prepared using a membrane emulsification technique. AB - Iodinated poppy-seed oil (IPSO) accumulates selectively in hepatocellular carcinoma (HCC) when injected into the hepatic artery. This virtue has been applied to the hepatic arterial injection chemotherapy for the disease. We invented a new water-in-oil-in-water emulsion (W/O/W), in which IPSO microdroplets, 70 micrometer in diameter, were suspended in physiological saline enclosing numerous vesicles of an aqueous solution of epirubicin with remarkable stability. After hepatic arterial injection, the microdroplets accumulated only in HCC tissue and remained in the tissue for more than 3 weeks affecting tumor cells. Efficacy of the W/O/W has been fully proved clinically; the 6-year cumulative survival rate for 24 patients bearing HCC nodules recurrent after hepatectomy, including even 12 patients with four or more nodules, though prognosis of these patients is recognized very poor, was 24%. PMID- 11104898 TI - Preparation and evaluation of o/w type emulsions containing antitumor prostaglandin. AB - Antitumor prostaglandins(PGs) such as Delta12-PGJ2 and Delta7-PGA1 possess a cyclopentenone or cross-conjugated dienone structures. Antitumor PGs are actively incorporated through cell membrane and control gene expression. Very recent studies clarified that P53 independent expression of p21 and gadd 45, activation of PPARgamma are involved in antitumor mechanism of these PGs. At the low concentration, these PGs exhibit physiological or pathological activity such as osteoblast calcification, promotion of colon cancer cell proliferation. COMPARE PROGRAM using human 38 tumor cell lines suggested that antitumor mechanism of Delta7-PGA1 and 13, 14-dihydro-15-deoxy-Delta7-PGA1 methyl ester (TEI-9826) are quite different from other anticancer agents which are clinically used. Lipid microspheres and Lipiodol formulation were examined as dosage form of the PGs and lipid microspheres were selected for further study. At first lipid microspheres integrated TEI-9038 (Lipo TEI-9038) was chosen as a candidate for clinical trial. However Lipo TEI-9038 failed to exhibit substantial antitumor effect because of its enzymatic instability and toxicity in vivo. Lipo TEI-9826 was then selected as promising candidate for clinical trial because of its stability in serum. Lipo TEI-9826 exhibited marked antitumor effect in several animal models including CDDP resistant nude mice model. Pharmacokinetic and toxicological studies using rats suggested that continuous infusion is the most suitable administration method for Lipo TEI-9826. New type emulsifier, Controlled High Pressure Process Homogenizer (De-BEE 2000 and mini De-BEE) was developed during the preclinical studies on manufacturing process of Lipo TEI-9826. These results warrant the clinical trial for Lipo TEI-9826 in CDDP resistant cancer. PMID- 11104899 TI - Disposition characteristics of emulsions and incorporated drugs after systemic or local injection. AB - Lipid emulsions are useful tools for controlling the in vivo disposition of drugs and plasmid DNA. The dispositions of lipid emulsions are determined by their tissue interaction depending on the anatomical and physiological characteristics of each tissue and the physicochemical and biological properties of lipid emulsions. In addition, the retention of drugs is another issue, as too rapid a release of the drug would lead to failure of exerting its therapeutic potency. This review presents an overview about the disposition profiles and various physicochemical properties of lipid emulsions and incorporated drugs after systemic or local injection. Controlled biodistribution of lipid emulsions and incorporated drugs are also discussed. PMID- 11104900 TI - Microemulsion-based media as novel drug delivery systems. AB - Microemulsions are clear, stable, isotropic mixtures of oil, water and surfactant, frequently in combination with a cosurfactant. These systems are currently of interest to the pharmaceutical scientist because of their considerable potential to act as drug delivery vehicles by incorporating a wide range of drug molecules. In order to appreciate the potential of microemulsions as delivery vehicles, this review gives an overview of the formation and phase behaviour and characterization of microemulsions. The use of microemulsions and closely related microemulsion-based systems as drug delivery vehicles is reviewed, with particular emphasis being placed on recent developments and future directions. PMID- 11104901 TI - The instability within: problems in current analyses of microsatellite instability. AB - Microsatellite instability is regarded as one of the phenotypes of defective DNA mismatch repair and, consequently, as a marker of high risk for cancer. Despite numerous studies, the reported rates for positive microsatellite instability differ widely in each human malignancy. These discrepancies may relate to problems in the methods used. To establish a methodology for an accurate microsatellite instability analysis, technical requirements for a precise assay and biological conditions required for positive microsatellite instability were discussed. First, to describe microsatellite changes in detail, a sensitive detection system with linear detection characteristics and electrophoresis with standardised migration and minimised migration errors are considered to be necessary. Therefore, systems using fluorescent labelling and laser scanning are recommended. For reproducible polymerase chain reactions, it is essential to control the terminal deoxynucleotidyl transferase activity in Taq polymerase. Second, as a biological condition for positive microsatellite instability, feasible selection and combination of microsatellite markers, mutations in specific DNA mismatch repair genes and existence of monoclonal populations enriched sufficiently in a sample are essential. Finally, one possible diagnostic criterion for positive microsatellite instability is proposed, that is the existence of one of the patterns shown in the panel (see Fig. 6) at one or more loci in a set of more than five microsatellite markers. PMID- 11104902 TI - Fanconi anemia lymphocytes: effect of DL-alpha-tocopherol (Vitamin E) on chromatid breaks and on G2 repair efficiency. AB - The high frequency of chromosomal breaks in Fanconi anemia (FA) lymphocytes has been related to the increased oxidative damage shown by these cells. The effect of 100 microM DL-alpha-tocopherol (Vitamin E) on the level of chromosomal damage in mitosis was studied in lymphocytes from five FA patients and from age matched controls, both under basal conditions and when G2 repair was prevented by 2.5 mM caffeine (G2 unrepaired damage). In addition, the effect of this antioxidant on G2 duration and the efficiency of G2 repair was also evaluated in the sample. alpha-Tocopherol (AT) decreased the frequency of chromosomal damage (under basal and inhibited G2 repair conditions) and the duration of G2 in FA cells. This antioxidant protective effect, expressed as the decrease in chromatid breaks, was greater in FA cells (50.8%) than in controls (25%). The efficiency of the G2 repair process (G2 R rate) defined as the ratio between the percentage of chromatid breaks repaired in G2 and the duration of this cell cycle phase was lesser in FA cells (10.6) than in controls (22.6). AT treatment slightly increased this G2 R rate, both in FA cells and controls. These results suggest that an increased oxidative damage and a lower G2 repair rate may be simultaneously involved in the high frequency of chromatid damage detected in FA cells. PMID- 11104903 TI - The XRCC1 399 glutamine allele is a risk factor for adenocarcinoma of the lung. AB - Defects in the repair and maintenance of DNA increase risk for cancer. X-ray cross-complementing group 1 protein (XRCC1) is involved with the repair of DNA single-strand breaks. A nucleotide substitution of guanine to adenine leading to a non-conservative amino acid change was identified in the XRCC1 gene at codon 399 (Arg/Gln). This change is associated with higher levels of aflatoxin B1 adducts and glycophorin A somatic mutations. A case-control study was conducted to test the hypothesis that the 399Gln allele is positively associated with risk for adenocarcinoma of the lung. XRCC1 genotypes were assessed at codon 399 in 172 cases of lung adenocarcinoma and 143 cancer-free controls. Two ethnic populations were represented, non-Hispanic White and Hispanic. The distribution of XRCC1 genotypes differed between cases and controls. Among cases, 47.7% were Arg/Arg, 35.5% were Arg/Gln, and 16.9% were Gln/Gln. Among controls, XRCC1 allele frequencies were 45.5% for Arg/Arg, 44.8% for Arg/Gln, and 9.8% for Gln/Gln. Logistic regression analysis was used to assess the association between lung adenocarcinoma and the G/G genotype relative to the A/A or A/G genotypes. In non Hispanic White participants, the lung cancer risk associated with the G/G genotype increased significantly after adjustment for age (OR=2.81; 95% CI, 1.2 7.9; P=0.03) and increased further after adjustment for smoking (OR=3.25; 95% CI, 1.2-10.7; P=0.03). Among all groups, a significant association was found between the G/G homozygote and lung cancer (OR=2.45; 95% CI, 1.1-5.8; P=0.03) after adjustment for age, ethnicity, and smoking. This study links a functional polymorphism in the critical repair gene XRCC1 to risk for adenocarcinoma of the lung. PMID- 11104904 TI - Induced mutagenic effects in the nucleotide excision repair deficient Drosophila mutant mus201(D1), expressing a truncated XPG protein. AB - Defects in nucleotide excision repair (NER) as defined by the UV sensitivity of xeroderma pigmentosum (XP), Cockayne syndrome (CS) and trichothiodystrophy (TTD) patients has lead to the identification of most of the genes involved: XPA through XPG, CSA and CSB. Whereas XP patients often show an increased risk for skin cancer after exposure to sunlight, this is not the case for patients with CS and TTD. Several CS patients have been shown to carry a defect in the XPG gene. The XPG, a structure specific endonuclease makes the incision 3' of damage and is also involved in the subsequent 5'incision during the NER process. In addition, XPG plays a role in the removal of oxidative DNA damage. The Drosophila XPG gene was isolated and based on the molecular defect of a spontaneous (insertion) and an EMS induced mutant, it was shown that a mutated XPG is responsible for the Drosophila mutagen-sensitive mutants mus201. One of these mutants, mus201(D1) has been used extensively in studies of the effects and mechanisms of many chemical mutagens as well as X-rays. The results of these studies are discussed in the light of the finding that mus201p is the Drosophila homologue of XPG. PMID- 11104905 TI - tert-Butoxyl radicals generate mainly 7,8-dihydro-8-oxoguanine in DNA. AB - Like hydroxyl radicals, alkoxyl radicals have been implicated in the generation of cellular oxidative DNA damage under physiological conditions; however, their genotoxic potential has not yet been established. We have analyzed the DNA damage induced by a photochemical source of tert-butoxyl radicals, the water soluble peroxy ester [4-(tert-butyldioxycarbonyl)benzyl]triethylammonium chloride (BCBT), using various repair endonucleases as probes. The irradiation (UV(360)) of BCBT in the presence of bacteriophage PM2 DNA was found to generate a DNA damage profile that consisted mostly of base modifications sensitive to the repair endonuclease Fpg protein. Approximately 90% of the modifications were identified as 7,8-dihydro-8-oxoguanine (8-oxoGua) residues by HPLC/ECD analysis. Oxidative pyrimidine modifications (sensitive to endonuclease III), sites of base loss (AP sites) and single-strand breaks were only minor modifications. Experiments with various scavengers and quenchers indicated that the DNA damage by BCBT+UV(360) was caused by tert-butoxyl radicals as the ultimate reactive species. The mutagenicity associated with the induced damage was analyzed in the gpt gene of plasmid pSV2gpt, which was exposed to BCBT+UV(360) and subsequently transfected into Escherichia coli. The results were in agreement with the specific generation of 8-oxoGua. Nearly all point mutations (20 out of 21) were found to be GC-->TA transversions known to be characteristic for 8-oxoGua. In conclusion, alkoxyl radicals generated from BCBT+UV(360) induce 8-oxoGua in DNA with a higher selectivity than any other reactive oxygen species analyzed so far. PMID- 11104906 TI - Endonuclease V of Escherichia coli prevents mutations from nitrosative deamination during nitrate/nitrite respiration. AB - Endonuclease V (Endo V) of Escherichia coli participates in the excision repair of hypoxanthine and xanthine (deaminated adenine and guanine) in DNA. It thereby reduces the mutagenic effects of nitrous acid by attacking lesions caused by nitrosative deamination. Nitrosating agents may be produced endogenously when E. coli is grown in oxygen-poor cultures, during which nitrate and nitrite replace oxygen as preferred electron acceptors. In this study, the protective effect of Endo V was observed under such conditions. During micro-aerobic growth, an nfi (Endo V) mutation enhanced the frequency of nitrate- and nitrite-induced A:T- >G:C and G:C-->A:T transition mutations, which are consistent with a defect in the removal of DNA hypoxanthine and xanthine, respectively. Similar effects were observed in saturated, aerobic cultures but not in well-aerated, logarithmically growing ones. A narG (nitrate reductase) mutation blocked the mutagenesis of the nfi mutant by nitrate but not by nitrite. These results differed from those of previous studies in which cell suspensions generated an exogenous nitrosating agent from nitrite, but not from nitrate, in a reaction that was narG-dependent. Nitrate/nitrite metabolism is also known to generate endogenous alkylating agents through N-nitrosation. However, an nfi mutation did not appreciably enhance mutagenesis by N-methyl-N-nitrosourea, suggesting that the mutator effect of nfi is not due to a defect in alkylation repair. The overall results indicate that Endo V functions during normal growth by helping to repair nitrosatively deaminated bases in DNA, which are by-products of anaerobic nitrate/nitrite respiration. PMID- 11104907 TI - Characterization of RAD52 homologs in the fission yeast Schizosaccharomyces pombe. AB - The RAD52 gene of Saccharomyces cerevisiae is essential for repair of DNA double strand breaks (DSBs) by homologous recombination. Inactivation of this gene confers hypersensitivity to DSB-inducing agents and defects in most forms of recombination. The rad22+ gene in Schizosaccharomyces pombe (here referred to as rad22A+) has been characterized as a homolog of RAD52 in fission yeast. Here, we report the identification of a second RAD52 homolog in Schizosaccharomyces pombe, called rad22B+. The amino acid sequences of Rad22A and Rad22B show significant conservation (38% identity). Deletion mutants of respectively, rad22A and rad22B, show different phenotypes with respect to sensitivity to X-rays and the ability to perform homologous recombination as measured by the integration of plasmid DNA. Inactivation of rad22A+ leads to a severe sensitivity to X-rays and a strong decrease in recombination (13-fold), while the rad22B mutation does not result in a decrease in homologous recombination or a change in radiation sensitivity. In a rad22A-rad22B double mutant the radiation sensitivity is further enhanced in comparison with the rad22A single mutant. Overexpression of the rad22B+ gene results in partial suppression of the DNA repair defects of the rad22A mutant strain. Meiotic recombination and spore viability are only slightly affected in either single mutant, but outgrowth of viable spores is almost 31-fold reduced in the rad22A-rad22B double mutant. The results obtained imply a crucial role for rad22A+ in repair and recombination in vegetative cells just like RAD52 in S. cerevisiae. The rad22B+ gene presumably has an auxiliary role in the repair of DSBs. The drastic reduced spore viability in the double mutant suggests that meiosis in S. pombe is dependent on the presence of either rad22A+ or rad22B+. PMID- 11104908 TI - Sequence variation in the human uracil-DNA glycosylase (UNG) gene. AB - Spontaneous deamination of cytosine results in a premutagenic G:U mismatch that may result in a GC-->AT transition during replication. The human UNG-gene encodes the major uracil-DNA glycosylase (UDG or UNG) which releases uracil from DNA, thus, initiating base excision repair to restore the correct DNA sequence. Bacterial and yeast mutants lacking the homologous UDG exhibit elevated spontaneous mutation frequencies. Hence, mutations in the human UNG gene could presumably result in a mutator phenotype. We screened all seven exons including exon-intron boundaries, both promoters, and one intron of the UNG gene and identified considerable sequence variation in cell lines derived from normal fibroblasts and tumour tissue. None of the sequence variants was accompanied by significantly reduced UDG activity. In the UNG gene from 62 sources, we identified 12 different variant alleles, with allele frequencies ranging from 0.01 to 0.23. We identified one variant allele per 3.8kb in non-coding regions, but none in the coding region of the gene. In promoter B we identified four different variants. A substitution within an AP2 element was observed in tumour cell lines only and had an allele frequency of 0.10. Introduction of this substitution into chimaeric promoter-luciferase constructs affected transcription from the promoter. UDG-activity varied little in fibroblasts, but widely between tumour cell lines. This variation did not however correlate with the presence of any of the variant alleles. In conclusion, mutations affecting the function of human UNG gene are seemingly infrequent in human tumour cell lines. PMID- 11104909 TI - Should the patella be resurfaced at total knee replacement? AB - Total knee replacement (TKR) presumably is replacement of the total knee articular surface. Sometimes it is and sometimes it is not. It is this author's firm conviction that the patella should be resurfaced in the vast majority of cases. Such advocacy must be critically justified and the potential drawbacks and alternatives examined. PMID- 11104910 TI - Patella or not?. (Is the patella component the 'Cinderella' of total knee arthroplasty?). PMID- 11104911 TI - Topical preparations for chronic disorders of the knee: a review. PMID- 11104912 TI - A mid-term follow-up of medial patellofemoral ligament reconstruction using an artificial ligament for recurrent patellar dislocation. AB - A prospective study on medial patellofemoral ligament reconstruction for recurrent patellar dislocation was performed. At an average follow-up of 5.9 years, 27 MPFL reconstructions using a mesh-type artificial ligament and medial retinaculum slip coverage were reviewed using the Crosby and Insall grading system. Fifteen knees (55%) were classified as excellent, 11 knees (41%) as good, 1 knee (4%) as fair/poor and none as worse. At a mid-term follow-up, medial patellofemoral ligament reconstruction should be considered as a safe and effective operation. PMID- 11104913 TI - Assessment of articular cartilage of the lateral tibial plateau in varus osteoarthritis of the knee. AB - This study describes the soft X-ray examinations of 24 lateral tibial plateaus obtained during total knee arthroplasty for varus osteoarthritis. The average thickness of the articular cartilage was 3.5 min and ranged from 2.1 to 5.0. We considered that 21 out of the 24 lateral tibial plateaus had well preserved articular cartilage. Within the well preserved articular cartilage, bony protuberances of various sizes were found in five cases. All lateral tibial plateaus except one showed osteophyte formation. We considered that 12 of the 24 lateral tibial plateaus had large osteophytes. Ten of these 12 lateral tibial plateaus had well preserved articular cartilage. Large osteophyte formation may not necessarily be a contra-indication of high tibial osteotomy (HTO) or unicompartmental knee arthroplasty (UKA). Cases with a bony protuberance may not be suitable for HTO or UKA, because the overlying articular cartilage is thin and inadequate for supporting load. PMID- 11104914 TI - Three-dimensional knee analyzer validation by simple fluoroscopic study. AB - Introduction: The complexity of the knee articulation makes its clinical evaluation extremely difficult. Insufficiency of existing instruments for knee evaluation prevents physicians from providing a diagnosis of injury and/or an evaluation of different treatments. To this end, our research group has developed a functional knee analyzer, which allows a three-dimensional evaluation of the knee in motion. The goal of this study is to scientifically validate the functional knee analyzer before using it in clinical setting. Materials and methods: The three-dimensional knee analyzer includes an orthoplastic exoskeleton attachment system, a kinematic tracking device, a screen for graphical display and a C(++) program with a user interface for calculating kinematic indices. A fluoroscopic study was performed on five healthy subjects with a mean age of 28. The experiment was set-up to determine the reduction of skin movement with respect to the underlying bone by using a knee exoskeleton attachment system. The root mean square (RMS) errors of markers movement about the abduction-X (RMSRx) and tibial rotation-Z (RMSRz) axes and displacement in the XZ plane (RMSpxpz) were calculated, once by placing markers directly on the skin and once on the exoskeleton attachment system. Results: Our results demonstrated that RMSpxpz, RMSRx and RMSRz were reduced by a factor of 6 (min 1.8, max 26), 4.3 (min 0.75, max 21) and 6.2 (min 2, max 26.4) on average, respectively, for four subjects out of five when the exoskeleton attachment was used. PMID- 11104915 TI - The outcome of cemented vs. cementless fixation of a femoral component in total knee replacement (TKR) with the identification of radiological signs for the prediction of failure. AB - A survival analysis and radiological review were performed on a series of femoral total knee arthroplasty (TKA) prostheses either cemented (150 cases) or cementless, press-fit (201 cases). The internal surface of the femoral components were shot-blast CoCr alloy. The incidence of loosening of the femoral component at 6 years was 9.8% with cementless fixation and 0.6% with cement (P<0.05) at 6 years. Amongst uncemented prostheses, there was no difference in the survival or radiological outcome with the use of a stem as against two condylar pegs. The clinical need for revision may be predicted radiologically 3 years after operation in symptomless patients by noting a change in component position combined with progressive radiolucent lines and osteolysis. Thus, radiological follow-up should be continued for a minimum of 3 years after knee replacement. The use of a stem enabled the detection of radiolucent lines which we believe were missed around prostheses with condylar pegs. Thus, the use of a stem improves the prediction of failure (but does not improve fixation). PMID- 11104916 TI - Staple vs. subcuticular vicryl skin closure in knee replacement surgery: a spectrophotographic assessment of wound characteristics. AB - Staple closure is a popular method of skin closure for patients undergoing knee replacement surgery. There are no guidelines regarding spacing of staples and some concern exists with regard to wound oxygenation in knees subject to early movement. We compared cutaneous wound characteristics in terms of blood oxygenation and blood content, using two types of skin closure. Staples or 4/0 subcuticular vicryl were used. We found favourable blood perfusion characteristics when using stapled closure. Our results also suggest that optimum wound oxygenation requires staple spacing of 6 mm or more. PMID- 11104917 TI - Should I scan or should I scope? AB - This paper proposes a clinical points-based method that can be used by less experienced orthopaedic surgeons who are faced with the dilemma 'Should I scan or should I scope?' By considering the history, the age of the patient and the clinical findings one can expect diagnostic performance equivalent to magnetic resonance imaging. The cost of MRI services must be considered when selecting patients for this investigation. We have suggested a threshold for requesting an MRI scan depending on the relative costs. PMID- 11104918 TI - Recurrent non-Hodgkin's lymphoma presenting as pathological fracture of the patella. AB - We report a case of recurrent non-Hodgkin's lymphoma presenting as anterior knee pain due to pathological fracture of the patella. Anterior knee pains is an extremely common presenting complaint at orthopaedic clinics and the patella is an infrequent site both for primary and recurrent tumours. This case, treated by patellectomy, illustrates the difficulty of treatment of these lesions and highlights the importance of adequate radiological assessment in sites of previous tumour involvement. PMID- 11104919 TI - Angiomyoma of the patellar fat pad. AB - We report a case of solitary angiomyoma arising in the infra-patellar fat pad and presenting with anterior knee pain. PMID- 11104920 TI - An investigation of the suitability of the peri-articular osteophyte as an autologous graft for the repair of articular surface defects. PMID- 11104921 TI - [Precarious life: an anthropological concept]. PMID- 11104922 TI - [Pneumopathies caused by inhalation of hydrocarbons: apropos of 3 cases]. AB - We report three personal cases of hydrocarbide aspiration pneumonia. High viscosity non-volatile hydrocarbides (paraffin oil, for instance) cause often pseudotumoral exogenous fat-aspiration lung disease. Low-viscosity volatile hydrocarbides (petroleum, gasoline, white spirit, for instance) cause acute pseudo-infectious lung disease with dyspnea and fever which usually resolves within a few weeks but which may also be life-threatening. Purely symptomatic treatment has greatly progressed with advances in intensive ventilatory assistance. Gastric emptying with emetic agents or lavage procedures is dangerous and must be avoided except for exceptional cases. When required, the airways must be protected with tracheal intubation. Volatile hydrocarbides should be stored in protected areas in containers with safety stoppers which children cannot open. PMID- 11104923 TI - Brain tuberculomas. AB - Although brain tuberculomas have become rare in developed countries, diagnosis should be kept in mind when confronted with brain space-occupying lesion(s), particularly in immigrants from endemic countries. Surprisingly, Human Immunodeficiency Virus (HIV) infection does not seem to have affected the incidence of this tuberculous lesion. Clinical, biological and radiological signs are only suggestive. Thus, when no other active extracranial tuberculous process is found, the diagnosis should be confirmed by a biopsy before beginning antituberculous treatment which is rapidly effective. Adjunction of steroids may be warranted owing to the common paradoxical enlargement of lesions during the first weeks of therapy, as is now well described in lymph node tuberculosis. Cure can be obtained in more than 85% of the cases, but neurological sequelae are not rare. PMID- 11104925 TI - [Erysipelas and necrotizing fasciitis: management (short text). Consensus conference. French Society of Dermatology]. PMID- 11104924 TI - [Infective endocarditis related to pacemaker leads. A review]. AB - Infective endocarditis is a rare but serious complication of permanent cardiac pacing with high mortality ranging from 10 to 30%. Clinical symptoms are sometimes acute but more often poor and aspecific in subacute and chronic forms causing prolonged diagnostic delay. In order to make endocarditis on pacemaker leads clearer, we conducted a medline search of all published literature. Analysis of this literature shows that the initial infective source is often local and that Staphylococcus species are the most often pathogens isolated. Clinicians have to search carefully for local inflammatory signs, past or ongoing, and pulmonary embolism because their presence will be helpful for diagnosis. Transoesophageal echocardiography is essential; it shows vegetations in more than 90% and must be repeated when the examination is negative. Treatment has a double goal: a prophylactic treatment in order to reduce risk factors of infection related to implantation of the pacemaker and a curative treatment associating prolonged antibiotic therapy with extraction of the material. PMID- 11104926 TI - [Therapeutic strategies for osteoporosis]. AB - Therapeutic strategy in osteoporosis should rely on the results of good quality randomized controlled trials. When a fracture has already occurred, first line treatments are alendronate or hormone replacement therapy. Alternatives are cyclical etidronate, or calcitonin in some countries. General measures, such as adequate nutrition and calcium and vitamin D intake, and prevention of falls should be associated to drugs. When there is no history of fracture, the decision will rely on the existence of clinical risk factors and the bone density. Women with a densitometric osteoporosis will be treated as those who have already fractured. Those who have osteopenia can receive hormone replacement therapy, alendronate or raloxifene. No treatment is indicated in women who have normal bone density. PMID- 11104927 TI - [Calcium and vitamin D deficiencies and their therapeutic indications]. PMID- 11104928 TI - Past and future of anabolic agents. PMID- 11104929 TI - [Estrogens and selective estrogen receptor modulators in the treatment of osteoporosis]. AB - Estrogen deficiency is the major determinant of bone loss, not only in the first years postmenopause, but also throughout the entire life and in the elderly. Major progress in the knowledge of cellular actions of estrogens has been made leading to a better understanding of the underlying mechanisms of different estrogen-deficiency related diseases such as osteoporosis, atherosclerosis and also maybe cerebral aging. Estrogen replacement therapy remains the first choice treatment in the prevention of postmenopausal osteoporosis, but the continuous aging process of the female population raises the question of a better strategy of action in a more efficient prevention of hip fractures. Moreover, the potential gynecological effects of estrogens are likely to limit their indications or long-term use. The development of new compounds, called SERMs (selective estrogen receptor modulators), with both agonist and antagonist estrogen actions, in particular with no negative effects on the uterus and the breast opens new therapeutical insights into the prevention of postmenopausal osteoporosis. PMID- 11104930 TI - [Secondary osteoporosis and glucocorticoid-induced osteoporosis]. AB - In order to assess properly the diagnosis of osteoporosis, a short clinical investigation is required to address potential causes for bone loss. Osteoporosis used to be suspected in a patient with vertebral demineralization, but nowadays it is often diagnosed in a patient with a low bone mass on a screening dual energy X-ray absorptiometry (DEXA). In this setting, it is important for the clinician to look for secondary osteoporosis, especially in men in whom secondary osteoporosis is more frequent than in women, before discussing any specific therapy. The major causes are longterm glucocorticoid therapy, endocrine (hypogonadism, primary hyperparathyroidism, hyperthyroidism), or digestive diseases. PMID- 11104931 TI - Bisphosphonates in the treatment of osteoporosis. Principles and efficacy. PMID- 11104933 TI - [Lymphoma involving the temporal artery]. AB - Involvement of the temporal arteries, considered to be the hallmark of giant cell arteritis, is rather rare in other pathologic processes. We describe a patient with lymphoma involving temporal artery. A 68-year-old man has been followed closely without therapy since 1989 for a low-grade non-Hodgkin lymphoma. He presented in 1995 with asymptomatic nodules on the temporal artery with preservation of the pulse. Temporal artery biopsy showed periarterial infiltration of mononuclear cells in keeping with follicular mixed cell lymphoma. The differential diagnosis of temporal arteritis must therefore, include other vasculitides, light chain amyloidosis but also lymphoma and emphasize the need for a temporal artery biopsy. PMID- 11104932 TI - [Familial Waldmann's disease]. AB - We report the observation of a mother and her daughter who presented edema, hypoprotidemia and lymphopenia due to protein-losing enteropathy. Radiological, endoscopic and histological investigations revealed the diagnosis of primary intestinal lymphangiectasis or Waldmann's disease. Dietary treatment with middle chained triglycerides was effective. Familial cases are rarely described. PMID- 11104935 TI - [Association of sarcoidosis and multiple autoimmune syndrome]. PMID- 11104934 TI - [Sarcoidosis-induced pleural thickening: a case report]. PMID- 11104936 TI - [A clear look]. PMID- 11104937 TI - [Risks incurred by first-injection intravenous drug users] [In Process Citation] AB - Aims. - The objectives of the study were to describe the circumstances surrounding the intiation of intravenous drug use, the role of the introducer and to evaluate intravenous drug users risk behaviors at the first injection of drug. Design.- In 1997, we conducted a cross-sectional survey using a structured questionnaire concerning the initiation process into intravenous drug abuse. IDUs were interviewed in four treatment drug abuse and psychosocial centers in Paris and in one prison. Participants.- Of the 152 consecutive IDUs interviewed, 143 completed the questionnaire, 83 were male. Findings. - The mean age at first opiate use and at first injection were 19 years (SD: 4.3) and 20 years (SD: 4.3). At first injection, heroin was the main used drug (91%), the subject was with others persons (91%), asked himself for injection (70%) albeit had not planned this injection (40%). The subject injected at a friend's home (31%). The introducer was an IDU (93%), mean age 23.4 (SD: 5.2). He or she was a friend (61%) or a sexual partner (14%). The preparation of the first injection and the injection were made by the introducer in 72% and 74% of cases. The injecting equipment had been borrowed (22%) from an IDU whose HIV status and HCV status were unknown in 83% and 85% of cases. Conclusion. - Our study shows novel results about the first injection, they are of prime importance for harm reduction. The introducer plays a major role in preventing risk-behavior at the first injection and for education about safe injecting practices. PMID- 11104938 TI - Comparison of high dose buprenorphine treatments of opiate dependent outpatients in four healthcare networks. AB - AIMS: The aim of this study was to compare the various clinical practices in four health care networks and to access how the variations in treatment effected the outcome in opiate-dependent patients. METHODS: A retrospective study was carried out with 71 participating general practitioners. These were chosen from a group of 354 practitioners from four health care networks. Each practitioner could enroll up to 5 patients who were currently undergoing treatment with high-dose buprenorphine(HDB). The patients treatment had to have been initiated between the 1(st) of February 1996 and the 31(st) of October 1996, and excluded any patients who had lapsed on their treatment during the first month. Patients were selected until a total of 75 cases were enrolled from each network. Data were then collected retrospectively between June and December 1997. Information collected concerned the initial stage of treatment, the stabilizing stage or level of treatment and followed up data on the most recent prescriptions. RESULTS: The final patient maintenance totals were high for all four care networks (82.7 to 96% of patients were still being followed by their doctor at the final evaluation). A positive outcome as indicated by reduction of risk and decreased social vulnerability was also observed in all networks. Additionally, in each network there was a clear correlation between prescription practices and patient behavior. For example, the prescription of HDB at a daily dose of less than 6.2mg was associated with a higher rate of benzodiazepine use; and prescription of several daily doses of HDB was associated with a higher percentage of injecting patients. CONCLUSION: This retrospective study provides evidence that general practitioner care of drug-dependent patients as outpatients, within a health care network helps to stabilize patient visits, allows treatment of associated comorbidities and favors social rehabilitation. The prescription of HDB as a single daily dose, individually adapted for each patient, optimizes the outcome and reduces misuse. PMID- 11104939 TI - [Intoxications by hallucinogenic mushrooms]. AB - In the context of excessive use of natural or synthetic psychoactive substances, with stimulating, psychedelic and hallucinogenic effects, an increase, if not a recurrence, in consumption of Psilocybe semilanceata has been observed in France over some fifteen years. Psilocybin and psilocin are the active compounds, responsible for the hallucinogenic effects and are a part of the substance group, derived from tryptamine and characterized by an indole nucleus. We report a clinical case observed in an emergency unit and review the historical, epidemiological and pharmacological data now available for these intoxications. Of particular interests in hallucinogenic mushroom abuse are: increased consumption in the context of youth cultural and entertainment movements; possible difficulties in the diagnosis in emergency conditions; possibility of severe and fatal complications. PMID- 11104940 TI - [Pregnancy and drug abuse: current situation and therapeutic strategies]. AB - Drug abuse, a general public health problem, concerns a growing number of the pregnant women. Pregnancy in a drug addict is a high risk condition. The prevention of neonatal and pediatric complications involving both organic and psychological conditions, requires early individual medical, psychological and social support adapted to each pregnant addict. The risk for the mother and the fetus warrant prescription of substitution therapy during pregnancy. Published results concerning pregnancies in women on substitution therapy have been encouraging and clearly show a decrease in maternal and fetal complications. These studies are however difficult to conduct and carry a certain number of biases related to the social and economical context of the patients and also to the type of drug abuse (multiple drugs, drug combinations, psychostimulants, smoking, alcohol). PMID- 11104941 TI - [Late-onset alcohol withdrawal syndrome]. AB - The alcoholic withdrawal syndrome (AWS) arises variably within hours following the hospitalization of an alcohol dependent patient. Delay usually observed between admission and the first symptoms depends above all on alcohol serum level concentration at arrival and on the degree of severity of physical dependence. The case reported here describes the very late onset severe alcoholic withdrawal syndrome observed in a 57-year-old alcohol dependent patient hospitalized for leg trauma and operated within hours followed admission. The first symptoms of AWS appeared only the 4-th day after hospitalization and the patient quickly developed a clinical state of delirium tremens. Delay in the onset of this AWS is discussed. PMID- 11104942 TI - [Neonatal withdrawal syndrome in twins born to a mother on methadone substitution]. AB - We report the cases of two female twins whose mother was taking methadone substitution therapy. These cases demonstrate the unpredictable nature of the neonatal withdrawal syndrome. One baby developed signs of withdrawal late 10 days after birth. She had a less severe syndrome than her twin sister who was hypotrophic and developed signs on day 1 which persisted for 6 weeks. PMID- 11104943 TI - [Tramadol dependence in a previous heroin addict]. PMID- 11104944 TI - [The concept of addictology and addiction: the patient's point of view. A study on 90 persons followed for addiction]. AB - The concept of addiction has gained recognition in French semantics, standing for a pathological condition that was formerly designated solely on the basis of specific drug abuse. We questioned 90 patients followed at there Parisian outpatient units about their understanding of the term "addiction". We also looked for signs that this term is used to federate notions since it is supposed to describe a general condition involving a large number of singular phenomena. One part of our questionnaire focused on treatment modes (several practitioners for several addictions, one institution for all addictions, specialized consultations with a practitioner other than the usual primary care physician). Our findings would illustrated areas where further details are needed in order to consider patient expectations when designing specialized care for outpatients or inpatients. PMID- 11104945 TI - Coca leaf chewing as therapy for cocaine maintenance. AB - Major ethnic groups in Bolivia (Aymaras and Quechuas) have chewed the coca leaf for generations upon generations without health problems. The effects of coca leaf chewing produce a level of social and economic adaptation that is beyond what is normally possible. This was a major factor during the Spanish colonization of Bolivia, when forced native labor was used extensively. The cocaine base, or "pasta", may be seen as a type of South American crack. Its obligatory method of administration is smoking. A primary condition of the "pasta" smoker is compulsive drug-search behavior and addiction to cocaine base destroys emotional and mental balance. Socio-economic maladjustment is the norm amongst "pasta" addicts. Since 1984 I have recommended the chewing of the coca leaf, between 100 to 200 grams of coca leaf per week for the treatment of cocaine dependence. Since this treatment was dispensed on an ad hoc basis, it was not possible to measure the relapses. However, an assessment was conducted on the basis of mental condition and level of social and economic adaptation before and after treatment. The patent's level of social acceptance, before treatment, only reached 60% at most, and after treatment, 26% improved their level of adaptation. Four patients among 50 reached an adaptation level of 100%. Upon final assessment, the level of social adaptation prior to treatment was only 28%, after treatment as many as 48.8% of the patients were socially adapted. PMID- 11104946 TI - [New risks of addiction for new populations: the example of hackers]. AB - Our purpose was to examine recent social and technical habits related to high tech environments. Our goal was to show that the prevention of risk behaviors due to training in data processing, requires an interdisciplinary approach where medical anthropology could benefit from and exchange of complementary information sources (particularly from psychiatrics and psychoanalysis). We used this approach to search for solutions regarding new kinds of addiction. When identifying pathological conditions and proposing appropriate care, these solutions must take into consideration the progressive loss of human nature in data processing environments and the very important and highly sophisticated relationship established between the human being and the computer. We looked at the hacker population as a modern tribe and marginal group. Our analysis led to a better understanding of this kind of artificial culture, sometimes called a "high tech" or "cyber" culture. The hacker population is integrating new rituals, languages and special rhythms which induce addictions. We show how high-tech environments operating in e-time and e-life induce addictions. This work illustrates a classical anthropological approach to the question (ethnological fields, interviews, literature analysis). The major challenge is to explain how high-tech environments present high risks for dependency in the hacker population and other, unwarned, computer (ab)users. PMID- 11104947 TI - [Pertinence of the addiction concept in eating behavior disorders]. AB - From a psychodynamic perspective, dependence disorders, irrespective of the object of addiction, can be seen as the expression of the subject's neurobiological, psychopathological, cultural and social vulnerability. Since vulnerability strengthens and reorganizes the personality, it can drive these subjects to perpetuate pathological behaviors. In this light, behavior disorders belong to the field of addiction diseases, especially considering that the underlying psychopathological structures are close to those observed in addiction, that depression plays a central role, and that their development into toxic addictive behavior (drugs, alcohol, psychotrope) is frequent. PMID- 11104949 TI - [ [In Process Citation] PMID- 11104948 TI - [The Rorschach's test for heroin addicts treated by methadone: from reality to imaginary]. AB - The aim of this study was to analyze the role of the imaginary and its link with reality among 30 subjects treated by methadone. The Rorschach test was proposed to 30 heroin addicts treated by methadone, over an average 7 months. The formal answers (used preferentially in the 30 results, though often inappropriately) and socializing landmarks involving considerable individual investment, we were able to identify two ways of working within the same protocol. The first relies most on conformity and adaptation and tries to put the imaginary aside. The second is a more projective, unorganized approach evidencing the influence of the imaginary. This specific imaginary activity could not be assimilated with the traditional "lack of fantasy" observed in heroin addicts. It would be interesting to examine the course of this approach over a longer period. PMID- 11104951 TI - [Neonatal consequences of cesarean section]. PMID- 11104950 TI - [Maternal consequences of cesarean as related to vaginal delivery]. PMID- 11104952 TI - [Is cesarean section indicated for suspected macrosomia?]. PMID- 11104953 TI - [Is cesarean section indicated for breach presentation?]. PMID- 11104954 TI - [Is cesarean section indicated for twin birth?]. PMID- 11104955 TI - [Is cesarean section indicated for the cicatricial uterus?]. PMID- 11104956 TI - [Indications for cesarean section in case of dystocia]. PMID- 11104957 TI - [Can (or should) the patient's desire for cesarean section be accepted if there is no medical indication?]. PMID- 11104958 TI - [Operative technique for cesarean section]. PMID- 11104960 TI - [ [In Process Citation] PMID- 11104959 TI - [Cesarean section: anesthesia techniques and postoperative care]. PMID- 11104961 TI - Why should neuroradiology be a specialty? The Italian situation. PMID- 11104962 TI - Functional cranial neuronavigation. Direct integration of fMRI and PET data. AB - OBJECTIVE: We report our first experiences with the direct integration of fMRI data into cranial neuronavigation. METHOD: For navigation we used the MKM system and thin-sliced T1 contrast enhanced images. As a first step 21 patients had fMRI for localization of the precentral gyrus, 2 patients for Broca area detection. By anatomical correlation, these functional data were indirectly compared to the intraoperative findings using cortical SSEP (n=20) or cortical stimulation (n=3). Encouraged by these preliminary results, we started the direct integration of fMRI into neuronavigation in June 1999, followed by PET in January 2000, enabling us to compare functional images with intraoperative findings directly. fMRI and PET data were integrated by landmark matching referring on skin fiducials. Meanwhile, fMRI data of 8 patients (6 motorcortex, 2 Broca) and PET images of 1 patient were directly integrated into neuronavigation. Six out of 8 patients had additional cortical monitoring, 2/8 were exclusively operated on by functional neuronavigation. RESULTS: Using indirect comparison between fMRI and intraoperative findings we observed a good correlation in every case for the motorcortex, but only in 1/2 for the speech area. In all 6 direct integrated fMRI cases, these findings corresponded well to the conventional ones. Both patients with sole functional navigation did not have any postoperative neurological deficit. The inaccuracy of the fMRI ifT1 matching was 2. 7 mm (sigma=0.9 mm) and 1.3 mm (sigma=0.4 mm) of the subsequent referenciation of the navigation. The tumor delinement shown by 11C-methionine PET could be proven by intraoperative biopsy outside its indicated tumor margin. The inaccuracy of the PET matching was 0. 8 mm. CONCLUSION: Functional neuronavigation enables to visualize and preserve relevant brain areas. Other functional areas like short-term memory, which solely can be detected by fMRI might also be monitored in the future. The integration of PET data expect to gain a better differentiation of tumor and edema. PMID- 11104963 TI - [MRI of drug-resistant epilepsies: contribution of FLAIR sequence in a series of 150 patients]. AB - We studied the usefulness of the Fluid-Attenuated Inversion-Recovery (FLAIR) pulse sequence for patients with intractable seizures by reviewing the MR images of 150 consecutive patients including a standard imaging protocol and a FLAIR sequence. FLAIR images revealed a cortical lesion in 8 of the 81 patients with no lesion detected on the standard MR imaging protocol. In addition, FLAIR images provided additional informations for 13 patients of the 69 patients for whom a lesion was already detected on the standard MR imaging protocol. Therefore, our results indicated that FLAIR sequences were a useful imaging tool for patients with intractable seizures since it improved the MR diagnosis in 21 of 150 patients (14%). PMID- 11104964 TI - Comparison of a T2* w. 3D CISS and a T2 w. 3D turbo spin echo sequence for the anatomical study of facial and vestibulocochlear nerves. AB - Thirty healthy volunteers were examined with a T2* w. 3D CISS and a T2 w. 3D turbo spin echo (TSE) sequence in order to compare the facial and vestibulocochlear nerve detectability in the cerebellopontine angle and the internal auditory canal. CISS was significantly better than 3D TSE for nerve detectability in the cerebellopontine angle and equally as good as 3D TSE in the internal auditory canal. We would therefore recommend the inclusion of CISS in an MR imaging protocol of the facial and vestibulocochlear nerves. PMID- 11104965 TI - Rasmussen's encephalitis: proton MR spectroscopy and diffusion MR findings. AB - Rasmussen's encephalitis is a chronic inflammation of the brain that leads to progressive neurologic deficits. The condition has previously been studied by various imaging modalities including MR imaging and MR spectroscopy. We studied three patients presenting with Rasmussen's encephalitis by using proton MR spectroscopy, and diffusion-weighted MR imaging. In these patients, on diffusion weighted MR imaging, mean apparent diffusion coefficient (ADC) value was 1.74 x 10(-3) mm(2)/sec within the parenchyma, apparently higher than that of the normal parenchyma (0.88 x 10(-3) mm(2)/sec) of a control group of five healthy subjects. Proton MR spectra were obtained with a TR value of 1,500 msec and differing TE values (135, 40, and 20 msec), and were compared with a control group of fourteen cases. In the affected regions of the brain, MR spectroscopy revealed decreased NAA, and increased Cho peaks associated with apparently decreased NAA/Cho, NAA/Cr ratios, and increased Cho/Cr ratios. Slightly increased mI peaks, and increased mI/NAA ratios were noted. A prominent lactate peak was noted in one of the patients. PMID- 11104966 TI - Neurosarcoidosis: evaluation with MRI. AB - Clinical studies report a rate of 5% and autopsy results a rate of 25% of brain involvement in sarcoidosis. The aim of this study was to evaluate the role of magnetic resonance imaging (MRI) in the diagnosis of patients with neurosarcoidosis. The MRI brain scans of 22 patients with sarcoidosis were retrospectively reviewed, along with the clinical information that was provided in the request form. All patients had signs and symptoms referable to the head and were examined with gadolinium enhancement. Cranial (facial) nerve paralysis was the most common clinical manifestation identified in 10 patients. A wide spectrum of MR findings was noted: Periventricular and white matter lesions on T2W spin echo images, mimicking multiple sclerosis (46%); multiple supratentorial and infratentorial brain lesions, mimicking metastases (36%); solitary intraaxial mass, mimicking high grade astrocytoma (9%); solitary extraaxial mass, mimicking meningioma (5%); leptomeningeal enhancement (36%). These findings are not specific for sarcoidosis and one must consider appropriate clinical circumstances in arriving at the correct diagnosis. In selected cases with isolated brain involvement, meningeal or cerebral biopsy may be required. PMID- 11104967 TI - [Spontaneous regression of an acute spinal subdural hematoma. MR imaging]. AB - We report a case of acute spinal subdural hematoma which developed then regressed spontaneously. MR imaging contributed essential information for diagnosis and follow-up after conservative treatment. We made a short review of the literature and discuss the causes, pathogenesis, clinical presentation and usual treatment of acute spinal subdural hematoma. The differential diagnosis with the more frequent and extradural hematoma requiring immediate surgical evacuation is fundamental. PMID- 11104968 TI - [Intracranial hydatid cyst: contribution of scanography]. AB - We reviewed 14 cases of intracranial hydatid cysts, describing the clinical features and radiographic findings. The clinical presentation was dominated by signs of increased intracranial pressure and neurological manifestations. Diagnosis was established on the basis of CT findings, most valuable as a diagnostic tool. PMID- 11104969 TI - [An unusual cerebral hydatid cyst]. AB - Hydatid cysts of the brain are very rare. Typically, the cyst is large, spherical, with sharply defined borders. Its may calcify, and there is no surrounding edema and no rim enhancement. Sometimes, one side of the cyst wall is very close to the calvarium, with thinning of the internal table. We report a case of hydatid cyst of the brain with particular scanographic signs, bilobular with thin membranes in the posterior component. There was a destruction of the opposite calvarium with exteriorisation into hypodermic soft tissue. PMID- 11104970 TI - [A rare primitive neuroectodermal tumor: the medulloepithelioma]. AB - Primitive neuroectodermal tumors (PNET) is a name used to describe rare and highly malignant tumors composed of undifferentiated cells resembling germinal or matrix cells of the embryonic neural tube. These tumors occur most commonly in the first decade of life, and have a particular radiological, histological and evolution features. We report a case of a PNET in an 8-year-old boy who presented intracranial hypertension studied with computed tomography scan, which demonstrated a sharply circumscribed expansive mass in the frontoparietal deep white matter, with a large cystic component, which was considered at first as a glial tumor. It was treated by total excision, and the histologic study demonstrated a medulloepithelioma. We studied the concept of PNET and their pathological, radiological and prognostic features. PMID- 11104971 TI - [Syphilitic spinal cord gumma]. AB - Syphilitic spinal cord gumma Syphilitic gummas of the central nervous system are exceptional and are in general described in the brain. We report the case of a Brown-Sequard syndrome in a 25-year-old patient. The cervical myelography and the brain CT as well as the vertebro-occipital junction were normal. The diagnosis of multiple sclerosis was evoked and corticosteroid therapy was initiated. The patient experienced clinical improvement. Two years later, the patient was readmitted. Immunological reactions for syphilis were positive, in serum and CSF. Tests for HIV were negative. The MRI showed a cervical spinal cord process at the C3 level with adhesive spinal associated arachnoiditis. Penicillin therapy was started prior to surgery for the spinal process. The syphilitic nature was confirmed by pathology. To our knowledge, the MR appearance of a syphilitic gumma of the spinal cord has not been described previously in the scientific literature. PMID- 11104972 TI - Spinal dural arteriovenous fistula associated with syringomyelia. AB - The previously undescribed association of a spinal dural arteriovenous fistula with syringomyelia was found in a 60-year-old male, who developed increasing paresis, numbness of both lower extremities and sphincteric dysfunction. Symptoms and signs were attributed to a syringomyelia at T5-L1 and an arteriovenous spinal dural fistula at L1. The fistula was successfully immobilised with N-butyl-cyano acrylate. Six months after the procedure, all abnormalities had nearly disappeared. Whether the relation between the fistula and the syringomyelia was coincidental or causative could not be determined. PMID- 11104973 TI - [Hepatocellular carcinoma screening in patients with cirrhosis: a large French multicentric study (HCC)]. PMID- 11104974 TI - [Ultrasonic features of congenital anomalies of the gallbladder]. AB - Congenital anomalies of the gallbladder (GB) correspond to the ultimate manifestation of the wide range of anatomical variations that may affect the GB. Anomalies may be numerical: duplication, accessory GB, and, rarely, agenesis. Anomalies of position are more common: ectopic GB, floating or wandering GB. Morphological anomalies (septations) are the most common GB anomalies. Most of these anomalies may result in pitfalls at US imaging. PMID- 11104975 TI - [Multislice CT: principles and new CT-scan applications]. AB - The introduction of new array detector technology for multislice CT improves CT scan capabilities. Compared to single-slice helical CT, this technique offers three significant advantages: the pitch can be increased by a factor of 4, resulting in shorter acquisition times and contrast media saving, the temporal and spatial resolution are improved and the slice thickness can be freely and retrospectively selected. This technique promises to revolutionize radiological practice, just as spiral systems did a decade ago. Multislice CT is particularly suitable for exploring the chest, the heart and the vessels. It is also specially useful for musculo-skeletal explorations and trauma patients. PMID- 11104976 TI - [Cyclic filling cystourethrography in the study of febrile urinary tract infection in children]. AB - PURPOSE: To assess the value of cyclic voiding cystourethrography (VCUG) for the detection of reflux in a large population of children with urinary tract infection. MATERIALS AND METHODS: 234 patients (67% less than 3 years of age) underwent VCUG with two fillings in 214 cases and three fillings in 139 cases. Appearance or increase in the grade of reflux compared with previous filling was expressed as "modification of the radiology report" (MRR). The amount of contrast material, the duration of fluoroscopy and the number of films were recorded. RESULTS: The rate of reflux was 18.4% at the first filling, 16% at the second filling with 9.8% MRR and 14.7% at the third filling with 10% MRR. These results were the same for children younger and older than 3 years. For children under three years, if one considered reflux grade higher than 1, the MRR was 9.6% at the second filling and 7.5% at the third while it was 5.8% and 3% for older children. On average, the use of contrast material increased 50% with a third filling, number of films was not modified and the time of fluoroscopy increased by 6 sec per filling. CONCLUSION: VCUG is recommended in all children. The detection of reflux higher than grade 1 is more frequent with multiple fillings in children under 3 years. The increase in radiation exposure and cost seems negligible. PMID- 11104977 TI - [Comparative assessment of helical CT-angiography, 2D TOF MR-angiography and 3D gadolinium enhanced MRA in aorto-iliac occlusive disease]. AB - PURPOSE: To compare helical CT-angiography (CTA) and two techniques of MR angiography (MRA) to conventional angiography in aorto-iliac occlusive disease. MATERIALS AND METHODS: The abdominal aorta and iliac arteries in 22 patients (4 for preoperative assessment of abdominal aortic aneurysm and 18 for peripheral vascular disease) were imaged using four techniques: digital subtraction angiography ("gold standard"), 2D TOF MR angiography, 3D Gd-enhanced MR angiography and helical CT angiography. Source (CTA and 2D TOF MRA) and MIPed images (after subtraction measures before and after gadolinium injection for 3D Gd-MRA) were reviewed. RESULTS: Sensitivity, specificity and accuracy for the detection of significant (>50%) stenosis and occlusion of aorto-iliac arteries were respectively: 38%, 89%, 77% for 2D TOF MRA; 75%, 71%, 72% for 3D Gd-MRA and 95%, 90%, 92% for CTA. Excluding the internal iliac arteries, results were 54%, 96%, 88% for 2D TOF MRA; 96%, 80%, 83% for 3D Gd-MRA and 92%, 93%, 95% for CTA. CONCLUSION: 3D Gd-MRA, a technique with potential for further improvement, is superior to 2D TOF MRA for detecting significant stenosis and occlusion of aorto iliac arteries. Results at Gd-MRA are nearly similar to those at CTA (after excluding internal iliac arteries). Results at Gd-MRA are not affected by calcified plaque. PMID- 11104978 TI - [Percutaneous cholecystostomy in non-surgical patients]. AB - PURPOSE: To assess the efficacy and complications of percutaneous cholecystostomy (PC) in the treatment of acute cholecystitis in non-surgical patients. MATERIALS AND METHODS: Retrospective study of 25 cases (16 males and 9 females) of PC. The average age was 82 years (range: 59-95). Eight had acute acalculous cholecystitis (AAC) and 17 had acute calculous cholecystitis (ACC). US-guided percutaneous cholecystostomy was performed in most cases; CT-guidance was required in 5 cases. RESULTS: One technical failure and one complication (abdominal wall hematoma) occurred. PC was successful for sepsis control in 21 patients (5 AAC and 16 ACC): delayed cholecystectomy was performed in one patient, and one patient had recurrent acute cholecystitis at one month that responded to medical management. For the 4 remaining patients: 1 corresponded to the technical failure, and failure of sepsis control was observed in the 3 others patients (2 AAC, 1 ACC). PC was the definitive treatment or resulted in sepsis control in 84% of cases. PC was the definitive treatment, without recurrence, in 76% of cases. CONCLUSION: US or CT guided percutaneous cholecystostomy is an effective treatment, with a low rate of complication, in elderly or critically ill patients. PC can be used as a definitive treatment or as a temporizing measure in critically ill patients allowing for delayed definitive surgical/endoscopic management. PMID- 11104979 TI - [Radiography scale in rheumatology. State of the art and consequences for clinical practice]. AB - PURPOSE: Realize a "state of the art" of numerization and reduction or magnification scale of musculoskeletal radiography. Looking for the factors associated with this scale variation. How do they influence rheumatology practice. MATERIALS AND METHODS: The study recenced all the numerized (RN) and conventional radiography (RC) of one hundred consecutive patients consulting a rheumatologist practitioner. The scale was read directly or on gradations on the side of the radio. All the measure was made with a graduated magnifying glass. RESULTS: This study confirms that RN gradually replace RC in musculoskeletal radiology (0% before 1994, 65% in 1999). It also confirms that reducing scale is more frequent (50% in 1994, 82% in 1999). The scale of reduction is significantly different depending on some anatomic area, some diagnosis and some geographical provinces. These scale modifications disturb some recommended measures in rheumatology practice. The significant correlation between reducing scale and joint space (r=-0,32; p=0, 009; n=67) tend to show that scale reduction provide an overestimation of joint space. CONCLUSION: Given the perturbation induced in rheumatology practice it would be better to keep the scale of 100% until relevance of reducing scale has been demonstrated. PMID- 11104980 TI - [Gas-containing gallstones: value of the "Mercedes-Benz" sign at CT examination]. AB - Gas-containing gallstones are well-known in vitro. The typical triradiate arrangement of fissures filled with gas, first described on abdominal plain films, was named by Meyers the "Mercedes-Benz" sign. This sign is absent of the recent literature. We report a case where gas was the only CT sign suggesting the presence of gallstones in the gallbladder. PMID- 11104981 TI - [CT and MR imaging of a solitary fibrous tumor of the thigh]. AB - Solitary fibrous tumor is a rare mesenchymal tumor usually involving the pleura. Extrapleural lesions may also occur. We report the CT and MRI appearance of a solitary fibrous tumor of the thigh. The imaging features as well as the histological and immunohistochemical characteristics are presented, especially its reactivity to the CD34 antigen. The tumor is most often benign, particularly in extra-pleural location, with good prognosis after total resection. Imaging is mainly useful to locate the tumor and assess its extension prior to surgery. PMID- 11104982 TI - [Pneumoblastoma in an adult: a case report]. AB - Pneumoblastoma of the adult is rare. It presents more often as a large heterogenous and peripheral parenchymal mass. In this case, chest radiographs showed a small homogeneous mass in the right lung apex. The solid nature but non specific appearance of the mass was confirmed at CT. The histological diagnosis was obtained from surgical biopsy. The interest of this observation is the rare occurrence of the lesion in adults and the atypical imaging features in this patient. PMID- 11104983 TI - [Epithelioid hemangioendothelioma of the foot]. AB - Epithelioid hemangioendothelioma is a rare bone tumor. The authors report a case of a grade 1 multifocal epithelioid hemangioendothelioma of the foot in a 24-year old man treated by large surgical resection of the two lesions. PMID- 11104984 TI - [What is your diagnosis? Adventitial cyst of the right popliteal artery]. PMID- 11104985 TI - [Prize awarded by the national union of medical publishers and health professions to the Journal de Radiologie]. PMID- 11104986 TI - [Diagnostic and therapeutic management of common lumbago and sciatica of less than 3 months of duration. Recommendations of the ANAES. Agence Nationale d'Accreditation et d'Evaluation en Sante]. PMID- 11104987 TI - [AFIP--information. Armed Forces Institute of Pathology]. PMID- 11104988 TI - [Partial replantation following proximal limb injury]. AB - PURPOSE OF THE STUDY: Proximal replantation is a technically feasible but life threatening procedure. Indications must be restricted to patients in good condition with a good functional prognosis. The goal of replantation must be focused not only on reimplanting the amputated limb but also on achieving a good functional outcome. For the lower limb, simple terminalization remains the best choice in many cases. When a proximal amputation is not suitable for replantation, the main aim of the surgical procedure must be to reconstruct a stump long enough to permit fitting a prosthesis preserving the function of the adjacent joint. If the proximal stump beyond the last joint is very short, it may be possible to restore some length by partial replantation of spared tissues from the amputated part. We present here the results we obtained following this policy. MATERIALS AND METHODS: This series included 16 cases of partial replantations, 14 involving the lower limb and 2 the upper limb. All were osteocutaneous microsurgical transfers. For the lower limb, all transfers recovered protective sensitivity following tibial nerve repair. The functional calcaeoplantar unit was used in 13 cases. The transfer of this specialized weight bearing tissue provided a stable distal surface making higher support unnecessary. In one case, we raised a 13-cm vascularized tibial segment covered with foot skin for additional length. For the upper limb, the osteocutaneous transfer, based on the radial artery, was not reinnervated, but this lack of sensitivity did not impair prosthesis fitting. RESULTS: One vascular failure was finally amputated. This was the only unsuccessful result. For all other patients, the surgical procedure facilitated prosthesis fitting and preserved the proximal joint function despite an initially very proximal amputation. DISCUSSION: The advantages of partial replantation are obvious compared with simple terminalization or secondary reconstruction. There is no secondary donor site and, because there is no major muscle mass in the distal fragment, the overall risk is very low compared with the risk of total proximal leg replantation. PMID- 11104989 TI - [Conservative surgical treatment of osteogenic sarcoma of the limb in children and adolescents]. AB - PURPOSE OF THE STUDY: Advances in chemotherapy protocols over the last 20 years have considerably improved the prognosis and functional outcome in patients with osteogenic sarcoma. We report here the results of a cooperative study conducted under the auspices of the French Society of Pediatric Oncology (SFOP). Twenty nine oncology centers participated in this retrospective national multicentric study. MATERIALS AND METHODS: The study included 153 patients with osteogenic sarcoma of the limb who were treated by the OS87 protocol with conservative surgery between 1987 and 1994. The OS87 protocol consisted in conservative or non conservative surgery combined with pre- and postoperative chemotherapy. The following inclusion criteria were used: age under 20 years, tumor localization in a limb (pelvis and spine excluded), no metastasis at diagnosis, biopsy proven osteogenic sarcoma. RESULTS: Mean age at diagnosis was 13 years. The knee localization predominated (80 p. 100). 82.5 p. 100 of the patients had grade IIB disease (Enneking classification). For the 187 patients included in the protocol surgery was non-conservative in 20 p. 100 of the cases and conservative in 80 p. 100. The choice of the surgical technique (arthroplasty, allograft, autograft, resection without reconstruction) depended on the patient's age and school situation. Data analyzed here concerned only those patients who had conservative treatment. Mean follow-up was 64 months. The actuarial survival curve plateaued at 71 p. 100 at more than 6 years. Early and late complications were numerous and variable (mechanical, infectious, local recurrence). Secondary amputation was required in 10 p. 100 of the patients. The overall functional outcome of the preserved limbs was nevertheless good with rapid restoration of self-sufficiency despite major surgery and a high number of reoperations (about 65 p. 100 of cases). DISCUSSION: In light of the frequency and the seriousness of the complications, these results are modest. Patients and family should be advised of the risk, particularly the risk of secondary amputation which may be required early due to contaminated excision or at mid term due to major non-cancerological complications. As survival has been improved, functional capacity must be preserved for several years. This orients surgery towards more "biological" reconstruction which can provide greater longevity than arthroplasty. PMID- 11104990 TI - [Sacral chordoma: retrospective review of 11 surgically treated cases]. AB - PURPOSE OF THE STUDY: Chordoma is a malignant neoplasm believed to arise from notochord remnants. It accounts for approximately 3 to 4 p. 100 of primary bone tumors and is localized along the axial skeleton, 50 p. 100 being sacrococcygeal. Clinical, radiographical and histological findings have been well established since the first description by Ribbert in 1894. Sacral chordomas are however difficult to manage and remain a challenge for surgeons and radiotherapists alike. The purpose of this study was to evaluate the long-term results of surgical treatment and patterns of failure in patients treated for chordoma of the sacrum in our department. MATERIALS AND METHODS: This retrospective study included 11 cases of sacral chordomas treated from 1973 to 1998. Patient age ranged from 36 to 77 years (mean 59 years). Six patients were female and five male. The initial treatment was surgery in all cases including intralesional removal in two cases, marginal resection in seven and complete en bloc resection in two. RESULTS: Median follow-up was 6 years (1 month to 14 years). Tumoral recurrences were observed in nine cases 5 months to 8 years after treatment. In two cases, recurrence was observed 8 years after radical sacrectomy. Treatment of recurrences was partial surgical removal with radiotherapy (40 to 70 Grays). Three patients developed metastases in lungs, liver and bone, respectively. Seven patients died, two from metastatic disease. The 5-year overall survival was 64 p. 100 but only 18 p. 100 of the patients survived 10 years. Average disease-free survival was 18 p. 100 at 5 years and 0 p. 100 at 10 years. DISCUSSION: Chordoma is a slow-growing tumor allowing survival for several years despite recurrent disease. However, only 10 to 20 p. 100 of the patients survive free of disease at 5 years. Recurrences are frequent (45 to 80 p. 100) and often multiple. Chordoma inevitably recurs and eventually leads to death after intralesional removal or marginal resection. Radical surgery should be attempted whenever technically feasible. When performed early, particularly for smaller lesions, it offers the best chance for cure. However, tumoral recurrence can occur postoperatively despite a macroscopically complete resection. Because radiation therapy seems to be more successful in controlling microscopic disease, it should be considered as a pre- or postoperative adjuvant to a macroscopically complete resection. PMID- 11104991 TI - [Revision in non-infected total knee arthroplasty: an analysis of 69 consecutive cases]. AB - PURPOSE OF THE STUDY: We reviewed 69 consecutive cases of total knee arthroplasty revisions to analyze the causes of failure. MATERIAL AND METHODS: Sixty-nine total knee arthroplasty revisions were required between 1990 and 1997 for non septic failure. Five categories of failures were identified: 30 loosenings including 11 with an initial malposition (varus position of the tibial component in 8 cases), 14 laxities (medial in 5, lateral in 5 and anteroposterior in 4), 11 stiff knees with no other clinical or radiological anomaly, 6 patellar failures (2 dislocations, 2 cases of excessive wear, 2 painful knees with a Freeman prosthesis), and 8 cases of painful knees with no other detectable anomaly. RESULTS: A three-phase reconstruction procedure was used after removing the failing TKA: 1) reconstruction of the tibia with replacement of lost bone, 2) reconstruction of the femur with balanced flexion determining the size of the implant, 3) balanced extension determining the distal/proximal position of the femoral component. A "simple" sliding prosthesis was used in 16 cases, a modular reconstruction prosthesis in 40 cases and a hinge prosthesis in 13 cases. Mean follow-up for functional and radiographic assessment after revision surgery was 37 months (59 cases) with a minimum follow-up of 1 year. The best outcome was observed in the "loosening", "laxity", and "stiffness" patients. Outcome was less favorable for the group "isolated pain" with IKS functional scores of 35.5 +/- 16 and 52.5 +/- 21. DISCUSSION: In 36 p. 100 of cases, TKA failure was related to a technical mistake (component malposition, poor ligament alignment). In 33 p. 100, failure was patient related (multiple procedures, congenital hip dysplasia, rheumatoid arthritis.). Outcome after revision TKA was less favorable than after primary TKA, particularly in case of painful knees with no other detectable anomaly. CONCLUSION: Surgical revision of TKA must follow a rigorous procedure with a detailed preoperative work-up. The decision for revision must not be made unless a precise anomaly has been identified. PMID- 11104992 TI - [Screws versus screw-plate fixation of type 2 Schatzker fractures of the lateral tibial plateau. Cadaver biomechanical study. Arthroscopy French Society]. AB - PURPOSE OF THE STUDY: We compared in vitro the efficacy of screw-plate fixation versus double screw fixation on a model of type 2 Schatzker fracture of the lateral tibial plateau. MATERIALS AND METHODS: Ten screw-plate fixations using a lateral prebent plate and 10 double-screw fixations (6.5 mm screws) were made on 10 pairs of non-embalmed cadaver knees after simulation of type 2 Schatzker fractures. The strength of each fixation was tested with a compression device. Criteria indicating failure were displacements greater than 2 mm of one or more fracture lines. The force applied at rupture and the stiffness of each type of fixation were compared. Wilcoxon's test was used for statistical analysis. RESULTS: Force at rupture and stiffness of the fixation were similar for the two types of fixation. There was no statistical difference (p > 0.05) between the screw-plate and the double-screw fixations. DISCUSSION: Our findings on a model of type 2 Schatzker fractures are in agreement with previous data obtained by other authors working on models of type 1 Schatsker fractures. The biomechanical stability of the double-screw fixation is as good as that obtained with screw plate fixation for the treatment of fractures of the lateral tibial plateau. PMID- 11104993 TI - [Stretching the triceps surae muscle after 40 degrees C warming in patients with cerebral palsy]. AB - PURPOSE OF THE STUDY: Equinus in patients with cerebral palsy results from at least two factors: excessive contracture of the triceps surae and muscle retraction. Tendon surgery and progressive lengthening techniques using plaster walking boots can provide variable improvement in retraction. We compared the effect of this technique when applied with or without prior 40 degrees C warming in the same patients. We also assessed the efficacy of this treatment method in terms or degree of retraction, patient age, puberty maturity, and sex. MATERIALS AND METHODS: This series included 70 muscles in 52 patients with cerebral palsy aged 2 years 11 months to 21 years (mean 8 years 3 months). Common features in these patients were: - equinus mainly explained by triceps retraction, - no history of prior surgery on the triceps tendon, - knee flexion less than 15 degrees in the upright position, - easily reduced lateral deformation of the foot, - absence of mediotarsal dislocation, - triceps stretching could be achieved without triggering unacceptably intense contracture. The retraction of the triceps surae was measured from the maximal passive dorsal flexion angle of the foot, before and after applying each stretching boot. The difference between these measurements gave the gain obtained with the plaster boot. Protocol R- (stretching with plaster boot) consisted in a series of slow stretchings for 10 minutes before making the boot which was worn 7 days. Recurrent retraction in these same patients warranted another treatment within a delay of 3 to 17 months (mean delay 8.7 months). The same treatment then followed protocol R+ where the stretching was preceded by immersion of the segment in a 40 degrees C water bath for 10 minutes. RESULTS: Mean gain obtained with protocol R+ (warming) was 6.8 degrees knee extended and 7.1 degrees knee flexed. These differences were highly significant in both cases (p <0.0001). We had no failures with protocol R+ while with protocol R- (stretching without warming) the gain was nil or less than 5 degrees for 29 muscles knee extended and for 32 muscles, knee flexed. The gain was not related to age, sex or puberty maturity. It was not related to the angle of dorsal flexion of the foot prior to stretching. DISCUSSION: Our findings demonstrate that when the conditions allowing prolonged stretching of the triceps surae are present, prior warming at 40 degrees C for 10 minutes leads to an improvement in muscle lengthening in all patients, even in those for whom prior treatment had been unsuccessful without warming. This observation would indicate that the mechanisms allowing greater lengthening are present in all patients with cerebral palsy but that they cannot be triggered due to abnormal muscle viscosity related to distal vasomotor disorders frequently observed in this condition. Further research is needed to detail this point. PMID- 11104994 TI - [Surgical treatment of chronic Achilles tendiopathies. Report of 52 cases]. AB - PURPOSE OF THE STUDY: We reviewed a series of 52 cases of chronic Achilles tendinopathy treated surgically by release of the fascia cruris, resection of peritendon, longitudinal incision of the tendon and occasional excision of intratendinous lesions. MATERIALS AND METHODS: The mean course prior to surgery was about 18 months. Twenty-six patients practiced sports. Complaints were bilateral in 12 cases. Pain was always present. Ultrasound exploration evidenced paratendinitis (n=21), tendinosis (n=22) and paratendinitis with tendinosis (n=9) (Puddu classification). Patients were reviewed after a minimal 2-year follow-up. Results were assessed on the basis of clinical findings. RESULTS: Mean follow-up was 5 years 6 months. Twenty-nine patients were free of pain. The range of motion was normal in 48 cases and 29 patients resumed sports activities at the same level as prior to surgery. Outcome was very good in 29 patients, good in 14 average in 6 and poor in 3. DISCUSSION: Stiffness of the tibio-tarsal joint can be avoided by proper mobilization. Outcome appears to be better in middle-aged patients. Poor outcome is closely related to amyotrophy. The presence of a foot deformity does not appear to have an unfavorable influence on outcome. The Achilles tendon must not be infiltrated. Ultrasound is highly contributive, but MRI provides a more accurate analysis. CONCLUSION: Surgical treatment of chronic Achilles tendinopathies can be proposed when conservative treatment has been unsuccessful. Outcome is better in young active patients and in cases where paratendinitis predominates. PMID- 11104995 TI - [Simple screw fixation for calcaneal fractures: 60 cases with preoperative computed tomography analysis]. AB - PURPOSE OF THE STUDY: In accordance with the conclusions established at the SOFCOT symposium in 1988, we propose surgical treatment of displaced fractures of the calcaneus with screw fixation after reduction. We developed an original classification system of 3D computed tomography images which allows a precise description of the fractures and guides joint and calcaneal body reconstructions. The purpose of this work was to provide a precise analysis of operated fractures in order to identify prognostic factors and validate use of exclusive screw fixation for calcaneal fractures. MATERIALS AND METHODS: This series included 60 operated articular fractures of the calcaneus. The Utheza classification was: 12 vertical, 7 horizontal with 1 fracture line, 3 horizontal with 2 fracture lines, 23 mixed with 1 fracture line and 15 mixed with 2 fracture lines. 3D computed tomography evidenced the fundamental fracture lines and their anterior extension. Fixation was achieved with one screw inserted in a transverse position under the posterior facet and one oblique screw from the greater tuberosity to the sustentaculum tali. The medial and lateral Bohler angles were measured on plain x rays. The analysis included search for a double line on the posterior talocalcaneal facet, secondary body displacement, the position of the oblique screw and the degree of posttraumatic subtalar wear. The clinical criteria established in the 1988 SOFCOT guidelines were recorded. Analysis of variance, Pearson and Spearman coefficients, and RIDITS analysis (the most powerful method available for evidencing a relationship between two qualitative variables one of which is ordinal) were used to search for prognostic elements and correlations. RESULTS: No severe complications were encountered with the wide lateral access. A negative medial Bohler angle was significantly correlated with an additional posterior facet line. A mean 80 p. 100 reduction in the lowering of the medial part of the posterior facet and an 87 p. 100 reduction in lateral pivoting were achieved irrespective of the type of fracture. Minimal secondary body displacements were significantly related to anchorage of the oblique screw outside the sustentaculum tali. Functional outcome was satisfactory (very good + good + average) in 75 p. 100 of the cases and physical outcome in 50 p. 100 (very good + good) irrespective of the type of fracture. Outcome was significantly correlated with reduction in the Bohler angle, double lines on the posterior facet, secondary displacement and osteoarthritis. DISCUSSION: The 3D analysis of posterior facet fractures using our classification was useful in guiding reconstruction with correction of the medial lowering and the lateral pivoting. A negative medial Bohler angle was a factor of poor prognosis: more posterior facet lines, joint wear and deterioration of the functional and physical outcome. Good outcome required good reduction of the Bohler angle and good anchorage of the oblique screw in the sustentaculum tali. Good subtalar mobility was associated with pain relief. Uniform anatomic and pathologic classifications and precise analysis criteria are needed for pertinent comparison between series and proper definition for indications for first-line reconstruction-arthrodesis. CONCLUSION: Measurement of the medial Bohler angle improves the sensitivity of revision criteria for articular fractures of the calcaneus. Screw fixation has proven its reliability. PMID- 11104996 TI - [Neurofibromatosis of the lower cervical spine: an operative case report]. AB - PURPOSE OF THE STUDY: We report a case of type 1 neurofibromatosis (von Recklinghausen's disease) of the lower cervical spine in a 13-year-old girl. CASE REPORT: There was no neurological deficit. Plain films showed dysplastic 82 degrees kyphosis centered on the C4-C5 disc. Surgical treatment consisted in anterior multilevel interbody grafting and plate osteosynthesis combined with posterior arthrodesis. Good bone fusion was obtained with acceptable cervical mobility. The residual cervical kyphosis was 18 degrees. DISCUSSION: An evaluation of the cervical spine should be proposed for patients with neurofibromatosis even if there is no thoracic scoliosis. Severe cervical deformities can lead to serious neurological complications. Circumferential arthrodesis appears to provide optimum results. PMID- 11104997 TI - [Absence of the radial artery: case report and review of the literature]. AB - PURPOSE OF THE STUDY: We report a case of complete unilateral absence of the radial artery in the forearm and reviewed the pertinent literature. CASE REPORT: An 18-year-old girl was admitted for multiple fractures after a car accident. She presented with a comminuted fracture of the left distal humerus, an open grade I fracture according to the Gustilo classification involving the right ulna and radius, a mediodiaphyseal fracture of the right femur and an open grade II fracture of the proximal and distal left tibia. After open reduction and internal fixation of the bones of the right forearm, she presented transient ischemia of her right hand, the radial pulse not being detectable at the end of surgery. An arteriography showed a complete absence of the right radial artery, which was thought to be caused by arterial thrombosis. Surgical exploration evidenced the complete absence of the radial artery. DISCUSSION: Absence of the radial artery is observed in radial preaxial hemimelia, in specific genetic and chromosomal disorders (Fanconi's anemia, Holt-Oram syndrome) and in association with other malformations. Unilateral absence of the radial artery has been described in association with other vascular abnormalities such as a larger anterior interosseous artery or the presence of a medial artery. Our case presented an isolated anatomical variation of the radial artery. This vascular anomaly was asymptomatic and discovered fortuitously. The incidence of this anatomic anomaly may be underestimated in the general population. PMID- 11104998 TI - [Re: Pseudotumoral subacute osteomyelitis: a series of 41 children]. PMID- 11104999 TI - The Korean Academy of Prosthodontics--founded in 1959. PMID- 11105000 TI - American Academy of Implant Prosthodontics--founded in 1982. PMID- 11105001 TI - News from the editorial council of the journal of prosthetic dentistry PMID- 11105002 TI - Five-year follow-up of InCeram laminate restorations: a clinical report. PMID- 11105003 TI - Direct tissue stops for distal extension removable partial dentures. AB - Supplemental impression++ strategies may be used to capture optimal registration of residual ridge tissues in distal extension-base RPDs. A procedure is described to adapt tissue stops in vivo and positively position the framework to the master cast when clinically using corrected impressions. This method is simple and cost effective, and it promotes accurate prosthetic-tissue relationships during clinical and laboratory phase of RPD fabrication. PMID- 11105004 TI - Periodontal tissue responses after insertion of artificial crowns and fixed partial dentures. AB - PURPOSE: The purpose of this review was, first, to critically evaluate published evidence on the effects of artificial crowns and fixed partial dentures (FPDs) on adjacent periodontal tissue health, and second to synthesize this evidence into meaningful summaries. Restoration qualities that contribute to inflammatory responses were identified based on strength of evidence, and variables that should be controlled in future investigations were outlined. Such information is necessary to accurately predict the prognosis of periodontal tissues adjacent to crowns or FPDs. METHODS: Clinical trial and epidemiologic evidence published in English was collected. The effects of crowns or FPDs on gingival inflammation, probing depths, and bone loss were evaluated based on accuracy of measurement, reliability of measurement, and/or appropriateness of data analysis. RESULTS: Crowns and FPDs increased the incidence of advanced gingival inflammation adjacent to restorations, particularly if restorations had intracrevicular finish line placement, poor marginal adaptation, or rough surfaces. However, because of the limitation in the accuracy and reliability of probing depth measurements, reports of greater mean probing depths of crowned teeth, which tended to be less than 1 mm greater than control teeth, should be questioned. Finally, crowns and FPDs in general did not accelerate the rate of adjacent bone loss. CONCLUSION: Clinically deficient restorations, as well as clinically acceptable restorations, can contribute to gingival inflammation. However, with the limitations of the applied methods of measurement, current evidence has not shown an increased attachment loss adjacent to crowns or FPDs. Future trials should document periodontal health before therapy and periodically after restoration insertion so that each tooth serves as its own control. In future studies, the periodontal disease history of the patient, the influence of the restoration on plaque formation, and the composition of the crevicular microflora must be recorded. PMID- 11105005 TI - Effect of variable light intensity on composite shrinkage. AB - STATEMENT OF PROBLEM: Polymerization shrinkage is a critical limitation of dental composites and may contribute to postoperative pain, tooth fracture, microleakage, and secondary caries. Polymerization with high-intensity light sources has been related to increased depth of cure and improved mechanical properties. However, high-intensity light initiation has also been associated with greater polymerization shrinkage. PURPOSE: The purpose of this study was to investigate the effect of sequentially increasing light intensity on the polymerization shrinkage of 2 composites, a hybrid and a microfil. A Knoop hardness test was used to evaluate effectiveness of the cure with each intensity increase. MATERIAL AND METHODS: Four groups of 12 samples were measured for polymerization shrinkage by using a linometer. Light intensity curing sequences were as follows: full-intensity control (100% intensity for 40 seconds), low intensity control (25% intensity for 40 seconds), test group 1 (25% intensity for 20 seconds, 50% for 10 seconds, 100% for 10 seconds), and test group 2 (25% intensity for 10 seconds, 50% for 10 seconds, 100% for 20 seconds). Statistical comparisons were made using a 1-factor ANOVA and a Tukey multiple comparisons test within each material. RESULTS: Results showed a significant difference (P<. 05) in mean linear shrinkage between the full-intensity control group and the other 3 sequences for both composites. No difference existed within the other 3 groups for either composite. Knoop hardness was similar for the full-intensity control and test group 2. The low-intensity control group and test group 1 were also similar but significantly lower. CONCLUSION: Curing composites for 10 seconds at 25% intensity, 10 seconds at 50%, and 20 seconds at 100% significantly reduced polymerization shrinkage while not compromising depth of cure. PMID- 11105006 TI - Marginal fit and short-term clinical performance of porcelain-veneered CICERO, CEREC, and Procera onlays. AB - STATEMENT OF PROBLEM: Onlay preparations are very complex surfaces for computer surface digitization, CAD, and CAM of all-ceramic onlay cores. PURPOSE: This study tested the hypothesis that onlays can be fabricated with CICERO, CEREC, and Procera core technologies. MATERIAL AND METHODS: Fifteen mandibular and 10 maxillary molars were prepared for onlays in 17 patients (11 women and 6 men). The onlay design was experimental. Molars were prepared with deep gingival chamfers in the proximal boxes and around the functional cusps. The nonfunctional cusps were prepared with broad bevels. Eight stone dies of preparations were measured with a laser beam (CICERO), 10 dies with a light beam (CEREC), and 7 dies with a contact probe (Procera). Two onlay cores were produced for the same stone die. One core was used to analyze fit on the stone die, and the other core was porcelain veneered for optimizing anatomy, esthetics, and fit of the onlay and cemented. The fit of the onlay core on the stone die and the cement width on a stone cast were measured by a microscopic digital imaging system. The onlays were evaluated for function every 6 months for 2 years. RESULTS: Measurements of the margins by the CICERO system were (1) precise (error <4%) and (2) accurate with an SD of less than 9 microm. The proposed onlay preparation design met the requirement that all points of the surface be visible from a single point of view for optical 3-dimensional mapping by the CEREC system. For the surface measurements by the Procera contact probe, the orientation of the sapphire tip toward the preparation surface was critical, and it was necessary to apply wax to smooth internal edges. The marginal gaps of the CICERO, CEREC, and Procera cores on the stone dies were 74 microm (SD 15), 85 microm (SD 40), and 68 microm (SD 53), respectively. The cement width was 81 microm (SD 64). No fractures occurred. CONCLUSION: Marginal gaps for the onlay cores were no more than 85 microm. The cement width of the semicomputer-produced onlays of 81 microm was a favorable measurement value for a clinically acceptable, strong all-ceramic onlay. However, this value as well as anatomy and esthetics of the onlay depended on the craftsmanship of the porcelain veneering by the dental technician. PMID- 11105007 TI - Prospective clinical 5-year study of ceramic-veneered titanium restorations with the Procera system. AB - STATEMENT OF PROBLEM: The biocompatibility of titanium has been well documented, but clinical outcomes of ceramic-veneered titanium restorations were not conclusive. PURPOSE: The study presents the results of a 5-year clinical study of individual crowns and fixed partial dentures (FPDs) veneered with a low-fusing ceramic of the Procera system. MATERIAL AND METHODS: All patients at one clinic who required crowns or FPDs during a 2-year period were invited to participate. A total of 260 patients received 333 ceramic-veneered Procera restorations (242 single crowns and 91 FPDs). Clinical registrations were performed annually, and the restorations were evaluated according to the California Dental Association rating system. At the 5-year follow-up, 198 (76%) patients were examined. Most of the loss of patients could be explained. RESULTS: Practically all Procera restorations were judged as satisfactory both at baseline and follow-up examinations. One artificial crown and 1 FPD were remade because of extensive fractures of ceramic veneers. Two FPDs had fractures of a soldered joint, but only 1 FPD was replaced. Some minor complications occurred, such as small porcelain fractures that could be polished (6% of single crowns, 13% of FPDs) and or loosened restorations that were recemented. Other recorded complications were not related to the Procera system but to dental caries, loosened posts and cores, and root fractures. CONCLUSION: Clinical outcomes over a 5-year period for ceramic-veneered titanium restorations with the Procera system were favorable. PMID- 11105008 TI - A role for surface topography in creating and maintaining bone at titanium endosseous implants. AB - STATEMENT OF PROBLEM: A variety of claims are made regarding the effects of surface topography on implant osseointegration. Many in vivo and in vitro experimental observations have key limitations in their interpretations. PURPOSE: This review considers the major claims made concerning the effects of commercially pure (cp) titanium implant surface topography on osseointegration. Important findings of consensus are highlighted, and existing controversies are revealed. MATERIAL AND METHODS: This review considers many of the research publications listed in MEDLINE and presented in biomedical research publications and textbooks. RESULTS: Implant surface topography is not well defined in the marketplace or consistently reported among experimental studies. Many in vitro evaluations are not predictive of or correlated with in vivo outcomes. In some culture models, increased surface topography positively affects pro-osteogenic cellular activities. Animal models reveal modest increases in bone-to-implant contact and increases in the biomechanical interlock of the implant with bone for implants of increased surface topography. Existing information fails to define increased surface topography as a risk factor for peri-implant inflammation. CONCLUSION: Increased cp titanium implant surface topography improves the bone-to implant contact and the mechanical properties of the enhanced interface. Growing clinical evidence for increased bone-to-implant contact at altered cp titanium implants confirms the temporally limited observations made in preclinical studies. In the absence of controlled comparative clinical trials, the aggregate experimental evidence supports the use of cp titanium implants with increased surface topography. PMID- 11105009 TI - In vivo force measurements on maxillary implants supporting a fixed prosthesis or an overdenture: a pilot study. AB - STATEMENT OF PROBLEM: In this preliminary study, an attempt was made to measure in vivo forces simultaneously on 5 maxillary implants with different types of superstructure. MATERIAL AND METHODS: Force measurements were carried out on 1 test patient with 5 ITI implants in the edentulous maxilla. A screw-retained fixed complete denture and an overdenture were fabricated for comparative measurements of forces. The overdenture could be mounted to 2 different types of bars. The measuring method was used with piezo-electric force transducers that were directly mounted onto the implants. This allowed for simultaneous measurements of forces in 3 dimensions, ie, in axial and transverse directions. Static and functional forces such as maximum biting (clenching), biting on a bite plate, and chewing food were registered. All measurements were repeated in the same way 2 years later. RESULTS: The registered forces exhibited similar force patterns with both types of superstructure and both types of bars for overdenture connection. The force magnitudes were significantly different for the 3 dimensions (P<.05) with highest forces along the implant axis. On the posterior implants, force magnitudes were significantly higher (P<.05) than on the anterior implants in all 3 dimensions. On the anterior implants, under some test conditions, the transverse force components reached up to 100% of the axial force or even exceeded it during the chewing of food. During maximum biting, no upward force (tensile force) was found on any implant with the fixed complete denture, but upward force was found on one anterior implant with the overdenture. When chewing food, small force magnitudes in upward directions were regularly found with both superstructures. The force patterns between the first and second registrations showed similar trends, and no obvious differences were found. CONCLUSION: From these results it was concluded that similar patterns of force transmission onto the implants are observed with a fixed complete denture and an overdenture connected to maxillary implants. The bar design did not significantly influence the force pattern. PMID- 11105010 TI - Measurement of the dimensions and abutment rotational freedom of gold-machined 3i UCLA-type abutments in the as-received condition, after casting with a noble metal alloy and porcelain firing. AB - STATEMENT OF PROBLEM: Laboratory processing of implant-supported prostheses may alter the surface of the abutment in contact with the implant head and thus the interface fit. PURPOSE: This study assessed changes at the implant interface of gold-machined UCLA abutments after casting and porcelain baking in the case of single-tooth restorations. MATERIAL AND METHODS: The depth (d) and width (w) of the hexagonal portion of the abutment, the apical diameter (D) of the abutment, and the abutment rotational freedom (R) were assessed for 30 gold-machined UCLA abutments before casting procedures (time 0), after casting with a noble metal alloy (time 1), and after the addition of porcelain (time 2) to detect any eventual fitting change in the abutments on the top of the implant hexagon. RESULTS: No significant differences relative to all study parameters (d, w, D, and R) were observed between times 0, 1, and 2 (P=.576). CONCLUSION: The results of this investigation suggest that, if all laboratory steps are observed carefully, changes at the implant interface of gold-machined UCLA abutments do not occur. PMID- 11105011 TI - Effect of a new metal primer on the bond strength between a resin cement and two high-noble alloys. AB - STATEMENT OF PROBLEM: With the development of new adhesive resin cements, the question of surface treatment of noble metal castings with primers has become an important issue. PURPOSE: This study compared the tensile bond strength and its durability of a new metal primer (Alloy Primer, Kuraray) to 2 noble metal alloys (Au-Ag-Cu-Pt and Au-Pt-Pd-Ag-In). MATERIAL AND METHODS: Sixty cast disk specimens of each alloy were polished, grit blasted with 50 microm Al(2)O(3), and ultrasonically cleaned in 96% isopropanol. Then, they were either nonprimed or primed only with the Alloy Primer or Alloy Primer combined with ED Primer (Kuraray). Plexiglas tubes filled with self-curing composite resin (Clearfil FII, Kuraray) were bonded to the metal samples with the use of an alignment apparatus and a self-curing luting cement (Panavia 21 Ex). The samples were stored in water, either for 3 days with no thermal cycling or for 150 days with 37,500 thermal cycles. After the different storage conditions, the tensile bond strengths of the specimens were determined. RESULTS: The mean bond strengths increased over storage time for all groups, except for the grit-blasted Au-Pt-Pd Ag-In group. However, only in the grit-blasted and the primed groups for the Au Ag-Cu-Pt alloy was this increase significantly different (P<.01). After 150 days of storage, the mean bond strength to Au-Ag-Cu-Pt alloy was 38.8 MPa without priming, whereas it was 40.6 to 40.8 MPa with the use of the primers. After the same time, the mean bond strength to the Au-Pt-Pd-Ag-In alloy was 20.6 MPa without priming, whereas it was 31. 9 to 37.8 MPa with the use of the primers. When comparing the different bonding methods and different storage times for the alloys, the superiority of the usage of both primers in combination was determined. Conclusion. The tested Alloy Primer significantly improved the bond strength of the dental adhesive resin cement (Panavia 21 Ex) to noble alloys. However, this effect depended on the alloy composition and was much greater for the Au-Pt-Pd-Ag-In alloy than for the Au-Ag-Cu-Pt alloy. PMID- 11105012 TI - Comparison of the tensile bond strength of high-noble, noble, and base metal alloys bonded to enamel. AB - STATEMENT OF PROBLEM: Although the bond strengths of various resin composite luting materials have been reported in the literature, the evaluation of these systems with various cast alloys of different compositions has not been completely clarified. PURPOSE: To evaluate the tensile bond strength of sandblasted high-noble, noble, and base metal alloys bonded to etched enamel by 2 different bonding agents of different chemical composition: Panavia-Ex (BIS-GMA) and Super-Bond (4-META acrylic). MATERIAL AND METHODS: Flat enamel surfaces were prepared on buccal surfaces of 60 extracted noncarious human incisors. Teeth were divided into 3 groups of 20 each. Twenty circular disks of 5 mm diameter were prepared for casting for each group. Group I was cast with a high-noble, group II with a noble, and group III with a base metal alloy. The surfaces of the disks were sandblasted with 250 microm Al(2)O(3). Ten disks of each group were bonded to exposed enamel surfaces with Super-Bond and 10 disks with Panavia-Ex as recommended by the manufacturer. The tensile bond strength was measured with an Instron universal testing machine with a crosshead speed of 0.5 mm/min until failure occurred. RESULTS: Two-way ANOVA was used to evaluate the results. The differences in bond strengths of Super-Bond and Panavia-Ex with different alloys were not significant. The highest bond strengths were obtained in base metal alloys, followed by noble and high-noble alloys. These results were significant. CONCLUSION: Panavia-Ex and Super-Bond exhibited comparable tensile bond strengths. For both luting agents, the highest bond strengths were achieved with base metal alloys and the lowest with high-noble alloys. PMID- 11105013 TI - Bond strength of three porcelains to two forms of titanium using two firing atmospheres. AB - STATEMENT OF PROBLEM: Problems with casting and porcelain bonding are encountered when titanium is used in metal-porcelain restorations. The oxidation characteristics of titanium are the main problem. The bonding mechanisms in titanium-porcelain systems are complex and poorly understood. PURPOSE: An in vitro investigation was performed to evaluate the bonding characteristics of 3 titanium-porcelain systems in various firing conditions. MATERIAL AND METHODS: This study evaluated the bonding strength of 3 commercial titanium porcelains fired in a vacuum and in an argon atmosphere to cast and noncast commercially pure titanium, using a 3-point bending test according to DIN 13927 and SEM with energy-dispersive spectrometry analysis. The results were compared with an Ni-Cr alloy and a conventional porcelain that was chosen as a control. RESULTS: The Ni Cr-conventional porcelain system fired in an argon atmosphere had significantly higher bond strength than the other systems (P<.001). In addition, the bond strength of the titanium-spark erosion-Noritake Ti22 combination, fired in an argon atmosphere, was significantly higher than the other titanium-porcelain groups, which had results similar to those obtained with the vacuum-fired, Ni-Cr conventional porcelain and argon-fired titanium-cast-Noritake Ti22 groups. On the other hand, the bond strength of the titanium-TiBond and titanium-Vita Titankeramik groups was below the lower limit value in the DIN 13927 standard for the 3-point bending test (25 MPa). Although the results of the Duncan multiple range test showed that firing in an argon atmosphere did not affect the bond strength of the titanium-Vita Titankeramik groups, the titanium-spark erosion TiBond group, or the titanium-cast-Noritake Ti22 group, argon firing improved the bond strengths of the Ni-Cr-conventional porcelain group, the titanium-cast TiBond group, and the titanium-spark erosion-Noritake Ti22 porcelain group. It was also found that there were no significant differences between the bond strengths of cast and non-cast titanium groups; an exception was the titanium TiBond groups in which the porcelain was fired in a vacuum. CONCLUSION: The oxide layer produced on titanium was considered to have a potentially adverse effect on titanium-porcelain bonding. It was also concluded that matching the titanium porcelain combination is the main determinant for optimal bonding. Firing in an argon atmosphere that limited the oxidation of titanium improved the titanium porcelain bond in some of the groups. PMID- 11105014 TI - Effect of pressure of helium, argon, krypton, and xenon on the porosity, microstructure, and mechanical properties of commercially pure titanium castings. AB - STATEMENT OF PROBLEM: Porosity is a frequently observed casting defect in dental titanium alloys. PURPOSE: This study evaluated the effect of pressure of helium, argon, krypton, and xenon on the porosity, microstructure, and mechanical properties of commercially pure titanium (cp Ti) castings. MATERIAL AND METHODS: Eight groups (A-H) of 16 rectangular wax patterns each (30 mm in length, 3 mm in width, and 1 mm in depth) were prepared. The wax patterns were invested with a magnesia-based material and cast with cp Ti (grade II). Groups A, C, E, and G were cast under a pressure of 1 atm, and groups B, D, F, and H were cast under a pressure of 0.5 atm of He, Ar, Kr, and Xe, respectively. The extent of the porosity of the cast specimens was determined radiographically and quantified by image analysis. Three specimens of each group and 3 cylinders of the as-received cp Ti used as a reference were embedded in resin and studied metallographically after grinding, polishing, and chemical etching. These surfaces were used for determination of the Vickers hardness (VHN) as well. Eight specimens from each group were fractured in the tensile mode, and the 0.2% yield strength, fracture stress, and percentage elongation were calculated. Porosity was analyzed with 2 way ANOVA and the Newman-Keuls multiple range test. VHN measurements and tensile properties for specimen groups were compared with 1-way ANOVA and the Newman Keuls multiple range test (95% significance level). RESULTS: The porosity levels per group were (%): A = 5.50 +/- 4.34, B = 0.77 +/- 1.27, C = 2.44 +/- 3.68, D = 0.06 +/- 0.12, E-H = 0. Two-way ANOVA showed that there was no detectable interaction (P<.05) between gas type and applied pressure. Metallographic examination revealed no differences in microstructure among the groups studied. A finer grain size was observed in all cast groups compared with the original cp Ti. The VHN of the as-received cp Ti was significantly greater than all the cast groups tested. Groups cast under He showed the highest VHN, yield strength, and fracture stress. No significant differences were found in percentage elongation values among the groups. CONCLUSION: Porosity and mechanical properties of cp Ti castings are dependent on the gas type and pressure, whereas the microstructure remains unaffected. PMID- 11105015 TI - Immediate lingual flange extension. PMID- 11105016 TI - Adaptation of a relocatable head frame for stereotactic radiotherapy. AB - An important role of the maxillofacial prosthodontist is to support the radiotherapist in the administration of therapy. For example, a device can be made that properly and repeatedly positions the patient for each radiation treatment. This article describes a procedure for adapting a relocatable head frame to be used during stereotactic radiotherapy, a treatment modality for malignant intracranial tumors. PMID- 11105017 TI - Automated segmentation and measurement of global white matter lesion volume in patients with multiple sclerosis. AB - A fully automated magnetic resonance (MR) segmentation method for identification and volume measurement of demyelinated white matter has been developed. Spin-echo MR brain scans were performed in 38 patients with multiple sclerosis (MS) and in 46 healthy subjects. Segmentation of normal tissues and white matter lesions (WML) was obtained, based on their relaxation rates and proton density maps. For WML identification, additional criteria included three-dimensional (3D) lesion shape and surrounding tissue composition. Segmented images were generated, and normal brain tissues and WML volumes were obtained. Sensitivity, specificity, and reproducibility of the method were calculated, using the WML identified by two neuroradiologists as the gold standard. The average volume of "abnormal" white matter in normal subjects (false positive) was 0.11 ml (range 0-0.59 ml). In MS patients the average WML volume was 31.0 ml (range 1.1-132.5 ml), with a sensitivity of 87.3%. In the reproducibility study, the mean SD of WML volumes was 2.9 ml. The procedure appears suitable for monitoring disease changes over time. J. Magn. Reson. Imaging 2000;12:799-807. PMID- 11105018 TI - MR imaging of pituitary morphology in idiopathic intracranial hypertension. AB - The aim of this study was to investigate the morphologic changes of the pituitary gland in patients with the clinical diagnosis of idiopathic intracranial hypertension (IIH). Qualitative and quantitative analyses of pituitary morphology were performed in normal subjects (n = 23), patients with the clinical diagnosis of IIH (n = 40), and patients with acute increased intracranial pressure (AICP; n = 37) caused by acute head trauma. The loss of pituitary height (concavity) on the sagittal T1-weighted image was classified into five categories: I = normal, II = superior concavity that was mild (<(1/3) the height of the sella), III = moderate (between (1/3) and (2/3) concavity of height of sella), IV = severe (>(2/3) concavity of height of sella), and V = empty sella. The area ratio of pituitary gland to sella turcica measured in the midsagittal plane was quantified. Clinical records were retrospectively reviewed to correlate with magnetic resonance (MR) findings. Using moderate concavity (>(1/3)) as the minimum criterion for abnormality, IIH patients had an 85% incidence of morphologic changes with 80% sensitivity and 92% specificity. Empty sella (almost complete concavity of the sella) was found in only 2.5% of patients with IIH. Quantitative analysis of the pituitary gland/sella turcica area ratio showed a significant decrease in patients with IIH (P < 0.0001) but no significant difference between the normal subjects and AICP patients. A posterior deviation of the pituitary stalk was seen in 43% of patients. No enlargement of the ventricles or sulcal effacement was seen in IIH patients. Routine brain MR examination of patients with IIH frequently shows morphologic changes of the pituitary gland ranging from various degrees of concavity to (rarely) the extreme case of an empty sella. The etiology is unknown and may be related to the severity and duration of elevated CSF pressure. Such findings may be useful to facilitate the diagnosis of IIH, particularly in patients with equivocal clinical findings or when IIH is not suspected. J. Magn. Reson. Imaging 2000;12:808-813. PMID- 11105019 TI - Comparison of three-dimensional fast spin echo and gradient echo sequences for high-resolution temporal bone imaging. AB - T2-weighted high-resolution gradient and fast spin echo sequences are widely used as an alternative or adjunct to contrast-enhanced T1-weighted temporal bone imaging. However, to date no systematic comparison has been presented. The purpose of this work is to identify optimal acquisition parameters and to compare volume gradient and fast spin echo techniques. Signal intensities and scan efficiency were computed for gradient echo segment-interleaved motion-compensated acquisition into steady state (SIMCAST), standard fast spin echo (FSE), and fast recovery fast spin echo (FR-FSE). Computations were compared with inner ear images acquired with cubic voxel sizes of 0.35-0.40 mm(3)in 5-8 minutes. Given otherwise identical conditions, the FR-FSE sequence produces images with improved SNR in shorter scan times than standard FSE. For FR-FSE, the scan efficiency is optimal for specific pairs of TR and echo train length, eg, 400 ms/8, 735 ms/16, and 2,050 ms/48. FR-FSE images with large TR and echo trains, while achieving better SNR, are severely compromised by blurring. Imaging with echo train lengths of 16-24 and TR of 800-1,200 ms is a good compromise, and FR-FSE signal-to-noise ratio (SNR) and scan efficiency become comparable to SIMCAST. In vivo image quality is excellent with both FR-FSE and SIMCAST, but SIMCAST images have slightly higher SNR and are significantly more crisp. J. Magn. Reson. Imaging 2000;12:814-825. PMID- 11105020 TI - Tumor volume measurements of acoustic neuromas with three-dimensional constructive interference in steady state and conventional spin-echo MR imaging. AB - The purpose was to compare three-dimensional (3D) constructive interference in steady state (CISS) and conventional spin-echo (SE) MR imaging in tumor volume measurements of acoustic neuromas. Twenty-two patients with acoustic neuromas were examined using high-resolution 3D-CISS and SE imaging at a 1.5-T system. Tumor volume determined by SE imaging with the ellipsoid formula was overestimated by 692 mm(3)(35%) on average as compared with that at 3D-CISS with the voxel-count method (the reference standard). Intra- and interobserver variations in SE imaging were poor as compared with 3D-CISS imaging. However, tumor volume results with SE imaging showed a high correlation with those using 3D-CISS imaging (P <. 0001). On the basis of diameters shown on SE images, the tumor volume could be assessed using the following equation (P <.0001): (Tumor volume) = -26.407 + 0.387 x (maximum diameter along the pyramid) x(maximum diameter perpendicular to the pyramid) x (maximum height). J. Magn. Reson. Imaging 2000;12:826-832. PMID- 11105021 TI - Random-grid stereologic volumetry of MR head scans. AB - Point count stereology is a useful tool in obtaining volumetric measures of objects in three-dimensional (3D) images when the segmentation of objects is not feasible. Presently, fixed-grid 3D stereology is being used where a 3D parallelepiped grid is randomly placed for sampling the image space in order to generate test points. Although this is a popular technique, the use of a fixed grid introduces errors in the final estimate in practice and makes the technique inefficient. Random-grid 3D stereology is introduced to improve the efficiency of the volume estimates in stereology. In this manuscript, we prove random-grid stereology as a more consistent technique than fixed-grid stereology and use it for volumetry of the brain and ventricles in magnetic resonance (MR) head scans. We demonstrate superior efficiency and accuracy of random-grid stereology with experiments. Also, the effects of grid sizes, the optimal directions of sectioning the object for volume estimates of the brain and ventricles, and the reliability of the technique are investigated. J. Magn. Reson. Imaging 2000;12:833-841. PMID- 11105022 TI - Multispectral analysis of the temporal evolution of cerebral ischemia in the rat brain. AB - A major difficulty in staging and predicting ischemic brain injury by magnetic resonance (MR) imaging is the time-varying nature of the MR parameters within the ischemic lesion. A new multispectral (MS) approach is described to characterize cerebral ischemia in a time-independent fashion. MS analysis of five MR parameters (mean diffusivity, diffusion anisotropy, T2, proton density, and perfusion) was employed to characterize the progression of ischemic lesion in the rat brain following 60 minutes of transient focal ischemia. k-Means (KM) and fuzzy c-means (FCM) classification methods were employed to define the acute and subacute ischemic lesion. KM produced an estimate of lesion volume that was highly correlated with postmortem infarct volume, independent of the age of the lesion. Overall classification rates for KM exceeded FCM at acute and subacute time points as follows: KM, 90.5%, 94.4%, and 95. 9%; FCM, 82.4%, 90.6%, and 82.6% (for 45 minutes, 180 minutes, and 24-120 hours post MCAO groups). MS analysis also offers a formal method of combining diffusion and perfusion parameters to provide an estimate of the ischemic penumbra (KM classification rate = 70.3%). J. Magn. Reson. Imaging 2000;12:842-858. PMID- 11105023 TI - Changes in cerebral metabolism are detected prior to perfusion changes in early HIV-CMC: A coregistered (1)H MRS and SPECT study. AB - Human immunodeficiency virus-cognitive motor complex (HIV-CMC), a common complication of the acquired immunodeficiency syndrome (AIDS), is characterized by progressive cognitive impairment and motor dysfunction. Functional imaging methods, such as single-photon emission computed tomography (SPECT) and proton magnetic resonance spectroscopy ((1)H-MRS), have been applied to assess the severity of brain injury. However, it is unclear which of these two methods is more sensitive in detecting brain abnormalities in patients with early HIV-CMC. Twenty-four HIV-CMC patients were compared with 34 healthy subjects; each had quantitative SPECT ((133)Xenon-calibrated (99m)Tc-HMPAO) and quantitative (1)H MRS. Both modalities were co-registered in order to assess regional cerebral blood flow (rCBF) and metabolite concentrations within the same voxel of interest in four brain regions (midfrontal and midparietal gray matter, temporoparietal white matter, and basal ganglia). On SPECT, only the temporoparietal white matter showed a trend for decreased rCBF in HIV-CMC patients (-13%, P = 0.06). On MRS, HIV-CMC patients showed significantly reduced creatine concentration in the basal ganglia (-8%, P = 0.008), as well as increased myoinositol concentrations in the basal ganglia (+25%, P = 0.01) and the temporoparietal white matter (+18%, P = 0.08). There was no significant correlation between SPECT and MRS variables in the patients in any region. (1)H MRS showed abnormal neurochemistry in the basal ganglia, whereas rCBF on SPECT was normal in the same region. This finding suggests that metabolite concentrations on (1)H MRS are better surrogate markers than rCBF measurements with SPECT for the evaluation of brain injury in early HIV CMC. J. Magn. Reson. Imaging 2000;12:859-865. PMID- 11105024 TI - Evaluation of nonperfused myocardial ischemia with MRI and an intravascular USPIO contrast agent in an ex vivo pig model. AB - The ultrasmall superparamagnetic iron oxide (USPIO) preparation NC100150 Injection (Clariscan; Nycomed Imaging, Oslo, Norway) was tested for its ability to delineate nonperfused myocardium under steady-state conditions. An experimental animal model of focal myocardial ischemia induced by ligation of the distal part of the left anterior descending artery was used. The contrast agent was administered in four doses: 0, 4, 8, and 12 mg Fe/kg body weight. Magnetic resonance examination ex vivo, including T1-, T2-, and T2*-weighted sequences, was performed. Nonperfused myocardium was determined by fluorescein. The best delineation of nonperfused myocardium was found with a T1-weighted inversion recovery/turbo spin-echo sequence and doses of 4 and 8 mg Fe/kg body weight, where 95% of the volume was discernible at the dose of 4 mg Fe/kg body weight. The results suggest that steady-state imaging by T1-weighted sequence with the use of NC100150 Injection to delineate nonperfused myocardium is feasible. J. Magn. Reson. Imaging 2000;12:866-872. PMID- 11105025 TI - Investigating intrinsic myocardial mechanics: the role of MR tagging, velocity phase mapping, and diffusion imaging. AB - Assessment of myocardial mechanics is an integral part of understanding and predicting heart disease. This review covers the two most common magnetic resonance (MR) methods used to measure myocardial motion: myocardial tagging and myocardial velocity mapping. Myocardial tagging has been well established in clinical research, despite its time-consuming postprocessing procedure. Myocardial velocity mapping uses the phase shifts of the spins to encode the velocity into the MR signal. This means that once the myocardial contours have been segmented, the data can be automatically processed to obtain quantitative measurements. Diffusion MR also has found applications in cardiac imaging, with preliminary results of myocardial fiber architecture being obtained recently. These three different MR techniques have provided valuable insights into the assessment of intrinsic cardiac mechanics. J. Magn. Reson. Imaging 2000;12:873 883. PMID- 11105026 TI - MR angiography of the vascular tree from the aorta to the foot: combining two dimensional time-of-flight and three-dimensional contrast-enhanced imaging. AB - A composite approach for magnetic resonance (MR) angiography of the lower extremities is described. Thirty patients were studied with this approach, which combined a two-dimensional (2D) time-of-flight (TOF) technique with a 3D contrast enhanced technique. A head/neck coil was selected for imaging mid-foot to upper calf, and the body coil was used for the remainder of the peripheral vascular tree. Acquired data were transferred to a workstation for postprocessing. The final maximum intensity projection images, which display the entire vascular anatomy from aortic bifurcation to foot, were created using a 1024 x 1024 matrix. Very small arteries can be differentiated in critical regions like the calf and foot. Compared with TOF-2D alone, the scan time was reduced. This method offers another option for MR angiography of the lower extremities. J. Magn. Reson. Imaging 2000;12:884-889. PMID- 11105027 TI - Blood pool MR contrast agents for cardiovascular imaging. AB - Currently available magnetic resonance (MR) contrast agents are not confined to the intravascular space because of their small molecular size. These agents produce peak vascular enhancement for only a short period. Conversely, blood pool agents have longer intravascular residence time and higher relaxivity. Therefore these agents provide MR angiography with flexibility, versatility, and accuracy. With blood pool agents, the timing of contrast injection becomes less significant because the optimal imaging window is in tens of minutes rather than seconds. In addition, larger anatomic regions can be imaged optimally. Preliminary evidence appears to support the notion that blood pool agents may play a diagnostic role in coronary, peripheral, and pulmonary angiography. Besides their ability to increase vascular contrast, blood pool agents provide physiologic information, including rate of entry, rate of accumulation, and rate of elimination. MR imaging with blood pool agents also have proven to be of significant value in the assessments of myocardial perfusion and microvascular permeability. In anticipation of broad clinical use, blood pool agents are currently being evaluated in human trails. Examples include gadolinium-chelate that binds in vivo to albumin to form blood pool agents and ultrasmall superparamagnetic iron oxide particles. This review discusses the applications of MR blood pool agents in the cardiovascular system. J. Magn. Reson. Imaging 2000;12:890-898. PMID- 11105028 TI - MR imaging of mediastinal lymph nodes: evaluation using a superparamagnetic contrast agent. AB - The purpose of this study was to determine whether intravenous injection of a magnetic resonance (MR) contrast agent, ultrasmall superparamagnetic iron oxide (ferumoxtran-10), can be useful in characterizing lymph nodes in patients with lung cancer. Twelve patients with known or suspected lung cancer were studied. Pre- and postcontrast injection of ferumoxtran-10 MR scans of the chest were obtained. Analysis of the signal intensities and bronchoscopic fine needle aspiration of a single node were performed in each patient. Six of 12 patients had a final diagnosis of lung cancer. T1-weighted images were best for localizing mediastinal lymph nodes. Signal intensity changes before and after contrast were best visualized on T2-weighted and gradient-echo images. All four patients with lung cancer who had nodes positive for malignancy at biopsy had no change in signal intensity of the nodes on T2 images. The signal intensity decreased in the remaining two patients, and the nodes were benign. Of the eight patients with benign disease, five had no change in signal intensity of the nodes. Therefore the sensitivity for tumor involvement of the nodes is 100%, but the specificity is only 37.5%. Ferumoxtran-10 is a contrast agent that can alter the signal intensity of lymph nodes. Lack of signal change may be due to malignant or inflammatory change. Studies in a larger population of lung cancer patients may help to characterize the utility of this agent further. J. Magn. Reson. Imaging 2000;12:899-904. PMID- 11105029 TI - New generation of monomer-stabilized very small superparamagnetic iron oxide particles (VSOP) as contrast medium for MR angiography: preclinical results in rats and rabbits. AB - The purpose of this study was to evaluate the signal enhancement characteristics of very small superparamagnetic iron oxide particles (VSOP)-C63, a new monomer coated, iron oxide-based magnetic resonance (MR) blood pool contrast medium with a very small particle size and optimized physical properties. Equilibrium MR angiography (MRA) of rats (thoracic and abdominal vessels) was performed at 1.5 T with a three-dimensional gradient-recalled echo (3D GRE) technique (TR/TE 6.6/2.3 msec, flip angle 25 degrees ) before and after (every 3-5 minutes up to 50 minutes) i.v. injection of VSOP-C63 [dosages: 15, 30, 45, 60, 75, and 90 micromol Fe/kg; diameter: 8 nm; relaxivities at 0.47 T: R1 = 30 l/(mmol * s); R2 = 39 l/(mmol * s)]. First-pass MRA images (3D-GRE, TR/TE 4.5/1.7 msec, flip angle 25 degrees ) were obtained with 45 micromol Fe/kg VSOP-C63 in comparison with 0.2 mmol Gd/kg of gadolinium diethylene triamine pentaacetic acid (Gd DTPA; before and every 5 seconds p.i.). MRA (3D GRE, TR/TE 4.5/1.7 msec, flip angle 25 degrees) of coronary vessels in rabbits was performed after i.v. injection of 45 micromol Fe/kg of VSOP-C63. In rats maximal S/N ratio in thoracic and abdominal arteries directly after i.v. injection of VSOP-C63 was 25 +/- 1, 43 +/- 2, 49 +/- 4, 57 +/- 3, 64 +/- 3, and 63 +/- 3 for the different dosages. Blood half-life was dose dependent (15 +/- 2, 20 +/- 3, 29 +/- 6, 37 +/- 5, 61 +/- 16, and 86 +/- 21 minutes). At a dose of 30 micromol Fe/kg even small intrarenal arteries were sharply delineated. First-pass MRA showed no significant difference in the S/N ratio between Gd-DTPA (71.5 +/- 11.5) and VSOP-C63 (65.1 +/- 18. 3). The proximal segments of the coronary arteries in rabbits were clearly depicted at a dose of 45 micromol Fe/kg. The monomer-coated, iron oxide-based contrast medium VSOP-C63 exhibits favorable properties as a blood pool agent for both equilibrium and first-pass MRA. J. Magn. Reson. Imaging 2000;12:905-911. PMID- 11105030 TI - Contrast-enhanced breath-hold three-dimensional magnetic resonance angiography in the evaluation of renal arteries: optimization of technique and pitfalls. AB - The authors describe the optimization of a contrast-enhanced, breath-held, three dimensional magnetic resonance angiography (CE-BH-3DMRA) technique in the assessment of the renal arteries and compare its utility with conventional x-ray angiography (XRA). Signal optimization using specific pulse sequence parameters was based on the patient's circulatory conditions, injection rate, and pulse sequence timing. Fifty-one patients (27 M, 24 F; mean age 69.7 years) were evaluated with CE-BH-3DMRA and XRA. All patients had an MR angiogram 3 months either before or after XRA. A test bolus study was performed for accurate assessment of transit time in each patient. A total of 51 patients (115 vessels) were studied in which the sensitivity and specificity for all renal artery stenoses including the proximal and mid-renal arterial segments were 96% and 92%, respectively. In-stent stenosis could only be diagnosed by quantifying flow beyond the stent using an additional triggered phase contrast cine pulse sequence. A total of 11 accessory renal arteries were correctly identified. In addition, fibromuscular dysplasia in two patients and stents in three patients were correctly identified on MRA. J. Magn. Reson. Imaging 2000;12:912-923. PMID- 11105031 TI - Motion of the proximal renal artery during the cardiac cycle. AB - In 48 hypertensive patients, the motion of the proximal renal artery during the cardiac cycle was quantified using two-dimensional quantitative flow (QF) measurements and automatic contour detection. Substantial translational motion was observed with an amplitude ranging from 1 to 4 mm. Since motion effectively reduces spatial resolution, the use of motion suppression techniques should be strongly considered for renal MR angiography. J. Magn. Reson. Imaging 2000;12:924 928. PMID- 11105032 TI - Dynamic in vivo oxymetry using overhauser enhanced MR imaging. AB - A noninvasive method for in vivo measurement of the oxygen concentration has been developed. By introducing a novel contrast medium (CM) based on a single electron substance, it is possible to enhance the proton signal through the Overhauser effect. A low-field magnetic resonance scanner is used to image the proton nuclei of the object. The electron spin transition of the CM is saturated using rf irradiation. As a consequence, the nuclear polarization becomes enhanced through dipole-dipole interaction. The signal enhancement is a function of rf power and of the EPR line width of the substance, which is influenced by the oxygen concentration. The maximum in vivo enhancement has been measured to 60. Image data, generated with different scanning parameters, is used in a postprocessing method to generate images showing pO(2) and the contrast medium concentration, respectively. The mathematical foundation of the postprocessing algorithm is outlined. The results from phantom experiments and animal experiments, in which the oxygen content of the inspired gas was varied, are presented. The potential for human imaging is discussed. J. Magn. Reson. Imaging 2000;12:929-938. PMID- 11105033 TI - Measuring flow reattachment lengths downstream of a stenosis using MRI. AB - Flow reattachment lengths (l(r)) are measured downstream of an abrupt axisymmetric 75% stenosis, located inside a cylindrical channel, for steady flow using ultra-fast magnetic resonance imaging (MRI). The MRI results are compared with those from other similar (non-MRI) studies. The MRI data confirm the existence of three flow reattachment regimes (laminar, fully turbulent, and transition) related to the flow Reynolds number (Re) measured inside the stenosis. Based on the MRI experiments, the laminar regime occurs at a stenotic Reynolds number below 250 with a slope (l(r)/Re) of 0.086. The fully developed turbulence occurs at a stenotic Reynolds number above 3600 with a minimum observed reattachment length of 5 step heights. The transition regime (occurring between the laminar and fully turbulent regimes) is characterized by a reattachment length plateau and then a drop with Re(-1.1). J. Magn. Reson. Imaging 2000;12:939-948. PMID- 11105034 TI - Three cases of spinal dural AVF: evaluation with first-pass, gadolinium-enhanced, three-dimensional MR angiography. AB - A first-pass, contrast-enhanced three-dimensional (3D) magnetic resonance (MR) angiography with short acquisition time was performed for three consecutive cases with suspected spinal arteriovenous fistula (AVF). This MR technique demonstrated the feeding arteries and draining veins of the spinal dural AVF, and was very useful for the definite diagnosis of the lesions as a disease with arteriovenous shunt. With this MR technique, we could localize the feeding artery and shunt before conventional angiography. J. Magn. Reson. Imaging 2000;12:949-952. PMID- 11105035 TI - Utilization of "used" vials: cost-effective technique for MR arthrography. AB - Because full vials of commercially available MR arthrographic contrast are expensive, we hypothesized that the small residual contrast in a "used" vial would be adequate for MR arthrography. After sterility testing and quantity analysis of the residual contrast in 28 vials, this method was successfully used in 10 patients. J. Magn. Reson. Imaging 2000;12:953-955. PMID- 11105037 TI - Heavily T1-weighted images without respiratory artifacts: partial angle inversion recovery fast spin-echo imaging (PAIR-FSE). AB - We obtained T1-weighted images in the abdominal region using the partial angle inversion recovery fast spin echo (PAIR-FSE) with the respiratory triggering (RT) method and compared the image quality with that of conventional SE (TR/TE 500/10 msec) with the partial angle inversion recovery (PEAR) method. The signal difference to noise ratio of the PAIR-FSE was 1.6 times higher (6.94 +/- 3.08) than that of SE (4.30 +/- 1.88). Respiratory motion-induced ghost artifacts were reduced by half in PAIR-FSE with RT (1.01 +/- 0.47) in comparison with SE with PEAR (2.24 +/- 0.70). J. Magn. Reson. Imaging 2000;12:960-964. PMID- 11105036 TI - A B(0) shift correction method based on edge RMS reduction for EPI fMRI. AB - Shifting of echoplanar images (EPI) in the phase-encoding direction during functional magnetic resonance imaging (fMRI) experiments can be observed due to B(0) drift. These shifts can cause artifacts in functional activation maps that can be corrected using a navigator echo (NE) technique, but the NE correction requires pulse sequence modifications not available on many clinical systems. A fast, postprocessing correction method based on edge root-mean-square error reduction (ERMSR) is introduced and shown to provide an equivalent correction. J. Magn. Reson. Imaging 2000;12:956-959. PMID- 11105038 TI - Dynamic image interpretation of MRI of the breast. AB - Dynamic breast MRI provides information on both lesion cross-sectional morphology and functional lesion features such as vascularity/perfusion and vessel permeability. This review gives an overview of the historical background of dynamic contrast-enhanced breast MRI. It explains the technique's pathophysiological basis, describes the various technical approaches that have been pursued and the corresponding interpretation guidelines that have been proposed (including their respective diagnostic accuracies), and presents established and evolving clinical applications of the "dynamic approach" to breast MRI. J. Magn. Reson. Imaging 2000;12:965-974. PMID- 11105039 TI - Magnetic resonance imaging of breast cancer: clinical indications and breast MRI reporting system. AB - Magnetic resonance imaging (MRI) is well suited to the investigation of breast cancer by virtue of its noninvasive nature and its multiplanar imaging abilities. MRI investigations showed high sensitivity but modest specificity for breast cancer detection and diagnosis. Most early studies tested the ability of MRI to evaluate and diagnose findings in the breast discovered by other imaging tests or by breast physical examination (1-4). When it was discovered that MRI identified small breast cancers undetected by mammography or breast ultrasound, MRI was used to estimate breast cancer extent in known cancer cases for surgical planning (5,6). These investigations led to the use of MRI in a multitude of breast imaging applications, raising further questions about the use of MRI in everyday practice: What are the indications for breast MRI in general practice? What is its role in light of other imaging tests? What are its benefits and limitations in each setting? How do I report these studies? The purpose of this article is to review the clinical background regarding indications for the use of MRI and relevant cases in which MRI can impact patient management in breast disease, and to describe new developments in reporting breast MRI studies. J. Magn. Reson. Imaging 2000;12:975-983. PMID- 11105040 TI - Bilateral open breast coil and compatible intervention device. AB - Dynamic contrast-enhanced breast MRI is an extremely sensitive method for breast lesion detection. For MR-only detected lesions it is essential that needle biopsy or localization prior to surgery is carried out under MR guidance. This work describes a bilateral open breast coil and prototype intervention device, which may be used in these situations. Results demonstrate that the open coil provides images superior to those obtained with a conventional closed breast coil. Initial phantom tests with the intervention device indicate a potential for this system to be used in the MR-guided localization of breast lesions. J. Magn. Reson. Imaging 2000;12:984-990. PMID- 11105041 TI - Simultaneous MRI measurement of blood flow, blood volume, and capillary permeability in mammary tumors using two different contrast agents. AB - A technique for the simultaneous measurement of three vascular parameters: blood flow (Frho), blood volume (v(b)), and the capillary permeability-surface area product (PSrho) in breast tumors using dynamic contrast-enhanced magnetic resonance imaging (MRI) is presented. Features of the technique include measurement of precontrast tumor T(1), rapid temporal sampling, measurement of the arterial input function, and use of a distributed parameter tracer kinetic model. Parameter measurements are compared that were determined using two contrast agents of different molecular weights, gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA; 0.6 kDa) and Gadomer-17 (17 kDa), in 18 spontaneous canine mammary tumors. Measurements of Frho and v(b) corresponded well with literature values, and the mean PSrho measured using Gd-DTPA was a factor of 15 higher than that measured using Gadomer-17. J. Magn. Reson. Imaging 2000;12:991 1003. PMID- 11105042 TI - Semiquantitative assessment of uterine perfusion using first pass dynamic contrast-enhanced MR imaging for patients treated with uterine fibroid embolization. AB - The feasibility of using first pass dynamic contrast-enhanced MRI to monitor semiquantitatively the perfusion changes of the uterus after uterine arterial embolization is demonstrated. Ten women, who underwent uterine arterial embolization for fibroid treatment, were included in this study. To derive a perfusion index, an additional axial slice through the abdominal aorta was obtained simultaneously when acquiring MR perfusion data. This technique may prove valuable in monitoring the outcome of uterine arterial embolization and documentation of preserved uterine perfusion after this procedure. J. Magn. Reson. Imaging 2000;12:1004-1008. PMID- 11105043 TI - Magnetic resonance hysterography and hysterosalpingography using hyperpolarized (3)He: demonstration of feasibility in an animal model. AB - Assessment of the uterine cavity and patency of the fallopian tubes remains a difficult goal with magnetic resonance imaging (MRI). The purpose of this paper is to describe the development of a new magnetic resonance hysterography (MR-HG) and hysterosalpingography (MR-HSG) technique employing hyperpolarized (3)He. Two dimensional (2D) and 3D gradient-echo imaging sequences were developed and optimized using a phantom. An optimized sequence was then applied in swine cadavers. J. Magn. Reson. Imaging 2000;12:1009-1013. PMID- 11105044 TI - Diffusion-weighted EPI of cystic ovarian lesions: evaluation of cystic contents using apparent diffusion coefficients. AB - Diffusion-weighted echoplanar imaging (EPI) was performed to evaluate the contents of cystic ovarian lesions [33 endometrial cysts, 16 ovarian cysts, 5 serous cystadenomas, 6 mucinous cystadenomas, 13 malignant cystic ovarian tumors, and 3 benign cystic lesions mimicking malignant cystic ovarian tumor (BMMs)]. Apparent diffusion coefficient (ADC) values were calculated for the cystic contents. Many lesions showed unrealistically high ADC greater than water at body temperature (0.00324 mm(2)/sec), especially in lesions more than 12 cm in diameter. The higher ADC values were attributed to the sloshing effect: intermittent compression of large ovarian lesions by abdominal breathing before the breath-hold scan. When limited to lesions with ADC less than 0. 00324 mm(2)/sec, endometrial cysts and malignant cystic ovarian tumors showed lower ADC values than ovarian cysts and serous cystadenomas (P < 0.03). When further limited to lesions less than 12 cm in axial diameter, an additional ADC difference between malignant cystic ovarian BMMs was demonstrated (P < 0.03). Diffusion-weighted EPI showed the possibility that calculated ADC values might be useful in evaluating small to medium cystic ovarian lesions. J. Magn. Reson. Imaging 2000;12:1014-1019. PMID- 11105045 TI - Magnetic resonance imaging of the primary site in stage I cervical carcinoma: A comparison of endovaginal coil with external phased array coil techniques at 0.5T. AB - OBJECTIVE: To compare endovaginal with pelvic phased array coil magnetic resonance imaging (MRI) in detection of Stage I cervical carcinoma by correlating the findings with histopathology. PATIENTS AND METHODS: Forty consecutive patients with Stage I cervical carcinoma confirmed histologically were studied using an endovaginal coil alone immediately followed by a pelvic phased array coil. T1-W transverse and T2-W FSE sagittal images made with each coil were analyzed independently by two radiologists noting the presence and size of a mass within the cervix and any parametrial extension or involvement of adjacent organs. Tumor volumes were measured using the electronic calliper to compute tumor area on each slice and multiplying by the slice thickness. Thirty patients underwent radical hysterectomy, one a trachylectomy, one simple hysterectomy and four extended cone biopsies. Four patients had radiotherapy to the primary tumor. Following surgery, histopathologic findings were recorded and tumor volumes measured. RESULTS: Tumor volumes ranged from 0-106 cm(3)(median 1.4 cm(3), mean 9 +/- 19.4 cm(3)). Thirty-six patients had correlation of the primary site with the surgical specimen. Agreement between observers was excellent for both endovaginal (k = 0.90) and pelvic phased array (k = 0.96) techniques. Combined sensitivity and specificity for both observers of endovaginal MR imaging for detection of tumor was 96% and 70%, respectively; for pelvic phased array imaging sensitivity was substantially less at 54%. Specificity was higher at 83.7%, probably because small abnormalities were seldom visible. In patients treated surgically, early parametrial involvement in four women on endovaginal MRI was confirmed histologically in two. Pelvic phased array imaging showed early parametrial involvement in four women and was confirmed in one. CONCLUSION: Endovaginal MRI adds substantially to information from pelvic phased array images in the preoperative assessment of patients with early cervical cancer. J. Magn. Reson. Imaging 2000;12:1020-1026. PMID- 11105047 TI - Gadolinium-enhanced MR angiography in the evaluation of congenital cardiovascular disease pre- and postoperative states in infants and children. AB - The purpose of this study was to evaluate the utility of dynamic contrast magnetic resonance (MR) angiography under sedation for assessing congenital cardiovascular disease in infants and small children before and after cardiac surgery. In 38 patients with cardiovascular abnormalities, thoracic vasculatures were evaluated in either the preoperative (group 1; 23 patients, median age 9 months old) or the postoperative (group 2; 15 patients median age 1. 3 years old) state using gadolinium-enhanced dynamic MR angiography. Acquired data were processed with a multiprojection volume reconstruction. Image quality (grades 1 5, undiagnostic to excellent), presence or absence of the extracardiac vasculature, its pathology (group 1), and the patency of the postsurgical shunt or reconstructed vasculature (group 2) were evaluated. All images were of diagnostic image quality (mean grade 4.4-3.2). Recognition of the thoracic vasculature was correctly made in all cases (38/38, 100%). In group 1, anomalies and pathologic changes were correctly identified in 22 of 23 cases (95.7%). In one patient with dextrocardia and the cardiac type of total anomalous pulmonary venous return, the abnormality was not recognized. In group 2, the patency of the shunt flow after Glenn (n = 6), modified Fontan (n = 4), Rastelli (n = 1), and Blalock-Taussig (n = 3) operations was well demonstrated in each case. The reconstructed pulmonary artery (n = 1), veins (n = 2), and aorta (n = 1) were correctly visualized. In one case, stenosis of the reconstructed left pulmonary vein was missed by MR angiography. In 14 of 15 cases in group 2 (93.3%), MR angiography correctly gave information on the operated areas. In conclusion, dynamic contrast MR angiography under sedation is useful for evaluation of the thoracic vasculature of infants and small children in the pre- and postoperative states. J. Magn. Reson. Imaging 2000;12:1034-1042. PMID- 11105046 TI - Pixel analysis of MR perfusion imaging in predicting radiation therapy outcome in cervical cancer. AB - The purpose of this study was to assess heterogeneity of tumor microcirculation determined by dynamic contrast-enhanced magnetic resonance (MR) imaging and its prognostic value for tumor radiosensitivity and long-term tumor control using pixel-by-pixel analysis of the dynamic contrast enhancement. Sixteen patients with advanced cervical cancer were examined with dynamic contrast-enhanced MR imaging at the time of radiation therapy. Pixel-by-pixel statistical analysis of the ratio of post- to precontrast relative signal intensity (RSI) values in the tumor region was performed to generate pixel RSI distributions of dynamic enhancement patterns. Histogram parameters were correlated with subsequent tumor control based on long-term cancer follow-up (median follow-up 4.6 years; range 3.8-5.2 years). The RSI distribution histograms showed a wide spectrum of heterogeneity in the dynamic enhancement pattern within the tumor. The quantity of low-enhancement regions (10th percentile RSI < 2.5) significantly predicted subsequent tumor recurrence (88% vs. 0%, P = 0.0004). Discriminant analysis based on both 10th percentile RSI and pixel number (reflective of tumor size) further improved the prediction rate (100% correct prediction of subsequent tumor control vs. recurrence). These preliminary results suggest that quantification of the extent of poor vascularity regions within the tumor may be useful in predicting long-term tumor control and treatment outcome in cervical cancer. J. Magn. Reson. Imaging 2000;12:1027-1033. PMID- 11105048 TI - MRI-Advance in patient care PMID- 11105049 TI - Cre recombinase expression in cerebellar Purkinje cells. AB - The cerebellar cortex and its sole output, the Purkinje cell, have been implicated in motor coordination, learning and cognitive functions. Therefore, the ability to generate Purkinje cell-specific mutations in physiologically relevant genes is of particular neurobiological interest. A suitable approach is the Cre/loxP strategy that allows temporally and spatially controlled gene inactivation. Here, we present the characterization of transgenic mouse strains expressing Cre recombinase controlled by the L7/pcp-2 gene. Endogenous L7/pcp-2 protein is expressed exclusively in Purkinje cells and retinal bipolar neurones. Recombination was detected by beta-galactosidase histochemistry in tissues from crosses of the L7/pcp-2:Cre transgenic lines with two different indicator strains, GtROSA26 and ACZL. Purkinje cells in all folia of the cerebellum displayed intense beta-galactosidase staining, whereas only few blue cells were observed in the retina and other parts of the CNS. Thus, these transgenic lines are potentially of great importance for genetic manipulations in cerebellar Purkinje cells. PMID- 11105050 TI - The Fugu rubripes tyrosinase gene promoter targets transgene expression to pigment cells in the mouse. AB - The regulation of the mouse tyrosinase gene expression is controlled by a highly conserved element at -100 bp, the M-box, and an enhancer at -12 kb. In most vertebrates, the length of intergenic sequences makes it difficult to analyze the whole gene and the complete regulatory region. We took advantage of the compact Fugu genome to identify regulatory regions involved in pigment cell-specific expression. We isolated the Fugu tyrosinase gene, and identified putative cis acting regulatory elements within the promoter. We then asked whether the Fugu promoter sequence functions in mouse pigment cells. We showed that E11.5 transgenic embryos bearing 6 kb or 3 kb of Fugu tyrosinase 5' sequence fused to the reporter gene lacZ revealed melanoblast and RPE-specific expression. This is the first evidence that the tyrosinase promoter is active at midgestation in melanoblasts, long before the onset of pigmentation. PMID- 11105051 TI - Widespread recombinase expression using FLPeR (flipper) mice. AB - As conditional genetic strategies advance, the need for multiple site-specific recombinase systems has emerged. To meet this need in part, we have targeted the constitutive ROSA26 locus to create a mouse strain with generalized expression of the enhanced version of the site-specific recombinase FLP (FLPe). This strain is designated FLPeR ("flipper"). Using this strain, extensive target gene recombination can be achieved in most tissue types, including cells of the developing germ line. FLPeR mice therefore serve two important functions: as a source of many different FLPe-expressing primary cell lines and as a deleter strain. Moreover, because the FLPeR mouse is a 129-derived strain, a 129 genetic background can be preserved when crossed to most ES cell-derived mice. This enables conditional genetic alterations to be maintained on a standard background, a feature important for obtaining reproducible results and genetically defined controls. PMID- 11105053 TI - Developmental effects of a chimeric ultraspiracle gene derived from Drosophila and Chironomus. AB - The ultraspiracle (usp) gene encodes a nuclear receptor that forms a heterodimer with the ecdysone receptor (EcR) to mediate transcriptional responses to the insect steroid hormone, 20-hydroxyecdysone (20HE). The responses ultimately elicit changes associated with molting and metamorphosis. Although Ultraspiracle (USP) is required at several developmental times, it is unclear whether USP plays stage-specific roles in Drosophila. A chimeric transgene (d/cusp), produced by replacing the ligand-binding domain (LBD) of Drosophila USP with the equivalent domain from another Diptera, Chironomus tentans, was tested for its ability to rescue Drosophila usp mutants from early larval lethality. A single copy of the d/cusp was sufficient to rescue transformants from several lines through larval development but they died suddenly during the late third instar. Additional doses of d/cusp were required to allow survival through the adult stage, but they did not restore a normal prepupal contraction. Thus, the arrest at the onset of metamorphosis apparently is caused by the impaired ability of the chimeric USP to mediate a stage-specific function associated with the LBD. PMID- 11105052 TI - Related function of mouse SOX3, SOX9, and SRY HMG domains assayed by male sex determination. AB - Sox genes encode proteins related to each other, and to the sex determining gene Sry, by the presence of a DNA binding motif known as the HMG domain. Although HMG domains can bind to related DNA sequences, Sox gene products may achieve target gene specificity by binding to preferred target sequences or by interacting with specific partner proteins. To assess their functional similarities, we replaced the HMG box of Sry with the HMG box of Sox3 or Sox9 and tested whether these constructs caused sex reversal in XX mice. Our results indicate that such chimeric transgenes can functionally replace Sry and elicit development of testis cords, male patterns of gene expression, and elaboration of male secondary sexual characteristics. This implies that chimeric SRY proteins with SOX HMG domains can bind to and regulate SRY target genes and that potential SRY partner factor interactions are not disrupted by HMG domain substitutions. genesis 28:111-124, 2000. PMID- 11105054 TI - DNA chips and development: prospects and problems. PMID- 11105055 TI - Function of Rx, but not Pax6, is essential for the formation of retinal progenitor cells in mice. AB - Rx plays a critical role in eye formation. Targeted elimination of Rx results in embryos that do not develop eyes. In this study, we have investigated the expression of Otx2, Six3, and Pax6 in Rx deficient embryos. We find that these genes show normal activation in the anterior neural plate in Rx-/- embryos, but they are not upregulated in the area of the neural plate that would form the primordium of the optic vesicle. In contrast, in homozygous Small eye embryos that lack Pax6 function, Rx shows normal activation in the anterior neural plate and normal upregulation in the optic vesicle/retinal progenitor cells. This suggests that neither Rx expression nor the formation of retinal progenitor cells is dependent on a functional copy of the Pax6 gene, but that Pax6 expression and the formation of the progenitor cells of the optic cup is dependent on a functional copy of the Rx gene. PMID- 11105056 TI - Insectivores await new explorations for developmental biology. PMID- 11105057 TI - Z/EG, a double reporter mouse line that expresses enhanced green fluorescent protein upon Cre-mediated excision. AB - The Cre/loxP system has become an important tool in designing postintegrational switch mechanisms for transgenes in mice. The power and spectrum of application of this system depends on transgenic mouse lines that provide Cre recombinase activity with a defined cell type-, tissue-, or developmental stage-specificity. We have developed a novel mouse line that acts as a Cre reporter. The mice, designated Z/EG (lacZ/EGFP), express lacZ throughout embryonic development and adult stages. Cre excision, however, removes the lacZ gene, which activates expression of the second reporter, enhanced green fluorescent protein. We have found that the double-reporter Z/EG line is able to indicate the occurrence of Cre excision from early embryonic to adult lineages. The advantage of the Z/EG line is that Cre-mediated excision can be monitored in live samples and that live cells with Cre-mediated excision can be isolated using a single-step FACS. It will be a valuable reagent for the increasing number of investigators taking advantage of the powerful tools provided by the Cre/loxP site-specific recombinase system. PMID- 11105058 TI - Mouse embryonic stem (ES) cell lines established from neuronal cell-derived cloned blastocysts. AB - We have established mouse embryonic stem (ES) cell lines from blastocysts derived by transfer of nuclei of fetal neuronal cells. These neuronal cell-derived embryonic cell lines had properties that characterize them as ES cells, including typical cell markers and alkaline phosphatase activity. Moreover, the cells had a normal karyotype and were pluripotent, as they were capable of differentiating into all three germ layers. Although they were derived from neuronal donor nuclei, the cells no longer expressed neuronal markers; however, they were capable of differentiating into cells with neuronal characteristics. These results suggest that the clone-derived cells have fully acquired an ES cell character. Thus, ES cells can be derived from embryos resulting from nuclear transfer, which results in reprogramming of the genetic information and acquisition of pluripotency. ES cells established from somatic cell-derived blastocysts could be useful not only as research tools for studying reprogramming but also as models for cell-based transplantation therapy. PMID- 11105059 TI - A marker assisted selection protocol (MASP) to generate C57BL/6J or 129S6/SvEvTac speed congenic or consomic strains. AB - A marker assisted selection protocol is presented that allows for the generation of congenic or consomic strains derived from a C57BL/6J:129S6/SvEvTac mixed strain background. The protocol uses defined primer pairs to generate amplicons that can be distinguished by non-denaturing agarose electrophoresis. Use of this application should result in substantial savings in time, effort, and cost for investigators in all areas of transgenic mouse research. PMID- 11105060 TI - Cre-mediated recombination in the pituitary gland. AB - Organ-specific expression of a cre recombinase transgene allows for the analysis of gene function in a particular tissue or cell type. Using a 4.6 kb promoter from the mouse glycoprotein hormone alpha-subunit (alphaGSU or Cga) gene, we have generated and characterized a line of transgenic mice that express cre recombinase in the anterior and intermediate lobes of the pituitary gland. Utilizing a cre-responsive reporter transgene, alphaGSU-cre transgene expression was detected in the pituitary primordium and in all five cell types of the adult anterior pituitary. alphaGSU-cre transgene activity was also detected in the cardiac and skeletal muscle. Little or no activity was evident in the gonads, adrenal glands, brain, ventromedial hypothalamus, or kidneys. The alphaGSU-cre transgenic mice characterized here will be a valuable tool for examining gene function in the pituitary gland. PMID- 11105061 TI - Huntington disease: DNA analysis in Brazilian population. AB - Huntington disease (HD) is associated with expansions of a CAG trinucleotide repeat in the HD gene. Accurate measurement of a specific CAG repeat sequence in the HD gene in 92 Brazilian controls without HD, 44 Brazilian subjects with clinical findings suggestive of HD and 40 individuals from 6 putative HD families, showed a range from 7 to 33 repeats in normal subjects and 39 to 88 repeats in affected subjects. A trend between early age at onset of first symptoms and increasing number of repeats was seen. Major increase of repeat size through paternal inheritance than through maternal inheritance was observed. Data generated from this study may have significant implications for the etiology, knowledge of the incidence, diagnosis, prognosis, genetic counseling and treatment of HD Brazilian patients. PMID- 11105062 TI - Motor neuron diseases in the university hospital of Fortaleza (Northeastern Brazil): a clinico-demographic analysis of 87 cases. AB - In this retrospective (1980-1998) study, we have analyzed clinico demographically, from the records of the University Hospital of Fortaleza (Brazil), a group of 87 patients showing signs and symptoms of motor neuron diseases (MNDs). Their diagnosis was determined clinically and laboratorially. The WFN criteria were used for amyotrophic lateral sclerosis (ALS) diagnosis. The clinico-demographic analysis of the 87 cases of MNDs showed that 4 were diagnosed as spinal muscular atrophy (SMA), 5 cases as ALS subsets: 2 as progressive bulbar paralysis (PBP), 2 as progressive muscular atrophy (PMA) and 1 as monomelic amyotrophy (MA), and 78 cases of ALS. The latter comprised 51 males and 27 females, with a mean age of 42.02 years. They were sub-divided into 4 groups according to age: from 15 to 29 years (n= 17), 30 to 39 years (n= 18), 40 to 69 years (n= 39) and 70 to 78 years (n= 4). From the 78 ALS patients, 76 were of the classic sporadic form whilst only 2 were of the familial form. The analysis of the 87 patients with MNDs from the University Hospital of Fortaleza showed a predominance of ALS patients, with a high number of cases of juvenile and early onset adult sporadic ALS. PMID- 11105063 TI - Congenital destructive hemispheric lesions and epilepsy: clinical features and relevance of associated hippocampal atrophy. AB - We studied the clinical, EEG and MRI findings in 19 patients with epilepsy secondary to congenital destructive hemispheric insults. Patients were divided in two groups: 10 with cystic lesions (group 1), and 9 with atrophic lesions (group 2). Seizure and EEG features, as well as developmental sequelae were similar between the two groups, except for the finding that patients of group 2 more commonly presented seizures with more than one semiological type. MRI showed hyperintense T2 signal extending beyond the lesion in almost all patients of both groups, and it was more diffuse in group 2. Associated hippocampal atrophy (HA) was observed in 70% of group 1 patients and 77.7% of group 2, and it was not correlated with duration of epilepsy or seizure frequency. There was a good concordance between HA and electroclinical localization. The high prevalence of associated HA in both groups suggests a common pathogenesis with the more obvious lesion. Our findings indicate that in some of these patients with extensive destructive lesions, there may be a more circumscribed epileptogenic area, particularly in those with cystic lesions and HA, leading to a potential rationale for effective surgical treatment. PMID- 11105064 TI - Clinical characteristics and surgical outcome of patients with temporal lobe tumors and epilepsy. AB - This is a retrospective study of 21 surgically treated patients with temporal lobe tumors and epilepsy. Evaluation included clinical data, EEG findings, structural scans, pathological diagnosis and post-surgical follow-up. There were 9 cases of ganglioglioma, 5 pilocytic astrocytoma, 3 ganglioneuroma, 2 dysembryoplastic neuroepithelial tumor, 1 pleomorphic xantoastrocytoma, and 1 meningioangiomatosis. Mean follow-up time was 22 months and outcome was evaluated according to Engel's classification; 76.2% were classified in class I and 23.8% in II and III. All patients classes II and III had been submitted to mesial and neocortical resections. There were no differences related to clinical characteristics, pathological diagnosis or duration of follow-up in patients seizure-free or not. All patients had abnormal MRI and ten of these had normal CT; the MRI characteristics were compared to pathological diagnosis and specific histological characteristics of the tumors were not discernible by MRI. We concluded that MRI was essential for the diagnosis and precise location of TL tumors. Ganglioglioma was the most frequent tumor and lesionectomy associated to mesial resection doesn't guarantee a better prognosis. PMID- 11105065 TI - Results of surgery in patients with bilateral independent temporal lobe spiking (BITLS) with normal MRI or bilateral mesial temporal sclerosis (MTS) investigated with bilateral subdural grids. AB - PURPOSE: The introduction of new technologies in the clinical practice have greatly decreased the number of patients submitted to invasive recordings. On the other hand, some patients with refractory temporal lobe epilepsy have normal MR scans or bilateral potentially epileptogenic lesions. This paper reports the results of invasive neurophysiology and surgical outcome in such patients. METHOD: Sixteen patients were studied. Eleven had normal MRI (Group I) and five had bilateral mesial temporal sclerosis (Group II). All patients had BITLS and non-localizatory seizures on video-EEG monitoring. All patients were implanted bilaterally with 32-contacts subdural grids. They were submitted to a cortico amygdalo-total hippocampectomy at the side defined by chronic electrocorticography (ECoG). RESULTS: In Group I, seizures came from a single side in nine patients. In nine patients, seizures started at one side, spread to the ipsolateral contacts and contralaterally afterwards. On the other hand, in two Group I patients seizures started in one mesial region and spread to the contralateral parahippocampus and neocortex before spreading to ipsolateral contacts. All patients in Group II had seizures starting unilaterally with focal EcoG onset in the mesial regions. Eight Group I patients are seizure-free and three are in Engel's class II. Eighty percent of Group II patients are seizure free after surgery and one patient is in Engel's class II. CONCLUSION: Good surgical results can be obtained in patients with BITLS. Patients with normal MRI seem to have a worse prognosis when compared to patients with unilateral or even bilateral MTS. Extensive subdural coverage is essential in patients with normal MRI. PMID- 11105066 TI - Cysticidal therapy: impact on seizure control in epilepsy associated with neurocysticercosis. AB - OBJECTIVE: To evaluate the clinical features and seizure control of epilepsy related to neurocysticercosis. METHOD: 18 patients with partial epilepsy and neurocysticercosis were treated with albendazol or praziquantel and followed from 3 months to 12 years. We analyzed results from the CSF exam, interictal electroencephalogram (EEG), head computerized tomography and/or magnetic resonance imaging. RESULTS: The patients' mean age was 36.4 years. The mean duration of epilepsy was 16 years. 83% patients had simple partial seizures; 17% had complex partial seizures. All patients underwent routine EEGs: 62% had abnormalities and 38% were normal. A relationship was observed between focal EEG abnormality and the location of cyst in 28% of the patients. The CSF exams showed pleocytosis in 33% of the patients, and 28% had elevated protein levels. Only 22% of patients had positive titer for cysticercosis in the CSF. In all patients who had somatosensory and special sensory seizures there was a relationship between location of the cysts and seizure semiology (n=11). After cysticidal therapy, 83% patients had a significant improvement in controlling seizures. CONCLUSION: In this group, we found a predominance of simple partial seizures and a relationship between somatosensory and special sensory seizures and the location of the cysts. Cysticidal therapy was effective in controlling seizures in these patients and should be considered for patients with partial seizures and semiology related to cyst location. PMID- 11105067 TI - Predisposition to metabolic acidosis induced by topiramate. AB - RATIONALE: Metabolic acidosis induced by topiramate is a well documented but infrequent adverse event. The objective was to demonstrate the lowering of carbon dioxide serum levels, which is usually asymptomatic but may facilitate the occurrence of metabolic acidosis in patients using topiramate. METHODS: We evaluated, prospectively, the carbon dioxide serum levels of 18 patients seen at the epilepsy clinic of our university hospital, before and 3 months after introducing topiramate. RESULTS: Five patients were female and 13 were male, age ranging from 2 to 16 years old (mean=9. 3). Carbon dioxide mean serum levels were 25 and 21.2 mmol/L (normal = 22 to 30), before and 3 months after introducing topiramate, respectively. Dose ranged from 2.08 to 11.76 mg/kg/day (mean=6. 7mg/kg/day). Adverse events were anorexia, nausea and somnolence. CONCLUSION: We conclude that the lowering of carbon dioxide serum levels induced by topiramate is mostly asymptomatic, but may facilitate the occurrence of metabolic acidosis. Since patients in use of topiramate have refractory epilepsy, they may need epilepsy surgery, and must be carefully monitored for the risk of metabolic acidosis during surgery. PMID- 11105068 TI - Double-blind clonazepam vs placebo in panic disorder treatment. AB - OBJECTIVE: To assess the effectiveness of clonazepam, in a fixed dose (2 mg/day), compared with placebo in the treatment of panic disorder patients. METHOD: 24 panic disorder patients with agoraphobia were randomly selected. The diagnosis was obtained using the structured clinical interview for DSM-IV. All twenty-four subjects were randomly assigned to either treatment with clonazepam (2 mg/day) or placebo, during 6 weeks. Efficacy assessments included: change from baseline in the number of panic attacks; CGI scores for panic disorder; Hamilton rating scale for anxiety; and panic associated symptoms scale. RESULTS: At the therapeutic endpoint, only one of 9 placebo patients (11.1%) were free of panic attacks, compared with 8 of 13 (61.5%) clonazepam patients (Fisher exact test; p=0,031). CONCLUSION: the results provide evidence for the efficacy of clonazepam in panic disorder patients. PMID- 11105069 TI - Risk factors of neurological lesions in low cervical spine fractures and dislocations. AB - Eighty-nine patients with lower cervical spine fractures or dislocations were evaluated for risk factors of neurological lesion. The age, sex, level and pattern of fracture and sagittal diameter of the spinal canal were analysed. There were no significant differences on the age, gender, level and Torg's ratio between intact patients and those with nerve root injury, incomplete or complete spinal cord injuries. Bilateral facet dislocations and burst fractures are a significant risk factor of spinal cord injury. PMID- 11105070 TI - Streptozotocin-induced diabetes duration is important to determine changes in the number and basophily of myenteric neurons. AB - The aim of present study was to evaluate the number and basophily of cell bodies of myenteric neurons in the ileum of rats with diabetes mellitus induced by streptozotocin. Four groups of rats were used: diabetes was induced in two (D) whereas the other two worked as controls (N). Animals were sacrificed six (6N, 6D) or nineteen (19N, 19D) weeks after diabetes induction. A segment of the terminal portion of the ileum of each rat was obtained and stained with Giemsa's solution, for whole-mount preparation studies. Forty fields were analyzed in each animal, and the number and basophily intensity of cell bodies were recorded. After counting, the following mean numbers of neurons/mm2 were obtained: 6N=593.1 +/- 95.75, 6D=639.1 +/- 130.8, 19N=580.1 +/- 175.6 and 19D=402.0 +/- 144.8. The analysis of basophily shown that highest frequency of neurons with weak/intermediary basophily was verified in 6D group (55.3%), whereas the groups 6N, 19N e 19D presented 38%, 36% e 40% respectively. The statistical analysis showed that a long period is necessary to decrease the number of neurons/mm2 in the rat ileum after diabetes induction, and that there was a reduction in basophily intensity in diabetic rats after 6 weeks of treatment, and such cells do not recover after a longer period (19 weeks). PMID- 11105071 TI - Histomorphometrical study of the elastic fiber system in the anterior cerebral artery of man. AB - The aim of the present study was to quantify the distribution of the elastic fiber system within the wall of the anterior cerebral artery. The study is based on the works of Glynn (1940) and Stehbens (1989) concerning the incidence and origin of brain aneurysms and recent studies of the elastic fibers. The anterior cerebral artery was divided into three segments, S1, S2 and S3: S1 corresponds to the origin of the anterior cerebral artery, S2 is located at the junction of the anterior cerebral artery with the anterior communicating artery, and S3 at the junction of the rostrum and genu of the corpus callosum,which were submitted to routine histological procedures. A histomorphometrical study was undertaken using an estimation of the linear density (Ld) of the components of the fibrous elastic system which evaluates their full length in each segment. Data were analyzed using first order linear regression methods. The results show a decreasing quantity of elastic fibers in the three segments (S1>S2>S3). Study of the elastic fiber system may originate new concepts regarding the genesis of cerebral artery aneurysm. PMID- 11105072 TI - [Focal cerebral ischaemia induced by middle cerebral artery occlusion and the neuroprotective effect of ketoprofen in rats]. AB - Cerebral ischaemia is eventualy observed during neurosurgical procedures and in several clinical entities that may cause severe neurological deficits and even death. Because it is a severe and complex problem, several studies have been done aiming to elucidate the mechanisms of the ischemic phenomenon and aiming to abolish or to diminish its effects, using drugs that protect the neurons from ischaemia-induced damage. Several neurotransmitters play a role in cerebral ischaemia with emphasis to glutamate by its high concentration in the central nervous system. The purpose of this study was to evaluate the effect of focal cerebral ischaemia in the rat through the dosage of the glutamate and morphological findings, and to evaluate a possible protective effect of the ketoprofen to ischemic neurons. Thirty-six rats Wistar were divided into four groups. The first was a control group, the second a sham group and the animals of the third and fourth groups were submitted to induced cerebral ischaemia through selective obstruction of the midlle cerebral artery during 15, 30 and 45 minutes. Animals of the fourth group were previously treated with ketoprofen 15 minutes before the ischaemia. The ischaemia was evaluated through the histopathological examination and through dosage of the extracellular glutamate in vitro. The histopathological examination showed that there was no difference between the animals of the control and of the sham groups. In the animals submitted to ischemia histopathological alterations appeared at 30 minutes and become more intense at 45 minutes of ischaemia. The main findings were interstitial edema, chromatinic disorganization, vacuolization and nuclear desintegration. The animals treated with ketoprofen showed similar alterations, but they were less intense. Decrease in the dosage of glutamate in the parietal cortex of the animals submitted to ischaemia started at 30 minutes and became more intense at 45 minutes of ischaemia and was similar for animals previously treated or not with ketoprofen, indicating that this drug seems not to interfere with the metabolism of the glutamate at the synapses. The morphological findings in the parietal cortex of the animals submitted to ischaemia, previously treated or not with ketoprofen, suggest that this drug has a neuroprotective effect. PMID- 11105073 TI - [Pituitary adenomas: relationship between invasiveness and proliferative cell nuclear index]. AB - We evaluated clinically, radiologically and surgically a series of 76 pituitary adenomas. All cases were assessed immunohistochemically and in 49 patients the PCNA monoclonal antibody was measured. The most frequent types found were the bihormonal adenomas, followed by prolactinomas and non secreting adenomas. The bihormonal adenomas, non secreting adenonas and the sub unit alfa producing adenomas were proportionally more invase as determined by radiological criteria (CTscan or MRI). In 59 patients a transphenoidal approach was used, six cases were operated on transcranially and in 11 patients we used a combination of both approach. Total resection were achieved in 32 cases, most of which were microadenomas, in 15 cases the resection was subtotal and partial in 29 cases. Diabetes insipidus was the most frequent endocrine complication. It was observed that secreting adenomas tend to be associated with an increased PCNA and invasive adenomas correlated with PCNA 3 and 4. An improvement in vision was observed in 85% of macroadenomas seen after a total, subtotal or partial resection. PMID- 11105074 TI - [Etiology of the epileptic seizures in Recife city, Brazil: study of 249 patients]. AB - The etiological causes of the epileptic seizures presenting by 249 patients were studied, in a neurological clinic in Recife City, Brazil. The cause of the seizure was not identified in 43.0% of the patients. As suspected causal factors we found: ischemic cerebrovascular disease (ICVD, 17.3%), cysticercosis (8.8%), head trauma (HT, 6.8%), brain tumors (6.8%), hemorrhagic cerebrovascular disease (HCVD, 4.8%), vascular malformation (3.6%), other causes (8. 4%). In the patients with age less than 15-year-old, the most frequent cause was cysticercosis (10.3%), following vascular malformation (5.1%), and ICVD (5.1%). In the group between 15 and 45 years of age the major cause of seizure was cysticercosis (11.6%), following HT (10.7%), ICVD (4.5%), and brain tumors (3.6%). On the other hand, in the patients with more than 45-year-old the main cause was ICVD (36.7%), following brain tumors (12.3%), HCVD (11.2%), and cysticercosis (5.1%). Im conclusion, cerebrovascular disease was the most prevalent cause of epileptic seizures, considering all groups of patients. Although, cysticercosis was main cause found in the patients with less than 45 years of age. PMID- 11105075 TI - [Alternative diagnoses among suspected herpes simplex encephalitis patients with negative polymerase chain reaction]. AB - The aim of this study was to analyze the diagnosis found in a series of patients in which the diagnosis of Herpes simplex encephalitis (HSE) was ruled out by a negative polymerase chain reaction (PCR) result for HSV DNA in cerebrospinal fluid (CSF) samples. Forty three out of 61 HSE suspected patients had negative PCR. An alternative diagnosis was established in 41.9% of these patients. These patients were diagnosed as having viral (2 cases-11.1%) and non viral (5 cases 27.2%) CNS infections, vascular (4 cases-22.2%) and demyelinating diseases (3 cases-16.7%), metabolic disturbances (3 cases-16.7%), and CNS tumor (1 case 5.6%). The non specific clinical presentation of this disease and the availability of an efficient treatment for HSE explain why several patients with other diseases were initially treated with acyclovir. The early use of PCR in CSF was considered essential for the evaluation of the acute encephalitis cases in this study. PMID- 11105076 TI - [Intravenous immunoglobulin in children with Guillain-Barre syndrome]. AB - We report our experience with intravenous immunoglobulin (IVIG), plasmapheresis and supportive care in 13 patients with the Guillain-Barre syndrome. Seven of 13 patients received IVIG, 2 plasmapheresis and 4 supportive care. At 15th day after IVIG administration, all patients in this group had improved at least one disability grade. In the plasmapheresis group, 1 improved at 5th day after the procedure. Two of the 4 patients that received supportive care improved at 20th day of evaluation. In the IVIG group, the final scores were lower and had no relapses. These results suggest faster clinical improvement with IVIG when compared with supportive measures. PMID- 11105077 TI - [Sport as integration factor of the physically handicapped in our society]. AB - The objective of this study was to make use of sports as a rehabilitation method, as well as to assess the physical, psychological, and social aspects of those present some physical handicap, particularly those who have some kind of chronic disease and are no longer taking part in any rehabilitation program. Thirty handicapped people were evaluated: fifteen started with basketball and fifteen with swimming, according either to the specific preference of each one of them or to the degree and kind of physical impairment. They were submmited to the following evaluations: clinical examination, physiotherapy assessment, social interview and use of the Rivermead Social Scale, functional classification of the sport, use of the Barthel and Rivermead Functional Scales, and the psychological profile test (POMS). After two years, no relevant change in the moving evolution of the athletes were reported. Concerning the POMS psychological test, both basketball and swimming groups presented with high vigor and low depression levels. Considering the social aspects, both groups presented substantial improvement, specially regarding their relationship to one person or more people and also in the everyday activities (be it social, leisure, or domestic), thus leading them to better social integration. This essay shows that sport can bring people who are physically impaired a better social integration and physical conditions. PMID- 11105078 TI - [Central nervous system neurocytomas: clinicopathological analysis of tree cases]. AB - Central nervous system neurocytoma is a rare benign tumor of neuronal origin. Because of some clinical and radiological findings CNS neurocytomas were confused with other intraventricular lesions (ependymomas, choroid plexus papilloma, oligodendrogliomas, subependymal astrocytomas). Pathological diagnosis improved with immunohistochemical and electron microscopic studies. We present three cases of intraventricular neurocytomas confirmed by immunohistochemical studies. According to the literature clinical signs, radiological features, surgical and pathological findings are discussed. PMID- 11105079 TI - Toxoplasma gondii myelitis in a patient with adult T-cell leukemia-lymphoma. AB - Adult T cell leukemia-lymphoma (ATL) caused by HTLV-I may be associated with severe immunosupression and several opportunistic infections. Toxoplasmic encephalitis is a common central nervous system opportunistic infection in severely immunosupressed patients, however spinal cord involvement by this parasite is rare. In this paper, we report a case of toxoplasmic myelitis in a patient with ATL. PMID- 11105081 TI - [Broca and the beginning of modern neurosurgery] [In Process Citation] AB - This study presents Broca's pioneering efforts on cerebral localizations and craniotopography and their application on the first craniotomy based on cerebral localization. PMID- 11105080 TI - Invasive medullary thymoma associated with Myasthenia gravis: an unusual case. AB - Thymomas are tumors characterized by a remarkable morphological heterogeneity and variable clinical behavior. This tumor has unique clinical associations, most notably with hematological abnormalities and myasthenia gravis. According with the Muller-Hermelink criteria, there are significant differences between the histological types of thymomas and the association with myasthenia gravis. Among the different histological types, medullary thymoma is the least frequent variant associated with this autoimmune disease. In this report we describe a case of medullary thymoma presenting in a 71-year- old woman with a myasthenic syndrome. PMID- 11105082 TI - [Miller fisher syndrome and optic neuritis: case report]. AB - We report a case of Miller Fisher syndrome and bilateral demyelinating optic neuropathy suggesting the possible involvement of central nervous system in this syndrome. The optic neuritis was confirmed by visual evoked potential. PMID- 11105083 TI - [Lafora's disease: diagnosis by muscle biopsy (case report)]. AB - A 16-year-old female patient had myoclonic epilepsy caused by Lafora's disease. Muscle biopsy showed a prominent splitting pattern in muscle fibers with the nicotinamide adenine nucleotide dehydrogenase-tetrazolium reductase reaction, hematoxylin-eosin, and PAS stains. This morphologic appearance of the tissue permits diagnosis using the benign technique of muscle biopsy. The ultrastructural examination of muscle may be necessary to confirm the diagnosis of Lafora myoclonus epilepsy if light microscopical findings are equivocal. PMID- 11105084 TI - Emery-Dreifuss muscular dystrophy: anatomical-clinical correlation (case report). AB - We report on a man that had weakness of humeroperoneal distribution associated with limited range of motion of the cervical spine and elbows since he was 5 years old. At age 26 he developed tachycardia episodes. A complex arrhythmia was discovered, and a nodal ablation was done with a cardiac pacemaker implanted. The patient had an arrhythmia and sudden death followed this. Emery-Dreifuss muscular dystrophy is a rare recessive X-linked muscular disorder where mixed patterns in electromyography and muscle histology (neurogenic and/or myopathic) have caused nosological confusion. The autopsy findings are here described and correlated to the clinical features in an attempt to better understand the ambiguous findings concerning the process etiology. PMID- 11105085 TI - [Susacs syndrome: case report]. AB - We present the case of a 34-year-old woman with clinical picture suggestive of Susacs syndrome, or retinocochlear vasculopathy. This syndrome, which was described for the first time in 1979, is characterized by a clinical triad of encephalopathy, neurosensorial deafness and visual deficit. Its pathogenesis and treatment are still disputed. We have called special attention to differential diagnosis, since this entity has not yet been described in Brazil, and is probably underdiagnosed. PMID- 11105086 TI - [Multiple sclerosis, spinal cord ependymoma and intracranial meningioma: case report]. AB - We report the association a multiple sclerosis (MS), spinal cord tumour and intracranial tumor in a 63 years-old female patient with a 10 years history of relapsing/remitting MS. Symptoms usually remitted in response to costicosteroid therapy. In 1997 the patient presented with paraparesis and paresis of right arm which did not respond to corticotherapy. A spinal RMI revealed in the cervical spinal an intra spinal cord tumour, further diagnosed as ependymoma, and a parietal region meningioma. We call attention to this rare association of central nervous system tumour and MS, enphasizing the need for investigation of new and uncommon symptoms during the evolution of MS. PMID- 11105087 TI - [Unusual cause for bilateral trigeminal neuralgia: unilateral racemous cysticercus of cerebellopontine angle. Case report]. AB - We report the case of a 42-year-old woman with a racemous cystecercus in the right cerebellopontine angle (CPA), who presented with bilateral trigeminal neuralgia. The parasite was completly removed via a right suboccipital craniotomy. On the first postoperative day, the patient indicated that the pain disappeared. The neuralgia was caused by two probable mechanisms: a distortion of the brain stem and compression of the nerve against an arterial loop at the entry zone or arachnoiditis caused by the parasite in the both CPA cisternae. This case demonstrates the advisability of obtaining imaging studies in all patients with trigeminal neuralgia before starting any management. We must always remind that the cysticercus may be a differential diagnosis of CPA lesions. PMID- 11105088 TI - [Carpal tunnel syndrome: controversies regarding clinical and electrodiagnosis and its work-relatedness]. AB - The diagnosis of carpal tunnel syndrome (CTS) may be difficult because paresthesias and tingling in the upper limbs are commonly reported in the general population. These symptoms are poorly correlated with changes of nerve conduction studies of the median nerve. CTS should be diagnosed only when typical symptoms are associated with significant electrophysiological abnormalities. The association of CTS with work is highly controversial. PMID- 11105089 TI - Machado de Assis's own writings about his epilepsy: a brief clinical note. AB - Machado de Assis's own writings about his epilepsy are here given. They come from his correspondence with his friend Mario de Alencar during the last 8 months of Machado de Assis's life. These are the only places where Machado de Assis dealt clearly with his epilepsy during his entire life. PMID- 11105090 TI - The pathogenesis of tropical spastic paraparesis/human T-cell leukemia type I associated myelopathy. AB - Tropical spastic paraparesis/human T-cell leukemia type I-associated myelopathy (TSP/HAM) is caused by a human T-cell leukemia virus type I (HTLV-I) after a long incubation period. TSP/HAM is characterized by a chronic progressive paraparesis with sphincter disturbances, no/mild sensory loss, the absence of spinal cord compression and seropositivity for HTLV-I antibodies. The pathogenesis of this entity is not completely known and involves a multivariable phenomenon of immune system activation against the presence of HTLV-I antigens, leading to an inflammatory process and demyelination, mainly in the thoracic spinal cord. The current hypothesis about the pathogenesis of TSP/HAM is: 1) presence of HTLV-I antigens in the lumbar spinal cord, noted by an increased DNA HTLV-I load; 2) CTL either with their lytic functions or release/production of soluble factors, such as CC-chemokines, cytokines, and adhesion molecules; 3) the presence of Tax gene expression that activates T-cell proliferation or induces an inflammatory process in the spinal cord; 4) the presence of B cells with neutralizing antibody production, or complement activation by an immune complex phenomenon, and 5) lower IL-2 and IFN-gamma production and increased IL-10, indicating drive to a cytokine type 2 pattern in the TSP/HAM subjects and the existence of a genetic background such as some HLA haplotypes. All of these factors should be implicated in TSP/HAM and further studies are necessary to investigate their role in the development of TSP/HAM. PMID- 11105091 TI - Aggrecan structure in amphibian cartilage. AB - The structure of the large proteoglycan present in the bullfrog epiphyseal cartilage was studied by immunochemical and biochemical methods. The isolated monomer showed a polydisperse behavior on Sepharose CL2B, with a peak at Kav = 0.14. Chondroitin sulfate chains were identified by HPLC analysis of the products formed by chondroitinase digestion and mercuric acetate treatment. These chains have approximately 38 disaccharides, a Di45:Di68 ratio of 1.6 and GalNAc4S + GalNAc4,6S are the main non-reducing terminals. Keratan sulfate was identified by the use of two monoclonal antibodies in Western blots after chondroitinase ABC treatment. A keratan sulfate-rich region (approximately 110 kDa) was isolated by sequential treatment with chondroitinase ABC and proteases. We also employed antibodies in Western blotting experiments and showed that the full length deglycosylated core protein is about 300 kDa after SDS-PAGE. Domain-specific antibodies revealed the presence of immunoreactive sites corresponding to G1/G2 and G3 globular domains and the characterization of this large proteoglycan as aggrecan. The results indicate the high conservation of the aggrecan domain structure in this lower vertebrate. PMID- 11105092 TI - Important amino acid residues of potato plant uncoupling protein (StUCP). AB - Chemical modifications were used to identify some of the functionally important amino acid residues of the potato plant uncoupling protein (StUCP). The proton dependent swelling of potato mitochondria in K(+)-acetate in the presence of linoleic acid and valinomycin was inhibited by mersalyl (K(i) = 5 microM) and other hydrophilic SH reagents such as Thiolyte MB, iodoacetate and 5, 5'-dithio bis-(2-nitrobenzoate), but not by hydrophobic N-ethylmaleimide. This pattern of inhibition by SH reagents was similar to that of brown adipose tissue uncoupling protein (UCP1). As with UCP1, the arginine reagent 2,3-butadione, but not N ethylmaleimide or other hydrophobic SH reagents, prevented the inhibition of StUCP-mediated transport by ATP in isolated potato mitochondria or with reconstituted StUCP. The results indicate that the most reactive amino acid residues in UCP1 and StUCP are similar, with the exception of N-ethylmaleimide reactive cysteines in the purine nucleotide-binding site. PMID- 11105093 TI - A high-fructose diet induces changes in pp185 phosphorylation in muscle and liver of rats. AB - Insulin stimulates the tyrosine kinase activity of its receptor resulting in the tyrosine phosphorylation of pp185, which contains insulin receptor substrates IRS 1 and IRS-2. These early steps in insulin action are essential for the metabolic effects of insulin. Feeding animals a high-fructose diet results in insulin resistance. However, the exact molecular mechanism underlying this effect is unknown. In the present study, we determined the levels and phosphorylation status of the insulin receptor and pp185 (IRS-(1/2)) in liver and muscle of rats submitted to a high-fructose diet evaluated by immunoblotting with specific antibodies. Feeding fructose (28 days) induced a discrete insulin resistance, as demonstrated by the insulin tolerance test. Plasma glucose and serum insulin and cholesterol levels of the two groups of rats, fructose-fed and control, were similar, whereas plasma triacylglycerol concentration was significantly increased in the rats submitted to the fructose diet (P<0.05). There were no changes in insulin receptor concentration in the liver or muscle of either group. However, insulin-stimulated receptor autophosphorylation was reduced to 72 +/- 4% (P<0.05) in the liver of high-fructose rats. The IRS-1 protein levels were similar in both liver and muscle of the two groups of rats. In contrast, there was a significant decrease in insulin-induced pp185 (IRS-(1/2)) phosphorylation, to 83 +/- 5% (P<0.05) in liver and to 77 +/- 4% (P<0.05) in muscle of the high-fructose rats. These data suggest that changes in the early steps of insulin signal transduction may have an important role in the insulin resistance induced by high-fructose feeding. PMID- 11105094 TI - Bone mineral density in Brazilian men 50 years and older. AB - Bone mineral density (BMD) in the lumbar spine (LSBMD), femoral neck (FNBMD) and whole body (WBBMD) and whole body tissue composition were evaluated in 288 Brazilian men 50 years and older, 80% white and 20% Mulattoes. Age was inversely correlated with WBBMD (r = -0.20) and FNBMD (r = -0.21) but not with LSBMD (r = 0.03). Body mass index and weight showed a strong positive correlation with WBBMD (r = 0.48 and 0.54), LSBMD (r = 0.37 and 0.45) and FNBMD (r = 0.42 and 0.48). Correlation with height was positive but weaker. No significant bone loss at the lumbar spine level was observed as the population aged. FNBMD and WBBMD decreased significantly only in the last decade (age 70-79) studied. BMD was higher for Brazilian men as compared to Brazilian women at all sites. No significant differences were observed between Brazilian and the US/European male population for BMD in the femoral neck. BMD measured by dual-energy X-ray absorptiometry in South American men is reported here for the first time. A decrease in FNBMD was detected only later in life, with a pattern similar to that described for the US/European male population. PMID- 11105095 TI - Studies of the small bowel surface by scanning electron microscopy in infants with persistent diarrhea. AB - We describe the ultrastructural abnormalities of the small bowel surface in 16 infants with persistent diarrhea. The age range of the patients was 2 to 10 months, mean 4.8 months. All patients had diarrhea lasting 14 or more days. Bacterial overgrowth of the colonic microflora in the jejunal secretion, at concentrations above 10(4) colonies/ml, was present in 11 (68.7%) patients. The stool culture was positive for an enteropathogenic agent in 8 (50.0%) patients: for EPEC O111 in 2, EPEC O119 in 1, EAEC in 1, and Shigella flexneri in 1; mixed infections due to EPEC O111 and EAEC in 1 patient, EPEC O119 and EAEC in 1 and EPEC O55, EPEC O111, EAEC and Shigella sonnei in 1. Morphological abnormalities in the small bowel mucosa were observed in all 16 patients, varying in intensity from moderate 9 (56.3%) to severe 7 (43.7%). The scanning electron microscopic study of small bowel biopsies from these subjects showed several surface abnormalities. At low magnification (100X) most of the villi showed mild to moderate stunting, but on several occasions there was subtotal villus atrophy. At higher magnification (7,500X) photomicrographs showed derangement of the enterocytes; on several occasions the cell borders were not clearly defined and very often microvilli were decreased in number and height; in some areas there was a total disappearance of the microvilli. In half of the patients a mucus fibrinoid pseudomembrane was seen partially coating the enterocytes, a finding that provides additional information on the pathophysiology of persistent diarrhea. PMID- 11105096 TI - Hypomagnesemia in critically ill cancer patients: a prospective study of predictive factors. AB - Hypomagnesemia is the most common electrolyte disturbance seen upon admission to the intensive care unit (ICU). Reliable predictors of its occurrence are not described. The objective of this prospective study was to determine factors predictive of hypomagnesemia upon admission to the ICU. In a single tertiary cancer center, 226 patients with different diagnoses upon entering were studied. Hypomagnesemia was defined by serum levels <1.5 mg/dl. Demographic data, type of cancer, cause of admission, previous history of arrhythmia, cardiovascular disease, renal failure, drug administration (particularly diuretics, antiarrhythmics, chemotherapy and platinum compounds), previous nutrition intake and presence of hypovolemia were recorded for each patient. Blood was collected for determination of serum magnesium, potassium, sodium, calcium, phosphorus, blood urea nitrogen and creatinine levels. Upon admission, 103 (45.6%) patients had hypomagnesemia and 123 (54.4%) had normomagnesemia. A normal dietary habit prior to ICU admission was associated with normal Mg levels (P = 0.007) and higher average levels of serum Mg (P = 0.002). Postoperative patients (N = 182) had lower levels of serum Mg (0.60 +/- 0.14 mmol/l compared with 0.66 +/- 0.17 mmol/l, P = 0.006). A stepwise multiple linear regression disclosed that only normal dietary habits (OR = 0.45; CI = 0.26-0.79) and the fact of being a postoperative patient (OR = 2.42; CI = 1. 17-4.98) were significantly correlated with serum Mg levels (overall model probability = 0.001). These findings should be used to identify patients at risk for such disturbance, even in other critically ill populations. PMID- 11105097 TI - A comparative study of aging of the elastic fiber system of the diaphragm and the rectus abdominis muscles in rats. AB - In the present study the age-related changes of the striated muscle elastic fiber system were investigated in the diaphragm and rectus abdominis muscles of 1-, 4-, 8- and 18-month-old rats. The activation patterns of these muscles differ in that the diaphragm is regularly mobilized tens of times every minute during the entire life of the animal whereas the rectus abdominis, although mobilized in respiration, is much less and more irregularly activated. The elastic fibers were stained by the Verhoeff technique for mature elastic fibers. Weigert stain was used to stain mature and elaunin elastic fibers, and Weigert-oxone to stain mature, elaunin and oxytalan elastic fibers. The density of mature and elaunin elastic fibers showed a progressive increase with age, whereas the amount of oxytalan elastic fibers decreased in both diaphragm and rectus abdominis muscles and their muscular fasciae. These age-related quantitative and structural changes of the elastic fiber system may reduce the viscoelastic properties of the diaphragm and rectus abdominis muscles, which may compromise the transmission of tensile muscle strength to the tendons and may affect maximum total strength. PMID- 11105098 TI - CD4(+) T cells participate in the nephropathy of canine visceral leishmaniasis. AB - Renal involvement in visceral leishmaniasis (VL) is very frequent. The renal lesions of humans and dogs are similar but their pathogenesis has not been clearly elucidated. There is growing evidence that the cellular immune response is involved in the pathogenesis of immunologically mediated glomerulonephritis. Since T cells could participate in the pathogenesis of nephropathy, in the present study we investigated the possible involvement of CD4(+) and CD8(+) T cells in the nephropathy of canine VL. Six dogs naturally infected with Leishmania (Leishmania) chagasi from the endemic area in the Northeast of Brazil, the town of Teresina in the State of Piaui, were studied. An expressive inflammatory infiltrate of CD4(+) T cells both in glomeruli and in interstitium was present in 4 animals and absent in 2. CD8(+) T cells were detected only in one animal. CD4(+) T cells alone were observed in 3 animals; when CD8+ T cells were present CD4(+) T cells were also present. CD4(+) T cells were observed in cases of focal segmental glomerulosclerosis, diffuse membranoproliferative glomerulonephritis, diffuse mesangial proliferative glomerulonephritis and crescentic glomerulonephritis. CD8(+) T cells were present only in a case of crescentic glomerulonephritis. Leishmania antigen was detected in glomeruli and in interstitial inflammatory infiltrate in 4 animals and immunoglobulins were observed in 4 dogs. In this study we observed that T cells, in addition to immunoglobulins, are present in the renal lesion of canine VL. Further studies are in progress addressing the immunopathogenic mechanisms involving the participation of immunoglobulins and T cells in canine VL nephropathy. PMID- 11105099 TI - Production and characterization of monoclonal antibodies to Brazilian isolates of bovine viral diarrhea virus. AB - Three Brazilian isolates of bovine viral diarrhea virus (BVDV), antigenically distinct from the standard North American isolates, were selected to immunize BALB/c mice in order to obtain hybridoma cells secreting anti-BVDV monoclonal antibodies (mAbs). Two hybridoma clones secreting mAbs, reacting specifically with BVDV-infected cells (mAbs 3.1C4 and 6.F11), were selected after five fusions and screening of 1001 hypoxanthine-aminopterin-thymidine-resistant clones. These mAbs reacted in an indirect fluorescent antibody (IFA) assay with all 39 South and North American BVDV field isolates and reference strains available in our laboratory, yet failed to recognize other pestiviruses, namely the hog cholera virus. The mAbs reacted at dilutions up to 1:25,600 (ascitic fluid) and 1:100 (hybridoma culture supernatant) in IFA and immunoperoxidase (IPX) staining of BVDV-infected cells but only mAb 3.1C4 neutralized virus infectivity. Furthermore, both mAbs failed to recognize BVDV proteins by IPX in formalin-fixed paraffin-embedded tissues and following SDS-PAGE and immunoblot analysis of virus infected cells, suggesting they are probably directed to conformational-type epitopes. The protein specificity of these mAbs was then determined by IFA staining of CV-1 cells transiently expressing each of the BVDV proteins: mAb 3. 1C4 reacted with the structural protein E2/gp53 and mAb 6.F11 reacted with the structural protein E1/gp25. Both mAbs were shown to be of the IgG2a isotype. To our knowledge, these are the first mAbs produced against South American BVDV isolates and will certainly be useful for research and diagnostic purposes. PMID- 11105100 TI - Non-neuronal cells are not the limiting factor for the low axonal regeneration in C57BL/6J mice. AB - Peripheral axonal regeneration was investigated in adult male mice of the C57BL/6J (C), BALB/cJ (B) and A/J (A) strains and in their F1 descendants using a predegenerated nerve transplantation model. Four types of transplants were performed: 1) isotransplants between animals of the C, B and A strains; 2) donors of the C strain and recipients of the C x B and C x A breeding; 3) donors of the B strain and recipients of the C x B breeding, and 4) donors of the A strain and recipients of the C x A breeding. Donors had the left sciatic nerve transected and two weeks later a segment of the distal stump was transplanted into the recipient. Four weeks after transplantation the regenerated nerves were used to determine the total number of regenerated myelinated fibers (TMF), diameter of myelinated fibers (FD) and myelin thickness (MT). The highest TMF values were obtained in the groups where C57BL/6J mice were the donors (C to F1 (C x B) = 4658 +/- 304; C to F1 (C x A) = 3899 +/- 198). Also, A/J grafts led to a significantly higher TMF (A to F1 (C x A) = 3933 +/- 565). Additionally, isotransplant experiments showed that when the nerve is previously degenerated, C57BL/6J mice display the largest number of myelinated fibers (C to C = 3136 +/- 287; B to B = 2759 +/- 170, and A to A = 2835 +/- 239). We also observed that when C57BL/6J was the graft donor, FD was the highest and MT did not differ significantly when compared with the other groups. These morphometric results reinforce the idea that Schwann cells and the nerve environment of C57BL/6J provide enough support to the regenerative process. In this respect, the present results support the hypothesis that the non-neuronal cells, mainly Schwann cells, present in the sciatic nerve of C57BL/6J mice are not the main limiting factor responsible for low axonal regeneration. PMID- 11105101 TI - Early myelin breakdown following sural nerve crush: a freeze-fracture study. AB - In this study we describe the early changes of the myelin sheath following surgical nerve crush. We used the freeze-fracture technique to better evaluate myelin alterations during an early stage of Wallerian degeneration. Rat sural nerves were experimentally crushed and animals were sacrificed by transcardiac perfusion 30 h after surgery. Segments of the nerves were processed for routine transmission electron microscopy and freeze-fracture techniques. Our results show that 30 h after the lesion there was asynchrony in the pattern of Wallerian degeneration, with different nerve fibers exhibiting variable degrees of axon disruption. This was observed by both techniques. Careful examination of several replicas revealed early changes in myelin membranes represented by vacuolization and splitting of consecutive lamellae, rearrangement of intramembranous particles and disappearance of paranodal transverse bands associated or not with retraction of paranodal myelin terminal loops from the axolemma. These alterations are compatible with a direct injury to the myelin sheath following nerve crush. The results are discussed in terms of a similar mechanism underlying both axon and myelin breakdown. PMID- 11105102 TI - Enhancement of declarative memory associated with emotional content in a Brazilian sample. AB - Several studies have documented that emotional arousal may enhance long-term memory. This is an adaptation of a paradigm previously used in North American and European samples in investigations of the influence of emotion on long-term retention. A sample of 46 healthy adults of high and low educational levels watched a slide presentation of stories. A randomly assigned group watched a story with an arousing content and another group watched a neutral story. The stories were matched for structure and comprehensibility and the set and order of the 11 slides were the same in both conditions. Immediately after viewing the slide presentation, the participants were asked to rate the emotionality of the narrative. The arousing narrative was rated as being more emotional than the neutral narrative (t(44) = -3.6, P<0.001). Ten days later subjects were asked to remember the story and answer a multiple-choice questionnaire about it. The subjects who watched the arousing story had higher scores in the free recall measure (t(44) = -2.59, P<0. 01). There were no differences between groups in the multiple-choice test of recognition memory (t(44) = 0.26). These findings confirm that an emotional arousing content enhances long-term declarative memory and indicate the possibility of applying this instrument to clinical samples of various cultural backgrounds. PMID- 11105103 TI - Acute AT1 receptor blockade does not improve the depressed baroreflex in rats with chronic renal hypertension. AB - To assess the role of angiotensin II in the sensitivity of the baroreflex control of heart rate (HR) in normotensive rats (N = 6) and chronically hypertensive rats (1K1C, 2 months, N = 7), reflex changes of HR were evaluated before and after (15 min) the administration of a selective angiotensin II receptor antagonist (losartan, 10 mg/kg, iv). Baseline values of mean arterial pressure (MAP) were higher in hypertensive rats (195 +/- 6 mmHg) than in normotensive rats (110 +/- 2 mmHg). Losartan administration promoted a decrease in MAP only in hypertensive rats (16%), with no changes in HR. During the control period, the sensitivity of the bradycardic and tachycardic responses to acute MAP changes were depressed in hypertensive rats (approximately 70% and approximately 65%, respectively) and remained unchanged after losartan administration. Plasma renin activity was similar in the two groups. The present study demonstrates that acute blockade of AT1 receptors with losartan lowers the MAP in chronic renal hypertensive rats without reversal of baroreflex hyposensitivity, suggesting that the impairment of baroreflex control of HR is not dependent on an increased angiotensin II level. PMID- 11105104 TI - Oxygen consumption of rats with broad intestinal resection. AB - The study was performed to investigate possible alterations in oxygen consumption in an animal model with broad intestinal resection. Oxygen consumption and the thermal effect of a short meal were measured in rats subjected to short bowel syndrome. Four groups of rats were used. Group I was the control group, group II was sham operated, group III was submitted to 80% jejunum-ileum resection, and group IV was submitted to 80% jejunum-ileum resection with colon interposition. Ninety days after surgery, oxygen consumption was measured over a period of 6 h with the animals fasted overnight. The thermal effect of feeding was determined in another session of oxygen consumption measurement in animals fasted for 12 h. A 12-kcal meal was then introduced into the animal chamber and oxygen consumption was measured for a further 4 h. No differences in fasting oxygen consumption or in the thermal effect of the meal were detected among the groups studied. It is concluded that short bowel syndrome does not affect the overall energy expenditure of rats. PMID- 11105105 TI - Correlation between HIV and HCV in Brazilian prisoners: evidence for parenteral transmission inside prison. AB - OBJECTIVE: It is an accepted fact that confinement conditions increase the risk of some infections related to sexual and/or injecting drugs practices. Mathematical techniques were applied to estimate time-dependent incidence densities of HIV infection among inmates. METHODS: A total of 631 prisoners from a Brazilian prison with 4,900 inmates at that time were interviewed and their blood drawn. Risky behavior for HIV infection was analyzed, and serological tests for HIV, hepatitis C and syphilis were performed, intended as surrogates for parenteral and sexual HIV transmission, respectively. Mathematical techniques were used to estimate the incidence density ratio, as related to the time of imprisonment. RESULTS: Prevalence were: HIV - 16%; HCV - 34%; and syphilis - 18%. The main risk behaviors related to HIV infection were HCV prevalence (OR=10.49) and the acknowledged use of injecting drugs (OR=3.36). Incidence density ratio derivation showed that the risk of acquiring HIV infection increases with the time of imprisonment, peaking around three years after incarceration. CONCLUSIONS: The correlation between HIV and HCV seroprevalence and the results of the mathematical analysis suggest that HIV transmission in this population is predominantly due to parenteral exposure by injecting drug, and that it increases with time of imprisonment. PMID- 11105106 TI - [Treatment abandonment and tuberculosis recurrence: aspects of previous episodes, Brazil, 1993-1994]. AB - OBJECTIVE: Recurrent patients due to treatment dropout or disease relapse have been congesting health centers and impeding treatment routines and services. The purpose is to study aspects of previous treatment and irregularities concerning the patient, medication and service organization and to evaluate the outcome of retreatment. METHODS: A descriptive study of patient's personal characteristics, previous treatment and retreatment was carried out at Campinas, Sao Paulo State, in 1993 and 1994. Statistics analyses were performed using 95% confidence interval with Yates correction, exact Fisher test and Mantel Haenszel qui-square for stratification. RESULTS: Retreatment sequence corroborated with the default rates (63%) that were lower, but still high among relapsed cases (28.4%). Only 34.1% of relapsed cases did not present irregularities at previous treatment. CONCLUSIONS: High proportions of retreatment and inadequate previous treatment are favouring drug resistance. Efforts should be taken to improving tuberculosis services efficiency because many irregularities are due to "failures" in health services routine. PMID- 11105107 TI - [Evaluation of bacterial contamination in disinfectants for domestic use]. AB - OBJECTIVE: To evaluate disinfectants for domestic use for the presence of bacteria, identify them, and determine their tolerance level to benzalkonium chloride. METHODS: Fifty-two samples of commercially available disinfectants for domestic use were acquired at random in the metropolitan area of Sao Paulo, Brazil, and analyzed to detect the presence of bacterial contaminants. The isolated organisms were identified and their tolerance level to benzalkonium chloride was determined by broth macrodilution method. RESULTS: Sixteen (30.77%) of fifty-two disinfectants sampled were contaminated by Gram-negative bacteria, with counts varying between 10(4) and 10(6) UFC/ml. Alcaligenes xylosoxidans, Burkholderia cepacia and Serratia marcescens were the predominant organisms found. The minimum inhibitory concentration (MIC: mg/ml) of benzalkonium chloride for these bacteria were 2.48, 1.23 and 0.30 to S. marcescens, A. xylosoxidans and B. cepacia, respectively. CONCLUSIONS: The disinfectant formulation containing quaternary ammonium compounds (QACs) may be exposed to contamination by Gram negative bacteria. The MICs of benzalkonium chloride against the isolated bacteria were low, indicating that the bacteria grown in culture media without QACs lost their tolerance to this biocide. PMID- 11105108 TI - Health financing changes in the context of health care decentralization: the case of three Latin American countries. AB - OBJECTIVE: The results of an evaluative longitudinal study, which identified the effects of health care decentralization on health financing in Mexico, Nicaragua and Peru are presented in this article. METHODS: The methodology had two main phases. In the first, secondary sources of data and documents were analyzed with the following variables: type of decentralization implemented, source of financing, funds for financing, providers, final use of resources, mechanisms for resource allocation. In the second phase, primary data were collected by a survey of key personnel in the health sector. RESULTS: Results of the comparative analysis are presented, showing the changes implemented in the three countries, as well as the strengths and weaknesses of each country in matters of financing and decentralization. CONCLUSIONS: The main financing changes implemented and quantitative trends with respect to the five financing indicators are presented as a methodological tool to implement corrections and adjustments in health financing. PMID- 11105109 TI - [Adult Aedes albopictus and Ae. scapularis behavior (Diptera: Culicidae in Southeastern Brazil]. AB - OBJECTIVE: Aedes albopictus and Ae. scapularis were found living together in the Pedrinhas Village, Southeastern of Sao Paulo State, Brazil. This finding was a good opportunity to make observations about the mosquitoes' behavior. METHODS: From October 1996 to January 2000 observations were carried out through systematic collections with human bait, environment aspirations and Shannon trap utilization. Synanthropy was estimated by the Nuorteva index and synanthropic ratios. RESULTS: The 87 collections with human bait yield 872 females adults. Williams' means, multiplied by 100, were 118 and 21 for Ae. albopictus at the 7 AM-6PM and 6PM-8PM hours, respectively, 100 and 106 for Ae. scapularis at the same timetable but there was an evening peak. Through environmental aspirations, a total of 1,124 adults samples was collected, 226 Ae. albopictus samples and 898 Ae. scapularis samples. The period between the months of January-May was the one with higher yield for both mosquitoes. There was no Ae. albopictus in the Shannon trap operated inside the adjacent forest. Regarding the sinanthropy, that culicid showed the higher index values, while Ae. scapularis was ubiquitous. CONCLUSIONS: The data obtained allows to form the hypothesis that Ae. scapularis females may have a diapause phase in the resting places and after that period they will retake the hematophagy habit. That might explain the higher activity at the human bait during the dry months, corresponding to the period of July-October. PMID- 11105110 TI - [Psychiatric emergency service in a university general hospital: a prospective study]. AB - OBJECTIVES: The aim was to carry out a prospective study about the characteristics of the public seen at a psychiatric emergency room and of its service. METHODS: The data were acquired though a protocol developed for this study and applied to all the patients seen during two months. RESULTS: 600 protocols were filled out, corresponding to 96.5% (487 patients) of the attendance during the study period. Most of the patients seen were males, single, with a low educational level, professionally inactive and living with their families. The most frequent diagnoses were psychoactive substance use disorders (26.3%), schizophrenia (15.5%), manic episode (11.8%), major depression (10.9%) and non-psychotic disorders (10.9%). There were differences between gender in some diagnostic categories. After initial evaluation, 2/3 were medicated, (1/2) stayed under observation, and (1/4) stayed more than 10 hours in the service unit. About 20% of the attendance resulted in hospitalization and 60% in referrals to outpatient services. Discharges due to evasion represented only 2.0% of the total. Returning service users did not differ from those seen only once to what concern marital status, professional situation and household conditions. However, returning users presented a higher frequency of previous hospitalization and psychotic disorders. CONCLUSIONS: Individuals with severe psychiatric disorders were seen in an actual emergency situation. The psychiatric emergency service has been expanding its actions and has been an effective part of the mental health service network. PMID- 11105111 TI - [Psychiatric cases identification by multi-step epidemiological studies: methods, problems and applicability]. AB - OBJECTIVE: To discuss methodological aspects of the two stages in the identification of psychiatric cases in epidemiological studies. METHODS: Analyze the methodology used in the Multicentric Psychiatric Morbidity Study, which was conducted in three Brazilian cities (Sao Paulo, Brasilia and Porto Alegre). In the first stage of that study, a random sample (6,740 individuals) of the population was drawn and all the participants were screened with the Questionnaire of Psychiatric Morbidity of the Adult (QMPA). In the second stage, a sample (775 individuals) of this population was drawn and these individuals were submitted to the Inventory of Symptoms of DSM-III, carried out by psychiatrists and trained psychologists. RESULTS: The study procedure for estimating the prevalence is described in details, showing that though the screening scales are a weak tool, they don't interfere with the methodology. CONCLUSION: The advantage of this methodology is to correct any distortions caused by the current tools used in the identification of psychiatric cases. PMID- 11105112 TI - Causal attributions in Brazilian children's reasoning about health and illness. AB - INTRODUCTION: At a time when a great number of diseases can be prevented by changing one's habits and life style, investigations have focused on understanding what adults and children believe to be desirable health practices and uncovering the factors associated with successful adherence to such practices. For these, causal attributions for health and illness were investigated among 96 Brazilian elementary school students. METHODS: Ninety six subjects, aged 6 to 14, were interviewed individually and their causal attributions were assessed through 14 true-false items (e.g. people stay well [healthy] because they are lucky). The relationship between the children's causal attributions and demographic characteristics were also examined. RESULTS: Overall, the results were consistent with previous researches. "Taking care of oneself" was considered the most important cause of good health. "Viruses and germs" and "lack of self-care" were the most selected causes of illness. Analyses revealed significant relationship between subjects' causal attribution and their age, school grade level, socioeconomic status and gender. CONCLUSIONS: The study findings suggest that there may be more cross-cultural similarities than differences in children's causal attributions for health and illness. Finding ways to help individuals engage in appropriate preventive-maintenance health practices without developing an exaggerated notion that the individuals can control their own health and illness is a challenge which remains to be addressed by further research. PMID- 11105113 TI - [Asthmatic children's risk factors for emergency room visits, Brazil]. AB - OBJECTIVES: To study a sample of asthmatic children to get to know how the disease is managed by caretakers and to identify predictive factors associated with attendance in emergency room for asthma. METHODS: A cross-sectional study nested in a cohort was undertaken in the urban area of Pelotas, Southern Brazil. 981 children aged 4-5 years, who belong to the cohort of 1993, participated in this study. RESULTS: The asthma prevalence in the children sample was 25.4%. Morbidity for asthma was quite high: 31% of the children were seen in emergency rooms in the last year, 57% attended medical clinics and 26% were hospitalized in the first 4 years of life. The crude analysis identified the following predictive factors for emergency room visits: low educational level (RO=4.1), low family income (RO=6. 5), 3 or more children sleeping in the same room (RO=2.2), severity of asthma attacks (RO=2.7), use of asthma medicines in the last year (RO=1.9) and hospitalizations due to asthma (RO=3.0). Multivariate analyses using logistic regression were used to adjust each variable for the effect of the remainder. CONCLUSIONS: The asthma prevalence among preschool children in Pelotas is high, resulting therefore in high morbidity. The predictor factors for emergency room visits due to asthma found, after multivariate analysis, were mother's low educational level, severity of the asthma attacks and hospitalization. PMID- 11105114 TI - [Lipid profile among school children in Campinas, Brazil]. AB - OBJECTIVE: To describe the lipid profile and the prevalence of hypercholesterolemia among schoolchildren aged 7 to 14 years in Campinas, Sao Paulo State, Brazil. METHODS: Plasma cholesterol levels, fractions, ratios and triglycerides were determined according to age and gender in a total of 1,600 schoolchildren. Hypercholesterolemia was considered borderline for 170 mg/dl/=200 mg/dl. RESULTS: Schoolchildren presented a cholesterol mean of 160 mg/dl, HDL-cholesterol mean of 49 mg/dl, LDL-cholesterol mean of 96 mg/dl, VLDL-cholesterol mean of 16 mg/dl, triglycerides mean of 79 mg/dl, cholesterol/HDL-cholesterol mean of 3.5 and LDL-cholesterol/HDL cholesterol mean of 2.1. In general, females had higher cholesterol and triglycerides values than males. The prevalence of hypercholesterolemia was 35.0%: 15.6% was borderline high, 9.8% moderate and 9.5% severe. Females presented higher prevalence of hypercholesterolemia than males. CONCLUSIONS: The results pointed to the emergence of hypercholesterolemia as a public health problem in Brazil. PMID- 11105115 TI - [Diagnosis of overweight in adolescents: comparative study of the performance different criteria for body mass index]. AB - OBJECTIVE: In an attempt to simplify the screening process for detecting obesity in adolescence, the performance of different cutoff values for body mass index (BMI) was assessed in a population-based cohort in Southern Brazil. METHODS: A total of 493 adolescents aged 15-16 years who lived in the city of Pelotas, Brazil, were studied. Obesity was defined according to the WHO criteria taking into account age and sex (a BMI equal to or greater than the 85th percentile of the NHANES I reference, plus subscapular and triceps skinfold equal to or greater than the 90th percentile of the same reference). Different BMI cutoff values were used to assess their specificity and sensitivity. RESULTS: For boys, BMI>/=25 kg/m(2) showed the best performance for detecting obesity, with a sensitivity of 90% and only 5% of false positives. The Brazilian proposed criteria that was used had 100% sensitivity but up to 23% of false positives. Higher cutoff values were also tested, but there was a slight increase in specificity, accompanied by a marked reduction in sensitivity. CONCLUSIONS: The BMI cutoff of 25 kg/m(2) presented the best performance for screening obesity in the studied sample, and it is recommended for adolescents aged 15 and more in populations with similar characteristics. It provides a single cutoff value to be used in primary health services, eliminating the need for age and sex-specific values and skinfold measurements, and it is also consistent with the cutoff value proposed to identifying overweight adults. PMID- 11105116 TI - [Differential mortality between metropolitan areas of Brazil, 1985-1995]. AB - OBJECTIVE: To analyze differential changes of rates and stratification of mortality by gender and causes of death in the metropolitan area of Belo Horizonte (RMBH) and Salvador (RMS) between 1985 and 1995. METHODS: The Ministry of Health's Mortality Information System (SIM) provides data on death causes by age and sex that was used for this study. The groups of death causes were classified according to two major groups (preventable and non-preventable) and the decomposition method presented by Pollard was applied to analyze the contribution of each group of death causes in the changes in life expectation. RESULTS: There have been changes in the pace of the current mortality rate decline in RMBH and RMS, which have resulted in a reduction in the differences between the mortality rates in both areas. In both areas there was a substantial reduction in the mortality rates in the group of preventable causes, especially among women. CONCLUSIONS: There is still a structure of death causes, which seems to indicate that the improvement in mortality among the poor has been lower than it was expected. PMID- 11105117 TI - [Validity of occupational histories from proxy respondents]. AB - OBJECTIVE: Occupational studies often use data obtained from proxy respondents. However, few investigations have been conducted on the validity of occupational data provided by workers' surrogates. This study aimed to compare self-reported occupational data to information obtained from next-of-kin, as a contribution to assess the validity of using proxy respondents to obtain information about workers. METHODS: Worker/next-of-kin pairs, residents in Southeastern Brazil, were interviewed separately in 1998 about worker's occupational past history. The concordance, sensitivity, and specificity of proxy reports about workers' occupations were examined comparing to self-reports. RESULTS: A total of 2.163 worker/next-of-kin pairs were interviewed. The Kappa statistic for the agreement between worker's and next-of-kin's report about the worker's main occupation was 0.86 (CI95%; 0.85 - 0.88). The sensitivity of proxy reports on occupations ranged from 77.5% (64.6% - 90.4%) to 98.9% (97.3% - 100.0%), and specificity ranged from 96. 9% (96.0% - 97.7%) to 99.9% (99.7% - 100.0%). CONCLUSIONS: These are encouraging findings for the use of proxy respondents in occupational studies when occupations are considered as categorical variables. However, caution is required when assessing occupational exposition by means of cumulative work time because next-of-kin may underestimates this information. PMID- 11105118 TI - [Assessment of the chronic risk for ingestion of pesticide residues in the Brazilian diet]. AB - OBJECTIVE: To conduct a chronic dietary risk assessment of the pesticides registered in Brazil up until 1999. METHODS: The Theoretical Maximum Daily Intake (TMDI) for each pesticide was calculated using the Brazilian maximum residue limits and food consumption data from IBGE, the Brazilian Statistical Institute. The risk characterization was done comparing the TMDI with the acceptable daily intakes (ADI) from other countries and from the Codex Alimentarius. RESULTS: The TMDI was higher than the ADI (%ADI>100) at least in one Brazilian metropolitan region for 23 pesticides. Sixteen compounds are organophosphate insecticides, with methyl parathion having the TMDI exceeding the most toxicological parameter (%ADI N=9,300). Rice, beans, citrus and tomato were the commodities which most contributed to the ingestion. From the compounds under higher risk, only 6 were registered according to the Law 98.816/90, which concerns the use of pesticides in the country. CONCLUSIONS: The compounds identified in the study as presenting a potential health concern to the Brazilian consumers, and the commodities which most contributed to the ingestion, should be prioritized by the government in pesticide residue monitoring programs and in the re-registration process. In addition, residue data in food as consumed, processing factors and appropriate consumption data should be generated to allow further studies. PMID- 11105119 TI - [Morphology of the eggs of Triatoma circummaculata and Triatoma rubrovaria (Hemiptera, Reduviidae)]. AB - OBJECTIVE: To study morphologically the eggs of T. circummaculata and T. rubrovaria. METHODS: Forty eggs of the two species were measured through Nikon model 6C profile projector. Student test was utilized for the statistical analysis and scanning electron microscopy for the morphological study of the eggs. RESULTS/CONCLUSIONS: The statistical analysis shows that the T. rubrovaria eggs are larger that those of T. circummaculata. Microscopical observations revealed that the chorial rim and the spermatic groove are less evident in T. circummaculata than in T. rubrovaria. Both species, the majority of exochorion cells are hexagonal. It was also observed the embryo eclosion and details of the embryonic molt. PMID- 11105120 TI - [Finding of Aedes aegypti breeding in bromeliad]. AB - A breeding place of Aedes aegypti immature forms were found in bromeliads domesticated for decorative purposes. Implications for the control measures were considered. PMID- 11105121 TI - [Pachycrepoideus vindemiae (Hymenoptera: Pteromalidae) as parasite of Ophyra aenescens (Diptera: Muscidae) in Brazil]. AB - The first occurrence of the parasitoid Pachycrepoideus vindemiae on pupae of Ophyra aenescens, a fly of medical-sanitary importance, is reported. A swine carcass was used as bait to collect the insects. In the study, 302 pupae of Ophyra aenescens (Wiedemann) (Diptera: Muscidae) were obtained, 6 (1.98%) of them yielded the parasitoid Pachycrepoideus vindemiae (Hymenoptera: Pteromalidae PMID- 11105122 TI - [Evaluation of health programs, services and technologies]. AB - The field of program, services and technology evaluation in general, and in health care in particular, is going through an important growth and conceptual and methodological diversification. It is also the object of an increasing demand for its participation as an effective supportive instrument in the decision making process, and a constant need in the dynamics of health systems and services. In this paper, based on literature review, nuclear criteria involved in the organization of all evaluation processes are identified, and articulated with the existing institutionalized evaluation practices in developed countries, that is, program evaluation, quality assessment and management and technology assessment. In conclusion, the incipient development of a methodological evaluation output in Brazil is analyzed. PMID- 11105124 TI - [The national health card: a tool for a new model of care]. PMID- 11105123 TI - [Underlying causes of death and maternal mortality]. PMID- 11105125 TI - Vaccine Adjuvants: A New Hope for Effective Immunizations. AB - Two important issues regarding the use of immunization to control infections and malignancies in the future are: 1) the need to render poorly immunogenic, often highly purified, antigens more effective; and 2) the desire to direct the immune response in specific ways to achieve the most relevant response for each disease. The first issue can be solved by understanding and then recreating and enhancing the antigen presentation process by a broad range of vaccine adjuvants. The second issue requires careful selection among the adjuvants to allow directing of the immune response in the most appropriate manner. For example, in different settings expansion of a B cell response, cytotoxic T cell response, or enhancement of either a Thl or Th2 subset response may be desired. These goals are accomplished by the use of several newly developed non-cytokine adjuvants, or by direct injection of the relevant cytokines. Some non-cytokine molecular adjuvants and cytokines used as adjuvants have already been proven effective in animal models and/or in clinical trials. Here, we review the present state of the art in the use of vaccine adjuvants for control of various infectious diseases. PMID- 11105126 TI - Use of Standard Therapy for Tuberculosis is Associated with Increased Adverse Reactions in Patients with HIV. AB - Hepatitis due to anti-tuberculosis therapy is an infrequent, but potentially devastating event. In HIV positive patients with tuberculosis (TB), the consequences are likely to be even greater, as they frequently require other hepatotoxic medications. The object of our study was to determine the frequency of toxic hepatitis during therapy for TB. Included were 198 patients with a presumed or confirmed diagnosis of tuberculosis; of whom, 69 were HIV positive (35%), 75 were negative (38%) and 54 had unknown HIV status (27%). Toxic hepatitis occurred in 15/198 (8%) patients. The incidence of hepatitis in HIV patients was much greater than in HIV negative/unknown [RR=7.5 (2.2-25.6); p=0.0001] and the onset of hepatitis was short (median 7 days in HIV patients). During TB therapy, 1 in S (12.5%) patients taking ketoconazole developed hepatitis; 9/53 (17%) taking sulfamethoxazole-trimethoprim [RR=3.4 (1.1-9.3); p=0.03]. Among the 15 patients who developed hepatitis 11 required hospitalization (mean 19 days), 5 died (33.3%), 2/15 (13%) due to hepatitis. HIV positive patients had a significantly higher rate of toxic hepatitis during anti tuberculosis therapy than those without HIV infection. Hepatitis occurred just after initiation of TB treatment. Clinical findings were non-specific and hepatic enzyme elevations were moderate, yet hospitalization and mortality rates were high. This suggests that in settings where careful monitoring of patients early in their course of TB treatment is routine, morbidity and mortality may be low, but poor monitoring would have potentially serious consequences. There is a need for new drug treatments (schedules or regimens) for TB in an effort to reduce these adverse events. PMID- 11105127 TI - Prospective Evaluation of In Vitro Production of HIV-1 Specific Antibodies (IVAP), p24 Antigenemia and Viral Culture for the Diagnosis of HIV Infection in Children Born to HIV-1 Infected Mothers. AB - Three methods (IVAP, p24 antigenemia and viral cultivation) for the diagnosis of HIV-1 infection among children born to HIV-1 infected mothers were prospectively evaluated to determine the applicability of IVAP as a useful technique for that purpose. We tested 15 children (8 p0 and 7 pII) and 19 adults with well established serological status for HIV infection. The children were followed for at least 1 year, unless tbey developed symptoms of clinical AIDS, or their HIV serology became negative. The IMP method was more sensitive and specific than the other 2 tests in determining whether or not the infants were infected with HIV. All negative test results (5/8) were confirmed during the follow-up period, while two-thirds of the IVAP positive children developed AIDS during the same time period. Despite the small sample studied, we conclude that IVAP is an inexpensive and simple technique potentially useful to establish whether or not HIV seropositive children born to infected mothers are HIV infected. PMID- 11105128 TI - Malaria Transmission Associated with Airplane Travel. AB - Plasmodium falciparum malaria was diagnosed in 3 patients in Sa o Paulo during a 5 day period between August 31, and September 4, 1996, at a time and place where malaria transmission does not occur. After investigation of the 3 cases it was determined that the infections were acquired as a result of an international airplane flight from Lebanon to Sao Paulo on August 16, which included a 30 minute stop-over in Abidjan, Ivory Coast, Africa. During the epidemiological evaluation, it was found that each of the 3 patients had been seated in the first class cabin. Entomological investigation at the airport revealed the presence of 4 specimens of Anopheles gambiae in airplanes (3 in the first class cabin and 1 in the luggage compartment) used on this route. The species of mosquito identified is predominant in Africa. Two of the patients were seriously ill, but all recovered after treatment with either mefloquine (1 patient) or artesunate (2 patients). A survey of other passengers on the same flight or on similar Aights did not reveal any other eases of malaria. Malaria was not considered during initial evaluation by the attending physicians at the three different hospitals where the patients were admitted. These cases reveal the existence of vector borne disease transmission during airplane travel, and emphasize the importance of obtaining a travel history during the evaluation of an ill patient. In addition, the cases reinforce the need for vigilance in the control of vectors of disease around seaports, airports and hospitals. PMID- 11105129 TI - The ratio of plasma levels of IL-10/TNF-alpha and its relationship to disease severity and survival in patients with leptospirosis. AB - Twelve patients with leptospirosis were enrolled in a study to evaluate whether a correlation exists between disease severity and plasma levels of TNF-alpha and IL 10 as determined by enzyme-linked immunoassay. Six patients had less severe disease by analysis of liver, kidney and lung involvement, and 6 patients had severe disease, 2 of whom died. The results of the study confirm an association between high levels of TNF-alpha and poor outcome in leptospirosis. Levels of IL 10 were in a similar range in the 10 surviving and the 2 non-surviving patients. The ratio of IL-10/TNF-a correlated with disease severity in that a high ratio was associated with less severe disease and survival. The possibility is raised that treatment approaches to change this cytokine profile may be more effective in this type of inflammatory condition than it has been in bacterial sepsis. PMID- 11105130 TI - Cutaneous Leishmaniasis Coinfection in AIDS Patients: Case Report and Literature Review. AB - A fatal case of mucosal leishmaniasis in a patient with AIDS is presented. Seventeen cases of L. brasiliensis coinfection in AIDS patients in Brazil have been presented at the meetings of The Brazilian Society of Infectious Diseases. As in the case presented here, 4 cases with mucosal involvement also had negative delayed type hypersensitivity reactions to leishmania antigens. This lack of immunologic responsiveness, the invasive nature of the infection, and the poor response to therapy indicate that the combination of these two infections should be viewed as a new disease entity. PMID- 11105131 TI - When Leishmania and HIV Interact, a New Broad Spectrum of Leishmaniasis Occurs. PMID- 11105132 TI - Can We Alter the Cytokine Balance in the Management of Infectious Diseases? PMID- 11105133 TI - Emerging Pathogens Associated with Infectious Diarrhea. AB - In recent years, emerging microbial pathogens associated with infectious diarrhea have caused significant morbidity and mortality. Although sporadic cases of infectious diarrhea have occurred worldwide in the past, recent outbreaks in the United States traced to contaminated water or food have raised concerns about the safety of the water supply and the adequacy of surveillance of the food supply and foodborne diseases. In 1993, Cryptosporidium parvum, an important cause of unrelenting diarrhea and severe weight loss in AIDS patients, was associated with the largest outbreak of infectious diarrhea caused by contaminated municipal water that has ever been reported in the U.S. During the early summer of 1996, a major outbreak of Cyclospora cayetanensis that infected approximately 1,500 persons in 20 states, Washington, D.C. and two Canadian provinces was reported from North America. The suspected food vehicle in this outbreak was contaminated raspberries imported from Guatemala. In addition to these coccidian protozoa,Escherichia coli 0157:H7, first recognized in 1982 as a cause of hemorrhagic colitis, has recently been responsible for a multi-state outbreak in the U.S. due to contamination of commercial ground beef, and an outbreak in Japan that infected over 9,500 persons, two-thirds of whom were children. The contaminated food vehicle in the latter outbreak, although suspected to be radish sprouts, remains unknown. These recent massive outbreaks underscore the importance of a well-established public health infrastructure and an effective surveillance system for the early identification and reporting of infected patients that will lead to appropriate epidemiologic investigations and the rapid detection of contaminated vehicles. PMID- 11105134 TI - Detection of Toxoplasma gondii Antigen in Cerebrospinal Fluid Samples Using a Dot Enzyme-Linked Immunosorbent Assay. AB - Toxoplasma encephalitis (TE) is the most common manifestation of a recurrent Toxoplasma gondii infection in immunocompromised individuals, especially in AIDS patients. The infection is associated with considerable morbidity and mortality, even with improved diagnostic procedures and adequate treatment. The diagnosis of TE is difficult to establish by the demonstration of the parasite in the central nervous system or in cerebrospinal fluid (CSF). In order to diagnose TE we have developed a TGA-Dot-ELISA for antigen detection using rabbit anti-toxoplasma antiserum. We have applied this test to specimens of cerebrospinal fluid from patients with toxoplasma encephalitis (180 patients), neurocysticercosis (15), or bacterial meningitis (15). The relative sensitivity and specificity obtained for the TGA-Dot-ELISA were 0.930 and 0.730, respectively. TGA-Dot-ELISA is a promising method for the rapid diagnosis and effective prophylaxis of toxoplasma encephalitis. PMID- 11105135 TI - Central Venous Catheter-Related Infections in Intensive Care Units. AB - Objective: To determine the rates of colonization and infection related to central venous catheter (CVC), the causative microorganisms, and the influence of various factors. Methods: From June to August 1993, all CVC in 4 Intensive Care Units were evaluated from their insertion to removaL Data were collected by 3 nurses. Blood and catheter tips were cultured. Results: Of 84 catheters, 29.8% were colonized, 9.5% of patients showed evidence of local infection, and 4.8% had primary bloodstream infections. The internal jugular vein was the most common site for catheter insertion (81%). Causes of removal were: end of need (48.3%), suspected infection (23.3%), malfunction (20%), routine change (8.3%). Among removals because of suspected infection, 50% presented evidence of local infection, 43% were colonized(>15cfu), but there were no bloodstream infections. The average time of catheter use for those which became colonized was longer than for catheters that did not become colonized (p=0.008). The average time of catheter use associated with removal for infection (local and bloodstream) was longer than for removal for other reasons (p=0.042). Among colonized catheters, 16% developed bloodstream infection and 20% local infection. Immunosupressive drugs, cancer, diabetes mellitus, HIV-infection, and neutropenia were not associated with infection or colonization. The most common microorganisms were gram-negative rods and S.aureus. Conclusions: The duration of venous catheter use increased the risk of colonization and infection. This observation suggests that physicians must strive for the shortest time of use of venous catheters, but it does not indicate a need for routine central venous catheter removal. PMID- 11105136 TI - Calculation of HIV Infection Rates and Projection of the Number of Cases of AIDS in Sao Paulo, Brazil Using a Backcalculation Method. AB - Reconstruction of HIV infection rates and a projection of the number of AIDS cases in the City of Sao Paulo were determined using a backcalculation method. AIDS cases reported to the Centro de Vigilancia Epidemioloica, the State Surveillance Department in Sao Paulo were adjusted for reporting delays through 1993 using a non-parametric delay distribution. The incubation period distribution was derived from a multicenter cohort study in the United States and modelled in two stages: infection to CD4(+) 200/mm(3) and CD4(+) 200/mm(3) to AIDS diagnosis. After 1990, a factor reflecting the influence of treatment on the incubation distribution was used. Estimates were generated for the following risk cathegories: homosexual/bisexual men, heterosexuals, and IV drug users (IDU). HIV infection rates peaked at approximately 10,000 new infections annually in the mid 1980s, followed by a tendency to stabilize around 4,000 new annual infections in the early 1990s. HIV infection curves varied among risk cathegories with a decrease in infections in homosexual and bisexual men, but a continuous rise in incidence rates estimated for heterosexuals. By projecting AIDS incidence rates a slight upward trend was seen with approximately 4,600 new cases expected in 1997. It is notable that over 4,400 cases would be expected in 1997, even if no new HIV infections occurred. Backcalculation methods are expected to be useful for evaluating changes in infection and disease in various risk groups, and the effects of therapy. PMID- 11105137 TI - Antimicrobial Susceptibility of Klebsiella pneumoniae Producing Extended-Spectrum beta-lactamase (ESBL) Isolated in Hospitals in Brazil. AB - The prevalence of klebsiella pneumoniae producing extended-spectrum beta lactamase (ESBL) has been increasing all over the world. Infections caused by ESBL producing isolates are difficult to detect with current susceptibility tests, and are difficult to treat. ESBLs confer resistance to all currently available beta-lactam, except carbapenems. In addition, ESBL production is usually associated with resistance to other classes of antimicrobial agents such as aminoglycosides and quinolones. The objective of this study was to evaluate the in vitro susceptibility patterns of ESBL producing K pneumoniae isolated in Brazil. Seventy-two strains were tested using E test against 30 antimicrobial agents, including carbapenems, second and third generation cephalosporins, aminoglycosides, quinolones, and some new compounds. The most active compounds (i.e. 100% susceptibility) were meropenem (MIC90, 0.125ug/mL), imipenem (MIC90, 0.25ug/mL), and cefotetan (MIC90, 2ug/mL). Ciprofloxacin (MIC90, 1ug/mL, 94% susceptibility) and cefepime (MIC90, 6ug/mL, 92% susceptibility), were also very active against our collection of ESBL producing K pneumoniae. None of the six aminoglycosides showed good activity against these strains (16% to 41% susceptibility) and only 39% of the isolates were susceptible to piperacillin/tazobactam. The results of our study indicated that the carbapenems are the most active compounds against ESBL producing L pneumoniae in Brazil, and ciprofloxacin remains very active against these strains. Cefotetan and cefepime were also very active against ESBL producing K.pneumoniaein Brazil; however, further studies are necessary to evaluate the role of these cephalosporins in the treatment of infections due to ESBL producing strains. PMID- 11105138 TI - Rhodotorula glutinis Fungemia: a Case Report and Literature Review. AB - A fatal case of Rhodotorula glutinis fungemia in an 11-year-old boy with acute lymphoblastic leukemia undergoing third reinduction chemotherapy is reported. This is the first case of fungemia by Rhodotorula glutinis reported in Brazil. PMID- 11105139 TI - The Challenge of Surveillance and Control of Emerging Communicable Diseases. PMID- 11105140 TI - Clinical Microbiology: an Essential Tool for the Infectious Diseases Clinician. PMID- 11105141 TI - C-Reactive Protein: Re-evaluation of a Diagnostic Laboratory Test. AB - C-reactive protein has been a measure of acute phase reactions to inflammation for 40 years. Recently improved quantitative assays in serum and cerebrospinal fluid (CSF) have allowed a re-evaluation of its potential as a diagnostic laboratory test. The main advances in the newer methods have been that they can provide rapid (hours) information on the hepatocyte synthesis of this molecule during immune response. We have tested its value in patients with presumed bacterial meningitis. Based on our experience, newer standardized, quantitative assessments of C-reactive protein can be very useful in distinguishing between bacterial and other forms of meningeal irritation during the first few days of hospitalization. Other investigators have indicated that by serial measurements important information on the resolution or continuation of inflammatory processes can be obtained. We recommend improved standardization of this test, and recalculation of its usefulness as a diagnostic laboratory test. PMID- 11105142 TI - Protease Inhibitors Therapy for AIDS Patients. AB - Protease inhibitors have had a significant clinical impact on treatment of previously treated and untreated patients with HIV infection. In the treatment of previously treated patients, the addition of a protease inhibitor and at least one other antiretroviral agent likely to be sensitive to that particular isolate, has resulted in significant increases in CD(4)(+) cell counts, decreases in plasma viral burden and substantial decreases in the rate of development of new clinical events and mortality. Dramatic and sustained reductions in viral burden have been noted in a majority of patients treated with such regimens. In addition to combining a protease inhibitor with one or more nucleoside analogs, pilot studies of combinations of two protease inhibitors or a protease inhibitor plus a non-nucleoside reverse transcriptase inhibitor, have also demonstrated impressive drug efficacy and tolerance. PMID- 11105143 TI - Efficacy and Tolerability of Liposomal Amphotericin B (Ambisome) in the Treatment of Visceral Leishmaniasis in Brazil. AB - Thirty-two patients were enrolled in an open-label, dose/schedule ranging clinical trial to evaluate the efficacy and tolerability ofliposomal amphotericin B (Ambisome) in the treatment of visceral leishmaniasis. All patients received a dose of 2mg/kg daily for the first 4 days, followed by a single repeat dose of 2mg/kg at day 10 in 4 patients (total dose 10mg/kg); repeat doses on days 5, 6, and 10 in 13 patients (total dose 14mg/kg); or daily doses were continued on days 5 through 10 in 15 patients (total dose 20mg/kg). Patients had a mean age of 9 years, ranging between 3 and 26 years. Their mean weight was 25.9kg, ranging between 9.5kg and 75kg. All patients had splenomegaly, 31/32 had hepatomegaly, and 20 patients tested had leishmania documented on splenic aspirate. Six of the 32 patients were treated after relapse following antimony therapy. The duration of illness prior to therapy was a mean of 2 months, ranging between 2 weeks and 23 months. During and after treatment, there were significant reductions in liver and spleen sizes, and significant increases in body weight, hemoglobin levels and white blood cell counts. All patients showed initial cure at the 1 month follow up. Seven patients relapsed between 2 and 6 months after the start of treatment. There was no dose relationship to the occurrence of relapse. The relapse rate in children 5 years of age or less was 7/15 (47%). Associated causes of relapse were refractory disease (i.e., previous relapses) in 2, severe malnutrition in 1, and concurrent disease (meningococcal meningitis) in 1. In the other 2 cases, no associated event was observed except young age (ages 3 and 5 years). One relapsed patient was treated successfully with 14 days of lipid amphotericin B, and the others were cured by use of antimony for 20 to 30 days. There were no dose related adverse events. The most common event was fever which occurred in 13/32 patients (41%); 3/4 patients in the 10mg dose group, 7/13 in the 14mg dose group, and 3/15 in the 20mg dose group. Three patients had cardiac arrhythmia, one also with myocarditis diagnosed 2 weeks after therapy was discontinued. One patient developed hepatitis after dose 3 and the drug was discontinued. We concluded that liposomal amphotericin B is effective in a daily dose of 2mg/kg given for 5-10 doses as an initial cure, but that relapse occurs in young children, particularly those with documented treatment resistant disease or concurrent malnutrition or infection. Patients should be carefully monitored for these risk factors before and during the months alter therapy, and for the occurrence of arrhythmia, cardio pulmonary effects or hepatotoxicity. This treatment provides an important advance over previously used antimony therapy and appears to be more effective and well tolerated than non-lipid amphotericin B. PMID- 11105144 TI - Evaluation of Secnidazole Gel and Tinidazole Suspension in the Treatment of Giardiasis in Children. AB - Giardiasis is a cosmopolitan parasitosis. Diarrhea, abdominal colic, and flatulence are the main clinical symptoms, however, malabsorption, and impairment of growth of children may occur. The 5-nitroimidazoles are the drugs of choice in the treatment of giardiasis. Methods: The efficacy and tolerability of secnidazole and tinidazole were evaluated in a randomized, open-label, clinical trial performed with 267 Giardia lamblia-positive children. Secnidazole, in a new gel formulation, and tinidazole suspension were prescribed as single oral doses of 30mg/kg and 50mg/kg, respectively. Clinical and parasitological follow-up was carried out before, and at 7, 14, and 21 days after treatment. Results: Clinical cure was observed in 77.3% and 75.7% of the patients in the secnidazole and tinidazole groups, respectively. Parasitological cure was obtained in the 91.3% and 89.6% in the secnidazole and tinidazole groups, respectively. A metallic taste after drug ingestion was more commonly reported in the tinidazole group than in the secnidazole group (p<0.05). Conclusions: The authors conclude that both secnidazole gel and tinidazole administered as a single oral dose are effective treatments for children with giardiasis since both high cure rates and good tolerability were observed. PMID- 11105145 TI - Hepatitis B Vaccination of Health Care Workers is Not Yet a Reality. AB - It is well documented that health care workers (HCW) have a higher prevalence of hepatitis B markers, and a higher risk of acquiring hepatitis than the general population does. In this study, we obtained the prevalence of vaccination against hepatitis B among HCW in a tertiary hospital, evaluated the reasons why hospital personnel did not use the vaccine, and we determined the prevalence of accidents which carried the potential of infection among the professional staff. HCW at possible risk were included in the study. The investigation showed that 39.3% of HCW were completely vaccinated, 12.9% received only partial vaccination, 8.4% were in the process of a vaccination series, 36.5% were never vaccinated, and 2.8% had passive immunization against hepatitis B virus. Lack of opportunity and difficulty in obtaining the vaccine were the main reasons given for non vaccination. Of the non-vaccinated, 84.5% declared an intention to take the vaccine. Accidents with a potential for infection were observed in 57.9% of the population (sharp object accidents in 48.6%, accidents involving contact with blood or body fluids on mucous membranes in 27.6% and incision related accidents in 5.3%). The accidents were grouped into categories of risk of infection according to profession or medical specialty. Surgeons had the highest number of accidents. Strategies for vaccination campaign plans include an emphasis on the risks of exposure to the virus, discussion about the efficacy and safety of vaccination, and counselling to eliminate resistance to immunization. PMID- 11105146 TI - Detection of Herpes Virus (KSHV) DNA Sequences in Brazilian Patients With AIDS Associated Kaposi's sarcoma. AB - To explore the possible involvement of herpes virus (KSHV) in AIDS-associated Kaposi's sarcoma (KS) in 7 patients in Brazil, we analyzed 7 AIDS-KS lesions. Using PCR, we found KSHV specific sequences in 3 cases and by using nested PCR, we identified sequences in each of the 7 cases. Direct sequencing on nested-PCR products showed a certain degree of variability in relation to classic KSHV sequences, and identified alterations similar to those described in some endemic cases from Africa and in AIDS-associated KS specimens from North America. This mixed pattern of KSHV sequences observed in AIDS-associated KS from Brazil may reflect the geographic origin of the samples, consistent with the environmental and epidemiological backgrounds of people in this country. It is apparent that, just as in other countries in the world, Kaposi's sarcoma in HIV patients is related to herpes virus infection. PMID- 11105147 TI - Bone Paracoccidioidomycosis in an HIV-Positive Patient. AB - AIDS patients are vulnerable to infection by opportunistic microbes, including various fungi such as Pneumocystis carinii, Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans and many others. However, the association of AIDS and infection with Paracoccidioides brasiliensis has been rarely recorded. We report a case of an HIV-positive patient with bone infection by this fungus with a clinical form not previously published. This clinical presentation included primarily a massive bone lesion, but it did not include the lymphatic and disseminated disease described in HIV-positive patients. The patient responded well to medical and surgical treatment. We suggest that patients with moderate, rather than severe, immunosuppression may have forms of paracoccidioidomycosis with a pathologic process intermediate to those seen in the immunologically normal host and the full AIDS syndrome. PMID- 11105148 TI - Ethical Issues Regarding Randomized, Placebo-Controlled Clinical Trials And Their Publication in Medical Journals. PMID- 11105149 TI - The Lipoprotein Profile in HIV Infected Patients. AB - Infection can change plasma lipoproteins by increasing the triglycerides and decreasing the cholesterol plasma levels. This process is thought to be the result of alterations in lipoprotein metabolism produced by cytokines that mediate the immune response, including tumor necrosis factor, interleukin-1 and the interferons. The acquired immunodeficiency syndrome (AIDS) has been shown to be accompanied by increased plasma triglyceride levels and a trend toward decreased plasma cholesterol levels. Plasma low density lipoprotein (LDL) patterns are also changed by infection and patients with AIDS have increased levels of small dense LDL particles. Decreases in high density lipoprotein cholesterol (HDL-C) and apolipoprotein A-I, and an increase in lipoprotein (a) [Lp(a)] are the usual lipidic disorders during HIV infection, even in those patients with CD, lymphocyte counts above 400 cells/ mm(3). Plasma lipoproteins possess well-recognized transport functions. Certain classes of these lipoproteins can also regulate selected metabolic functions of a variety of cell types. Among these bioregulatory properties is the regulation of lymphocyte function and regulation of immune response. Thus, a number of immune functions may be significantly influenced by the lipoprotein alterations present in AIDS. Furthermore, the decreased HDL-C and increased triglycerides and Lp(a) are associated with an increased risk of myocardial infarction and some cardiovascular events have been reported in HIV positive individuals. This paper describes lipoprotein alterations during HIV infection, and evaluates their relationship to immune function and atherogenic profile. PMID- 11105150 TI - Rotavirus Infection in Brazil: Epidemiology, Immunity, and Potential Vaccination. AB - Worldwide, it is estimated that rotaviruses cause more than 125 million diarrheal episodes and nearly 1 million deaths per year among infants and young children. In Brazil, these agents have been associated with 13%-40% of cases of acute diarrhea affecting pediatric in-patients and outpatients. Longitudinal community based studies involving children followed from birth to the age of three years, yielded an average of 0.3 rotavirus-related episodes of diarrhea per child per year. While seasonality of rotavirus infections is not evident in the more northern tropical areas, a peak incidence during the driest months (May to September) has been noted in Brazil's central and southern states. All four epidemiologically important rotavirus serotypes have been identified in Brazil, with Gl and G2 accounting for about two-thirds of typed strains. The recent characterization of G- and P- genotypes has demonstrated that predominant strains are essentially the same as those most commonly identified worldwide: taken together, P[S]G1 and P[4]G2 rotaviruses account for over 50% of genotyped strains. In addition, the occurrence of unusual types, such as P[8]G5, and mixed infections, have been reported in about 10% and 20% of diarrheal cases, respectively. Outbreaks of rotavirus diarrhea have been reported in both urban and remote communities, G2 type being reported to cause severe gastroenteritis among adults and children. Among Indian populations the clinical attack rate has approached 90%. Rotavirus group C has been associated with outbreaks of diarrhea in day-care centers, yielding clinical attack rates of nearly 50%. Seroprevalence studies in several regions in Brazil indicate that at least 70% of children aged 4-5 years have antibodies to rotavirus. It has also been demonstrated in longitudinal studies that heterotypic immune response in a primary infection may be an intrinsic property of the rotavirus strain. A tetravalent rhesus-human reassortant rotavirus vaccine (4 x 10,000 pfu/dose) has been evaluated in northern Brazil with the following main results: a) the vaccine was safe with only low-grade fever on days 3-5 in 2%-3% of vaccinees after the first dose; b) an IgA antibody response occurred in 58% of vaccinees and 33% of placebo recipients; and c) the overall vaccine efficacy was 35% (p = 0.03) against any rotavirus diarrhea for the 2-year study period, but reached 57% (p = 0.008) during the first year of follow-up. In view of findings indicating strain diversity throughout Brazil, a country-wild surveillance system is urgently needed to monitor circulating rotavirus strains. PMID- 11105152 TI - Molecular Epidemiology of Lyme Disease Spirochetes Based on a Probe Complementary to Ribosomal RNA. AB - Lyme disease is caused by the spirochete, Borrelia burgdorferi, a bacteria which infects many vertebrates including humans. Borrelia have been isolated from many parts of the world, and there is interest to identify common genetic markers to improve molecular methods of diagnosis, and to aid in understanding varied manifestations of the disease. A total of 48 Borrelia burgdorferi strains, including: 38 isolated from ticks (Ixodes dammini, I. persulcatus, I. ricinus and I. pacificus), 3 from animals (dog, bird and hamster), and 7 from human clinical cases (skin, CSF, plasma and blood) from different geographic areas, were studied by DNA/DNA hybridization and rRNA gene restriction patterns by using a biotinylated pKK3535 probe (Altewegg M., Mayer L.W., 1989). The migration patterns of rRNA gene-restriction fragments after clevage by Hind III separate these strains into 5 ribotypes of Borrelia burgdorferi: Type I (38 American,2 European strains); Type II (13 American strains); Type III (3 Asian and 1 European strains); Type IV (1 European and 2 Asian strains) and Type V (1 Asian strain). The use of ribotyping has provided an additional tool to investigate the differences or common patterns which cause various Lyme disease syndromes. PMID- 11105151 TI - Comparative in vitro Activity of Meropenem Versus Other Extended-Spectrum Antimicrobial Agents Against 2,085 Clinical Isolates Tested in 13 Brazilian Centers. AB - Meropenem is a parenteral carbapenem antibacterial agent with a very broad spectrum of antibacterial activity. It is the second agent of its class to become available in Brazil. The in vitro antibacterial activity of meropenem was compared with imipenem and four other antimicrobial agents in a multicenter study. This study involved 13 clinical microbiology laboratories, 10 of which came from 8 Brazilian states. A total of 2,085 clinical isolates consecutively collected between December 1995 and March 1996 were susceptibility tested using the Etest and following the NCCLS procedures. Meropenem inhibited more than 90% of isolates of Enterobacteriaceae at 0.5 ug/mL, except for Citrobacter sp. (1 ug/ml). Generally, meropenem was slightly more active than imipenem against Gram negative organisms and its spectrum of antimicrobial activity was broader than those of all other drugs tested. Against Pseudomonas aeruginosa, meropenem (MIC50, 0.38 ug/ml) was approximately 8-fold more active than imipenem (MIC 50,3 ug/mL). Imipenem was two-to eight-fold more active than meropenem against some Gram-positive specees oxacillin, including Enterococcus faecalis (MIC 50 of 0.75 ug/mL and 2 ug/mL respectively), oxacillin-susceptible Staphylococcus aureus (MIC 50 of 0.47 &mul;g/mL and 0.094 ug/mL), oxacillin-susceptible Staphylococcus epidermidis (MIC 50 of 0.064 ug/mL and 0.5mg/mL). Against Streptococcus sp. meropenem was slightly more active than imipenem (MIC 50, 0.016 ug/mL). The results of this study may be used to guide empiric therapy in Brazil and indicates that meropenem may have an important role in the treatment of infections caused by multiresistant strains of bacteria. PMID- 11105154 TI - A Tribute to Two International Clinicians, Teachers and Field Researchers: Dr. Philip D. Marsden and Dr. Kenneth E. Mott. PMID- 11105153 TI - Cladophialophora bantiana (Previously Cladosporium trichoides): First Report of a Case in Brazil. AB - Cladophialophora bantiana (Cladosporium trichoides) is a black fungus recorded rarely as a cause of brain abscess. Only 21 cases have been reported in the literature world-wide. We describe the first case seen in Brazil. A 30 year old, previously healthy female, HIV negative, came to the hospital with a clinical diagnosis of brain tumor. After biopsy and culture of the lesion, it was found that she had an abscess due to Cladosporium trichoides. During the following five months, the patient underwent three more surgical brain interventions to totally remove the area of compromised tissue. In addition to surgery, amphotericin B, both intravenously and intrathecally, was used followed by itraconazole orally, without success. Six months after the first surgical intervention, the patient died. The worldwide experience with diagnosis and treatment of patients with this diseases is reviewed. PMID- 11105155 TI - A piece of my mind: the harm of "first, do no harm". PMID- 11105156 TI - JAMA 100 years ago: MODERN EMPIRICAL INVENTIONS.* PMID- 11105157 TI - Consensus panel recommendations for treatment of early breast cancer. PMID- 11105158 TI - Setting priorities and budgets to fight against global AIDS. PMID- 11105160 TI - From the Food and Drug Administration. PMID- 11105161 TI - From the food and drug administration: shock wave for heel pain PMID- 11105162 TI - From the food and drug administration: preventive use for ramipril PMID- 11105163 TI - From the food and drug administration: drug product problem reports PMID- 11105164 TI - Slow wave sleep and release of growth hormone. PMID- 11105165 TI - Slow wave sleep and release of growth hormone PMID- 11105166 TI - Impact of the Brady Act on homicide and suicide rates. PMID- 11105167 TI - Impact of the Brady Act on homicide and suicide rates. PMID- 11105168 TI - Impact of the Brady Act on homicide and suicide rates. PMID- 11105169 TI - Impact of the Brady Act on homicide and suicide rates. PMID- 11105171 TI - Impact of the brady act on homicide and suicide rates PMID- 11105170 TI - Impact of the Brady Act on homicide and suicide rates. PMID- 11105172 TI - The art of physical diagnosis. PMID- 11105173 TI - The art of physical diagnosis PMID- 11105174 TI - The art of physical diagnosis. PMID- 11105175 TI - Adverse effects associated with use of nevirapine in HIV postexposure prophylaxis for 2 health care workers. PMID- 11105176 TI - Adverse effects associated with use of nevirapine in HIV postexposure prophylaxis for 2 health care workers. PMID- 11105177 TI - A prospective study of back belts for prevention of back pain and injury. AB - CONTEXT: Despite scientific uncertainties about effectiveness, wearing back belts in the hopes of preventing costly and disabling low back injury in employees is becoming common in the workplace. OBJECTIVE: To evaluate the effectiveness of using back belts in reducing back injury claims and low back pain. DESIGN AND SETTING: Prospective cohort study. From April 1996 through April 1998, we identified material-handling employees in 160 new retail merchandise stores (89 required back belt use; 71 had voluntary back belt use) in 30 states (from New Hampshire to Michigan in the north and from Florida to Texas in the south); data collection ended December 1998, median follow-up was 6(1/2) months. PARTICIPANTS: A referred sample of 13,873 material handling employees provided 9377 baseline interviews and 6311 (67%) follow-up interviews; 206 (1.4%) refused baseline interview. MAIN OUTCOME MEASURES: Incidence rate of material-handling back injury workers' compensation claims and 6-month incidence rate of self-reported low back pain. RESULTS: Neither frequent back belt use nor a belt-requirement store policy was significantly associated with back injury claim rates or self-reported back pain. Rate ratios comparing back injury claims of those who reported wearing back belts usually every day and once or twice a week vs those who reported wearing belts never or once or twice a month were 1.22 (95% confidence interval [CI], 0.87-1.70) and 0.95 (95% CI, 0.56-1.59), respectively. The respective odds ratios for low back pain incidence were 0.97 (95% CI, 0.83-1.13) and 0.92 (95% CI, 0.73 1.16). CONCLUSIONS: In the largest prospective cohort study of back belt use, adjusted for multiple individual risk factors, neither frequent back belt use nor a store policy that required belt use was associated with reduced incidence of back injury claims or low back pain. JAMA. 2000;284:2727-2732. PMID- 11105178 TI - A large rubella outbreak with spread from the workplace to the community. AB - CONTEXT: Childhood vaccination has reduced rubella disease to low levels in the United States, but outbreaks continue to occur. The largest outbreak in the past 5 years occurred in Nebraska in 1999. OBJECTIVES: To examine risk factors for disease, susceptibility of the risk population, role of vaccine failure, and the need for new vaccination strategies in response to the Nebraska rubella outbreak. DESIGN, SETTING, AND PATIENTS: Investigation of 83 confirmed rubella cases occurring in Douglas County, Nebraska, between March 23 and August 24, 1999; serosurvey of 413 pregnant women in the outbreak locale between October 1998 and March 1999 (prior to outbreak) and April and November 1999 (during and after outbreak). MAIN OUTCOME MEASURES: Case characteristics, compared with that of the general county population; area childhood rubella vaccination rates; and susceptibility among pregnant women before vs during and after the outbreak. RESULTS: All 83 rubella cases were unvaccinated or had unknown vaccination status and fell into 3 groups: (1) 52 (63%) were young adults (median age, 26 years), 83% of whom were born in Latin American countries where rubella vaccination was not routine. They were either employed in meatpacking plants or were their household contacts. Attack rates in the plants were high (14.4 per 1000 vs 0. 19 per 1000 for general county population); (2) 16 (19%), including 14 children (9 of whom were aged <12 months) and 2 parents, were US-born and non-Hispanic, who acquired the disease through contacts at 2 day care facilities (attack rate, 88.1 per 1000); and (3) 15 (18%) were young adults (median age, 22 years) whose major disease risk was residence in population-dense census tracts where meatpacking related cases resided (R(2) = 0.343; P<.001); 87% of these persons were born in Latin America. Among pregnant women, susceptibility rates were 13% before the outbreak and 11% during and after the outbreak. Six (25%) of 24 susceptible women tested were seropositive for rubella IgM. Rubella vaccination rates were 90.2% for preschool children and 99.8% for school-aged children. CONCLUSIONS: A large rubella outbreak occurred among unvaccinated persons in a community with high immunity levels. Crowded working and living conditions facilitated transmission, but vaccine failure did not. Workplace vaccination could be considered to prevent similar outbreaks. JAMA. 2000;284:2733-2739. PMID- 11105179 TI - Individual cholesterol variation in response to a margarine- or butter-based diet: A study in families. AB - CONTEXT: The effectiveness of dietary modification in reducing low-density lipoprotein cholesterol (LDL-C) levels can be reliably predicted for populations, but not for individuals. OBJECTIVE: To determine whether individual variation in cholesterol response to dietary modification is a familial trait. DESIGN: Two period, outpatient crossover trial conducted from September 1997 to September 1999. SETTING AND PARTICIPANTS: Fifty-six families from the Dallas-Ft Worth, Tex, area with 2 biological parents and at least 2 children aged 5 years or older volunteered; 46 families (n = 92 adults and n = 134 children) completed the study. INTERVENTION: All families followed two 5-week dietary regimens that included individualized daily dietary prescriptions and emphasized a low saturated fat diet supplemented with specially manufactured baked goods and spreadable fat. One regimen used butter only and the other used margarine only. MAIN OUTCOME MEASURE: Mean LDL-C levels during the last 2 weeks of each dietary period. RESULTS: Margarine intake compared with butter intake lowered LDL-C levels 11% in adults (95% confidence interval [CI], 13% to 9%) and 9% in children (95% CI, 12% to 6%) (P<.001 for both adults and children). The distribution of individual responses were peaked around the mean response. For adults and children together, family membership accounted for 19% of variability in response (P =.007). In children, family membership accounted for 40% of variability in response of percent change in LDL-C levels (P =.002). Body mass index and change in cholesterol ester (CE) 18:2/18:1 ratio accounted for 26% of variation, leaving 26% still attributable to family membership. In all participants, BMI predicted response-heavier individuals had higher LDL-C levels, less excursion in CE fatty acids, and less LDL-C response to dietary change. CONCLUSIONS: Our results suggest that individual variation in response to a cholesterol-lowering diet is a familial trait. Body weight is an important modifiable factor that influences response. JAMA. 2000;284:2740-2747. PMID- 11105180 TI - Health and economic benefits of increased beta-blocker use following myocardial infarction. AB - CONTEXT: beta-blockers are underused in patients who have myocardial infarction (MI), despite the proven efficacy of these agents. New evidence indicates that beta-blockers can have benefit in patients with conditions that have been considered relative contraindications. Understanding the consequences of underuse of beta-blockers is important because of the implications for current policy debates over quality-of-care measures and Medicare prescription drug coverage. OBJECTIVE: To examine the potential health and economic impact of increased use of beta-blockers in patients who have had MI. DESIGN AND SETTING: We used the Coronary Heart Disease (CHD) Policy Model, a computer-simulation Markov model of CHD in the US population, to estimate the epidemiological impact and cost effectiveness of increased beta-blocker use from current to target levels among survivors of MI aged 35 to 84 years. Simulations included 1 cohort of MI survivors in 2000 followed up for 20 years and 20 successive annual cohorts of all first-MI survivors in 2000-2020. Mortality and morbidity from CHD were derived from published meta-analyses and recent studies. This analysis used a societal perspective. MAIN OUTCOME MEASURES: Prevented MIs, CHD mortality, life years gained, and cost per quality-adjusted life-year (QALY) gained in 2000-2020. RESULTS: Initiating beta-blocker use for all MI survivors except those with absolute contraindications in 2000 and continuing treatment for 20 years would result in 4300 fewer CHD deaths, 3500 MIs prevented, and 45,000 life-years gained compared with current use. The incremental cost per QALY gained would be $4500. If this increase in beta-blocker use were implemented in all first-MI survivors annually over 20 years, beta-blockers would save $18 million and result in 72,000 fewer CHD deaths, 62,000 MIs prevented, and 447,000 life-years gained. Sensitivity analyses demonstrated that the cost-effectiveness of beta-blocker therapy would always be less than $11,000 per QALY gained, even under unfavorable assumptions, and may even be cost saving. Restricting beta-blockers only to ideal patients (those without absolute or relative contraindications) would reduce the epidemiological impact of beta-blocker therapy by about 60%. CONCLUSIONS: Our simulation indicates that increased use of beta-blockers after MI would lead to impressive gains in health and would be potentially cost saving. JAMA. 2000;284:2748-2754. PMID- 11105181 TI - Technical appendix PMID- 11105182 TI - Electroacupuncture for control of myeloablative chemotherapy-induced emesis: A randomized controlled trial. AB - CONTEXT: High-dose chemotherapy poses considerable challenges to emesis management. Although prior studies suggest that acupuncture may reduce nausea and emesis, it is unclear whether such benefit comes from the nonspecific effects of attention and clinician-patient interaction. OBJECTIVE: To compare the effectiveness of electroacupuncture vs minimal needling and mock electrical stimulation or antiemetic medications alone in controlling emesis among patients undergoing a highly emetogenic chemotherapy regimen. DESIGN: Three-arm, parallel group, randomized controlled trial conducted from March 1996 to December 1997, with a 5-day study period and a 9-day follow-up. SETTING: Oncology center at a university medical center. PATIENTS: One hundred four women (mean age, 46 years) with high-risk breast cancer. INTERVENTIONS: Patients were randomly assigned to receive low-frequency electroacupuncture at classic antiemetic acupuncture points once daily for 5 days (n = 37); minimal needling at control points with mock electrostimulation on the same schedule (n = 33); or no adjunct needling (n = 34). All patients received concurrent triple antiemetic pharmacotherapy and high dose chemotherapy (cyclophosphamide, cisplatin, and carmustine). MAIN OUTCOME MEASURES: Total number of emesis episodes occurring during the 5-day study period and the proportion of emesis-free days, compared among the 3 groups. RESULTS: The number of emesis episodes occurring during the 5 days was lower for patients receiving electroacupuncture compared with those receiving minimal needling or pharmacotherapy alone (median number of episodes, 5, 10, and 15, respectively; P<.001). The electroacupuncture group had fewer episodes of emesis than the minimal needling group (P<.001), whereas the minimal needling group had fewer episodes of emesis than the antiemetic pharmacotherapy alone group (P =.01). The differences among groups were not significant during the 9-day follow-up period (P =.18). CONCLUSIONS: In this study of patients with breast cancer receiving high-dose chemotherapy, adjunct electroacupuncture was more effective in controlling emesis than minimal needling or antiemetic pharmacotherapy alone, although the observed effect had limited duration. JAMA. 2000;284:2755-2761. PMID- 11105183 TI - Caring for the critically ill patient. Current and projected workforce requirements for care of the critically ill and patients with pulmonary disease: can we meet the requirements of an aging population? AB - CONTEXT: Two important areas of medicine, care of the critically ill and management of pulmonary disease, are likely to be influenced by the aging of the US population. OBJECTIVE: To estimate current and future requirements for adult critical care and pulmonary medicine physicians in the United States. DESIGN, SETTING, AND PARTICIPANTS: Analysis of existing population, patient, and hospital data sets and prospective, nationally representative surveys of intensive care unit (ICU) directors (n = 393) and critical care specialists (intensivists) and pulmonary specialists (pulmonologists) (n = 421), conducted from 1996 to 1999. MAIN OUTCOME MEASURES: Influence of patient, physician, regional, hospital, and payer characteristics on current practice patterns; forecasted future supply of and demand for specialist care through 2030. Separate models for critical care and pulmonary disease. Base-case projections with sensitivity analyses to estimate the impact of future changes in training and retirement, disease prevalence and management, and health care reform initiatives. RESULTS: In 1997, intensivists provided care to 36.8% of all ICU patients. Care in the ICU was provided more commonly by intensivists in regions with high managed care penetration. The current ratio of supply to demand is forecast to remain in rough equilibrium until 2007. Subsequently, demand will grow rapidly while supply will remain near constant, yielding a shortfall of specialist hours equal to 22% of demand by 2020 and 35% by 2030, primarily because of the aging of the US population. Sensitivity analyses suggest that the spread of current health care reform initiatives will either have no effect or worsen this shortfall. A shortfall of pulmonologist time will also occur before 2007 and increase to 35% by 2020 and 46% by 2030. CONCLUSIONS: We forecast that the proportion of care provided by intensivists and pulmonologists in the United States will decrease below current standards in less than 10 years. While current health care reform initiatives and modification of existing practice patterns may temporarily forestall this problem, most anticipated effects are minor in comparison with the growing disease burden created by the aging US population. JAMA. 2000;284:2762 2770. PMID- 11105184 TI - Clinical features of and recent advances in therapy for Fabry disease. AB - Fabry disease is an X-linked recessive lysosomal storage disorder caused by a deficiency of alpha-galactosidase A. Intracellular accumulation of globotriaosylceramide, the glycolipid substrate of this enzyme, leads to severe painful neuropathy with progressive renal, cardiovascular, and cerebrovascular dysfunction and early death. Men are predominantly affected but many female carriers have similar clinical involvement, including increased risk of stroke. Physical stigmata, such as angiokeratomas in skin and mucous membranes and characteristic benign corneal abnormalities, facilitate identification of Fabry disease. The finding of a marked decreased activity of alpha-galactosidase A in white blood cells or cultured skin fibroblasts confirms the diagnosis. Treatment thus far has been symptomatic only. Etiology-based therapies are being developed that include enzyme replacement therapy, gene therapy, and substrate deprivation. Our recently completed double-blind, placebo-controlled trial of intravenous infusions of alpha-galactosidase A in patients with Fabry disease demonstrated the safety and efficacy of this treatment. JAMA. 2000;284:2771-2775. PMID- 11105185 TI - The Supreme Court and bedside rationing. PMID- 11105186 TI - Back belts in the workplace. PMID- 11105187 TI - Msjama: on the cover PMID- 11105188 TI - MSJAMA: modernizing disease management: A question of priorities. PMID- 11105189 TI - MSJAMA: improving the practice of pain management. PMID- 11105190 TI - MSJAMA: drugs and therapeutics in the age of the genome. PMID- 11105191 TI - MSJAMA: bringing gene therapy to the clinic. PMID- 11105192 TI - MSJAMA: the new white plague. PMID- 11105193 TI - The cell function analyser (CFA) - a physiological in vitro vascular model and potential alternative to animal experiments. AB - Cell-vessel wall interactions (adhesion, emigration) and cell-cell cohesion (aggregation) have been assessed primarily in animal experiments. The cell function analyser (CFA) is an in vitro vascular model, in which the three components of Virchow's triad are present in a highly standardised and variable form. The CFA permits visual and quantitative analysis of cellular adhesion, emigration and aggregation under physiologically relevant flow conditions (i.e. to the arteries and to the microcirculation). Although the method does not entail the use of a living animal or of animal tissue, as is true for animal experiments, with the CFA specimen fixation and histomorphological analysis after the experiment is possible. The efficacy of the method for platelet function testing has been verified by numerous clinical studies. The wide variability of test parameters make CFA suitable for in vitro analysis of other cell-vessel-wall mediated processes, such as inflammation, wound healing and tumour metastasis. We present: 1) a description of the CFA method and underlying hemodynamic principles, 2) a review of clinical and experimental results with platelets and, 3) the first results of convective flow-mediated leukocyte-endothelial interactions. The CFA provides an in vitro alternative to animal experiments, can be classified as a replacement method and possesses an analysis spectrum that will greatly reduce the overall need for the previous. PMID- 11105194 TI - [The ZEBET database on alternative methods to animal experiments in the Internet- a concrete contribution to the protection of animals]. AB - Up from February of the year 2000 ZEBET (German Centre for the Documentation and Validation of Alternative Methods) at the Federal Institute for Consumer Health Protection and Veterinary Medicine (BgVV) put the ZEBET-database on alternative methods to animal experiments on the Internet in English via DIMDI, the German Institute for Medical Documentation and Information (http://gripsdb.dimdi.de/engl/guieng.html). The access is free, moreover DIMDI's complete service is available to visitors of the ZEBET-database. The ZEBET database contains documents on alternatives to testing in animals, which have been carefully evaluated by ZEBET's staff according to the "3Rs"-concept established by Russel and Burch in 1959. Therefore, methods documented in the ZEBET database must meet at least one of the following criteria: "replacement" of an animal experiment by a non-animal method, "reduction" of the number of animals used, "refinement" of an experiment by minimising pain and suffering of animals. In addition, the ZEBET-database provides information on the current stage of development and validation of a method and on the acceptance for either scientific or regulatory purposes. Each document is characterised by the following criteria: the title of a method, keywords, assessment, summary and bibliographic references. To search DIMDI<Pt(II) + Pt(III), etc. take place. In the sonolysis of aqueous solutions of SDS, DBS or PEG-MS, two kinds of organic reducing radicals, R(ab) and R(py), are proposed to contribute to the reduction. Radical R(ab) is formed from the reaction of the surfactants with primary radicals such as hydroxyl radicals and hydrogen atoms originated from the sonolysis of water, and radical R(py) is formed from the direct thermal decomposition of surfactants in the interfacial region between the collapsing cavities and the bulk water. R(ab) is effective for both the reduction steps, whereas R(py) is involved only in the reduction step (1). This fact coincides with the previous reported sonochemical reduction of Pt(II) ions. Hydrogen atoms themselves scarcely participate in the reduction. The average diameter (1.0 nm) of platinum particles prepared from the system of PEG MS is smaller than those from the aqueous solution of anionic surfactant SDS (3.0 nm) and DBS (3.0 nm). PMID- 11105316 TI - Simultaneous irradiation of ultrasound and UV light. Ultrasonic acceleration of the photochemical disappearance of 4,4'-dihalogenated benzils in 1,4-dioxane. AB - Irradiation of ultrasound accelerated the rate of the photochemical disappearance of 4,4'-dihalogenated benzils in 1,4-dioxane in the order of halogen = I, Br > Cl > F. 4,4'-dimethoxy and unsubstituted benzils did not show acceleration. Possible mechanisms of the acceleration are discussed. PMID- 11105317 TI - Simple quantification of ultrasonic intensity using aqueous solution of phenolphthalein. AB - Aqueous phenolphthalein solution under sonication was investigated for use as a chemical dosimeter. The fading time of aqueous phenolphthalein solution under sonication depended on the concentration of phenolphthalein and the pH values of solutions. The fading time was correlated to the ultrasonic intensity in a reaction vessel that is estimated on the basis of decomposition of porphyrin. The relation between the fading time and the ultrasonic intensity for different frequencies is expressed by a single curve. From these results, it is indicated that aqueous solutions of phenolphthalein is useful for simple quantification of ultrasonic intensity for practical use, and one can regard it as one of the ultrasonic intensity indicators. PMID- 11105318 TI - ESR-spin trapping study on the sonochemistry of liquids in the presence of oxygen. Evidence for the superoxide radical anion formation. AB - The sonolysis of water and some organic liquids such as ethylene glycol, methanol and chloroform in the presence of oxygen, at 20 and 475 kHz ultrasound frequencies has been investigated by the ESR-spin trapping technique. 5,5 Dimethyl-1-pyrroline-N-oxide (DMPO), 3,3,5,5-tetramethylpyrroline-N-oxide (TMPO) and N-tert-butyl-alpha-phenyl nitrone (PBN) were able to trap superoxide radical anion, generated as the result of the sonication of the organic media. The addition of superoxide dismutase (SOD) resulted in a dramatic decrease of the ESR signal intensity of the superoxide radical adduct. In addition, the thermolysis of the liquids under ultrasound was shown by ESR detection of the spin adducts of the radicals formed by homolytic fragmentation. Occasionally, the nature of the detected spin adduct was dependent on the sonication time or on the frequency of the ultrasonic radiation. Experiments carried out in the presence of 2-methyl-2 nitrosopropane (MNP) resulted in the detection of radicals originating from thermal decomposition of the spin trap, showing its lability under ultrasonic radiation. PMID- 11105319 TI - Mixing time analysis of a sonochemical reactor. AB - Mixing time measurements have been carried out in a cylindrical reactor irradiated with an ultrasonic horn fitted with different size tips. Liquid phase bulk velocities induced by the vibrating horn surface have been estimated from the mixing time measurements. A relationship has been established between the mean horn surface velocities (frequency x amplitude) and the mean velocities estimated from the mixing time measurements. A correlation has been developed for the prediction of the mixing time using a method similar to that used for liquid jet mixing. This could be the first step in defining the overall flow field, the information about which can then be used to get realistic numerical solutions of the Rayleigh-Plesset equation for a travelling cavity to understand the cavity dynamics in the various parts of the ultrasonic horn reactor. PMID- 11105320 TI - Surfactant-assisted organic reactions in water. Effect of ultrasound on condensation reactions between active methylene compounds and arylaldehydes. AB - A series of quaternary ammonium salts has been tested as phase transfer agents to promote condensation reactions in an aqueous solution of sodium hydroxide in the absence of any organic solvent. Methyltrioctylammonium chloride (Aliquat 336) emerges as the most efficient catalyst. Sonication of the reaction media has a poor but positive kinetic effect. PMID- 11105321 TI - Ultrasonic irradiation effect in the impregnation-reduction process of preparing Pt/Nafion NH(4)(+) sensor. AB - A systematic study on the ultrasonic irradiation effect in the impregnation reduction (I-R) process for preparing a Pt/Nafion electrode was carried out in a flow-injection system of ammonium ion detection. Both the impregnation and the reduction stages were affected by ultrasonic irradiation which increased the sensing currents of electrodes. Moreover, the effect of ultrasonic irradiation was found more significant in the reduction process than in the impregnation process. The relationship between sensing current and power of ultrasonic irradiation was also obtained. The specific active surface area of the Pt/Nafion electrodes were evaluated by the cyclic voltametric technique. Meanwhile, the surfaces of the electrodes were characterized by XRD and SEM. PMID- 11105322 TI - Cavitation damage on metallic plate surfaces oscillating at 20 kHz. AB - Plates of aluminium (UNI 4507) of about 2.55 cm2 and 1.0 mm thick, of different shapes, vibrating in a liquid at a frequency of about 20 kHz and subjected to strong cavitation damage on their central parts exhibit, on the zone near the edge, a reduction of cavitative effect up to inhibition. We named this phenomenon, the "border effect". Evidence is reported which demonstrates that the border effect, depending on the plate velocity, is related to the angle between the plate surface and its direction displacement, that is to the value of the component of the liquid velocity tangent to the plate. PMID- 11105323 TI - Sonophotocatalytic decomposition of water using TiO(2) photocatalyst. AB - Sonophotocatalytic reaction is a photocatalytic reaction with ultrasonic irradiation or the simultaneous irradiation of ultrasound and light with a photocatalyst. The possibility of the effect of hybrid of sonochemical and photocatalytic reactions was examined. Liquid water was hardly decomposed to H(2) and O(2) by photocatalysis or sonolysis, independently. In order to decompose water, powdered TiO(2) photocatalyst suspended in distilled water should be simultaneously irradiated by light and ultrasound. This sonophotocatalytic reaction was effective on the decomposition of water to H(2) and O(2). PMID- 11105324 TI - Modelling of cavitational erosion in the area of surfaces of smooth contact. AB - A qualitative and numerical analysis of the differential equation, describing pulsation of a symmetrical cavity in the area of the contact of smooth surfaces with different curvatures, forming a slot is presented in this article. This particular model is a slot between a plane and a sphere. Qualitative reasons of high erosion activity of such a toroidal bubble are defined. The boundary values were found when solutions of the equation are either non-harmonic periodic and non-vanishing, or vanishing (i.e., the bubble bursts). The results of this article can be applied to the controlling tasks of ultrasonic cavitation processes. PMID- 11105325 TI - ["Magnesium in the cardiology"]. PMID- 11105326 TI - [Free intracellular magnesium in myocardium--measurement and physiological role- state of the art]. AB - Magnesium is an important intracellular cation which is known to participate in over 300 biochemical reactions of the cell. Most of the cell's Mg2+ is bound to intracellular structures, and only 5% of the total Mg2+ are unbound and thus biologically active. Clinical data concerning the therapeutic value of Mg2+ in various conditions like myocardial infarction are conflicting, experimental support for a specific therapy is still lacking, because values of free intracellular myocardial Mg2+ as well as behaviour of (Mg2+)i during ischemia or changes of extracellular ion-concentrations have not been elucidated as yet. Various techniques like Mg2+ sensitive dyes, nuclear magnetic resonance spectroscopy or Mg2+ selective microelectrodes have been employed, each of these techniques, however, harbours serious limitations. PMID- 11105327 TI - [Magnesium in coronary artery disease--is there evidence?]. AB - The role of magnesium in coronary artery disease has been evaluated extensively during the last three decades. The intravenous application of magnesium in acute coronary syndromes is of major importance, the beneficial effects of magnesium in acute myocardial infarction have been underlined in several studies. The promising results of LIMIT-2 could not be confirmed by the data of ISIS-4. A world-wide, multicenter trial (MAGIC) has been set up in order to evaluate the optimal patient cohort as well as the ideal dose regimen for the application of intravenous magnesium in patients with acute MI. Furthermore, magnesium is of significance in the pathomechanism of reperfusion injury and reduction of malign arrhythmia in the critical acute phase of MI, if applied intravenously. In stable coronary artery disease oral magnesium therapy has proven beneficial effects too. PMID- 11105328 TI - [Significance of magnesium in cardiac arrhythmias]. AB - Magnesium is of great importance in cardiac arrhythmias. It increases the ventricular threshold for fibrillation. Sinus node refractoriness and conduction in the AV node are both prolonged. Main indications for intravenous application of magnesium are Torsade de pointes tachycardias, digitalis toxicity induced tachyarrhythmias and multifocal atrial tachycardias. Additionally, patients with ventricular arrhythmias due to overdoses of neuroleptics or tricyclic antidepressants may profit from i.v. magnesium. Monomorphic ventricular tachycardias and ventricular arrhythmias refractory to class III antiarrhythmics have been shown to respond to i.v. magnesium. Recent publications have documented that perioperative use of magnesium can reduce the incidence of arrhythmic events on the atrial and ventricular level. Oral magnesium has been used for many years in patients with symptomatic extrasystoles. Studies show that the incidence of extrasystoles as well as patients' symptoms are reduced during oral magnesium therapy. PMID- 11105329 TI - Interrelationship of magnesium and congestive heart failure. AB - Congestive heart failure is a most arrhythmogenic disease that is responsible for many unexpected sudden deaths. Reduction of sudden cardiac death rates among patient populations receiving long-term diuretic therapy, when given potassium- and magnesium-sparing diuretics as well as magnesium supplements, substantiates the premise that maintenance of adequate magnesium status is important in the prevention and management of cardiovascular diseases that predispose to congestive heart failure. PMID- 11105330 TI - The role of magnesium as antithrombotic therapy. AB - Meta-analysis of previous relatively small clinical trials, comparing intravenous magnesium with placebo in acute myocardial infarction (AMI) patients, mainly without thrombolytic therapy, demonstrated that magnesium reduced in-hospital mortality by 19%, mainly by reducing the incidence of serious arrhythmias and left ventricular heart failure by one quarter. These findings have led us to hypothesize that magnesium treatment inhibits platelet-dependent thrombosis in patients with coronary artery disease (CAD). In a prospective, double blind, and crossover study, we have recently demonstrated that oral magnesium treatment inhibits thrombus formation measured by platelet-dependent thrombosis in stable CAD patients by 35%. This effect appears to be independent of platelet aggregation and activation, and is additive to that of aspirin. High dose of intravenous magnesium can inhibit thrombus formation and is associated with suppression of platelet aggregation. Magnesium treatment can dose-dependently inhibit a wide variety of agonists of platelet aggregation, such as thromboxane A2 and stimulate prostacyclin synthesis. The molecular basis for these effects is likely modulated via reduction of intracellular calcium mobilization. Hypomagnesemia also selectively impaired the release of nitric oxide from the coronary endothelium. We have recently demonstrated that oral magnesium treatment can improve endothelium-dependent vasodilation in CAD patients with optimal lipid values. Because nitric oxide is a potent endogenous vasodilator and inhibitor of platelet aggregation and adhesion, hypomagnesemia could promote vasoconstriction and coronary thrombosis in hypomagnesemic states. These findings suggest a potential mechanism whereby magnesium may beneficially alter outcomes in CAD patients. PMID- 11105331 TI - [From a herd of sheep to pharmacoeconomics]. PMID- 11105332 TI - External cholesteaoma and fibrous dysplasia of temporal bone. AB - Cholesteatoma is a disease that involves almost exclusively the middle ear structures and the mastoid bone. In rare cases it involves the external auditory canal. The author would like to present case report of a patient affected by external ear canal cholesteatoma associated to fibrous dysplasia of the temporal bone. The problems related to the pathogenesis and the diagnosis of the disease are presented and discussed. PMID- 11105333 TI - [Synovial sarcoma of the tongue. Case report and review of the literature]. AB - Primary synovial sarcoma of the head and neck is a rare entity, nevertheless the literature count on 80 published cases, among them 7 linguals. We contribute with another case: a synovial sarcoma of the tongue, in a 26-year-old man; having a biphasic tumor pattern, with two malignant constituents, epithelial and sarcomatous, similar to other of the same location. Laboratory tests: histochemical, immunohistochemical and electron microscopic were done in order to state the tumor's histogenesis. Our results suggest a pluripotential mesenchymal origin instead of a synovial origin, because the epithelial character of one of the tumor constituents was obviously epithelial. PMID- 11105334 TI - [Mixed tumor (pleomorphic adenoma) of head and neck. Typical and atypical patterns]. AB - Pleomorphic adenoma is a benign growth frequently encountered in major salivary glands, although minor salivary glands sometimes can be affected. In the latter non-specific clinical signs allows only the postoperative diagnosis. Nevertheless the existence of lesser salivary glands outside the oral cavity, and therefore pleomorphic adenomas, justify the specialist's need of its knowledge because of the difficulties arising in differential diagnosis. The AA. present a review of 22 cases in various sites treated in their Department in a 9 year-term. PMID- 11105335 TI - [The acoustic distortion products application in noise pathology]. AB - We study a total of 40 individuals exposed to noise in 3 forms of presentation: continuous (metal-workers), impact (hunters and policemen) and acute (bombs or fireworks). All them underwent a study with otoscopy, pure-tone audiometry, speech audiometry, tympanometry and recording of DP 2f1-f2 in the DPgram form. The results showed an early damage in DP (3 to 6 kHz). That are demonstrated by a decrease in recording in direct relation with increased audiometric threshold. Being obvious at 6 kHz among subjects exposed to continuous noise, though the smaller extent data are expressed in the records of 3 and 4 kHz, independently of noise form of exposure. PMID- 11105336 TI - [Pilomatrixoma and differential diagnosis of parotid area tumors]. AB - A case report of pilomatrixoma, benign neoplasy, originated of the skin annexes, which localisation force us to rule out a parotid tumor. Our intention is to include the pilomatrixoma among the possible differential diagnosis of calcified masses inside the growths of the parotid gland. PMID- 11105337 TI - [Epidemiologic and descriptive study covering one year of consultation in otoneurology department]. AB - The spread on Otoneurologic Units are increasing in number, because of the specific pathologic subjects it deals. But there are scanty epidemiologic studies on patients concurrent to these Unities. We report the data from a descriptive statistic pertaining to the Otoneurological Unit of our Hospital, in the wholly 1996. During this period of time 914 patients were attended, middle-aged 49.55, and the relation men/women being 0.61. The majority housewives, living in rural districts, prevailing the married women patients. The two ways of sanitary access more employed in so cases were: the otologist of the Social Security or reporting to the Urgency-Units of Hospitals. And the more complaining trouble was imbalance. Diagnosis more repeated was labyrinthine pathology due to ischemia and secondary vestibular conditions because cervical muscles contractures. We consider the profile of our patient group enhances the special risk in order to endure otoneurological pathology. And also we point out the fact that the most diagnosis are not related specially to vestibular pathology, but to other clinical entities not directly linked to an Otoneurologic dispensary. PMID- 11105338 TI - [Adaptable radial flap for oropharynx reconstructive microsurgery after oncologic resection]. AB - The free radial forearm flap has become the most common used flap in recreative head and neck. The tolerable complications rate and also the achievements rate offers one of the best choices in pharynx repair. PMID- 11105339 TI - [Warthin tumor. Report of eight cases and review of the literature]. AB - We have studied 8 patients with Warthin's tumor of the parotid gland (WT). Seven of them were men and smoked more than a packet daily. Seven were operated with conservative superficial parotidectomy and the other one with total parotidectomy with facial preservation. Two had postoperative peripheral facial paresia which disappeared with medical treatment in few days. One of the operated suffered from postoperative Frey's syndrome. PMID- 11105340 TI - [The usefulness of sucralfate in postoperative improvement of children's tonsillectomy]. AB - INTRODUCTION: Sucralfate is an effective agent in the treatment of peptic ulcer, mixing up the fibrinous exudate of duodenal ulcer to form a protective barrier promoting its healing. Similarly the deprived muscle in the tonsillar bed results protected and consequently bring down the post-surgical morbidity. OBJECTIVE: The aim of this study has been to ascertain the suitability of sucralfate in alleviating symptoms after tonsil's removal. MATERIAL AND METHODS: 205 children were included in the study and randomly postoperative treated with sucralfate. A lot of different parameters were controlled afterwards such as days with pain, taken of analgesics, return to normal diet, fever, bloody saliva, halitosis or vomits. RESULTS: There was a significant association between the use of sucralfate and less days with sore throat, less pain and analgesia required. CONCLUSIONS: The use of sucralfate means and efficient measure in order to alleviate the pain and postoperative discomfort associated with tonsils removal, being a cheap drug and without having topical after-effects. PMID- 11105341 TI - [Can the surgeon contribute to reducing postoperative pain in abdominal surgery?]. PMID- 11105342 TI - [Radiochemotherapy in anal canal epidermoid cancer]. AB - Epidermoid anal canal carcinomas are radio- and chemosensitive tumors. Within a few decades, conservative treatment has replaced mutilating surgery as standard therapeutic practice. Exclusive high-dose radiotherapy remains the standard treatment of early stages T1-T2 N0 smaller than 4 cm. Three phase III trials demonstrated the benefits of combined chemotherapy and irradiation on the local efficacy and colostomy-free survival. Mitomycin C and 5 FU were delivered on weeks 1 and 5 of the irradiation. Concomitant 5 FU and CDDP regimen and neoadjuvant chemotherapy were evaluated on phase II trials. Ongoing intensification phase III trials are described. PMID- 11105343 TI - [Laparoscopic treatment of perforated duodenal ulcers. Results of a retrospective multicentric study. French Society of Laparoscopic Surgery]. AB - STUDY AIM: The aim of this multicentric retrospective study was to report procedures, mortality and morbidity rate in a series of patients operated on for perforated duodenal ulcer with a laparoscopic approach. PATIENTS AND METHODS: Four-hundred and nineteen patients from 18 centers were included. The duration of the study was ten years (1990 to 1999). There were 299 men and 120 women aged from 19 to 98 years (mean: 48 years). The ASA scores were as follows: I (48.7%), II (31.3%), III (17.5%), IV (2.5%). The mean duration between the onset of perforation and the time of operation was 13.4 hours (range: 1-70). The surgical procedures were suture (76.7%), epiploplasty (9.9%), only irrigation of the abdominal cavity (2.7%). RESULTS: Conversion into laparotomy was performed in 10.6% of the patients. Mean operative time was 85 minutes. The morbidity and mortality rates were 13.4 and 1.4% respectively. Seventeen patients were reoperated because of fistula (n = 5), intra-abdominal abscess (n = 5), small bowel obstruction (n = 4), bleeding ulcer (n = 1), iatrogenic perforation of the gallbladder (n = 1) and small bowel (n = 1). Mean hospital stay was 8.5 days. All patients were discharged with a medical treatment of the peptic ulcer disease and in most of the cases, with antibiotics for Helicobacter pylori eradication. Six patients out of 96 with a medical history of chronic peptic ulcer underwent a vagotomy. CONCLUSION: Laparoscopic repair of perforated duodenal ulcer is a safe option providing low rates of morbidity, reoperation and mortality, and can be considered the treatment of choice. PMID- 11105344 TI - [Pancreatic intubation ifn pancreas divisum]. AB - AIM: Long-term results of endoscopic pancreatic stenting in pancreas divisum is still debated. The aim of this retrospective study was to evaluate the efficacy of dorsal duct stenting in patients presenting with acute recurrent pancreatitis. PATIENTS AND METHODS: Between 1980 and 1998, among 34 patients presenting with recurrent acute pancreatitis associated with pancreas divisum, 21 were treated by pancreatic stenting during a mean time of 11 months. There were 13 men and eight women (mean age: 50 years). RESULTS: The median follow-up was 50 (range 11-105) months. The number of patients presenting with acute pancreatitis before pancreatic stenting, at the end of stenting and at the end of the follow-up was respectively 21/21 (100%), 2/19 (10%) and 2/18 (11%) (P < 0.01). The number of patients presenting with chronic pain before stenting, at the end of stenting and at the end of the follow-up was respectively 17/21 (80%), 6/19 (31%) and 5/18 (27%) (P = 0.07). The overall morbidity rate was 8/21 patients (38%) including mainly acute pancreatitis (three cases); all but one complication were managed conservatively. CONCLUSION: In patients with pancreas divisum, dorsal duct stenting decreases the rate of recurrent acute pancreatitis but the improvement of chronic pain appears less obvious. PMID- 11105345 TI - [Percutaneous cholecystostomy for acute cholecystitis in high-risk patients]. AB - AIM OF THE STUDY: The aim of this retrospective study was to report the results of percutaneous cholecystostomy in a selected group of high-risk patients with contraindications of general anesthesia. PATIENTS AND METHODS: From October 1995 to December 1999, a percutaneous cholecystostomy was performed in 29 patients with acute cholecystitis. There were 20 women and nine men with a mean age of 80.6 years (range: 59 to 95 years). All the patients were ASA III (N = 23) or ASA IV (N = 6). Ultrasound-guided percutaneous cholecystostomy was performed in 24 cases and computed tomography-guided cholecystostomy in five cases. RESULTS: Percutaneous cholecystostomy was easily performed in 28 cases; there was one failed procedure. The drainage was not efficient in three patients who were operated on with one postoperative death of a patient who had a necrotic cholecystitis. There was no mortality in relation with cholecystostomy. One patient died at day 15 from myocardia infarction. The morbidity rate was 3.4% (one case). Postoperative length of hospital stay was 13 days (range: 7-30 days). The duration of the entire procedure ranged from 9 to 60 days (mean: 20 days). The mean follow-up of patients was 17 months (range: 4-40 months). One patient had recurrent acute cholecystitis and another one had angiocholitis; two patients underwent delayed elective laparoscopic cholecystectomy; 20 patients remained asymptomatic and 16 were still alive at the time of this study (13 with biliary stones and three without). CONCLUSION: Percutaneous cholecystostomy is a valuable alternative procedure for high-risk patients with acute cholecystitis. It's a safe and usually effective procedure without mortality and with a low morbidity. Whenever possible, percutaneous cholecystostomy should be followed by laparoscopic cholecystectomy. PMID- 11105346 TI - [Stomach adenocarcinoma. Evolution of surgical treatment in a series of 350 cases]. AB - STUDY AIM: The aim of this prospective study was to evaluate, in a series of 350 gastric adenocarcinomas, the evolution of their clinical and histological features and the evolution of their surgical management. PATIENTS AND METHODS: From 1970 to 1996, 350 patients with gastric carcinoma (cardiac cancer excluded) were operated on in the same center. Mean age was 68.8 years and the sex ratio 1:4. These patients were divided into three groups which were analysed and compared (group 1 operated on between 1970 and 1988, group 2 between 1979 and 1987, group 3 between 1988 and 1996). RESULTS: Antropyloric cancer was the most common (56% in group 3). Tumor size decreased but there were in group 3 more undifferentiated tumors (54.2%) and more tumors with lymph node involvement (72%). Stage III and IV tumors were still common (70% in group 3) and early gastric cancer incidence very low (9.9%). After 1980, surgical management was more radical, with more total gastrectomies and larger lymph node dissections. Adjuvant intraoperative and postoperative radiotherapy has been associated since 1985. Postoperative mortality rate decreased (13.8% in group 1 vs 6.1% in group 3) (P = 0.04) as did the postoperative morbidity rate (31.8% in group 1 vs 17.5% in group 3). The five-year actuarial survival rate was respectively 18.9% and 39.2% for groups 1 and 2 (P = 0.003); it was respectively 59.8% and 43.6% for all patients treated by adjuvant radiotherapy and for those with lymph node involvement. CONCLUSION: Prognosis of gastric adenocarcinoma is still poor. A more radical surgical treatment did not increase the postoperative morbidity rate, but increased the survival rate. New adjuvant treatments including radiotherapy have to be evaluated in order to improve the prognosis. PMID- 11105347 TI - [Isolated popliteal arteries: results of surgical treatment and causes of failure]. AB - Isolated popliteal artery is defined as an obstruction of a superficial femoral artery with a patent popliteal segment followed by an obstructed distal popliteal artery or a patent leg artery less than 5 cm long. PURPOSE: The aim of this retrospective study was to report the results of surgical treatment and the causes of failures. PATIENTS AND METHODS: From 1988 to 1996, 31 patients with isolated popliteal artery were operated on with femoropopliteal bypass. The age of the patients ranged from 45 to 92 years, (mean: 79 years); all had critical ischemia that threatened limb viability. All underwent preoperative arteriography and diagnosis was confirmed by intraoperative arteriography. RESULTS: In the postoperative course, there were 22 patent bypasses (68%) with minor amputation in five patients, and nine thromboses that required a major amputation in seven patients, a trans-metatarsal amputation in one, and a medical treatment in one. With a mean 37-month follow-up, seven thromboses required a major amputation in five patients, a new bypass in one and a medical treatment in one. The death rate was 34% at two years. The actuarial patency rates of the bypasses were 51% at one year, 38% at two years and 25% at five years. The limb salvage rate was identical. The patency rates were 65% at one, two and five years for venous bypasses and 38%, 13% and 0% respectively for PTFE bypasses. Statistical analysis showed two causes of failure: the absence of a run-off branch and the use of PTFE prostheses. No other statistically significant cause of failure was demonstrated among those analysed. Favourable anatomic conditions for a bypass to a leg artery were not predictive of failure of a femoro-popliteal bypass on the isolated arterial segment. CONCLUSION: Bypass to isolated popliteal artery is indicated in patients whose limb viability is jeopardized. Results may be considered as satisfactory especially if there is a run-off branch and if a venous graft is available for the bypass. PMID- 11105348 TI - [Burch laparoscopic colposuspension. Results of 30-month follow-up]. AB - Burch colpo-suspension, which is the present gold standard for treatment of stress urinary incontinence, may be performed laparoscopically. STUDY AIM: The aim of this retrospective study was to report the results of laparoscopic Burch colpo-suspension with a 30-month follow-up and to assess the reason for the unsuccessful results. PATIENTS AND METHOD: From 1990 to 1999, 118 patients (mean age: 46 years) were operated on for stress urinary incontinence with laparoscopic colpo-suspension. Urinary incontinence was classified grade 1 (6%), 2 (67%) and 3 (27%). The Burch colpo-suspension was performed through extraperitoneal approach in 51% and transperitoneal in 49%. A genital prolapse was associated in 31% of the patients and treated with sacropexy. A subtotal hysterectomy was performed in 25% of the patients and a vaginal hysterectomy in 46%. RESULTS: Global morbidity rate was 19%, including four cases of bladder injury. With a 30-month follow-up, 76/118 (64.4%) had no more urinary incontinence. Parity, age, previous pelvic surgery, detrusor instability and low urethral closure pressure were not predictive of recurrent stress urinary incontinence after treatment. Associated sacropexy was only correlated with a high risk of failure (P = 0.04). Patients with hysterectomy had significantly better results (72% vs 41.9%) (P = 0.05). Trans- and extraperitoneal techniques had similar results (P = 0.7). CONCLUSION: With a 30-month follow-up, 64.4% of the patients had satisfactory results with strictly no more stress urinary incontinence. There was no significant difference between the trans- and the extraperitoneal approach. Sacropexy was only associated with a higher rate of failure. PMID- 11105349 TI - [Horseradish peroxidase retrograde labeling of primary sensory axons in rats: a comparison between three different intraspinal injections methods]. AB - STUDY AIM: In order to improve the results of intraspinal retrograde labeling of post-ganglionic primary sensory axons by horseradish peroxidase (HRP), the authors compared three different intraspinal injection methods of this tracer into the inferior thoracic spinal cord in the rat. MATERIAL AND METHOD: 'Open field' method (group 1, N = 8); stereotactic injection, needle tip diameter = 0.72 mm (group 2, N = 8); stereotactic injection, needle tip diameter = 0.24 mm (group 3, N = 8). Histological features of the spinal injection site showed that tissue damages due to injection was more extensive and deeper than expected. HRP transported in retrograde fashion from injection site to sensory body cells located in dorsal root ganglia (DRG) was revealed by the Mesulam histochemical technique. RESULTS: The mean number of labeled neurons per DRG was 652 in group 3, 116 in group 2, and 77 in group 1. Differences were statistically significant, especially between groups 1 and 3 (P = 4.10(-16)) and groups 2 and 3 (P = 2.10( 17)). CONCLUSION: Retrograde labeling of primary sensory axons by HRP (or another axonal tracer) with fine needle stereotactic intraspinal injection may represent an alternative to anterograde labeling. This reliable and reproducible method may be useful in studies dealing with regeneration of post-ganglionic primary sensory axons. PMID- 11105350 TI - [Costal chondroma and chondrosarcoma]. AB - The aim of this study was to report two cases of chondrosarcoma located on the chest wall, in order to emphasize the difficulty encountered by the pathologist to differentiate a chondrosarcoma from a chondroma and the importance, in our opinion, of performing a large resection with wide margins in all cases. PMID- 11105351 TI - [Video-laparoscopic diagnosis and follow-up of a peritoneal tuberculosis]. AB - A 21-year-old woman suffering from abdominal pain and a fever of 39 degrees C was hospitalized. Ultrasonography and computed tomographic scan showed a large amount of ascites and one hepatic node. The serum CA 125 level was elevated. Protein Chain Reaction (PCR) searching tuberculosis antigen in ascitic fluid was normal. A diagnosis of peritoneal tuberculosis was supposed and an exploratory laparoscopic procedure performed. Peroperative observation of the ascites, with multiple sites of adhesion, and pathological examination of the hepatic nodule and peritoneum confirmed initial diagnosis. Antituberculous treatment was given for one year. A second laparoscopic procedure was performed and found no disease remaining. PMID- 11105352 TI - [Surgical treatment of anterior rectoceles in women. The transanal approach]. AB - Anterior rectocele is a herniation of the anterior rectal wall into the vagina, which may be either isolated or associated with other pelvic floor disorders. Rectocele could result in outlet obstruction with dyschezia, manual extraction of faeces and/or false incontinence. Rectocele is diagnosed clinically, and can be confirmed by defecography. Other tests may demonstrate associated causes of constipation. Symptomatic rectoceles can be treated via a transrectal route, with two or three layers of plication of the rectal wall and excision of the redundant mucosal flap. The results of transrectal repair are good: short hospital stay, no mortality, morbidity less than 5%, good short- and mid-term results in approximately 80% of cases. Selection criteria in favour of the transrectal approach have not been clearly identified. PMID- 11105353 TI - [Surgical treatment of anterior rectoceles in women. The perioneal-vaginal approach]. AB - Rectoceles are best repaired via a perineal approach. The transperineovaginal approach provides access to the outer side of the rectocele: the rectal hernia is repaired with two or three purse-string sutures and suture of the rectal fascia. Levatorplasty, performed without narrowing of the vagina, reinforces the repair and strengthens the lax pelvic floor. Unilateral sacro-spinofixation of the vagina is a useful adjunct to restore normal anatomy. Rectocele repair via a perineovaginal approach has a low morbidity rate and achieves good functional results. Concomitant sphincteroplasty may be performed in the case of symptomatic rupture of the anal sphincter, treating as well urinary incontinence or prolapse of the uterus. Surgery is indicated in symptomatic rectocele when retraining the pelvic floor by biofeedback and medical therapy have failed to relieve symptoms. There are no clear predictive factors of outcome and the patient must be informed about the risk of persisting symptoms or failure. PMID- 11105354 TI - [Percy at the battle of Eylau]. AB - The battle of Eylau was the bloodiest of the Napoleonic era: around 40,000 Russian and French victims littered the battlefield. It was icy cold. Percy, in charge of the army's health service, was horrified "at the view of corpses heaped in the snow," and organized first aid for the numerous wounded. PMID- 11105355 TI - [Stenosis of the common bile duct by migration of metallic clips]. PMID- 11105356 TI - [Gastrointestinal hemorrhage due to jejunal metastases from kidney cancer]. PMID- 11105357 TI - [Perforated diverticulitis of the transverse colon]. PMID- 11105358 TI - [Primary malignant tumors of the appendix]. PMID- 11105359 TI - [Small intestine angiosarcoma associated with a foreign body]. PMID- 11105360 TI - [Keloids. 52 cases treated at Ouagadougou]. PMID- 11105361 TI - Porphyria cutanea tarda. AB - Porphyria cutanea tarda (PCT) is a metabolic disorder of haem biosynthesis caused by decreased activity of uroporphyrinogen decarboxylase. Porphyria cutanea tarda is manifest by fragility, erosions, bullae, milia and scars on sun-exposed skin. Excess porphyrins in the skin interact with light of approximately 400 nm wavelength radiant energy, forming reactive oxygen species. Porphyria cutanea tarda is categorized as familial, acquired or toxic. Factors that may induce clinical expression of PCT in susceptible individuals include alcohol, oestrogen, iron, polyhalogenated compounds and viral infections. Porphyria cutanea tarda is associated with an increased incidence of the haemochromatosis gene. Treatments for PCT include withdrawal of aggravating factors, phlebotomy and oral antimalarial medications. PMID- 11105362 TI - Getting ahead of head lice. AB - Dermatologists are the nominal experts in the management of head lice in Australia, yet many dermatologists infrequently treat patients with this condition. Most people are managed in the community by school nurses, local council health officers, pharmacists, paediatricians or general practitioners. Only a small number will present to the dermatologist and commonly these patients will have tried a variety of treatments and failed to respond. Resistance is reported to all of the currently available insecticide treatments and this makes management of this common community-acquired infestation more involved. PMID- 11105363 TI - Genetics of alopecia areata. AB - Alopecia areata is a common disorder with a genetic predisposition where interaction with environmental factors leads to episodes of terminal hair loss. In this review article, we examine the evidence for a genetic basis to this disorder and discuss the prospects for future research into genetic susceptibility areas and the problems that are likely to be encountered in such research. PMID- 11105364 TI - Mixed immunobullous disease: is this linear IgA disease? AB - We report five patients who demonstrated clinical, histological and direct immunofluorescence (IF) features typical of linear IgA disease (LAD), but who also displayed IgG anti-basement membrane zone (BMZ) antibodies on indirect IF. The presence of circulating IgG anti-BMZ antibodies is often said to exclude the diagnosis of LAD. Unable to confidently classify these patients, we reviewed their clinical progress for unifying features. This revealed an almost universal benefit from dapsone therapy. We therefore propose that when strong linear IgA deposition is observed at the BMZ, a first line trial of dapsone is indicated, irrespective of the presence of circulating IgG. The class of antibody fixed in vivo appears to influence the clinical picture more than the class of circulating antibody. PMID- 11105365 TI - Occupational epoxy resin allergic contact dermatitis. AB - Sixteen cases of occupational contact dermatitis to epoxy resins were seen over a 5-year period. All were men. Six cases worked in the construction industry, two worked as painters, two as engineers, two as car windscreen repairers, and one each worked in a timber yard, a car yard, on a farm and as a cane-furniture salesman. Most presented with rashes on their faces (56%), hands (50%) or arms (37%). Two patients were allergic to the reactive diluent phenyl glycidyl ether, and one was allergic to the epoxy hardener isophorone diamine. The rest were allergic to the epoxy resin itself. Outcome in this series was poor because most continued to be exposed to epoxy resins in their workplace environment. PMID- 11105366 TI - Adult-onset atopic dermatitis. AB - Atopic dermatitis beginning in adult life is not mentioned in the medical literature. In a review of 2604 patients attending a contact dermatitis clinic, 243 patients (9%) were diagnosed with atopic dermatitis which began for the first time at 20 years of age or older with no contact factors present. This compares with 213 patients (8%) who had atopic dermatitis and contact dermatitis. Patients with purely atopic dermatitis had negative patch testing to relevant allergens and the diagnosis was based on a personal or family history of atopy as well as elevated IgE levels and multiple positive skin prick tests. A broad range of age of onset was found, as well as a female preponderance. The commonest sites of dermatitis were generalized involvement, dermatitis of the hands or eczema involving the face. PMID- 11105367 TI - Epidemiology of cutaneous lupus erythematosus in a tertiary referral centre in Singapore. AB - The aim of this retrospective study was to investigate the epidemiology, yield of investigations and proportion of patients who develop systemic lupus erythematosus (SLE) among the subsets of cutaneous lupus erythematosus (LE) in the Singapore Asian population. One hundred and twenty-five patients were diagnosed with cutaneous LE on clinico-pathological correlation, of which 73 had discoid lupus erythematosus (DLE), eight had subacute cutaneous LE (SCLE), 22 had acute LE lesions and the remainder had other less common forms of cutaneous LE. Histology was consistent with LE in 94.4% and suggestive in 4.8%. Direct immunofluorescence was positive in 61% of DLE, 86% of SCLE and 80% of acute LE cases. Antinuclear antibody (ANA) was present in the majority of acute LE (85%) and SCLE (88%) but only in 25% of DLE. Eight patients (11%) presenting with DLE had definite SLE at first presentation and two (2.7%) subsequently several months later. Of these patients, six had only mucocutaneous and serological criteria but two had major organ involvement. Five SCLE patients (63%) fulfilled the criteria for SLE, including two with major organ involvement. PMID- 11105368 TI - Bullous systemic lupus erythematosus. AB - A 19-year-old woman with a 6 month history of systemic lupus erythematosus (SLE) developed a widespread urticated, erythematous eruption associated with tense, fluid-filled blisters, erosions and crusting. Biopsy showed subepidermal blistering with a prominent neutrophilic infiltrate. Direct immunofluorescence showed markedly positive granular IgG deposition with weak IgM, IgA and C3 at the dermoepidermal junction. No circulating antibodies were detected on indirect immunofluorescence. A diagnosis of bullous systemic erythematosus was made. Treatment with prednisone was ineffective. Subsequent treatment with dapsone led to rapid sustained remission of skin symptoms. Bullous SLE is a rare manifestation of SLE. We review the recent literature and discuss the distinctive features of this condition and contrast them with cutaneous SLE with blisters and the subepidermal blistering disorders. PMID- 11105369 TI - Disseminated granuloma annulare following erythema multiforme minor. AB - A 44-year-old woman presented with erythema multiforme minor followed by disseminated granuloma annulare 4 weeks later. The patient was not taking any medication and had no history of herpes simplex infection. Involvement of a delayed-type hypersensitivity reaction in the pathogenesis of these two well known disorders, as suggested by immunological investigations, may explain their concurrence in our patient. The substitution of the erythema multiforme minor lesions by an eruption of disseminated granuloma annulare at the same sites suggests the possibility of a Koebner phenomenon or an isotopic response. PMID- 11105370 TI - Drug-related pemphigus and angiotensin converting enzyme inhibitors. AB - A 105-year-old woman developed pemphigus foliaceus. She had been on fosinopril, an angiotensin-converting enzyme inhibitor (ACE inhibitor) for 4 years. Anti intercellular cement substance antibodies were positive with titre > 160. She died during admission of an unrelated illness. A 57-year-old man developed pemphigus vulgaris after 11 months treatment with quinapril. At 14 months after developing pemphigus, this man continues on prednisone and azathioprine. We speculate that these are cases of ACE-inhibitor-related pemphigus and we review ACE-inhibitor-related pemphigus. PMID- 11105371 TI - Keratosis lichenoides chronica: report of a case developing after erythroderma. AB - A 66-year-old male presented with keratosis lichenoides chronica after a presumed drug-induced erythroderma. After resolution of the erythroderma, slightly scaly erythematous and violaceous papules in a reticular arrangement over the trunk and limbs developed in association with hoarseness, palmoplantar keratoderma, onycholysis and subungual keratosis. Histology from a lichenoid lesion showed pseudo-epitheliomatous hyperplasia, hyperorthokeratosis, parakeratosis, dyskeratosis, neutrophil exocytosis and focal vacuolar degeneration of the basal layer of the epidermis. There was a band-like chronic inflammatory infiltrate in the upper dermis. The skin improved with prednisone 40 mg/day for 15 days, leaving atrophic hypopigmented scars. A diagnosis of keratosis lichenoides chronica was made. PMID- 11105372 TI - Persistent head lice following multiple treatments: evidence for insecticide resistance in Pediculus humanus capitis. AB - Viable head lice were found on the scalps of two family members following multiple topical insecticide treatments. The possibility of reinfestation had been reliably excluded. Persistent infestation could be diagnosed only after cutting the hair and combing repeatedly, which allowed visualization of juvenile (nymphal) and adult lice. Insecticide-resistant headlouse infestations are probably much more common than is generally realised and may persist unnoticed, so that more aggressive approaches will be needed to eradicate these ectoparasites from individuals and communities. PMID- 11105373 TI - Nail staining from hydroquinone cream. AB - Topical hydroquinone is used in the treatment of a number of skin conditions. A 33-year-old male presented with brown discolouration of the fingernails following the application of 4% hydroquinone in sorbolene cream and 0.1% tretinoin cream to the face intermittently for 9 months. The discolouration resolved when the creams were ceased. PMID- 11105374 TI - Erythema multiforme: a case with unusual histopathological features. AB - A 14-year-old boy presented with widespread cutaneous and mucosal lesions clinically consistent with erythema multiforme. He gave a history of previous episodes of a similar eruption. Histological examination of a representative lesion showed changes consistent with erythema multiforme. It also, however, contained large numbers of eosinophils, forming a dermal interstitial infiltrate and epidermal microabscesses. The full blood examination showed a persistent eosinophilia. The appearances initially confused two experienced dermatopathologists. PMID- 11105375 TI - V-Y-S-plasty closure of circular defects. AB - Standard ellipse excision of lesions followed by direct closure results in a linear scar. Linear scar contracture may disrupt adjacent anatomy, rendering negative functional and cosmetic outcomes. This is of particular relevance when operating in the vicinity of paired or symmetrical anatomic structures. The V-Y-S plasty closure of circular defects conserves tissue and all but eliminates the need for undermining of wound edges. It is ideally suited for the closure of defects adjacent to the canthi, eyelids, eyebrows, lip commissures and nasal alae. The V-Y-S-plasty is also useful for the extremities where skin is under considerable tension. The technique and our refinements are described. PMID- 11105376 TI - Contact dermatitis to Asparagus officinalis. AB - A 53-year-old farm worker presented with a 3-year history of an occupational allergic contact dermatitis to asparagus. The dermatitis cleared quickly with courses of systemic corticosteroids but relapsed within days of further exposure to asparagus. The genera Asparagus is made up of some 300 species. It belongs to the family Liliaceae which includes tulips, onions and garlic. Asparagus contains asparagin, coniferin and the glucoside vanillin. The allergen may be a plant growth inhibitor, 1,2,3-Trithiane-5-carboxylic acid, which is present in young shoots. PMID- 11105377 TI - Warts and their treatment. 1950. PMID- 11105378 TI - Warts and their treatment. 1950. PMID- 11105379 TI - Treatment of warts. 1950. PMID- 11105380 TI - Treatment of warts at the turn of the millennium. AB - Gunpowder may no longer be a recommended treatment for warts, but most of the therapies used in 1950 are still used to some extent now. The problems, now as then, are the multiplicity of treatment options and the lack of a cure. Several treatments popular in the past decade are discussed. PMID- 11105381 TI - Acne vulgaris: its aetiology and treatment. 1951. PMID- 11105382 TI - A physician's prospect of the aetiology of acne. Narcissus not parsley. 1951. PMID- 11105383 TI - Some observations on follicular obstruction in acne. 1951. PMID- 11105384 TI - Biochemistry in relation to the aetiology of acne vulgaris. 1951. PMID- 11105385 TI - Acne vulgaris: yesterday, today and tomorrow. AB - Acne vulgaris is one of the commonest diseases known to humanity, affecting up to 98% of all adolescents. This review examines important aspects of its epidemiology, aetiology and management in Australia in the year 2000, in comparison with a symposium in the inaugural volume of the Australian Journal of Dermatology in 1951. PMID- 11105386 TI - Angiomata and their treatment. 1952. PMID- 11105387 TI - Treatment of problem cases of angiomata of infants. 1952. PMID- 11105388 TI - The physical aspects of radiation treatment of angiomata. 1952. PMID- 11105389 TI - Treatment of 'angiomas': a modern commentary. AB - The terminology and classification of vascular birthmarks has been simplified and clarified since 1951. Where treatment is indicated, this also has changed from radiotherapy being the treatment of choice to now having a range of therapeutic options including corticosteroids, alpha-interferon, alginate and hydrocolloid dressings, antibiotics, laser and surgery. Another important aspect of the management of haemangiomas is the recognition that certain patterns can be associated with other abnormalities. PMID- 11105390 TI - Identification and time dependence of quantitative trait loci for basal locomotor activity in the BXD recombinant inbred series and a B6D2 F2 intercross. AB - A complimentary two-phase strategy was used to detect and map quantitative trait loci (QTLs) associated with the basal locomotor response to a saline challenge (10 ml/kg). In phase 1, putative QTLs, significant at p < 0.01 or better, were identified by analysis of the strain means for 25 strains of the B x D recombinant inbred series. QTLs were identified on chromosomes 1, 3, 5, 9, 10, 16, and 18. Some of these QTLs were detected across the entire experimental period (0-20 min), while others were associated with specific 5-min blocks. Eighteen hundred C57BL/6J (B6) x DBA/2J (D2) F2 intercross animals were phenotyped for the basal locomotor response, and of this group, 500 to 700 individuals, pseudo-randomly selected, were used for a genomewide scan to confirm the RI-generated QTLs and to detect new QTLs. No new QTLs were detected but the QTLs on chromosome 1 were confirmed at p < 10(-5) to p < 10(-9), depending on the time interval. In addition, the QTLs on chromosomes 5 and 9 were confirmed at p < 0.001, providing a combined probability (RI + F2) which exceeds the threshold for a significant association. Two additional phenotypes which showed significant RI strain differences were examined--adaptation and thigmotaxis. Adaptation mapped to the same region of chromosome 9 and thigmotaxis to the same region of chromosome 1 as the distance-traveled QTL. Overall, the data presented here and elsewhere (Flint et al., 1995; Gershenfeld et al., 1997) illustrate that QTLs for basal activity are both robust and reliable. PMID- 11105391 TI - MPTP susceptibility in the mouse: behavioral, neurochemical, and histological analysis of gender and strain differences. AB - To investigate the impact of strain and sex in the l-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) animal model of Parkinson's disease, C57BL/6 and BALB/c mice were treated with either systemic MPTP-HCl (4 x 15 mg/kg) or saline and were examined in a number of behavioral tests. Furthermore, neostriatal and ventral striatal monoamine contents were determined, and the numbers of tyrosine hydroxylase-immunostained cells were counted in the substantia nigra and ventral tegmental area. Open-field testing showed that locomotor activity was drastically reduced as an acute effect of MPTP in both strains; however, subsequent recovery to control levels was faster in BALB/c mice than in C57BL/6. Nest building also indicated strain-dependent effects, since it was delayed only in C57BL/6 mice treated with MPTP. The other tests (grip test, pole test, rotarod, elevated plus maze), although partly sensitive for over-all strain or gender differences, turned out not to be useful to compare MPTP effects in these two strains. Neurochemically, MPTP led to more severe neostriatal dopamine depletions in C57BL/6 (-85%) than in BALB/c mice (-58%). Histologically, a loss of tyrosine hydroxylase immunoreactivity (-25%) was observed only in the substantia nigra of C57BL/6 animals. Thus, our analysis consistently showed that the C57BL/6 mouse strain is more susceptible to MPTP than the BALB/c strain. Sex differences in MPTP sensitivity were not observed in our mice. The implications of these findings for the search for genes related to susceptibility to neurodegeneration are discussed. PMID- 11105392 TI - Confirmation of contextual fear conditioning QTLs by short-term selection. AB - A short-term selection study for contextual fear conditioning was conducted as a confirmational strategy to analyze the chromosomal locations of five previously mapped contextual fear conditioning quantitative trait loci (QTLs). The founding population was a C57BL/6 (B6) x DBA/2 (D2) F2 intercross. High and low lines were selected for three generations based on contextual fear conditioning scores. Fear conditioning was quantified as the percentage of time spent in a "frozen" posture when placed back into the chamber, where a mild footshock and a tone had been paired with exposure to the context 24 h earlier. Allele frequencies of at least three SSLP DNA markers linked to each of the five QTLs were determined in each generation. As the selection progressed, the frequency of D2 alleles decreased in the low line and increased in the high line for chromosomes 1 and 16, while the opposite was observed in chromosomes 2, 3, and 10. The direction of divergence for alleles on these five chromosomes is consistent with the original QTL mapping study. Differences between the lines in D2 allele frequencies were found to be significant for markers on chromosomes 2, 3, and 16 but did not reach significance on chromosomes 1 or 10. In general, the results are in good agreement with our original fear conditioning QTL mapping study and provide further evidence that these QTLs regulate variation in contextual fear conditioning in crosses of B6 and D2 mice. PMID- 11105393 TI - Breaking through artificial selection limits of an adaptive behavior in mice and the consequences for correlated responses. AB - Previous divergent selection for nest-building behavior at 22 +/- 1 degrees C resulted in a 40-fold difference between the high and the low lines in amount of cotton used to build a nest. Correlated responses to selection indicated positive genetic correlations with body weight, nest-building at 4 +/- 1 degrees C, and litter size and negative genetic correlations with food consumption. At generation 46, the replicate high-selected (High 1 x High 2), randomly bred control (Control 1 x Control 2), and low-selected (Low 1 x Low 2) lines were crossed and the F1 showed significant heterosis for nest-building behavior. Regression of the F3 on the F2 generation gave heritability estimates of 0.16 +/- 0.10 for the high and 0.07 +/- 0.10 for the low cross, revealing a potential to break the selection limit (at least in the high direction), which had been reached at about 20 generations of selection. Indeed, renewed selection resulted in responses in both the high and the low directions of nesting, yielding realized heritabilities of 0.29 +/- 0.02 and 0.30 +/- 0.004, respectively. Replicated renewed selection, using the F3 generation as the base population, in the high direction of nesting resulted in correlated increases in nest-building at 4 +/- 1 degrees C, litter size, and food consumption. Body weight did not change. The positive correlation with food consumption is opposite in sign compared to the original selection experiment. This indicates that the evolutionary potential of a population to adapt to a changing environment not only depends on its current genetic variability in one adaptive trait, but may be constrained by genetic correlations changing over the course of selection. PMID- 11105394 TI - Variation of female preference for male coloration in the eastern mosquitofish Gambusia holbrooki. AB - The preference for melanistic males was studied in two populations of eastern mosquitofish (Gambusia holbrooki, Pisces: Poeciliidae), one from Florida and one from northern Italy. Melanism in the eastern mosquitofish is a Y--linked character, expressed in males only. Melanistic males have black spots varying in size and number. In the Florida population, melanistic males are common, whereas in the Italian population they have never been observed. Females were male deprived for at least 2 months before being tested in a dichotomous choice chamber. Italian females showed a significant preference for unpigmented males from their own population, whereas Florida females preferred melanistic males. When given the choice between males with few (< 10% of the body surface) and males with many (> 50%) black spots, Italian females preferred males with few black spots and Florida females those with many black spots. The preference of the Italian females for unpigmented males was confirmed in females reared from birth to maturity in the presence of only melanistic males. The preference of Florida females for melanistic males was also confirmed in females reared from birth to maturity in the presence of only unpigmented males. Altogether, these results demonstrate that in the eastern mosquitofish there is polymorphism in female preference and that this preference does not have an environmental basis. PMID- 11105395 TI - The heritability of personality factors in chimpanzees (Pan troglodytes). AB - Human personality and behavior genetic studies have resulted in a growing consensus that five heritable factors account for most variance in human personality. Prior research showed that chimpanzee personality is composed of a dominance-related factor and five human-like factors--Surgency, Dependability, Emotional Stability, Agreeableness, and Openness. Genetic, shared zoo, and nonshared environmental variance components of the six factors were estimated by regressing squared phenotypic differences of all possible pairs of chimpanzees onto 1 - Rij, where Rij equals the degree of relationship and a variable indicating whether the pair was housed in the same zoo. Dominance showed significant narrow-sense heritability. Shared zoo effects accounted for only a negligible proportion of the variance for all factors. PMID- 11105396 TI - Further evidence against the environmental transmission of individual differences in neuroticism from a collaborative study of 45,850 twins and relatives on two continents. AB - We examine the hypothesis that environmental transmission is a significant factor in individual differences for Neuroticism among 45,850 members of extended twin kinships from Australia (N = 20,945) and the United States (N = 24,905). To this large data set we fitted a model estimating genetic and environmental components of variance and gene-environmental covariance to examine the causes of individual differences in Neuroticism. For the combined sample we reject models including environmental transmission, shared environment, and a special twin environment in favor of more parsimonious genetic models. The best-fitting model involved only modest assortative mating, nonshared environment, and both additive and nonadditive genetic components. We conclude, first, that there is no evidence for environmental transmission as a contribution to individual differences in Neuroticism in these replicated samples, drawn from different continents, and, second, that a simple genetic structure underlies familial resemblance for the personality trait of Neuroticism. It is interesting that, despite the opportunity provided by the elaborate design and extensive power of our study, the picture revealed for the causes of individual differences in Neuroticism is little more complex than that found from earlier, simpler designs applied to smaller samples. However, this simplicity could not have been confirmed without using a highly informative design and a very large sample. PMID- 11105397 TI - Some properties of a variance components model for fine-mapping quantitative trait loci. AB - Identifying etiological variants for multifactorial traits by allelic association holds promise when many markers are available in close proximity. However, evidence for or against association at any particular marker does not provide any direct information about the influence of causal variants or the frequency of the etiologic allele(s). Recently, a variance components model of linkage and association was developed for quantitative traits which is sufficiently flexible to provide some insights into these issues. We show that this combined linkage/association model provides an estimate of the additive genetic variance of a trait that is attributable to disequilibrium between the marker and QTL. We use this estimate to construct approximate boundaries of the minimum level of disequilibrium between an observed marker and unobserved QTL and to delimit the permissible range of allele frequencies at the QTL based on available data at nearby markers. This information may facilitate fine-mapping studies of complex traits that aim to localize QTLs by assessment of association with many markers in a candidate region of interest. PMID- 11105398 TI - Stoolmiller on restriction of range in adoption studies: a comment. AB - Stoolmiller has recently presented a model featuring an inferred dimension of family quality which, he suggests, is severely truncated in adoption studies such as the Texas Adoption Project, resulting in gross underestimation of the effects of shared family environment. We discuss some potential limitations of his approach in general and as he applies it to the Texas adoption data on IQ. PMID- 11105399 TI - Developing technology applications for intervention research: a case study. AB - The REACH for TLC (telephone-linked care) project is a randomized study to assess the feasibility and usefulness of a computer-mediated telecommunications system for family caregivers of persons with Alzheimer's disease. The development of this automated system created technical challenges compounded by the participation in a national multisite research study. Issues arose due to the difference of cultures between researchers and technology developers. Strategic approaches were implemented proactively and during the course of the intervention to balance the competing demands of technology development and outcomes research. Key issues are discussed and recommendations made to assist others interested in developing technology-based applications for intervention research. PMID- 11105400 TI - Formative evaluation of a computer-assisted learning program in pharmacology for nursing students. AB - Previous literature on evaluation of computer-assisted learning (CAL) programs has generally not emphasized the importance of evaluation during the design and development phases. A tendency toward an objective model of evaluation rather than a naturalistic model has also meant that there is little consideration given to the context in which students learn. The aim of this study was to demonstrate the benefits of using a combination of objective and naturalistic models when undertaking a formative evaluation of a computer-assisted learning program. During the design and development phases, the program, Pharmacology Resource for Nurses (PRN), was evaluated using observation of student pairs, student questionnaires, and student focus group interviews to address the complex issues underlying program effectiveness. This study confirmed the importance and value of collecting a variety of evaluation data in order to produce a useful learning program for students. PMID- 11105401 TI - The informatics nurse specialist as change agent. Application of innovation diffusion theory. AB - The informatics nurse specialist (INS) is often the primary change agent in facilitating the implementation of clinical information systems (CIS) in healthcare settings. The INS has a unique understanding of the nursing issues that can affect the change process, and thus is in a key position to facilitate positive implementation outcomes. Innovation-diffusion theory is particularly useful in its application to the change agent role of the INS. With this theoretical knowledge, the INS can design CIS training interventions according to the psychological phenomena of Rogers' Innovation-Decision Process. An understanding of the decision-making process and the distribution of different rates of innovation adoption within a given population enable the INS to anticipate and address influential factors that affect the implementation process. Thus, Innovation-Diffusion Theory may be used as a powerful cognitive tool for the INS in facilitating the diffusion process and nurses' adoption of the technology in practice. PMID- 11105402 TI - The effectiveness of computer-assisted instruction in teaching nursing students about congenital heart disease. AB - This study compared the effectiveness of three instructional intervention strategies for teaching nursing students about congenital heart disease (CHD). They are: (1) computer-assisted instruction (CAI), (2) traditional class room lecture (TCL); and (3) the combination of CAI and TCL. The subjects were associate degree nursing (ADN) students enrolled in a pediatric nursing course at the University of Cincinnati Raymond Walters College. Differences between pre- and post-scores on a 20-item multiple choice test were analyzed by analysis of variance. There was a significant improvement in scores for all groups but no significant difference in improvement in scores between the CAI group and the TCL group. The CAI/TCL group showed significant improvement in scores compared with the other two groups. The researcher concluded that when teaching strategies are comparable, CAI is as effective as TCL. Used together, a significant improvement is seen in student performance than when either strategy is used alone. PMID- 11105403 TI - Electropalatographic and cephalometric assessment of tongue function in open bite and non-open bite subjects. AB - Anterior open bite (AOB) and tongue thrust swallowing are frequently associated, but the relationship between the two remains unclear. Electropalatography (EPG), which is used in speech pathology to measure dynamic tongue function for diagnostic, therapeutic, and research purposes, is a suitable technique for the investigation of this relationship. The present clinical study examined the dentofacial pattern and tongue function in AOB and non-open bite children. EPG recordings of speech and swallowing, and lateral head radiographs were obtained from eight 10-year-old boys with tongue thrust swallowing behaviour and AOB, and from eight age-matched non-open bite controls. Analysis of data from the two groups indicated that although differences were small, the open bite children displayed trends for longer face morphology and greater upper incisor proclination, less consistent production of closures during speech, a more posterior pattern of EPG contact, and relatively sparse EPG contact during swallowing. The discovery of differing patterns of contact for the /d[symbol: see text]/ and /t[symbol: see text]/ phonemes indicates that these should be included when speech is used to test for the presence of fronted tongue behaviour. PMID- 11105404 TI - Effect of a static magnetic field on orthodontic tooth movement in the rat. AB - Orthodontic tooth movement may be enhanced by the application of a magnetic field. Bone remodelling necessary for orthodontic tooth movement involves clastic cells, which are tartrate-resistant acid phosphatase (TRAP) positive and which may also be regulated by growth hormone (GH) via its receptor (GHR). The aim of this study was to determine the effect of a static magnetic field (SMF) on orthodontic tooth movement in the rat. Thirty-two male Wistar rats, 9 weeks old, were fitted with an orthodontic appliance directing a mesial force of 30 g on the left maxillary first molar. The appliance incorporated a weight (NM) or a magnet (M). The animals were killed at 1, 3, 7, or 14 days post-appliance insertion, and the maxillae processed to paraffin. Sagittal sections of the first molar were stained with haematoxylin and eosin (H&E), for TRAP activity or immunohistochemically for GHR. The percentage body weight loss/gain, magnetic flux density, tooth movement, width of the periodontal ligament (PDL), length of root resorption lacunae, and hyalinized zone were measured. TRAP and GHR-positive cells along the alveolar bone, root surface, and in the PDL space were counted. The incorporation of a SMF (100-170 Gauss) into an orthodontic appliance did not enhance tooth movement, nor greatly alter the histological appearance of the PDL during tooth movement. However significantly greater root resorption (P = 0.016), increased width of the PDL (P = 0.017) and greater TRAP activity (P = 0.001) were observed for group M at day 7 on the compression side. At day 14 no differences were observed between the appliance groups. PMID- 11105405 TI - Functional magnetic resonance imaging of temporomandibular joint disorders. AB - Fifty-eight temporomandibular joints (TMJs) from 40 patients with TMJ-related symptoms were examined by means of magnetic resonance scans with modified gradient echo sequences and a special double coil. This technique yielded a good spatial resolution of the intra-articular soft tissues, especially the articular disc and the bone structure of the TMJ. In combination with an incremental jaw opener, the disc-condyle complex was analysed in various closed and open mouth positions, depending on the clinical examination. Open mouth movement with differentiation of disc-condyle rotational and translation movement was demonstrated. Disturbances of TMJ motion showed interrupted condylar translation combined with mandibular deviation during open mouth movement (n = 8/58). Early phases of internal derangement of the TMJ with partial anterior disc displacement with (n = 12/58) or without (n = 2/58) reduction, total anterior disc displacement without reduction (n = 10/58), disc deformation (n = 10/58), disc adhesion (n = 2/58), condylar hypermobility (n = 6/58), condylar displacement (n = 8/58), and late phases of internal derangement of the TMJ with osteoarthrosis (n = 14/58) were clearly identified. Bilateral TMJ disorder was found in 72.5 per cent of the patients. By using motion-adapted, semi-dynamic magnetic resonance imaging (MRI), it is possible to improve the understanding of the complexity of TMJ movements. PMID- 11105406 TI - An evaluation of active shape models for the automatic identification of cephalometric landmarks. AB - This paper describes an evaluation of the application of active shape models to cephalometric landmarking. Permissible deformations of a template were established from a training set of hand-annotated images and the resulting model was used to fit to unseen images. An evaluation of this technique in comparison to the accuracy achieved by previous methods is presented. Sixty-three randomly selected cephalograms were tested using a drop-one-out method. On average, 13 per cent of 16 landmarks were within 1 mm, 35 per cent within 2 mm, and 74 per cent within 5 mm. It was concluded that the current implementation does not give sufficient accuracy for completely automated landmarking, but could be used as a time-saving tool to provide a first-estimate location of the landmarks. The method is also of interest because it provides a framework for a range of future improvements. PMID- 11105407 TI - Relationship between occlusion and satisfaction with dental appearance in orthodontically treated and untreated groups. A longitudinal study. AB - The aims of this study were to assess the relationship between occlusion, satisfaction with dental appearance, and self-esteem at the ages of 11 (T1) and 15 years (T2), and to study perceived treatment effects. Separate questionnaires were completed by children and their parents to determine their attitude. The dental casts of 224 children were collected at T1 and T2, and assessed by the Aesthetic Component (AC) and Dental Health Component (DHC) of the Index of Orthodontic Treatment Need (IOTN), and Peer Assessment Rating (PAR) Index. At T2, 16 children had been treated with removable and 51 with fixed appliances, while 157 were untreated. The children in the fixed appliance group had better dental aesthetics (AC) and occlusion (DHC) than those in the two other groups. Average PAR score reduction was 71.6 per cent (T1-T2) and satisfaction with own or child's dental appearance increased significantly. The untreated group showed increased malocclusions. In spite of that, the children expressed higher satisfaction with their own dental appearance at T2 than at T1, while the parents' satisfaction level was unchanged. For the total group, orthodontic concern at T1, AC at T2, and gender accounted for 18.0 per cent of the variation in the children's satisfaction with their own dental appearance. Parents' concern at T1 and AC at T2 accounted for 32.2 per cent of the variation in parents' satisfaction. Improvement in self-esteem from 11 to 15 years was not correlated with treatment changes. A gender difference was found. The answers to the questionnaire indicated that both children and parents rate pleasant aesthetics as an important factor for psychosocial well being. PMID- 11105408 TI - Effect of rapid maxillary expansion on skeletal, dental, and nasal structures: a postero-anterior cephalometric study. AB - The purpose of this study was to compare the transverse dimensions of skeletal, dental, and nasal structures of a group of patients with maxillary narrowness before and after rapid maxillary expansion (RME) with an untreated control group using postero-anterior (PA) cephalometric radiographs. The material consisted of PA cephalograms of 25 children with a posterior crossbite (mean age 13 years 4 months), and 25 age- and sex-matched controls (mean age 13 years 11 months). Both groups consisted of 20 females and five males. Thirty-four reference points were digitized using the Dentofacial Planner software program. The 17 variables studied consisted of six skeletal, four dental, and seven intra-nasal linear measurements. Student's t-tests were used to compare the differences between the groups, and the effect of RME on skeletal, dental, and nasal structures. RME produced small, but statistically significant changes in maxillary width, upper and lower molar widths, the width between upper central incisor apices, and intra nasal width. When compared with previous studies, the changes observed were similar for patients of a similar age group, but less than reported for a younger population. There is some evidence that the pattern of expansion produced by RME will vary depending on the age and maturity of the subject. PMID- 11105409 TI - The Class II Division 2 craniofacial type is associated with numerous congenital tooth anomalies. AB - The aim of the present study was to examine whether a putative relationship exists between the Class II division 2 craniofacial type and congenital anomalies of the dentition, such as missing teeth, peg-shaped laterals, transpositions, supernumerary teeth and canine impactions. Two hundred and sixty-seven untreated patients with Class II division 2 malocclusion were examined. The results show that 56.6 per cent of the patients exhibited some form of congenital tooth anomaly, 13.9 per cent agenesis of the upper lateral incisors, 7.5 per cent peg shaped upper laterals, while impacted canines were present in 33.5 per cent of the subjects. Transpositions were present in 1.1 per cent of the patients and in all cases the canine was involved. No patient exhibited a supernumerary tooth. Comparing the results of the present study with existing data on the percentage of congenital tooth anomalies in the general population, it can be concluded that Class II division 2 malocclusions are closely associated with congenital tooth anomalies. PMID- 11105410 TI - Temporomandibular dysfunction in patients treated with orthodontics in combination with orthognathic surgery. AB - Fifty-two patients with malocclusions underwent orthodontic treatment in combination with orthognathic surgery involving a Le Fort I and/or sagittal split osteotomy. Approximately 5 years after surgery, the patients were examined for signs and symptoms of temporomandibular disorders (TMD). The frequencies were found to be low in comparison with epidemiological studies in this field. The aesthetic outcome and chewing ability were improved in most patients (about 80 per cent). Some of the patients had reported recurrent and daily headaches before treatment. At examination, only two patients had reported having a headache once or twice a week, while all the others suffered from headaches less often or had no headache at all. Eighty-three per cent of the patients reported that they would be prepared to undergo the orthodontic/surgical treatment again with their present knowledge of the procedure. This study shows that orthodontic/surgical treatment of malocclusions not only has a beneficial effect on the aesthetic appearance and chewing ability, but also results in an improvement in signs and symptoms of TMD, including headaches. PMID- 11105411 TI - Craniofacial growth of immature rats following administration of vincristine and doxorubicin. AB - The aim of the present study was to investigate the possible short-term effect of two anti-neoplastic drugs, vincristine and doxorubicin, on the craniofacial skeleton in young rats. On the basis of findings from pilot experiments, one dose of 0.0375 mg/kg vincristine or 1.0 mg/kg doxorubicin was given parenterally to inbred Long-Evans/Turku rats at 10 or 30 days of age, and followed up until 30 or 50 days, respectively. Some 30-day-old rats received two additional doses of the drugs, 3 and 6 days after the first injection. Controls were given physiological saline. A total of 310 rats were used: 40 for the pilot study, 180 medicated, and 90 control animals for the experiment itself. The weights of the rats were recorded, a number of craniofacial dimensions were measured, and the neurocranial volume determined in the case of the most severely affected rats. The weight gain of the younger rats was retarded, as was that of the older rats that received repeated drug injections. Most dimensions of the craniofacial skeleton were significantly smaller in the vincristine-treated young animals, and following multiple injections of vincristine or doxorubicin also in the older ones when compared with the controls. Contrary to the general pattern, the measurements of the foramen magnum increased in the older rats, a feature associated with the decrease in brain cavity volume observed in those that received vincristine. These findings indicate that anti-neoplastic agents can have a short-term adverse effect on the craniofacial growth and that the morphological changes are differential, rather than uniform. PMID- 11105412 TI - The effect of early static loading on the in vitro shear/peel bond strength of a 'no-mix' orthodontic adhesive. AB - This study addressed the question of whether shear and tensile loads applied 15 minutes after bonding metal brackets to enamel affected the shear/peel bond strength of the adhesive. Ninety standard 0.022-inch stainless steel edgewise premolar mesh-backed brackets were bonded using a no-mix chemical-cured adhesive to 90 teeth, which had been prepared in a standardized manner. After 15 minutes three groups of 30 teeth were subjected to the following regimes: no applied load, tensile static load of 0.77 N (78 g), and shear static load of 0.77 N. After 14 days storage in 100 per cent relative humidity at 37 degrees C, the shear/peel strength of the adhesive bond was measured using a purpose built jig mounted on a universal testing machine. Shear/peel bond strengths were analysed using Weibull statistics. The Weibull moduli of the three groups indicated that the adhesive performed consistently despite early static loading. Characteristic strengths were 9.22, 9.27, and 9.05 MPa for the control, tensile, and shear groups, respectively. The findings indicate that static loads (such as tying in of archwires) can be placed on brackets 15 minutes after cementation, without a clinically significant reduction in bond strength of the tested adhesive. PMID- 11105413 TI - Medical malpractice among physicians: who will be sued and who will pay? AB - This paper examines whether a physician's future claims of medical malpractice are predictable from information on the physician's recent claims history, training credentials, practice characteristics, and demographics. Data on the medical malpractice experience of 8,733 Michigan physicians between 1980 and 1989 is analyzed. We find strong evidence of repetition over time regarding who was sued and who paid claims. The worse a physician's malpractice litigation record during 1980-1984, the worse was his record during 1985-1989. Training credentials were also highly predictive of future malpractice experience. Physicians trained at lower ranked medical schools or who went through lower-ranked residency programs faced higher odds of developing adverse malpractice records, even after controlling for their previous litigation record. Growing internet access to information on these characteristics will help inform prospective patients if they wish to avoid physicians likely to be sued and likely to make payments in the future for malpractice. PMID- 11105414 TI - Managed care, deficit financing, and aggregate health care expenditure in the United States: a cointegration analysis. AB - We applied a battery of cointegration tests comprising those of Johansen and Juselius [19], Phillips and Hansen [35], and Engle and Granger [6], to model aggregate health care expenditure using 1960-96 US data. The existence of a stable long-run economic relationship or cointegration is confirmed, in the United States, between aggregate health care expenditure and real GDP, population age distribution, managed care enrollment, number of practicing physicians, and government deficits. The evidence of cointegration among these variables, chosen on the theoretical basis of prior studies, implies that while they are individually non-stationary in levels, together they are highly correlated and move, in the long run to form an economic equilibrium relationship of US aggregate health care expenditure. More specifically, and for the first time in this line of inquiry, (i) managed care enrollment is found to be negatively associated with the level of health care spending, (ii) supply disinduced demand effects of physicians tend to moderate health expenditure, and (iii) government deficit financing is positively related to health care spending. The observed sign and magnitude of the income coefficient are consistent with health care being a luxury good. PMID- 11105415 TI - Diagnosis of MRSA with neural networks and logistic regression approach. AB - Antibiotic-resistant pathogens are increasingly prevalent in the hospitals and community. A timely and accurate diagnosis of the infection would greatly help physicians effectively treat patients. In this research we investigate the potential of using neural networks (NN) and logistic regression (LR) approach in diagnosing methicillin-resistant Staphylococcus aureus (MRSA). Receiver-Operating Characteristic (ROC) curve and the cross-validation method are used to compare the performances of both systems. We found that NN is better than the logistic regression approach, in terms of both the discriminatory power and the robustness. With modeling flexibility inherent in its techniques, NN is effective in dealing with MRSA and other classification problems involving large numbers of variables and interaction complexity. On the other hand, logistic regression in our case is slightly inferior, offers more clarity and less perplexity. It could be a method of choice when fewer variables are involved and/or justification of the results is desired. PMID- 11105416 TI - Technical efficiency and economies of diversification in health care. AB - In national health services, where there is a tendency towards a lack of resources and a continuous increase in demand, it is necessary to implement decisions that promote efficiency. In this paper we focus on potential diversification economies as a strategy to increase efficiency levels. We evaluate the change in efficiency in Catalan hospitals between 1987 and 1992, and analyse the presence of possible diversification economies in each hospital. We use Data Envelopment Analysis, which does not need information on either input or output prices. The results are that the majority of hospitals could increase their efficiency and reduce their costs by diversification to the output-mix offered. Potential productivity gains are between 29% and 46%. PMID- 11105417 TI - The impact of the Austrian hospital financing reform on hospital productivity: empirical evidence on efficiency and technology changes using a non-parametric input-based Malmquist approach. AB - The 1997 hospital financing reform has been supposed to reduce considerable inefficiencies in the provision of hospital care in Austria. This paper focuses on the changes in hospital productivity between 1994 and 1998, thus including three years before the reform and two years after the reform. Using Data Envelopment Analysis we calculated the input-based Malmquist index, which is then decomposed into indices of pure technical efficiency change, scale efficiency change and technology change. The results illustrated a considerably positive shift in technology between 1996 and 1998, whereas the intended enhancement in technical efficiency has not yet taken place. PMID- 11105418 TI - The planning of cervical cancer screening programmes in eastern Europe: is viral testing a suitable alternative to smear testing? AB - Cervical cancer screening with human papillomavirus (HPV) DNA testing has potential advantages over conventional, smear testing in that it can predict cases in which invasive cancers are more likely to develop, may be cheaper to implement and improve compliance. In areas of the world where little formalized cervical cancer screening takes place, or where health resources are limited, HPV testing has been suggested as a possible alternative for primary screening. In this paper we demonstrate the use of mathematical modelling to evaluate the effects of setting up screening programmes in Eastern Europe with HPV DNA testing as the primary screening tool and compare it with conventional smear testing. The impact of screening is measured in terms of the life years gained and the resulting resource usage and cost. We investigate several screening options with different screening intervals and age ranges for the target population. PMID- 11105419 TI - Factors predicting occult bacteremia in young children. AB - A febrile child without a definite localizing sign of infection may be in initial phase of bacteremia which unless treated would result in systemic complication. These instances are referred to as "Occult bacteremia". The common pathogens isolated in these children are Streptococcus pneumoniae, Hemophilus influenzae and Neisseria meningitidis. A hundred consecutive children in the age group of 3 36 months attending pediatric outpatient department and casualty were clinically evaluated using AIOS (acute illness observation scale) score and were subjected to complete blood counts, smear for malarial parasites, ESR and blood culture. In the 19-month study period, 4 instances of occult bacteremia were identified. Streptococcus pneumoniae was cultured in 3 cases and H. influenzae in one. A febrile and toxic child in the age group of 3-36 months has a high risk of occult bacteremia. High fever of temperature > or = 102 degrees F, ESR > or = 15 mm/hour, and total leukocyte count > or = 15,000/mm3, in a child with AIOS score of > or = 10 may be considered for more detailed investigations and early intervention with antimicrobial therapy. PMID- 11105420 TI - Allergenicity of common foods restricted in respiratory allergy. AB - Although hypersensitivity to foods is often linked to exacerbations of symptoms of respiratory allergy, no such information is available regarding the foods traditionally considered to play a probable etiological role in respiratory allergy in India, which are in fact quite different from the ones implicated in the West. The present study was undertaken to investigate whether the practice of withholding certain common foods by parents and practitioners of indigenous systems of medicine (i.e. Ayurvedic and Unani systems of medicine) in children suffering from respiratory allergy had any scientific basis or explanation as judged by modern techniques of investigation. Skin prick tests were performed on 64 children with symptoms pertaining to respiratory allergy (32 each in study and control group) using crude antigenic food extracts. Oral food challenges were administered to children to confirm or rule out allergenicity of food (s) incriminated on the basis of the clinical history and/or a positive skin test. Parental history of food restriction alone, in absence of positive skin prick test was of little value in predicting a positive response to the food challenges (1 challenge positive out of 77 based on food restriction: 1.29%). Only 27.02% and 18.75% of positive skin tests were found to be clinically significant in study and control groups respectively. Traditionally, food beliefs were upheld in only 12.5% children for immediate onset clinical reactions (with 5.31% of the foods restricted in their diet) and 9.37% children for delayed onset clinical reactions (with 3.19% of the foods restricted in their diet). The present study shows that even though food restriction is a common practice in patients with respiratory allergy in India, objective documentation of Type I reactions due to these foods cannot be obtained in a majority of such children. PMID- 11105421 TI - Ocular and systemic lesions in children with HIV. AB - The spectrum of ocular lesions in children with HIV infection is different from that seen in adults. Ocular lesions in pediatric AIDS patients have not been studied in India. We analyzed the clinical profile, demographic characteristics, ocular and systemic lesions in children with AIDS seen in a referral eye institute in India. The clinical profile and demographic features were studied and complete ocular examination was done. Systemic findings were evaluated at an AIDS care center and recorded in a precoded proforma. Out of the 218 cases of HIV infection seen at our hospital between December 1993 and October 1999, 12 (5.50%) were below 15 years of age. Seven (58.33%) were males and 5 (41.66%) were females with the mean age of 6.5 years and median age of 6.2 years. Vertical transmission was the most common mode of infection (58.33%). Seven (58.33%) of these patients had systemic infection, the most common being pulmonary tuberculosis (42.85%). Ocular lesions were found in 6 (50%) patients. The most common ocular lesions were anterior uveitis and cytomegalovirus retinitis (CMV) (33%) followed by retinal detachment (16.66%) and vitreous hemorrhage (16.66%). High prevalence of ocular lesions in pediatric AIDS patients in India in a referral eye centre was observed. The most common lesions were anterior uveitis followed by CMV retinitis. The management in such cases was often challenging in a developing country like India. PMID- 11105422 TI - Homocystinuria with congenital/developmental cataract. AB - The aim of the study is to screen patients for homocystinuria with and without cataract and analyse for homocystine and methionine. Fifty-eight samples from 29 patients, i.e., plasma and urine collected after overnight fasting were analysed by the screening test for homocystine, and paper chromatography for homocystine and methionine. Out of 29 homocystinuric patients, 24 had cataract. Only one had appreciable amounts of methionine in his serum. He also had mental retardation as expected and belongs to Type I. The other types did not have methionine but had only homocystine. There was no mental retardation or ectopia lentis. So they belonged to Types II, III or IV. As there is excess methionine in Type I, with low cystine, cataract may be due to deficiency of cysteine and reduced glutathione and might be averted by suitable therapy, i.e., high cystine-low methionine diet with B6. In other types with low methionine, cataract may be due to decreased availability of amino acids for the synthesis of lens proteins; the treatment of choice should be B12, and folate with methionine. PMID- 11105423 TI - Maternal and cord plasma selenium levels in full-term neonates. AB - Selenium is a part of the enzyme glutathione peroxidase and has an important role in the prevention of oxygen free radical injury. Hence good selenium nutrition is of special relevance to the neonate. The present study evaluated plasma selenium levels in cord plasma of 82 full term, appropriate for gestational age babies and their mothers at delivery. The plasma selenium levels in babies were 54.17 +/- 1.34 ppb which was significantly lower than 70.63 +/- 1.62 ppb seen in their mothers. Anemic mothers with a Hb < 8 g/dl had a plasma selenium level (60.74 +/- 4.57 ppb) which was lower than those with a Hb > 8 g/dl i.e. 74.19 +/- 2.17 ppb. Maternal age, parity, literacy and socio-economic status did not affect the plasma selenium levels. PMID- 11105424 TI - Managing the assessment of neonatal jaundice: importance of timing. AB - In view of the limitations in the accurate visual assessment of jaundice and its potential role as a predictive vector for serious neurologic sequelae, we propose that a universal screening of bilirubin be considered concurrent to the routine pre-discharge metabolic screening. Universal bilirubin screening in the term and near-term newborns when plotted on "Hour-specific Bilirubin Nomogram" in lieu of the usual "day-specific" value will predict the high-risk and the low-risk groups and facilitate cost-effective and individualized follow-up of those babies at risk. A percentile based bilirubin nomogram for the first week of age was constructed from hour-specific pre- and post-discharge bilirubin values of 2840 healthy term and near-term babies. The accuracy of the pre-discharge bilirubin values was determined as a predictive vector. Pre-discharge (18-72 hours age), 6.1% of the study population had bilirubin values in the high-risk zone (> 95th percentile). Of these, 39.5% remained in that zone (likelihood ratio ?LR? = 14.08). Pre-discharge, 32.1% of the study population had bilirubin values in the intermediate risk zone (40-75th percentiles). In a clinically significant minority of these babies (6.4%), the post-discharge values moved to the high-risk zone (L-R = 3.2 for the move from the upper-intermediate zone and 0.48 from the lower-intermediate zone). In the remainder 61.8% of the population who were identified to be at low risk, there was no measurable risk for significant hyperbilirubinemia (L-R = 0). The bilirubin nomogram can predict which infant is at high, intermediate, and low risk for subsequent excessive hyperbilirubinemia and allows for the individualized follow-up of these high-risk babies with particular attention to those who may need evaluation and intervention. Whereas, identification of the low risk group allows for a less intense bilirubin follow up and in whom a visual check by an experienced care-provider may suffice. PMID- 11105425 TI - Prognostic factors for persistent diarrhoea managed in a community setting. AB - Two hundred and five cases (mean age 13.4, SD 9.5) of persistent diarrhoea (PD) of 14-28 days duration, attending an urban slum clinic and treated according to standard WHO guidelines, were monitored at weekly intervals to obtain an estimate of treatment failure rates and to identify its clinical predictors. Vitamin and micronutrients (daily 2RDA) were additionally provided. Only 9 (8.2%) of 109 children with criteria for hospital care accepted in-patient care. Weight gain was considered inadequate if the daily increment between enrollment and day 7 of follow up was < 10 g at age 0-3 months, < 5 g at 4-6 months, and any weight loss for those older than 6 months. Recovery was considered delayed if diarrhoea ceased 7 days after enrollment. Overall, 28.3% cases had inadequate weight gain and 25.6% had delayed recovery. The non-breast milk calorie intake was 11.2% during infancy and 40.6% at later ages of the recommended intakes. In a logistic regression model, initial watery stool frequency greater than median (adjusted OR 2.30, p = 0.01), age < or = 6 months (adjusted OR 2.24, p = 0.04) and low consumption of micronutrient mixture (adjusted OR 2.62, p = 0.01) were associated with an increased risk of delayed recovery. In a Cox proportional hazards model for time to recovery from diarrhoea, low consumption of the micronutrient mixture and age < or = 6 months reduced the chances of recovery by 29% and 37% respectively. Low consumption of the prescribed micronutrient mixture (adjusted OR 2.21, p = 0.04), fever (adjusted OR 1.91, p = 0.05) and diarrhoea continuing beyond study day 7 (adjusted OR 2.29, p = 0.03) increased the risk of inadequate weight gain. Breast feeding status and animal milk consumption did not influence weight gain or recovery. Due to the low compliance for advised hospitalisation, approaches for care at community level itself need to be evolved. Focus should be on increasing the overall dietary intake and provision of generous but safe amount of micronutrients; our findings do not support need for routine elimination of animal milk. The efficacy of individual micronutrients needs evaluation in controlled trials. PMID- 11105426 TI - Inflammatory bowel disease. AB - Till about 3 decades ago, inflammatory bowel disease (IBD) was considered as non existent in our country. However, since that time several reports of IBD, mainly ulcerative colitis have been published. More recently, Crohn's disease is also being reported from the country. This trend of UC appearing first in a population followed by CD also appears to be true in other developing nations. A substantial increase in the rates of CD over UC in the last few decades is reported from developed nations as well. Of the other epidemiological factors, an increased risk of CD and lower risk of UC in smokers is established in adults. However, it appears that smoking increases the risk of IBD in children. The etiology of IBD remains elusive. Within the triad of genetics, immunity and antigen responsible for the development of IBD, maximum advances have been made in the field of immune aberrations and this is being exploited to treat the disease. It is well established that IBD results from a disordered immune system in the gut, in response to an unidentified antigen in a predisposed individual. The immune response is enhanced and revolves around antigen-presenting cells, CD 4 T lymphocytes and tumor necrosis factor alpha. CD results from an enhanced Th1 activity. The pathogenesis of UC is less clear but appears to be humoral. Advances in diagnostics include the availability of serology, ultrasound and nuclear scans, none of which have been tried in our setting where infectious diseases and tuberculosis is rampant. Growth failure and the importance of nutrition in IBD, especially CD, cannot be underemphasized. In many situations nutritional interventions have been used solely as a form of therapy for CD. Newer steroid molecules with minimal systemic effects are also being considered. Other treatment options highlighted are the use of immunosuppressive agents, biologic agents and role of surgery. PMID- 11105427 TI - Celiac disease--a worldwide problem. AB - Celiac disease and dermatitis herpetiformis are caused by the alcohol soluble fractions of wheat, barley, and rye. Reliable serological tests are available for both mass and risk group screening and recent epidemiological studies on celiac disease suggest that the prevalence varies between 1:100-300 in different continents. The clinical manifestations of the disease has changed in the West and the classical symptomatic cases represent only approximately 1/7th of all diagnosed cases. Symptoms such as, anemia, short stature, dental enamel defect or osteoporosis can be the only manifestations of the atypical disease. There is an increased prevalence of celiac disease in patients with autoimmune diseases. Recent data suggest that there is a correlation between the prevalence of autoimmune diseases and the number of years that an individual consumes gluten containing foods. Genetic studies revealed a high prevalence of certain HLA antigens in celiac patients, however, there is likelihood that non-HIA genes are also important in the pathomechanism. An interesting new development is the recognition of tissue transglutaminase (tTG), an enzyme that probably forms an autoantigen with gluten. It is generally accepted that antibodies to tTG are identical to the previously described antiendomysium antibodies. Whether or not tTG is responsible for the initiation of an immunoreaction against prolamines or just exacerbates the immune response is a subject of further investigations. PMID- 11105428 TI - Clinicopathological profile of Wilms' tumor. AB - The profile of renal tumors in children less than 15 years of age during the period 1991-1997 is presented. Among the 37 children with kidney tumors, 29 (78.4%) had Wilms' tumor. There was also a 20-year-old female with Wilms' tumor. The median age at presentation was 2.6 years (range 2.5 months to 20 years). 66.7% of the cases diagnosed were < or = 3 years and 90% were < or = 6 years. Five cases were under one year of age. The male to female ratio was 2:1. Twenty two cases (73.3%) were triphasic and 7 (23.3%) were biphasic. Only one case was monophasic with blastemal component. Five cases (16.7%) showed nephrogenic rests in the uninvolved renal parenchyma and one case had nephroblastomatosis. The tumor was favorable in 26 cases (86.7%) and unfavorable in 4. Fourteen cases were in-patients while 16 were outside referrals. The pathological (10 cases whose specimens were sent from other centers) and clinicopathological (13 hospitalized patients) staging showed 10 cases (43.5%) with stage 1, 4 cases (17.4%) with stage 2, and 7 cases (30.4%) with stage 3. In two cases (8.7%), there was stage 4 disease. The length of the follow-up period in the 13 hospitalized patients ranged from 7 days to 5 years 5 months (median 14 months). There was one recurrence and one death after 2 years of diagnosis. PMID- 11105429 TI - Infantile-onset leukoencephalopathy with discrepant mild clinical course. AB - Four children characterised by megalencephaly and cerebral leukoencephalopathy with infantile onset, defined on the basis of clinical and neuroimaging findings are reported. The course of the disease is characterised by stabilization of the macrocephaly and slow clinical deterioration. The CT scan findings include supratentorial diffuse hypodensities in the white matter and swelling. The characteristic MRI findings include the discrepant severity in comparison with the clinical picture, diffuse supratentorial white matter abnormalities with subcortical cysts. The basic defect of the disease is unknown. Considering the high rate of consanguinity among the parents and the presence of two affected sibs in one family, an autosomal recessive inheritance is assumed. We report four unrelated cases of this entity. PMID- 11105430 TI - Fraser-cryptophthalmos syndrome. AB - Fraser or Cryptophthalmos syndrome is a variable syndrome to the extent that cryptophthalmos might not be present in all cases. However, the main features are a "hidden eye", other craniofacial abnormalities, renal abnormalities, syndactyly and abnormal genitalia. It may be classified as isolated cryptophthalmos or cryptophthalmos sequence and cryptophthalmos syndrome. The cryptophthalmos syndrome has an autosomal recessive mode of inheritance. Isolated cryptophthalmos has been reported as an autosomal dominant trait. Prenatal diagnosis is possible using ultrasonography and fetoscopy. We report three cases of cryptophthalmos. One with renal agenesis had cryptophthalmos syndrome and the other two had isolated cryptophthalmos or cryptophthalmos sequence. PMID- 11105431 TI - Cystic hygroma of the gluteal region. AB - Cystic hygromas occur most commonly in the neck. Rarely are they known to involve the axilla, groin, mediastinum, retroperitoneum, pelvis, mesentery, omentum and spleen. We successfully managed a case of cystic hygroma of gluteal region in a one and half year old child who presented with a cystic, non transilluminant swelling in this region since birth. The diagnosis of cystic hygroma was made by surgery and subsequently confirmed after histopathological examination. Because of rarity of cystic hygroma in gluteal region this case in being reported. PMID- 11105432 TI - Balantidiasis in an infant. PMID- 11105433 TI - Comments on congenital methemoglobinemia. PMID- 11105435 TI - SIDS--a defect in circulatory control. PMID- 11105434 TI - Meningococcus C vaccination programme. PMID- 11105436 TI - Water fluoridation: pollutant or panacea? PMID- 11105437 TI - Safe disposal of waste, not a waste of resources. PMID- 11105438 TI - Psychosocial and cancer support services in Ireland. PMID- 11105439 TI - A study of the knowledge that school rugby coaches have in the management and prevention of serious neck injury. AB - The following study is an analysis of the knowledge school rugby coaches responsible for Senior Cup Team in Leinster, have in the area of neck injury prevention and management. When serious neck injuries affect schoolboys it is particularly tragic and deserves special attention. The study targeted all the coaches of Senior Cup Teams in Leinster. Results showed that coaches lack vital knowledge in the area of neck injury prevention, recognition and management. Only 50% (n = 18) of coaches have a first aid qualification and only 47% (n = 17) carry first aid equipment to deal with neck injuries to matches. More than half of the schools in the study sample do not have neck collars available and a staggering 83% (n = 30) of schools do not have a stretcher available to SCT players. The study highlights the need to provide better first aid facilities in schools and demonstrates the need for further education of school rugby coaches. PMID- 11105440 TI - Does maternal smoking increase the risk of neonatal polycythaemia? AB - The objective of this observational study was to determine the relationship between tobacco smoking during pregnancy and neonatal Polycythaemia, and to assess the dose-response relationship. Thirty two pregnant women who smoked tobacco (cases), and ninety pregnant women who did not smoke (controls), were randomly selected from the annual obstetrics population in the Erinville hospital in Cork. This study was carried out over eighteen months and the subjects were seen three times, at 28, 32, and 36 weeks gestation. At each visit, a smokalyser test was preformed and the results were recorded. The subjects were also given charts to fill in the number of cigarettes they smoked each day for the four week period. Nicotine consumption milligrams per day was calculated depending on the brand they smoked. Finally, at labour, cord blood samples were obtained and sent for haemoglobin and haematocrit estimation. At the end of the study it was found that both cord blood haemoglobin and haematocrit were statistically significantly higher in smoking mothers, p < 0.01 and p < 0.001 respectively. The dose-response relationship was also statistically significant. PMID- 11105441 TI - Cervical cytology history in Irish doctors and midwives. AB - In Ireland we do not have a national cervical screening programme. It is hoped that such a service will soon be implemented and a pilot programme is currently underway. In our department there was a perception that many women working in the area of women's health had a "laissez-faire" attitude towards their own cervical cytology. This prompted us to perform an anonymous survey of a sample of Irish gynaecologists, general practitioners and midwives in advance of a national cervical screening programme. Overall, 80% of the questionnaires were returned completed--72% from gynaecologists, 68% from GPs and 100% from midwives. 60% of those who returned completed questionnaires were "up-to-date", 21% were "late" and 19% had never had a cervical smear. PMID- 11105442 TI - Establishing a mobile coronary care service in a rural setting. AB - A mobile coronary care unit (MCCU) service was introduced at Sligo General Hospital in 1992. Initially, a medical registrar and CCU staff nurse drove a fully equipped MCCU car to the patient location (Phase 1). When the patient was medically stabilised and had received thrombolysis, if appropriate, the doctor and nurse travelled with him in the conventional ambulance to the hospital. Subsequently, on a pilot basis, the doctor, nurse and equipment were transported by taxi to the patient (Phase 2). Since 1995, the MCCU service has been provided routinely by Sligo General Hospital (Phase 3). During each Phase the proportion of patients with acute myocardial infarction (AMI) was as follows: Phase 1--16 of 35 (46%), Phase 2--7 of 19 (36%), Phase 3--53 of 154 (34%). The incidence of unstable angina (UA) was 25%, 32% and 26% during each of the three Phases. Five patients had successful out of hospital (OOH) defibrillation and 56(74%) of the 76 AMI patients had OOH thrombolysis. The MCCU reduced the median "Call to needle" time by 64 minutes in Phase 1 and 102 minutes in Phase 2 compared to the conventional service. Median delay to MCCU treatment was 46, 40 and 80 minutes during each of the three Phases. In an Irish setting, a MCCU service provided rapid safe and effective care, including thrombolysis and defibrillation for patients with suspected AMI. PMID- 11105443 TI - Leptospire-induced acute rhabdomyolysis in a patient with previously undiagnosed mitochondrial myopathy. PMID- 11105444 TI - Femoral hernia in children. PMID- 11105445 TI - Phenomenology of panic attacks reflects human evolutionary history of separation anxiety. PMID- 11105446 TI - Phencyclidine, ketamine, and khat phencyclidine (PCP, DOA, 'angel dust', 'crystal', 'hog') PMID- 11105447 TI - Euthanasia and assisted suicide. PMID- 11105448 TI - Hip replacement. PMID- 11105449 TI - Individualizing treatment plan and combining approaches key to pain management and holistic care: a student's perspective. PMID- 11105450 TI - Objectivity and accuracy of mammogram interpretation using the BI-RADS final assessment categories in 40- to 49-year-old women. AB - To determine if use of the five final assessment categories of the American College of Radiology's Breast Imaging Reporting and Data System (BI-RADS) improved objectivity or accuracy of mammographic evaluation in 40- to 49-year-old women, fifty mammograms of 40- to 49-year-old women that were obtained at a tertiary referral teaching hospital were classified according to those five final assessment categories. The mammograms were blinded to six American Osteopathic Board of Radiology-certified radiologists who were asked to classify each mammogram within the five final BI-RADS categories based on the mediolateral oblique and craniocaudal views presented. No history was allowed. Use of the BI RADS five final assessment categories provided moderate interobserver objectivity, moderately high agreement among the radiologists' interpretation (reliability), and moderate accuracy of interpretation (validity) when compared to criterion. Moderate interobserver reliability and accuracy has been previously identified; however, no scientific review of the BI-RADS five final assessment categories in 40- to 49-year-old females was discovered in the current literature. No overall improvement of objectivity or accuracy was demonstrated using the five final assessment categories of the BI-RADS lexicon in 40- to 49 year-old women. PMID- 11105451 TI - Glycation as the glucose link to diabetic complications. AB - Hyperglycemia is considered a key causal factor in the development of diabetic vascular complications and can mediate its adverse effects through multiple pathways. This was confirmed for microangiopathy in the Diabetes Control and Complications Trial study for type 1 diabetes and corroborated for type 2 diabetes by the United Kingdom Prospective Diabetes Study published in 1998. Prevention of diabetic complications requires at least control of glycemia. This article briefly summarizes the evidence that strongly supports the role of hyperglycemia in vascular complications. After outlining the role of the polyol pathway, protein kinase C, and oxidative stress, the author focuses on one of the key biochemical mechanisms for this pathologic process: the direct deleterious action of glucose and other sugars on proteins, known as glycation or nonenzymatic glycosylation. Results of animal studies and phase III clinical trials reveal that the inhibition of this process attenuates the development of a range of these complications. PMID- 11105452 TI - Galbreath technique: a manipulative treatment for otitis media revisited. AB - Otitis media is a common disorder that results in numerous visits to the physician each year. Antimicrobials, antihistamines, steroids, and surgery have all been used to treat otitis media; however, the literature makes little mention of osteopathic manipulative treatment in this regard. This article describes a technique that was first described in 1929 by William Otis Galbreath, DO. By simple mandibular manipulation, the eustachian tube is made to open and close in a "pumping action" that allows the ear to drain accumulated fluid more effectively. Physicians can easily teach this procedure to parents for use at home. PMID- 11105453 TI - Spirituality in history taking. AB - Andrew Taylor Still, MD, DO, included in his founding postulates of osteopathy the concept that a patient's health includes the health of a patient's spirit. In the recent past, medicine as a whole, and osteopathic medicine specifically, has neglected this postulate. Recent research has confirmed the validity of Still's postulate, and many medical training institutions have received grants and established programs to incorporate spirituality into their curriculum. As with any patient evaluation, the history and physical examination is the starting platform. This article describes several tools that can be easily incorporated into the history and physical examination, along with some of the obstacles in evaluating the health of the patient's spirit. PMID- 11105454 TI - Influencing the vegetative nervous system through manipulation. 1945. PMID- 11105455 TI - The autonomic nervous system in osteopathic therapy. 1948. PMID- 11105456 TI - Update: the latest on hormone replacement therapy. AB - Hormone replacement therapy in menopausal women offers many potential health benefits. Only 15% of patients who are eligible for therapy continue treatment past 3 months. Fear and misinformation prevent them from initiating therapy, and side effects prompt them to discontinue taking their medication. This article reviews menopause and hormone replacement with emphasis on the risks, benefits, and hazards of therapy. New options for treatment are explored. PMID- 11105457 TI - Clinical use of selective estrogen receptor modulators. AB - Selective estrogen receptor modulators (SERMs) are an exciting category of drugs for physicians providing healthcare to women. This article explores their pharmacology, effects on target organs and clinical effectiveness. PMID- 11105458 TI - Osteoporosis. AB - Human bone thickness is genetically determined and influenced by lifestyle. Deterioration of bones affects the entire body's ability to function. Initially, osteoporosis is asymptomatic but may lead to bone fractures, altered mobility, and decreased functional ability. Identifying individuals at high risk for problems caused by low bone mass and evaluating bone density are key factors in fighting this major health problem. This article reviews the tests used to diagnose and follow the progression of the disease. It also discusses the benefits of vitamin D and calcium supplementation in bone formation and preservation, as well as treatment and prevention of osteoporosis. PMID- 11105459 TI - Managing common problems in perimenopausal women. AB - The perimenopause can be a time of many changes in the lives of women. Uncomfortable, nuisance, and sometimes serious symptoms and medical problems often accompany it. Some of these problems include concerns about fertility or contraception (or both), hot flashes, sleep disturbances, urogenital changes, and abnormal uterine bleeding. Common treatment options for perimenopausal problems include hormone replacement therapy with estrogen and progestins and surgical modes of therapy. In addition, many women will notice a change in their health during the perimenopausal years and a strategy for screening and health maintenance needs to be established. PMID- 11105460 TI - Vascular medicine and osteopathic medicine: treating the whole patient. AB - Education and instruction in the care of the patient with peripheral vascular diseases is, at best, fragmented during the first years of medical training. Attention to the issues of peripheral arterial, venous, and lymphatic disorders deserves a more formal approach with respect to physician education, patient evaluation and treatment, knowledge and application of various diagnostic modalities, and involvement of our physician colleagues in complementary disciplines. The vascular medicine internist is an invaluable resource in these areas. The aging of our general population will lead to an increase in manifest peripheral vascular disease within our patient population. Having received additional comprehensive training in the management of the complex patient with peripheral vascular disease, the vascular medicine internist may serve as a complete resource for their care. PMID- 11105461 TI - Hypertension and renal artery stenosis: a complex clinical scenario. AB - Hypertension remains the most common reason for patients to visit physicians in the United States. Although awareness of hypertension among patients continues to increase, adequate control of hypertension remains poor. In addition, as the population of patients with hypertension ages, atherosclerosis becomes increasingly prevalent. Atherosclerotic renal artery stenosis is the most common secondary cause of hypertension and can cause hypertension to be difficult to control. Atherosclerotic renal artery stenosis may also result in chronic renal insufficiency. The physician must be aware of the clinical scenarios in which renal artery stenosis may occur, methods of diagnosis, and indications for intervention. PMID- 11105462 TI - Diagnosis and medical management of patients with intermittent claudication. AB - Intermittent claudication is a symptom complex associated with atherosclerosis of the aorta and lower extremities. It is a clinical marker of systemic atherosclerosis, and therefore, management cannot be considered isolated from treatment of underlying risk factors of atherosclerosis. The focus of the management is twofold. The first is to reduce morbidity and mortality from cardiovascular events, including myocardial infarction and stroke. The second focus is to improve the functional status of patients who have impairment of daily activities secondary to symptoms of claudication through pharmacologic and rehabilitative means, that is, exercise. Exercise is the cornerstone of therapy. A conservative approach is favored in patients who have mild and moderate symptoms of claudication. Intervention with percutaneous techniques or surgery is generally reserved for patients who have severe impairment of lifestyle or threatened tissue. PMID- 11105463 TI - Endovascular treatment of peripheral arterial disease. AB - Atherosclerosis is a systemic disease affecting quality and length of life. Endovascular revascularization can be used to improve quality of life. The benefit is greatest in patients with subclavian, renal, or iliac artery symptomatic disease. The advent of stents improves the initial technical success rates of angioplasty to more than 90% in most locations. The development of stent: grafts has altered the treatment of abdominal aortic aneurysms and should be strongly considered as an alternative to open surgery. PMID- 11105464 TI - Cognitive and linguistic correlates of children's discourse after closed head injury: a three-year follow-up. AB - The discourse of 91 children who had sustained severe (n = 68) or mild (n = 23) closed head injury (CHI) was examined at least three years postinjury. The groups' retellings of a narrative story were analyzed according to two domains, information and language. In comparison to the mild CHI group, the severe group produced stories characterized by reduced content and information, impaired organization, fewer words, and less complex sentences. The relationships between discourse production and the groups' performance on measures of language, executive function, memory, and processing speed were examined. Correlations were found between discourse production and general verbal ability including verbal fluency. Correlations were also found for discourse performance and executive function measures associated with problem solving and working memory. Site and extent of lesion were not useful in predicting discourse production. These findings indicate that children who sustain a severe closed head injury during early to middle childhood are at risk for persisting deficits in discourse processing and other cognitive abilities. PMID- 11105465 TI - Evidence for a deficit in procedural learning in children and adolescents with autism: implications for cerebellar contribution. AB - To examine the hypothesis that abnormalities in those cognitive functions for which cerebellar components have been implicated contribute to the pathophysiology of autism, tests of judgment of explicit time intervals and procedural learning were administered to 11 participants with autism and 17 age and-IQ-matched controls. Results indicated that the group with autism demonstrated significant impairments in procedural learning compared with the group of controls. No significant difference in judgment of explicit time intervals was found. The data suggest that deficits in procedural learning may contribute to the cognitive and behavioral phenotype of autism; these deficits may be secondary to abnormalities in cerebellar-frontal circuitry. PMID- 11105466 TI - Category fluency test: normative data for English- and Spanish-speaking elderly. AB - Category fluency tasks are an important component of neuropsychological assessment, especially when evaluating for dementia syndromes. The growth in the number of Spanish-speaking elderly in the United States has increased the need for appropriate neuropsychological measures and normative data for this population. This study provides norms for English and Spanish speakers, over the age of 50, on 3 frequently used measures of category fluency: animals, vegetables, and fruits. In addition, it examines the impact of age, education, gender, language, and depressed mood on total fluency scores and on scores on each of these fluency measures. A sample of 702 cognitively intact elderly, 424 English speakers, and 278 Spanish speakers, participated in the study. Normative data are provided stratified by language, age, education, and gender. Results evidence that regardless of the primary language of the examinee, age, education, and gender are the strongest predictors of total category fluency scores, with gender being the best predictor of performance after adjusting for age and education. English and Spanish speakers obtained similar scores on animal and fruit fluency, but English speakers generated more vegetable exemplars than Spanish speakers. Results also indicate that different fluency measures are affected by various factors to different degrees. PMID- 11105467 TI - Speed and memory in WAIS-R-NI Digit Symbol performance among healthy older adults. AB - Although roles have been proposed for both graphomotor speed and learning in the execution of Digit Symbol, few data have been available concerning performance across the adult lifespan on the Symbol Copy, paired associates, or free recall measures derived from Digit Symbol and recommended in the WAIS-R-NI. We report findings on 177 healthy older adults (ages 50-90), providing normative data by age group, education level, and gender. As previously reported, Digit Symbol scores decline steeply with age (r = -.64). Symbol Copy speed declines almost as steeply (r = -.58). Incidental learning, however, declines only modestly (r = .26 on both measures). Symbol Copy is a far stronger correlate of Digit Symbol (r = .72) than are paired associates or free recall (r = .26 and r = .28, respectively). The 2 incidental learning measures do, however, offer valuable supplementary information as part of a comprehensive individual assessment. When low Digit Symbol scores are produced by slowing on Symbol Copy, further evaluation of perceptual and motor speed and dexterity are indicated. When low incidental learning scores are obtained, further evaluation of memory is warranted. Qualitative analysis of errors (e.g., rotations) made on the incidental learning procedures may also be valuable. PMID- 11105468 TI - Theory of mind and executive functions in normal human aging and Parkinson's disease. AB - Although the majority of research in theory of mind (TOM) has focused on young children or individuals with autism, recent investigations have begun to look at TOM throughout the lifespan and in other neurological and psychiatric populations. Some have suggested that TOM may represent a dissociable, modular brain system that is related to, but separable, from other brain functions including executive functions (EF). Recently, studies have shown that TOM performance can be compromised following an acquired brain insult (e.g., damage to the right hemisphere). However, the relationship of such impaired TOM performance to other brain functions in these cases has not been explored. This study investigated the effects of both normal human aging and Parkinson's disease on TOM. The relationship of TOM performance and EF in these groups was also examined. The results suggested that although TOM performance appeared compromised in the group of individuals with Parkinson's disease, the elderly control participants were relatively unimpaired relative to younger individuals. Significant relationships between several measures of TOM and EF were also found. The implications of these findings, and also the finding that failure on one measure of TOM did not necessarily predict failure on all measures of TOM, are discussed. PMID- 11105469 TI - Learning to read is much more than learning to read: a neuropsychologically based reading program. AB - Departing from the observation that illiterates significantly underscore in some neuropsychological tests, a learning-to-read method named NEUROALFA was developed. NEUROALFA is directed to reinforce these underscored abilities during the learning-to-read process. It was administered to a sample of 21 adult illiterates in Colima (Mexico). Results were compared with 2 control groups using more traditional procedures in learning to read. The NEUROPSI neuropsychological test battery was administered to all the participants before and after completing the learning-to-read training program. All 3 groups presented some improvement in the test scores. Gains, however, were significantly higher in the experimental group in Orientation in Time, Digits Backward, Visual Detection, Verbal Memory, Copy of a Semi-Complex Figure, Language Comprehension, Phonological Verbal Fluency, Similarities, Calculation Abilities, Sequences, and all the recall subtests, excluding Recognition. Performance in standard reading tests was also significantly higher in the experimental group. Correlations between pretest NEUROPSI scores and reading ability were low. However, correlations between posttest NEUROPSI scores and reading scores were higher and significant for several subtests. Results are interpreting as supporting the assumption that reinforcement of those abilities in which illiterates significantly underscore results in a significant improvement in neuropsychological test scores and strongly facilitates the learning-to-read process. The NEUROALFA method of teaching reading to adult illiterates is beginning to be used extensively in Mexico. To our knowledge, this is the first attempt to apply neuropsychological principles to social problems. PMID- 11105470 TI - The Boston Diagnostic Aphasia Examination-Spanish version: the influence of demographic variables. AB - The Boston Diagnostic Aphasia Examination Battery (BDAE) is one of the most widely used aphasia tests worldwide. Information about general population performance, however, is limited. This paper analyzes the effects of gender, age, socioeconomic status (SES), academic achievement, and occupation on the BDAE Spanish version. The BDAE was administered to a randomized sample of 156 occupationally active 19-60-year-old participants (75 male and 81 female) from two SES groups. Gender and age had a significant effect on some reading and writing subtests. Body-part naming and mechanics of writing scores were significantly decreased in the low SES group. Education had a significant impact over most of the BDAE subtests. A stepwise regression model showed that academic achievement was best able to predict the variance in BDAE scores with a low (< 15%) to modest (> 17%) but significant capability (F MANOVA p < .01). A factor analysis disclosed 7 factors that explained 67% of the total variance. PMID- 11105471 TI - Neurobehaviors and psychotic symptoms in Alzheimer's disease. AB - Psychotic symptoms are common in Alzheimer's disease (AD) and clinicoanatomical and neuropsychological evidence indicate an association between these symptoms and frontal lobe dysfunction. Neuro-behaviors associated with frontal dysfunction were assessed in Alzheimer's disease (AD) patients with (n = 20) and without psychotic symptoms (n = 21) matched for mean age, education, gender, and dementia severity. The Frontal Lobe Personality Scale (FLOPs) was completed by patient caregivers to measure behaviors typically associated with frontal dysfunction. Findings indicated that AD patients with psychotic symptoms exhibited significantly greater neurobehavioral dysfunction (FLOPs M = 130.69, SD = 24.70) than AD patients without psychotic symptoms (FLOPs M = 111.10, SD = 25.83). Subscale analyses indicated that psychotic AD patients were more dis-inhibited (M = 28.28, SD = 7.54) than patients without psychotic symptoms (M = 20.92, SD = 4.9). Findings are consistent with and contribute to previous neuropsychological and clinicoanatomical research suggesting increased frontal dysfunction in AD with psychotic symptoms and lend additional empirical support to subtyping AD based on the presence of psychotic symptoms. Furthermore, findings provide preliminary evidence indicating which specific type of neurobehavioral abnormalities are related to the presence of distressing psychotic symptoms. PMID- 11105472 TI - "Subcortical" cognitive impairment in patients with systemic lupus erythematosus. AB - Studies of cognitive functioning in patients with systemic lupus erythematosus (SLE) have found deficits even in patients without other evidence of neurologic involvement. The present study used scores on the 11 items of the Mini-Mental State Exam (MMSE) to classify the cognitive impairment of 93 SLE patients as suggestive of "cortical" or "subcortical" dysfunction using a validated statistical algorithm. Ninety-five percent of patients were categorized as having "subcortical" deficits, and 5% were categorized as having "cortical" deficits. When the analysis was limited to only those with total MMSE scores < or = 24, 81% were classified as "subcortical" and 19% as "cortical." These results suggest that SLE patients can have psychomotor and mental tracking deficits of a type seen in patients with subcortical brain disease, even in the absence of gross neurologic involvement. PMID- 11105473 TI - Current medical and surgical treatment options for gastroesophageal reflux disease. AB - Gastroesophageal reflux disease (GERD) is one of the most common disorders in medicine. The options of medical versus surgical therapies have been highlighted by more potent acid suppression medications and by the introduction of minimally invasive surgery for GERD. Many factors will impact on the treatment selection for each individual patient: the underlying pathophysiology, typical vs atypical symptoms, presence of reflux complications, and success and limitations of medical and surgical treatments. Medical antireflux therapy is very effective and safe, but long-term maintenance therapy is required for most patients. Minimally invasive antireflux surgery has provided excellent results, but outcome is dependent on patient selection and surgical expertise. Careful pre-operative evaluation is essential to determine the optimal treatment and surgical approach. PMID- 11105474 TI - Public access defibrillators in Kentucky. PMID- 11105475 TI - Building mental health professionals' decisional models into tests of predictive validity: the accuracy of contextualized predictions of violence. AB - To safely manage potentially violent patients in the community, mental health professionals (MHPs) must assess when and under what conditions a patient may be involved in a violent act. This study applies a more ecologically sensitive approach than past research by building the conditions that MHPs believe make patient violence more likely into tests of their predictive validity. In specific, the accuracy of MHPs' predictions that patients were more likely to become violent when they consumed alcohol was assessed based on a sample of 714 patients. The results indicate that MHPs do not discriminate well between patients who are likely to become violent during periods in which they drink from those who are not. MHPs' predictions appear more descriptive of the drinking behavior of a high-risk group than predictive of alcohol-related violent incidents. Thus, even when their apparent decisional processes are considered in tests of accuracy, MHPs' predictions of violence are only moderately more accurate than chance. This paper analyzes the implications of these findings for risk assessment practice and for conducting further clinically relevant research. PMID- 11105476 TI - Lawyers' questioning: the effect of confusing questions on witness confidence and accuracy. AB - The present study investigated the effect on witness confidence and accuracy of confusing questions often used by attorneys in court. Participants viewed a videotaped film and were individually questioned about the incident 1 week later. Half the participants were asked questions using six categories of confusing questions (negatives, double negatives, leading, multiple questions, complex syntax, and complex vocabulary); the remaining half were asked for the same information using simply phrased equivalents. Confusing questions reduced participant-witnesses' accuracy and suppressed confidence-accuracy relationships compared with the condition where simplified alternatives were asked. Witness performance was impaired by the fact that mock-witnesses rarely asked for a confusing question to be explained or qualified their answers. This experiment demonstrates the importance of ensuring that lawyers ask witnesses simple, clear, questions. PMID- 11105477 TI - Truth, lies, and videotape: an investigation of the ability of federal parole officers to detect deception. AB - The ability of a group of Canadian federal parole officers to detect deception was investigated over the course of 2 days of lie detection training. On the first day of training, 32 officers judged the honesty of 12 (6 true, 6 fabricated) videotaped speakers describing personal experiences, half of which were judged before and half judged after training. On the second day, 5 weeks later, 20 of the original participants judged the honesty of another 12 videotapes (again, 6 pre- and 6 posttraining). To isolate factors relating to detection accuracy, three groups of undergraduate participants made judgments on the same 24 videotapes: (1) a feedback group, which received feedback on accuracy following each judgment, (2) a feedback + cue information group, which was given feedback and information on empirically based cues to deception, and (3) a control group, which did not receive feedback or cue information. Results indicated that at baseline all groups performed at or below chance levels. However, overall, all experimental groups (including the parole officers) became significantly better at detecting deception than the control group. By the final set of judgments, the parole officers were significantly more accurate (M = 76.7%) than their baseline performance (M = 40.4%) as well as significantly more accurate than the control group (M = 62.5%). The results indicate that detecting deceit is difficult, but training and feedback can enhance detection skills. PMID- 11105478 TI - Incapacitation and just deserts as motives for punishment. AB - What motivates a person's desire to punish actors who commit intentional, counternormative harms? Two possible answers are a just deserts motive or a desire to incarcerate the actor so that he cannot be a further danger to society. Research participants in two experiments assigned punishments to actors whose offenses were varied with respect to the moral seriousness of the offense and the likelihood that the perpetrator would commit similar future offenses. Respondents increased the punishment as the seriousness of the offense increased, but their sentences were not affected by variations in the likelihood of committing future offenses, suggesting that just deserts was the primary sentencing motive. Only in a case in which a brain tumor was identified as the cause of an actor's violent action, a case that does not fit the standard prototype of a crime intentionally committed, did respondents show a desire to incarcerate the actor in order to prevent future harms rather than assigning a just deserts based punishment. PMID- 11105479 TI - Witnessing-condition heterogeneity and witnesses' versus investigators' confidence in the accuracy of witnesses' identification decisions. AB - Undergraduate participants were tested in 144 pairs, with one member of each pair randomly assigned to a "witness" role and the other to an "investigator" role. Each witness viewed a target person on video under good or poor witnessing conditions and was then interviewed by an investigator, who administered a photo line up and rated his or her confidence in the witness. Witnesses also (separately) rated their own confidence. Investigators discriminated between accurate and inaccurate witnesses, but did so less well than witnesses' own confidence ratings and were biased toward accepting witnesses' decisions. Moreover, investigators' confidence made no unique contribution to the prediction of witnesses' accuracy. Witnesses' confidence and accuracy were affected in the same direction by witnessing conditions, and there was a substantial confidence accuracy correlation when data were collapsed across witnessing conditions. Confidence can be strongly indicative of accuracy when witnessing conditions vary widely, and witnesses' confidence may be a better indicator than investigators'. PMID- 11105480 TI - Accuracy of investigators' verbatim notes of their forensic interviews with alleged child abuse victims. AB - Verbatim contemporaneous accounts of 20 investigative interviews were compared with audiotaped recordings thereof. More than half (57%) of the interviewers' utterances along with 25% of the incident-relevant details provided by the children were not reported in the "verbatim" notes. The structure of the interviews was also represented inaccurately in these accounts. Fewer than half (44%) of the details provided by the children were attributed to the correct eliciting utterance type. Investigators systematically misattributed details to more open rather than more focused prompts. These results underscore the superiority of electronic recording when the content and structure of investigative interviews must be preserved. PMID- 11105481 TI - Remaining challenges towards elimination of leprosy. PMID- 11105482 TI - The Global Alliance for the Elimination of Leprosy (GAEL) PMID- 11105483 TI - Leprosy research in the new millennium in the special programme for research and training in tropical diseases (TDR) PMID- 11105484 TI - Monitoring motor nerve function in leprosy patients. AB - Manual muscle strength testing has an important function in the management of leprosy patients. Its importance was first recognized in the 1960s, especially when following patients who were started on steroid treatment to monitor the nerve function and the effect of treatment. In those days, and still in many centres today, many or all muscles were tested that are innervated by the nerves that can be at risk in leprosy. The author argues that not all muscles innervated by the nerves at risk need to be tested and also that many muscles cannot be tested in isolation. A muscle charting form is presented which is suitable for screening purposes, and that also allows for more detail when motor function is impaired. PMID- 11105485 TI - Are cytokines our key to immunity against mycobacterial diseases? PMID- 11105486 TI - A prospective cohort study comes of age. PMID- 11105487 TI - The ALERT MDT Field Evaluation Study (AMFES): a descriptive study of leprosy in Ethiopia. Patients, methods and baseline characteristics. AB - The ALERT MDT Field Evaluation Study (AMFES) is a long-term prospective study of 650 patients (594 new cases and 56 relapses after dapsone monotherapy), treated with fixed-duration multiple-drug therapy (MDT), as recommended by WHO. Follow-up has continued for up to 11 years from the start of treatment. This paper presents the methodology of the study and the baseline characteristics of the cohort, while accompanying papers examine the incidence of, and possible risk factors for, the various complications of leprosy, including relapse, reactions and nerve function impairment. The methods of diagnosis, classification and treatment with MDT are described; nerve function was assessed at every visit to the clinic using a standardized methodology, so that reactions and new impairment could be detected early and treated. Eighty-four per cent of new case had at least one thickened nerve, with the ulnar nerve most commonly involved. Seventy-seven per cent of cases completed treatment and only one adverse reaction to the MDT drugs was noted. Twenty-eight per cent of all patients were given steroids at one time or another, almost always for new nerve function impairment, and 3% of these developed significant complications of steroid treatment. Twenty-nine patients (5%) received hospital care, including 14 patients who underwent major surgery. Sixty-one per cent of the women over 19 years of age had at least one pregnancy, but pregnancies were much less common after leprosy was diagnosed. PMID- 11105488 TI - The pattern of leprosy-related neuropathy in the AMFES patients in Ethiopia: definitions, incidence, risk factors and outcome. AB - The ALERT MDT Field Evaluation Study (AMFES) began in 1988 and followed patients prospectively for up to 10 years after release from treatment (RFT). This paper presents the findings from this cohort with regard to neuropathy and nerve damage. Five hundred and ninety-four new cases of leprosy are included in the study, 300 multibacillary (MB) and 294 paucibacillary (PB) cases. Fifty-five percent of patients had some degree of impairment at diagnosis and a further 73 (12%) developed new nerve function impairment (NFI) after starting multiple drug therapy (MDT). The overall incidence rate for neuropathy was 39 episodes per 100 PYAR in the first year after diagnosis, gradually declining to 12 episodes per 100 PYAR in the sixth year. In those patients without impairment at diagnosis, the incidence rate of neuropathy was 25 episodes per 100 PYAR for MB cases and 11 per 100 PYAR for PB cases in the first year; in 33% of MB cases whose first episode of neuropathy occurred after diagnosis, that first episode took place after the first year, or after the normal period of treatment with MDT. Seventy three patients with neuropathy developing after diagnosis are reported more fully: 34 (47%) had only one nerve involved and of these 25 (73%) had a single, acute episode of neuropathy. Nine (27%) had further episodes. Thirty-nine (53%) had more than one nerve involved and of these 16 (41%) had a single, acute episode, while 23 (59%) had further episodes. The terms 'chronic' and 'recurrent' neuropathy are defined and used to describe the pattern of neuropathy in those with repeated attacks. In patients with no impairment at the start of the study, treatment with steroids resulted in full recovery in 88% of nerves with acute neuropathy but only 51% of those with chronic or recurrent neuropathy. The median time to full recovery from acute neuropathy was approximately 6 months, but in a few cases recovery occurred gradually over 2-3 years. Severe neuropathy was less likely to be followed by a complete recovery than mild or moderate neuropathy. Forty-two percent of nerves with acute neuropathy that were not treated with steroids also fully recovered. In the group of patients who were thought to have old, permanent impairments at diagnosis, full recovery of nerve function occurred in 87/374 (23%) of the nerves involved. The overall outcome is illustrated by examining the average EHF score for groups of patients. Patients with no new neuropathy after diagnosis show a gradual improvement in their EHF score, while those with any episodes of neuropathy after diagnosis show a gradual deterioration after completion of MDT. Possible explanations for these findings are discussed. Risk factors for neuropathy, for chronic and recurrent neuropathy, and for a poor outcome 5 years after release from treatment, are examined. Impairment at diagnosis was the main risk factor for a poor outcome, accompanied by the occurrence of chronic/recurrent neuropathy or a reversal reaction. PMID- 11105489 TI - Reversal reactions in the skin lesions of AMFES patients: incidence and risk factors. AB - Reversal reactions affect the skin and/or nerves of leprosy patients. This paper looks at reversal reactions involving the skin in 594 new patients in central Ethiopia, followed for between 6 and 11 years after the start of treatment. The incidence of reversal reaction declines steadily after the start of treatment, but the first episode may occur as long as 5 years after diagnosis in both paucibacillary (PB) and multibacillary (MB) patients. Recurrent episodes occurred up to 6 years after diagnosis. PB patients were at greatest risk for reversal reaction in the first year after diagnosis and MB patients in the first 4 years. The highest incidence rate was 18 episodes per 100 person years in MB patients during the first year after diagnosis. The ratio of the incidence rates for the first 3 years in MB versus PB patients is 2.4 (95% CI 1.6-3.8). This study confirms that starting effective treatment and borderline classification are risk factors for reversal reactions. Pregnancy/delivery in the 6 months prior to diagnosis was a significant risk factor for presenting with a reversal reaction [relative risk (RR) 5.9 (95% CI 2.1-16.5)], but later pregnancies were not associated with an increased risk. Being female was a significant risk factor for the late appearance of the first episode of reversal reaction. Having a reversal reaction in the first year after diagnosis was a highly significant risk factor for the development of later reactions [RR in PB cases 11.9 (95% CI 3.4-41.7); in MB cases 6.4 (95% CI 3.8-10.6)]. Being HIV positive was a risk factor for developing recurrent reversal reactions, although only three out of 29 recurrent cases were HIV positive [RR 2.7 (95% CI 1.4-5.1)]. PMID- 11105490 TI - ENL reactions in the multibacillary cases of the AMFES cohort in central Ethiopia: incidence and risk factors. AB - Erythema nodosum leprosum (ENL), or type 2 leprosy reactions are an important complication of multibacillary leprosy. The AMFES cohort includes 300 new multibacillary cases that have been followed for up to 10 years from the start of treatment, in central Ethiopia. Sixteen (5.3%) patients had ENL reactions. The incidence of ENL was maximal in the second and third years after the start of treatment, reaching 6.9 episodes per 100 person years at risk. Factors associated with being lepromatous [LL classification and a high bacillary index (BI)] gave an increased risk of developing ENL; in the univariate analysis, LL classification gave a relative risk of 3.6 (95% CI 1.3-10) and a BI of 6 gave a relative risk of 8.6 (95% CI 2.3-32) for the development of ENL. HIV co-infection was found to be a risk factor in this cohort, but as the numbers involved are small (only two HIV positive patients had ENL), this finding must be confirmed in larger studies. Ten of the 16 cases had recurrent episodes and five had at least five episodes occurring over a period of more than 2 years. The management and prognosis of ENL reactions are discussed. PMID- 11105491 TI - Relapses after fixed duration multiple drug therapy: the AMFES cohort. AB - Relapse rates after multiple-drug therapy (MDT) have been low, although there remains a concern about the possibility of late relapse in those with an initially high bacterial load. In all, 502 patients in the AMFES cohort completed fixed-duration MDT and are included in this report. There have been no confirmed relapses in the AMFES cohort, in a follow-up period of up to 8 years after completion of treatment, even in the 57 cases with an initial average bacillary index of > or = 4.0, 20 of whom have been followed for more than 5 years after ceasing MDT. Methods of diagnosing a relapse are discussed. PMID- 11105492 TI - The pattern of decline in bacillary index after 2 years of WHO recommended multiple drug therapy: the AMFES cohort. AB - With effective antibiotic treatment, the bacillary index (BI) in multibacillary leprosy patients declines over a number of years. This can be quantified as a rate of decline in log-units per year or as the time until smear negativity is reached. In the AMFES cohort 220 cases had data on the changes in their BI over time, while 170 cases are documented until smear negativity. The average BI at the start was 3.3 (SD 1.5; range 0.3-5.5) and the mean rate of decline was 0.85 units per year (median 0.7 units per year); in the first 2 years after diagnosis, the mean rate of decline was 1.15 units per year. The rate of decline was not related to any clinical features of the disease except delay in diagnosis: patients presenting for treatment early had a significantly faster rate of clearing the bacilli (adjusted relative risk 2.3; 95% CI 1.0-5.1). Fifty-eight percent of cases took longer than 3 years to reach smear negativity, but this time interval is largely determined by the initial BI and classification, making it a less useful indicator of bacterial clearance. More severe impairment at the start of treatment was associated with a faster return to smear negativity, for which no obvious explanation can be given. Reversal reactions, which occurred in 25% of the cases reviewed, are not associated with a more rapid clearance of bacilli. PMID- 11105493 TI - The effect of HIV status on the clinical picture of leprosy: a prospective study in Ethiopia. AB - No major interaction between HIV infection and leprosy has been documented. The ALERT MDT Field Evaluation Study (AMFES) has allowed the examination of possible interactions in a prospective manner, although the total number of HIV-positive individuals was not high at 22 (3.8%) of 581 patients tested. There was an excess number of deaths in the HIV-positive group: 27% compared with 5.7% in the HIV negative group, although the causes of death were not recorded (relative risk 4.8; 95% CI 2.2-10.2). HIV-positive individuals had a higher risk of ENL reactions (relative risk 5.2; 95% CI 1.7-15.9). Reversal reactions and neuritis (both acute and chronic) were not significantly influenced by HIV status, although there was a possible increase in recurrent reversal reactions in HIV positive cases (relative risk 2.2; 95% CI 0.98-4.7). There was no evidence to suggest an increased risk of developing leprosy or of developing multibacillary rather than paucibacillary disease. There was no association between HIV positivity and the development of impairment. PMID- 11105494 TI - The hand-foot impairment score as a tool for evaluating prevention of disability activities in leprosy: an exploration in patients treated with corticosteroids. AB - The hand-foot (HF) impairment score in leprosy patients is the sum of the WHO disability grades for hands and feet. This retrospective study explored the possibility of using the HF score for evaluation of the effectiveness of corticosteroid treatment programmes for nerve function impairment (NFI). Changes in the score were compared with changes in sensory testing (ST) and voluntary muscle testing (VMT) for 42 leprosy patients who received corticosteroid treatment. The WHO grade did not change in 30/60 (50%) of extremities gaining, and in 4/10 (40%) extremities losing sensation and/or muscle strength. However, 18/24 (75%) patients with a definite gain in function improved in HF score, while the HF score remained unchanged in 10/11 (91%) patients with no change in nerve function. Five patients with impairment in multiple extremities showed both gain and loss of sensation and/or muscle strength in the same or different extremities. Overall, improvement, deterioration and absence of change in NFI, as indicated by changes in ST and VMT were reflected correctly by the HF score in 28 (76%) of the remaining 37 patients. It was also shown that the HF score does not give appropriate information on the extent of the effect of corticosteroid treatment. This study illustrates that the HF score can not be used to support management of corticosteroid treatment of individual patients, but indicates this score to be a promising device for the evaluation of the effectiveness of corticosteroid treatment programmes. This study used the HF score because information on (changes in) eye impairment was not considered reliable. However, in principle, we consider the EHF score, which is the sum of the WHO disability grades for hands, feet and eyes, preferable for evaluation purposes. We strongly recommend further validation of the EHF score as a tool for evaluation of corticosteroid treatment programmes for patient groups with different distributions of NFI through prospective studies. PMID- 11105495 TI - High dose prednisolone treatment of leprosy patients undergoing reactions is associated with a rapid decrease in urinary nitric oxide metabolites and clinical improvement. AB - Evidence is accumulating that nitric oxide (NO) produced by macrophages has a role in the pathogenesis of reactions in leprosy. We followed the urinary levels of the metabolites of NO [nitrite (NO2-) and nitrate (NO3-)] and the clinical response to prednisolone treatment in leprosy patients (n = 9) admitted to ALERT leprosy hospital Addis Ababa, Ethiopia, because of reversal reaction (RR) or erythema nodosum leprosum (ENL). In untreated reactional leprosy patients, the levels of urinary NO metabolites (1645 +/- 454 microM, n = 9, ENL = 4, RR = 5) decreased significantly 2 weeks after high dose prednisolone treatment (1075 +/- 414 microM, P < 0.05), and remained stable 4 (895 +/- 385 microM, P < 0.02) and 6 weeks following treatment initiation (1048 +/- 452 microM, P < 0.02). This decrease was also present when the reactional patients were subdivided according to the type of reaction (ENL, RR) and coincided with a clinical improvement. In patients showing a poor clinical response to steroids, no or minor effects on the urinary NO metabolite levels were observed. We conclude that there is a correlation between the decrease in urinary NO metabolites and a favourable clinical response after high dose prednisolone treatment of reactional leprosy patients. PMID- 11105496 TI - Predisposing factors for recurrent skin ulcers in leprosy. AB - This study was designed to determine the factors associated with recurrence of leprosy ulcers. Between April and August 1992, 55 consecutive leprosy patients admitted with skin ulcers were studied. Factors predisposing to recurrence, e.g. patient's age, disease duration, ulcer site, ulcer depth and physical deformity (taking into account neuromuscular and skeletal damage) were evaluated. Ulcer recurrence occurred in 40/55 (75%) patients. Recurrent ulceration was associated with location in the lower extremity (P = 0.02), where recurrences were more common in the midfoot and heel (P = 0.01). Recurrence was also associated with severity of physical deformity (P = 0.01), which increased the odds of recurrent ulceration by 4.2 times (95% confidence interval, 1.01-18.3). The severity of physical deformity itself was associated with the age of the patient (P = 0.04) and the disease duration (P = 0.02). In conclusion, there is a need to focus on identification of risk factors for recurrent leprosy ulceration. Targeted prevention strategies would be required if morbidity associated with recurrent skin ulceration is to be avoided. PMID- 11105497 TI - Towards an understanding of non-compliance. An assessment of risk factors for defaulting from leprosy treatment. AB - Within the Eastern Leprosy Control Project of Nepal, a retrospective case control study looked for simple factors that might be used operationally to predict non compliant behaviour in patients. Patients with these factors would then become the targets of measures such as intensified health education messages and home visits in order to reduce the risk of defaulting. A study of 1442 patient cards (half defaulters, half treatment completed) revealed occasional small but significant demographic and clinical differences, but none was of a sufficient magnitude to be operationally useful. Review of the attendance of patients in the first few months of treatment suggested that eventual defaulting was strongly associated with irregularity from the commencement of treatment. It is possible that an early indicator based on attendance over the first months can be used to target patients who are in danger of non-completion of treatment. PMID- 11105498 TI - Modified leprosy elimination campaign (MLEC) for case detection in a remote tribal area in the State of Orissa, India. AB - A leprosy project was established in a difficult to reach area under guidelines of Government of India. The leprosy services were provided by Koraput Leprosy Eradication Project (KORALEP) and general health services by Primary Health Care (PHC). Leprosy elimination campaigns (LECs) were suggested by WHO to detect more cases in the community. A modified leprosy elimination campaign (MLEC), carried out utilizing the services of primary health care workers is discussed in this paper. Apart from the trained health workers, Anganwadi workers along with some literate people from the district were also included in the search teams. In all, 1543 cases were shortlisted from the suspects identified and on re-examination 576 cases were confirmed as active cases. Sixty percent of the cases detected were very early cases with two to three skin lesions. This could be achieved with a very brief training of health workers and involving village voluntary workers. MLEC was found to be a useful tool for case finding in such areas. PMID- 11105499 TI - Acro-osteolysis prior to diagnosis of leprosy. AB - lysis (bone resorption) has been observed in a heterogeneous group of congenital and acquired bone disorders. Leprosy is the main cause of peripheral neuropathy leading to acro-osteolysis in endemic countries. Pure neuritic leprosy, a less common form of the disease, is difficult to diagnose. Two unrelated leprosy patients with acropathy whose disease began as pure neuritic are discussed. PMID- 11105500 TI - 'Wall Journal' on leprosy--a novel method to educate medical students. PMID- 11105501 TI - WHO leprosy elimination campaign--beyond 2005. PMID- 11105502 TI - Blocking natural killer cell activity. PMID- 11105503 TI - Quality improvement: new contributions from the field of health services research. PMID- 11105504 TI - Are managed care plans organizing for quality? AB - This article examines the degree to which managed care organizations (MCOs) are reorganizing to take responsibility for the quality of care and service they provide. Specifically, factors prompting plans to focus on quality improvement (QI) and how they may be building the capacity to improve quality are considered. The authors' analysis is based on executive interviews with the plan medical directors, QI directors, and chief executive officers (CEOs) in a sample of 24 health plans. The overall response rate was 58.3 percent (medical director = 62.5 percent, QI director = 79.2 percent, CEO = 33.3 percent). The authors queried respondents about (1) perceived drivers and obstacles to the development of an effective QI program, (2) plan organizational structure for QI, and (3) technical capacities for data collection, management, and performance measurement. The results suggest that MCOs are responding to outside pressures to engage in QI. They are reorganizing their management structures and more slowly and tentatively are building technical capacity for QI. PMID- 11105505 TI - Employers: quality takers or quality makers? AB - This article provides a synthesis of past research to help understand the extent to which employers are using their considerable market power to drive health care quality. Are employers quality takers or quality makers? The literature provides some clues about aspects of quality employers are attempting to influence, strategies they are pursuing to influence quality, and their impact. Some employers are interested in some indicators of quality and are incorporating them in a variety of different purchasing strategies. The indicators most frequently used by employers, however, probably are not the ones that clinical experts and policy makers would select as most reflective of clinical quality. It appears that employers as a group are becoming more informed quality takers but are not yet quality makers--with the exception of a few well-resourced outliers. Recent events provide mixed signals about whether the future employer role in influencing quality will diminish, stall, or flourish. PMID- 11105506 TI - Translating behavioral health services research into benefits policy. AB - This article uses a 4-pronged statistical approach to examine the impact of a mental health carve-out at a major employer. To examine net financial impact of the carve-out, the authors perform a pre-post, multivariate regression analysis of changes in costs. Using a random-effects model, the authors explore the ultimate financial impact of the carve-out for patients and for the firm. Using a multinomial logistic regression, they examine differing program effects by intensity of use. A fixed-effects negative binomial regression models the episodic nature of outpatient care, controlling for patient-specific unobserved characteristics that influence health care utilization. The carve-out slightly reduced overall mental health costs and utilization while expanding entry-level access to routine services. At the same time, the specific carve-out shifted financial burdens from the firm onto high-utilization patients. Therefore, this carve-out appears poorly suited to the care of individuals experiencing severe and debilitating psychiatric disorders. PMID- 11105507 TI - Are women being counseled about estrogen replacement therapy? AB - The U.S. Preventive Services Task Force and several medical professional associations have published guidelines recommending that all women be counseled around the time of menopause about the benefits and risks of estrogen replacement therapy (ERT) so that they may make an informed decision about its use. Despite the proliferation of ERT counseling guidelines, little is known about whether these guidelines are being followed. There were 1,500 female members (aged 40 to 69) of a Northeastern U.S. Independent Practice Association--model Health Maintenance Organization who were surveyed, and 51 percent reported that a health care provider had talked with them about the benefits and risks of ERT. In multivariate analyses, a woman's demographic characteristics (age, race, income), stage of menopause, severity of menopausal symptoms, and body weight were the major correlates of receipt of ERT counseling. Women at greater risk for osteoporosis or heart disease were no more likely to be counseled, although those with diagnosed osteoporosis were. What appear to be selective ERT counseling practices will need to be modified if the goal of providing universal ERT counseling to midlife women is to be attained. PMID- 11105508 TI - Wrestling with typology: penetrating the "black box" of managed care by focusing on health care system characteristics. AB - The health care system has undergone a fundamental transformation undermining the usefulness of the typology of the health maintenance organization, the independent practice association, the preferred provider organization, and so forth. The authors present a new approach to studying the health care system. In matrix form, they have identified a set of organizational and delivery characteristics with the potential to influence outcomes of interest, such as access to services, quality, health status and functioning, and cost. The matrix groups the characteristics by domain--financial features, structure, care delivery and management policies, and products--and by key roles in the health care system--sponsor, plan, provider intermediary organization, and direct services provider. The matrix is a tool for researchers, administrators, clinicians, data collectors, regulators, and other policy makers. It suggests a new set of players to be studied, emphasizes the relationships among the players, and provides a checklist of independent, control, and interactive variables to be included in analyses. PMID- 11105509 TI - Assessing the relationship between quality of care and the characteristics of health care organizations. AB - In the past two decades, relationships among health plans, medical groups, and providers have grown more complex and the number of clinical management strategies has increased. In this context, determining the independent effect of a particular organizational strategy on quality of care has become more difficult. The authors review some of the issues a researcher must address when studying the relationship between organizational characteristics and quality of care. They offer criteria for selecting a research question, list organizational characteristics that may influence quality, and suggest sampling and study design techniques to reduce confounding. Since this type of research often requires a health care organization as collaborator, the authors discuss strategies for developing research partnerships and collecting data from the partner organization. Finally, they offer suggestions for translating research into policy. PMID- 11105510 TI - Quality measures for mental health care: results from a national inventory. AB - The National Inventory of Mental Health Quality Measures was funded by the Agency for Healthcare Research and Quality to (1) inventory process measures for assessing the quality of mental health care; (2) identify clinical, administrative, and quality domains where measures have been developed; and (3) identify areas where further research and development is needed. Among the 86 measures identified, most evaluated treatment of major mental disorders, for example, schizophrenia (24 percent) and major depression (21 percent). A small proportion focused on children (8 percent) or the elderly (9 percent). Domains of quality included treatment appropriateness (65 percent), continuity (26 percent), access (26 percent), coordination (13 percent), detection (12 percent), and prevention (6 percent). Few measures were evaluated for reliability (12 percent) or validity (3 percent). Measures imposing a lower burden were more likely to be in use (chi 2 = 4.41, p = .036). Further measures are needed to assess care for several priority clinical and demographic groups. Research should focus on measure validity, reliability, and implementation costs. In order to foster quality improvement activities and use of common measures and specifications for mental health care, the inventory of quality measures will be made available at www.challiance.org/cqaimh. PMID- 11105511 TI - Selection bias in HMOs: a review of the evidence. AB - Early reviews found that health maintenance organizations (HMOs) attracted healthier beneficiaries in the Medicare program and healthier employees in the market for employer-based insurance. This review finds that HMOs still attract healthier Medicare beneficiaries, that HMOs no longer attract healthier employees, and that HMOs attract healthier Medicaid recipients. This review also found conflicting evidence about whether Medicare HMOs are overpaid, no evidence that HMOs are overpaid in the market for employer-based insurance, and evidence that concerns about overpaying Medicaid HMOs have diminished because many states are adopting mandatory programs. PMID- 11105512 TI - Managed care litigation: legal doctrine at the boundary of tort and contract. AB - This article summarizes the various approaches to how the law should assign responsibility in a system where health care financing and delivery are combined. Health law scholars have been debating whether conflicts in managed care between individual patient needs and preserving assets for the patient population should be resolved by tort or contract law. Until recently, the literature has been dominated by scholars arguing that managed care should be guided by contractual arrangements and concepts to stimulate the market changes occurring in health care delivery. We summarize the arguments for and against both contract and tort, along with recent attempts to bridge the gap between the two approaches. The case in favor of a contract regime fails to account for the hybrid nature of managed care delivery and the context in which managed care litigation arises. Thus, tort law retains a fundamental monitoring role in the managed care era. PMID- 11105513 TI - The implications of plan design options for Medicare beneficiaries. AB - To make an informed selection between traditional Medicare and a Medicare managed care plan, a consumer needs to understand the implications of choosing one over the other. What are the implications of plan design for care, cost, and patient autonomy? Consumers need information about these questions. However, a barrier to developing this consumer information is the lack of a consistent body of evidence. An intermediate step is to tap expert knowledge. The purpose of this study is to use expert consensus (across a spectrum of health care experts) to identify the implications of plan design. Experts were surveyed and the degree to which there is consensus provides an initial picture of what experts judge to be important to the consumer. The findings show that experts agree on several implications associated with choosing managed care over the traditional Medicare plan. They also agree that many of these attributes vary considerably across health plans. PMID- 11105514 TI - Determinants of antidepressant treatment compliance: implications for policy. AB - Depression is among the most prevalent, devastating, and undertreated disorders in our society. Treatment with antidepressant medications is effective in controlling symptoms, but treatment beyond the point of symptom resolution is necessary to restore functional status and prevent recurrent episodes. An important step in improving compliance is to identify the determinants of antidepressant treatment compliance. A broader motivation for our study is to examine compliance by patients with a chronic but treatable disease. With claims data between 1990 and 1993, this study uses logistic regression analysis to examine the determinants of compliance among 2,012 antidepressant recipients. The results show that initiating treatment with a tricyclic antidepressant reduces the probability of antidepressant treatment compliance. Initiating treatment with a selective serotonin reuptake inhibitor and undergoing family, group, or individual psychotherapy treatments increase the probability of compliance. Case management does not meaningfully affect compliance. Implications for policy and clinical practice are discussed. PMID- 11105515 TI - What is free association and what does it measure? AB - This paper reports the results of a study of free association in which participants were asked to produce the first two words to come to mind. The findings were used to estimate the reliability of indices of strength and set size for different types of items and to model free association as a retrieval task. When confined to first responses, reliability was generally high for both indices, particularly for words with smaller sets of associates and stronger primaries. When second responses were included, reliability declined. A second response added new but weak items to the set, and, when the primary associate was not produced on the first opportunity, it tended not to be produced on the second. Relative to when multiple responses are requested, first-response free association provides more reliable indices of the relative strength and set size for a word's strongest associates. A model of free association assuming that a strength distribution underlies each response provided a good fit to the data. PMID- 11105516 TI - On associations between computers and restaurants: rapid learning of new associations on a conceptual implicit memory test. AB - A novel event-based conceptual implicit memory test was designed to tap the development of new associations between objects and ad hoc categories. At study, participants were presented with a plausible story that linked an incongruous object (computer) with an ad hoc category (restaurant). At test, participants judged whether a given object was typically found in a restaurant. In Experiment 1, judgment time was significantly slower for the incongruous object (computer) when the story had previously linked the computer to the restaurant, relative to when it had not. Experiment 2 replicated this effect and ruled out the alternative interpretation that this interference effect was attributable to a general slowing of responses to all studied items. Unlike in prior studies, this demonstration of associative priming cannot be attributed to perceptual priming or to test awareness in memory-intact participants. The paradigm therefore offers a unique opportunity to study single-trial conceptual learning in memory-intact and memory-impaired populations. PMID- 11105517 TI - Associative recognition: a case of recall-to-reject processing. AB - Two-process accounts of recognition memory assume that memory judgments are based on both a rapidly available familiarity-based process and a slower, more accurate, recall-based mechanism. Past experiments on the time course of item recognition have not supported the recall-to-reject account of the second process, in which the retrieval of an old item is used to reject a similar foil (Rotello & Heit, 1999). In three new experiments, using analyses similar to those of Rotello and Heit, we found robust evidence for recall-to-reject processing in associative recognition, for word pairs, and for list-discrimination judgments. Put together, these results have implications for two-process accounts of recognition. PMID- 11105518 TI - Brain potentials of recollection and familiarity. AB - It is widely hypothesized that separate recollection and familiarity processes contribute to recognition memory. The present research measured event-related brain potentials (ERPs) from 128 head locations to identify patterns of brain activity related to recollection and familiarity. In two experiments, subjects performed a recognition memory task requiring discrimination between previously studied words, similar words that changed plurality between study and test, and new words (following Hintzman & Curran, 1994). The FN400 ERP component (300-500 msec) varied with the familiarity of words (new > studied = similar). The parietal component (400-800 msec) was associated with the recollection of plurality (studied > similar = new). Differences in the timing and spatial topography of the FN400 and parietal effects support the view that familiarity and recollection arise from distinct neurocognitive processes. PMID- 11105519 TI - "Aha" effects in the generation of pictures. AB - An "aha" effect in memory was first reported by Auble, Franks, and Soraci (1979). They demonstrated that recall was greater for sentences that were initially incomprehensible but which were eventually comprehended, as compared with sentences that were understood from the outset. The present studies extend this "aha" effect to memory for pictorial stimuli. In Experiment 1, a recall advantage for pictures encoded by connecting the dots as compared with those encoded by tracing or visual scanning occurred only in the absence of foreknowledge of the picture (i.e., an "aha" effect). In Experiment 2, we replicated this finding and obtained evidence that conceptually based, verbal foreknowledge does not function in a similar manner as does pictorial foreknowledge in suppressing the "aha" recall advantage. These results place important constraints on previous research on generation effects for visual stimuli and attest to the cross-modal generalizability of the "aha" effect. PMID- 11105520 TI - Perceptual interference decays over short unfilled intervals. AB - The perceptual interference effect refers to the fact that object identification is directly related to the amount of information available at initial exposure. The present article investigated whether perceptual interference would dissipate when a short, unfilled interval was introduced between exposures to a degraded object. Across three experiments using both musical and pictorial stimuli, identification performance increased directly with the length of the unfilled interval. Consequently, significant perceptual interference was obtained only when the interval between exposures was relatively short (< 500 msec for melodies; < 300 msec for pictures). These results are consistent with explanations that attribute perceptual interference to increased perceptual noise created by exposures to highly degraded objects. The data also suggest that perceptual interference is mediated by systems that are not consciously controlled by the subject and that perceptual interference in the visual domain decays more rapidly than perceptual interference in the auditory domain. PMID- 11105521 TI - Verbal coding in olfactory versus nonolfactory cognition. AB - Two paired-associate memory experiments were conducted to investigate verbal coding in olfactory versus nonolfactory cognition. Experiment 1 examined the effects of switching/not switching odors and visual items to words between encoding and test sessions. Experiment 2 examined switching/not switching perceptual odors and verbal-imagine versions of odors with each other. Experiment 1 showed that memory was impaired for odors but not visual cues when they were switched to their verbal form at test. Experiment 2 revealed that memory was impaired for both odors and verbal-imagine cues when they were switched in format at test and that odor sensory imagery was not accessed by the instruction to imagine a smell. Together, these findings suggest that olfaction is distinguished from other sensory systems by the degree of verbal coding involved in associated cognitive processing. PMID- 11105522 TI - Asymmetry between encoding and retrieval processes: evidence from divided attention and a calibration analysis. AB - Two experiments provide further information on the effects of divided attention (DA) on encoding and retrieval processes. The first experiment examined the effects of decision and motor difficulty of a concurrent reaction time task. A calibration analysis was used in the second experiment to test the hypothesis that shifting attentional emphasis away from encoding to the secondary task reduces the level of processing the to-be-remembered items receive. Overall, the results confirm and extend the conclusions of Craik, Govoni, Naveh-Benjamin, and Anderson (1996) and Naveh-Benjamin, Craik, Guez, and Dori (1998), by pointing to clear differences between encoding and retrieval processes: Encoding is affected by simultaneous task demands, especially those associated with "central" resources involved in conscious decision making, whereas retrieval is obligatory in that it is largely immune to the effects of simultaneous demands. The results of the calibration analysis suggest that one reason for the poorer memory performance as a result of DA at encoding is a qualitative shift to less deep, elaborative strategies. PMID- 11105523 TI - The categorical perception of colors and facial expressions: the effect of verbal interference. AB - A series of five experiments examined the categorical perception previously found for color and facial expressions. Using a two-alternative forced-choice recognition memory paradigm, it was found that verbal interference selectively removed the defining feature of categorical perception. Under verbal interference, there was no longer the greater accuracy normally observed for cross-category judgments relative to within-category judgments. The advantage for cross-category comparisons in memory appeared to derive from verbal coding both at encoding and at storage. It thus appears that while both visual and verbal codes may be employed in the recognition memory for colors and facial expressions, subjects only made use of verbal coding when demonstrating categorical perception. PMID- 11105524 TI - Functionality and spatial relations in memory and language. AB - We examined whether the functionality of spatial relations affects the construction and memory of information in situation models. A functional relationship involves the interaction of entities that is implied by either typical use or contextual demands. Previous research has shown that spatial relations are less likely to be encoded during comprehension unless there is extensive prior knowledge, explicit instructions to attend to spatial information, or a clear emphasis on spatial information. If the construction of a situation model is guided by a need to understand the functional structure of a situation, then functional spatial relations should be more likely to be encoded. The results of our study showed that sentences with functional spatial relations were read faster and remembered better in both recall and recognition tests than sentences with nonfunctional spatial relations. PMID- 11105525 TI - Letter-detection patterns in German: a window to the early extraction of sentential structure during reading. AB - Letters are more difficult to detect in function words than in content words, presumably because function words serve to cue sentential structure but recede to the background as meaning unfolds. This function disadvantage was found for the definite article in German for all three genders and all four cases, but it was more pronounced when the article appeared in a nominative noun phrase than in an object noun phrase. It was also more pronounced for the typical subject-predicate object sentential format than for the object-predicate-subject sentential format and also when the definite article unequivocally specified the case of a phrase than when it was ambiguous. The results suggest that the structural frames established on line in reading are finely tuned to both phrase-level and sentence level organization. PMID- 11105526 TI - The rereading effect: metacomprehension accuracy improves across reading trials. AB - Guided by a hypothesis that integrates principles of monitoring from a cue-based framework of metacognitive judgments with assumptions about levels of text representation derived from theories of comprehension, we discovered that rereading improves metacomprehension accuracy. In Experiments 1 and 2, the participants read texts either once or twice, rated their comprehension for each text, and then were tested on the material. In both experiments, correlations between comprehension ratings and test scores were reliably greater for participants who reread texts than for participants who read texts only once. Furthermore, in contrast to the low levels of accuracy typically reported in the literature, rereading produced relatively high levels of accuracy, with the median gamma between ratings and test performance being +.60 across participants from both experiments. Our discussion focuses on two alternative hypotheses--that improved accuracy is an artifact of when judgments are collected or that it results from increased reliability of test performance--and on evidence that is inconsistent with these explanations for the rereading effect. PMID- 11105527 TI - Effects of titles on the processing of text and lexically ambiguous words: evidence from eye movements. AB - Providing titles for passages improves the comprehension and memorability of text. Titles have generally been thought to facilitate comprehension at later stages of processing. Consistent with prior research, we found that passages presented with titles were better recalled than those without titles. Furthermore, in Experiment 1, the presence of titles led to fewer regressive eye movements, shorter end-of-sentence reading times, and shorter fixation times on target nouns. Experiments 2 and 3, using ambiguous target words, indicated that except when a very infrequent sense of a word is required, titles provide a strong enough context to allow for ambiguous words to be processed as quickly as control words. The results of the three experiments suggest that titles affect processing at both integrative and lexical stages of reading. PMID- 11105528 TI - The presence of an event in the narrated situation affects its availability to the comprehender. AB - Narrative descriptions of events often depart from how these events would have occurred in "real time." For example, narratives often contain time shifts in which events that are irrelevant to the plot are omitted. Zwaan (1996) has shown that these time shifts may affect on-line comprehension. Specifically, they are associated with increases in processing load and a deactivation of previous information. The experiments in the present article show that the situation is more complex. Specifically, there is only a deactivation of previous events if they are not assumed to be ongoing after a time shift. Furthermore, explicit discontinuations of events, as in he stopped walking also lead to deactivations when compared with explicit continuations and resumptions. PMID- 11105529 TI - Resource allocation during spoken discourse processing: effects of age and passage difficulty as revealed by self-paced listening. AB - The allocation of processing resources during spoken discourse comprehension was studied in a manner analogous to self-paced reading using the auditory moving window technique (Ferreira, Henderson, Anes, Weeks, & McFarlane, 1996). Young and older participants listened to spoken passages in a self-paced segment-by-segment fashion. In Experiment 1, we examined the influence of speech rate and passage complexity on discourse encoding and recall performance. In Experiment 2, we examined the influence of speech rate and presentation mode (self-paced vs. full passage presentation) on recall performance. Results suggest that diminished memory performance in the older adult group relative to the young adult group is attributable to age-related differences in how resources were allocated during the initial encoding of the spoken discourse. PMID- 11105530 TI - Plan formation, retention, and execution in prospective memory: a new approach and age-related effects. AB - Existing laboratory paradigms of prospective memory instruct subjects to remember to perform a single, isolated act at an appropriate point in the experiment. These paradigms do not completely capture many everyday complex prospective memory situations in which a series or set of delayed actions is planned to be executed in some subsequent period of time. We adapted a laboratory paradigm within which to study these prospective memory processes, and we investigated age related influences on these prospective memory processes. Age-related declines were found in the planning, initiation, and execution of the set of tasks. In contrast, there were no age differences in plan retention or in the fidelity with which the plan was performed. PMID- 11105531 TI - How to eliminate illusions in quantified reasoning. AB - The mental model theory postulates that reasoners build models of situations described in premises. These models normally make explicit only what is true according to the premises. The theory has an unexpected consequence. It predicts the existence of illusions in inferences: Certain inferences should have compelling but erroneous conclusions. Previous studies have corroborated the existence of such illusions. The present study reports the first effective antidote to them. For example, most people incorrectly answer "yes" to the following problem: Only one of the following statements is true ... /At least some of the plastic beads are not red./None of the plastic beads are red./Is it possible that none of the red beads are plastic? In two experiments, we progressively eliminated this fallacy and others by using instructions designed to overcome the bias toward truth. The difference between the illusory and the control problems disappeared when the participants were instructed to work out both the case in which the first premise was true and second premise was false and the case in which the second premise was true and the first premise was false. PMID- 11105532 TI - Reasoning versus text processing in the Wason selection task: a nondeontic perspective on perspective effects. AB - We argue that perspective effects in the Wason four-card selection task are a product of the linguistic interpretation of the rule in the context of the problem text and not of the reasoning process underlying card selection. In three experiments, participants recalled the rule they used in either a selection or a plausibility rating task. The results showed that (1) participants tended to recall rules compatible with their card selection and not with the rule as stated in the problem and (2) recall was not affected by whether or not participants performed card selection. We conclude that perspective effects in the Wason selection task do not concern how card selection is reasoned about but instead reflect the inferential text processing involved in the comprehension of the problem text. Together with earlier research that showed selection performance in nondeontic contexts to be indistinguishable from selection performance in deontic contexts (Almor & Sloman, 1996; Sperber, Cara, & Girotto, 1995), the present results undermine the claim that reasoning in a deontic context elicits specialized cognitive processes. PMID- 11105533 TI - Counterfactual thinking about controllable events. AB - When people think about what might have been, they mentally undo controllable rather than uncontrollable events. We report the results of two experiments in which we examined this controllability effect in counterfactual thinking. The experiments show that the mutability of controllable events is influenced by the perceived appropriateness or inappropriateness of the events. The first experiment shows that people change inappropriate controllable actions more than appropriate controllable ones. The second experiment shows that people mutate inappropriate controllable events whether the outcome is exceptional or normal with respect to intrapersonal habitual norms, and whether the outcome is positive or negative. We discuss the implications for alternative theories of counterfactual thinking. PMID- 11105534 TI - ADA Title III and the Internet: technology and civil rights. PMID- 11105535 TI - Review of comparative studies between conventional and liposomal amphotericin B (Ambisome) in neutropenic patients with fever of unknown origin and patients with systemic mycosis. AB - Fungal infections are an important cause of morbidity and mortality in immunocompromised patients. Treatment with amphotericin B is the main therapeutic approach. However, this treatment is limited by the substantial toxicity. We present the data of the first randomized prospective comparative trial in adults (134 patients with fever of unknown origin) with conventional amphotericin B and a liposomal formulation of amphotericin B (AmBisome, published in 1997 by Prentice et al. (Br. J. Haematol. 98, 711-718) and the data of adults with documented fungal infections (59 patients), treated in this trial. Patients received either conventional amphotericin B 1 mg kg-1 per day, liposomal amphotericin B 1 mg kg-1 per day or liposomal amphotericin B 3 mg kg-1 per day. Patients were entered if they had fever of unknown origin (FUO), defined as temperature of 38 degrees C or more, not responding to 96 h of systemic broad spectrum antibiotic treatment, and neutropenia (< 0.5 x 10(9) l-1). Efficacy of treatment was assessed, with success defined as resolution of fever for three consecutive days (< 38 degrees C) in the group of patients with FUO and the freedom of clinical signs and/or the elimination of fungus in the group of patients with documented fungal infections. The safety of treatment and renal and hepatic toxicity of liposomal and conventional amphotericin B were compared. No statistically significant difference was found in the treatment efficacy in the three study arms. However, there is a tendency of better treatment results in the two groups of patients, who received liposomal amphotericin B. Thirty-five per cent of patients with documented fungal infections and 46% of patients with FUO responded to amphotericin B. In the patients group, that received 1 mg kg-1 liposomal amphotericin B it was 63 and 49%, in the group of patients that received 3 mg kg-1 liposomal amphotericin B it was 47 and 64%. Evidence of toxicity due to amphotericin B was seen in 50 patients (83%), toxicity due to liposomal amphotericin B, 1 mg kg-1, was seen in 35 patients (50%), and due to liposomal amphotericin B 3 mg kg-1 in 34 patients (54%). This was a statistically significant difference (P = 0.001). It was concluded that liposomal amphotericin B was safer than conventional amphotericin B, but both formulations are equivalent in treatment efficacy. The prophylactic use of amphotericin B in these immunocompromised patients is discussed. PMID- 11105536 TI - Experimental coccidioidomycosis in hamsters. Disease kinetics and death curve in relation to infective dose. AB - A study of experimental coccidioidomycosis in hamsters (Mesocricetus auratus) is presented. Two experiments were conducted on 75 animals inoculated intracardially with the mycelial form of Coccidioides immitis. The first research (experiment I) studied the kinetics of experimental disease in 15 hamsters inoculated with 300 C. immitis arthroconidia. The parameters studied were: (a) presence of macroscopic lesions in the brain, lungs, liver, spleen and kidneys; (b) microscopic identification of spherules in wet mount preparations of these specimens; (c) samples from all organs cultured at 37 degrees C on Sabouraud glucose agar; (d) blood cultures drawn every 24 h during the first week and subsequently every 48 h and (e) histopathological studies of all organs. The second experiment (experiment II) determined the relationship between the inoculum size and death curve in six groups of 10 animals each, which had received doses of 10, 50, 100, 150, 200 and 300 arthroconidia, respectively. On day 14 post-inoculation, all the animals underwent skin tests and 1 ml of blood was obtained by cardiac puncture to detect antibodies. Disseminated disease with persistent fungaemia developed in all the studied animals. Coccidioides immitis was recovered from all organs, with the lungs being the first to present disease. Death occurred in all groups, regardless of the dose of arthroconidia and 83.3% died between day 22 and day 28 post-infection. The use of this model is proposed for the biological standardization of antigens, the study of prophylactic measures and the "in vivo" evaluation of new antifungal treatments. PMID- 11105537 TI - Multiple forms of the serine protease Alp of Aspergillus fumigatus. AB - Alkaline proteases were produced by a virulent strain of Aspergillus fumigatus during growth on media containing glucose and proteins or peptides. After isoelectric focusing, six bands of proteolytic activity were detected with synthetic substrates after blotting on nitrocellulose membranes. The main protease (pI = 8.6) corresponded to the known subtilisin-like protease Alp of A. fumigatus and five minor components had lower isoelectric points (8.1 to 5.2). All proteases were produced on different media and in various phases of growth with only small quantitative variations. They also had identical pH optima, were denatured above 45 degrees C and stabilized by Ca2+ ions, were affected by the inhibitors of serine proteases only and had nearly identical substrate specificity against 13 synthetic substrates. On gel chromatography the three most acidic components had higher molecular weights than the main enzyme Alp. It remains to be determined if the enzymes under study arise through posttranslational processing of the main protease or are true isoenzymes, products of a gene family. PMID- 11105538 TI - The frequency of Candida parapsilosis in onychomycosis. An epidemiological survey in Israel. AB - Candida albicans is regarded as the major pathogen in yeast-induced onychomycosis. Based on our impression of an increasing prevalence of Candida parapsilosis in this disease, we examined the data of two mycology laboratories in the same geographic location, from 1994 to 1996 in one (centre A) and for 1995 (6 months) in the other (centre B). A total of 954 and 230 toenails and 621 and 190 fingernails, respectively, underwent KOH microscopy and culture studies in each centre. Positive findings were noted in 45 and 65% of the toenails and 44 and 72% of the fingernails, respectively. In the toenails, Candida spp. were found in 22 and 15%, respectively, and in the fingernails, in 77 and 63%, respectively. The most frequent Candida species was C. parapsilosis (39.5% in toenails, 36.7% in fingernails), followed by C. albicans (19.5% in toenails, and 34.4% in fingernails). These results demonstrate a higher frequency of isolation of C. parapsilosis compared with C. albicans in onychomycosis. This might have important therapeutic implications. PMID- 11105539 TI - Double-blind randomized dose-finding study in acute vulvovaginal candidosis. Comparison of flutrimazole site-release cream (1, 2 and 4%) with placebo site release vaginal cream. AB - A double-blind randomized comparative phase II study of flutrimazole site-release vaginal cream (1, 2 and 4%) with placebo site-release vaginal cream was undertaken in patients with acute vulvovaginal candidosis. Vaginitis was demonstrated by both positive findings on microscopic examination of vaginal smears and positive culture as well as by the presence of clinical signs and symptoms. The vaginal monodose treatment was inserted in the evening at bedtime using a vaginal applicator and, in addition, all four groups of patients received additional topical external cream for application to the vulva twice-daily for 7 days; the placebo group received a placebo cream and the active therapy groups all received a 2% flutrimazole cream. A total of 133 patients who were seen over a 10-month period were screened and randomized: five patients did not take the allocated medication, and four patients whose menstrual period began shortly after study entry were excluded from the study, leaving 124 patients who were randomly allocated to receive a monodose vaginal 1% cream (regimen A, 28 patients), a monodose vaginal 2% cream (regimen B, 32 patients), a monodose vaginal 4% cream (regimen C, 31 patients) or a monodose vaginal placebo cream (regimen D, 33 patients). At the assessment 9 days after the end of therapy the proportion of patients who were cured was 82% in group A, 87.4% in group B, 83.8% in group C and 63.5% in group D. Three patients (10.7%) in group A, four (12.5%) in group B, one (3.2%) in group C and 12 (36.36%) in group D did not respond to the treatment. One patient (3.5%) in group A, and two patients (6.4%) in group C terminated the treatment prematurely due to intolerance. There was a significant association between Candida glabrata and treatment failure (P < 0.04) and C. glabrata and carrier state (P = 0.01) in vagina (chi 2 test, P = 0.01) and vulvovagina (chi 2 test, P = 0.00001). At the assessment 4 weeks after the end of therapy the proportion of cured patients was 60.6% in group A, 78% in group B, 80.6% in group C and 48.4% in group D. Group D (placebo) versus group B (2%) and group C (4%) showed a significant difference (P = 0.01 and P = 0.007, respectively). Although there were no significant differences in clinical and mycological activity between the three active groups, group B (flutrimazole 2% site-release vaginal cream) was chosen for clinical use due to its tolerance profile. Seven patients (25%) in group A, three (9.3%) in group B, two (6.4%) in group C and five (15.1%) in group D relapsed 4 weeks after the end of therapy; the relapse rate was not significantly associated with positive culture results 9 days after treatment. There was a significant association between C. glabrata and the carrier state (P < 0.01). The overall ineffective treatment (includes failures at control 1, relapses at control 2 and premature terminations) was 39% in group A, 21.7% in group B, 16% in group C and 51.3% in group D. There was a significant difference in the overall ineffective treatment when C and D groups were compared with placebo (P = 0.01 and P = 0.003, respectively). PMID- 11105540 TI - Case report. Pathohistological findings in a clinical case of disseminated infection with Fusarium oxysporum. AB - Despite appropriate antimicrobial and antifungal therapy (amphotericin B), a disseminated infection with Fusarium oxysporum in a 75-year-old immunocompromised patient (acute myeloid leukaemia, minimal leucocyte count of 0.5 giga l-1) led rapidly to death. A similarly fatal course of an F. oxysporum infection has been reported in several cases. Fusarium oxysporum could be isolated shortly before death from blood cultures and from a swab taken from skin efflorescences. An autopsy revealed histopathologically typical fungal infiltrates in the mucosa of the pharynx, epiglottis, trachea, and oesophagus and in the parenchyma of the spleen, the lung and both kidneys. Because of the high risk of a fatal outcome of this infection, the clinician should aim at maximum diagnostic enforcement. We propose both analysis of blood cultures and immediate skin biopsy--with PAS staining--of suspicious dermal efflorescences for microscopic examination. The treatment of choice is discussed controversially but a beneficial effect has been reported from granulocyte transfusion, subcutaneous administration of GM-CSF and concomitant treatment with amphotericin B. PMID- 11105541 TI - [Morphometric changes in the choriocapillaris and choroid in eyes with advanced glaucoma damage]. AB - BACKGROUND: In addition to elevated intraocular pressure, a compromised ocular blood supply has been implicated in the pathogenesis of primary open-angle glaucoma (PCOG). METHODS AND MATERIALS: We analyzed 20 eyebank eyes with end stage PCOG and compared these with 20 age-matched controls. The following variables were measured: density and diameter of large choroidal vessels in the macular and equatorial choroid; thickness of the choroid in the macular and equatorial region; and density and thickness of choriocapillaris in the macular, peripapillary, and equatorial choroid. RESULTS: Eyes with glaucoma displayed a lower density of the capillaries of the choriocapillaris than control eyes in the macular, temporal peripapillary, and equatorial choroid, with 0.50 vs. 0.55 (P = 0.018), 0.46 vs. 0.51 (P = 0.016), and 0.50 vs. 0.55 (P = 0.038), respectively. Assessment of large choroidal vessels in the macular choroid showed that eyes with glaucoma had less density of veins (11.7 vs. 38.9/mm2; P < 0.001) and arteries (7.7 vs. 12.4/mm2; P = 0.005) and arteries with a higher diameter (45.6 vs. 28.2 microns; P < 0.001) than control eyes. The large vessels in the equatorial choroid displayed no significant difference in diameter but a lower density (21.2 vs. 44.1/mm2; P = 0.017) in eyes with glaucomatous damage than controls. CONCLUSION: Eyes with advanced glaucomatous damage after long-standing PCOG exhibit many changes in the choroidal vasculature. We cannot conclude from our study whether the observed vascular changes in the choroid are primary pathogenic factors or secondary phenomena. PMID- 11105542 TI - [Intra-individual comparison of intraocular lenses of highly refractive silicone (Allergan SI40NB) and hydrophobic acrylate (Alcon Acrysof MA60BM). 1-year follow up]. AB - BACKGROUND: A prospective, randomized study was performed to evaluate intra individually the biocompatibility of foldable, highly refractive silicone and hydrophobic acrylic intraocular lenses (IOL). MATERIALS AND METHODS: We studied 35 patients who underwent phacoemulsification using a self-sealing tunnel incision. In a randomized fashion one eye received a 6-mm optic IOL made of high refractive index silicone (Allergan SI40NB) and the other eye a hydrophobic acrylic 6-mm optic IOL (Alcon AcrySof MA60BM). All patients were examined 7 days, 1-3 and 6 months, and 1 year postoperatively. RESULTS: The mean best-corrected visual acuity (BCVA) was 0.9 +/- 0.12 vs. 0.89 +/- 0.13 (SI40NB vs. MA60BM) after 1-3 months. One-year postoperatively BCVA was still 0.9 +/- 0.12 vs. 0.87 +/- 0.14. The flare values (photon counts/ms) increased slightly 7 days after surgery (14.2 +/- 8.68 vs. 15.49 +/- 7.2, n.s.). Three months after surgery these values were again in the normal range. The mean IOL decentration was 0.29 +/- 0.14 vs. 0.3 +/- 0.15 mm 1 year postoperatively. Scheimpflug slit photography showed 40% of MA60BM IOLs to have "glistenings." No significant difference regarding posterior capsular opacification was found. CONCLUSION: One year after implantation of foldable, highly refractive silicone and hydrophobic acrylic IOLs using a self-sealing tunnel incision and phacoemulsification, no significant functional or morphological differences between the two IOL types were observed. PMID- 11105543 TI - [Stereoscopic vision before and after cataract extraction with artificial lens implantation]. AB - BACKGROUND: Based on current insurance data patients with pseudophakia can be expected to suffer reduced stereoacuity. A prospective study was carried out to test this relationship. PATIENTS AND METHODS: Stereoacuity was measured by Titmus and Lang tests in 50 patients with cataracts who had no other ocular complaints and a minimal visual acuity of the more impaired eye of 20/50. Examinations were carried out before and for 3 days after cataract surgery (phacoemulsification, posterior chamber lens). RESULTS: Median stereoacuity improved from 60" preoperatively to 50" postoperatively in patients with bilateral phakia, from 90" to 45" in those with unilateral phakia. Results of the Lang test were difficult to interpret. CONCLUSION: Near stereoacuity is not worsened by unilateral or bilateral pseudophakia if there are no other complaints, and if the eyes are corrected for reading distance. PMID- 11105544 TI - [A retinoid constituent of lipofuscin, A2-E, is a photosensitizer in human retinal pigment epithelial cells]. AB - BACKGROUND: A fluorescent compound of lipofuscin, A2-E, has been shown to impair lysosomal function and to increase the intralysosomal pH of human retinal pigment epithelial (RPE) cells. This study addressed the phototoxic potential of A2-E on RPE cells. METHODS: A2-E accumulation was confirmed by fluorescence microscopy and fluorescence-activated cell sorter analysis. Acridine orange staining allowed assessment of lysosomal integrity and intralysosomal pH. Phototoxic properties of A2-E were determined by exposing A2-E-free and A2-E-fed RPE cell cultures to short-wavelength visible light and assessing cell viability and lysosomal integrity. RESULTS: Intralysosomal accumulation of A2-E was confirmed. Acridine orange staining showed that the A2-E was located in the lysosomal compartment and induced an elevation of intralysosomal pH. Exposure of A2-E fed cells to light resulted in a significant loss of cell viability by 72 h which was not observed in either RPE cells maintained in the dark or A2-E-free cultures exposed to light. Toxicity was associated with a loss of lysosomal integrity. CONCLUSION: A2 E is detrimental to RPE cell function by a variety of mechanisms including inhibition of lysosomal degrading capacity, loss of membrane integrity, and phototoxicity. Such mechanisms could contribute to retinal aging and to retinal diseases associated with excessive lipofuscin accumulation. PMID- 11105545 TI - [Acute retinpathia praematurorum. Is plasma prorenin level of prognostic value?]. AB - BACKGROUND: In patients with diabetes mellitus an elevated level of plasma prorenin (PP) may be associated with proliferative diabetic retinopathy. Although retinopathy of prematurity (ROP) is also characterized by retinal vasoproliferation, no study on PP in ROP appears to have been carried out. This study investigated PP prospectively in preterm infants with high risk of ROP. PATIENTS AND METHODS: In 304 preterm infants (gestational age 24-36 weeks, mean +/- SD 29.8 +/- 2.6 weeks; birth weight 570-1750 g, 1180 +/- 294 g) PP was examined prospectively between 3 and 14 weeks postnatally. Renin and total renin (after cryoactivation) were determined by radioimmunoassay. Total renin minus renin is the PP level; PP was correlated with the presence of ROP, stage of ROP, gestational age, birth weight, and postnatal age. RESULTS: There was no significant difference between mean PP in 112 preterm infants with ROP (682 +/- 666 ng/l) and that in 192 preterm infants without ROP (622 +/- 454 ng/l). There was no correlation between PP and the stage of ROP, gestational age, or birth weight. Mean PP decreased with increasing postnatal age (postnatal age 3-4 weeks: 906 +/- 587 ng/l; 7-8 weeks: 585 +/- 423 ng/l; 13-14 weeks: 326 +/- 205 ng/l). CONCLUSION: This study found no significant difference in PP between preterm infants with and those without ROP. Thus PP is not a valid predictor or indicator of ROP. However, the study showed a hitherto unknown correlation between PP and postnatal age in preterm infants. PMID- 11105546 TI - [Laser canaliculoplasty]. AB - BACKGROUND: Laser canalicular plastic surgery using the erbium-YAG laser is a new method for treating canaliculus stenosis. We compared results of this method with those of conventional surgical operations on canaliculi. PATIENTS AND METHODS: Between 1996 and 1998 a total of 44 cases of symptomatic canaliculus stenosis were treated by laser. An erbium-YAG laser with a maximum power of 100 mJ was used in conjunction with a sapphire fiber 3 cm long and 325 microns in diameter. Endoscopic diagnosis of the other parts of the lacrimal passage was carried out before and after laser treatment and after tubular intubation. After an average of 12 months the results were assessed via questionnaires and follow-up examinations. RESULTS: The success rate for reducing epiphora was 67%. Overall 86% of patients reported an improvement in canaliculus communis stenosis. CONCLUSIONS: Laser canalicular plasty as minimally invasive surgical treatment is an excellent surgical method for treating point-focal, noninflammatory canalicular stenosis. PMID- 11105547 TI - [Can rhegmatogenous retinal detachment be prevented? Reflections on the history of "prophylactic" treatment of retinal detachment]. AB - BACKGROUND: Less than 50% of clinical detachments are preventable, and the only known methods achieving this goal are, to promptly examine both retinas of every patient with symptoms of acute posterior vitreous detachment, and to promptly treat every definite tractional tear in such patients. PMID- 11105548 TI - [Sterilization of phacoemulsification and vitrectomy instruments. Contamination and evaluation]. AB - BACKGROUND: Contamination of automated surgical equipment is widely disregarded as a potential source of perioperative infection. We investigated the possibility of contamination of the aspiration fluid by the vacuum control manifold (VCM). The normal, unsterile internal VCM was compared with a modified external VCM that was regularly disinfected. MATERIALS AND METHODS: We investigated 37 aspiration fluid specimens from routine cataract and vitrectomy operations performed with automated evacuation systems. There were 25 specimens from three automated evacuation systems equipped with an internal VCM (experimental groups) and 12 specimens from one system equipped with a modified external VCM (control group). No hygiene procedures were used with the hidden internal VCM, but the modified external VCM was regularly rinsed and filled with 70% isopropanol overnight. Specimens were collected under sterile conditions, centrifuged, cultured for bacterial growth on blood agar and MacConkey agar for 24-48 h at 37 degrees C, and analyzed microbiologically. RESULTS: Aspiration fluids of irrigation/aspiration systems used for intraocular surgery were found to be severely contaminated with bacteria originating from the VCM. In all aspiration fluid specimens from internal VCM systems, 2(+)-4+ bacterial growth was found. Stenotrophomonas maltophilia (17), Comamonas acidovorans (8), and Agrobacterium radiobacter (13) were found most frequently. All specimens from the modified external VCM system remained sterile. There was a significant difference with regard to the frequency of contamination of the aspiration fluid between experimental and control groups (P = 0.0001, chi 2). CONCLUSIONS: We found that the aspiration fluid of common phaco- and vitrectomy systems was strongly contaminated by bacteria originating from the internal VCM. The technical modification of an external VCM allows easy disinfection and prevents contamination of the aspiration fluid. PMID- 11105549 TI - [Diagnosis and therapy of diffuse lamellar keratitis]. PMID- 11105550 TI - [Bilateral papilledema. Pseudotumor cerebri]. PMID- 11105551 TI - [Reduced secretion of tears after craniocerebral trauma. Keratoconjunctivitis sicca after skull base fracture by damage of parasympathetic innervation of the lacrimal gland]. PMID- 11105552 TI - [HIV infection and the eye]. PMID- 11105553 TI - [Effects of growth hormone replacement therapy in adults with severe growth hormone deficiency]. AB - The aim of the study was to analyse the effects of GH replacement therapy (1 year duration) on body composition, carbohydrate metabolism, thyroid hormone metabolism and bone mineral density in 8 adults with growth hormone deficiency (5 women, 3 men; mean age 40 years). Mean maintenance dose of GH was 1.5 IU/day-1.76 IU/day for women and 1.07 IU/day for men, respectively--determined according to individual patient requirements. Serum insulin-like growth factor-I standard deviation score increased from -5.4 to 0.0 (p < 0.001). There was a significant negative relationship between serum insulin-like growth factor-I standard deviation score at the start of therapy and the increase in this score (r = 0.85; p < 0.05). The waist:hip ratio decreased after 12 months by 0.039 (p < 0.05). The glycosylated hemoglobin increased (4.43 +/- 0.56% vs. 5.86 +/- 0.27; p < 0.05), and a negative correlation of the baseline glycosylated hemoglobin to the glycosylated hemoglobin increase was found (r = -0.88; p < 0.01). Both the free triiodothyronine and free triiodothyronine:free thyroxine ratio increased (3.09 +/- 0.22 vs. 4.17 +/- 0.40; p < 0.05, and 0.234 +/- 0.02 vs. 0.324 +/- 0.04; p < 0.01), and a positive relationship was observed between this ratio at the start of therapy and the increase in the ratio (r = 0.76, p < 0.05). The bone mineral density of lumbar spine and femoral neck expressed as z-score increased ( 1.18 +/- 0.56 vs. -0.75 +/- 0.48; p < 0.01 and -0.06 +/- 0.60 vs. 0.43 +/- 0.43; p < 0.05), while the bone mineral density of forearm was unchanged. CONCLUSIONS: Growth hormone replacement leads to a decrease in visceral fat, modulates the thyroid hormone levels by increasing peripheral conversion of thyroxine to triiodothyronine and probably is a physiological regulator of peripheral thyroxine metabolism, slightly deteriorates the carbohydrate metabolism, and results in an increase of bone mineral density of lumbar spine and femoral neck. PMID- 11105554 TI - [Growth hormone replacement therapy in adulthood]. PMID- 11105555 TI - [Threshold parameters in clinical practice]. AB - Certain parameters above a given intensity of an incremental exercise load are diverging from the linearity of the load and oxygen uptake. This intensity in Watts, speed or percent of the maximal oxygen uptake is called as "anaerobic threshold". The not fully understood events can reflect a relative local oxygen deficiency, levelling off the elimination of lactic acid, consecutive changes in ventilation, levelling off of the stroke volume and/or the macroerg phosphate turnover, uprising of the sympathoadrenal activation, changes in the activation of motor units etc. Empirically the "threshold" has been effectively applied by the performance physiologist and a growing body supports its use in the clinical diagnostics and (cardiac) rehabilitation. Some of the "threshold" parameters (acid-base status, lactic acid level, heart rate deflection) were easily available. Connected to the (spiro)ergometry, "thresholds" offer a valid, objective measure for fitness assessment and for monitoring of the effects of medicaments. It is rational to standardise certain diagnostic parameters to the "threshold" load. PMID- 11105556 TI - [Follow-up study of children after functional endoscopic sinus surgery]. AB - In the pathology of recidive infection of upper and lower respiratory tract the chronic diseases of the paranasal sinuses play an important role. The authors ordered for their FESS patients a control examination, 1 year after operation. They present examination data of 30 patients. They analyzed the respiratory complaints, the results of anterior rhinoscopy and CT, the allergies and the performed surgery. 17 patients were healed, 10 got better and in 3 cases the complaints were unchanged. Allergic rhinitis or bronchial asthma were found in more then 2/3 of their patients. According to their opinion the FESS is effective method in treatment of he chronic respiratory tract infections. In the background of the recidive respiratory tract inflammation the chronic diseases of the paranasal sinuses are often found if they are looked for. PMID- 11105557 TI - [Human insulin-induced lipoatrophy]. AB - The medical history of a 14-year-old diabetic adolescent female patient is presented. The patient has been exclusively treated by human insulin since the manifestation of diabetes at age of 11. As a clinical curiosity lipoatrophy developed at different sites of insulin injections (upper arm, thigh, abdominal wall, buttock). The insulin administration technique by pen-devices was correct. The patient proved to be non-atopic without signs of insulin allergy on intracutan tests. On histological examination, "lipoblastoma-like" alterations without signs of local immune mechanisms and no inflammatory cell infiltrates were found at the site of lipoatrophy. The histological findings suggested dedifferentiation of adipose tissue mediated probably by elevated local tumor necrosis factor-alpha. Immunological consequences of previous human insulin treatment were documented by high insulin-specific IgG and IgE antibody titer, however, no clinical signs of immunogenic insulin resistance were found. Switching to insulin analogue (insulin lispro) before main meals no further lipoatrophic areas were observed despite of using human NPH-insulin for basal insulin supplementation. Insulin analogue (insulin lispro) may be useful for treating diabetic patients with lipoatrophy secondary to previous human insulin treatment. PMID- 11105558 TI - [Vascular surgery at the turn of the Millennium]. PMID- 11105559 TI - [The year 1900 volume of the (Hungarian) Medical Weekly]. PMID- 11105560 TI - Inhibition of the transcription factor NF-kappa B by sesquiterpene lactones from Podachaenium eminens. AB - Investigation of Podachaenium eminens afforded nine sesquiterpene lactones (Sls) from which costunolide, 7-hydroxycostunolide, santamarin as well as 3 chlorodehydroleucodin are new for this plant and 3,4-dehydro-4 dehydroxypodachaenin (= 3-costoyloxydehydroleucodin) is found for the first time in nature. All isolated Sls were studied for their anti-inflammatory activity using the transcription factor NF-kappa B as a molecular target. NF-kappa B is involved in the synthesis of inflammatory mediators, such as cytokines and chemokines. Except for podachaenin, all compounds completely inhibited NF-kappa B DNA binding in an electrophoretic mobility shift assay at concentrations between 5 and 200 microM without showing any cytotoxic effects. 3,4-Epoxydehydroleucodin possessing an alpha-methylene-gamma-butyrolactone and a second reactive structure element by its epoxy ring alpha,beta to a carbonyl group was most active. Although the majority of the Sls tested in this study were monofunctional only low concentrations of 50 microM were often needed for complete inhibition. Possible reasons are discussed for this result. PMID- 11105561 TI - Effects of prenylated flavonoids and biflavonoids on lipopolysaccharide-induced nitric oxide production from the mouse macrophage cell line RAW 264.7. AB - Certain flavonoid derivatives possess anti-inflammatory activity in vitro and in vivo. Besides their antioxidative properties and effects on the arachidonic acid metabolism including cyclooxygenase/lipoxygenase inhibition, some flavones and flavonols were previously found to show inhibitory activity on nitric oxide production by inducible nitric oxide synthase (iNOS; NOS type 2) through suppression of iNOS induction. As part of our continuing investigations, the effects of unique and minor flavonoids (prenylated flavonoids and biflavonoids) on nitric oxide production from lipopolysaccharide-induced macrophage cell line (RAW 264.7) were evaluated in order to establish their inhibitory activity on NO production and correlate this action with their in vivo anti-inflammatory potential. Among the derivatives tested, prenylated compounds including morusin, kuwanon C, and sanggenon D and biflavonoids such as bilobetin and ginkgetin were found to inhibit NO production from lipopolysaccharide (LPS)-induced RAW 264.7 cells at > 10 microM. Inhibition of nitric oxide production was mediated by suppression of iNOS enzyme induction but not by direct inhibition of iNOS enzyme activity. An exception was echinoisoflavanone that inhibited iNOS enzyme activity (IC50 = 83 microM) and suppressed iNOS enzyme induction as well. While most prenylated derivatives showed cytotoxicity to RAW cells at 10-100 microM, all biflavonoids tested were not cytotoxic. Since nitric oxide (NO) produced by inducible NO synthase (iNOS) plays an important role in inflammatory disorders, inhibition of NO production by these flavonoids may contribute, at least in part, to their anti-inflammatory and immunoregulating potential in vivo. PMID- 11105562 TI - Kavain inhibits murine airway smooth muscle contraction. AB - This study examined the effect of kavain, the principle biologically active component of kava, on murine airway smooth muscle. In isolated isometrically contracted tracheal ring preparations, kavain was noted to diminish the maximal contractile response to both muscarinic receptor activation and voltage-operated calcium channel activation. The IC50 for kavain in rings precontracted with carbachol was found to be 177 microM +/- 53.1, and, in rings precontracted with KCl, it was found to be 59.6 microM +/- 10.1. In addition, pretreatment with kavain attenuated airway smooth muscle contraction evoked with either carbachol or KCl. The EC50 for KCl was not affected by kavain pretreatment. However, the EC50 for carbachol was significantly affected by a high kavain pretreatment dose. Nitric oxide mediated relaxation was not observed to play a role in kavain's smooth muscle relaxing properties. Similarly, prostaglandin pathways are not likely involved in these effects since pretreatment of tracheal rings with indomethacin before carbachol contraction did not reduce the relaxant effect of kavain. The mechanism of kavain-induced relaxation and inhibition of contraction is likely due to a mechanism common to both contractile agonists that were employed in our study. PMID- 11105563 TI - Pharmacological effects of urinary products obtained after treatment with saiboku to, a herbal medicine for bronchial asthma, on type IV allergic reaction. AB - To define the anti-allergic components in Saiboku-To, a herbal medicine for bronchial asthma, we examined the effects of 11 compounds found in post administrative urine of Saiboku-To on concanavalin A-induced human lymphocyte blastogenesis in vitro and picryl chloride (PC)-induced mouse ear swelling in vivo. The urinary products of Saiboku-To were flavonoids and lignans derived from the constitutional herbs and their hydrogenated metabolites. Medicarpin derived from Glycyrrhiza glabra, magnolol and 8,9-dihydroxydihydromagnolol from Magnolia officinalis, baicalein, wogonin and oroxylin A from Suctellaria baicalensis inhibited lymphocyte blastogenesis in dose-dependent fashion with IC50 values ranging from 3.0 to 7.7 micrograms/mL, which corresponded to 20-100 times that of prednisolone IC50 (0.08 microgram/mL). Davidigenin, dihydrowogonin and dihydrooroxylin A, which are hydrogenated metabolites of liquiritigenin, wogonin and oroxylin A, respectively, had no or little effects on lymphocyte blastogenesis. Oral administration of Saiboku-To, medicarpin, baicalein, magnolol and baicalin (100 mg/kg), inhibited PC-induced ear swelling significantly by 23.5, 40.1, 30.5, 23.6 and 20.9%, respectively, though the effects were weaker than that of 5 mg/kg of prednisolone (52.9%). The results suggested that flavonoids and lignans tested in the present study were implicated in anti asthmatic effect of Saiboku-To through suppression of type IV allergic reaction. PMID- 11105564 TI - Intra-specific variability of feverfew: correlations between parthenolide, morphological traits and seed origin. AB - Parthenolide, a biologically active sesquiterpene lactone found in feverfew [Tanacetum parthenium (L.) Schultz Bip., Asteraceae], has been indirectly linked to the anti-migraine action of feverfew preparations. Commercial preparations of feverfew leaves are known to vary widely in parthenolide content. Thirty-one feverfew accessions of diverse origin were examined for morphological traits and parthenolide content. Significant variation in parthenolide content was found among the populations. Mean parthenolide levels ranged from non-detectable to 1.68% +/- 0.97 (per dry weight) based upon HPLC-UV-MS. In general, feverfew plants grown from wild-collected seed from botanical gardens and the USDA accessions had higher mean parthenolide levels (0.72% +/- 0.57) than plants from commercial sources, including the generic material (0.34% +/- 0.23) and cultivars (0.35% +/- 0.40). Feverfew varieties with a light green/yellow leaf color had significantly higher mean parthenolide levels (1.61% +/- 0.61%) than darker leafed varieties. A significant positive correlation between days to anthesis and parthenolide content was observed. Parthenolide levels did not correlate with floral morphology. This study shows that further selection for improved horticultural attributes and natural product content has potential to improve feverfew for the botanical/medicinal plant industry. PMID- 11105565 TI - Coumarins and antiplatelet constituents from the root bark of Zanthoxylum schinifolium. AB - Further study on the chloroform-soluble portion of the root bark of Zanthoxylum schinifolium led to the isolation of eight new coumarins: methylschinilenol (1), hydroxyepoxycollinin I (2), 8-methoxyanisocoumarin H (3), hydroxyschininallylol (4), hydroxyepoxycollinin II (5), schinitrienin (6), schininallylone (7), and isoschinilenol (8), along with twenty-six known compounds including fourteen coumarins, and nine alkaloids. The structural elucidation was determined by spectroscopic data. Among the isolates, terpenyl-coumarins and furoquinolines were the active constituents with antiplatelet aggregation in vitro and collinin (10) showed significant anti-HBC DNA replication activity. PMID- 11105566 TI - Gallotannins and related polyphenols from Pistacia weinmannifolia. AB - Two new gallotannins, pistafolins A (1) and B (2), were isolated from the leaf extract of Pistacia weinmannifolia. Their structures were determined by spectral methods. Four known gallotannins (3-6), seven known flavonoid glycosides (7-13), along with 1-O-beta-D-(6'-O-galloyl)-glucopyranosyl-3-methoxy-5-hydroxybenzen e (14), gallic acid (15), methyl gallate (16), (+)-catechin (17), and (+) gallocatechin (18), were also isolated. Some of these compounds were tested for their cytotoxicity toward K562 cells, and two small molecular phenolic compounds, 15 and 18, showed significant inhibitory effects with IC50 values less than 5 micrograms/ml. PMID- 11105567 TI - Cardioactive terpenoids and a new rearranged diterpene from Salvia syriaca. AB - From the roots of Salvia syriaca L., in addition to known terpenoids, a new rearranged diterpene, named salvisyrianone, was isolated. The structure of the new compound was assigned from spectral data. The crude extract and the single compounds were tested for cardiovascular parameters using Wistar Albino rats. Antihypertensive activity was established in the crude extract of the roots as well as in two compounds, ferruginol and 3 beta-hydroxystigmast-5-en-7-one. PMID- 11105568 TI - Bioactive triterpenoids from the stem bark of Picea glehni. AB - Two new triterpenoids, 3 alpha-methoxyserrat-14-en-21 beta-yl formate (1), and 24 methylcycloartenone (2), were isolated from the stem bark of Picea glehni (Fr. Schm.) Masters together with three known triterpenoids, 3 alpha-methoxyserrat-14 en-21 beta-ol, 3 beta-methoxyserrat-14-en-21 beta-ol, and piceanonol A. Compounds 1, 2, and a synthetic sample, 3 alpha-methoxyserrat-13-en-21 beta-yl formate showed potent inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). PMID- 11105569 TI - Direct analysis and identification of triterpene glycosides by LC/MS in black cohosh, Cimicifuga racemosa, and in several commercially available black cohosh products. AB - A method to directly identify triterpene glycosides using reversed-phase liquid chromatography with positive atmospheric pressure chemical ionization mass spectrometry (LC/(+)APCIMS) was developed. Based on the analysis of the molecular weight, fragment ions, selected ion chromatograms, a number of triterpene glycosides, including actein, 27-deoxyactein, cimicfugoside M, and cimicifugoside, from Cimicifuga racemosa were studied. A chromone, cimifugin, from C. foetida was also identified. Cimicifugoside M and cimifugin can specifically serve as indicators for species identification. The method can, therefore, be used to distinguish black cohosh products from among different plant species for quality control purposes. PMID- 11105570 TI - Inhibitory activity of stilbenes from medicinal plants on the expression of cell adhesion molecules on THP1 cells. AB - The inhibitory activity of stilbenes isolated from medicinal plants on cell adhesion molecules on the surface of THP-1 human monocytic cell lines was investigated. Among ten stilbenes tested, four stilbenes displayed a significant inhibitory activity on the expression of both intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). A cell-to-cell adhesion assay showed that 3,5-dihydroxy-4'-methoxystilbene and 2,3,4',5 tetrahydroxystilbene-2-O-beta-D-glucopyranoside as well as resveratrol blocked significantly TNF-alpha-inducing cell-cell adhesion between human umbilical vein endothelial cells (HUVEC) and THP-1 cells. PMID- 11105571 TI - Inducible nitric oxide synthase inhibitors from Saposhnikovia divaricata and Panax quinquefolium. AB - A series of polyacetylenes, falcarinone, panaxynol, falcarindiol, panaxydol, and panaxytriol, were isolated from Saposhnikovia divaricata (Turcz.) Schischk and Panax quinquefolium L. These polyacetylenes were identified as active principles on the inhibition of nitrite production by inducible nitric oxide synthase (iNOS). Treatment with 10 microM of panaxynol, falcarindiol, panaxydol and panaxytriol decreased the LPS/IFN-gamma-stimulated accumulation of nitrite by 71.92 +/- 3.07, 69.95 +/- 3.68, 45.48 +/- 6.11 and 36.85 +/- 8.80%, respectively. The IC50 value of falcarinone, panaxynol, falcarindiol, panaxydol and panaxytriol was > 20, 2.23, 1.98, 6.58 and 9.85 microM, respectively. PMID- 11105572 TI - Antimicrobial activity and chemical composition of the bark oil of Croton stellulifer, an endemic species from S. Tome e Principe. AB - The composition and the antimicrobial activity of the bark oil of Croton stellulifer, an endemic and rare species of these islands (S. Tome and Principe) are reported. Analysis was carried out by GC, GC/MS and 13C-NMR. The major constituents were alpha-phellandrene (15.4-18.6%), p-cymene (14.4-17.7%), linalool (12.0-12.6%) and alpha-pinene (8.1-9.1%). Kessane, a sesquiterpenoid oxide, not yet reported in the genus Croton, was identified by NMR. The essential oil of C. stellulifer was active against both bacterial and fungal strains, except Aspergillus niger. PMID- 11105573 TI - Relaxant effects of petasins in isolated guinea pig trachea and their structure activity relationships. AB - In the present study, we attempted to compare four petasins, isolated from Petasites formosanus Kitamura, and to look for structure-activity relationships, which may be helpful for synthesizing more active compounds for the treatment of asthma. Four petasins, including petasin, isopetasin, S-petasin and S-isopetasin, concentration-dependently relaxed histamine (10 microM)-, carbachol (0.2 microM) , KCl (30 mM)-, and leukotriene D4 (10 nM)-induced precontractions of isolated guinea pig trachealis. The IC50 values strongly showed that the relaxant effects of the sulfur-containing petasins, S-petasin and S-isopetasin, were more potent than those of non-sulfur-containing petasins, petasin and isopetasin. S isopetasin, with IC50 values around 10 microM, selectively relaxed carbachol- and KCl-induced precontractions, and had almost no effects (IC50s > 300 microM) on histamine- and leukotriene D4-induced precontractions. However, S-petasin, with IC50 values about 6-9 microM, non-selectively relaxed the precontractions induced by all these contractile agents. The influence of isomerization of either petasin to isopetasin or S-petasin to S-isopetasin on the relaxant effects is not clear. PMID- 11105574 TI - Acute intoxication of cyclosporin caused by coadministration of decoctions of the fruits of Citrus aurantium and the Pericarps of Citrus grandis. AB - In order to investigate the effects of Citrus herbs on cyclosporin absorption and disposition, swine were given cyclosporin (10 mg/kg) with or without decoctions of Citri Aurantii Fructus (CAF) or Citri Grandis Pericarpium (CGP) in a crossover design. FPIA method (fluorescence polarization immunoassay) was used to determine the blood concentration of cyclosporin. The decoctions were characterized by their flavanone contents. Our results indicated that the coadministration of CAF and CGP significantly increased the Cmax of cyclosporin by 64% and 79%, respectively. The AUC of cyclosporin was significantly elevated by 97% when coadministered with CGP. Among the swine, 1/5 and 3/5 exhibited acute toxicity of cyclosporin after concomitant intake of CAF and CGP, respectively. This indicates an interaction of Citrus compounds with a commonly used drug. We suggest when cyclosporin is coadministered with these Citrus decoctions, the blood concentration of cyclosporin should be carefully monitored for dose adjustment to avoid cyclosporin intoxication. PMID- 11105575 TI - Divergence of the indole alkaloid pattern in two somatic hybrid plant cell subcultures of Rauvolfia serpentina x Rhazya stricta. AB - The alkaloid pattern in Rauvolfia serpentina x Rhazya stricta somatic hybrid cell subcultures R x R17 K was studied and 11 compounds were identified on the basis of their spectral data. Among them, 1,2-dehydroaspidospermidine, rhazinilam, stemmadenine and tabersonine were reported as typical of Rhazya species while vomilenine and sarpagine are characteristic of Rauvolfia alkaloid metabolism. The alkaloid pattern in R x R17 K subcultures was compared with that in the other hybrid cell subcultures (R x R17 M) which were developed from the same origin hybrid cultures but have been maintained separately for about ten years. The data presented here exhibit pronounced divergence of the alkaloid patterns in R x R17 K and R x R17 M cell subcultures. PMID- 11105576 TI - Genetic heterogeneity of ribosomal RNA gene and matK gene in Panax notoginseng. AB - Previously, 185 ribosomal RNA gene and matK gene sequences of Chinese herbal medicines, Ginseng Radix, Panacis Japonici Rhizoma and Panacis Quinquefolli Radix were shown to correspond with those of the original plants, Panax ginseng, P. japonicus and P. quinquefolius, respectively, with the species-specific sequences especially for 18S rRNA gene sequences. In P. notoginseng and its derivative, Notoginseng Radix, however, we found two genetic groups with respect to both gene sequences. Five base substitutions were detected on both gene sequences and the homology between two groups was 99.7% for the 18S rRNA gene and 99.6% for the matK gene, respectively. One genetic group was found to have the identical sequences as those of P. ginseng. PMID- 11105577 TI - Phenylpropanes from Acorus tatarinowii. AB - In addition to a number of known compounds, four new phenylpropanes isoacoramone, (cis) epoxyasarone, (threo) 1',2'-dihydroxyasarone and (erythro) 1',2' dihydroxyasarone were obtained from the roots of Acorus tatarinowii ("Shi-Chang Pu" in Chinese). The later two isomers were obtained as a mixture. However, all the chemical shifts of the protons and carbons for these two components were assigned by 1D- and 2D-NMR techniques. PMID- 11105578 TI - Three lycopodium alkaloid N-oxides from Huperzia serrata. AB - Three lycopodium alkaloid N-oxides, huperzine J (1), huperzine K (2) and huperzine L (3) were obtained from Huperzia serrata (Thunb.) Trev. Their structures were elucidated by spectroscopic methods and the relative configurations of 1 and 3 were established via NOESY NMR observations; optical rotation values and CD data are presented. PMID- 11105579 TI - Naphthoquinone glucosides of Drosera gigantea from in vitro cultures. AB - From the shoots of Drosera gigantea propagated under in vitro culture, the rare naphthoquinone glucosides droserone (3,5-dihydroxy-2-methyl-1,4-naphthoquinone) and hydroxydroserone (3,5,8-trihydroxy-2-methyl-1,4-napthoquinone) 5-O-beta glucosides, together with the free naphthoquinones droserone, hydroxydroserone and plumbagin were isolated. The structures of the glucosides and hydroxydroserone were studied by 2D NMR techniques. The glucosides appear to be responsible for the brown-red (maroon) colour of this plant. Of the other naphthoquinones typical for the family Droseraceae only hydroplumbagin glucoside could be detected, whereas the presence of 7-methyljuglone and rossoliside (= 7 methylhydrojuglone glucoside) was excluded. PMID- 11105580 TI - Essential oil of Phlomis lanata growing in Greece: chemical composition and antimicrobial activity. AB - The essential oil obtained from the aerial parts of Phlomis lanata has been analyzed by GC/MS. 48 compounds representing 96.85% of the oil were identified; alpha-pinene, limonene and trans-caryophyllene were found as its main components. The essential all showed a moderate in vitro activity against six Gram (+/-) bacteria and a stronger one against the three tested pathogenic fungi. PMID- 11105581 TI - Composition and antimicrobial activity of the essential oil of Scutellaria albida ssp. albida from Greece. AB - Steam distilled essential oil from aerial parts of Scutellaria albida ssp. albida was analyzed by GC and GC/MS. Fifteen compounds were identified of which linalool (52.63%) and trans-nerolidol (9.03%) were the major constituents. Furthermore, the oil was tested against four bacteria and two yeasts and was found to be moderately active against all microorganisms tested. PMID- 11105582 TI - A new bisabolene derivative from the essential oil of Prangos uechtritzii fruits. AB - The fruits of endemic Prangos uechtritzii Boiss. & Hausskn. (Umbelliferae) were subjected to hydrodistillation and microdistillation. The resulting volatiles were investigated by GC-MS to determine the composition of the essential oils. 109 compounds representing 86.7% and thirty-two compounds representing 90.0% were identified and isolated by two different techniques, respectively. Column chromatography of the essential oil yielded a new bisabolene ether (7-epi-1,2 dehydrosesquicineole), which was characterized by spectral methods (GC-FTIR, 1D-, 2D NMR and HRESIMS). PMID- 11105583 TI - On the stability of sesquiterpene lactones in the officinal Arnica tincture of the German pharmacopoeia. AB - An investigation of sesquiterpene lactone content in Arnica tincture (German Pharmacopoeia) after storage for three years at different temperatures (+4, +25 and +30 degrees C) was carried out by GC and GC-MS analysis. A decrease (13, 32 and 37%, respectively) in the content of the main active compounds in this preparation, 11 alpha,13-dihydrohelenalin esters, correlating with storage temperature was found. This change was shown to be caused by addition of ethanol to the cyclopentenone structure of these molecules leading to 2-ethoxy-2,3,11,13 tetrahydrohelenalin derivatives. PMID- 11105584 TI - New lanostanoids from Ganoderma lucidum that induce NAD(P)H:quinone oxidoreductase in cultured hepalcic7 murine hepatoma cells. AB - Two new lanostanoids were isolated from the basidiocarp of Ganoderma lucidum and were identified as 26,27-dihydroxy-5 alpha-lanosta-7,9(11),24-triene-3,22-dione (1) and 26-hydroxy-5 alpha-lanosta-7,9(11),24-triene-3,22-dione (2) by their respective spectral data. Crude extracts and the isolated compounds were tested for their potential to induce NAD(P)H:quinone oxidoreductase (QR), a phase 2 drug metabolizing enzyme, as an approach to detect potential cancer chemopreventive activity. Compound 2 doubled the specific activity of QR at a concentration of 3.0 micrograms/ml, whereas compound 1 was significantly less active (1.7-fold induction at 20 micrograms/ml). In addition, both compounds weakly inhibited sheep vesicle cyclooxygenase 1 activity at a test concentration of 40 micrograms/ml. PMID- 11105585 TI - Triterpenoids, p-coumaric acid esters and flavonoids from Artemisia igniaria. AB - Twenty-eight components were detected from the extract of Artemisia igniaria, which included four triterpenoids, eight p-coumaric acid long chain alkyl esters, seven flavonoids and nine common plant constituents. Their structures were determined by spectroscopic methods. This is the first recorded instance of beta glutinanol and cis-p-coumaric acid eicosanyl ester occurring in nature. PMID- 11105586 TI - Gene transfer strategies for improving radiolabeled peptide imaging and therapy. AB - Utilization of molecular biology techniques offers attractive options in nuclear medicine for improving cancer imaging and therapy with radiolabeled peptides. Two of these options include utilization of phage-panning to identify novel tumor specific peptides or single chain antibodies and gene transfer techniques to increase the number of antigen/receptor sites expressed on malignant cells. Our group has focused on the latter approach for improving radiolabeled peptide imaging and therapy. The most widely used gene transfer vectors in clinical gene therapy trials include retrovirus, cationic lipids, and adenovirus. We have utilized adenovirus vectors for gene transfer because of their ability to accomplish efficient in vivo gene transfer. Adenovirus vectors encoding the genes for a variety of antigens/receptors (carcinoembryonic antigen, gastrin-releasing peptide receptor, somatostatin receptor subtype 2 (SSTr2)) have all shown that their expression is increased on cancer cells both in vitro and in vivo following adenovirus infection. Of particular interest has been the adenovirus encoding for SSTr2 (AdCMVSSTr2). Various radioisotopes have been attached to somatostatin analogues for imaging and therapy of SSTr2-positive tumors both clinically and in animal models. The use of these analogues in combination with AdCMVSSTr2 is a promising approach for improving the detection sensitivity and therapeutic efficacy of these radiolabeled peptides against solid tumors. In addition, we have proposed the use of SSTr2 as a marker for imaging the expression of another cancer therapeutic transgene (e.g. cytosine deaminase, thymidine kinase) encoded within the same vector. This would allow for non-invasive monitoring of gene delivery to tumor sites. PMID- 11105587 TI - Engineering membrane proteins for nuclear medicine: applications for gene therapy and cell tracking. AB - Nuclear imaging techniques such as PET and SPECT imaging are expected to play major roles in evaluating the efficacy of in vivo gene therapy. In particular, the quantification of vector delivery and imaging the efficacy of gene expression are of key interests in testing new treatment paradigms and in designing novel vectors. In this review article we illustrate how nuclear imaging can be used to image novel cell-surface expressed fusion proteins and how this strategy can be used to probe for phenotypic changes in genetically manipulated cells. Since the described approach uses new fusion proteins, typically not present on eukaryotic cells, such "artificial receptors" can be designed to bind radioisotopes currently in clinical use. The described fusion proteins consist of 1) a binding domain such as a peptide based chelator that binds 99mTc oxotechnetate and 2) a membrane anchoring domain. A variety of fusion proteins have been tested so far and the most promising one to date consists of a metallothionein (MT)-derived C terminal peptide fused to a type II membrane protein markers containing the N terminal membrane anchoring domain of neutral endopeptidase (PEP). Cell-surface expression of MT in transfected cells has been demonstrated using monoclonal antibodies in vitro. Both in vitro and in vivo transchelation experiments have confirmed expression of 99mTc-binding sites in eukaryotic cells. We expect the described approach to evolve into a useful strategy to "tag" transfected cells with 99mTc and thus assessing efficiency of gene delivery and expression. PMID- 11105588 TI - In vivo antisense imaging. AB - Antisense oligonucleotides, in short antisense, are small chains of nucleic acids capable to bind to cellular ribonucleic acid (RNA) by a hybridization mechanism. In vitro, antisense are widely used as reagents to detect or block specific RNA sequences. The use of antisense as in vivo diagnostic agents is attractive because it would bring molecular imaging at the level of gene expression. However, oligonucleotides are non-canonical radiopharmaceuticals and much progress is needed to adapt them to in vivo imaging. The requirements to reach this goal include improvements in radiosynthesis, stability, targeting, and specific and non-specific binding. They will be examined in this review together with the current achievements in the applications of antisense as nuclear medicine radiopharmaceuticals. PMID- 11105589 TI - Development of DNA-based radiopharmaceuticals carrying Auger-electron emitters for anti-gene radiotherapy. AB - Targeting of radiation damage to specific DNA sequences is the essence of antigene radiotherapy. This technique also provides a tool to study molecular mechanisms of DNA repair on a defined, single radiodamaged site. We achieved such sequence-specific radiodamage by combining the highly localized DNA damage produced by the decay of Auger-electron-emitters such as 125I with the sequence specific action of triplex-forming oligonucleotides (TFO). TFO complementary to polypurine-polypyrimidine regions of human genes were synthesized and labeled with 125I-dCTP by the primer extension method. 125I-TFO were delivered into cells with several delivery systems. In addition, human enzymes capable of supporting DNA single-strand-break repair were isolated and assessed for their role in the repair of this lesion. Also, the mutagenicity and repairability of 125I-TFO induced double strand breaks (DSB) were assessed by repair of a plasmid possessing a site-specific DSB lesion. Using plasmids containing target polypurine-polypyrimidine tracts, we obtained the fine structure of sequence specific DNA breaks produced by decay of 125I with single-nucleotide resolution. We showed that the designed 125I-TFO in nanomolar concentrations could bind to and introduce double-strand breaks into the target sequences in situ, i.e., within isolated nuclei and intact digitonin-permeabilized cells. We also showed 125I-TFO-induced DSB to be highly mutagenic lesions resulting in a mutation frequency of nearly 80%, with deletions comprising the majority of mutations. The results obtained demonstrate the ability of 125I-TFO to target specific sequences in their natural environment--within eucaryotic nucleus. Repair of 125I-TFO induced DNA damage should typically result in mutagenic gene inactivation. PMID- 11105590 TI - Designer genes: recombinant antibody fragments for biological imaging. AB - Monoclonal antibodies (MAbs), with high specificy and high affinity for their target antigens, can be utilized for delivery of agents such as radionuclides, enzymes, drugs, or toxins in vivo. However, the implementation of radiolabeled antibodies as "magic bullets" for detection and treatment of diseases such as cancer has required addressing several shortcomings of murine MAbs. These include their immunogenicity, sub-optimal targeting and pharmacokinetic properties, and practical issues of production and radiolabeling. Genetic engineering provides a powerful approach for redesigning antibodies for use in oncologic applications in vivo. Recombinant fragments have been produced that retain high affinity for target antigens, and display a combination of rapid, high-level tumor targeting with concomitant clearance from normal tissues and the circulation in animal models. An important first step was cloning and engineering of antibody heavy and light chain variable domains into single-chain Fvs (molecular weight, 25-27 kDa), in which the variable regions are joined via a synthetic linker peptide sequence. Although scFvs themselves showed limited tumor uptake in preclinical and clinical studies, they provide a useful building block for intermediate-sized recombinant fragments. Covalently linked dimers or non-covalent dimers of scFvs (also known as diabodies) show improved targeting and clearance properties due to their higher molecular weight (55 kDa) and increased avidity. Further gains can be made by generation of larger recombinant fragments, such as the minibody, an scFv-CH3 fusion protein that self-assembles into a bivalent dimer of 80 kDa. A systematic evaluation of scFv, diabody, minibody, and intact antibody (based on comparison of tumor uptakes, tumor:blood activity ratios, and calculation of an Imaging Figure of Merit) can form the basis for selection of combinations of recombinant fragments and radionuclides for imaging applications. Ease of engineering and expression, combined with novel specificities that will arise from advances in genomic and combinatorial approaches to target discovery, will usher in a new era of recombinant antibodies for biological imaging. PMID- 11105591 TI - Antibody phage display applications for nuclear medicine imaging and therapy. AB - Antibody-based constructs genetically engineered from genes of diverse origin provide a remarkable opportunity to develop functional molecular imaging techniques and specific molecular targeted radionuclide therapies. Phage display libraries of antibody fragment genes can be used to select antibody-based constructs that bind any chosen epitope. A large naive human antibody-based library was used to illustrate binding of antibody constructs to a variety of common and unique antigens. Antibody-based libraries from hybridoma cells, lymphocytes from immunized humans or from mice and human antibody repertoires produced in transgenic mice have also been described. Several orders of magnitude of affinity enhancement can be achieved by random or site specific mutations of the selected binding peptide domains of the scFv. Affinities (Kd) as high as 10( 11) M (10 pM) for affinity-matured scFv have been documented. Such gene libraries thus offer an almost limitless variety of antibody-based molecular binding peptide modules that can be used in creative ways for the construction of new targeting agents for functional or molecular imaging and therapy. PMID- 11105592 TI - [Image of the month. Lipoma of the trigeminal nerve]. PMID- 11105593 TI - [How I treat: from specialized pharmacology to drug therapy: a plea for an optimal educational program for rational therapeutics, from decision making to drug prescription]. AB - Clinical pharmacology and therapeutics are two complementary disciplines which should lead the medical student, through an optimized training, to a rational prescription of drugs, ultimate and important step of the medical approach. Such a learning should occur progressively throughout the medical education, focusing, first, on the therapeutic reasoning ("why?") and, second, on the practical application leading to the prescription ("how?"). The medical student should learn the difficult task of integrating disease, drug and patient, in order to optimize the benefit/risk ratio, while being informed about new concepts such as "Evidence-Based Medicine" and pharmacoeconomics. PMID- 11105594 TI - [Clinical case of the month. Association between medullary thyroid cancer and type 3 autoimmune thyroiditis: discussion of an unusual case]. AB - A 67 year old woman was found to have Grave's disease associated with medullary carcinoma of the thyroid. She was treated by surgery and hormone replacement therapy by Elthyrone. This case shows possible associations between thyroid carcinoma and Grave's disease. PMID- 11105595 TI - [An update of diagnostic and therapeutic procedures in Alzheimer disease]. AB - Early diagnosis of Alzheimer's disease is difficult, but neuropsychological tests become quite sensitive, and neuroimaging gives useful information. Criteria for differential diagnosis appear in the literature, but vascular dementia remains an ill-defined concept. There are ongoing researches on possible therapeutic interferences with formation of histological brain lesions. Meanwhile, treatments are symptomatic and should be started as early as possible. Both inhibitors of acetylcholinesterase and cognitive rehabilitation in day-care centres fulfill therapeutic criteria for dementia, i.e. improvement or stabilization of cognitive impairment and daily life activities. PMID- 11105596 TI - [Treatment of Crohn disease in adults with tumor necrosis factor-alpha (TNF alpha) antibodies]. AB - Crohn's disease (CD) is a chronic inflammatory disease of the bowel characterized by segmental transmural inflammation and granulomatous changes. TNF alpha is a member of a large family of proteins and receptors that are involved in immune regulation. It is a proinflammatory and immunoregulatory cytokine synthesized by monocytes, macrophages, and T cells. TNF alpha plays an early central role in the cytokine cascade of the inflammatory process. Recently, chimeric monoclonal antibodies that inhibits TNF alpha have been used in the treatment of Crohn's disease. Infliximab has been the most largely used antibody. It is commercialized in the USA and has recently obtained an European marketing approvement. Infliximab is indicated for the treatment of moderately to severely active CD in patients having an inadequate response to conventional therapy. Clinical trials have demonstrated efficacy when the agent is initiated as a 5 mg/kg single intravenous infusion. In patients with fistulizing CD, administration of 2 subsequent 5 mg/kg doses 2 and 6 weeks after the initial dose appears to be efficacious. Limited clinical data also suggest that infliximab retreatment regimen restores response and maintains remission rates. Infliximab appears to be well tolerated. To date, very little is known about the potential for long-term toxicity with infliximab therapy. PMID- 11105597 TI - [Cochlear implants]. AB - A cochlear implant is a technologically advanced medical device that simulates sound in the cochlea by electrically stimulating the hearing nerve. Cochlear implants are designed to help severely or totally deaf individuals who gain little or no benefit from hearing aids. Hearing aids and assistive listening devices amplify sounds. The sounds produced by even the most sophisticated hearing aids may not offer much benefit to people with a severe-to-profound or profound hearing loss in both ears. A cochlear implant is an electronic device that provides the function of the damaged or absent hair cells by providing electrical stimulation to the remaining nerve fibers. The implant provides useful hearing and improved communication abilities to the implant user. PMID- 11105598 TI - [The value and limits of intervertebral cages in lumbar arthrodesis]. AB - Cages are not disk prosthesis, they not preserve intervertebral mobility. In selected cases they are useful to improve immediate intervertebral stability when lumbar interbody fusion is performed, they also avoid the risk of bone graft crushing before fusion is obtained thanks to their rigidity. In most cases, additional instrumentation is mandatory. A perfect surgical procedure is required to avoid specific complication and failure. The use of cages does not preclude the use of more classical spinal fusion procedures, they have specific indications and do not replace former techniques. PMID- 11105599 TI - [Tattoos. From ritual to ornament with their complications]. AB - Tattoos are performed for distinct purposes which range from social rituals to aesthetic aims. They are permanent or transient according to the site of the pigments being located either in the dermis or in the stratum corneum, respectively. Tattoos may be responsible for allergic reactions, infections or psychological distress of the subject who requests to be relieved of it. PMID- 11105600 TI - [Post-Doc (post-graduate training for medical doctors in Europe). The use of the Internet in the continuing education of general practice physicians: a European project of multimedia continuing medical education. The University of Aachen, Maastricht, Catholic University Leuven and the Central University Limburg (Diepenbeek) and EUREGIONET]. AB - The continuing formation of the General Practitioners (GP) must stay in stride with the rapid evolution of society, technology, science and needs of the population. The Web allows on-line a rapid access to a pertinent and practical information whenever needed. PostDoc is a joint venture of the Universities of Aachen, Maastricht, Leuven and Diepenbeek. The Department of general medicine is associated with the Service of Technology of Education in this project. The interactivity of an Internet Website introduces a cooperative dimension in the work of the GP's centred on formation and discussion of problems encountered. This tool allows each GP to contribute to clinical cases and information. PMID- 11105601 TI - [Current therapeutics 2000]. AB - The most important drugs registered and/or launched in Belgium during the last year in the various disciplines of internal medicine will be briefly described. The originality of each molecule as well as its modalities of appropriate use in clinical practice will be emphasized. PMID- 11105602 TI - [Melatonin. II. Physiological and therapeutic effects]. AB - Melatonin is a hormone mainly secreted by the pineal gland during the dark phase of the light-dark cycle. The most known function of melatonin in mammals is to transmit information concerning light-dark cycles playing the role of an active neuroendocrine transducer of environmental information. Given this chronobiologic role of pineal melatonin, it seems to be useful in the management of shift work, jet lag and some sleep disorders. In vitro like in vivo melatonin seems to be effective as an antioxidant and oncostatic agent. Melatonin may provide protection against aging process, degenerative diseases, cancer and play a role also in sexual maturation, reproduction, immune function and psychiatric illness. The administration of melatonin in the jet-lag syndrome is well codified. Further clinical research is needed for a better understanding and definition of other indications, treatment regimens and safety of the hormone. The aim of this paper is to review the current knowledge on its clinical implications. PMID- 11105603 TI - [Ethological approach to seasonal depression and manic-depressive psychoses]. AB - Some psychotherapists disagree with the importance of seasonal factors and sunlight in the etiology of depressions and bipolar disorders. Depression in spring and summer, mania in autumn and winter do not seem consistent with the expected effects of light on mood. Ethology offers an explanation: the variations of photoperiod can have opposite effects, depending on individuals, on territorial and hierarchical tendencies which constitute biological roots of mood disorders. PMID- 11105604 TI - [The Hemovigilance Commission of the University Hospital Center]. AB - As suggested by the National Blood Council, a Hemovigilance Committee was set up in the University Hospital of Liege in 1995. A multidisciplinary discussion takes place on any action aiming at the improvement of transfusion safety, and the follow-up of its implementation. The first issue to be discussed was the set up of a detailed documentation of all blood transfusions. The data are now recorded on a single document allowing proper identification of people and products involved, and of the eventual incidents. This document has lead to a better transfusion safety and to an improved administrative management of blood transfusion. The Commission has been coordinating two multi-centric studies analyzing the consumption of fresh blood products and the incidence of transfusion reactions. Among blood-saving policies, autologous transfusion and volume reduction of samples drawn for laboratory purposes have been discussed. Other measures were taken to improve the labeling of samples for cross-mach and to actively follow-up transfusion reactions. By its actions and advises, the Commission aims to direct strategies towards a safe and rational use of blood products. PMID- 11105605 TI - [How I study anomalies of glucose metabolism, insulin secretion and the insulin sensitivity with an oral glucose tolerance test]. AB - Oral glucose tolerance test (OGTT) has been widely used for the diagnosis of diabetes mellitus, gestational diabetes, impaired glucose tolerance or reactive hypoglycaemia. Simultaneous measurements of plasma glucose and insulin levels also allow to derive indices of insulin secretion and insulin sensitivity. Whereas OGTT is not considered anymore as the first choice for the diagnosis of diabetes mellitus, it remains useful for studying abnormalities of glucose metabolism as well as of insulin secretion and insulin action. PMID- 11105606 TI - [Estrogen therapy after menopause and cardiovascular protection: discordance between epidemiologic observations and the results of therapeutic trials] . AB - Based upon the positive results of a large epidemiological study and the negative results of an interventional clinical trial recently published in the New England Journal of Medicine, we will briefly discuss the impact of estrogen replacement therapy on the protection against coronary heart disease in postmenopausal women. PMID- 11105607 TI - [Filling repair and repair fillings]. AB - The possibility of repair of restorations have been regarded for long time as "patchwork dentistry" in standard textbook literature and university education. The current knowledge of the biohazards of dental restorations do not allow an unreflected attitude towards repair techniques. Since there is a lack of investigations on the indication and longevity of repair restorations this technique remains empiric in character. The survey discusses the indication of repair and re-restoration. PMID- 11105608 TI - [Trigeminal neuralgia. Its implications in dentistry. A clinical and therapeutic review]. PMID- 11105609 TI - [Effect of controlled reconditioning training on performance and mood in patients after aortocoronary bypass operation]. AB - The cardiovascular rehabilitation after coronary artery bypass surgery (CABS) is carried out in different intensities and intervals after the intervention in an inpatient or outpatient manner. Studies, which prospectively evaluate the influence of such programs concerning the physical and the psychological status are missing. The following questions were therefore pursued in the present prospective study (66 patients with recently performed myocardial revascularisation): 1. Is it possible by a controlled, regular physical training to normalize the functional capacity in patients after CABS in comparison with a healthy, not specially trained control collective? 2. Do psychological factors and/or age, sex and postoperative anaemia influence the recovery of the reduced functional capacity after the intervention? To answer these questions 66 patients were enrolled 16 +/- 1 days after CABS in a controlled, inpatient average three week-rehabilitation program. During this program the functional capacity of the patients doubled, however without reaching the values of the normal collective. Furthermore, physical and psychological wellbeing and disability as well as the initially mentioned feelings of despondency normalized in men and women. The improvement of fitness during the rehabilitation program was mainly dependent on the improvement of the social integration in women, whereas it correlated with the increase of the physical and psychological well-being in men. Moreover, only in the male patient group the correction of anaemia was an important factor. Age, sex and the preoperative left ventricular function didn't have any effect on the rapidity of recovery after the operation. In summary patients (aged between 30 and 75) of both sexes can already be enrolled in a regular training program two weeks after CABS, leading to a good recovery of their physical and psychological capacity. PMID- 11105610 TI - [Smoking in psychiatry, a neglected problem]. AB - The prevalence of smoking and the associated health risk are especially high in psychiatric patients. Smoking prevention has, however, been neglected until now in psychiatry. Nicotine withdrawal may be associated with some difficulties in psychiatric patients. Existing programs for the prevention and treatment of tobacco dependence do not take into account these special problems. PMID- 11105611 TI - [Evaluation of an internet site on evidence-based medicine]. AB - The present study evaluated a Swiss internet provider of Evidence-based Medicine (EBM) with regard to its utilization and function for medical practitioners. The internet provider under study (www.evimed.ch) primarily provides abstracts of original articles relevant to medical practice that are presented according to the criteria of EBM and includes information about EBM itself. In March 1999 a survey was conducted to better appraise the benefits gained from the information provided from the website visitors' point of view. Around 400 persons who had entered their names in the homepage guest book were informed about the survey by e-mail. A total of 167 questionnaires were filled in online, which is equivalent to the reply rate of close to 42%. The majority of the replies (63.5%) were from private-practice physicians, 22.2% from hospital-based physicians. The average age ranged between 40 and 49 years. 67.7% of the 167 respondents had internet access at their workplace, 72.5% had private internet access. For their own practical work, 61.1% of the respondents rated the information provided by www.evimed.ch as generally useful. The clinical relevance of the studies presented in the Journal Club was rated as good by 55.7% and as very good by 26.9%. The reliability of the information provided was rated as good by 56.3% and as very good by 35.3%. The majority regarded the following homepage sites as personally important: Journal Club (55.7%), articles about EBM (46.1%), MEDLINE access (37.7%) and article citations/links (41.3%). The homepage was visited at an average frequency of 1-3 times a month. 50.3% preferred electronic media (40.1% using various internet providers, 10.2% www.evimed.ch) and 44.3% preferred print media to search for specialist information on a specific medical subject. With regard to new medical findings, 44.9% of the respondents stated that they used print media, 17.4% the www.evimed.ch homepage and 28.7% other internet sources as their primary information medium. Based on this survey, the majority of the respondents gave a positive rating of the www.evimed.ch homepage. Information about EBM and critically appraised studies were evaluated as particularly useful. PMID- 11105612 TI - [Eating at night--eating during sleep]. AB - This 45-years old female, who was referred to us for controlled weight-reduction reports an unusual eating behaviour: No food intake during daytime, uncontrolled eating in the evening and repetitive eating during the night. Eating in the night is a heterogeneous symptom regarding pathogenesis and phenotypic expression. Possible entities are the so called "night-eating syndrome" or "sleep-related eating disorders" with attenuated awareness. Referring to a case report the differential diagnosis of nightly eating disorders will be given and the symptoms and signs of selected pathologies will be discussed shortly. PMID- 11105613 TI - [Pancytopenia]. PMID- 11105614 TI - Polymorphisms of surfactant protein A genes and the risk of bronchopulmonary dysplasia in preterm infants. AB - The pathophysiology of bronchopulmonary dysplasia (BPD) as an inflammatory disorder secondary to neonatal respiratory distress syndrome (RDS) is not yet fully understood and still represents a major complication of prematurity. The main pathophysiologic feature of RDS is a primary surfactant deficiency in a structurally immature lung. Pulmonary surfactant contains 90 percent phospholipids and 10 percent proteins (surfactant proteins A, B, C, and D). As surfactant protein A (SP-A) has several major immunological and metabolic intrapulmonary functions, we aimed at investigating an association of polymorphisms of SP-A1 and SP-A2 encoding genes and the risk of BPD. We performed a case-control study exclusively including Caucasian preterm infants below 32 weeks of gestation matched for the degree of immaturity and the year of birth. Venous cord blood was taken prospectively and analyzed by polymerase chain reaction (PCR), single-strand conformation polymorphism (SSCP), cloning and sequencing. BPD was defined as oxygen dependency or need for mechanical ventilation at day 28. Twenty-three infants with BPD were enrolled (mean gestational age 26.2 weeks; mean birth weight 760.4 g) and compared with 23 infants matched on the basis of gestational age (mean gestational age 27.9 weeks; mean birthweight 1015 g). We observed a significantly increased frequency of the SP-A1 polymorphism 6A6 in infants with BPD compared with controls. In addition to previously established risk factors for BPD, 6A6 polymorphism for SP-A1 gene is an independent co-factor. We believe treatment of neonatal RDS should also include stratification according to genetic risk factors. PMID- 11105615 TI - Dual-probe pH monitoring for the assessment of gastroesophageal reflux in the course of chronic hoarseness in children. AB - The purpose of our study was to assess gastroesophageal reflux (GER) by dual probe pH monitoring in children suffering from chronic hoarseness for more than six months. Seventeen children (aged between 2 and 12 years, 10 boys and 7 girls) were enrolled. All children underwent a laryngoscopy and a 24-hour dual-probe pH monitoring. At both sensor, distal and proximal esophageal, a pathological GER was defined as the presence of episodes of acid reflux with pH < 4 during a fraction of the total recording time greater than 5.2 percent. The computer considered the child was supine when asleep and upright when awake. Laryngoscopy revealed interarytenoid erythema and/or edema with vocal cord nodules or granulomas in 13 cases (76.4%), isolated vocal nodules or granulomas in three cases (17.6%) and a normal appearance in one case (5.8%). At both sensors, the majority of refluxes occurred when the child was upright, as analyzed by the percentage of time the intra-esophageal pH was below four (% time pH < 4), number of refluxes, reflux episodes/hour and longest reflux episode, p < 0.05 between upright and supine for each parameter. The median total % time pH < 4 on the proximal and distal probes was respectively 1.62 percent (95% CI 1.50-3.79) and 11.49 percent (95% CI 8.81-27.17), p < 0.0003. Among the 17 hoarse children, a pathological GER was observed in 12 (70.5%) at the distal sensor and in three (17.5%) at both sensors. Among the 16 hoarse children with abnormal findings on laryngoscopy, two (12.5%) had diagnosed pathological GER at the proximal and 11 (68.7%) at the distal sensor. The only child with normal findings on laryngoscopy exhibited a pathological GER at both sensors. Our results suggest that chronic hoarseness is associated with a pathological GER. The majority of these documented refluxes occurred when the child was awake. PMID- 11105616 TI - Allogeneic bone marrow transplantation for children with myelodysplastic syndrome. AB - Six children with myelodysplastic syndrome underwent allogeneic bone marrow transplantation (BMT) from their HLA-identical siblings. Ages ranged from six to 16 years. French-American British (FAB) diagnosis was refractory anemia with excess blasts (RAEB) in three, RAEB in transformation (RAEB-t) in one and chronic myelomonocytic leukemia (CMML) in two cases. Two patients had progressed to leukemia before BMT. All patients received busulfan and cyclophosphamide as a conditioning regimen. Antithymocyte globulin (ATG) was administered to two of them due to the multiple transfusion history. Graft versus host disease (GvHD) prophylaxis consisted of cyclosporine-methotrexate. Engraftment was documented in all patients except one who underwent a second infusion of bone marrow cells. She died in the early post-transplant period with pancytopenia and veno-occlusive disease of the liver. Two patients died from disease recurrence. Three patients are alive > 12 months post-transplant, two are in remission and one just relapsed at +16 months and is now being prepared for a second bone marrow transplant. The only significant factor for favorable outcome was short duration between diagnosis to transplant in the two patients in remission. PMID- 11105617 TI - Fludarabine, cytarabine, G-CSF and idarubicin (FLAG-IDA) for the treatment of relapsed or poor risk childhood acute leukemia. AB - The prognosis of relapsed acute leukemia or chronic leukemia in acute blast crisis is poor and new chemotherapeutic regimens could be useful for these patients. Six relapsed acute lymphoblastic leukemia (ALL), nine relapsed acute myeloblastic leukemia (AML), one chronic myelomonocytic leukemia (CMML) and one chronic myeloid leukemia (CML) in acute blast crisis between three to 18 years (median 10 years) received fludarabine, cytarabine, G-CSF and idarubicin (FLAG IDA) chemotherapy (CT). Five of the AML relapses were after bone marrow transplantation (BMT) and four were recurrent relapses. At the end of the second course only three patients (2 AML, 1 ALL) were in complete remission (CR). Of the three patients in CR, one patient with AML had her first donor lymphocyte transfusion (DLT) on the 7th day of the second FLAG-IDA course and she is disease free on the 30th month of the second remission. The remaining two patients were transplanted from unrelated donors in a BMT center abroad on the 5th and 8th month of the last remission and both died with BMT-related complications. Out of 25 courses, seven resulted in fatal infections. The regimen was ineffective in B cell ALL as in acute blastic crisis of CMML and CML. We could not evaluate the remission-inducing effect accurately in most of the patients due to induction failure. FLAG-IDA appears to be a myelotoxic therapy for relapsed or poor risk leukemia in a developing country. It is not cost-effective; dose modifications or a regimen without IDA may be tried if there is an available marrow donor. PMID- 11105618 TI - Hemophilic arthropathy: evaluation of clinical and radiological characteristics and disability. AB - Hemophilia is an inherited bleeding disorder which produces its greatest morbidity in the musculoskeletal system. Musculoskeletal complications of hemophilia include acute hemarthrosis, chronic arthritis and hemophilic arthropathy. We studied the clinical and radiological characteristics of joint involvement in hemophiliacs. Functional loss was also demonstrated. There were 25 patients with a mean age of 17; the mean coagulant factor level was 4.2 percent. All the patients had hemarthrotic attacks. Knees were the most commonly affected joints followed by elbows, ankles and hips. The mean number of involved joints was 3.3. Even the patients with moderate disease had arthropathy. Sixty-four percent of the patients had pain and motion restrictions of the involved joints. Four patients developed intramuscular hematoma. We had patients at several radiological stages of severity. Radiological scores best correlated with age and the total number of involved joints. Various degrees of functional loss, namely disability, were observed. The disability score significantly correlated with the radiological score and age. The results of this study suggest that hemophilic arthropathy is an important problem and that multidisciplinary management is needed. Musculoskeletal care as well as appropriate and timely rehabilitation programs could prevent the development of sequelae. PMID- 11105619 TI - Clinical features of 21 patients with lissencephaly type I (agyria-pachygyria). AB - Lissencephaly (agyria-pachygyria) is the most severe neuronal migration disorder, characterized by total or partial absence of gyri. In this study, 21 patients with lissencephaly type I (9 girls, 12 boys) with a mean age of 19 +/- 21 months (2 weeks-8 years) were evaluated clinically and graded according to neuroradiological findings (19 patients by magnetic resonance imaging MRI and 2 by computed tomography CT). Three patients were classified as lissencephaly grade 2 and 18 patients as grade 3 or 4. Clinically, 12 patients (57%) had microcephaly, and eight (38%) had facial dysmorphism. All the patients had prominent psychomotor retardation, moderate to severe; the most frequent neurological findings were spastic guadriplegia (36.4%) and hypotonia with exaggerated tendon reflexes (27.3%). Seventy-eight percent of the patients had epileptic seizures resistant to conventional treatment. Lissencephaly is a cerebral cortical malformation that should be considered in children with developmental delay with or without microcephaly and facial dysmorphism. In addition, it should be investigated in the etiology of early-onset childhood epilepsy. PMID- 11105620 TI - Microdeletion of 22q11 (CATCH 22) in children with conotruncal heart defect and extracardiac malformations. AB - CATCH 22 is a medical acronym for cardiac defects, abnormal facies, thymic hypoplasia, cleft palate, and hypocalcemia, and a variable deletion on chromosome 22q11. The deletion within the chromosome region of 22q11 may occur in patients with dysmorphologic and cardiological syndromes: DiGeorge syndrome (DGS), velocardiofacial syndrome (VCFS), and conotruncal anomaly face syndrome (CAFS). In this study, using N25 (D22S75) DiGeorge chromosome region probe. fluorescence in situ hybridization (FISH) analyses were performed on 32 patients with congenital heart diseases. Twenty-nine of 32 patients had conotruncal heart disease. A 22q11 deletion was detected in two patients (6.9%) of the 29 patients with conotruncal heart disease. One of our 22qdel (+) patients had unilateral facial nerve palsy. Although it is not a frequent finding, unilateral facial nerve palsy will be included among the symptoms of CATCH 22 syndrome. After careful clinical evaluation of patients with conotruncal cardiac anomalies, only syndromic cases should be screened for this deletion. PMID- 11105621 TI - Serratia marcescens: an emerging microorganism in the neonatal intensive care unit. AB - As smaller babies survive in neonatal intensive care units, late-onset septicemia with unusual pathogens appears. Between 1 January and 31 December 1998, in Hacettepe University Ihsan Dogramaci Children's Hospital Neonatal Intensive Care Unit, seven infants had S. marcescens isolates. Four babies had septicemia with the microorganism. The case fatality rate was 50 percent in infants with S. marcescens septicemia. The combination of ceftazidime or imipenem with amikacin appears appropriate for the treatment of newborns with Serratia infection. PMID- 11105622 TI - Ventricular diastolic filling indices in pulmonary stenosis. AB - Children with valvar pulmonary stenosis have right ventricular diastolic filling abnormalities that may be due to either right ventricular hypertrophy or right ventricular outflow obstruction. In order to investigate the reason for this abnormality, 23 consecutive cases with pulmonary stenosis (mean age 7.94 +/- 3.33 years) undergoing transluminal pulmonary balloon valvuloplasty without significant tricuspid or pulmonary valvar regurgitation were studied prospectively. Right ventricular diastolic filling indices and pulmonary valvar systolic gradients were measured in these children one day before and after pulmonary balloon valvuloplasty and were re-examined six months later. Right ventricular diastolic indices based on rapid early diastolic filling peak velocity (peak E), peak velocity during atrial contraction (peak A), and ratio of E/A were determined by pulsed Doppler echocardiography. In conclusion, right ventricular diastolic filling indices in patients with pulmonary stenosis did not improve after pulmonary balloon valvuloplasty in the first day but when re examined by the sixth month there was a significant improvement. These data suggest that diastolic filling abnormalities are more likely a result of right ventricular hypertrophy than of right ventricular outflow obstruction. PMID- 11105623 TI - Unguarded tricuspid orifice diagnosed by echocardiography: a clinical study. AB - Complete absence of tricuspid valve tissue and apparatus with a normal orifice between the right atrium and the right ventricle is defined as "unguarded tricuspid orifice". Very few case report of this anomaly have appeared in the literature. In this article, we present five cases of unguarded tricuspid orifice, isolated or in combination with other anomalies. All patients were males, aged six days to five years; only one case is alive at present. In our opinion, this anomaly is not so infrequent as it is believed to be, and the diagnosis can be made easily with echocardiography if it is kept in mind. PMID- 11105624 TI - Hereditary angioedema: case report of a family. AB - Hereditary angioedema (HAE) is a rare disease resulting from deficiency of complement 1 esterase inhibitor (C1-INH). The clinical manifestations of this disease include recurrent attacks of self-limiting edema affecting face, extremities, gastrointestinal system and upper airways. In this report, we present eleven members of a family with HAE. Edema of the extremities was the most common symptom, occurring in ten patients. Three patients experienced severe laryngeal edema that required tracheotomy. Three patients developed facial and scrotal edema. Three patients experienced severe abdominal pain. The mean age at onset of symptoms was 11 years. C1-INH levels were undetectable in two patients and low in nine patients. CH50 was undetectable in all of the patients. C4 level for all patients was low. HAE in our first case, a 10-year-old boy, was diagnosed on the basis of low C1-INH, CH50 and C4, in addition to his familial history. Eleven members of this family, for whom laboratory studies could not be done, had similar symptoms and course. Two patients died as a result of laryngeal edema before establishment of diagnosis. This case report indicates the importance of recognition and early treatment of HAE to prevent a potentially fatal outcome. PMID- 11105625 TI - Ivemark syndrome: asplenia with kidney collecting duct cysts and polysplenia with cerebellar cyst. AB - Two newborns, one male and one female, from two different families, with Ivemark syndrome proven at autopsy are reported. One of them had asplenia and another had polysplenia. Both newborns had complex cardiac defects with isomerism of the lungs. The newborn with asplenia had dextrocardia, transposition of the great vessels, stenosis of the pulmonary artery, common atrioventricular canal and patent ductus arteriosus. The newborn with polysplenia had a common atrium, hypoplastic left ventricle and patent ductus arteriosus. The patient with asplenia had cystic dilated collecting ducts of the kidney and the patient with polysplenia had cerebellar cyst. These associate malformations have not been reported previously. Both cases were sporadic. PMID- 11105626 TI - A case of isotretinoin embryopathy with bilateral anotia and Taussig-Bing malformation. AB - We report a newborn infant with multiple congenital anomalies (anotia and Taussig Bing malformation) due to exposure to isotretinoin within the first trimester. In this paper we aim to draw to the fact that caution is needed when prescribing vitamin A-containing drugs to women of childbearing years. PMID- 11105627 TI - Teratoid Wilms' tumor: a case report. AB - Teratoid Wilms' tumor is rarely seen and is a description used only recently. The term describes classical nephroma with a diversity of cell types and tissues. In this reported case, the epithelial component consisting of squamous areas made up 70 percent of the tumor; no criteria of dysplasia nor any nephroblastomatosis areas or endodermal elements were presented. Although it is reported that teratoid Wilms' tumor is not usually aggressive or metastatic, a case of unilateral teratoid Wilms' tumor in a 2.5-year-old-boy who died because of metastatic disease is presented and the literature reviewed. PMID- 11105628 TI - Progressive encephalopathy with edema, hypsarrhythmia, and optic atrophy (PEHO syndrome) in a Turkish child. AB - We report a Turkish boy with PEHO syndrome (progressive encephalopathy with edema, hypsarrhythmia, and optic atrophy). He had generalized hypotonia and abnormal eye movements during early infancy. Infantile spasms were seen in the second year of life. Arrest of psychomotor development and blindness were noticed early in childhood. Serial magnetic resonance imaging revealed progressive infratentorial atrophy with association of cortical atrophy and corpus callosum hypoplasia. This is an additional case of PEHO syndrome, to our knowledge the first such case from Turkey. PMID- 11105629 TI - Body stalk anomaly in monozygotic twinning: a case report. AB - We describe a case of concordant body stalk anomaly in a monozygotic twin. Autopsy of the fetus showed abnormalities compatible with the maldevelopment of embryonic folding. Abdominal viscera were in a sac covered by the amnion and were attached directly to the placenta. The anus was not visible and no discernible external genitalia were noted. Other findings included a neural tube defect and a rectal duplication as an enteric cyst. Umbilical cord had only one vein and an artery. No abnormalities were found on pathologic examination of the placenta. Although we encountered cases previously with gastroschsis and omphalocele, this was the first case of body stalk anomaly that we recognized as an enteric cyst, which is extremely rare in twins. PMID- 11105630 TI - Thoracic ectopic kidney in a child: a case report. AB - Congenital thoracic ectopic kidney is a very rare developmental anomaly and the rarest form of all ectopic kidneys. It is usually asymptomatic and discovered incidentally on a routine chest radiography. We report a thoracic ectopic kidney in a 19-month-old boy, which initially presented as a well demarcated mass at the base of the right lung on chest x-ray. Intravenous pyelography (IVP) and thoraco abdominal computed tomography (CT) demonstrated a normal functioning transdiaphragmatic thoracic ectopic right kidney, but technetium-99m DTPA and DMSA scintigraphy demonstrated pelvic stasis. We hereby discuss the features of congenital thoracic ectopic kidney and review the literature. Although it is extremely rare, thoracic ectopic kidney should be considered in differential diagnosis of a mass with a well demarcated superior margin in the lower part of the thorax, and renal scintigraphy must be performed even if CT and IVP results are normal. PMID- 11105631 TI - Acute lymphoblastic leukemia in a child with Wilson disease. AB - Wilson disease is an autosomal recessively inherited disease of copper metabolism and is characterized by liver and central nervous system dysfunction. The heterozygote carrier state rate is about one in 90 persons and the incidence of the disease is about 30 in 1,000,000. Although leukemia is the most common form of childhood malignancies, the probability of the presence of Wilson disease and acute lymphoblastic leukemia in the same patient is very low. We report an unusual case of a child with Wilson disease who developed acute lymphoblastic leukemia in three months. PMID- 11105632 TI - Biphasic pulmonary blastoma in a child. AB - Pulmonary blastoma (PB) is a rare malignant pulmonary tumor composed of immature mesenchyme and/or epithelium that resembles an embryonic lung at 10-16 weeks gestation. PBs constitute only 0.25 to 0.5 percent of all primary malignant lung tumors. Approximately 20 percent of the reported cases have occurred in pediatric patients. A seven-year-old girl presented with fever, cough, respiratory distress and chest pain on the left side. An x-ray, ultrasonography and a computed tomographic scan of the chest showed a large mass consisting of solid and cystic components almost completely occupying the left hemithorax associated with pleural effusion. The diagnosis of biphasic PB was established by histological examination of thoracotomy material. The patient was considered inoperable due to tumor involvement of the mediastinum, and she died two days after the initiation of chemotherapy. We report this case of PB to raise attention to the clinical, radiological and pathological features of PB in childhood because of its rarity. PMID- 11105633 TI - Juvenile hyaline fibromatosis in one Turkish child. AB - We describe a case of juvenile hyaline fibromatosis (JHF) in a Turkish child. Only about 40 cases of juvenile hyaline fibromatosis had been reported in English literature as of March 1998, and it had not been reported in English literature from Turkey as of November 1998. Juvenile hyaline fibromatosis characterized by multiple cutaneous masses is a rare hereditary disorder. This disease is usually found in children, and a malfunction of collagen synthesis is considered as the pathogenetic cause. In the presented case, light microscopy demonstrated an abundance of a homogeneous, amorphous, eosinophilic extracellular matrix in which fibroblasts were embedded. Well-formed collagen fibers could not be demonstrated with Gieson's method or with reticulin preparation. The hayalin material periodic acid-Schiff-positive and diastase-resistant, whereas the Congo red method was negative. Immunohistochemically, the spindle-shaped cells were actin (smooth muscle) negative. PMID- 11105634 TI - Leukocytoclastic vasculitis associated with methotrexate therapy. PMID- 11105635 TI - Concurrent presence of HBsAg and anti-HBs in children. PMID- 11105636 TI - The prefrontal cortex and the integration of sensory, limbic and autonomic information. PMID- 11105637 TI - Neurobiological mechanisms of emotionally influenced, long-term memory. PMID- 11105638 TI - Hypothalamic connections with the cerebral cortex. PMID- 11105640 TI - Functional anatomy of arousal and attention systems in the human brain. PMID- 11105639 TI - Functional neuroanatomy of the prefrontal cortex: autonomic interactions. PMID- 11105641 TI - Attentional processes and learning and memory in rats: the prefrontal cortex and hippocampus compared. PMID- 11105642 TI - Limitations in information processing in the human brain: neuroimaging of dual task performance and working memory tasks. PMID- 11105643 TI - Role of the prefrontal cortex of the rat in learning and decision making: effects of transient inactivation. PMID- 11105644 TI - The integration of stress by the hypothalamus, amygdala and prefrontal cortex: balance between the autonomic nervous system and the neuroendocrine system. PMID- 11105645 TI - Dopamine and noradrenaline release in the prefrontal cortex in relation to unconditioned and conditioned stress and reward. PMID- 11105646 TI - Locus coeruleus and regulation of behavioral flexibility and attention. PMID- 11105647 TI - Stress impairs prefrontal cortical function in rats and monkeys: role of dopamine D1 and norepinephrine alpha-1 receptor mechanisms. PMID- 11105648 TI - Involvement of basal ganglia and orbitofrontal cortex in goal-directed behavior. AB - An impressive array of neural processing appears to be dedicated to the extraction of reward-related information from environmental stimuli and use of this information in the generation of goal-directed behaviors. While other structures are certainly involved in these processes, the characteristics of activations seen in mesencephalic dopamine neurons, striatal neurons and neurons of the orbitofrontal cortex provide distinct examples of the different ways in which reward-related information is processed. In addition, the differences in activations seen in these three regions demonstrate the different roles they may play in goal-directed behavior. A principal role played by dopamine neurons is that of a detector of an error in reward prediction. The homogeneity of responsiveness across the population of dopamine neurons indicates that this error signal is widely broadcast to dopamine terminal regions where it could provide a teaching signal for synaptic modifications underlying the learning of goal-directed appetitive behaviors. The responses of these same neurons to conditioned stimuli associated with reward could also serve as a signal of prediction error useful for the learning of sequences of environmental stimuli leading to reward. Dopamine neuron responses to both rewards and conditioned stimuli are not contingent on the behavior executed to obtain the reward and thus appear to reflect a relatively pure signal of a reward prediction error. It is not yet clear whether these activations, and responses to novel stimuli, have an additional function in engaging neural systems involved in the representation and execution of goal-directed behaviors. This representation of goal-directed behaviors may involve the striatal regions studied, where processing of reward related information appears to be much more heterogeneous. Different subpopulations of striatal neurons are activated at different stages in the course of goal-directed behaviors, with largely separate populations activated following presentation of conditioned stimuli, preceding reinforcers, and following reinforcers. Neurons exhibiting each of these types of activation appear to differentiate between rewarding and non-rewarding outcomes of behavioral acts and, as a population, appear to be biased towards processing reward vs. non-reward. These activations observed in the striatum were often contingent on the behavioral act associated with obtaining reward, reflecting an integration of information not observed in dopamine neurons. Another difference between reward processing in striatal neurons and dopamine neurons is the influence of predictability on neuronal responsiveness. Unlike dopamine neurons, many striatal neurons respond to predicted rewards, although at least some may reflect the relative degree of predictability in the magnitude of the responses to reward. Thus, striatal processing of reward-related information is in some ways more complex than that observed in dopamine neurons, incorporating information on behavior and potentially providing more detailed information regarding predictability. These activations could serve as a component of the neural representation of the goal, and/or the behavioral aspects of goal-directed behaviors. As such they would be of use for the execution of appropriate goal directed behaviors in response to known environmental stimuli, as well as for generating behaviors in response to novel stimuli that may be associated with desirable goals. Neuronal activations in the orbitofrontal cortex appear to involve less integration of behavioral and reward-related information, but rather incorporate another aspect of reward, the relative motivational significance of different rewards. These activations would serve a function similar to those striatal neurons that encode exclusively reward-related information in situations in which only a single outcome is obtainable. (ABSTRACT TRUNCATED) PMID- 11105649 TI - Reward-dependent learning in neuronal networks for planning and decision making. AB - Neuronal network models have been proposed for the organization of evaluation and decision processes in prefrontal circuitry and their putative neuronal and molecular bases. The models all include an implementation and simulation of an elementary reward mechanism. Their central hypothesis is that tentative rules of behavior, which are coded by clusters of active neurons in prefrontal cortex, are selected or rejected based on an evaluation by this reward signal, which may be conveyed, for instance, by the mesencephalic dopaminergic neurons with which the prefrontal cortex is densely interconnected. At the molecular level, the reward signal is postulated to be a neurotransmitter such as dopamine, which exerts a global modulatory action on prefrontal synaptic efficacies, either via volume transmission or via targeted synaptic triads. Negative reinforcement has the effect of destabilizing the currently active rule-coding clusters; subsequently, spontaneous activity varies again from one cluster to another, giving the organism the chance to discover and learn a new rule. Thus, reward signals function as effective selection signals that either maintain or suppress currently active prefrontal representations as a function of their current adequacy. Simulations of this variation-selection have successfully accounted for the main features of several major tasks that depend on prefrontal cortex integrity, such as the delayed-response test, the Wisconsin card sorting test, the Tower of London test and the Stroop test. For the more complex tasks, we have found it necessary to supplement the external reward input with a second mechanism that supplies an internal reward; it consists of an auto-evaluation loop which short-circuits the reward input from the exterior. This allows for an internal evaluation of covert motor intentions without actualizing them as behaviors, by simply testing them covertly by comparison with memorized former experiences. This element of architecture gives access to enhanced rates of learning via an elementary process of internal or covert mental simulation. We have recently applied these ideas to a new model, developed with M. Kerszberg, which hypothesizes that prefrontal cortex and its reward-related connections contribute crucially to conscious effortful tasks. This model distinguishes two main computational spaces within the human brain: a unique global workspace composed of distributed and heavily interconnected neurons with long-range axons, and a set of specialized and modular perceptual, motor, memory, evaluative and attentional processors. We postulate that workspace neurons are mobilized in effortful tasks for which the specialized processors do not suffice; they selectively mobilize or suppress, through descending connections, the contribution of specific processor neurons. In the course of task performance, workspace neurons become spontaneously co-activated, forming discrete though variable spatio-temporal patterns subject to modulation by vigilance signals and to selection by reward signals. A computer simulation of the Stroop task shows workspace activation to increase during acquisition of a novel task, effortful execution, and after errors. This model makes predictions concerning the spatio temporal activation patterns during brain imaging of cognitive tasks, particularly concerning the conditions of activation of dorsolateral prefrontal cortex and anterior cingulate, their relation to reward mechanisms, and their specific reaction during error processing. PMID- 11105650 TI - The glutamate hypothesis of reinforcement learning. PMID- 11105651 TI - Interactions between medial prefrontal cortex and meso-limbic components of brain reward circuitry. PMID- 11105652 TI - Limbic cortical-ventral striatal systems underlying appetitive conditioning. PMID- 11105653 TI - Plasticity of neuronal firing in deep layers of the medial prefrontal cortex in rats engaged in operant conditioning. PMID- 11105654 TI - Activity patterns in mesolimbic regions in rats during operant tasks for reward. PMID- 11105656 TI - Evidence of fronto-thalamic involvement in schizophrenia. PMID- 11105655 TI - Reward deficiency syndrome: genetic aspects of behavioral disorders. AB - The dopaminergic and opioidergic reward pathways of the brain are critical for survival since they provide the pleasure drives for eating, love and reproduction; these are called 'natural rewards' and involve the release of dopamine in the nucleus accumbens and frontal lobes. However, the same release of dopamine and production of sensations of pleasure can be produced by 'unnatural rewards' such as alcohol, cocaine, methamphetamine, heroin, nicotine, marijuana, and other drugs, and by compulsive activities such as gambling, eating, and sex, and by risk taking behaviors. Since only a minority of individuals become addicted to these compounds or behaviors, it is reasonable to ask what factors distinguish those who do become addicted from those who do not. It has usually been assumed that these behaviors are entirely voluntary and that environmental factors play the major role; however, since all of these behaviors have a significant genetic component, the presence of one or more variant genes presumably act as risk factors for these behaviors. Since the primary neurotransmitter of the reward pathway is dopamine, genes for dopamine synthesis, degradation, receptors, and transporters are reasonable candidates. However, serotonin, norepinephrine, GABA, opioid, and cannabinoid neurons all modify dopamine metabolism and dopamine neurons. We have proposed that defects in various combinations of the genes for these neurotransmitters result in a Reward Deficiency Syndrome (RDS) and that such individuals are at risk for abuse of the unnatural rewards. Because of its importance, the gene for the [figure: see text] dopamine D2 receptor was a major candidate gene. Studies in the past decade have shown that in various subject groups the Taq I A1 allele of the DRD2 gene is associated with alcoholism, drug abuse, smoking, obesity, compulsive gambling, and several personality traits. A range of other dopamine, opioid, cannabinoid, norepinephrine, and related genes have since been added to the list. Like other behavioral disorders, these are polygenically inherited and each gene accounts for only a small per cent of the variance. Techniques such as the Multivariate Analysis of Associations, which simultaneously examine the contribution of multiple genes, hold promise for understanding the genetic make up of polygenic disorders. PMID- 11105657 TI - The importance of a human 3D database and atlas for studies of prefrontal and thalamic functions. PMID- 11105658 TI - Interaction of prefrontal cortical and hypothalamic systems in the pathogenesis of depression. PMID- 11105659 TI - Histopathology of the prefrontal cortex in major depression: what does it tell us about dysfunctional monoaminergic circuits? PMID- 11105660 TI - Functional anatomical abnormalities in limbic and prefrontal cortical structures in major depression. AB - Neuroimaging studies of major depression have identified neurophysiological abnormalities in multiple areas of the prefrontal cortex (PFC), the amygdala, and related parts of the striatum and thalamus. Some of these abnormalities are mood state-dependent, and appear in regions where cerebral blood flow (CBF) increases during other normal and pathological emotional states. These neurophysiological differences between depressives and non-depressed controls may thus locate areas where physiological activity changes to mediate or respond to the emotional, behavioral and cognitive manifestations of major depressive episodes (MDE). Other abnormalities persist following symptom remission, and are found in orbital and medial PFC areas where post mortem studies demonstrate reductions in cortex volume and/or histopathological changes in primary mood disorders. These orbital and medial PFC areas have been shown by other types of evidence to modulate emotional behavior and stress responses, suggesting that dysfunction involving these regions may be involved in the pathogenesis of depressive symptoms. Finally, physiological activity is decreased during MDE in dorsal PFC areas implicated in language, selective attention, visuospatial or mnemonic processing, but these abnormalities reverse with symptom remission. These areas of 'deactivation' during the depressed state may reflect neurophysiological interactions between cognitive and emotional processing, and may relate to the subtle cognitive impairments associated with MDE. PMID- 11105661 TI - Role for dopamine in the behavioral functions of the prefrontal corticostriatal system: implications for mental disorders and psychotropic drug action. AB - We have discussed the role of dopamine in modulating the interactions between cortical and striatal regions that are involved in behavioral regulation. The evidence reviewed seems to suggest that dopamine acts, overall, to promote stimulus-induced responding for conditioned or reward-related stimuli by integrative actions at multiple forebrain sites. It is thus not surprising that dopaminergic dysfunction has been implicated in a number of neuropsychiatric disorders that involve abnormal cognitive and affective function. Future studies aimed at pinpointing the precise anatomical sites of action and molecular mechanisms involved in dopaminergic transmission within the corticolimbic circuit are critical for trying to disentangle the cellular mechanisms by which dopamine exerts its actions. Moreover, the afferent control of dopamine neurons from brainstem and forebrain sites need to be fully explored in order to begin to understand what mechanisms are involved in regulating the dopaminergic response to stimuli with incentive value. Finally, the post-synaptic consequences of prolonged and supranormal dopaminergic activation need to be investigated in order to understand what persistent neuroadaptations result from chronic activation of this neuromodulatory system (e.g. in drug addiction). Answers to these sorts of questions will undoubtedly provide important insights into the nature of dopaminergic function in the animal and human brain. PMID- 11105662 TI - Eighth C.U. Ariens Kappers Lecture. The fabric of the mind: a neurobiological perspective. PMID- 11105663 TI - From arousal to cognition: the integrative position of the prefrontal cortex. PMID- 11105664 TI - Strategies for spinal cord injury repair. PMID- 11105665 TI - Cell death and plasticity after experimental spinal cord injury. PMID- 11105666 TI - The multi-domain structure of extracellular matrix molecules: implications for nervous system regeneration. PMID- 11105667 TI - Spinal cord injury-induced inflammation: a dual-edged sword. PMID- 11105668 TI - Immunological regulation of neuronal degeneration and regeneration in the injured spinal cord. PMID- 11105669 TI - Plasticity of neuronal networks in the spinal cord: modifications in response to altered sensory input. PMID- 11105670 TI - Neural plasticity as revealed by the natural progression of movement expression- both voluntary and involuntary--in humans after spinal cord injury. PMID- 11105671 TI - Laufband (LB) therapy in spinal cord lesioned persons. PMID- 11105672 TI - Spinal and supraspinal plasticity after incomplete spinal cord injury: correlations between functional magnetic resonance imaging and engaged locomotor networks. PMID- 11105673 TI - Impact of neuroprosthetic applications on functional recovery. PMID- 11105674 TI - Nerve cuffs for nerve repair and regeneration. PMID- 11105675 TI - Cortical motor areas and their properties: implications for neuroprosthetics. PMID- 11105676 TI - Network level properties of short-term plasticity in the somatosensory system. PMID- 11105677 TI - The reorganization of somatosensory and motor cortex after peripheral nerve or spinal cord injury in primates. PMID- 11105678 TI - Neurotrophins and activity-dependent plasticity. PMID- 11105679 TI - Differences in the regulation of neuropeptide Y, somatostatin and parvalbumin levels in hippocampal interneurons by neuronal activity and BDNF. PMID- 11105680 TI - Long-term regulation of excitatory and inhibitory synaptic transmission in hippocampal cultures by brain-derived neurotrophic factor. PMID- 11105681 TI - Neurotrophins and activity-dependent inhibitory synaptogenesis. PMID- 11105682 TI - Modulation of hippocampal synaptic transmission and plasticity by neurotrophins. PMID- 11105683 TI - Neurotrophin-evoked rapid excitation of central neurons. PMID- 11105684 TI - Candidate cells for transplantation into the injured CNS. PMID- 11105685 TI - Autoimmune involvement in CNS trauma is beneficial if well controlled. PMID- 11105686 TI - Olfactory ensheathing glia transplantation into the injured spinal cord. PMID- 11105687 TI - Precursor cells for transplantation. PMID- 11105688 TI - Potential use of marrow stromal cells as therapeutic vectors for diseases of the central nervous system. PMID- 11105689 TI - Neurobiology of human neurons (NT2N) grafted into mouse spinal cord: implications for improving therapy of spinal cord injury. AB - Emerging data suggest that current strategies for the treatment of spinal cord injury might be improved or augmented by spinal cord grafts of neural cells, and it is possible that grafted neurons might have therapeutic potential. Thus, here we have summarized recent studies of the neurobiology of clonal human (NT2N) neurons grafted into spinal cord of immunodeficient athymic nude mice. Postmitotic human NT2N neurons derived in vitro from an embryonal carcinoma cell line (NT2) were transplanted into spinal cord of neonatal, adolescent and adult nude mice where they became integrated into the host gray and white matter, did not migrate from the graft site, and survived for > 15 months after implantation. The neuronal phenotype of the grafted NT2N cells was similar in gray and white matter regardless of host age at implantation, and some of the processes extended by the transplanted NT2N neurons became ensheathed by oligodendrocytes. However, there were consistent differences between NT2N processes traversing white versus gray matter. Most notably, NT2N processes with a trajectory in white matter extended over much longer distances (some for > 2 cm) than those confined to gray matter. Thus, NT2N neurons grafted into spinal cord of nude mice integrated into gray as well as white matter, where they exhibited and maintained the morphological and molecular phenotype of mature neurons for > 15 months after implantation. Also, the processes extended by grafted NT2N neurons differentially responded to cues restricted to gray versus white matter. Further insight into the neurobiology of grafted human NT2N neurons in the normal and injured spinal cord of experimental animals may lead to novel and more effective strategies for the treatment of spinal cord injury. PMID- 11105690 TI - Grafting of genetically modified fibroblasts into the injured spinal cord. PMID- 11105691 TI - Delivery of therapeutic molecules into the CNS. PMID- 11105692 TI - Neurotrophin small-molecule mimetics. PMID- 11105693 TI - Tissue engineering strategies for nervous system repair. PMID- 11105694 TI - In vivo neuroprotection of injured CNS neurons by a single injection of a DNA plasmid encoding the Bcl-2 gene. PMID- 11105695 TI - Association of renal agenesis and mullerian duct anomalies. AB - PURPOSE: The purpose of this work was to determine the association of renal agenesis with the different types of mullerian duct anomalies (MDAs). METHOD: A 5 year retrospective review of MR records identified 57 patients with MDAs. Associated renal anomalies were correlated with the various types of MDAs. RESULTS: Renal agenesis was found in 17 (29.8%) of 57 patients. No other renal anomalies were identified. Renal agenesis was more frequent in patients with uterus didelphys (13/16 cases). Renal agenesis was also seen in patients with uterine agenesis (2/5 cases) and unicornuate uterus (2/7 cases). All 11 cases of obstructed uterus didelphys were associated with renal agenesis ipsilateral to the side of the obstructing transverse hemivaginal septum. CONCLUSION: Renal agenesis is more commonly seen in uterus didelphys than in other types of MDAs. Renal agenesis in patients with uterus didelphys is often ipsilateral to an obstructing, transverse, hemivaginal septum. PMID- 11105696 TI - Double-phase helical CT of small renal parenchymal neoplasms: correlation with pathologic findings and tumor angiogenesis. AB - PURPOSE: To correlate the enhancement pattern of double-phase helical computed tomography (CT) of small renal parenchymal neoplasms with pathologic findings and tumor angiogenesis, and evaluate whether the enhancement pattern would be useful in differentiating the histomorphologic types of small renal parenchymal neoplasms. MATERIALS AND METHODS: Double-phase helical CT (5 mm slice) of the corticomedullary phase (CMP) and late nephrographic phase (NP) was performed in 40 surgically resected renal neoplasms <3.5 cm. The patterns of CT attenuation value and homogeneity were correlated with the subtypes of neoplasms, microvessel density, and the existence of intratumoral necrosis or hemorrhage. RESULTS: Clear cell renal cell carcinomas (RCC) (n = 29) showed a peak attenuation value in the CMP of >100 HU [Hounsfield units]. Chromophobe cell RCC (n = 2) showed a peak attenuation value in the CMP of <100 HU. Papillary RCC (n = 5) showed a gradual enhancement with the attenuation value in the CMP of <100 HU. However oncocytomas (n = 2) and metanephric adenomas (n = 2) also showed patterns similar to these subtypes of RCC. The degree of enhancement in the CMP correlated with microvessel density (r = 0.87). All tumors with an homogeneous enhancement pattern did not show necrosis or hemorrhage on histologic specimen. CONCLUSION: The enhancement pattern in double-phase helical CT was different among the subtypes of RCC, and correlated with microvessel density or the existence of intratumoral necrosis or hemorrhage. However it did not differentiate between RCC and other solid tumors. PMID- 11105697 TI - Incidental detection of preclinical renal tumors with electron beam computed tomography: report of 26 consecutive operated patients. AB - PURPOSE: The purpose of this work was to describe the positive predictive value of electron beam CT (EBCT) for diagnosis of solid renal tumors. METHOD: Among 11,932 consecutive patients undergoing screening EBCT, 27 cases met EBCT criteria for solid renal tumors. Twenty-six of 27 patients underwent surgery. RESULTS: Surgical pathology identified 25 solid renal tumors and 1 adrenal hemorrhage with thrombus. Twenty tumors were classified as T1N0M0, one was T2N0M0, and one was T3aN0M0. All tumor patients are clinically well at 1-41 months (mean 17 months) postoperatively. None of the patients had clinical signs or symptoms characteristic of renal malignancy. CONCLUSION: EBCT is an effective tool for detection of solid renal tumors in a healthy outpatient population (positive predictive value 0.96). The detection rate is low [0.2% (26/11,932) at our facility] in patients undergoing EBCT for other indications. The cost effectiveness and sensitivity of this technique for solid renal tumor detection among various populations remain to be determined. PMID- 11105698 TI - CT findings after uterine artery embolization. AB - Asymptomatic uterine leiomyoma can be detected on routine computed tomography (CT) of the pelvis. Leiomyomas have been described as low attenuation masses that can disrupt the smooth contour of a normal uterus. Four women underwent uterine artery embolization for the treatment of uterine leiomyoma. CT findings include initial retention of contrast in fibroids the day of the procedure and central necrosis of the fibroid with subsequent cavitation as early as 1 month postprocedure. PMID- 11105699 TI - Multidetector CT angiography in the evaluation of pancreatic carcinoma: preliminary observations. AB - Multidetector CT (MDCT) provides unparalleled capabilities for combining narrow scan collimation with rapid data acquisition protocols. When combined with CT angiographic techniques and 3D-volume rendering we are able to create unique displays for evaluating a range of clinical pathologies. In this pictorial review we present the potential advantages of using MDCT angiography for the evaluation of pancreatic cancer and its role in the accurate staging of these patients. The use of dual-phase CT scanning in both the arterial phase and portal phase is addressed with the role of 3D CT angiography clearly defined. Numerous case studies are presented to show the advantages of these techniques over simple axial CT imaging. PMID- 11105700 TI - Lipomatous tumors of the stomach: CT findings and differential diagnosis. AB - This article reviews the computed tomography imaging features of a variety of gastric tumors containing fatty tissue. Lipoma, angiolipoma, liposarcoma, and teratoma are described. Differential diagnosis includes primary and reactive lipomatosis, carcinoma engulfing the perivisceral fat thus mimicking differentiated liposarcoma, and mesenchymal gastric and peritoneal neoplasms. PMID- 11105701 TI - Reevaluation of spiral CT cholangiography: basic considerations and reliability for detecting choledocholithiasis in 80 patients. AB - PURPOSE: The purpose of this work was to reevaluate the characteristics and diagnostic accuracy of spiral CT cholangiography (CTC) for detecting biliary calculi. METHOD: Spiral CTC was performed in 133 patients with suspected biliary or pancreatic diseases. All source images were reviewed by two radiologists who were unaware of final diagnoses. Attenuation values of bile were correlated with biochemical data and visualization of anatomic detail. The statistical measures in detecting the presence of choledocholithiasis were calculated in 80 patients with confirmed diagnoses. RESULTS: Statistically significant correlations were found between the degree of biliary enhancement and both serum bilirubin and alkaline phosphatase levels. Of the 80 patients, 18 (23%) had choledocholithiasis and 62 did not. Observers diagnosed them with a sensitivity of 89% and a specificity of 98%. A mild adverse reaction to contrast material was observed in three (2.3%) patients. CONCLUSION: Spiral CTC is a reliable, noninvasive, and accessible technique for detecting choledocholithiasis. PMID- 11105702 TI - CT appearance of hepatic parenchymal changes after percutaneous microwave coagulation therapy for hepatocellular carcinoma. AB - PURPOSE: The purpose of this study was to evaluate the changes in the CT appearance of the hepatic parenchyma surrounding the necrotic area in the early period after percutaneous microwave coagulation therapy (PMCT) for hepatocellular carcinoma (HCC). METHOD: We reviewed enhanced CT scans obtained before and within 2 weeks, at 1 month, and at 3 months after PMCT of 61 lesions in 47 patients with HCC. RESULTS: On dynamic CT, early enhancement of the hepatic parenchyma around the treated area was a frequent finding within 1 (87%) or 2 (68%) weeks after PMCT, but such enhancement disappeared on follow-up. Arterioportal shunts were also demonstrated by enhanced CT after treatment (21% at < or =2 weeks), and these shunts tended to persist for >1 month. CONCLUSION: We should evaluate the effect of PMCT by performing dynamic enhanced CT not only within 2 weeks to determine the end-point of treatment but also at 1 month or more after finishing treatment for definite assessment of tumor necrosis. PMID- 11105703 TI - Dynamic contrast-enhanced subtraction and delayed MRI of gastric tumors: radiologic-pathologic correlation. AB - PURPOSE: Our goal was to determine whether dynamic MR subtraction images could be used to detect and stage gastric tumors. METHOD: Dynamic MR subtraction images were prospectively performed in 20 patients without gastric lesions and in 39 patients with gastric tumors. The flat- or depressed-type early gastric cancers were excluded. The MR findings were assessed for layered pattern of the normal gastric wall, detectability of tumors, enhanced pattern of tumor, and depth of the tumor invasion. Surgical specimens were obtained from 30 of the patients with tumors, and histopathologic sections were made in the dynamic MR scanning direction. RESULTS: The three-layered structure of the normal gastric wall was apparent in more of the dynamic MR subtraction images (60%) than of the nonsubtraction images (30%) in the control group. All 39 gastric tumors were detected by MRI. The intact inner layers overlying stromal tumors and outer layers interrupted by advanced gastric cancers were clear on the subtracted images. MRI accurately T-staged 88% of the gastric cancers. CONCLUSION: Dynamic MR subtraction images can be used to identify gastric tumors and to stage gastric cancers. PMID- 11105704 TI - Value of hepatic arterial phase CT versus lipiodol ultrafluid CT in the detection of hepatocellular carcinoma. AB - OBJECTIVE: To evaluate the role of hepatic arterial phase (HAP) spiral computed tomography (CT), as compared with iodized oil (Lipiodol ultrafluid [LUF]) CT for revealing nodular hepatocellular carcinomas (HCC). METHODS: Twenty-four cirrhotic patients underwent two-phase HCT examination: HAP 25 seconds and portal phase 70 seconds after injection of 1.5 mL/Kg contrast medium. All patients also underwent hepatic angiography and intraarterial infusion of iodized oil; LUF CT was performed 3-4 weeks after infusion. HCT images were compared with LUF CT images for detection of hepatic nodules. RESULTS: We found no significant difference between the sensitivity of HAP CT and LUF CT for nodules >10 mm, while HAP CT was more sensitive than LUF CT in revealing nodules <10 mm (47 vs. 27, p < 0.001). CONCLUSIONS: HCT should be considered as the first method for the detection of HCC, whereas LUF CT should be used only for therapy. PMID- 11105705 TI - Amyand's hernia: prospective CT diagnosis. AB - We report two cases of Amyand's hernia, which is the development of acute appendicitis within an inguinal hernia. Both patients were clinically thought to have incarcerated inguinal hernias, but were correctly prospectively diagnosed as having Amyand's hernia on the basis of preoperative computed tomography (CT) examinations. Our cases again show the utility of CT of the acute abdomen and pelvis in revealing a previously unsuspected diagnosis and rapidly triaging patients to the appropriate management. PMID- 11105706 TI - Radiographic findings of ischemic hepatitis in a cirrhotic patient. AB - We report the radiographic findings of ischemic hepatitis in a patient with cirrhosis. The abdominal ultrasound exam showed multiple hypoechoic nodules in the liver measuring up to 2 cm, suggestive of diffuse metastatic disease. Abdominal computed tomography (CT) scan revealed multiple hypodense masses throughout the liver with no enhancement. Liver biopsy revealed coagulative hepatocyte necrosis at the center of the regenerative nodules. Repeat CT scan obtained 5 months later showed complete resolution of the hypodense nodules. Ischemic necrosis of regenerative nodules should be differentiated from diffuse hepatic metastatic disease in the setting of ischemic hepatitis in cirrhotic patients. PMID- 11105707 TI - MR arteriography using an implantable port system: a new method in assessing perfusion abnormalities during hepatic arterial infusion chemotherapy. AB - We present a case in which MR arteriography (MRA) with an indwelling catheter was used in a perfusion study of intrahepatic arterial chemotherapy for liver metastases. After embolization of collateral vessels using platinum coils, CT imaging was disturbed by strong artifact. However, platinum coils produced no MR artifact. In addition, MRA had greater advantages in depicting perfusion defects than perfusion scintigraphy. We consider MRA useful in assessing perfusion abnormalities during intrahepatic arterial chemotherapy. PMID- 11105708 TI - Fat replacement with absence of acinar and ductal structure in the pancreatic body and tail. AB - We report an unusual case of fat replacement of the pancreatic body and tail. Findings on contrast-enhanced computed tomography and ERCP could be confused with dorsal pancreas agenesis. Histopathologic examination of the resected specimen revealed massive fat replacement with complete absence of the acinar and ductal tissue and scattered islets of Langerhans. PMID- 11105709 TI - The use of clinical CT for baseline bone density assessment. AB - PURPOSE: Many patients having an abnormal initial bone densitometry study have had a previous abdominal/pelvic computed tomography (CT) for other clinical reasons. This study evaluates if a nondedicated quantitative CT (QCT) abdominal/pelvic CT scan could be used as reliable baseline data for subsequent dedicated bone density studies. SUBJECTS AND METHODS: Twenty-six patients (13 men, 13 women) undergoing clinically-indicated non-i.v. and i.v. contrast abdominal/pelvic CT had dedicated QCT performed immediately following scans of the L1, L2, and L3 vertebral bodies. QCT was then performed on all three scans. A repeated measures analysis of variance model was used to analyze the data in order to compare noncontrast clinical CT with QCT and noncontrast clinical with contrast clinical CT. RESULTS: The mean bone mineral density for the noncontrast clinical study was 98.51 (mg/cc) versus 90.56 (mg/cc) for QCT (p = 0.0003; 95% confidence interval: 3.90 to 13.71). There was no significant difference (p = 0.085) between QCT performed from non-i.v. and i.v. contrast clinical CT scans. CONCLUSION: Bone densitometry can be performed from either non-i.v. or i.v. contrast clinical CT scans if a conversion factor is applied. This can be determined by utilizing a formula Daverage = -7.83 + (0.99 x NCaverage), where Daverage and NCaverage are the abbreviations of "dedicated" and "noncontrast clinical" BMD averaged over vertebral bodies L1-L3, respectively. PMID- 11105710 TI - MRI of destructive achilles tendon rupture associated with skin ulceration. AB - We present a case series of the MR findings of destructive tears of the Achilles tendon secondary to overlying soft tissue ulcerations. PMID- 11105711 TI - Optimization of contrast-enhanced MR angiography of the hands with a timing bolus and elliptically reordered 3D pulse sequence. AB - Our objective was to optimize bolus administration and sequence setting in gadolinium-enhanced magnetic resonance (MR) angiography of the hands. Elliptically reordered three-dimensional (3D) spoiled gradient-echo sequence with non-slab-selective radio frequency excitation was optimized according to the measurements of arterial and venous time-signal curves in 21 patients. Great variations in bolus arrival time and arterio-venous transit time could be observed. In most patients high-quality arterial depiction could be obtained with minor venous contamination. Contrast-to-noise, spatial resolution, and selective arterial filling is still a challenge for 3D MR angiography of the hand but can be optimized using Gadolinium-BOPTA and a dedicated pulse sequence setting with exact bolus timing. PMID- 11105712 TI - MR-guided and MR-monitored neurosurgical procedures at 1.5 T. AB - A combined MR suite and operating room (MR-OR) has been developed and extensively assessed for its use in a wide spectrum of therapeutic applications. Equipped with a 1.5 T short bore clinical MR scanner and standard neurosurgical OR equipment, in this MR surgical suite, surgeons can obtain intraoperative planar and volumetric MR images with superior soft tissue contrast and spatial resolution for surgical planning, guidance, and monitoring. Besides MR morphologic imaging capability, blood oxygen level-dependent functional MRI and proton MR spectroscopic imaging have been demonstrated intraoperatively in the same MR-OR to aid in surgical planning and guide tumor resections. A perspective surgical navigation device and remotely operated instrument have been developed and successfully used to assist surgeons in aligning and introducing biopsy needles under fluoroscopic MRI in brain biopsy procedures. Furthermore, surgical complications can be assessed immediately before the closure. There are numerous advantages offered by this unprecedented MR-guided surgical approach, most of which are demonstrated and presented herein. Since 1997, >270 neurosurgical cases (42% brain biopsies, 25% tumor resections, 11% functional neurosurgeries, 10% cyst drainages and shunt placements, and 12% others) have been performed in the MR-OR with a <1% overall complication rate. The tumor recurrence rate for the MR guided surgical approach is significantly less than that of the conventional one. Exemplary neurosurgical cases that have been performed in the MR-OR suite within the last 24 months are included. Overall, this high magnetic field approach to the MR-guided minimally invasive surgical procedures has been shown to be practical and acceptable to neurosurgeons as well as to neuroradiologists for a wide range of neurosurgical and neuroradiologic applications. PMID- 11105713 TI - The significance of bilateral CSI changes for the postoperative outcome in temporal lobe epilepsy. AB - PURPOSE: In a prospective study, we evaluated the significance of preoperative bilateral chemical shift spectroscopy imaging (CSI) changes for the prognosis of postoperative seizure outcome in the surgical treatment of patients with temporal lobe epilepsy (TLE). METHOD: CSI using multivoxel spectroscopy was performed. Twenty-six consecutive TLE patients scheduled for epilepsy surgery were included. To evaluate the value of the CSI with respect to postoperative seizure outcome, discriminant analysis with ipsilateral and contralateral CSI was performed. RESULTS: The discriminant analysis showed that the contralateral metabolic changes alone are able to predict seizure outcome whereby 92.3% of cases were correctly classified. Upon comparison of the two groups of seizure-free and non seizure-free patients with respect to contralateral metabolic changes, the difference proved to be highly significant (paired t test: t = -6.3, df = 24, p < 0.001). CONCLUSION: Bilateral metabolic CSI changes have a predictive value for the postoperative outcome in patients with TLE. In patients with severe bilateral metabolic changes, poor seizure outcome is a likely result. PMID- 11105714 TI - fMRI for preoperative neurosurgical mapping of motor cortex and language in a clinical setting. AB - PURPOSE: Identification of the precentral gyrus can be difficult in patients with brain tumors. The purpose of the current study was to evaluate the clinical usefulness of functional MRI (fMRI) in identifying motor cortex and speech areas as a part of preoperative neurosurgical planning. METHOD: fMRI was performed using a 1.5 T MR unit in 41 patients with brain tumors. The motor paradigm was finger tapping and foot movement, whereas the language paradigm consisted of a two word semantic test. Statistical analysis of the data was done using the Kolmogorow-Smirnow test. Plots of signal intensities over time were created. RESULTS: The precentral gyrus was identified in 38 of 41 patients. In two patients, fMRI was not of acceptable quality due to motion artifacts. Speech areas were localized in 33 patients. In a typical clinical setting, the value of the method was graded "high." CONCLUSION: fMRI's efficacy in the preoperative localization of language and motor areas is high. The method should become a routine adjunct for preoperative evaluation of brain tumors in the near future. PMID- 11105715 TI - Investigation of alternating and continuous experimental task designs during single finger opposition fMRI: a comparative study. AB - The purpose of this study was to empirically investigate and compare the effects of alternating and continuous experimental task designs on blood oxygenation level dependent (BOLD) signal contrast. Six healthy volunteers underwent single finger opposition functional magnetic resonance imaging (fMRI) using T2*-weighted echo planar imaging technique on a 1.5 T MR scanner. Two different acquisition patterns were tested: alternating (ABABAB) and continuous (AAABBB), rest: A, activation: B. The BOLD signal contrast within a primary motor cortex region of interest (ROI) was evaluated using normalized t-values (z-scores) and mean region of interest (ROI) intensity for the two patterns. Analysis of variance (ANOVA) on ROI mean z-score and signal intensities demonstrate that the alternating pattern of administering rest and activation epochs produced a more robust statistical difference than a continuous pattern. The results showed that different patterns of acquisition yield differences in the BOLD signal at field strength of 1.5 T, and that an alternating task design can be considered more optimal than a continuous task design. PMID- 11105716 TI - The impact of peak saturation of the arterial input function on quantitative evaluation of dynamic susceptibility contrast-enhanced MR studies. AB - PURPOSE: The purpose of this work was to investigate systematic errors in dynamic contrast-enhanced MR perfusion studies due to peak saturation of the arterial input function (AIF) and to introduce a simple correction algorithm. METHOD: Computer simulations were performed to evaluate the influence of AIF peak saturation and to demonstrate the effectiveness of the presented correction algorithm. To compare the computer simulations with real MR data, MR perfusion measurements were performed on volunteers. RESULTS: The computer simulations show that AIF peak saturation leads to a systematic overestimation of cerebral blood volume (CBV) and cerebral blood flow (CBF) values, which was confirmed by comparing the obtained MR data with PET results. With use of an improved calculation algorithm correcting for AIF peak saturation, a significant improvement of the obtained CBV and CBF values could be demonstrated. CONCLUSION: Our results suggest that AIF peak saturation leads to a significant systematic error in the determination of CBV and CBF values and has necessarily to be taken into account for dynamic contrast-enhanced MR perfusion studies. PMID- 11105717 TI - High-resolution MR venography at 3.0 Tesla. AB - PURPOSE: The aim of this study was to investigate the visualization of small venous vessels in the normal human brain at a field strength of 3 Tesla. METHODS: T2*-weighted, three-dimensional gradient-echo images were acquired by exploiting the magnetic susceptibility difference between oxygenated and deoxygenated hemoglobin in the vasculature and microvasculature. The spatial resolution was 0.5 x 0.5 x 1 mm3, and sequence parameters were varied to obtain good vessel delineation. Improved visibility of venous vessels was obtained by creating phase mask images from the magnetic resonance phase images and multiplying these by the magnitude images. Venograms were created by performing a minimum intensity projection over targeted volumes. RESULTS: Highly detailed visualization of venous structures deep in the brain and in the superficial cortical areas were obtained without administration of an exogenous contrast agent; compared with similar studies performed at 1.5 T, the echo time could be reduced from typically 40-50 ms to 17-28 ms. CONCLUSION: Imaging at high-field strength offers the possibility of improved resolution and the delineation of smaller vessels compared with lower field strengths. PMID- 11105718 TI - Quantification and minimization of magnetic susceptibility artifacts on GRE images. AB - PURPOSE: The purpose of this work was to determine the optimal imaging parameters for minimization of metallic susceptibility artifacts during gradient echo (GRE) imaging. METHOD: We performed GRE imaging of titanium screws in a nickel-doped agarose gel phantom, systematically varying several parameters to characterize and quantify susceptibility artifacts. RESULTS: The greatest reduction in artifact size came from using a short TE; increasing the frequency matrix and decreasing the slice thickness also contributed substantially to reducing the artifact size. Whenever possible, implanted prostheses should be aligned with the main magnetic field to minimize artifact size. Parameters with negligible effect on artifact size included bandwidth, phase encode matrix, and field of view. CONCLUSION: Radiologists can easily adjust the above parameters in their imaging protocols to improve GRE image quality in patients with implanted metallic devices. PMID- 11105719 TI - Hypersensitivity pneumonitis: patterns on high-resolution CT. AB - Hypersensitivity pneumonitis, also known as extrinsic allergic alveolitis, is caused by inhalation of specific environmental organic antigens. This disease may have typical high-resolution CT findings that, in the appropriate clinical setting, can be sufficiently characteristic to allow a confident diagnosis without the need for a lung biopsy. In this pictorial essay, the high-resolution CT patterns of hypersensitivity pneumonitis are illustrated. The authors emphasize the correlation among the radiologic presentation, functional abnormalities, and pathologic findings. PMID- 11105720 TI - Anomalous systemic arterialization to normal basal segments of the left lower lobe: helical CT and CTA findings. AB - PURPOSE: The purpose of this work was to evaluate the helical CT and CT angiography (CTA) findings of anomalous systemic artery (ASA) to the basal segments of the left lower lobe (LLL). METHOD: Three patients (two had hemoptysis, one was asymptomatic) with blotchy nodular density in the LLL revealed on chest radiographs underwent helical CT and CTA. Bronchoscopy was performed in two of these patients. Angiography and surgery were performed in one patient. RESULTS: All three patients demonstrated characteristic helical CT and CTA findings including 1) a sigmoid-shaped ASA originating from the lower descending thoracic aorta, with a distal bulbous configuration and four arterial branches supplying the basal segments of the LLL; 2) absence of an interlobar pulmonary artery or presence of a small artery lateral to the truncus basalis; 3) engorged vascular markings in the basal segments of the LLL; and 4) normal tracheobronchial tree and lung parenchyma. CONCLUSION: The findings in the present three cases suggest that the use of invasive studies such as angiography or bronchoscopy may be obviated in the diagnosis of ASA to the LLL because diagnosis can be provided through a clear set of criteria on helical CT and CTA. PMID- 11105721 TI - Gemcitabine pulmonary toxicity: CT features. AB - The purpose of this case report is to describe the CT imaging features of pulmonary toxicity from gemcitabine, a relatively new chemotherapeutic agent, in three patients. CT features of gemcitabine pulmonary toxicity include ground glass opacity (n = 3), thickened septal lines (n = 3), and reticular opacities (n = 3). Distribution is diffuse and bilateral, and may be symmetric (n = 2) or asymmetric (n = 1). Clinical symptoms and imaging findings are potentially reversible with steroid therapy. PMID- 11105722 TI - Transthoracic lung hernia following esophagectomy. PMID- 11105723 TI - Sectional neuroanatomy of the upper limb I: brachial plexus. AB - This series of three articles is structured to provide anatomically accurate functional schematics of the motor and sensory innervation of the shoulder and upper limb. This first paper provides radiographically oriented sagittal sections through the brachial plexus to assist in directly identifying a plexal lesion. A coronal schematic of the brachial plexus and summary table allows prediction of unique patterns of denervation from 19 lesion sites. Correlation between the lesion and the denervation pattern ensures the lesion is, in fact, clinically significant. The next two articles will present a color-coded atlas that allows the radiologist to quickly assess patterns of denervated muscles and thereby indirectly localize the lesion site. Thus, the three articles can be used together to predict the clinical picture for a given nerve lesion or extrapolate lesion location when a constellation of denervated muscles are seen on an upper limb magnetic resonance imaging or electromyographic study. PMID- 11105724 TI - Medical and genetic privacy. PMID- 11105725 TI - The November elections: what's at stake for the health of African Americans. PMID- 11105726 TI - Recommendations of the clinical trials consensus panel. National Medical Association. PMID- 11105727 TI - The intersection of race, gender, and primary care: results from the Women Physicians' Health Study. AB - The Women Physicians' Health Study is a nationally distributed mailed questionnaire survey of a random sample of 4501 female physicians. We examined differences in the professional characteristics and personal health habits of minority women physicians compared to other women physicians, with regard to the choice of primary care specialties, type or location of practice site, and career satisfaction. Most women physicians were self-described as non-Hispanic white (77.4%), with 13% Asians, and few blacks (4.3%) or Hispanics (5.2%). Blacks and Hispanics were more likely to choose primary care specialties (61.6% and 57.9%, respectively, vs. 49.3% of whites, p < 0.05). Black and Hispanic physicians were most likely to practice in urban areas (71.8% and 72.2%, respectively, p < 0.001). Minority physicians were most likely to report spending some time each week on clinical work for which they did not expect compensation. Black physicians were least likely to report high levels of work control and were least likely to be satisfied with their careers. While most physicians were compliant with the examined recommendations of the U.S. Preventive Services Task Force, we did find significant differences by ethnicity in compliance with clinical breast exams, mammograms, and pap smears. In conclusion, there continues to be fewer blacks and Hispanics in the U.S. physician workforce than in the general population. Minority women physicians are more likely to provide primary care services in communities that have been traditionally underserved and may also report higher rates of career dissatisfaction. PMID- 11105728 TI - Parameters of obesity in African-American women. AB - Non-Hispanic African-American women have the highest incidence of overweight in the United States, at 48%. For non-Hispanic white females, the prevalence is 32.9%. This striking difference can be expected to have a great impact on morbidity and mortality within this culture. The purpose of this study is to ascertain, by descriptive analysis of data derived from a questionnaire, whether there are modifiable factors specific to African-American women that could lead to an increased prevalence of obesity. Forty adult, African-American obese women were given a questionnaire covering personal socioeconomics, dietary habits, educational level, exercise patterns, childhood exposures, and stress management. Data from the questionnaire were grouped and collated to determine whether specific trends could be discerned. This descriptive evaluation found that hair care issues had some effect on exercise patterns. In addition, a lack of childhood role models for exercise and an adult pattern of sedentary lifestyles appeared to be significant factors contributing to obesity. After this pilot study, a comparative study with a white group of obese women or a group of nonobese African-American women should be the next step in evaluation to further define and understand these observations. PMID- 11105729 TI - Association between increased levels of TNF-alpha, decreased levels of prealbumin and retinol-binding protein, and disease outcome. AB - We determined whether there is an association between tumor necrosis factor-alpha (TNF-alpha), undernutrition [prealbumin (PA) <160 mg/L, retinol binding protein (RBP) <30 mg/L], disease stage, outcome (death or survival), and race in children with leukemia. TNF-alpha, PA, and RBP were measured in 52 patients (0.8 to 17 years old): 18 African Americans, 34 whites; 27 newly diagnosed (ND), and 25 in clinical remission (CR). Mean levels of TNF-alpha were higher in patients than in 46 healthy children (p < 0.05), but were not different between ND and CR groups. Mean acute phase proteins (APP) were different among groups: ND > CR > controls (p < 0.05). Mean levels of PA and RBP were lower in patients than in controls (p < 0.051, and tended to be higher in CR than in ND patients. African-American patients had lower concentrations of TNF-alpha, PA, and RBP but higher APP than white patients (p < 0.05). CR patients and African-American patients who died tended to have higher levels of TNF-alpha and APP, but lower PA and RBP than those who survived. A higher percentage of ND African Americans (45%) than of ND whites (13%) died. Results suggest that undernutrition and inflammation in CR patients and African Americans were associated with poor survival, and that ND African Americans have a poorer outcome than whites independently of TNF-alpha levels. PMID- 11105730 TI - Colorectal cancer risk perceptions and screening intentions in a minority population. AB - This is a 2-year follow-up to a previously reported baseline paper. We focused on a predominantly low-income African-American population from a community health center and investigated the relationships among perceptions of perceived risks for colorectal cancer (CRC), concerns about getting CRC, screening intentions, and whether participants had a fecal occult blood test (FOBT) on schedule at follow-up. Baseline absolute risk did not predict screening intentions or being on schedule (15% of sample), nor did it predict follow-up perceived absolute risk, comparative risk, or CRC concerns. Participants who expressed greater perceived absolute risk, comparative risk, and concerns at follow-up were more likely to report thinking about or definitely planning to get an FOBT within the next 2 years (49% of the sample). In addition, baseline absolute risk and whether or not a person had an FOBT on schedule at baseline did not predict being on schedule at follow-up. A significant percentage of the population (20%) were not able to state whether their CRC risk was below average, average, or above average. In addition, 44% of the population viewed their risks as lower than their peers, and 58% reported being not at all or slightly concerned about getting CRC. These results suggest that educational efforts are needed especially for low-income minority populations to enhance knowledge and accuracy of risk perceptions for CRC and interventions that explicitly manipulate risk are needed to assess to what extent risk perceptions can be modified and subsequently affect screening. PMID- 11105731 TI - On the need for race- or ethnic- focused scientific research in the Journal of the National Medical Association. PMID- 11105732 TI - The new cholinesterase inhibitors for Alzheimer's disease, Part 2: illustrating their mechanisms of action. PMID- 11105733 TI - Adverse neuropsychiatric reactions to herbal and over-the-counter "antidepressants". AB - BACKGROUND: Many unregulated over-the-counter agents for the treatment of depression are now available to patients and consumers. The potential for adverse neuropsychiatric effects with these agents has not been systematically studied in most cases. DATA SOURCES: The author performed a MEDLINE search on a variety of herbal and nonherbal over-the-counter agents said to be useful in the treatment of depression. The Physicians' Desk Reference for Herbal Medicines was also consulted. DATA SYNTHESIS: Although many of the herbal agents said to have benefits in depression appear to be safe, serious neuropsychiatric side effects and interactions have been reported for several over-the-counter "antidepressants." There is reason to suspect underreporting of those adverse events. Moreover, there is very little evidence from systematic studies regarding the potential for drug-drug or herb-drug interactions with these over-the-counter agents. Vitamins and amino acids touted for the treatment of depression are also not without risk. CONCLUSION: Although some over-the-counter remedies for depression are probably safe and effective for as-yet unidentified subgroups of depressed individuals, more research is required before these agents can be recommended for routine use. Stricter U.S. Food and Drug Administration oversight of these agents is indicated. PMID- 11105734 TI - Treatment of dysthymia with sertraline: a double-blind, placebo-controlled trial in dysthymic patients without major depression. AB - BACKGROUND: The selective serotonin reuptake inhibitor sertraline has been shown to be efficacious and well tolerated for the treatment of major depressive disorder. Relatively few trials, however, have examined the role of pharmacotherapy in dysthymia without concurrent major depression. The current investigation focuses on the use of sertraline for the treatment of dysthymia. METHOD: In this 12-week, multicenter, double-blind study, 310 patients with a DSM III-R diagnosis of dysthymic disorder without concurrent major depression were randomly assigned to receive either sertraline (N = 158) or placebo (N = 152). Sertraline was initiated at a dose of 50 mg daily, with titration permitted to a maximum of 200 mg daily. The primary evaluation criteria were the Structured Interview Guide for the Hamilton Depression Rating Scale, Seasonal Affective Disorder Version (SIGH-SAD), the Montgomery-Asberg Depression Rating Scale (MADRS), and the Clinical Global Impressions-Severity of Illness (CGI-S) and Improvement (CGI-I) scales. RESULTS: Mean percentage reductions for the intent-to treat population in SIGH-SAD scores were 44.6% for the sertraline-treated group and 33.2% for the placebo-treated group (p = .03); MADRS scores, 43.6% and 33.0% (p = .02); and CGI-S scores, 32.8% and 22.8% (p = .02). A significantly greater proportion of the sertraline-treated group was classified as responders (defined for HAM-D and MADRS scores as a 50% score reduction and for CGI-I as a score of 1 or 2 by the final visit) and remitters (SIGH-SAD score < or = 8) relative to the placebo-treated group by the final visit. In addition, sertraline-treated patients experienced greater improvements in all 9 domains of the Battelle Quality of Life Questionnaire than placebo-treated patients did, with a significant difference observed in favor of sertraline in 8 of the 9 domains. The life satisfaction and social interaction quality of life domains showed significantly greater response in sertraline responders compared with placebo SIGH-SAD responders. Sertraline was well tolerated. Thirteen percent of the sertraline-treated group versus 8% of the placebo-treated group withdrew from therapy owing to adverse events (p = .14). CONCLUSION: Sertraline is efficacious and well tolerated in the short-term treatment of dysthymia without concurrent major depression. PMID- 11105735 TI - Paroxetine levels in postpartum depressed women, breast milk, and infant serum. AB - BACKGROUND: The purpose of this study was to determine the concentrations of paroxetine in maternal serum, breast milk, and infant serum samples and to estimate infant exposure through breastfeeding. METHOD: A total of 25 sample sets was obtained: I sample set each from 23 mother-infant dyads and 2 sample sets from 1 mother-infant dyad. All mothers met DSM-IV criteria for major depressive disorder. The maternal fixed dosage of paroxetine was 10, 20, or 40 mg/day for a minimum of 30 days before the samples were drawn. Samples were collected 6 hours after dose intake, and the concentration of paroxetine in each sample was determined using gas chromatography/mass spectrometry. The analytic method employed in this study is the most sensitive to date, with the ability to detect drug concentrations as low as 0.1 ng/mL. RESULTS: Detectable levels of paroxetine were present in all maternal serum samples and in 24 of the 25 breast milk samples. In all of the infant serum samples, the paroxetine concentrations were below the lower limit of quantification. No unusual adverse effects were reported in any of the infants. CONCLUSION: The results of this study demonstrate that paroxetine, like the other selective serotonin reuptake inhibitors studied to date, is excreted into the breast milk of nursing mothers. The mean infant dose of paroxetine was 1. 1% of the maternal dose. Although no short-term adverse effects were reported in any of the infants in this study, future studies are needed to address a more systematic method for observing and recording any adverse effects. In addition, future studies should incorporate follow-up studies in order to evaluate possible long-term effects of paroxetine exposure. PMID- 11105736 TI - Strategies for switching from conventional antipsychotic drugs or risperidone to olanzapine. AB - BACKGROUND: This study compared the efficacy and safety of 4 therapeutically relevant strategies for switching clinically stable patients from a conventional antipsychotic drug or risperidone to olanzapine. METHOD: Two hundred nine outpatients with a DSM-IV diagnosis of schizophrenia or schizo-affective disorder who were clinically stable while being treated with a conventional antipsychotic drug or risperidone were openly randomly assigned to either abrupt or gradual discontinuation of their prior antipsychotic drug. Patients were further randomly assigned in a double-blind fashion to immediate olanzapine initiation (olanzapine, 10 mg q.d. for 3 weeks) or stepwise initiation (a sequence of 1 week each on placebo; olanzapine, 5 mg q.d.; and olanzapine, 10 mg q.d.). The efficacy of these 4 switching paradigms was assessed using the Clinical Global Impressions (CGI)-Improvement scale, Patient's Global Impressions (PGI)-Improvement scale, and Positive and Negative Syndrome Scale (PANSS). Safety assessments included ratings for extrapyramidal symptoms, cognitive impairment, adverse events, laboratory parameters, weight change, and vital signs. RESULTS: The paradigm of gradual antipsychotic drug discontinuation combined with an initial full dose of olanzapine, 10 mg/day, had the most favorable efficacy and tolerability profile overall. By week 3, the majority of completing patients on all 4 switching paradigms were either improved or clinically unchanged (> 90%). No clinically significant differences between switching paradigms were seen in laboratory values or vital signs. CONCLUSION: In this study, switching clinically stable outpatients with a diagnosis of schizophrenia or schizoaffective disorder to olanzapine was most successful when a full therapeutic dose of olanzapine was immediately initiated while gradually discontinuing prior conventional antipsychotic drug or risperidone treatment. Overall, switching was achieved without increased vulnerability to relapse or to occurrence of clinically burdensome antipsychotic drug withdrawal symptoms in the majority of patients. PMID- 11105738 TI - The efficacy and safety of a new enteric-coated formulation of fluoxetine given once weekly during the continuation treatment of major depressive disorder. AB - BACKGROUND: A simple, once-weekly dosing regimen could be a convenient alternative for many patients during long-term treatment of depression. Such a strategy might also be effective for improving medication compliance and the outcome of continuation treatment. The safety and effectiveness of a new formulation of enteric-coated fluoxetine (90 mg) given once weekly was tested during the continuation treatment of major depressive disorder. METHOD: Patients meeting DSM-IV criteria for major depressive disorder with modified 17-item Hamilton Rating Scale for Depression (HAM-D-17) scores > or = 18 and Clinical Global Impressions-Severity of Illness scale (CGI-S) scores > or = 4 were treated 13 weeks with open-label 20 mg/day of fluoxetine in a multicenter U.S. study. Responders (N = 501) were randomly assigned to receive 20 mg of fluoxetine daily, placebo, or 90 mg of enteric-coated fluoxetine weekly for 25 weeks of double blind continuation treatment. The primary efficacy measure was the percentage of patients who relapsed. Time to relapse was tested over the 25-week continuation period using log-rank analyses of the Kaplan-Meier estimates of relapse rates. Additional analyses of efficacy included comparison of change from baseline to endpoint for the HAM-D-17, CGI-S, and HAM-D-28 subscales by last observation carried forward (LOCF). Safety measures included comparison of treatment-emergent adverse events, both spontaneous and solicited (using the Association for Methodology of Documentation in Psychiatry-Module 5), vital signs, and laboratory measures. RESULTS: Relapse rates for patients assigned to fluoxetine, either 20 mg daily or 90 mg weekly, were significantly lower than for placebo by log-rank analysis and LOCF analyses of secondary efficacy measures. Efficacy did not significantly differ between the 2 active drug groups by these measures. Enteric coated fluoxetine at a once-weekly dose of 90 mg was well tolerated, and its safety profile was similar to that of daily 20 mg of fluoxetine. CONCLUSION: The formulation of enteric-coated fluoxetine taken once weekly is effective, safe, and well tolerated for continuation treatment of depression in patients who responded to acute treatment with 20 mg/day of fluoxetine. Monitoring during long term treatment for evidence of sustained remission is important regardless of dosing regimen. PMID- 11105737 TI - A double-blind, placebo-controlled, prophylaxis study of lamotrigine in rapid cycling bipolar disorder. Lamictal 614 Study Group. AB - BACKGROUND: Patients with rapid-cycling bipolar disorder are often treatment refractory. This study examined lamotrigine as maintenance monotherapy for rapid cycling bipolar disorder. METHOD: Lamotrigine was added to patients' current psychotropic regimens and titrated to clinical effect during an open-label treatment phase. Stabilized patients were tapered off other psychotropics and randomly assigned to lamotrigine or placebo monotherapy for 6 months. Time to additional pharmacotherapy for emerging symptoms was the primary outcome measure. Secondary efficacy measures included survival in study (time to any premature discontinuation), percentage of patients stable without relapse for 6 months, and changes in the Global Assessment Scale and Clinical Global Impressions-Severity scale. Safety was assessed from adverse event, physical examination, and laboratory data. RESULTS: 324 patients with rapid-cycling bipolar disorder (DSM IV criteria) received open-label lamotrigine, and 182 patients were randomly assigned to the double-blind maintenance phase. The difference between the treatment groups in time to additional pharmacotherapy did not achieve statistical significance in the overall efficacy population. However, survival in study was statistically different between the treatment groups (p = .036). Analyses also indicated a 6-week difference in median survival time favoring lamotrigine. Forty-one percent of lamotrigine patients versus 26% of placebo patients (p = .03) were stable without relapse for 6 months of monotherapy. Lamotrigine was well tolerated; there were no treatment-related changes in laboratory parameters, vital signs, or body weight. No serious rashes occurred. CONCLUSION: This was the largest and only prospective placebo-controlled study of rapid-cycling bipolar disorder patients to date; results indicate lamotrigine monotherapy is a useful treatment for some patients with rapid-cycling bipolar disorder. PMID- 11105739 TI - Bupropion SR reduces periodic limb movements associated with arousals from sleep in depressed patients with periodic limb movement disorder. AB - BACKGROUND: Antidepressant-induced periodic limb movement disorder (PLMD) may limit the tolerability of some antidepressant medications and interfere with treatment response. Given the role of dopamine in PLMD and the effects of bupropion sustained-release (SR) on central dopaminergic function, we hypothesized that bupropion SR would not be associated with antidepressant induced PLMD. METHOD: In an expanded case-series design, we compared the effects of bupropion SR, after about 10 weeks of treatment, on measures of PLMD, depression, and sleep in 5 depressed (Research Diagnostic Criteria) patients who also met criteria for having pretreatment PLMD. Depression was measured using the Beck Depression Inventory and the Hamilton Rating Scale for Depression. Patients were considered to have PLMD if polysomnographic recordings showed > 5 periodic limb movements/hour of sleep that were associated with arousals from sleep. RESULTS: Bupropion SR treatment was associated with a reduction in measures of PLMD and an improvement in depression. CONCLUSION: These results show that bupropion SR is not associated with antidepressant-induced PLMD. Rather, bupropion SR treatment reduces objective measures of PLMD in depressed patients with the disorder. PMID- 11105740 TI - Fluoxetine versus sertraline and paroxetine in major depressive disorder: changes in weight with long-term treatment. AB - BACKGROUND: The effects of extended selective serotonin reuptake inhibitor (SSRI) treatment on weight are not well characterized. Also unknown is whether different agents have differential effects. To examine these questions, we assessed weight changes in patients randomly assigned to long-term treatment with fluoxetine, sertraline, or paroxetine. METHOD: Patients (N = 284) with major depressive disorder (DSM-IV) were randomly assigned to double-blind treatment with fluoxetine (N = 92), sertraline, (N = 96), or paroxetine (N = 96) for a total of 26 to 32 weeks. The mean percent change in weight was compared for each group, as was the number of patients who had > or = 7% weight increase from baseline. RESULTS: Patients (fluoxetine, N = 44; sertraline, N = 48; paroxetine, N = 47) who completed the trial were included in these analyses. Paroxetine-treated patients experienced a significant weight increase, fluoxetine-treated patients had a modest but nonsignificant weight decrease, and patients treated with sertraline had a modest but nonsignificant weight increase. The number of patients whose weight increased > 7% from baseline was significantly greater for paroxetine-treated compared with either fluoxetine-treated or sertraline-treated patients. CONCLUSION: Risk of weight gain during extended SSRI treatment differs depending on which SSRI is used. PMID- 11105741 TI - Schizophrenia-associated idiopathic unconjugated hyperbilirubinemia (Gilbert's syndrome). AB - BACKGROUND: Idiopathic unconjugated hyperbilirubinemia (Gilbert's syndrome) is a benign hyperbilirubinemia found in the general population. There has been only 1 previous report of Gilbert's syndrome occurring in schizophrenic patients. The present study was conducted to determine the frequency of Gilbert's syndrome in schizophrenic patients relative to patients with other psychiatric disorders. METHOD: Plasma bilirubin concentrations of every patient admitted to the psychiatric hospital during a 3-year period were collected, and patients were examined to exclude all other causes of hyperbilirubinemia. In addition, the psychiatric symptoms of schizophrenic patients (ICD-10 criteria) with hyperbilirubinemia were evaluated by the Positive and Negative Syndrome Scale (PANSS). RESULTS: Schizophrenic patients showed a significantly higher incidence of hyperbilirubinemia (p < .05) relative to patients suffering from other psychiatric disorders, and schizophrenic patients with hyperbilirubinemia showed significantly higher scores on the positive and general psychiatric subscales of the PANSS (p < .0001) than patients without hyperbilirubinemia. CONCLUSION: The apparently higher frequency of Gilbert's syndrome in schizophrenic patients may reflect a relationship between hyperbilirubinemia and schizophrenic psychosis. Hypothetical explanations, such as a possible genetic disposition for Gilbert's syndrome, an increased vulnerability of red cell membranes, and the role of estrogens in schizophrenic patients, are discussed. PMID- 11105742 TI - Reboxetine treatment of depression in Parkinson's disease. PMID- 11105743 TI - Olanzapine-induced neutropenia in patients with history of clozapine treatment: two case reports from a state psychiatric institution. PMID- 11105744 TI - Venlafaxine versus sertraline for major depressive disorder. PMID- 11105745 TI - Adverse events of fluoxetine: postmarketing compared with premarketing clinical trials. PMID- 11105746 TI - Diagnosing melancholia. PMID- 11105747 TI - Differences in quality of life domains and psychopathologic and psychosocial factors in psychiatric patients. AB - BACKGROUND: Although treatment of severe mental disorders should strive to optimize quality of life (QOL) for the individual patient, little is known about variations in QOL domains and related psychopathologic and psychosocial factors in patients suffering from schizophrenia, schizoaffective disorder, and/or mood disorders. We hypothesized that QOL in severe mental disorder patients would have a more substantial relationship with psychosocial factors than with illness associated factors. METHOD: A case-control, cross-sectional design was used to examine QOL of 210 inpatients who met DSM-IV criteria for a severe mental disorder and who were consecutively admitted to closed, open, and rehabilitation wards. Following psychiatric examination, 210 inpatients were assessed using standardized self-report measures of QOL, insight, medication side effects, psychological distress, self-esteem, self-efficacy, coping, expressed emotion, and social support. QOL ratings for patients and a matched control group (175 nonpatients) were compared. Regression and factor analyses were used to compare multidimensional variables between patients with schizophrenia and schizoaffective and mood disorders. RESULTS: In all QOL domains, patients were less satisfied than nonpatient controls. Patients with schizophrenia reported less satisfaction with social relationships and medication when compared with patients with schizoaffective and/or mood disorders. Regression analysis established differential clusters of predictors for each group of patients and for various domains of QOL. On the basis of the results of factor analysis, we propose a distress protection model to enhance life satisfaction for severe mental disorder patients. CONCLUSION: Psychosocial factors rather than psychopathologic symptoms affect subjective QOL of hospitalized patients with severe mental disorders. The findings enable better understanding of the combining effects of psychopathology and psychosocial factors on subjective life satisfaction and highlight targets for more effective intervention and rehabilitation. PMID- 11105749 TI - Sequence bias in edited kinetoplastid RNAs. AB - Uridylate residues (Us) are inserted and deleted at precise positions in mitochondrial transcripts of Trypanosoma brucei. These sequence changes are determined by interactions with small guide RNAs (gRNAs) that are complementary to edited sequence. Adenylate (A) and guanylate (G) residues in gRNAs across from editing sites pair with inserted Us. We evaluate whether sequence bias exists in the bases surrounding insertion sites. Upon analyzing all reported insertion sites in T. brucei, we find that the predicted base pairs flanking insertion sites show a strong bias. Specifically, guiding As and Gs tend to be flanked by cytosine residues and Us. This bias is expected if precise base-pair interactions at the editing site determine the number of inserted Us. PMID- 11105748 TI - Coupled nucleotide covariations reveal dynamic RNA interaction patterns. AB - Evolutionarily conserved structures in related RNA molecules contain coordinated variations (covariations) of paired nucleotides. Analysis of covariations is a very powerful approach to deduce phylogenetically conserved (i.e., functional) conformations, including tertiary interactions. Here we discuss conserved RNA folding pathways that are revealed by covariation patterns. In such pathways, structural requirements for alternative pairings cause some nucleotides to covary with two different partners. Such "coupled" covariations between three or more nucleotides were found in various types of RNAs. The analysis of coupled covariations can unravel important features of RNA folding dynamics and improve phylogeny reconstruction in some cases. Importantly, it is necessary to distinguish between multiple covariations determined by mutually exclusive structures and those determined by tertiary contacts. PMID- 11105750 TI - Bypassing the rRNA processing endonucleolytic cleavage at site A2 in Saccharomyces cerevisiae. AB - Rrp5p is the only ribosomal RNA processing trans-acting factor that is required for the synthesis of both 18S and 5.8S rRNAs in Saccharomyces cerevisiae. Mutational analyses have characterized modified forms of Rrp5p that either affect formation of 18S rRNA by inhibiting cleavage at sites A0/A1/A2, or synthesis of 5.8S rRNA by inhibiting cleavage at site A3. Here, we examine the rRNA maturation process associated with a RRP5 bipartite allele that codes for two noncontiguous parts of the protein. This slow-growing bipartite mutant has a unique rRNA processing phenotype that proceeds without endonucleolytic cleavage at site A2. In wild-type cells, the A2 cleavage takes place on the 32S pre-rRNA and is responsible for the formation of 20S and 27SA2 species, the precursors of mature 18S and 5.8S/25S rRNAs, respectively. In the bipartite strain, such precursors were not detectable as judged by Northern analysis or in vivo labeling. They were replaced by the aberrant 21S species and the bypassing 27SA3 precursor, both descended from direct cleavage of 32S pre-rRNA at site A3, which provides an alternative rRNA maturation pathway in this strain. The 21S pre-rRNA is the sole detectable and most likely available precursor of 18S rRNA in this particular strain, indicating that 18S rRNA can be directly produced from 21S. Furthermore, 21S species were found associated with 43S preribosomal particles as similarly observed for the 20S pre-rRNA in the wild-type cells. PMID- 11105751 TI - Deletion of a conserved dinucleotide inhibits the second step of group II intron splicing. AB - Few point mutations have been described that specifically inhibit the second step of group II intron splicing. Furthermore, the effects of such mutations are typically not apparent unless the mutations are studied in the context of a substrate that harbors a very short 5' exon. Truncation of the 5' exon slows the second step of splicing. Once the second step has been slowed, the effects of point mutations can be seen. We report the unexpected observation that the deletion of a conserved GA dinucleotide dramatically inhibits the second step of splicing, even when the mutation is studied in the context of a full-length substrate. In contrast, we find that this mutation does not significantly affect the first step of splicing, unless the mutation is studied in combination with a second point mutation that is known to inhibit the first step. Even in that context, the effect of the GA deletion mutation on the first step is modest. These observations, together with the inferred location of the GA dinucleotide in the three-dimensional structure of the intron, suggest that this dinucleotide plays a particularly important role in the second step of splicing. PMID- 11105752 TI - Crystal structure of an RNA duplex containing phenyl-ribonucleotides, hydrophobic isosteres of the natural pyrimidines. AB - Chemically modified nucleotide analogs have gained widespread popularity for probing structure-function relationships. Among the modifications that were incorporated into RNAs for assessing the role of individual functional groups, the phenyl nucleotide has displayed surprising effects both in the contexts of the hammerhead ribozyme and pre-mRNA splicing. To examine the conformational properties of this hydrophobic base analog, we determined the crystal structure of an RNA double helix with incorporated phenyl ribonucleotides at 1.97 A resolution. In the structure, phenyl residues are engaged in self-pairing and their arrangements suggest energetically favorable stacking interactions with 3' adjacent guanines. The presence of the phenyl rings in the center of the duplex results in only moderate changes of the helical geometry. This finding is in line with those of earlier experiments that showed the phenyl analog to be a remarkably good mimetic of natural base function. Because the stacking interactions displayed by phenyl residues appear to be similar to those for natural bases, reduced conformational restriction due to the lack of hydrogen bonds with phenyl as well as alterations in its solvent structure may be the main causes of the activity changes with phenyl-modified RNAs. PMID- 11105753 TI - Tissue-specific autoregulation of Drosophila suppressor of forked by alternative poly(A) site utilization leads to accumulation of the suppressor of forked protein in mitotically active cells. AB - The Suppressor of forked protein is the Drosophila homolog of the 77K subunit of human cleavage stimulation factor, a complex required for the first step of the mRNA 3'-end-processing reaction. We have shown previously that wild-type su(f) function is required for the accumulation of a truncated su(f) transcript polyadenylated in intron 4 of the gene. This led us to propose a model in which the Su(f) protein would negatively regulate its own accumulation by stimulating 3'-end formation of this truncated su(f) RNA. In this article, we demonstrate this model and show that su(f) autoregulation is tissue specific. The Su(f) protein accumulates at a high level in dividing tissues, but not in nondividing tissues. We show that this distribution of the Su(f) protein results from stimulation by Su(f) of the tissue-specific utilization of the su(f) intronic poly(A) site, leading to the accumulation of the truncated su(f) transcript in nondividing tissues. Utilization of this intronic poly(A) site is affected in a su(f) mutant and restored in the mutant with a transgene encoding wild-type Su(f) protein. These data provide an in vivo example of cell-type-specific regulation of a protein level by poly(A) site choice, and confirm the role of Su(f) in regulation of poly(A) site utilization. PMID- 11105754 TI - Specificities of Caenorhabditis elegans and human hairpin binding proteins for the first nucleotide in the histone mRNA hairpin loop. AB - The 3' ends of animal replication-dependent histone mRNAs are formed by endonucleolytic cleavage of the primary transcripts downstream of a highly conserved RNA hairpin. The hairpin-binding protein (HBP) binds to this RNA element and is involved in histone RNA 3' processing. A minimal RNA-binding domain (RBD) of approximately 73 amino acids that has no similarity with other known RNA-binding motifs was identified in human HBP [Wang Z-F et al., Genes & Dev, 1996, 10:3028-3040]. The primary sequence identity between human and Caenorhabditis elegans RBDs is 55% compared to 38% for the full-length proteins. We analyzed whether differences between C. elegans and human HBP and hairpins are reflected in the specificity of RNA binding. The C. elegans HBP and its RBD recognize only their cognate RNA hairpins, whereas the human HBP or RBD can bind both the mammalian and the C. elegans hairpins. This selectivity of C. elegans HBP is mostly mediated by the first nucleotide in the loop, which is C in C. elegans and U in all other metazoans. By converting amino acids in the human RBD to the corresponding C. elegans residues at places where the latter deviates from the consensus, we could identify two amino acid segments that contribute to selectivity for the first nucleotide of the hairpin loop. PMID- 11105755 TI - The human endogenous retrovirus K Rev response element coincides with a predicted RNA folding region. AB - Human endogenous retrovirus K (HERV-K) is the name given to an approximately 30 million-year-old family of endogenous retroviruses present at >50 copies per haploid human genome. Previously, the HERV-K were shown to encode a nuclear RNA export factor, termed K-Rev, that is the functional equivalent of the H-Rev protein encoded by human immunodeficiency virus type 1. HERV-K was also shown to contain a cis-acting target element, the HERV-K Rev response element (K-RRE), that allowed the nuclear export of linked RNA transcripts in the presence of either K-Rev or H-Rev. Here, we demonstrate that the functionally defined K-RRE coincides with a statistically highly significant unusual RNA folding region and present a potential RNA secondary structure for the approximately 416-nt K-RRE. Both in vitro and in vivo assays of sequence specific RNA binding were used to map two primary binding sites for K-Rev, and one primary binding site for H-Rev, within the K-RRE. Of note, all three binding sites map to discrete predicted RNA stem-loop subdomains within the larger K-RRE structure. Although almost the entire 416-nt K-RRE was required for the activation of nuclear RNA export in cells expressing K-Rev, mutational inactivation of the binding sites for K-Rev resulted in the selective loss of the K-RRE response to K-Rev but not to H-Rev. Together, these data strongly suggest that the K-RRE, like the H-RRE, coincides with an extensive RNA secondary structure and identify specific sites within the K-RRE that can recruit either K-Rev or H-Rev to HERV-K RNA transcripts. PMID- 11105756 TI - Functional analyses of interacting factors involved in both pre-mRNA splicing and cell cycle progression in Saccharomyces cerevisiae. AB - Through a genetic screen to search for factors that interact with Prp17/Cdc40p, a protein involved in both cell cycle progression and pre-mRNA splicing, we identify three novel factors, which we call Syf1p, Syf2p, and Syf3 (SYnthetic lethal with cdc Forty). Here we present evidence that all three proteins are spliceosome associated, that they associate weakly or transiently with U6 and U5 snRNAs, and that Syf1p and Syf3p (also known as Clf1p) are required for pre-mRNA splicing. In addition we show that depletion of Syf1p or Syf3p results in cell cycle arrest at the G2/M transition. Thus, like Prp17/Cdc40p, Syf1p and Syf3p are involved in two distinct cellular processes. We discuss the likelihood that Syf1p, Syf2p, and Syf3p are components of a protein complex that assembles into spliceosomes and also regulates cell cycle progression. PMID- 11105757 TI - Polysome distribution of phospholipid hydroperoxide glutathione peroxidase mRNA: evidence for a block in elongation at the UGA/selenocysteine codon. AB - The translation of mammalian selenoprotein mRNAs requires the 3' untranslated region that contains a selenocysteine insertion sequence (SECIS) element necessary for decoding an in-frame UGA codon as selenocysteine (Sec). Selenoprotein biosynthesis is inefficient, which may be due to competition between Sec insertion and termination at the UGA/Sec codon. We analyzed the polysome distribution of phospholipid hydroperoxide glutathione peroxidase (PHGPx) mRNA, a member of the glutathione peroxidase family of selenoproteins, in rat hepatoma cell and mouse liver extracts. In linear sucrose gradients, the sedimentation velocity of PHGPx mRNA was impeded compared to CuZn superoxide dismutase (SOD) mRNA, which has a coding region of similar size. Selenium supplementation increased the loading of ribosomes onto PHGPx mRNA, but not CuZn SOD mRNA. To determine whether the slow sedimentation velocity of PHGPx mRNA is due to a block in elongation, we analyzed the polysome distribution of wild-type and mutant mRNAs translated in vitro. Mutation of the UGA/Sec codon to UGU/cysteine increased ribosome loading and protein synthesis. When UGA/Sec was replaced with UAA or when the SECIS element core was deleted, the distribution of the mutant mRNAs was similar to the wild-type mRNA. Addition of SECIS-binding protein SBP2, which is essential for Sec insertion, increased ribosome loading and translation of wild-type PHGPx mRNA, but had no effect on the mutant mRNAs. These results suggest that elongation is impeded at UGA/Sec, and that selenium and SBP2 alleviate this block by promoting Sec incorporation instead of termination. PMID- 11105758 TI - Alternative designs for construction of the class II transfer RNA tertiary core. AB - The structural requirements for assembly of functional class II transfer RNA core regions have been examined by sequence analysis and tested by reconstruction of alternative folds into the tertiary domain of Escherichia coli tRNA(2)Gln. At least four distinct designs have been identified that permit stable folding and efficient synthetase recognition, as assessed by thermal melting profiles and glutaminylation kinetics. Although most large variable-arm tRNAs found in nature possess an enlarged D-loop, lack of this feature can be compensated for by insertion of nucleotides either 3' to the variable loop or within the short acceptor/D-stem connector region. Rare pyrimidines at nt 9 in the core region can be accommodated in the class II framework, but only if specific nucleotides are present either in the D-loop or 3' to the variable arm. Glutaminyl-tRNA synthetase requires one or two unpaired uridines 3' to the variable arm to efficiently aminoacylate several of the class II frameworks. Because there are no specific enzyme contacts in the tRNAGln core region, these data suggest that tRNA discrimination by GlnRS depends in part on indirect readout of RNA sequence information. PMID- 11105759 TI - The Ct-RAE1 protein interacts with Balbiani ring RNP particles at the nuclear pore. AB - RAE1 is an evolutionarily conserved protein that associates with both mRNPs and nucleoporins, and may bridge the interaction between mRNP export cargoes and the nuclear pore complex (NPC). However, the mechanism by which RAE1 functions in mRNA export is still unknown and the time point at which RAE1 interacts with the exported RNP has not been directly investigated. Here we have addressed this question in the Balbiani ring (BR) system of Chironomus tentans using immunoelectron microscopy. The RAE1 protein of C. tentans, Ct-RAE1, is 70% identical to human RAE1/mrnp41 (hRAE1) and is recognized by antibodies raised against hRAE1. As in vertebrate cells, Ct-RAE1 is concentrated at the nuclear envelope and also dispersed throughout the nuclear interior. Here we show that Ct RAE1 does not bind to the BR particle either cotranscriptionally or in the nucleoplasm. Instead, the interaction between Ct-RAE1 and the exported BR particle occurs at the NPC. Moreover, the localization of Ct-RAE1 at the NPC is correlated with the presence of an exported RNP in the NPC. Finally, the anti RAE1 antibody does not label the cytoplasmic side of BR particles in transit through the central channel, which indicates that Ct-RAE1 either remains anchored at the nuclear side of the NPC during translocation of the RNP through the central channel or becomes transiently associated with the RNP but is rapidly released into the cytoplasm. PMID- 11105760 TI - RBP45 and RBP47, two oligouridylate-specific hnRNP-like proteins interacting with poly(A)+ RNA in nuclei of plant cells. AB - Introns in plant nuclear pre-mRNAs are highly enriched in U or U + A residues and this property is essential for efficient splicing. Moreover, 3'-untranslated regions (3'-UTRs) in plant pre-mRNAs are generally UA-rich and contain sequences that are important for the polyadenylation reaction. Here, we characterize two structurally related RNA-binding proteins (RBPs) from Nicotiana plumbaginifolia, referred to as RBP45 and RBP47, having specificity for oligouridylates. Both proteins contain three RBD-type RNA-binding domains and a glutamine-rich N terminus, and share similarity with Nam8p, a protein associated with U1 snRNP in the yeast Saccharomyces cerevisiae. Deletion analysis of RBP45 and RBP47 indicated that the presence of at least two RBD are required for interaction with RNA and that domains other than RBD do not significantly contribute to binding. mRNAs for RBP45 and RBP47 and mRNAs encoding six related proteins in Arabidopsis thaliana are constitutively expressed in different plant organs. Indirect immunofluorescence and fractionation of cell extracts showed that RBP45 and RBP47 are localized in the nucleus. In vivo UV crosslinking experiments demonstrated their association with the nuclear poly(A)+ RNA. In contrast to UBP1, another oligouridylate-binding nuclear three-RBD protein of N. plumbaginifolia (Lambermon et al., EMBO J, 2000, 19:1638-1649), RBP45 and RBP47 do not stimulate mRNA splicing and accumulation when transiently overexpressed in protoplasts. Properties of RBP45 and RBP47 suggest they represent hnRNP-proteins participating in still undefined steps of pre-mRNA maturation in plant cell nuclei. PMID- 11105761 TI - Nascent 60S ribosomal subunits enter the free pool bound by Nmd3p. AB - Nmd3p from yeast is required for the export of the large (60S) ribosomal subunit from the nucleus (Ho et al., 2000). Here, we show that Nmd3p forms a stable complex with free 60S subunits. Using an epitope-tagged Nmd3p, we show that free 60S subunits can be coimmunoprecipitated with Nmd3p. The interaction was specific for 60S subunits; 40S subunits were not coimmunoprecipitated. Using this coprecipitation technique and pulse-chase labeling of ribosomal subunit proteins we showed that Nmd3p bound nascent subunits, consistent with its role in export. However, under conditions in which ribosome biogenesis was inhibited (e.g., inhibition of transcription with thiolutin, inhibition of transcription of ribosomal protein and RNA genes in a sly1-1 mutant at nonpermissive temperature, and inhibition of translation in a conditional prt1 mutant), Nmd3p remained associated with 60S subunits. In addition, Nmd3delta120, a truncated protein that lacked a nuclear localization signal, retained 60S binding. These results suggest that Nmd3p recruits nascent 60S subunits into the pool of free 60S subunits and exchanges on 60S subunits as they recycle during translation. PMID- 11105762 TI - Dual role for the RNA-binding domain of Xenopus laevis SLBP1 in histone pre-mRNA processing. AB - The replication-dependent histone mRNAs end in a conserved 26-nt sequence that forms a stem-loop structure. This sequence is required for histone pre-mRNA processing and plays a role in multiple aspects of histone mRNA metabolism. Two proteins that bind the 3' end of histone mRNA are found in Xenopus oocytes. xSLBP1 is found in the nucleus, where it functions in histone pre-mRNA processing, and in the cytoplasm, where it may control histone mRNA translation and stability. xSLBP2 is a cytoplasmic protein, inactive in histone pre-mRNA processing, whose expression is restricted to oogenesis and early development. These proteins are similar only in their RNA-binding domains (RBD). A chimeric protein (1-2-1) in which the RBD of xSLBP1 has been replaced with the RBD of xSLBP2 binds the stem-loop with an affinity similar to the original protein. The 1-2-1 protein efficiently localizes to the nucleus of the frog oocyte, but is not active in processing of histone pre-mRNA in vivo. This protein does not support processing in a nuclear extract, but inhibits processing by competing with the active SLBP by binding to the substrate. The 1-2-1 protein also inhibits processing of synthetic histone pre-mRNA injected into frog oocytes, but has no effect on processing of histone pre-mRNA transcribed from an injected histone gene. This result suggests that sequences in the RBD of xSLBP1 give it preferential access to histone pre-mRNA transcribed in vivo. PMID- 11105763 TI - Mapping of the RNA recognition site of Escherichia coli ribosomal protein S7. AB - Bacterial ribosomal protein S7 initiates the folding of the 3' major domain of 16S ribosomal RNA by binding to its lower half. The X-ray structure of protein S7 from thermophilic bacteria was recently solved and found to be a modular structure, consisting of an alpha-helical domain with a beta-ribbon extension. To gain further insights into its interaction with rRNA, we cloned the S7 gene from Escherichia coli K12 into a pET expression vector and introduced 4 deletions and 12 amino acid substitutions in the protein sequence. The binding of each mutant to the lower half of the 3' major domain of 16S rRNA was assessed by filtration on nitrocellulose membranes. Deletion of the N-terminal 17 residues or deletion of the B hairpins (residues 72-89) severely decreased S7 affinity for the rRNA. Truncation of the C-terminal portion (residues 138-178), which includes part of the terminal alpha-helix, significantly affected S7 binding, whereas a shorter truncation (residues 148-178) only marginally influenced its binding. Severe effects were also observed with several strategic point mutations located throughout the protein, including Q8A and F17G in the N-terminal region, and K35Q, G54S, K113Q, and M115G in loops connecting the alpha-helices. Our results are consistent with the occurrence of several sites of contact between S7 and the 16S rRNA, in line with its role in the folding of the 3' major domain. PMID- 11105764 TI - Yeast Rrp9p is an evolutionarily conserved U3 snoRNP protein essential for early pre-rRNA processing cleavages and requires box C for its association. AB - Pre-rRNA processing in eukaryotic cells requires participation of several snoRNPs. These include the highly conserved and abundant U3 snoRNP, which is essential for synthesis of 18S rRNA. Here we report the characterization of Rrp9p, a novel yeast U3 protein, identified via its homology to the human U3-55k protein. Epitope-tagged Rrp9p specifically precipitates U3 snoRNA, but Rrp9p is not required for the stable accumulation of this snoRNA. Genetic depletion of Rrp9p inhibits the early cleavages of the primary pre-rRNA transcript at A0, A1, and A2 and, consequently, production of 18S, but not 25S and 5.8S, rRNA. The hU3 55k protein can partially complement a yeast rrp9 null mutant, indicating that the function of this protein has been conserved. Immunoprecipitation of extracts from cells that coexpress epitope-tagged Rrp9p and various mutant forms of U3 snoRNA limits the region required for association of Rrp9p to the U3-specific box B/C motif. Box C is essential, whereas box B plays a supportive role. PMID- 11105765 TI - Use of terbium as a probe of tRNA tertiary structure and folding. AB - Lanthanide metals such as terbium have previously been shown to be useful for mapping metal-binding sites in RNA. Terbium binds to the same sites on RNA as magnesium, however, with a much higher affinity. Thus, low concentrations of terbium ions can easily displace magnesium and promote phosphodiester backbone scission. At higher concentrations, terbium cleaves RNA in a sequence-independent manner, with a preference for single-stranded, non-Watson-Crick base-paired regions. Here, we show that terbium is a sensitive probe of human tRNALys,3 tertiary structure and folding. When 1 microM tRNA is used, the optimal terbium ion concentration for detecting Mg2+-induced tertiary structural changes is 50-60 microM. Using these concentrations of RNA and terbium, a magnesium-dependent folding transition with a midpoint (KMg) of 2.6 mM is observed for unmodified human tRNALys,3. At lower Tb3+ concentrations, cleavage is restricted to nucleotides that constitute specific metal-binding pockets. This small chemical probe should also be useful for detecting protein induced structural changes in RNA. PMID- 11105766 TI - Drug-susceptible tuberculosis outbreak in a state correctional facility housing HIV-infected inmates--South Carolina, 1999-2000. AB - During 1999-2000, South Carolina's Department of Corrections (SCDC), Department of Health and Environmental Control (DHEC), and CDC investigated an outbreak of drug-susceptible tuberculosis (TB) that occurred in a state correctional facility housing human immunodeficiency virus (HIV)-infected inmates. All culture confirmed case-patients have been linked by IS6110-based DNA fingerprinting of Mycobacterium tuberculosis isolates. This report describes the outbreak investigation and illustrates the need for increased vigilance for TB in settings in which HIV-infected persons congregate. PMID- 11105767 TI - Update: West Nile Virus activity--Eastern United States, 2000. AB - Data reported to CDC through the West Nile Virus (WNV) Surveillance System have shown an increase in the geographic range of WNV activity in 2000 compared with 1999, the first year that WNV was reported in the Western Hemisphere. In response to this occurrence of WNV, 17 states along the Atlantic and Gulf coasts, New York City, and the District of Columbia conducted WNV surveillance, which included monitoring mosquitoes, sentinel chicken flocks, wild birds, and potentially susceptible mammals (e.g., horses and humans). In 1999, WNV was detected in four states (Connecticut, Maryland, New Jersey, and New York) . In 2000, epizootic activity in birds and/or mosquitoes was reported from 12 states (Connecticut, Delaware, Maryland, Massachusetts, New Hampshire, New Jersey, New York, North Carolina, Pennsylvania, Rhode Island, Vermont, and Virginia) and the District of Columbia. Of the 13 jurisdictions, seven also reported severe neurologic WNV infections in humans, horses, and/or other mammal species. This report presents surveillance data reported to CDC from January 1 through November 15. PMID- 11105768 TI - Measles, rubella, and congenital rubella syndrome--United States and Mexico, 1997 1999. AB - In 1996, the Immunization Working Group of the Mexico-United States Binational Commission was established to enhance coordination of disease surveillance, assure high vaccination coverage in both countries, and hasten the elimination of vaccine-preventable diseases. The United States and Mexico share the Pan American Health Organization (PAHO) goal of measles elimination by 2000. The United States also established a goal of eliminating indigenous rubella and congenital rubella syndrome (CRS) by 2000. This report summarizes the measles and rubella vaccination and surveillance data for the United States and Mexico for 1997-1999. PMID- 11105769 TI - A comparative study of the ability of different solvents and adsorbents to extract aroma compounds from alcoholic beverages. AB - Seven liquid solvent systems--dichloromethane, dichloromethane-pentane (1:1), freon 113, diethyl ether-pentane (1:1 and 1:9), ethyl acetate-pentane (with and without an additional salting-out effect) (1:3 and 1:20), and seven solid-phase extraction (SPE) systems (Amberlite XAD-2, 4, 7, and 16; Porapak Q; C8; and C18)- are comparatively studied. The distribution coefficients between the extraction system and a hydroalcoholic solution (12% v/v in ethanol, pH = 3.2) of 14 selected volatile compounds belonging to different chemical families and polarities are calculated. The results are processed by factor analysis and cluster analysis, and the following conclusions are reached. First, the efficiency of extraction decreases in this order: polymeric sorbents > silica based sorbents > liquid-liquid systems with salting-out effect approximately dichloromethane > rest of liquid solvents. Second, the addition of salt mainly increases the recovery of compounds with Lewis acid properties. Third, the efficiency of the extraction of a liquid solvent depends not only on its polarity but also on its solubility in water. Fourth, in regards to the selectivity of the SPE systems, Porapak Q is the best to extract nonpolar compounds, Amberlite XAD 4 and 16 provide the least selective extraction profiles, and C8 and C18 have a special ability to extract compounds with a Bronstedt-Lowry character. Results indicate that in all cases liquid solvents can be replaced satisfactorily by SPE systems. PMID- 11105771 TI - An improved configuration of a moving wire transport detector. AB - An improved design of the transport detector is described that uses a pre oxidized titanium ribbon as the transport medium. The titanium ribbon has a high loading capacity that permits a large proportion of the total column eluent to be taken into the sensing system. The solute is sensed by pyrolysis and the subsequent detection of pyrolysis products by a miniature argon detector. The pyrolyzer and sensor system is designed to ensure that all the pyrolysis products enter the detector with minimum dilution and band dispersion. As a result, the sensitivity of the detector (or minimum detectable concentration) has been reduced by approximately two orders of magnitude compared with the original design. The sensitivity of the system described to sucrose is 8 x 10(-8) g/mL, which is similar to the sensitivity of the fixed-wavelength UV detector to benzene (approximately 5 x 10(-8) g/mL). It would appear that the new design has potentially a sensitivity at least an order of magnitude lower than that reported here. PMID- 11105770 TI - Feasibility of on-line supercritical fluid extraction of steroids from aqueous based matrices with analysis via gas chromatography-mass spectrometry. AB - The solubility of testosterone, boldenone, androstenone, etiocholanolone, and epitestosterone are measured in pure supercritical CO2. Testosterone exhibited the highest solubility in supercritical CO2. The solubility of all steroids except epitestosterone increased by one order of magnitude with increasing pressure from 100 to 400 atm. Epitestosterone had the lowest solubility in supercritical CO2 and its solubility was not affected by pressure. The extraction efficiency of steroids from an aqueous saline environment exceeded 95%. Because of the partial solubility of water in supercritical CO2, the addition of a moisture trap after the aqueous vessel is necessary to prevent the plugging and deterioration of the gas chromatographic (GC) column. It is demonstrated that on line supercritical fluid extraction-GC-mass spectrometry is feasible for the quantitative extraction and analysis of steroids from both saline and urine solutions. However, it is determined that the adsorbent vessel filled with Hydromatrix is not sufficient to trap all the moisture, and after 3 to 4 extractions, the GC column efficiency lowered. PMID- 11105772 TI - Application of Convective Interaction Media (CIM) disk monolithic columns for fast separation and monitoring of organic acids. AB - The separation of organic acids on the anion-exchange monolithic support, commercially available as Convective Interaction Media (CIM), is presented in this study. It is demonstrated that citric, isocitric, pyruvic, fumaric, malic, and alpha-ketoglutaric acid can be successfully separated using a CIM monolithic column of suitable user-adjustable length. The effect of the mobile phase composition on the separation is investigated. CIM monolithic columns of adjustable length from 3 to 18 mm are compared regarding the resolution and the back pressure. It is shown that the CIM monolithic column of 12 mm in length enables a good separation of all six organic acids within 3 min and exhibits a linear dependence of back pressure versus flow rate. The resolution and the dynamic binding capacity are found to be flow-unaffected. A filtrated sample of bioprocess supernatant is analyzed without previous pretreatment, which indicates the possibility of online monitoring of small molecules during the bioprocess using CIM monolithic columns. PMID- 11105773 TI - A new bonded porous polymer PLOT U column with increased polarity. AB - The separation features of a new type of PLOT U column are presented through many applications. This type of PLOT U column is coated with a divinylbenzene-ethylene glycol dimethacrylate copolymer. It has an increased polarity when compared with a conventional PLOT Q type column. The stationary phase of the PLOT U column is truly bonded, thus providing column rinsability and low column bleed. PMID- 11105774 TI - Characterizing the selectivity of stationary phases and organic modifiers in reversed-phase high-performance liquid chromatographic systems by a general solvation equation using gradient elution. AB - Retention data for a set of 69 compounds using rapid gradient elution are obtained on a wide range of reversed-phase stationary phases and organic modifiers. The chromatographic stationary phases studied are Inertsil (IN)-ODS, pentafluorophenyl, fluoro-octyl, n-propylcyano, Polymer (PLRP-S 100), and hexylphenyl. The organic solvent modifiers are 2,2,2-trifluoroethanol (TFE); 1,1,1,3,3,3-hexafluoropropan-2-ol (HFIP); isopropanol; methanol (MeOH); acetonitrile (AcN); tetrahydrofuran; 1,4-dioxane; N,N-dimethylformamide; and mixed solvents of dimethylsulfoxide (DMSO) with AcN and DMSO with MeOH (1:1). A total of 25 chromatographic systems are analyzed using a solvation equation. In general, most of the systems give reasonable statistics. The selectivity of the reversed phase-high-performance liquid chromatographic (HPLC) systems with respect to the solute's dipolarity-polarity, hydrogen-bond acidity, and basicity are reflected in correspondingly large coefficients in the solvation equation. We wanted to find the most orthogonal HPLC systems, showing the highest possible selectivity difference in order to derive molecular descriptors using the gradient retention times of a compound. We selected eight chromatographic systems that have a large range of coefficients of interest (s, a, and b) similar to those found in water-solvent partitions used previously to derive molecular descriptors. The systems selected are IN-ODS phases with AcN, MeOH, TFE, and HFIP as mobile phase, PLRP-S 100 phase with AcN, propylcyano phase with AcN and MeOH, and fluorooctyl phase with TFE. Using the retention data obtained for a compound in the selected chromatographic systems, we can estimate the molecular descriptors with the faster and simpler gradient elution method. PMID- 11105776 TI - What causes column voiding and is there any way of preventing or minimizing it from occurring? PMID- 11105775 TI - Improved analysis of a gas oil using a high-performance thin-layer chromatographic system. PMID- 11105777 TI - High levels of endorphin and related pathologies of veterinary concern. A review. AB - The authors report information about endogenous opioid peptides (EOP), receptors, antagonists and their interference with pain, stress, endocrine and immune system. A relationship between EOP and calcium homeostasis, both at extracellular and intracellular level, has been observed. In vitro, beta-endorphin exerts different actions through calcium channel functionality in epithelial cells. In rat aorta and cerebral cortex: beta-endorphin or Naloxone alternatively influence oocyte maturation through the mu-receptor gene expression and intracellular calcium concentration in granulosa and cumulus cells. Calcium channel block is removed by administrating Naloxone and calcium. In vivo, Naloxone and calcium removes EOP induced apoptosis in granulosa cells; is the most safe therapy in cow's milk fever; allow to remove ovarian follicular cysts. A negative influence of opioids on immune response after vaccination was established; EOP-related metabolic problems in post-partum cows. Abnormal intestinal motility, in which a Ca++ influence is well known, can be removed by Naloxone and calcium administration. Calcium-related function and neuromodulation must be re-evaluated since high level of EOP are involved in many pathologies through their influence on calcium activity. The use of calcium salts and Naloxone offers a safe and supplementary therapeutical possibility, active in any condition of altered endogenous opioids. PMID- 11105778 TI - TNF-alpha blockade by a dimeric TNF type I receptor molecule selectively inhibits adaptive immune responses. AB - Tumor necrosis factor-alpha (TNF-alpha) is a mediator of severe inflammatory processes, including rheumatoid arthritis. Suppression of TNF with a soluble type I or type II receptor molecule (TNF-RI or TNF-RII) has the potential to decrease cytokine levels and modulate inflammatory diseases in humans. However, it has recently been reported that treatment of mice with a TNF-RI:Fc immunoadhesin protein augmented Gram positive infections and subsequent mortality. To determine if TNF-alpha blockade with soluble TNF-alpha receptors might alter immune system function, assays were assessed in rodents treated with a dimeric form of the p55 TNF-RI, Tumor Necrosis Factor-binding protein (TNFbp). Administration of TNFbp resulted in suppression of primary and secondary IgG antibody responses and cell mediated immune function. No treatment-related differences were detected in immune-enhancing assays or non-specific immune function parameters. Bacterial host resistance assays with Listeria monocytogenes, Staphylococcus aureus or Escherichia coli showed an increase in tissue colony counts only with L. monocytogenes challenged animals following TNFbp administration. These results suggest that TNFbp has the capacity to inhibit adaptive immune function in experimental animal models. Studies suggest that while reducing TNF-alpha is important in controlling cytokine-dependent disease states, maintenance of a threshold level may be critical for normal immune function. PMID- 11105779 TI - Effect of a synthetic lipid immunomodulator on the regulation of the transcription factor NF-kappaB. AB - Macrophage activation plays a central role in host defense against a variety of pathogens via inducible messengers. The transcription factor NF-kappaB controls the synthesis of cytokines involved in immune responses. In quiescent cells, NF kappaB is located in the cytosol bound to an inhibitor IkappaB. Upon appropriate signal, NF-KB translocates to the nucleus and binds to DNA. The present study investigated the involvement of an immunomodulator, (diHDA-glycerol) on the NF kappaB/IkappaB complex. Results were compared to those obtained with lipopolysaccharide (LPS) as a major virulence factor in bacterial sepsis. Data showed that exposure of J774.1 cells either to LPS or diHDA-glycerol substantially increased with time the nuclear levels of NF-kappaB complexes. Antibodies to various NF-kappaB proteins supershifted p50, p65 and to a lesser extent c-rel. Western blot analyses showed a rapid cytosolic IkappaB-alpha turn over following LPS exposure in contrast to diHDA-glycerol treatment. Further experiments investigated the involvement of protein kinase C (PKC) by using two inhibitors, staurosporine and H7. Pretreatment of J774.1 with either inhibitor prior to diHDA-glycerol or LPS exposure decreased NF-kappaB activation. Our results indicate that diHDA-glycerol was acting on NF-kappaB through IkappaB regulative mechanisms differing from those used by LPS. DiHDA-glycerol is likely acting on many other transcription factors targeting distinct genes implied in up regulation of the immune system. PMID- 11105780 TI - Neutrophil peripheral count and human leukocyte elastase during chronic lithium carbonate therapy. AB - Plasma levels of human polymorphonuclear elastase (PMN-E) are considered a marker of granulocyte activation and can potentially complement the peripheral neutrophil count in laboratory and pathophysiological settings. Neutrophilic leukocytosis is a well known effect of lithium therapy, but there is no information about the concomitant behaviour of PMN-E in these patients. The aim of this study was to evaluate both polymorphonuclear leukocyte count and plasma PMN-E levels in depression patients undergoing chronic lithium therapy. Absolute and differential leukocyte count in venous peripheral blood was determined by an automated method, and PMN-E evaluated by enzyme immunoassay. 39 patients (11 males, 28 females; mean age 43. +/- 6.02) with depression disorders were studied, during lithium carbonate therapy. Neutrophilia (neutrophil count > 7.500x10(9) cells per liter) was found in 7 (18%) patients and an increase in plasma PMN-E levels (PMN-E > 56 microg per liter ) in 6 (15%). No correlations were found between neutrophil count, plasma concentration of PMN-E, plasma level of lithium and duration of therapy. The results show that in these patients, not only the PMN count but also elastase levels can exceed the normal range. The absence of correlation between these two parameters suggests that the state of PMN activation is not linked to their number in peripheral blood. PMID- 11105781 TI - Mercury exposure and murine response to Plasmodium yoelii infection and immunization. AB - Malaria has re-emerged in Amazonia over the past two decades. Many factors have been proposed for this, among them changes in population distribution, failures of vector control and pharmacologic management, and local as well as global environmental changes. Among the latter factors, we have studied the potential role of increasing exposures to the immunotoxic metal mercury, which is widely used in Amazonia for artisanal extraction of alluvial gold deposits. We report here that Hg impairs host resistance to malaria infection at exo-erythrocytic stages. Hg exposed mice have higher parasitemia following infection with sporozoites, but not after transfusion of infected red cells. In mice inoculated with irradiated sporozoites, Hg blocks acquisition of immunity. In addition Hg affects immunologic parameters that are known to be involved in host response to malaria infection. These results have potential implications for the incidence and prevalence of malaria among populations exposed to mercury from artisanal goldmining and consumption of contaminated fish regions with high rates of malaria and other infectious diseases. PMID- 11105782 TI - Potentiation of tumor necrosis factor-alpha-induced apoptosis by mistletoe lectin. AB - Mistletoe lectins (MLs) constitute the active principle in extract preparations from mistletoe, commonly used as immunomodulator in adjuvant tumor therapy. MLs, classified as type II ribosome inactivating proteins, inhibit protein synthesis. Inhibitors of protein synthesis may modify cancer cell response to tumor necrosis factor-alpha (TNF). In the present study, we have hypothesized that the anticancer efficacy of TNF may be potentiated by MLs. In deed, simultaneous treatment of human cervix carcinoma HeLa or breast carcinoma MCF-7 cells with MLs isolated from European or Korean mistletoe rendered them more sensitive to induction of apoptosis by TNF. The mechanism by which MLs amplify the effect of TNF may involve suppression of the survival protein synthesis. PMID- 11105783 TI - Immunomodulation of Bu-Zhong-Yi-Qi-Tang on in vitro granulocyte colony stimulating-factor and tumor necrosis factor-alpha production by peripheral blood mononuclear cells. AB - Bu-Zhong-Yi-Qi-Tang (BZYQT) is a Chinese medicine, and has been used for the treatment of hepatocellular carcinoma (HCC) patients. At present, we still do not fully understand the effects of BZYQT on the cellular physiology. Present in vitro study demonstrated that BZYQT is capable of increasing granulocyte colony stimulating-factor (G-CSF) and tumor necrosis factor-alpha (TNF-alpha) production by peripheral blood mononuclear cells (PBMC) in healthy volunteers and patients with HCC. The productions of G-CSF and TNF-alpha by PBMC of volunteers were significantly stimulated by more than 125 microg/ml of BZYQT. G-CSF levels stimulated by PBMC of healthy volunteers were higher than in PBMC of the HCC patients when more than 625 microg/ml of BZYQT was administrated. The reason may be due to the impaired immunologic reactivity of mononuclear cells in HCC patients. However, the production levels of TNF-alpha in HCC patients can be stimulated to levels as high as those in healthy volunteers. When adding high concentration (3.125 mg/ml) of BZYQT to the cultured PBMC, the increments of G CSF and TNF-alpha production decreased although there were no obvious changes in the number of metabolic active PBMC changed. TNF-alpha andG-CSF are known to play important roles in the biological defensive mechanism. These findings show that BZYQT is a unique formula for the stimulation of PBMC to produce G-CSF and TNF alpha. Administration of BZYQT may be beneficial for patients with HCC to modulate these cytokines. PMID- 11105784 TI - Stimulatory action of Pluchea quitoc extract on the hematopoietic response during murine listeriosis. AB - The importance of both granulocytes and macrophages in the response to Listeria monocytogenes infection make this infection a suitable choice to investigate the effects of Pluchea quitoc on hematopoiesis. A significant depletion of bone marrow granulocyte-macrophage progenitor cells (CFU-GM) was observed at 48 and 72 h after intraperitoneal infection of mice with 1 x 10(4) L. monocytogenes. However, the treatment of infected animals with P. quitoc ethanolic extract (250, 500 or 1000 mg/kg) given orally for 3 consecutive days prior to infection produced a stimulatory effect on myelopoiesis, restoring the number of CFU-GM to normal. This same dose-schedule also increased colony formation in normal mice as compared to controls. In addition, P. quitoc significantly enhanced survival of infected mice. Thus, it is probable that the ability of P. quitoc to induce a higher reserve of granulocyte-macrophage precursors in the bone marrow is of major significance in determining early resistance to infection. PMID- 11105785 TI - What is your diagnosis? Canine eosinophilic furunculosis (folliculitis) of the face. PMID- 11105786 TI - Oral medications for treating diabetes mellitus in dogs and cats. AB - Five classes of oral hypoglycaemic drugs and two trace minerals used to treat diabetes mellitus in humans are reviewed and current knowledge on the use of these drugs in diabetic dogs and cats is presented. Oral sulphonylurea drugs stimulate insulin secretion and have been used successfully to treat diabetes in cats but not dogs. Preliminary studies evaluating the efficacy of the biguanide, metformin, in diabetic cats have not been promising. Pharmacokinetic studies have been performed in healthy cats, but clinical studies evaluating the efficacy of the insulin-sensitising drugs, thiazolidinediones, have not been reported. Treatment with the alpha-glucosidase inhibitor, acarbose, improved control of glycaemia in diabetic dogs; similar studies have not been reported in cats. Although chromium picolinate did not improve control of glycaemia in diabetic dogs, vanadium has improved control of the abnormality in diabetic cats. PMID- 11105787 TI - Osteopenia and other radiographic signs in canine hyperadrenocorticism. AB - The specificity of conventional radiography in assessing canine hyperadrenocorticism was evaluated by comparing the Incidence of related radiographic findings in 24 hyperadrenocorticoid, 15 diabetic and 20 hypothyroid dogs. Hyperadrenocorticoid dogs showed significantly more perihilar bronchial mineralisation than other groups. There was no significant variation between the disease groups with respect to obesity, hepatomegaly, contour of the caudoventral hepatic margin, peripheral bronchial mineralisation or osteopenia. Adrenal mineralisation and calcinosis cutis were rare findings observed only in hyperadrenocorticoid dogs. The effect of obesity on the radiographic appearance of bone was studied using a dissected lumbar spine from a canine cadaver. An osteopenic effect could be demonstrated by superimposition of a 10 cm-thick fat block. The low specificity of almost all common signs in canine hyperadrenocorticism and the low incidence of characteristic findings demonstrate the limited potential of radiography in assessing this condition. Radiographic assessment of bone density is unreliable because of artefactual osteopenic effects of high kVp settings necessary in obese dogs. PMID- 11105788 TI - Propofol for treatment of refractory seizures in dogs and a cat with intracranial disorders. AB - Twelve dogs and one cat that were presented with seizures due to various disorders of intracranial origin were treated with one or several boluses of propofol (2 to 8 mg/kg). All the animals had received previous medication, including diazepam alone or in combination with phenobarbital and/or pentobarbital. Seizure control with prevention of further convulsions was achieved in 11 patients. In one dog, seizures kept recurring after periods of successful control following administration of propofol. Another dog with continuing seizures resistant to barbiturate therapy died following administration of propofol. This retrospective study suggests that propofol may be an effective drug in controlling status epilepticus resistant to conventional medication and may be used as an alternative to pentobarbital administration. PMID- 11105789 TI - Diagnostic features, confirmation and disease progression in 28 cases of lethal acrodermatitis of bull terriers. AB - Lethal acrodermatitis (LAD) is a genetically determined metabolic disease of bull terriers first described in the USA in the 1980s. In this study, the largest so far reported, 28 bull terriers born in the UK were diagnosed as suffering from LAD, and the clinical findings and the progression of the disease with time are described. The main characteristics of LAD are stunting, splayed digits, eating difficulties, skin disease of the face and feet, and increased susceptibility to microbial infections. In older dogs, paronychia, nail disease and hyperkeratosis of the footpads develops, becoming severe in dogs over six months of age. A diagnosis of LAD can be strongly suspected in any bull terrier showing a combination of the aforementioned signs from an early age. Dermatohistopathological demonstration of marked parakeratotic hyperkeratosis is strongly supportive of the diagnosis of LAD and, in association with the typical clinical findings, is sufficient to confirm a diagnosis. Although many of the clinical signs and the pathology of this condition suggest zinc deficiency, the measurement of blood zinc levels as a diagnostic aid is of limited value. PMID- 11105790 TI - Hypospadias in a Himalayan cat. AB - A one-year-old male Himalayan cat was presented with a history of chronic cystitis. Physical examination revealed that the cat had hypospadias. It was postulated that the abnormal urethral opening on the ventral aspect of the penis permitted faecal contamination of the preputial area and gave rise to the ascending infection. The hypospadias was surgically corrected and a complete recovery was achieved. PMID- 11105791 TI - Lateral glenohumeral ligament rupture in three dogs. AB - Three adult large breed dogs were evaluated for chronic forelimb lameness. Clinical examination localised pain to the area of the shoulder joint. Traditional imaging methods, including radiography, arthrography and ultrasonography, were unrewarding. Arthroscopy performed via a lateral portal demonstrated complete tears of the proximal part of the lateral glenohumeral ligament in all cases. Two of the three cases responded to treatment with intra articular methylprednisolone and rest with a resolution of the lameness, while the third failed to improve. Surgical intervention in this third case involved lateral capsulorraphy, and re-examination at five weeks postoperatively showed the dog to be without lameness. Tearing of the lateral glenohumeral ligament should be considered in the differential diagnosis of shoulder lameness. Surgical stabilisation should be considered in cases refractory to conservative treatment. PMID- 11105792 TI - Traumatic subcutaneous translocation of the spleen in an Old English sheepdog. AB - A 14-month-old entire male Old English sheepdog was presented with a slowly enlarging subcutaneous soft tissue swelling caudal to the last rib. Radiographic and ultrasonographic Investigations demonstrated a soft tissue mass closely adherent to the body wall, with a coarse hypoechoic pattern. Surgical exploration revealed a mass with the gross appearance of splenic tissue. The mass was adhered to a healed paracostal abdominal tear. Histological examination confirmed the resected tissue to be normal spleen. PMID- 11105793 TI - Resolution of skin lesions and long-term survival in a dog with superficial necrolytic dermatitis and liver cirrhosis. AB - A nine-year-old, neutered female Shetland sheepdog was presented with crusted, ulcerative skin lesions affecting the footpads, commissures of the lips and the lateral canthi of the eyes. Histopathological examination of skin biopsies revealed changes consistent with superficial necrolytic dermatitis and biochemical analysis demonstrated elevated liver enzymes. Abdominal radiography revealed a small liver which, on ultrasonography, appeared diffusely mottled and showed changes suggestive of periportal fibrosis. On exploratory laparotomy, the pancreas appeared normal, but the liver was small and had multiple nodules throughout the parenchyma. This appearance was confirmed as cirrhosis on histopathological examination. The dog was placed on a hepatic support diet and treated with colchicine, essential fatty acid supplementation and raw egg yolks. After four weeks, the skin lesions had resolved and the dog remained free of clinical signs over a 22-month follow-up period. PMID- 11105794 TI - Risk factors for obesity in the cat. PMID- 11105795 TI - Congress 2001: a taste of what's in store. PMID- 11105797 TI - Mild renal dysfunction is associated with insulin resistance in chronic glomerulonephritis. AB - BACKGROUND: Insulin resistance is associated with advanced and moderate chronic renal failure (CRF). However, insulin resistance in chronic glomerulonephritis (CGN) before onset of frank renal dysfunction is not fully evaluated. We attempted to investigate the association of insulin resistance with mild renal dysfunction and with abnormal calcium homeostasis. PATIENTS AND METHODS: Eighteen young, lean non-diabetic male patients with biopsy-proven CGN (age 30 +/- 7 years, body mass index 23.0 +/- 2.5 kg/m2) were enrolled. Insulin sensitivity was estimated by the glucose infusion rate (M value) during euglycemic hyperinsulinemic clamping for 60 to 120 min. Calcium-related parameters including intracellular calcium concentrations ([Ca2+]i) in platelets were also measured. Renal function was normal or slightly impaired (serum creatinine, 1.0 +/- 0.2 mg/dl; glomerular filtration rate (GFR), 68 to 131 ml/min/1.48 m2). We divided subjects into an insulin-sensitive (IS) group (M value > 7.3 mg/kg/min, the overall mean) and an insulin-resistant (IR) group (M value < 7.3 mg/kg/min). RESULTS: During a 75 g oral glucose tolerance test, the plasma glucose concentration at 120 min after glucose loading and the immunoreactive insulin concentration at 60 min were significantly higher in the IR group. GFR was notably lower in the IR group than in the IS group (p = 0.0003), and was significantly correlated with insulin sensitivity (p < 0.02, r = 0.58). The basal [Ca2+]i was significantly higher in the IR than in the IS group (39 +/- 9 vs. 30 +/- 9 nM, p < 0.05). CONCLUSION: Mild renal dysfunction and elevated basal [Ca2+]i are associated with insulin resistance in CGN. PMID- 11105796 TI - Blood pressure reduction associated with preservation of renal function in hypertensive patients with IgA nephropathy: a 3-year follow-up. AB - BACKGROUND: The relative importance of hypertension in the progression of renal failure is well understood. Recently, several studies have provided evidence that antihypertensive therapy enhances renal survival. However, the specific antihypertensive drug regimens that are most effective in generating such long term effects remain controversial. PATIENTS AND METHODS: Forty-nine hypertensive IgA nephropathy (IgAN) patients (39 +/- 7 years old, serum creatinine 1.1 +/- 0.2 mg/dl) with proteinuria received antihypertensive therapy with angiotensin converting enzyme inhibitors (ACEi: 2.5-10 mg of benazapril daily) and calcium channel blockers (CCBs: 2.5-10 mg amlodipine daily) for 3 years. The patients' blood pressures in group one were controlled below 140/85 mmHg by increases in their first drug dose or by addition of the second drug in group 1. Blood pressures for patients in group 2 were controlled using the same two options, except to levels below 130/70 mmHg. Patients within the two groups were selected and controlled with regard to sex, age, and serum creatinine. The renal protective effects of each protocol were evaluated in terms of reductions in creatinine clearance. After 3 years of the above outlined blood pressure control regimens, the reductions in creatinine clearance were compared. RESULTS: Creatinine clearances decreased in group 1 patients (from 85.7 +/- 2.4 ml/min to 72.9 +/- 2.4 ml/min, p < 0.05). On the other hand, creatinine clearance remained essentially unchanged for patients in group 2 (from 87.2 +/-4.7 ml/min to 85.9 +/ 5.9 ml/min). Although creatinine clearance in both groups was almost the same at the start of study, there was a significant difference between them by the conclusion of the study (p < 0.05). Proteinuria and hematuria did not change significantly throughout the study and there were no significant differences in these respects between these two corresponding groups. There were no significant differences between the groups with reference to side-effects or complications. CONCLUSION: These data provide evidence that reducing blood pressure has protective renal effects in cases of mild renal insufficiency with hypertension in IgA nephropathy. PMID- 11105798 TI - Comparison of cystatin C versus creatinine for detection of mild renal failure. AB - AIM: Serum cystatin C (SCyst) has been proposed as a novel indicator of GFR. PATIENTS AND METHODS: We compared SCyst, serum creatinine (SCreat) and Cockcroft and Gault's estimated clearance (CCG) using inulin clearance (Cin) as gold standard. 140 subjects (161 samples; aged 39 +/- 14; male/female: 79/82) underwent simultaneous measurements. RESULTS: A highly significant correlation r = 0.70, 0.74, 0.77 (p < 0.0001) was found between 1/SCyst, 1/SCreat, C(CG), respectively, and Cin. Receiver-operating characteristic (ROC) analysis was performed on SCyst, SCreat and C(CG) using a Cin cut-off of 90 ml/min/1.73 m2. Best fit for SCyst was 0.90 mg/l with a sensitivity of 75% and a specificity of 92%. The area under the ROC curve was not significantly greater for SCreat or C(CG) than for SCyst (p = 0.91,0.13, respectively). When relationship between Cin and SCyst was plotted, experimental data deviated from the theoretical model, suggesting that cystatin C may not be solely filtered. Additional patients were selected in our database on the basis of discordant SCreat/GFR values: false negative (n = 46 samples, 31 patients) and false positive (n = 16 samples, 9 patients). In this highly selected subgroup, 38% of the SCreat false positive had normal SCyst values and 48% of the false negative SCreat had abnormally elevated SCyst. CONCLUSION: This study suggests that SCyst is not more sensitive than SCreat or C(CG) for detecting renal failure, however, SCyst could be proposed as a confirmatory test for patients with elevated SCreat. PMID- 11105799 TI - The incidence of thrombovenous and thromboembolic complications in kidney transplant patients with recurrent glomerulonephritis is dependent on the occurrence of severe proteinuria. AB - BACKGROUND: Patients with recurrent glomerulonephritis (RG) after kidney transplantation are at high risk for thromboembolic events but it is unclear when the risk begins to increase. PATIENTS AND METHODS: We evaluated the risk for thrombovenous and thromboembolic complications in relation to the occurrence of severe proteinuria (> or = 2 g protein in 24-hour urine) in 15 renal allograft recipients with biopsy-proven RG, who had received 20 allografts RG. The total period of observation was 53 (10-91) months. The post-transplant period before the occurrence of severe proteinuria lasted 18 (1-34) months and the subsequent proteinuric period until the end of the study, 35 (9-85) months. RESULTS: The monthly incidence of thrombovenous and thromboembolic complications was only 1/18 in the first period before and in contrast, 11/35 in the subsequent period after the occurrence of severe proteinuria. The mean urinary protein excretion increased from 0.4 +/- 0.1 g/day immediately after transplantation to 6.1 +/- 4.8 g/day at the end of the study (p < 0.001). During the same period there was a 1.2 fold increase of fibrinogen (from 366 +/- 88 to 442 +/- 120 mg/dl, p < 0.025) and a 1.2-fold decrease of antithrombin III (from 110 +/- 12 to 92 +/- 12%, p < 0.001). All thrombotic complications occurred in 6 patients with 9 grafts; at the end of the study this group showed higher fibrinogen concentrations (454 +/- 155 versus 433 +/- 89 mg/dl, NS) m and lower antithrombin III levels (88 +/- 11 versus 97 +/- 11%, p < 0.05) than the group without thrombotic complications. CONCLUSION: In kidney transplant patients with RG a high risk for thrombovenous and thromboembolic complications can be obs- served after the occurrence of severe proteinuria; this can mainly be explained by high fibrinogen and low antithrombin III levels. Anticoagulation therapy should be started in patients with RG immediately after the occurrence of severe proteinuria. PMID- 11105800 TI - Comparison of patients hemodialyzed for lithium poisoning and those for whom dialysis was recommended by PCC but not done: what lesson can we learn? AB - AIMS: To compare patients for whom hemodialysis was done for lithium poisoning and those for whom it was recommended by the poison control centre (PCC) but not done and to evaluate the effect of withholding hemodialysis on outcomes. METHODS: All lithium overdoses brought to the attention of the PCC were prospectively followed from January 1 to December 31, 1996. Patients for whom hemodialysis was done were compared with those for whom it was recommended but not done in terms of clinical presentation, lithium elimination half-life, need for transfer to another centre for hemodialysis, and outcome (death, or sequel or recovery). RESULTS: A total of 205 cases of lithium overdoses were collected including 110 with levels higher than 1.5 mmol/l. There were 12 acute lithium overdoses; no patients required hemodialysis and there were no sequel or deaths. There were 174 acute on chronic overdoses; hemodialysis was recommended in 9 patients but only 6 underwent hemodialysis; one patient died during hemodialysis but no other had sequel. There were 19 chronic poisonings; hemodialysis was recommended in 9 patients but only 2 had hemodialysis, a third patient underwent hemodialysis despite it not being recommended; one patient died without hemodialysis and one other had sequel after hemodialysis. No difference were observed between the groups for age, sex, type of poisoning (acute on chronic/chronic), levels (initial/peak/6 hours/extrapolated at 30 hours), time of presentation post ingestion, presence of co-ingestants, symptoms and signs, Hansen and Amdisen grade, initial creatinine, time of recommendation to perform hemodialysis (daytime or nighttime), need to transfer patients to another centre to perform hemodialysis, and outcome. Patients with acute on chronic poisoning that were not hemodialyzed had longer elimination half-life than those for whom hemodialysis was done even before hemodialysis was performed: 50.1 +/- 13.6 h (n = 3) versus 12.9 +/-12.1 (n = 3) (p = 0.007), respectively. CONCLUSION: No difference was observed between patients for whom hemodialysis was done and those for whom it was recommended by PCC but not done. Despite the death of one patient clearly associated with voluntary withholding hemodialysis, sequel was not seen in that group. The indications for hemodialysis in lithium poisoning should be reconsidered to include only the more severe cases. PMID- 11105801 TI - Predictive value of access blood flow and stenosis in detection of graft failure. AB - AIMS: Low access flow and the diagnosis of high degrees of venous stenosis have been recommended as indications for prophylactic angioplasty. However, recent studies have shown that prophylactic angioplasty for > 50% stenosis did not prolong graft patency, and that a single flow measurement may not accurately predict graft failure. In this study we compared the value of monthly measurement of access flow and of the maximal degree of stenosis in the detection of graft failure over a three-month period. METHODS: Thirty-nine hemodialysis patients with polytetrafluoroethylene (PTFE) grafts were evaluated by Doppler ultrasound at monthly intervals for three months. Graft failures were defined as thrombosis, or surgical and angioplastic revisions required because of the presence of access recirculation, and patients with graft failure were followed within the subsequent one-month periods of observation. RESULTS: Twelve graft failures occurred during the three-month period of observation. The risk for subsequent graft failure significantly increased at flows < 300 ml/min. Nine (20%) graft failures occurred with stenoses of 30 to 50%, and three (13%) with stenoses of> 50%. The grafts that failed in the second and the third study months had a 25.8% (380 +/- 62 vs. 287 +/- 190 ml/min, p < 0.05) and a 36.5% (393 +/- 142 vs. 226 +/ 41 ml/min, p < 0.05) decrease in access flow, respectively. There was no significant change in access flow for the grafts patent throughout the study (911 +/- 333, 794 +/- 302, and 919 +/383 ml/min, p = ns). No significant increases in maximal stenosis were found for the grafts that failed in the second month (44 +/ 6.1 vs. 48 +/- 15%, p = ns) and the third month (48 +/- 9 vs. 51 +/- 16%, p = ns). There were no significant changes in the maximum stenosis for the grafts patent throughout the three-month study periods (37 +/- 15,43 +/- 11, and 44 +/- 15%, p = ns). CONCLUSIONS: Access flow is a more sensitive predictor of graft failure than stenosis. Examination of trend in decline of access flow is a more powerful indicator to detect graft dysfunction than an individual single flow value. PMID- 11105802 TI - Serum creatinine can predict adequacy of peritoneal dialysis--preliminary report. AB - AIM: To determine the value of creatinine clearance, estimated using the Cockcroft and Gault formula, in assessing adequacy of peritoneal dialysis. METHODS: We undertook a retrospective analysis of creatinine clearance results derived from a conventional 24-hour collection in 35 stable outpatients on peritoneal dialysis and compared them with those calculated from the corresponding serum creatinine using the Cockcroft and Gault formula. RESULTS: There was a strong correlation between the 2 methods (r = 0.82, p < 0.0001), although the formula tended to over-estimate clearances. The formula had a positive predictive value of 88% and a negative predictive value of 86% for detecting inadequate dialysis. CONCLUSION: The creatinine clearance calculated using the Cockcroft and Gault formula can be used in patients on peritoneal dialysis to estimate adequacy of dialysis. We believe that this method, which is far less expensive and time-consuming, deserves further testing in a larger population in order to define more accurately its role in the management of PD patients. PMID- 11105803 TI - Nephrotic syndrome due to focal glomerulosclerosis and undifferentiated carcinoma. AB - In neoplastic disorder-related nephrotic syndrome, focal glomerulosclerosis (FGS) has been reported mainly in hematological disorders like minimal change nephrotic syndrome (MCNS) in association with presumed T lymphocyte dysfunction. The association of FGS with cancer or solid tumor is rare. We report a case of nephrotic syndrome due to FGS in a patient with undifferentiated adenocarcinoma of the cystic duct. Although the underlying mechanism is unclear, the development of FGS seemed to be related to the poor histological differentiation of the cancer in the possibility of production of an active peptide. PMID- 11105804 TI - Crescentic glomerulonephritis complicating the course of a hypocomplementemic urticarial vasculitis. AB - The authors report a case of crescentic glomerulonephritis leading to end-stage renal failure in a patient affected by a hypocomplementemic urticarial vasculitis (HUV). The patient's clinical course was characterized by the survey of several episodes of extra-renal vasculitis. We emphasize that HUV commonly considered as a limited form of vasculitis might in some cases have a dramatic evolution. PMID- 11105805 TI - Different pathological findings in each of four parathyroid glands in a long standing hemodialysis patient. AB - A 51-year-old male patient with chronic renal failure, who had required dialysis for 22 years, presented with a cervical mass. Laboratory data were consistent with secondary hyperparathyroidism due to chronic renal failure. Cervical exploration was performed with excision of four parathyroid glands and autotransplantation of the normal gland into the forearm. The cervical tumor of the right inferior gland demonstrated parathyroid carcinoma histologically. Adenoma of the right superior gland and hyperplasia of the left superior gland were also recognized. The left inferior gland was normal. A few cases of parathyroid carcinoma in patients on maintenance hemodialysis have been previously reported. However, this is the first report in which all four parathyroid glands revealed different pathological findings: carcinoma, adenoma, hyperplasia and normal gland. Chronic stimulation of the parathyroid glands to release parathyroid hormone might have caused the variety of findings in the four parathyroid glands. PMID- 11105806 TI - Fatal cytomegalovirus pneumonia after preemptive antiviral therapy in a renal transplant recipient. AB - Cytomegalovirus (CMV) infections occur with an incidence of up to 70% in renal transplant patients and mortality is low due to effective antiviral drugs. We report here the case of a patient who suffered from an uncommonly severe and therapy-resistant pulmonary CMV infection. During the disease course, CMV-PCR from alveolar cells and lung biopsy material was repeatedly negative despite high CMV pp65 antigenemia. CMV pneumonia was finally diagnosed from a biopsy obtained by open thoracotomy revealing positive CMV immunostaining of lung tissue. The patient died of respiratory failure though double-treatment using both ganciclovir and foscavir was administered. Post mortem, the clinically suspected resistance to both antiviral drugs, but not to cidofovir, could be proven by bioassay testing of in vitro growth responses using viral cultures. CMV pneumonia may thus not be diagnosed by standard PCR techniques in rare cases and may be resistant to the available antiviral therapy. Severe CMV pneumonia may benefit from novel antiviral drugs such as cidofovir, which is currently used in the treatment of CMV retinitis in HIV patients. PMID- 11105807 TI - Beta-interferon-induced nephrotic syndrome in a patient with multiple sclerosis. PMID- 11105808 TI - A comparative study of losartan and enalapril on erythropoiesis and renal function in hypertensive patients with renal parenchymal disease. PMID- 11105809 TI - Late onset oncogenic osteomalacia-associated with neurofibromatosis type II. PMID- 11105810 TI - Hereditary distal renal tubular acidosis associated with deafness and cataract. PMID- 11105811 TI - RPA's strategy to enhance and improve continuous quality improvement in the ESRD Program. Part 1 of 2. Renal Physicians Association. PMID- 11105812 TI - Barriers to HCQIP as the vehicle for national CQI in the ESRD. PMID- 11105813 TI - [123I]VIP receptor scintigraphy in patients with pancreatic adenocarcinomas. PMID- 11105814 TI - Does thyroid stunning exist? A model with benign thyroid disease. AB - With regard to the treatment of differentiated non-medullary thyroid carcinoma, there is controversy over whether radiation from a diagnostic radioiodine (131I) application really does have a suppressive effect on the uptake of subsequent therapeutic 131I (so-called thyroid stunning). However, inherent difficulties in exact remnant/metastatic tissue volumetry make it difficult to quantify how much diagnostic 131I is actually absorbed (absorbed energy dose) and hence to decide whether a threshold absorbed dose exists beyond which such stunning would occur. Since in benign thyroid disease the target volume can be readily quantified by ultrasonography, we sought to determine definitely whether stunning of thyroid cells occurs upon a second application of radioiodine 4 days following the first one. We therefore studied 171 consecutive patients with benign thyroid disease (diffuse goitre, Graves' disease, toxic nodular goitre) who received two-step 131I therapy during a single in-patient stay. For application of both calculated 131I activities we performed kinetic dosimetry of 131I uptake, effective half life and absorbed dose. At the second application, patients showed significant stunning (a 31.7% decrease in 131I uptake, from 34.7% +/- 15.4% at first application to 23.7% +/- 12.3% at second application, P < 0.0005) without a significant difference in effective half-life (4.9 +/- 1.3 vs 5.0 +/- 1.7 days, P > 0.2). ANOVA showed that the extent of stunning was influenced significantly only by the absorbed energy dose at first application (F = 13.5, P < 0.0005), while first-application 131I activity, target volume, gender and thyroid function had no influence (all F < or = 0.71, all P > 0.4). There was no significant correlation between extent of thyroid stunning and first-application 131I activity ( r = 0.07, P > 0.3), whereas there was a highly significant correlation between thyroid stunning and first absorbed energy dose (r = 0.64, P < 0.00005), the latter correlation fitting a logarithmic model best. Multivariate factor analysis also revealed first absorbed energy dose to be the only decisive stunning factor. In conclusion, our study confirms that stunning exists in benign thyroid conditions and that it is a purely radiobiological inhibitory phenomenon related to absorbed dose. PMID- 11105815 TI - Cost-effectiveness of FDG-PET for the management of potentially operable non small cell lung cancer: priority for a PET-based strategy after nodal-negative CT results. AB - Decision analysis is used here to establish the most cost-effective strategy for management of potentially operable non-small cell lung cancers (NSCLCs). The strategies compared were conventional staging (strategy A), dedicated systems of positron emission tomography (PET) using fluorine-18 fluorodeoxyglucose (FDG) in patients with normal-sized (strategy B) or in patients with enlarged mediastinal lymph nodes (part of strategy C), and FDG-PET followed by exclusion from surgical procedures when both computed tomography (CT) and PET were positive for mediastinal lymph nodes (strategy D) or when PET alone was positive (strategy E). Based on published data, the sensitivity and specificity of FDG-PET were estimated at 0.74 and 0.96 for detecting metastasis in normal-sized mediastinal lymph nodes, and at 0.95 and 0.76 when these lymph nodes were enlarged. The calculated probability of up-staging to M1 by using PET was 0.05. The costs quoted correspond to the cost reimbursed in 1999 by the public health provider in Germany. The incremental cost-effectiveness ratio (ICER) of strategy B was much more favourable (143 EUR/LYS; LYS = life year saved) than the ICER of strategy C (36,667 EUR/LYS). In strategy B, the use of PET did not raise the overall costs because the costs of PET were almost balanced by a better selection of patients for beneficial cancer resection. The exclusion from biopsy confirmation in strategies D and E led to cost savings that did not justify the expected reduction in life expectancy. In sensitivity analyses, the ICERs of strategy B were robust to the pretest likelihood of N2/N3, to penalized test parameters of PET and to reimbursement of PET. However, the ICER of strategy B would be raised to 28,000 EUR/LYS through use of thoracic PET without whole-body scanning. To conclude, the implementation of whole-body PET with a full ring of detectors in the preoperative staging of patients with NSCLC and normal-sized lymph nodes is clearly cost-effective. However, patients with nodal-positive PET results should not be excluded from biopsy. PMID- 11105816 TI - Reliability of DMSA for the diagnosis of renal parenchymal abnormality in children. AB - The objective of this study was to evaluate the variability of technetium-99m dimercaptosuccinic acid (DMSA) scintigraphy interpretation by four nuclear medicine physicians for the diagnosis of renal parenchymal abnormality in children, and to compare variability among three different DMSA methods in clinical use: planar alone, single-photon emission tomography (SPET) alone, and planar with SPET. One hundred consecutive DMSA studies were independently interpreted 3 times by four participating nuclear medicine specialists from different departments and in random order. All scans were classified by the presence or absence of renal parenchymal abnormality using the modified four level grading system of Goldraich. Indices of agreement were the percentage of agreement and the kappa statistic. Disagreement was analysed using children, kidneys and kidney zones (three zones per kidney). Using patients as the unit of analysis, agreement for planar and planar with SPET methods was 87%-88% (kappa 0.74) for the normal-abnormal scan classification. The corresponding agreement value for the SPET alone method was 78% (kappa 0.56). Similarly, substantial disagreement (disagreement > or = 2 categories) occurred in 2.5% and 1.3% of comparisons between observers for planar alone and planar with SPET, respectively, but in 5.2% of comparisons for SPET alone. These results did not vary appreciably whether interpretation of patients, kidneys or kidney zones was compared. It is concluded that the four experienced nuclear medicine physicians showed substantial agreement in the interpretation of planar alone and planar with SPET DMSA scintigraphic images. Interpretation of SPET DMSA images, without planar images, was significantly more variable than interpretation using the two other methods, disagreement occurring in more than 20% of comparisons. SPET DMSA scintigraphy, when used without planar images, does not provide a firm basis for clinical decision making in the care of children who may have renal damage. There is no apparent benefit of reduced variability from the extra provision of SPET data to nuclear medicine physicians who already have planar images. PMID- 11105817 TI - Fever of unknown origin: prospective comparison of [18F]FDG imaging with a double head coincidence camera and gallium-67 citrate SPET. AB - Gallium-67 citrate is currently considered as the tracer of first choice in the diagnostic workup of fever of unknown origin (FUO). Fluorine-18 2'-deoxy-2-fluoro D-glucose (FDG) has been shown to accumulate in malignant tumours but also in inflammatory processes. The aim of this study was to prospectively evaluate FDG imaging with a double-head coincidence camera (DHCC) in patients with FUO in comparison with planar and single-photon emission tomography (SPET) 67Ga citrate scanning. Twenty FUO patients underwent FDG imaging with a DHCC which included transaxial and longitudinal whole-body tomography. In 18 of these subjects, 67Ga citrate whole-body and SPET imaging was performed. The 67Ga citrate and FDG images were interpreted by two investigators, both blinded to the results of other diagnostic modalities. Forty percent (8/20) of the patients had infection, 25% (5/20) had auto-immune diseases, 10% (2/20) had neoplasms and 15% (3/20) had other diseases. Fever remained unexplained in 10% (2/20) of the patients. Of the 20 patients studied, FDG imaging was positive and essentially contributed to the final diagnosis in 11 (55%). The sensitivity of transaxial FDG tomography in detecting the focus of fever was 84% and the specificity, 86%. Positive and negative predictive values were 92% and 75%, respectively. If the analysis was restricted to the 18 patients who were investigated both with 67Ga citrate and FDG, sensitivity was 81% and specificity, 86%. Positive and negative predictive values were 90% and 75%, respectively. The diagnostic accuracy of whole-body FDG tomography (again restricted to the aforementioned 18 patients) was lower (sensitivity, 36%; specificity, 86%; positive and negative predictive values, 80% and 46%, respectively). 67Ga citrate SPET yielded a sensitivity of 67% in detecting the focus of fever and a specificity of 78%. Positive and negative predictive values were 75% and 70%, respectively. A low sensitivity (45%), but combined with a high specificity (100%), was found in planar 67Ga imaging. Positive and negative predictive values were 100% and 54%, respectively. It is concluded that in the context of FUO, transaxial FDG tomography performed with a DHCC is superior to 67Ga citrate SPET. This seems to be the consequence of superior tracer kinetics of FDG compared with those of 67Ga citrate and of a better spatial resolution of a DHCC system compared with SPET imaging. In patients with FUO, FDG imaging with either dedicated PET or DHCC should be considered the procedure of choice. PMID- 11105818 TI - Semi-quantitative evaluation of gallium-67 scintigraphy in lupus nephritis. AB - Within nuclear medicine there is a trend towards quantitative analysis. Gallium renal scan has been reported to be useful in monitoring the disease activity of lupus nephritis. However, only visual interpretation using a four-grade scale has been performed in previous studies, and this method is not sensitive enough for follow-up. In this study, we developed a semi-quantitative method for gallium renal scintigraphy to find a potential parameter for the evaluation of lupus nephritis. Forty-eight patients with lupus nephritis underwent renal biopsy to determine World Health Organization classification, activity index (AI) and chronicity index (CI). A delayed 48-h gallium scan was also performed and interpreted by visual and semi-quantitative methods. For semi-quantitative analysis of the gallium uptake in both kidneys, regions of interest (ROIs) were drawn over both kidneys, the right forearm and the adjacent spine. The uptake ratios between these ROIs were calculated and expressed as the "kidney/spine ratio (K/S ratio)" or the "kidney/arm ratio (K/A ratio)". Spearman's rank correlation test and Mann-Whitney U test were used for statistical analysis. Our data showed a good correlation between the semi-quantitative gallium scan and the results of visual interpretation. K/S ratios showed a better correlation with AI than did K/A ratios. Furthermore, the left K/S ratio displayed a better correlation with AI than did the right K/S ratio. In contrast, CI did not correlate well with the results of semi-quantitative gallium scan. In conclusion, semi-quantitative gallium renal scan is easy to perform and shows a good correlation with the results of visual interpretation and renal biopsy. The left K/S ratio from semi-quantitative renal gallium scintigraphy displays the best correlation with AI and is a useful parameter in evaluating the disease activity in lupus nephritis. PMID- 11105819 TI - A comparison of the overall first-pass kinetics of thallium-201 and technetium 99m MIBI in normoxic and low-flow ischaemic myocardium. AB - The specific impact of ischaemia on the myocardial kinetics of thallium-201 and technetium-99m 2-methoxy-2-isobutylisonitrile (MIBI) remains a matter of debate. Using an isolated heart model perfused with red blood cell-enhanced perfusate, we compared the overall first-pass kinetics of 201Tl and MIBI under haemodynamically stable conditions of low-flow ischaemia (> 50% reduction in normal coronary flow and a > or = 20 mmHg fall in systolic contraction pressure, n = 10) and normoxia (n = 11). For both 201Tl and MIBI, we found that under ischaemic conditions (as compared with normoxia) there was a higher initial net extraction fraction (201Tl: 0.78 +/- 0.03 vs 0.72 +/- 0.06, P = 0.006; MIBI: 0.49 +/- 0.10 vs 0.39 +/ 0.11, P = 0.03), a lower clearance rate in the 30 min following extraction (% decrease in cardiac uptake: 201Tl: 30 +/- 12 vs 47 +/- 14, P = 0.02; MIBI: 5 +/- 5 vs 13 +/- 11, P = 0.02) and a higher retention fraction at 30 min (20lTl: 0.54 +/- 0.10 vs 0.39 +/- 0.12, P = 0.004; MIBI: 0.46 +/- 0.08 vs 0.33 +/- 0.12, P = 0.01). Multivariate analyses, however, revealed that all myocardial kinetic parameters of both tracers were dependent only on coronary flow rates, without any additional significant impact of the presence of ischaemia or states of contractility or oxidative metabolism. We conclude that the myocardial fractional retention of both 201Tl and MIBI is strongly correlated with the decrease in coronary flow during ischaemia. This inverse relationship with coronary flow derives from both the flow-dependent increase in the initial myocardial extraction and the decrease in the subsequent myocardial washout of the tracers. PMID- 11105820 TI - Cardiac sympathetic dysfunction contributes to left ventricular remodeling after acute myocardial infarction. AB - To investigate the role of the cardiac sympathetic nervous system in left ventricular remodelling, 50 patients with first-time acute myocardial infarction (AMI) and patency of the infarct-related artery after reperfusion underwent quantitative iodine-123 metaiodobenzylguanidine (MIBG) imaging at 4 days and 4 weeks (n=42), and quantitative technetium-99m tetrofosmin imaging at 2 days after AMI. They also underwent both ventriculography and coronary angiography on admission and about 4 weeks after AMI. On the basis of left ventricular end systolic volume (LVESV), patients were divided into two groups. Patients with LVESV dilatation (n=20) had a significantly lower ejection fraction (P<0.003) and a significantly higher severity score of 99mTc-tetrofosmin (P<0.04), and total severity (P<0.01), delta extent (P<0.007) and delta severity (P<0.0008) scores of MIBG than patients without LVESV dilatation (n=30). delta severity score of MIBG was directly correlated with change in LVESV at 4 weeks (r=0.63, P<0.0001). Stepwise linear discriminant function analysis showed that delta severity score of MIBG (P<0.0002) was the only discriminator of LVESV dilatation. Patients with LVESV dilatation had higher regional washout rates in both the infarct and the non-infarct zones than patients without such dilatation. Furthermore, no MIBG parameters changed significantly between 4 days and 4 weeks after AMI. In reperfused AMI, delta severity score of MIBG was related to the degree of ventricular dilatation and was the only powerful discriminator of ventricular dilatation. These results suggest that cardiac sympathetic nervous abnormality might contribute to left ventricular remodelling in reperfused AMI. MIBG imaging may allow identification of reperfused AMI patients at high risk for left ventricular remodelling. PMID- 11105821 TI - Non-invasive assessment of the effect of cardiac sympathetic innervation on metabolism of the human heart. AB - The role of cardiac sympathetic nerves in the regulation of myocardial metabolism is not well defined. Owing to the presence of incomplete reinnervation, heart transplant recipients provide a unique model to study the effects of efferent sympathetic innervation. Using this model, we sought to determine the influence of cardiac sympathetic signals on substrate utilisation and overall oxidative metabolism. In 21 transplant recipients, positron emission tomography was applied to determine sympathetic innervation with the noradrenaline analogue carbon11 hydroxyephedrine, oxidative metabolism with carbon11 acetate (n=14), and glucose utilisation with fluorine-18 fluorodeoxyglucose (n=7). The reinnervated area comprised 22% +/- 20% of the left ventricle. Oxidative metabolism was similar in denervated and reinnervated myocardium [0.06 +/- 0.01 vs 0.06 +/- 0.01/min for k(mono)], while glucose uptake was significantly higher in denervated myocardium (6.9 +/- 6.6 vs 6.0 +/- 6.2 micromol/min/100 g; P=0.03). Reinnervation mainly occurred in the territory of the left anterior descending artery, where retention of 11C-hydroxyephedrine (6.8 +/- 2.7%/min) was higher compared with territories of the left circumflex (4.1 +/- 1.7%/min; P<0.01) and right coronary (3.8 +/- 1.1%/min; P<0.01) arteries. Oxidative metabolism was similar in all three territories, but compared with the reinnervated territory of the left anterior descending artery (53% +/- 16% of maximum), relative FDG uptake was higher in territories of the left circumflex (76% +/- 6%, P<0.01) and right coronary (67% +/- 10%, P<0.05) arteries. Similar degrees of regional heterogeneity were not observed in normals. Thus, while overall energy production through oxidative metabolism remains unaffected, cardiac utilisation of glucose in the fasting state is increased in the absence of catecholamine uptake sites. Innervated myocardium, however, may preferentially utilise free fatty acids, suggesting a role for sympathetic tone in substrate utilisation. PMID- 11105822 TI - Evaluation of ictal brain SPET using statistical parametric mapping in temporal lobe epilepsy. AB - An automated voxel-based analysis of brain images using statistical parametric mapping (SPM) is accepted as a standard approach in the analysis of activation studies in positron emission tomography and functional magnetic resonance imaging. This study aimed to investigate whether or not SPM would increase the diagnostic yield of ictal brain single-photon emission tomography (SPET) in temporal lobe epilepsy (TLE). Twenty-one patients (age 27.14 +/- 5.79 years) with temporal lobe epilepsy (right in 8, left in 13) who had a successful seizure outcome after surgery and nine normal subjects were included in the study. The data of ictal and interictal brain SPET of the patients and baseline SPET of the normal control group were analysed using SPM96 software. The t statistic SPM?t? was transformed to SPM?Z? with various thresholds of P<0.05, 0.005 and 0.001, and corrected extent threshold P value of 0.05. The SPM data were compared with the conventional ictal and interictal subtraction method. On group comparison, ictal SPET showed increased uptake within the epileptogenic mesial temporal lobe. On single case analysis, ictal SPET images correctly lateralized the epileptogenic temporal lobe in 18 cases, falsely lateralized it in one and failed to lateralize it in two as compared with the mean image of the normal group at a significance level of P<0.05. Comparing the individual ictal images with the corresponding interictal group, 15 patients were correctly lateralized, one was falsely lateralized and four were not lateralized. At significance levels of P<0.005 and P<0.001, correct lateralization of the epileptogenic temporal lobe was achieved in 15 and 13 patients, respectively, as compared with the normal group. On the other hand, when comparison was made with the corresponding interictal group, only 7 out of 21 patients were correctly lateralized at the threshold of P<0.005 and five at P<0.001. The result of the subtraction method was close to the single case analysis on SPM at P<0.05. However, at higher thresholds (P<0.005 and 0.001) the subtraction method was comparable to the SPM results only when individual ictal images were compared with the normal control group, and not when comparison was with the interictal group. It is concluded that SPM is an alternative diagnostic method for the localization or lateralization of the seizure focus in temporal lobe epilepsy and that interictal SPET could be omitted if a normal brain SPET database were to be established. The medical cost of seizure localization would thereby be reduced. PMID- 11105823 TI - Measurement of extrastriatal D2-like receptor binding with [11C]FLB 457--a test retest analysis. AB - [11C]FLB 457 is a radioligand for positron emission tomography (PET) that possesses high affinity to D2/D3 receptors. It has been suggested to be useful for quantification of low-density dopamine D2 receptor populations, e.g. in cortical and limbic brain areas. We explored the reproducibility of five methods for measuring extrastriatal D2-like receptor binding potential with [11C]FLB 457. Seven healthy male volunteers were examined twice with [11C]FLB 457 (high specific radioactivity) on the same day, at least 3 h apart. Four brain areas, frontal cortex, nucleus thalamus, temporal cortex and cerebellar cortex, were examined. Binding potentials (BPs) were derived from (1) a target to cerebellum distribution volume ratio, (2/3) two reversible reference tissue compartment models and (4) a transient equilibrium approach. For comparison, BP values were also calculated with the standard three-compartment kinetic model that does not assume a receptor-free reference region. The use of the standard three compartment model did not result in reproducible BP estimates. The distribution volume (DV) ratio, reference tissue compartment models and the transient equilibrium method all had good to excellent intraclass correlation coefficients (ICCs) in the studied brain areas ranging from 0.56 to 0.93. Absolute variability was also relatively low, ranging from 5.3% to 10.4%. There were no marked differences in the ICC or absolute and relative variability between the four methods based on a reference tissue (cerebellum). In addition, we did not observe systematic differences in the BP between the first and the second scan. These data indicate that the reproducibility of the DV ratio, reference tissue models and the transient equilibrium method is good or excellent. However, each of these methods includes assumptions affecting their validity. Thus, the choice of method will be critically dependent on the purpose of the study. PMID- 11105824 TI - [11C]flumazenil metabolite measurement in plasma is not necessary for accurate brain benzodiazepine receptor quantification. AB - In this work, a mathematical correction for metabolites has been validated which estimates the relative amount of [11C]flumazenil ([11C]FMZ) in the total plasma curve from the tissue kinetic data without the need for direct metabolite measurement in blood plasma samples. Kinetic data were obtained using a 90-min three-injection protocol on five normal volunteers. First, the relative amount of [11C]FMZ in plasma was modelled by a two-parameter exponential function. The parameters were estimated either directly by fitting this model to the blood plasma metabolite measurements, or indirectly from the simultaneous fitting of tissue time activity curves from several brain regions with a non-linear FMZ kinetic model. Second, the direct and indirect metabolite corrections were fixed and the FMZ compartmental parameters were determined on a regional basis in the brain. The validation was performed by comparing the regional values of benzodiazepine receptor density Bmax and equilibrium dissociation constant Kd obtained with the direct metabolite correction with those values obtained with the indirect correction. For Bmax, the correlation coefficient r2 was above 0.97 for all subjects and the slope values of the linear regression were within the interval [0.97, 1.2]. For Kd, r2 was above 0.96, and the slope values of the linear regression were within the interval [0.99, 1.1]. Simulation studies were performed in order to evaluate whether this metabolite correction method could be used in a clinical protocol where only a single [11C]FMZ injection and a linear compartmental model are used. The resulting [11C]FMZ distribution volume estimates were found to be linearly correlated with the true values, with r2=1.0 and a slope value of 1.1. The mathematical metabolite correction proved to be a feasible and reliable method to estimate the relative amount of [11C]FMZ in plasma and the compartmental model parameters for three-injection protocols. Although validation with real data is necessary, simulation results suggest that our analysis method may also be applied to single-injection protocols. PMID- 11105825 TI - Vasoactive intestinal peptide receptor scintigraphy in patients with pancreatic adenocarcinomas or neuroendocrine tumours. AB - Human adenocarcinomas of the gastroenteropancreatic system overexpress vasoactive intestinal peptide (VIP) receptors and therefore represent logical diagnostic targets for receptor scintigraphy. Using iodine-123 labelled VIP, the newly employed diagnostic procedure termed VIP receptor scintigraphy (VIP-RS) appears to detect tumour tissue, especially pancreatic metastatic tumours, in almost all cases. So far, however, only a single centre has demonstrated convincing positive results. The aim of this study was to compare the sensitivity and specificity of VIP-RS with those of computer tomography (CT) and transabdominal ultrasound in patients with extensive pancreatic metastatic adenocarcinomas and neuroendocrine tumours. VIP was radiolabelled with carrier-free 123I using the chloramine T method and preparative high-performance liquid chromatography for purification. Patients with metastatic pancreatic (n=12) and colorectal (n=3) carcinomas (adenocarcinoma: n=13, neuroendocrine tumour: n=2) were studied by VIP-RS, CT, ultrasound and, in one case, also by radioligand receptor autoradiography. Carrier-free radioiodinated VIP of maximum specific radioactivity maintained a high biological activity as determined by cAMP formation in receptor-expressing tumour cell lines. Intravenous injection of 123I-VIP did not cause any side effects. Biodistribution, determined over 24 h, was high in the lungs and low in abdominal organs. Although all patients had extensive metastatic disease as evidenced by CT and ultrasound, VIP-RS was unable to detect either primaries or metastases in these patients. Only in two patients could a significant uptake of radiolabel be detected in organs directly infiltrated by the primary. To exclude false-negative findings, tumour tissue in one patient with a large primary, undetectable by VIP-RS, was analysed by radioligand receptor autoradiography and shown to be receptor positive. Moreover, in vitro receptor determinations showed that pancreatic carcinomas usually have fewer VIP receptors than the normal tissues to which they metastasize, like the liver. It is concluded that VIP can be radioactively labelled with maximum specific radioactivity while maintaining biological activity. Intravenous administration leads to a biodistribution almost identical to that reported previously. However, in contrast to these reports, very low sensitivity and specificity were observed for the detection of pancreatic cancers. In retrospect, these findings are not surprising since VIP receptor expression was observed to be higher in normal tissues than in neoplastic ones. PMID- 11105826 TI - Technetium-99m RP527, a GRP analogue for visualisation of GRP receptor-expressing malignancies: a feasibility study. AB - Gastrin-releasing peptide (GRP) receptor scintigraphy could allow prediction of response to GRP receptor-targeted treatment options, early non-invasive diagnosis and in vivo prognostic stratification of GRP receptor-positive tumours. This study reports on the imaging characteristics and efficacy for tumour detection of technetium-99m RP527, a 99mTc chelated targeting peptide derived from bombesin, which binds GRP receptors with high affinity. Ten patients (four men and six women, mean age 56.4 years) either suffering from metastasised prostate (n, number of patients = 4) or breast carcinoma (n=1) or presenting with a clinical diagnosis highly suggestive for breast carcinoma (n=5) were included in the study. In the latter five patients, 99mTc-RP527 scintigraphy was performed prior to diagnostic, e.g. biopsy, and staging examinations. Final diagnosis in these patients was breast carcinoma in all five. In all patients, whole-body planar scans and tomographic images were acquired 1 h and 5-6 h post injection of 555 MBq 99mTc-RP527 and tumour to normal tissue (T/N) ratios determined. 99mTc-RP527 showed specific uptake in four of six breast and one of four prostate carcinomas. T/N ratios derived from planar and tomographic images increased significantly (P<0.01) from 1.65 (SD 1.53) and 3.35 (SD 3.04) to 2.58 (SD 1.26) and 7.23 (SD 8.46), respectively. T/N ratios derived from tomographic images were consistently higher (P<0.01). The data presented suggest that 99mTc-RP527 results in specific tumour localisation and exhibits good imaging characteristics with a good T/N ratio that may be further enhanced by single-photon emission tomography. PMID- 11105827 TI - Influence of different chelators (HYNIC, MAG3 and DTPA) on tumor cell accumulation and mouse biodistribution of technetium-99m labeled to antisense DNA. AB - We have shown recently that cell accumulation in culture of antisense DNA is strongly influenced by the presence of a 99mTc-MAG3 group for radiolabeling. We have now compared the in vitro and mouse in vivo behavior of 99mTc when radiolabeled to one antisense phosphorothioate DNA by three different methods. The 18-mer antisense DNA against the RIalpha subunit of PKA was conjugated via a primary amine on the 5'-end with the NHS esters of HYNIC and MAG3 and by the cyclic anhydride of DTPA. Surface plasmon resonance measurements revealed that the association rate constant for hybridization was unchanged for all three chelators as compared with that of the native DNA. Size exclusion HPLC showed rapid and quantitative protein binding for all three chelators upon incubation of labeled DNAs in 37 degrees C serum and cell culture medium. However, in each case, radiolabeled and intact oligonucleotide was still detectable after 24 h. Cellular uptake was tested in an RIalpha mRNA-positive cancer cell line. The order of cellular accumulation of 99mTc was DTPA>HYNIC(tricine) >MAG3, with the differences increasing with time between 4 and 24 h. The rate of 99mTc egress from cells was found to be MAG3>HYNIC>DTPA, which may explain the order of cellular accumulation. The biodistribution in normal mice was heavily influenced by the labeling method and followed a pattern similar to that seen previously by us for peptides labeled with the same chelators. In conclusion, although these studies concerned only one antisense DNA in one cell line, the results suggest that the success of antisense imaging may depend, in part, on the method of radiolabeling. PMID- 11105828 TI - "FUR"--one size suits all. AB - This work used amalgamated data from previous projects in order to test the concept that when organ function is expressed in terms of tracer kinetics, the results are independent of patient size or gender. Dynamic gamma camera studies were analysed by measuring the rate of movement of tracers from the blood into various organs. These rates were expressed as a "fractional uptake rate" (FUR), which is the fraction of tracer in the blood taken up by the organ per unit time. As these values were small, it was convenient to express the FUR per million seconds. The FUR was calculated using the expression FUR = SLOPE (of Rutland Patlak plot), multiplied by B(0) (the blood curve value back-extrapolated to time zero), and divided by the TOTAL amount of tracer injected. Data were used from adult patients between the ages of 20 and 49 years who had normal organ function. Organ/tracer groups studied were the skeletal uptake of 99mTc-MDP, the renal uptake of 99mTc-MAG3, the renal uptake of 99mTc-MDP, the renal uptake of 99mTc DTPA, the hepatic uptake of 99mTc-colloid, the splenic uptake of 99mTc-colloid, and the hepatic uptake of 99mTc-DISIDA. Each organ/tracer group was divided into three subgroups according to patient size (smallest, middle and largest), and also into subgroups according to gender. Comparison of these subgroups did not show any significant size- or gender-related differences in FUR values. It is concluded that for patients with normally functioning organs the FUR is independent of patient size or gender. Thus, the FUR is a valuable way of expressing organ function, particularly in patients with unusual or rapidly changing body size, such as children. PMID- 11105829 TI - Simplified quantification of dopamine transporters in humans using [99mTc]TRODAT 1 and single-photon emission tomography. AB - Quantification of dopamine transporters (DAT) using [99mTc]TRODAT-1 and single photon emission tomography (SPET) requires full kinetic modeling of the data, using complex and invasive arterial blood sampling to provide an input function to the model. We have shown previously that a simpler reference tissue model provides accurate quantitative results, using a reference region devoid of DAT as the input to the model and thereby obviating the need for blood sampling. We now extend this work into humans, and develop further simplifications to make the imaging protocol much more practical as a routine procedure. Fourteen healthy subjects (age 29.8 +/- 8.4 years, range 18.7-45.5 years) underwent dynamic SPET for 6 h following injection of 752 +/- 28 MBq [99mTc]TRODAT-1. The kinetic data were analyzed using nonlinear regression analysis (NLRA) and Logan-Patlak graphical analysis. In addition, simple average ratios of striatal-to-background counts were obtained for three 1-h periods (3-4 h, 4-5 h, 5-6 h), and compared against the kinetic models. All methods gave an index of specific binding, proportional to the binding potential, known as the distribution volume ratio (DVR). The reference tissue NLRA gave mean values of k3=0.013 +/- 0.003 min(-1), k4=0.011 +/- 0.002 min(-1), and DVR=2.29 +/- 0.17. Graphical analysis gave a value of DVR=2.28 +/- 0. 16, and the three ratio values of DVR were: 3-4 h, 2.18 +/- 0.15; 4-5 h, 2.34 +/- 0.13; and 5-6 h, 2.46 +/- 0.19. Graphical analysis was highly correlated with NLRA (R2=0.91, slope=0.90 +/- 0.08). The ratio methods correlated well with NLRA (3-4 h, R2=0.71, slope= 0.73 +/- 0.13; 4-5 h, R2=0.86, slope=0.73 +/- 0.09; 5-6 h, R2=0.80, slope=1.00 +/- 0.15), and also with graphical analysis (3-4 h, R2=0.65, slope=0.74 +/- 0.16; 4-5 h, R2=0.85, slope=0.78 +/- 0.09; 5-6 h, R2=0.88, slope=1.11 +/- 0.12). The optimum equilibrium time point for obtaining a simple ratio was approximately 4.5-5.5 h. In conclusion, the simple ratio techniques for obtaining a quantitative measure of specific binding correlated well with the reference tissue kinetic models, using both NLRA and graphical analysis. The optimum time for obtaining a ratio appeared to be in the range 4.5-5.5 h. Earlier time points, while still relatively accurate, had a lower sensitivity and may not be optimized for measuring small changes in DAT concentrations. PMID- 11105830 TI - Imaging the serotonin transporter with positron emission tomography: initial human studies with [11C]DAPP and [11C]DASB. AB - Two novel radioligands, N,N-dimethyl-2-(2-amino-4-methoxyphenylthio) b enzylamine (DAPP) and (N,N-dimethyl-2-(2-amino-4-cyanophenylthio) benzylamine (D ASB), were radiolabeled with carbon-11 and evaluated as in vivo probes of the serotonin transporter (SERT) using positron emission tomography (PET). Both compounds are highly selective, with nanomolar affinity for the serotonin transporter and micromolar affinity for the dopamine and norepinephrine transporters. Six volunteers were imaged twice, once with each of the two radioligands. Both ligands displayed very good brain penetration and selective retention in regions rich in serotonin reuptake sites. Both had similar brain uptake and kinetics, but the cyano analogue, [11C]DASB, had a slightly higher brain penetration in all subjects. Plasma analysis revealed that both radiotracers were rapidly metabolized to give mainly hydrophilic species as determined by reverse-phase high-performance liquid chromatography. Inhibition of specific binding to the SERT was demonstrated in three additional subjects imaged with [11C]DASB following an oral dose of the selective serotonin reuptake blocker citalopram. These preliminary studies indicate that both these substituted phenylthiobenzylamines have highly suitable characteristics for probing the serotonin reuptake system with PET in humans. PMID- 11105831 TI - Imaging the glutamatergic system in vivo--relevance to schizophrenia. AB - Schizophrenia is a devastating psychiatric illness. Its pathophysiology is not fully clarified. Animal data, in vitro and indirect in vivo imaging support glutamatergic N-methyl-D-aspartate (NMDA) receptor hypofunction in the disorder. A lack of suitable ligands has obstructed direct evaluation of the NMDA receptor hypofunction hypothesis of schizophrenia. Many research groups are working towards developing appropriate single-photon emission tomography and positron emission tomography ligands for the NMDA receptor. This paper briefly presents evidence for links between glutamatergic system dysfunction and schizophrenia. It reviews the radioligands to evaluate glutamatergic receptors in vivo and discusses issues in developing novel ligands for the glutamatergic system. PMID- 11105832 TI - Quantifying the radiation exposure of nursing staff delivering hyperbaric oxygen therapy to grade IV neuroblastoma patients post 131I-MIBG therapy. PMID- 11105833 TI - Neuropsychological and neurophysiological evaluation after anterior communicating artery (ACoA) aneurysm surgery. AB - BACKGROUND: We evaluated high cerebral functions 6 months after surgery for bleeding ACoA aneurysms comparing neurophysiological and neuropsycological tests. METHODS: Twelve patients were chosen among a series of cases operated on in the first 48 hours after ACoA aneurysm bleeding. All of them were in Hunt-Hess grade I or II. We excluded patients over 65 years, or with intracranial haematomas, intraventricular haemorrhage, hydrocephalus, or with multiple or giant aneurysms. All of them underwent neurophysiological evaluation with recording and mapping of long latency (P300) auditory and visual event-related potentials (ERPs) and a neuropsychological assessment for memory, intelligence, frontal lobe functions and language. RESULTS: Neuropsychological assessment: All patients were severely damaged on phonemic fluency. In a first group (group A: 3 cases) tests were all in a normal range. In a second (group B: 3 cases) the tests showed severe impairment on learning and long term memory. In a third (group C: 6 cases) tests showed memory and "frontal lobe" deficits. Neurophysiological assessment: The whole group of patients showed significant delay in ERPs recordings compared to controls. ERPs of patients in group A and B showed no significant differences from controls, while being significantly delayed in 5 patients out of 6 of group C. CONCLUSIONS: All patients had difficulties in the phonemic task in which a notable cognitive effort is necessary, while intelligence, short term memory, attention and language were within normal limits. Patients in group C showed severe frontal lobe type cognitive impairment. Those ones in groups A and B did not present cognitive derangements (A) or only memory and learning impairment (B). ERPs may be an objective parameter in the follow-up of cases with cognitive impairment, even if neurophysiological tests cannot be replaced. PMID- 11105834 TI - Shoulder-hand syndrome in neurosurgical patients treated with barbiturates. A long term evaluation. AB - OBJECTIVES: To assess the incidence of shoulder-hand syndrome (SHS) in neurosurgical patients (head injuries, intracranial ruptured aneurysms and intracranial meningiomas), treated with barbiturates. SHS is a chronic condition characterized by intense tenderness and functional impairment affecting one hand, the shoulder or both. Barbiturates have been identified as cause of SHS, although there is controversial evidence on the incidence of this disorder in patients started on long-term Phenobarbital (PB) therapy. METHODS: One hundred and twenty six neurosurgical patients, treated with barbiturates, and a control group of 108 patients, treated with carbamazepine or phenytoin, were enrolled. Both groups were followed up for at least 24 to 36 months. RESULTS: Thirty-five PB-treated patients (27.6%) experienced SHS. In these patients SHS developed during the first 7 months of therapy and regressed after PB discontinuation or, in 2 cases, after dosage reduction. None of the patients in the control group developed SHS. CONCLUSIONS: The occurrence of SHS in the study group was much more common than that reported previously. This higher incidence should depend upon the coexistence of separate risk factors such as age over 50 years, surgery and intracranial pathology. Early diagnosis and rapid withdrawl of treatment are important for symptomatic relief and full functional recovery. PMID- 11105835 TI - Positive predictive values of selected clinical signs associated with skull base fractures. AB - BACKGROUND: The goal of this study was to determinate the positive predictive values of selected clinical signs for skull base fractures and associated intracranial lesions. METHODS: EXPERIMENTAL DESIGN: Clinical and radiological data were collected prospectively for all patients with selected clinical signs of skull base fractures, and their admission criteria were: 1) recent head injury story; 2) presence of one or more of following clinical signs: unilateral or bilateral blepharohaematoma, bloody otorrhea, and Battle's sign. SETTING: Emergency service of a institutional hospital. PATIENTS: One hundred forty two patients with the selected clinical signs for skull base fracture. RESULTS: Frontal bone fractures were the most frequent in patients with selected clinical signs. Battle's sign (100%) and unilateral blepharohaematoma (90%) were the signs with higher positive predictive values for skull base fractures; bilateral blepharohaematoma (70%) and bloody otorrhea (70%) were those with less values. The positive predictive values of the selected signs for intracranial lesions (acute extradural haematoma, pneumocephalus, brain contusion, brain sweLling, and acute subdural haematoma) were: unilateral and bilateral blepharohaematoma with positive predictive values of 85% and 68%, respectively; Battle's sign was 66%; and bloody otorrhea was 46%. For patients at admission on the 13-15 Glasgow Coma Scale only, the positive predictive values for that intracranial lesions were: blepharohaematoma=78%; Battle's sign=66%; and bloody otorrhea=41%. CONCLUSIONS: Our data demonstrated that the selected signs of skull base fractures have high positive predictive values for the presence of skull fracture and intracranial lesions, even in those patients classified in the Glasgow Coma Scale between 13 and 15. This indicates that all patients with the selected clinical signs should be submitted to computerized tomography of skull and with bone window, with the aim to detect associated lesions. PMID- 11105836 TI - Peri- and postoperative pain valutation in carpal tunnel release of median nerve compression. AB - We analysed 108 patients, operated on day surgery, for carpal tunnel release of median nerve compression, to evaluate peri- and postoperative pain. We made in all cases a short intertenarian incision (25 mm) with microsurgical technique and local anaesthesia using mepivacaine 2% without vasoconstrictor. We evaluated pain for local anaesthetic infiltration as VRS (Verbal Rating Scale) 6,3 median-time to the first possible analgesic assumption (in all cases paracetamol 500 mg), total analgesic assumption, pressure algometry (to evaluate "allodiny") after the first 48 hours and subjective pain intensity by a numerical pain scale. Pain intensity on first drug assumption (after a mean time of 7 hours from the end of surgery) had a mean VAS value of 2,15; while after a second assumption of analgesic (after a mean time of 15 hours from surgery) had a mean VAS value of 2. Mean total analgesic assumption was 1,64 tablets of paracetamol 500 mg. From these data we may deduce that peri- and postoperative pain following median nerve decompression with this technique and anaesthesia, has a moderate intense peak of brief duration, for local anaesthetic infiltration (that seems to be the most painful event) and modest and not constant pain in the postoperative time (more evident 7 and 15 hours from the end of surgery). It may be useful association with mepivacaine bicarbonate solutions or injecting less painful local anaesthetic. PMID- 11105837 TI - Glial cyst of the pineal gland: case report and considerations about surgical management. AB - Symptomatic glial cyst of the pineal gland are rare lesions. Origin, natural history and factors leading to cyst enlargement are not completely clear; thus management remain uncertain in some cases. We report a case of symptomatic glial cyst and analyze the implication for surgery. Surgical management is indicated in patients presenting hydrocephalus, mass effect or symptoms related to mesencephalic dysfunction. The infratentorial supracerebellar approach represent the first choice for this condition allowing easy orientation with wide exposure of the tumor and good visibility of deep venous systems that may be preserved. Size of the tumor is a key element in evaluation of the treatment and the appropriate course for asymptomatic cyst less than 1 cm in size consist of conservative management. Periodic follow up is always indicated. PMID- 11105838 TI - Spinal cord ependymoma presenting with acute paraplegia due to tumoral bleeding. AB - Acute paraplegia is a rare presentation for a spinal cord ependymoma Among spinal cord tumors ependymomas are most commonly associated with subarachnoid hemorrhage and there is evidence that some have had intratumoral hemorrhage, but most of these bleedings pass without symptoms. In this report, a case of spinal cord ependymoma debutting with acute neurological deterioration due to tumoral bleeding is presented. We discuss the clinical and neuroradiological findings and review the literature related to this unusual presentation. PMID- 11105839 TI - Intramedullary spinal cord metastasis. A case report. AB - This 54-year-old patient with a breast carcinoma of one year's evolution presented a progressive paraparesis and sphincter disregulation of a week evolution; MRI image showed a tumor in the medullary conus. She improved after removal of the conus mass. The histologic diagnosis was metastasis of adenocarcinoma. Metastasis at this level is infrequent and represents less than 1% of all spinal metastases. When the patients' general condition is good, surgery can relieve the neurologic deficit produced by the medullary mass. PMID- 11105840 TI - Fatal carotid dissection after blunt head trauma. AB - Occurrence of internal carotid artery injuries associated with skull base fracture has been reported. A. report a case of fatal intracranial carotid dissection related to petrous fracture involving the carotid canal. Identification of carotid lesions may be difficult and generally related to appearance of unexpected neurological deficit. Skull base fractures may be considered an indirect sign for detection of vascular injury. Patterns of the fracture are of paramount importance; routine CT scan may fail to detect basilar fractures and high definition fine-cut CT scan should be executed to carefully identify and evaluate fractures. Temporal and sphenoid bone fractures are common in head trauma and involvement of the course of the carotid artery is frequent. The involvement of the intracranial carotid artery course represents a direct risk factor for lesions of the petrous, lacerum and cavernous segments of the carotid artery. Early diagnosis of post-traumatic vascular injury may lead to prognosis improvement because of effectiveness of heparin anticoagulant therapy. Then vascular screening is recommendable in cases with complex fractures of the skull base and particularly fracturing along the course of the carotid artery. Magnetic resonance angiography may be considered the first line diagnostic tools for vascular screening. Angiography may be reserved for patients with a proven lesion or rapid neurological deterioration taking into account the possibility of interventional treatment. PMID- 11105841 TI - Cranio-orbital missile wound and bullet migration. Case report. AB - An unusual case of craniocerebral missile injury, with orbital roof perforation and spontaneous bullet migration into the maxillary sinus, is reported. Emergency treatment consisted in wide craniectomy around the bullet entry point, blood and foreign bodies debridement. Subsequent procedures were necessary for abscess evacuation, transmaxillary bullet removal and later cranial vault reconstruction. Challenging aspects were the treatment of the infectious complications, following cerebrospinal fluid fistula through the wound, and the onset of post-traumatic epilepsy, scarcely responsive to common antiepileptic drugs. The treatment of the abscess by combined systemic and intracavitary antibiotic therapy and of the chronic seizures by progressive adjustment with new protocols of antiepileptic drugs under EEG and brain mapping revealed successful. PMID- 11105842 TI - Determination of morphine by capillary electrophoresis immunoassay in thermally reversible hydrogel-modified buffer and laser-induced fluorescence detection. AB - In this paper thermally reversible hydrogel used as a replaceable packed material for capillary electrophoresis was examined. A simple and rapid method of detecting morphine was developed, which demonstrated the potential of strong affinity antibodies as a selector for immunologically-based separations in serum by capillary electrophoresis. Polyclonal antibodies were linked to hydrogel and applied to the separation of free fluorescein isothiocyanate (FITC)-labeled antigen and bound FITC antigen. The separation was monitored with laser-induced fluorescence detection. Different separation conditions were studied. The results indicated that poly-N-isopropylacrylamide hydrogel (PNIPA) is a kind of steady, replaceable gel. The specific determination of morphine did not require a long incubation time and PNIPA hydrogel-modified antibodies can be stockpiled at 4 degrees C before assay. It can be used to determine morphine with good precision and a detection limit lower than 8.5 ng/ml. Details of the preparation of hydrogel cross-linked polyclonal antibody and of typical separations of bound and free antigen are presented. PMID- 11105843 TI - Micellar electrokinetic and high-performance liquid chromatographic determination of potential manufacturing impurities in pholcodine. AB - Quantitative high-performance liquid chromatographic (HPLC) and micellar electrokinetic chromatographic (MEKC) methods have been developed for the determination of four structurally related potential manufacturing impurities, including morphine, of the opiate derivative pholcodine. Pholcodine and the four impurities were separated by MEKC in less than 14 min using a 70 cm x 75 microm I.D. uncoated fused-silica capillary (25 kV at 30 degrees C) and a running buffer consisting of 10% acetonitrile (v/v) in 20 mM borate-phosphate buffer pH 8.0 containing 40 mM sodium dodecyl sulphate (SDS). The MEKC method was compared to a HPLC method using a 5 microm Luna phenyl-hexyl column (150 x 4.6 mm I.D.) eluted with a mobile phase consisting of a mixture of 10% (v/v) acetonitrile, 7% (v/v) tetrahydrofuran in 20 mM phosphate buffer pH 8.0. Both methods were fully validated and a comparison was made regarding selectivity, linearity, precision, robustness and limits of detection and quantitation. The presence of the impurities in different samples of pholcodine drug substance was investigated using both methods. PMID- 11105844 TI - Effects of aliphatic amines on capillary electrochromatographic performance of tricyclic antidepressants on octadecylsilica. AB - Adding aliphatic amines to the mobile phase improves peak symmetry and efficiency in capillary electrochromatography of tricyclic antidepressants on octadecylsilica. The most hydrophobic aliphatic amine studied, dimethyloctylamine (DMOA), was the most efficient. Despite the fact that the amine additives substantially reduced the electroosmotic flow, the retention of the analytes decreased indicating a strong competitive effect of the additives. DMOA gave the largest retention decrease, and simultaneously reduced the resolution, indicating that silanophilic interaction is significant to the separation. Highest efficiencies were obtained at the lowest pH (2.8). Acetonitrile influenced both efficiency and peak symmetry, and best results were obtained at 60%. PMID- 11105845 TI - Adaptation of capillary electrophoresis to the determination of selected cephalosporins for injection. AB - The migration behaviour of cephazolin, cefuroxime sodium, ceftriaxone sodium, cefoperazone sodium and ceftazidime in a mixture was studied. Phosphate-borate buffer pH 5-8 alone and with addition of sodium dodecylsulfate (SDS) was used. In capillary zone electrophoresis of all research compounds separation was not achieved. It was observed that supplementation buffer pH 6.5 with SDS (10 g/l) improved resolution of cephalosporins, but addition of pentanesulfonic acid (17.4 g/l) to the running buffer at pH 6.5 results in separation of each cephalosporin. In this condition good repeatability of migration times as well as repeatability of peak area were confirmed. PMID- 11105846 TI - Optimization of capillary electrophoretic separation of several inhibitors of the angiotensin-converting enzyme. AB - Capillary electrophoretic separation of eight inhibitors of the angiotensin converting enzyme, viz., enalapril, lisinopril, quinapril, fosinopril, perindopril, ramipril, benazepril and cilazapril, was investigated with respect to the following parameters: pH of the running buffer, organic modifiers and surfactants. The most critical parameter is the pH of the running buffer. The addition of sodium dodecyl sulfate had a negative influence on the peak symmetry, and selectivity was not improved. The separation of the eight compounds can be performed by means of two phosphate buffers (each 100 mM) at pH 7.0 and pH 6.25, respectively. This combination is necessary for the selective identification of structurally related substances because of their similar pKa values. PMID- 11105847 TI - Analysis of doxycycline by capillary electrophoresis. Method development and validation. AB - An optimized capillary electrophoresis method for the analysis of doxycycline is described. The influence of methanol as organic modifier, buffer pH, buffer concentration, capillary length, column temperature, Triton X-100 and methyl-beta cyclodextrin was systematically investigated. A central composite design was performed in order to optimize the method. The optimal separation conditions were: capillary, uncoated fused-silica [40 cm (32 cm effective length) x 50 microm I.D.]; background electrolyte, a solution of 145 mM sodium carbonate and 1 mM EDTA brought to pH 10.3-methanol (89:11, v/v); temperature, 15 degrees C; voltage, 12 kV. The method showed good selectivity, repeatability, linearity and sensitivity. Six commercial samples were quantitatively analyzed. The results were compared with those established by the liquid chromatography method from the European Pharmacopoeia. PMID- 11105848 TI - Chiral capillary electrophoresis as predictor for separation of drug enantiomers in continuous flow zone electrophoresis. AB - Separation of the enantiomers of chlorpheniramine and methadone in acidic buffers containing carboxymethyl-betacyclodextrin (CMCD) as chiral selector was investigated by capillary zone electrophoresis. For a range of pH and CMCD concentrations, the mobility difference and resolution of the enantiomers were determined. Then, conditions known to provide well resolved enantiomers and optimized chiral separation were applied to chiral continuous flow electrophoresis. In that approach, a thin film of fluid flowing between two parallel plates is employed as carrier for electrophoresis. The electrolytes and the sample are continuously admitted at one end of the electrophoresis chamber and are fractionated by an array of outlet tubes at the other. The number of pure enantiomeric fractions obtained by chiral continuous flow electrophoresis was found to be directly dependent on the enantiomeric mobility difference. For racemic chlorpheniramine separated in a betaine-acetic acid buffer at a total throughput of 5 mg/h, complete enantiomeric separation is shown to require a mobility difference of about 3 x 10(-9) m2/V s. Furthermore, compared to the previous investigations with hydroxypropyl-beta-cyclodextrin, CMCD was found to permit improved fractionation of methadone enantiomers. With a total racemic drug throughput of about 15 mg/h, continuous flow zone electrophoresis processing with CMCD as chiral selector is shown to have the potential of providing pure enantiomers on a mg/h scale. The results indicate that chiral capillary zone electrophoresis data can be employed as predictor for preparative scale chiral separations based upon continuous flow zone electrophoresis. PMID- 11105849 TI - Capillary electrophoresis analysis of gentamicin sulphate with UV detection after pre-capillary derivatization with 1,2-phthalic dicarboxaldehyde and mercaptoacetic acid. AB - A selective, sensitive, and rapid pre-capillary derivatization method for determination of the multicomponent aminoglycoside antibiotic gentamicin is described. The derivatization reagents 1,2-phthalic dicarboxaldehyde and mercaptoacetic acid were used and the thioisoindole derivative was UV detected at 330 nm. A central composite experimental design was performed to optimize selectivity and derivatization conditions. Baseline separation of gentamicin C1, C1a, C2, C2a, C2b, sisomicin and several minor components was achieved with a background electrolyte containing 30 mM sodium tetraborate, 7.5 mM beta cyclodextrin and 12.5% (v/v) methanol at pH 10. Quantitative analysis was performed and illustrated the potential use of capillary electrophoresis for the identification and quantitation of gentamicin as an alternative to methods prescribed in the United States Pharmacopeia and European Pharmacopoeia. PMID- 11105850 TI - Non-aqueous capillary electrophoresis of tamoxifen and its acid hydrolysis products. AB - Tamoxifen and its acid hydrolysis products were separated and tentatively identified by non-aqueous capillary electrophoresis with thermooptical absorbance and electrospray ionization mass spectrometry. Acid hydrolysis is a convenient method of generating tamoxifen degradation products. The parent compound and seven hydrolysis products were separated in 9 min. PMID- 11105851 TI - Analysis of 2beta-carbomethoxy-3beta-(4-fluorophenyl)-N-(3-iodo-E allyl)nortropane in rat plasma. I. Method development and validation by capillary electrophoresis. AB - Altropane, 2beta-carbomethoxy-3beta-(4-fluorophenyl)-N-(3-iodo-E-allyl)nor tropane, is an imaging agent that was developed recently for early detection of Parkinson's disease. Its promise as a useful radiopharmaceutical for single photon emission computed tomography or positron emission tomography imaging of the brain has been well demonstrated, and it is currently undergoing clinical trials. This paper presents methods development and validation of capillary electrophoresis (CE) techniques to analyze Altropane in aqueous environments as well as in rat plasma, using an internal standard, nicotinamide. N Allylaltropane, 2beta-carbomethoxy-3beta-(4-fluorophenyl)-N-allylnortropane, which is a known degradation product of the Altropane precursor (tributyltinaltropane), was used to verify the method's specificity. A solid phase extraction method for extraction of Altropane from rat plasma is also described. The results presented in this paper demonstrate the applicability of CE methods to study the pharmacokinetic properties of Altropane in animal models. The results of the pharmacokinetic study will be published later, as Part II. PMID- 11105852 TI - Determination at ppb level of an anti-human immunodeficiency virus nucleoside drug by capillary electrophoresis-electrospray ionization tandem mass spectrometry. AB - Capillary electrophoresis coupled with tandem mass spectrometry was used to indirectly separate and quantify the active metabolite of the anti-human immunodeficiency virus (anti-HIV) didanosine drug. The influence of several parameters (pH and ionic strength of volatile formic acid-ammonia buffer) upon electroosmotic flow, electrophoretic mobility and peak efficiency of several nucleosides (A, dA, ddA, C) has been studied. This paper illustrates the current importance in CE-MS technique as a complementary or substituted method to the known HPLC-radioimmunoassay or HPLC-UV method to measure levels of anti-HIV drugs. The limit of detection for 2',3'-dideoxyadenosine by this method is 2 microg 1(-1) in a formic acid-ammonia buffer (pH 2.5, 10 mM ionic strength). This methodology could be used to perform simultaneous detection of two or more anti HIV nucleosides, such as stavudine or didanosine in combination therapy. PMID- 11105853 TI - Simultaneous analysis of some amphetamine derivatives in urine by nonaqueous capillary electrophoresis coupled to electrospray ionization mass spectrometry. AB - A nonaqueous capillary electrophoresis method, coupled to UV and electrospray mass spectrometry (ESI-MS), is described for the simultaneous analysis of Ecstasy and other related derivatives. Several electrophoretic and ESI-MS parameters were systematically investigated, such as electrolyte nature and concentration, organic solvent and sheath liquid compositions, nebulization gas pressure and drying gas flow-rate. The best results were achieved with an acetonitrile methanol (80:20, v/v) mixture containing 25 mM ammonium formate and 1 M formic acid, an applied voltage of 30 kV and a separation temperature of 15 degrees C. Under optimized CE-ESI-MS conditions, separation of the investigated drugs was performed in less than 6 min, with a high efficiency. Method precision based on migration time and peak area was determined and the limits of detection, which depend on the tested compound, were established between 20 and 70 ng ml(-1) in the selected ion monitoring mode. Finally, the described method was successfully applied to the analysis of amphetamines in urine after a liquid-liquid extraction. PMID- 11105854 TI - Capillary electrochromatography and capillary electrochromatography-electrospray mass spectrometry for the separation of non-steroidal anti-inflammatory drugs. AB - In this study capillary electrochromatography (CEC) was utilized for the separation of ten non-steroidal anti-inflammatory drugs (NSAIDs). Experiments were carried out in a commercially available CE instrument using a packed capillary with RP-18 silica particles where the stationary phase completely filled the capillary. The mobile phase consisted of a mixture of ammonium formate buffer pH 2.5 and acetonitrile. Selectivity and resolution were studied changing the pH and the concentration of the buffer, the acetonitrile content mobile phase and the capillary temperature. The optimum experimental conditions for CEC separation of the studied drug mixture were found using 50 mM ammonium formate pH 2.5-acetonitrile (40:60) at 25 degrees C. The CEC capillary was coupled to an electrospray mass spectrometer for the characterization of the NSAIDs. A mobile phase composed by the same buffer but with a higher concentration of acetonitrile (90%) was used in order to speed up the separation of analytes. PMID- 11105855 TI - Analysis of codeine, dihydrocodeine and their glucuronides in human urine by electrokinetic capillary immunoassays and capillary electrophoresis-ion trap mass spectrometry. AB - Screening for and confirmation of illicit, abused and banned drugs in human urine is a timely topic in which capillary separation techniques play a key role. Capillary electrophoresis (CE) represents the newest technology employed in this field of analysis. Two rapid competitive binding, electrokinetic capillary-based immunoassays are shown to be capable of recognizing the presence, but not the identity, of urinary opioids, namely codeine (COD), codeine-6-glucuronide, dihydrocodeine (DHC), dihydrocodeine-6-glucuronide, morphine (MOR), morphine-3 glucuronide and ethylmorphine (EMOR). In these approaches, aliquots of urine and immunoreagents of a commercial, broadly cross-reacting fluorescence polarization immunoassay for opiates were combined and analyzed by capillary zone electrophoresis or micellar electrokinetic capillary chromatography with laser induced fluorescence detection. With the fluorescent tracer solution employed, the former method is shown to provide simple electropherograms which are characterized by an opioid concentration dependent magnitude of the free tracer peak. In presence of dodecyl sulfate micelles, however, two tracer peaks with equal opioid concentration sensitivity are monitored. These data suggest the presence of two fluorescent tracers which react competitively with the urinary opioids for the binding sites of the antibody. Assay sensitivities for COD and MOR are comparable (10 ng/ml), whereas those for DHC and EMOR are about four-fold lower. Furthermore, glucuronides are shown to react like the corresponding free opioids. Analysis of urines that were collected after administration of 7 mg COD and 25 mg DHC tested positively in both assay formats. The presence of the free and conjugated codeinoids in these urines and their identification was accomplished by capillary electrophoresis-ion trap mass spectrometry (CE-MS). This confirmatory assay is based upon solid-phase extraction using a mixed-mode polymer cartridge followed by CE hyphenated to the LCQ mass spectrometer with electrospray ionization in the positive ion mode. With this technology, MS2 is employed for proper identification of COD (m/z 300.4) and DHC (m/z 302.4) whereas MS3 provides unambiguous identification of the glucuronides of COD (m/z 476.5) and DHC (m/z 478.5) via their fragmentation to COD and DHC, respectively. MSn (n > or = 2) is shown to be capable of properly identifying the urinary codeinoids on the 100-200 ng/ml concentration level. PMID- 11105856 TI - Capillary electrophoresis with laser-induced fluorescence in clinical drug development routine application and future aspects. AB - The clinical bioanalytical setting is characterized by sample volumes of < 1 ml biological fluid (e.g. plasma, urine), a range of 3-4 decades of concentrations to be quantified and a limit of quantitation in the microg/l-ng/l range for sets of 100-5000 individual samples. Setup of capillary electrophoresis (CE) for routine application in this analytical field was successful for analytes accessible to fluorescence detection by using laser-induced fluorescence (LIF) detection. Empowerment of CE-LIF for routine serial analysis of thousands of samples includes improvement in autosampler techniques, thorough procedures for capillary treatment and particularly more advanced detection technology. Introduction of multi-capillary systems with charge-coupled device cameras and frequency doubled Ar-ion laser (lambda = 257 nm) offers this technique the chance of superiority over classical analytical assays - especially in the field of (new) low volume samples e.g. capillary blood or microdialysate encouraging clinicians to search for meaningful non-invasive samples. PMID- 11105857 TI - Determination of biologically active low-molecular-mass thiols in human blood. I. Fast qualitative and quantitative, gradient and isocratic reversed-phase high performance liquid chromatography with photometric and fluorescence detection. AB - The fast isocratic and gradient reversed-phase high-performance liquid chromatographic methods employing photometric and/or fluorescence detection are described for the precise reproducible simultaneous measurement of total homocysteine, cysteine, and glutathione in human blood. Sample preparation involves conversion of disulfides to free thiols with triphenylphosphine, precipitation of proteins with sulfosalicylic acid, and conjugation of thiols with monobromobimane. The aminothiol assay is optimized by reduction and derivatization step conditions (pH, temperature and time of reactions), as well as by chromatographic conditions to obtain reliable quantitative results within the concentration range corresponding to the levels of these thiols in human blood in norm and pathology. Its sensitivity allows the detection of aminothiol quantities >2 pmol. PMID- 11105858 TI - Determination of biologically active low-molecular-mass thiols in human blood. II. High-performance capillary electrophoresis with photometric detection. AB - A new high-performance capillary electrophoresis assay for aminothiols in human blood, including homocysteine, a marker of several human metabolism disorders, has been developed. Sample preparation involves conversion of disulfides to free thiols with triphenylphosphine, precipitation of proteins with sulfosalicylic acid, and conjugation of the thiols with monobromobimane. Derivatized thiols were separated in a sodium phosphate buffer using a fused-silica capillary (65 cm x 50 microm I.D.) at 30 degrees C. With the electric field of 250 V cm(-1), separation of homocysteine, glutathione and cysteine occurred at less than 10 min. Detection at 250 or 234 nm was used to confirm the monobimane-thiols peaks. The detection limit was approximately 5 nmol/ml for all labeled aminothiols. The proposed method for these compounds' analysis included simple sample preparation, high selectivity, good linearity (r2>0.999), high reproducibility (within-run precision for derivatized aminothiol peaks area RSD<5% for three times consequently injected sample); high reliability and the small volumes required for analysis made it suitable for clinical studies. PMID- 11105859 TI - Assays for total homocysteine and other thiols by capillary electrophoresis-laser induced fluorescence detection. I. Preanalytical condition studies. AB - In recent papers, we presented a new analytical method for thiol quantification in serum. It is based on the use of capillary electrophoresis and laser-induced fluorescence to analyze thiol 6-iodoacetamidofluoresceine (IAF) derivatives. Quantitative results of homocysteine, glutathione, cysteine-glycin, and cysteine were shown (Clin. Chem. 45 (1999) 412). A comprehensive comparison of the quantitation of homocysteine in serum, using high-performance liquid chromatography/conventional fluorescence detection and fluorescence polarization immunoassay was also used (E. Causse et al., Electrophoresis 21 (2000) 2074). Sample preparation prior to derivatization with IAF had never been investigated. In this work we present the results of quantitation of thiols in serum and plasma with three different anticoagulants widely used: ethylenediaminetetraacetic acid (EDTA), heparin, and sodium citrate. We show that serum and EDTA plasma gave the same results. Then serum protein precipitations by acetonitrile, acetone, sulfosalicylic acid, perchloric acid and trichloracetic acid, prior to derivatization by IAF, were also investigated. Their influence on the concentrations of the thiols were determined. Sulfosalicylic acid and acetonitrile precipitations are well adapted, whereas acetone cannot be used. PMID- 11105860 TI - Specific thiol determination by micellar electrokinetic chromatography and on column detection reaction with 2,2'-dipyridyldisulfide. AB - A new method for specific determination of glutathione using micellar electrokinetic chromatography and on-column reaction with 2,2'-dipyridyldisulfide is described. 2,2'-Dipyridyldisulfide and a sample of glutathione are injected consecutively into the capillary as two discrete plugs separated with a short plug of background electrolyte. Due to the differences in the mobilities of the 2,2'-dipyridyldisulfide and glutathione, on-column mixing and reaction occur. Glutathione is in this reaction quantitatively transformed into a mixed disulfide concomitantly with formation of an equimolar amount of the 2-thiopyridone which is further separated by micellar electrokinetic chromatography and determined spectrophotometrically at 343 nm. The concentration of glutathione is thus estimated indirectly from the result of 2-thiopyridone determination. PMID- 11105861 TI - Determination of soluble anions and cations from waters of pulp and paper mills with on-line coupled capillary electrophoresis. AB - When the degree of closure of the paper machine wet end waters increases, wet end problems also become more difficult to control without specific and selective on line measurements. The need to measure the concentrations of individual compounds in order to explain wet end phenomena is growing. This study was performed to set up a CE system to a paper machine and to determine soluble inorganic and organic ions in different locations of pulp and paper process waters with real time analyses by two on-line CE methods. A reconstructed commercial CE apparatus was connected to a papermaking machine via an apparatus, which was a combined sampling and sample pretreatment instrument, the role of which was to filter and dilute the samples before on-line determination by CE. The on-line procedures were optimized for simultaneous determination of anions as chloride, sulfate, oxalate, formate and acetate and for determination of cations as potassium, calcium, sodium, magnesium and traces of aluminium. The quantification was performed with external standard methods using the programs available in the commercial CE instrument. The concentrations of the ions were transferred by using a computerized transfer algorithm exporting the results from the analysis instrument to the process control unit. The developed on-line procedures were tested three times in paper and paperboard mills for 1 month at a time. Correlations were observed between the CE results and changes in the processes. PMID- 11105862 TI - Determination of aromatic amines in water samples by capillary electrophoresis with electrochemical and fluorescence detection. AB - Two capillary electrophoresis methods have been compared for the determination of aniline derivatives in environmental water samples. With the first method the anilines were separated as cations by free zone electrophoresis at low pH, and detected by amperometry. For this, the separation capillary was connected through a palladium field decoupler to an electrochemical detection cell which had been modified to match the volume scale of the separation. Most anilines tested, except chlorinated compounds, could be detected with full sensitivity at a detection potential of +0.7 V. Detection limits with this detection scheme were on a low microg/l level. The alternative method involved the derivatization of the anilines with fluorescamine, the separation of the derivatives formed by micellar electrokinetic chromatography, and fluorescence detection. For detection a lamp-based, fibre optics instrument was used. Detection limits with fluorimetry were comparable with those obtained with amperometric detection (in the order of 1 microg/l). Still, this method was preferred since it gave a higher separation efficiency and shorter analysis times (approximately 4 min). The most important argument, however, was its higher reliability and ease-of-handling. Preliminary experiments with water samples collected in areas where pollution with anilines may be expected showed that the method is highly specific, with few interferences showing up in the electropherograms. PMID- 11105863 TI - Simultaneous determination of alkali, alkaline earth and transition metal ions by capillary electrophoresis with indirect UV detection. AB - Determination of metal ions in aqueous samples using capillary electrophoresis can be accomplished with indirect UV detection. For optimal determination of alkali, alkaline earth, and transition metal ions, several electrolyte systems were studied. Detection at 214 nm was performed with a background electrolyte containing reagents with inherent absorbance in the UV range: imidazole, 4 methylbenzylamine and 4-aminopyridine. Glycolic acid and alpha-hydroxyisobutyric acid were used as complexing reagents. A mixture of 16 metal ions was successfully separated. The detection limits were between 92 ppb for Ca and 454 ppb for Cu with hydrostatic injection. All peaks were completely resolved and well separated. A separation efficiency of about 650,000 theoretical plates per meter was achieved for the Mg ion. The described methods can be used successfully in routine analysis of real samples. One of the methods was applied to an environmental water sample from the Georgian river Kasretula. PMID- 11105864 TI - Separation of bisphenol A and three alkylphenols by micellar electrokinetic chromatography. AB - Analytical conditions of pH, surfactants, and additives were investigated for the simultaneous separation of bisphenol A and alkylphenols by micellar electrokinetic chromatography. Reproducibility of migration time was improved at higher pH (pH 8.0). When five surfactants having linear alkyl chains or four bile salts were used, the separation of hydrophobic phenols and 4-nonylphenol isomers was not achieved. In order to improve the separation, the use of additives with sodium dodecyl sulfate solution was investigated. The separation of hydrophobic phenols was improved by the addition of organic solvents, however, isomers were not separated. Their separation was achieved by the addition of beta- or gamma cyclodextrin. PMID- 11105865 TI - Determination of haloalkane dehalogenase activity by capillary zone electrophoresis. AB - A new sensitive method has been developed for the determination of haloalkane dehalogenase activity. The enzymatic reactions were carried out directly in thermostatted autosampler vials and the formation of product - bromide or chloride ions - was monitored by sequential capillary zone electrophoresis runs. The determinations were performed in a 75 microm fused-silica capillary using 5 mM chromate, 0.5 mM tetradecyltrimethylammonium bromide (pH 8.4) as a background electrolyte, separation voltage 15 kV (negative polarity) and indirect detection at sample wavelength 315 nm, reference wavelength 375 nm for brominated and chlorinated substrates, respectively 0.1 M beta-alanine-HCl (pH 3.50) as a background electrolyte, separation voltage 18 kV (negative polarity) and direct detection at 200 nm for brominated substrates. The temperature of capillary was in both cases 25 degrees C. The method is rapid, can be automated, and requires only small amount of enzyme preparation and substrate. PMID- 11105866 TI - Determination of cationic surfactants by capillary zone electrophoresis and micellar electrokinetic chromatography with deoxycholate micelles in the presence of large organic solvent concentrations. AB - Mixtures of the cationic surfactants benzalkonium chloride (BKC) and cetylpyridinium chloride (CPC) were quickly resolved and reproducibly and reliably determined by using background electrolytes (BGEs) containing 80 mM borate, pH 8.5, bile salts and large concentrations of an organic solvent. When the bile salt is present, the separation mechanism changes from capillary zone electrophoresis (CZE) to a mixed micellar electrokinetic chromatography (MEKC) CZE, with predominant MEKC interactions, which lead to an excellent resolution of all the solutes, including the C12-C18 homologues of BKC and CPC. A BGE containing 50 mM sodium deoxycholate and 30% ethanol for an extreme resolution, or 20% tetrahydrofuran for an adequate resolution within a much shorter analysis time, is recommended. The procedure was applied to the determination of the surfactants in industrial and household formulations, with excellent resolution between the homologues, detection limits of a few microg ml(-1) and reproducibilities below 2%. PMID- 11105867 TI - Separation of two groups of oestrogen mimicking compounds using micellar electrokinetic chromatography. AB - Two groups of compounds are being investigated due to their reported oestrogen mimicking characteristics in the environment. Separation of phenolic compounds and synthetic oestrogens using micellar electrokinetic chromatography is reported. Photodiode array detection is used for both separations. A standard separation buffer can be used for both groups of compounds including zwitterionic buffer cyclohexylamino-1-propanesulfonic acid, 20 mM at pH 11.5. It was found necessary to include 15% acetonitrile and 25 mM sodium dodecyl sulfate to aid separation and maintain analytes in solution. Optimum separations are achieved using 20 kV with hydrodynamic injection for 5 s. The relative standard deviation (RSD) for reproducibility was investigated for a mixture of phenols and synthetic oestrogens. For these compounds RSD was found to be <0.6% in all cases. Peak efficiencies ranged from 76,000 to 150,000 theoretical plates for different analytes. Application to environmental samples is discussed. PMID- 11105868 TI - Capillary electrophoretic separation of the enantiomers of organophosphates with a phosphorus stereogenic center using the sodium salt of octakis(2,3-diacetyl-6 sulfo)-gamma-cyclodextrin as resolving agent. AB - The sodium salt of the single-isomer, chiral resolving agent, octakis(2,3 diacetyl-6-sulfo)-gamma-cyclodextrin (ODAS-gammaCD) has been used for the capillary electrophoretic separation of the enantiomers of alkylarylphosphates which carry a phosphorus-based stereogenic center. The effective mobilities and separation selectivities were measured at different ODAS-gammaCD and methanol concentrations to find the conditions under which the minor enantiomers could be adequately quantitated in samples obtained by chemical resolution of the racemic mixtures. This work extends the utility of ODAS-gammaCD to a hitherto unexplored field, the capillary electrophoretic separation of the enantiomers of organophosphorus compounds. PMID- 11105869 TI - Separation selectivity of anionic metal complexes of N,N' bis(hydroxybenzyl)ethylenediamine-N,N'-diacetic acid in ion and ion electrokinetic chromatography. AB - The complexes of Fe(III), Co(III), Mn(III), Al(III), Cu(II), Ni(II), Cd(II) and Zn(II) with N,N'-bis(hydroxybenzyl)ethylenediamine-N,N'-diacetic acid (HBED) were separated by ion exchange in different modes: ion chromatography (IC) and ion electrokinetic chromatography (IEKC). In column IC these complexes were separated on an IonPac AS4a anion-exchange column (Dionex, USA). Parameters of the background electrolyte that were examined in IEKC mode include polymer, competing ion concentration and pH. The use of poly(diallyldimethylammonium chloride) (PDADMACl) as a modifier in IEKC provides separation selectivity only slightly different from that observed in IC on the IonPac AS4a column. Optimal separation conditions were found to be: 0.1 mM HBED, 50 mM PDADMAOH, 10 mM Na2 B4 O7, pH adjusted to 10 with acetic acid. The use of an aromatic ligand allowed a 10-fold decrease in detection limits of metal ions in comparison with previously studied EDTA. A separation efficiency up to 400,000 theoretical plates was demonstrated for IEKC. PMID- 11105870 TI - Separation and direct UV detection of lanthanides complexed with cupferron by capillary electrophoresis. AB - Separation and detection of lanthanides by capillary zone electrophoresis in the presence of cupferron (N-nitroso-N-phenylhydroxylamine) as UV absorbing complexing agent were investigated. The resolution of partially complexed positively charged cupferron complexes is improved by using a buffer ligand competing with cupferron for metal ions. When hydroxyisobutyric acid (HIBA) is used as buffer and competing ligand, it provides complete separation of all 14 lanthanides with good peak shapes. An on-column separation of 14 lanthanides was achieved in only 7 min using 0.1 mmol/l cupferron, 15 mmol/1 HIBA at pH 4.9. The separation efficiencies for the optimum separation condition are between 77,000 and 208,000 theoretical plates. Determination of lanthanide complexes was performed by direct UV detection at 210 nm. Detection limits (signal-to-noise ratio=3) are ca. 0.24-0.47 microg/ml for lanthanides. Under optimum conditions, the complete separation of thorium and uranium from mixed lanthanides was achieved. PMID- 11105871 TI - Simultaneous determination of inorganic anions and cations by capillary electrophoresis with indirect UV detection. AB - The development of a separation system for the simultaneous determination of inorganic anions and cations, low-molecular-mass organic acids and aliphatic amines by capillary electrophoresis with indirect UV detection using new electrolyte systems is described. Different principles of the experimental enforcement are compared. The principle of both-side injection was investigated using two different electrolyte systems. In order to avoid system peaks caused by the presence of different electrolyte co-ions, the selection of useful electrolyte components is more difficult than the choice of electrolytes for separate anion or cation analysis and special preparation procedures are necessary. The applicability of the method is shown by investigations of reproducibility, linearity of the calibration and by the analysis of drinking water including a comparison with results of measurements carried out with atomic absorption spectrometry for the cation determination, and ion chromatography for the anion determination. PMID- 11105872 TI - Analysis of enzymatically glucosylated flavonoids by capillary electrophoresis. AB - HPCE with UV detection was applied to the analyses of enzymatically glucosylated flavonoids, which are used as natural food additives in Japan. Four items, which have flavonol or flavanone as aglycone, were analyzed. Each of these items is a mixture of glycosides with various lengths of maltooligosaccharide chain. On capillary zone electrophoresis with an untreated fused-silica capillary at alkaline pH, glycosides with longer sugar chains migrated more rapidly. Flavonol glycosides with 1-13 glucose units were distinguished with the borate buffer (pH 10.0). Flavanone glycosides needed higher pH values for good separation than flavonol glycosides. PMID- 11105873 TI - Effects of amine modifiers on the separation of tetramethylrhodamine-labeled mono and oligosaccharides by capillary zone electrophoresis. AB - In this work, nine tetramethylrhodamine (TMR) labeled isomeric oligosaccharide derivatives of betaGal(1 --> 4) betaGlcNAc-O-TMR were separated by capillary zone electrophoresis coupled with laser-induced fluorescence detection. Charged species were created in situ by complexation with borate and phenylborate. Micellar separation was achieved by addition of 10 mM sodium dodecylsulfate to the running buffer. We have investigated the effects of adding a homologous series of monoamine modifiers on the separation efficiency of these oligosaccharides. The separation was significantly improved in the presence of the organic modifiers methyl- and ethylamines, but worsened in the presence of propyl- and butylamines. Possible mechanisms of the amine additives are discussed. PMID- 11105874 TI - Capillary electrophoresis and off-line capillary electrophoresis-electrospray ionization quadrupole time-of-flight tandem mass spectrometry of carbohydrates. AB - The use of off-line high-performance capillary electrophoresis in connection with nanospray electrospray ionization quadrupole time-of-flight tandem mass spectrometry for identification of complex carbohydrates of biological origin is presented. The method was applied to the identification of O-glycosylated amino acids and -glycopeptides from the urine of patients suffering from a hereditary disease - N-acetylhexosaminidase deficiency. Structural elements typical for O glycosylation of proteins, like expression of core 1 and 2 type O-glycans with different numbers of N-acetyllactosaminyl repeats and different degrees of sialylation, can be directly detected. PMID- 11105875 TI - Determination of optical purity of phosphonic acid analogues of aromatic amino acids by capillary electrophoresis with alpha-cyclodextrin. AB - A simple and efficient method for the determination of enantiomeric purity of structurally diverse phosphonic and phosphinic acid analogues of phenylalanine and phenylglycine using capillary electrophoresis is presented. These preliminary studies indicated that the enantiomer separation is strongly dependent on the structure of the aminophosphonic acid. PMID- 11105876 TI - Analysis of carnitine and acylcarnitines in urine by capillary electrophoresis. AB - A capillary electrophoresis method is described for the simultaneous analysis of carnitine and short-chain acylcarnitines in aqueous standard solutions and urine samples. Samples were worked up using silica gel extraction and derivatization with 4'-bromophenacyl trifluoromethanesulfonate. Separation was performed in less than 8 min using a binary buffer system containing phosphate/phosphoric acid and sodium dodecyl sulfate. 3-(2,2,2-Trimethylhydrazinium)propionate (mildronate) was used as an internal standard. The method was developed with aqueous standard solutions and then applied successfully to spiked and unspiked human urine samples. The limit of detection for both carnitine and acetylcamitine is 3 microM. PMID- 11105877 TI - Determination of urinary catecholamines with capillary electrophoresis after solid-phase extraction. AB - The stabilities of 3,4-dihydroxybenzylamine (DHBA), dopamine, 3-methoxytyramine, normetanephrine and metanephrine standards under acid, base and enzymatic hydrolysis conditions were studied. Basic incubation media were not suitable for 3,4-dihydroxy compounds, but acid and enzymatic hydrolysis conditions were applicable to all the compounds. The results of acid and enzymatic hydrolysis were comparable and the enzymatic hydrolysis was applied to a urine matrix. A method including solid-phase extraction (SPE) with a copolymer sorbent was developed for purification of the urine samples. Due to poor recovery of DHBA, the most frequently used internal standard in catecholamine analysis, this compound was replaced with the 3-O-methoxy structure. The recoveries of the compounds in spiked urine samples in SPE were between 96.4 and 124.4%. The repeatability of the combination of enzymatic hydrolysis and SPE pretreatment was good for all the compounds, except for dopamine and 3-methoxytyramine due to some matrix compounds still interfering with the separation. The analyses were performed with capillary electrophoresis in an ammonium acetate buffer with UV detection. The validation data for the compounds including limit of detection, limit of quantification, linearity and repeatability of the method are presented. PMID- 11105878 TI - Capillary zone electrophoresis of a recombinant adenovirus. AB - Adenovirus preparations are used as vectors in a number of gene therapy clinical development programs. The success of commercial production of adenovirus will strongly depend on the development of methods to define the recombinant virus product by analysis as opposed to being defined by the manufacturing process. While most analytical techniques examine portions of the virus, e.g. proteins or DNA, ion-exchange chromatography has been used to separate intact virus at low efficiency. A free zone capillary electrophoretic method was developed for high efficiency separations of adenovirus 5. Experimental conditions such as buffer pH and concentration were explored which produced a high-efficiency separation in less than 20 min. The virus band was identified by collection of CE fractions and examination using a cell based assay. Initially, a single virus peak is found in fresh virus samples. After as little as one freeze-thaw in 1 x phosphate-buffered saline with 2% sucrose, the active virus migrates as a regular series of peaks. The nature of the virus modification leading to the differing electrophoretic mobilities is presently under investigation. PMID- 11105879 TI - Sodium dodecylsulfate capillary gel electrophoretic measurement of the concentration ratios of albumin and alpha2-macroglobulin in cerebrospinal fluid and serum of patients with neurological disorders. AB - Sodium dodecylsulfate capillary gel electrophoresis (SDS-CGE) was applied to measure the concentration ratios of albumin (Alb) and alpha2-macroglobulin (alphaMG) in the cerebrospinal fluid (CSF) and concurrent serum samples from patients with various neurological disorders. The values of the alphaMG index in individual patients were calculated on the basis of the peak area ratios of Alb and an alphaMG subunit on the CSF and serum electropherograms. The alphaMG index value thus obtained was most prominently raised in patients with inflammatory diseases of the brain and/or meninges, suggesting that the function of the blood brain barrier (BBB) was disturbed under the pathological conditions in the central nervous system. The measurement of the concentration ratios of Alb and alphaMG in CSF and the concurrent serum samples by the present SDS-CGE system seems to be useful as an aid in the biochemical examination of the BBB function in patients with neurological disorders. PMID- 11105880 TI - Highly reproducible capillary zone electrophoresis of humic acids in cyclodextrin or oligosaccharide-modified background electrolytes. AB - Capillary zone electrophoresis has been used for the characterization and separation of humic acids. It was found that addition of saccharides like alpha-, beta-, gamma-cyclodextrins, maltose, hydroxyethylcellulose or dextran sulfate in the background electrolyte (50 mM Na2 B4 O7, pH 9.6) yields better separation patterns and highly reproducible electropherograms. Electropherograms with higher numbers of peaks and high reproducibility were obtained with alpha- and beta cyclodextrins or with a mixture of alpha- + gamma-cyclodextrin-modified background electrolytes. Separation was carried out with the cathode at the detector end of the column. Adsorption of humic acids to the capillary wall was diminished using an epoxy-coated capillary tube. PMID- 11105881 TI - Kidney transplantation in dogs with naturally occurring end-stage renal disease. PMID- 11105882 TI - Veterinary ethics and animal welfare. PMID- 11105883 TI - Dolasetron: a new option for nausea and vomiting. PMID- 11105884 TI - Recurrent and persistent urinary tract infections in dogs: 383 cases (1969-1995). AB - Laboratory records of bacterial urine cultures from 383 dogs with recurrent or persistent urinary tract infections (UTI) diagnosed at the University of California Veterinary Medical Teaching Hospital (VMTH) between 1969 and 1995 were reviewed retrospectively to characterize the bacteria involved and their association with age, gender, and breed of dogs affected. Sixty-eight breeds and a mixed-breed group were represented. Escherichia coli was the most common isolate, although mixed-bacterial infections were seen in 58% of the female and 55% of the male dogs. Recurrent and persistent UTI were most prevalent in middle aged to older German shepherd dogs, miniature/toy poodles, and Labrador retrievers, with no apparent sex predilection. Criteria fitting recurrent and persistent UTI were present in 0.3% of all dogs seen at the VMTH during this 26 year period. PMID- 11105885 TI - Feline cytauxzoonosis: a case report and literature review. AB - A 5.5-year-old Siamese presented for evaluation of a three-day history of anorexia and lethargy. Upon physical examination, the cat was depressed, dehydrated, pyrexic, had injected conjunctiva and sclera, pale mucous membranes, and a grade II/VI systolic heart murmur. Thoracic radiographs revealed moderate to severe, diffuse, bronchointerstitial pulmonary changes with enlarged and tortuous pulmonary vessels. With continued hospitalization, the cat became dyspneic and died. The postmortem cytopathological examination of the liver, spleen, and lung impressions revealed reticuloendothelial cell infection with Cytauxzoon felis. PMID- 11105886 TI - Arterial blood pressure measurement in a population of healthy geriatric dogs. AB - The purpose of this study was to evaluate healthy geriatric dogs for the presence of systemic hypertension. Thirty-three geriatric dogs (i.e., dogs exceeding the geriatric age range for their weight group) and 22 control dogs (i.e., dogs less than six years of age) were evaluated by measuring blood pressure with an oscillometric monitor. Five consecutive blood pressure measurements were taken in each dog, averaged, and compared. Diastolic and mean blood pressure measurements were significantly lower in the geriatric group as compared to the control group. Systolic blood pressure measurements were not significantly different between the two groups. Systemic hypertension does not appear to be a common clinical problem in the healthy geriatric dog. PMID- 11105887 TI - Metastatic digital carcinoma in the cat: a retrospective study of 36 cats (1992 1998). AB - Thirty-six cats with bronchogenic carcinoma metastatic to the digit were identified. The mean age was 12.7 years, with no breed or sex predilection. Records from 19 cases were available for review. These cats presented with a primary complaint of lameness that involved primarily weight-bearing digits and the third phalanx. None had respiratory signs, despite the presence of pulmonary carcinoma. Course of disease was consistent in the 19 cases, with a mean survival time of 58 days from initial presentation. Amputation of affected digits was rarely palliative due to development of lesions in other digits and progressive nonrespiratory disease. PMID- 11105888 TI - Hyperadrenocorticism and hyperprogesteronemia in a cat with an adrenocortical adenocarcinoma. AB - A seven-year-old, neutered male domestic shorthair cat was evaluated for poorly regulated diabetes mellitus and increased skin fragility. Imaging studies revealed a right adrenal gland tumor, but cortisol testing did not support a diagnosis of hyperadrenocorticism. Serum concentrations of progesterone and testosterone were increased compared with a group of normal cats, and the clinical signs were attributed to hyperprogesteronemia. At necropsy, a diagnosis of adrenocortical adenocarcinoma was confirmed, and immunohistochemical staining confirmed the presence of progesterone within the tumor. Clinical signs of hyperadrenocorticism in cats may occur due to increased serum concentrations of hormones other than cortisol. PMID- 11105889 TI - Feline osteosarcoma: 145 cases (1990-1995). AB - Feline osteosarcoma (OSA) is a rare tumor in cats. Ninety (62%) of feline OSAs detailed in this study arose from the skeleton, and 55 (38%) arose from extraskeletal sites. Fifty OSAs originated in the appendicular skeleton, and 40 OSAs originated in the axial skeleton. Extraskeletal OSA sites included subcutaneous sites (n=44), with an apparent prevalence for sites commonly used for vaccination. Other locations included ocular/orbital (n=4), oral (n=3), intestinal/omental (n=3), and mammary sites (n=1). Survival data was available for 74 cases. When considered as a group, cats with either appendicular (mean, 11.8 mos) or extraskeletal (mean, 12.67 mos) OSA survived longer than those with axial (mean, 6.07 mos) OSA. Regardless of the type of feline OSA, aggressive surgical excision with or without ancillary therapy appeared to be the most effective form of treatment. PMID- 11105890 TI - Histopathological, radiographic, and arthrographic comparison of the biceps tendon in normal dogs and dogs with biceps tenosynovitis. AB - In dogs surgically treated for biceps tenosynovitis, the most common histopathological findings were fibrosis and collagen degeneration (n=13), synovial villous or vascular hyperplasia (n=10), lymphocytic-plasmacytic infiltrates (n=10), cartilaginous metaplasia (n=8), and ischemic necrosis (n=5). Degree of histopathological changes was associated with degree (p equals 0.000), but not duration (p equals 0.543), of lameness. Furthermore, there was no association between histopathological changes and age or radiographic and arthrographic findings. Cartilage metaplasia was the only histopathological finding in both affected tendons (8/18) and normal control dogs (13/13). Age and size of the control dogs were not determined; however, since all these dogs were clinically normal, fibrocartilaginous metaplasia can be present as an incidental finding in the biceps tendon of origin in dogs. PMID- 11105891 TI - Case examples demonstrating the clinical utility of obtaining both right and left lateral abdominal radiographs in small animals. AB - Seven case examples are presented which illustrate the utility of using both right to left and left to right laterolateral abdominal radiographs (left lateral and right lateral recumbent radiographs, respectively) when evaluating gastrointestinal disease. These cases demonstrate the benefits of obtaining both lateral abdominal radiographs in disease of the stomach, small intestine, and large intestine. A review of the literature concerning this technique is provided. PMID- 11105892 TI - Compression radiography: an old technique revisited. AB - Five case examples are provided to illustrate the use of compression radiography in the dog and cat. Abdominal compression radiography provides for evaluation of the size, shape, location, and opacity of a specific area without the degree of superimposition encountered with survey radiographs. The following cases illustrate compression techniques of the intestinal tract, uterus, kidney, bladder, and spleen. A review of the technique as well as a discussion of the advantages and limitations are presented. PMID- 11105893 TI - Use of skin staples for rapid closure of gastrointestinal incisions in the treatment of canine linear foreign bodies. AB - A technique was developed for closure of gastrotomy and enterotomy incisions using disposable skin staples. The technique was used successfully in three dogs with gastrointestinal linear foreign bodies that required a gastrotomy and one or more enterotomies. The method allows for secure closure of gastrointestinal incisions and minimizes the surgical time in patients requiring multiple gastrointestinal incisions. PMID- 11105894 TI - Retrospective evaluation of occlusion of patent ductus arteriosus with hemoclips in 20 dogs. AB - Twenty dogs with patent ductus arteriosus occluded with Hemoclips were evaluated with a mean follow-up time of 799.4 days (range, 83 to 3,580 days). Significant decreases were found between pre- and postsurgical means for vertebral heart size and for echocardiographic left atrial/aortic-root ratios and percent fractional shortening (%FS). Despite a good clinical outcome, six of 20 dogs had persistent cardiomyopathy of overload with diminished %FS (28% or less) at follow-up. One dog had residual ductal flow identified five days postoperatively. Subsequent evaluations in this case at 60, 144, 226, 344, and 560 days postoperatively demonstrated gradually diminishing ductal flow. The remaining 19 dogs did not recanalize. PMID- 11105895 TI - Shrinkage in the horizontal dimensions of the vulva (vulvar shrinkage) as an indicator of standing heat in the beagle. AB - During the proestrous and estrous periods in 12 beagles, the following parameters were measured daily: the horizontal dimensions of the vulva, vaginal cytology, and serum luteinizing hormone (LH) and progesterone concentrations. Measurements of serum LH concentrations allowed for the identification of the LH surge and the optimal time for artificial insemination (AI). Nine out of the 12 beagles became pregnant through AI and completed a gestation. Shrinkage in the horizontal dimensions of the vulva (i.e., vulvar shrinkage) was primarily observed prior and subsequent to the LH surge. In six of the nine (66.7%) beagles that became pregnant, vulvar shrinkage was observed on the days in which the LH surge was confirmed, and the rate of vulvar shrinkage tended to be greater at higher serum LH concentrations. Further vulvar shrinkage was identified in all nine beagles within two days of the LH surge. An increase in the serum progesterone concentration was observed after the LH surge in each of the beagles that became pregnant, together with clinical signs of estrous behavior (i.e., standing heat) as well as a change in vulva condition from swollen to soft. This demonstrates that vulvar shrinkage is induced in response to the onset of the LH surge and that the LH surge can be predicted through the measurement of the horizontal dimensions of the vulva, vaginal cytology, and the assessment of serum progesterone concentrations in beagles. PMID- 11105896 TI - Runt and Lozenge function in Drosophila development. AB - Runt and Lozenge (LZ) are members of the Runt domain family of transcriptional regulators and control a large number of developmental processes in Drosophila. Runt is a pair-rule gene, and is part of the network of genes that control pattern formation in the embryo. In the central nervous system, Runt function is necessary for the development of a subset of neurons. Runt is also a key regulator of sex determination, and directly controls Sex-lethal, a master gene that determines sex of the animal and controls dosage compensation. The LZ protein also participates in several key processes. LZ controls pre-patterning and cell-fate choices in the development of the visual system by regulating the expression of several fate-specifying transcription factors, and works in conjunction with general signaling pathways. LZ function is also required in hematopoiesis for the specification of a Drosophila blood cell lineage. PMID- 11105897 TI - Potential roles for RUNX1 and its orthologs in determining hematopoietic cell fate. AB - Runx1 (also known as AML1, Cbfa2 and Pebpa2b) and Cbfb encode a DNA-binding alpha subunit and the non-DNA-binding beta subunit of a mammalian core-binding factor (CBF). The discovery of RUNX1 and CBFB as genes rearranged in human leukemias prompted predictions that both genes would play important roles in normal hematopoiesis. These predictions were borne out, as indeed Runx1 and its Xenopus and Drosophila homologs, Xaml and lozenge (lz), appear to determine hematopoietic cell fate during development. We will review what is known about Runx1 function in hematopoiesis in three model organisms, mouse, frog and fly, focusing on the earliest events of hematopoietic cell emergence in the embryo. PMID- 11105898 TI - Role of Cbfa1 in osteoblast differentiation and function. AB - Among the multiple cell lineages whose differentiation is affected by a runt related gene the osteoblast is a relative newcomer. Molecular biology, developmental biology and mouse and human genetic studies have demonstrated that Cbfa1 is a critical regulator of osteoblast differentiation in vertebrates. Cbfa1 is not only a differentiation factor but also a regulator of bone formation by differentiated osteoblasts beyond development. Thus, Cbfa1 controls osteogenesis at multiple stages. PMID- 11105899 TI - Alterations of the AML1 transcription factor in human leukemia. AB - The identification of clonal chromosomal translocations in human leukemias provided one of the first insights into the underlying pathogenesis of this clinically heterogeneous disease. Over the last decade a large number of these chromosomal rearrangements have been molecularly cloned and the involved genes identified. A surprising finding that has emerged from this work is that many of these chromosomal alterations target the genes encoding the AML1/CBFbeta transcription factor complex, a critical regulator of normal hematopoiesis. In this review, we summarize our present understanding of the mechanisms through which alterations of AML1/CBFbeta contribute to leukemogenesis. PMID- 11105900 TI - Function of CBFbeta/Bro proteins. AB - Mammalian core binding factor beta (CBFbeta) and Drosophila Brother (Bro) and Big brother (Bgb) proteins are transcription factors that dimerize with mammalian Runx and Drosophila Runt and Lozenge proteins and augment their DNA binding affinity and transcriptional potency. CBFbeta is essential for development and sustenance of definitive hematopoiesis during mouse embryogenesis. Bro and Bgb are required for Runt/Lozenge functions in Drosophila development. CBFbeta contributes to leukemogenesis since the CBFB gene is specifically and consistently mutated by a chromosome 16 inversion found in patients with acute myeloid leukemia subtype M4Eo. The ubiquitous expression pattern of the CBFB gene suggests that it may play important roles in many other organ systems. PMID- 11105901 TI - Mechanisms of transcriptional regulation by Runt domain proteins. AB - Runt domain proteins have vital roles in regulating transcription in developmental pathways extending from sex determination and segmentation in fruit fly embryos to the development of blood and bone in mammals. Many of the insights into the mechanisms by which these proteins act to regulate transcription originate either from studies on the Drosophila runt gene, the founding member of this family, or from work on the mammalian PEBP2/CBF transcription factor. Genetic experiments in the Drosophila system reveal that runt functions both to activate and to repress transcription of different downstream target genes and indicate that different mechanisms are used in the regulation of different specific downstream target genes. These studies have also identified other nuclear factors that work with Runt in some of these pathways. Studies in mammalian systems have provided additional evidence for the complexity of transcriptional regulation by Runt domain proteins and have identified other transcription factors that cooperate with Runt domain proteins to regulate the activity of different specific cis-regulatory enhancers. The emerging view from studies in both systems is that these proteins act as context-dependent regulators of transcription, activating or repressing gene expression dependent upon the constititution of a particular promoter/enhancer in a particular cell type. These results have yielded new insights into the molecular mechanisms that control animal development and provide a framework for investigating fundamental issues in eukaryotic transcriptional regulation. PMID- 11105902 TI - CBF--a biophysical perspective. AB - Core binding factor (CBF) is a heterodimeric transcription factor consisting of a DNA-binding subunit (Runx, also referred to as CBFA, AML 1, PEBP2alpha) and a non DNA-binding subunit (CBFB). Biophysical characterization of the two proteins (and their interactions is providing a detailed understanding of this important transcription factor at the molecular level. Measurements of the relevant binding constants are helping to elucidate the mechanism of leukemogenesis associated with altered forms of these proteins. Determination of the 3D structures of CBFB and the DNA- and CBFB-binding domain of Runx, referred to as the Runt domain, are providing a structural basis for the functioning of the two proteins of CBF. PMID- 11105903 TI - Targeted disruption of the mouse prosaposin gene affects the development of the prostate gland and other male reproductive organs. AB - The prosaposin gene encodes a 65-70 kilodalton (kd) protein, which is secreted or targeted to lysosomes. In lysosomes, prosaposin is the precursor of 4 activator proteins, designated saposins A, B, C, and D, which promote by acidic hydrolases, the degradation of glycosphingolipids with short oligosaccharide chains. Mutations of the prosaposin gene have been linked to several lysosomal storage disorders. An animal model was recently developed by creating a null allele in embryonic stem cells through gene targeting in order to investigate the phenotypic diversity of prosaposin mutations, the involvement of this protein in lysosomal storage diseases, and to develop potential therapeutic approaches. Mutant homozygous mice die at 35-40 days of age and neurological disorders contribute to their early death. Secreted prosaposin is present in milk and in cerebrospinal and seminal fluids. In the nervous system, prosaposin exhibits a trophic activity. Examination of reproduc-tive organs in homozygous mutant males shows several abnormalities such as a decrease in testis size with reduced spermiogenesis, and an involution of the prostate, seminal vesicle, and epididymis, although levels of testosterone in blood remain normal. In the prostate of homozygous mutants, only basal cells appear to be present, whereas secretory cells are absent. The epithelia in efferent ducts is formed by ciliated cells, whereas heterozygotes exhibit a majority of nonciliated cells. Our data indicate that prosaposin is involved in the development and maintenance of male reproductive organs. In prostatic epithelium, targeted disruption of the prosaposin gene appears to inactivate the mitogen-activated protein kinase pathway and to interfere with differentiation of secretory cells. PMID- 11105904 TI - All you wanted to know about spermatogonia but were afraid to ask. PMID- 11105905 TI - Phenotypes of sperm pathology: genetic and acquired forms in infertile men. PMID- 11105906 TI - Adults with cystic fibrosis and (in)fertility: how has the health care system responded? PMID- 11105907 TI - Clinical and laboratory management of male infertility: an opinion on its current status. PMID- 11105908 TI - Mouse Spam1 (PH-20): evidence for its expression in the epididymis and for a new category of spermatogenic-expressed genes. AB - The gene for the sperm adhesion molecule 1 (PH-20), SPAM1, has been known to be testis-specific and exclusively haploid expressed. We show that in mice, the 2 common isoforms of the protein (Spam1) observed in sperm are also present in the caput, corpus, and cauda epididymides. Both qualitative and quantitative variation of expression of the protein were observed in epididymis with the highest expression detected in the corpus. The endogenous production of enzymatically active (via hyaluronidase) Spam1 by epididymal cells is supported by the detection of steady-state Spam1 epididymal messenger RNA in both wild type and germ cell-deficient mice. In situ transcript hybridization shows the transcript to be localized to the principal cells of the epithelium. The protein was similarly immunolocalized to these cells, predominantly in vesicles near the apical region. The results suggest a mechanism for transportation of Spam1 from the epididymal epithelium to sperm during their transit and storage in the cauda. None of the current categories of spermatogenic-expressed genes shows the dual transcription pattern (haploid testicular/diploid epididymal) observed for Spam1. The work also confirms and extends the finding that Spam1 is expressed in the kidney. PMID- 11105909 TI - Effects of long-term administration of androgens and estrogen on rhesus monkey prostate: possible induction of benign prostatic hyperplasia. AB - Rhesus monkeys were used to investigate the role of androgenic steroids and estradiol in the induction of hyperplastic changes in stromal and glandular prostate tissues. Adult male rhesus monkeys were procured from the wild and, after routine quarantine procedures, were randomly divided into 5 groups of 5 animals each. Gluteus maximus muscles were injected with 2.5 mg of androstenedione (Group II), 2.5 mg of dihydrotestosterone (DHT) or 0.25 mg of estradiol (Group II), 2.5 mg androstanediol (Diol; Group IV), or Diol in combination with 0.25 mg of estradiol (Group V). Group I consisted of untreated controls. Animals were injected with steroids 3 times a week for 2 years. Treatment with androstenedione (Group II) resulted in stromal hyperplasia in the caudal lobe and an increase in epithelial cell height in all zones except in the central zone of the caudal lobe. In monkeys treated with DHT and estradiol (Group III), stromal hyperplasia in both lobes, a decrease in tubular size, and degranulation and vacuolation of epithelial cells were noticed. Injection of Diol alone (Group IV) or in combination with estradiol (Group V) resulted in a widening of stroma in the central and peripheral zones of cranial and caudal lobes, whereas the tubular size decreased. Diol also induced epithelial cell hypercellularity in the central and peripheral zones of the caudal lobe and in the peripheral zone of the cranial lobe. Prostate-specific antigen levels in Group IV animals gradually increased from 6 months of treatment and were maximal after 18 months of injections. Serum estradiol levels increased to detectable levels in all groups except Group IV. Serum testosterone levels decreased to very low or undetectable levels in all groups, whereas prostate-specific acid phosphatase increased in all treated groups. Prolactin levels were elevated in all treated groups except in animals injected with androstenedione. These results indicate that repeated long-term injections of androstenedione or DHT and estradiol induced stromal hyperplasia, which may be an estrogen-related effect. Androstanediol-induced hypercellularity and stratification of glandular epithelium is comparable to human prostatic intraepithelial neoplasia. These results also suggest that the rhesus monkey is a suitable animal model for experimental induction of prostate diseases. PMID- 11105910 TI - Epididymal epithelial cells cultured in vitro prolong the motility of bovine sperm. AB - It is well known that the epididymis is an excellent environment to maintain sperm viability. Therefore, we used different sections of bovine epididymis (caput, corpus, and cauda) to develop epithelial cell culture monolayers to identify factors that will increase sperm survival in the freezing-thawing process. Each epididymal section was dissected and treated with collagenase to obtain epithelial cell clusters. The cells were cultured in RPMI-1640 medium with 10% serum at 38.5 degrees C. A confluent monolayer was obtained after 5-7 days in culture and preliminary characterization using cytokeratin antibody indicated that the cell culture contained 85%-95% of epithelial cells. These cellular cultures were tested for their ability to maintain motility of epididymal and frozen-thawed spermatozoa. Washed spermatozoa were added to obtain a final dilution of 1 x 10(6) spermatozoa/mL. The motility of frozen-thawed spermatozoa was also recorded after incubation in conditioned media. Our results show that cocultures of spermatozoa and epididymal cell monolayers for 24 and 48 hours were beneficial for maintaining epididymal and frozen-thawed sperm motility (36.0% and 20.4%) compared with spermatozoa cultured with fibroblast cells or in the absence of a cell monolayer (0%; P < .01). The conditioned medium provides favorable conditions for sperm motility. Results with conditioned medium on maintenance of frozen-thawed sperm motility suggest that epididymal cells in vitro secrete beneficial factors that prolong the sperm survival. PMID- 11105911 TI - Stage and region-specific localization of lipocalin-type prostaglandin D synthase in the adult murine testis and epididymis. AB - Lipocalin-type prostaglandin D synthase in semen has been associated with male fertility, although this relationship is not well defined. To gain insight into potential mechanisms, the objective of the present study was to immunocytochemically localize lipocalin-type prostaglandin D synthase within the testis, efferent ducts, and 4 segments of mouse epididymis. In the testis, immunoperoxidase staining was localized within the Sertoli cells only at stages VI-VIII of the spermatogenic cycle, which is just prior to spermiation. Intense staining was also evident throughout the interstitial tissue, including Leydig cells. The entire epithelium of the efferent ducts, including ciliated and nonciliated cells, was immunoreactive. A distinct pattern of immunostaining for lipocalin-type prostaglandin D synthase was observed in different regions of epididymis, suggesting a possible role in sperm maturation. Staining for lipocalin-type prostaglandin D synthase was strikingly absent in the initial segment. In caput epididymidis, staining was evident throughout the cell cytoplasm of principal cells with some cells more intensely stained than adjacent ones. In the corpus region, overall staining intensity decreased and appeared to be concentrated in the apical region of principal cells, but some cells were completely unreactive. Reaction product in the cauda region was heavily concentrated on microvilli and within the epididymal lumen. In all epididymal regions, expression of lipocalin-type prostaglandin D synthase was specific to epithelial principal cells; no immunoreactivity was apparent in other cell types. The specific localization of lipocalin-type prostaglandin D synthase within the testicular interstitial tissue, Sertoli cells, and principal cells of caput epididymidis strongly suggests that this protein plays an integral role in both the development and maturation of sperm. PMID- 11105912 TI - Interleukin-1beta regulates nitric oxide production and gamma-glutamyl transpeptidase activity in sertoli cells. AB - Several cytokines have been involved in the regulation of Sertoli cell function. Further investigations are required to elucidate the role of interleukin-1beta (IL1beta) in Sertoli cell physiology. Twenty-day-old rat Sertoli cell cultures were used to investigate a possible role of IL1beta in the regulation of gamma glutamyl transpeptidase (gammaGTP) and to elucidate the signaling pathway utilized by this cytokine. GammaGTP is a membrane-bound enzyme that has been involved in amino acid transport across the plasma membrane and in protection from oxidative stress through its importance in the regulation of glutathione levels. Previous studies suggested that IL1beta stimulates NO biosynthesis in other cell types. Therefore, we investigated whether IL1beta modified the level of nitrite, a stable metabolite of NO, in Sertoli cells. Dose-response curves to IL1beta for gammaGTP activity and nitrite production were observed. The increments observed in gammaGTP activity and nitrite production were partially and completely blocked by simultaneous treatment with the NO synthase inhibitor aminoguanidine. Treatment of Sertoli cell cultures with the NO donors sodium nitroprusside and S-nitroso-N-acetylpenicillamine resulted in an increase in gammaGTP activity. The presence of neural, endothelial, and inducible isoforms of NO synthase (NOS) was investigated by a immunohistochemical technique using specific antibodies. The 2 constitutive isoforms were present under basal conditions, and the inducible protein appeared in IL1beta-treated cultures. Finally, translocation of NF-kappaB p65 subunit to the nucleus in IL1beta-treated cultures was observed. These findings suggest that the action of IL1beta on Sertoli cell gammaGTP activity is partially mediated via activation of NF-kappaB and increments in iNOS and cellular production of NO. PMID- 11105913 TI - Evaluation of poly(styrene-4-sulfonate) as a preventive agent for conception and sexually transmitted diseases. AB - A commercial preparation of a sodium polystyrene sulfonate (designated as N-PSS; its molecular weight is 500000 daltons) was tested as an inhibitor of sperm function and as a preventive agent for conception and the transmission of sexually transmitted diseases. The polymer is an irreversible inhibitor of hyaluronidase and acrosin; its IC50 values are 5.7 microg/mL and 0.5 microg/mL, for hyaluronidase and acrosin, respectively. N-PSS is also a stimulus of human sperm acrosomal loss. It produces maximal acrosomal loss at 2.5 microg/mL. Contraception in rabbits is nearly complete when rabbit spermatozoa are pretreated with 0.5 mg/mL of N-PSS before artificial insemination; however, N-PSS does not immobilize spermatozoa at concentrations as high as 50 mg/mL. N-PSS has broad spectrum antiviral and antibacterial activities. Infection by human immunodeficiency virus and herpes simplex virus are inhibited by N-PSS; 3-log reductions are produced by 7 microg/mL and 3 microg/mL, respectively. N-PSS is active against Chlamydia trachomatis and Neisseria gonorrhoeae. At 1 mg/mL, N-PSS inhibits chlamydial infectivity by more than 90%. N-PSS produces a 3-log reduction in gonococcal growth at 15 microg/mL. In contrast, N-PSS (5 mg/mL) does not affect the growth of Lactobacillus (normal component of the vaginal flora). N PSS can be classified as a noncytotoxic contraceptive antimicrobial agent. These properties justify bringing a polystyrene sulfonate into clinical trials for its evaluation as a preventive agent for conception and several sexually transmitted diseases. PMID- 11105914 TI - Addition of specific metabolites to bovine epididymal cell culture medium enhances survival and motility of cryopreserved sperm. AB - We have developed a cell culture system of bovine epididymal epithelium in which cryopreserved bovine sperm motility was efficiently maintained for many hours. The culture conditions to maintain viable epididymal cells are quite different from conditions normally used to incubate sperm cells. Thus, we have modified a previously described principal cell medium (PCM; Moore et al, 1992) using HEPES as a buffer and supplemented media with myo-inositol, pyruvate, lactate, glycerol, and carnitine to mimic epididymal intraluminal conditions. In the first experiments the effects of PCM and our epididymal cell medium (ECM) on sperm motility were compared in the absence of cells and evaluated by microscopic analysis under a phase contrast microscope or using the Hamilton Thorn Image Analyzer System. Our results showed that motility of cauda epididymal sperm was significantly higher in ECM than in PCM during a 48-hour incubation period when both media were supplemented with 10% fetal bovine serum (FBS). We then replaced FBS with bovine serum albumin (BSA) or no proteins at all to verify if ECM was able to enhance sperm survival. To test this aspect we used frozen-thawed sperm, which survived up to 48 hours when sperm cells were coincubated with epididymal cell monolayers. Hence, PCM, ECM, and different media containing each metabolite of ECM were supplemented with 0.5% BSA to assess motility of thawed sperm after an incubation period of 6 hours. A positive effect on sperm motility was observed in all fresh and unconditioned media containing 1 mM pyruvate. Motion parameters were more efficiently maintained in all conditioned media than in unconditioned media. Our results showed, however, that pyruvate was almost completely oxidized or consumed by epididymal cells during preincubation of culture media. We conclude that motility of frozen-thawed bovine spermatozoa can be improved using a culture medium or a medium conditioned by epididymal cell cultures without carnitine but containing mainly pyruvate, inositol, glycerol, and lactate. PMID- 11105915 TI - Correlation between clusterin-positive spermatozoa determined by flow cytometry in bull semen and fertility. AB - The objectives were to 1) develop a rapid and accurate method for detection of clusterin-positive spermatozoa (CPS) in bull semen and 2) determine the utility of incidence of CPS for prediction of fertility of bull semen in comparison to routine semen quality traits. Semen from 3 bulls was immunostained with anti bovine clusterin antibody and with FITC-conjugated anti-rabbit IgG for method development. Clusterin-positive spermatozoa were determined by flow cytometry (FCM) and fluorescence microscopy, and results were compared by paired t test. There was no difference between FCM and microscopic techniques (P = .81). Flow cytometry was then used for determination of CPS in semen of 48 bulls with known fertility. Significant inverse relationships were found between the percentage of CPS and raw nonreturn rate (r = -.30), adjusted nonreturn rate (r = -.58), and estimated relative conception rate (ERCR; r = -.60). Estimated relative conception rate is potentially a very accurate method for determining fertility, and it resulted in highest correlation with CPS. An inverse relationship was observed between the percentage of CPS and prefreeze and postfreeze motility (r = -.51), whereas a direct relationship was found between CPS and primary, secondary, tertiary, and total sperm abnormalities (r = .52, .77, .32, and .58, respectively). The fractions of motile and abnormal spermatozoa, with the exception of tertiary abnormalities, were inversely correlated with 2 or more of the fertility estimates, but none of them showed the characteristic increase in correlation with improvement of accuracy of fertility estimate as demonstrated by CPS. We conclude that FCM is useful for objective and efficient detection of CPS in bull semen. The results suggest that the percentage of CPS in bull semen is potentially a better predictor of fertility than sperm motility or abnormal morphology. PMID- 11105916 TI - The effect of reactive oxygen species on equine sperm motility, viability, acrosomal integrity, mitochondrial membrane potential, and membrane lipid peroxidation. AB - The objective of this study was to examine the influence of reactive oxygen species (ROS), generated through the use of the xanthine (X)-xanthine oxidase (XO) system, on equine sperm motility, viability, acrosomal integrity, mitochondrial membrane potential, and membrane lipid peroxidation. Equine spermatozoa were separated from seminal plasma on a discontinuous Percoll gradient, and spermatozoa were incubated with 0.6 mM X and 0.05 U/mL XO for 30 minutes. Catalase (150 U/mL), superoxide dismutase (SOD, 150 U/mL), or glutathione (GSH, 1.5 mM) were evaluated for their ability to preserve sperm function in the presence of the induced oxidative stress. At the end of the 30 minute incubation, sperm motility was determined by computer-assisted semen analysis. Viability and acrosomal integrity were determined by Hoechst-Pisum sativum staining, and mitochondrial membrane potential was determined by staining with JC-1. Incubation with the X-XO system led to a significant (P < .01) increase in hydrogen peroxide production and an associated decrease (P < .01) in motility parameters. Total motility was significantly (P < .01) lower in the presence of X-XO compared with the case of the control (29%+/-9% vs 73%+/-1%, respectively). Catalase, but not SOD, prevented a decline in motility secondary to oxidative stress (71%+/-4% vs 30%+/-3%, respectively). The addition of glutathione had an intermediate effect in preserving sperm motility at the end of the 30-minute incubation (53%+/-3%). No influence of X-XO could be determined on viability, acrosomal integrity, or mitochondrial membrane potential. In order to promote lipid peroxidation, samples were incubated with ferrous sulfate (0.64 mM) and sodium ascorbate (20 mM) for 2 hours after the X-XO incubation. No increase in membrane lipid peroxidation was detected. This study indicates that hydrogen peroxide is the major ROS responsible for damage to equine spermatozoa. The decrease in sperm motility associated with ROS occurs in the absence of any detectable decrease in viability, acrosomal integrity, or mitochondrial membrane potential or of any detectable increase in lipid peroxidation. PMID- 11105917 TI - Functional and ultrastructural features of DNA-fragmented human sperm. AB - The functional significance of deoxyribonucleic acid (DNA) fragmentation in ejaculated human sperm is unclear. In this study the extent of DNA strand breakage in swim-up selected spermatozoa was evaluated by terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick end labeling (TUNEL) coupled flow cytometry and correlated with several functional and morphological sperm parameters. The extent of DNA fragmentation (mean = 11.07%+/-8.00%, range = 0.79%-42.64%, n = 140) was positively related to abnormal morphology and associated with defects of the sperm tail. A negative correlation was found between DNA breakage and progressive motility. When a stepwise multiple linear regression model was used to analyze the relationship between DNA fragmentation and the aforementioned parameters, only motility results were included in the model. The presence of spermatozoa showing submicroscopic characteristics resembling those of somatic apoptosis has been reported in human ejaculate. To verify whether sperm DNA fragmentation was associated with the presence of such apoptotic-like cells, we performed electron microscopy and TUNEL-coupled flow cytometry in a limited number of sperm samples (n = 24). Although we did not observe any significant relationship between DNA breakage and the characteristics that are suggestive of apoptosis, an association was found with several ultrastructural features, indicating an impaired motility. Hence, we conclude that in ejaculated sperm, DNA fragmentation does not correspond to the apoptosis like phenomenon and that it is associated with defects of motility. PMID- 11105918 TI - Quantification of the nonenzymatic fast and slow TRAP in a postaddition assay in human seminal plasma and the antioxidant contributions of various seminal compounds. AB - Total radical-trapping antioxidant potential (TRAP) measurements of human seminal plasma (N = 25) were performed by using a post-addition assay based on trapping 2,2' Azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radicals. This method enables the antioxidant capacity of human seminal plasma and its constituents to be quantified. The standard procedure consisted of determination of the Trolox equivalent antioxidant capacity (TEAC) after incubating the test sample in the ABTS radical solution for 10 seconds (fast TRAP) and 300 s (total TRAP). Interestingly, seminal plasma showed a fast TRAP and a high slow TRAP (Total TRAP - Fast TRAP). The final total TRAP of seminal plasma is about 10 times higher than that of blood plasma. Various components of seminal plasma contribute to its fast TRAP; 37% can be attributed to vitamin C, uric acid, and tyrosine; proteins and polyphenolic compounds contribute a further 57%. In contrast, the slow TRAP was attributed to vitamin C (1%), uric acid (2%), and tyrosine (15%) and to proteins and polyphenolic compounds (33%). It was not possible to account for the remaining 49%. Neither known putative antioxidants, such as spermine, pyruvate, and taurine, nor other seminal compounds, such as carnitine, sialic acid, fructose, spermidine, glycerophosphorylcholine, and hyaluronic acid, contributed to any significant radical-trapping activity at a standard concentration of 1 mM. Of the amino acids, only tyrosine possessed a slow TRAP, and it is present at a high concentration in seminal plasma. Glutathione and hypotaurine show high fast and slow TRAPs, respectively. However, because of their low concentration in seminal plasma, their contribution to the TRAP is negligible. In conclusion, seminal plasma possesses a high antioxidant buffer capacity that protects spermatozoa from oxidative stress. Moreover, these findings suggest that the fast and slow TRAPs may have an important role as infertility markers and treatment targets in future antioxidant therapies. PMID- 11105919 TI - Separation of ram spermatozoa bearing X and Y chromosome by centrifugal countercurrent distribution in an aqueous two-phase system. AB - The availability of reliable quantification techniques of X and Y chromosome bearing spermatozoa in a given insemination dose would allow further approaches to their separation, which is a topic of unquestionable interest in animal production. The aim of the current work was the development of a combined approach of polymerase chain reaction and countercurrent distribution to address both objectives. First, using Sac/polymorphisms for ZFX/ZFYloci in sheep deoxyribonucleic acid, a linear correlation has been established between the densitometric quantification of the restricted fragment length polymorphisms corresponding to the amplified loci ZFX/ZFY by polymerase chain reaction and the theoretical proportions of X and Y chromosomes in standard solutions. The method, subsequently applied to semen samples, estimated an equal proportion of spermatozoa bearing each chromosome. Second, by using centrifugal countercurrent distribution in a sensitive-charge aqueous two-phase system, we achieved the separation of a sperm population enriched in Y chromosome-bearing ram spermatozoa (75%) with a high viability rate (57%). PMID- 11105920 TI - The cyclic GMP-specific phosphodiesterase inhibitor, sildenafil, stimulates human sperm motility and capacitation but not acrosome reaction. AB - Capacitation is the series of transformations that spermatozoa undergo in the female genital tract in order to bind to the zona pellucida, initiate the acrosome reaction, and fertilize an egg. Cyclic adenosine monophosphate (cAMP) plays an important role in this process and its levels are regulated by 2 key enzymes, adenylyl cyclase and cyclic nucleotide phosphodiesterase (PDE), the latter being involved in cAMP degradation. Evidence was provided for the involvement of PDE in sperm motility and capacitation. Of the 10 gene families of PDE that exist in mammalian tissues, the calcium-calmodulin-dependent (type 1) and the cAMP-specific (type 4) have been found in human spermatozoa. Using sildenafil, we investigated a highly potent cyclic guanosine monophosphate (cGMP) specific PDE (type 5) inhibitor and whether this PDE is present in human spermatozoa and is involved in sperm functions. Sildenafil inhibited PDE activity of Percoll-washed spermatozoa with an IC50 of 97+/-3 and 33+/-3 microM when cAMP and cGMP, respectively, were used as substrates. Because the IC50 of sildenafil obtained for PDE type 5 is much lower (2 to 6 nM) than that obtained with sperm PDE, the data suggest that PDE type 5 represents only a small fraction of the whole PDE activity of spermatozoa. Sildenafil causes dose-dependent increases in sperm cAMP levels and capacitation, which are associated with an increase in the levels of tyrosine phosphorylation of 2 fibrous sheath proteins (p105/81). Sperm velocity, amplitude of lateral head displacement, and hyperactivation were increased at 30-180 minutes. Sildenafil did not trigger the acrosome reaction in capacitated spermatozoa. These results suggest that under our experimental conditions, sildenafil triggers human sperm motility and capacitation, probably via its inhibitory action on PDE activity other than type 5 with a resultant rise in cAMP levels. PMID- 11105921 TI - Sperm calcium levels and chlortetracycline fluorescence patterns are related to the in vivo fertility of cryopreserved bovine semen. AB - Cryopreserved bovine semen is less fertile than fresh semen for reasons that have not been fully elucidated. Cryopreservation is known to disrupt the sperm plasma membrane and it induces premature capacitation of a sperm subpopulation, which may be a result of the increased internal calcium levels after thawing. To test the hypothesis that sperm intracellular calcium level is correlated with in vivo fertility, we used the fluorescent calcium indicator, indo-1, and flow cytometry to assess intracellular calcium levels in frozen-thawed sperm from bulls of varying degrees of fertility. We also tested a second hypothesis that the physiological status of sperm, as assessed by the chlortetracycline (CTC) fluorescent assay, is correlated with fertility. As detected by indo-1 fluorescence, the intracellular calcium level is negatively correlated with bull fertility immediately after thawing (P = .0362; n = 3 ejaculates from each of 10 animals). Moreover, there was a significant difference between the 3 most and least fertile bulls over 4 hours of incubation (P < .05; n = 3 ejaculates per bull). Finally, there was a positive correlation between sperm displaying the CTC acrosome reaction pattern and fertility (P = .0014; n = 3 ejaculates from each of 10 bulls). PMID- 11105923 TI - Expression and cellular localization of contraception-associated protein. AB - Treatment of male rats with ornidazole results in reversible infertility, which is associated with the detection of the contraception-associated protein 1 (CAP1) in epididymal fluid. The protein, which is present in sperm but not detectable in epididymal fluid of fertile rats, seems to be shed from sperm during ornidazole administration. Cloning and characterization of the gene revealed a high degree of similarity between CAP1 and DJ-1 (Wagenfeld et al, 1998b) a protein that was recently found in humans and which has been classified as a novel oncogene. Reverse transcription of total ribonucleic acid (RNA) from various species indicated that a gene similar to CAP1 was also expressed in the testes of hamsters, mice, cynomolgus monkeys, and humans. Detection of RNA expression in rats at the cellular level by in situ hybridization revealed a stage-specific CAP1 expression in the cytoplasm of pachytene spermatocytes (stages IX-XIII), secondary spermatocytes (stage XIV), and round spermatids (stages I-VII). Immunolocalization of CAP1 in rat testis showed a strong staining of elongating spermatids (stages VI-VIII), indicating a translational delay of CAP1 expression. The location of CAP1 on sperm depended on the method of fixation used, with CAP1 being exhibited on the equatorial segment of the sperm head and cytoplasmic droplets. Flow cytometric analysis of sperm from ornidazole-fed rats revealed a significant decline (of 22%-24%) in the amount of sperm surface CAP1 compared with controls, which is associated with an altered location on the sperm head. These observations support a putative role of the protein in the fertilization process. PMID- 11105922 TI - Targeted disruption of the cation-dependent or cation-independent mannose 6 phosphate receptor does not decrease the content of acid glycosidases in the acrosome. AB - The acrosome is a unique organelle containing acid hydrolases common to lysosomes as well as unique enzymes. Its ultimate exocytosis, as well as the absence of several lysosomal markers, has led to the speculation that it should be considered a secretory or zymogen vesicle rather than a specialized lysosome. The basic targeting machinery for eukaryotic lysosomal acid glycosidases are the two mannose 6-phosphate receptors. Mouse testicular germ cells are known to express both the cation-independent (CI-MPR) and cation-dependent (CD-MPR) forms of the mannose 6-phosphate receptors, but the CD-MPR is predominant. In this report, we utilized the recent targeted disruption of the CD-MPR and CI-MPR genes to determine whether these mutations affect targeting of acid glycosidases to the acrosome. Antibody to luminal fluid beta-D-galactosidase was used to examine the targeting of immunoreactive product within the acrosome of permeabilized spermatozoa and testicular spermatids. No obvious changes in acrosomal immunoreactivity in either MPR homozygous mutant were observed when compared with the case of wild-type littermates. In addition, targeted disruption of either MPR did not result in decreased levels of beta-D-galactosidase, alpha-D-mannosidase, or N-acetylglucosaminidase activities in spermatozoa from either MPR-homozygous mutant. These results suggest that the targeted disruption of either MPR does not result in decreased acrosomal targeting efficiency. PMID- 11105924 TI - Toxic effects of polychlorinated biphenyls on cultured rat Sertoli cells. AB - Polychlorinated biphenyls (PCBs) are ubiquitous and persistent environmental contaminants. In mammals, PCBs affect spermatogenesis and may be associated with Sertoli cell changes. Therefore, our aim was to evaluate in vitro toxic effects of hydroxylated PCB (PCB-22; 2',3',4',5'-tetrachloro-4-biphenylol) and PCB congener (PCB-77; 3,3',4,4'-tetrachlorobiphenyl) on Sertoli cells isolated from 19- to 21-day-old male rats. Sertoli cells incubated for 24 hours in 10(-7) M PCB 22 and 10(-8) M PCB-77, but not in 0.05% ETOH or 10(-7) M 17beta-estradiol (E2) showed morphological changes. Sertoli cells demonstrated progressive damage with higher concentrations of PCB-77 (10(-7) M). After 24 hours, 10(-7) M PCB-22 killed 20% of the Sertoli cells and equimolar PCB-77 killed 45% of the Sertoli cells in culture. At 10(-8) M, PCB-22 did not kill any significant number of Sertoli cells, whereas PCB-77 killed 40% of cells in culture. This result showed differential effects of PCB compounds, with PCB-77 being more cytotoxic than PCB 22 as tested on Sertoli cells. Because PCB-77 produced greater toxic effects, we further tested this congener on Sertoli cell lactate production. After 24 hours, lactate production by Sertoli cells treated with 10(-7) or 10(-8) M PCB-77 was significantly increased. Finally, Sertoli cells exposed to 10(-7) M PCB-77 showed disorganized and less intense F-actin staining. The results demonstrate that PCBs, but not E2, is directly toxic to Sertoli cells in vitro, and suggest this toxic effect is independent of estrogenic action. PMID- 11105925 TI - The future of andrology is easy: the future is bright indeed. PMID- 11105926 TI - Instrumentation for coincidence imaging with multihead scintillation cameras. AB - Positron emission tomography (PET) has been used as a tool by investigators for many years to study metabolic processes in the body primarily with the radiopharmaceutical 18-fluordeoxyglucose. However, use of this technology has not been widespread because of the high expense of the equipment and its limitation to the imaging of positron emitters only. Recent improvements in scintillation camera technology have now made it possible to produce hybrid multihead cameras that can function in a coincidence mode for the detection of the annihilation radiation from positron emitters and in the normal mode for routine single-photon imaging. Although still limited in sensitivity, these camera systems continue to be improved and the recent addition of iterative reconstruction algorithms and attenuation correction capability have resulted in significant improvements in image quality. The integration of a low resolution computed tomography (CT) scanner with a dualhead camera by 1 manufacturer now makes it possible to perform attenuation correction and image fusion of anatomy and function into 1 image to improve the anatomic localization of abnormalities detected with coincidence imaging. Investigators continue to work on improved electronics and new types of detectors to further improve the sensitivity of these systems. These developments coupled with continued improvements in PET technology have resulted in the availability of a broad spectrum of systems for the investigator to consider when purchasing a system with positron imaging capability. PMID- 11105927 TI - Attenuation correction in hybrid positron emission tomography. AB - Attenuation effects are more severe for coincidence imaging than for single photon imaging. The capability to measure and correct attenuation now exists with dedicated positron emission tomography (PET) scanners. Attenuation correction may or may not improve lesion detection in various situations, but it definitely produces a more realistic radioactivity distribution and is essential for quantitation, which is an important PET capability. For hybrid PET systems, though, which are relatively new and in which neither the performance nor the cost of the scanner can be compromised much compared with the conventional nuclear medicine device alone, attenuation correction is still novel. Just as the entire modality of PET imaging on hybrid gamma cameras has expanded very rapidly, the capability to achieve attenuation correction has quickly followed. Both radioactive source-based and x-ray-based systems exist that provide adequate maps for attenuation correction, and the x-ray systems go even further to provide anatomic detail to aid in image interpretation. PMID- 11105928 TI - The role of hybrid cameras in oncology. AB - The rapid advances in imaging technologies are a challenge for nuclear medicine physicians, radiologists, and clinicians who must integrate these technologies for optimal patient care and outcome at minimal cost. Multiple indications for functional imaging using F-18-fluorodeoxyglucose (FDG) are now well accepted in the field of oncology, including differentiation of benign from malignant lesions, staging malignant lesions, detection of malignant recurrence, and monitoring therapy. The use of FDG imaging was first shown using dedicated positron emission tomography (PET) with multiple full rings of bismuth germanate detectors. Most manufacturers now have available hybrid gamma cameras capable of imaging conventional single-photon emitters, as well as positron emitters such as FDG. This new technology was developed to make FDG imaging more widely accessible, first using single photon emission computed tomography (SPECT) with high-energy collimators, and then using dualhead coincidence (DHC) detection with multihead gamma cameras that improved spatial resolution. Most hybrid gamma cameras are now equipped with thicker NaI(TI) crystals to improve sensitivity. Technical developments are still evolving with correction for attenuation and new iterative reconstruction algorithms to improve the quality of the images. Users need to be familiar with the rapid developments of the technology as well as its limitations. Currently, one model of hybrid gamma camera is equipped with an integrated x-ray transmission system for attenuation correction, anatomic mapping, and image fusion. This powerful tool has promising clinical applications including intensity-modulated radiation therapy. PMID- 11105929 TI - 18-Fluorodeoxyglucose imaging with positron emission tomography and single photon emission computed tomography: cardiac applications. AB - The assessment of myocardial viability has become an important aspect of the diagnostic and prognostic work-up of patients with ischemic cardiomyopathy. Although revascularization may be considered in patients with extensive viable myocardium, patients with predominantly scar tissue should be treated medically or evaluated for heart transplantation. Among the many viability tests, noninvasive assessment of cardiac glucose use (as a marker of viable tissue) with F18-fluorodeoxyglucose (FDG) is considered the most accurate technique to detect viable myocardium. Cardiac FDG uptake has traditionally been imaged with positron emission tomography (PET). Clinical studies have shown that FDG-PET can accurately identify patients with viable myocardium that are likely to benefit from revascularization procedures, in terms of improvement of left ventricular (LV) function, alleviation of heart failure symptoms, and improvement of long term prognosis. However, the restricted availability of PET equipment cannot meet the increasing demand for viability studies. As a consequence, much effort has been invested over the past years in the development of 511-keV collimators, enabling FDG imaging with single-photon emission computed tomography (SPECT). Because SPECT cameras are widely available, this approach may allow a more widespread use of FDG for the assessment of myocardial viability. Initial studies have directly compared FDG-SPECT with FDG-PET and consistently reported a good agreement for the assessment of myocardial viability between these 2 techniques. Additional studies have shown that FDG-SPECT can also predict improvement of LV function and heart failure symptoms after revascularization. Finally, recent developments, including coincidence imaging and attenuation correction, may further optimize cardiac FDG imaging (for the assessment of viability) without PET systems. PMID- 11105930 TI - The economics of creating a positron emission tomography center. AB - Positron emission tomography (PET) scanning has been a powerful research tool since its inception. Changes in the marketplace that have allowed PET to move into the clinical environment include the commercial availability of appropriate radiopharmaceuticals, reimbursement of procedures by insurance companies, and increasing awareness of physicians of the benefits of PET. Facilities that are interested in clinical PET need to develop a process to purchase equipment with an appropriate business plan. This is necessary to assure financial viability and to convince hospital administrators of the viability. The creation of a successful PET program requires an understanding of all aspects relating to a center. The process begins with reviewing the mission statement of the facility. The next step is to prepare the feasibility study, which includes reviewing the existing marketplace and determining the volume, level of referring physicians' interest, and availability of radiopharmaceuticals. Finally, an appropriate pro forma needs to be constructed to facilitate the final decision concerning the potential financial viability of such an endeavor. PMID- 11105931 TI - Muscle uptake of 18-fluorine fluorodeoxyglucose. PMID- 11105932 TI - Unusual intestinal and urinary tract accumulation on bone scan: a case with Indiana pouch. PMID- 11105933 TI - Ecstasy (MDMA): a review of its possible persistent psychological effects. AB - RATIONALE: Recreational use of "ecstasy" (3,4-methylenedioxymethamphetamine; MDMA) has become increasingly widespread. Until recently, however, little was known about the possible persistent psychological effects of extensive use of this drug. OBJECTIVE: The aim of the present review is to evaluate recent empirical evidence concerning the persistent psychological sequelae of recreational ecstasy use. METHODS: The methodologies of open trial studies of recreational ecstasy users are evaluated and reports of the presence or absence of persistent psychological problems are related to the extent of past exposure to ecstasy. RESULTS: There is growing evidence that chronic, heavy, recreational use of ecstasy is associated with sleep disorders, depressed mood, persistent elevation of anxiety, impulsiveness and hostility, and selective impairment of episodic memory, working memory and attention. There is tentative evidence that these cognitive deficits persist for at least 6 months after abstinence, whereas anxiety and hostility remit after a year of abstinence. The possibility that some of these psychological problems are caused by ecstasy-induced neurotoxicity is supported by preclinical evidence of MDMA-induced neurotoxicity and behavioural deficits, evidence of depleted serotonin in heavy ecstasy users, and by dose response relationships between the extent of exposure to ecstasy and the severity of cognitive impairments. CONCLUSIONS: An increasing number of young, heavy ecstasy users are at significant risk of persistent cognitive impairments and disturbances of affect and personality. Some of these problems may remit after abstinence, but residual neurotoxicity and decline of serotonergic function with age may result in recurrent psychopathology and premature cognitive decline. PMID- 11105934 TI - Effect of benzodiazepine on temporal integration in object perception. AB - RATIONALE: Though various psychometrical tests indicate that benzodiazepines affect vigilance, few studies have been conducted to assess the effect of benzodiazepines on attentional processes. OBJECTIVE: We used a RSVP (Rapid Serial Visual Presentation) procedure to investigate the effect of benzodiazepines on the attentional blink effect. It refers to the difficulty in detecting a probe following identification of a target within a temporal window of 500 ms. METHOD: Three experimental groups were tested (placebo, lorazepam and diazepam). Sequences of 15 pictures were centrally displayed for 50 ms each. In a dual-task condition, observers were instructed (1) to identify the target (the single picture on a blue background) and (2) to detect the presence of a probe. In the single-task condition, subjects were asked to detect the probe. The serial position of the probe relative to the target was varied. RESULTS: Performance was equivalent for the three groups in the single-task condition. In the dual-task condition, the attentional blink was increased in magnitude and duration for benzodiazepine-treated subjects, especially diazepam, than for placebo-treated subjects. A large number of intrusions (a tendency to report as target the name of a picture preceding the target) were observed in the benzodiazepine-treated groups. CONCLUSION: The results indicate that benzodiazepines impair visual integration in the temporal domain. This extends previous findings that benzodiazepine impairs visual integration in the spatial domain. The results also suggest that benzodiazepine increase time to disengage attention from a first to a second target. PMID- 11105935 TI - Similarities in the action of Ro 60-0175, a 5-HT2C receptor agonist and d fenfluramine on feeding patterns in the rat. AB - RATIONALE: Activation of 5-HT2C receptors is thought to enhance satiety and to mediate the action of the prototypical anorectic drug d-fenfluramine. OBJECTIVE: Four experiments investigated the role of the 5-HT2C receptor in the modulation of feeding by comparison of the effects of the putative selective 5-HT2C receptor agonist Ro 60-0175 and d-fenfluramine on feeding behaviour. METHODS: Microstructural analyses of meal patterning and drinking of a palatable solution were made over a range of drug doses administered to male Lister hooded rats. RESULTS: Ro 60-0175 increased the latency to the first meal (3 mg/kg) and reduced meal size (1 mg/kg). d-Fenfluramine (1 mg/kg) produced a similar behavioural pattern, but 3 mg/kg produced a more profound hypophagia that persisted for 10-12 h. Ro 60-0175 (1, 3 mg/kg) and d-fenfluramine (1.5 mg/kg) reduced ingestion of a palatable glucose/saccharin solution, by a reduction in the number of bouts of licking, with little effect on the size of individual bouts. d-Fenfluramine induced hypophagia (2.1 mg/kg) was challenged by the administration of the selective 5-HT2C receptor antagonist SB 242084 (1, 3 mg/kg) in the meal patterning paradigm. SB 242084 significantly attenuated the decrease in feeding rate and increase in latency to feed produced by d-fenfluramine, but had no effect on the fenfluramine-induced reduction in meal size. A similar pattern of results was obtained when Ro 60-0175-induced hypophagia (3 mg/kg) was challenged by SB 242084 (1, 3 mg/kg). CONCLUSIONS: These results demonstrate that Ro 60-0175 is a useful probe of the importance of 5-HT2C activation in the control of food intake and support the hypothesis that activation of 5-HT2C receptors is a critical aspect of the hypophagic action of d-fenfluramine. The 5-HT2C receptor may prove to be a useful target in the development of clinically effective drugs for the treatment of obesity. PMID- 11105936 TI - Impact of family history of alcoholism on cocaine-induced subjective effects and pharmacokinetic profile. AB - RATIONALE: The importance of genetic factors in the development of alcoholism has been demonstrated repeatedly. However, the impact of a family history of alcoholism on the development of other drug use has been less thoroughly studied. OBJECTIVE: The present study was conducted to investigate whether individuals with a positive family history of alcoholism (FHP) differ from individuals without a history (FHN) in their pharmacokinetic profile, subjective and physiological response to an acute intranasal dose of cocaine (0.9 mg/kg). METHODS: Nine FHP and nine FHN male occasional cocaine users provided informed consent and participated in this double-blind, placebo-controlled, two-visit study. Responses to cocaine were assessed via a joystick device, the Addiction Research Center Inventory, visual analog scales and heart rate. Plasma concentrations of cocaine and its metabolites, benzoylecgonine and ecgonine methylester also were measured. RESULTS: There were no significant differences between FHP and FHN subjects in subjective reports of intoxication, physiologic responses or plasma cocaine and benzoylecgonine concentrations following cocaine administration. Plasma levels of the cocaine metabolite ecgonine methylester were significantly higher in FHP subjects from 50 to 120 min post-cocaine administration compared to FHN subjects. CONCLUSIONS: Our findings indicate that family history of alcoholism does not appear to influence the behavioral and physiological responses to acute cocaine administration, but that some aspects of cocaine metabolism may be different between the two groups. PMID- 11105937 TI - Pinealectomy blocks stress-induced motor stimulation but not sensitization and tolerance to a dopamine D2 receptor agonist. AB - RATIONALE: The motor stimulant effects of the selective dopamine D2 receptor agonist, (+)-4-propyl-9-hydroxynaphthoxazine (PHNO), develop both tolerance and sensitization depending on circadian rhythms and time of day. Daytime tolerance can be transiently reversed by stress. Given that only tolerance develops when rats are kept under constant light conditions, it seems plausible that the pineal hormone melatonin may determine the circadian rhythm in tolerance and sensitization. OBJECTIVE: The effects of pinealectomy on the development of sensitization and stress-induced reversal of tolerance to sensitization to the motor stimulant effects of PHNO were determined. METHODS: Sprague-Dawley rats were pinealectomized or given sham operations and administered continuously with PHNO (5 microg/h) via subcutaneously implanted mini-pumps. Injections of 2-iodo melatonin were subsequently administered to determine if sensitization to PHNO could be reinstated in the pinealectomized animals, assuming that sensitization would be reduced. RESULTS: Pinealectomy did not influence circadian rhythms in the development of sensitization and tolerance to PHNO. Pinealectomy blocked the motor activation effects of "injection-stress", and this effect was reinstated by treatment with 2-iodo-melatonin. CONCLUSIONS: Melatonin is not involved in the development of sensitization or tolerance to the behavioral effects of PHNO. However, melatonin modulates the stress-induced motor activity responses. PMID- 11105938 TI - Effect of a subanesthetic dose of ketamine on memory and conscious awareness in healthy volunteers. AB - RATIONALE: Ketamine is an NMDA receptor antagonist with psychotogenic and cognitive effects in healthy volunteers and schizophrenic patients which has been proposed to be a useful tool to investigate neurobiological basis of schizophrenia. OBJECTIVE: The present study characterized the effects of a subanesthetic dose of ketamine on memory and related subjective states of awareness in healthy volunteers. METHODS: Twenty-six subjects were given either a 60-min ketamine (0.5 mg/kg per hour) or a placebo infusion. To obtain constant plasma ketamine throughout the experiment, ketamine was administered using a computer-controlled infusion system. Subjects carried out episodic memory tasks involving words presented before and during infusion. Memory performance was assessed with recognition and free recall tasks. Subjective states of awareness were assessed using an experiential approach. Levels of psychopathology were evaluated with BPRS. RESULTS: Ketamine impaired performance in free recall and recognition of words presented during, but not before, infusion. There were no differences between groups concerning states of awareness associated with recognition memory. Subjects under ketamine had higher BPRS total scores as well as BPRS negative and positive cluster scores than control subjects. CONCLUSIONS: Ketamine decreases episodic memory performance by impairing encoding, but not retrieval processes. It does not selectively impair subjective states of awareness associated with recognition memory as it has been seen in patients with schizophrenia. Ketamine might mimic the memory impairment associated with acute, but not chronic, forms of schizophrenia. PMID- 11105939 TI - Nornicotine pretreatment decreases intravenous nicotine self-administration in rats. AB - RATIONALE: Nicotine has been shown to be effective as a treatment for reducing tobacco dependence. However, few studies have examined the effect of other nicotinic agonists to determine if they can also decrease nicotine self administration. OBJECTIVE: The present study determined if nornicotine, a tobacco alkaloid and major nicotine metabolite in brain, could reduce nicotine self administration in rats. METHODS: Each rat was prepared with an indwelling jugular catheter and trained to self-administer intravenous nicotine (0.03 mg/kg per infusion). After nicotine self-administration stabilized, rats were pretreated with either (-)-nicotine (0, 0.1, 0.3, and 1.0 mg/kg free base) or (+/-) nornicotine (0, 1, 3, 5.6, and 10.0 mg/kg free base) and assessed for nicotine self-administration. A separate group of rats was maintained on sucrose reinforced responding and pretreated with nornicotine to determine the specificity of the pretreatment effect. In another group of rats, the time course of the pretreatment effect of either (-)-nicotine (0.56 and 1.0 mg/kg) or (+/-) nornicotine (5.6 and 10.0 mg/kg) was examined. RESULTS: Nicotine and nornicotine each produced a dose-dependent decrease in nicotine self-administration. Furthermore, the decrease in nicotine self-administration in response to the 5.6 mg/kg nornicotine pretreatment was specific to nicotine self-administration, as this dose did not decrease sucrose reinforced responding in tolerant animals. In addition, within the dose range tested, the suppressant effect of nornicotine had a two-fold longer duration than that of nicotine (120 versus 60 min). CONCLUSION: These results suggest that nornicotine may be an effective treatment for tobacco dependence. PMID- 11105940 TI - Antidepressant-like effects of the subtype-selective nicotinic acetylcholine receptor agonist, SIB-1508Y, in the learned helplessness rat model of depression. AB - RATIONALE: Epidemiological studies of smokers suggest that there is a link between nicotine and depression. Nonetheless, few studies have examined the potential use of nicotinic ligands in the treatment of depression. OBJECTIVES: The goal of this study was to evaluate the effects of SIB-1508Y, a novel subtype selective ligand for high affinity nicotinic acetylcholine receptors (nAChRs), in the learned helplessness model of depression in rats. METHODS: In this model, exposure to inescapable foot-shock produces a lasting deficit in escape responses emitted in a subsequent conditioned avoidance procedure (learned helplessness). The effect of SIB-1508Y on learned helplessness was compared to the clinically used antidepressants, imipramine and fluoxetine, and the non-selective nAChR ligand, nicotine. RESULTS: Similarly to imipramine and fluoxetine, subchronic treatment (5 days) with SIB-1508Y reversed the escape deficit in the learned helplessness model in a dose dependent manner. The effect of SIB-1508Y on learned helplessness was still apparent 1 week following drug administration and was also maintained after 4 weeks of daily administration. In contrast, while nicotine was able to attenuate the learned helplessness deficit, this trend only reached statistical significance after chronic administration. The non-competitive ion channel blocker mecamylamine increased escape failures when administered alone and blocked the effects of SIB-1508Y but not imipramine. SIB- 1508Y also produced an increase in avoidance responding, which suggests an enhancement of learning. CONCLUSION: These results not only suggest a role for nAChRs in the development of a depressive-like syndrome, but also that subtype-selective nAChR agonists, such as SIB-1508Y, may offer a novel therapeutic approach to the treatment of depression. PMID- 11105941 TI - Locomotor sensitization to quinpirole in rats: effects of drug abstinence and sex. AB - RATIONALE: Behavioral sensitization, induced by the chronic administration of psychomotor stimulants, serves as an experimental model for the development of behavioral pathology. Although many factors are known to influence the sensitization produced by indirect dopamine agonists, such as cocaine and the amphetamines, less is known about factors that influence the behavioral sensitization produced by direct dopamine receptor agonists. OBJECTIVE: As the extent to which behavioral sensitization is expressed following the repeated administration of indirect dopamine agonists can depend upon a period of drug abstinence, the present study determined the effects of drug abstinence on the expression of locomotor sensitization to the D2/D3 receptor agonist, quinpirole (QNP). METHODS: Male and female rats were administered ten, twice weekly, injections of 0.5 mg/kg QNP or saline (SAL), and then received one of five QNP doses (0-1.0 mg/kg; n=7-10/dose) in two dose-response tests for locomotor sensitization, conducted at 3 and 15 days following the cessation of chronic treatment. RESULTS: The sensitized locomotor response of QNP-treated animals was similar on the 2 test days in both male and female subjects. Compared to males, female rats displayed greater locomotor responding to QNP, both during chronic treatment and on the dose-response tests for sensitization. CONCLUSIONS: QNP locomotor sensitization is (a) not influenced by 2 weeks of QNP abstinence and (b) can be influenced by the sex of the animal. It is suggested that direct and indirect dopamine agonists produce locomotor sensitization via distinct mechanisms that differ in sensitivity to the passage of time but are both influenced by sex-specific variables. PMID- 11105942 TI - Effects of noradrenaline depletion in the brain on response on novelty in isolation-reared rats. AB - RATIONALE: Social isolation from weaning in the rat produces a variety of neurochemical and behavioural effects in the adult that in part parallel changes seen in human schizophrenia. OBJECTIVES: The study investigated the effects of central noradrenaline (NA) depletion by the selective neurotoxin, N-(2 chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), on the behaviour of isolation reared rats. METHODS: Male Lister hooded rats were reared singly or in groups after weaning. During week 2, the rats were tested in photocell activity cages and were then injected with DSP-4 (25 mg/kg, IP). During week 4, rats were tested in the open field under the following conditions: open field alone, with two novel stimuli (T1), and with a familiar and a novel object (T2), and in the activity cages. RESULTS: DSP-4 significantly reduced cortical and hippocampal NA levels with no effect on the hypothalamus. Isolation-reared rats exhibited locomotor hyperactivity and reduced habituation to the testing arena, although their exploration of the novel objects in T1 was not significantly different from group-reared rats. DSP-4 treatment in group-reared rats increased inner zone activity in the open field but did not significantly affect the exploration of novel objects. DSP-4 treatment in isolates reduced exploration of objects at T2 while increasing exploration of the general environment. CONCLUSIONS: Isolation rearing influences the behavioural effects of central NA depletion. The results suggest isolation-induced changes in the central noradrenergic system in the isolated rat, supporting the view that early environmental factors can have long term effects on central noradrenergic function as well as other neurotransmitter systems. PMID- 11105943 TI - Tolerance to repeated nicotine administration on performance, subjective, and physiological responses in nonsmokers. AB - RATIONALE: When administered acutely to nonsmokers, nicotine's effects on performance are inconsistent, perhaps because of suboptimal dosing or initial dysphoria that could interfere with performance. OBJECTIVE: The purpose of this study was to determine if a range of nicotine doses administered for 8 days to nonsmokers would enhance psychomotor and cognitive abilities and to document the development of nicotine tolerance or sensitization. METHODS: Twelve male volunteers, who reported ever smoking five cigarettes or less, participated in 8 consecutive experimental days in which they were administered four doses of nicotine polacrilex gum each day in this order: 0, 2, 4, and 8 mg. Performance, subjective, and physiological measures were assessed before and after each dose. RESULTS: Plasma nicotine concentration ranged from 6.9 to 11.5 ng/ml following the 8 mg dose. Nicotine increased rate of responding and decreased response time on working memory (digit recall); however, accuracy was impaired. Nicotine also decreased accuracy on visual scanning and attention (two-letter search), and the 8 mg dose impaired gross motor coordination (circular lights). Tolerance did not develop to the performance impairing effects of nicotine. Nicotine produced dose related increases in ratings of dysphoria and negative mood, including tension, anxiety, nervousness, turning of stomach, and sedation. Tolerance developed to some, but not all, of these aversive effects. Tolerance also was not observed to the increased cardiovascular measures. CONCLUSION: Although tolerance developed to some of the aversive effects of nicotine, performance enhancement was not observed. These data do not support the hypothesis that nicotine-induced performance enhancement contributes to the reinforcing effects of tobacco use during the early stages of dependence development. PMID- 11105944 TI - Smoking-related cues elicit craving in tobacco "chippers": a replication and validation of the two-factor structure of the Questionnaire of Smoking Urges. AB - RATIONALE: The reliability and validity of the Questionnaire of Smoking Urges (QSU), a multi-factorial measure of cravings to smoke cigarettes has recently been called into question. OBJECTIVE: In the first phase of the present study, the reliability of the two-factor structure of QSU was examined. The new and original factor structures were then used to investigate the effects of 2 h of abstinence from smoking, and exposure to smoking-related cues, in a subset of the same sample of regular smokers. The present study also investigated the effects of smoking-related cues on QSU measures in tobacco "chippers" (occasional non dependent smokers). METHODS: The factor structure of the QSU was investigated with a factor analysis of responses to the 32 QSU items produced by a sample of 271 regular smokers. A subset of these data was used to assess the influence of abstinence and cues on QSU factor scores. The final study used a sample of 32 tobacco chippers to assess the effect of smoking-related cues on their QSU factor scores. RESULTS: The results revealed a two-factor structure almost identical to that published by Tiffany and Drobes. In the second experiment, a brief period of abstinence significantly elevated factor 1 scores, but not factor 2 scores, compared to the non-abstinent condition. Exposure to smoking-related cues significantly elevated factor 2 scores and also tended to elevate factor 1 scores. In experiment 3, exposure to smoking-related cues significantly elevated factor 1 scores, but not factor 2 scores, in tobacco chippers. CONCLUSION: The QSU is a psychometrically sound instrument for the assessment of urges to smoke, which shows good construct validity. The two factors of the QSU show differential discriminative properties. PMID- 11105945 TI - Lack of operant ethanol self-administration in dopamine D2 receptor knockout mice. AB - RATIONALE: Dopamine D2 receptors are postulated to play an important role in modulating the reinforcing effects of abused drugs including ethanol. OBJECTIVES: This experiment examined operant ethanol self-administration in dopamine D2 receptor knockout (KO) mice and wild-type (WT) mice using a continuous access procedure. METHODS: Adult male KO and WT mice were trained in 30-min sessions to perform a lever press response for access to 10% v/v ethanol. After training, the mice were placed in test chambers on a continuous (23 h/day) basis with access to food (one lever press, i.e., FR1), 10% v/v ethanol (four lever presses, i.e., FR4), and water from a sipper tube (phase 1). After 30 consecutive sessions, response patterns were determined for 0, 5, 10, 20 and 30% v/v ethanol (phase 2). Saccharin (0.2% w/v) was subsequently added to the ethanol mixture and responding was examined for 0, 5, 10 and 20% ethanol (phase 3). RESULTS: During phase 1, WT mice displayed higher ethanol-lever responding compared to KO mice. Food lever responding and water intake was the same in both genotypes. During phase 2, WT mice displayed concentration-dependent ethanol lever responding, whereas KO mice responded at low rates regardless of ethanol concentration. WT mice also responded more for food compared to KO mice. Each genotype showed similar water intakes except at the 20% ethanol concentration, where WT mice had lower intakes. During phase 3, WT mice continued to show higher responding for all concentrations including saccharin alone. WT mice also continued to respond more for food compared to KO mice, but drank less water. In each phase, WT mice displayed episodic (bout) responding on the ethanol lever. KO mice did not respond for ethanol in bouts. CONCLUSIONS: Reduced responding in the KO mice for several reinforcers including ethanol indicates a more general role for dopamine D2 receptors in motivated responding rather than a specific role in ethanol reinforcement. PMID- 11105946 TI - Morphologic and immunohistochemical features of experimentally induced allergic contact dermatitis in Gottingen minipigs. AB - Many preclinical studies in investigative dermatology are performed preferably in pigs because pig skin is more similar to human skin than is rodent skin. A frequently used model is allergic contact dermatitis (ACD); however, this T-cell mediated skin condition so far is not well characterized in pigs. The present study is aimed at the evaluation of morphologic and immunohistochemical features of experimentally induced acute ACD in Gottingen minipigs using 2,4 dinitrofluorobenzene (DNFB) as a hapten. Eight minipigs were sensitized with 10% DNFB and challenged 2 weeks later at different sites with 1% DNFB. In addition to clinical examinations, cutaneous blood flow was quantified by laser Doppler velocimetry (Periflux PF3). These examinations were performed before challenge and 8, 24, 48, and 72 hours after challenge. Skin biopsies were taken at the same time points, fixed, sectioned, and stained with Giemsa for histologic evaluation, or with mouse anti-swine monoclonal antibodies (CD1, CD2, CD4, CD5, CD8, CD25, CD45, MHCII) and with one mouse anti-human monoclonal antibody (CD62E) cross reacting with swine for immunohistochemical evaluation. Positively stained cells were counted per square millimeter of epidermis and dermis by using a video image analyzing system (Videoplan Kontron). Erythema and cutaneous blood flow peaked at 24 hours. The major epidermal changes most pronounced at 48 hours were acanthosis, spongiosis, intracellular edema, exocytosis, and abscesses mainly containing neutrophils and mononuclear cells (MNC). Perivascular infiltrates of MNC as well as neutrophils and eosinophils were the most significant dermal changes, with peak levels at 24-48 hours. In biopsies taken before challenge, CD1+ dendritic cells were found in similar numbers and locations as MHCII+ cells in the epidermis. In the epidermis the maximum CD1+ cell decrease occurred at 24 hours whereas in the dermis the maximum increase in CD1+ stained cells was seen at 72 hours. The dermal infiltrate (CD2+, CD5+, CD25+, and CD45+) was most dense at 48 hours. Between 8 and 48 hours more CD4+ were present than CD8+, cells, whereas at 72 hours CD4+ and CD8+ cells were similar in numbers. These findings closely resemble changes in human ACD. Therefore, DNFB-induced ACD in Gottingen minipigs is considered to be an appropriate animal model to study immunopathologic mechanisms and pharmacologic intervention. PMID- 11105947 TI - Two genetic defects in alphaIIb are associated with type I Glanzmann's thrombasthenia in a Great Pyrenees dog: a 14-base insertion in exon 13 and a splicing defect of intron 13. AB - Glannzmann's thrombasthenia (GT) is an autosomal recessive bleeding disorder caused by qualitative or quantitative deficiencies of the platelet membrane glycoprotein alphaIIbbeta3. This is the first report of a molecular genetic basis for type I GT in dogs. As previously reported, a thrombasthenic Great Pyrenees dog (dog No. 1) experienced uncontrolled epistaxis despite results of coagulation screening tests, platelet quantitation, and von Willebrand factor quantitation that were within reference ranges. Platelet aggregation was minimal in response to agonists. Flow cytometry, autoradiography, and immunoblot experiments demonstrated either marked reduction or absence of glycoproteins alphaIIb and beta3. In this study, we report the presence of a 14-base insertion in exon 13 and defective splicing of intron 13 in the alphaIIb gene of two thrombasthenic dogs (Nos. 1 and 8). The insertion disrupted the fourth alphaIIb calcium-binding domain, caused a shift in the reading frame and resulted in a premature termination codon. Possible consequences of this mutation include decreased alphaIIb mRNA stability and production of truncated alphaIIb protein that lacks the transmembrane and cytoplasmic domains and a large portion of the extracellular domain. We identified the dam, sire, and three littermates of dog No. 8 as carriers of the alphaIIb mutation. Canine alphaIIb and beta3 genes share significant homology with the genes in human beings, making canine GT an excellent translational model for human GT. A defined molecular basis for canine GT will enhance ongoing gene therapy research and increase the understanding of structure-function relationships of this integrin. PMID- 11105948 TI - Gastritis and intestinal metaplasia in Syrian hamsters infected with Helicobacter aurati and two other microaerobes. AB - Chronic gastritis and intestinal metaplasia associated with naturally occurring colonization by Helicobacter aurati and two other microaerobic species were observed in Syrian hamsters. Thirty-five hamsters, between 7 and 12 months of age, were evaluated from two research and three commercial facilities. Microaerobic bacteria were cultured from the hamster stomachs. These bacteria included H. aurati, a fusiform, urease-positive species; a second novel helical, urease-negative Helicobacter sp.; as well as a smaller, urease-negative Campylobacter sp. Southern blot analysis detected Helicobacter spp. DNA in the gastric tissues of all 35 hamsters; 15 hamsters also had Campylobacter sp. DNA in their gastric tissues. When examined by light microscopy, argyrophilic bacteria consistent with H. aurati or the second Helicobacter sp. were present in antral sections of 12 out of the 15 hamsters where bacteria were seen, while 9 out of the 15 hamsters had bacteria resembling the Campylobacter sp. The presence of Helicobacter spp. but not the presence of Campylobacter sp. was significantly correlated to gastritis severity (P < 0.0001 for Helicobacter spp., P = 0.6025 for Campylobacter sp.) and intestinal metaplasia, as measured by numbers of goblet cells (P = 0.0239 for Helicobacter spp., P = 0.5525 for Campylobacter sp.). Severely affected hamsters also had Giardia sp. within their metaplastic gastric pits. Hamsters with naturally occurring helicobacter-associated gastritis provide a model for studying the development of intestinal metaplasia and gastric giardiasis in H. pylori-infected humans. PMID- 11105949 TI - Retrospective study of 338 canine oral melanomas with clinical, histologic, and immunohistochemical review of 129 cases. AB - Diagnostic records from 338 canine oral melanomas in 338 dogs received at the Veterinary Medical Diagnostic Laboratory (1992-1999) were reviewed. Of these tumors, 122 plus an additional 7 metastatic melanomas of unknown origin were selected for clinical follow-up, histologic review, and immunohistochemistry. Chow Chow, Golden Retriever, and Pekingese/Poodle mix breeds were overrepresented, whereas Boxer and German Shepherd breeds were underrepresented. There was no gender predisposition and the average age at presentation was 11.4 years. Forty-nine dogs were euthanized due to recurrence or metastasis. The average postsurgical survival time was 173 days. The gingiva and the labial mucosa were the most common sites. Most tumors were composed of either polygonal cells (27 cases, 20.9%), spindle cells (44 cases, 34.1%), or a mixture of the two (polygonal and spindle) (54 cases, 41.9%). Clear cell (3 cases, 2.3%) and adenoid/papillary (1 case, 0.8%) patterns were uncommon. The metastases of 6/6 oral melanomas had morphologic and immunohistochemical features similar to those of the primary tumors. Immunohistochemically, Melan A was detected in 113/122 oral (92.6%) and 5/7 (71.9%) metastatic melanomas. Only 4/163 nonmelanocytic tumors were focally and weakly positive for Melan A. Antibodies against vimentin, S100 protein, and neuron-specific enolase stained 129 (100%), 98 (76%), and 115 (89.1%) of 129 melanomas, respectively. Antibodies against other melanocytic associated antigens (tyrosinase, glycoprotein 100) did not yield adequate staining. We conclude that Melan A is a specific and sensitive marker for canine melanomas. PMID- 11105950 TI - Common metastatic carcinoma of California sea lions (Zalophus californianus): evidence of genital origin and association with novel gammaherpesvirus. AB - Tissues from 10 adult California sea lions (Zalophus californianus, seven females and three males) that had metastatic carcinoma in sublumbar area lymph nodes were examined histologically. A distinctive epithelial proliferative lesion interpreted as intraepithelial neoplasia was found in genital tracts of all ten animals; in vagina (5/7), cervix (7/7), uterus (3/7), penis (3/3) and prepuce (3/3). Intraepithelial neoplasia closely resembled metastatic carcinomas and was directly contiguous with invasive carcinoma in one animal. Rare eosinophilic intranuclear inclusion bodies were found in penile and preputial intraepithelial neoplasia (one animal), cervical intraepithelial neoplasia (one animal), invasive cervical carcinoma (one animal) and metastatic carcinoma (two animals). Electron microscopic examination of tissues from two sea lions (one with intraepithelial neoplasia and one with metastatic carcinoma) demonstrated viral particles consistent with a herpesvirus. An immunohistochemical stain for the latent membrane protein of Epstein-Barr virus was positive in intraepithelial neoplasia in one sea lion. Herpesvirus DNA sequences were detected by consensus primer polymerase chain reaction (PCR) in metastatic carcinomas from all four sea lions from which unfixed tumor samples were available. Results of sequencing were consistent with a novel gammaherpesvirus in the genus Rhadinovirus. DNA extracted from the four metastatic carcinomas also was tested for papillomavirus by Southern blot and PCR with consensus papillomavirus primers; all samples were negative by both methods. These findings support the genital origin of the sea lion carcinoma and implicate a novel gammaherpesvirus as a possible cause. PMID- 11105951 TI - In situ hybridization detection of bovine respiratory syncytial virus in the lung of experimentally infected lambs. AB - We studied the distribution of bovine respiratory syncytial virus (BRSV) RNA in lungs of experimentally inoculated lambs by in situ hybridization at different times postinoculation. The probe used for in situ hybridization was prepared by reverse transcription of BRSV RNA, followed by polymerase chain reaction (PCR) amplification of the cDNA. Twenty-five Merino lambs of both sexes with a live weight of 17 +/- 3 kg received an intratracheal inoculation of 20 ml saline solution containing 1.26 X 10(6) TCID50 BRSV (strain NMK7)/ml. Lambs were slaughtered 1, 3, 7, 11, and 15 days postinoculation (PID). Bronchial and bronchiolar epithelial cells were positive for BRSV nucleic acid by ISH at 1, 3, 7, and 11 PID. However, alveolar epithelial cells contained positive cells at 1, 3, and 7 PID. Cells containing viral RNA were detected from 1 to 11 PID in exudate within bronchial and bronchiolar lumina and from 3 to 7 PID in alveolar exudates. Positive hybridization signals were identified in interstitial mononuclear cells and in bronchi-associated lymphoid tissue from 3 to 11 PID. Mononuclear cells were located in peribronchiolar tissue and interalveolar septa. The highest signal intensity in positive cells was observed at 3 and 7 PID, coinciding with the most important histopathological findings. PMID- 11105952 TI - Mouse model of sublethal and lethal intraperitoneal glanders (Burkholderia mallei). AB - Sixty male BALB/c mice were inoculated intraperitoneally with either a sublethal or a lethal dose of Burkholderia mallei China 7 strain, then killed at multiple time points postinoculation. Histopathologic changes were qualitatively similar in both groups and consisted of pyogranulomatous inflammation. In sublethal study mice, changes were first seen at 6 hours in mediastinal lymph nodes, then in spleen, liver, peripheral lymph nodes, and bone marrow at day 3. These changes generally reached maximal incidence and severity by day 4 but decreased by comparison in all tissues except the liver. Changes were first seen in lethal study mice also at 6 hours in mediastinal lymph nodes and in spleens. At day 1, changes were present in liver, peripheral lymph nodes, and bone marrow. The incidence and severity of these changes were maximal at day 2. In contrast to sublethal study mice, the incidence and severity of the changes did not decrease through the remainder of the study. The most significant difference between the two groups was the rapid involvement of the spleen in the lethal study mice. Changes indicative of impaired vascular perfusion were more frequently seen in the sublethal study mice. Our findings indicate that mice are susceptible to B. mallei infection and may serve as an appropriate model for glanders infection in a resistant host such as human beings. Additionally, by immunoelectron microscopy, we showed the presence of type I O-antigenic polysaccharide (capsular) antigen surrounding B. mallei. PMID- 11105953 TI - Clinical, hematologic, and immunophenotypic characterization of canine large granular lymphocytosis. AB - Clinical, hematologic, and immunophenotypic data were studied in 25 dogs with large granular lymphocyte (LGL) lymphocytosis. Primarily large-breed dogs were affected, with an average age at initial diagnosis of 10 years (range 5-14 years). All dogs had persistent (>4 months) LGL lymphocytosis except for three that were euthanized with aggressive disease. Splenomegaly was reported in 12 of 20 dogs in which splenic size was evaluated. The clinical course was heterogeneous and dogs were divided into four groups based on similar clinical and hematologic findings: acute leukemia (3/25), persistent lymphocytosis with anemia (12/25), persistent lymphocytosis without anemia (8/25), and reactive lymphocytosis (2/25). Immunophenotypes varied within groups but were homogeneous among cells from the same patient except in the two dogs classified as reactive LGL lymphocytosis. Analysis of T-cell receptor (TCR) usage identified three main LGL lineages. TCRalphabeta was expressed in 15/25 (60%) cases. TCRgammadelta was expressed in 8/25 (32%) cases, and 2/25 (8%) cases were CD3-, compatible with NK cells. beta2 integrin expression was distinctive. CD11a was consistently expressed, while CD11b was absent. CD11c was expressed only weakly in 16/25 (64%) cases. The leukointegrin alphadbeta2 was highly prevalent on all LGL lineages, being expressed in 23/25 (92%) cases. Prominent involvement of the spleen, relative sparing of bone marrow, an unexpectedly large proportion of gammadelta T cell LGLs, and the distinctive beta2 integrin expression pattern on diverse lineages of LGLs suggest the disease arises from unique populations of lymphocytes that preferentially localize in the splenic red pulp. PMID- 11105954 TI - Parvovirus infection of keratinocytes as a cause of canine erythema multiforme. AB - Erythema multiforme major was diagnosed in a dog with necrotizing parvoviral enteritis. Skin lesions consisted of ulceration of the footpads, pressure points, mouth, and vaginal mucosa; vesicles in the oral cavity; and erythematous patches on the abdomen and perivulvar skin. Microscopic examination of mucosal and haired skin specimens revealed lymphocyte-associated keratinocyte apoptosis at various levels of the epidermis. Basophilic cytoplasmic inclusions were seen in basal and suprabasal keratinocytes. Immunohistochemical staining, performed with canine parvovirus-2-specific monoclonal antibodies, confirmed the parvovirus nature of the inclusions in the nucleus and cytoplasm of oral and skin epithelial cells. This is the first case of canine erythema multiforme reported to be caused by a viral infection of keratinocytes. This case study indicates that the search for epitheliotropic viruses should be attempted in cases of erythema multiforme in which a drug cause cannot be identified. PMID- 11105955 TI - Pyogranulomatous meningoencephalitis due to Actinomyces sp. in a dog. AB - Actinomyces sp. are commensal, filamentous, gram-positive, acid-fast-negative bacteria that can cause pyogranulomatous inflammation in animals and humans. Central nervous system (CNS) disease is a rare presentation of actinomycosis and is usually due to extension from infected wounds or seeding from distant sites. A dog with progressive, poorly localized neurologic signs had primary CNS actinomycosis without history or evidence of previous trauma or other organ involvement. Histologically, there was a severe pyogranulomatous meningoencephalitis with intralesional filamentous bacteria that were also visible on cytology of the cerebral spinal fluid (CSF) postmortem. Actinomyces sp. was cultured postmortem from the CSF, confirming the diagnosis. This case demonstrates that Actinomyces sp. can be a causative agent of primary CNS disease in dogs. PMID- 11105956 TI - Meningeal osteosarcoma in a dog. AB - A meningeal osteosarcoma was diagnosed in a dog displaying neurologic signs compatible with a space-occupying cerebellar lesion. Gross lesions, restricted to the brain, consisted of a solitary, compressive mass attached to the dura mater overlying the left cerebellum. The mass was composed of single and multinucleated, atypical polygonal cells that lined or rested within lacuna surrounded by eosinophilic, mineralized matrix. The matrical component stained dark green-yellow to blue with Movat's pentachrome stain, deep blue to red with Heidenhain aniline blue stain, and brown-black with Von Kossa stain. Results of these stains were interpreted as tumor osteoid. Foci of dural mineralization and osseous metaplasia were present at the point of tumor attachment. The microscopic observations were interpreted as an osteosarcoma of extraskeletal origin. To our knowledge, these findings represent the first documented case of a meningeal osteosarcoma in a domestic animal species. PMID- 11105957 TI - Spontaneously occurring hepatocellular neoplasia in adolescent cynomolgus monkeys (Macaca fascicularis). AB - Spontaneous hepatic neoplasms were identified in two adolescent (<5 years of age) male cynomolgus monkeys (Macaca fascicularis). Monkey No. 1 had a solitary hepatocellular carcinoma (HCC). Monkey No. 2 had multiple discrete tumors consisting of several poorly circumscribed HCCs and a mixed hepatocholangiocellular carcinoma (MHC). Metastases were not evident in either monkey. Histochemical and immunohistochemical stains were used to assess phenotypic alterations in the tumors. Many or most neoplastic hepatocytes (NHs) of both monkeys stained positive for low-molecular-weight cytokeratin (LMWCK), cytokeratin (CK) 8, and CK 18. In monkey No. 1, small aggregates of NHs were positive for carcinoembryonic antigen (CEA), glutathione S-transferase-pi (GST), and alpha-fetoprotein (AFP), but NHs were uniformly negative for CK 7. NHs in monkey No. 2 were negative for CEA and AFP but were multifocally positive for GST and CK 7. Broad-spectrum cytokeratin (BSCK), high-molecular-weight cytokeratin (HMWCK), and CK 19 did not react with NHs of either animal. Neoplastic cells forming ductlike structures in the MHC of monkey No. 2 stained with LMWCK, CK 7, CK8, CK 18, BSCK, and GST but not with HMWCK or CK 19. Tumors in both monkeys had enhanced pericellular fibronectin staining. Nonneoplastic parenchyma of both monkeys contained multiple discrete foci of cellular alteration and scattered aggregates of hepatocytes with strong cytoplasmic staining for fibronectin. Staining patterns of these tumors demonstrate immunophenotypic heterogeneity of the neoplastic cells within individual tumors and variability among tumors. This information may serve as a useful reference for others encountering similar lesions in primates. PMID- 11105958 TI - Primary pulmonary artery leiomyosarcoma in an adult dog. AB - A 7-year-old neutered female English Setter presented with syncope, anemia, and weight loss. Clinical examination revealed a systolic murmur and echocardiography demonstrated a mass on the pulmonic valve. Postmortem examination confirmed the presence of a pulmonic valve mass that extended along the pulmonary trunk and into the left pulmonary artery. Multiple pale nodules were observed in the right lung. Microscopic examinations of the pulmonary artery mass and the lung nodules revealed a pleomorphic population of spindle cells often arranged in broad bands containing strap-like nuclei and eosinophilic cytoplasm devoid of cross striations. The neoplastic cells expressed vimentin and alpha-smooth muscle actin but did not express desmin, CD31, factor VIII, or S100. The presentation, histological features, immunocytochemical profiles, and behavior of this tumor were indicative of a primary pulmonary artery leiomyosarcoma with lung metastasis. PMID- 11105959 TI - Retrospective study of porcine circovirus 2 infection in Japan: seven cases in 1989. AB - Retrospectively, we demonstrated that seven pigs necropsied in 1989 had characteristic porcine circovirus 2 (PCV-2) lymphoid tissue lesions of severe lymphoid depletion, syncytial giant cell formation, and intracytoplasmic inclusion bodies in macrophages. Immunohistochemically, PCV-2 antigen was detected in these tissues and the distribution of positive staining corresponded to the distribution of inclusion bodies. Electron microscopy demonstrated viral particles compatible with porcine PCV-2. Therefore, the disease occurred in Japan as early as 1989. PMID- 11105960 TI - Congenital hepatic fibrosis and cystic bile duct formation in Swiss Freiberger horses. AB - Congenital hepatic fibrosis with autosomal recessive or dominant inheritance has been described in humans, cats, piglets, and dogs. In horses, only two cases of congenital hepatic fibrosis have been previously reported. This retrospective study of records from the Institute for Animal Pathology, University of Berne, identified 30 foals with liver lesions compatible with congenital hepatic fibrosis. Anamnestic data revealed clinical signs of severe liver injury in most affected animals. Pathologic examination showed severely enlarged, firm livers with thin-walled cysts. Histologically, the livers showed diffuse porto-portal bridging fibrosis with many small, irregularly formed and sometimes cystic bile ducts. All foals belonged to the Swiss Freiberger breed. Pedigree analysis revealed that the diseased animals could be traced back to one stallion. These results strongly suggest that congenital hepatic fibrosis in Swiss Freiberger horses is a recessively inherited autosomal genetic defect. PMID- 11105961 TI - Fatal nonneurological EHV-1 infection in a yearling filly. AB - A case of fatal nonneurological equine herpesvirus 1 (EHV-1) infection in a yearling filly is described. Gross lesions included extensive pulmonary edema, prominent laryngeal lymphoid follicles, and congestion and edema of the dorsal third ventricle choroid plexus. Histologically, there was vasculitis, hemorrhage, and edema in the lungs and dorsal third ventricle choroid plexus as well as mild intestinal crypt necrosis with occasional intranuclear inclusion bodies. The perivascular and vascular inflammatory infiltrates were comprised mainly of T lymphocytes and macrophages. EHV-1 antigen was identified within the nucleus and cytoplasm of endothelial cells, dendritic-like cells of the pharyngeal lymphoid follicles, pharyngeal glandular epithelium, crypt enterocytes, and monocytes. Attempted virus isolation was negative. Weak seroconversion for EHV-1 was observed. Herpesvirus-like particles were identified within pharyngeal endothelial cells by transmission electron microscopy. Polymerase chain reaction amplified 369 and 188 base-pair fragments specific for EHV-1. The scarcity of pathognomonic viral inclusions and lesions in this case suggests that this disease may not be recognized, particularly in situations when ancillary laboratory procedures are limited. PMID- 11105962 TI - Axonal and neuronal amyloid precursor protein immunoreactivity in the brains of guinea pigs given tunicamycin. AB - Amyloid precursor protein (APP) immunocytochemistry was used to study axonal and neuronal changes in guinea pig brains exposed to tunicamycin. Substantial axonal injury was found in ischemic-hypoxic foci and more generally, but this injury was not readily appreciable in conventionally stained sections. Neuronal perikaryal APP expression was also widely distributed, possibly as an acute phase response to this neurotoxin. PMID- 11105963 TI - Malignant teratoid medulloepithelioma in a llama. AB - A lesion was identified in the eye of a juvenile llama, and preliminary clinical findings included anterior uveitis and an exudative retinal detachment suggestive of infectious disease. However, histopathologic evaluation of the enucleated globe revealed an intraocular neoplasm composed of primitive neuroepithelium forming ribbons, cords, and rosettes, heteroplastic elements including spindle cells in a loose myxomatous matrix, and islands of well-differentiated hyaline cartilage. Immunohistochemically, neoplastic cells were positive for vimentin and neuron-specific enolase. Spindle cells were multifocally positive for desmin and muscle specific actin, indicating differentiation towards myofibers. These findings are consistent with a diagnosis of malignant teratoid medulloepithelioma, an extremely rare ocular neoplasm that affects children and young animals. PMID- 11105964 TI - Cholesterol granulomas in three meerkats (Suricata suricatta). AB - Cholesterol granulomas are uncommon pathologic lesions in animals, although they are important intracranial tumors in humans. This report describes cholesterol granulomas associated with multiple organ systems of three captive meerkats. In the most severe case, meerkat No. 1, the pathologic behavior of the cholesterol granuloma was unique in that it appeared to locally invade the cerebrum and calvarium, possibly contributing to neurological deficits observed antemortem. A review of other meerkat necropsies revealed incidental, asymptomatic cholesterol granulomas in organs of two other individuals, meerkat Nos. 2 and 3. Histologically, all lesions were composed of cholesterol clefts admixed with large, foamy macrophages containing hemosiderin, multinucleated giant cells, lymphocytes, plasma cells, and foci of mineralization. Hypercholesterolemia was documented in two of the three meerkats. PMID- 11105965 TI - Hepatitis and staging of hepatic damage in pigs naturally infected with porcine circovirus type 2. AB - A total of 100 liver samples from pigs with postweaning multisystemic wasting syndrome (PMWS) were studied. All livers were evaluated microscopically and were staged based on the severity and localization of lesions. Presence of porcine circovirus type 2 (PCV-2) was evaluated using an in situ hybridization technique. Eighty-eight of 100 livers (88%) had a variable degree of lymphohistiocytic hepatitis, with apoptotic bodies, disorganization of hepatic plates, and/or perilobular fibrosis. Twelve pigs did not have microscopic liver lesions. Four stages of hepatic damage were established based on intensity and distribution of the lesions. Viral nucleic acid was detected in 70 of 100 livers (70%). Target cells for PCV-2 infection included Kupffer cells, hepatocytes, and inflammatory infiltrates. According to distribution of PCV-2 nucleic acid, four basic labeling patterns were identified. This study shows that liver damage is a frequent microscopic finding in cases of PMWS and hepatocytes are a target cell for PCV-2 infection and replication. Therefore, PCV-2 should be considered a new hepatitis inducing viral agent in pigs. PMID- 11105966 TI - Transitional cell carcinoma of the urinary bladder in a Thoroughbred, with intra abdominal dissemination. AB - A 14-year-old Thoroughbred gelding with a history of acute onset of hematuria was presented for necropsy. Transitional cell carcinoma of the urinary bladder with intra-abdominal dissemination was diagnosed. Tumor masses were observed on the splenic capsule and surrounding the distal abdominal aorta. Tumor cells showed diffuse cytoplasmic reactivity for cytokeratin but were negative for epithelial membrane antigen, carcinoembryonic antigen, tumor-associated glycoprotein 72, and vimentin. PMID- 11105967 TI - Gestational diabetes mellitus: metabolic control during labour. AB - The purpose of this study was to assess, in women with gestational diabetes mellitus (GDM): 1) metabolic control during labour using a standardised protocol; 2) the influence of therapy during pregnancy in intrapartum metabolic control and insulin requirements; and 3) the impact of maternal glycaemia during labour on neonatal hypoglycaemia. An observational study of 85 women with GDM (54 insulin treated) was performed. Intrapartum metabolic management included i.v. glucose and insulin infusions, urinary ketone measurement and hourly capillary blood glucose (CBG) monitoring. Mean CBG from arrival to delivery was 4.7 +/- 1.1 mmol/l with 83% of mean CBG values within the target range (2.8-6.9 mmol/l). Mean CBG and insulin requirements were unrelated to therapy during pregnancy, but hypoglycaemia (CBG<2.8 mmol/l) was more frequent in women receiving insulin during pregnancy (40.7 vs 19.4 %, p<0.01). In several logistic regression models, CBG during labour was predictive of neonatal hypoglycaemia. We conclude that in women with GDM, the use of a standardised intrapartum management protocol is associated to fair metabolic control, that insulin requirements during labour are unrelated to therapy during pregnancy and that high CBG during labour increases the risk of neonatal hypoglycaemia. PMID- 11105968 TI - Low adherence of General Practitioners to National Cholesterol Education Program guidelines for the management of hyperlipidaemia. AB - Aim of our study was to assess adherence to the National Cholesterol Education Program Adult-Treatment Panel II (NCEP-ATP II) in patients cared for by General Practitioners (GPs) in an Italian community. The design of the work was cross sectional cohort study; the base was an unselected cohort of 1,168 patients cared for by GPs and screened at our lipid clinic in 1994-1995 in the Province of Turin (Italy). Blood samples were collected after 12-hr fast to measure plasma levels of total cholesterol, triglycerides, HDL-cholesterol, glucose and thyroid stimulating hormone (TSH). LDL-cholesterol was calculated using Friedewald's formula. In patients with body mass index (BMI) >30 kg/m2, an oral glucose tolerance test was performed. Blood pressure was measured in all patients, and a baseline ECG or a stress test was performed in those with unknown cardiovascular disease (CVD), then they were classified following the NCEP-ATP II criteria. Primary hyperlipidaemia accounted for 86.9% of the cohort with most patients requiring pharmacological treatment; in 34.4% of the patients, LDL-cholesterol values were > or = 6.46 mmol/l (250 mg/dl) and in 23.7% with established CVD, LDL cholesterol levels were > or = 5.68 mmol/l (220 mg/dl). In only 7.3% of patients the NCEP treatment goals were achieved, with 1.3% among those in secondary prevention. We observed great discrepancies between clinical practice and international recommendations for the management of hyperlipidaemia. PMID- 11105969 TI - Fibronectin and lipoprotein(a) are inversely related to plasminogen activator inhibitor type-1 levels in Type 2 diabetic patients without complications. AB - Plasminogen activator inhibitor type-1 (PAI-1), the most important physiological fibrinolysis inhibitor, is considered an independent factor of cardiovascular risk in Type 2 diabetes mellitus (T2DM). In previous papers we demonstrated that a T2DM population without complications presents: 1) PAI-1 not increased with respect to a control group; and 2) a negative correlation between PAI-1 and lipoprotein(a) [Lp(a)], suggesting that in these subjects PAI-1 levels could be modulated by the "endothelial stress" induced by Lp(a) and diabetes. This work has been performed in order to better verify this intriguing hypothesis, and the endothelial stress has been evaluated through a marker of endothelial damage, fibronectin (FNC). For this purpose we chose a T2DM population without complications (n=73) and a control group (n=46). Plasma concentrations of FNC, Lp(a), PAI-1 antigen and activity, and the main parameters of lipo- and glycometabolic balance were determined. Fibronectin was significantly higher in diabetics with respect to controls (p<0.01). As expected, significant correlation between PAI-1 antigen, PAI-1 activity and Lp(a) (r=-0.54,p<0.01 and r= 0.39,p<0.01, respectively) was found only in diabetic patients. In the same group FNC showed a significant correlation with PAI-1 antigen and activity (r= 0.49,p<0.01 and r=-0.47; p<0.01, respectively), while no relationship was found between Lp(a) and FNC. Multiple regression analysis showed statistically significant correlation between PAI-1 antigen and PAI-1 activity with FNC and Lp(a) in diabetic patients without complications (p<0.05). These data suggest that in absence of complications, the endothelium is able to modulate PAI-1 levels, favouring in that way the fibrinolytic pathway and, subsequently, the recovery of the endothelial integrity. This modulation seems to be related to parameters such as Lp(a) and FNC, although the mechanisms of the endothelial stress of these two molecules seem to be different. PMID- 11105970 TI - Relationship between patient practice-oriented knowledge and metabolic control in intensively treated Type 1 diabetic patients: results of the validation of the Knowledge and Practices Diabetes Questionnaire. AB - AIMS: To validate a newly developed questionnaire for the measurement of patients' knowledge and practices, with particular attention to its ability in predicting HbA1c levels. RESEARCH DESIGN AND METHODS: The Knowledge and Practices Diabetes Questionnaire (KPDQ) is a questionnaire composed of two scales, investigating patient knowledge and practices. Twenty-two questions, 12 dealing with patients' knowledge and 10 relative to patients' practices, were initially identified. Factor analysis and reliability analysis were used to validate the questionnaire. The ability of the two scales in predicting metabolic control was then evaluated. The questionnaire was administered to a population of Type 1 diabetic subjects intensively treated and regularly attending the diabetes outpatient clinic of Pescara General Hospital. The mean of all HbA1c measurements performed after patients were taken in charge by the clinic was used as an indicator of metabolic control. RESULTS: Out of 133 Type 1 patients identified, 77 (58 %) filled in the questionnaire. Respondents had a mean age (+/- SD) of 37 +/- 13 years and a mean diabetes duration of 13 +/- 9 years. The application of factor and reliability analyses led to the definition of two final scales composed of 10 (Knowledge Score, KS) and 5 items (Practice Score, PS), respectively. Item-scale correlation was > or = 0.40 for all the items investigated. Cronbach's alpha coefficient exceeded the value of 0.70 for both scales. The mean number of HbA1c determinations during a median period of observation of 4 years was of 11 +/- 5. The mean HbA1c value for the whole population was of 7.0% +/- 1.4, while the proportion of patients with values < or = 7.0% was of 57%. After adjusting for clinical and patient-related characteristics, the KS was the only independent predictor of metabolic control. Patients in the lowest quartile of the KS showed a more than 20-fold increased risk of having mean HbA1c values > or = 7.0% as opposed to those in the highest quartile (odds ratio, OR=23.3;p=0.009). No association emerged between metabolic control and PS. CONCLUSIONS: The KPDQ presents excellent psychometric properties. The KS also shows a very impressive association with the mean HbA1c values over a period of 4 years. These findings are particularly remarkable in that many studies have failed in documenting such a relationship. The KS can thus be considered as a quick and efficient screening tool to be used in an ambulatory setting to monitor the level of practice-oriented knowledge of patients with Type 1 diabetes as well as to identify those subjects who need individualized educational interventions. PMID- 11105971 TI - Plasma concentrations of leptin and selected minerals do not differ in Type 2 diabetic patients with or without sulfonylurea inefficacy. AB - The oral hypoglycemic agent sulfonylurea (SU) can increase plasma leptin concentrations. Additionally, diabetic subjects frequently have an altered plasma status of selected minerals. However, whether these described plasma parameters are changed in Type 2 diabetic patients who exhibit SU inefficacy has not yet been evaluated. In this preliminary study, fasting blood samples were collected from 16 Type 2 diabetic patients with secondary SU failure. As controls, 16 sex-, age- and adiposity-matched diabetic patients, who had similar diabetic duration and optimal glycemic control by SU, were also recruited. The results show that plasma values of leptin, C-peptide, calcium, magnesium, copper, and zinc did not significantly differ in these diabetic patients with or without secondary SU failure. However, the gender effect on plasma leptin level and the correlations between leptin and adiposity and C-peptide were retained. This study indicates that there is no relation between SU inefficacy and the plasma status of leptin and selected minerals. PMID- 11105973 TI - Vitamin A status in diabetic children. PMID- 11105972 TI - Endothelial nitric oxide in diabetes mellitus: too much or not enough? AB - Vascular complications are the leading cause of increased mortality in patients with diabetes mellitus. Endothelial dysfunction, characterised by impaired endothelium-dependent vasoreactivity, is the first sign of blood vessel damage that precedes morphological changes of the vessel wall. With other factors altered bioavailability of nitric oxide, the most potent endothelium-derived vasodilator, contributes to the changes in vascular tone and integrity. In addition to the impairment of vascular reactivity and permeability, other anti atherosclerotic functions of nitric oxide are also diminished, which may result in activated monocyte, leukocyte and platelet adhesion to the endothelium, increased platelet aggregation, lipoprotein influx to the subendothelial space and smooth muscle proliferation. Hyperglycaemia-induced increased oxidative stress and impairment of antioxidant defence are suggested to play a role in the pathomechanism of vascular damage, partly by influencing nitric oxide. Both in experimental and clinical Type 1 and Type 2 diabetes mellitus the apparently conflicting data of increased, unaltered or diminished nitric oxide action suggests a complex, time and localisation-dependent alteration of vascular function. The understanding of the mechanisms that lead to diabetic endothelial dysfunction and its early detection are necessary to establish appropriate intervention to prevent irreversible atherosclerotic vessel damage in patients with diabetes mellitus. PMID- 11105974 TI - The effects of whole-body restriction on task performance. AB - Many occupations, particularly involving maintenance operations, require individuals to perform both physical tasks and mental tasks in restricted spaces. Researchers have examined physical task performance under various restrictions; however, little research has investigated the effects of restricted space on cognitive tasks. Cognitive task performance in restricted spaces presents cognitive demands (i.e. the task itself) as well as additional physical demands (e.g. awkward postures), which may adversely affect task performance or operator workload. This research focused on the effects of whole-body restrictions on cognitive task performance. An experiment was conducted that examined 9 levels of restriction created in a laboratory: an unrestricted control, 6 single whole-body restrictions at two severity levels (2 lateral, 2 sagittal and 2 vertical) and 2 multiple restrictions (sagittal/vertical, lateral/sagittal/vertical). An inspection task served as the cognitive task. Behavioural, physiological and psychophysical measures were collected and analysed to measure the operator and performance effects. Operator response differences were found among the various forms of restriction as well as the severity level of similar forms of restriction. Increasing restriction significantly affected the behavioural and physiological operator response as opposed to the cognitive response. PMID- 11105975 TI - Stabilographic analysis of unconstrained standing. AB - Natural standing is characterized by postural changes and several hypotheses have been proposed to explain these changes. In this paper, four hypotheses were investigated by quantifying the number of postural changes in the centre of pressure data from unconstrained standing in different experimental conditions, studying the effects of mechanical loading, visual conditions, and type of support surface and sole of the shoes. The subjects stood for 30 min with no specific instructions other than not to step off a force plate. There were no significant effects on the number of centre of pressure patterns associated with the postural changes due to load, vision, surface and shoes during standing; on average, approximately two centre of pressure patterns per minute were observed in all conditions. The analysis of the centre of pressure data by the commonly used statistical parameters (standard deviation, velocity, and mean frequency of the centre of pressure displacement and area of the stabilogram) also did not reveal any effect of the different conditions. PMID- 11105976 TI - Performance with tables and graphs: effects of training and a Visual Search Model. AB - After more than 70 years of research it is still not clear under what conditions graphic presentations of information have an advantage over tables. A minimum assumption Visual Search Model (VSM) was designed to predict the performance of various tasks with tables and graphs that show data with different levels of complexity. An experiment tested the performance of five tasks with tables, bargraphs and line-graphs, showing data with various levels of complexity, over the course of nine experimental sessions in order to assess possible changes in the relative efficiency of the displays after practice. Tables had an initial advantage over graphs for all tasks, and there were complex interactions between the variables. The initial differences between the displays disappeared for some tasks after users gained experience with the displays, while for other tasks the differences continued to exist even after extended practice. The VSM predicted the results for tables well. For graphs the model was adequate for tasks that involve single data points, such as reading values or comparing pairs of values. The performance of tasks that require the analysis of data configurations, such as reading a trend, could not be predicted with the VSM. Hence the VSM can predict task performance with tables and graphs for low-integration tasks. PMID- 11105977 TI - Prospective validation of a low-back disorder risk model and assessment of ergonomic interventions associated with manual materials handling tasks. AB - The evaluation of low-back disorder risk associated with materials handling tasks can be performed using a variety of assessment tools. Most of these tools vary greatly in their underlying logic, yet few have been assessed for their predictive ability. It is important to document how well an assessment tool realistically reflects the job's injury risk, since only valid and accurate tools can reliably determine whether a given ergonomic intervention will result in a future reduction in back injuries. The goal of this study was to evaluate how well a previously reported low-back disorder (LBD) risk assessment model (Marras et al. 1993) could predict changes in LBD injury rates as the physical conditions to which employees are exposed were changed. Thirty-six repetitive materials handling jobs from 16 different companies were included in this prospective cohort study. Of these 36 jobs, 32 underwent an ergonomic intervention during the observation period, and four jobs in which no intervention occurred served as a comparison group. The trunk motions and workplace features of 142 employees performing these jobs were observed both before and after workplace interventions were incorporated. In addition, the jobs' LBD rates were documented for these pre and post-intervention periods. The results indicated that a statistically significant correlation existed between changes in the jobs' estimated LBD risk values and changes in their actual low-back incidence rates over the observation period. Linear and Poisson regression models also were developed to predict a change in a job's incidence rate and the number of LBD on ajob respectively, as a function of the job's risk change using this assessment model. Finally, this prospective study showed which ergonomic interventions consistently reduced the jobs' mean low-back incidence rates. These results support use of the LBD risk model to assess accurately a job's potential to lead to low-back injuries among its employees. PMID- 11105978 TI - Off-road machine controls: investigating the risk of carpal tunnel syndrome. AB - Occupationally induced hand and wrist repetitive strain injuries (RSI) such as carpal tunnel syndrome (CTS) are a growing problem in North America. The purpose of this investigation was to apply a modification of the wrist flexion/ extension models of Armstrong and Chaffin (1978, 1979) to determine if joystick controller use in off-road machines could contribute to the development of CTS. A construction equipment cab in the laboratory was instrumented to allow force, displacement and angle measurements from 10 operators while they completed an approximately 30-min joystick motion protocol. The investigation revealed that both the external fingertip and predicted internal wrist forces resulting from the use of these joysticks were very low, indicating that the CTS risk associated with this factor was slight. However, the results also indicated that, particularly for the 'forward' and 'left' right side motions and for all left side motions, force was exerted by other portions of the fingers and hand, thereby under-predicting the tendon tension and internal wrist forces. Wrist angles observed were highest for motions that moved the joysticks to the sides rather than front to back. Thus, the 'right' and 'left' motions for both hands posed a higher risk for CTS development. When the right hand moved into the 'right' position and the left hand moved into the 'left' position, the wrist went into extension in both cases. Results indicate that neither learning nor fatigue affected the results. PMID- 11105979 TI - Muscular rest and gap frequency as EMG measures of physical exposure: the impact of work tasks and individual related factors. AB - Owing to an orderly recruitment of motor units, low threshold type I fibres are presumed to be vulnerable in contractions of long duration. To study load on these fibres muscular rest was registered as the time fraction of electromyographic (EMG) activity below a threshold. Moreover, the frequency of periods with muscular rest, EMG gaps, was derived, since a low gap frequency has been shown to be a risk factor for musculoskeletal disorders. Trapezius EMG was registered in 24 female hospital cleaners, 21 female office workers and 13 male office workers during one working day. Cleaners have a high risk of neck/shoulder pain and had much less muscular rest than office workers measured as a percentage of total registered time (median value = 1.5%, range = 0.2-13% vs. median value = 12%, range = 0.0-32%, respectively). Gap frequency showed no difference between the two occupational groups. Both measures displayed a wide inter-individual variation. For the cleaners, some of the variance was explained by body mass index (BMI) and age, with lower values of muscular rest for older subjects with a high BMI. Among the office workers, low values of muscular rest and a high gap frequency were registered in subjects with a low subjective muscular tension tendency. Gender, strength, smoking, job strain, employment time and musculoskeletal symptoms had no impact on either EMG measure. PMID- 11105980 TI - Assessing noise annoyance: an improvement-oriented approach. AB - A concept for practice-oriented assessment of noise annoyance at the workplace is presented. Employees evaluated the noise situation at their workplace by characterizing the loudest noise event with respect to relevant noise characteristics. The results from a first use of the questionnaire for Subjective Evaluation of Noise Characteristics in Office Workplaces (SENO) show (1) a general need for an additional constructive measure of subjective noise annoyance, (2) that evaluation of the loudest noise event is representative for the overall workplace situation, and (3) that coping plays a crucial role and should be explicitly controlled. Finally, examples of how to use SENO for improving the workplace situation are given. PMID- 11105981 TI - 'Ergonomics in the Nordic European countries--a historical perspective': paper presented at IEA 2000/HFES 2000 Congress. International Ergonomics Association. AB - The Nordic Ergonomic Society was founded more than 30 years ago to represent Sweden, Finland, Norway and Denmark. Recently Iceland has also been included. The Nordic Ergonomics Society has, to a large extent, traditionally been oriented towards work in the area of physiology, and a large number of its members have backgrounds within such areas as work physiology, physiotherapy and rehabilitation. However, from its inception the society has had members who are experienced within, such fields as work psychology, design, engineering and occupational health and safety. Over the last decade we have also had new members from such areas as work sociology, organizational psychology, leadership, training, information technology and cybernetics. Members have been concerned mainly with the application and practice of ergonomics. The number of members involved in research has also increased over the years. The principal areas of application for such research have been within industry and government. PMID- 11105982 TI - Cytotoxic effect of gossypol on colon carcinoma cells. AB - Gossypol, a male contraceptive drug extracted from cottonseeds, has been found to have antiproliferative activity on tumour cells and is thought to be a potential anticancer drug. The aim of this study was to investigate the mechanisms of gossypol-induced cell death on two colon carcinoma cell lines, HT29 and LoVo. Firstly, we studied the effect of gossypol on the colony forming ability of these tumour cells, which is the main target of chemotherapeutic drugs. Using clonogenic assays, flow cytometry and DNA gel electrophoresis techniques, we have found that gossypol not only inhibited colony forming ability of these tumour cells, but we also observed cellular internucleosomal DNA fragmentation in the cells treated with 3 doses of gossypol and this was accompanied by the appearance of a sub-G1 apoptotic peak and morphological characteristics of apoptosis. Our results suggest that the gossypol induced cell death is via an apoptotic pathway and the effect of gossypol may not be cell cycle specific. Using Western blotting analysis, we found that the gossypol-induced apoptosis may not be involved in the regulation of p53 but possibly associated with the regulation of bcl-2 and Bax expression. Our evidence indicates that gossypol may provide a potential therapeutic benefit for the treatment of colon carcinoma and understanding the mechanisms of gossypol-induced cytotoxicity on tumour cells is essential for including this drug in clinical use. PMID- 11105983 TI - Neuroprotective effects of Ginkgo biloba extract in brain ischemia are mediated by inhibition of nitric oxide synthesis. AB - We studied the effects of pre-treatment (15 days) with oral administration of Ginkgo biloba extract (Ph-Gb 37.5-150 mg/kg) on brain malonildialdehyde (MDA), brain edema, brain nitrite and nitrate and delayed neuronal death following transient cerebral ischemia in the Mongolian gerbil. Survival was not modified, however, pre-treatment with Ginkgo biloba significantly and in a dose-dependent way reduced post-ischemic brain MDA levels and post-ischemic brain edema. Delayed neuronal death in the CA1 of the hippocampus was attenuated by the highest dose of the extract. Increase of nitrite and nitrate was observed after cerebral ischemia in the hippocampus and it was dose-dependently reduced in animals pretreated with Ph-Gb, thus suggesting that neuroprotective effects of Ginkgo biloba may be due to an inhibitory action on nitric oxide formation. PMID- 11105984 TI - [125I]-galanin binding sites in the human nodose ganglion. AB - The cell bodies of centrally-projecting vagal afferent neurons are contained in the inferior vagal (nodose) ganglion. Although binding sites for a number of different neuropeptides/modulators have been detected in the human nodose ganglion, the presence of galanin binding sites has not been reported. In vitro receptor autoradiography using [125I]-galanin enabled visualisation of binding sites for galanin in the human nodose ganglion. The presence of such binding sites suggests a potential role for galanin in the neuromodulation of vagal transmission in humans. PMID- 11105985 TI - Effects of acute alcohol intoxication on growth axis in human adolescents of both sexes. AB - We previously reported the deleterious effects of acute alcohol intoxication (AAI) on pituitary-gonadal and pituitary-adrenal axes hormones in human adolescents. In the present paper we studied the effects of AAI on the growth axis hormones, and the possible contribution of the insulin-glucose axis to the alcohol-induced dysfunction of the growth axis in human adolescents. Blood samples were drawn from adolescents that arrived at the emergency department with evident behavioural symptoms of drunkenness (AAI) or with nil consumption of alcohol (controls [C]). AAI produced in the adolescents of both sexes in our series: a decrease in growth hormone (GH) levels, without significant alteration of either insulin-like growth factor-I (IGF-I) or insulin-like growth factor binding protein-3 (IGFBP3); an increase in plasma glucose and a decrease in insulin in the female adolescents but not in the males. Males and females undergo a significant period of bone growth during adolescence. Growth axis hormones play an important role in the pubertal spurt. Thus, ethanol consumption during adolescence could have long-lasting deleterious effects on this aspect of development. In industrialised countries, around 35% of alcohol drinkers are under 16 years old, therefore the result of this study should be made known to adolescents and the appropriate authorities. PMID- 11105986 TI - Brain-to-blood efflux transport of estrone-3-sulfate at the blood-brain barrier in rats. AB - Efflux transport of estrogens such as estrone-3-sulfate (E1S), and estrone (E1) across the blood-brain barrier (BBB) was evaluated using the Brain Efflux Index (BEI) method. The apparent BBB efflux rate constant (Keff) of [3H]E1S, and [3H]E1 was 6.63 x 10(-2) +/- 0.77 x 10(-2) min(-1), and 6.91 x 10(-2) +/- 1.23 x 10(-2) min(-1), respectively. The efflux transport of [3H]E1S from brain across the BBB was a saturable process with Michaelis constant (Km) of 96.0 +/- 34.4 microM and 93.4 +/- 22.0 microM estimated by two different methods. By determining [3H]E1S metabolites using high performance liquid chromatography (HPLC) after intracerebral injection, significant amounts of [3H]E1S were found in the jugular venous plasma, providing direct evidence that most of [3H]E1S is transported from brain across the BBB in intact form. To compare the apparent efflux clearance across the BBB of E1S with that of E1, the brain distribution volume of E1S and E1 was estimated using the brain slice uptake method. The apparent efflux clearance of [3H]E1S was determined to be 74.9 +/- 3.8 microl/(min x g brain) due to the distribution volume of 1.13 +/- 0.06 ml/g brain. By contrast, the apparent efflux clearance of E1 was more than 227 +/- 3 microl/(min x g brain), since the distribution volume of [3H]E1 at 60 min was 3.28 +/- 0.13 ml/g. The E1S efflux transport process was inhibited by more than 40% by coadministration of bile acids (taurocholate, and cholate), and organic anions (sulfobromophthalein, and probenecid), whereas other organic anions did not affect the E1S efflux transport. The [3H]E1S efflux was significantly reduced by 48.6% after preadministration of 5 mM dehydroepiandrosterone sulfate. These results suggest that E1S is transported from brain to the circulating blood across the BBB via a carrier-mediated efflux transport system. PMID- 11105987 TI - Effects of mifepristone (RU486) treatment on the development of uterine adenomyosis induced by pituitary grafting in mice. AB - To evaluate the effects of mifepristone (RU486) on the development of uterine adenomyosis induced by pituitary grafting (PG), 3 groups of mice receiving pituitary grafts at 7 weeks of age were given RU486 in food (20 mg/kg chow) from 3-14 (RU486-3 group) or 10-14 (RU486-10 group) weeks of age, or were given no further treatment (PG control group), respectively. All the mice were killed at 14 weeks of age. The uterine weight was significantly decreased in both RU486 treated groups compared with the PG control group. The incidence of adenomyosis was also decreased significantly in both the RU486-3 group (0/10 mice) and RU486 10 group (2/10 mice) compared with the PG control group (7/9 mice). To look for vascular changes in the uterine tissues, which have been reported to be related to the development of adenomyosis, immunohistochemical staining of von Willebrand factor in the blood vessels was performed. The mean surface area and minor axis of blood vessels in the uterus were thereby found to be significantly decreased in the RU486-10 group compared to the PG control group. The results clearly indicated that RU486, a potent antiprogestin, could inhibit the genesis of uterine adenomyosis in mice, and at the same time caused shrinkage of the vascular system. As in humans, progesterone as well as the vascular system therefore appear to be important factors in the pathogenesis of uterine adenomyosis in this mouse model. PMID- 11105988 TI - TNF-alpha, leptin, and lymphocyte function in human aging. AB - Aging is associated with increased inflammatory activity and concomitant decreased T cell mediated immune responses. Leptin may provide a link between inflammation and T cell function in aging. The aim of the study was to investigate if plasma levels of tumor necrosis factor (TNF)-alpha were associated with leptin, circulating interleukin-2 receptors (sIL-2R), and phytohaemagglutinin (PHA) induced IL-2 production in whole blood in elderly humans. Circulating levels of TNF-alpha and sIL-2R were higher in elderly humans (N=42) compared to a young control group (N=37) whereas there was no difference with regard to IL-2 production. Furthermore, there were no age-related differences in serum levels of leptin. However, women had higher levels than men. In the elderly people, serum levels of leptin were correlated with TNF-alpha in univariate regression analysis and in a multiple linear regression analysis adjusting for the effect of gender and body mass index. Furthermore, TNF-alpha, but not leptin, was positively correlated to sIL-2R and negatively correlated to IL-2 production. In conclusion, increased plasma levels of TNF-alpha in aging is associated with poor IL-2 production ex vivo and lymphocyte activation in vivo. These associations do not seem to involve leptin. PMID- 11105989 TI - Activation of G-proteins in the mouse pons/medulla by beta-endorphin is mediated by the stimulation of mu- and putative epsilon-receptors. AB - The activation of mu-, delta- and kappa1-opioid receptors by their respective agonists increases the binding of the non-hydrolyzable GTP analog guanosine-5' (gamma-thio)-triphosphate (GTPgammaS) to G proteins. Beta-endorphin is an endogenous opioid peptide which binds nonselectively to mu-, delta- and putative epsilon-opioid receptors. The present experiment was designed to determine which opioid receptors are involved in the stimulation of [35S]GTPgammaS binding induced by beta-endorphin in the mouse pons/medulla. The mouse pons/medulla membranes were incubated in an assay buffer containing 50 pM [35S]GTPgammaS, 30 microM GDP and various concentrations of beta-endorphin. Beta-endorphin (0.1 nM 10 microM) increased [35S]GTPgammaS binding in a concentration-dependent manner, and 10 microM beta-endorphin produced a maximal stimulation of approximately 260% over baseline. This stimulation of [35S]GTPgammaS binding by beta-endorphin was partially attenuated by the mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA), but not by the delta-opioid receptor antagonist naltrindole (NTI) or the kappa1-opioid receptor antagonist nor-binaltorphimine (nor-BNI). Beta endorphin stimulated [35S]GTPgammaS binding by about 80% in the presence of 10 microM beta-FNA, 30 nM NTI and 100 nM nor-BNI. The same concentrations of these antagonists completely blocked the stimulation of [35S]GTPgammaS binding induced by 10 microM [D-Ala2,NHPhe4,Gly-ol]enkephalin, [D-Pen(2,5)]enkephalin and U50,488H, respectively. Moreover, the residual stimulation of [35S]GTPgammaS binding induced by beta-endorphin in the presence of the three opioid receptor antagonists was significantly attenuated by 100 nM of the putative epsilon-opioid receptor partial agonist beta-endorphin (1-27). These results indicate that the stimulation of [35S]GTPgammaS binding induced by beta-endorphin is mediated by the stimulation of both mu- and putative epsilon-opioid receptors in the mouse pons/medulla. PMID- 11105990 TI - First demonstration of an inhibitory activity of milk proteins against human immunodeficiency virus-1 reverse transcriptase and the effect of succinylation. AB - A variety of milk proteins including lactoferrin, angiogenin-1, alpha lactalbumin, beta-lactoglobulin, lactoperoxidase, casein and the novel whey proteins lactogenin and glycolactin were tested for inhibitory activity toward human immunodeficiency virus-1 reverse transcriptase (HIV-1 RT), alpha glucosidase, beta-glucosidase and beta-glucuronidase. Lactoferrin exerted the most potent inhibitory action with an IC50 of about 6 microM. Lactoperoxidase, lactogenin, angiogenin-1 and glycolactin inhibited HIV-1 RT activity with decreasing potencies. Beta-lactoglobulin, alpha-lactalbumin and casein displayed little or no inhibitory effect. Succinylation with succinic anhydride augmented the inhibitory effect of glycolactin, beta-lactoglobulin, alpha-lactalbumin, casein and human lactoferrin. The inhibitory effect of the various milk proteins on the activities of alpha-glucosidase, beta-glucosidase and beta-glucuronidase was meager. Succinylation tended to increase the alpha-glucosidase-inhibitory effect of milk proteins but neither their beta-glucosidase-inhibitory nor beta glucuronidase-inhibitory effect was affected. PMID- 11105992 TI - Interaction of lidocaine with the cardiac sodium channel: effects of low extracellular pH are consistent with an external blocking site. AB - Brief extracellular application of millimolar concentrations of lidocaine affected sodium currents recorded in isolated rat ventricular myocytes in two ways: 1) a reduction of the maximum current consistent with a channel blocking action, and 2) a negative shift in the voltage dependence of inactivation consistent with an interaction with the inactivated state of the channel. Both effects occurred very rapidly (<< 1 s). Decreasing extracellular pH to 6.4 increased the potency for channel block (EC50 1.8 +/- 0.2 mM at pH 7.4 and 0.8 +/ 0.1 mM at pH 6.5) and decreased the potency to shift inactivation (V(1/2) shift 42 mV by 1 mM lidocaine at pH 7.4 and -12.6 mV at pH 6.5). Channel block was slightly less at +90 mV compared to -40 mV at either pH (not statistically significant). The increase in potency for block at decreased extracellular pH, while intracellular pH is buffered, and the lack of voltage dependence of block, suggest that the charged form of lidocaine can block the channel by interacting with a site near the extracellular mouth, although alternative explanations are discussed. PMID- 11105991 TI - Effects of salmeterol on muscarinic inhibition of adenylyl cyclase in bovine trachealis cells. AB - The goal was to assess whether salmeterol, a potent and long-acting beta-2 adrenergic agonist used in the treatment of asthma, also has non-beta-2 adrenergic effects on the stimulation or inhibition of adenylyl cyclase activity. Salmeterol (100 nM) maximally stimulated cAMP accumulation in enzyme dispersed bovine trachealis cells and this was entirely inhibited by propranolol, as expected for beta-adrenergic stimulation. However, the same concentration of salmeterol also antagonized carbachol inhibition of cAMP accumulation and altered binding of carbachol to muscarinic receptors. These effects of salmeterol were sensitive to washing of the cells and this was not consistent with a beta-2 adrenergic mechanism. The findings suggested that the maximal, beta-2-adrenergic stimulation of cAMP accumulation by salmeterol was accompanied by a non-beta-2 adrenergic interaction of salmeterol with muscarinic receptors that attenuated muscarinic inhibition of adenylyl cyclase. PMID- 11105993 TI - Pranlukast protects leukotriene C4- and D4-induced epithelial cell impairment of the nasal mucosa in vitro. AB - To investigate the effect of pranlukast on leukotriene- induced airway mucosal epithelial dysfunction, samples of human nasal mucosa obtained during surgery for facial trauma were exposed to leukotriene C4 and/or D4 and observed on a TV screen magnified x 2,500. Leukotriene C4- and D4-induced ciliary inhibition and delayed mucosal surface alterations appeared several hours later. Pranlukast prevented both the mucosal epithelial cell dysfunction and the delayed epithelial cell alteration. PMID- 11105994 TI - Hepatic histidase and muscle branched chain aminotransferase gene expression in experimental nephrosis. AB - Protein and amino acid metabolism is altered during nephrotic syndrome. However, the expression of the amino acid degrading enzymes has not been well studied. The objective of this work was to assess the expression of hepatic histidase (Hal) and skeletal muscle mitochondrial branched chain amino transferase (BCATm) in rats with experimental nephrotic syndrome induced by a single injection of puromycin aminonucleoside (150 mg/kg). Six days after the injection rats were killed and hepatic Hal and skeletal muscle BCATm activities were measured. Also, total mRNA from both tissues was isolated and Hal and BCATm mRNA expression were analyzed by Northern blot. Rats with NS showed a reduction in food intake with respect to the control group. Hepatic Hal activity increased significantly in nephrotic and pair fed rats by 59% compared to control group. This change in activity was associated with a corresponding increase in Hal mRNA abundance. On the other hand, skeletal muscle BCATm activity and mRNA abundance were similar in the three groups studied. These results suggest that the increase in Hal expression was associated with the reduced food intake and not to the NS. However, BCAT expression did not change indicating the importance of BCAA in body nitrogen conservation. PMID- 11105995 TI - Occurrence of orally administered curcuminoid as glucuronide and glucuronide/sulfate conjugates in rat plasma. AB - Curcuminoids, curcumin and its structurally related compounds, constitute the phenolic yellowish pigment of turmeric. We investigated the absorption and metabolism of orally administered curcuminoids (curcumin, demethoxycurcumin and bisdemethoxycurcumin) in rats. HPLC and LC-MS analyses after enzymatic hydrolyses showed that the predominant metabolites in plasma following administration were glucuronides and glucuronide/sulfates (conjugates with both glucuronide and sulfate) of curcuminoids. The plasma concentrations of conjugated curcuminoids reached a maximum one hour after administration. The conjugative enzyme activities for glucuronidation and sulfation of curcumin were found in liver, kidney and intestinal mucosa. These results indicate that orally administered curcuminoids are absorbed from the alimentary tract and present in the general blood circulation after largely being metabolized to the form of glucuronide and glucuronide/sulfate conjugates. PMID- 11105996 TI - Ethanol inhibits IgE-induced degranulation and cytokine production in cultured mouse and human mast cells. AB - Activated mast cells (MC) can produce a wide variety of potent inflammatory mediators. Excessive alcohol consumption is known to lead to immune deficiency and propensity for pneumonias in particular. As MCs are important in the first line of defence of mucosal membranes we have studied the effect of ethanol (EtOH) on several MC functions. EtOH attenuated dose dependently IgE-induced degranulation of mouse bone marrow derived mast cells (mBMMC) as reflected by the release of granule associated beta-hexosaminidase (beta-hex). A mean of 26 +/- 7% inhibition of beta-hex release was observed in the presence of 5/1000 (86 mM) EtOH and nearly complete inhibition in the presence of 20/1000 (344 mM) ethanol. The IgE-induced degranulation of mBMMC cultured with EtOH for seven days was inhibited to a similar degree as the degranulation of mBMMC exposed to EtOH for only one hour. Inclusion of 5/1000 (86 mM) ethanol in the medium reduced tumour necrosis factor (TNF)-alpha and interleukin (IL)-8 production in human mast cell line (HMC-1) cells by 55 +/- 7% and 19 +/- 5%, respectively, and the presence of 20/1000 (344 mM) ethanol inhibited the expression 81 +/- 12% and 59 +/- 14% respectively. These results suggest that, in contrast to previous assumption, ethanol inhibits several critical MC functions at least in vitro. This inhibition of mediator, and cytokine release in particular, could contribute to the immune deficiency associated with chronic alcohol consumption. PMID- 11105997 TI - Characterization of a platelet activating factor acetylhydrolase from rat adipocyte. AB - Platelet activating factor-acetylhydrolases (PAF-AHs) are a family of enzymes with the common property of hydrolyzing and inactivating PAF and thus regulating its levels. In the course of studying the role of PAF in rat adipocytes and its possible implication in body weight regulation and immune response, conditions in which adipocytes are involved, we investigated the existence of PAF-AH in these cells. We detected PAF-AH activity in rat adipocytes which is mainly distributed in the cytosol. The behaviour of the enzyme during hydrophobic chromatography, together with the fact that part of the enzyme activity was found in the fat cake of adipocyte homogenate suggests the hydrophobic nature of rat adipocyte PAF-AH. The enzyme activity was distinct from the Ca2+-dependent and independent phospholipase A2, the lysophospholipase, the lecithin:cholesterol acyltransferase (LCAT) and from non-specific acetylhydrolases. We partially purified PAF-AH from rat adipocyte cytosolic fraction and the purified enzyme revealed a major protein band of 66 kDa and a minor one of 37 kDa on SDS/PAGE. The purified PAF-AH has an apparent Km value of 4.9 microM and the enzyme activity was inactivated by PMSF and 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and moderately stimulated by dithiothreitol (DTT). Furthermore, in this study we identified PAF in rat adipocytes and determined its concentration. PMID- 11105998 TI - Effects of experimental hypothyroidism on the development of the hypothalamo pituitary-adrenal axis in the rat. AB - Hypothyroid pups were obtained by adding methimazole in the mother's drinking water from day 15 of gestation and sacrificed at 4, 8 or 15 days. Circulating corticosterone decreased at all ages, while CBG concentrations diminished at day 4, increased at day 8 and did not change at day 15 in hypothyroid rats. As opposed to controls, plasma ACTH concentrations decreased steadily with age while there was an accumulation of ACTH in the anterior pituitary of hypothyroid 15-day old rats. Anterior pituitary POMC contents were unaffected by the treatment. In the hypothalamic PVN, CRF mRNA levels in the total population of CRF-synthesizing cells and in the CRF+/AVP+ subpopulation were below those of controls whatever the age considered while AVP mRNA in the CRF+/AVP+ subpopulation did not change at day 4 and decreased at day 8 and 15 in hypothyroid animals. Both the number of cell bodies expressing detectable levels of CRF mRNA and the percentage of CRF and AVP colocalization decreased at day 4 and were unchanged thereafter. CRF and AVP immunoreactivity in the zona externa of the median eminence increased with age but was not affected by methimazole treatment. The concentration of AVP mRNA in the magnocellular cell bodies of the PVN and the SON as well as AVP immunoreactivity in the zona interna of the median eminence were not changed by the treatment at days 4 and 8. In hypothyroid 15-day-old rats, SON AVP mRNA increased, AVP immunoreactivity decreased while plasma osmolality was enhanced. In conclusion, our data demonstrate that experimental hypothyroidism impairs specifically the maturation of hypothalamic parvocellular CRF and AVP gene expression during the stress hyporesponsive period. These observations suggest that the physiological peak in plasma thyroxine concentrations that occur between day 8-12 may participate in the maturation of hypothalamic CRF- and AVP synthesizing cells. PMID- 11105999 TI - Effect of fumonisin B1 on inducible nitric oxide synthase and cyclooxygenase-2 in LPS-stimulated J774A.1 cells. AB - Fumonisin B1 (FB1) is a water-soluble fungal metabolite that elicits a wide spectrum of toxicological effects. Cellular targets of FB1 include immune cells and in particular macrophages. In the present study the cytotoxic effect of FB1 (1-100 microM) was evaluated using a murine macrophage cell line (J774A.1) as model system. The effect of FB1 on nitric oxide (NO) and prostaglandin E2 (PGE2) production induced by lipopolysaccharide (LPS, 10 and 100 ng/ml) was also investigated. Macrophages were pretreated with FB1 for 72 h and then stimulated with LPS for 24 h. The increase of LPS-induced production of these inflammatory mediators was observed at increasing concentrations of FB1 (0.1-10 microM) and was found to be concentration dependent. By western blot analysis we demonstrated that the observed increase of NO and PGE2 production by FB1 was related to an enhancement of iNOS and COX-2 expression. PMID- 11106000 TI - Differential inhibitory effects of sophoricoside analogs on bioactivity of several cytokines. AB - Effects of sophoricoside and its analogs on proinflammatory cytokines have been investigated. Sophoricoside, genistein and orobol exhibited inhibitory effects on IL-5, IL-3, GM-CSF and IL-6 bioactivities. Genistin showed inhibitory effects on IL-5 and IL-3 bioactivities, but did not inhibit GM-CSF and IL-6 bioactivities. None of the sophoricoside analogs showed inhibitory effects on both IL-1beta and TNF-alpha bioactivities. Among the compounds, sophoricoside exhibited the highest inhibitory effects on IL-5, IL-3 and IL-6 bioactivities with IC50 values of 1.9 microM, 6.9 microM and 6.0 microM, respectively and orobol did show on GM-CSF bioactivity with an IC50 value of 18.0 microM. The result would provide an additional mechanism by which the compounds exert immunosuppressive and anti inflammatory effects. PMID- 11106001 TI - The effect of the antipsychotic drug mosapramine on the expression of Fos protein in the rat brain: comparison with haloperidol, clozapine and risperidone. AB - In this study, we examined the effect of the acute p.o. administration of the antipsychotic drug mosapramine, as well as the antipsychotic drugs clozapine, haloperidol and risperidone, on the expression of Fos protein in the medial prefrontal cortex, nucleus accumbens and dorsolateral striatum of rat brain. The administration of mosapramine (1 or 3 mg/kg) significantly increased the number of Fos protein positive neurons in the medial prefrontal cortex, but not in the dorsolateral striatum. In addition, mosapramine (1, 3 or 10 mg/kg) produced a dose-dependent increase in the number of Fos protein positive neurons in the nucleus accumbens. The acute administration of 10 mg/kg of mosapramine significantly increased the number of Fos protein positive neurons in all brain regions. The acute administration of clozapine (30 mg/kg), similarly to mosapramine at lower doses (1 or 3 mg/kg), significantly increased the number of Fos protein positive neurons in the medial prefrontal cortex and nucleus accumbens, but not dorsolateral striatum. In contrast, haloperidol (0.3 mg/kg) significantly increased the number of Fos protein positive neurons in the nucleus accumbens and dorsolateral striatum, but not medial prefrontal cortex. The acute administration of risperidone (0.3 or 1 mg/kg) did not affect the number of Fos protein positive neurons in the medial prefrontal cortex, nucleus accumbens or dorsolateral striatum of rat brain, whereas a 3 mg/kg dose of risperidone significantly increased the number of Fos protein positive neurons in all brain regions. These results suggest that the ability of mosapramine to enhance expression of Fos protein in the medial prefrontal cortex may contribute to a clozapine-like profile with respect to actions on negative symptoms in schizophrenia. Furthermore, the lack of effect of low doses of mosapramine on Fos protein expression in the dorsolateral striatum, an area believed to play a role in movement, suggests that it may have a lower tendency to induce neurological side effects. PMID- 11106002 TI - The effect of suramin on bleomycin-induced lung injury. AB - Since transforming growth factor beta (TGF-beta) is presumed to play a role in lung fibrosis, we evaluated the effect of suramin (Sur), a substance with an anti TGF-beta effect, in vivo on bleomycin (Bleo)-induced pulmonary injury in mice and in vitro on human lung fibroblasts. Four groups of C57BL/6 mice each received one of four treatments: (1) intratracheal (i.t.) instillation of Bleo and intraperitoneal (i.p.) injections of Sur, every other day, starting one day before i.t. instillation of Bleo (Bleo-Sur); (2) i.t. Bleo and i.p. injections of saline (Bleo-Sal); (3) i.t. saline and i.p. Sur (Sal-Sur); and (4) i.t. and i.p. saline (Sal-Sal). Animals were sacrificed 14 days after i.t. treatment. Lung injury was evaluated by analysis of bronchoalveolar lavage (BAL) fluid, histologically by the semiquantitative morphological index, and biochemically by analysis of lung hydroxyproline content. In vitro, Sur did not affect TGF-beta induced increase of alpha1 (I) collagen mRNA in human lung fibroblasts. In vivo treatment of mice with Sur did not affect Bleo-induced lung injury. These results indicate that despite its potential anti TGF-beta and lymphocytotoxic effects, Sur is not a therapeutic candidate drug for rescue of lung fibrosis. PMID- 11106003 TI - Previous anesthesia can temporarily overshadow the expression of a withdrawal syndrome in opiate dependent rats. AB - We hypothesized that induction of opiate antagonist-precipitated withdrawal under anesthesia can decrease the expression of later withdrawal signs. Three groups of morphine-dependent rats were compared in different experimental conditions of withdrawal precipitation using naloxone. We showed that anesthesia can temporarily overshadow the expression of withdrawal signs, but that some signs can be delayed and increased in intensity. This can be explained by a parallel and temporary effect of anesthesia on arousal and pain threshold. This carries important implications on the use of anesthesia in detoxification procedures. PMID- 11106004 TI - A comparison of human immunodeficiency virus type-1 protease inhibition activities by the aqueous and methanol extracts of Chinese medicinal herbs. AB - The aqueous and methanol extracts of thirty-one herbs traditionally used as anti fever remedies in China were screened for their in vitro inhibition on human immunodeficiency virus type-1 protease (HIV-1 PR). The activity of recombinant HIV-1 protease was determined by sequence-specific cleavage at the Tyr-Pro bond of the fluorogenic substrate (Arg-Glu(EDANS)-Ser-Gln-Asn-Tyr-Pro-Ile-Val-Gln Lys(DABCYL)- Arg) or by HPLC anaylsis of the cleavage products after incubation of the enzyme with a synthetic peptide substrate (Acetyl-Ser-Gln-Asn-Tyr-Pro-Val Val-amide). Among the herbal extracts examined, the aqueous extracts of Prunella vulgaris and Scutellaria baicalensis and the methanol extracts of Woodwardia unigemmata, Paeonica suffruticosa and Spatholobus suberectus elicited significant inhibition (>90%) at a concentration of 200 microg/ml. PMID- 11106005 TI - Extended frequency range measurements for determining the Kneser-type acoustic relaxation time. AB - In the present paper, the authors discuss studies carried out for many years dealing particularly with two compounds: benzene and carbon disulphide and compare them with the results obtained by numerous acoustics researchers. These compounds are typical liquids in which acoustic Kneser-type relaxation occurs, caused by an irreversible vibrational and translational (VT) transition. Since magnitudes describing the relaxation process were diverse in many papers, we have undertaken an attempt to clarify these differences and to indicate how to avoid errors resulting from instrumental imperfections and the disregard of the considerable measurement errors when investigating velocity dispersion in the hypersonic (GHz) range. The results of these researches changed the interpretation of previous papers. PMID- 11106006 TI - Measurement of the effect of high-power ultrasound on wetting of paper. AB - High-power ultrasound has been known to promote penetration of liquids into porous materials. This work presents results of experimental investigations of the influence of 40 kHz high-power ultrasound on wetting processes of papers. Wetting was monitored by the measurement of the attenuation of 0.5-9 MHz ultrasound transmitted through the immersed sample. The samples were sized papers with contact angles 40 degrees, 70 degrees and 110 degrees, and the immersion liquids were isopropanol-water mixtures of 0-100% isopropanol concentration. The investigation showed that application of high-power ultrasound resulted in faster sorption processes. However, the absolute reduction in time to reach the stationary state was not very great. PMID- 11106007 TI - Application of adaptive time-frequency decomposition in ultrasonic NDE of highly scattering materials. AB - In the paper, adaptive time-frequency decomposition by basis pursuit (BP) is utilized to improve ultrasonic flaw detection in highly-scattering materials as an alternative to the Wavelet Transform technique. The detection of ultrasonic pulses using the BP is described. Computer simulation was performed to verify the signal detection improvements for an ultrasonic wave embodied in white noise, and numerical results show good detection even for signal-noise ratio (SNR) of -18 dB. The improvement in detection is experimentally verified using cast steel samples with artificial flaws. PMID- 11106008 TI - Generation of very high pressure pulses at the surface of a sandwiched piezoelectric material. AB - New clinical concepts in lithotripsy demand small shock heads. Reducing the size of piezoelectric shock heads will only be possible if the pressure generated at the surface of each transducer can be increased so that the total pressure at the focus remains very high. We propose for the first time to increase the pressure without increasing the transducer voltage by using sandwiched transducers, which are a combination of several stacked transducers. When excited at appropriate time intervals, the pressure waves generated by each one reinforce when they reach the load. This new technique has been successfully tested. A pressure of 2.5 MPa was generated with two stacked, 5 mm-thick 1-3 piezocomposite transducers operating at an excitation voltage of 8 kV. No transducer damage was detected after 10(6) shocks, which corresponds approximately to the treatment of 500 patients. PMID- 11106009 TI - Study on the lifetime and attenuation properties of microbubbles coated with carboxylic acid salts. AB - Four kinds of surfactants, sodium laurate, sodium myristate, sodium palmitate and sodium oleate were used to study the effects of surfactant coatings on the lifetime and attenuation of microbubbles. The changes in the size distribution of microbubbles prepared with these surfactants in saline were measured with a Coulter Multisizer (Coulter Electronics Ltd., Luton, UK). Frequency characteristics of ultrasonic attenuation of the microbubble suspensions were measured between 400 kHz and 6 MHz. From the changes in attenuation in the microbubble suspensions over time, it was found that the lifetime of microbubbles in a suspension also depends on the frequency of the irradiating ultrasound. The effect of surfactants on the frequency characteristics of attenuation was also studied, and characteristics of the surfactant coating, including shell elasticity and shell friction parameters were calculated from the measurement results. Microbubbles produced with sodium palmitate had the longest lifetime and the smallest average size. The shell had very little effect on the ultrasonic properties of microbubbles produced with sodium palmitate, suggesting that the sodium palmitate microbubbles behaved ultrasonically as free microbubbles. PMID- 11106010 TI - The performance and design of ultrasonic vibration system for flexural mode. AB - The matrix method for analyzing flexural vibration system is outlined. The displacement, stress and velocity gain of ultrasonic flexural vibration systems are investigated by transfer matrix method. A design method is given for flexural transducer and vibration system. PMID- 11106011 TI - The common risk factor approach: a rational basis for promoting oral health. AB - Conventional oral health education is not effective nor efficient. Many oral health programmes are developed and implemented in isolation from other health programmes. This often leads, at best to a duplication of effort, or worse, conflicting messages being delivered to the public. In addition, oral health programmes tend to concentrate on individual behaviour change and largely ignore the influence of socio-political factors as the key determinants of health. Based upon the general principles of health promotion this paper presents a rationale for an alternative approach for oral health policy. The common risk factor approach addresses risk factors common to many chronic conditions within the context of the wider socio-environmental milieu. Oral health is determined by diet, hygiene, smoking, alcohol use, stress and trauma. As these causes are common to a number of other chronic diseases, adopting a collaborative approach is more rational than one that is disease specific. The common risk factor approach can be implemented in a variety of ways. Food policy development and the Health Promoting Schools initiative are used as examples of effective ways of promoting oral health. PMID- 11106012 TI - Nutritional variables related to gingival health in adolescent girls. AB - In order to study the nutritional variables associated with gingival health, a case-control study was designed to control strong variables whose effect on gingival status may obscure the potential effect of weaker ones, such as nutrition. Two groups of 27 gingivitis-affected and -unaffected female adolescents were selected. All were aged 17-19 years, with mean age of the two groups statistically not different. All were non-smokers, all reported daily toothbrushing frequency of twice/day or more, and none had clinical signs of hyponutrition. Mean DMFT of the two groups was statistically not different. The effect of nutritional variables, obtained by a three-day food record and by assessing the nutritional status of the girls, on presence/absence of gingivitis was evaluated by a variety of stepwise logistic regression analyses. Age (positive correlation), riboflavin, calcium and frequency of fibre intake (negative correlations) significantly explained the risk for gingivitis. Strong intercorrelation between riboflavin and calcium was also found, due to the high quantity of milk consumed by the girls, since this food provided the main source of riboflavin and calcium. The data suggest that some dietary measures may be useful for the maintenance of healthy gingival status. PMID- 11106013 TI - Dental neglect and dental health among 26-year-olds in the Dunedin Multidisciplinary Health and Development Study. AB - OBJECTIVES: To test a modification of a previously-reported six-item dental neglect scale and examine its association with dental health and service-use among young adults. METHODS: Of the 980 26-year-old participants in the Dunedin Multidisciplinary Health and Development Study, 973 completed the scale and 930 underwent dental examination. Sociodemographic and dental service-use data were collected using a self-report questionnaire. RESULTS: Factor analysis showed that five of the original six items loaded on the scale, and responses to those items were summed to give a dental neglect scale score for each participant. Scale scores were normally distributed (mean=13.0; SD=3.6; range 5 to 25), and a median split created higher and lower dental neglect groups. A higher proportion of the higher neglect group group: (i) were male; (ii) rated their dental health and dental appearance as below average; (iii) brushed their teeth infrequently; (iv) had extensive plaque deposits; (v) used dental services only when they had a problem; (vi) had not recently seen a dentist; (vii) had lost at least one permanent tooth because of caries, and (viii) had a greater number of decayed tooth surfaces. CONCLUSIONS: Although further examination of its validity and reliability is indicated, the dental neglect scale appears to hold promise for use in dental health promotion, not only in highlighting population groups or individuals who would benefit from intensive health promotion efforts, but also in the evaluation of health promotion interventions. PMID- 11106014 TI - Caries prevention with chlorhexidine-thymol varnish in high risk schoolchildren. AB - In a prospective, randomised and blinded cohort study, the acceptance and effectiveness of a 3-monthly chlorhexidine-thymol varnish application was assessed in 8- to 10-yr-old children (n=29) with high caries incidence after brushing with a 1.23% fluoride gel. The children of the control group (n=25) brushed with a fluoride gel only. The standardised interview showed good acceptance of the varnish applications in spite of the bitter taste. Caries incidence in one year was 1.2 DMFS (SD 1.5, median 1; 95% confidence interval 0 1) for the test group and 2.1 DMFS (SD 2.3, median 2; 95% confidence interval 0 3) for the control group. Due to a slightly lower reduction in caries incidence (42.3%) than in other studies and a higher standard deviation than in unselected study groups, the power of the present study was not high enough to prove this difference in caries incidence to be statistically significant in the Mann Whitney test (P=0.11). This was also found for the difference of 0.4 initial lesions. The distribution patterns of caries incidence for tooth groups (70% in the first permanent molar) and surfaces (58% on occlusal surfaces) did not differ between the test and the control group. The reductions of caries incidence which have been found in highly caries active children of the present study should be evaluated further with a larger study sample. PMID- 11106015 TI - Self-assessed oral health status of ethnic minority residents of South London. AB - OBJECTIVE: To determine the self-assessed oral health status of individuals from minority ethnic communities living in South London. METHOD: A sample of 1,057 individuals from minority ethnic communities (as defined by Office of Censuses and Surveys categories) resident in South London were asked to complete measures of their oral and facial symptoms, the impact of their oral health on their daily functioning and of their satisfaction with the appearance of their teeth and gums. MEASURES: Self-assessed oral health status was determined by means of two short scales addressing oral symptoms and the impact of oral health on activities of daily living. Satisfaction with the appearance of the teeth and gums was also assessed. FINDINGS: No significant differences were found between minority ethnic communities in the number of symptoms reported, in the level of impact which such symptoms cause, or in their dissatisfaction with the appearance of their teeth and gums. Reporting of symptoms and impact were mildly though significantly correlated with dissatisfaction. CONCLUSIONS: There appears to be little difference between ethnic communities in their reporting of oral and facial symptoms, though these groups report higher levels of dissatisfaction with the appearance of their teeth and gums. Social and demographic factors play an important role in determining perceptions of oral health. The findings should be interpreted with caution given the difficulty of sampling minority ethnic communities and the subsequent limited representativeness of the sample. PMID- 11106016 TI - Dental anxiety reduction and dental attendance after treatment in a dental fear clinic: a follow-up study. AB - OBJECTIVES: The aim of the present study was to assess treatment outcome in terms of dental anxiety reduction at a post-treatment assessment and dental anxiety reduction and dental attendance one year later. Furthermore, it was determined to what extent psychopathological characteristics were related to treatment outcome. METHODS: Questionnaires were sent to 280 patients treated with one of three treatment modes (i.e., behavioral management (BM), nitrous oxide sedation (NOS), and intravenous sedation (IVS)) at a dental fear clinic in The Netherlands. Dental anxiety before (T1) and after (T2) treatment was assessed using the Dental Anxiety Scale (DAS) and the Short version of the Dental Anxiety Inventory (S DAI); the Symptom Checklist 90 (SCL-90) was used to assess general psychopathology. Dental anxiety was assessed again a year later and patients were questioned about their dental attendance pattern (T3). RESULTS: ANOVA showed that the DAS and S-DAI scores at T2 and T3 were statistically significant lower than the initial scores. In addition, IVS patients showed less anxiety reduction than BM patients at both T2 and T3. Of the 145 patients whose last visit to the clinic was at least one year ago, 62% had visited a GDP at T3. A regression analysis revealed that, beside treatment mode, somatization, number of visits to clinic for dental treatment, and number of months between first and last visit to the clinic predicted dental anxiety at follow-up. CONCLUSIONS: It is concluded that, although a reduction in dental anxiety level was present, a relatively large proportion of patients did not improve, in terms of both dental anxiety and dental attendance. PMID- 11106017 TI - Oral hygiene and sugar consumption among urban and rural adolescents in Ghana. AB - OBJECTIVES: The purpose of this study was to compare categories of self-reported oral health behavior among adolescents raised in urban and rural areas of Ghana. METHODS: Second year students (n=583) aged 14-18 years were selected from the five secondary schools in the Awutu-Effutu Senya district, using a stratified random sampling procedure. A total of 504 (86%) students completed questionnaires under supervision at school. RESULTS: Cross-tabulation analyses revealed that most urban and rural residents (97% and 96%) reported daily tooth brushing, whereas small and moderate proportions reported use of toothpicks and intake of sugared snacks, respectively. Multiple logistic regression analyses, controlling for gender and parents' education, showed that the socio-regional context (urban/rural) was significantly related to intake of cakes/biscuits (odds ratio (OR)=2.6, 95% CI 1.7-4.4), chocolate/sweets (OR=3.5, 95% CI 2.0-6.0) and use of toothpicks (OR=2.6, 95% CI 1.3-5.5), indicating true differences between urban and rural residents. CONCLUSION: As predicted, urban residents were more likely than their rural counterparts to use toothpicks and to consume sugared snacks. Among both urban and rural residents, males and those having less educated parents reported oral hygiene and sugar intake less frequently than did females and the highly educated. Whereas the gender difference with respect to intake of sugared snacks was larger among urban than rural residents, the socio-economic disparity with respect to use of toothpicks was most pronounced among rural residents. The results appear to imply that in addition to targeting specific oral health behaviors, bath individual characteristics and the wider socio regional context should be addressed when planning oral health intervention among Ghanaian adolescents. PMID- 11106018 TI - Factors associated with dental anxiety and attendance in middle-aged and elderly women. AB - The aim of this study was to analyze the association between dental anxiety, dental attendance, health status and social factors. Our previous studies have shown that dental anxiety declines with age and is associated with poor oral health. In addition, correlations between dental anxiety, dental utilization and dental status have been revealed. However, the association of these factors with general health and social factors has not been analyzed in our previous studies. In a study of women's health in Goteborg, Sweden in 1992, 1.017 urban women aged 38 to 84 years took part in a series of investigations including clinical examinations, interviews and questionnaires. In addition to descriptive and simple inference statistics, a two-part multiple logistic regression model was utilized to investigate dental anxiety and dental utilization. Dental fear was less prevalent among older women, dentate or not, although 10% of females 62 years of age and older still reported high dental anxiety (DAS > or = 12). 94% of the younger (< or = 62 yr) and 76% of the older (> or = 70 yr) women reported regular dental attendance. When separating all women into dentate and edentulous groups, 94% of the dentate and 11% of the edentulous respondents reported regular dental care. Due to the large difference in dental attendance between dentate and edentulous women, these groups were analyzed separately. Multiple logistic regression analyses showed that the following factors were associated with irregular dental utilization among dentate women: high dental anxiety, fewer teeth and restorations, more caries, poorer chewing ability and dissatisfaction with dental esthetics. In the multiple regression for dental anxiety, high fear was shown to be associated with irregular dental care, age (younger), fewer teeth, dissatisfaction with dental esthetics and lower scores on the SF-36 mental health scale. A separate analysis showed that individuals with high fear and regular, as opposed to irregular, dental attendance had more teeth at a statistically significant level, which were less often decayed and more often restored. In spite of the group with high fear and irregular attendance having fewer teeth, their level of decay was seven times higher. Overall, the results indicate a strong association between dental fear and dental attendance. Weak associations were found among socio-economic, dental health and general health factors. PMID- 11106019 TI - Factors of patient satisfaction/dissatisfaction in a dental faculty outpatient clinic in Turkey. AB - OBJECTIVES: Being service providers, dental professionals should satisfy their consumers/dental patients. This study investigates satisfaction with dental care among the patients of a dental faculty outpatient clinic of a major university in Izmir, Turkey. METHOD: The study was performed on 1001 patients of whom 674 filled out the questionnaire containing sociodemographic items and open-ended questions to determine the factors of (dis)satisfaction. The open-ended questions were content analyzed, and each patient was scored according to his comments: "complaining: (0)", "both praising and complaining: (1)", "no comment: (2)", and "praising: (3)". Each factor that has an impact on the decision of the study sample was determined by statistical analyses of data, using student t-test, chi square test, and multiple regression analysis. RESULTS: Most of the patients were highly educated (74.7%), had a high income (48.7%), and were young to middle-aged (73.1%). There was a well-balanced gender representation. The patient sample had sought care mostly for dental caries, periodontal diseases, problems with old restorations, and prosthetic rehabilitation. Of 1,001 patients, 38.6% were satisfied with the dental care they received, 23.8% were both satisfied and dissatisfied, 5% were dissatisfied, and 32.7% failed to comment. No significant differences were observed between the satisfaction/dissatisfaction scores and sociodemographic variables of the patients in the groups (P>0.05). The most important components of satisfaction were found to be "relationship between dentists and patients" (P<0.001), "organized service system" (P<0.001), and "scientific ability of dental personnel" (P<0.001). The most prominent complaints were "long treatment span" (P<0.001), "disorganized service system" (P<0.001), and "slowness of radiographical examination procedures" (P<0.001). CONCLUSIONS: Despite the significant variations among the cultural and ethnic structures of different societies, personal interactions have priority in establishing satisfying dental service. PMID- 11106020 TI - Differential regulation of fixN-reiterated genes in Rhizobium etli by a novel fixL-fixK cascade. AB - Among the complexities in the regulation of nitrogen fixation in the Rhizobiaceae are reiteration of regulatory components as well as variant roles for each component between species. For Rhizobium etli CFN42, we reported that the symbiotic plasmid (pCFN42d) contains a key regulatory gene (fixKd) and genes for a symbiotic cytochrome oxidase (fixNOQPd). Here we discuss the occurrence of reiteration of these genes (fixKf and fixNOQPf) and the finding of an unusual fixL homolog on a plasmid previously considered cryptic (pCFN42f). The structure of the deduced FixL polypeptide is suggestive of a fusion of the receiver and transmitter modules of a two-component regulatory system as described in R. leguminosarum bv. viciae VF39. Gene fusion analysis, coupled with mutation of each regulatory element, revealed that free-living expression of FixKf was dependent fully on FixL. In contrast, synthesis of FixKd was not detected under the conditions tested. The FixKf protein is needed for microaerobic expression of both fixN reiterations, whereas the FixKd protein appears to be dispensable. Interestingly, expression of the fixN reiterations exhibits a differential dependence for FixL, where transcription of fixNf was suppressed in the absence of FixL but expression of fixNd still showed significant levels. This suggests the existence of a FixL-independent mechanism for expression of the fixNd reiteration. Surprisingly, mutations in fixL, fixKd, or fixKf (either singly or in combination) did not alter symbiotic effectiveness. A mutation in fixNd (but not in fixNf) was, however, severely affected, indicating a differential role for these reiterations in nitrogen fixation. PMID- 11106021 TI - Chromosomal insertion of phenazine-1-carboxylic acid biosynthetic pathway enhances efficacy of damping-off disease control by Pseudomonas fluorescens. AB - A disarmed Tn5 vector (pUT::Ptac-phzABCDEFG) was used to introduce a single copy of the genes responsible for phenazine-1-carboxylic acid (PCA) biosynthesis into the chromosome of a plant-growth-promoting rhizobacterium Pseudomonas fluorescens. The PCA gene cluster was modified for expression under a constitutive Ptac promoter and lacked the phzIR regulators. PCA-producing variants significantly improved the ability of the wild-type P. fluorescens to reduce damping-off disease of pea seedlings caused by Pythium ultimum, even under conditions of heavy soil infestation. Under conditions of oxygen limitation that are typical of the rhizosphere, PCA production per cell in vitro was greater than that recorded in fast-growing, nutrient-rich cultures. Similarly, when the in vitro nutrient supply was limited, P fluorescens::phz variants that produced the most PCA effectively competed against P. ultimum by suppressing mycelial development. Soil-based bioassays confirmed that the level of PCA biosynthesis correlated directly with the efficacy of biological control and the persistence of inocula in soil microcosms. They also showed that soil pretreatment with bacteria provides a suitable method for plant protection by reducing infection, effectively decontaminating the soil. These data demonstrate that the insertion of a single chromosomal copy of the genes for a novel antifungal compound, PCA, enhances the ecological fitness of a natural isolate already adapted to the rhizosphere and capable of suppressing fungal disease. PMID- 11106022 TI - Effects of targeted replacement of the tomatinase gene on the interaction of Septoria lycopersici with tomato plants. AB - Many plants produce constitutive antifungal molecules belonging to the saponin family of secondary metabolites, which have been implicated in plant defense. Successful pathogens of these plants must presumably have some means of combating the chemical defenses of their hosts. In the oat root pathogen Gaeumannomyces graminis, the saponin-detoxifying enzyme avenacinase has been shown to be essential for pathogenicity. A number of other phytopathogenic fungi also produce saponin-degrading enzymes, although the significance of these for saponin resistance and pathogenicity has not yet been established. The tomato leaf spot pathogen Septoria lycopersici secretes the enzyme tomatinase, which degrades the tomato steroidal glycoalkaloid alpha-tomatine. Here we report the isolation and characterization of tomatinase-deficient mutants of S. lycopersici following targeted gene disruption. Tomatinase-minus mutants were more sensitive to alpha tomatine than the wild-type strain. They could, however, still grow in the presence of 1 mM alpha-tomatine, suggesting that nondegradative mechanisms of tolerance are also important. There were no obvious effects of loss of tomatinase on macroscopic lesion formation on tomato leaves, but trypan blue staining of infected tissue during the early stages of infection revealed more dying mesophyll cells in leaves that had been inoculated with tomatinase-minus mutants. Expression of a defense-related basic beta-1,3 glucanase gene was also enhanced in these leaves. These differences in plant response may be associated with subtle differences in the growth of the wild-type and mutant strains during infection. Alternatively, tomatinase may be involved in suppression of plant defense mechanisms. PMID- 11106023 TI - The Pseudomonas syringae avrRpt2 gene product promotes pathogen virulence from inside plant cells. AB - Several bacterial avr genes have been shown to contribute to virulence on susceptible plants lacking the corresponding resistance (R) gene. The mechanisms by which avr genes promote parasitism and disease, however, are not well understood. We investigated the role of the Pseudomonas syringae pv. tomato avrRpt2 gene in pathogenesis by studying the interaction of P. syringae pv. tomato strain PstDC3000 expressing avrRpt2 with several Arabidopsis thaliana lines lacking the corresponding R gene, RPS2. We found that PstDC3000 expressing avrRpt2 grew to significantly higher levels and often resulted in the formation of more severe disease symptoms in ecotype No-0 plants carrying a mutant RPS2 allele, as well as in two Col-0 mutant lines, cpr5 rps2 and coil rps2, that exhibit enhanced resistance. We also generated transgenic A. thaliana lines expressing avrRpt2 and demonstrated, by using several different assays, that expression of avrRpt2 within the plant also promotes virulence of PstDC3000. Thus, AvrRpt2 appears to promote pathogen virulence from within the plant cell. PMID- 11106024 TI - Xanthomonas oryzae pv. oryzae avirulence genes contribute differently and specifically to pathogen aggressiveness. AB - Genomic copies of three Xanthomonas oryzae pv. oryzae avirulence (avr) genes, avrXa7, avrXal0, and avrxa5, and four homologous genes, aB3.5, aB3.6, aB4.3, and aB4.5, were mutagenized individually or in combination to study the roles of avr genes in one component of pathogen fitness, i.e., aggressiveness or the amount of disease X. oryzae pv. oryzae causes in susceptible rice lines. These X. oryzae pv. oryzae genes are members of the highly related Xanthomonas avrBs3 gene family. Compared to the wild-type strain, X. oryzae pv. oryzae strains with mutations in avrXa7, avrxa5, and the four homologous genes caused shorter lesions on rice line IR24, which contains no resistance genes relevant to the wild-type strain. The contribution of each gene to lesion length varied, with avrXa7 contributing the most and avrXal0 showing no measurable effect on aggressiveness. The functional, plasmidborne copies of avrXa7, aB4.5, and avrxa5 restored aggressiveness only to strains with mutations in avrXa7, aB4.5, and avrxa5, respectively. Mutations in avrXa7 were not complemented by plasmids carrying any other avr gene family members. These data indicate that some, but not all, avr family members contribute to pathogen aggressiveness and that the contributions are quantitatively different. Furthermore, despite their sequence similarity, the aggressiveness functions of these gene family members are not interchangeable. The results suggest that selection and pyramiding resistance genes can be guided by the degree of fitness penalty that is empirically determined in avr gene mutations. PMID- 11106025 TI - Mating-type genes from asexual phytopathogenic ascomycetes Fusarium oxysporum and Alternaria alternata. AB - Mating-type (MAT) loci were cloned from two asexual (mitosporic) phytopathogenic ascomycetes, Fusarium oxysporum (a pyrenomycete) and Alternaria alternata (a loculoascomycete), by a polymerase chain reaction (PCR)-based strategy. The conserved high mobility group (HMG) box domain found in the MAT1-2-1 protein was used as a starting point for cloning and sequencing the entire MAT1-2 idiomorph plus flanking regions. Primer pairs designed to both flanking regions were used to amplify the opposite MAT1-1 idiomorph. The MAT1-1 and MAT1-2 idiomorphs were approximately 4.6 and 3.8 kb in F. oxysporum and approximately 1.9 and 2.2 kb in A. alternata, respectively. In both species, the MAT1-1 idiomorph contains at least one gene that encodes a protein with a putative alpha box domain and the MAT1-2 idiomorph contains one gene that encodes a protein with a putative HMG box domain. MAT-specific primers were used to assess the mating type of F. oxysporum and A. alternata field isolates by PCR. MAT genes from A. alternata were expressed. The A. alternata genes were confirmed to be functional in a close sexual relative, Cochliobolus heterostrophus, by heterologous expression. PMID- 11106026 TI - Root colonization by phenazine-1-carboxamide-producing bacterium Pseudomonas chlororaphis PCL1391 is essential for biocontrol of tomato foot and root rot. AB - The phenazine-1-carboxamide-producing bacterium Pseudomonas chlororaphis PCL1391 controls tomato foot and root rot caused by Fusarium oxysporum f. sp. radicislycopersici. To test whether root colonization is required for biocontrol, mutants impaired in the known colonization traits motility, prototrophy for amino acids, or production of the site-specific recombinase, Sss/XerC were tested for their root tip colonization and biocontrol abilities. Upon tomato seedling inoculation, colonization mutants of strain PCL1391 were impaired in root tip colonization in a gnotobiotic sand system and in potting soil. In addition, all mutants were impaired in their ability to control tomato foot and root rot, despite the fact that they produce wild-type levels of phenazine-1-carboxamide, the antifungal metabolite previously shown to be required for biocontrol. These results show, for what we believe to be the first time, that root colonization plays a crucial role in biocontrol, presumably by providing a delivery system for antifungal metabolites. The ability to colonize and produce phenazine-1 carboxamide is essential for control of F. oxysporum f. sp. radicis-lycopersici. Furthermore, there is a notable overlap of traits identified as being important for colonization of the rhizosphere and animal tissues. PMID- 11106028 TI - Restriction enzyme-mediated integration used to produce pathogenicity mutants of Colletotrichum graminicola. AB - We have developed a restriction enzyme-mediated insertional mutagenesis (REMI) system for the maize pathogen Colletotrichum graminicola. In this report, we demonstrate the utility of a REMI-based mutagenesis approach to identify novel pathogenicity genes. Use of REMI increased transformation efficiency by as much as 27-fold over transformations with linearized plasmid alone. Ninety-nine transformants were examined by Southern analysis, and 51% contained simple integrations consisting of one copy of the vector integrated at a single site in the genome. All appeared to have a plasmid integration at a unique site. Sequencing across the integration sites of six transformants demonstrated that in all cases the plasmid integration occurred at the corresponding restriction enzyme-recognition site. We used an in vitro bioassay to identify two pathogenicity mutants among 660 transformants. Genomic DNA flanking the plasmid integration sites was used to identify corresponding cosmids in a wild-type genomic library. The pathogenicity of one of the mutants was restored when it was transformed with the cosmids. PMID- 11106027 TI - Xv4-vrxv4: a new gene-for-gene interaction identified between Xanthomonas campestris pv. vesicatoria race T3 and wild tomato relative Lycopersicon pennellii. AB - Strains of tomato race 3 (T3) of Xanthomonas campestris pv. vesicatoria elicit a hypersensitive response (HR) in leaves of Lycopersicon pennellii LA716. Genetic segregation of the resistance exhibited ratios near 3:1 in F2 populations, which confirmed that a single dominant gene controlled the inheritance of this trait. With the aid of a collection of introgression lines, restriction fragment length polymorphism, and cleaved amplified polymorphic sequence markers, the resistance locus was located on chromosome 3 between TG599 and TG134. An avirulence gene named avrXv4 was also isolated by mobilizing a total of 600 clones from a genomic DNA library of the T3 strain 91-118 into the X. campestris pv. vesicatoria strain ME90, virulent on L. pennellii. One cosmid clone, pXcvT3-60 (29-kb insert), induced HR in resistant plants. The avirulent phenotype of pXcvT3-60 was confirmed by comparing growth rates in planta and electrolyte leakages among transconjugants carrying a mutated or intact clone with the wild-type T3 strain 91-118. A 1.9-kb DNA fragment contained within a 6.8-kb active subclone was sequenced and was determined to carry an open reading frame of 1,077 bp. The predicted AvrXv4 protein exhibits high similarity to members of an emerging new family of bacterial proteins from plant and mammalian pathogens comprising AvrRxv, AvrBsT, YopJ, YopP, AvrA, and YL40. PMID- 11106029 TI - Elicitation of hypersensitive cell death by extracellularly targeted HrpZPsph produced in planta. AB - The ability of the Pseudomonas syringae pv. phaseolicola harpin (HrpZPsph) to elicit hypersensitive response was investigated in three Nicotiana genotypes. The hrpZPsph gene was placed under chemical regulation (tetracycline induction) in TetR+ Nicotiana tabacum cv. Wisconsin 38 (W38) or was transiently expressed in N. benthamiana following infection with a PVX-derived vector and in three Nicotiana genotypes by agroinfiltration. The constructs were designed to express either the canonical form of harpin (HrpZPsph) or an N-terminally extended version of the protein carrying the signal peptide portion of the tobacco pathogenesis-related protein PR1a (SP-HrpZPsph). Stable transformants of N. tabacum cv. W38 did not develop necrosis upon induction with tetracycline, probably as a result of insufficient harpin accumulation. In contrast, N. benthamiana plants infected with the PVX constructs produced high concentrations of harpin in biologically active form, but only those expressing the secretable form of harpin developed necrotic symptoms. These symptoms were less severe than those caused by PVX::avrPto; however, they were accompanied by induction of hsr203J, a hypersensitive response-specific gene transcript. These results suggest that the plant cellular receptor(s) for harpin is extracellular. PMID- 11106030 TI - Avirulence in the wheat septoria tritici leaf blotch fungus Mycosphaerella graminicola is controlled by a single locus. AB - Segregation of avirulence in Mycosphaerella graminicola, a heterothallic ascomycete that causes wheat septoria tritici leaf blotch, was studied in F1, BC1, and F2 populations by inoculation assays on five wheat cultivars in the seedling stage and by amplified fragment length polymorphism and random amplified polymorphic DNA analyses. F1 was generated by crossing isolates IPO323 (avirulent) and IPO94269 (virulent). All F1, BC1, and F2 progeny isolates were virulent on the susceptible check cultivar Taichung 29 and were avirulent on the resistant check cultivar Kavkav-K4500. Avirulence segregation was observed in F1 and in several BC1 and F2 generations on the differential cultivars Shafir, Kavkaz, and Veranopolis at a 1:1 ratio. Avirulence for the three differential cultivars always cosegregated. We conclude that avirulence in isolate IPO323 is controlled by a single, seemingly complex locus. PMID- 11106031 TI - Nitric oxide inhibition of tobacco catalase and ascorbate peroxidase. AB - We used a variety of nitric oxide (NO) donors to demonstrate that NO inhibits the activities of tobacco catalase and ascorbate peroxidase (APX). This inhibition appears to be reversible because removal of the NO donor led to a significant recovery of enzymatic activity. In contrast, APX and catalase were irreversibly inhibited by peroxynitrite. The ability of NO and peroxynitrite to inhibit the two major H2O2-scavenging enzymes in plant cells suggests that NO may participate in redox signaling during the activation of defense responses following pathogen attack. PMID- 11106032 TI - Lipochito-oligosaccharide nodulation factors stimulate cytoplasmic polarity with longitudinal endoplasmic reticulum and vesicles at the tip in vetch root hairs. AB - Vetch root hair development has four stages: bulge, growing, growth terminating, and full-grown hair. In the assay we used, the nodulation factor induced swellings and outgrowths in growth-terminating hairs. Bulges, swellings, and full grown hairs have transverse endoplasmic reticulum (ER) and no tip-accumulated vesicles. Growing hairs and outgrowths show vesicle accumulation in the tip and longitudinal subapical ER. Bulge walls and walls of swellings appear mottled. PMID- 11106033 TI - Transmission of human immune deficiency virus type-1 from mother to infant. PMID- 11106034 TI - Breathing patterns, oxygen and carbon dioxide levels during infancy. PMID- 11106035 TI - Don't discard your indomethacin yet. PMID- 11106036 TI - Breastmilk and skin-to-skin kangaroo care for premature infants. Avoiding bonding failure. PMID- 11106037 TI - Breathing patterns, oxygen and carbon dioxide levels in sleeping healthy infants during the first nine months after birth. AB - Data on arterial oxygen saturation (SaO2), transcutaneous PO2, pCO2 (tcpO2, tcpCO2) and breathing patterns in sleeping healthy term infants were obtained during the first 9 mo after birth. Forty-four healthy infants, mean GA at birth 40 +/- 1.0 wk, mean BW 3520 +/- 562 g were examined between 2 wk and 9 mo postnatally in a cross-sectional study. SaO2, tcpO2, tcpCO2, heart rate (HR), rib cage and abdominal respiratory movements were recorded during natural nocturnal sleep, stratified for sleep states (active sleep (AS), indeterminate sleep (IS), quiet sleep (QS)). The data on AS and IS were pooled as in previous studies. The variables were analysed with respect to age. SaO2 in AS + IS and QS was 96.1 +/- 1.3%, 96.6 +/- 1.4%, respectively. TcpO2 in AS + IS was 10.6 +/- 1.1 kPa and 10.7 +/- 1.3 kPa in QS, while tcpCO2 in AS + IS was 5.4 +/- 0.3 kPa and 5.4 +/- 0.4 kPa in QS. Neither SaO2 nor tcpO2 was influenced by age. TcpCO2 decreased significantly postnatally. Five infants (11.3%) experienced episodes of hypoxaemia with a mean decrease in SaO2 to 86 +/- 1.5%. In four infants these hypoxaemic episodes were linked to upper airway obstructions. Episodes of SaO2 < 90% in conjunction with a decrease in HR to < 100 bpm were detected in one infant only. Periodic breathing (PB) was observed in 38.6% of infants. CONCLUSION: Oxygenation and carbon dioxide levels in sleeping healthy term infants were comparable to those reported in older children. Hypoxaemic episodes, if present, are associated with upper airway obstruction. PB, often assumed to be a pathological feature, is a normal breathing pattern in this age group. PMID- 11106038 TI - Effect of training on peak oxygen uptake and blood lipids in 13 to 14-year-old girls. AB - The aim of this study was to investigate the effects of exercise training on the peak oxygen uptake (peak VO2) and blood lipid profile of 13 to 14-y-old postmenarcheal girls. Treadmill determined peak VO2, total cholesterol, high density lipoprotein cholesterol, low density cholesterol, and triglycerides were the outcome measures assessed at baseline and following exercise training. Twenty girls completed a 20-wk programme of exercise training which involved maintaining the heart rate at 75-85% maximum for 20 min, three times per week. Heart rate was rigorously monitored using telemetry throughout each training session. Eighteen girls acted as the control group. There were no significant (p > 0.05) changes in the outcome measures following the training programme. CONCLUSIONS: These findings suggest that exercise training of this frequency, intensity and duration for a period of 20 wk has no significant effect on either the peak VO2 or blood lipid and lipoprotein profile of normolipidaemic, postmenarcheal girls. PMID- 11106040 TI - Spontaneous growth in German children and adolescents with genetically confirmed Prader-Willi syndrome. AB - Height and weight in children with Prader-Willi syndrome, diagnosed by standard clinical criteria, follow a specific developmental pattern resulting in early childhood obesity, absent pubertal growth spurt and adolescent short stature. New molecular techniques (methylation analysis, fluorescence in situ hybridization) now allow the unequivocal diagnosis of Prader-Willi syndrome (PWS). We investigated the possibility of a bias in syndrome-specific growth standards based on clinically diagnosed patients by comparing these standards with new standards derived from 100 German patients with molecularly confirmed PWS, none of whom had received a growth-promoting therapy. Height centile curves of the German patients fall in the tall range of standards derived from American patients. This is mainly due to an elevation of the lower centile ranges in both sexes. When the height standards derived from German patients are compared to those of a large multinational cohort of patients, 78% of whom were not confirmed by genetic testing, only minor differences in the height centiles become apparent. The population background therefore does not appear to play a major role for the observed differences. In a marked proportion of patients a decreased sitting height/height ratio is found. This was usually associated with scoliosis. Weight standards from our study group show that after 14 y of age German girls with PWS are heavier than their American counterparts. Standards for the body mass index of German patients of both sexes are increased over normal reference standards (p < 0.0001) and do increase with age (boys: p = 0.0038; girls: p = 0.0004). PWS genotypes or sex had no apparent influence on SDS for height, weight and body mass index. CONCLUSIONS: Because of the observed differences to other growth standards, use of the newly constructed centile curves is advocated in German patients with molecularly confirmed PWS to avoid delay in the diagnosis of additional growth-compromising conditions. PMID- 11106039 TI - Final height and body disproportion in thalassaemic boys and girls with spontaneous or induced puberty. AB - The aim of the study was to evaluate whether sex hormone replacement therapy adversely affected final height and body disproportion in thalassaemic boys and girls. Thirty-six patients with spontaneous (SP) or induced puberty (IP) were studied in order to define the pattern of height growth through three observations: the first (A) at the age of 7-9; the second (B) at onset of spontaneous or induced puberty; and the third (C) when final height was reached. We examined 14 females with SP (f-SP) and 8 with IP (f-IP); 7 males with SP (m SP) and 7 with IP (m-IP). Girls with IP reached the same final height of girls with SP (f-IP 153.8 (4.3) versus f-SP 154.4 (5.5) cm); p > 0.05) close to target height (f-IP 155.9 (5.2) cm versus f-SP 155.5 (3.6) cm). Girls with IP reached the final height at older chronological age (CA) (17.0 (0.6) y) than girls with SP (CA of 15.3 (0.7) y), but at the same bone age (BA) (f-IP 15.1 (0.9) y versus f-SP 14.8 (0.6) y). There was no difference between the two groups for pubertal growth (f-SP 16.2 (7.7) cm versus f-IP 12.2 (7.4) cm (p > 0.05)) that was negatively correlated with both prepubertal growth and BA at onset of puberty in both groups. Values of sitting height (sds) with respect to BA (SHsdsBA) were not significantly different between the two groups, and showed a worsening from the first observation to final height, reaching values around -2 SD, in both groups. Values of subischial leg length (sds) with respect to BA (SLLsdsBA) were in the normal range at both observations in all girls. High serum ferritin levels were observed in both groups (f-SP 3189 (2296) ng/ml and f-IP 3998 (2545) ng/ml; p > 0.05). Also boys with induced puberty reached the same final height of those with spontaneous one (m-IP 160.9 (5.5) cm versus m-SP 161.8 (2.4) cm; p > 0.05), but it was lower than target height in both groups (m-IP 168.1 (4.1) cm versus m-SP 169.6 (3.2) cm). Boys with IP reached final height at CA of 18.6 (1.1) y slightly older than boys with SP (CA 17.2 (0.9) y), but at the same BA (m-IP 15.9 (1.5) y versus m-SP 16.3 (0.8) y). Pubertal growth values were significantly different between boys with SP 18.9 (5.3) cm and those with IP 13.8 (4.9) cm (p < 0.05), but they were negatively correlated with prepubertal growth values in both groups (m-SP r = -0.91; p < 0.002 and m-IP r = -0.51; p < 0.05). SHsdsBA showed a worsening from the first observation to final height, reaching values around -3 SD in both groups, while SLLsdsBA were always in the normal range in all patients. Serum ferritin levels were higher in boys with IP (3400 (1179) ng/ml) than in those with SP (2020 (496) ng/ml). CONCLUSIONS: Our data showed that: (a) patients of both sexes with induced puberty reached the same final height of patients with spontaneous puberty; (b) all patients showed a body disproportion with truncal shortening and normal leg length that was more severe in boys of both groups at final height; (c) body disproportion was independent of pubertal or prepubertal period of greater height gain, suggesting that sexual steroids replacement therapy did not adversely affect either final height or body disproportion. Further studies, focused on the pathogenesis of the truncal shortening, are necessary in order to acquire more insight into the causes of this impairment. PMID- 11106041 TI - CagA seropositivity and the severity of Helicobacter pylori infection in dyspeptic children. AB - AIM: To assess the incidence of cagA (cytotoxin-associated protein) and to evaluate its correlation with endoscopic-histologic findings and with eradication rate in a series of children affected by Helicobacter pylori (H. pylori) gastritis. METHODS: Fifty consecutive H. pylori gastritis children (27M; median age 10 y and 11 mo) were tested for IgG cagA protein (Western Blot technique). Pretreatment H. pylori infection was assessed on the grounds of endoscopic antral biopsy specimens by means of rapid urease test and histologic examination (Giemsa staining). All the children were treated with omeprazole (1 mg/kg/d), clarithromycin (15 mg/kg/d) and amoxycillin (50 mg/kg/d) for 2 wk. According to universally accepted clinical practice, outcome of treatment was assessed by 13C urea breath test at least 6 wk after the end of therapy. RESULTS: Thirty-five children (70%) were seropositive to cagA+ protein (median age 11 y and 1 mo). Endoscopic findings of cagA+ patients were similar to those of cagA- patients. In cagA seropositive patients the severity of histologic gastritis was higher (p < 0.05) and the granulocytic infiltration more marked (p < 0.01) than in seronegative ones. In cagA+ children, H. pylori eradication rate was significantly lower (p < 0.02). CONCLUSIONS: cagA testing may be of useful clinical interest because its positivity can imply a more severe gastritis and a lower susceptibility to eradication treatment. PMID- 11106042 TI - A single intramuscular dose of ceftriaxone changes nasopharyngeal bacterial flora in children with acute otitis media. AB - The increasing prevalence of drug-resistant bacteria is attributed to the extensive use of antibiotics, which causes selective pressure on the nasopharyngeal bacterial flora. Shortened courses of antibiotics have been proposed to decrease the development of resistant strains. We determined the effect of a single intramuscular dose of ceftriaxone (50 mg/kg) on the nasopharyngeal bacterial flora in 167 children (median age 13 mo) with acute otitis media. Nasopharyngeal samples for bacterial culture were obtained before and 5 d after treatment with ceftriaxone. Before treatment, Moraxella catarrhalis was isolated in 99 (59%) children, Streptococcus pneumoniae in 87 (52%), and Haemophilus influenzae in 53 (32%). After treatment, M. catarrhalis was found in 62 (37%) children, which constitutes a 37% decrease in the colonization rate by this pathogen (p < 0.001). S. pneumoniae was isolated in 50 (30%; 43% decrease) and H. influenzae in 17 (10%; 68% decrease) children after treatment (p < 0.001 for both). Before treatment, 60% of pneumococcal isolates were sensitive to penicillin, 26% were of intermediate susceptibility, and 14% were penicillin resistant. Eradication of S. pneumoniae occurred mainly in children with penicillin-sensitive isolates. As a consequence, only 24% of pneumococcal isolates that remained after treatment were sensitive to penicillin, 59% were penicillin-intermediate, and 16% were penicillin-resistant. A single dose of ceftriaxone resulted in significant changes in the nasopharyngeal bacterial flora, increasing the relative prevalence of pneumococcal strains with decreased susceptibility to penicillin. PMID- 11106043 TI - Cervical lymph node infections with non-tuberculous mycobacteria in preschool children: interferon gamma deficiency as a possible cause of clinical infection. AB - Epidemiological studies have demonstrated a prevalence of positive PPD reactions to non-tuberculous mycobacteria (NTM) in 9% of 4-5-y-old children in the Goteborg area. Very few children of this age develop suppurative infections in lymph nodes that require surgical procedures. The hypothesis was that these children might have T cell deficiencies with abnormalities of macrophage functions, particularly with type 1 cytokines. Twenty-four children who needed operations were investigated immunologically and compared to 10 children of the same age operated on for non-infectious reasons. The methods used were flow cytometry analysis of lymphocytes in blood, blood lymphocyte stimulation assays and interferon gamma analyses. The patients had significantly lower levels of interferon gamma than the controls after stimulation with Candida antigens or Con A. The numbers of T and B lymphocytes were higher in patients than in controls. CONCLUSION: Children with necrotic lymph node infections in the cervical region due to NTM had lower interferon gamma production after stimulation than healthy age-matched controls. PMID- 11106044 TI - Heart rate variability in infants with apparent life-threatening events. AB - Heart rate variability (HRV) is often used as an index of sympatho-vagal balance. A decreased HRV has been observed in patients with central hypoventilation and in infants who have later succumbed to sudden infant death syndrome (SIDS). The aim of the present study was to investigate whether HRV is altered in infants with apparent life-threatening events (ALTE), a group with an increased risk of SIDS. Fifty infants with ALTE were compared with 50 age- and sex-matched controls. ECG was recorded overnight in all infants. Two sequences of RR intervals free of artefacts were selected from each sleep state and spectral analysis of RR variability was performed. The mean and SD of RR and the low (LFPow) and high (HFPow) frequency power were analysed. In active sleep (AS) the LF/HF ratio was lower in ALTE infants, but no differences were seen in either the LFPow or the HFPow. In quiet sleep (QS), however, ALTE infants had higher SD-RR (p = 0.006), greater HFPow (p = 0.02) and VLFPow (very low frequency power, p = 0.02) than the control infants. The same results were seen when the two sleep states were combined for analysis, ALTE infants had higher SD-RR (p = 0.004), HFPow (p = 0.006) and VLFPow (p = 0.04). CONCLUSION: The different HRV pattern in ALTE infants compared to healthy controls suggests an altered autonomic control. PMID- 11106045 TI - Abnormal fetal aortic velocity waveform and postnatal growth. AB - Postnatal growth from birth up to 7 y of age was evaluated in 151 children with varying degrees of intrauterine growth retardation who were previously examined in their intrauterine life with Doppler velocimetry of the thoracic descending aorta. The children with abnormal fetal aortic blood flow class (BFC), of which 39/46 (85%) had a birthweight > or = 2 SD below the mean of the population, were lean at birth and had a high rate of catch-up growth in weight and length during the first 3 and 6 mo, respectively. After the initial phases of rapid catch-up in weight and length, mean values of SD scores for weight and height remained relatively unchanged up until 2 y of age, thereafter increasing gradually up to 7 y of age, leaving 4/46 (8%) and 4/46 (8%) below -2 SD for weight and height, respectively. The pattern of changes in length/height and weight over time did not differ between those infants with abnormal BFC and those with normal BFC. The abnormal fetal aortic waveform was not related to rate of early catch-up growth or to height or weight at 7 y of age after adjustment for deviation in growth at birth. The magnitude of deficit in weight and length at birth was more predictive of subsequent growth. PMID- 11106046 TI - Potential role of colour-Doppler cystosonography with echocontrast in the screening and follow-up of vesicoureteral reflux. AB - Primary vesicoureteral reflux is a predisposing factor for urinary tract infections in children. The first-choice technique for the diagnosis of vesicoureteral reflux is voiding cystourethrography, followed by cystoscintigraphy; cystoscintigraphy, however, has the advantage of only minor irradiation of the patient, but it does not allow the morphological evaluation of bladder and vesicoureteral reflux grading. Colour-Doppler cystosonography with echocontrast is a recently introduced method for imaging vesicoureteral reflux. The aim of our study is to evaluate the role of colour-Doppler cystosonography with echocontrast in the diagnosis of vesicoureteral reflux. Twenty children (11M, 9F) aged between 0.4 and 4.9 y underwent colour-Doppler cystosonography using a diluted solution of Levovist (Schering, Germany), after filling up the bladder with saline. In all patients, vesicoureteral reflux diagnosis and grading had been performed previously by voiding cystourethrography within 5 d from ultrasonography. Our data showed high accuracy in the detection of medium to severe vesicoureteral reflux (grades III-V), confirmed by radiological features in 9/9 patients. Conversely, in the 11 patients with mild vesicoureteral reflux (grades I-II), this technique showed extremely low sensitivity, allowing diagnosis in only four cases. CONCLUSIONS: Colour-Doppler cystosonography, because of the absence of ionizing radiations, has great advantages, particularly in patients needing prolonged monitoring. Despite experiences reported in the literature, this technique has a role in the diagnosis of vesicoureteral reflux. Our group chooses colour-Doppler cystosonography for the follow-up of medium severe grade vesicoureteral reflux already diagnosed by radiology and/or scintigraphy. Cystoscintigraphy is employed only to confirm cases resulting negative at ultrasonography. PMID- 11106047 TI - Efficacy and safety of rectal thiopental sedation in outpatient echocardiographic examination of children. AB - The efficacy and safety of rectal thiopental administration in sedation for paediatric echocardiographic examination were prospectively investigated in infants with known or suspected congenital heart disease in an outpatient manner. A total of 1150 patients (546F, 604M) were studied; 264 were 7 d to 6 mo old (group I), 572 were 6 mo to 2-y-old (group II), and 314 were 2 to 6-y-old (group III). Thiopental sodium dissolved in 10 ml of water in a syringe to which a 6-F feeding catheter was attached was administered prior to echocardiographic examination to patients in groups I, II and III with doses of 50, 35 and 25 mg/kg, respectively in an emergency care environment. Length of time to achieve sedation (induction time), duration of sedation, length of time to return to normal activity (recovery time), whether sedation was successful and side effects were recorded. In the overall study population, sedation was successful in 1094 (95.1%) of the patients, the induction time was 16.34 +/- 3.69 min, the duration of sedation was 35.07 +/- 7.04 min, the recovery time was 63.25 +/- 10.17 min and the overall side-effect prevalence was 2%. Sedation was significantly more successful, the induction time was significantly shorter, the recovery time was significantly longer and side effects significantly more prominent in groups I and II compared to group III. CONCLUSION: Rectally administered thiopental is a safe and efficacious agent for sedation of infants and young children with known or suspected congenital heart disease who are undergoing echocardiography in an outpatient cardiology clinic, provided that it is used in an emergency care setting considering the risk of respiratory depression even though the prevalence of this side effect is significantly low. PMID- 11106048 TI - Outcome of congenital heart defects--a population-based study. AB - In a population-based study including 35,218 infants born alive during the 15-y period 1982-96, 360 (1%) were diagnosed as having a congenital heart defect (CHD). At a follow-up 3-18 y after birth (median 9.5 y) 154 patients (42.8%) were spontaneously cured; of these, 142 (92.2%) had ventricular septal defects (VSDs). Forty-two patients (11.7%) died, 22 of these (52.4%) during the neonatal period (0-28 d after birth). A total of 119 patients (33.1%) underwent therapeutic procedures (surgery, catheter interventions), 24 (20.2%) of whom died. Of the 95 children surviving therapeutic procedures, 54 (56.8%) had their defects completely repaired, while 41 (43.2%) had residual defects or cardiac sequelae, often of minor importance. In 69 children (19.2%) with persistent non-operated defects, 43 (62.3%) had VSDs. A chromosomal disorder, syndrome or associated extracardiac malformation occurred in 72 children (20%). CONCLUSIONS: The study underlines the broad variety in severity of CHDs, with a high neonatal mortality rate as well as a high rate of spontaneous cure. It is estimated that 25% of infants born with a CHD will grow into adult age with persistent non-operated defects, residual defects or cardiac sequelae after therapeutic procedures. PMID- 11106049 TI - Comparison of self-administration and face-to-face interview for surveys of low back pain in adolescents. AB - The aim of this study was to compare self-administration with face-to-face interview in the investigation of low back pain in adolescents. Fifty-seven adolescents with low back pain (mean age 17.3 y; range 16-18) first completed a questionnaire and were then invited to an interview. Analysis included item completion, percentages of agreement and weighted kappa. Item completion rates were high and comparable between self-administration and interview. Information between both methods was analogous for severity and localization of problems, sleep behaviour, medical consultation and sports/leisure activities. Onset, progression, duration of low back pain and some items on the influence of movement/positions presented less comparable data. CONCLUSION: Even when two different methods of data acquisition are used, results suggest that the method used does not change the interpretation of results in the case of most items. PMID- 11106050 TI - Does early hospital discharge with home support of families with preterm infants affect breastfeeding success? A randomized trial. AB - The aim of the present study was to determine if earlier discharge of preterm infants (<37 wk) from hospital is safe and if it affects breastfeeding rates. In a pilot observational study, premature infants received full oral (sucking) feeds for a mean (SD) 7.7 +/- 7.9 d before discharge. In the main study, 308 preterm infants were randomly assigned to either Early Discharge (148 infants) when fully orally fed but not yet gaining weight or Routine Discharge (160 infants) when fully orally fed and also gaining weight before discharge. A further 122 mothers declined randomization. The Early Discharge group was followed by Visiting Nurse Specialists who were available 24 h a day, while the Routine group was followed by the Home Care Nurses available on week days. There were no significant differences between the groups in birthweight or gestational age. The Early Discharge group were discharged 2.5 +/- 2 d after full oral feeding compared to 4.4 +/- 2.7 d for the Routine group (p < 0.001) and 6.1 +/- 5 d for those who declined. However, there was no significant difference between the Early and Routine groups for breastfeeding either at discharge (80 vs 83%), or 6 wk (55 vs 60%) or 6 mo after discharge (36 vs 36%), or for weight gain, or rates of re hospitalization (8.8% vs 11.9% at 6 wk, p = 0.37). CONCLUSION: Early discharge from hospital once a preterm infant can take full oral feeds does not alter later breastfeeding rates when adequate visiting nursing support is available. PMID- 11106051 TI - Central apnoea and endogenous prostaglandins in neonates. AB - AIM: Central apnoeas without an identifiable precipitating cause frequently occur in the neonatal period. Serious apnoeas should be treated with ventilation enhancing methylxanthines. Drugs such as opioids or prostaglandins (PGE2) are known to induce apnoea. PGE2 is an endogenous hormone that plays an important role in the regulation of neural activity and a relationship between PGE2 and central apnoeas has been postulated. METHODS: In order to test the hypothesis that the incidence of central apnoeas in preterm infants is related to endogenous PGE concentration, we measured the urinary concentration of PGE2 and PGE-M and determined the number of central apnoeas >10 s/12 h in overnight polygraphy in 18 preterm infants with apnoeas, bradycardias and desaturations, and 18 normal controls. RESULTS: We found 80.6 (SE 6.9) central apnoeas in the study group, and 52.9 (SE 4.1) in the control group (p = 0.002). Urinary PGE2 concentration was 25.9 (SE 6.1) ng/h/1.73 m2 in the control, 31.2 (SE 15.8) ng/h/1.73 m2 in the study group (p = n.s.), PGE-M concentration was 486 (SE 35) ng/h/1.73 m2 in the control and 1132 (SE 131) ng/h/1.73 m2 in the study group (p < 0.0001). There was a significant correlation between the number of central apnoeas and the PGE-M concentration in the study group (r = 0.68, p < 0.0001). CONCLUSION: Our results suggest a relationship between PGE and the respiratory system and open potential therapeutic options for the treatment of central apnoeas in neonates. PMID- 11106052 TI - Prophylaxis of patent ductus arteriosus with ibuprofen in preterm infants. AB - The aim of our study was to evaluate whether the prophylactic use of ibuprofen would reduce the incidence of significant patent ductus arteriosus (PDA) and to confirm the effectiveness of ibuprofen as rescue treatment in closing PDA. Eighty preterm infants with gestational age less than 34 wk with infant respiratory distress syndrome (iRDS) were randomized to receive intravenous ibuprofen lysine (10 mg/kg, followed by 5 mg/kg after 24 and 48 h) either within 24 h of life (group A) or after echocardiographic diagnosis of PDA (group B). To evaluate the severity of RDS in each patient, we calculated the initial and highest values of Oxygenation Index (O.I. = mean airway pressure x FiO2 x 100/PaO2) and Ventilatory Index (V.I. = O.I. x mechanical respiratory rate). Other studied variables were ventilatory support, renal function, biochemical and haematological profiles, frequency of bronchopulmonary dysplasia (BPD), intraventricular haemorrhage (IVH), necrotizing enterocolitis (NEC) and retinopathy of prematurity (ROP). On the 3rd day of life, 8% (3/40) of patients of group A and 53% of patients (21/40) of group B (p < 0.0001) developed a significant PDA. Between patients of group B who presented PDA at 3 d of life 90% (19/21) had a closure of ductus arteriosus after ibuprofen treatment. Initial and highest values of O.I. and V.I. were similar in both groups A and B. No significant differences between the groups were observed in regard to respiratory support, renal function and frequency of BPD, IVH, NEC and ROP. Ibuprofen was not associated with adverse effects. CONCLUSION: Prophylactic treatment with ibuprofen reduces PDA occurrence in preterm infants with iRDS at 3 d of life in comparison with rescue treatment, but both modes are effective in closing the ductus without significant adverse effects. PMID- 11106053 TI - Downregulatory cytokines in tracheobronchial aspirate fluid from infants with chronic lung disease of prematurity. AB - Chronic lung disease of prematurity (CLD) is associated with an inflammatory response in the preterm lung and increased levels of proinflammatory cytokines in tracheobronchial aspirate fluid (TAF). We investigated TAF levels of transforming growth factor-beta1 (TGF-beta1), interleukin-10 (IL-10), interleukin-4 (IL-4) and interleukin-12 (IL-12) cytokines possibly important in downregulating the proinflammatory response and/or inducing lung fibrosis in infants with developing and established CLD. Infants with CLD (n = 24) were compared with preterm infants with RDS that resolved (n = 22) and postoperative infants without lung disease (n = 23). TAF levels of TGF-beta1, IL-10, IL-4 and IL-12 were studied by quantitative enzyme immunoassay. Levels of TGF-beta1 were significantly higher during the first week of life in infants who developed CLD, remained high at 2 wk and past 4 wk of age. TAF levels of TGF-beta1 did not decrease significantly in six infants with CLD after treatment with steroids. TAF IL-10 was detected in 12/46 (26%) preterm infants. Infants with CLD or RDS were more likely to have measurable TAF levels of IL-10, compared with the postoperative infants without lung disease (p < 0.02 and 0.04, respectively). TAF levels of IL-4 or IL-12 were below the detection limits in all samples. CONCLUSIONS: We have demonstrated a sustained increase of TGF-beta1 levels in TAF from preterm infants who develop CLD, suggesting an important role for TGF-beta1 in the fibrotic response in the CLD lung. The elevated TGF-beta1 levels, combined with an absent or irregular secretion of IL-4, IL-10 and IL-12, can have importance for the increased tendency for the development of CLD in preterm infants. PMID- 11106054 TI - Ultrasonographic detection of very thin percutaneous central venous catheter in neonates. AB - To assess the ability of ultrasonography to detect the tip of a very thin (0.4 mm outer diameter) percutaneous central venous catheter (PCVC) in neonates, the PCVC tip location was assessed by ultrasonography (US) and compared to the location estimated by standard radiography for 57 PCVCs in 44 neonates. Of 57 occasions, the examiner could not find the PCVC tip in three cases (5%). In the remaining 54 instances, in 87% of cases, the PCVC tip position was consistent with the location implied by skeletal landmarks on standard radiographs. On 24 occasions we also assessed catheter tip dislodgement according to flexion and extension of the infant's arm. US could detect 78% of cases of catheter tip dislodgement. The PCVC tip was sometimes visualized as a dot and parallel lines as well as mere parallel lines. In a large population of cases, US is a reliable method for detection of a thin PCVC tip. US provides precise information about the PCVC tip position in relation to vascular structure and contributes to safer positioning of the PCVC than traditional radiography alone. PMID- 11106055 TI - Human immunodeficiency virus type-1: mother-to-child transmission. Meeting of World Federation of Scientists in Erice, Italy, August 2000. Joint report of AIDS/Infectious Diseases PMP and Mother and Child PMP. PMID- 11106056 TI - Severe adenovirus bronchiolitis in children. AB - Severe adenoviral infections such as the necrotizing adenovirus bronchiolitis occur sporadically in infants. Ascertaining the etiologic role of adenovirus in cases of lung disease can pose a diagnostic problem. We present two cases of severe bronchiolitis in previously healthy children in which adenovirus could be shown to be the causing agent. Both children received immunosuppressive therapy with steroids and Cyclosporin for 3 mo and a course of intravenous Ribavirin for 10 d. The results were conflicting: despite therapy Patient 1 died due to respiratory failure, Patient 2 improved notably. CONCLUSIONS: Adenovirus can cause severe bronchiolitis in previously healthy children. Diagnosis may be difficult to achieve. The role of antiviral therapy in the treatment of adenoviral infections remains to be cleared. PMID- 11106057 TI - Smoking preceded pulmonary involvement in adults with Langerhans cell histiocytosis diagnosed in childhood. AB - Two patients with childhood Langerhans cell histiocytosis (LCH) (aged 2 and 6 y at diagnosis) in whom pulmonary involvement was diagnosed in adulthood, 23 and 12 y later, respectively, are presented. In each patient, smoking preceded the diagnosis of pulmonary involvement by 3 y, providing further evidence that smoking is a risk factor in the development of pulmonary LCH. PMID- 11106058 TI - Multiorgan failure induced by intravenous immunoglobulin. PMID- 11106059 TI - Sex difference in hospital attendance rates. PMID- 11106060 TI - Fatty acid composition of human milk in Hungary. PMID- 11106061 TI - What's a scientific journal for? PMID- 11106062 TI - Pseudorabies virus and the functional architecture of the circadian timing system. AB - Transneuronal tracing of neuronal circuitry with neurotropic viruses has provided valuable insights in the way in which the nervous system imposes temporal organization on physiological processes and behavior. The swine alpha herpes virus known as pseudorabies virus, or PRV, has been particularly useful in this regard. Early studies identified attenuated mutants with selective tropism for visual circuitry involved in circadian regulation, and subsequent experiments employing this virus have provided considerable insight into the polysynaptic organization of the suprachiasmatic nuclei and associated circuitry. This literature, which has emerged during the past decade, is the subject of this mini review. PMID- 11106063 TI - The period E-box is sufficient to drive circadian oscillation of transcription in vivo. AB - The minimum element from the Drosophila period promoter capable of driving in vivo cycling mRNA is the 69 bp circadian regulatory sequence (CRS). In cell culture, an 18 bp E-box element from the period promoter is regulated by five genes that are involved in the regulation of circadian expression in flies. This E-box is a target for transcriptional activation by bHLH-PAS proteins dCLOCK (dCLK) and CYCLE (CYC), this activation is inhibited by PERIOD (PER) and TIMELESS (TIM) together, and inhibition of dCLK/CYC by PER and TIM is blocked by CRYPTOCHROME (CRY) in the presence of light. Here, the same 18 bp E-box region generated rhythmic expression of luciferase in flies under both light-dark cycling and constant conditions. Flies heterozygous for the Clke(jrk) mutation maintained rhythmic expression from the E-box although at a lower level than wild type. Homozygous mutant Clk(jrk) animals had drastically lowered and arrhythmic expression. In a per01 background, expression from the E-box was high and not rhythmic. Transcription mediated by the per E-box was restricted to the same spatial pattern as the CRS. The per E-box DNA element and cognate binding proteins can confer per-like temporal and spatial expression. This demonstrates in vivo that the known circadian genes that form the core of the circadian oscillator in Drosophila integrate their activities at a single DNA element. PMID- 11106065 TI - Squaring up the E-box. PMID- 11106064 TI - Specific sequences outside the E-box are required for proper per expression and behavioral rescue. AB - A 69 bp circadian regulatory sequence (CRS) upstream of the per gene is sufficient to drive circadian transcription, mediate proper spatial expression, and rescue behavioral rhythmicity in per01 flies. Within the CRS, an E-box is required for transcriptional activation by two basic-helix-loop-helix (bHLH) PERARNT-SIM (PAS) transcription factors, dCLOCK (dCLK) and CYCLE (CYC). To define sequences within the CRS that are required for spatial expression, circadian expression, and behavioral rhythmicity, a series of mutants that alter blocks of 3 to 12 nucleotides across the entire CRS were used to drive lacZ or per expression in vivo. As expected, the E-box within the CRS is necessary for high level expression and behavioral rhythmicity, but sequences outside the E-box are also required for transcriptional activation, proper spatial expression, and behavioral rhythmicity. These results indicate that the dCLK-CYC target site extends beyond the E-box and that factors other than dCLK and CYC modulate per transcription. PMID- 11106066 TI - Effects of light intensity and restraint on dark-pulse-induced circadian phase shifting during subjective night in Syrian hamsters. AB - Dark pulses presented on a background of constant light (LL) result in phase advances during midsubjective day and early subjective night, and phase delays during late subjective night, as shown in the dark-pulse phase response curve. In hamsters, the phase-shifting effects of dark pulses are thought to be mediated by increased activity, as previous studies have shown that restraining animals during dark pulses blocks the phase shifts observed in midsubjective day and late subjective night. This study focuses on dark-pulse-induced phase shifting during early subjective night, examining the influence of both LL intensity and restraint on the magnitude of these phase shifts. Syrian hamsters were maintained in LL of four different illumination levels (1, 10, 100, or 600 lux) and periodically presented with 6-h pulses (dark pulse alone, restraint alone, or dark pulse plus restraint) beginning at circadian time 11. Phase advances were observed in response to dark pulses alone, and the magnitude of these shifts was dependent on background illumination, with significantly larger advances seen under higher intensities. No relationship was found between the amount of activity displayed during dark pulses and phase shift magnitude. Six-hour periods of restraint resulted in phase delays, the magnitude of which was also dependent on background illumination. Restraining hamsters during dark pulses reduced the magnitude of phase advances, but the extent of this reduction could be predicted from the additive effects of the dark-pulse-alone and restraint-alone conditions. These results indicate that the phase-shifting effects of dark pulses during early subjective night are not mediated by behavioral activation and may instead reflect a mirror image of the phase-delaying effects of light pulses at this phase. PMID- 11106067 TI - Tau differences between short-day responsive and short-day nonresponsive white footed mice (Peromyscus leucopus) do not affect reproductive photoresponsiveness. AB - In laboratory-bred rodent populations, intraspecific variation in circadian system organization is a known cause of individual variation in reproductive photoresponsiveness. The authors sought to determine whether circadian system variation accounted for individual variation in reproductive photoresponsiveness in a single, highly genetically variable population of Peromyscus leucopus recently derived from the wild. Running-wheel activity patterns of male and female mice, aged 70 to 90 days, from artificially selected lines of reproductively photoresponsive (R) and nonresponsive (NR) lines were monitored under short-day photoperiod (8 h light, 16 h dark), long-day photoperiod (16 h light, 8 h dark), and constant darkness (DD). NR mice displayed a significantly longer mean free-running period (24.08 h) in DD compared with R mice (23.75 h), due in large part to a difference between NR and R females (24.25 h vs. 23.74 h, respectively). All other entrainment characteristics (alpha, phase angle of activity) under short days, long days, and DD were similar between R and NR mice. Variation in free-running period and entrainment characteristics has been shown to affect photoresponsiveness in other rodent species by altering the manner in which the circadian system interprets short days. To determine whether variation in photoresponsiveness in P. leucopus is due to differences in free-running period instead of variation downstream from the central circadian clock in the pathway controlling photoresponsiveness, the authors exposed young R and NR mice to DD and measured the effect on reproductive organ development. If variation in free-running period affected how the circadian system of mice interpreted short days, then both R and NR mice exposed to DD should have exhibited a delay in gonadal development. Only R mice exhibited pubertal delay in DD. NR mice exhibited large paired testes, paired seminal vesicles, paired ovaries, and uterine weight typical of mice nonresponsive to short days, whereas R mice exhibited reproductive organ weight typical of mice responsive to short days. These data suggest that despite significant differences in free-running period between R and NR mice, individual variation in photoresponsiveness is not due to differences in how the circadian systems of R and NR mice interpret the LD cycle. PMID- 11106068 TI - The coincidence of light and melatonin with a specific phase of the circadian pacemaker is important for the timing of seasonal breeding in the ewe. AB - The timing of reproductive activity in seasonal breeding sheep relies on daily photoperiodic signals being relayed to provide information on the time of year. Although light and melatonin are involved, the exact mechanism is not understood. In this experiment, three groups of 6 Romney Marsh ewes, a highly seasonal breed, were provided with 8 weeks of short nights (9.6-9.8 h, by artificially advancing dawn) around the winter solstice, near the end of their natural breeding season. One group of animals was infused to a physiological level with melatonin for 5 h during the afternoon prior to the onset of dark, while a second group was identically infused but for 5 h from the time of lights on. A third group received the short-night treatment only. Following the short-night treatment, all groups were exposed to long nights (> 14 h, by delaying dawn) until the summer solstice. Ovarian activity, assessed by progesterone monitoring twice weekly, showed that the noninfused and the morning-infused groups displayed renewed reproductive activity in response to the short-night/long-night treatment. There was no renewed ovarian activity in the afternoon-infused group, indicating that the time of day that melatonin is present, rather than the duration of melatonin exposure, is an important signal in the control of reproductive timing. Measurements of a marker of the endogenous circadian pacemaker, by melatonin measurements under acutely extended darkness, revealed that the short-night treatments phase advanced the onset of the pacemaker in all groups such that the afternoon phase of the pacemaker was coincident with light. The results provide strong support for the model that proposes that an afternoon-located sensitive phase of the pacemaker is responsible for the relay of photoperiodic signals in the timing control of seasonal breeding. The model proposes that the reproductive axis be primed during short nights when the sensitive phase is coincident with light in the afternoon so ovarian activity can be induced when the sensitive phase is located within the longer nights of autumn and coincident with endogenous melatonin. PMID- 11106070 TI - A journal by any other name. PMID- 11106069 TI - The timing of the human circadian clock is accurately represented by the core body temperature rhythm following phase shifts to a three-cycle light stimulus near the critical zone. AB - A double-stimulus experiment was conducted to evaluate the phase of the underlying circadian clock following light-induced phase shifts of the human circadian system. Circadian phase was assayed by constant routine from the rhythm in core body temperature before and after a three-cycle bright-light stimulus applied near the estimated minimum of the core body temperature rhythm. An identical, consecutive three-cycle light stimulus was then applied, and phase was reassessed. Phase shifts to these consecutive stimuli were no different from those obtained in a previous study following light stimuli applied under steady state conditions over a range of circadian phases similar to those at which the consecutive stimuli were applied. These data suggest that circadian phase shifts of the core body temperature rhythm in response to a three-cycle stimulus occur within 24 h following the end of the 3-day light stimulus and that this poststimulus temperature rhythm accurately reflects the timing of the underlying circadian clock. PMID- 11106071 TI - Small lymphocytic lymphoma with perifollicular, marginal zone, or interfollicular distribution. AB - Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) typically involves nodal or extranodal tissues as a diffuse proliferation with pseudofollicular growth centers obliterating normal architecture. We describe 16 cases of CLL/SLL in which the neoplasm was confined to the marginal zone, perifollicular, or interfollicular regions surrounding benign lymphoid follicles in either nodal or extranodal sites. Twelve of 12 (100%) patients with adequate staging data had disseminated disease (Stage III or IV) at presentation. Eight of the 16 (50%) patients had absolute peripheral lymphocytosis (range, 5 to 30 x 10(9)/L). Pseudofollicular growth centers were identified in 14 of 16 cases (87.5%). Immunophenotypic studies revealed that the tumor cells were positive for CD20 (16/16) and CD5 (11/11) in all cases examined. CD23 was positive in 12 of 14 (86%) interpretable cases. IgM and IgD were positive in 13 of 14 (93%) and 10 of 10 (100%) interpretable cases, respectively. All cases were negative for CD3 (16/16), CD45RO (16/16), CD10 (15/15), and cyclin D1 (15/15). We conclude that CLL/SLL can have unusual patterns of involvement, including marginal zone, perifollicular, and interfollicular patterns that can be difficult to recognize histologically. Thirteen of 16 (81%) cases in this study were misinterpreted by the referring pathologists. Recognition of proliferation centers coupled with demonstration of a CD5+ CD23+ B-cell immunophenotype establishes the correct diagnosis of CLL/SLL. PMID- 11106072 TI - The expression of cyclins D1 and E in predicting short-term survival in squamous cell carcinoma of the lung. AB - Cyclins D1 (cD1) and E (cE) are G1 phase cyclins believed to participate in the pathogenesis of malignancy. Overexpression of cD1 has been reported to influence prognosis in squamous cell carcinomas (SCC) of the larynx, but was not significant in a limited study of non-small cell lung cancers (NSCLC). Altered expression of cE has been proposed as another potential prognostic marker in malignancy but its possible role in NSCLC has not been elucidated. In order to determine the prognostic value of cD1 and cE in NSCLC, paraffin-embedded sections of 467 NSCLC were immunostained with monoclonal antibody to cD1 (1:500, PharMingen, San Diego, CA) and 400 NSCLC with MA to cE (1:2500, PharMingen) using an enhanced sensitivity avidin-biotin complex technique. The number of tumor cells with nuclear and/or cytoplasmic immunopositivity was graded on a scale of: 0 = less than 1%, 1 = 1 to 10%, 2 = 10 to 25%, 3 = 25 to 50%, 4 = 50 to 75%, 5 = more than 75%. Results were correlated with survival by Kaplan-Meier survival plot using Stat-View software (Abacus Concepts, Berkeley, CA). Overall, 426 NSCLC with cD1 and 360 NSCLC with cE had adequate follow-up (median, 76 mo) for survival analysis. Both cyclins independently showed significance in prognosis of SCC but not other cell types. For cD1, absence of immunostaining was associated with worse prognosis than any immunopositivity for all stages of SCC (P = .025). For cE, Stage I and II SCC with less than 50% immunopositivity had a worse prognosis (P = .029). Of 70 Stage I and II SCC immunostained for both monoclonal antibodies, 55% of patients with tumors that demonstrated both absence of cD1 staining and cE immunopositivity in less than 50% of cells were dead at 5 years compared to 35% of patients with tumors that demonstrated positive staining with cD1 and cE immunopositivity in more than 50% of cells. These results strongly suggest cD1 and cE can independently predict prognosis in early stage SCC. Worse prognosis was associated with loss of expression, consistent with mechanisms other than overexpression of these cyclins in the progression of SCC. PMID- 11106073 TI - Correlation of p53 mutations in ThinPrep-processed fine needle breast aspirates with surgically resected breast cancers. AB - Mutations of the p53 gene are one of the most common genetic changes found in cancer; their presence may be prognostic and even influence treatment for breast cancer. In this study, we investigated whether DNA could be extracted from the residual cells left in ThinPrep-processed breast fine-needle aspirates and whether p53 gene changes could be detected in the DNA. The results were then correlated with DNA extracted from the matched formalin-fixed, paraffin-embedded, surgically resected breast cancer when available. DNA was successfully extracted from 54 of 62 aspirates and all 31 surgical specimens. p53 gene mutations were detected in 10 of the 54 cytology specimens (18.5%) and consisted of base pair substitutions or deletions. Silent or intronic p53 changes were found in five additional aspirates. One of the aspirates had two gene alterations, resulting in a total of six gene changes. Five of these changes were located in introns 6 or 9 and the sixth was a silent (no amino acid change) change in exon 6. p53 Polymorphisms were detected in nine aspirates (16.3%) and were located in codon 47 (one aspirate), codon 72 (six aspirates), and codon 213 (two aspirates). All cases with surgical material available showed identical p53 mutations, alterations, and polymorphisms in the resected tumors compared with those detected in the corresponding aspirates. The results of this study show that DNA suitable for analysis of p53 gene sequence changes can be successfully extracted from ThinPrep-processed breast fine-needle aspirates, and that identical alterations are detected in both the cytology and surgical specimens. PMID- 11106074 TI - Microsatellite analysis in post-transplantation lymphoproliferative disorder to determine donor/recipient origin. AB - Post-transplantation lymphoproliferative disorders (PTLD) are a group of heterogeneous diseases that occur after organ transplantation. Determination of the origin of the tumor cells not only provides clues to its possible pathogenetic mechanism, but also gives prognostic guidance in the clinical management of patients. We reviewed the clinicopathological features of four cases of PTLD that developed after solid organ transplantation. Using microsatellite analysis performed on paraffin-embedded tissue and using multiple, highly polymorphic markers, we have successfully determined the recipient/donor origin of the tumor cells in all of them. The time of onset of the PTLD ranged from 5 to 11 mo. All cases were diffuse large cell lymphomas of B-cell lineage, and the two cases that have been tested for EBV by in situ hybridization were positive. Three of the 4 PTLD were of donor origin and these three patients died of diseases unrelated to PTLD. The single patient with PTLD of recipient origin died of disseminated PTLD. The mean survival length of the three patients with donor origin was 26.3 mo, whereas that of the patient with recipient origin was 12 mo. Our results indicate a relatively high incidence of PTLD of donor origin among our patients with solid organ transplantation, as compared to other reported series. Moreover, the finding of the relatively indolent nature of PTLD of donor origin supports that determination of the donor/recipient origin of PTLD is of prognostic significance. PMID- 11106075 TI - Immunophenotype of high-grade prostatic adenocarcinoma and urothelial carcinoma. AB - Morphologic features alone can usually be used to distinguish prostatic adenocarcinoma and urothelial carcinoma of the urinary bladder. Poorly differentiated tumors, however, can occasionally have features of both neoplasms, making determination of site of origin difficult. No study has provided a panel of antibodies to assist in the distinction of these two tumors. For this study, 73 examples of moderately and poorly differentiated prostatic adenocarcinoma and 46 examples of high-grade urothelial carcinoma were obtained from radical resection specimens. Immunohistochemical studies were performed using the following panel of antibodies: cytokeratin (CK) 7, CK 20, 34betaE12, Leu M1, carcinoembryonic antigen (CEA)m, CEAp, p53, Leu 7, prostate-specific acid phosphatase (PSAP), prostate-specific antigen (PSA), and B72.3. Mucicarmine was also performed. Intermediate and high-grade prostatic carcinoma were compared and then high-grade prostatic carcinoma was compared with high-grade urothelial carcinoma. PSA and PSAP each stained 94% of prostatic adenocarcinomas, but no urothelial carcinomas. Leu 7 stained 94% of prostate and 17% of urothelial carcinomas. Over half of the urothelial carcinomas showed positivity for 34betaE12 (65%), as did two cases of prostatic carcinoma (6%). Eighty-three percent of urothelial carcinomas and 12% of prostatic adenocarcinomas stained with CK 7. Forty-one percent of urothelial carcinomas and 12% of prostatic carcinomas were reactive for CEAm, and p53 stained 33% and 3% of urothelial and prostatic adenocarcinomas, respectively. No significant difference was seen in the expression of CEAp, CK 20, B72.3, Leu M1, or mucicarmine between prostate and urothelial carcinoma. We propose a panel of six antibodies to assist in the distinction of high-grade prostatic adenocarcinoma from high grade urothelial carcinoma: PSA, PSAP, 34betaE12, Leu 7, CK 7, and p53. The first three antibodies should be used initially; if results are negative, the remaining antibodies may be employed. PMID- 11106076 TI - Hemosiderotic fibrohistiocytic lipomatous lesion: ten cases of a previously undescribed fatty lesion of the foot/ankle. AB - We address the clinicopathologic features of a previously undescribed heavily pigmented spindle cell proliferation within a circumscribed benign lipomatous lesion that occurs mainly in the ankle region of older females. Patients with "lipoma with fibrohistiocytic proliferation" were retrieved from our files. Slides and clinical information were reviewed, and immunohistochemistry was performed (n = 5). Ten patients with hemosiderotic fibrohistiocytic lipomatous lesions were identified. All cases demonstrated a well-circumscribed fatty lesion with random focal proliferations of plump, slightly pleomorphic spindled cells, scattered inflammatory cells, and abundant iron pigment. The spindled cells had vesicular nuclei with indistinct nucleoli; occasional hyperchromatism was observed. No nuclear cytoplasmic inclusions were identified. The spindled component had a reactive appearance. In most cases, the fatty component, with homogeneously sized adipocytes, predominated. The lesions occurred in the foot/ankle region (8/10, one each cheek and hand) of primarily females (8/10) with a mean age of 50.6 years (range 42-63 years), size of 7.7 cm (range 2.5-17 cm), and prior duration of 3.1 years. Seven of eight patients had a history of prior trauma. The spindled component was positive for vimentin, calponin, CD34, and occasionally KP-1 or lysozyme and negative for caldesmon, S100, and desmin. Follow-up on eight patients revealed four with recurrences or residual disease over three years, requiring re-excision. No cases metastasized or caused patient death (mean 12 years, range 1-23 years). We describe a predominantly fatty lesion that is hemosiderin rich with a "fibrohistiocytic" proliferation, composed of histiocytes, myofibroblasts, and C34-positive fibroblasts, which occurs predominantly in the ankle region of middle-aged females. We believe that this is a reactive process due to antecedent trauma, the inflammatory cells, hemosiderin, mixed spindled cells, and homogeneous non-neoplastic appearance of the fat. HFLL can be distinguished from previously described lesions. Correct identification of hemosiderotic fibrohistiocytic lipomatous lesion is important, as it may locally recur. PMID- 11106077 TI - Acute encephalomyelitis during an outbreak of enterovirus type 71 infection in Taiwan: report of an autopsy case with pathologic, immunofluorescence, and molecular studies. AB - We report a fatal case of enterovirus type 71 (EV 71) infection in an 8-year-old girl during a summer outbreak of hand, foot, and mouth disease in 1998 in Taiwan. The clinical course was rapidly progressive, with manifestations of hand, foot, and mouth disease, aseptic meningitis, encephalomyelitis, and pulmonary edema. The patient died 24 hours after admission. Postmortem study revealed extensive inflammation in the meninges and central nervous system and marked pulmonary edema with focal hemorrhage. Brain stem and spinal cord were most severely involved. The inflammatory infiltrates consisted largely of neutrophils involving primarily the gray matter with perivascular lymphocytic cuffing, and neuronophagia. The lungs and heart showed no evidence of inflammation. EV 71 was isolated from the fresh brain tissues and identified by immunofluorescence method with type-specific EV 71 monoclonal antibody. It was also confirmed by neutralization test and reverse-transcriptase polymerase chain reaction with sequence analysis. The present case was the first example in which EV 71 was demonstrated to be the causative agent of fatal encephalomyelitis during its epidemic in Taiwan. PMID- 11106078 TI - Cytogenetic-morphologic correlations in aneurysmal bone cyst, giant cell tumor of bone and combined lesions. A report from the CHAMP study group. AB - Aneurysmal bone cyst and giant cell tumor of bone are relatively rare bone tumors that sometimes coexist. We examined the karyotypes of 3 aneurysmal bone cysts, 12 giant cell tumors, and 3 combined lesions. All aneurysmal bone cysts showed involvement of chromosome segments 17p11-13 and/or 16q22. In addition, in 1 of the 3 giant cell tumors with secondary aneurysmal bone cyst, both chromosome bands were rearranged as well, albeit not in a balanced translocation. Seven out of 12 giant cell tumors were characterized by telomeric associations. One giant cell tumor showed a dup(16)(q13q22), suggesting the presence of a (minor) secondary aneurysmal bone cyst component, despite the absence of histological proof. Our results, combined with literature data further substantiate that segments 16q22 and 17p11-13 are nonrandomly involved in at least some aneurysmal bone cysts, irrespective of subtype (primary, secondary, intra/extraosseous, solid or classic). These findings strongly suggest that some aneurysmal bone cysts are true neoplasms. In addition, telomeric associations are the most frequent chromosomal aberrations in giant cell tumor of bone, the significance of which remains elusive. In combined giant cell tumor/aneurysmal bone cyst each component seems to retain its own karyotypic abnormality. PMID- 11106079 TI - Rhabdoid features in leiomyosarcoma of soft tissue: with special reference to aggressive behavior. AB - The presence of rhabdoid cells has been reported in various types of malignant neoplasms and has been determined to be a predictor of aggressive behavior of neoplasms regardless of tumor histogenesis. One hundred and thirteen cases of leiomyosarcoma, selected from 1800 soft tissue sarcomas, were reviewed on hematoxylin and eosin sections, and immunohistochemical staining when available, and seven cases with rhabdoid features were retrieved. Clinicopathologic differences were analyzed to compare between cases with rhabdoid features and those without rhabdoid features. In the seven cases with rhabdoid features, two were intra-abdominal, and the others arose in external soft tissues including muscle, subcutis, and cutis. Patient age ranged from 33 to 84 years, three were female, and four were male. Tumor size ranged from 3 to 22 cm. Clinical evidence showed no differences from those cases without rhabdoid features. Histologically, one of the abdominal cases was epithelioid leiomyosarcoma. Two of the 7 cases were better subclassified as pleomorphic leiomyosarcoma, in which rhabdoid cells are diffusely scattered. In cases other than those with pleomorphic leiomyosarcomas, foci of anaplastic areas were observed, and collections of rhabdoid cells were present in those areas. Immunohistochemical examination of the cases confirmed myogenic differentiation, and showed rhabdoid cells being positive for vimentin and desmin in the inclusion bodies, and diffusely so for muscle actin in the cytoplasm. After dividing all the cases of leiomyosarcoma by their location, prognostic analysis was performed. Leiomyosarcoma of external soft tissue with rhabdoid cells showed a tendency for poorer prognoses than cases without rhabdoid features. On the contrary, retroperitoneal cases did not. This study indicates that rhabdoid features are associated with aggressive biological behavior in leiomyosarcoma of the external soft tissue. PMID- 11106080 TI - Immunophenotypic and genotypic markers of follicular center cell neoplasia in diffuse large B-cell lymphomas. AB - Diffuse large B-cell lymphomas (DLBCL) are a biologically and clinically heterogeneous entity. Although some DLBCL represent transformation of follicular lymphomas (FL), the proportion that is of follicular center cell (FCC) origin remains uncertain. Immunophenotypic and genotypic markers used to suggest a FCC origin for a lymphoma (bcl-6 and CD10 expression, lack of CD138 expression, bcl-2 rearrangements [R]) or to subdivide DLBCL (bcl-2 expression, bcl-6 R) were therefore investigated in 22 FL and 44 DLBCL using paraffin section immunostains and Southern blot/polymerase chain reaction analysis. All FL tested were bcl-6+ (19) and CD138- (22) with 16/19 also bcl-2 and CD10+ (classic phenotype), one bcl2+, CD10- (grade III) and two bcl2-, CD10+ (grade II or III). Bcl-2R was identified in 4/5 FL-GrI, 3/6 FL-GrII, and 1/3 FL-GrIII. Bcl-6R was found in 0/5, 2/4, and 0/3 FL, respectively. All but 3/41 DLBCL were bcl-6+ with 17/37 also bcl 2+ and CD10+. Three of these cases were also CD138+. Twelve bcl-6+ cases were bcl 2+, CD10-, six bcl-2-, CD10+, and two bcl-2-, CD10-. The three bcl-6- cases were bcl-2+, CD138- and two were CD10+. Bcl-2R was identified in 5/27 DLBCL with 4/5 bcl-2+, 3/4 tested CD10+ and 4/4 bcl-6+. Bcl-6R was identified in 7/26 including three with a classic FL phenotype. The vast majority of DLBCL in this study have an immunophenotype that supports a FCC origin. Although the proportion of DLBCL that co-expressed bcl-6, CD10 and bcl-2 was lower than for the FL, absence of bcl 2 or CD10 may be associated with higher grade FL It is also possible that bcl-6 expression is not completely specific for a FCC origin. Only a minority of cases suggested postfollicular differentiation. Only a minority of DLBCL show bcl-2R, suggesting that many have a different molecular pathogenesis than most low-grade FL. Bcl-6R did not exclude a FCC origin. PMID- 11106081 TI - Cutaneous dendritic cells are main targets in acute HIV-1-infection. AB - Acute human immunodeficiency virus (HIV) infection is a transient illness that typically presents with mucocutaneous and constitutional symptoms. It is soon followed by seroconversion with the detection of anti-HIV antibodies in the peripheral blood. To better understand the pathogenetic events leading to this clinical picture, we sought to investigate the (immuno)histologic features of the skin rash occurring in an acutely infected person. A skin biopsy of an acutely infected person was investigated histologically and immunohistologically using paraffin-embedded tissue sections. Interface dermatitis with pronounced vacuolization of the basal keratinocytes was a prominent histological finding. The inflammatory infiltrate was composed of CD3+/CD8+ T cells with coexpression of Granzyme B7 and TIA-1, and CD68+ histiocytes/dendritic cells. CD1a+ intraepidermal Langerhans cells (LC) were significantly decreased and individual LC coexpressed HIV-p24 antigens as evidenced in double labeling experiments. HIV infected LC were demonstrated in close apposition to cytotoxic T cells. This study provides the first definitive evidence for infection of LC at extramucosal sites in this very early stage of disease. Our findings emphasize the critical role of dendritic cells as a virus reservoir and the skin as a major site of HIV replication during the course of the disease. PMID- 11106082 TI - Strong correlation between results of fluorescent in situ hybridization and immunohistochemistry for the assessment of the ERBB2 (HER-2/neu) gene status in breast carcinoma. AB - ERBB2 (HER-2/neu) amplification and/or overexpression are associated with poor prognosis in node-positive breast carcinoma. Its prognostic value in node negative cases and its predictive value for response to chemotherapy remain controversial. This may be related to the use of molecular methods, which are sensitive to dilution of tumor material by normal cells, or the use of nonstandardized immunohistochemistry (IHC) procedures, for the determination of the ERBB2 gene status. In addition, new therapeutic approaches that target the cells overexpressing ERBB2 are under development. These perspectives necessitate a reliable evaluation of the status of ERBB2 in individual tumors before the application of specific therapeutic strategies. Fluorescent in situ hybridization (FISH) and IHC allow the evaluation of the ERBB2 status specifically in tumor cells on archival material. We have analyzed a series of 100 invasive ductal breast carcinomas without lymph node invasion both by IHC, using the CB11 monoclonal antibody and a sensitive Auidin Biotin Complex (ABC) immunodetection system, and by FISH, using the Oncor Inform HER-21neu (ERBB2) gene amplification detection system as reference technique. Complete concordance between the results of FISH and IHC was seen in 98% of the cases. ERBB2 amplification (more than four signals per nucleus) was observed in 12 of the 100 cases, and all but one showed an overexpression of the protein (membrane staining) by IHC. Conversely, ERBB2 expression was present in one case without gene amplification. In conclusion, ERBB2 overexpression detected by IHC is highly correlated to gene amplification detected by FISH. Thus, under standardized conditions, IHC is a reliable and economical test to assess the ERBB2 status in tumors. The use of FISH could be limited to the verification of the status of tumors displaying a weak membrane immunostaining. PMID- 11106083 TI - CD20-Positive peripheral T-cell lymphoma: report of a case after nodular sclerosis Hodgkin's disease and review of the literature. AB - We present a case of peripheral T-cell lymphoma co-expressing CD3 and CD20, as well as demonstrating T-cell receptor gene rearrangement, in a patient who had been diagnosed with nodular sclerosis Hodgkin's disease 5 years previously. Although 15 cases of CD20-positive T-cell neoplasms have been previously reported in the literature, this is the first report of CD20-positive T-cell lymphoma occurring subsequent to treatment of Hodgkin's disease. The current case affords an opportunity to review the rarely reported expression of CD20 in T-cell neoplasms as well as the relationship between Hodgkin's disease and subsequently occurring non-Hodgkin's lymphomas. In addition, the identification of this case supports the suggestion that the use of CD20 antibodies alone in paraffin sections may lead to an incorrect determination of cell lineage in some cases. PMID- 11106084 TI - Malignant peripheral nerve sheath tumors with t(X;18). A pathologic and molecular genetic study. AB - Spindle cell sarcomas often present the surgical pathologist with a considerable diagnostic challenge. Malignant peripheral nerve sheath tumor, leiomyosarcoma, fibrosarcoma, and monophasic synovial sarcoma may all appear similar histologically. The application of ancillary diagnostic modalities, such as immunohistochemistry and electron microscopy, may be helpful in the differentiation of these tumors, but in cases in which these adjunctive techniques fail to demonstrate any more definitive evidence of differentiation, tumor categorization may remain difficult. Cytogenetic and molecular genetic characterization of tumors have provided the basis for the application of molecular assays as the newest components of the diagnostic armamentarium. Because the chromosomal translocation t(X;18) has been observed repeatedly in many synovial sarcomas, it has been heralded as a diagnostic hallmark of synovial sarcoma. To formally test the specificity of this translocation for the diagnosis of synovial sarcoma, RNA extracted from formalin-fixed, paraffin-embedded tissue from a variety of soft tissue and spindle cell tumors was evaluated for the presence of t(X;18) by reverse transcriptase-polymerase chain reaction. Although 85% of the synovial sarcomas studied demonstrated t(X;18), 75% of the malignant peripheral nerve sheath tumors in our cohort also demonstrated this translocation. We conclude that the translocation t(X;18) is not specific to synovial sarcoma and discuss the implications of the demonstration of t(X;18) in a majority of malignant peripheral nerve sheath tumors. PMID- 11106085 TI - Clarithromycin resistance in Helicobacter pylori infection: does it matter? PMID- 11106086 TI - Herbal hepatotoxicity: an expanding but poorly defined problem. AB - Alternative therapies, including herbal remedies, are popular in the general population and even more so among patients with liver disease. The use of such products is now well established in western society and is no longer confined to traditional medicine practitioners in Asia, Africa and the Middle-East. Their perceived benefits remain generally unproven and concern about adverse effects is leading to closer scrutiny of these products. Herbal hepatotoxicity has been recognized for many years, but new agents are constantly being identified. The varied manifestations of liver injury include steatosis, acute and chronic hepatitis, hepatic fibrosis, zonal or diffuse hepatic necrosis, bile duct injury, veno-occlusive disease, acute liver failure requiring liver transplantation and carcinogenesis. Potential interactions between herbal medicines and conventional drugs may interfere with patient management. Concurrent use of such products is not often disclosed unless specifically sought after and can lead to perpetuation of the liver injury. The present review focuses on emerging herbal hepatotoxins, newer patterns of liver injury among the older agents and provides an updated tabulation of the adverse effects of major herbal hepatotoxins. Key issues of diagnosis and prevention of this growing problem are addressed. Continued public education, physician awareness and more stringent licensing are required to tackle this growing problem. PMID- 11106087 TI - Botulinum toxin for achalasia in children. AB - BACKGROUND: Injection of botulinum toxin (BTx) into the lower esophageal sphincter (LES) of adult patients with achalasia results in the effective relief of symptoms. The aim of the present study was to examine the effectiveness of BTx in pediatric patients suffering from achalasia. METHODS: Seven patients suffering from achalasia with or without prior treatment were treated with intrasphincteric injection of BTx. The median duration of follow up was 15 months. RESULTS: All seven patients improved. The median interval before recurrence of symptoms was 4 months (range 1-14 months). There was an inverse relationship between the pretreatment LES pressure and the duration of response (r=-0.6). The mean pretreatment LES pressure in the subgroup with a response greater than 6 months was 38+/-10 mmHg compared with 61+/-12 mmHg in the other four patients (P= 0.05). All seven patients required retreatment. CONCLUSION: Botulinum toxin is effective in relieving symptoms in pediatric patients suffering from achalasia, producing a sustained response beyond 6 months in 43% of patients. PMID- 11106088 TI - Association of Helicobacter pylori infection with atrophic gastritis and intestinal metaplasia. AB - AIMS: To evaluate the effect of Helicobacter pylori infection and aging on atrophy and intestinal metaplasia of the gastric mucosa. METHODS: One hundred and sixty-three patients were divided into three age groups and underwent an upper gastrointestinal endoscopy where no esophagitis, peptic ulcers, or malignancies were detected. Two biopsy specimens were obtained from the anterior and posterior walls of the antrum and of the fundus. These were used to evaluate the grade of gastritis, bacterial culture and histologic evidence of H. pylori infection. RESULTS: Helicobacter pylori infection was found to be directly associated with an increased risk of gastritis grade (odds ratio (OR) = 90 (95% CI; 30-270)). An age of 60 years and older along with H. pylori infection was also strongly associated with an increased risk of atrophy (OR = 6.6, (95% CI; 2.9-15.2)); OR = 9.8, (95% CI; 2.7-35.4)), as was intestinal metaplasia of the gastric mucosa (OR = 5.5, (95% CI; 1.7-17.6)); OR = 7.9, (95% CI; 2.8-46.1)). The prevalence of atrophic gastritis increased with advancing age in H. pylori-infected patients, but no such phenomenon was observed in H. pylori-uninfected patients. The prevalence of intestinal metaplasia significantly increased with advancing age, irrespective of the presence of H. pylori infection. In addition, H. pylori uninfected female patients had a decreased risk of intestinal metaplasia. CONCLUSIONS: These results suggest that atrophic gastritis is not a normal aging process, but instead is likely to be the result of H. pylori infection, while intestinal metaplasia is caused by both the aging process and H. pylori infection. A decreased risk of intestinal metaplasia found in uninfected female subjects may partly explain the lower prevalence of gastric cancer in females than in males. PMID- 11106089 TI - Endoscopic characteristics of low-grade gastric mucosa-associated lymphoid tissue lymphoma after eradication of Helicobacter pylori. AB - BACKGROUND AND AIMS: It was recently reported that low-grade gastric lymphoma of mucosa-associated lymphoid tissue (MALT) was regressed by the eradication of Helicobacter pylori. The aim of this study was to confirm the effect of H. pylori eradication on low-grade gastric MALT lymphoma and to investigate the whitish mucosa that appeared with regression of the lesions. METHODS: Forty-seven H. pylori-positive patients with low-grade gastric MALT lymphoma were treated by using triple therapy. Biopsy specimens were histologically graded and B cell clonality was examined by using reverse transcription-polymerase chain reaction before and after eradication treatment. The relationship between the appearance of whitish mucosa and the degree of gastric gland loss was evaluated. RESULTS: Histologic regression was observed 2 months after eradication therapy in 42 of 47 patients. However, B cell monoclonality changed to polyclonality in only 23 patients during the follow-up period. The appearance of whitish mucosa in patients who showed histologic regression became more frequent as the degree of gastric gland loss increased (P< 0.001). CONCLUSIONS: Most low-grade gastric MALT lymphoma histologically regressed after H. pylori eradication. The appearance of whitish mucosa after histologic regression reflected the degree of gastric gland loss. Whitish mucosa is an endoscopic characteristic and may be an endoscopic marker for regression of low-grade gastric MALT lymphoma. PMID- 11106090 TI - Vasoactive intestinal polypeptide appears to be one of the mediators in misoprostol-enhanced small intestinal transit in rats. AB - BACKGROUND AND AIMS: Prostaglandin analogs have the pharmacologic effect of speeding up small intestinal transit (SIT). It remains unknown whether some gut peptides also mediate this enhancement. We studied the effect of misoprostol on rat SIT and looked at the role of vasoactive intestinal polypeptide (VIP) release during its action. METHODS: A group of rats initially received oral misoprostol treatment of 1, 10, 50 and 100 microg/kg, respectively. By using orally fed charcoal as a motility marker, the SIT was assessed at 30 min following oral misoprostol treatment. Another group of rats received misoprostol as an intraperitoneal injection in similar doses to the group above. The small intestinal transit was computed for this group at 30 min following misoprostol injection via an instilled radiochromium motility marker that went through a previously placed intraduodenal catheter. The plasma VIP level was measured by using a radioimmunoassay kit. RESULTS: Neither charcoal evaluated transit nor the plasma VIP level was influenced by the lower doses of oral misoprostol treatment (1 and 10 microg/kg), whereas other doses enhanced SIT and diminished the plasma VIP level (P< 0.01).The similar effects on radiochromium computed SIT (P< 0.01) and plasma VIP levels were obtained in tubed rats following misoprostol intraperitoneal treatment. The SIT results correlated negatively with plasma VIP levels. CONCLUSIONS: Enhanced SIT and diminished VIP levels are found in rats following misoprostol treatment. It appears that inhibited VIP release is one of the mechanisms in misoprostol-enhanced SIT. PMID- 11106091 TI - Endogenous endothelin in a rat model of acute colonic mucosal injury. AB - BACKGROUND: Endothelin (ET) is involved in various biologic activities in non vascular and vascular tissues. While ET has some significant effects on gastrointestinal functions, the possible role of endogenous ET in the host response to mucosal injury has not been well clarified. METHODS: The present study describes an investigation of the effects of an endothelin A receptor antagonist, BQ-123, on lactate dehydrogenase (LDH), mucus and albumin flux into the perfusate in a rat model of acute colonic injury, induced by acetic acid perfusion. The present study also examined localization of ET in damaged rat colons by using immunohistochemistry. RESULTS: A 4% acetic acid treatment induced mild mucosal damage of perfused rat colon and increased LDH as well as albumin and protein-bound hexose release into the perfusate. Pretreatment with BQ-123 significantly reduced LDH activity and protein-bound hexose concentration in the perfusate and delayed the reduction of albumin leakage from damaged mucosa. Vascular endothelial, neural and surface epithelial cells of the colon showed strong ET-like immunoreactivity. Mucosal damage markedly influenced ET expression by epithelial cells. Mild mucosal damage decreased the ET expression by surface epithelial cells while moderate mucosal damage induced a mosaic location of ET positive epithelial cells in the crypt. Severe mucosal damage abolished the ET expression by epithelial cells. CONCLUSIONS: Endothelin may play a role in the host response to acute mucosal damage. Mucosal ET production is significantly affected by mucosal injury. PMID- 11106092 TI - Divergence of mucosal and motor effects of insulin-like growth factor (IGF)-I and LR3IGF-I on rat isolated ileum following abdominal irradiation. AB - BACKGROUND AND AIMS: In addition to its beneficial effects on small intestinal mucosal development and repair, insulin-like growth factor (IGF)-I has also been reported to improve neural function in toxic neuropathies. It has recently been recognized that enteric neural abnormalities contribute to the small intestinal dysmotility observed during and after abdominal radiotherapy for gynecological and pelvic malignancy. The aim of the present study was to evaluate the effects of IGF-I (5 mg/kg per day) and the more potent analog LR3IGF-I (5 mg/kg per day) on neurally mediated ileal dysmotility following irradiation. METHODS: Intestinal motor activity was recorded from 6-8 cm segments of explanted rat ileum using a miniaturized manometric technique during arterial perfusion with oxygenated fluorocarbon solution. Studies were performed 4 days after treatment with 10 Gy abdominal irradiation. At the time of irradiation, all rats underwent implantation of an osmotic mini-pump that contained 100 mmol/L acetic acid vehicle (n = 8), IGF-I (n = 8) or LR3IGF-I (n = 7). For each experiment, the total number of pressure waves, high-amplitude long-duration (defined as > 20 mmHg, > 6 s; HALD) pressure waves and long bursts (> 20) of pressure waves were determined. Ileal segments from 12 non-irradiated rats were used as controls for manometric studies. In radiotherapy treated animals, the degree of mucosal damage was determined using a standardized histologic scoring system. RESULTS: The HALD pressure waves were infrequent in non-irradiated rats but occurred in all irradiated animals. Insulin-like growth factor-I and LR3IGF-I had no effect on the frequency, amplitude or migration characteristics of HALD pressure waves compared with vehicle. Histologic damage was reduced in animals that received IGF I and LR3IGF-I compared with vehicle-treated animals. CONCLUSIONS: In radiation enteritis, IGF-I has no effect on neurally mediated small intestinal dysmotility while improving mucosal histology. The disparity between these results suggests that parallel but separate pathologic processes underlie mucosal and motor abnormalities in radiation enteritis. PMID- 11106093 TI - Activation of mucosal phospholipase D in a rat model of colitis. AB - BACKGROUND AND AIMS: Phospholipase D (PLD) hydrolyzes phosphatidylcholine and produces lipid second messengers. Although cellular PLD has recently been recognized as an important signal-transmitting enzyme, the role of PLD in pathophysiologic conditions is largely unknown. In particular, the regulation of PLD in intestinal inflammation has not been previously investigated. The aim of the present study was to elucidate the role of PLD in experimental colitis. METHODS: Rats were intracolonically administered acetic acid and assessed for mucosal damage, mucosal PLD activity, mucosal myeloperoxidase activity, mucosal chemiluminescence and luminal concentration of leukotriene B4. Acetic acid treatment induced acute mucosal injury that was maximal at 24 h after treatment. RESULTS: Mucosal PLD activity was significantly elevated and correlated with mucosal damage. Chemiluminescence in colitic mucosa was inhibited by the addition of ethanol which suppresses the formation of phosphatidic acid catalyzed by PLD. CONCLUSION: These results suggest that PLD is activated in experimental colitis in rats and that PLD may play a role in mucosal damage induced by reactive oxygen metabolites. PMID- 11106094 TI - Detection of Listeria monocytogenes by polymerase chain reaction in intestinal mucosal biopsies from patients with inflammatory bowel disease and controls. AB - BACKGROUND AND AIMS: Components of the intestinal microflora are believed to play an important role in the pathogenesis of inflammatory bowel disease (IBD) in genetically susceptible hosts acting either as a non-specific antigenic stimulus or as a specific pathogen. Listeria monocytogenes has been suggested as an organism with the potential to cause IBD. The objective of the present study was to investigate the prevalence of L. monocytogenes DNA in intestinal biopsies from patients with IBD and from non-IBD controls by using nested polymerase chain reaction (PCR). METHODS: The DNA was extracted from 274 colonoscopic biopsies, which were obtained from 23 patients with Crohn's disease (CD), 28 with ulcerative colitis (UC) and 39 non-IBD control patients. Nested PCR amplification was used to detect the presence of the L. monocytogenes listeriolysin O (hly) gene. The sequences of positive PCR products were determined and compared with databases. RESULTS: The sensitivity of our nested PCR was 10 fg L. monocytogenes DNA. Overall, L. monocytogenes DNA was detected in 13.0% patients with CD, 17.9% patients with UC and 25.6% non-IBD control patients or in 29 of 274 (10.6%) endoscopic biopsies. Among them, L. monocytogenes DNA was detected in four of 67 (6%) biopsies from patients with CD, five of 94 (5.3%) biopsies from patients with UC and 20 of 113 biopsies (17.7%) from non-IBD control patients. Sequence analysis of positive PCR products demonstrated more than 95% similarity to the hly gene sequence of L. monocytogenes, confirming the authenticity of our PCR products. CONCLUSION: Listeria monocytogenes DNA was detected in the intestine of both patients with IBD and in non-IBD control patients, probably reflecting the widespread presence of this organism in the environment. The low yield of positive biopsies in our IBD patients (5-6%) and the fact that the detection rate of L. monocytogenes DNA was similar in endoscopic biopsies from IBD patients and non-IBD controls does not support a direct role for L. monocytogenes in the pathogenesis of IBD, at least in New Zealand patients. PMID- 11106095 TI - Analysis of K-ras codon 12 mutations and p53 overexpression in colorectal nodule aggregating tumors. AB - BACKGROUND AND AIMS: Morphologically, colorectal nodule-aggregating tumors are quite different from polypoid-type colorectal tumors that develop via the adenoma carcinoma sequence. Although polypoid-type colorectal tumors are well known to have a high incidence of K-ras gene mutation and p53 overexpression, colorectal nodule-aggregating tumors have not been examined in terms of genetic changes and clinicopathological features. In the present study, therefore, we analysed the clinicopathological features, genetic changes in K-ras codon 12, and p53 overexpression in colorectal nodule-aggregating tumors. METHODS: A total of 18 colorectal nodule-aggregating tumors were surgically resected and then analysed clinicopathologically. Immunohistochemistry and polymerase chain reaction-single stranded conformational polymorphism were performed to analyse p53 abnormalities in the tumors. K-ras codon 12 mutations were screened out by the polymerase chain reaction-restriction fragment length polymorphism method and analysed by fluorescence direct sequencing. RESULTS: p53 overexpression was observed in six lesions (33%). p53-overexpressing cells were observed in parts of carcinoma or adenoma showing high-grade atypia. Four of the 10 (40%) samples had a p53 gene mutation. Nine of the 18 (50%) samples had a K-ras codon 12 point mutation. In eight cases (89%), the mutations of the K-ras codon 12 were of the same type: GGT (glycine) to GTT (valine). CONCLUSIONS: The colorectal nodule-aggregating tumor has distinctive characteristics showing a morphological phenotype of the superficial-type tumors and genotype of the polypoid tumors in terms of K-ras gene mutation and p53 overexpression. PMID- 11106096 TI - Folate deficiency diminishes the occurrence of aberrant crypt foci in the rat colon but does not alter global DNA methylation status. AB - BACKGROUND AND AIMS: It has been suggested that a diminished folate status may enhance colorectal carcinogenesis by causing DNA hypomethylation. The aims of the present study were to assess the impact of different levels of folate depletion on azoxymethane (AOM)-induced aberrant crypt foci (ACF) formation and DNA hypomethylation in the colon of male Sprague-Dawley rats. METHODS: Rats, aged 4 weeks, were divided into four groups and were fed semipurified diets either containing adequate folate (control), devoid of folate (FD) or FD + 1% succinylsulfathiazole before AOM treatment (FD1) or during the last 4 weeks of the study (FD2). At 8 weeks of age, all animals received subcutaneous injections of AOM once weekly for 3 weeks at a dose rate of 15 mg/kg bodyweight. Animals were necropsied 6 weeks after the last AOM injection and the ACF were visualized under light microscopy in formalin-fixed, methylene blue-stained colons. RESULTS: Blood folate concentrations were significantly depleted (P < 0.001) in the treatment groups consuming folate deplete diets, with the FD2 treatment group having significantly lower folate levels compared with all other groups. Higher plasma homocysteine concentrations (P < 0.001) were observed in the groups that exhibited diminished blood folate levels. There were no significant differences in global DNA methylation in the liver or colonic mucosa between the four groups, despite some groups exhibiting marked folate depletion. Animals with the most severe folate deficiency (FD2) had a lower final bodyweight and had significantly fewer ACF in their colon (P < 0.05) compared with control animals. Total (mean +/ SEM) ACF counts were as follows: control 286+/-24; FD 290+/-25; FD1 218+/-32; and FD2 205+/-27. CONCLUSIONS: In this model, folate deficiency diminished the occurrence of ACF but did not alter global DNA methylation status in the colon. PMID- 11106097 TI - A multicenter, randomized, controlled trial of interferon alfacon-1 compared with alpha-2a-interferon in Chinese patients with chronic hepatitis C virus infection. AB - BACKGROUND: Alpha-interferons are the accepted therapy for patients infected with chronic hepatitis C virus (HCV) in China. However, consensus interferon (CIFN) for HCV treatment is effective in patients with chronic hepatitis C from Western countries. METHODS: This randomized, controlled trial was conducted to determine the safety and efficacy of CIFN at two doses, and to compare it with alpha-2a interferon (IFN-alpha-2a) in Chinese patients with chronic HCV. Interferon-naive patients with chronic HCV infection (n = 187) were randomly chosen to receive 15 microg CIFN or 9 microg or 3 MU IFN-alpha-2a subcutaneously, three times a week for 24 weeks, followed by a 24 week observation period. Efficacy was evaluated by the normalization of serum alanine aminotransferase (ALT) and the non detectability disappearance of serum HCV-RNA by using reverse-transcription polymerase chain reaction. The safety of CIFN was evaluated by recording the type and severity of adverse effects. RESULTS: The combined ALT and HCV-RNA end-of treatment and sustained responses were observed to be greater for treatment with 15 microg CIFN (59.0% and 55.7%, respectively) compared to IFN alpha-2a (36.1% and 39.3%, respectively; P = 0.01 for the end-of-treatment, P = 0.07 for the sustained response). The combined ALT and HCV-RNA end-of-treatment and sustained responses for treatment with 9 microg CIFN (both 49.2%) were higher than those for IFN-alpha-2a (not statistically significant). Data were analyzed by using a logistic-multiple-variate regression model, which indicated that the higher IFN dose (15 microg or 9 microg CIFN vs 3 MU IFN-alpha-2a; P < 0.01) appeared to be associated with a better sustained response. The type, frequency and severity of adverse effects were comparable across treatment groups. CONCLUSIONS: Consensus interferon appears to be safe and effective at concentrations of 9 and 15 microg, but 15 microg CIFN may be more effective than 3 MU IFN-alpha-2a, without increased toxicity. PMID- 11106098 TI - Seroprevalence of GB virus C/hepatitis G virus-RNA and anti-envelope antibody in high-risk populations in Taiwan. AB - BACKGROUND: GB Virus C (GBV-C)/hepatitis G virus (HGV) was identified in 1995 1996 as a transfusion-transmissible virus. The diagnosis of GBV-C/HGV infection is based on the detection of GBV-C/HGV-RNA by using polymerase chain reaction. Recently, an enzyme immunoassay detecting the antibodies to the viral protein, E2 envelope protein (anti-envelope) of GBV-C/HGV, has been developed. METHODS: Serum GBV-C/HGV-RNA and anti-envelope antibody were determined in 76 cases of intravenous drug users (IVDU), 76 patients with regular hemodialysis and in 80 prostitutes to evaluate the GBV-C/HGV infection rate among high-risk populations in Taiwan. Seventy-six healthy blood donors were randomly selected and were used as a control group. RESULTS: The prevalence of GBV-C/HGV-RNA in high-risk populations was 33% for IVDU, 16% for patients with hemodialysis and 13% for prostitutes, which was significantly higher than the 3% obtained in the control group (P < 0.05 for all groups). The prevalence of anti-envelope antibody was 13% for IVDU, 21% for patients with hemodialysis and 23% for prostitutes, which was not significantly different from the control group (11%). Among the 99 subjects who had positive GBV-C/HGV markers, 97 were tested for exclusive positivity for either GBV-C/HGV-RNA or anti-envelope antibody. CONCLUSIONS: The presence of serum anti-envelope antibody usually indicates the clearance of serum GBV-C/HGV RNA in patients infected with GBV-C/HGV. GBVirus-C/HGV infection in high-risk populations, determined by the presence of serum GBV-C/HGV-RNA, may underestimate the true level of past and present infection. PMID- 11106099 TI - Frequency and significance of antibodies to chromatin in autoimmune hepatitis type I. AB - AIMS AND METHODS: To assess the frequency and clinical significance of antibodies to chromatin (ACA) in autoimmune hepatitis (AIH), 36 Japanese patients with AIH type I were studied for serum reactivity with chromatin by using an ELISA. RESULTS: Antibodies to chromatin were detected in 19 of 36 patients with AIH type I. There was a significantly higher frequency of ACA in patients with AIH type I than in patients with primary biliary cirrhosis, chronic hepatitis C and B (52.8 vs 13.2, 5.4 and 6.7%, respectively; P<0.01). None of the 19 healthy subjects had positive reactions. Sixteen of 19 patients with seropositive sera (44.4%) had reactivities with other nuclear antigens (recombinant nucleoproteins U1RNP-A, U1RNP-70; recombinant ribonucleoprotein complexes SSA/Ro 52K, SSA/Ro 60K; recombinant centromere Cenp-B; dsDNA and histones). Adsorption with double stranded DNA (dsDNA) and histones could not remove the majority of antichromatin reactivity as 81.9% of the antibody reactivity still remained. In five sera samples from AIH type I patients positive for anti-dsDNA and antihistones, the antibody activities for dsDNA and histones were inhibited after the absorbtion of sera with chromatin. The patterns of antinuclear antibodies (ANA) detected by using indirect immunofluorescence were similar between patients with and without ACA. Patients with ACA had significantly high serum levels of gamma-globulin and immunoglobulin G. The ACA titers dropped significantly after corticosteroid treatment (P< 0.01). CONCLUSIONS: Antibodies to chromatin are frequently present in patients with AIH type I and they are one of the dominant autoantibodies associated with ANA reactivity in AIH type I. Antibodies to chromatin cannot be used to characterize distinct clinical subgroups of AIH type I. PMID- 11106100 TI - Troglitazone prevents fatty changes of the liver in obese diabetic rats. AB - BACKGROUND AND AIMS: Troglitazone is a newly developed antidiabetic drug and is indicated to be useful for the treatment of patients with type II diabetes mellitus. Recently, however, it became clear that troglitazone could cause liver dysfunction in some patients. In addition, a relationship between the activation of the peroxisome proliferator-activated receptor gamma receptor by troglitazone and colon tumorigenesis has been suggested. The present study was undertaken to examine the effects of long-term administration of troglitazone on the liver and intestine in genetically obese and diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) and control Long-Evans Tokushima Otsuka (LETO) rats. METHODS: A troglitazone-rich diet (200 mg/100 g normal chow) or a standard rat chow, free of troglitazone (control), was given to OLETF and LETO rats from 12 or 28 weeks of age until 72 weeks of age. Serum levels of glucose, insulin, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were determined at several time points. In addition, histology of the liver and intestine and serum levels of cholesterol and triglycerides were examined at 72 weeks of age. RESULTS: Troglitazone prevented age-related increases in fasting glucose and insulin concentrations in OLETF rats, but had no significant influences on serum levels of AST and ALT in both strains of rats. The liver weights in the control OLETF rats were significantly heavier than in the LETO rats. Troglitazone significantly reduced serum cholesterol and triglyceride levels and the liver weight. However, it had no influence on the large intestine weight and the number of colonic polyps in both OLETF and LETO rats. Sections of the liver from the untreated OLETF rats showed mild fatty changes in the central zone of the hepatic lobule, whereas those from the troglitazone-treated OLETF rats appeared normal with no fat deposition in the hepatocytes. Troglitazone in LETO rats also caused no significant histopathologic changes of the liver tissue. CONCLUSION: Our present study demonstrated that long-term administration of troglitazone prevents the progress of the metabolic derangement and fatty changes of the liver in genetically determined obese diabetes. PMID- 11106101 TI - Clinicopathologic analysis of stage II-III hepatocellular carcinoma showing early massive recurrence after liver resection. AB - BACKGROUND AND AIMS: Prognosis after hepatectomy for hepatocellular carcinoma (HCC) has been improved by progress in the evaluation of hepatic functional reserve, surgical techniques and perioperative management. However, even when curative resection is performed at a relatively early stage, a considerable number of patients develop early intrahepatic and/or extrahepatic recurrence postoperatively. This study analyzed the clinicopathologic features of HCC with early recurrence. METHODS: We reviewed records of 513 consecutive patients who had undergone liver resection for HCC. There were 48 deaths within a year after surgery from recurrence, including 21 patients with stage II or III HCC (group I). Clinicopathologic parameters of group I patients were compared with those of 188 patients (group II) who developed recurrence following resection of stage II or III HCC and died more than 1 year after surgery. RESULTS: On univariate analysis, age, tumor diameter (phi), alpha-fetoprotein (AFP):phi and protein induced by vitamin K absence or antagonist II (PIVKA-II):phi were significantly greater in group I than in group II. Macroscopic portal vein invasion, microscopic vascular invasion, intrahepatic metastasis, poor differentiation, pleomorphism, sarcomatous change, vascular lake, and angiographic condensed pooling were more frequently observed in group I than group II. Five independent determinants were selected by multivariate analysis: AFP:phi, histologic pleomorphism, sarcomatous change, vascular lake and angiographic condensed pooling. CONCLUSIONS: Highly malignant HCC with extremely poor prognosis exhibits peculiar clinicopathologic characteristics, particularly histologic immaturity, and can be predicted by preoperative indicators such as markedly elevated tumor marker concentrations and condensed pooling on angiography. PMID- 11106102 TI - Prevention of proteolysis in cold-stored rat liver by addition of amino acids to the preservation solution. AB - BACKGROUND: One process identified as detrimental in liver preservation is proteolysis. METHODS: We tested the effects of adding antiproteolytic amino acids (L-alanine, L-glutamine, L-histidine, L-leucine, L-methionine, L-phenylalanine, L proline, L-tryptophan) to the preservation medium, in a model of reperfusion of 24 h cold-stored rat livers. RESULTS: During the preservation period, antiproteolytic amino acids inhibited the proteolysis observed in stored livers as shown by branched-chain amino acid fluxes, which switched from release to uptake. During reperfusion, cold storage of lives without the addition of antiproteolytic amino acids resulted in a decrease in the total amino acid and branched-chain amino acid uptake and a lower perfusion flow rate. The addition of antiproteolytic amino acids during liver storage resulted in the maintenance of total amino acid and branched-chain amino acid uptake and a significant improvement in the perfusion flow rate during reperfusion. CONCLUSIONS: The presence of antiproteolytic amino acids in the preservation medium might be of interest in improving hepatic graft viability in transplantation. PMID- 11106103 TI - Clinical features and etiology of hepatocellular carcinoma arising in patients with membranous obstruction of the inferior vena cava: in reference to hepatitis viral infection. AB - BACKGROUND AND AIMS: Budd-Chiari syndrome (BCS) comprises hepatic vein thrombosis and inferior vena cava (IVC) obstruction known as membranous obstruction of the IVC (MOVC). The latter is frequently complicated by hepatocellular carcinoma (HCC). The etiology of MOVC-associated HCC in relation to hepatitis viral infection is not known. In this study, we investigated the clinical features and etiology of HCC in MOVC. METHODS: Membranous obstruction of IVC and HCC were diagnosed and studied by using imaging techniques. Sera from patients with MOVC, complicated by HCC, were examined for hepatitis viral antigens and antibodies (hepatitis B surface antigen (HBsAg), antibody to HBsAg (anti-HBs), antibody to hepatitis B core antigen (anti-HBc) and third generation antibody to hepatitis C virus (anti-HCV)) and for hepatitis viral nucleic acids (hepatitis B virus (HBV) DNA, hepatitis C virus (HCV)-RNA, hepatitis G virus (HGV)-RNA and TT virus DNA). RESULTS: We studied 12 patients with BCS who were seen between April 1968 and February 1999. All of them had MOVC. Hepatocellular carcinoma developed in three (25%) of them. There were no obvious differences in the clinical features and imaging findings concerning MOVC between patients with and without HCC. Hepatocellular carcinoma in these three patients showed no clear trend in clinical features and imaging findings. Of the hepatitis viral markers examined, HBsAg, anti-HBc and HBV-DNA were positive in only one of three patients with HCC and all of the viral markers were negative in the other two patients. CONCLUSIONS: Chronic congestion in the liver, caused by an outflow block of hepatic veins and subsequent histopathologic change, must have led to HCC in two patients without any hepatitis viral markers. Patients with MOVC should be followed closely as a high-risk group for HCC. PMID- 11106104 TI - Detection of intracellular interleukin-2 production in peripheral T lymphocytes by flow cytometry in patients with pancreatobiliary malignancies. AB - BACKGROUND AND AIMS: To date, it has been reported that cellular immunity is decreased in patients with cancer and investigations into cytokine production has been insufficient. Therefore, we examined intracellular cytokine production by using flow cytometry in patients with cancer and discussed the reasons for the impairment of their immune system. METHODS: Eleven patients with hepatobiliary malignancies (68.5+/-11.8 years of age), eight age-matched controls (70.0+/-12.0 years of age) and 10 young volunteers (31.9+/-3.1 years of age) were used in the present study. Stimulated peripheral blood mononuclear cells from these patients were stained with fluorescence-labeled anticytokine monoclonal antibodies and analyzed with a Fluorescence activated cell sorter (FAC)Scan. RESULTS: The percentage of positively stained T cells was calculated and compared with controls. Repeated measured ANOVA was used for statistical analysis. Interleukin (IL)-2 production was significantly decreased in patients with cancer compared to controls (P=0.0122), and it may suggest decreased cellular immune activity of the patients. Simultaneously, spontaneous intracellular IL-4 production was observed in patients and age-matched controls, but levels were significantly increased when compared with the young volunteers (P=0.0052, P=0.031, respectively). CONCLUSIONS: It was of interest that spontaneous intracellular IL-4 production was detected in elderly subjects. PMID- 11106105 TI - Images of interest. Hepatobiliary and pancreatic: abdominal skin injury following arterial chemoembolization for hepatocellular carcinoma. PMID- 11106106 TI - Images of interest. Gastrointestinal: complications of fundoplication. PMID- 11106107 TI - Clinical relapse in Whipple's disease despite maintenance therapy. AB - Whipple's disease is a multisystem disorder that was first reported just over 100 years ago. Only recently, the bacillus responsible for the condition was identified and subsequently cultured. However, differences of opinion remain regarding the best antibiotic regimen and duration of therapy at primary diagnosis and there is also great uncertainty about the management of disease relapse. We report a case of clinical relapse of Whipple's disease in a man who was on a prolonged therapy with trimethoprim-sulfamethoxazole. We describe his management and review the literature on the treatment of this condition, with particular reference to the recurrence of the disease. PMID- 11106108 TI - Mitoxantrone-related acute myelocytic leukemias. PMID- 11106109 TI - Molecular aspects of multiple myeloma. AB - Multiple myeloma (MM) is a B-cell neoplasm characterized by bone marrow infiltration with malignant plasma cells, which synthesize and secrete monoclonal immunoglobulin (Ig) fragments. Despite the considerable progress in the understanding of MM biology, the molecular basis of the disease remains elusive. The initial transformation is thought to occur in a postgerminal center B-lineage cell, carrying a somatically hypermutated Ig heavy chain (IGH) gene. This plasmablastic precursor cell colonizes the bone marrow, propagates clonally and differentiates into a slowly proliferating myeloma cell population, all under the influence of specific cell adhesion molecules and cytokines. Production of interleukin-6 by stromal cells, osteoblasts and, in some cases, neoplastic cells is an essential element of myeloma cell growth, with the cytokine stimulus being delivered intracellularly via the Jack-STAT and ras signaling pathways. While karyotypic changes have been identified in up to 50% of MM patients, recent molecular cytogenetic techniques have revealed chromosomal abnormalities in the vast majority of examined cases. Translocations mostly involve illegal switch rearrangements of the IGH locus with various partner genes (CCND1, FGFR3, c-maf). Such events have been assigned a critical role in MM development. Mutations in coding and regulatory regions, as well as aberrant expression patterns of several oncogenes (c-myc, ras) and tumor suppressor genes (p16, p15) have been reported. Key regulators of programmed cell death (BCL-2, Fas), tumor expansion (metalloproteinases) and drug responsiveness (topoisomerase II alpha) have also been implicated in the pathogenesis of this hematologic malignancy. A tumorigenic role for human herpesvirus 8 (HHV8) was postulated recently, following the detection of viral sequences in bone marrow dendritic cells of MM patients. However, since several research groups were unable to confirm this observation, the role of HHV8 remains unclear. Translation of the advances in MM molecular biology into novel therapeutic strategies is essential in order to improve disease prognosis. PMID- 11106110 TI - Locoregional immunotherapy in cancer patients: review of clinical studies. AB - Many patients with invasive cancer have a compromised immune system. This immune dysfunction does appear to start at the site of the tumor. Locoregional immunotherapy is given to stimulate the immune system in order to kill tumor cells either indirectly via a specific or a non-specific way or directly via cell transfer therapy. Advantages to give this immunotherapy locoregionally in stead of systemically are a higher concentration of the immunomodulator at the site of the tumor, to attract or activate effector cells, and diminished toxicity. In this review we have summarised the clinical studies using loco-regional immunotherapy in patients with cancer. Only phase I and II studies were performed. Clinical responses were seen. No single locoregional treatment has become a standard therapy. Relatively few investigations were performed to estimate the influence on the locally effector mechanisms or immune dysfunction. In future clinical trials it is essential to get a better insight in these mechanisms. PMID- 11106111 TI - Dose-escalation study of CHOP with or without prophylactic G-CSF in aggressive non-Hodgkin's lymphoma. AB - BACKGROUND: CHOP is accepted as the gold standard for first line chemotherapy of aggressive non-Hodgkin's lymphoma (NHL). A dose-escalation study of CHOP was conducted to determine the maximal tolerated dose (MTD) and toxicity profile of CHOP at three-week intervals with or without prophylactic recombinant human granulocyte colony-stimulating factor (rHuG-CSF) in patients with aggressive NHL. PATIENTS AND METHODS: The doses of drugs were escalated from 50 mg/m2 to 70 mg/m2 for doxorubicin and from 750 mg/m2 to 2250 mg/m2 for cyclophosphamide, with conventional doses of vincristine and oral prednisolone. After the MTD was determined without rHuG-CSF, dose escalation was conducted with prophylactic rHuG CSF. RESULTS: Thirty-three patients with NHL were enrolled into the study. The MTD without prophylactic rHuG-CSF was 70 mg/m2 of doxorubicin and 1250 mg/m2 of cyclophosphamide, with neutropenia as a dose-limiting toxicity. The MTD with prophylactic rHuG-CSF was 70 mg/m2 of doxorubicin and 2250 mg/m2 of cyclophosphamide. The overall response rate was 100% (76% complete response and 24% partial response). Progression-free survival and overall survival at five years were 45% and 66%, respectively. CONCLUSIONS: Significant dose escalation of doxorubicin and cyclophosphamide was feasible with prophylactic rHuG-CSF. The efficacy of dose-escalated CHOP should be compared with that of standard CHOP. PMID- 11106112 TI - Front-line treatment of advanced breast cancer with docetaxel and epirubicin: a multicenter phase II study. AB - PURPOSE: In a previous phase I trial we evaluated the toxicity and determined the maximum tolerated doses of the docetaxel (D)-epirubicin (Epi) combination. We conducted a multicenter phase II study to evaluate the efficacy and tolerability of this regimen as front-line treatment in women with advanced breast cancer (ABC). PATIENTS AND METHODS: Fifty-four women with ABC stage IIIB (4 patients) or IV (50 patients) received front-line treatment with Epi 70 mg/m2 on day 1 and D 90 mg/m2 on day 2. The median age was 55 years, performance status (WHO) was 0-1 in 49 patients and visceral disease was present in 45 (83%). RESULTS: All patients were evaluable for toxicity and 50 for response. In an intent-to-treat analysis complete remission was observed in 5(9%) patients, partial remission in 31 (57%) (overall response rate 66%, 95% confidence interval: 54% 79%), stable disease in 9 (17%) and disease progression in 9 (17%). After a median follow-up of 11.5 months, the median duration of responses was 8 months, the median time to disease progression 11.5 months and the median survival has not yet been reached. The probability of one-year survival was 65%. Three hundred six cycles of treatment were administered (median 6 cycles per patient). Grade 3 and 4 neutropenia was observed in 8 (15%) and 31 (57%) patients, respectively, and febrile neutropenia in 19 (35%). Prophylactic rh-G-CSF was used in 45 (83%) patients or 226 (74%) cycles. Other hematologic or non-hematologic toxicities were usually mild. In five (9%) patients the left ventricular ejection fraction (LVEF) was decreased by more than 10% with the treatment. Two patients died during the treatment of respiratory failure without associated neutropenia. CONCLUSIONS: The combination of docetaxel epirubicin is an effective and well tolerated front-line treatment in patients with ABC. PMID- 11106113 TI - Breast cancer in women < or = 35 years: review of 1002 cases from a single institution. AB - BACKGROUND: Early-onset breast cancer may differ with respect to etiology, clinical features and outcome compared with breast cancer in older women. To gain further insight, we retrospectively reviewed the clinical features and outcome of women < or = 35 years with primary breast cancer seen at our institution over a 30-year period. PATIENTS AND METHODS: Charts were reviewed for women with operable breast cancer diagnosed < or = 35 years of age seen at the Princess Margaret Hospital (PMH), Toronto from 1965-1994. RESULTS: One thousand eighty-six women with non-metastatic invasive breast cancer, aged 18.3-35.6 years (median 32.1 years) were referred to PMH. Symptoms at presentation included: self detected breast lump (83%), other breast symptom (10%), physician diagnosis (4%) and unknown (3%). Tumor size was known in 936 (>2 cm in 61%) and nodal status in 888 (lymph node positive in 52%). Modified radical mastectomy was performed in 568 (57%) and breast-conservation surgery (BCS) in 422 (42%). Five hundred sixteen (51%) patients received adjuvant radiotherapy and five hundred thirty four (53%) adjuvant systemic therapy. Two hundred ninety-three (29%) patients had a family history of breast cancer (FH). Contralateral breast cancer (CBC) occurred more frequently in women with FH (P range 0.042-0.008). Local recurrence (LR) was 37% and 73% at 10 years in those treated by BCS with and without radiotherapy, respectively. At 10 years, disease-free survival (DFS) was 30% and overall patient survival 48%. CONCLUSIONS: In this cohort, breast cancer was usually self-diagnosed and tumors were > 2 cm at presentation in approximately two-thirds of cases, suggesting the possibilities of a delay in diagnosis, more aggressive tumors or both. Our results are compatible with the known association of breast cancer FH with increased CBC. Our data also corroborates the suggestion that positive genetic testing in this age group should lead to consideration of more aggressive ipsilateral and contralateral breast management. In those receiving adjuvant irradiation after BCS, the LR rate was high, but did not impact on overall survival. PMID- 11106115 TI - A phase II trial of gemcitabine in combination with 5-fluorouracil (24-hour) and folinic acid in patients with chemonaive advanced pancreatic cancer. AB - BACKGROUND: Gemcitabine (Gemzar) and 5-fluorouracil (5-FU) plus folinic acid (FA) both have proven activity in the treatment of patients with advanced pancreatic cancer. The present study was initiated to investigate the efficacy of gemcitabine in combination with 5-FU-FA. PATIENTS AND METHODS: Thirty-eight patients, median age 60 years (range 34-70) with inoperable, stage IV, pancreatic cancer were enrolled into the study and treated on an outpatient basis. All except one patient received at least one cycle of treatment with gemcitabine (1000 mg/m2), followed by FA (200 mg/m2) and 5-FU (750 mg/m2) administered as a 24-hour continuous infusion on days 1, 8, 15 and 22 of a 42-day schedule. No patient had received prior chemotherapy or radiotherapy. All 38 patients were assessed for efficacy, toxicity and time to progressive disease. RESULTS: Two patients (5%), achieved a partial response and thirty-four patients (89%) achieved stable disease. There were two early deaths (< or = 4 weeks). The median time to progression was 7.1 months (range 0.4-18.1+; 95% confidence interval (95% CI): 5.3-7.9 months). Three patients had a progression-free interval of greater than 12 months and 12 of 38 patients (32%) survived longer than 12 months. The median overall survival was 9.3 months (range 0.5-26.5; 95% CI: 7.3-13.0 months). The incidence of grade 3 and 4 toxicities was low. CONCLUSIONS: The combination of gemcitabine and 5-FU-FA is active and well tolerated and seems to offer an improvement in progression-free interval over both gemcitabine monotherapy and 5 FU-FA therapy. PMID- 11106114 TI - A phase II study of weekly docetaxel as salvage chemotherapy for advanced gastric cancer. AB - BACKGROUND: Docetaxel has shown some activity in advanced gastric cancer. Recent phase I studies found low hematologic toxicity and a favourable toxicity profile when docetaxel was administered on a weekly schedule. In this study, we explored the activity of weekly docetaxel in patients with advanced gastric cancer who failed first-line chemotherapy. MATERIALS AND METHODS: Patients with stable or progressing disease after first-line chemotherapy received 36 mg/m2 weekly docetaxel. One cycle consisted of six administrations followed by a two-weeks rest, patients were re-evaluated at week eight. The optimal two-stage design was adopted for early stopping of the trial if responses were one or less in 21 patients (< 20% response rate with alpha and beta error probabilities 0.05 and 0.010 respectively). RESULTS: Twenty-one patients have been enrolled and they are fully evaluable for response and toxicity. One patient achieved partial response, 8 patients had stable disease and 12 patients progressed. Median overall survival from the onset of salvage chemotherapy was 3.5 months. Hematologic toxicity was observed in two patients who experienced grade III leukopenia. Beginning from the third week of treatment, most of the patients (90%) showed grade II asthenia which resulted the commonest side-effect. CONCLUSIONS: This schedule of weekly docetaxel did not show significant activity in pretreated patients with advanced gastric cancer. PMID- 11106116 TI - 2-Fluorine-18-fluoro-2-deoxy-D glucose positron emission tomography in the pretreatment staging of Hodgkin's disease: influence on patient management in a single institution. AB - PURPOSE: Optimum therapy for patients with Hodgkin's disease (HD) is determined by a number of prognostic factors, one of which is an accurate definition of extent of disease (stage). Computerised tomography is widely used in staging but cannot reliably evaluate normal sized lymph nodes and some extranodal sites, e.g., liver, spleen and bone marrow. 2-Fluorine-18-fluoro-2-deoxy-D glucose (FDG) has been shown to concentrate preferentially in lymphoma sites (whether in nodal or extranodal tissue) and therefore may have a useful role in staging patients with HD. This study compares concurrent computerized tomography (CT) and FDG positron emission tomography (PET) in the staging of Hodgkin's disease and assesses the frequency of stage migration and possible changes in therapy related to the use of PET scanning. PATIENTS AND METHODS: This was a single centre retrospective study of 44 patients with Hodgkin's disease who underwent both staging CT and PET prior to treatment between September 1993 and August 1998 at St. Thomas' Hospital. The number and sites of disease were assessed for each patient, documenting any stage and therapy modification prompted by PET findings. RESULTS: One hundred fifty-nine sites of disease were demonstrated in forty-four patients by FDG-PET compared with eighty-four by CT. As a result, 18 (40.9%) patients were upstaged, nine of these by FDG-uptake in splenic or extranodal sites not visualised on CT. Only three patients were downstaged by PET results. Eleven patients (25%) had treatment modified by PET scan findings. CONCLUSIONS: Significantly more sites of disease were identified by PET than CT resulting in stage changes and a modification of therapy in 25% of patients. This has important implications not only for current patient management but also for the design of future clinical trials. PMID- 11106117 TI - Streptozocin and o,p'DDD in the treatment of adrenocortical cancer patients: long term survival in its adjuvant use. AB - BACKGROUND: To evaluate the efficacy of streptozocin and o.p'DDD (SO) in adrenocortical cancer (ACC) patients since other chemotherapeutic regimens have limited effects. PATIENTS AND METHODS: We performed a phase II study with SO therapy in 40 ACC patients (median age 44 years). Oral o,p'DDD administration (1 4 g/d, every day) was given together with intravenous streptozocin (1 g/d for five days, thereafter 2 g once every three weeks). 5HT3-receptor blocker was used as standard premedication for streptozocin. RESULTS: The SO therapy was found to have significant effects on disease-free interval (P = 0.02) as well as on survival (P = 0.01) in adjuvantly treated cases (n = 17) in comparison to the patients who did not get any therapy after complete resection (n = 11). Complete or partial response was obtained in 36.4% of patients with measurable disease (n = 22). The overall two-year and five-year survival rates were 70% and 32.5%, respectively. The presence of metastases at diagnosis was identified as a poor prognostic factor (P = 0.02). CONCLUSIONS: The present study necessitates further randomized clinical study of SO therapy in the treatment of ACC, mainly as adjuvant treatment immediately after curative intended surgery, and could be developed into a regular treatment regimen. PMID- 11106118 TI - Early secondary acute myelogenous leukemia in breast cancer patients after treatment with mitoxantrone, cyclophosphamide, fluorouracil and radiation therapy. AB - BACKGROUND: The topoisomerase II-targeted drugs, epipodophyllotoxins and anthracyclines, have been shown to induce therapy-related AML (t-AML) characterized by a short latency period after chemotherapy, the absence of prior myelodysplastic syndrome and stereotyped chromosome aberrations. Few reports have been published on patients treated with the anthracenedione mitoxantrone which also targets topoisomerase II. We observed 10 cases of such t-AML over a 7-year period in breast cancer patients treated with mitoxantrone combined with fluorouracil, cyclophosphamide and regional radiotherapy, and in three cases with vindesine. PATIENTS AND METHODS: We retrospectively analyzed patients referred to our hospital for AML with a past history of polychemotherapy for breast cancer, including mitoxantrone, either as adjuvant (8 patients)/neoadjuvant (1 patient) therapy or for metastatic disease (1 patient). We studied the probability of developing t-AML in a prospective series of 350 patients treated with an adjuvant FNC regimen (mitoxantrone, fluorouracil, cyclophosphamide) and radiation therapy. RESULTS: The median age was 45 years (range 35-67). t-AML developed 13-36 months (median 16) after beginning chemotherapy for breast cancer, and 4-28 months (median 10.5) after ending treatment. As described in t-AML following treatment with epipodophyllotoxins or anthracyclines, we found a majority of FAB M4, M5 and M3 phenotypes (7 of 10), and characteristic karyotype abnormalities that also can be found in de novo AML: breakpoint on chromosome 11q23 (3 patients), inv(16)(p13q22) (2 patients), t(15;17)(q22;q11) (1 patient), t(8;21)(q22;q22) (1 patient) and del(20q)(q11) (1 patient). The prognosis was poor. All patients died of AML shortly after diagnosis. Since two patients had been enrolled in a prospective trial for the treatment of breast cancer which included 350 patients, the probability of developing t-AML was calculated to be 0.7% from 25-40 months, using the Kaplan-Meier method (95%, confidence interval (95% CI): 0.1-4.5). CONCLUSIONS: The combination of mitoxantrone with cyclophosphamide, fluorouracil, and radiation therapy can induce t-AML, as with other topoisomerase II-targeted drugs. Despite a low incidence, the prognosis appears to be poor. PMID- 11106119 TI - A three-week schedule of gemcitabine-cisplatin in advanced non-small-cell lung cancer with two different cisplatin dose levels: a phase II randomized trial. AB - BACKGROUND: To explore a new schedule of gemcitabine-cisplatin (GP) combination therapy using two different cisplatin doses in patients with advanced non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: From May to December 1997, 92 chemonaive patients entered the study and 88 (28 with locally advanced and 60 with disseminated NSCLC) were evaluable for response and toxicity (45 in arm A and 43 in arm B). Patients were randomly assigned to arm A or arm B. Gemcitabine 1000 mg/m2 was given on days 1-8 plus cisplatin 100 mg/m2 in arm A and cisplatin 70 mg/m2 in arm B on day 2 of every 21-day cycle. RESULTS: The overall response rates in arms A and B were 42% (95% confidence interval (CI): 27.8%-56.7%) and 47% (95% CI: 31.6%-61.5%), respectively. Median duration of response was 9.7 months (range 1.8 to 30.9 months; 13.1 and 9.5 months for arm A and B, respectively), and median survival was 12 months (range 0.2 to 31.1 months; 15.4 and 11.5 months for arm A and B, respectively). Major WHO grade 3-4 toxicities in arm A vs. arm B included: thrombocytopenia (23% vs. 17% of courses), leukopenia (15%, vs. 4% of courses), anemia (7% vs. 6% of courses), and nausea-vomiting (20% vs. 7% of patients). Grade 1-2 nephrotoxicity occurred in 20% of patients in arm A and in 7% of patients in arm B, with one grade 4 episode in arm A. Six patients discontinued treatment because of toxicities, 5 in arm A and I in arm B. CONCLUSIONS: Results of this trial indicate that both schedules are feasible and active, with a milder toxicity in the arm with the lower cisplatin dose. PMID- 11106120 TI - Evaluation of drug interactions in the established FEC regimen in primary cultures of tumour cells from patients. AB - BACKGROUND: Chemotherapy using multi-drug regimens is considered more active than single-agent therapy. This may be due to synergistic interactions or, simply, a higher probability of administering an active agent. We investigated in vitro the type of drug interactions in a recognized regimen in relationship to tumour type and drug sensitivity. PATIENTS AND METHODS: The possibility of synergistic and additive interactions between individual cytotoxic drugs was investigated for the component drugs of the established FEC regimen, i.e., 5-fluorouracil, epirubicin and cyclophosphamide, in 243 patient tumour samples representing various drug sensitivity using the non-clonogenic fluorometric microculture cytotoxicity assay. RESULTS: Using a cell survival of < or = 50% as a limit for drug activity and sample sensitivity, the overall response rates to the most active single drug (Dmax) and the combination were 56% and 64%, respectively, with a distribution among diagnoses similar to that in the clinic. For 86% of the samples there was concordance with respect to judgement of activity using either Dmax or the combination. For samples being sensitive to at least one single drug, 95% were also sensitive to the combination whereas for samples with insignificant Dmax effect, only 2% were sensitive to the combination. In samples with modest Dmax effects, i.e., cell survival in the range > 50%- < or = 80%, 45% responded to the combination. The effect of the combination was generally well predicted from the Dmax effect. CONCLUSIONS: The superior antitumour effect of drug combinations compared with single drugs may be due to the higher chance of selecting an active agent. However, for intermediately sensitive tumours, additional interaction effects of a combination may be of clinical significance. PMID- 11106121 TI - A combination of a fixed dose rate infusion of gemcitabine associated to a bolus 5-fluorouracil in advanced pancreatic cancer, a report from the Italian Group for the Study of Digestive Tract Cancer (GISCAD). AB - BACKGROUND: Laboratory evidences suggest the possibility that an infusion rate of 10 mg/m2/min may be more effective than the standard 30-min infusion of Gemcitabine (GEM). PATIENTS AND METHODS: Thirty-four patients with histologically verified locally unresectable and/or metastatic pancreatic carcinoma received GEM at the dose of 1,500 mg/m2 with an infusion rate of 10 mg/m2/min, associated to 5 fluorouracil (5-FU) at the dose of 600 mg/m2. Both drugs were administered weekly for two consecutive weeks out of every three weeks. RESULTS: One complete and five partial responses have been observed for an overall response rate of 17% (95% CI: 3%-27%). The time to progression was 3.7 months with a median survival of 5.7 months. A clinical benefit was obtained in 5 of 29 patients (17%). Grade 3 4 WHO toxicities included neutropenia (35%) and thrombocytopenia (10%). CONCLUSION: It is unlikely that a fixed dose rate infusion of GEM, at least with this dose, can improve palliation in comparison with the standard 30-min infusion schedule in advanced pancreatic cancer. PMID- 11106122 TI - Pharmacokinetics and bioequivalence of a combined oral formulation of eniluracil, an inactivator of dihydropyrimidine dehydrogenase, and 5-fluorouracil in patients with advanced solid malignancies. AB - BACKGROUND: This study was performed to evaluate the pharmacokinetics, bioequivalence, and feasibility of a combined oral formulation of 5-flurouracil (5-FU) and eniluracil (Glaxo Wellcome Inc., Research Triangle Park, North Carolina), an inactivator of dihydropyrimidine dehydrogenase (DPD). The rationale for developing a combined eniluracil/5-FU formulation oral dosing form is to simplify treatment with these agents, which has been performed using separate dosing forms, and decrease the probability of severe toxicity and/or suboptimal therapeutic results caused by inadvertently high or conversely insufficient 5-FU dosing. PATIENTS AND METHODS: The trial was a randomized, three-way crossover bioequivalence study of three oral dosing forms of eniluracil/5-FU tablets in adults with solid malignancies. Each period consisted of two days of treatment and a five- to seven-day washout phase. Eniluracil at a dose of 20 mg, which results in maximal DPD inactivation, was administered twice daily on the first day and in the evening on the second day of each of the three treatments. On the morning of the second day, all patients received a total eniluracil dose of 20 mg orally and a total 5-FU dose of 2 mg orally as either separate tablets (treatment A) or combined eniluracil/5-FU tablets in two different strengths (2 tablets of eniluracil/5-FU at a strength (mg/mg) of 10/1 (treatment B) or 8 tablets at a strength of 2.5/0.25 (treatment C)). The pharmacokinetics of plasma 5-FU, eniluracil, and uracil, and the urinary excretion of eniluracil, 5-FU, uracil, and alpha-fluoro-beta-alanine (FBAL), were studied. To determine the bioequivalence of the combined eniluracil/5-FU dosing forms compared to the separate tablets, an analysis of variance on pharmacokinetic parameters reflecting eniluracil and 5-FU exposure was performed. RESULTS: Thirty-nine patients with advanced solid malignancies had complete pharmacokinetic studies performed during treatments A, B, and C. The pharmacokinetics of eniluracil and 5 FU were similar among the three types of treatment. Both strengths of the combined eniluracil/5-FU dosing form and the separate dosing forms were bioequivalent. Mean values for terminal half-life, systemic clearance, and apparent volume of distribution for oral 5-FU during treatments A/B/C were 5.5/5.6/5.6 hours, 6.6/6.6/6.5 liters/hour, and 50.7/51.5/50.0 liters, respectively. The intersubject coefficient of variation for pharmacokinetic variables reflecting 5-FU exposure and clearance in treatments ranged from 23% to 33%. The urinary excretion of unchanged 5-FU over 24 hours following treatments A, B, and C averaged 52.2%, 56.1%, and 50.8'%, of the administered dose of 5-FU, respectively. Parameters reflecting DPD inhibition, including plasma uracil and urinary FBAL excretion following treatments A, B, and C were similar. Toxicity was generally mild and similar following all three types of treatments. CONCLUSIONS: The pharmacokinetics of 5-FU and eniluracil were similar and met bioequivalence criteria following treatment with the separate oral formulations of 5-FU and eniluracil and two strengths of the combined formulation. The availability of a combined eniluracil/5-FU oral dosing form will likely simplify dosing and decrease the probability of severe toxicity or suboptimal therapeutic results caused by an inadvertent 5-FU overdose or insufficient 5-FU dosing in the case of separate oral formulations, thereby enhancing the overall feasibility and 0therapeutic index of oral 5-FU therapy. PMID- 11106123 TI - Biweekly irinotecan or raltitrexed plus 6S-leucovorin and bolus 5-fluorouracil in advanced colorectal carcinoma: a Southern Italy Cooperative Oncology Group phase II-III randomized trial. AB - PURPOSE: The aim of this randomised trial was to evaluate the activity and toxicity of a biweekly regimen including 6S-leucovorin-modulated 5-fluorouracil (LFA-5-FU), combined with either irinotecan (CPT-11 + LFA 5-FU) or raltitrexed (Tomudex) (TOM + LFA-5-FU), in advanced colorectal cancer patients, and to make a preliminary comparison of both these experimental regimens with a biweekly administration of LFA-5-FU modulated by methotrexate (MTX + LFA-5-FU). PATIENTS AND METHODS: One hundred fifty-nine patients with advanced colorectal carcinoma previously untreated for the metastatic disease (34 of them previously exposed to adjuvant 5-FU) were randomly allocated to receive: CPT-11, 200 mg/m2 i.v. on day 1, followed on day 2 by LFA, 250 mg/m2 i.v. infusion and 5-FU, 850 mg/m2 s i.v. bolus (arm A); TOM, 3 mg/m2 i.v. on day 1, followed on day 2 by LFA, 250 mg/m2 i.v. infusion and 5-FU, 1050 mg/m2 i.v. bolus (arm B); or MTX, 750 mg/m2 i.v. on day 1, followed on day 2 by LFA, 250 mg/m2 i.v. infusion and 5-FU, 800 mg/m2 i.v. bolus (arm C). Courses were repeated every two weeks in all arms of the trial. Response rate (RR) was evaluated after every four courses. The sample size was defined to have an 80% power to detect a 35% RR for each experimental treatment, and to show a difference of at least 4% in RR with the standard treatment if the true difference is 15% or more. RESULTS: The RRs were: 34% (95% confidence interval (95%, CI): 21%-48%) in arm A, including 3 complete responses (CRs) and 15 partial responses (PRs), 24% (95% CI: 14%-38%) in arm B, including 2 CRs and 11 PRs, and 24% (95% CI: 14%-38%), with 2 CRs and 11 PRs, in arm C. After a median follow-up time of 62 (range 18-108) weeks, the median time to progression was 38, 25, and 27 weeks for arm A, B, and C, respectively. With 94 patients still alive, the one-year probability of survival was 61%, 54%, and 59%, respectively. WHO grade 3 or 4 neutropenia and diarrhoea affected 46% and 16%, respectively, of patients treated with CPT-11 + LFA 5-FU. Median relative dose intensity over eight cycles (DI8) was 78% for CPT-11 and 82% for 5-FU. Severe toxicities of TOM + LFA-5-FU were neutropenia (16%) and diarrhoea (16%), but median relative DI8 was 93% for TOM, and 82% for 5-FU. CONCLUSIONS: CPT-11 + LFA 5-FU compares favorably in term of activity and toxicity with other combination regimens including CPT-11 and continuous infusional 5-FU. The hypothesis of a RR 15% higher than the MTX + LFA-5-FU treatment can not be ruled out after this interim analysis. The TOM + LFA 5-FU regimen showed a RR and a toxicity profile very close to the MTX + LFA 5-FU combination, and dose not deserve further evaluation in advanced colorectal cancer patients. PMID- 11106124 TI - Pemetrexed: a multitargeted antifolate agent with promising activity in solid tumors. AB - Pemetrexed disodium (ALIMTA, LY231514 (MTA)) is a novel multitargeted antifolate, that inhibits at least three enzymes involved in folate metabolism and DNA synthesis. These enzymes are thymidylate synthase, dihydrofolate reductase, and glycinamide ribonucleotide formyltransferase. This agent has shown broad antitumor activity in phase II trials in a wide variety of solid tumors, including non-small cell lung cancer, breast, colon, pancreas, bladder, head and neck, and cervical carcinomas, and is undergoing randomized phase III studies. MTA has also shown promising activity in a broad range of tumors in combination with other active agents such as gemcitabine and cisplatin. The clinical pharmacology, toxicity and clinical activity of MTA are reviewed in this report. PMID- 11106125 TI - Metastatic secretory breast cancer. Non-responsiveness to chemotherapy: case report and review of the literature. AB - Secretory carcinoma of the breast is a rare and indolent tumour originally described in children but occurring equally in the adult population. The principal management problems following primary surgical treatment are local recurrence and axillary lymph node metastases. Distant metastases are extremely rare. We present the case of a 27-year-old woman with pulmonary metastases from a secretory breast cancer treated by mastectomy and axillary lymph node dissection 12 years previously. There was no response to chemotherapy; however, the patient remained alive and active two years from presentation with metastatic disease and one year from cessation of all cytotoxic chemotherapy. She eventually died of respiratory failure two and a half years after presentation. To our knowledge, this is only the fourth reported case of distant metastases from secretory breast cancer and the second reported case in which current active chemotherapy has been used. We review the literature and discuss the apparent chemoresistance of this tumour including the lack of membrane staining for Her2/neu. In the absence of any proven effective chemotherapy we believe that symptom control becomes the focus of management and offers patients with metastatic secretory breast cancer the greatest chance of a functional and good quality existence. PMID- 11106126 TI - Tumor lysis syndrome following hemi-body irradiation for metastatic breast cancer. AB - Tumor lysis syndrome (TLS) is a rare serious acute complication of cancer therapy, reported mainly following chemotherapy in patients with large tumor load and chemosensitive disease. These are mainly patients with non-Hodgkin's lymphoma, leukemia and rarely in solid tumors. It is less frequently described after radiotherapy for lymphoid and hematological malignancies. TLS following radiotherapy for solid tumors is a very rare complication. In this report/review we describe a seventy-three-year-old male patient with progressive metastatic carcinoma of the breast to the lungs, liver and bone. He was referred for radiotherapy because of generalized bony pains. The patient was planned for sequential hemi-body irradiation starting with the more symptomatic upper half body. After premedication, he was given 8.5 Gy to the mid point at the maximum chest separation with anterior lung attenuator limiting uncorrected lung dose to 6.15 Gy. A further 3.5 Gy electron boost to the fungating breast tumor was given to the 100%. Forty-eight hours after irradiation he developed hyperkalemia, hyperphosphatemia, hyperuricemia, hypocalcemia and renal failure. These clinical and biochemical changes are typical of tumor lysis syndrome (TLS). Despite hydration, and treating the hyperuricemia, the patient developed coma and died eight days after irradiation. The prophylaxis and management of TLS and in high risk patients are described to avoid this frequently fatal complication. PMID- 11106127 TI - Breast involvement in immunolymphoproliferative disorders: report of two cases of multiple myeloma of the breast. AB - Breast involvement by immunolymphoproliferative disorders is rare. Primary and secondary malignant lymphomas of the breast are much more common than multiple myeloma, of which only 16 cases have been described. We report two cases of patients with an established diagnosis of multiple myeloma in whom breast involvement appeared during the course of the disease. We underline diagnostic difficulties caused by the lack of clinical and radiological features which allow us to differentiate between breast carcinoma and breast involvement by lymphoproliferative disease. Only fine needle aspiration and/or excisional biopsy can differentiate between immunolymphoproliferative disorders and epithelial or mesenchymal tumors of the breast. PMID- 11106128 TI - High-dose chemotherapy with autologous stem-cell transplantation in patients with pretreated advanced malignant melanoma. PMID- 11106129 TI - Concerns about clinical drug trials. PMID- 11106130 TI - Comparative abuse liability of intravenously administered remifentanil and fentanyl. AB - Remifentanil is a short-acting, esterase-metabolized opioid analgesic. This study compared the abuse liability of remifentanil with that of fentanyl and placebo in a randomized, double-blind, crossover study. Twelve recreational users of opioids received increasing doses of remifentanil (0.6, 1.2, 1.8, 2.4, and 3.0 microg/kg), fentanyl (0.4, 0.8, 1.3, 2.0, 3.0, and 4.5 microg/kg) and placebo via an intravenous infusion pump. Subjective measures (Cole/Addiction Research Center Inventory [ARCI] scales and visual analog scale [VAS] items such as "High" and "Good Effects") and physiologic variables (blood pressure, O2 saturation, pupil diameter) were recorded. For each measure, the differences from baseline were reduced to an area under the response curve (AUC) and a peak, and each subject's response to the maximum tolerable dose for each of the two active drug classes and mean response to several placebo infusions were entered into a 12 x 3 analysis of variance. All differences in drug versus placebo effects were significant. Although a majority of the peak effects that were measurable within 4 minutes after drug infusion reflected greater remifentanil effects, only one, High VAS, was statistically significant. In contrast, observations that could only be made > or = 5 minutes after drug infusion predominantly indicated significantly greater fentanyl peak effects, including High VAS, Liking VAS, Good Effects VAS, and Cole/ARCI Abuse Potential. Fentanyl AUCs were generally significantly larger than the corresponding remifentanil AUCs. A drug abuser seeking longer-lasting drug effects might select fentanyl over remifentanil, but these data do not completely rule out remifentanil abuse by some individuals with access to both the drug and the infusion equipment or by those who prefer briefer, repeated effects. PMID- 11106131 TI - A placebo-controlled study of lamotrigine and gabapentin monotherapy in refractory mood disorders. AB - There is a pressing need for additional treatment options for refractory mood disorders. This controlled comparative study evaluated the efficacy of lamotrigine (LTG) and gabapentin (GBP) monotherapy versus placebo (PLC). Thirty one patients with refractory bipolar and unipolar mood disorders participated in a double-blind, randomized, crossover series of three 6-week monotherapy evaluations including LTG, GBP, and PLC. There was a standardized blinded titration to assess clinical efficacy or to determine the maximum tolerated daily dose (LTG 500 mg or GBP 4,800 mg). The primary outcome measure was the Clinical Global Impressions Scale (CGI) for Bipolar Illness as supplemented by other standard rating instruments. The mean doses at week 6 were 274 +/- 128 mg for LTG and 3,987 +/- 856 mg for GBP. Response rates (CGI ratings of much or very much improved) were the following: LTG, 52% (16/31); GBP, 26% (8/31); and PLC, 23% (7/31) (Cochran's Q = 6.952, df = 2, N = 31, p = 0.031). Post hoc Q differences (df = 1, N = 31) were the following: LTG versus GBP (Qdiff = 5.33, p = 0.011); LTG versus PLC (Qdiff = 4.76, p = 0.022); and GBP versus PLC (Qdiff = 0.08, p = 0.70). With respect to anticonvulsant dose and gender, there was no difference between the responders and the nonresponders. The agents were generally well tolerated. This controlled investigation preliminarily suggests the efficacy of LTG in treatment-refractory affectively ill patients. Further definition of responsive subtypes and the role of these medications in the treatment of mood disorders requires additional study. PMID- 11106132 TI - Therapeutic effects of imipramine are counteracted by its metabolite, desipramine, in patients with generalized anxiety disorder. AB - Imipramine has been shown to reduce anxiety in patients with generalized anxiety disorder (GAD). However, some properties of imipramine may diminish or counteract its anxiolytic effects. The authors previously found that the greater the reduction in cardiac vagal control after 6 weeks of imipramine treatment, the smaller the improvement in anxiety-related symptoms. The purpose of this study was to determine whether the authors' previous findings were replicable and to gather information on the plasma levels of imipramine, desipramine (the major metabolite of imipramine), and anticholinergic levels. Fourteen patients with GAD were administered imipramine for 6 weeks. Their scores from self-administered and investigator-administered rating scales were obtained before and after the treatment, and the changes in these scores were contrasted with the changes in cardiac vagal tone, along with the absolute plasma levels of imipramine, desipramine, and anticholinergic activity at the end of week 6. The authors observed a greater improvement in symptoms of anxiety in those who showed the smallest decreases in cardiac vagal tone and in those who showed the smallest increases in desipramine and anticholinergic plasma levels. Moreover, strong relationships were observed between desipramine and anticholinergic levels. These results demonstrate that imipramine not only has therapeutic effects, but it may also have properties that result in physiologic states that counteract its therapeutic effects. Future research should investigate the direct anticholinergic effects of desipramine and determine whether there is a parallel between the anticholinergic effects and the clinical outcome of other pharmacologic treatments, including antidepressants with predominantly norepinephrine or serotonin reuptake inhibitory properties. PMID- 11106133 TI - Do antidepressants selectively suppress spontaneous (unexpected) panic attacks? A replication. AB - The purpose of this study was to test the following interrelated hypotheses in a larger sample by attempting to replicate supportive results from a small therapeutic study: (1) the pathogenesis of panic disorder includes at least two identifiable components: a biological component represented by spontaneous (unexpected) panic attacks, and a cognitive component represented by situational attacks and especially by phobias; (2) these components respond differently to treatment; (3) many biological processes respond to an effective intervention in proportion to their deviance from "normal" prior to treatment ("Law of Initial Value"); and (4) the response of spontaneous panic attacks to an effective treatment conforms to that model. Previously, the authors reanalyzed an 8-week therapeutic study of panic disorder that included groups treated with placebo and with imipramine (225 mg daily). The criteria of response were spontaneous panic attacks (biological component), situational panic attacks (both components), and agoraphobia ratings (cognitive component). The analyses compared the regression lines for posttreatment status on pretreatment status in the imipramine and placebo groups. The effect of imipramine on spontaneous panic attacks fitted the hypothesized model: the pre-post slope in the placebo group was approximately 1 (45 degrees), whereas the slope in the imipramine group was approximately 0. There was no significant difference in pre-post slopes between the imipramine and placebo groups for situational panic attacks or agoraphobia ratings. For this report, the authors applied the same approach to another larger data set from a study using a similar design, but a different antidepressant. In this multicenter, double-blind study, patients with panic disorder were randomly assigned to receive 10 weeks of treatment with placebo (N = 78) or fluoxetine 10 mg (N = 84) or 20 mg (N = 81) daily. Spontaneous and situational panic attacks were registered in a daily diary, and agoraphobia was rated at each visit. Using baseline and endpoint data, fluoxetine had a statistically significant, dose dependent, suppressive effect on spontaneous panic attacks, as measured by the pre-post slopes in the three treatment groups. The placebo group showed some response (slope = 0.69). There were no significant drug effects on situational panic attacks. On ratings of agoraphobia, the slopes in the placebo and the fluoxetine 20 mg groups did not differ, but the slope in the fluoxetine 10 mg group was significantly less than that in the placebo group, suggesting a therapeutic drug effect on agoraphobia only at the lower dose. These results are consistent with the stated hypotheses. They suggest that the therapeutic effects of antidepressants on panic disorder may be due primarily to the specific suppression of spontaneous panic attacks among patients with high baseline pathologic findings. Implications of these results for concepts of pathogenesis, clinical practice, and therapeutic research regarding panic disorder are discussed. PMID- 11106134 TI - A comparative pharmacokinetic and dynamic evaluation of alprazolam sustained release, bromazepam, and lorazepam. AB - Sustained-release (SR) alprazolam may facilitate compliance with oral benzodiazepine treatment of panic disorders that currently requires doses administered three or four times daily. To compare the pharmacokinetic, psychomotor performance, and subjective effects of alprazolam SR (1.5 mg), bromazepam (3 mg taken three times daily), and lorazepam (1 mg taken three times daily), 13 male volunteers (aged 20-45 years) randomly received on four separate occasions one of these medications or placebo. Once before and 11 times after drug administration, the subjects were tested using psychomotor performance tests (manual tracking and digit-symbol substitution test [DSST]) and computerized questionnaires (such as the Tufts University Benzodiazepine Scale [TUBS], the Addiction Research Center Inventory, and the visual analog scales) to determine the subjective effects of the drugs. Blood samples for the determination of the plasma levels of the drugs were collected before and 17 times after the drug was administered. A peak plateau of plasma alprazolam began approximately 6 hours after the dose, which was later than the initial peaks for lorazepam and bromazepam (1-2 hours after the dose). Once this plateau had begun, alprazolam SR sustained that concentration better than did the other two formulations. Of the 10 measures on which the response averaged for the first 14 hours differed among drugs (p < 0.05), bromazepam differed from placebo on two measures, lorazepam on four (including DSST Performance and TUBS Sedation), and alprazolam SR on nine (including all four affected by lorazepam). Lorazepam and alprazolam, but not bromazepam, produced significantly more sedation than placebo. The doses of the three drugs were not equipotent in sedation and mood effects. None of the drugs tested differed from placebo on measures relevant to abuse liability. PMID- 11106135 TI - Prevention of relapse in generalized social phobia: results of a 24-week study in responders to 20 weeks of sertraline treatment. AB - The aim of this study was to evaluate the efficacy, tolerability, and effects on quality of life of sertraline, a selective serotonin reuptake inhibitor, in the prevention of relapse of generalized social phobia. Fifty adult outpatients with generalized social phobia who were rated much or very much improved on the Clinical Global Impression Scale of Improvement (CGI-I) after 20 weeks of sertraline treatment (50-200 mg/day) were randomly assigned in a one-to-one ratio to either continue double-blind treatment with sertraline or immediately switch to placebo for another 24 weeks. The initial 20-week study was placebo controlled, and 15 responders to placebo also continued to receive double-blind placebo treatment in the continuation study. Eighty-eight percent of patients in the sertraline-continuation group and only 40% of patients in the placebo-switch and placebo-responder groups completed the study. In intent-to-treat endpoint analyses, 1 (4%) of 25 patients in the sertraline-continuation group and 9 (36%) of 25 patients in the placebo-switch group had relapsed at study endpoint (chi2 = 8.0, Fisher exact test, p = 0.01). The relative risk (hazards ratio) for relapse associated with placebo-switch relative to sertraline-continuation treatment was 10.2 (95% confidence interval, 1.3-80.7). Mean CGI-Severity, Marks Fear Questionnaire (MFQ) Social Phobia subscale, and Duke Brief Social Phobia Scale (BSPS) total scores were reduced by 0.07, 0.34, and 1.86 in the Sertraline Continuation group and increased by 0.88, 4.09, and 5.99 in the Placebo-Switch group (all F > 5.3, p < 0.03), respectively. CGI-Severity, MFQ Social Phobia subscale, and BSPS scores also increased in the Placebo-Responder group. Discontinuations because of lack of efficacy were 4% in the sertraline continuation group, 28% in the placebo-switch group (chi2 = 5.36, Fisher exact test, p = 0.049), relative to sertraline, and 27% in the placebo-responder group. Sertraline was effective in preventing relapse of generalized social phobia. Future research should assess whether improvements may be maintained or further increased by longer periods of treatment or through the addition of cognitive behavioral techniques. PMID- 11106136 TI - Sertraline versus paroxetine in major depression: clinical outcome after six months of continuous therapy. AB - reuptake inhibitors (SSRIs) during continuation therapy. This investigation reports the differential effect of 6 months of treatment with sertraline versus paroxetine for symptoms of depression, quality of life, and personality outcomes. Outpatients with unipolar major depression (DSM-III-R) were randomly assigned to receive 24 weeks of double-blind treatment with flexible doses of paroxetine (20 40 mg) or sertraline (50-150 mg). Assessments included the Montgomery-Asberg Depression Rating Scale (MADRS), the Clinical Global Impression Scale, the Battelle Quality of Life Questionnaire, and the Structured Clinical Interview for DSM-III-R Personality Disorders screen questionnaire. One hundred seventy-six patients (mean age, 43 years; 64% female; baseline MADRS, 30.3) were treated with sertraline and 177 patients (mean age, 42 years; 71% female; MADRS, 30.7) with paroxetine. Antidepressant efficacy during continuation therapy was sustained, with only 2% of patients receiving sertraline and 9% of patients receiving paroxetine suffering a relapse. Continuation therapy resulted in a substantial conversion of responders during short-term treatment to full remission: remitter rates increased from 52% to 80% for sertraline and from 57% to 74% for paroxetine. The improvements in quality of life were related to a reduced depression score. SSRI treatment had significant beneficial effects on both categorical and dimensional measures of personality. A logistic regression analysis identified early response (25% reduction in MADRS scores at week 2) as the most important predictor of treatment response, whereas high severity, chronicity, and poor baseline quality of life had no effect. Both treatments were well-tolerated, with sertraline having a somewhat lower side effect profile. Sertraline and paroxetine demonstrated comparable efficacy during short-term and continuation therapy. Treatment was associated with significant improvement in quality of life and with reductions in axis II personality psychopathology. PMID- 11106137 TI - Hematologic reference ranges in a population of patients with schizophrenia. AB - The potential hematotoxic effects of antipsychotic drugs are well known and may limit the use of some effective therapies. Although some previous studies have suggested that patients with schizophrenia may have altered "normal" values, only limited data were available. It is now believed that biological values do not usually follow a normal distribution; therefore, reference ranges are frequently used when interpreting laboratory tests in clinical practice and in research. However, it may be important to use disease-specific hematologic reference ranges when evaluating laboratory test results for patients with schizophrenia. In this study, data taken from patients with schizophrenia prior to treatment in previous phase II and phase III pharmaceutical studies were analyzed to produce reference ranges for a variety of hematologic parameters. An increased variability was shown in the reference ranges for all white blood cell indices between patients with schizophrenia and the population without schizophrenia. Certain reference values also showed heterogeneity for gender, age, and racial descent. This study suggests that abnormal hematologic findings in patients with schizophrenia should be assessed in the context of a valid reference range. This information will be of value to psychiatrists, laboratory scientists, and other physicians who encounter hematologic problems in patients with schizophrenia, as well as in the assessment of the adverse effects of new therapeutic agents. PMID- 11106138 TI - Side effects and time course of response in a placebo-controlled trial of fluoxetine for the treatment of geriatric depression. AB - A high rate of improvement among patients who receive placebo in controlled trials of antidepressants can complicate the evaluation of true drug effect. Placebo response may be a reaction to the psychosocial factors of study participation or a function of changes in the natural course of depression. Drug side effects may also influence patients' expectations, and they should be distinguished from the somatic symptoms associated with major depression. The authors reanalyzed data from a large, multicenter, placebo-controlled clinical trial of fluoxetine treatment of geriatric depression to evaluate similarities and differences between responders and nonresponders in both treatment groups. Specifically, the authors examined weekly somatic complaints as possible predictors of response and of dropout, as well as the time course and onset of response. Fluoxetine was superior to placebo on all outcome measures. Among somatic complaints associated with fluoxetine response, headache before and after randomization was associated with a good response and anxiety after randomization was associated with a poor response. Somnolence before and after randomization was associated with a good placebo response. Early and persistent improvement occurred among similar proportions of responders in both groups. The difference between fluoxetine and placebo seemed to be a persistent response beginning during the 4th week. Pretreatment somnolence was associated with early, persistent improvement in both groups and may serve as a marker for placebo response. PMID- 11106139 TI - Changes in energy during treatment of depression: an analysis of fluoxetine in double-blind, placebo-controlled trials. AB - More than two thirds of patients with depression present with symptoms of fatigue, low energy, and listlessness. Because daytime sedation may be a concern in such patients, a "nonsedating" antidepressant should be considered. The authors examined the effects of fluoxetine on depression-related disturbances in energy. Data from seven double-blind, placebo-controlled clinical trials in 2,075 patients with major depression were retrospectively analyzed. The Hamilton Rating Scale for Depression (HAM-D) Retardation factor score (total of items 1, 7, 8, and 14) was used as the primary measure of energy improvement, whereas the HAM-D 17 total score was used to assess changes in overall depression. Elderly patients (aged 60 years and older) were included in the overall group and were also analyzed separately. In addition, a subgroup analysis was performed using the HAM D Retardation factor score to categorize patients as having low (score < 8) or high (score > or = 8) levels of retardation at baseline. Beginning at week 3, fluoxetine-treated patients experienced statistically significant reductions in their HAM-D Retardation factor score compared with placebo-treated patients. The reductions for the elderly subgroup were less than those for the overall population, but they were still statistically significant beginning at week 4. Patients in both the low and high baseline retardation groups improved significantly. HAM-D-17 total scores for fluoxetine-treated patients in all groups (total, elderly, high retardation, and low retardation) improved significantly compared with placebo-treated patients. These findings demonstrate that fluoxetine-treated patients experience an improvement in energy symptoms as their overall depression improves. PMID- 11106140 TI - The drug-placebo response curve: a new method for assessing drug effects in clinical trials. AB - Unlike many other areas of medicine, psychiatric clinical trials have no gold standard laboratory tests with which to measure drug effects. Although reliable and valid psychometric measures are available, the standard analysis of such measures obscures the clinical relevance of drug effects. In this study, the authors sought to address this problem by developing a graphical display called the drug-placebo response curve. The features of the drug-placebo response curve are described by applying it to a previously published, controlled clinical trial of desipramine in the treatment of adults with attention-deficit/hyperactivity disorder. Unlike the standard analysis, the drug-placebo response curve showed that desipramine was effective over the full range of response and that it prevented worsening, in addition to increasing the likelihood of improvement. The drug-placebo response curve provides clinically relevant information about the effect of drugs on continuous outcomes in controlled clinical trials. PMID- 11106141 TI - A double-blind, placebo-controlled study on the effects of Gotu Kola (Centella asiatica) on acoustic startle response in healthy subjects. AB - Investigations of the pharmacologic profile of medicinal plants have revealed that a number of plants with purported anxiolytic activity bind to cholecystokinin (CCK) receptors. This finding is intriguing in view of the proposed involvement of CCK in the pathophysiology of fear and anxiety. This double-blind, placebo-controlled study was undertaken to evaluate the anxiolytic activity of Gotu Kola (Centella asiatica) in healthy subjects. Gotu Kola has been used for centuries in Ayurvedic and traditional Chinese medicine to alleviate symptoms of depression and anxiety. Recent studies in the rat have shown that long-term pretreatment with Gotu Kola decreases locomotor activity, enhances elevated-plus maze performance, and attenuates the acoustic startle response (ASR). In this study, the authors evaluated the effects of Gotu Kola on the ASR in humans. Subjects were randomly assigned to receive either a single 12-g orally administered dose of Gotu Kola (N = 20) or placebo (N = 20). The results revealed that compared with placebo, Gotu Kola significantly attenuated the peak ASR amplitude 30 and 60 minutes after treatment. Gotu Kola had no significant effect on self-rated mood, heart rate, or blood pressure. These preliminary findings suggest that Gotu Kola has anxiolytic activity in humans as revealed by the ASR. It remains to be seen whether this herb has therapeutic efficacy in the treatment of anxiety syndromes. PMID- 11106142 TI - Epidemiologic investigation of the relative clearance of haloperidol by mixed effect modeling using routine clinical pharmacokinetic data in Japanese patients. AB - The steady-state trough concentrations of haloperidol were studied to clarify the role of the characteristics of Japanese patients in estimating haloperidol dosing regimens by using routine therapeutic drug-monitoring data. Nonlinear mixed effects modeling (NONMEM) was used to estimate the effect of a variety of developmental and demographic factors on haloperidol clearance values using 270 serum level measurements obtained from 191 patients during their clinical course. The final model describing haloperidol's relative clearance was CL = 0.74 x TBW(0.594) x DOSE(0.326) x 1.32CO1 x 0.867AGE, where CL is clearance (measured in liters per hour), TBW is the total body weight (in kilograms), DOSE is the daily dose of haloperidol (in grams per kilogram per day), CO1 = 1 for concomitant administration of antiepileptic drugs (phenobarbital, phenytoin, or carbamazepine) and CO1 = 0 otherwise, and AGE = 1 for patients aged 55 years or older and AGE = 0 otherwise. Concomitant administration of haloperidol and antiepileptic drugs resulted in a 32% increase in haloperidol clearance. Patients aged 55 years or older showed a 13.3% reduction in clearance values compared with the younger population. PMID- 11106143 TI - First-night effect of melatonin treatment in patients with chronic schizophrenia. AB - The first-night effect (FNE) is the tendency for individuals to sleep worse than normal during their first night of polysomnographic sleep evaluation. FNE reflects the adaptive increase of alertness and perhaps the stress resulting from an unfamiliar sleeping environment. This effect is usually absent in patients with chronic schizophrenia. Melatonin (N-acetyl-5-methoxy-tryptamine), the hormone secreted by the pineal gland at night, has been found to improve sleep in elderly patients with insomnia and recently in patients with chronic schizophrenia. The authors used FNE as a marker to explore the neurobehavioral responses of patients with chronic schizophrenia to melatonin treatment. In a randomized, double-blind, crossover trial, 14 patients with chronic schizophrenia were administered melatonin (2 mg in a controlled-release formulation) or placebo for 3 weeks with a 1-week washout between treatment periods. Polysomnography was performed during the last two consecutive nights of each treatment period. The following significant FNEs were observed with melatonin treatment: (1) rapid eye movement sleep latency was longer; (2) sleep efficiency was lower; and (3) the duration of wakefulness during sleep was lower on the first night than on the second night. These effects were not found when the patients received a placebo. The FNE was manifested regardless of whether melatonin was administered before or after the placebo treatment period. For the first time, these results show that melatonin treatment exaggerates FNE in patients with chronic schizophrenia, thereby suggesting an improved ability of these patients to mobilize alertness in unfamiliar surroundings. PMID- 11106144 TI - Off-label use of antipsychotic drugs. AB - Despite the fact that most antipsychotics have only been formally evaluated for the treatment of schizophreniform disorder, schizophrenia, mania, and schizoaffective disorder (defined as "classical indications"), antipsychotics are widely used for the treatment of a broad range of symptoms and disorders. In this study, 173 patients who were having their prescriptions for antipsychotics filled at local pharmacies were interviewed. In 115 patients (66.5%), an antipsychotic was prescribed for off-label indications. Patients most often stated that they took antipsychotics as a tranquilizer or an anxiolytic. Neither gender, education, duration of treatment, nor efficacy of treatment showed an influence on the prescription practices for antipsychotics. In contrast, family status and side effects showed a significant influence. A classical indication was more often found in married and widowed patients than in unmarried or divorced ones. Patients in whom antipsychotics were prescribed for the treatment of schizophrenia, schizophreniform disorder, mania, or schizoaffective disorder experienced side effects more often than others. Age was also important for the indication of antipsychotics. Classical indications of antipsychotics were most often found in patients aged 30 to 49 years. In older patients (49-70 years), antipsychotics were almost exclusively used for off-label indications. In classical indications, clozapine was used more frequently (50%) than other antipsychotics. Melperone was primarily prescribed for off-label use. PMID- 11106145 TI - Selection bias in clinical trials with antipsychotics. AB - Although the selection of patients is known to be a powerful factor affecting the results of clinical trials, little is known about recruitment issues. Many patients with schizophrenia who are screened for a clinical trial of an investigational antipsychotic are ultimately not included in the study. Therefore, the question arises of whether the results obtained by studying a selected group of patients are really representative of the general population of patients with schizophrenia. The authors studied possible reasons for selective sampling in 200 patients who were consecutively admitted to inpatient units of Innsbruck's Department of Psychiatry with a diagnosis of schizophreniform or schizophrenic disorder over a time period of 33 months. Apart from demographic data and a psychopathologic rating (using the Brief Psychiatric Rating Scale), the authors recorded whether or not a patient was included in a phase III study and whether or not those were not included would have theoretically been eligible for such a study. Twenty-seven patients were finally recruited for a clinical trial. These patients were younger, on average, had a more recent onset of illness, and had experienced fewer psychotic episodes in the past. A history of noncompliance with previous treatment and the refusal of consent were the most common reasons for not including theoretically eligible patients in a clinical trial. PMID- 11106146 TI - Olanzapine prolongation of granulocytopenia after clozapine discontinuation. PMID- 11106147 TI - Neuropletic malignant syndrome and olanzapine. PMID- 11106148 TI - Regional c-fos expression in rat brain may predict antipsychotic therapeutic window. PMID- 11106149 TI - Mecamylamine increases cigarette smoking in psychiatric patients. PMID- 11106151 TI - Risperidone treatment of aggressive behavior in children with Tourette syndrome. PMID- 11106150 TI - Fluvoxamine increases plasma and urinary levels of clozapine and its major metabolites in a time- and dose-dependent manner. PMID- 11106152 TI - A case of paroxetine-induced dyskinetic movements. PMID- 11106153 TI - Serotonin syndrome after small doses of citalopram or sertraline. PMID- 11106154 TI - The effects of fluoxetine versus nortriptyline on body weight in depression. PMID- 11106155 TI - Orlistat in the treatment of psychopharmacologically induced weight gain. PMID- 11106156 TI - Nefazodone in the breast milk of nursing mothers: a report of two patients. PMID- 11106157 TI - Successful molecular dynamics simulation of the zinc-bound farnesyltransferase using the cationic dummy atom approach. AB - Farnesyltransferase (FT) inhibitors can suppress tumor cell proliferation without substantially interfering with normal cell growth, thus holding promise for cancer treatment. A structure-based approach to the design of improved FT inhibitors relies on knowledge of the conformational flexibility of the zinc containing active site of FT. Although several X-ray structures of FT have been reported, detailed information regarding the active site conformational flexibility of the enzyme is still not available. Molecular dynamics (MD) simulations of FT can offer the requisite information, but have not been applied due to a lack of effective methods for simulating the four-ligand coordination of zinc in proteins. Here, we report in detail the problems that occurred in the conventional MD simulations of the zinc-bound FT and a solution to these problems by employing a simple method that uses cationic dummy atoms to impose orientational requirement for zinc ligands. A successful 1.0 ns (1.0 fs time step) MD simulation of zinc-bound FT suggests that nine conserved residues (Asn127alpha, Gln162alpha, Asn165alpha, Gln195alpha, His248beta, Lys294beta, Leu295beta, Lys353beta, and Ser357beta) in the active site of mammalian FT are relatively mobile. Some of these residues might be involved in the ligand-induced active site conformational rearrangement upon binding and deserve attention in screening and design of improved FT inhibitors for cancer chemotherapy. PMID- 11106158 TI - Exchanging the active site between phytases for altering the functional properties of the enzyme. AB - By using a novel consensus approach, we have previously managed to generate a fully synthetic phytase, consensus phytase-1, that was 15-26 degrees C more thermostable than the parent fungal phytases used in its design (Lehmann et al., 2000). We now sought to use the backbone of consensus phytase-1 and to modify its catalytic properties. This was done by replacing a considerable part of the active site (i.e., all the divergent residues) with the corresponding residues of Aspergillus niger NRRL 3135 phytase, which displays pronounced differences in specific activity, substrate specificity, and pH-activity profile. For the new protein termed consensus phytase-7, a major - although not complete - shift in catalytic properties was observed, demonstrating that rational transfer of favorable catalytic properties from one phytase to another is possible by using this approach. Although the exchange of the active site was associated with a 7.6 degrees C decrease in unfolding temperature (Tm) as measured by differential scanning calorimetry, consensus phytase-7 still was >7 degrees C more thermostable than all wild-type ascomycete phytases known to date. Thus, combination of the consensus approach with the selection of a "preferred" active site allows the design of a thermostabilized variant of an enzyme family of interest that (most closely) matches the most favorable catalytic properties found among its family members. PMID- 11106159 TI - Two-state vs. multistate protein unfolding studied by optical melting and hydrogen exchange. AB - A direct conflict between the stabilization free energy parameters of cytochrome c determined by optical methods and by hydrogen exchange (HX) is quantitatively explained when the partially folded intermediates seen by HX are taken into account. The results support the previous HX measurements of intermediate populations, show how intermediates can elude the standard melting analysis, and illustrate how they confuse the analysis when they are significantly populated within the melting transition region. PMID- 11106160 TI - The acid-induced folded state of Sac7d is the native state. AB - Sac7d unfolds at low pH in the absence of salt, with the greatest extent of unfolding obtained at pH 2. We have previously shown that the acid unfolded protein is induced to refold by decreasing the pH to 0 or by addition of salt (McCrary BS, Bedell J. Edmondson SP, Shriver JW, 1998, J Mol Biol 276:203-224). Both near-ultraviolet circular dichroism spectra and ANS fluorescence enhancements indicate that the acid- and salt-induced folded states have a native fold and are not molten globular. 1H,15N heteronuclear single quantum coherence NMR spectra confirm that the native, acid-, and salt-induced folded states are essentially identical. The most significant differences in amide 1H and 15N chemical shifts are attributed to hydrogen bonding to titrating carboxyl side chains and through-bond inductive effects. The 1H NMR chemical shifts of protons affected by ring currents in the hydrophobic core of the acid- and salt-induced folded states are identical to those observed in the native. The radius of gyration of the acid-induced folded state at pH 0 is shown to be identical to that of the native state at pH 7 by small angle X-ray scattering. We conclude that acid-induced collapse of Sac7d does not lead to a molten globule but proceeds directly to the native state. The folding of Sac7d as a function of pH and anion concentration is summarized with a phase diagram that is similar to those observed for other proteins that undergo acid-induced folding except that the A-state is encompassed by the native state. These results demonstrate that formation of a molten globule is not a general property of proteins that are refolded by acid. PMID- 11106162 TI - Crystal structure of an in vivo HIV-1 protease mutant in complex with saquinavir: insights into the mechanisms of drug resistance. AB - Saquinavir is a widely used HIV-1 protease inhibitor drug for AIDS therapy. Its effectiveness, however, has been hindered by the emergence of resistant mutations, a common problem for inhibitor drugs that target HIV-1 viral enzymes. Three HIV-1 protease mutant species, G48V, L90M, and G48V/L90M double mutant, are associated in vivo with saquinavir resistance by the enzyme (Jacobsen et al., 1996). Kinetic studies on these mutants demonstrate a 13.5-, 3-, and 419-fold increase in Ki values, respectively, compared to the wild-type enzyme (Ermolieff J, Lin X, Tang J, 1997, Biochemistry 36:12364-12370). To gain an understanding of how these mutations modulate inhibitor binding, we have solved the HIV-1 protease crystal structure of the G48V/L90M double mutant in complex with saquinavir at 2.6 A resolution. This mutant complex is compared with that of the wild-type enzyme bound to the same inhibitor (Krohn A, Redshaw S, Richie JC, Graves BJ, Hatada MH, 1991, J Med Chem 34:3340-3342). Our analysis shows that to accommodate a valine side chain at position 48, the inhibitor moves away from the protease, resulting in the formation of larger gaps between the inhibitor P3 subsite and the flap region of the enzyme. Other subsites also demonstrate reduced inhibitor interaction due to an overall change of inhibitor conformation. The new methionine side chain at position 90 has van der Waals interactions with main chain atoms of the active site residues resulting in a decrease in the volume and the structural flexibility of S1/S1' substrate binding pockets. Indirect interactions between the mutant methionine side chain and the substrate scissile bond or the isostere part of the inhibitor may differ from those of the wild-type enzyme and therefore may facilitate catalysis by the resistant mutant. PMID- 11106161 TI - Structure of a rat alpha1-macroglobulin receptor-binding domain dimer. AB - Alpha-macroglobulin inhibits a broad spectrum of proteinases by forming macromolecular cages inside which proteinases are cross-linked and trapped. Upon formation of a complex with proteinase, alpha-macroglobulin undergoes a large conformational change that results in the exposure of its receptor-binding domain (RBD). Engagement of this domain by alpha-macroglobulin receptor permits clearance of the alpha-macroglobulin: proteinase complex from circulation. The crystal structure of rat alpha1-macroglobulin RBD has been determined at 2.3 A resolution. The RBD is composed of a nine-stranded beta-sandwich and a single alpha-helix that has been implicated as part of the receptor binding site and that lies on the surface of the beta-sandwich. The crystallographic asymmetric unit contains a dimer of RBDs related by approximate twofold symmetry such that the putative receptor recognition sites of the two monomers are contiguous. By gel filtration and ultracentrifugation, it is shown that RBD dimers form in solution with a dissociation constant of approximately 50 microM. The structure of the RBD dimer might mimic a conformation of transformed alpha-macroglobulin in which the proposed receptor binding residues are exposed on one face of the dimer. A pair of phenylalanine residues replaces a cystine that is conserved in other members of the macroglobulin family. These residues participate in a network of aromatic side-chain interactions that appears to stabilize the dimer interface. PMID- 11106163 TI - The binding of myristoylated N-terminal nonapeptide from neuro-specific protein CAP-23/NAP-22 to calmodulin does not induce the globular structure observed for the calmodulin-nonmyristylated peptide complex. AB - CAP-23/NAP-22, a neuron-specific protein kinase C substrate, is Nalpha myristoylated and interacts with calmodulin (CaM) in the presence of Ca2+ ions. Takasaki et al. (1999, J Biol Chem 274:11848-11853) have recently found that the myristoylated N-terminal nonapeptide of CAP-23/NAP-22 (mC/N9) binds to Ca2+ bound CaM (Ca2+/CaM). In the present study, small-angle X-ray scattering was used to investigate structural changes of Ca2+/CaM induced by its binding to mC/N9 in solution. The binding of one mC/N9 molecule induced an insignificant structural change in Ca2+/CaM. The 1:1 complex appeared to retain the extended conformation much like that of Ca2+/CaM in isolation. However, it could be seen that the binding of two mC/N9 molecules induced a drastic structural change in Ca2+/CaM, followed by a slight structural change by the binding of more than two but less than four mC/N9 molecules. Under the saturated condition (the molar ratio of 1:4), the radius of gyration (Rg) for the Ca2+/CaM-mC/N9 complex was 19.8 +/- 0.3 A. This value was significantly smaller than that of Ca2+/CaM (21.9 +/- 0.3 A), which adopted a dumbbell structure and was conversely 2-3 A larger than those of the complexes of Ca2+/CaM with the nonmyristoylated target peptides of myosin light chain kinase or CaM kinase II, which adopted a compact globular structure. The pair distance distribution function had no shoulder peak at around 40 A, which was mainly due to the dumbbell structure. These results suggest that Ca2+/CaM interacts with Nalpha-myristoylated CAP-23/NAP-22 differently than it does with other nonmyristoylated target proteins. The N-terminal amino acid sequence alignment of CAP-23/NAP-22 and other myristoylated proteins suggests that the protein myristoylation plays important roles not only in the binding of CAP-23/NAP-22 to Ca2+/CaM, but also in the protein-protein interactions related to other myristoylated proteins. PMID- 11106164 TI - Divalent metal cofactor binding in the kinetic folding trajectory of Escherichia coli ribonuclease HI. AB - Proteins often require cofactors to perform their biological functions and must fold in the presence of their cognate ligands. Using circular dichroism spectroscopy. we investigated the effects of divalent metal binding upon the folding pathway of Escherichia coli RNase HI. This enzyme binds divalent metal in its active site, which is proximal to the folding core of RNase HI as defined by hydrogen/deuterium exchange studies. Metal binding increases the apparent stability of native RNase HI chiefly by reducing the unfolding rate. As with the apo-form of the protein, refolding from high denaturant concentrations in the presence of Mg2+ follows three-state kinetics: formation of a rapid burst phase followed by measurable single exponential kinetics. Therefore, the overall folding pathway of RNase HI is minimally perturbed by the presence of metal ions. Our results indicate that the metal cofactor enters the active site pocket only after the enzyme reaches its native fold, and therefore, divalent metal binding stabilizes the protein by decreasing its unfolding rate. Furthermore, the binding of the cofactor is dependent upon a carboxylate critical for activity (Asp10). A mutation in this residue (D10A) alters the folding kinetics in the absence of metal ions such that they are similar to those observed for the unaltered enzyme in the presence of metal. PMID- 11106165 TI - Lipoylating and biotinylating enzymes contain a homologous catalytic module. AB - Biotin and lipoic acid moieties are the covalently attached coenzyme cofactors of several multicomponent enzyme complexes that catalyze key metabolic reactions. Attachment of these moieties to the biotinyl- and lipoyl-dependent enzymes is post-translationally catalyzed by specific biotinylating and lipoylating protein enzymes. In Escherichia coli, two different enzymes, LplA and LipB, catalyze independent pathways for the lipoylation of the relevant enzymes, whereas only one enzyme, the BirA protein, is responsible for all the biotinylation. Counterparts of the E. coli BirA, LplA, and LipB enzymes have been previously identified in many organisms, but homology among the three families has never been reported. Computational analysis based on PSI-BLAST profiles and secondary structure predictions indicates, however, that lipoylating and biotinylating enzymes are evolutionarily related protein families containing a homologous catalytic module. Sequence conservation among the three families is very poor, but a single lysine residue is strictly conserved in all of them, which, according to the available X-ray crystal structure of the E. coli BirA protein, is expected to contribute to the binding of lipoic acid in the LplA and LipB enzymes. PMID- 11106166 TI - The PA domain: a protease-associated domain. AB - We have identified a similarity between the apical domain of the human transferrin receptor and several other protein families. This domain is found associated with two different families of peptidases. Therefore, we term it the PA domain for protease-associated domain. The PA domain is found inserted within a loop of the peptidase domain of family M8/M33 zinc peptidases. The PA domain is also found in a vacuolar sorting receptor and a ring finger protein of unknown function that may be a cell surface receptor. The PA domain may mediate substrate determination of peptidases or form protein-protein interactions. PMID- 11106167 TI - Constraint-based assembly of tertiary protein structures from secondary structure elements. AB - A challenge in computational protein folding is to assemble secondary structure elements-helices and strands-into well-packed tertiary structures. Particularly difficult is the formation of beta-sheets from strands, because they involve large conformational searches at the same time as precise packing and hydrogen bonding. Here we describe a method, called Geocore-2, that (1) grows chains one monomer or secondary structure at a time, then (2) disconnects the loops and performs a fast rigid-body docking step to achieve canonical packings, then (3) in the case of intrasheet strand packing, adjusts the side-chain rotamers; and finally (4) reattaches loops. Computational efficiency is enhanced by using a branch-and-bound search in which pruning rules aim to achieve a hydrophobic core and satisfactory hydrogen bonding patterns. We show that the pruning rules reduce computational time by 10(3)- to 10(5)-fold, and that this strategy is computationally practical at least for molecules up to about 100 amino acids long. PMID- 11106168 TI - Modification of the substrate specificity of porcine pepsin for the enzymatic production of bovine hide gelatin. AB - The substrate specificity of porcine pepsin has been altered by site-directed mutagenesis in an attempt to selectively cleave bovine hide collagen at only a few sites, similar to cathepsin D, for the production of high quality gelatin. Kinetic parameters were determined using chromogenic peptide substrates based on the sequence Lys-Pro-Xaa-Yaa-Phe*Nph-Arg-Leu (where Xaa is Ile or Pro, Yaa is Glu. Leu, Gln or Lys, Nph is p-nitrophenylalanine, and * is the site of cleavage). Substitution of Thr222 and Glu287 within the S2 subsite of pepsin by Val and Met, respectively, produced a double mutant with a two- to fourfold higher kcat/Km, compared with wild-type pepsin, for the chromogenic peptides with residues Leu, Gln, and Glu at position P2 (Yaa). The results suggest that the functional group of the P2 side chain may be exposed to solvent, while the aliphatic portion interacts with hydrophobic residues comprising S2. Wild-type pepsin cleaved a peptide corresponding to the carboxy-terminal telopeptide region of bovine type I collagen alpha1 chain, SGGYDLSFLPQPPQE, predominantly at three sites (Asp-Leu, Leu-Ser, and Phe-Leu) and at a significantly lower rate at Ser Phe. However, Thr222Val/Glu287Met cleaved site Ser-Phe at a rate 20-fold higher than the wild-type. Significantly, enzymes containing the double substitution Phe111Thr/Leu112Phe cleaved this peptide predominantly at one site Leu-Ser (similar to cathepsin D) and at a rate 23-fold higher than the wild-type. These mutants can potentially enhance the rate of solubilization of bovine hide collagen under conditions mild enough to maintain the triple helix structure and hence minimize the rate of subsequent denaturation and proteolytic cleavage. PMID- 11106169 TI - Formation of insulin amyloid fibrils followed by FTIR simultaneously with CD and electron microscopy. AB - Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), and electron microscopy (EM) have been used simultaneously to follow the temperature induced formation of amyloid fibrils by bovine insulin at acidic pH. The FTIR and CD data confirm that, before heating, insulin molecules in solution at pH 2.3 have a predominantly native-like alpha-helical structure. On heating to 70 degrees C, partial unfolding occurs and results initially in aggregates that are shown by CD and FT-IR spectra to retain a predominantly helical structure. Following this step, changes in the CD and FTIR spectra occur that are indicative of the extensive conversion of the molecular conformation from alpha-helical to beta-sheet structure. At later stages, EM shows the development of fibrils with well-defined repetitive morphologies including structures with a periodic helical twist of approximately 450 A. The results indicate that formation of fibrils by insulin requires substantial unfolding of the native protein, and that the most highly ordered structures result from a slow evolution of the morphology of the initially formed fibrillar species. PMID- 11106170 TI - New insight on beta-lactoglobulin binding sites by 1-anilinonaphthalene-8 sulfonate fluorescence decay. AB - The fluorescence time decay parameters of the beta-lactoglobulin-1 anilinonaphthalene-8-sulfonate complex have been investigated under physical and chemical perturbations (2 < pH < 8 and added electrolyte 0 < NaCl < 0.5 M) to obtain new insight on the nature of the protein binding interactions. A double exponential decay of the bound probe lifetime has been confirmed by the presence of a longer component, 11 to 14.5 ns, and a shorter component, 2.5 to 3.5 ns. The two lifetimes are ascribed to different binding modes associated also with different exposure to the solvent; in particular, the longer component is attributed to binding inside the hydrophobic beta barrel, while a "surface" site is suggested for the shorter component. A detailed analysis of the lifetime fractional intensities correlates the binding constants with ionic strength and supports the presence of electrostatic effects at both sites. A Debye-Huckel approach, applied to extrapolate the electrostatic free energy contribution vs. pH at vanishing ionic strength, gives interesting clues on the effective charge felt by the ANS ligands in the proximity of each site. In particular, binding is found to parallel the aspartate and glutamate titrations between pH 3 and pH 4.5; the "surface" site mainly responds to the presence of these local titrating charges while the "internal" site more closely follows the overall protein net charge. PMID- 11106171 TI - Comparison of binding energies of SrcSH2-phosphotyrosyl peptides with structure based prediction using surface area based empirical parameterization. AB - The prediction of binding energies from the three-dimensional (3D) structure of a protein-ligand complex is an important goal of biophysics and structural biology. Here, we critically assess the use of empirical, solvent-accessible surface area based calculations for the prediction of the binding of Src-SH2 domain with a series of tyrosyl phosphopeptides based on the high-affinity ligand from the hamster middle T antigen (hmT), where the residue in the pY+ 3 position has been changed. Two other peptides based on the C-terminal regulatory site of the Src protein and the platelet-derived growth factor receptor (PDGFR) are also investigated. Here, we take into account the effects of proton linkage on binding, and test five different surface area-based models that include different treatments for the contributions to conformational change and protein solvation. These differences relate to the treatment of conformational flexibility in the peptide ligand and the inclusion of proximal ordered solvent molecules in the surface area calculations. This allowed the calculation of a range of thermodynamic state functions (deltaCp, deltaS, deltaH, and deltaG) directly from structure. Comparison with the experimentally derived data shows little agreement for the interaction of SrcSH2 domain and the range of tyrosyl phosphopeptides. Furthermore, the adoption of the different models to treat conformational change and solvation has a dramatic effect on the calculated thermodynamic functions, making the predicted binding energies highly model dependent. While empirical, solvent-accessible surface area based calculations are becoming widely adopted to interpret thermodynamic data, this study highlights potential problems with application and interpretation of this type of approach. There is undoubtedly some agreement between predicted and experimentally determined thermodynamic parameters: however, the tolerance of this approach is not sufficient to make it ubiquitously applicable. PMID- 11106172 TI - A 16-amino acid peptide from human alpha2-macroglobulin binds transforming growth factor-beta and platelet-derived growth factor-BB. AB - Alpha2-macroglobulin (alpha2M) is a major carrier of transforming growth factor beta (TGF-beta) in vitro and in vivo. By screening glutathione S-transferase (GST) fusion proteins with overlapping sequences, we localized the TGFbeta binding site to aa 700-738 of the mature human alpha2M subunit. In separate experiments, we screened overlapping synthetic peptides corresponding to aa 696 777 of alpha2M and identified a single 16-mer (718-733) that binds TGF-beta1. Platelet-derived growth factor-BB (PDGF-BB) bound to the same peptide, even though TGF-beta and PDGF-BB share almost no sequence identity. The sequence of the growth factor-binding peptide, WDLVVVNSAGVAEVGV, included a high proportion of hydrophobic amino acids. The analogous peptide from murinoglobulin, a human alpha2M homologue that does not bind growth factors, contained only three nonconservative amino acid substitutions; however, the MUG peptide failed to bind TGF-beta1 and PDGF-BB. These results demonstrate that a distinct and highly restricted site in alpha2M, positioned near the C-terminal flank of the bait region, mediates growth factor binding. At least part of the growth factor binding site is encoded by exon 18 of the alpha2M gene, which is notable for a 5' splice site polymorphism that has been implicated in Alzheimer's Disease. PMID- 11106173 TI - Role of a solvent-exposed aromatic cluster in the folding of Escherichia coli CspA. AB - Escherichia coli CspA is a member of the cold shock protein family. All cold shock proteins studied to date fold rapidly by an apparent two-state mechanism. CspA contains an unusual cluster of aromatic amino acids on its surface that is necessary for nucleic acid binding and also provides stability to CspA (Hillier et al., 1998). To elucidate the role this aromatic cluster plays in the determining the folding rate and pathway of CspA, we have studied the folding kinetics of mutants containing either leucine or serine substituted for Phe 18, Phe20, and/or Phe31. The leucine substitutions are found to accelerate folding and the serine substitutions to decelerate folding. Because these residues exert effects on the free energy of the folding transition state, they may be necessary for nucleating folding. They are not responsible, however, for the very compact, native-like transition state ensemble seen in the cold shock proteins, as the refolding rates of the mutants all show a similar, weak dependence of unfolding rate on denaturant concentration. Using mutant cycle analysis, we show that there is energetic coupling among the three residues between the unfolded and transition states, suggesting that the cooperative nature of these interactions helps to determine the unfolding rate. Overall, our results suggest that separate evolutionary pressures can act simultaneously on the same group of residues to maintain function, stability, and folding rate. PMID- 11106174 TI - Protein renaturation by the liquid organic salt ethylammonium nitrate. AB - The room-temperature liquid salt, ethylammonium nitrate (EAN), has been used to enhance the recovery of denatured-reduced hen egg white lysozyme (HEWL). Our results show that EAN has the ability to prevent aggregation of the denatured protein. The use of EAN as a refolding additive is advantageous because the renaturation is a one-step process. When HEWL was denatured reduced using routine procedures and renatured using EAN as an additive, HEWL was found to regain 75% of its activity. When HEWL was denatured and reduced in neat EAN, dilution resulted in over 90% recovery of active protein. An important aspect of this process is that renaturation of HEWL occurs at concentrations of 1.6 mg/mL, whereas other renaturation processes occur at significantly lower protein concentrations. Additionally, the refolded-active protein can be separated from the molten salt by simple desalting methods. Although the use of a low temperature molten salt in protein renaturation is unconventional, the power of this approach lies in its simplicity and utility. PMID- 11106175 TI - Engineering the substrate specificity of Escherichia coli asparaginase. II. Selective reduction of glutaminase activity by amino acid replacements at position 248. AB - The use of Escherichia coli asparaginase II as a drug for the treatment of acute lymphoblastic leukemia is complicated by the significant glutaminase side activity of the enzyme. To develop enzyme forms with reduced glutaminase activity, a number of variants with amino acid replacements in the vicinity of the substrate binding site were constructed and assayed for their kinetic and stability properties. We found that replacements of Asp248 affected glutamine turnover much more strongly than asparagine hydrolysis. In the wild-type enzyme, N248 modulates substrate binding to a neighboring subunit by hydrogen bonding to side chains that directly interact with the substrate. In variant N248A, the loss of transition state stabilization caused by the mutation was 15 kJ mol(-1) for L glutamine compared to 4 kJ mol(-1) for L-aspartic beta-hydroxamate and 7 kJ mol( 1) for L-asparagine. Smaller differences were seen with other N248 variants. Modeling studies suggested that the selective reduction of glutaminase activity is the result of small conformational changes that affect active-site residues and catalytically relevant water molecules. PMID- 11106176 TI - Purification and refolding of vascular endothelial growth factor-B. AB - Vascular endothelial growth factor (VEGF)-A interacts with the receptor tyrosine kinases VEGF-R1 and R2, and the importance of this interaction in endothelial cell (EC) function and blood vessel development has been well documented. Other ligands that interact differentially with these receptors and that are structurally related to VEGF-A include VEGF-B, VEGF-C, VEGF-D, and placenta growth factor (PLGF). Compared with VEGF-A, relatively little is known about the biological role of the VEGF-R1 specific ligand, VEGF-B. Two splice variant isoforms that differ at the COOH-terminus and which retain unique solubility characteristics are widely expressed throughout embryonic and postnatal development. Recent analysis of mice with a targeted deletion of the VEGF-B gene has revealed a defect in heart development and function consistent with an important role in vascularization of the myocardium (Bellomo D et al., 2000, Circ Res 86:E29-E35). To facilitate further characterization of VEGF-B, we have developed a protocol for expression and purification of refolded recombinant protein from Escherichia coli inclusion bodies (IBs). The approach developed resolves a number of significant issues associated with VEGF-B, including the ability to heterodimerize with endogenous VEGF-A when co-expressed in mammalian cells, a complex secondary structure incorporating inter- and intrachain disulfide bonds and hydrophobic characteristics that preclude the use of standard chromatographic resins. The resulting purified disulfide-linked homodimer was demonstrated to bind to VEGF-R1 and to compete with VEGF-A for binding to this receptor. PMID- 11106178 TI - The three-dimensional structure of the ternary complex of Corynebacterium glutamicum diaminopimelate dehydrogenase-NADPH-L-2-amino-6-methylene-pimelate. AB - The three-dimensional (3D) structure of Corynebacterium glutamicum diaminopimelate D-dehydrogenase in a ternary complex with NADPH and L-2-amino-6 methylene-pimelate has been solved and refined to a resolution of 2.1 A. L-2 Amino-6-methylene-pimelate was recently synthesized and shown to be a potent competitive inhibitor (5 microM) vs. meso-diaminopimelate of the Bacillus sphaericus dehydrogenase (Sutherland et al., 1999). Diaminopimelate dehydrogenase catalyzes the reversible NADP+ -dependent oxidation of the D-amino acid stereocenter of mesodiaminopimelate, and is the only enzyme known to catalyze the oxidative deamination of a D-amino acid. The enzyme is involved in the biosynthesis of meso-diaminopimelate and L-lysine from L-aspartate, a biosynthetic pathway of considerable interest because it is essential for growth of certain bacteria. The dehydrogenase is found in a limited number of species of bacteria, as opposed to the alternative succinylase and acetylase pathways that are widely distributed in bacteria and plants. The structure of the ternary complex reported here provides a structural rationale for the nature and potency of the inhibition exhibited by the unsaturated L-2-amino-6-methylene-pimelate against the dehydrogenase. In particular, we compare the present structure with other structures containing either bound substrate, meso-diaminopimelate, or a conformationally restricted isoxazoline inhibitor. We have identified a significant interaction between the alpha-L-amino group of the unsaturated inhibitor and the indole ring of Trp144 that may account for the tight binding of this inhibitor. PMID- 11106177 TI - Dimer formation by a "monomeric" protein. AB - Dimeric proteins can arise by the swapping of structural domains between monomers. The prevalence of this occurrence is unknown. Ribonuclease A (RNase A) is assumed to be a monomer near physiological conditions. Here, this hypothesis is tested and found to be imprecise. The two histidine residues (His12 and His119) in the active site of RNase A arise from two domains (S-peptide and S protein) of the protein. The H12A and H119A variants have 10(5)-fold less ribonucleolytic activity than does the wild-type enzyme. Incubating a 1:1 mixture of the H12A and H119A variants at pH 6.5 and 65 degrees C results in a 10(3)-fold increase in ribonucleolytic activity. A large quantity of active dimer can be produced by lyophilizing a 1:1 mixture of the H12A and H119A variants from acetic acid. At pH 6.5 and 65 degrees C, the ribonucleolytic activity of this dimer converges to that of the dimer formed by simply incubating the monomers, as expected for a monomer-dimer equilibrium. The equilibrium dissociation constant for the dimer is near 2 mM at both 65 and 37 degrees C. This value of Kd is only 20-fold greater than the concentration of RNase A in the cow pancreas, suggesting that RNase A dimers exist in vivo. The intrinsic ability of RNase A to form dimers under physiological conditions is consistent with a detailed model for the evolution of homodimeric proteins. Dimers of "monomeric" proteins could be more prevalent than is usually appreciated. PMID- 11106179 TI - Pyramidalization of backbone carbonyl carbon atoms in proteins. AB - The high accuracy of X-ray analyses at atomic resolution is now able to display subtle deformations from standard geometry of building blocks in proteins. From the analysis of nine ultra-high resolution protein structures, we derived the first experimental evidence that a significant pyramidalization at the main-chain carbonyl carbon atom occurs in proteins. Our findings also show that this pyramidalization is related to the main-chain psi torsion angle. The carbonyl carbon atoms of residues that adopt alphaR and extended conformations show a clear preference for positive and negative pyramidalization, respectively. The agreement between our data and those previously obtained from small molecule structures demonstrates that carbon pyramidalization is an intrinsic property of the peptide structure. Although small in magnitude, the pyramidalization is well preserved in the complex folded state of a macromolecular structure that results from the interplay of many different forces. In addition, this property of the peptide group may have interesting implications for the enzymatic reactions involving the carbonyl carbon atoms. PMID- 11106180 TI - A logical sequence search for S100B target proteins. AB - The EF-hand calcium-binding protein S100B has been shown to interact in vitro in a calcium-sensitive manner with many substrates. These potential S100B target proteins have been screened for the preservation of a previously identified consensus sequence across species. The results were compared to known structural and in vitro properties of the proteins to rationalize choices for potential binding partners. Our approach uncovered four oligomeric proteins tubulin (alpha and beta), glial fibrillary acidic protein (GFAP), desmin, and vimentin that have conserved regions matching the consensus sequence. In the type III intermediate filament proteins (GFAP, vimentin, and desmin), this region corresponds to a portion of a coiled-coil (helix 2A), the structural element responsible for their assembly. In tubulin, the sequence matches correspond to regions of alpha and beta tubulin found at the alpha beta tubulin interface. In both cases, these consensus sequence matches provide a logical explanation for in vitro observations that S100B is able to inhibit oligomerization of these proteins. PMID- 11106182 TI - Oculoplastic surgery in cyberspace. PMID- 11106181 TI - Chemical modification of a variant of human MIP-1alpha; implications for dimer structure. AB - A sequence variant of human MIP-1alpha, in which Asp26 has been replaced by Al alpha, has been chemically modified by the addition of 13C-labeled methyl groups at each of the lysine residues and the N-terminus. The sites of methylation have been verified by a combination of MALDI-TOF mass spectrometric experiments and tryptic digestion followed by N-terminal mapping. The effect of the modification on the structure and activity of the protein have been determined by analytical ultra-centrifugation, 13C NMR spectroscopy and receptor binding studies. The results of these experiments suggest that huMIP-alpha D26A (BB10010), when present as a dimer, adopts a globular structure, like MCP-3, rather than the elongated or cylindrical structure determined for dimers of huMIP-1beta and RANTES. PMID- 11106183 TI - The 1999 Wendell Hughes lecture. Surgery, service, and soul. PMID- 11106184 TI - Electrosurgical modification of orbicularis oculi hypertrophy. AB - PURPOSE: To assess two electrosurgical approaches for the modification of orbicularis hypertrophy that may be used in conjunction with, or separate from, lower lid blepharoplasty. The hypothesis to be tested is that purely electrosurgical nonexcisional techniques may be used to modify orbicularis oculi muscle. METHODS: Electrosurgical techniques to treat orbicularis hypertrophy with an "open" and a "closed" technique are described. The open technique is performed in conjunction with transconjunctival blepharoplasty. The closed technique requires a 1-mm to 2-mm dermal incision, 2 minutes of surgical time per eyelid, and a specially insulated and formed electrosurgical needle. A review and case series are presented to illustrate and describe the techniques and results. RESULTS: Results for both techniques were rated by both patients and surgeons using the categories of poor, fair, good, or excellent. The open technique was performed in conjunction with transconjunctival blepharoplasty on 23 patients during 2 years with a minimum follow-up of 6 months. Results for the open technique were considered "excellent" by 14 patients and "good" by 9 patients. The operating surgeons evaluated the improvement as "excellent" in 4, "good" in 11, and "fair" in 8 patients. The closed technique was performed on eight patients. Results for patient satisfaction for the closed technique were considered "good" by 4, "excellent" by 2, "fair" by 1, and the final patient abstained from categorization. Operating surgeon evaluation of the closed technique revealed "excellent" outcomes in 3, "good" in 3, and "fair" for 2 patients. CONCLUSION: Electrosurgical techniques may be used to modify orbicularis hypertrophy. Drawbacks include a significant learning curve, potential cutaneous ulceration, and occasional temporary anatomic distortion as manifested by scleral show. Complications are minimal, and the technique was safe in all patients studied. PMID- 11106185 TI - Complications of laser resurfacing and their management. AB - PURPOSE: To describe complications associated with laser resurfacing along with specific treatment recommendations. METHODS: The authors' experiences with laser resurfacing complications are discussed in conjunction with a review of published reports. Current preoperative and postoperative regimens are also presented. RESULTS: Postoperative erythema occurs in all patients and is considered a transient side effect, not a complication. Postinflammatory hyperpigmentation, hypopigmentation, scarring, wound infections, milia, ectropion, pain, acneiform eruptions, pruritus, and contact dermatitis are reported by multiple authors. Specific interventions combined with the passage of time allow most of these complications to resolve, leaving the patient with an acceptable final result. CONCLUSIONS: Although laser resurfacing is a safe and effective method of facial rejuvenation, the cosmetic surgeon must be aware of the various complications that may be encountered. Prompt recognition of complications and appropriate management provide the best opportunity for an acceptable aesthetic outcome. PMID- 11106186 TI - The geometrical basis of the eyelid contour. AB - PURPOSE: To derive a two-dimensional, frontal-view model of eyelid contour. METHODS: Observational study. Palpebral fissure images of 110 normal subjects were acquired with a charge-coupled device camera and processed with National Institutes of Health Image software on a Macintosh computer. Monocular frontal view images of the palpebral fissures were recorded and second-degree polynomial functions were fitted to both upper and lower eyelid contours for two areas: the whole eyelid margin (ciliated and inner canthal portions) and the ciliated portion alone. In addition, frontal and lateral palpebral fissure images were obtained. From the frontal view, the upper and lower ciliated contours were fitted with quadratic functions. From the lateral view, the upper and lower lateral angles, formed by the upper and lower eyelid margins and the axial axis, were measured. RESULTS: Exclusion of the inner canthal portion of the eyelid contour led to a much better quadratic fit for the contours. The sine (sin) of the upper lateral angle was strongly correlated with the parameter A of the quadratic function fitted to the upper eyelid (the parameter A determines the curvature of the function around its extremum point). For the lower eyelid, this correlation was not significant. CONCLUSIONS: The parabolic shape of the upper ciliated contour seen in two-dimensional images can be justified geometrically in a simple way, allowing a precise quantification of its shape. The same was not true for the lower eyelid. The parabolic shape of the upper eyelid can be demonstrated, using the Taylor series, to be a close approximation of the arc of a circle. PMID- 11106187 TI - Burn scar malignancies of the eyelids. AB - PURPOSE: To study the clinicopathologic characteristics and treatment of eyelid carcinomas developing in thermal burn scars. METHODS: A review of eight cases of eyelid burn scar malignancies: two from our own experience and six from published reports. RESULTS: Reported cases of burn scar malignancy of the eyelid are short latency basal cell carcinomas. All carcinomas arose from small superficial burns. These potentially aggressive tumors respond well to local excision. CONCLUSION: As with other areas of the body, eyelid burn scars may undergo neoplastic degeneration. These carcinomas are predominately short latency basal cell carcinomas, rather than long-latency squamous cell carcinomas that are more common elsewhere in the body, including the head and neck region. Clinicians should be diligent in the long-term surveillance of all eyelid burns. PMID- 11106188 TI - Surgical resection of giant papillae and autologous conjunctival graft in patients with severe vernal keratoconjunctivitis and giant papillae. AB - PURPOSE: Giant papillae (GP) in patients with vernal keratoconjunctivitis (VKC) refractory to clinical treatment may cause serious corneal complications, such as shield ulcer. We propose a surgical treatment--resection of GP--in conjunction with free autologous conjunctival graft to treat severe cases of VKC with GP. METHODS: Six eyes of five patients with VKC, characterized by GP and shield ulcer refractory to clinical treatment, underwent surgical resection of GP associated with free autologous conjunctival graft. RESULTS: No recurrence of GP over the graft was observed during follow-up intervals ranging from 9 months to 27 months. Corneal shield ulcers healed during the first week after treatment and did not recur. CONCLUSIONS: Patients with refractory VKC and GP associated with corneal shield ulcer may benefit from resection of GP and autologous conjunctival graft. PMID- 11106189 TI - Relation between p53 overexpression and clinical behavior of ocular/orbital invasion of conjunctival squamous cell carcinoma. AB - PURPOSE: To investigate the p53 gene as a prognostic indicator in conjunctival squamous cell carcinoma. METHODS: Medical records were reviewed and histopathology slides were examined to verify the diagnosis and grade of tumor differentiation in a retrospective case series of 12 patients with "invasive" and 11 patients with "noninvasive" conjunctival squamous cell carcinoma. The p53 antigen was detected using the streptavidin biotin-alkaline phosphatase immunostaining method. Statistical analysis was performed using the Fisher exact test. RESULTS: p53 overexpression was present in 14 cases (approximately 60%). Eight and 6 of 14 p53-positive tumors were invasive and noninvasive, respectively. All patients with recurrence (two), recurrence and metastasis (two), metastasis (three), and death with tumor dissemination (three) had p53 overexpression. Of all adverse outcomes, only two cases with recurrence were p53 negative: no metastasis or death with dissemination was seen in p53-negative tumor cases. CONCLUSIONS: There was no statistical relationship between p53 overexpression and age, sex, location, invasiveness, or histopathologic differentiation of the tumor; however, a significant association existed between p53 positivity and adverse clinical behavior (p = 0.014). PMID- 11106190 TI - Squamous cell carcinoma with perineural invasion presenting as a Tolosa-Hunt-like syndrome: a potential pitfall in diagnosis. AB - PURPOSE: To describe a case of perineural invasion resulting from squamous cell carcinoma of forehead. METHODS: Case report. RESULTS: Perineural invasion resulting from squamous cell carcinoma of the periocular skin can present as a Tolosa-Hunt-like syndrome with lack of radiologic findings on magnetic resonance imaging (MRI) in its early stages. CONCLUSION: A high level of suspicion for perineural invasion is required when assessing multiple cranial nerve palsies in patients with a history of cutaneous malignancy, despite negative sequential MRI. Perineural invasion must be ruled out by a biopsy of the involved nerves, whenever possible, before empiric therapy with systemic steroids is contemplated. PMID- 11106191 TI - Merkel cell carcinoma: clinicopathologic correlation, management, and follow-up in five patients. AB - PURPOSE: To review the clinicopathologic features, management and follow-up in five patients with periocular Merkel cell carcinoma (MCC). METHODS: In this case series study we reviewed the clinical records and histopathologic findings of five consecutive patients with MCC, treated and followed between May 1991 and November 1998. RESULTS: Four patients were female and one was male. Their mean age at the time of surgery was 80 years (range: 69-86 years). Patients presented with a painless, nonulcerated, rapidly growing, solitary reddish or violaceous nodule, ranging in size from 11 to 21 mm, located in the upper eyelid in 4 patients and in the left eyebrow in one patient. Management included excision with frozen section control in three patients, and excision with wide surgical margins in two patients. Histopathologically, all tumors exhibited round cells of intermediate size and scanty cytoplasm, and large, round to oval vesicular nuclei with finely dispersed chromatin and one to three inconspicuous nucleoli. Mitotic figures were numerous. No patient had regional or extraregional metastases at diagnosis. Local recurrence or distant metastases were not detectable during a mean follow-up period of 45 months (range: 32-61 months). This outcome contrasts with the high incidence of local recurrence and early nodal metastasis reported in previous publications. CONCLUSIONS: Early diagnosis and aggressive, histologically controlled surgical treatment of this rare, highly malignant tumor may provide a longer disease-free period. Further investigation is necessary to determine the prognostic factors for recurrence and survival. PMID- 11106192 TI - Collagen vascular diseases: cutaneous manifestations in ophthalmology. AB - PURPOSE: To describe the effects of collagen vascular diseases on the eyelids and periorbital tissues. METHODS: Retrospective review of dermatologic pathology slides at Massachusetts General Hospital and eye pathology slides at Massachusetts Eye and Ear Infirmary, Boston, Massachusetts. RESULTS: A spectrum of dermatologic manifestations of collagen vascular diseases was observed, affecting the eyelids and periorbital region. CONCLUSIONS: Collagen vascular diseases may present complicated diagnostic and clinical challenges for the practicing ophthalmologist. Familiarity with the cutaneous periocular manifestations of these diseases may facilitate early recognition, diagnosis, and ophthalmologic intervention where necessary. PMID- 11106193 TI - Lacrimal canalicular diverticulum: a cause of epiphora and discharge. AB - PURPOSE: To report a rare cause of epiphora and chronic discharge: lacrimal canalicular diverticulum. METHODS: Case report. RESULTS: A distended left lower eyelid canaliculus was associated with chronic epiphora and discharge. A suspected canalicular diverticulum was noted on dacryocystogram. CONCLUSIONS: Canalicular diverticula should be considered in the differential diagnosis in patients that present with epiphora and discharge with a patent nasolacrimal system. Dacryocystography is diagnostic. PMID- 11106194 TI - Standards for infant respiratory function testing: what(ever) next? PMID- 11106195 TI - Improving standards of clinical care in cystic fibrosis. PMID- 11106196 TI - Urinary excretion of leukotriene E4 and eosinophil protein X in children with atopic asthma. AB - Measurement of leukotriene E4 (LTE4) in urine is a noninvasive method for assessing changes in the rate of total body cysteinyl leukotriene production. Eosinophil protein X (EPX) has been used to assess eosinophil activity and monitor inflammation in bronchial asthma. The aim of the study was to look for differences in urinary LTE4 and EPX concentrations between children with stable atopic asthma and healthy controls and to compare asthmatic children with different disease severity. In addition the relationship was evaluated between urinary LTE4 and EPX levels and lung function. LTE4 was also measured (enzyme immunoassay) together with EPX (radioimmunoassay) in urine and lung function tests were carried out in children with mild asthma (steroid-naive) (n=49), moderate to severe asthma (using inhaled steroids) (n=31) and healthy control subjects (n=28). Urinary leukotriene E4 (LTE4) was significantly higher in children with asthma than in controls (median [25-75 percentile] 238.5 (126.5 375.7) SD 191.8 versus 189 (51-253.2) SD 131.7 pg.mg(-1) creatinine; p=0.021). Urinary EPX was also significantly increased in asthmatic children compared with controls (85.5 [64-131.5] SD 76.2 versus 48.5 [43.2-90] 112.1 microg x mmol(-1) creatinine; p=0.006). There were no differences in urinary LTE4 and EPX between the group of mild and the group of moderate to severe asthmatic children. There were significant associations between the urinary LTE4 and intrathoracic gas volume (ITGV), residual volume (RV), forced expiratory volume in one second (FEV1), forced expiratory capacity (FVC) and maximum expiratory flow rate at 25% of vital capacity (MEF25). Urinary EPX was only correlated with maximum expiratory flow rate at 75% of vital capacity (MEF75). Thus measurement of urinary LTE4 may predict the degree of airflow obstruction in asthmatic children. Urinary LTE4 and EPX are useful markers of airway inflammation and can be helpful in guiding asthma management. There was no correlation between LTE4 and EPX levels. PMID- 11106197 TI - When a "wheeze" is not a wheeze: acoustic analysis of breath sounds in infants. AB - Epidemiological studies indicate that the prevalence of "wheeze" is very high in early childhood. However, it is clear that parents and clinicians frequently use the term "wheeze" for a range of audible respiratory noises. The commonest audible sounds originating from the lower airways in infancy are ruttles, which differ from classical wheeze in that the sound is much lower in pitch, with a continuous rattling quality and lacking any musical features. The aim of this study was to clearly differentiate wheeze and ruttles objectively using acoustic analysis. Lung sounds were recorded in 15 infants, seven with wheeze and eight with ruttles, using a small sensitive piezoelectric accelerometer, and information relating to the respiratory cycle was obtained using inductive plethysmography. The acoustic signals were analysed using a fast fourier transformation technique (Respiratory Acoustics Laboratory Environment programme). The acoustic properties of the two noises were shown to be quite distinct, the classical wheeze being characterized by a sinusoidal waveform with one or more distinct peaks in the power spectrum display; the ruttle is represented by an irregular nonsinusoidal waveform with diffuse peaks in the power spectrum and with increased sound intensity at a frequency of <600 Hz. It is important for clinicians and epidemiologists to recognize that there are distinct types of audible respiratory noise in early life with characteristic acoustic properties. PMID- 11106198 TI - Lung function measured by the oscillometric method in prematurely born children with chronic lung disease. AB - Premature birth is related to a chronic respiratory morbidity, which may persist until school-age. In these children, the forced oscillation technique would be suitable for evaluation of lung function even at preschool age, since it requires only minimal patient cooperation. In order to investigate the oscillometric findings related to premature birth, using the oscillation technique and conventional lung function methods 49 school-aged children born prematurely with (n=15) or without (n=34) chronic lung disease (CLD), and 18 healthy children born at full term were studied. Children with CLD had higher respiratory resistance (Rrs,5) and lower reactance (Xrs,5) than prematurely born children without CLD or healthy controls. Both Rrs,5 (r=-0.55, p<0.0001) and Xrs,5 (r=0.76, p<0.0001) were significantly associated with forced expiratory volume in one second (FEV1), the agreement with spirometry being better in Xrs,5 than in Rrs,5 (p=0.02). Rrs,5 was significantly related to airway resistance (Raw) measured by body plethysmography (r=0.63, p<0.0001), but underestimated resistance at high values of Raw. There was no significant relationship between the pulmonary diffusing capacity and the oscillometric findings. Compared to conventional methods, the oscillometric method yields concordant information on the severity of lung function deficit in children born prematurely, with or without chronic lung disease. In these children, the oscillometric findings are probably due to peripheral or more widespread airway obstruction. As conventional methods are not usually suitable for preschool children, oscillometry may serve as an alternative for early evaluation of chronic lung disease among children with premature birth in clinical or research settings. PMID- 11106199 TI - Lack of association between adult asthma and the tumour necrosis factor alpha-308 polymorphism gene. AB - Tumour necrosis factor (TNF)alpha is a cytokine endowed with potent inflammatory properties that may contribute to airway inflammation in asthma. It has previously been shown that the single base pair polymorphism-308 (G to A substitution) in the promoter of TNFalpha gene results in enhanced cytokine secretion. Whether this polymorphism is associated with the presence of phenotypic expression of asthma is questioned. In this study the relative frequency of TNF1 and TNF2 alleles in a population of adult healthy subjects (n=98) and adult Caucasian asthmatics (n=95) was compared taking into account their disease severity, atopic status and their smoking habit. For the whole group of asthma the genotype frequency for 1/1, 1/2, 2/2 were 67%, 33% and 0%, respectively, and not significantly different from those found in the control group that reached 70%, 28% and 2% respectively (p>0.05). The allele frequencies in asthma were 86% and 14% for TNF1 and TNF2 respectively while the corresponding figures were 85% and 15% in the control group (p>0.05). Furthermore, subdividing asthmatics into severe forced expiratory volume in one second <60% pred), atopic or smoking patients did not show any significant association with this TNFalpha polymorphism. To conclude the polymorphism -308 in the promoter of the TNFalpha gene does not confer a susceptibility to develop asthma nor to grade its severity. PMID- 11106200 TI - Elevation of total serum immunoglobulin E is associated with asthma in nonallergic individuals. AB - Elevated serum immunoglobulin (Ig)E is the hallmark of atopy, and contributes to asthma and bronchial hyperresponsiveness in atopic individuals. In contrast, the significance of IgE in nonallergic subjects is less clear. The aim of the present study is to clarify a potential association of IgE and asthma in absence of clinical allergy. To this purpose 1,219 consecutive patients of a pulmonary practice were evaluated. Nonallergic patients were defined by negative skin prick test, history of atopy and specific IgE, 509 subjects (42%) were nonallergic. Among these, 80 patients (16%) had elevated total IgE levels (>150 U x mL(-1)). Prevalence and severity of asthma in nonallergic subjects with IgE>150 U x mL(-1) were compared with subjects with normal IgE levels, and lung function parameters were correlated with serum IgE in all nonallergic subjects and asthmatics. Asthma was more prevalent in nonallergic subjects with elevated IgE levels than in nonallergic subjects with normal IgE (39% versus 14%; p<0.001). Lung function values of nonallergic asthmatics were lower for forced expiratory volume in one second (FEV1)% predicted (66+20% versus 83+/-17%; p<0.001), FEV1% forced vital capacity (FVC) (70+/-14% versus 81+/-8%; p<0.001) and forced mid expiratory flow (FEF25-75) (1.7+/-0.9 L x s(-1) versus 2.8+/-0.9 L x s(-1); p=0.002) in patients with high IgE compared to asthmatics with normal IgE, and were negatively correlated with log IgE levels in all nonallergic asthmatics. (FEVI % pred: r= 0.5, p<0.001; FEV1% FVC: r=-0.53, p<0.001; FEF25-75: r=-0.52, p<0.001). In the whole study population, multivariate analysis showed a greater than fivefold asthma risk for nonallergic individuals with serum IgE>150 U x mL(-1). These data support the role of IgE as risk factor for asthma independent of allergy, and they further challenge the definition of intrinsic asthma as "non-IgE mediated" entity. PMID- 11106201 TI - Using Global Initiative for Asthma guidelines to assess asthma severity in populations. AB - The classification of asthmatics into severity categories is a crucial issue for assessing the asthma burden within a community, in which a proportion of patients is currently treated. There is no epidemiological method currently available. The Global Initiative for Asthma (GINA) was used to classify 4,362 patients aged 16 45 yrs (49% males, 42% taking inhaled corticosteroids), enrolled by 545 chest specialists in France with short standardized questionnaires including forced expiratory volume in one second (FEV1) measurements. Two independent GINA classifications were combined, one based only on symptoms and FEVI, and the other based only on current medication, to construct a final "symptom-FEV1 medication" classification. Almost 40% of the patients classed as step 1, 30% of those classed as step 2 and 13% of those classed as step 3 in the initial symptom-FEV1 classification, were allocated to categories of higher severity in the final classification. The approach was validated by showing that the proportions of: 1) patients considered by the physicians as having severe or moderately severe asthma; 2) patients with a history of hospital admission for asthma; and 3) patients with a history of emergency department visits for asthma, increased with severity steps in the final classification, for each step of the two initial independent classifications. The treatment manage plan in the Global Initiative for Asthma was not developed for assessing severity of asthma but rather to describe the recommended therapy for asthma with different severity. This is the first attempt to assess the severity of asthma in a large population of asthmatics mostly taking treatment, based on the Global Initiative for Asthma guidelines. The authors propose this simple and pragmatic procedure for a potential classification which should be put to the test in other studies. PMID- 11106202 TI - Magnesium levels in plasma and erythrocytes before and after histamine challenge. AB - Previous studies have assessed the protective effect of nebulized magnesium sulphate on bronchial hyperreactivity. This study investigated the effect of histamine challenge on intracellular (erythrocytes) and extracellular (plasma) levels of magnesium and the possible relationship between degree of bronchial hyperreactivity and levels of Mg in plasma and erythrocytes. The authors studied 42 mildly asthmatic patients (10 on inhaled steroids) and 20 healthy subjects. Histamine challenge was performed by the dosimeter method and provocative dose causing a 20% fall in forced expiratory volume in one second (PD20) (FEV1) was calculated. Mg levels were measured with a calmagite colourimetric assay, both at baseline and when FEV1 had fallen by 20%. The results showed that Mg levels in plasma did not significantly change after histamine challenge (from 2.06+/-0.02 mg x dL(-1) to 2.08+/-0.02 mg x dL(-1) respectively, p=0.14). Conversely there was a statistically significant decrease in Mg levels in erythrocytes between these two time points (from 1.84+/-0.02 fmmol x cell to 1.78+/-0.02 fmmol x cell p<0.0001). Similar results were observed when the subgroups were studied separately. There was no significant correlation between PD20, the difference in both magnesium concentrations (baseline-PD20 time) or the initial values of Mg levels in erythrocytes and plasma. To conclude, histamine challenge reduces magnesium levels in erythrocytes while plasma levels remain unchanged. This histamine-induced decrease in magnesium levels occurs regardless of the diagnosis of asthma, and it is not correlated with the degree of bronchial hyperreactivity. PMID- 11106203 TI - Nasal inflammatory and respiratory parameters in human volunteers during and after repeated exposure to chlorine. AB - The objectives of this study were: 1) to determine if chlorine exposure at low levels induces nasal effects in humans as it does in rodents; and 2) to establish a possible occurrence of respiratory effects in human volunteers exposed to chlorine vapour at concentrations of 0, 0.1, 0.3 and 0.5 ppm. The study was conducted in a double-blind fashion in 8 male volunteers using a repeated measures design, with randomly selected exposure sequences. Subjects were exposed for 6 h x day(-1) on 3 consecutive days to each of the 4 exposure conditions. In nasal lavage, interleukin-8 (IL-8), albumin, total cell number, and percentages of neutrophils, lymphocytes, monocytes, eosinophils, and epithelial cells were determined. The lung function parameters that were analysed included forced vital capacity (FVC), forced expiratory volume in first second (FEV1), FEV1/FVC ratio, and maximal mid expiratory flow (MMEF). Data analysis was limited to 7 subjects since one volunteer decided to stop participating for reasons not related to the study. Nasal lavage measurements did not support an inflammatory response or irritant effects on the nasal epithelium. For FVC, FEV1, and FEV1/FVC, no significant differences were found. MMEF was significantly different between the 0 and 0.5 ppm exposure, but this was attributed to an unexplained shift in baseline values during control (0 ppm) exposure. The present data does not support an inflammatory effect in the nose nor shows changes in respiratory function at repeated exposure up to 0.5 ppm. This discrepancy with previous data in rodents can be attributed at least in part to differences in respiratory tract airflow characteristics. PMID- 11106204 TI - Chronic cough and gastro-oesophageal reflux: a double-blind placebo-controlled study with omeprazole. AB - Gastro-oesophageal reflux (GOR) is an important cause of chronic cough. There has been a lack of placebo-controlled trials treating GOR related chronic cough with antireflux therapy. The aim of this study was to determine the efficacy of omeprazole on GOR related chronic cough. After excluding other common causes of cough, oesophageal pH monitoring was performed on 48 patients with chronic cough. Twenty-nine patients found to have GOR were randomized in a double-blind fashion to receive omeprazole 40 mg o.d. or placebo for 8 weeks. After a 2-week washout period, patients were crossed over to the other treatment. Symptoms were recorded daily in a diary. Twenty-one patients completed both treatment periods. Cough (p=0.02) and gastric symptoms (p=0.003) improved significantly during the omeprazole treatment in twelve patients who received placebo during the first and omeprazole during the second 8-week period. In nine patients who received omeprazole during the first 8-week period, amelioration in cough reached statistical significance only after cessation of omeprazole. Gastric symptoms also remained minor during placebo in these nine patients. Omeprazole 40 mg o.d. seems to improve chronic cough in patients with gastrooesophageal reflux and the effect of omeprazole in ameliorating both cough and reflux symptoms continues after treatment ceases. PMID- 11106205 TI - Nasal obstruction as a risk factor for sleep apnoea syndrome. AB - Nasal obstruction has frequently been mentioned as a possible risk factor in obstructive sleep apnoea syndrome (OSAS). Over a 2-yr period, 541 unselected consecutive snorers referred for suspected breathing disorders during sleep were included to undergo posterior rhinomanometry. In addition cephalometric landmarks and body mass index (BMI) were obtained. Polysomnography was used to determine the number of abnormal respiratory events that occurred during sleep. OSAS was defined as 15 episodes, or more, of apnoea or hypopnoea per hour of sleep (AHI). Of the 541 consecutive snorers 528 underwent nasal resistance measurement by posterior rhinomanometry (failure rate: 2.4%). Patients with OSAS (259 patients) had higher nasal resistance than patients without OSAS (2.6+/-1.6 hPa x L x s(-1) versus 2.2+/-1.0 hPa x L x s(-1), respectively, p<0.005). A stepwise multiple regression analysis showed that BMI, male sex, nasal resistance, and cephalometric parameters were contributing factors to the AHI. The r2-value of the multiple regression analysis was 0.183. Nasal resistance contributed 2.3% of the variance (p<0.0001), whereas mandibular plane-hyoid distance, BMI, male sex and age contributed 6.2%, 4.6%, 3% and 1.3% of the variance, respectively. To conclude, daytime nasal obstruction is an independent risk factor for OSAS. PMID- 11106206 TI - Abnormal lipid peroxidation in patients with sleep apnoea. AB - The prevalence of cardiovascular diseases is increased in patients with the obstructive sleep apnoea syndrome (OSAS). The fall and rise of arterial oxygenation that follows each apnoea may increase lipid peroxidation and contributes to explaining this association. In the present study, the authors determined lipid peroxidation in patients with OSAS and the effect of treatment with continuous positive airway pressure (CPAP). Fourteen male patients with severe OSAS (59+/-5 apnoea x h(-1)) (+/-SEM) and 13 healthy nonsmoking, male volunteers of similar age were studied. Patients were studied at diagnosis and after treatment with CPAP for more than 1 yr (>4 h x night(-1)). A venous blood sample was obtained early in the morning after fasting all night. In patients with OSAS, a sample before and during sleep was also obtained. Low density lipoprotein (LDL) particles were isolated by sequential ultracentrifugation. Their level of oxidation was determined by the thiobarbituric acid assay (TBARs), and their susceptibility to oxidation by the lag phase measurement. Patients with OSAS showed higher TBARs (28.1+/-2.8 versus 20.0+/-1.8 nmol x malondialdehyde x mgLDL protein(-1), p=0.02) and shorter lag phase values (83.8+/-3.4 versus 99.7+/ 3.4 min, p=0.005) than controls. These differences were not due to the smoking status of the patient. Likewise, these values did not change significantly throughout the night yet, the lag phase value was significantly improved by treatment with CPAP (124.9+/-8.5 min; p<0.001). These results indicate that obstructive sleep apnoea syndrome is associated with abnormal lipid peroxidation and that this is improved by chronic use of Continuous positive airway pressure. These results can contribute towards explaining the high prevalence of cardiovascular diseases seen in Obstructive sleep apnoea syndrome. PMID- 11106207 TI - Platelet function in patients with obstructive sleep apnoea syndrome. AB - Patients with obstructive sleep apnoea syndrome (OSAS) are subject to an increased cardiovascular morbidity including myocardial infarction and stroke. Platelets play an important role in the pathogenesis and triggering of acute cardiovascular syndromes. So far, the influence of OSAS on platelet function is not fully understood. Platelet aggregability to epinephrine, collagen, arachidonic acid, and adenosine diphosphate in vitro was measured in 17 consecutive male patients (53.0+/-2.1 yrs) with polysomnographically verified OSAS and compared with that of 15 male controls (50.1+/-3.6 yrs) at 20:00 h, 24:00 h, and 06:00 h. In addition, the long-term effects of continuous positive airway pressure (CPAP) therapy on platelet aggregability was assessed after 6 months. Platelet aggregation in vitro induced by epinephrine showed a slight increase overnight in the untreated OSAS patients (NS) whereas it decreased slightly (NS) in the controls and in the treated OSAS patients. Pretherapeutic platelet aggregability was significantly lowered by CPAP therapy both at 24:00 h (64.0+/-6.5 versus 55.3+/-6.7%, p<0.05) and at 06:00 h (64.1+/-6.5 versus 45.8+/ 7.6%; p=0.01). Platelet aggregability during sleep in the controls resembled that found in patients with OSAS during CPAP therapy. The results suggest that obstructive sleep apnoea syndrome contributes, at least in part, to platelet dysfunction and that long-term continuous positive airway pressure treatment may reduce platelet aggregability. PMID- 11106208 TI - Influence of sampling interval on the evaluation of nocturnal blood pressure in subjects with and without obstructive sleep apnoea. AB - Blood pressure (BP) variability during sleep is high in obstructive sleep apnoea syndrome (OSAS). How BP sampling interval affects the estimate of mean nocturnal BP in OSAS and control subjects was investigated. Nine subjects with apnoea/hypopnoea index (AHI) <5 and 18 OSAS patients with AHI >30 underwent nocturnal polysomnography with beat-by-beat BP monitoring. Mean nocturnal BP was evaluated averaging: a) all systolic (Ps) and diastolic (Pd) BP values; b) Ps and Pd sampled every 5, 10, 15, 20, and 30 min. The sampling starting point was repeatedly shifted, and several mean BP estimates for each sampling interval were obtained. Differences (deltaPs and deltaPd) between means obtained by sampling BP and by averaging all BP values were calculated. In both groups deltaPs and deltaPd scatter increased as sampling interval increased; their variance was always higher in OSAS subjects (p<0.001). Over 95% of deltaPs and deltaPd were <5% of the beat-by-beat mean values at all sampling intervals in controls, but this occurred only at sampling intervals < or =10 min in OSAS subjects. To conclude, for each blood pressure sampling time, a larger number of inaccurate nocturnal mean blood pressure estimates are obtained in obstructive sleep apnoea syndrome than in control subjects. Obstructive sleep apnoea syndrome subjects require more frequent blood pressure measurements to obtain a similar accuracy in nocturnal blood pressure evaluation. PMID- 11106209 TI - Partitioning of dead space--a method and reference values in the awake human. AB - Although dead space is often increased in disease, it is not frequently measured in the clinic. This may reflect that an adequate method as well as reference values are missing. Healthy males and females, n=38, age 20-61 yrs, were connected to a pneumotachograph and a fast CO2 analyser after radial artery catheterization. The physiological dead space was partitioned into airway and alveolar dead space using a delineation principle denoted the pre-interface expirate. Physiological dead space was 201+/-41 mL in males and 150+/-34 mL in females. Dead space values were depending upon parameters reflecting lung size (predicted total lung capacity), breathing pattern and age. After multiple correlation the variation decreased and differences between males and females disappeared. The residual SD was then for physiological dead space 18.9 mL. The clinical use of the new method for determination of dead space can be based upon reference values, with a more narrow range than previous data. PMID- 11106210 TI - Volume and time dependence of respiratory system mechanics in normal anaesthetized paralysed humans. AB - The purpose of the present investigation was to assess the effect of large tidal volumes and mean lung volumes on the viscoelastic properties of the respiratory system in normal humans; and to verify if in this case the results could be satisfactorily described by a simple linear viscoelastic model of the respiratory system. Twenty-eight subjects (7 females), aged 14-28 yrs, were studied before orthopaedic surgery on the lower limbs. None were obese, or had clinical evidence of cardiopulmonary disease. The interrupter conductance and the viscoelastic constants of the respiratory system were assessed using the rapid end-inspiratory airway occlusion method during mechanical ventilation with tidal volumes up to 3 L and applied end-expiratory pressures up to 23 cmH2O. It was found that the interrupter conductance increased linearly with lung volume over a larger range than used previously; and the viscoelastic resistance and time constant did not change over the entire range of tidal volumes and end-expiratory pressures studied. In conclusion, in normal anaesthetized, paralysed subjects a simple linear viscoelastic model satisfactorily described the viscoelastic behaviour of the respiratory system over the whole range of volume studied. PMID- 11106211 TI - Can peak expiratory flow be measured accurately during a forced vital capacity manoeuvre? AB - Spirometry and peak flow measurements traditionally depend on different forced expiratory manoeuvres and have usually been performed on separate, dedicated equipment. As spirometry becomes more widely used in primary care settings, the authors wished to determine whether there was a systematic difference between peak expiratory flow (PEF) derived from a short sharp exhalation (PEF manoeuvre) and from a full forced vital capacity (FVC) manoeuvre, using the same turbine spirometer (Microloop, Micro Medical, Kent, UK). Eighty children (38 with current asthma) aged 7-16 yrs were asked to perform 2 blocks of PEF and FVC manoeuvres, the order being randomly assigned. PEF obtained from a peak flow manoeuvre (PEFPF) was significantly greater than that from a forced vital capacity manoeuvre (PEFVC) in both healthy (group mean difference 20 L x min(-1); p<0.001) and asthmatic children (group mean difference 9 L.min(-1); p<0.004). For clinical purposes, a mean difference of about 3% for children with asthma is of no practical significance, and peak expiratory flow data can usefully be obtained during spirometric recordings. PMID- 11106212 TI - Detection of rhinovirus in induced sputum at exacerbation of chronic obstructive pulmonary disease. AB - Common colds are associated with exacerbations of chronic obstructive pulmonary disease (COPD). However, the role of the common cold virus (human rhinovirus) in the production of symptoms and lower airway inflammation at COPD exacerbation is unknown. Thirty three patients with moderate-to-severe COPD were seen at baseline, when the number of chest infections in the previous year was noted, and acutely at COPD exacerbation. Within 48 h after the onset of the exacerbation and at baseline, nasal aspirates and induced sputum were taken for rhinovirus reverse transcriptase polymerase chain reaction (RT-PCR) analysis and determination of cytokine levels. Symptoms, recorded on diary cards, were noted and forced expiratory volume in one second (FEV1) and forced vital capacity (FVC) measured. At exacerbation, mean FEV1 and FVC fell significantly from baseline (p<0.001). Ten of 43 exacerbations were associated with rhinovirus infection, detected in induced sputum. In four of these, nasopharyngeal samples contained no detectable rhinovirus. All baseline samples were negative for rhinovirus. The simultaneous presence of increased nasal discharge/nasal congestion (in 26 of the 43 exacerbations) and increased sputum (29 exacerbations) was strongly associated with the presence of rhinovirus (odds ratio 6.15; p=0.036). Total symptom scores were greater for rhinovirus as compared to nonrhinovirus exacerbations (p=0.039). Median baseline sputum interleukin-6 levels rose from 90.2 to 140.3 pg x mL(-1) at exacerbation (p=0.005); the change was greater in the presence of rhinovirus infection (p=0.008). Rhinovirus infection can be detected at chronic obstructive pulmonary disease exacerbation. This is associated with elevation of lower airway interleukin-6 levels, which may mediate lower airway symptom expression during chronic obstructive pulmonary disease exacerbations. PMID- 11106213 TI - Expiratory muscle pressure and breathing mechanics in chronic obstructive pulmonary disease. AB - Expiratory muscle recruitment is common in stable chronic obstructive pulmonary disease (COPD) patients. Due to airway obstruction, there is little reason to believe that active expiration in COPD would be mechanically effective in lowering operating lung volume. The physiological significance of expiratory muscle recruitment in COPD, therefore, remains unknown. The purpose of this study was to assess, in COPD patients breathing at rest, the effect of expiratory muscle contraction on force generating ability of the diaphragm. The force generating ability of the diaphragm was evaluated from its pressure swing (Pdi) for a given diaphragm electrical activity (Edi), where Edi was normalized as % of its maximal value (Pdi/Edi/Edi,max). Phasic expiratory muscle contraction was measured as the total expiratory rise in gastric pressure (Pga,exp.rise). Nineteen seated patients with moderate to severe COPD, participated in the study and 10 exhibited phasic rise in Pga during expiration with a mean Pga,exp.rise of 1.91+/-0.89 cmH2O. The patients were thus divided into passive expiration (PE) and active expiration (AE) groups. There was no significant difference in various lung function and breathing pattern parameters between the two groups. Pdi/Edi/Edi,max was 0.63+/-0.07 and 0.54+/-0.07 cmH2O/% in PE and AE groups, respectively, and was not significantly different between each other. Compared with PE group, AE group not only recruited expiratory muscles, but also preferentially recruited inspiratory rib cage muscles and derecruited the diaphragm. The results do not support a significant improvement of the force generating ability of the diaphragm by phasic contraction of expiratory muscles at rest in chronic obstructive pulmonary disease patients. PMID- 11106214 TI - Up-regulation of circulating adhesion molecules in bronchiectasis. AB - Adhesion molecules are expressed on the surface of endothelial cells and leukocytes and are responsible for mediating the migration of intravascular leukocytes into inflamed tissue. Intensive recruitment of neutrophils into the airways occurs in bronchiectasis, although little is known about the role of adhesion molecules in this process. The authors, therefore, determined serum levels of E-selectin, intercellular adhesion molecule (ICAM)-1 and vascular adhesion molecule (VCAM)-1 in stable bronchiectasis patients (n=37) and healthy control subjects (n=17), and evaluated their relationship with clinical markers of disease severity in bronchiectasis. Serum levels of E-selectin, ICAM-1 and VCAM-1 in bronchiectasis patients were significantly higher than those in control subjects (p=0.02, <0.0001 and 0.0002 respectively). Both E-selectin and ICAM-1 levels were inversely related to forced expiratory volume in one second (FEV1)% predicted (r=-0.57, p<0.001; and r=-0.53, p=0.001 respectively), and FVC% predicted (r=-0.52, p=0.002; and r=-0.46, p=0.005). This was not the case for VCAM-1 levels. There was a correlation between serum ICAM-1 levels and 24 h sputum volume (r=0.34, p= 0.04). Serum E-selectin and ICAM-1, but not VCAM-1, levels showed correlation with the number of lung lobes affected by bronchiectasis (r=0.35, p=0.04 and r=0.34, p=0.04 respectively). These original observations strongly suggest that E-selectin, intercellular adhesion molecule-1 and Vascular adhesion molecule-1 could play a significant role in the pathogenesis of bronchiectasis. PMID- 11106215 TI - Modulation by pentobarbital of neutrophil responses to inhaled E. coli endotoxin in sheep: role of lung epithelium. AB - Neutrophils (PMNs) are implicated in the pathogenesis of acute respiratory distress syndrome (ARDS). The role of the epithelium in the modulation of PMN migration within the lungs was examined. Epithelial integrity and PMN concentrations in the lung air spaces and lymph were measured in sheep anaesthetized with either halothane (1-2.5%) or intravenous pentobarbital (12+/-4 mg x kg(-1) x h(-1)). Ventilation with an aerosol containing 25 mg Escherichia Coli endotoxin (lipopolysaccharide; LPS) effected neutrophil recruitment to the air spaces. Lymphatic clearance of aerosolized 99mTc-DTPA provided an index of epithelial integrity. Three hours after the deposition of LPS, the lung lining fluid of sheep anaesthetized with halothane (n=7) had 4.9+/-3.2x10(6) PMN.mL(-1), but the lung lymph had almost no PMNs (3+/-8%). Sheep anaesthetized with pentobarbital (n=6) had fewer PMNs in the air spaces (2.4+/-1.2X10(6) mL(-1)) and more PMNs in the lung lymph (30+/-20%). Control sheep (n=5) that received no LPS had almost no PMNs in the airspaces or lung lymph, regardless of the anaesthesia. Three additional sheep that remained awake after receiving LPS also had no PMNs in the lung lymph. The PMN fraction in the lung lymph correlated well with the extra-alveolar epithelial permeability measured by lymphatic clearance of aerosolized diethylenetriamine penta-acetic acid (r=0.81, p<0.001). Aerosolized lipopolysaccharide recruits neutrophils into the lungs of sheep, but they appear to remain in the airspaces unless extra-alveolar permeability is increased by agents such as pentobarbital. PMID- 11106216 TI - Evaluation of a method for assessing respiratory mechanics during noninvasive ventilation. AB - Noninvasive assessment of respiratory resistance (Rrs) and elastance (Ers), which is not easy with conventional methods, could be useful in the optimization of pressure support ventilation. The aim of this study was to evaluate a simple noninvasive method (Delta-inst) of measuring Rrs during nasal pressure support ventilation. Rrs and Ers (Delta-inst) were computed from inspiratory mask pressure, flow and volume recorded during pressure support ventilation. The Delta inst method was compared with the forced oscillation technique (FOT) in seven patients with chronic obstructive pulmonary disease (COPD) and in eight healthy subjects without and with added resistance (3.1 cmH2O x s x L(-1)). Rrs measured by Delta-inst (5.2+/-1.7, 7.2+/-0.5 and 6.9+/-1.2 cmH2O x s x L(-1)) and by FOT (5.0+/-0.7, 7.6+/-0.9 and 8.1+/-2.7 cmH2O x s x L(-1)) in healthy subjects without and with added resistance and COPD, respectively, were not significantly different (p>0.05). Rrs measured by both techniques showed a significant coefficient of linear correlation (r=0.70 s) (p<0.01). In the COPD patients, the variability of Delta-inst Rrs (30%) was greater than that of FOT Rrs (21%). The agreement between Ers obtained by Delta-inst and by FOT was less than that found for Rrs. Delta-inst is a noninvasive and simple method for reliably assessing resistance. Therefore, it is useful for monitoring airway obstruction and is potentially helpful in adapting the settings for pressure support ventilation in accordance with patient mechanics. PMID- 11106217 TI - Noninvasive ventilation for acute respiratory failure. Quite low time consumption for nurses. AB - Methods of noninvasive pressure support ventilation (NIPSV) are not always easy to apply in patients with acute exacerbations of chronic obstructive pulmonary disease (COPD). The assistance time spent by nurses in relation to ventilatory time was prospectively studied, when NIPSV was used, in a sequential mode, in COPD patients with either acute exacerbations (58 patients, group I) or postextubation hypercapnic respiratory insufficiency (42 patients, group II) in a medical intensive care unit. During the first 24 h after enrolment, NIPSV was used for 6.7+/-3.2 h (mean+/-SD) in group I and 5.6+/-3.1 h in group II; the duration of NIPSV sessions and the nurse time consumption per session were respectively 47+/-12 and 11+/-7 min in group 1, and 46+/-12 and 11+/-6 min in group II. After the first 24 h of the study, the duration of NIPSV was 4.7+/-3.2 h x day(-1) in group I and 4.9+/-3.5 h x day(-1) in group II, and the nurse time consumption dropped significantly: the duration of NIPSV sessions and the nurse time consumption per session were respectively 44+/-10 and 7+/-4 min in group I, and 47+/-14 and 7+/-3 min in group II. Between the first 24 h and the subsequent period of 24 h, the nursing time dropped significantly (98 versus 59 min in group I (p<0.05), and 85 versus 52 min in group II (p<0.05)). There was no difference in the duration of NIPSV sessions, or in the overall assistance time per session, between the two groups of patients. In conclusion, the study seems to favour a quite low assistance time spent by nurses in relation to ventilatory time when noninvasive pressure support ventilation is used in chronic obstructive pulmonary disease patients with either acute exacerbations or postextubation hypercapnic respiratory insufficiency. PMID- 11106218 TI - Smoking cessation with four nicotine replacement regimes in a lung clinic. AB - Smoking cessation is a key intervention for prevention of several lung diseases. The aim of the present study was to compare the effect of smoking cessation with nicotine replacement in a lung clinic in a low resource set-up suitable for implementation in other lung clinics. This was an open, randomized trial with 4 different nicotine replacement regimes combined with minimal behavioural support in daily routine. A total of 446 smokers (>9 cigarettes x day(-1)) were allocated to a nurse-conducted smoking cessation programme with 4 treatment arms: a 5-mg nicotine patch ("placebo"), a 15-mg nicotine patch, nicotine inhaler, and a 15-mg nicotine patch plus nicotine inhaler. Recommended use of the nicotine products were 3 months with the possibility of continuing use up to 9 months on an individual basis. Individual follow-up studies were scheduled after 2 and 6 weeks, 3, 6, 9 and 12 months. The 12-month point prevalence was 6% (5-mg patch (placebo)), 16% (15-mg patch) (p<0.05), 9% (inhaler) and 11% (15-mg patch plus inhaler), respectively. To conclude, the set-up investigated in this study which included minimal behavioural support with nicotine patches should be evaluated in other lung clinics, as it doubled success rate when compared to a placebo with a 1-yr point prevalence of 16% and also the resources used are limited. PMID- 11106219 TI - Do respiratory epidemics confound the association between air pollution and daily deaths? AB - Daily deaths are associated with air pollution. This association might be con*hhy;founded by uncontrolled risk factors. In order to estimate the potential confounding caused by respiratory epidemics of the association between air pollution and health effects, a time series study of air pollution and daily deaths was carried out. Daily records of deaths for all ages were obtained from five US cities: Chicago, IL; Detroit, MI; Minneapolis, MN; Pittsburgh, PA; and Seattle, WA. Daily levels of particles with a 50% cut-off aerodynamic diameter of 10 microm (PM10) and weather measurements were obtained. City-specific analysis was carried out using Poisson regression, adjusting for time trend, ambient temperature, dew point, barometric pressure and day of the week. A cubic polynomial was used for each epidemic period (> or =10 days of excessive pneumonia hospital admissions), and a dummy variable was used to control for isolated epidemic days. A 10-microg x m(-3) increase in PM10 concentration (lag 0 1) was associated with increased daily deaths in Chicago (0.81%, 95% confidence internal (CI) 0.54-1.09); Detroit (0.87%, 95% CI 0.60-1.15), Minneapolis (1.34%, 95% CI 0.78-1.90), Pittsburgh (0.84%, 95% CI 0.51-1.18) and Seattle (0.52%, 95% CI 0.11-0.94). When controlling for respiratory epidemics, small decreases in the PMlo effect were observed (Chicago 9%, Detroit 11%, Minneapolis 3%, Pittsburgh 5%, and Seattle 15%). The overall effect of PM10 concentration was 0.85% (95% CI 0.60-1.10) per 10 microg x m(-3) before controlling for epidemics and 0.78% (95% CI 0.51-1.05) after. This study showed that the association between air pollution and daily deaths is not due to failure to control for influenza or pneumonia epidemics. PMID- 11106220 TI - What is the respiratory retention of inhaled hexamethylene di-isocyanate? AB - Isocyanates are a frequent cause of occupational asthma. Specific inhalation challenges are often required to confirm the diagnosis. The inhaled concentration has to be assessed during this procedure. However, the respiratory retention of di-isocyanate has not, to the authors knowledge, been evaluated in humans. The existence of a closed-circuit apparatus, designed to carry out these challenges, makes it possible to assess the dose. The respiratory retention of hexamethylene di-isocyanate (HDI) generated in vapour form, in both normal subjects (n=4) and subjects referred for the investigation of occupational asthma due to HDI in whom the diagnosis was excluded (n=5), was assessed. The latter group included four subjects with nonoccupational asthma. The HDI was generated at concentrations varying 5.1-15.2 ppb. The expired concentrations of HDI during such challenges varied 1.4-5.3 ppb. Therefore, the respiratory retention was 61-90%. To conclude, the majority of inhaled hexamethylene di-isocyanate vapour is retained within the airways and/on lung parenchyma. PMID- 11106221 TI - Specifications for equipment used for infant pulmonary function testing. ERS/ATS Task Force on Standards for Infant Respiratory Function Testing. European Respiratory Society/ American Thoracic Society. AB - The aim of this position paper is to define minimal performance criteria for the separate items comprising equipment used to measure respiratory function in infants together with overall performance criteria for the assembled pieces of such equipment. These guidelines cover numerous aspects including: 1) safety, 2) documentation and maintenance of equipment, 3) physical characteristics of mechanical parts and signal transducers, and 4) data acquisition. Further, validation procedures for individual components as well as for the integrated equipment are recommended. Adherence to these guidelines should ensure that infant lung function measurements can be performed with an acceptable degree of safety, precision and reproducibility. They will also facilitate multicentre collection of data and performance of clinical investigations. Manufacturers of infant respiratory function equipment should make every effort to comply with these guidelines, which represent the current standards of paediatric health professionals in this field. PMID- 11106222 TI - Tidal forced expirations. ERS/ATS Task Force on Standards for Infant Respiratory Function Testing. European Respiratory Society/American Thoracic Society. AB - The progress of infant lung function testing has been retarded by both the lack of user-friendly, widely available and affordable equipment and the lack of standardized methodology. The European Respiratory Society/American Thoracic Society Task Force on Standards for Infant Respiratory Function Testing was formed in an attempt to address these deficiencies. This document represents the consensus of investigators with vast experience in the measurement of lung function in infants. The present recommendations deal with equipment requirements, study procedures and reporting of data for measurements of forced expiration at end-tidal inspiration. They represent the "state of the art" in 1999. They are not meant to inhibit further developments in this technique. The authors anticipate that these guidelines will be updated regularly as knowledge progresses. PMID- 11106223 TI - Antibiotic therapy against Pseudomonas aeruginosa in cystic fibrosis: a European consensus. AB - Cystic fibrosis (CF) is the most common lethal hereditary disorder with autosomal recessive heredity in caucasians. The majority of CF patients suffer from chronic respiratory infection with the opportunistic bacterial pathogen Pseudomonas aeruginosa. No consensus among clinicians has been reached so far concerning antibiotic treatment against P. aeruginosa in CF patients. Consensus answers to 24 important questions in this context, based on current evidence, are presented, given by a panel of 34 European experts. Questions addressed and answered are: The diagnosis of P. aeruginosa lung colonization in CF; The impact of P. aeruginosa on the clinical state of CF patients; The assessment of P. aeruginosa susceptibility against antibiotics and the importance of these results for the clinician; The use of monotherapy versus combination therapy; The development of microbial resistance; The achievement of optimal airway concentrations; The effects of subinhibitory concentrations of antibiotics on P. aeruginosa; Statements on the pharmacokinetics of antibiotics in CF patients; Recommendations for doses and dosing intervals and length of treatment regimens; and Toxic side effects due to repeated antibiotic therapy was addressed. The expert panel answered further questions on the use of fluoroquinolones in children with CF, on the administration of nebulized antibiotics and whether prevention of P. aeruginosa lung colonization is possible in CF using antibiotic therapy. Problems of antibiotic therapy at home and in the hospital were addressed, a consensus statement on regular maintenance treatment, or treatment on demand, was given and different routes of administration of antibiotics were recommended for different clinical situations. Finally, the factors which determine the choice of the antibiotic, the dosage, and the duration of the treatment in cystic fibrosis patients were addressed and the design of future antibiotic studies in the context of Pseudomonas aeruginosa lung infection in cystic fibrosis patients were recommended. PMID- 11106224 TI - Genetic aspects in sarcoidosis. AB - Sarcoidosis is an immune-mediated, multiorgan, granulomatous disorder thought to be triggered by an intricate combination of environmental and genetic factors. Two robust lines of evidence support the hypothesis of a genetic component in the pathogenesis of sarcoidosis: racial variation in its epidemiology and familial clustering of cases. The relationship between epidemiology and environmental factors affecting variations in sarcoidosis incidence/prevalence and presentation are reviewed, as well as strategies to be pursued in the search for susceptibility genes for the disorder. Pathogenic processes leading to sarcoid granuloma formation and maintenance have prompted investigators interested in the genetics of sarcoidosis to focus mainly on major histocompatibility complex genes, and indeed a remarkable amount of data has been accumulated during the last two decades. Whilst in contrast with some autoimmune disorders a clear association between human leukocyte antigen (HLA) and sarcoidosis is still a controversial issue, there is, however, a general agreement that some HLA genes are related to phenotypic variations of the disease. Some genetic investigators have focused on T-cell receptor genes, immunoglobulin genes, angiotensin converting enzyme gene, chemokine genes and others. From a review of studies performed in different racial and ethnic groups, a reasonable suggestion arises that genetic factors are the major determinant in the racial variations in the epidemiology of the disorder. This assumption is, however, so far limited by lack of studies considering both genetic and environmental factors simultaneously. PMID- 11106225 TI - Clinical aspects of exhaled nitric oxide. AB - There has been intense research into the role nitric oxide (NO) plays in physiological and pathological mechanisms and its clinical significance in respiratory medicine. Elevated levels of exhaled levels of exhaled NO in asthma and other inflammatory lung diseases lead to many studies examining NO as potential markers of airway inflammation, enabling repeated noninvasive and standardized monitoring of airway inflammation. In airway inflammation, NO is not merely a marker but may have anti-inflammatory and pro-inflammatory effects. Significant correlation has been found between exhaled NO and skin test scores in steroid naive asthmatic patients, allowing to discriminate patients with and without airway responsiveness. Exhaled NO is significantly elevated in acute asthma, or steroid-resistant severe asthma, or when the maintenance dose of inhaled steroids is reduced, and quickly reduced down to the levels in patients with stable asthma after steroid treatment. Exhaled NO has been successfully used to monitor anti-inflammatory treatment with inhaled corticosteroids in asthma. Exhaled NO is extremely sensitive and rapid marker of the dose-dependent effect of steroid treatment, or asthma deterioration, which is increased to any changes in lung function, provocative concentration causing a 20% fall in forced expiratory volume, sputum eosinophilia or asthma symptoms. Exhaled NO is not increased in stable chronic obstructive pulmonary disease (COPD), but patients with unstable COPD, or bronchiectasis have high NO levels. Exhaled and nasal NO are diagnostically low in cystic fibrosis and primary pulmonary dyskinesia. Analysis of exhaled air, including nitric oxide, is feasible and could provide a noninvasive method for use in monitoring and management of lung diseases. PMID- 11106226 TI - Bronchial hyperresponsiveness and airway inflammatory markers in nonasthmatics with allergic rhinitis. PMID- 11106227 TI - Circulating endothelin-1 and obstructive sleep apnoea. PMID- 11106228 TI - Detection and quantitation of human papillomavirus (HPV) DNA in the sera of patients with HPV-associated head and neck squamous cell carcinoma. AB - The human papillomavirus (HPV) has been implicated as an etiological factor in a subset of head and neck squamous cell carcinoma (HNSCC). Because circulating tumor DNA has previously been detected in the sera of patients with advanced HNSCC (stage III or IV), we hypothesized that HPV DNA might be present in the sera of HPV-positive HNSCC patients. Serum DNA extracts from 70 patients with HNSCC were screened for HPV using conventional PCR and a real-time quantitative assay. All samples subjected to conventional PCR were further tested by dot blot hybridization, and positives were confirmed by Southern blotting. Paired tumor DNA from archived tissues was then similarly screened for HPV genomic material (n = 51) or tested by in situ hybridization (n = 19). HPV-16 DNA was detected with L1 primers in 0 of 65 sera and in 15 of 70 (21%) tumors. Conventional PCR with E7 primers and Southern blot hybridization detected HPV-16 DNA in four (6%) sera. Using real-time quantitative PCR, six samples were found to contain various levels of circulating HPV DNA (mean, 12 copies/ml; range, <1-35.) All six serum positive patients had corresponding tumors positive for E7. Four of these patients with HPV-positive tumors later developed distant metastases, suggesting that HPV DNA in serum may represent occult hematogenous spread of cancer cells in this subset of patients. Although a much larger prospective trial is required, the presence of HPV genomic material in serum DNA of HPV-positive HNSCC patients may serve as a useful marker of early metastatic disease. PMID- 11106229 TI - Clinical significance of micrometastases in axillary lymph nodes assessed by reverse transcription-polymerase chain reaction in breast cancer patients. AB - We evaluated the clinical significance of micrometastases in axillary lymph nodes (AxLNs) of breast cancer patients for prediction of prognosis. Archived formalin fixed paraffin-embedded AxLN specimens from 129 node-negative breast cancer patients diagnosed by routine H&E staining between 1986 and 1990 were subjected to carcinoembryonic antigen-specific reverse transcription-PCR analysis. Micrometastases were detected in 40 of 129 (31.0%) node-negative breast cancer patients. After a median follow-up period of 105.6 months, log-rank test analysis indicated that 10-year disease-free and overall survival rates by Kaplan-Meier methods were significantly better in patients without micrometastases than in patients with micrometastases [disease-free survival, 87.6% versus 66.1% (P = 0.0008); overall survival, 93.7% versus 67.8% (P = 0.0024)]. The presence of micrometastases in AxLNs was revealed by multivariate analyses to be an independent and significant predictor of clinical outcome. The hazard ratio was 3.992 (95% confidence interval, 1.293-12.323; P = 0.0161) for relapse and 4.293 (95% confidence interval, 1.043-17.675; P = 0.0436) for cancer-related death. The molecular staging of AxLNs using reverse transcription-PCR is useful for prediction of clinical outcome in early-stage breast cancer patients and can provide a powerful and sensitive complement to routine histopathological analysis. PMID- 11106230 TI - Phase I trial of XR9576 in healthy volunteers demonstrates modulation of P glycoprotein in CD56+ lymphocytes after oral and intravenous administration. AB - XR9576 is a novel inhibitor of P-glycoprotein (P-gp) that has been shown to reverse P-gp-dependent multidrug-resistance in tumor cell lines and tumor-bearing animals. Here we report the first i.v. and p.o. administration to healthy volunteers of XR9576 in dose-escalating studies with the aim of investigating its effects on safety, its pharmacokinetics, and a surrogate marker of efficacy. XR9576 was administered as a single dose-upward titration of 0.1, 0.2, 0.5, 1.0, and 2 mg/kg XR9576 i.v. or 50, 100, 200, 500, and 750 mg/volunteer p.o. The surrogate marker for in vivo efficacy examined the accumulation of the P-gp substrate Rhodamine-123 (Rh-123) in P-gp-expressing CD56+ lymphocytes by flow cytometry. Addition of Rh-123 to blood samples from subjects given XR9576 or a placebo demonstrated drug-dependent modulation of P-gp activity. Even at the lowest doses, significant effects were observed on Rh-123 accumulation in CD56+ cells. Maximal effects were seen during the i.v. infusion or 4-6 h after oral administration. As the dose was increased, a concomitant rise in the level and duration of P-gp blockade was observed. A dose of 2.0 mg/kg i.v. and > or = 200 mg/volunteer p.o. gave approximately 100% inhibition of P-gp for in excess of 24 h. All doses of XR9576 were well tolerated. Inhibition increased with XR9576 plasma concentration, and maximal activity was achieved at 150-200 ng/ml XR9576. In conclusion, XR9576 has demonstrated sustained inhibition of P-gp after i.v. and oral administration and, supported by the elimination half-life of about 24 h, XR9576 is being taken into Phase II as a once-daily agent. PMID- 11106231 TI - Peptide priming of cytolytic activity to HER-2 epitope 369-377 in healthy individuals. AB - The presence of tumor-reactive CTLs in tumor infiltrates and in the peripheral blood of cancer patients demonstrates an immune response against tumors that apparently cannot control disease spread. This raises concerns as to whether amplification of this response may be useful during disease progression. Induction of tumor-reactive CTLs in healthy donors at risk, as well as in patients free of disease, may be therapeutically important, based on the hypothesis that CTLs that recognize tumors early may be more effective in containing their progression than CTLs that expand only when the disease progresses. To address the feasibility of priming cytolytic activity in healthy donors, we used the HER-2 peptide E75 (369-377) as an immunogen and autologous peripheral blood mononuclear cell-derived dendritic cells as antigen-presenting cells. We found that of 10 healthy donors tested, two responded at priming with E75 presented on autologous dendritic cells by induction of E75-specific CTL activity. Three other responders were identified after two additional restimulations. Of these five responders, three recognized E75 presented on the ovarian tumor line SKOV3.A2, as demonstrated by cold-target inhibition experiments. Induction of cytolytic activity at priming was enhanced in responders by tumor necrosis factor-alpha and interleukin 12 but not in the nonresponders. AlphaB7.1 monoclonal antibody added at priming enhanced induction of lytic activity in only one of the four nonresponding donors tested, suggesting that in the majority of donors, E75-precursor CTLs were not tolerized. Because of the possibility that disease may develop in nonresponders, strategies to improve the immunogenicity of tumor antigens for healthy donors may be required for development of cancer vaccines. PMID- 11106232 TI - Phase I trial of paclitaxel plus megestrol acetate in patients with paclitaxel refractory ovarian cancer. AB - Increased expression of P-glycoprotein has been proposed as one important mechanism for inherent or acquired drug resistance of malignant disease to cytotoxic chemotherapy. In experimental systems, hormonal agents, including megestrol acetate (MA), have been shown to be capable of reversing P-glycoprotein mediated multidrug resistance to natural products, including paclitaxel. Because paclitaxel is one of the most active cytotoxic agents in ovarian cancer (OC), we sought to determine whether retreating patients with well-defined paclitaxel resistant OC with a combination of paclitaxel and MA would result in clinically relevant reversal of that resistant state. In this Phase I trial, 44 patients with OC or primary peritoneal carcinoma received paclitaxel (135-175 mg/m2 over 3 h) plus an oral loading dose (800-9600 mg over 24 h) and subsequent maintenance dose (800-3200 mg/day x 3 days) of micronized MA. Both the loading dose and maintenance therapy were delivered in four equal daily doses. Therapy was repeated every 21 days, assuming recovery from the toxicity of the prior course. There were no intrapatient dose escalations. The major toxicity of the regimen was peripheral neuropathy (32% of patients; 11% grade 2-3), although four individuals developed major venous blood clots and one suffered a stroke. Four patients exhibited biological evidence of antineoplastic effects, although only one patient experienced improvement in clinically relevant symptoms. Although pharmacokinetic studies were not performed as a component of this study, prior evaluation of MA pharmacokinetics and in vitro data examining the concentrations of the agent required to reverse P-glycoprotein-mediated paclitaxel resistance suggest that the majority of our patient population achieved levels of MA theoretically capable of producing this desired effect. We conclude that the level of activity and toxicity pattern observed in this trial, associated with the combination of paclitaxel and MA, does not provide strong support for further exploration of the regimen as a treatment strategy to overcome paclitaxel resistance in OC. PMID- 11106233 TI - Phase II study of dolastatin-10 in patients with hormone-refractory metastatic prostate adenocarcinoma. AB - Dolastatin-10 is a natural, cytotoxic peptide with microtubule-inhibitory and apoptotic effects. It has demonstrated in vitro and in vivo efficacy in the DU 145 human prostate cancer model. A Phase II clinical trial was designed in patients with hormone-refractory prostate cancer. Dolastatin-10 was administered at a dose of 400 microg/m2 i.v. every 3 weeks. Dose escalation to 450 microg/m2 was permitted. Toxicity evaluation was conducted every 2 weeks, and assessment of response was done at the end of every two cycles. Sixteen patients were enrolled between October 1998 to December 1999. The median age was 71 years (range, 59-79 years). Median prostate-specific antigen value was 108 ng/ml (range, 15.3-1672 ng/ml). Of the 15 eligible patients, 7 were Caucasian and 8 were African American. Eight patients had bone-only metastases, and seven had measurable disease with or without bone metastases. A total of 56 cycles have been administered. Only 2 patients required dose adjustment because of toxicity, and in 5 patients, dose escalation was feasible to 450 microg/m2. The major toxicities observed were grade 3 and 4 neutropenia in 8 patients and grade 3 neuropathy in 1 patient. All 15 patients are evaluable for response. Three patients demonstrated stable disease; 2 of these had bone disease, and 1 had nodal metastasis. All others had disease progression. Dolastatin-10 is very well tolerated in this elderly, pretreated population but lacks significant clinical activity as a single agent. PMID- 11106234 TI - Clinical phase I dose escalation and pharmacokinetic study of high-dose chemotherapy with treosulfan and autologous peripheral blood stem cell transplantation in patients with advanced malignancies. AB - A Phase I dose escalation and pharmacokinetic study of the alkylating cytotoxic agent treosulfan was conducted to evaluate the maximum tolerated dose and the dose-limiting toxicities in patients with advanced malignancies rescued by autologous peripheral blood stem cell transplantation. Twenty-two patients (15 ovarian and 7 other carcinomas/lymphomas) with a median age of 48 years were treated with 28 high-dose courses. Treosulfan was infused over 2 h at escalating doses from 20 to 56 g/m2, and pharmacokinetic parameters were analyzed. At 56 g/m2, three of six patients experienced dose-limiting toxicities: diarrhea grade III/IV in three patients; mucositis/stomatitis grade III in one patient; toxic epidermal necrolysis in one patient; and grade III acidosis in one patient. Other low-grade side effects, including erythema, pain, fatigue, and nausea/vomiting, were recorded. Two patients died within 4 weeks after treatment because of rapid tumor progression and fungal infection, respectively. Plasma half-life, distribution volume, and renal elimination of treosulfan were independent of dose, whereas the increase in area under the curve was linear up to 56 g/m2 treosulfan. The maximum tolerated dose of high-dose treosulfan is 47 g/m2. A split-dose or continuous infusion regimen is recommended for future high-dose trials. In consideration of antineoplastic activity and limited organ toxicity, inclusion of high-dose treosulfan in combination protocols with autologous peripheral blood stem cell transplantation seems worthwhile. PMID- 11106235 TI - Prognostic value of the type I growth factor receptors in a large series of human primary breast cancers quantified with a real-time reverse transcription polymerase chain reaction assay. AB - We measured the expression of the type I growth factor receptor gene family [epidermal growth factor receptor (EGFR), c-erbB-2, c-erbB-3 and c-erbB-4] in a series of 365 unselected primary breast cancers. The expression was quantified with a real-time one-step reverse transcriptase-PCR (RT-PCR) assay, based upon the 5' nuclease activity of the Taq polymerase and using an Abi Prism 7700 Sequence Detector System (Perkin-Elmer, Courtaboeuf, France). c-erbB-3 and c-erbB 4 were positively correlated to each other (Spearman test) and negatively correlated to EGFR. EGFR and c-erbB-2 were inversely correlated to the presence of estradiol receptors (ER) and progesterone receptors (PgR), and positively correlated to the histoprognostic grading (HPG). Conversely, c-erbB-3 and c-erbB 4 were positively correlated to the presence of ER and PgR, and inversely correlated to the grading HPG. EGFR was inversely related (chi2 test) to the presence of ER and PgR, and positively associated with HPG. In contrast, both c erbB-3 and c-erbB-4 were inversely related to HPG, and positively associated with the presence of ER and PgR. The expression level of EGFR and c-erbB-2 was significantly higher in ER- and PgR-negative tumors compared with ER- and PgR positive tumors (Student's t test), and in tumors with higher grade compared with tumors with lower grade. The expression level of c-erbB-3 and c-erbB-4 was significantly higher in ER- and PgR-positive tumors compared with ER- and PgR negative tumors and in tumors with lower grade compared with tumors with higher grade. In overall survival studies, Cox univariate analyses showed prognostic values of EGFR [> or = median; P = 0.026; risk ratio (RR), 1.6], c-erbB-3 (> or = median; P = 0.0093; RR, 0.58), c-erbB-4 (> or = median; P = 0.0024; RR, 0.52), HPG, node involvement, tumor diameter, ER, and PgR. In Cox multivariate analyses, tumor diameter, ER, and PgR had a prognostic value. In relapse-free survival studies, univariate analyses demonstrated prognostic values of tumor diameter, node involvement, and c-erbB-4 (P = 0.015; RR, 0.65). These three parameters maintained their prognostic value in multivariate analyses (c-erbB-4, P = 0.035; RR, 0.67). This study confirms that EGFR expression and c-erbB-2 expression are markers of tumor aggressiveness in breast cancer. Conversely, we demonstrate that c-erbB-3 and c-erbB-4 elevated expressions are associated with a better prognosis. PMID- 11106236 TI - Human herpesvirus 8 open reading frame 26 and open reading frame 65 sequences from multiple myeloma patients: a shared pattern not found in Kaposi's sarcoma or primary effusion lymphoma. AB - Human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus, has been implicated in the pathogenesis of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), multicentric Castleman's disease, and recently multiple myeloma (MM). DNA sequence analyses of HHV-8 suggest that multiple HHV-8 strains exist. We extracted DNA from 24 patients with MM and 3 patients with monoclonal gammopathy of undetermined significance and compared HHV-8 open reading frames (ORFs) 26 and 65 sequences with those derived from patients with KS, PEL, and two HHV-8-positive PEL cell lines KS-1 and BC-1. ORF26 sequence data suggest that MM patients are consistently carriers of HHV-8 strain subtype C3. All MM patients also consistently revealed either a single bp deletion or substitution at position 112197 in ORF65. This unique alteration is not present in patients with KS or PEL or in PEL cell lines. It occurs in the portion of ORF65 that is known to be responsible for a serological response to HHV-8. PMID- 11106237 TI - Inverse expression of S100A4 and E-cadherin is associated with metastatic potential in gastric cancer. AB - S100A4 is known to be involved in cancer cell motility by virtue of its ability to activate nonmuscle myosin. E-cadherin has an important role in the homophilic cell-cell adhesion and is called an invasion suppressor gene. In the current study, we investigate the histological type and metastatic potential of gastric cancer from the aspect of the interrelationship of E-cadherin and S100A4 expression. Expression of E-cadherin and S100A4 in gastric cancer cell lines, primary gastric cancers, and their normal counterparts were analyzed by reverse transcription-PCR, Western blot, and immunohistochemical methods. S100A4 protein and E-cadherin were expressed in five of eight gastric cancer cell lines, and inverse expression of the two proteins are found in four cell lines. In the clinical specimens, E-cadherin mRNA expression in differentiated adenocarcinomas (88%, 14 of 16) was significantly more frequent than that in poorly differentiated adenocarcinomas (50%, 22 of 44; P = 0.015). Western blot analysis demonstrates that S100A4 protein expression in poorly differentiated adenocarcinomas was 1.6-fold higher than in well differentiated adenocarcinoma. Immunohistochemically, S100A4 expression was detected in 51 (55%) of 92 primary gastric cancers. Reduced expression of E-cadherin in primary tumors was found in 66 (72%) of 92 tumors. S100A4 expression in the poorly differentiated adenocarcinomas had a strong relation to positive lymph node involvement or peritoneal dissemination. Reduced E-cadherin expression showed a strong relationship with positive serosal involvement and infiltrating type. Tumors classified as a group with reduced E-cadherin and high expression of S100A4 reveal positive peritoneal dissemination, serosal involvement, and infiltrating type in the growth pattern. Furthermore, these tumors showed a strong correlation with the poorly differentiated adenocarcinoma. In contrast, tumors with preserved E-cadherin and low expression of S100A4 have a close relation to the well differentiated adenocarcinoma and a favorable prognosis. By the Cox proportional hazard model, S100A4 and E-cadherin tissue status was judged as an independent prognostic factor. S100A4 and E-cadherin tissue status may be a powerful aid in evaluating metastatic potential or the prognosis of patients with gastric cancer. PMID- 11106238 TI - E-cadherin expression is silenced by 5' CpG island methylation in acute leukemia. AB - E-Cadherin is a transmembrane glycoprotein that mediates Ca2+-dependent intercellular adhesion in normal epithelium. In tumors of epithelial origin, E cadherin expression frequently is reduced, an event that contributes to tumor invasion and metastasis. The role of E-cadherin in hematopoietic tissues is less clear. In normal bone marrow, E-cadherin is expressed on erythroid progenitors, CD34+ stem cells, and stromal cells, where it likely contributes to intercellular interactions during hematopoiesis. In this study, we used a nested-PCR approach to examine the methylation status of the E-cadherin 5' CpG island in blood and bone marrow samples from normal donors and in bone marrow from patients with acute leukemia. In normal peripheral blood mononuclear cells and bone marrow, E cadherin was completely unmethylated. In peripheral blood mononuclear cells, expression was evident by reverse transcription-PCR. Immunoblotting confirmed E cadherin protein expression in two lymphoblastoid cell lines derived from normal donors. In contrast, E-cadherin was aberrantly methylated in 4 of 4 (100%) leukemia cell lines, 14 of 44 (32%) acute myelogenous leukemias, and 18 of 33 (53%) acute lymphoblastic leukemias. Genomic bisulfite sequencing of primary leukemias confirmed dense methylation across the CpG island. Methylation was associated with loss of E-cadherin RNA and protein in leukemia cell lines and primary leukemias. Following treatment with 5-aza-2'-deoxycytidine, a methylated leukemia cell line expressed both E-cadherin transcript and protein. Our results show that methylation of E-cadherin occurs commonly in acute leukemia and suggests a hypothesis for E-cadherin down-regulation in leukemogenesis. PMID- 11106239 TI - CYFRA 21-1 serum analysis in patients with esophageal cancer. AB - This study was conducted to determine a potential use of CYFRA 21-1 in patients suffering from carcinoma of the esophagus. CYFRA 21-1 serum concentrations of 50 patients with histologically proven malignant lesion of the esophagus were compared with 50 healthy persons, 50 patients with benign esophageal disease, and 50 patients with benign lung disease. Additional analysis of serum carcinoembryonic antigen, CA 72-4, and squamous cell carcinoma-antigen serum concentrations were performed. The patients with esophageal carcinoma underwent follow-up tumor marker examinations every three months for 1 year. Analysis to detect statistically significant differences was conducted to estimate a cutoff and to evaluate tumor entity, tumor stage, survival, and tumor-free survival. CYFRA 21-1 at a cutoff of 1.40 ng/ml showed an overall sensitivity to esophageal carcinoma of 36% (45.5% to squamous cell carcinoma, 17.6% to adenocarcinoma) at a specificity of 97.3%. CYFRA 21-1 concentrations showed a tendency to higher serum levels depending on local tumor burden. A correlation of CYFRA 21-1 with various N- or M-stage disease was not observed. Postoperative development in terms of survival and tumor-free survival showed significant correlation to preoperative CYFRA 21-1 concentrations. Clinical tumor recurrence was preceded by CY-FRA 21-1 elevation by 3.4 months. For prognosis and follow-up, this marker is justified for additional analysis in a larger series of patients suffering from carcinoma of the esophagus. PMID- 11106240 TI - Prognostic significance of cyclin E overexpression in laryngeal squamous cell carcinomas. AB - Cyclin E plays a pivotal role in the regulation of G1-S transition and relates to malignant transformation of cells. However, the clinical significance of cyclin E in patients with laryngeal squamous cell carcinoma (LSCC) remains unknown. We examined the expression of cyclin E in 102 patients with LSCC and analyzed its relation to clinicopathological parameters, cell proliferation, and clinical outcome. Cyclin E overexpression was observed in 54 cases (52.94%) of LSCC and was significantly correlated with the tumor site (P = 0.012), tumor size (P = 0.006), poor differentiation (P = 0.026), lymph node metastasis (P = 0.012), and advanced stage (P = 0.002). A positive correlation between the cyclin E expression and proliferative activity of tumor cells was found (r = 0.896; P < 0.0001). Kaplan-Meier analysis showed that shorter disease-free and overall survival was significantly associated with proliferating cell nuclear antigen (PCNA) overexpression and cyclin E overexpression. When PCNA and cyclin E are combined, the patients with both PCNA overexpression and cyclin E overexpression had the poorest prognoses when compared with the other cases. Additionally, in early stage (I-II) cases, cyclin E was also revealed to possess a significant prognostic role. By multivariate analysis, lymph node metastasis and cyclin E overexpression were independent prognostic factors for disease-free survival, and tumor size, lymph node metastasis, advanced stage, as well as cyclin E overexpression were independent prognostic factors for overall survival. These findings indicate that cyclin E overexpression is associated with unfavorable clinicopathological parameters and represents an independent marker for cell proliferation and prognosis of LSCC. PMID- 11106241 TI - BRCA1 and BRCA2 mutations in breast cancer families with multiple primary cancers. AB - Ninety-eight women ascertained from high-risk breast/ovarian cancer clinics with breast cancer reporting at least one other primary cancer in themselves or in a relative with breast cancer were compared with 99 women with breast cancer who reported a family history of breast cancer only. All DNA was screened for coding region mutations in BRCA1 and BRCA2 using heteroduplex analysis, followed by direct sequencing. Our data indicate that 42.9% of families reporting breast and any second nonbreast type of primary cancer in the same individual had a BRCA1 or BRCA2 mutation, as compared with the 12.1% of families reporting breast cancer only (P < 0.001). Among the 66 women reporting breast cancer and a nonovarian second primary cancer, 15 (22.7%) had mutations in BRCA1 or BRCA2 (P = 0.04). Among the 32 families where ovarian cancer was the second primary cancer, 27 (84.4%) had a mutation in BRCA1 or BRCA2 (P < 0.001). BRCA1 and BRCA2 mutations were twice as common in the presence of a reported second nonovarian cancer. These data suggest that the presence of multiple primary cancer of any kind may predict for an increased likelihood of finding a BRCA1 or BRCA2 mutation and supports previous studies suggesting that BRCA1 and BRCA2 mutations may be associated with an increased susceptibility to cancers other than breast and ovarian cancer. PMID- 11106242 TI - Expression and prognostic value of Wilms' tumor 1 and early growth response 1 proteins in nephroblastoma. AB - Wilms' tumor is one of the most common solid tumors of children. The protein product of the tumor-suppressor gene, Wilms' tumor 1 (WT-1), binds to the same DNA sequences as the protein product of the early growth response 1 (EGR-1) gene. There is experimental evidence that EGR-1 is involved in controlling cell growth. The expression of both genes in Wilms' tumor was studied by others, mainly at the mRNA level. The present study evaluates the prognostic value of WT-1 and EGR-1 in 61 Wilms' tumors of chemotherapeutically treated patients at the protein level, using an immunohistochemical approach. WT-1 was expressed in normal kidney tissues and in the blastemal and epithelial component of Wilms' tumor, whereas stromal tissue was negative. EGR-1 was expressed in normal kidney tissues and in the three main cell types of Wilms' tumor. In 59 and 56% of Wilms' tumor, the blastemal cells stained for WT-1 and EGR-1, respectively. The blastemal expression of WT-1 and EGR-1 and the epithelial expression of WT-1 were statistically significantly correlated with clinical stage. WT-1 immunoreactivity correlated with EGR-1 expression. Univariate analysis showed that blastemal WT-1 and EGR-1 expression were indicative for clinical progression and tumor-specific survival, whereas epithelial staining was of no prognostic value. Multivariate analysis showed that blastemal WT-1 expression is an independent prognostic marker for clinical progression other than stage. We conclude that a relationship exists between WT-1 and EGR-1 expression in clinical nephroblastomas. Blastemal WT-1 and EGR-1 expression is related to prognosis. PMID- 11106243 TI - Cyclin D1 overexpression is a critical event in gallbladder carcinogenesis and independently predicts decreased survival for patients with gallbladder carcinoma. AB - This study was designed to test the hypothesis that cyclin D1 overexpression is involved in the multistep process of gallbladder carcinogenesis and can be used to predict poor prognosis for patients with gallbladder carcinoma (GBC). Cyclin D1 expression was examined immunohistochemically in a series of specimens, including 8 normal epithelia, 8 benign adenomyoma lesions, 6 precancerous adenomas, and 37 carcinomas of the gallbladder. Four of the 6 (67%) adenomas and 15 of the 37 (41%) adenocarcinomas demonstrated cyclin D1 overexpression (>5% nuclear staining), whereas all normal epithelia and adenomyoma lesions were negative for cyclin D1. Kaplan-Meier curves showed that cyclin D1 overexpression was significantly related to decreased overall survival (P < 0.05) in patients with GBCs. The Cox proportional hazards model identified cyclin D1 overexpression as an independent prognostic marker for death (P = 0.024; risk ratio, 4.2; 95% confidence interval, 1.2-14.7). To test whether cyclin D1 overexpression is a critical event in gallbladder neoplasms, cyclin-dependent kinase inhibitor p27Kip1 was introduced to ascertain how cyclin D1 affects clinical outcomes. Subsequently, neoplasms were divided into three groups on the basis of the combination of cyclin D1 expression and p27Kip1 status, which had been determined previously. Group 1 showed no abnormality in either cyclin D1 or p27Kip1 expression. Group 2 showed aberrant expression of one of the two proteins, whereas group 3 showed concurrent abnormalities in both proteins. Results indicated that overall survival was greatest in group 1, followed by a significant decrease in group 2 and a more precipitous decrease in group 3. In conclusion, cyclin D1 overexpression is an early event in gallbladder carcinogenesis and independently predicts decreased survival for patients with GBC. PMID- 11106244 TI - The short form of the alternatively spliced flt-4 but not its ligand vascular endothelial growth factor C is related to lymph node metastasis in human breast cancers. AB - Angiogenesis is essential for tumor growth and metastasis. It is regulated by numerous angiogenic factors, one of the most important being vascular endothelial growth factor (VEGF). Recently, VEGF-C, a new VEGF family member, has been identified that binds to the tyrosine kinase receptors flt-4 [VEGF receptor (VEGFR) 3] and KDR (VEGFR2). Although the importance of VEGF has been shown in many human tumor types, the contribution of VEGF-C and its primary receptor flt-4 to tumor progression is less well understood. We have therefore measured the level of VEGF-C, flt-4, and KDR mRNA by RNase protection assay and the pattern of VEGF-C expression by immunohistochemistry in 11 normal breast tissue samples and 61 invasive breast cancers. No significant difference in VEGF-C expression was observed between normal and neoplastic breast tissues (P = 0.11). There was a significant correlation between VEGF-C and both flt-4 (P = 0.02) and KDR (P = 0.0002), but no association was seen between VEGF-C and either lymph node status (P = 0.66) or number of involved nodes (P = 0.88), patient age (P = 0.83), tumor size (P = 0.20), estrogen receptor status (P = 0.67), or tumor grade (P = 0.35). No significant relationship was present between VEGF-C and vascular invasion (P = 0.30), tumor vascularity (P = 0.21), VEGF-A (P = 0.62), or thymidine phosphorylase expression (P = 1.00). VEGF-C was expressed predominantly in the cytoplasm of tumor cells, although occasional stromal components including fibroblasts were also positive. We could demonstrate no association between lymph node metastasis and either VEGF-C (P = 0.66) or flt-4 (P = 0.4). However, we did observe a significant loss of the long but not the short isoform of flt-4 in tumors compared with normal tissues (P = 0.02 and P = 0.25, respectively), and this difference was largely accounted for by the reduction of long flt-4 in node positive tumors. These findings strongly support a role for VEGF-C/flt-4 signaling in tumor growth by enhancement of angiogenesis and/or lymphangiogenesis and suggest that differential regulation of these processes may be controlled via flt-4 isoform transcription. They further suggest that the measurement of flt-4 isoform expression may identify a patient group that is likely to have node positive disease and therefore benefit from additional treatment and also emphasize an additional ligand interaction that could be exploited by anti-VEGFR therapy. PMID- 11106245 TI - Relationship between vessel density and expression of vascular endothelial growth factor and basic fibroblast growth factor in small cell lung cancer in vivo and in vitro. AB - In 21 human small cell lung cancer (SCLC) cell lines, we determined the expression of mRNA and secreted protein levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). The VEGF expression was highly variable between cell lines, with a > 100-fold variation, under identical in vitro conditions. The bFGF expression in cell lines was generally very low. Nine of the cell lines were further analyzed during growth as solid tumor xenografts in nude mice (in vivo). A more uniform VEGF protein expression was present in vivo. Compared with the variable in vitro expression, VEGF was relatively up-regulated in the tumor lines CPH 54A and CPH 54B and down-regulated in GLC 3. One line, DMS 79, had a high VEGF expression in vivo as well as in vitro. The vessel density was determined by Chalkley point counting on CD31 immunostained cryosections of tumors of each of the nine SCLC lines. We found a strong positive correlation between vessel density and tissue VEGF protein expression (r(s) = 0.75; P = 0.02) and a comparatively strong negative correlation (r(s) = -0.80; P = 0.01) between vessel density and tissue bFGF expression. No significant correlation was present between vessel density and in vitro VEGF expression. We conclude that VEGF and bFGF expression is dependent on microenvironmental conditions, as well as cell line-specific factors, and that a strong positive correlation exists between in vivo VEGF expression and vessel density, whereas high tissue levels of bFGF are not correlated with higher vessel densities in SCLC xenografts. PMID- 11106246 TI - High preoperative plasma tissue inhibitor of metalloproteinase-1 levels are associated with short survival of patients with colorectal cancer. AB - The objective of the present study was to measure preoperative plasma tissue inhibitor of metalloproteinase (TIMP)-1 levels in colorectal cancer patients and relate these values to clinical and biochemical patient characteristics. TIMP-1 levels were determined by ELISA in EDTA plasma samples collected preoperatively from 588 colorectal cancer patients. Plasma TIMP-1 levels were distributed with a median value of 141.1 microg/liter (range, 53.7-788.7 microg/liter). Whereas no significant differences were found in the median plasma TIMP-1 levels among patients with Dukes' stage A, B, and C disease, patients with Dukes' stage D disease had significantly higher plasma TIMP-1 levels (P < 0.0001); however, high plasma TIMP-1 levels were not restricted to advanced disease. A relatively weak correlation between plasma TIMP-1 level and age was found (r = 0.35; P < 0.0001). There was no significant difference in TIMP-1 levels between males and females (P = 0.97). Univariate analysis demonstrated an increasing risk of mortality with increasing TIMP-1 levels [scored as the log(e)(TIMP-1); hazard ratio = 3.3; 95% confidence interval, 2.6-4.2; P < 0.0001]. Including covariates (Dukes' stage, primary tumor location, gender, age, plasminogen activator inhibitor type 1, and soluble urokinase plasminogen activator receptor) in a multivariate analysis, TIMP-1 was retained in the final model (hazard ratio = 2.5; 95% confidence interval, 1.7-3.7; P < 0.0001). This study showed a highly significant association between preoperative plasma TIMP-1 levels and survival in colorectal cancer patients, with higher plasma TIMP-1 levels being associated with poor outcome. Independent of clinical parameters including Dukes' stage, plasma TIMP-1 levels were found to strongly predict prognosis of colorectal cancer patients. Additional studies are needed to validate the clinical usefulness of plasma TIMP 1 measurements. PMID- 11106247 TI - FRA-1 expression in hyperplastic and neoplastic thyroid diseases. AB - fra-1 gene overexpression has been shown to represent a general event in thyroid cell transformation in vitro and in vivo. Moreover, inhibition of FRA-1 protein synthesis by stable transfection with a fra-1 antisense construct significantly reduces the malignant phenotype of the transformed thyroid cells, indicating a pivotal role of the fra-1 gene product in the process of cellular transformation. In the attempt to define the potential use of FRA-1 protein detection in the diagnosis of thyroid diseases, we analyzed Fra-1 expression by a combination of immunohistochemistry and reverse transcription-PCR (RT-PCR) assay in 174 samples of thyroid nodules (22 nodular hyperplasias, 102 follicular adenomas, 34 papillary carcinomas, 12 follicular carcinomas, and 4 anaplastic carcinomas) representative of the spectrum of thyroid tumor pathology. FRA-1 protein was abundant in all of the carcinoma samples (50/50, 100%), with an intense staining in the nucleus and the cytoplasm. Positive staining was also found in most of the adenomas (90 of 102; 88%), but in this case, the staining was restricted to the nucleus. Similar results were obtained from the analysis of thyroid goiters; however, the number of positive cases is lower than adenomas (8 of 22; 36%); moreover, the staining was not observed in all of the cells. Conversely, no FRA-1 protein was detectable in 12 normal thyroid tissue samples used as controls. RT PCR analysis confirmed a higher fra-1 expression in papillary and follicular carcinomas compared with goiters and adenomas. fra-1 expression was also analyzed on 10 fine needle aspiration biopsy (FNAB) samples by RT-PCR. fra-1-specific mRNA was detected in seven of the eight FNABs corresponding to thyroid nodules that were eventually diagnosed as adenomas (three of four) and carcinomas (four of four) after surgery. Conversely, no fra-1 gene expression was observed in two FNABs derived from normal thyroid. Further studies are required before suggesting FRA-1 protein detection as a useful tool for the diagnosis of hyperplastic and neoplastic disorders of the thyroid gland. PMID- 11106248 TI - Increase in the frequency of p16INK4 gene inactivation by hypermethylation in lung cancer during the process of metastasis and its relation to the status of p53. AB - The p16INK4 gene, which is a tumor suppressor gene, is frequently altered in lung cancers. Hypermethylation of the promoter region of the p16INK4 gene seems to be the major mechanism through which p16INK4 become inactivated. Hypermethylation of the p16INK4 gene was reported to occur at an early stage in lung cancer. To determine whether the change in p16INK4 methylation status occurs at the late stage in the progression of primary lung cancers, we analyzed the primary and metastatic tumor tissues and normal lung samples from 29 cases of advanced lung cancer with distant metastasis. In each tissue sample, we analyzed the p16INK4 and p15INK4b genes for mutations and the methylation status of both genes using PCR-single strand conformation polymorphism, direct sequencing, and methylation specific PCR analysis. We also analyzed a subset of the samples for p16INK4 protein expression. Genetic mutations in the coding region of the p16INK4 and p15INK4b genes were not found in any of the examined specimens. The promoter region of the p16INK4 gene was hypermethylated in the tumor samples of the primary or metastatic site of 37.0% (10 of 27) of the subjects. The promoter region of the p16INK4 gene was hypermethylated at both the primary and metastatic sites in two of the 10 cases and at only the metastatic site in 8 cases. By immunohistochemical analysis, we confirmed the presence of p16INK4 protein at the primary site of all cases in which the promoter region of the p16INK4 gene was hypermethylated at only the metastatic site. Interestingly, all 8 cases with a hypermethylated p16INK4 promoter region, at only the metastatic site, did not have p53 mutation. The results of this study indicate that tumor cells in which the p16INK4 gene has been inactivated by hypermethylation of the promoter region could have an advantage in progression and metastasis in non-small cell lung cancers, especially in the tumors with normal p53, and that the frequency of p16INK4 gene inactivation by hypermethylation could vary in clinical course. PMID- 11106249 TI - Construction and characterization of bispecific costimulatory molecules containing a minimized CD86 (B7-2) domain and single-chain antibody fragments for tumor targeting. AB - Efficient T-cell activation requires two signals. The first signal, which confers specificity, is provided by interaction of the T-cell receptor with peptides presented by MHC molecules. One of the second costimulatory signals is induced by binding of B7 proteins on the surface of antigen-presenting cells to CD28 on the T-cell surface. Expression of B7 molecules on tumor cells can result in the activation of tumor specific T lymphocytes and induce protective antitumor immunity. However, at present such gene-therapeutic approaches are limited by the inability to selectively target B7 gene expression to cancer cells. As an alternative approach we exploited recombinant antibody fragments to localize a costimulatory B7 molecule to the surface of tumor cells. We constructed chimeric proteins that contain in a single polypeptide chain a portion of human B7-2 (CD86) genetically fused to single-chain (sc) Fv antibody domains specific for the tumor-associated antigens epidermal growth factor receptor and the closely related ErbB2 receptor tyrosine kinase. A small recombinant fragment of human CD86 was characterized that corresponds to amino acid residues 1-111 (CD86(111)) of the mature protein. CD86(111) produced in the yeast Pichia pastoris and CD86(111) expressed in bacteria was functionally active and displayed specific binding to B7 counter receptors. Bacterially expressed CD86(111)-scFv fusion proteins specifically localized to the respective target antigens on the surface of tumor cells and markedly enhanced the proliferation of primary T cells when bound to immobilized tumor antigen. PMID- 11106250 TI - Selectivity of TAG-72-targeted adenovirus gene transfer to primary ovarian carcinoma cells versus autologous mesothelial cells in vitro. AB - Efficient gene transfer by recombinant adenovirus (Ad) vectors depends on expression of CAR and alpha(v) integrin on target cells. Because Ad may also infect nearby nontarget cells expressing these receptors, such as peritoneal mesothelial cells after i.p. injection, we hypothesized that targeting Ad gene delivery to a receptor overexpressed on most ovarian carcinoma cells, such as TAG 72, would enhance the selectivity of Ad gene transfer when used in this context. A monoclonal antibody that has been investigated clinically for immunotherapy and immunodetection of ovarian carcinomas, namely CC49, was used to construct a bispecific conjugate with the Fab fragment of a neutralizing anti-knob mAb to target Ad binding via TAG-72. This conjugate facilitated TAG-72-specific, CAR independent Ad reporter gene transfer to both ovarian cancer cell lines and primary ovarian cancer cells cultured from malignant ascites fluid. Fab-CC49 was very selective for tumor cells, augmenting Ad gene transfer to primary ovarian cancer cells 2- to 28-fold relative to untargeted Ad, while also decreasing gene transfer to autologous cultured mesothelial cells 4- to 9-fold. These data suggest that targeting Ad via TAG-72 may improve the selectivity of Ad gene transfer for ovarian tumors 8- to 252-fold on i.p. vector injection. These results also define the requirements for a candidate target receptor in the rational design of a targeted Ad vector for ultimate clinical utility, one that selectively infects tumor cells and spares normal cells on i.p. injection. Such a vector may increase gene transfer and decrease the toxicity of Ad vectors, which would improve the therapeutic index of cytotoxic gene therapy for ovarian cancer in clinical trials. PMID- 11106251 TI - Histone deacetylase inhibitors decrease proliferation and modulate cell cycle gene expression in normal mammary epithelial cells. AB - Full-term pregnancy early in reproductive life is protective against breast cancer in women. The protective effects of parity have variously been attributed to the differentiation that accompanies pregnancy and lactation, alterations in ovarian hormone receptor levels, and altered sensitivity to ovarian hormones. Butyrate, a short-chain fatty acid, induces differentiation in breast cancer cell lines and decreases hormone receptor expression. Butyrate also inhibits proliferation in breast cancer cell lines and modulates expression of key cell cycle-regulatory proteins including cyclin D1. Given these properties, butyrate could be considered a promising agent for breast cancer prevention. Therefore, this study aimed to determine the effects of butyrate on normal human breast epithelial cells and to compare the effects of two stable butyrate derivatives with more favorable pharmacological properties: phenylacetate and its p.o. active precursor phenylbutyrate. Treatment with each agent resulted in concentration dependent growth inhibition in a normal breast epithelial cell line and two breast cancer cell lines (MCF-7 and MDA-MB-231). Phenylbutyrate and butyrate inhibited proliferation to a similar extent, but phenylacetate was less effective in all of the cell lines. All three of the agents induced differentiation (accumulation of lipid droplets) in normal as well as in breast cancer cells and caused a decrease in estrogen receptor (ER) mRNA in MCF-7 cells. The butyrates decreased expression of cyclin D1, increased expression of p21(Waf1/Cip1), and hypophosphorylated pRB in the normal mammary epithelial cells. The effects on cyclin D1 expression correlated with the effects on cell proliferation, which suggests that modulation of cyclin D1 expression may underpin the antiproliferative effects of butyrates. We have shown that butyrate and butyrate like agents are able to decrease proliferation and induce differentiation in normal breast cells as well as in malignant breast cells (ER-positive and ER negative) and, as such, may be considered as candidate chemopreventative agents for women at high risk of developing breast cancer. PMID- 11106252 TI - Antitumor activity of combined treatment of human cancer cells with ionizing radiation and anti-epidermal growth factor receptor monoclonal antibody C225 plus type I protein kinase A antisense oligonucleotide. AB - Recent studies have suggested that selective inhibition of mitogenic pathways may improve the antitumor activity of ionizing radiation. The epidermal growth factor receptor (EGFR) is overexpressed and is involved in autocrine growth control in the majority of human carcinomas. Protein kinase A type I (PKAI) plays a key role in neoplastic transformation and is overexpressed in cancer cells in which an EGFR autocrine pathway is activated. We used two specific inhibitors of EGFR and PKAI that are under clinical evaluation in cancer patients: C225, an anti-EGFR chimeric human-mouse monoclonal antibody (MAb); and a mixed-backbone antisense oligonucleotide targeting the PKAI RIalpha subunit (PKAI AS). We tested in human colon cancer (GEO) and ovarian cancer (OVCAR-3) cell lines the antiproliferative activity of MAb C225 and/or PKAI AS in combination with ionizing radiation. In vivo antitumor activity was evaluated in nude mice bearing established GEO xenografts. Dose-dependent inhibition of soft agar growth was observed in both cancer cell lines with ionizing radiation, C225, or PKAI AS oligonucleotide. A cooperative antiproliferative effect was obtained when cancer cells were treated with ionizing radiation followed by MAb C225 or PKAI AS oligonucleotide. This effect was observed at all doses tested in both GEO and OVCAR-3 cancer cell lines. A combination of the three treatments at the lowest doses produced an even greater effect than that observed when two modalities were combined. Treatment of mice bearing established human GEO colon cancer xenografts with radiotherapy (RT), MAb C225, or PKAI AS oligonucleotide produced dose-dependent tumor growth inhibition that was reversible upon treatment cessation. A potentiation of the antitumor activity was observed in all mice treated with RT in combination with MAb C225 or PKAI AS oligonucleotide. Long-term GEO tumor growth regression was obtained following treatment with ionizing radiation in combination with MAb C225 plus PKAI AS oligonucleotide, which produced a significant improvement in survival compared with controls (P < 0.001), the RT-treated group (P < 0.001), or the group treated with MAb C225 plus PKAI AS oligonucleotide (P < 0.001). All mice of the RT + MAb C225 + PKAI AS group were alive 26 weeks after tumor cell injection. Furthermore, 50% of mice in this group were alive and tumor-free after 35 weeks. This study provides a rationale for evaluating in cancer patients the combination of ionizing radiation and selective drugs that block EGFR and PKAI pathways. PMID- 11106253 TI - Antitumor therapeutic potential of activated human umbilical cord blood cells against leukemia and breast cancer. AB - In this study, in vitro and in vivo antitumor effects of mononuclear cells from human umbilical cord blood cells (UCBCs) and peripheral blood stem cells (PBCs) harvest obtained by leukapheresis were compared. Interleukin 2 (IL-2)-activated mononuclear cells from UCBCs showed increased cytotoxicity against K562 and Raji hematopoietic malignant cells compared with PBCs (P < 0.05). After IL-2 activation, both UCBCs and PBCs showed significant cytotoxicity against MDA-231 human breast cancer cells. The UCBC population involved in this antitumor activity appeared to be CD56+ natural killer precursors. The cytotoxicity of UCBCs was inhibited in the absence of Ca2+ (P < 0.05), supporting a perforin/granzyme-mediated target of cell lysis. In addition, antibodies to Fas ligand blocked cytotoxic activity, suggesting that some of the antitumor cytotoxicity was Fas ligand mediated. In vivo antitumor effects of UCBCs and PBCs were studied using a human leukemic cell-bearing severe combined immunodeficient mouse model. There was a significant increase in the survival of K562 leukemia bearing mice that also received 5 million in vitro IL-2-activated UCBCs or PBCs i.v. on days 3 and day 5 after tumor transplantation compared with untreated mice (P < 0.01). Similar antitumor cytotoxicity of UCBCs and PBCs was also observed against MDA-231 human breast cancer grown in severe combined immunodeficient mice (P < 0.01). These studies suggest that IL-2-activated UCBCs may be a useful source of cellular therapy for patients with hematological malignancies and breast cancer. PMID- 11106254 TI - Tamoxifen inhibits angiogenesis in estrogen receptor-negative animal models. AB - Inhibition of tumor angiogenesis is a therapeutic strategy that can inhibit tumor growth and metastases. The aim of this study was to determine whether the estrogen receptor (ER) ligand drug tamoxifen has antiangiogenic effects. We used three different models of angiogenesis, including measurement of microvessel densities in murine tumors, ex vivo aortic ring assays, and corneal pocket assays. ER-negative fibrosarcoma tumors in tamoxifen-treated ovariectomized rats had significantly less vessel formation compared with untreated animals (median microvessel density, 53.6 versus 94.3 counts/per x 200 field; P = 0.002). Rat aortic rings treated with tamoxifen at several different concentrations demonstrated significantly less vascular sprouting than control rings (P = 0.0001). Corneal pocket assays performed in tamoxifen-treated rats compared with control and estrogen-treated rats demonstrated decreased vascular length (0.88 mm versus 1.26 mm versus 1.47 mm; P = 0.022) and vessel area (21% versus 34% versus 47%; P = 0.018). These three animal models all showed significant inhibition of angiogenesis by tamoxifen and suggest a possible contributory mechanism of ER independent manipulation by tamoxifen in the treatment and prevention of breast cancer. These studies raise the question as to whether or not newer ER ligand drugs might possess even more potent antiangiogenic effects, which in turn could lead to the broadening of the clinical usefulness of these compounds in a number of diseases. More importantly, these studies suggest that the antiangiogenic effects of tamoxifen are due, in part, to ER-independent mechanisms. PMID- 11106255 TI - The aromatic-L-amino acid decarboxylase inhibitor carbidopa is selectively cytotoxic to human pulmonary carcinoid and small cell lung carcinoma cells. AB - The carcinoid tumor is an uncommon neuroendocrine neoplasm the hallmark of which is excessive serotonin production. In studying kinetics of tryptophan hydroxylase and aromatic-L-amino acid decarboxylase (AAAD) in human carcinoid hepatic metastases and adjacent normal liver (J. A. Gilbert et al, Biochem. Pharmacol., 50: 845-850, 1995), we identified one significant difference: the Vmax of carcinoid AAAD was 50-fold higher than that in normal liver. Here, we report Western and Northern analyses detecting large quantities of AAAD polypeptide and mRNA in human carcinoid primary as well as metastatic tumors compared with normal surrounding tissues. To assess the feasibility of targeting these high AAAD levels for chemotherapy, AAAD inhibitors carbidopa (alpha-methyl-dopahydrazine), alpha-monofluoromethyldopa (MFMD), and 3-hydroxybenzylhydrazine (NSD-1015) were incubated (72 h) with NCI-H727 human lung carcinoid cells. Carbidopa and MFMD were lethal (IC50 = 29 +/- 2 microM and 56 +/- 6 microM, respectively); NSD-1015 had no effect on proliferation. On exposure to other human tumor lines, carbidopa was lethal only to NCI-H146 and NCI-H209 small cell lung carcinoma (SCLC) lines (IC50 = 12 +/- 1 microM and 22 +/- 5 microM, respectively). Carbidopa (100 microM) decreased growth of (but did not kill) SK-N-SH neuroblastoma and A204 rhabdomyosarcoma cells and did not affect proliferation of DU 145 prostate, MCF7 breast, or NCI-H460 large cell lung carcinoma lines. The rank order of lines by AAAD activity was NCI-H146 > NCI-H209 > SK-N-SH > NCI-H727, whereas A204, DU 145, MCF7, and NCI-H460 had no measurable activity. For lung tumor lines (carcinoid, two SCLC, and one large cell lung carcinoma), AAAD activity was correlated with the potency of carbidopa-induced cytotoxicity. However, carcinoid cell death was not solely attributable to complete inhibition of either AAAD activity or the serotonin synthetic pathway. In further evaluating potential applications of these findings with carbidopa, we determined that sublethal doses of carbidopa produced additive cytotoxic effects in carcinoid cells in combination with etoposide and cytotoxic synergy in SCLC cells when coincubated with topotecan. PMID- 11106256 TI - Rapid development of tamoxifen-stimulated mutant p53 breast tumors (T47D) in athymic mice. AB - MCF-7 cells are used routinely to study tamoxifen-stimulated drug resistance in vivo. However, unlike MCF-7 cells, T47D cells express mutant p53 protein and lose the estrogen receptor (ER) during long-term estrogen deprivation in vitro [Pink et al., Br. J. Cancer, 74: 1227-1236, 1996 (erratum, Br. J. Cancer, 75: 1557, 1997)]. As a result, T47D tumors may respond differently from MCF-7 tumors to long-term tamoxifen treatment. Ovariectomized athymic mice were given injections bilaterally with T47D cells (5 x 10(5)) into the mammary fat pads. A rapidly growing estradiol responsive tumor (T47D:E2) was established and 0.5 mg of tamoxifen given daily blocked estrogen-stimulated growth. In subsequent experiments, low doses of tamoxifen (0.17 mg or 0.5 mg) did not produce tamoxifen stimulated tumors at 14 weeks, whereas high-dose tamoxifen (1.5 mg) consistently produced tamoxifen-stimulated tumors (T47D:Tam; 17 tumors/20 sites) at 8 weeks. In contrast, 1.5 mg of tamoxifen produced tamoxifen-stimulated MCF-7 tumors (MCF 7: Tam2) at a slower rate (20 weeks) and less consistently (14 tumors/26 sites). When the T47D:Tam tumor was passaged, it grew maximally with either 1.5 mg of tamoxifen or a 1-cm estradiol (premenopausal levels) capsule, and similar results were obtained with MCF-7:Tam2 tumors. Interestingly, when T47D:Tam tumors were treated with the 0.5 mg of tamoxifen, tumors grew only to 50% maximum. All of the tumors originating from MCF-7 and T47D cells expressed ER at similar levels; therefore, tamoxifen did not select for an ER-negative tumor. In conclusion, we have shown that tamoxifen-stimulated T47D p53 mutant tumors can be developed rapidly with high-dose therapy (1.5 mg daily). The results from this model provide new opportunities to investigate the rapid development of drug resistance to adjuvant tamoxifen in patients with mutant p53 breast tumors. PMID- 11106258 TI - The importance of p53-independent apoptosis in the intestinal toxicity induced by raltitrexed (ZD1694, Tomudex): genetic differences between BALB/c and DBA/2 mice. AB - The thymidylate synthase inhibitor raltitrexed (ZD1694, Tomudex) induces greater intestinal toxicity, manifested as diarrhea and weight loss, in BALB/c than in DBA/2 mice. No convincing pharmacokinetic or pharmacodynamic reason for this strain difference has been established. We have investigated whether this strain difference in response to raltitrexed is related to differential susceptibilities of intestinal mucosae to undergo apoptosis and also whether p53 expression, a critical factor in 5-fluorouracil-induced intestinal apoptosis and toxicity, modulates this response. Ten mg/kg or 100 mg/kg raltitrexed were administered as single or double i.p. injections 24 h apart to BALB/c, DBA/2, and p53-/- mice. Apoptosis, mitosis, and tissue damage were assessed in intestinal epithelium, and animal weight was recorded. BALB/c mice developed diarrhea and weight loss following 100 mg/kg x2 raltitrexed, whereas DBA/2 mice did not. BALB/c mice were more sensitive than DBA/2 to induction of small-intestinal and colonic apoptosis 24 h following 100 mg/kg raltitrexed. Inhibition of mitosis was equivalent in both strains. Both strains showed histopathological damage to the small intestine after 100 mg/kg x2 raltitrexed, but only BALB/c mice demonstrated colonic damage. p53-null mice showed the same level of small intestinal apoptosis as their wild type counterparts 24 h after 100 mg/kg x1 raltitrexed and also the same levels of intestinal toxicity 3, 5, and 7 days after 100 mg/kg x2 raltitrexed. Thus, BALB/c mice were more susceptible to induction of intestinal apoptosis by raltitrexed than DBA/2 mice and also demonstrated more histopathological damage in the colon correlating with the induction of diarrhea and weight loss. In contrast to 5 fluorouracil, the intestinal apoptosis and toxicity induced by raltitrexed were p53-independent. PMID- 11106257 TI - Induction of antitumor immunity with combination of HER2/neu DNA vaccine and interleukin 2 gene-modified tumor vaccine. AB - The therapeutic effects of both cytokine-secreting tumor vaccine and DNA vaccine were studied using mouse MBT-2 bladder cancer cells as a model. Cytokine secreting MBT-2 cells were obtained by infecting cells with retroviral particles containing interleukin (IL) 2-, IL-4-, or granulocyte-macrophage colony stimulating factor (GM-CSF)-expression vector. The MBT-2-IL-2 cells were not tumorigenic in syngenic C3H mice at all. Tumor formation decreased significantly for the MBT-2-GM-CSF cells. MBT-2-IL-2, -IL-4, and -GM-CSF cells were killed by irradiation and tested as tumor vaccines. The irradiated MBT2-IL-2 cells could complete protect mice from the growth of the preexisting tumor cells, and the immune memory lasted for 8 months. On the other hand, irradiated MBT-2-IL-4 and MBT-2-GM-CSF cells were less effective. When the loading tumor mass increased, all tumor vaccines lost protective effects. DNA vaccine encoding the tumor antigen neu was additionally tested to improve the therapeutic efficacy. Coinjection of 60 microg pSV-neu DNA was effective in enhancing the antitumor effects of MBT2-IL-2; however, DNA vaccine alone cannot prevent the progression of the preexisting tumor. Immunohistochemical analysis of tumor infiltrate revealed massive increase of CD4+ lymphoid cells in the group of mice treated with both DNA vaccine and IL-2-secreted tumor vaccine. Western blotting demonstrated the presence of anti-neu antibody in the serum from immunized mice. In contrast, combination of DNA vaccine and MBT-2-GM-CSF has no additive effect. The results indicate the combination of DNA vaccine and IL-2-secreting tumor vaccine can additionally improve therapeutic efficacy, and the efficacy is correlated with the increase of CD4+ T lymphocytes and anti-neu antibody. PMID- 11106259 TI - Specific distribution of TOP-53 to the lung and lung-localized tumor is determined by its interaction with phospholipids. AB - We have investigated the mechanism of TOP-53 distribution to the lung and lung localized tumor. In contrast to etoposide (VP-16), TOP-53 contains a basic aminoalkyl group that may predispose it to interact specifically with phospholipids, consequently leading to an increase of drug accumulation in the tissues. Therefore, we have studied its interaction with phospholipids in vitro using an organic solvent-water partition system. TOP-53 appeared to have the most potent binding affinity (Ka = 563 x 10(-2) microM) to phosphatidylserine (PhS), whereas VP-16 showed no interaction with any phospholipid tested. PhS content determined after HPLC separation varied among tested tissues; however, large quantities were found in normal lung and lung cancer tissues far exceeding those present in the liver and kidney. The predicted tissue-to-plasma partition coefficient values, estimated based on PhS content and its binding affinity, resembled those experimentally determined. We concluded that tissue distribution of TOP-53 is determined by PhS content in the tissues and by binding affinity. As a result of specific accumulation in the lung, TOP-53 appeared to show a strong antitumor activity (increase of life span = 171%) against cancer metastasizing to the lung, whereas VP-16 was less effective (increase of life span = 78%). These results suggest that TOP-53 may have an advantage over VP-16 in the treatment of lung cancers in patients. PMID- 11106260 TI - Prostate cancer radiosensitization in vivo with adenovirus-mediated p53 gene therapy. AB - An adenovirus 5 vector containing wild-type p53 cDNA (Ad5-p53) and a cytomegalovirus promoter was used to generate p53 transgene expression. Control vector (Ad5-pA) contained the poly-adenosine sequence. PC3 cells (2 x 10(6)) were injected s.c. into the legs of nude mice. Treatment with Ad5-p53 was initiated at a tumor volume of 200 mm3. Three intratumoral injections (days 1, 4, and 7) were given with 3 x 10(8) plaque-forming units, followed by 5 Gy pelvic irradiation (day 8) in one fraction using a cobalt-60 source. Tumor volume measurements were obtained every 2 days. LNCaP cells (2 x 10(6)) were injected orthotopically into the prostates of nude mice, and tumor weight was approximated using serum prostate-specific antigen (PSA) obtained from weekly tail vein bleedings. The target PSA for the start of the studies was 5 ng/ml. The intraprostatic injections of Ad5-p53 were done twice (days 1 and 2) and followed by 5 Gy pelvic irradiation on day 3. The PC3 tumor volume growth curves were log transformed and fitted using linear regression. The times (in days) for the tumors to reach 500 mm3 were calculated as 10.7 +/- 0.7 (+/- SE) for the saline control (no virus), 9.8 +/- 2.1 for Ad5-pA, 15.6 +/- 1.6 for Ad5-p53, 14.6 +/- 1.5 radiation therapy (RT; 5 Gy), 14.6 +/- 1.5 for Ad5-pA plus RT, and 31.4 +/- 5.3 for Ad5-p53 plus RT. The Ad5-p53 plus RT times were significantly different from the other groups. An enhancement factor of 3.4 was calculated, indicating supra-additivity. LNCaP tumor growth was determined via weekly serum PSA measurements. Treatment failure was determined using two PSA-based methods; a serum PSA of > 1.5 ng/ml or two rises in PSA during 6 weeks posttreatment. The results were similar using either end point. Treatment with Ad5-p53 plus 5 Gy resulted in significantly fewer PSA failures (<30%), as compared with Ad5-p53 alone (64-73%) and the other controls (approximately 80-100%) These results are also consistent with a supra-additive inhibition of tumor growth. Tumor growth in vivo was inhibited supra-additively when p53null and p53wildtype prostate tumors were treated with Ad5-p53 and 5 Gy radiation. PMID- 11106261 TI - Bioactivation of tegafur to 5-fluorouracil is catalyzed by cytochrome P-450 2A6 in human liver microsomes in vitro. AB - Tegafur is a prodrug of 5-fluorouracil (5-FU) consisting of a new class of oral chemotherapeutic agents, tegafur/uracil and S-1, which are classified as dihydropyrimidine dehydrogenase inhibitory fluoropyrimidines. It is bioactivated to 5-FU via 5'-hydroxylation mediated by cytochrome P-450 (CYP). However, which isoform(s) of CYP is responsible for the bioactivation process of tegafur remains unclear. The purpose of the present study was to identify the human CYP isoform(s) involved in the metabolic activation of tegafur using human liver microsomes and cDNA-expressed human CYPs. The formation of 5-FU from tegafur in human liver microsomes showed biphase kinetics with Km and Vmax values for the high-affinity component of 0.43 +/- 0.05 mM and 4.02 +/- 1.70 nmol/mg/min (mean +/- SD, n = 4), respectively. In the correlation study using a panel of 10 human liver microsomes, the formation of 5-FU from tegafur showed a significant correlation (r = 0.98; P < 0.001) with coumarin 7-hydroxylation, a marker activity of CYP2A6. In addition, a specific substrate of CYP2A6 and anti-CYP2A6 antibody inhibited the formation of 5-FU by 90% in human liver microsomes. Moreover, cDNA-expressed CYP2A6 showed the highest activity for the formation of 5-FU among 10 cDNA-expressed CYPs, with a Km value similar to that found for the high-affinity component in human liver microsomes. These findings clearly suggest that CYP2A6 is a principal enzyme responsible for the bioactivation process of tegafur in human liver microsomes. However, to what extent the bioactivation of tegafur by CYP2A6 accounts for the formation of 5-FU in vivo remains unclear, because the formation of 5-FU from tegafur is also catalyzed by the soluble fraction of a 100,000 x g supernatant and also derived from spontaneous degradation of tegafur. PMID- 11106262 TI - Increased oral bioavailability of paclitaxel by GF120918 in mice through selective modulation of P-glycoprotein. AB - Previous studies in mice with disrupted mdr1a P-glycoprotein genes have shown that the oral bioavailability of paclitaxel is very low because of the presence of this drug-transporting protein in the intestinal wall. Additional studies with cyclosporin A have shown that this P-glycoprotein-inhibiting agent is able to increase the bioavailability of paclitaxel in mouse models and in patients. However, the potential immune-suppressive side effects of cyclosporin A renders this compound less suitable for chronic use in cancer patients. In this paper we present the results obtained with GF120918, an experimental P-glycoprotein inhibitor, on the oral bioavailability of paclitaxel in both wild-type and mdrlab knockout mice. GF120918 (25 mg/kg) was administered p.o. by gavage 15 min or 2 h before oral or i.v. dosing of paclitaxel, respectively. Paclitaxel plasma levels were quantified by high-performance liquid chromatography. GF120918 increased the plasma values for areas under the concentration-time curve of oral paclitaxel in wild-type mice by 6.6-fold from 408 to 2701 ng x ml(-1) h. Calculated relative to their respective values for area under the concentration-time curve after i.v. administration, GF120918 increased the oral bioavailability of paclitaxel in wild type mice from 8.5 to 40.2%. The plasma pharmacokinetics of paclitaxel in mdr1ab knockout mice was not altered by GF120918, whereas the pharmacokinetics of paclitaxel in wild-type mice receiving GF120918 became comparable with mdr1ab knockout mice. This result indicates that GF120918 at this dose-level selectively and completely blocks P-glycoprotein in the intestines and does not notably interfere in the elimination of paclitaxel by metabolism or other transporters. On the basis of this result, GF120918 has been selected for additional study in humans. PMID- 11106263 TI - Gene therapy of human bladder cancer with adenovirus-mediated antisense basic fibroblast growth factor. AB - We previously investigated the role of basic fibroblast growth factor (bFGF) as a mediator of angiogenesis, tumorigenicity, and metastasis of transitional cell carcinoma (TCC) of the bladder. In the present study, we determined whether adenoviral-mediated antisense bFGF gene transfer therapy (Ad bFGF-AS) would inhibit TCCs growing in the subcutis of nude mice. In vitro, Ad bFGF-AS inhibited endothelial cell proliferation and enhanced apoptosis. The highly metastatic human TCC cell line 253J-BV(R) was implanted ectopically in the subcutis of athymic nude mice, and therapy was begun when the tumors reached a diameter between 5 and 7 mm. Intralesional therapy with Ad bFGF-AS decreased the in vivo expression of bFGF and matrix metalloproteinase type 9 mRNA and protein, and reduced microvessel density and enhanced endothelial cell apoptosis. Tumor growth was significantly inhibited by Ad bFGF-AS (mean, 58 mg) compared with controls [saline (mean, 562 mg), beta-galactosidase adenovirus (mean, 586 mg), and sense bFGF adenoviral therapy (Ad bFGF-S; mean, 3012 mg)]. These results suggest that Ad bFGF-AS therapy affects endothelial cells directly and tumor cells indirectly through down-regulation of bFGF and matrix metalloproteinase type 9, resulting in endothelial cell apoptosis and significant tumor growth inhibition. Furthermore, these studies confirm that bFGF expression is a valid target for the therapy of bladder cancer. PMID- 11106264 TI - p53 dependence of Fas induction and acute apoptosis in response to 5-fluorouracil leucovorin in human colon carcinoma cell lines. AB - We examined the patterns of induction of apoptosis, Fas expression, and the influence of the status of the p53 tumor suppressor gene, in response to treatment of human colon carcinoma cell lines to 5-fluorouracil (FUra) combined with leucovorin (LV) under conditions of both DNA-directed (HT29, VRC5/c1, and RKO) and RNA-directed (HCT8 and HCT116) cytotoxicity. Acute apoptosis was induced in cell lines expressing wtp53 (RKO, HCT8, and HCT116), independent of the mechanism of FUra action. In HT29 cells that expressed mp53, apoptosis was a delayed event. Cell lines undergoing DNA-directed FUra cytotoxicity demonstrated marked accumulation of cells in S-phase (HT29 and RKO), whereas those lines undergoing RNA-directed cytotoxicity (HCT8 and HCT116) demonstrated marked cell cycle phase arrest in G2-M, both reversible by dThd. dThd partially protected HCT8 and HCT116 cells from FUra-LV-induced apoptosis but had no influence on FUra LV-induced loss in clonogenic survival. In cells expressing wtp53, the Fas death receptor was induced in response to FUra-LV treatment. FUra-LV sensitized RKO cells to the anti-Fas monoclonal antibody CH-11 that was completely reversed by dThd, demonstrating the involvement of DNA damage in FUra-LV-induced, Fas dependent sensitization to CH-11. In contrast, FUra-LV sensitized HCT116 cells to CH-11-induced apoptosis, which was not dThd reversible. Transduction of HT29 cells with Ad-wtp53 induced elevated Fas expression and sensitized the cells to FUra-LV-induced apoptosis. Data indicate that the presence of a wtp53 gene determines FUra-LV-induced Fas expression, the kinetics of FUra-LV-induced apoptosis and not the extent of apoptosis induced, both being independent of the mechanism of FUra action. Therefore, in colon carcinomas that express wtp53, the approach to sensitize tumors to Fas-mediated apoptosis may be further enhanced from the effect of FUra-LV in elevating Fas expression in a p53-dependent manner. PMID- 11106265 TI - Genetically modified CD34+ cells exert a cytotoxic bystander effect on human endothelial and cancer cells. AB - We and others have proposed mammalian cells as gene delivery vehicles with the potential for overcoming physiological barriers to viral vectors. To that end, we previously have shown the potential of CD34+ endothelial progenitors for systemic gene delivery in a primate angiogenesis model. Here we seek to explore the utility of CD34+ cells of human origin as vehicles for toxin genes and, in particular, to measure their capacity to effect a cytotoxic bystander effect in human endothelium and tumor cells. To this end, CD34+ cells were transduced with TOZ.1, a nonreplicative herpes simplex vector encoding thymidine kinase. To test the capacity of CD34+ cells to induce a cytotoxic bystander effect in target cells, we performed mixing experiments, whereby TOZ.1-transduced CD34+ cells were mixed with either human vascular endothelial cells or human ovarian tumor cells (SKOV3.ip1). Cell viability was measured by the MTS assay. Lastly, mixtures of TOZ.1-transduced CD34+ cells and SKOV3.ip1 tumor cells were injected s.c. to evaluate the bystander effect in vivo. After transduction of CD34+ cells with TOZ.1, treatment with ganciclovir induced the killing of 99% of cells. In cell mixing experiments, a linear correlation was observed between the percentages of TOZ.1-transduced CD34+ cells and total cell killing. For example, when 50% of CD34+ transduced cells were mixed with nontransduced SKOV3.ip1, >70% of all cells died. Similarly, when the same percentage was mixed with human vascular endothelial cells, >80% of the total number of cells died. In vivo studies showed an abrogation of tumor formation when TOZ.1-transduced CD34+ cells and ganciclovir were administered. Our observations establish the feasibility of a method for cell-based toxin gene delivery into disseminated areas of tumor angiogenesis. PMID- 11106266 TI - Kupffer cells do not play a role in governing the efficacy of liposomal mitoxantrone used to treat a tumor model designed to assess drug delivery to liver. AB - A tumor model designed to assess liposome-mediated drug delivery to liver has been used in an attempt to better understand the mechanism of activity of liposomal mitoxantrone, a liposomal anticancer drug formulation that appears to be uniquely effective in treating this tumor model. Reductions in liposomal mitoxantrone accumulation in the liver were achieved either by use of poly(ethylene)glycol (PEG)-modified lipids or by methods designed to deplete liver phagocytes, a method referred to as hepatic mononuclear phagocytic system (MPS) blockade. A 2-fold reduction in mitoxantrone delivery to the liver was obtained using a mitoxantrone formulation with PEG-modified lipids, and a 3-fold reduction was obtained when liposomal mitoxantrone was given to animals pretreated to induce hepatic MPS blockade. Results demonstrate that the liposomal mitoxantrone formulation prepared with PEG-modified lipids was significantly less active than the formulations that did not contain PEG lipids, with median survival times of 17 days and 100% 60-day survival, respectively. In contrast, hepatic MPS blockade had no effect on the therapeutic activity of 1,2-dimyristoyl phosphatidylcholine/cholesterol (DMPC/Chol) mitoxantrone (100% 60-day survival). These data suggest that the hepatic MPS does not play a role in mediating the therapeutic activity of DMPC/Chol mitoxantrone in the treatment of liver localized disease. Results with formulations prepared with a PEG-stabilized surface, however, suggest that nonspecific methods to decrease liposome cell interactions inhibit the therapeutic activity of DMPC/Chol mitoxantrone. PMID- 11106267 TI - Correspondence re: A. E. Biemer-Huttmann et al., Mucin core protein expression in colorectal cancers with high levels of microsatellite instability indicates a novel pathway of morphogenesis. Clin. Cancer Res., 6: 1909-1916, 2000. PMID- 11106268 TI - Identifying the genetic basis for neurologic disease: the pediatrician's call to action. PMID- 11106269 TI - Genetics of brain development and malformation syndromes. AB - The identification of the specific genes responsible for several childhood neurologic disorders has provided a framework with which to understand key development stages in human brain development. Common genetic disorders of brain development include septo-optic dysplasia, schizencephaly, holoprosencephaly, periventricular heterotopia, lissencephaly, and Joubert syndrome. For each of these disorders, a critical step in brain development is interrupted. The identification of the responsible genes is providing scientists a window into the key modulators of brain development, and providing clinicians the opportunity to offer genetic testing to individual patients and their families. PMID- 11106270 TI - Genetics of mental retardation. AB - Aim of this review is to present the latest advances in the identification of the genetic determinants of intellectual deficiency. Mental retardation (MR) is often associated with other neurologic symptoms, metabolic disorders, or malformation syndromes. The purpose of the review is to subdivide the large field of MR into categories that may help professionals in making a diagnosis. Nonspecific MR can also segregate in families and the mapping and cloning of corresponding mutant genes will eventually advance our understanding of normal and abnormal brain functioning. Several genes responsible for nonspecific X-linked mental retardation have been identified in the last 12 to 24 months and are being intensively investigated. This will hopefully lead to new possibilities of either genetic or pharmacological therapy. PMID- 11106271 TI - Genetics of the epilepsies. AB - Recent molecular insights into the human idiopathic epilepsies have suggested the central role of ligand-gated and voltage-gated ion channels in their etiology. So far, genes coding for sodium and potassium channel subunits as well as a nicotinic cholinergic receptor subunit have been identified for Mendelian idiopathic epilepsies. In vitro and in vivo studies of mutations demonstrate functional changes, allowing new insights in mechanisms underlying hyperexcitability. Similarly, spontaneous murine epilepsy models have been associated with calcium channel molecular defects. The major challenge before us in understanding the genetics of the epilepsies is to identify genes for common forms of epilepsy following complex inheritance. Once such genes are discovered, the gene-gene-environmental interactions producing specific epilepsy syndromes can be explored. PMID- 11106272 TI - Genetics of brain tumors. AB - Brain tumors are among the most common forms of cancer in children and account for most cancer-related deaths in this age group. The incidence of brain tumors appears to be increasing in children, while therapeutic advances have been modest. Few genetic studies exist on pediatric brain tumors, in part because tissue from low-grade and brain stem tumors is not readily available, and also because individual centers have relatively few cases. Genetic changes in infiltrating astrocytomas involve genes in the p53 and RB pathways, and show alterations that are similar to infiltrating astrocytomas in adults. The PTC gene is mutated in a subgroup of medulloblastomas, and may lead to increased proliferation in granule cells that normally express this receptor. Further studies are needed to identify genetic alterations in pilocytic and low-grade astrocytomas, which account for 40% of brain tumors in children. PMID- 11106273 TI - Genetics of pediatric neuromuscular disease. AB - Our understanding of the neuromuscular disorders of childhood has been rapidly expanding. This is mostly because of the discovery of the underlying genetic loci for the vast majority of these diseases and the abnormal proteins produced caused by these mutations. Spinal muscular atrophy is the second most frequent autosomal recessive disease of childhood and the most fatal. It has been mapped to chromosome 5q11.2-13.3, an area with three distinct genes associated with spinal muscular atrophy. Charcot-Marie-Tooth is the most common inherited peripheral neuropathy. Three genes encoding for myelin proteins and one for a nuclear protein have been associated with this group of disorders. Finally, since dystrophin was cloned in 1986, many proteins assisting dystrophin in anchoring the muscle cytoskeleton to the extracellular matrix have been discovered. Mutations in these genes lead to various forms of muscular dystrophy. PMID- 11106274 TI - Allergy, immunology, and related disorders. PMID- 11106275 TI - Gene therapy of lymphoid primary immunodeficiencies. AB - Gene therapy offers an attractive option to the most severe forms of primary immunodeficiency diseases. Identification of disease-associated genes as well as advances in the technology of gene transfer into hematopoietic progenitor cells have set the basis for the first clinical trials. Settings characterized by the potential for a selective advantage provided to transduced cells are the first diseases to target. The recent example of successful treatment of severe combined immunodeficiency-X1 (gammac deficiency) illustrates this potential. PMID- 11106276 TI - Periodic fever syndromes. AB - The term periodic fever syndrome has been used in a restricted sense to denote two diseases in which episodic fevers occur with a regular periodicity: cyclic neutropenia and the periodic fever, aphthous stomatitis, pharyngitis, and adenopathy (PFAPA) syndrome. Other authors have used the term in a more general sense to encompass a larger group of disorders characterized by recurrent episodes of fever that do not necessarily follow a strictly periodic pattern. These include familial Mediterranean fever, the autosomal dominant familial fevers (also known as Hibernian fever), and the hyperimmunoglobulin D syndrome. This article follows the latter usage, and reviews recent advances in our understanding of the genetics and molecular pathology of this group of diseases, as well as their clinical characterization and treatment. PMID- 11106277 TI - Peanut and tree nut allergy. AB - Among foods causing allergic reactions in children, peanut (a legume) and tree nuts (ie, walnut, hazel nut, Brazil nut, pecan) have attracted considerable attention for several reasons. Allergies to these foods are common, frequently have an onset in the first few years of life, generally persist, and account for severe and potentially fatal allergic reactions. Furthermore, the ubiquity of these foods in the diet makes avoidance difficult and accidental ingestions, with reactions, common. This review discusses recent and emerging information on the prevalence, clinical characteristics, natural history, genetic basis, and current treatment of these allergies. In addition, recent advances in the molecular and immunologic characteristics of these allergens, and novel therapeutic options under investigation in animal models, are reviewed. PMID- 11106278 TI - Immunotherapy of asthma and allergic diseases. AB - The goals of therapy for allergic disease and asthma, which have increased dramatically during the past 2 decades, are to relieve and prevent symptoms. Currently, allergen immunotherapy is the only available treatment that can reduce symptoms, alter the natural course of disease, and induce long-term clinical remission effectively and safely in patients with allergic rhinitis, asthma, and insect venom anaphylaxis. Allergen immunotherapy may even prevent the evolution towards polysensitization and prevent the development of asthma in allergic children. In the long run, it is more cost-effective than pharmacotherapy and environmental control measures alone. Future developments, such as using alternate routes of administration, peptide fragments of allergen, adjuvants, and DNA vaccines, may improve its efficiency in inducing long-term clinical relief of symptoms. PMID- 11106279 TI - Pediatric asthma among minority populations. AB - Minority children in the United States are at higher risk for asthma and related hospitalizations than white children, and their asthma tends to be more severe. Empirical studies have yet to demonstrate a definitive cause for their high risk and severity. The strongest candidate-predictors include cockroach allergens, household smoking, air pollution, poor access to quality health care, and underutilization of inhaled anti-inflammatory medications. In particular, recent studies have shown that black and Latino children continue to misuse health care and medications because of lack of access to culturally sensitive pediatricians who understand their needs and barriers, which contributes to more severe, poorly controlled asthma. It has been suggested that interventions for minority asthmatic children focus on improving access to asthma medical homes that deliver culturally appropriate and relevant care tailored to the needs of the family, improving family-provider communication, and improving knowledge and acceptance of asthma clinical practice guidelines, particularly for providers who work in community-based clinics. PMID- 11106280 TI - Integration of genetics into medical practice: ethical, legal, and social perspective. PMID- 11106282 TI - Inherited iron overload disorders. AB - Iron is an essential nutrient that is highly toxic in excess. Normal iron balance is maintained primarily by regulation of intestinal absorption of the metal from the diet. Iron overload generally results from a chronic increase in intestinal absorption. During the past 5 years, it has become apparent that there are at least eight inherited disorders of iron metabolism characterized by the toxic accumulation of iron. This review provides an update for pediatricians on the clinical features and pathogenesis of these disorders. PMID- 11106281 TI - Genetics of Rett syndrome: properties of the newly discovered gene and pathobiology of the disorder. AB - The recent identification of mutations in the gene methyl-Cpg-binding protein-2 (MECP2) in girls with Rett syndrome (RS) has firmly established the molecular genetic basis of this unique, X-linked, dominant disorder and provides a dramatic conclusion to an intensive, decade-long search. This finding has ramifications far beyond establishing the gene for RS. Recent data indicate that the clinical phenotypes for MECP2 mutations range from mild disability in the mother of a girl with RS to rapidly progressive encephalopathy in her brother. Further, the pathobiology of MECP2 could be a prototype for other disorders of neurodevelopment. MECP2 encodes a methyl-CpG-binding protein (MeCP2), which is critical for transcriptional silencing of an as yet unknown number and type of genes responsible for the pathobiology of RS. As such, this discovery opens up completely new vistas as to fundamental neurobiologic processes, to disease mechanisms in the neurodevelopmental disabilities, and to potential new therapeutic strategies for RS and related disorders. PMID- 11106283 TI - Osteogenesis imperfecta: perspectives and opportunities. AB - The last 2 years have seen additions proposed to the very limited armamentarium of treatments for osteogenesis imperfecta. These include the use of bisphosphonates to decrease bone resorption, growth hormone to augment growth and collagen production, and bone marrow transplantation to create chimeras at the level of the collagen production unit in bone. Although there are optimistic proponents for each strategy, the lack of well-controlled studies and the absence of clearly defined objectives for therapy hinder clear assessment. PMID- 11106284 TI - Genetics of Hirschsprung disease. AB - Hirschsprung disease (HSCR), or congenital intestinal aganglionosis, is a relatively common disorder of neural crest migration. It has a strong genetic basis, although simple Mendelian inheritance is rarely observed. Hirschsprung disease is associated with several other anomalies and syndromes, and animal models for these conditions exist. Mutations in the RET gene are responsible for approximately half of familial cases and a smaller fraction of sporadic cases. Mutations in genes that encode RET ligands (GDNF and NTN); components of another signaling pathway (EDNRB, EDN3, ECE-1); and the transcription factor, SOX10, have been identified in HSCR patients. A subset of these mutations is associated with anomalies of pigmentation and/or hearing loss. For almost every HSCR gene, incomplete penetrance of the HSCR phenotype has been observed, probably due to genetic modifier loci. Thus, HSCR has become a model of a complex polygenic disorder in which the interplay of different genes is currently being elucidated. PMID- 11106285 TI - Office laboratory procedures, office economics, parenting and parent education, and urinary tract infection. AB - We again review four areas of interest to office-based pediatricians: office laboratory procedures, office economics, parenting and patient education, and urinary tract infections. Sean Elliott provides an update on the Clinical Laboratories Improvement Amendments (CLIA) and their impact of office practice. Eve Shapiro reviews office economics, focusing on measuring quality of care, use of performance data, costs of new technologies, and the impact of managed care on the medical marketplace. John Walter offers an update on parenting and parent education, with approaches to counseling families about overuse of antibiotics, teen pregnancy, hyperactivity, violence, and asthma. Richard Wahl reviews the recent research on urinary tract infection, with special attention paid to office diagnosis and management, longitudinal studies of children with urinary tract infections, and the controversy surrounding the American Academy of Pediatrics Task Force on Circumcision report. PMID- 11106286 TI - Bibliography. Current world literature. Neurology. PMID- 11106287 TI - Bibliography. Current world literature. Allergy, immunology and related disorders. PMID- 11106288 TI - Bibliography. Current world literature. Genetics. PMID- 11106289 TI - Use of diphtheria toxoid-tetanus toxoid-acellular pertussis vaccine as a five dose series. Supplemental recommendations of the Advisory Committee on Immunization Practices (ACIP). AB - Four vaccines containing diphtheria and tetanus toxoids and acellular pertussis vaccine (DTaP) are currently licensed in the United States for use among infants and young children. As of October 2000, two products, ACEL-IMUNE (a product of Lederle Laboratories) and Tripedia (Aventis Pasteur, Inc.) were licensed for the five-dose DTaP vaccination series. Two other vaccines, Infanrix (SmithKline Beecham Biologicals) and Certiva (North American Vaccine, Inc.) are licensed for the first four doses of the vaccination series, beginning with the primary series at ages 2, 4, and 6 months, and for completing the DTaP series among children who began the series with diphtheria and tetanus toxoids and whole-cell pertussis vaccine. This report supplements the statement from CDC's Advisory Committee on Immunization Practices regarding use of acellular pertussis vaccines and summarizes data regarding reactogenicity of acellular pertussis vaccines when administered as the fourth and fifth consecutive doses. Increases in the frequency and magnitude of local reactions at the injection site with increasing dose number have occurred for all currently licensed DTaP vaccines. Extensive swelling of the injected limb, sometimes involving the entire thigh or upper arm, after receipt of the fourth and fifth doses of DTaP vaccines has been demonstrated for multiple products from different manufacturers. Because data are insufficient regarding the safety, immunogenicity, and efficacy of using DTaP vaccines from different manufacturers in a mixed sequence, ACIP continues to recommend that, whenever feasible, the same brand of DTaP vaccine be used for all doses in the vaccination series. When the vaccine provider does not know or does not have available the type of DTaP vaccine previously administered, any of the licensed DTaP vaccines can be used to complete the vaccine series. PMID- 11106290 TI - Measurement and thermal modeling of high-Q piezoelectric resonators AB - This paper presents experimental data from the determination of the equivalent electrical circuit parameters of AT-cut quartz resonators and the measurement technique used. From an analysis of the experimental data it is possible to determine the equations which represent the variations of the parameters versus temperature. The models can then be used in simulation programs to determine the thermal behavior of the frequency of devices which use quartz crystal in their configuration and which require high stability. PSpice circuit simulation and experimental results are also included. PMID- 11106291 TI - Probability density estimation using incomplete data AB - An unsupervised method of learning probability density function parameters in the framework of mixture densities from incomplete data is developed. Unsupervised learning can be considered as learning from observations as they are received. In real-world processes, there are many imperfections in the observations. The Expectation-Maximization algorithm is used to iteratively find the maximum likelihood estimate of the missing values and the parameters of the probability density function. Reliability of the Expectation-Maximization algorithm and its convergence properties during the learning process from incomplete data are presented. Three examples of learning mixture probability density function parameters from an incomplete data set are presented and the boundaries of properly estimating the mixture probability density function parameters as the percentage of missing data increases are examined. PMID- 11106292 TI - Design of robust discrete control with desirable quadratic stability AB - In this paper, a design of robust discrete control with desirable quadratic stability is proposed. The design procedure is the extended discrete version of the continuous robust quadratic stabilization technique proposed by Gu et al. [K. Gu, Y.H. Chen, M.A. Zohdy, N.K. Loh, Quadratic stabilizability of uncertain systems: a two level optimization setup, Automatica 27 (1) (1991) 161-165.]. The effect of the sampling time selection, and the effect of the domain on the robustness, is examined. The presented algorithm is applied to a discrete mass spring system, and a discrete simplified car steering system to demonstrate the feasibility of the procedure, and the effect of the time sampling on the robustness. The robustness increases in both examples considered, as the sampling time decreases to some degree. PMID- 11106293 TI - Auto-tuning of TITO decoupling controllers from step tests AB - This paper considers auto-tuning of simple lead-lag decoupler plus decentralized PI/PID controllers for effective control of two-input and two-output (TITO) processes. A new robust identification method from step tests is presented first for SISO processes and then sequentially applied to TITO processes. The resulting 1st-order plus dead-time model is used for decoupler design and the 2nd-order one is used for decentralized PID sequential tuning. The simulation is given for illustration of the proposed tuning. PMID- 11106294 TI - Robust PID tuning strategy for uncertain plants based on the Kharitonov theorem AB - In this paper, the Kharitonov theorem for interval plants is exploited for the purpose of synthesizing a stabilizing controller. The aim here is to develop a controller to simultaneously stabilize the four Kharitonov-defined vortex polynomials. Different from the prevailing works, the controller is designed systematically and graphically through the search of a non-conservative Kharitonov region in the controller coefficient parameter plane. The region characterizes all stabilizing PID controllers that stabilize an uncertain plant. Thus the relationship between the Kharitonov region and the stabilizing controller parameters is manifest. Extensively, to further guarantee the system with certain robust safety margins, a virtual gain phase margin tester compensator is added. Stability analysis is carried out. The control system is proved to maintain robustness at least to the pre-specified margins. The synthesized controller with coefficients selected from the obtained non conservative Kharitonov region can stabilize the concerned uncertain plants and fulfill system specifications in terms of gain margins and phase margins. PMID- 11106295 TI - Identifying three-phase induction motor faults using artificial neural networks AB - This paper presents an artificial neural network (ANN) based technique to identify faults in a three-phase induction motor. The main types of faults considered are overload, single phasing, unbalanced supply voltage, locked rotor, ground fault, over-voltage and under-voltage. Three-phase currents and voltages from the induction motor are used in the proposed approach. A feedforward layered neural network structure is used. The network is trained using the backpropagation algorithm. The trained network is tested with simulated fault current and voltage data. Fault detection is attempted in the no fault to fault transition period. Off-line testing results on a 3 HP induction motor model show that the proposed ANN based method is effective in identifying various types of faults. PMID- 11106296 TI - A methodology for development of configurable remote access measurement system AB - A configurable remote access measurement system (CRAMS) is designed using an object oriented methodology (OOM) and implemented using the integration of an object oriented language, JAVA, a relational database management system, MS ACCESS, and an instrumentation software package (LabVIEW). OOM is a powerful technique that is used to manage the complexity of large systems. It allows for easy maintenance and upgrading of the developed systems. The main focus of this paper is to present a detailed procedure for the analysis, design and implementation of CRAMS. The functionality of CRAMS is demonstrated by creating a remotely accessible laboratory environment using a set of programmable and virtual instruments connected to a PC server. PMID- 11106297 TI - Preclinical psychopharmacology of AIDS-associated dementia: lessons to be learned from the cognitive psychopharmacology of other dementias. AB - Following a brief discussion of the epidemiology, underlying neuropathological mechanisms, neuropsychological symptoms and present treatment strategies of AIDS associated dementia (AAD), parallels are drawn between the longer standing research on drugs for the treatment of other cognitive disorders, particularly senile dementia, and ongoing efforts to develop psychopharmacological approaches for the treatment of the cognitive impairments in AAD. Important aspects of hypotheses designed to guide such a research are indicated with the help of a speculative, paradigmatic hypothesis concerning the role of cortical cholinergic inputs in AAD. Furthermore, aspects of validity of animal models, and cognition as a crucial intervening variable in the effects of potential treatments, are evaluated. PMID- 11106298 TI - The feline model of neuroAIDS: understanding the progression towards AIDS dementia. AB - Feline immunodeficiency virus (FIV) is a neurotropic lentivirus that produces a protracted state of immunodeficiency and encephalopathy in the cat. Recent evidence has shown several similarities to the natural progression of human immunodeficiency virus infection (HIV-1) associated degenerative effects on the central and peripheral nervous systems. Similar to HIV-1, FIV-induced encephalopathy neurovirulence is strain dependent, results in progressive immunodeficiency and increasing early peripheral but not brain viral load, preferentially affects the developing nervous system, produces quantifiable behavioural and neurophysiological impairment that is not directly linked to neuronal infectivity, and induces neuronal injury and loss both in vivo and in vitro. This paper highlights the cumulative scientific body of evidence supporting the use of the feline model of neuroAIDS. PMID- 11106299 TI - HIV dementia: the role of the basal ganglia and dopaminergic systems. AB - The clinical features of human immunodeficiency virus (HIV) dementia exhibit the hallmarks of a subcortical dementia. These features include psychomotor slowing, apathy, bradykinesia and altered posture and gait similar to those observed in advanced Parkinson's disease. The dementia has the hallmarks attributed to subcortical dementia. The exquisite sensitivity of many of these patients to dopamine receptor blockade suggested a profound and, perhaps, selective abnormality of striatal dopaminergic systems. Additional investigations, electrophysiological, pathological, virological, metabolic and radiological studies, indicate that the basal ganglia are a major target of HIV infection. In this review, we describe the evidence for involvement of basal ganglia and, in particular, the dopaminergic systems, in HIV dementia. We also suggest novel therapeutic strategies that may be beneficial in the treatment of this disorder. PMID- 11106300 TI - Neurotoxicity and dysfunction of dopaminergic systems associated with AIDS dementia. AB - Infection with the human immunodeficiency virus (HIV) selectively targets the basal ganglia resulting in loss of dopaminergic neurons. Although frequently asymptomatic, some patients may develop signs of dopamine deficiency de novo. Accordingly, they are highly susceptible to drugs that act on dopaminergic systems. Both neuroleptics and psychostimulants may exacerbate these symptoms. Experimental evidence suggests that viral proteins such as gp120 and Tat can cause toxicity to dopaminergic neurons, and this toxicity is synergistic with compounds such as methamphetamine and cocaine that also act on the dopaminergic system. In addition, other neurotransmitters that modulate dopaminergic function, such as glutamate and opioids, may also modify the susceptibility of the dopamine system to HIV. Therefore, a thorough understanding of the mechanisms that lead to this selective neurotoxicity of dopaminergic neurons would also likely lead to the development of therapeutic modalities for patients with HIV dementia. PMID- 11106301 TI - Neurobehavioural consequences of substance abuse and HIV infection. AB - Although our understanding of how human immunodeficiency virus (HIV)-related neurobehavioural deficits develop is nascent and preliminary, some clues have emerged which may clarify lingering uncertainties. In particular, HIV seems to yield brain dysfunction by mediating pathological changes upon neuronal function. HIV also compromises immunological integrity, thereby resulting in secondary infections that may further increase brain dysfunction. Notably, many individuals with HIV tend to be current or past abusers of drugs, and, in some cases, their drug use may have actually presented a pathway for initial HIV infection. Similar to HIV, many drugs tend to yield pathological changes upon neuronal function. Further paralleling HIV, some drugs seem to compromise immune function, which in turn may yield secondary detrimental effects upon the brain. Yet, despite the relatively high comorbidity rates of HIV infection and substance abuse, few investigations have addressed the potential interaction between these two factors upon neurobehavioural status. Towards this end, the present paper reviews the existing literature concerning neuropsychological effects of HIV and substance use, and suggests potential mechanisms whereby substance use may potentiate and exacerbate the onset and severity of neurobehavioural abnormalities in HIV infection. PMID- 11106303 TI - Methamphetamine and HIV-1: potential interactions and the use of the FIV/cat model. AB - The interaction of methamphetamine with human immunodeficiency virus (HIV), the aetiologic agent of Acquired Immune Deficiency Syndrome (AIDS), has not been thoroughly investigated. However, increasingly, a larger proportion of HIV infected individuals acquire the virus through methamphetamine use or are exposed to this drug during their disease course. In certain populations, there is a convergence of methamphetamine use and HIV-1 infection; yet our understanding of the potential effects that simultaneous exposure to these two agents have on disease progression is extremely limited. Studying the interactions between methamphetamine and lentivirus in people is difficult. To thoroughly understand methamphetamine's effects on lentivirus disease progression, an animal model that is both clinically relevant and easily manipulated is essential. In this report, we identified potential problems with methamphetamine abuse in individuals with a concurrent HIV-1 infection, described the Feline Immunodeficiency Virus (FIV)/cat model for HIV-1, and reported our early findings using this modelling system to study the interaction of methamphetamine and lentivirus infections. PMID- 11106302 TI - Morphine inhibits human microglial cell production of, and migration towards, RANTES. AB - The beta-chemokine RANTES has recently been implicated in the neuropathogenesis of the human immunodeficiency virus. Based upon previous studies of the effects of morphine on microglial cell production of cytokines and chemotaxis towards the activated complement component C5a, we tested the hypothesis that this opiate would alter the production of and migration towards RANTES by human microglia. Treatment of highly purified microglial cell cultures with morphine (10(-8)-10( 6) M) potently inhibited RANTES production by lipopolysaccharide- and interleukin 1beta-stimulated cells. Using a chemotaxis chamber to assess directed migration towards RANTES, treatment of microglial cells with morphine (10(-10)-10(-6) M) was found to suppress chemotaxis. The inhibitory effects of morphine on RANTES production and on chemotaxis were blocked by naloxone and beta-funaltrexamine, indicating that morphine mediated its suppressive effects via activation of microglial p-opioid receptors. Morphine's inhibitory effect on chemotaxis did not appear to be associated with an alteration in RANTES-induced [Ca2+]i mobilization. While the clinical significance of these in-vitro findings is unknown, they suggest that mu-opioid receptor agonists could alter certain neurodegenerative and inflammatory processes within the brain. PMID- 11106304 TI - Pharmacological frontiers in the treatment of AIDS dementia. AB - Even in the era of highly active antiretroviral therapy, AIDS dementia remains an important and devastating complication of human immunodeficiency virus (HIV-1) infection. Based on the 1997 AIDS case rate of 56 per 100 000 population in the USA, a reasonable estimated incidence of AIDS dementia is 3-8 per 100000, similar to that of multiple sclerosis. The pharmacology of AIDS dementia has been dominated by antiretroviral therapies, the best studied of which is azidothymidine. New and specific therapies are needed to treat and prevent brain injury in the setting of HIV infection. Rational therapy has been limited by the absence of large, adequate and well-controlled clinical trials using neuroprotective agents or those with disease-modifying potential, as well as by an incomplete understanding of the pathophysiology of AIDS dementia. In this review, a summary of evidence-based hypotheses of HIV-associated brain injury is followed by information on current nonantiretroviral therapeutic trials and their scientific rationale. PMID- 11106305 TI - Effects of acute tryptophan depletion on prepulse inhibition of the acoustic startle (eyeblink) response and the N1/P2 auditory evoked response in man. AB - Contraction of the orbicularis oculi muscle in response to a sudden loud sound (acoustic startle response) and the N1/P2 component of the auditory evoked potential are both attenuated when a brief low-intensity stimulus is presented 30 500 ms before the 'startle-eliciting' stimulus (prepulse inhibition). Here, we report the effect of acute tryptophan depletion on prepulse inhibition of these responses. Thirteen males (21-52 years) participated in two sessions separated by 7 days, in which they ingested a drink containing a mixture of amino-acids, which either included (+ TP) or did not include (- TP) tryptophan, according to a balanced double-blind design. Electromyographic (EMG) responses of the orbicularis oculi muscle and N1/P2 auditory evoked potentials were recorded in a 20-min session, 6 h after ingestion of the mixture. Subjects received 40 trials in which 1-kHz sounds were presented: (i) 40 ms, 115 dB ('pulse alone' trials) and (ii) 40 ms, 85 dB, followed after 120 ms by 40 ms, 115 dB ('prepulse/pulse' trials). Mean amplitudes of the EMG response and the N1/P2 potential were derived from the pulse-alone trials and, in each case, percentage prepulse inhibition was calculated. Plasma tryptophan levels were measured from blood samples taken before and 7 h after each treatment. Under the + TP condition, both the EMG response and the N1/P2 complex showed > 60% prepulse inhibition. The - TP condition was associated with (i) significant suppression of prepulse inhibition of the EMG response, with no significant change in response amplitude and (ii) reduction of the amplitude of the N1/P2 potential, with no significant change in prepulse inhibition of this response. Tryptophan levels rose by 90+/-15% under the + TP condition and fell by 81+/-3% under the - TP condition. The suppression of prepulse inhibition of the acoustic startle response under the - TP condition suggests that central 5-hydroxytryptaminergic mechanisms may be involved in regulating prepulse inhibition of this response. The lack of effect of tryptophan depletion on prepulse inhibition of the N1/P2 potential suggests that different mechanisms are involved in prepulse inhibition of the startle response and the N1/P2 complex. PMID- 11106306 TI - Does 'rebound mania' occur after stopping carbamazepine? A pilot study. AB - Withdrawal of lithium prophylaxis in patients with bipolar affective disorder has been shown to precipitate 'rebound mania', an effect which may negate its benefits in the poorly compliant. No studies have looked for similar effects on withdrawal of carbamazepine, an alternative and adjunctive prophylactic treatment. This retrospective study examined the effects of withdrawal of carbamazepine prophylaxis in patients with bipolar disorder. A systematic search for patients with bipolar disorder who stopped carbamazepine therapy whilst in remission was conducted, followed by case note review and interview. In a case series of six patients who stopped carbamazepine, four remained well for at least 3 months, one developed an episode of moderate depression and one remained well before resuming treatment after 1 month. None required admission or suffered a manic episode in the 3 months following cessation. This study does not support the existence of a carbamazepine 'rebound' effect. It raises the possibility that recurrence after stopping carbamazepine may be less severe than the 'rebound mania' seen on lithium withdrawal. If this is the case, it may be a better choice of mood stabilizer in the poorly compliant. To date, there is insufficient evidence on which to base this choice. There is a need to examine this issue further through larger prospective and experimental studies on the effects of anticonvulsant withdrawal. PMID- 11106307 TI - The serotonin uptake inhibitor citalopram reduces acute cardiovascular and vegetative effects of 3,4-methylenedioxymethamphetamine ('Ecstasy') in healthy volunteers. AB - MDMA (3,4-methylenedioxymethamphetamine) or 'Ecstasy' is a widely used recreational drug that produces a state of heightened mood but also cardiovascular and vegetative side-effects. In animals, MDMA releases serotonin and, to a lesser extent, dopamine and norepinephrine. The release of serotonin can be blocked by serotonin uptake inhibitors such as citalopram. It is unknown to what extent this mechanism is also responsible for the physiological side effects of MDMA seen in humans. We investigated the effect of citalopram pretreatment (40 mg i.v.) on vegetative and cardiovascular effects of MDMA (1.5 mg/kg p.o.) in a double-blind placebo-controlled study in 16 healthy volunteers. MDMA moderately increased blood pressure and heart rate, slightly elevated body temperature and produced a broad range of acute and short-term side-effects. Citalopram reduced all these MDMA-induced physiological changes except for body temperature. These findings suggest that physiological effects of MDMA in humans are partially due to an interaction of MDMA with the serotonin carrier and a subsequent release of serotonin. PMID- 11106308 TI - Comparison of donepezil-, tacrine-, rivastigmine- and metrifonate-induced central and peripheral cholinergically mediated responses in the rat. AB - There are now several acetylcholinesterase inhibitors in clinical use for the treatment of Alzheimer's disease, however, no systematic comparative studies of their central and peripheral cholinergic mediated effects in rats appear to have been reported. The present study investigated the dose-response characteristics of donepezil, tacrine, rivastigmine and metrifonate in inducing tremor, lacrimation, salivation and hypothermia and the duration of action of these compounds in Lister hooded rats. Data obtained were compared with the clinical observations on these drugs. Three doses of each compound were given orally to establish a dose-response curve for each behaviour, Tremor and lacrimation were scored, salivation was measured by weighing swabs applied to the mouth area and hypothermia was measured with a rectal probe. ED50 values were calculated for tremor. Using a just sub-maximal tremorigenic dose, the duration of response was examined. All four compounds produced dose-dependent increases in tremor and hypothermia. Only tacrine also produced marked salivation and lacrimation. The order of potency (ED50 value in micromol/kg) was rivastigmine (3.7), donepezil (18.0), tacrine (37.5), metrifonate (470). Tremor following tacrine (150 micromol/kg) and donepezil (20 micromol/kg) was prolonged (> 6 h) with a similar hypothermic response. The duration of these responses following metrifonate (777 micromol/kg) and rivastigmine (12.5 micromol/kg) did not exceed 3 h. Tacrine had poor selectivity for central (tremor) versus peripheral (salivation/lacrimation) effects compared to the other compounds. Donepezil also had a sustained duration of action. The data are consistent with clinical results and indicate that simple in-vivo models may assist in the selection of acetylcholinesterase inhibitors with a suitable response profile for use in the symptomatic treatment of Alzheimer's disease. PMID- 11106309 TI - Chronic administration of the cholesterol reducing drug gemfibrozil fails to alter 5-HT1A and 5-HT2A mediated receptor behaviours in rats. AB - Some studies have suggested that reductions in plasma cholesterol might be associated with suicidal behaviour. Serotonergic systems are thought to be involved in suicidal ideation and behaviour and links with altered 5-HT1A and 5 HT2A receptors have been proposed. We have therefore examined the effects of cholesterol reduction using gemfibrozil, upon 5-HT2A and 5-HT1A receptor-related behaviours in rats. Rats were treated chronically (57 days) with gemfibrozil (50 mg/kg p.o.) or gum acacia vehicle and challenged sequentially with the 5-HT1A agonist 8-hydroxy-d-n-aminopropyl tetralin, to elicit 5-HT1A syndrome behaviours, and the 5-HT2 receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)2-amino-propane to establish their head-shake frequency. Significant cholesterol reduction, within the clinical range, failed to induce any changes in either 5-HT1A or 5-HT2A mediated behaviours. These data suggest that cholesterol reduction fails to induce changes in 5-HT1A and 5-HT2A receptor tone suggesting that the reduction of plasma cholesterol, within the human clinical range, does not result in neuroplasticity of the 5-HT1A and 5-HT2A receptors in rats. PMID- 11106310 TI - Depressive relapse following acute tryptophan depletion in patients with major depressive disorder. AB - Acute tryptophan depletion (ATD) lowers serotonin synthesis and elicits depressive symptoms in some, though not all, remitted patients with major depressive disorder (MDD). In the present study, eight medication-free remitted patients with MDD, seasonal pattern, were tested twice, once following the ingestion of a tryptophan-containing mixture, once following ATD. ATD significantly increased Hamilton depression scores (p < 0.001). Four of the patients had a family history of psychiatric disorders: substance abuse (n = 4), mood disorders (n = 3) or Cluster B personality disorders (n = 3). The mood lowering response to ATD was significantly greater in those patients with, than without, affected relatives (p < 0.001). These preliminary findings (1) support the hypothesis that depressed states are related to disturbed serotonin neurotransmission and (2) suggest that depressive symptoms following ATD might identify a subgroup of patients at high genetic risk for disorders associated with affective lability and dysregulated impulse-control, conditions thought to be related to low serotonin neurotransmission. PMID- 11106311 TI - A double-dissociation of behavioural and event-related potential effects of two benzodiazepines with similar potencies. AB - This study was designed to explore the role of benzodiazepine affinity to benzodiazepine binding site on acute psychomotor, subjective and memory effects, as well as auditory Event Related Potential (ERP) latencies, in healthy volunteers. Two benzodiazepines with similar affinity to benzodiazepine binding sites, or potency, were compared: the atypical compound lorazepam (2.0 mg), which has been reported to impair priming, and a standard benzodiazepine, flunitrazepam (0.6 mg, 0.8 mg, 1.0 mg). The study followed a placebo-controlled, double-blind, parallel-group design. Sixty subjects completed a test battery before treatment and at theoretical peak plasma concentration of drugs. Lorazepam and 1.0 mg of flunitrazepam led to comparable alterations on psychomotor, subjective and auditory episodic memory measures. A double-dissociation was found for lorazepam and the equipotent dose of flunitrazepam (1.0 mg): lorazepam was more deleterious than flunitrazepam in time taken to identify fragmented shapes. Lorazepam also impaired direct and indirect stem-completion in comparison to placebo, but this effect was abolished when time to identify shapes was used as a covariate. By contrast, 1.0 mg of flunitrazepam prolonged auditory ERP latencies to a greater extent than lorazepam. High affinity to the benzodiazepine binding sites does not seem to explain the consistent lorazepam-induced impairment of indirect stem completion. Differences in impairment profile between the benzodiazepines employed may relate to the modality (visual or not) of the tasks used. PMID- 11106312 TI - Amelioration of specific working memory deficits by methylphenidate in a case of adult attention deficit/hyperactivity disorder. AB - Cognitive neuroscience has provided an extensive literature on the neuroanatomy and psychopharmacology of working memory. However, while it has been shown that children with attention deficit/hyperactivity disorder (AD/HD) have deficits in working memory, relatively little is known about working memory functions in adults diagnosed with AD/HD. Furthermore, it remains to be seen whether methylphenidate (Ritalin), which is used in the treatment of childhood AD/HD can improve performance deficits in adult AD/HD patients. We have used three paradigms of spatial working memory validated in cortical lesion patients, and psychopharmacological and neuroimaging studies, in order to examine the effects of methylphenidate administration in a case of an adult diagnosed with AD/HD. In the AD/HD patient at baseline testing, performance on a test of spatial recognition memory and on a task of self-ordered spatial working memory was shown to be impaired. Importantly, the impairments on the self-ordered spatial working memory task were ameliorated by an acute oral dose of methylphenidate (0.5 mg/kg). These findings provide insights into the possible neurochemical and neuroanatomical substrates of the action of methylphenidate in AD/HD and suggest a useful methodology for further research into this potentially debilitating disorder. PMID- 11106313 TI - Hypnopompic hallucinations with donepezil. AB - A case of hypnopompic hallucinations associated with donepezil is described. Electroencephalogram (EEG) and sleep EEG changes are common in Alzheimers Disease and acetylcholinesterase inhibitor drugs can affect rapid eye movement sleep and alertness. The importance of assessing sleep in patients treated with these drugs is discussed. PMID- 11106315 TI - Possible role of cellular immunity: a case of cellulitis. AB - On the basis of the observation that there was a "skip" area in an otherwise diffuse drug eruption where cellulitis had previously occurred, it is theorized that both delayed hypersensitivity type of dermatologic drug reaction and cellulitis share pathogenic mechanisms. PMID- 11106316 TI - Infection as a trigger of diabetic ketoacidosis in intensive care-unit patients. AB - We determined the prevalence and indicators of infection in intensive care unit (ICU) patients with diabetic ketoacidosis (DKA) by performing a retrospective analysis of 123 episodes of DKA (in 113 patients) managed in a medical ICU between 1990 and 1997. In univariate analysis, features associated with infection were female sex, neurological symptoms at admission, fever during the week before admission, a need for colloids, a high blood lactate level at admission, and lack of complete clearance of ketonuria within 12 h. Multivariate analysis identified 3 independent predictors of infection: female sex (odds ratio [OR], 2.31; confidence interval [CI], 1.05-5.35), neurological symptoms at admission (OR, 2.83; CI, 1.18-6.8), and lack of complete clearance of ketonuria within 12 h (OR, 3.73; CI, 1.58-9.09). Infection is the leading trigger of DKA in ICU patients. Neurological symptoms at admission and lack of complete clearance of ketonuria within 12 h are useful warning signals of infection. PMID- 11106317 TI - Renal involvement of human parvovirus B19 in an immunocompetent host. AB - Human parvovirus B19, which is most commonly known to cause erythema infectiosum in children, is also known to cause infection in adults, with complications ranging from a self-limited polyarthropathy in immunocompetent patients to hydrops fetalis in pregnant women, transient aplastic crises in patients with chronic hemolytic anemias, and chronic aplastic anemia in immunocompromised hosts. We describe a previously healthy immunocompetent woman who presented with manifestations of acute parvovirus B19 infection. PMID- 11106318 TI - The role of the clinical laboratory in managing chemical or biological terrorism. AB - BACKGROUND: Domestic and international acts of terrorism using chemicals and pathogens as weapons have recently attracted much attention because of several hoaxes and real incidents. Clinical laboratories, especially those affiliated with major trauma centers, should be prepared to respond rapidly by providing diagnostic tests for the detection and identification of specific agents, so that specific therapy and victim management can be initiated in a timely manner. As first-line responders, clinical laboratory personnel should become familiar with the various chemical or biological agents and be active participants in their local defense programs. APPROACH: We review the selected agents previously considered or used in chemical and biological warfare, outline their poisonous and pathogenic effects, describe techniques used in their identification, address some of the logistical and technical difficulties in maintaining such tests in clinical laboratories, and comment on some of the analytical issues, such as specimen handling and personal protective equipment. CONTENT: The chemical agents discussed include nerve, blistering, and pulmonary agents and cyanides. Biological agents, including anthrax and smallpox, are also discussed as examples for organisms with potential use in bioterrorism. Available therapies for each agent are outlined to assist clinical laboratory personnel in making intelligent decisions regarding implementation of diagnostic tests as a part of a comprehensive defense program. SUMMARY: As the civilian medical community prepares for biological and chemical terrorist attacks, improvement in the capabilities of clinical laboratories is essential in supporting counterterrorism programs designed to respond to such attacks. Accurate assessment of resources in clinical laboratories is important because it will provide local authorities with an alternative resource for immediate diagnostic analysis. It is, therefore, recommended that clinical laboratories identify their current resources and the extent of support they can provide, and inform the authorities of their state of readiness. PMID- 11106319 TI - Should we use carbohydrate-deficient transferrin instead of gamma glutamyltransferase for detecting problem drinkers? A systematic review and metaanalysis. AB - BACKGROUND: Carbohydrate-deficient transferrin (CDT) has been used as a test for excessive alcohol consumption in research, clinical, and medico-legal settings, but there remain conflicting data on its accuracy, with sensitivities ranging from <20% to 100%. We examined evidence of its benefit over a conventional and less expensive test, gamma-glutamyltransferase (GGT), and compared the accuracy of different CDT assay methods. METHODS: We performed a systematic review using summary ROC analysis of 110 studies prior to June 1998 on the use of CDT in the detection of alcohol dependence or hazardous/harmful alcohol use. RESULTS: We identified several potential sources of bias in studies. In studies examining CDT and GGT in the same subjects, subject characteristics were less likely to influence the comparison. In such paired studies, the original Pharmacia CDT assay was significantly more accurate than GGT, but the modified CDTect assay did not perform as well as the original and was not significantly better than GGT. The accuracy of the AXIS %CDT assay was statistically indistinguishable from modified CDTect. Several CDT assay methods appeared promising, in particular, liquid chromatography (chromatofocusing, HPLC, fast protein liquid chromatography) and isoelectric focusing, but there were insufficient paired studies from which to draw firm conclusions. CONCLUSIONS: In studies published before June 1998, the results obtained with commercially available CDT assays were not significantly better than GGT as markers of excessive alcohol use in paired studies. Further high-quality studies comparing CDTect (modified) and other CDT assays with GGT in the same subjects are needed. PMID- 11106320 TI - Fetal DNA in maternal plasma: biology and diagnostic applications. AB - BACKGROUND: Molecular analysis of plasma DNA during human pregnancy has led to the discovery that maternal plasma contains both fetal and maternal DNA. This valuable source of fetal DNA opens up new possibilities for noninvasive prenatal diagnosis. APPROACH: Published data from the last 3 years demonstrating the feasibility and utility of analyzing fetal DNA in maternal plasma are reviewed. CONTENT: The detection of fetal DNA in maternal plasma is much simpler and more robust than detecting fetal nucleated cells in maternal blood, and does not require prior enrichment. This approach has been shown to have application in the prenatal diagnosis of fetal rhesus D status, sex-linked disorders, and other paternally inherited genetic disorders. Abnormal fetal DNA concentrations in maternal plasma and serum have been found in common pregnancy-associated disorders, including preterm labor and preeclampsia, as well as in pregnancies complicated by fetal trisomy 21. After delivery, fetal DNA is cleared rapidly from maternal plasma, with a half-life in the order of minutes. These clearance kinetics exhibit an important difference from fetal cell clearance, where long term persistence has been demonstrated. SUMMARY: It has been only 3 years since fetal DNA was first detected in maternal plasma, and much remains to be learned about the biology of this phenomenon. In addition, additional diagnostic applications beyond those discussed here can be expected in the near future. PMID- 11106321 TI - Implementation of reference systems in laboratory medicine. PMID- 11106322 TI - Synergistic effect between apolipoprotein E and angiotensinogen gene polymorphisms in the risk for early myocardial infarction. AB - BACKGROUND: Several studies based on different populations worldwide have described an association between cardiovascular diseases and genetic variations in the apolipoprotein E (A:POE), angiotensinogen (A:GT), angiotensin receptor type 1 (A:T1R), and angiotensin-converting enzyme (A:CE) genes. In addition, there is growing evidence of an interaction between hypercholesterolemia and the renin-angiotensin system in the risk for hypertension and atherosclerosis. METHODS: To determine whether the DNA polymorphisms in A:POE (epsilon2, epsilon3, and epsilon4 alleles), A:GT (M235T), A:T1R (1166 A:/C:), and ACE (I:/D:) are associated with early onset of myocardial infarction (MI), we genotyped 220 patients and 200 controls <55 years of age. Patients and controls were males from the same homogeneous Caucasian population. Data concerning hypertension, diabetes, and tobacco consumption were recorded. The lipid profiles of patients and controls were also determined. RESULTS: APOE, ACE, AGT, and AT1R allele and genotype frequencies did not differ between patients and controls. None of these polymorphisms was related to the biochemical values in patients or controls. The frequency of individuals who were both APOE epsilon4 allele carriers and AGT-TT homozygotes was significantly higher in patients than in controls (11% vs 3.5%; P: = 0.0037). In patients, the frequency of epsilon4 carriers was significantly higher (P: <0.00001) in those who were AGT-TT (46%) than those who were AGT-MT/MM (14%). Mean cholesterol was significantly higher in AGT-TT + APOE epsilon34/44 patients than in the TM/MM + epsilon34/44 or TT + epsilon23/33 genotypes (P: = 0. 029). CONCLUSIONS: Our data suggest a synergistic effect between the APOE and AGT polymorphisms and early MI. The increased risk could be mediated in part through higher cholesterol concentrations among individuals who are AGT-TT + APOE epsilon4 allele carriers. PMID- 11106323 TI - Defects in pyrimidine degradation identified by HPLC-electrospray tandem mass spectrometry of urine specimens or urine-soaked filter paper strips. AB - BACKGROUND: Urinary concentrations of thymine, uracil, and their degradation products are useful indicators of deficiencies of enzymes of the pyrimidine degradation pathway. We describe a rapid, specific method to measure these concentrations to detect inborn errors of pyrimidine metabolism. METHODS: We used urine or urine-soaked filter-paper strips as samples and measured thymine, uracil, and their degradation products dihydrothymine, dihydrouracil, N:-carbamyl ss-aminoisobutyric acid, and N:-carbamyl-ss-alanine. Reversed-phase HPLC was combined with electrospray ionization tandem mass spectrometry, and detection was performed by multiple-reaction monitoring. Stable-isotope-labeled reference compounds were used as internal standards. RESULTS: All pyrimidine degradation products could be measured in one analytical run of 15 min. Detection limits were 0.4-4 micromol/L. The intraassay imprecision (CV) of urine samples with added compounds was 1.3-12% for liquid urines and 1. 0-10% for filter-paper extracts of the urines. The interassay imprecision (CV) was 3-11% (100-200 micromol/L). Recoveries were 89-99% at 100-200 micromol/L and 95-106% at 1 mmol/L in liquid urines, and 93-103% at 100-200 micromol/L and 100-106% at 1 mmol/L in filter paper samples. Correct identifications of deficiencies of the pyrimidine degrading enzymes were readily made with urine samples from patients with known defects. CONCLUSIONS: HPLC with electrospray ionization tandem mass spectrometry allows rapid testing for disorders of the pyrimidine degradation pathway, and filter-paper samples allow easy collection, transport, and storage of urine samples. PMID- 11106324 TI - Quantification of melanoma cell-specific MART-1 mRNA in peripheral blood by a calibrated competitive reverse transcription-PCR. AB - BACKGROUND: Reverse transcription-PCR (RT-PCR) amplification of melanoma cell specific mRNA can detect melanoma cells in the peripheral blood of patients with malignant melanoma. We present a method to quantify mRNA coding for the melanoma specific melanoma antigen recognized by T cells #1 (MART-1) in RNA isolated from peripheral blood. METHODS: To establish a calibration curve, we measured the concentration of MART-1 mRNA in SK-MEL-28 melanoma cells grown in vitro by competitive RT-PCR. Serial dilutions of these cells were used as calibrators in the assay. The assay was conducted by adding a fixed amount of a RNA internal standard to RNA isolated from either peripheral blood or the calibrators before RT-PCR amplification with MART-1 primers in a nested PCR design. The amount of MART-1 mRNA in blood samples was calculated from the calibration curve. RESULTS: Addition of melanoma cells grown in vitro to blood from healthy donors demonstrated that the method can detect a single SK-MEL-28 melanoma cell in 1 mL of blood (1.5 x 10(-21) mol MART-1 mRNA/mL). MART-1 mRNA was observed in 4 of 12 blood samples from patients with malignant melanoma, at concentrations of 3-18 x 10(-21) mol MART-1 mRNA/mL of blood. No MART-1 mRNA was detected in blood samples from 25 controls without malignant melanoma. Intra- and interassay CVs were 15% (n = 12; mean = 44 x 10(-21) mol MART-1 mRNA/mL) and 33% (15 samples analyzed in two different analytical runs; mean = 30 x 10(-21) mol MART-1 mRNA/mL), respectively. CONCLUSIONS: Our method is the first competitive RT-PCR assay for quantification of melanoma cells in blood samples that compensates for the variation of both the reverse transcription and PCR reactions. The method allows the inclusion of control samples for continuous quality assessment. PMID- 11106325 TI - K-ras point mutation detection in lung cancer: comparison of two approaches to somatic mutation detection using ARMS allele-specific amplification. AB - BACKGROUND: The use of sensitive molecular techniques to detect rare cells in a population is of increasing interest to the molecular pathologist, but detection limits often are poorly defined in any given molecular assay. We combined the approaches of real-time quantitative PCR with ARMS(TM) allele-specific amplification in a novel assay for detecting mutant K-ras sequences in clinical samples. METHODS: ARMS reactions were used to detect seven commonly occurring mutations in the K-ras oncogene. These mutations produce amino acid changes in codon 12 (Gly to Ala, Arg, Asp, Cys, Ser, or Val) and codon 13 (Gly to Asp). A control reaction was used to measure the total amount of amplifiable K-ras sequence in a sample so that the ratio of mutant to wild-type sequence could be measured. Quantitative data were confirmed for a selection of samples by an independent cloning and sequencing method. The assay was used to analyze 82 lung tumor DNA samples. RESULTS: The assay detected K-ras mutations in 44% of adenocarcinomas, which is equivalent to frequencies reported in the literature using ultrasensitive techniques. Forty-six percent of squamous carcinomas were also positive. The ratio of mutant sequence in the tumor DNA samples was 0.04 100%. CONCLUSIONS: The assay is homogeneous, with addition of tumor DNA sample being the only step before results are generated. The quantitative nature of the assay can potentially be used to define the analytical sensitivity necessary for any specified diagnostic application of K-ras (or other) point mutation detection. PMID- 11106326 TI - DNA base bulge vs unmatched end formation in probe-based diagnostic insertion/deletion genotyping: genotyping the UGT1A1 (TA)(n) polymorphism by real time fluorescence PCR. AB - BACKGROUND: Gilbert syndrome is a clinically inconsequential entity of mild unconjugated hyperbilirubinemia caused by an A(TA)(n)TAA insertion polymorphism (UGT1A1*28) in the promoter region of the gene coding for the enzyme UDP glucuronosyltransferase 1 (EC 2.4.1. 17; UGT1A1). Present methods for genotyping this polymorphism are laborious. METHODS: Hybridization probes were designed complementary to the wild type (TA)(6) and to alleles with (TA)(7) and (TA)(8) repeats in the promoter region. Melting points were measured in samples representing all currently known alleles with (TA)(5) to (TA)(8) repeats. Probe melting points were predicted with a thermodynamic nearest-neighbor model for Watson-Crick paired probes. The dominant secondary structures resulting from probe hybridization were predicted by thermodynamic free energy calculations. Alternatively samples were genotyped based on amplicon size resolved by high resolution polyacrylamide gel electrophoresis. RESULTS: Only short probes (22-24 bases) could be successfully used for genotyping this locus because of the very low stability of this TA repeat. Assays based on (TA)(7) or (TA)(8) genotype compatible hybridization probes effectively discriminated five to eight TA repeats. The consecutive use of two different detection probes was necessary for better discrimination of some heterozygous genotypes. All results were in concordance with the alternative genotyping method. Of 100 investigated Caucasians (50 males, 50 females), 9 (9%) were homozygous for the (TA)(7) allele. CONCLUSIONS: The presented method for genotyping the (TA)(n) promoter polymorphism of the UGT1A1 gene with the LightCycler has the potential to genotype all currently known (TA)(n) repeats in a single assay and is sensitive toward possible new genotypes. Our findings also show that thermodynamic calculations are of practical value for the design of hybridization probe assays for the genotyping of insertion/deletion polymorphisms. PMID- 11106327 TI - A modified, optimized kinetic photometric assay for the determination of blood coagulation factor XIII activity in plasma. AB - BACKGROUND: Blood coagulation factor XIII (FXIII) is a zymogen that is transformed into an active transglutaminase by thrombin and Ca(2+). FXIII plays an essential role in fibrin stabilization and in the protection of fibrin from proteolytic degradation. No convenient method has been available for the measurement of FXIII activity in plasma. The aim of the present study was to improve and optimize a kinetic photometric FXIII assay originally developed in our laboratory. METHODS: In the assay, FXIII was activated by thrombin and Ca(2+). Fibrin polymerization was prevented by an inhibitory tetrapeptide. Glycine-ethyl ester and a glutamine residue of a synthetic dodecapeptide served as acyl acceptor and acyl donor transglutaminase substrates, respectively. The amount of ammonia released during the reaction was monitored using glutamate dehydrogenase and NADPH. RESULTS: The use of a new glutamine substrate and optimization of activator and substrate concentrations increased sensitivity. Substitution of NADPH for NADH and introduction of an appropriate blank eliminated systemic overestimation of FXIII activity. The recovery of FXIII was 96%, the assay was linear up to 470 U/L, the detection limit was 1 U/L, and the imprecision (CV) was <8% even at very low FXIII activities. A reference interval of 108-224 U/L (69-143%) was established. The results correlated well with results obtained by an immunoassay specific for plasma FXIII. CONCLUSIONS: The optimized FXIII assay is a simple, rapid method for the diagnosis of inherited or acquired FXIII deficiencies and increased FXIII concentrations. It can be easily adapted to clinical chemistry analyzers. PMID- 11106328 TI - Use of a reference material proposed by the International Federation of Clinical Chemistry and Laboratory Medicine to evaluate analytical methods for the determination of plasma lipoprotein(a). AB - BACKGROUND: As part of the NIH/National Heart, Lung and Blood Institute Contract for the Standardization of Lipoprotein(a) [Lp(a)] Measurements, a study was performed in collaboration with the IFCC Working Group for the Standardization of Lp(a) Assays. The aims of the study, performed with the participation of 16 manufacturers and 6 research laboratories, were to evaluate the IFCC proposed reference material (PRM) for its ability to transfer an accuracy-based value to the immunoassay calibrators and to assess concordance in results among different methods. METHODS: Two different purified Lp(a) preparations with protein mass concentrations determined by amino acid analysis were used to calibrate the reference method. A Lp(a) value of 107 nmol/L was assigned to PRM. After uniformity of calibration was demonstrated in the 22 evaluated systems, Lp(a) was measured on 30 fresh-frozen sera covering a wide range of Lp(a) values and apolipoprotein(a) [apo(a)] sizes. RESULTS: The among-laboratory CVs for these samples (6-31%) were, in general, higher than those obtained for PRM (2.8%) and the quality-control samples (14%, 12%, and 9%, respectively), reflecting the broad range of apo(a) sizes in the 30 samples and the sensitivity of most methods to apo(a) size heterogeneity. Thus, although all of the assays were uniformly calibrated through the use of PRM, no uniformity in results was achieved for the isoform-sensitive methods. CONCLUSIONS: Linear regression analyses indicated that to various degrees, apo(a) size heterogeneity affects the outcome of the immunochemical methods used to measure Lp(a). We have also shown that the inaccuracy of Lp(a) values determined by methods sensitive to apo(a) size significantly affects the assessment of individual risk status for coronary artery disease. PMID- 11106329 TI - Determination of the designer drugs 3, 4-methylenedioxymethamphetamine, 3,4 methylenedioxyethylamphetamine, and 3,4-methylenedioxyamphetamine with HPLC and fluorescence detection in whole blood, serum, vitreous humor, and urine. AB - BACKGROUND: The popular designer drugs 3, 4-methylenedioxymethamphetamine (MDMA) and 3, 4-methylenedioxyethylamphetamine (MDEA) can be determined in serum, whole blood, and urine, but also in vitreous humor. The latter matrix is interesting when dealing with decomposed bodies in a toxicological setting. METHODS: After extraction, chromatographic separation was achieved on a narrow-bore C(18) column by gradient elution with fluorometric detection; results were confirmed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: The method was linear over the range of 2-1000 microg/L for whole blood, serum, and vitreous humor, and 0.1-5 mg/L for urine. Extraction recoveries were >70%, imprecision (CV) was 2.5-19%, and analytical recoveries were 95.5-104.4%. The limit of detection (LOD) and the limit of quantification (LOQ) were 0.8 and 2 microg/L, respectively, for whole blood, serum, and vitreous humor, and 2.5 microg/L and 0.1 mg/L, respectively, for urine. Excellent correlations between the quantitative LC-fluorescence and LC-MS/MS results were obtained. We found the following concentrations in a thanatochemical distribution study in rabbits: in serum, 5.3-685 microg/L for MDMA and from the LOQ to 14.5 microg/L for 3, 4 methylenedioxyamphetamine (MDA); in whole blood, 19.7-710 microg/L for MDMA and from the LOQ to 17.8 microg/L for MDA; in vitreous humor, 12.1-97.8 microg/L for MDMA and from the LOQ to 3.86 microg/L for MDA. In routine toxicological urine samples, concentrations ranged from LOQ to 14.62 mg/L for MDA, from LOQ to 157 mg/L for MDMA, and from LOQ to 32.54 mg/L for MDEA. CONCLUSIONS: The HPLC method described is sensitive, specific, and suitable for the determination of MDMA, MDEA, and MDA in whole blood, serum, vitreous humor, and urine. PMID- 11106330 TI - Erythrocyte folate analysis: saponin added during lysis of whole blood can increase apparent folate concentrations, depending on hemolysate pH. AB - BACKGROUND: The analysis of red cell folate (RCF) depends on complete hemolysis of erythrocytes, and it is assumed that complete hemolysis is achieved by 10-fold dilution of whole blood with hypotonic solutions of 10 g/L ascorbic acid/ascorbate. This report challenges this assumption. METHODS: The conventional method of erythrocyte lysis was modified to include saponin, a known effective hemolyzing agent. The influence of saponin was determined at various lysate pHs, using the microbiological (Lactobacillus rhamnosus) folate assay. The effect of saponin during lysate preparation was subsequently compared with either the effect of 30 s of sonication or a single 1-h freeze-thaw cycle. RESULTS: Saponin addition was found to increase assayable RCF up to ninefold, depending on lysate pH. Sonication of lysates had no effect, and freezing-thawing lysates once did not always guarantee complete hemolysis. Lysates created with 10 g/L ascorbic acid (a historically widely used diluent) without pH adjustment produced assayable folate concentrations significantly lower than optimal. CONCLUSIONS: A lysing agent should be incorporated into RCF assays to guarantee complete hemolysis. Ten-fold dilution of blood with 10 g/L ascorbic acid, without pH adjustment, produces lysates with pHs (pH 4.0) below the point (pH 4.7) at which hemoglobin can denature irreversibly. The optimum pH for hemolysates is approximately 5.0. PMID- 11106331 TI - High-speed detection of blood-borne hepatitis C virus RNA by single-tube real time fluorescence reverse transcription-PCR with the LightCycler. PMID- 11106332 TI - Analytical and clinical performance of the Immulite cardiac troponin I assay. PMID- 11106333 TI - Detection of multiple allergen-specific IgEs on microarrays by immunoassay with rolling circle amplification. PMID- 11106334 TI - Macroprolactin and the Roche Elecsys prolactin assay: characteristics of the reaction and detection by precipitation with polyethylene glycol. PMID- 11106335 TI - Rapid detection of the C3435T polymorphism of multidrug resistance gene 1 using fluorogenic hybridization probes. PMID- 11106336 TI - Comparison of two automated adrenocorticotropic hormone assays. PMID- 11106337 TI - Rapid homogeneous immunoassay for human ferritin in the Cobas Mira using colloidal gold as the reporter reagent. PMID- 11106338 TI - Quantification of hTERT mRNA and telomerase activity in bladder washings of patients with recurrent urothelial cell carcinomas. PMID- 11106339 TI - Prolonged hyperlipasemia attributable to a novel type of macrolipase. AB - BACKGROUND: We present the case of an 80-year-old woman who was admitted to hospital with an intermittent volvulus of the right colon. A total colectomy was performed. Initially, serum amylase and lipase increased concordantly, but after a few weeks amylase normalized (85 U/L), whereas lipase increased to 3764 U/L. This discrepancy and persistence of hyperlipasemia suggested a macromolecular form of lipase. METHODS: The nature of the macromolecular complex was studied using high-pressure liquid gel-permeation chromatography, affinity chromatography, (immuno)electrophoresis, and immunodiffusion. RESULTS: Gel permeation chromatography revealed a macrolipase, with a molecular mass >900 kDa, that contributed up to 56% of total serum lipase activity. Butanol extraction of the specimen did not alter the elution profile. The thermostabilities of pancreatic lipase and the macroform were similar, whereas activation energy (E:(a)) was lower in the macromolecular lipase (28 +/- 4 kJ. mol(-1). K(-1) vs 48 +/- 7 kJ. mol(-1). K(-1) (P: = 0.02). Agarose electrophoresis showed a broad band of lipase activity at the application site. Protein A-Sepharose affinity gel chromatography excluded IgG-linked lipase. Agarose electrophoresis and immunofixation excluded linkage to other immunoglobulins. Radial immunodiffusion did not show lipase activity in the immunoglobulin precipitation bands. Radial immunodiffusion with alpha(2)-macroglobulin (alpha(2)-MG) antibodies showed a diffuse spot of lipase activity within the precipitation band, suggesting a macromolecular association between lipase and alpha(2)-MG. Affinity gel chromatography against alpha(2)-MG showed lipase activity in the alpha(2)-MG bound fractions. CONCLUSION: This is the first report of a macrolipase in which an association between alpha(2)-MG and lipase is described. PMID- 11106340 TI - Detection of telomerase activity in urine as a tool for noninvasive detection of recurrent bladder tumors is poor and cannot be improved by timing of sampling. PMID- 11106341 TI - Erythrocyte folate does not accurately reflect folate status in sickle cell disease. PMID- 11106342 TI - Increases of creatine kinase MB and cardiac troponin T in serum of a patient with uterine leiomyosarcoma. PMID- 11106343 TI - Failure of assay to identify low cobalamin concentrations. PMID- 11106344 TI - Failure of assay to identify low cobalamin concentrations. Representatives of Bayer Diagnostics respond. PMID- 11106345 TI - Long-term stability of nelfinavir mesylate in human plasma. PMID- 11106346 TI - Discrimination of the nature of doping with 19-norsteroids through hair analysis. PMID- 11106347 TI - Detection of macroprolactin causing hyperprolactinemia in commercial assays for prolactin. PMID- 11106348 TI - MMP1 and MMP3 polymorphisms in promoter regions and cancer. PMID- 11106349 TI - Diagnosis and monitoring of hepatic injury. I. Performance characteristics of laboratory tests. AB - PURPOSE: To review information on performance characteristics for tests that are commonly used to identify acute and chronic hepatic injury. DATA SOURCES AND STUDY SELECTION: A MEDLINE search was performed for key words related to hepatic tests, including quality specifications, aminotransferases, alkaline phosphatase, gamma-glutamyltransferase, bilirubin, albumin, ammonia, and viral markers. Abstracts were reviewed, and articles discussing performance of laboratory tests were selected for review. Additional articles were selected from the references. Guideline Preparation and Review: Drafts of the guidelines were posted on the Internet, presented at the AACC Annual Meeting in 1999, and reviewed by experts. Areas requiring further amplification or literature review were identified for further analysis. Specific recommendations were made based on analysis of published data and evaluated for strength of evidence and clinical impact. The drafts were also reviewed by the Practice Guidelines Committee of the American Association for the Study of Liver Diseases and approved by the committee and the Association's Council. RECOMMENDATIONS: Although many specific recommendations are made in the guidelines, some summary recommendations are discussed here. Alanine aminotransferase is the most important test for recognition of acute and chronic hepatic injury. Performance goals should aim for total error of <10% at the upper reference limit to meet clinical needs in monitoring patients with chronic hepatic injury. Laboratories should have age-adjusted reference limits for enzymes in children, and gender-adjusted reference limits for aminotransferases, gamma-glutamyltransferase, and total bilirubin in adults. The international normalized ratio should not be the sole method for reporting results of prothrombin time in liver disease; additional research is needed to determine the reporting mechanism that best correlates with functional impairment. Harmonization is needed for alanine aminotransferase activity, and improved standardization for hepatitis C viral RNA measurements. PMID- 11106350 TI - Diagnosis and monitoring of hepatic injury. II. Recommendations for use of laboratory tests in screening, diagnosis, and monitoring. AB - PURPOSE: To review information on the use of laboratory tests in screening, diagnosis, and monitoring of acute and chronic hepatic injury. DATA SOURCES AND STUDY SELECTION: A MEDLINE search was performed for key words related to hepatic diseases, including acute hepatitis, chronic hepatitis, alcoholic hepatitis, cirrhosis, hepatocellular carcinoma, and etiologic causes. Abstracts were reviewed, and articles discussing use of laboratory tests selected for review. Additional articles were selected from the references. Guideline Preparation and Review: Drafts of the guidelines were posted on the Internet, presented at the AACC Annual Meeting in 1999, and reviewed by experts. Areas requiring further amplification or literature review were identified for further analysis. Specific recommendations were made based on analysis of published data and evaluated for strength of evidence and clinical impact. RECOMMENDATIONS: Although many specific recommendations are made in the guidelines, only some summary recommendations are listed here. In acute hepatic injury, prothrombin time and, to a lesser extent, total bilirubin are the best indicators of severity of disease. Although ALT is useful for detecting acute and chronic hepatic injury, it is not related to severity of acute hepatic injury and only weakly related to severity of chronic hepatic injury. Specific tests of viral markers should be the initial differential tests in both acute and chronic hepatic injury; when positive, they are also useful for monitoring recovery from hepatitis B and C. PMID- 11106351 TI - The clinical chemist. PMID- 11106352 TI - Neurofibromatosis type 2. AB - Neurofibromatosis type 2 is an often devastating autosomal dominant disorder which, until relatively recently, was confused with its more common namesake neurofibromatosis type 1. Subjects who inherit a mutated allele of the NF2 gene inevitably develop schwannomas, affecting particularly the superior vestibular branch of the 8th cranial nerve, usually bilaterally. Meningiomas and other benign central nervous system tumours such as ependymomas are other common features. Much of the morbidity from these tumours results from their treatment. It is now possible to identify the NF2 mutation in most families, although about 20% of apparently sporadic cases are actually mosaic for their mutation. As a classical tumour suppressor, inactivation of the NF2 gene product, merlin/schwannomin, leads to the development of both NF2 associated and sporadic tumours. Merlin/schwannomin associates with proteins at the cell cytoskeleton near the plasma membrane and it inhibits cell proliferation, adhesion, and migration. PMID- 11106354 TI - The ALX4 homeobox gene is mutated in patients with ossification defects of the skull (foramina parietalia permagna, OMIM 168500). AB - Foramina parietalia permagna (FPP) (OMIM 168500) is caused by ossification defects in the parietal bones. Recently, it was shown that loss of function mutations in the MSX2 homeobox gene on chromosome 5 are responsible for the presence of these lesions in some FPP patients. However, the absence of MSX2 mutations in some of the FPP patients analysed and the presence of FPP associated with chromosome 11p deletions in DEFECT 11 (OMIM 601224) patients or associated with Saethre-Chotzen syndrome suggests genetic heterogeneity for this disorder. Starting from a BAC/P1/cosmid contig of the DEFECT 11 region on chromosome 11, we have now isolated the ALX4 gene, a previously unidentified member of the ALX homeobox gene family in humans. Mutation analysis of the ALX4 gene in three unrelated FPP families without the MSX2 mutation identified mutations in two families, indicating that mutations in ALX4 could be responsible for these skull defects and suggesting further genetic heterogeneity of FPP. PMID- 11106353 TI - Chromatin modification and disease. PMID- 11106355 TI - Epigenotype-phenotype correlations in Beckwith-Wiedemann syndrome. AB - Beckwith-Wiedemann syndrome (BWS) is a model imprinting disorder resulting from mutations or epigenetic events involving imprinted genes at chromosome 11p15.5. Thus, germline mutations in CDKN1C, uniparental disomy (UPD), and loss of imprinting of IGF2 and other imprinted genes have been implicated. Many familial BWS cases have germline CDKN1C mutations. However, most BWS cases are sporadic and UPD or putative imprinting errors predominate in this group. We have identified previously a subgroup of sporadic cases with loss of imprinting (LOI) of IGF2 and epigenetic silencing of H19 proposed to be caused by a defect in a distal 11p15.5 imprinting control element (designated BWSIC1). However, many sporadic BWS patients show biallelic IGF2 expression in the presence of normal H19 methylation and expression patterns. This and other evidence suggested the existence of a further imprinting control element (BWSIC2) at 11p15. 5. Recently, we showed that a subgroup of BWS patients have loss of methylation (LOM) at a differentially methylated region (KvDMR1) within the KCNQ1 gene centromeric to the IGF2 and H19 genes. We have now analysed a large series of sporadic cases to define the frequency and phenotypic correlates of epigenetic abnormalities in BWS. LOM at KvDMR1 was detected by Southern analysis or a novel PCR based method in 35 of 69 (51%) sporadic BWS without UPD. LOM at KvDMR1 was often, but not invariably associated with LOI of IGF2. KvDMR1 LOM was not detected in BWS patients with putative BWSIC1 defects and cases with KvDMR1 LOM (that is, putative BWSIC2 defects) invariably had a normal H19 methylation pattern. The incidence of exomphalos in putative BWSIC2 defect patients was not significantly different from that in patients with germline CDKN1C mutations (20/29 and 13/15 respectively), but was significantly greater than that in patients with putative BWSIC1 defects (0/5, p=0.007) and UPD (0/22, p<0.0001). These findings are consistent with the hypothesis that LOM of KvDMR1 (BWSIC2 defect) results in epigenetic silencing of CDKN1C and variable LOI of IGF2. BWS patients with embryonal tumours have UPD or a BWSIC1 defect but not LOM of KvDMR1. This study has further shown how (1) variations in phenotypic expression of BWS may be linked to specific molecular subgroups and (2) molecular analysis of BWS can provide insights into mechanisms of imprinting regulation. PMID- 11106356 TI - Identification of cathepsin C mutations in ethnically diverse papillon-Lefevre syndrome patients. AB - INTRODUCTION: Papillon-Lefevre syndrome (PLS) is an autosomal recessive disorder characterised by palmoplantar keratoderma and severe, early onset periodontitis, which results from deficiency of cathepsin C activity secondary to mutations in the cathepsin C gene. To date, 13 different cathepsin C mutations have been reported in PLS patients, all of which are homozygous for a given mutation, reflecting consanguinity. AIM: To evaluate the generality of cathepsin C mutations in PLS, we studied an ethnically diverse group of 20 unrelated families. METHODS: Mutations were identified by direct automated sequencing of genomic DNA amplified for exonic regions and associated splice site junctions of the cathepsin C gene. Long range PCR was performed to determine the genomic structure of the cathepsin C gene. RESULTS: The cathepsin C gene spans over 46 kb, with six introns ranging in size from 1.6 to 22.4 kb. Eleven novel mutations and four previously reported mutations were identified in affected subjects from 14 families. Missense mutations were most common (9/15), followed by nonsense mutations (3/15), insertions (2/15), and deletions (1/15). Among these 14 probands, two were compound heterozygotes. Affected subjects with transgressions of the dermal lesions onto the knees or elbows or both had mutations in both the pro- and mature regions of the enzyme, although most were in the mature region. CONCLUSION: Mutations in the mature region of cathepsin C were more likely to be associated with the transgressions of the dermatological lesions, although the results were not statistically significant. A comprehensive list of all cathepsin C mutations described to date, representing 25 mutations from 32 families with PLS and related conditions, is also presented. PMID- 11106357 TI - Growth in North American white children with neurofibromatosis 1 (NF1). AB - OBJECTIVE: To analyse the distributions of and generate growth charts for stature and occipitofrontal circumference (OFC) in neurofibromatosis 1 (NF1) patients. DESIGN: Cross sectional database survey. SETTING: The National Neurofibromatosis Foundation International Database (NFDB) includes clinical information on NF1 patients from 14 participating centres in North America. SUBJECTS: A total of 569 white, North American, NF1 patients, 55% female and 45% male. MAIN OUTCOME MEASURES: Stature and OFC measurements of NF1 patients were compared to age and sex matched population norms using z score standardisation and centile curves. RESULTS: The distributions of stature and OFC are shifted and unimodal among NF1 patients; 13% of patients have short stature (>/=2 standard deviations below the population mean) and 24% have macrocephaly (OFC >/=2 standard deviations above the population mean). CONCLUSIONS: Alterations of stature and OFC are not limited to NF1 patients with frank short stature or macrocephaly. PMID- 11106358 TI - Cryptic von Hippel-Lindau disease: germline mutations in patients with haemangioblastoma only. AB - OBJECTIVES: Central nervous system haemangioblastoma (HAB) is a major feature of von Hippel-Lindau (VHL) disease, and it is estimated that about 30% of HAB patients have VHL disease. Consequently, it is widely recommended that sporadic HAB patients are screened for clinical and radiological features of VHL disease because of the risk of multiple tumours. We investigated the frequency of VHL germline mutations in patients with HAB only with no clinical or radiological evidence of VHL disease to define the role of molecular genetic analysis in the management of such patients. METHODS: Eighty four patients with a single HAB (23 Dutch, 61 UK) and four with multiple HAB (two Dutch, two UK) were studied by direct sequencing of the coding region and quantitative Southern blotting. RESULTS: A VHL germline mutation was found in three of 69 (4.3%) single HAB patients aged 50 years or less (three of 84 (3.6%) total single HAB patients). A germline VHL mutation was detected in a 44 year old woman with a solitary cerebellar HAB, as well as in four clinically unaffected close relatives, and in two single HAB cases presenting at the ages of 29 and 36 years. Germline VHL mutations were detected in two of four cases with multiple HAB. CONCLUSIONS: Early detection of VHL disease is important to reduce morbidity and mortality and therefore we recommend that, in addition to conventional clinical and radiological investigations, VHL gene mutation analysis should be offered to all HAB patients younger than 50 years. HAB patients aged >50 years will have a lower a priori risk of VHL disease and further data are required to evaluate the role of routine molecular genetic investigations in late onset HAB cases. The failure to detect germline VHL mutations in some patients with multiple HAB may indicate the presence of somatic mosaicism or additional HAB susceptibility genes. PMID- 11106359 TI - Spectrum of mutations in the MECP2 gene in patients with infantile autism and Rett syndrome. PMID- 11106360 TI - 2157delG: a frequent mutation in BRCA2 missed by PTT. PMID- 11106361 TI - A clinical assessment of neurofibromatosis type 1 (NF1) and segmental NF in Northern Finland. PMID- 11106362 TI - Exclusion of a disease relevant role of PAX4 in the aetiology of Silver-Russell syndrome: screening for mutations and determination of imprinting status. PMID- 11106363 TI - Recurrence risks in undiagnosed mental retardation. PMID- 11106364 TI - Bardet-Biedl and Cohen syndromes: differential diagnostic criteria. PMID- 11106365 TI - CHARGE association in a child with de Novo chromosomal aberration 46, X,der(X)t(X;2)(p22.1;q33) detected by spectral karyotyping. PMID- 11106366 TI - Identification of motifs in cholera toxin A1 polypeptide that are required for its interaction with human ADP-ribosylation factor 6 in a bacterial two-hybrid system. AB - The latent ADP-ribosyltransferase activity of cholera toxin (CT) that is activated after proteolytic nicking and reduction is associated with the CT A1 subunit (CTA1) polypeptide. This activity is stimulated in vitro by interaction with eukaryotic proteins termed ADP-ribosylation factors (ARFs). We analyzed this interaction in a modified bacterial two-hybrid system in which the T18 and T25 fragments of the catalytic domain of Bordetella pertussis adenylate cyclase were fused to CTA1 and human ARF6 polypeptides, respectively. Direct interaction between the CTA1 and ARF6 domains in these hybrid proteins reconstituted the adenylate cyclase activity and permitted cAMP-dependent signal transduction in an Escherichia coli reporter system. We constructed improved vectors and reporter strains for this system, and we isolated variants of CTA1 that showed greatly decreased ability to interact with ARF6. Amino acid substitutions in these CTA1 variants were widely separated in the primary sequence but were contiguous in the three-dimensional structure of CT. These residues, which begin to define the ARF interaction motif of CTA1, are partially buried in the crystal structure of CT holotoxin, suggesting that a change in the conformation of CTA1 enables it to bind to ARF. Variant CTA polypeptides containing these substitutions assembled into holotoxin as well as wild-type CTA, but the variant holotoxins showed greatly reduced enterotoxicity. These findings suggest functional interaction between CTA1 and ARF is required for maximal toxicity of CT in vivo. PMID- 11106367 TI - In search of ant ancestors. PMID- 11106368 TI - Inorganic polyphosphate kinase and adenylate kinase participate in the polyphosphate:AMP phosphotransferase activity of Escherichia coli. AB - Polyphosphate kinase (PPK), responsible for the processive synthesis of inorganic polyphosphate (polyP) from ATP in Escherichia coli, can transfer in reverse the terminal phosphate residue of polyP to ADP to yield ATP. PolyP also serves as a donor in a polyP:AMP phosphotransferase (PAP) activity observed in extracts of Acinetobacter johnsonii and Myxococcus xanthus. We have found that overexpression of the gene encoding PPK results in a large enhancement of PAP activity in E. coli. The PAP activity requires both PPK and adenylate kinase in equimolar amounts. PPK and adenylate kinase form a complex in the presence of polyphosphate. We discuss a phosphotransfer mechanism that involves both enzymes and enables polyP to be a phospho-donor to AMP. PMID- 11106369 TI - Polygyny, mate-guarding, and posthumous fertilization as alternative male mating strategies. AB - Alternative male mating strategies within populations are thought to be evolutionarily stable because different behaviors allow each male type to successfully gain access to females. Although alternative male strategies are widespread among animals, quantitative evidence for the success of discrete male strategies is available for only a few systems. We use nuclear microsatellites to estimate the paternity rates of three male lizard strategies previously modeled as a rock-paper-scissors game. Each strategy has strengths that allow it to outcompete one morph, and weaknesses that leave it vulnerable to the strategy of another. Blue-throated males mate-guard their females and avoid cuckoldry by yellow-throated "sneaker" males, but mate-guarding is ineffective against aggressive orange-throated neighbors. The ultradominant orange-throated males are highly polygynous and maintain large territories; they overpower blue-throated neighbors and cosire offspring with their females, but are often cuckolded by yellow-throated males. Finally, yellow-throated sneaker males sire offspring via secretive copulations and often share paternity of offspring within a female's clutch. Sneaker males sire more offspring posthumously, indicating that sperm competition may be an important component of their strategy. PMID- 11106370 TI - A maize sesquiterpene cyclase gene induced by insect herbivory and volicitin: characterization of wild-type and mutant alleles. AB - Plants can defend themselves from herbivorous insects by emitting volatile chemical signals that attract natural enemies of the herbivore. For example, maize seedlings attacked by beet armyworm larvae (Spodoptera exigua) produce a mixture of terpenoid and indole volatiles that serve to attract parasitic wasps. A key step in terpenoid biosynthesis is the conversion of acyclic prenyl diphosphates to terpenoid compounds by specific terpenoid synthases (cyclases). We have cloned a maize sesquiterpene cyclase gene, stc1, by transposon tagging and have identified two deletion mutations of the gene. The stc1 gene is located on chromosome 9S and does not seem to have a closely related ortholog in the maize genome. It is induced 15- to 30-fold in maize leaves by beet armyworm larvae feeding or by application of purified volicitin, the insect-derived elicitor, at a mechanically wounded site. stc1 induction is systemic, because undamaged leaves of the same plant show a similar increase in stc1 transcription. Analysis of volatiles from volicitin-treated seedlings revealed that a major naphthalene-based sesquiterpene was present in wild-type seedlings but absent in the Ac-insertion and x-ray-deletion mutants. Therefore, we have identified a maize gene that responds to caterpillar herbivory by producing a chemical defense signal that most likely serves to attract natural enemies of the herbivore. PMID- 11106371 TI - Water-borne cues induce chemical defense in a marine alga (Ascophyllum nodosum). AB - It is well known that herbivores can induce chemical defenses in terrestrial vascular plants, but few examples of inducible production of defense chemicals have been reported for aquatic macrophytes. Furthermore, it is well established that water-borne chemical cues from predators or predator-wounded conspecifics can induce defensive changes of aquatic prey animals, but no such communication between aquatic herbivores and seaweeds has been reported. Here we show that water-borne cues from actively feeding herbivorous gastropods, flat periwinkles (Littorina obtusata), can serve as external signals to induce production of defense chemicals (phlorotannins) in unharmed individuals of seaweeds, knotted wrack (Ascophyllum nodosum), and that the increased levels of defense chemicals deter further feeding by periwinkles. Because seaweeds have poorly developed internal-transport systems and may not be able to elicit systemic-induced chemical defenses through conveyance of internal signals, this mechanism ensures that seaweeds can anticipate future periwinkle attacks without receiving direct damage by herbivores. PMID- 11106372 TI - Antisense imaging of gene expression in the brain in vivo. AB - Antisense radiopharmaceuticals could be used to image gene expression in the brain in vivo, should these polar molecules be made transportable through the blood-brain barrier. The present studies describe an antisense imaging agent comprised of an iodinated peptide nucleic acid (PNA) conjugated to a monoclonal antibody to the rat transferrin receptor by using avidin-biotin technology. The PNA was a 16-mer antisense to the sequence around the methionine initiation codon of the luciferase mRNA. C6 rat glioma cells were permanently transfected with a luciferase expression plasmid, and C6 experimental brain tumors were developed in adult rats. The expression of the luciferase transgene in the tumors in vivo was confirmed by measurement of luciferase enzyme activity in the tumor extract. The [(125)I]PNA conjugate was injected intravenously in anesthetized animals with brain tumors and killed 2 h later for frozen sectioning of brain and film autoradiography. No image of the luciferase gene expression was obtained after the administration of either the unconjugated antiluciferase PNA or a PNA conjugate that was antisense to the mRNA of a viral transcript. In contrast, tumors were imaged in all rats administered the [(125)I]PNA that was antisense to the luciferase sequence and was conjugated to the targeting antibody. In conclusion, these studies demonstrate gene expression in the brain in vivo can be imaged with antisense radiopharmaceuticals that are conjugated to a brain drug targeting system. PMID- 11106373 TI - Detection of glutamic acid decarboxylase-activated T cells with I-Ag7 tetramers. AB - CD4(+) T cells selected by the type 1 diabetes associated class II MHC I-A(g7) molecules play a critical role in the disease process. Multivalent MHC/peptide tetramers have been used to directly detect antigen-specific T cells. Detection of autoantigen-activated CD4(+) T cells with tetramers should be very helpful in the study of the roles of these cells in diabetes. We report here the generation of tetramers of I-A(g7) covalently linked to two glutamic acid decarboxylase (GAD) peptides and the detection of GAD peptide-activated T cells from nonobese diabetic (NOD) mice. The I-A(g7) heterodimers can form stable complexes with a covalently bound GAD peptide and can stimulate antigen specific T cells. Furthermore, I-A(g7)/GAD peptide tetramer can detect most if not all of the antigen-specific CD4(+) T cells from immunized NOD mice. Antigen-specific T cells detected by the tetramers can up-regulate their CD4 expression on the cell surface after being restimulated with the GAD peptides in vitro. In contrast, the tetramers can detect a percentage of T cells in lymph nodes and spleens and T cells infiltrating islets from nonimmunized mice that is not significantly above the background. Therefore, T cells specific for the GAD peptides are present in NOD mice at a frequency too low to be detected, but immunization of NOD mice can facilitate their detection by tetramers. PMID- 11106374 TI - Silent information regulator 2 family of NAD- dependent histone/protein deacetylases generates a unique product, 1-O-acetyl-ADP-ribose. AB - Conflicting reports have suggested that the silent information regulator 2 (SIR2) protein family employs NAD(+) to ADP-ribosylate histones [Tanny, J. C., Dowd, G. J., Huang, J., Hilz, H. & Moazed, D. (1999) Cell 99, 735-745; Frye, R. A. (1999) Biochem. Biophys. Res. Commun. 260, 273-279], deacetylate histones [Landry, J., Sutton, A., Tafrov, S. T., Heller, R. C., Stebbins, J., Pillus, L. & Sternglanz, R. (2000) Proc. Natl. Acad. Sci. USA 97, 5807-5811; Smith, J. S., Brachmann, C. B., Celic, I., Kenna, M. A., Muhammad, S., Starai, V. J., Avalos, J. L., Escalante-Semerena, J. C., Grubmeyer, C., Wolberger, C. & Boeke, J. D. (2000) Proc. Natl. Acad. Sci. USA 97, 6658-6663], or both [Imai, S., Armstrong, C. M., Kaeberlein, M. & Guarente, L. (2000) Nature (London) 403, 795-800]. Uncovering the true enzymatic function of SIR2 is critical to the basic understanding of its cellular function. Therefore, we set out to authenticate the reaction products and to determine the intrinsic catalytic mechanism. We provide direct evidence that the efficient histone/protein deacetylase reaction is tightly coupled to the formation of a previously unidentified acetyl-ADP-ribose product (1-O-acetyl-ADP ribose). One molecule of NAD(+) and one molecule of acetyl-lysine are readily catalyzed to one molecule of deacetylated lysine, nicotinamide, and 1-O-acetyl ADP-ribose. A unique reaction mechanism involving the attack of enzyme-bound acetate or the direct attack of acetyl-lysine on an oxocarbenium ADP-ribose intermediate is proposed. We suggest that the reported histone/protein ADP ribosyltransferase activity is a low-efficiency side reaction that can be explained through the partial uncoupling of the intrinsic deacetylation and acetate transfer to ADP-ribose. PMID- 11106375 TI - Identification of brain-specific and imprinted small nucleolar RNA genes exhibiting an unusual genomic organization. AB - We have identified three C/D-box small nucleolar RNAs (snoRNAs) and one H/ACA-box snoRNA in mouse and human. In mice, all four snoRNAs (MBII-13, MBII-52, MBII-85, and MBI-36) are exclusively expressed in the brain, unlike all other known snoRNAs. Two of the human RNA orthologues (HBII-52 and HBI-36) share this expression pattern, and the remainder, HBII-13 and HBII-85, are prevalently expressed in that tissue. In mice and humans, the brain-specific H/ACA box snoRNA (MBI-36 and HBI-36, respectively) is intron-encoded in the brain-specific serotonin 2C receptor gene. The three human C/D box snoRNAs map to chromosome 15q11-q13, within a region implicated in the Prader-Willi syndrome (PWS), which is a neurogenetic disease resulting from a deficiency of paternal gene expression. Unlike other C/D box snoRNAs, two snoRNAs, HBII-52 and HBII-85, are encoded in a tandemly repeated array of 47 or 24 units, respectively. In mouse the homologue of HBII-52 is processed from intronic portions of the tandem repeats. Interestingly, these snoRNAs were absent from the cortex of a patient with PWS and from a PWS mouse model, demonstrating their paternal imprinting status and pointing to their potential role in the etiology of PWS. Despite displaying hallmarks of the two families of ubiquitous snoRNAs that guide 2'-O ribose methylation and pseudouridylation of rRNA, respectively, they lack any telltale rRNA complementarity. Instead, brain-specific C/D box snoRNA HBII-52 has an 18-nt phylogenetically conserved complementarity to a critical segment of serotonin 2C receptor mRNA, pointing to a potential role in the processing of this mRNA. PMID- 11106376 TI - Femtosecond direct observation of charge transfer between bases in DNA. AB - Charge transfer in supramolecular assemblies of DNA is unique because of the notion that the pi-stacked bases within the duplex may mediate the transport, possibly leading to damage and/or repair. The phenomenon of transport through pi stacked arrays over a long distance has an analogy to conduction in molecular electronics, but the mechanism still needs to be determined. To decipher the elementary steps and the mechanism, one has to directly measure the dynamics in real time and in suitably designed, structurally well characterized DNA assemblies. Here, we report our first observation of the femtosecond dynamics of charge transport processes occurring between bases within duplex DNA. By monitoring the population of an initially excited 2-aminopurine, an isomer of adenine, we can follow the charge transfer process and measure its rate. We then study the effect of different bases next to the donor (acceptor), the base sequence, and the distance dependence between the donor and acceptor. We find that the charge injection to a nearest neighbor base is crucial and the time scale is vastly different: 10 ps for guanine and up to 512 ps for inosine. Depending on the base sequence the transfer can be slowed down or inhibited, and the distance dependence is dramatic over the range of 14 A. These observations provide the time scale, and the range and efficiency of the transfer. The results suggest the invalidity of an efficient wire-type behavior and indicate that long range transport is a slow process of a different mechanism. PMID- 11106377 TI - On the origin of metacentric, attached-X (A-X) chromosomes in Drosophila melanogaster males. AB - We describe here the isolation and cytogenetic characterization of a mutation inseparabile which generates in males a high frequency of A-X females. The mutation, segregating in low frequency in a laboratory stock, maps to cytological location 82F7-11 in the third chromosome. The mutation acts premeiotically in the male germ line. Disrupting the X chromosome centromeric heterochromatin suppresses the formation of A-X chromosome, implying that the mutation is involved in chromatid cohesion. The inseparabile mutation also affects disjunction of the chromosome 4 in males. We suspect that the mutation was responsible for the original A-X female found by L. V. Morgan in 1921. PMID- 11106378 TI - Insect herbivory accelerates nutrient cycling and increases plant production. AB - Ecologists hold two views about the role of herbivory in ecosystem dynamics. First, from a food web perspective in population/community ecology, consumption by herbivores reduces plant abundance. Second, from a nutrient cycling perspective in ecosystem ecology, herbivory sometimes slows down cycling, which decreases plant abundance, but at other times speeds up cycling, which possibly increases plant abundance. The nutrient cycling perspective on herbivory has been experimentally addressed more thoroughly in aquatic systems than in terrestrial systems. We experimentally examined how grasshoppers influence nutrient cycling and, thereby, plant abundance and plant species composition over a period of 5 years. We examined how grasshoppers influence nutrient (nitrogen) cycling (i) by their excrement, (ii) by changing the abundance of and the decomposition rate of plant litter, and (iii) by both. Grasshoppers may speed up nitrogen cycling by changing the abundance and decomposition rate of plant litter, which increases total plant abundance (up to 32.9 g/m(2) or 18%), especially, the abundance of plants that are better competitors when nitrogen is more available. However, whether grasshoppers enhance plant abundance depends on how much they consume. Consequently, ecosystems and food web perspectives are not mutually exclusive. Finally, under some conditions, grasshoppers may decrease nutrient cycling and plant abundance. PMID- 11106379 TI - Femtosecond studies of protein-ligand hydrophobic binding and dynamics: human serum albumin. AB - In this contribution, we report studies of the nature of the dynamics and hydrophobic binding in protein-ligand complexes of human serum albumin with 2-(2' hydroxyphenyl)-4-methyloxazole. With femtosecond time resolution, we examined the orientational motion of the ligand, its intrinsic nuclear motions, and the lifetime changes in the hydrophobic phase. For comparisons, with similar but chemical nanocavities, we also studied the same ligand in micelles and cyclodextrins. The hydrophobic interactions in the binding crevice are much stronger than those observed in cyclodextrins and micelles. The confined geometry restrains the nonradiative decay and significantly lengthens the excited-state lifetime. The observed dynamics over the femtosecond-to-nanosecond time scale indicate that the binding structure is rigid and the local motions of the ligand are nearly "frozen" in the protein. Another major finding is the elucidation of the directed dynamics by the protein. Proton transfer and intramolecular twisting of 2-(2'-hydroxyphenyl)-4-methyloxazole were observed to evolve along two routes: one involves the direct stretching motion in the molecular plane (approximately 200 fs) and is not sensitive to the environment; the second, less dominant, is related to the twisting motion (approximately 3 ps) of the two heterocyclic rings and drastically slows down in the protein hydrophobic pocket. PMID- 11106380 TI - Maternal germ-line transmission of mutant mtDNAs from embryonic stem cell-derived chimeric mice. AB - We report a method for introducing mtDNA mutations into the mouse female germ line by means of embryonic stem (ES) cell cybrids. Mitochondria were recovered from the brain of a NZB mouse by fusion of synaptosomes to a mtDNA-deficient (rho degrees ) cell line. These cybrids were enucleated and the cytoplasts were electrofused to rhodamine-6G (R-6G)-treated female ES cells. The resulting ES cell cybrids permitted transmission of the NZB mtDNAs through the mouse maternal lineage for three generations. Similarly, mtDNAs from a partially respiratory deficient chloramphenicol-resistant (CAP(R)) cell line also were introduced into female chimeric mice and were transmitted to the progeny. CAP(R) chimeric mice developed a variety of ocular abnormalities, including congenital cataracts, decreased retinal function, and hamaratomas of the optic nerve. The germ-line transmission of the CAP(R) mutation resulted in animals with growth retardation, myopathy, dilated cardiomyopathy, and perinatal or in utero lethality. Skeletal and heart muscle mitochondria of the CAP(R) mice were enlarged and atypical with inclusions. This mouse ES cell-cybrid approach now provides the means to generate a wide variety of mouse models of mitochondrial disease. PMID- 11106381 TI - Chemical synthesis and spontaneous folding of a multidomain protein: anticoagulant microprotein S. AB - Because of recent high-yield native ligation techniques, chemical synthesis of larger multidomain bioactive proteins is rapidly coming within reach. Here we describe the total chemical synthesis of a designed "microprotein S," comprising the gamma-carboxyglutamic acid-rich module, the thrombin-sensitive module, and the first epidermal growth factor-like module of human plasma protein S (residues 1-116). Synthetic microprotein S expressed anticoagulant cofactor activity for activated protein C in the down-regulation of blood coagulation, and the anticoagulant activity of microprotein S was not neutralized by C4b-binding protein, a natural inhibitor of native protein S in plasma. The correct folding of this complex multidomain protein was enhanced compared with individual modules because the gamma-carboxyglutamic acid-rich module and the thrombin-sensitive module markedly facilitated correct folding of the first epidermal growth factor like module compared with folding of the first epidermal growth factor-like module alone. These results demonstrate that total chemical synthesis of proteins offers an effective way to generate multidomain biologically active proteins. PMID- 11106383 TI - Algebraic orbifold conformal field theories. AB - The unitary rational orbifold conformal field theories in the algebraic quantum field theory and subfactor theory framework are formulated. Under general conditions, it is shown that the orbifold of a given unitary rational conformal field theory generates a unitary modular category. Many new unitary modular categories are obtained. It is also shown that the irreducible representations of orbifolds of rank one lattice vertex operator algebras give rise to unitary modular categories and determine the corresponding modular matrices, which has been conjectured for some time. PMID- 11106382 TI - Highly conserved glutamic acid in the extracellular IV-V loop in rhodopsins acts as the counterion in retinochrome, a member of the rhodopsin family. AB - Retinochrome is a member of the rhodopsin family having a chromophore retinal and functioning as a retinal photoisomerase in squid photoreceptor cells. Unlike vertebrate rhodopsins, but like many invertebrate rhodopsins, retinochrome does not have a glutamic acid at position 113 to serve as a counterion for the protonated retinylidene Schiff base. Here we investigated possible counterions in retinochrome by site-specific mutagenesis. Our results showed that the counterion is the glutamic acid at position 181, at which almost all the pigments in the rhodopsin family, including vertebrate and invertebrate rhodopsins, have a glutamic or aspartic acid. The remarkable exceptions are the long-wavelength visual pigments that have a histidine that, together with a nearby lysine, serves as a chloride-binding site. Replacement of Glu-181 of bovine rhodopsin with Gln caused a 10-nm red-shift of absorption maximum. Because the position at 181 is in the extracellular loop connecting the transmembrane helices VI and V, these results demonstrate the importance of this loop to function for spectral tuning in the rhodopsin family. PMID- 11106384 TI - The structure of aspartyl dipeptidase reveals a unique fold with a Ser-His-Glu catalytic triad. AB - The three-dimensional structure of Salmonella typhimurium aspartyl dipeptidase, peptidase E, was solved crystallographically and refined to 1.2-A resolution. The structure of this 25-kDa enzyme consists of two mixed beta-sheets forming a V, flanked by six alpha-helices. The active site contains a Ser-His-Glu catalytic triad and is the first example of a serine peptidase/protease with a glutamate in the catalytic triad. The active site Ser is located on a strand-helix motif reminiscent of that found in alpha/beta-hydrolases, but the polypeptide fold and the organization of the catalytic triad differ from those of the known serine proteases. This enzyme is a member of a family of serine hydrolases and appears to represent a new example of convergent evolution of peptidase activity. PMID- 11106385 TI - Acyl-CoA hydrolysis by the high molecular weight protein 1 subunit of yersiniabactin synthetase: mutational evidence for a cascade of four acyl-enzyme intermediates during hydrolytic editing. AB - Yersiniabactin (Ybt) synthetase is a three-subunit, 17-domain [7 domains in high molecular weight protein (HMWP)2, 9 in HMWP1, and 1 in YbtE] enzyme producing the virulence-conferring siderophore yersiniabactin in Yersinia pestis. The 350-kDa HMWP1 subunit contains a polyketide synthase module (KS-AT-MT(2)-KR-ACP) and a nonribosomal peptide synthetase module (Cy(3)-MT(3)-PCP(3)-TE). The full-length HMWP1 was heterologously overexpressed in Escherichia coli and purified to near homogeneity. The purified HMWP1 showed thioesterase activity toward acyl-CoAs, such as acetyl-CoA, benzoyl-CoA, and malonyl-CoA, with saturation kinetics and relative catalytic efficiencies of 172:50:1. A chain-releasing thioesterase (TE) activity is ascribed to the C-terminal TE domain, and this was substantiated by the fact that acyl-N-acetylcysteamines were hydrolyzed by the didomain PCP(3)-TE fragment of HMWP1. However, PCP(3)-TE failed to hydrolyze any of the acyl-CoAs, suggesting the TE domain does not recognize CoA moiety, thus the acyl-CoA hydrolysis by HMWP1 must involve other domains. Ser-to-Ala mutants in each of the AT, ACP, PCP(3), and TE domains reduced hydrolysis rates of the two fastest substrates, acetyl-CoA and benzoyl-CoA, by more than two orders of magnitude. Thus, the acyl-CoA hydrolysis activity requires 4 of the 9 domains of HMWP1, and it is consistent with autoacylation of the AT domain active site serine and subsequent passage of the itinerant acyl chain from AT to ACP to PCP(3) to the TE domain, a cascade of four sequential acyl-enzyme intermediates, for hydrolytic turnover. This could represent an editing pathway for this polyketide synthase/nonribosomal peptide synthetase assembly line. PMID- 11106386 TI - Species-specific polyamines from diatoms control silica morphology. AB - Biomineralizing organisms use organic molecules to generate species-specific mineral patterns. Here, we describe the chemical structure of long-chain polyamines (up to 20 repeated units), which represent the main organic constituent of diatom biosilica. These substances are the longest polyamine chains found in nature and induce rapid silica precipitation from a silicic acid solution. Each diatom is equipped with a species-specific set of polyamines and silica-precipitating proteins, which are termed silaffins. Different morphologies of precipitating silica can be generated by polyamines of different chain lengths as well as by a synergistic action of long-chain polyamines and silaffins. PMID- 11106387 TI - Cell injury releases endogenous adjuvants that stimulate cytotoxic T cell responses. AB - General immunostimulants (adjuvants) are essential for generating immunity to many antigens. In bacterial infections, adjuvants are provided by components of the microorganism, e.g., lipopolysaccharide. However, it is unclear what provides the adjuvant effect for immune responses that are generated to tumors and many viruses. Here we show that cell injury and death of tumor or even normal cells provide a potent adjuvant effect for the stimulation of cytotoxic T lymphocyte responses. This adjuvant activity is constitutively present in the cytoplasm of cells and is increased in the cytoplasm of cells dying by apoptosis. The release of these components stimulates immune responses both locally and at a distance, and provides a simple mechanism to alert the immune system to potential danger in almost all pathological situations. PMID- 11106388 TI - Structural characterization of the human respiratory syncytial virus fusion protein core. AB - Human respiratory syncytial virus (HRSV) is a major cause of a number of severe respiratory diseases, including bronchiolitis and pneumonia, in infants and young children. The HRSV F protein, a glycoprotein essential for viral entry, is a primary target for vaccine and drug development. Two heptad-repeat regions within the HRSV F sequence were predicted by the computer program learncoil-vmf. These regions are thought to form trimer-of-hairpins-like structures, similar to those found in the fusion proteins of several enveloped viruses. The hairpin structure likely brings the viral and cellular membranes into close apposition, thereby facilitating membrane fusion and subsequent viral entry. Here, we show that peptides, denoted HR-N and HR-C, corresponding to the heptad-repeat regions from the N-terminal and C-terminal segments of the HRSV F protein, respectively, form a stable alpha-helical trimer of heterodimers. The HRSV N/C complex was crystallized and its x-ray structure was determined at 2.3-A resolution. As anticipated, the complex is a six-helix bundle in which the HR-N peptides form a three-stranded, central coiled coil, and the HR-C peptides pack in an antiparallel manner into hydrophobic grooves on the coiled-coil surface. There is remarkable structural similarity between the HRSV N/C complex and the fusion protein core of other viruses, including HIV-1 gp41. In addition, earlier work has shown that HRSV HR-C peptides, like the HIV-1 gp41 C peptides, inhibit viral infection. Thus, drug discovery and vaccine development strategies aimed at inhibiting viral entry by blocking hairpin formation may be applied to the inhibition of HRSV. PMID- 11106389 TI - An herbivore elicitor activates the gene for indole emission in maize. AB - Maize and a variety of other plant species release volatile compounds in response to herbivore attack that serve as chemical cues to signal natural enemies of the feeding herbivore. N-(17-hydroxylinolenoyl)-l-glutamine is an elicitor component that has been isolated and chemically characterized from the regurgitant of the herbivore-pest beet armyworm. This fatty acid derivative, referred to as volicitin, triggers the synthesis and release of volatile components, including terpenoids and indole in maize. Here we report on a previously unidentified enzyme, indole-3-glycerol phosphate lyase (IGL), that catalyzes the formation of free indole and is selectively activated by volicitin. IGL's enzymatic properties are similar to BX1, a maize enzyme that serves as the entry point to the secondary defense metabolites DIBOA and DIMBOA. Gene-sequence analysis indicates that Igl and Bx1 are evolutionarily related to the tryptophan synthase alpha subunit. PMID- 11106390 TI - Time-resolved x-ray diffraction reveals multiple conformations in the M-N transition of the bacteriorhodopsin photocycle. AB - We measured the M-N transition of wild-type bacteriorhodopsin (pH 9, 10 degrees C) by time-resolved x-ray diffraction study at SPring8 BL45XU-A. We confirmed the accumulation of M and N intermediates by absorbance measurements, and we found that the time resolution of x-ray diffraction experiments (244 ms) was sufficient to resolve the M-N transition. From the x-ray diffraction data, three components were decomposed by singular value decomposition analysis. The existence of three components in the M-->N-->BR reaction revealed that BR changes its structure during the M-N transition. Moreover, the difference Fourier maps of reconstituted fast and slow decay components clearly showed that the electron density distributions of the F helix changes in the M-N transition. The observed structural change at the F helix will increase access of the Schiff base and D96 to the cytoplasmic surface and facilitate the proton transfer steps that begin with the decay of the M state. PMID- 11106391 TI - Language-related cortex in deaf individuals: functional specialization for language or perceptual plasticity? PMID- 11106392 TI - Structural basis of the abscess-modulating polysaccharide A2 from Bacteroides fragilis. AB - Zwitterionic capsular polysaccharides from pathogenic bacteria have peculiar immunological properties. They are capable of eliciting T-cell proliferation and modulating the course of abscess formation. To understand the molecular basis of this characteristic immune response, we are conducting detailed structure function studies on these polysaccharides. We have identified, purified, and characterized an abscess-modulating polysaccharide, PS A2, from the clinical strain Bacteroides fragilis 638R. Here, we report the elucidation of both the chemical and three-dimensional structures of PS A2 by NMR spectroscopy, chemical methods, gas chromatography-mass spectrometry, and restrained molecular dynamics calculations. PS A2 consists of a pentasaccharide repeating unit containing mannoheptose, N-acetylmannosamine, 3-acetamido-3,6-dideoxyglucose, 2-amino-4 acetamido-2,4,6-trideoxygalactose, fucose, and 3-hydroxybutanoic acid. PS A2 is zwitterionic and carries one cationic free amine and one anionic carboxylate in each repeating unit. It forms an extended right-handed helix with two repeating units per turn and a pitch of 20 A. Positive and negative charges are exposed on the outer surface of the polymer in a regularly spaced pattern, which renders them easily accessible to other molecules. The helix is characterized by repeated large grooves whose lateral boundaries are occupied by the charges. The three dimensional structure of PS A2 explicitly suggests mechanisms of interaction between zwitterionic polysaccharides and proteins. PMID- 11106393 TI - Improved barley broiler feed with transgenic malt containing heat-stable (1,3 1,4)-beta-glucanase. AB - The low nutritional value of barley for poultry is because of the absence of an intestinal enzyme for efficient depolymerization of (1, 3-1,4)-beta-glucan, the major polysaccharide of the endosperm cell walls. This leads to high viscosity in the intestine, limited nutrient uptake, decreased growth rate, and unhygienic sticky droppings adhering to chickens and floors of the production cages. Consequently, the 7.5 billion broiler chickens produced annually in the United States are primarily raised on corn-soybean diets. Here we show that addition to normal barley of 6.2% transgenic malt containing a thermotolerant (1,3-1,4)-beta glucanase (4.28 microg.g(-1) soluble protein) provides a weight gain equivalent to corn diets. The number of birds with adhering sticky droppings is drastically reduced. Intestines and excrements of chickens fed the barley control diet contained large amounts of soluble (1,3-1,4)-beta-glucan, which was reduced by 75 and 50%, respectively, by adding transgenic malt to the diet. The amount of active recombinant enzyme in the small intestine corresponded to that present in the feed, whereas an 11-fold concentration of the enzyme was observed in the ceca, and a 7.5-fold concentration occurred in the excrement. Glycosylation of the beta-glucanase isolated from the ceca testified to its origin from the transgenic barley. Analysis of the data from this trial demonstrates the possibility of introducing individual recombinant enzymes into various parts of the gastrointestinal tract of chickens with transgenic malt and thereby the possibility of evaluating their effect on the metabolism of a given ingredient targeted by the enzyme. PMID- 11106394 TI - The mechanism of substrate (aglycone) specificity in beta -glucosidases is revealed by crystal structures of mutant maize beta -glucosidase-DIMBOA, DIMBOAGlc, and -dhurrin complexes. AB - The mechanism and the site of substrate (i.e., aglycone) recognition and specificity were investigated in maize beta-glucosidase (Glu1) by x-ray crystallography by using crystals of a catalytically inactive mutant (Glu1E191D) in complex with the natural substrate 2-O-beta-d-glucopyranosyl-4-hydroxy-7 methoxy-1,4-benzoxazin-3-one (DIMBOAGlc), the free aglycone DIMBOA, and competitive inhibitor para-hydroxy-S-mandelonitrile beta-glucoside (dhurrin). The structures of these complexes and of the free enzyme were solved at 2.1-, 2.1-, 2.0-, and 2.2-A resolution, respectively. The structural data from the complexes allowed us to visualize an intact substrate, free aglycone, or a competitive inhibitor in the slot-like active site of a beta-glucosidase. These data show that the aglycone moiety of the substrate is sandwiched between W378 on one side and F198, F205, and F466 on the other. Thus, specific conformations of these four hydrophobic amino acids and the shape of the aglycone-binding site they form determine aglycone recognition and substrate specificity in Glu1. In addition to these four residues, A467 interacts with the 7-methoxy group of DIMBOA. All residues but W378 are variable among beta-glucosidases that differ in substrate specificity, supporting the conclusion that these sites are the basis of aglycone recognition and binding (i.e., substrate specificity) in beta-glucosidases. The data also provide a plausible explanation for the competitive binding of dhurrin to maize beta-glucosidases with high affinity without being hydrolyzed. PMID- 11106395 TI - Molecular basis for discriminating between normal and damaged bases by the human alkyladenine glycosylase, AAG. AB - The human 3-methyladenine DNA glycosylase [alkyladenine DNA glycosylase (AAG)] catalyzes the first step of base excision repair by cleaving damaged bases from DNA. Unlike other DNA glycosylases that are specific for a particular type of damaged base, AAG excises a chemically diverse selection of substrate bases damaged by alkylation or deamination. The 2.1-A crystal structure of AAG complexed to DNA containing 1,N(6)-ethenoadenine suggests how modified bases can be distinguished from normal DNA bases in the enzyme active site. Mutational analyses of residues contacting the alkylated base in the crystal structures suggest that the shape of the damaged base, its hydrogen-bonding characteristics, and its aromaticity all contribute to the selective recognition of damage by AAG. PMID- 11106396 TI - Targeted modification and transportation of cellular proteins. AB - Peptide aptamers are proteins selected from combinatorial libraries that display conformationally constrained variable regions. Peptide aptamers can disrupt specific protein interactions and thus represent a useful method for manipulating protein function in vivo. Here, we describe aptamer derivatives that extend the range of functional manipulations. We isolated an aptamer with increased affinity for its Cdk2 target by mutagenizing an existing aptamer and identifying tighter binding mutants with calibrated two-hybrid reporter genes. We used this and other anti-Cdk2 aptamers as recognition domains in chimeric proteins that contained other functional moieties. Aptamers fused to the catalytic domain of a ubiquitin ligase specifically decorated LexA-Cdk2 with ubiquitin moieties in vivo. Aptamers against Cdk2 and another protein, Ste5, that carried a nuclear localization sequence transported their targets into the nucleus. These experiments indicate that fusion proteins containing aptameric recognition moieties will be useful for specific modification of protein function in vivo. PMID- 11106397 TI - A family of peptidoglycan recognition proteins in the fruit fly Drosophila melanogaster. AB - Peptidoglycans from bacterial cell walls trigger immune responses in insects and mammals. A peptidoglycan recognition protein, PGRP, has been cloned from moths as well as vertebrates and has been shown to participate in peptidoglycan-mediated activation of prophenoloxidase in the silk moth. Here we report that Drosophila expresses 12 PGRP genes, distributed in 8 chromosomal loci on the 3 major chromosomes. By analyzing cDNA clones and genomic databases, we grouped them into two classes: PGRP-SA, SB1, SB2, SC1A, SC1B, SC2, and SD, with short transcripts and short 5'-untranslated regions; and PGRP-LA, LB, LC, LD, and LE, with long transcripts and long 5'-untranslated regions. The predicted structures indicate that the first group encodes extracellular proteins and the second group, intracellular and membrane-spanning proteins. Most PGRP genes are expressed in all postembryonic stages. Peptidoglycan injections strongly induce five of the genes. Transcripts from the different PGRP genes were found in immune competent organs such as fat body, gut, and hemocytes. We demonstrate that at least PGRP-SA and SC1B can bind peptidoglycan, and a function in immunity is likely for this family. PMID- 11106398 TI - Correlation between plasmid content and infectivity in Borrelia burgdorferi. AB - Infectivity-associated plasmids were identified in Borrelia burgdorferi B31 by using PCR to detect each of the plasmids in a panel of 19 clonal isolates. The clones exhibited high-, low-, and intermediate-infectivity phenotypes based on their frequency of isolation from needle-inoculated C3H/HeN mice. Presence or absence of 21 of the 22 plasmids was determined in each of the clones by using PCR primers specific for regions unique to each plasmid, as identified in the recently available genome sequence. Southern blot hybridization results were used to confirm the PCR results in some cases. Plasmid lp25 exhibited a direct correlation with infectivity in that it was consistently present in all clones of high or intermediate infectivity and was absent in all low-infectivity clones. lp28-1, containing the vmp-like sequence locus, also correlated with infectivity; all clones that lacked lp28-1 but contained lp25 had an intermediate infectivity phenotype, in which infection was primarily restricted to the joints. Plasmids cp9, cp32-3, lp21, lp28-2, lp28-4, and lp56 apparently are not required for infection in this model, because clones lacking these plasmids exhibited a high infectivity phenotype. Plasmids cp26, cp32-1, cp32-2 and/or cp32-7, cp32-4, cp32 6, cp32-8, cp32-9, lp17, lp28-3, lp36, lp38, and lp54 were consistently present in all clones examined. On the basis of these results, lp25 and lp28-1 appear to encode virulence factors important in the pathogenesis of B. burgdorferi B31. PMID- 11106399 TI - Transplanted fetal striatum in Huntington's disease: phenotypic development and lack of pathology. AB - Neural and stem cell transplantation is emerging as a potential treatment for neurodegenerative diseases. Transplantation of specific committed neuroblasts (fetal neurons) to the adult brain provides such scientific exploration of these new potential therapies. Huntington's disease (HD) is a fatal, incurable autosomal dominant (CAG repeat expansion of huntingtin protein) neurodegenerative disorder with primary neuronal pathology within the caudate-putamen (striatum). In a clinical trial of human fetal striatal tissue transplantation, one patient died 18 months after transplantation from cardiovascular disease, and postmortem histological analysis demonstrated surviving transplanted cells with typical morphology of the developing striatum. Selective markers of both striatal projection and interneurons such as dopamine and c-AMP-related phosphoprotein, calretinin, acetylcholinesterase, choline acetyltransferase, tyrosine hydroxylase, calbindin, enkephalin, and substance P showed positive transplant regions clearly innervated by host tyrosine hydroxylase fibers. There was no histological evidence of immune rejection including microglia and macrophages. Notably, neuronal protein aggregates of mutated huntingtin, which is typical HD neuropathology, were not found within the transplanted fetal tissue. Thus, although there is a genetically predetermined process causing neuronal death within the HD striatum, implanted fetal neural cells lacking the mutant HD gene may be able to replace damaged host neurons and reconstitute damaged neuronal connections. This study demonstrates that grafts derived from human fetal striatal tissue can survive, develop, and are unaffected by the disease process, at least for 18 months, after transplantation into a patient with HD. PMID- 11106400 TI - Speech-like cerebral activity in profoundly deaf people processing signed languages: implications for the neural basis of human language. AB - For more than a century we have understood that our brain's left hemisphere is the primary site for processing language, yet why this is so has remained more elusive. Using positron emission tomography, we report cerebral blood flow activity in profoundly deaf signers processing specific aspects of sign language in key brain sites widely assumed to be unimodal speech or sound processing areas: the left inferior frontal cortex when signers produced meaningful signs, and the planum temporale bilaterally when they viewed signs or meaningless parts of signs (sign-phonetic and syllabic units). Contrary to prevailing wisdom, the planum temporale may not be exclusively dedicated to processing speech sounds, but may be specialized for processing more abstract properties essential to language that can engage multiple modalities. We hypothesize that the neural tissue involved in language processing may not be prespecified exclusively by sensory modality (such as sound) but may entail polymodal neural tissue that has evolved unique sensitivity to aspects of the patterning of natural language. Such neural specialization for aspects of language patterning appears to be neurally unmodifiable in so far as languages with radically different sensory modalities such as speech and sign are processed at similar brain sites, while, at the same time, the neural pathways for expressing and perceiving natural language appear to be neurally highly modifiable. PMID- 11106401 TI - Novel features of the XRN-family in Arabidopsis: evidence that AtXRN4, one of several orthologs of nuclear Xrn2p/Rat1p, functions in the cytoplasm. AB - The 5'-3' exoribonucleases Xrn1p and Xrn2p/Rat1p function in the degradation and processing of several classes of RNA in Saccharomyces cerevisiae. Xrn1p is the main enzyme catalyzing cytoplasmic mRNA degradation in multiple decay pathways, whereas Xrn2p/Rat1p functions in the processing of rRNAs and small nucleolar RNAs (snoRNAs) in the nucleus. Much less is known about the XRN-like proteins of multicellular eukaryotes; however, differences in their activities could explain differences in mRNA degradation between multicellular and unicellular eukaryotes. One such difference is the lack in plants and animals of mRNA decay intermediates like those generated in yeast when Xrn1p is blocked by poly(G) tracts that are inserted within mRNAs. We investigated the XRN-family in Arabidopsis thaliana and found it to have several novel features. First, the Arabidopsis genome contains three XRN-like genes (AtXRNs) that are structurally similar to Xrn2p/Rat1p, a characteristic unique to plants. Furthermore, our experimental results and sequence database searches indicate that Xrn1p orthologs may be absent from higher plants. Second, the lack of poly(G) mRNA decay intermediates in plants cannot be explained by the activity of the AtXRNs, because they are blocked by poly(G) tracts. Finally, complementation of yeast mutants and localization studies indicate that two of the AtXRNs likely function in the nucleus, whereas the third acts in the cytoplasm. Thus, the XRN-family in plants is more complex than in other eukaryotes, and, if an XRN-like enzyme plays a role in mRNA decay in plants, the likely participant is a cytoplasmic Xrn2p/Rat1p ortholog, rather than an Xrn1p ortholog. PMID- 11106402 TI - Transforming growth factor-beta PMID- 11106403 TI - Signaling of transforming growth factor-beta family members through Smad proteins. AB - Smads are pivotal intracellular nuclear effectors of transforming growth factor beta (TGF-beta) family members. Ligand-induced activation of TGF-beta family receptors with intrinsic serine/threonine kinase activity trigger phosphorylation of receptor-regulated Smads (R-Smads), whereas Smad2 and Smad3 are phosphorylated by TGF-beta, and activin type I receptors, Smad1, Smad5 and Smad8, act downstream of BMP type I receptors. Activated R-Smads form heteromeric complexes with common partner Smads (Co-Smads), e.g. Smad4, which translocate efficiently to the nucleus, where they regulate, in co-operation with other transcription factors, coactivators and corepressors, the transcription of target genes. Inhibitory Smads act in most cases in an opposite manner from R- and Co-Smads. Like other components in the TGF-beta family signaling cascade, Smad activity is intricately regulated. The multifunctional and context dependency of TGF-beta family responses are reflected in the function of Smads as signal integrators. Certain Smads are somatically mutated at high frequency in particular types of human cancers. Gene ablation of Smads in the mouse has revealed their critical roles during embryonic development. Here we review the latest advances in our understanding of the Smad mechanism of action and their in vivo functions. PMID- 11106404 TI - GDNF - a stranger in the TGF-beta superfamily? AB - Glial cell line-derived neurotrophic factor (GDNF) family, consisting of GDNF, neurturin, artemin and persephin are distant members of the transforming growth factor-beta (TGF-beta) superfamily. Unlike other members of the TGF-beta superfamily, which signal through the receptor serine-threonine kinases, GDNF family ligands activate intracellular signalling cascades via the receptor tyrosine kinase Ret. GDNF family ligands first bind to the glycosylphosphatidylinositol (GPI)-anchored GDNF family receptor alpha (GFRalpha) and then the GDNF family ligand-GFRalpha complex binds to and stimulates autophosphorylation of Ret. Alternatively, a preassociated complex between GFRalpha and Ret could form the binding site for the GDNF family ligand. GFRalpha1, GFRalpha2, GFRalpha3 and GFRalpha4 are the physiological coreceptors for GDNF, neurturin, artemin and persephin, respectively. Although all GDNF family ligands signal via activated Ret, GDNF can signal also via GFRalpha1 in the absence of Ret. GPI-anchored GFRalpha receptors are localized in plasma membrane to lipid rafts. GDNF binding to GFRalpha1 also recruits Ret to the lipid rafts and triggers association with Src, which is required for effective downstream signalling, leading to differentiation and neuronal survival. GDNF family ligands are potent survival factors for midbrain dopamine neurons, motoneurons, noradrenergic neurons, as well as for sympathetic, parasympathetic and sensory neurons. However, for most neuronal populations, except for motoneurons, TGF-beta is required as a cofactor for GDNF family ligand signalling. Because GDNF and neurturin can rescue dopamine neurons in the animal models of Parkinson disease, as well as motoneurons in vivo, hopes have been raised that GDNF family ligands may be new drugs for the treatment of neurodegenerative diseases. GDNF also has distinct functions outside the nervous system, promoting ureteric branching in kidney development and regulating spermatogenesis. PMID- 11106405 TI - Functions of transforming growth factor-beta isoforms in the nervous system. Cues based on localization and experimental in vitro and in vivo evidence. AB - This review briefly describes the cellular distribution and documented roles of the transforming growth factor (TGF)-beta isoforms TGF-beta2 and -beta3 in the central and peripheral nervous system. TGF-beta2 and -beta3 are coexpressed in developing radial glial and mature astroglial and Schwann cells, as well as in subpopulations of differentiated neurons, most prominently in cortical, hippocampal, and brainstem/spinal cord motor neurons. In vitro studies have suggested a number of potential, physiologically relevant functions for TGF-betas including regulation of astroglial cell proliferation, expression of adhesion molecules, survival promoting roles for neurons in combination with established neurotrophic factors, and differentiative actions on neurons. PMID- 11106406 TI - Evidence for an evolutionary conserved role of bone morphogenetic protein growth factors and phox2 transcription factors during noradrenergic differentiation of sympathetic neurons. Induction of a putative synexpression group of neurotransmitter-synthesizing enzymes. AB - The noradrenergic transmitter phenotype in postganglionic sympathetic neurons is induced early during embryonic development in avian and mammalian primary sympathetic ganglia. The simultaneous expression of tyrosine hydroxylase and dopamine beta-hydroxylase, enzymes of the noradrenaline biosynthesis pathway, indicates that different genes contributing to the noradrenergic transmitter phenotype are regulated as a synexpression group. This conclusion is supported by the demonstration of bone morphogenetic protein (BMP) growth factors and Phox2 transcription factors being necessary for the expression of both tyrosine hydroxylase and dopamine beta-hydroxylase in differentiating sympathetic neurons. The close similarity in the expression patterns of the relevant genes as well as in the function of BMPs and Phox2s between avian and mammalian embryos strongly suggests that noradrenergic induction occurs along a conserved signalling pathway in these vertebrate classes. PMID- 11106407 TI - Targeted mutations of transforming growth factor-beta genes reveal important roles in mouse development and adult homeostasis. AB - Transforming growth factors-beta (TGF-beta) are multifunctional molecules with profound biological effects in many developmental processes including regulation of cell proliferation, differentiation, cell adhesion, skeletal development, haematopoiesis, inflammatory responses, and wound healing. To learn about the role of TGF-beta in vivo, phenotypes of targeted mutations of molecules within the TGF-beta signalling pathway, TGF-beta1, -beta2, -beta3, TGF-beta receptor (TbetaR-II) and the signalling molecules SMAD2, SMAD3 and SMAD4, are discussed in this review. The three individual TGF-beta mutants show distinct and only partially overlapping phenotypes. In mice, targeted disruption of the TGF-beta1 gene results in diffuse and lethal inflammation about 3 weeks after birth, suggesting a prominent role of TGF-beta in the regulation of immune cell proliferation and extravasation into tissues. However, just half of the TGF-beta1 (-/-) conceptuses actually reach partuition due to defective haematopoiesis and endothelial differentiation. Targeted disruption of both TGF-beta2 and TGF-beta3 genes results in perinatal lethality. TGF-beta2 null mice exhibit a broad range of developmental defects, including cardiac, lung, craniofacial, limb, eye, ear and urogenital defects, whereas TGF-beta3 gene ablation results exclusively in defective palatogenesis and delayed pulmonary development. The TbetaR-II null phenotype closely resembles that of TGF-beta1 (-/-) conceptuses, which die in utero by E10.5. Loss of SMAD2 or SMAD4 results in related phenotypes: the mutants fail to form an organized egg cylinder, lack mesoderm required for gastrulation and die prior to E8.5. Together, gene ablation within the TGF-beta signalling pathway supports the notion of a prominent role of TGF-beta during development. PMID- 11106408 TI - Identification of low-density Triton X-100-insoluble plasma membrane microdomains in higher plants. AB - Low density Triton X-100-insoluble plasma membrane microdomains can be isolated from different mammalian cell types and are proposed to be involved in membrane trafficking, cell morphogenesis and signal transduction. Heterotrimeric G proteins and their receptors are often associated with such domains, suggesting that these structures are involved in G-protein-coupled signaling. Here we report that detergent-insoluble plasma membrane microdomains also exist in higher plants and contain about 15% of membrane-bound heterotrimeric G-protein beta-subunit (Gbeta). Plasma membrane microdomains were isolated from tobacco leaves. They have low buoyant density relative to the surrounding plasma membrane, and are insoluble in Triton X-100 at 4 degrees C. Detergent-insoluble vesicles were examined by freeze-fracture electron microscopy. They have sizes in the range 100 400 nm, and often contain aggregated protein complexes. The majority of plasma membrane proteins cannot be detected in the Triton X-100-insoluble fraction, while few polypeptides are highly enriched. We identified six proteins with molecular masses of 22, 28, 35, 60, 67 and 94 kDa in detergent-insoluble fractions that are glycosylphosphatidylinositol (GPI)-anchored. PMID- 11106409 TI - Differential mechanism-based labeling and unequivocal activity assignment of the two active sites of intestinal lactase/phlorizin hydrolase. AB - Milk lactose is hydrolysed to galactose and glucose in the small intestine of mammals by the lactase/phlorizin hydrolase complex (LPH; EC 3.2.1.108/62). The two enzymatic activities, lactase and phlorizin hydrolase, are located in the same polypeptide chain. According to sequence homology, mature LPH contains two different regions (III and IV), each of them homologous to family 1 glycosidases and each with a putative active site. There has been some discrepancy with regard to the assignment of enzymatic activity to the two active sites. Here we show differential reactivity of the two active sites with mechanism-based glycosidase inhibitors. When LPH is treated with 2',4'-dinitrophenyl 2-deoxy-2-fluoro-beta-D glucopyranoside (1) and 2', 4'-dinitrophenyl-2-deoxy-2-fluoro-beta-D galactopyranoside (2), known mechanism-based inhibitors of glycosidases, it is observed that compound 1 preferentially inactivates the phlorizin hydrolase activity whereas compound 2 is selective for the lactase active site. On the other hand, glycals (D-glucal and D-galactal) competitively inhibit lactase activity but not phlorizin hydrolase activity. This allows labeling of the phlorizin site with compound 1 by protection with a glycal. By differential labeling of each active site using 1 and 2 followed by proteolysis and MS analysis of the labeled fragments, we confirm that the phlorizin hydrolysis occurs mainly at the active site located at region III of LPH and that the active site located at region IV is responsible for the lactase activity. This assignment is coincident with that proposed from the results of recent active site mutagenesis studies [Zecca, L., Mesonero, J.E., Stutz, A., Poiree, J.C., Giudicelli, J., Cursio, R., Gloor, S.M. & Semenza, G. (1998) FEBS Lett. 435, 225 228] and opposite to that based on data from early affinity labeling with conduritol B epoxide [Wacker, W., Keller, P., Falchetto, R., Legler, G. & Semenza, G. (1992) J. Biol. Chem. 267, 18744-18752]. PMID- 11106410 TI - Hev b 9, an enolase and a new cross-reactive allergen from hevea latex and molds. Purification, characterization, cloning and expression. AB - Natural rubber latex allergy is an IgE-mediated disease that is caused by proteins that elute from commercial latex products. A complementary DNA (cDNA) coding for Hev b 9, an enolase (2-phospho-D-glycerate hydrolyase) and allergen from latex of the rubber tree Hevea brasiliensis, was amplified by PCR. The PCR primers were designed according to conserved regions of enolases from plants. The obtained cDNA amplification product consisted of 1651 bp and encoded a protein of 445 amino-acid residues with a calculated molecular mass of 47.6 kDa. Sequence comparisons revealed high similarities of the Hevea latex enolase to mold enolases that have been identified as important allergens. In addition, the crucial amino-acid residues that participate in the formation of the catalytic site and the Mg2+ binding site of enolases were also conserved. Hevea latex enolase was produced as a recombinant protein in Escherichia coli with an N terminal hexahistidyl tag, and purified by affinity chromatography. The yield amounted to 110 mg of purified Hev b 9 per litre of bacterial culture. The recombinant allergen bound IgE from latex, as well as mold-allergic patients, in immunoblot and ELISA experiments. The natural enolase was isolated from Hevea latex by (NH4)2SO4 precipitation and ion exchange chromatography. The natural and the recombinant (r)Hev b 9 showed equivalent enzymatic activity. Patients' IgE antibodies preincubated with rHev b 9 lost their ability to bind to natural (n) Hev b 9, indicating the identity of the B-cell epitopes on both molecules. Cross reactivity with two enolases from Cladosporium herbarum and Alternaria alternata was determined by inhibition of IgE-binding to these enolases by rHev b 9. Therefore, enolases may represent another class of highly conserved enzymes with allergenic potentials. PMID- 11106411 TI - Overproduction of spinach betaine aldehyde dehydrogenase in Escherichia coli. Structural and functional properties of wild-type, mutants and E. coli enzymes. AB - Betaine aldehyde dehydrogenase (BADH) catalyzes the last step in the synthesis of the osmoprotectant glycine betaine from choline. Although betaine aldehyde has been thought to be a specific substrate for BADH, recent studies have shown that human and sugar beet BADHs also catalyze the oxidation of omega-aminoaldehydes. To characterize the kinetic and stability properties of spinach BADH, five kinds of expression vectors encoding full length, mature, E103Q, E103K, and chimera BADHs were constructed. These enzymes together with Escherichia coli BADH were expressed in E. coli and purified. The affinities for betaine aldehyde were similar in the spinach and E. coli BADHs, whereas those for omega-aminoaldehydes were higher in spinach BADH than in E. coli BADH. A chimera BADH in which part of the Rossmann type fold in the spinach BADH was replaced with that of E. coli BADH, showed properties which resembled spinach BADH more than E. coli BADH. The spinach E103K mutant was almost inactive, whereas the E103Q mutant showed a similar activity for the oxidation of betaine aldehyde to that of wild type BADH, but a lower affinity for omega-aminoaldehydes. All spinach BADHs were dimers whereas E. coli BADH was a tetramer. E. coli BADH was more stable at high temperature than spinach BADHs. The E103Q mutant was most labile to high temperature. These properties are discussed in relation to the structure of spinach BADH. PMID- 11106412 TI - Peptide bond formation mediated by substrate mimetics. Structure guidedoptimization of trypsin for synthesis. AB - Substrate mimetics are excellent tools for protease-mediated peptide synthesis that enable the coupling of peptides independently of the primary specificity of the enzyme without undesired cleavages of the newly formed peptide bonds. However, the synthetic utility of this beneficial approach is limited to reactions with nonspecific amino-acid-containing peptides while the coupling of specific ones leads to unwanted cleavages due to the native proteolytic activity of the biocatalyst. This paper reports on the use of site-directed mutagenesis to design trypsin variants with decreased cleavage activity. Starting from the variant D189S, which is known for its low proteolytic potential, Ser189 and Ser190 were exchanged for Ala to further repress the inherent amidase activity of trypsin D189S. The effect of mutations was analysed by model synthesis reactions using specific amino-acid-containing peptides and substrate mimetics as the reactants. Finally, computer-assisted protein-ligand docking studies were performed to get closer insight into the molecular basis of the experimental results. PMID- 11106413 TI - Cloning and characterization of the rainbow trout (Oncorhynchus mykiss) type II interleukin-1 receptor cDNA. AB - A homologue of mammalian type II interleukin-1 receptor (IL-1RII) was isolated from a rainbow trout cDNA library by differential hybridization using a suppression subtractive hybridization generated probe enriched for sequences upregulated after immune stimulation. The trout cDNA has an ORF encoding 441 amino acids, and represents the first piscine IL-1 receptor described. The predicted amino-acid sequence has 29 and 26% identity with human and mouse IL 1RII, respectively. The trout IL-1 receptor has a domain organization similar to that of mammalian type II receptor, with a short cytoplasmic tail of 24 amino acids. These results suggest that type II receptor is also present in lower vertebrates, and therefore the duplication of an ancestral gene that generated type I and type II IL-1 receptors occurred prior to the time mammals emerged. PMID- 11106414 TI - Role of arginine 177 in the MnII binding site of manganese peroxidase. Studies with R177D, R177E, R177N, and R177Q mutants. AB - Previously, we reported that Arg177 is involved in MnII binding at the MnII binding site of manganese peroxidase isozyme 1 (MnP1) of Phanerochaete chrysosporium by examining two mutants: R177A and R177K. We now report on additional mutants: R177D, R177E, R177N, and R177Q. These new mutant enzymes were produced by homologous expression in P. chrysosporium and were purified to homogeneity. The molecular mass and the UV/visible spectra of the ferric and oxidized intermediates of the mutant enzymes were similar to those of the wild type enzyme, suggesting proper folding, heme insertion, and preservation of the heme environment. However, steady-state and transient-state kinetic analyses demonstrate significantly altered characteristics of MnII oxidation by these new mutant enzymes. Increased dissociation constants (Kd) and apparent Km values for MnII suggest that these mutations at Arg177 decrease binding of MnII to the enzyme. These lowered binding efficiencies, as observed with the R177A and R177K mutants, suggest that the salt-bridge between Arg177 and the MnII binding ligand Glu35 is disrupted in these new mutants. Decreased kcat values for MnII oxidation, decreased second-order rate constants for compound I reduction (k2app), and decreased first-order rate constants for compound II reduction (k3) indicate that these new mutations also decrease the electron-transfer rate. This decrease in rate constants for compounds I and II reduction was not observed in our previous study on the R177A and R177K mutations. The lower rate constants suggest that, even with high MnII concentrations, the MnII binding geometries may be altered in the MnII binding site of these new mutants. These new results, combined with the results from our previous study, clearly indicate a role for Arg177 in promoting efficient MnII binding and oxidation by MnP. PMID- 11106415 TI - Molecular heterogeneity of the hemocyanin isolated from the king crab Paralithodes camtschaticae. AB - Native Paralithodes camtschaticae hemocyanin is found as a mixture of dodecamers (24S; 80%) and hexamers (16S; 20%). Removal of Ca2+ ions by dialysis against EDTA containing buffer solution at neutral pH induces complete dissociation of the 24S form into the 16S form. Under these conditions, a further increase in pH to 9.2 produces complete dissociation of the hexamers into monomers (5S). In both cases, the dissociation process is reversible. The dodecamer (24S) is composed of two different hexamers which can be discriminated only by ion-exchange chromatography in the presence of Ca2+ ions. At alkaline pH and in the presence of EDTA, two major monomeric fractions can be separated by ion-exchange chromatography: ParcI (60%) and ParcII (40%). The reassociation properties of the two fractions were studied separately to define their ability to form hexamers and dodecamers. The oxygen-binding properties of the different aggregation states were investigated. Native hemocyanin binds O2 co-operatively (nH = 3) and with low affinity (p50 approximately 103 Torr). The two monomeric fractions, ParcI and ParcII, are not co-operative and the affinity is twice that of the native protein (p50 approximately 65 and 52 Torr). Oxygen-binding measurements of native hemocyanin carried out at different pH values indicate a strong positive Bohr effect within the pH range 6.5-8.0 and an increase in oxygen affinity at pH below 6.5. PMID- 11106416 TI - Temporal secretion of a multicellulolytic system in Myxobacter sp. AL-1. Molecular cloning and heterologous expression of cel9 encoding a modular endocellulase clustered in an operon with cel48, an exocellobiohydrolase gene. AB - The Gram-negative soil micro-organism Myxobacter sp. AL-1 possesses at least five extracellular cellulases, the production of which is regulated by the growth cycle. We cloned the complete gene for one of these cellulases, termed cel9, which encoded a 67-kDa modular family 9 endoglycohydrolase, which was produced during the stationary phase of growth and was strongly enhanced by avicel. The predicted product of cel9 matches the structural architecture of family 9 cellulases such as Thermonospora fusca endo/exocellulase E4. Cel9 protein was synthesized in Escherichia coli from a multicopy plasmid and in Bacillus subtilis from the isopropyl thiogalactoside-inducible Pspac promoter and was purified from the culture medium. Thermal stability, optimum pH and temperature dependence of Cel9 were similar when expressed from either source, and were indistinguishable from related cellulases produced by thermophilic bacteria. Downstream from cel9 was found a partial ORF, designated cel48, the deduced product of which was highly similar to bacterial exocellobiohydrolases and processive endoglucanases belonging to family 48 of the glycosyl hydrolases. The cel9 and cel48 genes appear to be arranged as part of an operon. PMID- 11106417 TI - Characterization of active-site mutants of Schizosaccharomyces pombe phosphoglycerate mutase. Elucidation of the roles of amino acids involved in substrate binding and catalysis. AB - The roles of a number of amino acids present at the active site of the monomeric phosphoglycerate mutase from the fission yeast Schizosaccharomyces pombe have been explored by site-directed mutagenesis. The amino acids examined could be divided broadly into those presumed from previous related structural studies to be important in the catalytic process (R14, S62 and E93) and those thought to be important in substrate binding (R94, R120 and R121). Most of these residues have not previously been studied by site-directed mutagenesis. All the mutants except R14 were expressed in an engineered null strain of Saccharomyces cerevisiae (S150 gpm:HIS) in good yield. The R14Q mutant was expressed in good yield in the transformed AH22 strain of S. cerevisiae. The S62A mutant was markedly unstable, preventing purification. The various mutants were purified to homogeneity and characterized in terms of kinetic parameters, CD and fluorescence spectra, stability towards denaturation by guanidinium chloride, and stability of phosphorylated enzyme intermediate. In addition, the binding of substrate (3 phosphoglycerate) to wild-type, E93D and R120,121Q enzymes was measured by isothermal titration calorimetry. The results provide evidence for the proposed roles of each of these amino acids in the catalytic cycle and in substrate binding, and will support the current investigation of the structure and dynamics of the enzyme using multidimensional NMR techniques. PMID- 11106418 TI - Refolding, structural transition and spermatozoa-binding of recombinant bonnet monkey (Macaca radiata) zona pellucida glycoprotein-C expressed in Escherichia coli. AB - An internal cDNA fragment (978 bp) corresponding to bonnet monkey (Macaca radiata) zona pellucida glycoprotein-C (bmZPC), excluding the N-terminal signal sequence and the C-terminal transmembrane-like domain, was cloned in pQE-30 vector and the protein expressed as inclusion bodies in Escherichia coli. Recombinant bmZPC (r-bmZPC) was solubilized from purified inclusion bodies in the absence of a high concentration of chaotropic agents and was subsequently refolded. Use of a low concentration of urea (2 M) during solubilization of r bmZPC helped to minimize the extent of protein aggregation during refolding of the recombinant protein, and retain the existing native-like secondary structure that was essential for proper folding. Purified r-bmZPC appeared as a dominant band of 43 kDa on SDS/PAGE and Western blot. Although it lacked carbohydrate moieties, the purified and refolded r-bmZPC bound to the head region of bonnet monkey spermatozoa, confirming the existence of a native-like conformation. CD revealed a maximum at 200 nm and a single broad minimum extending from 209 to 216 nm, indicating the presence of both alpha-helical and beta-sheet conformations in the refolded r-bmZPC. Two different phases of transition were observed by urea gradient electrophoresis, suggesting the existence of multiple intermediate stages during the unfolding of r-bmZPC. The availability of refolded r-bmZPC will help in elucidating its role during the complex cascade of events during fertilization. PMID- 11106419 TI - The iron-sulfur clusters in 2-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans. Biochemical and spectroscopic investigations. AB - The reversible dehydration of (R)-2-hydroxyglutaryl-CoA to (E)-glutaconyl-CoA is catalysed by the combined action of two oxygen-sensitive enzymes from Acidaminococcus fermentans, the homodimeric component A (2 x 27 kDa) and the heterodimeric component D (45 and 50 kDa). Component A was purified to homogeneity (specific activity 25-30 s-1) using streptavidin-tag affinity chromatography. In the presence of 5 mM MgCl2 and 1 mM ADP or ATP, component A could be stabilized and stored for 4-5 days at 4 degrees C without loss of activity. The purification of component D from A. fermentans was also improved as indicated by the 1.5-fold higher specific activity (15 s-1). The content of 1.0 riboflavin 5'-phosphate (FMN) per heterodimer could be confirmed, whereas in contrast to an earlier report only trace amounts of riboflavin (< 0.1) could be detected. Each active component contains an oxygen sensitive diamagnetic [4Fe 4S]2+ cluster as revealed by UV-visible, EPR and Mossbauer spectroscopy. Reduction of the [4Fe-4S]2+ cluster in component A with dithionite yields a paramagnetic [4Fe-4S]1+ cluster with the unusual electron spin ground state S = 3/2 as indicated by strong absorption type EPR signals at high g values, g = 4-6. Spin-Hamiltonian simulations of the EPR spectra and of magnetic Mossbauer spectra were performed to determine the zero field splitting (ZFS) parameters of the cluster and the 57Fe hyperfine interaction parameters. The electronic properties of the [4Fe-4S]2+, 1+ clusters of component A are similar to those of the nitrogenase iron protein in which a [4Fe-4S]2+ cluster bridges the two subunits of the homodimeric protein. Under air component A looses its activity within seconds due to irreversible degradation of its [4Fe-4S]2+ cluster to a [2Fe-2S]2+ cluster. The [4Fe-4S]2+ cluster of component D could not be reduced to a [4Fe 4S]1+ cluster, even with excess of Ti(III)citrate or dithionite. Exposure to oxic conditions slowly converts the diamagnetic [4Fe-4S]2+ cluster of component D to a paramagnetic [3Fe-4S]+ cluster concomitant with loss of activity (30% within 24 h at 4 degrees C). PMID- 11106420 TI - Oxidative degradation of bilirubin produces vasoactive compounds. AB - Subarachnoid haemorrhage is often followed by haemolysis and concomitant oxidative stress, and is frequently complicated by pathological vasoconstriction or cerebral vasospasm. It is known that upregulation of haem oxygenase (HO-1) is induced by oxidative stress and results in release of biliverdin and bilirubin (BR), which are scavengers of reactive oxygen species (ROS). Here we report biomimetic studies aimed at modelling pathological conditions leading to oxidative degradation of BR. Oxidative degradation products of BR, formed by reaction with hydrogen peroxide (an ROS model system), demonstrated biological activity by stimulating oxygen consumption and force development in vascular smooth muscle from porcine carotid artery. Analogous biological activity was observed with vasoactive cerebrospinal fluid from subarachnoid haemorrhage patients. Three degradation products of BR were isolated: two were assigned as isomeric monopyrrole (C9H11N2O2) derivatives, 4-methyl-5-oxo-3-vinyl-(1, 5 dihydropyrrol-2-ylidene)acetamide and 3-methyl-5-oxo-4-vinyl-(1, 5-dihydropyrrol 2-ylidene)acetamide and the third was 4-methyl-3-vinylmaleimide (MVM), a previously isolated photodegradation product of biliverdin. Possible mechanisms of oxidative degradation of BR are discussed. Tentative assignment of these structures in the cerebrospinal fluid (CSF) of cerebral vasospasm patients has been made. It is proposed that one or more of the degradation products of biliverdin or bilirubin are involved in complications such as vasospasm and or pathological vasoconstriction associated with haemorrhage. PMID- 11106421 TI - Negative regulation of the platelet Na+/H+ exchanger by trimeric G-proteins. AB - Human platelets contain a Na+/H+ exchanger (NHE) that regulates the cytosolic pH. The role of trimeric G-proteins in NHE control was investigated in plasma membrane vesicles by measuring exchange of intravesicular protons for extravesicular Na+. Exchange was saturable, independent of membrane potential and inhibited by ethylisopropyl amiloride (Ki 0.05 micromol.L-1), demonstrating the involvement of NHE-1. The G-protein activators AlF4- and GMP-P(NH)P reduced exchange by increasing the Km for Na+ from 11.3 +/- 2.1 mM to 21.6 +/- 1.4 mM (AlF4-) and 19.8 +/- 1.1 mM (GMP-P(NH)P), leaving Vmax and the Hill coefficient unchanged. This effect was abolished by inhibitors of Gi-proteins (N ethylmaleimide, holoenzyme- and A-protomer of pertussis toxin) and by an anti Galpha Ig and GDP(beta)S. Activation of Gi-proteins by mastoparan and its synthetic analogue Mas7 also strongly reduced NHE activity. These data show that in platelets NHE-1 is under negative control of the Gi-family of trimeric G proteins. PMID- 11106422 TI - Cloning and expression of a distinct subclass of plant thioredoxins. AB - mRNAs encoding a novel thioredoxin were isolated from pollen RNA of Lolium perenne (LpTrx), Hordeum bulbosum (HbTrx), Phalaris coerulescens (PTrx) and Secale cereale (ScTrx). The cDNAs contain a single ORF of 393 bp encoding a protein of 131 amino acids. The predicted proteins showed highest homology to plant thioredoxins of the h class yet form a distinct subgroup that is characterized by a high level of sequence conservation (95.4-97.7% identity). GenBank searches revealed additional members of this subclass in tomato, soybean, rice and pine. LpTrx and PTrx were expressed as recombinant proteins in Escherichia coli and tested for thioredoxin activity. Both proteins displayed typical thioredoxin activity in the nonspecific insulin reduction assay, however, were not reduced by E. coli NADPH-dependant thioredoxin reductase. PMID- 11106423 TI - Involvement of calcium-independent phospholipase A2 in uterine stromal cell phospholipid remodelling. AB - The role of Ca2+-independent phospholipase A2 (iPLA2) in arachidonic (AA) and docosahexaenoic (DHA) acid incorporation and phospholipid remodelling in rat uterine stromal cells (UIII cells) was studied. Incorporation of AA and DHA into UIII cell phospholipids was Ca2+-independent. Bromoenollactone (BEL), a potent inhibitor of iPLA2, reduced lysophosphatidylcholine level and AA incorporation into phospholipids by approximately 20%. DHA incorporation was not affected by BEL, indicating that the pathways for AA and DHA incorporation are partially different. In control cells, the transfer of AA occurred mainly from diacyl glycerophosphocholine (GroPCho) to alkenylacyl-glycerophosphoethanolamine (GroPEtn) and to a lesser extent from diacyl-GroPCho to diacyl-GroPEtn. [3H]DHA was redistributed from diacyl-GroPCho and alkylacyl-GroPEtn to alkenylacyl GroPEtn. BEL treatment inhibited completely the redistributrion of AA within diacyl-GroPCho and diacyl -GroPEtn and reduced the [3H]DHA content of diacyl GroPEtn, indicating that a BEL-sensitive iPLA2 controls the redistribution of polyunsaturated fatty acids to diacyl-GroPEtn. In contrast the redistribution of radioactive AA and DHA to alkenylacyl-GroPEtn was almost insensitive to BEL. The analysis of substrate specificity and BEL sensitivity of iPLA2 activity indicates that UIII cells exhibit at least two isoforms of iPLA2, one of which is BEL sensitive and quite selective of diacyl species, and another one that is insensitive to BEL and selective for alkenylacyl-GroPEtn. Taken together, these results suggest that several iPLA2 participate independently in the remodelling of UIII cell phospholipids. PMID- 11106424 TI - Limited heme synthesis in porphobilinogen deaminase-deficient mice impairs transcriptional activation of specific cytochrome P450 genes by phenobarbital. AB - Heme is not only a very important prosthetic group that modulates the structure and activity of heme proteins but also a regulatory molecule that controls metabolic pathways and the biosynthesis of various proteins. However, investigation into heme regulatory effects in higher vertebrates has been hampered by the lack of a suitable animal model. A knockout mouse with targeted disruption of porphobilinogen deaminase, the third enzyme of the heme pathway, has been generated in our laboratory and used in the present study as an in vivo model of heme deficiency to explore diverse heme regulatory properties. In this model with a defined heme disturbance, we observed a superinductive response of delta-aminolevulinate synthase, the first enzyme in heme synthesis, after phenobarbital treatment. We also found that limited heme is associated with decreased induction of cytochrome P450 by phenobarbital as a consequence of impaired gene transcription. This inhibitory effect is isoenzyme-specific, being significant for cyp2a5. The activity and mRNA level of this particular cytochrome P450 are significantly lower in the phenobarbital-induced porphobilinogen deaminase-deficient mice (55% and 43%, respectively), but its expression can be restored to normal values when exogenous heme is administered. Other heme proteins, namely neuronal nitric oxide synthase and soluble guanylate cyclase, function normally in mice with limited heme. Our results demonstrate that the expression of various heme proteins is differentially regulated in conditions of reduced heme availability. Moreover, our findings emphasize the importance of heme protein function in the genesis of pathophysiological manifestations in acute intermittent porphyria. PMID- 11106425 TI - Conserved amino acids at the C-terminus of rat phospholipase D1 are essential for enzymatic activity. AB - Rat brain phospholipase D1 (rPLD1) has two highly conserved motifs [H(X)K(X)4D, denoted HKD] located at the N-terminal and C-terminal halves, which are required for activity. Association of the two halves is essential for rPLD1 activity, which probably brings the two HKD domains together to form a catalytic center. In the present study, we find that an intact C-terminus is also essential for the catalytic activity of rPLD1. Serial deletion of the last four amino acids, EVWT, which are conserved in all mammalian PLD isoforms, abolished the catalytic activity of rPLD1. This loss of catalytic activity was not due to a lack of association of the N-terminal and C-terminal halves. Mutations of the last three amino acids showed that substitutions with charged or less hydrophobic amino acids all reduced PLD activity. For example, mutations of Thr1036 and Val1034 to Asp or Lys caused marked inactivation, whereas mutation to other amino acids had less effect. Mutation of Trp1035 to Leu, Ala, His or Tyr caused complete inactivation, whereas mutation of Glu1033 to Ala enhanced activity. The size of the amino acids at the C-terminus also affected the catalytic activity of PLD, reduced activity being observed with conservative mutations within the EVWT sequence (such as T/S, V/L or W/F). The enzyme was also inactivated by the addition of Ala or Val to the C-terminus of this sequence. Interestingly, the inactive C-terminal mutants could be complemented by cotransfection with a wild type C-terminal half to restore PLD activity in vivo. These data demonstrate that the integrity of the C-terminus of rPLD1 is essential for its catalytic activity. Important features are the hydrophobicity, charge and size of the four conserved C-terminal amino acids. It is proposed that these play important roles in maintaining a functional catalytic structure by interacting with a specific domain within rPLD1. PMID- 11106426 TI - Structures of two cell wall-associated polysaccharides of a Streptococcus mitis biovar 1 strain. A unique teichoic acid-like polysaccharide and the group O antigen which is a C-polysaccharide in common with pneumococci. AB - The cell wall of Streptococcus mitis biovar 1 strain SK137 contains the C polysaccharide known as the common antigen of a closely related species Streptococcus pneumoniae, and a teichoic acid-like polysaccharide with a unique structure. The two polysaccharides are different entities and could be partially separated by gel chromatography. The structures of the two polysaccharides were determined by chemical methods and by NMR spectroscopy. The teichoic acid-like polymer has a heptasaccharide phosphate repeating unit with the following structure: The structure neither contains ribitol nor glycerol phosphate as classical teichoic acids do, thus we have used the expression teichoic acid-like for this polysaccharide. The following structure of the C-polysaccharide repeating unit was established: where AAT is 2-acetamido-4-amino-2,4, 6-trideoxy D-galactose. It has a carbohydrate backbone identical to that of one of the two structures of C-polysaccharide previously identified in S. pneumoniae. C polysaccharide of S. mitis is characterized by the presence, in each repeating unit, of two residues of phosphocholine and both galactosamine residues in the N acetylated form. Immunochemical analysis showed that C-polysaccharide constitutes the Lancefield group O antigen. Studies using mAbs directed against the backbone and against the phosphocholine moiety of the C-polysaccharide revealed several different patterns of these epitopes among 95 S. mitis and Streptococcus oralis strains tested and the exclusive presence of the group O antigen in the majority of S. mitis biovar 1 strains. PMID- 11106427 TI - Sites of limited proteolysis in the pyruvate decarboxylase component of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus and their role in catalysis. AB - The E1 component (pyruvate decarboxylase) of the pyruvate dehydrogenase complex of Bacillus stearothermophilus is a heterotetramer (alpha2beta2) of E1alpha and E1beta polypeptide chains. The domain structure of the E1alpha and E1beta chains, and the protein-protein interactions involved in assembly, have been studied by means of limited proteolysis. It appears that there may be two conformers of E1alpha in the E1 heterotetramer, one being more susceptible to proteolysis than the other. A highly conserved region in E1alpha, part of a surface loop at the entrance to the active site, is the most susceptible to cleavage in E1 (alpha2beta2). As a result, the oxidative decarboxylation of pyruvate catalysed by E1 in the presence of dichlorophenol indophenol as an artificial electron acceptor is markedly enhanced, but the reductive acetylation of a free lipoyl domain is unchanged. The parameters of the interaction between cleaved E1 and the peripheral subunit-binding domain of the dihydrolipoyl acetyltransferase E2 component are identical to those of the wild-type E1. However, a pyruvate dehydrogenase complex assembled in vitro with cleaved E1p exhibits a markedly lower overall catalytic activity than that assembled with untreated E1. This implies that active site coupling between the E1 and E2 components has been impaired. This has important implications for the way in which a tethered lipoyl domain can interact with E1 in the assembled complex. PMID- 11106428 TI - Identification of IkappaBalpha as a substrate of Fas-associated phosphatase-1. AB - Fas (APO-1/CD95), a member of the tumor necrosis factor receptor (TNFR)/nerve growth factor receptor (NGFR) superfamily, is a cell-surface molecule that induces apoptosis upon activation. Fas-associated phosphatase-1 (FAP-1) is a 250 kDa protein tyrosine phosphatase (PTP) that is associated with the negative regulatory domain of Fas (C-terminal 15 amino acids). Human tumor cell lines become resistant to Fas-mediated apoptosis when transfected with FAP-1, indicating that FAP-1 functions as a negative regulator in Fas-mediated death signaling. However, the mechanisms by which FAP-1 inhibits apoptosis are still unclear. In order to determine how FAP-1 affects the signaling mediated by Fas, we set out to identify substrates of FAP-1. Toward this end, we prepared synthetic proteins with either the catalytic domain of FAP-1 (C-terminal 399 amino acids) or its inactive form (Cys2408-->Ser) fused to glutathione-S transferase (GST). Using an in vitro dephosphorylation reaction, we found that FAP-1 dephosphorylates IkappaBalpha. Furthermore, a substrate trapping mutant was found to bind tyrosine-phosphorylated IkappaBalpha. Taken together, our data confirm that IkappaBalpha is a substrate of FAP-1. PMID- 11106430 TI - Dissection of the enzymatic and immunologic functions of macrophage migration inhibitory factor. Full immunologic activity of N-terminally truncated mutants. AB - Macrophage migration inhibitory factor (MIF) is a cytokine with broad regulatory functions in innate immunity. MIF belongs to the few cytokines displaying catalytic activities, i.e. MIF has a Pro2-dependent tautomerase and a Cys-Ala-Leu Cys (CALC) cysteine-based thiol-protein oxidoreductase activity. Previous studies have addressed the roles of the catalytic site residues and the C-terminus. The two activities have not been directly compared. Here we report on the N-terminal mutational analysis and minimization of MIF and on a dissection of the two catalytic activities by comparing mutants P2AMIF, Delta4MIF, Delta5MIF, Delta6MIF, Delta7MIF, Delta8MIF, and Delta10MIF with the cysteine mutants of MIF. As N-terminal deletion was predicted to interfere with protein structure due to disruption of the central beta sheet, it was surprising that deletion of up to six N-terminal residues resulted in normally expressed proteins with wild-type conformation. Strikingly, such mutants exhibited full MIF-specific immunologic activity. While mutation of Pro2 eliminated tautomerase activity, the CALC cysteine residues had no influence on this activity. However, mutant C81SMIF, which otherwise has full biologic activity, only had 32% tautomerase activity. Deletion of four N-terminal residues did not interfere with insulin reduction by MIF. By contrast, reduction of 2-hydroxyethyldisulfide (HED) was markedly affected by N-terminal manipulation, with P2AMIF and Delta2MIF exhibiting 40% activity, and Delta4MIF completely failing to reduce HED. This study constitutes the first comparison of the two catalytic activities of MIF and should assist in understanding the molecular links between the catalytic and immunologic activities of this cytokine and in providing guidelines for N-terminal protein minimization. PMID- 11106429 TI - Cloning, expression and characterization of a cDNA (6A8) encoding a novel human alpha-mannosidase. AB - A 3300-bp cDNA (6A8) has been isolated from a human tonsil cell lambdagt11 cDNA library (GenBank accession number: AF044414). The 6A8 gene is localized on human chromosome 13q31-32. Its cDNA has an open reading frame from position 57 bp to 3243 bp, encoding a 1062 amino-acid polypeptide. The sequence of the polypeptide has 89% identity to rat liver ER alpha-mannosidase. Homogenates of COS-7 cells transfected with 6A8 cDNA showed an enhanced enzymatic activity with p-nitro phenyl-alpha-D-mannopyranoside, which was not inhibited by swainsonine. These data suggest that 6A8 alpha-mannosidase belongs to the class II alpha mannosidase. Western blot analysis showed a band for 6A8 cDNA encoded protein of approximately 120 kDa. Northern blot analysis revealed two 6A8 mRNA transcripts with different tissue distribution. Enhanced concanavalin A (ConA) binding to CNE 2L2 cells transfected with a reverse 6A8 DNA was observed, indicating that the 6A8 protein is an important cellular alpha-mannosidase. PMID- 11106431 TI - Purification and characterization of a new, ubiquitously distributed class of microtubule-associated protein with molecular mass 250 kDa. AB - A heat-stable microtubule-associated protein (MAP) with relative molecular mass 250 000, termed 250-kDa MAP, was purified from bovine adrenal cortex. It is classified as a MAP subspecies distinct from MAP1, MAP2, tau, and MAP4, as judged from its electrophoretic mobility, heat stability and immunoreactivity. Purified 250-kDa MAP was able to bind to taxol-stabilized microtubules, although it lacked the ability to polymerize purified tubulin into microtubules. Western-blot analysis showed that this MAP was expressed ubiquitously in mammalian tissues. Immunofluorescence microscopy revealed that polyclonal antibodies raised against 250-kDa MAP stained many punctate structures in the cytoplasm of cultured cells. Blurry cytosolic staining was also observed. Judging from the result of nocodazole treatment, the punctate structures were associated with the microtubule network throughout the cytoplasm, while cytosolic 250-kDa MAP colocalized with free tubulin. Under electron microscopy, 250-kDa MAP has the appearance of a hollow sphere of about 12 nm diameter. PMID- 11106432 TI - Mercuric chloride activates the Src-family protein tyrosine kinase, Hck in myelomonocytic cells. AB - Hck is a member of the Src-family of protein tyrosine kinases that appears to function in mature leukocytes to communicate a number of extracellular signals including various cytokines. In this study we show that the thiol-reactive heavy metal, mercuric chloride (HgCl2) induces rapid and robust activation of tyrosine phosphorylation within human myelomonocytic cells. This increase in tyrosine phosphorylated proteins requires the activity of Hck because both kinase inactive alleles of Hck and pharmacological inhibitors selective for the Src-family kinases are able to abrogate the cellular response to HgCl2. Furthermore, ectopic expression of Hck in murine fibroblasts is able to confer HgCl2 responsiveness, as indicated by an increase in tyrosine-phosphorylated proteins to a normally nonresponsive cell line. Concomitant with the activation of Hck, there is a physical association of Hck with another cytoplasmic protein tyrosine kinase, Syk. The ability of HgCl2 to activate Src-family kinases such as Hck in hematopoietic cells may help explain why exposure to the heavy metal is associated with immune system dysfunction in rodents as well as humans. PMID- 11106434 TI - Modulation of activity and substrate specificity by modifying the backbone length of the distant interdomain loop of D-amino acid aminotransferase. AB - The activity and substrate specificity of D-amino acid aminotransferase (D-AAT) (EC 2.6.1.21) can be rationally modulated by replacing the loop core (P119-R120 P121) with glycine chains of different lengths: 1, 3, or 5 glycines. The mutant enzymes were much more active than the wild-type enzyme in the overall reactions between various amino acids and pyruvate. The presteady-state kinetic analyses of half-reactions revealed that the 5-glycine mutant has the highest affinity (Kd) among all mutant enzymes and the wild-type enzyme towards various amino acids except D-aspartate. The 5-glycine mutant was much more efficient as a catalyst than the wild-type enzyme because the mutant enzyme showed the highest value of specificity constant (kmax/Kd) for all amino acids except D-aspartate and D glutamate. The kmax/Kd values of the three mutants decreased with decrease in glycine chain length for each amino acid examined. Our findings may provide a new approach to rational modulation of enzymes. PMID- 11106433 TI - Cloning and functional characterization of the 5' regulatory region of the human mitochondrial glycerol-3-phosphate dehydrogenase gene. Lack of 3,5,3' triiodothyronine responsiveness in adipose tissue. AB - We report data on the structural and functional characterization of the 5' flanking region of the human mitochondrial glycerol-3-phosphate dehydrogenase (mtGPDH) gene. We found two regions upstream of 5'-untranslated sequences exhibiting promoter activity in transient transfection assays. Transcription start sites and potential regulatory sites in both promoter regions were defined. The proximal promoter was approximately sevenfold more active than the distal one in most cell lines, but it was only twice as active in a neuroblastoma cell line. These observations seem to indicate that the rate of transcription, as well as the tissue-specific expression of the human mtGPDH gene, is the result of a combinatorial effect of transcription factors on at least two promoters. 3,5,3' Triiodothyronine failed to alter the transcriptional activity of human mtGPDH promoter(s) constructs in transient transfection assays. Although this finding seems to be in conflict with the reported effect of 3,5,3'-triiodothyronine in rodents, it is consistent with our observation of 3,5, 3'-triiodothyronine stimulation of mtGPDH activity in primary cultures of rat adipocytes, but not human cultured adipocytes, suggesting distinct regulation of this gene in both species. PMID- 11106435 TI - Identification and characterization of a novel transcriptional regulator, MatR, for malonate metabolism in Rhizobium leguminosarum bv. trifolii. AB - A novel gene, matR, located upstream of matABC, transcribed in the opposite direction, and encoding a putative regulatory protein by sequence analysis was discovered from Rhizobium leguminosarum bv. trifolii. The matA, matB, and matC genes encode malonyl-CoA decarboxylase, malonyl-CoA synthetase, and a presumed malonate transporter, respectively. Together, these enzymes catalyze the uptake and conversion of malonate to acetyl-CoA. The deduced amino-acid sequence of matR showed sequence similarity with GntR from Bacillus subtilis in the N-terminal region encoding a helix-turn-helix domain. Electrophoretic mobility shift assay indicated that MatR bound to a fragment of DNA corresponding to the mat promoter region. The addition of malonate or methylmalonate increased the association of MatR and DNA fragment. DNase I footprinting assays identified a MatR binding site encompassing 66 nucleotides near the mat promoter. The mat operator region included an inverted repeat (TCTTGTA/TACACGA) centered -46.5 relative to the transcription start site. Transcriptional assays, using the luciferase gene, revealed that MatR represses transcription from the mat promoter and malonate alleviates MatR-mediated repression effect on the expression of Pmat-luc+ reporter fusion. PMID- 11106436 TI - The structure of the nonreducing terminal groups in the O-specific polysaccharides from two strains of Bordetella bronchiseptica. AB - The structures of the polysaccharide chains of the LPS from Bordetella bronchiseptica strains 110H and Bp512 were analysed by NMR spectroscopy and mass spectrometry. The polysaccharides consist of alpha-(1-4)-linked 2,3-diacetamido 2,3-dideoxy-L-galacturonic acid repeating units. Polysaccharides from both strains have 2,3, 4-triamino-2,3,4-trideoxy-alpha-galacturonamide derivatives at their nonreducing ends, a monosaccharide identified for the first time in nature. The polymers from the two strains differ in the nature of the acylation of the amino groups of this monosaccharide. In the strain 110H, the residue is formylated at positions 3 and 4, and has N-formyl-L-alanyl or L-alanyl substituents at N-2. In the strain Bp512, the amino group at position 2 is acetylated, at position 3 it is formylated, and the amino group at position 4 bears a 2-methoxypropionyl substituent. The distribution of the acyl groups was determined from long range 1H-13C correlation (HMBC) NMR spectra. Measurement of the spectra under different pH conditions showed that carboxyl groups of the inner uronic acid residues of the polymeric chain are free, and that carboxyl groups of the terminal residues are amidated. These conclusions were confirmed by the results of mass spectrometric analysis. PMID- 11106437 TI - The rat ortholog of the presumptive flounder antifreeze enhancer-binding protein is a helicase domain-containing protein. AB - The expression of winter flounder liver-type antifreeze protein (wflAFP) genes is tissue-specific and under seasonal and hormonal regulation. The only intron of the major wflAFP gene was demonstrated to be a liver-specific enhancer in both mammalian cell lines and flounder hepatocytes. Element B, the core enhancer sequence, was shown to interact specifically with a liver-enriched transcription factor, CCAAT/enhancer-binding protein alpha (C/EBPalpha), as well as a presumptive antifreeze enhancer-binding protein (AEP). In this study, the identity of the rat AEP ortholog was revealed via its DNA-protein interaction with element B. It is a helicase-domain-containing protein, 988 amino acids in length, and is homologous to mouse Smubp-2, hamster Rip-1 and human Smubp-2. The specific binding between element B and AEP was confirmed by South-Western analysis and gel retardation assays. Residues in element B important to this interaction were identified by methylation interference assays. Mutation on one of the residues disrupted the binding between element B and AEP and its enhancer activity was significantly reduced, suggesting that AEP is essential for the transactivation of the wflAFP gene intron. The rat AEP is ubiquitously expressed in various tissues, and the flounder homolog is present as shown by genomic Southern analysis. The potential role of AEP in regulating the flounder AFP gene expression is discussed. PMID- 11106438 TI - Expression and characterization of truncated forms of humanized L243 IgG1. Architectural features can influence synthesis of its oligosaccharide chains and affect superoxide production triggered through human Fcgamma receptor I. AB - The properties of IgG and its subcomponents are being exploited to generate new therapeutics with selected biological activities. In this study, a series of truncated, humanized IgG1 antibodies was expressed in Chinese hamster ovary cells, to evaluate the contribution of structural components to glycosylation and function. The series includes L243 IgG1 (alpha-MHC Class II) lacking a CH3 domain pair (DeltaCH3-IgG1), single-chain Fv fusion proteins with Fc or a hinge-CH2 domain, Fc with/out a hinge, and a single CH2 domain. Glycosylation of IgG Fc is important for recognition by effector ligands such as Fcgamma receptors. HPLC analysis of released and pyridylaminated oligosaccharides indicates that intact IgG1 and scFvFc antibodies are galactosylated and sialylated to levels similar to those observed previously for normal human IgG1. The truncated forms express increased levels of digalactosylated (30-83%) or sialylated (9-21%) oligosaccharide chains with the highest levels observed for the single CH2 domain. These data show which architectural components influence IgG glycosylation processing and that the (CH3)2 pair is particularly influential. When MHC Class II bearing (JY) cells were sensitized with L243 DeltaCH3-IgG1, scFvFc, or scFvhCH2 they elicited superoxide production, from U937 cells, at levels of 35-45% relative to that obtained for intact L243 IgG1 (100%). Mild reduction and alkylation of the hinge disulphide bonds of scFvhCH2 greatly decreased its capacity to trigger superoxide production. Thus, the L243 scFvhCH2 homo-dimer constitutes the minimal truncated form that binds the MHC Class II antigen and triggers superoxide production through FcgammaRI. PMID- 11106439 TI - Functional characterization of isoschizomeric His-Cys box homing endonucleases from Naegleria. AB - Several species within the amoeboflagellate genus Naegleria harbor an optional ORF containing group I introns in their nuclear small subunit ribosomal DNA. The different ORFs encode homing endonucleases with 65 to 95% identity at the amino acid level. I-NjaI, I-NanI and I-NitI, from introns in Naegleria jamiesoni, N. andersoni and N. italica, respectively, were analyzed in more detail and found to be isoschizomeric endonucleases that recognize and cleave an approximal 19-bp partially symmetrical sequence, creating a pentanucleotide 3' overhang upon cleavage. The optimal conditions for cleavage activity with respect to temperature, pH, salt and divalent metal ions were investigated. The optimal cleavage temperature for all three endonucleases was found to be 37 degrees C and the activity was dependent on the concentration of NaCl with an optimum at 200 mM. Divalent metal ions, primarily Mg2+, are essential for Naegleria endonuclease activity. Whereas both Mn2+ and Ca2+ could substitute for Mg2+, but with a slower cleavage rate, Zn2+ was unable to support cleavage. Interestingly, the pH dependence of DNA cleavage was found to vary significantly between the I-NitI and I-NjaI/I-NanI endonucleases with optimal pH values at 6.5 and 9, respectively. Site-directed mutagenesis of conserved I-NjaI residues strongly supports the hypothesis that Naegleria homing endonucleases share a similar zinc-binding structure and active site with the His-Cys box homing endonuclease I-PpoI. PMID- 11106440 TI - Making a difference: a day in the life of a flood relief volunteer in Mozambique. PMID- 11106441 TI - Emergency nursing: promises to keep! PMID- 11106442 TI - Acute coronary syndromes in the emergency department: new millennium, new mentality. PMID- 11106443 TI - More on the RN shortage and the use of UAPs. PMID- 11106444 TI - Policies hinder nursing staff. PMID- 11106445 TI - More on the RN shortage and the use of UAPs. PMID- 11106446 TI - In saving others' lives, ED nurses risk their own. PMID- 11106447 TI - Hearing after age 50. PMID- 11106448 TI - More on treating an agitated, irate minor against her will. PMID- 11106449 TI - Taking a stand on gun safety. PMID- 11106450 TI - More on music therapy in the emergency department. PMID- 11106452 TI - ED "hold" patients: is their care also being held? AB - INTRODUCTION: Patient care should be governed by the same standards of care regardless of location within a hospital system. Little is known about adherence to standards of care for admitted patients who are "held" in the emergency department because of an unavailable inpatient bed. For example, is there a difference in the timeliness of nursing assessments and initial antibiotic administration for patients with pneumonia who are held in the emergency department compared with those who are directly admitted to an inpatient bed? METHODS: A descriptive comparison research design with 2 known groups ("ED hold" patients and "ED direct-admit" patients) was used. A convenience sample of 104 closed medical records was obtained from a Midwestern hospital for a retrospective chart audit. RESULTS: Patients held in the emergency department had their blood pressure, pulse, respiratory rate, and oxygen saturation recorded with greater frequency compared with patients who were directly admitted. Directly admitted patients had their intake and output and their temperature noted with greater frequency. Although there was no statistically significant difference in the timeliness of respiratory assessments or antibiotic administration, antibiotics were delivered to the ED direct-admit group an average of 1 hour earlier than to the ED hold group. DISCUSSION: When measuring only the frequency and/or timeliness of taking vital signs, recording intake and output, and administering antibiotics, there were statistical differences between "hold" patients and "direct-admit" patients. In some instances, ED "hold" patients actually received more timely assessment than did "direct-admit" patients. Direct-admit patients received antibiotics an average of 1 hour sooner. More studies are necessary to evaluate other aspects of care that may or may not be compromised when admitted patients are "held" in the emergency department. PMID- 11106451 TI - A 30-year-old woman with possible unknown ingestion of date rape drugs. PMID- 11106453 TI - Seeking care for nonurgent medical conditions in the emergency department: through the eyes of the patient. AB - INTRODUCTION: The policy goal of shifting nonurgent visits from the emergency department to nonemergency health care settings is commonly devised, planned, and implemented without considering patients' perspectives. The purpose of this study was to gain an understanding of the context in which patients choose to seek health care in an emergency department. Human science provided the framework for this exploratory descriptive research study. METHODS: This study was conducted at an urban, university emergency department in Denver, Colo. Uninsured adult patients triaged as nonurgent who were being discharged home were eligible to participate. Eligible patients from 15 randomly selected shifts were asked to participate. Following their ED visit, open-ended interviews began with the question, "Can you tell me the story, or the chain of events, that led to your coming to the emergency department today?" Each interview was audiotaped. Transcripts were analyzed to identify common themes. Patients also rated their severity of illness from 1 (not severe) to 5 (life-threatening), and they rated their satisfaction with the health care they received from 1 (not satisfied) to 5 (extremely satisfied). RESULTS: The 30 study participants ranged in age from 17 to 60 years; 22 participants (73%) were women. Most patients (73%) rated their severity of illness as 3 or less and their satisfaction with the health care they received as 4 or more (83%). Five themes for seeking care were identified: (1) toughing it out, (2) symptoms overwhelming self-care measures, (3) calling a friend, (4) nowhere else to go, and (5) convenience. Despite the fact that the patients had nonurgent medical problems, their stories revealed that distress in their lives had influenced their need for emergency care. CONCLUSIONS: Access was prominent in the minds of uninsured patients seeking ED care for nonurgent medical diagnoses. Typically, patients did not perceive themselves as having an urgent problem, had been unsuccessful in gaining access to alternative non-ED health care settings, and found the emergency department to be a convenient and quality source of health care. The patients' stories relayed a context for ED visits that goes beyond medical diagnoses. This perspective has important implications for quality care delivery and for including patients in planning ways to access emergency health care. PMID- 11106454 TI - A successful emergency angioplasty program for patients with acute coronary syndrome in a community hospital setting: Cape Cod Hospital's 3-year experience. PMID- 11106455 TI - Nursing--THEN and NOW. PMID- 11106457 TI - The nursing shortage: what can we do? PMID- 11106458 TI - Asthma and latex allergy. PMID- 11106459 TI - Nine assessment points for interfacility transports. PMID- 11106461 TI - Volunteer? Who, me? PMID- 11106462 TI - Two nurses and a doctor: health care workers allege retaliation for blowing the whistle on understaffing. PMID- 11106464 TI - Assessing critical thinking of applicants to the Master of Science in Nursing Emergency Nurse Practitioner Option with clinical case scenario-based interviewing. PMID- 11106465 TI - Body piercing: old traditions creating new challenges. PMID- 11106466 TI - One RN's journey into policy work. PMID- 11106467 TI - It can happen to you. PMID- 11106468 TI - Workers' compensation, disability insurance, and Social Security benefits clarified: an interview with a disability expert by Joanne Venturella. PMID- 11106469 TI - Conducting research in the emergency department: respect ED nurses' workload and recognize their contribution. PMID- 11106470 TI - Identification and documentation of bite marks. PMID- 11106471 TI - A quick examination of the lower extremity. PMID- 11106472 TI - An adult male with facial swelling, erythema, and sensation of arm tightness. PMID- 11106476 TI - Emergency management of acute coronary syndromes. PMID- 11106473 TI - Doug Bouldin-an advocate for patients' bill of rights. PMID- 11106477 TI - The structure of UDP-N-acetylglucosamine 2-epimerase reveals homology to phosphoglycosyl transferases. AB - Bacterial UDP-N-acetylglucosamine 2-epimerase catalyzes the reversible epimerization at C-2 of UDP-N-acetylglucosamine (UDP-GlcNAc) and thereby provides bacteria with UDP-N-acetylmannosamine (UDP-ManNAc), the activated donor of ManNAc residues. ManNAc is critical for several processes in bacteria, including formation of the antiphagocytic capsular polysaccharide of pathogens such as Streptococcus pneumoniae types 19F and 19A. We have determined the X-ray structure (2.5 A) of UDP-GlcNAc 2-epimerase with bound UDP and identified a previously unsuspected structural homology with the enzymes glycogen phosphorylase and T4 phage beta-glucosyltransferase. The relationship to these phosphoglycosyl transferases is very intriguing in terms of possible similarities in the catalytic mechanisms. Specifically, this observation is consistent with the proposal that the UDP-GlcNAc 2-epimerase-catalyzed elimination and re addition of UDP to the glycal intermediate may proceed through a transition state with significant oxocarbenium ion-like character. The homodimeric epimerase is composed of two similar alpha/beta/alpha sandwich domains with the active site located in the deep cleft at the domain interface. Comparison of the multiple copies in the asymmetric unit has revealed that the epimerase can undergo a 10 degrees interdomain rotation that is implicated in the regulatory mechanism. A structure-based sequence alignment has identified several basic residues in the active site that may be involved in the proton transfer at C-2 or stabilization of the proposed oxocarbenium ion-like transition state. This insight into the structure of the bacterial epimerase is applicable to the homologous N-terminal domain of the bifunctional mammalian UDP-GlcNAc "hydrolyzing" 2-epimerase/ManNAc kinase that catalyzes the rate-determining step in the sialic acid biosynthetic pathway. PMID- 11106478 TI - An examination of the role of asp-177 in the His-Asp catalytic dyad of Leuconostoc mesenteroides glucose 6-phosphate dehydrogenase: X-ray structure and pH dependence of kinetic parameters of the D177N mutant enzyme. AB - The role of Asp-177 in the His-Asp catalytic dyad of glucose 6-phosphate dehydrogenase from Leuconostoc mesenteroides has been investigated by a structural and functional characterization of the D177N mutant enzyme. Its three dimensional structure has been determined by X-ray cryocrystallography in the presence of NAD(+) and in the presence of glucose 6-phosphate plus NADPH. The structure of a glucose 6-phosphate complex of a mutant (Q365C) with normal enzyme activity has also been determined and substrate binding compared. To understand the effect of Asp-177 on the ionization properties of the catalytic base His-240, the pH dependence of kinetic parameters has been determined for the D177N mutant and compared to that of the wild-type enzyme. The structures give details of glucose 6-phosphate binding and show that replacement of the Asp-177 of the catalytic dyad with asparagine does not affect the overall structure of glucose 6 phosphate dehydrogenase. Additionally, the evidence suggests that the productive tautomer of His-240 in the D177N mutant enzyme is stabilized by a hydrogen bond with Asn-177; hence, the mutation does not affect tautomer stabilization. We conclude, therefore, that the absence of a negatively charged aspartate at 177 accounts for the decrease in catalytic activity at pH 7.8. Structural analysis suggests that the pH dependence of the kinetic parameters of D177N glucose 6 phosphate dehydrogenase results from an ionized water molecule replacing the missing negative charge of the mutated Asp-177 at high pH. Glucose 6-phosphate binding orders and orients His-178 in the D177N-glucose 6-phosphate-NADPH ternary complex and appears to be necessary to form this water-binding site. PMID- 11106479 TI - Delineation of the roles of amino acids involved in the catalytic functions of Leuconostoc mesenteroides glucose 6-phosphate dehydrogenase. AB - The roles of particular amino acids in substrate and coenzyme binding and catalysis of glucose-6-phosphate dehydrogenase of Leuconostoc mesenteroides have been investigated by site-directed mutagenesis, kinetic analysis, and determination of binding constants. The enzyme from this species has functional dual NADP(+)/NAD(+) specificity. Previous investigations in our laboratories determined the three-dimensional structure. Kinetic studies showed an ordered mechanism for the NADP-linked reaction while the NAD-linked reaction is random. His-240 was identified as the catalytic base, and Arg-46 was identified as important for NADP(+) but not NAD(+) binding. Mutations have been selected on the basis of the three-dimensional structure. Kinetic studies of 14 mutant enzymes are reported and kinetic mechanisms are reported for 5 mutant enzymes. Fourteen substrate or coenzyme dissociation constants have been measured for 11 mutant enzymes. Roles of particular residues are inferred from k(cat), K(m), k(cat)/K(m), K(d), and changes in kinetic mechanism. Results for enzymes K182R, K182Q, K343R, and K343Q establish Lys-182 and Lys-343 as important in binding substrate both to free enzyme and during catalysis. Studies of mutant enzymes Y415F and Y179F showed no significant contribution for Tyr-415 to substrate binding and only a small contribution for Tyr-179. Changes in kinetics for T14A, Q47E, and R46A enzymes implicate these residues, to differing extents, in coenzyme binding and discrimination between NADP(+) and NAD(+). By the same measure, Lys-343 is also involved in defining coenzyme specificity. Decrease in k(cat) and k(cat)/K(m) for the D374Q mutant enzyme defines the way Asp-374, unique to L. mesenteroides G6PD, modulates stabilization of the enzyme during catalysis by its interaction with Lys-182. The greatly reduced k(cat) values of enzymes P149V and P149G indicate the importance of the cis conformation of Pro 149 in accessing the correct transition state. PMID- 11106480 TI - Consequences of cAMP-binding site mutations on the structural stability of the type I regulatory subunit of cAMP-dependent protein kinase. AB - The regulatory (R) subunit of cAMP-dependent protein kinase (cAPK) is a multidomain protein with two tandem cAMP-binding domains, A and B. The importance of cAMP binding on the stability of the R subunit was probed by intrinsic fluorescence and circular dichroism (CD) in the presence and absence of urea. Several mutants were characterized. The site-specific mutants R(R209K) and R(R333K) had defects in cAMP-binding sites A and B, respectively. R(M329W) had an additional tryptophan in domain B. Delta(260-379)R lacked Trp260 and domain B. The most destabilizing mutation was R209K. Both CD and fluorescence experiments carried out in the presence of urea showed a decrease in cooperativity of the unfolding, which also occurred at lower urea concentrations. Unlike native R, R(R209K) was not stabilized by excess cAMP. Additionally, CD revealed significant alterations in the secondary structure of the R209K mutant. Therefore, Arg209 is important not only as a contact site for cAMP binding but also for the intrinsic structural stability of the full-length protein. Introducing the comparable mutation into domain B, R333K, had a smaller effect on the integrity and stability of domain A. Unfolding was still cooperative; the protein was stabilized by excess cAMP, but the unfolding curve was biphasic. The R(M329W) mutant behaved functionally like the native protein. The Delta(260-379)R deletion mutant was not significantly different from wild-type RIalpha in its stability. Consequently, domain B and the interaction between Trp260 and cAMP bound to site A are not critical requirements for the structural stability of the cAPK regulatory subunit. PMID- 11106481 TI - Ubiquinone binding, ubiquinone exclusion, and detailed cofactor conformation in a mutant bacterial reaction center. AB - The X-ray crystal structure of a Rhodobacter sphaeroides reaction center with the mutation Ala M260 to Trp (AM260W) has been determined. Diffraction data were collected that were 97.6% complete between 30.0 and 2.1 A resolution. The electron density maps confirm the conclusions of a previous spectroscopic study, that the Q(A) ubiquinone is absent from the AM260W reaction center (Ridge, J. P., van Brederode, M. E., Goodwin, M. G., van Grondelle, R., and Jones, M. R. (1999) Photosynthesis Res. 59, 9-26). Exclusion of the Q(A) ubiquinone caused by the AM260W mutation is accompanied by a change in the packing of amino acids in the vicinity of the Q(A) site that form part of a loop that connects the DE and E helices of the M subunit. This repacking minimizes the volume of the cavity that results from the exclusion of the Q(A) ubiquinone, and further space is taken up by a feature in the electron density maps that has been modeled as a chloride ion. An unexpected finding is that the occupancy of the Q(B) site by ubiquinone appears to be high in the AM260W crystals, and as a result the position of the Q(B) ubiquinone is well-defined. The high quality of the electron density maps also reveals more precise information on the detailed conformation of the reaction center carotenoid, and we discuss the possibility of a bonding interaction between the methoxy group of the carotenoid and residue Trp M75. The conformation of the 2-acetyl carbonyl group in each of the reaction center bacteriochlorins is also discussed. PMID- 11106482 TI - Hybrid-cluster protein (HCP) from Desulfovibrio vulgaris (Hildenborough) at 1.6 A resolution. AB - The three-dimensional structure of the hybrid cluster protein from Desulfovibrio vulgaris (Hildenborough) has been determined at 1.6 A resolution using synchrotron X-ray radiation. The protein can be divided into three domains: an N terminal mainly alpha-helical domain and two similar domains comprising a central beta-sheet flanked by alpha-helices. The protein contains two 4Fe clusters with an edge-to-edge distance of 10.9 A. Four cysteine residues at the N-terminus of the protein are ligands to the iron atoms of a conventional [4Fe-4S] cubane cluster. The second cluster has an unusual asymmetric structure and has been named the hybrid cluster to reflect the variety of protein ligands, namely two mu sulfido bridges, two mu(2)-oxo bridges, and a further disordered bridging ligand. Anomalous differences in data collected at 1.488 A and close to the iron edge at 1.743 A have been used to confirm the identity of the metal and sulfur atoms. The hybrid cluster is buried in the center of the protein, but is accessible through a large hydrophobic cavity that runs the length of domain 3. Hydrophobic channels have previously been identified as access routes to the active centers in redox enzymes with gaseous substrates. The hybrid cluster is also accessible by a hydrophilic channel. The [4Fe-4S] cubane cluster is close to an indentation on the surface of the protein and can also be approached on the opposite side by a long solvent channel. At the present time, neither the significance of these channels nor, indeed, the function of the hybrid cluster protein is known. PMID- 11106483 TI - Guanine specific binding at a DNA junction formed by d[CG(5-BrU)ACG](2) with a topoisomerase poison in the presence of Co(2+) ions. AB - The structure of the duplex d[CG(5-BrU)ACG](2) bound to 9-bromophenazine-4 carboxamide has been solved through MAD phasing at 2.0 A resolution. It shows an unexpected and previously unreported intercalation cavity stabilized by the drug and novel binding modes of Co(2+) ions at certain guanine N7 sites. For the intercalation cavity the terminal cytosine is rotated to pair with the guanine of a symmetry-related duplex to create a pseudo-Holliday junction geometry, with two such cavities linked through the minor groove interactions of the N2/N3 guanine sites at an angle of 40 degrees, creating a quadruplex-like structure. The mode of binding of the drug is shown to be disordered, with the major conformations showing the side chain bound to the N7 position of adjacent guanines. The other end of the duplex exhibits a terminal base fraying in the presence of Co(2+) ions linking symmetry-related guanines, causing the helices to intertwine through the minor groove. The stabilization of the structure by the intercalating drug shows that this class of compound may bind to DNA junctions as well as duplex DNA or to strand-nicked DNA ('hemi-intercalated'), as in the cleavable complex. This suggests a structural basis for the dual poisoning of topoisomerase I and II enzymes by this family of drugs. PMID- 11106484 TI - High-resolution structure of the HNF-1alpha dimerization domain. AB - The N-terminal dimerization domain of the transcriptional activator hepatocyte nuclear factor-1alpha (HNF-1alpha) is essential for DNA binding and association of the transcriptional coactivator, DCoH (dimerization cofactor of HNF-1). To investigate the basis for dimerization of HNF-1 proteins, we determined the 1.2 A resolution X-ray crystal structure of the dimerization domain of HNF-1alpha (HNF p1). Phasing was facilitated by devising a simple synthesis for Fmoc selenomethionine and substituting leucine residues with selenomethionine. The HNF 1 dimerization domain forms a unique, four-helix bundle that is preserved with localized conformational shifts in the DCoH complex. In three different crystal forms, HNF-p1 displays subtle shifts in the conformation of the interhelix loop and the crossing angle between the amino- and carboxyl-terminal helices. In all three crystal forms, the HNF-p1 dimers pair through an exposed hydrophobic surface that also forms the binding site for DCoH. Conserved core residues in the dimerization domain of the homologous transcriptional regulator HNF-1beta rationalize the functional heterodimerization of the HNF-1alpha and HNF-1beta proteins. Mutations in HNF-1alpha are associated with maturity-onset diabetes of the young type 3 (MODY3), and the structure of HNF-p1 provides insights into the effects of three MODY3 mutations. PMID- 11106485 TI - Structural origins of the interfacial activation in Thermomyces (Humicola) lanuginosa lipase. AB - The already known X-ray structures of lipases provide little evidence about initial, discrete structural steps occurring in the first phases of their activation in the presence of lipids (process referred to as interfacial activation). To address this problem, five new Thermomyces (formerly Humicola) lanuginosa lipase (TlL) crystal structures have been solved and compared with four previously reported structures of this enzyme. The bias coming from different crystallization media has been minimized by the growth of all crystals under the same crystallization conditions, in the presence of detergent/lipid analogues, with low or high ionic strength as the only main variable. Resulting structures and their characteristic features allowed the identification of three structurally distinct species of this enzyme: low activity form (LA), activated form (A), and fully Active (FA) form. The isomerization of the Cys268-Cys22 disulfide, synchronized with the formation of a new, short alpha(0) helix and flipping of the Arg84 (Arginine switch) located in the lid's proximal hinge, have been postulated as the key, structural factors of the initial transitions between LA and A forms. The experimental results were supplemented by theoretical calculations. The magnitude of the activation barrier between LA (ground state) and A (end state) forms of TlL (10.6 kcal/mol) is comparable to the enthalpic barriers typical for ring flips and disulfide isomerizations at ambient temperatures. This suggests that the sequence of the structural changes, as exemplified in various TlL crystal structures, mirror those that may occur during interfacial activation. PMID- 11106486 TI - Cationic porphyrins promote the formation of i-motif DNA and bind peripherally by a nonintercalative mechanism. AB - Telomeric C-rich strands can form a noncanonical intercalated DNA structure known as an i-motif. We have studied the interactions of the cationic porphyrin 5,10,15,20-tetra-(N-methyl-4-pyridyl)porphine (TMPyP4) with the i-motif forms of several oligonucleotides containing telomeric sequences. TMPyP4 was found to promote the formation of the i-motif DNA structure. On the basis of (1)H NMR studies, we have created a model of the i-motif-TMPyP4 complex that is consistent with all the available experimental data. Two-dimensional NOESY data prompted us to conclude that TMPyP4 binds specifically to the edge of the intercalated DNA core by a nonintercalative mechanism. Since we have shown that TMPyP4 binds to and stabilizes the G-quadruplex form of the complementary G-rich telomeric strand, this study raises the intriguing possibility that TMPyP4 can trigger the formation of unusual DNA structures in both strands of the telomeres, which may in turn explain the recently documented biological effects of TMPyP4 in cancer cells. PMID- 11106487 TI - Externally added aFGF mutants do not require extensive unfolding for transport to the cytosol and the nucleus in NIH/3T3 cells. AB - Acidic fibroblast growth factor (aFGF) is transported to the cytosol and the nucleus when added to cells expressing FGF receptors, implying that aFGF must cross cellular membranes. Since protein translocation across membranes commonly requires extensive unfolding of the protein, we were interested in testing whether this is also necessary for membrane translocation of aFGF. We therefore constructed mutant growth factors with intramolecular disulfide bonds to prevent complete unfolding. Control experiments demonstrated that translocation of aFGF by the diphtheria toxin pathway, which requires extensive unfolding of the protein, was prevented by disulfide bond formation, indicating that the introduced disulfide bonds interfered with the unfolding of the growth factor. On the other hand, when the growth factor as such was added to cells expressing FGF receptors, the disulfide-bonded mutants were translocated to the cytosol and the nucleus equally well as wild-type aFGF. The possibility that the translocation of the mutants was due to reduction of the disulfide bonds prior to translocation was tested in experiments using an irreversibly cross-linked mutant. Also this mutant was transported to the cytosol and to the nucleus. The results suggest that extensive unfolding is not required for membrane translocation of aFGF. PMID- 11106488 TI - New photointermediates in the two photon signaling pathway of sensory rhodopsin I. AB - Sensory rhodopsin-I (SRI) functions as a color discriminating receptor in halobacterial phototaxis. SRI exists in the membrane as a molecular complex with a signal transducer protein. Excitation of its thermally stable form, SRI(587), generates a long-lived photointermediate of its photocycle, S(373), and an attractant phototactic response. S(373) decays thermally in a few seconds into SRI(587.) However, when S(373) is excited by UV-blue light, it photoconverts into SRI(587) in less than a second, generating a repellent phototactic response. Only one intermediate of this back-photoreaction, S(b)(510), is known. We studied the back-photoreaction in both native SRI and its transducer free form fSRI by measuring laser flash induced absorption changes of S(373) photoproducts from 100 ns to 1 s in the 350-750 nm range. Using global exponential fitting, we determined the spectra and kinetics of the photointermediates. S(373) and fS(373) when pumped with 355 nm laser light generate in less than 100 ns two intermediate species: a previously undetected species that absorbs maximally at about 410 nm, S(b)(410), and the previously described S(b)(510). These two intermediates appear to be in a rapid equilibrium, which probably entails protonation change of the Schiff base chromophore. At pH 6 this system relaxes to SRI(587) via another intermediate absorbing maximally around 550 nm, which thermally decays back to the ground state. The same intermediates are seen in the presence and absence of transducer; however, the kinetics are affected by binding of the transducer. PMID- 11106489 TI - Identification of unique amino acids that modulate CYP4A7 activity. AB - A multifamily sequence alignment of the rabbit CYP4A members with the known structure of CYP102 indicates amino acid differences falling within the so-called substrate recognition site(s) (SRS). Chimeric proteins constructed between CYP4A4 and CYP4A7 indicate that laurate activity is affected by the residues within SRS1 and prostaglandin activity is influenced by SRS2-3. Site-directed mutant proteins of CYP4A7 found laurate and arachidonate activity markedly diminished in the R90W mutant (SRS1) and somewhat decreased in W93S. While PGE(1) activity was only slightly increased, the mutant proteins H206Y and S255F (SRS2-3), on the other hand, exhibited remarkable increases in laurate and arachidonate metabolism (3 fold) above wild-type substrate metabolism. Mutant proteins H206Y, S255F, and H206Y/S255F but not R90W/W93S, wild-type CYP4A4, or CYP4A7 metabolized arachidonic acid in the absence of cytochrome b(5). Stopped-flow kinetic experiments were performed in a CO-saturated environment performed to estimate interaction rates of the monooxygenase reaction components. The mutant protein H206Y, which exhibits 3-fold higher than wild-type substrate activity, interacts with CPR at a rate at least 10 times faster than that of wild-type CYP4A7. These experimental results provide insight regarding the residues responsible for modulation of substrate specificity, affinity, and kinetics, as well as possible localization within the enzyme structure based on comparisons with homologous, known cytochrome P450 structures. PMID- 11106490 TI - Cloning, expression, and characterization of human cytosolic aminopeptidase P: a single manganese(II)-dependent enzyme. AB - The mammalian bradykinin-degrading enzyme aminopeptidase P (AP-P; E. C. 3.4.11.9) is a metal-dependent enzyme and is a member of the peptidase clan MG. AP-P exists as membrane-bound and cytosolic forms, which represent distinct gene products. A partially truncated clone encoding the cytosolic form was obtained from a human pancreatic cDNA library and the 5' region containing the initiating Met was obtained by 5' rapid accumulation of cDNA ends (RACE). The open reading frame encodes a protein of 623 amino acids with a calculated molecular mass of 69,886 Da. The full-length cDNA with a C-terminal hexahistidine tag was expressed in Escherichia coli and COS-1 cells and migrated on SDS-PAGE with a molecular mass of 71 kDa. The expressed cytosolic AP-P hydrolyzed the X-Pro bond of bradykinin and substance P but did not hydrolyze Gly-Pro-hydroxyPro. Hydrolysis of bradykinin was inhibited by 1,10-phenanthroline and by the specific inhibitor of the membrane-bound form of mammalian AP-P, apstatin. Inductively coupled plasma atomic emission spectroscopy of AP-P expressed in E. coli revealed the presence of 1 mol of manganese/mol of protein and insignificant amounts of cobalt, iron, and zinc. The enzymatic activity of AP-P was promoted in the presence of Mn(II), and this activation was increased further by the addition of glutathione. The only other metal ion to cause slight activation of the enzyme was Co(II), with Ca(II), Cu(II), Mg(II), Ni(II), and Zn(II) all being inhibitory. Removal of the metal ion from the protein was achieved by treatment with 1,10-phenanthroline. The metal-free enzyme was reactivated by the addition of Mn(II) and, partially, by Fe(II). Neither Co(II) nor Zn(II) reactivated the metal-free enzyme. On the basis of these data we propose that human cytosolic AP-P is a single metal ion dependent enzyme and that manganese is most likely the metal ion used in vivo. PMID- 11106491 TI - Identification of critical residues in the active site of porcine membrane-bound aminopeptidase P. AB - The membrane-bound form of mammalian aminopeptidase P (AP-P; EC 3.4. 11.9) is a mono-zinc-containing enzyme that lacks any of the typical metal binding motifs found in other zinc metalloproteases. To identify residues involved in metal binding and catalysis, sequence and structural information was used to align the sequence of porcine membrane-bound AP-P with other members of the peptidase clan MG, including Escherichia coli AP-P and methionyl aminopeptidases. Residues predicted to be critical for activity were mutated and the resultant proteins were expressed in COS-1 cells. Immunoelectrophoretic blot analysis was used to compare the levels of expression of the mutant proteins, and their ability to hydrolyze bradykinin and Gly-Pro-hydroxyPro was assessed. Asp449, Asp460, His523, Glu554, and Glu568 are predicted to serve as metal ion ligands in the active site, and mutagenesis of these residues resulted in fully glycosylated proteins that were catalytically inactive. Mutation of His429 and His532 also resulted in catalytically inactive proteins, and these residues, by analogy with E. coli AP P, are likely to play a role in shuttling protons during catalysis. These studies indicate that mammalian membrane-bound AP-P has an active-site configuration similar to that of other members of the peptidase clan MG, which is compatible with either a dual metal ion model or a single metal ion in the active site. The latter model is consistent, however, with the known metal stoichiometry of both the membrane-bound and cytosolic forms of AP-P and with a recently proposed model for methionyl aminopeptidase. PMID- 11106492 TI - Evidence for two distinct effector-binding sites in threonine deaminase by site directed mutagenesis, kinetic, and binding experiments. AB - A three-dimensional structure comparison between the dimeric regulatory serine binding domain of Escherichia coli D-3-phosphoglycerate dehydrogenase [Schuller, D. J., Grant, G. A., and Banaszak, L. J. (1995) Nat. Struct. Biol. 2, 69-76] and the regulatory domain of E. coli threonine deaminase [Gallagher, D. T., Gilliland, G. L., Xiao, G., Zondlo, J., Fisher, K. E., Chinchilla, D. , and Eisenstein, E. (1998) Structure 6, 465-475] led us to make the hypothesis that threonine deaminase could have two binding sites per monomer. To test this hypothesis about the corresponding plant enzyme, site-directed mutagenesis was carried out on the recombinant Arabidopsis thaliana threonine deaminase. Kinetic and binding experiments demonstrated for the first time that each regulatory domain of the monomers of A. thaliana threonine deaminase possesses two different effector-binding sites constituted in part by Y449 and Y543. Our results demonstrate that Y449 belongs to a high-affinity binding site whose interaction with a first isoleucine induces conformational modifications yielding a conformer displaying a higher activity and with enhanced ability to bind a second isoleucine on a lower-affinity binding site containing Y543. Isoleucine interaction with this latter binding site is responsible for conformational modifications leading to final inhibition of the enzyme. Y449 interacts with both regulators, isoleucine and valine. However, interaction of valine with the high affinity binding site induces different conformational modifications leading to reversal of isoleucine binding and reversal of inhibition. PMID- 11106493 TI - Reactivity of cysteine-49 and its influence on the activation of microsomal glutathione transferase 1: evidence for subunit interaction. AB - Microsomal glutathione transferase 1 is a homotrimeric detoxication enzyme protecting against electrophiles. The enzyme can also react with electrophiles, and when modification occurs at a unique Cys49 the reaction often results in activation. Here we describe the characterization of the chemical properties of this sulfhydryl (kinetic pK(a) was 8.8 +/- 0.3 and 9.0 +/- 0.1 with two different reagents) and we conclude that the protein environment does not lower the pK(a). Upon a direct comparison of the reactivity of Cys49 and low molecular weight thiols [L-Cys and glutathione (GSH)], the protein sulfhydryl displayed a 10-fold lower reactivity. The reactivity was correlated to reagent concentration in a linear fashion with a polar reagent, whereas the reactivity toward a hydrophobic reagent displayed saturation behavior (at low concentrations). This finding indicates that Cys49 is situated in a hydrophobic binding pocket. In a series of related quinones, activation occurs with the more reactive and less sterically hindered compounds. Thus, activation can be used to detect reactive intermediates during the metabolism of foreign compounds but certain intermediates can (and will) escape undetected. The reactivities of the three cysteines in the homotrimer were shown not to differ dramatically as the reaction of the protein with 4, 4'-dithiodipyridine could be fitted to a single exponential. On the basis of this result, a probabilistic expression could be used to relate the overall degree of modification to fractional activation. When N-ethylmaleimide activation (determined by the 1-chloro-2, 4-dinitrobenzene assay) was plotted against modification (determined with 4,4'-dithiodipyridine), a nonlinear relation was obtained, clearly showing that subunits do not function independently. The contribution to activation by single-, double-, and triple-modified trimers, were 0 +/- 0.06, 0.74 +/- 0.09, and 0.97 +/- 0.06, respectively. The double-modified enzyme appears partly activated, but this conclusion is more uncertain due to the possibility of independent modification of the purified enzyme upon storage. It is, however, clear that the single-modified enzyme is not activated whereas the triple-modified enzyme is fully activated. These observations together with the fact that MGST1 homotrimers bind only one substrate molecule (GSH) strongly support the view that subunits must interact in a functional manner. PMID- 11106494 TI - Regulation of the successive reaction catalyzed by rat neuronal nitric oxide synthase. AB - The rat neuronal nitric oxide synthase (nNOS) catalyzes two monooxygenase reactions successively from L-arginine (L-Arg) to L-citrulline (L-Cit) via N(omega)-hydroxy-L-arginine (OH-Arg) without most of OH-Arg leaving the substrate binding site. In the steady-state reaction conditions, the amount of OH-Arg produced is about 1/30-1/50 that of L-Cit. We found in this study using nNOS purified from an Escherichia coli expression system that the ratio of the amount of OH-Arg to L-Cit (OH-Arg/L-Cit) increased to about 1 at low concentration of NADPH. In one cycle of the nNOS reaction, the decrease in NADPH concentration was found to reduce the rates of two monooxygenase reactions but had little effect on the rate constant of OH-Arg dissociation from the enzyme. The addition of NADP(+), the competitive inhibitor for NADPH, caused the decrease in the rates of monooxygenase reactions in a single cycle of the reaction and the increase in the ratio of OH-Arg/L-Cit in the steady state. At low CaM concentrations, the ratio of OH-Arg/L-Cit was about the same as that at high CaM. In a single cycle of the nNOS reaction, the rate of monooxygenation was not altered by the CaM concentration but the amount of metabolized L-Arg decreased with the decrease in CaM concentration, showing that the amount of active nNOS was regulated by complex formation between nNOS and CaM. It becomes clear that there are two regulatory mechanisms for the successive reaction of nNOS. One controls the rates of monooxygenations and the other controls the amount of active species of nNOS. PMID- 11106495 TI - Engineered eglin c variants inhibit yeast and human proprotein processing proteases, Kex2 and furin. AB - We engineered eglin c, a potent subtilisin inhibitor, to create inhibitors for enzymes of the Kex2/furin family of proprotein processing proteases. A structural gene was synthesized that encoded "R(1)-eglin", having Arg at P(1) in the reactive site loop in place of Leu(45). Ten additional variants were created by cassette mutagenesis of R(1)-eglin. These polypeptides were expressed in Escherichia coli, purified to homogeneity, and their interactions with secreted, soluble Kex2 and furin were examined. R(1)-eglin itself was a modest inhibitor of Kex2, with a K(a) of approximately 10(7) M(-)(1). Substituting Arg (in R(4)R(1) eglin) or Met (in M(4)R(1)-eglin) for Pro(42) at P(4) created potent Kex2 inhibitors exhibiting K(a) values of approximately 10(9) M(-)(1). R(4)R(1)-eglin inhibited furin with a K(a) of 4.0 x 10(8) M(-)(1). Introduction of Lys at P(1), in place of Arg in R(4)R(1)-eglin reduced affinity only approximately 3-fold for Kex2 but 15-fold for furin. The stabilities of enzyme-inhibitor complexes were characterized by association and dissociation rate constants and visualized by polyacrylamide gel electrophoresis. R(4)R(1)-eglin formed stable 1:1 complexes with both Kex2 and furin. However, substitution of Lys at P(2) in place of Thr(44) resulted in eglin variants that inhibited both Kex2 and furin but which were eventually cleaved (temporary inhibition). Surprisingly, R(6)R(4)R(1)-eglin, in which Arg was substituted for Gly(40) in R(4)R(1)-eglin, exhibited stable, high-affinity complex formation with Kex2 (K(a) of 3.5 x 10(9) M(-)(1)) but temporary inhibition of furin. This suggests that enzyme-specific interactions can alter the conformation of the reactive site loop, converting a permanent inhibitor into a substrate. Eglin variants offer possible avenues for affinity purification, crystallization, and regulation of proprotein processing proteases. PMID- 11106496 TI - Escherichia coli LipA is a lipoyl synthase: in vitro biosynthesis of lipoylated pyruvate dehydrogenase complex from octanoyl-acyl carrier protein. AB - The Escherichia coli lipA gene product has been genetically linked to carbon sulfur bond formation in lipoic acid biosynthesis [Vanden Boom, T. J., Reed, K. E., and Cronan, J. E., Jr. (1991) J. Bacteriol. 173, 6411-6420], although in vitro lipoate biosynthesis with LipA has never been observed. In this study, the lipA gene and a hexahistidine tagged lipA construct (LipA-His) were overexpressed in E. coli as soluble proteins. The proteins were purified as a mixture of monomeric and dimeric species that contain approximately four iron atoms per LipA polypeptide and a similar amount of acid-labile sulfide. Electron paramagnetic resonance and electronic absorbance spectroscopy indicate that the proteins contain a mixture of [3Fe-4S] and [4Fe-4S] cluster states. Reduction with sodium dithionite results in small quantities of an S = 1/2 [4Fe-4S](1+) cluster with the majority of the protein containing a species consistent with an S = 0 [4Fe 4S](2+) cluster. LipA was assayed for lipoate or lipoyl-ACP formation using E. coli lipoate-protein ligase A (LplA) or lipoyl-[acyl-carrier-protein]-protein-N lipoyltransferase (LipB), respectively, to lipoylate apo-pyruvate dehydrogenase complex (apo-PDC) [Jordan, S. W., and Cronan, J. E. (1997) Methods Enzymol. 279, 176-183]. When sodium dithionite-reduced LipA was incubated with octanoyl-ACP, LipB, apo-PDC, and S-adenosyl methionine (AdoMet), lipoylated PDC was formed. As shown by this assay, octanoic acid is not a substrate for LipA. Confirmation that LipA catalyzes formation of lipoyl groups from octanoyl-ACP was obtained by MALDI mass spectrometry of a recombinant PDC lipoyl-binding domain that had been lipoylated in a LipA reaction. These results provide information about the mechanism of LipA catalysis and place LipA within the family of iron-sulfur proteins that utilize AdoMet for radical-based chemistry. PMID- 11106497 TI - Modulation by Ca(2+) and by membrane binding of the dynamics of domain III of annexin 2 (p36) and the annexin 2-p11 complex (p90): implications for their biochemical properties. AB - The modulation of the local structure and dynamics of domain III of annexin 2 (Anx2), in both the monomeric (p36) and heterotetrameric forms (p90), by calcium and by membrane binding was studied by time-resolved fluorescence intensity and anisotropy measurements of the single tryptophan residue (W212). The results yield the same dominant excited-state lifetime (1.4 ns) in both p36 and p90, suggesting that the conformation and environment of W212 are very similar. The fluorescence anisotropy decay data were analyzed by associative (two-dimensional) as well as nonassociative (one-dimensional) models. Although no statistical criterion is decisive for one model versus the other, only the associative model allows recovery of a physically relevant value of the Brownian rotational correlation of the protein. Using the associative model, a nanosecond flexibility is detectable in p90 but not in p36. When Ca(2+) binds in the millimolar concentration range to both forms of Anx2, a conformational change takes place leading to an increase of the major excited-state lifetime (2.6 ns) and to a suppression of the W212 local flexibility of p90. Binding to membranes of either p36 or p90 in the presence of Ca(2+) does not induce any conformational change other than that provoked by Ca(2+) binding alone. The W212 local flexibility in both proteins increases significantly, however, in their membrane-bound forms. In the presence of membranes, the conformation change of domain III in p90 displays a sensitivity to Ca(2+) 2 orders of magnitude higher than that of p36, reaching intracellular sub-micromolar concentration ranges. This higher Ca(2+) sensitivity correlates with the Ca(2+)-dependent membrane aggregation but not with their Ca(2+)-dependent binding to membranes. The significance of these structural and dynamical changes for the function of the protein is discussed. PMID- 11106498 TI - N-Terminal domain of annexin 2 regulates Ca(2+)-dependent membrane aggregation by the core domain: a site directed mutagenesis study. AB - Annexin 2 binds and aggregates biological membranes in a Ca(2+)-dependent manner. This protein exists as a monomer (p36) or as a heterotetramer (p90) in which two p36 chains are associated with a dimer of p11, a member of the S100 protein family. Protein kinase C phosphorylates the protein at the level of the N terminal tail on serines 11 and 25, thereby modifying its oligomeric structure and its properties of membrane aggregation. To analyze these effects, the properties of a series of mutants in which serines 11 and 25 were replaced by alanine and/or glutamic acid were investigated. The affinity for p11 light chain was decreased in the S11E mutants. Glutamic acid residues in positions 11 or 25 did not change membrane binding, either in the tetrameric or in the monomeric form. On the other hand, these mutations affected the aggregation properties of the two forms. For the tetramer, the aggregation efficiency was decreased but not the Ca(2+) sensitivity, whereas the latter was affected in the case of the monomer. The effects were stronger in the S11E mutants, and they were cumulative in the double mutant. They suggest a different conformation of the N-terminal domain in the mutants (and in the phosphorylated protein), a hypothesis which is supported by proteolysis experiments. This conformational change would affect aggregation by the monomer through a dimerization step. PMID- 11106499 TI - Activation of phosphatidylinositol-specific phospholipase C by HDL-associated lysosphingolipid. Involvement in mitogenesis but not in cholesterol efflux. AB - Our earlier studies demonstrated that high-density lipoproteins (HDLs) stimulate multiple signaling pathways, including activation of phosphatidylcholine-specific phospholipases C and D (PC-PLs) and phosphatidylinositol-specific phospholipase C (PI-PLC). However, only activation of PC-PLs was linked to the HDL-induced cholesterol efflux. In the study presented here, the role of HDL-induced PI-PLC activation was studied. In human skin fibroblasts, HDL potently induced PI-PLC as inferred from enhanced phosphatidylinositol bisphosphate (PtdInsP(2)) turnover and Ca(2+) mobilization. The major protein component of HDL, apo A-I, did not induce PtdInsP(2) turnover or Ca(2+) mobilization in these cells. Both HDL and apo A-I promoted cellular cholesterol efflux, whereas only HDL induced fibroblast proliferation. Inhibition of PI-PLC with U73122 or blocking intracellular Ca(2+) elevation with Ni(2+) or EGTA markedly reduced the extent of HDL-induced cell proliferation but had no effect on cholesterol efflux. In fibroblasts from patients with Tangier disease which are characterized by defective cholesterol efflux, neither HDL-induced PtdInsP(2) breakdown and Ca(2+) mobilization nor cell proliferation was impaired. HDL-induced fibroblast proliferation, PtdInsP(2) turnover, and Ca(2+) mobilization were fully mimicked by the lipid fraction isolated from HDL. Analysis of this fraction with high-performance liquid chromatography (HPLC) and time-of-flight secondary ion mass spectroscopy (TOF SIMS) revealed that the PI-PLC-inducing activity is identical with two bioactive lysosphingolipids, namely, lysosulfatide (LSF) and sphingosylphosphorylcholine (SPC). Like native HDL, LSF and SPC induced PtdInsP(2) turnover, Ca(2+) mobilization, and fibroblast proliferation. However, both compounds did not promote cholesterol efflux. In conclusion, two agonist activities are carried by HDL. Apo A-I stimulates phosphatidylcholine breakdown and thereby facilitates cholesterol efflux, whereas LSF and SPC trigger PI-PLC activation and thereby stimulate cell proliferation. PMID- 11106500 TI - Effects of guanidine hydrochloride on the proton inventory of proteins: implications on interpretations of protein stability. AB - The DeltaG degrees (N)(-)(D) value obtained from extrapolation to zero denaturant concentration by the linear extrapolation method (LEM) is commonly interpreted to represent the Gibbs energy difference between native (N) and denatured (D) ensembles at the limit of zero denaturant concentration. For DeltaG degrees (N)( )(D) to be interpreted solely in terms of N and D, as is common practice, it must be shown to be independent of denaturant concentration. Because DeltaG degrees (N)(-)(D) is often observed to be dependent on the nature of the denaturant, it is necessary to determine the circumstances under which DeltaG degrees (N)(-)(D) can be interpreted as a property solely of the protein. Here, we use proton inventory, a thermodynamic property of both the native and denatured ensembles, to monitor the thermodynamic character of denaturant-dependent aspects of N and D ensembles and the N right arrow over left arrow D transition. Use of a thermodynamic rather than a spectral parameter to monitor denaturation provides insight into the manner in which denaturant affects the meaning of DeltaG degrees (N)(-)(D) and the nature of the N right arrow over left arrow D transition. Three classes of proteins are defined in terms of the thermodynamic behaviors of their N right arrow over left arrow D transition and N and D ensembles. With guanidine hydrochloride as a denaturant, the classification of protein denaturations by these procedures determines when the LEM gives readily interpretable DeltaG degrees (N)(-)(D) values with this denaturant and when it does not. PMID- 11106501 TI - Calculation of z-coordinates and orientational restraints using a metal binding tag. AB - We introduce a new simple methodology allowing the measurement of (1)H-(15)N residual dipolar couplings, dipolar shifts, and unpaired electron-amide proton distances. This method utilizes a zinc finger tag fused at either the N- or the C terminus of a protein. We have demonstrated this fusion strategy by incorporating the zinc finger of the retroviral gag protein onto the C-terminus of barnase, a ribonuclease produced by Bacillus amiloliquifaciance. We show that this tag can be substituted with cobalt and manganese. Binding of cobalt to the gag zinc finger-barnase fusion protein introduced sufficient anisotropic paramagnetic susceptibility for orientation of the molecule in the magnetic field. Partial alignment permitted measurement of (1)J(HN) scalar couplings along with dipolar couplings. Replacement of bound cobalt with diamagnetic zinc removes the paramagnetic-induced orientation of barnase, permitting the measurement of only (1)J(HN) scalar couplings. Dipolar couplings, ranging from -0.9 to 0.6 Hz, were easily measured from the difference in splitting frequencies in the presence of cobalt and zinc. The observed paramagnetic anisotropy induced by cobalt binding to the metal binding tag also permitted measurement of dipolar shifts. Substitution of manganese into the metal binding tag permitted the measurement of unpaired electron-amide proton distances using paramagnetic relaxation enhancement methodology. The availability of both amide proton dipolar shifts and unpaired electron to amide proton distances permitted the direct calculation of z coordinates for individual amide protons. This approach is robust and will prove powerful for global fold determination of proteins identified in genome initiatives. PMID- 11106502 TI - Light-induced conformational changes of rhodopsin probed by fluorescent alexa594 immobilized on the cytoplasmic surface. AB - A novel fluorescence method has been developed for detecting the light-induced conformational changes of rhodopsin and for monitoring the interaction between photolyzed rhodopsin and G-protein or arrestin. Rhodopsin in native membranes was selectively modified with fluorescent Alexa594-maleimide at the Cys(316) position, with a large excess of the reagent Cys(140) that was also derivatized. Modification with Alexa594 allowed the monitoring of fluorescence changes at a red excitation light wavelength of 605 nm, thus avoiding significant rhodopsin bleaching. Upon absorption of a photon by rhodopsin, the fluorescence intensity increased as much as 20% at acidic pH with an apparent pK(a) of approximately 6.8 at 4 degrees C, and was sensitive to the presence of hydroxylamine. These findings indicated that the increase in fluorescence is specific for metarhodopsin II. In the presence of transducin, a significant increase in fluorescence was observed. This increase of fluorescence emission intensity was reduced by addition of GTP, in agreement with the fact that transducin enhances the formation of metarhodopsin II. Under conditions that favored the formation of a metarhodopsin II-Alexa594 complex, transducin slightly decreased the fluorescence. In the presence of arrestin, under conditions that favored the formation of metarhodopsin I or II, a phosphorylated, photolyzed rhodopsin Alexa594 complex only slightly decreased the fluorescence intensity, suggesting that the cytoplasmic surface structure of metarhodopsin II is different in the complex with arrestin and transducin. These results demonstrate the application of Alexa594-modified rhodopsin (Alexa594-rhodopsin) to continuously monitor the conformational changes in rhodopsin during light-induced transformations and its interactions with other proteins. PMID- 11106503 TI - Mechanisms of inactivation of human S-adenosylhomocysteine hydrolase by 5',5',6',6'-tetradehydro-6'-deoxy-6'-halohomoadenosines. AB - In an effort to design more specific and potent inhibitors of S adenosylhomocysteine (AdoHcy) hydrolase, we investigated the mechanisms by which 5',5',6', 6'-tetradehydro-6'-deoxy-6'-halohomoadenosines (X = Cl, Br, I) inactivated this enzyme. The 6'-chloro (a) and 6'-bromo (b) acetylenic nucleoside analogues produced partial ( approximately 50%) loss of enzyme activity with a concomitant ( approximately 50%) reduction of E-NAD(+) to E-NADH. In addition, Ade and halide ions were released from the inhibitors in amounts suggestive of a process involving enzyme catalysis. AdoHcy hydrolase, which was inactivated with compound a, was shown to contain 2 mol of the inhibitor covalently bound to Lys318 of two subunits of the homotetramer. These data suggest that the enzyme mediated water addition at the 5' position of compound a or b produces an alpha halomethyl ketone intermediate, which is then attacked by a proximal nucleophile (i.e., Lys318) to form the enzyme-inhibitor covalent adduct (lethal event); in a parallel pathway (nonlethal event), addition of water at the 6' position produces an acyl halide, which is released into solution and chemically degrades into Ade, halide ion, and sugar-derived products. In contrast, compound c completely inactivated AdoHcy hydrolase by converting 2 equiv of E-NAD(+) to E-NADH and causing the release of 2 equiv of E-NAD(+) into solution. Four moles of the inhibitor was shown to be tightly bound to the tetrameric enzyme. These data suggest that compound c inactivates AdoHcy hydrolase by a mechanism similar to the acetylenic analogue of Ado described previously by Parry et al. [(1991) Biochemistry 30, 9988-9997]. PMID- 11106504 TI - Aminotransferase activity and bioinformatic analysis of 1-aminocyclopropane-1 carboxylate synthase. AB - The mechanistic fate of pyridoxal phosphate (PLP)-dependent enzymes diverges after the quinonoid intermediate. 1-Aminocyclopropane-1-carboxylate (ACC) synthase, a member of the alpha family of PLP-dependent enzymes, is optimized to direct electrons from the quinonoid intermediate to the gamma-carbon of its substrate, S-adenosyl-L-methionine (SAM), to yield ACC and 5' methylthioadenosine. The data presented show that this quinonoid may also accept a proton at C(4)' of the cofactor to yield alpha-keto acids and the pyridoxamine phosphate (PMP) form of the enzyme when other amino acids are presented as alternative substrates. Addition of excess pyruvate converts the PMP form of the enzyme back to the PLP form. C(alpha)-deprotonation from L-Ala is shown by NMR monitored solvent exchange to be reversible with a rate that is less than 25-fold slower than that of deprotonation of SAM. The rate-determining step for transamination follows the formation of the quinonoid intermediate. The rate determining step for alpha, gamma-elimination from enzyme-bound SAM is likewise shown to occur after C(alpha)-deprotonation, and the quinonoid intermediate accumulates during this reaction. BLAST searches, sequence alignments, and structural comparisons indicate that ACC synthases are evolutionarily related to the aminotransferases. In agreement with previously published reports, an absence of homology was found between the alpha and beta families of the PLP-dependent enzyme superfamily. PMID- 11106505 TI - Thermooptic effect in chloroplast thylakoid membranes. Thermal and light stability of pigment arrays with different levels of structural complexity. AB - In chloroplast thylakoid membranes, chiral macrodomains, i.e., large arrays of pigment molecules with long-range chiral order, have earlier been shown to undergo light-induced reversible and irreversible structural changes; such reorganizations did not affect the short-range, excitonic pigment-pigment interactions. These structural changes and similar changes in lamellar aggregates of the main chlorophyll a/b light-harvesting complexes exhibited a linear dependence on the intensity of light that was not utilized in photosynthesis. It has been hypothesized that the light-induced rearrangements are driven by a thermooptic effect, i.e., thermal fluctuations due to the dissipation of excess excitation energies [Barzda, V., et al. (1996) Biochemistry 35, 8981-8985]. To test this hypothesis, we have utilized circular dichroism (CD) spectroscopy to investigate the structural stability of the chiral macrodomains and the constituent bulk pigment-protein complexes of granal thylakoid membranes against heat and prolonged, intense illumination. (i) In intact thylakoid membranes, the chiral macrodomains displayed high stability below 40 degrees C, but they were gradually disassembled between 50 and 60 degrees C; the thermal stability of the chiral macrodomains could be decreased substantially by suspending the membranes in reaction media that were hypotonic or had low ionic strength. (ii) The chiral macrodomains were also susceptible to high light: prolonged illumination with intense white light (25 min, 2500 microE m(-)(2) s(-)(1), 25 degrees C) induced similar, irreversible disassembly to that observed at high temperatures; in different preparations, lower thermal stability was coupled to lower light stability. (iii) The light stability depended significantly on the temperature: between about 5 and 15 degrees C, the macrodomains in the intact thylakoids were virtually not susceptible to high light; in contrast, the same preillumination at 35-40 degrees C almost completely destroyed the chiral macrodomains. (iv) As testified by the excitonic CD bands, the molecular organization of the pigment protein complexes in all samples exhibited very high thermal stability between about 15 and 65 degrees C, and virtually total immunity against intense illumination. These data are fully consistent with the hypothesis of a thermooptic effect, and are interpreted within the frame of a simple model. PMID- 11106506 TI - Structure-activity relations in the oxidation of phenethylamine analogues by recombinant human liver monoamine oxidase A. AB - The interaction of recombinant human liver monoamine oxidase A (MAO A) with a series of phenethylamine substrate analogues has been investigated by steady state and stopped-flow kinetic techniques. Substrate analogues with para substituents exhibit large deuterium kinetic isotope effect on k(cat), on k(cat)/K(m), and on the limiting rate of enzyme reduction in reductive half reaction experiments. These kinetic isotope effect values range from 5 to 10 with the exception of tyramine, which exhibited smaller steady-state isotope effects (2.3-3.5) than that observed on the rate of flavin reduction (6.9). The stopped flow data show that imine release from the reduced enzyme is slower than the rate of catalytic turnover. Phenethylamine oxidation by MAO A can be described as the C-H bond cleavage step being rate limiting in catalysis and with oxygen reacting with the reduced enzyme-imine complex. In the case of tyramine, the product release from the oxidized enzyme-imine complex contributes to the rate limitation in catalysis. The binding affinities of a series of para-substituted phenethylamine analogues to MAO A show an increase in affinity of the deprotonated amine with increasing van der Waals volume of the substituent. The limiting rate of enzyme reduction decreases with increasing van der Waals volume of the substituent in a linear manner with no observable electronic contribution as observed previously with benzylamine reduction of MAO A [Miller, J. R., and Edmondson, D. E. (1999) Biochemistry 38, 13670-13683]. Examination of side chain analogues of phenethylamine show 3-phenylpropylamine to be oxidized 2.5-fold more slowly and bound 75-fold more tightly than phenethylamine. 4-Phenylbutylamine is not a substrate for MAO A but is a good competitive inhibitor with a K(i) value of 31 +/- 5 microM. Analysis of the effect of alkyl side chain alterations on binding affinities of a series of arylalkylamine analogues taken from this study and from the literature show a linear correlation with the Taft steric value (E(s)) of the side chain. These results suggest that the binding site for the aryl ring is identical for phenethylamine and for benzylamine analogues and that steric interactions of the alkyl side chain with the enzyme strongly contribute to the binding affinities of a series of reversible inhibitors of MAO A. PMID- 11106507 TI - Involvement of phylogenetically conserved acidic amino acid residues in catalysis by an oxidative DNA damage enzyme formamidopyrimidine glycosylase. AB - Formamidopyrimidine glycosylase (Fpg) is an important bacterial base excision repair enzyme, which initiates removal of damaged purines such as the highly mutagenic 8-oxoguanine. Similar to other glycosylase/AP lyases, catalysis by Fpg is known to proceed by a nucleophilic attack by an amino group (the secondary amine of its N-terminal proline) on C1' of the deoxyribose sugar at a damaged base, which results in the departure of the base from the DNA and removal of the sugar ring by beta/delta-elimination. However, in contrast to other enzymes in this class, in which acidic amino acids have been shown to be essential for glycosyl and phosphodiester bond scission, the catalytically essential acidic residues have not been documented for Fpg. Multiple sequence alignments of conserved acidic residues in all known bacterial Fpg-like proteins revealed six conserved glutamic and aspartic acid residues. Site-directed mutagenesis was used to change glutamic and aspartic acid residues to glutamines and asparagines, respectively. While the Asp to Asn mutants had no effect on the incision activity on 8-oxoguanine-containing DNA, several of the substitutions at glutamates reduced Fpg activity on the 8-oxoguanosine DNA, with the E3Q and E174Q mutants being essentially devoid of activity. The AP lyase activity of all of the glutamic acid mutants was slightly reduced as compared to the wild-type enzyme. Sodium borohydride trapping of wild-type Fpg and its E3Q and E174Q mutants on 8 oxoguanosine or AP site containing DNA correlated with the relative activity of the mutants on either of these substrates. PMID- 11106508 TI - Evidence for a four-strand exchange catalyzed by the RecA protein. AB - Strand exchange between two duplexes is usually initiated as a three-strand event that requires the presence of a single-stranded overhang or gap in one of the two molecules. Here we show that the RecA protein can catalyze a four-strand exchange. Specifically, it can recombine short hairpin substrates with homologous stems provided that one of the hairpins possesses a chimeric DNA/RNA backbone. This four-strand exchange reaction goes to completion in the presence of ATPgammaS and releases a stable heteroduplex upon removal of the RecA protein. Under identical conditions, strand exchange between two DNA hairpins is incomplete and generates a nascent heteroduplex that rapidly dissociates when the RecA protein is denatured. Since presynaptic filament formation does not appear to melt either type of hairpin, we propose that exchange occurs between homologously aligned duplexes that are extended and unwound within a RecA filament. The first reaction provides a mechanism for gene targeting by chimeric double-hairpin oligonucleotides while the second reaction explains the ability of the RecA protein to transiently align double-stranded DNA molecules. PMID- 11106509 TI - Protein kinase C regulation of intracellular and cell surface amyloid precursor protein (APP) cleavage in CHO695 cells. AB - Cleavage of amyloid precursor protein (APP) by beta-secretase generates beta amyloid (Abeta), the major component of senile plaques in Alzheimer's disease. Cleavage of APP by alpha-secretase prevents Abeta formation, producing nonamyloidogenic secreted APPs products. PKC-regulated APP alpha-secretase cleavage has been shown to involve tumor necrosis factor alpha (TNF-alpha) converting enzyme (TACE). To determine the location of APP cleavage, we examined PKC-regulated APPs secretion by examining cell surface versus intracellular APP in CHO cells stably expressing APP(695) (CHO695). We demonstrate that PKC regulates cell surface and intracellular APP cleavage. The majority of secreted APPs originates from the intracellular compartment, and PKC does not cause an increase in APP trafficking to the cell surface for cleavage. Therefore, intracellular APP regulated by PKC must be cleaved at an intracellular site. Experiments utilizing Brefeldin A suggest APP cleavage occurs at the Golgi or late in the secretory pathway. Experiments using TAPI, an inhibitor of TACE, demonstrate PKC-regulated APPs secretion from the cell surface is inhibited after pretreatment with TAPI, and APPs secretion from the intracellular pool is partially inhibited after pretreatment with TAPI. These findings suggest PKC regulated APP cleavage occurs at multiple locations within the cell and both events appear to involve TACE. PMID- 11106510 TI - PROFILE. PMID- 11106511 TI - PRESENTATION OF CHURCHILL LIVINGSTONE MEDAL BEST CANDIDATE FESSH DIPLOMA EXAMINATION 1999. PMID- 11106512 TI - Should anatomic reduction be pursued in distal radial fractures? AB - Two articles on distal radial fractures in young adults are published in this issue of the journal. One reports the results of open reduction and internal fixation with the pi -plate in a group of patients in which the majority of fractures were complex and intraarticular. The other retrospectively assessed 169 fractures with an average follow-up of 4.9 years, based on the radiographic evaluation of the patient's X-rays until discharge and on the answers provided by a patient-based subjective outcome questionnaire. My task has been to comment, criticize and analyse the findings and results reported in both articles, in order to highlight areas of uncertainty and controversy regarding the current management of fractures of the distal radius in the younger age group. PMID- 11106513 TI - Open reduction and internal fixation of intra articular and unstable fractures of the distal radius using the AO distal radius plate. AB - This study reports the results of open reduction and internal fixation of 25 dorsally displaced distal radial fractures using a specifically designed plate for the distal radius, the AO pi plate (Synthes Ltd, Paoli, USA). Twenty-one of these fractures were complex and intra-articular (AO Type 'C'). Measurement of range of motion of the affected wrist at an average follow-up of 16 months revealed a median return of 60 degrees of wrist extension, 40 degrees of wrist flexion, 90 degrees of pronation and 90 degrees of supination. Radiographic assessment revealed restoration of normal radial length, inclination and palmar tilt in all but six cases. The final outcome, as assessed by the Gartland and Werley scale, was excellent in four cases, good in 11, and fair in ten cases. Complications were seen in five patients. PMID- 11106514 TI - Outcome of distal radial fractures in young adults. AB - The outcome of 169 fractures of the distal radius in adults under the age of 50 were assessed at least 18 months after injury (mean follow-up, 4.9 years) using a validated, patient-based outcome questionnaire. The questionnaire responses demonstrated that neither the Frykman nor the Mayo classifications of distal radial fractures predicted outcome. Fracture union with more than 10 degrees of dorsal tilt was associated with increased difficulty with everyday activities and work, while union with a step in the radiocarpal articular surface was associated with loss of wrist mobility and difficulty with fine dextrous tasks. No measure of either intra- or extra-articular malunion influenced the severity or frequency of persistent wrist pain. PMID- 11106515 TI - T incision for exposure of the distal radius and wrist. AB - A retrospective review of 66 T-shaped incisions for exposure of the dorsal distal radius and wrist was performed. The incision provided excellent exposure in all cases and no additional incisions were required. Cosmesis was considered acceptable by all patients. Complications occurred in 6% and were more likely in patients undergoing fixation of acute distal radius fractures using Kirschner wires which protruded through the skin flaps. PMID- 11106516 TI - The mechanical properties of locking and grasping suture loop configurations in four-strand core suture techniques. AB - We have compared the effect of locking and grasping suture loop configurations in four-strand core suture techniques for tendon repair. Forty canine flexor digitorum profundus tendons were repaired with one of four suture techniques (the grasping cruciate, the double-modified grasping Kessler, the locking cruciate and the double-modified locking Kessler) and tested to failure in a tensile testing machine. The mode of failure in all the locking suture specimens was breakage of the sutures in the locking loops or at suture knots. The sutures did not pull out of the tendon, as was seen in the grasping suture specimens. The greatest tensile strength was found with the double-modified locking Kessler technique which incorporated eight rectangular locking loop configurations. PMID- 11106517 TI - Flexor tendon blood vessels. AB - The aim of this study was to assess rabbit long flexor tendon vascularity in a qualitative and quantitative manner using immunohistochemistry. The endothelial cell surface marker CD31 was targeted with a specific monoclonal mouse-anti-human antibody with good species cross-reactivity. Subsequent signal amplification and chromogen labelling allowed vessel visualization. Computer image analysis was performed. Values for vessel number and total vessel area per section, as well as the sections' cross-sectional tendon areas, were obtained. There was a consistent deep tendon avascular zone between the A2 and A4 pulley in the rabbit forepaw. This was not the case in the hindpaw, with dorsally orientated longitudinal vessels coursing the length of the intrasynovial tendon. The area of least vascularity in the hindpaw was around the metacarpophalangeal joint. We therefore recommend the use of hindpaw tendons when using the rabbit as a flexor tendon experimental model. This is because its vascular pattern is similar to that of the human flexor digitorum profundus. PMID- 11106518 TI - Strength of the intrinsic muscles of the hand measured with a hand-held dynamometer: reliability in patients with ulnar and median nerve paralysis. AB - The aim of this study was to assess the reliability of a technique to measure the strength of the intrinsic hand muscles. Intraclass Correlation Coefficients showed an excellent level of reliability for the comparison of muscle strength between groups of patients. However, for the results of individual patients, the calculated Standard Error of Measurements (10-16%) and the Smallest Detectable Differences for intraobserver (31-36%) and interobserver (37-52%) values indicate that only relatively large changes in strength can be confidently detected with this technique. The results of the present study were compared with those of four previous grip strength studies. PMID- 11106519 TI - Ganglia: the patient's perception. AB - This study investigates the concerns of 50 patients with ganglia and their reasons for primary care consultation and referral to a hand unit. Although a minority of patients sought advice and treatment because of pain, more (38%) were concerned about the cosmetic appearance and a significant number (28%) were concerned that their ganglion was a malignant growth. The general practitioners referred 70% of patients to the hand clinic for "excision of the ganglion" and 30% for further "advice and treatment". However, 74% of patients were satisfied with aspiration of the ganglion and general advice. PMID- 11106520 TI - The Compass Elbow Hinge: indications and initial results. AB - The Compass Elbow Hinge uses Illizarov's methods of fixation to externally hold the elbow reduced and allow both passive and active motion. Eleven patients with degenerative disease, contracture or instability were treated with the Compass Elbow Hinge and were retrospectively evaluated at an average follow-up of 29 months (range: 18-62 months). One was lost to follow-up. Patients with degenerative changes underwent fascia lata interposition while those treated for contractures underwent anterior and posterior capsular release with or without fascia lata interposition. Those with elbow instability underwent ligament reconstruction. The device was removed after 6 weeks and seven of the 11 patients were satisfied with the outcome of the operation. Stability could not be achieved in two patients with coronoid fractures that were not reconstructed. One patient did not tolerate the device and requested its removal with subsequent subluxation. We conclude that patient selection and compliance are key elements in achieving a satisfactory outcome with the device. PMID- 11106521 TI - Efficacy of forearm versus upper arm tourniquet for local anaesthetic surgery of the hand. AB - A prospective study was undertaken to compare the use of forearm and upper arm tourniquets for local anaesthetic procedures on the hand. One hundred consecutive patients with an upper arm tourniquet were compared with a further consecutive 100 patients in whom a forearm tourniquet was used. The tourniquet time was always less than 20 minutes and the scoring of perceived pain was not significantly different in the two groups. Use of a forearm tourniquet was well tolerated and was not associated with an increase in complications. PMID- 11106522 TI - A modification of the technique for intravenous regional blockade for hand surgery. AB - A prospective study was conducted to assess a modification to Bier's intravenous regional anaesthesia which introduced a third temporary distal forearm tourniquet. This confines the injected lignocaine to the hand, resulting in a higher local lignocaine concentration. Subsequent exsanguination of the limb then channels the remaining intravascular lignocaine under the distal cuff of a double tourniquet. Of the 18 patients, none experienced pain during operation and all tolerated the tourniquet without significant discomfort. Mild postoperative giddiness was noted in one patient. No other anaesthetic complications were encountered. In a subjective assessment of the bloodlessness of the operating field, two were ranked satisfactory, ten good and six excellent. None of the patients required re-exsanguination when using this technique. PMID- 11106523 TI - Rhys-Davies exsanguinator: pressure characteristics and technique for improving performance. AB - This paper reports the results of an investigation into the pressure exerted by the Rhys-Davies exsanguinator on the palm and dorsum of the hand. We hypothesised that, due to the shape of the hand, the palm is shielded from the full effect of the exsanguinator, but our study showed that it is compressed as well as the dorsum. However, placing a bag of fluid in the patient's palm significantly increased the pressure applied to the palm. PMID- 11106524 TI - Adhesion formation after nerve repair: an experimental study of early protected mobilization in the rabbit. AB - The common peroneal nerve and its surrounding muscles were cut and repaired in 14 rabbits. The injured limb was then either immobilized for 3 weeks or passively mobilized within a "safety range" every day. At 3 weeks after operation, the "stretch test" and "peel test" showed no difference in the biomechanical features of the adhesions between the nerve repair and the surrounding soft tissues. PMID- 11106525 TI - Peripheral nerve motion measurement with spectral Doppler sonography: a reliability study. AB - This study evaluates single operator test-retest reliability of spectral Doppler ultrasound measurement of median nerve excursion during wrist extension. Longitudinal motion of the median nerve was measured at the elbow on three occasions in both upper limbs of 16 healthy subjects using a standard colour Doppler ultrasound system. The mean of the three maximum velocity time integrals was calculated from the spectral Doppler sonogram of each test. Analysis of data with intraclass correlation coefficient indicated a high degree of repeatability (0.92). Spectral Doppler ultrasound may provide a valuable method for measurement of peripheral nerve motion and may have a role in the clinical assessment of entrapment syndromes. PMID- 11106526 TI - The effectiveness of ADCON-T/N, a new anti-adhesion barrier gel, in fresh divisions of the flexor tendons in Zone II. AB - In a prospective randomized clinical trial, ADCON-T/N was investigated with regard to its effectiveness in fresh traumatic injuries of the flexor tendons in Zone II of the hand. Thirty patients participated in the trial. Following a standardized technique of tendon repair, the total active motion (TAM) and total extension lag (TEL) were determined after 12 weeks and evaluated according to the Buck-Gramcko score. Excellent results were achieved in 15 out of 16 patients in the ADCON-T/N group and 12 out of 14 in the control group. However, no statistically significant difference was found between the mean TAM and TEL in the two groups. PMID- 11106527 TI - Validity and reliability of three generic outcome measures for hand disorders. AB - This is an assessment of three different outcome measures for the hand: the Hand Clinic Questionnaire (HCQ); the Patient Evaluation Measure (PEM); and the Hand Outcome Survey Sheet (HOSS). Each measure has been tested for its reliability and validity. The results suggest that the PEM and the HCQ are comparably consistent but the PEM is more reproducible. Both the PEM and HOSS are valid questionnaires. The PEM is suitable for use in an outpatient clinic and as a postal questionnaire. The HOSS may be used for research or audit especially when the injury has been measured using the Hand Injury Severity Score. PMID- 11106528 TI - Outcome measures following metacarpophalangeal joint replacement. AB - We used the Jebsen-Taylor hand function assessment system to prospectively study the effect of metacarpophalangeal joint replacement on overall hand function in 29 hands. In addition pain relief, subjective improvement in hand function, appearance and overall patient satisfaction were assessed. There was modest improvement in the number of Jebsen-Taylor tasks performed (1.8 to 3. 1), and pain relief was good or excellent in 18 of 24 patients. Eleven patients felt their hand function had improved by more than 50%, and the majority of patients (22 of 24) were very satisfied with the procedure. This study demonstrates that despite limited improvements in objective outcome measures, this procedure is reliable in producing a very high rate of patient satisfaction. PMID- 11106529 TI - Schwannomas of the upper extremity. AB - This study presented the clinical characteristics, MRI features and postoperative results of 20 schwannomas in the arms of 13 patients. Twelve tumours had a positive Tinel's sign, one caused weakness of the wrist and another in Guyon's canal caused hypothenar muscle atrophy. Of the nine cases which underwent magnetic resonance imaging preoperatively, six were correctly diagnosed as schwannomas. All masses were excised using microsurgical techniques and two transient neurological complications occurred. PMID- 11106530 TI - The first toe-to-hand transfer: a thirty-year follow-up. AB - In April 1968 the first successful free toe-to-hand transfer was performed by Mr. J. R. Cobbett at the Queen Victoria Hospital to reconstruct the thumb of a woodworker. Details of the case are discussed and the patient's current level of sensation, power and function are presented over 30 years following reconstruction. PMID- 11106532 TI - CONTINUING MEDICAL EDUCATION. PMID- 11106531 TI - Digital arterial occlusion in scleroderma: is there a role for digital arterial reconstruction? AB - A 47-year-old patient with Raynaud's phenomenon secondary to scleroderma developed long finger ischaemic pain. This was successfully treated with a palmar sympathectomy and a long finger digital artery reconstruction using a reversed small calibre vein graft between a perforator from the deep palmar arch and the radial digital artery at the level of the distal interphalangeal joint. PMID- 11106534 TI - Letters to the Editor. PMID- 11106533 TI - Re: Horton, TC (2000). Isolated paralysis of the extensor pollicis longus muscle: a further variation of the posterior interosseous nerve palsy. Journal of Hand Surgery, 25B 2, 225-6. PMID- 11106535 TI - Epstein-Barr virus (EBV) load and cytokine gene expression in activated T cells of chronic active EBV infection. AB - To identify the role of T cells in chronic active Epstein-Barr virus (EBV) infection, EBV and cytokine gene expression was quantified by use of real-time polymerase chain reaction (PCR) among 6 patients who fulfilled the diagnostic criteria for chronic active EBV infection. Four of these patients showed clonal expansion of EBV-infected T cells. Quantitative PCR for EBV DNA in peripheral blood of patients with symptomatic chronic active EBV infection showed higher copy numbers of virus (mean, 1.45 x 10(5) copies/mL) than were seen in blood from patients with infectious mononucleosis (3.08 x 10(3) copies/mL) or with EBV associated hemophagocytosis (2.95 x 10(4) copies/mL). Fractionated CD3(+) HLA DR(+) cells from patients with chronic active EBV infection contained higher copy numbers than did CD3(+) HLA-DR(-) cells. Quantitative PCR for cytokines revealed that interferon-gamma, interleukin (IL)-2, IL-10, and transforming growth factor beta genes were expressed at higher levels in HLA-DR(+) than in HLA-DR(-) T cells. These results suggest that activated T cells in chronic active EBV infection expressed high levels of EBV DNA and both Th1 and Th2 cytokines. EBV infected T cells may contribute to the unbalanced cytokine profiles of chronic mononucleosis. PMID- 11106536 TI - Early human immunodeficiency virus (HIV) infection in the HIV Network for Prevention Trials Vaccine Preparedness Cohort: risk behaviors, symptoms, and early plasma and genital tract virus load. AB - Risk behaviors, symptoms, and virologic characteristics were studied among 103 human immunodeficiency virus (HIV) seroconverters in vaccine preparedness cohorts during 1995-1998. Overall, 83% of subjects were men who had sex with men; most reported multiple risk episodes and symptoms (84%, > or =1 symptom) during seroconversion. Acute HIV was diagnosed in only 8 of 50 who sought medical care. Median initial pretreatment plasma virus load was 25,800 copies/mL (range, undetectable-262,000 copies/mL) a mean of 4 months after seroconversion, and 9.7% had nucleoside-associated mutations; none had multidrug resistance. Semen virus load was more variable, 1.3 log(10) lower and modestly correlated (r=.28; 95% confidence interval, 0.16-0.42) with plasma among untreated men. When the plasma RNA level was <5000 copies/mL, 32% of untreated men, 13% on nucleoside regimens, and 7% on protease inhibitor-containing regimens had detectable seminal RNA. Acute HIV was seldom diagnosed, representing missed opportunities for early treatment and prevention. Most subjects had several relatively stable virus loads before initiation of antiretrovirals, indicating feasibility of assessing HIV vaccines on virus set point in efficacy trials. PMID- 11106537 TI - Human immunodeficiency virus type 1 quasi species that rebound after discontinuation of highly active antiretroviral therapy are similar to the viral quasi species present before initiation of therapy. AB - In an effort to identify the sources of the viruses that emerge after discontinuation of therapy, analyses of human immunodeficiency virus (HIV) quasi species were done for 3 patients with sustained levels of HIV RNA of <50 copies/mL for 1-3 years. The sequences found in the rebounding plasma virus were closely related to those of the actively replicating form of viruses present before the initiation of combination therapy. All quasi species found in the rebounding plasma virus were also present in proviral DNA, cell-associated RNA in peripheral blood mononuclear cells (PBMC), and virion RNA derived from PBMC coculture during periods when plasma HIV RNA levels were <50 copies/mL. These findings suggest that the rapid resurgence of plasma viremia observed after discontinuation of therapy and the viruses cocultured from PBMC are derived from a relatively stable pool of the replicating form of virus rather than from activation of a previously latent pool. PMID- 11106538 TI - Phylogenetic distribution of extraintestinal virulence-associated traits in Escherichia coli. AB - The 72 member strains of the Escherichia coli Reference collection were assessed as to genotype for 31 putative extraintestinal virulence factor (VF) genes and DNA sequence for papA, the P fimbrial structural subunit gene. Although most VFs were concentrated in phylogenetic group B2 or jointly in groups B2 and D, others were concentrated primarily in group D, were broadly distributed (without group specific associations), and/or occurred only outside of group B2. Statistical correlations among VFs suggested linkage on pathogenicity-associated islands or plasmids. Isolates from humans and nonhuman primates had more VFs than did isolates from other animals. Sequence diversity was minimal within each F type specific papA allele group but was substantial among different papA allele groups. The distribution patterns of papA variants and other VFs suggested multiple horizontal transfer events. These findings provide new insights into the phylogenetic origins of extraintestinal VFs in E. coli. PMID- 11106539 TI - Distinct distribution of rare US genotypes of Kaposi's sarcoma-associated herpesvirus (KSHV) in South Texas: implications for KSHV epidemiology. AB - Genotypes of Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) from patients with KS in South Texas were examined. Open-reading frame (ORF)-K1 and ORF-K15 DNA segments from 16 KSHV isolates were amplified by polymerase chain reaction, and KSHV subtypes were assigned on the basis of sequence variations. K1 genotyping showed that 75% exhibited C subtype and 25% exhibited A subtype. K15 genotyping showed that 56% exhibited M form, of which 89% exhibited C3 K1 subtype and 44% exhibited P form. A unique isolate was found and was classified as C6 clade. All of the M KSHV isolates had been obtained from human immunodeficiency virus negative classic KS patients >50 years of age, of whom 78% were Hispanic. Conversely, all KS patients with AIDS were <36 years of age and exhibited P form KSHV. These findings indicate that C3/M KSHV genotypes are more prevalent in South Texas (50%) than in other US regions (3%) and that M form KSHV likely existed in this region long before the AIDS epidemic. PMID- 11106540 TI - Identification of non-B human immunodeficiency virus type 1 subtypes in rural Georgia. AB - As part of an ongoing molecular epidemiological investigation of human immunodeficiency virus type 1 (HIV-1) in rural Georgia, the 5' half of reverse transcriptase (RT) genotypes from 30 patients was sequenced and phylogenetically analyzed. Two patients, GA132 and GA169, were infected with pol sequences of non B subtype origin that were found to cluster phylogenetically with subtype A-E of Thai origin. Sliding window bootstrap analysis of GA169 showed clear evidence of A/B recombination within the pol gene segment, whereas in the other patient, GA132, no break point within RT could be identified. Interestingly, pairwise comparisons between these 2 patients' C2-V3 env region revealed a 13.5% divergence. However, similar comparisons within the non-B pol segments yielded a 1.23% nucleotide divergence, which suggests a complex phylogenetic and epidemiological history of the subtype A pol genotype in this region. These data demonstrate an increasing diversity of HIV-1 subtypes and the potential emergence of previously unidentified HIV-1 A-E/B recombinants in the rural United States. PMID- 11106541 TI - Vascular endothelial growth factor in bacterial meningitis: detection in cerebrospinal fluid and localization in postmortem brain. AB - Vascular endothelial growth factor (VEGF) is a potent vascular permeability factor and a mediator of brain edema. To assess the role of VEGF during bacterial meningitis, VEGF was measured in cerebrospinal fluid (CSF) and blood of 37 patients with bacterial meningitis and 51 control patients, including 16 patients with viral meningitis. Circulating VEGF levels were similar in bacterial meningitis patients and control patients. VEGF(CSF) was detected in 11 (30%) of 37 of bacterial meningitis patients (range, <25-633 pg/mL) but in none of the control patients. The median VEGF index was 6.2 (range, 0.6-42), indicating intrathecal production. Median CSF cell counts, protein levels, and CSF: serum albumin ratios were higher for patients with detectable VEGF(CSF), although the difference was not statistically significant. VEGF immunoreactivity in autopsy brain specimens was found in the inflammatory infiltrate of patients with bacterial meningitis. These results indicate that inflammatory cells secrete VEGF during bacterial meningitis and that VEGF may contribute to blood-brain barrier disruption. PMID- 11106542 TI - Molecular comparison of extraintestinal Escherichia coli isolates of the same electrophoretic lineages from humans and domestic animals. AB - Molecular typing methods were used to characterize 38 Escherichia coli strains that originally were isolated from extraintestinal infections and represented 5 multilocus enzyme electrophoretic types (ETs) recovered from both humans and animals. Within each ET, the human and animal isolates did not consistently segregate by host group, according to individual virulence factors (VFs), composite VF-serotype profiles, or pulsed-field gel electrophoresis profiles. Several close matches with respect to VF-serotype profiles were identified between human and canine isolates from different locales. One canine and 2 human isolates of serogroup O6 closely resembled archetypal human pyelonephritis isolate 536 (O6:K15:H31), according to papA sequence and VF-serotype profile. These findings support the hypothesis that certain pathogenic lineages of E. coli cause disease in both humans and animals and that humans may acquire pathogenic E. coli from domestic pets. PMID- 11106543 TI - Use of tissue culture-amplified human immunodeficiency virus type 1 to study evolutionary changes in vivo. PMID- 11106546 TI - Editor's farewell PMID- 11106545 TI - Use of ESAT-6 and CFP-10 antigens for diagnosis of extrapulmonary tuberculosis. PMID- 11106548 TI - p63 is a prostate basal cell marker and is required for prostate development. AB - The p53 homologue p63 encodes for different isotypes able to either transactivate p53 reporter genes (TAp63) or act as p53-dominant-negatives (DeltaNp63). p63 is expressed in the basal cells of many epithelial organs and its germline inactivation in the mouse results in agenesis of organs such as skin appendages and the breast. Here, we show that prostate basal cells, but not secretory or neuroendocrine cells, express p63. In addition, prostate basal cells in culture predominantly express the DeltaNp63alpha isotype. In contrast, p63 protein is not detected in human prostate adenocarcinomas. Finally, and most importantly, p63(-/ ) mice do not develop the prostate. These results indicate that p63 is required for prostate development and support the hypothesis that basal cells represent and/or include prostate stem cells. Furthermore, our results show that p63 immunohistochemistry may be a valuable tool in the differential diagnosis of benign versus malignant prostatic lesions. PMID- 11106547 TI - Uteroplacental blood flow. The story of decidualization, menstruation, and trophoblast invasion. PMID- 11106549 TI - Matrix metalloproteinase 9 promoter activity is induced coincident with invasion during tumor progression. AB - Matrix metalloproteinase 9 (MMP-9, also known as gelatinase B or 92-kd Type IV collagenase) is overexpressed in many human and murine cancers. We induced carcinomas in mice carrying a transgene that links the MMP-9 promoter to the reporter ss-galactosidase so that activation of the MMP-9 promoter would be indicated by ss-galactosidase. Mammary carcinomas were induced by mating the MMP 9 promoter reporter transgenic mice with mice carrying a transgene for murine mammary tumor virus promoter linked to polyoma middle T antigen, a transgene that leads to rapid development of mammary tumors in female mice. None of the hyperplastic mammary glands and none of the carcinomas in situ expressed ss galactosidase. However, all invasive tumors had evidence of ss-galactosidase expression. In addition to the breast carcinomas, a malignant teratoma in a female and a papillary adenocarcinoma in the pelvic region of a male arose and were also ss-galactosidase positive. We also induced skin tumors in the mice with the MMP-9 reporter transgene with 7, 12-dimethylbenz[a]anthracene (DMBA) treatment followed by phorbol 12 myristate 13-acetate (TPA). None of the papillomas or in situ carcinomas showed any ss-galactosidase expression, but expression was seen in invasive carcinoma. Although normal skin epithelial cells did not express ss-galactosidase, we did find staining in a few cells at the duct of the sebaceous gland at the base of the hair follicles. The MMP-9 reporter transgene did not lead to expression in the alveolar macrophages, confirming that additional upstream sequences are required for expression in macrophages. These experiments have revealed that MMP-9 promoter activity is induced coincident with invasion during tumor progression. Furthermore, this indicates that the more proximal upstream elements of the promoter are sufficient for MMP-9 transcription during tumor progression. PMID- 11106550 TI - Ezrin immunoreactivity is associated with increasing malignancy of astrocytic tumors but is absent in oligodendrogliomas. AB - The actin-binding protein ezrin has been associated with motility and invasive behavior of malignant cells. To assess the presence of this protein in human glial cells of the brain and its potential role in benign and malignant glial tumors, we studied ezrin immunoreactivity (IR), proliferation (MIB-1-IR), and apoptosis (terminal dUTP nick-end labeling) in normal human brain tissues from 10 autopsies and tissues from 115 cases of human glial tumors including astro cytomas, ependymomas, oligodendrogliomas, and glioblastomas. We found weak staining of peripheral processes in normal human brain astrocytes and in World Health Organization grade II benign astrocytomas. Staining was markedly increased in anaplastic astrocytomas (World Health Organization grade III) and clearly strongest in glioblastomas (World Health Organization grade IV). The increase of ezrin-IR correlated significantly with increasing malignancy of astrocytic tumors (P < 0.0001). Statistical analysis revealed a stronger association with increasing malignancy for ezrin-IR than for MIB-1-IR or terminal dUTP nick-end labeling staining. Ezrin-IR was absent in normal oligodendrocytes and in oligodendrogliomas, but pronounced in normal ependymal cells and ependymomas. Ezrin-IR seems to be specific for astrocytes and ependymal glia in the normal brain. Our results indicate that ezrin-IR may provide a useful tool for the distinction of oligodendrogliomas and astrocytomas and for the grading of astrocytic tumors. PMID- 11106551 TI - Deregulation of the Rb and p53 pathways in uveal melanoma. AB - Uveal melanoma is the most common primary eye cancer, yet its molecular pathogenesis is poorly understood. In this study, we investigated the immunohistochemical expression of proteins in the Rb and p53 tumor suppressor pathways in 33 uveal melanomas from enucleated eyes. Strong nuclear staining for Rb was present in most tumors. However, a few cases displayed weak nuclear staining and strong cytoplasmic staining (possibly indicating Rb mutation), and this aberrant staining correlated strongly with failed radiotherapy or thermotherapy before enucleation. Staining for cyclin D1 was positive in most tumors and was associated with advanced age and larger tumor size, which are both poor prognostic factors. Generally, immunostaining for p53 was weak (suggesting a lack of p53 mutations), although p53 positivity correlated strongly with staining for phosphorylated Rb, supporting the notion that inappropriate phosphorylation of Rb can induce p53. Strong immunostaining for MDM2, which can functionally block p53 activity, was observed in most tumors and correlated significantly with female sex. Strong cytoplasmic staining was observed for Bcl2, which can inhibit both p53-dependent and -independent apoptosis. We conclude that Rb and p53 are mutated infrequently in uveal melanoma, but their respective pathways may be functionally inactivated. PMID- 11106552 TI - Consistent patterns of allelic loss in natural killer cell lymphoma. AB - Natural killer (NK) cell lymphomas are a group of rare but highly aggressive malignancies. Clinically, they can be divided into nasal NK cell lymphomas, nonnasal NK cell lymphomas, and aggressive NK cell lymphoma/leukemia. To determine the patterns of genetic deletions in these tumors, we performed loss of heterozygosity (LOH) analysis on 15 cases (11 nasal and four nonnasal), and fluorescence in situ hybridization on three cases of aggressive lymphoma/leukemia. A panel of 41 microsatellite loci on chromosomes 6q, 11q, 13q, and 17p were investigated. LOH at chromosomes 6q and 13q was frequently detected in NK cell lymphomas, being found in 80 and 66.7% of cases, respectively. LOH at chromosomes 11q and 17p was less common, being found in 28.6 and 30.8% of cases, respectively. Most tumors showed multiple loci deletions at different chromosomal regions, but several patterns of LOH could be defined. LOH at chromosome 6q was found in 90.9% of nasal NK cell lymphomas, but only in 50% of nonnasal NK cell lymphomas. LOH at chromosome 13q was found in 63.6% of nasal NK cell lymphomas and 75% of nonnasal NK cell lymphomas. For nasal NK cell lymphomas, LOH at 13q was found in 33.3% of cases at presentation, but 100% of cases at relapse. Five tumors showed LOH in only one chromosomal region, involving 6q in three cases (two nasal and one nonnasal), and 13q in two cases (both nonnasal). For the three cases of aggressive NK cell lymphoma/leukemia studied by fluorescence in situ hybridization, DNA loss at 13q14 and 17p13 regions were demonstrated. 17p13 seemed to be more commonly involved in aggressive than nasal and nonnasal NK cell lymphomas. Our results suggested that consistent patterns of LOH could be defined in NK cell malignancies. These deleted loci may contain genes important in the initiation and progression of this lymphoma. PMID- 11106553 TI - Up-regulation of CCR5 expression in the placenta is associated with human immunodeficiency virus-1 vertical transmission. AB - The role of placenta in vertical transmission is not yet fully understood. A protective role of the placenta during gestation is suggested by the finding that caesarian sections reduce the risk of transmission of human immunodeficiency virus (HIV)-1 from mother to child three- to fourfold. Here we investigated whether the immunological milieu of the placenta might be important in HIV-1 transmission. In situ imaging of immunohistochemically stained placenta sections and reverse transcriptase-polymerase chain reaction demonstrated a fourfold increase in CCR5:CXCR4 expression ratio in placentae from transmitting women compared to placentae from nontransmitting women. This chemokine receptor repertoire was consistent with an up-regulation of interleukin-4 and interleukin 10 expression in placentae from nontransmitting placentae compared to transmitting placentae. In situ imaging demonstrated that CCR5 and CXCR4 were expressed on placental macrophages and lymphocytes but not in trophoblasts. Simultaneous immunofluorescence/ultrasensitive in situ hybridization for HIV-1 gag-pol mRNA revealed that HIV-1 infects primarily CXCR4-expressing cells in placentae from nontransmitting women whereas predominantly CCR5-expressing cells were infected in placentae from transmitting women. These data are consistent with transmission of a homogeneous population of nonsyncytium-inducing HIV-1 isolates that use CCR5 as co-receptor. PMID- 11106554 TI - Exogenous interferon-gamma enhances atherosclerosis in apolipoprotein E-/- mice. AB - A role for interferon-gamma (IFN-gamma) has been implied in the atherogenic process. To determine whether exogenously administered IFN-gamma exerts an effect on the development of atherosclerosis, we intraperitoneally administered either recombinant IFN-gamma (100 U/g body weight) or phosphate buffered saline daily for 30 days to atherosclerosis-susceptible apolipoprotein E-/- mice (16-week-old male mice, n = 11 per group) fed a normal diet. Atherosclerotic lesion size was quantified in the ascending aorta. The number of T lymphocytes and major histocompatibility complex (MHC) class II-positive cells within lesions were also quantified in this region. IFN-gamma administration reduced serum cholesterol concentrations by 15% (P = 0.02). For both groups, the majority of cholesterol was present in very low density lipoproteins, which were modestly reduced in mice receiving IFN-gamma. Despite the decrease in serum cholesterol concentrations, IFN-gamma injections significantly increased lesion size twofold compared to controls (119,980 +/- 18, 536 vs. 59,396 +/- 20,017 micrometer(2); P = 0.038). IFN-gamma also significantly increased the mean number of T lymphocytes (19 +/- 4 vs. 7 +/- 1 cells; P = 0.03) and MHC class II-positive cells (10 +/- 3 vs. 3 +/- 1 cells; P = 0.04) within lesions. These data lend further support to a pro atherogenic role of IFN-gamma. PMID- 11106555 TI - Molecular evidence for multicentric development of thyroid carcinomas in patients with familial adenomatous polyposis. AB - Familial adenomatous polyposis is characterized by multiple colorectal adenomas and an increased incidence of colorectal carcinomas. Patients also develop various extracolonic tumors, of which, thyroid carcinoma is common in young females. The occurrence of multiple carcinomas in one thyroid is frequently observed, although some carcinomas are solitary. To clarify whether each carcinoma develops independently or metastatically spreads from the first one formed, we analyzed the adenomatous polyposis coli (APC) gene mutation in each carcinoma. We found that each carcinoma had a different somatic mutation of the APC gene. This is molecular confirmation for the multicentric development of thyroid carcinomas in familial adenomatous polyposis through biallelic inactivation of the APC gene. PMID- 11106556 TI - Co-localization of multiple antigens and specific DNA. A novel method using methyl methacrylate-embedded semithin serial sections and catalyzed reporter deposition. AB - Co-localization of proteins and nucleic acid sequences by in situ hybridization and immunohistochemistry is frequently difficult as the process necessary to detect the target structure of one technique may negatively affect the target of the other. Morphological impairment may also limit the application of the two techniques on sensitive tissue. To overcome these problems we developed a method to perform in situ hybridization and immunohistochemistry on semithin sections of methyl methacrylate-embedded tissue. Microwave-stimulated antigen retrieval, signal amplification by catalyzed reporter deposition, and fluorescent dyes were used for both techniques, yielding high sensitivity and excellent morphological preservation compared to conventional paraffin sections. Co-localization of in situ hybridization and immunohistochemistry signals with high morphological resolution was achieved on single sections as well as on adjacent multiple serial sections, using computerized image processing. The latter allowed for the co localization of multiple antigens and a specific DNA sequence at the same tissue level. The method was successfully applied to radiation bone marrow chimeric rats created by transplanting wild-type Lewis rat bone marrow into TK-tsa transgenic Lewis rats, in an attempt to trace and characterize TK-tsa transgenic cells. It also proved useful in the co-localization of multiple antigens in peripheral nerve biopsies. PMID- 11106557 TI - Apolipoprotein E inhibits neointimal hyperplasia after arterial injury in mice. AB - The potential cytostatic function of apolipoprotein (apo) E in vivo was explored by measuring neointimal hyperplasia in response to vascular injury in apoE deficient and apoE-overexpressing transgenic mice. Results showed a significant increase in medial thickness, medial area, and neointimal formation after vascular injury in both apoE knockout and wild-type C57BL/6 mice. Immunochemical analysis with smooth muscle alpha-actin-specific antibodies revealed that the neointima contained proliferating smooth muscle cells. Neointimal area was 3.4 fold greater, and the intima/medial ratio as well as stenotic luminal area was more pronounced in apoE(-/-) mice than those observed in control mice (P < 0.05). The human apoE3 transgenic mice in FVB/N genetic background were then used to verify a direct effect of apoE in protection against neointimal hyperplasia in response to mechanically induced vascular injury. Results showed that neointimal area was reduced threefold to fourfold in mice overexpressing the human apoE3 transgene (P < 0.05). Importantly, suppression of neointimal formation in the apoE transgenic mice also abolished the luminal stenosis observed in their nontransgenic FVB/N counterparts. These results documented a direct role of apoE in modulating vascular response to injury, suggesting that increasing apoE level may be beneficial in protection against restenosis after vascular surgery. PMID- 11106559 TI - Modulation of human colon tumor-stromal interactions by the endothelin system. AB - Tumor neovascularization is considered to be a critical step in the development of a malignant tumor. Endothelin (ET)-1 is a powerful vasoconstrictor and mitogenic peptide that is produced by many cancer cell lines. The cellular distribution of the ET components was evaluated in human colon tumors and compared to normal colon. There was more of the ET components (preproET-1, endothelin-converting enzyme-1, and ETA and ETB receptors) in adenomas and adenocarcinomas than in the normal colon. There was overproduction of preproET-1 and endothelin-converting enzyme-1 in carcinoma cells and stromal vessels, suggesting that they are a local source of ET-1. ETA receptors were present in stromal myofibroblasts of neoplastic tissue, and there were large amounts of ETB receptors in the endothelium and myofibroblasts. There was also a redistribution of alpha-smooth muscle actin-positive cells in the vascular structures of tumors. An experimental rat model of induced colon cancer treated for 30 days with bosentan, a mixed antagonist of both ET receptors, confirmed the morphological changes observed during the tumor vascularization. Our data suggest that ET-1 and its receptor play a role in colon cancer progression, with ET-1 functioning as a negative modulator of the stromal response. PMID- 11106558 TI - Inhibition of the tissue factor-thrombin pathway limits infarct size after myocardial ischemia-reperfusion injury by reducing inflammation. AB - Functional inhibition of tissue factor (TF) has been shown to improve coronary blood flow after myocardial ischemia/reperfusion (I/R) injury. TF initiates the coagulation protease cascade, resulting in the generation of the serine protease thrombin and fibrin deposition. Thrombin can also contribute to an inflammatory response by activating various cell types, including vascular endothelial cells. We used a rabbit coronary ligation model to investigate the role of TF in acute myocardial I/R injury. At-risk areas of myocardium showed increased TF expression in the sarcolemma of cardiomyocytes, which was associated with a low level of extravascular fibrin deposition. Functional inhibition of TF activity with an anti-rabbit TF monoclonal antibody administered either 15 minutes before or 30 minutes after coronary ligation reduced infarct size by 61% (P = 0.004) and 44% (P = 0.014), respectively. Similarly, we found that inhibition of thrombin with hirudin reduced infarct size by 59% (P = 0.014). In contrast, defibrinogenating the rabbits with ancrod had no effect on infarct size, suggesting that fibrin deposition does not significantly contribute to infarct size. Functional inhibition of thrombin reduced chemokine expression and inhibition of either TF or thrombin reduced leukocyte infiltration. We propose that cardiomyocyte TF initiates extravascular thrombin generation, which enhances inflammation and injury during myocardial I/R. PMID- 11106560 TI - Facilitated wound healing by activation of the Transglutaminase 1 gene. AB - Transglutaminase 1 (TGase 1) is a Ca(2+)-dependent enzyme which catalyzes epsilon (gamma-glutamyl)lysine cross-linking of substrate proteins such as involucrin and loricrin to generate the cornified envelope at the cell periphery of the stratum corneum. We have shown that disruption of the TGase 1 gene in mice results in neonatal lethality, absence of the cornified envelope, and impaired skin barrier function. Based on the importance of TGase 1 in epidermal morphogenesis, we have now assessed its role in wound healing. In neonatal mouse skin, TGase 1 mRNA as well as keratin 6alpha was induced in the epidermis at the wound edges as early as 2 hours after injury and that expression continued in the migrating epidermis until completion of re-epithelialization. The TGase 1 enzyme co-localized on the plasma membrane of migrating keratinocytes with involucrin, but not with loricrin, which suggests the premature assembly of the cornified envelope. Similar injuries to TGase 1 knockout mouse skins grafted on athymic nude mice showed substantial delays in wound healing concomitant with sustained K6alpha mRNA induction. From these results, we suggest that activation of the TGase 1gene is essential for facilitated repair of skin injury. PMID- 11106561 TI - Iron overload and heart fibrosis in mice deficient for both beta2-microglobulin and Rag1. AB - Genetic causes of hereditary hemochromatosis (HH) include mutations in the HFE gene, a ss2-microglobulin (ss2m)-associated major histocompatibility complex class I-like protein. Accordingly, mutant ss2m(-/-) mice have increased intestinal iron absorption and develop parenchymal iron overload in the liver. In humans, other genetic and environmental factors have been suggested to influence the pathology and severity of HH. Previously, an association has been reported between low numbers of lymphocytes and the severity of clinical expression of the iron overload in HH. In the present study, the effect of a total absence of lymphocytes on iron overload was investigated by crossing ss2m(-/-) mice (which develop iron overload resembling human disease) with mice deficient in recombinase activator gene 1 (Rag1), which is required for normal B and T lymphocyte development. Iron overload was more severe in ss2mRag1 double deficient mice than in each of the single deficient mice, with iron accumulation in parenchymal cells of the liver, in acinar cells of the pancreas, and in heart myocytes. With increasing age ss2mRag1(-/-) mice develop extensive heart fibrosis, which could be prevented by reconstitution with normal hematopoietic cells. Thus, the development of iron-mediated cellular damage is substantially enhanced when a Rag1 mutation, which causes a lack of mature lymphocytes, is introduced into ss2m(-/-) mice. Mice deficient in ss2m and Rag1 thus offer a new experimental model of iron-related cardiomyopathy. PMID- 11106562 TI - Production of experimental malignant pleural effusions is dependent on invasion of the pleura and expression of vascular endothelial growth factor/vascular permeability factor by human lung cancer cells. AB - We determined the molecular mechanisms that regulate the pathogenesis of malignant pleural effusion (PE) associated with advanced stage of human, non small-cell lung cancer. Intravenous injection of human PC14 and PC14PE6 (adenocarcinoma) or H226 (squamous cell carcinoma) cells into nude mice yielded numerous lung lesions. PC14 and PC14PE6 lung lesions invaded the pleura and produced PE containing a high level of vascular endothelial growth factor (VEGF) localized vascular hyperpermeability. Lung lesions produced by H226 cells were confined to the lung parenchyma with no PE. The level of expression of VEGF mRNA and protein by the cell lines directly correlated with extent of PE formation. Transfection of PC14PE6 cells with antisense VEGF165 gene did not inhibit invasion into the pleural space but reduced PE formation. H226 cells transfected with either sense VEGF 165 or sense VEGF 121 genes induced localized vascular hyperpermeability and produced PE only after direct implantation into the thoracic cavity. The production of PE was thus associated with the ability of tumor cells to invade the pleura, a property associated with expression of high levels of urokinase-type plasminogen activator and low levels of TIMP-2. Collectively, the data demonstrate that the production of malignant PE requires tumor cells to invade the pleura and express high levels of VEGF/VPF. PMID- 11106563 TI - Role of nephrin in cell junction formation in human nephrogenesis. AB - Nephrin is a cell adhesion protein located at the slit diaphragm area of glomerular podocytes. Mutations in nephrin-coding gene (NPHS1) cause congenital nephrotic syndrome (NPHS1). We studied the developmental expression of nephrin, ZO-1 and P-cadherin in normal fetal kidneys and in NPHS1 kidneys. We used in situ hybridization and immunohistochemistry at light and electron microscopic levels. Nephrin and zonula occludens-1 (ZO-1) were first expressed in late S-shaped bodies. During capillary loop stage, nephrin and ZO-1 localized at the basal margin and in the cell-cell adhesion sites between developing podocytes, especially in junctions with ladder-like structures. In mature glomeruli, nephrin and ZO-1 concentrated at the slit diaphragm area. P-cadherin was first detected in ureteric buds, tubules, and vesicle stage glomeruli. Later, P-cadherin was seen at the basal margin of developing podocytes. Fetal NPHS1 kidneys with Fin major/Fin-major genotype did not express nephrin, whereas the expression of ZO-1 and P-cadherin was comparable to that of control kidneys. Although early junctional complexes proved structurally normal, junctions with ladder-like structures and slit diaphragms were completely missing. The results indicate that nephrin is dispensable for early development of podocyte junctional complexes. However, nephrin appears to be essential for formation of junctions with ladder like structures and slit diaphragms. PMID- 11106564 TI - Reduced heparan sulfate accumulation in enterocytes contributes to protein-losing enteropathy in a congenital disorder of glycosylation. AB - Intestinal biopsy in a boy with gastroenteritis-induced protein-losing enteropathy (PLE) showed loss of heparan sulfate (HS) and syndecan-1 core protein from the basolateral surface of the enterocytes, which improved after PLE subsided. Isoelectric focusing analysis of serum transferrin indicated a congenital disorder of glycosylation (CDG) and subsequent analysis showed three point mutations in the ALG6 gene encoding an alpha1,3-glucosyltransferase needed for the addition of the first glucose to the dolichol-linked oligosaccharide. The maternal mutation, C998T, causing an A333V substitution, has been shown to cause CDG-Ic, whereas the two paternal mutations, T391C (Y131H) and C924A (S308R) have not previously been reported. The mutations were tested for their ability to rescue faulty N:-linked glycosylation of carboxypeptidase Y in an ALG6-deficient Saccharomyces cerevisiae strain. Normal human ALG6 rescues glycosylation and A333V partially rescues, whereas the combined paternal mutations (Y131H and S308R) are ineffective. Underglycosylation resulting from each of these mutations is much more severe in rapidly dividing yeast. Similarly, incomplete protein glycosylation in the patient is most severe in rapidly dividing enterocytes during gastroenteritis-induced stress. Incomplete N:-linked glycosylation of an HS core protein and/or other biosynthetic enzymes may explain the selective localized loss of HS and PLE. PMID- 11106565 TI - The role of up-regulated serine proteases and matrix metalloproteinases in the pathogenesis of a murine model of colitis. AB - Proteinases are important at several phases of physiological and pathological inflammation, mediating cellular infiltration, cytokine activation, tissue damage, remodeling, and repair. However, little is known of their role in the pathogenesis of inflammatory bowel disease. The aim of this study was to assess the role of tissue proteases in a mouse model of colitis. Proteolytic activity was analyzed, using gel and in situ zymography, in colonic tissues from severe combined immunodeficient mice with colitis induced by transfer of CD4(+) T lymphocytes. Serine proteinase levels increased in colitic tissue, with major species of 23 kd, 30 kd, and 45 kd. Co-migration and inhibition studies indicated that the 23-kd proteinase was pancreatic trypsin and that the 30-kd species was neutrophil elastase. Matrix metalloproteinase (MMP)-9 expression, and MMP-2 and MMP-9 activation, was elevated in colitic tissues. Proteinase levels followed a decreasing concentration gradient from proximal to distal colon. Proteolysis was localized to infiltrating leukocytes in diseased severe combined immunodeficient mice. Transmural inflammation was associated with serine proteinase and MMP activity in overlying epithelium and with marked subepithelial proteolytic activity. The results demonstrate a clear elevation in the levels and activation of proteases in colitis, potentially contributing to disease progression through loss of epithelial barrier function. PMID- 11106566 TI - Inhibition of MAP kinase kinase causes morphological reversion and dissociation between soft agar growth and in vivo tumorigenesis in angiosarcoma cells. AB - Activated ras causes increased activity of several signal transduction systems, including the mitogen-activated protein kinase kinase (MAPKK) pathway and the phosphoinositol-3-kinase (PI-3-K) pathway. We have previously shown that the PI-3 K pathway plays a major role in regulation of ras-mediated tumor angiogenesis in angiosarcoma cells. However, the contribution of the MAPKK pathway to tumorigenesis and angiogenesis is not fully understood. Overexpression of constitutively active forms of MAPKK has previously been shown to transform nonmalignant NIH3T3 fibroblasts, but the effect of down-regulation of MAPKK on tumorigenesis and angiogenesis in a well established tumor has not been fully explored. We introduced a dominant negative MAPKK gene into SVR murine angiosarcoma cells. Introduction of a dominant negative MAPKK causes a significant decrease in proliferation rate in vitro and morphological reversion. Cells expressing the dominant negative MAPKK have a greatly decreased ability to form colonies in soft agar compared with wild-type cells. Despite the decreased cell growth in vitro and inability to grow in soft agar, the cells were equally tumorigenic in nude mice. Our results suggest that the MAPKK pathway is required for soft agar growth of angiosarcoma cells, and separates the phenotypes of soft agar growth versus in vivo tumorigenicity. These findings have implications in the development of signal transduction modulators as potential antineoplastic agents. PMID- 11106567 TI - Human colorectal cancers with an intact p16/cyclin D1/pRb pathway have up regulated p16 expression and decreased proliferation in small invasive tumor clusters. AB - A systematic spatial heterogeneity with high proliferative activity at the luminal border and low activity at the invasive margin is an unexpected behavior that has been observed in colorectal cancer (CRC). To clarify this phenomenon and possible underlying regulatory mechanisms, we have by immunohistochemistry elucidated the proliferative activity and the expression of G1/S regulatory proteins in small and large tumor cell clusters at the invasive margin in 97 CRCs. By identifying small tumor clusters at the tumor front, actually invading cancer cells could be characterized and analyzed separately. These cells could then be compared with the main tumor mass represented by the larger tumor clusters. The proliferation was significantly lower in small tumor clusters compared with larger clusters (P < 0.001) and the decrease in proliferation was correlated with a p16 up-regulation (r(s) = -0.41, P < 0.001). Interestingly, CRCs lacking p16 expression (18%) or tumors with other aberrations in the p16/cyclin D1/pRb pathway had a less pronounced decrease in proliferation between large and small clusters (P < 0.001), further strengthening the association between p16 and ceased proliferation at the invasive margin. This contrasts to tumors with low p27 or abnormal p53 levels showing sustained proliferation in small tumor clusters. Our findings imply that invading CRC cells generally have low proliferative activity, and this phenomenon seems to be mediated through p16 and the p16/cyclin D1/pRb pathway. PMID- 11106568 TI - Ligation of alpha4ss1 integrin on human intestinal mucosal mesenchymal cells selectively Up-regulates membrane type-1 matrix metalloproteinase and confers a migratory phenotype. AB - Human intestinal lamina propria mesenchymal cells show high surface expression of the alpha4ss1 integrin. Ligation of alpha4ss1 on mesenchymal cell lines with an activating monoclonal anti-alpha4 antibody or vascular cell adhesion molecule immunoglobulin (VCAM-IgG) leads to the appearance of activated forms of gelatinase A in culture supernatants, and the de novo expression of activated membrane type-1-matrix metalloproteinase (MT1-MMP). In functional assays, signaling through alpha4ss1 results in an increased capacity of mesenchymal cells to migrate through an artificial extracellular matrix, an effect inhibitable by excess tissue inhibitor of metalloproteinase-2. In organ cultures of human intestine, VCAM-IgG also up-regulates MT1-MMP, and in mucosal ulcers of inflammatory bowel disease patients, MT1-MMP transcripts are abundant, coincident with expression of VCAM-1 on cells at the ulcer margin. Collectively these results suggest that alpha4ss1-induced up-regulation of MT1-MMP may be a crucial factor in the migration of mesenchymal cells into ulcer beds during restitution of diseased gut mucosa. PMID- 11106569 TI - Hepatocyte transplantation into diseased mouse liver. Kinetics of parenchymal repopulation and identification of the proliferative capacity of tetraploid and octaploid hepatocytes. AB - To examine the process of liver repopulation by transplanted hepatocytes, we developed transgenic mice carrying a mouse major urinary protein-urokinase-type plasminogen activator fusion transgene. Expression of this transgene induced diffuse hepatocellular damage beginning at 3 weeks of age, and homozygous mice supported up to 97% parenchymal repopulation by healthy donor hepatocytes transplanted into the spleen. Using this transplantation model, we determined that 1) a mean of 21% of splenically injected hepatocytes engraft in liver parenchyma; 2) a mean of 6.6% of splenically injected hepatocytes (or one-third of engrafted cells) can give rise to proliferating hepatocyte foci; 3) transplanted cells in proliferating foci display an initial cell-doubling time of 28 hours, and focus growth continues through a mean of 12 cell doublings; 4) hepatocytes isolated from young and aged adult mice display similar focus repopulation kinetics; 5) the extent of repopulated parenchyma remains stable throughout the life of the recipient mouse; and 6) tetraploid and octaploid hepatocytes can support clonal proliferation. PMID- 11106570 TI - Loss of Fas-ligand expression in mouse keratinocytes during UV carcinogenesis. AB - Skin cells containing excessive ultraviolet (UV) radiation-induced DNA damage are eliminated by apoptosis that involves the p53 pathway and Fas/Fas-Ligand (Fas-L) interactions. To determine whether dysregulation of apoptosis plays a role in skin cancer development through disruption of Fas/Fas-L interactions, hairless SKH-hr1 mice were exposed to chronic UV irradiation from Kodacel-filtered FS40 lamps for 30 weeks. Their skin was analyzed for the presence of sunburn cells (apoptotic keratinocytes) and for Fas and Fas-L expression at various time points. A dramatic decrease in the numbers of morphologically identified sunburn cells and TUNEL-positive cells was detected as early as 1 week after chronic UV exposure began. After 4 weeks of chronic UV exposure, these cells were barely detectable. This defect in apoptosis was paralleled by an initial decrease in Fas L expression during the first week of chronic UV irradiation and a complete loss of expression after 4 weeks. Fas expression, however, increased during the course of chronic UV exposure. p53 mutations were detected in the UV-irradiated epidermis as early as 1 week after irradiation began and continued to accumulate with further UV exposure. Mice exposed to chronic UV began to develop skin tumors after approximately 8 weeks, and all mice had multiple skin tumors by 24 weeks. Most of the tumors expressed Fas but not Fas-L. We conclude that chronic UV exposure may induce a loss of Fas-L expression and a gain in p53 mutations, leading to dysregulation of apoptosis, expansion of mutated keratinocytes, and initiation of skin cancer. PMID- 11106571 TI - The cell cycle Cdc25A tyrosine phosphatase is activated in degenerating postmitotic neurons in Alzheimer's disease. AB - The Cdc25 phosphatases play key roles in cell-cycle progression by activating cyclin-dependent kinases. The latter are absent from neurons that are terminally differentiated in adult brain. However, accumulation of mitotic phosphoepitopes, and re-expression and activation of the M phase regulator, Cdc2/cyclin B, have been described in neurons undergoing degeneration in Alzheimer's disease (AD). To explain this atypical mitotic activation in neurons we investigated the Cdc2 activating Cdc25A phosphatase in human brain. The structural hallmarks of AD neurodegeneration, neurofibrillary tangles and neuritic plaques, were prominently immunolabeled with Cdc25A antibodies. In addition numerous neurons without visible structural alterations were also intensely stained, whereas control brain was very weakly positive. After immunoprecipitation from control and AD tissue, we found that the tyrosine dephosphorylating activity of Cdc25A against exogenous Cdc2 substrate was elevated in AD. Accordingly, Cdc25A from AD tissue displayed increased immunoreactivity with the mitotic phosphoepitope-specific antibody, MPM 2, and co-localized with MPM-2 immunoreactivity in AD neurons. These data suggest that Cdc25A participates in mitotic activation during neurodegeneration. The involvement of Cdc25A in cellular transformation, modulation of the DNA damage checkpoint, and linkage of mitogenic signaling to cell cycle machinery, also implicates one of these cell-cycle pathways in AD pathogenesis. PMID- 11106572 TI - Intracerebral recruitment and maturation of dendritic cells in the onset and progression of experimental autoimmune encephalomyelitis. AB - Dendritic cells (DCs) are thought to be key elements in the initiation and maintenance of autoimmune diseases. In this study, we sought evidence that DCs recruited to the central nervous system (CNS), a site that is primarily devoid of resident DCs, play a role in the effector phase and propagation of the immune response in experimental autoimmune encephalomyelitis (EAE). After immunization of SJL mice with proteolipid protein 139-151 peptide, process-bearing cells expressing the DC markers DEC-205 and CD11c appeared early in the spinal cord. During acute, chronic, and relapsing EAE, DEC-205(+) DCs expressing a lymphostimulatory phenotype (including the mature DC marker MIDC-8, major histocompatibility complex class II, CD40, and CD86 molecules) accumulated within the CNS inflammatory cell infiltrates. More prominent infiltration of the spinal cord parenchyma by mature DCs was observed in mice with relapsing disease. Macrophage inflammatory protein 3alpha, a chemokine active on DCs and lymphocytes, and its receptor CCR6 were up-regulated in the CNS during EAE. These findings suggest that intracerebral recruitment and maturation of DCs may be crucial in the local stimulation and maintenance of autoreactive immune responses, and that therapeutic strategies aimed at manipulating DC migration could be useful in the treatment of CNS autoimmune disorders. PMID- 11106573 TI - Astroglial expression of human alpha(1)-antichymotrypsin enhances alzheimer-like pathology in amyloid protein precursor transgenic mice. AB - Proteases and their inhibitors play key roles in physiological and pathological processes. Cerebral amyloid plaques are a pathological hallmark of Alzheimer's disease (AD). They contain amyloid-ss (Ass) peptides in tight association with the serine protease inhibitor alpha(1)-antichymotrypsin.(1,2) However, it is unknown whether the increased expression of alpha(1)-antichymotrypsin found in AD brains counteracts or contributes to the disease. We used regulatory sequences of the glial fibrillary acidic protein gene(3) to express human alpha(1) antichymotrypsin (hACT) in astrocytes of transgenic mice. These mice were crossed with transgenic mice that produce human amyloid protein precursors (hAPP) and Ass in neurons.(4,5) No amyloid plaques were found in transgenic mice expressing hACT alone, whereas hAPP transgenic mice and hAPP/hACT doubly transgenic mice developed typical AD-like amyloid plaques in the hippocampus and neocortex around 6 to 8 months of age. Co-expression of hAPP and hACT significantly increased the plaque burden at 7 to 8, 14, and 20 months. Both hAPP and hAPP/hACT mice showed significant decreases in synaptophysin-immunoreactive presynaptic terminals in the dentate gyrus, compared with nontransgenic littermates. Our results demonstrate that hACT acts as an amyloidogenic co-factor in vivo and suggest that the role of hACT in AD is pathogenic. PMID- 11106574 TI - Expression and characterization of trypsinogen produced in the human male genital tract. AB - Trypsinogen is a serine proteinase produced mainly by the pancreas, but it has recently been found to be expressed also in several cancers such as ovarian and colon cancer and in vascular endothelial cells. In this study, we found that trypsinogen-1 and -2 are present at high concentrations (median levels, 0.4 and 0.5 mg/L, respectively) in human seminal fluid and purified them to homogeneity by immunoaffinity and anion exchange chromatography. Purified trypsinogen isoenzymes displayed a M(r) of 25 to 28 kd in sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blotting. Most of the trypsinogen 1 purified from seminal fluid was enzymatically active whereas trypsinogen-2 occurred as the proform, which could be activated by enteropeptidase in vitro. Immunohistochemically, trypsinogen protein was detected in the human prostate, urethra, utriculus, ejaculatory duct, seminal vesicles, deferent duct, epididymal glands, and testis. Expression of trypsinogen mRNA in the same organs was demonstrated by in situ hybridization. Trypsinogen mRNA was also detected in the prostate and seminal vesicles by reverse transcriptase-polymerase chain reaction and Northern blotting. Isolated trypsin was shown to activate the proenzyme form of prostate-specific antigen. These results suggest that trypsinogen isoenzymes found in seminal fluid are produced locally in the male genital tract and that they may play a physiological role in the semen. PMID- 11106575 TI - Human nasopharyngeal-associated lymphoreticular tissues. Functional analysis of subepithelial and intraepithelial B and T cells from adenoids and tonsils. AB - Subepithelial and intraepithelial lymphocytes of human adenoids and tonsils were characterized and directly compared to determine the potential contribution of these tissues to mucosal and systemic immune responses. The distribution of T and B cell subsets, cytokine patterns, and antibody (Ab) isotype profiles were similar for adenoids and tonsils. Both tissues contained predominantly B cells ( approximately 65%), approximately 5% macrophages, and 30% CD3(+) T cells. The T cells were primarily of the CD4(+) subset ( approximately 80%). Tonsillar intraepithelial lymphocytes were also enriched in B cells. The analysis of dispersed cells revealed a higher frequency of cells secreting IgG than IgA and the predominant Ig subclass profiles were IgG1 > IgG3 and IgA1 > IgA2, respectively. In situ analysis also revealed higher numbers of IgG- than IgA positive cells. These IgG-positive cells were present in the epithelium and in the subepithelial zones of both tonsils and adenoids. Mitogen-triggered T cells from tonsils and adenoids produced both Th1- and Th2-type cytokines, clearly exhibiting their pluripotentiality for support of cell-mediated and Ab responses. Interestingly, antigen-specific T cells produced interferon-gamma and lower levels of interleukin-5. These results suggest that adenoids and tonsils of the nasopharyngeal-associated lymphoreticular tissues represent a distinct component of the mucosal-associated lymphoreticular tissues with features of both systemic and mucosal compartments. PMID- 11106576 TI - The CD44v7/8 epitope as a target to restrain proliferation of fibroblast-like synoviocytes in rheumatoid arthritis. AB - CD44 is a receptor for the glycosaminoglycan hyaluronan. It exists in a large range of isoforms because of variability in the pattern of glycosylation (both N- and O-linked) and of multiple splice variants. Human fibroblast-like synoviocytes derived from patients with rheumatoid arthritis express certain CD44 splice variants and we have investigated the functional implications of their expression. We found that the rate of proliferation of fibroblast-like synoviocytes expressing the CD44v7/8 epitope (average doubling time 55 hours) exceeds those obtained from the same synovial specimen but lacking this particular epitope (69 hours). Antibodies against CD44v7/8, but not against other exons, inhibit cell proliferation with concomitant induction of the cell cycle inhibitors GADD45, GADD153 and the cyclin-dependent protein-kinase inhibitors p21Waf/Cip. These data show that expression of CD44v7/8 contributes to the transformed phenotype of fibroblast-like synoviocytes. More importantly, they reveal the presence of a target that might be amenable to pharmacological intervention in the treatment of rheumatoid arthritis. PMID- 11106577 TI - Immunohistochemical analysis of endothelial-monocyte-activating polypeptide-II expression in vivo. AB - Endothelial-monocyte activating polypeptide (EMAP)-II is a novel molecule with cytokine-like pro-inflammatory properties, inducing procoagulant activity on the surface of endothelial cells and monocyte/macrophages in vitro, as well as up regulating E- and P-selectin expression. EMAP-II is chemotactic for monocytes/macrophages and neutrophils, and stimulates myeloperoxidase release from neutrophils. Injection of EMAP-II into the mouse footpad induces an acute inflammatory response, although some regression occurs in response to direct injection of EMAP-II into murine tumors. Very little is known about the expression of EMAP-II in normal tissues of mice or humans, or about its function in vivo. We developed polyclonal antibodies against EMAP-II using recombinant protein produced in Escherichia coli, and used these antibodies to carry out an immunohistochemical study of the occurrence and distribution of EMAP-II in human tissues. The distribution of EMAP-II protein is relatively restricted, occurring primarily in endocrine organs, in cells of neuroendocrine origin, but also in tissues with high turnover. EMAP-II is strongly expressed in secretory epithelial cells of the thyroid, pancreas, adrenal and salivary glands, among others, as well as in neurons and subsets of monocytes/macrophages. It is also found in the epithelium of the small and large intestines. We conclude that EMAP-II expression is usually, but not always, associated with tissues that display high turnover and high levels of protein synthesis. PMID- 11106578 TI - Chemokine receptor 1 knockout abrogates natural killer cell recruitment and impairs type-1 cytokines in lymphoid tissue during pulmonary granuloma formation. AB - Mice with targeted mutation of chemokine receptor 1 (CCR1) were used to assess the contribution of CCR1 agonists to local, regional, and systemic inflammatory related events during experimental pulmonary granuloma formation. Models of Th1 (type-1) and Th2 (type-2) cell-mediated lung granulomas were induced in wild-type (CCR+/+) and knockout (CCR1-/-) mice by embolizing Sepharose beads coupled to the purified protein derivative of Mycobacterium bovis or soluble antigens derived from Schistosoma mansoni eggs. Morphometric analysis indicated that granuloma sizes were unchanged in CCR1-/- mice, but flow cytometric analyses of dispersed granulomas revealed that natural killer cell recruitment to type-1 lesions was abrogated by 60%. Analysis of cytokine production by draining lymph node cultures showed altered expression in CCR1-/- mice characterized by reduced interleukin-2 and interferon-gamma in the type-1 response, and enhanced interleukin-5 and interleukin-13 in the type-2 response. Peripheral blood leukocytosis was also enhanced in the type-1 but not the type-2 response. These findings suggest that CCR1 agonists contribute to multiple immunoinflammatory events in the type-1 granulomatous response with natural killer cell accumulation being particularly sensitive to CCR1 disruption. Although functional efficacy of granulomas may be altered, chemokine redundancy and cytokine reserve seem to make the bulk of the exudative response resistant to CCR1 disruption. PMID- 11106579 TI - Calpain inhibitor I reduces the development of acute and chronic inflammation. AB - There is limited evidence that inhibition of the activity of the protease calpain I reduces inflammation. Here we investigate the effects of calpain inhibitor I in animal models of acute and chronic inflammation (carrageenan-induced pleurisy and collagen-induced arthritis). We report here for the first time that calpain inhibitor I (given at 5, 10, or 20 mg/kg i.p. in the pleurisy model or at 5 mg/kg i.p every 48 hours in the arthritis model) exerts potent anti-inflammatory effects (eg, inhibition of pleural exudate formation, mononuclear cell infiltration, delayed the development of the clinical signs and histological injury) in vivo. Furthermore, calpain inhibitor I reduced (1) the staining for nitrotyrosine and poly (ADP-ribose) polymerase (immunohistochemistry) and (2) the expression of inducible nitric oxide synthase and cyclooxygenase-2 in the lungs of carrageenan-treated rats and in joints from collagen-treated rats. Thus, prevention of the activation of calpain I reduces the development of acute and chronic inflammation. Inhibition of calpain I activity may represent a novel therapeutic approach for the therapy of inflammation. PMID- 11106580 TI - Involvement of IL-6, apart from its role in immunity, in mediating a chronic response during experimental arthritis. AB - Interleukin-6 (IL-6) is highly produced during arthritis but its exact function is still unknown. In this study we examined if IL-6, apart from its role in immunity, was involved in the local inflammatory response in experimental arthritis. IL-6 deficient (IL-6(-/-)) and wild-type mice were first compared in the antigen-induced arthritis model. IL-6 deficiency resulted in a mild, transient inflammation whereas wild-type mice developed a chronic, destructive synovitis. Wild-type mice immunized with one-tenth of the normal antigen dose still developed chronic arthritis despite low antibody levels, excluding reduced humoral immunity in IL-6(-/-) mice as a crucial phenomenon. In addition, passive immune-complex-induced arthritis did not differ between wild-type and IL-6(-/-) mice. Another option is reduced levels of Th1 cells in IL-6(-/-) mice. However, transfer of antigen-specific wild-type lymph node cells to IL-6(-/-) mice enhanced acute joint inflammation and increased cartilage damage but still could not sustain chronic inflammation, suggesting involvement of nonimmune elements of IL-6 activity in chronicity. In line with this, nonimmunologically mediated zymosan-induced arthritis developed similarly in the first week, but only wild type mice developed chronic synovitis. These results indicate an important role for IL-6 in propagation of joint inflammation, potentially independent of its role in immunity. PMID- 11106581 TI - Effect of apolipoprotein E allele epsilon4 on the initial phase of amyloid beta protein accumulation in the human brain. AB - Deposition of amyloid ss-protein (Ass), a hallmark of Alzheimer's disease, occurs to some extent in the brains of most elderly individuals. We sought to learn when Ass deposition begins and how deposition is affected by apolipoprotein E allele epsilon4, a strong risk factor for late-onset Alzheimer's disease. Using an improved extraction protocol and specific enzyme-linked immunosorbent assay, we quantified the levels of Ass40 and Ass42 in the insoluble fractions of brains from 105 autopsy cases, aged 22 to 81 years at death, who showed no signs of dementia. Ass40 and Ass42 were detected in the insoluble fractions from all of the brains examined; low levels were even found in the brains of patients as young as 20 to 30 years of age. The incidence of significant Ass accumulation increased age-dependently, with Ass42 levels beginning to rise steeply in some patients in their late 40's, accompanied by much smaller increases in Ass40 levels. The presence of the apolipoprotein E epsilon4 allele was found to significantly enhance the accumulation of Ass42 and, to a lesser extent, that of Ass40. These findings strongly suggest that the presence of epsilon4 allele results in an earlier onset of Ass42 accumulation in the brain. PMID- 11106582 TI - S20G mutant amylin exhibits increased in vitro amyloidogenicity and increased intracellular cytotoxicity compared to wild-type amylin. AB - Human amylin, a major constituent of pancreatic amyloid deposits, may be a pathogenetic factor for noninsulin-dependent diabetes mellitus (NIDDM). We demonstrated that the human amylin S20G gene mutation (S20G) was associated with a history of early onset, more severe type of NIDDM, linking the amylin gene to this disease. Also, we demonstrated that expression of human wild-type (WT) amylin in COS-1 cells leads to intracellular amyloidogenesis and induction of apoptosis, suggesting a possible mechanism for disease induction. Therefore we compared the abilities of S20G and WT amylin to induce apoptosis in transfected COS-1 cells and form amyloid in vitro. We transfected the rat (RAT), mutated human (MUT), WT, and S20G amylin genes into COS-1 cells and measured apoptosis using fluorescent-activated cell sorting analysis at 48, 72, and 96 hours. At 96 hours apoptosis increased significantly (P < 0.01) in cells transfected with WT and S20G over RAT or MUT (WT, 19%; S20G, 25%; RAT, 13%; and MUT, 12%) and the difference between WT and S20G was significant (P < 0.05). Synthetic WT and S20G monomeric peptides were used to generate amyloid fibrils in vitro as measured by the thioflavin T binding assay. The S20G amylin formed approximately twofold more amyloid at a rate approximately threefold higher than WT. Electron micrography indicated that the in vitro amyloid generated by WT and S20G amylins were morphologically indistinguishable. The results suggest that increased cytotoxicity by S20G is because of increased amyloidogenicity, which may be a causative factor in the early development of NIDDM, possibly through loss of ss cell mass. PMID- 11106583 TI - Onset of maternal arterial blood flow and placental oxidative stress. A possible factor in human early pregnancy failure. AB - The aim was to measure changes in the oxygen tension within the human placenta associated with onset of the maternal arterial circulation at the end of the first trimester of pregnancy, and the impact on placental tissues. Using a multiparameter probe we established that the oxygen tension rises steeply from <20 mmHg at 8 weeks of gestation to >50 mmHg at 12 weeks. This rise coincides with morphological changes in the uterine arteries that allow free flow of maternal blood into the placenta, and is associated with increases in the mRNA concentrations and activities of the antioxidant enzymes catalase, glutathione peroxidase, and manganese and copper/zinc superoxide dismutase within placental tissues. Between 8 to 9 weeks there is a sharp peak of expression of the inducible form of heat shock protein 70, formation of nitrotyrosine residues, and derangement of the mitochondrial cristae within the syncytiotrophoblast. We conclude that a burst of oxidative stress occurs in the normal placenta as the maternal circulation is established. We speculate that this may serve a physiological role in stimulating normal placental differentiation, but may also be a factor in the pathogenesis of pre-eclampsia and early pregnancy failure if antioxidant defenses are depleted. PMID- 11106584 TI - The metastatic ability of Ewing's sarcoma cells is modulated by stem cell factor and by its receptor c-kit. AB - Ewing's sarcoma is a primitive highly malignant tumor of bone and soft tissues usually metastasizing to bone, bone marrow, and lung. Growth factor receptors and their ligands may be involved in its growth and dissemination. We analyzed the expression of c-kit and its ligand stem cell factor (SCF) in a panel of six Ewing's sarcoma cell lines. All cell lines exhibited substantial levels of surface c-kit expression, and five of six displayed transmembrane SCF on the cell surface. Expression of c-kit was down-modulated in all lines by exposure to exogenous SCF. The SCF treatment was able to confer to cells a growth advantage in vitro, due both to an increase in cell proliferation and to a reduction in the apoptotic rate. When used in the lower compartment of a migration chamber, SCF acted as a strong chemoattractant for Ewing's sarcoma cells. The pretreatment of cells with SCF reduced their chemotactic response to SCF. In athymic nude mice, Ewing's sarcoma cells injected intravenously metastasized to the lung and to a variety of extrapulmonary sites, including bone and bone marrow. Metastatic sites resembled those observed in Ewing's sarcoma patients and corresponded to SCF-rich microenvironments. The in vitro pretreatment of cells with SCF strongly reduced the metastatic ability of Ewing's sarcoma cells, both to the lung and to extrapulmonary sites. This could be dependent on the down-modulation of c-kit expression observed in SCF-pretreated cells, leading to a reduced sensitivity to the chemotactic and proliferative actions of SCF. Our results indicate that the response to SCF mediated by c-kit may be involved in growth, migration, and metastatic ability of Ewing's sarcoma cells. PMID- 11106585 TI - Different subtypes of human lung adenocarcinoma caused by different etiological factors. Evidence from p53 mutational spectra. AB - Human lung adenocarcinomas are only relatively weakly associated with tobacco smoke, and other etiological factors need to be clarified. These may also vary with the histopathology. Because the p53 mutation status (frequency and spectrum) of a carcinoma can provide clues to causative agents, we subclassified 113 adenocarcinomas into five cell types: hobnail, columnar/cuboidal, mixed, polygonal, and goblet (54, 23, 18, 13, and 5, respectively) and investigated relationships with p53 mutations and smoking history. In the hobnail cell type, a low mutational frequency (37%) and a high proportion of transitions (65%), especially G:C to A:T transitions at CpG dinucleotides (45%) associated with spontaneous mutations, were found with a weak relation to tobacco smoke. In contrast, a high mutation frequency (70%) with a higher proportion of transversions (50%), especially G:C to T:A (44%) on the nontranscribed DNA strand, caused by exogenous carcinogenic agents like tobacco smoke, were observed for the columnar cell type, as with squamous cell carcinomas. These results indicate that two major subtypes of lung adenocarcinoma exist, one probably caused by tobacco smoke, and the other possibly due to spontaneous mutations. For the prevention of lung adenocarcinomas, in addition to stopping tobacco smoking, the elucidation of endogenous mechanisms is important. PMID- 11106586 TI - A feline model of experimentally induced islet amyloidosis. AB - The study of the pathogenesis of islet amyloidosis and its relationship to the development and progression of type 2 diabetes mellitus has been hampered by the lack of an experimentally inducible animal model. The domestic cat, by virtue of the fact that it is one of the few species that spontaneously develop a form of diabetes mellitus that closely resembles human type 2 diabetes, including the formation of amyloid deposits derived from islet amyloid polypeptide (IAPP), was considered to be an excellent candidate species in which to attempt to develop a nontransgenic animal model for this disease process. To develop the model, 8 healthy domestic cats were given a 50% pancreatectomy, which was followed by treatment with growth hormone and dexamethasone. Once a stable diabetic state was established, cats were randomly assigned to groups treated with either glipizide or insulin at doses appropriate to control hyperglycemia. Cats were maintained on this treatment regimen for 18 months and then euthanized. Based on light microscopic examination of Congo red-stained sections of pancreas, all cats were negative for the presence of islet amyloid at the time of pancreatectomy. At the end of the study all 4 glipizide-treated cats had islet amyloid deposits, whereas only 1 of 4 insulin-treated cats had detectable amyloid. In addition, the glipizide treated cats had threefold higher basal and fivefold higher glucose stimulated plasma IAPP concentrations than insulin-treated cats, suggesting an association between elevated IAPP secretion and islet amyloidosis. Blood glycosylated hemoglobin concentrations were not significantly different between the two treatment groups. This study documents for the first time an inducible model of islet amyloidosis in a nontransgenic animal. PMID- 11106587 TI - Development of spontaneous mammary tumors in BALB/c p53 heterozygous mice. A model for Li-Fraumeni syndrome. AB - Breast cancer is the most frequent tumor type among women in the United States and in individuals with Li-Fraumeni syndrome. The p53 tumor suppressor gene is altered in a large proportion of both spontaneous breast malignancies and Li Fraumeni breast cancers. This suggests that loss of p53 can accelerate breast tumorigenesis, yet p53-deficient mice rarely develop mammary tumors. To evaluate the effect of p53 loss on mammary tumor formation, the p53(null) allele was back crossed onto the BALB/c genetic background. Median survival was 15.4 weeks for BALB/c-p53(-/-) mice compared to 54 weeks for BALB/c-p53(+/-) mice. Sarcomas and lymphomas were the most frequent tumor types in BALB/c-p53(-/-) mice, whereas 55% of the female BALB/c-p53(+/-) mice developed mammary carcinomas. The mammary tumors were highly aneuploid, frequently lost the remaining wild-type p53 allele, but rarely lost BRCA1. Although mammary tumors were rarely detected in BALB/c p53(-/-) female mice, when glands from BALB/c-p53(-/-) mice were transplanted into wild-type BALB/c hosts, 75% developed mammary tumors. The high rate of mammary tumor development in the BALB/c background, but not C57Bl/6 or 129/Sv, suggests a genetic predisposition toward mammary tumorigenesis. Therefore, the BALB/c-p53(+/-) mice provide a unique model for the study of breast cancer in Li Fraumeni syndrome. These results demonstrate the critical role that the p53 tumor suppressor gene plays in preventing tumorigenesis in the mammary gland. PMID- 11106588 TI - Calcium buffers in flash-light. PMID- 11106590 TI - Engineering aspects of enzymatic signal transduction: photoreceptors in the retina. AB - Identifying the basic module of enzymatic amplification as an irreversible cycle of messenger activation/deactivation by a "push-pull" pair of opposing enzymes, we analyze it in terms of gain, bandwidth, noise, and power consumption. The enzymatic signal transduction cascade is viewed as an information channel, the design of which is governed by the statistical properties of the input and the noise and dynamic range constraints of the output. With the example of vertebrate phototransduction cascade we demonstrate that all of the relevant engineering parameters are controlled by enzyme concentrations and, from functional considerations, derive bounds on the required protein numbers. Conversely, the ability of enzymatic networks to change their response characteristics by varying only the abundance of different enzymes illustrates how functional diversity may be built from nearly conserved molecular components. PMID- 11106589 TI - Metabolically derived potential on the outer membrane of mitochondria: a computational model. AB - The outer mitochondrial membrane (OMM) is permeable to various small substances because of the presence of a voltage-dependent anion channel (VDAC). The voltage dependence of VDAC's permeability is puzzling, because the existence of membrane potential on the OMM has never been shown. We propose that steady-state metabolically derived potential (MDP) may be generated on the OMM as the result of the difference in its permeability restriction for various charged metabolites. To demonstrate the possibility of MDP generation, two models were considered: a liposomal model and a simplified cell model with a creatine kinase energy channeling system. Quantitative computational analysis of the simplified cell model shows that a MDP of up to -5 mV, in addition to the Donnan potential, may be generated at high workloads, even if the OMM is highly permeable to small inorganic ions, including potassium. Calculations show that MDP and DeltapH, generated on the OMM, depend on the cytoplasmic pH and energy demand rate. Computational modeling suggests that MDP may be important for cell energy metabolism regulation in multiple ways, including VDAC's permeability modulation and the effect of electrodynamic compartmentation. The osmotic pressure difference between the mitochondrial intermembrane space and the cytoplasm, as related to the electrodynamic compartmentation effects, might explain the morphological changes in mitochondria under intense workloads. PMID- 11106591 TI - A method for parameter optimization in computational biology. AB - Models in computational biology, such as those used in binding, docking, and folding, are often empirical and have adjustable parameters. Because few of these models are yet fully predictive, the problem may be nonoptimal choices of parameters. We describe an algorithm called ENPOP (energy function parameter optimization) that improves-and sometimes optimizes-the parameters for any given model and for any given search strategy that identifies the stable state of that model. ENPOP iteratively adjusts the parameters simultaneously to move the model global minimum energy conformation for each of m different molecules as close as possible to the true native conformations, based on some appropriate measure of structural error. A proof of principle is given for two very different test problems. The first involves three different two-dimensional model protein molecules having 12 to 37 monomers and four parameters in common. The parameters converge to the values used to design the model native structures. The second problem involves nine bumpy landscapes, each having between 4 and 12 degrees of freedom. For the three adjustable parameters, the globally optimal values are known in advance. ENPOP converges quickly to the correct parameter set. PMID- 11106592 TI - Analysis and implications of equivalent uniform approximations of nonuniform unitary synaptic systems. AB - Real synaptic systems consist of a nonuniform population of synapses with a broad spectrum of probability and response distributions varying between synapses, and broad amplitude distributions of postsynaptic unitary responses within a given synapse. A common approach to such systems has been to assume identical synapses and recover apparent quantal parameters by deconvolution procedures from measured evoked (ePSC) and unitary evoked postsynaptic current (uePSC) distributions. Here we explicitly consider nonuniform synaptic systems with both intra (type I) and intersynaptic (type II) response variability and formally define an equivalent system of uniform synapses in which both uePSC and ePSC amplitude distributions best approximate those of the actual nonuniform synaptic system. This equivalent system has the advantage of being fully defined by just four quantal parameters: n, the number of equivalent synapses;p, the mean probability of quantal release; mu, mean; and sigma(2), variance of the uePSC distribution. We show that these equivalent parameters are weighted averages of intrinsic parameters and can be approximated by apparent quantal parameters, therefore establishing a useful analytical link between the apparent and intrinsic parameters. The present study extends previous work on compound binomial analysis of synaptic transmission by highlighting the importance of the product of p and mu, and the variance of that product. Conditions for a unique deconvolution of apparent uniform synaptic parameters have been derived and justified. Our approach does not require independence of synaptic parameters, such as p and mu from each other, therefore the approach will hold even if feedback (i.e., via retrograde transmission) exists between pre and postsynaptic signals. Using numerical simulations we demonstrate how equivalent parameters are meaningful even when there is considerable variation in intrinsic parameters, including systems where subpopulations of high- and low-release probability synapses are present, therefore even under such conditions the apparent parameters estimated from experiments would be informative. PMID- 11106593 TI - A combined molecular dynamics and diffusion model of single proton conduction through gramicidin. AB - We develop a model for proton conduction through gramicidin based on the molecular dynamics simulations of Pomes and Roux (Biophys. J. 72:A246, 1997). The transport of a single proton through the gramicidin pore is described by a potential of mean force and diffusion coefficient obtained from the molecular dynamics. In addition, the model incorporates the dynamics of a defect in the hydrogen bonding structure of pore waters without an excess proton. Proton entrance and exit were not simulated by the molecular dynamics. The single proton conduction model includes a simple representation of these processes that involves three free parameters. A reasonable value can be chosen for one of these, and the other two can be optimized to yield a good fit to the proton conductance data of, Ann. N.Y. Acad. Sci. 339:8-20) for pH > or = 1.7. A sensitivity analysis shows the significance of this fit. PMID- 11106594 TI - Fluorescence intensity multiple distributions analysis: concurrent determination of diffusion times and molecular brightness. AB - Fluorescence correlation spectroscopy (FCS) has proven to be a powerful technique with single-molecule sensitivity. Recently, it has found a complement in the form of fluorescence intensity distribution analysis (FIDA). Here we introduce a fluorescence fluctuation method that combines the features of both techniques. It is based on the global analysis of a set of photon count number histograms, recorded with multiple widths of counting time intervals simultaneously. This fluorescence intensity multiple distributions analysis (FIMDA) distinguishes fluorescent species on the basis of both the specific molecular brightness and the translational diffusion time. The combined information, extracted from a single measurement, increases the readout effectively by one dimension and thus breaks the individual limits of FCS and FIDA. In this paper a theory is introduced that describes the dependence of photon count number distributions on diffusion coefficients. The theory is applied to a series of photon count number histograms corresponding to different widths of counting time intervals. Although the ability of the method to determine specific brightness values, diffusion times, and concentrations from mixtures is demonstrated on simulated data, its experimental utilization is shown by the determination of the binding constant of a protein-ligand interaction exemplifying its broad applicability in the life sciences. PMID- 11106595 TI - Lipid-mediated interactions between intrinsic membrane proteins: a theoretical study based on integral equations. AB - This study of lipid-mediated interactions between proteins is based on a theory recently developed by the authors for describing the structure of the hydrocarbon chains in the neighborhood of a protein inclusion embedded in a lipid membrane [Lague et al., Farad. Discuss. 111:165-172, 1998]. The theory involves the hypernetted chain integral equation formalism for liquids. The exact lateral density-density response function of the hydrocarbon core, extracted from molecular dynamics simulations of a pure dipalmitoylphosphatidylcholine bilayer based on an atomic model, is used as input. For the sake of simplicity, protein inclusions are modeled as hard repulsive cylinders. Numerical calculations were performed with three cylinder sizes: a small cylinder of 2.5-A radius, corresponding roughly to an aliphatic chain; a medium cylinder of 5-A radius, corresponding to a alpha-helical polyalanine protein; and a large cylinder of 9-A radius, representing a small protein, such as the gramicidin channel. The calculations show that the average hydrocarbon density is perturbed over a distance of 20-25 A from the edge of the cylinder for every cylinder size. The lipid-mediated protein-protein effective interaction is calculated and is shown to be nonmonotonic. In the case of the small and the medium cylinders, the lipid mediated effective interaction of two identical cylinders is repulsive at an intermediate range but attractive at short range. At contact, there is a free energy of -2k(B)T for the 2.5-A-radius cylinder and -9k(B)T for the 5-A-radius cylinder, indicating that the association of two alpha-helices of both sizes is favored by the lipid matrix. In contrast, the effective interaction is repulsive at all distances in the case of the large cylinder. Results were obtained with two integral equations theories: hypernetted chain and Percus-Yevick. For the two theories, all results are qualitatively identical. PMID- 11106597 TI - A quantitative approximation scheme for the traveling wave solutions in the Hodgkin-Huxley model. AB - We introduce an approximation scheme for the Hodgkin-Huxley model of nerve conductance that allows calculation of both the speed and shape of the traveling pulses, in quantitative agreement with the solutions of the model. We demonstrate that the reduced problem for the front of the traveling pulse admits a unique solution. We obtain an explicit analytical expression for the speed of the pulses that is valid with good accuracy in a wide range of the parameters. PMID- 11106596 TI - The phantom burster model for pancreatic beta-cells. AB - Pancreatic beta-cells exhibit bursting oscillations with a wide range of periods. Whereas periods in isolated cells are generally either a few seconds or a few minutes, in intact islets of Langerhans they are intermediate (10-60 s). We develop a mathematical model for beta-cell electrical activity capable of generating this wide range of bursting oscillations. Unlike previous models, bursting is driven by the interaction of two slow processes, one with a relatively small time constant (1-5 s) and the other with a much larger time constant (1-2 min). Bursting on the intermediate time scale is generated without need for a slow process having an intermediate time constant, hence phantom bursting. The model suggests that isolated cells exhibiting a fast pattern may nonetheless possess slower processes that can be brought out by injecting suitable exogenous currents. Guided by this, we devise an experimental protocol using the dynamic clamp technique that reliably elicits islet-like, medium period oscillations from isolated cells. Finally, we show that strong electrical coupling between a fast burster and a slow burster can produce synchronized medium bursting, suggesting that islets may be composed of cells that are intrinsically either fast or slow, with few or none that are intrinsically medium. PMID- 11106598 TI - Protein motions at zero-total angular momentum: the importance of long-range correlations. AB - A constant-energy molecular dynamics simulation is used to monitor protein motion at zero-total angular momentum. With a simple protein model, it is shown that overall rotation is possible at zero-total angular momentum as a result of flexibility. Since the rotational motion is negligible on a time scale of 1000 reduced time units, the essentially rotation-free portion of the trajectory provides an unbiased test of the common approximate methods for separating overall rotation from internal motions by optimal superposition. Removing rotation by minimizing the root-mean-square deviation (RMSD) for the entire system is found to be more appropriate than using the RMSD for only the more rigid part of the system. The results verify the existence of positive cross correlation in the motions of atoms separated by large distances. PMID- 11106599 TI - Heparan sulfate biosynthesis: a theoretical study of the initial sulfation step by N-deacetylase/N-sulfotransferase. AB - Heparan sulfate N-deacetylase/N-sulfotransferase (NDST) catalyzes the deacetylation and sulfation of N-acetyl-D-glucosamine residues of heparan sulfate, a key step in its biosynthesis. Recent crystallographic and mutational studies have identified several potentially catalytic residues of the sulfotransferase domain of this enzyme (, J. Biol. Chem. 274:10673-10676). We have used the x-ray crystal structure of heparan sulfate N-sulfotransferase with 3'-phosphoadenosine 5'-phosphate to build a solution model with cofactor 3' phosphoadenosine 5'-phosphosulfate (PAPS) and a model heparan sulfate ligand bound, and subsequently performed a 2-ns dynamics solution simulation. The simulation results confirm the importance of residues Glu(642), Lys(614), and Lys(833), with the possible involvement of Thr(617) and Thr(618), in binding PAPS. Additionally, Lys(676) is found in close proximity to the reaction site in our solvated structure. This study illustrates for the first time the possible involvement of water in the catalysis. Three water molecules were found in the binding site, where they are coordinated to PAPS, heparan sulfate, and the catalytic residues. PMID- 11106600 TI - The effects of non-identifiability on testing for detailed balance in aggregated Markov models for ion-channel gating. AB - Aggregated Markov models are a widely used tool to model patch clamp data measured from single ion channels. These channels must obey the principle of detailed balance in thermodynamic equilibrium; otherwise, the channel is driven by an external source of energy. We investigate the power of a likelihood ratio test for detailed balance for a number of data points which is in the order of magnitude of patch clamp experiments. We show that for certain models with nearly equal dwell times, a test for detailed balance suffers from a loss of power to detect violations of detailed balance which is due to the non-identifiability of the transition rates for models with equal dwell times. PMID- 11106602 TI - Fluctuations in repressor control: thermodynamic constraints on stochastic focusing. AB - The influence of fluctuations in molecule numbers on genetic control circuits has received considerable attention. The consensus has been that such fluctuations will make regulation less precise. In contrast, it has more recently been shown that signal fluctuations can sharpen the response in a regulated process by the principle of stochastic focusing (SF) (, Proc. Natl. Acad. Sci. USA. 97:7148 7153). In many cases, the larger the fluctuations are, the sharper is the response. Here we investigate how fluctuations in repressor or corepressor numbers can improve the control of gene expression. Because SF is found to be constrained by detailed balance, this requires that the control loops contain driven processes out of equilibrium. Some simple and realistic out-of-equilibrium steps that will break detailed balance and make room for SF in such systems are discussed. We conclude that when the active repressors are controlled by corepressor molecules that display large ("coherent") number fluctuations or when corepressors can be irreversibly removed directly from promoter-bound repressors, the response in gene activity can become significantly sharper than without intrinsic noise. A simple experimental design to establish the possibility of SF for repressor control is suggested. PMID- 11106601 TI - Modeling zymogen protein C. AB - A solution structure for the complete zymogen form of human coagulation protein C is modeled. The initial core structure is based on the x-ray crystallographic structure of the gamma-carboxyglutamic acid (Gla)-domainless activated form. The Gla domain (residues 1-48) is modeled from the x-ray crystal coordinates of the factor VII(a)/tissue factor complex and oriented with the epidermal growth factor 1 domain to yield an initial orientation consistent with the x-ray crystal structure of porcine factor IX(a). The missing C-terminal residues in the light chain (residues 147-157) and the activation peptide residues 158-169 were introduced using homology modeling so that the activation peptide residues directly interact with the residues in the calcium binding loop. Molecular dynamics simulations (Amber-particle-mesh-Ewald) are used to obtain the complete calcium-complexed solution structure. The individual domain structures of protein C in solution are largely unaffected by solvation, whereas the Gla-epidermal growth factor-1 orientation evolves to a form different from both factors VII(a) and IX(a). The solution structure of the zymogen protein C is compared with the crystal structures of the existing zymogen serine proteases: chymotrypsinogen, proproteinase, and prethrombin-2. Calculated electrostatic potential surfaces support the involvement of the serine protease calcium ion binding loop in providing a suitable electrostatic environment around the scissile bond for II(a)/thrombomodulin interaction. PMID- 11106603 TI - Kinetics of association of anti-lysozyme monoclonal antibody D44.1 and hen-egg lysozyme. AB - Association rate constants for antigen/antibody associations have been computed by Brownian Dynamics simulations of D. L. Ermak and J. A. McCammon, J. Chem. Phys. 69:1352-1360, 1978. The model of monoclonal antibody (mAb) D44.1 is based on crystallographic data (B. C. Braden et al., J. Mol. Biol. 243:767-781, 1994). Electrostatic forces that steer the antigen to the antibody-combining site are computed by solving the linearized Poisson-Boltzmann equation. D44. 1-HEL complex displays very similar association motifs to a related anti-lysozyme antibody, HyHEL-5-HEL system. The computed association rate constants are comparable in the two systems, although the experimental affinity constants differ by three orders of magnitude (D. Tello et al., Biochem. Soc. Trans. 21:943-946, 1993; K. A. Hibbits et al., Biochemistry. 33:3584-3590, 1994). Simulations suggest that the origin of the differences in the affinity come from dissociation rate constants. We have also carried out simulation experiments on a number of mutant antibody fragment-HEL associations to address the role of electrostatics and, to a limited extent, the orientational aspects of association. PMID- 11106604 TI - Residence times of water molecules in the hydration sites of myoglobin. AB - Hydration sites are high-density regions in the three-dimensional time-averaged solvent structure in molecular dynamics simulations and diffraction experiments. In a simulation of sperm whale myoglobin, we found 294 such high-density regions. Their positions appear to agree reasonably well with the distributions of waters of hydration found in 38 x-ray and 1 neutron high-resolution structures of this protein. The hydration sites are characterized by an average occupancy and a combination of residence time parameters designed to approximate a distribution of residence times. It appears that although the occupancy and residence times of the majority of sites are rather bulk-like, the residence time distribution is shifted toward the longer components, relative to bulk. The sites with particularly long residence times are located only in the cavities and clefts of the protein. This indicates that other factors, such as hydrogen bonds and hydrophobicity of underlying protein residues, play a lesser role in determining the residence times of the longest-lived sites. PMID- 11106605 TI - Na(+)-dependent Ca(2+) transport modulates the secretory response to the Fcepsilon receptor stimulus of mast cells. AB - Immunological stimulation of rat mucosal-type mast cells (RBL-2H3 line) by clustering of their Fcepsilon receptors (FcepsilonRI) causes a rapid and transient increase in free cytoplasmic Ca(2+) ion concentration ([Ca(2+)](i)) because of its release from intracellular stores. This is followed by a sustained elevated [Ca(2+)](i), which is attained by Ca(2+) influx. Because an FcepsilonRI induced increase in the membrane permeability for Na(+) ions has also been observed, and secretion is at least partially inhibited by lowering of extracellular sodium ion concentrations ([Na(+)](o)), the operation of a Na(+)/Ca(2+) exchanger has been considered. We found significant coupling between the Ca(2+) and Na(+) ion gradients across plasma membranes of RBL-2H3 cells, which we investigated employing (23)Na-NMR, (45)Ca(2+), (85)Sr(2+), and the Ca(2+)-sensitive fluorescent probe indo-1. The reduction in extracellular Ca(2+) concentrations ([Ca(2+)](o)) provoked a [Na(+)](i) increase, and a decrease in [Na(+)](o) results in a Ca(2+) influx as well as an increase in [Ca(2+)](i). Mediator secretion assays, monitoring the released beta-hexosaminidase activity, showed in the presence of extracellular sodium a sigmoidal dependence on [Ca(2+)](o). However, the secretion was not affected by varying [Ca(2+)](o) as [Na(+)](o) was lowered to 0.4 mM, while it was almost completely inhibited at [Na(+)](o) = 136 mM and [Ca(2+)](o) < 0.05 mM. Increasing [Na(+)](o) caused the secretion to reach a minimum at [Na(+)](o) = 20 mM, followed by a steady increase to its maximum value at 136 mM. A parallel [Na(+)](o) dependence of the Ca(2+) fluxes was observed: Antigen stimulation at [Na(+)](o) = 136 mM caused a pronounced Ca(2+) influx. At [Na(+)](o) = 17 mM only a slight Ca(2+) efflux was detected, whereas at [Na(+)](o) = 0.4 mM no Ca(2+) transport across the cell membrane could be observed. Our results clearly indicate that the [Na(+)](o) dependence of the secretory response to FcepsilonRI stimulation is due to its influence on the [Ca(2+)](i), which is mediated by a Na(+)-dependent Ca(2+) transport. PMID- 11106606 TI - Actin protofilament orientation in deformation of the erythrocyte membrane skeleton. AB - The red cell's spectrin-actin network is known to sustain local states of shear, dilation, and condensation, and yet the short actin filaments are found to maintain membrane-tangent and near-random azimuthal orientations. When calibrated with polarization results for single actin filaments, imaging of micropipette deformed red cell ghosts has allowed an assessment of actin orientations and possible reorientations in the network. At the hemispherical cap of the aspirated projection, where the network can be dilated severalfold, filaments have the same membrane-tangent orientation as on a relatively unstrained portion of membrane. Likewise, over the length of the network projection pulled into the micropipette, where the network is strongly sheared in axial extension and circumferential contraction, actin maintains its tangent orientation and is only very weakly aligned with network extension. Similar results are found for the integral membrane protein Band 3. Allowing for thermal fluctuations, we deduce a bound for the effective coupling constant, alpha, between network shear and azimuthal orientation of the protofilament. The finding that alpha must be about an order of magnitude or more below its tight-coupling value illustrates how nanostructural kinematics can decouple from more macroscopic responses. Monte Carlo simulations of spectrin-actin networks at approximately 10-nm resolution further support this conclusion and substantiate an image of protofilaments as elements of a high-temperature spin glass. PMID- 11106607 TI - Interferon-gamma and sinusoidal electric fields signal by modulating NAD(P)H oscillations in polarized neutrophils. AB - Metabolic activity in eukaryotic cells is known to naturally oscillate. We have recently observed a 20-s period NAD(P)H oscillation in neutrophils and other polarized cells. Here we show that when polarized human neutrophils are exposed to interferon-gamma or to ultra-low-frequency electric fields with periods double that of the NAD(P)H oscillation, the amplitude of the NAD(P)H oscillations increases. Furthermore, increases in NAD(P)H amplitude, whether mediated by interferon-gamma or by an oscillating electric field, signals increased production of reactive oxygen metabolites. Hence, amplitude modulation of NAD(P)H oscillations suggests a novel signaling mechanism in polarized cells. PMID- 11106609 TI - Block of wild-type and inactivation-deficient cardiac sodium channels IFM/QQQ stably expressed in mammalian cells. AB - The role of inactivation as a central mechanism in blockade of the cardiac Na(+) channel by antiarrhythmic drugs remains uncertain. We have used whole-cell and single channel recordings to examine the block of wild-type and inactivation deficient mutant cardiac Na(+) channels, IFM/QQQ, stably expressed in HEK-293 cells. We studied the open-channel blockers disopyramide and flecainide, and the lidocaine derivative RAD-243. All three drugs blocked the wild-type Na(+) channel in a use-dependent manner. There was no use-dependent block of IFM/QQQ mutant channels with trains of 20 40-ms pulses at 150-ms interpulse intervals during disopyramide exposure. Flecainide and RAD-243 retained their use-dependent blocking action and accelerated macroscopic current relaxation. All three drugs reduced the mean open time of single channels and increased the probability of their failure to open. From the abbreviation of the mean open times, we estimated association rates of approximately 10(6)/M/s for the three drugs. Reducing the burst duration contributed to the acceleration of macroscopic current relaxation during exposure to flecainide and RAD-243. The qualitative differences in use dependent block appear to be the result of differences in drug dissociation rate. The inactivation gate may play a trapping role during exposure to some sodium channel blocking drugs. PMID- 11106608 TI - Binding kinetics of calbindin-D(28k) determined by flash photolysis of caged Ca(2+) AB - We have used UV flash photolysis of DM-nitrophen in combination with model-based analysis of Oregon Green 488 BAPTA-5N fluorescence transients to study the kinetics of Ca(2+) binding to calbindin-D(28K). The experiments used saturated DM nitrophen at a [Ca(2+)] of 1.5 microM. Under these conditions, UV laser flashes produced rapid steplike increases in [Ca(2+)] in the absence of calbindin-D(28K), and in its presence the decay of the flash-induced fluorescence was due solely to the Ca(2+) buffering by the protein. We developed a novel method for kinetic parameter derivation and used the synthetic Ca(2+) buffer EGTA to confirm its validity. We provide evidence that calbindin-D(28K) binds Ca(2+) in at least two distinct kinetic patterns, one arising from high-affinity sites that bind Ca(2+) with a k(on) comparable to that of EGTA (i.e., approximately 1 x 10(7) M(-1) s( 1)) and another with lower affinity and an approximately eightfold faster k(on). In view of the inability of conventional approaches to adequately resolve rapid Ca(2+) binding kinetics of Ca(2+) buffers, this method promises to be highly valuable for studying the Ca(2+) binding properties of other biologically important Ca(2+) binding proteins. PMID- 11106610 TI - The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels. AB - Intercellular channels formed of members of the gene family of connexins (Cxs) vary from being substantially cation selective to being anion selective. We took advantage of the ability of Cx46 to function as an unopposed hemichannel to examine the basis of Cx charge selectivity. Previously we showed Cx46 hemichannels to be large pores that predominantly conduct cations and inwardly rectify in symmetric salts, properties suggesting selectivity is influenced by fixed negative charges located toward the extracellular end of the pore. Here we demonstrate that high ionic strength solutions applied to the extracellular, but not the intracellular, side of Cx46 hemichannels substantially reduce the ratio of cation to anion permeability. Substitution of the first extracellular loop (E1) domain of Cx32, an anion-preferring Cx, reduces conductance, converts Cx46 from cation to anion preferring, and changes the I-V relation form inwardly to outwardly rectifying. These data suggest that fixed negative charges influencing selectivity in Cx46 are located in E1 and are substantially reduced and/or are replaced with positive charges from the Cx32 E1 sequence. Extending studies to Cx46 cell-cell channels, we show that they maintain a strong preference for cations, have a conductance nearly that expected by the series addition of hemichannels, but lack rectification in symmetric salts. These properties are consistent with preservation of the fixed charge region in E1 of hemichannels, which upon docking, become symmetrically placed near the center of the cell-cell channel pore. Furthermore, heterotypic cell-cell channels formed by pairing Cx46 with Cx32 or Cx43 rectify in symmetric salts in accordance with the differences in the charges we ascribed to E1. These data are consistent with charged residues in E1 facing the channel lumen and playing an important role in determining Cx channel conductance and selectivity. PMID- 11106611 TI - Mg(2+) block unmasks Ca(2+)/Ba(2+) selectivity of alpha1G T-type calcium channels. AB - We have examined permeation by Ca(2+) and Ba(2+), and block by Mg(2+), using whole-cell recordings from alpha1G T-type calcium channels stably expressed in HEK 293 cells. Without Mg(o)(2+), inward currents were comparable with Ca(2+) and Ba(2+). Surprisingly, three other results indicate that alpha1G is actually selective for Ca(2+) over Ba(2+). 1) Mg(2+) block is approximately 7-fold more potent with Ba(2+) than with Ca(2+). With near-physiological (1 mM) Mg(o)(2+), inward currents were approximately 3-fold larger with 2 mM Ca(2+) than with 2 mM Ba(2+). The stronger competition between Ca(2+) and Mg(2+) implies that Ca(2+) binds more tightly than Ba(2+). 2) Outward currents (carried by Na(+)) are blocked more strongly by Ca(2+) than by Ba(2+). 3) The reversal potential is more positive with Ca(2+) than with Ba(2+), thus P(Ca) > P(Ba). We conclude that alpha1G can distinguish Ca(2+) from Ba(2+), despite the similar inward currents in the absence of Mg(o)(2+). Our results can be explained by a 2-site, 3-barrier model if Ca(2+) enters the pore 2-fold more easily than Ba(2+) but exits the pore at a 2-fold lower rate. PMID- 11106612 TI - Rhodopsin activation affects the environment of specific neighboring phospholipids: an FTIR spectroscopic study. AB - Rhodopsin is a member of a superfamily of G-protein-coupled receptors that transduce signals across membranes. We used Fourier-transform infrared (FTIR) difference spectroscopy to study the interaction between rhodopsin and lipid bilayer upon receptor activation. A difference band at 1744 cm(-1) (+)/1727 cm( 1) (-) was identified in the FTIR-difference spectrum of rhodopsin mutant D83N/E122Q in which spectral difference bands arising from the carbonyl stretching frequencies of protonated carboxylic acid groups were removed by mutation. As the band was abolished by detergent delipidation, we suggested that it arose from carbonyl groups of phospholipid fatty acid esters. Rhodopsin and the D83N/E122Q mutant were reconstituted into various (13)C-labeled 1-palmitoyl-2 oleoyl-sn-glycero-3-phosphocholine vesicles and probed. The 1744-cm(-1) (+)/1727 cm(-1) (-) band could be unequivocally assigned to a change in the lipid ester carbonyl stretch upon receptor activation, with roughly equal contribution from both lipid esters. The band intensity scaled with the amount of rhodopsin but not with the amount of lipid, excluding the possibility that it was due to the bulk lipid phase. We also excluded the possibility that the lipid band represents a change in the number of boundary lipids or a general alteration in the boundary lipid environment upon formation of metarhodopsin II. Instead, the data suggest that the lipid band represents the change of a specific lipid-receptor interaction that is coupled to protein conformational changes. PMID- 11106613 TI - The anomalous mole fraction effect in Chara: gating at the edge of temporal resolution. AB - The anomalous mole fraction effect (AMFE) of the K(+) channel in excised patches of the tonoplast of Chara showed a minimum of apparent open-channel current at 20 mM Tl(+) and 230 mM K(+). Time series obtained at a sampling rate of 100 kHz (filter 25 kHz) were analyzed by three methods to find out whether the AMFE results from an effect on gating or on the conductivity of the open state. Fitting the amplitude histograms by a superposition of gaussians showed a broadening in the presence of Tl(+). Dwell-time analysis based on an O-O-C-C-C model failed to evaluate rate constants above the filter frequency. Thus, the absence of any reduction of apparent open-channel current in time series simulated with the evaluated rate constants could not be taken as evidence against the hypothesis of gating. Finally, a direct fit of the measured time series using five different 5-state Hidden Markov models revealed that the presence of Tl(+) changed the rate constants in such a way that the number of transitions into the short-lived open state (30 micros) increased strongly compared to those in the absence of Tl(+). These models explain 25% reduction of apparent single-channel current amplitude through a rapid gating mechanism. PMID- 11106614 TI - Modeling and docking the endothelin G-protein-coupled receptor. AB - A model of the endothelin G-protein-coupled receptor (ET(A)) has been constructed using a segmented approach. The model was produced using a bovine rhodopsin model as a template for the seven transmembrane alpha-helices. The three cytoplasmic loop regions and the C-terminal region were modeled on NMR structures of corresponding segments from bovine rhodopsin. The three extracellular loops were modeled on homologous loop regions in other proteins of known structure. The N terminal region was modeled as a three-helix domain based on its homology with a hydrolase protein. To test the model, the FTDOCK algorithm was used to predict the ligand-binding site for the crystal structure of human endothelin. The site of docking is consistent with mutational and biochemical data. The principal sites of interaction in the endothelin ligand all lie on one face of a helix that has been implicated by structure-activity relationship studies as being essential for binding. As further support for the model, attempts to dock bigET, an inactive precursor to endothelin that does not bind to the receptor, found no sites for tight binding. The model of the receptor-ligand complex produced forms a basis for rational drug design of agonists and antagonists for this G-protein coupled receptor. PMID- 11106615 TI - Determinants of excitability in cardiac myocytes: mechanistic investigation of memory effect. AB - The excitability of a cardiac cell depends upon many factors, including the rate and duration of pacing. Furthermore, cell excitability and its variability underlie many electrophysiological phenomena in the heart. In this study, we used a detailed mathematical model of the ventricular myocyte to investigate the determinants of excitability and gain insight into the mechanism by which excitability depends on the rate and duration of pacing (the memory effect). RESULTS: i) The primary determinant of excitability depends upon the duration (T) of the stimulus. ii) For a short T, excitability is determined by the difference between the threshold membrane potential and the resting membrane potential. iii) For a long T, excitability is determined by the resting membrane resistance, R(m). iv) In the case of long T, pacing induced changes in [Na(+)](i) and [Ca(2+)](i) over time affect R(m) and excitability by shifting the current voltage (IV) curve in the vertical direction and are responsible for the memory effect. CONCLUSIONS: The results have important implications during an arrhythmia, where a cardiac cell may be subjected to rapid repetitive excitation for an extended period of time. Effective anti-arrhythmic strategies may be developed to exploit the R(m) dependence of excitability for a long T. PMID- 11106616 TI - Heme structure and orientation in single monolayers of cytochrome c on polar and nonpolar soft surfaces. AB - Polarized x-ray absorption fine structure (XAFS) spectroscopy has been performed in fluorescence mode under total external reflection conditions on frozen hydrated single monolayers of yeast cytochrome c (YCC). The protein molecules were vectorially oriented within the monolayer by tethering their naturally occurring and unique surface cysteine residues to the sulfhydryl-endgroups at the surface of a mixed organic self-assembled monolayer, itself covalently attached to an ultrapure silicon wafer. The sulfhydryl-endgroups were isolated by dilution with either methyl- or hydroxyl-endgroups, producing macroscopically nonpolar or uncharged-polar soft surfaces, respectively. Independent information on the heme plane orientation relative to the monolayer plane was obtained experimentally via optical linear dichroism. The polarized XAFS data have been analyzed both qualitatively and by a global mapping approach limited to systematically altering the various iron-ligand distances within a model for the local atomic environment of the heme prosthetic group, and comparing the theoretically generated XAFS spectra with those obtained experimentally. A similar analysis of unpolarized XAFS data from a frozen solution of YCC was performed using either the heme environment from the NMR solution or the x-ray crystallographic data for YCC as the model structure. All resulting iron-ligand distances were then used in molecular dynamics (MD) computer simulations of YCC in these three systems to investigate the possible effects of anisotropic ligand motions on the fits of the calculated to the experimental XAFS spectra. PMID- 11106617 TI - Molecular dynamics generation of nonarbitrary membrane models reveals lipid orientational correlations. AB - This report addresses the following problems associated with the generation of computer models of phospholipid bilayer membranes using molecular dynamics simulations: arbitrary initial structures and short equilibration periods, an Ewald-induced strong coupling of phospholipids, uncertainty regarding which value should be used for surface tension to alleviate the problem of the small size of the membrane, and simultaneous realization of both order parameters and the surface area. We generated a computer model of the liquid-crystalline L-alpha dimyristoylphosphatidylcholine (DMPC) bilayer, starting from a configuration based on a crystal structure (rather than from an arbitrary structure). To break the crystalline structure, a 20-ps high-temperature pulse of 510 K (but not 450 or 480 K) was effective. The system finally obtained is an all-atom model, with Ewald summation to evaluate Coulombic interactions and a constant surface tension of 35 dynes/cm/water-membrane interface, equilibrated for 12 ns (over 50 ns total calculation time), which reproduces all of the experimentally observed parameters examined in this work. Furthermore, this model shows the presence of significant orientational correlations between neighboring alkyl chains and between shoulder vectors (which show the orientations of the lipids about their long axes) of neighboring DMPCs. PMID- 11106618 TI - Use of a single glycine residue to determine the tilt and orientation of a transmembrane helix. A new structural label for infrared spectroscopy. AB - Site-directed dichroism is an emerging technique for the determination of membrane protein structure. However, due to a number of factors, among which is the high natural abundance of (13)C, the use of this technique has been restricted to the study of small peptides. We have overcome these problems through the use of a double C-deuterated glycine as a label. The modification of a single residue (Gly) in the transmembrane segment of M2, a protein from the Influenza A virus that forms H(+)-selective ion channels, has allowed us to determine its helix tilt and rotational orientation. Double C-deuteration shifts the antisymmetric and symmetric stretching vibrations of the CD(2) group in glycine to a transparent region of the infrared spectrum where the dichroic ratio of these bands can be measured. The two dichroisms, along with the helix amide I dichroic ratio, have been used to determine the helix tilt and rotational orientation of M2. The results are entirely consistent with previous site directed dichroism and solid-state NMR experiments, validating C-deuterated glycine (GlyCD(2)) as a structural probe that can now be used in the study of polytopic membrane proteins. PMID- 11106619 TI - Design of supported membranes tethered via metal-affinity ligand-receptor pairs. AB - Model lipid layers are very promising in investigating the complex network of recognition, transport and signaling processes at membranes. We have developed a novel and generic approach to create supported lipid membranes tethered by metal affinity binding. By self-assembly we have generated various interfaces that display histidine sequences (6xHis) via polymer spacers. These histidine functionalized interfaces are designed to allow specific docking and fusion of vesicles containing metal-chelating lipids. By means of surface plasmon resonance and atomic force microscopy we analyzed the formation and subsequently the structure of these solid-supported membranes. Although the affinity constant of single ligand-receptor pairs is only in the micromolar range, very stable immobilization of these membranes was observed. This behavior can be explained by multivalent interactions resembling many features of cell adhesion. The process is highly specific, because vesicle docking and bilayer formation are strictly dependent on the presence of metal-affinity ligand-receptor pairs. The surface accessibility and geometry of these tethered membranes were probed by binding of histidine-tagged polypeptides. The supported membranes show adsorption kinetics and values similar to planar supported monolayers. Using various combinations of metal-chelating and histidine-tagged lipids or thiols these metal-affinity tethered membranes should make a great impact on probing and eventually understanding the dynamic dialog of reconstituted membrane proteins. PMID- 11106620 TI - From liposomes to supported, planar bilayer structures on hydrophilic and hydrophobic surfaces: an atomic force microscopy study. AB - The sequence of events involved in the transition from attached liposomes to bilayer patches on hydrophilic and hydrophobic solid supports were visualized in situ by Tapping Mode atomic force microscopy in liquid. In a smooth manner, the attached liposomes spread and flattened from the outer edges toward the center until the two membrane bilayers were stacked on top of each other. The top bilayer then either rolls or slides over the bottom bilayer, and the adjacent edges join to form a larger membrane patch. This is clearly visible from the apparent height of 6.0-7.5 nm of the single bilayer, measured in situ. The addition of calcium appeared to increase the rate of the processes preventing the visualization of the intermediate stages. The same intermediate steps appeared to be present on hydrophobic surfaces, although the attached liposomes seemed to be distorted and the resultant membrane edges were uneven. This work has provided visual and detailed information on liposome coalescence (fusion) onto solid supports and demonstrated how the atomic force microscope can be used to study the process. PMID- 11106621 TI - The role of surfactant proteins in DPPC enrichment of surface films. AB - A pressure-driven captive bubble surfactometer was used to determine the role of surfactant proteins in refinement of the surface film. The advantage of this apparatus is that surface films can be spread at the interface of an air bubble with a different lipid/protein composition than the subphase vesicles. Using different combinations of subphase vesicles and spread surface films a clear correlation between dipalmitoylphosphatidylcholine (DPPC) content and minimum surface tension was observed. Spread phospholipid films containing 50% DPPC over a subphase containing 50% DPPC vesicles did not form stable surface films with a low minimum surface tension. Addition of surfactant protein B (SP-B) to the surface film led to a progressive decrease in minimum surface tension toward 1 mN/m upon cycling, indicating an enrichment in DPPC. Surfactant protein C (SP-C) had no such detectable refining effect on the film. Surfactant protein A (SP-A) had a positive effect on refinement when it was present in the subphase. However, this effect was only observed when SP-A was combined with SP-B and incubated with subphase vesicles before addition to the air bubble containing sample chamber. Comparison of spread films with adsorbed films indicated that refinement induced by SP-B occurs by selective removal of non-DPPC lipids upon cycling. SP-A, combined with SP-B, induces a selective adsorption of DPPC from subphase vesicles into the surface film. This is achieved by formation of large lipid structures which might resemble tubular myelin. PMID- 11106622 TI - Area per lipid and acyl length distributions in fluid phosphatidylcholines determined by (2)H NMR spectroscopy. AB - Deuterium ((2)H) NMR spectroscopy provides detailed information regarding the structural fluctuations of lipid bilayers, including both the equilibrium properties and dynamics. Experimental (2)H NMR measurements for the homologous series of 1, 2-diacyl-sn-glycero-3-phosphocholines with perdeuterated saturated chains (from C12:0 to C18:0) have been performed on randomly oriented, fully hydrated multilamellar samples. For each lipid, the C-D bond order parameters have been calculated from de-Paked (2)H NMR spectra as a function of temperature. The experimental order parameters were analyzed using a mean-torque potential model for the acyl chain segment distributions, and comparison was made with the conventional diamond lattice approach. Statistical mechanical principles were used to relate the measured order parameters to the lipid bilayer structural parameters: the hydrocarbon thickness and the mean interfacial area per lipid. At fixed temperature, the area decreases with increasing acyl length, indicating increased van der Waals attraction for longer lipid chains. However, the main effect of increasing the acyl chain length is on the hydrocarbon thickness rather than on the area per lipid. Expansion coefficients of the structural parameters are reported and interpreted using an empirical free energy function that describes the force balance in fluid bilayers. At the same absolute temperature, the phosphatidylcholine (PC) series exhibits a universal chain packing profile that differs from that of phosphatidylethanolamines (PE). Hence, the lateral packing of phospholipids is more sensitive to the headgroup methylation than to the acyl chain length. A fit to the area per lipid for the PC series using the empirical free energy function shows that the PE area represents a limiting value for the packing of fluid acyl chains. PMID- 11106623 TI - Electrostatic control of phospholipid polymorphism. AB - A regular progression of polymorphic phase behavior was observed for mixtures of the anionic phospholipid, cardiolipin, and the cationic phospholipid derivative, 1, 2-dioleoyl-sn-glycero-3-ethylphosphocholine. As revealed by freeze-fracture electron microscopy and small-angle x-ray diffraction, whereas the two lipids separately assume only lamellar phases, their mixtures exhibit a symmetrical (depending on charge ratio and not polarity) sequence of nonlamellar phases. The inverted hexagonal phase, H(II,) formed from equimolar mixtures of the two lipids, i.e., at net charge neutrality (charge ratio (CR((+/-))) = 1:1). When one type of lipid was in significant excess (CR((+/-)) = 2:1 or CR((+/-)) = 1:2), a bicontinuous cubic structure was observed. These cubic phases were very similar to those sometimes present in cellular organelles that contain cardiolipin. Increasing the excess of cationic or anionic charge to CR((+/-)) = 4:1 or CR((+/ )) = 1:4 led to the appearance of membrane bilayers with numerous interlamellar contacts, i.e., sponge structures. It is evident that interactions between cationic and anionic moieties can influence the packing of polar heads and hence control polymorphic phase transitions. The facile isothermal, polymorphic interconversion of these lipids may have important biological and technical implications. PMID- 11106624 TI - Molecular dynamics and (2)H-NMR study of the influence of an amphiphilic peptide on membrane order and dynamics. AB - A molecular dynamics simulation of a fully hydrated model membrane consisting of 12 molecules of 1, 2-dimyristoyl-sn-glycero-3-phosphocholine, one amphiphilic peptide with the sequence acetyl-Lys-Lys-Gly-Leu(16)-Lys-Lys-Ala-amide, and 593 water molecules was performed for 1.06 ns (Belohorcova, K., J. H. Davis, T. B. Woolf, and B. Roux. 1997. Biophys. J. 73:3039-3055). The analysis presented here is primarily focused on the phospholipid component and the results are compared with experimental (2)H-NMR studies of the lipid component of mixtures of the same peptide and lipid at a molar ratio of 1:32, and with earlier studies of closely related peptide/lipid mixtures. The phospholipid chain and headgroup isomer populations and isomerization rates compare favorably with previous simulations and experimental measurements. Of particular interest is the effect of the peptide on the phospholipid headgroup and hydrocarbon chain orientational order calculated from the simulation, which also agree well with experimental measurements performed on this and closely related systems. Comparison of the experimental results with the simulations not only shows that there is significant agreement between the two methods, but also provides new insight into the effect of the peptide on the lipid dynamics. In particular, these results confirm that a membrane spanning peptide has little effect on lipid chain order, and bilayer thickness if its hydrophobic length closely matches the lipid hydrocarbon thickness. In addition, we find that the peptide can have a strong ordering effect if it is longer than the lipid hydrophobic thickness. PMID- 11106625 TI - Alteration of tropomyosin function and folding by a nemaline myopathy-causing mutation. AB - Mutations in the human TPM3 gene encoding gamma-tropomyosin (alpha-tropomyosin slow) expressed in slow skeletal muscle fibers cause nemaline myopathy. Nemaline myopathy is a rare, clinically heterogeneous congenital skeletal muscle disease with associated muscle weakness, characterized by the presence of nemaline rods in muscle fibers. In one missense mutation the codon corresponding to Met-8, a highly conserved residue, is changed to Arg. Here, a rat fast alpha-tropomyosin cDNA with the Met8Arg mutation was expressed in Escherichia coli to investigate the effect of the mutation on in vitro function. The Met8Arg mutation reduces tropomyosin affinity for regulated actin 30- to 100-fold. Ca(2+)-sensitive regulatory function is retained, although activation of the actomyosin S1 ATPase in the presence of Ca(2+) is reduced. The poor activation may reflect weakened actin affinity or reduced effectiveness in switching the thin filament to the open, force-producing state. The presence of the Met8Arg mutation severely, but locally, destabilizes the tropomyosin coiled coil in a model peptide, and would be expected to impair end-to-end association between TMs on the thin filament. In muscle, the mutation may alter thin filament assembly consequent to lower actin affinity and altered binding of the N-terminus to tropomodulin at the pointed end of the filament. The mutation may also reduce force generation during activation. PMID- 11106626 TI - Extensibility of isoforms of cardiac titin: variation in contour length of molecular subsegments provides a basis for cellular passive stiffness diversity. AB - Titin is a giant polypeptide that spans between the Z- and M-lines of the cardiac muscle sarcomere and that develops force when extended. This force arises from titin's extensible I-band region, which consists mainly of three segment types: serially linked immunoglobulin-like domains (Ig segments), interrupted by the PEVK segment, and the N2B unique sequence. Recently it was reported that the myocardium of large mammals co-expresses small (N2B) and large (N2BA) cardiac isoforms and that the passive stiffness of cardiac myocytes varies with the isoform expression ratio. To understand the molecular basis of the differences in passive stiffness we investigated titin's extensibility in bovine atrium, which expresses predominantly N2BA titin, and compared it to that of rat, which expresses predominantly N2B titin. Immunoelectron microscopy was used with antibodies that flank the Ig segments, the PEVK segment, and the unique sequence of the N2B element. The extension of the various segments was then determined as a function of sarcomere length (SL). When slack sarcomeres of bovine atrium were stretched, the PEVK segment extended much more steeply and the unique N2B sequence less steeply than in rat, while the Ig segments behaved similarly in both species. However, the extensions normalized with the segment's contour length (i.e., the fractional extensions) of Ig, PEVK, and unique sequence segments all increase less steeply with SL in cow than in rat. Considering that fractional extension determines the level of entropic force, these differences in fractional extension are expected to result in shallow and steep passive force-SL curves in myocytes that express high levels of N2BA and N2B titin, respectively. Thus, the findings provide a molecular basis for passive stiffness diversity. PMID- 11106627 TI - Thermal behavior of long wavelength absorption transitions in Spirulina platensis photosystem I trimers. AB - In photosystem I trimers of Spirulina platensis a major long wavelength transition is irreversibly bleached by illumination with high-intensity white light. The photobleaching hole, identified by both absorption and circular dichroism spectroscopies, is interpreted as the inhomogeneously broadened Q(y) transition of a chlorophyll form that absorbs maximally near 709 nm at room temperature. Analysis of the mean square deviation of the photobleaching hole between 80 and 300 K, in the linear electron-phonon frame, indicates that the optical reorganization energy is 52 cm(-1), four times greater than that for the bulk, short-wavelength-absorbing chlorophylls, and the inhomogenous site distribution bandwidth is close to 150 cm(-1). The room temperature bandwidth, close to 18.5 nm, is dominated by thermal (homogeneous) broadening. Photobleaching induces correlated circular dichroism changes, of opposite sign, at 709 and 670 nm, which suggests that the long wavelength transition may be a low energy excitonic band, in agreement with its high reorganization energy. Clear identification of the 709-nm spectral form was used in developing a Gaussian description of the long wavelength absorption tail by analyzing the changing band shape during photobleaching using a global decomposition procedure. Additional absorption states near 720, 733, and 743 nm were thus identified. The lowest energy state at 743 nm is present in substoichiometric levels at room temperature and its presence was confirmed by fluorescence spectroscopy. This state displays an unusual increase in intensity upon lowering the temperature, which is successfully described by assuming the presence of low-lying, thermally populated states. PMID- 11106628 TI - The dynamics of protein hydration water: a quantitative comparison of molecular dynamics simulations and neutron-scattering experiments. AB - We present results from an extensive molecular dynamics simulation study of water hydrating the protein Ribonuclease A, at a series of temperatures in cluster, crystal, and powder environments. The dynamics of protein hydration water appear to be very similar in crystal and powder environments at moderate to high hydration levels. Thus, we contend that experiments performed on powder samples are appropriate for discussing hydration water dynamics in native protein environments. Our analysis reveals that simulations performed on cluster models consisting of proteins surrounded by a finite water shell with free boundaries are not appropriate for the study of the solvent dynamics. Detailed comparison to available x-ray diffraction and inelastic neutron-scattering data shows that current generation force fields are capable of accurately reproducing the structural and dynamical observables. On the time scale of tens of picoseconds, at room temperature and high hydration, significant water translational diffusion and rotational motion occur. At low hydration, the water molecules are translationally confined but display appreciable rotational motion. Below the protein dynamical transition temperature, both translational and rotational motions of the water molecules are essentially arrested. Taken together, these results suggest that water translational motion is necessary for the structural relaxation that permits anharmonic and diffusive motions in proteins. Furthermore, it appears that the exchange of protein-water hydrogen bonds by water rotational/librational motion is not sufficient to permit protein structural relaxation. Rather, the complete exchange of protein-bound water molecules by translational displacement seems to be required. PMID- 11106629 TI - The mechanics of F-actin microenvironments depend on the chemistry of probing surfaces. AB - To understand the microscopic mechanical properties of actin networks, we monitor the motion of embedded particles with controlled surface properties. The highly resolved Brownian motions of these particles reveal the viscoelastic character of the microenvironments around them. In both non-cross-linked and highly cross linked actin networks, particles that bind F-actin report viscoelastic moduli comparable to those determined by macroscopic rheology experiments. By contrast, particles modified to prevent actin binding have weak microenvironments that are surprisingly insensitive to the introduction of filament cross-links. Even when adjacent in the same cross-linked gel, actin-binding and nonbinding particles report viscoelastic moduli that differ by two orders of magnitude at low frequencies (0.5-1.5 rad/s) but converge at high frequencies (> 10(4) rad/s). For all particle chemistries, electron and light microscopies show no F-actin recruitment or depletion, so F-actin microheterogeneities cannot explain the deep penetration (approximately 100 nm) of nonbinding particles. Instead, we hypothesize that a local depletion of cross-linking around nonbinding particles explains the phenomena. With implications for organelle mobility in cells, our results show that actin binding is required for microenvironments to reflect macroscopic properties, and conversely, releasing actin enhances particle mobility beyond the effects of mere biochemical untethering. PMID- 11106630 TI - Biomolecular interactions measured by atomic force microscopy. AB - Atomic force microscopy (AFM) is nowadays frequently applied to determine interaction forces between biological molecules. Starting with the detection of the first discrete unbinding forces between ligands and receptors by AFM only several years ago, measurements have become more and more quantitative. At the same time, theories have been developed to describe and understand the dynamics of the unbinding process and experimental techniques have been refined to verify this theory. In addition, the detection of molecular recognition forces has been exploited to map and image the location of binding sites. In this review we discuss the important contributions that have led to the development of this field. In addition, we emphasize the potential of chemically well-defined surface modification techniques to further improve reproducible measurements by AFM. This increased reproducibility will pave the way for a better understanding of molecular interactions in cell biology. PMID- 11106632 TI - Fluorescence correlation spectroscopy in small cytosolic compartments depends critically on the diffusion model used. AB - Fluorescence correlation spectroscopy (FCS) is a powerful technique for measuring low concentrations of fluorescent molecules and their diffusion constants. In the standard case, fluorescence fluctuations are measured in an open detection volume defined by the confocal optics. However, if FCS measurements are carried out in cellular processes that confine the detection volume, the standard FCS model leads to erroneous results. In this paper, we derive a modified FCS model that takes into account the confinement of the detection volume. Using this model, we have carried out the first FCS measurements in dendrites of cultured neurons. We further derive, for the case of confined diffusion, the limits within which the standard two- and three-dimensional diffusion models give reliable results. PMID- 11106631 TI - Ultrastructural organization of amyloid fibrils by atomic force microscopy. AB - Atomic force microscopy has been employed to investigate the structural organization of amyloid fibrils produced in vitro from three very different polypeptide sequences. The systems investigated are a 10-residue peptide derived from the sequence of transthyretin, the 90-residue SH3 domain of bovine phosphatidylinositol-3'-kinase, and human wild-type lysozyme, a 130-residue protein containing four disulfide bridges. The results demonstrate distinct similarities between the structures formed by the different classes of fibrils despite the contrasting nature of the polypeptide species involved. SH3 and lysozyme fibrils consist typically of four protofilaments, exhibiting a left handed twist along the fibril axis. The substructure of TTR(10-19) fibrils is not resolved by atomic force microscopy and their uniform appearance is suggestive of a regular self-association of very thin filaments. We propose that the exact number and orientation of protofilaments within amyloid fibrils is dictated by packing of the regions of the polypeptide chains that are not directly involved in formation of the cross-beta core of the fibrils. The results obtained for these proteins, none of which is directly associated with any human disease, are closely similar to those of disease-related amyloid fibrils, supporting the concept that amyloid is a generic structure of polypeptide chains. The detailed architecture of an individual fibril, however, depends on the manner in which the protofilaments assemble into the fibrillar structure, which in turn is dependent on the sequence of the polypeptide and the conditions under which the fibril is formed. PMID- 11106633 TI - Direct observation of the transition from calcite to aragonite growth as induced by abalone shell proteins. AB - The mixture of EDTA-soluble proteins found in abalone nacre are known to cause the nucleation and growth of aragonite on calcite seed crystals in supersaturated solutions of calcium carbonate. Past atomic force microscope studies of the interaction of these proteins with calcite crystals did not observe this transition because no information about the crystal polymorph on the surface was obtained. Here we have used the atomic force microscope to directly observe changes in the atomic lattice on a calcite seed crystal after the introduction of abalone shell proteins. The observed changes are consistent with a transition to (001) aragonite growth on a (1014) calcite surface. PMID- 11106634 TI - Solution conformations of a trimannoside from nuclear magnetic resonance and molecular dynamics simulations. AB - N-linked oligosaccharides often act as ligands for receptor proteins in a variety of cell recognition processes. Knowledge of the solution conformations, as well as protein-bound conformations, of these oligosaccharides is required to understand these important interactions. In this paper we present a model for the solution conformations sampled by a simple trimannoside, methyl 3, 6-di-O-(alpha D-mannopyranosyl)-alpha-D-mannopyranoside, which contains two of the most commonly found glycosidic linkages in N-linked oligosaccharides. This model was derived from simulated annealing protocols incorporating distance restraints extracted from NOESY spectra along with torsional restraints computed from three bond (1)H-(13)C coupling constants measured across the glycosidic bonds. The model was refined in light of unrestrained molecular dynamics simulations conducted in the presence of solvent water. The resulting model depicts a molecule undergoing conformational averaging in solution, adopting four major and two minor conformations. The four major conformations arise from a pair of two state transitions, one each at the alpha(1-->3) and alpha(1-->6) linkages, whereas the minor conformations result from an additional transition of the alpha(1-->6) linkage. Our data also suggest that the alpha(1-->3) transition is fast and changes the molecular shape slightly, whereas the alpha(1-->6) is much slower and alters the molecular shape dramatically. PMID- 11106635 TI - First-principles determination of hybrid bilayer membrane structure by phase sensitive neutron reflectometry. AB - The application of a new, phase-sensitive neutron reflectometry method to reveal the compositional depth profiles of biomimetic membranes is reported. Determination of the complex reflection amplitude allows the related scattering length density (SLD) profile to be obtained by a first-principles inversion without the need for fitting or adjustable parameters. The SLD profile so obtained is unique for most membranes and can therefore be directly compared with the SLD profile corresponding to the chemical compositional profile of the film, as predicted, for example, by a molecular dynamics simulation. Knowledge of the real part of the reflection amplitude, in addition to enabling the inversion, makes it possible to assign a spatial resolution to the profile for a given range of wavevector transfer over which the reflectivity data are collected. Furthermore, the imaginary part of the reflection amplitude can be used as a sensitive diagnostic tool for recognizing the existence of certain in-plane inhomogeneities in the sample. Measurements demonstrating the practical realization of this phase-sensitive technique were performed on a hybrid bilayer membrane (self-assembled monolayer of thiahexa (ethylene oxide) alkane on gold and a phospholipid layer) in intimate contact with an aqueous reservoir. Analysis of the experimental results shows that accurate compositional depth profiles can now be obtained with a spatial resolution in the subnanometer range, primarily limited by the background originating from the reservoir and the roughness of the film's supporting substrate. PMID- 11106636 TI - Development of separable electron spin resonance-computed tomography imaging for multiple radical species: an application to .OH and .NO. AB - A method of separable ESR-CT (electron spin resonance-computed tomography) imaging for multiple radical species was developed and applied to imaging of .OH and .NO. The algorithm was improved by combining filtered back-projection with a modified algebraic reconstruction technique to enhance accuracy and shorten calculation time. With this algorithm, spectral-spatial images of the phantom consisting of 3-carbamoyl-2,2,5,5,-tetramethylpyrrolidine-N-oxyl and 2-phenyl 4,4,5,5,-tetramethylimidazoline-3-oxide-1-oxyl could be obtained in different directions by rotating the spatial axis. The spatial function of individual radicals was extracted by each of the two methods from each spectral-spatial image. The separative 2D images of each radical were individually constructed using the spatial function obtained with the two methods. By comparing the separative images with the phantom sample, the algorithm for separable ESR-CT imaging was established. This ESR-CT technique was combined with L-band ESR spectroscopy and applied to the separative imaging of .OH and .NO, which were spin trapped with 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) and Fe(2+)-N-methyl-D glucamine dithiocarbamate complex, respectively. The ESR signal of DMPO-OH decreased gradually during data acquisition, and the decrease was calibrated by extrapolating the signal intensity to the beginning of data sampling. Both the position and size of the individual images for .OH and .NO were in very good agreement with the findings for the sample. PMID- 11106637 TI - Mapping of glucocorticoid receptor DNA binding domain surfaces contributing to transrepression of NF-kappa B and induction of apoptosis. AB - Glucocorticoids (GCs) function, in part, through the ability of the glucocorticoid receptor (GR) to activate gene expression and in part through the transrepression of AP-1 and NF-kappaB. Here we characterize the effect of GR DNA binding domain (DBD) mutations, previously analyzed for changes in the ability to activate gene expression or transrepress AP-1. We have identified a GR mutant capable of distinguishing between transrepression of NF-kappaB and AP-1. Using circular dichroism spectroscopy, we show that this mutation does not appreciably alter GR DBD conformation, suggesting that functional differences between the mutant and wild type protein are the result of an alteration of a specific interaction surface. These data suggest that transrepression of NF-kappaB and AP 1 occurs through distinct protein-protein interactions and argue against the hypothesis that transrepression occurs through competition for a single coactivator protein. Introduction of these mutations into GC-resistant CEM lymphoblastic T cells restored dexamethasone (DEX)-mediated apoptosis as did wild type GR regardless of whether these mutants were transrepression or activation defective. Thus, DEX-mediated apoptosis in transformed T cells is more complex than originally appreciated. PMID- 11106638 TI - Lack of a role of the interferon-stimulated response element-like region in interferon alpha -induced suppression of Hepatitis B virus in vitro. AB - The antiviral effect of interferon-alpha (IFNalpha) on hepatitis B virus (HBV) is well documented in vitro and in vivo, but the mechanisms involved are elusive. Recently, an interferon-stimulated response like element (ISRE) competent for binding of interferon-stimulated gene factor-3gamma (p48) has been identified in the HBV enhancer I region. Mutation of this element was shown to abrogate IFNalpha-mediated reduction of HBV X-gene promoter-driven reporter gene expression. This suggested a role of the ISRE and of p48 in IFNalpha-induced antiviral activity against productive HBV infection. Here, we analyzed the antiviral effect of both IFNalpha and enhanced p48 expression on complete HBV genomes containing the wild-type or mutated ISRE. In human hepatoma cells transfected with both genomes, viral RNA and replicative intermediates were reduced by IFNalpha treatment to a similar degree. Enhanced p48 expression increased IFNalpha-induced suppression of HBV RNA significantly from 75 +/- 22.5% to 46 +/- 9.8%, but this was independent of the integrity of the ISRE-like region. These data imply that p48 neither mediates the antiviral activity of IFNalpha against HBV nor down-regulates enhancer I activity by binding directly to the HBV ISRE-like region, but rather argue for an indirect role of p48. PMID- 11106639 TI - Crucial role of the high-loop lysine for the catalytic activity of arginyl-tRNA synthetase. AB - The presence of two short signature sequence motifs (His-Ile-Gly-His (HIGH) and Lys-Met-Ser-Lys (KMSK)) is a characteristic of the class I aminoacyl-tRNA synthetases. These motifs constitute a portion of the catalytic site in three dimensions and play an important role in catalysis. In particular, the second lysine of the KMSK motif (K2) is the crucial catalytic residue for stabilization of the transition state of the amino acid activation reaction (aminoacyl adenylate formation). Arginyl-tRNA synthetase (ArgRS) is unique among all of the class I enyzmes, as the majority of ArgRS species lack canonical KMSK sequences. Thus, the mechanism by which this group of ArgRSs achieves the catalytic reaction is not well understood. Using three-dimensional modeling in combination with sequence analysis and site-directed mutagenesis, we found a conserved lysine in the KMSK-lacking ArgRSs upstream of the HIGH sequence motif, which is likely to be a functional counterpart of the canonical class I K2 lysine. The results suggest a plausible partition of the ArgRSs into two major groups, on the basis of the conservation of the HIGH lysine. PMID- 11106640 TI - COUP-TF1 antagonizes Nkx2.5-mediated activation of the calreticulin gene during cardiac development. AB - Calreticulin, a Ca(2+) binding chaperone of the endoplasmic reticulum, is also highly expressed in the embryonic heart, and knockout of the calreticulin gene is lethal during embryogenesis because of impaired cardiac development. The protein is down-regulated after birth, and elevated expression of calreticulin in newborn hearts is associated with severe cardiac pathology and death. Here we show that the transcription factor Nkx2.5 activates expression of the calreticulin gene in the heart. Binding of chicken ovalbumin upstream promoter-transcription factor 1 to the Nkx2.5 binding site suppresses transcription from the calreticulin promoter. Nkx2.5 and chicken ovalbumin upstream promoter-transcription factor 1 play antagonistic roles in regulating the expression of calreticulin during cardiac development. These studies indicate that cardiac-specific transcription factor Nkx2.5 plays a central role in activating calreticulin expression and that there is a cooperation between chicken ovalbumin upstream promoter-transcription factor 1 and Nkx2.5 at the calreticulin promoter. PMID- 11106641 TI - DjlA is a third DnaK co-chaperone of Escherichia coli, and DjlA-mediated induction of colanic acid capsule requires DjlA-DnaK interaction. AB - DjlA is a 30-kDa type III membrane protein of Escherichia coli with the majority, including an extreme C-terminal putative J-domain, oriented toward the cytoplasm. No other regions of sequence similarity aside from the J-domain exist between DjlA and the known DnaK (Hsp70) co-chaperones DnaJ (Hsp40) and CbpA. In this study, we explored whether and to what extent DjlA possesses DnaK co-chaperone activity and under what conditions a DjlA-DnaK interaction could be important to the cell. We found that the DjlA J-domain can substitute fully for the J-domain of DnaJ using various in vivo functional complementation assays. In addition, the purified cytoplasmic fragment of DjlA was shown to be capable of stimulating DnaK ATPase in a manner indistinguishable from DnaJ, and, furthermore, DjlA could act as a DnaK co-chaperone in the reactivation of chemically denatured luciferase in vitro. DjlA expression in the cell is tightly controlled, and even its mild overexpression leads to induction of mucoid capsule. Previous analysis showed that DjlA-mediated induction of the wca capsule operon required the RcsC/RcsB two component signaling system and that wca induction by DjlA was lost when cells contained mutations in either the dnaK or grpE gene. We now show using allele specific genetic suppression analysis that DjlA must interact with DnaK for DjlA mediated stimulation of capsule synthesis. Collectively, these results demonstrate that DjlA is a co-chaperone for DnaK and that this chaperone-co chaperone pair is implicated directly, or indirectly, in the regulation of colanic acid capsule. PMID- 11106642 TI - Escherichia coli MutS,L modulate RuvAB-dependent branch migration between diverged DNA. AB - This study examines the interaction between Escherichia coli MutS,L and E. coli RuvAB during E. coli RecA-promoted strand exchange. RuvAB is a branch migration complex that stimulates heterologous strand exchange. Previous studies indicate that RuvAB increases the rate at which heteroduplex products are formed by RecA, that RuvA and RuvB are required for this stimulation, and that RuvAB does not stimulate homologous strand exchange. This study indicates that MutS,L inhibit the formation of full-length heteroduplex DNA between M13-fd DNA in the presence of RuvAB, such that less than 2% of the linear substrate is converted to product. Inhibition depends on the time at which MutS,L are added to the reaction and is strongest when MutS,L are added during initiation. The kinetics of the strand exchange reaction suggest that MutS,L directly inhibit RuvAB-dependent branch migration in the absence of RecA. The inhibition requires the formation of base base mismatches and ATP utilization; no effect on RuvAB-promoted strand exchange is seen if an ATP-deficient mutant of MutS (MutS501) is included in the reaction instead of wild-type MutS. These results are consistent with a role for MutS,L in maintaining genomic stability and replication fidelity. PMID- 11106643 TI - Repression of human reduced folate carrier gene expression by wild type p53. AB - The relationship between loss of functional p53 and human reduced folate carrier (hRFC) levels and function was examined in REH lymphoblastic leukemia cells, which express wild type p53, and in p53-null K562 cells (K562(pTet-on/p53)) engineered to express wild type p53 under control of a tetracycline-inducible promoter. Activation of p53 in REH cells by treatment with daunorubicin was accompanied by decreased ( approximately 5-fold) levels of hRFC transcripts and methotrexate transport. Treatment of K562(pTet-on/p53) cells with doxycycline resulted in a dose-dependent expression of p53 protein and transcripts, increased p21 protein, decreased dihydrofolate reductase, and G(1) arrest with decreased numbers of cells in S-phase. p53 induction was accompanied by up to 3-fold decreases in hRFC transcripts transcribed from the upstream hRFC-B promoter and similar losses of hRFC protein and methotrexate uptake capacity. Expression of p15 in an analogous inducible system in K562 cells resulted in a nearly identical decrease of S-phase cells and dihydrofolate reductase without effects on hRFC levels or activity. When the hRFC-B promoter was expressed as full-length and basal promoter-luciferase reporter constructs in K562(pTet-on/p53) cells, induction of p53 with doxycycline resulted in a 3-fold loss of promoter activity, which was reversed by cotransfection with a trans-dominant-negative p53. These studies show that wild type p53 acts as a repressor of hRFC gene expression, via a mechanism that is independent of its effects on cell cycle progression. PMID- 11106644 TI - Physiochemical properties of rat liver mitochondrial ribosomes. AB - In the present study, the physiochemical properties of rat liver mitochondrial ribosomes were examined and compared with Escherichia coli ribosomes. The sedimentation and translational diffusion coefficients as well as the molecular weight and buoyant density of rat mitochondrial ribosomes were determined. Sedimentation coefficients were established using the time-derivative algorithm (Philo, J. S. (2000) Anal. Biochem. 279, 151-163). The sedimentation coefficients of the intact monosome, large subunit, and small subunit were 55, 39, and 28 S, respectively. Mitochondrial ribosomes had a particle composition of 75% protein and 25% RNA. The partial specific volume was 0.688 ml/g, as determined from the protein and RNA composition. The buoyant density of formaldehyde-fixed ribosomes in cesium chloride was 1.41 g/cm(3). The molecular masses of mitochondrial and E. coli ribosomes determined by static light-scattering experiments were 3.57 +/- 0.14 MDa and 2.49 +/- 0.06 MDa, respectively. The diffusion coefficient obtained from dynamic light-scattering measurements was 1.10 +/- 0.01 x 10(-7) cm(2) s(-1) for mitochondrial ribosomes and 1.72 +/- 0.03 x 10(-7) cm(2) s(-1) for the 70 S E. coli monosome. The hydration factor determined from these hydrodynamic parameters were 4.6 g of water/g of ribosome and 1.3 g/g for mitochondrial and E. coli ribosomes, respectively. A calculated hydration factor of 3.3 g/g for mitochondrial ribosomes was also obtained utilizing a calculated molecular mass and the Svedberg equation. These measurements of solvation suggest that ribosomes are highly hydrated structures. They are also in agreement with current models depicting ribosomes as porous structures containing numerous gaps and tunnels. PMID- 11106645 TI - Cbfa1 contributes to the osteoblast-specific expression of type I collagen genes. AB - Type I collagen is composed of two chains, alpha1(I) and alpha2(I), encoded by two distinct genes, the alpha1(I) and alpha2(I) collagen genes, that are highly expressed in osteoblasts. In most physiological situations, alpha1(I) and alpha2(I) collagen expression is coregulated, suggesting that identical transcription factors control their expression. Here, we studied the role of Cbfa1, an osteoblast-specific transcription factor, in the control of alpha1(I) and alpha2(I) collagen expression in osteoblasts. A consensus Cbfa1-binding site, termed OSE2, is present at the same location in the alpha1(I) collagen promoter at approximately -1347 base pairs (bp) of the rat, mouse, and human genes. Cbfa1 can bind to this site, as demonstrated by electrophoretic mobility shift assay (EMSA) and supershift experiments using an anti-Cbfa1 antibody. Mutagenesis of the alpha1(I) collagen OSE2 at -1347 bp reduced the activity of a alpha1(I) collagen promoter fragment 2- to 3-fold. Moreover, multimers of this OSE2 at 1347bp confer osteoblast-specific activity to a minimum alpha1(I) collagen promoter fragment in DNA transfection experiments as well as in transgenic mice. An additional Cbfa1-binding element is present in the alpha1(I) collagen promoter of mouse, rat, and human at approximately position -372. This site binds Cbfa1 only weakly and does not act as a cis-acting activator of transcription when tested in DNA transfection experiments. Similar to alpha1(I) collagen, the mouse alpha2(I) collagen gene contains multiple OSE2 sites, of which one is conserved across multiple species. In EMSA, Cbfa1 binds to this site and multimers of this alpha2(I) OSE2 element confer osteoblast-specific activity to the minimum alpha1(I) collagen promoter in DNA transfection experiments. Thus, our results suggest that Cbfa1 is one of the positive regulators of the osteoblast-specific expression of both type I collagen genes. PMID- 11106646 TI - Hierarchy of merlin and ezrin N- and C-terminal domain interactions in homo- and heterotypic associations and their relationship to binding of scaffolding proteins EBP50 and E3KARP. AB - The neurofibromatosis 2 tumor suppressor gene product merlin has strong sequence identity to the ezrin-radixin-moesin (ERM) family over its approximately 300 residue N-terminal domain. ERM proteins are membrane cytoskeletal linkers that are negatively regulated by an intramolecular association between domains known as NH(2)- and COOH-ERM association domains (N- and C-ERMADs) that mask sites for binding membrane-associated proteins, such as EBP50 and E3KARP, and F-actin. Here we show that merlin has self-association regions analogous to the N- and C ERMADs. Moreover, the N-/C-ERMAD interaction in merlin is relatively weak and dynamic, and this property is reflected by the ability of full-length recombinant merlin to form homo-oligomers. Remarkably, the merlin C-ERMAD has a higher affinity for the N-ERMAD of ezrin than the N-ERMAD of merlin. Both the ezrin and merlin N-ERMAD bind EBP50. This interaction with the ezrin N-ERMAD can be inhibited by the presence of the ezrin C-ERMAD, whereas interaction with the merlin N-ERMAD is not inhibited by either C-ERMAD. E3KARP binds tightly to the ezrin N-ERMAD but has little affinity for the merlin N-ERMAD. The implications of these associations and the hierarchies of binding for the function and regulation of merlin and ERM proteins are discussed. PMID- 11106647 TI - The modulation of oxygen radical production by nitric oxide in mitochondria. AB - Biological systems that produce or are exposed to nitric oxide (NO radical) exhibit changes in the rate of oxygen free radical production. Considering that mitochondria are the main intracellular source of oxygen radicals, and based on the recently documented production of NO(radical) by intact mitochondria, we investigated whether NO(radical), produced by the mitochondrial nitric-oxide synthase, could affect the generation of oxygen radicals. Toward this end, changes in H(2)O(2) production by rat liver mitochondria were monitored at different rates of endogenous NO(radical) production. The observed changes in H(2)O(2) production indicated that NO(radical) affected the rate of oxygen radical production by modulating the rate of O(2) consumption at the cytochrome oxidase level. This mechanism was supported by these three experimental proofs: 1) the reciprocal correlation between H(2)O(2) production and respiratory rates under different conditions of NO(radical) production; 2) the pattern of oxidized/reduced carriers in the presence of NO(radical), which pointed to cytochrome oxidase as the crossover point; and 3) the reversibility of these effects, evidenced in the presence of oxymyoglobin, which excluded a significant role for other NO(radical)-derived species such as peroxynitrite. Other sources of H(2)O(2) investigated, such as the aerobic formation of nitrosoglutathione and the GSH-mediated decay of nitrosoglutathione, were found quantitatively negligible compared with the total rate of H(2)O(2) production. PMID- 11106648 TI - Essential tyrosine residues for interaction of the non-receptor protein-tyrosine phosphatase PTP1B with N-cadherin. AB - Expression of a dominant-negative, catalytically inactive form of the nonreceptor protein-tyrosine phosphatase PTP1B in L-cells constitutively expressing N cadherin results in loss of N-cadherin-mediated cell-cell adhesion. PTP1B interacts directly with the cytoplasmic domain of N-cadherin, and this association is regulated by phosphorylation of tyrosine residues in PTP1B. The following three tyrosine residues in PTP1B are potential substrates for tyrosine kinases: Tyr-66, Tyr-152, and Tyr-153. To determine the tyrosine residue(s) that are crucial for the cadherin-PTP1B interaction we used site-directed mutagenesis to create catalytically inactive PTP1B constructs bearing additional single, double, or triple mutations in which tyrosine was substituted by phenylalanine. Mutation Y152F eliminates binding to N-cadherin in vitro, whereas mutations Y66F and Y153F do not. Overexpression of the catalytically inactive PTP1B with the Y152F mutation in L-cells constitutively expressing N-cadherin has no effect on N cadherin-mediated adhesion, and immunoprecipitation reveals that the mutant Y152F PTP1B does not associate with N-cadherin in situ. Furthermore, among cells overexpressing the Y152F mutant endogenous PTP1B associates with N-cadherin and is tyrosine-phosphorylated. PMID- 11106649 TI - Distinct arachidonate-releasing functions of mammalian secreted phospholipase A2s in human embryonic kidney 293 and rat mastocytoma RBL-2H3 cells through heparan sulfate shuttling and external plasma membrane mechanisms. AB - We analyzed the ability of a diverse set of mammalian secreted phospholipase A(2) (sPLA(2)) to release arachidonate for lipid mediator generation in two transfected cell lines. In human embryonic kidney 293 cells, the heparin-binding enzymes sPLA(2)-IIA, -IID, and -V promote stimulus-dependent arachidonic acid release and prostaglandin E(2) production in a manner dependent on the heparan sulfate proteoglycan glypican. In contrast, sPLA(2)-IB, -IIC, and -IIE, which bind weakly or not at all to heparanoids, fail to elicit arachidonate release, and addition of a heparin binding site to sPLA(2)-IIC allows it to release arachidonate. Heparin nonbinding sPLA(2)-X liberates arachidonic acid most likely from the phosphatidylcholine-rich outer plasma membrane in a glypican-independent manner. In rat mastocytoma RBL-2H3 cells that lack glypican, sPLA(2)-V and -X, which are unique among sPLA(2)s in being able to hydrolyze phosphatidylcholine rich membranes, act most likely on the extracellular face of the plasma membrane to markedly augment IgE-dependent immediate production of leukotriene C(4) and platelet-activating factor. sPLA(2)-IB, -IIA, -IIC, -IID, and -IIE exert minimal effects in RBL-2H3 cells. These results are also supported by studies with sPLA(2) mutants and immunocytostaining and reveal that sPLA(2)-dependent lipid mediator generation occur by distinct (heparanoid-dependent and -independent) mechanisms in HEK293 and RBL-2H3 cells. PMID- 11106650 TI - Interferon-gamma-induced regulation of peroxisome proliferator-activated receptor gamma and STATs in adipocytes. AB - Interferon-gamma (IFN-gamma) is known primarily for its roles in immunological responses but also has been shown to affect fat metabolism and adipocyte gene expression. To further investigate the effects of IFN-gamma on fat cells, we examined the effects of this cytokine on the expression of adipocyte transcription factors in 3T3-L1 adipocytes. Although IFN-gamma regulated the expression of several adipocyte transcription factors, IFN-gamma treatment resulted in a rapid reduction of both peroxisome proliferator-activated receptor (PPAR) protein and mRNA. A 48-h exposure to IFN-gamma also resulted in a decrease of both CCAAT/enhancer-binding alpha and sterol regulatory element binding protein (SREBP-1) expression. The short half-life of both the PPARgamma mRNA and protein likely contributed to the rapid decline of both cytosolic and nuclear PPARgamma in the presence of IFN-gamma. Our studies clearly demonstrated that the IFN-gamma-induced loss of PPARgamma protein is partially inhibited in the presence of two distinct proteasome inhibitors. Moreover, IFN-gamma also inhibited the transcription of PPARgamma, which was accompanied by a decrease in PPARgamma mRNA accumulation. In addition, exposure to IFN-gamma resulted in a substantial increase in STAT 1 expression and a small increase in STAT 3 expression. IFN-gamma treatment of 3T3-L1 adipocytes (48-96 h) resulted in a substantial inhibition of insulin-sensitive glucose uptake. These data clearly demonstrate that IFN-gamma treatment results in the development of insulin resistance, which is accompanied by the regulation of various adipocyte transcription factors, in particular the synthesis and degradation of PPARgamma. PMID- 11106651 TI - The DNA binding properties of the parsley bZIP transcription factor CPRF4a are regulated by light. AB - The common plant regulatory factors (CPRFs) from parsley are transcription factors with a basic leucine zipper motif that bind to cis-regulatory elements frequently found in promoters of light-regulated genes. Recent studies have revealed that certain CPRF proteins are regulated in response to light by changes in their expression level and in their intracellular localization. Here, we describe an additional mechanism contributing to the light-dependent regulation of CPRF proteins. We show that the DNA binding activity of the factor CPRF4a is modulated in a phosphorylation-dependent manner and that cytosolic components are involved in the regulation of this process. Moreover, we have identified a cytosolic kinase responsible for CPRF4a phosphorylation. Modification of recombinant CPRF4a by this kinase, however, is insufficient to cause a full activation of the factor, suggesting that additional modifications are required. Furthermore, we demonstrate that the DNA binding activity of the factor is modified upon light treatment. The results of additional irradiation experiments suggest that this photoresponse is controlled by different photoreceptor systems. We discuss the possible role of CPRF4a in light signal transduction as well as the emerging regulatory network controlling CPRF activities in parsley. PMID- 11106652 TI - Inefficient bypass of an abasic site by DNA polymerase eta. AB - DNA polymerase eta (Pol eta) bypasses a cis-syn thymine-thymine dimer efficiently and accurately, and inactivation of Pol eta in humans results in the cancer-prone syndrome, the variant form of xeroderma pigmentosum. Also, Pol eta bypasses the 8 oxoguanine lesion efficiently by predominantly inserting a C opposite this lesion, and it bypasses the O(6)-methylguanine lesion by inserting a C or a T. To further assess the range of DNA lesions tolerated by Pol eta, here we examine the bypass of an abasic site, a prototypical noninstructional lesion. Steady-state kinetic analyses show that both yeast and human Pol eta are very inefficient in both inserting a nucleotide opposite an abasic site and in extending from the nucleotide inserted. Hence, Pol eta bypasses this lesion extremely poorly. These results suggest that Pol eta requires the presence of template bases opposite both the incoming nucleotide and the primer terminus to catalyze efficient nucleotide incorporation. PMID- 11106653 TI - Amyloid-beta interactions with chondroitin sulfate-derived monosaccharides and disaccharides. implications for drug development. AB - In Alzheimer's disease, the major pathological features are diffuse and senile plaques that are primarily composed of the amyloid-beta (A beta) peptide. It has been proposed that proteoglycans and glycosaminoglycans (GAG) facilitate amyloid fibril formation and/or stabilize the plaque aggregates. To develop effective therapeutics based on A beta-GAG interactions, understanding the A beta binding motif on the GAG chain is imperative. Using electron microscopy, fluorescence spectroscopy, and competitive inhibition ELISAs, we have evaluated the ability of chondroitin sulfate-derived monosaccharides and disaccharides to induce the structural changes in A beta that are associated with GAG interactions. Our results demonstrate that the disaccharides GalNAc-4-sulfate(4S), Delta UA-GalNAc 6-sulfate(6S), and Delta UA-GalNAc-4,6-sulfate(4S,6S), the iduronic acid-2 sulfate analogues, and the monosaccharides d-GalNAc-4S, d-GalNAc-6S, and d-GalNAc 4S,6S, but not d-GalNAc, d-GlcNAc, or Delta UA-GalNAc, induce the fibrillar features of A beta-GAG interactions. The binding affinities of all chondroitin sulfate-derived saccharides mimic those of the intact GAG chains. The sulfated monosaccharides and disaccharides compete with the intact chondroitin sulfate and heparin GAGs for A beta binding, as illustrated by competitive inhibition ELISAs. Therefore, the development of therapeutics based on the model of A beta chondroitin sulfate binding may lead to effective inhibitors of the GAG-induced amyloid formation that is observed in vitro. PMID- 11106654 TI - Interaction with p53 enhances binding of cisplatin-modified DNA by high mobility group 1 protein. AB - A nonhistone chromosomal protein, high mobility group (HMG) 1, is ubiquitous in higher eukaryotic cells and binds preferentially to cisplatin-modified DNA. HMG1 also functions as a coactivator of p53, a tumor suppressor protein. We investigated physical interactions between HMG1 and p53 and the influence of p53 on the ability of HMG1 to recognize damaged DNA. Using immunochemical coprecipitation, we observed binding of HMG1 and p53. Interaction between HMG1 and p53 required the HMG A box of HMG1 and amino acids 363-376 of p53. Cisplatin modified DNA binding by HMG1 was significantly enhanced by p53. An HMG1-specific antibody that recognized the A box of this protein also stimulated cisplatin modified DNA binding. These data suggest that an interaction with either p53 or antibody may induce conformational change in the HMG1 A box that optimizes DNA binding by HMG1. Interaction of p53 with HMG1 after DNA damage may promote activation of specific HMG1 binding to damaged DNA in vivo and provide a molecular link between DNA damage and p53-mediated DNA repair. PMID- 11106655 TI - Mechanisms underlying preferential assembly of heparan sulfate on glypican-1. AB - Glypicans are major cell surface heparan sulfate proteoglycans, the structures of which are characterized by the presence of a cysteine-rich globular domain, a short glycosaminoglycan (GAG) attachment region, and a glycosylphosphatidylinositol membrane anchor. Despite strong evolutionary conservation of the globular domains of glypicans, no function has yet been attributed to them. By using a novel quantitative approach for assessing proteoglycan glycosylation, we show here that removal of the globular domain from rat glypican-1 converts the proteoglycan from one that bears approximately 90% heparan sulfate (HS) to one that bears approximately 90% chondroitin sulfate. Mutational analysis shows that sequences at least 70 amino acids away from the glypican-1 GAG attachment site are required for preferential HS assembly, although more nearby sequences also play a role. The effects of the glypican-1 globular domain on HS assembly could also be demonstrated by fusing this domain to sequences representing the GAG attachment sites of other proteoglycans or, surprisingly, simply by expressing the isolated globular domain in cells and analyzing effects either on an exogenously expressed glypican-1 GAG attachment domain or on endogenous proteoglycans. Quantitative analysis of the effect of the globular domain on GAG addition to proteoglycan core proteins suggested that preferential HS assembly is achieved, at least in part, through the inhibition of chondroitin sulfate assembly. These data identify the glypican-1 globular domain as a structural motif that potently influences GAG class determination and suggest that an important role of glypican globular domains is to ensure a high level of HS substitution of these proteoglycans. PMID- 11106656 TI - The C-terminal tail of the metabotropic glutamate receptor subtype 7 is necessary but not sufficient for cell surface delivery and polarized targeting in neurons and epithelia. AB - Complex neuronal functions rely upon the precise sorting, targeting, and restriction of receptors to specific synaptic microdomains. Little is known, however, of the molecular signals responsible for mediating these selective distributions. Here we report that metabotropic glutamate receptor subtype 7a (mGluR7a) is polarized at the basolateral surface when expressed in Madin-Darby canine kidney (MDCK) epithelial cells but is not polarized when expressed in cultured hippocampal neurons. Truncation of the mGluR7 cytoplasmic tail produces a protein that is restricted to a perinuclear intracellular compartment in both neurons and MDCK cells, where this protein colocalizes with a trans-Golgi network antigen. The mGluR7 cytoplasmic domain appended to the transmembrane portion of the vesicular stomatitis virus G protein and the ectodomain of human placental alkaline phosphatase is distributed over the entire cell surface in cultured neurons. When expressed in MDCK cells, this construct remains in an intracellular compartment distinct from endosomes or lysosomes. Thus, the cytoplasmic tail domain of mGluR7 is necessary but not sufficient for polarized targeting in MDCK monolayers, whereas in neurons the cytoplasmic tail is sufficient for cell surface expression but not polarization. Additional mechanisms are likely required to mediate mGluR7 neuronal polarization and synaptic clustering. PMID- 11106657 TI - Differentiating embryonal stem cells are a rich source of haemopoietic gene products and suggest erythroid preconditioning of primitive haemopoietic stem cells. AB - The difficulties associated with studying molecular mechanisms important in hemopoietic stem cell (HSC) function such as the problems of purifying homogeneous stem cell populations, have prompted us to adapt the murine ES cell system as an in vitro model of HSC generation and function. We now report that careful analysis of the time course of HSC generation in differentiating ES cells allows them to be used as a source of known and novel hemopoietic gene products. We have generated a subtracted library using cDNA from ES cells collected just prior to and just following the emergence of HSCs. Analysis of this library shows it to be a rich source of known hemopoietic and hemopoietic related gene products with 44% of identifiable cDNAs falling into these camps. We have demonstrated the value of this system as a source of novel genes of relevance to HSC function by characterizing a novel membrane protein encoding cDNA that is preferentially expressed in primitive hemopoietic cells. Intriguingly, further analysis of the known components of the subtracted library is suggestive of erythroid preconditioning of the ES cell-derived HSC. We have used dot-blot and in situ analysis to indicate that this erythroid preconditioning is probably restricted to primitive but not definitive HSC. PMID- 11106658 TI - c-Jun NH2-terminal kinase activation leads to a FADD-dependent but Fas ligand independent cell death in Jurkat T cells. AB - Persistent c-Jun NH2-terminal kinase (JNK) activation induces cell death. Different mechanisms are ascribed to JNK-induced cell death. Most of the JNK apoptosis studies employ stress stimuli known to activate kinases other than JNK. Here we used overexpression of mitogen-activated protein kinase kinase 7 (MKK7) to activate selectively JNK in T lymphoma Jurkat cells. Similar to that reported previously, Fas ligand (FasL) expression was up-regulated by JNK activation. Dominant negative-FADD and caspase-8 inhibitor benzyloxycarbonyl-Ile-Glu-Thr-Asp effectively inhibited MKK7-induced cell death, supporting a major involvement of FADD cascade. However, MKK7-induced cell death was not prevented by antagonist antibody ZB4 and Fas-Fc, indicating that Fas-FasL interaction is minimally involved. Confocal microscopy revealed that persistent JNK activation led to clustering of Fas. Our results suggest that, in contrast to that reported previously, JNK alone-induced death in Jurkat cells is FADD-dependent but is not triggered by Fas-FasL interaction. PMID- 11106659 TI - Involvement of DNA topoisomerase IIbeta in neuronal differentiation. AB - Two isoforms of DNA topoisomerase II (topo II) have been identified in mammalian cells. While topo IIalpha is essential for chromosome segregation in mitotic cells, in vivo function of topo IIbeta remains to be clarified. Here we demonstrate that the nucleoplasmic topo IIbeta, highly expressed in differentiating cerebellar neurons, is the catalytically competent entity operating directly on chromatin DNA in vivo. When the cells reached terminal differentiation, this in vivo activity decreased to a negligible level with concomitant loss of the nucleoplasmic enzyme. Effects of topo II-specific inhibitors were analyzed in a primary culture of cerebellar granule neurons that can mimic the in vivo situation. Only the beta isoform was expressed in granule cells differentiating in vitro. ICRF-193, a catalytic topo II inhibitor, suppressed the transcriptional induction of amphiphysin I which is essential for mature neuronal activity. The effect decreased significantly as the cells differentiate. Expression profiling with a cDNA macroarray showed that 18% of detectable transcripts were up-regulated during the differentiation and one-third of them were susceptible to ICRF-193. The results suggest that topo IIbeta is involved in an early stage of granule cell differentiation by potentiating inducible neuronal genes to become transcribable probably through alterations in higher order chromatin structure. PMID- 11106660 TI - Evaluation of the mutagenic potential of the principal DNA adduct of acrolein. AB - Acrolein is produced extensively in the environment by incomplete combustion of organic materials, and it arises endogenously in humans as a metabolic by product. Acrolein reacts with DNA at guanine residues to form the exocyclic adduct, 8-hydroxypropanodeoxyguanosine (HOPdG). Acrolein is mutagenic, and a correlation exists between HOPdG levels in Salmonella typhimurium treated with acrolein and a resultant increase in mutation frequency. Site-specifically modified oligonucleotides were used to explore the mutagenic potential of HOPdG in Escherichia coli strains that were either wild-type for repair or deficient in nucleotide excision repair or base excision repair. Oligonucleotides modified with HOPdG were inserted into double-stranded bacteriophage vectors using the gapped-duplex method or into single-stranded bacteriophage vectors and transformed into SOS-induced E. coli strains. Progeny phage were analyzed by oligonucleotide hybridization to establish the mutation frequency and the spectrum of mutations produced by HOPdG. The correct base, dCMP, was incorporated opposite HOPdG in all circumstances tested. In contrast, in vitro lesion bypass studies showed that HOPdG causes misincorporation opposite the modified base and is a block to replication. The combination of these studies showed that HOPdG is not miscoding in vivo at the level of sensitivity of these site-specific mutagenesis assays. PMID- 11106662 TI - Quaternary structure of rice nonsymbiotic hemoglobin. AB - Plant nonsymbiotic hemoglobins are hexacoordinate heme proteins found in all plants. Although expression is linked with hypoxic environmental conditions (Taylor, E. R., Nie, X. Z., Alexander, W. M., and Hill, R. D. (1994) Plant Mol. Biol. 24, 853-862), no discrete physiological function has yet been attributed to this family of proteins. The crystal structure of a nonsymbiotic hemoglobin from rice has recently been determined. The crystalline protein is homodimeric and hexacoordinate with two histidine side chains coordinating the heme iron atom. Despite the fact that the amino acids responsible for the subunit interface are relatively conserved among the nonsymbiotic hemoglobins, previous work suggests that this group of proteins might display variability in quaternary structure (Duff, S. M. G., Wittenberg, J. B., and Hill, R. D. (1997) J. Biol. Chem. 272, 16746-16752; Arredondo-Peter, R., Hargrove, M. S., Sarath, G., Moran, J. F., Lohrman, J., Olson, J. S., and Klucas, R. V. (1997) Plant Physiol. 115, 1259 1266). Analytical ultracentrifugation and size exclusion high pressure liquid chromatography were used to investigate the quaternary structure of rice nonsymbiotic hemoglobin at various states of ligation and oxidation. Additionally, site-directed mutagenesis was used to test the role of several interface amino acids in dimer formation and ligand binding. Results were analyzed in light of possible physiological functions and indicate that the plant nonsymbiotic hemoglobins are not oxygen transport proteins but more closely resemble known oxygen sensors. PMID- 11106661 TI - WD repeat domains target dictyostelium myosin heavy chain kinases by binding directly to myosin filaments. AB - Myosin heavy chain kinase (MHCK) A phosphorylates mapped sites at the C-terminal tail of Dictyostelium myosin II heavy chain, driving disassembly of myosin filaments both in vitro and in vivo. MHCK A is organized into three functional domains that include an N-terminal coiled-coil region, a central kinase catalytic domain unrelated to conventional protein kinases, and a WD repeat domain at the C terminus. MHCK B is a homologue of MHCK A that possesses structurally related catalytic and WD repeat domains. In the current study, we explored the role of the WD repeat domains in defining the activities of both MHCK A and MHCK B using recombinant bacterially expressed truncations of these kinases either with or without their WD repeat domains. We demonstrate that substrate targeting is a conserved function of the WD repeat domains of both MHCK A and MHCK B and that this targeting is specific for Dictyostelium myosin II filaments. We also show that the mechanism of targeting involves direct binding of the WD repeat domains to the myosin substrate. To our knowledge, this is the first report of WD repeat domains physically targeting attached kinase domains to their substrates. The examples presented here may serve as a paradigm for enzyme targeting in other systems. PMID- 11106664 TI - T Cell Receptor Binding to a pMHCII Ligand Is Kinetically Distinct from and Independent of CD4. AB - Immune recognition of pMHCII ligands by a helper T lymphocyte involves its antigen-specific T cell receptor (TCR) and CD4 coreceptor. We have characterized the binding of both molecules to the same pMHCII. The D10 alphabeta TCR heterodimer binds to conalbumin/I-A(k) with virtually identical kinetics and affinity as the single chain ValphaVbeta domain module (scD10) (Kd = 6-8 microm). The CD4 ectodomain does not alter either interaction. Moreover, CD4 alone demonstrates weak pMHCII binding (Kd = 200 microm), with no discernable affinity for the alphabeta TCR heterodimer. Hence, rather than providing a major contribution to binding energy, the critical role for the coreceptor in antigen specific activation likely results from transient inducible recruitment of the CD4 cytoplasmic tail-associated lck tyrosine kinase to the pMHCII-ligated TCR complex. PMID- 11106663 TI - Substrate-binding clusters of the K+-transporting Kdp ATPase of Escherichia coli investigated by amber suppression scanning mutagenesis. AB - The Kdp-ATPase of Escherichia coli is a four-subunit P-type ATPase that accumulates K(+) with high affinity and specificity. Residues clustered in four regions of the KdpA subunit of Kdp were implicated as critical for K(+) binding from the analysis of mutants with reduced affinity for K(+) (Buurman, E., Kim, K. T., and Epstein, W. (1995) J. Biol. Chem. 270, 6678-6685). K(+) binding by this pump has been analyzed in detail by site-directed mutagenesis. We have examined 83 of the 557 residues in KdpA, from 11 to 34 residues in each of four binding clusters known to affect K(+) binding. Amber mutations were constructed in a plasmid carrying the kdpFABC structural genes. Transferring these plasmids to 12 suppressor strains, each inserting a different amino acid at amber codons, created 12 different substitutions at the mutated sites. This study delineates the four clusters and confirms that they are important for K(+) affinity but have little effect on the rate of transport. At only 21 of the residues studied did at least three substitutions alter affinity for K(+), an indication that a residue is in or very near a K(+) binding site. At many residues lysine was the only substitution that altered its affinity. The effect of lysine is most likely a repulsive effect of this cationic residue on K(+) and thus reflects the effective distance between a residue and the site of binding or passage of K(+) in KdpA. Once a crystallographic structure of Kdp is available, this measure of effective distance will help identify the path of K(+) as it moves through the KdpA subunit to cross the membrane. PMID- 11106665 TI - Characterization of transsulfuration and cysteine biosynthetic pathways in the protozoan hemoflagellate, Trypanosoma cruzi. Isolation and molecular characterization of cystathionine beta-synthase and serine acetyltransferase from Trypanosoma. AB - Sulfur-containing amino acids play an important role in a variety of cellular functions such as protein synthesis, methylation, and polyamine and glutathione synthesis. We cloned and characterized cDNA encoding cystathionine beta-synthase (CBS), which is a key enzyme of transsulfuration pathway, from a hemoflagellate protozoan parasite Trypanosoma cruzi. T. cruzi CBS, unlike mammalian CBS, lacks the regulatory carboxyl terminus, does not contain heme, and is not activated by S-adenosylmethionine. T. cruzi CBS mRNA is expressed as at least six independent isotypes with sequence microheterogeneity from tandemly linked multicopy genes. The enzyme forms a homotetramer and, in addition to CBS activity, the enzyme has serine sulfhydrylase and cysteine synthase (CS) activities in vitro. Expression of the T. cruzi CBS in Saccharomyces cerevisiae and Escherichia coli demonstrates that the CBS and CS activities are functional in vivo. Enzymatic studies on T. cruzi extracts indicate that there is an additional CS enzyme and stage-specific control of CBS and CS expression. We also cloned and characterized cDNA encoding serine acetyltransferase (SAT), a key enzyme in the sulfate assimilatory cysteine biosynthetic pathway. Dissimilar to bacterial and plant SAT, a recombinant T. cruzi SAT showed allosteric inhibition by l-cysteine, l-cystine, and, to a lesser extent, glutathione. Together, these studies demonstrate the T. cruzi is a unique protist in possessing both transsulfuration and sulfur assimilatory pathways. PMID- 11106666 TI - Both subunits of the dimeric plant photoreceptor phytochrome require chromophore for stability of the far-red light-absorbing form. AB - The dimeric plant photoreceptor phytochrome is converted from its inactive red light-absorbing form (Pr) into the active far-red light-absorbing form (Pfr) upon light absorption. Dynamics of Pfr generation and of thermal Pfr-to-Pr conversion are of fundamental importance for inducing adequate responses to light signals. Here, we analyzed the role of subunit interactions on spectroscopic properties of dimeric phytochrome A. Using a coexpression system and affinity chromatography, we prepared mixed phytochrome dimers that can incorporate the essential chromophore only in one subunit. We demonstrate that such mixed dimers have unaltered difference spectra. In contrast, dark reversion differed greatly between Pfr-Pfr homodimers and Pfr-Pr heterodimers, the former being about 100 fold more stable. Temperature dependence of reaction rates revealed an additional stabilization of about 4 kcal/mol in homodimers. Consequences of these findings are discussed in relation to the biological function of, and functional diversification between, phytochrome family members. PMID- 11106667 TI - Pet111p, an inner membrane-bound translational activator that limits expression of the Saccharomyces cerevisiae mitochondrial gene COX2. AB - The protein specified by the Saccharomyces cerevisiae nuclear gene PET111 specifically activates translation of the mitochondrially coded mRNA for cytochrome c oxidase subunit II (Cox2p). We found Pet111p specifically in mitochondria of both wild-type cells and cells expressing a chromosomal gene for a functional epitope-tagged form of Pet111p. Pet111p was associated with mitochondrial membranes and was highly resistant to extraction with alkaline carbonate. Pet111p was protected from proteolytic digestion by the mitochondrial inner membrane. Thus, it is exposed only on the matrix side, where it could participate directly in organellar translation and localize Cox2p synthesis by virtue of its functional interaction with the COX2 mRNA 5'-untranslated leader. We also found that Pet111p is present at levels limiting the synthesis of Cox2p by examining the effect of altered PET111 gene dosage in the nucleus on expression of a reporter gene, cox2::ARG8(m), that was inserted into mitochondrial DNA. The level of the reporter protein, Arg8p, was one-half that of wild type in a diploid strain heterozygous for a pet111 deletion mutation, whereas it was increased 2.8-fold in a strain bearing extra copies of PET111 on a high-copy plasmid. Thus, Pet111p could play dual roles in both membrane localization and regulation of Cox2p synthesis within mitochondria. PMID- 11106668 TI - c-IAP1 is cleaved by caspases to produce a proapoptotic C-terminal fragment. AB - Although human c-IAP1 and c-IAP2 have been reported to possess antiapoptotic activity against a variety of stimuli in several mammalian cell types, we observed that full-length c-IAP1 and c-IAP2 failed to protect cells from apoptosis induced by Bax overexpression, tumor necrosis factor alpha treatment or Sindbis virus infection. However, deletion of the C-terminal RING domains of c IAP1 and c-IAP2 restored antiapoptotic activity, indicating that this region negatively regulates the antiapoptotic function of the N-terminal BIR domain. This finding is consistent with the observation by others that the spacer region and RING domain of c-IAP1 functions as an E3 ligase, promoting autoubiquitination and degradation of c-IAP1. In addition, we found that c-IAP1 is cleaved during apoptosis to 52- and 35-kDa fragments. Both fragments contain the C-terminal end of c-IAP1 including the RING finger. In vitro cleavage of c-IAP1 with apoptotic cell extracts or with purified recombinant caspase-3 produced similar fragments. Furthermore, transfection of cells with the spacer-RING domain alone suppressed the antiapoptotic function of the N-terminal BIR domain of c-IAP1 and induced apoptosis. Optimal death-inducing activity of the spacer-RING required both the spacer region and the zinc-binding RING domain of c-IAP1 but did not require the caspase recruitment domain located within the spacer region. To the contrary, deletion of the caspase recruitment domain increased proapoptotic activity, apparently by stabilizing the C-terminal fragment. PMID- 11106669 TI - HER2 in prostate cancer--a viable target or innocent bystander? PMID- 11106670 TI - Preventing cancer by disrupting progression of precancerous lesions. PMID- 11106671 TI - Consensus panel endorses range of adjuvant therapies for breast cancer. PMID- 11106673 TI - For more information PMID- 11106672 TI - Flurry of activity follows report on quality of cancer care. PMID- 11106674 TI - Prostate cancer: numbers may not tell the whole story. PMID- 11106676 TI - Stat bite: prostate cancer incidence and mortality in U.S. blacks and whites, 1973-1997. PMID- 11106675 TI - New study results highlight role of radiation therapy in prostate cancer. PMID- 11106677 TI - Children's oncology group looks to increase efficiency, numbers in clinical trials. PMID- 11106678 TI - Targeted cancer therapies attempt to hit the bull's-eye. PMID- 11106679 TI - Chemoprevention of gastric dysplasia: randomized trial of antioxidant supplements and anti-helicobacter pylori therapy. AB - BACKGROUND: Previous research has identified a high risk of gastric carcinoma as well as a high prevalence of cancer precursor lesions in rural populations living in the province of Narino, Colombia, in the Andes Mountains. METHODS: A randomized, controlled chemoprevention trial was conducted in subjects with confirmed histologic diagnoses of multifocal nonmetaplastic atrophy and/or intestinal metaplasia, two precancerous lesions. Individuals were assigned to receive anti-Helicobacter pylori triple therapy and/or dietary supplementation with ascorbic acid, beta-carotene, or their corresponding placebos. Gastric biopsy specimens taken at baseline were compared with those taken at 72 months. Relative risks of progression, no change, and regression from multifocal nonmetaplastic atrophy and intestinal metaplasia were analyzed with multivariate polytomous logistic regression models to estimate treatment effects. All statistical tests were two-sided. RESULTS: All three basic interventions resulted in statistically significant increases in the rates of regression: Relative risks were 4.8 (95% confidence interval [CI] = 1.6-14.2) for anti-H. pylori treatment, 5. 1 (95% CI = 1.7-15.0) for beta-carotene treatment, and 5.0 (95% CI = 1.7-14.4) for ascorbic acid treatment in subjects with atrophy. Corresponding relative risks of regression in subjects with intestinal metaplasia were 3.1 (95% CI = 1.0 9.3), 3.4 (95% CI = 1.1-9.8), and 3.3 (95% CI = 1.1-9.5). Combinations of treatments did not statistically significantly increase the regression rates. Curing the H. pylori infection (which occurred in 74% of the treated subjects) produced a marked and statistically significant increase in the rate of regression of the precursor lesions (relative risks = 8.7 [95% CI = 2.7-28.2] for subjects with atrophy and 5.4 [95% CI = 1.7-17.6] for subjects with intestinal metaplasia). CONCLUSIONS: In the very high-risk population studied, effective anti-H. pylori treatment and dietary supplementation with antioxidant micronutrients may interfere with the precancerous process, mostly by increasing the rate of regression of cancer precursor lesions, and may be an effective strategy to prevent gastric carcinoma. PMID- 11106680 TI - Cigarette smoking and colorectal cancer mortality in the cancer prevention study II. AB - BACKGROUND: Recent studies suggest that long-term cigarette smoking is associated with an increased risk of colorectal cancer. Whether the association is causal or due to confounding remains unclear. METHODS: We examined cigarette smoking in relation to colorectal cancer mortality, evaluating smoking duration and recency and controlling for potential confounders in the Cancer Prevention Study II. This prospective nationwide mortality study of 1 184 657 adults (age > or =30 years) was begun by the American Cancer Society in 1982. After exclusions, our analytic cohort included 312 332 men and 469 019 women, among whom 4432 colon or rectal cancer deaths occurred between 1982 and 1996 among individuals who were cancer free in 1982. Rate ratios (RRs) and 95% confidence intervals (CIs) were estimated by fitting Cox proportional hazards models. All statistical tests were two-sided. RESULTS: Multivariate-adjusted colorectal cancer mortality rates were highest among current smokers, were intermediate among former smokers, and were lowest in lifelong nonsmokers. The multivariate-adjusted RR (95% CI) for current compared with never smokers was 1.32 (1.16-1.49) among men and 1.41 (1.26-1.58) among women. Increased risk was evident after 20 or more years of smoking for men and women combined as compared with never smokers. Risk among current and former smokers increased with duration of smoking and average number of cigarettes smoked per day; risk in former smokers decreased significantly with years since quitting. If the multivariate-adjusted RR estimates in this study do, in fact, reflect causality, then approximately 12% of colorectal cancer deaths among both men and women in the general U.S. population in 1997 were attributable to smoking. CONCLUSIONS: Long-term cigarette smoking is associated with increased risk of colorectal cancer mortality in both men and women. Clear reduction in risk is observed with early smoking cessation. PMID- 11106681 TI - Synergistic cytotoxicity in solid tumor cell lines between N-(4 hydroxyphenyl)retinamide and modulators of ceramide metabolism. AB - BACKGROUND: We previously reported that N-(4-hydroxyphenyl)retinamide (4-HPR, fenretinide) treatment caused large increases of ceramide levels in neuroblastoma cell lines and induced cell death by a combination of apoptosis and necrosis through p53 (also known as TP53)-independent and caspase-independent pathways. Our goal was to determine if several molecules that inhibit enzymes involved in ceramide metabolism-L-threo-dihydrosphingosine (safingol), d, l-threo-1-phenyl-2 hexadecanoylamino-3-morpholino-1-propanol (PPMP), and tamoxifen-enhanced 4-HPR mediated cytotoxicity and/or affected ceramide levels. METHODS: Cellular lipids were quantified by radiolabeling and thin-layer chromatography. Cytotoxicity and cytotoxic synergy (expressed as combination index, where combination index <1 indicates synergy and >1 indicates antagonism) were measured in cultured cancer cell lines with the use of a fluorescence-based assay of cell viability employing digital imaging microscopy. Statistical tests were two-sided. RESULTS: 4-HPR increased ceramide levels by de novo synthesis. Safingol (1-4 microM) was incorporated into a stereochemical variant of ceramide and synergized with a 3:1 molar ratio of 4-HPR (3-12 microM), to produce a 100-fold to 10 000-fold (2 to 4 logs) increase in cytotoxicity relative to 4-HPR alone in neuroblastoma (combination index <0.1), lung (combination index <0.1-0.2), melanoma (combination index <0.1-0.2), prostate (combination index <0.1-1.0), colon (combination index 0.1-0.3), breast (combination index = 0.1-0.5), and pancreas (combination index = 0.2) cell lines, including p53 mutant and alkylator resistant cell lines. The 4-HPR and safingol combination was cytotoxic in low oxygen conditions and was minimally toxic to normal fibroblasts and bone marrow myeloid progenitor cells. Addition of agents that retard ceramide glucosylation and/or acylation, such as PPMP or tamoxifen, to 4-HPR or to the combination of 4 HPR and safingol further increased cytotoxicity to tumor cells. CONCLUSIONS: Combinations of 4-HPR and modulators of ceramide metabolism may form the basis for a novel chemotherapy that is functional under hypoxic conditions (e.g., such as those within tumors) and is p53 independent and caspase independent. PMID- 11106682 TI - Plasma insulin-like growth factor-I, insulin-like growth factor-binding proteins, and prostate cancer risk: a prospective study. AB - BACKGROUND: Recent studies have suggested that men with elevated plasma levels of insulin-like growth factor-I (IGF-I) may have an increased risk of prostate cancer. Furthermore, IGF-binding proteins (IGFBPs) and insulin can modulate the activity of IGF-I. In this study, we sought to determine the role of IGF-I as well as IGFBPs-1, -2, and -3 and insulin as possible etiologic factors for prostate cancer. METHODS: We conducted a nested case-control study within the Northern Sweden Health and Disease Cohort Study. We measured levels of IGF-I, IGFBP-1, IGFBP-2, IGFBP-3, and insulin in plasma samples from 149 men who had a diagnosis of prostate cancer between 1 month and 10 years after blood collection and among 298 control men. All statistical tests are two-sided. RESULTS: Case subjects had statistically significantly higher mean levels of IGF-I than control subjects (229 ng/mL; 95% confidence interval [CI] = 218-240 ng/mL] versus 214 ng/mL [95% CI = 208-221 ng/mL]; P =.02) and IGFBP-3 (2611 ng/mL [95% CI = 2518 2704 ng/mL] versus 2498 ng/mL [95% CI = 2437-2560 ng/mL]; P =.04). Conditional logistic regression analyses showed increases in prostate cancer risk with rising levels of IGF-I (P:(for trend) =.02) and IGFBP-3 (P(for trend) =.03). In case subjects younger than 59 years at the time of blood collection, the risk associated with increased IGF-I was higher (P:(for trend) =.01), whereas the risk associated with increased IGFBP-3 was lower (P(for trend) =.44) than the corresponding risks in the full cohort. Prostate cancer risk was not associated with levels of IGFBP-1, IGFBP-2, or insulin. CONCLUSIONS: Prostate cancer risk is increased in men with elevated plasma IGF-I. This association was particularly strong in younger men in this study, suggesting that circulating IGF-I may be specifically involved in the early pathogenesis of prostate cancer. PMID- 11106683 TI - Her-2-neu expression and progression toward androgen independence in human prostate cancer. AB - BACKGROUND: Human prostate cancers are initially androgen dependent but ultimately become androgen independent. Overexpression of the Her-2-neu receptor tyrosine kinase has been associated with the progression to androgen independence in prostate cancer cells. We examined the expression of Her-2-neu in normal and cancerous prostate tissues to assess its role in the progression to androgen independence. METHODS: Prostate cancer tissue sections were obtained from 67 patients treated by surgery alone (UNT tumors), 34 patients treated with total androgen ablation therapy before surgery (TAA tumors), and 18 patients in whom total androgen ablation therapy failed and who developed bone metastases (androgen-independent [AI] disease). The sections were immunostained for Her-2 neu, androgen receptor (AR), prostate-specific antigen (PSA), and Ki-67 (a marker of cell proliferation) protein expression. Messenger RNA (mRNA) levels and gene amplification of Her-2-neu were examined by RNA in situ hybridization and fluorescent in situ hybridization(FISH), respectively, in a subset of 27 tumors (nine UNT, 11 TAA, and seven AI). All statistical tests were two-sided. RESULTS: Her-2-neu protein expression was statistically significantly higher in TAA tumors than in UNT tumors with the use of two different scoring methods (P =.008 and P =.002). The proportion of Her-2-neu-positive tumors increased from the UNT group (17 of 67) to the TAA group (20 of 34) to the AI group (14 of 18) (P<.001). When compared with UNT tumors, tumor cell proliferation was higher in AI tumors (P =.014) and lower in TAA tumors (P<.001). All tumors expressed AR and PSA proteins. Although Her-2-neu mRNA expression was high in TAA and AI tumors, no Her-2-neu gene amplification was detected by FISH in any of the tumor types. CONCLUSIONS: Her-2-neu expression appears to increase with progression to androgen independence. Thus, therapeutic targeting of this tyrosine kinase in prostate cancer may be warranted. PMID- 11106684 TI - Oxidative stress and AP-1 activity in tamoxifen-resistant breast tumors in vivo. AB - BACKGROUND: Most breast cancers, even those that are initially responsive to tamoxifen, ultimately become resistant. The molecular basis for this resistance, which in some patients is thought to involve stimulation of tumor growth by tamoxifen, is unclear. Tamoxifen induces cellular oxidative stress, and because changes in cell redox state can activate signaling pathways leading to the activation of activating protein-1 (AP-1), we investigated whether tamoxifen resistant growth in vivo is associated with oxidative stress and/or activation of AP-1 in a xenograft model system where resistance is caused by tamoxifen stimulated growth. METHODS: Control estrogen-treated, tamoxifen-sensitive, and tamoxifen-resistant MCF-7 xenograft tumors were assessed for oxidative stress by measuring levels of antioxidant enzyme (e.g., superoxide dismutase [SOD], glutathione S-transferase [GST], and hexose monophosphate shunt [HMS]) activity, glutathione, and lipid peroxidation. AP-1 protein levels, phosphorylated c-jun levels, and phosphorylated Jun NH(2)-terminal kinase (JNK) levels were examined by western blot analyses, and AP-1 DNA-binding and transcriptional activities were assessed by electrophoretic mobility shift assays and a reporter gene system. All statistical tests are two-sided. RESULTS: Compared with control estrogen-treated tumors, tamoxifen resistant tumors had statistically significantly increased SOD (more than threefold; P=.004) and GST (twofold; P=.004) activity and statistically significantly reduced glutathione levels (greater than twofold; P<.001) and HMS activity (10-fold; P<.001). Lipid peroxides were not significantly different between control and tamoxifen resistant tumors. We observed no differences in AP-1 protein components or DNA binding activity. However, AP-1-dependent transcription (P=.04) and phosphorylated c-Jun and JNK levels (P<.001) were statistically significantly increased in the tamoxifen-resistant tumors. CONCLUSION: Our results suggest that the conversion of breast tumors to a tamoxifen-resistant phenotype is associated with oxidative stress and the subsequent antioxidant response and with increased phosphorylated JNK and c-Jun levels and AP-1 activity, which together could contribute to tumor growth. PMID- 11106685 TI - Analysis of the MRP4 drug resistance profile in transfected NIH3T3 cells. AB - BACKGROUND: Multidrug resistance-associated protein (MRP) 1 and canalicular multispecific organic anion transporter (cMOAT or MRP2) are adenosine triphosphate-binding cassette transporters that confer resistance to anticancer agents. In addition to these two transporters, there are at least four other human MRP subfamily members (MRP3 through MRP6). We and others reported previously that MRP3 is capable of conferring resistance to certain anticancer agents. In this study, we investigated whether MRP4 (MOAT-B), whose transcript accumulates to the highest levels in prostate tissue, has the capacity to confer drug resistance. METHODS: MRP4-transfected NIH3T3 cells were generated, and their drug sensitivity was analyzed. The subcellular localization of MRP4 was assessed by immunohistochemical analysis in transfected cells and in prostate tissue. Statistical tests were two-sided. RESULTS: MRP4 was detected as a 170-kd protein that was localized in the plasma membrane and cytoplasm of transfected cells. The MRP4 transfectants displayed 5.5-fold increased resistance to methotrexate in short-term drug-exposure assays (P=.022) and exhibited decreased cellular accumulation of this agent at 4 hours (P=.006) and 24 hours (P<.001). In continuous-exposure assays, however, the MRP4 transfectants did not display increased resistance for either methotrexate or natural product cytotoxic agents (anthracyclines, etoposide, vinca alkaloids, and paclitaxel [Taxol]). However, the transfectants did show increased resistance (2.3-fold) for the anti-acquired immunodeficiency syndrome nucleoside analogue 9-(2-phosphonylmethoxyethyl)adenine (PMEA) (P=.022) in continuous-exposure assays. Consistent with MRP4's plasma membrane localization in transfected cells, analysis of prostate tissue showed that MRP4 protein was localized primarily in the basolateral plasma membranes of tubuloacinar cells. CONCLUSIONS: These results indicate that MRP4 confers resistance to short-term methotrexate and continuous PMEA treatment. Given its structure, drug resistance profile and subcellular localization, MRP4 probably functions as an amphipathic anion efflux pump whose substrate range includes glutamate and phosphate conjugates. PMID- 11106686 TI - Expression of nuclear antigen Ki-67 in prostate cancer needle biopsy and radical prostatectomy specimens. PMID- 11106687 TI - Phenolphthalein laxatives and risk of cancer. PMID- 11106688 TI - Comprehensive cancer care: integrating alternative, complementary and conventional therapies PMID- 11106689 TI - Management of prostate cancer PMID- 11106690 TI - Current therapy in cancer, 2nd edition PMID- 11106691 TI - More about: prospective study of colorectal cancer risk in men and plasma levels of insulin-like growth factor (IGF)-I and IGF-binding protein-3. PMID- 11106692 TI - Re: population-based, case-control study of HER2 genetic polymorphism and breast cancer risk. PMID- 11106693 TI - RESPONSE: more about: prospective study of colorectal cancer risk in men and plasma levels of insulin-like growth factor (IGF)-I and IGF-binding protein-3 PMID- 11106694 TI - RESPONSE: more about: prospective study of colorectal cancer risk in men and plasma levels of insulin-like growth factor (IGF)-I and IGF- binding protein-3 PMID- 11106695 TI - Re: effect of hormone replacement therapy on breast cancer risk: estrogen versus estrogen plus progestin. PMID- 11106696 TI - RESPONSE: re: effect of hormone replacement therapy on breast cancer risk: estrogen versus estrogen plus progestin PMID- 11106697 TI - Quality improvement and accountability in the treatment of tobacco dependence: the need for a national training and certification programme. PMID- 11106699 TI - Hungary: the complete consumer service PMID- 11106698 TI - Uzbekistan: who's in charge now? PMID- 11106701 TI - Pakistan: that boat again PMID- 11106700 TI - Gambia: hey, big boy! PMID- 11106702 TI - SE Asia: Rockefeller's new programme. PMID- 11106703 TI - Canada: warnings with colour pictures required. PMID- 11106704 TI - UK: disarming truth about NRT PMID- 11106705 TI - Heterogeneity among smokers and non-smokers in attitudes and behaviour regarding smoking and smoking restrictions. AB - OBJECTIVE: To determine if smokers and non-smokers cluster into meaningful, discrete subgroups with distinguishable attitudes and behaviours regarding smoking and smoking restrictions. DESIGN: Qualitative research with 45 smokers guided development of questionnaire items applied in a population based telephone survey of 432 current smokers and 1332 non-smokers in Ontario, Canada. METHODS: Cluster analysis of questionnaire items used to categorise adult smokers and non smokers; comparison of clusters on sociodemographic characteristics and composite knowledge and attitude scores. RESULTS: Smokers clustered in three groups. "Reluctant" smokers (16%) show more concern about other people discovering that they smoke, but parallel "easygoing" smokers (42%) in supporting restrictions on smoking and not smoking around others. "Adamant" smokers (42%) feel restrictions have gone too far, and are less likely to accommodate non-smokers. Significant gradients across categories in the expected direction were observed with respect to smoking status, stage of change, knowledge, and attitude scores, and predicted compliance with restrictions, validating the proposed typology. Non-smokers also clustered into three groups, of which the "adamant" non-smokers (45%) are the least favourably disposed to smoking. "Unempowered" non-smokers (34%) also oppose smoking, but tend not to act on it. "Laissez-faire" non-smokers (21%) are less opposed to smoking in both attitude and behaviour. A significant gradient across categories in the expected direction was observed with respect to composite scores regarding knowledge of the health effects of active and passive smoking and a composite score on support for restrictions on smoking in public places. CONCLUSION: Recognition and consideration of the types of smokers and non-smokers in the population and their distinguishing characteristics could inform the development of tobacco control policies and programmes and suggest strategies to assist implementation. PMID- 11106706 TI - Development of a state wide tobacco treatment specialist training and certification programme for Massachusetts. AB - OBJECTIVE: To describe the research conducted to structure and develop a statewide tobacco training and certification programme for tobacco treatment specialists (TTSs) in Massachusetts. DESIGN: Qualitative research strategies were used to obtain information on certification development and opinions regarding TTS training and certification from key informants. A role definition and validation study was then conducted to determine the core competencies for TTSs. A comprehensive training programme was developed, piloted, and finalised, and a certification programme was initiated. PARTICIPANTS: Key informants included: individuals involved in the development of their professional certification programmes; tobacco treatment providers from across Massachusetts; and national tobacco treatment experts. MAIN OUTCOME MEASURES: Participants' opinions about the need for and structure of a training and certification programme for individuals specialising in the provision of moderate to intensive tobacco treatment; delineation of core competencies for TTSs, using the Agency for Health Care Policy and Research (now the Agency for Healthcare Research and Quality) clinical practice guideline as a foundation for the development of evidence based standards of practice for the treatment of nicotine dependence. RESULTS: The data support a comprehensive training and certification programme for TTSs in Massachusetts. Main concerns include the cost of obtaining certification, the potential to exclude uncertified healthcare professionals from delivering basic tobacco treatment, and the role of the TTS in the healthcare delivery system and the community. The training programme developed for Massachusetts was piloted, and the structure of a statewide training and certification programme for TTSs was finalised. CONCLUSIONS: The research provides support for the need and acceptance of a training and certification programme for TTSs in Massachusetts, and presents the challenges to be addressed. We demonstrated the feasibility of developing and implementing an evidence based training programme, and of initiating a statewide certification programme in Massachusetts. This work will add to a national dialogue on the development of a training and certification programme for tobacco treatment providers and encourage further research into the potential impact of statewide and national certification. PMID- 11106707 TI - Patient recall versus physician documentation in report of smoking cessation counselling performed in the inpatient setting. AB - OBJECTIVES: To determine rates of patient reported and physician documented counselling; to identify predictors of each report; and to identify the impact of each report on smoking cessation attempts after discharge from the hospital. DESIGN: Stickers on subjects' charts prompted physicians to give brief smoking cessation counselling to patients in the hospital. Patients reported counselling received and quit attempts in a phone interview conducted 7-18 days after discharge. Rates of counselling and correlations were calculated, and multivariate analysis identified predictors of patient report, physician documentation, and quit attempts. SETTING: Four hospitals in the Minneapolis/St Paul metropolitan area. SUBJECTS: 682 hospital patients who had smoked more than 100 cigarettes in their lifetime and had smoked in the last three months. RESULTS: 71.0% of patients reported counselling, and physicians documented counselling in the charts of 46.2% of patients (correlation = 0.15, kappa = 0.13). Patient report was predicted by specific hospital, belief that their hospitalisation was smoking related, diagnosis of a smoking related disease, and physician documentation of counselling. Physician documentation was predicted by female patient, specific hospital, longer hospital stay, and marginally predicted by smoking related disease. Quit attempts were predicted by patient report of counselling, but not physician documentation. CONCLUSIONS: Physicians document counselling in the hospital at a lower rate than patients report it, and the correlation between reports is very low, making an accurate assessment of true rates of counselling difficult. While it is important to increase physician documentation, it is even more important to increase patient recall, as this is the only report that predicts a quit attempt. PMID- 11106708 TI - Short term effects of cigarette smoking on hospitalisation and associated lost workdays in a young healthy population. AB - OBJECTIVE: There are relatively few published studies conducted among people of younger ages examining short term outcomes of cigarette smoking, and only a small number with outcomes important to employers. The present study was designed to assess the short term effects of smoking on hospitalisation and lost workdays. DESIGN: Retrospective cohort study. SETTING: Military population. SUBJECTS: 87 991 men and women serving on active duty in the US Army during 1987 to 1998 who took a health risk appraisal two or more times and were followed for an average of 2.4 years. MAIN OUTCOME MEASURES: Rate ratios for hospitalisations and lost workdays, and fraction of hospitalisations and lost workdays attributable to current smoking (population attributable fraction). RESULTS: Compared with never smokers, men and women who were current smokers had higher short term rates of hospitalisation and lost workdays for a broad range of conditions. Population attributable fractions (PAFs) for outcomes not related to injury or pregnancy were 7.5% (men) and 5.0% (women) for hospitalisation, and 14.1% (men) and 3.0% (women) for lost workdays. Evidence suggests that current smoking may have been under reported in this cohort, in which case the true PAFs would be higher than those reported. CONCLUSIONS: In this young healthy population, substantial fractions of hospitalisations and lost workdays were attributable to current smoking, particularly among men. PMID- 11106709 TI - Modelling the short term consequences of smoking cessation in England on the hospitalisation rates for acute myocardial infarction and stroke. AB - OBJECTIVES: To estimate the short term event and cost consequences of achieving two smoking cessation targets for England among a cohort of 35-64 year olds, in terms of the number of hospitalised acute myocardial infarctions (AMIs) and strokes avoided. DESIGN: A spreadsheet model based on previous work and using data for England was constructed to simulate the effects of achieving the target set out in the government's tobacco white paper (target 1). We also examined the consequence of achieving the intensive smoking reduction witnessed in California (target 2). RESULTS: Target 1 would result in 347 AMI and 214 stroke hospitalisations avoided in the year 2000, and by 2010 this would be 6386 AMI and 4964 strokes avoided. Achieving target 2 would result in 739 AMI and 455 stroke hospitalisations avoided in 2000, and 14 554 AMI and 11 304 strokes avoided by 2010. Achieving target 1 would save pound524 million ( pound423 million discounted at a rate of 2.67% for stroke and 2.31% for AMI) and target 2 would save pound1.14 billion ( pound921 million discounted) in terms of National Health Service costs. CONCLUSION: In the short term (11 years), reductions in the prevalence of smoking will produce sizeable reductions in both events and hospital costs. PMID- 11106710 TI - Adults' response to Massachusetts anti-tobacco television advertisements: impact of viewer and advertisement characteristics. AB - OBJECTIVE: To assess adults' receptivity to the Massachusetts television anti tobacco campaign. Reactions were examined as a function of respondents' demographics, baseline tobacco control attitudes, changes in smoking status during the campaign, and advertisements' affective qualities. DESIGN: A random digit dial telephone survey in 1993 at the start of the media campaign and re interview in 1996 of respondents to the baseline survey. PARTICIPANTS: Respondents were 1544 adults who completed the baseline and follow up interview. INTERVENTION: By the time the follow up survey was completed, approximately $49 million had been spent on the media campaign. Approximately 66 spots had been aired. MAIN OUTCOME MEASURES: Reported exposure to television advertisements; perceived effectiveness of nine specific advertisements each. RESULTS: 56% of respondents reported seeing anti-tobacco advertisements at least once a week during the preceding three years. The average effectiveness rating for all advertisements recalled on a 0-10 scale was 7.29, and did not differ by smoking status group. Advertisements eliciting strong negative emotions (sadness and fear) were rated most effective by quitters, non-smokers, and by smokers who at baseline were planning to quit soon. Humorous, entertaining advertisements were seen as ineffective by all groups. CONCLUSION: The Massachusetts anti-tobacco campaign achieved high levels of penetration into the population and was well received by both smokers and non-smokers. The results suggest that advertisements depicting suffering as a result of tobacco use may be instrumental in promoting cessation or reinforcing the decision to quit. Further research is needed to lend additional support to the link between perceived effectiveness and smoking behaviour change. PMID- 11106711 TI - Tobacco cessation skills certification in Arizona: application of a state wide, community based model for diffusion of evidence based practice guidelines. AB - OBJECTIVE: To describe the development and preliminary results from a community based certification model for training in tobacco cessation skills in Arizona. DESIGN: A programme evaluation using both quantitative pre-post measures and qualitative methods. SETTING: Arizona's comprehensive tobacco control programme of state funded, community based local projects and their community partners providing tobacco treatment services for geographically, socioeconomically, and ethnically diverse communities. INTERVENTION: A three tiered model of skills based training emphasising Agency for Health Care Policy and Research guidelines, and utilising a training of trainers approach to build community capacity. Certification roles addressed basic tobacco cessation skills, tobacco cessation specialist, and tobacco treatment services manager. PARTICIPANTS: Initial target audience was community based local project personnel and their community partners, with later adoption by community organisations unaffiliated with local projects, and the general public. MAIN EVALUATION MEASURES: Process measures: participant satisfaction, knowledge, skills, and self-efficacy. OUTCOME: participant demographics, community organisations represented, post-training, cessation related activities. RESULTS: During the model's implementation year, 1075 participants attended certification training, 947 participants received basic skills certificates and 82 received specialist certificates. Pre, post, and three month measures of self efficacy showed significant and durable increases. Analysis of participant characteristics demonstrated broad community representation. At post-training follow up, 80.9% of basic skills trainees had performed at least one brief intervention and 74.8% had made a referral to intensive services. Among cessation specialists, 48.8% were delivering intensive services and 69.5% were teaching basic skills classes. CONCLUSIONS: Initial experience with Arizona's state wide, community based model for certification of tobacco cessation skills training suggests this model may be a promising method for broad, population based diffusion of evidence based tobacco cessation guidelines. PMID- 11106712 TI - Do social support interventions ("buddy systems") aid smoking cessation? A review. AB - OBJECTIVE: To provide an overview of the role of social support in smoking cessation and to critically review evidence regarding the use of "buddy systems" (where smokers are specifically provided with someone to support them) to aid smoking cessation. DATA SOURCES: Studies were located by searching Medline and Psyclit using the key words "smoking", "smoking cessation", "social support", and "buddy". Additional studies were identified through reference lists. Only studies reported in English and published since 1980 were included. STUDY SELECTION: Studies were selected on four criteria: publication in a peer reviewed journal; randomised controlled trial using smokers who wanted to stop; the use of a social support intervention, including a "buddy"; dependent variable of smoking abstinence. Most research in this area does not use a randomised design so only a small proportion of the originally identified studies were included. DATA SYNTHESIS: In view of the diverse nature of the studies, a meta-analysis was not attempted. Ten studies were identified: nine were clinic based smoking trials, eight used a group format, and nine used buddies from among smokers' existing relationships. Support training varied from role play and rehearsal to a simple instruction to call each other regularly. Intervention and follow up periods varied between studies. Two studies showed a significant benefit of the intervention in the short term. CONCLUSIONS: Research methodology in many cases was poor. The evidence would suggest that in the context of a smokers clinic the use of buddies may be of some benefit. There is a lack of evidence regarding the efficacy of the use of buddies in community interventions. This is an important area for future research. PMID- 11106714 TI - Civil warriors PMID- 11106713 TI - Evolution of a comprehensive tobacco control programme: building system capacity and strategic partnerships--lessons from Massachusetts. AB - BACKGROUND: Since the passage of a voter approved state referendum in 1992 to establish a 25 cent increase on the excise tax on cigarettes and smokeless tobacco, Massachusetts has received an average of $40 million annually for its tobacco control programme. This funding allowed Massachusetts to expand and develop its tobacco control programme to become one of the most comprehensive in the world. OBJECTIVES: The development of the Massachusetts Tobacco Control Program is outlined, focusing on three stages of development: formation, strategic partnership building, and shared leadership. METHODS: The development of management structures, programmatic infrastructure, communication and partnership networks, and advisory structures are tracked throughout the three phases. RESULTS: The use of pre-existing public health resources, implementation of a strong training component, a geographical management structure, the creation of public and private partnerships, and the development of a shared leadership model contributed to building consensus and provided the foundation for coordinated approaches to tobacco control. CONCLUSION: Other states and countries can use lessons learned from Massachusetts about the organisational development of a comprehensive tobacco control programme as they embark upon similar efforts. PMID- 11106715 TI - Peginterferon alfa-2a in patients with chronic hepatitis C. AB - BACKGROUND: Covalent attachment of a 40-kd branched-chain polyethylene glycol moiety to interferon alfa-2a results in a compound (peginterferon alfa-2a) that has sustained absorption, a slower rate of clearance, and a longer half-life than unmodified interferon alfa-2a. We compared the clinical effects of a regimen of peginterferon alfa-2a with those of a regimen of interferon alfa-2a in the initial treatment of patients with chronic hepatitis C. METHODS: We randomly assigned 531 patients with chronic hepatitis C to receive either 180 microg of peginterferon alfa-2a subcutaneously once per week for 48 weeks (267 patients) or 6 million units of interferon alfa-2a subcutaneously three times per week for 12 weeks, followed by 3 million units three times per week for 36 weeks (264 patients). All the patients were assessed at week 72 for a sustained virologic response, defined as an undetectable level of hepatitis C virus RNA (<100 copies per milliliter). RESULTS: In the peginterferon group, 223 of the 267 patients completed treatment and 206 completed follow-up. In the interferon group, 161 of the 264 patients completed treatment and 154 completed follow-up. In an intention to-treat analysis in which patients who missed the examination at the end of treatment or follow-up were considered not to have had a response at that point, peginterferon alfa-2a was associated with a higher rate of virologic response than was interferon alfa-2a at week 48 (69 percent vs. 28 percent, P=0.001) and at week 72 (39 percent vs. 19 percent, P=0.001). Sustained normalization of serum alanine aminotransferase concentrations at week 72 was also more common in the peginterferon group than in the interferon group (45 percent vs. 25 percent, P=0.001). The two groups were similar with respect to the frequency and severity of adverse events, which were typical of those associated with interferon alfa. CONCLUSIONS: In patients with chronic hepatitis C, a regimen of peginterferon alfa-2a given once weekly is more effective than a regimen of interferon alfa-2a given three times weekly. PMID- 11106716 TI - Peginterferon alfa-2a in patients with chronic hepatitis C and cirrhosis. AB - BACKGROUND: Chronic hepatitis C virus (HCV) infection in patients with cirrhosis is difficult to treat. In patients with chronic hepatitis C but without cirrhosis, once-weekly administration of interferon modified by the attachment of a 40-kd branched-chain polyethylene glycol moiety (peginterferon alfa-2a) is more efficacious than a regimen of unmodified interferon. We examined the efficacy and safety of peginterferon alfa-2a in patients with HCV-related cirrhosis or bridging fibrosis. METHODS: We randomly assigned 271 patients with cirrhosis or bridging fibrosis to receive subcutaneous treatment with 3 million units of interferon alfa-2a three times weekly (88 patients), 90 microg of peginterferon alfa-2a once weekly (96), or 180 microg of peginterferon alfa-2a once weekly (87). Treatment lasted 48 weeks and was followed by a 24-week follow-up period. We assessed efficacy by measuring HCV RNA and alanine aminotransferase and by evaluating liver-biopsy specimens. A histologic response was defined as a decrease of at least 2 points on the 22-point Histological Activity Index. RESULTS: In an intention-to-treat analysis, HCV RNA was undetectable at week 72 in 8 percent, 15 percent, and 30 percent of the patients treated with interferon alfa-2a and with 90 microg and 180 microg of peginterferon alfa-2a, respectively (P=0.001 for the comparison between 180 microg of peginterferon alfa-2a and interferon alfa-2a). At week 72, alanine aminotransferase concentrations had normalized in 15 percent, 20 percent, and 34 percent of patients, respectively (P=0.004 for the comparison between 180 microg of peginterferon alfa-2a and interferon alfa-2a). In the subgroup of 184 patients with paired liver-biopsy specimens, the rates of histologic response at week 72 were 31 percent, 44 percent, and 54 percent, respectively (P=0.02 for the comparison between 180 microg of peginterferon alfa-2a and interferon alfa-2a). All three treatments were similarly tolerated. CONCLUSIONS: In patients with chronic hepatitis C and cirrhosis or bridging fibrosis, 180 microg of peginterferon alfa-2a administered once weekly is significantly more effective than 3 million units of standard interferon alfa-2a administered three times weekly. PMID- 11106717 TI - The risk of menstrual abnormalities after tubal sterilization. U.S. Collaborative Review of Sterilization Working Group. AB - BACKGROUND: The existence of a post-tubal-ligation syndrome of menstrual abnormalities has been debated for decades. We used data from the U.S. Collaborative Review of Sterilization to determine whether the likelihood of persistent menstrual abnormalities was greater among women who had undergone tubal sterilization than among women who had not. METHODS: A total of 9514 women who underwent tubal sterilization and 573 women whose partners underwent vasectomy were followed in a multicenter, prospective cohort study for up to five years by means of annual telephone interviews. All women were asked the same questions about six characteristics of their menstrual cycles in the presterilization and follow-up interviews. Multiple logistic-regression analysis was used to assess the risk of persistent menstrual changes. RESULTS: The women who had undergone sterilization were no more likely than those who had not undergone the procedure to report persistent changes in intermenstrual bleeding or the length of the menstrual cycle. They were more likely to have decreases in the number of days of bleeding (odds ratio, 2.4; 95 percent confidence interval, 1.1 to 5.2), the amount of bleeding (odds ratio, 1.5; 95 percent confidence interval, 1.1 to 2.0), and menstrual pain (odds ratio, 1.3; 95 percent confidence interval, 1.0 to 1.8) and to have an increase in cycle irregularity (odds ratio, 1.6; 95 percent confidence interval, 1.1 to 2.3). Among women who had had very heavy bleeding at base line, women who had undergone sterilization were more likely than women who had not undergone the procedure to report decreased bleeding (45 percent vs. 33 percent, P=0.03). CONCLUSIONS: Women who have undergone tubal sterilization are no more likely than other women to have menstrual abnormalities. PMID- 11106718 TI - Mutations in sarcomere protein genes as a cause of dilated cardiomyopathy. AB - BACKGROUND: The molecular basis of idiopathic dilated cardiomyopathy, a primary myocardial disorder that results in reduced contractile function, is largely unknown. Some cases of familial dilated cardiomyopathy are caused by mutations in cardiac cytoskeletal proteins; this finding implicates defects in contractile force transmission as one mechanism underlying this disorder. To elucidate this important cause of heart failure, we investigated other genetic causes of dilated cardiomyopathy. METHODS: Clinical evaluations were performed in 21 kindreds with familial dilated cardiomyopathy. A genome-wide linkage study prompted a search of the genes encoding beta-myosin heavy chain, troponin T, troponin I, and alpha tropomyosin for disease-causing mutations. RESULTS: A genetic locus for mutations associated with dilated cardiomyopathy was identified at chromosome 14q11.2-13 (maximal lod score, 5.11; theta=0), where the gene for cardiac beta-myosin heavy chain is encoded. Analyses of this and other genes for sarcomere proteins identified disease-causing dominant mutations in four kindreds. Cardiac beta myosin heavy-chain missense mutations (Ser532Pro and Phe764Leu) and a deletion in cardiac troponin T (deltaLys210) caused early-onset ventricular dilatation (average age at diagnosis, 24 years) and diminished contractile function and frequently resulted in heart failure. Affected persons had neither antecedent cardiac hypertrophy (average maximal left-ventricular-wall thickness, 8.5 mm) nor histopathological findings characteristic of hypertrophy. CONCLUSION: Mutations in sarcomere protein genes account for approximately 10 percent of cases of familial dilated cardiomyopathy and are particularly prevalent in families with early-onset ventricular dilatation and dysfunction. Because distinct mutations in sarcomere proteins cause either dilated or hypertrophic cardiomyopathy, the effects of mutant sarcomere proteins on muscle mechanics must trigger two different series of events that remodel the heart. PMID- 11106720 TI - Images in clinical medicine. Astrocytoma following the pyramidal tract. PMID- 11106719 TI - Treatment of Menetrier's disease with a monoclonal antibody against the epidermal growth factor receptor. PMID- 11106721 TI - Immunodeficiency diseases caused by defects in phagocytes. PMID- 11106722 TI - The diagnosis and treatment of cough. PMID- 11106723 TI - Conquering hepatitis C, step by step. PMID- 11106724 TI - Tubal sterilization--safe and effective. PMID- 11106726 TI - Correction: Hepatic Iron Concentration and Total Body Iron Stores in Thalassemia Major. PMID- 11106725 TI - Correction: Inactivation of the DNA-Repair Gene MGMT and the Clinical Response of Gliomas to Alkylating Agents. PMID- 11106727 TI - Motor protein receptors: moonlighting on other jobs. PMID- 11106728 TI - A novel motor, KIF13A, transports mannose-6-phosphate receptor to plasma membrane through direct interaction with AP-1 complex. AB - Intracellular transport mediated by kinesin superfamily proteins (KIFs) is a highly regulated process. The molecular mechanism of KIFs binding to their respective cargoes remains unclear. We report that KIF13A is a novel plus end directed microtubule-dependent motor protein and associates with beta 1-adaptin, a subunit of the AP-1 adaptor complex. The cargo vesicles of KIF13A contained AP 1 and mannnose-6-phosphate receptor (M6PR). Overexpression of KIF13A resulted in mislocalization of the AP-1 and the M6PR. Functional blockade of KIF13A reduced cell surface expression of the M6PR. Thus, KIF13A transports M6PR-containing vesicles and targets the M6PR from TGN to the plasma membrane via direct interaction with the AP-1 adaptor complex. PMID- 11106729 TI - Kinesin-dependent axonal transport is mediated by the sunday driver (SYD) protein. AB - A broadly conserved membrane-associated protein required for the functional interaction of kinesin-I with axonal cargo was identified. Mutations in sunday driver (syd) and the axonal transport motor kinesin-I cause similar phenotypes in Drosophila, including aberrant accumulations of axonal cargoes. GFP-tagged mammalian SYD localizes to tubulovesicular structures that costain for kinesin-I and a marker of the secretory pathway. Coimmunoprecipitation analysis indicates that mouse SYD forms a complex with kinesin-I in vivo. Yeast two-hybrid analysis and in vitro interaction studies reveal that SYD directly binds kinesin-I via the tetratricopeptide repeat (TPR) domain of kinesin light chain (KLC) with K(d) congruent with 200 nM. We propose that SYD mediates the axonal transport of at least one class of vesicles by interacting directly with KLC. PMID- 11106730 TI - Enhancement of memory-related long-term facilitation by ApAF, a novel transcription factor that acts downstream from both CREB1 and CREB2. AB - The memory for sensitization of the gill withdrawal reflex in Aplysia is reflected in facilitation of the monosynaptic connection between the sensory and motor neurons of the reflex. The switch from short- to long-term facilitation requires activation of CREB1, derepression of ApCREB2, and induction of ApC/EBP. In search for genes that act downstream from CREB1, we have identified a transcription activator, ApAF, which is stimulated by protein kinase A and can dimerize with both ApC/EBP and ApCREB2. ApAF is necessary for long-term facilitation induced by five pulses of serotonin, by activation of CREB1, or by derepression of ApCREB2. Overexpression of ApAF enhances the long-term facilitation further. Thus, ApAF is a candidate memory enhancer gene downstream from both CREB1 and ApCREB2. PMID- 11106731 TI - Genetic ablation and restoration of the olfactory topographic map. AB - In the olfactory sensory system, neurons expressing a given odorant receptor project with precision to two of 1800 spatially invariant glomeruli creating a topographic map within the olfactory bulb. Olfactory sensory neurons have a half life of about 90 days and are continually renewing. This poses the problem of how this precise spatial map is maintained throughout the life of the organism. We have developed a genetic approach to effect the synchronous ablation of subpopulations of neurons expressing a given receptor. The axons of newly generated neurons can then be followed as they enter the brain and converge on glomerular targets during adult life. The observation that following neuronal cell killing, the spatial map is faithfully restored, demonstrates that the information necessary for the establishment of the sensory map persists throughout the life of the organism. PMID- 11106732 TI - Structure of the molecular chaperone prefoldin: unique interaction of multiple coiled coil tentacles with unfolded proteins. AB - Prefoldin (GimC) is a hexameric molecular chaperone complex built from two related classes of subunits and present in all eukaryotes and archaea. Prefoldin interacts with nascent polypeptide chains and, in vitro, can functionally substitute for the Hsp70 chaperone system in stabilizing non-native proteins for subsequent folding in the central cavity of a chaperonin. Here, we present the crystal structure and characterization of the prefoldin hexamer from the archaeum Methanobacterium thermoautotrophicum. Prefoldin has the appearance of a jellyfish: its body consists of a double beta barrel assembly with six long tentacle-like coiled coils protruding from it. The distal regions of the coiled coils expose hydrophobic patches and are required for multivalent binding of nonnative proteins. PMID- 11106733 TI - Crystal and solution structures of an HslUV protease-chaperone complex. AB - HslUV is a "prokaryotic proteasome" composed of the HslV protease and the HslU ATPase, a chaperone of the Clp/Hsp100 family. The 3.4 A crystal structure of an HslUV complex is presented here. Two hexameric ATP binding rings of HslU bind intimately to opposite sides of the HslV protease; the HslU "intermediate domains" extend outward from the complex. The solution structure of HslUV, derived from small angle X-ray scattering data under conditions where the complex is assembled and active, agrees with this crystallographic structure. When the complex forms, the carboxy-terminal helices of HslU distend and bind between subunits of HslV, and the apical helices of HslV shift substantially, transmitting a conformational change to the active site region of the protease. PMID- 11106734 TI - Structure of Bax: coregulation of dimer formation and intracellular localization. AB - Apoptosis is stimulated by the insertion of Bax from the cytosol into mitochondrial membranes. The solution structure of Bax, including the putative transmembrane domain at the C terminus, was determined in order to understand the regulation of its subcellular location. Bax consists of 9 alpha helices where the assembly of helices alpha1 through alpha 8 resembles that of the apoptosis inhibitor, Bcl-x(L). The C-terminal alpha 9 helix occupies the hydrophobic pocket proposed previously to mediate heterodimer formation and bioactivity of opposing members of the Bcl-2 family. The Bax structure shows that the orientation of helix alpha 9 provides simultaneous control over its mitochondrial targeting and dimer formation. PMID- 11106735 TI - Solution structure of the interacting domains of the Mad-Sin3 complex: implications for recruitment of a chromatin-modifying complex. AB - Gene-specific targeting of the Sin3 corepressor complex by DNA-bound repressors is an important mechanism of gene silencing in eukaryotes. The Sin3 corepressor specifically associates with a diverse group of transcriptional repressors, including members of the Mad family, that play crucial roles in development. The NMR structure of the complex formed by the PAH2 domain of mammalian Sin3A with the transrepression domain (SID) of human Mad1 reveals that both domains undergo mutual folding transitions upon complex formation generating an unusual left handed four-helix bundle structure and an amphipathic alpha helix, respectively. The SID helix is wedged within a deep hydrophobic pocket defined by two PAH2 helices. Structure-function analyses of the Mad-Sin3 complex provide a basis for understanding the underlying mechanism(s) that lead to gene silencing. PMID- 11106736 TI - Ordered recruitment of chromatin modifying and general transcription factors to the IFN-beta promoter. AB - Here, we show that the IFN-beta enhanceosome activates transcription by directing the ordered recruitment of chromatin modifying and general transcription factors to the IFN-beta promoter. The enhanceosome is assembled in the nucleosome-free enhancer region of the IFN-beta gene, leading to the modification and remodeling of a strategically positioned nucleosome that masks the TATA box and the start site of transcription. Initially, the GCN5 complex is recruited, which acetylates the nucleosome, and this is followed by recruitment of the CBP-PolII holoenzyme complex. Nucleosome acetylation in turn facilitates SWI/SNF recruitment by CBP, resulting in chromatin remodeling. This program of recruitment culminates in the binding of TFIID to the promoter and the activation of transcription. PMID- 11106737 TI - Retroviral entry mediated by receptor priming and low pH triggering of an envelope glycoprotein. AB - Avian leukosis virus (ALV) has been used as a model system to understand the mechanism of pH-independent viral entry involving receptor-induced conformational changes in the viral envelope (Env) glycoprotein that lead to membrane fusion. Here, we report the unexpected finding that ALV entry depends on a critical low pH step that was overlooked when this virus was directly compared to the classical pH-dependent influenza A virus. In contrast to influenza A virus, receptor interaction plays an essential role in priming ALV Env for subsequent low pH triggering. Our results reveal a novel principle in viral entry, namely that receptor interaction can convert a pH-insensitive viral glycoprotein to a form that is responsive to low pH. PMID- 11106738 TI - The mouse Spo11 gene is required for meiotic chromosome synapsis. AB - The Spo11 protein initiates meiotic recombination by generating DNA double-strand breaks (DSBs) and is required for meiotic synapsis in S. cerevisiae. Surprisingly, Spo11 homologs are dispensable for synapsis in C. elegans and Drosophila yet required for meiotic recombination. Disruption of mouse Spo11 results in infertility. Spermatocytes arrest prior to pachytene with little or no synapsis and undergo apoptosis. We did not detect Rad51/Dmc1 foci in meiotic chromosome spreads, indicating DSBs are not formed. Cisplatin-induced DSBs restored Rad51/Dmc1 foci and promoted synapsis. Spo11 localizes to discrete foci during leptotene and to homologously synapsed chromosomes. Other mouse mutants that arrest during meiotic prophase (Atm -/-, Dmc1 -/-, mei1, and Morc(-/-)) showed altered Spo11 protein localization and expression. We speculate that there is an additional role for Spo11, after it generates DSBs, in synapsis. PMID- 11106739 TI - Chromosome synapsis defects and sexually dimorphic meiotic progression in mice lacking Spo11. AB - Spo11, a protein first identified in yeast, is thought to generate the chromosome breaks that initiate meiotic recombination. We now report that disruption of mouse Spo11 leads to severe gonadal abnormalities from defective meiosis. Spermatocytes suffer apoptotic death during early prophase; oocytes reach the diplotene/dictyate stage in nearly normal numbers, but most die soon after birth. Consistent with a conserved function in initiating meiotic recombination, Dmc1/Rad51 focus formation is abolished. Spo11(-/-) meiocytes also display homologous chromosome synapsis defects, similar to fungi but distinct from flies and nematodes. We propose that recombination initiation precedes and is required for normal synapsis in mammals. Our results also support the view that mammalian checkpoint responses to meiotic recombination and/or synapsis defects are sexually dimorphic. PMID- 11106740 TI - Chromatin opening of DNA satellites by targeted sequence-specific drugs. AB - There are few tools available for dissecting and elucidating the functions of DNA satellites and other nongenic DNA. To address this, we have explored the experimental potential of DNA sequence-specific drugs containing pyrrole and imidazole amino acids (polyamides). Compounds were synthesized that target different Drosophila melanogaster satellites. Dimeric oligopyrroles were shown to target the AT-rich satellites I, III, and SARs (scaffold associated regions). One polyamide (P31) specifically binds the GAGAA satellite V. Specificity of targeting was established by footprinting, epifluorescence of nuclei, and polytene chromosomes stained with fluorescent derivatives. These polyamides were shown to mediate satellite-specific chromatin opening of the chromatin fiber. Remarkably, certain polyamides induced defined gain or loss-of-function phenotypes when fed to Drosophila melanogaster. PMID- 11106741 TI - Specific gain- and loss-of-function phenotypes induced by satellite-specific DNA binding drugs fed to Drosophila melanogaster. AB - DNA-binding pyrrole-imidazole compounds were synthesized that target different Drosophila melanogaster satellites. Compound P31 specifically binds the GAGAA satellite V, and P9 targets the AT-rich satellites I and III. Remarkably, these drugs, when fed to developing Drosophila flies, caused gain- or loss-of-function phenotypes. While polyamide P9 (not P31) suppressed PEV of white-mottled flies (increased gene expression), P31 (not P9) mediated three well-defined, homeotic transformations (loss-of-function) exclusively in brown-dominant flies. Both phenomena are explained at the molecular level by chromatin opening (increased accessibility) of the targeted DNA satellites. Chromatin opening of satellite III by P9 is proposed to suppress PEV of white-mottled flies, whereas chromatin opening of satellite V by P31 is proposed to create an inopportune "sink" for the GAGA factor (GAF). PMID- 11106742 TI - A chromatin insulator determines the nuclear localization of DNA. AB - Chromatin insulators might regulate gene expression by controlling the subnuclear organization of DNA. We found that a DNA sequence normally located inside of the nucleus moved to the periphery when the gypsy insulator was placed within the sequence. The presence of the gypsy insulator also caused two sequences, normally found in different regions of the nucleus, to come together at a single location. Alterations in this subnuclear organization imposed by the gypsy insulator correlated with changes in gene expression that took place during the heat-shock response. These global changes in transcription were accompanied by dramatic alterations in the distribution of insulator proteins and DNA. The results suggest that the nuclear organization imposed by the gypsy insulator on the chromatin fiber is important for gene expression. PMID- 11106743 TI - Histone acetylation and hSWI/SNF remodeling act in concert to stimulate V(D)J cleavage of nucleosomal DNA. AB - The ordered assembly of immunoglobulin and TCR genes by V(D)J recombination depends on the regulated accessibility of individual loci. We show here that the histone tails and intrinsic nucleosome structure pose significant impediments to V(D)J cleavage. However, alterations to nucleosome structure via histone acetylation or by stable hSWI/SNF-dependent remodeling greatly increase the accessibility of nucleosomal DNA to V(D)J cleavage. Moreover, acetylation and hSWI/SNF remodeling can act in concert on an individual nucleosome to achieve levels of V(D)J cleavage approaching those observed on naked DNA. These results are consistent with a model in which regulated recruitment of chromatin modifying activities is involved in mediating the lineage and stage-specific control of V(D)J recombination. PMID- 11106744 TI - ATP-driven chromatin remodeling activity and histone acetyltransferases act sequentially during transactivation by RAR/RXR In vitro. AB - Using a "crude" chromatin-based transcription system that mimics transactivation by RAR/RXR heterodimers in vivo, we could not demonstrate that chromatin remodeling was required to relieve nucleosomal repression. Using "purified" chromatin templates, we show here that, irrespective of the presence of histone H1, both ATP-driven chromatin remodeling activities and histone acetyltransferase (HAT) activities of coactivators recruited by liganded receptors are required to achieve transactivation. DNA footprinting, ChIP analysis, and order of addition experiments indicate that coactivator HAT activities and two ATP-driven remodeling activities are sequentially involved at distinct steps preceding initiation of transcription. Thus, both ATP-driven chromatin remodeling and HAT activities act in a temporally ordered and interdependent manner to alleviate the repressive effects of nucleosomal histones on transcription by RARalpha/RXRalpha heterodimers. PMID- 11106746 TI - An RNA-binding chameleon. AB - The arginine-rich RNA binding motif is found in a wide variety of proteins, including several viral regulatory proteins. Although related at the primary sequence level, arginine-rich domains from different proteins adopt different conformations depending on the RNA site recognized, and in some cases fold only in the context of RNA. Here we show that the RNA binding domain of the Jembrana disease virus (JDV) Tat protein is able to recognize two different TAR RNA sites, from human and bovine immunodeficiency viruses (HIV and BIV, respectively), adopting different conformations in the two RNA contexts and using different amino acids for recognition. In addition to the conformational differences, the JDV domain requires the cyclin T1 protein for high-affinity binding to HIV TAR, but not to BIV TAR. The "chameleon-like" behavior of the JDV Tat RNA binding domain reinforces the concept that RNA molecules can provide structural scaffolds for protein folding, and suggests mechanisms for evolving distinct RNA binding specificities from a single multifunctional domain. PMID- 11106745 TI - Competitive recruitment of CBP and Rb-HDAC regulates UBF acetylation and ribosomal transcription. AB - RNA polymerase I (PolI) transcription is activated by the HMG box architectural factor UBF, which loops approximately 140 bp of DNA into the enhancesome, necessitating major chromatin remodeling. Here we show that the acetyltransferase CBP is recruited to and acetylates UBF both in vitro and in vivo. CBP activates PolI transcription in vivo through its acetyltransferase domain and acetylation of UBF facilitates transcription derepression and activation in vitro. CBP activation and Rb suppression of ribosomal transcription by recruitment to UBF are mutually exclusive, regulating in vivo PolI transcription through an acetylation-deacetylation "flip-flop." Thus, PolI transcription is regulated by protein acetylation, and the competitive recruitment of CBP and Rb. PMID- 11106747 TI - Functional anatomy of a dsRNA trigger: differential requirement for the two trigger strands in RNA interference. AB - In RNA-mediated interference (RNAi), externally provided mixtures of sense and antisense RNA trigger concerted degradation of homologous cellular RNAs. We show that RNAi requires duplex formation between the two trigger strands, that the duplex must include a region of identity between trigger and target RNAs, and that duplexes as short as 26 bp can trigger RNAi. Consistent with in vitro observations, a fraction of input dsRNA is converted in vivo to short segments of approximately 25 nt. Interference assays with modified dsRNAs indicate precise chemical requirements for both bases and backbone of the RNA trigger. Strikingly, certain modifications are well tolerated on the sense, but not the antisense, strand, indicating that the two trigger strands have distinct roles in the interference process. PMID- 11106748 TI - The apoptosis-promoting factor TIA-1 is a regulator of alternative pre-mRNA splicing. AB - We report here that the apoptosis-promoting protein TIA-1 regulates alternative pre-mRNA splicing of the Drosophila melanogaster gene male-specific-lethal 2 and of the human apoptotic gene Fas. TIA-1 associates selectively with pre-mRNAs that contain 5' splice sites followed by U-rich sequences. TIA-1 binding to the U-rich stretches facilitates 5' splice site recognition by U1 snRNP. This activity is critical for activation of the weak 5' splice site of msl-2 and for modulating the choice of splice site partner in Fas. Structural and functional similarities with the Saccharomyces cerevisiae splicing factor Nam8 suggest striking evolutionary conservation of a mechanism of pre-mRNA splicing regulation that controls biological processes as diverse as meiosis in yeast, dosage compensation in fruit flies, or programmed cell death in humans. PMID- 11106749 TI - Regulated translation initiation controls stress-induced gene expression in mammalian cells. AB - Protein kinases that phosphorylate the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha) are activated in stressed cells and negatively regulate protein synthesis. Phenotypic analysis of targeted mutations in murine cells reveals a novel role for eIF2alpha kinases in regulating gene expression in the unfolded protein response (UPR) and in amino acid starved cells. When activated by their cognate upstream stress signals, the mammalian eIF2 kinases PERK and GCN2 repress translation of most mRNAs but selectively increase translation of Activating Transcription Factor 4 (ATF4), resulting in the induction of the downstream gene CHOP (GADD153). This is the first example of a mammalian signaling pathway homologous to the well studied yeast general control response in which eIF2alpha phosphorylation activates genes involved in amino acid biosynthesis. Mammalian cells thus utilize an ancient pathway to regulate gene expression in response to diverse stress signals. PMID- 11106750 TI - JunD protects cells from p53-dependent senescence and apoptosis. AB - JunD is the most broadly expressed member of the Jun family and the AP-1 transcription factor complex. Primary fibroblasts lacking JunD displayed p53 dependent growth arrest, upregulated p19(Arf) expression, and premature senescence. In contrast, immortalized cell lines lacking JunD showed increased proliferation and higher cyclinD1 levels. These properties are reminiscent of the effects of oncogenic Ras expression on primary and established cell cultures. Furthermore, JunD(-/-) fibroblasts exhibited increased p53-dependent apoptosis upon ultraviolet irradiation and were sensitive to the cytotoxic effects of TNF alpha. The antiapoptotic role of JunD was confirmed using an in vivo model of TNF mediated hepatitis. We propose that JunD protects cells from senescence, or apoptotic responses to stress stimuli, by acting as a modulator of the signaling pathways that link Ras to p53. PMID- 11106751 TI - A PKC-eta/Fyn-dependent pathway leading to keratinocyte growth arrest and differentiation. AB - Growth control of epithelial cells differs substantially from other cell types. Activation of Fyn, a Src kinase family member, is required for normal keratinocyte differentiation. We report that increased Fyn activity by itself suppresses growth of keratinocytes, but not dermal fibroblasts, through downmodulation of EGF receptor (EGFR) signaling. Protein kinase C-eta has also been implicated in keratinocyte growth/differentiation control. We show that growth suppression of keratinocytes by PKC-eta depends mostly on Fyn. PKC-eta activity is both necessary and sufficient for Fyn activation, PKC-eta and Fyn are found in association, and recombinant PKC-eta directly activates Fyn. Thus, our findings reveal a direct cross talk between PKC-eta and Fyn, which presides over the decision between keratinocyte (epithelial) cell growth and differentiation. PMID- 11106752 TI - Two critical hits for promyelocytic leukemia. AB - Acute promyelocytic leukemia (APL) is associated with chromosomal translocations that always involve the RARalpha gene, which variably fuses to one of several distinct loci, including PML or PLZF (X genes). Due to the reciprocity of the translocation, X-RARalpha and RARalpha-X fusion proteins coexist in APL blasts. PLZF-RARalpha transgenic mice (TM) develop leukemia that lacks the differentiation block at the promyelocytic stage that characterizes APL. We generated TM expressing RARalpha-PLZF and PLZF-RARalpha in their promyelocytes. RARalpha-PLZF TM do not develop leukemia. However, PLZF-RARalpha/RARalpha-PLZF double TM develop leukemia with classic APL features. We demonstrate that RARalpha-PLZF can interfere with PLZF transcriptional repression and that this is critical for APL pathogenesis, since leukemias in PLZF(-/-)/PLZF-RARalpha mutants and in PLZF-RARalpha/RARalpha-PLZF TM are indistinguishable. Thus, both products of a cancer-associated translocation are crucial in determining the distinctive features of the disease. PMID- 11106753 TI - Combgap relays wingless signal reception to the determination of cortical cell fate in the Drosophila visual system. AB - The dorsoventral axis of the Drosophila visual cortex is patterned by nonautonomous signals expressed at its dorsal and ventral margins. wingless (wg) expression at the margins induces decapentaplegic (dpp), optomotor blind (omb), and aristaless in adjacent domains. We show that Combgap, a zinc finger protein, represses Wg target gene expression in the visual cortex. Wg signal reception downregulates combgap expression and derepresses target gene transcription. Combgap participates in a Hedgehog-controlled circuit in the developing wing and leg by regulating the expression of Cubitus interruptus. Combgap is thus a tissue specific relay between Wingless and its target genes for the determination of cell fate in the visual cortex. PMID- 11106754 TI - Phosphorylation of the Cdc42 exchange factor Cdc24 by the PAK-like kinase Cla4 may regulate polarized growth in yeast. AB - Rho-type GTPases control many cytoskeletal rearrangements, but their regulation remains poorly understood. Here, we show that in S. cerevisiae, activation of the CDK Cdc28-Cln2 at bud emergence triggers relocalization of Cdc24, the GEF for Cdc42, from the nucleus to the polarization site, where it is stably maintained by binding to the adaptor Bem1. Locally activated Cdc42 then polarizes the cytoskeleton in a manner dependent on its effectors Bni1 and the PAK-like kinase Cla4. In addition, Cla4 induces phosphorylation of Cdc24, leading to its dissociation from Bem1 at bud tips, thereby ending polarized bud growth in vivo. Our results thus suggest a dynamic temporal and spatial regulation of the Cdc42 module: Cdc28-Cln triggers actin polarization by activating Cdc42, which in turn restricts its own activation via a negative feedback loop acting on its GEF Cdc24. PMID- 11106755 TI - The molecular basis of FHA domain:phosphopeptide binding specificity and implications for phospho-dependent signaling mechanisms. AB - Forkhead-associated (FHA) domains are a class of ubiquitous signaling modules that appear to function through interactions with phosphorylated target molecules. We have used oriented peptide library screening to determine the optimal phosphopeptide binding motifs recognized by several FHA domains, including those within a number of DNA damage checkpoint kinases, and determined the X-ray structure of Rad53p-FHA1, in complex with a phospho-threonine peptide, at 1.6 A resolution. The structure reveals a striking similarity to the MH2 domains of Smad tumor suppressor proteins and reveals a mode of peptide binding that differs from SH2, 14-3-3, or PTB domain complexes. These results have important implications for DNA damage signaling and CHK2-dependent tumor suppression, and they indicate that FHA domains play important and unsuspected roles in S/T kinase signaling mechanisms in prokaryotes and eukaryotes. PMID- 11106756 TI - Crystal structure of eukaryotic DNA ligase-adenylate illuminates the mechanism of nick sensing and strand joining. AB - Chlorella virus DNA ligase is the smallest eukaryotic ATP-dependent ligase known; it has an intrinsic nick-sensing function and suffices for yeast cell growth. Here, we report the 2.0 A crystal structure of the covalent ligase-AMP reaction intermediate. The conformation of the adenosine nucleoside and contacts between the enzyme and the ribose sugar have undergone a significant change compared to complexes of T7 ligase with ATP or mRNA capping enzyme with GTP. The conformational switch allows the 3' OH of AMP to coordinate directly the 5' PO(4) of the nick. The structure explains why nick sensing is restricted to adenylated ligase and why the 5' phosphate is required for DNA binding. We identify a metal binding site on ligase-adenylate and propose a mechanism of nick recognition and catalysis supported by mutational data. PMID- 11106757 TI - Crystal structure of yeast Esa1 suggests a unified mechanism for catalysis and substrate binding by histone acetyltransferases. AB - Esa1 is the catalytic subunit of the NuA4 histone acetylase (HAT) complex that acetylates histone H4, and it is a member of the MYST family of HAT proteins that includes the MOZ oncoprotein and the HIV-1 Tat interacting protein Tip60. Here we report the X-ray crystal structure of the HAT domain of Esa1 bound to coenzyme A and investigate the protein's catalytic mechanism. Our data reveal that Esa1 contains a central core domain harboring a putative catalytic base, and flanking domains that are implicated in histone binding. Comparisons with the Gcn5/PCAF and Hat1 proteins suggest a unified mechanism of catalysis and histone binding by HAT proteins, whereby a structurally conserved core domain mediates catalysis, and sequence variability within a structurally related N- and C-terminal scaffold determines substrate specificity. PMID- 11106758 TI - The beta-slip: a novel concept in transthyretin amyloidosis. AB - Transthyretin is a tetrameric plasma protein associated with two forms of amyloid disease. The structure of the highly amyloidogenic transthyretin triple mutant TTRG53S/E54D/L55S determined at 2.3 A resolution reveals a novel conformation: the beta-slip. A three-residue shift in beta strand D places Leu-58 at the position normally occupied by Leu-55 now mutated to serine. The beta-slip is best defined in two of the four monomers, where it makes new protein-protein interactions to an area normally involved in complex formation with retinol binding protein. This interaction creates unique packing arrangements, where two protein helices combine to form a double helix in agreement with fiber diffraction and electron microscopy data. Based on these findings, a novel model for transthyretin amyloid formation is presented. PMID- 11106759 TI - The structure of ribosome-channel complexes engaged in protein translocation. AB - Cotranslational translocation of proteins requires ribosome binding to the Sec61p channel at the endoplasmic reticulum (ER) membrane. We have used electron cryomicroscopy to determine the structures of ribosome-channel complexes in the absence or presence of translocating polypeptide chains. Surprisingly, the structures are similar and contain 3-4 connections between the ribosome and channel that leave a lateral opening into the cytosol. Therefore, the ribosome channel junction may allow the direct transfer of polypeptides into the channel and provide a path for the egress of some nascent chains into the cytosol. Moreover, complexes solubilized from mammalian ER membranes contain an additional membrane protein that has a large, lumenal protrusion and is intercalated into the wall of the Sec61p channel. Thus, the native channel contains a component that is not essential for translocation. PMID- 11106760 TI - Mechanism of membrane insertion of a multimeric beta-barrel protein: perfringolysin O creates a pore using ordered and coupled conformational changes. AB - Perfringolysin O, a bacterial cytolytic toxin, forms unusually large pores in cholesterol-containing membranes by the spontaneous insertion of two of its four domains into the bilayer. By monitoring the kinetics of domain-specific conformational changes and pore formation using fluorescence spectroscopy, the temporal sequence of domain-membrane interactions has been established. One membrane-exposed domain does not penetrate deeply into the bilayer and is not part of the actual pore, but is responsible for membrane recognition. This domain must bind to the membrane before insertion of the other domain into the bilayer is initiated. The two domains are conformationally coupled, even though they are spatially separated. Thus, cytolytic pore formation is accomplished by a novel mechanism of ordered conformational changes and interdomain communication. PMID- 11106761 TI - Vesicular stomatitis virus matrix protein inhibits host cell gene expression by targeting the nucleoporin Nup98. AB - Vesicular stomatitis virus matrix protein (VSV M) has been shown to inhibit both transcription and nucleocytoplasmic transport. We have isolated a mutant form of M, termed M(D), lacking both inhibitory activities. HeLa cells expressing M, but not M(D), accumulate polyadenylated RNAs within the nucleus. Concomitantly, a fraction of M, but not of the M(D) mutant, localizes at the nuclear rim. Additionally, the nucleoporin Nup98 specifically interacts with M but not with M(D). In Nup98(-/-) cells, both the levels of M at the nuclear envelope and its inhibitory effects on host cell-directed expression of reporter genes were significantly reduced. Together, our data demonstrate that VSV M inhibits host cell gene expression by targeting a nucleoporin and primarily blocking nuclear export. PMID- 11106762 TI - Phosphorylation of CPEB by Eg2 mediates the recruitment of CPSF into an active cytoplasmic polyadenylation complex. AB - The release of Xenopus oocytes from prophase I arrest is largely driven by the cytoplasmic polyadenylation-induced translation of dormant maternal mRNAs. Two cis elements, the CPE and the hexanucleotide AAUAAA, and their respective binding factors, CPEB and a cytoplasmic form of CPSF, control polyadenylation. The most proximal stimulus for polyadenylation is Eg2-catalyzed phosphorylation of CPEB serine 174. Here, we show that this phosphorylation event stimulates an interaction between CPEB and CPSF. This interaction is direct, does not require RNA tethering, and occurs through the 160 kDa subunit of CPSF. Eg2-stimulated and CPE-dependent polyadenylation is reconstituted in vitro using purified components. These results demonstrate that the molecular function of Eg2 phosphorylated CPEB is to recruit CPSF into an active cytoplasmic polyadenylation complex. PMID- 11106763 TI - Structural basis for nucleotide exchange and competition with tRNA in the yeast elongation factor complex eEF1A:eEF1Balpha. AB - The crystal structure of a complex between the protein biosynthesis elongation factor eEF1A (formerly EF-1alpha) and the catalytic C terminus of its exchange factor, eEF1Balpha (formerly EF-1beta), was determined to 1.67 A resolution. One end of the nucleotide exchange factor is buried between the switch 1 and 2 regions of eEF1A and destroys the binding site for the Mg(2+) ion associated with the nucleotide. The second end of eEF1Balpha interacts with domain 2 of eEF1A in the region hypothesized to be involved in the binding of the CCA-aminoacyl end of the tRNA. The competition between eEF1Balpha and aminoacylated tRNA may be a central element in channeling the reactants in eukaryotic protein synthesis. The recognition of eEF1A by eEF1Balpha is very different from that observed in the prokaryotic EF-Tu:EF-Ts complex. Recognition of the switch 2 region in nucleotide exchange is, however, common to the elongation factor complexes and those of Ras:Sos and Arf1:Sec7. PMID- 11106764 TI - Polycystin-1, the gene product of PKD1, induces resistance to apoptosis and spontaneous tubulogenesis in MDCK cells. AB - The major form of autosomal dominant polycystic kidney disease (ADPKD) results from mutation of a gene (PKD1) of unknown function that is essential for the later stages of renal tubular differentiation. In this report, we describe a novel cell culture system for studying how PKD1 regulates this process. We show that expression of human PKD1 in MDCK cells slows their growth and protects them from programmed cell death. MDCK cells expressing PKD1 also spontaneously form branching tubules while control cells form simple cysts. Increased cell proliferation and apoptosis have been implicated in the pathogenesis of cystic diseases. Our study suggests that PKD1 may function to regulate both pathways, allowing cells to enter a differentiation pathway that results in tubule formation. PMID- 11106766 TI - New insight into the structure and function of the alternative oxidase. AB - The alternative oxidase is a ubiquinol oxidase found in plant mitochondria, as well as in the mitochondria of some fungi and protists. It catalyzes a cyanide resistant reduction of oxygen to water without translocation of protons across the inner mitochondrial membrane, and thus functions as a non-energy-conserving member of the respiratory electron transfer chain. The active site of the alternative oxidase has been modelled as a diiron center within a four-helix bundle by Siedow et al. (FEBS Lett. 362 (1995) 10-14) and more recently by Andersson and Nordlund (FEBS Lett. 449 (1999) 17-22). The cloning of the Arabidopsis thaliana IMMUTANS (Im) gene, which encodes a plastid enzyme distantly related to the mitochondrial alternative oxidases (Wu et al. Plant Cell 11 (1999) 43-55; Carol et al. Plant Cell 11 (1999) 57-68), has now narrowed the range of possible ligands to the diiron center of the alternative oxidase. The Im protein sequence suggests a minor modification to the recent model of the active site of the alternative oxidase. This change moves an invariant tyrosine into a conserved hydrophobic pocket in the vicinity of the active site, in a position analogous to the long-lived tyrosine radical at the diiron center of ribonucleotide reductase, and similar to the tyrosines near the diiron center of bacterioferritin and rubrerythrin. The Im sequence and modified structural model yield a compelling picture of the alternative oxidase as a diiron carboxylate protein. The current status of the relationship of structure to function in the alternative oxidase is reviewed. PMID- 11106767 TI - Electron transport through photosystem II in leaves during light pulses: acceptor resistance increases with nonphotochemical excitation quenching. AB - Light response curves of photosystem (PS) II electron transport from oxygen evolving complex to plastoquinone (PQ) were measured in sunflower (Helianthus annuus L.), cotton (Gossypium hirsutum L.) and tobacco (Nicotiana tabacum L.) leaves by recording O(2) evolution and fluorescence in 5-200 ms light pulses of 500-13500 micromol absorbed quanta m(-2) s(-1). The leaves were pre-adapted at 60 2000 micromol quanta m(-2) s(-1) for 60 min to obtain different nonphotochemical excitation quenching, which was predominantly of reversible q(I) type (relaxation time 30 min). PQ was completely oxidized by turning the actinic light off and illuminating with far-red light for 2 s before the pulse was applied in the dark, 4 s after the actinic light was turned off. Electron transport rate calculated from fluorescence transients considering PS II donor side resistance (V. Oja, A. Laisk, submitted) was maximal at the beginning of pulses (J(Fi)) and decreased immediately. The dependences of J(Fi) on pulse absorbed flux density were rectangular hyperbolas with K(m) about 7500 micromol m(-2) s(-1). Both the extrapolated plateau J(Fm) and initial slope (intrinsic quantum yield of PS II, Y(m)) decreased proportionally when q(I) increased from minimum to maximum (J(Fm) from 2860 to 1450 micromol e(-) m(-2) s(-1) and Y(m) from 0.41 to 0.23). The time constant for electron transfer away from the PS II acceptor side, calculated from a model of PS II electron transport for 2 micromol PS II m(-2), increased from 607 to 1315 microseconds with the activation of q(I) while the donor side time constant changed from 289 to 329 microseconds. These results show that changes in the electron transfer processes on the acceptor side of PS II occur in parallel with nonphotochemical (predominantly reversible q(I) type) excitation quenching. PMID- 11106768 TI - Are mitochondria a permanent source of reactive oxygen species? AB - The observation that in isolated mitochondria electrons may leak out of the respiratory chain to form superoxide radicals (O(2)(radical-)) has prompted the assumption that O(2)(radical-) formation is a compulsory by-product of respiration. Since mitochondrial O(2)(radical-) formation under homeostatic conditions could not be demonstrated in situ so far, conclusions drawn from isolated mitochondria must be considered with precaution. The present study reveals a link between electron deviation from the respiratory chain to oxygen and the coupling state in the presence of antimycin A. Another important factor is the analytical system applied for the detection of activated oxygen species. Due to the presence of superoxide dismutase in mitochondria, O(2)(radical-) release cannot be realistically determined in intact mitochondria. We therefore followed the release of the stable dismutation product H(2)O(2) by comparing most frequently used H(2)O(2) detection methods. The possible interaction of the detection systems with the respiratory chain was avoided by a recently developed method, which was compared with conventional methods. Irrespective of the methods applied, the substrates used for respiration and the state of respiration established, intact mitochondria could not be made to release H(2)O(2) from dismutating O(2)(radical-). Although regular mitochondrial respiration is unlikely to supply single electrons for O(2)(radical-) formation our study does not exclude the possibility of the respiratory chain becoming a radical source under certain conditions. PMID- 11106769 TI - An elementary kinetic model of energy coupling in biological membranes. AB - The purpose of this work is to contribute to the understanding of the fundamental kinetic properties of the processes of energy coupling in biological membranes. For this, we consider a model of a microorganism that, in its plasma membrane, expresses two electrogenic enzymes (E(1) and E(2)) transporting the same monovalent cation C and electrodiffusive paths for C and for a monovalent anion A. E(1) (E(2)) couples transport C to the reaction S(1)<-->P(1) (S(2)<-->P(2)). We developed a mathematical model that describes the rate of change of the electrical potential difference across the membrane, of the internal concentrations of C and A, and of the concentrations of S(2) and P(2). The enzymes are incorporated via two-state kinetic models; the passive ionic fluxes are represented by classical formulations of electrodiffusion. The microorganism volume is maintained constant by accessory regulatory devices. The model is utilized for stationary and dynamic studies for the case of bacteria employing the electrochemical gradient of Na(+) as energetic intermediate. Among other conclusions, the results show that the membrane potential represents the relevant kinetic intermediate for the overall coupling between the energy donor reaction S(1)<-->P(1) and the synthesis of S(2). PMID- 11106770 TI - Oxygen yield from single turnover flashes in leaves: non-photochemical excitation quenching and the number of active PSII. AB - O(2) evolution from single turnover flashes of up to 96 micromol absorbed quanta m(-2) and from multiple turnover pulses of 8.6 and 38.6 ms duration and 12800 and 850 micromol absorbed quanta m(-2) s(-1) intensity, respectively, was measured in sunflower leaves with the help of zirconium O(2) analyser. O(2) evolution from one flash could be measured with 1% accuracy on the background of 10-50 micromol O(2) mol(-1). Before the measurements leaves were pre-adapted either at 30-60 or 1700 micromol quanta m(-2) s(-1) to induce different non-photochemical excitation quenching (q(N)). Short (1 min) exposures at the high light that created only energy-dependent, q(E) type quenching, caused no changes in the O(2) yield from saturating flashes or pulses that could be related to the q(E) quenching, but the yield from low intensity flashes and pulses decreased considerably. Long 30-60 min exposures at the high light induced a reversible inhibitory, q(I) type quenching that decreased the O(2) yield from both, saturating and limiting flashes and pulses (but more from the limiting ones), which reversed within 15 min under the low light. The results are in agreement with the notion that q(E) is caused by a quenching process in the PSII antenna and no changes occur in the PSII centres, but the reversible (15-30 min) q(I) quenching is accompanied by inactivation of a part of PSII centres. PMID- 11106771 TI - Definition of crucial structural factors of acetogenins, potent inhibitors of mitochondrial complex I. AB - Some natural acetogenins are the most potent inhibitors of bovine heart mitochondrial complex I. These compounds are characterized by two functional units (i.e. hydroxylated tetrahydrofuran (THF) and alpha,beta-unsaturated gamma lactone ring moieties) separated by a long alkyl spacer. To elucidate which structural factors of acetogenins including their active conformation are crucial for the potent inhibitory effect, we synthesized a series of novel acetogenin analogues possessing bis-THF rings. The present study clearly demonstrated that the natural gamma-lactone ring is not crucial for the potent inhibition, although this moiety is the most common structural unit among a large number of natural acetogenins and has been suggested to be the only reactive species that directly interacts with the enzyme (Shimada et al., Biochemistry 37 (1998) 854-866). The presence of free hydroxy group(s) in the adjacent bis-THF rings was favorable, but not essential, for the potent activity. This was probably because high polarity (or hydrophilicity), rather than hydrogen bond-donating ability, around the bis-THF rings is required to retain the inhibitor in the active conformation. Interestingly, length of the alkyl spacer proved to be a very important structural factor for the potent activity, the optimal length being approximately 13 carbon atoms. The present study provided further strong evidence for the previous proposal (Kuwabara et al., Eur. J. Biochem. 267 (2000) 2538-2546) that the gamma-lactone and THF ring moieties act in a cooperative manner on complex I with the support of some specific conformation of the spacer. PMID- 11106772 TI - Light-induced changes of NADPH fluorescence in isolated chloroplasts: a spectral and fluorescence lifetime study. AB - Isolated chloroplasts show a light-induced reversible increase in blue-green fluorescence (BGF), which is only dependent on NADPH changes. In the present communication, we report a time-resolved and spectral analysis of this BGF in reconstituted chloroplasts and intact isolated chloroplasts, in the dark and under actinic illumination. From these measurements we deduced the contribution of the different forms of NADPH (free and bound to proteins) to the light-induced variation of BGF and conclude that this variation is due only to the redox change of the NADP pool. A simple model estimating the distribution of NADPH between the free and bound form was designed, that explains the differences measured for the BGF of reconstituted chloroplasts and intact chloroplasts. From the decay associated spectra of the chloroplast BGF, we also deduced the participation of flavins to the green peak of chloroplast fluorescence emission spectrum, and the existence of excitation energy transfer from proteins to bound NADPH in chloroplasts. In addition, we re-examined the use of chloroplast BGF as a quantitative measure of NADPH concentration, and confirmed that chloroplast BGF can be used for non-destructive, continuous and probably quantitative monitoring of light-induced changes in NADP redox state. PMID- 11106773 TI - The position of cytochrome b(559) relative to Q(A) in photosystem II studied by electron-electron double resonance (ELDOR). AB - The electron-electron double resonance (ELDOR) method was applied to measure the dipole interaction between cytochrome (Cyt) b(+)(559) and the primary acceptor quinone (Q(-)(A)), observed at g=2.0045 with the peak to peak width of about 9 G, in Photosystem II (PS II) in which the non-heme Fe(2+) was substituted by Zn(2+). The paramagnetic centers of Cyt b(+)(559)Y(D)Q(-)(A) were trapped by illumination at 273 K for 8 min, followed by dark adaptation for 3 min and freezing into 77 K. The distance between the pair Cyt b(+)(559)-Q(-)(A) was estimated from the dipole interaction constant fitted to the observed ELDOR time profile to be 40+/-1 A. In the membrane oriented PS II particles the angle between the vector from Q(A) to Cyt b(559) and the membrane normal was determined to be 80+/-5 degrees. The position of Cyt b(559) relative to Q(A) suggests that the heme plane is located on the stromal side of the thylakoid membrane. ELDOR was not observed for Cyt b(+)(559) Y(D) spin pair, suggesting the distance between them is more than 50 A. PMID- 11106774 TI - Energy transfer and charge separation in the purple non-sulfur bacterium Roseospirillum parvum. AB - The antenna reaction centre system of the recently described purple non-sulfur bacterium Roseospirillum parvum strain 930I was studied with various spectroscopic techniques. The bacterium contains bacteriochlorophyll (BChl) a, 20% of which was esterified with tetrahydrogeranylgeraniol. In the near-infrared, the antenna showed absorption bands at 805 and 909 nm (929 nm at 6 K). Fluorescence bands were located at 925 and 954 nm, at 300 and 6 K, respectively. Fluorescence excitation spectra and time resolved picosecond absorbance difference spectroscopy showed a nearly 100% efficient energy transfer from BChl 805 to BChl 909, with a time constant of only 2.6 ps. This and other evidence indicate that both types of BChl belong to a single LH1 complex. Flash induced difference spectra show that the primary electron donor absorbs at 886 nm, i.e. at 285 cm(-1) higher energy than the long wavelength antenna band. Nevertheless, the time constant for trapping in the reaction centre was the same as for almost all other purple bacteria: 55+/-5 ps. The shape as well as the amplitude of the absorbance difference spectrum of the excited antenna indicated exciton interaction and delocalisation of the excited state over the BChl 909 ring, whereas BChl 805 appeared to have a monomeric nature. PMID- 11106775 TI - Resistance of isolated pulmonary mitochondria during in vitro anoxia/reoxygenation. AB - The aim of the study was to investigate the effect of in vitro anoxia/reoxygenation on the oxidative phosphorylation of isolated lung mitochondria. Mitochondria were isolated after harvesting from fresh pig lungs flushed with Euro-Collins solution. Mitochondrial respiratory parameters were determined in isolated mitochondria before anoxia (control), after 5-45 min anoxia followed by 5 min reoxygenation, and after 25 or 40 min of in vitro incubation in order to follow the in vitro aging of mitochondria during respiratory assays. Respiratory parameters measured after anoxia/reoxygenation did not show any oxidative phosphorylation dysfunction, indicating a high resistance of pulmonary mitochondria to in vitro anoxia/reoxygenation (up to 45 min anoxia). These results indicate that mitochondria are not directly responsible of their oxidative phosphorylation damage observed after in vivo ischemia (K. Willet et al., Transplantation 69 (2000) 582) but are a target of others cellular injuries leading to mitochondrial dysfunction in vivo. PMID- 11106776 TI - Interactions between redox partners in various cytochrome P450 systems: functional and structural aspects. AB - The various types of redox partner interactions employed in cytochrome P450 systems are described. The similarities and differences between the redox components in the major categories of P450 systems present in bacteria, mitochondria and microsomes are discussed in the light of the accumulated evidence from X-ray crystallographic and NMR spectroscopic determinations. Molecular modeling of the interactions between the redox components in various P450 mono-oxygenase systems is proposed on the basis of structural and mutagenesis information, together with experimental findings based on chemical modification of key residues likely to be associated with complementary binding sites on certain typical P450 isoforms and their respective redox partners. PMID- 11106777 TI - A lysine residue involved in the inhibition of vacuolar H(+)-pyrophosphatase by fluorescein 5'-isothiocyanate. AB - Vacuolar proton pumping pyrophosphatase (H(+)-PPase; EC 3.6.1.1) plays a central role in the electrogenic translocation of protons from cytosol to the vacuole lumen at the expense of PP(i) hydrolysis. A fluorescent probe, fluorescein 5' isothiocyanate (FITC), was used to modify a lysine residue of vacuolar H(+) PPase. The enzymatic activity and its associated H(+) translocation of vacuolar H(+)-PPase were markedly decreased by FITC in a concentration-dependent manner. The inhibition of enzymatic activity followed pseudo-first-order rate kinetics. A double-logarithmic plot of the apparent reaction rate constant against FITC concentration yielded a straight line with a slope of 0.89, suggesting that the alteration of a single lysine residue on the enzyme is sufficient to inhibit vacuolar H(+)-PPase. Changes in K(m) but not V(max) values of vacuolar H(+)-PPase as inhibited by FITC were obtained, indicating that the labeling caused a modification in affinity of the enzyme to its substrate. FITC inhibition of vacuolar H(+)-PPase could be protected by its physiological substrate, Mg(2+) PP(i). These results indicate that FITC might specifically compete with the substrate at the active site and the FITC-labeled lysine residue locates probably in or near the catalytic domain of the enzyme. The enhancement of fluorescence intensity and the blue shift of the emission maximum of FITC after modification of vacuolar H(+)-PPase suggest that the FITC-labeled lysine residue is located in a relatively hydrophobic region. PMID- 11106778 TI - Kinetic characterization of His-tagged CP47 photosystem II in Synechocystis sp. PCC6803. AB - Recently, construction of strains of Synechocystis sp. PCC6803 having a His(6) extension (His-tag) of the carboxyl terminus of the CP47 protein has been reported (T.M. Bricker et al, Biochim. Biophys. Acta 1409 (1998) 50; M.J. Reifler et al., in: Garab, Pusztai (Eds.) Proc. XIth International Congress on Photosynthesis, 1998). While these initial reports suggest a minimal impact of the His-tag upon Photosystem (PS) II function, a more thorough analysis of the kinetic properties of the modified complex is essential. This communication reports on a more detailed kinetic analysis to assess possible perturbations of PS II due to the genetic addition of the His-tag on the CP47 protein. It was found that: (1) Patterns of flash O(2) yield exhibited normal period four oscillations and the associated fits of the Kok-Joliot S-state cycling parameters were virtually identical to the wild type; (2) O(2) release kinetics during the S(3)-S(0) transition were experimentally indistinguishable from the wild type; (3) S-state decay measurements indicate slightly faster decays of the S(2) and S(3) states compared to the wild type; (4) fluorescence measurements indicate that the kinetics of the forward reaction of electron transfer from Q(A)(-) to Q(B) and back-reactions of Q(A)(-) with PS II electron donors are similar in the His-tag and wild-type strains. It is therefore concluded that the addition of the His-tag results in a minimal perturbation of PS II function. PMID- 11106779 TI - A historical overview of chemical research on cannabinoids. AB - The chemical research on the plant cannabinoids and their derivatives over two centuries is concisely reviewed. The tortuous path leading to the discovery of the endogenous cannabinoids is described. Future directions, which will probably be followed are delineated. PMID- 11106780 TI - Conformational requirements for endocannabinoid interaction with the cannabinoid receptors, the anandamide transporter and fatty acid amidohydrolase. AB - Anandamide (N-arachidonoylethanolamine) has been identified as an endogenous ligand of the G-protein coupled cannabinoid CB(1) receptor. Recent studies have postulated the existence of carrier-mediated anandamide transport which is involved in the termination of the biological effects of anandamide. A membrane bound amidohydrolase (fatty acid amide hydrolase, FAAH), located intracellulary, hydrolyzes and inactivates anandamide and other endogenous cannabinoids such as 2 arachidonoylglycerol (2-AG). Structure-activity relationships (SARs) for endocannabinoid interaction with the CB receptors, the anandamide transporter and FAAH are currently emerging in the literature. This review considers the divergences between these SARs and focuses upon the conformational implications for endocannabinoid recognition at each of these biological targets. PMID- 11106781 TI - Molecular probes for the cannabinoid receptors. AB - Cannabinoids produce most of their biochemical and pharmacological effects by interacting with CB1 and CB2 cannabinoid receptors, both of which are G-protein coupled membrane-bound functional proteins. CB1 is found in the central nervous system and in a variety of other organs including heart, vascular endothelium, uterus, vas deferens, testis and small intestine. Conversely, the CB2 receptor appears to be associated exclusively with the immune system and is found in the periphery of the spleen and other cells associated with immunochemical functions. Although both CB1 and CB2 have been cloned and the primary sequences are known, their three dimensional structures and the amino acid residues at the active site, critical for ligand recognition, binding and activation have not been characterized. In the absence of any X-ray crystallographic and NMR data, information on the structural requirements for ligand-receptor interactions is obtained with the help of suitably designed molecular probes. These ligands either interact with the receptor in a reversible fashion (reversible probes) or, alternatively, attach at or near the receptor active site with the formation of a covalent bond (irreversible probes). Subsequently, information related to ligand binding and receptor activation is further amplified with the help of receptor mutants and computer modeling. This review focuses on molecular probes related to the classical and non-classical cannabinoids that have been reported since the discovery of the first cannabinoid receptor over a decade ago. PMID- 11106782 TI - Cellular signal transduction by anandamide and 2-arachidonoylglycerol. AB - Anandamide (arachidonylethanolamide) and 2-arachidonoylglycerol mediate many of their actions via either CB(1) or CB(2) cannabinoid receptor subtypes. These agonist-receptor interactions result in activation of G proteins, particularly those of the G(i/o) family. Signal transduction pathways that are regulated by these G proteins include inhibition of adenylyl cyclase, regulation of ion currents (inhibition of voltage-gated L, N and P/Q Ca(2+)-currents; activation of K(+) currents); activation of focal adhesion kinase (FAK), mitogen activated protein kinase (MAPK) and induction of immediate early genes; and stimulation of nitric oxide synthase (NOS). Other effects of anandamide and/or 2 arachidonoylglycerol that are not mediated via cannabinoid receptors include inhibition of L-type Ca(2+) channels, stimulation of VR(1) vanilloid receptors, transient changes in intracellular Ca(2+), and disruption of gap junction function. Cardiovascular regulation by anandamide appears to occur by a variety of receptor-mediated and non-receptor-mediated mechanisms. This review will describe and evaluate each of these signal transduction pathways and mechanisms. PMID- 11106783 TI - Pathways and mechanisms of N-acylethanolamine biosynthesis: can anandamide be generated selectively? AB - Long-chain N-acylethanolamines (NAEs) and their precursors, N-acylethanolamine phospholipids, are ubiquitous trace constituents of animal and human cells, tissues and body fluids. Their cellular levels appear to be tightly regulated and they accumulate as the result of injury. Saturated and monounsaturated congeners which represent the vast majority of cellular NAEs can have cytoprotective effects while polyunsaturated NAEs, especially 20:4n-6 NAE (anandamide), elicit physiological effects by binding to and activating cannabinoid receptors. It is the purpose of this article to review published data on the pathways and mechanisms of NAE biosynthesis in mammals and to evaluate this information for its physiological significance. The generation and turnover of NAE via N-acyl PE through the transacylation-phosphodiesterase pathway may represent a novel cannabinoid receptor-independent signalling system, analogous to and possibly related to ceramide-mediated cell signalling. PMID- 11106784 TI - 2-Arachidonoylglycerol and the cannabinoid receptors. AB - 2-Arachidonoylglycerol (2-AG) is a unique molecular species of monoacylglycerol isolated from rat brain and canine gut as an endogenous cannabinoid receptor ligand (Sugiura, T., Kondo, S., Sukagawa, A., Nakane, S., Shinoda, A., Itoh, K., Yamashita, A., Waku, K., 1995. 2-Arachidonoylglycerol: a possible endogenous cannabinoid receptor ligand in brain. Biochem. Biophys. Res. Commun. 215, 89-97; Mechoulam, R., Ben-Shabat, S., Hanus, L., Ligumsky, M., Kaminski, N. E., Schatz, A.R., Gopher, A., Almog, S., Martin, B.R., Compton, D.R., Pertwee, R.G., Giffin, G., Bayewitch, M., Brag, J., Vogel, Z., 1995. Identification of an endogenous 2 monoglyceride, present in canine gut, that binds to cannabinoid receptors. Biochem. Pharmacol. 50, 83-90). 2-AG binds to the cannabinoid receptors (CB1 and CB2) and exhibits a variety of cannabimimetic activities in vitro and in vivo. Recently, we found that 2-AG induces Ca(2+) transients in NG108-15 cells, which express the CB1 receptor, and in HL-60 cells, which express the CB2 receptor, through a cannabinoid receptor- and Gi/Go-dependent mechanism. Based on the results of structure-activity relationship experiments, we concluded that 2-AG but not anandamide is the natural ligand for both the CB1 and the CB2 receptors and both receptors are primarily 2-AG receptors. Evidences are gradually accumulating that 2-AG is a physiologically essential molecule, although further detailed studies appear to be necessary to determine relative importance of 2-AG and anandamide in various animal tissues. In this review, we described mainly our previous and current experimental results, as well as those of others, concerning the tissue levels, bioactions and metabolism of 2-AG. PMID- 11106785 TI - The fatty acid amide hydrolase (FAAH). AB - The topic of this review is fatty acid amide hydrolase (FAAH), one of the best characterized enzymes involved in the hydrolysis of bioactive lipids such as anandamide, 2-arachidonoylglycerol (2-AG), and oleamide. Herein, we discuss the nomenclature, the various assays that have been developed, the relative activity of the various substrates and the reversibility of the enzyme reactions catalyzed by FAAH. We also describe the cloning of the enzyme from rat and subsequent cDNA isolation from mouse, human, and pig. The proteins and the mRNAs from different species are compared. Cloning the enzyme permitted the purification and characterization of recombinant FAAH. The conserved regions of FAAH are described in terms of sequence and function, including the amidase domain which contains the serine catalytic nucleophile, the hydrophobic domain important for self association, and the proline rich domain region, which may be important for subcellular localization. The distribution of FAAH in the major organs of the body is described as well as regional distribution in the brain and its correlation with cannabinoid receptors. Since FAAH is recognized as a drug target, a large number of inhibitors have been synthesized and tested since 1994 and these are reviewed in terms of reversibility, potency, and specificity for FAAH and cannabinoid receptors. PMID- 11106786 TI - The movement of N-arachidonoylethanolamine (anandamide) across cellular membranes. AB - This review presents and explores the hypothesis that N-arachidonoylethanolamine (AEA, also called anandamide) is transported across cellular membranes by a process that is protein-mediated. Support for this hypothesis comes from experiments demonstrating that cellular accumulation of extracellularly applied AEA is saturable, time and temperature dependent and exhibits selective inhibition by various structural analogs of AEA. The accumulation of AEA is cell specific; data is presented demonstrating that several cell types, including the bovine adrenal zona glomerulosa cell, exhibit very high capacity for AEA accumulation while others, such as the HeLa cell, have a very low capacity. The transport process has the characteristics of facilitated diffusion; it is bi directional, not dependent on either ATP or extracellular sodium and exhibits the trans effect of flux coupling. Several important questions remain to be answered regarding the carrier, including its molecular structure and its role in the release and inactivation of endogenously produced AEA. PMID- 11106787 TI - N-Acylethanolamines and precursor phospholipids - relation to cell injury. AB - The present review focuses on the relationship between formation of N acylethanolamine phospholipids (NAPEs) and N-acyletransferase (NAEs) catalyzed by N-acyltranferase and NAPE-hydrolyzing phospholipase D, respectively, and cell injury in tissues like brain, heart, and testis. A number of mechanisms are proposed by which these two groups of lipids may have cytoprotective properties. The mechanisms may involve activation of cannabinoid receptors, as well as non receptor-mediated effects such as stabilization of membrane bilayers, antioxidant mechanisms, inhibition of calcium leakage from mitochondria, and direct inhibition of ceramidase. Anandamide (20:4-NAE) is formed as a minor component along with other NAEs during cell injury. Whether 20:4-NAE has a separate physiological role is at present not known, but some data suggest that 20:4-NAE may be formed, e.g. in the uterus, by a more selective mechanism without being accompanied by a vast majority of saturated and monounsaturated NAEs. PMID- 11106788 TI - The endocannabinoid system: a physiological perspective on its role in psychomotor control. AB - The discovery of cannabinoid receptors has led to the identification of two natural activators for these receptors, anandamide and 2-arachidonoylglycerol, and to the elucidation of their biochemical pathways of formation and inactivation. Although the physiological significance of the endogenous cannabinoid system is still poorly understood, important information is becoming available on the possible functional roles of this system in the basal ganglia, a forebrain region that is involved in the control of sensorimotor and motivational aspects of behavior. These discoveries - which are going to enrich the way in which we look at basal ganglia functions - are summarized in this mini-review. The role of the endocannabinoids as modulators of psychomotor behaviors and the potential therapeutic perspectives deriving from the pharmacological manipulation of the endogenous cannabinoid system are also discussed. PMID- 11106789 TI - Endocannabinoids as cardiovascular modulators. AB - Cannabinoids, the bioactive constituents of the marijuana plant and their synthetic and endogenous analogs cause not only neurobehavioral, but also cardiovascular effects. The most important component of these effects is a profound decrease in blood pressure and heart rate. Although multiple lines of evidence indicate that the hypotensive and bradycardic effects of anandamide and other cannabinoids are mediated by peripherally located CB1 cannabinoid receptors, anandamide can also elicit vasodilation in certain vascular beds, which is independent of CB1 or CB2 receptors. Possible cellular mechanisms underlying these effects and the cellular sources of vasoactive anandamide are discussed. PMID- 11106790 TI - Cannabinoid receptors and the regulation of immune response. AB - Cannabinoid research underwent a tremendous increase during the last 10 years. This progress was made possible by the discovery of cannabinoid receptors and the endogenous ligands for these receptors. Cannabinoid research is developing in two major directions: neurobehavioral properties of cannabinoids and the impact of cannabinoids on the immune system. Recent studies characterized the cannabinoid induced response as a very complex process because of the involvement of multiple signalling pathways linked to cannabinoid receptors or effects elicited by cannabinoids without receptor participation. The objective of this review is to present this complexity as it applies to immune response. The functional properties of cannabinoid receptors, signalling pathways linked to cannabinoid receptors and the modulation of immune response by cannabinoid receptor ligands are discussed. Special attention is given to 'endocannabinoids' as immunomodulatory molecules. PMID- 11106791 TI - Endocannabinoids and fatty acid amides in cancer, inflammation and related disorders. AB - The long history of the medicinal use of Cannabis sativa and, more recently, of its chemical constituents, the cannabinoids, suggests that also the endogenous ligands of cannabinoid receptors, the endocannabinoids, and, particularly, their derivatives may be used as therapeutic agents. Studies aimed at correlating the tissue and body fluid levels of endogenous cannabinoid-like molecules with pathological conditions have been started and may lead to identify those diseases that can be alleviated by drugs that either mimic or antagonize the action of these substances, or modulate their biosynthesis and degradation. Hints for the therapeutic applications of endocannabinoids, however, can be obtained also from our previous knowledge of marijuana medicinal properties. In this article, we discuss the anti-tumor and anti-inflammatory activity of: (1) the endocannabinoids anandamide (arachidonoylethanolamide) and 2-arachidonoyl glycerol; (2) the bioactive fatty acid amides palmitoylethanolamide and oleamide; and (3) some synthetic derivatives of these compounds, such as the N-acyl vanillyl-amines. Furthermore, the possible role of cannabimimetic fatty acid derivatives in the pathological consequences of cancer and inflammation, such as cachexia, wasting syndrome, chronic pain and local vasodilation, will be examined. PMID- 11106792 TI - Ligand-receptor signaling with endocannabinoids in preimplantation embryo development and implantation. AB - Although adverse effects of cannabinoids on pregnancy have been indicated for many years, the mechanisms by which they exert their actions were not clearly understood. Only recently, molecular and biochemical approaches have led to the identification of two types of cannabinoid receptors, brain-type receptors (CB1 R) and spleen-type receptors (CB2-R), which mediate cannabinoid effects. These findings were followed by the discovery of endocannabinoids, anandamide and 2 arachidonoylglycerol (2-AG). The natural cannabinoids and endocannabinoids exert their effects via cannabinoid receptors and share similar pharmacological and physiological properties. Recent demonstration of expression of functional CB1-R in the preimplantation embryo and synthesis of anandamide in the pregnant uterus of mice suggests that cannabinoid ligand-receptor signaling is operative in the regulation of preimplantation embryo development and implantation. This review describes recent observations and their significance in embryo-uterine interactions during implantation and future research directions in this emerging area of interest. PMID- 11106793 TI - Emerging physiological roles for N-acylphosphatidylethanolamine metabolism in plants: signal transduction and membrane protection. AB - The activation of N-acylphosphatidylethanolamine (NAPE) metabolism in plants appears to be associated mostly with cellular stresses. In response to pathogen elicitors, NAPE is hydrolzyed by phospholipase-D (PLD), and corresponding medium chain, saturated N-acylethanolamines (NAEs) are released by plant cells where they act as lipid mediators to modulate ion flux and activate defense gene expression. In desiccated seeds of higher plants, long-chain, saturated and unsaturated NAEs are prevalent, but are rapidly metabolized during the first few hours of imbibition, a period of substantial osmotic stress. NAPE synthesis is increased in seeds during this same period of rapid rehydration. A membrane-bound enzyme designated NAPE synthase has been purified from imbibed cottonseeds and its unusual biochemical properties suggest that it may scavenge free fatty acids in vivo. This feature of NAPE metabolism may be unique to higher plants a may be a mechanism for the rapid recycling of fatty acids back into membrane-associated NAPE. Altogether, increasing evidence indicates that NAPE metabolism in plants shares functional similarities with NAPE metabolism in animal systems, including signal transduction and cellular protection. In particular, the emerging role of released NAEs as lipid mediators in plant defense signaling represents an intriguing parallel to 'endocannabinoid signaling' in several mammalian cell types. PMID- 11106794 TI - Identification of mucAB-like homologs on two IncT plasmids, R394 and Rts-1. AB - Recent phylogenetic analysis of the superfamily of lesion-replicating DNA polymerases suggest that they can be broadly divided into four sub-groups comprised of UmuC-like, DinB-like, Rev1-like and Rad30-like proteins. The UmuC like sub-family is best characterized at the genetic level and sequence analysis of eleven umu orthologs, residing on bacterial chromosomes or on self transmissible R-plasmids allows further subdivision into five sub-groups (UmuDC, MucAB, ImpAB, RumAB and RulAB) based on amino acid sequence conservation. Some of these orthologs are apparently inactive in situ, but may promote increased mutagenesis and survival when subcloned and expressed from high-copy number plasmids. We were, therefore, interested in devising an assay that would identify umuC-like genes in situ in the absence of a functional assay. To this end, degenerate primers directed towards conserved amino acid regions within the UmuC like sub-family of DNA polymerases were designed and used to identify mucAB-like operons on the IncT plasmids, R394 and Rts-1.Interestingly, DNA sequence analysis of an approximately 7kb region of R394 identified two LexA-regulated genes immediately downstream of mucAB((R394)) that are similar to the chromosomally encoded Escherichia coli tus gene and the IncI plasmid-encoded impC gene, respectively. Analysis of the R394 and Rts-1 mucB genes revealed that both contain insertions which result in the expression of a truncated inactive MucB protein. While R394 was unable to restore mutagenesis functions to a DeltaumuDC E. coli strain, Rts-1 surprisingly promoted significant levels of MMS-induced SOS mutagenesis, raising the possibility that Rts-1 encodes another, yet unidentified, umu-like homolog. PMID- 11106795 TI - Induction of chromosome aberrations in vitro by phenolphthalein: mechanistic studies. AB - Phenolphthalein induces tumors in rodents but because it is negative in assays for mutation in Salmonella and in mammalian cells, for DNA adducts and for DNA strand breaks, its primary mechanism does not seem to be DNA damage. Chromosome aberration (Ab) induction by phenolphthalein in vitro is associated with marked cytotoxicity. At very high doses, phenolphthalein induces weak increases in micronuclei (MN) in mouse bone marrow; a larger response is seen with chronic treatment. All this suggests genotoxicity is a secondary effect that may not occur at lower doses. In heterozygous TSG-p53((R)) mice, phenolphthalein induces lymphomas and also MN, many with kinetochores (K), implying chromosome loss. Induction of aneuploidy would be compatible with the loss of the normal p53 gene seen in the lymphomas. Here we address some of the postulated mechanisms of genotoxicity in vitro, including metabolic activation, inhibition of thymidylate synthetase, cytotoxicity, oxidative stress, DNA damage and aneuploidy. We show clearly that phenolphthalein does not require metabolic activation by S9 to induce Abs. Inhibition of thymidylate synthetase is an unlikely mechanism, since thymidine did not prevent Ab induction by phenolphthalein. Phenolphthalein dramatically inhibited DNA synthesis, in common with many non-DNA reactive chemicals that induce Abs at cytotoxic doses. Phenolphthalein strongly enhances levels of intracellular oxygen radicals (ROS). The radical scavenger DMSO suppresses phenolphthalein-induced toxicity and Abs whereas H(2)O(2) potentiates them, suggesting a role for peroxidative activation. Phenolphthalein did not produce DNA strand breaks in rat hepatocytes or DNA adducts in Chinese hamster ovary (CHO) cells. All the evidence points to an indirect mechanism for Abs that is unlikely to operate at low doses of phenolphthalein. We also found that phenolphthalein induces mitotic abnormalities and MN with kinetochores in vitro. These are also enhanced by H(2)O(2) and suppressed by DMSO. Our findings suggest that induction of Abs in vitro is a high-dose effect in oxidatively stressed cells and may thus have a threshold. There may be more than one mechanism operating in vitro and in vivo, possibly indirect genotoxicity at high doses and also chromosome loss, both of which would likely have a threshold. PMID- 11106796 TI - Benzo(a)pyrene and X-rays induce reversions of the pink-eyed unstable mutation in the retinal pigment epithelium of mice. AB - The pink-eyed unstable (p(un)) mutation is the result of a 70kb tandem duplication within the murine p gene. Homologous deletion/recombination of the locus to wild-type occurs spontaneously in embryos and results in pigmented spots in the fur and eye that persist for life. Such deletion events are also inducible by a variety of DNA damaging agents, as we have observed previously with the fur spot assay. Here, we describe the use of the retinal pigment epithelium (RPE) of the eye to detect reversion events induced with two differently acting agents. Benzo(a)pyrene (B(a)P) induces a high frequency, and X-ray exposure a more modest increase, of p(un) reversion in both the fur and the eye. The eye-spot assay requires fewer mice for significant results than the fur spot assay. Previous work had elucidated the cell proliferation pattern in the RPE and a position effect variegation phenotype in the pattern of p(un) reversions, which we have confirmed. Acute exposure to B(a)P or X-rays resulted in an increased frequency of reversion events. The majority of the spontaneous reversions lie toward the periphery of the RPE whereas induced events are found more centrally, closer to the optic nerve head. The induced distribution corresponds to the major sites of cell proliferation in the RPE at the time of exposure, and further advocates the proposal that dividing cells are at highest risk to develop deletions. PMID- 11106797 TI - Mutagenicity in lung of big Blue((R)) mice and induction of tandem-base substitutions in Salmonella by the air pollutant peroxyacetyl nitrate (PAN): predicted formation of intrastrand cross-links. AB - Peroxyacetyl nitrate (PAN) is a ubiquitous air pollutant formed from NO(2) reacting with acetoxy radicals generated from ambient aldehydes in the presence of sunlight and ozone. It contributes to eye irritation associated with photochemical smog and is present in most urban air. PAN was generated in a chamber containing open petri dishes of Salmonella TA100 (gas-phase exposure). After subtraction of the background mutation spectrum, the spectrum of PAN induced mutants selected at 3.1-fold above the background mutant yield was 59% GC ->TA, 29% GC-->AT, 2% GC-->CG, and 10% multiple mutations - primarily GG-->TT tandem-base substitutions. Using computational molecular modeling methods, a mechanism was developed for producing this unusual tandem-base substitution. The mechanism depends on the protonation of PAN near the polyanionic DNA to release NO(2)(+) resulting in intrastrand dimer formation. Insertion of AA opposite the dimerized GG would account for the tandem GG-->TT transversions. Nose-only exposure of Big Blue((R)) mice to PAN at 78ppm (near the MTD) was mutagenic at the lacI gene in the lung (mutant frequency +/-S.E. of 6.16+/-0.58/10(5) for controls versus 8.24+/-0.30/10(5) for PAN, P=0.016). No tandem-base mutations were detected among the 40 lacI mutants sequenced. Dosimetry with 3H-PAN showed that 24h after exposure, 3.9% of the radiolabel was in the nasal tissue, and only 0.3% was in the lung. However, based on the molecular modeling considerations, the labeled portion of the molecule would not have been expected to have been bound covalently to DNA. Our results indicate that PAN is weakly mutagenic in the lungs of mice and in Salmonella and that PAN produces a unique signature mutation (a tandem GG-->TT transversion) in Salmonella that is likely due to a GG intrastrand cross-link. Thus, PAN may pose a mutagenic and possible carcinogenic risk to humans, especially at the high concentrations at which it is present in some urban environments. PMID- 11106798 TI - 3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) induces apoptosis in rat splenocytes and thymocytes by different mechanisms. AB - 3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) is a potent carcinogen present in cooked meat. Although the target of this carcinogen is mainly in the liver, Trp-P-1 is distributed in the thymus and spleen as well as in the liver after administration. However, the cytotoxic effect of Trp-P-1 on lymphocytes has not been examined in detail. In the present study, we investigated the cytotoxicity of Trp-P-1 against rat splenocytes and thymocytes. Trp-P-1 reduced viability of both types of cells in the same manner, the LD(50) at 6h in culture was 15 microM, and the time for the 50% decrease in cell viability (t(1/2)) at 20 microM was 3h. In both types of cells, Trp-P-1 caused the activation of caspase-3 like proteases and the cleavage of poly(ADP-ribose) polymerase, both of which are biochemical markers of apoptosis. On the other hand, DNA fragmentation occurred in splenocytes, but not in thymocytes although Trp-P-1 activated 32-34kDa nucleases that may not be able to degrade DNA into nucleosomal units. These results indicated that Trp-P-1 induces apoptosis in both splenocytes and thymocytes by different mechanisms in which distinct apoptotic pathways may exist downstream of the caspase cascade. PMID- 11106799 TI - Ethnic differences in the expression of Epstein-Barr virus latent membrane protein-1 mutations in nasopharyngeal carcinoma. AB - The latent membrane protein-1 (LMP1) of Epstein-Barr virus (EBV) is a viral oncoprotein implicated in several EBV-associated pathologies. Many studies have characterized carboxy-terminal mutations within LMP1, errors in this area are critical since this portion contains sequences responsible for LMP1 targeting, half-life and association with host cell proteins. Although, data suggests that mutations in this area extend LMP1 half-life and increase its oncogenesis, some studies have not shown this to be true for all EBV-associated tumors. In order to evaluate 3'-end LMP1-DNA mutations in three different ethnic populations with nasopharyngeal carcinoma (NPC), we examined EBV-DNA in 34 patients of various origins (Caucasian, Chinese and Inuit). While 68% of the total group expressed EBV-antigens, only 56% of Caucasians but 86% of Inuit expressed this viral protein. Over 67% of Inuit NPC tissue contained the characteristic 30 bp deletion that was observed in only 20% of Caucasians and 33% of Chinese samples. DNA sequencing revealed that the Inuit population showed the most frequent DNA mutations and corresponding amino acid alterations in LMP1. Our results suggest that EBV-associated NPC-DNA mutations in LMP1 do not occur at equal rates in different racial groups and are more common at distinct sites in NPC tissue from Chinese and Inuit sources. PMID- 11106800 TI - No correlation between germline mutation at repeat DNA and meiotic crossover in male mice exposed to X-rays or cisplatin. AB - To test the hypothesis that mouse germline expanded simple tandem repeat (ESTR) mutations are associated with recombination events during spermatogenesis, crossover frequencies were compared with germline mutation rates at ESTR loci in male mice acutely exposed to 1Gy of X-rays or to 10mg/kg of the anticancer drug cisplatin. Ionising radiation resulted in a highly significant 2.7-3.6-fold increase in ESTR mutation rate in males mated 4, 5 and 6 weeks after exposure, but not 3 weeks after exposure. In contrast, irradiation had no effect on meiotic crossover frequencies assayed on six chromosomes using 25 polymorphic microsatellite loci spaced at approximately 20cM intervals and covering 421cM of the mouse genome. Paternal exposure to cisplatin did not affect either ESTR mutation rates or crossover frequencies, despite a report that cisplatin can increase crossover frequency in mice. Correlation analysis did not reveal any associations between the paternal ESTR mutation rate and crossover frequency in unexposed males and in those exposed to X-rays or cisplatin. This study does not, therefore, support the hypothesis that mutation induction at mouse ESTR loci results from a general genome-wide increase in meiotic recombination rate. PMID- 11106801 TI - Role in tumorigenesis of silent mutations in the TP53 gene. AB - Over 10,000 mutations in the TP53 suppressor gene have been recorded in the International Agency for Research on Cancer (IARC) tumor data base. About 4% of these mutations are silent. It is a question whether these mutations play a role in tumor development. In order to approach this question, we asked whether the reported silent mutations are randomly distributed throughout the TP53 gene. The p53 data base was searched exon by exon. From the frequency of codons with no silent mutations, the average number of silent mutations per codon for each exon was calculated using the Poisson distribution. The results indicate the distribution to be non-random. About one-third of all silent mutations occur in "hot-spots" and after subtraction of these hot-spots, the remaining silent mutations are randomly distributed. In addition, the percentage of silent mutations among the total in the silent mutation hot-spots is close to that expected for random mutation. We conclude that most of the silent mutations recorded in tumors play no role in tumor development and that the percentage of silent mutation is an indication of the amount of random mutation during tumorigenesis. Silent mutations occur to a significantly different extent in different tumor types. Tumors of the esophagus and colon have a low frequency of silent mutations, tumors of the prostate have a high frequency. PMID- 11106802 TI - Intravenous injection of cycloheximide induces apoptosis and up-regulates p53 and Fas receptor expression in the rat liver in vivo. AB - A single administration of protein synthesis inhibitor, cycloheximide (CHX) induces apoptosis of hepatocytes in vivo. We investigated the underlying mechanisms of this phenomenon and the role of p53 and Fas receptor using terminal dUTP nick end labeling (TUNEL), quantitative reverse transcription polymerase chain reaction and immunohistochemistry. Rat liver tissue specimens were obtained at different time intervals after injection of CHX. The proportion of TUNEL positive apoptotic hepatocytes increased with time and reached a plateau at 2.5h after the injection. The p53 and Fas receptor mRNAs and the proportion of immunoreactive p53-positive and Fas receptor-positive hepatocytes increased markedly with time from 1h after the administration. Since the time course of increased proportion of apoptotic hepatocytes does not parallel that of p53- or Fas receptor-positive hepatocytes and apoptotic hepatocytes appeared prior to up regulation of p53 and Fas receptor expression, it is likely that the enhanced expression of p53 and Fas receptor is not involved directly in CHX-induced apoptosis of hepatocytes in vivo. Rats injected with a single intravenous dose of CHX, however, provide a simple and useful model for investigating the apoptotic machinery and the molecular mechanism of transcriptional up-regulation of p53 and Fas receptor in hepatocytes. PMID- 11106803 TI - Primary mouse fibroblasts deficient for c-Fos, p53 or for both proteins are hypersensitive to UV light and alkylating agent-induced chromosomal breakage and apoptosis. AB - The important regulatory proteins, c-Fos and p53 are induced by exposure of cells to a variety of DNA damaging agents. To investigate their role in cellular defense against genotoxic compounds, we comparatively analysed chromosomal aberrations and apoptosis induced by ultraviolet (UV-C) light and the potent alkylating agent methyl methanesulfonate (MMS) in primary diploid mouse fibroblasts knockout for either c-Fos or p53, or double knockout for both genes. We show that c-Fos and p53 deficient fibroblasts are more sensitive than the corresponding wild-type cells as to the induction of chromosomal aberrations and apoptosis. Double knockout fibroblasts lacking both c-Fos and p53 are viable and were even more sensitive, showing additivity of the chromosomal breakage effects observed in the single knockouts. Regarding the endpoint apoptosis, double knockout fibroblasts displayed a sensitivity similar to c-Fos and p53 deficient cells. The data indicate that (a) both c-Fos and p53 are involved in cellular protection against the clastogenic effect of genotoxic agents, (b) p53 is not required for induction of apoptosis by UV light and MMS, but rather prevents fibroblasts from undergoing apoptotic cell death upon DNA damage, and (c) c-Fos and p53 seem to act independently in determining genotoxic resistance, which is hypothesized to be achieved by impaired DNA repair or differential cell cycle check point control. PMID- 11106804 TI - The in vivo dose rate effect of chronic gamma radiation in mice: translocation and micronucleus analyses. AB - The in vivo effects of chronic, ultra low dose rates of gamma radiation in mice were evaluated using fluorescence in situ hybridization and the in vivo micronucleus test. SWRxC57BL/6 mice were divided into nine exposure groups and continuously exposed to 0.5, 2.0 or 4.0cGy 137Cs per day for 30, 60 or 90 days; unexposed control mice were also included. Following exposure, blood samples were taken from each animal and the frequencies of micronucleated polychromatic erythrocytes (MPCE) and micronucleated normochromatic erythrocytes (MNCE) were determined using flow cytometry. Peripheral blood lymphocytes were cultured and analyzed by chromosome painting to determine translocation frequencies. A significant dose rate response was seen in translocations and both MPCE and MNCE. Comparisons were made between the three chronic dose rates and it was determined that there was no significant difference among translocation frequencies for each rate. However, a significant difference was found between the chronic exposures reported here and the fractionated daily exposures reported previously. Dose rate reduction effects, ranging from 3 at low doses to 14 at high doses, were found for chronic versus acute exposures. The possibility of gender effects was investigated in both micronucleus and translocation data. No gender effect was found in translocation induction, but a slight effect was suggested in micronucleus induction. PMID- 11106805 TI - Protein structural dynamics by single-molecule fluorescence polarization. PMID- 11106806 TI - Single molecule force spectroscopy in biology using the atomic force microscope. AB - The importance of forces in biology has been recognized for quite a while but only in the past decade have we acquired instrumentation and methodology to directly measure interactive forces at the level of single biological macromolecules and/or their complexes. This review focuses on force measurements performed with the atomic force microscope. A general introduction to the principle of action is followed by review of the types of interactions being studied, describing the main results and discussing the biological implications. PMID- 11106807 TI - Mechanical design of proteins studied by single-molecule force spectroscopy and protein engineering. AB - Mechanical unfolding and refolding may regulate the molecular elasticity of modular proteins with mechanical functions. The development of the atomic force microscopy (AFM) has recently enabled the dynamic measurement of these processes at the single-molecule level. Protein engineering techniques allow the construction of homomeric polyproteins for the precise analysis of the mechanical unfolding of single domains. alpha-Helical domains are mechanically compliant, whereas beta-sandwich domains, particularly those that resist unfolding with backbone hydrogen bonds between strands perpendicular to the applied force, are more stable and appear frequently in proteins subject to mechanical forces. The mechanical stability of a domain seems to be determined by its hydrogen bonding pattern and is correlated with its kinetic stability rather than its thermodynamic stability. Force spectroscopy using AFM promises to elucidate the dynamic mechanical properties of a wide variety of proteins at the single molecule level and provide an important complement to other structural and dynamic techniques (e.g., X-ray crystallography, NMR spectroscopy, patch-clamp). PMID- 11106808 TI - The importance of molecular structure and conformation: learning with scanning probe microscopy. AB - Molecular structure holds a key to understanding Nature's intricate design mechanisms and blueprints. If we can understand her blueprints and basic materials, perhaps we can begin to mimic her beautiful products more cost effectively and with less detrimental environmental consequences. Higher resolution instrumentation has allowed us to study single molecules. Indeed, many stellar contributions to the field have come forth in the last couple of years. We can measure the forces required to unravel individual domains of biological molecules such as titin or DNA to a few picoNewtons resolution. This review will attempt to provide a general overview of the field of single molecule analysis using scanning force microscopy. PMID- 11106810 TI - Single-molecule biochemistry coming of age. PMID- 11106809 TI - Twisting and stretching single DNA molecules. AB - The elastic properties of DNA are essential for its biological function. They control its bending and twisting as well as the induction of structural modifications in the molecule. These can affect its interaction with the cell machinery. The response of a single DNA molecule to a mechanical stress can be precisely determined in single-molecule experiments which give access to an accurate measurement of the elastic parameters of DNA. PMID- 11106811 TI - Analysis of genetically modified oils. AB - Genetically modified oils with altered functional or nutritional characteristics are being introduced into the marketplace. A wide array of analytical techniques has been utilized to facilitate developing these oils. This article attempts to review the utilization of these analytical procedures for characterizing both the chemistry and some functionality of these oils. Although techniques to assess oxidative stability in frying and food applications are covered, measurement of nutritional characteristics are not. PMID- 11106812 TI - Chemistry and biochemistry of palm oil. PMID- 11106813 TI - Screening for specific chromosome involvement in hematological malignancies using a set of seven chromosome painting probes. An alternative approach for chromosome analysis using standard FISH instrumentation. AB - We report the application of multi-color fluorescence in situ hydribidization (FISH) for bone marrow metaphase cell analysis of hematological malignancies using a sub-set of the human karyotype for chromosome painting. A combination of chromosome probes labeled with three haptens enabled the construction of a "painting probe" which detects seven different chromosomes. The probe was used to screen three chronic myeloid leukemia (CML) derived cell lines and ten CML patient bone marrow samples for aberrations, additional to the Ph rearrangement, that are associated with the onset of blast crisis of CML. This approach was shown to identify karyotype changes commonly seen by conventional karyotyping, and in addition revealed chromosome changes unresolved or undetected by conventional cytogenetic analysis. The seven-color painting probe provides a useful, fast, and reliable complementary tool for chromosome analysis, especially in cases with poor chromosome morphology. This is a simple approach, since the probes can be displayed in a standard red/green/blue format accessible to standard fluorescence microscopes and image-processing software. The proposed approach using panels of locus-specific probes as well as chromosome paints will be useful in all diagnostic routine environments where analysis is directed towards screening for genetic rearrangements and/or specific patterns of chromosome involvement with diagnostic/prognostic value. PMID- 11106814 TI - TEL/AML-1 fusion gene. its frequency and prognostic significance in childhood acute lymphoblastic leukemia. AB - TEL gene rearrangement due to the 12;21 chromosome translocation is believed to be the most common molecular genetic abnormality in childhood acute lymphoblastic leukemia (ALL). A study was conducted to investigate the frequency and prognostic significance of TEL/AML-1 fusion gene resulting from a cryptic t(12;21)(p13;q22). Bone marrow samples from 86 patients diagnosed over the past 5 years at Columbus Children's Hospital were analyzed by fluorescence in situ hybridization (FISH) technique for TEL/AML-1 fusion gene, using LSI((R)) DNA probes. The positive cases were analyzed for clinical outcome. Patients in this study received treatment according to Children's Cancer Group (CCG) protocols. Fifteen of the 86 cases (17%) were positive for the fusion gene. All were B-cell lineage and except for one, all were CD10 positive. TEL/AML-1 was not found in any T-cell ALL. The mean overall survival (OS) following diagnosis for the TEL/AML-1-positive group was significantly longer than for the TEL/AML-1-negative group by log-rank = 7.84, P = 0.005. Similarly, the event-free survival (EFS) after remission for the positive group (median 94.5 months) was longer than the negative group (median 57 months) by log-rank = 7.19, P = 0.007. This study confirms that the TEL/AML-1 fusion gene may be the most common genetic event in childhood ALL, occurring in 17% of the patients. It appears restricted to the B-cell lineage. In this study, the presence of a TEL/AML-1 fusion gene was statistically significant in predicting both OS and EFS, indicating a favorable clinical outcome for these patients. Screening for TEL/AML-1 should become routine at diagnosis and a useful biological variable for risk stratification in future clinical trials. PMID- 11106815 TI - Translocation (2;8)(p12;q24) associated with a cryptic t(12;21)(p13;q22) TEL/AML1 gene rearrangement in a child with acute lymphoblastic leukemia. AB - We report a case of childhood acute lymphoblastic leukemia with the simultaneous occurrence of a t(2;8)(p12;q24) typically associated with mature B cell or Burkitt leukemia, and a t(12;21)(p13;q22) exclusively associated with pre-B cell ALL. The lymphoblasts were characterized as L2 morphology by the French-American British classification. However, there were atypical morphologic findings for L2 ALL, including vacuolization in some cells. The lymphoblasts were periodic acid Schiff positive and myeloperoxidase negative. Immunophenotypic analysis revealed that the majority of lymphoblasts were TdT+, CD10+, CD19+, CD20-, and cytoplasmic mu+. These features were consistent with an immature pre-B cell leukemia phenotype with some characteristics of a mature B-cell leukemia. A t(2;8)(p12;q24)(p12;q24), characteristic of mature B-cell leukemia or Burkitt type leukemia, was detected by conventional cytogenetics with no other cytogenetic abnormalities. However, diagnostic peripheral blood and bone marrow specimens demonstrated simultaneous occurrence of a cryptic t(12;21)(p13;q22) by both FISH and RT-PCR. The simultaneous occurrence of these translocations in a pediatric patient have implications for the pathogenesis of leukemias with t(2;8)(p12;q24) as well as t(12;21)(p12;q22). Analysis of additional cases of leukemia with translocations involving the MYC locus on 8q24 will be required to determine the frequency of association with the cryptic t(12;21)(p13;22), and the prognostic significance of the simultaneous occurrence of the translocations. PMID- 11106816 TI - Variant intra philadelphia translocation with rearrangement of BCR-ABL and ABL BCR within the same chromosome in a patient with cALL. AB - A unique variant Philadelphia translocation accompanied by the loss of the short arm of chromosome 9 in a 32-year-old female with common acute lymphoblastic leukemia (cALL) is described. Furthermore, supernumerary chromosome 8 material was found as an insertion into the long arm of chromosome 2 and/or as ring chromosomes in addition to two normal chromosomes 8. The chromosomal abnormalities were identified by combined conventional chromosome banding analysis and fluorescence in situ hybridization (FISH). The BCR-ABL rearrangement was confirmed by FISH and reverse transcriptase-polymerase chain reaction (RT PCR) studies. Possible mechanisms leading to this variant intra Philadelphia translocation are discussed. The aberrations found have prognostic implications, because 9p anomalies confer an adverse effect to the already poor prognosis of Philadelphia-positive ALL. PMID- 11106817 TI - Molecular and cytogenetic analysis of glioblastoma multiforme. AB - Glioblastoma multiforme (GBM) is the most common primary tumor occurring in the central nervous system of adults. Although progress has been made in clinical management of this tumor, little is known about the molecular defects underlying the initiation and progression of GBM. To address these issues, we have characterized five cases of GBM using cytogenetics, comparative genomic hybridization (CGH), fluorescence in situ hybridization (FISH), and direct sequencing. All of these tumors were observed to have clonal chromosome aberrations. Complicated chromosome translocations including der(18)t(2;4;12;18), der(X)t(X;10)(q27.1;p12.1) and der(10)t(10;15)(p11.23;q11.2), and der(1) (:1p31- >1q44::7q11. 3-->7qter) were seen in three tumors. Loss of the CDKN2 gene was noted in four tumors. A gain of copy number of the Cathepsin L gene was seen in two tumors. Amplification of the CDK4, MDM2, and GLI/CHOP genes was noted in two tumors, and amplification of the PDGFR gene was detected in one tumor. Mutation of exon 5 of the TP53 gene was found in three tumors. No mutation of the BCL10 gene was detected in five cases of GBM analyzed, although deletion of chromosome 1p was seen in two tumors. These results provide information for further investigation of GBM. PMID- 11106818 TI - Evaluation at single cell level of residual Philadelphia negative hemopoietic stem cells in chronic phase CML patients. AB - In chronic myeloid leukemia, accurate determination of Ph(-) Hemopoietic stem cells (HSC) in peripheral blood (PB), bone marrow (BM) and leukapheresis products is important for the selection of patients for whom mobilization, collection, and autografting of Ph(-) HSC are envisaged. To this effect, the BCR/ABL fusion was assessed at the single cell level in 25 sets of PB and BM samples using dual color I-FISH in immunophenotyped CD34(+) cells and RT-PCR of individual CFU-GM colonies. In 15 cases found to be 100% Ph(+), the respective BCR/ABL gene was absent in 30% of CD34(+) cells, while the respective transcripts could not be identified in 17% of CFU-GM. The mean percentage of BCR/ABL(-) CD34(+) cells and CFU-GM cells was higher (38% and 29%, respectively) in untreated patients than in treated patients (24% and 7%, respectively). In eight cases with cytogenetic response (CgR), the percentage of Ph(-) metaphases correlated with the level of BCR/ABL(-) colonies in BM and PB and with the proportion of BCR/ABL(-) CD34(+) cells in the BM. Immunophenotyping and FISH was fast, easy, always informative, and quantitative for the BCR/ABL(-) CD34(+) cells. Our results show that (a) at early diagnosis a high frequency of BCR/ABL(-) HSC circulate in the PB and that Ph(-) hematopoiesis is not completely suppressed; (b) although normal clonogenic cells decline rapidly within a few months after diagnosis, appreciable numbers of normal CD34(+) cells survive in chronic phase, especially in patients with CgR. PMID- 11106819 TI - Chromosome abnormalities in malignant melanoma: clinical significance of nonrandom chromosome abnormalities in 206 cases. AB - We report the cytogenetic abnormalities from a series of 206 primary malignant melanoma specimens referred to a single institution. A total of 169 out of 206 unique cases had chromosome breakpoints. A previously described statistical method was used to detect nonrandom distribution of chromosome breakpoints at the level of chromosome regions. Nonrandom occurrence of chromosome breakpoints (indicating that the observed number of breaks significantly exceeded the expected number of breaks) was detected in 28 regions, suggesting a hierarchy of genetic abnormalities in melanoma. Clinical variables and tumor characteristics were analyzed for associations with the presence of any nonrandom chromosome breakpoints; with individual, nonrandomly involved chromosome regions; and with paired, nonrandomly involved chromosome regions. No nonrandomly involved chromosome regions or pairs of regions appeared to significantly affect survival. These results identify recurring, nonrandom chromosome abnormalities in malignant melanoma. These results suggest that recurring, nonrandom chromosome alterations play a key role in the etiology and/or progression of malignant melanoma and identify targets within the genome for molecular genetic studies. PMID- 11106820 TI - A simple strategy for breakpoint fragment determination in chronic myeloid leukemia. AB - Molecular characterization is considered a part of the routine work-up of chronic myeloid leukemia (CML) cases. Southern blot analysis using the universal BCR (UBCR) probe on BglII-digested DNA samples is the most commonly used technique, while employing the human 3' bcr probe (PR-1) is usually considered a complementary tool. In this study, we tried to develop a simple and economic strategy for molecular characterization of CML using the 3' probe as it has been shown to be the one capable of locating the breakpoint site. Seventy-eight cases of CML were studied. Molecular analysis was performed using the Southern blot technique. DNA was digested with Bam HI, BglII, EcoRI, and XbaI. Hybridization was performed using the human 3' bcr (PR-1) probe. BamHI and BglII could differentiate fragment 1 (F1) showing rearrangement (R) with Bam HI and germline configuration (G) with BglII; F2/3 showing R with both, and F4 showing R with BamHI and G with BglII. F2/3 cases were further divided by HindIII enzyme into F2 showing (G) and F3 showing (R). Fragment 0 showed G with both, but R with EcoRI and/or XbaI, while 3' deletion gave G with all four enzymes. Our results showed a relative incidence of 6.4% for F0, 20.5% for F1, 32.1% for F2, 19.2% for F3, 15.4% for F4, and 6.4% for 3' deletion. Sixty cases were evaluated clinically and hematologically and were followed up for disease evolution and survival. They included 32 cases in early chronic phase, 24 in late chronic phase, two in acceleration, and two in blastic crisis. No significant correlation was encountered between the breakpoint site and any of the clinical and hematological data except those patients with 3' deletion who showed a very short survival. The study emphasizes Southern blotting as the method of choice for molecular characterization of CML and offers a simple and economic strategy for diagnosis and determination of breakpoint fragment. PMID- 11106821 TI - Primary CD30/Ki-1 positive anaplastic large cell lymphoma of skeletal muscle with der(17)t(1;17)(q11;p11). AB - CD30/Ki-1 positive anaplastic large cell lymphoma (Ki-1 ALCL) frequently exhibits extranodal disease and chromosomal t(2;5)(p23;q35). An 11-year-old girl presented with an intramuscular tumor of the right upper arm. Tumors of the chest wall, left arm and leg, hepatomegaly, pleural effusion, and enlarged lymph nodes then developed. The intramuscular tumor and pleural effusion showed a diffuse infiltration of large atypical cells with abundant amphophilic cytoplasms. The tumor cells were positive for CD30, CD2, CD45RO, and p80, but were negative for other T-cell, B-cell, and myeloid cell antigens. She was diagnosed as having Ki-1 ALCL with a T-cell origin. Cytogenetic studies showed an abnormal karyotype including a der(17)t(1;17)(q11;p11). She received seven cycles of intensive chemotherapy followed by an autologous peripheral blood stem cell transplantation, and has been in complete remission for more than two years. The primary involvement of skeletal muscle is quite uncommon in ALCL, and an abnormal karyotype including t(1;17)(q11;p11) has not been reported previously. Since a high frequency of aberrations of 1p36/1q12 or 17p13.3 was detected in sarcoma cells, the presence of suppressor genes is suggestive in these sites. PMID- 11106822 TI - Sister chromatid exchanges in peripheral lymphocytes from women with carcinoma of the uterine cervix. AB - Sister chromatid exchanges (SCE) are reciprocal exchanges between sister chromatids. It has been reported that in patients with cervical cancer, the frequency of SCE in peripheral lymphocytes is significantly higher than that in normal individuals; however, other studies have shown no significant difference. The aim of this unmatched case-control study was to compare the mean number of SCE per metaphase in lymphocytes from women with and without carcinoma of the cervix uteri. The SCE specimens were prepared by the fluorescence plus giemsa technique in peripheral lymphocytes from 28 women with carcinoma of cervix uteri and 28 controls. The mean number of SCE per metaphase in women with carcinoma of cervix uteri (7.80 +/- 1.05) was higher than the control group (6.98 +/- 1.13) (P < 0.05; t-test). This study had a statistical power of 0.80 and an alpha value of 0.05. This finding suggests that an increased number of SCE in peripheral lymphocytes is associated with cervical cancer. We consider that the lack of reported association of SCE and cervical cancer might be attributed to the none determination of the statistical power and sample size. PMID- 11106823 TI - Prognostic value of tumoral ploidy in a series of spanish patients with acute lymphoblastic leukemia. AB - The prognostic value of tumoral ploidy and its relation with nonrandom chromosome alterations were analyzed in 89 cases of acute lymphoblastic leukemia. Ploidy is associated with the number of nonrandom chromosome alterations and survival. The pseudodiploid and hypodiploid groups had a high incidence of nonrandom alterations and poor survival while the diploid and high hyperdiploid groups had a lower incidence of nonrandom alterations and longer survivals. The patients in the low hyperdiploid group with random alterations received the same treatment as the diploid and high hyperdiploid groups but had poor survivals. Our analysis confirms that ploidy is a very important prognostic factor and suggests that patients with low hyperdiploidy should receive intensive therapies similar to those of patients in the groups with a high number of nonrandom alterations. PMID- 11106824 TI - Body mass, age, and the APC I1307K allele in Ashkenazi Jewish prostate cancer patients. AB - The I1307K mutation of the adenopolyposis coli gene (APC), located on chromosome 5q21-q22, is associated with an increased risk of cancer in Ashkenazi Jews. In the present study, we analyzed age and body mass of Ashkenazi Jewish prostate cancer patients, with and without the APC I1307K mutation. Participants in our study were found through urology and radiation oncology clinics, and all eligible patients were asked to take part. A familial history was obtained by interview or self-report questionnaire. Histological confirmation of diagnosis was obtained for all subjects. The I1307K allele of the APC gene was detected by amplification of lymphocyte DNA from peripheral blood according to standard polymerase chain reaction (PCR) and dot blot procedures. We studied 135 Ashkenazi Jewish men with prostate cancer. The youngest was 49, the oldest 80, average age 68 +/- 6.88 (mean +/- SD). The older patients carrying the wild type APC allele tended to have a lower body mass than the younger ones (r = -.27, P =.002). Of 71 patients under 70 years old, 65 carried the wild type APC allele, and had a body mass index of 28. 7 +/- 4.23 kg/m(2). The six men under age 70 carrying the I1307K APC allele had a body mass index of 26.87 +/- 1.44 kg/m(2). The difference in body mass index of the two groups is significant (P =. 032, t test for unequal variance). Increased body mass is a prostate cancer risk factor, and hereditary prostate cancer is associated with younger patients. Therefore, our finding, that patients under age 70 carrying the I1307K allele are significantly thinner than those carrying the wild type allele, suggests that the APC I1307K allele is also a prostate cancer risk factor. Our results are in accord with other studies indicating that APC mutations increase the risk of prostate cancer. PMID- 11106825 TI - Myelodysplastic syndrome and a nonrandom chromosomal abnormality t(1;19): an indolent pathologic entity. AB - The chromosomal abnormality t(1;19) is an infrequent finding in adult hematopoietic malignancies. This is only the second report of t(1;19) in association with myelodysplastic syndrome in which there was an apparent excellent response to oral cyclosporin A and a very indolent clinical course. PMID- 11106826 TI - CALM-AF10 fusion gene in leukemias: simple and inversion-associated translocation (10;11). AB - A translocation (10;11)(p12;q14) was observed in two children, one with acute eosinophilic leukemia and the other with acute T-cell lymphoblastic leukemia. The presence of CALM-AF10 fusion was ascertained by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Fluorescence in situ hybridization (FISH) analysis showed that AF10 gene splitting was associated with partial inversion of chromosome 11 in the first patient. In addition, FISH analysis also determined the orientation of the CALM gene, 5' telomere to 3' centromere on 11q. PMID- 11106827 TI - Identification of two new translocations that disrupt the AML1 gene. AB - The AML1 gene, located at chromosome 21q22, encodes a component (CBFalpha2) of a heterodimeric transcription factor complex termed core binding factor (CBF), which binds to DNA and activates gene expression. Chromosomal rearrangements may lead to disruption of this gene and development of acute leukemia. Twelve AML1 translocations have been identified to date, and include sites on chromosomes 1, 2, 3, 5, 8, 12, 14, 15, 16, 17, 18, and 19. Here we report two new translocations involving AML1 in acute myeloid leukemia, in which the disruption of the AML1 gene was documented by GTG banding cytogenetic studies and metaphase and interphase FISH analysis. These chromosomal breakpoints identified as harboring new fusion partners for AML1 are at 2p11.2 and 20q13.1. The two patients in who these translocation were identified were elderly males with newly diagnosed AML. These patients shared the same poor outcomes reported for other rare AML1 translocations. PMID- 11106828 TI - Low grade fibromyxoid sarcoma. a further low-grade soft tissue malignancy characterized by a ring chromosome. AB - Supernumerary rings in the context of a simple karyotype characterize several low grade malignant tumors of soft tissue and bone. Low-grade fibromyxoid sarcoma is an uncommon low-grade sarcoma, the cytogenetics of which has not yet been reported. Here we describe the first molecular-cytogenetic characterization of a pulmonary metastasis of low-grade fibromyxoid sarcoma. The histology of the primary and recurrent tumors was consistent with the diagnosis of low-grade fibromyxoid sarcoma of the usual type, whereas the pulmonary metastasis was of the "giant rosettes" variant. Cytogenetic analysis revealed a ring chromosome. Because gain of material of chromosomes 7 and 16 was detected by CGH, the ring chromosome is assumed to be composed of material from these respective chromosomes. PMID- 11106829 TI - Complex rearrangement of chromosomes 6 and 11 as the sole anomaly in atypical teratoid/rhabdoid tumors of the central nervous system. AB - Atypical teratoid/rhabdoid tumor of the central nervous system is a rare childhood tumor with a distinct histologic appearance and an aggressive clinical course. Few tumors have been analyzed cytogenetically. The only consistent chromosomal abnormality identified in some of these tumors has been monosomy or deletions of chromosome 22; in others, a normal chromosome 22 was present. The authors report an atypical teratoid/rhabdoid neoplasm of the central nervous system with a novel complex rearrangement affecting chromosomes 6 and 11 as the sole anomaly. The involvement of region 11p15 could be important in the pathogenesis of this entity. PMID- 11106830 TI - Five fold increase of insulin concentration delays the absorption of subcutaneously injected human insulin suspensions in pigs. AB - Human NPH U100 (100 IU/ml), given once daily, is often absorbed too fast to cover the basal insulin demand throughout 24 h. The aim of the present study was to examine whether the absorption of human insulin suspensions would be delayed by increasing the insulin concentration from U100 to U500. In each experiment 10 IU of corresponding human Ul00 and U500 preparations, labelled with 125I-insulin, were injected subcutaneously and contralaterally in the neck of a pig followed by monitoring residual radioactivity over the injection sites. The time until 75, 50 and 25% residual radioactivity (T(75%), T(50%) and T(25%)) using NPH U500 was compared with NPH U100 in 14 experiments: T(75%): 5.0+/-0.5 (mean+/-SEM) vs 3.8+/ 0.3 h (P=0.007, paired t-test), T(50%): 12. 2+/-0.9 vs 9.0+/-0.6 h (P=0.003) and T(25%). 24.2+/-1.2 vs 17.9+/-1. 2 h (P=0.001). The corresponding values for semilente U500 compared with semilente U100 in eight experiments were: T(75%): 2.8+/-0.4 vs 1.6+/-0.2 h (P=0.02), T(50%): 5.6+/-0.6 vs 3.4+/-0.3 h (P=0.01) and T(25%): 10.9+/-1.1 vs 7.2+/-0.7 h (P=0.009). Thus, the absorption of a given dose of human NPH or human semilente insulin in pigs is substantially delayed by changing the insulin concentration from Ul00 to U500. Human NPH U500 appears to be more appropriate than human NPH U100 for injection once daily in basal insulin therapy. PMID- 11106831 TI - Identification of glucokinase mutation in subjects with post-renal transplantation diabetes mellitus. AB - Mutations in the glucokinase (GCK) gene are considered to be a possible cause of maturity-onset diabetes of the young. The purpose of this study was to evaluate the contribution of this gene to the development of post-renal transplantation diabetes mellitus (PTDM). Identification of the GCK mutation was attempted in 58 selected renal allograft recipients with PTDM and 45 normal controls. The exons in the GCK gene were examined using polymerase chain reaction (PCR), followed by an analysis of single-stranded DNA conformational polymorphism (SSCP). The abnormal bands were then confirmed by DNA sequencing analysis. The family members of the patients affected with GCK mutation were also examined. Two of the 58 PTDM patients (3. 4%) were found to have GCK mutations. One had the mutation on exon 5 and the other on intron 7. One control subject had the mutation on intron 9. The mutation on exon 5 was identified as a substitution of CCT (proline) for CTT (leucine) at codon 164, which has never been reported before. The family members of the PTDM patient with a mutation on exon 5 were analyzed by PCR, followed by SSCP, and two of them had the same mutation. The abnormal band seen on SSCP analysis of exon 7 was identified as the C-->T substitution at the 39th nucleotide in intron 7. Two of the family members also displayed the same bands on the SSCP. One of the 45 normal controls had a known polymorphism located at the 8th nucleotide in intron 9. We found a GCK mutation on the exon in subjects with PTDM and we speculate that this mutation may be one of the possible contributing factors of PTDM, although variations of the GCK gene are not common causes of PTDM. PMID- 11106832 TI - Homocysteine and endogenous markers of renal function in type 2 diabetic patients without coronary heart disease. AB - The aim of this study was to assess parameters of renal function and other determinants of plasma homocysteine in type 2 diabetic patients without coronary heart disease (CHD). Fasting plasma homocysteine, serum cystatin C and serum creatinine were determined in 183 (75 men, 108 women) Type 2 diabetic patients without clinical evidence of CHD. Creatinine clearance was calculated and parameters such as blood pressure, body mass index (BMI), and glycated haemoglobin (HbA(1c)) were assessed. The urine albumin:creatinine ratio was used to classify patients as normo-, micro- or macroalbuminuric. One hundred and ten patients were normoalbuminuric, 67 patients were microalbuminuric and six patients were macroalbuminuric. There was no statistically significant difference in plasma homocysteine concentration between patients with normoalbuminuria and microalbuminuria. There was a trend towards increasing plasma homocysteine with decreasing glomerular filtration rate (GFR) (r=-0.46; P<0.0001). There was statistically significant correlation between plasma homocysteine and age (r=0.37), serum cystatin C (r=0.47), and serum creatinine (r=0.56). Plasma homocysteine concentration was significantly higher in patients with BMI<30 kg/m(2) and showed significant inverse correlation with weight (r=-0.16; P=0.03) and body mass index (r=-0.24; P=0.001). Homocysteine and serum creatinine were significantly higher in males than females and higher in smokers than non smokers but was not associated with glycemic control and duration of diabetes. In conclusion, elevated homocysteine concentration in patients with type 2 DM without CHD is related to age, gender, smoking, BMI and GFR. Follow up studies will provide further information on the association between hyperhomocysteinemia and the development of cardiovascular disease. PMID- 11106833 TI - Sex differences in the growth of diabetic children. AB - To investigate the effect of diabetes on the growth of children and to detect possible impairment of their final height, 58 Chinese subjects (22 boys; 36 girls) with childhood diabetes were studied longitudinally from diagnosis. Mean onset age at presentation was 8. 70 years. All patients were measured and weighed at diagnosis and every 3-4 months during the follow up period. Their height and weight measurements were converted to standard deviation scores (S.D. S.) using normal height and weight-for-height reference standards for Chinese children established in Hong Kong. The mean height S.D.S. for boys and girls at diagnosis were +0.76 and -0.07 (P=0.015). The mean final height S.D.S. for boys and girls were +0.14 and -0.57. The final heights of girls were significantly shorter than their target heights. At attainment of final height for diabetic girls, their mean weight-for-height S.D.S. was +0.76 indicating that they tended to become overweight. This study demonstrates the sex differences in the growth of diabetic children. Diabetic boys were tall for age at presentation but they achieved average final heights while diabetic girls attained below average adult stature and they tended to become obese. PMID- 11106834 TI - Detection of the association between a deletion polymorphism in the gene encoding angiotensin I-converting enzyme and advanced diabetic retinopathy. AB - We investigated the relationship between advanced diabetic retinopathy (ADR) and an angiotensin-converting enzyme (ACE) gene polymorphism in subjects with type 2 diabetes and ADR, pre-proliferative (PrePDR) or proliferative diabetic retinopathy (PDR) without overt nephropathy. Polymerase chain reactions were used to detect insertion/deletion (I/D) polymorphisms of the ACE gene. There was no difference in the frequency of II, ID, or DD genotypes, or of I and D alleles among subjects with type 2 diabetes without diabetic retinopathy (NDR) or with simple diabetic retinopathy (SDR) and non-diabetic controls. There was also no difference in the frequency of ACE genotypes among subjects with type 2 diabetes with NDR, or SDR and ADR. However, the frequency of the ACE DD genotype in ADR was significantly higher than that in controls (chi(2)=6.64, P=0.036). On the other hand, the frequency of the D allele in ADR was significantly higher than that in controls (chi(2)=6.33, P=0.012), NDR (chi(2)=4.18, P=0.041) and SDR (chi(2)=4. 89, P=0.027), respectively. These results indicate a significant relationship between the presence of the D allele polymorphism in the ACE gene and ADR in Japanese subjects with type 2 diabetes and no overt nephropathy. PMID- 11106835 TI - Cognitive dysfunction in older subjects with diabetes mellitus: impact on diabetes self-management and use of care services. All Wales Research into Elderly (AWARE) Study. AB - OBJECTIVE: To determine whether cognitive impairment is associated with changes in self-care behaviour and use of health and social services in older subjects with diabetes mellitus. RESEARCH DESIGN AND METHODS: This was a community based, case-control study of subjects registered with general practices participating in the All Wales Research into Elderly (AWARE) Diabetes Study. The 396 patients aged 65 years or older with known diabetes mellitus were compared with 393 age- and sex-matched, non-diabetic controls. Adjusted odds ratio estimates of normal performance on Mini-Mental State Examination (MMSE) and Clock Drawing Test (numbers and hands) were determined. Information on self-care behaviours and use of services was obtained. RESULTS: A total of 283 (71%) diabetic subjects scored 24 or more on MMSE, compared with 323 (88%) of controls (OR 0.54, P<0.0005). The mean (S.D.) scores were 24.5 (5.1) and 25.7 (4.3), respectively (difference between means 1.22; 95% CI 0.56, 1.88; P<0. 001). Clock testing demonstrated that 257 (65%) and 286 (72%) diabetic subjects correctly placed the numbers and hands, respectively, compared with 299 (76%) and 329 (84%) of controls (OR 0.59, P<0.001 and P<0.52, P<0.0005, respectively). Both test scores declined with increasing age, earlier school leaving age and deteriorating visual acuity. Of other variables examined, only need for oral hypoglycaemic drugs or insulin, history of stroke, dementia or Parkinson's disease and symptoms of autonomic neuropathy significantly impaired one or more cognitive test scores. The odds ratios (95% CI) for normal cognitive test results in subjects with diabetes after adjusting for all significant variables was 0.74 (0. 56, 0.97), P=0.029 for MMSE scores and 0.63 (0.44, 0.93), P=0.019, and 0.58 (0.38, 0.89), P=0.013, for the numbers and hands parts of the clock test, respectively. In comparison with diabetic subjects with no evidence of cognitive impairment, diabetic subjects with an MMSE score <23 were significantly less likely to be involved in diabetes self-care (P<0.001) and diabetes monitoring (P<0.001). A low MMSE score was also significantly associated with higher hospitalisation in the previous year (P=0.001), reduced ADL (activities of daily living) ability (P<0.001) and increased need for assistance in personal care (P=0.001). CONCLUSIONS: Elderly subjects with predominantly Type 2 diabetes mellitus display significant excess of cognitive dysfunction, associated with poorer ability in diabetes self-care and greater dependency. Routine screening of cognition in older subjects with diabetes is recommended. PMID- 11106836 TI - Oxidative protein damage in early stage Type 1 diabetic patients. AB - To examine the influence of oxidative stress on oxidative protein damage, we studied 51 young Type 1 diabetic patients clinically free of complications and 48 healthy normolipidaemic age-matched controls. We determined: (1) plasma carbonyl (PCO), plasma total thiol (T-SH), and nitrotyrosine (NT) levels as markers of oxidative protein damage; (2) plasma lipid hydroperoxide (LHP), and nitric oxide (NO) levels as markers of oxidative stress; (3) plasma total antioxidant capacity (TAO), ceruloplasmin (Cp), transferrin (TRF), unsaturated iron binding capacity (UIBC), erythrocyte glutathione (GSH), and erythrocyte superoxide dismutase (SOD) as markers of free radical scavengers. There were no significant differences in the levels of these markers between prepubertal diabetic patients and the controls. The levels of both of PCO and LHP were increased in adolescent and young adult Type 1 diabetic patients with respect to their controls. In the adolescent group, patient versus control values for PCO were 1.04+/-0.067 versus 0.67+/-0.0274 nmol/mg and for LHP they were 2. 10+/-1.09 versus 1.00+/-0.4 nmol/mg. In the young adult group, patient versus control values for PCO were 0.99+/-0.054 versus 0. 66+/-0.02 nmol/mg and for LHP they were 1.96+/-0.78 versus 1.15+/-0. 4 nmol/mg. TAO levels were significantly decreased in adolescent diabetic patients compared to their controls (0.92+/-0.27 vs. 1. 86+/-0.37) and in young adult diabetic patients compared to their controls (0.80+/-0.27 vs. 2.11+/-0.54 nmol/mg). T-SH was not different between diabetic patients and the controls. Serum NT, NO, and erythrocyte SOD levels were not different either between three groups of diabetic patients or between the patients and their controls. We attribute this lack of difference to limited disease duration. Changes in markers of oxidative stress other than NT, NO, and SOD observed in adolescent and young adult early stage Type 1 diabetic patients contribute to the imbalance in the redox status of the plasma. We attribute this imbalance to metal catalyzed protein oxidation in both groups of Type 1 diabetic patients clinically free from complications. PMID- 11106837 TI - Correlation among fasting plasma glucose, two-hour plasma glucose levels in OGTT and HbA1c. AB - A study was made on the association among 2-h plasma glucose (PG) in oral glucose tolerance test (OGTT), fasting plasma glucose (FPG) using correlation and regression equation. Subjects were 13174 OGTT examinees tested between 1980 and 1998. Blood glucose was determined by the glucose oxidase method and glycated hemoglobin (HbA1c) by the HPLC method. As for correlation between 2-h PG and FPG, regression equation of the <60 year group was y=57.1+0.336x (r=0.866, P<0.0001) and that of the >==60 year group was y=61.5+0.286x (r=0.814, P<0. 0001). FPG was calculated at 124.3 in the <60 year group and 118.7 mg/dl in the >==60 year group for 2-h PG of 200 mg/dl, 2-h PG were calculated at 199.5 and 210.7 mg/dl for FPG of 126 mg/dl, respectively. In the <60 year group, FPG were calculated at 121.7 and 124.4 mg/dl and 2-h PG at 193.2 and 199.3 mg/dl for HbA1c of 6.0 and 6.1%, respectively. As for associations between HbA1c and FPG or 2-h PG being high correlation, it is possible to estimate a prevalence of DM in a group using HbA1c>==6.1%. High correlations were demonstrated among all the three measures; FPG, 2-h PG, HbA1c. If 2-h PG is used in diagnosing diabetes mellitus, an FPG of 126 mg/dl proposed by ADA and World Health Organization (WHO) as a diagnostic level of FPG is an acceptable value for the Japanese. PMID- 11106839 TI - Alkaloid levels in Duncecap (Delphinium occidentale) and Tall larkspur (D. barbeyi) grown in reciprocal gardens: separating genetic from environmental influences. AB - The objective of this study was to determine whether differences in toxicity between Tall larkspur (Delphinium barbeyi) and Duncecap larkspur (D. occidentale) were genetically inherent within the species, or due to environmental influences unique to the different regions where they grow. There was no difference in the concentration of the toxic alkaloids between the two species when grown in common gardens. However, both species had higher concentration of toxic alkaloids when grown in the southern Rocky Mountain region which is characterized by summer monsoonal thunderstorms, compared to summer drought in the north. In a unique occurrence where Duncecap larkspur grew beside Tall larkspur, toxic alkaloids were very low in Duncecap larkspur, but total alkaloid concentration was often higher than in Tall larkspur. PMID- 11106838 TI - Antihypertensive drugs as predictors of type 2 diabetes among subjects with impaired glucose tolerance. AB - AIMS: to examine the incidence rate of progression to Type 2 diabetes and baseline prognostic risk factors, focusing on hypertension and antihypertensive medication, in a cohort (n=207) with impaired glucose tolerance (IGT). METHODS: after 2 and 4.6 (1. 9-6.4) years new cases of diabetes were diagnosed by the oral glucose tolerance test (OGTT). Hypertension (BP 160/95 or antihypertensive medication) was included in multiple regression analyses to assess the effect of risk factors on the development of diabetes. RESULTS: diabetes developed in 32 subjects (19%), an incidence of 41/1000 (95% CI 28-57/1000) person-years. In univariate analyses, progression to diabetes was associated with a high (>9.0 mmol/l) 2-h OGTT value (P=0.008), a high fasting insulin (>12.0 mU/l) level (P=0.000), a high triglyceride (>/=1.3 mmol/l) level (P=0.028), a high BMI (>/=28.0 kg/m(2)) (P=0.013) and hypertension (P=0.003). The risk for the development of diabetes was not increased in hypertensive subjects without antihypertensive medication compared with normotensive subjects (OR 0.8, 95% CI 0.3-2. 6). However, it was increased in subjects with on medication, especially diuretics alone or in combination with other drugs. Hypertensive subjects on diuretics had higher levels of fasting insulin and triglycerides and higher BMIs at baseline than normotensive subjects. After adjustment for 2-h OGTT, fasting insulin, triglycerides and BMI, the OR for diabetes was 7.7 (95% CI 2.1-28.2) in hypertensive subjects using diuretics alone or in combination with other drugs and 2.6 (95% CI 1.0-6.7) in those using other drugs compared with normotensive subjects. The OR of diabetes corresponding to a one-unit increase in the 2-h OGTT concentration was 2.5 (95% CI 1.6-4.0) in the whole cohort. CONCLUSIONS: the rate of progression from IGT to Type 2 diabetes in this population was similar to that seen in other studies among Caucasian populations. The use of antihypertensive medication, especially diuretics, and a high 2-h OGTT level were significant predictors of subsequent deterioration to diabetes. PMID- 11106840 TI - Species discrimination and population differentiation in ants using microsatellites. AB - This study was conducted to establish the regional scale of population differentiation of ants in the wheat belt of central western New South Wales. Microsatellite variation was surveyed at five loci in two morphologically similar ant species (designated "A" and "B") from the Camponotus ephippium complex. Three of the five scored microsatellite loci were highly variable with totals, in the two species, of 11, 13 and 42 alleles. The other loci had two and three alleles. The mean number of alleles per locus per sample ranged from 2.0 to 4.6 for species A and from 1.4 to 3.8 in species B. Mean observed heterozygosity was 0.385 for species A and 0.363 for species B. The geographic distribution of genotypes was significantly non-random for all tested loci in both species. Eight of 47 alleles in species A and 15 of 28 in species B were restricted to a single site. Allelic accumulation percentages were calculated for several orderings of samples - level of heterozygosity, sample size and geographic position. In all orderings three or more samples must be included for more than three-quarters of alleles to be represented. PMID- 11106841 TI - Quinolizidine alkaloids from Genista ephedroides. AB - Alkaloids retamine, anagyrine, lupanine, 17-oxoretamine, 12-alpha-hydroxylupanine were detected, along with four others unidentified compounds in the aerial parts of Genista ephedroides D.C. PMID- 11106842 TI - Non-toxic pyrrolizidine alkaloids from Eupatorium semialatum. AB - The leaves of Eupatorium semialatum were investigated for the occurrence of pyrrolizidine alkaloids. Although this type of alkaloids generally occurs in the Eupatorieae, only unusual non-toxic pyrrolizidines of the tussilagin type were identified. All compounds are methyl esters of the corresponding beta-amino acids. PMID- 11106843 TI - Exudate flavonoid aglycones in the alpine species of Achillea sect. Ptarmica: Chemosystematics of A. moschata and related species (Compositae-Anthemideae). AB - In completion of our studies on Achillea exudate flavonoids, 11 alpine species of Achillea sect. Ptarmica were analyzed for their aglycone profiles. The study focuses on species commonly associated with A. moschata. The major flavonoid constituents found in exudates of most taxa were 6-hydroxyflavonol 3,6,4' trimethyl ethers, except in A. ageratifolia and its subspecies, which are characterized by the accumulation of the 3,6,7-trimethoxy and 6-hydroxyflavones. Infraspecific variation was particularly high in A. abrotanoides and A. moschata. Results are discussed in relation to published data for related species and within the context of evolutionary aspects in the genus Achillea. The 3,6,4' trimethoxy substitution is regarded as a basic chemical trend within the genus Achillea. Geographical and ecological aspects are briefly addressed, and a summary on known exudate aglycone composition of species from all sections of Achillea is included. PMID- 11106844 TI - Capsaicinoid profiles are not good chemotaxonomic indicators for Capsicum species. AB - Capsaicinoids have been suggested as an aid in identifying Capsicum species. The distribution of seven capsaicinoids and their chemotaxonomic significance were examined within nearly 200 accessions of six Capsicum species. The seven capsaicinoids were separated and quantified using high-performance liquid chromatography. The capsaicinoid profiles were not consistent when examined within a species, therefore they have limited use as a chemotaxonomic indicator. In addition, the generalization that capsaicin and dihydrocapsaicin are always the major capsaicinoids was not true, exceptions were found for some of the accessions studied. PMID- 11106845 TI - Hypericin and pseudohypericin in some Hypericum species. AB - Hypericin and pseudohypericin were found in 27 of the 36 evaluated species from Hypericum L., belonging to 17 sections of the genus. Pseudohypericin is reported by us in 15 taxa for the first time. Most of the species contained both components and the amount of pseudohypericin usually exceeded that of hypericin. In H. hirsutum and H. empetrifolium only hypericin was found, whereas H. formosissimum yielded pseudohypericin only. The total content of hypericins varied widely from 0.009% in H. empetrifolium to 0.512% in H. boissieri and the largest amounts were established in taxa of sections Drosocarpium, Hypericum and Thasia. The distribution of hypericin and pseudohypericin in Hypericum species has an important taxonomic value for infrageneric classification of the genus. These components were not found in the primitive sections Ascyreia, Androsaemum, Inodora, Roscyna, Bupleuroides and Spachium but occur widely in Hypericum, Adenosepalum and the sections from Olympia group. Although the genera of subfamily Hypericoideae are characterized by the presence of anthrone derivatives, condensed anthrones such as hypericin and pseudohypericin have not been found in these genera and the remaining subfamilies of the Guttiferae. PMID- 11106846 TI - Intraspecific chemical variability of the leaf essential oil of Juniperus phoenicea subsp. turbinata from Corsica. AB - The composition of 50 samples of essential oil of individual plants of Juniperus phoenicea subsp. turbinata from Corsica was investigated by GC, GC-MS and 13C NMR. alpha-Pinene, beta-phellandrene, alpha-terpinyl acetate, Delta-3-carene, myrcene and alpha-phellandrene were found to be the main constituents. The results were submitted to cluster analysis and discriminant analysis which allowed two groups of essential oils to be distinguished with respect to the content of alpha-pinene, beta-phellandrene and alpha-terpinyl acetate. PMID- 11106847 TI - Volatile oil variability in Thymus serpylloides ssp. gadorensis growing wild in Southeastern Spain. AB - Volatile oils from single plants of Thymus serpylloides ssp. gadorensis were collected from Southeastern Spain and studied to check for chemical variability using gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). Many of the samples showed a phenolic chemotype, while another important group had significant levels of linalool. Geraniol, myrcene, caryophyllene oxide, terpinen-4-ol and 1,8-cineole were commonly present. Principal Component Analysis (PCA) and Cluster Analysis (CA) of this chemical variability separated two groups of plants characterized by either phenols or linalool, and an isolated third type with geraniol. A few samples were found to have both phenolic and non-phenolic compounds in high quantities, thus showing a mixed chemotype. Multidimensional scaling analysis (MDS) of the percentage concentration for each component of the essential oil showed that thymol, linalool, 1,8-cineole, borneol and geraniol have clear divergent vectors. PMID- 11106848 TI - Constituents of the sponge Thorecta reticulata. PMID- 11106849 TI - Chemical constituents of Packera coahuilensis and Packera bellidifolia. PMID- 11106850 TI - Aristolactams from the stem bark of Uvaria hamiltonii. PMID- 11106852 TI - A minor coumarino-lignoid from Jatropha gossypifolia. PMID- 11106851 TI - N-trans-feruloyltyramine from two species of the Solanaceae. PMID- 11106853 TI - Totarol, totaradiol and ferruginol: three diterpenes from Thuja plicata (Cupressaceae). PMID- 11106854 TI - Gynocardin and cyclopentenylglycine in Rawsonia lucida. PMID- 11106855 TI - Ethanol neurobehavioral teratogenesis and the role of the hippocampal glutamate-N methyl-D-aspartate receptor-nitric oxide synthase system. AB - The purpose of this review is to evaluate a proposed mechanism for ethanol neurobehavioral teratogenesis in the hippocampus, involving suppression of the glutamate-N-methyl-D-aspartate (NMDA) receptor-nitric oxide synthase (NOS) system. It is postulated that suppression of this signal transduction system in the fetus by chronic maternal consumption of ethanol plays a key role in hippocampal dysmorphology and dysfunction in postnatal life. This mechanism is evaluated critically based on the current literature and our research findings. In view of the apparent time course for loss of CA1 pyramidal cells in the hippocampus produced by chronic prenatal ethanol exposure that manifests in early postnatal life, it is proposed that therapeutic intervention, which targets the glutamate-NMDA receptor-NOS system, may prevent or lessen the magnitude of postnatal hippocampal dysfunction. PMID- 11106856 TI - Effects of prenatal cocaine exposure on dopamine system development: a meta analysis. AB - Several studies have investigated the effects of prenatal cocaine (PCOC) exposure on the nigrostriatal dopaminergic system in animal models of maternal drug abuse, yet independent examinations of striatal dopamine (DA) receptors and tissue DA levels have produced equivocal results. The current meta-analysis provides a quantitative review of the literature on these topics, and analyzes potential moderators of the effects of PCOC exposure on these variables. The results indicate that the effects of PCOC exposure on striatal DA levels, D1 and D2 receptor-binding densities, and D2 receptor-binding affinity are negligible when collapsed over age, sex, species, and several other methodological variables. However, effects of PCOC exposure on some dopaminergic measures were significantly influenced by factors such as age and sex. As expected, and as suggested by the selectivity and specificity of PCOC-induced changes reported in the published literature, the direction and magnitude of differences between genders or age groups in this study were not systematic across all dependent measures. Generally, PCOC exposure was more often linked to decreases, rather than increases, in the selected dependent measures. These findings indicate that PCOC exposure produces selective alterations in striatal dopaminergic system function which do not appear under all experimental circumstances, but which may be important factors in behavioral alterations seen in selected groups after PCOC exposure. PMID- 11106857 TI - Attention deficit/hyperactivity disorder: characteristics, interventions and models. AB - An epidemiological study of Attention Deficit/Hyperactivity Disorder (ADHD) suggests that the prevalence may be two to three times higher than the figure of 3-5% often cited. In addition, the data suggest that both underdiagnosis and overdiagnosis occur frequently. Rodent animal models of ADHD, like the Spontaneously Hypertensive Rat (SHR) and other rat models such as those with chemical and radiation-induced brain lesions and cerebellar stunting, and the Coloboma mouse model exhibit clear similarities with several aspects of the human disorder and should prove useful in studying specific traits. Operant behavioral tasks that model learning, short-term memory and simple discriminations are sensitive to ADHD and methylphenidate has been shown to normalize ADHD performance in a short-term memory task. Recent findings challenge not only the current postulate that response inhibition is a unique deficit in ADHD, but also the concepts of ADHD and its treatment, which presume intact perceptual abilities. Time perception deficits may account, in part, for the excessive variability in motor response times on speeded reaction time tasks, motor control problems and motor clumsiness associated with ADHD. The Multimodality Treatment Study of ADHD (MTA) provided data suggesting that pharmacological interventions that included systematic and frequent follow-up with parents and teachers, with or without psychosocial interventions, are superior to psychosocial interventions or standard community care alone. Additionally, the MTA was one of the first studies to demonstrate benefits of multimodal and pharmacological interventions lasting longer than 1 year. Imaging studies have demonstrated differences in brain areas in children with ADHD: anterior corpus callosum, right anterior white matter, and cerebellar volumes are all decreased in children with ADHD and there is less brain asymmetry in ADHD subjects. Additionally, functional imaging studies, coupled with pharmacological manipulations, suggest decreased blood flow and energy utilization in prefrontal cortex and striatum and the dysregulation of catecholamine systems in persons with ADHD. PMID- 11106858 TI - Neurobehavioral outcomes of cocaine-exposed infants. AB - The present study investigated the neurobehavioral outcomes of fetal cocaine exposure. Attempts were made to control, by design or statistical analysis, for significant confounders. Timing and amount of drug exposures were considered, and biologic measures of exposure were quantified to classify exposure severity. One hundred sixty-one non-cocaine and 158 cocaine-exposed (82 heavily and 76 lightly exposed) infants were seen at a mean-corrected age of 43 weeks post-conception and administered the Neurobehavioral Assessment (NB Assessment). Heavily cocaine exposed infants had more jitteriness and attentional problems than lightly and non-exposed infants. They also had more movement and tone abnormalities, and sensory asymmetries than non-exposed infants. Heavily exposed infants were more likely to be identified with an abnormality than non-exposed infants and there was a trend toward heavily exposed infants being more likely to be identified with an abnormality than lightly exposed infants. Furthermore, there was a trend for heavily exposed infants to be less likely to be testable than non-exposed infants. After the confounding and mediating factors were considered, heavily cocaine-exposed infants were four times as likely to be jittery and nearly twice as likely to demonstrate any abnormality than lightly and non-exposed infants, but all other effects were no longer significant. Higher concentrations of the cocaine metabolites of cocaine, cocaethylene, and benzoylecgonine (BZE) were related to higher incidence of movement and tone abnormalities, jitteriness, and presence of any abnormality. Higher cocaethylene levels were related to attentional abnormalities and higher meta-hydroxybenzoylecgonine (m-OH-BZE) was related to jitteriness. Drug effects on attention were mediated by maternal psychological distress, suggesting that this factor should be considered in future studies of drug exposure effects. PMID- 11106859 TI - Quantitative trait loci for acute behavioral sensitivity to paraoxon. AB - Genetic mechanisms responsible for organophosphate (OP)-induced behavioral changes remain obscure. In the present study, provisional quantitative trait loci (QTL) associated with acute sensitivity or insensitivity to hypolocomotion produced by the OP paraoxon were identified. Naive adult male and female mice of the BXD/Ty series (22 different BXD strains plus C57BL/6J and DBA/2J progenitor strains) received 0 or 0.25 mg/kg paraoxon (IP), immediately before placement in an activity chamber for a 30-min test. As expected, based on dose-response and time course studies with Swiss-Webster, C57BL/6, and DBA/2 mice, paraoxon treatment reduced locomotor activity in most, but not all BXD strains. Heritability (proportion of phenotypic variability attributed to genetic differences) was 0. 58 for the paraoxon treatment effect. Difference scores (strain mean for vehicle activity minus strain mean for paraoxon activity), and percent change in activity of paraoxon-treated mice compared to vehicle-treated mice were calculated for each BXD strain. QTL analyses using activity difference scores and percentage change in activity were conducted using a database with over 1300 unique genetic markers. Several provisional QTL found on different chromosomes were associated with the activity phenotype. Of these, several markers attained p<0.01 or greater. These were as follows: Chr 1: Ly9, p<0.006; Chr 6: D6Ncvs44, p<0.0005; Chr 9: D9Mit15, p<0. 003; Chr 11: D11Ncvs76, p<0.002; Chr 15: Tstap198, p<0.008. In addition, several markers on chromosome 3 approached p<0.01. Identified genes found near these regions include two plasma carboxylesterase alleles on chromosomes 6 and 9, a glutamate receptor subtype on chromosome 11 and a glycine receptor subunit on chromosome 11, raising the possibility that these genes could be the basis for these provisional QTLs. PMID- 11106860 TI - Gestational and lactational exposure to TCDD or coplanar PCBs alters adult expression of saccharin preference behavior in female rats. AB - Previous studies have shown that maternal doses of 1 microg/kg or less of 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) in late gestation can demasculinize and feminize reproductive behavior in male rats. However, it was not known whether coplanar polychlorinated biphenyls (PCBs) had similar effects, or whether non reproductive sexually dimorphic behaviors such as saccharin preference behavior were also altered. We determined the effects of TCDD or coplanar PCBs on saccharin consumption and saccharin preference in male and female rats. Sprague Dawley rats were dosed with 3,3',4, 4'-tetrachlorobiphenyl (PCB 77; 2 or 8 mg/kg/day), 3,3',4,4', 5-pentachlorobiphenyl (PCB 126; 0.25 or 1.0 microg/kg/day), TCDD (0. 025 or 0.10 microg/kg/day), or corn oil vehicle on days 10-16 of gestation. Maternal exposure to TCDD or coplanar PCBs did not change saccharin consumption or saccharin preference in male rats. However, TCDD and coplanar PCB-exposed females showed decreased saccharin consumption and saccharin preference. The results indicate that saccharin consumption is masculinized in female rats exposed to TCDD or coplanar PCBs during perinatal development. This effect could be related to the anti-estrogenic actions of these chemicals. PMID- 11106861 TI - Developmental aspects of delayed matching-to-sample task performance in children. AB - The influence of age, sex, and intelligence (IQ) on performance of a delayed matching-to-sample (DMTS) task, commonly used with animals and adult human subjects to study aspects of short-term memory, was examined for 674 children, 5 to 13 years old. The data suggest that younger children were less accurate at short delays and displayed a greater decrease in accuracy as recall delay increased than older children. Children with lower IQs demonstrated consistent impairment in recall of information when compared to children with higher IQs. No significant differences in task performance were observed between boys and girls. These normative data provide insights into the developmental time course of behaviors thought to serve as metrics of short-term memory. These data will be critical for ongoing and future studies in determining whether specific clinical diagnoses, drug treatments, or other risk factors (e.g., perinatal drug exposure, pregnancy complications, exposure to toxicants) are associated with differences on specific aspects of task performance. The use of tasks that are also applicable to animal models provides great opportunities for the conduct of important comparative studies. PMID- 11106862 TI - Nicotine exposure during the neonatal brain growth spurt produces hyperactivity in preweanling rats. AB - Despite warning labels and increases in evidence of the adverse effects of tobacco use, women continue to use tobacco products during pregnancy. Cigarette smoking has been linked to increased prenatal mortality, increased incidence of SIDS, reductions in birth weight, and disruptions in CNS and behavioral development. Animal model systems have critically established the causal relationship between nicotine and adverse developmental outcome. The present study examines the behavioral effects of nicotine exposure in the rat during the third trimester equivalent of the human brain growth spurt, a period of rapid development of the cholinergic systems and a period during which the CNS is particularly vulnerable to a number of insults. Sprague-Dawley rat pups were exposed to nicotine (6.0 mg/kg/day) from postnatal days (PD) 4-9 via an artificial rearing procedure. This procedure ensures that observed effects are not due to nutritional deficits. Two control groups were employed, an artificially reared control group and a normally reared control group. Activity level was measured on PD 18-19. Nicotine-exposed subjects were significantly overactive compared to both control groups, which did not differ significantly from one another. This behavioral alteration was observed in the absence of nicotine-induced body weight deficits. These results suggest that women who use tobacco products during late gestation may place their fetuses at risk for hyperactivity later in life, particularly during early adolescence. PMID- 11106863 TI - Neonatal choline supplementation ameliorates the effects of prenatal alcohol exposure on a discrimination learning task in rats. AB - Prenatal alcohol exposure can disrupt brain development and lead to a myriad of behavioral alterations, including motor coordination deficits, hyperactivity, and learning deficits. There remains a need, however, to identify treatments and interventions for reducing the severity of alcohol-related neurodevelopmental disorders. Some of the alcohol-induced deficits in learning may be related to alterations in cholinergic functioning. Interestingly, there is a growing literature demonstrating that pre- and/or early postnatal choline supplementation can lead to long-term enhancement in learning and memory and cholinergic activity in rats. The present study examined whether such early choline supplementation might counter the effects of prenatal alcohol treatment on a visuospatial discrimination task. Pregnant Sprague-Dawley rats were randomly assigned to one of three prenatal treatment groups. One group received a liquid diet containing 35% ethanol-derived calories (EDC) from gestational day (GD) 6-20. A second group served as a pair-fed (PF) control group and the third group served as an ad lib lab chow (LC) control. On postnatal day (PD) 2, pups were assigned within-litter to one of three postnatal treatments: choline, saline vehicle, or no treatment. Choline and vehicle pups were intubated with a choline chloride solution or vehicle daily from PD 2 to 21, whereas the non-treated pups were handled daily but not intubated. On PD 45, subjects were tested on a visuospatial discrimination task. Ethanol-exposed subjects who were not treated neonatally with choline committed a significantly greater number of errors both during acquisition and during delayed discrimination training compared to both PF and LC controls. Neonatal choline treatment significantly improved performance on the discrimination task in all groups; however, the beneficial effects of choline were significantly larger in ethanol-exposed subjects. Indeed, the performance of ethanol-exposed pups treated with neonatal choline did not differ from any of the PF or LC groups on any measure. Thus, early postnatal choline supplementation significantly attenuated the effects of prenatal alcohol on this learning task. Importantly, these effects were not due to the acute effects of choline, but rather to long-term changes in brain and behavioral development. These data suggest that early dietary interventions may reduce the severity of fetal alcohol effects. PMID- 11106864 TI - Dose-response and time-course of neurobehavioral changes following oral chlorpyrifos in rats of different ages. AB - Young rats have been shown in several laboratories to be more sensitive to the neurotoxic effects of acute exposure to chlorpyrifos. To examine the neurobehavioral effects of chlorpyrifos as a function of age and dose, we conducted dose-response and time-course assessments in rats of three different ages (postnatal day, or PND, 17, 27, and adults). Doses were selected to span the effective dose range in each age group: PND17 - 4, 10, 20 mg/kg; PND27 - 10, 25, 50 mg/kg; adult - 10, 50, 100 mg/kg. Rats were tested at the time of peak effect on the day of dosing, and again at 1 and 3 days, and at 1 and 2 weeks after a single oral dose. There were age- and sex-related differences in the recovery of these behavioral effects; the adult males recovered from the behavioral effects more quickly than the other age groups, and the adult females showed the slowest recovery (up to at least 3 days). Although these doses had been shown previously to produce a similar degree of cholinesterase inhibition, the neurobehavioral alterations fell into the following three patterns of effect as a function of age. (1) Some endpoints (e.g., gait abnormalities, tremor) showed a dose-response curve that was shifted to the right in the older animals. Calculated ED50 values indicated that the PND17 rats were three- to five-fold more sensitive than the adults. (2) Some measures showed less effect in the youngest rats; for example, maximal motor activity decreases were half as great as with adults. (3) A few effects that were typically observed in adults, e.g., salivation, were not seen at all in the PND17 rats. Thus, differential responses on these neurobehavioral endpoints were observed as a function of age. These data suggest that, for some endpoints, young rats are more sensitive to a range of chlorpyrifos doses; however, the magnitude of age-related differences depends on the specific endpoint and time of assessment, as well as age and sex of the test subject. PMID- 11106865 TI - Inhalation exposure to white spirit causes region-dependent alterations in the levels of glial fibrillary acidic protein. AB - Enhanced expression of glial fibrillary acidic protein (GFAP) is known to be associated with toxicant-induced gliosis, a homotypic response of the central nervous system to neural injury. A variety of neurochemical and neurophysiological effects have been observed in experimental animals exposed to white spirit, but a linkage of such effects to neural damage has not been established. Here we evaluated the regional levels of GFAP to assess potential sites of CNS damage in the rat, following exposure to dearomatized and aromatic white spirit. Samples from rats exposed to dearomatized white spirit were assayed for GFAP levels in the United States and Denmark. The results were remarkably similar between countries. Small region-dependent increases and decreases in GFAP were observed with the cerebellum showing the most consistent effects (increases). In contrast, samples from rats exposed to aromatic white spirit showed large (as much as 150% of control) increases in regional levels of GFAP; again, the cerebellum showed the most consistent effects. The data are indicative of an aromatic white-spirit-induced astrogliosis in several regions of the rat CNS and suggest that chronic exposure to this solvent may be associated with underlying neural damage. PMID- 11106866 TI - Chronic prenatal exposure to paroxetine (Paxil) and cognitive development of mice offspring. AB - This study investigated the impact on cognitive development in CD-1 mice from chronic prenatal exposure to the antidepressant paroxetine. CD-1 mice were given either paroxetine as 30 mg/kg/day or a placebo in food bars for 2 weeks before mating and throughout gestation. One offspring per gender from each litter was tested on each of the following tasks: tube runway, spatial maze, passive avoidance chamber, and water straight runway followed by an unforced decision maze. Learning occurred in both genders in all tasks (p<0.001) with no significant differences between treatment groups at the final learning session. Juvenile runway was the only task in which the paroxetine-exposed males demonstrated a learning rate that was slower than the placebo-exposed offspring (p=0.06). Post learning sessions did not show any significant treatment differences during the juvenile and adult periods during the water straight runway, mazes, and avoidance chamber tasks. In conclusion, chronic prenatal exposure in mice of paroxetine did not impact cognition on select tasks. PMID- 11106867 TI - Two components of long-term depression are impaired by chronic lead exposure in area CA1 and dentate gyrus of rat hippocampus in vitro. AB - Previous studies have demonstrated that low-level lead exposure can impair the induction of long-term depression (LTD) in area CA1 and dentate gyrus (DG) of rat hippocampus in vitro and in vivo. The induction of LTD in area CA1 and DG has been shown to associate with N-methyl-D-aspartate receptors (NMDARs) and voltage gated calcium channel (VGCC). In this study, the relative contributions of NMDARs dependent and VGCC-dependent components in the induction of LTD in the hippocampus and the impairments of these two components of LTD by chronic low level lead exposure were investigated. Neonatal Wistar rats were exposed to lead from parturition to weaning via milk of dams drinking 0.2% lead acetate solution. Field excitatory postsynaptic potentials (EPSPs) were recorded in area CA1 and DG before and after two 15-min trains of 1-Hz low-frequency stimulation (LFS) (2x900 pulses). In area CA1, the amplitude of NMDARs-dependent LTD (NMDA-LTD), in the presence of 10 microM nimodipine (a blocker of L-type Ca(2+) channels), was 80.05+/-2.54% (n=8) and 94.58+/-10.57% (n=8) in the control and lead-exposed rats, respectively. The amplitude of VGCC-dependent LTD (VGCC-LTD), in the presence of 50 microM (-)-2-amino-5-phosphonopentanoic acid (AP5), was 80.36+/ 4.08% (n=10) and 93.91+/-7.85% (n=10) in the control and lead-exposed rats, respectively. In area DG the amplitude of NMDA-LTD, with both 50 microM Ni(2+) (a blocker of T-type Ca(2+) channels) and 10 microM nimodipine present, in the control rats (79. 97+/-4.30%, n=8) was significantly larger than that in the lead exposed rats (91.24+/-11.08%, n=10, P<0.001). The amplitude of VGCC-LTD, with 50 microM AP5 present, was significantly larger in the control rats (70.80+/-3.64%, n=9) than that in the lead-exposed rats (87.60+/-9.00%, n=10, P<0.001). The results suggested that chronic lead exposure affected two components of LTD induction in area CA1 and DG. Furthermore, the impairment of two components by lead exposure might be similar in area CA1, while the impairment of VGCC-LTD might be more serious in DG of hippocampus. PMID- 11106869 TI - Valproate enhances N-cadherin production in Xenopus embryos. AB - Xenopus embryos were exposed to valproate (0, 20, 40, 80 mg/l) either before or after neural tube closure. The embryos were then homogenized and fractionated by gel electrophoresis, and N-cadherin was detected and measured with quantitative immunoblotting. Findings indicated that valproate exposure increased N-cadherin production in a dose-dependent manner. Embryos exposed prior to neural tube closure tended to be more sensitive to the effects of valproate. These findings suggest that alterations in N-cadherin-mediated adhesion or morphogenesis may partially explain the teratogenic mechanism of valproate. PMID- 11106868 TI - Preweaning treatment with methamphetamine induces increases in both corticosterone and ACTH in rats. AB - Treatment with methamphetamine (MA) on postnatal days P11-20 induces adult spatial learning and memory deficits without affecting monoamine levels in various brain regions. In this study, we examined the pituitary and adrenal response of animals administered MA four times daily on P11, P11-15, or from P11 to P20. Corticosterone (CORT) and adrenocorticotropin hormone (ACTH) levels were assessed over a 1-hour period following MA exposure. On P11, MA produced marked elevations of both CORT and ACTH; this is during the stress hyporesponsive period (SHRP). On P15 and P20, the maximal effect of MA on CORT titers was observed at 30 min, with lower, but still significantly increased, levels at 60 min compared to controls. Males receiving MA on P15 had higher levels of ACTH than did control males, while no differences were noted among females. On P20, MA treatment resulted in higher levels of ACTH relative to vehicle-injected controls, but levels were not different from controls that were only weighed at each drug administration. MA treatment inhibited body, but not brain weight gain, resulting in hippocampal weights that were heavier in the MA-treated animals when expressed as a percent of body weight. The elevations of adrenal steroids by MA, during late phases of hippocampal neurogenesis, may contribute to neuronal alterations that are later manifested in deficits of learning and memory. PMID- 11106870 TI - CRH signalling in the bed nucleus of the stria terminalis is involved in stress induced cardiac vagal activation in conscious rats. AB - The bed nucleus of the stria terminalis (BNST) is involved in autonomic and behavioral reactions to fearful stimuli and contains corticotropin-releasing hormone (CRH) fibers and terminals. The role of CRH in the medial part of the BNST in the regulation of heart rate (HR) and PQ interval of the electrocardiogram was studied under resting conditions and conditioned fear stress in freely moving rats. Microinfusion of CRH (0.2 microg/0.6 microl) in the medial BNST under resting conditions significantly enhanced HR as compared to saline treatment, but did not reduce the PQ interval, indicating that exogenous CRH in the medial BNST can activate both the sympathetic and parasympathetic cardiac outflow. In addition, CRH induced a slight increase in gross locomotor activity, an effect that succeeded the tachycardiac response, indicating that the HR response was not a consequence of increased locomotor activity, but likely a direct effect of CRH. CF was induced by 10-min forced exposure to a cage in which the rat had experienced footshocks (5 x 0.5 mA x 3s) the day before. alpha helical CRH(9-41) (alphahCRH; 5 microg/0.6 microl), a non-selective CRH receptor antagonist, or saline was infused into the medial BNST of rats prior to CF. CF induced freezing behavior, associated with an increase in HR and PQ interval, indicating activation of sympathetic and vagal outflow to the heart. alphahCRH significantly reduced the PQ response, but enhanced the tachycardia, suggesting inhibition of vagal activity. In addition, alpha-helical CRH(9-41) reduced the freezing response. Taken together, the data provide first evidence that CRH, released in the medial BNST during stress, contributes to cardiac stress responses, particularly by activating vagal outflow. PMID- 11106871 TI - Desensitization of 5-HT(1A) autoreceptors by a low chronic fluoxetine dose effect of the concurrent administration of WAY-100635. AB - Using microdialysis, receptor autoradiography and in situ hybridization, we examined the effects of fluoxetine alone or with WAY-100635 on: (a) extracellular 5-HT in frontal cortex; and (b) density and sensitivity of 5-HT(1A) autoreceptors in rat brain. WAY-100635 (0.3 mg/kg, s.c.) doubled the increase in extracellular 5-HT produced by fluoxetine (3 mg/kg, i.p.) in frontal cortex. Two-week minipump treatments with these daily doses significantly raised extracellular 5-HT to 275 +/- 33% (fluoxetine) and 245 +/- 10% (fluoxetine plus WAY-100635) of controls. Fluoxetine 3 mg/kg.day desensitized dorsal raphe 5-HT(1A) autoreceptors, an effect prevented by the concurrent WAY-100635 administration. However, WAY-100635 (alone or with fluoxetine) did not change 5-HT(1A) autoreceptor sensitivity. The density of 5-HT(1A) receptors and its encoding mRNA, was unaffected by these treatments. These results suggest that prolonged blockade of 5-HT(1A) receptors in vivo prevents the autoreceptor desensitization induced by fluoxetine but does not result in receptor sensitization. PMID- 11106872 TI - Modifications in brain CaM kinase II after long-term treatment with desmethylimipramine. AB - The present study investigated the effect of long-term (15 mg/kg for 15 days) and acute (15 mg/kg, single administration) treatment with desmethylimipramine, a tricyclic antidepressant drug, on calcium/calmodulin-dependent protein kinase II (CaMKII), a kinase implicated in the mechanism of antidepressant drug action. Similar to selective and non-selective serotonin reuptake inhibitors, long-term, but not acute, treatment with desmethylimipramine markedly increased the activity of CaMKII in the hippocampal synaptic vesicle fraction (+51.9%). The kinase activity was also increased in the same fraction of frontal cortex (+24.2%) and in the striatum (+45.9%), although in this last area the mechanism appeared to be different because the protein level of the kinase was also markedly increased (+43.7%). However, the effect of treatment was not restricted to the presynaptic kinase, because CaMKII activity was also increased in the total cellular cytosol in cortical areas. The autonomous (calcium-independent) activity of CaMKII was assayed for the first time after antidepressant treatment, and found to be increased in synaptic vesicles of all three areas. These results confirmed the involvement of CaMKII in antidepressant drug action and suggested that modulation of transmitter release is a primary component in the action of psychotropic drugs. PMID- 11106873 TI - Low versus standard dose mCPP challenge in obsessive-compulsive patients. AB - In several reports, the acute oral administration of the partial serotonergic agonist meta-chlorophenylpiperazine (mCPP) in dose of 0. 5 mg/kg induced a significant worsening of obsessive-compulsive (OC) symptoms in a number of patients. The aim of our study was to test the 0.25 mg/kg mCPP dose, which was hypothesized to be more specific for OC symptoms and was until now tested only on healthy subjects. In a double-blind, controlled crossover study, 12 OC patients participated on three test days, receiving one of the following on each day: oral 0.5 mg/kg mCPP (standard dose), 0.25 mg/kg mCPP (low dose), or placebo. Behavioral ratings were obtained by means of Visual Analogue Scale (VAS) ratings. The low dose mCPP induced a significant worsening of OC symptoms in 50% (6/12) of the patients, whereas 8.3% (1/12) of the patients showed a worsening after the standard dose. On the other hand, only the standard dose mCPP induced a worsening, although not statistically significant, of anxiety ratings. Our data show that the 0.25 mg/kg dose mCPP induces a specific response in OC symptoms, with little anxiogenic effect. To confirm these preliminary data, future studies will be needed on larger samples and with more sensitive rating scales. PMID- 11106874 TI - Lower baseline plasma cortisol and prolactin together with increased body temperature and higher mCPP-induced cortisol responses in men with pedophilia. AB - There is some evidence that hormonal and serotonergic alterations may play a role in the pathophysiology of paraphilias. The aims of the present study were to examine: 1) baseline plasma cortisol, plasma prolactin, and body temperature; and 2) cortisol, prolactin, body temperature, as well as behavioral responses to meta chlorophenylpiperazine (mCPP) and placebo in pedophiles and normal men. Pedophiles showed significantly lower baseline plasma cortisol and prolactin concentrations and a higher body temperature than normal volunteers. The mCPP induced cortisol responses were significantly greater in pedophiles than in normal volunteers. In normal volunteers, mCPP-induced a hyperthermic response, whereas in pedophiles no such response was observed. mCPP induced different behavioral responses in pedophiles than in normal men. In pedophiles, but not in normal men, mCPP increased the sensations "feeling dizzy, " "restless," and "strange" and decreased the sensation "feeling hungry". The results suggest that there are several serotonergic disturbances in pedophiles. It is hypothesized that the results are compatible with a decreased activity of the serotonergic presynaptic neuron and a 5-HT2 postsynaptic receptor hyperresponsivity. PMID- 11106875 TI - The effect of clozapine on caudate nucleus volume in schizophrenic patients previously treated with typical antipsychotics. AB - Typical antipsychotics have been reported to enlarge the caudate nucleus in schizophrenic patients. The atypical antipsychotic, clozapine, is associated with a decrease in caudate size in patients previously treated with typical antipsychotics. The present study investigates whether a change in caudate volume after switching from treatment with typical antipsychotics to treatment with clozapine is related to improvement in symptoms or tardive dyskinesia (TD). Twenty-six schizophrenic patients participated in this open study. Caudate nucleus volume and TD were assessed before discontinuing typical antipsychotics and after 24 weeks of treatment with clozapine. After discontinuing typical antipsychotics, symptoms were assessed in a 3 days drug-free period and subsequently once a month. Treatment with clozapine resulted in a decrease in caudate volume, improvement in symptoms and amelioration of TD. However, no difference in caudate volume changes was found between responders and non responders to clozapine and no correlations were found between caudate volume changes and reduction of TD. In conclusion, this study replicates earlier findings that clozapine decreases caudate volume in patients previously treated with typical antipsychotics and suggests that this effect is unrelated to treatment response or to amelioration of TD. PMID- 11106876 TI - Fatty acid derivatives of clozapine: prolonged antidopaminergic activity of docosahexaenoylclozapine in the rat. AB - Stable amides of clozapine derived from fatty acids prominent in cerebral tissue might enhance the central activity of clozapine and reduce its exposure to peripheral tissues. Such derivatives might enhance the safety of this unique drug, which is the only agent with securely established superior antipsychotic effectiveness, but with a risk of potentially lethal systemic toxicity. Amide derivatives of clozapine were prepared from structurally varied fatty acid chlorides and evaluated for ability to inhibit behavioral arousal in rat induced by dopamine agonist apomorphine and to induce catalepsy. Their duration-of-action and potency were compared to free clozapine, and concentrations of clozapine were assayed in brain and blood. Selected agents were also evaluated for affinity at dopamine receptors and other potential drug-target sites. Clozapine-N-amides of linoleic, myristic, oleic, and palmitic acids had moderate initial central depressant activity but by 6 h, failed to inhibit arousal induced by apomorphine. However, the docosahexaenoic acid (DHA) derivative was orally bioavailable, 10 times more potent (ED(50) 5.0 micromol/kg) than clozapine itself, and very long acting (>/= 24 h) against apomorphine, and did not induce catalepsy. DHA itself was inactive behaviorally. Clozapine showed expected dopamine receptor affinities, but DHA-clozapine was inactive at these and other potential target sites. After systemic administration of DHA-clozapine, serum levels of free clozapine were very low, and brain concentrations somewhat lower than after administering clozapine. DHA-clozapine is a long-acting central depressant with powerful and prolonged antidopaminergic activity after oral administration or injection without inducing catalepsy, and it markedly reduced peripheral exposure to free clozapine. It lacked the receptor-affinities shown by clozapine, suggesting that DHA-clozapine may be a precursor of free, pharmacologically active clozapine. Such agents may represent potential antipsychotic drugs with improved central/peripheral distribution, and possibly enhanced safety. PMID- 11106877 TI - Impaired inhibition of conditioned responses produced by subchronic administration of phencyclidine to rats. AB - Several recent investigations have suggested that an important function of the frontostriatal system is inhibitory response control, and we previously reported that subchronic exposure to phencyclidine (PCP) produced deficits in inhibitory control in monkeys. The current studies were designed to examine whether subchronic administration of PCP to rats would subsequently affect the ability to inhibit conditioned responses when relationships between reward and stimuli of affective significance change. First, the effects of long-term exposure to PCP on acquisition of a novel, concurrent discrimination or reversal learning were assessed; PCP-treated rats were selectively impaired in the ability to acquire the reversal of an already-learned stimulus-reward association. Furthermore, there were no effects of PCP treatment on the learning of a novel instrumental response; however, PCP-treated rats produced more responses during extinction of instrumental responding than did control subjects. Finally, PCP-treated rats produced more responses for a conditioned reinforcer than did control rats. These data suggest that PCP-treated rats are impaired in their ability to modulate behavior based upon new or changing information about stimulus-reward associations, possibly due to an inability to inhibit conditioned responding towards incentive stimuli. These effects may have relevance to mental disorders involving affective impairments and impulsivity, including schizophrenia, obsessive-compulsive disorders, and drug abuse. PMID- 11106878 TI - Augmented accumbal serotonin levels decrease the preference for a morphine associated environment during withdrawal. AB - Recent studies have found that acute morphine administration increases serotonin (5-HT) transmission within the nucleus accumbens and other forebrain regions. In contrast, 5-HT transmission is depressed during withdrawal from chronic morphine. We show that pharmacological agents that increase brain 5-HT levels (fluoxetine or 5-hydoxytryptophan, 5-HTP) abolish the preference of chronically morphine treated, withdrawn rats for a morphine-associated environment. Similar results were seen when fluoxetine was microinjected into the nucleus accumbens. Conversely, rats given morphine acutely showed an enhanced preference for a morphine-associated environment when pretreated with these agents. Fluoxetine also decreased the heightened anxiety found in morphine withdrawn rats. The results of our study indicate that drugs that augment 5-HT levels may reduce the desire for morphine during withdrawal. PMID- 11106880 TI - The use of clinical information in diagnosing chronic heart failure: a comparison between general practitioners, cardiologists, and students. AB - In a clinical judgement analysis study, 27 general practitioners, 22 cardiologists, and 21 medical students assessed 40 case vignettes with regard to the probability of heart failure, in order to study the weights of different kinds of information (cues) measured by the regression coefficients in a multiple regression model. The vignettes were based on actual patients. We found that diagnostic accomplishment and diagnostic strategies were surprisingly similar on the group level, but very different on the individual level. The most important cues for the participants were cardiac enlargement and pulmonary stasis. Strategies in which cardiac enlargement was the predominating cue led to a higher diagnostic accomplishment; a third of the participants used such strategies. The cues given in the vignettes could have been utilized more efficiently; cardiac enlargement seems to be more important and "classical" symptoms less important for predicting heart failure than the participants realize. PMID- 11106879 TI - Interleukin-6 enhances expression of adenosine A(1) receptor mRNA and signaling in cultured rat cortical astrocytes and brain slices. AB - The inhibitory neuromodulator adenosine is released in the brain in high concentrations under conditions of exaggerated neuronal activity such as ischemia and seizures, or electroconvulsive treatment. By inhibiting neural overactivity, adenosine counteracts seizure activity and promotes neuronal survival. Since stimulation of adenosine A(2b) receptors on astrocytes induces increased synthesis and release of interleukin-6, which also exerts neuroprotective effects, we hypothesized that the effects of interleukin-6 and of adenosine might be related. We report here that stimulation with interleukin-6 of cultured astrocytes, of cultured organotypic brain slices from newborn rat cortex, and of freshly prepared brain slices from rat cortex induces a concentration- and time dependent upregulation of adenosine A(1) receptor mRNA. This increased adenosine A(1) receptor mRNA expression is accompanied in astrocytes by an increase in adenosine A(1) receptor-mediated signaling via the phosphoinositide-dependent pathway. Since upregulation of adenosine A(1) receptors leads to increased neuroprotective effects of adenosine, we suggest that the neuroprotective actions of interleukin-6 and adenosine are related and might be mediated at least in part through upregulation of adenosine A(1) receptors. These results may be of relevance for a better understanding of neuroprotection in brain damage but also point to a potential impact of neuroprotection in the mechanisms of the antidepressive effects of chronic carbamazepine, electroconvulsive therapy, and sleep deprivation, which are all accompanied by adenosine A(1) receptor upregulation. PMID- 11106881 TI - The association between exposure to a rear-end collision and future neck or shoulder pain: a cohort study. AB - Neck pain is the most frequently reported feature in connection with whiplash injury, but it is also a common complaint in the general population. Therefore it is crucial to include an unexposed comparison group when evaluating the association between neck pain and a previous motor vehicle crash (MVC). To determine whether exposure to a rear-end collision, without or with whiplash injury, is associated with future neck or shoulder pain, a cohort study was conducted. The study population consisted of persons covered by traffic insurance at one of the largest insurance companies in Sweden. Claim reports were collected from the period November 1987 to April 1988. Drivers exposed to a rear-end collision were divided into two subgroups, without reported whiplash injury (n = 204) and with reported whiplash injury (n = 232). Two comparison groups, unexposed to MVCs, consisting of 1599 and 2089 persons, were selected with consideration taken to the age and gender distribution in the exposed subgroups. A questionnaire concerning neck or shoulder pain and other subjective health complaints was mailed to all the study subjects at follow-up in 1994, 7 years after the rear-end collision. The relative risk of neck or shoulder pain at follow-up was 1.3 (95% CI 0.8-2.0) in the exposed subjects without whiplash injury compared with the unexposed. The corresponding relative risk in subjects with whiplash injury was 2.7 (95% CI 2.1-3. 5). We conclude that there is no increased risk of future neck or shoulder pain in drivers who did not report whiplash injury in connection with a rear-end collision 7 years earlier. In drivers with reported whiplash injury, the risk of neck or shoulder pain 7 years after the collision was increased nearly three-fold compared with that in unexposed subjects. PMID- 11106882 TI - Construction of an algorithm for quick detection of patients with low bone mineral density and its applicability in daily general practice. AB - OBJECTIVE: To construct a quick algorithm to detect patients with low bone mineral density (BMD) and osteoporosis and determine its applicability in daily general practice. DESIGN: Cross-sectional study in all 9107 postmenopausal women, aged 50-80, registered at 12 general practice centers. SUBJECTS AND MEASUREMENTS: All healthy women (5303) and 25% of the remaining group (943/3804) were invited to participate. Of 6246 invited women, 4725 (76%) participated. The women were questioned (state of health, medical history, family history, and food questionnaire) and examined [weight, height, body mass index (BMI), and BMD of the lumbar spine]. STATISTICS: Multivariable, stepwise backward and forward logistic regression analyses were performed, with BMD of the lumbar spine (L2-L4, cut-off points at 0.800 g/cm(2) for osteoporosis and 0.970 g/cm(2) for low BMD) as the dependent variable. An algorithm was constructed with those variables that correlated statistically significantly and clinically relevant with the presence of both osteoporosis and low BMD. RESULTS: The prevalence of osteoporosis was 23%, that of low BMD was 65%. Only three variables (age, BMI, and fractures) were statistically significant and clinically relevant correlated with the presence of both osteoporosis and low BMD. Age (OR 2.70 for osteoporosis and OR 1.77 for low BMD) and fractures during the past five years (OR 3.60 for osteoporosis and OR 2.85 for low BMD) were found to be the key predictors. From the algorithm the absolute risks varied from 9% to 51% for osteoporosis and from 48% to 84% for low BMD. The corresponding relative risks varied from 1.0 to 5.7 and from 1.0 to 1.8. CONCLUSIONS: Using an algorithm with age, BMI, and fracture history subgroups at high risk could be identified. However, in whatever combination, many women with osteoporosis could not be identified. Despite the differences in methods, we found predictors for osteoporosis which were comparable with the results of other cross-sectional studies, meaning that the first selection of patients at high risk for low BMD can be done adequately by both specialists and general practitioners. PMID- 11106883 TI - The Tromso study: determinants of precision in bone densitometry. AB - Studies of precision determinants in bone densitometry are scarce. A total of 111 subjects recruited from the population-based multipurpose Tromso Study (Norway), 27-75 years of age, had repeated forearm bone single X-ray absorptiometry (SXA) measurements. Measurement conditions were systematically varied in series up to eight scans. Median coefficients of variation (CV) for two scans performed 1 week apart, by two different operators were 0.79% and 0.98% at distal and ultradistal sites, respectively. The CV distribution was skewed: 5% of the subjects had individual CVs above 2.2% (distal) and 3.4% (ultradistal). Age (P = 0.0097) and repositioning were important determinants of precision. The SXA bone mineral density (BMD)-measurement method is sufficiently precise to establish BMD level. The minimal individual percentage BMD change that can be detected with 95% certainty was 2% and 3% at distal and ultradistal sites, respectively. Detection of BMD changes less than this should rely on multiple repeat measurements at each point in time. PMID- 11106884 TI - Predicting non-elective hospital readmissions: a multi-site study. Department of Veterans Affairs Cooperative Study Group on Primary Care and Readmissions. AB - OBJECTIVE: To determine clinical and patient-centered factors predicting non elective hospital readmissions. DESIGN: Secondary analysis from a randomized clinical trial. CLINICAL SETTING: Nine VA medical centers. PARTICIPANTS: Patients discharged from the medical service with diabetes mellitus, congestive heart failure, and/or chronic obstructive pulmonary disease (COPD). MAIN OUTCOME MEASUREMENT: Non-elective readmission within 90 days. RESULTS: Of 1378 patients discharged, 23.3% were readmitted. After controlling for hospital and intervention status, risk of readmission was increased if the patient had more hospitalizations and emergency room visits in the prior 6 months, higher blood urea nitrogen, lower mental health function, a diagnosis of COPD, and increased satisfaction with access to emergency care assessed on the index hospitalization. CONCLUSIONS: Both clinical and patient-centered factors identifiable at discharge are related to non-elective readmission. These factors identify high-risk patients and provide guidance for future interventions. The relationship of patient satisfaction measures to readmission deserves further study. PMID- 11106885 TI - Publication and related bias in meta-analysis: power of statistical tests and prevalence in the literature. AB - Publication and selection biases in meta-analysis are more likely to affect small studies, which also tend to be of lower methodological quality. This may lead to "small-study effects," where the smaller studies in a meta-analysis show larger treatment effects. Small-study effects may also arise because of between-trial heterogeneity. Statistical tests for small-study effects have been proposed, but their validity has been questioned. A set of typical meta-analyses containing 5, 10, 20, and 30 trials was defined based on the characteristics of 78 published meta-analyses identified in a hand search of eight journals from 1993 to 1997. Simulations were performed to assess the power of a weighted regression method and a rank correlation test in the presence of no bias, moderate bias or severe bias. We based evidence of small-study effects on P < 0.1. The power to detect bias increased with increasing numbers of trials. The rank correlation test was less powerful than the regression method. For example, assuming a control group event rate of 20% and no treatment effect, moderate bias was detected with the regression test in 13.7%, 23.5%, 40.1% and 51.6% of meta-analyses with 5, 10, 20 and 30 trials. The corresponding figures for the correlation test were 8.5%, 14.7%, 20.4% and 26.0%, respectively. Severe bias was detected with the regression method in 23.5%, 56.1%, 88.3% and 95.9% of meta-analyses with 5, 10, 20 and 30 trials, as compared to 11.9%, 31.1%, 45.3% and 65.4% with the correlation test. Similar results were obtained in simulations incorporating moderate treatment effects. However the regression method gave false-positive rates which were too high in some situations (large treatment effects, or few events per trial, or all trials of similar sizes). Using the regression method, evidence of small-study effects was present in 21 (26.9%) of the 78 published meta-analyses. Tests for small-study effects should routinely be performed in meta-analysis. Their power is however limited, particularly for moderate amounts of bias or meta-analyses based on a small number of small studies. When evidence of small-study effects is found, careful consideration should be given to possible explanations for these in the reporting of the meta-analysis. PMID- 11106887 TI - Combined evidence from multiple outcomes in a clinical trial. AB - Clinical investigators are encouraged to apply recently developed statistical methodology. For each patient in a trial, favorable and unfavorable results from multiple outcomes may be summarized in a suitable summary measure. This summary measure may be used in a two-sample t-test to decide which treatment is best. An example illustrates how the evidence from the main outcome criteria may be combined. The required study size depends on the mean treatment effect on the outcomes in the summary measure. When separate outcomes are considered, there is a multiple comparisons problem, for which Hochberg offered a simple solution. Evaluation of a single-summary measure may require a larger or a smaller study size than evaluation of separate outcomes, depending on whether treatment effects are about the same or very different. PMID- 11106886 TI - Weighting bias in meta-analysis of binary outcomes. AB - This article demonstrates that the weighting-according-to-the-variance method may introduce biases in meta-analyses of binary outcomes. The weighting favors studies that have certain frequencies of outcome events and weights given to studies of the same size may differ tens to thousands of times merely because of variations in the frequency. It also applies different standards to different measures of effect. Thus, the weighting may distort the combined result or even lead to contradictory conclusions when different measures of effect are used. Generally, the bias is more likely to arise when the effect is heterogeneous across the combined trials, the trials are conducted in populations of highly varied risks, the relative risk is used as the effect measure, the effect to be combined is small, any of the trials falls beyond the risk range of 20% and 80%, and/or the number of trials is small. Suggestions for detection and control of the bias are also given. PMID- 11106888 TI - Competing risks in absence of independence: impact of AIDS on liver function failure mortality, and lung cancer on ischemic heart disease mortality. AB - The increase in lung cancer (LC) mortality can produce a decrease in mortality from other causes, including ischemic heart disease (IHD). This problem (called the competing risks problem) has been addressed usually assuming independence between the competing causes of death. Our purpose is to show that assuming dependence of causes of death allows obtaining a better estimation of cumulative mortality. We use a clinical epidemiological example on the impact of AIDS in liver function failure in a cohort of drug users. The competing effect under dependence is 47% higher than under independence. This result is compared with a population-based example on LC and IHD mortalities in Spanish people in 1992. LC and IHD share tobacco smoking as a common risk factor, so independence cannot be assumed. Under the independence assumption, both life expectancy and number of deaths from IHD are underestimated. The difference is small compared to the model computed under dependence and it occurs mainly in the elderly (0.3% more deaths in people aged 70 and over). PMID- 11106889 TI - A community-based epidemiological survey of female urinary incontinence: the Norwegian EPINCONT study. Epidemiology of Incontinence in the County of Nord Trondelag. AB - OBJECTIVES: The aim was to assess the prevalence of any urinary leakage in an unselected female population in Norway, and to estimate the prevalence of significant incontinence. METHODS: The EPINCONT Study is part of a large survey (HUNT 2) performed in a county in Norway during 1995-97. Everyone aged 20 years or more was invited. 27,936 (80%) of 34,755 community-dwelling women answered a questionnaire. A validated severity index was used to assess severity. RESULTS: Twenty-five percent of the participating women had urinary leakage. Nearly 7% had significant incontinence, defined as moderate or severe incontinence that was experienced as bothersome. The prevalence of incontinence increased with increasing age. Half of the incontinence was of stress type, 11% had urge and 36% mixed incontinence. CONCLUSIONS: Urinary leakage is highly prevalent. Seven percent have significant incontinence and should be regarded as potential patients. PMID- 11106890 TI - Estimating the prevalence of hypertension corrected for the effect of within person variability in blood pressure. AB - The objective of our study was to assess the applicability of using estimates of within-person variance (WPV) from reproducibility studies for a correction of blood pressure values in another study to improve the accuracy of the prevalence estimation of hypertension. Data were collected from two cross-sectional population-based studies on cardiovascular disease risk factors conducted from 1987 to 1995 among 55,026 subjects aged 20-59 years. Correction factors were calculated from a reproducibility study among 924 subjects who were examined in 1989 and 1990. All other studies with repeated measurements of blood pressure were searched in MEDLINE from 1966 onward. Six studies satisfied the inclusion criteria. The prevalence of hypertension, uncorrected and corrected with factors from other studies, were compared with the prevalence of hypertension corrected with the factor from our study. The uncorrected prevalence of hypertension was 17.3% [95%CI:17.0-17.7]. The prevalence of hypertension after correction for WPV with the factor from our study was 13.5% [95%CI:13.2-13.8]. Correction for WPV with factors from the appropriate studies (depending on factors such as number of measurements taken per visit, and time interval between visits) resulted in prevalences ranging from 13.9% to 14.7%. The bias that occurs when no correction for WPV is performed is much larger (29% overestimation) than the bias that occurs when correction factors are derived from other studies (3.1-8.4% overestimation). When repeated measurements of blood pressure are not available in a population study for a sample of that same study, it is advisable to use data from another study to correct for WPV. PMID- 11106891 TI - Primary hyperparathyroidism detected in a health screening. The Tromso study. AB - Serum calcium was measured in 12,339 men and 13,394 women ages 25 to 75. Primary hyperparathyroidism, defined as a combination of serum calcium and parathyroid hormone (PTH) levels within the extreme or upper normal range, was diagnosed in 17 men and 47 women. The prevalence in both sexes increased with age. When 42 subjects with asymptomatic primary hyperparathyroidism were followed for 3 years, no significant increase in serum calcium or PTH was seen. In a subgroup of 473 men and 517 women ages 50 to 75, serum PTH was measured along with serum calcium. Depending on the criteria used to define primary hyperparathyroidism, the prevalence in older women within this subgroup ranged from 3.6% to 13.9%. The study concluded that a high prevalence of primary hyperparathyroidism exists in older women, although the progression of the disease, judging by serum calcium and PTH measurements, appears to be very slow. PMID- 11106893 TI - Cholecystectomy in Sweden 1989 and 1994: long admissions assessed by the inpatient registry. AB - The purpose of this study was to compare cholecystectomy in Sweden (pop. 8.9 million) 1989 to 1994 when the diffusion of laparoscopic cholecystectomy (LC) was completed, focusing on long hospital admissions as a proxy indicator of adverse events. This was an observational study of all patients operated on with cholecystectomy in 1989 and 1994 (n = 19,432) from the National Inpatient Registry. The risk of a long admission was analyzed by multivariate analyses. Odds ratios of long admissions were computed considering gender, age groups, acute or chronic gallstone disease, 1989 and 1994, county level of operations per 1000 inhabitants, and hospital categories. Stratified analyses were performed by acuteness of disease, and year. Long admissions were defined as lasting longer than 20 days in 1989 and 14 days in 1994. Odds ratios of a long admission increased steeply with age and acute gallstone disease. The county level of operations per 1000 inhabitants had no influence on risk nor did hospital category. The absolute number of those operated on with an acute gallstone disease changed little between 1989 and 1994, whereas operations for chronic disease increased significantly. Stratification revealed that their risk of a long admission was increased both in 1989 and 1994, particularly for women. Those with chronic gallstone disease had no increased risk. After the introduction of the laparoscope and a rise in the number of cholecystectomies, patients with chronic gallstone disease seem to have a constant risk of long hospital stay. However, because patients with acute disease had an increased risk in both 1989 and 1994, further longitudinal analyses are needed to analyze the level of complications in this group. PMID- 11106892 TI - Menstrual patterns and risk of adult-onset diabetes mellitus. AB - We examined the association between menstrual patterns and risk of developing adult-onset diabetes in a prospective study of 668 white, college-educated women who completed menstrual diaries throughout their reproductive years. We calculated summary measures of cycle length and variability and bleeding duration for ages < or = 22, 23-27, 28-32, and 33-37 years. The analysis included 35,418 person-years of follow-up and 49 self-reported cases of diabetes (median age at diagnosis, 63 years). There was no association between diabetes risk and age at menarche, mean cycle length, cycle variability, or frequency of long cycles (> 42 days). Longer bleeding periods in the mid- and late reproductive years were somewhat associated with an increased risk of diabetes (adjusted rate ratio 1.4, 95% confidence interval 1.0-1.8 per day increase in bleeding duration for menses during ages 28-32). These results do not support the association of long or irregular menstrual cycles with post-menopausal diabetes incidence, but do suggest a possible association of longer bleeding duration with subsequent onset of diabetes. PMID- 11106895 TI - PTGS in plants. PMID- 11106894 TI - Do patient factors alter the relationship between physician characteristics and use of long-acting benzodiazepines? AB - Despite known hazards associated with their use, long-acting benzodiazepines are frequently used in the treatment of older adults. While such use has been linked to physician characteristics, the effect of patient factors has not been considered. To investigate this, data from 1423 Quebec community-dwelling subjects of the Canadian Study of Health and Aging were linked to records of prescriptions billed to the provincial health insurance program during the year following study entry. The standardized one-year period prevalence of any use of long-acting benzodiazepines was 12.2%. Among benzodiazepine users, long-acting benzodiazepine use was more common among male patients and patients of earlier graduating prescribers and specialist prescribers. However, the effect of the latter two factors were modified by patient self-reported anxiety. This study demonstrates that consideration of patient factors may be necessary to obtain an accurate estimate of the association between at least some physician factors and use of long-acting benzodiazepines. PMID- 11106896 TI - Milk as a fungicide. PMID- 11106897 TI - USDA estimate of GM crops. PMID- 11106898 TI - 40% more GM crops in 1999. PMID- 11106900 TI - Saving crop diversity. PMID- 11106899 TI - Health benefits of GM beet. PMID- 11106901 TI - New website and pesticide research updates. PMID- 11106902 TI - Strawberry fruit genes. PMID- 11106904 TI - Biodiversity as insurance. PMID- 11106903 TI - Growing rice in Canada. PMID- 11106905 TI - GM plants communicate with bacteria. PMID- 11106906 TI - Detoxification confers bacterial resistance. PMID- 11106907 TI - US grain group wants GM-crop tests. PMID- 11106908 TI - Single-copy transgenes. PMID- 11106909 TI - Transient expression in tobacco leaves. PMID- 11106910 TI - Potato website announcement. PMID- 11106911 TI - Mussel power. PMID- 11106912 TI - Insect enzymes for bioremediation. PMID- 11106913 TI - The assessment of growth hormone deficiency in children and adults with particular reference to the transitional period. PMID- 11106914 TI - Cabergoline and quinagolide therapy for prolactinomas. PMID- 11106915 TI - Elderly people with hypothalamic-pituitary disease and growth hormone deficiency: lipid profiles, body composition and quality of life compared with control subjects. AB - OBJECTIVE: In healthy adults the secretion of growth hormone (GH) and insulin like growth factor 1 (IGF-1) declines with ageing and body composition alters, particularly with an increase in total body fat. In elderly people, hypothalamic pituitary disease can cause GH deficiency (GHD), compared with age matched controls. This study aimed to clarify whether GHD in the elderly is associated with differences in body composition, circulating lipid levels and quality of life (QOL) compared with control subjects. SUBJECTS: Twenty-seven elderly patients (14 males, mean age 71 years, range 65-83) with hypothalamic-pituitary disorders (23 pituitary tumours) and GHD (mean (SD) peak stimulated GH response 1.6 mIU/l (1.03) range 0.6-5) were studied. Twenty-five patients had been treated surgically (six cranial surgery, 19 transsphenoidal) and eight patients had received external cranial irradiation. Twenty-seven control subjects (14 males, mean age 72 years, range 65-86) were also studied. METHODS: Weight, body mass index (BMI), total fat mass (FM, bioelectrical impedance), waist to hip ratio (WHR), serum IGF-1, fasting blood glucose and lipid profile were measured. QOL was assessed in both groups using five interviewer administered self-rating questionnaires: The Nottingham Health Profile, Short-Form 36, Hospital Anxiety and Depression Scale, Mental Fatigue Questionnaire and Life Fulfilment Scale. The GHD group also completed the Disease Impact Scale. RESULTS: The data (mean (SD)) from males and females were analyzed separately. The male patients had a higher BMI than controls, 28.9(4.5) vs. 25.2(2.3) kg/m2 (P = 0.01) but the BMI in the female patients and controls was similar. In the female patients compared with the controls, FM was higher 39. 4(6) vs. 33.1(8.3) % (P = 0.02), WHR was also higher 0.9(0.08) vs. 0. 83(0.09) (P = 0.03) and serum IGF-1 levels were lower 10.8(6.4) vs. 18.2(6.5) nmol/l (P = 0.01). However, in the male patients, FM, WHR and IGF-1 levels were similar to the controls. Fasting blood glucose was similar in both male and female patients and the controls. Two female patients and one male control subject were taking lipid-lowering agents and were therefore excluded from the analysis of lipid profiles. Total cholesterol, triglyceride, LDL cholesterol, HDL cholesterol and total cholesterol/HDL cholesterol ratio were not significantly different for both male and female patients compared with the controls. The 27 patients with GHD reported significantly less energy (P < 0.05), mobility (P < 0.05) and personal life fulfillment (P < 0.01) than the 27 controls. There were significantly more problems with emotional reaction (P < 0.01), social isolation (P < 0.05) and mental fatigue (P < 0.05). Additionally the GHD group reported more impairment in areas of social functioning (P < 0.05), general health (P < 0.05) and mental health (P < 0.05). The GHD group reported a modest degree of disease impact (mean score of 14.1). There were no significant differences in the domains of material life fulfillment, pain, sleep, physical functioning, vitality, anxiety, depression, self-esteem or role physical functioning compared with the controls. CONCLUSION: Compared with control subjects, the elderly female patients with hypothalamic-pituitary disease and GHD had a significantly higher total fat mass, with the WHR indicating a more central fat distribution and lower female serum IGF-1 levels. In contrast, elderly male patients had similar total fat mass, WHR and IGF-1 levels compared to the controls. There were no significant differences in the lipid profiles between male or female patients compared to controls. However, many of the male patients were receiving androgen replacement which might have influenced these results. Low HDL cholesterol concentrations are probably a better predictor of future cardiovascular disease than raised LDL cholesterol levels in the elderly population and these were similar in patients and controls for both PMID- 11106916 TI - Effects of DHEA replacement on bone mineral density and body composition in elderly women and men. AB - OBJECTIVE: Dehydroepiandrosterone (DHEA) is a precursor for both oestrogens and androgens. Its marked decline with ageing may influence age-related changes in tissues influenced by sex hormones. The aim of this study was to determine the effects of DHEA replacement on bone mineral density (BMD) and body composition in elderly women and men with low serum DHEA sulphate (DHEAS) levels. DESIGN: Prospective 6 month trial of oral DHEA replacement, 50 mg/day. PATIENTS: Experimental subjects were 10 women and eight men, aged 73 +/- 1 years. Control subjects were 10 women and eight men, aged 74 +/- 1 years. MEASUREMENTS: BMD, body composition, serum markers of bone turnover, serum lipids and lipoproteins, oral glucose tolerance, serum IGF-I, total serum oestrogens and testosterone. RESULTS: BMD of the total body and lumbar spine increased (mean +/- SEM; 1.6 +/- 0.6% and 2.5 +/- 0.8%, respectively; both P < or = 0.05), fat mass decreased (- 1.3 +/- 0.4 kg; P < 0.01) and fat-free mass increased (0.9 +/- 0.4 kg; P < or = 0. 05) in response to DHEA replacement. DHEA replacement also resulted in increases in serum IGF-I (from 108 +/- 8 to 143 +/- 7 microg/l; P < 0.01) and total serum testosterone concentrations (from 10.7 +/- 1.2 to 15.6 +/- 1.8 nmol/l in the men and from 2.1 +/- 0.2 to 4.5 +/- 0.4 nmol/l in the women; both P < or = 0.05). CONCLUSIONS: The results provide preliminary evidence that DHEA replacement in those elderly women and men who have very low serum DHEAS levels can partially reverse age-related changes in fat mass, fat-free mass, and BMD, and raise the possibility that increases in IGF-I and/or testosterone play a role in mediating these effects of DHEA. PMID- 11106917 TI - Beta cell function declines with age in glucose tolerant Caucasians. AB - OBJECTIVE: As type 2 diabetes results from an imbalance between insulin sensitivity and beta cell function, either or both may worsen with age. However, existing data are controversial on the effect of ageing on both insulin sensitivity and beta cell function. SUBJECTS AND DESIGN: We enrolled 149 healthy, glucose tolerant and normotensive Caucasians (age 35 +/- 1 years, body mass index 26.07 +/- 0.44 kg/m2, waist-hip ratio 0.842 +/- 0.009 cm/cm, mean +/- standard error). A cross-sectional study was designed to examine the impact of age on insulin sensitivity and beta cell function. Their beta cell function (percentage B [%B]) and insulin sensitivity (percentage S [%S]) were estimated using the homeostasis model assessment. RESULTS: Simple regression analysis revealed that %B declined with age (P = 0.008) while no relation was found between %S and age (P = 0.769). A stepwise regression analysis revealed that body mass index and diastolic blood pressure explained 14.7% of variation in %S, while age, waist-hip ratio, gender, and systolic blood pressure had no influence on %S. Age, body mass index and diastolic blood pressure together accounted for 21.7% of variation in %B, with age being an independent variable. CONCLUSIONS: In the present study, we showed that beta cell function declined with age at a rate of about 1% per year. In contrast, insulin sensitivity was not affected by ageing. Our observations suggest that the age-related decline in glucose tolerance is primarily related to the loss of beta-cell function. PMID- 11106918 TI - Comparison of octreotide acetate LAR and lanreotide SR in patients with acromegaly. AB - BACKGROUND AND OBJECTIVE: The most effective option for the medical treatment of patients with acromegaly is the use of somatostatin analogues. Long-acting depot formulations for intramuscular injection of two somatostatin analogues have recently become available: octreotide acetate LAR (Sandostatin LAR, Novartis Pharma AG) and lanreotide SR (Somatuline, Ipsen Biotech). We wished to compare efficacy of octreotide LAR and lanreotide SR in acromegalic patients. PATIENTS AND METHODS: A group of 125 patients with acromegaly (67 females; mean age, 47 years; 59 patients had previous pituitary irradiation) from 26 medical centres in France, Spain and Germany were studied. Before the study, all patients had been treated with intramuscular injections of lanreotide SR (mean duration, 26 months) at a dose of 30 mg which was injected every 10 days in 64 and every 14 days in 61 patients, respectively. All patients were switched from lanreotide SR to intramuscular injections of 20 mg of octreotide LAR once monthly for three months. In order to obtain efficacy and safety data of lanreotide SR under study conditions, it was decided to randomly assign at day 1, in a 3 : 1 ratio, the time point of the treatment switch; 27 of the patients were randomly assigned to continue the lanreotide SR treatment for the first 3 months of the study (group A); they were on octreotide LAR 20 mg from month 4-6. The other 98 patients were assigned to be switched to treatment with octreotide LAR 20 mg at day 1 (group B). In group B patients, octreotide LAR treatment was continued until month 6, with an adjustment of the dose based on GH levels obtained at month 3. RESULTS: The mean GH concentration decreased from 9.6 +/- 1.3 mU/l at the last evaluation on lanreotide SR to 6.8 +/- 1.0 mU/l after three injections of octreotide LAR (P < 0.001). The percentages of patients with mean GH values < or = 6.5 mU/l (2.5 microg/l) and < or = 2.6 mU/l (1.0 microg/l) at the last evaluation on lanreotide SR were 54% and 14%, and these values increased after 3 months treatment with octreotide LAR to 68% and 35% (P < 0.001), respectively. IGF-I levels were normal in 48% at the last evaluation on lanreotide SR and in 65% after 3 months on octreotide LAR (P < 0.001). Patients with pre-study pituitary irradiation had lower mean GH and IGF-I concentrations. But the effects of the treatment change did not differ between the irradiated and the nonirradiated patients. In general both drugs were well tolerated. CONCLUSION: Octreotide LAR 20 mg administered once monthly was more effective than lanreotide SR 30 mg administered 2 or 3 times monthly in reducing GH and IGF-I in patients with acromegaly. PMID- 11106919 TI - Cholinergic modulation of growth hormone responses to growth hormone-releasing hormone in uraemic patients on peritoneal dialysis. AB - BACKGROUND: Hypothalamic cholinergic neurotransmission plays a major role in the regulation of GH secretion. Pyridostigmine, a cholinesterase inhibitor, is able to decrease hypothalamic somatostatinergic tone and release GH in normal subjects. Blockade of muscarinic receptor with pirenzepine blunts the GH release in several clinical situations. However, little information is available on the role played by central cholinergic pathways in GH regulation in uraemic patients. OBJECTIVE: We aimed to assess GH responses to GHRH after pretreatment with pyridostigmine and pirenzepine in a group of uraemic patients undergoing peritoneal dialysis (PD). GH responses of the patients treated with recombinant human erythropeitin (rhEPO) were compared to patients without treatment. DESIGN: We studied 14 male patients on PD and nine control subjects. All subjects underwent three endocrine test in random order after an overnight fast. Each subject received GHRH (100 microg, i.v. in bolus at 0 minutes). Sixty minutes before the injection of GHRH subjects were given oral placebo, pyridostigmine (120 mg), or pirenzepine (100 mg). MEASUREMENTS: Blood samples for GH were collected at -60, 0, 15, 30, 45, 60 and 90 minutes The hormonal secretory responses were studied by a time-averaged (area under the curves, AUC) and time independent (peak values) analysis. RESULTS: Baseline GH concentrations were similar in patients and controls. GH responses to placebo plus GHRH were also comparable in patients and controls (peak 26.6 +/- 3.8 vs. 33.2 +/- 4.4 mU/l, AUC 28.2 +/- 3.4 vs. 27.8 +/- 4.6 mU/h/l). Pyridostigmine administration induced a significant potentiation of GH responses to GHRH both in patients (peak 43.2 +/- 5.2 mU/l, AUC 47.6 +/- 6.0 mU/h/l; P < 0.01) and in control subjects (peak 79.2 +/- 8.6 mU/l, AUC 78.0 +/- 9.4 mU/h/l; P < 0.01). However, the increment in GH peak and AUC was significantly (P < 0.05) greater in controls in relation to values found in patients. Pirenzepine administration induced an abolishment of GH release after GHRH stimulation both in PD patients (peak 5.4 +/- 2.6 mU/l, AUC 6.0 +/- 2.4 mU/h/l; P < 0.01) and in healthy controls (peak 3.8 +/- 0.6 mU/l, AUC 4.0 +/- 0.4 mU/h/l; P < 0.05). Responses to pyridostigmine plus GHRH and pirenzepine plus GHRH were similar in patients on chronic therapy with recombinant human erythropeitin and in patients without rhEPO therapy. CONCLUSION: These results suggest that the cholinergic regulation of GH release is preserved in uraemic patients on peritoneal dialysis. The significantly lower increase in GH response to GHRH induced by pyridostigmine suggests that cholinergic stimulatory tone is attenuated in patients in relation to control subjects. Long-term therapy with rhEPO seems not to affect GH responses to cholinergic stimulation or blockade. PMID- 11106920 TI - Recovery of growth hormone secretion following cabergoline treatment of macroprolactinomas. AB - OBJECTIVE: Cabergoline therapy normalizes prolactin levels and reduces the size of macroprolactinomas. However there are no data indicating whether cabergoline can normalize growth hormone secretion in patients who were growth hormone deficient at the time of diagnosis of a macroprolactinoma. SUBJECTS AND METHODS: We studied nine patients with biochemical and radiological evidence of a macroprolactinoma who were also growth hormone deficient (peak growth hormone response to insulin-induced hypoglycaemia < 10 mU/l). Patients were assessed before and after cabergoline therapy to assess their growth hormone secretory status, IGF-I levels, cortisol response and change in tumour size. RESULTS: Treatment with cabergoline was associated with a significant reduction in prolactin concentration (74341 +/- 31939 mU/l vs. 265.9 +/- 86.3, P = 0.009). The mean change in peak growth hormone response to insulin-induced hypoglycaemia was significantly greater following cabergoline therapy compared with pretreatment levels (33.5 +/- 11.8 mU/l vs. 4. 34 +/- 1.21 mU/l, P = 0.022). However IGF-I levels were not different after treatment when compared with baseline although a nonsignificant trend towards improvement was noted (24.2 +/- 3.97 nmol/l vs. 18.4 +/- 4.94 nmol/l, P = 0.058). The mean peak cortisol concentration was 407.7 +/- 64.1 nmol/l before treatment with a nonsignificant rise to 477.4 +/- 84.8 nmol/l, P = 0.813 after treatment. These changes were associated with a significant reduction in mean maximal tumour diameter (21.2 +/- 2.9 mm vs. 29.1 +/- 2.8 mm, P = 0.009). There was no significant difference in either prolactin concentration or tumour size pre- or post-treatment between those who recovered growth hormone secretion and those that did not. Six of the nine (67%) patients recovered a normal growth hormone response (> 10 mU/l) after cabergoline therapy. Those that remained growth hormone deficient after treatment were all panhypopituitary at baseline while those that recovered showed only partial anterior hypopituitarism. CONCLUSION: These data indicate that growth hormone secretion may recover following successful reduction of prolactin levels after cabergoline therapy for a mean of 22 months (range 6-28 months) in most but not all subjects with a macroprolactinoma. It is therefore advisable that individuals with a macroprolactinoma in whom growth hormone replacement therapy is being considered undergo repeat assessment of growth hormone secretion following medical treatment. PMID- 11106921 TI - Sweat secretion rates in growth hormone disorders. AB - BACKGROUND: While increased sweating is a prominent symptom in patients with active acromegaly, reduced sweating is gaining status as part of the growth hormone deficiency (GHD) syndrome. DESIGN AND SUBJECTS: Sweat secretion rate (SSR), as measured by pilocarpine iontophoresis represents the maximal capacity for stimulated sweat secretion in a localized skin area. SSR was studied in 37 patients with a history of acromegaly, 20 adult patients with GHD before and during long-term GH substitution of GHD adults, and 58 control subjects. RESULTS: Acromegaly: Patients with acromegaly had significantly higher SSR than healthy controls (Z-score + 1.9 (+/- 1.1) mean (+/- SD) (P < 0.001)). SSR was increased irrespective of current clinical disease activity. Thus, the SSR Z-scores in 16 clinically inactive patients were + 2.1 (+/- 1.2), in 10 slightly or doubtfully active patients + 1.5 (+/- 0.7) and in 11 active patients + 1.8 (+/- 1.3). There was no correlation between SSR and IGF-I. GHD: Twenty adult patients participated in an 18-month randomised, placebo controlled, double blinded study of physiological dose GH substitution, followed by 18 months of open GH treatment. SSR at baseline was reduced in male but not in female GHD patients. Mean SSR (95% confidence interval) for 11 male patients was 89.0 mg/30 minutes (51.9-126.1) as compared to 133.5 mg/30 minutes (59.2-259.9) (P = 0.01) in 24 male controls, and for 11 female patients 48.2 mg/30 minutes (25.9-70.6) as compared to 49.2 mg/30 minutes (12.6-93. 9) in 34 female controls. GH treatment in physiological substitution doses for up to 36 months had no effect on SSR. CONCLUSION: We have demonstrated that longstanding GH hypersecretion in patients with acromegaly induces irreversible changes of sweat gland function, with persistently elevated SSR despite treatment and clinical cure. In GHD patients, SSR was reduced in males but not in females, which together with the established gender difference in normal controls emphasises the role of androgen deficiency as a cofactor for reduced sweating in hypopituitary patients. Sweat gland development seems to be more susceptible to lack of hormones in childhood and adolescence than in adulthood, whereas growth hormone excess can modify sweat function later in life. PMID- 11106922 TI - The effect of distinct activating mutations of the luteinizing hormone receptor gene on the pituitary-gonadal axis in both sexes. AB - OBJECTIVE: Familial or sporadic male-limited precocious puberty is a distinct and unusual gonadotrophin-independent form of sexual precocity caused by constitutively activating mutations of the luteinizing hormone receptor (LHR). In the present study, we evaluated the effect of known activating mutations at different sites of the LHR gene on the pituitary-gonadal axis in both sexes. PATIENTS: Four unrelated Brazilian boys (I-IV) with gonadotrophin-independent precocious puberty and two asymptomatic females (V-VI), a sister and mother of two of the affected boys, were studied. Patients I, II and V carried the Ala568Val mutation located at the third intracellular loop of the LHR. Patient III carried the Leu457Arg mutation at the third transmembrane helix, and patients IV and VI carried the Thr577Ile mutation at the sixth transmembrane helix of the LHR. MEASUREMENTS: Serum levels of LH, FSH, testosterone, and oestradiol under basal and GnRH-stimulated conditions were determined in all patients. Testosterone levels were also measured after a hCG stimulation test in patient III. RESULTS: Basal LH and FSH levels were prepubertal in all boys studied. The GnRH-stimulated serum LH and FSH levels were prepubertal in three boys (I, II and IV), whereas patient III showed totally suppressed LH and FSH levels at ages 2 and 7 years (bone ages 6 and 14 years, respectively). Serum testosterone levels ranged from 3.8 to 69.5 nmol/l in the four boys. Patient III had the highest testosterone levels that did not respond to hCG stimulation. The 4 year-old girl (patient V) was phenotypically normal and the acute response to GnRH was indicative of prepubertal status. Patient VI had normal menstrual cycles and fertility. CONCLUSIONS: These findings indicate variable effects of LHR activating mutations on the pituitary-gonadal axis in boys that can result in lack of normal LH and FSH release. In contrast, prepubertal and adult females were asymptomatic and had normal basal and GnRH-stimulated LH and FSH levels. PMID- 11106923 TI - Salivary, but not serum or urinary levels of progesterone are elevated after topical application of progesterone cream to pre-and postmenopausal women. AB - OBJECTIVE: The use of topically applied micronised ('natural') progesterone as a substitute for synthetic oestrogens and progestogen preparations is controversial. The aim of this study was to examine the changes in blood and salivary concentrations of progesterone following a single topical application of a progesterone cream. PATIENTS AND MEASUREMENTS: We investigated six premenopausal women in the luteal phase and six postmenopausal women to determine the short-term changes in serum, urinary and salivary progesterone concentrations following a single 64 mg topical application of micronised progesterone. RESULTS: Serum progesterone concentrations did not increase during the first 3 hours after application of progesterone cream, however, salivary values rose significantly in both premenopausal and postmenopausal women, consistent with the view that progesterone is absorbed and transported through the body. Salivary progesterone concentrations were significantly elevated above basal levels by 30-60 minutes and reached peak levels at 1-4 h, with mean levels approximately fivefold higher in premenopausal, than in menopausal women. CONCLUSIONS: Salivary progesterone measurements confirm that topically applied progesterone is absorbed, despite the lack of change in serum progesterone concentrations. However, at the dose administered, serum progesterone levels do not reach those observed after oral or vaginally delivered progesterone preparations. Higher doses may be required to induce biological responses within the endometrium. PMID- 11106924 TI - Parathyroid hormone-related protein in the aetiology of fibrous dysplasia of bone in the McCune Albright syndrome. AB - OBJECTIVE: Fibrous dysplasia, observed in bone lesions in the McCune Albright syndrome (MAS), is thought to result from abnormalities in cells of the osteogenic lineage associated with over-activation of the cAMP signalling pathway in affected cells. The aim of this study was to investigate the role of parathyroid hormone-related protein (PTHrP) in the aetiology of MAS, and to determine a possible therapeutic role for 1,25-dihydroxy vitamin D(3) (1,25(OH)(2)D(3)). DESIGN: The effects of 1,25(OH)(2)D(3) on PTHrP production and mRNA expression were determined in vitro. 1,25(OH)(2)D(3) therapy was administered to three patients with MAS. PATIENTS: Clinical data from four MAS patients (MAS1, 2, 3 and 4), and in vitro studies using bone from three MAS patients (MAS1, 2, and 3), are presented. MEASUREMENTS: Immunoradiometric assay and low-cycle number reverse transcriptase-linked PCR were used to determine PTHrP production and mRNA expression in vitro. Standard clinical biochemistry was recorded pre and post commencement of 1,25(OH)(2)D(3) treatment. RESULTS: We report the elevated secretion of PTHrP, and a concomitant rise in PTHrP mRNA expression, in cultured osteoblasts from three MAS patients. Treatment with 1,25(OH)(2)D(3) produced a dose-dependent decrease in PTHrP protein secretion and mRNA expression. Marked improvement in bone biochemistry in MAS1, 2 and 3 post treatment with 1,25(OH)(2)D(3) is documented. CONCLUSION: This study provides the first evidence suggesting that PTHrP may contribute to the aetiology of fibrous dysplasia in MAS. In addition, the therapeutic administration of 1,25(OH)92)D(3) may provide clinicians with an important new regime for symptomatic relief of bone pain and fracture in some patients with MAS. PMID- 11106925 TI - Determinants of thyroid volume as measured by ultrasonography in healthy adults randomly selected. AB - OBJECTIVES: The relationship between thyroid volume and anthropometric characteristics is a matter of controversy. The aim of this study was to investigate thyroid volume and its determinants in healthy adult subjects from a noniodine-deficient area. DESIGN AND PATIENTS: Of the 280 000 inhabitants of the city, served by L'Hospitalet de Llobregat, we randomly selected 880 subjects from the census of the city. The participation rate in the study was 44%; after application of several exclusion criteria, a further 28 subjects were excluded because of previously diagnosed thyroid disease. We finally studied 268 subjects representative of the census of the city: 134 male and 134 female, without thyroid disease. We determined the anthropometric characteristics, body mass index, waist-hip ratio, body surface area; body composition by bioelectrical impedance analyser; thyroid volume by ultrasonography; basal TSH, antithyroid antibodies and urinary iodine excretion. RESULTS: Thyroid volume in our population was higher in males (9.19 ml, CI 9.09-10.65) than in females (6.19 ml, CI 6.02-6.92), P = 0.001. Significant correlations were found among thyroid volume and body weight (r = 0.39, P = 0.0001), height (r = 0.44, P = 0.0001), body mass index (r = 0.13, P = 0.02), waist-hip ratio (r = 0.38, P = 0.0001), body surface area (r = 0.48, P = 0.0001), total body water (r = 0.14, P = 0.02), free fat mass (r = 0.47, P = 0.0001), fat mass (r = 0.37, P = 0.001) and body fat (r = 0.32, P = 0.001). Negative correlation was found between thyroid volume and basal TSH (r = -0.26, P = 0.001). No correlations were found among thyroid volume and iodine excretion, previous pregnancies in women, cigarette smoking and alcohol consumption. In a multiple regression analysis with thyroid volume as the dependent variable, body surface area was demonstrated to account for the 44% of variation of thyroid volume (P = 0.0001). CONCLUSION: It is important to know the reference values of the thyroid volume in a population free of iodine deficiency and its determinants. Body surface area accounts for much of the variation of thyroid volume. Age, gender, anthropometric variables, body composition variables and biological variables, do not significantly influence the thyroid volume when considered as possible additions to this baseline model. PMID- 11106926 TI - Over-expression of hepatocyte growth factor/scatter factor (HGF/SF) and the HGF/SF receptor (cMET) are associated with a high risk of metastasis and recurrence for children and young adults with papillary thyroid carcinoma. AB - OBJECTIVE: The study determined if hepatocyte growth factor/scatter factor (HGF/SF) or the HGF/SF receptor (cMET) might be important for metastasis in thyroid cancer. DESIGN: We examined HGF/SF and cMET expression by immunohistochemistry in a retrospective group of benign and malignant thyroid lesions from children and young adults, and correlated the intensity of expression with clinical outcome. PATIENTS: Patients included 42 children and young adults with papillary thyroid carcinomas (PTC), seven with follicular thyroid carcinomas (FTC), two with medullary thyroid carcinomas (MTC), 14 with benign thyroid disorders, and two with normal thyroids. MEASUREMENTS: Expression of cMET was graded from 0 (absent) to 4 (intense); and HGF/SF expression was graded from 0 (absent-minimal) to 3 (diffuse and intense). RESULTS: cMET staining was greater in PTC (mean intensity 2.3 +/- 0.4 vs. 0.8 +/- 0.2, P < 0.005) and FTC (2.4 +/- 0.6 vs. 0.8 +/- 0.2, P = 0.04) than benign lesions (0.8 +/- 0.2) or normal thyroids (0.4 +/- 0.5). PTC with intense cMET staining had shorter disease free survival (P = 0.05) and increased HGF/SF staining (r = 0.39, P = 0.017). HGF/SF correlated with the extent of disease at diagnosis (r = 0.33, P = 0.049). Patients with PTC were stratified into quartiles based on combined cMET and HGF/SF staining. Those with intense cMET and HGF/SF staining were younger (P = 0.05), and had reduced disease free survival (P = 0.03). CONCLUSIONS: We conclude that increased cMET and HGF/SF expression is associated with a high risk for metastasis and recurrence in children and young adults with papillary thyroid carcinoma. PMID- 11106927 TI - Placental growth hormone and IGF-I in a pregnant woman with Pit-1 deficiency. AB - The respective contributions of pituitary and placental GH to circulating IGF-I in pregnant women have not been well established. We measured the serum concentrations of placental growth hormone (PGH) and IGF-I in a woman with pit-1 deficiency before, during and after pregnancy, resulting in the birth of a healthy child (not pit-1 deficient). Both PGH and IGF-I concentrations were below the assay detection limit before and after pregnancy. During pregnancy, PGH and IGF-I levels increased steadily; the concentrations of PGH and IGF-I in late pregnancy were comparable with levels previously measured in normal pregnancies. PGH and IGF-I concentrations were strongly correlated throughout pregnancy (r = 0.90; P = 0.002). PGH was undetectable in cord serum, whilst the IGF-I concentration was within the normal range. The findings of this case study corroborate the notion that PGH is the prime regulator of maternal serum IGF-I during pregnancy. PMID- 11106928 TI - Simultaneous peripubertal onset of multireactive autoimmune diseases with an unusual long-lasting remission of type 1 diabetes mellitus. AB - Although it is well known that patients with type 1 diabetes mellitus are susceptible to other autoimmune diseases, the simultaneous occurrence of clustered distinct autoimmune diseases is uncommon. We report a 16-year-old girl, previously diagnosed as having coeliac disease and IgA deficiency, who at 13 years of age developed a clustering of distinct autoimmune diseases, including type 1 diabetes mellitus, rheumatoid arthritis (RA) and euthyroid autoimmune thyroiditis, eventually resulting in a simultaneous long-term remission. The clinical picture was associated with a functional immunodeficiency characterized by a defect in proliferative responses to T cell predominant mitogens and a normal response to the B cell predominant mitogen. In addition, the T cell activation markers HLA-DR, IL-2 receptor and transferrin receptor) were not upregulated. The clinical course of this immunodeficiency paralleled the outcome of the autoimmune diseases. After the abrupt onset, spontaneous clinical remission of both diabetes mellitus and RA was observed. Insulin was first reduced in dose and then discontinued completely at 15 months, in the presence of normal C peptide secretion and normal metabolic control (HbA1c 5.8%). Anti glutamate decarboxylase (GAD65) and anti-IA-2 antibodies remained persistently high. During the remission phase a normalization of the functional immune defect was observed. The gradual resolution of the multisystemic diseases as well as the normalization of immune function in our patient is unusual. This case may be of considerable value in furthering our knowledge of the immunological mechanisms implicated in these rare multireactive syndromes. PMID- 11106929 TI - Influence of thyroid pathology on the results of parathyroid gammagraphy with Tc(99m)-sestamibi. PMID- 11106930 TI - Cutaneous pseudolymphoma associated with bromocriptine therapy. PMID- 11106931 TI - Review article: molecular signals and genetic reprogramming in peripheral T-cell differentiation. AB - Rearrangement of gene segments occurs in T lymphocytes during thymic development as the T-cell receptor (TCR) is first expressed, allowing T cells to become central regulators of antigen specificity in the acquired immune system. However, further development of T cells occurs after population of peripheral lymphoid tissues, which can result in T-cell expansion and differentiation into effectors of various immune function, or progression to memory T cells, anergic cells or death by apoptosis. This review focuses on more recent developments concerning the choices that peripheral T cells make between first encountering antigen through TCR recognition and death. These decisions are associated with a process of genetic reprogramming that alters the behaviour of cells so that immune responses are appropriately regulated. PMID- 11106932 TI - Nitric oxide and virus infection. AB - Nitric oxide (NO) has complex and diverse functions in physiological and pathophysiological phenomena. The mechanisms of many events induced by NO are now well defined, so that a fundamental understanding of NO biology is almost established. Accumulated evidence suggests that NO and oxygen radicals such as superoxide are key molecules in the pathogenesis of various infectious diseases. NO biosynthesis, particularly through expression of an inducible NO synthase (iNOS), occurs in a variety of microbial infections. Although antimicrobial activity of NO is appreciated for bacteria and protozoa, NO has opposing effects in virus infections such as influenza virus pneumonia and certain other neurotropic virus infections. iNOS produces an excessive amount of NO for long periods, which allows generation of a highly reactive nitrogen oxide species, peroxynitrite, via a radical coupling reaction of NO with superoxide. Thus, peroxynitrite causes oxidative tissue injury through potent oxidation and nitration reactions of various biomolecules. NO also appears to affect a host's immune response, with immunopathological consequences. For example, overproduction of NO in virus infections in mice is reported to suppress type 1 helper T-cell-dependent immune responses, leading to type 2 helper T-cell-biased immunological host responses. Thus, NO may be a host response modulator rather than a simple antiviral agent. The unique biological properties of NO are further illustrated by our recent data suggesting that viral mutation and evolution may be accelerated by NO-induced oxidative stress. Here, we discuss these multiple roles of NO in pathogenesis of virus infections as related to both non-specific inflammatory responses and immunological host reactions modulated by NO during infections in vivo. PMID- 11106933 TI - The calcium-independent protein kinase C participates in an early process of CD3/CD28-mediated induction of thymocyte apoptosis. AB - Thymocyte negative selection eliminates self-reactive clones and involves both a T-cell receptor (TCR)/CD3-mediated signal and a costimulatory signal, which can be delivered via CD28. Anti-CD3/anti-CD28-triggered apoptosis in isolated CD4+CD8+ thymocytes in vitro provides a basic model for negative selection. Effects of isoform-selective and non-isoform-selective inhibitors of protein kinase C (PKC) on this apoptotic process suggest that activation of Ca2+ independent PKC isoforms during the first 2-3 hr of culture is essential for inducing apoptosis, and that Ca2+-dependent PKC isoforms may be influential, but not essential, for apoptosis. To assess the CD3/CD28-mediated activation of PKC in the apoptotic process, we prepared CD4+CD8+ thymocytes (without contamination with cells that had received negative or positive selection signals in vivo) by establishing TCR-transgenic mice with RAG-2-deficient and non-selecting major histocompatibility complex (MHC) backgrounds, in addition to a CD4+CD8+ thymocyte enriched population from normal mice. Translocation of Ca2+-independent PKC from the cytosolic fraction to the particulate fraction of CD4+CD8+ thymocytes was induced by CD3/CD28-mediated stimulation, but not by CD3- or CD28-mediated stimulation alone, and peaked 2 hr after the start of culture. The kinase activity of the translocated Ca2+-independent PKC was dependent on cofactors in vitro, indicating that novel (n)PKC, but not atypical (a)PKC or a proteolytic PKC fragment, was responsible for the activity. Immunoblotting analysis indicated that the nPKC-theta isoform was the major contributor among nPKC isoforms, and that the classical (c)PKC-alpha isoform was the major contributor among cPKC isoforms. These results suggest that activation of nPKC (especially the theta isoform) in CD4+CD8+ thymocytes is involved in a pathway for negative selection. PMID- 11106934 TI - Selection and characterization of MUC1-specific CD8+ T cells from MUC1 transgenic mice immunized with dendritic-carcinoma fusion cells. AB - Mice transgenic for the human MUC1 carcinoma-associated antigen (MUC1.Tg) are tolerant to immunization with MUC1 antigen. Recent studies, however, have demonstrated that immunization of MUC1.Tg mice with fusions of MUC1-positive tumour and dendritic cells (FC/MUC1) reverses MUC1 unresponsiveness and results in rejection of established MUC1-positive pulmonary metastases. Here we demonstrate that lymph node cells from MUC1.Tg mice immunized with the FC/MUC1 fusion cells proliferate in response to MUC1 antigen by a mechanism dependent on the function of CD4, major histocompatibility complex (MHC) class II, B7-1, B7-2, CD28, CD40 and CD40 ligand. The findings demonstrate that stimulation of lymph node cells with MUC1 results in selection of MUC1-specific CD8+ T cells. We show that the CD8+ T cells exhibit MUC1-specific cytotoxic T lymphocyte (CTL) activity by recognition of MUC1 peptides presented in the context of MHC class I molecules Kb and Db. The MUC1-specific CD8+ T cells also exhibit antitumour activity against MUC1-positive metastases, but with no apparent reactivity against normal tissues. These results indicate that immunization of MUC1.Tg mice with FC/MUC1 reverses immunological unresponsiveness to MUC1 by presentation of MUC1 peptides in the presence of costimulatory signals and generates MHC-restricted MUC1 specific CD8+ T cells. PMID- 11106935 TI - An alphabeta T-cell-independent immunoprotective response towards gut coccidia is supported by gammadelta cells. AB - Although gammadelta cells are commonly hypothesized to provide a 'first line of defence', gammadelta-cell-deficient mice are generally only marginally more susceptible to pathogens. Because gammadelta cells are enriched within epithelia, it is important to resolve whether immunoprotective capacity towards epithelial tropic pathogens is absent from the gammadelta-cell compartment, or whether such activity is present but simply redundant with that of alphabeta T cells. In this work, following infection of the intestinal epithelium of alphabeta T-cell deficient mice with the coccidian parasite, Eimeria vermiformis, gammadelta cells were shown to support the rapid activation of other lymphoid cells and to confer a transferable antipathogen effect that could be eradicated by neutralization of interferon-gamma. However, unlike alphabeta T cells, these effects of gammadelta cells showed no evidence of functional immunological memory. These results are directly relevant to coccidiosis, an economically significant disease of livestock, and should have general relevance to infections involving alphabeta T cell deficiencies, e.g. cryptosporidiosis in patients with acquired immune deficiency syndrome (AIDS). PMID- 11106936 TI - Antigen-specific proliferation of porcine CD8alphaalpha cells to an extracellular bacterial pathogen. AB - A vaccine inducing protective immunity to a spirochaete-induced colitis of pigs predominantly stimulates expansion of CD8+ cells in vivo and in antigen stimulated lymphocyte cultures. CD8+ cells, however, are rarely considered necessary for protection against extracellular bacterial pathogens. In the present study, pigs recovering from colitis resulting from experimental infection with Brachyspira (Serpulina) hyodysenteriae had increased percentages of peripheral blood CD4- CD8+ (alphaalpha-expressing) cells compared with non infected pigs. CD8alphaalpha+ cells proliferated in antigen-stimulated cultures of peripheral blood mononuclear cells from B. hyodysenteriae-vaccinated pigs. Proliferating CD8alphaalpha+ cells consisted of CD4-, CD4+ and gammadelta T-cell receptor-positive cells. CD4- CD8alphabeta+ cells from vaccinated or infected pigs did not proliferate upon in vitro antigen stimulation. Of the CD8alphaalpha cells that had proliferated, flow cytometric analysis indicated that the majority of the CD4+ CD8+ cells were large (i.e. lymphoblasts) whereas the CD4- CD8+ cells were predominantly small. Addition of monoclonal antibodies (mAb) specific for either porcine major histocompatibility complex (MHC) class I or class II antigens diminished B. hyodysenteriae-specific proliferative responses whereas addition of mAb to porcine MHC II, but not porcine MHC I, reduced the CD8alphaalpha response. In vitro depletion of CD4+ cells by flow cytometric cell sorting diminished, but did not completely abrogate, the proliferative response of cells from vaccinated pigs to B. hyodysenteriae antigen stimulation. These results suggest that CD8alphaalpha cells are involved in recovery and possibly protection from a spirochaete-induced colitis of pigs; yet, this response appears to be partially dependent upon CD4+ cells. PMID- 11106937 TI - A novel subset of murine B cells that expresses unmasked forms of CD22 is enriched in the bone marrow: implications for B-cell homing to the bone marrow. AB - CD22 is a B-cell-restricted transmembrane protein, which acts as a negative regulator of B-cell signalling. CD22 also has lectin-like adhesive properties. When expressed on transfected fibroblasts, it is capable of mediating adhesion to other cells via recognition of cell-surface glycoconjugates terminating in alpha2,6-linked sialic acids. In previous studies in the mouse, CD22 was implicated as a bone marrow homing receptor for recirculating immunoglobulin D+ (IgD+) B cells through recognition of sialylated ligands on marrow sinusoidal endothelium. As the adhesive function of CD22 can be masked when alpha2,6-linked sialic acids are co-expressed at the cell surface, the aim of the present study was to investigate whether recirculating B cells have unmasked forms of CD22 that could be involved in bone marrow homing. Using alpha2,6-sialyllactose coupled to biotinylated polyacrylamide as a probe for detection of unmasked CD22, we showed that approximately 2-5% of IgD+ murine B cells in the spleen and mesenteric lymph nodes were able to bind this synthetic ligand. In the bone marrow, however, the fraction of IgD+ B cells with unmasked CD22 was increased by two- to fivefold. B cells from CD22-deficient mice were not stained with the polyacrylamide probe, confirming that staining of B cells in wild-type mice was caused by CD22 and not by other potential sialic acid-binding lectins. In conclusion, we have identified a new subset of mature B cells in the mouse with unmasked CD22. This subset of recirculating B cells may bind to CD22 ligands on bone marrow sinusoidal endothelium, leading to their selective homing and subsequent enrichment in this tissue. PMID- 11106938 TI - Mapping of the chicken immunoglobulin heavy-chain constant region gene locus reveals an inverted alpha gene upstream of a condensed upsilon gene. AB - Chicken antibodies are increasingly being used as diagnostic and therapeutic tools. As only the genomic organization of the micro encoding gene was previously known, we analysed the chicken immunoglobulin Y (IgY) and IgA (upsilon and alpha chain) immunoglobulin heavy-chain constant region (IGHC) genes and the organization of the chicken IGHC locus. The alpha gene is encoded by four separate exons, whereas, surprisingly, there is no intervening DNA sequence between the CH1 and CH2 domains of the IgY heavy chain, which is thus encoded by three exons separated by two introns. DNA sequence analysis shows that the exon boundaries of the chicken IGHC genes are not consistent with published domain borders. Furthermore, differences in DNA sequence confirm the existence of IgY, IgA and IgM allotypes in chickens. Finally, our results show that the IGHC genes of chicken (IgY, IgA and IgM) are all colocated on chromosome E18C15W15, where the alpha gene is located upstream of the upsilon gene in an inverted transcriptional orientation. The distances between the mu and alpha genes and the alpha and upsilon genes are about 18 and 15 kilobases, respectively, and thus, the size of the whole chicken IGHC locus is approximately 67 kilobases. PMID- 11106939 TI - Immunoglobulin A cell distribution in the human small intestine: phenotypic and functional characteristics. AB - We compared B-cell phenotypes in Peyer's patches and solitary lymphoid follicles (organized gut-associated lymphoid tissue, GALT) with those in jejunal or ileal lamina propria. In situ, immunostaining showed that small B cells of naive [surface immunoglobulin D-positive (sIgD+) CD27-] and memory (sIgD+/- CD27+) phenotypes occurred almost exclusively in GALT, whereas the lamina propria contained only scattered sIgA+ CD27+ memory cells. In contrast, B-cell blasts and plasma cells negative for CD20 and often also for CD19 but with strong expression of CD38, CD27 and cytoplasmic IgA (cIgA), dominated in the lamina propria but were scarce in GALT. By flow cytometry, the proportion of dispersed CD19+ B lymphocytes varied from 4 to 42% among jejunal mucosal samples; between 5 and 50% of these were sIgD+, suggesting a variable contamination with GALT cells. B-cell blasts and plasma cells, identified by their large size and strong expression of CD38, were regularly found (25-35% of the total mononuclear cell population). Distinction between B-cell blasts and mature plasma cells was made by the presence or absence of human leucocyte antigen (HLA) class II molecules, CD45RA, CD19 and surface immunoglobulin. No CD19+ B cells outside GALT expressed CD5, but a very small portion of the lamina propria B-cell blasts were positive for CD28. Dispersed sIgA+ lamina propria cells expressed low levels of CD40, proliferated on CD40 ligation and constitutively secreted IgA in vitro. We concluded that the lamina propria B-cell compartment consists mainly of B-cell blasts and plasma cells but also has scattered, small sIgA+ cells that can proliferate in response to CD40 ligation and may therefore function as local memory cells for recall antigens. PMID- 11106940 TI - Natural autoantibodies against heat-shock proteins hsp70 and gp96: implications for immunotherapy using heat-shock proteins. AB - Immunization of mice with cognate cancer-derived heat-shock protein (hsp) preparations leads to protection from cancer growth. As hsp used for vaccination or therapy are derived from autologous cancers, questions of pathological autoimmunity are of immense significance for the ongoing translation of this approach to therapy of human cancer. Employing the sera of normal adult mice as the first antibody, highly sensitive immunoblotting revealed the presence of anti hsp natural autoantibodies in healthy animals. Natural autoantibodies of the immunoglobulin D (IgD) isotype bind to gp96, whereas hsp70 was recognized by IgD and IgM autoantibodies. Neither hsp was recognized by the IgA, IgE or IgG immunoglobulins contained in the serum. The antigen-antibody recognition was titratable and dependent on the integrity of the IgD molecule. Sera from only a subset of the animals tested were found to be positive for autoantibodies against gp96 and hsp70, and individual and strain-specific variations were detected. Injection of gp96 into healthy mice did not show sustained or consistent anti gp96 IgD antibody response, class switching, toxicity or pathological autoimmunity. IgD autoantibodies against gp96 and hsp70 were also not detected in the autoimmune lpr mice. These observations show the existence of a measured and tightly regulated natural immune response to hsp. PMID- 11106941 TI - Specificity and binding kinetics of murine lupus anti-DNA monoclonal antibodies implicate different stimuli for their production. AB - The origin and relative biological importance of the many different DNA-reactive antibodies that appear in systemic lupus erythematosus are not well understood. A detailed analysis of their fine specificity and binding characteristics with DNA is a necessary step in understanding their biology. We have examined here two monoclonal antibodies (mAb) IV-228 and V-88 that are, respectively, characteristic of antibodies, which bind exclusively to single-stranded (ss) DNA and to both double-stranded (ds) DNA and ssDNA. By surface plasmon resonance (SPR) on BIAcore, we characterized the kinetics of binding of each antibody to synthetic ss and ds oligonucleotides. Antibody V-88 and IV-228 showed different patterns of reactivity for both ss and ds oligonucleotides, characterized by distinctly different kinetic parameters. Analysis of their binding kinetics indicates the importance of base composition in defining DNA epitopes, and shows that some epitopes, such as that recognized by mAb V-88, are expressed on dsDNA and ssDNA, whereas others, as recognized by IV-228, are not. The base preferences of V-88 for ds GC-rich structures over AT-rich, and of IV-228 for ss T-rich structures, also reveal distinct differences between these antibodies. We conclude that the different binding properties of the antibodies will relate to their biological activities. The base preferences of the antibodies suggest that they might be induced by different immunological stimuli, such as those that could be provided by the various DNA fragments and structures released during programmed cell death. PMID- 11106942 TI - T helper 1-type cytokine transcription in peripheral blood mononuclear cells of pseudorabies virus (Suid herpesvirus 1)-primed swine indicates efficient immunization. AB - The induction of porcine cytokines, which are believed to be important for the regulation of T helper (Th)1- and Th2-specific immune responses of pigs, was analysed after in vitro restimulation with a herpesvirus, Suid herpes 1 (pseudorabies virus [PRV]), in peripheral blood mononuclear cells (PBMC). To this end, quantitative, competitive reverse transcription-polymerase chain reaction (RT-qcPCR) was established using constructed heterologous DNA MIMICS, which contain cytokine- or glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-specific primer-binding sites. This is a simple method that allows reliable determination of the differing regulation of cytokine mRNAs specific for porcine interleukin (IL)-2, -4 and -10, interferon gamma (IFN-gamma) and the housekeeping gene, GAPDH, as an endogenous control. PBMC derived from naive (innate response) and PRV-primed (memory response) outbred swine were analysed comparatively. The results demonstrated that restimulation with PRV significantly enhanced the transcription of Th1-type cytokines (IL-2 and IFN-gamma) but not of Th2-type cytokines (IL-4 and IL-10). This virus-specific cytokine response was only found with PBMC from swine protected against lethal PRV challenge infection, but not with naive PBMC or with PBMC from pigs immunized with plasmid DNA encoding PRV glycoprotein gC. Notably, PBMC derived from immune and naive pigs constitutively produced relatively high amounts of IL-10-specific mRNA, exceeding that of GAPDH mRNA, independently of the addition of viral antigen or the mitogen concanavalin A (Con A). The results of this work should help to provide a better understanding of the effector cell/cytokine network response to infection with, or vaccination against, PRV. Additionally, the simple, reliable and sensitive RT-qcPCR, when used to determine the porcine cytokine pattern, might be of prognostic value for the induction of protective immunity. PMID- 11106943 TI - Recombinant adenovirus vectors expressing interleukin-5 and -6 specifically enhance mucosal immunoglobulin A responses in the lung. AB - In this study, we have examined the in vivo effects of interleukin-5 (IL-5) and IL-6 over-expression on systemic and mucosal immune responses using recombinant human type 5 adenoviruses capable of expressing these cytokines upon infection. A recombinant adenovirus containing the murine IL-5 gene within the E3 region was constructed and found to express high levels of IL-5 protein both in vitro and in vivo. Intranasal inoculation of mice with this vector or a vector expressing murine IL-6 increased adenovirus-specific immunoglobulin A (IgA) titres in lung lavage fluid threefold compared with those elicited by control virus. The simultaneous expression of both cytokines by co-inoculation altered the kinetics of the mucosal anti-adenovirus IgA response and resulted in a more than additive increase in antibody titres. The co-expression effect on IgA synthesis was not due to an increase in numbers of antigen-specific resident lung tissue lymphocytes. When mucosal IgG responses were examined, IL-6 expression had the largest impact on anti-adenovirus levels, whereas co-expression produced an intermediate response. Systemic immune responses were also affected by IL-6 expression as a twofold increase in serum IgG anti-adenovirus titres was observed after a secondary challenge with wild-type adenovirus. These results demonstrate a relevant role for IL-5 and IL-6 in the development of mucosal immune responses in vivo and suggest that the incorporation of either IL-5 and/or IL-6 into recombinant adenovirus vectors may be a useful tool in the development of mucosal vaccines. PMID- 11106944 TI - Adherent dendritic cells expressing high levels of interleukin-10 and low levels of interleukin-12 induce antigen-specific tolerance to experimental autoimmune encephalomyelitis. AB - We have previously shown that tolerance can be induced against acute experimental autoimmune encephalomyelitis (EAE) in Lewis rats by bone marrow-derived dendritic cells (DC) that have been pulsed in vitro with encephalitogenic myelin basic protein peptide 68-86 (MBP 68-86), and injected subcutaneously into healthy rats prior to immunization with MBP 68-86 plus complete Freund's adjuvant. To elucidate better the properties of tolerogenic DC, we here compared plastic adherent DC with floating, non-adherent DC, which were cultured for 7 days in the presence of granulocyte-macrophage colony-stimulating factor plus interleukin-4 (IL-4). Adherent DC expressed high levels of IL-10 mRNA and protein, and low levels of IL-12 mRNA and showed high expression of CD54 compared with floating DC. Proliferation, nitrite concentration and capacity for antigen presentation were lower in adherent DC than in floating DC. There were no differences between adherent and floating DC regarding expression of CD11c, OX62, major histocompatibility complex class II, CD80, or CD86. Most importantly, we observed that adherent DC induced tolerance to EAE in vivo when injected subcutaneously into Lewis rats prior to immunization, while floating DC did not. Adherent DC mediated tolerance to EAE was associated with augmented proliferation, nitric oxide production and frequency of apoptotic cells as well as with up-regulation of transforming growth factor-beta (TGF-beta) -expressing cells in T-cell areas of lymph nodes. Tolerance induction by adherent DC seems to be related to a nitric oxide-apoptosis pathway and to up-regulation of TGF-beta-expressing cells. PMID- 11106945 TI - Induction of specific transplantation immunity by oral immunization with allogeneic cells. AB - Oral administration of antigen has been shown to be effective for both positive and negative modulation of immune responses. In the present study we characterized changes in the reactivity of the immune system after oral immunization with allogeneic spleen cells. Mice were orally immunized for 10 consecutive days with fresh allogeneic spleen cells, and the phenotype, proliferative response, cytotoxic activity and cytokine production profile of recipient spleen cells were assessed 1 or 7 days after the last immunization dose. Although no significant changes in the proportion of CD4+, CD8+ or CD25+ cells were observed in the spleen of orally immunized mice, significant activation of alloreactivity in spleen cells was found. Cells from orally immunized mice exhibited enhanced proliferation and cytotoxic activity after stimulation with specific allogeneic cells in vitro, and produced considerably higher concentrations of interferon-gamma (IFN-gamma) and significantly less interleukin (IL)-4 than did cells from control mice. The production of IL-2 was essentially unchanged and that of IL-10 was only slightly increased. The systemic allosensitization induced by oral immunization was demonstrated in vivo by increased resistance to the growth of allogeneic tumours induced by subcutaneous inoculation of high doses of tumour cells. In addition, orthotopic corneal allografts in orally immunized recipients were rejected more rapidly (in a second set manner) than in control, untreated recipients. These data show that oral immunization with allogeneic cells modulates individual components of the immune response and that specific transplantation immunity, rather than tolerance, is induced in the treated recipients. PMID- 11106946 TI - Hydrogen peroxide augments eosinophil adhesion via beta2 integrin. AB - During eosinophil (EOS) accumulation at sites of allergic inflammation, an initial step is the binding of EOS to adhesion molecules expressed on vascular endothelial cells (EC). We have previously observed that adhesion of peripheral blood EOS to recombinant human vascular cell adhesion molecule-1 (rh-VCAM-1) stimulates the respiratory burst of EOS. Although the biological consequence of this activation remains to be elucidated, reactive oxygen species such as hydrogen peroxide (H2O2) may modify the adhesive property of EOS. In the present study, we examined whether H2O2 modifies the adhesive property of EOS. EOS were isolated from the peripheral blood of healthy subjects. Adhesion of the EOS to paraformaldehyde-fixed human umbilical vein EC (HUVEC), stimulated or not stimulated with tumour necrosis factor-alpha (TNF-alpha; 100 pM for 24 hr), was examined in the presence or absence of H2O2. H2O2 significantly enhanced adhesion of EOS to both resting and TNF-alpha-stimulated fixed HUVEC (P < 0.01, respectively). Such enhancing effects were inhibited by anti-beta2 integrin antibody or anti-CD11b antibody, but not by anti-CD11a or anti-alpha4 integrin antibody. H2O2 also enhanced EOS adhesion to rh-intracellular cell adhesion molecule-1 (ICAM-1) but not to rh-VCAM-1. Finally, H2O2 enhanced the expression of both CD11b and CD18 on EOS. These results indicate that H2O2 directly augments the adhesive property of EOS through beta2 integrin. PMID- 11106947 TI - Interleukin-4 and RANTES expression in maturing eosinophils derived from human cord blood CD34+ progenitors. AB - Eosinophils elaborate a number of proinflammatory mediators, including immunoregulatory cytokines and chemokines. Interleukin (IL)-4 and RANTES are important cytokines that have previously been shown to be expressed by mature eosinophils. We hypothesized that de novo synthesis of IL-4 and RANTES occurs in nascent eosinophils, leading to storage of newly produced proteins in crystalloid granule-like structures. Cytokine mRNA and protein expression were examined in cultured eosinophil colonies, which were derived from purified cord blood CD34+ cells and generated in semisolid media (methylcellulose) in the presence of recombinant human (rh)IL-3 and rhIL-5. Cytokine mRNA profiles were analysed by the reverse transcription-polymerase chain reaction (RT-PCR) to determine transcription of IL-4 and RANTES in cells on days 0, 7, 14, 21 and 28 of culture. The expression of translated cytokine products and granule major basic protein (MBP) was confirmed, from day 23 onwards, for colonies cultured in semisolid media, by immunofluorescent labelling and confocal laser-scanning microscopy (CLSM). We found that mRNA sequences encoding IL-4 and RANTES were expressed in freshly prepared, non-differentiated CD34+ cells. Furthermore, RANTES mRNA localized to carbol chromotrope 2R-positive colony cells, as assessed using in situ RT-PCR on day 21 of culture in semisolid media, and was found to gradually decrease (relative to beta2-microglobulin) in rhIL-3- and rhIL-5-treated colony cells (comprising > 90% eosinophil-like cells) up to day 28. Immunoreactivity for IL-4 and RANTES co-localized with MBP in maturing colony eosinophils on day 23 of culture in semisolid media, as judged by CLSM. These results suggest that synthesis and storage of immunoregulatory cytokines, essential for processes associated with adaptive immunity, occurs in nascent eosinophils during their growth and differentiation. PMID- 11106949 TI - Multiphoton microscopy in life sciences. AB - Near infrared (NIR) multiphoton microscopy is becoming a novel optical tool of choice for fluorescence imaging with high spatial and temporal resolution, diagnostics, photochemistry and nanoprocessing within living cells and tissues. Three-dimensional fluorescence imaging based on non-resonant two-photon or three photon fluorophor excitation requires light intensities in the range of MW cm(-2) to GW cm(-2), which can be derived by diffraction limited focusing of continuous wave and pulsed NIR laser radiation. NIR lasers can be employed as the excitation source for multifluorophor multiphoton excitation and hence multicolour imaging. In combination with fluorescence in situ hybridization (FISH), this novel approach can be used for multi-gene detection (multiphoton multicolour FISH). Owing to the high NIR penetration depth, non-invasive optical biopsies can be obtained from patients and ex vivo tissue by morphological and functional fluorescence imaging of endogenous fluorophores such as NAD(P)H, flavin, lipofuscin, porphyrins, collagen and elastin. Recent botanical applications of multiphoton microscopy include depth-resolved imaging of pigments (chlorophyll) and green fluorescent proteins as well as non-invasive fluorophore loading into single living plant cells. Non-destructive fluorescence imaging with multiphoton microscopes is limited to an optical window. Above certain intensities, multiphoton laser microscopy leads to impaired cellular reproduction, formation of giant cells, oxidative stress and apoptosis-like cell death. Major intracellular targets of photodamage in animal cells are mitochondria as well as the Golgi apparatus. The damage is most likely based on a two-photon excitation process rather than a one-photon or three-photon event. Picosecond and femtosecond laser microscopes therefore provide approximately the same safe relative optical window for two-photon vital cell studies. In labelled cells, additional phototoxic effects may occur via photodynamic action. This has been demonstrated for aminolevulinic acid-induced protoporphyrin IX and other porphyrin sensitizers in cells. When the light intensity in NIR microscopes is increased to TW cm(-2) levels, highly localized optical breakdown and plasma formation do occur. These femtosecond NIR laser microscopes can also be used as novel ultraprecise nanosurgical tools with cut sizes between 100 nm and 300 nm. Using the versatile nanoscalpel, intracellular dissection of chromosomes within living cells can be performed without perturbing the outer cell membrane. Moreover, cells remain alive. Non-invasive NIR laser surgery within a living cell or within an organelle is therefore possible. PMID- 11106950 TI - Adaptive aberration correction in a two-photon microscope AB - We demonstrate aberration correction in two-photon microscopy. Specimen-induced aberrations were measured with a modal wavefront sensor, implemented using a ferro-electric liquid crystal spatial light modulator (FLCSLM). Wavefront correction was performed using the same FLCSLM. Axial scanned (xz) images of fluorescently labelled polystyrene beads using an oil immersion lens show restored sectioning ability at a depth of 28 &mgr;m in an aqueous specimen. PMID- 11106948 TI - Control of chemokine production at the blood-retina barrier. AB - Chemokine production at the blood-retina barrier probably plays a critical role in determining the influx of tissue-damaging cells from the circulation into the retina during inflammation. The blood-retina barrier comprises the retinal microvascular endothelium and the retinal pigment epithelium. Chemokine expression and production by human retinal microvascular endothelial cells (REC) have never been reported previously, so we examined the in vitro expression and production of monocyte chemoattractant protein-1 (MCP-1), regulated on activation of normal T-cell expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, interleukin (IL)-8, epithelial cell-derived neutrophil activating protein-78 (ENA-78) and growth related oncogene alpha (GROalpha) in these cells, both unstimulated and stimulated by cytokines likely to be present during the evolution of an inflammatory response. We compared this to expression and production of these chemokines in vitro in human retinal pigment epithelial cells (RPE). MCP-1 was expressed and produced constitutively by REC but all the chemokines were produced in greater amounts upon stimulation with the proinflammatory cytokines IL-1beta and tumour necrosis factor-alpha (TNF-alpha). MCP-1 and IL-8 were produced at much higher levels than the other chemokines tested. MIP-1alpha and MIP-1beta were present only at low levels, even after stimulation with IL-1beta and TNF-alpha. Cytokines with greater anti-inflammatory activity, such as IL-4, IL-10, IL-13, transforming growth factor-beta (TGF-beta) and IL-6, had little effect on chemokine production either by REC alone or after stimulation with IL-1beta and TNF-alpha. RPE, although a very different cell type, showed a similar pattern of expression and production of chemokines, indicating the site-specific nature of chemokine production. Chemokine production by REC and RPE is probably significant in selective leucocyte recruitment during the development of inflammation in the retina. PMID- 11106951 TI - Adhesion artefacts in atomic force microscopy imaging AB - Artefacts that affect contrast and arise from adhesion forces in atomic force microscopy images of aramid fibres (both fresh and plasma-treated) are investigated. It is demonstrated that these stem not only from variations in the chemical composition of the surface but also from certain topographical features (which may appear hidden or enhanced in the images), resulting in changes in the lateral forces that are detected by the cantilever and are comparable to the vertical forces. It is also shown that both types of contribution to the forces can be uncoupled to yield images free from these artefacts, thus allowing more accurate quantitative measurements. These artefactual effects are also generally applicable to many other materials. PMID- 11106953 TI - New controlled environment vitrification system for cryo-transmission electron microscopy: design and application to surfactant solutions. AB - A newly designed controlled environment vitrification system for cryo transmission electron microscopy of vitrified samples (cryo TEM) is presented. It significantly reduces the lag-time from blotting to vitrification by means of an optimized blotting procedure. Furthermore, a modified transfer system allows rapid transfer of the vitrified sample into the microscope and reduces the equilibration time. The practicable throughput of samples is thereby markedly increased. This new equipment is used to determine the overall size of individual polymer-like mixed lecithin-bile salt micelles, which very critically depends on the composition of the sample. The quantitative agreement with the results from independent light scattering experiments on the same sample supports the validity and performance of the presented sample preparation and handling procedures. In addition, these experiments demonstrate that cryo TEM experiments not only provide direct structural information, but furthermore permit a quantitative determination of micellar properties. PMID- 11106952 TI - Blind deconvolution of 3D transmitted light brightfield micrographs. AB - The blind deconvolution algorithm for 3D transmitted light brightfield (TLB) microscopy, published previously [Holmes et al. Handbook of Biological Confocal Microscopy (1995)], is summarized with example images. The main emphasis of this paper is to discuss more thoroughly the importance and usefulness of this method and to provide more detailed evidence, some being quantitative, of its necessity. Samples of horseradish peroxidase (HRP)-stained pyramidal neurones were prepared and evaluated for the ability to see fine structures clearly, including the dendrites and spines. It is demonstrated that the appearance of fine spine structure, and means of identifying spine categories, is made possible by using blind deconvolution. A comparison of images of the same sample from reflected light confocal microscopy, which is the conventional light microscopic way of viewing the 3D structure of these HRP-stained samples, shows that the blind deconvolution method is far superior for clearly showing the structure with less distortion and better resolution of the spines. The main significance of this research is that it is now possible to obtain clear images of 3D structure by light microscopy of absorbing stains. This is important because the TLB microscope is probably the most widely used modality in the life-science laboratory, yet, until now, there has been no reliable means for it to provide visualization of 3D structure clearly. The main importance of the blind deconvolution approach is that it obviates the need to measure the point spread function of the optical system, so that it now becomes realistic to provide a 3D light microscopic deconvolution method that can be pervasively used by microscopists. PMID- 11106954 TI - Detection of shape changes in biological features. AB - The question of analysing shape changes over time, such as during growth and during the progress of disease, is an important fundamental issue for many applications. Recent mathematical developments in the understanding of the detailed structure of shape spaces have made possible the quantitative study of shape variation. In this paper, we combine the classical multidimensional scaling technique with knowledge of the geometry of shape spaces to examine the role played by the geodesics in shape spaces. We present some promising early results answering questions about shape changes over time. PMID- 11106955 TI - Non-uniform systematic sampling in stereology. AB - Non-uniform systematic sampling designs in stereology are studied. Various methods of constructing non-uniform systematic sampling points from prior knowledge of the measurement function are presented. As an example, we consider area estimation from lengths of linear intercepts. The efficiency of two area estimators, based on non-uniform sampling of parallel lines, is compared to that of the classical 2D Cavalieri estimator, based on uniform sampling, in a sample of planar profiles from transverse sections of 41 small myelinated axons. The comparison is based on simulations. It is concluded that for profiles of this type one of the non-uniform sampling schemes is more efficient than the traditional uniform sampling scheme. Other examples where non-uniform systematic sampling may be used are in area estimation from lines emanating from a fixed point, area estimation from concentric circles or spirals and curve length estimation from sweeping lines. It is shown that proportional-to-size sampling is a special case of non-uniform systematic sampling. Finally, the effect of noise in the observations is discussed. PMID- 11106956 TI - Improved estimation of the pair correlation function of random sets. AB - The texture of binary spatial structures can be characterized by second-order methods of spatial statistics. The pair correlation function, which describes the structure in terms of spatial correlation as a function of distance, is of central importance in this context. Conventionally, the pair correlation function of stationary and isotropic random sets is estimated as the ratio of the covariance to the square of volume fraction of the phase of interest. In the present paper, an improved estimator of the pair correlation function is presented, where the covariance is divided by the square of a distance-adapted estimator of volume fraction. The new estimator is explained mathematically and applied to simulated images of the Boolean model and to microscopic images from neoplastic and non-neoplastic human glandular tissues. It leads to a considerable reduction of bias and variance of estimated pair correlation functions, in particular for large distances. PMID- 11106957 TI - Application of a laser scalpel to sectioning fragile tissues prior to optical sectioning microscopy and 3D reconstruction. AB - A method is described for the cutting of fragile material with a laser scalpel which minimizes damage to friable materials, making the interior of structures accessible for optical sectioning microscopy or for high resolution X-ray microtomography followed by 3D reconstruction. PMID- 11106958 TI - 'Analysis of spherical aberration of a water immersion objective: application to specimens with refractive index 1.33-1.40'. PMID- 11106959 TI - Penetration pathways of fluorescent dyes in human hair fibres investigated by scanning near-field optical microscopy. AB - Thin cross-sections of human hairs were investigated by scanning near-field optical microscopy (SNOM) and confocal laser scanning microscopy (CLSM) after penetration of a fluorescent dye. The same samples were measured with both techniques to compare the observed structures. The images obtained from the two methods show nearly identical structures representing pathways of the dye molecules in hairs. The SNOM images provide a higher resolution than the CLSM images. Therefore, SNOM is believed to be a suitable method for investigations at a resolution of 100 nm on penetration pathways of fluorescent dyes such as the cell membrane complex pathway in cross-sections of hairs. PMID- 11106960 TI - Off-axis electron holography of patterned magnetic nanostructures. AB - Magnetization reversal processes in lithographically patterned magnetic elements that have lateral dimensions of 70-500 nm, thicknesses of 3-30 nm and a wide range of shapes and layer sequences have been followed in situ using off-axis electron holography in the transmission electron microscope. This technique allows domain structures within individual elements and the magnetic interactions between them to be quantified at close to the nanometre scale. The behaviour of 30 nm-thick Co elements was compared with that of 10 nm-thick Ni and Co elements, as well as with Co/Au/Ni trilayers. The hysteresis loops of individual elements were determined directly from the measured holographic phase images. The reproducibility of an element's domain structure in successive cycles was found to be affected by the out-of-plane component of the applied magnetic field and by the exact details of its initial magnetic state. Close proximity to adjacent elements led to strong intercell coupling, and remanent states with the in-plane magnetic field removed included domain structures such as solenoidal (vortex) states that were never observed during hysteresis cycling. Narrow rectangular bars reversed without the formation of end domains, whereas closely separated magnetic layers within individual elements were observed to couple to each other during field reversal. PMID- 11106961 TI - An efficient algorithm for measurement and correction of chromatic aberrations in fluorescence microscopy. AB - Even the best optical microscopes available on the market exhibit chromatic aberrations to some extent. In some types of study, chromatic aberrations of current optics cannot be neglected and a software correction is highly desirable. This paper describes a novel method of chromatic aberration measurement and software correction using sub-resolution bead imaging and computer image analysis. The method is quick, precise and enables the determination of both longitudinal and lateral chromatic aberrations. Correction function can be computed in about half an hour, including image acquisition. Using this approach, chromatic aberrations can be reduced to 10-20 nm laterally and 10-60 nm axially depending on the type of optical set-up. The method is especially suitable for fluorescence microscopy, where a limited number of wavelengths are observed. PMID- 11106962 TI - A comparison of optical geometries for combined flash photolysis and total internal reflection fluorescence microscopy. AB - Total internal reflection fluorescence (TIRF) microscopy, used in conjunction with flash photolysis, provides a useful way of investigating the kinetics of macromolecular interactions. We compare different TIRF optical geometries to establish an optimal combination. Excitation light was introduced via four different arrangements: (1) a prism positioned on the microscope optical axis, (2) an offset prism with propagation through a silica slide trans to the objective lens, (3) an offset prism with propagation through a silica coverslip cis to a water-immersion objective lens and (4) a prismless arrangement using a high NA oil-immersion objective lens. Photolysis was achieved using a Xe flash lamp and a customised silica condenser lens. Single myosin molecules labelled with a Cy3 fluorophore were used as a test sample. Although the offset trans prism gave the best signal-to-background ratio, a customised thin rhombic prism incorporated, on axis, into the flash condenser assembly was almost as good and was more practical for scanning multiple fields. An oil-immersion lens gave the brightest image for sample depths < 30 micrometer but above this limit, a water immersion lens was better. The prismless arrangement may offer advantages in other situations but it is important to check the actual numerical aperture of the objective lens. PMID- 11106963 TI - The continuum normalization method for quantification of X-ray spectra in biological microanalysis. 1. Generalized bremsstrahlung production cross-sections and analysis using standards. AB - The thin self-supporting biological specimens used for quantitative X-ray microanalysis are problematical because the sections are most unlikely to be uniform in thickness or density, so the intensities of the characteristic lines alone are not a good measure of composition. The method developed to overcome these problems was introduced by T. A. Hall in 1971 and uses the bremsstrahlung or continuum intensity recorded in the X-ray spectrum to normalize each characteristic line, and hence is frequently referred to as the continuum normalization (CN) procedure. Reformulating the CN method of quantification in terms of generalized cross-sections and calculating more accurate values of bremsstrahlung production using a formula allows us a better understanding of the options open to the analyst of biological thin sections by which the errors in the measurement may be reduced. If one chooses to use the original Hall (1971) method using Kramers cross-sections, the window measuring the continuum for normalization should be set in the 4-7 keV region for typical scanning electron microscope and microprobe beam energies, 20-40 kV, and above 10 keV for transmission electron microscope energies of 80 kV and above. Although it is clear that peak counts must not contribute to the white count, the window should be as wide as possible to reduce statistical errors. PMID- 11106964 TI - Development of an X-ray photoemission electron microscopy system with multi probes, and its application to surface imaging at static and dynamic states. AB - We have developed a new X-ray photoemission electron microscopy system combined with low energy electron microscopy, photoemission electron microscopy, mirror electron microscopy (MEM), secondary electron emission microscopy (SEEM) and Auger electron emission microscopy, which provides multi-angle information on the distribution and change of element, chemical state, structure, etc. at solid surfaces under the working conditions such as high temperature and gas atmosphere. The performance of each microscopical method was examined and typical images are presented. The dynamic behaviour of fabricated surfaces has been imaged in real time by SEEM and MEM. PMID- 11106965 TI - Image analysis of insulation mineral fibres. AB - We present two methods for measuring the diameter and length of man-made vitreous fibres based on the automated image analysis of scanning electron microscopy images. The fibres we want to measure are used in materials such as glass wool, which in turn are used for thermal and acoustic insulation. The measurement of the diameters and lengths of these fibres is used by the glass wool industry for quality control purposes. To obtain reliable quality estimators, the measurement of several hundred images is necessary. These measurements are usually obtained manually by operators. Manual measurements, although reliable when performed by skilled operators, are slow due to the need for the operators to rest often to retain their ability to spot faint fibres on noisy backgrounds. Moreover, the task of measuring thousands of fibres every day, even with the help of semi automated image analysis systems, is dull and repetitive. The need for an automated procedure which could replace manual measurements is quite real. For each of the two methods that we propose to accomplish this task, we present the sample preparation, the microscope setting and the image analysis algorithms used for the segmentation of the fibres and for their measurement. We also show how a statistical analysis of the results can alleviate most measurement biases, and how we can estimate the true distribution of fibre lengths by diameter class by measuring only the lengths of the fibres visible in the field of view. PMID- 11106966 TI - A simplified application of systematic field sampling and low-cost video recording set-up for viewing disector pairs - exemplified in the rat cochlear nucleus. AB - A description of a simple and efficient way of systematic field sampling along with a low-cost set-up for simultaneous viewing of physical disector pairs at the light microscopic level is presented. So far the use of a programmable motorized stage for the former purpose and two projection microscopes for the latter have proved to be the most efficient and fastest solutions, but the relatively high expense of purchasing such equipment constitutes a major impediment to the widespread use of such stereological methods. We describe practical applications of two new cost-effective alternative approaches derived from existing ones. These approaches are used in estimating the total number of neurones in the cochlear nucleus of the rat and involve the systematic selection of section fields at a magnification significantly lower than the magnification used for counting and a video recording set-up for comparing section pairs. Apart from providing a cheap way for the implementation of the physical disector, the main feature of the application, which may also encourage its use, is that it provides an efficient and simple design for systematic area sampling without requiring any specialized equipment. The same is true for optical disector designs, where the procedure is simplified even further. PMID- 11106967 TI - Efficient estimation of number density in opaque material microstructures: the large-area disector. AB - Unbiased and efficient estimation of number density of features in opaque material microstructures has been quite difficult. In this contribution a montage based efficient serial sectioning technique is presented as a practical solution for efficient estimation of number density in such microstructures. The new technique utilizes a combination of digital image analysis and unbiased disector sampling procedures. PMID- 11106968 TI - Monitoring age-related changes of collagen content and vascularity in ganglia using unbiased stereological methods. AB - We describe for the first time application of unbiased stereological techniques to estimate total volume and volume fractions of interest in individual dorsal root ganglia (DRG). Volume estimates were obtained using a two-stage sampling design. Sections were systematically sampled following a random start, from DRG which were embedded in methacrylate and exhaustively sectioned. We further examined the efficiency of point counting irregular volume fractions housed in a regular reference volume. We found that the precision of volume estimates was relatively unaffected by exhaustive sampling, and that the magnitude of error was, in large part, determined by object shape. PMID- 11106969 TI - Neuroendocrinology briefings 12: The maternal brain. AB - The mother's brain is prepared by the hormones of pregnancy to show the strong maternal feelings that ensure the newborn is cared for. These hormones induce a cascade of changes in the brain, reducing stress reactions, evoking maternal behaviour and preparing the neuroendocrine circuits that drive the birth process and ensure that the suckling infant gets milk. The nerve cells that make oxytocin are involved in all of these aspects of motherhood, and details are emerging of how their performance is adapted by pregnancy. PMID- 11106970 TI - Variations in maternal behaviour are associated with differences in oxytocin receptor levels in the rat. AB - Female Long-Evans rats exhibit stable individual differences in maternal behaviours such as pup licking/grooming and arched-back nursing posture (LG-ABN). These variations in maternal behaviour are accompanied by differences in lactation-induced increases in oxytocin receptor levels in brain regions known to mediate the expression of maternal care in this species (i.e. the bed nucleus of the stria terminalis, the medial preoptic area and the lateral septum). Oxytocin receptor levels in the central nucleus of the amygdala were significantly higher in high compared to low LG-ABN females regardless of reproductive status. These findings suggest that individual differences in maternal behaviour may be directly related to variations in oxytocin receptor expression. PMID- 11106971 TI - Chronic iodine deprivation attenuates stress-induced and diurnal variation in corticosterone secretion in female Wistar rats. AB - Many millions of people throughout the world are at risk of developing iodine deficiency-associated disorders. The underlying effects of iodine deficiency on neuroendocrine function are poorly defined. We have studied stress-induced and diurnal variation in corticosterone secretion in female rats rendered chronically hypothyroid by feeding them an iodine-free diet for 6 months. Corticosterone secretory responses in iodine deficient animals were compared to those seen in animals rendered hypothyroid with propylthiouracil and untreated controls. By using a well-validated, automated blood sampling system to collect small samples of blood over the complete daily cycle in unrestrained animals, we have demonstrated for the first time that the normal diurnal rhythm of corticosterone secretion is lost in chronic iodine deficiency and that the corticosterone secretory response to the psychological stress of 10 min exposure to white noise is attenuated. Despite restoration of circulating triiodothyronine and thyrotropin releasing hormone- and thyroid stimulating hormone beta-transcript prevalence in the hypothalamus and pituitary, respectively, 1 month after restoration of normal iodine-containing diet both the diurnal variation in corticosterone levels and the corticosterone secretory response to the noise stress remained reduced in amplitude compared to control animals. Thus, chronic hypothyroidism induced by iodine deficiency significantly attenuates hypothalamo pituitary-adrenal axis activity, an effect that persists after functional recovery of the thyroid axis. PMID- 11106972 TI - Substance P downregulates basal and gonadotropin-releasing hormone-induced gonadotropin in vitro secretion by pituitary gland of crested newt, Triturus carnifex. AB - The possible role of Substance P (SP) was studied in the modulation of basal and gonadotopin-releasing hormone (GnRH)-induced gonadotropin secretion in the urodele crested newt, Triturus carnifex. During prereproduction, reproduction (noncourtship and courtship), refractory, recovery and aestivation, male and female pituitaries were incubated with medium-alone, GnRH, SP, GnRH receptor antagonist (antide), and SP receptor antagonist (L-703606). Since antisera raised against gonadotropins are not available for this species, we measured these hormones indirectly through their effects on the secretion of testicular androgens and ovarian progesterone from gonads superfused with the preincubated pituitaries. Pituitaries of both sexes preincubated with medium-alone, GnRH, GnRH plus L-703606, and GnRH plus SP plus L-703606 increased steroid secretion during prereproduction, noncourtship, courtship, and recovery; the increase induced by the pituitaries incubated with medium-alone was lower during prereproduction, noncourtship, and recovery. Pituitaries preincubated with SP, GnRH plus SP, GnRH plus SP plus antide, and SP plus antide did not change basal steroid secretion in any of the reproductive phases considered. Antide, L-703606, GnRH plus antide, GnRH plus SP plus antide plus L-703606, SP plus L-703606, and antide plus L 703606 experimental groups showed the same results as those with medium-alone. These results suggest that SP downregulates gonadotropin release in both Triturus carnifex sexes. In addition, an antagonist role, through receptor-independent mechanisms, exists between GnRH (upregulation) and SP (downregulation) in the modulation of pituitary. PMID- 11106973 TI - Antagonistic oxytocin/alpha2-adrenoreceptor interactions in the nucleus tractus solitarii: relevance for central cardiovascular control. AB - The modulation of the central cardiovascular effects of alpha2-adrenoceptor activation by oxytocin in the nucleus tractus solitarii has been evaluated by cardiovascular analysis and by quantitative receptor autoradiography. Microinjections in the nucleus tractus solitarii of a threshold dose of oxytocin effectively and significantly counteracted the vasodepressor and bradycardic actions of an ED50 dose of the alpha2-adrenoceptor agonist clonidine. The coinjection of a threshold dose of oxytocin with a threshold dose of clonidine did not produce any changes in the mean arterial pressure but a tachycardic response was observed. Receptor autoradiographical experiments showed that oxytocin (3 nM) significantly increased the Kd and Bmax values of [3H]p aminoclonidine binding sites in the nucleus tractus solitarii compatible with a possible antagonistic interaction with the alpha2-adrenoceptors, and this effect was blocked by the presence of the specific oxytocin receptor antagonist 1 deamino-2-D-Tyr-(OEt)-4-Thr-8-Orn-oxytocin. These findings suggest the existence of an antagonistic oxytocin/alpha2-adrenoceptor interaction in nucleus tractus solitarii that may be of relevance for the demonstrated modulation of alpha2 adrenoceptor induced cardiovascular responses by oxytocin. PMID- 11106974 TI - Effects of orexins on the hypothalamic-pituitary-adrenal system. AB - The effects of the recently identified neuropeptides orexin-A and orexin-B on the hypothalamic-pituitary-adrenal (HPA) system were investigated. An in vivo system was used to assess the central effects of both orexin-A and orexin-B. Different doses of the orexins (2.8-560 pmol) were administered intracerebroventricularly (i.c.v.) to adult male rats, and plasma corticosterone was used as an index of the degree of the activation of the HPA system. Both peptides exhibited a clear dose-response action, although orexin-B proved to be less effective than orexin A. Pretreatment with the corticotropin-releasing hormone (CRH) antagonist alpha helical CRH9-41 completely prevented the action of the orexins. Orexin-A, orexin B or adrenocorticotropic hormone (ACTH) was further administered intraperitoneally (i.p.). While ACTH evoked a significant adrenal response, the orexins did not influence the basal secretion. Adrenal slices, oxygenized and perifused with Krebs' solution, were also treated with orexin-A, orexin-B or ACTH. Both orexins failed to modify the release of corticosterone, but ACTH induced a marked adrenal response. This study suggests that these appetite regulating peptides might activate the HPA system at a central level but neither orexin-A nor orexin-B appears to modulate directly the adrenal corticosterone release. PMID- 11106975 TI - Vasopressin (V1a) receptor binding, mRNA expression and transcriptional regulation by androgen in the Syrian hamster brain. AB - Arginine vasopressin plays an important role in the regulation of social behaviours in rodents. In the Syrian hamster, vasopressin injected directly into the brain stimulates scent marking and aggressive behaviour in a steroid dependent manner and is therefore a useful model for investigating steroid peptide-behaviour interactions. In this study, we used in situ hybridization and radioligand binding assays on adjacent sections of hamster brains to compare the relative distribution of vasopressin (V1a) receptor mRNA and V1a receptor binding. V1a receptor mRNA and binding are abundant in the lateral septum, bed nucleus of the stria terminalis, medial preoptic nucleus, anterodorsal thalamus and suprachiasmatic nucleus. Moderate receptor binding and low levels of receptor mRNA are present in the central nucleus of the amygdala and a lateral zone from the medial preoptic area through the anterior hypothalamus. V1a receptor mRNA is anatomically more restricted in several areas compared to the ligand binding pattern, which is consistent with significant spread of receptor protein along neuronal processes. Comparison of V1a receptor ligand binding and mRNA in intact, castrated, and castrated-testosterone treated animals reveals that V1a receptors in the medial preoptic nucleus are regulated by androgen, most likely by an upregulation of V1a receptor gene expression in a cluster of neurones concentrated in the ventromedial part of this nucleus. This study confirms the presence of the V1a subtype of vasopressin receptors in behaviourally important regions of the hamster brain and suggests that transcriptional regulation by gonadal steroids may play a role in modulating behavioural sensitivity to vasopressin. PMID- 11106976 TI - Priming with interleukin-1beta suppresses experimental allergic encephalomyelitis in the Lewis rat. AB - Lewis rats exhibit multiple defects in their hypothalamus-pituitary-adrenal (HPA) system that are considered to play a causal role in the susceptibility of this strain to autoimmune diseases, i.e. experimental allergic encephalomyelitis (EAE). In the present study, we aimed to modulate the HPA response of the Lewis rat and establish its consequences for the susceptibility to EAE. Because in Wistar rats, single administration of interleukin (IL)-beta (priming) is known to induce long-lasting (weeks) sensitization of HPA responses to stressors and immune stimuli, Lewis rats were given a single dose of hIL-1beta or vehicle 1 week prior to induction of EAE by immunization with myelin basic protein (MBP). Subsequently, neurological deficits were monitored once daily. The results show that IL-1 priming markedly suppresses the neurological symptoms of EAE, without affecting the onset or duration of the disease. Measurement of vasopressin and corticotropin releasing hormone (CRH) in the external zone of the median eminence revealed that, as compared to Wistar rats, Lewis rats exhibit low vasopressin but identical CRH, and that IL-1 priming increases (0.001) vasopressin without affecting CRH stores, which is consistent with a shift to vasopressin-dominated control of adrenocorticotropic hormone (ACTH) secretion as described in Wistar rats under conditions of HPA hyper(re)activity. However, IL-1 priming did not affect a.m. corticosterone levels following immunization with MBP or during the clinical phase of EAE. IL-1 priming of Lewis rats attenuated the ACTH responses to an IL-1 challenge 11 days later, which may relate to an increase in resting corticosterone levels. Thus, the mechanisms underlying IL-1 induced suppression of EAE are not related to enhanced HPA responses. In addition, we did not find IL 1 priming-induced alterations in MBP-specific immunoglobulin (Ig)M, IgG1, IgGa and IgGb plasma titres, or gross alterations in T cell activation as reflected in spontaneous or concanavalin-induced T cell proliferation. We therefore speculate that IL-1-induced elevation of resting corticosterone levels may influence the development of EAE. PMID- 11106977 TI - Intracerebroventricular administration of the rat growth hormone (GH) receptor antagonist G118R stimulates GH secretion: evidence for the existence of short loop negative feedback of GH. AB - Pulsatile growth hormone (GH) secretion is regulated by three hypothalamic factors, growth hormone-releasing hormone (GHRH), somatostatin and the natural ligand for the GH secretagogue receptor (Ghrelin). These factors and their effects are, in turn, affected by short loop feedback of GH itself. To test the hypothesis that hypothalamic GH receptors are involved in the ultradian rhythmicity of pituitary GH secretion, the rat GH receptor antagonist (G118R) was administered to adult male rats by intracerebroventricular (i.c. v.) injection and the effects on spontaneous GH secretion were studied. Normal saline was administered i.c.v. to eight control rats. Mean GH concentrations increased significantly in the rat treated with G118R compared to rats that received normal saline. The pulse amplitude rose by a mean of 33.3 ng/ml and the total area under the curve increased by a mean of 15 061 ng/ml x min. The number of GH peaks did not change significantly following G118R. These data suggest that GH regulates its own secretion by acting directly on hypothalamic GH receptors. PMID- 11106978 TI - Effects of female pheromones on gonadotropin-releasing hormone gene expression and luteinizing hormone release in male wild-type and oestrogen receptor-alpha knockout mice. AB - Pheromones are an important class of environmental cues that affect the hypothalamic-pituitary-gonadal axis in a variety of vertebrate species, including humans. When male mice contact female-soiled bedding, or urine, they display a reflexive luteinizing hormone (LH) surge within 30 min. Aside from the requirement that males have gonads to show this response, the physiological mechanisms that underlie this pituitary response are unknown. In this experiment, we asked if female pheromones acted at the level of gonadotropin-releasing hormone (GnRH) gene expression to affect this hormone response. In addition, we also examined the contribution of one of the oestrogen receptors (ERalpha) by studying this neuroendocrine reflex in wild-type and oestrogen receptor-alpha knockout (ERalphaKO) males. Both ERalphaKO and wild-type males showed the expected LH surge, 45 and 90 min after contact with female pheromones. Males housed in clean bedding or bedding soiled by another adult male did not display the LH elevation. Interestingly, this dramatic change in LH concentrations was not accompanied by any alterations in GnRH mRNA expression or levels of primary transcript in the preoptic area-anterior hypothalamus. The one exception to this was a significant increase in GnRH mRNA expression in tissue collected from wild type males exposed to bedding from another male. This is particularly intriguing since LH was not elevated in these males. These data replicate and extend our previous finding that ERalphaKO males do exhibit an LH surge in response to female pheromones. Thus, this neuroendocrine response is regulated by a steroid receptor other than ERalpha and does not require alterations in GnRH mRNA expression. PMID- 11106979 TI - The gonadotropin-releasing hormone neurosecretory system of the jerboa (Jaculus orientalis) and its seasonal variations. AB - The distribution of cells expressing gonadotropin-releasing hormone (GnRH) immunoreactivity was examined in the brain of adult jerboa during two distinct periods of the reproductive cycle. During spring-summer, when the jerboa is sexually active, a high density of cell bodies and fibres immunoreactive (IR) for GnRH was observed at the level of separation of the frontal lobes, in the medial septal nucleus (MS) and in the diagonal band of Broca (DBB), in the preoptic area (POA), in the organum vasculosum laminae terminalis (OVLT), in the retrochiasmatic area and hypothalamus. In autumn, when the jerboa is sexually inactive, GnRH-immunoreactivity was less intense than during spring-summer. In the POA, we noted a 55% decrease in the number of GnRH containing cells with no change in cell numbers in the MS-DBB. Furthermore, a lower density of GnRH immunopositive axon fibres is observed in all the previously mentioned structures and the immunoreaction intensity was very weak particularly within the median eminence and OVLT. Independently of the season, the GnRH immunoreactivity within neurones and fibres was similar in jerboas living in captivity and in jerboas living in their natural biotope. The effects of photoperiod on the density of POA GnRH and arcuate nucleus beta-endorphin-containing cells were studied in jerboas maintained in long day [(LD) 16-h light, 8-h dark] and short day [(SD) 8-h light, 16-h dark] for 8 weeks. In the POA, the GnRH-IR cell number was not significantly altered by the photoperiod. Similarly, in the mediobasal hypothalamus, the number of beta-endorphin-IR neurones was not affected by such a parameter. Consequently, the GnRH seasonal variations cannot be correlated to changes in the photoperiod alone. PMID- 11106980 TI - Central administration of orexin A suppresses basal and domperidone stimulated plasma prolactin. AB - Orexin immunoreactive fibres are abundant in the hypothalamus suggesting a neuroendocrine regulatory role. Intracerebroventricular (ICV) administration of orexin A suppressed plasma prolactin in male rats by 71% at 20 min post-injection and 83% at 90 min post-injection (P < 0.005 vs saline at both time points). To investigate whether this effect was through the tuberoinfundibular dopaminergic (TIDA) system, a supra-maximal dose of domperidone, a dopamine receptor antagonist, was injected intraperitoneally (i.p.) prior to ICV injection of orexin A. ICV orexin A significantly suppressed domperidone (9 mg/kg)-stimulated plasma prolactin levels, by up to 40% (i.p. domperidone + ICV orexin A 3 nmol 34.5 +/- 7.4 ng/ml and i.p. domperidone + ICV orexin A 20 nmol 43.5 +/- 4.3 ng/ml, both P < 0.005 vs i.p. domperidone + ICV saline 57.9 +/- 2.7 ng/ml). Orexin A, 100 nM, significantly stimulated release of neurotensin, vasoactive intestinal polypeptide, somatostatin, corticotropin releasing factor and luteinizing hormone releasing hormone, but had no effect on release of dopamine, thyrotropin releasing hormone (TRH), vasopressin or melanin-concentrating hormone from hypothalamic explants in vitro. Orexin A did not alter basal or TRH stimulated prolactin release in dispersed pituitary cells harvested from male rats. The data suggest that ICV administration of orexin A suppresses plasma prolactin in part through a pathway independent of the dopaminergic system. PMID- 11106981 TI - Modulation of pituitary dopamine D1 or D2 receptors and secretion of follicle stimulating hormone and luteinizing hormone during the annual reproductive cycle of female rainbow trout. AB - The two gonadotrophins follicle stimulating hormone (FSH) and luteinizing hormone (LH) have distinct temporal expression and release profiles in fish, but little is known regarding their neuroendocrine control, especially for FSH. The present experiments were performed on previtellogenic, mature and preovulatory female trout. The catecholamine synthesis inhibitor, alpha-methyl-p-tyrosine, increased plasma LH and FSH concentrations of mature fish. The dopamine agonist apomorphine decreased and the dopamine antagonist domperidone increased plasma LH concentration of preovulatory fish and delayed ovulation, but did not modify plasma FSH concentration. The dopamine D2 agonist bromocryptine inhibited LH release in cultured gonadotrophs from mature and preovulatory fish, but not from previtellogenic fish. Bromocryptine also significantly inhibited basal and salmon gonadotrophin releasing-hormone (sGnRH)-induced FSH release from cultured gonadotrophs of mature fish, but not of preovulatory fish, and increased FSH release from gonadotrophs of previtellogenic fish. The dopamine D1 agonist SKF 38393 had no observed effect on the release of FSH and LH, at any reproductive stage studied. The D1 agonist SKF 38393, the D2 agonist bromocriptine and sGnRH had no observed effects on cell contents of FSH and LH. Taken together, these data suggest that, at the level of the pituitary, dopamine inhibits LH release as vitellogenesis proceeds, via activation of dopamine D2 receptors. We demonstrate for the first time in fish a control of FSH release (a dopamine control), especially in mature fish which have low circulating concentrations of FSH. PMID- 11106983 TI - Considerations in measuring the electrical potentials of metallic restorations in vivo. AB - Many variables are believed to affect the accurate measuring of metallic restoration electrical potentials. This study examined the effects of intra- versus extra-oral location of the reference electrode, the type of metallic probe used to make contact with the restoration, and scratching and brushing of the restoration surface. Dental amalgam restorations were measured in 40 human subjects. Results showed that only the location of the reference electrode affected the central tendency of the potential. The study discusses the significance of some experimental variables in the accurate measuring of metallic potentials and the need to consider individual subject differences when statistically analysing for the central tendency of a sample. PMID- 11106982 TI - Influence of silanization and filler fraction on aged dental composites. AB - The effect of silanization and filler fraction on the mechanical properties of aged dental composites was investigated. Experimental composites (75/25 Bis GMA/TEGDMA resin reinforced with 0, 12.6, 30.0, and 56.5 vol% 8 microm silanized/unsilanized BaSiO6) were fabricated into 4.7 mm diameter x 2.2 mm thick discs and 3.5 mm diameter x 7.3 mm thick discs for diametral tensile and compressive tests, respectively. The effect of immersion in 75% ethanol at 37 degrees C for 0-30 days on the diametral tensile strength (DTS) and compressive strength (CS) of the samples was evaluated and analysed by ANOVA and Tukey LSD test. The fracture interface between filler and resin matrix was then examined by scanning electron microscope. Results and subsequent statistical evidence from DTS (18.6+/-7.6 MPa, silanized versus 11.7+/-2.6 MPa, unsilanized) and CS (85.1+/ 29.7 MPa, silanized versus 56.0+/-11.3 MPa, unsilanized) strongly implies that silanization may greatly enhance the mechanical properties of the resin composites. Furthermore, it also shows that both DTS and CS increased proportionally as the filler fraction of the composites increased. However, in the unsilanized groups, DTS decreased (up to 40%) as the filler fraction increased, and CS showed no relevance to the filler fraction at all. As for the influence of aging, it was found that both DTS and CS showed a significant decrease after immersion in 75% ethanol, and silanization heavily correlated with the filler fraction of aged-resin composites. Microscopic examination of the fractured samples showed that failure primarily occurred within the resin matrix per se for silanized composites and adjacent to the filler particles for unsilanized composites. All the evidence points to the conclusion that mechanical properties of aged-resin composites can be greatly influenced by silanization and the filler fraction. PMID- 11106984 TI - Dimensional measurement and finite element analysis of I-bar clasps in clinical use. AB - An I-bar clasp is a popular retainer for distal-extension removable partial dentures. However, there have been almost no evidence-based criteria on the mechanically preferable shape. The present study aimed to investigate the variations of dimension in I-bar clasps used in patients, and to clarify the effect of the variations on stiffness and stress of I-bar clasps by finite element analysis. Dimensions (thickness, width, taper, radius of curvature, length, relation to oral structures) of 23 I-bar clasps were measured. A three dimensional finite element model was made for each measured I-bar clasp with vertical and horizontal straight sections connected with a curved section. A concentrated load of 5 N was applied at the lowest point of the tip that contacted the abutment in the buccal direction. Maximal equivalent stress and stiffness of each clasp were evaluated. The measured dimension, stiffness, and maximum stress showed wide variations. Mean stiffness was far from the proper one, and mean stress was near the proportional limit of Co-Cr alloy. Considering the stiffness and stresses in this study, only six clasps out of 23 were appropriate. These results suggest that evidence-based criteria of preferable shape of I-bar clasps should be determined. PMID- 11106985 TI - Biomechanical calculation of human TM joint loading with jaw opening. AB - A three-dimensional, static mathematical calculation of the stomatognathic system was done to predict total temporomandibular joint (TMJ) loading at different levels of jaw opening. The model assumed that muscle forces acting on the mandible could be simulated by a combination of contractile components (CCs) and elastic components (ECs) and that static equilibrium existed within the body of the mandible. The model also imposed the constraint that any generated joint reaction force would act on the centre of the condyle. The results of the model demonstrated that under all conditions of opening and for all values of the elastic modulus selected, the forces between the TMJ condyle and the articular eminence were compressive in nature. The compressive force magnitude increased from 2.7 to 27.6 N incrementally as the jaw opened from 10 to 40 mm. Overall data in this study indicated that the TMJ tissues undergo low levels of compression at open positions up to 40 mm. Finally, the condition of trismus (increased jaw closing activation with opening) was simulated, the joint reaction force at 20 mm opening increased from 7.7 to 64.9 N with only a 20% activation of the closers. PMID- 11106986 TI - A method for quantifying overall satisfaction of complete denture patients. AB - Reproducible and quantitative evaluation of patient satisfaction with their complete dentures is of great importance for preoperative diagnosis, treatment planning and assessment of complete denture treatment outcome. This study attempted (1) to clarify the degree of contribution of various factors to overall satisfaction, and (2) to develop a method for quantitative assessment of overall satisfaction with complete dentures. Twelve satisfaction factors and a three grade scale were used to assess 302 complete denture patients. The contribution of each grade of the 12 factors to the overall satisfaction level was determined by multiple regression analysis. Seven factors were highly correlated to the overall satisfaction. Based on the level of contribution by these seven significant factors, a scoring method for satisfaction was established. Category scores for these factors were calculated and the sum of the category scores was converted to an integer between 0 and 100. The resulting quantification score was closely correlated with overall satisfaction. A protocol for scoring overall satisfaction was developed based on the scientifically analysed contribution of each satisfaction factor. PMID- 11106987 TI - The effect of venting on pulpward pressure transmission and seating on crown cementation: a laboratory study. AB - Measurable hydraulic forces towards the pulp chamber can occur whilst cementing a crown. This study investigated the effect of crown venting and the use of two different seating forces (100 and 2. 5 N) on pulpward pressure transmission. Forty single-rooted premolar teeth were collected from patients requiring extractions for orthodontic reasons. A standardized crown preparation was made and impressions taken to construct individual base-metal alloy crowns. The teeth were divided into four groups based on the type of crown (vented or non-vented) and force used (100 or 2.5 N). The teeth mounted on a specially constructed stand and connected to a 0-104 kPa pressure transducer. The pulp chamber was perfused with saline driven by nitrogen gas at 83 kPa for up to 2 h until a steady state perfusion rate was achieved. Crowns were cemented to the teeth on a uniaxial seating device using zinc phosphate cement. Simultaneous recordings of both pressure pulse and seating discrepancies were recorded. The 40-fold increase in seating force from 2.5 to 100 N resulted in a significant increase (P<0.001) in pulpward pressure in both vented and non-vented crowns. Seating was also significantly improved for the vented crowns regardless of which force was used. Venting had no significant effect on pulpward pressure transmission. It was concluded that a significant relationship existed between seating force and pulpward pressure transmission. Venting improved crown seating but did not have a demonstrable effect on pulpward pressure transmission. PMID- 11106988 TI - Changes in jaw-jerk on different levels of jaw closure and teeth-clenching in humans. AB - We investigated how the jaw-jerk in the human masseter muscle is modulated in relation to the level of jaw closure (JC) and teeth clenching. Electromyographic (EMG) activity was recorded with surface electrodes. Background EMG activity of the masseter muscle was kept at three low teeth clenching levels with visual feedback. The level of JC was changed in six steps along the habitual path of closure relative to the mean maximal jaw opening during gum chewing by inserting a bite block between the upper and lower molars. The jaw-jerk was evoked by applying mechanical stimulation of about 20 N with a hammer to the bite-fork placed on the lower molars on one side in each condition of combination of a level of JC with a level of teeth-clenching. At the resting condition the excitability of the jaw-jerk increased with JC, while at weak voluntary teeth clenching it then decreased and increased again as the jaw was progressively closed. It is suggested that the excitability of the jaw-jerk would increase toward the occlusal position, which in turn would contribute to smooth masticatory movements. In addition, the mode of modulation of the jaw-jerk was studied in a subject with skeletal malocclusion. PMID- 11106989 TI - Titanium for removable partial dentures (III): 2-year clinical follow-up in an undergraduate programme. AB - This study examined the success of titanium (Ti) removable partial dentures (RPDs) compared with that of cobalt-chromium (Co-Cr) RPDs using a randomized controlled clinical trial. Thirty-eight RPD patients were provided with either Co Cr (20 patients) or Ti (18 patients) RPDs. The total numbers of dentures was 31 (13 maxillary, 18 mandibular) for Co-Cr and 23 (11 maxillary, 12 mandibular) for Ti. Patients were reviewed for 24 months following denture issue. After the initial 12 months, 20 clinical problems were recorded and became the criteria for subsequent assessment. Incidence of failure was analysed using both Fisher's exact test and the chi square test at a significance level of P<0.05. Fracture of retainers in both metals occurred only in the first 12 months. Some failure types presented at significantly higher levels in the first 12 months but there were no significant differences between the two in all the criteria examined between the 12- and the 24-month reviews. Although differences existed in failure types between Co-Cr and Ti RPDs during the early review stages, the overall success rate of Ti RPDs was comparable with that of Co-Cr RPDs after 24 months. The higher incidence of failures in Ti RPDs prior to the 12-month review suggests the importance of taking its lower rigidity into account when designing RPDs. PMID- 11106990 TI - Electromyographic activity in patients with temporomandibular disorders. AB - Evaluation of masticatory muscle activity by surface electromyography (EMG) is a valuable tool for diagnosing dysfunction of the masticatory apparatus. However, controversy exists with regard to the usefulness of the EMG for patients with temporomandibular disorders (TMD). Forty patients with TMD were subjected to surface EMG of the masticatory muscles. These patients had consulted because of temporomandibular pain and clicks. In most cases (75%), the symptoms affected the patient's left side. Overall mean resting activity was 2.52 microV+/-1.25 microV (s.d.), which is slightly higher than in comparable healthy subjects (1.92+/-1.20 microV). Mean resting activity was highest in the anterior digastric muscle (3.49 microV) on the left side. Overall mean activity during clenching was 66.77+/ 35.22 microV, which is about half that observed in healthy subjects (110.30+/ 82.97 microV). During leftward movement of the jaw, activity was on average highest in the left digastric, while during rightward movement, activity was on average highest in the right anterior temporal (AT). Our results thus indicate that patients with temporomandibular joint (TMJ) disorder show: (1) a slight increase in basal tone; (2) a significantly reduced capacity for clenching; and (3) an apparently paradoxical inhibition of the dysfunctional-side AT during movement of the mandible towards that side. PMID- 11106991 TI - Saliva tannin interactions. AB - Many plant foods contain tannins, compounds that bind proteins, such as mammalian enzymes. Although described as tasteless, tannins can be detected orally by their astringency. However, the actual mechanism of oral detection and the effect of tannins on mastication and swallowing have been little investigated. Here, we show from in vitro tests that tannic acid, a common standard in tests used to detect tannins, significantly reduces the lubricating qualities of human saliva both by decreasing its viscosity and increasing friction, both factors lending support to the notion that astringency is a tactile phenomenon. From the literature, it is clear that this effect depends on the presence of salivary proline-rich proteins (PRP). In a mammalian context, ingestion of tannin-rich foods in a species with salivary PRP will be signalled by interference with bolus formation during mastication while the increase in friction may also be detectable and lead to increased tooth wear if the signal is ignored. In a human context, cross-cultural preferences for tannin-rich beverages such as tea, coffee and red wine at the end of meals may be explained by reduction in adhesion of food particles to the oral mucosa allowing their rapid oral clearance. PMID- 11106992 TI - Measuring system for lip movements using two video trackers. AB - Movements of soft tissues surrounding the oral cavity, especially lips and cheeks, have a strong influence on mastication and phonetics. They also influence the relationship between a denture and its oral environment. The purpose of this study was to develop a three-dimensional measuring system for soft tissue movement. This system consisted of two video trackers placed stereographically and a computer. In addition, one video tracker was connected for measuring mandibular movements. The accuracy of this system was evaluated using computerized XYZ pulse stage. The resolution of this system was 0.10 x 0.10 x 0.10 mm, when the measurement was carried out in the area of the 24.0 (X) x 20.0 (Y) x 20.0 (Z) mm with a working distance of 500 mm and a frequency of 120 Hz. In the present study, the lip movements of a dentulous subject with mandibular movements during chewing peanuts were analysed using this system. The new system demonstrated its value for analysing soft tissue movement. PMID- 11106993 TI - Difference in tracks between habitual open and close mandibular movements at the condyle in children. AB - Although previous studies have paid much attention to the condylar movement in adults with permanent dentition, little attention has been paid to such movement in children. In this study, we therefore clarified the difference in habitual open and close movements at the condyle in children. Three groups of subjects were used; primary (10 children), early mixed (10 children), and permanent dentition (10 adults). The habitual open and close mandibular movement of each subject was measured using a TRIMET, which can three-dimensionally analyse the simultaneous movements of multiple points on the mandible of a subject. The measurements were then compared among the three groups. The three-dimensional analysis detected significant difference in all directions between children with primary dentition and adults with permanent dentition: primary dentition had the smallest anterior-posterior and superior-inferior directions, and the largest left-right direction. Coincidence of the open and close tracks occurred in the adults (adult group) but not in the children (primary and early-mixed dentition groups). The early-mixed dentition group showed tracks that were between those for the primary dentition group and the adult group. These results suggest that the regularity of the condylar track might be well established with dental development. PMID- 11106994 TI - Influence of protrusive tooth contact on tapping point distribution. AB - This study investigated the influence of protrusive tooth contacts (tooth contacts during mandibular protrusion) on the tapping point distribution. Nine healthy subjects volunteered for this study and the protrusive tooth contact pattern, as well as the retrusive tooth contact pattern, was altered on four maxillary occlusal splints. The first splint was adjusted to make the sagittal incisal path of protrusion and retrusion equivalent to that of the natural dentition. The second and third splints had partial and complete elimination of the protrusive tooth contact, respectively. The fourth splint had complete elimination of both protrusive and retrusive tooth contacts. The subjects were asked to use each splint continuously for 1 week. The tapping point distribution was measured on the 7th day after insertion of each splint. The four experimental occlusal conditions were found to have a significant effect on the tapping point distribution. The complete elimination of the protrusive tooth contact caused an anterior tapping point location and an increase in the tapping point area. The former tendency was found to be independent of the presence of the retrusive tooth contact. In conclusion, it was suggested that the protrusive tooth contact plays a significant role in maintaining the consistency and stability of the tapping point. PMID- 11106995 TI - The behaviour of doramectin in the gastrointestinal tract, its secretion in bile and pharmacokinetic disposition in the peripheral circulation after oral and intravenous administration to sheep. AB - Sheep were 'compartmentalized' by surgically implanting cannulae in the rumen, abomasum and terminal ileum with a re-entrant cannula inserted between the cystic duct and the duodenum to monitor bile secretion. Doramectin, containing a trace of [3H]-doramectin, was administered both intravenously (i.v.) and intraruminally (i.r.) at a dosage of 150 microg/kg. The pharmacokinetic behaviour of [3H] labelled products was determined in these pools, and also in peripheral plasma, urine and faeces. Parent doramectin was also determined in plasma, abomasal digesta fluid and bile. Following i.r. administration, [3H] compounds were almost entirely associated with particulate digesta. A 14.5 h half-life in the rumen prolonged the presence of [3H] in the abomasum. Doramectin appeared to be degraded in abomasal digesta because only 24% of abomasal [3H] was attributed to the parent drug. Absorption of doramectin resulted in a systemic availability of 35%, of which 1.6 and 23.6% of the dose was contained in urine and biliary secretions, respectively. Following i.v. administration, almost negligible quantities of [3H] were secreted into the rumen or abomasum and only 2.7% of the dose was excreted in urine, whereas 132% was secreted in bile. This indicated that approximately one-third of biliary metabolites were enterohepatically recycled with biliary metabolites, elevating the proportion of [3H] in fluid digesta in the small intestine. Passage of the i.r.-administered drug through the gastrointestinal tract (GIT) resulted in virtually complete faecal excretion of [3H] within 5 days, whereas the continued secretion of i.v.-administered [3H] in bile prolonged the presence of [3H] in the GIT, with faecal clearance not being complete for at least 10 days. This multi-compartmental study has provided more information on the behaviour of doramectin than can be obtained from examining drug disposition in the peripheral circulation alone. With this knowledge, it is anticipated that opportunities for improving drug performance will be identified. PMID- 11106996 TI - Identification of lidocaine and its metabolites in post-administration equine urine by ELISA and MS/MS. AB - Lidocaine is a local anesthetic drug that is widely used in equine medicine. It has the advantage of giving good local anesthesia and a longer duration of action than procaine. Although approved for use in horses in training by the American Association of Equine Practitioners (AAEP), lidocaine is also an Association of Racing Commissioners International (ARCI) Class 2 drug and its detection in forensic samples can result in significant penalties. Lidocaine was observed as a monoprotonated ion at m/z 235 by ESI+ MS/MS (electrospray ionization-positive ion mode) analysis. The base peak ion at m/z 86, representing the postulated methylenediethylamino fragment [CH2N(CH2CH3)2]+, was characteristic of lidocaine and 3-hydroxylidocaine in both ESI+ and EI (electron impact-positive ion mode) mass spectrometry. In addition, we identified an ion at m/z 427 as the principal parent ion of the ion at m/z 86, consistent with the presence of a protonated analog of 3-hydroxylidocaine-glucuronide. We also sought to establish post administration ELISA-based 'detection times' for lidocaine and lidocaine-related compounds in urine following single subcutaneous injections of various doses (10, 40, 400 mg). Our findings suggest relatively long ELISA based 'detection times' for lidocaine following higher doses of this drug. PMID- 11106997 TI - Iontophoresis of dexamethosone-phosphate into the equine tibiotarsal joint. AB - In human rehabilitation medicine, dexamethasone-phosphate is theoretically iontophoresed to localized subcutaneous tissue where conversion to dexamethasone occurs. This delivery system has recently been introduced into veterinary medicine for the same purpose. However, the pharmacokinetic justification for parenteral delivery of this prodrug remains undocumented. Utilizing iontophoretic methods that are relevant to both human and veterinary clinical practice, the present investigation compared injection and iontophoresis of dexamethasone phosphate into the equine tibiotarsal joint, also known as the tarsocrual joint. The tibiotarsal joints of seven horses were injected with 4 mL of 6 mg/mL dexamethasone-phosphate. With a similar drug concentration and over the same application site, six different horses underwent simultaneous cathodic iontophoresis (4 mA, 40 min) or passive application (0 mA, 40 min) on contralateral limbs. Following all applications, tibiotarsal joint synovium was collected. Local venous blood samples were also collected from the iontophoretic and passive application sites for analysis of plasma drug concentrations. Because of the potential for conversion of dexamethasone-phosphate to dexamethasone, an extraction and analysis protocol was developed for both chemicals. The technique demonstrated a linear range of detection (0.39-12 microg/mL) and a capability for measuring both chemicals in plasma and synovium. Conversion of dexamethasone phosphate to dexamethasone occurred during synovial incubation (37 degrees C) and following freeze-thaw cycles. In contrast to the measurable synovial concentrations of dexamethasone-phosphate (2.3 +/- 0.96 mg/mL) and dexamethasone (0.27 +/- 0.07 mg/mL) following injection, neither drug was detected in the synovium or the local venous blood following iontophoretic or passive applications. In conclusion, these results do not confirm iontophoretic or passive delivery of measurable dexamethasone-phosphate into the tibiotarsal joint using current clinical methods. PMID- 11106998 TI - Intratracheal clenbuterol in the horse: its pharmacological efficacy and analytical detection. AB - Clenbuterol, a beta2 agonist/antagonist, is the only bronchodilator approved by the US Food and Drug Administration for use in horses. The Association of Racing Commissioners International classifies clenbuterol as a class 3 agent, and, as such, its identification in post-race samples may lead to sanctions. Anecdotal reports suggest that clenbuterol may have been administered by intratracheal (IT) injection to obtain beneficial effects and avoid post-race detection. The objectives of this study were (1) to measure the pharmacological efficacy of IT dose of clenbuterol and (2) to determine the analytical findings in urine in the presence and absence of furosemide. When administered intratracheally (90 microg/horse) to horses suffering from chronic obstructive pulmonary disease (COPD), clenbuterol had effects that were not significantly different from those of saline. In parallel experiments using a behavior chamber, no significant effects of IT clenbuterol on heart rate or spontaneous locomotor activity were observed. Clenbuterol concentrations in the urine were also measured after IT dose in the presence and absence of furosemide. Four horses were administered i.v. furosemide (5 mg/kg), and four horses were administered saline (5 mL). Two hours later, all horses were administrated clenbuterol (IT, 90 microg), and the furosemide-treated horses received a second dose of furosemide (2.5 mg/kg, i.v.). Three hours after clenbuterol dose (1 h after hypothetical 'post-time'), the mean specific gravity of urine samples from furosemide-treated horses was 1.024, well above the 1.010 concentration at which furosemide is considered to interfere with drug detection. There was no interference by furosemide with 'enhanced' ELISA screening of clenbuterol equivalents in extracted and concentrated samples. Similarly, furosemide had no effect on mass spectral identification or quantification of clenbuterol in these samples. These results suggest that the IT dose of clenbuterol (90 microg) is, in pharmacological terms, indistinguishable from the dose of saline, and that, using extracted samples, clenbuterol dose is readily detectable at 3 h after dosing. Furthermore, concomitant dose of furosemide does not interfere with detection or confirmation of clenbuterol. PMID- 11106999 TI - Chiral inversion of R(-) fenoprofen and ketoprofen enantiomers in cats. AB - The chiral inversion process is a characteristic metabolic pathway for different aryl-2-propionic acids or profens. Important variations have been observed between these individual compounds as well as between animal species. In this study, R(-) fenoprofen [R(-)FPF] and R(-) ketoprofen [R(-) KTF] were used to investigate their comparative stereoconversion in cats. After intravenous (i.v.) administration of R(-) FPF, the percentage of chiral inversion was 93.20+/ 13.70%. A highly significant correlation (r: 0.978) was observed between the clearance of R(-) FPF and the chiral inversion process. After i.v. administration of R(-) KTF, the percentage of inversion was only 36.73+/-2.8%. No correlation between the clearance of R(-) KTF and this process was observed. R(-) FPF was metabolized by the pathways of thioesterification - chiral inversion processes. For R(-) KTF, the competitive metabolic pathways, glucuronidation and hydroxylation may be involved. However, these metabolic steps are saturable or less functional in cats. Moreover, the thioesterification of R(-) KTF in in vitro studies has been shown to be important in carnivores. The lack of correlation between clearance and chiral inversion process of R(-) KTF may be finally explained by deviation of thioesterification to other metabolic pathways of lipids and/or aminoacid conjugation, particulary glicine derivatives. PMID- 11107000 TI - Comparison of plasma pharmacokinetics and bioequivalence of ceftiofur sodium in cattle after a single intramuscular or subcutaneous injection. AB - Ceftiofur sodium, a broad-spectrum cephalosporin, is active against gram-positive and gram-negative pathogens of veterinary importance. This study was designed to compare the bioequivalence of the sodium salt in cattle after a single intramuscular (i.m.) or subcutaneous dose (s.c.) of 2.2 mg ceftiofur equivalents/kg body weight. The criteria used to evaluate bioequivalence were (1) the area under the curve from time of injection to the limit of quantitation (LOQ) of the assay (AUC0-LOQ), and (2) time concentrations remained above 0.2 microg/mL (t>0.2). Twelve crossbred beef cattle were enrolled in a three-period, two-treatment crossover trial, with a minimum 2-week washout period between doses of 2.2 mg ceftiofur equivalents/kg. Blood samples were collected serially for up to 72 h post-injection. Plasma samples were then analyzed using a validated assay that measures ceftiofur, and all desfuroylceftiofur-related metabolites, by high performance liquid chromatography (HPLC) as the stable derivative, desfuroylceftiofur acetamide. A maximum plasma concentration (Cmax) of 13.9+/ 3.55 microg/mL was observed from 0. 67-2.0 h after i.m. administration, whereas a Cmax of 13.6+/-3.85 microg/mL was observed from 0.67-3.0 h after s.c. administration. The AUC0-LOQ was 108+/-35.0 microg. h/mL after i.m. dosing, compared with 105+/-29.8 microg. h/mL after s.c. dosing. The pre-established criterion for equivalence of the AUC0-LOQ for the i.m. and s.c. routes of administration was satisfied. The t>0.2 was 49.2+/-8.55 h after i.m. administration, compared with 47.0+/-9.40 h after s.c. administration. The pre established criterion for equivalence of the t>0.2 for i.m. and s.c. administration was satisfied. The equivalence of AUC0-LOQ and t>0.2 for i.m. and s.c. administration of 2.2 mg ceftiofur equivalents (CE)/kg doses of ceftiofur sodium suggest similar therapeutic efficacy and systemic safety for the two routes of administration. PMID- 11107001 TI - Pharmacokinetics and residual behaviour in milk of oxytetracycline in cows following administration of uterine pessaries. AB - The plasma kinetics and residual depletion in milk of cows treated by the intrauterine route with pessaries containing oxytetracycline (OTC) were evaluated. The antibiotic was administered to five healthy Friesian cows at a dosage of 3g/head in the early post partum phase. Blood samples were collected before and at different time intervals (3, 6, 12, 24, 48, 72, 84, and 96 h) after treatment. Milk was drawn before treatment and at 12-h intervals for 4 consecutive days. Samples were analysed by a high-performance liquid chromatography method and the pharmacokinetic parameters were processed using the minimum Akaike information criterion estimation (MAICE) test. The mean values obtained indicated a relatively low area under the concentration time curve (25.19+/-12.61 microg/mg per h) and maximum plasma concentration (Cmax) (0.549+/ 0.278 microg/mL) with delayed time to Cmax (11.71+/-4.15 h) and elimination half life (21.96+/-4.42 h). A similar pattern could be shown for milk, in which measurable residual levels are found in two out of five animals until the 72nd hour after treatment. Data obtained demonstrate that OTC administered as a solid form is poorly and slowly absorbed from the uterus of cows. PMID- 11107002 TI - Pharmacokinetics and metabolic effects of triamcinolone acetonide and their possible relationships to glucocorticoid-induced laminitis in horses. AB - Experiments were performed to establish the pharmacokinetics of triamcinolone acetonide and the effects of the glucocorticoid on glucose metabolism in horses. The pharmacokinetics after intravenous (i.v.) dosing was best described by a three-compartment open model. There was rapid distribution from the central compartment followed by two phases of elimination. The half-life of the rapid elimination phase was 83.5 min and of the slower phase was 12 h. The term (Vss/Vc)-1was 12.3 indicating extensive distribution into the tissues. Triamcinolone acetonide given i.v. or intramuscularly (i.m. ) induced a prolonged period of hyperglycaemia, hyperinsulinaemia and hypertriglyceridaemia. Significant changes in plasma glucagon and serum non-esterified fatty acids were not observed. These observations suggest that the hyperglycaemia was a result of decreased glucose utilization by tissues and increased gluconeogenesis. The effects on glucose metabolism persisted for 3-4 days after triamcinolone was given i.m. at 0.05 mg/kg, the upper limit of the recommended dose range, and for 8 days when given at 0. 2 mg/kg. These observations, together with recent evidence implicating inhibition of glucose metabolism in the pathogenesis of equine laminitis, indicated that triamcinolone-induced laminitis may be associated with the long duration of action of the glucocorticoid when higher than recommended doses or when repeated doses are given. PMID- 11107004 TI - Tissue distribution of benzylpenicillin after intramammary administration in the isolated perfused bovine udder. AB - Udders from previously healthy lactating cows were perfused with warmed and gassed Tyrode solution in vitro. Benzylpenicillin was administered in three formulations: an oily suspension with micronized particles of <10 microm diameter, an oily suspension with average particle size of 40 microm and an aqueous solution (3 million IU benzylpenicillin-potassium, volume 15 mL). The antibiotics were administered intracisternally to six front and six rear quarters each. Moreover, a dry-off-ointment (100 000 IU benzylpenicillin-potassium and 100 000 IU benzylpenicillin-benzathine, volume 7.5 mL) was tested in four udder halves. Perfusate samples were collected over 3 h. Furthermore, glandular tissue at different vertical distances from the teat base and the regional lymph node were sampled after 3 h. The determination of benzylpenicillin was performed by high pressure liquid chromatography with UV detection. With increasing distance from the teat base, the concentration of benzylpenicillin in tissue exponentially decreased. Using the aqueous solution or oily suspension that contained micronized active principle, higher concentrations were reached compared to the formulation with particle sizes of 40 microm. In udder lymph nodes, the concentration was highest after treatment with the coarse suspension. The transfer from the dry-off-ointment with benzathine-salt into perfusate was very low. These results suggest that it is possible to study tissue distribution of antibiotics in the isolated perfused bovine udder. PMID- 11107003 TI - Comparison of fluoroquinolone pharmacokinetic parameters after treatment with marbofloxacin, enrofloxacin, and difloxacin in dogs. AB - Plasma, urine, and skin drug concentrations were determined for dogs (n=12) given five daily oral doses of marbofloxacin (MAR) (2.75 mg/kg), enrofloxacin (ENR) (5.0 mg/kg) or difloxacin (DIF) (5.0 mg/kg). Concentrations of the active metabolite of ENR, ciprofloxacin (CIP), were also determined. The three-period, three-treatment crossover experimental design included a 21-day washout period between treatments. Area under the plasma drug concentration vs. time curve (AUC0 last, microg/mLxh of MAR was greater than for ENR, CIP, ENR/CIP combined, and DIF. Maximum concentration (Cmax) of MAR was greater than ENR, CIP, and DIF. Time of maximum plasma concentration (Tmax) was similar for MAR and DIF; Tmax occurred earlier for ENR and later for CIP. Plasma half-life (t1/2) of MAR was longer than for ENR, CIP, and DIF. Urine concentrations of DIF were less than MAR or ENR/CIP combined, but urine concentrations of MAR and ENR/CIP combined did not differ. DIF skin concentrations were less than the concentrations of MAR or ENR/CIP combined 2 h after dosing, but skin concentrations of MAR and ENR/CIP combined did not differ. PMID- 11107005 TI - Non-linear pharmacokinetics of ofloxacin after a single intravenous bolus dose in pigs. AB - The pharmacokinetics of ofloxacin (OFLX) was investigated after intravenous administration of 3, 10 and 30 mg/kg of body weight in pigs. Plasma OFLX concentration-time course collected from the highest dosage showed a convex decline, indicating a capacity-limited process in drug elimination (Michaelis Menten elimination). Dose-normalized area under curve (AUC/Dose) and mean resident time (MRT) were dose-dependent, indicating a classical pattern of non linear elimination pharmacokinetics. Based on simultaneous curve fitting from three doses, non-linear pharmacokinetic parameters were as follows: 0.87 mg/h/kg for maximum velocity, 2.20 microg/mL in Michaelis-Menten constant and 2.06 L/kg for apparent volume of distribution. Based on a model-independent analysis, the apparent volume of distribution at steady-state (Vdss) was dose-independent whereas total body clearance (CLtot) was dose-dependent, mainly contributed by renal clearance (CLr) with the regression line of CLtot=1.14xCLr+0.09 (r=0.92). The intercept of the regression line indicates non-renal clearance (CLnr), corresponding to the value of observed CLnr without dose-dependency. Because of a higher CLr compared with glomerular filtration rate (GFR) in spite of drug reabsorption, the CLr must contain the renal active tubular secretion. With increasing dosage, the level of saturation of tubular secretion of OFLX decreased the CLr, resulting in the decrease in CLtot. The plasma protein binding to OFLX was dose-independent: mean free fraction (fp)=0.73, with probably no influential effect on OFLX disposition. In conclusion, the degree of saturation in the renal active tubular secretion of OFLX could be a major causal factor in the alteration of CLr in an increasing dosage of OFLX. Accordingly, the alteration of CLr could directly induce the non-linear pharmacokinetics of OFLX in pigs, an important consideration in clinical therapeutics. PMID- 11107006 TI - Intravenous pentoxifylline does not affect the exercise-induced pulmonary arterial, capillary or venous hypertension in Thoroughbred horses. AB - The present study was carried out to examine whether intravenously administered pentoxifylline-a phosphodiesterase inhibitor which increases red blood cell deformability and decreases blood viscosity-would attenuate the magnitude of exercise-induced pulmonary capillary hypertension in healthy, fit Thoroughbred horses and in turn, diminish the occurrence of exercise-induced pulmonary hemorrhage (EIPH). Experiments were carried out on six healthy, sound, exercise trained Thoroughbred horses. Hemodynamic data were collected at rest, and during exercise performed at 8 and 14 m/sec on 3.5% uphill grade in the control (no medications) and the pentoxifylline (8.5 mg/kg, i.v.) experiments. The sequence of treatments was randomized for every horse and 7 days were allowed between treatments. Galloping at 14 m/sec on 3.5% uphill grade elicited maximal heart rate. In both treatments, simultaneous measurements of phasic and mean right atrial and pulmonary arterial, capillary and wedge pressures were made using catheter-tip-manometers whose signals were carefully referenced at the point of the left shoulder. In the control study, exercise resulted in progressive significant increments in heart rate, right atrial and pulmonary arterial, capillary and venous pressures; thereby, confirming that exercising Thoroughbreds develop significant pulmonary hypertension. All horses experienced exercise induced pulmonary hemorrhage (EIPH) in the control experiments. Pentoxifylline administration to standing horses caused anxiety, tachycardia, muscular fasciculations/tremors and mild sweating, but statistically significant changes in right atrial and pulmonary arterial, capillary and venous pressures were not detected. Exercise in the pentoxifylline treatment also resulted in progressive significant increments in heart rate and right atrial as well as pulmonary vascular pressures, but these data were not statistically significantly different from those in the control study and the incidence of EIPH remained unchanged. Thus, it was concluded that i.v. pentoxifylline is ineffective in attenuating the exercise-induced pulmonary arterial, capillary and venous hypertension in healthy, fit Thoroughbred horses. PMID- 11107008 TI - In this issue PMID- 11107007 TI - Involvement of platelet activating factor in the endotoxin-induced effects on gastrointestinal electrical activity and some haematological parameters in the conscious miniature pig. AB - In conscious miniature pigs the influence of intravenous dose of lipopolysaccharide (LPS), 10 microg/kg over 10 min, with and without pretreatment with a platelet activating factor (PAF) antagonist, SAH 63-675 10 mg/kg, on gastrointestinal electrical activity, arterial pressure and clinical and haematological parameters was studied. Dose of LPS provoked mild clinical signs and hypotension, which were prevented by PAF antagonism. The LPS induced leukocytosis and increase in mature neutrophils, however, were PAF independent. Pretreatment with the PAF antagonist attenuated the LPS-provoked inhibition of electrical activity in the antrum, jejunum, ileum and caecum. These results suggest a beneficial effect of PAF antagonism in porcine endotoxaemia. PMID- 11107009 TI - Developing the journal PMID- 11107010 TI - Reflecting on levels of confidence and competence in skills acquisition. PMID- 11107011 TI - On theory in medical education. PMID- 11107012 TI - American medical students in Israel: stress and coping--a follow-up study. AB - BACKGROUND: Medical students studying abroad face the double stress of adjusting to a new cultural environment while at the same time, coping with the usual stresses of medical school. In a previous article, we examined the perceived stress and coping of American medical students studying in Israel. AIMS: The current study was designed to follow up changes in made in response to the original study. PARTICIPANTS: First year students, NY/American Program, Sackler School of Medicine, Tel Aviv University, Israel. METHODS: Ways of Coping Checklist (WCCL), Appraisal Dimension Scale (ADS) and two instruments specifically designed for the study. RESULTS: Students' coping with their adjustment to Israel was highly correlated to their adjustment to medical school. There was significant improvement in student mental health and student satisfaction and a corresponding reduction in dysfunctional defence mechanism and a previous pattern of heavy drinking. DISCUSSION: The results are discussed in terms of improvements in the student support system proposed at the time of the initial study as well as changes in the student body. Limitations and future directions for research are also discussed. PMID- 11107013 TI - Becoming professional: when and how does it start? A comparative study of first year medical and law students in the UK. AB - AIM: The purpose of this study is to investigate whether differences identified between first-year law and medical students in North America in the 1950s apply in the UK in the 1990s. First-year law and medical students are compared in terms of commitment to career, alternative career choices and length of time the student has wished to study for his/her chosen profession. METHOD: Questionnaires were administered to first-year law students at the University of East Anglia and Essex University and to first-year medical students at Liverpool University Medical School and St George's Hospital Medical School. A total of 162 questionnaires were completed by law students and 195 questionnaires from medical students. ANALYSIS: The questionnaire responses provided by law and medical students were analysed using a series of two-sample comparisons. Differences between the two groups were examined using t and chi-squared tests. In each of the seven questions answered by students, the differences between the law and medical students were found to be significant. This suggests a difference in career aspirations and perceptions between the two groups. RESULTS AND CONCLUSIONS: The study shows a greater commitment of medical students than law students to their chosen career. This is demonstrated by medical students' greater desire to pursue their career, their greater satisfaction with their choice of career and finding that more medical students would persist with reapplying for medicine than law students would in reapplying for law. It is also shown that medical students are twice as likely as law students to have a family member within the profession. PMID- 11107014 TI - Clarifying the concepts of confidence and competence to produce appropriate self evaluation measurement scales. AB - INTRODUCTION: This paper reviews the literature on self-evaluation and discusses the findings of a small-scale qualitative study which explored the terms 'confidence' and 'competence' as useful measures in a self-evaluation scale. Four pre-registration house officers took part in interviews and completed a provisional instrument to assess their perceived competence. FINDINGS: Competence and confidence are useful terms for house officers expressing beliefs about their ability to perform their job but the terms should not be used synonymously. In our study, 'competent' represented what individuals knew about their ability and was based on the individual's previous experience of the task. 'Confident' described a judgement which influenced whether an individual was willing or not to undertake an activity. Confidence was not necessarily based on known levels of competence and therefore performance of tasks which were unfamiliar to the house officer also involved the assessment of risk. The authors give examples of task and skill scales which may be useful in the process of self-evaluation by pre registration house officers. CONCLUSIONS: The authors suggest that the process of assessing oneself is complicated, and by its very nature can never be objective or free from the beliefs and values individuals hold about themselves. Therefore self-evaluation instruments are best used to help individuals analyse their work practices and to promote reflection on performance. They should not be used to judge the 'accuracy' of the individual's evaluation. PMID- 11107015 TI - Academic careers in general practice and primary care. AB - In the past 10 years, significant developments in general practice teaching and research have led to the considerable growth of academic general practice as a discipline. This paper reviews issues relating to these developments, particularly career pathways and training aspects. The need to extend these advances to the broadening arena of primary health care has given further impetus for the development of academic careers. General practice will need to work closely with secondary care, community health, and social services to develop primary health care in its broadest sense, and an evidence base, generated by relevant research and evaluation, must underpin all of this. Structural and funding changes to undergraduate education, postgraduate training and primary care research have created a range of opportunities for general practice clinicians to define career pathways, not formerly available, within multiprofessional and multidisciplinary departments and groups. Education for future general practice and primary care must underpin developments as much as a research base. Relevant masters' degrees and diplomas are now widely available, and extended vocational training and higher professional education will enable general practitioners in their formative years to consider academic opportunities. PMID- 11107016 TI - The diffusion of the health agenda and the fundamental need for partnership in medical education. AB - This paper explores the fundamental reasons for partnership in health care and medical education. It reviews the philosophical and policy contexts of health care trends and suggests that many of these trends can be summarized as a process of diffusion relating to: (a) what is on the health agenda, (b) who sets the health agenda and (c) the increasing indeterminacy of the health agenda. Various aspects of the 'social turn' in health care are introduced and offered as a partial explanation for the diffusion of the health agenda. Finally, some of the implications of these discussions for medical education are set out, in particular the need for partnerships within and beyond the academy. PMID- 11107017 TI - Putting teamwork in context. AB - Multidisciplinary teamwork is becoming more important in both the delivery of health care and in the organization and management of that delivery. The first of these has been accepted but traditional professional education has done little to address the challenge it presents to professionals. Recent reforms in the British NHS have made the challenge more urgent. Professionals must work together but in increasingly flexible and innovatory ways. They are also required to play more formal roles in NHS management and policy. Where teamwork has been addressed in professional education it has concentrated on the inter-personal dynamics of working teams. This remains important but to respond effectively to the new challenges curricula and educational practice will have to be clearer about the variety of teams involved and the importance of the context within which teams work. One view is offered as to how that context might be understood in order to map team diversity. Two models are offered to help develop multidisciplinary team learning. One of these deals with key aspects of the organizational setting and the other with factors that affect team processes. It is argued that both should help to facilitate multidisciplinary curriculum development but also suggest learning needs to be met within unidisciplinary professional education. Concentration on team dynamics alone will not deliver the teamwork required in the new NHS. PMID- 11107018 TI - The effect of the general practice registrar year on perceived skills in palliative care in the West Midlands. AB - OBJECTIVES: To ascertain the effect of 12 months spent as a GP registrar on perceived skills in palliative care. DESIGN: A previously validated questionnaire for use with medical undergraduates is modified and used to survey perceived skills in five aspects of providing palliative care in five different scenarios at two points during the 12-month period of general practice vocational training where no specific teaching intervention is conducted. SETTING: The West Midlands. PARTICIPANTS: 210 GP registrars. RESULTS: Perceived skill ratings were seen to significantly increase during the 12-month period, but anxiety in caring for the dying did not significantly decrease. Ratings of skills were lowest when caring for a child dying with leukaemia or a young adult dying with AIDS. In addition, other important variables which had a statistically significant influence were gender and age, but interestingly not the number of previous senior house officer (SHO) posts undertaken or whether the respondent had had formal teaching on the subject in the past. CONCLUSION: It might therefore be postulated that training as a GP registrar has an important impact on the development of perceived skills in palliative care. PMID- 11107019 TI - Learning in primary care--a report. AB - A symposium on Learning in Primary Care was held in Cape Town, South Africa, as a pre-conference workshop to the 9th International Ottawa Conference on Medical Education. The aim of this report is to inform medical educationalists of important issues in learning in primary care and to stimulate further debate. Four international speakers gave presentations on their experiences in teaching and learning in primary care. Objective positive outcome measures include acquiring clinical skills equally well in general practice as in hospital, and improved history taking, physical examination and communication skills learning. Students regard the course as an essential requirement for learning and are appreciative of the wider aspect to learning provided by the community, giving a more holistic view of health. A SWOT analysis (strengths, weaknesses, opportunities and threats) of teaching and learning in primary care identified that learning in primary care is of a generalist nature and reality based, but is hampered by a lack of resources. The increased professionalization of teaching in primary care results in better training, cost containment, and improved quality of health care at community level. It is important to focus on turning threats into opportunities. Academic credibility needs to be established by conducting research on learning in primary care and developing the conceptual basis of primary care. PMID- 11107020 TI - The Objective Structured Video Exam for assessment of communication skills. AB - OBJECTIVES: (i) To design a new, quick and efficient method of assessing specific cognitive aspects of trainee clinical communication skills, to be known as the Objective Structured Video Exam (OSVE) (Study 1); (ii) to prepare a scoring scheme for markers (Study 2); and (iii) to determine reliability and evidence for validity of the OSVE (Study 3). METHODS: Study 1 describes how the exam was designed. The OSVE assesses the student's recognition and understanding of the consequences of various communication skills. In addition, the assessment taps the number of alternative skills that the student believes will be of assistance in improving the patient-doctor interaction. Study 2 outlines the scoring system that is based on a range of 50 marks. Study 3 reports inter-rater consistency and presents evidence to support the validity of the new assessment by associating the marks from 607 1st year undergraduate medical students with their performance ratings in a communication skills OSCE. SETTING: Medical school, The University of Liverpool. RESULTS: Preparation of a scoring scheme for the OSVE produced consistent marking. The reliability of the marking scheme was high (ICC=0.94). Evidence for the construct validity of the OSVE was found when a moderate predicted relationship of the OSVE to interviewing behaviour in the communication skills OSCE was shown (r=0.17, P < 0.001). CONCLUSION: A new video-based written examination (the OSVE) that is efficient and quick to administer was shown to be reliable and to demonstrate some evidence for validity. PMID- 11107021 TI - Call for papers - really good stuff 2001 really good stuff: new ideas in medical education PMID- 11107022 TI - Reports of new ideas in medical education annual, peer-reviewed collection of reports on innovative approaches to medical education PMID- 11107024 TI - Introduction PMID- 11107023 TI - Contents PMID- 11107025 TI - International review panel PMID- 11107026 TI - The visualization and modification of the body in art and medicine-- how an innovative special study module explored the humanity of medicine. PMID- 11107027 TI - The 'Critical thinking Module'--a grant proposal simulation exercise. PMID- 11107028 TI - Health promotion in medical undergraduate education--are special study modules pragmatic options? PMID- 11107029 TI - Assessment--putting it all together. PMID- 11107030 TI - Integrating the art and science of medicine at early stages of medical training. PMID- 11107031 TI - Expanding the goals of an early clinical experience for first-year medical students. PMID- 11107032 TI - Residents as teachers--a faculty development approach to programme development. PMID- 11107033 TI - Using the Myers-Briggs Type Indicator (MBTI) in the teaching of leadership skills. PMID- 11107034 TI - Teaching laboratory management in the Department of Laboratory Medicine and Pathobiology at the University of Toronto, Toronto, Canada. PMID- 11107035 TI - OSCE logistics--handheld computers replace checklists and provide automated feedback. Objective structured clinical examination. PMID- 11107036 TI - Effectiveness of PBL curricula. PMID- 11107037 TI - Benign sadness, malignant neglect. PMID- 11107038 TI - Undergraduate medical students' views on the value of dissecting. PMID- 11107039 TI - Doctors and medical students with disabilities. PMID- 11107040 TI - Computers and conference organization. PMID- 11107041 TI - Hospital clinicians' views on training as examiners for undergraduate regulatory clinical examinations. PMID- 11107042 TI - Student support programmes in Australian medical schools. PMID- 11107043 TI - Better use of abbreviations--a lesson from a stroke unit. PMID- 11107044 TI - Computer literacy in medical students. PMID- 11107045 TI - Face to face--Gordon Page. PMID- 11107046 TI - A new era for pathology international PMID- 11107047 TI - Citation index for pathology journals. PMID- 11107048 TI - Molecular characteristics of eight gastric cancer cell lines established in Japan. AB - Molecular characterization of eight gastric cancer cell lines established in Japan are summarized according to the genetic and epigenetic alterations and growth factor status. TMK-1 poorly differentiated adenocarcinoma cell line harbors mutant p53 tumor suppressor gene and rearrangement of p15MTS2. MKN-1 adenosquamous carcinoma line with mutant p53 reveals silencing of E-cadherin by promoter CpG hypermethylation. MKN-7 well-differentiated adenocarcinoma cell line has amplification of c-erbB2 oncogene and cyclin E gene. MKN-28 well differentiated adenocarcinoma cell line reveals mutations in p53 and APC tumor suppressor genes and silencing of CD44. The MKN-45 poorly differentiated adenocarcinoma cell line with wild-type p53 is characterized by homozygous deletion of p16CDKN2/MTS1/INK4A and p15MTS2, amplification of c-met oncogene and promoter mutation of E-cadherin. MKN-74 derived from moderately differentiated tubular adenocarcinoma has wild-type p53. KATO-III signet ring cell carcinoma line has genomic deletion of p53, amplification of K-sam and c-met oncogene and mutation of E-cadherin. HSC-39 signet ring cell carcinoma cell line harboring p53 missense mutation has homozygous deletion of p16CDKN2/MTS1/INK4A and p15MTS2, amplifications of c-myc, c-met, K-sam and CD44 gene and mutation in beta-catenin gene. PMID- 11107050 TI - Alternatively spliced MDM2 transcripts in human breast cancer in relation to tumor necrosis and lymph node involvement. AB - Several short forms of alternatively spliced Murine double minute 2 (MDM2) transcripts have recently been shown to correlate with high-grade malignancy in a number of human tumors. We examined the frequency of splice variants and their correlation with clinicopathological features in 60 cases of human breast cancer. Seven short forms coexpressed with wild-type mRNA were detected by nested RT-PCR. Sequencing of all the MDM2 variants demonstrated mRNA splicing which disrupted not only the conserved p53-binding domain but also, further towards the carboxy terminus, the conserved nuclear localization sequence and/or the acidic and zinc finger domains. There was no significant correlation between the coexpression of splice variants and tumor size, histologic type or hormone (estrogen and progesterone) receptor status. However, cases with spliced MDM2 transcripts tended to be of a more aggressive type with axillary lymph node involvement and extensive necrosis in the tumors. Although the functional significance of MDM2 variants remains obscure, we anticipate that these variants will be confirmed as a novel prognostic marker in human breast cancer. PMID- 11107049 TI - Correlation between genetic alterations and histopathological subtypes in bronchiolo-alveolar carcinoma and atypical adenomatous hyperplasia of the lung. AB - Bronchiolo-alveolar carcinoma (BAC) is a type of lung adenocarcinoma characterized by growth along the alveolar wall. It is divided into two subtypes: sclerosing BAC (SBAC), which has central fibrosis, and non-sclerosing BAC (NSBAC), which lacks central fibrosis. We compared the genetic alterations in these two types of BAC with those in atypical adenomatous hyperplasia (AAH). There were 39 cases of SBAC, 19 of NSBAC and 20 of AAH. To detect the loss of heterozygosity (LOH) we used the microsatellite markers D3S1234 and D3S1300 on chromosome 3p, IFNA and D9S144 on 9p, and TP53 on 17p. We also used polymerase chain reaction-SSCP analysis and direct sequencing to examine a point mutation of the p53 gene at exons 5-8. At the TP53 locus, the frequencies of LOH showed a statistical rank-difference correlation among AAH, NSBAC and SBAC. On chromosomes 3p and 9p there were no statistical differences of LOH among AAH, NSBAC and SBAC. We detected a significant statistical rank-difference correlation in the p53 mutation among AAH, NSBAC and SBAC. These findings suggest that a process of multistep carcinogenesis from AAH through NSBAC to SBAC might occur in some cases of adenocarcinoma, and LOH of 3p and 9p might be an early event of carcinogenesis, while the p53 mutation might be a later event. PMID- 11107051 TI - Clinicopathological characteristics of atypical cystic duct (ACD) of the breast: assessment of ACD as a precancerous lesion. AB - To clarify the clinicopathological features of an atypical cystic duct (ACD) as defined by Tsuchiya's criteria as a precancerous lesion of the breast, we used 200 whole mammary gland serial sections of breast cancer. Forty-four (22%) of the 200 breast cancer patients had ACD breast lesions. The frequency of patients with ACD increased in premenopausal women (P = 0.001). There was no correlation between the ACD-present group and the ACD-absent group for immunohistochemical status of the estrogen receptor (ER), progesterone receptor (PgR), p53, or c erbB2; Ki-67 labeling index of cancer tissues; size of tumor, or lymph node metastases. A number of ACD lesions displayed continuity to cancer lesions. In 500 serial sections of a paraffin-embedded tissue of a ACD case at 3 microm intervals, an apparent transition from ACD into ductal carcinoma in situ was observed. Immunohistochemical analysis using alpha-smooth muscle actin showed that myoepithelial cells of ACD stained strongly, and their nuclei and cytoplasm were thinning. In 16 of the 44 (36%) ACD-present patients, carcinoma cells stained positive for p53. Within those 16 cases, 12 cases (75%) were positive for p53 in ACD lesions. There was a significant correlation between the expression of p53 protein in malignant cells and ACD (P = 0.001). All 44 ACD lesions had no staining of c-erbB2, regardless of staining in malignant lesions. The mean Ki-67 labeling index of ACD lesions was low (0.3%), suggesting that ACD had a low proliferative rate. We suggest that ACD is the precancerous breast lesion because of a histologic continuum between ACD and malignancy, and because of p53 protein expression in ACD. PMID- 11107052 TI - Expression of cytokeratin 1, 5, 14, 19 and transforming growth factors-beta1, beta2, beta3 in osteofibrous dysplasia and adamantinoma: A possible association of transforming growth factor-beta with basal cell phenotype promotion. AB - To elucidate the precise origin and characteristics of the epithelial components of osteofibrous dysplasia (OF) and adamantinoma (AD), the expression of transforming growth factor (TGF)-beta1, beta2 and beta3 and cytokeratin (CK) subtypes were studied in five cases of AD and 18 cases of OF by immunohistochemistry. CK1 was expressed in 10 out of 18 OF cases; CK5 was expressed in one OF case; CK14 was positively stained in 10 cases of OF; CK19 was positively stained in 16 OF cases; CK1 was expressed in three out of five AD cases; CK5 was expressed in one case of AD; CK14 was positively stained in four AD cases; and CK19 was positively stained in five AD cases. In OF, TGF-beta1, beta2 and beta3 were expressed in both fibroblasts and osteoblasts. In AD, TGF beta1, beta2 and beta3 were expressed in both epithelial and fibrous components. These results suggest that epithelial components of AD and OF share epidermal characteristics, CK1, express basal cell phenotype and cytokeratins 5, 14 and 19. In addition to these epithelial characteristics, strong immunoreactivity for TGF beta poses the possibility of TGF-beta promotion of basal cell phenotype expression for the epithelial components in OF and AD. PMID- 11107053 TI - Clinicopathological features of solitary fibrous tumor of the meninges: An immunohistochemical reappraisal of cases previously diagnosed to be fibrous meningioma or hemangiopericytoma. AB - Cases of solitary fibrous tumor (SFT) of the meninges are increasingly being reported. However, the real incidence of SFT among meningeal tumors has yet to be determined. We therefore clinicopathologically re-examined 64 meningeal tumors originally diagnosed to be either fibrous meningioma (FM group, n = 46) or hemangiopericytoma (HPC group, n = 18) while paying special attention to SFT. We thus reclassified one case from the FM group (2%) and one case from the HPC group (6%) to be SFT, both of which showed diffuse CD34-immunoreactivity and dense intercellular reticulin fibers but neither epithelial membrane antigen nor S-100 protein expression. The MIB-1 staining index of these cases were 6. 2% and 3.9%, respectively. The former recurred 15 years after the initial surgery and the patient underwent a second removal of the tumor. The patient has been alive with no evidence of recurrence for 7 years after the second surgery. The latter patient has been alive with no evidence of recurrence for 3 years postoperatively. The results confirmed that the incidence of SFT among meningeal tumors is relatively low, however, because of its clinically indolent nature, a careful histochemical examination is necessary to differentiate SFT from other neoplasms with a more aggressive nature. Our findings emphasize the need to clinically recognize this lesion as a distinct entity. PMID- 11107054 TI - Synovial sarcomas of three children in the first decade: clinicopathological and molecular findings. AB - Synovial sarcoma in children below the age of 10 years is rare. We report on three cases of synovial sarcoma which were diagnosed in three children aged 3, 8 and 8 years, respectively. These tumors were located in the hip of the 8-year old, the foot of the 3-year-old, and the elbow of the other 8-year-old. Histologically, one tumor was a biphasic synovial sarcoma, and the other two, which had been initially diagnosed as infantile fibrosarcoma, were of the monophasic fibrous type. In the three cases, a reverse transcription-polymerase chain reaction (RT-PCR) using ribonucleic acid extracted from formalin-fixed, paraffin-embedded tissues detected SYT-SSX1 fusion gene transcripts resulting from translocation t(X;18)(p11.2;q11.2), which is specific for synovial sarcoma. ETV6-NTRK3 fusion gene transcripts that result from t(12;15)(p13;q25), which is characteristic of congenital/infantile fibrosarcoma, were not demonstrated. In conclusion, other pediatric soft tissue sarcomas, such as congenital/infantile fibrosarcoma, spindle cell rhabdomyosarcoma, leiomyosarcoma and malignant peripheral nerve sheath tumor, should be distinguished from synovial sarcoma in children, especially the monophasic fibrous type. RT-PCR analysis is a useful approach to the final diagnosis of synovial sarcoma arising at such an early age. PMID- 11107055 TI - The expression of a novel natural killer inhibitory molecule, Cho-1, on the chorionic cytotrophoblast cells of successful pregnancy, but not of spontaneous abortion. AB - The regulatory mechanism of the recognition and cytotoxicity by natural killer (NK) cells in placental tissue remains unclarified. Previous reports indicated that monoclonal antibody Cho-1-defined molecule (Cho-1 molecule) may act as the negative regulator in the cytotoxicity by human NK cells. The Cho-1 molecule is composed of non-covalently associated cell surface molecules of approximately 200 kDa and 40 kDa. In the present study we analyzed the expression of this novel molecule in extravillous cytotrophoblast cells, which are presumed to be exposed to the cytotoxic action by maternal NK cells, from clinical cases of successful pregnancy and spontaneous abortion. By using monoclonal antibody Cho-1, our immunohistochemical data indicated that the Cho-1 molecule is clearly expressed in the cytotrophoblast cells of the early phase of successful pregnancy, but only weakly expressed in those from spontaneous abortion. The cytotrophoblast cells in the late phase (9-10 months) of pregnancy also expressed this molecule. Fluorescence-activated cell sorter analysis also showed that it is expressed on the cytotrophoblast cell surface of successful pregnancy but not on that of spontaneous abortion, suggesting that Cho-1 antigen may act as a negative regulator of the cytotoxicity by NK cells in successful pregnancy of the fetus. PMID- 11107056 TI - Embryonal sarcoma of adult and pediatric kidneys: report of a case with localized submucosal invasion of the renal pelvis and long-term survival. AB - A case of cystic embryonal sarcoma of the kidney (CESK) with a rapidly fatal outcome was recently reported.1 Here, we report another case of a 12-year-old boy with a localized but ill-defined submucosal lesion of CESK in the right renal pelvis. The tumor consisted principally of small mesenchymal cells with oval to spindle nuclei and scanty cytoplasm, infiltrating in dense arrangements. Two growth patterns were distinguished in the tumor cells: (i) a diffuse infiltrating pattern without an epithelial component; and (ii) a foliated pattern with an epithelial lining over the surface. Foci of the diffuse pattern predominated over those that were lobular, infiltrating superficial layers of renal sinuses and along pyramids, in both of which remarkable intravenous invasion was evident. Foci of the foliated pattern invaded deeper portions of a few sinuses and frequently penetrated into their veins, producing together with their epithelial lining a characteristic foliated structure. Lining epithelial cells around lobular foci often appeared hob-nailed or eosinophilic in the cytoplasm. Despite the remarkable intravenous encroachment, the patient has remained well without a recurrence for more than 26 years after a simple nephrectomy. The present case report expands our understanding of the biological nature of CESK. PMID- 11107057 TI - Leiomyosarcoma of the pulmonary vein. AB - A case of a 74-year-old man with leiomyosarcoma of the pulmonary vein is reported. The patient felt transient chest oppression while playing golf 1 week before he visited a clinic with a common cold. He underwent an ultrasonographic examination of the heart, which showed a mass lesion in the left atrium. The preoperative clinical diagnosis was myxoma of the left atrium. Cardiac surgery revealed the mass to be a leiomyosarcoma, probably extending from the left inferior pulmonary vein. The patient underwent a left lower lobectomy of the lung, and the tumor was confirmed to have originated from the wall of the left inferior pulmonary vein. Although the patient had a metastatic lesion in the right axillary lymph node 11 months later, which was excised, he remained free of disease 14 months after the initial operation. Histologically, the tumors were composed of pleomorphic cells with bizarre nuclei and spindle cells with blunt ended nuclei with 1-4 mitotic figures in 10 high power fields. Immunohistologically, the tumor cells were positive for alpha-smooth muscle actin and desmin. We reviewed 17 cases of leiomyosarcoma of the pulmonary vein (six males and 11 females with a mean age of 50 years in each group). The present case was the oldest in age and to our knowledge was the first reported case with metastasis in a distant lymph node. PMID- 11107058 TI - Alpha-fetoprotein producing uterine corpus carcinoma: A hepatoid adenocarcinoma of the endometrium. AB - A case of alpha-fetoprotein (AFP) producing endometrial carcinoma in a 60-year old Japanese woman is presented. The patient complained of abnormal vaginal bleeding of 10 days' duration. On admission a uterine corpus mass and high serum AFP concentration (31950 ng/mL) was noted. There was no tumorous lesion in any other organ radiographically and endoscopically. Histologically, the biopsy specimen taken from the uterine mass showed a poorly differentiated endometrial carcinoma and a radical hysterectomy was subsequently performed. The postoperative serum AFP value transiently decreased with chemotherapy, however, lung metastases were found and the patient died 12 months following surgery. The resected uterus had a necrotic tumor, 6 x 5 x 4 cm in size, filling the endometrial cavity, characterized by exophytic growth with infiltration in the myometrium. Histologically, the tumor was composed of the main medullary carcinoma area with microcysts and admixed small areas of well-differentiated endometrioid adenocarcinoma, accompanied by a smooth transition with one another. In both the areas, the tumor cells had immunoreactive AFP, alpha-1-antitripsin, albumin, transferrin, carcinoembryonic antigen, CA19-9, and epithelial membrane antigen. There was no histologic evidence for a germ cell tumor. Based on these findings, this uterine corpus tumor was regarded as hepatoid variant of endometrial carcinoma. Although the histogenesis remains controversial, we assume the hypothesis that the tumor may arise in the endometrium per se in association with abnormal differentiation of muellerian duct elements. PMID- 11107059 TI - Granzyme B-positive primary gastric T-cell lymphoma: gastric T-cell lymphoma with the possibility of extrathymic T cell origin. AB - A case of primary gastric T-cell lymphoma, which was positive for granzyme B, is reported. The patient was a 47-year-old Japanese female who complained of a dull upper abdominal pain. Radiographic and endoscopic examinations revealed an ulcerative infiltrative lesion in her stomach. Following the confirmation of a high-grade malignant lymphoma, a distal gastrectomy with regional lymph nodal dissection was performed. The histology of the gastric lesion revealed a malignant lymphoma of the diffuse pleomorphic type without lymph nodal involvement. Immunohistochemistry revealed that the tumor cells were positive for LCA, CD3, TIA-1 and granzyme B, but were negative for CD4, CD8, CD56, CD30, L-26, EMA, TCR alpha/beta and TCR gamma/delta. Because the tumor cells showed T cell nature with cytotoxic activity proved by TIA-1 and granzyme B, and without evidence of further maturation of T cell, a malignant lymphoma originating from extrathymic-derived T cells was suggested. PMID- 11107060 TI - A case of cytophagic histiocytic panniculitis associated with exertional rhabdomyolysis. AB - An 18-year-old man who suffered from panniculitis involving the entire left lower limb after exertional rhabdomyolysis is reported. A high fever (>39 degrees C) and leukocytosis (>20,000/microL) persisted for 1 week, and his general status deteriorated rapidly into pre-disseminated intravascular coagulation, complicated by pleural effusion and prolonged clotting time. His condition was dramatically improved by steroid pulse therapy and he has remained in good health for the 20 months since discharge. Histologic examination of subcutaneous tissue from the swollen left lower limb revealed pleomorphic small, medium or large lymphocytes, macrophages and neutrophils infiltrating the edematous subcutaneous adipose tissue in a lobular panniculitis-like pattern. The majority of inflammatory cells were T lymphocytes, with equal proportions of CD4+ and CD8+ cells. As polymerase chain reaction did not show bands suggesting T cell receptor gamma gene rearrangement, the proliferation of T lymphocytes was considered to be polyclonal. The T lymphocytes also expressed Fas ligand, suggesting the involvement of Fas-mediated cytotoxicity. This case may represent a new category of cytophagic histiocytic panniculitis induced by exertional rhabdomyolysis. PMID- 11107061 TI - Molecular mechanism of monocyte predominant infiltration in chronic inflammation: mediation by a novel monocyte chemotactic factor, S19 ribosomal protein dimer. AB - A novel monocyte chemotactic factor, a cross-linked homodimer of S19 ribosomal protein (RP S19) was initially isolated from a rheumatoid arthritis synovial lesion. The RP S19 dimer causes the monocyte specific chemotaxis in vitro and the monocyte predominant infiltration in vivo, via its agonistic and antagonistic effects on the C5a receptors of monocytes and polymorphonuclear leukocytes, respectively. The agonistic effect is attributed to the similarity of regional structures between RP S19 and C5a, the complement C5-derived leukocyte chemotactic factor, although overall homology of the amino acid sequence between these molecules is only 4%. The antagonistic effect depends upon the C-terminal portion of RP S19. The RP S19 dimer is produced and released by apoptotic cells, and this dimer recruits monocytes from the circulation to the apoptotic lesion. The infiltrated monocytes/macrophages engulf the apoptotic cells, translocate to regional lymph nodes via lymphatics and present the antigenic information of the apoptotic cells to the T cell repertoire. In this manner, the apoptotic cell clearance system connects to the acquired immune system. The innate and acquired immune mechanisms, mediated by the RP S19 dimer, participate in the pathology of inveterate chronic inflammation such as rheumatoid arthritis. PMID- 11107062 TI - Chromophobe renal cell carcinoma: clinical, pathological and molecular biological aspects. AB - Chromophobe renal cell carcinoma (RCC), a newly established subtype of renal neoplasm, is composed of tumor cells with characteristically cloudy, weakly eosinophilic and reticular cytoplasm. The tumor should be distinguished from the common clear cell RCC, because of the unique clinicopathological and molecular biological features. The tumor does not show gender bias. Patient ages are similar to those of clear cell RCC, but might occur in the 20- to 40-year-old age group. Grossly, the tumor tends to be beige in color, which is different from the yellowish color of common RCC. Electron microscopy and immunohistochemistry indicate the intercalated cell of the collecting duct as the cellular origin. Cytogenetic study shows non-random multiple chromosome loss, with mitochondrial DNA rearrangement. Alteration of the von Hippel-Lindau (VHL) gene, a cancer suppressor gene relating with clear cell RCC, has not yet been observed. In order to adopt the most appropriate treatment, including gene therapy, recognition and correct pathological diagnosis of chromophobe RCC are extremely important. PMID- 11107063 TI - Effects of subtotal resection of the fundus on development of intestinal metaplasia induced by X-ray irradiation in Donryu rats. AB - Eight-week-old male Crj:Donryu rats underwent subtotal resection of the fundus and X-ray irradiation. Six months later the animals were autopsied and examined for intestinal metaplasia. The numbers of alkaline phosphatase-positive foci with the two treatments in combination were significantly increased, compared to the operation alone and non-treatment groups. Histologically assessed intestinal metaplasia was also increased in the combined treatment group. In conclusion, subtotal resection of the fundus combined with X-ray irradiation is an effective induction protocol for intestinal metaplasia. PMID- 11107064 TI - Abnormal distribution of collagen type IV in extrahepatic bile duct carcinoma. AB - The present study investigated the pathogenesis of desmoplastic stroma formation, which is characteristic of most bile duct carcinomas and other scirrhous carcinomas. Using immunohistochemical analysis, the expression of collagen types I and IV, laminin and TGF-beta1 was examined in human extrahepatic bile duct carcinoma and compared with gastric and colon carcinoma. In addition to delineating the basement membranes of carcinoma nests and blood vessels, collagen type IV was present along the thick bundles of collagenous fibers in the stroma of extrahepatic bile duct carcinoma and scirrhous gastric carcinoma. The immunoreactivity of collagen type IV was strong in the adjacent or surrounding interstitium of tumor cell nests, but was absent or weak in older, more central portions of the tumor that contained sclerotic collagen. In situ hybridization demonstrated active expression of collagen alpha1(IV) mRNA in extrahepatic bile duct carcinoma and scirrhous gastric carcinoma cells. These results suggest that, although collagen type IV is typically a component of the basement membrane, it is expressed in the interstitial stroma of extrahepatic bile duct carcinoma and scirrhous gastric carcinoma where it may play a role in desmoplastic stroma formation. PMID- 11107065 TI - Correlation between morphological heterogeneity and genetic alteration within one tumor in adenocarcinomas of the lung. AB - In some human cancers, multistep carcinogenesis has been advocated on the basis of morphological and genetic analysis. In adenocarcinoma of the lung, a carcinogenetic process from atypical adenomatous hyperplasia (AAH) to bronchiolo alveolar carcinoma (BAC) and/or more malignant adenocarcinoma has been recently suggested. In the present study, we selected 13 lung tumors which had AAH-like or BAC-like areas at the periphery, and poorly differentiated areas at other sites, and examined their loss of heterozygosity (LOH) on chromosome 3p, 9p and 17p and point mutation of the p53 gene. A heterogeneous pattern of LOH and/or point mutation of the p53 gene was detected in five of 13 cases, and genetic alterations were frequent in the areas of poorer differentiation. These findings suggest that some adenocarcinomas of the lung occur through multistep carcinogenesis. PMID- 11107066 TI - Intrathyroidal branchial cleft-like cyst in chronic thyroiditis. AB - An extremely rare case of intrathyroidal branchial cleft-like cyst is reported. A 71-year-old man complained of a growing mass in the right lateral neck. A cystic mass in the upper lobe of the right thyroid was demonstrated by ultrasonography and computed tomography. The surgical specimen revealed a cystic mass with dense fibrous capsule, 22 x 20 x 10 mm in size. Microscopically, the cyst walls and the surrounding thyroid tissue contained severe lymphoid cell infiltration with lymphoid follicle. Squamous epithelium lined the cyst wall. Immunohistochemically, squamous epithelium was positive for keratin, cytokeratin 19, carcinoembryonic antigen, and epithelial membrane antigen, but negative for calcitonin and chromogranin A. The patient is currently well with no evidence or recurrence for 43 months. PMID- 11107067 TI - Supradiaphragmatic ectopic adrenocorticotropic hormone-secreting adenoma. AB - A 22-year-old woman with Cushing's syndrome, caused by an extremely rare suprasellar ectopic pituitary adenoma, is presented. Magnetic resonance imaging and computed tomography revealed a well-circumscribed mass in the right suprasellar region. Endocrinological tests showed elevated s-adrenocorticotropic hormone level and hypercortisolemia. The tumor was totally removed by right subfrontal approach. At the time of the operation, the tumor was in continuity with the distal pituitary stalk but not with the pituitary gland. The diaphragma sellae was intact. Histologic diagnosis of the tumor specimen was confirmed as a pituitary adenoma. After surgical removal of the tumor, continued improvement in the patient's laboratory results and disappearance of her endocrine symptoms strongly indicated the absence of adenoma cells in the pituitary gland or stalk. Six years post-surgery, there was no evidence of recurrence in the patient's clinical and laboratory examination. This tumor probably originated from aberrant anterior pituitary cells of the pituitary stalk. PMID- 11107068 TI - Oncocytic mucoepidermoid carcinoma of the parotid gland: report of a case with DNA ploidy analysis and review of the literature. AB - A 44-year-old female presented with a painful mass in the left parotid gland. Histologic examination revealed the characteristic picture of oncocytic mucoepidermoid carcinoma (OMEC) composed mainly of sheets of oncocytic cells with uniform nuclei and eosinophilic cytoplasm, focally smaller epidermoid cells surrounding poorly formed glandular spaces, and a few cystic structures lined by well-differentiated mucous cells with intracytoplasmic mucin. Immunohistochemical staining with antimitochondrial antibody showed granular cytoplasmic positivity in oncocytic cells. The resulting histogram for DNA ploidy analysis was of diploid type. OMEC of the parotid gland is a recently described rare neoplasm. Only six cases have been previously reported in the literature. For an accurate approach in the management of patients, OMEC should be considered in the differential diagnosis of oncocytic lesions of the parotid gland, most of which are benign. PMID- 11107069 TI - Pulmonary adenocarcinoma with heterotopic bone formation. AB - Adenocarcinoma of the lung is a common malignancy. Frequently, this tumor can cause calcification within a primary tumor. However, an extremely rare occurrence in lung carcinomas is ossification within a primary tumor, and to our knowledge only three cases have been reported. We report a case of pulmonary adenocarcinoma with ossification, and discuss the pathogenesis of intratumoral ossification with a review of the literature. PMID- 11107070 TI - Primary small cell (oat cell) carcinoma of the breast: report of a case and review of the literature. AB - A case of primary small cell (oat cell) carcinoma of the breast in a 41-year-old woman is presented. The patient was alive and well without disease 16 months after modified radical mastectomy and subsequent chemotherapy. The tumor cells revealed morphologic similarity to oat cell carcinoma of the lung and immunohistochemical expression of neuroendocrine markers. In ultrastructural examination, the tumor cells had neurosecretory granules. Review of nine previously reported cases and this case of primary small cell carcinoma of the breast has revealed that this type of tumor shows prominent vascular invasion, frequent lymph node metastasis, infrequent expression of estrogen receptor, and also very poor prognosis. Immunohistochemical study for the c-kit proto-oncogene product, which has been reported to be a specific marker for pulmonary small cell carcinoma, demonstrated positive reactivity in approximately 80% of the tumor cells of this case, which is the first report according to our knowledge. The expression of c-kit might be some aid to the diagnosis of primary small cell carcinoma of the breast. PMID- 11107071 TI - Sarcomatoid hepatocellular carcinoma with chondroid variant: case report with immunohistochemical findings. AB - A case of sarcomatoid hepatocellular carcinoma (HCC) with a chondroid variant in a 72-year-old woman is reported. Histologically, the tumor consisted of typical HCC of adult type, with an adenocarcinoma-like structure and a sarcomatoid component containing multinodular chondroid foci. Transition from carcinoma to sarcomatoid component was observed. The cells containing mucus were extremely few in adenocarcinoma-like structures. Immunohistochemically, other than typical HCC, some of the chondroid cells, adenocarcinoma-like cells and sarcomatoid cells were positive for alpha-fetoprotein. PMID- 11107072 TI - Histopathological characterization of aortic intimal sarcoma with multiple tumor emboli. AB - A case of aortic intimal sarcoma with multiple tumor emboli and distal metastasis is reported. All metastasis (adrenal, spleen) were via the arteries. This case also had independent lung cancer. Macroscopically, the aortic tumor did not form a bulged mass, but had linear ulceration with abundant mural thrombi. Poorly cohesive large atypical cells were seen in the intima of the abdominal aorta without invasion into the media. Tumor cells were disseminated into the mural thrombi on the aorta and embolized its branches. In the metastatic tumor or tumor emboli of the distal artery, there were not only large atypical cells, but also the foci of spindle-shaped cells or epithelioid differentiation. Tumor cells in the aorta were immunohistochemically positive for only vimentin. Muscle-specific actin was positive focally for spindle-shaped cells of tumor emboli and metastatic tumors. Furthermore, cytokeratin-positive cells were scatteredly seen. All tumor cells were negative for factor VIII and did not have a histologic or phenotypic analogy with lung cancer. The primary intimal sarcoma in the present case was of undifferentiated non-endothelial intimal stromal cell origin, and may have had multipotential for differentiation. Investigation of the metastatic site was useful for recognizing the features of this tumor. PMID- 11107073 TI - Longevity possible in metastatic duodenal somatostatinoma. PMID- 11107074 TI - Reply PMID- 11107075 TI - Frequency of postoperative carotid duplex surveillance and type of closure: results from a randomized trial. AB - BACKGROUND/PURPOSE: In several nonrandomized studies investigators have reported on the value of postoperative carotid duplex surveillance (PCDS) with mixed results; however the type of closure was not analyzed in these studies. In this study we analyze the frequency and timing of postoperative carotid duplex ultrasound scanning according to the type of closure from a randomized carotid endarterectomy (CEA) trial comparing primary closure (PC) versus patching. PATIENT POPULATION AND METHODS: We randomized 399 CEAs into 135 PCs, 134 polytetrafluoroethylene (PTFE) patch closures, and 130 vein patch closures (VPCs) with a mean follow-up of 47 months. PCDS was done at 1, 6, and 12 months and every year thereafter (a mean of 4.0 studies per artery). Kaplan-Meier analysis was used to estimate the rate of > or = 80% restenosis over time and the time frame of progression from < 50%, to 50%-79% and > or = 80% stenosis. RESULTS: Restenoses of > or =80% developed in 24 (21%) arteries with PC and nine (4%) with patching. Kaplan-Meier estimate of freedom of > or = 80% restenosis at 1, 2, 3, 4, and 5 years was 92%, 83%, 80%, 76%, and 68% for PC, respectively, and 100%, 99%, 98%, 98%, and 91% for patching, respectively, (P <.01). Of 56 arteries with 20% to 50% restenosis, two of 28 patch closures and 10 of 28 PCs progressed to 50% to < 80% restenosis (P =.02); none of the patch closures and six of 28 PCs progressed to > or =80% (P =.03). In PCs, the median time to progression from <50% to 50%-79%, < 50% to > or =80%, and 50%-79% to > or = 80% was 42, 46, and 7 months, respectively. Of the 24 arteries with > or =80% restenosis in PC, 10 were symptomatic. Thus, assuming th symptomatic restenosis would have undergone duplex scan examinations regardless, there were 14 asymptomatic arteries (12%) that could have been detected only with PCDS (estimated cost, $139, 200), and those patients would have been candidates for redo CEA. Of the 9 arteries (3 PTFE closures and 6 VPCs) with > or =80% restenosis with patch closures, 6 asymptomatic (4 VPCs and 2 PTFE closures) arteries (3%) could have been detected with PCDS. In patients with normal duplex scan findings at the first 6 months, only four (2%) of 222 patched arteries (two asymptomatic) developed > or = 80% restenosis versus five (38%) of 13 in patients with abnormal duplex scan examination findings (P<.001). CONCLUSIONS: PCDS is beneficial in patients with PC, but is less beneficial in patients with patch closure. PCDS examinations at 6 months and at 1- to 2-year intervals for several years after PC are adequate. For patients with patching, a 6-month postoperative duplex scan examination with normal results is adequate. PMID- 11107076 TI - Eversion endarterectomy of the carotid artery: technical considerations and recurrent stenoses. AB - PURPOSE: The purpose of this study was to examine the characteristics of residual and recurrent lesions after eversion endarterectomy of the carotid artery (E-CE) and compare these results with those following endarterectomy and patch closure (CE-P). METHODS: We reviewed 274 patients who underwent carotid endarterectomy in 1998 with electroencephalographic monitoring, general anesthesia, completion duplex scan, and 1-year follow-up. CE-P was preferred for patients who required temporary shunting. In the E-CE group an additional proximal 2-cm arteriotomy was made in the common carotid artery (CCA) in 79 patients, a longer arteriotomy was made for extensive involvement of the CCA in 14 patients, and the internal carotid artery was advanced proximally as a patch for the CCA arteriotomy closure in 14 patients. Stenoses of > 50% that were present at 1 month were considered residual, and those of > 50% that were present at 1 year but not at 1 month were considered recurrent. RESULTS: There were five (1.8%) postoperative strokes (four after CE-P and one after E-CE, P = not significant). At 30 days there were 28 patients (10.2%) with residual stenoses > 50% (11 patients [10.2%] in the E-CE group and 17 patients [10.1%] in the CE-P group; P = not significant). The incidence of recurrent lesions of more than 50% was similar (4.6% for E-CE vs 4.7% for CE-P). CONCLUSION: The pattern of residual lesions and recurrent stenoses differs with each technique of endarterectomy. Proximal stenoses are more common after E-CE, and distal stenoses are more common after CE-P at both 1 month and 1 year. The frequency of proximal lesions is reduced in E-CE when either the internal carotid artery is advanced proximally onto the CCA or a long CCA arteriotomy is made. Distal recurrences do not seem to be a problem after eversion endarterectomy. PMID- 11107077 TI - Immediate reexploration for the perioperative neurologic event after carotid endarterectomy: is it worthwhile? AB - PURPOSE: When managing a new neurologic deficit after carotid endarterectomy (CEA), the surgeon is often preoccupied with determining the cause of the problem, requesting diagnostics tests, and deciding whether the patient should be surgically reexplored. The goal of this study was to analyze a series of perioperative neurologic events and to determine if careful analysis of their timing and mechanisms can predict which cases are likely to improve with reoperation. METHODS: A review of 2024 CEAs performed from 1985 to 1997 revealed 38 patients who manifested a neurologic deficit in the perioperative period (1.9%). These cases form the focus of this analysis. RESULTS: The causes of the events included intraoperative clamping ischemia in 5 patients (13.2%); thromboembolic events in 24 (63.2%); intracerebral hemorrhage in 5 (13.2%); and deficits unrelated to the operated artery in 4 (10.5%). Neurologic events manifesting in the first 24 hours after surgery were significantly more likely to be caused by thromboembolic events than by other causes of stroke (88.0% vs. 12.0%, P<.002); deficits manifesting after the first 24 hours were significantly more likely to be related to other causes. Of 25 deficits manifesting in the first 24 hours after surgery, 18 underwent immediate surgical reexploration. Intraluminal thrombus was noted in 15 of the 18 reexplorations (83. 3%); any technical defects were corrected. After the 18 reexplorations, in 12 cases there was either complete resolution of or significant improvement in the neurologic deficit that had been present (66.7%). CONCLUSIONS: Careful analysis of the timing and presentation of perioperative neurologic events after CEA can predict which cases are likely to improve with reoperation. Neurologic deficits that present during the first 24 hours after CEA are likely to be related to intraluminal thrombus formation and embolization. Unless another etiology for stroke has clearly been established, we think immediate reexploration of the artery without other confirmatory tests is mandatory to remove the embolic source and correct any technical problems. This will likely improve the neurologic outcome in these patients, because an uncorrected situation would lead to continued embolization and compromise. PMID- 11107078 TI - Pedal branch artery bypass: a viable limb salvage option. AB - PURPOSE: We reviewed our experience with pedal branch artery (PBA) bypass to confirm the role of these target arteries for limb salvage and to identify patient and technical factors that may be associated with graft patency and limb salvage. METHODS: In this retrospective study we analyzed 24 vein grafts to PBAs performed from 1988 to 1998 for limb salvage in 23 patients who had no suitable tibial, peroneal, or dorsal pedal target arteries. These PBA grafts were compared with 133 perimalleolar posterior tibial, defined at or below the ankle, or dorsalis pedis bypass grafts performed contemporaneously; the Kaplan-Meier life table was used in the analysis of graft patency and limb salvage. Life table analyses and logistic regression analysis of prognostic patient variables were also performed. RESULTS: The PBA bypass represented 3% of infrainguinal revascularizations for chronic critical limb ischemia at our institution over the study period. Patients who received PBA bypasses were more likely to be male (92% vs. 69%, P =.02) with lower incidences of overt coronary artery disease (33% vs. 50%, P =.12) and stroke (0% vs 15%, P =.04), and a higher incidence of end-stage renal disease (21% vs 8%, P =.06) than those undergoing perimalleolar bypass. Seventeen percent of PBA bypasses were performed with the anterior lateral malleolar artery, a vessel not previously described as a common bypass target. Two-year primary patency and limb salvage for PBA versus perimalleolar bypass was 70% versus 80% (P =.16) and 78% versus 91% (P = .28), respectively. Patency and limb salvage rates were no different in bypasses with above-knee or below-knee inflow arteries. CONCLUSION: An autogenous vein bypass to the PBA, though rarely required, provides acceptable primary patency and limb salvage when compared with perimalleolar tibial artery bypass when no suitable, more proximal target arteries are available. The PBA bypass should be considered before major amputation is undertaken. PMID- 11107079 TI - A comparative study of alternative conduits for lower extremity revascularization: all-autogenous conduit versus prosthetic grafts. AB - PURPOSE: In the absence of an adequate ipsilateral greater saphenous vein, various alternative conduits have been used for the performance of lower extremity revascularization. In this study we compared the effectiveness of all autogenous arm vein bypass grafts with that of prosthetic grafts. METHODS: Seven hundred forty lower extremity revascularization procedures (506 arm vein, 234 prosthetic) performed between 1990 and 1999 were followed prospectively by means of a computerized vascular registry. RESULTS: Bypass graft configurations were femoro-above-knee-popliteal (26 arm vein, 100 prosthetic); femoro-below-knee popliteal (38 arm vein, 29 prosthetic); femorotibial (174 arm vein, 55 prosthetic); femoropedal (23 arm vein, 2 prosthetic); popliteotibial/pedal (101 arm vein, 1 prosthetic); and extension "jump" grafts (144 arm vein, 47 prosthetic). The indications for surgery were limb salvage (98.0% arm vein, 89.7% prosthetic) and disabling claudication (2.0% arm vein, 10.3% prosthetic). The mean follow-up was 23.4 months (range, 1 month-7.4 years). Overall patient survival at 4 years was 54% (arm vein) and 69% (prosthetic). Cumulative patency varied with graft configuration. The 1-year primary patency rates for femorotibial grafts were 81.6% +/- 3.6% (arm vein) and 58.0% +/- 8.4% (prosthetic); the 3-year rates were 68.3% +/- 6.1% (arm vein) and 41.1% +/- 9.8% (prosthetic) (P<.01). The 1-year limb salvage rates for femorotibial grafts were 91.1% +/- 2.8% (arm vein) and 69.1% +/- 8. 8% (prosthetic); the 3-year rates were 81.4% +/- 5.6% (arm vein) and 63.2% +/- 10.3% (prosthetic) (P =.02). The 1-year primary patency rates for femoro-below-knee-popliteal grafts were 92.9% +/- 5.1% (arm vein) and 83.4% +/- 8.0% (prosthetic); the 3-year rates were 72.8% +/- 10.1% (arm vein) and 55.5% +/- 12.1% (prosthetic) (P=.05). The 1-year limb salvage rates for femoro-below-knee-popliteal grafts were 100% (arm vein) and 91.3% +/- 7.0% (prosthetic); the 3-year rates were 94.7% +/- 7.3% (arm vein) and 75.3% +/- 14.6% (prosthetic) (P = NS). CONCLUSION: In this study autogenous arm vein grafts demonstrated increased patency and limb salvage, compared with prosthetic grafts. These increases achieved statistical significance in the femoro-below-knee popliteal and femorotibial configurations. An effort to use an all-autogenous vein conduit is justified on the basis of these results; however, if no autogenous vein is available, prosthetic grafts provide a reasonable alternative to primary amputation. PMID- 11107080 TI - Two decades of abdominal aortic aneurysm repair: have we made any progress? AB - PURPOSE: Over the past 20 years, there have been numerous advances in our ability to detect and to treat abdominal aortic aneurysms (AAAs). We hypothesized that these advances would lead to (1) an increase in the rate of elective repair and a decrease in the incidence of ruptured AAA (rAAA) and (2) a decrease in operative deaths for both elective AAA (eAAA) and rAAA. METHODS: To test these hypotheses, we investigated the incidence and outcomes of eAAA and rAAA surgery between 1979 and 1997, using the National Hospital Discharge Survey. This data set is a randomized, stratified sample representing discharges from the nation's acute care, nonfederally funded hospitals. Codes from the International Classification of Diseases, Ninth Revision were used to identify our study population. RESULTS: Over the past 19 years, there has been no change in the incidence rate of eAAA repair (range, 44.1-77.9 per 100,000). Moreover, the incidence of rAAAs presenting to the nation's hospitals has not changed (range, 6.6-16.3 per 100,000). There has been no consistent improvement over time in operative deaths associated with either eAAA or rAAA repair (average rates over the study period: eAAA, 5.6%; rAAA, 45.7%). Significant predictors of death from eAAA in patients included an age older than 80 years, African American race, congestive heart failure (CHF), and diabetes (P<.0001 for all). Significant predictors of death from rAAA in patients included age older than 70 years, African American race, female sex, renal failure, and a hospital bed size more than 500 (P<.05 for all). CONCLUSION: On a national level, over the past 19 years, our ability to identify and to treat patients with AAA has not improved. Advances in technology and critical care have not affected outcome. Regionalization of care, screening of high-risk populations, and endovascular repair are strategies that might allow further improvement in the outcome of patients with aneurysmal disease. PMID- 11107081 TI - Management of synchronous renal neoplasm and abdominal aortic aneurysm. AB - OBJECTIVES: Renal neoplasm (RN) and abdominal aortic aneurysm (AAA) are occasionally discovered concurrently. The approach to synchronous malignancy and aortic aneurysm is controversial. METHODS: Between 1981 and 1999, concurrent RN and AAA were diagnosed in 50 patients at the Cleveland Clinic Foundation. Twenty three patients were managed conservatively because of small asymptomatic AAA or metastatic disease; these patients were excluded from the study. The remaining 27 patients underwent operative management of both entities with a staged or simultaneous approach, and they form the basis of this article. RESULTS: AAA diameter ranged from 4.8 to 13 cm (mean, 6.0+/-1.8 cm). RNs were managed with radical nephrectomy in 11 patients (41%), partial nephrectomy in 10 patients (37%), or both in 6 patients with bilateral renal tumors (22%). The AAA repair was performed at the time of the urologic procedure in 11 patients (41%), before the urologic procedure in 13 patients (48%), or after the urologic procedure in 3 patients (11%). The AAA was addressed with open surgical repair in 24 patients (89%); recently, three patients (11%) underwent endovascular repair of the aneurysm and staged partial nephrectomy. The incidence of major perioperative complications was 23% (6 patients). Acute renal failure was the most common complication (3 [11%]) followed by acute respiratory failure (2 [7.4%]), pulmonary embolism (1 [3.7%]), and stroke (1 [3.7%]). At the mean follow-up of 57 months, there were no graft infections reported. The 5-year overall and cancer specific survival rates were 62% and 81%, respectively. There was a significant difference in 5-year cancer-specific survival when comparing patients managed simultaneously versus staged (80% versus 35%, P =.007). CONCLUSIONS: The concurrent presentation of RN and AAA should not discourage one from treating both entities simultaneously because long-term survival is common. Endovascular repair of AAA holds promise as an attractive strategy in these complex patients. PMID- 11107082 TI - Adenoviral-mediated uteroglobin gene transfer inhibits neointimal hyperplasia after balloon injury in the rat carotid artery. AB - OBJECTIVE: Uteroglobin is a protein with potent anti-inflammatory and immunomodulatory effects. We hypothesize that induction of uteroglobin expression in the artery wall by local adenoviral gene transfer will decrease neointimal hyperplasia in the rat carotid artery after balloon injury. METHODS: Seven male Sprague-Dawley rats underwent balloon injury of the common carotid artery. After the injury, with flow occluded, the artery was instilled with 50 microL of the adenoviral vector encoding uteroglobin gene (Ad.UG) at a concentration of 1.35 x 10(11) pfu/mL (n = 7) or 0.68 x 10(11) pfu/mL (n = 7) (n = 7). Control animals were similarly treated: either an adenovirus encoding for beta-galactosidase gene (Ad.LacZ) at 1 x 10(11) pfu/mL (n = 7) or the phosphate-buffered saline (PBS) vehicle (n = 6) was used. The solution was allowed to dwell for 20 minutes. The rats were humanely killed after 14 days by perfusion fixation, and the carotid arteries were sectioned for analysis with computerized planimetry. The intima media area ratios were calculated for each artery and compared with analysis of variance with Bonferroni/Dunn post hoc testing. One additional rat from the PBS, Ad.LacZ, and Ad.UG (1.35 x 10(11) pfu/mL) groups was humanely killed 4 days after treatment for carotid artery protein extraction and Western blotting. RESULTS: Uteroglobin protein production was confirmed in the Ad.UG-treated arteries with Western blotting. Morphometric analysis showed that the Ad.UG group at 1.35 x 10(11) pfu/mL had a significantly lower intima-media area ratio than both the Ad.LacZ (P =.002) and PBS (P =.004) controls. The Ad.UG group at 0.68 x 10(11) pfu/mL was also significantly different from the Ad. LacZ (P =.003) and PBS (P =.006) controls. There was no statistical difference between the two control groups or between the two Ad.UG groups. CONCLUSION: Adenoviral gene transfer of uteroglobin, delivered intraluminally after arterial injury causes the production of uteroglobin protein and has an inhibitory effect on neointimal accumulation in the rat model. PMID- 11107083 TI - A novel insulin mimetic without a proliferative effect on vascular smooth muscle cells. AB - BACKGROUND: Insulin induces vascular smooth muscle cell (VSMC) proliferation, which is an important step in the atherosclerotic process. Recently, a nonpeptidyl fungal metabolite originally referred to as L-783,281, but also known as demethylasterriquinone B-1 (DMAQB-1), was found to have hypoglycemic activity in diabetic mice through interaction with the intracellular beta subunit of the insulin receptor. This study was designed to determine whether DMAQB-1 has an insulin-like proliferative effect on human infragenicular VSMCs. METHODS: Human infragenicular VSMCs were isolated from diabetic patients undergoing amputations. DMAQB-1 cell culture dose response was measured in both serum-free media and media with 1% fetal bovine serum (FBS). A working concentration of DMAQB-1 that ranged from 0.5 to 500 nmol/L was studied in the presence of varying concentrations of glucose and insulin. The ability of DMAQB-1 to stimulate glucose transport at less than or equal to 100 nmol/L was determined by [(14)C]-2 deoxyglucose uptake. DNA synthesis was used as the marker for proliferative stimulus and detected by [(3)H]-thymidine uptake measured at 24 hours. Analysis of variance was used to compare the results among the groups; a P value less than.05 was considered significant. Polynomial regression was used to calculate the median lethal dose. RESULTS: In normal glucose media (100 mg/dL), various concentrations of DMAQB-1 demonstrated a small but statistically significant decrease in DNA synthesis at 0.5 nmol/L in serum-free media and at 5 nmol/L in media supplemented with 1% FBS. The corresponding median lethal dose was 107 nmol/L in serum-free media and 650 nmol/L in media supplemented with 1% FBS. A DMAQB-1 concentration of 5 nmol/L induced glucose transport that was equivalent to an insulin concentration of 100 microU/mL. In serum-free, high glucose media (200 mg/dL), DMAQB-1 concentrations up to 500 nmol/L did not cause a statistically significant change in DNA synthesis. When serum-free, high glucose media was combined with mild (100 microU/mL) or moderate (250 microU/mL) concentrations of insulin, DMAQB-1 caused no statistically significant increase in DNA synthesis. CONCLUSION: Nontoxic doses of DMAQB-1 can induce glucose transport equivalent to insulin in the physiologic range. However, DMAQB-1 does not have an insulin-like proliferative effect on human VSMCs in normal-glucose, high-glucose, or high-insulin environments. PMID- 11107084 TI - Initial evaluation of carotid angioplasty and stenting with three different cerebral protection devices. AB - OBJECTIVE: The purpose of the study was to assess the effectiveness of cerebral protection devices during carotid artery angioplasty and stent placement. METHODS: Between September 1998 and September 1999, carotid angioplasty and stenting were performed in 46 patients with symptomatic (39.1%) or asymptomatic (60.9%) severe carotid artery stenosis. Wallstents were used in all patients with selective predilatation. Cerebral protection devices were used in 25 of these patients. Primary end points were perioperative neurologic complications and mortality. Data were collected prospectively. RESULTS: The overall combined end point of all neurologic deficits and death rate was 4.34%. Two neurologic events (one transient ischemic attack and one minor stroke) occurred in the unprotected group (9.53%) versus none in the group with cerebral protection. This difference is not statistically significant. The mortality rate was 0% for both groups. On an intention to treat basis, the overall technical success rate for carotid angioplasty was 97.8%, and for placement of cerebral protection devices it was 100%. An important number of particles of different sizes were captured in all cases in which cerebral protection devices were used. CONCLUSION: Experience has shown that cerebral protection during carotid angioplasty and stenting is technically feasible and appears to be effective in preventing procedure-related neurologic complications. Further investigation is warranted. PMID- 11107085 TI - Brachial artery catheterization to facilitate endovascular grafting of abdominal aortic aneurysm: safety and rationale. AB - PURPOSE: Endovascular treatment of abdominal aortic aneurysms (AAAs) is a technically demanding procedure that is based on the complexity and multiplicity of steps and the guidewire and catheter manipulations required. Brachial artery catheterization is an adjunctive technique that can facilitate the placement of an endoluminal prosthesis. METHODS: Brachial access was used during endoluminal AAA repair in 79 of 103 consecutive patients with a modular-design stent-graft prosthesis at two institutions. RESULTS: Left brachial access facilitated (1) angiography to guide juxtarenal device deployment, (2) antegrade contralateral limb access, (3) device delivery through disadvantaged iliac arteries by means of a brachial femoral wire, (4) access to renal arteries when necessary, and (5) catheter exchanges and a reduction in fluoroscopic positional changes. Complications included one puncture-site pseudoaneurysm, seven hematomas, and 29 patients with extensive ecchymosis. The length of stay was not prolonged in any case. There were no embolic, oculocerebral, or ischemic upper extremity events. CONCLUSIONS: Brachial artery catheterization, as an adjunctive technique to endoluminal AAA repair, offers noteworthy technical advantages with few, but self limiting complications. PMID- 11107086 TI - Duplex ultrasound scanning versus computed tomographic angiography for postoperative evaluation of endovascular abdominal aortic aneurysm repair. AB - OBJECTIVE: The purpose of this study was to compare duplex ultrasound scanning and computed tomographic (CT) angiography for postoperative imaging and surveillance after endovascular repair of abdominal aortic aneurysm (AAA). METHODS: One hundred consecutive patients with AAA underwent endovascular (Medtronic AneuRx, stent graft) aneurysm repair and were imaged with both CT angiography and duplex ultrasound scanning at regular intervals after the procedure. Each imaging modality was evaluated for technical adequacy and for documentation of aneurysm size, endoleak, and graft patency. In concurrent scan pairs, accuracy of duplex scanning was compared with CT. RESULTS: A total of 268 CT scans and 214 duplex scans were obtained at intervals of 1 to 30 months after endovascular aneurysm repair (mean follow-up interval, 9+/-7 months). All CT scans were technically adequate, and 198 (93%) of 214 duplex scans were technically adequate for the determination of aneurysm size, presence of endoleak, and graft patency. Concurrent (within 7 days of each other) scan pairs were obtained in 166 instances in 76 patients (1-6 per patient). The maximal transverse aneurysm sac diameter measured with both methods correlated closely (r = 0.93; P <.001) without a significant difference on paired analysis. In 92% of scans, measurements were within 5 mm of each other. Diagnosis of endoleak on both examinations correlated closely (P <.001), and compared with CT, duplex scanning had a sensitivity of 81%, a specificity of 95%, a positive predictive value of 94%, and a negative predictive value of 90%. Discordant results occurred in 8% of examinations, and in none of these was the endoleak close to the attachment sites or associated with aneurysm expansion. An endoleak was demonstrated on both tests in all eight patients who had an endoleak judged severe enough to warrant arteriography. Graft patency was documented in each instance, without discrepancy, with both modalities. CONCLUSIONS: High-quality duplex ultrasound scanning is comparable to CT angiography for the assessment of aneurysm size, endoleak, and graft patency after endovascular exclusion of AAA. PMID- 11107087 TI - The effect of a venous anastomosis Tyrell vein collar on the primary patency of arteriovenous grafts in patients undergoing hemodialysis. AB - PURPOSE: Vein collars and patches are used at the distal anastomoses of infrainguinal prosthetic grafts to improve graft patency. We initiated a randomized, prospective study to determine whether a Tyrell vein collar at the venous anastomosis of forearm loop arteriovenous grafts (AVGs) would improve patency. METHODS: Patients who required new forearm AVGs were randomized to (1) a standard end-to-side graft-vein anastomosis (control group) or (2) a Tyrell vein collar between the graft and the vein (study group). End points were (1) graft thrombosis, (2) graft removal and ligation, or (3) inadequate graft function. Randomization of 75 subjects was planned. The study was terminated early for ethical reasons. RESULTS: Seventeen patients (eight men, nine women) with a mean age of 52.8 years (range, 31-79 years) had 17 grafts placed (control group, n = 10; study group, n = 7). Comorbidities were not different between the groups (P>.05). Six (86%) of seven study grafts failed by 9 months (mean, 4.6 months). Four (66%) failed study grafts had venous outflow tract stenosis from intimal hyperplasia. This was confirmed at surgery in three and by angiography in one. The 9-month primary patency was 80% for the control group versus 17% for the study group (P =.015). Smaller outflow vein diameter in the study group (P =. 048) did not account for this inferior graft patency. CONCLUSION: A Tyrell vein collar at the venous anastomosis of a forearm AVG resulted in premature graft failure. The use of a Tyrell vein collar may accelerate venous anastomosis intimal hyperplasia. PMID- 11107088 TI - Effects of a venous cuff at the venous anastomosis of polytetrafluoroethylene grafts for hemodialysis vascular access. AB - INTRODUCTION AND METHODS: The most frequent complication of polytetrafluoroethylene (PTFE) arteriovenous grafts for hemodialysis is thrombotic occlusion due to stenosis caused by intimal hyperplasia. This complication is also known for peripheral bypass grafts. Because the use of a venous cuff at the distal anastomosis improves the patency of peripheral bypass grafts, we considered that it might also improve the patency of PTFE arteriovenous grafts. Therefore, a randomized multicenter trial was carried out to study the effect of a venous cuff at the venous anastomosis of PTFE arteriovenous grafts on the development of stenoses and the patency rates. RESULTS: Of the 120 included patients, 59 were randomized for a venous cuff. The incidence of thrombotic occlusion was lower in the cuff group (0.68 per patient year) than in the no-cuff group (0. 88 per patient-year; P =.0007). However, the primary and secondary patency rates were comparable. The cuff group tended to have fewer stenoses at the venous and arterial anastomoses when examined with duplex scan. Graft failure was higher in patients with an initial anastomosing vein diameter smaller than 4 mm (7 of 18 [39%]) than in those with a vein diameter of 4 mm or larger (16 of 88 [18%]; P =. 052). Local edema, skin atrophy, and obesity yielded a higher risk on graft failure (23% vs 11%). CONCLUSION: A venous cuff at the venous anastomosis of PTFE arteriovenous grafts for hemodialysis reduced the incidence of thrombotic occlusions; stenosis at the venous anastomosis was reduced. However, this did not result in a better patency rate. Therefore, the venous cuff should not be used routinely. Initial vein diameter and local problems (edema, obesity, or skin atrophy) appear to be the most important risk factors for graft failure. PMID- 11107089 TI - Lower ankle/brachial index, as calculated by averaging the dorsalis pedis and posterior tibial arterial pressures, and association with leg functioning in peripheral arterial disease. AB - OBJECTIVE: We compared three commonly used methods of ankle/brachial index (ABI) calculation to determine their relative association with objective measures of leg functioning in peripheral arterial disease. METHOD: The study design was cross-sectional; the setting was an academic medical center. The participants were 244 men and women, aged 55 years and older, with and without peripheral arterial disease, from a noninvasive vascular laboratory and a general medicine practice. The main outcome measures were walking velocity and endurance, measured with the 4-m walk and the 6-minute walk, respectively. Three methods of ABI calculation were assessed: using the highest arterial pressure within each leg (method #1), using the lowest pressure in each leg (method #2), and averaging the dorsalis pedis and posterior tibial pressures within each leg (method #3). For each method, we established the prevalence of peripheral arterial disease. We then used regression analyses to identify the ABI calculation method most closely associated with leg functioning. The ABI with the greatest statistical significance and largest regression coefficient was considered most closely associated with leg functioning. RESULTS: Peripheral arterial disease prevalence ranged from 47% when method #1 was used to 59% when method #2 was used. When the right and left legs were compared, the leg with the lower ABI, as identified through use of method #3, was most associated with leg functioning. Within the leg with the lower ABI, method #3 was more closely associated with 6-minute walk distance (regression coefficient = 811.5 feet per 1 unit ABI; P<.001) and 4-m walking velocity (regression coefficient = 0.353 m/s per 1 unit ABI; P<.001) than method #1 or method #2. CONCLUSION: The lower ABI, determined by averaging the dorsalis pedis and posterior tibial arterial pressures in each leg, is most predictive of walking endurance and walking velocity in peripheral arterial disease. PMID- 11107090 TI - The usefulness of a laser Doppler in the measurement of toe blood pressures. AB - OBJECTIVE: The purpose of this study was to evaluate the clinical value and reproducibility of laser Doppler (LD) versus photoplethysmography (PPG) in the measurement of the systolic toe blood pressure. METHODS: Toe blood pressure was measured in 60 patients in different stages of peripheral vascular disease with simultaneous digital sampling of PPG and two LD signals, each with a different filter setting (3 second [LD(3)] and 0.03 second [LD(0.03)]), and cuff pressure. These measurements were repeated after 1 week. The signals were analyzed with previous results ignored. The agreement of the PPG and LD pressures and reproducibility after 1 week were assessed by calculating the intraclass correlation coefficient (ICC). The agreement variation across the range of pressure values was visually explored by means of difference plots. RESULTS: In 19 legs with a very low pressure only LD could adequately measure the pressure, whereas PPG did not. The ICCs between PPG and LD(3) and LD(0.03) were 0.95 or more. The ICCs of the 1-week reproducibility of the PPG, LD(3), and LD(0.03) pressures were 0.92, 0.88, and 0.86, respectively. The variation was equally distributed across the range of pressures in all three methods. CONCLUSION: LD is a reliable alternative to PPG to measure toe blood pressures. Furthermore, LD is able to measure low pressures, which is relevant in the assessment of the presence of critical ischemia. PMID- 11107091 TI - Incidentally detected stenoses proximal to grafts originating below the common femoral artery: do they affect graft patency or warrant repair in asymptomatic patients? AB - OBJECTIVE: Stenoses in infrageniculate arteries proximal to a lower extremity vein graft may reduce flow velocity through the bypass graft and are thought to predispose to graft occlusion. Repair of these lesions has been recommended to preserve graft function. This study was undertaken to better define the natural history of grafts below inflow lesions and to evaluate the necessity of repair to preserve graft patency. METHODS: From 1994 through 1999, patients undergoing lower extremity vein grafts by a single surgeon at a university hospital and an affiliated teaching hospital were placed in a prospective protocol for proximal infrageniculate native artery and graft surveillance through use of duplex scanning. The records of those patients with grafts originating distal to the common femoral artery were evaluated; they form the basis for this report. Arteriograms were obtained before bypass grafting, and no patient had a stenosis greater than 50% diameter reduction proximal to the graft origin. Follow-up scans were obtained from the common femoral artery through the graft and outflow artery. The peak systolic velocity and velocity ratio in an infrageniculate native artery proximal to the graft origin were recorded, as were the location and the time interval since the bypass graft. Repair of these proximal lesions was not performed during the course of this study. Revision of the bypass graft or its anastomoses was undertaken according to preestablished duplex scan criteria. RESULTS: During this time, 288 autogenous infrainguinal bypass grafts were performed, of which 159 originated below the common femoral artery; of these, 74 were from the superficial femoral artery, 29 from the profunda femoris artery, 49 from the popliteal artery, and 7 from a tibial artery. The maximum peak systolic velocity proximal to the graft origin was more than 250 in 38 arteries (25%) and more than 300 in 26 arteries (16%). The velocity ratio was 3.0 or more in 32 arteries at the same location as the peak systolic velocity and 3.5 or more in 23 arteries (15%), confirming hemodynamically significant stenoses at these sites. The location of peak systolic velocity was the common femoral artery in 81 patients (51%), the superficial femoral artery in 50 (31%), the popliteal artery in 22 (14%), and a tibial artery in 6 (4%). Follow-up ranged from 8 to 60 months (mean, 35 months). During follow-up, 19 patients died, 18 with patent grafts. Overall, nine grafts occluded. One of the occluded grafts had a velocity ratio greater than 3.0; this may have contributed to graft thrombosis. The other occlusions resulted from an unrepaired graft lesion in 2 patients, graft infection in 2 patients, and graft ligation necessitated by below-knee amputation in 2 patients. No cause for the occlusion could be identified in two of the grafts (neither had evidence of proximal arterial stenosis). Assisted primary patency rates were 95% and 91% at 3 and 5 years, respectively. CONCLUSIONS: For grafts originating distal to the common femoral artery, stenoses proximal to the graft do not affect bypass graft patency and do not require repair to prevent graft occlusion. Surveillance of these lesions may therefore be unnecessary, inasmuch as the repair of proximal lesions should not be undertaken to preserve graft function. PMID- 11107092 TI - Adventitial versus intimal liposome-mediated ex vivo transfection of canine saphenous vein grafts with endothelial nitric oxide synthase gene. AB - PURPOSE: Experiments were designed (1) to evaluate liposome-mediated endothelial constitutive nitric oxide synthase (ecNOS) transfection in vein grafts and (2) to compare intimal and adventitial routes of transfection. METHODS: Male mongrel dogs (N = 36) underwent bilateral femoral artery bypass grafting with the lateral saphenous vein. In each animal one vein was transfected with plasmid (pVR1012) containing the ecNOS gene, and another vein was transfected with plasmid alone (control). Gene transfer was performed from either the intimal surface (Group I, n = 18) or the adventitial surface (Group II, n = 18). In each group there were three transfection subgroups (n = 6 each): (a ) 10 microg/mL naked plasmid DNA, (b ) 10 microg/mL plasmid DNA + liposome (LipofectAMINE PLUS), and (c ) 100 microg/mL plasmid DNA + LipofectAMINE PLUS. Grafts were harvested on the third postoperative day, and the transfection was assessed with molecular techniques and enzyme assay for activity of NOS by conversion of tritiated l-arginine to tritiated l-citrulline. Proliferating cells were quantified with a digital analysis of histologic sections after nuclear antigen Ki-67 (MIB1) immunohistochemistry. RESULTS: Transgene was identified with polymerase chain reaction in all ecNOS-transfected grafts, regardless of transfection modality. However, significant transcription of the ecNOS transgene was observed only in Group IIc (mean ecNOS messenger RNA, 8.7+/-1.7 vs. 3.1+/-0.7 x 10(-2) attomole/microL, in transfected compared with control grafts, respectively, P =.01). NOS activity increased approximately twofold in this group (11.58+/-2.1 and 6.3+/-1.0 pmol tritiated l-citrulline per milligram protein per hour in transfected and control grafts, respectively, P = .05). Numbers of proliferating cells did not differ among ecNOS-transfected and control grafts in any transfection group. CONCLUSION: These results suggest that ecNOS transfection of vein grafts is feasible through intimal and adventitial routes with naked DNA or a liposomal vector. However, efficient transcription of the transgene is evident at postoperative day 3 only after adventitial transfection of 100 microg/mL of the gene. PMID- 11107093 TI - Increased amount of type III pN-collagen in human abdominal aortic aneurysms: evidence for impaired type III collagen fibrillogenesis. AB - PURPOSE: This study aimed to characterize the distribution of structural domains of type I and III collagens in the wall of abdominal aortic aneurysms (AAAs), by the use of undilated atherosclerotic aortas (aortoiliac occlusive disease [AOD]) and healthy abdominal aortas as controls. METHODS: Immunohistochemical staining was applied with antibodies for the aminoterminal propeptides of type I (PINP) and type III (PIIINP) procollagens, which represent newly synthesized type I and III pN-collagens. In addition, an antibody against the aminoterminal telopeptide of type III collagen (IIINTP) was used as a means of detecting maturely cross linked type III collagen fibrils. RESULTS: The newly synthesized type III procollagen detected by means of PIIINP staining was concentrated in the media in aneurysmal aortas, whereas type I pN-collagen was localized in the intima in both AAAs and AODs. The healthy aortas showed no immunoreactivity for either PIIINP or PINP. The cross-linked type III collagen, detected by means of IIINTP staining, stained transmurally in all study groups, but appeared more abundant in the media in AAAs. CONCLUSION: Our results strongly suggest that the metabolism of type III collagen is enhanced in AAAs. Intensive type III pN-collagen staining was present mainly in the media layer in AAAs, suggesting a role of type III collagen in aneurysm formation, whereas type I pN-collagen was present in the intima in both AAAs and AODs, suggesting that type I collagen synthesis is a fibroproliferative response related to the atherosclerotic process. The increased type III pN collagen in AAAs may result in impaired fibril formation and, thus, in decreased tensile strength of aneurysmal tissue. PMID- 11107094 TI - Kommerell's diverticulum and aneurysmal right-sided aortic arch: a case report and review of the literature. AB - Right-sided aortic arch is a rare variant of the thoracic vascular anatomy that may be accompanied by an aberrant origin of the left subclavian artery. We report a true aneurysm of the distal arch and descending thoracic aorta in a patient with right-sided arch and review previous descriptions of aneurysms of anomalous right-sided aortas. In our patient, the left subclavian artery originated at the junction between the distal arch and the descending thoracic aorta located in the right chest and was aneurysmal (Kommerell's diverticulum); the thoracic aorta was also aneurysmal. Extra-anatomic left subclavian-to-carotid transposition was performed before the intrathoracic procedure. Subsequently, a right thoracotomy provided adequate exposure for repairing the aortic aneurysm and oversewing the aneurysmal origin of the subclavian artery. Because the distal aortic arch was involved, deep hypothermia and circulatory arrest were used. Only five previous instances of true aneurysms of a right-sided aortic arch have been reported; four of these patients underwent operative repair (via bilateral thoracotomy, median sternotomy, or right thoracotomy). We believe that a right thoracotomy provides good exposure and avoids the morbidity associated with bilateral thoracotomy. The reconstruction of the subclavian artery has not previously been reported in this setting. Performing subclavian reconstruction as an extrathoracic procedure before the intrathoracic repair would be expected to reduce the subsequent risk of distal ischemia or subclavian steal without increasing the overall morbidity associated with the procedure. PMID- 11107095 TI - Repair of a saccular aortic aneurysm with superficial femoral-popliteal vein in the presence of a pancreatic abscess. AB - When one is faced with impending rupture, repair of an aortic aneurysm cannot be delayed. In the presence of coexisting intra-abdominal sepsis, traditional therapy would call for aneurysm exclusion and axillofemoral bypass grafting. Consequences of this choice of treatment include limited long-term graft patency and recurrent prosthetic infection. Autogenous deep veins from the lower extremities have demonstrated exceptional patency and resilience to infection when used to replace infected aortic grafts. We now report a case of concomitant open drainage of a pancreatic abscess and repair of a saccular abdominal aortic aneurysm using the superficial femoral-popliteal vein as a conduit. PMID- 11107096 TI - Survival of an aortic trauma patient with Ehlers-Danlos syndrome type IV: a case report. AB - Ehlers-Danlos syndrome type IV is the most lethal variant of that illness and is associated with fatal large vessel arterial hemorrhages. The literature reports only two survivors of elective aortic surgery and two survivors of spontaneous aortic hemorrhage. This article presents a 14-year-old boy who had aortic and vena cava blunt trauma and survived. PMID- 11107097 TI - Endovascular repair of aortic allograft aneurysmal degeneration: a case report. AB - Aortoenteric graft fistula remains a dreadful complication of aortic surgery. Good results have been reported using in situ graft replacement with arterial allografts. Late aneurysmal degeneration of the graft itself may necessitate further repair. We report the case of such an aneurysmal degeneration 7 years after implantation of the allograft. Endovascular repair was performed with a Vanguard device; complete exclusion was obtained immediately. At 6-month follow up, the patient was alive and well. Duplex and computed tomography scans showed an excluded aneurysm with a slight reduction in size. Endovascular stent grafting may be a therapeutic option for treating patients with late allograft degeneration. PMID- 11107098 TI - Arteriovenous fistulae complicating cardiac pacemaker lead extraction: recognition, evaluation, and management. AB - Transvenous pacemaker lead extraction has become a commonly performed procedure that is associated with a small but significant risk. We report two cases where lead extraction was complicated by arteriovenous fistulae between branches of the aortic arch and the left brachiocephalic vein. Presenting signs and symptoms included severe chest or back pain, persistent or copious bleeding from the venous puncture site, unexplained hypotension or anemia, superior vena cava syndrome, and signs of central venous hypertension or acute heart failure. One patient whose injury was not recognized immediately and who did not undergo repair died rapidly, whereas the other patient who was diagnosed quickly underwent successful repair. Immediate diagnosis with arteriography and rapid intervention with surgery or percutaneous techniques are indicated and may prevent mortality. PMID- 11107099 TI - Distal emboli as an unusual late complication of a thrombosed arteriovenous hemodialysis graft. AB - The creation of an arteriovenous fistula for long-term hemodialysis access is one of the most commonly performed procedures in vascular and transplantation surgery. Prosthetic conduits are frequently prone to failure within their first year of construction, and after one or two revisions, they are left in their thrombosed state as permanent subcutaneous foreign bodies in the extremities. Conventional teaching has regarded these chronically thrombosed grafts to have a benign natural history, and their removal has been considered unnecessary. We describe an unusual late complication of distal thromboemboli from a chronically occluded arteriovenous graft that was implanted 10 years before and appeared as acute hand ischemia. PMID- 11107100 TI - Banding of the common iliac artery: an expedient in endoluminal correction of aortoiliac aneurysms. AB - Dilatation of the common iliac arteries is one of the most frequent causes for exclusion of patients in a series of endovascular correction of abdominal aortic aneurysms (AAAs). In this article we describe the banding technique we use to constrict the large iliac arteries. Four patients underwent endovascular treatment for AAAs with bifurcated grafts. Five of the eight common iliac arteries were 16 to 20 mm in diameter and were constricted around the endoprosthesis by banding with two cotton tapes through a retroperitoneal access. An angioplasty balloon was used as a counterresistance inside the graft. Completion angiogram and postoperative computed tomographic scans showed no endoleak in all cases. No complications occurred in the follow-up (3-10 months). Banding of the common iliac artery is an efficient procedure for endoluminal correction of AAAs when the diameter of the common iliac arteries is greater than 16 mm and less than 20 mm. PMID- 11107101 TI - Regarding "the effect of venous anastomosis Tyrell vein patch collar on the primary patency of arteriovenous grafts in patients undergoing hemodialysis" and "effects of a venous cuff at the venous anastomosis of polytetrafluoroethylene grafts for hemodialysis vascular access". PMID- 11107102 TI - Lifeline Foundation and National Institutes of Health. PMID- 11107103 TI - Extended outcome assessment in the care of vascular diseases: revising the paradigm for the 21st century. Ad Hoc Committee to Study Outcomes Assessment, Society for Vascular Surgery/International Society for Cardiovascular Surgery, North American Chapter. PMID- 11107104 TI - Advances in Neuroblastoma Research, 2000 Conference. Philadelphia, Pennsylvania, USA. May 15-18, 2000. Proceedings and abstracts. PMID- 11107105 TI - Identification of a 1-megabase consensus region of deletion at 1p36.3 in primary neuroblastomas. AB - BACKGROUND: Deletion of the distal short arm of chromosome 1 occurs frequently in neuroblastoma. In addition, neuroblastoma has been described in children with constitutional deletions within 1p36, supporting the existence of one or more neuroblastoma suppressor genes within this region. PROCEDURE: We have pursued a 1p36 tumor suppressor gene identification strategy that has included deletion mapping of 566 primary neuroblastomas and 46 neuroblastoma-derived cell lines, and have determined the parental origin of the deleted 1p homologue in 44 cases to determine whether there is evidence for genomic imprinting within this region. RESULTS AND CONCLUSIONS: We have identified a 1-Mb consensus region of deletion within 1p36.3 defined by primary tumor deletions, constructed a physical map of the region that is being sequenced to completion, and have identified and prioritized candidate genes within this region for further analyses. PMID- 11107106 TI - Identification of the homozygously deleted region at chromosome 1p36.2 in human neuroblastoma. AB - BACKGROUND: We have identified for the first time a homozygously deleted region within the smallest region of overlap at 1p36.2-3 in two neuroblastoma cell lines. PROCEDURE: The 800-kb PAC contig covering the entire homozygously deleted region was made and sequenced. To date, approximately 70% of sequencing has been accomplished, and the estimated length of the deleted region was 500 kb. RESULTS: Currently, we have found six genes within the region, which include three known genes as well as three other genes that have been reported during processing of our present project for the last 3(1/2) years. We report here the results of expression and mutation analyses of those genes. CONCLUSIONS: Full sequencing for the region of homozygous deletion as well as further analyses of the genes mapped within the region may reveal whether or not there is a neuroblastoma suppressor gene as proposed by the Knudson's two-hit hypothesis. PMID- 11107107 TI - Deletion mapping of 14q32 in human neuroblastoma defines an 1,100-kb region of common allelic loss. AB - BACKGROUND: We analyzed loss of heterozygosity (LOH) in 54 primary neuroblastomas (NBs) using 12 microsatellite markers on 14q32, and found that 31% (17/54) NBs showed LOH. PROCEDURE: The smallest region of overlap (SRO) was identified between D14S62 and D14S987. RESULTS: There was no statistical correlation between LOH and any clinicopathologic features, including age, stage, amplification of MYCN, and ploidy. A sequence-ready bacterial artificial chromosome (BAC) contig was constructed, and the minimum tiling path of six BACs covered the SRO; the physical length of this region was no larger than 1,000 kb. CONCLUSIONS: Our findings support the existence of a putative tumor-related gene on 14q32 for the tumorigenesis of NB. PMID- 11107108 TI - Localization of a hereditary neuroblastoma predisposition gene to 16p12-p13. AB - BACKGROUND: Hereditary predisposition to develop neuroblastoma segregates as an autosomal dominant Mendelian trait. PROCEDURE: We have performed linkage analysis on 10 families with neuroblastoma to localize a hereditary neuroblastoma predisposition gene (HNB1). RESULTS: A single genomic interval at chromosome bands 16p12-p13 was consistent with linkage (lod = 3.46), and identification of informative recombinants defined a 25.9-cM critical region between D16S748 and D16S3068. Loss of heterozygosity was identified in 5/12 familial (42%) and 55/259 nonfamilial (21%) neuroblastomas at multiple 16p polymorphic loci. A 12.8-cM smallest region of overlap of deletions was identified within the interval defined by linkage analysis (tel-D16S764-D16S412-cen). CONCLUSIONS: Taken together, these data suggest that HNB1 is located at 16p12-p13 and that inactivation of this gene may contribute to the pathogenesis of nonfamilial neuroblastomas. PMID- 11107109 TI - Analysis of loss of heterozygosity at 16p12-p13 (familial neuroblastoma locus) in 470 neuroblastomas including both sporadic and mass screening tumors. AB - BACKGROUND: Neuroblastoma (NBL) in children usually occurs in a sporadic form. However, it rarely occurs in families. Recently, the familial neuroblastoma (FNB) locus has been mapped to 16p12-p13 by linkage analysis. PROCEDURE: Here we show the result of loss of heterozygosity in the region spanning 16p12-p13 (D16S406 D16S409, 46 cM) in 470 NBLs including both sporadic and mass screening cases. RESULTS: Allelic loss was found in 61(13%) tumors. Deletion of 16p was associated with mass screening tumor (P = 0.035) and <1 year of age at diagnosis (P = 0.048). We found two commonly deleted regions: the sizes of the region were approximately 2 cM and approximately 6 cM. CONCLUSIONS: Our data suggested that allelic loss of 16p was common in favorable NBLs, and there may be at least two candidate loci within the region of FNB. PMID- 11107110 TI - Laser-capture microdissected schwannian and neuroblastic cells in stage 4 neuroblastomas have the same genetic alterations. AB - BACKGROUND: Neuroblastoma (NB) is a heterogeneous tumor composed of mixed populations of neuroblastic, Schwannian, and gangliocytic cells. PROCEDURE: We examined the genetic lineage of Schwannian and neuroblastic cells in stage 4 NB using laser-capture microdissection (LCM) and allelic analysis of microsatellite markers of chromosomes 1p, 11q, and 14q. RESULTS: In 18 out of 19 cases, individual cell types exhibited the same allelotype. CONCLUSION: Our results provide evidence that the majority of Schwannian and neuroblastic cells in stage 4 NB are of neoplastic origin, and support the hypothesis of a tumor stem cell capable of development along multiple lineages. PMID- 11107111 TI - Chromosome 2 short arm translocations revealed by M-FISH analysis of neuroblastoma cell lines. AB - PROCEDURE: M-FISH analysis was performed on 18 neuroblastoma cell lines, which were previously studied with cytogenetic, standard FISH and CGH data. RESULTS: One of the most striking findings of this study was the detection of chromosome 2 short arm rearrangements in 61% of the investigated cell lines. These rearrangements resulted from translocations with various partner chromosomes. All translocations, except one were unbalanced, leading to the consistent gain of chromosome segment 2pter-p22. A cryptic balanced translocation t(2;4) was observed with a breakpoint located in the vicinity of MYCN in cell line NBL-S. CONCLUSIONS: Combination of M-FISH results together with cytogenetic, standard FISH and CGH data yielded the most comprehensive description of chromosome 2 short arm rearrangements, leading to a consistent gain of chromosome 2 short arm material. PMID- 11107112 TI - Absent or reduced expression of the caspase 8 gene occurs frequently in neuroblastoma, but not commonly in Ewing sarcoma or rhabdomyosarcoma. AB - PROCEDURE: To clarify whether the caspase 8 gene is involved in the pathogenesis of neuroblastoma (NB), we examined alterations of the caspase 8 gene in 15 NB, seven Ewing sarcoma (ES), and eight rhabdomyosarcoma (RMS) cell lines, using reverse transcription-polymerase chain reaction (RT-PCR) and RT-PCR single-strand conformation polymorphism (SSCP) analyses. RESULTS: The caspase 8 gene was not expressed in 11 (73%) of 15 NB cell lines, it was absent in only one of seven ES cell lines, but was present in all eight RMS cell lines examined. No mutations were detected in any cell lines examined. CONCLUSIONS: Inactivation of the caspase 8 gene is considered to be involved in the pathogenesis of NB, but not ES or RMS. PMID- 11107113 TI - Deletion of 11q23 is a frequent event in the evolution of MYCN single-copy high risk neuroblastomas. AB - BACKGROUND: Deletions of the long arm of chromosome 11 are frequently identified in human neuroblastomas. PROCEDURE: We screened 394 primary neuroblastomas and 52 tumor-derived cell lines with a panel of 11q and 11p polymorphic markers to determine the frequency of chromosome 11 allelic deletion, and to differentiate partial deletions of chromosome 11q (unb[11q] LOH) from whole chromosome loss. RESULTS: Allelic deletion occurred most frequently at cytogenetic band 11q23 and was detected in 161 primary neuroblastomas (41%) and 18 cell lines (35%). Eighty seven tumors (22%) had unb[11q] LOH with a heterogeneous distribution of deletion breakpoints. Unb[11q] LOH was highly correlated with age > 1 year at diagnosis (P = 0.008), stage 4 disease (P = 0.001), unfavorable Shimada histopathology (P < 0.001), and assignment to a high-risk therapeutic protocol (P < 0.001), and was inversely correlated with MYCN amplification (P = 0.018). Patients whose tumors showed unb[11q] LOH were less likely to survive (P < 0.001), but there was only a trend towards an independent prognostic influence in multivariate analyses. CONCLUSIONS: Thus, structural rearrangements resulting in unb[11q] LOH commonly occur during the malignant evolution of high-risk neuroblastomas with single-copy MYCN. PMID- 11107114 TI - Hunting the subset-specific genes of neuroblastoma: expression profiling and differential screening of the full-length-enriched oligo-capping cDNA libraries. AB - BACKGROUND: Neuroblastoma (NBL) has a distinct nature in different prognostic subgroups. PROCEDURE: To understand the molecular mechanism of NBL's genesis and biology as well as that of the neural crest development, we constructed full length-enriched cDNA libraries by an oligo-capping method from two different subsets of primary NBL, one with favorable biology and the other with MYCN amplification. RESULTS: Sequencing analysis of these libraries revealed that the expression profile was markedly different between both subsets. To identify the genes differentially expressed between the subsets, semi-quantitative RT-PCR analyses are proceeding. CONCLUSION: So far, 54 transcripts have been found to be expressed at high levels in favorable NBLs, and significantly at low levels in unfavorable NBLs. PMID- 11107115 TI - Role of survivin, whose gene is mapped to 17q25, in human neuroblastoma and identification of a novel dominant-negative isoform, survivin-beta/2B. AB - PROCEDURE: We investigated the expression of survivin (SVV) and its isoform (SVV beta/2B) during different biological properties in neuroblastoma (NBL). RESULTS: High levels of SVV mRNA expression were significantly associated with advanced stages of NBL, diagnosis at over 1 year of age, low levels of TrkA expression, and sporadic tumors. Expression of a novel isoform, SVV-beta/2B, which had an insertion of 23 amino acids within the unique BIR domain was predominant in some favorable NBLs, while it was low and ubiquitous in most normal and malignant tissues. The SVV expression wasdown-regulated during apoptosis induced by retinoic acid (RA) in CHP134 NBL cells, which was inhibited by forced expression of SVV. In contrast, SVV-beta was constantly expressed during apoptosis. Like SVV,SVV-beta was also highly expressed during G(2)/M in a cell cycle-dependent manner, and was associated with but competed against SVV for binding with polymerized tubulin. CONCLUSION: These data suggest that expression of SVV is a poor prognostic indicator in human NBL, and it promotes growth and survival by regulating the levels of both isoforms. PMID- 11107116 TI - SAGE analysis of neuroblastoma reveals a high expression of the human homologue of the Drosophila Delta gene. AB - BACKGROUND: Serial Analysis of Gene Expression (SAGE) is an efficient method to establish a complete mRNA expression profile of a tissue. PROCEDURE: We applied SAGE to identify expression of developmental control genes in neuroblastoma. Results. The human homologue of the Drosophila Delta gene Delta like-1 (DLK1) was shown to have an unusually high expression in a SAGE library of the SK-N-FI neuroblastoma cell line. Northern blot analysis confirmed high DLK1 expression in SK-N-FI and several other neuroblastoma cell lines. Signalling between Delta and its receptor Notch controls many differentiation steps in Drosophila and man, including neural crest cell fate decision. CONCLUSIONS: Our data therefore suggest a role for the Delta-Notch pathway in neuroblast differentiation. PMID- 11107117 TI - BIN1 inhibits colony formation and induces apoptosis in neuroblastoma cell lines with MYCN amplification. AB - BACKGROUND: MYCN amplification and overexpression occurs in 25% of neuroblastomas and independently predicts for poor prognosis disease, an effect thought to be mediated by its role as a transcriptional activator of growth promoting genes. However, in many mammalian cells, deregulated expression of MYC family genes (including MYCN) induces apoptosis. We hypothesized that BIN1, a MYC interacting protein capable of inducing apoptosis, may be an important regulator of MYCN in neuroblastoma. RESULTS: BIN1 expression was found to be reduced in MYCN-amplified cell lines. Further, forced expression of BIN1 markedly reduced colony formation in MYCN-amplified, but not single-copy, cell lines. This effect appeared to be caused by an increase in apoptosis, and was augmented by serum deprivation and concurrent cytotoxic drug therapy in cell culture CONCLUSION: BIN1 inactivation may be necessary for MYCN overexpression to lead to cellular proliferation rather than programmed cell death in neuroblastomas with MYCN amplification. PMID- 11107118 TI - p53 mutations and loss of p53 function confer multidrug resistance in neuroblastoma. AB - BACKGROUND: Neuroblastomas often acquire sustained drug resistance during therapy. Sensitivities to carboplatin, etoposide, or melphalan were determined for 18 neuroblastoma cell lines; eight were sensitive and ten were resistant. As p53 mutations are rare in neuroblastomas studied at diagnosis, we determined if acquired p53 mutations and loss of function conferred multidrug resistance. RESULTS: Loss of p53 function (p53-LOF), defined as a failure to induce p21 and/or MDM2 in response to melphalan, was seen in 1/8 drug-sensitive and 6/10 drug-resistant cell lines. In four cell lines p53-LOF was associated with mutations in the DNA binding region of p53, while three cell lines with LOF and four cell lines with functional p53 had no evidence of p53 muta-tions. Nonfunctional and mutated p53 was detected in one resistant cell line, while a sensitive cell line derived from the same patient prior to treatment had functional and wild type (wt) p53. We transfected HPV 16 E6 (which mediates degradation of p53, causing LOF) into two drug-sensitive neuroblastoma cell lines with functional p53. LC(90) values of HPV 16 E6 transfected cell lines were 3-7 fold (melphalan), 8-109-fold (carboplatin), and 2-158-fold (etoposide) greater than that of LXSN-transfected controls. CONCLUSIONS: These data suggest that some neuroblastomas acquire p53 mutations during therapy, which is associated with a loss of p53 function, and can confer high-level multidrug resistance. PMID- 11107119 TI - Expression of neurotrophin receptor TrkA inhibits angiogenesis in neuroblastoma. AB - BACKGROUND: Mechanisms regulating the expression of angiogenic factors in tumor cells are largely unknown. High expression of the neurotrophin receptor TrkA in neuroblastomas (NB) is associated with favorable prognosis, whereas TrkB is expressed on aggressive, MYCN-amplified NB. PROCEDURE: To investigate the biological effects of TrkA and TrkB expression on angiogenesis in NB, we examined the expression of angiogenic factors in the human NB cell line SY5Y and its TrkA and TrkB transfectants. RESULTS: In comparison to parental SY5Y cells, mRNA and protein levels of angiogenic factors were significantly reduced in SY5Y-TrkA cells, whereas SY5Y-TrkB cells did not demonstrate a significant change. Conditioned medium (CM) of parental SY5Y and SY5Y-TrkB cells induced endothelial cell proliferation, but this effect was completely absent in SY5Y-TrkA cells. TrkA expression also resulted in severely impaired tumorigenicity in a mouse xenograft model, and was associated with reduced angiogenic factor expression and less vascularization of tumors, as determined by immunohistochemistry and an in vivo Matrigel assay. PMID- 11107120 TI - p75 mediated apoptosis in neuroblastoma cells is inhibited by expression of TrkA. AB - BACKGROUND: Neurotrophins mediate their effects by binding to members of the Trk family of receptor tyrosine kinases and to the low-affinity nerve growth factor receptor p75. Nerve growth factor (NGF) has been demonstrated to support survival and differentiation of neuroblastoma (NB) cells by activation of the TrkA receptor. The p75 receptor belongs to the tumor necrosis factor (TNF) family of death receptors and has been suggested as a receptor that mediates apoptosis in neuronal and NB cells. PROCEDURE: To investigate the effect of p75 expression in NB, we transfected the p75 cDNA into SH-SY5Y cells, an NB cell line lacking expression of both p75 and TrkA. RESULTS: Cell clones expressing elevated levels of p75 showed a high degree of apoptosis even in 10% serum-supplemented medium. Apoptotic signaling by p75 was ligand-independent and only partly caspase dependent. The level of apoptosis correlated directly with the expression level of the receptor, indicating that p75 may activate the cell death program directly. However, additional transfection of TrkA into SY5Y-p75 cells resulted in a significantly reduced rate of apoptosis even in the absence of NGF. CONCLUSIONS: Thus, expression of the TrkA receptor itself inhibits p75 mediated apoptosis in NB cells. PMID- 11107121 TI - Histone deacetylase inhibitors and retinoic acids inhibit growth of human neuroblastoma in vitro. AB - BACKGROUND: Neuroblastoma is a common childhood cancer with a poor overall prognosis. Retinoic acids (RAs) have been studied as a potential therapy, showing promise in recurrent disease. The histone deacetylase inhibitor (HDACI) M carboxycinnamic acid bishydroxamide (CBHA) is another potential therapy, which we recently described. Combinations of RAs and HDACIs currently under investigation display synergy in certain neoplasms. In this study, we evaluate the effect of combinations of RAs and HDACIs on human neuroblastoma cells. PROCEDURE: Established cell lines were cultured in increasing concentrations of HDACIs, RAs, and combinations thereof. Following exposure, viable cell number was quantified by trypan blue dye exclusion on a hemacytometer. Cell cycle analysis was performed by propidium iodide staining and FACS. RESULTS: All assayed HDACIs and RAs decreased viable cell number. Lower concentrations of each agent were effective when the two were combined. The primary reason for decreased cell number appears to be apoptosis following HDACI exposure and G1 arrest following RA exposure. Both effects are seen with cotreatment. Caspase inhibition abrogates the apoptotic response. CONCLUSIONS: CBHA causes apoptosis of human neuroblastoma in vitro, an effect that can add to the effects of RA. HDACIs and RAs inhibit neuroblastoma in significantly lower concentrations when used together than when used individually. Combination therapy may improve the ultimate efficacy while reducing the side effects of these agents in clinical use. PMID- 11107122 TI - MycN sensitizes neuroblastoma cells for drug-triggered apoptosis. AB - BACKGROUND: Amplification of the MYCN gene is found in a large proportion of neuroblastomas and is associated with a poor prognosis. PROCEDURE: To investigate the effect of ectopic MycN expression on the susceptibility of neuroblastoma cells to cytotoxic drugs, we used a human neuroblastoma cell line with tetracycline-controlled expression of MycN. RESULTS: Neither conditional expression of MycN alone nor low drug concentrations induced apoptosis. However, MycN and cytotoxic drugs cooperated to induce cell death. Apoptosis triggered by MycN and doxorubicin was mediated by cleavage of caspases and involved activation of the CD95 system. MycN overexpression and cytotoxic drugs also synergized to induce p53 and Bax protein expression and to trigger mitochondrial permeability transition and cytochrome c release. CONCLUSION: In that amplification of MYCN is considered an adverse prognostic factor, these findings suggest that dysfunctions in apoptosis pathways may be a mechanism by which MycN-induced apoptosis of neuroblastoma cells is inhibited. PMID- 11107123 TI - Expression of N-myc and MRP genes and their relationship to N-myc gene dosage and tumor formation in a murine neuroblastoma model. AB - BACKGROUND: Although the association between N-myc gene amplification and poor clinical outcome in neuroblastoma is well established, the mechanism by which amplification influences prognosis is not well defined. PROCEDURE: We used a human N-myc transgenic mouse model to investigate the role of N-myc in neuroblastoma, including its relationship to the multidrug-resistance-associated protein (MRP) gene. We developed a rapid real-time PCR method to distinguish homozygous and hemizygous N-myc mice that is comparable to Southern analysis. RESULTS: A highly significant correlation (P < 0.0001) between N-myc and MRP expression was demonstrated in murine tumors. Amplification of the transgene was observed in the majority of tumors, highlighting the clinical relevance of this model. However, no correlation between N-myc expression and transgene dosage or tumor latency was observed. CONCLUSIONS: The data suggest that increased N-myc dosage contributes to increased tumor incidence and decreased latency by mechanisms independent of N-myc expression. PMID- 11107124 TI - Schwann cell-conditioned medium inhibits angiogenesis in vitro and in vivo. AB - BACKGROUND: Neuroblastomas are biologically heterogeneous tumors that consist of two main cell populations: neuroblastic/ganglionic cells and Schwann cells. The amount of Schwannian stroma strongly impacts prognosis. Low tumor vascularity, localized stage, and favorable outcome are associated with tumors that are Schwannian stroma-rich/stroma-dominant. PROCEDURE: To investigate if Schwann cells play a role in inhibiting angiogenesis in neuroblastoma tumors, we examined the ability of human Schwann cell-conditioned medium to affect bFGF- and VEGF induced endothelial cell proliferation and migration, and in vivo angiogenesis. RESULTS: Schwann cell-conditioned medium significantly inhibited bFGF- and VEGF induced endothelial cell proliferation and migration. This effect appears to be specific for endothelial cells as smooth muscle cell and fibroblast proliferation were not inhibited by this medium. Schwann cell-conditioned medium also inhibited in vivo angiogenesis in rat corneal assays. CONCLUSIONS: Schwann cells produce a potent inhibitor(s) of angiogenesis that may be responsible for the low level of vascularity and more benign clinical behavior of Schwannian stroma-rich/stroma dominant neuroblastoma tumors. Studies to identify the inhibitor(s) are ongoing. PMID- 11107125 TI - Human polyomavirus BK (BKV) and neuroblastoma: mechanisms of oncogenic action and possible strategy for novel treatment. AB - BACKGROUND: We reported previously that nearly all human neuroblastomas analyzed contain and express genomic DNA sequences deriving from the human polyomavirus BK (BKV) [Flaegstad et al.: Cancer Res 59:1160-1163, 1999]. PROCEDURE: Here we show that the BKV large T antigen is expressed and bound to p53 in neuroblastoma cells and that this interference compromises the tumor suppressor function of p53. RESULTS: Treatment of neuroblastoma cells with large T antigen antisense constructs relocated active p53 to the nucleus. The relocation event was accompanied by enhanced p21(waf1/cip1) expression as well as induced apoptosis. CONCLUSIONS: Continuous antisense oligonucleotide treatment of nude rats with human neuroblastoma xenografts resulted in a significant but incomplete reduction of tumor growth compared to rats treated with saline. PMID- 11107126 TI - Retinoic-acid-resistant neuroblastoma cell lines show altered MYC regulation and high sensitivity to fenretinide. AB - BACKGROUND: High-dose, pulse-13-cis-retinoic acid (13-cis-RA) given after intensive cytotoxic therapy improves event-free survival for high-risk neuroblastoma (NB), but more than 50% of patients have tumor recurrence. PROCEDURE: We conducted multistep selection for resistance to all-trans-retinoic acid (ATRA) in NB cell lines with (SMS-KCNR and LA-N-5) or without (SMS-LHN) MYCN genomic amplification. RESULTS: After 12 exposures to 10 microM ATRA, the two MYCN-amplified cell lines (KCNR 12X RR and LA-N-5 12X RR) showed partial resistance to the cytostatic/differentiation effects of ATRA; complete resistance was seen in LHN 12X RR. ATRA-selected cells showed general RA resistance (cross resistance to 13-cis-RA). Transient (KCNR 12 X RR, LA-N-5 12X RR) or sustained (LHN 12X RR) novel overexpression of c-myc was associated with RA resistance. RA insensitive overexpression of MYCN by transduction in SMS-LHN also conferred RA resistance. Both parental and RA-resistant lines showed 2-4 logs of cell kill in response to N-(4-hydroxyphenyl)retinamide (4- HPR, fenretinide). Compared to parental lines, 4-HPR achieved 1-3 log greater cell kills in RA-resistant LHN 12X RR, LA-N-5 12X RR, KCNR 12X RR, and MYCN-transduced SMS-LHN or SK-N-RA. NB cell lines (n = 26) from 21 different patients showed that 16 of 26 (62%) were sensitive to 4-HPR (LC(90) < 10 microM), including lines established at relapse after myeloablative and/or 13-cis-RA therapy. CONCLUSION: Thus, RA-resistant NB cell lines can be sensitive (and in some cases collaterally hypersensitive) to 4 HPR. PMID- 11107127 TI - Resistance to TRAIL-induced apoptosis in neuroblastoma cells correlates with a loss of caspase-8 expression. AB - BACKGROUND: Disruption of apoptotic pathways may be involved in tumor formation, regression, and treatment resistance of neuroblastoma (NB). TNF-related apoptosis inducing ligand (TRAIL) is a potent inducer of apoptosis in cancer cell lines. PROCEDURE: In this study we analyzed the expression and function of TRAIL, its agonistic and antagonistic receptors, and important intracellular signaling elements in 18 NB cell lines. RESULTS: Semiquantitative RT-PCR revealed that TRAIL-R2 and TRAIL-R3 are the main TRAIL-receptors used by NB cells. Sensitivity to TRAIL-induced apoptosis did not correlate with mRNA expression of TRAIL receptors or cFLIP. Surprisingly, caspase-8 and caspase-10 mRNA was detected in only 5 of 18 NB cell lines. Interestingly, only these five NB cell lines were susceptible to TRAIL-induced apoptosis in a time- and dose-dependent manner. CONCLUSIONS: Treatment with 5-aza-2'-deoxycytidine restored mRNA expression of caspase-8 and -10 and TRAIL sensitivity of resistant cell lines, suggesting that gene methylation is involved in caspase inactivation. Since many cytotoxic drugs induce caspase-dependent apoptosis, failure to express caspase-8 and/or caspase 10 might be an important mechanism of resistance to chemotherapy in NB. PMID- 11107128 TI - Loss of caspase-8 expression in neuroblastoma is related to malignancy and resistance to TRAIL-induced apoptosis. AB - Background and Procedures NB-derived cell lines were tested for their sensitivity to apoptosis induced by the tumor-selective apoptotic ligand TRAIL. Noninvasive S type cell lines are highly sensitive to TRAIL, whereas invasive N-type cell lines are resistant. RESULTS: Although both S- and N-type cell lines express TRAIL-R2, FADD, and caspases-3 and -10, only S-type cells express caspase-8. Reduced levels of caspase-8 protein were also observed in a stage IV NB tumor when compared to a ganglioneuroma. The caspase-8 gene is not deleted in either N-type NB cell lines or high-stage tumors, and expression can be induced by demethylation. CONCLUSIONS: Therefore, caspase-8 expression is silenced in malignant NB, which correlates to tumor severity and resistance to TRAIL-induced apoptosis. PMID- 11107129 TI - 6-fluorodopamine selectively destroys neuroblastoma cells expressing the noradrenaline transporter. AB - BACKGROUND: 6-Hydroxydopamine (6-OHDA) was used for ex vivo purging of bone marrow from neuroblastoma cells before autologous transplantation. However, this concept failed because of the rapid autoxidation of 6-OHDA, which leads to the generation of cytotoxic reactive oxygen species (ROS), mainly in the incubation medium before 6-OHDA can be incorporated by neuroblastoma cells. PROCEDURE: We based our experiments on the theory that, in contrast, 6-fluorodopamine (6-FDA), which is slowly converted to 6-OHDA at neutral pH, is able to enter neuroblastoma cells via the noradrenaline transporter (NA-T). Therefore, most ROS are generated inside the target cells. RESULTS: Small amounts of ascorbate prevent the extracellular conversion of 6-FDA to 6-OHDA without affecting its cytotoxicity, leading to an even more selective effect of 6-FDA. CONCLUSIONS: We conclude that 6-FDA is a promising substance for selective destruction of NA-T-positive neuroblastoma cells. PMID- 11107130 TI - Betulinic acid induces apoptosis through a direct effect on mitochondria in neuroectodermal tumors. AB - BACKGROUND AND PROCEDURE: We identified BetA as a new cytotoxic agent active against neuroectodermal tumor cells including neuroblastoma, medulloblastoma, glioblastoma and Ewing sarcoma cells, representing the most common solid tumors of childhood. RESULTS: BetA induced apoptosis by a direct effect on mitochondria independent of accumulation of wild-type p53 protein and independent of death inducing ligand/receptor systems such as CD95. Mitochondrial perturbations on treatment with BetA resulted in the release of soluble apoptogenic factors such as cytochrome c or AIF from mitochondria into the cytosol, where they induced activation of caspases. Overexpression of the anti-apoptotic proteins Bcl-2 or Bcl-X(L) that blocked loss of the mitochondrial membrane potential and cytochrome c release from mitochondria also conferred resistance to BetA. Most importantly, BetA exhibited potent antitumor activity on neuroblastoma cells resistant to CD95 or doxorubicin-triggered apoptosis and on primary tumor cells from patients with neuroectodermal tumors. CONCLUSIONS: Thus, BetA may be a promising new agent in the treatment of neuroectodermal tumors including neuroblastoma in vivo. PMID- 11107131 TI - Multidrug resistance-associated protein 1 (MRP1) expression in neuroblastoma cell lines and primary tumors. AB - BACKGROUND AND PROCEDURE: MRP1 expression by neuroblastomas was evaluated by Northern blot analysis in 21 cell lines and 90 primary untreated tumors. Cytotoxicity assay in cell lines was performed for five anticancer drugs used in treating neuroblastoma. RESULTS: MRP1 expression did not correlate with drug resistance or with MYCN RNA expression in cell lines. MRP1 expression was higher in drug-sensitive cell lines established after chemotherapy relative to cell lines at diagnosis, but highly drug-resistant cell lines showed low MRP1 expression. Positive expression of MRP1 RNA in primary tumors was associated with a poorer survival relative to MRP1-negative tumors. However, MRP1 expression levels did not correlate with age, stage, MYCN amplification, or MYCN expression, and higher MRP1 expression was not associated with a worse outcome. CONCLUSIONS: In neuroblastoma, positive MRP1 RNA expression at diagnosis has prognostic significance, but high drug resistance is conferred by mechanisms other than MRP1. PMID- 11107132 TI - Different effects of TrkA expression in neuroblastoma cell lines with or without MYCN amplification. AB - BACKGROUND: Expression of the neurotrophin receptor TrkA is associated with a favorable prognosis in neuroblastoma (NB) and promotes growth inhibition and neuronal differentiation. Aggressive, MYCN-amplified NB tumors express little or no TrkA mRNA, suggesting that MYCN overexpression may inhibit TrkA expression. PROCEDURE: To study the interactions of TrkA expression and MYCN amplification in NB, we stably expressed the TrkA receptor in the MYCN single copy cell lines SH SY5Y and NB69 as well as in the MYCN amplified cell lines CHP134 and IMR5. RESULTS: All four transfected cell lines demonstrated high TrkA expression and similar activation of the TrkA receptor and of mitogen-activated protein kinases as well as induction of immediate-early genes in response to nerve growth factor (NGF). Introduction of TrkA restored NGF responsiveness of SH-SY5Y and NB69 cells, as demonstrated by morphologic differentiation, growth inhibition, and enhanced survival in serum-free medium. However, no morphologic, growth, or survival responses to NGF were detected in MYCN-amplified CHP134 and IMR5 TrkA transfectants. CONCLUSIONS: Thus, transfection of TrkA into MYCN amplified NB cell lines only partly restored the TrkA/NGF signaling pathway, suggesting additional inhibitory effects of MYCN overexpression on TrkA signaling. PMID- 11107133 TI - Association between favorable neuroblastoma and high expression of the novel metalloproteinase gene, nbla3145/XCE, cloned by differential screening of the full-length-enriched oligo-capping neuroblastoma cDNA libraries. AB - BACKGROUND: The biology of neuroblastoma (NBL) is at least partly regulated by neurotrophic factors and their receptors. PROCEDURE: To identify novel NBL related genes that affect growth, differentiation, and programmed cell death of the tumor, we constructed full-length-enriched cDNA libraries by oligo-capping method. RESULTS: Semi-quantitative and quantitative real-time RT-PCR showed that the nbla3145 gene was significantly highly expressed in favorable subset of NBL. The nucleo-tide sequence of nbla3145 revealed that it was a novel member of metalloproteinase, endothelin-converting enzyme, and was the same gene recently reported as XCE. We also cloned nbla3145beta which was a novel truncated form of nbla3145 (or nbla3145alpha). CONCLUSIONS: Our results suggested that nbla3145/XCE plays an important role in regulating growth and differentiation of NBL. PMID- 11107134 TI - Prognostic significance of GAGE detection in bone marrows on survival of patients with metastatic neuroblastoma. AB - BACKGROUND: Previously, we reported the utility of GAGE as a molecular marker for neuroblastoma (NB) and malignant melanoma in blood and bone marrow (BM). Among patients with stage III melanoma rendered disease-free by surgery, GAGE expression was a strong prognostic factor for patient survival. PROCEDURE: All patients with advanced NB diagnosed at > 1 year of age initially treated with protocol N6 (n = 24) and N7 (n = 38) at Memorial Sloan-Kettering Cancer Center were included in this study. Their BM cells at 12, 18, and 24 months (median time after diagnosis) were evaluated for the presence of GAGE. RESULTS: GAGE positivity at 12 months (25%), when patients were still on treatment, did not predict progression-free survival (PFS) and overall survival from the time of sampling. Positivity at 18 months (29%) was associated with poorer PFS and survival (but P > 0.05). By 24 months, the presence of GAGE (26%) was a very strong predictor of out-come (P < 0.001). When only remission marrows at 24 months were analyzed, PFS was 4.7-fold lower among GAGE-positive patients. Thirty seven percent of N6 patients were positive for GAGE, in contrast to 17% of the patients in the more current regimen N7. CONCLUSIONS: The detection of GAGE by RT PCR in marrow may have utility in molecular staging of patients in clinical remission. It may allow earlier identification of patients at risk, such that appropriate intervention can be given before clinical relapse. GAGE may also serve as a surrogate endpoint for adjuvant treatment strategies, and to determine the duration of therapy. PMID- 11107135 TI - Correlation of anti-idiotype network with survival following anti-G(D2) monoclonal antibody 3F8 therapy of stage 4 neuroblastoma. AB - BACKGROUND: A transient human anti-mouse antibody response was associated with significantly longer survival [Cheung et al. (1998): J Clin Oncol 16:3053] following antibody 3F8 (Ab1) treatment. We postulate that the induction of an idiotype network which included anti-anti-idiotypic (Ab3) and anti-G(D2) (Ab3') responses is associated with tumor control. PROCEDURE: Thirty-four patients with stage 4 neuroblastoma (NB) diagnosed at > 1 year of age were treated with anti G(D2) monoclonal antibody 3F8 at the end of chemotherapy RESULTS: Long-term progression-free survival and overall survival correlated significantly with Ab3' andAb3, but not with non-idiotypic antibody responses. Only one of six individual specificities showed significant correlations with patient survival. CONCLUSIONS: As in vitro correlates of idiotype network initiated by Ab1 treatment, Ab3 and Ab3' may provide convenient biologic endpoints for monoclonal antibody therapy of advanced NB, and a rationale for choosing specific anti-idiotypic antibodies for vaccine development. PMID- 11107136 TI - Retroviral vector-producer cell mediated angiogenesis inhibition restricts neuroblastoma growth in vivo. AB - BACKGROUND: The purpose of this study was to determine whether gene therapy mediated delivery of an angiogenesis inhibitor, a truncated, soluble vascular endothelial growth factor receptor (Flk-1/KDR, VEGFR-2), could suppress tumor growth in a murine model of neuroblastoma. METHODS: Murine fibroblasts producing a replication-defective retrovirus encoding this mutant form of flk-1 were made. These producer cells were mixed with neuroblastoma cells and injected subcutaneously into SCID mice. Subsequent tumor growth was then measured. RESULTS: Murine neuroblastoma growth was decreased by 95% after 25 days. Similar tumor growth inhibitory effects were observed when the flk-1 producer cells were co-injected with cells from two different human neuroblastoma cell lines. CONCLUSIONS: Neuroblastoma growth can be significantly restricted in vivo with a single injection of cells that produce a retroviral vector encoding the gene for an angiogenesis inhibitor. This suggests that gene therapy-mediated delivery can be an effective alternative to chronic administration of these cytostatic anticancer agents. PMID- 11107137 TI - Tyrosine hydroxylase-based DNA-vaccination is effective against murine neuroblastoma. AB - BACKGROUND: The disruption of self-tolerance against neuroblastoma is the ultimate goal of an effective DNA-vaccine. PROCEDURE: Here we demonstrate the induction of a protective immunity against syngeneic murine NXS2 neuroblastoma in A/J mice, following vaccination with tyrosine hydroxylase (TH) derived antigens. Oral gene delivery was accomplished using an attenuated strain of Salmonella typhimurium as a carrier harboring vectors encoding for mTH antigens. RESULTS: Vaccination was effective in protecting animals from a lethal challenge with wild type NXS2 tumor cells. CONCLUSIONS: These results provide the first evidence of the TH self antigen being recognized by T-cells and demonstrate that a TH-based DNA vaccine is a potentially useful immunotherapeutic strategy for neuroblastoma. PMID- 11107138 TI - Telomerase activity by TRAP assay and telomerase RNA (hTR) expression are predictive of outcome in neuroblastoma. AB - BACKGROUND: Recent studies have associated telomerase with prognostic factors and survival in neuroblastoma. PROCEDURE: We examined telomerase activity by telomere repeat amplification protocol (TRAP) and expression of the RNA component of telomerase (hTR) by Northern blotting in 106 primary neuroblastoma tumors and 22 established cell lines. RESULTS: Overall survival at 5 years for all 106 tumors was significantly better for patients with undetectable TRAP (75% vs. 59%; P = 0.03) or low hTR expression (84% vs. 43%; P < 0.0001), and especially for patients whose tumors had both low hTR expression and undetectable TRAP (all patients, 91% vs. 54%, P = 0.0002; for 17 stage IV-S tumors, 100% vs. 72%, P = 0.04). Strong expression of hTR was seen in 22 cell lines from aggressive tumors, and all maintained telomere length, but 3/22 were TRAP negative. CONCLUSIONS: These data suggest that both hTR expression and telomerase activity via the TRAP assay should be performed concurrently to predict survival in neuroblastoma patients, particularly in stage 4-S. PMID- 11107139 TI - Telomerase is a strong indicator for assessing the proneness to progression in neuroblastomas. AB - BACKGROUND: As traditional parameters do not ensure completely accurate prognostic grouping in neuroblastoma (NB), new molecular markers are needed for assessing the individual patient's prognosis more precisely. PROCEDURE, RESULTS, AND CONCLUSIONS: Based on 133 NB, we show that telomerase activity (TA) is a powerful, independent prognostic marker for all stages and is capable of differentiating between good and poor outcome in putative 'favorable' clinical or biological subgroups of NB patients. Analysis of gene and protein expression of telomerase subunits suggests that the presence or absence of TA in NB is strongly correlated with expression levels of both the catalytic subunit hTERT and the internal RNA component (hTR). PMID- 11107140 TI - Prognostic significance of EPHB6, EFNB2, and EFNB3 expressions in neuroblastoma. AB - BACKGROUND: EPH family receptor tyrosine kinases and their ligand ephrins play pivotal roles in development. High-level expression of transcripts encoding EPHB6 receptors (EPHB6), its ligands ephrin-B2 and ephrin-B3 (EFNB2, EFNB3) is predictive of favorable disease outcome of neuroblastoma (NB). When combined with TrkA expression, the expression of EPHB6, EFNB2, or EFNB3 predicts more accurately the disease outcome than each of the four variables alone. PROCEDURE: Cox regression and Kaplan-Meier analyses were used to assess the prognostic significance of EPHB6, EFNB2, EFNB3, and TrkA expressions in NB without MYCN amplification. RESULTS: High-level expression of EFNB3 or TrkA predicted favorable NB outcome of NB without MYCN amplification (p < 0.03). As found in the general NB population, EPHB6, EFNB2, or EFNB3 expression in combination with TrkA expression was significantly predictive of the disease outcome of normal MYCN NB (p < 0.01). CONCLUSIONS: EPHB6, EFNB2, and EFNB3 expressions may permit further refinement of the prognostic stratification of NB into favorable and unfavorable groups. PMID- 11107141 TI - Synergism of buthionine sulfoximine and melphalan against neuroblastoma cell lines derived after disease progression. AB - BACKGROUND: Despite intensive-alkylator based regimens, >50% of patients with high-risk neuroblastoma (NB) die from recurrent disease that is probably due, in part, to acquired alkylator resistance. PROCEDURE: Using buthionine sulfoximine (BSO)-mediated, glutathione (GSH) depletion to modulate melphalan (L-PAM) resistance, we examined six NB cell lines established after progressive disease following either standard chemotherapy, BSO/L-PAM therapy, or myeloablative therapy and autologous hematopoietic stem cell transplant (AHSCT). RESULTS: Four of the six cell lines (three p53-nonfunctional and one p53-functional) showed high-level L-PAM resistance. CONCLUSIONS: Fixed ratio analysis demonstrated BSO/L PAM synergy (combination index >1) for all cell lines tested. In L-PAM-resistant cell lines, the minimal cytotoxicity observed for BSO combined with nonmyeloablative concentrations of L-PAM was markedly enhanced (>4 logs total cell kill) when BSO was combined with myeloablative concentrations of L-PAM. In alkylator-resistant NB, the optimal use of BSO may require dose escalation of L PAM to levels requiring AHSCT. PMID- 11107142 TI - Distinct properties of fenretinide and CD437 lead to synergistic responses with chemotherapeutic reagents. AB - The RARbeta/gamma-selective retinoids fenretinide and CD437 induce caspase dependent apoptosis but generate free radicals independently of caspases. Apoptosis, but not free radical generation, induced by these retinoids is inhibited by RARbeta/gamma-specific antagonists. Both fenretinide and CD437 induce apoptosis synergistically with cisplatin, carboplatin, or etoposide. However, antioxidants inhibit this synergy to the level obtained with chemotherapeutic drugs alone, and this implies that free radical generation is important in the synergistic response. Since apoptosis induced by fenretinide or CD437 is mediated by apoptotic pathways involving RARs and/or mitochondria and differs from mechanisms of chemotherapy-induced apoptosis this may explain the strong synergistic response seen between these synthetic retinoids and chemotherapeutic drugs. These results suggest that fenretinide or CD437 may be useful adjuncts to neuroblastoma therapy. PMID- 11107143 TI - Differential expression in an antisense MYCN neuroblastoma model. AB - BACKGROUND: A previously published antisense MYCN-expressing model system was utilized to identify genes whose expression is altered by the down-regulation of MYCN by AS MYCN. RESULTS: Differential display comparing nontransfected human NB NBL-S cells to three AS MYCN-expressing cell lines (NBAS-4, -5, and -6) yielded nine differentially expressed cDNAs designated NDDE:1-9, for MYCN-dependent differential expression genes. A GenBank search revealed matches for seven of the nine cDNAs. Differential expression was confirmed for five of the cDNAs by Northern blot analysis. RESULTS: NDDE:8 is up-regulated in the AS MYCN-expressing clones and shares homology with the EB1 clone p53-induced gene (PRG3). NDDE:2 is up-regulated in the high-expressing N-myc protein NBL-S cells and shares homology with a 27 kD heat shock protein PAN1 clone. Further analysis of all five cDNAs might further elucidate the targets of MYCN in NB. PMID- 11107144 TI - Inhibition of tumor growth in a human neuroblastoma xenograft model with TNP-470. AB - Background and Procedure High-risk neuroblastoma disease features are correlated with tumor vascularity, suggesting that angiogenesis inhibitors may be a useful addition to current therapeutic strategies. We therefore examined the efficacy of TNP-470 (TAP Pharmaceuticals, Deerfield, IL) in human neuroblastoma xenograft models. RESULTS: Tumor growth rate was markedly inhibited in mice receiving TNP 470 administered alone with a treatment to control ratio (T/C) at day 21 = 0.4 (P <.001). TNP-470 also significantly inhibited tumorigenicity when administered shortly after xenograft inoculation (T/C at day 30 = 0.1, P <.001) and when administered following cyclophosphamide (T/C at day 35 = 0.1, P <.001). CONCLUSIONS: These data show that TNP-470 is a potent inhibitor of human neuroblastoma growth both alone and when given with conventional chemotherapy, suggesting that it may be a useful adjunctive therapy for high-risk neuroblastoma patients. PMID- 11107145 TI - CD34 selection as a stem cell purging strategy for neuroblastoma: preclinical and clinical studies. AB - BACKGROUND: The suitability of CD34 selection for purging peripheral blood progenitor cells (PBPC) collected from patients with neuroblastoma (NB) has been called into question, largely because of reports of detection of low levels of CD34 on the surface of some NB cell lines and tumors. PROCEDURE: We used three approaches to address the issue of purging of NB from stem cell specimens and possible labeling of NB: 1) Flow cytometric detection of CD34 on NB cell lines. We assessed CD34 expression using a panel of anti-CD34 monoclonal antibodies (MoAbs) including 9C5, 12.8, and QBend10 and showed no increase in labeling over secondary-only control. 2) Spiking experiments with the Isolex 50 system. NB cell lines were used to contaminate aliquots of PBPC collections, after which the products were purified using the Isolex 50. Purging of NB was assessed by quantitative multiplex RT-PCR (TaqMan system) using a tumor-specific transcript, GAGE. We demonstrated >2 logs of tumor cell depletion from these specimens. 3) Analysis of clinical specimens. PBPC pre- and post-CD34 selection were analyzed from patients treated on the CHP-594 transplant trial. RESULTS: In nine specimens selected using the Ceprate LC CD34 selection system where tumor was detectable by immunocytochemistry preselection, we observed >2.4 to >4.6 logs of NB purging after selection. We then analyzed 23 aliquots of PBPC infused into patients post CD34 selection and compared them to the product preselection; 20/23 specimens showed depletion of NB, although some level of GAGE message was observed in most post-CD34 selection specimens. CONCLUSION: These data show that purging of NB from PBPC specimens using CD34 selection is feasible, yielding infused products that are negative at the level of ICC but often positive at the level of RT-PCR. PMID- 11107146 TI - Metastatic neuroblastoma in infancy: what does the pattern of metastases contribute to prognosis? AB - BACKGROUND: The purpose was to investigate the influence of metastatic pattern and primary extension over midline on prognosis of infants with metastatic neuroblastoma. PROCEDURE: Data of 317 consecutive infants with metastatic neuroblastoma were analyzed. RESULTS: The amount of bone marrow infiltration (<10% vs. >10%) proved to be the most important factor and was more important than the presence of bone metastases. A disadvantage in outcome for patients with distant lymph node, intracranial, or atypical metastases or for patients with primary extension over midline could not be demonstrated. However, in the subgroup of patients treated with limited treatment, primary extension over midline proved a risk factor. CONCLUSION: A redefinition of Stage 4S on an international basis is suggested. PMID- 11107147 TI - Combined (111)In-pentetreotide scintigraphy and (123)I-mIBG scintigraphy in neuroblastoma provides prognostic information. AB - BACKGROUND: High-affinity somatostatin receptors (SRs) have been characterized in neuroblastomas and may be used as target structures for in vivo detection of SR. PROCEDURE: Eighty-eight children with histologically proven neuroblastoma were investigated at diagnosis or relapse by (123)I-mIBG and (111)In-pentetreotide scintigraphy. All tumors were investigated for MYCN copy number, chromosome 1p36 status, and 68/88 also for DNA content, followed for a median follow-up of 35 months (range 1-88 months). RESULTS: SR expression was detected in 56/88 tumors and (123)I-mIBG showed positivity in 83/88. (111)In-pentetreotide was less sensitive in detecting tumor tissue than was (123)I-mIBG (64% vs. 94%, P = 0.005). Survival (SUR) and event-free survival probability (EFS) according to Kaplan-Meier was significantly better for children with positive SR scintigraphy than for the children with a negative SR scan (SUR: 90% vs. 48% at 4 years log rank P < 0.003, EFS: 83% vs. 39% at 4 years, log rank P < 0.0002). CONCLUSIONS: (123)I-mIBG scintigraphy remains the best scintigraphic method for detecting neuroblastoma tumor tissue, whereas additional SR scintigraphy is able to provide significant prognostic information with a minimum of invasiveness. PMID- 11107148 TI - Selection of human antitumor single-chain Fv antibodies from the B-cell repertoire of patients immunized against autologous neuroblastoma. AB - BACKGROUND: We used phage display technology to clone human recombinant antitumor antibodies from the antibody repertoire of neuroblastoma patients immunized with cytokine-gene transduced tumor cells. PROCEDURE: Lymphocytes obtained from neuroblastoma patients either at diagnosis or after immunization with an autologous interleukin-2 gene transduced tumor vaccine were used to construct two human single-chain Fv (scFV) phage libraries. Tumor-reactive phage were characterized using ELISA, flow cytometry, and sequencing analysis. RESULTS: The initial screening after panning on neuroblastoma cells yielded a substantially higher proportion of selectivity tumor-binding phage clones derived from the immunized patients library (12.9%) than from the unvaccinated patients library (0.8%). The antibodies stained the cells from several additional pediatric malignancies, including Ewing sarcoma and rhabdomyosarcoma, in the absence of binding to any normal tissue cultures or epithelial tumor cell lines. The pattern of reactivity was different from that of antibodies recognizing other widely distributed neuroblastoma-associated antigens, suggesting recognition of a novel shared tumor antigen. CONCLUSION: The human recombinant scFV antibodies reported here appear to represent a tumor-specific B-cell response induced by autologous tumor immunization and are potentially useful targeting moieties for the treatment of selected childhood tumors. PMID- 11107149 TI - Rapid-sequence tandem transplant for children with high-risk neuroblastoma. AB - BACKGROUND: The majority of patients with high risk neuroblastoma (NB) still relapse. PROCEDURE: We designed a Phase II trial for children with advanced NB utilizing a program of induction chemotherapy followed by tandem high-dose chemoradiotherapy with stem cell rescue (HDC/SCR) in rapid sequence. Fifty-five patients were evaluable, ages 1-14 years, and 97 cycles of HDC/SCR have been completed to date. Pheresis was possible for every patient, despite their young age, with an average of 7.2 x 10(6) CD34+ cells/kg available to support each HDC/SCR cycle. RESULTS: Engraftment was rapid, with median time to neutrophil engraftment of 11 days. Five patients who completed the first HDC course did not complete the second and there were four toxic deaths. With a median follow-up of 24 months from diagnosis, 38 of 55 patients (3-year EFS 59%) remain event-free. A subset of the patients received stem cells purged by CD34 selection. The engraftment and EFS of these patients are similar to the overall group. CONCLUSION: This work demonstrates that a tandem transplant regimen for high-risk NB is a feasible treatment strategy in children and may improve disease-free survival. PMID- 11107150 TI - Population-based and controlled study to evaluate neuroblastoma screening at one year of age in Germany: interim results. AB - BACKGROUND: The German Neuroblastoma Screening Project is the first controlled and population-based screening study to evaluate the presumed benefit of neuroblastoma mass screening at 1 year of age (10-18 months). PROCEDURE: Screening takes place in 6 of the 16 German states; children from the remainder serve as controls. The German Childhood Cancer Registry enables a mostly complete follow-up and detection of false-negative patients. RESULTS: Up to December, 1999, 1,199,165 children were examined for urinary catecholamine metabolites and 124 cases of neuroblastoma were detected preclinically, giving a detection rate of 10.3/100,000. Within this cohort, 33 false-negative cases were found. CONCLUSIONS: The results of this screening program will be crucial for further implementation of neuroblastoma screening. PMID- 11107151 TI - Risk of unfavorable character among neuroblastomas detected through mass screening. The Japanese Infantile Neuroblastoma Cooperative Study. AB - Current study shows that about 50% of neuroblastomas (NBs) detected through mass screening had factor(s) indicating an unfavorable biological nature and that early intervention after the screening might improve clinical outcome of the patients. On the other hand, favorable properties were detected in the remaining half of the mass-screening NBs. Some of them might have the ability to regress spontaneously. Therapeutic modality should be determined according to their biological nature. Further investigation for their biologic properties is necessary to evaluate the benefits of the mass screening. PMID- 11107152 TI - A gene therapy approach to enhance the targeted radiotherapy of neuroblastoma. AB - BACKGROUND: The aims of this study were to determine whether the introduction and expression of the noradrenaline transporter (NAT) gene into NAT-negative neuroblastoma cell lines would make them amenable to targeted radiotherapy using [(131)I]MIBG. PROCEDURE: Neuroblastoma cell lines were transfected with a eukaryotic expression vector containing the bovine noradrenaline transporter cDNA under the expression of the CMV promoter. Stable transfectants were created by selection in geneticin (G418) and were characterised for their MIBG uptake ability and susceptibility to [(131)I]MIBG therapy. RESULTS: The cell line SK-N MC, which normally shows no ability to take up MIBG, was successfully transfected with bNAT. SK-N-MC.bNAT transfectants exhibited uptake and release kinetics similar to those of the natural NAT-expressing cell line SK-N-BE(2c). Levels of [(131)I]MIBG uptake were 33% of those of the highest naturally NAT-expressing cell line SK-N-BE(2c). Growth delay assays using multicellular spheroids indicated that this degree of [(131)I]MIBG uptake was sufficient to inhibit growth at radioactive concentrations of 4 Mbq/ml. CONCLUSIONS: These results demonstrate the feasibility of combining gene therapy with targeted radiotherapy to enhance uptake, and hence radiation dose, to neuroblastoma tumours using [(131)I]MIBG. With the appropriate delivery vehicle and tumour-specific control of expression, the introduction of noradrenaline transporter molecules may be a viable means of enhancing the response of neuroblastoma tumours to [(131)I]MIBG therapy. PMID- 11107153 TI - Humoral response to vaccination with interleukin-2-expressing allogeneic neuroblastoma cells after primary therapy. AB - BACKGROUND: Immunotherapy using cytokine-expressing tumor cells has shown promise as an anticancer strategy. We have recently begun a trial of interleukin-2 (IL-2) gene-modified allogeneic neuroblastoma cells administered in a sequence of eight injections to patients with high-risk neuroblastoma following completion of primary therapy. Six patients to date have completed treatment. PROCEDURE: We examined humoral responses to the immunizing cell line and, when available, to the patients' autologous tumor cells using an in vitro binding assay. RESULTS: Five of six patients developed a rise in antitumor antibodies to the immunizing neuroblastoma cell line following vaccination. Two of these patients had autologous tumor available; both demonstrated a humoral response to these cells as well. CONCLUSIONS: Our results demonstrate that vaccination with IL-2 expressing allogeneic tumor cells after intensive primary therapy can elicit a humoral response to the immunizing line. These antibodies are cross-reactive with the patients' own tumor cells in the two cases in which autologous cells were available. This suggests that different patients' tumors may share common antigens that can be exploited in immunotherapy strategies and supports the continued exploration of allogeneic tumor cells as tumor vaccines. PMID- 11107155 TI - Phase I topotecan preparative regimen for high-risk neuroblastoma, high-grade glioma, and refractory/recurrent pediatric solid tumors. AB - We evaluated the toxicity and maximum tolerated dose of topotecan in a novel myeloablative regimen as treatment for high-risk pediatric tumors. Patients received an assigned topotecan dosage in combination with fixed doses of carboplatin and thiotepa, followed by autologous hematopoietic stem cells infusion. Topotecan dose was escalated in cohorts of four patients until the maximum tolerated dose of topotecan was defined or until accrual of 30 patients. Pharmacokinetics of topotecan were examined, and event-free survival was estimated. We describe preliminary results following treatment of 25 pediatric patients with high-risk solid tumors. PMID- 11107154 TI - Targeted radioimmunotherapy for leptomeningeal cancer using (131)I-3F8. AB - BACKGROUND: Intrathecal antibody-based targeted therapies may have clinical potential for patients with leptomeningeal (LM) cancer. PROCEDURE: Five patients with GD2-positive LM tumors were injected with 1-2 mCi intra-Ommaya (131)I-3F8, a murine IgG3 antibody specific for GD2. Serial cerebrospinal fluid (CSF) and serum samples and SPECT imagings (4, 24, and 48 hr) were performed to predict radiation doses to the tumor and normal brain and blood prior to the administration of larger therapeutic doses. RESULTS: Side effects included self-limited fever, headache, and vomiting. Focal (131)I-3F8 uptake consistent with tumors was seen along the craniospinal axis in four patients. Calculated radiation dose to the CSF was 14.9-56 cGy/mCi and to blood and other organs outside the CNS less than 2 cGy/mCi. CONCLUSIONS: Intraventricular (131)I-3F8 successfully detected LM disease and resulted in a large favorable CSF/blood ratio. Intraventricular (131)I-3F8 may have clinical utility in the diagnosis and radioimmunotherapy of GD2-positive LM cancers. Med. Pediatr. Oncol. 35:716-718. 2000. PMID- 11107156 TI - Results of the cooperative protocol (N-III-95) for metastatic relapses and refractory neuroblastoma. AB - BACKGROUND: Prognosis of relapsed and refractory neuroblastoma is uniformly fatal; new therapeutic approaches are needed. PROCEDURE: Relapsed and refractory neuroblastoma patients were treated with continuous infusion chemotherapy combined with MIBG. RESULTS: Over 4 years, 35 heavily pretreated patients were registered, 29 with bone or/and bone marrow metastases. Grade 3 or 4 hematologic toxicity was frequent, without toxic deaths. Sixteen patients responded. The probability of 5-year overall survival was 0.19. CONCLUSIONS: This approach is feasible and toxicity manageable; it rescued some patients and prolonged their survival. It merits assay in newly diagnosed high-risk neuroblastoma patients. PMID- 11107158 TI - Erratum PMID- 11107157 TI - Meeting summary for Advances in Neuroblastoma Research--2000. PMID- 11107159 TI - Neurofibromatosis type 1. I. General overview. PMID- 11107160 TI - Evidence for a cytoskeleton attachment domain at the N-terminus of the NF2 protein. AB - Neurofibromatosis type 2 is a hereditary cancer syndrome characterized by the development of bilateral vestibular schwannomas. Underlying the disease are inactivating mutations of the NF2 tumor suppressor gene, located on chromosome 22, encoding a 595-amino-acid protein. The NF2 protein, also known as merlin or schwannomin, is reported to act as a membrane-cytoskeleton linking protein. This assumption is based on the homology of the NF2 protein to a group of band 4.1 related proteins, ezrin, radixin, and moesin. The cytoskeletal association of the NF2 protein has in part been confirmed by its ability to resist extraction from cells by nonionic detergents. We performed detergent extraction on COS cells transfected with NF2 cDNA constructs. The extracts were analyzed by Western blotting and immunofluorescent staining with monoclonal anti-NF2 antibodies. The results provide evidence for a high-affinity cytoskeleton attachment domain at amino acids 29-131 and a putative lower affinity domain between amino acids 321 and 470. PMID- 11107161 TI - Oligodendrocytes in aging mice lacking myelin-associated glycoprotein are dystrophic but not apoptotic. AB - Although MAG-null mice myelinate relatively normally except for subtle structural abnormalities in the periaxonal region of myelin sheaths, they develop more severe pathological changes as they age. The purpose of this study was to further define the biochemical aspects of CNS pathology caused by an absence of MAG. Proteins associated with myelin and oligodendrocytes were quantified by densitometry of western blots in whole brain homogenates, as well as in isolated myelinated axons and myelin. Neither myelin yields, nor levels of myelin basic protein and proteolipid protein, were decreased in comparison to control levels in 14-month-old MAG-null mice. On the other hand, 2',3'-cyclic nucleotide 3' phosphodiesterase (CNPase) and the 120 kD neural cell adhesion molecule (N-CAM) were substantially reduced in whole brain, myelinated axons, and myelin. Tubulin, Na(+)K(+)ATPase and Fyn tyrosine kinase were also reduced significantly in myelin related fractions, but not in whole brain homogenate. The decreased levels of these proteins suggest pathological abnormalities in oligodendrocytes. Furthermore, significant reductions of CNPase and 120 kD NCAM were also present at 2 months, indicating that the oligodendroglial abnormalities begin at a relatively early age. Neither TUNEL assays nor multiplex RT-PCR for mRNAs of apoptosis-related proteins in the aging MAG-null mice provided evidence for apoptotic oligodendrocytes. These biochemical findings suggest oligodendroglial damage in MAG-null mice and support the morphological observations pointing to a progressive "dying-back oligodendrogliopathy" as a consequence of MAG deficiency. PMID- 11107162 TI - Release of plasminogen activator inhibitor-1 from human astrocytes is regulated by intracellular ceramide. AB - The present study underscores a regulatory role of intracellular ceramide in astrocytes for the release of an extracellular serine protease, tissue-type plasminogen activator (t-PA), and its inhibitor, plasminogen activator inhibitor 1 (PAI-1). Treatment of cultured human astrocytes with N-acetylsphingosine, a cell-permeable short-chain ceramide analogue or daunorubicin that could increase intracellular ceramide via activation of ceramide synthase or sphingomyelin hydrolysis increased the release of t-PA and conversely decreased the PAI-1 release. Interestingly, treatment of the astrocytes with tumor necrosis factor (TNF)-alpha also increased the intracellular ceramide levels but caused the elevation of PAI-1 release without altering the t-PA release. These data suggest that the generation of ceramide in astrocytes is linked at least with the regulation of PAI-1 release. We also demonstrate that the suppression of PAI-1 release with daunorubicin accelerates the cell death of neuronally differentiated PC12 cells and suggest an antiapoptotic role of PAI-1 in the nervous system. PMID- 11107163 TI - Corticotropin releasing factor induces proliferation of cerebellar astrocytes. AB - In the adult cerebellum, corticotropin releasing factor (CRF), that is localized in climbing fibers, mossy fibers, and a fine varicose plexus along the Purkinje cell layer, modulates the responsiveness of Purkinje cells to excitatory amino acids. During development, CRF has been detected in the primitive cerebellar anlage as early as embryonic day (E)10, and is continuously expressed throughout embryonic and postnatal cerebellar ontogeny. To investigate a possible trophic role for CRF during cerebellar development, cerebellar culture studies using E18 mouse embryos were carried out. In our culture paradigm, that used serum-free defined medium to suppress cell proliferation, CRF induced proliferation of cells in a dose-dependent manner in a range of concentrations between 0.1-10 microM. The proliferating cells were identified as astrocytes based on their expression of vimentin and GFAP. BrdU incorporation studies supported the proposed mitogenic effect of CRF on developing astrocytes. The mitogenic effects of CRF seemed to be primarily on immature astrocytes determined by their differential expression of vimentin and GFAP. Astrocytes at more advanced stages of development, as determined by the extent of process outgrowth and GFAP expression, incorporated less BrdU compared to immature astrocytes. CRF receptors were localized in astrocytes, and the proliferation of astrocytes induced by CRF was inhibited by astressin, a competitive CRF receptor antagonist. In conclusion, CRF induces proliferation of astrocytes derived from the developing cerebellum, that suggests a gliotrophic role for CRF during cerebellar development. PMID- 11107164 TI - Akt negatively regulates the cJun N-terminal kinase pathway in PC12 cells. AB - The protein serine/threonine kinase Akt is a target of phosphatidylinositol 3 kinase that mediates many of the trophic actions of growth factors on cells. In PC12 cells, complete removal of serum leads to rapid stimulation of the cJun N terminal kinase (JNK) pathway. Inclusion of insulin-like growth factor-1, a stimulator of Akt in PC12 cells, inhibits JNK activation in this setting, whereas addition of wortmannin to PC12 cells in the presence of serum stimulates JNK activity, suggesting that growth factor-mediated signaling through the phosphatidylinositol 3-kinase/Akt pathway chronically inhibits the JNK pathway in PC12 cells. To explore the possible role of Akt as a negative regulator of JNK activity in PC12 cells, a myristoylated, gain-of-function Akt polypeptide (Myr Akt) was expressed by retrovirus-mediated gene transfer. Stimulation of JNK activity by serum withdrawal or UV irradiation in PC12 cell clones stably expressing Myr-Akt was inhibited approximately 95% or 50%, respectively, relative to control transfected PC12 cells. Phosphorylation of both JNKs and a proximal activator, MAP kinase kinase 4 (MKK4), in response to UV irradiation was inhibited in Myr-Akt-expressing PC12 cells. Furthermore, transient expression of Myr-Akt strongly inhibited cJun transactivation mediated by MEKK1 or MKK7-JNK3, a gain-of-function MKK7-JNK fusion protein. Interestingly, inhibited JNK activation in the Myr-Akt-expressing PC12 cells is associated with marked induction of JNK interacting protein-1 (JIP-1). We propose that negative regulation of the JNK pathway through Akt-dependent induction of specfic JIP proteins contributes to the antiapoptotic actions of Akt in neuronal cell types. PMID- 11107165 TI - TGF-beta 2 attenuates the injury-induced death of mature motoneurons. AB - The distributions of transforming growth factor-betas (TGF-betas) and their receptors suggest that the TGF-betas regulate motoneuron survival. This hypothesis was tested by avulsing the hypoglossal nerve of adult rats and perfusing either TGF-beta 2 or vehicle adjacent to the hypoglossal nucleus. By 4 weeks, half of the avulsed motoneurons had died. Infusion of 6 ng of TGF-beta 2 adjacent to the avulsed motor nucleus caused a significant attenuation of this death. This dose of TGF-beta 2 is low compared to that used with GDNF or BDNF in previous studies of avulsed motoneurons, indicating that TGF-beta 2 may be one of the more potent survival factors for adult motoneurons. TGF-beta 2 was, however, unable to prevent or reduce the axotomy-induced down regulation of choline acetyltransferase. Other motoneuron survival factors also have a narrow-spectrum of actions, suggesting that the homeostasis of motoneurons is regulated by a cocktail of growth factors with distinct but partially overlapping actions. PMID- 11107166 TI - Metabolic effects of 1-methyl-4-phenylpyridinium (MPP(+)) in primary neuron cultures. AB - Disruption of mitochondrial function has been proposed as an action of 1-methyl-4 phenylpyridinium (MPP(+)) that is responsible for its toxicity. In order to characterize effects of MPP(+) on energy metabolism in primary culture neurons, we monitored levels of several metabolites in cultured rat cerebellar granule cells exposed to MPP(+). The toxin produced a rapid concentration-dependent reduction in intracellular phosphocreatine (PCr), amounting to a 50-80% decrease within 30-60 min at 50 microM, that was maintained through the 1 week exposure interval examined. In contrast, ATP levels remained comparable to those of untreated neurons for approximately 4 days, at that time a 50% reduction in ATP was observed in association with a decrease in cell viability. Acute decreases in PCr were accompanied by increases in creatine such that the total creatine levels were maintained. Lactate levels in the culture medium were significantly increased (from 4.5 to 6.0 mM) within 6 hr after addition of MPP(+), with a concentration dependence similar to that observed for the reduction in PCr. Increased lactate production in the presence of MPP(+) coincided with a more rapid depletion of glucose in the culture medium. MPP(+) induced a rapid and sustained decrease in intracellular pH calculated from the creatine kinase equilibrium, and this acidification is considered primarily responsible for the observed decrease in PCr. These studies provide direct evidence that toxic concentrations of MPP(+) have acute effects on energy metabolism in primary culture neurons, consistent with an increased dependence on glycolysis to meet metabolic demand, but indicate that toxicity is not associated with overt, immediate failure to maintain cellular ATP. PMID- 11107167 TI - Motoneurons of the adult marmoset can grow axons and reform motor endplates through a peripheral nerve bridge joining the locally injured cervical spinal cord to the denervated biceps brachii muscle. AB - Reconnection of the injured spinal cord (SC) of the marmoset with the denervated biceps brachii muscle (BB) was obtained by using a peripheral nerve (PN) bridge. In 13 adult males, a 45 mm segment of the peroneal nerve was removed: one end was implanted unilaterally into the cervical SC of the same animal (autograft), determining a local injury, although the other end was either directly inserted into the BB (Group A) or, alternatively, sutured to its transected motor nerve, the musculocutaneous nerve (Group B). From 2-4 months post-surgery, eight out of the 10 surviving animals responded by a contraction of the BB to electrical stimulations of the PN bridge. All ten were then processed for a morphological study. As documented by retrograde axonal tracing studies using horse radish peroxidase or Fast Blue (FB), a mean number of 314 (Group A) or 45 (Group B) spinal neurons, mainly located close to the site of injury and grafting, re expressed a capacity to grow and extend axons into the PN bridge. Most of these regenerated axons were able to grow up to the BB and form or reform functional motor endplates. Many of the spinal neurons that were retrogradely labeled with FB simultaneously displayed immunoreactivity for choline acetyl-transferase and consequently were assumed to be motoneurons. Reinnervation and regeneration of the BB were documented by methods revealing axon terminals, endplates and myofibrillary ATPase activity. Our results indicate that motoneurons of the focally injured SC of a small-sized primate can, following the example of the adult rat, re-establish a lost motor function by extending new axons all the way through a PN bridge connected to a denervated skeletal muscle. PMID- 11107168 TI - Isolation and characterization of substance P-containing dense core vesicles from rabbit optic nerve and termini. AB - In neurons, neuropeptides and other synaptic components are transported down the axon to the synapse in vesicles using molecular motors of the kinesin family. In the synapse, these neuropeptides are found in dense core vesicles (DCVs), and, following calcium-mediated exocytosis, they interact with receptors on the target cell. We have developed a rapid, large-scale technique for purifying peptide containing DCVs from specific nuclei in the central nervous system. By using differential velocity gradient and equilibrium gradient centrifugation, neuropeptide-containing DCVs can be separated by size and density from optic nerve (ON) and its termini, the lateral geniculate nuclei and the superior colliculi. Isolated DCVs contain neuropeptides (substance P and brain-derived neurotrophic factor), synaptic vesicle (SV) membrane proteins (SV2, synaptotagmins, synaptophysin, Rab3 and synaptobrevin), SV-associated proteins (alpha-synuclein), secretory markers for DCVs previously isolated (secretogranin II), and beta-amyloid precursor protein. By using electron microscopic techniques, DCV were also visualized and shown to be immunoreactive for neuropeptides, neurotrophins, and SV membrane proteins. Because of the interesting group of physiological and potentially pathophysiological proteins associated with these vesicles; this isolation procedure, applicable to other CNS nuclei, should represent an important research tool. PMID- 11107169 TI - Ca(2+) dynamics in synaptosomes isolated from the squid optic lobe. AB - Synaptosomes from the optic lobes of squid (Loligo forbesi) were prepared by homogenization and allowed to settle onto glass coverslips. Synaptosomes were loaded with Ca(2+) sensitive dyes (Fura-2 AM, Calcium Green-1 AM and Calcium Green-5N AM), visualized by light microscopy and Ca(2+) sensitive fluorescence signals recorded and analyzed. With Fura-2, resting Ca(2+) was found to be 80 nM (n = 10, SEM 5.7). Addition of K(+) (30 mM), caffeine (3 mM) and thapsigargin (10 microM) evoked transient increases in cytoplasmic Ca(2+). Addition of BAPTA-AM (20 microM) decreased intrasynaptosomal free Ca(2+). Similar results were obtained with Calcium Green-1 AM but not with Calcium Green-5N AM. We conclude that synaptosomes from the squid optic lobe posses intact membranes and mechanisms to regulate intrasynaptosomal free [Ca(2+)], as well as caffeine sensitive Ca(2+) stores. The results of this study are discussed with respect to the role of Ca(2+) in presynaptic protein synthesis. PMID- 11107170 TI - Molecular mechanisms underlying human synovial sarcoma development. AB - Synovial sarcomas are rather common among soft-tissue tumors, occurring at any age but affecting mainly young adults. The vast majority of synovial sarcomas carries a t(X;18)(p11.2;q11.2) chromosomal translocation, in about one-third of the cases as the sole cytogenetic anomaly. Several studies have indicated that the t(X;18) translocation arises exclusively in synovial sarcomas, therefore being an excellent tool to diagnose this malignancy. The breakpoint-associated genes were recently isolated: SYT, from chromosome 18, and SSX1 and SSX2, both from the X chromosome. This discovery enabled the detection of SYT-SSX fusion transcripts by specific reverse transcriptase-polymerase chain reactions. This molecular genetics methodology has now been applied to numerous tumor samples and has led to the finding that, in contrast to tumors carrying SYT-SSX2 fusions, SYT SSX1-positive tumors more often exhibit a biphasic histology, show a higher proliferation rate, and are associated with a poorer clinical outcome. It has also been shown that the SYT and SSX proteins are localized in the nucleus, where they appear to play a role in transcriptional regulation, SYT as an activator of transcription and the SSX proteins as transcriptional repressors. It was also found that SYT interacts and colocalizes in the nucleus with the BRM protein, a transcriptional coactivator, and that the SSX proteins colocalize in the nucleus with polycomb group proteins, which are transcriptional corepressors. Together, these studies have provided mechanistic clues about how the SYT-SSX fusion proteins may trigger synovial sarcoma development. PMID- 11107171 TI - Cross-species color banding in ten cases of myeloid malignancies with complex karyotypes. AB - Cross-species color banding is a multiple-color fluorescence in situ hybridization (FISH) technique using probes developed from other animal species. Hybridization to human metaphases produces color banding patterns specific for each homologous chromosome pair. The technique has been evaluated in a complementary manner with G-banding and chromosome painting in a series of 10 myeloid malignancies with complex or unresolved karyotypes. Color banding detected the majority of chromosomal abnormalities, which had been identified by G-banding and in each case revealed chromosomal changes that G-banding had not identified. Painting was necessary to confirm these abnormalities due to the limitation of only seven colors in the color-banded karyotype. At the same time, painting fortuitously uncovered cryptic abnormalities in 6 of 10 cases that had not been detected by color banding. Insertions were visible by painting only. This study has demonstrated that in the analysis of complex karyotypes, the application of color banding revealed the involvement of the long arm of chromosome 3, indicating a poor risk, in two cases not identified by G-banding. Therefore, these techniques applied together have revealed cryptic chromosomal abnormalities with prognostic significance, which in some cases may have implications for patient management. PMID- 11107172 TI - Comparative genomic hybridization detects novel amplifications in fibroadenomas of the breast. AB - Comparative genomic hybridization analysis was performed for identification of chromosomal imbalances in 23 samples of fibroadenomas of the breast. Chromosomal gains rather than losses were a feature of these lesions. Only two cases with a familial and/or previous history of breast lesions had gain of 1q or 16q as the sole abnormality. The most frequently overrepresented segments were 5p14 (10/23 cases), 5q34-qter (6/23 cases), 13q32-qter (6/23 cases), 10q25-qter (5/23 cases), and 18q22 (4/23 cases). Some of these regions have previously been associated with breast carcinoma, but this study indicates that gain of these regions can also occur in benign breast lesions. Our findings may provide a basis for conducting further investigations to locate and identify genes associated with proliferation that may be involved in the early steps of tumorigenesis of the breast. PMID- 11107173 TI - Frequent activation of the beta-catenin-Tcf signaling pathway in nonfamilial colorectal carcinomas with microsatellite instability. AB - It has been reported that wild-type APC protein forms a complex with beta-Catenin and GSK3beta, inducing degradation of beta-Catenin in normal cells. Both beta Catenin and APC gene mutations have recently been shown to activate the same signaling pathway. Frequent mutations of beta-Catenin in hereditary nonpolyposis colorectal carcinomas have also been reported. It was, however, controversial whether the mutation of the beta-Catenin gene was frequent in nonfamilial colorectal carcinomas with high-frequency microsatellite instability (MSI-H). We analyzed the mutations of the APC and beta-Catenin genes in 56 nonfamilial colorectal carcinomas stratified according to the presence or absence of microsatellite instability (MSI). APC mutations were identified in 11 of 22 (50%) cases of MSI-H and 14 of 34 (41%) cases of microsatellite-stable (MSS)/low frequency microsatellite instability (MSI-L). In contrast, the frequency of beta Catenin mutations was significantly higher in MSI-H (6/22; 27%) than in MSS/MSI-L (1/34; 3%) (P = 0.01). beta-Catenin mutations were not detected in carcinomas with APC mutation. APC mutation occurred irrespective of MSI status. beta-Catenin mutation, however, occurred frequently in MSI-H carcinomas. Our data suggest that activation of the beta-Catenin-Tcf signaling pathway, through either beta-Catenin or APC mutation, frequently contributes to MSI-H nonfamilial colorectal carcinomas (17/22; 77%). PMID- 11107174 TI - Aberrant hypermethylation of the major breakpoint cluster region in 17p11.2 in medulloblastomas but not supratentorial PNETs. AB - Deletions of 17p have been consistently reported in up to 50% of medulloblastomas (MBs), and the major breakpoint interval has been localized to chromosome segment 17p11.2. Based on several reports linking aberrant DNA methylation and chromosomal disruption, we examined the methylation pattern in this region by employing restriction landmark genomic scanning (RLGS). Several CpG islands located in the major breakpoint cluster region were identified using a bacterial artificial chromosome (BAC) contig of the breakpoint region. A long-range methylation map was established for 20 MBs and 5 supratentorial primitive neuroectodermal tumors (stPNETs). Selected CpG islands were examined using Southern and bisulfite sequencing analysis. Aberrantly hypermethylated CpG islands in 17p11. 2 were found in 33% of MBs. Interestingly, one CpG island was methylated in MBs, but not in any of the examined stPNETs. A BAC clone covering three of the methylated CpG islands was partially sequenced in the search for a potential tumor suppressor gene. None of the expressed sequence tag sequences and full-length mouse/human cDNAs that were associated with aberrant methylation showed a change in expression levels due to methylation. The potential link between chromosomal instability in 17p11.2 and hypermethylation in this region is discussed. PMID- 11107175 TI - Establishment and characterization of chromosomal aberrations in human cholangiocarcinoma cell lines by cross-species color banding. AB - Cholangiocarcinoma (CC), a malignant neoplasm of the biliary epithelium, is usually fatal because of difficulty in early diagnosis and lack of availability of effective therapy. Furthermore, little is known about the genetics and biology of CC. Only a few reports concerning cytogenetic studies of CC have been published, and few cell lines have been established. We recently established four CC cell lines, designated as SCK, JCK, Cho-CK, and Choi-CK, and report the first application of cross-species color banding (RxFISH) and multiple chromosome painting for the characterization of the chromosomal rearrangements of these CC cell lines. Each cell line had unique modal karyotypic characteristics and showed a variable number of numerical and structural clonal cytogenetic aberrations. Chromosomes 3, 6, 7, 8, 12, 14, 17, and 18 were commonly involved in structural abnormalities. Homogeneously staining regions were determined in SCK and JCK, and double minute chromosomes were found in Cho-CK. The chromosomal aberrations of the four CC cell lines were effectively analyzed by RxFISH and FISH with multiple chromosome painting probes. The nonrandom rearrangements suggest candidate regions for isolation of genes related to CC. PMID- 11107176 TI - Multicolor fluorescence in situ hybridization with peptide nucleic acid probes for enumeration of specific chromosomes in human cells. AB - In previous studies, we showed that peptide nucleic acid (PNA) probes have significant advantages over conventional synthetic RNA or DNA probes in FISH procedures for detecting telomeric and trinucleotide repeat sequences. Here, we report that directly labeled PNA probes recognizing chromosome-specific repeat sequences are also powerful tools for detecting and enumerating specific chromosomes in interphase and metaphase cells. This is illustrated by multicolor FISH experiments with cells from normal individuals and patients with numerical sex chromosome aberrations. PMID- 11107177 TI - Transcriptional activation of short interspersed elements by DNA-damaging agents. AB - Short interspersed elements (SINEs), typified by the human Alu repeat, are RNA polymerase III (pol III)-transcribed sequences that replicate within the genome through an RNA intermediate. Replication of SINEs has been extensive in mammalian evolution: an estimated 5% of the human genome consists of Alu repeats. The mechanisms regulating transcription, reverse transcription, and reinsertion of SINE elements in genomic DNA are poorly understood. Here we report that expression of murine SINE transcripts of both the B1 and B2 classes is strongly upregulated after prolonged exposure to cisplatin, etoposide, or gamma radiation. A similar induction of Alu transcripts in human cells occurs under these conditions. This induction is not due to a general upregulation of pol III activity in either species. Genotoxic treatment of murine cells containing an exogenous human Alu element induced Alu transcription. Concomitant with the increased expression of SINEs, an increase in cellular reverse transcriptase was observed after exposure to these same DNA-damaging agents. These findings suggest that genomic damage may be an important activator of SINEs, and that SINE mobility may contribute to secondary malignancy after exposure to DNA-damaging chemotherapy. PMID- 11107178 TI - Early genetic events in HPV immortalised keratinocytes. AB - Cancer of the uterine cervix (CaCx) is the second most common cancer in women worldwide. More than 99% of all cervical cancers contain high-risk human papillomaviruses (HPVs), with type 16 predominating. HPV infection alone is not sufficient for neoplastic progression; the HPV-infected cell must undergo additional genetic changes. Cytogenetic analysis of CaCx has been limited due to difficulties in obtaining good-quality banded chromosome preparations. Oncogenic HPVs immortalise primary genital keratinocytes in vitro, and evidence suggests that the molecular genetic and cytogenetic abnormalities observed in HPV immortalised cells reflect the in vivo changes. Therefore, these lines represent suitable models for HPV-induced carcinogenesis. We have used both spectral karyotyping (SKY) and multiplex-FISH (M-FISH) analysis to identify karyotypic changes in HPV-16 immortalised keratinocyte cell lines and established CaCx lines. SKY and M-FISH identified chromosomal abnormalities in all cell lines examined, with a translocation of chromosome 10 or i(10q) occurring in 9 of the 12 cell lines investigated. Further studies with chromosome 10 band-specific probes identified the translocation event as involving 10q with the breakpoint at 10p11.2 in some cell lines or 10q11.2 in others. The pericentric region of chromosome 10 is known to contain duplicated sequences flanking the centromeric satellites. The duplicated sequences contain many zinc finger transcription factor encoding genes and disruption of these in HPV immortalised cell lines may alter the transcription with consequences for both cellular and viral gene expression. PMID- 11107179 TI - Correlation of histologic subtypes and replication error phenotype with comparative genomic hybridization in gastric cancer. AB - To characterize phenotypic and genotypic changes in gastric cancer (GC), DNA copy number aberrations (CNAs) were assessed in 53 tumors using comparative genomic hybridization (CGH) and correlated with clinicopathologic characteristics and status of TP53 and replication error (RER). The number of CNAs per tumor was 6.8 (gain 5.3, loss 1.5), and the number of changes was significantly higher in tumors with advanced stage, TP53 mutation, and without RER than in those with early stage (7.7 vs. 3.0), no TP53 mutations (12.4 vs. 4.8) or RER phenotype (8.2 vs. 2.6). Frequent abnormalities included gains on chromosomal arms 8q (43%), 6q (26%), 11q (26%), 13q (24%), 7p (23%), 17q (23%), and 20q (23%), and losses on chromosomal arms 16q (26%), 19p (23%), 5q (19%), 3p (15%), 4q(15%), and 1p (15%). Advanced GC demonstrated a higher prevalence of gains of 8q (51% vs. 10%, P < 0.05) and loss of 16q (33% vs. 0%, P < 0.05) than early GC. Gains on 8q (64% vs. 20%, P < 0.05), 17q (39% vs. 4%, P < 0.05) and losses on 3p (25% vs. 4%, P = 0.05) and 5q (32% vs. 4%, P < 0.05) were higher in intestinal GC than in diffuse GC. On the other hand, gains on 13q were more common in the diffuse type (40% vs. 11%, P < 0.05). As compared with noncardia cancer, cardia cancer showed more gains on 7p (58% vs. 12%, P < 0.05) and 20q (58% vs. 12%, P < 0.05) and more losses on 4q (50% vs. 5%, P < 0.05). The finding of histology-related aberrations and the combination of CGH and molecular data thus provide additional evidence suggesting genetic heterogeneity of GC. PMID- 11107180 TI - MYCN amplification and 17q in neuroblastoma: evidence for structural association. AB - MYCN oncogene amplification in neuroblastoma is statistically associated with gain of chromosome segment 17q21-qter. In neuroblastoma cell lines and primary tumors with MYCN amplification in the form of homogeneously staining regions (hsrs), juxtaposition of chromosome 17 material with MYCN sequences has occasionally been reported, raising the possibility of a physical affinity between MYCN and chromosome arm 17q. We used FISH to test for association between chromosome 17 segments and MYCN in eight neuroblastoma cell lines and two neuroblastoma primary tumors known to include hsrs. Evidence of an association was found in the chromosomes of both primary tumors; in one, a MYCN hsr was inserted into a structurally abnormal chromosome 17, in the other, an hsr in 16p was shown to be flanked by 17 material. In cell line NCG, hsrs in 4q and 16p were flanked by 17q material. These observations confirm the juxtaposition of 17q material with MYCN sequences in some neuroblastomas, and imply that there may be a physical or functional relationship between these two features in MYCN amplified neuroblastoma. PMID- 11107181 TI - Contiguous arrangement of p45 NFE2, HnRNP A1, and HP1 alpha on mouse chromosome 15 and human chromosome 12: evidence for suppression of these genes due to retroviral integration within the Fli-2 locus. AB - Fli-2 is a common site of proviral integration in multistage erythroleukemia cells induced by Friend murine leukemia virus (F-MuLV) or the polycythemia strain of Friend leukemia virus (FV-P). Previously, we reported that integration of Friend virus into the Fli-2 locus in CB3, an erythroleukemia cell line that harbors a homozygous inactivation of the Fli-2 locus, results in the loss of expression of two genes encoding the 45-kDa subunit of the erythroid-specific nuclear factor p45 NFE2 and the splicing factor HnRNP A1. Here, we report the identification of a third gene, Heterochromatin protein 1 (HP1alpha, also known as CBX5), which is located downstream of HnRNP A1, and p45 NFE2. Northern blot analysis revealed that the expression of HP1alpha, along with p45 NFE2 and HnRNP A1, is either undetectable or substantially reduced in CB3 cells, suggesting that HP1alpha expression is also regulated by proviral insertion within the Fli-2 locus in CB3 cells. Because p45 NFE2 was previously mapped to mouse chromosome 15, our results demonstrate that HP1alpha and HnRNP A1 are also located on mouse chromosome 15 and that the p45 NFE2, HnRNP A1, and HP1alpha genes are arranged contiguously. Contiguous arrangement of these three genes was also detected in man; this consequently localizes HP1alpha to human chromosome band 12q13. PMID- 11107183 TI - Comparative genomic hybridization in pineal parenchymal tumors. AB - Nine pineal parenchymal tumors were studied by comparative genomic hybridization. These consisted of three pineocytomas (WHO grade II), three pineal parenchymal tumors of intermediate differentiation (WHO grade III), and three pineoblastomas (WHO grade IV). An average of 0 chromosomal changes per pineocytoma, 5.3 per pineal parenchymal tumor of intermediate differentiation (3.3 gains vs. 2.0 losses), and 5.6 per pineoblastoma (2.3 gains vs. 3.3 losses) were found. The most frequent DNA copy number changes among pineal parenchymal tumors of intermediate differentiation and pineoblastomas were gains of 12q (3/6 cases), 4q, 5p, and 5q (2/6 cases each), as well as losses of 22 (4/6 cases), 9q, and 16q (2/6 cases each). Among pineal parenchymal tumors of intermediate differentiation, the most common chromosomal imbalances were +4q, +12q, and -22 (2/3 cases each), and in pineoblastomas -22 (2/3 cases). Five high level gains were identified, all of them in pineoblastomas; these were found on 1q12-qter, 5p13.2-14, 5q21-qter, 6p12-pter, and 14q21-qter. Clinically, all patients with pineocytomas and pineal parenchymal tumors of intermediate differentiation were alive after a mean observation time of 142 and 55 months, respectively, whereas all patients with pineoblastomas had died after an average of 17 months. Our findings suggest that pineal parenchymal tumors of intermediate differentiation are cytogenetically more similar to pineoblastomas and prognostically more similar to pineocytomas. Furthermore, imbalances in higher-grade pineal parenchymal tumors mainly affect gains of 12q and losses of chromosome 22. PMID- 11107182 TI - Absence of post-transcriptional RNA modifications of BCL10 in human malignant mesothelioma and colorectal cancer. AB - The BCL10 gene, located at 1p22, has been implicated in a number of human malignancies, including malignant mesotheliomas (MMs) and colorectal carcinomas. Subsequent reports, however, have revealed an absence of BCL10 mutations in genomic DNA from such tumors. It has been proposed that some abnormalities of this gene may be found only in RNA and not in genomic DNA, suggesting that BCL10 may be mutated post-transcriptionally, rather than at the genomic level. To explore this possibility, we performed SSCP mutation analysis and direct sequencing of cDNA from 17 MM cell lines displaying LOH in 1p22, 12 MM tumor specimens, and 11 colon carcinoma cell lines. SSCP revealed several different band shifts in these samples. The nucleotide changes observed in the cDNA samples were also seen in matched genomic DNA and corresponded to known polymorphisms in the general population. Thus, we conclude the BCL10 mutations are absent at the cDNA level, and that this gene does not undergo "molecular misreading." Since BCL10 also does not possess mutations at the genomic DNA level, it can be ruled out as a gene involved in the pathogenesis of MM and colorectal cancer. PMID- 11107184 TI - Chromosome arm-specific multicolor FISH. AB - Several systems for 24-color fluorescence in situ hybridization (FISH) have been developed and applied to karyotyping and detection of chromosomal abnormalities. We have developed a 42-color multicolor FISH (mFISH) technique (armFISH), which permits the detection of chromosomal aberrations at the resolution of chromosome arms. The armFISH uses a commercially available mFISH reagent kit (24XCyte, MetaSystems GmbH) supplemented with a set of differentially labeled chromosome arm-specific painting probes (arm-kit, comprising either p- or q-arms of all human chromosomes, except the p-arm of the acrocentric- and Y chromosomes). The mFISH-probe cocktail and the arm-kit are combined and hybridized together to metaphase chromosomes. The armFISH is analyzed in two steps; first, the conventional mFISH image analysis is performed, followed by the arm-kit analysis to reveal the chromosome arms involved. The examples demonstrate the utility of armFISH in defining chromosomal rearrangements of human cancers. PMID- 11107185 TI - Recurrent chromosome alterations in hepatocellular carcinoma detected by comparative genomic hybridization PMID- 11107186 TI - Simian virus-40 sequences are a negative prognostic cofactor in patients with malignant pleural mesothelioma PMID- 11107187 TI - Overview: pediatric epilepsy. PMID- 11107188 TI - Neonatal seizures: early-onset seizure syndromes and their consequences for development. AB - The determination of the developmental consequences of seizure syndromes in the neonate is based upon a number of factors which include: understanding of the clinical and electroencephalographic (EEG) features of neonatal seizures; current theories of the mechanisms by which neonatal seizures are generated; a current classification of neonatal seizures; potential etiologic and risk factors for seizures; and therapies. In addition, different seizure types, mechanisms of generation and etiologies of cerebral dysfunction may vary with conceptional age of the infant. There are a few distinct neonatal epileptic syndromes, which are rare, have been well described: benign neonatal convulsions; benign neonatal familial convulsions; early myoclonic encephalopathy and early infantile epileptic encephalopathy. The prognosis for the first two is relatively good while the outcome for the other two with encephalopathy is catastrophic. However, the majority of neonatal seizures occur as acute, reactive events in association with a wide range of etiologic factors. These etiologic factors, as well as those of the more traditionally defined syndromes, are the main determinants of eventual developmental outcome of neonates who experience seizures. Although experimental data suggests that some epileptic seizures eventually may have physiological, histological, metabolic, or behavioral consequences, there is yet direct evidence in humans to suggest that the occurrence of seizures themselves in the neonate is the main determinant of long-term outcome. PMID- 11107189 TI - Age-dependent consequences of seizures: relationship to seizure frequency, brain damage, and circuitry reorganization. AB - Seizures in the developing brain pose a challenge to the clinician. In addition to the acute effects of the seizure, there are questions regarding the impact of severe or recurrent seizures on the developing brain. Whether provoked seizures cause brain damage, synaptic reorganization, or epilepsy is of paramount importance to patients and physicians. Such questions are especially relevant in the decision to treat or not treat febrile seizures, a common occurrence in childhood. These clinical questions have been addressed using clinical and animal research. The largest prospective studies do not find a causal connection between febrile seizures and later temporal lobe epilepsy. The immature brain seems relatively resistant to the seizure-induced neuronal loss and new synapse formation seen in the mature brain. Laboratory investigations using a developmental rat model corresponding to human febrile seizures find that even though structural changes do not result from hyperthermic seizures, synaptic function may be chronically altered. The increased understanding of the cellular and synaptic mechanisms of seizure-induced damage may benefit patients and clinicians in the form of improved therapies to attenuate damage and changes induced by seizures and to prevent the development of epilepsy. PMID- 11107190 TI - Developmental seizures induced by common early-life insults: short- and long-term effects on seizure susceptibility. AB - The immature brain is highly susceptible to seizures induced by a variety of insults, including hypoxia, fever, and trauma. Unlike early life epilepsy associated with congenital dysplasias or genetic abnormalities, insults induce a hyperexcitable state in a previously normal brain. Here we evaluate the epileptogenic effects of seizure-inducing stimuli on the developing brain, and the age and regional specificity of these effects. PMID- 11107191 TI - Cellular abnormalities and synaptic plasticity in seizure disorders of the immature nervous system. AB - The nervous system has an enhanced capacity to generate seizures during a restricted phase of postnatal development. Studies in animals and particularly in in vitro brain slices from hippocampus and neocortex have been instrumental in furthering an understanding of the underlying processes. Developmental alterations in glutaminergic excitatory synaptic transmission appear to play a key role in the enhanced seizure susceptible of rodents during the second and third week of life. Prior to this period, the number of excitatory synapses is relatively low. The scarcity of connections and the inability of the existing synapses to release glutamate when activated at high frequencies likely contribute importantly to the resistance of neonates to seizures. However, at the beginning of week 2, a dramatic outgrowth of excitatory synapses occurs, and these synapses are able to faithfully follow activation at high frequencies. These changes, coupled with the prolonged nature of synaptic potentials in early life, likely contribute to the ease of seizure generation. After this time, seizure susceptibility declines, patterns of local synaptic connectivity remodel, and some synapses are pruned. Concurrently, the duration of excitatory postsynaptic potentials shortens due at least in part to a switch in the subunit composition of postsynaptic receptors. Other studies have examined the mechanisms underlying chronic epilepsy initiated in early life. Models of both cortical dysplasia and recurrent early-life seizures suggest that alterations in the normal development of excitatory synaptic transmission can contribute importantly to chronic epileptic conditions. In the recurrent early-life seizure model, abnormal use-dependent selection of subpopulations of excitatory synapses may play a role. In experimental cortical dysplasia, alterations in the molecular composition of postsynaptic receptor are observed that favor subunit combinations characteristic of infancy. PMID- 11107192 TI - Cortical malformations and epilepsy. AB - Brain malformations, resulting from aberrant patterns of brain development, are highly correlated with childhood seizure syndromes, as well as with cognitive disabilities and other neurological disorders. The structural malformations, often referred to as cortical dysplasia, are extremely varied, reflecting diverse underlying processes and critical timing of the developmental aberration. Recent studies have revealed a genetic basis for many forms of dysplasia. Gene mutations responsible for such common forms of dysplasia as lissencephaly and tuberous sclerosis have been identified, and investigators are beginning to understand how these gene mutations interrupt and/or misdirect the normal developmental pattern. Laboratory investigations, using animal models of cortical dysplasia, are beginning to elucidate how these structural malformations give rise to epilepsy and other functional pathologies. PMID- 11107193 TI - Epilepsy genes: the link between molecular dysfunction and pathophysiology. AB - Our understanding of the genetic basis of epilepsy is progressing at a rapid pace. Gene mutations causing several of the inherited epilepsies have been mapped, and several more are likely to be added in coming years. In this review, we summarize the available information on the genetic basis of human epilepsies and epilepsy syndromes, emphasizing how genetic defects may correlate with the pathophysiological mechanisms of brain hyperexcitability. Mutations leading to epilepsy have been identified in genes encoding voltage- and ligand-gated ion channels (benign familial neonatal convulsions, autosomal dominant nocturnal frontal lobe epilepsy, generalized epilepsy with febrile seizures "plus"), neurotransmitter receptors (Angelman syndrome), the molecular cascade of cellular energy production (myoclonic epilepsy with ragged red fibers), and proteins without a known role in neuronal excitability (Unverricht-Lundborg disease). Gene defects can lead to epilepsy by altering multiple and diverse aspects of neuronal function. PMID- 11107194 TI - Behavioral aspects of epilepsy in children with mental retardation. AB - Epilepsy and mental retardation, two relatively common childhood conditions, are both associated with a wide range of behavioral disorders. This article reviews the behavioral disturbances found in children with epilepsy, mental retardation, and both conditions. The behavioral disturbances found in children with epilepsy are associated with seizure-related, cognitive, developmental, and psychosocial factors. Although children with mental retardation also demonstrate a broad spectrum of behavioral disturbances, children with specific mental retardation syndromes have better-defined patterns of psychopathology. The presence of epilepsy and mental retardation seems to increase the severity of psychopathology. Further studies are needed, however, to define better the interaction of these two conditions and how they impact the behavior of children. PMID- 11107195 TI - Epilepsy and epileptiform EEG: association with autism and language disorders. AB - The relationship between epilepsy, language, behavior, and cognition is not well understood. Developmental and acquired disabilities such as autistic spectrum disorders, Landau-Kleffner Syndrome, electrical status epilepticus in sleep, and developmental dysphasias have been associated with epileptiform abnormalities. These disorders share many common features and raise important questions regarding this intricate relationship. This article reviews these disorders and discusses the proposed interaction between epileptiform abnormalities and cognitive dysfunction. Diagnostic and treatment issues will also be reviewed. PMID- 11107196 TI - Treatment of epilepsy in the multiply handicapped. AB - The medical management of epilepsy in the multi-handicapped patient requires careful evaluation, classification, and pharmacologic treatment. It is estimated that 20-40% of patients with mental retardation and cerebral palsy have epilepsy. This review reports the clinical trial data and personal experience related to the use of newer AEDs in the chronic management of epilepsy syndromes in children and adults, as well as information available on the treatment of seizures in individuals with mental retardation and associated handicaps. Furthermore, clusters of seizures, prolonged seizures and status epilepticus are more commonly seen in the multiply handicapped and mentally retarded population and require special attention. The new antiepileptic drugs felbamate, gabapentin, lamotrigine, levetiracetam, oxcarbazepine, tiagabine, topiramate, vigabatrin and zonisamide show specific advantage in some multiply handicapped patients, be it for seizure control or medication tolerance. Furthermore, new modalities of treatment for prolonged seizures allow better efficacy both outside of hospital and within hospital facilities. The treatment of epilepsy in multi-handicapped and retarded adults and children has significantly advanced in the past few years, and much of this improvement can be attributed to improved knowledge and monitoring of new antiepileptic drugs. Conventional anticonvulsants remain first line therapy for most clinicians, but newer AEDs must broaden the therapeutic option and do allow improved therapy for some multiply handicapped patients. PMID- 11107197 TI - In vitro validation of a new approach for quantitating regurgitations using proximal isovelocity surface area. AB - The present work has been designed to validate the calculation of the effective regurgitant orifice (ERO) area with the use of a new formula that takes into account the velocity profile (V(r) vs r) and that is insensitive to errors in the determination of the position of the orifice. Assuming a hemispheric model, ERO = 2 pi r(2). V(r)/V(o) (with V(o) = velocity at the orifice) and (V(o)/V(r))(0.5) = (2 pi/ERO)(0.5) r. Thus, the slope of the corresponding linear regression allows ERO to be calculated as: ERO = 2 pi/slope(2). This approach was tested in vitro in pulsatile conditions on circular, conical, and slit-like orifices. The calculated ERO was compared with the actual jet cross sectional area derived from the transverse velocity profile at the jet origin. For the purpose of comparison, the "classical" ERO was calculated for all the configurations, angulations, and threshold velocities. The relationship between (V(o)/V(r))(0.5) was linear (r > 0.98) over a wide range of velocities. The nonhemispheric components were found to modify the constant and not the slope. The mean variation of the calculated ERO was 6.5%. The correlation between the calculated and the actual ERO was very close (>0.97) with slope equal to 0.96. By comparison with the new method, the classical formula gave an underestimation of the ERO that dramatically increased when studying the flow closer to the orifice or in the case of error on the measurement of r. In conclusion, a method using velocity profiles instead of isolated values improves the accuracy of the proximal isovelocity surface area (PISA) method for measuring the ERO. PMID- 11107198 TI - Echocardiographic imaging of stentless aortic valve prostheses. AB - Homografts and stentless xenografts are increasingly used in aortic valve surgery. Echocardiography technicians and cardiologists have to know what they will find when performing an echo-Doppler examination in patients who received a stentless valve. We therefore evaluated echocardiographic images of 74 patients who received a Freestyle stentless bioprosthesis with three techniques and a follow-up of 2 years in two high-volume hospitals. Of the patients studied, 81% were operated using the subcoronary technique, 12% using the root-inclusion technique, and 7% using the full-root technique. RESULTS: Transvalvular gradients across the stentless valves were low: 8.0 mmHg when implanted with the subcoronary technique, 8.2 +/- 5.1 mmHg using the root-inclusion technique, and 6.5 mmHg using the full-root technique. Trivial aortic insufficiency (grade 1) was observed in 10.7% of the patients (8.9% for the subcoronary technique, 13% for the root-inclusion technique, and 0% for the full-root technique). When the bioprosthesis was implanted using the subcoronary technique or the root-inclusion technique, the prosthesis was placed inside the recipient aortic root. Using these techniques, a lumen between the double layer of the xenograft and the aortic wall could be observed. With the root-replacement technique, the porcine root became the most proximal part of the ascending aorta. As the native aortic wall was removed, in most cases, no double lumen could be observed with imaging of the ascending aorta. PMID- 11107199 TI - Reproducibility of left ventricular measurements with acoustic quantification: the influence of training. AB - Acoustic quantification (AQ) of two-dimensional (2-D) echocardiograms provides online estimation of left ventricular (LV) size and function. However, edge detection with AQ is influenced by gain settings and is therefore operator dependent. Our purpose was to compare AQ and conventional 2-D echo measurements of LV size and function obtained by different operators and to evaluate the influence of training on these measurements. A cardiac sonographer without previous experience with the AQ system was trained by an experienced operator. Twenty-two normal males (age, 28 +/- 4 years) participated in the study. Images were recorded with conventional 2-D and AQ echo from the short-axis and apical four-chamber views. During the initial training period, five subjects were imaged by the sonographer under the supervision of the trainer. At the initial study session, 12 subjects were imaged independently by the two operators. Following a second training period with five different subjects, the same initial 12 subjects were again imaged at a second study session. LV cavity areas were traced from the conventional 2-D echocardiograms and measured from the AQ waveforms at end diastole and end-systole. Volumes were calculated using the single-plane area length method. Ejection fraction (EF) was calculated from volumes. Reproducibility was determined by comparing the variability of AQ and conventional 2-D echo measurements obtained at the two sessions. A second training session reduced the operator variability only of the short-axis end diastolic area measurement (17 +/- 11% vs 6 +/- 5%, P < 0.025). We conclude that a single training session may be adequate for the reproducible estimation of ventricular volumes with the AQ method. PMID- 11107200 TI - A comparison of regional myocardial velocity information derived by pulsed and color Doppler techniques: an in vitro and in vivo study. AB - The objective was to compare velocity information derived from either a tissue mimicking phantom or normal contracting myocardium by both pulsed wave and color Doppler myocardial imaging (PWDMI and CDMI). Both CDMI and PWDMI allow quantitative assessment of regional myocardial contraction and relaxation velocities, but their potential clinical applications have not yet been investigated. Moreover, no information is available as to whether they can be used interchangeably for regional velocity assessment. For the in vitro study, a rotating, circular-shaped, tissue-mimicking sponge driven by a motor at speeds of 15, 30, 60, 90 rpm was used to derive velocity data from the same eight points of interest by using PWDMI or CDMI techniques. For the in vivo study, 25 normal subjects were examined at rest using parasternal and apical approaches. Velocity profiles were derived from the same 26 areas of interest (18 left ventricular segments, 3 right ventricular segments, and 5 measurement points for the tricuspid and mitral annuli) for each technique. Peak maximal velocities were detected by PWDMI and peak mean velocities were measured using CDMI. The results of the in vitro study phantom showed excellent correlation (r = 0.99, P < 0. 001) and satisfactory agreement (0.04 cm/sec; 3.3 cm/sec) between both Doppler techniques. PWDMI velocities were higher than CDMI velocities by up to 20% and overestimated the real velocity value (0. 37 +/- 0.29 cm/sec) while CDMI underestimated predicted velocity by 1.35 +/- 0.36 cm/sec. Good correlation (r = 0.87, P < 0.001), but poor agreement (-2.1 cm/sec; 5.4 cm/sec) was shown in vivo for all segments with regard to peak systolic and diastolic velocities. Both Doppler techniques cannot be used interchangeably for comparing peak velocities in the clinical situation. However, with adequate temporal resolution, they can be used interchangeably for velocity profile recording and for timing of events. PMID- 11107201 TI - Left ventricular inflow normal or pseudonormal. A new echocardiographic method: diastolic change of left atrial diameter. AB - Mitral flow Doppler study has been used to evaluate left ventricle (LV) diastolic function. Through its use, greater A wave than E wave, pseudonormal pattern, and restrictive pattern were observed progressively in patients with more LV diastolic dysfunction. Differentiation of normal or pseudonormal mitral flow is very important. In this study, left atrium (LA) diameter change during diastole was used as a new method for the differentiation of normal and pseudonormal mitral flow. METHOD: Sixty-eight patients (30 men, 38 women; mean age 53 +/- 13 years) with echocardiographically determined diastolic dysfunction and 60 healthy volunteers (36 men, 24 women; mean age 49 +/- 12 years) were included in the study. Mitral flow E/A ratio, isovolumetric relaxation time (IVRT), and deceleration time (DT) of E wave were used for determination of the diastolic dysfunction. Thirty of 68 diastolic dysfunction patients had A>E wave, 20 had pseudonormal mitral flow pattern, and 18 had restrictive mitral flow pattern. Left parasternal long-axis echocardiographic window was used for the measurement of LA diameter. Left atrium emptying fraction (LAEF) was defined as ratio of end diastolic LA diameter to end-systolic diameter. RESULTS: LAEF was found 0.69 +/- 0.01 (mean +/- SE) in the control group, 0.76 +/- 0.01 in the A>E group (P < 0.05, control vs A > E group), 0.83 +/- 0. 05 in the pseudonormal pattern group (P < 0.05, control vs pseudonormal pattern group), and 0.87 +/- 0.01 in the restrictive pattern group (P < 0.001, control vs restrictive pattern group). CONCLUSION: (1) LV diastolic dysfunction reduces the filling of LA content to the LV during diastole; (2) LA diameter changes during diastole as a new and practical method for the differentiation of the normal-pseudonormal mitral flow pattern. PMID- 11107202 TI - Restrictive left ventricular filling pattern after myocardial infarction: significance of concomitant preserved systolic function. AB - Restrictive left ventricular (LV) filling identifies a high risk subgroup, following myocardial infarction (MI). The extent and significance of systolic dysfunction in this group is not clear. The aim of our study was to determine the incidence and extent of systolic dysfunction in patients with restrictive filling and nonrestrictive filling examining the prognostic implications. Doppler parameters of LV diastolic function were measured in 102 post-MI subjects within 4 days. Restrictive filling was defined as the presence of E:A ratio > 2 or E:A ratio 1-2 with MDT < or = 140 msec. Follow-up was to a median of 11 months. Restrictive filling (group A) was found in 19 (19%) of 102 patients. Patients with this pattern were more likely to have systolic dysfunction than those without (group B); 63% and 35%, respectively, P = 0.024. Eight (42%) of 19 patients in group A had relatively preserved systolic function. At 11 months 14 patients had developed heart failure (HF), 6 in group A (32%) and 8 in group B (10%), P = 0.012. There were two deaths (11%) in group A and 7 (8%) in group B, P = ns. Seven (88%) of 8 patients in group A with relatively preserved systolic function were alive and free of heart failure at follow-up compared to 4 of 11 patients (36%) in group A with systolic dysfunction (P = 0.026). Restrictive filling can be associated with relatively preserved systolic function after MI and these patients have a relatively good outcome. Patients with restrictive filling post-MI are a heterogeneous group emphasizing the evaluation of both systolic and diastolic function. PMID- 11107203 TI - Determination of left ventricular mass by real-time three-dimensional echocardiography: in vitro validation. AB - Twenty-one explanted fixed hearts (14 dogs and 7 pigs) were examined to validate newly developed real-time three-dimensional (RT3D) echocardiography for measurement of left ventricular (LV) mass in vitro and to compare its accuracy and variability with those of conventional echocardiographic measurements. There was an excellent correlation and high degree of agreement for the determination of LV mass between RT3D echocardiography and true mass measurement (r = 0.98; standard error of the estimate [SEE] = 7.3 g; absolute difference [AD] = 2.8 g; y = 1.00 x -4.0, interobserver variability; 5.0%). The conventional echocardiographic methods yielded weaker correlations, larger standard errors, and interobserver variability (area-length method: r = 0.90; SEE = 13.3 g; AD = 13.2 g; 13.3 % / truncated ellipsoid method: r = 0.91; SEE = 14.7 g; AD = 10.5 g; 7. 9% / M-mode: r = 0.91; SEE = 16.2 g; AD = 9.4 g; 15.3%). Determination of LV mass by RT3D echocardiography has a high degree of accuracy and is superior to conventional one- and two-dimensional echocardiographic methods. PMID- 11107204 TI - Diagnosis of coronary artery disease in patients with permanent cardiac pacemaker by dobutamine stress echocardiography or exercise thallium-201 myocardial tomography. AB - This study evaluated the use of dobutamine stress echocardiography and exercise thallium-201 myocardial computed tomography (CT) in the diagnosis of coronary artery disease (CAD) in patients with permanent transvenous pacemaker with the electrode implanted in the right ventricle (RV). Twenty-nine consecutive patients with pacemaker underwent dobutamine stress echocardiography, exercise thallium 201 myocardial CT, and coronary arteriography over a period of 8 +/- 1 days. None of these patients had suffered a myocardial infarction (MI). The cardiac rhythm of every patient was electrically paced during echocardiography and tomography. Sixteen (55%) patients showed CAD on angiography (stenosis > or = 50% of the luminal diameter of a major epicardial vessel). The detection sensitivity for CAD was 94% for the tomography and 88% for the echocardiography (P = NS). The difference between the sensitivities of the two techniques in detecting CAD based on the affected coronary artery was not statistically significant. Of the 13 patients without CAD, tomography showed a positive result in nine cases, i.e., a specificity of 31%, whereas echocardiography showed a positive result in only one case, i.e., a specificity of 92% (P < 0. 01). Exercise thallium-201 myocardial computed tomography produces an increased rate of false-positive results in patients with permanent transvenous cardiac pacemaker (PCP) implanted in the right ventricle (RV). Dobutamine stress echocardiography can thus be used to reduce considerably the level of false-positive results in these patients and still retain a detection sensitivity for CAD equal to that of myocardial tomography. PMID- 11107205 TI - Evaluation of ventricular septal defect repair using intraoperative transesophageal echocardiography: frequency and significance of residual defects in infants and children. AB - Intraoperative transesophageal echocardiography (IOTEE) is commonly used to assess for residual defect and the need to return to bypass after repair of ventricular septal defect (VSD). The frequency and significance of residual septal defects as noted on IOTEE has not been well defined. We evaluated the frequency of residual VSD via IOTEE and the relationship between size of a residual VSD and rate of reoperation. In addition, we looked at the relationship between the presence of a residual VSD via IOTEE and the presence of residual VSD at follow-up transthoracic echocardiography (TTE). Residual VSD was measured via the largest width of the Doppler color jet diameter originating at the left ventricular septal surface. Of the 294 patients evaluated with IOTEE after VSD repair, one-third had a residual defect by IOTEE Doppler color flow mapping. Two thirds of these defects closed spontaneously on TTE by the time of hospital discharge. There was no difference in frequency of residual VSD between simple (VSD closure alone, n = 90) and complex (VSD with associated lesions, n = 204) repair. Return to bypass with immediate reoperation was undertaken in nine patients, all of whom had significant shunt via oximetry (Qp/Qs > 1.5:1.0). All had residual VSD color jet diameters > 3 mm. Seven patients had residual color jet equal to 3 mm; however, hemodynamic studies did not reveal a significant shunt and none of these had reoperation. Seven patients with no VSD or < 3 mm residual VSD via had late reoperation to close residual VSD at 4 days to 5 months after initial operation. These were due to patch dehiscence or development of an "intramural" VSD in patients with conotruncal anomaly. A residual defect on IOTEE color Doppler measuring > or = 4 mm predicts the need for immediate reoperation, while a 3 mm defect may be significant and requires additional intraoperative hemodynamic evaluation. The majority of small defects noted on IOTEE are not present at discharge TTE. Patients with conotruncal defect repair should be followed closely for development of late significant "intramural" defects. PMID- 11107206 TI - Aortic valve endocarditis due to Staphylococcus capitis. AB - Native valve endocarditis due to Staphylococcus capitis is uncommon and is usually managed conservatively with good outcome. Of the nine previously reported cases there has been only one mortality. We report a case of native aortic valve endocarditis due to S capitis in an elderly diabetic that had a fatal outcome despite appropriate therapy with antibiotics. A review the literature is also presented. PMID- 11107207 TI - Transesophageal echocardiographic diagnosis of thrombus in accessory lobes of the left atrial appendage. AB - Fifty-four percent of left atrial appendages have two lobes. The number ranges from one to four lobes. We describe three patients with accessory lobes of the left atrial appendage studied with multiplanar transesophageal echocardiography (TEE). In one patient there was evidence of thrombi in the accessory lobe. PMID- 11107208 TI - Interventricular septal hydatid cyst. AB - In this report we describe an unusual case of cardiac echinococcus located in the interventricular septum invaded by a cystic mass. It was demonstrated by using transthoracic echocardiography (TTE) and confirmed with magnetic resonance imaging (MRI). Surgical excision (cystopericystectomy) was performed on the patient as a curative therapy. Early recurrence was observed despite additional medical therapy with albendazole. PMID- 11107209 TI - Intramyocardial metastatic renal cell cancer. PMID- 11107210 TI - Quadricuspid aortic valve. PMID- 11107211 TI - Fibrotic aortic stenosis in a patient with dwarfism. AB - In this report, we present an adult patient with dwarfism who had severe aortic stenosis with markedly thickened fibrotic valve leaflets without calcification. These findings were well demonstrated by both two- and three-dimensional transesophageal echocardiography and confirmed at surgery and by pathological examination. PMID- 11107212 TI - Dyspnea in the ventilated patient: a call for patient-centered mechanical ventilation. PMID- 11107213 TI - The respiratory therapist and palliative care. PMID- 11107214 TI - Palliative home care for advanced lung disease. PMID- 11107215 TI - Palliation for the dying patient with lung cancer. PMID- 11107216 TI - Organizational change and delivery of multidisciplinary palliative care. PMID- 11107218 TI - The limitations of protocols for end-of-life care. PMID- 11107217 TI - Death and the practitioner. PMID- 11107219 TI - Palliative care in respiratory care: conference summary. PMID- 11107220 TI - Historical and political overview. AB - There are more than 10 million local government workers in the United States. Municipal workers are exposed to a wide variety of serious chemical, biological, physical, ergonomic, and safety hazards. Decisions made by legislative bodies and policy makers historically have had a severe impact on the health and safety of local government workers. In over half of the states, city workers are not covered by safety and health programs approved by the Occupational Safety and Health Administration. They also lack universal coverage under laws providing union recognition and the right to collectively bargain over working conditions. Collective action in the workplace and the political arena remains the most important vehicle for municipal workers to secure safe working conditions. PMID- 11107221 TI - Occupational health services for municipal employees. AB - Most municipalities have departments responsible for fire, policing, public works, parks, sanitation, health, water, administration, and communications. A comprehensive occupational medicine program must address the potential and actual health hazards associated with each of these fields, as well as pre employment/placement medical evaluations, medical surveillance, wellness programs, work-related injury, case management, return-to-work accommodations, and drug and alcohol testing. While no standard for municipal occupational heath services exists, a comprehensive approach to the variety of work settings that are inherent even in small municipalities is important for the service provider. PMID- 11107222 TI - Sewage workers: toxic hazards and health effects. AB - Municipal sewage workers provide an essential service in the protection of public health. The wastewater treatment process brings the worker in contact with multiple pathogens, toxic gases, chemicals, and physical hazards. Issues such as the prevalence of hepatitis A among wastewater treatment workers in the U.S. have not been well studied. There remains a controversy on the need to offer hepatitis A pre-exposure immunization. Health effects to some exposures, such as gram negative bacteria and endotoxins, have been well studied among other workers, and preventive measures, such as permissible endotoxin levels, that have been established for these workers should be adopted for the wastewater treatment environment. Further study into mortality and morbidity rates among sewage workers and the relationship to exposures and the development of preventive measures is needed. PMID- 11107223 TI - Police and corrections. AB - The author summarizes important occupational hazards of police and corrections work. Included in this chapter are morbidity and mortality data of police officers that describes increased risks of suicide, homicide, and ischemic heart disease. Information on tuberculosis exposure in correctional workers and of bloodborne pathogen exposure in police and corrections work is presented. The discussion concludes with strategies for the prevention of illness and injury among police officers and corrections workers based on these data. PMID- 11107224 TI - Occupational health problems of bridge and tunnel officers. AB - Bridge and tunnel officers (BTOs) sustain potential exposure to a number of physical, chemical, and work-organizational factors. They are at risk for both fatal and non-fatal occupational injuries due to moving vehicles, workplace violence, vehicular fires, and physical hazards, such as slippery walking surfaces due to oil or ice on roadways. This chapter describes the spectrum of occupational injuries and illnesses which may be seen in BTOs, focusing on: 1) vehicular exhaust and air pollution, 2) ergonomic hazards, 3) job strain, 4) noise, 5) blood-borne pathogens, 6) chemicals used in road work and maintenance (e.g., lead-based paint), and 7) with the recent advent of electronic traffic sensors, microwave radiation. Special emphasis is given to respiratory disease and cardiovascular disease. Finally, some recommendations for focused health surveillance and preventive efforts in this population are made. PMID- 11107225 TI - School employees: the forgotten municipal workers. AB - The occupational health and safety issues of public school employees in large urban areas are many and complex. Crumbling school infrastructure and crowded classrooms are associated with inadequate indoor air quality, asbestos exposure, noisy environments, and enhanced transmission of communicable and infectious diseases. Poor ventilation in vocational education classrooms, duplicator rooms, kitchens, and science laboratories may also contribute to hazardous exposures. Ergonomic hazards may be responsible for increasing rates of musculoskeletal disorders. Other work-related illnesses and injuries now documented among school employees include asthma, mesothelioma, asbestos-related lung cancer, violent assault, voice disorders, and depression. Such a diverse industry with many potentially hazardous activities and conditions calls for a comprehensive research and intervention agenda. PMID- 11107226 TI - Local government workers' health and safety programs. AB - Public sector workers are denied the broad legal protections of the Occupational Safety and Health (OSH) Act of 1970. The OSH Act provides incentives for states to operate a State OSHA Plan, but they must cover private and non-Federal public sector workers. Public sector workers in states not operating a State OSHA Plan are not protected by the OSH Act. A review of enforcement activities by State OSHA Plans suggests preferential treatment for public employers compared to private. Public sector safety and health programs vary widely in the 27 states without State OSHA Plans. The Department of Labor determined that the majority of these states operate unacceptable programs. Regardless of program type, public sector workersi injuries and illnesses frequently exceed those found in the private sector. PMID- 11107227 TI - Occupational health for firefighters. AB - Occupational health and safety programs for firefighters have received increasing attention over the last several years, due to the growing recognition of potential long-term health risks for firefighters. These workers not only face severe physical and psychological demands, but also risks of chronic or delayed adverse job-related health consequences. Firefighters are routinely exposed to a large number of toxic substances (e.g., carbon monoxide, benzene, particulate, asbestos, polynuclear aromatic compounds, hydrogen chloride, and cyanide) as well as physical hazards such as heat and noise. Their emergency medical response duties also put them at risk of exposure to infectious agents. Firefighters are at increased risk of cardiovascular disease, pulmonary disease, cancer, and noise induced hearing loss. Occupational medical care for firefighters needs to monitor for these long-term health risks. PMID- 11107228 TI - Workplace violence in municipal occupations. AB - Although workplace homicide rates are decreasing, violent events in the workplace remain a high risk in many occupations. The number of violent victimizations in the workplace has been estimated to be as high as 2 million per year among workers. Many municipal workers are at high risk because they provide services and enforce regulations, often in unsecured environments. Compared to other causes of workplace death, homicides and other violent events are relatively new areas of formal study for occupational safety and health specialists. However, there is a growing body of knowledge about the types of interventions that are effective in reducing workplace violence. This chapter examines trends in workplace violence, risk factors for municipal workers, and approaches to prevention. PMID- 11107229 TI - Occupational hazards of municipal solid waste workers. AB - The removal of municipal solid waste is a job associated with a variety of physical, chemical, and biological hazards. Municipal solid waste workers (MSWWs) have a risk of fatal occupational injuries that is much higher than for the general workforce. Among this group of workers, non-fatal injuries are mainly musculoskeletal. Other common injuries are fractures, ocular trauma, and bites, and diseases include skin and gastrointestinal disorders. Workers at municipal solid waste incinerators are exposed to a variety of concerning substances, such as heavy metals, respirable quartz dust, dioxins, furans, and mutagens. Workers can be protected by using safety procedures on and around garbage trucks and with personal protective equipment. The burden of morbidity due to occupational exposure to bioaerosols and carcinogens among MSWWs is unknown. PMID- 11107230 TI - Parks and golf course workers. AB - Most of the work done by parks and golf course workers is performed outside, and they are subject to risks similar to those of other outdoor workers, such as temperature and weather-related problems, infectious diseases, and poisonous plants, snakes, and spiders. They are also exposed to physical hazards related to noise and operation of heavy equipment. Chemical hazards result from use of pesticides and solvents. This chapter reviews pertinent OSHA regulations, pre placement strategies, and medical surveillance exams, and recommends preventive programs. More research is needed to determine specific hazards and work-related injury and illness rates for parks and golf course workers, so that effective preventive programs can be designed. PMID- 11107231 TI - Municipal healthcare workers: work-related health hazards. AB - Municipal healthcare workers are at risk for a variety of occupational health hazards as a result of daily exposure to work-related biological, chemical, enviromechanical, physical, and psychosocial agents. In addition, this population of workers has not been well studied, so the actual risk may be substantially underreported. This chapter describes these risks and hazards as well as prevention and control strategies. A clear and concerted emphasis on specifically targeting this at-risk workforce both in terms of hazard identification and developing and measuring strategies for risk reduction should be a high priority. PMID- 11107232 TI - Health and safety concerns of zoo and aquarium workers and animal control officers. AB - There are 183 zoos and aquariums accredited by the American Zoo and Aquarium Association and scores of smaller, nonaccredited zoos in the U.S. Thousands of individuals work at these zoos in many different roles. Likewise, most counties and many larger cities employ animal control officers and have animal shelters. The authors discuss the numerous hazards encountered by animal control officers as well as employees of zoo and aquarium facilities. Suggested protocols for ensuring a safe workplace are presented. However, a thorough industrial hygiene and safety inspection should be performed on a regular basis, as additional protocols may be needed periodically. PMID- 11107233 TI - A Historical Overview of The Brazilian Society of Infectious Diseases. PMID- 11107234 TI - Emerging Pathogens Associated with Tick-Borne Infections. AB - In recent years, several microbial agents have been identified that result in significant morbidity and mortality. The newly recognized tick borne infections, babesiosis and ehrlichiosis, may be transmitted by the same tick that transmits Borrelia burgdorferi and simultaneous infections may occur. Babesia are intraerythrocytic protozoa that may cause severe hemolytic anemia, whereas Ehrlichia, depending on the species, may infect either monocytes or granulocytes, with associated leukopenia, thrombocytopenia and anemia. Improved laboratory surveillance is urgently needed to assess the prevalence of these worldwide pathogens in order to institute appropriate infection control efforts. PMID- 11107235 TI - Herpes Simplex Virus Shedding in Bone Marrow Transplant Recipients During Low Dose Oral Acyclovir Prohylaxis. AB - A 400mg dose twice-a-day oral acyclovir prophylaxis regimen was evaluated in 50 allogeneic transplant recipients. Twenty (40%) patients experienced 24 episodes of herpes simplex virus (HSV) shedding; l7 (70.8%) occurring during prophylaxis. Thirteen of such episodes were asymptomatic and, in three, it was difficult to differentiate severe mucositis from viral lesions. In the remaining one, HSV pneumonia was suspected after a bronchoalveolar lavage (BAL) procedure performed in an attempt to early detection of cytomegalovirus (CMV). All cases responded to acyclovir therapy or dose adjustment suggesting that acyclovir resistance did not account for the occurrence of infection in our patients. These data demonstrated that oral acyclovir prophylaxis, 400mg dose twice-a-day, was inadequate to suppress viral shedding. The bronchoalveolar lavage procedure in a patient with HSV shedding could precipitate HSV spread to the lungs and the occurrence of pneumonia. PMID- 11107236 TI - Co-Infection of Tuberculosis and HIV/HTLV Retroviruses: Frequency and Prognosis Among Patients Admitted in a Brazilian Hospital. AB - During 2 1/2 year period,378 patients diagnosed with tuberculosis and admitted to a general hospital for care of the poor in Salvador, Bahia, were tested serologically for HIV-1, HTLV-I, and HTLV-II. The patients' mean age was 41.8 (range 14-89); they were hospitalized for a mean of 62 +/- 43 days; 70% were being treated for the first time; most of the remainder were being retreated after non-compliance with previously recommended anti-tuberculosis medication and a few required second-line therapy for relapsed disease. None had had previous serologic testing for retroviruses. Among the study population, 59 (16%) were found to be positive for retroviral infection. The distribution was as follows: 18 (4.8%) had HIV-1, 32 (8.5%) had HTLV-I, 2 of these had both HTLV-l and HTLV II, 9 (2.4%) had both HIV-1 and HTLV-I. The rates of positive serologic tests for retroviral infection in this Salvador is 0.2% for HIV-1 and 1.0% for HTLV-I. Thus, there is a higher than expected frequency of retroviral infections among patients hospitalized for treatment of tuberculosis. The prognosis for treated patients was determined by recording the cause of death and the mortality rate. In the 319 patients with negative serologic testing for retroviruses there were 25 deaths (8%). In 32 patients with HTLV-I infection there were 8 deaths (25%), and in 18 patients with HIV-1 infection there were 6 deaths (33%). In 9 patients with both HIV-1 and HTLV-I there were 5 deaths (56%). The causes of death in each serological group were primarily related to progression of tuberculosis rather than complications of rapid progression of the retroviral infection. We conclude that co-infection and disease due to either HIV-1 or HTLV-I/II infection and tuberculosis is common, that the occurrence of HTLV-I in this population is higher than previously recognized, and that prognosis associated with the management of tuberculosis is adversely affected by the presence of either retroviral infection. In a few patients with both retroviral infections, mortality was very high. All patients with tuberculosis should be tested for retroviral infection because of the prognostic and therapeutic implications. PMID- 11107237 TI - Aedes aegypti Surveillance and Correlation with the Occurrence of Dengue Fever in Bahia, Brazil. AB - Due to the high frequency of dengue fever cases and the presumed association of such an epidemic with an increase in the population of the mosquito vector, Aedes aegypti, we examined the records of the Ministry of Health in the state of Bahia, Brazil, regarding the monitoring of domestic mosquito larvae in municipalities throughout the state. The "House Index" number for larvae in domestic water reservoirs was determined for each municipality based on annual surveys from 1990 to 1994, and in 1996. In 1996, 69% of the municipalites surveyed in Bahia were positive, and 30% had indices above 5%. During 1990 and 1991, the level of larvae identified was low and stable; however, during November and December, 1992, a dramatic increase was recorded. The increase continued until 1996, when over 100 fold increases in house indices were recorded in Feira de Santana and Ilheus, and a 60-fold increase in Salvador. The dengue fever epidemic was documented in the region beginning in 1994. A strong correlation has been demonstrated between an increase in the mosquito larvae population and the emergence of dengue fever. PMID- 11107238 TI - Epidemiological Characteristics of HTLV-I and II Co-Infection in Brazilian Subjects Infected by HIV-1. AB - Serum samples from 895 HIV-1 infected individuals were tested for antibodies against HTLV-I and HTLV-II. The overall prevalence of the co-infection was high (16.3%). Epidemiological information was obtained from each subject including gender, age, intravenous drug use (IVDU), blood transfusion, previous diagnosis of sexually transmitted diseases (STD) and sexual behavior. The risks for acquiring retroviral infections other than HIV-1 were evaluated and the prevalence of co-infection was compared according to the AIDS clinical status. We detected seven cases (0.9%) of triple infection. HTLV-I co-infection was associated with blood transfusion (p=0.02), IVDU (p=0.0001) and female gender (p=0.0001). Co-infection by HTLV-II was primarily associated with a history of past blood transfusion (p=0.009). Women co-infected by HTLV-I or HTLV-II had a higher risk of AIDS than those infected only by HIV-1 (RR=2.04; 95% CI: 127-327, p=0.007 and RR=3.09; 95% CI: 1.07-8.91, p=0.04, respectively). These findings suggest that co-infection by HTLVI or II in Bahia, Brazil, may modify the clinical course of HIV-1 infection. PMID- 11107239 TI - Disseminated Strongyloides stercoralis Infection in an AIDS Patient: The Role of Suppressive Therapy. AB - Patients with AIDS are prone to develop infections caused by opportunistic pathogens. Unusual agents, such as Strongyloides stercoralis, are being described in this syndrome, resulting in disseminated disease which is always severe and, in some cases, fatal. We describe a case of a patient with AIDS and Strongyloides stercoralis infection involving the gastrointestinal tract and the lungs. Therapy with thiabendazole for ten days led to resolution of the acute episode. Preventive therapy with 3g of thiabendazole once a week was then prescribed, and repeated fecal examinations were negative for larvae. Following discontinuation of treatment, however, the patient again had a positive fecal examination for Strongyloides stercoralis larvae, even though reinfection was considered to be very unlikely. The patient was retreated with a shorter course of therapy and once per week preventive therapy was reintroduced. After four months of follow up, repeated fecal examinations were negative. When the treatment was changed to thiabendazole given once every two weeks, however, pulmonary Strongyloides stercoralis recurred. Subsequently, because of intolerance to thiabendazole, the patient was treated with cambendazole. The patient died three months later due to Pseudomonas aeruginosa pneumonia. Prolonged therapy for Strongyloides stercoralis infection may be necessary. Although further evaluation is needed, 3g of thiabendazole once a week may be adequate for this purpose. Cambendazole may be a useful alternative for disseminated Strongyloides stercoralis. PMID- 11107240 TI - Introduction to The New Journal, "The Brazilian Journal of Infectious Diseases" An International Perspective. PMID- 11107241 TI - Cryptococcal Meningitis. AB - Meningitis is the most common manifestation of disseminated cryptococcosis. Cryptococcal meningitis is the most common systemic mycotic infection in AIDS, which is the most frequent predisposing condition. The most severe complication is acute cerebral hypertension. Medical options for therapy have broadened considerably, but generally include initial intensive induction treatment followed, in patients with AIDS, by chronic suppression. With aggressive management of cryptococcal meningitis, mortality may be reduced to 10% or less. PMID- 11107242 TI - A Role for Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) in the Treatment of Neutropenic Patients with Pneumonia. AB - Pneumonia is a serious, difficult to manage, and often fatal infection in neutropenic patients. The availability of hematopoietic growth factors has made it possible to evaluate the role of reversing the neutropenic state. Granulocyte macrophage colony-stimulating factor (GM-CSF) has been used in an investigator initiated, open-label clinical trial in approximately 1200 patients. Data collected on each patient was reviewed to identify all patients who had the combination of neutropenia and pneumonia. Sixty-eight patients (5% of the patients for whom GM-CSF was requested) met the criteria for having neutropenic pneumonia. In this patient population there were 45 males and 20 females (gender was not indicated in 3). Ages ranged from 3 to 83 years (mean 39 +/- 18 years). The underlying diseases included: 7 patients who were receiving chemotherapy for solid malignant tumors, 14 for lymphoma, and 22 for leukemia; 15 post bone marrow transplantation primarily for hematologic malignancy; 3 idiosyncratic drug induced neutropenia; and 7 with other causes of neutropenia. The type of pneumonia was predominantly fungal in 21 patients, bacterial in 23, viral in 2, protozoal in 1, and uncertain in 21 (presumed to be bacterial in 19 and viral in 2). Patients received a mean of 5u/kg GM-CSF (range 1.3-12.5ug/kg) daily for a mean of 13 +/- 10 (range 2-57) days. The mean leukocyte count at start of treatment was 600 +/- 500 cells/mm(3), and at the end of treatment was 5600+/ 9200 cells/mm(3) (P=0.001). The time between start of GM-CSF and a leukocyte level in excess of 1500/mm(3) was a median of 13 days. Hematopoietic recovery was shown in 46/62 (74%), 40/64 (63%) showed good clinical and/ or radiologic improvement, and 41/68 (60%) survived. Four illustrative case reports are provided. By comparing the hematologic responders to non-responders, it is clear that persistent neutropenia contributed significantly to poor clinical outcome and mortality. Only 3/22 (13%) of non-responders survived, whereas 38/46 (83%) of responders survived. There were 7 adverse events (rash 1, fever/chills 2, malaise 1, myalgia/bone pain 2, increased myeloblasts 1) which were considered to be related to use of the cytokine. Aggravation of the pulmonary inflammation or sepsis syndrome was not observed. Tolerability was good or very good in 89% of patients. Based on this open-label study, the use of GM-CSF in combination with appropriate antibiotics is effective and safe for the management of patients with pneumonia and severe hematopoietic dysfunction. PMID- 11107243 TI - Once Daily Fleroxacin and Twice Daily Ciprofloxacin are Both Effective in the Treatment of Complicated Urinary Tract Infections. AB - In a multicenter randomized double-blind, trial, 90 patients received either fleroxacin 400mg pa once/day or ciprofloxacin 500mg po twice/day for treatment of complicated urinary tract infections (UTI). Treatment was administered orally and presumptively. Bacteriological efficacy was assessed 7 days post-treatment. In total, 78 patients were available for efficacy testing: 40 in the fleroxacin group and 38 in the ciprofloxacin group. The bacteriological cure rate was 92.5% and 94.7% in the fleroxacin and ciprofloxacin groups, respectively. The most commonly isolated pathogen (E.coli) was erradicated in 94.1% and 95.8% of the cases in fleroxacin and ciprofloxacin groups, respectively. Eight patients in the fleroxacin group had some adverse events, two of them severe (insomnia and photodermatitis). In the ciprofloxacin group, 11 patients had adverse events of mild to moderate intensity, mainly affecting the digestive system. ln conclusion, fleroxacin 400mg po once/day and ciprofloxacin 500mg po twice/day were both effective in the treatment of complicated urinary tract infections. PMID- 11107244 TI - In Vitro Antimicrobial Activity Against Enterococci Isolated in a University Hospital in Sao Paulo, Brazil. AB - The prevalence of multiresistant enterococci (MRE) is rapidly increasing and becoming an important problem in several countries. Sao Paulo Hospital is a 600 bed tertiary hospital located in Sao Paulo, Brazil. The use of vancomycin is very high in the hospital due to the high prevalence of multi-resistant S. aureus (around 70%). We susceptibility tested 250 isolates of Enterococci consecutively collected between March, 1994 and June, 1995. Isolates were susceptibility tested using agar dilution disc diffusion BHI screen plating, and E test. In addition to vancomycin and teicoplanin, the isolates were tested against ampicillin, gentamicin, streptomycin and RP 59-500. Methods used for susceptibility testing were compared. None of the isolates showed high-level resistance to vancomycin or teicoplanin. The MIC90s for teicoplanin were <e;1ug/mL for E. faecalis (EF, n=216), E. faecium (EFM, n=23) and for the non-faecalis-non-faecium (NFNF, n=11) species. The MlC90s for vancomycin were 2ug/mL, 4ug/mL and 4ug/mL for EF, EFM and NFNF respectively. Eight isolates (3.2%), 5 E. faecalis, 2 E. casseliflavus and 1 E. gallinarum presented intermediate MICs for vancomycin (6 - 12ug/mL), but they were highly susceptible to teicoplanin (MIC 0.19 - 1ug/mL). The percentage of resistance to ampicillin and high-level resistance to gentamicin and streptomycin were, respectively: 4.8%, 26.4%, and 24.8%. In spite of the high usage of vancomycin in our hospital, the prevalence of glycopeptide resistance among enterococci seems to be low. Teicoplanin appears to be more potent than vancomycin, RP 59-500, gentamicin, streptomycin and ampicilin against this genus, especially EFM and NFNF species. PMID- 11107245 TI - Blastocystis hominis as a Potential Cause of Diarrhea in AIDS Patients: a Report of Six Cases in Bahia, Brazil. AB - The role of Blastocystis hominis as a human pathogen is controversial, although there is some evidence suggesting that it is the agent implicated in causing diarrhea in immunocompromised patients. We report 6 cases of AIDS patients presenting with diarrhea with no agents identified in their stools exceptB. hominis. In all cases, treatment was followed by complete recovery from symptoms and clearance of B. hominis from the patients' stools. In 2 cases, relapse of diarrhea was followed by a positive stool examination for B. hominis oocysts, which again disappeared after treatment. These findings provide additional evidence for considering B. hominis a potential intestinal pathogen in AIDS patients. PMID- 11107246 TI - Leprosy and AIDS: Report of a Fatal Case and Literature Review. AB - We report a severe fatal lepromatous leprosy case in an adult homosexual male 2 years after AIDS class IV was diagnosed. Multidrug therapy including thalidomide for erythema nodosum leprosum (ENL) was ineffective. The patient died with multiple large skin ulcerations due to multibacillary leprosy. Leprosy in AIDS patients may present as a severe uncontrolled disease, with extensive ENL unresponsive to therapy. PMID- 11107247 TI - Blastocytis hominis - More than 70 Years of Debate Regarding Pathogenicity! PMID- 11107248 TI - A review of the biology and control of the coffee berry borer, Hypothenemus hampei (Coleoptera: Scolytidae). AB - The coffee berry borer, Hypothenemus hampei Ferrari, is a serious problem for the majority of the world's coffee growers and has proved to be one of the most intractable of present day pests. Despite a great deal of research, control still depends largely on the application of the organochlorine insecticide endosulfan, which is damaging to the environment, or a series of cultural and biological control methods which give variable and unpredictable results. This review summarizes the most important aspects of the biology and ecology of H. hampei and its control and identifies weak points in the knowledge about this pest. Emphasis is placed upon an analysis of the non-chemical control methods available and suggestions are offered for novel ecological and environmental factors worthy of further research, in the search for viable and sustainable control methods. PMID- 11107249 TI - Evaluation and host specificity of two seed flies Mesoclanis polana and M. magnipalpis (Diptera: Tephritidae): biological control agents for Chrysanthemoides monilifera (Asteraceae) in Australia. AB - Larvae of the South African tephritid flies Mesoclanis polana Munro and M. magnipalpis Bezzi feed in the developing seeds of Chrysanthemoides monilifera. Host specificity evaluation using 109 plant species from 25 families indicated that complete development was restricted to their natural host C. monilifera. Minor feeding and limited development was detected on 18 species, but was of no ecological or economic significance. Mesoclanis polana and M. magnipalpis have been released in Australia and M. polana has established and dispersed widely. Mesoclanis magnipalpis has not yet become naturalized. Parasitism of M. polana in Australia by several species of Hymenoptera has been detected, but is not expected to limit the establishment and impact of these flies. PMID- 11107250 TI - The diversity of beetle assemblages in different habitat types in Sabah, Malaysia. AB - The diversity of beetle assemblages in different habitat types (primary forest, logged forest, acacia plantation and oil palm plantation) in Sabah, Malaysia was investigated using three different methods based on habitat levels (Winkler sampling, flight-interception-trapping and mist-blowing). The overall diversity was extremely high, with 1711 species recorded from only 8028 individuals and 81 families (115 family and subfamily groups). Different degrees of environmental changes had varying effects on the beetle species richness and abundance, with oil palm plantation assemblage being most severely affected, followed by acacia plantation and then logged forest. A few species became numerically dominant in the oil palm plantation. In terms of beetle species composition, the acacia fauna showed much similarity with the logged forest fauna, and the oil palm fauna was very different from the rest. The effects of environmental variables (number of plant species, sapling and tree densities, amount of leaf litter, ground cover, canopy cover, soil pH and compaction) on the beetle assemblage were also investigated. Leaf litter correlated with species richness, abundance and composition of subterranean beetles. Plant species richness, tree and sapling densities correlated with species richness, abundance and composition of understorey beetles while ground cover correlated only with the species richness and abundance of these beetles. Canopy cover correlated only with arboreal beetles. In trophic structure, predators represented more than 40% of the species and individuals. Environmental changes affected the trophic structure with proportionally more herbivores (abundance) but fewer predators (species richness and abundance) in the oil palm plantation. Biodiversity, conservation and practical aspects of pest management were also highlighted in this study. PMID- 11107251 TI - Suitability of two root-mining weevils for the biological control of scentless chamomile, Tripleurospermum perforatum, with special regard to potential non target effects. AB - The biology and host range of the two root-mining weevils Diplapion confluens Kirby and Coryssomerus capucinus (Beck), two potential agents for the biological control of scentless chamomile Tripleurospermum perforatum (Merat) Lainz, were studied in the field in southern Germany and eastern Austria, and in a common garden and under laboratory conditions in Delemont, Switzerland from 1993 to 1999. Both weevils were univoltine, and females started to lay eggs in early spring. Diplapion confluens had three and C. capucinus five instars. Larvae of both species were found in the field from mid-April until the end of July; later instars preferentially fed in the vascular cylinder of the shoot base, root crown or root. Although larvae of both species occupy the same temporal and spatial niche within their host plants, they occurred at all investigated field sites together, and showed a similar distribution within sites. No negative or positive interspecific association was detected. Host-specificity tests including no choice, single-choice, and multiple-choice tests under confined conditions, as well as tests under field conditions with natural and augmented insect densities revealed that both herbivores were specific to plant species in the tribe Anthemideae. However, their development to mature larva or adult on several cultivated plants, as well as on one plant species native to North America, rendered them unsuitable for field release in North America. It was concluded that to investigate non-target effects reliably, host-specificity tests with biological control agents should be carried out under a variety of conditions, particularly with augmented insect densities, as are expected to occur naturally after release. PMID- 11107252 TI - Stabling and the protection of horses from Culicoides bolitinos (Diptera: Ceratopogonidae), a recently identified vector of African horse sickness. AB - The stabling of horses at night reportedly offers protection from African horse sickness and the most significant vector of the disease, Culicoides imicola Kieffer, has been shown to be exophilic. In certain high-lying regions of South Africa, however, C. bolitinos Meiswinkel, may be the major vector of the disease but its entry behaviour into stables is unknown. Accordingly, in the eastern Free State province of South Africa, light trap catches of C. bolitinos inside stables and outside, were compared. Two horse-baited stables, one traditional, and one modern, were used and combinations of stable (old/new), ceiling fans (on/off) and accessibility to Culicoides (stable doors open/closed or windows gauzed/ungauzed) were investigated as treatments. A total of 111,452 Culicoides of 26 species was collected on 60 trap nights; C. bolitinos was dominant (89.1% overall) with C. imicola second in abundance (2.9%). Outside catches were greater on warmer, drier, evenings but were suppressed by high wind speeds. Catches of C. imicola inside stables with doors open, or with windows ungauzed, were less than the numbers captured outside. In contrast, more C. bolitinos were caught in open stables than outside, i.e. open structures may protect horses from the exophilic C. imicola, but may increase attack rates from the endophilic C. bolitinos. The closing of doors and the gauzing of windows, however, led to a 14-fold reduction in numbers of C. bolitinos and C. imicola entering stables. A well-gauzed 'traditional' stable was as effective as a closed 'modern' stable. Ceiling fans had no suppressant effect. PMID- 11107253 TI - Parasitoids of medfly, Ceratitis capitata, and related tephritids in Kenyan coffee: a predominantly koinobiont assemblage. AB - Arabica coffee was sampled from two sites in the central highlands of Kenya (Rurima, Ruiru) and one site on the western side of the Rift Valley (Koru). Three species of ceratitidine Tephritidae, Ceratitis capitata (Wiedemann), C. rosa Karsch and Trirhithrum coffeae Bezzi, were reared from sites in the central highlands, and an additional species, C. anonae Graham, was recovered from the western-most site. Ten species of parasitic Hymenoptera were reared from these tephritids. The parasitoid assemblage was dominated by koinobionts. Eight of the species are koinobiont endoparasitoids, but only one idiobiont larval ectoparasitoid was reared, and only one idiobiont pupal endoparasitoid. The effects of sampling bias on determination of parasitoid assemblage size associated with concealed hosts are discussed. The potential for use of these parasitoids in biological control is also discussed. Most of the parasitoid species recovered during this study are capable of developing on C. capitata, while several also attack C. rosa. Both flies are notorious pests of tropical and subtropical fruits. PMID- 11107254 TI - The occurrence of Dialeurodes citrifolii and Aleurochiton pseudoplatani (Hemiptera: Aleyrodidae) in Lebanon--important new records in the Europe Mediterranean region. PMID- 11107309 TI - [Tridimensional skin models recording percutaneous absorption] AB - Today there is a lack of generally accepted alternatives to animal experiments for the evaluation of percutaneous absorption or skin penetration. To evaluate the potential of commercially available reconstructed epidermis we compared the barrier function of the three dimensional human skin equivalents Episkintrade mark, EpiDermtrade mark, and Skinethictrade mark to those of excised human skin. Moreover the batch-to-batch variation of skin equivalents was determined. Methods for evaluation included the determination of transepidermal water loss (TEWL) and of the prednisolone (PD) penetration. Whereas TEWL values of Skinethictrade mark and EpiDermtrade mark were very close (15 g.m-2.h-1), PD-penetration showed a clearly superior barrier function with the Skinethictrade mark model. Drug penetration was only 50% of the EpiDermtrade mark model and PD concentrations in the acceptor medium amounted to 150 ng/ml as compared to 300 ng/ml. The highest values were obtained with the Episkintrade mark model (20 g.m-2.h-1, 500 ng/ml). Therefore this model has the poorest barrier function. The TEWL value for human skin was 9 g.m-2.h-1 and PD concentrations in the acceptor medium were below the limit of detection (10.1 ng/ml). Because of the superior barrier function of the Skinethictrade mark the following evaluation was only performed using this model. Batch-to-batch variation was less than with human skin (interindividual variation of 2 h TEWL 12% and > 19%). Despite of the slightly less well-developed barrier, this skin model clearly appears superior with respect to the reproducibility of data. Therefore future investigations on the potential of human skin equivalents for the determination of percutaneous absorption appear very promising. PMID- 11107310 TI - [Development of prediction models for three in vitro embryotoxicity tests which are evaluated in an ECVAM validation study] AB - In 1997 ZEBET started the co-ordination of a study funded by the European Centre for the Validation of Alternative Methods (ECVAM) with the aim of pre-validation and validation of three in vitro embryotoxicity tests. These tests employ the cultivation of post-implantation whole rat embryos (WEC test), cultures of primary limb bud cells of rat embryos (micromass test, MM-Test), and cultures of a pluripotent mouse embryonic stem cell line (embryonic stem cell test, EST). In the current Validation Study each of the tests is evaluated in four laboratories under blind conditions. In an initial phase of the validation study six out of 20 test chemicals comprising different embryotoxic potential (non, weak and strong embryotoxic) were tested. The results were used to improve the prediction models (PM) for the WEC test and the MM-Test in order to identify the embryotoxic potential of test chemicals. In addition, the existing PM for the EST was evaluated using the results from testing of the initial six chemicals. The PM for the EST was developed using the results of a previous pre-validation study (Scholz et al.,1999), in which 94% of the learning sample were classified correctly. Applying this PM to the results of the initial phase of the current validation study, 80% of the experiments were classified correctly according to the embryotoxic potential of the tested chemicals in vivo. Applying the PM for the MM-Test and the WEC test that were developed during the current validation study in both tests, 79% correct classifications were achieved. Since the PM of the WEC-Test took into account only parameters of growth and development, but not cytotoxicity data, a second PM (PM2) for the WEC was developed that was improved by incorporating cytotoxicity data of the differentiated mouse fibroblast cell line 3T3 derived from the EST. This approach, which has previously never been used as an adjunct to the WEC test, resulted in an increase of correct classifications to 96%. PMID- 11107312 TI - [Cocultures between primary parenchymal and nonparenchymal liver cells improve the reliability of results from in vitro toxicity testing] AB - Conventional homotypic hepatocyte cultures do not include the possible contribution of nonparenchymal liver cells, particularly Kupffer cells, to the pharmacological and toxicological consequences after exposure to xenobiotics. Therefore the exchange of soluble factors between liver cells was investigated in cocultures between primary, freshly isolated cultured rat hepatocytes and Kupffer cells. Cocultures were exposed to endotoxins in combination with phenobarbital, 3 methylcholanthrene or lead. A strong but selective down-regulation of xenobiotic induced cytochrome P450 isoforms was detectable, mediated exclusively by TNFalphareleased from the Kupffer cells. Pb synergistically increased this endotoxin induced TNFalpha-release. The results indicate that cocultures improve the reliability of data obtained from organ specific cell cultures and that they simulate much closer the situation in the intact liver. PMID- 11107311 TI - [An in vitro test system for evaluation of human-immunotoxicological actions of polychlorinated biphenyls] AB - An in vitro test system using isolated human leukocytes for evaluation of immunotoxicological actions of polychlorinated biphenyls (PCB) is described. Immunotoxic effects of PCB 77, PCB 126 could be demonstrated by evaluation of the production of interleukin IL-1alpha, IL-2 and the tumor necrosis factor (TNF alpha) of stimulated human leukocytes. The release of IL-1alpha, IL-2 and TNF alpha was suppressed up to 60-80% by 0,05 ng of the PCB. PMID- 11107313 TI - [The isolated normothermic hemoperfused porcine leg as model for pharmacological and toxicological investigations] AB - In this short communication we present the isolated normothermic hemoperfused porcine leg as an alternative model for pharmacological and toxicological investigations. Both legs and blood were obtained at a slaughter house. Legs were connected to the perfusion apparatus Mediport Biotechnik and provided with room air and nutrients via a dialysis module over a seven hour period. The perfusion pressure, the perfusion flow, the response to noradrenergic stimuli, the glucose consumption influenced by insulin dosage and the absorption of estradiol were investigated. Once noradrenalin was given perfusion pressure increased as expected and dropped back to normal only a few minutes after administration. Moreover, in the legs treated with insulin, a glucose consumption was detected. Only the legs treated with estradiol showed an increasing concentration of estradiol in plasma throughout the experiment. From the results obtained, it can be concluded that the isolated normothermic hemoperfused porcine leg seems to be a suitable alternative model for the testing of transdermal absorption as well as for the investigation of acute vasoactive substances. Further studies will be performed in our laboratory in order to determine the metabolic condition of this system as well as to ascertain other possibilities of this model in pharmacological and toxicological investigations. PMID- 11107314 TI - [Das Wissenschaftsjournal Nature falscht Aussagen des Deutschen Tierschutzbundes] PMID- 11107315 TI - [Kommentar zum Entwurf der Verordnung uber die Meldung zu Versuchszwecken und zu bestimmten anderen Zwecken verwendeter Wirbeltiere in Deutschland] PMID- 11107316 TI - [Tierverbrauch im Studium] PMID- 11107317 TI - Progress in the Research of Atherosclerosis and Restenosis Using an in vitro Method: Transfilter Cocultures with Human Vascular Cells. AB - Excessive proliferation and migration of human arterial smooth muscle cells (haSMC) are prominent features of both primary atherogenesis, as well as restenosis following interventions, such as balloon angioplasty (PTCA) or stent implantation. Thus, in the last two decades many efforts are made to establish a therapeutic strategy with antiproliferative and antimigratory compounds. A great variety of substances belonging to different drug classes were already tested both in vitro and in vivo but there is still no breakthrough in the prevention or treatment of human arterial vessel diseases. Among the choice of more potent compounds, the main reasons for this failure are the use of unappropriate animal models or animal cell cultures allowing no direct transfer of the results to the human situation. For that reason, the more complex transfilter coculture model was developed and established for the cocultivation of human vascular cells and blood cells to imitate the morphology of the arterial vessel wall in vitro. The present paper describes the morphology of fibromuscular-like and atheromatous like proliferates induced in this model in comparison to human plaques, as well as its practicability for the pre-screening of antiarteriosclerotic compounds. As examples, two chemically different compounds are shown. We found that the described transfilter coculture system is a suitable and well-established model allowing fast and reproducible studies with antiproliferative and antimigratory drugs. The transferral of the results to the human situation on the basis of these complex in vitro-studies seems to be improved when compared to most animal models. PMID- 11107318 TI - Potency Testing of Swine Erysipelas Vaccines by Serology Results of a Pre validation Study. AB - The European Pharmacopoeia (Ph. Eur.) monograph on Swine Erysipelas Vaccine (inactivated) (Ph. Eur., 1997) requires the potency of each batch to be demonstrated in a mouse protection test. In this parallel-line bioassay, mice are challenged with a virulent strain of Erysipelothrix (E.) rhusiopathiae after immunisation with different doses of either the standard preparation or of the test vaccine. More than one hundred animals are necessary for the routine testing of a single batch. In previous studies (Beckmann und Cubetaler, 1994; Rosskopf Streicher et al., 1998), we have shown that an indirect enzyme-linked immunosorbent assay (ELISA) may be used to quantify the humoral response of mice. This method can replace the challenge model for the purposes of potency testing in the following manner: Ten mice are immunised subcutaneously with 1/10 of the vaccine dose required for pig vaccination. After three weeks, the mice are bled under anaesthesia. Serum samples are pooled and the antibody content is compared to that of a reference serum. In view of animal welfare the advantages of the alternative model are obvious: A highly reduced number of animals and the replacement of challenge exposure. This includes the discontinuation of the control group with a mortality rate of 100%. A pre-validation study was initiated to evaluate the performance of the serological method. Eight laboratories used the test kits to evaluate pooled serum samples from vaccinated mice. Both the reagents and the test protocol were shown to be satisfactory. The intra- and inter-laboratory reproducibility that was achieved indicates that the method is a strong candidate for validation. PMID- 11107319 TI - Acute Toxic Class Methods: Alternatives to LD/LC50 Tests. AB - The Acute Toxic Class (ATC) Methods for the oral, dermal and inhalation routes were developed as alternatives to the respective LD/LC50 Tests. The ATC Methods use at least 40% (and up to 90%) fewer animals and their reliability with respect to classification probabilities into one of several toxicity classes is in the same order of magnitude or even better in comparison to the LD/LC50 Tests. The ATC Methods are stepwise procedures with the use of three fixed starting doses/concentrations, three animals per step and a maximum of 6 animals per dose/concentration. The outcome (numbers of dead or moribund animals) will determine if further testing is necessary or if the test is terminated. Like the previous tests the biometric basis of the ATC Methods is the Probit model. The oral ATC Method is already an official Test Guideline of the OECD and the EU and it is widely used for the notification of new chemicals in the EU. This oral ATC Method was validated by two animal ring studies together with biometric evaluations. With respect to classification probabilities and expected animal numbers the agreement between animal data and biometric evaluations was very good. Therefore for the dermal and inhalation routes of administration the ATC Methods were entirely developed on a biometric basis. These two methods have still to be adopted by OECD. However, in view of the recently agreed Global Harmonisation System (GHS) of classification for acute toxicity all three ATC Methods have to be revised in order to include these new classification systems. PMID- 11107320 TI - Embryonic Stem Cells in vitro - Prospects for Cell and Developmental Biology, Embryotoxicology and Cell Therapy. AB - Embryonic stem (ES) cells are able to differentiate in vitro via embryo-like aggregates, so-called "embryoid bodies", into derivatives of the endodermal, ectodermal and mesodermal lineage. We established standardised protocols for cardiogenesis, myogenesis, neurogenesis and vascular smooth muscle cell differentiation in vitro. The developmentally controlled expression of tissue specific genes, proteins, ion channels and receptors during ES cell differentiation is the basis of several in vitro approaches: (1) "Loss of function" assays with ES cells containing homozygous mutations of specific genes, (2) "Gain of function" assays with ES cells overexpressing exogenous genes, (3) Developmental analysis of teratogenic/embryotoxic compounds in vitro, (4) Pharmacological assays and the establishment of model systems for pathological cell functions, and (5) The application of differentiation and growth factors for induction of selectively differentiated cells which, in the future, may be used as a source for tissue grafts. We propose the ES cell technology as valuable in vitro system to substitute and reduce the use of animals in basic and applied research. PMID- 11107321 TI - [Genetic algorithms in 3D-QSAR: The use of multiple ligand orientations for improved predictions of toxicity] AB - In a recent article in this journal, we discussed the application of a 3D-QSAR technique to the prediction of the toxicity of dibenzodioxins, dibenzofurans, and biphenyls (Vedani et al., 1999a). The use of such methods is legitimate because there is strong evidence that the toxicity is mediated by the Aryl hydrocarbon (Ah) receptor, a regulatory element involved in the mammalian metabolism of xenobiotics. In an extention to a concept developed at our laboratory (Vedani et al., 1998), we now show that safer predictions are possible if instead of a single - and, therefore, necessarily biased - assumption about the mutual orientation of the toxins, an ensemble of possible orientations is used for model construction. The contribution of a single entity within this ensemble to the toxin-receptor interaction energy is determined by a Boltzmann criterion. While in the single-orientation model two of the 26 toxins of an external test set were predicted false positive or false negative, all test substances are correctly predicted in the multiple-orientation model - including up to four different orientations per molecule - within a factor 10 of the experimental binding affinity. These results demonstrate that 3D-QSAR techniques based on a genetic algorithm can be used to predict the toxicity of chemical and pharmacological substances "in computo" if a receptor-mediated mechanism can be assumed. Consequently, the method can be used in toxicological screening assays, thereby replacing stressful tests on animals. PMID- 11107322 TI - New Applications of the Human Whole Blood Pyrogen Assay (PyroCheck). AB - The absence of pyrogens in injectable drugs is an indispensable safety control because contaminants causing fever pose a life-threatening risk to the patient resulting in the worst case in death by shock. When fever- inducing agents, i.e.pyrogens, come into contact with the immunocompetent cells in blood, these cells release mediators which transmit the fever signal to the thermoregulatory centre of the brain. The Phamocopoeia lists currently two test systems for pyrogenicity: 1. The in vivo rabbit pyrogen test which measures the fever reaction following injection of the sample to the animals. 2. The in vitro Limulus Amebocyte Lysate assay (LAL) which measures the coagulation in a lysate prepared from the blood of the horseshoe crab specifically initiated by endotoxins, i.e. cell wall components from Gram-negative bacteria. The new test presented here (PyroCheck) exploits the reaction of monocytes/macrophages for the detection of pyrogens: human whole blood taken from healthy volunteers is incubated in the presence of the test sample in any form, be it a solution, a powder or even solid material. Pyrogenic contaminations initiate the release of the "endogenous pyrogen" Interleukin-1beta determined by ELISA after a fixed incubation time. The technology presently listed in the Pharmacopoeia is limited to parenteralia (rabbit test: biologicals and pharmaceuticals, LAL: predominantly pharmaceuticals). In the EU Medical Devices Directive from 1995 the rabbit pyrogen test for medical products is in some cases requested. (in some cases LAL of an eluate from the device). However, pyrogen-testing needs to cover also innovative high-tech products such as medical devices (implants, medical plastic materials, dialysis machines), cellular therapies and species-specific agents (e.g. recombinant proteins). Here we report that the human blood test PyroCheck is suitable for testing filters in air quality control as well as for assessing medical devices and biocompatibility (dialysate fluid), i.e. that it can be extended to a wide spectrum of applications. PMID- 11107323 TI - Validation of in vitro Tests for Skin Corrosivity. AB - There have been several highly significant achievements in the area of alternatives for skin corrosivity testing in the last three years, most notably: (a) the validation, and subsequent endorsement, of two replacement alternative tests for skin corrosivity (the rat skin transcutaneous electrical resistance [TER] and EPISKINtrade mark human skin model assays) by the European Centre for the Validation of Alternative Methods (ECVAM); (b) an evaluation of the proposed OECD testing strategy for skin corrosion/irritation as it relates to the classification of corrosives; (c) completion of a successful prevalidation study on the use of the EpiDermtrade mark human skin model for corrosivity testing; (d) a review of CORROSITEXtrade mark by the Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAM) in the US; and (e) the submission of draft new test guidelines on skin corrosion to the OECD Secretariat and EU National Coordinators (Annex V test methods). It is now hoped that regulatory acceptance of the validated in vitro tests for skin corrosivity, at both EU and OECD levels, will be secured as quickly as possible. PMID- 11107324 TI - [Man and his fellow-creatures under ethical aspects] AB - Preliminary remarks Preceding the detailed literary review, here a few events, topics and publications for the busy reader including * The decision of the Bundesverfassungsgericht (Federal Constitutional Court) to declare unconstitutional and invalid chicken owner ordinances permitting caging * The decision of the German parliament to declare animal protection a national goal * Publication of a number of books which are likely to influence discussions for years to come since they bring to a close developments having emerged over an extended period of time. Without claiming to be exhaustive the following should be mentioned: Marc Bekoff and Carron A. Meany, ed.: "Encyclopedia of animal rights and animal welfare", an extensive work being reviewed by Peter Thornton in chapter 3.6 of this report. For the first time, animal protection of the Anglo American tradition is being summarised, in theory as well as in practice, and, on high standards: "The list of contributors reads like a Who"s Who of experts in their chosen fields and includes philosophers such as Peter Singer, Tom Regan, Tom Beauchamp and Bernard Rollin and welfare scientists such as Don Broom, David Fraser, Temple Grandin. Others involved in examining the role of animals in society and our relationship with them, such as Andrew Linzey, Richard Ryder, James Serpell and David Morton (to name just a few), have also provided entries" (Peter Thornton). The comprehensive monograph by Johannes Caspar: "Tierschutz im Recht der modernen Industriegesellschaft. Eine rechtliche Neukonstruktion auf philosophischer und historischer Grundlage" (Animal Protection in the Law of modern industrial Society. A new legal Construct on a philosophical and historical Base) represents an extensive critical incorporation of animal protection in Germany under legal and ethical aspects including a detailed rendering of the actual treatment of animals. The term "critical incorporation" was used on purpose since never before was cruelty to animals presented in such completeness and thoroughness, together with the distance and objectivity originating in the legal stance. The book is of invaluable significance in particular for the commentators of the German animal protection law as amended in 1998: For the first time, they are now in a position to draw from an abundance of facts and evaluations relevant to animal protection. It appears as a fortunate circumstance in this context, that not only the traditional Lorz-commentary is being continued (Albert Lorz and Ernst Metzger: "Tierschutzgesetz" [Animal Protection Law], commentary, 5th edition) but also a completely new commentary is being developed by the team of Hans Georg Kluge, ed., Antoine F. Goetschel, Jorg Hartung, Eisenhart von Loeper, Jost Dietrich Ort and Kerstin Reckwell: "Kommentar zum Tierschutzgesetz" (Commentary to the Animal Protection Law). "Moral Status" is not only the title of an important investigation by Mary Anne Warren (in the introductory chapter she describes and evaluates the basics of the Status discussion to date) but also a topic singularly dominating the scientific discourse. Out of rejection or reservation regarding the biocentrical position of Albert Schweitzer or Paul W. Taylor, time after time new restricting concepts have been introduced. In the same way, the criterion of the ability to suffer as developed by Bentham was considered by some as too far reaching. In considering animals ethically the line can be drawn so tightly that, aside from apes and maybe some large marine mammals, the whole of the animal kingdom can be degraded to at best, the ecological worthy resources of man; and all this without being suspected of an adversary position to animal protection. At times, the impression arises that science is seeking and finding ever new hurdles in order to keep the number of morally considered fellow-creatures as small as possible. Following common opinion, even animals with moral status are only entitled to protection from pain or suffering but not from being killed, as long as it is performed without causing fear or pain. In the prevailing opinion, only animals demonstrating a conscious will to live have the right to be protected and not the host of "lower" animals who are granted only a kind of life-impulse. But why is a life-impulse so much less powerful, so much less worthy? Does not even an oyster summon up all of it"s force to defend itself, instinctively? However, it is true that we cannot protect all of life or all animals to the same degree. But there is no sufficient reason to exclude this multitude of fellow-creatures from any consideration at all just because they are only demonstrating a life-impulse. Maybe, it would be meaningful to adopt a gradually increasing moral status, granted any living creature by virtue of it"s being alive. This would mean a protection from thoughtless and random harm and annihilation. This "biophilia", regarding all life as valuable (even if it"s protection varies in range), could reconcile two formally differing ethical positions: * pathocentrism, which presumes the ability to feel in all of life, and * biocentrism, intending to do justice to each life-form according to it"s abundance and intensity. PMID- 11107325 TI - [What is "ethical weighing" in animal testing?] AB - Ethical justification of legally required testing procedures with respect to pain, suffering or harm inflicted on animals are frequently being trivialised to meaningless phrases with a predictable outcome. Most applicants will not realise what kind of burdens they are placing upon ethics as an instrument for justification. Also, in ethical theory "defensibility" of animal testing is much less rigorously contemplated as it is, or would be, in comparable human testing. For these reasons, the article presented is critically investigating judicial and legal interpretations of the term "ethical weighing". In cases where true progress for the cause of animals becomes evident it chooses a rather unusual route based on the ground work of other scientists. The centre of attention is not given to more or less fictions cost/benefit calculations or point-systems but rather to the moral self- examination or possible self-justification of the scientist, meaning: a true reflection on ethics. PMID- 11107326 TI - [Is brain surgery on primates in basic research morally acceptable?] AB - In the contemporary controversy about the legitimacy of vivisection a few basic assumptions are shared by nearly all participants of the discussion. (I) Pure Research in the service of medicine is of great value for humankind. It contributes to prolonging human life and the alleviation and prevention of human suffering. (II) Brain surgery for the sole purpose of pure research is morally unacceptable in the case of any human being. (III) Primates are sensitive beings which lead a rich social life and are endowed with remarkable intellectual capacities. (IV) Primates have a moral standing, possibly to a lesser degree compared with human beings, certain acts are therefore an injustice toward them. The controversy then is about the question whether premise (I) outweighs (IV), i.e. whether the benefit of the pure research is from a moral point of view more important than the suffering of innocent primates. I shall present four arguments against such a conclusion. 1) According to premise (I) brain surgery on human beings for the sole purpose of pure research is morally unacceptable. Since this prohibition is meant to include all human beings it cannot rest on the exclusive human possession of reason because e.g. some mentally handicapped human beings lack this faculty. All other properties which may be named as basis for the ascription of a moral status which forbids brain surgery for pure research, are possessed also by some animals, especially primates; therefore it is impossible to deny them the same moral status. 2) Brain surgery on primates is confronted with an insoluble dilemma: If the characteristics of the primate brain are very similar to that of human beings, the scientific benefit is obvious, but the procedure appears to be morally unacceptable exactly because of this similarity. If, on the other hand, the characteristics differ significantly, brain surgery may seem legitimate but the scientific benefit becomes doubtful at best. 3) We could quite easily save hundreds of human lives if e.g. speed limits would be reduced (say) by half. Most of us, however, are unwilling to accept such a loss of quality of life in order to save a certain number of human lives. Since we are no prepared to pay this comparatively modest price, we have, in my eyes, no moral right to impose considerable pain and suffering on a primate to save human lives. 4) Pure research in the service of human medicine is from a moral point of view of great importance. Since most of the work in this area is done or financed by private corporations and not by state institutions, from a economical point of view the aim consists in making profit. Since the latter aspect has gained more and more weight in the last years the moral worth of pure research cannot rule out any other moral concern. PMID- 11107327 TI - [Experience of suffering. Disputable considerations on animal ethics] AB - In the current debate on the protection of animals, sentientist approaches dominate. Recourse to the ability of animals to suffer is intended to indicate and demonstrate their moral relevance. Although sentientist arguments appear in very different argumentative contexts, they all presuppose the particular significance of suffering, or rather, the ability to suffer for the imperative to take animals into moral consideration. This article questions this basic presupposition with a view to determining more precisely the structure of the relationship between suffering and moral relevance. Initially, a number of basic issues with respect to ethics and morality are considered. Then the essential presuppositions of sentientism are investigated. On the one hand, the question is addressed as to how the notion of suffering can be defined meaningfully with respect to its appropriateness as a moral criterion. On the other, it is shown that the moral relevance of suffering requires its own proof and should not be assumed a priori. Finally, a systematically developed ethical position on the protection of animals is introduced and sketched out with respect to its practical consequences. PMID- 11107328 TI - [Animal welfare and church] AB - In the author"s opinion the potential and the social importance of churches are underestimated especially regarding the practical animal welfare. Nevertheless the protestant and catholic church are still associations with a great number of members and have an infrastructure, which is unmatched. Even concerning the content there are clear positions from the church concerning the "relations between human being and animal" or rather the "responsibility which man takes for the animal as a co-creature", as numerous publications show. As a result each sort of co-operation with parishes should be tried for the purposes of the animal welfare, whenever it is possible. PMID- 11107329 TI - ECVAM and the Promotion of International Co-operation in the Development, Validation and Acceptance of Replacement Alternative Test Methods. AB - The European Centre for the Validation of Alternative Methods (ECVAM) was established by the European Commission in 1991, to co-ordinate the validation of alternative methods at the European Union level, to establish a database on alternative methods, and to promote dialogue among all the interested parties, in order to secure the international recognition and acceptance of validated alternative test methods. All ECVAM activities, including workshops, task forces, in-house laboratory studies, and contracted external pre-validation and validation studies, involve international co-operation. Of particular importance has been the role played by ECVAM, notably in partnership with ZEBET, in establishing the principles of test development, validation and acceptance, defining the practical procedures necessary to optimise the progress of new tests toward acceptance, and in successfully validating the 3T3 NRU PT test for phototoxic potential. It is vital that international co-operation goes beyond mere discussion and leads to definite collaborative projects, which lead, in turn, to genuine achievements. PMID- 11107330 TI - The Funding of Research on the Three Rs in the EU. AB - In a number of European countries, notably Germany, The Netherlands, Sweden and the UK, there has been a significant commitment to research designed to achieve one or more of the Three Rs. This work has been funded by industry, by government, and by the animal welfare movement. The cosmetic industry, ZEBET in Germany and the Dutch Platform in The Netherlands, and FRAME in the UK, deserve particular mention. The European Commission also plays a major role, in two main ways. Firstly, research is supported via DGXII, particularly through programmes such as BRIDGE, BIOTECH and BIOMED in the Third and Fourth Framework Programmes (1991-1998). In the Fifth Framework Programme (1999-2002), major support for international collaborative studies on the development of replacement alternative test methods will be provided as parts of programmes concerned with the Cell Factory (novel in vitro testing as alternatives to animal testing) and Environment and Health (improvement of predictive toxicity testing, with emphasis on in vitro test systems and alternative screening and testing protocols). The second kind of funding is by competitive contracts for specific studies required by various Services of the Commission, including, for example, pre-validation and validation studies conducted for ECVAM. PMID- 11107331 TI - [Tierversuchsverbot fur Kosmetika in der EU - Quo vadis?] PMID- 11107332 TI - [Grunes Licht fur klinische Xenotransplantations-Versuche in der Schweiz] PMID- 11107333 TI - Benefit of Bt corn hybrids. PMID- 11107335 TI - Flavonoid biosynthetic pathway. PMID- 11107334 TI - Wanted: misinterpreted science. PMID- 11107336 TI - Compromised resistance in saponin-deficient plants. PMID- 11107337 TI - Policy briefs. PMID- 11107339 TI - Myths of gene patents. PMID- 11107338 TI - Plant virus RNA in glacial ice. PMID- 11107341 TI - Fungus as weedkiller. PMID- 11107340 TI - Crop gene diversity declining. PMID- 11107342 TI - Growing pains for education. PMID- 11107343 TI - NP students and the Internet. PMID- 11107345 TI - Multistate licensure for APNs. PMID- 11107344 TI - Juggling obligations. PMID- 11107346 TI - Genital herpes in adolescents. PMID- 11107347 TI - Schizophrenia in the elderly. PMID- 11107348 TI - Nurse practitioner education in the new century. PMID- 11107349 TI - Faculty practice. One nursing school's experience. PMID- 11107350 TI - Essential to the profession's future: clinical preceptors. PMID- 11107351 TI - Women, HIV and violence. A dangerous triad. PMID- 11107352 TI - Vulvar vestibulitis. PMID- 11107353 TI - Ship to shore. Heading off health problems for international travelers. PMID- 11107354 TI - Gastroesophageal reflux disease. Primary care management issues. PMID- 11107355 TI - Weighing in on clinical guidelines for obesity. Tools to assist your patients. PMID- 11107356 TI - Premenstrual syndrome. Self-care measures are part of the solution. PMID- 11107357 TI - Adult anemia. PMID- 11107358 TI - Sustaining life twice. Will umbilical cord blood replace bone marrow? PMID- 11107359 TI - Isn't it time for education reform? PMID- 11107361 TI - [Physicians and nurses compared: results of a clinical study]. AB - The results of the most recent trials show that nurses can perform some medical activities with results equal to doctors and better patients satisfaction. The 8 trials analyze patients outcomes and costs of care and on each aspect, on average, nurses' performance is satisfactory. The trials provide useful material for a wider discussion on the actual and future role of nurses. PMID- 11107362 TI - [Debate in the USA on nurses' delegation of duties to assistants]. AB - In the today's rapidly changing practice environment, more nurses than ever are facing delegation dilemmas. The main issues of the debate on delegation of task and activities from nurses to less qualified personnel are reported. Nurses can delegate tasks but remain accountable for the final result. The main rules for delegation are discussed and the need for a correct communication and education to delegate is stressed. PMID- 11107363 TI - [Meta-analysis of the Cochrane Collaboration. Promoting collaboration between nurses and physicians]. AB - The aim of this systematic review of the Cochrane Collaboration was to assess the impact of interventions designed to change nurse-doctor collaboration on collaboration itself, on patient satisfaction, and on the effectiveness and efficiency of the health care provided. There are no good trials on interventions to improve collaboration between doctors and nurses. There are numerous strategies suggested to improve inter-disciplinary collaboration between doctors and nurses, such as joint workshops, meetings, development of team systems and strategies, and training in collaboration. However, no studies of these interventions that met the reviewers' criteria could be found. More research is needed to determine any impact of these strategies on interprofessional collaboration, and on the outcomes for patients. PMID- 11107364 TI - [Nurses' perception of specific nursing activities that can be delegated to other health care personnel]. AB - In order to understand the nurses' perception of the activities specific to nursing care, and to assess what they are ready to delegate to other health care workers, a complex activity of analysis was started. Head nurses and nurses representative of the different NHS hospital and district services (hospital wards, home care and district, preventive services, psychiatric services and first aid services in the district, operating theatre and clinics) of the Medio Friuli area (North of Italy) were asked to identify a. the main activities performed in the patients' care, b. the nature of the activities (whether administrative, hotel, health, support-non health), c. their level of standardization (i.e. whether their performance required discretionality and the tailoring of the activity to the patient's situation) and d. if they considered it delegable and to whom (doctors, support personnel, secretary, other health care personnel). Final aim of the work was to acquire elements in order to reorganize the nursing care. In order to leave the nurses free to identify areas that better allowed the description of their activities, (and due to the large and different numbers of sectors and areas involved), no strict instructions were given. Therefore the model used and level of detail in the description of activities and processes varied from group to group. Sixty nurses participated in the work. On average, from 38.9% (Operating Theatre and clinics) to 55.7% (Psychiatric services) of the activities identified were perceived as delegable to nurses aids and/or secretaries. The 89.2% of delegable activities (183/205) were considered highly standardized. Up to 23.4% (39/166) were classified as health care activities (i.e. prevention of pressure sores with mobilization, risk assessment with standardized scales, application of ointments, the care for patients during mobilization etc.). The remaining were classified as administrative or support activities. Although this does not necessarily imply that with the introduction in the health care team of a secretary and of the nurse-aid half of the time of the nurse could be freed, nurses perceive that on average half of their routine and basic activities could be well delegated to other figures. PMID- 11107365 TI - [Nurses' assistants: problems and perspectives concerning the changes in the nursing profession]. AB - The 'new' figure of the nurse-assistant is at the same time desired and feared by the nursing profession. In fact nurses could easily delegate some of their tasks and at the same time this would reduce their workload but also the number of nurses needed. The discussion on what could be delegated is still on the floor and there is urgent need of redefining the content of the work of the nurses. An open discussion also with the doctors is warranted, together with a careful analysis of how the problem has been dealt with in other countries. PMID- 11107366 TI - [Comparison of two different protocols on change of medication in central venous catheterization in patients with bone marrow transplantation: results of a randomized multicenter study]. AB - Care of central venous catheter (CVC) in patients undergoing bone marrow transplantation (BMT) raises significant problems related to the high risk of local infections, to the immunodeficient status, which in itself is a predisposing factor for systematic blood stream infections. Although frequent changes of CVC dressing might theoretically reduce the incidence of infections, they are also accompanied by significant skin toxicity and patient discomfort. No study has yet addressed these points. The objective of this study was to compare two different time interval protocols for CVC dressing, in order to assess the effects on local infections and toxicity. In a multicentre study, 339 bone marrow transplant (BMT) patients with a tunnelled CVC (group A, 230 pts) or a non tunnelled one (Group B, 169 patients) were randomly allocated to receive CVC dressing changes every 5 or 10 days if belonging to group A or 2 or 5 days if in group B. Transparent impermeable polyurethane dressings were used for all patients. The rate of local infection at the site of CVC insertion was assessed by microbiological assay every 10 days, while severity of skin toxicity was measured according to the ECOG scale. Sixty-five per cent of enrolled patients were finally evaluable. Patients (in both groups) receiving CVC dressing changes at longer intervals did not show a significant increase in the rate of local infections, while those who received dressing every two days had a significant increase in local skin toxicity. Longer intervals were accompanied by a reduction in costs. The results of this study demonstrate that the increase in time interval between CVC dressing changes in BMT patients did not increment the risk of local infections, while significantly reducing patients discomfort and costs. PMID- 11107367 TI - [Estimating the grade of patient satisfaction at the bone marrow transplantation department in Florence hospitals]. AB - The satisfaction of the patients admitted to the bone marrow transplant unit of Careggi Hospital was evaluated by the nursing team. The aim of the evaluation was to measure the level of satisfaction for the nursing care and services and the areas of improvement. The questionnaire, with 23 questions referring to 5 areas (hotel care, Nurses' reliability, Ability to reassure, to answer to patients' needs and Empathy) derived from the conceptual model of Servqual. Ninety patients were given (or mailed) the questionnaire during a follow-up visit. Patients were asked to answer the questions evaluating each aspect on a scale from 1 (falls short of expectation) to 10 (exceeds all expectations). The answers show a very high satisfaction (> 8) for all the areas except for the food that reported a medium score of 5.2. Further analysis will allow a better understanding of the causes of dissatisfaction. PMID- 11107368 TI - [Alzheimer disease: drug therapy and/or nursing care]. PMID- 11107369 TI - [The role of control of household mites in the treatment of asthma: a meta analysis]. PMID- 11107370 TI - Midwifery policies, position statements and standards--how do we know that we meet them? PMID- 11107371 TI - They were different and few: an Australian study of midwives' attitudes to research and computerised research findings. AB - This study sought to determine factors influencing the utilisation of research findings by Australian midwives before and after exposure to an on-line research database within their practice setting. The setting was a large maternity hospital in South Australia and data were collected via pre and post intervention questionnaires. 14 midwives from the Delivery Suite and Birth Centre of this hospital participated in the study. Midwives showed an increased receptivity to the concept of computerised research findings after exposure to the database. They rated research as highly important to providing quality midwifery care and for the midwifery profession, and saw midwives as the key players in maternity care research. These findings are not in keeping with previous research--largely accounted for, we believe, by the particular, and small number of, midwives who volunteered to participate in the study (24% of those invited). PMID- 11107373 TI - Midwives you've got m@il! PMID- 11107372 TI - Anxiety and functional status after childbirth. PMID- 11107374 TI - Breastfeeding duration of Thai women. AB - This quantitative study aimed to determine Thai working women's planned intention to breastfeed and to identify why the mothers chose to breastfeed their infants, how long they initially planned to breastfeed and how long they actually breastfeed. Ninety-nine Thai working mothers who had breastfed within the last 18 months, or who were still breastfeeding their infants and who used the services at three clinics in Ubon Ratchathani Province, Northeast Thailand were asked to participate in the study. The study findings show a strong correlation between the women's planned duration of breastfeeding and between the length of time she actually breastfed. The authors recommend that breastfeeding educational programs include partners and that the work environment provides support facilities to encourage and support the continuation of breastfeeding. PMID- 11107375 TI - Tribute to Henny Ligtermoet--birth activist and midwife's friend. PMID- 11107376 TI - Does evidenced-based practice medicalize midwifery care? Part 1. AB - In this paper evidence-based care is defined. The evidence to support the provision of care by midwives is presented, as is the evidence to support home birth for those women at low obstetric risk. In conclusion midwives are challenged to be political and use this evidence to support changes to improve the quality of care provided to women and their families. PMID- 11107377 TI - Do statistics affect patients' decisions? PMID- 11107378 TI - Gynecologic cancers: Part. I--Risk factors. AB - Gynecologic cancers will account for approximately 81,000 new cases of cancer this year. Although much is known about the risk factors for cervical, ovarian, and endometrial cancers, less is known about vaginal and vulvar cancer risk factors. Generally, risk factors and associations for gynecologic cancers are behavioral, reproductive, hormonal, and genetic related. Research continues to verify and refute the impact of certain factors. All nurses must be knowledgeable about the risk factors and associations for these cancers. PMID- 11107379 TI - Gynecologic cancers: Part. II--risk assessment and screening. AB - Screening interventions for gynecologic cancers involve identifying risk factors and high-risk groups of women, counseling for risk factor reduction, and recommending early detection strategies. All nurses can conduct gynecologic risk assessments, and advanced practice nurses can perform physical exams. All women must be knowledgeable about risk factors because gynecologic cancers are curable when diagnosed in the early stages. Nursing interventions include developing culturally sensitive programs and educational materials for targeted populations and educating women in all groups to raise awareness about gynecologic cancer risks. PMID- 11107381 TI - The impact of an ostomy on sexuality. AB - Creation of an ostomy results in physiological and psychological changes that affect sexuality. Major physiological complications for men include erectile dysfunction and ejaculatory difficulties. For women, dyspareunia is the most common physiological complication. The presence of an ostomy can alter a person's body image, which, in turn, influences the desire for sexual activity. Sexuality concerns should be addressed with all patients undergoing ostomy placement. The PLISSIT model, which outlines four stages of interventions used in sexual counseling, can be used to guide nursing care. PMID- 11107380 TI - Development of a protocol to prevent opioid-induced constipation in patients with cancer: a research utilization project. AB - Opioids are the major class of analgesics used in the management of moderate to severe cancer pain, and constipation is a common side effect of opioid administration. While monitoring for quality-assurance, nurses found that 95% of patients interviewed on a 28-bed oncology unit of a Midwestern hospital reported constipation as the major side effect of their opioid regimen for pain control. Through the efforts of a nursing research utilization committee, a protocol to prevent opioid-induced constipation in patients with cancer was developed and implemented. PMID- 11107382 TI - Carotid artery rupture. PMID- 11107383 TI - Herbs. PMID- 11107384 TI - Thalidomide. PMID- 11107385 TI - Controlling lymphedema-related swelling. PMID- 11107386 TI - Managing patients with a distended bladder. AB - A distended bladder is a serious medical situation that requires prompt intervention to drain urine and promote patient comfort. The presence of suprapubic pain distinguishes acute urinary retention from chronic retention. After decompression of the bladder, patients must be monitored closely for post decompression complications. Additional nursing management may include preparing patients for surgery or additional diagnostic tests. PMID- 11107387 TI - Tackling the stereotypes. PMID- 11107388 TI - Invisible barriers. PMID- 11107389 TI - Poor relations. PMID- 11107390 TI - Everyday heroes. PMID- 11107391 TI - Dobson's returners. PMID- 11107392 TI - Working breakfast. PMID- 11107393 TI - Money's not everything. PMID- 11107394 TI - Not so 'babywise'. PMID- 11107395 TI - Hand in hand. PMID- 11107396 TI - Working it out. PMID- 11107397 TI - Lessons in teamwork. PMID- 11107398 TI - The role of the clinical nurse specialist: a study. PMID- 11107399 TI - Clinical supervision for professional practice. PMID- 11107400 TI - Understanding oesophageal varices. AB - This article describes the aetiology of oesophageal varices, the variety of treatment options available and the physiological and psychological nursing needs of patients undergoing these treatment regimes. Its purpose is to produce a balanced overview, looking at nursing care in the acute period and the long-term support required to meet the complex needs of these patients and their carers. After reading this article you should be able to: Identify what lifestyle risk factors are associated with patients who have oesophageal varices. Describe the underlying physiology that promotes the development of oesophageal varices. State what treatment options are available and their associated complications. Recognise the needs of carers and staff when attending to this client group. Prepare an action plan for the long-term support of a patient prone to further bleeding episodes. PMID- 11107401 TI - Using antibiotics. PMID- 11107402 TI - States of emergency. PMID- 11107404 TI - Nursing thrives on humour, and the camaraderie nurses share makes the job worth doing. PMID- 11107403 TI - A high-flying career. PMID- 11107405 TI - Bills, skills and councils. PMID- 11107406 TI - Can things only get better? PMID- 11107407 TI - One hundred years of attitude. PMID- 11107408 TI - Pro-life organisations are using shock tactics to stop women going ahead with an abortion. PMID- 11107409 TI - The justified outrage over the removal of dead children's organs threatens to do serious damage to medical research. PMID- 11107410 TI - Read those labels carefully. PMID- 11107411 TI - The last laugh. PMID- 11107412 TI - It's a funny old job. PMID- 11107413 TI - Santa strikes back. PMID- 11107414 TI - Stress busters. PMID- 11107415 TI - Nurse heroes of the century. Interview by Jeanette McDonald. PMID- 11107416 TI - Return of the natives. PMID- 11107417 TI - NT/3M National Nursing Awards. Family favourite. PMID- 11107419 TI - The heart, Part Six: Coronary artery disease--2. PMID- 11107418 TI - Nurses in Essex are training hard to be ready for the millennium bug. PMID- 11107420 TI - Smoothing the journey home. PMID- 11107421 TI - Research and practice: making connections. PMID- 11107422 TI - Different routes to professional nursing. PMID- 11107424 TI - Coaching for improving job performance and satisfaction. PMID- 11107423 TI - A strategy for recruitment and retention in perioperative care. PMID- 11107425 TI - Primary care. Keep your eyes on the prize. PMID- 11107426 TI - Primary care. What a difference a day makes. PMID- 11107427 TI - Primary care. Can you keep a secret? PMID- 11107428 TI - [Anesthesia in the aged]. PMID- 11107429 TI - [Treatment of carotid stenosis in the aged]. PMID- 11107430 TI - [Laparoscopic surgery in the aged]. PMID- 11107431 TI - [Shoulder prosthesis in the aged]. PMID- 11107432 TI - [Artificial sphincter in urinary incontinence]. PMID- 11107433 TI - [The kitchen environment, a useful therapeutic tool]. PMID- 11107434 TI - [The project of "nutrition host" in the geriatric unit of community hospices in Beaune]. PMID- 11107435 TI - [Therapeutic touch as part of the care for the aged]. PMID- 11107436 TI - [How to facilitate self care for the aged?]. PMID- 11107437 TI - [Living with an Alzheimer patient]. PMID- 11107438 TI - [Paget disease, essential clinical features]. PMID- 11107439 TI - Synergism of cytotoxic effects of vinorelbine and paclitaxel in vitro. AB - Empiric combinations of vinca alkaloids with taxanes have been recently used in clinical oncology. To enhance the activity of these two classes of agents, we evaluated the sequence and duration of exposure, looking for synergistic effects. Cell lines DU 145, PC 3, LnCaP, LL 86, MCF7wt, and MCF7/ADR (NCI/ADR-RES) were incubated with varying concentrations of paclitaxel or vinorelbine. Cytotoxicity was evaluated by a semiautomated MTT (3-[4,5-dimethylthiazole-2-yl]-2,5 diphenyltetrazolium bromide) method. Synergism or antagonism of these two agents either sequentially or in combination was determined by median effect analysis. Prolonged exposure of cells to either drug enhanced cytotoxic effect. Synergism or antagonism with vinorelbine and paclitaxel were both sequence dependent and cell line specific. In the case of MCF7wt, synergism was seen when a 48-hr exposure to vinorelbine preceded paclitaxel, whereas antagonism was noted when both agents were applied simultaneously or when the sequence was reversed. Concurrent vinorelbine and paclitaxel were synergistic in four of six cell lines when the exposure was extended to 96 hr but not for shorter durations of exposure. Sequential exposure of vinorelbine preceding paclitaxel or prolonged exposure to both agents concurrently needs to be tested clinically to determine whether the antitumor activity of this combination can be enhanced. In addition, these studies suggest concurrent administration of these two agents may lead to a less than optimal cytotoxic result. PMID- 11107440 TI - Apoptosis and cell proliferation capacity in AKR lymphoma malignancy variants. AB - Dysregulation in apoptotic cell death has recently emerged as a factor in tumorigenesis, but its effect in tumor progression is not yet established. In the present study we evaluated the levels of proliferative and apoptotic cell fractions in a T-cell lymphoma tumor progression model. We compared these features and the expression of apoptosis-related genes in primary tumors of several AKR lymphoma malignancy variants. According to DNA flow cytometry, a considerable proportion of cells (35-40%) was in the proliferative (S + G2/M) phase in all variants, but a slight augmentation with increasing malignancy was noted. Apoptotic cell content was, unexpectedly, the lowest in the less malignant variant. This might be due to the higher content in macrophages observed in this variant, which possibly partly eliminated apoptotic bodies. We found an increase in bcl-2 level with increasing malignancy that was probably counterbalanced by the simultaneous increase observed in the Fas receptor. PMID- 11107442 TI - The phosphoprotein Op18/stathmin is differentially expressed in ovarian cancer. AB - To identify potential prognostic indicators of ovarian cancer and identify targets for therapeutic strategies, mRNA differential display was used to analyze gene expression differences in normal, benign, and cancerous ovarian tissue. One cDNA isolated by this technique, Op18/stathmin, is a highly conserved gene that is reported to have many different functions within a cell, including signal transduction, control of the cell cycle, and the regulation of microtubules. Quantitative Northern blot analysis of 12 malignant ovarian samples, 8 benign ovarian tumors, and 10 normal ovarian tissue samples demonstrated overexpression of Op18/stathmin mRNA in the malignant cancers. Immunohistochemistry showed an apparent overexpression of Op18/stathmin protein level and an association with proliferating cells. PMID- 11107441 TI - Expression of c-jun and c-fos in apoptotic cells after DNA damage. AB - Apoptosis was induced in HeLa cells by exposure to 50 microM N-methyl-N'-nitro-N nitrosoguanidine (MNNG) for various time intervals (up to 120 min). Apoptotic death was confirmed by the microscopic observation of cell blebbing, cell granulation, and cell aggregation. Cells also showed loss of phospholipid symmetry as judged by immunofluorescent microscopy with fluorescently labeled phosphatidyl serine-specific annexin V. In addition, staining of cells with ethidium bromide showed the presence of genomic DNA apoptotic bodies. The protein expression levels of c-jun and c-fos increased in DNA-damaged HeLa cells after MNNG treatment in a time-dependent fashion. Although the levels of c-fos increased rapidly during the first 30 min and remained high for 2 hr, the increase in c-jun expression was more gradual and slower (60-120 min) after MNNG treatment. These results are consistent with the conclusion that c-fos is important in the initial stages (commitment phase) of apoptosis and c-jun is involved in the late stages (execution phase) of apoptosis induced with alkylating agents. PMID- 11107443 TI - Protein kinase C modulation and anticancer drug response. PMID- 11107444 TI - Gene therapy in human cancer: report of human clinical trials. PMID- 11107445 TI - Oral fluoropyrimidines in cancer treatment. PMID- 11107446 TI - Palliative medicine: an emerging field of specialization. PMID- 11107448 TI - Spontaneously occurring tumors of companion animals as models for human cancer. AB - Spontaneous tumors in companion animals (dog and cat) offer a unique opportunity as models for human cancer biology and translational cancer therapeutics. The relatively high incidence of some cancers, similar biologic behavior, large body size, comparable responses to cytotoxic agents, and shorter overall lifespan are the factors that contribute to the advantages of the companion animal model. The tumor types that offer the best comparative interest include lymphoma/leukemia, osteosarcoma, STS, melanoma, and mammary tumors. With the increase in new therapeutic agents (traditional chemotherapy, gene therapy, biologic agents, etc.), the companion animal model can provide useful populations to test new agents where efficacy and toxicity can be examined. PMID- 11107447 TI - Molecular and clinical advances in core binding factor primary acute myeloid leukemia: a paradigm for translational research in malignant hematology. AB - Clonal chromosomal abnormalities are the most important prognostic indicators in acute myeloid leukemia (AML). Recent advances in molecular biology have allowed structural and functional characterization of many of these genomic rearrangements and have provided evidence for their primary role in leukemogenesis. Two of the most prevalent cytogenetic subtypes of adult primary or de novo AML, t(8;21)(q22;q22) and inv(16)(p13q22), are characterized by disruption of the AML1(CBF alpha 2) gene at 21q22 and the CBF beta gene at 16q22, respectively. Both genes encode a subunit of core binding factor (CBF), a regulator of normal hematopoiesis. At the molecular level, t(8;21)(q22;q22) and inv(16)(p13q22) result in the creation of novel fusion genes, AML1/ETO and CBF beta/MYH11, whose structures and functions are being successfully characterized by in vitro studies and transgenic animal models. Detection of t(8;21)(q22;q22) or inv(16)(p13q22) in adult patients with primary AML is a favorable independent prognostic indicator for achievement of cure after intensive chemotherapy or bone marrow transplantation and may serve as a paradigm for risk-adapted treatment in AML. The purpose of this review is to summarize the recent advances in the molecular biology and clinical management of t(8;21)(q22;q22) and inv(16)(p13q22) primary AML, collectively referred to here as CBF AML. PMID- 11107449 TI - Should women receiving tamoxifen be screened for endometrial cancer? An argument for screening. PMID- 11107450 TI - Should women receiving tamoxifen for breast cancer be screened for endometrial cancer? PMID- 11107451 TI - Greater expectations in a cancer trial: absolute more than relative survival increases, community more than academic clinicians. AB - There is no consensus on how the difference between control and experimental outcome rates, the clinically important difference, should be estimated when designing a clinical trial. We sought to determine whether community and academic clinicians had different perceptions as to what would constitute a clinically important increase in survival, when asked to respond in absolute or relative terms, before a trial was started rather than when the results were already known. A telephone survey of 25 practicing Canadian oncologists was performed. Questions were asked as to the importance of acceptable and minimally acceptable improvements in survival for a hypothetical trial of pancreatic cancer where the baseline survival was expected to be between 2 and 8 months. Responses were sought for absolute (additional months) or relative gains (percent improvement) in survival. The mean absolute additional survival expectations corresponded to at least a doubling of baseline survival and tended to be greatest when the prognosis was poorest (p = 0.06). Relative expectations for improved survival varied with baseline survival (p < 0.001). When improvement in survival was requested in relative terms, the median expected improvement was 25%. This is highly significantly different than when survival improvements were requested in absolute terms (p < 0.0001). Median absolute survival expectations were greater for community as compared with academically affiliated physicians (p = 0.046). We found that physicians are inconsistent in their interpretation of qualitative data. What constitutes a potentially clinically important treatment effect differs whether viewed in relative or absolute terms before the performance of a trial. Expectations were greatest when the prognosis was poorest and differed between community and academic physicians. PMID- 11107452 TI - Timing of antitubulin therapy: does it matter? PMID- 11107453 TI - Politics and the common good. PMID- 11107454 TI - Validity of utilization review tools. PMID- 11107455 TI - Validity of utilization review tools. PMID- 11107456 TI - Validity of utilization review tools. PMID- 11107457 TI - Validity of utilization review tools. PMID- 11107458 TI - Validity of utilization review tools. PMID- 11107459 TI - Intervening to reduce weight gain in pregnancy and gestational diabetes mellitus in Cree communities: an evaluation. AB - BACKGROUND: A high prevalence of gestational diabetes mellitus and type 2 diabetes has been observed among the Cree of James Bay, Quebec. To address this problem, a diet and activity intervention during pregnancy, which was based on social learning theory, was initiated in 4 Cree communities. METHODS: A prospective intervention compared dietary, weight and glycemic indicators for 107 control subjects and for 112 women who received the intervention during the course of their pregnancy. A control period in 4 communities (July 1995-March 1996) was followed by an intervention period (April 1996-January 1997) when subjects were offered regular, individual diet counselling, physical activity sessions and other activities related to nutrition. RESULTS: The intervention and control groups did not differ at baseline regarding their mean age (24.3 years [SD 6.29] v. 23.8 years [SD 5.86]), mean prepregnancy weight (81.0 kg [SD 19.46] v. 78.9 kg [SD 17.54]) and mean gestational age at recruitment (17.1 weeks [SD 7.06] v. 18.5 weeks [SD 6.92]). The intervention did not result in differences in diet measured at 24-30 weeks' gestation, rate of weight gain over the second half of pregnancy (0.53 kg per week [SD 0.32] v. 0.53 kg per week [SD 0.27]) or plasma glucose level (50 g oral glucose screen) between 24 and 30 weeks (7.21 mmol/L [SD 2.09] v. 7.43 mmol/L [SD 2.10]). Mean birth weights were similar (3741 g [SD 523] v. 3686 g [SD 686]), as was maternal weight at 6 weeks post partum (88.1 kg [SD 16.8] v. 86.4 kg [SD 19.0]). The only changes in dietary intake were a reduction in caffeine (pregnancy) and an increase in folate (post partum). INTERPRETATION: This intervention had only a minor impact on diet; finding ways of encouraging appropriate body weight and activity levels remains a challenge. PMID- 11107460 TI - Discussions of "code status" on a family practice teaching ward: what barriers do family physicians face? AB - BACKGROUND: Patients want physicians to ascertain their wishes related to resuscitation, yet such discussions of "code status" are often delayed in the hospital setting, which compromises patient autonomy. Few studies have examined family physicians' views on this topic. Our objectives were to explore the experiences of family physicians and family practice residents in establishing code status with their patients who had been admitted to hospital and to identify barriers to these discussions. METHODS: Semistructured, in-depth interviews were conducted with 5 family physicians and 5 family practice residents admitting patients to a family practice teaching ward in a university-affiliated urban tertiary care hospital. Interview transcripts were analysed inductively, and grounded theory was used to identify conceptual categories and recurring themes. Key findings were validated by means of member checking with participants, consensus meetings of the research team and consultation with qualitative researchers. RESULTS: Barriers to code-status discussions included personal discomfort with confronting mortality, fear of damaging the doctor-patient relationship or harming the patient by raising the topic of death, limited time to establish trust, and difficulty in managing complex family dynamics. In spite of these challenges, family physicians and residents viewed discussions of resuscitation as a significant part of their role. INTERPRETATION: Family physicians and residents need to develop personal awareness about difficulties in confronting mortality, enhance their communication strategies for broaching the topic of code status in the context of a trusting doctor-patient relationship and sharpen their skills in understanding and managing family dynamics related to end of-life decisions. Awareness of the barriers to code-status discussions can inform research, education and hospital policy. Consultation with patients is needed to develop effective communication strategies. PMID- 11107462 TI - Planning research for greater community involvement and long-term benefit. Special working group of the Cree Regional Child and Family Services Committee. PMID- 11107461 TI - Lipid screening to prevent coronary artery disease: a quantitative evaluation of evolving guidelines. AB - BACKGROUND: There is strong evidence to support the treatment of abnormal blood lipid levels among people with cardiovascular disease. Primary prevention is problematic because many individuals with lipid abnormalities may never actually develop cardiovascular disease. We evaluated the 1998 Canadian lipid guidelines to determine whether they accurately identify high-risk adults for primary prevention. METHODS: Using data from the Lipid Research Clinics and receiver operating characteristic (ROC) curves, we compared the diagnostic performance of the 1998 lipid guidelines when risk factors for coronary artery disease (CAD) were counted versus calculating risk using Framingham risk equations. We also compared the diagnostic accuracy of the 1998 guidelines with guidelines previously published by the National Cholesterol Education Program in the United States and the 1988 Canadian Consensus Conference on Cholesterol and then used Canadian Heart Health Survey data to forecast lipid screening and treatment rates for the Canadian population. RESULTS: The Framingham risk equations were more accurate than counting risk factors for predicting CAD risk (areas under the ROC curves, 0.83 [standard deviation (SD) 0.02] v. 0.77 [SD 0.03], p < 0.05). Risk counting was a particularly poor method for predicting risk for women. The 1998 Canadian guidelines identified high-risk individuals more accurately than the earlier guidelines, but the increased accuracy was largely due to a lower false positive rate or a higher true-negative rate (i.e., increased test specificity). Using the 1998 lipid guidelines we estimate that 5.9 million Canadians currently free of cardiovascular disease would be eligible for lipid screening and 322,705 (5.5%) would require therapy. INTERPRETATION: Calculating risk using risk equations is a more accurate method to identify people at high risk for CAD than counting the number of risk factors present, especially for women, and the 1998 Canadian lipid screening guidelines are significantly better at identifying high risk patients than the 1988 guidelines. Many of our findings were incorporated into the new 2000 guidelines. PMID- 11107463 TI - Screening colonoscopy: is it time? PMID- 11107464 TI - Clinical practice guidelines and the translation of knowledge: the science of continuing medical education. PMID- 11107465 TI - Do you relish interesting times? Call for applicants for the 2001 CMAJ Editorial Fellowship. PMID- 11107466 TI - Rheumatology: 10. Joint replacement of the hip and knee--when to refer and what to expect. PMID- 11107467 TI - The primary care clinic as a setting for continuing medical education: program description. AB - The Mexican Institute of Social Security (IMSS) is Mexico's Largest state financed health care system, providing care to 50 million people. This system comprises 1450 family medicine clinics staffed by 14,000 family physicians, as well as 240 secondary care hospitals and 10 tertiary care medical centres. We developed a program of continuing medical education (CME) for IMSS family physicians. The program had 4 stages, which were completed over a 7-month period: development of clinical guidelines, training of clinical instructors, an educational intervention (consisting of interactive workshops, individual tutorials and peer group sessions), and evaluation of both physicians' performance and patients' health status. The pilot study was conducted in an IMSS family medicine clinic providing care to 45,000 people; 20 family physicians and 4 clinical instructors participated. The 2 main reasons for visits to IMSS family medicine clinics are acute respiratory infections and type 2 diabetes mellitus. Therefore, patients being treated at the clinic for either of these illnesses were included in the study. The sources of data were interviews with physicians and patients, clinical records and written prescriptions. A 1-group pretest posttest design was used to compare physicians' performance in treating the 2 illnesses of interest. We found that the daily activities of the clinic could be reorganized to accommodate the CME program and that usual provision of health care services was maintained. Physicians accepted and participated actively in the program, and their performance improved over the course of the study. We conclude that this CME strategy is feasible, is acceptable to family physicians and may improve the quality of health care provided at IMSS primary care facilities. The effectiveness and sustainability of the strategy should be measured through an evaluative study. PMID- 11107468 TI - Cutaneous blastomycosis in New Brunswick: case report. AB - Blastomycosis is a fungal infection of immunocompetent hosts. We present a case of cutaneous blastomycosis acquired in New Brunswick, which provides evidence that this disease is endemic in Atlantic Canada. This case also demonstrates that the diagnosis of blastomycosis may be elusive. Perseverance, a high index of clinical suspicion and close cooperation with the microbiology laboratory may be required to diagnose this uncommon condition. PMID- 11107469 TI - Novel cases of blastomycosis acquired in Toronto, Ontario. AB - Blastomycosis a potentially fatal fungal disease, is well known from defined areas of endemicity in Ontario, primarily in the northern part of the province. We present 2 unusual cases that appear to extend the area of endemicity into urban southern Ontario, specifically Toronto. Both patients presented to a dermatology clinic with skin lesions. Chest radiography, history and general physical evaluation indicated no disease at other body sites. Both cases appeared to represent "inoculation blastomycosis" connected with minor gardening injuries and a cat scratch respectively. Atypical dissemination could not be completely excluded in either case. Neither patient had travelled recently to a known area of high endemicity for blastomycosis, nor had the cat that was involved in one of the cases. Physicians must become aware that blastomycosis may mimic other diseases, including dermal infections, and may occur in patients whose travel histories would not normally suggest this infection. PMID- 11107470 TI - The election cometh: CMAJ polls the federal parties. PMID- 11107471 TI - Private MRI clinics flourishing in Quebec. PMID- 11107472 TI - Debating Medicare from the left and right. PMID- 11107473 TI - Coping with Legionella. PMID- 11107474 TI - [Tpwards a validated use of the endocoronary stent]. PMID- 11107475 TI - [French translation and validation of the Edinburgh Questionnaire for the diagnosis of intermittent claudication]. AB - Obliterative arterial disease of the lower limbs is diagnosed by simple, reproducible, sensitive and non-invasive methods. One of these, a questionnaire for the diagnosis on intermittent limping, is a method of choice. Until recent years, the only validated questionnaire was the one proposed by the World Health Organisation. This was criticised a lot, especially for its lack of sensitivity. Recently, a Scottish group proposed an improvement in the diagnostic performance of this questionnaire by carrying out several changes. This new version, called the Edinburgh Questionnaire, has promising diagnostic qualities. The authors present a French version of this questionnaire. This French translation was validated in 105 patients referred for diagnosis of obliterative lower limb arterial disease. A sensitivity of 86.5%, a specificity of 95.6%, a positive predictive value of 91.4% and a negative predictive value of 92.9% of this French version are comparable to the results obtained with the English version. Therefore, the authors suggest using this questionnaire in epidemiological and public health studies in France. PMID- 11107476 TI - [Incidence and prognosis of atrioventricular block induced by radiofrequency ablation of intranodal reentrant tachycardia. A multicenter study]. AB - The object of this study was to assess the incidence and significance of atrioventricular block (AVB) induced by radiofrequency ablation of intranodal reentrant tachycardias. The study population was 18 patients aged 44 to 83, selected from a total population of 144 patients treated for recurrent, refractory tachycardias. These patients developed complete AVB (9 cases), 2nd degree ABV (3 cases) and 1st degree AVB (6 cases) either immediately or in the chronic phase after radiofrequency ablation. The outcomes were as follows: 1. In the 9 patients with complete AVB, the block regressed in a period ranging from 7 seconds to 5 minutes. It recurred as complete AVB 1 to 4 days later in 2 patients, and regressed again after a maximum of 10 days. One 47 year old woman had definitive complete AVB; 2. In the 3 patients with 2nd degree AVB, the block regressed within 7 days; 3. In the 6 cases of 1st degree AVB, 2 patients developed transient complete AVB the following day. The possible causes of AVB were: increased vagal tone in 1 case, ablation of the rapid pathway located in a postero-septal site in 8 cases and, in the remainder, pre-existing conduction defects. The authors conclude that transient complete AVB is common and usually has a good prognosis. Definitive complete AVB is a rare but possible (0.7%) complication of radiofrequency ablation of reentrant intranodal tachycardias; other forms of AVB generally regress quickly and, although they may recur within days, they carry a good prognosis in the following months. However, long-term follow-up remains necessary. PMID- 11107477 TI - [Long-term survival of elderly amputated vascular patients]. AB - The object of this study was to assess the 10 year outcome of patients over 70 years of age who underwent amputation for vascular diseases. The secondary objective was to determine the prognostic risk factors. One hundred and four consecutive patients having undergone a leg (16 cases) or through-thigh amputation (88 cases) were reviewed. The average age at the time of surgery was 80.7 years (+/- 6.5 years, range 70-98 years). At the time of the enquiry, there were 4 survivors (operated on average 107.7 months previously +/- 14.6 months). The survival rates at one, six, twelve months and two years were 74.1%, 48.1%, 38.5% and 27% respectively. The mean survival time was 19.2 months with a median of 6 months. Univariate analysis showed the following criteria to be statistically correlated with a poor prognosis: female gender (p = 0.008), previous psychiatric disease (p = 0.007), cachexia (p = 0.004), age of 80 or over (p = 0.025), absence of diabetes (p = 0.025). Multivariate analysis showed that men had a lower risk of death (RR: 0.591--95% CI: 0.394-0.888--p = 0.011). The comparison of subjects who died during the first year with the survivors, showed a deleterious effect of proximal amputations (p = 0.032) and absence of diabetes (p = 0.021). These results confirm the very mediocre prognosis of elderly amputated vascular patients during the first postoperative year. Thereafter, the outlook is not as bad. Female gender would seem to be a poor prognostic factor whereas the presence of diabetes could identify a subgroup with a better outlook. PMID- 11107478 TI - [Human valvular substitutes for the treatment of complex progressive endocarditis. Application to aortic, mitral and tricuspid valves]. AB - The aim of this study was to assess the immediate and long-term results of human valvular substitutes (homografts and autografts) in the treatment of complex progressive endocarditis in aortic, mitral and tricuspid valves. Since 1992, 80 patients (64 men, 16 women) aged 44 +/- 16 years (range 15 to 76 years), were treated. In 53 patients, the endocarditis involved native valves, 4 on previously plastified valves, or prosthetic valves in 27 patients. The endocarditis was recurrent 6 patients. The lesions were situated on the aortic valve (N = 59), mitral valve (N = 5), aortic and mitral valves (N = 12), aortic and tricuspid valves (N = 3), mitral and tricuspid valves (N = 1). The peroperative findings confirmed the lesions diagnosed at echocardiography: prosthetic valve dehiscence (27 patients), prosthetic cusp tear (N = 7), vegetations (N = 61), perforations (N = 48), periannular abscess (N = 47), aorto-ventricular discontinuity (N = 12), aorto-mitral discontinuity (N = 7), right ventricular aortic fistula (N = 1), aorto-pulmonary fistula (N = 1), pseudo-aneurysm (N = 1), ventricular septal defect (N = 1). Eighty-six human valvular substitutes were used (double homograft in 6 patients): aortic homograft (N = 63), pulmonary in the aortic position (N = 1), the mitral position (N = 12), of which 8 were in the mitral and 4 in the tricuspid position, pulmonary autograft (N = 10). Ten mitral valve repairs were performed on infected lesions. Associated procedures included mitral valve repair (N = 5), tricuspid valve repair (N = 1) for non-infarcted valve lesions, replacement of the ascending aorta (N = 2), the aortic arch (N = 1), coronary bypass surgery (N = 2) and one nephrectomy. The hospital mortality was 5% (4 patients). The causes of death were: infarction (N = 2), myocardial failure (N = 1) and multiorgan failure (N = 1). Four early reoperations were required for technical problems, none for endocarditis. Seventy-three of the 76 survivors were followed up for 43 +/- 24 months (range 1 to 84 months). Eight patients died during follow-up, but only 1 of cardiac causes (operation for recurrent endocarditis in a drug abuser). Seven operations were performed, 3 for technical problems or structural failure, 4 for recurrent endocarditis. At 5 years' follow up, the survival was 81 +/- 5%; 88 +/- 6% of patients were free of endocarditis, 77 +/- 6% had no reoperation: no patient had thromboembolic complications. These results show that human valvular substitutes are adapted for the treatment of complex, progressive aortic, mitral and tricuspid valve endocarditis when techniques of valvular repair are no longer feasible. PMID- 11107479 TI - [Physical reproduction of cardiac sutures. A new field of investigation in cardiology]. AB - A new technique of physical reproduction of cardiac anatomy has been developed from volumetric data and its practical value assessed in cardiological practice. The acquisition of the volumetric data was by 3D echocardiography. Parallel and equidistant 2D views were selected from this information. The images were printed at a scale adjusted to the true dimensions of the structures of interest and then stuck on a support, the thickness of which was identical to the distance between the views, and the slices were superimposed while respecting the initial orientation. This technique has been adapted secondarily to modern industrial processes of rapid prototyping (3D printing and powdering) allowing automatic tooling of models. Several physical models have been made: whole heart in end diastole, mitral valve stenosis and prolapse, atrial septal defect with insertion of a percutaneous prosthetic device, great vessels at the base of the heart. There are many possible cardiological applications of physical models: investigation of complex cardiac disease, pre- and per-operative simulation of surgical procedures, elaboration of prosthetic material, physiopathological studies, teaching and training, patient information. PMID- 11107480 TI - [Alterations of left ventricular mechanics after anterior myocardial infarction. Quantitative analysis by tissue doppler echocardiography]. AB - Myocardial ischemia is usually responsible for alterations of regional left ventricular function which are not quantified by routine echocardiography. We used tissue Doppler echocardiography to quantitate the three main components (radial, longitudinal and angular) of left ventricular function, and their alterations after acute anterior myocardial infarction. Radial function was assessed by the myocardial velocity gradient derived from M-mode color tissue Doppler echocardiography, and longitudinal function by the pulsed wave tissue Doppler echocardiography, in 26 patients who experienced a first acute anterior myocardial infarction and in 11 matched healthy subjects. We describe a new tissue Doppler method for the measurement of septal angular deformation during cardiac contraction. In healthy subjects, our results showed heterogeneous regional myocardial contraction. Septal angular deformation occurred in the same direction and was of the same order as predicted by previous MRI studies. After anterior infarction, systolic and diastolic myocardial velocity gradients were dramatically decreased in the anterior wall when compared with controls (p < 0.01), whereas the systolic velocity gradient was significantly higher in the opposite wall (p < 0.05). Pulsed wave tissue Doppler echocardiography revealed significant alterations of longitudinal left ventricular function. Septal angular deformation was decreased after anterior infarct, and a good correlation was found between the angle of rotation and the left ventricular ejection fraction by cineangiography. These results indicate that tissue Doppler echocardiography has the potential for measuring the three main components of regional left ventricular function, and for quantitatively assessing their functional alterations induced by acute myocardial ischemia. PMID- 11107481 TI - [Definitions of second degree atrioventricular block. An exercise in logic in clinical electrocardiography]. AB - Second degree atrioventricular block has received different definitions which make it difficult for physicians to interpret certain cases. This review of the literature provides an accurate definition of the criteria of Mobitz I and Mobitz II atrioventricular block, and proposes simple diagnostic methods based on conventional electrocardiography. The importance of the definitions is underlined and the consequences in terms of permanent cardiac pacing are emphasised. PMID- 11107482 TI - [Indications for automatic implantable ventricular defibrillators. French version. Bureau of Heart Rhythm Group of the French Cardiology Society]. PMID- 11107483 TI - [Information and consent card for stimulator implantation]. PMID- 11107484 TI - [Esophageal perforation during transesophageal echocardiography]. AB - Known for its reliability, transoesophageal echocardiography is an investigation which is increasingly used in cardiology, cardiac surgery and intensive care units. It is a semi-invasive investigation of which oesophageal perforation is a very rare but serious complication. Two cases of oesophageal perforation after transoesophageal echocardiography are reported out of a series of 87 oesophageal perforations treated between January 1981 and February 1999. In both cases, transoesophageal echocardiography was performed in conscious patients without known pre-existing oesophageal pathology. The presentations were acute. Both patients underwent emergency surgery. One patient is alive and the other one died one month after a second operation related to the perforation. Nine cases of oesophageal perforation have been reported after transoesophageal echocardiography. The pathogenesis, means of prevention and treatment of oesophageal perforation are discussed. PMID- 11107485 TI - [Buerger's disease and coronary disease]. AB - The authors report the case of a 33 year old man with distal occlusive arterial disease diagnosed as Buerger's disease, with two previous transient ischaemic attacks and coronary disease resulting in myocardial infarction. Coronary angiography showed narrowing of the second segment of the left anterior descending artery, occluded distally and not suitable for revascularisation. The observation of coronary artery disease is very rare in Buerger's disease and data of coronary angiography are very sparse in this context. The occurrence of myocardial infarction and the angiographic appearances of the left anterior descending artery raise the question of coronary involvement of Buerger's disease. PMID- 11107486 TI - [Septic false aneurysm of the ascending thoracic aorta. Value of covering omentum repair]. AB - The incidence of post-sternotomy mediastinitis is 1 to 5% but the mortality is about 40%. The formation of a pseudo-aneurysm is a rare complication and an infectious origin is exceptionally rare. Diagnosis must be early because of the risk of rupture. In this case, a 47 year old patient developed slowly progressive mediastinitis presenting with a pseudo-aneurysm of the aorta. Surgery comprised a retrosternal transposition of the greater omentum. PMID- 11107487 TI - [Iatrogenic dissection of the left main artery treated by direct stenting]. AB - A 70-year-old woman was referred to the department due to a symptomatic severe calcific-aortic stenosis. During pre-operative trans-radial approach coronagraphy occurred a iatrogenic dissection of the left main artery with total occlusion and cardiac arrest (asystole). Successful resuscitation is achieved with an emergency strategy of percutaneous direct stenting revascularization combined with cardiopulmonary resuscitation manoeuvres. PMID- 11107488 TI - The role of cognitive vulnerability and stress in the prediction of postpartum depressive symptomatology. AB - OBJECTIVES: The present study was designed to test the diathesis-stress components of Beck's cognitive theory of depression and the reformulated learned helplessness model of depression in the prediction of postpartum depressive symptomatology. DESIGN AND METHODS: The research used a two-wave longitudinal design--data were collected from 65 primiparous women during their third trimester of pregnancy and then 6 weeks after the birth. Cognitive vulnerability and initial depressive symptomatology were assessed at Time 1, whereas stress and postpartum depressive symptomatology were assessed at Time 2. RESULTS: There was some support for the diathesis-stress component of Beck's cognitive theory, to the extent that the negative relationship between both general and maternal specific dysfunctional attitudes associated with performance evaluation and Time 2 depressive symptomatology was strongest for women who reported high levels of parental stress. In a similar vein, the effects of dysfunctional attitudes (general and maternal-specific) associated with performance evaluation and need for approval (general measure only) on partner ratings of emotional distress were evident only among those women whose infants were rated as being temperamentally difficult. CONCLUSION: There was no support for the diathesis-stress component of the reformulated learned helplessness model of depression; however, there was some support for the diathesis-stress component of Beck's cognitive theory. PMID- 11107489 TI - Diabetes mellitus and the rate of cognitive ageing. AB - OBJECTIVES: There is some evidence that besides affecting peripheral neural function diabetes may also cause more widespread changes in the central nervous system which reduce cognitive efficiency and so, also, independence and quality of life. The present study explores whether diabetes mellitus is a compounding factor in average declines in cognitive performance observed in old age. DESIGN: A sample of diabetics and controls were compared on a battery of cognitive tasks previously used in cognitive ageing research. METHODS: Thirty-three insulin dependent (IDDM), 135 non-insulin dependent (NIDDM) diabetics and 2191 non diabetics aged between 50 and 91 years were compared on two tests of general intellectual ability, and on three tests of verbal memory. RESULTS: Overall, the combined IDDM and NIDDM groups had significantly lower average scores than the controls group on all cognitive tasks. Detailed analyses revealed most cognitive impairment for the NIDDM sub-group whose condition was managed by hypoglycaemic drugs, slightly less for those managed by diet, and no impairment for the IDDM group. These effects were independent of age, depression, socio-economic status, and presence of other illnesses. CONCLUSIONS: Together with other recent studies these data emphasize the need for early detection and effective management of diabetes in older patients. PMID- 11107490 TI - Adapting the Rivermead Behavioural Memory Test Extended Version (RBMT-E) for people with restricted mobility. AB - OBJECTIVES: The study aimed to facilitate use of the RBMT-E with people who have impaired mobility by devising substitute tasks for the route and message subtests that do not require the individual to move around, and by presenting normative data for these substitute tasks. DESIGN: A within-subjects design was used to assess a standardization sample. Participants completed two parallel versions of the tasks in counter-balanced order in two test sessions approximately 1 week apart. METHODS: Substitute versions of the route and message subtests not requiring the individual to move around the test room were devised using commercially available materials. These were administered to participants along with other RBMT-E subtests. Scores for each version were compared for the group as a whole and for subgroups divided according to age, gender and IQ. Based on the results, profile scores were derived for each task using box plot analysis. The participants (N = 111) were part of the standardization sample for the RBMT-E and comprised consecutive series of recruits in two centres, Cambridge (UK) and Sydney (Australia). RESULTS: Normative data are presented in the form of mean scores for the group as a whole, for men and women, for people aged under 30, 30 to 50, and over 50 years, and for people with an IQ of either 90-110 or over 110. Profile score transformations for the substitute tasks are provided. CONCLUSIONS: Where mobility problems preclude the use of the route and message subtests of the RBMT-E, the 'model' tasks described here can be substituted and profile scores calculated. This allows clinicians to obtain a full spectrum of subtest scores for the RBMT-E with mobility-impaired patients, thus allowing the calculation of a total profile score. PMID- 11107491 TI - The transtheoretical stages of change as a predictor of premature termination, attendance and alliance in psychotherapy. AB - OBJECTIVE: The Transtheoretical Model proposes that clients' stage of change will predict their engagement in therapy. This study tested the hypotheses that clients with high Precontemplation scores would prematurely terminate from therapy and that clients with high Action scores would attend a greater number of sessions and establish a more positive therapeutic alliance. DESIGN: Using a within-subjects design, baseline measures of stage of change were used to predict the number of sessions attended, premature drop-out from therapy, and the therapeutic alliance after sessions one and three. METHODS: Sixty clients completed the Stages of Change Scales and the Brief Symptom Inventory prior to therapy. The therapeutic alliance was assessed after the first and third therapy session using the Agnew Relationship Measure. RESULTS: The specific hypotheses were not supported. Premature termination from therapy was predicted by high level of initial symptom severity and low Contemplation scores. Positive therapeutic alliance at session one and session three was predicted by high Contemplation at baseline. CONCLUSIONS: These results suggest that the Transtheoretical Model, which was developed in the context of interventions for behavioural change, may not be directly generalizable to outpatient psychotherapy populations. However, Contemplation did predict premature termination and engagement. This implies that with adaptation the model may be a useful adjunct to psychotherapy assessment. PMID- 11107492 TI - Patient-rated therapeutic relationship and outcome in general practitioner treatment of psychological problems. AB - OBJECTIVE: To explore the association between therapeutic relationship and clinical outcome in general medical practitioner (GP) treatment of emotional problems. DESIGN: Correlational analyses of patient and observer measures of GP consultations with 3-month clinical outcomes. METHOD: Patients of nine GPs, presenting with emotional problems as assessed by the 30-item General Health Questionnaire (GHQ-30), were interviewed and completed a questionnaire about their experience of their consultation with the GP. The audiotaped consultations were coded by external observers for patient involvement, doctor empathy and amount of the consultation that involved listening interactions (patient talking and doctor listening). After 3 months the patients completed a postal follow-up questionnaire including the GHQ-30 and a single item measure of patient perceived change. RESULTS: Fifty-seven patients provided follow-up data. Interview and questionnaire measures of patient perceptions of their relationship with the GP in the consultation predicted both GHQ-30 outcome at 3 months (partial correlations controlling for initial GHQ-30, r = .30 and r = .36) and patient perceived change on the single item measure. Observer ratings of patient involvement and doctor empathy predicted patient rated change on the single item measure, but not follow-up GHQ-30. Observer coded listening interactions were unrelated to both outcome measures. CONCLUSION: The results indicate an association between patient perceptions of how GP and patient related in the consultation and reduction in symptom severity 3 months later. PMID- 11107493 TI - Perception, mental imagery and reality discrimination in hallucinating and non hallucinating schizophrenic patients. AB - OBJECTIVES: In this study the hypothesis was tested that hallucinations result from confusing external and internal stimulus sources, i.e., perception and imagery, respectively. DESIGN AND METHODS: Thirteen hallucinating and 19 non hallucinating schizophrenic patients, as well as 14 control participants performed multiple tests of perception, vividness of mental imagery and the ability to discriminate between them (reality discrimination). These functions were tested in both the auditory and the visual modalities. RESULTS: There were no group differences on perceptual acuity. The results on one imagery task indicated that for the hallucinating patients, the relative, but not the absolute, level of vividness of mental images might be higher in the auditory modality, which was the modality in which 12 of the patients also experienced hallucinations, than in the visual modality. Finally, there was a positive relationship between severity of (auditory) hallucinations and reality discrimination problems. CONCLUSIONS: Hallucinations may result from increased vividness of mental imagery, and their severity increases with larger impairments in reality discrimination. It is recommended that research into, and cognitive behavioural therapy for, hallucinations should also focus on their sensory qualities. PMID- 11107494 TI - Comments on the content of persecutory delusions: does the definition need clarification? AB - Diagnostic criteria for subtypes of delusional beliefs based upon content have rarely been the subject of comment. In this article, several influential accounts of persecutory delusions are reviewed; differences and difficulties are noted, and their potential effect on cognitive psychological investigations discussed. One method of ensuring that researchers study similar phenomena is to use a more detailed definition than currently available, and therefore a new set of criteria is offered. Finally, related methodological problems in this emerging research area are highlighted. The issues discussed may stimulate further research on the content of delusional beliefs. PMID- 11107495 TI - Symptom severity, personal and social variables after armed robbery. AB - OBJECTIVES: To investigate the relative importance of personal and social variables on post-trauma symptom recovery. DESIGN: Prospective survey of armed robbery victims. METHODS: Fifty-one consecutive armed robbery victims were assessed immediately and at 1 month post-raid for post-traumatic stress. One month post-raid, crisis support, causal attribution and coping were also measured. Thirty-one of the sample were assessed for symptoms 6 months after the raid. RESULTS: Both the main and follow-up samples had high levels of post traumatic stress symptoms immediately after the raid which reduced significantly 1 month later and, for the follow-up subsample, further still 6 months later. Higher levels of symptoms at 1 month and poor crisis support were associated with higher levels of symptoms at follow-up. CONCLUSIONS: Purposeful coping, symptom severity and social support effect post-trauma recovery. PMID- 11107496 TI - Complement deficiency. PMID- 11107497 TI - Defects in adhesion molecules. PMID- 11107498 TI - Genetics of IgA deficiency and common variable immunodeficiency. PMID- 11107499 TI - Chronic granulomatous disease and other disorders of neutrophil function. PMID- 11107500 TI - Common variable immunodeficiency. PMID- 11107503 TI - Influence of clinical and angiographic factors on development of collateral channels. AB - BACKGROUND: The degree of coronary collateralization is believed to be related to several clinical and angiographic factors. The duration and frequency of angina may be important factors in determining development of collateral channels. OBJECTIVE: To assess these factors for a consecutive series of patients suspected to have coronary artery disease. METHODS: Patients without at least one stenosis of < 50% and patients who had previously undergone bypass surgery were excluded from our study. Severity of stenosis was quantified by digital analysis, antegrade flow in terms of TIMI grade, and collaterals using the Rentrop classification. RESULTS: We reviewed 106 patients [mean age 61 years (range 35 84), 77.6% men]. Of these, 22 (21%) had presented with an acute coronary syndrome on this admission, whilst 46 patients (43%) had previously had an acute coronary syndrome. Collaterals were more likely in patients with stenoses of > 90% (Spearman correlation 0.65, P < 0.001) in patients with lower than normal TIMI flow grade (Spearman correlation 0.86, P < 0.01) and were related to regions of hypokinesis (Spearman correlation 0.35, P < 0.01). Significant collaterals were present in 14 patients (13%) despite their having TIMI grade II/III flow. Two of these patients had grade 2/3 collaterals with TIMI grade II/III antegrade flow. Degree of collateralization was not related to chronicity and frequency of symptoms, age, risk factors for atherosclerosis and nature of presentation (i.e. acute or stable symptoms). CONCLUSION: The likelihood of coronary collateralization cannot be prospectively predicted from clinical history alone, but appears to be largely a function of severity of stenosis and level of antegrade flow. A few patients develop high-grade collateral channels despite the presence of good antegrade flow. PMID- 11107504 TI - Intracoronary ultrasound measurements in women with myocardial infarction without significant coronary lesions. AB - METHODS: Morphologic characteristics of coronary arteries in eight women with myocardial infarction and angiographically normal or not significantly stenosed vessels were investigated with intracoronary ultrasound. The infarct-related vessel was assessed by three-dimensional volumetric analysis and compared with a control vessel from a noninfarcted area. RESULTS: Atherosclerosis was found in all infarct-related arteries. The plaques were predominantly soft, eccentric, poorly calcified, and with little lipid pools or none at all. Although the average area and thickness of plaques and cross-sectional narrowing of the infarct-related arteries were greater than those of control arteries, there were no pathognomonic characteristics of plaques in the infarct-related vessels. CONCLUSION: The possibility that atherosclerosis is the main etiologic factor for myocardial infarction can not be excluded even for women without an angiographically obvious coronary stenosis in the infarct-related vessels. PMID- 11107502 TI - X-linked hyper IgM syndrome. AB - X-linked hyper IgM syndrome is largely caused by defects in the CD40L (CD154). The patients have low levels of IgG and have difficulty with respiratory infections as well as opportunistic infections. The morbidity of this syndrome is high; however, early diagnosis and therapy should reduce morbidity and mortality. PMID- 11107501 TI - X-linked agammaglobulinemia. PMID- 11107505 TI - Increased coronary effects of stimulation of endothelin-B receptor in experimental hypercholesterolemia. AB - BACKGROUND: Vasoconstriction in response to endothelin-1 has been shown to be primarily related to its effects on the endothelin-A receptor. Experimental hypercholesterolemia is associated with an increase in coronary vasoconstrictor response to endothelin-1 in vivo, although the relative contributions of subtypes of endothelin receptor in this model remain unknown. OBJECTIVE: To test the hypothesis that there is an increase in coronary vasoconstriction in response to stimulation of endothelin-B receptor in hypercholesterolemia, which might be related to attenuation of activity of endothelin-derived relaxing factor. METHODS: We infused 5 ng/kg/min sarafotoxin, a specific endothelin-B receptor agonist, or 50 micrograms/kg/min NG-monomethyl-L-arginine (L-NMMA), a competitive inhibitor of nitric oxide synthase, into the left anterior descending coronary arteries of pigs before and after feeding them a cholesterol-rich diet for 10 weeks. RESULTS: There was a significant increase in serum level of cholesterol. After 10 weeks, infusion of sarafotoxin resulted in an accentuated decrease in coronary blood flow (CBF) compared with baseline (decreases by 60 +/- 7 versus 34 +/- 6%, P < 0.05). There was no significant difference between the effects on diameter of coronary arteries for the two time periods. The effect of L-NMMA on CBF was attenuated after 10 weeks (by 5 +/- 10.1 versus 45.6 +/- 4.7%, P < 0.05). Endothelin-receptor status of epicardial coronary arteries remained unchanged. Sarafotoxin and L-NMMA were co-infused at the above-mentioned doses into normolipidemic animals; the decrease in CBF in response to this co-infusion was comparable to the decrease observed with sarafotoxin alone in hypercholesterolemic animals (decreases of 67 +/- 5 versus 60 +/- 7, NS). CONCLUSIONS: The present results demonstrate that selective stimulation of the endothelin-B receptor increases coronary vasoconstriction in experimental hypercholesterolemia, primarily at the level of the microvasculature. These findings may be related to the attenuation of activity of endothelin-derived relaxing factor in this model, and support the hypothesis that endothelin-B receptor plays a role in the regulation of coronary vascular tone in pathophysiologic states. PMID- 11107506 TI - Myocardial reperfusion injury in neuronal nitric oxide synthase deficient mice. AB - BACKGROUND: A substantial amount of data suggesting that endothelial cell nitric oxide synthase (eNOS) plays a cardioprotective role in animal models of ischemia reperfusion injury has amassed. We have previously demonstrated that eNOS deficient (-/-) mice exhibit significantly larger myocardial infarcts than do wild-type mice. Few investigations have examined the neuronal form of nitric oxide synthase in the heart. The two constitutive isoforms have been demonstrated to play differing roles in studies of cerebral ischemia-reperfusion. OBJECTIVE: To characterize the role of neuronal nitric oxide synthase (nNOS) in myocardial ischemia-reperfusion injury. METHODS: Wild-type and nNOS -/- mice were subjected to 20 min of coronary artery occlusion and 120 min of reflow. RESULTS: We found no significant difference between the two groups in terms of infarct size. Microscopic cross-sections from both groups were examined for infiltration of polymorphonuclear leukocyte. Hearts of nNOS -/- mice exhibited significantly (P < 0.05) more polymorphonuclear leukocytes than did hearts of wild-type mice. CONCLUSION: Despite the fact that eNOS plays a cardioprotective role in the ischemic-reperfused myocardium, we observed no change in size of myocardial infarcts when nNOS was genetically disrupted. PMID- 11107507 TI - Detection of dissection and remodeling of atherosclerotic lesions in rabbits after balloon angioplasty by magnetic-resonance imaging. AB - OBJECTIVE: To assess the usefulness of magnetic-resonance imaging (MRI) for monitoring acute changes after angioplasty of preexisting lesions in rabbits with basal lesions similar to those observed in humans. METHODS: A combination of Fogarty balloon injury (1 week after initiation of diet) and a mildly hypercholesterolemic diet (0.2% cholesterol and 5% peanut oil) was used to promote the rapid formation of atherosclerotic lesions in 16 New Zealand white rabbits. After 5 months of the diet, angioplasty was performed on these lesions with a Gruntzig catheter in both iliac arteries and the abdominal aorta. MRI was used to monitor the initial formation of lesions after 3 and 5 months of the diet, and 2 days, 2 weeks, and 1 and 2 months after angioplasty. RESULTS: The combination of early Fogarty injury and mildly hypercholesterolemic diet induced fibroproliferative lesions similar to type Vb atherosclerotic lesions seen in humans. Angioplasty induced deep dissections at the shoulders of lesions in the majority of animals. These dissections often extended into the media. The cellular, proliferative response after angioplasty was localized and limited to sites of dissection. A significant increase in area of arterial wall was observed after angioplasty at sites of dissection without any loss of lumen. In contrast, proximal and distal to the sites of injury, there was no change in wall area but a transient reduction in lumen area. CONCLUSIONS: Comparison of MRI results with histology confirmed that changes in the wall and lumen, including small linear dissections in the lesions and arterial remodeling, are detectable by MRI. PMID- 11107509 TI - Catheter-based plasmid-mediated transfer of genes into ischemic myocardium using the pCOR plasmid. AB - BACKGROUND: Direct transfer of genes holds promise for the sustained delivery of therapeutic proteins to treat cardiovascular diseases. This can be accomplished by several approaches, including use of adenoviral vectors and naked plasmid DNA vectors. We previously demonstrated achieval of effective delivery of genes into the myocardium with a left ventricular-guided catheter-based approach using an adenoviral vector. OBJECTIVE: To evaluate the levels and duration of expression of genes induced after injection of a specific plasmid vector, using the same delivery platform as that in our previous work. METHODS: The pCOR plasmids are narrow-host-range plasmid vectors designed for nonviral gene therapy. We tested the ability of the pCOR plasmid vector to express its transgene after injection into the myocardium of pigs with chronic experimental ischemia using a catheter based transendocardial delivery system. Four animals were subjected to transendocardial injections of the luciferase reporter pCOR gene into ischemic and nonischemic zones using the Biosense intramyocardial injection catheter. Injections (1 mg per animal, 50 micrograms per injection site) were performed at 20 sites in ischemic and nonischemic zones. Measurements of luciferase activity were performed 3 and 7 days thereafter. RESULTS: We observed high levels of expression of luciferase gene in ischemic and nonischemic regions (on days 3 and 7, respectively, in ischemic zone 58,237 and 33,709 pg; in nonischemic zone 39,928 and 46,036 pg). Control noninjected samples from the left and right ventricles contained no detectable luciferase activity. CONCLUSIONS: With a catheter-based approach, the pCOR plasmid was successfully used to deliver genes into designated myocardial regions, and provides sustained expression of protein for at least 7 days, of roughly similar magnitudes in ischemic and nonischemic myocardium. PMID- 11107508 TI - Intracoronary administration of dipyridamole prior to percutaneous transluminal coronary angioplasty provides a protective effect exceeding that of ischemic preconditioning. AB - BACKGROUND: Ischemic preconditioning renders hearts more resistant to the deleterious consequences of ischemia. Adenosine is an important mediator in the induction and maintenance of ischemic preconditioning. Percutaneous transluminal coronary angioplasty (PTCA) allows the investigation of the consequences of ischemia in humans. The severity of myocardial ischemia decreases with subsequent balloon inflations during the course of PTCA. OBJECTIVE: To compare the effect of intracoronary administration of dipyridamole with the effect of consecutive balloon inflations. METHODS: We investigated 30 patients undergoing PTCA of the left anterior descending coronary artery in the setting of stable angina pectoris. Patients were randomly allocated to be administered either 0.5 mg/kg body weight dipyridamole intracoronarily or an equal amount of saline. Patients administered saline served as a control group. All patients were subjected to three consecutive balloon inflations. Severity of myocardial ischemia was assessed in terms of severity of chest pain, electrocardiographic signs of ischemia, and duration of balloon inflation tolerated. RESULTS: Patients administered dipyridamole intracoronarily tolerated significantly longer durations of balloon inflation than did patients in the control group. Severity of anginal pain and extent of electrocardiographic signs of ischemia were significantly lower after intracoronary administration of dipyridamole. The reductions in anginal pain and ST-segment shift caused by intracoronary administration of dipyridamole during the first balloon inflation were even more pronounced than the protection that was afforded by the third balloon inflation for patients in the control group. CONCLUSIONS: Intracoronary administration of dipyridamole prior to PTCA is associated with a significant gain in tolerance of ischemia. The protection afforded by intracoronary administration of dipyridamole is even more pronounced than the effect of ischemic preconditioning. PMID- 11107510 TI - Systolic hypertension in older patients: are calcium channel antagonists safe? AB - The Editor and Associate Editors are pleased that our readership will have the opportunity to benefit from a series of contributions by Dr John Rutherford addressing current issues in pharmacology and therapeutics pertinent to coronary artery disease. We are pleased also that Dr Rutherford will provide authoritative perspectives on topics of immediate interest on a regular basis. PMID- 11107511 TI - Bibliography. Current world literature. PMID- 11107512 TI - Total care of psoriasis. PMID- 11107513 TI - Epidermolytic hyperkeratosis with ichthyosis hystrix. PMID- 11107514 TI - Hemangiomas: update on classification, clinical presentation, and associated anomalies. AB - Infantile hemangiomas occur in 10% of children and are 3 times more common in female infants. The majority of hemangiomas are small, superficial tumors that require little, if any, treatment. During the last several years, new information regarding the classification, presentation, associations, and differential diagnosis of hemangioma has emerged and altered the management of these tumors. The purpose of this article is to briefly review some of these clinically relevant findings. A discussion of the pathogenesis and management of these potentially problematic tumors is beyond the scope of this article, but these topics have been addressed in several excellent reviews. PMID- 11107515 TI - Photo quiz. Drug-induced pemphigus foliaceus. PMID- 11107516 TI - Botanical briefs: dumb-cane--Dieffenbachia picta (Lodd.) Schott. PMID- 11107517 TI - A simple pediatric restraint. AB - This simple method for restraining movement makes procedures with pediatric patients much easier. The method uses a standard readily available bed sheet, is easy to learn, can be modified for use on various body areas, and makes short procedures possible with minimal nursing assistance. Procedures are made more safe by keeping children still without sedatives. Because wrapping in a bed sheet is less threatening to a young child than a standard Velcro restraint system, the entire procedure proceeds more smoothly than with commercially available restraints. Short procedures that might otherwise need to be referred out can now be performed in the office. PMID- 11107519 TI - Pterygium of the nail. PMID- 11107518 TI - Vaccine era measles in an adult. AB - Measles should be included in the differential diagnosis of patients with fever and the characteristic viral exanthem, even if a history of adequate immunization is obtained. We present the case of a 23-year-old white female who developed high fever (103 degrees F), brightly erythematous eruptions on the face, sore throat, dry cough, and myalgia 5 days after her return to the United States following a trip to Calcutta, India. The patient had extensive facial erythema from the hairline to the neck, but some areas beneath the chin were spared. Fine erythematous papules extended down the anterior neck, and white papules were seen on the buccal mucosa. The erythematous macules spread to the trunk and extremities, eventually becoming confluent and desquamating over a period of 1 week. Defervescence occurred with desquamation. Measles serology revealed the IgM antibody as positive and the IgG antibody as negative despite 2 measles, mumps, and rubella (MMR) vaccinations at ages 15 months and 7 years. Skin biopsy was consistent with viral infection. PMID- 11107520 TI - Psoriasis risk factors: role of lifestyle practices. AB - Psoriasis is a complex, multifactorial chronic skin disease. As in other chronic disorders, various lifestyle factors have been associated with its morbidity. We have pointed to the significance of the patient's lifestyle practices as they relate to psoriasis outcome. Several reports in the literature suggest that exogenous and endogenous factors, including emotional stress, alcohol use, smoking, and obesity, may have deleterious effects on the increased morbidity of psoriasis. In this study, we carried out a comprehensive evaluation to assess the effects of stress, alcohol use, smoking, obesity, and exercise on the natural history of psoriasis. PMID- 11107521 TI - Neuroimmunologic aspects of psoriasis. PMID- 11107522 TI - Cyclosporine in the treatment of psoriasis. AB - The efficacy of cyclosporine is well documented in multiple clinical trials. Patients with severe psoriasis who require systemic treatment should be considered for this therapeutic agent. The need for both a careful evaluation prior to therapy and strict monitoring of the patient cannot be over emphasized because untreated hypertension, renal dysfunction, and other disorders may lead to serious, nonreversible morbidity. Physicians are encouraged to follow the published guidelines for controlling psoriasis in their patients and lowering the risk of potential drug toxicity. PMID- 11107523 TI - Classical conditioning in the treatment of psoriasis. AB - It has been argued that the placebo effect represents a learned response. Research is suggested to address the utility of applying principles derived from classical (Pavlovian) conditioning to the design of drug treatment protocols. In the present instance, it is hypothesized that, by capitalizing on conditioned pharmacotherapeutic responses, it may be possible to reduce the cumulative amount of corticosteroid medication used in the treatment of psoriasis. PMID- 11107524 TI - Argyria following the use of dietary supplements containing colloidal silver protein. AB - The onset of argyria following the use of dietary supplements containing colloidal silver protein is presented. The patient was using a silver-containing product for cold and allergy prophylaxis. We review the past and present medicinal roles of silver and include a differential diagnosis for argyria. The hyperpigmentation of argyria is usually permanent, and it follows a sun-exposed distribution. This case report highlights the potential for toxicity following the use of dietary supplements and demonstrates the importance of physician inquiry regarding alternative medicines. Finally, we examine the limited role of the Food and Drug Administration (FDA) in regulating alternative medicines marketed as dietary supplements. PMID- 11107525 TI - Methotrexate and the photodermatitis reactivation reaction: a case report and review of the literature. AB - Methotrexate is an antimitotic and immunosuppressive agent used for the treatment of cancer, psoriasis, and rheumatologic disorders and for the termination of ectopic pregnancies. Physicians advising patients on the use of methotrexate need to be aware of its possible side effects, including photosensitivity. We present a patient who received methotrexate for the termination of an ectopic pregnancy and experienced a severe reactivation of her sunburn. The literature was reviewed on the types of photosensitivity and their relationship to methotrexate. PMID- 11107526 TI - Unusual presentation of secondary syphilis in 2 HIV-1 positive patients. AB - Due to diverse clinical and histopathological presentations, diagnosis of secondary syphilis can occasionally prove challenging. This is especially true in the setting of human immunodeficiency virus (HIV) infection. Variable clinical presentations of secondary syphilis in HIV disease may result in an incorrect diagnosis and an inappropriate treatment regimen. Similarly, the histology of secondary syphilitic lesions may show considerable variation, depending on the clinical morphology of the eruption. We report 2 cases of secondary syphilis in HIV-1-infected patients with cutaneous lesions of variable clinical presentation and an unusual lymphoid infiltrate simulating mycosis fungoides. PMID- 11107527 TI - Can granuloma annulare evolve into cutaneous sarcoidosis? AB - We report the unique case of a 50-year-old African American female with pulmonary sarcoidosis who presented with a new ichthyosiform eruption symmetrically located on the anterior shins and surrounded by red, translucent, intradermal papules. A skin biopsy of a new red papule showed features consistent with granuloma annulare (GA) with positive mucin staining, and an older hyperpigmented papule showed classic dermal noncaseating granulomas consistent with sarcoidosis. Recent reports have clearly demonstrated GA occurring in association with sarcoidosis, but this is the first report that suggests that a GA lesion may develop into a sarcoidal granuloma. We propose that GA may act as a precursor lesion to the more mature sarcoidal granuloma. This case further underscores the importance of careful clinicopathologic correlation. PMID- 11107528 TI - [Rebound of prostate specific antigen after discontinuation of antiandrogen therapy for benign prostatic hyperplasia]. AB - We compared the prostate specific antigen (PSA) levels in benign prostatic hyperplasia (BPH) patients with antiandrogen chlormadinone acetate (CMA) or allylestrenol (AE) before and during the treatment. We also investigated the serial change of PSA levels before, during and after discontinuation of antiandrogen therapy. Fifty-one BPH patients with normal PSA levels were treated with CMA or AE for 16 weeks. The mean serum PSA level significantly decreased after the treatment from 1.9 +/- 1.0 ng/ml to 1.1 +/- 0.7 ng/ml (M +/- SD). We discontinued medication with informed consent and the patients were carefully monitored for another 16 weeks. Nineteen patients were followed for 32 weeks. The mean serum PSA level decreased significantly from 2.0 +/- 1.0 ng/ml to 1.1 +/- 0.5 ng/ml (M +/- SD) and recovered approximately to the pretreatment level (1.7 +/- 1.1 ng/ml) at the end of this study. We found only one patient whose PSA was slightly elevated to a subnormal range (4.3 ng/ml) after discontinuation of therapy. The other BPH patients with normal PSA levels showed no excessive increase in PSA levels beyond the normal limit after discontinuation of antiandrogen therapy compared with the pretreatment baseline. In conclusion, BPH patients with a marked increase in PSA after discontinuation of antiandrogen therapy should be checked for prostate cancer. PMID- 11107529 TI - [Screening for prostatic diseases in one-day total health check-up by using prostate specific antigen, international prostate symptom score and QOL index]. AB - We preliminarily studied screening for prostatic diseases in one-day total health check-up by employing prostate specific antigen (PSA), international prostate symptom score (IPSS) and quality of life (QOL) index. From January 6 to March 31, 1998, a total of 390 men were included in this study, whose age ranged from 50 to 78 years with the mean of 57.5 years. The questionnaires, IPSS and QOL index, were mailed to the participants in advance. PSA (IMx: Dainapack) was measured at the end of the health check-up and the results of tests were explained on the same day. Participants who showed more than 8 points in IPSS, more than 4 points in QOL index and/or more than 4.1 ng/ml in PSA were given a referral to urologists of corresponding hospitals for further examination. A total of 116 men (29.7%) were judged to need thorough examination. Among 106 men who were referred to urologists, only 34 (32.1%) had visited the urologists by the end of July 1998. Two men (0.51% in all participants) were diagnosed with prostate cancer, 10 received some pharmacotherapy, and 2 underwent transurethral resection of prostate. The results indicate that screening for prostatic diseases in total health check-up is useful, even in an institute without staff urologists, in close association with urologists. PMID- 11107530 TI - [Retroperitoneal leiomyosarcoma found 5 cm in size: a case report]. AB - A 76-year-old woman presented with dull pain in left flank. Excretory urogram showed no function of left kidney. Retrograde pyelography, ultrasonography, computed tomography and magnetic resonance imaging demonstrated hydronephrosis of the left kidney and a mass lesion surrounding the ureter. The tumor was removed together with the upper region of the left ureter and kidney. The tumor was 5 x 3 x 2.5 cm in diameter and the pathological diagnosis was a leiomyosarcoma. Although recurrent tumor arose in the pelvis at 14 months after the operation, the patient has been in good general condition without showing any other evidence of recurrent lesion. This case may indicate the importance of early resection of retroperitoneal leiomyosarcoma. PMID- 11107531 TI - [A case of psoas cold abscess in a young tuberculosis patient]. AB - A 28-year-old man was referred to our hospital with a complaint of painful induration of right epididymis accompanied with right back pain and persistent low-grade fever. He was finally diagnosed with tuberculosis by sputum culture. Abdominal computed tomography (CT) revealed right psoas abscess and vertebral caries. He underwent a percutaneous drainage of the abscess followed by multidrug (streptomycin, pyrazinamide, refanpicin, isoniazide) combination therapy. Immediately after the drainage, symptoms began to improve with these therapies. However, four months later, abdominal CT showed a worsening of the abscess. Recently there is a stagnation in the decline of incidence of tuberculosis. It is still necessary to examine young people carefully bearing urogenital tuberculosis in mind. The pathogenesis and management of this rare condition are discussed. PMID- 11107532 TI - [A case of mesoblastic nephroma in an adult patient]. AB - We treated a rare case of adult mesoblastic nephroma. The patient was a 52-year old Japanese man with the chief complaint of intermittent gross hematuria and left lumbar pain. Abdominal ultrasonography, computed tomography, excretory urography, retrograde pyelography and angiography revealed a left renal tumor suspected to be a left pelvic tumor. A left nephroureterectomy was performed. The histologic examination showed a mesoblastic nephroma. A total of 38 adult mesoblastic nephroma cases were reviewed. PMID- 11107533 TI - [A case of ammonium urate urinary stones with anorexia nervosa]. AB - A 27-year-old woman had been suffering from bulimia and habitual vomiting for about 7 years and was incidentally found to have right renal stones by computed tomography. She was referred to our hospital for the treatment of these caluculi. On admission, she presented with hypokalemia, hypochloremia and metabolic alkalosis and was diagnosed with anorexia nervosa. Following successful removal by percutaneous nephrolithotripsy and extracorporeal shockwave lithotripsy the stones were found to consist of pure ammonium urate. Since the urine of an anorexia nervosa patient tends to be rich in uric acid and ammonium, anorexia nervosa seems to be associated with ammonium urate urinary stones. PMID- 11107534 TI - [Transitional cell carcinoma of urachus: a case report]. AB - We report a case of transitional cell carcinoma of urachus in a 72-year-old man. At follow up cystoscopy for past history of bladder cancer, we found a papillary tumor in the right orifice that came out to the bladder cavity intermittently. Although there was no cancerous lesion on the surface of the bladder mucosa, a submucosal eminence at the dome of bladder was observed. Sagittal magnetic resonance imaging (MRI) revealed an extravesical tumor (2 cm) at the position of urachus. Under the diagnosis of right ureteral cancer and urachal cancer, we performed right distal ureterectomy, ureteral reimplantation and total resection of urachus. Pathological examination revealed transitional cell carcinoma in the urachus and right ureter. The urachal cavity was isolated completely from the bladder cavity. Tumor infiltrated to the muscularis of the bladder dome from the urachal cavity, but there was no cancerous lesion on the surface of the bladder mucosa. Therefore, our diagnosis was urachal transitional cell carcinoma and right ureteral carcinoma. PMID- 11107535 TI - [A case of urachal abscess mimicking a tumor in the retrovesical space]. AB - A 14-year-old girl was admitted to our hospital because of umbilical erythema and discharge. She had had an appendectomy at the age of twelve. Abdominal ultrasonography and cystoscopy revealed a large tumor-like mass at the posterior wall of the bladder. Computed tomography and magnetic resonance imaging revealed urachal sinus. The diagnosis of urachal abscess had been confirmed and conservative treatment had been continued by drainage via umbilicus and the administration of antibiotics. Total excision of the urachus was performed about one month later because the bladder mass was not reduced. Pathological findings revealed an inflammatory thickened wall of the urachus and no evidence of malignancy. We report this rare case of urachal abscess with a large mass in the retrovesical space. PMID- 11107536 TI - [Urethral diverticula surrounding the urethra in women: report of 2 cases]. AB - We report two cases of female urethral diverticula. A 49-year-old woman (case 1) complained of perineal pain when she voided urine. A 36-year-old woman (case 2) complained of perineal pain. In both cases, intravenous pyerography and urethrography revealed diverticula around their urethra, and we diagnosed them with urethral diverticula which surrounded their urethra by magnetic resonance imaging. We treated them by transvaginal diverticulectomy. Case 2 was successfully treated, but the diverticulum recurred after one year in case 1. There are over 200 reported cases of female diverticula in the Japanese literature, but a urethral diverticulum surrounding the urethra is rare. PMID- 11107537 TI - [Urological procedures for progressive renal dysfunction due to polycystic kidney disease]. AB - Urological procedures for progressive renal dysfunction due to polycystic kidney disease (PKD) such as percutaneous puncture of renal cysts are merely symptomatic treatments and have little effect on renal function. At present, the two most effective methods of preventing renal dysfunction are blood pressure management and dietetic therapy, which are more effective with early initiation. Moreover, as PKD is an autosomal dominant disease, there is a high risk that family members of the patient may have asymptomatic PKD. It is essential to identify and treat such potential patients at an early stage in order to prevent progressive renal dysfunction. In place of the traditional nephrectomy, we attempted transcutaneous renal arterial embolization (TRAE) for hemorrhage into renal cysts, hematuria and obstruction of intestine due to proliferation of cysts after the introduction of hemodialysis. When TRAE was carried out on one kidney, the cysts in the other kidney proliferated and even though the renal arteries were completely embolized, it required 5 to 6 weeks for the kidney to contract. Our conclusions are TRAE is effective with no adverse reactions for PKD. These results suggest that in the future TRAE may become the preferred treatment for PKD in place of nephrectomy. PMID- 11107538 TI - [Acquired renal cystic disease]. AB - In 1977, Dunnill et al. described a new disorder, bilateral multiple renal cystic disease. It occurred among hemodialysis patients whose original illness had not been cyst-related. Acquired cystic disease of the kidney (ACDK) is commonly observed in patients undergoing hemodialysis. The incidence of ACDK is 40-50% in reports of autopsy and surgical specimens, rising to more than 90% after 5-10 years of dialysis. The volume of the kidneys decreases in the first 3 years of dialysis and then increases as the rate of cyst formation increases. In male patients undergoing long-term hemodialysis the incidence of ACDK is markedly high. ACDK is also found in patients before hemodialysis. The primary concern in patients with ACDK is the increased incidence (5-19%) of renal cell carcinoma (RCC). The incidence is about twelve to eighteen times higher than that in the general population and the cancers may be asymptomatic. Therefore, screening is essential if carcinomas are to be detected early. Regular screening by ultrasonic examination or CT scan is needed. A patient requires nephrectomy when the kidney cancer exists or is suspected by dynamic CT scan. Nephrectomy is performed only on the side with renal mass. It has been argued that RCC associated with ACDK are innocuous and do not predispose the patient to an increased risk of death from RCC. RCC arising from ACDK is considered to be a tumor of low malignant potential, compared with classic RCC. However, RCC has been reported to metastasize in 16% of the patients on dialysis and to be the cause of death in 2% of the kidney transplant recipients. The etiology of ACDK is unclear and its incidence increases with the duration of dialysis. ACDK patients have a propensity to develop adenocarcinoma. The increased incidence of RCC in ACDK patients warrants careful radiologic monitoring of end-stage kidneys in selected patients. PMID- 11107539 TI - [Renovascular hypertension]. AB - Renovascular disease is one of the most common causes of secondary hypertension. Recent technical advances have changed the management principles, which include a more aggressive approach to the diagnosis and treatment of renovascular hypertension (RVH). We experienced a total of 95 cases with RVH between 1958 and 1999. The mean age of all patients was 31.8 years old, ranging from 3 to 64 years. The three major basal diseases that caused RVH were fibromuscular dysplasia (34/95), arteriosclerosis (26/95), and aortitis (12/95). Ninety-two kidneys were treated in 79 of the 95 patients. The major therapeutic modalities performed were reconstruction of renal artery (6/79), nephrectomy (21/79), autotransplantation (26/79), and percutaneous transluminal angioplasty (PTA) (25/79). PTA is now the treatment of choice for the initial management of patients with RVH. Surgical treatment is generally reserved for patients in whom PTA fails. Pharmacotherapy is used on patients awaiting angioplasty or revascularization, those who are too ill for intervention, and those who have failed to respond to intervention. PMID- 11107540 TI - [Long-term follow-up of children with surgically treated primary vesicouretral reflux]. AB - We reviewed the symptoms of hypertension and proteinuria and evaluated the renal function in patients aged 15 and over with surgically treated primary vesicoureteral reflux. Twenty-two patients were enrolled in this study. There were no patients with hypertension except one patient with proteinuria. The extraction factor by Heyman's method in these patients tended to increase with age at operation and decrease with the grade of reflux at operation. There findings suggest that long-term follow-up is particularly essential to patients who had severe reflux and underwent anti-reflux surgery in the early stage of life. PMID- 11107541 TI - [The preoperative function of the affected kidney and the outcome of endopyelotomy for ureteropelvic junction obstruction]. AB - Between July 1988 and February 1998, we treated 132 patients with 133 procedures of endopyeloureterotomy via the transpelvic extraureteral approach and followed them up for more than one year. Of 9 unsuccessful cases, 4 were confirmed to have the patency of the stenotic segment on retrograde pyeloureterograms. We compared these 4 cases with another 4 cases which were subsequently treated successfully despite poor renal function. Pre- and post-operative changes in renal function in 8 cases with unilateral hydronephrosis due to ureteropelvic junction obstruction were studied based on 99mTc-DMSA renal uptake rate and creatinine clearance. In successful cases, DMSA renal uptake rates compared with the healthy side were 9% to 59%. Unilateral creatinine clearance was 6 to 34 ml/min. All cases were improved slightly in renal function. In unsuccessful cases, DMSA the renal uptake rate was 2% to 89% and creatinine clearance 10 to 16 ml/min. No improvement in renal function was noted. Patients with differential renal function of less than 20 ml/min would thus appear to have little chance of postoperative improvement of differential function. PMID- 11107542 TI - [Genes and environment]. AB - Many quantitative characters depend on the action of a large number of genes and environmental factors. The mode of inheritance of these characters is polygenic. The phenotypic variance of the character is the sum of the components, thus the genetic and the environmental variances (VP = VG + VE). The degree of genetic determination VG/VP and VE/VP are difficult to estimate in man. The heritability a related coefficient to VG/VP can be estimated from the degree of ressemblance between relatives. The heritability is the additive genetique variance as a proportion of the phenotypic variance. Polygenic threshold inheritance can account for the familial non mendelian distribution of multifactorial diseases. PMID- 11107543 TI - [Pharmacogenetics or "farm-ecogenetics"?]. AB - Pharmacogenetics and ecogenetics constitute the study of the contribution of genetic factors in interindividual variations in drugs and xenobiotics metabolism respectively and also its physiopathology (toxicity, therapeutic failure and pejorative effect). However, the frontiers between these two fields are interpenetrant with no defined limits. Among the enzymes involved in transformation of drugs/xenobiotics, the cytochrome P450 superfamily plays a major role. Most of the "poor metabolisers" either homozygotes or compound heterozygotes for mutant cytochrome P450 alleles have no detectable enzyme activity. Although phenotyping using a marker drug is usefull, it suffers from several constraints and is influenced by several environmental factors and physiopathological state of the individual. To overcome not only these difficulties but also to improve the overall predictiveness of a drug biotransformation, genotype technology offers an advantage and pharmaco-economic technology further expands the horizon. These fast moving fields must, in near future, allow tailoring of drug prescription on a personalized basis and should, in principle, help in the design and development of novel drug molecules. PMID- 11107544 TI - [Genetic predisposition to bilharziasis in humans: research methods and application to the study of Schistosoma mansoni infection]. AB - The development of genetic epidemiology methods using recent human genetic mapping information together with the growing availability of candidate genes has led to major advances in the identification of host genes in human schistosomiasis. Two phenotypes have been studied so far in the infection by Schistosoma mansoni: infection levels by the parasite as measured by the faecal egg counts, and the severe hepatic fibrosis caused by S. mansoni assessed by ultrasound examination. The first study was performed on Brazilian pedigrees and provided strong evidence for a major gene controlling infection levels by S. mansoni denoted as SM1 which was mapped to chromosome 5q31-q33. This region contains several candidate genes involved in the regulation of the Th1/Th2 response, and the direct role of polymorphisms located within these genes is under investigation. The second study conducted in Sudan also showed the presence of a major gene influencing the development of severe hepatic fibrosis due to S. mansoni infection denoted as SM2. This gene is not located in the 5q31-q33 region, but maps to chromosome 6q22-q23 and is closely linked to the IFN-gamma R1 gene encoding the receptor of the strongly anti-fibrogenic cytokine Interferon gamma. These findings indicate that two distinct genetic loci control human predisposition to schistosomiasis, SM1 located in the 5q31-q33 region which is likely to play a role in the Th1/Th2 differentiation, and SM2 in 6q22-q23 influencing disease progression with a possible involvement in the regulation of IFN-gamma. PMID- 11107545 TI - [Selective pressure in host-parasite systems]. AB - Selective pressures in host-parasite systems are the result of a continuous conflict between the divergent interests of each partner, on the long run. Whereas the fitness (lifetime reproductive success) of parasites is usually increased by a higher frequency of encounters with susceptible hosts and a better survival rate after infection, the fitness of hosts is increased by opposite processes, avoidance of encounters with infective stages and destruction of the parasites. These selective processes, often referred to as coevolution or arms races are in agreement with the Red Queen hypothesis of Van Valen, which assumes indefinite adaptive changes in both partners, in order to set up counter-measures against the weapons of "the other". Arms races in host-parasite systems thus suggest a gradualistic evolution, but this does not contradict the present day ideas on the tempo changes in the course of evolution (punctuated equilibria). Numerous factors, either genetic (evolutionary lag...), environmental (nutritional status...) or cultural (prevention, vaccination, therapy...) influence the severity of infections at an individual scale. The "terrain", which is a component of the individual phenotype, is thus at the cross-roads of genes, environment and culture. Humans must count more on their intelligence than on natural selection to prevent and cure infectious and parasitic diseases. PMID- 11107546 TI - [Mendelian predisposition to mycobacterial infections in humans]. AB - Selective susceptibility to poorly pathogenic mycobacteria, such as bacille Calmette-Guerin (BCG) vaccine and environmental non-tuberculous mycobacteria (NTM), bas long been suspected to be a mendelian disorder but its molecular basis has remained elusive. Recently, recessive mutations in the interferon gamma receptor ligand-binding chain (IFN gamma R1), interferon gamma receptor signalling chain (IFN gamma R2), interleukin 12 p40 subunit (IL-12 p40), and interleukin 12 receptor beta 1 chain (IL-12R beta 1) genes have been identified in a number of patients with disseminated BCG or NTM infection. Although genetically distinct, these conditions are immunologically related and highlight the essential role of interferon gamma-mediated immunity in the control of mycobacteria in man. PMID- 11107548 TI - [Genetics of asthma and atopy]. AB - The genetics of asthma and atopy has made tremendous progresses over the last couple of years. It has been known for ages that allergies and asthma are concentrating within families. Modern genetics has pinpointed some gene areas such as 5q31.1, 6p21.3, 11q13, 12q15, 14q11.1 and 16p12. Much of the information available today has been fragmentary and not always confirmed. Alleles at multiple loci are likely to be involved and the ultimate picture is most likely determined by many genetic and environmental factors. PMID- 11107549 TI - [Contribution of genetics to the concept of risk status for alcohol dependence]. AB - The factors involved in the variability of one's own risk for alcohol dependence is multifactorial and mostly unknown. Nevertheless, genetic factors are clearly involved in the risk for the disorder, the impact of addictive genetic factors being evaluated between 30% and 50%. Aggregation studies underline the difficulties of delimiting the boundaries of the phenotype, as some subgroups of alcohol-dependent patients have genetic factors with an increased weight (severe and young-onset disorder, presence of antisocial behavior...). Furthermore, familial studies also showed that the genetic of "addiction" may be more relevant than studying the genetic of one specific dependence. The use of one substance lately abused being probably more dependent of familial and/or environmental features. The discover of susceptibility genes had a greater impact on defining the phenotype than proposing new treatments. Three examples are given in this review. Firstly, le gene coding for the second dopamine receptor may be more specifically involved in severe and comorbid alcohol-dependence. Secondly, the gene coding for the serotonin transporter may increase the suicidal risk in alcohol-dependent patients. Thirdly, the quality of the withdrawal process is partly explained by the existence of a specific genotype of the dopamine transport gene. PMID- 11107547 TI - [Prevention of renal carcinoma: the nutri-genetic approach]. AB - The development of renal cell carcinoma (RCC) has been associated with both genetic and environmental factors, with somatic and germline mutations in the von Hippel-Lindau (VHL) tumor suppressor gene and with tobacco smoking, obesity, long term exposure to some nutrients, pollutants, and industrial solvents such as trichloroethylene. Intra and interfamilial variability of expression of germline mutations in the VHL gene and variable susceptibility to carcinogens in the sporadic forms strongly suggest the involvement of conditional modifier genes. In order to identify sub groups of individuals at increased risk because of susceptibility genotypes, we have collected a series of 460 patients who developed an RCC and 79 families with the von Hippel Lindau disease. To collect clinical and mutational data for correlation analysis we have developed a unique tool the Universal Mutation Database. Comparison of the spectrum of germline and somatic mutations in the VHL gene showed that: 1) in sporadic RCC mutations lead more often to truncated proteins (83%), while the remaining mutations (17%), include 3/4 of transversions and 1/4 of transitions. This high proportion of transversions supports the involvement of carcinogens the impact of which is conditioned by the genetic variability of xenobiotic metabolizing enzymes; 2) whereas in familial cases missense mutations are more common; this difference allowed us to define a prognostic factor for the occurrence of RCC in a VHL context. In order to look for genotypes conferring a higher risk we genotyped the RCC patients for 8 different genes (50 genotypes). A significant relationship was observed for several combinations of alleles including CYP1A1 ("variant"), NAT2 and NAT1 (slow) and GSTM1 (null allele). Associations between specific mutational profiles and at risk genotypes at different tumoral stages should allow us to: 1) define more precisely the nature of specific patterns of mutations in relation with the deficiency or overexpression of such or such enzymes in presence of particular carcinogens; 2) demonstrate that certain combinations of genotypes confer a particular risk to develop a specific type of tumor in VHL patients. Thus tracking of potentially carcinogenic substances, through their footprints and through identification of conditionally detrimental genotypes of genes participating in their detoxification should permit a better prevention through an appropriate nutrition adapted to each individual. PMID- 11107550 TI - [Clinical investigation of inhomogeneity of washout rate in dipyridamole stress thallium scintigraphy: implication of new parameter for the indication of coronary intervention]. AB - OBJECTIVES: The standard deviation of the washout rate in dipyridamole stress thallium-201 myocardial single photon emission computed tomography (SPECT) was correlated with the severity of coronary artery lesions and the viability of ischemic myocardium to provide a new quantitative parameter for judging indications for coronary intervention therapy. METHODS: Dipyridamole stress thallium-201 SPECT was performed in 233 patients for differential diagnosis of angina pectoris during the 40 months beginning in October 1995, and 57 patients were investigated who underwent coronary angiography within 6 months of the SPECT. The washout rate standard deviation (WRSD) in 720 fractions in the bull's eye view of the SPECT was determined. The conventional washout rate extent score (WRES) and washout rate severity score (WRSS) on the washout rate map were also determined. Based on the coronary angiography findings, patients were divided into 3 groups: zero-vessel group (zero-vessel disease, n = 20), one-vessel group (one-vessel disease, n = 18), and multivessel group (two- or three-vessel disease, n = 19). The patients were also divided into 2 other groups: Int group (n = 21), who underwent coronary intervention therapy, and Med group (n = 36), in whom intervention therapy was not indicated. RESULTS: All 3 parameters, WRSD, WRES and WRSS, showed significant differences between the 3 groups, and more coronary arteries affected by coronary artery stenosis were associated with higher WRSD (zero-vessel group: 5.4 +/- 1.5, one-vessel group: 7.0 +/- 3.7, multivessel group 11.4 +/- 6.7; p < 0.001), WRES (3.3 +/- 5.0, 15.5 +/- 18.1, 23.0 +/- 25.4; p < 0.01), and WRSS (1.4 +/- 2.8, 25.4 +/- 40.2, 84.8 +/- 114.5; p < 0.01). WRSD (Med group: 5.9 +/- 2.7, Int group: 11.3 +/- 6.4; p < 0.001), WRES (7.3 +/- 12.0, 24.7 +/- 24.9; p < 0.01), and WRSS (9.9 +/- 29.3, 82.9 +/- 108.2; p < 0.01) were all significantly higher in the Int group compared with the Med group. There were significant correlations between Gensini's score and WRSD (r = 0.51, p = 0.00005), WRES (r = 0.37, p = 0.005), and WRSS (r = 0.29, p = 0.03). The cutoff values for the indications for coronary intervention therapy were established for each of the 3 parameters as the maximum value of average sensitivity and specificity as follows: WRSD > 9.0 (sensitivity 0.62, specificity 0.89, positive predictive value 0.76, negative predictive value 0.24); WRES > 10.0 (0.62, 0.69, 0.54, 0.46, respectively); WRSS > 13.0 (0.62, 0.83, 0.68, 0.32, respectively). WRSD > 9.0 had the highest specificity and positive predictive value for judging indications. CONCLUSIONS: A new quantitative parameter, WRSD > 9.0, suggests the presence of viable and curable ischemic myocardium as an indication for coronary intervention therapy. PMID- 11107551 TI - [Diagnostic accuracy of carotid ultrasonography in screening for coronary artery disease]. AB - OBJECTIVES: This study investigated the diagnostic accuracy of carotid ultrasonography in screening for significant coronary artery disease (% diameter stenosis > or = 75%). METHODS: Five hundred sixty patients (342 males, 218 females, mean age 66.4 years) underwent both coronary angiography and carotid ultrasonography. Gensini's coronary score was calculated as a quantitative parameter of coronary atherosclerosis. The most hypertrophic intimal-medial complex thickness (IMT) of the bilateral common carotid arteries (distal and proximal to the echo probe in each artery) was measured within 2 or 3 cm from the carotid bifurcation. The mean IMT (mean of these 4 sites), the maximum IMT (maximum of these 4 sites), and number of plaques (localized hypertrophy of IMT > or = 1.1 mm) were calculated as a quantitative parameter of carotid atherosclerosis. RESULTS: The screening parameters were determined as 0.85 mm mean IMT, 1.1 mm maximum IMT, and at least 2 sites of plaque. The sensitivity, specificity and accuracy rate for the detection of coronary artery disease were 57.3%, 61.6% and 59.6% for mean IMT, 43.5%, 71.1% and 58.6% for maximum IMT, and 60.8%, 70.5% and 66.1% for number of plaques. Furthermore, the overall results (except maximum IMT) were 73.3%, 49.2% and 60.2%. CONCLUSIONS: These results suggest that carotid ultrasonography is useful as a non-invasive and easy screening method for coronary artery disease. Furthermore, carotid ultrasonography will allow routine observations to follow the progression of coronary atherosclerosis. PMID- 11107552 TI - Cardiac troponin I level correlates with chronic-phase left ventricular function after successful direct percutaneous transluminal coronary angioplasty in patients with acute myocardial infarction. AB - OBJECTIVES: The relationships between cardiac troponin I, various biochemical markers, and chronic-phase left ventricular ejection fraction (LVEF) after successful direct percutaneous transluminal coronary angioplasty (PTCA) were examined in 36 patients with acute myocardial infarction. METHODS: Biochemical markers were measured on admission, immediately after, and from 6 hours to 9 days after PTCA. RESULTS: The time to peak values were: creatine kinase-MB 9.7 hours, cardiac troponin I 9.8 hours, myoglobin 10.7 hours, creatine kinase 10.6 hours, cardiac troponin T 18.6 hours, and myosin light chain 68.9 hours. Cardiac troponin T, cardiac troponin I and myosin light chain levels were elevated over 9 days after successful direct PTCA. Chronic-phase LVEF inversely correlated with peak values of creatine kinase-MB (r = -0.519, p < 0.01), cardiac troponin T (r = -0.500, p < 0.01), cardiac troponin I (r = -0.441, p < 0.05) and creatine kinase (r = -0.411, p < 0.05). The values of cardiac troponin I, cardiac troponin T, creatine kinase and creatine kinase-MB at each sampling point were significantly inversely related to chronic-phase LVEF. The value of cardiac troponin I at each time point for 7 days correlated well with chronic-phase LVEF. CONCLUSIONS: Cardiac troponin I has high specificity for predicting long-term cardiac function after successful direct PTCA when early values are unavailable. PMID- 11107553 TI - [Clinicopathologic study of calcified aortic valve stenosis in the aged, and evaluation of the indications for percutaneous aortic balloon valvuloplasty]. AB - OBJECTIVES: We performed clinicopathologic study of 56 aged patients with calcified aortic valve stenosis and investigated the indications for percutaneous aortic balloon valvuloplasty. METHODS: The patients were 24 men and 32 women with a mean age of 81.9 years, who were classified into the following 3 types by etiology: 33 patients (58.9%) had senile aortic stenosis, 10 patients (17.9%) had bicuspid aortic stenosis, and 13 patients (23.2%) had rheumatic aortic stenosis. The sites of calcification were divided into the following 3 categories: cusp bases (base type), free edges (edge type), and both bases and edges (mixed type). RESULTS: Among the 33 patients with senile aortic stenosis, 10 (30.3%) had calcification of base type, 2 (6.1%) of edge type and 21 (63.6%) of mixed type. Among the 10 patients with bicuspid aortic stenosis, one (10%) had calcification of base type and 9 (90%) of mixed type. Among the 13 patients with rheumatic aortic stenosis, 3 (23.1%) had calcification of edge type and 10 (76.9%) of mixed type. In addition, 2 or 3 commissures were fused in patients with rheumatic aortic stenosis. The cusps of the aortic valves in the bicuspid type were the most severely thickened among the 3 groups. Soft X-ray imaging showed the aortic valves of rheumatic aortic stenosis were the most severely calcified (calcification score: 2.4), followed by those of bicuspid aortic stenosis (1.9) and senile aortic stenosis (1.6). CONCLUSIONS: Percutaneous aortic balloon valvuloplasty is most suitable in patients with calcified senile aortic stenosis because of the milder calcification, compared with those of the other 2 types. PMID- 11107554 TI - [Novel extracorporeal granulocyte apheresis column inhibits myocardial reperfusion injury: future clinical applications]. AB - OBJECTIVES: Leukocyte infiltration is very important in myocardial ischemia/reperfusion injury. A new extracorporeal circulation therapy using a granulocyte-apheresis column (G-1) has recently been proved to be effective and safe in patients with rheumatoid arthritis and with ulcerative colitis. This study investigated whether this therapy would reduce myocardial ischemia/reperfusion injury. METHODS: Rabbits were subjected to 30 min ischemia followed by 180 min reperfusion with no treatment (control group: n = 12); or treated with extracorporeal blood circulation (2 ml/min) passing through the G-1 (G-1 group: n = 12), or sham column (sham group: n = 12). Infarct size was determined by tetrazolium staining and expressed as a percentage of the area at risk. RESULTS: Infarct size in the G-1 group (15.9 +/- 3.4%) was significantly (p < 0.01) smaller than in the control (34.7 +/- 3.9%) and sham (35.2 +/- 4.1%) groups. Granulocyte emigration into the ischemic myocardium assessed by histopathological score was significantly (p < 0.01) less in the G-1 group (0.58 +/- 0.17) than in the sham group (1.42 +/- 0.26). There was a significant (r = 0.78, p < 0.0001) positive correlation between this score and infarct size. Flow cytometry showed alterations in the adhesion molecule expression on granulocytes during the passage through the G-1, but not sham, columns as a low L-selectin and high Mac-1 form, which is associated with the inability to home to the inflamed site. CONCLUSIONS: Extracorporeal circulation with the G-1 column, a newly developed clinically applicable therapy, appears to reduce infarct size at least in part by limiting the emigration capacity of granulocytes into the ischemic myocardium. PMID- 11107555 TI - [Percutaneous transluminal septal myocardial ablation combined with permanent dual-chamber pacing in hypertrophic obstructive cardiomyopathy: a case report]. AB - A 74-year-old woman suffering from dyspnea on effort and chest pain (New York Heart Association functional class III) was admitted to our hospital. Echocardiography and left ventriculography showed severe hypertrophy of the left ventricular wall and a severe intraventricular pressure gradient of about 150 mmHg. The diagnosis was hypertrophic obstructive cardiomyopathy. The intraventricular pressure gradient was decreased from 157 to 68 mmHg by percutaneous transluminal septal myocardial ablation (PTSMA), but this reduction was not adequate. Thereafter, dual-chamber pacing was additionally performed. The intraventricular pressure gradient was decreased to 26 mmHg with no complication. PTSMA is a certain way to decrease pressure gradient, but has some complications. Permanent dual-chamber pacing also can show the same effect, but is inferior to PTSMA. Combination of these 2 methods succeeded in abolishing the intraventricular pressure gradient with no complication. PMID- 11107556 TI - Massive deep vein thrombosis after cesarean section treated with a temporary inferior vena cava filter: a case report. AB - A 25-year-old woman suffered a massive deep vein thrombus in her left common iliac vein extending to the inferior vena cava after an abdominal cesarean section. The massive and floating inferior vena cava thrombus was considered to pose a high risk of pulmonary thromboembolism. After placement of a temporary inferior vena cava filter via the left brachial vein, thrombolytic therapy and anticoagulation therapy were instituted. The filter successfully prevented pulmonary thromboembolism during thrombolytic therapy. This patient was confined to bed because the filter moved vertically with left shoulder joint abduction. Although a temporary inferior vena cava filter is very useful for the prevention of pulmonary thromboembolism in a patient with deep vein thrombus, the appropriate range of activity for such a patient needs careful consideration. PMID- 11107557 TI - [Impaired cardiac function in a 61-year-old woman with coarse facial features, enlarged hands and feet]. PMID- 11107558 TI - [Serial observation for 6 years of "so-called" complex lesion in a patient with mild coronary stenosis]. PMID- 11107559 TI - [Achievements, disappointments and hopes in neurological therapy in the 20th century]. AB - Only in the second half of the 20th century a breakthrough occurred in the traditional neurological therapeutic methods based up to that time mainly on bromine with valerian extract and vitamins B. Later on in that century several great discoveries were made which improved greatly the effectiveness of the neurological therapy: psychopharmacology which began with the introduction of chlorpromazine and reserpine, the use of corticosteroids for which the Nobel award was given, levodopa introduction for Parkinson's disease, non-steroid antiinflammatory agents and the demonstration of their action mechanism /also Nobel award/, immunotherapy, botulin toxin for the treatment of dystonias and thrombolytic drugs possibly the drugs of the future. The main disappointment is the broad chasm between the progress made in diagnostic methods and the low effectiveness of therapy in strokes, amyotrophic lateral sclerosis, Alzheimer's disease and other degenerative neurological diseases. Many problems arose with the introduction of levodopa changing the course and clinical pattern of Parkinson's disease. The problem of our times are the adverse effects of pharmacotherapy. The low effectiveness of the new drugs used in epilepsy is also disappointing. A hope for the future is the new direction in therapy--the use of genes and also the use of monoclonal antibodies and neurotrophic agents. It is to be expected that in the near future medicine will find effective methods for the treatment of malignant neoplasms. PMID- 11107560 TI - [Pain--physiological or medical phenomenon]. AB - The word "pain" is ambiguous, symbolic and it is understood differently by the patient and by the doctor or physiologist. It describes a kind of sensation evoked by harmful stimuli which is a physiological phenomenon indispensable for protection and also suffering caused by injury or disease. The second component of the phenomenon is usually not recognized--this is the reaction to pain. The primary component is centripetal and ascending to brain centers, the second is centrifugal, descending and they both form the reflex arc. This is the pain or nociceptive reflex. Commonly, when we speak of pain we mean only the centripetal part of the reflex which cannot be objectively assessed in medical practise. This part is blocked by anaesthesia before surgical procedures. The second part is the subject of a separate consideration which begins with psychic reaction to pain and pain tolerance, and suffering depend on it. The motor reaction to pain is more spectacular and possible for recording. Three types of this reaction are discerned: flight, defense and suffering expression. Nociceptive sensation is a physiological receptor-mediated sensation, while pathological pain can derive from receptors as well as from nerves /conduction pain, neuropathic differentiation pain/ or from centres/central pain. The pathological pain has clinical features making possible the recognition of its origin, its mechanism for undertaking of appropriate measures. Neuropathic pain is the one most difficult to treat. The receptor-mediated pain continues as long as the stimulus is active, the neuropathic pain is longer lasting. PMID- 11107561 TI - [Selected problems for discussion of procedures in epilepsy]. AB - In neurological practice epilepsy treatment has a special place. Among the problems encountered by us in this connection every day the following are worth mentioning: Differential diagnosis of epileptic seizure against psychogenic fits and fainting. Management of the first seizure /treatment starting?/. How EEG tracings should be related to clinical status? Should imaging examination be suggested? Treatment programme /doses, mono- or polytherapy, new generation drugs/ Can anticonvulsants worsen epilepsy? Indications to surgical treatment. Whether and when anticonvulsants should be withdrawn in case of remission? Should treatment be given prophylactically after injury or stroke? How to deal with late onset epilepsy? What is the significance of concomitant diseases? The purpose of this lecture is to incite personal consideration of these matters and discussion on these problems in which it is difficult to take optimal decision. PMID- 11107562 TI - [Drug rebound headaches]. AB - The term "drug rebound headache" refers to headaches occurring every day in patients with migraine and tension headaches as a consequence of taking analgesics or ergotamine every day. Their cause is a vicious circle mechanism. This form of headache has been discovered in recent years and it is supposed that about 20% of patients with chronic headaches belong to that category. The only way to disrupt this vicious circle is immediate complete abandoning of these drugs. The abstinence period with associated troublesome symptoms lasts several to up to 20 days. Antidepressants are given for their alleviation. The author prescribes opipramol /Pramolan/ which is taken by increments from half tablet up to three in 6 days and the treatment is then continued for 4-6 weeks. From the 6th day on the patient should completely discontinue taking of analgesics. The material observed by the author comprises 47 patients /45 women/ aged 19-57 years, mean age 41 years, with these headaches continuing since 1-12 years /mean 3.0 years/. The above described method gave good results in 32 cases. In 10 cases complete withdrawal of analgesics was not possible by this method was abandoned accepting that their headaches were not due to drug abuse. PMID- 11107563 TI - [Emergency treatment of migraine attacks with particular reference to agonists of 5-HT1B/1D receptor]. AB - The author discusses the modern methods of emergency controlling of migraine attacks, especially the recently introduced drugs agonists of the 5-HT 1B/1D receptor /Sumatriptan, Zolmitriptan/ and prophylactic treatment. In light and moderately severe attacks analgesics are usually effective as well as non-steroid antiinflammatory drugs /paracetamol, acetylsalicylic acid, naproxen, diclofenac, ibuprofen, ketoprofen etc./ with or without addition of caffeine and codeine, and metoclopramide /for suppression of nausea and vomiting/ or Torecan. Ergotamine is still in use, although it can produce serious adverse effects. An essential advance in the treatment of more severe attacks was achieved with the introduction of Sumatriptan, a selective 5-HT1D receptor agonist in 1988. In pursuing this direction further indole derivatives /so called triptans/ were introduced. Zolmitriptan introduced in 1994 is an agonists of 5-HT 1B/1D receptor, is active both peripherally and centrally, is well absorbed from the digestive tract and has a good bioavailability index /40%/. In 2.5 mg doses it causes regression or marked alleviation of migraine attack within 2 hours in 70% of cases. Administration of a second dose after that time increases the percentage of successes. Adverse effects are usually mild, coronary complication have not yet been described. Prophylactic treatment is given to patients with attack frequency over 2 in a month. For that treatment usually dihydroergotamine, pisotifen, propranolol, metoprolol, flunarizin, valproic acid, iprasochrom and oxetoron are given. PMID- 11107564 TI - [Insomnia and its treatment]. AB - Insomnia is a very frequent complaint /periodically or permanently it affects about 35% of the population/ and is a serious sociological problem. This term covers at least four types of sleep disorders: difficult falling asleep, frequent awakening, too early awakening and impairment of sleep quality--sleep quantitatively sufficient but failing to produce a feeling of rest. Insomnia may be sporadic, short-lasting and chronic. The last type requires particularly medical assistance since impairment of sleep quality can lead to drug dependence. In every case of insomnia it should be tried to explain its cause /neurosis, depression, somatic diseases with symptoms leading to sleep disturbances, toxic factors such as alcohol, drugs, inappropriate sleep hygiene etc./. In the treatment the basic role is played by removal of causes and better observation of sleep hygiene. Hypnotic drugs are indicated in sporadic and short-lasting insomnia, but in chronic insomnia they should be used cautiously and not continuously. Barbiturates have been abandoned recently and benzodiazepines have replaced them. They are, however, fraught with numerous faults. Cyclopyrolones /Zolpidem, Zopiklon/ are the new generation of hypnotic drugs in which the negative features of benzodiazepines have been partly excluded. Their half-life is short, they cause no rebound effect, adverse effects are better tolerated and are less frequent, drug dependence is not produced. PMID- 11107565 TI - [Multiple sclerosis--certain clinical and diagnostic problems]. AB - The diagnosis of MS is based exclusively on clinical examination disclosing at least two focal lesions, repeatedly occurring recurrences and ruling out of other causes. This is the generally accepted principle based on the diagnostic criteria of Poser. The presence of the lesions can be demonstrated in clinical examinations by magnetic resonance imaging /MRI/ or the study of evoked potentials /EP/ which detect these changes in about 90% of cases of clinically certain MS. In 10% of cases the results are negative--but can this exclude MS? It happens also that MRI or EP carried out for another reason demonstrate the presence of such lesions--is it MS in such cases? The diagnosis of SM is supported by age below 40 years and the presence of at least four focal lesions in the white matter of the hemispheres. After the age of 50 years such lesions can be the consequences of vascular disease. Contrast administration makes possible the detection of new foci and their differentiation from the old ones. In 65-85% of cases the initial phase is marked by remitting course with more or less evident worsening after each exacerbation. Out of them, in 40-70% of cases the remitting course changes to secondary progressing course. In only 10% of cases the course is progressing from the beginning. The prognosis is better if the disease begins with optic nerve involvement and is worse if pyramidal or cerebellar signs appear first. CSF examination is less important than MRI or EP. In biochemical tests no sure markers of the disease are found. In the treatment of acute exacerbations steroids are still most effective. In clinically confirmed MS interferon beta given in the first 2 years is effective in 30% of cases. PMID- 11107566 TI - [Back pain syndromes]. AB - In the preface the causes are discussed of back-originated pains at different levels. Remark is made of three periods of lesions developing in intervertebral discs. The possibilities are discussed of the induction of back pain syndromes by overloading, and two mechanisms of this effect are described. Special attention is paid to distinguishing radicular from pseudo-radicular syndromes, and the characteristic features of both are specified. These basic features are discussed with reference to individual spinal segments. Five phases of the development of overloading disease are presented. In the closing chapter the causes of acute back pain syndromes are discussed as dependent on age. Also the risk factors of the occurrence of back pain syndromes are listed. PMID- 11107567 TI - [Prophylactic treatment of migraine]. AB - Migraine belongs to the most frequent idiopathic headaches affecting 5-10% of the population. The knowledge of migraine pathogenesis is as yet insufficient for complete elucidation of its causes. There is no effective drug for all patients which could interrupt the attack or prevent its development. The methods of migraine prevention could be divided into pharmacological and non pharmacological. Among the drugs used for attack prevention drugs are mentioned known and used since a long time and drugs less commonly used such as riboflavin, sulpiride, alpha-adrenolytic agents. Own experiences are presented with the use of iprasochrom for migraine prevention. Besides pharmacological methods also non pharmacological methods are discussed which are applied for attack prevention. PMID- 11107568 TI - [Treatment of Parkinson's disease--from theory to practice]. AB - The symptomatic treatment of Parkinson's disease is based on assumption that its symptoms are a consequence of damage to the dopaminergic system at the level of substantia nigra. However, not all symptoms can be explained by this damage, moreover, substitution therapy /with preparations containing dopamine precursor- levodopa and agonists of the dopaminergic receptor /not only fails to remove all disease symptoms but can even produce adverse effects. Theoretical bases of treatment are discussed, including the non-motor signs /dementia, depression, autonomic system disturbances/ with the analysis of how these theoretical assumptions come true in practise, including the author's practice. a schema is proposed of diagnosis establishing and decision taking of treatment with stress laid on the most frequent diagnostic and therapeutic errors. Expectations and hopes are discussed also concerning the search for the cause and better therapeutic methods in the future. PMID- 11107569 TI - [Pathophysiology and treatment of spinal cord medulla injuries]. AB - In view of the ever increasing incidence of spinal cord injuries and their very high socioeconomic costs studies are conducted for reduction of their consequences. In recent years considerable advances have been achieved in their treatment. The contribution of various mechanisms damaging spinal cord is known presently rather well, with isolation of two groups of causes: one is the primary spinal cord injury as a result of direct force acting on it during trauma, the other is secondary damage caused by vascular changes following trauma, free radicals, calcium distribution changes, participation of opioid receptors and inflammatory process. For counteracting these mechanisms in secondary cord damage treatment with drugs is justified. Among the drugs the main role is played by steroids--both glucocorticoids and non-glucocorticoids /lazaroids or aminosteroids/. Other drugs include calcium channel blockers, opioid receptor antagonists, serotonin antagonists, cyclo-oxygenase inhibitors, osmotically active drugs, antioxidants, NMDA receptor antagonists. Besides drugs hypervolaemia, haemodilution and hypothermia are tried. Proper diagnostic procedures and effective treatment of cord injury consequences depend on the knowledge of the mechanisms of later consequences of cord injury, this enables achieving of ever more effective treatment results. PMID- 11107570 TI - [Epidemiology of cerebral stroke in Poland]. AB - In Poland the annual incidence of cerebral stroke is 60,000 and half these patients die within one year, while the remaining one-half has permanent consequences of central nervous system damage. The incidence of stroke is in Poland similar to that in other European countries /the incidence rate is 177/100,000 in males and 125/100,000 in females/ but the death rate of 106/100,000 in males and 79/100,000 in females is among the highest in Europe. One of the main causes of this high death rate is the high mortality in early phase of stroke. On the basis of prospective epidemiological studies of the population in a district of Warsaw it was shown that extracerebral complication were one of the main causes of death in stroke patients. The analysis of the results of these studies served for construction of a model of independent factors increasing the risk of early death after stroke. These factors include age, stroke type, intensity of neurological signs and coexistence of other diseases. In Poland the high prevalence of the risk factors for vascular diseases and frequent coexistence of heart diseases are among the main causes of the high death rate in stroke. Cardiovascular diseases in stroke patients affect both the early and the late mortality through increasing the risk of systemic complication and second stroke. PMID- 11107571 TI - [General cerebral and systemic metabolic disturbances occurring in ischemic stroke with special emphasis on glucose and its metabolites]. AB - Ischaemic stroke is a stressogenic factor triggering a complex defensive reaction called "alarm reaction" by Selye. Stress gives rise to liberation of catecholamines, dopamine beta-hydroxylase in the blood, cerebro-spinal fluid and urine. Patients with ischaemic stroke were found to have increased adrenaline, noradrenaline, and 3-metoxy-4-hydroxymandelic acid level in urine and increased cortisol level in blood serum. Patients, especially those with severe ischaemic stroke have increased concentrations of glucose metabolites in blood and cerebro spinal fluid: pyruvate acid, lactic acid, acetylacetic acid and hydroxybutyric acid. PMID- 11107572 TI - [Evaluation of lactic acid levels in blood of patients with ischemic stroke in the earliest stage of the disease]. AB - The aim of the study was the assessment of concentrations of lactic acid in the blood of patients with mild course and severe course of ischaemic stroke in the earliest stage of the disease. The subject of the study were 20 patients with a mild ischaemic stroke and 20 patients with a severe one on its 1st, 3rd and 7th day. Enzymatic method for determining lactic acid content in the blood was used (the kits Test-Combination Lactate fully enzymatic of Boehringer Mannheim). In the group of patients with mild ischaemic stroke increase values of lactic acid were shown on all days of the disease, but the differences were statistically insignificant. In patients with severe ischaemic stroke these values were higher than in controls, and the differences were statistically significant. The study showed a positive correlation between high concentrations of lactic acid and severe clinical condition of patients. PMID- 11107573 TI - [Evaluation of pyruvic acid concentration in blood of patients with ischemic stroke in the earliest stage of the disease]. AB - The aim of the study was the assessment of concentrations of pyruvic acid in the blood of patients with mild course and severe course of ischaemic stroke in the earliest stage of the disease. The subject of the study were 20 patients with a mild ischaemic stroke and 20 patients with a severe one on its 1st, 3rd and 7th day. Enzymatic method for determining pyruvic acid content in the blood was used (ready made reagents Test-Combination Pyruvate by Boerhinger Mannheim). In the group of patients with mild ischaemic stroke the concentrations of pyruvic acid were increased on all days of the disease, compared with controls, but the differences were statistically insignificant. In the group of patients with severe ischaemic stroke the concentrations of pyruvic acid were higher than the ones in control group, but only on the 1st and 3rd day of the disease the differences were statistically significant. The results of determinations indicate that there is a positive correlation between levels of pyruvic acid and severity of the clinical course of the disease. PMID- 11107574 TI - [Binswanger's disease--diagnostic criteria and analysis of cases]. AB - Binswanger's disease(BD) in an effect of a vascular destruction of deep white matter structures following multiple infarcts leading to disturbances of perception functions, recent memory and characterological disorders. BD is a disease occurring in late 6th decade of life. Computerized tomography and magnetic resonance imaging demonstrate foci of microinfarcts which coexist with arterial hypertension, atheromatous lesions and general cerebral atherosclerosis. BD leads to dementia, certain Parkinson-like signs, pyramidal system involvement, depression, organic brain damage syndrome, dysbasia and sphincter control loss. Ten patients observed by the author are reported. PMID- 11107575 TI - [Evaluation of anxiety and depression in patients with vascular brain damage]. AB - In 90 patients with vascular damage to the CNS an assessment was carried out of the intensity of anxiety on the basis of Anxiety State and Trait Inventory, of the degree of symptom intensity of depression syndrome on the basis of Hamilton Depression Scale and of personality and socioeconomic conditions on the basis of history data in' and Eysenck Personality Inventory. In the studies the hemispheric laterality of brain damage was taken into account. The results were analysed statistically comparing them with those obtained in the control group of healthy men aged 18-24 years. On the basis of the studies and calculations the following conclusions were drawn: 1--the intensity of anxiety and depression in patients treated for vascular CNS damage is greater than in the control group. 2- statistically higher depression intensity was found in patients with left hemisphere damage as compared with the group with right hemisphere damage. 3--in patients living in good social conditions and regarding their family relations as good, the intensity of anxiety and depression was statistically lower than in the group of patients with poor family and socioeconomic conditions. PMID- 11107576 TI - [Stroke in the course of cerebral arteritis]. AB - In this paper we wanted to present cases of stroke occurring in the course of cerebral vasculitis. Cerebral vasculitis occurs most often in patients with infections, tumours and connective tissue diseases. Idiopathic vasculitis is observed very rarely and making the diagnosis meets great difficulties. Three patients (2 women and 1 man) with idiopathic cerebral arteritis aged 28-41 who underwent treatment in our department are prevented. There were no known stroke risk factors in anamnesis and diagnostic investigations. All patients had headaches before the occurrence of stroke. Electrocardiography, echocardiography were normal. Two patients had arteriography--small arterial occlusions and changes in arteries' walls were seen. Ischaemic changes in both cerebral hemispheres were shown in CT and MRI scans. One patient had high serum level of antinuclear antibodies one year later the diagnosis of lupus erythematosus was made. In patients with cerebral vasculitis extensive laboratory investigations should be carried out and combined with a detailed follow-up study. Neurological changes could be the first symptom of connective tissue disease. PMID- 11107577 TI - [A case of hypoxic encephalopathy in the course of chronic spastic bronchitis and pulmonary emphysema]. AB - Respiratory insufficiency appearing during chronic lung diseases leads to hypoxemia, hypercapnia, acidosis, right ventricular failure and secondary polyglobulia. These disturbances lead to respiratory encephalopathy which is characterized by the appearance of various types of neurological syndromes. We present here the case of a patient suffering from chronic spastic bronchitis accompanied by pulmonary emphysema, whose consciousness disturbances, a generalized epileptic seizure and hemiparesis were connected with his respiratory insufficiency intensifying during the basic disease. Removal of metabolic disturbances caused by respiratory insufficiency has a key role in preventing secondary neurological syndromes. PMID- 11107578 TI - [Ischemic cerebral stroke and anabolic steroids (case report)]. AB - Anabolic steroid use is widespread and has been associated with a variety of pathological conditions. The subject of this case is a 22 years old man with cerebral ischaemic stroke. He had no previous medical complaints but had a history of anabolic steroid abuse. This case presents a side effect of long term abuse of these drugs. PMID- 11107579 TI - [Primary and secondary intraventricular hemorrhage--clinical analysis]. AB - In the recent decades an evident increase in the number of diagnosed intraventricular haemorrhages has been noted. The trial was undertaken of assessing of the incidence of primary and secondary intraventricular haemorrhages, the role of aetiological factors in various types of haemorrhages, the risk factors and the results of imaging procedures, as well as the clinical symptoms and treatment methods. The analysed group comprised 90 patients in hospital with the diagnosis of intraventricular haemorrhage in the years 1989 1996. The diagnosis was based on the results of computerized tomography /CT/ and on autopsy in one case. The clinical course of the cases varied considerably: very serious mainly in secondary intraventricular haemorrhages with high mortality. No signs pathognomic for intraventricular haemorrhage were found. The prognosis in primary intraventricular haemorrhage was better. Hypertension as risk factor was present in 71 cases. Pathological changes of blood vessels were found in 23 cases. PMID- 11107580 TI - [Headache in systemic lupus erythematosus]. AB - This study was conducted to analyse the prevalence and features of headache in patients with SLE. 44 patients with SLE were reviewed retrospectively. The mean age of the patients was 43.8 +/- 11.5 years (r: 22-68), mean time of evolution of SLE was 11.8 +/- 9.0 years (r: 1-30). 28 patients (27 woman and 1 man) had clinical evidence of central nervous system involvement, 18 patients (40.9%) had headache; 6 (33.4%) being vascular, 3 (16.7%) migraine, 9 (50%) muscle contraction headache. In 4 patients headache started before the other signs or symptoms of SLE, in 8 patients headache appeared in the first year of the disease. In the patients with headache other manifestations of nervous system involvement were present very often. Most patients had more than one symptom or sign during the course of the illness. The results of our examination showed that headache is frequent in the course of SLE and probably it is connected with vascular lesion of the nervous system. We failed to find a relationship between headache and other manifestations of SLE and the treatment of SLE. PMID- 11107581 TI - [Empty sella syndrome--diagnostic difficulties]. AB - The authors present three middle-aged patients (1 male, 2 females) presenting with empty sella on magnetic resonance or computed tomography imaging. Their main complaints were severe headache and dizziness. Neurological examination showed no abnormalities, except for local tenderness on palpation and percussion in the fronto-parietal or fronto-temporal regions. Endocrine evaluation showed no functional deficit of the pituitary gland. Visual fields, fundi and EEG records were normal as were the CSF composition and pressure, a skull X-ray radiograms showed an enlarged sella with thinned floor and blurred posterior oblique processes. MRI revealed a liquid space below the plane of the sellar diaphragm which pressed on the pituitary gland. The diagnostic and therapeutic problems of empty sella syndrome are briefly discussed. PMID- 11107582 TI - [Levels of interleukin-6 in cerebrospinal fluid of patients with meningitis]. AB - Interleukin-6 (IL-6) is a multifunctional cytokine that regulates the immune response, differentiation of B cells, activation of T lymphocytes and acute phase reactions. AIM OF THE STUDY: Quantitation of interleukin-6 in cerebrospinal fluid from patients with purulent bacterial meningitis (PBM) and viral meningitis (VM) on the first day of hospitalization. PATIENTS AND METHODS: IL-6 activity was measured in cerebrospinal fluid (CSF) of 15 children with VM (predominantly 13/15 caused by mumps virus) in age ranging from 2.5 to 15 years and 13 patients with PBM (age ranging from 18 to 52 years). The control group consisted of 10 patients with meningeal syndrome and normal laboratory estimation of CSF (white cells count, concentration of glucose and protein). The IL-6 assays were performed using immunoenzymatic method, ELISA (Amersham) technique. RESULTS: Increased levels of IL-6 were detected in the CSF of patients with bacterial meningitis and with viral meningitis. CSF IL-6 levels were 6 times greater in patients with purulent bacterial versus viral meningitis. The highest level of IL-6 was observed in a 18 years old patient with pneumococcal meningitis who died. CONCLUSION: We suggest that cytokine levels may be valuable in distinguishing patients with purulent bacterial meningitis from viral meningitis and may help in prognosis of patients with purulent meningitis. PMID- 11107583 TI - [Measurement of acetylcholine receptor antibodies in serum of patients with myasthenia gravis]. AB - The aim of this study is the evaluation of the level of antiacetylcholine receptor antibodies in patients with different types of myasthenia (according to Osserman's classification) and establishing of correlation between the level of such antibodies and the clinical state of the patients. 63 patients with diagnosed myasthenia and 30 healthy controls without immunological diseases were evaluated. In all of them standard neurological examination was performed and the levels of antiacetylcholine receptor antibodies were measured. In 45 patients electrophysiological investigations were carried out. In 43 cases elevated levels of antiacetylcholine receptor antibodies were noted. In 20 patients the levels were within normal range. The level of antibodies showed correlation with the clinical type of myasthenia according to Osserman (the more severe was myasthenia, the higher was the level of antibodies). But the clinical state of the patient at the moment of examination did not show any clear correlation with the level of antibodies. Acetylcholine receptor antibodies measurement has significant diagnostic value in myasthenia. Nevertheless it should be interpreted with other diagnostic techniques. PMID- 11107584 TI - [A case of polymyositis with flaccid paraparesis]. AB - Polymyositis is characterized by inflammatory process in muscles and muscle weakness is the main clinical sign of the disease. We report a case of a 65-years old woman suffering from polymyositis with flaccid paraparesis. Forms of polymyositis where only proximal muscles of lower extremities are involved are very rare, so it may cause diagnostic mistakes. Because of that, it is very important to pay a special attention when making the diagnose of paresis. PMID- 11107585 TI - [Post lumbar puncture syndrome and the manner of needle insertion]. AB - We have analysed the correlation between the direction of needle bevel insertion and the occurrence of post-puncture syndrome appearing after diagnostic lumbar puncture. Post-puncture headache was observed in 38 of 380 patients (13.6%). The syndrome occurred in 11 patients (7.9%) in whom the needle bevel had been inserted parallel to the patient's backbone and in 27 patients (19.3%) in whom the needle bevel had been inserted perpendicularly to the backbone. The complications after lumbar puncture could be reduced by inserting the needle bevel parallel to the patient's backbone. PMID- 11107586 TI - [Brain functional imaging using magnetoencephalography]. AB - Neuromagnetism, the study of neural function by the measurement of magnetic fields, is a new, quickly developing branch of science. Magnetoencephalography is the measurement of weak magnetic fields generated by neuronal activity in the human brain. By measuring the magnetic field evoked around the head, a map of the functional organisation of the brain can be deduced with a sub-centimetre spatial resolution and a millisecond temporal resolution. Although MEG has been used since the early 1970 s it is only recently that large multi channel facilities have been combined with sophisticated analysis techniques and Magnetic Resonance Imaging to provide a tool for fundamental study of the brain. The author presents 9 the other functional imaging tools such as Positron Emission Tomography /PET/, functional Magnetic Resonance Imaging /fMRI/ et Encephalography /EEG/. PMID- 11107587 TI - [Clinical applications of late negative evoked potentials--contingent negative variation (CNV)]. AB - Contingent negative variation /CNV/ is a slow negative potential described first in 1964 by Walter et al. It is a correlate of cerebral activity in the frontal lobes connected with expectation of stimulus and frontal cortex preparation for the stimulus to come. CNV develops in the time between the warning signal /S1/ and the commanding signal /S2/. CNV contains two main components connected directly with brain function: the first one, so called early component, is connected with the process of orientation or warning /it is called also orientation wave/, the second one /late component/ is connected with the preparation for movement /expectation wave or preparatory wave/. The clinical application of CNV is for the evaluation of the correlation of potential changes with changes in cognitive functions occurring in various diseases. Numerous studies reported recently have confirmed the applicability of CNV on the diagnosis of dementia, Parkinson's disease, epilepsy, schizophrenia, anxiety states, chronic pains, including migraine. PMID- 11107588 TI - [Correlation between results of transcranial doppler ultrasonography and SPECT in the brain of patients with ischemic cerebral stroke]. AB - Single photon emission computerized tomography /SPECT/ and positron emission tomography /PET/ are used presently for the study of the cerebral blood flow /CBF/. The cost of these procedures limits the possibility of their use and makes them available mainly in large and rich clinical centres. Transcranial Doppler USG has no such drawbacks. The purpose of the present study was analysis of a possible correlation between marker cumulation /in SPECT/ in the vascularization area of the middle cerebral artery /MCA/ and the parameters Vs, Vm, Vd, PI, RI, R of the PUSG-G examination of these arteries in 50 patients aged 48-79 years treated for ischaemic cerebral episodes. Brain SPECT examination was done with Apex-SP-6-HR gamma camera assessing the distribution of 99mTc HMPAO marker in the vascularization area of both MCA. PUSG-D examination was done with Tc-2-64 unit. In both MCA the systolic Vs, the mean velocity Vm, the diastolic velocity Vd, the Gosling pulsation index PI, the Purcelot resistance index RI and the velocity amplitude R were measured. The following conclusions have been reached: 1/ in patients with ischaemic cerebral episodes a significant correlation was found between cerebral SPECT findings and the PUSG-D parameters; 2/ reduced perfusion of the cerebral tissue was correlated with lower values of Vs, Vm, Vd, and R and with higher values of PI and RI; 3/ slight disturbances of perfusion found in SPECT were not reflected in changed PUSG-D parameters; 4/ the results justify the use of PUSG-D for indirect assessment of blood flow in the MCA. PMID- 11107589 TI - [Use of transcranial doppler ultrasonography in patients with vascular dementia]. AB - The aim of the study was the evaluation of the clinical usefulness of transcranial Doppler ultrasonography /TDU/ in patients with vascular dementia. The study was carried out in a group of 69 patients divided into two groups: group i of 32 patients aged 45-78 years, mean age 58.4 years with two or more ischaemic foci on TDU and dementia symptoms. The cases in this group fulfilled the criteria of dementia according to DSM-IV, ICD 10, MMS scale and Hachinski ischaemic scale. In TDU the following parameters were analysed: maximal velocity /Vmax/, minimal velocity /Vd/, mean velocity /Vmean/ of blood flow in the middle cerebral arteries, pulsation index /PI/ and resistance index /RI/. In group II the patients were divided into three subgroups according to ICD-10 criteria and localization of ischaemic foci: with cortical, subcortical and mixed lesions. The following conclusion have been drawn: 1/ the parameters of TDU could serve for differentiation of patients with vascular dementia; 2/ the PI and RI parameters in TDU are most valuable for diagnosis establishing; 3/ the transcranial Doppler ultrasonography is insufficient for differential diagnosis of dementia. PMID- 11107590 TI - [Ectodermal method of Ryodorak--an attempt at clinical measurement for evaluation of physiotherapy effects in patients with low back pain]. AB - In the group of 132 patients with diagnosed pseudoradicular pain syndrome (57 subjects) or evident sciatic neuralgia (75 subjects) besides clinical and radiological examination, additional skin electro-conduction evaluation, being the sympathetic system activity exponent, was performed. Diagnostic part of empiric method of Yoshio Nakatani, was adapted, where the skin electro-conduction measurement of the direct current of 200 microA in 24 representative measuring points is the essence. The aim of this study was the analysis of electroconductivity in dermatomes corresponding to the level of discopathy and indirect trial of objectivisation of some widely applied in this group of patients physiotherapeutical methods: low frequency diadynamic currents (Bernard's), medium frequency interferential currents (Nemec's), low frequency impulse magnetic field and classic acupuncture. In the case of sciatic neuralgia statistically significant electroconductional differences were found in the range of dermatomes involved in pathological process in comparison to the other half of the body as well as other points of reference. PMID- 11107591 TI - [Spasticity and physical methods for controlling it]. AB - Spasticity is one of the greatest difficulties in patients with central nervous system injuries and diseases. Severe spasticity makes treatment, rehabilitation and care of patient very difficult and sometimes even impossible. It has been sought for many years for an objective method to evaluate the degree of spasticity, necessary to establish the results of treatment and rehabilitation. In this study we present subjective and objective methods of evaluating the spasticity in order to classify every patient to adequate therapeutic group. The authors present physical methods that not only contribute to control of spasticity together with pharmacotherapy and surgical treatment, but can be used alone. The big advantage of this therapy is a low invasiveness and the very few side effects. PMID- 11107592 TI - [Use of modified lateral nucleus pulposus herniation classification in low back pain]. AB - Nucleus pulposus herniation is one of frequent causes in spinal pains. The necessity of undertaking of appropriate treatment, often surgical, requires early establishing of precise diagnosis. The traditional classification of herniations into central, centrolateral and lateral seems to be insufficiently precise, especially from the surgical point of view. For that purpose, the authors, following the classification of Volle et al., applied in their analysis the new classification into central and lateral nucleus pulposus herniae and discerning among the latter ones four types with strictly defined localisation: mediolateral, lateral with recess involvement, intervertebral foramen hernia, outside intervertebral foramen hernia. Seventy-two cases were analysed. Hernia was diagnosed by means of CT. Particular attention was given to the most external hernias, that is into and outside the intervertebral foramen. The more accurate method for the assessment of hernias and bone changes was in CT the Reconstruction in sagittal and frontal planes and also in oblique projections. The authors believe that the diagnosis of lateral nucleus pulposus hernias in low spinal area, particularly those with herniation into and outside the intervertebral foramen should be based on CT in appropriate reconstructions. PMID- 11107593 TI - [Economics and policy of day surgery]. AB - With advances in technology, day surgery has become more efficient and has expanded remarkably due to the policies and economic incentives in some countries. In addition, day surgery could potentially serve as a model of explicit accountability for quality assurance and institutional processes for continuous improvement. It is recommended that Japan adapt its policies and systems to facilitate day surgery after a thorough analysis of the health effects and cost structure. Cost shifts to other services and parties should be considered carefully from a long-term, comprehensive perspective. It could be socially beneficial to subsidize start-up costs for the establishment of day surgery units, since significant capital and human resources are required for quality assurance. The encouragement of day surgery could be a driving force for the improvement of clinical technology and patient quality of life. It would foster collaboration between health service providers, including during preparation and follow-up, and allow patients to participate as partners in clinical processes and decisions. To ensure constant readiness, day surgery environments should be equipped with multisite, standardized databases on clinical and economic performance. An expansion of day surgery facilities could lead to the development of a new mechanism of professional quality improvement and to a new health insurance reimbursement system based on clinical achievements and resources. PMID- 11107594 TI - [Day surgery and anesthesia]. AB - Ambulatory surgery comprised up to 70% of scheduled surgery in the USA during the past decades. The main reason for the increase was economic. In Japan, day surgery has been performed in the pediatric field, such as for inguinal hernia. However, recently day surgery has been performed in adult patients, because of medical economics as well as to improve the quality of life of patients. Patient safety is the most important issue in the increasing number of day surgeries in Japan. Postoperative care occurs at home. To reduce postoperative complications, skilled surgery, prompt recovery from anesthesia, and postoperative care using sophisticated systems, manpower, and equipment are necessary. To ensure patient safety, the Japan Society for Ambulatory Anesthesia announced a set of "safety standards for ambulatory anesthesia" in 1999 and we continue to work to developing protocols that maximize the safety of and benefits to patients. PMID- 11107595 TI - [The day surgery system]. AB - The day surgery system was established in order to satisfy the demand of healthy patients, and we have performed more than 40 types of operation under this system so far. The critical path is an indispensable tool for ensuring safety and efficiency, and the care coordinator is the key person to manage the system for that purpose. In addition, the cost management technique focusing on DRG/PPS and risk management to increase patient satisfaction are particularly important because we must manage numerous surgical cases. The day surgery system is evaluated in relation to the primary education program for surgeons. Such education not only impacts medical and surgical skills, but also is a good opportunity for training in the basic bedside manner to offer the best service to patients. PMID- 11107596 TI - [Day surgery for breast cancer]. AB - In Japan, day surgery for breast cancer usually means partial mastectomy without axillary dissection for small carcinoma under local anesthesia in the outpatient clinic. If histopathological examination of the specimens by serial section reveals that the cancer is noninvasive and completely resected with negative surgical margins, no additional surgery under general anesthesia is needed, and the patient does not require hospitalization. We call this procedure "probe lumpectomy". From 1991 to 1998, 169 patients underwent probe lumpectomy in our institution, and there were no major complications. Of these 169 patients, 64 did not require hospitalization. Ipsilateral breast cancer was observed in two patients, and these tumors were diagnosed not as recurrence but as second primary cancers. No distant metastases were observed. As sentinel node biopsy, which is not always easy under local anesthesia, becomes more common, the indications for day surgery or short-stay surgery will expand. As breast cancer is a malignant disease, informed consent and careful follow-up are needed if the treatment is completed only in the outpatient clinic. PMID- 11107597 TI - [Day surgery for cholecyst lithiasis]. AB - Since 1997, laparoscopic cholecystectomy has been performed as one-day surgery (LC/DS) at our institution. Among the 122 patients enrolled in this program, 97 (80%) were successfully discharged within 24 hours after admission. Discharge was delayed for the other 25 patients, although 12 (48%) of them were discharged on postoperative day (POD) 2 or 3. This study not only verified the efficacy of LC/DS in shortening convalescence and allowing an early resumption of work but also confirmed the safety of LC/DS except in one patient with hemophilia A who required laparotomy for intraabdominal bleeding on POD 13. LC/DS is now the first choice of treatment for cholelithiasis regardless of symptoms. Discharge can be expected within 24 hours after admission in most cases, although the preference of patients should be considered when determining the timing of discharge. PMID- 11107598 TI - [Day surgery for adult inguinal hernia]. AB - The mesh plug technique for adult inguinal hernia repair is easy to perform and results in a good postoperative quality of life. It allows inguinal hernia surgery to be performed as day surgery, but some problems may occur. We performed day surgeries tp repair 110 adult inguinal hernias, and the results are reported here. The procedures are as follows. Under local anesthesia, the inguinal canal and hernial sac are freed. Then the internal ring and the weakness of the posterior wall are estimated. The plug is inserted and then the mesh is on layed. As a result, all of our cases were successful under local anesthesia. After surgery, 5 subcutaneous hematomas occurred. Six patients required subsequent hospitalization: one because of subcutaneous hematoma; and 5 for pain control. In summary, the mesh plug technique under local anesthesia for adult inguinal hernia repair is a useful method for day surgery, but some an on-call system after surgery is necessary and hospitalization for postoperative complications may be required. PMID- 11107599 TI - [Day surgery for pediatric inguinal hernia repair]. AB - Pediatric patients with inguinal hernia can be good candidates for day surgery. Since 1986, 1273 such patients have been treated under our day surgery system. Although 8 patients (0.6%) were unable to go home after surgery due to perioperative complications and 21 (1.6%) visited earlier than scheduled after discharge, no major complications were noted and postoperative complications did not differ from those seen in the inpatient setting. Our questionnaire survey demonstrated that 96% of the families were satisfied with the system. Based on our own experience and a literature review of day surgery for inguinal hernia repair in children, the problems surrounding the day care system are discussed. Patient selection, preoperative assessment, general anesthesia, postoperative care including oral intake and analgesia, and postoperative follow-up are considered the most important issues in the day care system. A team approach including pediatric surgeons, anesthesiologists, and pediatric nurses is considered indispensable for the safe and satisfactory day surgery treatment. PMID- 11107600 TI - [Day surgery for anal disease]. AB - Historically, patients with anal diseases treated on a day surgery basis had inadequate cure rates and a high complication rate. After World War II, modern treatment methods were learned from the UK and USA and improved in Japan. However, the improved radical methods were so complex that approximately 2 weeks' hospitalization was needed. Recently, day surgery for various diseases including hemorrhoids has been recommended by the Japanese ministry of Health and Welfare. However, the characteristics of anal anatomy and physiology make the smooth healing of wounds difficult and tend to cause postoperative pain, bleeding, infection, prolonged healing time, etc. To prevent such difficulties, care must be well planned following the critical path of informed consent, careful surgery, postoperative observation, and management at home. However, hospital staff in charge of such surgery are under so much stress that only patients with less severe anal disease without local or systemic complications should be selected for day surgery. PMID- 11107601 TI - [Day surgery for treatment of varicose veins in the leg]. AB - We had surgically treated varicose veins in 554 legs of 386 patients as of June 30, 2000. Varicose veins of the stem or segment type without skin changes were treated with sclerotherapy combined with high ligation, while a part of secondary varicose veins and the reticular or web type were treated with sclerotherapy alone. This paper describes our methods for day surgery for this condition. The most important therapeutic consideration in the surgical procedure is achieving sufficient venous collapse to prevent the occurrence of intravenous thrombus. In our 386 patients, a massive intravenous thrombus that was resected occurred in one limb (0.1%). Postoperative bleeding also occurred in one limb (0.1%) of a patient with severe liver cirrhosis. PMID- 11107602 TI - Arguments for a single-payer national health insurance program. PMID- 11107603 TI - Psychologists and prescription privileges. PMID- 11107604 TI - Prescribing behaviors of Colorado advanced practice nurses. PMID- 11107605 TI - Improving pediatric asthma outcomes using self-management skills. AB - To improve pediatric asthma care in practices with limited time and staff, patients and families should be taught self-management skills to avoid crisis asthma care. A trained asthma team in a private practice setting can educate families of children with asthma. Specialized forms help the team assess a family's initial asthma knowledge, track their learning progress, and guide the staff and family during their self-management skill-development discussions. Team members ensure that the family leaves each visit with the appropriate information and skills, leading the family toward a more stable and predictable lifestyle. PMID- 11107606 TI - Forms facilitating primary care documentation. AB - Documentation is a time-consuming but vital component of patient care. As health care providers function under increasing time constraints, well-designed forms can help simplify and improve the documentation process. Patient record forms can help clinicians identify essential points of the history and physical examination and can provide anticipatory guidance. One classification system frequently used by primary care providers is the evaluation and management (E/M) code system. E/M codes are a subset of the Current Procedural Terminology codes developed by the American Medical Association to standardize the terminology used to describe medical and surgical services. This article presents one documentation system that primary care providers may find effective in their documentation efforts. PMID- 11107607 TI - Culturally appropriate guidelines for alcohol and drug abuse prevention. AB - Information on the efficacy of wellness protocols in culturally diverse populations is meager. A study was conducted to develop culturally appropriate guidelines to prevent alcohol and drug abuse. Five groups of women participated in focus groups; based on the participants' perceptions, Put Prevention into Practice guidelines for the prevention of alcohol and drug abuse were adapted for each group. Further research studies evaluating these adapted guidelines and developing additional culturally appropriate tools are needed. PMID- 11107608 TI - Hypertension prevalence in a rural Haitian missionary clinic. PMID- 11107609 TI - Linezolid: a new class of antibiotic. PMID- 11107610 TI - Malnutrition in the older adult. PMID- 11107611 TI - Molecular epidemiology of bovine tuberculosis. I. Mycobacterium bovis genotyping. AB - The lack of a typing system for Mycobacterium bovis has, until recently, been an impediment to undertaking sophisticated epidemiological studies to assist in the control and eradication of tuberculosis in domestic animals. Molecular biology techniques for mycobacteria have been in development since the mid-1980s, leading to the availability of a number of genetic typing systems for M. bovis. The authors summarise the available techniques, identify those which are most useful at present and those which might prove useful in the future. The present recommendation is to use spoligotyping analysis for rapid, large scale screening of M. bovis isolates, and to use restriction fragment length polymorphism analysis using the polymorphic guanine and cytosine-rich repeat sequences probe where greater differentiation of isolates is required. In the future, systematic analysis of the genome sequence of M. bovis will allow the development of improved techniques that combine good discrimination with ease of use. PMID- 11107612 TI - Molecular epidemiology of bovine tuberculosis. II. Applications of genotyping. AB - The applications of genotyping of Mycobacterium bovis are reviewed. Published research to date has been conducted predominantly within the context of validating typing methods, and few studies have been specifically epidemiological. This is contrasted with the situation in human tuberculosis, where the application of restriction fragment length polymorphism typing using insertion sequence IS6110 has successfully led to insights into the epidemiology and molecular evolution of the pathogen. Based upon the medical experience, the adoption of an integrated approach which combines epidemiology and molecular biology is recommended for future studies. Accordingly, clear identification and explanation of type clustering should be possible, which should facilitate decisions related to disease control. PMID- 11107613 TI - The use of participatory appraisal by veterinarians in Africa. AB - The term 'participatory appraisal' refers to a range of methods for data collection, learning and facilitation, which enable local people to play an active role in defining, analysing and solving their problems. A questionnaire survey was used to obtain information on the use of participatory appraisal (PA) from veterinarians working in Africa. A low overall response rate of 28.6% was achieved. Within Africa, response rates varied from 15.6% from government veterinarians to 47.6% from veterinarians working with non-governmental organisations. Information is presented on preferred methods, specific uses, levels of training and perceived advantages and disadvantages of PA. While PA was considered by many informants to be a valuable approach to working with communities to analyse and solve local animal health problems, respondents also identified constraints to the wider use of PA. These constraints included lack of financial resources, low availability of relevant training courses and materials, lack of time to attend training courses, and negative attitudes among colleagues. The author concludes that greater institutional awareness of the role of PA in the development of Veterinary Services is required. Such awareness might be achieved by wider dissemination of experiences related to the use of PA and the development of veterinary-orientated training courses for centrally-based personnel and workers in the field. The latter should include attention to appropriate attitudes and behaviour for veterinary professionals who are attempting to develop services according to the priorities and capacity of the community. PMID- 11107614 TI - Designing serological surveillance programmes to document freedom from disease with special reference to exotic viral diseases of pigs in Denmark. AB - Surveillance programmes based on laboratory screening tests are increasingly used to document freedom from disease in order to facilitate trade. The following aspects must be considered when designing such programmes: diseases to be selected; epidemiology of the diseases; unit of analysis (animal or herd); target age group (or target farm type); test characteristics and sample size. Issues related to these aspects are discussed and illustrated using the example of serological surveillance for exotic viral diseases in the pig population of Denmark. Sampling designs based on individual animal samples are compared with herd-based sampling (two-stage sampling). While the latter is likely to require a larger sample size, the increased level of information and the reliability of the results obtained are considered to be worth the expense. Issues related to the development of international standards for declaring freedom from disease are discussed. The authors conclude that international standards are desirable, providing that these standards represent scientifically valid principles. PMID- 11107615 TI - Control of zoonoses in Cyprus. AB - The excellent results achieved in the control of animal diseases in Cyprus have allowed the Veterinary Services to take a leading role in the elimination, surveillance and investigation of important zoonoses. The programmes for the control of echinococcosis, brucellosis, bovine tuberculosis, anthrax and taeniasis, and the measures taken to prevent the importation of rabies are described. Public awareness of the risks posed by the presence of zoonoses and the efficient and effective intersectoral co-operation achieved between the veterinary, medical, public health and other Government services and non governmental organisations are considered to be the key to the successful control of zoonoses in Cyprus. PMID- 11107616 TI - [Information program applied to the temporal study of pathological processes in animal populations]. AB - WinEpi Tasas 2.0 is a new, globally accessible computer program, which has been developed as a research tool for the epidemiological study of the pathological processes which affect animal populations. This new version of the program, based on the experimental version, WinEpi Tasas 1.0, enables users to study the distribution and evolution of diseases in affected populations, over time. This information, combined with data on spatial distribution, is of fundamental importance for the establishment of appropriate control strategies and health policies. Such disease control measures are of particular significance in the current context of emerging and re-emerging diseases. The program can be run in either English or Spanish. PMID- 11107617 TI - Identification of antigenic epitopes on the foot and mouth disease virus isolate O1/Manisa/Turkey/69 using monoclonal antibodies. AB - A panel of mouse monoclonal antibodies (MAbs) was produced against a strain of type O foot and mouth disease virus (FMDV) from the Middle East, O1/Manisa/Turkey/69. Seven neutralising MAbs were fully characterised and all were found to react with conformational epitopes. Monoclonal antibody neutralisation-resistant mutants (MARMs) were generated from the parental virus stock and the complete capsid sequences of these MARMs were determined. Sequence analysis revealed that five of the MARMs had amino acid substitutions at either residue 72 or 73 of VP2 (beta B-beta C loop), indicating that five of the MAbs were directed against antigenic site 2. The sixth MARM contained a single amino acid substitution at position 198 of VP1 (carboxy-terminal region). The seventh MARM contained two amino acid substitutions, at position 72 of VP2 and position 149 of VP1 (beta G-beta H loop). These findings indicate that MAbs directed against a type O FMDV from the Middle East recognise residues in the same structural features to those raised against strains from Europe of the same serotype. PMID- 11107618 TI - Development and evaluation of a monoclonal antibody based competitive enzyme linked immunosorbent assay for the detection of rinderpest virus antibodies. AB - A competitive enzyme-linked immunosorbent assay has been standardised for the detection of antibodies to rinderpest virus in sera from cattle, sheep and goats. The test uses a neutralising monoclonal antibody (MAb) directed against the haemagglutinin protein of rinderpest virus. The test is specific for rinderpest, as it failed to detect antibodies to peste des petits ruminants virus in convalescent goat sera. A 45% inhibition of the binding of the MAb to the antigen was used as the cut-off point for deciding the rinderpest status of the test samples. The specificity and sensitivity of the test and the stability of the test reagents were determined and compared to the results obtained using a commercial kit with approximately 1,200 serum samples from cattle, sheep and goats in India. The current test compared very well with the commercial kit. The test is expected to be extremely useful for sero-monitoring and sero-surveillance of rinderpest in countries which are actively pursuing a rinderpest eradication programme. PMID- 11107619 TI - Spatial risk factors related to outbreaks of contagious bovine pleuropneumonia in northern Italy (1990-1993). AB - In October 1990, an outbreak of contagious bovine pleuropneumonia (CBPP) was reported in Italy after an absence of approximately one century. Since October 1990, ninety-four outbreaks have occurred in Italy, of which forty-seven were concentrated in three areas of northern Italy (Lombardy region). The disease was eradicated in September 1993. The data used for the analysis were obtained from the epidemiological investigations undertaken during the outbreaks of 1990-1993. The unit of interest for the analysis is the farm. Spatial segregation of infected and uninfected farms within the study area was determined through the Pielou index of segregation. Data from herds within the same set of outbreaks were analysed through logistic regression to identify factors which could be used to discriminate between infected and uninfected herds. The study indicated a clear spatial segregation between infected and uninfected herds. The results of the analysis do not indicate the mode of disease spread. However, the study demonstrated that both aerosol and indirect transmission of the infection could have occurred, as previously documented in Africa. The possibility that indirect transmission played a prominent role in the spread of CBPP in the region of Lombardy is a completely new suggestion. Further studies are required to understand the epidemiology of CBPP in regions with intensive farming and a relatively cold climate. In particular, three aspects require consideration; firstly, animal movement among neighbouring herds could produce a pattern of disease similar to the one produced by indirect transmission (this possibility was excluded in the epidemiological outbreak investigations conducted by field veterinarians). Secondly, the methods of spatial analysis used in this study have not been previously used in the field of veterinary epidemiology. Further validation of the efficacy of these methods is thus required. Thirdly, the epidemiology of CBPP under conditions of high animal density and a relatively cool climate, as encountered in Lombardy, requires further investigation. PMID- 11107620 TI - [Comparative study of the immunizing ability of some attenuated strains of sheep pox virus and of a sensitizing vaccine]. AB - The authors present the results of a study designed to compare the immunogenicity of several attenuated strains of sheep pox virus and a virus/immune serum vaccine. Two of the strains studied present immunogenic characteristics that make them particularly interesting for the manufacture of a vaccine. The first, named Djelfa, confers solid immunity to animals without provoking a vaccinal reaction; the second, known as Romania, can provide protection beyond twenty-four months, but causes persistent vaccinal lesions. In a country like Algeria, these two strains could be used for immunoprophylaxis of sheep pox: the first in those regions in which prevalence is low and sheep are vaccinated on a regular basis, and the second in regions of high prevalence where herds are moved to new pastures each season. PMID- 11107621 TI - Evaluation of control programmes for echinococcosis/hydatidosis in Cyprus. AB - In Cyprus, echinococcosis (Echinococcus granulosus) was an important public and animal health problem until the 1970s. In 1971, the Department of Veterinary Services instigated a control programme, which was based on an island model, using arecoline testing, euthanasia of positive dogs, control of dog breeding, elimination of stray dogs and control measures in slaughterhouses. The first programme ended in 1985 with excellent results and E. granulosus was considered to have been eradicated from both the definitive hosts (dogs) and the intermediate hosts (livestock). However, surveillance from 1985 to 1993 demonstrated that the life-cycle of E. granulosus was continuing at very low levels. Control measures were therefore reintroduced in 1993. The second programme commenced in 1993 and was based on surveillance of intermediate hosts, control of infected flocks, and testing and treatment of owned and stray dogs in infected areas. A continental model was employed for the second programme, due to the division of the island in 1974. The experience gained from the two programme is described. PMID- 11107622 TI - Reservoir hosts of Leptospira inadai in India. AB - Isolation of Leptospira from the kidneys of Rattus rattus wroughtoni hinton, Rattus rattus rufescens, Bandicota bengalensis and Bandicota indica was attempted in Bangalore in southern India. In total, 296 spirochaetes were isolated from 1,348 kidney cultures (an isolation rate of 22%). A batch of fifty-six isolates from India was identified, based on serological and polymerase chain reaction analysis, of which twenty-three isolates were identified as L. inadai by the World Health Organization/Food and Agriculture Organization Collaborating Centre for Reference and Research on Leptospirosis, in Brisbane. This is the first record of isolation of L. inadai from rodents. The preponderance of L. inadai in four different species of rodents suggests that these animals could be the natural reservoir hosts of L. inadai, and raises a critical question as to the likely impact of this species of Leptospira on the renal carrier status of other Leptospira pathogenic to humans and animals in this part of India. Virulence studies conducted at the University of Trieste in Italy, revealed that isolates of L. inadai from India were moderately or totally serum resistant when subjected to a serum killing test. To establish the possible seroprevalence of this species in the population, the inclusion of L. inadai in the battery of leptospiral antigens used for sero-epidemiological studies is recommended. PMID- 11107623 TI - [First description of concomitant infection with tuberculosis and paratuberculosis in dairy sheep in Argentina]. AB - The authors present the results of a comparative intradermal tuberculin test performed on dairy sheep to determine rates of reaction to bovine and avian purified protein derivative (PPD). Test reactions were interpreted by palpation and measurement with a calliper. Results were analysed by plotting the values obtained. Agar gel immunodiffusion tests, growth in culture and histopathological examinations were also performed to allow differential diagnosis between tuberculosis and paratuberculosis. In the first tuberculin test, performed on 288 sheep, 74 animals showed a positive reaction to avian PPD (25.69%), two animals showed a positive reaction to bovine PPD (0.69%), and one animal was positive with respect to both tuberculins (0.34%). In the second test, involving 247 sheep, 55 demonstrated a positive reaction to avian PPD (22.26%), whilst none reacted to bovine PPD. The presence of Mycobacterium avium subsp. paratuberculosis was confirmed by culture and serological testing. In addition, lesions compatible with those caused by M. bovis were found by histopathological examination. These findings highlight the importance of the accurate diagnosis of tuberculosis and paratuberculosis for the effective control of both diseases. PMID- 11107624 TI - [Validation of an indirect immunoenzyme assay for the detection of antibodies against Trypanosoma evansi in horses in Argentina]. AB - An indirect enzyme-linked immunosorbent assay (ELISA) to detect antibodies against Trypanosoma evansi was evaluated using 90 different sera, obtained from naturally-infected horses. As negative controls, 218 sera from the T. evansi-free zone of Argentina, and 90 uninfected sera from the enzootic zone were used. The results of the ELISA were expressed in terms of percent positivity (PP) when compared with a positive primary reference serum, obtained from a horse experimentally-infected with T. evansi. The inter-assay coefficient of variation (CV), expressed as PP, was 44.7% for the negative control serum, 8.8% for the mildly positive reference serum, and 9.2% for the secondary positive control serum, while the intra-assay CV for each of the above sera was 6%, 2.8% and 5%, respectively. Positive and negative serological results were differentiated using a histogram of the distribution of the results obtained using sera from infected and uninfected animals from the enzootic zone (expressed in PP). A PP of 50 indicated a sensitivity of 95.5% for a confidence interval (CI) of 91.3% to 99.7%, and a specificity of 98% for a CI between 95% and 100%. Positive and negative predictive values were established for each rate of prevalence between 0.01% and 25%. The use of reference control sera in each assay enabled reproducible results to be obtained. The author recommends that this methodology be used whenever certification of the T. evansi status of horses is required, and particularly when animals are to be moved from an infected to a disease-free area. PMID- 11107625 TI - Veterinary research at the Central Veterinary Laboratory, Weybridge, with special reference to scrapie and bovine spongiform encephalopathy. AB - A Veterinary Laboratory Service was commissioned by the Ministry of Agriculture, Fisheries and Food of the United Kingdom in 1894 and the Service commenced veterinary research in 1905. The Central Veterinary Laboratory (CVL) was opened in 1917 and has become known world-wide under the name 'Weybridge'. In 1922, a network of Veterinary Investigation Centres was established in England and Wales and these continue to make an important contribution to surveillance for animal diseases. Problems recognised at these centres provide an important stimulus for research at the CVL, a good example being the case of bovine spongiform encephalopathy (BSE). Research into many non-infectious and infectious diseases is conducted, and research into scrapie was already in progress when BSE was discovered in 1986. A research programme was commenced to investigate the clinical signs, diagnostic methods, pathology, pathogenesis and epidemiology of BSE and the transmission characteristics of the BSE agent in farm animals. Some of the results of these studies and prospects for the future are discussed. PMID- 11107626 TI - Macrophage recruitment and activation: a model for comparing resistance to Salmonella enteritidis in different broiler breeds. AB - A model for comparing resistance to Salmonella Enteritidis was evaluated in different broiler breeds. The recruitment and phagocytic activity of peritoneal macrophages were assessed in three different broiler breeds (A, B and C) which are farmed world-wide. Assessment was performed after three days of intraperitoneal (i.p.) administration of 3% Sephadex G-200 (10 ml), initiated at twenty-one days of age, followed by contact with i.p. live S. Enteritidis (10 ml, 1.2 x 10(8) colony forming units/ml) for 45 min. Assessment included determination of the number of i.p. macrophages recruited, the number of i.p. phagocytized S. Enteritidis cells per macrophage, the levels of degranulated i.p. beta-glucuronidase and beta-galactosidase, and the count of surviving S. Enteritidis cells. Confirmation of the significance of the model was obtained by comparing resistance to field infection by S. Enteritidis in the three broiler breeds. The recruitment of i.p. macrophages in response to challenge with Sephadex and S. Enteritidis was significantly higher (P < 0.05) in birds of breed A (mean cumulative i.p. macrophage count, in 10 fields of microscopic slide smear magnified at x1,000, was equal to 81.7), compared to recruitment in birds of breed B (33.3) or breed C (41.2). The mean number of phagocytized S. Enteritidis cells per i.p. macrophage in birds of breed A (2.68) was significantly higher (P < 0.05) than in breed B (0.83) and insignificantly higher (P > 0.05) than in breed C (2.35). In addition, the highest level of recruitment and phagocytic activity of macrophages, in birds of breed A, was associated with a higher significant mean i.p. beta-glucuronidase activity (10,425.5 units/ml) than in breed B (3,438.2 units/ml) or breed C (3,356.94 units/ml) (P < 0.05). Moreover, birds of breed A demonstrated a higher mean i.p. beta-galactosidase activity (2.225 units/ml) than birds of breed B (0.852 units/ml) or breed C (1.852 units/ml) (P > 0.05). The higher level of recruitment and activity of i.p. macrophages and the higher rate of degranulation of i.p. enzymes in breed A were associated with a greater number of surviving i.p. S. Enteritidis cells. In response to outbreaks of S. Enteritidis in the field, the average mortality was significantly higher in flocks of breed A (3.2%) than in flocks of breed B (1.2%) or breed C (0.96%) (P < 0.05). These data provide an indication of the significance of the model in reflecting the differences in resistance of S. Enteritidis of broiler breeds reared in a farm environment. PMID- 11107627 TI - Implementation of a system for the regional management of animal health emergencies. AB - A telematic system to support decisions and operations in case of animal health emergencies has been designed and implemented in the Abruzzo region of Italy. The system aims to improve decision-making by Veterinary Services in the event of an outbreak of exotic disease. The system has been tested, first by a simulated outbreak of foot and mouth disease, and then during an outbreak of swine vesicular disease. Critical problems were detected and corrected in both cases. PMID- 11107628 TI - Sero-surveillance of wild boar in The Netherlands, 1996-1999. AB - From 1996 to 1999, blood samples were collected from wild boar shot during the hunting season in Crown properties, national parks and the free wildlife belt in the Netherlands. Sera were screened for the presence of antibodies against classical swine fever virus (CSFV), swine vesicular disease virus (SVDV), Aujeszky's disease virus (ADV) and Trichinella spiralis. The results of the sero surveillance system indicate that CSFV, SVDV and ADV are uncommon within the wild boar population. Hence, the wild boar population is not thought to be an important reservoir of these viruses in the Netherlands. Infection with ADV and CSFV is endemic in the wild boar population in Germany. Since contact between the wild boar populations of Germany and the Netherlands cannot be excluded in the southern part of the Netherlands, continuation of the sero-surveillance system seems appropriate. In the decade before 1998, no antibodies to Trichinella spp. were found in the wild boar population of the Netherlands. The detection of some seropositive animals during the hunting season of 1998-1999 corresponds to the previous findings in wild boar before 1988. However, the recent data do not have consequences for the pig industry of the Netherlands, since the country has been considered Trichinella-free for many decades. PMID- 11107629 TI - Epizootiological aspects of peste des petits ruminants and rinderpest in sheep and goats in Saudi Arabia. AB - Epizootiological aspects of peste des petits ruminants (PPR) and rinderpest in sheep and goats in Saudi Arabia are examined. The presence of PPR has been suspected on occasions, but virus isolation has been successful only once. Information regarding PPR and rinderpest in sheep and goats in Saudi Arabia is scarce. The only survey conducted indicated that neither disease is endemic in the country. PMID- 11107630 TI - Mycobacterium paratuberculosis infection in two free-ranging Alpine ibex. AB - The authors report two cases of Mycobacterium paratuberculosis infection in free ranging Alpine ibex (Capra ibex) from two different herds in the Western Alps, Italy. One ibex, found dead in October 1998, was in poor condition. The second animal died due to trauma following capture with a dart gun. The only gross lesions observed were the enlargement of the mesenteric and iliac lymph nodes. Samples from both ibex tested positive to polymerase chain reaction for a primer set specific for the M. paratuberculosis insertion sequence IS900 and one ibex also tested positive to the Zielh-Nielsen stain. Isolation by bacterial culture was not successful. The infected ibex originated from herds in which seroreactors to M. paratuberculosis had been found previously. Seroreactors to M. paratuberculosis were also detected in sympatric cattle. PMID- 11107631 TI - Contagious ecthyma associated with myiasis in sheep. AB - A severe outbreak of contagious ecthyma (orf) is described in sheep in Saudi Arabia. In some of the affected sheep, the condition was highly aggravated by myiasis which appeared to have been favoured by the abundance of flies during the hot season. The outbreak is discussed in relation to the epidemiology of the disease in Saudi Arabia. PMID- 11107632 TI - Immune response of buffaloes to vaccination with Brucella abortus strain 19. AB - The immune response of buffalo calves and heifers to a full or half dose of Brucella abortus strain 19 vaccine was studied. Buffalo calves developed high serum agglutination test (SAT) titres following full dose vaccination. These titres declined more rapidly in calves of six months of age than in calves of eleven to twelve months of age. Specific immunoglobulin G titres, as measured by 2-mercaptoethanol-treated serum agglutination, declined much earlier than SAT titres. Buffalo heifers vaccinated with a full or half dose developed high SAT titres. The rate of decline of SAT titres in heifers was much slower than in calves. Vaccination with a half dose did not appear to offer any advantage in terms of disappearance of SAT titres. PMID- 11107633 TI - Genomic diversity and prevalence of Rotavirus in cow and buffalo calves in northern India. AB - Faecal samples were collected from seventy-eight diarrhoeic cow and buffalo calves between November 1998 and February 1999 to study the genomic diversity and prevalence of Rotavirus infection by ribonucleic acid polyacrylamide gel electrophoresis (RNA-PAGE) and enzyme-linked immunosorbent assay (ELISA). In the organised dairy farm (where daily production and health records were maintained), the overall prevalence of infection with Rotavirus, recorded by RNA-PAGE and ELISA, was 27.02% (10/37) in both cow and buffalo calves. In unorganised dairy herds (where no production or health records were maintained), RNA-PAGE and ELISA detected infection with Rotavirus in 26.8% (11/41) of cow and 19.5% (8/41) of buffalo calves. Five distinct electropherotypes were found to circulate in cow and buffalo calves. All were short electropherotypes except the single long electropherotype observed in a buffalo calf in an unorganised dairy herd. Some differences in RNA migration pattern were observed when these electropherotypes were compared with the neonatal calf diarrhoea virus strain of Rotavirus. Some electropherotypes were restricted to one farm while others were found in both organised and unorganised dairy herds and in both cow and buffalo calves. PMID- 11107634 TI - [Standards for human physiological requirements in food substances and energy: a retrospective analysis and a developmental outlook]. AB - The retrospective analysis of 4 variants of norms of human physiological needs in food substances and energy accepted in 1951, 1968, 1982 and 1991 is given. Dynamics of the scientifically proved specification of value of needs is revealed and the necessity of permanent development of conceptual base of normalization is determined. Last one is based on two natural laws of a balanced diet: 1) compulsion of conformity energy expense and energy consumption; 2) the value of consumption of the basic food substances (fibers, fats, carbohydrates) should be in limits of physiologically necessary quantity proportion and be accompanied by satisfaction of human needs in different nutrients. PMID- 11107635 TI - [The dynamic nature of the nutrition of the population of 1 of the districts of Moscow over a 10-year period]. AB - On the basis of three-multiple research of character of a feed of the inhabitants of one of Moscow district by the standardized method of the 24-th hour interrogation reveals significant changes in structure of a feed of the population from 1986 to 1996. The shifts have appeared more dynamical in the second five-year from 1991 to 1996. Nevertheless, atherogenicity of ration of a researched population with superfluous consumption of the saturated fats and simple carbohydrates remains. Is established, in a feed of the women there were large shifts, than at the men. The structure of a feed of the inhabitants of Moscow differed from structure of a feed of the inhabitants of Russia. PMID- 11107636 TI - [The actual nutrition of children in the pediatric institutions of Kemerovo]. AB - Dietary intake from kindergartens and boarding schools was studied. Analysis of nutrient and energetic content of rations was given. It is determined that there are a disbalance of the main components and deficient of some vitamins and minerals. Recommendations for correction of children's nourishment were proposed. PMID- 11107637 TI - [The determination of the double bounds in blood serum lipids by titration with ozone: the pathophysiology and diagnostic significance]. AB - In this article the method of definition of double binders in lipids of blood serum in patients with different diseases basically atherosclerosis and ischemic heart disease, with use titration by ozone is given. The received data testify to an opportunity of use of this method in the diagnostic purposes. PMID- 11107638 TI - [The vitamin and mineral composition of the diets of adolescent schoolchildren in the Far North]. AB - Screening examination of pupils in 13-16 ages of Nadym by method of 24h.dietary interrogatory detected complex vitamin insufficiency and imbalance of mineral composition of juveniles diets. It was showed necessity of correction of pupils nutrition which are living in the conditions of the Far North. PMID- 11107639 TI - [The adaptation characteristics of infants with a perinatal lesion of the central nervous system in relation to the nature and schedule of their feeding]. AB - We studied in dynamics 72 babies with cerebral perinatal disturbances aged up to 3 months (32 boys and 40 girls) in City Maternity Homes No 1 and No 4 and in New Born Pathology Department of City Infant Hospital No 1 in Krasnoyarsk. All babies were divided into four groups according to the character and schedule of feeding: I-st group--babies, fed with mammary milk at the first 2 hours and being fed more than 10 times during the day with obligatory night feeding; II-nd group--babies, fed with mammary milk at the first 2 hours and being fed 6 times during the day usually with night break; III-rd group--babies, fed with mammary milk in 6-8 hours after birth and being fed more than 10 times during the day with obligatory night feeding; IV-th group--babies, fed with mammary milk in 6-8 hours after birth and being fed 6 times during the day with usual night break. We have marked faster tempo of normalization in indices of neurological status and physical development in groups of infants, fed with mammary milk at the first 2 hours and being fed more that 10 times during a day with obligatory night feeding as compared to babies, which were fed according to common schedule. We didn't find the dependence from the stage of perinatal pathology of nerve system. Frequency of functional disturbances in cardio rhythm, malfunctions of gastrointestinal tract, allergy diseases and iron-deficiency anemia in babies, fed with special food in early age (not mammary milk) is evidently higher as compared to babies, fed with mammary milk only. We received the results of examination in dynamics which prove, that mammary milk creates favorable background for the development of adaptation processes in baby organism, which allows faster compensation of residual factors, connected with perinatal disturbances of nerve system. PMID- 11107640 TI - [The effect of an antiatherosclerotic diet including omega-3 polyunsaturated fatty acids of marine and plant origins on the indices of cellular and humoral immunity in patients with ischemic heart disease and a disordered carbohydrate tolerance]. AB - The dynamic of natural antibodies against catecholamines and alpha-2 macroglobulin, thrombin, antithrombin III and parameters of cellular immunity in 92 patients with ischemic heart disease and ischemic heart disease complicated by impaired glucose tolerance was studied of influence of antiatherosclerotic diet with fish and vegetable PUFA omega-3 from "Eicolen". Besides favorable influence to a clinical picture of the disease universal normalizing influence of antiatherosclerotic diet with addition Eicolen on parameters of humoral a cellular immunity. PMID- 11107641 TI - [Fluoroplastics: their properties, hygienic aspects and the possibility of their use in contact with food products]. AB - In the article the questions of fluoroplastes destruction, opportunity of migration in foodstuff non-polymerized monomers, additives and chemical substances formed as a result of thermal disintegration of polymer are considered. PMID- 11107642 TI - [Human ecology. The developmental outlook for the manufacture of products for functional use in Kaluga Province]. PMID- 11107643 TI - [An improvement in the hygienic instruction of workers in dairy industry enterprises]. AB - The article presents the experience of the work on the sanitary education of the workers of the milk industry plants in Moscow and Moscow Province. The Institute's specialists had worked out basing on the studied experience the program of the full-time and external differential sanitary education of the workers of the milk industry which was approved by the Department of the State Sanitary Control of the Ministry of Health of Russia the April 22, 1998. "An exercise book for the sanitary education of the workers of the milk industry plants" was also prepared and published in 1999. PMID- 11107644 TI - [Accelerated methods for the microbiological quality control of food products in the critical control point system in the analysis of the hazard factor]. AB - The rapid microbiological methods of quality control food in the HACCP systems. The paper contain the summary of the hazard analysis control point concept for the control of food microbiological safety. It characterises the express methods that can detect and identify the Food Poison Organisms, including the methods for the rapid detection and enumeration of microorganisms by means of a new impedance measuring systems, molecular-biologic methods and DNA-diagnostic methods. PMID- 11107645 TI - [Molecular basis and mechanisms of Influenza viruses adaptation to reproduction in murine lung]. AB - Analyzes modern data on the role of individual segments of genome in acquisition of virulence during influenza virus adaptation to reproduction in the lungs of mice. Discusses the significance of molecular mechanisms (loss of potential glycosylation site(s), optimum pH of HA-mediated fusion, beta-inhibitor sensitivity, pH-dependent association/dissociation of M1 protein with viral RNP, and host factors) involved in adaptation of influenza virus to a new host. PMID- 11107646 TI - [Genetic analysis of tick-borne encephalitis virus strains from West Siberia]. AB - Tick-borne encephalitis (TBE) virus strains were isolated in West Siberia in the forest-steppe region near the Ob river in 1981-1992. Hybridization of genome RNA of 46 TBE strains with [32P]cDNA of TBE Sofyin strain revealed essential differences in the genomes of West-Siberian and Far-Eastern Sofyin strains of TBE virus. Nucleotide sequences of 6 TBE strains (1348-1503 n.) have been determined. A 89-98% homology of Siberian TBE strains has been shown, while the similarity of the respective fragment of E gene for West Siberian and Sofyin strains was no more than 81%. No significant changes in E gene of TBE strains have been detected over a 12-year period. PMID- 11107648 TI - [Variability and prevalence characteristics of Influenza A virus (H1N1) in period 1990-1998]. AB - Circulation of influenza A(H1N1) viruses in Russia and CIS countries had a wave like pattern with period of silence in 1990-1995 and activation in 1995-1998, when these viruses were isolated together with A(H3N2) and B viruses. Antigenic drift of epidemic strains' hemagglutinin (HA) was directed to alteration of HA in reference strains A/Texas/36/91, A/Johannesburg/82/96, and A/Beijing/262/95. A/Moscow/17/98 strain similar to A/Beijing/262/95 was isolated on MDCK cells for the first time in European Russia. This means that A/Beijing/262/95 (H1N1) spread sporadically in the country at that time. Comparative analysis of HA thermosensitivity of influenza A(H1N1) strains of 1977-1998 showed a tendency to increase of their thermal stability. The sensitivity of erythrocytes of different animals to A(H1N1) strains isolated during the same epidemic season was different. Differences in amino acid sequences of epidemic strains' HA varied from 5 to 14 sites in comparison with the reference strains, depending on the reference strain and year of isolation. PMID- 11107647 TI - [Isolation of Getah virus (Togaviridae, Alfavirus) strains in North- Eastern Asia]. AB - Fifteen strains of Getah alfavirus were for the first time isolated from Aedes and Culex mosquitoes in Yakutia, Magadan region, Buryatia, and Khabarovsk region of the Russian Federation and in Mongolia. The area of this virus dissemination in the above regions was steppe, mixed forest, Northern taiga, and forest-tundra zones, reaching the tundra zone in the North. Getah virus is the only alfavirus occurring under such severe climatic conditions. PMID- 11107649 TI - [Criteria of primary screening of attenuated strains for live vaccine against Influenza A]. AB - Reassortant strains for live influenza vaccine (LIV) were selected using two additional markers: intensity of cytopathic effect (CPE) at 40 degrees C in MDCK cells and toxicity for mice (induction of acute hemorrhagic pulmonary edema after intranasal challenge with undiluted virus). All wild-type viruses induced a high CPE in MDCK cells, while the reassortants differed by this sign. Only vaccine strains and attenuation donors were characterized by a low CPE. Modern epidemic viruses are highly toxic for mice, causing the death of 60-100% animals from hemorrhagic pulmonary edema on days 3-4 after intranasal infection. Attenuation donors and vaccine strains were not toxic for mice, the level of toxic effect correlating with CPE in MDCK culture. Evaluation of CPE in MDCK culture and toxicity for mice can be used for primary screening of candidates for LIV. PMID- 11107650 TI - [Protective effect of cycloferon in experimental influenza]. AB - Protective effect of granulated cycloferon has been studied in albino mice infected with influenza. The drug induced interferon production and prolonged the life span, decreased the mortality, suppressed the virus reproduction in the lungs, and decreased the infective activity of the virus. Morphologically the drug decreased the intensity of viral lesions in the bronchiolar epithelium, impeded the infection dissemination, and stimulated the productive (cellular) component of local inflammatory reaction. No influence on the development of chronic lesions in the lungs was detected. PMID- 11107651 TI - [Isolation and characteristics of anti-adenovirus monoclonal antibodies in immunoenzyme and immunofluorescent reactions]. AB - New monoclonal antibodies (MAbs) to adenovirus hexon, highly active in ELISA and immunofluorescent analysis, were prepared. According to competitive ELISA, new MAbs differed in their blocking activity and were directed to 2 different hexon epitopes. MAb 3H8 did not modify antigen binding of the rest MAbs labeled with peroxidase (PAb x Pox), and none of unlabeled MAbs suppressed the reaction of MAb x Pox 3H8. MAbs 1E8 7F1, 1E11, and 3B1 reacted with each other but differed by the spectrum and level of competitive inhibition, which indicated that they were directed to different epitopes of adenovirus hexon. Comparison of the specific activity of MAbs 7F1 and 1E8 in direct immunofluorescent detection of adenovirus antigens in infected cell cultures and clinical materials from patients showed a good coincidence (90-97%) of the results with the IMAGEN Adenovirus test (Dako) and with polyclonal FITC conjugates to adenovirus hexon. PMID- 11107652 TI - [Morphology and formation of symplasts in cell cultures infected with measles virus]. AB - The morphology and formation of giant cells were studied in diploid r-68 and MRC 5 cells in comparison with Vero cells at different times after inoculation with measles Leningrad-16 virus by electron and light microscopy and morphometry. The virions were released mainly from mononuclear cells and small syncytia. The nuclei were positioned centrally in the syncytia formed by diploid cells and at the periphery in Vero cell culture. The examined cultures were similar in terms of virus titers but differed by morphology, size and number of syncytia per unit of surface and time course of their formation. In comparison with Vero cells, in diploid r-68 and MRC-5 cells the maximal number involved in giant cells was 5 and 3.5 times lesser and the number of syncytia per surface unit 20 and 13 times lesser, respectively. Human r-68 diploid cell culture was characterized by the least number and size of syncytia over the entire course of experiment and longest life of the monolayer. PMID- 11107653 TI - [Effect of experimental herpesvirus infection on morphofunctional state of mast cells]. AB - Mast cells are involved in the reaction of the organism to herpesvirus infection, which manifests by changes in their count, morphology, and function. The onset of infection is characterized by primary activation of mast cells, the peak of disease by suppressed function, and convalescence by secondary activation and stabilization of cell morphology and function. Viral infection running in the presence of immunosuppression caused by cyclophosphamide is characterized by deep persistent suppression of the morphology and functions of mast cells at the peak of disease. PMID- 11107654 TI - [Use of kinetic hemagglutination inhibition test in study of antigenic relationships of hantavirus strains circulating in foci of hemorrhagic fever with renal syndrome in Southern Far East of Russia]. AB - The kinetics of reaction between hemagglutinating antigens and immune sera to 10 hantavirus strains (7 isolated in the Primorye territory and 3 in other regions) was studied in the cross kinetic hemagglutination inhibition (KHAI) test. An A parameter is suggested for evaluation of the similarity and differences between the studied strains. This parameter helped detect antigenic differences between Hantaan and Seoul viruses and the inter-type differences between Hantaan-like strains. KHAI is more specific than HAI studies of antigenic relationships between strains of different hantavirus serotypes. PMID- 11107655 TI - [Cessation of circulation of wild-type poliomyelitis virus strains in the population is a principal condition for certifying the eradication of poliomyelitis]. PMID- 11107656 TI - [Two problems arising at the final stage of poliomyelitis elimination]. PMID- 11107657 TI - Invasive meningococcal disease in Canada, 1 January 1997 to 31 December 1998. PMID- 11107658 TI - Pseudo-outbreak of Pseudomonas putida in a hospital outpatient clinic originating from a contaminated commercial anti-fog solution--Vancouver, British Columbia. PMID- 11107659 TI - Recognizing and managing boundary issues in case management. AB - Much of the literature on worker-client boundaries in clinical practice is based on assumptions that the relationship between the two parties is structured and formal. These assumptions do not always apply in community-based case management practice, where the worker and client interact in a wide variety of settings and circumstances. The relative informality of case management makes the establishment of appropriate worker-client boundaries both critical and difficult. In this article key principles for recognizing and managing boundary issues are presented and discussed. PMID- 11107660 TI - Facilitating comparisons between evaluations of case management programs. AB - We need to know if, as an overall approach to service delivery, case management is delivering on its promises of improving quality, decreasing costs, and increasing access to services. Attempts to do this are hindered by the very limited comparability currently possible between evaluations of case management programs. One cause of this limited comparability is that few individual evaluation reports provide clear and explicit information on key aspects of the evaluation, and on the case management program itself. Consequently, there is a need to ensure that the key characteristics of individual case management programs and their evaluations are clearly described when the results are reported. As a first step in achieving this, a review of the evaluation theory literature and the case management literature was undertaken to find effective ways of describing the differences in evaluations and in case management programs. The result is a draft framework to support the acknowledgment, identification, and description of these differences. PMID- 11107661 TI - An insider's perspective of managed mental health care. AB - Managed mental health care has been examined from different perspectives in scholarly literature. It has been examined from the provider's perspective through scholarly writing. Executives of managed care companies have also provided their input in this arena. One group of professionals involved in managed care has not contributed to the literature. That is, the case managers employed by the managed mental health care companies themselves. Issues in managed mental health care are discussed from the perspective of the case managers employed by the managed care companies. This group can view managed mental health care from different standpoints. Thus, case managers can provide unique insights into managed mental health care. PMID- 11107662 TI - The language of memory: the influence of writing and reading on the lives and well-being of senior adults. PMID- 11107663 TI - Effects of the home health care interim payment system on access to home health care for people on Medicare. PMID- 11107664 TI - Creating the strategic future of long-term-care organizations. AB - The operating environment in the health care industry is turbulent--organizations are expected to adapt or die. This paper addresses the structure of a strategic planning process for long-term-care organizations. Nursing homes, assisted living (personal care) facilities, continuing care retirement communities, adult day services centers, hospice programs and home- and community-based agencies face both opportunities and threats. The authors recommend an eight-step process for strategy making: plan to plan; external analysis; internal analysis; vision; matching current and future strategies; strategy choice; action and linkage to operations and budget. A case example illustrates the concepts. Long-term-care leaders are encouraged to plan for their future or face a future planned by competitors and regulators. PMID- 11107665 TI - Home care in jeopardy. The impact of severe fiscal pressures on patients, management, and staff: a family member's perspective. PMID- 11107666 TI - Home care in jeopardy. The impact of severe fiscal pressures on patients, management, and staff: the perspective of management and staff. AB - This article has summarized staff experiences and reactions to a revolutionary financing methodology which has been mandated for the nation's Skilled Nursing Facilities. The true impact, beyond the initial observations outlined above, has yet to be realized. In an industry that is characterized as a large cost center of the nation's health bill, it is expected that severe measures will be taken to stem the rising costs; but it is also an industry that directly fulfills a nation's solemn obligation to its most vulnerable and needy constituents. The long-term-care industry has always suffered from marginal staffing ratios and heavy workloads for its providers, thus any new measure that gives nurses more to write with less time to write it means decreased morale, burnout, and career migration. This package will be more palatable when the computer technology, so prevalent elsewhere in our society, is effectively visited on this process. To date, none of these technologies has made the grade. It is of more than passing interest that the pressure brought by PPS and the heavier care patients in SNFs have produced legislation on mandatory staffing with bills in Arkansas, California, Florida, and Minnesota. It will be of great interest to see how the new U.S. President and Congress will address these vital issues. PMID- 11107667 TI - HGV: a "confusing" guest. PMID- 11107668 TI - [Ethical and deontological evaluation of the treatment of comatous patients]. AB - The caretaking of the patient in coma requires an anthropological and clinical approach. The ethics of well-done work suggests to reject futile medical treatment and euthanasia but, at the same time, to perform a correct palliative care and to support the family. PMID- 11107669 TI - [Ethical questions in the treatment of the person in persistent vegetative state. The symbolic case of Nancy Beth Cruzan]. AB - The article deals with a the ethical issues about treatment of Persistent Vegetative State (Pvs) patients. The Nancy Beth Cruzan case, the US woman who died after the withdrawal of tube feeding and hydration after seven years of Pvs is analysed as paradigmatic case. The ethical analysis face with the following issues: 1. Is the Pvs synonymous of cerebral death?; 2. How the tube feeding and hydration must be considered: ordinary or extraordinary, proportionate or disproportionate means? 3. The issue of the living will; 4. the economic impact of the management of the Pvs patients. The conclusion of the contribution is the following: the withdrawal of the artificial feeding and hydration of a Pvs patient must be considered as omissive euthanasia, and consequently it is an action ethically unacceptable, in the light of the Hippocratic medical tradition and of the person-centred ethics. PMID- 11107670 TI - [Fact-finding investigation of the social health conditions of the elderly]. AB - PURPOSE: The intent of the present contribution is that to verify the social health conditions of the elderly present on national territory. MATERIALS AND METHODS: The Authors have thought up an anonymous questionnaire, made up of 63 questions, concerning the life-style, the social conditions and health of the elderly over 65 years old, and consequently by now on pension. Two thousand questionnaires have been distributed to medical officers, present in north as well as in the centre and in the south of the Country; of these we have then received 1386 questionnaires for statistical processing. RESULTS: The results have been about the same for both sexes as well as for geographical distribution. The greatest part of the elderly are self-sufficient from an economical point of view, in spite of a certain economical dissatisfaction present among the males in the north. The women appear to be more active, inclined to travel. The voluptuary habits, of which smoking and drinking are almost absent in all the national territory and in both sexes. As far as prevention in concerned, consequently submitting oneself to medical check-ups, is more accepted in the south by both males and females and in the north and centre only by women. CONCLUSIONS: Our fact-finding study has given good results, and a good conformity in the portion of people interviewed. There could be a greater impulse given to the prevention of pathologies, perhaps with the assistance of free prevention campaigns revolted toward the elderly, and still dedicating more time to our "grandparents" that must not be considered a burden, but rather as a resource. PMID- 11107671 TI - [Quality of life in hemodialyis patients: the effect of educational status]. AB - INTRODUCTION: Hemodialysis has a major influence on the quality of life of chronic renal failure patients. Great attention is currently paid to the development of supporting programmes for this patient group. Aim of this study was to evaluate the quality of life in maintenance dialysis and to research the influence of various factors related to treatment and ESRD on quality of life, taking into account also the level of school instruction. PATIENTS AND METHODS: Reduced functional abilities, as measured by the Sickness Impact Profile (SIP), and Functional Living Test (FLT), derived by Karnofsky Activity Scale were assessed; Hospital Anxiety and Depression Scale (HAD) and semistructured interviews, including a clinical grading of symptoms were considered vs. age, duration of dialysis, level of school instruction. The study was performed with 40 hemodialysis patients, aged 57.4 +/- 14.9 years (range 22-79), treated since at least three years. RESULTS: Significant (P < 0.05) independent correlates with higher SIP scores (greater disability) and Functional Living Test were lower educational level, and the score of Hospital Anxiety and Depression Scale (HAD). No correlation was found for any of the three scales vs. age and vs. dialytic age; no gender difference was observed. DISCUSSION: A greater care in considering Quality of Life questionnaires is warranted, especially for the severe interference of instruction level of patients on results. QALY (Quality Adjusted Life Years), used as a tool for decision-making in clinical and political subsets, can include critical bias that invalidate conclusion. PMID- 11107672 TI - Detection and clinical evaluation of GBV-C/HGV in plasma from patients with chronic hepatitis of unknown etiology. AB - PURPOSE: The role of GBV-C/HGV virus infection in patients with chronic hepatitis of unknown etiology has been studied. Two groups of patients have been included: a) 50 consecutive patients with chronic hepatitis (mean age 47 yrs; M/F = 35/15), negative for markers of hepatitis viruses (HBV-HCV). These patients did not show evidence of metabolic, autoimmune, alcoholic or toxic liver disease; b) 50 healthy blood donors (mean age 35 yrs; M/F = 35/15). All the subjects were HIV seronegative. PATIENTS AND METHODS: The detection of GBV-C/HGV RNA was performed by reverse transcription-polymerase chain reaction techniques and capillary zone electrophoresis analysis (CZE). RESULTS: There was no difference between the two groups of patients (14% vs 6%; p = 0.18 NS). CONCLUSIONS: The etiopathogenetic role of GBV-C/HGV in chronic hepatitis of unknown etiology is yet to be shown. PMID- 11107673 TI - Apoptosis in recent myocardial infarction. AB - PURPOSE: Apoptosis is considered a common pathological feature in acute myocardial infarction (MI) and heart failure; however its role in the later phases post MI has not been characterized. The goal of our study was to investigate by pathological examination human hearts at 20 to 30 days post MI and identify signs of ongoing cell apoptosis. MATERIALS AND METHODS: Two hearts were collected at autopsy from patients who died 20 to 30 days from the onset of MI (Cases 1 and 2). Gross anatomy and light microscopy examination of the hearts was performed to define the infarcted area and the infarct-related artery. The in situ end-labeling of DNA fragmentation (TUNEL) was performed to identify apoptotic cells and the apoptotic rate (AR) was calculated. RESULTS: There were no signs of acute necrosis in any of the specimens examined. A high number of myocardiocyte were positive at TUNEL examination in specimens obtained at sites of infarction, mean AR = 44%, but not in specimens derived from the same patients at regions remote from the MI, AR = 0. CONCLUSIONS: High grade apoptosis is present at sites of infarction and not in regions remote from the infarcted area in the later phases post MI. These data support persistent myocardiocyte loss and identify a possible explanation of progressive left ventricular dysfunction in the subacute phases of MI. PMID- 11107674 TI - [Treatment of hypertension in the year 2000: do we all agree?]. PMID- 11107675 TI - [Migraine]. PMID- 11107676 TI - [Imaging of the breast tissue]. AB - Mammography remains the most important breast exam; mammography, know to be highly sensitive in detecting microcalcifications. Ultrasound is not suitable for screening, but it allows enough resolution to discriminate the very subtle differences of acoustic impedance among the breast tissue. The indications for MRI of the breast is far established may be defined as follows: 1) patients with silicone implants with or without mastectomy; 2) patients whose breast are difficult to evaluate by combined mammography and ultrasonography, who have: a) had breast conservation therapy, b) axillary lymph-node metastasis from an unknown primary tumor; c) postoperative scarring; d) proven carcinoma of one breast, MRI being performed to exclude multifocality. The authors recommend caution in the use of breast MRI in the assessment and management of suspected recurrent carcinoma. PMID- 11107677 TI - [Paraneoplastic syndromes of the central nervous system]. AB - Paraneoplastic syndromes of central nervous system are rare neurologic syndromes caused by cancer but not secondary to metastases. The physiopathologic mechanisms underlying these syndromes are still under debate. We report the biological and clinical features of the most frequent paraneoplastic syndromes involving the central nervous system. Their early clinical identification might be an useful marker of an otherwise unknown visceral malignancy. Furthermore, they might also be suggestive for the particular type of cancer present. Once, therefore, the diagnosis of these paraneoplastic syndromes has been established, an appropriate evaluation for the asymptomatic neoplasm in cancer-free individuals or investigation for the malignancy recurrences in oncologic patients might be performed. PMID- 11107678 TI - [Renal carcinoma: effective modulation of low-dose interferon-alpha and interleukin-2 with medroxyprogesterone acetate and 13-cis retinoic acid]. AB - We report a case history of a patient with clear renal carcinoma. After surgical treatment of primary tumor patient had been treated with medium-high doses IL-2 and alpha IFN for lung metastasis and paraaortic nodes, without significant response. Subsequently, low doses alpha IFN/IL-2 produced a response, and further response have been obtained by the combination of low-doses alpha IFN/IL-2 plus medroxyprogesterone acetate and cis-retinoic acid. We can therefore conclude that in immunogenic tumors, such as renal cancer, various immunologic strategies are justified, also employing in combination drugs not active as single agents, or modifying doses and schedules. PMID- 11107679 TI - [67 year old man with abdominal pain, anorexia and low-grade fever]. AB - A case of pericardial effusion presenting clinically with pretamponade is shown. TBC is not a rare cause of Pericardial effusion and at present Tuberculosis is a more and more frequent infection also in occidental countries. Guide lines for diagnosis and treatment are revised. PMID- 11107680 TI - [The success of chemotherapy in cancer. Cure of Hodgkin's disease. I]. PMID- 11107681 TI - [Platelet glycoprotein IIB-IIIA receptor inhibitors in patients with ischemic heart disease]. AB - Glycoprotein IIb-IIIa receptor inhibitors are the newest anti-platelets drugs currently used in patients with coronary artery disease. We examined mechanisms of their action and different pharmacokinetic and pharmacodynamic characteristics of the four glycoprotein IIb-IIIa antagonists evaluated in randomized, controlled and multicenter trials. We reviewed results of these trials in the settings of percutaneous revascularizations procedures or unstable coronary syndromes. Platelet glycoprotein IIb-IIIa receptor inhibitors reduced incidence of cardiac death and myocardial infarction during the short- and midterm, and benefit was greater in: a) patients undergoing coronary angioplasty with or without stent implantation, particularly in the presence of unstable angina, diabetes or complex and diffuse coronary artery disease; b) as a direct therapy of unstable coronary syndromes, particularly in patients with refractory angina, diabetes and elevated Troponin; more recently they have been used as adjuvant therapy in acute myocardial infarction. Infusion of these drugs was not associated with higher rates of major bleedings. PMID- 11107682 TI - [Ramipril and new therapeutic possibilities for ACE-inhibitors]. PMID- 11107683 TI - Fibrous dysplasia of the temporal bone. PMID- 11107684 TI - Vocal fold granuloma: the "ball-valve" phenomenon. PMID- 11107685 TI - Endoscopic view of a hematoma of the nasal septum. PMID- 11107686 TI - Electronystagmography: dizziness and vestibulocollic symptoms. PMID- 11107687 TI - Aggressive invasive fungal sinusitis. PMID- 11107688 TI - Sleep-disordered breathing: a survey of otolaryngologic practice at military hospitals. AB - We conducted a survey of otolaryngologists at all Veterans Administration and Department of Defense hospitals in the United States to ascertain the nature and scope of their treatment of sleep-disordered breathing. Questionnaire responses indicated that head and neck surgeons in military hospitals have a strong interest in the management of patients with snoring and sleep apnea. Because of the difficulty in obtaining timely sleep test results and the low number of referrals from physicians who perform such testing, many otolaryngologists expressed a desire to be able to perform their own sleep testing. PMID- 11107689 TI - Patients' perceived outcomes after stapedectomy for otosclerosis. AB - We conducted a retrospective study of 29 patients who had undergone stapedectomy for otosclerosis to determine how well their subjective perceptions of hearing improvement correlated with objective audiometric measurements. Patients expressed their assessments of hearing function by completing two versions of the Hearing Disability and Handicap Scale (HDHS). One version of the HDHS was based on patients' retrospective recollections of their hearing impairment prior to surgery, and the other reflected their assessment of their current function. We evaluated these HDHS data both separately and in conjunction with pre- and postoperative audiometric findings. Following surgery, the group's mean pure-tone average improved significantly, from 58 to 27 dB--that is, the average patient had a moderately severe hearing loss preoperatively and only a mild hearing loss postoperatively. Significant improvement was also reflected in the difference between the mean pre- and postoperative HDHS scores, although some patients indicated that they experienced almost no improvement. Overall, our findings indicated that there was a relationship between objective and subjective assessments of hearing improvement following surgery, but that it was weak. Although most patients perceived significant improvement, the degree of that perceived improvement cannot be predicted from the pure-tone audiogram. We conclude, therefore, that a significant difference between audiometric findings and HDHS self-assessments is useful in identifying patients who might benefit from additional counseling and/or aural rehabilitation. PMID- 11107690 TI - The histopathology of routine tonsillectomy specimens: results of a study and review of literature. AB - Controversy continues to attend the routine histologic examination of tonsillectomy specimens. We performed a retrospective evaluation of 400 tonsil specimens removed from 200 patients. We found that 68.3% of the specimens contained reactive lymphoid hyperplasia, 13.5% had follicular hyperplasia, 10.0% represented acute or chronic tonsillitis, and 7.5% were normal. Only one case of malignancy was detected: a non-Hodgkin's lymphoma in one tonsil of a patient who was a cigarette smoker and who had asymmetric tonsils. The results of our study, taken in combination with data already published in the literature, indicate that routine histologic examination of tonsillectomy specimens is unnecessary and results only in added costs and a loss of man-hours. However, in patients who have certain preoperative risk factors, a histopathologic evaluation of tonsillar specimens remains mandatory. PMID- 11107691 TI - Clinical experiences with acute mastoiditis--1988 through 1998. AB - The incidence of acute mastoiditis has declined dramatically during the postantibiotic era. Even so, antibiotic-resistant or unusual pathogens can still cause this disease entity. At our hospital, we documented an increase in antibiotic-resistant and atypical pathogens such as Actinomyces spp. and Mycobacterium tuberculosis. In this paper, we discuss the optimal diagnosis and treatment strategy for acute mastoiditis, and we describe our retrospective review of 13 patients with mastoiditis who were treated at our hospital from 1988 through 1998. Eight of these patients recovered following treatment with intravenous antibiotics, with or without myringotomy, and five who had complications of disease were managed surgically. Among these five, one developed chronic otitis media and one developed cholesteatoma 3 years later. For patients with acute mastoiditis, we emphasize the need to be aware of any unusual pathogens that do not respond to empiric antibiotic therapy. PMID- 11107692 TI - Gene transfer into the central nervous system using herpes simplex virus-1 vectors. AB - Manipulation of gene expression in developing or in mature central nervous systems (CNS) holds a promise for the resolution of many compelling neurobiological questions, including the feasibility of gene therapy to treat diseases of the brain. In this context, a number of viral vectors have been used in recent years to introduce and express genes into the CNS. This article discusses a gene transfer system based on the Herpes Simplex Virus-1 (HSV-1). We describe here the use of non-replicating, non-toxic HSV-1 vector, 8117/43, in a series of studies carried in our joint program. This vector proves further the utility of HSV-1 as a delivery vehicle to a number of distinct sites within the CNS. PMID- 11107693 TI - The origin of cells of the cochlear ganglion in early human embryos. AB - The study was conducted on 6 human embryos at stage 13. It was found that the facial-vestibulocochlear complex is closely related to the otic vesicle, and the particular components of that complex may be distinguished. They show different cellular arrangement and shape. The neural crest cells migrating from the dorsal hindbrain are continuous with cells forming the cochlear ganglion. This gives evidence for neural crest contribution to the cochlear ganglion. PMID- 11107694 TI - Three-dimensional appearance of bovine epidermal keratinocytes in different stages of differentiation revealed by cell maceration and scanning electron microscopic investigation. AB - The epidermis of the modified skin of the bovine hoof is a highly mechanical loaded tissue. Consequently, all cell connections have to withstand high mechanical forces. As an adaptation to this stress, the epidermal keratinocytes show characteristic surface modifications. Furthermore, the tissue displays a complex three-dimensional architecture which is difficult to appreciate from histological sections. SEM-observation of macerated tissue samples is a fast, easy to use and reliable tool to receive three-dimensional information about the appearance and spatial relationship of cells within a tissue. Using cell maceration, the aim of this study was to separate individual as well as smaller groups of keratinocytes in order to reveal the formations of the cell surface, the appearance of individual cells and the spatial relationship of cells within the tissue. A NaOH maceration method described in literature was modified and applied to tissue samples from the wall and bulbar segment of the hooves of six cows. The method facilitated separation between the epidermal cells. Single cells as well as cell groups were available for SEM observation which revealed a three dimensional appearance characteristic for different stages of differentiation of the keratinocytes. The observed findings suggest that throughout the process of differentiation the surface modifications provide the basis for a stable cell to cell adhesion which is established by desmosomes and the intercellular cementing substance. Additionally, the broadened cellular surface area is related to the supply of the highly metabolic active living epidermal cells with nutrients and oxygen. Longer cell processes typically found in the central surface area of the keratinocytes may carry gap junctions and may be involved in cell communication. This, however, has to be clarified by further electron microscopic studies. The demonstrated appearance of individual cells and the complex architecture enable the hoof epidermis to fulfill its unique biomechanical functions. PMID- 11107695 TI - Immunocytochemical evaluation of reorganisation of keratinocyte cytoskeleton induced by change in Ca2+ concentration in culture medium. AB - Ca2+ level-induced changes in the arrangement of cytoskeleton of cultured keratinocytes were estimated immunocytochemically, by evaluating expression of specific cytokeratins, desmoplakin and tubulin. Keratinocytes were isolated from fragments of skin of dead human foetuses. Culture of epidermal cells was performed in two phases: phase I yielded cells of high proliferation abilities in serum-free Keratinocyte SFM of low Ca2+ level (0.03 mM); in phase II differentiated cells were obtained in Dulbecco medium of a high Ca2+ concentration (1.2 mM). Immunocytochemical evaluation of phase I and II cells revealed an array of differences which involved mainly expression and distribution of specific cytokeratins, distribution of tubulin, testifying to a different microtubule arrangement and distribution of desmoplakin and indicating a tendency to form desmosomes. The changes were induced by the changes in Ca2+ level in the culture medium. PMID- 11107696 TI - Ultrastructural immunogold study on the various cell types of cultured pancreatic islets of adult rats. AB - Whereas several reports describing the ultrastructure of the intact pancreatic islets have been recorded, published experience with the ultrastructural integrity of the cultured pancreatic islets is limited. The present study was, therefore, undertaken to provide an ultrastructure identification of the different cells in the cultured islets of the adult rat pancreas, after marking their secretory granules with gold particles. Pancreatic islets were isolated from adult male Wistar rats by the intraductal perfusion of collagenase technique. The islets were cultured in RPMI-1640 medium for 3 days and processed for preparation of ultrathin sections. The sections were stained with the indirect immunogold technique for insulin, glucagon, somatostatin, and pancreatic polypeptide. Ultrastructural examination of the cultured islets clearly identified the presence of B, A, D and PP-cells, as indicated by the numerous gold particles concentrated predominantly over the secretory granules. The secretory granules of the various cell types of the cultured islets demonstrated several similarities as well as differences from the recorded results of the corresponding secretory granules of the intact islets. The differences probably reflect a deviation in the underlying mechanisms of synthesis, maturation and secretion of the different secretory products of the cells in the cultured islets as they adapt to the in vitro environment. PMID- 11107698 TI - The neuronal structure of the substantia nigra in the guinea pig: Nissl and Golgi study. AB - The studies were carried out on the mesencephalos of adult guinea pigs. The preparations were made by means of the Golgi technique, as well as the Nissl and Kluver-Barrera methods. Four types of neurons were distinguished in the substantia nigra (SN) of the guinea pig: 1. Bipolar neurons of two kinds: the neurons of the first kind have elongated, fusiform perikarya (25-40 microns), whereas the cells of the second kind have rounded and oval perikarya (15-22 microns). These neurons possess two dendritic trunks which arise from the opposite poles of the cell body and run for a relatively long distance. The bipolar neurons are the most numerous in the pars compacta of SN. 2. Triangular neurons with three primary dendrites arising conically from a perikaryon (20-35 microns). They are the most often observed type of neurons in the pars reticulata of SN. 3. Multipolar neurons with quadrangular or oval perikarya (22-35 microns) and 4-5 dendritic trunks which spread out in all directions. 4. Pear-shaped neurons (perikarya 15-25 microns), which have one or two primary dendritic trunks arising from one pole of the cell body. In all the types of neurons an axon originates either from the dendritic trunk or from the soma and is observed only in its initial segment. PMID- 11107697 TI - Cholinergic endings on various neurones containing calcium binding proteins and glutamic acid decarboxylase in the hippocampus of the rat. AB - Immunohistochemical study of the cholinergic innervation of the hippocampal cells containing glutamic acid decarboxylase (GAD) and calcium binding proteins: parvalbumin (PV), calbindin D28k (CB) and calretinin (CR) was conducted on 5 adult rat brains. Analysis of sections with double immunostaining for vesicular acetylcholine transporter (VAChT; the marker of cholinergic cells, fibres and terminals) and respectively either GAD or PV, CB, CR, using confocal laser scanning microscope shows that the intensive cholinergic innervations receive GAD, PV and CB-positive hippocampal cells. Cholinergic afferentiations of the CR positive neurones are considerably fewer. PMID- 11107699 TI - SEM demonstration of elastic fibres in the integument of small and densely-haired mammals. AB - The combination of SEM and autoclave methods gave a clear three-dimensional demonstration of integumental elastic fibres in small densely-haired mammals. The specific organisation of a fine and spongy elastic network was characterised by uniformly thin elastic fibres which were homogeneously distributed between both hair follicle types throughout the whole dermis. All the hair follicles were connected with each other by elastic fibres along their complete intradermal length. The advantage of such a specific elastic dermis construction is that all hair follicles can be moved together and simultaneously along the entire body, so that a better and rapid insulation is achieved after erection of the hair follicles during very low temperatures. PMID- 11107700 TI - Early development of the cruciate ligaments in staged human embryos. AB - Investigations were carried out on 43 serially sectioned human embryos of developmental stages 18 to 23. The homogeneous interzone of the future knee joint is observed in embryos at stage 18. During stage 19 this interzone is differentiated into dense, intensively stained, peripheral parts, which are the primordia of menisci and the medial portion, in which the cruciate ligaments are formed. All structures of the interior of the knee joint are more clearly delineated during stage 20, and they are well developed during the last embryonic week (stages 21-23). PMID- 11107701 TI - Sex differences in the bony pelvis of the fruit-eating bat, Eidolon helvum. AB - Investigation into the bony pelvis of the fruit-eating bat, Eidolon helvum, revealed differences in the pelvic dimensions. The pubic joint of the female consists of interpubic ligament that increased in length until puberty. In the juvenile male Eidolon the pubic joint consists of ligament. Ossification of the interpubic ligament in the male is not an adult phenomenon but begins at the juvenile stage. In the adult the distance from the ischial tuberosity to the iliac crest, from the ischial tuberosity to the medial end of the pubis, and the outlet anteroposterior diameter of the pelvis, were significantly higher in the male than in the female. This indicates that the hip bone in the male is longer in both the vertical (height) and transverse (width) measurements than in the female. The dimensions of the adult pelvic inlet and outlet transverse diameters were significantly higher in the female than in the male. This indicates that the female pelvic canal is wider than the male and this is a consequence of the female interpubic ligament. PMID- 11107702 TI - Prenatal development of coronary arteries in the rat: morphometric patterns. AB - The aim of this work was to address morphometric patterns of coronary artery (c.a.) development in the rat based on serial section analysis of hearts at different stages of prenatal development. Studies were performed on foetal hearts 15-21 days (ED) post-conception. Paraffin sections were stained with haematoxylin eosin (H&E) and frozen sections were labelled with Griffonia simplicifolia I (GSI) lectin (endothelial cell marker). Coronary arteries' luminal diameters were measured at different distances from the aortic roots and the main c.a. branch lengths were calculated from serial sections. All measured values were compared to heart length and to foetal stages. On ED15 precursors of c.a. were distinguished as tubes running on both sides of the outflow tract. Below the aortic valves the tubes had the largest diameter. Formation and development of c.a. proceeded by elongation of vascular tubes distally, ramification and formation of the media and the adventitia. During the prenatal period the c.a. length increased approximately 14-fold, while heart length increased about 4 fold, and crown-rump length about 2.5-fold. The lumen of the proximal part of c.a. increased 4-fold during ED18-21. An increase in c.a. length is the fastest compared to the heart growth, and crown-rump growth during the foetal life. PMID- 11107703 TI - Influence of low and high doses of fluoride on tooth germ development in rats. AB - The influence of fluoride on tooth germ development, especially mineralised tissue, is well documented in numerous dental publications, but there are few reports concerning the influence of fluoride on enamel organ and dental papilla cells. The aim of the study was to assess histologically the development of tooth germs of 20-day-old rat foetuses whose mothers drank water without fluoride or with low (10 mg) and high (110 mg) contents of natrium fluoride, starting from the 12th day of the pregnancy. The fluoride contained in drinking water in low as well as high concentration accelerated the development of enamel organ and dental papilla structures in rat foetuses. The acceleration was proportional to the content of fluoride in drinking water. No disturbances caused by high concentration of natrium fluoride were observed. PMID- 11107704 TI - The localisation of the electrode in permanently paced heart--an echocardiographical study. AB - Permanent cardiac pacing is a widely applied procedure in invasive cardiology. The aim of our study was the analysis of the localisation of the tip of the pacemaker lead and its course in the right ventricle. Research was carried out on a group of 12 patients (5F, 7M), from 40 to 93 years of age (average 70 +/- 15 yrs) with permanent cardiac pacing or implantable cardioverter-defibrillator (ICD). Subsequent echocardiographic views were applied: an apical four chamber view, a subcostal one and a parasternal right ventricular inflow tract view. At the level of the tricuspid annulus the electrode was positioned: the anterior leaflet--41.7% (5 pts), the anteroseptal commissure 25% (3 pts), the posterior leaflet 8.3% (1 pt) and the septal one--8.3% (1 pt). In 16.7% (2 patients) the lead was positioned centrally in the right atrioventricular orifice. Regarding the further positioning of the electrode in the ventricle, in 41.7% (5 pts) the leads were placed along the interventricular septum, in 16.7% (2 pts) along the anterior wall of right ventricle and in 41.7% (5 pts) across the centre of the right ventricle. The tip of the lead was positioned in the apex of the right ventricle in 83.4% (10 pts). In the remaining 16.7% (2 pts) the position was not apical--in 1 patient the anterior wall of the right ventricle and in 1 patient the interventricular septum. In the VVI pacing mode the electrode did not lie on the interventricular septum. In contrast to this in 80% of patients (4 pts) having the DDD pacing mode the lead was situated on the interventricular septum on its course downwards to the ventricle. CONCLUSIONS: 1) On the level of the leaflets of the tricuspid valve the lead most often was positioned at the level of the anterior leaflet and the anteroseptal commissure. 2) Most patients had an apical localisation of the tip of the lead. 3) Differences between morphological and echocardiographic studies are related to the intravital and the two dimensional character of echocardiography, and probably to the small population of the group examined. PMID- 11107705 TI - Somatic and skeleton development of rat foetuses following in-utero exposure to isopropylantipyrine (propyphenazone) during the second trimester of gestation. AB - Isopropylantipyrine (IPA, propyphenazone) is a pyrazolone derivative, widely used as an antipyretic and analgesic drug. The aim of the study was to evaluate the influence of propyphenazone on rat development. IPA was administered to pregnant rats from day 8 to day 14 of pregnancy once a day, orally by a stomach tube at doses of 2.10 (R1), 21.0 (R2), and 210.0 mg/kg/day (R3). The dams were sacrificed on day 21 of gestation and corpora luteum, implants, resorptions, and live foetuses were counted. The weight of foetuses and placentas, the length of foetuses and their tails were checked. The foetuses were fixed in alcohol and skeletons were stained with alizarin. There was a statistical difference in body length in R1, R2 and numbers of subcutaneous ecchymose in R1. External and skeletal examination of the foetuses revealed no evidence of teratogenesis. It can be concluded that IPA has no harmful effects on the prenatal development of the rat offspring at doses used in the present study. PMID- 11107706 TI - The anatomy of the median branches of the basilar artery. AB - The aim of study was the examination of the median branches (MB) of the basilar artery (BA). 783 MB were found in 100 human brainstems injected with coloured latex. The attachment of the MB to the quadrants of the BA's trunk, their structural shape, length of trunks, topographical arrangement (on three vascular levels) and the clinical importance of this vascular pattern was evaluated. The MB arise from Pr and Pl quadrants of the BA's trunk in 100% cases, represent 37.7% of BA's branches and are mostly located in the foramen caecum area (56%). The MB confine the BA's stem mobility on the V CN area and above, because of their shortest trunks especially on these vascular levels. In the examined specimens the MB varied in number from 0 to 22 (mean, 8). PMID- 11107707 TI - Substance-P of neural cells in human trigeminal ganglion. AB - Expression of substance-P in human neurons of trigeminal ganglia has been investigated by immunohistochemistry and morphometry. These neurons constituted 12.8% to 32.6% of the total neuronal population in the trigeminal ganglia. Substance-P positive granulations were concentrated around the nucleus, distributed focally in neuroplasm or dispersed over the neuroplasm. Morphometric analysis has indicated the presence of three populations of SP-positive cells: small, medium-sized and large. The results suggest a functional differentiation on the level of the first neurons of the afferent path of the stomatognathic system. Substance-P is likely to play a role in the transmission not only of nociceptive impulses but also of those involved in the mechano-functional stimulation of system activities. PMID- 11107708 TI - The cytoarchitectonic and neuronal structure of the red nucleus in guinea pig: Nissl and Golgi studies. AB - The present studies were carried out on the brains of adult guinea pigs, Dunkin Hartley strain. On the basis of preparations, they were stained according to the Nissl and the Kluver-Barrera method's; a short description of the cytoarchitectonics and the characteristics of the rubral cells were written. The red nucleus (RN) of the guinea pig is 1.2 mm in length. Three cellular parts in RN, and three classes (A, B, C) of the rubral cells were distinguished. Taking into consideration the predominant cell size, RN was divided into magnocellular part (RNm), parvocellular part (RNp) and intermediate part (RNi). On the basis of Golgi impregnated preparations four neuronal types (I, II, III, IV) were distinguished. To sum up, in the guinea pig were observed: the large, mainly multipolar (type I) and bipolar (type II) spiny being coarse (class A) in Nissl material; the medium-sized, triangular, aspiny (type III) corresponding to the fine cells (class B); and the small, both spiny and aspiny neurons (type IV), which are the fine or achromatic cells (classes B or C) in Nissl stained slices. The highest degree of dendritic branching was observed in type I, whereas the lowest in cells of types III and IV. PMID- 11107709 TI - The dermatopharmacologic profile of ciclopirox 8% nail lacquer. AB - Ciclopirox 8% nail lacquer is a recently introduced topical formulation of the hydroxypyridone antimycotic ciclopirox. In vitro data indicate that ciclopirox has activity against the important pathogenic dermatophytes responsible for onychomycosis. The lacquer delivery system provides a high concentration gradient for the transfer of the antifungal agent through the nail plate. Daily application to the toenail surface of healthy subjects resulted in good penetration and distribution within all nail layers. Systemic absorption of ciclopirox in five patients with onychomycosis was minimal. PMID- 11107710 TI - Ciclopirox 8% nail lacquer in the treatment of onychomycosis of the toenails in the United States. AB - Ciclopirox 8% nail lacquer has recently become the first topical antifungal agent to be approved by the US Food and Drug Administration for the treatment of onychomycosis. This article reviews the results of the two pivotal clinical trials of this drug that have been performed in the United States as well as those that have been carried out in other countries. The two US studies were both double-blind, vehicle-controlled, parallel-group, multicenter studies designed to determine the efficacy and safety of ciclopirox nail lacquer in the treatment of mild-to-moderate onychomycosis of the toenails caused by dermatophytes. The combined results show a 34% mycologic cure rate, as compared with 10% for the placebo. Data from the ten studies conducted worldwide show a meta-analytic mean (+/- SE) mycologic cure rate of 52.6% +/- 4.2%. As expected for a topical agent, ciclopirox nail lacquer was found to be extremely safe, with mild, transient irritation at the site of application reported as the most common adverse event. Ciclopirox nail lacquer may also have potential for use in combination or adjunctive therapy. Further studies will help to better position this agent for the treatment of this widespread podiatric condition. PMID- 11107711 TI - Ciclopirox nail lacquer and podiatric practice. AB - Ciclopirox 8% nail lacquer has recently been approved by the US Food and Drug Administration (FDA) for the management of mild-to-moderate dermatophytic onychomycosis not involving the lunula. Previously, the agents that were approved for the treatment of dermatophytic pedal onychomycosis--griseofulvin, itraconazole, and terbinafine--were administered orally. When ciclopirox nail lacquer is used, it is recommended that the infected nail undergo debridement by a health-care professional as frequently as monthly. It is important to be aware of the circumstances under which debridement of the mycotic nail may be considered medically necessary and therefore potentially eligible for reimbursement by third-party payers. For many nail presentations, nail debridement is an important component of a treatment protocol involving either the oral medications or the topical lacquer, as it serves to reduce the fungal load and ameliorate symptoms. With the availability of a new FDA-approved topical treatment alternative, it remains to be seen if podiatrists will embrace the definitive treatment of onychomycosis using the newer oral agents, the new nail lacquer, or both in combination with nail debridement to treat the disease. PMID- 11107712 TI - Disseminated coccidioidomycosis masquerading as tendinitis. AB - The authors report on a case of disseminated coccidioidomycosis of the medial cuneiform. This is a rare finding, with only a few reports of Coccidioides immitis being cultured from the bones of the foot. A brief overview of coccidioidomycosis, pertinent imaging studies, and histopathologic evaluation are presented. PMID- 11107713 TI - Overweight and obesity in Mississippi: a growing problem. AB - The difficulties, complexities, hazards, and failures of treatment for overweight and obesity argue strongly for an approach that prevents excessive weight gain. The prevalence and trends data support a preventive approach beginning in childhood and adolescence but also targeting college-age students and young adults: the prevalence of overweight is already high (30%) in adolescents prevalence rates increase markedly in the college years (compare the rates in the 18-24 and 25-34 year old age groups with the rates in the older age groups) the greatest average increase in prevalence rates over the period 1990-99 has been in the 18-24 and 25-34 year old age groups (data not shown). PMID- 11107714 TI - Management of pregnancy related carpal tunnel syndrome. PMID- 11107715 TI - Chronic biscuit poisoning. PMID- 11107716 TI - Plaintiff's attorney actively seeking out patients who have used various FDA approved medications and who "may be entitled to compensation for damages caused by these medications". PMID- 11107717 TI - Updated recommendations from the Advisory Committee on Immunization Practices in response to delays in supply of influenza vaccine for the 2000-01 season. PMID- 11107718 TI - Information and quality healthcare--HCFA quality data. PMID- 11107719 TI - Same song, second verse. PMID- 11107720 TI - Looking back: a unique case of obstetrics. PMID- 11107721 TI - [Corticosteroids are efficient in treatment of pseudocroup. Meta analysis confirms Swedish therapeutic tradition]. PMID- 11107722 TI - [The SBU report on back pain and neck pain. An ambitious survey of a big problem]. PMID- 11107723 TI - [A Cochrane report on carotid surgery: early endarterectomy in carotid stenosis is recommended]. PMID- 11107724 TI - [Analgesia for infants. A review of randomized trials]. PMID- 11107725 TI - [Collected knowledge about back pain and neck pain. What we know--and what we don't know]. PMID- 11107726 TI - [Ultrasound duplex an important tool in diagnosis of vascular diseases]. AB - Ultrasound duplex scanning has caused a revolution to noninvasive diagnostics of disease in arteries and veins. A duplex examination (duplex = gemini, twin) initially implied the use of a combination of black and white 2D image and spectral Doppler, but now also includes colored directional Doppler and energy (nondirectional) Doppler. Late ultrasound developments are: low frequency (tissue) Doppler, 2nd harmonics image and doppler, contrast enhancement of blood flow, 3- and 4 dimensional reconstruction of image and flow, and compound imaging, every modality with new and improved information. It is possible to image the arterial and venous circulation in detail, including the vessel wall and the atherosclerotic atheroma, viewing both anatomical, functional, dynamic and mechanical properties. Furthermore, it is possible to use low-energy diagnostic ultrasound for intervention, for example to enhance thrombolysis. Duplex scanners now are used as the prime diagnostic tool in vascular laboratories. Even for traditionally morphological diagnostic purposes, flow information is valuable. A duplex examination is normally entirely noninvasive and nontraumatic, and thus without risks or discomfort for the patient. The technic has improved gradually and now may complement or even replace angiography and venography, for example for detection of carotid, iliac and femoral stenosis, venous insufficiency and venous thrombosis. As the examination is performed in real time, momentary flow and wall movements may be studied, for example during exercise and in drug research. PMID- 11107727 TI - [Treatment of accidental hypothermia]. AB - New knowledge about accidental hypothermia acquired in recent years may simplify treatment and aid the evaluation of prognosis. Evidence of death or severe collapse due to the feared afterdrop has not been published. Afterdrop is a phenomenon of conductive heat loss. Evaluation of rewarming techniques shows that results from forced air rewarming techniques are equivalent to or better than results from invasive rewarming methods, except for rewarming with cardiopulmonary bypass. In hypothermia the most important differential diagnosis is death. Patients who are cold and could be resuscitated must be differentiated from patients, who are cold because they are dead. Experience from abroad has shown that extreme hyperkalaemia may be a useful diagnostic tool. PMID- 11107728 TI - [Inline skating--high fracture risk. Two of three injured are boys and young men. Wrist fractures are most common]. AB - In-line skating injuries have increased in recent years. Hospital based data from Umea concerning 135 persons injured in in line skating collisions were analyzed. The highest yearly incidence of injury was found in males 10-19 years of age, at 1.7 per 1,000 inhabitants; the corresponding figure for females was 0.5. Two thirds of incidents were caused by falls due to balance problems without the influence of any "external factor" such as rough road surfaces. No collisions with motor vehicles or other road users were registered. Nearly half of the injuries were fractures or dislocation injuries, most frequently of the upper extremities. Non-minor head injuries were rare. Protective gear for wrist and elbow may have the potential to reduce these injuries. PMID- 11107729 TI - [Four perspectives on women's breasts]. PMID- 11107730 TI - [Better integrated measures required to protect society against microorganisms]. PMID- 11107731 TI - [New guidelines for hypertension should be adapted to Swedish conditions!]. PMID- 11107732 TI - [Suspended leave in forensic psychiatric care--how are the patients to be rehabilitated?]. PMID- 11107733 TI - [The National Board of Health Welfare answers: attitude is against the intentions of the psychiatric reform]. PMID- 11107734 TI - [What to tell and not to tell]. PMID- 11107735 TI - [More about the administration of Turbuhaler]. PMID- 11107736 TI - [Difficult to see advantages of the geriatric care reform]. PMID- 11107737 TI - [A Swedish webside makes the calculation of the Crohn index simpler]. PMID- 11107738 TI - [A lot of confusion around the CSF]. PMID- 11107739 TI - [A world conference on tobacco: smoking is the most dangerous threat against life and health]. PMID- 11107740 TI - [Venous thromboembolism, warfarin and cancer--a well-known connection]. PMID- 11107741 TI - [Diagnostic ultrasound--background and possibilities of development]. PMID- 11107742 TI - [Children with asthma followed up for 21 years. Reduced severity, but patients seldom grow out of asthma by adulthood]. AB - The course of asthma severity, clinical allergies, allergic sensitization, lung function, changes in living conditions and social outcome were studied prospectively in 55 asthmatic children for 21 years, from a mean age of 9 to 30 years. Asthma severity improved, but only 16 percent were in remission at the final follow-up. After adolescence, clinical improvement continued among males but not among females. Lung function showed a similar gender difference with respect to clinical course. Generally, clinical allergies and sensitization to pollens and animal danders persisted in adulthood. In adulthood, asthma severity and degree of bronchial hyperresponsiveness correlated with the extent of sensitization to furred animals. PMID- 11107743 TI - [Asthma and allergy in schools--guidelines with strong support]. PMID- 11107744 TI - [High-dose melphalan with stem cell support is now an established myeloma therapy. Treatment of myeloma from a 30-year perspective]. AB - Melphalan and prednisone have been the backbone in myeloma therapy for more than 40 years. New developments in chemotherapy and supportive therapy, achieved during the two decades which preceded the use of high-dose chemotherapy with stem cell rescue, have not changed the overall prognosis. A study of high-dose melphalan with autologous stem cell support on 274 patients < 60 years, performed by the Nordic Myeloma Study Group, has shown a prolongation of the median survival by 1.5 years. The results confirm that this therapy is a major step forward in myeloma therapy. Cost-utility and quality-of-life studies show that high-dose therapy has acceptable costs and leads to a favorable long-term quality of-life. PMID- 11107745 TI - [Myasthenia gravis--an autoimmune neuromuscular disease]. PMID- 11107746 TI - [Digital imaging system are rapidly introduced in Swedish departments of radiography. This calls for new strategic planning]. AB - Diagnostic radiology in Sweden is changing rapidly to digital (filmless) technique. The advantages are more rapid delivery of radiologic service, better working conditions and less negative effects on the environment. Teleradiology is also facilitated. The Swedish Board of Health and Welfare has investigated the speed with which this change is taking place. In 1998, 26 of the 118 departments of diagnostic radiology had already turned digital; it is estimated that in the near future at least five departments will become fully digital each year. For planning purposes, less emphasis should be put on the supply of radiographic film, and more on telecommunications, computer hardware and digital storage. PMID- 11107747 TI - [Interleukin-18 and new drugs. Protective effect against tumor growth and infections]. AB - IL-18, originally identified as interferon-gamma inducing factor (IGIF), is related to the IL-1 family in terms of its structure as well as its processing, receptor, signal transduction pathway and pro-inflammatory properties. IL-18 is also functionally related to IL-12, as it induces the production of Th1 cytokines and participates in cell-mediated immune cytotoxicity. A summary is made of recent advances in the understanding of IL-18 structure, processing, receptor expression and immunoregulatory functions. It focuses on the role of IL-18 modulation in tumours, infections and autoimmune and inflammatory diseases. PMID- 11107748 TI - ["It was up to me to prove my sickness, but how? I don't have a zipper in my neck after all". Meeting insurance authorities is often a shocking experience for the client]. PMID- 11107749 TI - [English terminology can be adapted to Swedish language]. PMID- 11107750 TI - [Meniere and lemology]. PMID- 11107751 TI - [Both children and adults suffered of syphilis]. PMID- 11107752 TI - [Views on the MFR documentation on child health services: early diagnosis and early intervention improve prognosis of children with autistic disorder]. PMID- 11107753 TI - [Gardeners and the important congress]. PMID- 11107754 TI - [Claes Sundelin answers: No evidence of results in connection with screening for autism]. PMID- 11107755 TI - [Mobile phone industry on mobile phones and safety: arbitrary safety rules must not eliminate known permissible levels]. PMID- 11107756 TI - [A reply: the safety principle should be applied]. PMID- 11107757 TI - [Cervix screening--good scientific basis]. PMID- 11107758 TI - [Localized prostatic cancer--should the patient or the physician choose the treatment?]. PMID- 11107759 TI - [Prevalence of carpal tunnel syndrome has not been changed during a 20-year period]. PMID- 11107760 TI - [COX-2 inhibitors are similar or different--a reply]. PMID- 11107761 TI - Medical errors. PMID- 11107762 TI - The morbid fingerprints of Salmonella. PMID- 11107763 TI - From Atkins to Zone: the truth about high-fat, high-protein diets for weight loss. PMID- 11107764 TI - Cardiovascular disease and nutrition. PMID- 11107765 TI - It's only natural: use of "natural" products from the clinician's perspective. PMID- 11107766 TI - The role of fiber in the diets of children. PMID- 11107767 TI - Physician-delivered nutrition counseling: why and how? PMID- 11107769 TI - Medicare's first snapshot on quality: what does it mean? PMID- 11107768 TI - Typhlitis. PMID- 11107770 TI - Overweight and obesity among Rhode Island adults. PMID- 11107771 TI - The safety of calcium fortification. PMID- 11107772 TI - Physician "de-selection:" healthcare providers' rights when terminated from managed care plans. PMID- 11107773 TI - Medical malpractice tort actions provide two benefits: compensation for the victim of medical malpractice, and a reminder to the health care profession of the legal consequences of negligence. PMID- 11107774 TI - [Nonspecific adhesion of Staphylococcus aureus strains to solid surfaces]. AB - Autoagregating strains of bacteria are characterised by high surface hydrophobicity, which determines their ability to adhesion. An assessment was done of non-specific adhesion to solid surfaces of S. aureus strains isolated from blood, pus and nasopharynx of hospitalised people. The method used made possible differentiation of strains, which were studied, on the basis of their surface characteristics. Their properties decide about the abilities of strains to the colonisation of host tissues and at the same time they influence their potential virulence. In the study attention was also paid to the participation of surface proteins in the processes of adhesion cells to glass surfaces. PMID- 11107775 TI - [Influence of culture conditions on cell surface hydrophobicity of rods of genus Serratia]. AB - The cell surface hydrophobicity (CSH) is a non-specific adhesion factor that is important in the proliferation of microorganisms on solid surfaces. Serratia spp. is a bacterium that has been increasingly implicated as a primary pathogen in numerous human infections, particularly in urinary tract infections. CSH of 60 Serratia spp. strains isolated from clinical materials was evaluated using the ammonium sulfate salt aggregation test. Bacteria were grown for 24 h and 48 h at room temperature (22 degrees C) and 37 degrees C on enrichment broth and agar (Biomed), enrichment agar with 5% human blood and medium composed of agar granulated (Becton Dickinson), neopeptone (Difco) and 1% (v/v) glycerol. CSH was estimated most frequently when the analyzed strains in enrichment broth were cultured. When grown in enrichment broth cells of Serratia spp. at room temperature were more hydrophobic (43% after 24 h and 47% after 48 h) than those at 37 degrees C (30% after 24 h and 33% after 48 h). CSH of the examined Serratia spp. strains were depended on the temperature, time of the culture of bacteria and the kind of media. The influence of the culture conditions on the changes in CSH of the analyzed bacteria may suggest significance of these properties in the pathogenesis of Serratia spp. PMID- 11107776 TI - [Occurrence of beta-lactamase type ESBL in rods of the Serratia species]. AB - Serratia spp. has been identified as an important opportunistic pathogen agent in nosocomial infections. The aim of the study was the determination of extended spectrum beta-lactamases (ESBL) occurrence among 78 of Serratia spp. strains isolated in 1996-1998 from clinical specimens obtained from patients of State Clinical Hospital in Bydgoszcz. Identification of Serratia spp. strains was performed in automatic ATB system with ID 32GN strips (bioMerieux). The strains with ESBL activity were detected by double-disc method according to Jarlier et al. (10) with small modifications. Clavulanic acid, tazobactam and sulbactam were used as the inhibitors of ESBLs. Drug-susceptibility was determined by disc diffusion method according to NCCLS standards. Forty-five (57.7%) of the strains were ESBL (+). All of them belonged to S. marcescens species. The majority--91.1% of strains was derived from urine, 3 from wound and 1 from blood. The obtained results indicate the necessity of monitoring of ESBL-producing strains among gram negative rods from clinical specimen. The aims of such a procedure are to control and to prevent their dissemination within hospital, as well as to avoid therapeutic failures. PMID- 11107777 TI - [Evaluation of the usefulness of selected virulence markers for the identification of virulent Yersinia enterocolitica strains. I. Phenotypic markders associated with Plasmid pYV]. AB - The species Yersinia enterocolitica includes either pathogenic or non-pathogenic strains. Therefore it is necessary to differentiate virulent bacilli from other. It is well known that pathogenic strains of Y. enterocolitica bearing virulence associated plasmid called pYV, which could be demonstrated by its isolation or detected by the presence of specific, phenotypic properties directly related with this plasmid. The aim of the presented paper was to check the ability of some phenotypic virulence markers associated with pYV, to detection of pathogenic Y. enterocolitica strains. In the presented work 152 (130 carrying pYV) clinical strains of Y. enterocolitica O3 isolated mainly from stool were examined for the presence of phenotypic virulence markers such as: calcium dependency, Congo-red binding, autoagglutination and agglutination with Mangifera indica extract. Both first features were detected parallel, on the same plate, using CRMOX (Congo-red, Magnesium Oxalate) agar. The detection of the tested markers in the examined strains was compared with the presence of virulence plasmid. The obtained results confirmed the observations done by other authors that Y. enterocolitica strains, in which bacilli bearing the virulence plasmid predominate, exhibit all tested phenotypic properties whereas the plasmid-cured isogenic strains show no one of these features. Therefore all the tested markers could be useful for detection of virulent Y. enterocolitica strains directly isolated from patients. The most useful virulence markers in bacteriological study seems to be calcium dependency and Congo-red binding, examined together by the use of CRMOX agar, because they confirm the presence of the virulence plasmid by parallel detection of two physiologically different features associated with this plasmid. In addition CRMOX agar allows for the examination rough strains while agglutination tests do not. PMID- 11107778 TI - [Evaluation of the usefulness of selected virulence markers for identification of virulent Yersinia enterocolitica strains. II. Genotypic markers associated with the pYV plasmid]. AB - Pathogenic strains of Yersinia enterocolitica bear virulence associated plasmid pYV. Unfortunately plasmid pYV is easily lost by these bacteria incubated at elevated temperatures (37 degrees C) or long stored at room temperatures. This sometimes makes difficult the detection of the virulence plasmid, especially by its isolation or biochemical tests. On the other hand, observations done by some authors suggest that polymerase chain reaction (PCR) could be useful for demonstration of the pYV plasmid of Yersinia strains. Accordingly to this observation the aim of the presented study was to check the usefulness of plasmid localised genes virF and yadA, detected by PCR, for the identification of the virulent strains of Y. enterocolitica. In the presented study one hundred and fifty two clinical strains of Y. enterocolitica belonging to serogroup O3 were investigated by the PCR for the presence of genes virF and yadA. Bacterial strains were first tested for the presence of pYV plasmid. In addition the phenotypic features: calcium dependence, Congo red binding and autoagglutination were determined. In this way the virulence plasmid was found in 130 of 152 examined strains. For PCR studies also forty plasmid-cured strains of Y. enterocolitica and 32 non-Y. enterocolitica, Enterobacteriaceae strains were included. The obtained results show that the tested genes were present only in Yersinia strains possessing the pYV plasmid and no one non-specific PCR product was observed. The detection level of these genes in nested PCR permits to detect pathogenic Y. enterocolitica in suspension composed of 1 x 10(3) CFU/ml of pYV+ bacilli and 3 x 10(9) CFU/ml plasmid-cured, isogenic bacteria. In the study it was shown that genes virF and yadA were useful virulence markers, which could be helpful in clinical studies for the detection of the virulence plasmid in Y. enterocolitica strains long stored or incubated at elevated temperatures. PMID- 11107779 TI - [Evaluation of the usefulness of selected virulence markers for identification of virulent strains of Yersinia enterocolitica strains. III. Chromosome markers of virulence]. AB - It is well known that virulent strains of Y. enterocolitica bear the virulence associated plasmid pYV. Moreover some authors consider that the pathogenic strains of these bacteria have chromosome encoded phenotypic and genotypic features such as: genes ail and yst which could be used as virulence markers. The virulent strains of Y. enterocolitica do not produce pyrasinamidase and are not able to ferment salicin and cannot hydrolyse esculin. In addition these strains produce thermostable enterotoxin called YST and protein Ail (attachment invasion locus). In contrast to phenotypic virulence makers the biological function of proteins Ail, YST and nucleotide sequence of genes ail and yst is well described. In the presented study one hundred thirty virulence plasmid bearing Y. enterocolitica strains belonging to serogroup O3 were examined for the presence of genes ail, yst, and were tested for their inability to pyrasinamidase production, salicin fermentation and esculin hydrolysis. In addition forty pYV plasmid-cured isogenic strains were included in to the study. Genes ail and yst were detected by polymerase chain reaction (PCR). The obtained results indicate that all tested 130 pYV+ Y. enterocolitica strains as well as 40 plasmid-cured isogenic strains have carried ail and yst genes. All tested strains did not produce pyrasinamidase, hydrolyse esculin and ferment salicin. This generally was in agreement with the observations done by other authors and suggest that the chromosomal virulence markers, especially well described genes ail and yst, could be useful for excluding the potential virulence of Y. enterocolitica strains, which had lost pYV plasmid and have no ail or yst genes. Therefore, in clinical studies, Y. enterocolitica strains isolated directly from patients should be primarily tested for the presence of the virulence plasmid and secondarily, the negative ones could be examined for the presence of the chromosomal virulence markers. PMID- 11107780 TI - [Use of PCR methods for detection of selected genes of Mycoplasma pneumoniae]. AB - The aim of this study was standardization of PCR for the detection of gene encoding the P1 protein, 16S rRNA and elongation factor Tu of M. pneumoniae. A total of 13 strains of M. pneumoniae, 28 strains of other mycoplasmas and 14 strains of different bacteria causing respiratory tract infections were tested. In all of tested M. pneumoniae strains the presence of the sought genes was confirmed. The specificity of DNA was confirmed by the restriction endonuclease analysis with enzymes Hind III, Alu I and Hha I. With none of primers specific for the M. pneumoniae genes amplification of DNA from other bacteria was noted. The PCR method with the selected primers allowed to detect from 10(2) to 10(4) cfu M. pneumoniae/ml suspended in broth. The obtained results indicate that the PCR method can be used for detection of M. pneumoniae genes. A very good sensitivity and specificity predestine++ PCR as a potential quick diagnostic method for identification of M. pneumoniae in clinical specimens. PMID- 11107781 TI - [Use of the toxin-binding inhibition test for estimation of tetanus antitoxin in serum]. AB - The usefulness of the toxin binding inhibition test (ToBI) for the titration of tetanus antibody in human sera was assessed. Sera from 80 healthy people with different vaccination histories that had been previously tested by the in vivo toxin neutralization (TN) test were retested by the ToBI test. The lowest tetanus antibody titre which could be detected by using 0.1 Lf/ml tetanus toxin was 0.01 IU/ml. Comparison between the estimates obtained by the ToBI test and those obtained by the TN test (r = 0.93 and r = 0.7 for titration low titre sera) showed good correlation. No overestimation of antibody content was seen in titrating low titre sera by the ToBI test. It is concluded that the ToBI-test is a reliable alternative to toxin neutralization test in mice. PMID- 11107782 TI - [Immunogenic activity of temperature sensitive mutation of herpes simplex virus type-1 in mono- and polyvalent conditions with different attenuated strains of virus. I. Analysis of humoral immunity induced by mutant temperature sensitive herpes simplex virus type-1 in monovalent systems]. AB - The aim of the present study was the analysis of the immunogenic activity of temperature sensitive mutant of HSV-1 with significantly decreased pathogenicity for mice. The stability of immunogenic potency was also tested. The level of antibody-mediated response in guinea pigs after two doses of mutant ts HSV-1 or native strain was compared. Antibodies to HSV were measured using ELISA and CFM. Seroconversion rates in tested groups were high, from 80% to 100%. The level of antibody response at 4 week after inoculation measured by ELISA and CFM showed no significant differences between 4 passages of mutant ts HSV-1. Significant higher increase of antibodies level after two doses of viruses was observed for mutant ts HSV-1 as compared to native strain, both in ELISA and CFM. PMID- 11107783 TI - [Immunogenic activity of temperature sensitive mutation of herpes simplex virus type-1 in mono- and polyvalent conditions with different attenuated strains of virus. II. Analysis of humoral immunologic response induced by mutants of temperature sensitive herpes simplex type-1 virus in polyvalent systems]. AB - The ability of mutant ts HSV-1 on 45 passage to induction of IgG antibody response in the presence of other attenuated viral strain, like measles, mumps, rubella was evaluated, and the effect of mutant ts HSV-1 on humoral immunity induced by attenuated measles and mumps strains was also tested. The obtained results clearly indicated that even immunization with other RNA virus strains had no effect on IgG antibody response to mutant ts HSV-1. Significant differences were observed in IgG antibody responses to mumps and measles between groups of guinea pigs immunized with these viruses alone or together with other viruses. This was especially observed in the case of mumps virus. It clearly shows that change of virus concentration rates in polyvalent schema of immunization is needed. PMID- 11107784 TI - [Susceptibility of staphylococci to natural and synthetic iron chelators]. AB - A total of 237 strains of staphylococci belonging to 27 species were tested for susceptibility to natural and synthetic iron-chelators. Coagulase-negative staphylococci were much more susceptible than coagulase-positive ones: 82 out of 148 coagulase-negative strains were susceptible to desferrioxamine B, rhodotorulic acid and ovotransferrin and 7 out of 89 coagulase-positive strains. A correlation between the susceptibility to iron-chelators species affiliation and the origin of strain was not found. PMID- 11107785 TI - [Restriction analysis of Bordetella pertussis DNA isolated from patients with whooping cough in 1968 and 1995-98 and B. pertussis used for production of national vaccine strains]. AB - The genotypic and serotypic analysis of B. pertussis strains isolated from the nasopharynx of children with whooping cough in the years 1968 and 1995-98 and B. pertussis vaccine strains was the aim of this study. The genotyping of the examined strains was done by electrophoretic division of DNA in pulsed field. The 3 types (A, B, C) and 2 subtypes (A1 and A2) of DNA restriction patterns were determined for the B. pertussis strains isolated in 1968. The 2 types (D and E) and 10 subtypes (D1-D10) of DNA restriction patterns were identified for B. pertussis strains from the years 1995-98. The DNA restriction patterns of B. pertussis strains isolated in the years 1968 and 1995-98 were not identical what was the evidence of the fact that in the sixties and nineties whooping cough was caused by different B. pertussis clones. The different DNA profiles were also observed for vaccine strains as well as for vaccine strains and current isolates. Differences in DNA patterns of vaccine strains and B. pertussis strains isolated in the years 1995-98 indicated a relationship possibility in some cases while lack of relationship between these strains in other cases. Serotyping of the examined B. pertussis strains was performed by the agglutination method with the sera against B. pertussis agglutinogens 1, 2 and 3. Most strains--15 (75%) isolated in 1968 possessed only agglutinogens 1 and 3. Serotype 1, 2, 3 was most frequently observed among isolates from the years 1995-98. This study indicates the expediency of periodical change of B. pertussis vaccine strains in the aspect of whooping cough resurgence in the years 1994-95 and 1997-98. PMID- 11107786 TI - [Infectivity and resistance to antibiotics of uropathogenic gram-negative rods]. AB - The nature of bacterial isolates from children with clinically suspected urinary tract infections (UTI) was studied. The susceptibility of urinary pathogens to selected antibiotics was determined. The results clearly show that E. coli was identified as the main causative agent of UTI children (67% of isolates). The second commonest pathogen was P. mirabilis (10%). Over half E. coli isolates were resistant to amino-penicilins but almost all isolates (over 80%) were sensitive to antimicrobial agents combined with beta-lactamase inhibitors. We found significantly high percentage (32.5%) of ESBL strains among K. pneumoniae isolates. PMID- 11107787 TI - [Beta-lactamases with a wide substrate spectrum in gram negative strictly anaerobic rods]. AB - This study was performed to determine the susceptibility of the clinical strains of Gram-negative strictly anaerobic rods to newer beta-lactam antibiotics. Also, the trial was undertaken to detect strains producing extended-spectrum beta lactamases (ESBLs) and inducible beta-lactamases (IBLs) among Bacteroides spp. and Prevotella spp. rods isolated from hospitalized patients. One hundred strains of Gram-negative, obligatory anaerobic rods were applied in the study. The strains were identified in automatic ATB system using API 20 A strips. beta lactamase-positive strains were determined with disc nitrocefin test. ESBL producing strains were detected with double disc test according to Jarlier et al. (1988). Clavulanate was applied as the inhibitor of these beta-lactamases (AMO/CLAV disc). ESBL-positive strains were confirmed with the use of E test (TZ/TZL strip). Inducible beta-lactamases were determined by double disc method according to Sanders and Sanders (1979). Cefoxitin was the inducer of these beta lactamases (FOX disc). Among 93 Bacteroides spp. strains and 7 Prevotella spp. strains, 91 strains (91%) produced beta-lactamases. Two ESBL-producing strains (2%) were detected. Strains producing inducible beta-lactamases (IBL) were not found. A high activity of the examined beta-lactam antibiotics against strains of Gram-negative anaerobes was found. The majority of strains were susceptible to piperacillin (95%), piperacillin combined with tazobactam (99%), ticarcillin combined with clavulanic acid (99%), meropenem (97%) and imipenem (99%). The obtained results indicate the necessity of ESBL determination among strains of the genus Bacteroides, isolated from clinical specimens. Newer beta-lactam antibiotics, especially penicillins in combination with beta-lactamase inhibitors and carbapenems, are useful in empiric therapy of infections caused by Bacteroides spp. and Prevotella spp. anaerobic rods. PMID- 11107788 TI - [Evaluation of cytotoxic activity of Vibrio cholerae non-01 culture filtrate on established cell lines and human diploid cells]. AB - The cell-destroying effect of cell free filtrates of 90 V. cholerae non-01 cultures was measured by titration method in 3 established cell lines: CHO, HeLa and Vero and in 3 human diploid cells cultures: MRC-5, WI-38 and PZ. The vibrio strains differed in the titre of toxic effect. Most sensitive was CHO cell line, least sensitive were human diploid cell cultures. It was found that bacterial strains produced different substances toxic for various cell lines. Among them NAG-ST toxin produced by 41% of examined strains was identified and hemolysins/cytolysins activity was evaluated. Both may play a role in the pathogenicity of those strains for humans. PMID- 11107789 TI - [Evaluation of the usefulness for the latex agglutination test for serodiagnosis of Mycoplasma pneumoniae infections]. AB - The usefulness of latex agglutination test prepared in our laboratory for the diagnosis of M. pneumoniae infections was assessed. A total of 628 serum samples obtained from patients with respiratory tract infections were tested by complement fixation test and by latex test, from among them 274 serum samples were additionally tested by ELISA--Ig A/--IgG/--IgM and by immunoelectroprecipitation test. The highest sensitivity and specificity was displayed by the latex test in relation to ELISA when determining mycoplasmal antibodies of IgM class (respectively 82.1% and 89.6%) and to the complement fixation test (81.0% and 89.0%). Positive latex test results in our investigations were associated only with the presence of IgM antibodies and were not dependent on the IgA and IgG antibody classes. The latex agglutination test may be used in routine serodiagnosis of mycoplasmosis under condition that the results obtained in this test will be confirmed by the complement fixation test or ELISA. PMID- 11107790 TI - [Bactericidal properties of neutrophils from peripheral blood (PMN)of patients with Lyme borreliosis]. AB - In recent years in Poland, the interest has increased in studies about tick borne diseases, mainly Lyme borreliosis. Immune response and genotype of pathogen play an important role in the course of this disease. Phagocytic cells, especially PMN are dominant in defence mechanisms against bacterial infections. The main feature of PMN is their ability to destroy pathogenic microorganisms by phagocytosis. The aim of this study was to estimate the phagocytic activity of PMN connected with intracellular respiratory burst in patients with Lyme borreliosis. The PMN activity tests completed were: phagocytosis, spontaneous and reduced of nitrotetralizate blue test (NBT). Decreased phagocytic activity and oxygen metabolism of PMN from patients with borreliosis in comparison with values of controls were found. Normalization of these parameters after treatment was observed. Changed phagocytic activity connected with intracellular oxygen metabolism during the course of therapy was the main observation. Depression of phagocytic activity of PMN connected with oxygen metabolism can influence defence reactions in patients with Lyme borreliosis. It is suggested that changes observed are acquired and associated with Borrelia burgdorferi presence. PMID- 11107791 TI - [Phylogenetic relationship of street rabies virus strains and their antigenic reactivity with antibodies induced by vaccine strains. I. Analysis of phylogenetic relationship of street rabies virus strains isolated in Poland]. AB - The aims of these studies were: genetic characteristic of street rabies virus strains isolated from different animal species in Poland and determination of phylogenetic relationships to reference laboratory strains of the street rabies viruses belonging to genotype 1 and 5. The variability of rabies isolates and their phylogenetic relationship were studied by comparing the nucleotide sequence of the virus genome fragment. The Polish strains of genotype 1 belong to four phylogenetic groups (NE, CE, NEE, EE) corresponding to four variants: fox-racoon dog (F-RD); European fox 1 (F1); European fox 2 (F2) and European fox 3 (F3). On the Polish territories there are no rabies strains representing the variant dog wolf and typical for arctic fox variant. The similarity of nucleotide and amino acid sequences of street rabies strains belonging to genotype 1 and laboratory strain CVS is very high. It is about 91% similarity at nucleotide level and 95% at amino acid level. Rabies strain CVS is similar to genotype 5 bat strains (EBL 1) only in about 69% and 74% at nucleotide and amino acid level, respectively. The genetic divergence of rabies strains circulating in Poland raised the need of permanent epidemiological and virological surveillance. The genotype and variant of isolated strains should be determined (using PCR and RLFP methods). PMID- 11107792 TI - [Phylogenetic relationship of street rabies virus strains and their antigenic reactivity with antibodies induced by vaccine strains. II. Correlation between genetic distance and antibody reactivity]. AB - The aim of these studies was the estimation of the influence of genetic divergence of reactivity with sera of people vaccinated against rabies of Polish rabies strains. Genetic similarity between CVS strain and street rabies strains of genotype 1 is relatively high. However, CVS strain showed the highest reactivity with standard immunoglobulin and sera of antirabies vaccinated people (measured by western blot method). It was completely different from street viruses. Cluster method based on genetic and serologic features indicated high difference between CVS strain and street rabies strains belonging to genotype 1 and genotype 5. On this basis CVS strain was classified as a separate cluster. The genetic divergence of rabies strains circulating in Poland suggests the need of permanent epidemiological and virological surveillance. Strains different in their genotypic and biotypic characteristics should be estimated according to their antigenic similarity to vaccine strain. In practice neutralisation test using mono- and polyclonal sera should be performed. Strains isolated from new or atypical animal species should be studied first of all. PMID- 11107793 TI - [Proportion of fungi among isolated microorganisms from blood of patients treated in the teaching departments of The University Hospital in Cracow in 1993-1998]. AB - The study was carried out of 9742 blood cultures obtained in 1993-1998 from patients of selected departments of the University Hospital. The received samples were assessed from the standpoint of the participation of fungi strains of clinical importance as aetiological factors in systemic infections and the resistance status of a selected group of pathogens. Microbiological blood studies were conducted in the system of continuous monitoring of the obtained cultures by means of the automatic ATB system using a commercially available ID 32C set. ATB Fungus tests were used for drug resistance assessment. In all, 95 strains of yeasts (5.1%) were obtained in cultures, and an increasing variety of these pathogenic species was noted in sepsis. A systematic reduction was noted in the proportion of C. albicans and a steep rise of C. parapsilosis were observed among the aetiological factors of generalized nosocomial fungal infections. A tendency was noted for a continuous rise in the frequency of invasive candidemias with a most significant rise in their proportion in patients in general surgery and haematology departments. Among the fungi isolated from septic complications the proportion of strains resistant to antifungal drugs has been rising. PMID- 11107794 TI - [Henry Mosing (1910-1999]. PMID- 11107795 TI - [New therapy for the most severe rheumatism. Healing in sight]. PMID- 11107796 TI - [TNF-alpha blocker in rheumatoid arthritis. Who should be treated with the new substances?]. PMID- 11107797 TI - [After the HOPE Study. ACE inhibitor now for every diabetic patient?. Interview by Dr. Dirk Einecke]. PMID- 11107798 TI - [Chlamydia and coronary heart diseases--no acquittal. The puzzle fits increasingly better]. PMID- 11107799 TI - [Too much, too many fats, too sweet. Raised for obesity]. PMID- 11107800 TI - [Overview of drug interactions. Combining drugs correctly!]. AB - Drug interactions may be of two basic types: pharmacodynamic and pharmacokinetic. Pharmacodynamic interactions may be direct, in that two drugs intervene synergistically or antagonistically in the same biological system, or indirect, meaning that one drug may modify a parameter of central importance to a system, which in turn may influence the activity of the second drug. Of the two possibilities pharmacokinetic interactions are the more difficult to recognize since they have nothing to do with the biological action proper. Here, the absorption and metabolism of the drug are the most important factors. A compounding factor is the fact that the activity of a drug may show marked individual, genetically determined, differences. PMID- 11107801 TI - [Influenza and pneumococcal pneumonia. High time for vaccination]. PMID- 11107802 TI - [Bach flower therapy. What is the value of a water-brandy mixture?]. PMID- 11107803 TI - [Diabetic foot syndrome, 6. Diagnosis and treatment of mycoses]. PMID- 11107804 TI - [Dehydration in geriatric patients. Fluid substitution--also subcutaneous!]. AB - Signs of exsiccation are to be found in almost every fourth acutely ill patient admitted to the geriatric department. The major clinical signs are those associated with reduced general condition, together with somnolence, possibly agitation, oliguria, dry skin, orthostatic hypotension, and a tendency to be bedridden. In such cases, subcutaneous infusion is a simple-to-apply and for the patient stress-free, largely pain-free possibility for fluid replacement. Properly applied and with account being taken of the contraindications--in particular emergency situations, decompensated cardiac insufficiency and severe coagulation disturbances--the risks of s.c. infusion are negligible. PMID- 11107805 TI - [Diagnostic quiz. Dangerous infiltration treatment. Iatrogenic mantel pneumothorax bilaterally]. PMID- 11107806 TI - [Absolution for Schottdorf--scolding for health insurance. Are reimbursement tricks now legal?]. PMID- 11107807 TI - [Only what happens in your own practice counts. No fear of collective regress]. PMID- 11107808 TI - [This expert assessment also impacts the future of your practice. Do we have too much, too little or the wrong medicine?]. PMID- 11107809 TI - [How long must we still sacrifice ourselves? Up to 20% of our performance is not reimbursed]. PMID- 11107810 TI - [Acute seasonal respiratory tract infections. Even legionellas participate significantly]. PMID- 11107811 TI - [Rheumatism therapy, from the economic viewpoint. COX-2 inhibitor as cost saving drug]. PMID- 11107812 TI - [Recurrent colpitis. A vaccine could bring relief]. PMID- 11107813 TI - [News from transplantation medicine. Less rejection thanks to new monitoring]. PMID- 11107814 TI - [Chronic hepatitis B. Is long-term healing possible?]. PMID- 11107815 TI - [Therapy of type 2 diabetes. Blood lipids should not be neglected!]. PMID- 11107816 TI - [Blood pressure regulation. Also before waking, no treatment failures]. PMID- 11107817 TI - [Proper vaccination in general practice]. PMID- 11107818 TI - [Cell culture for the sick liver. Will cirrhosis soon be curable?. Interview by Petra Eiden]. PMID- 11107819 TI - [Chronic inflammatory bowel diseases. No fear with powerful preventive drugs]. PMID- 11107820 TI - [Alcohol and the pancreas. What the liver tolerates is already too much for the pancreas]. PMID- 11107821 TI - [Peptic ulcer hemorrhage: i.v. antacid prevents recurrence]. PMID- 11107822 TI - [Central or vestibular vertigo? Diagnostic look through Frenzel glasses]. AB - Vertigo is not a uniform symptom. The basis for a differentiated diagnosis is the history, which should record the frequency and duration of attacks. Further diagnostic investigations serve to differentiate between non-vestibular and vestibular (peripheral) forms of vertigo. Of essential importance is the determination of nystagmus with the aid of Frenzel lenses, with distinction being made between voluntary nystagmus and provoked nystagmus. These orientating examinations, which can be carried out by the family doctor, set the points for the further course. On account of the need for special equipment, such studies as the caloric tests, the swivel chair test, optokinetic tests or the tilting stage test, remain the domain of the specialist. They enable a definitive differentiation of an ENT illness from a neurological disorder. PMID- 11107823 TI - [Special forms of vertigo. Which therapy for which type of vertigo?]. AB - Paroxysmal vertigo, permanent vertigo, positional and postural vertigo, and transient vertigo are special forms, which are differentiated by their temporal course. Paroxysmal vertigo, together with hearing impairment and noises in the ear, is typical of Meniere's disease. Persistent vertigo often occurs after the loss of a peripheral vestibular end organ (e.g. after trauma of infection). In the case of positional and postural vertigo, a differentiation must be made between benign paroxysmal positional vertigo due to "wandering" otoliths, and orthostatic vertigo, which occurs on changing rapidly from a lying to a sitting position. The diagnosis is verified by Epley or Semont positional maneuvers. Confirmation of a cervical vertigo is provided by the de Kleijn and Nieuwenhuyse test. Brief episodes of vertigo (transient vertigo) occur after transient ischemia, for example when craning to look at a high building. In the acute stage, treatment of vestibular vertigo consists in the damping of the threshold for vestibular stimuli with antiemetics. For long-term treatment, antihistaminics and histamine-like substances have proven of value. PMID- 11107824 TI - [Hypertension in obesity. Weight reduction, then blood pressure returns to normal!. Interview by Eckhard Bottcher-Buhler]. PMID- 11107825 TI - [Nail-patella syndrome. Possible kidney failure as a complication]. PMID- 11107826 TI - [Mistletoe therapy. An alternative cancer treatment]. PMID- 11107827 TI - [Dermatologic manifestations of internal diseases]. PMID- 11107828 TI - [Diagnostic quiz. Chronic diarrhea with normal colonoscopy findings. Sprue]. PMID- 11107829 TI - [Budget-free zone. How much money are you ignoring?]. PMID- 11107831 TI - [Underestimated risk. Postprandial hyperglycemia induces early vascular damage]. PMID- 11107830 TI - [Botulinum toxin A in esthetic medicine. Poison for facial lines and crows feet]. PMID- 11107833 TI - [Anatomy in wax. An exhibit in Bonn of anatomic models of the "La Specola" collection]. PMID- 11107832 TI - [Complicated therapy of hypertension. The higher the risk, the lower the target blood pressure]. PMID- 11107834 TI - Sterility assurance and probability and steam quality assessment and HTM-2010. PMID- 11107835 TI - Steam sterilization and steam quality. PMID- 11107836 TI - Albumin as a carrier system for delivering drugs to solid tumors. PMID- 11107837 TI - Development of a dye ingress method to assess container-closure integrity: correlation to microbial ingress. AB - To demonstrate maintenance of parenteral product sterility, container-closure integrity over the shelf life of the product is critical. In the past, sterility testing has been used to ensure closure integrity. However, because of the limitations associated with sterility testing, there is a need for an improved method for evaluating container-closure integrity. This article describes the development of a physical test method (dye ingress) for the evaluation of container-closure integrity. FD&C Red No. 40 dye was used in dye ingress studies. The dye solution visual detection limit was similar to the spectophotometric detection limit. This limit was approximately 0.0025 microL of dye/mL, which corresponds to an absorbance of approximately 0.002 absorbance units at 506 nm. Breached vials with various sizes of microtubes were utilized to correlate the dye ingress method with a microbial ingress method. The inner diameter of the microtubes ranged from 2 to 75 microns. The dye ingress and microbial ingress methods had similar sensitivity to breached vials. One advantage of the dye method over microbial ingress is that it may be utilized with vials containing formulations that are cidal or static to microbes. Thus, the dye ingress method is considered an excellent test method for evaluating container-closure integrity. PMID- 11107838 TI - Organic solvents for pharmaceutical parenterals and embolic liquids: a review of toxicity data. AB - Non-aqueous solvents have long been used in subcutaneous or intramuscular pharmaceutical formulations to dissolve water-insoluble drugs. In recent years, the need for these vehicles was increased since the drug discovery process has yielded many poorly water-soluble drugs. Besides, preparations containing embolic materials dissolved in undiluted non-aqueous water-miscible solvents have been proposed for the intravascular treatment of aneurysms, arteriovenous malformations, or tumors. These organic solvents, regarded as chemically and biologically inert, may show pharmacological and toxicological effects. Therefore, knowledge of tolerance and activity of non-aqueous solvents is essential before they can be administered, especially when given undiluted. This paper focuses on thirteen organic solvents reported as possible vehicles for injectable products and details toxicological data when they have been administered intravascularly. These solvents can be subdivided into three groups according to their description in the literature either for intravenous pharmaceutical parenterals or for intravascular embolic liquids: well-documented organic solvents (propylene glycol, polyethylene glycols, ethanol), solvents described in specific applications (dimethyl sulfoxide, N-methyl-2-pyrrolidone, glycofurol, Solketal, glycerol formal, acetone), and solvents not reported in intravascular applications but potentially useful (tetrahydrofurfuryl alcohol, diglyme, dimethyl isosorbide, ethyl lactate). This review of the literature shows that toxicity data on intravascular organic solvents are insufficient because they concern solvents diluted with water and because of the lack of comparative evaluation using the same methodologies. PMID- 11107839 TI - Alternative microbial testing: a novel DNA-based detection system for specified microorganisms in pharmaceutical preparations. AB - Fluorescence-coupled PCR technology was employed to quantify DNA segments specific for Staphylococcus aureus, Pseudomonas aeruginosa, and Enterobacteriaceae. The PCR procedure is put forward as an alternative method for detecting microbial contaminations in pharmaceutical preparations and is compared to the tests for specified microorganisms described in European Pharmacopoeia (EP) 2, 2.6.13 and the USP, chapter 61. Data presented here describe the validation of this analytical method when used for proof of absence of specified microorganisms. The detection systems were specific for the microorganisms analyzed, and led to linear results over a wide range (more than 6-7 log intervals). The correlation coefficients lay above 0.99. The precision of replicate determinations within a single test was observed to be high, the relative standard deviation being between 0.39% and 1.53%. The precision between different tests was also high, with a relative standard deviation between 0.76% and 1.91%. The sensitivity without pre-enrichment amounted to 1-10 CFU. Since determination of the specified bacteria was performed following pre-enrichment, the limit of detection amounted to 1 CFU. Equivalent results were obtained in a study on nine batches of a milky hydrophilic cream (SH-No. M 440 A) with the conventional test for microbial contamination and the PCR procedure. The data presented here strongly indicate that the use of fluorescence-coupled PCR techniques can prove the absence of specified bacteria faster and more efficiently than conventional methods. PMID- 11107840 TI - A comparison of dry powder inhaler dose delivery characteristics using a power criterion. AB - A novel method is described that compares the in vitro drug delivery characteristics of different dry powder inhalers (DPIs) based on the power supplied by the air flow. Power is defined as the rate at which work is done by the air flow and is equal to the product of pressure gradient and air flow rate through the inhaler. Tests were conducted to compare the emitted dose and fine particle fraction of the total dose (FPFTD) of the Diskus, Diskhaler, and a proprietary powder dispersion device (PDD) containing fluticasone propionate/lactose powder blends. Three power levels were selected: 1, 2, and 3 W, representing a range of inspiratory efforts. The Diskus and PDD were independent of power, delivering consistent doses and fine particle fractions over the range of powers evaluated. Diskhaler was dependent on power, the emitted dose and FPFTD significantly decreasing at the lower power level (1W) (p < 0.005). Diskus and Diskhaler are trademarks of the Glaxo Wellcome group of companies. PMID- 11107841 TI - Absolute or sterilizing grade filtration--what is required? PMID- 11107842 TI - Mechanism of DNA damage photosensitized by trisbipyrazyl ruthenium complex. Unusual role of Cu/Zn superoxide dismutase. AB - Trisbipyrazyl ruthenium(II) (Ru[bpz]3(2+)) was examined as DNA photosensitizer. Damage resulting from the photolysis of synthetic oligonucleotides has been monitored by polyacrylamide gel electrophoresis. Photoadduct formation is found on both single- and double-stranded oligonucleotides. On oligonucleotide duplex, oxidative damage occurs selectively at the 5'G of the 5'GG3' site and to a lesser extent at the 5'G of a GA sequence. These findings suggest the involvement of electron transfer and show that this mechanism is the main DNA damaging process involved in Ru(bpz)3(2+) photosensitization. In addition, photoadducts and oxidative damage are both highly affected by an increase of salt concentration in the reaction medium, stressing the importance of direct interactions between nucleic acid bases and the excited ruthenium complex for efficient electron transfer. On single-stranded oligonucleotides, all the guanines are oxidized to the same extent. In this case, oxidative damage, which is not affected by an increase of salt in the solution, has been attributed, in part, to singlet oxygen. More importantly, Cu/Zn superoxide dismutase (SOD) strongly enhances the yield of all damage, correlated to an increase of both electron transfer and singlet oxygen production. This original activity of SOD is the first example of bioactivation of a polyazaaromatic ruthenium complex. PMID- 11107843 TI - Time-resolved thermodynamic changes photoinduced in 5,12-trans-locked bacteriorhodopsin. Evidence that retinal isomerization is required for protein activation. AB - Structural volume changes upon excitation of isomerization-blocked 5,12-trans locked bacteriorhodopsin (bR) (bacterio-opsin + 5-12-trans-locked retinal) were studied using photothermal methods. The very small prompt expansion detected using laser-induced optoacoustics (0.3 mL/mol of absorbed photons) is assigned to a charge reorganization in the chromophore protein pocket concomitant with the formation of the intermediate T5.12. The subsequent contraction associated with a 300 ns lifetime is assigned to protein movements required to reach the entire chromoprotein free energy minimum, after the 17 ps optical decay of T5.12. The volume changes comprise the entropy of medium rearrangement during T5.12 formation and decay. The slow changes detected in previous studies by atomic force microscopy might be explained by the slowing down of movements in films containing 5,12-trans-locked bR. Photothermal beam deflection data with the 5,12 trans-locked bR suspensions indicate no further changes in microseconds to hundreds of milliseconds. Thus, all the absorbed energy is either released to the solution as heat or used for entropy changes within the first 300 ns after the pulse, supporting the paradigm that isomerization is required for signal transduction in retinal proteins. Bacterio-opsin assembled with all-trans-retinal afforded (similar to data reported with wild-type bR) an expansion of 2.6 mL/mol (assigned to the production of KE) followed by a further expansion of 0.8 mL/mol (KE-->KL; KE, KL, early and late K's) involving no heat loss. For KL decay to L, a contraction of 6 mL/mol of phototransformed reconstituted all-trans bR was determined. PMID- 11107844 TI - Driving force and electronic coupling effects on photoinduced electron transfer in a fullerene-based molecular triad AB - Tuning thermodynamic driving force and electronic coupling through structural modifications of a carotene (C) porphyrin (P) fullerene (C60) molecular triad has permitted control of five electron and energy transfer rate constants and two excited state lifetimes in order to prepare a high-energy charge-separated state by photoinduced electron transfer with a quantum yield of essentially unity (> or = 96%). Excitation of the porphyrin moiety of C-P-C60 is followed by a combination of photoinduced electron transfer to give C-P(.+)-C60.- and singlet singlet energy transfer to yield C-P-1C60. The fullerene excited state accepts an electron from the porphyrin to also generate C-P(.+)-C60.-. Overall, this initial state is formed with a quantum yield of 0.97. Charge shift from the carotenoid to yield C(.+)-P-C60.- is at least 60 times faster than recombination of C-P(.+) C60.-, leading to the overall quantum yield near unity for the final state. Formation of a similar charge-separate species from the zinc analog of the triad with a yield of 40% is also observed. Charge recombination of C(.+)-P-C60.- in 2 methyltetrahydrofuran yields the carotenoid triplet state, rather than the ground state. Comparison of the results for this triad with those for related triads with different structural features provides information concerning the effects of driving force and electronic coupling on each of the electron transfer steps. PMID- 11107845 TI - Effect of pH on the fluorescence and absorption spectra of hypericin in reverse micelles. AB - The well-characterized, monodispersed nature of reverse micelles formed by sodium bis(2-ethylhexyl)sulfosuccinate/heptane and their usefulness in approximating a membrane-like environment have been exploited to investigate the effect of pH and water pool size on the photophysical properties of hypericin (Hyp). Our measurements reveal two titratable groups of pKa approximately 1.5 and approximately 12.5. These are assigned to the HypH+/Hyp equilibrium (the deprotonation of a carbonyl group) and the Hyp-/Hyp2- equilibrium (the deprotonation of a peri hydroxyl group). The low-energy absorbance maxima of HypH+, of Hyp and Hyp- and of Hyp2- are 583, 594 and 613 nm, respectively. Neither at pH 13 nor at 1 M HCl is the system entirely in the Hyp2- or the HypH+ forms. Ours is the first study of Hyp in reverse micelles as well as the first time-resolved study of Hyp as a function of pH. PMID- 11107846 TI - N-hydroxy-4-(4-chlorophenyl)thiazole-2(3H)-thione as a photochemical hydroxyl radical source: photochemistry and oxidative damage of DNA (strand breaks) and 2' deoxyguanosine (8-oxodG formation). AB - On irradiation of N-hydroxythiazole-2(3H)-thione 3 at 300 nm, the photoproducts disulfide 4, bisthiazole 5 and thiazole 6 are formed. During this photolysis, hydroxyl radicals are released, which have been detected by spin trapping with 5,5-dimethyl-1-pyrroline N-oxide (DMPO), coupled with electron paramagnetic resonance spectroscopy. In the presence of supercoiled pBR322 DNA, irradiation of thiazolethione 3 induces strand breaks through the photogenerated hydroxyl radicals, as confirmed by control experiment with the hydroxyl-radical scavenger isopropanol. Singlet oxygen appears not to be involved, as attested by the lack of a D2O isotope effect. During the photoreaction of thiazolethione 3 in the presence of 2'-deoxyguanosine (dG), the latter is photooxidized (ca 10% conversion after 2 h of irradiation) to the 7,8-dihydro-8-oxo-2'-deoxyguanosine as the main oxidation product. The dG conversion levels off after complete consumption of thiazolethione 3 and is suppressed by the addition of the hydroxyl radical scavenger 2,6-di-tert-butylcresol or DMPO. Since the photoproducts 4-6 are ineffective as sensitizers for the photooxidation of dG and DNA, the hydroxyl radicals released in the photolysis of thiazolethione 3 are the oxidizing species of DNA and dG. These results suggest that the thiazolethione 3 may serve as a novel and effective photochemical hydroxyl-radical source for photobiological studies. PMID- 11107847 TI - Effect of solvent on the excited-state photophysical properties of curcumin. AB - Photophysical properties of curcumin, 1,7-bis-(4-hydroxy-3-methoxy phenyl)-1,6 heptadiene-2,5-dione, a pigment found in the rhizomes of Curcuma longa (turmeric) have been studied in different kinds of organic solvent and also in Triton X-100 aqueous micellar media using time-resolved fluorescence and transient absorption techniques having pico and nanosecond time resolution, in addition to steady state absorption and fluorescence spectroscopic techniques. Steady-state absorption and fluorescence characteristics of curcumin have been found to be sensitive to the solvent characteristics. Large change (delta mu = 6.1 Debye) in dipole moments due to photoexcitation to the excited singlet state (S1) indicates strong intramolecular charge transfer character of the latter. Curcumin is a weakly fluorescent molecule and the fluorescence decay properties in most of the solvents could be fitted well to a double-exponential decay function. The shorter component having lifetime in the range 50-350 ps and percent contribution of amplitude more than 90% in different solvents may be assigned to the enol form, whereas the longer component, having lifetime in the range 500-1180 ps with less than 10% contribution may be assigned to the di-keto form of curcumin. Our nuclear magnetic resonance study in CDCl3 and dimethyl sulfoxide-D6 also supports the fact that the enol form is present in the solution by more than about 95% in these solvents. Excited singlet (S1) and triplet (T1) absorption spectrum and decay kinetics have been characterized by pico and nanosecond laser flash photolysis. Quantum yield of the triplet is low (phi T < or = 0.12). Both the fluorescence and triplet quantum yields being low (phi f + phi T < 0.18), the photophysics of curcumin is dominated by the energy relaxation mechanism via the internal conversion process. PMID- 11107848 TI - Photochromic behavior of 2,2-spiro-adamantylidene-2H-naphtho AB - The photokinetic behavior of a photochromic compound, 2,2-spiro-adamantylidene-2H naphtho[1,2-b]pyran (Py), has been investigated by using monochromatic irradiation in the UV and visible ranges. An unusually complex photochemistry occurs whereby two colored forms (o-quinone-allides) are produced, one of which is thermoreversible whereas the other is not thermoreversible but photoreversible. These are the result of ring opening of the pyran C-O bond to a short lived open-form intermediate which converts to the two different colored forms by twisting around different C-C bonds. Their spectra and molar absorption coefficients were obtained in acetonitrile solution. The kinetic parameters of the thermal bleaching (rate constant and activation energy) and photobleaching (quantum yield) were measured. A plausible reaction mechanism is proposed. Based on this mechanism, mathematical methods were devised which were capable of analyzing the system during its dynamic evolution and evaluating the quantum yields of the colorforming photoreactions. PMID- 11107849 TI - Probing the primary event in the photocycle of photoactive yellow protein using photochemical hole-burning technique. AB - Photochemical hole-burning spectroscopy was used to study the excited-state electronic structure of the 4-hydroxycinnamyl chromophore in photoactive yellow protein (PYP). This system is known to undergo a trans-to-cis isomerization process on a femtosecond-to-picosecond time scale, similar to membrane-bound rhodopsins, and is characterized by a broad featureless absorbance at 446 nm. Resolved vibronic structure was observed for the hole-burned spectra obtained when PYP in phosphate buffer at pH 7 was frozen at low temperature and irradiated with narrow bandwidth laser light at 431 nm. The approximate homogeneous width of 752 cm-1 could be calculated from the deconvolution of the hole-burned spectra leading to an estimated dephasing time of approximately 14 fs for the PYP excited state structure. The resolved vibronic structure also enabled us to obtain an estimated change in the C=C stretching frequency, from 1663 cm-1 in the ground state to approximately 1429 cm-1 upon photoexcitation. The results obtained allowed us to speculate about the excited-state structure of PYP. We discuss the data for PYP in relation to the excited-state model proposed for the photosynthetic membrane protein bacteriorhodopsin, and use it to explain the primary event in the function of photoactive biological protein systems. Photoexcitation was also carried out at 475 nm. The vibronic structure obtained was quite different both in terms of the frequencies and Franck-Condon envelope. The origin of this spectrum was tentatively assigned. PMID- 11107850 TI - Effects of UV irradiation on skin and nonskin-associated herpes simplex virus infections in rats. AB - Herpes simplex virus (HSV) normally causes vescular lesions on mucocutaneous surfaces but can also cause encephalitis. The virus can reactivate from the latent state in neurons to form recrudescent lesions. One common stimulus for reactivation is exposure to sunlight. In the present study, the effects of irradiating rats with suberythemal ultraviolet (UV) before or after infecting them epidermally with HSV was investigated. Preexposure to UV impaired HSV specific cellular immune responses, as indicated by delayed type hypersensitivity (DTH) and in vitro lymphoproliferation assays. However, the number and severity of the skin lesions were not altered. In contrast, exposure after infection did not affect cellular immunity but resulted in a large increase in the severity and number of lesions. In a second series of experiments, the effects of preirradiating with UV on HSV infection was examined using a route of inoculation which was not skin-associated, namely intranasal, allowing direct non-invasive access to the nervous system. It was found that suppressed DTH resulted, together with an increase in the incidence and severity of neurological symptoms and an increased viral load in the brain. Therefore, unlike the situation in the skin, irradiation of rats before intranasal inoculation led to a suppressed immune response to HSV which correlated with increased viral load and symptoms. These results indicate that the effects of UV may be dependent on whether the animal is exposed before or after the infection, and whether the infection is skin associated or systemic. PMID- 11107851 TI - Influence of UV radiation on four freshwater invertebrates. AB - Laboratory tests confirmed a negative and variable response of the following four species to artificial UV radiation: Cypridopsis vidua, an ostracode; Chironomus riparius, a midge larvae; Hyalella azteca, an amphipod; and Daphnia magna, a daphnid. Severe damage occurred at UV-B irradiance ranging from 50 to 80% of incident summer values. Under constant exposure to UV and photosynthetically active radiation (PAR) the acute lethal response was recorded at 0.3, 0.8, 0.8 and 4.9 W m-2 UV-B for D. magna, H. azteca, C. riparius and C. vidua, respectively. Sublethal UV-B damage to invertebrates included impaired movement, partial paralysis, changes in pigmentation and altered water balance (bloating). A series of UV-B, UV-A and PAR treatments, applied separately and in combination, revealed a positive role for both UV-A and PAR in slowing down UV-B damage. Mean lethal concentration values of the species typically more tolerant to UV and PAR (Cypridopsis, Chironomus) decreased conspicuously when both UV-A and PAR were eliminated. For UV-B-sensitive species (Hyalella, Daphnia) these differences were notably smaller. We suggest that this gradation of sensitivity among the tested species demonstrates potential differences in repairing mechanisms which seem to work more efficiently for ostracodes and chironomids than for amphipods and daphnids. Manipulations with a cellulose acetate filter showed that lower range UV-B (280-290 nm), produced by FS-40 lamps, may cause excessive UV damage to invertebrates. PMID- 11107852 TI - Photoactive protochlorophyllide regeneration in cotyledons and leaves from higher plants. AB - Chlorophyll accumulation during greening implies the continuous transformation of photoactive protochlorophyllide (Pchlide) to chlorophyllide. Since this reaction is a light-dependent step, the study of regeneration of photoactive Pchlide under a continuous illumination is difficult. Therefore this process is best studied on etiolated plants during a period of darkness following the initial photoreduction of photoactive Pchlide. In this study, the regeneration process has been studied using spinach cotyledons, as well as barley and bean leaves, illuminated by a single saturating flash. The regeneration was characterized using 77 K fluorescence emission and excitation spectra and high-performance liquid chromatography. The fluorescence data indicated that the same spectral forms of photoactive Pchlide are regenerated by different pathways: (1) photoactive Pchlide regeneration starts immediately after the photoreduction through the formation of a nonphotoactive Pchlide form, emitting fluorescence at approximately 651 nm. This form is similar to the large aggregate of photoactive Pchlide present before the illumination, but it contains oxidized form of nicotinamide adenine dinucleotide phosphate, instead of the reduced form (NADPH), in the ternary complexes; and (2) after the dislocation of the large aggregates of chlorophyllide-light-dependent NADPH:Pchlide a photooxidoreductase-NADPH ternary complexes, the regeneration occurs at the expense of the several nonphotoactive Pchlide spectral forms present before the illumination. PMID- 11107853 TI - Nanosecond laser photolysis studies of chlorosomes and artificial aggregates containing bacteriochlorophyll e: evidence for the proximity of carotenoids and bacteriochlorophyll a in chlorosomes from Chlorobium phaeobacteroides strain CL1401. AB - Time-resolved, laser-induced changes in absorbance, delta A(lambda; t), have been recorded with a view to probing pigment-pigment interactions in chlorosomes (control as well as carotenoid-depleted) and artificial aggregates of bacteriochlorophyll e (BChle). Control chlorosomes were isolated from Chlorobium phaeobacteroides strain CL1401, whose chromophores comprise BChle, bacteriochlorophyll a (BChla) and several carotenoid (Car) pigments; Car-depleted chlorosomes, from cells grown in cultures containing 2-hydroxybiphenyl. Artificial aggregates were prepared by dispersing BChle in aqueous phase in the presence of monogalactosyl diglyceride. In chlorosomes delta A(lambda; t) shows, besides a signal attributable to triplet Car (with a half-life of about 4 microseconds), signals in the Qy regions of both BChl. The BChla signal decays at the same rate as the Car signal, which is explained by postulating that some Car are in intimate contact with some baseplate BChla pigments, and that when a ground-state Car changes into a triplet Car, the absorption spectrum of its BChla neighbors undergoes a concomitant change (termed transient environment-induced perturbation). The signal in the Qy-region of BChle behaves differently: its amplitude falls, under reducing conditions, by more than a factor of two during the first 0.5 microsecond (a period during which the Car signal suffers negligible diminution), and is much smaller under nonreducing conditions. The BChle signal is also attributed to transient environment-induced perturbation, but in this case the perturber is a BChle photoproduct (probably a triplet or a radical ion). The absence of long-lived BChle triplets in all three systems, and of long-lived BChla triplets in chlorosomes, indicates that BChle in densely packed assemblies is less vulnerable to photodamage than monomeric BChle and that, in chlorosome, BChla rather than BChle needs, and receives, photoprotection from an adjacent Car. PMID- 11107854 TI - The effect of photodynamic action on two virulence factors of gram-negative bacteria. AB - Photodynamic therapy could provide an alternative to antibiotics for the treatment of local infections since a wide range of microorganisms have been shown to be susceptible to killing by photodynamic action (PDA) in vitro. The purpose of this study was to determine whether PDA was also able to affect the potency of two key bacterial virulence factors--lipopolysaccharide (LPS) and proteases. Suspensions of LPS from Escherichia coli and culture supernatants containing proteases of Pseudomonas aeruginosa were exposed to red light in the presence of toluidine blue O (TBO). The activity of each virulence factor was determined before and after irradiation. The limulus amoebocyte lysate (LAL) assay and the induction of proinflammatory cytokine (interleukin-8 and -6) release from human peripheral blood mononuclear cells (PBMC) were used for assessing the biological activity of LPS. Protease activity was quantified by azocasein hydrolysis. The biological activities of the LPS (both the LAL activity and its ability to induce cytokine release from PBMC) and the proteases were reduced significantly by irradiation with red light in the presence of TBO in a dose-dependent manner with respect to both the light energy dose and the TBO concentration. The ability of TBO-mediated PDA to reduce the activities of key virulence factors may be an additional benefit of using light-activated antimicrobial agents in the treatment of infectious diseases. PMID- 11107855 TI - Topical application of 5-aminolevulinic acid hexyl ester and 5-aminolevulinic acid to normal nude mouse skin: differences in protoporphyrin IX fluorescence kinetics and the role of the stratum corneum. AB - An important limitation of topical 5-aminolevulinic acid (ALA)-based photodetection and photodynamic therapy is that the amount of the fluorescing and photosensitizing product protoporphyrin IX (PpIX) formed is limited. The reason for this is probably the limited diffusion of ALA through the stratum corneum. A solution to this problem might be found in the use of ALA derivatives, as these compounds are more lipophilic and therefore might have better penetration properties than ALA itself. Previous studies have shown that ALA hexyl ester (ALAHE) is more successful than ALA for photodetection of early (pre)malignant lesions in the bladder. However, ALA pentyl ester slightly increased the in vivo PpIX fluorescence in early (pre)malignant lesions in hairless mouse skin compared to ALA. The increased PpIX fluorescence is located in the stratum corneum and not in the dysplastic epidermal layer. In the present study, ALA- and ALAHE-induced PpIX fluorescence kinetics are compared in the normal nude mouse skin, of which the permeability properties differ from the bladder. Application times and ALA(HE) concentrations were varied, the effect of a penetration enhancer and the effect of tape stripping the skin before or after application were investigated. Only during application for 24 h, did ALAHE induce slightly more PpIX fluorescence than ALA. After application times ranging from 1 to 60 min, ALA induced PpIX fluorescence was higher than ALAHE-induced PpIX fluorescence. ALA also induced higher PpIX production than ALAHE after 10 min of application with concentrations ranging from 0.5 to 40%. The results of experiments with the penetration enhancer and tape stripping indicated that the stratum corneum acts a barrier against ALA and ALAHE. Use of penetration enhancer or tape stripping enhanced the PpIX production more in the case of ALAHE application than in the case of ALA application. This, together with the results from the different application times and concentrations indicates that ALAHE diffuses more slowly across the stratum corneum than ALA. PMID- 11107856 TI - Fluorescence imaging during photodynamic therapy of experimental tumors in mice sensitized with disulfonated aluminum phthalocyanine. AB - A fluorescence imaging system was used to monitor the emission of disulfonated aluminum phthalocyanine (AlS2Pc) during the photodynamic therapy (PDT) of murine tumors. Cells of the MS-2 fibrosarcoma were injected in mice in two compartments in order to cause the development of tumors in different host tissues. Two drug doses and two uptake times were considered. Moreover, the fluorescence of the AlS2Pc was excited using two wavelengths on the opposite sides of the absorption peak to detect a possible change in the absorption spectrum of the sensitizer induced by the PDT. In the tumors, the treatment induces a variation of the fluorescence intensity: in some mice a mild photobleaching takes place, in others a fluorescence enhancement occurs. Which effect predominates depends on the experimental conditions, even though a large spread of data was found amongst mice of the same group. In all mice, independently of the drug dose, uptake time or tumor compartment, a marked increase in the fluorescence signal takes place at the borders of the irradiated area. To quantify this effect we evaluated the ratio between the fluorescence intensities in the peritumoral area and in the tumor itself. This ratio increases monotonically during the PDT, showing a different behavior with the two excitation wavelengths. This indicates that the AlS2Pc absorption spectrum shifts toward shorter wavelengths as a result of the irradiation. PMID- 11107857 TI - PEG-m-THPC-mediated photodynamic effects on normal rat tissues. AB - Photodynamic therapy (PDT) of malignancies uses light to activate a photosensitizer preferentially accumulated in cancer cells. The first pegylated photosensitizer, tetrakis-(m-methoxypolyethylene glycol) derivative of 7,8 dihydro-5,10,15,20-tetrakis(3-hydroxyphenyl)-21-23-[H]-porphyrin (PEG-m-THPC), was evaluated in non-tumor-bearing rats. The aim of this study was to assess the photodynamic threshold for damage and its sequelae in normal rat tissue. Thirty five Fischer rats were sensitized with 3, 9 or 30 mg/kg body weight PEG-m-THPC. Colon, vagina and perineum were irradiated with laser light of 652 nm wavelength and an optical dose of 50, 150 or 450 J/cm fiber length. Temperature in the pelvis was measured during PDT. Three days following PDT the effect on skin, vagina, colon, striated muscle, connective tissue, nerves and blood vessels was assessed by histology. The healing of the above-mentioned tissues was assessed on two rats 3 and 8 weeks after PDT using 9 mg/kg PEG-m-THPC activated with 450 J/cm laser light. No dark toxicity was observed. PDT using 30 mg/kg PEG-m-THPC induced severe necrosis irrespective of the optical dose. Body weight of 9 or 3 mg/kg activated with less than 450 J/cm induced moderate or no damage. No substantial increase in body temperature was seen during PDT. Tissues with severe PDT-induced damage seem to have a good tendency to regenerate. We conclude that within the dose required for tumor treatment PEG-m-THPC is a safe photosensitizer with promising properties. PDT of the colon mucosa below 9 mg/kg PEG-m-THPC and 150 J/cm seems to be safe. All other tissues can be exposed to 9 mg/kg PEG-m-THPC activated with less than 450 J/cm laser light with little side effects. PMID- 11107858 TI - The dose dependence of cyclobutane dimer induction and repair in UVB-irradiated human keratinocytes. AB - UVB and UVA components of the solar spectrum or from artificial UV-sources might be important etiological factors for the induction and development of skin cancer. In particular, deficiencies in the capacity to repair UV-induced DNA lesions have been linked to this phenomenon. However, until now only limited data are available on the biological and physical parameters governing repair capacity. We have, therefore, developed a flowcytometric assay using fluorescence labeled monoclonal antibodies to study the dose-dependence of induction and repair of UVB-induced cyclobutane pyrimidine dimers in a spontaneously immortalized keratinocytic cell line (HaCaT). Our results show that the kinetics of recognition and incision of UVB-induced DNA lesions slows down by a factor of about 3 in a dose range of 100-800 J m-2. Furthermore, a thorough analysis of repair kinetics indicates that this reduction in repair capacity might not be dependent on saturation of enzymatic repair capacity (Michaelis-Menten) but may be caused by a UV-induced impairment of enzymes involved in DNA repair. Because this effect is evident in vitro at doses comparable to the minimal erythemal dose in vivo, our results might have significant impact on risk assessment for UV induced carcinogenesis. PMID- 11107859 TI - Analyses of structure of photoreceptor organelle and blepharismin-associated protein in unicellular eukaryote Blepharisma. AB - In the ciliated protozoan Blepharisma, step-up photophobic response is believed to be mediated by a novel type of photosensory pigment known as "blepharismins" (BL) that are contained in the pigment granules located just beneath the plasma membrane. We examined the ultrastructure of the pigment granules by freeze fracture and thin-section electron microscopy and proposed a schematic diagram showing the granules' three-dimensional inner membranous structure. Some of the BL are suggested to be associated with 200 kDa membrane protein. High-pressure liquid chromatography analysis of pigment species associated with 200 kDa protein obtained from blue forms of Blepharisma (oxyblepharisma) revealed that the 200 kDa protein was associated with five types of oxyblepharismin. The fluorescence intensity was increased when the pigments were dissociated from the 200 kDa protein. The result supports the hypothesis that the pigment-200 kDa complex is able to transduce light energy into signals mediating the photobehavior of Blepharisma. PMID- 11107860 TI - Water and carboxyl group environments in the dehydration blueshift of bacteriorhodopsin. AB - The proton channels of the bacteriorhodopsin (BR) proton pump contain bound water molecules. The channels connect the purple membrane surfaces with the protonated retinal Schiff base at the membrane center. Films of purple membrane equilibrated at low relative humidity display a shift of the 570 nm retinal absorbance maximum to 528 nm, with most of the change occurring below 15% relative humidity. Purple membrane films were dehydrated to defined humidities between about 50 and 4.5% and examined by Fourier transform infrared difference spectroscopy. In spectra of dehydrated-minus-hydrated purple membrane, troughs are observed at 3645 and 3550 cm-1, and peaks are observed at 3665 and 3500 cm-1. We attribute these changes to water dissociation from the proton uptake channel and the resulting changes in hydrogen bonding of water that remains bound. Also, in the carboxylic acid spectral region, a trough was observed at 1742 cm-1 and a peak at 1737 cm-1. The magnitude of the trough to peak difference between 1737 and 1742 cm-1 correlates linearly with the extent of the 528 nm pigment. This suggests that a carboxylic acid group or groups is undergoing a change in environment as a result of dehydration, and that this change is linked to the appearance of the 528 nm pigment. Dehydration difference spectra with BR mutants D96N and D115N show that the 1737-1742 cm-1 change is due to Asp 96 and Asp 115. A possible mechanism is suggested that links dissociation of water in the proton uptake channel to the environmental change at the Schiff base site. PMID- 11107861 TI - [Graves-Basedow ophthalmopathy in light of current views]. AB - Ophthalmopathy developing in the course of Graves-Basedow disease, although known for years, still causes controversy both about classification of eye symptoms, treatment modalities and terminology. The paper gives an update view on the etiopathogenesis of ophthalmopathy, terminology, classification and dynamics of clinical symptoms, as well as it presents the methods for treating its most important symptoms. Particular attention was focused on surgical treatment of lid retraction and strabismus, as well as on the present requirements in pharmacological and surgical treatment of the most serious complication of ophthalmopathy i.e. optic neuropathy. The role of an ophthalmologist in the process of diagnosis and monitoring of Graves' ophthalmopathy was also emphasised. PMID- 11107862 TI - [Surgical correction of strabismus in Graves-Basedow ophthalmopathy. Personal experiences]. AB - The aim of the study was to evaluate long-term results of surgical treatment of strabismus and diplopia connected with Graves' ophthalmopathy. Surgery of extraocular muscles was performed in 64 patients, previously with different methods for their Graves' ophthalmopathy. All patients showed stabilization of the strabismus angle and diplopia as well as euthyroidism. Before the operation ultrasonography, orbital computed tomography and forced duction test were performed. Before the operation horizontal strabismus with vertical deviation and vertical strabismus were most frequent (36.6% and 35.2%, respectively). A total of 136 operations had been performed. Postoperative follow-up ranged from 6 months to 15 years, including 30 patients with the follow-up exceeding one-year. Recovery or recovery without binocular single vision after surgery was observed in 47 patients (73.4%), including 34 patients (53.1%) with single binocular vision. Improvement was seen in 15 patients (23.4%). No improvement was observed in two cases (3.1%) due to marked vertical strabismus before surgery, they were referred for further surgical treatment. Overcorrection after surgery in 3 cases was observed, and in one case we applied prismatic correction successfully, the two others required reoperation. CONCLUSIONS: 1. Surgical treatment of strabismus and diplopia in Graves' ophthalmopathy is very useful because of high percentage of positive results, confirmed in long-term observation. 2. Diplopia and a large angle of strabismus are a necessary indication for surgical correction of extraocular muscles. 3. A prerequisite for surgical treatment is stable euthyreosis and stabilization of the strabismus angle as well as diplopia and performation of forced duction test for evaluation of stage disease. 4. Prior treatment of ophthalmopathy and duration of strabismus as well as diplopia do not appear to influence this outcome. PMID- 11107863 TI - [Ophthalmologic aspects of treating optic neuropathy during the course of Graves Basedow disease]. AB - The aim of the present study is to evaluate the efficacy of conservative treatment of optic neuropathy in the course of Graves' disease. Nineteen patients with neuropathy out of 520 patients with Graves' disease, including 15 women and 4 men were treated. Age of the patients ranged from 36 to 69 years, mean age was 51.3. In all patients thyroid function was normalized, 4 patients underwent strumectomy. Bi- and unilateral neuropathy was found in 13 and 6 cases respectively. The therapy consisted of high-dose steroids (Prednisone in 12 cases or Dexamethasone i.v. in 3 cases) or they were combined with gamma cobalt-60 irradiation of orbit tissues (2000 c Gy in 10 fractions) in 8 cases. In 2 cases radiotherapy alone was performed. Two cases were excluded from treatment because of optic atrophy during the first examination. In all patients full ophthalmological investigation, ultrasonography and computed tomography of the orbit before and after the treatment were performed. Follow-up ranged from 5 to 46 months, mean 21.4. Improvement of visual acuity and perimetry (p < 0.05) after the treatment was observed and changes of the fundus after the treatment were significantly (p = 0.000) improved. Exophthalmos significantly decreased (p < 0.05) but only in the right eye. In all patients the therapy reduced the ophthalmopathy index from mean 7.5 to 4.1 (p = 0.000). IN CONCLUSION: 1. Early diagnosis and appropriate, constant conservative treatment of optic neuropathy in Graves' disease prevent from blindness and surgical intervention. 2. Corticotherapy in combination with radiotherapy of the orbit is a beneficial method of the treatment of optic neuropathy in Graves' disease. PMID- 11107864 TI - [Risk factors for occurrence of ophthalmopathy in the course of Graves-Basedow disease]. AB - Ophthalmopathy is a serious complication of Graves-Basedow disease. It is a common and the most significant symptom of this illness and usually persists even when other symptoms associated with abnormal thyroid function disappear. Despite of well-known auto-immunologic patho-genesis of ophthalmopathy it is still hard to predict when this severe complication will occur. This condition is thought to be associated with diagnosis and treatment errors. We studied the patients suffering from Graves-Basedow disease with and without ophthalmopathy. We concluded that the risk factors for ophthalmopathy were: many non-treated episodes of hyperthyroidism, delayed diagnosis and initiation of therapy, no thyroxine supplementation, improper preparation to strumectomy. PMID- 11107865 TI - [Optimizing computed tomography diagnosis in retrobulbar orbit tumors]. AB - CT is still one of the most commonly applied diagnostic procedure in cases of suspected neoplastic lesions in the retrobulbar region. Orbit tumors are rare, however due to their localisation close to the organ of sight, they make a serious clinical problem. The authors analysed retrospectively 127 tumors (interview, ophthalmological examination, CT, surgical procedure, histopathologic findings) with the aim of improving CT diagnostics. PMID- 11107866 TI - [Computed tomography images of selected retrobulbar orbit tumors]. AB - Accurate evaluation of the retrobulbar orbit tumors and obtaining similar radiologic and histopathologic diagnosis are the most important for the therapeutic management. The authors analysed a group of 124 patients undergoing orbital CT exams and subsequently surgical procedure. They differentiated and described typical CT images of cavernous angioma, malignant lymphoma, optic glioma and dermoid cyst of orbit in correlation with the clinical data. PMID- 11107867 TI - [Influence of gluten free diet on bone mineral density (BMD) in children with celiac disease]. AB - In children with celiac disease (CD) bone metabolism and mineralization are frequently disturbed. The present study aimed to assess the influence of gluten free diet (GFD) on bone mineral density (BMD) in 73 children with CD, mean age of 12.4 +/- 0.4 years and mean body mass index (BMI) 17.9 +/- 0.4 kg/m2 (mean +/- SEM). Diagnosis of CD was established according to ESPEGAN criteria. Compliance to the GFD was verified on the basis of interview and by estimation of antiendomysial antibodies (IgAEmA/IgGEmA) in blood serum. BMD was measured by dual energy X-ray absorptiometry (DEXA). Plasma calcium (Ca) and phosphorus (P) concentrations, alkaline phosphatase (AP) and its bone fraction (BAP) were estimated before BMD measurement. All children were divided into two groups. Group A consisted of 33 children where gluten free diet was strictly respected for 11.7 +/- 0.6 years. The second group (Group B) comprised 40 children without strictly respected GFD. Children who strictly followed GFD showed statistically higher BMI, AP-spine BMD and total body BMD in comparison with children without GFD (BMI 19 +/- 0.52 kg/m2 vs 17.3 +/- 0.4 kg/m2; p < 0.01, AP-spine BMD 0.951 +/ 0.04 g/cm2 vs 0.767 + 0.03 g/cm2; p < 0.005, Total Body BMD 1.013 +/- 0.02 g/cm2 vs 0.933 +/- 0.01 g/cm2; p < 0.05) respectively. No significant differences were found in plasma Ca, P, AP, BAP between both groups. A statistically significant positive correlation (p < 0.001) was found between duration of GFD and AP-spine BMD and total body BMD, respectively. A statistically significant positive correlation (p < 0.05) was also found between duration of GFD and BMI. CONCLUSION: Long-term GFD significantly improves BMD and BMI in children with CD. PMID- 11107868 TI - [Renal osteodystrophy in dialysis patients as estimated by three-point bone densitometry]. AB - The aim of a study was to estimate the renal osteodystrophy status using bone densitometry in relation to selected biochemical parameters of calcium-phosphate metabolism. The study population consisted of 123 patients with end-stage renal disease, including 24 patients treated with continuous ambulatory peritoneal dialysis (CAPD), aged between 22 and 73 years (mean 49.9 years), on dialysis program for mean period of 14.9 months and 99 patients on maintenance hemodialysis for mean period of 58.8 months, aged between 19 and 72 years (mean 46.6 years). Densitometric measurements using DEXA technique were performed in three different skeletal points: distal ends of both radial bones, lumbar spinal region and femoral neck. Concomitantly, serum concentrations of total and ionized calcium, phosphates and parathormone as well as alkaline phosphatase serum activity were measured. Among male patients treated with CAPD significantly higher BMD values in right forearm were found as compared to women treated with this method (0.769 vs. 0.616; p < 0.001). Higher values of BMD were also found in both forearms in whole CAPD population as compared to those on hemodialysis. However, there was no difference in densitometry results between CAPD and HD patients as well as between men and women within these groups, when measured in femoral neck and lumbar spinal region. Among hemodialysis patients higher levels of phosphates and PTH were found as compared to CAPD, doses of drugs used for treatment of osteodystrophy--calcium carbonate, aluminum hydroxide and active vitamin D were also higher in individuals on HD. In addition, in CAPD patients statistically significant, positive correlations were found between BMD value in lumbar spinal area as well as in femoral neck and amount of ingested calcium carbonate, between BMD in lumbar spinal area and aluminum hydroxide dose taken by patients and between BMD in both forearms and dose of active vitamin D. We failed to demonstrate any relationship between obtained densitometric results as well as biochemical markers of calcium-phosphate metabolism and quantitative parameters of dialysis adequacy in both treatment modes. Obtained results let us to conclude that renal osteodystrophy is less advanced in patients treated with peritoneal dialysis, however this may be related only to markedly shorter renal replacement therapy period in this group. Lack of significant abnormalities in densitometry measurements taken in lumbar spinal area and femoral neck, while they are present in forearms, may suggest that the latter point of skeleton may be most useful for identification of bone mass deficiency in dialyzed patients. PMID- 11107869 TI - [The impact of changes in levels of calcium, phosphate and magnesium during hemodialysis on autonomic system reactivity as measured by heart rate variability analysis]. AB - The aim of the study was to analyze the response of autonomic nervous system to dialysis related systemic volume reduction using heart rate variability (HRV) analysis. The possible relationship between changes in serum levels of calcium phosphate balance parameters and HRV measurements results was also analyzed. The study was performed in 32 patients (20 men, 12 women) aged between 27 and 71 years (mean 44 years) treated with maintenance dialysis due to end-stage renal disease from 3 months to 15 years (mean 4.4 years). All parameters mentioned above were analyzed during 4-hour dialysis session. Mean value of LF/HF ratio at the beginning of the procedure was 5.36, with continuous increase in consecutive measurements performed 30-minute intervals to the maximal value 8.2 in 120th minute of HD session (p < 0.05). In the next measurements continuous decrease in the mentioned parameter was noticed, to the mean value 6.99 in minute 240. The values of LF/HF ratio were also lower for the whole HD session in the group of patients with initial predialytic concentration of calcium lower than 2.35 mmol/l as compared to those with initial calcium concentration higher than 2.35 mmol/l. Higher values of LF/HF ratio and bigger oscillation amplitude of this parameter were also noted in those patients, in whom the percentage reduction of magnesium level during dialysis exceeded 20%. In addition, statistically significant relationship between percentage reduction in magnesium ion concentration and LF/HF ratio during HD was found. Obtained results let us to conclude, that hemodialysis leads to important change in the activity of both components of autonomic nervous system. The factors which may adversely influence the quality of this response may be, among others, low total calcium ion concentration as well as low percentage reduction in magnesium level during hemodialysis session. PMID- 11107870 TI - [Nutritional therapy in decompensated liver cirrhosis]. AB - In decompensated liver cirrhosis, all effort should be made to maintain normal nutritional status so long as possible. Early nutritional assessment and dietetic intervention in the management of malnutrition, encephalopathy, ascites, infection and bleeding from gastrointestinal tract markedly decreases mortality and improves survival after liver transplantation in this group of patients. PMID- 11107871 TI - [New hypoglycemic drugs in treatment of diabetes type 2]. AB - Diabetes mellitus type 2 is a syndrome with different etiopathology and clinical picture. Results of clinical studies show that hyperglycaemia increases during the time of diabetes even if it is intensively treated. At the present time some new hypoglycaemic drugs have been introduced for type 2 diabetes mellitus therapy. These drugs can increase insulin secretion by the inhibition of the ATP sensitive K+ channels (glimepyride, repaglinide) or by incretin action (glucagon like peptide-1). Other drugs can decrease insulin resistance acting on nuclear receptors (troglitazone). Normalization of stomach motoric activity can also lead to decreasing of blood glucose concentration (pramlintid). Also other mechanisms of action are taken into consideration, like changing of cytokine activity or catabolism regulation of beta-cells of the Langerhans' islets. PMID- 11107872 TI - [Iodine, sodium-iodine symporter and thyroid diseases]. AB - The Na+/I- symporter (NIS) is the plasma membrane protein that catalyzes the accumulation of iodine into thyroid cells against an electrochemical gradient. This iodine uptake is the first step in thyroid hormone biogenesis which is stimulated by TSH--the main hormone regulating thyroid--specific functions. This process requires energy from the action of Na+/K+ ATP-ase. TSH upregulates NIS gene expression, and this induction can be modulated by cytokines. Analysis of the distribution of NIS mRNA demonstrated variable levels of NIS transcription in different tissue samples. Increased NIS expression has been found in autonomous adenoma and Graves' disease, decreased levels of NIS protein and/or mRNA NIS were observed in Hashimoto's disease, scintigraphically cold nodules and most thyroid cancers. NIS mutation has been found in congenital hypothyroidism. Potential defects in the NIS gene are currently under investigation in various thyroid diseases. PMID- 11107873 TI - [Characteristic features of primary hyperparathyroidism caused by parathyroid cancer--based on 2 cases]. AB - Two cases of women with primary hyperparathyroidism caused by parathyroid cancer were presented. The authors noticed the following characteristic features of primary hyperparathyroidism in the course of the cancer: rich clinical symptomatology usually in form of considerable bone destruction, renal stones and nephrocalcinosis, biochemically very high level of calcium, above 14-16 mg%, threatening with hypercalcemic crisis and considerably higher parathormone serum concentration even up to twenty times above the norm. Parathyroid cancer, more often than adenoma, is a stiff and large neck tumour accessible for palpation. There are no specific biochemical and imagining examination techniques to recognise beyond any doubt the cancer character of primary hyperparathyroidism before operation. The histopathological diagnosis of this cancer is difficult and is not usually done intraoperatively. The recurrences of the malignancy are typical and they require reoperations after stating places of relapse or metastases. PMID- 11107874 TI - [Hemochromatosis--report of 2 cases]. AB - Hereditary hemochromatosis is an iron metabolism disorder known for a century, leading to iron overload and dysfunction of the major organs of the body. Early diagnosis and treatment result in a normal life expectancy. Authors present a report of two cases (the father and his son) in early stadium of the disease. PMID- 11107875 TI - [Liver transplantation in a patient with hemolytic syndrome in the course of fulminant Wilson's disease]. AB - The case report of a 20 year old woman with fulminant liver failure and hemolytic syndrome was described. Huge amounts of copper excreted in the urine (3555 mg/12 h without, and 5180 mg/12 h after d-penicillamine provocation, respectively) confirmed the diagnosis of fulminant Wilson's disease. Because the patient's general condition worsened rapidly (hemolysis, diathesis hemorrhagic, ascites, encephalopathy increased during 3 days of clinical observation) orthotopic liver transplantation was performed. After the transplantation, ischemic type biliary lesion (ITBL) II stage was diagnosed. The woman is still being treated with Prograf and Urso-Falk. The patient returned to her normal life, continues to work and was married. Two years after OLT she gave birth to a healthy boy. The liver function tests are normal with the exception of GGTP and FALK activities elevation. Copper and ceruloplasmin level, as well as copper excretion in the urine are within the normal values. PMID- 11107876 TI - Introduction to the special issue: psychology in the 21st century. PMID- 11107877 TI - DNA. AB - The authors predict that in a few years, many areas of psychology will be awash in specific genes responsible for the widespread influence of genetics on behavior. As the focus shifts from finding genes (genomics) to understanding how genes affect behavior (behavioral genomics), it is important for the future of psychology as a science that pathways between genes and behavior be examined not only at the molecular biological level of cells or the neuroscience level of the brain but also at the psychological level of analysis. After a brief overview of quantitative genetic research, the authors describe how genes that influence complex traits like behavioral dimensions and disorders in human and nonhuman animals are being found. Finally, the authors discuss behavioral genomics and predict that DNA will revolutionize psychological research and treatment early in the 21st century. PMID- 11107878 TI - Multilevel integrative analyses of human behavior: social neuroscience and the complementing nature of social and biological approaches. AB - Social and biological explanations traditionally have been cast as incompatible, but advances in recent years have revealed a new view synthesized from these 2 very different levels of analysis. The authors review evidence underscoring the complementing nature of social and biological levels of analysis and how the 2 together can foster understanding of the mechanisms underlying complex behavior and the mind. Specifically, they review the utility of considering social influences on biological processes that are often viewed as outside the social domain including genetic constitution, gene expression, disease, and autonomic, neuroendocrine, and immune activity. This research underscores the unity of psychology and the importance of retaining multilevel integrative research that spans molar and molecular levels of analysis. Especially needed in the coming years is more research on the mechanisms linking social and biological events and processes. PMID- 11107879 TI - The relationship context of human behavior and development. AB - The influence of social relationships on human development and behavior is receiving increased attention from psychologists, who are central contributors to the rapidly developing multidisciplinary field of relationship science. In this article, the authors selectively review some of the significant strides that have been made toward understanding the effects of relationships on development and behavior and the processes by which relationships exert their influence on these, with the purpose of highlighting important questions that remain to be answered, controversial issues that need to be resolved, and potentially profitable paths for future inquiry. The authors' thesis is that important advances in psychological knowledge will be achieved from concerted investigation of the relationship context in which most important human behaviors are developed and displayed. PMID- 11107880 TI - Cognitive neuroscience: origins and promise. AB - Both Freud and Wundt had hoped to base psychology on an understanding of the neural basis of mental events. Their efforts were unsuccessful because the structure and function of the human brain was not available for empirical study at the physiological level. Over the last part of this century, there has been amazing growth and vitality in the field of human brain function. In this paper, we trace critical developments in the fields of cognitive psychology, neuropsychology, and brain imaging related to the development of cognitive neuroscience. Cognitive Neuroscience has established that the decomposition of mental events can be united with an understanding of the mental and emotional computations carried out by the human brain. Cognitive neuroscience has the capability of influencing psychology in diverse areas from how children develop to how adults age; from how humans learn to how we imagine; from volitional control to psychopathologies. PMID- 11107881 TI - Emotion, plasticity, context, and regulation: perspectives from affective neuroscience. AB - The authors present an overview of the neural bases of emotion. They underscore the role of the prefrontal cortex (PFC) and amygdala in 2 broad approach- and withdrawal-related emotion systems. Components and measures of affective style are identified. Emphasis is given to affective chronometry and a role for the PFC in this process is proposed. Plasticity in the central circuitry of emotion is considered, and implications of data showing experience-induced changes in the hippocampus for understanding psychopathology and stress-related symptoms are discussed. Two key forms of affective plasticity are described--context and regulation. A role for the hippocampus in context-dependent normal and dysfunctional emotional responding is proposed. Finally, implications of these data for understanding the impact on neural circuitry of interventions to promote positive affect and on mechanisms that govern health and disease are considered. PMID- 11107882 TI - Choice and the relative pleasure of consequences. AB - Although pleasure played a central role in early theories of decision making, it gradually became peripheral, largely because of measurement concerns. Normative theories became more mathematical, and descriptive theories emphasized cognition over emotion. In recent years, there has been a renewed interest in emotions and choice. This article examines attempts to model pleasure and pain in terms of utilities, decision weights, and counterfactual comparisons. Research on disappointment and regret has provided both empirical and theoretical insights. Many researchers now realize that the predictability of the emotions that follow from decisions is as important as the predictability of choice. PMID- 11107883 TI - Beyond behaviorism: on the automaticity of higher mental processes. AB - The first 100 years of experimental psychology were dominated by 2 major schools of thought: behaviorism and cognitive science. Here the authors consider the common philosophical commitment to determinism by both schools, and how the radical behaviorists' thesis of the determined nature of higher mental processes is being pursued today in social cognition research on automaticity. In harmony with "dual process" models in contemporary cognitive science, which equate determined processes with those that are automatic and which require no intervening conscious choice or guidance, as opposed to "controlled" processes which do, the social cognition research on the automaticity of higher mental processes provides compelling evidence for the determinism of those processes. This research has revealed that social interaction, evaluation and judgment, and the operation of internal goal structures can all proceed without the intervention of conscious acts of will and guidance of the process. PMID- 11107884 TI - Toward DSM-V and the classification of psychopathology. AB - The Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) developed by the American Psychiatric Association (1994) is a compelling effort at a best approximation to date of a scientifically based nomenclature, but even its authors have acknowledged that its diagnoses and criterion sets are highly debatable. Well-meaning clinicians, theorists, and researchers could find some basis for fault in virtually every sentence, due in part to the absence of adequate research to guide its construction. Some points of disagreement, however, are more fundamental than others. The authors discuss issues that cut across individual diagnostic categories and that should receive particular attention in DSM-V: (a) the process by which the diagnostic manual is developed, (b) the differentiation from normal psychological functioning, (c) the differentiation among diagnostic categories, (d) cross-sectional vs. longitudinal diagnoses, and (e) the role of laboratory instruments. PMID- 11107885 TI - Research on psychotherapy efficacy and effectiveness: between Scylla and Charybdis? AB - Clinical researchers have recently begun to explore differences between psychotherapy outcome studies that focus on efficacy and those that focus on effectiveness. The authors provide concise descriptions of these research models, followed by more extended consideration of the most important conceptual and empirical distinctions between the two. Research on the efficacy/effectiveness distinction is then put into context: The common treatment variables that also influence treatment outcomes are reviewed. Fifty years of research on psychotherapy outcomes are next considered; contemporary research on the efficacy and effectiveness research models is emphasized. A description and evaluation of current efforts to heighten the value of technique-focused research to clinicians follow. The authors conclude by anticipating some promising future directions in this research domain. PMID- 11107886 TI - [Centennial of Pedro Kouri]. PMID- 11107887 TI - [Plesiomonas shigelloides, a Vibrionaceae to be taken into account]. AB - The antigenic structure and antimicrobial susceptibility were studied in 99 strains isolated from patients with acute diarrhea (6 strains from an outbreak of digestive transmission disease in Santiago de Cuba) and a strain isolated from a patient who died from infections neurological syndrome (INS, meningitis). Four new serotypes (093, 994, 095, 096), which had not been described in the world classification, were identified from the Cuban isolated strains and were included in the International Serotyping Scheme by the International Reference Center located in Prague, Czech Republic. For the first time in Cuba, the circulation of serotypes 017:H11, 011: H2, 023. H1alc, 057: H3 which show cross reaction to Shiguella species was proved. Those strains from the outbreak of digestive disease belonged to serotype 050: H11 and had a thermostable toxin. The first case of infectious neurologic syndrome with Plesionomas shigelloides etiology reported in Cuba was described; the strain corresponded to serotype 050: H11. The worldwide reported pattern of antimicrobial resistance was demonstrated. PMID- 11107888 TI - [Rapid detection of Enterovirus by a direct method of polymerase chain reaction]. AB - For the detection of Enterovirus, we devised a direct economical method of polymerase chain reaction which does not require a previous extraction of ribonucleic acid and uses infected cell culture supernatants. The system was developed on the basis of universal primers of Enterovirus and specific primers of vaccinal strain Sabin 1. The achieved results proved that the non-existence of methods of RNA extraction and purification prior to the reaction does not affect the susceptibility and specificity of the system, in the rapid detection of Enterovirus genomes and identification of vaccinal strains of poliovirus. PMID- 11107889 TI - [Folic acid deficiency and increased concentrations of formate in serum and cerebrospinal fluid of patients with epidemic optical neuropathy]. AB - We studied 62 patients aged 48 years as an average and diagnosed with bilateral optical neuropathy during an epidemics in Pinar del Rio province. Of these patients, 42 showed the optical form whereas 20 had the mixed form of optical neuropathy. We researched into the levels of formate and folate in serum and cerebrospinal fluid samples and we found a marked deficiency of folates in more than 50% of samples and high formate concentration levels in almost 25% of samples. We concluded that nutritional shortages that lead to a reduction of folates, and the intake of small amounts of methanol in alcoholic drinks could lead to lacking energetic states which would facilitate that the optical nerve be affected and the epidemic optical neuropathy appear. PMID- 11107890 TI - [Resistance to insecticides in Blattella germanica species strains from Santiago de Cuba]. AB - A study was conducted on the level of resistance to seven insecticides, namely, 3 organophosphate compounds (malathion, chlorpyrifos and pirimiphos-methyl), one carbamate (propoxur) and 1 pyrethroid (cypermethrin, deltamethrin and lambdacialotrine) of three field-collected strains of Blattella germanica (Linnaeus, 1767) from Santiago de Cuba. These strains showed high resistance levels to malathion, cypermethrin, deltamethrin and lambdacialotrine and low resistance to pirimiphos-methyl, and also they were susceptible to chlorpyrifos and propoxur. The levels of resistance to tested organophosphate insecticides such as malathion and pirimiphos-methyl and to pyrethroid compounds like cypermethrin, deltamethrin and lambdacialotrine may be related to the increased production of esterases as a mechanism of resistance. The value of frequency of the resistant genes for enzyme acetylcholinesterase was very low, therefore, the modified acetylcholinesterase is not involved in resistance to insecticides tested in the studied strains from Santiago de Cuba. PMID- 11107891 TI - [Specific IgE response to Blomia tropicalis mites in Cuban patients]. AB - We performed a prospective crosswise analytical study of 84 patients treated in the Allergy Service of "Calixto Garcia" General Hospital whose skin tests to allergenic Blomia tropicalis extract were positive. These patients' sera were used in assays for determining specific IgE antibodies and in the identification of allergen proteins in this extract by means of western blotting. Forty-four patients (55%) showed specific IgE levels higher than 0.1 U and 17 patients exhibited levels over 0.3 U. Bronchial asthma and rhinitis were the pathologies with elevated specific IgE levels. However, generally speaking, the specific IgE values were lower than those reported in other mites in spite of the fact that skin reactivity behaved the same. It was determined that specific IgE antibodies are mainly directed to allergens with a molecular weight from 13 to 15 D. PMID- 11107892 TI - [Introduction of Bacillus sphaericus strain-2362 (GRISELESF) for biological control of malaria vectors in Guatemala]. AB - Malaria continues to be an important health problem in a number of countries of Central and South America where it is considered as a highly prevent endemic disease. The objective of this paper is to assess the entomo-epidemiological impact of a pilot program for the biological control of malaria-transmitting vectors, which was implemented in 1998 in Escuintla, Republic of Guatemala. This program was based on the use of 20,000 L of biolarvicide Bacillus sphaericus- strain-2362 (GRISELESF) which was applied in the 46 localities of highest epidemiological risk at a rate of 10 mL/m2 of effective area of breeding. The entomologic effectiveness of this biolarvicide was monitored from the first 72 hours to 4 months after the application. There was a total larval reduction of 94.57 in the maturity stage of the water phase of Anopheles albimanus vector. The epidemiological analysis was carried out by comparing the rate of malaria prevalence (per 1000 pop) during 1997 and 1998. The five treated municipalities showed a statistically significant reduction of 50% (p 0.01). The results obtained in this paper coincided with those reported by comparable studies, so, this allowed us to recommend the use of the biolarvicide Bacillus sphaericus (strain-2362) as part of a comprehensive program of malaria-transmitting vector control in the Republic of Guatemala and other countries of the region. PMID- 11107893 TI - [Knowledge and practice concerning pediculosis in a health district]. AB - Pediculosis is an endemics that has recently intensified world wide and also in Cuba as of 1970. A study was performed in the schools located in the health area of the Vedado Polyclinics in the City of Havana to find out the level of knowledge acquired and the practices followed that may have an effect on the control of this disease. Two qualitative techniques i.e. group analysis and non participatory observation allowed to gather information. Wrong pieces of knowledge and ill practices were detected such as the belief that nits can fly, the use of a drug called Lindano 1% to treat children as a preventive method while they are in class, application of products in a wrong way, use of harmful products and no taking out of nits in a systematic way. These mistaken concepts and practices affect the analysis of cases and thus, they should be taken into consideration to increase the efficiency of the control program. PMID- 11107894 TI - [Work tool for the study of sexually transmitted diseases and HIV/AIDS in adolescents]. AB - A working instrument was designed to explore sex knowledge, practices and behaviours as well as factors related to sexually transmitted diseases and HIV/AIDS. The instrument consists of a questionnaire that should be asked by the interviewer. This questionnaire was validated in a test carried out in the City of Havana and submitted to the expert's criteria to improve its overall design. PMID- 11107895 TI - [Evaluation of surveillance in primary health care: a methodological proposal]. AB - Given the development of public health, hygiene and epidemiology in Cuba and their linking with primary care, and particularly, the novel approach to health surveillance in its different implementation alternatives, it is necessary to develop new forms and methods for surveillance evaluation at this level. The paper is aimed at presenting methodological proposal for assessing surveillance in primary health care. We made a literature review of known documents, guides or manuals which have been published in Cuba and in other countries and we interviewed a group of experts in this field. An initial instrument was worked out and tested in three health areas of the capital and afterwards, it was modified accordingly. The process of assessment was divided into 4 aspects: structure, process or functioning, results or usefulness and economic evaluation. To assess functioning of surveillance, we took the analysis of its attributes into account, i.e., simplicity, flexibility, acceptability, sensitivity positive predictive value, representativity, opportunity and also integrality and responsiveness capacity. We described steps to be implemented and elements to be considered. Generally, the evaluation should be carried out in a week. The cost of implementing each evaluating process is estimated at 40 Official Announcement. V Workshop of university libraries of Latin and the Caribbean. Upon finishing the evaluation, a report should be submitted with the comprehensive assessment and recommendations for the evaluated level. PMID- 11107896 TI - [Emergence of a new pathogen: Cyclospora cayetanensis in patients infected with human immunodeficiency virus]. AB - The emergence of a new pathogen Cyclospora cayetanensis as a cause of clinical disease in immunosuppressed hosts is related with prolonged, severe and highly recurrent diarrheas. This paper reports two Cuban cases of cyclosporiasis associated with infection from human immunodeficiency virus in which non sporulated oocysts of Cyclospora cayetanensis were detected in feces by modified Zielhl Neelsen's technique. The most significant clinical symptoms were chronic diarrheas and loss of body weight, with CD4 levels below 200 cells per mm3. The occurrence of severe digestive symptoms in patients with Cyclospora cayetanensi and important immune compromise backed up the concept that this pathogen may act as a new opportunistic pathogen in patients with HIV. PMID- 11107897 TI - [Giant inguinal condyloma (Buschke Lowenstein tumor) with a clinical aspect of squamous carcinoma]. AB - This paper presents a patient with multiple condyloma and a Buschke-Lowenstein tumor in the groin with clinical aspect of a squamous carcinoma. Malignity was histologically ruled out in this case. Lesions were treated by surgery and during the postoperative period, the patient was treated with interferon alpha i.m. at a rate of 9 x 10(6) UI/day three times a week for 3 weeks. One year after the treatment, the patient had not shown any relapse. PMID- 11107898 TI - [Hemorrhagic pneumonia caused by influenza virus: virological diagnosis]. AB - The paper presents the case of a female patient who was admitted to "Calixto Garcia" General Hospital with respiratory distress and hypovolemic or septic shock. She was diagnosed with viral hemorrhagic pneumonia. From the endotracheal secretion taken as a sample, the influenza virus was isolated as etiological agent, which, through the hemaglutination inhibition technique, was characterized as a strain belonging to H3N2 subtype, very similar to strain A/Johannesburg/33/94 from the antigenic viewpoint. The patient recovered satisfactorily after being treated with rivabirin. PMID- 11107899 TI - [The factors associated with noncompliance with antimalarial treatment in Ecuadorian patients]. AB - A total of 249 persons living in the northwest part of Ecuador with a clinical diagnosis of malaria confirmed by thick blood films were treated with chloroquine and primaquine according to the therapeutical system in force in the National Service for Eradication of Malaria. New clinical assessment and thick blood film were applied after 4 days in P. falciparum (n = 120) cases and after 8 days in P. vivax (n = 129) cases; patients were questioned about the compliance or non compliance with the treatment, and the reasons for their acting in either way were studied. EPI-INFO 6.04 and SPSS PC 7.0 packages served to process the information: "kind adjustment test" (bondad de ajuste) abd factorial analysis of correspondences were used. The patient who daily took his/her pills for the number of days indicated, at the established intervals and at the right time was defined as a patient complying with the drug therapy. For every 3 patients complying with treatment, there were 2 who did not; non-compliance was not significantly related to age, sex, educational level, ethnic group, urban or rural setting or level of income, but learning about seriousness of the infection did help to compliance with the therapy. The reasons for non-compliance were mainly associated with drugs (side effects/reluctancy to take drugs), with the fact of forgetting to take them and of "getting cured quickly". The profile of the patient who did not comply with treatment corresponded to male, teenager, mixed race, poor and rural setting. PMID- 11107900 TI - Analisis clinico y epidemiologico de los accidentes por mordeduras de serpientes del genero Bothrops en Venezuela [A clinical and epidemiological analysis of accidental bites by snakes of the genus Bothrops in Venezuela]. AB - Clinical register of 60 patients bitten by Bothrops snake who assisted at Leopoldo Manrique Hospital and the Institute of Tropical Medicine (HLM-IMT) in Caracas during 1996-1997 were analysed. The accident was more frequent in males (45/75%). In 32 cases (53.3%) the snake was classified and 26 were Bothrops lanceolatus, 4 Bothrops venezuelensis and 2 Bothrops atrox. Anatomic regions more frequent bitten were superior members (40/66.6%): hands (36/60%), forearm (2/3.3%), elbow (1/1.6%) and arm (1/1.6%). On inferior members (20/33.3%): legs (6/10%), feet (10/16.7%), ankle (2/3.3%), and the hip (2:3.3%). The most frequent clinical manifestations in moderate and severe cases (33 patient) were pain (100%), oedema (98%), ecchymosis (76%), blisters (20%), necrosis (12%), abscess (6%) bleeding (19%), heart failure (1/1.6%), renal failure (1/1.6%). The blood clotting was evaluated in 60 (100%) cases and it was altered in 33 (55%) patients. No deaths were recorded. PMID- 11107901 TI - [The need for the use of chromogens for quantifying Leishmania promastigotes on plates from 96 wells]. AB - The existing difficulties in the treatment of leishmaniasis justify the testing of the effect of new products on parasite forms in the search of a therapeutic alternative for the parasitosis. Given the need of establishing a method to evaluate the activity of natural synthetic products in vitro under the Cuban conditions, this paper was aimed at defining the usefulness of p nitrophenilphosphate as a chromogenic substance for quantification of parasites in plaques from 96 wells. To standardize this colorimetric method the stages of the parasite growth curve were set. The study of linearity and selection of the sample size, which was optional for these assays, showed that it was possible to obtain maximum linear determination coefficient with 20 mm. Likewise, the variation coefficient was compared with and without the chromogen and the effect of changes in culture medium on the reading of absorbances was analyzed. The set limit of quantification proved the need of using chromogen for the purposes of this paper and the general results allow to recommend this less subjective, simpler and quicker methodology to test products of interest in this field. PMID- 11107902 TI - [The production of a latex immunoglobulin conjugate for the diagnosis of Gardnerella vaginalis]. AB - To obtain an anti-Gadnerella vaginalis latex globulin, specific immunosera to reference strain (ATCC 14018) and to a clinical isolation strain pool were produced. These immunosera were characterized using PAGE-SDS and immunoblotting where a close antigenic relation between the clinical isolation strains and the reference strain was observed. The latex globulin conjugates obtained from these 2 immunosera were evaluated in vitro with a resulting level of detection of 10 ufc/mL of Gardnerella vaginalis. These two conjugates were also evaluated in clinical samples and compared with the 6 vaginalis culture and the criteria considered for diagnosing bacterial vaginosis. The anti-strain pool (II) latex globulin conjugate turned out to be more sensitive and specific than anti-G. vaginalis ATCC strain (I) latex globulin. PMID- 11107903 TI - [Toxigenic Vibrio cholerae non-01]. AB - The antimicrobial susceptibility and the presence of a heat-stable toxin were researched into 100 non-01 Vibrio cholerae strains sent by 7 different health centers to the National Reference Laboratory of Acute Diarrheal Diseases in "Pedro Kouri" Tropical Medicine Institute. The presence of 20% toxigenic non-01 Vibrio cholerae was detected, a figure substantially higher than that reported in other geographic areas, except for endemic areas. This result will make it possible to set epidemiological alert in Cuba because these strains can be infected by CTX phages (element transporting genes that encode for choleric toxin) which will give such strains an epidemic potential similar to that of the etiologic agent of cholera. The identified strains could be studied as possible cholera vaccine candidates. PMID- 11107904 TI - [The knowledge, attitudes and practice of population groups with respect to tuberculosis. 1994-1996]. AB - The development of successful tuberculosis control programs requires the people's involvement, hence a study was performed to identify knowledge, perceptions and practices of the population regarding occurrence, transmission, treatment and control of this disease. The focal group technique was used in 6 sets of persons aged 15 years and over from 6 municipalities of the City of Havana. These groups thought that tuberculosis had declined in the last ten years but had increased again in the last 2-3 years, that it was a contagious disease presenting symptoms such as cough, hemoptysis, loss of weight, fever. They considered it as a terrible, undesirable sickness associated with poverty and caught due to malnourishment, poor hygiene of the sick person and his/her relatives and smoking. Some thought that this disease was curable and other that it was not. Several other people believed that patients should be isolated in hospital whereas others stated that they could have a normal life at home, most preferred to be informed about the disease by TV and radio. It was concluded that a quantitative study should be performed based on these results. PMID- 11107905 TI - [Neutralizing antibodies to 5 HIV-1 strains in macaques immunized with the multi epitope polypeptide TAB9]. AB - Presence of neutralizing antibodies(Nab) to 5 laboratory strains of human immunodeficiency virus type-1 (HIV-1) was studied and compared in the sera of eight macaca (Macaca fascicularis) after the third and fourth immunizations with multi-epitope polypeptide TAB9, emulsified with adjuvant Montanide ISA 720. Four animals were inoculated 1 mg and the others 200 micrograms, 2 animals were used as controls and injected only with adjuvant. Although the presence of neutralizing antibodies to homologous strains of HIV-group group B was confirmed in most animals after immunization through antigen p24 capture ELISA (DAVIH Agp24, Cuba), no statistically significant differences were found neither in titers caused by antigen concentrations nor in the responses after the third and fourth immunizing doses. Controls did not develop neutralizing antibodies. PMID- 11107906 TI - [Monoclonal antibodies to dengue virus 4: the spectrum of the reactivity to 4 serotypes of dengue]. AB - These types of monoclonal antibodies 8E8, 3F7 and 1E9 to dengue 4 virus H-241 strain. These monoclonal antibodies show various patterns of reactivity to the four dengue serotypes and different antigen preparations of serotype 4 when they were tested in various serological methods. The monoclonal antibody 8E8 exhibited a specificity of serotype (D-2; by hemagglutination inhibition); subcomplex (D-2 and D-4 by immunofluorescence) and complex (by immunoperoxidase technique). It was able to neutralize by 80% homologous virus and it turned out to be the only reactive monoclonal antibody in the complement fixation test. The monoclonal 3F7 did not react to by hemagglutination inhibition, recognized serotypes D-1, D-2, D 3 and D-4 by immunofluorescence and only serotypes D1 and D4 by immunoperoxidase technique but it was unable to neutralize the homologous virus. The 1E9 antibody was reactive to serotypes D1, D-2, D-3 and D-4 only by hemagglutination inhibition and neutralized serotype D-4. All the monoclonal antibodies were able to react to various dengue antigens through an ELISA of double antibody and showed fluorescent activity against 38th pass in Beagle dog kidney culture; however, they could not react to a D-4 recombinant antigen. PMID- 11107907 TI - [Cuban epidemic neuropathy: the risk factors in the population]. AB - A cross-sectional study was conducted in 1995 in a representative sample of the Cuban population aged 15 years or over with the objective of describing prevalence and characteristics of smoking, alcohol consumption, and physical inactivity in the urban population which could have been affected by epidemic neuropathy from 1991-1993. The sampling was stratified at provincial and municipal levels and then by cluster samplings. 93% of the sample was surveyed (14 300 people). 30% of the population aged 17 years and over smoked; the highest proportion of smokers was located in 40-49 years age group; men smoked more than woman regardless of age. The prevalence of alcohol consumption was 45.2% in which Eastern provinces exceeded the domestic average with males predominating. The prevalence of physical inactivity at national level was 33%, 25.7% for males and 39.8% for females. 47.3% of males and 25.4% of females classified as physically active because of their useful extra activity. It was considered that irregular relationships between these 3 risk factors and the incidence of epidemic neuropathy at the ecological level make it think that, although they have a real influence on the determinants of the disease, other factors may also better account for the occurrence of these cases. PMID- 11107908 TI - [Entomological surveillance over Aedes (S) aegypti and other culicids in Ciudad de La Habana, Cuba 1991-1996]. AB - The results of the entomologic surveillance carried out from 1991 to 1996 in Boyeros municipality, City of Havana within the Program for Eradication of Aedes aegypti were analyzed. Data on mosquitoes fauna collected in the municipality by various sampling methods, larval survey, human bait, capture at rest and larval traps. Culex quinquefasciatus, Aedes mediovittatus and Aedes aegypti species were the most found by the different methods. It was proved that water tanks, man-made deposits, other kinds of tanks and low tanks were the most exploited resources by mosquitoes for their breeding in this municipality whereas larval surveys were the most sensitive method for detecting species since it contributed a greater variety of species. Reference is made to the search for a mechanism that help estimate adult populations from larval indices provided by surveys and the method of capture at rest is stressed as the most sensitive method for adult mosquitoes, particularly for dengue vector within the surveillance system aimed at this species. PMID- 11107909 TI - [The ecological succession of mosquito species in the town of Boyeros, Ciudad de la Habana 1994-1996]. AB - An analysis was made on the incidence of Culicidae in Boyeros municipality, City of Havana from 1994 to 1996 based on the requirements of the national program for eradication of Aedes aegypti. A total of 13 species was detected in which Aedes mediovittatus predominated during 3 years of study, followed by Culex quinquefasciatus whereas for the first time Aedes albopictus species was found in the territory. People's Councils Wajay, Santiago de las Vegas and Armada exhibited the highest values of infestation by Culicidae in the municipality, therefore, they required that anti-Aedes program, operators paid more attention to this situation. It was found that depots preferred by Culicidae for breeding were larval traps followed by low tanks in 1994 and 1995 and by other reservoirs in 1996. A mediovittatus preferred larval traps whereas Cx. quinquefasciatus preferred low tanks except in 1996 when this species was mostly found in other tanks. 93% of all test-positive reservoirs found were colonized by only one species, whereas 7% presented larvae from 2 species or more. A. mediovittatus and C. quinquefasciatus were the most related, above all in low tanks. PMID- 11107910 TI - [The diagnosis of fascioliasis of the biliary tract by imaging]. AB - Three patients with abdominal pain were studied and through ultrasonography, it was suspected that they had Fasciola hepatica in their gallbladder and choedochio. By using endoscopic retrograde cholangiopancreatography (ERCP), the presence of parasites in extrahepatic biliary passages of the cases was confirmed and then, it was taken out with a Dormia basket. It was concluded that imaging is a diagnostic means to be considered in this parasitosis. PMID- 11107911 TI - [Psoriasis and AIDS: a report of 2 cases]. AB - Even when there is not direct relation between psoriasis and AIDS, there have been reported impressive manifestations of psoriasis in AIDS diagnosis, difficulties in response to therapy, increase of serious forms of the disease, and clearing of lesions in terminal phase of AIDS. Two cases in which the two diseases are associated were presented. Both cases had outbreaking psoriasis guttata, one of them after being diagnosed with AIDS, in addition to have other dermatosis such as leukoplasia and ungual candidiasis; and the two patients also had scabies. Additionally, presentation of generalized lesions resistant to prescribed therapies was observed. PMID- 11107912 TI - [Reactive strips for the diagnosis of oxidase activity in bacteria]. PMID- 11107913 TI - [Outcasts in medical office]. PMID- 11107914 TI - [Current treatment of colorectal cancer]. PMID- 11107915 TI - [Alzheimer disease--new hope?]. PMID- 11107916 TI - [Successful cancer treatment--and so what?]. PMID- 11107917 TI - [Psychological, social and economic situation of women surgically treated for cancer]. AB - INTRODUCTION: As part of an international effort, a study of the psychosocial condition of women with cancer in Norway was performed in 1997. MATERIAL AND METHODS: 851 women with breast or gynaecological cancer in 27 Norwegian hospitals were asked to fill in a multi-choice questionnaire. RESULTS: 76% of the breast cancers were detected by the women themselves, 19% by mammography and 11% by clinical examination. Similar numbers for gynaecological cancer were 59% by the women and 41% by the doctors (13% screening smear). 25% of the women were informed about the cancer diagnosis by telephone and 9% by letter. 87% were satisfied with treatment and care, 7% dissatisfied. 62% were satisfied with the availability of doctors, 20% not. In 47% of the breast cases, therapy alternatives were discussed with the patients. In 23% of gynaecological, 50% of breast, and 72% of gynaecological cancer, patients had no wish to participate in the decision. 94 of a total of 850 patients (11%) experienced financial problems after the diagnosis had been established. More than 90% of the women had a better or unchanged relation to their partner, family and friends after treatment. However, 30% of the breast and 14% of the gynaecological cancer patients had problems with their body image, and 16% and 15% felt they were less sexually attractive. 89% felt their partner coped well with the situation, 9% had problems and only 2% lost their partner. 32% of the patients had used alternative medicine, 44% of those with recurrence and 28% under primary treatment. INTERPRETATION: Women operated for breast and gynaecological cancer face a variety of psychosocial and financial problems. PMID- 11107918 TI - [Rehabilitation of women with breast cancer]. AB - BACKGROUND: Previous studies on effects of rehabilitation programmes for women with breast cancer are rare, but promising. This study aimed to examine the physical and psychological conditions for these patients before and after a rehabilitation programme at Red Cross Haugland Rehabilitation Centre in Norway. MATERIAL AND METHODS: Included in the study were a total of 50 women, aged 31-66 (mean 49) years, who had undergone surgical treatment, chemotherapy and radiation therapy for cancer mammae stage 1 and 2 (limited to the breast only or spread to the axillary lymph nodes, respectively). They received a three-week rehabilitation programme, followed by a three-month period at home and a one-week follow-up at the rehabilitation centre. Examinations of physical and psychological status were performed before and after the three-week stay and at follow-up. RESULTS: Maximum oxygen uptake increased from 67% to 77% of predicted value, the mental status and subjective rating of life quality improved, the physical activity level increased, and 36 out of 46 subjects returned to their jobs during the three-month follow-up. The women themselves reported subjective positive effects of participating in the programme. INTERPRETATION: Although the present study was non-controlled, the positive results were so promising that further controlled studies should be encouraged, as well as rehabilitation programmes for women with breast cancer. PMID- 11107919 TI - [Childhood trauma and chronic pain]. AB - BACKGROUND: Chronic pain is a complex, multidimensional phenomenon. Medical intervention has often little effect. MATERIAL AND METHODS: The case histories of 100 patients evaluated by the multidisciplinary pain clinic team at Haukeland University Hospital in Bergen, Norway were examined. RESULTS: 44% of the patients reported serious trauma long before the development of pain. INTERPRETATION: Chronification of pain appears to be related to childhood trauma. PMID- 11107920 TI - [Postoperative wound infections in elective colorectal surgery]. AB - BACKGROUND: Surgical wound infections still represent a significant problem in colorectal surgery. We wanted to investigate the frequency of postoperative wound infections in patients who had elective colorectal surgery at Lillehammer County Hospital. By reviewing relevant literature we also wanted to assess the evidence base for using antimicrobial prophylaxis in colorectal surgery, which regimens are to be preferred, and the probable reduction in wound infections. MATERIAL AND METHODS: 99 patients were followed up at 14 days after surgery; presence or absence of wound infection was noted. Samples for culture were obtained in all patients with suspected wound infection. Literature on the subject was studied focusing on systematic reviews. RESULTS: Eight out of 99 patients (8%) had wound infections. In a systematic review of 147 studies comparing different antibiotic regimens, the overall infection rate for patients receiving antibiotic prophylaxis was 11%. INTERPRETATION: The surgical wound infection rate was comparable with the results from relevant systematic reviews. The effect of antimicrobial prophylaxis is well documented. A combination of an anaerobic and an aerobic drug should be used, but no gold standard can be identified. Two adequate regimens have been investigated in controlled studies carried out in Norway. PMID- 11107921 TI - [Borderline tumors of the ovary]. AB - Only within the last decade we have begun to fully appreciate the natural history and biologic behaviour of borderline tumours in the ovaries. In contradiction to invasive epithelial tumours, most borderline tumours are confined to the ovary(ies) (stage I). Because the prognosis of stage I serous borderline tumours is excellent, with five-year survival rates of almost 100%, some experts are advocating that this subset should be classified as benign. Although the standard treatment for older patients is abdominal hysterectomy and bilateral salpingo oophorectomy, many young patients who have not completed childbearing can be safely treated with unilateral salpingo-oophorectomy coupled with comprehensive surgical staging, thereby preserving their fertility potential. Another major controversy associated with borderline tumours is the clinical management of patients with advanced-stage disease or peritoneal implants. Many experts strongly believe that surgery is the only effective treatment for borderline tumours. Others routinely employ postoperative chemotherapy for at least some subset of patients with peritoneal implants. Several investigators have focused on DNA ploidy as a predictor of recurrence and survival, but their findings are conflicting. PMID- 11107922 TI - [Human papillomavirus as a risk factor in carcinogenesis]. AB - BACKGROUND: The causal link between Human papillomavirus (HPV) and cervical neoplasia has been established through clinical, epidemiological and molecular studies during the last two decades. In 1995, HPV 16 and 18 were classified as human carcinogens. MATERIALS AND METHODS: This paper discusses the aetiological role of HPV infection in carcinomas of the anogenital region, the respiratory and the upper digestive tract on the basis of a literature search and the author's own research. RESULTS: Almost all patients with cervical carcinomas and precursor lesions are HPV-positive. HPV is a necessary, but not sufficient factor in carcinogenesis. Several risk factors such as other sexually transmitted diseases, sexual activity, smoking, immunosuppression and diet may be important. High prevalences of HPV-DNA have also been found in carcinomas of other anogenital regions; however, HPV does not seem to be important in prostate carcinogenesis. Recent studies have shown that HPV may be a risk factor in carcinogenesis in the respiratory and upper digestive tract, but its aetiological role has yet to be proved. INTERPRETATION: Therapeutic and prophylactic vaccination is now being developed. In a few years time they may influence the treatment and prophylaxis of cervical carcinomas, some of the most common cancers among women worldwide. PMID- 11107923 TI - [New drugs in the treatment of advanced colorectal cancer]. AB - BACKGROUND: Colorectal cancer is one of the leading cancers in industrialised countries in terms of incidence and mortality. Advanced colorectal cancer has traditionally been treated with 5-fluorouracil, alone or in combination with leucovorin. This treatment has shown response rates of 10-25%; however, little effect has been observed on survival. MATERIAL, METHODS AND RESULTS: In recent years, a number of new drugs for advanced colorectal cancer have been developed and tested. We have made a comprehensive survey of the literature and find that phase II and phase III clinical studies show improved response rates compared to the traditional use of 5-fluorouracil and leucovorin. Thymidylate-synthase inhibitors, oxaliplatine and topoisomerase inhibitors, used singly or in combination, improve response rates and show a significant effect on survival in patients with metastatic colorectal disease. INTERPRETATION: Several new drugs now available in hospital treatment of metastatic colorectal cancer show improved response and increased survival compared to traditional 5-fluorouracil regimens. PMID- 11107924 TI - [Fluorinated pyrimidines in oral treatment of advanced colorectal cancer]. AB - BACKGROUND: For several years, fluorinated pyrimidines have been used in the treatment of advanced colorectal cancer, mainly in the form of intravenous injections of 5-fluorouracil (5-FU), alone or in combination with leucovorin. Oral treatment with 5-FU has been difficult because of high toxicity and low bioavailability. MATERIAL, METHODS AND RESULTS: Increased knowledge of the metabolism of 5-FU, reviewed in this article, has led to the development of a number of new oral drugs for the treatment of advanced colorectal cancer. Administration of prodrugs and inhibitors of 5-FU-catabolic enzymes has led to stable and high levels of active drug. Several drugs have shown promising results in new clinical trials. INTERPRETATION: New 5-FU related drugs for oral administration show results comparable to those of the cytotoxic drugs that have been administered in hospital. In the future, general practitioners could possibly treat and follow up a larger proportion of these patients. PMID- 11107925 TI - [What is actually Hodgkin disease?]. AB - For long, there has been controversy over the very nature of Hodgkin's disease: Is it a true neoplasm, an inflammatory, even infectious disease, an unusual immunologic reaction, or a combination of these pathogeneses? New information has been obtained during the last few years from immunohistology, immunohistochemistry, and molecular genetics. Epidemiologic, serologic and and direct detection studies all independently point to Epstein-Barr-virus as a potential aetiologic cofactor. Clonality studies show that viral infection precedes clonal expansion of tumour cells. Single cell analysis of rearrangement of immunoglobulin genes published 1996-98 proves the B cell nature of Reed Sternberg and Hodgkin cells, and indicates their origin from a single transformed B cell and subsequent monoclonal expansion. Hodgkin's disease is a tumour of cytokine-producing and cytokine-responding cells. Different quantities of a series of cytokines may explain differences in prognosis, clinical symptoms and histopathological features. PMID- 11107926 TI - [Professional cooperation and overview of the health status. Two important initiatives from Christian August Egeberg in 1838 and 1855]. AB - Christian August Egeberg (1809-74) was a Norwegian military surgeon who practised surgery and family medicine in the capital city of Christiania and in neighbouring rural Baerum. He saw the medical profession's need for establishing a scientific community, for information-seeking and knowledge updates. He was among the founders of the Norwegian Medical Society in 1833 and in 1838 initiated a series of Scandinavian scientific conferences which lasted until 1929. In 1855 he established a notification system for contagious diseases with reports submitted every month from practising physician. The system was intended to provide a basis for surveillance of the so-called epidemic constitution, so that appropriate measures might be taken against prevailing diseases. PMID- 11107927 TI - [Individualized drug therapy]. PMID- 11107928 TI - [Why is insulin release from beta cells insufficient in type 2 diabetes?]. AB - Insulin secretion is finely tuned to tissue requirements by tight links to prevailing blood glucose levels. The normal regulation of insulin secretion is linked to glucose metabolism in the pancreatic beta-cell, a major but not exclusive signal for secretion being closure of K+ ATP-dependent channels in the cell membrane through an increase in the cytosolic ATP/ADP. Insulin secretion in type 2 diabetes is abnormal in several respects, due to genetic causes, but also due to the metabolic environment of the pancreatic beta-cells. This environment may be particularly important for the deterioration of insulin secretion, which occurs with increasing duration of diabetes. Factors of the environment with potential importance include over-stimulation, a negative effect of hyperglycaemia per se ("glucotoxicity"), and adverse effects of elevated fatty acids ("lipotoxicity"). A better understanding of the mechanisms behind these factors and of their clinical importance will pave the way for treatment which could preserve beta-cell function in type 2 diabetic patients. PMID- 11107929 TI - [Genetic and immunologic risks for development of type 1 diabetes--experiences from an intervention trial]. AB - BACKGROUND: Type 1 diabetes is a common multi-factorial disease. Recently, promising attempts have been made at preventing the disease in individuals at risk. We present our experiences as participants in an international multicentre intervention trial, the European Nicotinamide Diabetes Intervention Trial (ENDIT). The aim is to prevent prediabetic individuals from becoming overtly diabetic by the use of nicotinamide. MATERIAL AND METHODS: First degree relatives of type 1 diabetes children attending paediatric clinics in Norway were recruited. The level of islet cell antibodies (ICA) was determined. Individuals with ICA titer above 20 Juvenile Diabetes Foundation Units (JDFU) were allocated to treatment with nicotinamide or placebo in a double-blind design. In the Norwegian patients in the trial HLA-DQ genotypes were also determined. RESULTS: 56 individuals had ICA > 20 JDFU; 36 agreed to participate in the trial. Assessment of genetic and immunological risk did not seem to emotionally upset the majority of participants; so far, no serious adverse events have been observed. The final results of this trial are expected in year 2003. INTERPRETATION: Prevention of type 1 diabetes may be feasible in the future. PMID- 11107930 TI - [Diabetes among immigrants from non-western countries]. AB - Around 15% of all inhabitants in Oslo, Norway, have non-western ethnic background. In downtown Oslo the proportion is around one third. Diabetes is more frequent in non-western ethnic groups than among westerners. Treatment of diabetes requires a collaboration between the patient and the health professional and a common understanding of the nature of the disease and the treatment objectives. In the encounter with patients of foreign origin there are several cultural obstacles for compliance: language problems, illiteracy, religious fatalism and reluctance by the patients to take responsibility for his/her own disease. Lovisenberg Diakonale Sykehus in Oslo has conducted education classes for groups of Pakistani persons with diabetes. PMID- 11107931 TI - [Different motives behind criticism of mass screening for cancer of the breast and cervix uteri]. PMID- 11107932 TI - [A wide perspective on meta-analysis--may be of crucial significance for the patients]. PMID- 11107933 TI - [DNA diagnosis of asylum applicants]. PMID- 11107934 TI - [Critical shortage of internship positions?]. PMID- 11107935 TI - [Resign, Mr chairman!]. PMID- 11107936 TI - [Emergency medical service in Oslo]. PMID- 11107937 TI - [All Norwegians will become physicians soon]. PMID- 11107938 TI - [NettDoktor.no and the Tidsskriftet]. PMID- 11107939 TI - [Do we follow guidelines for the treatment of asthma in children?]. PMID- 11107940 TI - [Microdissection of chromosomes]. AB - The technique of deciphering different chromosome abnormalities by the employment of specific DNA probes prepared by microdissection of normal or abnormal chromosomes is described. It is emphasised that in some families the diagnosis is possible only by "family specific" probes. PMID- 11107941 TI - [New insights into the etiology and pathogenesis of pre-eclampsia]. AB - Pre-eclampsia is a major cause of maternal and foetal morbidity and mortality but the etiology and pathogenesis are basically unknown. Pre-eclampsia is clearly associated to the placenta (trophoblast) alone. Epidemiological observations have raised one of the most popular hypotheses of the genesis of pre-eclampsia--an immune maladaptation between mother and foetus. Markedly raised incidences are seen in blood relatives (mothers, sisters, daughters) to pre-eclamptic women. In the genetic studies, the importance of specific combinations of mother-foetus genotypes are recognized. The pathological processes involve an aberrant trophoblast-invasion of the spiral arteries, placental dysfunction, abnormal levels of cytokines and endothelial cell dysfunction, resulting in systemic and organ dysfunction. Genetic linkage studies and the use of molecular biology will probably elucidate the predisposing factors, etiology and pathogenesis of pre eclampsia in more detail. There might be several pathways ending up with the same clinical manifestations. PMID- 11107942 TI - [Utilization of anti-asthmatic drugs among Danish children in 1998]. AB - INTRODUCTION: The aim of the study was to evaluate the use of anti-asthmatics among Danish children in 1998. METHODS: Patient specific data were collected on anti-asthmatics (ATC-group R03) prescribed for children aged 0-15 years in 1998. Data included a total of 381,557 prescriptions for 139,727 individuals. RESULTS: Anti-asthmatics were prescribed for 13.9% of all Danish children on one or several occasions in 1998. The highest one-year prevalence and incidence rate of drug use was found for children aged 0-2 years. Most children were exclusively treated with either a short acting beta 2-agonist (66.7%) or an inhaled steroid (6.5%). Only 26.2% received both types of anti-asthmatics. CONCLUSION: In conclusion, anti-asthmatics were predominantly prescribed for the youngest children. Most children were exclusively treated with a short acting beta 2 agonist in 1998, which is only recommended in the case of mild intermittent asthma. PMID- 11107943 TI - [How do patients evaluate the newly introduced system of substituting prescriptions?]. AB - In 1997 a new prescription system was introduced in Denmark. The pharmacist must now substitute the prescribed drug with a cheaper version either by a generic prescription (G-substitution) or by an original prescription (O-substitution) unless the prescribing doctor indicates that substitution is not allowed in the specific case. The purpose of this study was to obtain the patients' view on the new prescription system and to identify any related problems. The investigation was based on structured interviews. The interview guide was designed as a questionnaire, which was validated and tested before use. The response rate was 82%. The study showed that 84% of the patients were satisfied with the system and 85% of the patients thought that it should continue. Eighty-three percent of the patients had tried another version of the substituted medicine earlier. Out of these, 6% had experienced more side-effects from the substituted medicine, and 10% felt that the substituted medicine had a weaker effect. There was one case of erroneous medical treatment as a consequence of the substitution system. Only few problems such as more side-effects or less effect of the substituted medicine was experienced by the patients. It can be concluded that the patients in general are satisfied with the new prescription system. PMID- 11107944 TI - [How do practitioners evaluate the newly introduced system of substituting prescriptions?]. AB - AIMS: In 1997 a new prescription system was introduced in the Danish health care system. The pharmacist must now substitute a prescribed drug with a cheaper version, either generic (G-substitution) or original (O-substitution) unless the general practitioner (GP) indicates that substitution is not allowed. The purpose of this study was to obtain the GPs' views on the system and evaluate the problems related to the system. METHODS: The study was based on questionnaires to GPs developed via qualitative interviews with the GPs and afterwards pilot tested. RESULTS: Out of 300 GPs the response rate was 80%. The study showed that 61% of the GPs were dissatisfied with the system and thought that it should be removed. There were several reasons for this: the system was incomprehensible, the introduction and information about the system was insufficient and the extra workload was too heavy. All the GPs agreed that analogue substitution (substitution between drugs with the same effect obtained by different means) was medically unjustifiable and should not be introduced. PMID- 11107946 TI - [Standardisation of surgical mesh implantation in incisional hernia]. AB - PURPOSE: To describe the relation between risk factors, postoperative complications, recurrence rates and satisfaction before and after the introduction of Stoppas technique in patients operated for incisional hernia with implantation of a surgical mesh. MATERIAL AND METHODS: The records of 27 patients undergoing an incisional hernia repair using a surgical mesh in the period 1.1.1993-30.6.1998 were reviewed, and 18 living patients were examined. RESULTS: Among the eight patients operated using Stoppas technique there were no postoperative complications but one recurrence, and among the ten patients operated without using a specific standard technique there were five with complications and four with recurrence. CONCLUSION: Operation for incisional hernia using implantation of a surgical mesh seems to require a standard surgical technique and care must be taken in patients with risk factors. PMID- 11107945 TI - [How do pharmacists evaluate the newly introduced system of substituting prescriptions?]. AB - In 1997 a new prescription system was introduced in the Danish health care system. The pharmacist must now substitute a prescription with a cheaper version of the drug (either generic or original) unless the prescribing doctor indicates that substitution is not allowed in the specific case. The purpose of this study was to evaluate problems of the system and obtain the pharmacists' views on the system. The study was based on questionnaires to a representative sample of 75 pharmacists (a quarter of Denmark's pharmacists). The response rate was 72%. Half of the pharmacists were dissatisfied with the system, which primarily was due to the excessive workload imposed. In spite of this, about half the pharmacists wanted the system to be continued, because the overall purpose of finding the cheapest drug for the patient is good. Nearly all pharmacists thought that analogue substitution (substitution between drugs with the same overall effects but obtained by different means) should not be introduced. PMID- 11107947 TI - [Hereditary antithrombin deficiency resulting in severe neonatal thrombosis]. AB - We report a severe vena cava inferior thrombosis in a mature two day-old boy. Pregnancy and birth were event free. The mother had antithrombin deficiency. During pregnancy she was treated with enoxaparin and just before delivery with 2000 IE antithrombin concentrate. At admission the child had unmeasurable antithrombin and low protein C concentrations. He was treated with antithrombin concentrate and heparin. The thrombus disappeared, and now he is on maintenance treatment with warfarin. Risk factors and treatment of inherited antithrombin deficiency are discussed. PMID- 11107948 TI - [Postoperative analgesia. A question of prioritization?]. PMID- 11107949 TI - [Picture of the month: lung abscess]. PMID- 11107950 TI - [Is there an evidence to continue standard anticoagulation treatment in hip and knee alloplasties?]. PMID- 11107951 TI - [Physician staff at the Roskilde festival]. PMID- 11107952 TI - [The process of medical dissertations. New guidelines from the medical faculty in Copenhagen]. PMID- 11107953 TI - [Interferon beta treatment of disseminated sclerosis in Denmark--current status]. PMID- 11107954 TI - [What do we know about the morbidity among immigrants and their contacts with health care in Denmark?]. PMID- 11107955 TI - [Risk of suicide, can it be significantly reduced? Two reviews with a discussion]. PMID- 11107956 TI - [Genetic research--a research field in progress]. PMID- 11107957 TI - [Investigation of patients with recurrent syncope]. PMID- 11107958 TI - [Prevalence of dementia in Denmark]. PMID- 11107959 TI - [Flumazenil and hepatic failure]. PMID- 11107960 TI - [Food and cardiac death]. PMID- 11107961 TI - [Concerning the use of flaxseed as food supplementation]. PMID- 11107962 TI - [Risks for the recipients of donated blood or donated organs]. PMID- 11107963 TI - [Disease among immigrants and refugees]. PMID- 11107964 TI - [Rickets in young immigrants]. PMID- 11107965 TI - [Vitamin D deficiency among immigrants]. AB - Vitamin D deficiency in immigrants has been known in the UK for 30-40 years. In Denmark, we have become aware of the problem only recently. Of 69 randomly chosen Palestinian women living in Denmark 85% were found to have very low levels of 25 hydroxyvitamin D (< 10 nmol/l). Vitamin D deficiency is caused by inadequate exposure to sunlight and a low dietary content of vitamin D and calcium. Typical symptoms are muscle pain, muscle spasms, diminished muscular strength, deep bone pain, and paraesthesias. The diagnosis can be tested by three blood tests: serum 25-hydroxyvitamin D, serum PTH, and serum alkaline phosphatase. If a combination of low 25-hydroxyvitamin D and secondary hyperparathyroidism is found, the treatment should be high-dose ergocalciferol or cholecalciferol (100,000 IU weekly). If only (isolated) low 25-hydroxyvitamin D is found, treatment with 1000 IU of ergocalciferol or cholecalciferol in combination with one gram of calcium daily will be adequate. PMID- 11107966 TI - [Zoonoses among immigrants]. AB - During the last ten years Denmark has received an increasing number of immigrants, especially from the Balkans, the Middle East, and Somalia. Some of these may suffer from the zoonoses occurring in their country of origin. But apart from echinococcosis, zoonoses in these immigrants do not seem to pose a quantitatively greater problem than in Danes who have visited these areas. However it is important to have a knowledge of the symptoms and mode of transmission of zoonoses occurring in areas where the immigrants come from. PMID- 11107967 TI - [Leishmaniasis]. AB - Leishmania parasites are obligate intracellular protozoa, that produce clinical pictures, ranging from localised, self-healing ulcers to systemic, lethal diseases. The diseases caused by the parasites can be divided into cutaneous, mucocutaneous, and visceral leishmaniasis. Recovery from the infection often leaves lifelong immunity. Leishmaniasis may occur in individuals who have been to the Mediterranean countries, the countries on the Horn of Africa, the Arabian Peninsula, parts of Asia, and South and Central America. Co-infection of Leishmania parasites and HIV is a special problem. Leishmaniasis can be treated with pentavalent compounds of antimony, but other drugs, including amphotericin B, are also affective. PMID- 11107968 TI - [Diseases among refugee and immigrant children]. AB - Foreign adopted children and children of asylum applicants and refugees, newly arrived in Denmark, often have lived under conditions that make the following diagnostic considerations relevant: scabies, lice, impetigo and fungal skin infections, nutritional iron deficiency or bleeding, anaemia caused by hook worms in the gastrointestinal tract, malaria, tuberculosis, hepatitis B, HIV infection and various intestinal parasites. Haemoglobinopathies including sickle cell anaemia and talassaemia should also be kept in mind in anaemia. Immigrant children are admitted to hospital approximately twice as frequently as Danish children but with the same diagnoses apart from some increased frequency of psychological and behavioural disturbances and talassaemia. PMID- 11107969 TI - [Parasitologic examination of feces--how to interpret the question?]. PMID- 11107970 TI - [Immigration and tuberculosis. Can tuberculosis among immigrants be reduced?]. PMID- 11107971 TI - [Hepatitis B among immigrants]. PMID- 11107972 TI - [Mental disease among refugees and immigrants]. PMID- 11107973 TI - [Type 2 diabetes among immigrants]. PMID- 11107974 TI - [Eosinophilia among immigrants]. PMID- 11107975 TI - [Malaria--immunity gaps and how to stop them]. PMID- 11107976 TI - [Sleeping sickness is awake again]. PMID- 11107977 TI - [Paracetamol poisoning among immigrants in a department of hepatology]. AB - INTRODUCTION: An increased incidence of suicides and suicidal behaviour among immigrants has been described in other countries. In Denmark, misuse of paracetamol is suspected in some foreign-born minority groups, although no data have been produced to substantiate this suspicion. MATERIALS AND METHODS: A retrospective study of the incidence of paracetamol poisoning in patients admitted to a specialised department of hepatology from 1994 to 1999 was carried out. RESULTS: Of a total of 580 patients, 56 (9.7%) were immigrants, among whom a significant overrepresentation was found of immigrants from Turkey, Iran, Pakistan, and Lebanon (observed/expected ratios of 1.95, 4.14, 2.67, and 2.45 respectively). The immigrants differed from the Danish-born patients in that they were younger (21 vs 35 years of age), had a lower level of alcohol consumption (3% vs 30% with regular alcohol abuse), and were in general less severely intoxicated (3% vs 22% developing hepatic encephalopathy). Compared to the Danish born patients, the immigrants more frequently stated socio-economic problems as the reason for their self-poisoning (29% vs 10%). DISCUSSION: The study demonstrates an overrepresentation of immigrants among patients admitted with paracetamol poisoning in Denmark. PMID- 11107978 TI - [Hepatitis A in Denmark. Notified cases 1996-1999]. AB - INTRODUCTION: The aim of the study was to describe the incidence of hepatitis A in Denmark with the emphasis on the role of immigrants in relation to transmission and prevention of the disease. METHOD: A retrospective study of notified cases of hepatitis A during the period, 1.1.1996-31.12.1999. RESULTS: A total of 398 notified cases were examined, 45% of which occurred in immigrants. The average incidence per year was 13.2 per 100,000 for immigrants and 1.1 per 100,000 for Danes. The incidence for immigrants from Pakistan and Turkey was 4-5 times that for immigrants as a whole. The median age in immigrants was eight years and in Danes 29 years. Immigrants were hospitalised in 35% of the cases. Of children below ten years of age 31% were admitted. Danes were hospitalised in 43% of the cases, and of children below ten years of age 44% were admitted. Infection was acquired abroad for 71% of immigrants, 49% of whom visited Pakistan or Turkey. Of those infected in Denmark, person-to-person transmission was the most common mode of infection for both groups. Immigrants who had been travelling to endemic areas were involved in 21 of 34 outbreaks. CONCLUSION: The incidence of hepatitis A in Denmark seems to relate highly to the children of immigrants, who come from high endemic areas. Vaccination of those above one year of age is recommended when travelling abroad if anticipated. Economic compensation could be considered. PMID- 11107979 TI - [Sexually transmitted infections among immigrants in Denmark. Is it a problem?]. AB - INTRODUCTION: The aim of the study was to assess the incidence of sexually transmitted infections (STIs) in immigrants in Denmark. MATERIAL AND METHODS: Analysis of surveillance data from 1.8.1990-31.5.2000 for HIV infection and from 1.1.1994-31.5.2000 for gonorrhoea and syphilis. National data on the population mainly per 1.1.1998 were used. RESULTS: Overall, 28% of the notified, newly diagnosed HIV-infected persons were foreigners, a proportion which was 18% for both cases with gonorrhoea and syphilis; 488 (64%) were from Africa, whereas 110 (14%) were from Europe. The estimated annual incidence of first-time diagnosed HIV infection was five times higher in immigrants than in native Danes (22.3 vs 4.1 per 10(5)), namely 161.8 per 10(5) in Africans and 5.6 per 10(5) in Europeans. Similar differences were found for gonorrhoea (6.87 vs 2.14 per 10(5)) and syphilis (1.66 vs 0.16 per 10(5)). In addition, the annual incidence of newly diagnosed HIV was three times higher in male and 16 times higher in female immigrants than in Danes. The incidence of both gonorrhoea and syphilis in Danes was five times higher in men than in women, which was also found for gonorrhoea in immigrants. In immigrants, no real difference in the incidence of HIV and syphilis was found between the two genders. CONCLUSION: The incidence of diagnosed HIV infection, gonorrhoea, and syphilis in Denmark is generally low, but fairly high in certain groups of immigrants. Information, early diagnosis and treatment are central elements in the prevention of STIs and should be adapted for new sub-populations. PMID- 11107980 TI - [Incidence of methicillin-resistant Staphylococcus aureus among Kosovar-Albanian refugees at the refugee-center in Randers]. AB - INTRODUCTION: With the emergence of the war in Kosova, Europe faced a massive problem dealing with the refugees. The Danish quota was 3,000 refugees. Their health care was organised by the Danish Red Cross in collaboration with the District Hospital of Randers (DHR), the University Hospital of Arhus, and the Psychiatry Unit of the County of Arhus. The aim of the present retrospective study was to describe the prevalence of MRSA in this group of refugees. MATERIALS AND METHODS: Nasal screening culture for MRSA was performed on the first 50 refugees arriving at Randers. On admission to the DHR, the Kosovar-Albanian refugees were isolated until the MRSA culture showed negative. RESULTS: MRSA causing serious nosocomial infections has become a major problem in hospitals worldwide, with a higher incidence in the southern part of Europe than in Denmark. The initial nasal screening revealed no MRSA positive cultures. During the course of the subsequent 14 months, we found eight Kosovar-Albanian refugees infected with/colonised by MRSA (Table 1). We observed no spread of MRSA to other patient groups. CONCLUSION: We conclude that 1) the results of the initial screening of 50 refugees did not predict the succeding high incidence of MRSA; 2) the usual treatment with mupirocin nasal ointment and chlorhexidine wash did not prevent either reinfection or spread of MRSA in the refugee centre; 3) the rigorous isolation and screening strategy at DHR prevented the spread of MRSA to other patients and staff. PMID- 11107981 TI - [Ritual circumcision. A medicosocial problem]. AB - Circumcision is one of the oldest and most common operations, which has been practised for thousands of years by Moslems, Jews and various tribes in Africa, America, and Australia. Unfortunately, complications may occur during and after circumcision, ranging from trivial to tragic. Our investigation shows a higher incidence of complications when performed by a non-qualified surgeon. We therefore recommend that the operation should only be done by a surgeon in the public health service. PMID- 11107982 TI - [Skin diseases among refugees and immigrants. Four exotic case reports]. AB - The influx of immigrants from outside the Western world, has led to a wider spectrum of dermatological diseases seen by doctors in Denmark. We present four case histories, in which the disease was brought to Denmark from the patient's land of origin. Tropical diseases may present as a skin disease as such, or a generalised disease with skin manifestations, the commonest signs being ulcers, papules, exanthema, changes in pigmentation, and itching. PMID- 11107983 TI - [Severe rickets in puberty]. AB - Two adolescent brothers of Palestinian origin are described. They had lived an extreme indoor life for years, had low vitamin D intake and hypocalcaemia. Despite longstanding disabling symptoms the diagnosis was delayed. It is emphasized that rickets should be considered in such patients. PMID- 11107984 TI - [Osteomalacia--a frequently overlooked condition among refugees and immigrants]. AB - A case of osteomalacia due to vitamin D deficiency in a 25 year old female immigrant from Pakistan is described. Symptoms consisted of low back and hip pain, associated with proximal muscle weakness and waddling gait. Laboratory evaluation revealed low 25 hydroxy vitamin D, secondary hyperparathyroidism and high serum levels of alkaline phosphatases. PMID- 11107985 TI - [Thoracic hydatic cysts. An important differential diagnosis to tuberculosis among immigrants]. AB - Hydatid cysts, caused by Echinococcus granulosis, most commonly occur in the liver as a space-occupying lesion; the second most common are lung cysts, whereas rupture into the pleural space occurs very rarely. A case of intra-thoracic hydatid cysts with invasion of the pleural space and rupture into the thoracic muscle is described in a young immigrant from Morocco to Europe (Denmark). Hydatid cyst is an important differential diagnosis from tuberculosis in immigrants from areas with endemic echinococcosis and tuberculosis, such as North Africa and the Middle East. PMID- 11107986 TI - [Takayasu's arteritis. A rare differential diagnosis in fever of unknown origin]. AB - We report a case of Takayasu's arteritis as a cause of fever of unknown origin in a woman from Somalia. In patients with fever of unknown origin, it is important to consider the rare differential diagnoses and repeat physical examination despite easy access to high-technological examination methods. PMID- 11107987 TI - [Tuberculosis in immigrants]. AB - Three cases of tuberculosis in immigrants are described here. The manifestations of tuberculosis in this group are often different from those seen in Danes--in casu tuberculosis of the lymph nodes, skin, bone, and intestines. Diagnosis is difficult, as Danish doctors are not familiar with the clinical picture. Language problems are common and often necessitate the presence of an interpreter. Immigrants tend to move around without notifying the responsible doctor and the resources needed to find them may not be readily available. These factors may cause a considerable delay--from months to years--in the diagnosis and treatment. Doctors examining immigrants from Africa and Asia should be aware of tuberculosis as a possible cause of disease. Secretion and biopsies from fistulae, chronic ulcerations, lymph nodes, and abscesses should always be cultured for Mycobacterium tuberculosis. The Mantoux reaction can be misleadingly negative in patients suffering from severe tuberculosis. Identification of Mycobacterium tuberculosis is not necessary for treatment--that decision is made on the clinical picture. On the other hand material for a bacteriological diagnosis should always be obtained, because of possible resistance problems. PMID- 11107988 TI - [Tuberculous osteomyelitis in children--a problem of immigrants?]. AB - The incidence of tuberculosis in Denmark has increased in recent years, owing to immigrants from, primarily, Somalia. Bone and joint TB are mainly seen in young immigrants and a case of tuberculous ostitis of the tarsal bones and surrounding soft tissue is described. PMID- 11107989 TI - [HIV-positive refugee from Africa with low compliance. Interview about reasons for treatment failure]. AB - Strict compliance is required for a sustained response to antiretroviral therapy for HIV. Several studies from the US have found that African-Americans are less compliant and are more liable to treatment failure than caucasians. We describe a young, male refugee from Africa with treatment failure after non-compliance. A qualitative interview shows that his main reason for non-compliance is fear of disclosure and stigmatization. There seems to be a need for research into the reasons for non-compliance among vulnerable groups and targeted interventions. PMID- 11107990 TI - [Travel prophylaxis among immigrants]. AB - A female immigrant and her child acquired Plasmodium falciparum infection during a visit to her home country. They did not receive chemoprophylaxis during or after the journey. Immigrants comprise a high risk group for malaria as well as typhoid fever and hepatitis A. Increased attention towards this group of travellers is recommended. PMID- 11107991 TI - [Familial Mediterranean fever with pseudodominant inheritance]. AB - Familial Mediterranean fever (FMF) is an autosomal, recessively inherited disease, mainly affecting patients from the Mediterranean basin. Owing to the recessive transmission, the disease in most of the affected families only occurs in the members of one generation. However, high consanguinity rates in populations with carrier frequencies as much as 1:5 may account for the occurrence of FMF in two or more successive generations, so-called pseudodominant inheritance. We report a case of pseudodominant inheritance in a Turkish family living in Denmark. PMID- 11107992 TI - [Familial Mediterranean fever in a 26-year old Lebanese man]. AB - FMF is a hereditary disorder characterised by periodic fever and acute abdominal, chest, or joint pain. In the long term, amyloidosis may develop and eventually result in kidney failure. A 26-year-old man from Lebanon was diagnosed with FMF by genetic testing and treated with colchicine for two months. Colchicine reduced the frequency, duration, and intensity of his attacks, and thus minimised the risk of amyloidosis developing. PMID- 11107993 TI - [Treatment of familial Mediterranean fever. A not so rare diagnosis in Denmark]. AB - Familial Mediterranean fever (FMF) is characterised by recurrent episodes of fever and painful serositis. It is inherited as an autosomal recessive disease, predominantly in people from the Eastern Mediterranean area. The patient described here was followed-up for 6 years. He had FMF diagnosed primarily on clinical grounds, which is now verified by a gene test. PMID- 11107994 TI - [Circumcision performed by a hairdresser--formerly a circumciser]. AB - A case of circumcision undertaken by the newborn boy's grandfather is described. PMID- 11107995 TI - [Picture of the month. Leprosy]. PMID- 11107996 TI - ["Help me I'm to be married". Psychological cause of erectile dysfunction and its treatment]. PMID- 11107997 TI - [Ebola fever]. PMID- 11107998 TI - [How is health examination of adult asylum applicants arranged in Denmark?]. PMID- 11107999 TI - [Unproven therapy]. PMID- 11108000 TI - Progress towards interrupting indigenous measles transmission in the western hemisphere. PMID- 11108001 TI - Malaria. New technologies. PMID- 11108002 TI - Managing patients with chronic bronchitis: from primary prevention to advance directives. PMID- 11108003 TI - Slow pulse in an elderly woman. PMID- 11108004 TI - Intervening in atherogenesis: lessons from diabetes. AB - Age-related atherogenesis resembles diabetes, a model in which glucose looms as an important culprit. Within the body, the sugar becomes a glue able to harden and obstruct blood vessels while also activating such cells as macrophages and T lymphocytes. A new discovery is that large quantities of glycation endproducts may arrive preformed in foods prepared by routine cooking methods. PMID- 11108005 TI - Treating acute exacerbations of chronic bronchitis. AB - The value of medical therapy for acute exacerbations of chronic bronchitis generally increases with the severity of the episode. Antibiotics have a definite but limited benefit in severe cases. Systemic corticosteroids are valuable during hospitalization, but not after the patient has been discharged. Once recovery has occurred, pulmonary rehabilitation can improve quality of life. PMID- 11108006 TI - Appropriate use of carotid endarterectomy. AB - When stroke-threatening symptoms derive from the extracranial portion of a carotid artery, endarterectomy becomes a consideration. The available evidence indicates clear benefit for patients with a severe symptomatic stenosis. For asymptomatic carotid disease, the risk of a surgical complication may contraindicate the procedure. PMID- 11108007 TI - Respiration of arsenate and selenate by hyperthermophilic archaea. AB - A novel, strictly anaerobic, hyperthermophilic, facultative organotrophic archaeon was isolated from a hot spring at Pisciarelli Solfatara, Naples, Italy. The rod-shaped cells grew chemolithoautotrophically with carbon dioxide as carbon source, hydrogen as electron donor and arsenate, thiosulfate or elemental sulfur as electron acceptor. H2S was formed from sulfur or thiosulfate, arsenite from arsenate. Organotrophically, the new isolate grew optimally in the presence of an inorganic electron acceptor like sulfur, selenate or arsenate. Cultures, grown on arsenate and thiosulfate or arsenate and L-cysteine, precipitated realgar (As2S2). During growth on selenate, elemental selenium was produced. The G+C content of the DNA was 58.3 mol%. Due to 16S rRNA gene sequence analysis combined with physiological and morphological criteria, the new isolate belongs to the Thermoproteales order. It represents a new species within the genus Pyrobaculum, the type species of which we name Pyrobaculum arsenaticum (type strain PZ6*, DSM 13514, ATCC 700994). Comparative studies with different Pyrobaculum-species showed, that Pyrobaculum aerophilum was also able to grow organotrophically under anaerobic culture conditions in the presence of arsenate, selenate and selenite. During growth on selenite, elemental selenium was formed as final product. In contrast to P. arsenaticum, P. aerophilum could use selenate or arsenate for lithoautotrophic growth with carbon dioxide and hydrogen. PMID- 11108008 TI - Comparative analysis of the whole set of rRNA operons between an enterohemorrhagic Escherichia coli O157:H7 Sakai strain and an Escherichia coli K 12 strain MG1655. AB - Two primer sets for direct sequence determination of all seven rRNA operons (rrn) of Escherichia coli have been developed; one is for specific-amplification of each rrn operon and the other is for direct sequencing of the amplified operons. Using these primer sets, we determined the nucleotide sequences of seven rrn operons, including promoter and terminator regions, of an enterohemorrhagic E. coli (EHEC) O157:H7 Sakai strain. To elucidate the intercistronic or intraspecific variation of rrn operons, their sequences were compared with those for the K-12 rrn operons. The rrn genes and the internal transcribed spacer regions showed a higher similarity to each other in each strain than between the corresponding operons of the two strains. However, the degree of intercistronic homogeneity was much higher in the EHEC strain than in K-12. In contrast, promoter and terminator regions in each operons were conserved between the corresponding operons of the two strains, which exceeded intercistronic similarity. PMID- 11108009 TI - Characterization of acid phosphatase activities in the equine pathogen Streptococcus equi. AB - Acid phosphatases hydrolyse phosphomonoesters at acidic pH in a variety of physiological contexts. The recently defined class C family of acid phosphatases includes the 32 kDa LppC lipoprotein of Streptococcus equisimilis. To define further the distribution of acid phosphatases in the genus Streptococcus we have examined the equine pathogens Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus. Whole cell assays indicated that these organisms possess two acid phosphatases with activity optima at pH 5.0 and pH 6.0-6.5 and that only the former of these was, like LppC, resistant to EDTA. Western blotting with a polyclonal anti-LppC antiserum revealed the presence of a cross-reactive 32 kDa protein in both organisms. The cross-reactive protein in S. equi was shown to be a surface accessible lipoprotein as its processing was inhibited by the antibiotic globomycin and it was released from whole cells by treatment with trypsin. The presence of DNA sequences homologous to the S. equisimilis lppC gene were confirmed by PCR. These data strongly suggest that Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus produce a lipoprotein acid phosphatase homologous to LppC of S. equisimilis. PMID- 11108010 TI - Identification of methionine-processed HPr in the equine pathogen Streptococcus equi. AB - Using preparative electrophoresis, a low molecular weight protein has been partially purified from a cell extract of the equine pathogen Streptococcus equi susp. equi. N-terminal sequence analysis and Western blotting revealed the protein to be HPr, a central component of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). Interestingly, the only form of the HPr protein detected in S. equi was one with the amino-terminal methionine removed, a modification that has previously been associated with surface localization of streptococcal HPr proteins. PMID- 11108011 TI - The discovery of enfumafungin, a novel antifungal compound produced by an endophytic Hormonema species biological activity and taxonomy of the producing organisms. AB - In a screening of natural products with antifungal activity derived from endophytic fungi, we detected a potent activity in a culture belonging to the form-genus Hormonema, isolated from leaves of Juniperus communis. The compound is a new triterpene glycoside, showing an antifungal activity highly potent in vitro against Candida and Aspergillus and with moderate efficacy in an in vivo mouse model of disseminated candidiasis. The agent is especially interesting since its antifungal spectrum and its effect on morphology of Aspergillus fumigatus is comparable to that of the glucan synthase inhibitor pneumocandin B,,, the natural precursor of the clinical candidate MK-0991 (caspofungin acetate). An additional search for other Hormonema isolates producing improved titers or derivatives resulted in the isolation of two more strains recovered from the same plant host showing identical activity. The producing isolates were compared with other non producing Hormonema strains by DNA fingerprinting and sequencing of the rDNA internal transcribed spacers. Comparison of rDNA sequences with other fungal species suggests that the producing fungus could be an undetermined Kabatina species. Kabatina is a coelomycetous genus whose members are known to produce Hormonema-like states in culture. PMID- 11108012 TI - Paenibacillus granivorans sp. nov., a new Paenibacillus species which degrades native potato starch granules. AB - From a native potato starch-degrading enrichment culture, strain A30 had been isolated and had tentatively been identified as a member of the Bacillus firmus/lentus group (Wijbenga et al. Appl. Microbiol. Biotechnol. 35, 180-184, 1991). In this paper the isolate A30 is further characterized using phylogentic analysis of the 16S rDNA and determination of a number of additional phenotypic characteristics. These data are compared to those of Paenibacillus amylolyticus, P. chibensis, and P. thiaminolyticus. It is concluded that strain A30 is a new Paenibacillus species, for which the name Paenibacillus granivorans is suggested. PMID- 11108013 TI - 16S rDNA sequence analysis of Xylella fastidiosa strains. AB - The 16S rDNA encoding the small subunit ribosomal RNA were amplified by PCR, cloned, and sequenced from 16 strains of Xylella fastidiosa originating from nine different hosts. In pair-wise comparisons, X. fastidiosa strains showed a maximum variation of 1.0% or 14 nucleotide positions. When all 16 sequences were considered as a set, 54 variable positions were found. Analysis of the sequence data indicated that the X. fastdiosa strains formed three rDNA groups. Group one includes Pierce's disease and mulberry leaf scorch strains; Group two, periwinkle wilt, plum leaf scald, phony peach, oak leaf scorch, and elm leaf scorch strains; and Group three, citrus variegated chlorosis and coffee leaf scorch strains. All X. fastidiosa strains exhibited significantly higher levels of sequence heterogeneity (63 to 83 nucleotide positions) when compared to species from Xanthomonas and Stenotrophomonas. Our data demonstrate that 16S rDNA sequence data could provide valuable information for future classification of X. fastidiosa at the sub-species level. PMID- 11108014 TI - Phylogenetic evidence for novel and genetically different intestinal spirochetes resembling Brachyspira aalborgi in the mucosa of the human colon as revealed by 16S rDNA analysis. AB - Intestinal spirochetes (Brachyspira spp.) are causative agents of intestinal disorders in animals and humans. Phylogenetic analysis of cloned 16S rRNA genes from biopsies of the intestinal mucosa of the colon from two Swedish 60-years old adults without clinical symptoms revealed the presence of intestinal spirochetes. Seventeen clones from two individuals and 11 reference strains were analyzed and the intestinal spirochetes could be divided into two lineages, the Brachyspira aalborgi and the Brachyspira hyodysenteriae lineages. All of the clones grouped in the B. aalborgi lineage. Moreover, the B. aalborgi lineage could be divided into three distinct phylogenetic clusters as confirmed by bootstrap and signature nucleotide analysis. The first cluster comprised 6 clones and the type strain B. aalborgi NCTC 11492T. The cluster 1 showed a 16S rRNA gene similarity of 99.4 99.9%. This cluster also harbored the only other strain of B. aalborgi isolated so far, namely strain W1, which was subjected to phylogenetic analysis in this work. The second cluster harbored 9 clones with a 98.7 to 99.5% range of 16S rDNA similarity to the B. aalborgi cluster 1. Two clones branched distinct and early of the B. aalborgi line forming the third cluster and was found to be 98.7% similar to cluster 1 and 98.3-99.1% to cluster 2. Interestingly, this shows that considerable variation of intestinal spirochetes can be found as constituents of the colonic microbiota in humans, genetically resembling B. aalborgi. The presented data aid significantly to the diagnostic and taxonomic work on these organisms. PMID- 11108015 TI - Polyphasic characterization of poly-3-hydroxybutyrate-co-3-hydroxyvalerate (p(HB co-HV)) metabolizing and denitrifying Acidovorax sp. strains. AB - For the purpose of denitrification in small drinking water plants, a bacterial mixed population was isolated from a packed bed column bioreactor with poly-3 hydroxybutyrate-co-3-hydroxyvalerate (P(HB-co-HV)) as a substrate for the denitrification of ground water (10 degrees C). Isolates 2nIII from the mixed culture, with the ability to denitrify and metabolize P(HB-co-HV), were used as starter cultures for the elimination of nitrate in ground water. The strains were characterized by diverse techniques. Classical phenotypic studies lead to rRNA group III of the genus Pseudomonas. Results obtained by molecular techniques demonstrated that the 2nIII strains are members of the Comamonadaceae and shows similarities to the genus Acidovorax. However, an integration of the 2nIII isolates within one of the known Acidovorax species is not possible for the moment. The 2nIII starter cultures clustered close to Av. temperans according to their whole cell proteins and fatty acids, whereas in DNA/DNA hybridization no significant DNA binding (< 25%) was found. In contrast a significant but low degree of DNA/DNA hybridization was found between the 2nIII strains and Av. facilis and Av. delafieldii. Our polyphasic results lead to the conclusion that the 2nIII strains may constitute a separate Acicdovorax species. PMID- 11108016 TI - Vibrio pelagius: differences of the type strain deposited at various culture collections. AB - A critical evaluation of published and own taxonomic and phylogenetic studies on Vibrio pelagius showed substantial diversity of strains received as type strains from various Culture Collections. The comparison of data based upon 16S rRNA sequence analyses, earlier genomic DNA-DNA similarity studies as well as physiological investigations and the original description indicate that Vibrio pelagius strains CECT 4202T and ATCC 25916T really represent the originally described type species whereas strains NCIMB 1900T and CIP 102762T highly likely are representatives of Vibrio natriegens. PMID- 11108017 TI - Taxonomic analysis of extremely halophilic archaea isolated from 56-years-old dead sea brine samples. AB - A taxonomic study comprising both phenotypic and genotypic characterization, has been carried out on a total of 158 extremely halophilic aerobic archaeal strains. These strains were isolated from enrichments prepared from Dead Sea water samples dating from 1936 that were collected by B. E. Volcani for the demonstration of microbial life in the Dead Sea. The isolates were examined for 126 morphological, physiological, biochemical and nutritional tests. Numerical analysis of the data, by using the S(J) coefficient and UPGMA clustering method, showed that the isolates clustered into six phenons. Twenty-two out of the 158 strains used in this study were characterized previously (ARAHAL et al., 1996) and were placed into five phenotypic groups. The genotypic study included both the determination of the guanineplus-cytosine content of the DNA and DNA-DNA hybridization studies. For this purpose, representative strains from the six phenons were chosen. These groups were found to represent some members of three different genera - Haloarcula (phenons A, B, and C), Haloferax (phenons D and E) and Halobacterium (phenon F) - of the family Halobacteriaceae, some of them never reported to occur in the Dead Sea, such as Haloarcula hispanica, while Haloferax volcanii (phenons D and E) was described in the Dead Sea by studies carried out several decades later than Volcani's work. PMID- 11108018 TI - Simultaneous identification of five bifidobacterium species isolated from human beings using multiple PCR primers. AB - On the basis of 16S rRNA sequences, 5 species-specific forward primers were designed for the identification of 5 Bifidobacterium species isolated from human intestine, namely B. bifidum, B. adolescentis, B. infantis, B. breve and B. longum. As the 5 primers targeted at different sites of 16S rDNA, by using their mixture and a genus-specific reversed primer, the 5 Bifidobacterium species can be simultaneously identified in individual or in mixed culture through PCR amplification. The specificity of the primers was confirmed by the use of genomic DNAs from type strains of all 32 Bifidobacterium species and 6 other relatives. The 5-primer mixture was also applied to the identification of Bifidobacterium strains used commercially. The results turned out to be in accordance with those from conventional identification. This multiple-primer method provides a useful tool for rapid identification of the 5 Bifidobacterium species indicated. PMID- 11108019 TI - Specific detection of bifidobacterium strains in a pharmaceutical probiotic product and in human feces by polymerase chain reaction. AB - For PCR specific detection of the strains Bifidobacterium longum Y 10, B. infantis Y 1 and B. breve Y 8 used in a new probiotic product (VSL-3), strains specific rDNA primers have been developed. Spacer regions between the 16S and 23S rRNA genes (ITS) of the three strains were amplified by PCR with conserved primers and the nucleotide sequence of these ITSs were determined. On the basis of their comparison with the rDNA sequences retrieved from GenBank, we designed new primers which specifically recognize the species B. breve and the two strains B. infantis Y 1 and B. breve Y 8. Specificity of these primers was confirmed through the analysis of 60 bifidobacteria strains belonging to the more representative human species. The feasibility of this PCR method was investigated in commercial VSL-3 product and fecal samples collected from 4 patients affected by inflammatory bowel deseases and two healthy subjects before and after the VSL 3 administration. By PCR analysis of different VSL-3 commercial batches we were successful in differentiating and quantifying the strains B. longum Y 10, B. infantis Y 1 and B. breve Y 8. B. infantis Y 1 and B. breve Y 8 could be detected at high concentration in fecal specimens of both patients and subjects treated with the probiotic preparation, showing a different colonization behaviour. Seven days after the VSL-3 treatment suspension, no patients and subjects harbored B. infantis Y 1 and B. breve Y 8, indicating a transient presence of these exogenous strains. PMID- 11108020 TI - Development of molecular RAPD marker for the identification of Pediococcus acidilactici strains. AB - A RAPD analysis performed using a single primer targeted to the pediocin AcH/PA-1 gene was carried out on several P. acidilactici strains and on some related species of lactic acid bacteria. The high degree of genetic variability detected in P. acidilactici strains did not allow the selection of a common RAPD fragment that could be chosen as a potential species-specific DNA marker. Nevertheless a 700 bp fragment, that was found to be peculiar of all potential pediocin producer strains analyzed, was cloned and sequenced with the aim to develop a species specific PCR marker. Sequence analysis of the cloned 700 bp fragment showed one putative small open reading frame (ORF1), with no significant homology with known genes, and a partial putative second coding region (ORF2) with a high degree of similarity with several methionyl tRNA synthesis (metS) genes. The two coding regions were separated by a short spacer region. Primers targeted to ORF2 plus part of the spacer region and primers designed for the amplification of the entire cloned RAPD fragment were found to be species-specific for the detection of P. acidilactici strains. Furthermore primers designed on the ORF1 sequence allowed the amplification of a 439 bp fragment only in some P. acidilactici strains, including pediocin producing strains. PMID- 11108021 TI - Ribotyping of vibrio populations associated with cultured oysters (Ostrea edulis). AB - The intraspecific variability of Vibrio splendidus, V. harveyi and V. tubiashii recovered from oysters (Ostrea edulis) collected at the Mediterranean coast near Valencia, Spain, was analyzed by ribotyping. The two former species represented the most abundant ones, and the third one was the only species described as pathogenic for oysters. A total of 115 environmental strains were studied, 84 of V. splendidus, 23 of V. harveyi and 8 of V. tubiashii. Chromosomal DNA was digested with KpnI and hybridized with an oligonucleotide probe complementary to a highly conserved sequence in the 23S rRNA gene. Ribotyping among natural populations of the three species rendered 5 to 9 bands, and showed a high genetic diversity, with a ratio no. of strains/no. of ribotypes between 1.1 and 1.5. Cluster analysis of V. splendidus ribotypes suggests a seasonal pattern of incidence, with those ribotypes corresponding to winter and spring samples being maintained in the oysters over the year. PMID- 11108022 TI - Restriction fragment length polymorphism analysis of 16S rDNA and low molecular weight RNA profiling of rhizobial isolates from shrubby legumes endemic to the Canary islands. AB - Thirty-six strains of slow-growing rhizobia isolated from nodules of four woody legumes endemic to the Canary islands were characterised by 16S rDNA PCR-RFLP analyses (ARDRA) and LMW RNA profiling, and compared with reference strains representing Bradyrhizobium japonicum, B. elkanii, B. liaoningense, and two unclassified Bradyrhizobium sp. (Lupinus) strains. Both techniques showed similar results, indicating the existence of three genotypes among the Canarian isolates. Analysis of the combined RFLP patterns obtained with four endonucleases, showed the existence of predominant genotype comprising 75% of the Canarian isolates (BTA-1 group) and the Bradyrhizobium sp. (Lupinus) strains. A second genotype was shared by nine Canarian isolates (BGA-1 group) and the B. japonicum and B. liaoningense reference strains. The BES-5 strain formed an independent group, as also did the B. elkanii reference strains. LMW RNA profile analysis consistently resolved the same three genotypes detected by 16S ARDRA among the Canarian isolates, and suggested that all these isolates are genotypically more related to B. japonicum than to B. elkanii or B. liaoningense. Cluster analysis of the combined 16S ARDRA and LMW RNA profiles resolved the BTA-1 group with the Bradyrhizobium sp. (Lupinus) strains, and the BES-5 isolate, as a well separated sub-branch of the B. japonicum cluster. Thus, the two types of analyses indicated that the isolates related to BTA-1 conform a group of bradyrhizobial strains that can be clearly distinguishable from representatives of the tree currently described Bradyrhizobium species. No correlation between genotypes, host legumes, and geographic location was found. PMID- 11108023 TI - A thermophilic Bacillus isolated from an Eolian shallow hydrothermal vent, able to produce exopolysaccharides. AB - A thermophilic aerobic microorganism, able to produce two exocellular polysaccharides (EPS1 and EPS2), was isolated from a shallow hydrothermal vent at Vulcano island (Eolian Islands, Italy). EPS1 and EPS2 were based on mannose and glucose although in a different ratio. EPS2 possessed a trisaccharide repeating unit with a manno-pyranoside configuration. New isolate phenotype was studied by physiological and morphological observations, including biochemical and antimicrobial susceptibility tests (134). Previous analyses carried out on 87 field isolates and 8 thermophilic reference bacilli displayed low phenotypic similarity level (S(SM) = 65%) with Bacillus thermodenitrificans DSM 465. Optimal growth occurs at 65 degrees C and pH 7.0. Oxidase and catalase are negative. The guanine-plus-cytosine (G+C) content of DNA is 52.7%. Genotypic investigations demonstrated the diversity of the isolate with fifteen selected thermophilic Bacillus spp. when we compared the restriction patterns of the amplified 16S rDNA. The membrane lipids are based on fatty acids mainly belonging to the iso family. PMID- 11108024 TI - Degradation of latex and of natural rubber by Streptomyces strain La 7. AB - Streptomyces strain La 7 was isolated from the banquete of a city high way in Karlsruhe. According to partial 16S rRNA gene sequencing it was identical with Streptomyces albogriseolus and Streptomyces viridodiastaticus. DNA-DNA-similarity studies revealed 80.3-82.4% similarity between each of two of the three strains. Although phylogenetically closely related, Streptomyces strain La 7 differed from the two reference strains by morphological as well as physiological features and might represent a new species aside of S. albogriseolus and S. viridodiastaticus. The new Streptomyces strain La 7 was grown in a medium containing a latex emulsion or squares of natural rubber gloves as the only carbon source. On agar plates with a latex overlay agar, translucent halo formation around the colonies was observed. The unvulcanized latex was metabolized and the carbon from the isoprene units was apparently used for cell growth. In shake cultures with unlimited oxygen supply, during 60 days of incubation, 140 mg of the 175 mg totally emulgated latex were degraded exponentially. In sterile control flasks about 3% of the initial amount of latex could not be recovered after incubation on a shaker, presumably due to photochemical transformation. During static incubation of sterile medium, the latex formed a sticky layer at the surface of the medium and on the glass walls and recovery of the material was more difficult. Estimation of the protein content of cells from total nitrogen resulted in about 50% of the degraded latex being incorporated into cells, if a standard cell composition was assumed. Direct protein analysis according to Bradford (1976) gave much lower estimates, presumably due to a low content of aromatic amino acids. Stripes of natural rubber were degraded by Streptomyces strain La 7 during 70 days to an extent of about 30%. Scanning electron microscopy demonstrated, that hyphes of Streptomyces strain La 7 colonized and penetrated the latex surface with a concomitant deterioration of the latex material. PMID- 11108025 TI - Natural polyploidization of some cultured yeast Saccharomyces sensu stricto: auto and allotetraploidy. AB - Using genetic and flow cytometric analyses, we showed that wine strain S6U is an allotetraploid of S. cerevisiae x S. bayanus. Hybrid constitution of the strain and its meiotic segregants was confirmed by Southern hybridization analysis of their chromosomal DNAs using four S. cerevisiae cloned genes: LYS2 (chr. II), TRK1 (chr. X), ARG4 (chr. VIII), ACT1 (chr. VI) and PCR/RFLP analysis of the MET2 gene (chr. XIV). Monosporic progeny of strain S6U was highly viable in first generation but completely nonviable in the second one. According to the genetic analysis, sherry strain S. cerevisiae SBY 2592 was found to be an autotetraploid heterozygous for homo-heterothallism. PMID- 11108026 TI - Utilisation of differential killer toxin sensitivity patterns for fingerprinting and clustering yeast strains belonging to different genera. AB - The differential killer sensitivity of 103 yeast cultures belonging to 12 species (genera Debaryomyces, Kluyveromyces, Saccharomyces, and Zygosaccharomyces), all previously taxonomically certified by nDNA-nDNA reassociation, against a given panel of 39 killer yeasts was used as a fingerprinting tool. All strains, with the only exception of eight cultures belonging to the species Zygosaccharomyces bailii, were characterised by a specific, individual sensitivity pattern (killer formula). Cluster analysis of binary sequences based on killer sensitivity of strains belonging to different genera is presented and discussed. PMID- 11108027 TI - Atomic force microscopy in structural biology: from the subcellular to the submolecular. AB - Atomic force microscopy (AFM) is capable of generating images within ranges of resolution that are of particular interest in biology. Although atomic resolution may not be possible with biological samples, a great deal of information can still be obtained from images that provide structures at a slightly lower level of resolution. The submolecular resolution images of bacteriorhodopsin and the chaperonin GroES, which revealed, respectively, individual loops and beta-turns, confirmed and complemented other structural investigations, while the molecular level features in images of membrane-bound VacA, a cytotoxin from Helicobacter pylori, immediately suggested the possibility, subsequently proven, of channel forming ability. A series of images with macromolecular resolution directly provided details on the mechanisms by which RNA polymerase nonspecifically translocates along DNA, and images with subcellular resolving power of erythrocytic cellular membranes showed, with unambiguous clarity, linear arrays of molecular complexes. In this review, we will describe some of the most biologically relevant findings that have been obtained with AFM within ranges of resolution from the submolecular to the molecular, and from the macromolecular to the subcellular. Furthermore, we will describe some of the sample conditions and imaging environments that are likely important to achieve a particular level of resolution. PMID- 11108028 TI - Atomic force microscopy proposes a 'kiss and pull' mechanism for enhancer function. off. AB - The DNase I-hyper-sensitive sites (HS2-HS4) in the beta-globin gene enhancer region (locus control region; LCR) have been known as the target of Bach1/MafK heterodimers. We have demonstrated previously by utilizing atomic force microscopy (AFM) that Bach1/MafK mediates the formation of a looped-DNA structure in the LCR fragment. Here we perform further detailed analyses of the loop structure formed between each HSs by AFM, and propose a novel model for the enhancer/protein interaction: the Bach1/MafK heterodimer preferentially binds to HS2 with highest affinity and to HS3 with lower affinity. However, they assemble to each other to form a stable complex of four heterodimers and mediate a DNA loop formation. Once the DNA loop is formed between HS2 and HS3, the Bach1/MafK complex at the HS3 side leaves the HS3 and starts to slide along the DNA strand towards HS2 with the other side of the complex fixed at the HS2 region. This 'kiss and pull' model will contribute to understand the function of regulatory proteins at enhancer regions in terms of higher-order structure of DNA, e.g. nucleosomes and chromatin. PMID- 11108029 TI - Atomic force microscopy with carbon nanotube probe resolves the subunit organization of protein complexes. AB - Among many scanning probe microscopies, atomic force microscopy (AFM) is a useful technique to analyse the structure of biological materials because of its applicability to non-conductors in physiological conditions with high resolution. However, the resolution has been limited to an inherent property of the technique; tip effect associated with a large radius of the scanning probe. To overcome this problem, we developed a carbon nanotube probe by attaching a carbon nanotube to a conventional scanning probe under a well-controlled process. Because of the constant and small radius of the tip (2.5-10 nm) and the high aspect ratio (1:100) of the carbon nanotube, the lateral resolution has been much improved judging from the apparent widths of DNA and nucleosomes. The carbon nanotube probes also possessed a higher durability than the conventional probes. We further evaluated the quality of carbon nanotube probes by three parameters to find out the best condition for AFM imaging: the angle to the tip axis; the length; and the tight fixation to the conventional tip. These carbon nanotube probes, with high vertical resolution, enabled us to clearly visualize the subunit organization of multi-subunit proteins and to propose structural models for proliferating cell nuclear antigen and replication factor C. This success in the application of carbon nanotube probes provides the current AFM technology with an additional power for the analyses of the detailed structure of biological materials and the relationship between the structure and function of proteins. PMID- 11108030 TI - Atomic force microscopy of intact and digested collagen molecules. AB - The present study was performed to analyse the structure of non-digested and digested collagen type I molecules by atomic force microscopy (AFM). Collagen type I molecules from the bovine skin were diluted with 0.05 N acetic acid, spread on a mica plate, air-dried and observed by non-contact mode AFM in air. Collagen molecules digested with Clostridium histolyticum collagenase were also examined by AFM. Intact collagen type I molecules were observed as twisted threads ranging mainly between 280 and 310 nm in length. The surface of the molecules was uneven and both ends usually slightly bulged like a globule. Depressions on the molecules were found throughout the length, and were most prominent approximately 70 nm from one end of the molecules. The collagenase treated collagen molecules were degraded into fragments with various lengths, which corresponded to the data from sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The end of these fragments often appeared like a tuft, suggesting that the triple-helix unraveled at these regions. PMID- 11108031 TI - The application of the atomic force microscope to studies of medically important protozoan parasites. AB - Both living and fixed specimens of the medically-important parasitic protozoa, Trypanosoma cruzi, Toxoplasma gondii, Giardia lamblia, Entamoeba histolytica, and Acanthamoeba spp. were studied by atomic force microscopy (AFM). The preparation of fixed specimens was similar to methods used for either scanning or transmission electron microscopy. AFM scanning was performed using both contact and tapping modes. A classical fixation procedure utilizing glutaraldehyde followed by ethanol dehydration was not suitable for all parasite species. AFM images could not be obtained from fixed samples of T. cruzi, T. gondii or E. histolytica. However, excellent topographic images could be obtained from specimens of G. lamblia and Acanthamoeba under identical conditions. Critical point drying permitted AFM imaging of both trypomastigote and epimastigote stages of T. cruzi. Phase imaging of T. cruzi elucidated unique surface details at a level of resolution not visible using any other imaging modalities. AFM elasticity map imaging of T. cruzi-infected and T. gondii-infected cells demonstrated that both parasites were markedly firmer than the surrounding host cell cytoplasm. The parasitophorous vacuole surrounding replicating T. gondii tachyzoites was also visualized by elasticity map imaging. These data suggest that although much remains to be learned about preparing parasitic protozoa for AFM imaging, the technique has the potential of providing unique and important insights into these disease causing organisms. PMID- 11108032 TI - Changes in the surface structure of the hamster sperm head associated with maturation, in vitro capacitation and acrosome reaction: an atomic force microscopic study. AB - Structural changes of the hamster sperm head surface associated with maturation, capacitation and acrosome reaction were examined by atomic force microscopy. Spermatozoa were taken from the initial segment and distal cauda of the epididymis, washed in a modified Tyrode solution and fixed by glutaraldehyde. Some sperms taken from distal cauda epididymides were incubated with the capacitation medium before fixation. All samples were attached on the glass slide, dried in a critical point drier and observed by atomic force microscopy. The sperm head surface was characterized by the presence of numerous round particles, approximately 40 and 60 nm in diameter. The distribution and density of these particles on the sperm surface were significantly different between the equatorial segment and post-acrosomal region in each sperm, and also between sperms under different conditions. The surface of the equatorial segment was rather smooth in sperms from the initial segment of the epididymis, but had many large (60 nm) particles in sperms from the distal cauda epididymides, suggesting that the large particles were glycoproteins which were secreted from the epididymis and attached to the sperm surface during maturation. The number of these particles dramatically decreased in both capacitated acrosome-unreacted and acrosome-reacted sperms. This finding supports the idea that glycoproteins are removed from the sperm surface during capacitation. Atomic force microscopic studies of the sperm head surface are expected to be used for future molecular studies on the cell surface components involved in the mechanism of maturation, capacitation and acrosome reaction. PMID- 11108033 TI - Structural analysis of red blood cell membrane with an atomic force microscope. AB - To evaluate the usefulness of the atomic force microscope (AFM) for structural analysis of biomedical samples and to determine suitable sample preparation methods for AFM observation, the membrane of human erythrocytes prepared by various methods for electron microscopy was examined by the AFM. Strand-like elevations with 20-50 nm in width, 30-80 nm in length and 3-5 nm in height were observed, which formed networks composed of squares, pentagons and hexagons on the cytoplasmic or back surface of the erythrocyte membrane. Using colloidal gold labelled antibody, this network was found to contain spectrin molecules. Therefore it was very likely that the undercoat molecules of the plasma membrane were imaged by AFM. A large number of gentle elevations 300-400 nm in diameter and 2 nm in height were found to be distributed uniformly on the extracellular or true surface of intact erythrocyte, presumably reflecting the presence of undercoat membrane skeleton on the cytoplasmic surface. However, no structure that seemed to be derived from glycocalyces was discernible on the true surface. Structure corresponding to the unit membrane or lipid bilayer structure observable by electron microscopy was not demonstrated in the cross-section of the membrane. In freeze-fractured samples, a large number of small particles that corresponded to the intramembranous particle were also demonstrated on the membrane halves. Since AFM allows depiction of the fine structures of biological samples with very simple sample processing at a resolution comparable to or exceeding that of SEM, imaging technology using AFM can be applied to obtain biomedical information. However, several problems have to be solved in future development of the equipment. PMID- 11108034 TI - The cell biological application of carbon nanotube probes for atomic force microscopy: comparative studies of malaria-infected erythrocytes. AB - We describe the first cell biological application of carbon nanotube (CN) probes for atomic force microscopy studies. Topographic and phase images were collected from Plasmodium falciparum malaria-infected erythrocytes using both TappingMode Etched Silicon Probes (TESP probe) and CN probes. We estimate that the lateral resolution of a CN probe-generated topographic image is at least four-fold higher than that of the TESP probe. Carbon nanotube probe-generated phase images of P. falciparum-induced knobs on the surface of erythrocytes also show a markedly higher lateral resolution than comparable TESP probe-generate phase images of the same area. We conclude that CN probes are useful for cell biological atomic force microscopy studies and should play an increasingly important role in the future of this evolving discipline. PMID- 11108035 TI - Deformation of the envelope of a spherical gram-negative bacterium during the atomic force microscopic measurements. AB - A theoretical approach for the description of the deformation of the envelope of a spherical Gram-negative bacteria is presented. It is shown that the force displacement relation taken on top of bacteria is accurately approximated by a linear dependence. The bacterial rigidity is shown to be controlled mainly by its turgor pressure, while the lateral rigidity of the bacterial wall determines the distance from the tip at which the displacement vanishes. PMID- 11108036 TI - Quantitative analyses of topography and elasticity of living and fixed astrocytes. AB - The topography and elasticity of living and fixed astrocytes cultured from the rat cerebra were studied quantitatively by atomic force microscopy (AFM). Ridge like structures reflecting F-actin beneath the cell membrane were prominent in the contact-mode images of living astrocytes. Many of these ridges became unclear after fixation (2% glutaraldehyde). In addition, the ridge-like structures were invisible in the topography of living cells observed at zero-loading force in the force mapping mode, which is considered to show the real cell surface not pressed down by an AFM tip. The topography of fixed cells observed both in the contact mode and at zero-loading force in the force mapping mode was similar to that of living cells observed at zero-loading force in the force mapping mode, although some deformed areas were detected in the fixed cells. The elasticity map images of living astrocytes showed that the cell membrane above the nucleus was softer (2-3 kPa) than the surroundings, and that the cell membrane above F-actin was stiffer (10-20 kPa) than the surroundings. In the elasticity map images of fixed astrocytes, on the other hand, the elasticity of the cells was found to be relatively uniform (200-700 kPa) irrespective of the inner structures of cells. These results show that images observed by AFM should be carefully examined in consideration of the force introduced to specimens and the elasticity of specimens to find out the real surface topography. PMID- 11108037 TI - Time-lapse viscoelastic imaging of living fibroblasts using force modulation mode in AFM. AB - Using the force modulation mode in atomic force microscopy, we have succeeded in capturing time-lapse viscoelastic images of living mouse fibroblasts (NIH3T3) for several hours in a physiological condition without damaging the fibroblasts. Elongation of the lamellipodia and swelling of blebs were observed in time-lapse topographic images, which were taken every 10 min. The corresponding viscoelastic responses at a frequency of 600 Hz were visualized as consecutive images. The stiffer part of the cell body was fairly stable and did not show morphological changes for over 1 h. This is probably due to excess condensation of the actin network, hardening the cell cortex, and lowering the cytoskeletal activity. The nuclear portion of the cell body seems to be slightly less viscous than the peripheral region. PMID- 11108038 TI - Observations of xenon gas-treated barley cells in solution by atomic force microscopy. AB - Barley cells cut from a sprout were exposed to either air or high-pressure xenon gas for 3 days and the surface of those cells was observed by atomic force microscopy (AFM) to examine the effect of the gas treatment. This method enabled the direct observation of the fresh surface of the barley cells in solution at high resolution. The cuticle layer was preserved on the primary cell wall of 0.48 MPa xenon gas-treated barley cells, while air-treated barley cells lost the cuticle layer from the primary cell wall. These findings indicate that the high pressure xenon gas treatment is effective to preserve the cuticle layer attached to the primary cell wall. AFM is a powerful tool for the observation of the surface structure of living plant cells in solution. PMID- 11108039 TI - Structure of rice starch granules in nanometre scale as revealed by atomic force microscopy. AB - Atomic force microscopy (AFM) was performed for the analysis of the fine structure of rice starch granules in the nanometre scale which were prepared by a physical destruction method. The present study directly demonstrated that fine particles of approximately 30 nm in diameter were present inside each granule and occasionally formed straight chain arrangements. We considered that these fine particles correspond to the individual single cluster in the cluster model which has been proposed in previous studies on the starch granule structure. PMID- 11108040 TI - Microstructure of Cu(x)Mo6S8 Chevrel phase thin films on R-plane sapphire AB - Cu(x)Mo6S8 Chevrel phase thin films epitaxially grown on R-plane sapphire substrates have been studied using field emission scanning electron microscopy (FE-SEM) and high-resolution transmission electron microscopy (HREM). For samples grown in optimal conditions, the SEM images evidence oriented grains of about 100 nm average size; the HREM images of cross-section samples allow to deduce the local epitaxial relations between the substrates and films. HREM also evidences two grain families, previously deduced from X-ray and reflection high-energy electron diffraction studies. No defects have been observed in the films; a higher molybdenum content in the films results in Mo-precipitation at the film substrate interface. PMID- 11108041 TI - Histochemistry of food tissue by colour scanning electron microscopy. AB - A low-vacuum scanning electron microscope (SEM) allows microscopy of insulating specimens without metal coating, and so preserves the intact colour information on the specimen surface. We have attempted a new approach to characterize constituent distribution of food tissues by a histochemical method utilizing a colour SEM with an optical microscope and the low-vacuum SEM through digital image processing. To observe food tissues such as brown rice and adzuki bean, a colour SEM image of the specimen that has been stained by a modified method used in optical microscope histochemistry has proved to provide information of both the microscopic structure and constituent distribution on the specimen surface. PMID- 11108042 TI - Electron microscopy of satellite tobacco mosaic virus crystals: metal-coated, negatively stained and stereo pairs. AB - Highly purified virions of satellite tobacco mosaic virus (STMV) were found to crystallize at relatively low concentrations (300-500 microg ml(-1)) in pure water. Small crystals of these preparations were examined in the transmission electron microscope after either being rotary shadowcast with metal or negatively stained with 4% uranyl acetate. Stereo views were also obtained of both types of preparations. Stereo pairs of metal-coated crystals provided good three dimensional images. When stereo pairs of negatively stained crystals were printed from second negatives, they provided striking images although the three dimensional aspect was not so pronounced. Images of both types of preparations were compared with a computer-generated model of the virus. This model was based on data obtained in earlier X-ray diffraction crystallographic studies. Measurements of crystal axes on the EM images were somewhat lower than those of the computer model. It is assumed the reason for this is the dehydration of crystals during preparation for electron microscopy. The EM images did verify the type of crystal lattice determined in the X-ray diffraction studies. Conversely, knowing the exact unit cell parameters and the distribution of virions in the crystal from X-ray diffraction data aids in the further interpretation of electron micrographs of virus crystals. PMID- 11108043 TI - Simultaneous STM and UHV electron microscope observation of silicon nanowires extracted from Si(111) surface AB - A miniaturized scanning tunnelling microscope (STM) was fitted in a side-entry holder of an ultra-high vacuum electron microscope. The clean Si(111)7 x 7 surface was observed by both STM and reflection electron microscopy (REM) at atomic resolution. The tungsten tips were often rounded off upon tip-approach with a constant current, through a gentle touch with the sample surface. The apices of such rounded tips had radii of several tens of granometre with widths of about 3 x 3 nm. Atomically resolved STM of the Si(111)7 x 7 surface was obtainable when an atom or an atomic cluster sits on the tip surface. The rounded tips were used for fabrication of Si nanowires by the touch-and-away operation of the tip. The nanowires grew longer at higher substrate temperature and they reached as long as several tens of nanometre at 700 degrees C. The nanowire had many twins and the (111) twin lamellae were stacked in the direction of the wire axis. In another case, the twin planes were oblique to the wire axis so that the (112) direction was nearly parallel to the wire axis. PMID- 11108044 TI - Observation and nucleation control of Ge nanoislands on Si(111) surfaces using scanning reflection electron microscopy AB - Using a high-resolution scanning reflection electron microscope with multifunctions, we investigate Ge nucleation processes on clean Si(111) surfaces and form Ge islands on them by controlling step arrangements of Si(111) surfaces and by using focused electron beam (EB)-induced surface reactions. It is found that three-dimensional (3D) Ge islands grow selectively at step band areas on the surfaces without growth of the islands at terrace areas. Three-dimensional Ge nanoislands are formed at given points by stimulating the Ge wetting layer using focused electron beams and scanning tunnelling microscopy. Ge nanoislands are also formed by depositing Ge on Si windows in ultrathin SiO2 films and subsequent annealing of the sample. The islands are formed only at the window positions. These results imply new methods for forming Ge quantum dots or nanostructures at given areas. PMID- 11108045 TI - Control of island formation on silicon surfaces using ultra-high-vacuum scanning electron microscopy AB - In situ scanning electron microscopy has been used to control Au island formation on a patterned Si(111) surface with a periodic array of atomic-step bunches and holes. Liquid phase Au-Si islands were observed to redistribute on the patterned surface by annealing. The islands accumulate at a particular position of the step bunch in each pattern unit. This phenomenon is interpreted in terms of the energetic stability of a droplet on a patterned surface. PMID- 11108046 TI - Atomic resolution Z-contrast imaging of semiconductors AB - The direct interpretability of atomic resolution Z-contrast images obtained from a scanning transmission electron microscope (STEM) makes this imaging technique particularly powerful for the analysis of interfaces and defects in semiconductor materials and devices. In this paper, the principles of the technique are outlined and representative examples of its use are presented. In particular, we show the use of Z-contrast imaging to determine the polarity of a CdTe film grown on a Si substrate, the atomic structures of stacking faults and threading dislocation cores in GaN, and the atomistic structure of an ohmic metal/semiconductor contact of Au/GaAs. PMID- 11108047 TI - Quantitative HRTEM analysis of semiconductor quantum dots AB - Elastic strains and layer compositions of semiconductor quantum dots are quantified by the measurement of lattice fringe spacings from high-resolution micrographs. Analyses of simulated images, taking thin-specimen relaxation into account by application of finite element simulations, demonstrate that the local slopes of these strain profiles may contain severe artefacts mainly caused by local crystal tilts. Nevertheless, average strain values may be measured with sufficient accuracy and can be used to obtain an estimate on average layer compositions by application of the continuum theory of elasticity when analysing experimental micrographs. Focusing on In(x)Ga1-xAs/GaAs and Ge(x)Si1-x/Si heterostructures, it is demonstrated that elastic strains of nanoscale coherent islands are severely decreased due to an elastic relaxation mechanism compared to fully strained two-dimensional layers. The analysis of self-assembled quantum dots and two-dimensional wetting layers buried by capping layers gives clear evidence for a substantial reduction of the lattice strains compared to the values expected for the nominal layer stoichiometries. This observation originates most presumably from a compositional intermixing during epitaxial growth. PMID- 11108048 TI - Fluctuation microscopy: a probe of atomic correlations in disordered materials AB - Fluctuation microscopy is a new technique to study medium-range structure in disordered materials. Low-resolution dark-field transmission electron microscope images are interpreted as spatially resolved diffraction maps. Statistical analysis of intensity variations in these maps provides information about higher order atom position correlation functions. Here we give a qualitative description of the technique and explore the properties and utility of higher-order correlation functions in describing the structure of disordered materials. PMID- 11108049 TI - Near-edge conduction band electronic states in SiGe alloys AB - Spatially resolved electron energy loss spectroscopy (EELS) measurements in GeSi alloys illustrate the relationship of atomic structure to local electronic structure. Extending earlier measurements, where electronic structure was found to be controlled by composition in relaxed alloys, measurements in anisotropically strained alloys show splitting of normally degenerate band edges into two components. In a strained Si quantum well, this allows the engineered band offset to be followed from the GeSi substrate through the well to the alloy capping layer. In the high-mobility conduction channel, the band edge is found to be very sharp, in spite of obvious composition roughness. Near a misfit dislocation under the Si well, the band edge can shift by as much as 0.25 eV due to local strain. Within the core of the defect, however, strictly local behavior dominates the observations. Local conduction band splitting and in-gap states are both observed. PMID- 11108050 TI - Plasmon-loss imaging of chains of crystalline-silicon nanospheres and silicon nanowires AB - Nanostructures in chains of crystalline-silicon nanospheres and silicon nanowires were investigated using energy-filtered transmission electron microscopy (TEM). Observation of the shape of the silicon nanospheres in the chains provided the direct evidence that the chains were formed via the surface oxidation process, which may preferentially work at the necks. The diverse nanostructures in silicon nanowires were revealed, and we found smooth-shaped wires which have periodically modulated silicon cores. Nanostructures in wire-chain transition regions were also investigated for the first time. The wire-chain transition is not a simple junction of a silicon nanowire and a chain of silicon nanospheres, but has a periodically modulated silicon core in the wire region near the transition position. PMID- 11108051 TI - Direct observation of piezoelectric fields in GaN/ InGaN/GaN strained quantum wells AB - Off-axis electron holography is used to examine a single thin InGaN quantum well in GaN viewed in cross-section. The results show a phase offset across the well, which, under weakly diffracting conditions, is an approximately linear function of specimen thickness. This phase offset is ascribed to a change AV0 in the specimen mean inner potential V0 caused by a piezoelectric field induced by misfit strains in the InGaN layer. This paper examines the dependence of the phase offset on the diffracting conditions and on thin foil relaxation effects. It is shown that relaxation is negligible for the film thicknesses involved. Using a range of weakly diffracting conditions, the phase offset is measured as deltaV0/V0 = 0.042+/-0.012. Zone axis convergent beam electron diffraction patterns were taken and compared to simulations to determine the crystal polarity, showing the magnitude of the inner potential increased in the [0001] direction. By using dark-field displacement fringes to measure the InGaN layer thickness, and recent estimates of V0, the magnitude of the piezoelectric field is determined. This paper assesses the accuracy and limitations of electron holography for the studies of electric fields in other GaN structures. PMID- 11108052 TI - Quantitative TEM of point defects in Si AB - We review the use of transmission electron microscopy (TEM) to provide a quantitative measurement of both vacancy and interstitial clusters in ion implanted silicon. Interstitials agglomerate into rod-like defects on 131I) planes, and the evaporation of these defects can be directly correlated to the diffusion enhancements observed during annealing of ion-damaged silicon. Vacancy clusters are easily detected in TEM once they have been labelled using a Au diffusion technique. The combination of the two approaches provides a quantitative test for models of implantation and annealing in silicon. Detailed models for point defect behaviour, which include Ostwald-ripening and the surface recombination velocity, reproduce all of the crucial features of the observed defect annealing. PMID- 11108053 TI - Advances in SEM-based diffraction studies of defects and strains in semiconductors AB - Two scanning electron microscope (SEM)-based diffraction techniques, i.e. electron channelling contrast imaging (ECCI) and electron back-scatter diffraction (EBSD) are used to study lattice defect and local elastic strain distributions in Si1-xGe(x) epilayers grown on Si substrates patterned with mesas. The ECCI technique allows the misfit dislocations to be imaged in bulk samples. The misfit dislocations caused plastic relaxation of the strain in planar regions between mesas and in the wider mesas. In the narrower mesas the removal of lateral constraint at the mesa side faces had reduced the stress sufficiently to suppress the propagation of dislocations parallel to the closely spaced side faces. The measurements of small changes in the positions of two zone axes in EBSD patterns caused by variations in the local strain field were used to determine the strains and rotations making up the generalized plane strain tensor describing the deformation in the long mesa structures. The strain sensitivity of the method was determined to be approximately +/- 2 x 10(-4). Distributions of strains and rotations across mesas of several dimensions are reported and differ significantly between mesa for which the mesas width to epilayer thickness is high and low. PMID- 11108054 TI - Mechanism for secondary electron dopant contrast in the SEM AB - The growing use of secondary electron imaging in the scanning electron microscope (SEM) to map dopant distributions has stimulated an increasing interest in the mechanism that gives rise to so-called dopant contrast. In this paper a range of experimental results are used to demonstrate the wide applicability of the technique. These results are then incorporated into a model where, in particular, the effect of the surface barrier and the vacuum level are considered. It is found that the dominant contribution to the contrast mechanism is due to the three-dimensional variation of the vacuum level outside the semiconductor. PMID- 11108055 TI - Dislocation propagation in GaN films formed by epitaxial lateral overgrowth AB - We have investigated by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) the relationship between surface morphological evolution and dislocation propagation in GaN films formed by epitaxial lateral overgrowth (ELO) in hydride vapour phase epitaxy. The SEM observations revealed that step and terrace structures were formed on (0001) surfaces of the films both in the earlier and the later stages of growth, suggesting the occurrence of step flow growth during ELO. Bending dislocations with laterally propagated segments were frequently observed in the ELO films and their morphology led to a reduction in threading dislocation density in the film surface regions. Systematic TEM observations were performed to reveal the detailed structure of the bending dislocations. Comparison between the SEM and the TEM results showed that the lateral propagation of the dislocation was closely related to the appearance of the [1101) facets. A mechanism for dislocation propagation is discussed that explains the observed dislocation structure and surface step morphology. PMID- 11108056 TI - Gradual tilting of crystallographic orientation and configuration of dislocations in GaN selectively grown by vapour phase epitaxy methods AB - Cross-sectional transmission electron microscope (TEM) observation was performed for selectively grown gallium nitride (GaN) in order to examine the dependence of GaN microstructure on the growth conditions. The GaN films were grown by hydride vapour phase epitaxy (HVPE) or metalorganic vapour phase epitaxy (MOVPE) on GaN covered with a patterned mask. Thin foil specimens for TEM observation were prepared with focused ion beam (FIB) machining apparatus. It was demonstrated that the c-axis of GaN grown over the terrace of the mask tilts towards the centre of the terrace when the GaN is grown in a carrier gas of N2. The wider terrace results in a larger tilting angle if other growth conditions are identical. The tilting is attributed to 'horizontal dislocations' (HDs) generated during the overgrowth of GaN on the mask terrace. The HDs in HVPE-GaN have a semi loop shape and are tangled with one another, while those in MOVPE-GaN are straight and lined up to form low-angle grain boundaries. PMID- 11108057 TI - Derivation of growth mechanism of nano-defects in GaN from TEM data AB - Transmission electron microscopy (TEM) has been used to describe 'hollow' defects, e.g. nanotubes and pinholes (empty inside) formed in GaN hetero- and homoepitaxial layers. These defects are believed to be formed due to the presence of impurities and dopants. These defects appear to be even more deleterious than dislocations for the application of GaN in devices because their formation interrupts two-dimensional growth of quantum wells, and therefore will influence electrical and optical properties of the material. Experimental observations show that both these defects start from V-shaped indentations delineated by (1011) planes, and their density increases with higher concentrations of impurities or dopants. These 'hollow' defects are often found to be attached to dislocations, but the frequently observed termination of these defects inside the crystal eliminates dislocations as the source of these defects. Theoretical predictions also do not suggest that dislocations would have large-diameter empty cores like these of nanotubes and pinholes. PMID- 11108058 TI - Studies on the structure of crescent-shaped GaAs quantum wires by combination of electron microscopy and photoluminescence spectroscopy AB - Crescent-shaped GaAs quantum wires were fabricated on a V-grooved GaAs(001) substrate using a flow rate modulation epitaxy technique in metalorganic chemical vapor phase deposition method. The microstructures of the quantum wires were investigated using electron microscopy. The optimum growth conditions for the quantum wire superlattice structures were discussed from the observed microstructures. The optical properties of the fabricated single quantum wires were also investigated using photoluminescence spectroscopy. The relationship between the microstructures of the quantum wires and the observed optical properties is discussed on the basis of computer simulations of the electronics structure. PMID- 11108059 TI - Translocation of sLe(x) on the azurophilic granule membrane to the plasma membrane in activated human neutrophils. AB - Using immunogold electron microscopy, we found that human neutrophilic sialyl Lewis x (sLe(x)), an adhesive ligand for selectins, detectable by a monoclonal antibody, KM-93, is present in the sacculi of the Golgi apparatus as well as on the membranes of large electron-lucent azurophilic granules and the plasma membrane, including surface projections and microvilli. Neutrophilic sLe(x), however, was not detected on the membranes of specific granules. In comparison with the distribution of sLe(x), CD18 was localized on the plasma membrane and specific granule membrane but not on the azurophilic granule membrane. We also found by immunogold electron microscopy and flow cytometry that treatment of neutrophils with sialidase resulted in a loss of sLex on the plasma membrane. In contrast, intracellular sLex on the azurophilic granule membrane was not destroyed by sialidase. When sialidase-treated neutrophils were stimulated with N formyl-methionyl-leucyl-phenylalanine (fMLP), an inflammatory mediator peptide, in the presence of cytochalasin B, we observed by immunogold electron microscopy and flow cytometry that sLe(x) again appeared on the plasma membrane. These results indicate that stimulation by fMLP induces the up-regulation of sLe(x) on the cell surface by promoting translocation of sLe(x) from the azurophilic granule membrane to the plasma membrane in human neutrophils. PMID- 11108060 TI - Reproducibility and applicability of gallium replication as evaluated by biological specimen use. AB - Structures of biological surfaces pressed on to a pure liquid gallium surface were successfully traced on to the gallium surface by quick-freezing below the melting point (28.78 degrees C) in air or water for replication in scanning electron microscopy. Gallium's high surface tension (approximately 700 mN m(-1) at 30 degrees C) deteriorates the spatial resolution of replicas and destroys some types of specimens. Five different biological surfaces were replicated on to gallium surfaces to evaluate spatial resolution and specimen resistance, i.e. reproducibility and applicability. Gallium replication of jewel beetle wing and human hair demonstrated submicron spatial resolution in the horizontal direction at least. Trials of protozoa, bacteria, and culture cell replication showed that protozoa are suited to replication because the cell membrane has characteristic structures with sufficient resistance to the gallium surface. PMID- 11108061 TI - The effect of calcium ionophore A23187 on neurites from embryonic mouse spinal cord explants in culture. AB - The calcium channel ionophore A23187 has been shown to cause a striking reduction in both the rate of neurite outgrowth and in growth cone motility; it has also been shown that these changes may be reversed when the ionophore is removed from the culture medium. The evidence supports the view that a specific concentration range of calcium is essential for outgrowth and motility and that calcium influences neuronal growth by an effect on actin structures within the growth cone. Using time-lapse video microscopy techniques, the effects of calcium ionophore A23187 on the rate of neurite outgrowth and growth cone motility in cultured embryonic mouse spinal cord neurons were examined. Concentrations in the range of 1-100 microM resulted in an inhibition of neuronal outgrowth, a loss of filopodia and a significant reduction in survival after seven hours. The results show that although the treatment with A23187 inhibited growth at all concentrations used, motility was inhibited only at the highest concentration indicating that the optimum calcium concentration for motility is higher than that for growth and that calcium mediates its effects on growth and motility at a very local level. PMID- 11108062 TI - Enzyme-histochemical demonstration of lymphatic vessels in the golden hamster periodontium: an attempt to reactivate the 5'-nucleotidase activity with Mg++ ion supply. AB - Enzyme-histochemical demonstration of lymphatic vessels in the golden hamster periodontium was performed on cryostat sections using the 5'-nucleotidase (5' Nase) staining method by light microscopy and backscattered electron imaging of scanning electron microscopy. The inhibition of the 5'-Nase activity by decalcification was cancelled by the Mg++ ion supply. The reaction products of 5' Nase activity were produced on the lymphatic endothelial cells and the tubular structures of lymphatic vessels were seen as highlights by backscattered electron imaging. The invasion of 5'-Nase-positive lymphatic vessels into the alveolar bone from the periodontium was found in the present study. PMID- 11108063 TI - Sleep and the smoker. PMID- 11108064 TI - Therapies for chronic hepatitis B: emerging roles for nucleoside analogues. PMID- 11108065 TI - Management of Graves' disease in Australia. AB - BACKGROUND: Surveys of physicians in Europe, the USA and elsewhere have shown marked international differences in the management of Graves' disease. There are no comparable data on clinical practice in Australia. AIMS: To examine the current management of Graves' disease by Australian endocrinologists, particularly controversial aspects of management. METHODS: A questionnaire, modified from previous studies, was sent to members of the Endocrine Society of Australia, asking how they would manage a 43-year-old female with a first episode of Graves' disease. Eight variations on this index case (goitre size, age, sex, severity, recurrent disease) were then introduced. A novel ninth variation, recurrent Graves' disease accompanied by moderate ophthalmopathy, was added. RESULTS: Responses from 130 endocrinologists who regularly managed Graves' disease in adults were analysed. For the index case, medical treatment with antithyroid drugs was recommended by 81% of respondents and radioiodine by 19%. Most respondents also recommended medical treatment for a patient aged 19 years, a patient with a large goitre, no goitre or severe hyperthyroidism. For an older patient aged 71 years, however, 57% of endocrinologists recommended radioiodine, and the remainder medical treatment. For recurrent Graves' disease after previous medical treatment, 76% of respondents recommended radioiodine, 22% medical treatment and 2% surgery. By contrast, for an identical case accompanied by moderate ophthalmopathy, 54% recommended medical treatment, 27% surgery and only 19% radioiodine. CONCLUSIONS: Most endocrinologists in Australia recommend medical treatment for a first episode of Graves' disease. Radioiodine is used mainly in older patients and for recurrent disease. In the presence of significant ophthalmopathy, most endocrinologists avoid the use of radioiodine. PMID- 11108066 TI - The optimal use of allopurinol: an audit of allopurinol use in South Auckland. AB - BACKGROUND: Gout is a common and challenging problem in South Auckland, New Zealand. Allopurinol is widely used but urate reduction remains unsatisfactory. Allopurinol dosing guidelines and a therapeutic range for plasma oxypurinol levels have been published. AIMS: We aimed to determine the appropriateness of allopurinol dosing according to current guidelines and to assess the relationship between plasma creatinine, oxypurinol and urate. In addition, we assessed the clinical usefulness of the oxypurinol level. METHODS: Thirty-one patients, on a stable dose of allopurinol for at least three weeks, had plasma creatinine, urate and oxypurinol measured as part of routine clinical assessment. Relationships between the various methods were examined using regression analysis. Fisher's exact test was used to test associations with categorical variables. RESULTS: Fifty-five per cent of patients were on higher than recommended doses of allopurinol. There was a statistically significant relationship between calculated creatinine clearance and plasma oxypurinol level. Only 50% of patients with a plasma oxypurinol within the therapeutic range (30-100 micromol/L) had a plasma urate < 0.42 mmol/L and this did not increase significantly in the patients with an oxypurinol level > 100 micromol/L. CONCLUSIONS: There is poor adherence to the current recommended dosing guidelines for allopurinol. Creatinine clearance rather than plasma creatinine needs to be used to predict the dose of allopurinol. The current role of the oxypurinol level is to identify non-compliers with allopurinol therapy. We need further research to clarify whether increasing the dose of allopurinol outside the recommended dose range to reach an oxypurinol level of close to 100 micromol/L may be of benefit in those who have not had sufficient urate reduction. PMID- 11108067 TI - The diagonal ear lobe crease (Frank's sign) is not associated with coronary artery disease or retinopathy in type 2 diabetes: the Fremantle Diabetes Study. AB - BACKGROUND: The diagonal ear lobe crease (ELC) has been suggested as a simple marker of vascular disease in the general population but there are few data from diabetic patients despite their increased risk of angiopathy. AIM: To determine whether the ELC is a clinically useful sign of coronary artery disease (CAD) or retinopathy in type 2 diabetes. METHODS: One thousand and twenty-two patients from the multi-ethnic urban catchment area of Fremantle Hospital in Western Australia were studied. This sample represents 79% of the type 2 diabetic subjects recruited to the ongoing Fremantle Diabetes Study and 49% of all 2072 patients with type 2 diabetes identified through active case detection in a postcode-defined region of 120,097 people. In addition to other comprehensive data relating to diabetes and its management, the presence of an ELC and evidence of both CAD and retinopathy were ascertained for each patient. RESULTS: The prevalence of ELC was 55%. Patients with an ELC were more likely to have CAD than those without an ELC (p=0.019), but the proportions with retinopathy were not significantly different (p=0.085). The sensitivity and specificity of ELC for detecting CAD were 60% and 48%, and for retinopathy 61% and 47% respectively. The patients with an ELC were significantly older, more likely to be male and had a higher systolic blood pressure than those without (p<0.02). After adjusting for known vascular risk factors, socioeconomic variables and ethnicity in a logistic regression model, an ELC was neither a significant independent predictor of CAD (p=0.45) nor of retinopathy (p=0.14). CONCLUSIONS: The ELC is of little value as a sign of the presence of diabetic vascular complications. PMID- 11108068 TI - Non-urease producing Helicobacter pylori in chronic gastritis. AB - BACKGROUND: Helicobacter pylori infection is the commonest cause of gastritis. Different patterns of immune response to H. pylori infection and characteristics of bacteria are considered to contribute to clinical outcomes. AIM: To determine characteristics of the host H. pylori relationship in subjects with non-ulcer dyspepsia and a histological diagnosis of gastritis. METHODS: Thirty-five subjects with chronic gastritis undergoing endoscopy (mean age 53 years, range 24 82, 14 male and 21 female) were studied, none of whom was on nonsteroidal anti inflammatory drugs or antibiotics. H. pylori infection was determined by rapid urease test (CLOtest), culture, antibody and RT-PCR for Ure C, Cag A and 26 kDa gene and histology. Cytokine production of mucosal IL-6 and IL-8 were measured by ELISA. RESULTS: Fifteen subjects were positive by CLOtest and/or bacterial culture. In these subjects histology showed numerous helical forms of H. pylori (Group I). Nine subjects were negative by CLOtest, bacterial culture, and mRNA for urease C fragment, but positive by PCR for the 26 kDa protein encoding gene. Histology in these subjects showed the presence of either coccoid forms (four), or scant helical forms (two), or mixed coccoid/helical forms (three) (Group II). Eleven subjects were negative by all methods of detection (Group III). IgG and IgA antibody levels in serum (p<0.05) and gastric tissue culture supernatant (p<0.001) were significantly higher in Group I than those in Group II or III. There were significant differences in the IgG serum and IgA supernatant antibody levels (p<0.01 and p<0.05) when Group II was compared to Group III. Supernatant IL-6 levels were significantly higher in Group I (p<0.01) than those from Groups II and III. IL-8 levels were higher in Group I (p<0.01) and Group II (p<0.05) when compared to Group III. CONCLUSIONS: 'H. pylori-negative' gastritis can be associated with a non-urease producing form of H. pylori, with a reduction in both local and systemic antibody levels and mucosal pro-inflammatory cytokines. PMID- 11108069 TI - A telephone call reminder to improve outpatient attendance in patients referred from the emergency department: a randomised controlled trial. AB - BACKGROUND: Poor compliance with attendance at outpatient clinic appointments in patients referred from emergency departments (EDs) is a major problem in public hospitals. AIMS: To determine whether the intervention of a telephone call within three days of ED attendance would improve: 1. the proportion of patients making recommended outpatient appointments; and 2. the proportion of patients attending scheduled appointments. To characterise reasons for non-attendance at appointments made by patients referred from the ED. METHODS: A randomised controlled trial was undertaken of 400 patients recommended to make outpatient appointments during attendance at The Royal Melbourne Hospital ED in July-August 1999. INTERVENTION: a telephone call one to three days after attendance to remind the patient about the appointment (and its importance for medical follow-up) if one had been made and to offer to make an appointment if one had not been made. OUTCOME MEASURES: 1. making the recommended appointment; 2. attendance at the scheduled appointment; and 3. reasons for non-attendance at scheduled appointments. RESULTS: The telephone intervention improved attendance at scheduled appointments from 54.4% to 70.7% (p=0.002). The proportion of patients making appointments was not significantly affected. The commonest reasons given for non-attendance were: attended general practitioner (13%), attended private specialist (6.6%), inpatient in hospital at time of appointment (6.6%), too busy or inconvenient (5.3%), claimed to have attended (5.3%) and did not differ by intervention. CONCLUSIONS: A significant improvement in the proportion of patients attending outpatients appointments can be made by a simple reminder telephone call one to three days after attendance at the ED. PMID- 11108070 TI - Advances in the management of prostate cancer. PMID- 11108071 TI - Atherosclerosis and the vulnerable plaque--pathogenesis: Part I. PMID- 11108072 TI - Diabetes mellitus and coronary heart disease--from prevention to intervention: Part II. PMID- 11108073 TI - The potential impact of global environmental change on population health. AB - Due to rapid industrial changes and increased pressure of people on fragile ecosystems, large-scale environmental perturbations have been occurring on Earth. Major current environmental problems that can be expected to have a substantial effect on human health include human-induced climate change and stratosphere ozone depletion, because they threaten the ecological support systems on which human life depends. The most serious potential consequence of global environmental change is the erosion of Earth's life-support systems. The public health assessments of the present and future anthropogenic damage to the biosphere have important implications for human health and wellbeing. Medical practitioners have an important role to play in this field. PMID- 11108074 TI - Richard Rawdon Stawell--the complete physician. PMID- 11108075 TI - From the other side--a patient's perspective. PMID- 11108076 TI - Kikuchi's disease associated with parotidomegaly, thyroiditis and a rash in a young man. PMID- 11108077 TI - Clinical ergotism with severe bilateral upper limb ischaemia precipitated by an erythromycin--ergotamine drug interaction. PMID- 11108078 TI - Ataxia and toadfish poisoning. PMID- 11108079 TI - Implications of a rapid decrease in serum Troponin T levels after renal transplantation. PMID- 11108080 TI - Gouty arthritis in Australian Aborigines. PMID- 11108081 TI - Diltiazem-mediated inhibition of sildenafil metabolism may promote nitrate induced hypotension. PMID- 11108082 TI - The development of close relationships in Japan and the United States: paths of symbiotic harmony and generative tension. AB - Findings from research on parent-child and adult mate relationships suggest that there are different paths of development in Japan and the United States. In Japan, the path is one of symbiotic harmony, as seen in the emphasis on union in infancy, others' expectations in childhood, the stability of relationships with parents and peers in adolescence, and assurance about the mate relationship in adulthood. In the United States, the path is one of generative tension, as seen in the tug between separation and reunion in infancy, the emphasis on personal preferences in childhood, the transfer of closeness from parents to peers in adolescence, and the emphasis on trust-a faith and hope in new relationships-in adulthood. The notion that there are different paths of development challenges Western investigators' presumption that certain processes-separation individuation, use of the relational partner as a secure base for exploration, and conflict between partners-are central in all relationships. The notion of different paths also challenges the assumption of many cross-cultural investigators that relationships in the United States are less valued or weaker than those in Japan; this article highlights cultural differences in the meaning and dynamics, as opposed to the importance and strength, of relationships. The model suggests a need to investigate the processes underlying, and the adaptive consequences of, these two alternative paths. PMID- 11108083 TI - Collective construction of the self and social relationships: a rejoinder and some extensions. AB - This commentary elaborates on the basic thesis developed by Rothbaum, Pott, Azuma, Miyake, and Weisz and underscores the significance of the co-constructive process of the self and social relationship. Implications for future cultural psychological inquiry in this area are discussed. PMID- 11108084 TI - New insight and old dilemma: a cross-cultural comparison of Japan and the United States. AB - Conflict in close relationships, or "generative tension," characterizes both the United States and Japan, with differences only in the style and timing of its manifestation. The potentially fruitful strategy of Rothbaum et al.'s article is constrained by their cross-cultural comparative methodology. PMID- 11108085 TI - Developmental pathways in close relationships. AB - Multiple case study developmental pathway research is needed to substantiate the theoretical propositions of the target article. PMID- 11108086 TI - Human development in the United States and Japan: new ways to think about continuity across the lifespan. AB - The work of Rothbaum and colleagues integrates more than 200 studies conducted in two countries over four stages of development. Their method of integrating studies provides a promising way to overcome some of the most vexing methodological difficulties of cross-cultural research. PMID- 11108087 TI - Using "the Japanese problem" as a corrective to the ethnocentricity of Western theory. AB - This reflection on the essay by Rothbaum, Pott, Azuma, Miyake, and Weisz focuses on how knowledge about Japanese psychological development and culture can serve as a corrective to the ethnocentrism of Western theory. Particular attention is given to the Japanese cultural concepts of amae and kejime. PMID- 11108088 TI - Trade-offs in the study of culture and development: theories, methods, and values. AB - The commentators are unanimous in their support for our general orientation to culture and development, and for the pathways we have identified, and they suggest ways to enrich our approach to theory, methods, and values. We view their main suggestions as relating to trade-offs: between theories that highlight generalizations or exceptions; between methods that rely on one-, two-, or multiculture studies; and between values involving individuation or accommodation. Here, we describe ways to find an optimal balance in each instance. PMID- 11108089 TI - The cues that infants use to distinguish discontinuous quantities: evidence using a shift-rate recovery paradigm. AB - A novel experimental method, shift-rate recovery, was developed and used in a series of three experiments. These examined the extent to which 6-month-old infants (N = 131) find perceptual cues such as density and length useful in the discrimination of linearly arranged sets containing large numbers of objects. Results showed that infants discriminated between arrays that differ in number and density, with length held constant, when the arrays were presented either simultaneously or successively. On the other hand, infants discriminated only between arrays that differ in number and length, with density held constant, when the arrays were presented simultaneously. Infants were, however, able to perform a successive length discrimination when the arrays were continuous rather than consisting of discrete items. These findings support the conclusion that infants are able to discriminate between large number sets by relying on absolute cues such as density (but not length) and on relative cues such as optical one-to-one correspondence. PMID- 11108090 TI - The development of relational landmark use in six- to twelve-month-old infants in a spatial orientation task. AB - The ability to use the relations between visible landmarks to locate nonvisible goals (allocentric spatial coding) underlies success on a variety of everyday spatial orientation problems. Little is known about the development of true relational coding in infancy. Ninety-six 6-, 8.5- and 12-month-old infants were observed in a peekaboo paradigm in which they had to turn to a target location after displacement to a novel position and direction of facing. In a landmark condition, the target position was located between two landmarks, contrasted with a control condition in which no distinctive landmarks were provided. Six-month old infants performed poorly in both conditions, 8.5-month-olds were significantly better with the landmarks, and 12-month-olds solved the task with or without landmarks. A follow-up study confirmed that the 8.5-month-olds used both landmarks to solve the task. This demonstration of allocentric spatial coding in 8.5-month-old infants shows earlier competence than that found in previous work in which only infants at the end of the first year were able to use landmarks relationally. PMID- 11108091 TI - The development of visual expectations in the first year. AB - The development of expectations was investigated by using the Visual Expectations Procedure. In Experiment 1, 128 infants aged 6-, 9-, and 12-months-old saw two 40 trial sequences of a videotaped mechanical toy appearing in various locations. The sequences represented an alternation pattern (i.e., ABAB) or a complex pivot pattern (i.e., ABCBABCB). In Experiment 2, 76 infants aged 4-, 8-, and 12-months old saw either a left-right alternation or a top-bottom alternation. Reaction time improved and the percentage of anticipations increased between 6 and 9 months in Experiment 1 and between 4 and 8 months in Experiment 2 but not thereafter. Anticipations for the pivot sequence and for younger infants on both sequences were often incorrect (i.e., gaze shifts occurred before stimulus onset but were not directed toward the upcoming stimulus). We conclude that young infants have expectations that reflect some degree of general or procedural knowledge, but it is not clear that this behavior implies specific, articulated expectations about upcoming events. PMID- 11108092 TI - Detecting blickets: how young children use information about novel causal powers in categorization and induction. AB - Three studies explored whether and when children could categorize objects on the basis of a novel underlying causal power. To test this we constructed a "blicket detector," a machine that lit up and played music when certain objects were placed on it. First, 2-, 3- and 4-year-old children saw that an object labeled as a "blicket" would set off the machine. In a categorization task, other objects were demonstrated on the machine. Some set it off and some did not. Children were asked to say which objects were "blickets." In an induction task, other objects were or were not labeled as "blickets." Children had to predict which objects would have the causal power to set off the machine. The causal power could conflict with perceptual properties of the object, such as color and shape. In an association task the object was associated with the machine's lighting up but did not cause it to light up. Even the youngest children sometimes used the causal power to determine the object's name rather than using its perceptual properties and sometimes used the object's name rather than its perceptual properties to predict the object's causal powers. Children rarely categorized the object on the basis of the associated event. Young children also sometimes made interesting memory errors-they incorrectly reported that objects with the same perceptual features had had the same causal power. These studies demonstrate that even very young children will easily and swiftly learn about a new causal power of an object and spontaneously use that information in classifying and naming the object. PMID- 11108093 TI - Implicit learning in children is not related to age: evidence from drawing behavior. AB - Three experiments are reported on implicit learning in 432 children between the ages of 4 and 10 years, using a new paradigm ("the neutral parameter procedure") based on drawing behavior. The first two experiments demonstrated that children modified their drawing behavior following specially devised practice in such a way that these modifications could not be viewed as the result of deliberate adaptive strategies. The third experiment showed that these behavioral modifications lasted for at least 1 hr after the training phase. No age-related differences appeared in the experiments. A comparison of children's data with similar adults' data also failed to reveal any age differences. These results provide compelling evidence that implicit learning processes are age independent. Some implications of these results for developmental issues are discussed, notably the hypothesis of the formation of implicit knowledge in the course of learning. PMID- 11108094 TI - Infants' perception of the audible, visible, and bimodal attributes of multimodal syllables. AB - Three experiments investigated 4-, 6-, and 8-month-old infants' perception of the audible, visible, and combined attributes of bimodally specified syllables. Ninety-six infants in each experiment were habituated to a person mouthing and uttering a syllable and then tested for detection of changes of either the audible, visible, or combined attributes of the syllable. When the attributes of the syllable were produced in an adult-directed manner, all three age groups discriminated the audible and bimodal attribute changes but only the 8-month-olds discriminated the visible one. When the difference between the familiar and novel attributes of the syllable was enhanced by testing with a novel syllable produced in an infant-directed manner, all three age groups detected all three types of changes. Finally, to test the possible role of audiovisual synchrony in responsiveness, infants were tested with an asynchronous syllable spoken either by the same person or by a novel person following habituation to a synchronous syllable. Results suggested that at four months infants attended primarily to the featural information, at six months primarily to the asynchrony, and at eight months to both features independently. These results help identify some of the important dimensions of multimodal speech during early development. PMID- 11108095 TI - Prosodic correlates of stress in babbling: an acoustical study. AB - Prelinguistic babbling often seems remarkably speech-like, not because it has recognizable words but because it seems to have adult-like prosody. To quantify this impression, we compared disyllabic sequences from five infants and five adults in terms of the use of frequency, intensity, and duration to mark stress. Significantly larger values for the three acoustic variables were observed on stressed than on unstressed syllables independent of syllable position for both groups. Adults showed the correlates of utterance final syllables--lower f0, lower intensity, and longer duration; infants showed only decrease in intensity. Ratios for stressed to unstressed syllables and participation of the three variables in stress production in individual disyllables were highly similar in both groups. No bias toward the English lexical trochaic stress pattern was observed. We conclude that infants in English environments produce adult-like stress patterns before they produce lexical items, which specify stress. Acoustic and perceptual analyses are used to explore stress marking by prelinguistic infants in an English language environment. Results show that infants employ the three acoustic correlates of stress in individual syllables in a manner largely similar to that of adult speakers, although they do not show second-syllable declination effects or an English language trochaic stress bias. PMID- 11108096 TI - Two-year-olds will name artifacts by their functions. AB - Do young children take functional information into account in naming artifacts? In three studies of lexical categorization, 112 children 2 years of age learned new names for novel artifacts with novel functions and then extended the names to new objects. The objects were designed to have functions that were causally related in simple and compelling ways to perceptible aspects of their physical structure. Despite only minimal opportunity to familiarize themselves with the objects, children generalized the names in accordance with the objects' functions. This result obtained even when children had to discover the functions of the named objects on their own (Experiment 2) and when all the test objects had some discernible function (Experiment 3). Two-year-olds name by function when they can make sense of the relation between the appearances and the functions of artifacts. PMID- 11108097 TI - Taking decisions seriously: young children's understanding of conventional truth. AB - Research suggests that young children may see a direct and one-way connection between facts about the world and epistemic mental states (e.g., belief). Conventions represent instances of active constructions of the mind that change facts about the world. As such, a mature understanding of convention would seem to present a strong challenge to children's simplified notions of epistemic relations. Three experiments assessed young children's abilities to track behavioral, representational, and truth aspects of conventions. In Experiment 1, 3- and 4-year-old children (N = 30) recognized that conventional stipulations would change people's behaviors. However, participants generally failed to understand how stipulations might affect representations. In Experiment 2, 3-, 5 , and 7-year-old children (N = 53) were asked to reason about the truth values of statements about pretenses and conventions. The two younger groups of children often confused the two types of states, whereas older children consistently judged that conventions, but not pretenses, changed reality. In Experiment 3, the same 3- and 5-year-olds (N = 42) participated in tasks assessing their understanding of representational diversity (e.g., false belief). In general, children's performance on false-belief and "false-convention" tasks did not differ, which suggests that conventions were understood as involving truth claims (as akin to beliefs about physical reality). Children's difficulties with the idea of conventional truth seems consistent with current accounts of developing theories of mind. PMID- 11108098 TI - Remembering and understanding: the effects of changes in underlying knowledge on children's recollections. AB - This study was designed to explore the influence of changes in children's knowledge on earlier constructed memories. Kindergartners' (N = 102) recall of a series of stories was examined as a function of their interpersonal knowledge about the main story character. Children's knowledge about the protagonist was manipulated prior to presentation of the stories, and the effects of their impressions on story recall were examined. A change in some of the children's impressions was then promoted, and the impact of this second knowledge manipulation on recall of previously heard stories was assessed. The results indicated that children's story recall was affected by their prior impressions. Moreover, following the second knowledge manipulation, children revised their story reports in ways that were consistent with their newly acquired impressions, which suggests that they had reconstructed their memories of previously heard stories. These findings provide evidence for both prospective and retrospective effects of knowledge on memory. PMID- 11108099 TI - Same beginnings, different stories: a comparison of American and Chinese children's narratives. AB - This study examined social, emotional, and cognitive characteristics of American and Chinese children's narratives. Twenty-four American and 26 Chinese 6-year-old children participated. Each child was interviewed individually twice with a 1 week delay interval. During the two interviews, children were asked to tell 11 stories prompted by pictures and standard verbal leads and to recount 7 emotional memories. Content analyses were performed on children's stories and memories. In line with predictions, findings indicated that compared with American children, Chinese children showed greater orientation toward social engagement, greater concern with moral correctness, greater concern with authority, a less autonomous orientation, more expressions of emotions, and more situational details in both their stories and memories. A few gender differences were found. Findings are discussed in terms of different value systems and early socialization practices in these two cultures. PMID- 11108100 TI - Motivated scientific reasoning biases, epistemological beliefs, and theory polarization: a two-process approach to adolescent cognition. AB - Theory-motivated reasoning biases arise when different reasoning skills are invoked to evaluate evidence that is congruent or incongruent with individuals' belief systems. To explore this phenomenon, 66 early and 73 middle adolescents evaluated evidence relevant to their theories of social class or religion. In both conditions, reasoning biases were found, but in-group biases were evident only in the religion condition. In both conditions, higher order scientific reasoning was used to reject theory-incongruent evidence and judgmental heuristics (i.e., cognitive rules of thumb) were used to evaluate theory congruent evidence. In both conditions, subsequent to the evidence presentation, adolescents' theories became more extreme (i.e., polarized) than at the outset of the experiment. Beliefs regarding the origin, acquisition, and certainty of knowledge, however, appeared to moderate reasoning biases and theory polarization. Age differences emerged on only one index of bias: In the religion condition, middle adolescents were more likely to treat theory-incongruent evidence as implausible. These findings are pertinent to theories of cognitive development, decision making, rationality, and in-group favoritism. PMID- 11108101 TI - Prediction of elementary school children's externalizing problem behaviors from attentional and behavioral regulation and negative emotionality. AB - The purpose of this study was to examine the moderating role of individual differences in negative emotionality in the relations of behavioral and attentional (emotional) regulation to externalizing problem behaviors. Teachers' and one parent's reports of children's regulation (attentional and behavioral), emotionality, and problem behavior were obtained when children were in kindergarten to grade 3 and two years later (N = 169; 146 in major analyses); children's behavioral regulation also was assessed with a measure of persistence. According to the best fitting structural equation model, at two ages behavioral dysregulation predicted externalizing behavior problems for children both high and low in negative emotionality, whereas prediction of problem behavior from attentional control was significant only for children prone to negative emotionality. There were unique, additive effects of behavioral and attentional regulation for predicting problem behavior as well as moderating effects of negative emotionality for attentional regulation. PMID- 11108102 TI - Helping mothers discuss sexuality and AIDS with adolescents. AB - The current study was designed to alter experimentally mothers' style when discussing sexuality and AIDS with their adolescent children. Mothers of 11- to 15-year-olds (N = 50) were assigned to an intervention or control group, resulting in 20 dyads in each group. All dyads were assessed twice, on self reported and observed communication, AIDS knowledge, and perceived vulnerability to AIDS. Intervention group mothers received two training sessions. Observational data revealed that intervention group mothers reduced their amount of speaking, asked more open-ended questions, acted less judgmental, and discussed dating and sexuality more than did control group mothers. Intervention group adolescents reported increased discussions of birth control and increased daily comfort talking to their mothers. There was some evidence that intervention group girls increased in AIDS knowledge. There was no change in AIDS-related beliefs. PMID- 11108103 TI - The role of peers in the emergence of heterosexual romantic relationships in adolescence. AB - Adolescents' peer structures and the quality of their friendships were explored as antecedents of romantic relationships. Longitudinal data were gathered in a sample of 180 high school students over a 3-year period from grade 9 to grade 11. Consistent with Dunphy (1963), small groups of close friends were predictive of other-sex peer networks which were, in turn predictive of the emergence of future romantic relationships. Indirect effects were found for same-sex groups of close friends and same-sex networks. Consistent with Furman and Wehner (1994), the qualitative features of relationships with both friends and romantic partners were predictive of the qualitative features of subsequent romantic experiences. These linkages suggest ways in which peer relationships may support romantic development at this stage of the life cycle. PMID- 11108104 TI - Single mothers in low-wage jobs: financial strain, parenting, and preschoolers' outcomes. AB - Using data from an ongoing study of 93 single Black mothers of preschoolers who had been welfare recipients, but were employed in low-wage jobs at baseline, this study tests a model of how maternal education, economic conditions (earnings and financial strain), and the availability of instrumental support influence maternal psychological functioning, parenting, and child development. Results indicate that maternal educational attainment was positively associated with earnings, which, together with instrumental support, were negatively associated with financial strain. Financial strain, in turn, was implicated in elevated levels of depressive symptoms, which were directly and negatively implicated in parenting quality. The quality of parenting was associated with children's behavior problems and preschool ability. Specifically, mothers with higher scores on the HOME scale, our measure of involved, supportive parenting, had children with fewer behavior problems and better preschool ability. PMID- 11108105 TI - Mother-child discourse, attachment security, shared positive affect, and early conscience development. AB - The separate literatures on parental discipline, maternal discourse about emotion, and autobiographical memory support the idea that parent-child discourse in the context of a supportive relationship plays a role in a child's early conscience development, and this study was designed to examine this issue. Forty two preschool children and their mothers took part in a 45-min structured laboratory session, and at their homes, mothers completed the Attachment Q-Set. As part of the laboratory session, each mother was asked to discuss with her child one incident that occurred within the last week in which her child behaved well and one in which her child misbehaved. These conversations were transcribed verbatim and coded for maternal references to feelings, rules, consequences of the child's actions, and moral evaluatives. Each child also took part in a behavioral measure of internalization and several compliance tasks, and mothers completed a maternal report of the child's early conscience development. Consistent with attachment theory, attachment security predicted maternal and child references to feelings and moral evaluatives. Attachment security, shared positive affect between the mother and child, and maternal references to feelings and moral evaluatives also predicted specific aspects of early conscience development. PMID- 11108106 TI - Appraisal, social support, and life events: predicting outcome behavior in school age children. AB - To examine the relation between social support and appraisal of life events in predicting adaptive, externalizing, and internalizing behavior in 265 school-age children, child-report on both a global and a significant other measure of social support was used. Life event scores were separated into events endorsed as negative and events endorsed as positive by the child. Using hierarchical regression analyses, the present study tested two models: main effects and moderator models of the relation between life events, social support, and behavioral outcome. Support was found for global social support and positive life events in predicting adaptive, externalizing, and internalizing behavior. Gender differences were also found. Support was found for both the main effects and moderator models of the association between life events and global social support. Appraisal of life events as positive appears to compensate for lower levels of global social support. Appraisal is discussed as a possible protective factor from maladjustment after exposure to major life events. PMID- 11108107 TI - Comparisons of adopted and nonadopted adolescents in a large, nationally representative sample. AB - There are conflicting findings about whether adopted children have more psychological and behavioral problems than nonadoptees. Research results are discrepant partly because many previous studies were based on small clinical samples or on samples biased by self-selection. A nationally representative school survey (Add Health) was used to compare adopted (n = 1,587) and nonadopted adolescents (total N = 87,165) across a wide variety of measures. Standardized mean differences show that adopted adolescents are at higher risk in all of the domains examined, including school achievement and problems, substance use, psychological well-being, physical health, fighting, and lying to parents. Demographic and background variable breakdowns show that the effect sizes for differences between adopted and nonadopted adolescents were larger for males, younger or older adolescents, Hispanics or Asians, and adolescents living in group homes or with parents of low education. Distributional analyses revealed approximately a 1:1 ratio of adopted to nonadopted adolescents in the middle ranges of the outcome variables but a ratio of 3:1 or greater near the tails of the distributions. These data clearly show that more adopted adolescents have problems of various kinds than their nonadopted peers; effect sizes were small to moderate based on mean differences, but comparisons of distributions suggest much larger proportions of adopted than nonadopted adolescents at the extremes of salient outcome variables. PMID- 11108108 TI - Anomalies and variants of the endoscopic anatomy for third ventriculostomy. AB - OBJECTIVE: Endoscopic third ventriculostomy (ETV) is an alternative to shunt placement in occlusive hydrocephalus. The negative impact of anatomic anomalies and variants on ETV have been sporadically reported but not yet investigated systematically. Therefore, the objectives of the present study are 1) to evaluate the frequency of endoscopic anatomic anomalies of the ventricular system, 2) to define their potential to complicate the procedure and to compromise the surgical results, and 3) to investigate the value of preoperative magnetic resonance (MR) imaging for their detection. METHOD: The video recordings, the operative reports, and the preoperative MR images of 25 hydrocephalic patients who underwent ETV were reviewed. The surgical results were classified into completed and successful, completed, but failed, and unsuccessfully attempted ETV and were correlated with the absence or presence of anatomic variants. RESULTS: In 9 of the 25 patients, 10 anatomic anomalies or variants, respectively, were identified, accounting for an incidence rate of 36%. The single most common anatomic anomaly was a thickened third ventricular floor in 4 patients. Anatomic variants extended the operation time (n = 6), increased the stretching of floor and walls of the third ventricle during perforation (n = 4), were related to minor arterial bleeding (n = 3), and obscured the visual control of the basilar artery (n = 2). In 5 of the 9 patients, ETV was completed and successful, but in 2 patients, ETV was finally abandoned, and in an additional 2 patients, ETV was completed, but failed to cure the symptoms of hydrocephalus. In contrast, ETV was completed and successful in all 16 patients with normal anatomy. All anatomic anomalies had been detectable on preoperative MR imaging, with the exception of the thickened floor of the third ventricle. CONCLUSION: Anatomic anomalies are a frequent finding during ETV. Successful perforation and control of the hydrocephalus correlates with the absence of anatomic anomalies. Most anatomic variants have the potential to increase the operative risk. With the exception of the thickened third ventricular floor, MR imaging allows us to identify all anatomic anomalies preoperatively, and enables the neurosurgeon to weigh the operative risk in a patient with an anatomic anomaly against the chance to perform ETV successfully. PMID- 11108109 TI - Endoscopic treatment of colloid cysts of the third ventricle: 9 consecutive cases. AB - The purpose of this study is to evaluate the efficacy of the endoscopic technique for the treatment of the colloid cysts of the third ventricle. Between August 1995 and October 1997 a series of nine patients with colloid cyst of the third ventricle (6 males and 3 females) were treated with this method. The technique, consisting of cyst fenestration, aspiration of the colloid, and coagulation of the internal layer of the wall, was always effective in restoring CSF circulation. Operating time was 54-120 min (median 67 min). We recorded only one post-operative septic complication but no signs of direct surgical morbidity. Post-operation hospital stay was 2-30 days (median 5 days). Follow up was 14-40 months (mean 27 months). We did not observe any clinical or radiological recurrence. Endoscopic treatment of colloid cysts of the third ventricle is a safe and effective alternative to the well-established approaches of microsurgical removal and stereotactic aspiration. Only a very long follow-up will answer the question of the long-term effectiveness of this method. PMID- 11108110 TI - Cranial neuronavigation in neurosurgery: assessment of usefulness in relation to type and site of pathology in 284 patients. AB - OBJECTIVE: Neuronavigation improves intraoperative topographical orientation in neurosurgery. We wanted to better define the practical value of this technique in relation to the pathology operated on and the types of cranial surgery that profit the most from it. METHODS: Usefulness, interactive use and probably preventive effect of neuronavigation in cranial neurosurgery were assessed in a consecutive series of 284 patients on the basis of questionnaires with two- or five-point scale ratings by different neurosurgeons. RESULTS: Neuronavigation was most helpful in tumors of the hemispheres (particularly the central area) not visible at the cortical surface or resembling normal white matter, and in endoscopic procedures within small ventricles or cysts with non-translucent walls or when vision was blurred by cloudy CSF. In the same pathologies and surgical procedures, the device was interactively used, taking advantage of the specific possibilities of interactive image-guided neurosurgery. A probably preventive effect of neuronavigation was noted in operations in eloquent areas; highest scores were given for intraaxial tumors of the central region. The subjective assessments of usefulness, interactive use or preventive effect were not dependent on the involvement of the neurosurgeons in this study. CONCLUSION: We recommend this technique in resecting tumors in eloquent areas of the cortex or white matter, in approaching deep-seated processes not visible at the cortical surface, in defining borders of tumors resembling normal brain tissue, and in guiding endoscopes where ventricles are small or vision is blurred. This recommendation applies to any neurosurgeon familiar with the technique and managing neurosurgical cases requiring precise topographical orientation where normal anatomic landmarks are missing. PMID- 11108111 TI - Endoscopic third ventriculostomy with ultrasonic contact microprobe. AB - The authors describe their first clinical experiences in endoscopic third ventriculostomy (ETV) with the original ultrasonic contact microprobe (UCM) designed at the Department of Neurosurgery in Zagreb. The analysis includes the clinical course of disease in eight patients submitted to surgery from May to September 1999 (3 men and 5 women, from 14 to 61 years of age). Surgery was performed in patients with neurological symptoms of elevated intracranial pressure and neuroradiological evidence of non-communicating hydrocephalus caused by mesencephalic aqueduct stenosis. The perforation in the base of the third brain ventricle made by the ultrasonic contact microprobe was widened by a balloon catheter. The authors have come to conclusion that the ETV when performed by contact ultrasonic microprobe is a small risk procedure in case of non communicating hydrocephalus. For its small diameter (1.6 mm) and simple handling the newly designed contact ultrasonic microprobe is very suitable for use in neuroendoscopy as it enables fenestration of the third brain ventricle with minimal thermal and ultrastructural damage to the adjacent neurovascular structures. Further research will be focused on defining indications for the use of the device in other neuroendoscopic procedures as well. PMID- 11108112 TI - Traumatic subependymal hematoma during endoscopic third ventriculostomy in a patient with a third ventricle tumor: case report. AB - Endoscopic third ventriculostomy has become a routine intervention for the treatment of non-communicating hydrocephalus. This technique is largely considered safe and a very low incidence of complications is reported. However, hemorrhage in the course of neuroendoscopy is still a problem difficult to manage. The authors present a case in which endoscopic third ventriculostomy and tumor biopsy were performed in a young patient with a huge tumor growing in the posterior part of the third ventricle. The surgical approach to realize the stoma was difficult because the tumor size reduced the third ventricle diameter. Surgical manipulation produced a traumatic subependymal hematoma. This hematoma drained spontaneously after few minutes into the ventricle and the blood was washed away. The postoperative neurological course was uneventful and the ventriculostomy showed to work well by reducing the size of the lateral ventricles and the intracranial pressure in three days. This complication during endoscopic third ventriculostomy has never been reported before. We emphasize the difficulty of endoscopic procedures in patients with huge tumors in the third ventricle. Where reduction in size of the third ventricle and of the foramen of Monro ist present we suggest a careful approach to the third ventricle. PMID- 11108113 TI - Combined endovascular-microsurgical treatment of tentorial-incisural dural arteriovenous malformations. Report of five cases. AB - Dural arteriovenous malformations of the tentorial edge are rare lesions with an unfavorable clinical course if left untreated. As yet no optimum treatment has been established. We retrospectively evaluated the results of our therapy concept, which consists of endovascular embolization followed by microsurgical obliteration, and compared the results to the relevant literature in order to clarify which therapeutic regimen seems to be optimum at the present time. In all of our five patients the lesion was completely obliterated, as proven by angiography, with favorable clinical results and low morbidity. Several other clinical series confirm our results, so we conclude that combined endovascular therapy and microsurgery is the optimum treatment for these lesions. PMID- 11108114 TI - Practicability of intraoperative microvascular Doppler sonography in aneurysm surgery. AB - Inadvertent narrowing of parent or branching vessels is one major cause of unfavorable outcome from aneurysm surgery. Intraoperative micro-Doppler sonography of arterial brain vessels during surgery for cerebral aneurysms of the anterior circulation was performed in 50 patients and compared retrospectively with 50 patients, who were operated upon without micro-Doppler sonography. Intraoperative micro-Doppler sonography demonstrated the need for repositioning of the clip in 12 instances. Outcome after surgery with micro-Doppler sonography appeared slightly better than without. Micro-Doppler sonography is concluded to be a practicable adjunct to routine aneurysm surgery. PMID- 11108115 TI - The difference between ultrasound-guided and stereotactic-guided neurosurgical procedures. AB - We evaluate two different methods, ultrasound (US) guidance and stereotactic guidance, routinely used in our Department for navigation in various neurosurgical procedures. We have performed 53 US-guided and 101 stereotactic guided procedures. These procedures were intracranial lesion biopsies, intracranial cysts and abscesses puncture and evacuations, ventricular punctures for hydrocephalus shunt operations, stereotactic-guided microneurosurgical resections, and stereotactic-guided endoscopic operations. Advantages of the US guided operations are the shortness of the procedure, simplicity (no need for moving patient for additional CT scanning), no irradiation and the possibility of real-time imaging. The disadvantages of the US-guided procedures are worse resolution of the images in deep-seated and small lesions as well as the need for a bigger trepanation because of the transducer's dimensions. Stereotactic procedures are time-consuming but more precise and usually done in local anaesthesia because only a small trepanation is required. Main disadvantage of the stereotactic-guided procedures when compared with the US-guided procedures is a lack of real-time intraoperative control. According to our experience, both methods are complementary and safe and they do not cause any additional complications when used as a navigation tool in microneurosurgical operations. Both methods are highly reliable when used in properly selected patients. PMID- 11108116 TI - Endoscopic treatment of para- and intraventricular cerebrospinal fluid cysts. AB - This study has been made to define the role of endoscopy and the most appropriate approach and technique of endoscopic fenestration of paraventricular and intraventricular CSF cysts according to the cyst size and location. Twenty-two patients with intraventricular (13 cases) and paraventricular (9 cases) CSF cysts, operated upon by endoscopic technique in three Italian neurosurgical centers, are reviewed. Paraventricular hemispheric cysts have been treated by endoscopic fenestration from the cyst to the lateral ventricle. Midline intraventricular cysts (2 of the septum pellucidum and 4 of the velum interpositum) underwent fenestration from the right lateral ventricle to the cyst, with fenestration in both lateral ventricles in one case. Cysts of the choroid plexus have been fenestrated from the homolateral enlarged ventricle (4 cases) or from the contralateral compressed ventricle (2 cases). Twenty patients (more than 90%) were definitively cured by the endoscopic procedure, whereas only 2 patients required a shunt or a direct approach. We think that the endoscopic fenestration must be considered the treatment of choice of intraventricular and paraventricular CSF cysts. PMID- 11108117 TI - Spinal epidural cavernous angioma: case report. AB - Epidural cavernous angiomas are apparently rare lesions. We present a case of paraplegia with acute onset secondary to spinal epidural cavernous angioma at levels C7 and T1. Magnetic resonance imaging gave the clinician the opportunity to diagnose the lesion preoperatively. In this article, we present and discuss this rare case with a review of the literature. PMID- 11108118 TI - Non-resective management of pineoblastoma. AB - The results of a non-resective treatment approach for pineoblastoma comprising stereotactic biopsy, cerebrospinal fluid diversion, and fractionated radiotherapy in six patients over a period of six years are presented. There were three male and three female patients, with a median age at diagnosis of 20 years. Magnetic resonance imaging of the spine, ventricular cerebrospinal fluid cytology, and tumour markers in cerebrospinal fluid were negative. Tumour response to initial radiotherapy was complete in three patients and partial in three patients. Recurrences were treated with interstitial irradiation with iodine-125 seeds in four instances, repeat radiotherapy when time elapsed was more than five years in one instance, with surgical resection in two instances, and chemotherapy in two instances. The diagnostic and therapeutic effectiveness of this management strategy is assessed. There were no complications related to surgical procedures. The median follow-up time was 48 months (range 14-70 months). Five patients were alive at 14, 45, 51, 57, and 70 months of follow-up. One patient died of disease at 28 months following diagnosis. The overall survival rate was 80 % +/- 17.89 % at 28 months. The results of this study suggest that this non-resective treatment approach is acceptable as an initial treatment alternative to radical surgical resection of pineoblastomas. PMID- 11108119 TI - Friedrich August von Ammon (1799-1861). AB - As this year marks the 200th anniversary of the birth of Friedrich August von Ammon (1799-1861), it presents an opportunity to recall and celebrate Ammon's life and his many contributions that helped shape both ophthalmology and plastic surgery into independent specialties. Some 250 years after Georg Bartisch, Ammon reestablished Dresden as a major center of ophthalmology, through the creation of a teaching institute, the publication of influential periodicals, and his personal publication of important monographs and journal articles. Ammon's Zeitschrift fur die Ophthalmologie (founded 1830) was only the third journal in history that had been entirely devoted to ophthalmology. His prize-winning treatise, De Iritide (1835), correctly categorized iritis and gave a detailed description of sympathetic ophthalmia. His beautiful color-plate atlas, Klinische Darstellungen (1838-1841), contains landmark descriptions of congenital eye anomalies and has been described by Norman as 'the best summary of the knowledge of diseases of the eye prior to the introduction of the ophthalmoscope.' Ammon made most of his literary contributions in ophthalmology but he also contributed to another emerging field, plastic surgery. His monograph, Die plastische Chirurgie (1842), critically and comprehensively surveyed the entire history and current practice of plastic surgery, one of the very first textbooks to achieve this goal. In his last monograph (1858) Ammon returned to the subject of embryology of the human eye and drew upon his 30 years of study. Before his death, two of his last papers reported ophthalmoscopic observations, demonstrating his exploration of the frontiers of ophthalmology that characterized his entire career. PMID- 11108120 TI - Helena B. Fedukowicz: pioneer educator in ocular microbiology. PMID- 11108121 TI - Three reading aids painted by Tomaso da Modena in the chapter house of San Nicolo Monastery in Treviso, Italy. AB - In 1352 the artist Tomaso da Modena depicted three monks with reading aids in the chapter house of San Nicolo Monastery. One of them is wearing rivet spectacles, the second is using a reading glass, and the third has a reading glass on a stand. These rivet spectacles are often quoted in the literature as the first pictorial representation of spectacles, while the reading glass was found in only one reference in the literature with only one illustration, and the reading glass with stand was not found at all. PMID- 11108122 TI - Werner Ernst Reichardt Ph.D: founder of modern computational visual neurophysiology and anti-Nazi resistance fighter. AB - Werner Ernst Reichardt was born on January 30, 1924 in Berlin and at age 19 was drafted into the Luftwaffe and assigned to an electronic signals section laboratory. He became an active member of a resistance group and supplied radios for the movement in Germany. He emerged from the ashes of the Second World War and dedicated his scientific life to the development of the newborn specialty of biological physics. Following graduation from the Technische Hochschule Charlottenburg, he did a fellowship at CalTech under Max Delbruck. On returning to Germany he joined the Max Planck Institut and later became Director of the Max Planck Institut fur Biologische Kybernetik in Tubingen, West Germany. Reichardt was one of the founders of the quantitative study of visually controlled orientation in animals. His work is very nearly unique in its close dialectic between elegant non-linear mathematical theory and quantitative experimental test of their predictions. During the 1950s Reichardt and his collaborators jointly developed an autocorrelation model (i.e. the firing rate of the involved visual neurones is closely correlated with the features of the pattern stimulating them) of how moving patterns are perceived by motion detectors in the visual system of the fly. This was the first mathematical description of a biological abstraction process. His findings apply to vertebrate vision, including motion detection and figure-ground description in human vision. His Max Planck Institute became a world renowned center for the computational approach to information processing by the nervous system. At his retirement party from the Institute he founded, Reichardt died on the evening of September 11th, 1992. PMID- 11108123 TI - Harold Gifford, Sr., M.D. Omaha's premier turn of the century ophthalmologist. PMID- 11108124 TI - George Handel and his blindness. PMID- 11108125 TI - Retinal detachment surgery: historical notes. PMID- 11108126 TI - Julian John Chisholm M.D.: Confederate surgeon, ophthalmologist and hospital founder. PMID- 11108127 TI - Robert Koch's debt to Ferdinand Cohn. PMID- 11108128 TI - Eye surgery in Stockholm at the beginning and at the end of the 20th century: a comparative study. PMID- 11108129 TI - First known lenses originating in Egypt about 4600 years ago. PMID- 11108130 TI - Mechanisms of transcriptional activation of cAMP-responsive element-binding protein CREB. AB - The CREB-CREM transcription factors are the main gene regulatory effectors of the cAMP signaling pathway. The investigations of this family of transcription factors had a profound impact on the understanding of signaling-induced gene transcription. Here we discuss some key aspects of the underlying biology, review transcriptional activation by CREB proteins through transcription cofactors and present novel insights into the context- and position-specific function of CREB on complex genes. PMID- 11108131 TI - Transcriptional regulation by cAMP in the heart. AB - Stimulation of the cAMP-dependent signalling pathway by beta-adrenergic catecholamines is an important physiological mechanism to increase contractile force in the heart. In addition to this, long-term beta-adrenergic stimulation by elevated catecholamines also influences the expressional control of functionally relevant cardiac regulatory proteins in human heart failure. The regulation of transcription by the cAMP-response element (CRE) is an important mechanism for a cAMP-mediated control of gene expression involved e.g. in spermiogenesis and memory/learning processes. This article discusses recent data leading to the hypothesis that this mechanism also contributes to altered gene regulation in heart failure. PMID- 11108132 TI - Regulation of tumor growth and metastasis of human melanoma by the CREB transcription factor family. AB - The purpose of this study was to determine the role of CREB and its associated proteins in melanoma progression. We used MeWo human melanoma cells transfected with a dominant negative construct of CREB, KCREB. KCREB has a mutation in its DNA-binding domain and can not bind the CRE element. Expression of KCREB yields proper heterodimerization with CREB and its associated proteins, but the proteins associated with KCREB do not confer the same degree of transcriptional activity as they would in the case of wild-type CREB. Here, we demonstrate that expression of KCREB in MeWo melanoma cells leads to a decrease in their tumorigenicity and metastatic potential in nude mice. We identified two mechanisms that explain at least partially this effect of KCREB. The first, is one in which CREB and its associated proteins play an essential role in invasion. We showed that the invasive properties of KCREB-transfected MeWo cells were reduced due to the downregulation of the CRE-dependent expression of the type IV collagenase MMP-2 and the adhesion molecule MCAM/MUC18. In the second mechanism, CREB and its associated proteins act as survival factors for human melanoma cells. Here we demonstrated that expression of KCREB in MeWo cells rendered them susceptible to apoptosis induced by thapsigargin, which in turn increased the intracellular level of Ca2+. Thapsigargin induced CREB and ATF-1 phosphorylation and activated CRE-dependent transcription in MeWo cells. Collectively, our data demonstrate that CREB and its associated proteins play an important role in tumor growth and metastasis of human melanoma. PMID- 11108133 TI - CRE-decoy oligonucleotide-inhibition of gene expression and tumor growth. AB - Nucleic acid molecules with high affinities for a target transcription factor can be introduced into cells as decoy cis-elements to bind these factors and alter gene expression. This review discusses a synthetic single-stranded palindromic oligonucleotide, which self-hybridizes to form a duplex/hairpin and competes with cAMP response element (CRE) enhancers for binding transcription factors. This oligonucleotide inhibits CRE- and Ap-1-directed gene transcription and promotes growth inhibition in vitro and in vivo in a broad spectrum of cancer cells, without adversely affecting normal cell growth. Evidence presented here suggests that the CRE-decoy oligonucleotide can provide a powerful new means of combating cancers, viral diseases, and other pathological conditions by regulating the expression of cAMP-responsive genes. PMID- 11108134 TI - Attenuation of macrophage apoptosis by the cAMP-signaling system. AB - Previous studies revealed that expression and activation of cyclooxygenase-2 (Cox 2) conveyed a protective principle in murine macrophages, thus attenuating pro apoptotic actions of chemotherapeutic agents or programmed cell death as a result of massive nitric oxide (NO) generation. Expression of Cox-2 was achieved by treatment of cells with lipopolysaccharide/interferon-gamma or nontoxic doses of NO releasing agents. We reasoned E-type prostanoid formation, and in turn an intracellular cAMP increase as the underlying protective mechanism. To prove our hypothesis, we analyzed the effects of lipophilic cAMP-analogs on NO, cisplatin, or etoposide induced apoptosis in RAW 264.7 macrophages. Selected apoptotic parameters comprised DNA fragmentation (diphenylamine assay), annexin V staining of phosphatidylserine, caspase activity (quantitated by the cleavage of a fluorogenic caspase-3-like substrate Ac-DEVD-AMC), and mitochondrial membrane depolarisation (delta psi). Western blots detected accumulation of the tumor suppressor protein p53, relocation of cytochrome c to the cytosol, and expression of the anti-apoptotic protein Bcl-xL. Prestimulation with lipophilic cAMP-analogs attenuated apoptosis with the notion that cell death parameters were basically absent. To verify gene induction by cAMP in association with protection we established activation of cAMP response element binding protein (CREB) by gel shift analysis and moreover, treated macrophages with oligonucleotides containing a cAMP-responsive element (CRE) in order to scavenge CREB. Decoy oligonucleotides, but not control oligonucleotides, attenuated cAMP-evoked protection and reestablished pro-apoptotic parameters. We conclude that gene induction by cAMP protects macrophages towards apoptosis that occurs as a result of excessive NO formation or addition of chemotherapeutica. Attenuating programmed cell death by the cAMP-signaling system may be found in association with Cox-2 expression and tumor formation. PMID- 11108135 TI - Catecholamines induce IL-10 release in patients suffering from acute myocardial infarction by transactivating its promoter in monocytic but not in T-cells. AB - The anti-inflammatory cytokine IL-10 is up-regulated in response to TNF-alpha suggesting a control mechanism of inflammation. In addition, we recently found systemic IL-10 release in response to acute stress reactions in the absence of any systemic inflammation. In vitro and in vivo studies in experimental models suggest that catecholamines induce IL-10 release via a cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) dependent pathway. Here we studied patients for plasma IL-10 after acute myocardial infarction, a very stressful event without significant signs of systemic inflammation. In fact, the activation of the sympathetic system initiated by cardiac infarction was accompanied by a temporary systemic release of IL-10. Catecholamine induced IL-10 may be released by different cells. Recently, we demonstrated that catecholamines directly stimulate the IL-10 promoter/enhancer via a cAMP/PKA pathway in monocytic cells. A cAMP responsive element (CRE) was identified as major target. Here we show that there is no influence of catecholamines on the IL-10 promoter activity in T cells. In contrast to monocytic cells, in T-cells cAMP-induced PKA-dependent phosphorylation of the CRE-binding protein 1 (CREB-1) seems to play a marginal role in IL-10 induction, which was reflected by a low cAMP-dependent IL-10 promoter/enhancer stimulation in reporter gene assays. Thus, catecholamines are directly involved in the regulation of IL-10 expression in monocytic but not in T cells after acute stressful conditions. PMID- 11108136 TI - Regulation of tyrosine hydroxylase gene transcription by the cAMP-signaling pathway: involvement of multiple transcription factors. AB - The conversion of L-tyrosine to 3,4-dihydroxy-L-phenylalanine by tyrosine hydroxylase (TH) is the first and rate-limiting step in biosynthesis of catecholamine neurotransmitters. TH gene expression is regulated in a cell type specific and cAMP-dependent manner. Evidence from this laboratory and others indicates that the cAMP response element (CRE), residing at -45 to -38 bp upstream of the transcription initiation site, is essential for both basal and cAMP-inducible transcription of the TH gene. To understand the control mechanisms of TH gene transcription in greater detail, we sought to identify and characterize the transcription factors involved in recognition and activation of the CRE of the TH gene. Remarkably, electrophoretic mobility shift assay and antibody supershift experiments indicated that all three major CRE-binding protein factors, i.e. CREB, ATF1, and CREM, may participate in forming specific DNA/protein complexes with the CRE of the TH gene. To address the transcriptional activation function of individual factors, we replaced the TH CRE with a GAL4 binding site and cotransfected this modified TH promoter-reporter gene with an effector plasmid that encodes GAL4-fused transcription factor. Our results indicate that CREB but not ATF1 can support basal promoter activity while both can robustly induce the promoter activity in response to co-expression of the catalytic subunit of cAMP-dependent protein kinase (PKA). We further show that the coactivator CBP up-regulates PKA-mediated activation of the TH promoter and, if tethered to the TH promoter by a GAL4-fusion, can robustly transactivate the TH promoter even in the absence of PKA. Collectively, our results suggest that multiple CRE-binding factors interact with the CRE and regulate, in conjunction with the coactivator CBP, the transcriptional activity of the TH gene. PMID- 11108137 TI - Norepinephrine transporter expression and function in noradrenergic cell differentiation. AB - The classical view of norepinephrine transporter (NET) function is the re-uptake of released norepinephrine (NE) by mature sympathetic neurons and noradrenergic neurons of the locus ceruleus (LC; [1-3]). In this report we review previous data and present new results that show that NET is expressed in the young embryo in a wide range of neuronal and non-neuronal tissues and that NET has additional functions during embryonic development. Sympathetic neurons are derived from neural crest stem cells. Fibroblast growth factor-2 (FGF-2), neurotrophin-3 (NT 3) and transforming growth factor-beta1 (TGF-beta1) regulate NET expression in cultured quail neural crest cells by causing an increase in NET mRNA levels. They also promote NET function in both neural crest cells and presumptive noradrenergic cells of the LC. The growth factors are synthesized by the neural crest cells and therefore are likely to have autocrine function. In a subsequent stage of development, NE transport regulates differentiation of noradrenergic neurons in the peripheral nervous system and the LC by promoting expression of tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH). Conversely, uptake inhibitors, such as the tricyclic antidepressant, desipramine, and the drug of abuse, cocaine, inhibit noradrenergic differentiation in both tissues. Taken together, our data indicate that NET is expressed early in embryonic development, NE transport is involved in regulating expression of the noradrenergic phenotype in the peripheral and central nervous systems, and norepinephrine uptake inhibitors can disturb noradrenergic cell differentiation in the sympathetic ganglion (SG) and LC. PMID- 11108139 TI - cAMP increases the expression of human angiotensinogen gene through a combination of cyclic AMP responsive element binding protein and a liver specific transcription factor. AB - Angiotensinogen is the glycoprotein precursor of one of the most potent vasoactive hormones angiotensin-II which plays an important role in the regulation of blood pressure. We show here that the promoter activity of reporter constructs containing human angiotensinogen promoter is increased by cAMP treatment on transient transfection in HepG2 cells. We have identified a composite cAMP responsive element, located around 840 bases upstream from the transcriptional initiation site, in the promoter of human angiotensinogen gene. This element is recognized by members of CREB/ATF as well as C/EBP family of transcription factors. Another C/EBP binding site that is not recognized by CREB is located 10 bases upstream from this site. We show that co-transfection of CREB increases the promoter activity of reporter constructs containing human angiotensinogen gene promoter attached to the CAT gene. We also show that co transfection of DBP (which is a member of C/EBP family of transcription factors) increases promoter activity of these reporter constructs. PMID- 11108138 TI - Catecholamines and angiotensinogen gene expression in kidney proximal tubular cells. AB - To investigate the molecular mechanism(s) of action of catecholamines on the expression of the angiotensinogen (ANG) gene in kidney proximal tubular cells, we used opossum kidney (OK) cells with a fusion gene containing the 5'-flanking regulatory sequence of the rat ANG gene fused with a human growth hormone (hGH) gene as a reporter, pOGH (rANG N-1498/+18), permanently integrated into their genomes. The level of expression of the ANG-GH fusion gene was quantified by the amount of immunoreactive-hGH (IR-hGH) secreted into the medium. The addition of norepinephrine (NE), isoproterenol (a beta1/beta2-adrenergic receptor (AR) agonist) and iodoclonidine (an alpha2-AR agonist) stimulated the expression of the ANG-GH fusion gene in a dose-dependent manner, whereas the addition of epinephrine and phenylephrine (alpha1-AR agonist) had no effect. The stimulatory effect of NE was blocked by the presence of propranolol (beta-AR blocker), atenolol (beta1-AR blocker), yohimbine (alpha2-AR blocker), Rp-cAMP (an inhibitor of cAMP-dependent protein kinase AI & AII) and staurosporine (an inhibitor of protein kinase C), but was not blocked by ICI 118, 551 (beta2-AR blocker) and prazosin (alpha1-AR blocker). The addition of a combination of isoproterenol and iodoclonidine or a combination of 8-Bromo-cAMP (8-Br-cAMP) and phorbol 12 myristate (PMA) synergistically stimulated the expression of the ANG-GH fusion gene as compared to the addition of isoproterenol, iodoclonidine, 8-Br-cAMP or PMA alone. Furthermore, the addition of NE, 8-Br-cAMP or PMA stimulated the expression of pOGH (rANG N-806/-779/-53/+18), a fusion gene containing the putative cAMP responsive element (CRE, ANG N-806/-779) upstream of the ANG promoter (ANG N-53/+18) in OK cells, but had no effect on the expression of fusion genes containing the mutant of the CRE. Gel mobility shift assays revealed that the ANG-CRE binds with the DNA-binding domain (bZIP254-327) of the cAMP responsive binding protein (CREB). The binding of the labeled ANG-CRE to CREB (bZIP254-327) was displaced by unlabeled ANG-CRE and the CRE of the somatostatin gene but not by the mutants of the ANG-CRE. Finally, NE stimulated the phosphorylation of CREB in OK cells. These studies demonstrate that the molecular mechanism(s) of NE action on the expression of the ANG gene in OK cells may be mediated via both the PKA and PKC signalling pathways and via the phosphorylation of CREB. The phosphorylated CREB then interacts with the CRE in the 5'-flanking region of the ANG gene and subsequently stimulates the gene expression. PMID- 11108140 TI - The role of Ca2+ mobilization and heterotrimeric G protein activation in mediating tyrosine phosphorylation signaling patterns in vascular smooth muscle cells. AB - This work investigated the role of Ca2+ mobilization and heterotrimeric G protein activation in mediating angiotensin II-dependent tyrosine phosphorylation signaling patterns. We demonstrate that the predominant, angiotensin II dependent, tyrosine phosphorylation signaling patterns seen in vascular smooth muscle cells are blocked by the intracellular Ca2+ chelator BAPTA-AM, but not by the Ca2+ channel blocker verapamil. Activation of heterotrimeric G proteins with NaF resulted in a divergent signaling effect; NaF treatment was sufficient to increase tyrosine phosphorylation levels of some proteins independent of angiotensin II treatment. In the same cells, NaF alone had no effect on other cellular proteins, but greatly potentiated the ability of angiotensin II to increase the tyrosine phosphorylation levels of these proteins. Two proteins identified in these studies were paxillin and Jak2. We found that NaF treatment alone, independent of angiotensin II stimulation, was sufficient to increase the tyrosine phosphorylation levels of paxillin. Furthermore, the ability of either NaF and/or angiotensin II to increase tyrosine phosphorylation levels of paxillin is critically dependent on intracellular Ca2+. In contrast, angiotensin II mediated Jak2 tyrosine phosphorylation was independent of intracellular Ca2+ mobilization and extracellular Ca2+ entry. Thus, our data suggest that angiotensin II-dependent tyrosine phosphorylation signaling cascades are mediated through a diverse set of signaling pathways that are partially dependent on Ca2+ mobilization and heterotrimeric G protein activation. PMID- 11108142 TI - The inducible cAMP early repressor ICERIIgamma inhibits CREB and AP-1 transcription but not AT1 receptor gene expression in vascular smooth muscle cells. AB - Growth factors and agonists of both G alpha s and G alpha q-coupled receptor agonists each down regulate angiotensin AT1 receptor gene expression in vascular smooth muscle cells. To test whether the transcription factor CREB is in the pathway involved in modulation of AT1-receptor gene expression, AT1 receptor mRNA levels were measured after stimulation with forskolin, angiotensin II and PDGF-BB in VSMC expressing the inducible cAMP early repressor protein, ICERIIgamma, fused to the green fluorescent protein (eGFP). This fusion protein inhibits luciferase induction from promoters driven by either CREB response elements (CRE) or, unexpectedly, by tetradecanoylphorbol acetate response elements (TRE), which bind activator protein (AP-1). EGFP-ICERIIgamma expression inhibits occupancy of CRE and TRE elements by endogenous VSMC nuclear proteins. Nevertheless, agonist induced AT1-receptor mRNA down regulation is unaffected by eGFP-ICERIIgamma expression. Although CREB transcription is activated by multiple classes of agonists in VSMC, and ICER can inhibit agonist-induced transcription mediated by both CREB and AP-1 enhancers, these transcription factors are not obligate components of the pathways regulating AT1-R gene expression in rat VSMC. PMID- 11108141 TI - Functional cross-talk between the cyclic AMP and Jak/STAT signaling pathways in vascular smooth muscle cells. AB - Angiotensin II (Ang II), the primary effector of the renin-angiotensin system, is a multifunctional hormone that plays an important role in vascular function. In addition to its classical vasoconstrictor action, more recent studies demonstrated that Ang II stimulates the growth of a number of cell types, including vascular smooth muscle cells (SMC) (reviewed in [1-3]). In vivo studies have shown that chronic infusion of Ang II leads to the development of vascular hypertrophy in rats, whereas administration of angiotensin-converting enzyme (ACE) inhibitors or Ang II receptor antagonists prevents or regresses vascular hypertrophy in models of genetic and experimental hypertension [4]. Consistent with in vivo data, several laboratories have shown that Ang II stimulates protein synthesis and induces cellular hypertrophy, but not cell proliferation, in cultured aortic SMC [5-9]. Ang II also induces directed migration (chemotaxis) of vascular SMC [10, 11], although its effect is less prominent than that of platelet-derived growth factor (PDGF). The cellular mechanisms underlying these diverse actions of Ang II are not clearly understood but are likely to involve the activation of distinct signaling pathways. PMID- 11108143 TI - Angiotensin II-induced changes in G-protein expression and resistance of renal microvessels in young genetically hypertensive rats. AB - Altered regulation of cAMP may contribute to enhanced renal reactivity to angiotensin II (Ang II) in spontaneously hypertensive rats (SHR). Such a phenomenon may occur in renal preglomerular arterioles and may involve changes in expression of GTP-binding regulatory proteins. We have examined the effects of Ang II on steady state levels of G(alpha i-1,2), G(alpha i-3), G(alpha s) and G(alpha q) in preglomerular arterioles from young marginally hypertensive SHR and on mean arterial pressure (MAP), renal vascular resistance (RVR) and renal cAMP excretion (UcAMP.V). Young (5-6 week old) SHR and Wistar Kyoto (WKY) rats received Ang II (35 ng/kg/min, s.c.) or vehicle for 7 days via osmotic minipumps. Urine was collected over the last 24 h. On day seven, MAP and renal blood flow were measured in anesthetized rats and RVR was determined. Preglomerular arterioles were isolated by perfusing the kidneys with iron oxide and using a series of mechanical steps coupled with the use of a magnet to retain iron-laden vessels. Membranes were prepared and the expressions of G(alpha i-1,2), G(alpha i 3), G(alpha s) and G(alpha q) were evaluated by Western immunoblotting. Baseline MAP (124 +/- 6 mmHg) was only marginally (p > 0.05) higher in SHR when compared with WKY rats (110 +/- 4 mmHg). RBF (3.04 +/- 0.16 mL/min) was significantly lower and RVR (41.10 +/- 1.37 mmHg.min/mL) was significantly higher in SHR when compared to age-matched WKY rats (4.36 +/- 0.30 mL/min and 25.79 +/- 1.58 mmHg.min/mL, respectively). Ang II significantly increased MAP in SHR (17 mmHg) but not in WKY rats. These increases in MAP were accompanied by significant increases in RVR in SHR (48% over control) but not in WKY rats. Compared to WKY rats, preglomerular arterioles from SHR exhibited significantly higher basal expression of G(alpha i-1,2) (11- fold), G(alpha 1-3) (13-fold) and G(alpha s) (3 fold). Chronic infusion of Ang II, however, downregulated the expression of G(alpha s) (by 53%; p < 0.05), G(alpha i-1,2) (by 72%; p < 0.05) and G(alpha i-3) (by 35%; p > 0.05) in SHR preglomerular arterioles but significantly upregulated the expression of these proteins in WKY by 3-, 8- and 15-fold, respectively. Basal levels of G(alpha q) were not different in preglomerular arterioles from the two strains but were downregulated by Ang II in both WKY (74% of basal) and SHR (52% of control). Baseline UcAMP.V was significantly lower in SHR (31.22 +/- 6.51 nmol/24 h) compared with WKY rats (65.33 +/- 3.60 nmol/24 h). Chronic Ang II infusion significantly increased UcAMP.V in SHR as well as WKY rats. These data clearly demonstrate that expressions of Gi isoforms as well as Gs in renal microvessels are elevated during early stages of hypertension and suggest that the elevated levels of Gi proteins may be directly associated with a blunted adenylyl cyclase-cAMP cascade in the renal microvasculature. Furthermore, Ang II appears to directly downregulate the expression of Gs in young SHR but not in young WKY renal microvessels. Such diversity in its effect on G-protein expression may be important for enhanced renal sensitivity to Ang II in SHR. PMID- 11108144 TI - Angiotensin receptor II is present in dopaminergic cell line of rat substantia nigra and it is down regulated by aminochrome. AB - Angiotensin receptor II mRNA was found to be expressed in dopaminergic neuronal cell line RCSN3 of rat substantia nigra using RT-PCR reaction. Aminochrome (150 microM), a metabolite of the dopamine oxidative pathway, was found to down regulate the expression of angiotensin receptor mRNA in RCSN3 cells by 83% (p < 0.05). PMID- 11108145 TI - Control of cardiomyocyte gene expression as drug target. AB - Pressure overload of the heart is associated with a perturbed gene expression of the cardiomyocyte leading to an impaired pump function. The ensuing neuro endocrine activation results in disordered influences of angiotensin II and catecholamines on gene expression. To assess whether angiotensin II type 1 receptor inhibition can also counteract a raised sympathetic nervous system activity, spontaneously hypertensive rats fed a hypercaloric diet were treated with eprosartan (daily 90 mg/kg body wt) and cardiovascular parameters were monitored with implanted radiotelemetry pressure transducers. Both, blood pressure and heart rate were increased (p < 0.05) by the hypercaloric diet. Although eprosartan reduced (p < 0.05) the raised systolic and diastolic blood pressure, the diet-induced rise in heart rate was blunted only partially. In addition to drugs interfering with the enhanced catecholamine influence, compounds should be considered that selectively affect cardiomyocyte gene expression via 'metabolic' signals. PMID- 11108146 TI - Designing antisense to inhibit the renin-angiotensin system. AB - Overactive renin-angiotensin system has been indicated in numerous pathological situations. Current treatment is based on pharmaceutical compounds, which work on the proteins level. Undisputedly helpful, it is not, however, flawless. Some of the drawbacks include adverse effects and non-compliance problem, since in many cases medicine has to be taken at least once a day for a long time. Therefore it seems logical to try a different approach, for instance to correct the disease at the gene expression level, possibly having a choice of shorter or longer-lasting effects. This current review combines results, relevant to the angiotensin system, with the antisense approach, which decreases amount of target protein by interfering at the mRNA level. Dependent on the tool used--oligodeoxynucleotide, plasmid or viral vector, the antisense effect lasts from few days to months. PMID- 11108148 TI - The role of Jak/STAT signaling in heart tissue renin-angiotensin system. AB - The involvement of the Renin Angiotensin System (RAS) and the role of its primary effector, angiotensin II (Ang II), in etiology of myocardial hypertrophy and ischemia is well documented. In several animal models, the RAS is activated in cardiac cell types that express the receptor AT1, and/or AT2, through which the Ang II mediated effects are promoted. In this article, we briefly review recent experimental evidence on the critical role of a prominent signaling pathway, the Jak/STAT pathway in activation and maintenance of the local RAS in cardiac hypertrophy and ischemia. Recent studies in our laboratory document that the promoter of the prohormone angiotensinogen (Ang) gene serves as the target site for STAT proteins, thereby linking the Jak/STAT pathway to activation of heart tissue autocrine Ang II loop. STAT5A and STAT6, are selectively activated when the heart is subjected to ischemic injury, whereas activation of STAT3 and STAT5A is involved in myocardial hypertrophy. Blockage of RAS activation by treatment with specific inhibitor promotes a remarkable recovery in functional hemodynamics of the myocardium. Thus, activation of selective sets of STAT proteins constitutes the primary signaling event in the pathogenesis of myocardial hypertrophy and ischemia. PMID- 11108147 TI - Angiotensin II induces gene transcription through cell-type-dependent effects on the nuclear factor-kappaB (NF-kappaB) transcription factor. AB - The vasopressor octapeptide, angiotensin II (Ang II), exerts homeostatic responses in cardiovascular tissues, including the heart, blood vessel wall, adrenal cortex and liver (a major source of circulating plasma proteins). One of the effects of Ang II is to induce expression of regulatory, structural and cytokine genes that play important roles in long-term control of blood pressure, vascular remodeling, cardiac hypertrophy and inflammation. The identification of nuclear signaling pathways and target transcription factors has provide important insight into cellular responses and the spectrum of genes controlled by Ang II. Here we will review how Ang II activates the transcription factors, Activator Protein 1 (AP-1), Signal Transducer and Activator of Transcription (STATs), and Nuclear Factor-kappaB (NF-kappaB). NF-kappaB is of particular interest because it is an important mediator of resynthesis of the Ang II precursor, angiotensinogen AGT. Through this positive feedback loop, long-term changes in the activity of the renin angiotensin system occur. Although NF-kappaB is ubiquitously expressed, surprisingly the mechanism for Ang II-inducible NF-kappaB regulation differs between aortic smooth muscle cells (VSMCs) and hepatocytes. In VSMC, Ang II induces nuclear translocation of cytoplasmic transactivatory NF-kappaB proteins through proteolysis of its inhibitor, IkappaB. By contrast, in hepatocytes, Ang II induces large nuclear isoforms of NF-kappaB1 to bind DNA through a mechanism independent of changes in IkappaB turnover. NF-kappaB activation depends upon the activity of DAG-sensitive PKC isoforms and ROS signaling pathway. These observations indicate that significant differences exist in Ang II signaling depending upon cell-type involved and suggest the possibility that tissue selective modulation of Ang II effects is possible in the cardiovascular system. PMID- 11108149 TI - Rho plays an important role in angiotensin II-induced hypertrophic responses in cardiac myocytes. AB - Angiotensin II (Ang II) evokes a variety of hypertrophic responses such as activation of protein kinases, reprogramming of gene expressions and an increase in protein synthesis in cardiac myocytes. In this study, we examined the role of Rho family small GTP binding proteins (G proteins) in Ang II-induced cardiac hypertrophy. Ang II strongly activated extracellular signal-regulated protein kinases (ERKs) in cardiac myocytes of neonatal rats. Although Ang II-induced activation of ERKs was completely suppressed by an Ang II type 1 receptor antagonist, CV-11974, this activation was not inhibited by the pretreatment with C3 exoenzyme, which abrogates Rho functions. Overexpression of Rho GDP dissociation inhibitor (Rho-GDI), dominant negative mutants of Rac1 (D.N.Rac1), or D.N.Cdc42 had no effects on Ang II-induced activation of transfected ERK2. The promoter activity of skeletal alpha-actin and c-fos genes was increased by Ang II, and the increase was partly inhibited by overexpression of Rho-GDI and the pretreatment with C3 exoenzyme. Ang II increased phenylalanine incorporation into cardiac myocytes by approximately 1.4 fold as compared with control, and this increase was also significantly suppressed by the pretreatment with C3 exoenzyme. These results suggest that the Rho family small G proteins play important roles in Ang II-induced hypertrophic responses in cardiac myocytes. PMID- 11108150 TI - Mechanisms of angiotensin II-induced platelet-derived growth factor gene expression. AB - Angiotensin II induces the expression of platelet-derived growth factor A-chain and B-chain in cultured vascular smooth muscle cells at the level of transcription. The renin-angiotensin system has also been implicated in the increased expression of platelet-derived growth factor in the mechanically injured artery wall. This review is concerned with recent developments in our understanding of the signaling and transcriptional mechanisms mediating the inducible expression of one vasoconstrictor by another. PMID- 11108151 TI - Transactivation of EGF receptor induced by angiotensin II regulates fibronectin and TGF-beta gene expression via transcriptional and post-transcriptional mechanisms. AB - The signaling cascade elicited by angiotensin II (Ang II) resembles that characteristic of growth factor, and recent evidence indicates transactivation of epidermal growth factor receptor (EGF-R) by G protein-coupled receptors. Here, we report the involvement of EGF-R in Ang II-induced synthesis of fibronectin and TGF-beta in cardiac fibroblasts. Ang II stimulated fibronectin mRNA levels dose dependently with a maximal increase (approximately 5-fold) observed after 12 h of incubation. Ang II-, or calcium ionophore-induced fibronectin synthesis was completely abolished by tyrosine kinase inhibitors and intracellular Ca2+ chelating agents. Ang II-induced fibronectin mRNA was not affected by PKC inhibitors or PKC depletion, whereas specific inhibition of EGF-R function by a dominant negative EGF-R mutant and tyrphostin AG1478 abolished induction of fibronectin mRNA. We isolated the rat fibronectin gene including the 5'-flanking region and found that the AP-1 binding site present in the promoter region was responsible for the Ang II responsiveness of this gene. Gel retardation assay revealed the binding of nuclear protein to the AP-1 site, which was supershifted with anti-c-fos and anti-c-jun but not anti-ATF-2 antibodies. Conditioned medium from Ang II-treated cells contained TGF-beta bioactivity and addition of neutralizing TGF-beta antibody modestly (46%) inhibited induction of fibronectin. Ang II-induced synthesis of TGF-beta was also abolished by inhibition of EGF-R function. The effect of TGF-beta was exerted by stabilizing fibronectin mRNA without affecting the promoter activity and required de novo protein synthesis. We concluded that Ang II-induced expression of fibronectin and TGF-beta is mediated by downstream signaling of EGF-R transactivated by Ca2+-dependent tyrosine kinase, and that Ang II-induced fibronectin mRNA expression is regulated by two different mechanisms; transcriptional control by binding of c-fos/c-jun complex to the AP-1 site, and post-transcriptional control by mRNA stabilization due to autocrine and/or paracrine effects of TGF-beta. Thus, this study suggested that the action of Ang II on extracellular matrix formation should be interpreted in association with the EGF-R signaling cascade. PMID- 11108153 TI - Regulation of angiotensin II receptors in the medullary thick ascending limb. AB - Angiotensin II (Ang II) is an important regulator of the function of medullary thick ascending limb of loop of Henle (MTAL). Recent studies showed that changes in Ang II receptor expression occur and underlie changes in the function of proximal tubules during altered sodium intake. The present experiment was designed to determine (1) whether expression of the type 1 Ang II (AT1) receptor in the MTAL is regulated by altered sodium intake, and (2) the specific pathway(s) mediating sodium-induced AT1 expression in the MTAL. Wistar rats were fed a normal sodium (0.5%, NS), low sodium (0.07%, LS), or high sodium (4%, HS) diet for 2 weeks. Northern blot analysis and radioligand binding showed that in rats fed a normal sodium diet the rank of order for both AT1 mRNA expression and receptor density was outer medulla > cortex > inner medulla. Sodium restriction significantly increased both AT1 mRNA expression and receptor density in the outer medulla. In contrast, neither AT1 mRNA expression nor receptor density in the outer medulla was altered by sodium loading. Losartan treatment (3 mg/kg/per day by oral gavage for 2 weeks) prevented low sodium-induced upregulation of the AT1 receptor in the outer medulla, but it had no effect on AT1 expression in the outer medulla of rats fed a normal sodium diet. Highly purified suspensions of MTAL were isolated from rats fed a normal or low sodium diet. Low sodium intake significantly increased AT1 mRNA level by 184% and AT1 receptor density by 58% in MTALs. Primary cultures of MTAL cells were treated with PBS, Ang II (10(-8) M), and Ang II + 17 octadecynoic (17 ODYA, 10 microM). Ang II caused about 2-fold increase in AT1 mRNA levels, and this increase was diminished by about 30% by the addition of 17 ODYA. We conclude that (1) sodium restriction but not sodium loading increases AT1 receptor expression in the MTAL, (2) low sodium-induced upregulation of the AT1 receptor in the MTAL is Ang II-dependent, and (3) Ang II induced upregulation of the AT1 receptor in the MTAL is mediated, at least in part, by cytochrome P450 pathways. PMID- 11108152 TI - Expression of renin-angiotensin system and extracellular matrix genes in cardiovascular cells and its regulation through AT1 receptor. AB - Angiotensinogen (AGT) is a unique substrate of the renin-angiotensin system and fibronectin (FN) is an important component of the extracellular matrix. These play critical roles in the pathophysiological changes including cardiovascular remodeling and hypertrophy in response to hypertension. This study was performed to examine the regulation of AGT and FN gene in cardiac myocytes (CMs) and vascular smooth muscle cells (VSMCs) in response to mechanical stretch. Mechanical stretch significantly increased the AGT mRNA expression in CMs, while these stimuli did not affect FN mRNA levels. On the other hand, mechanical stretch upregulated FN mRNA levels in VSMCs, whereas no increase in AGT mRNA levels was observed in response to stretch stimuli. An angiotensin II type 1 (AT1) receptor antagonist (CV11974) significantly decreased these stretch mediated increases in mRNA level and promoter activity of the AGT and FN gene, whereas angiotensin II type 2 (AT2) receptor antagonist (PD 123319) did not affect the induction. These results indicate that mechanical stretch activates transcription of the AGT and FN gene mainly via AT1 receptor-pathway in CMs and VSMCs. Furthermore, mechanisms regulating AGT and FN gene seem to be different between CMs and VSMCs. PMID- 11108154 TI - Bradykinin (B2) independent effect of captopril on the development of pressure overload cardiac hypertrophy. AB - Besides the reduction of angiotensin II formation, locally increased kinins may play a role in the cardiovascular action of angiotensin converting enzyme (ACE) inhibitors. To characterize the contribution of bradykinin to the effects of ACE inhibition by captopril on the development of pressure overload hypertrophy, sham operated rats and rats with ascending aortic constriction were treated with captopril (80 mg/kg/day) or captopril and B2 kinin receptor antagonist HOE 140 (0.5 mg/kg/day) for 7 weeks. Left ventricular mass and geometry, hydroxyproline concentration and myosin isozymes (marker of a fetal phenotype) were assessed. Rats with aortic constriction exhibited a marked increase in left ventricular weight and diastolic pressure-volume relationship was shifted to smaller volumes. Signs of congestive heart failure were not apparent. The hydroxyproline concentration remained unaltered. However, the proportion of isomyosin V3 was increased (p < 0.05). Administration of captopril reduced (p < 0.05) systolic blood pressure, body and cardiac weight in all treated rats. The reduction of left ventricular weight was disproportionally higher in pressure overloaded rats, thus the relative left ventricular weight decreased by 15% (p < 0.05). Captopril augmented the isomyosin V1 expression (p < 0.05) in sham operated as well as pressure overloaded rats. The isomyosin V1 percentage was inversely related to the relative left ventricular weight. Two different (p < 0.05) correlation lines were detected for untreated and captopril treated rats. None of captopril associated effects were removed by simultaneously administered B, kinin receptor antagonist HOE 140. Thus, stimulation of bradykinin B2 receptor appears not to mediate the effects of captopril on cardiac growth and contractile proteins during the development of pressure overload hypertrophy. PMID- 11108155 TI - Role of cardiac renin-angiotensin system in sarcoplasmic reticulum function and gene expression in the ischemic-reperfused heart. AB - The aim of this study was to explore the possible participation of cardiac renin angiotensin system (RAS) in the ischemia-reperfusion induced changes in heart function as well as Ca2+-handling activities and gene expression of cardiac sarcoplasmic reticulum (SR) proteins. The isolated rat hearts, treated for 10 min without and with 30 microM captopril or 100 microM losartan, were subjected to 30 min ischemia followed by reperfusion for 60 min and processed for the measurement of SR function and gene expression. Attenuated recovery of the left ventricular developed pressure (LVDP) upon reperfusion of the ischemic heart was accompanied by a marked reduction in SR Ca2+-pump ATPase, Ca2+-uptake and Ca2+-release activities. Northern blot analysis revealed that mRNA levels for SR Ca2+-handling proteins such as Ca2+-pump ATPase (SERCA2a), ryanodine receptor, calsequestrin and phospholamban were decreased in the ischemia-reperfused heart as compared with the non-ischemic control. Treatment with captopril improved the recovery of LVDP as well as SR Ca2+-pump ATPase and Ca2+-uptake activities in the postischemic hearts but had no effect on changes in Ca2+-release activity due to ischemic-reperfusion. Losartan neither affected the changes in contractile function nor modified alterations in SR Ca2+-handling activities. The ischemia reperfusion induced decrease in mRNA levels for SR Ca2+-handling proteins were not affected by treatment with captopril or losartan. The results suggest that the improvement of cardiac function in the ischemic-reperfused heart by captopril is associated with the preservation of SR Ca2+-pump activities; however, it is unlikely that this action of captopril is mediated through the modification of cardiac RAS. Furthermore, cardiac RAS does not appear to contribute towards the ischemia-reperfusion induced changes in gene expression for SR Ca2+-handling proteins. PMID- 11108156 TI - Drug targeting by surface cationization. AB - Cationization of drug products and carriers involves a direct modification or attachment of conveying or accompanying components, either of which cause a charge modification. Cationization of macromolecules such as proteins and nucleotides and particulate drug carriers generally enhances their cellular uptake by endocytosis. The most common use of cationization today is in gene delivery. This is undertaken by either employing cationic polymers or entraping nucleotides in cationic carriers such as cationic liposomes. Cationized delivery systems are also used to overcome biological barriers and are suggested for drug targeting, in a nonspecific manner, to a variety of body organs, including brain, eyes, nose, and inflamed intestinal epithelium. Protein cationization is also suggested both for tumor immunotherapy and as a diagnostic tool in cancer therapy. Cationization has proven itself to be a straightforward tool for targeting to cells, tissues, and selected organs. This article reviews the extensive range of applications of cationization for improving drug and gene delivery and summarizes major technologies employed for that purpose. PMID- 11108157 TI - Regional variation in oral mucosal drug permeability. AB - The purpose of this article is to review regional variation in oral mucosal drug permeability. The structure and composition of the mucosa at different sites in the oral cavity, factors affecting mucosal permeability, selection of appropriate experimental systems for studying mucosal permeability, and formulation factors including penetration enhancement relevant to the design of systems for oral mucosal delivery are reviewed. PMID- 11108158 TI - Drug delivery by the intravaginal route. AB - The human vagina represents a potential, accessible space that offers a valuable route for drug delivery through the use of specifically designed carrier systems for both local and systemic applications. Intravaginal drug delivery is particularly appropriate for drugs associated with women's health issues but may also have applications in general drug delivery within the female population. A range of drug delivery platforms suitable for intravaginal administration are discussed in this review, including hydrogels, vaginal tablets, pessaries/suppositories, particulate systems, and intravaginal rings. Drug release mechanisms and absorption pathways are reviewed with respect to a range of therapeutic and prophylactic indications for intravaginal delivery. PMID- 11108159 TI - Evaluation of 62Cu labeled diacetyl-bis(N4-methylthiosemicarbazone) as a hypoxic tissue tracer in patients with lung cancer. AB - 62Cu labeled diacetyl-bis(N4-methylthiosemicarbazone) (62Cu-ATSM) has been proposed as a generator-produced, positron-emitting tracer for hypoxic tissue imaging. From basic studies, the retention mechanism of 62Cu-ATSM is considered to be closely related to cytosolic/microsomal bioreduction, a possible system for hypoxic bioreductive drug activation. In order to evaluate the characteristics of 62Cu-ATSM, PET studies were performed in 4 normal subjects and 6 patients with lung cancer. 62Cu-ATSM cleared rapidly from the blood with little lung uptake (0.43+/-0.09, uptake ratio; divided by the arterial input function) in normal subjects. Intense tumor uptake of 62Cu-ATSM was observed in all patients with lung cancer (3.00+/-1.50). A negative correlation was observed between blood flow and flow-normalized 62Cu-ATSM uptake in three of four patients. In contrast, 62Cu ATSM uptake was not related to that of 18F-fluorodeoxyglucose. The negative correlation between blood flow and flow normalized 62Cu-ATSM uptake suggests an enhancement of retention of 62Cu-ATSM by low flow. 62Cu-ATSM is a promising PET tracer for tumor imaging, which might bring new information for chemotherapeutic treatment as well as radiotherapy of hypoxic tumors. PMID- 11108160 TI - Estimation of 99mTc-MAG3 clearance by single-sample methods and camera-based methods. AB - We compared single-sample methods, proposed by Russell et al. and Bubeck et al., and camera-based methods in calculating 99mTc-MAG3 clearance, and determined camera-based methods that provide estimates comparable to those measured by the Russell method. Twenty-one patients underwent 99mTc-MAG3 renal scintigraphy, and clearance was measured by the Russell method and Bubeck method. Various renogram parameters were determined based on the slope of the renogram and area under the renogram, and correlated with the clearance measured by the Russell method. Camera-based clearance was calculated with the obtained regression equations and with equations determined previously using the Bubeck method as a standard. The Bubeck method provided lower measures than the Russell method in high renal function. Clearance measured by the Russell method was well correlated with renogram parameters, and clearance calculated with the obtained regression equation was comparable to that measured by the Russell method. When camera-based clearance was predicted with the previous equation, it was lower than the result obtained by the Russell method in high function. In conclusion, there are systematic differences in 99mTc-MAG3 clearance calculated by different methods. The camera-based methods obtained in this study appear to facilitate comparison of results obtained by the Russell method and camera-based method. PMID- 11108161 TI - Regional cardiac sympathetic reinnervation in transplanted human hearts detected by 123I-MIBG SPECT imaging. AB - The purpose of this study was to assess the regional cardiac sympathetic reinnervation late (> or = 1 year) after heart transplantation (HTX) by means of 123I-MIBG (MIBG) scintigraphy. Eight patients with a pretransplantation diagnosis of idiopathic dilated cardiomyopathy underwent MIBG scintigraphy more than one year after HTX. The presence or absence of regional MIBG uptake was evaluated in each SPECT image, and global MIBG uptake was semi-quantitatively assessed by the heart to mediastinum ratio (H/M). Five of 8 patients had visible MIBG uptake in both planar and SPECT images (PU group), whereas 3 of 8 patients had no uptake, 2 of them after a period of 2 years, and one of them as long as 5 years after HTX, respectively (NU group). Positive regional MIBG uptake involved the basal anterior region in all 5 patients, the basal septal region in 4 patients, the basal lateral region in 3 patients and the basal posterior region in 1 patient. The H/M value was 1.24+/-0.10 in the PU group and 1.09+/-0.03 in the NU group. In conclusion, MIBG SPECT can detect regional sympathetic reinnervation, indicating that basal septal and lateral regions next to the basal anterior are more likely to be reinnervated, but reinnervation is much less likely to occur in the midventricular and apical regions. PMID- 11108162 TI - 18F-FDG for the staging of patients with differentiated thyroid cancer: comparison of a dual-head coincidence gamma camera with dedicated PET. AB - Coincidence imaging with a dual-head gamma camera may offer a cost-effective alternative to dedicated PET. The aim of this study was to compare the diagnostic accuracy of coincidence imaging and PET in patients with differentiated thyroid cancer. Thirty-one patients were studied after thyroidectomy and radioiodine ablation. They were injected with a single dose of 300 MBq 18F-FDG. Scanning was performed on a dedicated PET system after 1 hr, and on a coincidence gamma camera after 4 hrs. Based on a lesion-by-lesion comparison, coincidence imaging and PET concurred in 69% of 118 lesions. Based on lesion size, concurrence was 96% in lesions larger than 1.5 cm, and 62% in those between 1 and 1.5 cm. Lesions smaller than 1 cm could not be identified with coincidence imaging. Identical staging was obtained with coincidence imaging and PET in 26/31 patients (84%). In four patients FDG accumulating lesions were shown by both the coincidence camera and the dedicated scanner, but not detectable with any other imaging means and were confirmed histologically on surgery. Although a coincidence camera is technically inferior to a dedicated PET scanner, it may provide clinically useful results in situations were a lesion of sufficient size and FDG uptake is to be expected, e.g. when evaluating a known lesion for malignancy. PMID- 11108163 TI - Estimation of the area at risk in myocardial infarction of rats by means of I-123 beta-methyliodophenyl pentadecanoic acid imaging. AB - Clinical investigations have suggested that the defects in SPECT images of a free fatty acid analog, I-123 beta-methyliodophenyl pentadecanoic acid (BMIPP) may indicate the ischemic risk area. To elucidate whether I-123 BMIPP can indicate the area at risk of ischemia, ex-vivo autoradiography was performed in rats whose left coronary artery was occluded for 60 min and then reperfused. I-123 BMIPP was injected at the acute stage (n = 10), or the subacute stage (7 days after reperfusion; n = 9). Infarction and the area at risk were identified by triphenyl tetrazolium chloride staining and injection of methylene blue during religation just before sacrifice, respectively. The BMIPP uptake in the risk area was significantly lower than that in the remote area at the acute (risk, 53.7+/-23.3% of the uptake at right ventricle, mean +/- SD; remote, 109.3+/-11.8%; p < 0.01) and subacute (risk, 52.5+/-11.5%; remote, 97.9+/-14.3%; p < 0.01) stages. In addition, the area with reduced uptake of I-123 BMIPP showed a significant correlation with the area at risk both at the acute (r = 0.98, p < 0.01) and subacute (r = 0.92, p < 0.01) stages. In conclusion, the area at risk can be evaluated by I-123 BMIPP both at the acute and subacute stages. PMID- 11108164 TI - Evaluation of coronary blood flow reserve by 13N-NH3 positron emission computed tomography (PET) with dipyridamole in the treatment of hypertension with the ACE inhibitor (Cilazapril). AB - PURPOSE: The purpose of this study was to evaluate the effect of treatment with an angiotensin-converting enzyme (ACE) inhibitor (Cilazapril) for early hypertensive patients in terms of coronary blood flow reserve evaluated by 13NH3 positron emission tomography (PET). METHODS: Before and after 12 weeks of ACE inhibitor treatment, 13NH3-PET with dipyridamole provocation test was performed, and definite myocardial perfusion and coronary flow reserve (CFR) were calculated. RESULTS: Compared to our normal subjects previously reported (2.61+/ 0.74), average coronary flow reserve was decreased (1.70+/-0.64 in hypertensive patients), and improved after treatment (1.77+/-0.52), but not significantly. Of 12 patients, five (42%) showed improved coronary flow reserve from 1.34 to 1.99 without a significant change in the resting flow. Only one patient (8%) showed deterioration after the ACE inhibitor treatment. The coronary vascular resistance (CVR) after ACE inhibitor treatment of the patients with CFR < 2.0 decreased significantly compared with those with CFR> or = 2.0 (p < 0.03). CONCLUSIONS: These results indicate that hypertensive patients at the early stage show decreased coronary flow reserve despite having normal resting flow. Treatment with an ACE inhibitor (Cilazapril) for 12 weeks improved coronary flow reserve in 42% of our patients. The CVR of the patients with CFR < 2.0 showed improvement compared to those with CFR> or = 2.0. This result indicates that an ACE inhibitor (e.g., Cilazapril) should be one of the choices for improving CFR if hypertensive patients in early stage show signs of ischemia or diastolic dysfunction, which may be one of the sequels of reserve restriction. PMID- 11108165 TI - Changes in perfusion and fatty acid metabolism of rat heart with autoimmune myocarditis. AB - To elucidate the change in perfusion and aerobic metabolism in myocarditis, tissue counting and dual tracer ex vivo autoradiography with Tl-201 and a free fatty acid analog, I-123- or I-125-labeled (p-iodophenyl)-methyl-pentadecanoic acid (BMIPP), were performed in rats with myocarditis induced by immunization with cardiac myosin. Inflammatory damage was classified histologically. At the acute stage (2-4 weeks after the antigen-injection), total heart uptakes of Tl and BMIPP and the ratio (BMIPP/Tl) were significantly reduced in myocarditis rats (N = 15) compared with the controls (N = 12). Myocardial distribution of Tl and BMIPP was not homogeneous. Relative uptake of Tl and BMIPP (N = 9, 128 regions) was gradually decreased with the extent of inflammation, and the regional BMIPP/Tl was smaller than the control. At the subacute stage (7 weeks after the antigen-injection), total Tl uptake in myocarditis rats (N = 5) recovered to the control level (N = 4), but that of BMIPP was still significantly lower than the control. BMIPP/Tl was still significantly lower in myocarditis. Myocardial distribution of Tl and BMIPP recovered to be more homogeneous. Relative uptake of Tl and BMIPP (N = 6, 78 regions) still gradually but significantly decreased with the extent of inflammation. Regional BMIPP/Tl was still depressed in myocarditis. These results indicate that myocardial perfusion and aerobic metabolism were discrepant and heterogeneously suppressed with severe inflammation during the acute stages, but the difference decreases with time. Examination with Tl-201 and BMIPP may provide information about the severity of myocarditis. PMID- 11108166 TI - Relationship between regional severity of emphysema and coronary heart disease. AB - We analyzed the relationship between regional severity of emphysema, which was evaluated by three-dimensional fractal analysis (3D-FA) of Technegas SPECT images, and coronary heart disease (CHD). For 22 patients with emphysema who underwent Technegas SPECT, we followed up CHD events. The follow-up period was 5.4+/-0.5 (mean +/- SD) years. We defined the upper-lung fractal dimension (U-FD) and lower-lung fractal dimension (L-FD) obtained with 3D-FA of Technegas SPECT images as the regional severity of emphysema. FD became greater with the progression of emphysematous change. During the follow-up period, CHD events occurred in 6 (27%) of the 22 patients. The ratio of U-FD to L-FD for patients with CHD events (0.87+/-0.22) was significantly smaller than for patients without CHD events (1.52+/-0.38) (p = 0.0015). These findings suggest that severer emphysema in the lower lung indicates a higher risk of CHD than that in the upper lung. PMID- 11108167 TI - A case of cavernous hemangioma of the small intestine diagnosed by scintigraphy with Tc-99m-labeled red blood cells. AB - Hemangioma of the small intestine is rare, and the preoperative diagnosis of it is difficult. We report a patient with gastrointestinal bleeding for whom Tc-99m labeled red blood cell scintigraphy was useful in diagnosing cavernous hemangioma of the small intestine. A 25-year-old man was referred to our hospital for recurrent iron deficiency anemia. Because of the patient's severe anemia, imaging was performed to locate the bleeding lesion in the gastrointestinal tract. Scintigraphy with Tc-99m-labeled red blood cells revealed pooling indicating a tumor and extravasation of blood from the tumor. Scintigraphy with Tc-99m pertechnetate revealed no abnormal accumulation. Partial resection of the small intestine was done, and cavernous hemangioma of the small intestine was diagnosed by using the specimen of resected tissue. PMID- 11108168 TI - Assessment of penile bone graft viability by bone scintigraphy: a case report. AB - A 20-year-old man, who had penile reconstruction surgery with an iliac bone graft a year ago due to malcircumcision at 6 years old underwent bone scintigraphy in order to detect bone graft viability. The accumulation of 99mTc-MDP in the penile region revealed the viability of the bone graft. This case report shows that bone scintigraphy can be used to assess the viability of a bone graft located inside the penis as well as bone grafts placed elsewhere in the extremities. PMID- 11108169 TI - Detecting meningeal carcinomatosis from breast cancer with thallium-201 SPECT. AB - Thallium-201 (201Tl) scintigraphy is one of the imaging methods used in the detection of various tumors including brain metastasis. We evaluated a patient with meningeal carcinomatosis from breast cancer by using 201Tl single-photon emission computed tomography (SPECT). Meningeal spread of a tumor was noted on enhanced CT. SPECT revealed tumor localization in meningeal carcinomatosis. These results suggest that SPECT with 201Tl may be useful in detecting meningeal carcinomatosis from breast cancer. PMID- 11108170 TI - A case of recurrent cholangitis after bile duct injury during laparoscopic cholecystectomy: value of scintigraphy with Tc-99m GSA and hepatobiliary scintigraphy for indication of lobectomy. AB - A 39-year-old woman with acute cholecystitis and gallstones underwent laparoscopic cholecystectomy. She suffered from recurrent episodes of cholangitis due to injury of the major bile ducts during laparoscopic cholecystectomy. Hepatobiliary scintigraphy with Tc-99m Sn-N-pyridoxyl-5-methyltryptophan was performed. Although normal bile excretion was found from the left hepatic duct to the percutaneous transhepatic biliary drainage (PTBD) tube, excretion from the right hepatic lobe was prolonged. Scintigraphy with Tc-99m diethylenetriaminepentaacetic acid-galactosyl human serum albumin demonstrated atrophy of the right hepatic lobe and enlargement of the left hepatic lobe. Cholangiography via the PTBD tube revealed complete obstruction of the left hepatico-jejunal anastomosis and could not enhance the right intrahepatic bile duct. A right hepatic lobectomy was performed because of the atrophy, glissonitis and the absence of an appropriate bile duct for reconstruction. Postoperatively she was active and exhibited no evidence of recurrent cholangitis. PMID- 11108172 TI - Gliosarcoma with thallium-201 SPECT. AB - Thallium-201 (201Tl) chloride scintigraphy is the imaging method use for the detection of various tumors including glioblastoma, but only limited information on 201Tl uptake in gliosarcoma is available. We investigated a patient with gliosarcoma by means of 201Tl single-photon emission computed tomography (SPECT) and MRI. SPECT imaging revealed high 201Tl uptake in the tumor, which was closely correlated with contrast-enhancement on MRI. These results suggest that SPECT with 201Tl may be useful for detecting gliosarcoma and provide physiological information on this tumor. PMID- 11108171 TI - A case of malignant lymphoma of the hand. AB - Malignant lymphoma is rarely seen in the hand. We present a case of malignant lymphoma of the hand with Ga-67 citrate and MR images, and discuss the usefulness of Ga-67 citrate scintigraphy for diagnosing this condition. PMID- 11108173 TI - Use of CA15-3, CEA and prolactin for the primary diagnosis of breast cancer and correlation with the prognostic factors at the time of initial diagnosis. AB - The main goals of the clinical use of tumor markers are to evaluate the adequacy of the treatment, monitor recurrence and follow up response to the treatment applied. For this purpose a baseline level for the commonly used tumor marker must be known at the time of initial diagnosis, before any therapy, in order to compare with the tumor marker levels which will be obtained after the treatment and during the clinical follow-up. The aim of this study was to investigate the correlation, if there is any, of the baseline levels of CA15-3, CEA and prolactin (PRL) in patients with breast cancer with the most commonly used prognostic factors, i) the presence of distant metastasis, ii) the presence of axillary lymphatic invasion, iii) the number of invaded axillary lymph nodes, iv) tumor size and v) stage of the disease, for breast cancer. Baseline serum CA15-3, CEA and PRL levels of 172 patients with breast masses were determined prior to biopsy. The sensitivity and specificity of baseline CA15-3, CEA and PRL were; 23.2% and 95.3%, 17.41% and 83.7%, 5.8% and 97.6%, respectively. At least one of the three tumor markers was high in 36% (31/86) of the breast cancer patients. Baseline CA15-3 levels were frequently higher than CEA in patients with bone metastasis (60% vs. 20%) and axillary lymphatic invasion (31.8% vs. 25%), and showed a better correlation with the stage of disease. Baseline tumor marker levels showed no statistically significant correlation with either the number of invaded axillary lymph nodes or tumor size. In conclusion, sensitivities and negative predictive values for baseline CA15-3, CEA and PRL were not satisfactory for primary diagnosis of breast cancer. Correlation of baseline CA15-3 was found superior to CEA and PRL in terms of stage of disease, presence of axillary invasion and distant metastasis. PMID- 11108174 TI - Outlining the body contours with scattered photons in lymphoscintigraphy for sentinel nodes. AB - Although lymphoscintigraphy is a useful method of detecting the sentinel nodes of malignancy, conventional lymphoscintigraphy images only the sentinel nodes without revealing their anatomical location. We, therefore, used scattered photons to attempt to outline the body contours of patients with either breast or esophageal cancer. Lymphoscintigraphy was performed 3 to 4 hours after the injection of 111 MBq of 99mTc tin colloid into the peritumoral region. Images were obtained with dual-energy windows of 130 to 150 keV for the primary photons and 70 to 110 keV for the scattered photons. The images constructed from the scattered photons clearly showed the contours of the body, and the fusion images constructed from the primary and scattered photons allowed for easy identification of the location of the sentinel nodes. The results of this study confirm that images obtained from scattered photons on lymphoscintigraphy are helpful in identifying the anatomical location of sentinel nodes. PMID- 11108175 TI - Reliability of two 2,400-m time-trial protocols for assessing performance of Standardbred racehorses. AB - OBJECTIVE: To evaluate the reliability of 2 time-trial protocols, 1 that involved use of a pacemaker and 1 that did not, for assessing performance of Standardbred racehorses. ANIMALS: 3- to 7-year-old Standardbred maiden pacers. PROCEDURE: 11 Standardbred pacers were used to determine the reliability of a time-trial protocol that involved use of a galloping pacemaker. All tests were performed by a single driver. Horses were paced for an initial 1,600 m then raced and timed for the final 800 m. They were retested 9 days later. Twenty-two horses were used to determine the reliability of a time-trial protocol that did not involve use of a pacemaker. All tests were performed by a second driver. Horses were paced for the first 1,200 m then raced and timed for the final 1,200 m. They were retested 3 and 8 days later. RESULTS: Reliability of performance time, expressed as the coefficient of variation (ie, the typical percentage variation in a horse's time between trials), was similar for the 2 protocols (1.0% and 1.3%, respectively). CONCLUSIONS: The small variations in performance times for the 2 time-trial protocols were similar to those of the best comparable laboratory and field tests of human and equine performance. Both protocols would be suitable for investigating factors affecting performance of Standardbred racehorses. PMID- 11108176 TI - Maturation of the auditory system in clinically normal puppies as reflected by the brain stem auditory-evoked potential wave V latency-intensity curve and rarefaction-condensation differential potentials. AB - OBJECTIVE: To evaluate auditory maturation in puppies. ANIMALS: Ten clinically normal Beagle puppies. PROCEDURE: Puppies were examined repeatedly from days 11 to 36 after birth (8 measurements). Click-evoked brain stem auditory-evoked potentials (BAEP) were obtained in response to rarefaction and condensation click stimuli from 90 dB normal hearing level to wave V threshold, using steps of 10 dB. Responses were added, providing an equivalent to alternate polarity clicks, and subtracted, providing the rarefaction-condensation differential potential (RCDP). Steps of 5 dB were used to determine thresholds of RCDP and wave V. Slope of the low-intensity segment of the wave V latency-intensity curve was calculated. The intensity range at which RCDP could not be recorded (ie, pre-RCDP range) was calculated by subtracting the threshold of wave V from threshold of RCDP RESULTS: Slope of the wave V latency-intensity curve low-intensity segment evolved with age, changing from (mean +/- SD) -90.8 +/- 41.6 to -27.8 +/- 4.1 micros/dB. Similar results were obtained from days 23 through 36. The pre-RCDP range diminished as puppies became older, decreasing from 40.0 +/- 7.5 to 20.5 +/ 6.4 dB. CONCLUSION AND CLINICAL RELEVANCE: Changes in slope of the latency intensity curve with age suggest enlargement of the audible range of frequencies toward high frequencies up to the third week after birth. Decrease in the pre RCDP range may indicate an increase of the audible range of frequencies toward low frequencies. Age-related reference values will assist clinicians in detecting hearing loss in puppies. PMID- 11108177 TI - Measurement of peptidase activity and evaluation of effectiveness of administration of minocycline for treatment of dogs with periodontitis. AB - OBJECTIVE: To determine clinical, enzymatic, and microbiologic effects of controlled-release localized administration of minocycline on dogs with periodontitis. ANIMALS: Five adult Beagles with periodontitis. PROCEDURE: After tooth scaling and root planing, 2 treatment, 1 placebo, and 1 control site were selected for each dog. Treatment sites (n = 10) received a periodontal formulation of minocycline hydrochloride, placebo sites (5) received ointment without minocycline, and control sites (5) did not receive ointment. Treatments were administered 4 times at weekly intervals. Peptidase activity and clinical and microbiologic effects were evaluated and compared among sites for 17 weeks. RESULTS: Bleeding of the gums on probing (BOP) and pocket depth (PD) improved at the treatment site and were maintained for 13 weeks after treatment. However, BOP and PD in placebo and control sites increased from weeks 9 to 17 Peptidase activity in the periodontal pocket decreased noticeably from week 1 to 17, compared with baseline values for the treatment site. However, peptidase activity for placebo and control sites increased and were above baseline values on week 9 and week 13, respectively. Total bacterial counts decreased by 90% for treatment sites and remained at that value for 13 weeks. However, for placebo and control sites, bacterial counts increased and reached the baseline value on week 17. CONCLUSIONS AND CLINICAL RELEVANCE: Increased peptidase activity is correlated with the progression of periodontitis in dogs. Treatment with minocycline, using a localized delivery system, was effective in dogs for at least 13 weeks after cessation of drug administration. PMID- 11108178 TI - Immediate urodynamic and anatomic response to colposuspension in female Beagles. AB - OBJECTIVE: To characterize urodynamic function and anatomy before and after colposuspension in anesthetized female Beagles. ANIMALS: 12 adult female Beagles. PROCEDURE; During general anesthesia (thiopental sodium induction and halothane maintenance), urethral pressure profiles, leak point pressure measurements with a 50-ml bladder volume, positive contrast cystograms, and retrograde vaginourethrocystograms were performed. A caudal midline laparotomy was used to perform colposuspension. Urodynamic and radiographic studies were repeated after surgery. RESULTS: Leak point pressures were increased (120 to 168.9 cm H2O), and maximum urethral closure pressures decreased (43.7 to 19.3 cm H2O ) after colposuspension. The urethra and bladder were moved cranially; the external urethral orifice was positioned closer to the pelvic cavity, and the neck of the bladder was positioned more cranially into the abdomen. Length of the urethra, as measured by use of vaginourethrocystograms, was increased by 3%. As measured by use of urethral pressure profiles, total profile length was increased by 19.9%, and functional profile length was increased by 19.2%. CONCLUSIONS AND CLINICAL RELEVANCE: Increased leak-point pressure correlated with the expected clinical improvement attributable to colposuspension. Increased exposure of the urethra to abdominal and pelvic cavity pressures may be the mechanism by which incontinent dogs become continent after colposuspension. Results of the leak-point pressure test may correlate with clinical behavior before and after colposuspension for treatment of incontinence. PMID- 11108180 TI - Hemodynamic and electrophysiologic effects of ontazolast in dogs. AB - OBJECTIVE: To determine whether QT interval is prolonged or sudden death is caused by ventricular fibrillation resulting from torsades de pointes and to identify hemodynamic effects of ontazolast. ANIMALS: 28 Beagles. PROCEDURE: Physiologic variables were measured for 2 hours in conscious dogs given ontazolast (0, 1, or 3 mg/kg of body weight, IV) and for 1 hour in anesthetized dogs given cumulative doses of ontazolast (0, 1, 3, 6, or 8 mg/kg, IV). RESULTS: Ontazolast prolonged QT interval and QT interval corrected for heart rate (QTc) at doses of 6 mg/kg in anesthetized dogs. At 8 mg/kg, both variables remained prolonged but tended to decrease. In conscious dogs, ontazolast increased QT interval and QTc 15 minutes after administration, but both variables returned to reference ranges by 60 minutes. In conscious dogs, ontazolast increased maximum rate of increase of left ventricular pressure and maximal velocity of fiber shortening, indicators of inotropy, and increased tau, indicating a decreased rate of relaxation. One conscious dog receiving 3 mg/kg developed nonfatal torsades de pointes, but another conscious dog developed ventricular fibrillation. Two anesthetized dogs receiving 6 mg/kg developed early afterdepolarizations, and all dogs developed secondary components in theirT waves. CONCLUSION AND CLINICAL RELEVANCE: Ontazolast possesses potent class-III antiarrhythmic properties and induces prolongation of QTc in a dose-dependent fashion. Because there was a clear dose-dependent prolongation of QT interval in all instances, ontazolast may serve as a positive-control compound for studying other compounds that are believed to prolong the QT interval. PMID- 11108179 TI - Genetic analysis of vesicular stomatitis virus-New Jersey from the 1995 outbreak in the western United States. AB - OBJECTIVE: To compare molecular associations between the vesicular stomatitis virus (VSV)-New Jersey isolates of the 1995 outbreak with those from previous outbreaks between 1982 and 1985 in the western United States. SAMPLE POPULATION: 23 virus isolates considered representative of the 1995 outbreak of vesicular stomatitis. PROCEDURE: Viral gene coding for surface-envelope protein G was evaluated by use of nucleotide sequencing and phylogenetic analysis. RESULTS: Changes in up to 0.77% of the nucleotide bases and 1.35% of the amino acids were detected among the 1995 viral isolates, whereas changes in up to 3.2 and 2.9% of the nucleotides and amino acids, respectively, were found, compared with the 1982 to 1985 viruses. Insertions or deletions were not found in the entire gene, which spanned 1,554 nucleotide bases. CONCLUSIONS AND CLINICAL RELEVANCE: Phylogenetic analysis indicated that the 1995 VSV-New Jersey belongs to a lineage distinct from that of the 1982 to 1985 viruses that caused previous outbreaks in the western United States. Furthermore, it also is distinct from strains from Central America and from the Georgian Hazelhurst strain. PMID- 11108181 TI - Interleukin-8 concentration and neutrophil chemotactic activity in bronchoalveolar lavage fluid of horses with chronic obstructive pulmonary disease following exposure to hay. AB - OBJECTIVE: To analyze effects of hay dust exposure on interleukin-8 (IL-8) concentration, percentage of neutrophils, and neutrophil chemotactic activity in bronchoalveolar lavage fluid (BALF) of horses with chronic obstructive pulmonary disease (COPD). ANIMALS: 16 healthy horses and 29 horses with COPD. PROCEDURE: IL 8 concentration, percentage of neutrophils, and neutrophil chemotactic activity in BALF were measured. Values were analyzed with respect to hay dust exposure. These variables were also measured in 5 asymptomatic horses with COPD after the induction of clinical signs by changing feed from silage to hay. RESULTS: L-8 concentrations and chemotactic activity in BALF were greater in horses with COPD, compared with healthy horses, and greater in horses with COPD exposed to hay dust, compared with nonexposed affected horses. An increase in IL-8 concentration accompanied by an increase in percentage of neutrophils in BALF and development of clinical signs of COPD were induced in asymptomatic horses with COPD by changing feed from silage to hay. CONCLUSIONS AND CLINICAL RELEVANCE: Exposure of horses with COPD to hay dust components resulted in an increase in IL-8 secretion at the bronchoalveolar surface. This chemokine may play a role in the pathogenesis of COPD, because it causes neutrophil accumulation in the bronchoalveolar space. Our results underscore the importance of eliminating dust sources for the treatment and prevention of COPD in horses. PMID- 11108182 TI - Results of a longitudinal study of the prevalence of Escherichia coli O157:H7 on cow-calf farms. AB - OBJECTIVE: To describe the frequency and distribution of Escherichia coli O157:H7 in the feces and environment of cow-calf herds housed on pasture. SAMPLE POPULATION: Fecal and water samples for 10 cow-calf farms in Kansas. PROCEDURE: Fecal and water samples were obtained monthly throughout a 1-year period (3,152 fecal samples from 2,058 cattle; 199 water samples). Escherichia coli O157:H7 in fecal and water samples was determined, using microbial culture. RESULTS: Escherichia coli O157:H7 was detected in 40 of 3,152 (1.3%) fecal samples, and 40 of 2,058 (1.9%) cattle had > or = 1 sample with E coli. Fecal shedding by specific cattle was transient; none of the cattle had E coli in more than 1 sample. Significant differences were not detected in overall prevalence among farms. However, significant differences were detected in prevalence among sample collection dates. Escherichia coli O157:H7 was detected in 3 of 199 (1.5%) water samples. CONCLUSIONS AND CLINICAL RELEVANCE: Implementing control strategies for E coli O157:H7 at all levels of the cattle industry will decrease the risk of this organism entering the human food chain. Devising effective on-farm strategies to control E coli O157:H7 in cow-calf herds will require an understanding of the epidemiologic characteristics of this pathogen. PMID- 11108183 TI - Tyrosine hydroxylase- and neuropeptides-immunoreactive nerves in canine trachea. AB - OBJECTIVE: To determine distribution of catecholaminergic and peptidergic nerve fibers in canine tracheas by use of immunohistochemistry. SAMPLE POPULATION: 10 tracheas collected from healthy adult dogs after euthanasia. PROCEDURE: Structure of the nerve network and distribution of tyrosine hydroxylase (TH)- and 6 types of neuropeptide-containing nerves in canine tracheas were immunohistochemically studied, using neurochemical markers. RESULTS: Intraepithelial free nerve endings with immunoreactivity for calcitonin gene-related peptide (CGRP) and substance P (SP) were observed. Tyrosine hydroxylase-, SP-, vasoactive intestinal peptide (VIP)-, and galanin (GAL)-immunoreactive nerve fibers were observed within and around the submucosal seromucous gland. In the smooth muscle layer, numerous TH- and GAL-immunoreactive nerve fibers, a moderate number of VIP- and neuropeptide Y (NPY)-immunoreactive nerve fibers, and a few SP- and methionine enkephalin (ENK) immunoreactive nerve fibers were observed. Numerous nerve cell bodies with VIP and GAL immunoreactivity and a few with SP ENK, and NPY immunoreactivity were observed. Many TH-immunoreactive fibers were arranged in a meshwork around blood vessels. Nerves with CGRP-, SP-, VIP-, GAL-, ENK-, and NPY-immunoreactivity were also observed around blood vessels. CONCLUSIONS: Complex innervation, including catecholamine- and neuropeptide-containing nerves, which may be related to regulation of muscle contraction and glandular secretion, are found in canine tracheas. PMID- 11108184 TI - Evaluation of local and systemic immune responses induced by intramuscular injection of a Mycoplasma hyopneumoniae bacterin to pigs. AB - OBJECTIVE: To evaluate immune responses induced by administration of Mycoplasma hyopneumoniae bacterin to pigs. Animals-60 healthy 7- to 10-day-old cross-bred boars. PROCEDURE: Pigs were assigned to 1 of 4 pig groups (15 pigs/group): vaccinated, challenged; vaccinated, nonchallenged; nonvaccinated, challenged; nonvaccinated, nonchallenged. Vaccinated pigs received IM injections of a mycoplasma bacterin on days 0 and 14, whereas nonvaccinated pigs received saline (0.9% NaCl) solution. Pigs in the challenged groups were inoculated intratracheally with M hyopneumoniae on day 42. Pigs were euthanatized and necropsied 41, 44, 48, and 70 days after the first vaccination, and proportion of lung surface with pneumonic lesions was determined. Percentage of lymphocyte subpopulations and number of interferon-gamma (IFN-gamma) secreting lymphocytes in blood and tissues, cytokine and antibody concentrations in bronchoalveolar lavage (BAL) fluid, and serum antibody concentrations were determined. RESULTS: Vaccination against and infection with M hyopneumoniae induced a local mucosal immune response in the respiratory tract of pigs. Proportion of lung surface with pneumonic lesions in vaccinated challenged pigs was reduced on day 70, compared with nonvaccinated challenged pigs. Vaccination stimulated the production of M hyopneumoniae-specific IFN-gamma secreting blood lymphocytes. Tumor necrosis factor-alpha concentration in BAL fluid on day 70 was increased in nonvaccinated challenged pigs, compared with vaccinated challenged pigs. CONCLUSIONS AND CLINICAL RELEVANCE: Vaccination against M hyopneumoniae induced local, mucosal, humoral, and cellular immune responses. Moreover, vaccination reduced the severity of lung lesions in challenged pigs, suggesting that mucosal antibodies, mediation of the inflammatory response, and cell-mediated immune responses are important for control of mycoplasmal pneumonia in pigs. PMID- 11108185 TI - Effect of ration and exercise on plasma creatine kinase activity and lactate concentration in Thoroughbred horses with recurrent exertional rhabdomyolysis. AB - OBJECTIVE: To determine the effects of 3 rations (low grain, fat, high grain) on plasma creatine kinase (CK) activity and lactate concentration in Thoroughbred horses with recurrent exertional rhabdomyolysis (RER). ANIMALS: 5 Thoroughbreds with RER and 3 healthy Thoroughbreds (control horses). PROCEDURES: Rations were formulated to meet (low-grain and fat rations) or exceed (high-grain ration) daily energy requirements. Each ration was fed to horses in a crossover design for 3 weeks. Horses were exercised on a treadmill Monday through Friday; maximum speed on Monday and Friday was 11 m/s (6% slope), on Tuesday and Thursday was 9 m/s, and on Wednesday was 4.5 m/s. Plasma CK activity and lactate concentration were determined before and after exercise. RESULTS: Horses with RER fed the high grain ration had significantly greater CK activity and change in CK activity 4 hours after exercise, compared with those fed the low-grain ration. Horses with RER exercised at the trot or canter had significantly greater increases in CK activity, compared with those exercised at the gallop. Plasma lactate concentrations after exercise were similar in control and affected horses. Lactate concentration and CK activity were not correlated in horses with RER. CONCLUSIONS AND CLINICAL RELEVANCE: Rations high in grain and formulated to exceed daily energy requirements may increase episodes of rhabdomyolysis in thoroughbred horses susceptible to RER. PMID- 11108186 TI - Evaluation of toxicokinetic variables and arthropathic changes in juvenile rabbits after oral administration of an investigational fluoroquinolone, PD 117596. AB - OBJECTIVE: To compare toxicokinetic variables and associated tissue drug concentrations with severity of articular lesions in weight-bearing joints of juvenile rabbits after oral administration of a fluoroquinolone. ANIMAL: Ten 6- to 7-week-old, 800- to 1,200-g, New Zealand White rabbits. PROCEDURES: Rabbits were gavaged daily with the fluoroquinolone PD 117596 at 500 mg/kg of body weight for 5 days. Blood samples were collected on day 4 at preestablished times, up to 24 hours after drug administration. On day 5 gross lesion severity and prevalence were evaluated in the major weight-bearing joints, and tissue specimens were collected (60 minutes after drug administration). Serum and tissue drug concentrations were determined by microbiologic plate assay. RESULTS: Macroscopically, treatment rabbits had a high prevalence of arthropathy with the distal portion of the femur having the highest prevalence and severity of lesions. Grossly, alterations to articular cartilage included 1 to 4 mm in diameter vesicles or erosions. Histologically, vesicles were identified in the midzone or close to the zone of calcified cartilage of treatment rabbits. Chondrocyte cellularity was reduced in affected areas, and perivesicular regions had reduced staining with Safranin O. Correlation analysis of area under the curve values with total scores for lesion severity had a significant positive relationship. CONCLUSIONS: Our findings support the use of juvenile rabbits as a model for arthropathic changes induced by fluoroquinolone administration. PMID- 11108187 TI - Conglutinin and immunoconglutinin titers in stressed calves in a feedlot. AB - OBJECTIVE: To determine whether increased conglutinin titers are evident in stressed calves that do not develop respiratory tract disease in feedlots, compared with respiratory tract disease, and to determine the increase in immunoconglutinin titers. ANIMALS: 101 mixed-breed beef calves. PROCEDURE: Calves were processed at 4 farms of origin and allowed to remain with their dams for another 100 days. Calves from each farm were brought to a centrally located order buyer barn. In a feedlot, 101 calves were assigned to pens and observed daily for clinical signs of acute respiratory tract disease. When sick calves were detected, they were treated with antibiotics and isolated in a pen for 4 days. Conglutinin and immunoconglutinin titers were determined for all calves. RESULTS: During the 28-day study, 73 calves developed respiratory tract disease, whereas 28 calves remained healthy. Mean conglutinin titers differed significantly among calves from the 4 farms. Significant differences were not detected in conglutinin titers among calves on the basis of sex, morbidity, or vaccination status against Mannheimia haemolytica at each farm, the order-buyer barn, or the feedlot on days 8, 15, and 28 after arrival. Immunoconglutinin titers in calves differed significantly among farms and morbidity status. CONCLUSIONS AND CLINICAL RELEVANCE: Mean conglutinin titers in calves do not appear to be associated with the incidence of acute respiratory tract disease; however, increased immunoconglutinin titers appear to be associated with recovery of stressed calves from respiratory tract disease during the first 15 days after arrival in a feedlot. PMID- 11108188 TI - Comparison of the effects of adrafinil, propentofylline, and nicergoline on behavior in aged dogs. AB - OBJECTIVE: To compare the efficacy of adrafinil, propentofylline, and nicergoline for enhancing behavior of aged dogs. ANIMALS: 36 Beagles between 9 and 16 years old. PROCEDURE: Dogs were randomly assigned to receive adrafinil (20 mg/kg of body weight, PO, q 24 h; n = 12), propentofylline (5 mg/kg, PO, q 12 h; 12), or nicergoline (0.5 mg/kg, PO, q 24 h; 12) for 33 days. Baseline behaviors in an open field and in kennels (home cage) were recorded before treatment. After treatment, behaviors in the open field were recorded 2 hours after drug administration on days 2, 15, and 28, and 10 hours after administration on days 7, 20, and 33. Behaviors in the home cage were recorded 2 and 7 hours after drug administration on days 4, 17, and 30. RESULTS: Treatment with adrafinil resulted in a significant increase in locomotion in each of the open-field tests and an increase in locomotion in the home cage. This latter increase was smaller and more variable than that in the open field. Locomotion was not affected by treatment with propentofylline or nicergoline. In the open field, sniffing decreased over time in all 3 groups, but the largest decline was observed in the propentofylline group. CONCLUSIONS AND CLINICAL RELEVANCE: Treatment with adrafinil may improve the quality of life of aged dogs by increasing exploratory behavior and alertness. PMID- 11108189 TI - Intracellular magnesium concentrations in dogs with gastric dilatation-volvulus. AB - OBJECTIVE: To quantify and compare intracellular magnesium concentrations (Mgi) in clinically normal dogs (control dogs) and dogs that have gastric dilatation volvulus (GDV dogs) and to determine whether there is a difference in Mgi and serum magnesium concentrations (Mgs) between GDV dogs with and without cardiac arrhythmias. ANIMALS: 41 control dogs and 21 GDV dogs. PROCEDURE: Rectus abdominis muscle specimens were obtained from control and GDV dogs for determination of Mgi. Blood samples were obtained from GDV dogs for determination of Mgs, and dogs were monitored for 48 hours for cardiac arrhythmias. Muscle specimens were frozen at -40 C, oven dried at 95 C, and digested with concentrated nitric acid. Multielemental analyses were performed by simultaneous/sequential inductively coupled plasma-atomic emission spectroscopy with fixed-cross flow nebulization. The Mg, was standardized to sulfur content to correct for the amount of fat and fascia in the muscle specimen. Mean (+/- SEM) values were recorded in parts per million (ppm). Results-There were no significant differences in Mgi between control (627 +/- 11.1 ppm) and GDV (597 +/ 20.5 ppm) dogs, in Mgi between GDV dogs with (590 +/- 34 ppm) and without (584 +/- 29 ppm) cardiac arrhythmias, and in Mgs between GDV dogs with (1.77 +/- 0.26 ppm) and without (1.51 +/- 0.09 ppm) cardiac arrhythmias. There was no correlation between Mgs and Mgi (R2 = 0.0001). CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that Mg depletion is not pathophysiologically important in dogs with GDV and does not play a role in the cardiac arrhythmias detected in these patients. PMID- 11108190 TI - Effect of sodium bisulfate on skin and hooves of horses. AB - OBJECTIVE: To evaluate the safety of sodium bisulfate for use in horse barn environments by determining its irritant effect on skin and hooves. ANIMALS: 6 female mixed-breed ponies. PROCEDURE: Sodium bisulfate was applied to clipped intact skin of 6 ponies to evaluate its irritant effect after single (48 hours) and repetitive (6 h/d for 10 days) applications; similar areas of skin were used as untreated control sites. In addition, sodium bisulfate was applied to the sole of both front hooves of each pony and covered with wet gauze, and the entire hoof was covered with adhesive tape for 48 hours. RESULTS: Contact with moistened sodium bisulfate for 48 hours had no effect on pony skin. Contact with sodium bisulfate for 6 hours on 10 consecutive days did not cause gross changes but did cause mild to moderate microscopic changes including epidermal necrosis, hyperkeratosis, capillary congestion, edema, and diffuse mixed inflammatory cell infiltrate. All changes were limited to the epidermis and superficial dermis. Gross changes in hoof sole, signs of lameness, and increase in digital pulse pressure or pulse intensity were not detected. CONCLUSIONS AND CLINICAL RELEVANCE: Duration of contact with sodium bisulfate in this study was in excess of that expected under typical husbandry conditions. Despite this fact, gross changes in skin and hooves were not detected. Microscopic lesions were confined to the epidermis and superficial dermis. Results suggest that contact with sodium bisulfate under these conditions is safe. PMID- 11108191 TI - Evaluation of results for Schirmer tear tests conducted with and without application of a topical anesthetic in clinically normal dogs of 5 breeds. AB - OBJECTIVE: To evaluate, for clinically normal dogs, results of Schirmer tear tests in eyes without topical anesthetic (STT) and to detect differences associated with breed, sex, age, day, and time of day in eyes in which STT was performed after use of topical anesthetic (STTa). ANIMALS: 41 Beagles, 43 Labrador Retrievers, 25 Golden Retrievers, 26 English Springer Spaniels, and 22 Shetland Sheepdogs. PROCEDURE: Beagles had STT and STTa values measured twice daily for 5 days. Client-owned dogs of 4 other breeds had STT and STTa values measured once. RESULTS: Mean +/- SD values of Beagles for STT and STTa were 20.2 +/- 2.5 and 3.8 +/- 2.7 mm/min. Mean values for STT and STTa were as follows: Labrador Retriever, 22.9 +/- 4.1 and 9.6 +/- 3.8 mm/min; English Springer Spaniel; 20.7 +/- 3.2 and 5.4 +/- 3.4 mm/min; Golden Retriever, 21.8 + 3.7 and 8.8 +/- 3.1 mm/min; and Shetland Sheepdog, 15.8 +/- 1.8 and 3.6 +/- 2.8 mm/min. Overall mean values for STT and STTa were 20.2 +/- 3.0 and 6.2 +/- 3.1 mm/min. Differences for STT and STTa were detected among breeds, but significant differences were not associated with sex or age within each breed or in overall values for all dogs. CONCLUSIONS AND CLINICAL RELEVANCE: Results for the STT reported here compare favorably with reported values, except for results of Shetland Sheepdogs; however, results for the STTa differ dramatically from reported values. Clinicians should consider effects attributable to breed when evaluating results of STT and STTa in dogs. PMID- 11108192 TI - Improved culture methods for isolation of Salmonella organisms from swine feces. AB - OBJECTIVE: To compare 3 alternative culture techniques for the detection of Salmonella organisms in swine feces with a modification of the International Standard Organization (ISO) 6579 standard protocol. SAMPLE POPULATION: Fecal samples from swine herds suspected of having Salmonella infections. PROCEDURE: 4 experiments were performed to evaluate the following: 1) diagnostic sensitivity of the selective preenrichment and rapid isolation novel technology (SPRINT) protocol, compared with that of the modified ISO protocol; 2) detection limit of the SPRINT protocol for Salmonella organisms; 3) use of tetrathionate-novobiocin (TTN) broth, compared with selenite cysteine (SC) broth for selective enrichment; and 4) use of universal preenrichment (UPE) broth, compared with buffered peptone water (BPW) for preenrichment of samples prior to the use of modified semisolid Rappaport-Vassiliadis (MSRV) plates. RESULTS: Comparing the Salmonella culture results of 183 swine fecal samples, the diagnostic sensitivity of the SPRINT protocol (0.86) was not significantly different than the diagnostic sensitivity of the modified ISO protocol (0.80), although it was 24 hours faster. The SPRINT protocol could detect 5 of the 6 investigated Salmonella serotypes at inoculation concentrations of < 10 colony-forming units (CFU)/25 g of uncontaminated feces. The TTN broth performed significantly better than the SC broth for selective enrichment of Salmonella organisms. There was no significant difference in results of preenrichment of samples between the use of UPE broth or BPW. CONCLUSIONS AND CLINICAL RELEVANCE: The SPRINT protocol may provide a faster alternative for isolation of Salmonella organisms from swine fecal samples. Furthermore, the use of TTN broth instead of SC broth may increase the sensitivity of the modified ISO 6579 protocol. PMID- 11108193 TI - Use of sevoflurane for anesthetic management of horses during thoracotomy. AB - OBJECTIVE: To evaluate sevoflurane as an inhalation anesthetic for thoracotomy in horses. ANIMALS: 18 horses between 2 and 15 years old. PROCEDURE: 4 horses were used to develop surgical techniques and were euthanatized at the end of the procedure. The remaining 14 horses were selected, because they had an episode of bleeding from their lungs during strenuous exercise. General anesthesia was induced with xylazine (1.0 mg/kg of body weight, IV) followed by ketamine (2.0 mg/kg, IV). Anesthesia was maintained with sevoflurane in oxygen delivered via a circle anesthetic breathing circuit. Ventilation was controlled to maintain PaCO2 at approximately 45 mm Hg. Neuromuscular blocking drugs (succinylcholine or atracurium) were administered to eliminate spontaneous breathing efforts and to facilitate surgery. Cardiovascular performance was monitored and supported as indicated. RESULTS: 2 of the 14 horses not euthanatized died as a result of ventricular fibrillation. Mean (+/- SD) duration of anesthesia was 304.9 +/- 64.1 minutes for horses that survived and 216.7 +/- 85.5 minutes for horses that were euthanatized or died. Our subjective opinion was that sevoflurane afforded good control of anesthetic depth during induction, maintenance, and recovery. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of sevoflurane together with neuromuscular blocking drugs provides stable and easily controllable anesthetic management of horses for elective thoracotomy and cardiac manipulation. PMID- 11108194 TI - Effect of dietary supplements containing antioxidants on attenuation of muscle damage in exercising sled dogs. AB - OBJECTIVE: To determine whether dietary antioxidants would attenuate exercise induced increases in plasma creatine kinase (CK) activity in sled dogs. ANIMALS: 41 trained adult sled dogs. PROCEDURE: Dogs, randomly assigned to 2 groups, received the same base diet throughout the study. After 8 weeks on that diet, 1 group (21 dogs) received a daily supplement containing vitamins E (457 U) and C (706 mg) and beta-carotene (5.1 mg), and a control group (20 dogs) received a supplement containing minimal amounts of antioxidants. After 3 weeks, both groups performed identical endurance exercise on each of 3 days. Blood samples were collected before and 3 weeks after addition of supplements and after each day of exercise. Plasma was analyzed for vitamins E and C, retinol, uric acid, triglyceride, and cholesterol concentrations, total antioxidant status (TAS), and CK activity. RESULTS: Feeding supplements containing antioxidants caused a significant increase in vitamin E concentration but did not change retinol or vitamin C concentrations orTAS. Exercise caused significantly higher CK activity, but did not cause a significant difference in CK activity between groups. Exercise was associated with significantly lower vitamin E, retinol, and cholesterol concentrations and TAS but significantly higher vitamin C, triglyceride, and uric acid concentrations in both groups. CONCLUSIONS AND CLINICAL RELEVANCE: Use of supplements containing the doses of antioxidants used here failed to attenuate exercise-induced increases in CK activity. Muscle damage in sled dogs, as measured by plasma CK activity, may be caused by a mechanism other than oxidant stress. PMID- 11108195 TI - Hematologic changes associated with the appearance of eccentrocytes after intragastric administration of garlic extract to dogs. AB - OBJECTIVE: To determine whether dogs given garlic extract developed hemolytic anemia and to establish the hematologic characteristics induced experimentally by intragastric administration of garlic extract. ANIMALS: 8 healthy adult mixed breed dogs. PROCEDURE: 4 dogs were given 1.25 ml of garlic extract/kg of body weight (5 g of whole garlic/kg) intragastrically once a day for 7 days. The remaining 4 control dogs received water instead of garlic extract. Complete blood counts were performed, and methemoglobin and erythrocyte-reduced glutathione concentrations, percentage of erythrocytes with Heinz bodies, and percentage of eccentrocytes were determined before and for 30 days after administration of the first dose of garlic extract. Ultrastructural analysis of eccentrocytes was performed. RESULTS: Compared with initial values, erythrocyte count, Hct, and hemoglobin concentration decreased to a minimum value on days 9 to 11 in dogs given garlic extract. Heinz body formation, an increase in erythrocyte-reduced glutathione concentration, and eccentrocytes were also detected in these dogs. However, no dog developed hemolytic anemia. CONCLUSIONS AND CLINICAL RELEVANCE: The constituents of garlic have the potential to oxidize erythrocyte membranes and hemoglobin, inducing hemolysis associated with the appearance of eccentrocytes in dogs. Thus, foods containing garlic should not be fed to dogs. Eccentrocytosis appears to be a major diagnostic feature of garlic-induced hemolysis in dogs. PMID- 11108196 TI - Methicillin-resistant coagulase-negative staphylococci isolated from healthy horses in Japan. AB - OBJECTIVE: To determine patterns of methicillin-resistant staphylococci isolated from apparently healthy horses. SAMPLE POPULATION: 44 horses from 8 riding clubs in Japan. PROCEDURE: Methicill in-resistant staphylococci were isolated from the skin or nares, using a selective medium containing a beta-(symboric) lactam antibiotic, ceftizoxime. Clonality of isolates was determined by use of pulsed field gel electrophoresis. Detection of mecA, mecl, and mecR1 genes was accomplished by use of polymerase chain reactions. RESULT: Of the 44 horses, 13 (29.5%) yielded 15 isolates of methicillin-resistant staphylococci. The 15 isolates were identified as 6 species (Staphylococcus epidermidis, S lentus, S saprophyticus, S xylosus, S sciuri, and S haemolyticus). However, methicillin resistant S aureus was seldom isolated. Each isolate contained the mecA gene and had a high resistance to beta-lactam antibiotics. Some isolates also were resistant to other antibiotics such as erythromycin and kanamycin. CONCLUSIONS AND CLINICAL RELEVANCE: Methicillin-resistant coagulase-negative staphylococci that were highly resistant to various antibiotics were isolated from apparently healthy horses in Japan. These organisms must be considered a potential threat to horses and veterinarians who care for them. PMID- 11108197 TI - Prevalence and relevance of antibodies to type-I and -II collagen in synovial fluid of dogs with cranial cruciate ligament damage. AB - OBJECTIVE: To measure and compare synovial fluid antibody titers to type-I and II collagen in stifle joints with instability caused by complete or partial cranial cruciate ligament (CCL) rupture and joints with osteoarthrosis secondary to other pathologic changes in dogs. ANIMALS: 82 dogs with diseased stifle joints. PROCEDURE: Synovial fluid samples were collected from 7 dogs with clinically normal stifles (control group) and 82 dogs with diseased joints (50 stifle joints with complete rupture of the CCL, 20 with partial damage of the CCL, and 12 joints with radiographic signs of osteoarthritis secondary to other arthropathies). Synovial fluid samples were tested for autoantibodies to type-I and -II collagen by an ELISA. RESULTS: In dogs with complete and partial CCL rupture, synovial fluid antibody titers to type-I and -II collagen were significantly increased, compared with control dogs. Forty-eight percent (24/50) of samples from dogs with complete CCL rupture and 35% (7/20) of samples from dogs with partial CCL rupture had antibody titers to type-I collagen that were greater than the mean plus 2 standard deviations of the control group titers. Synovial fluid antibody titers to type-II collagen were high in 40% of the dogs with partial or (8/20) complete (20/50) CCL rupture. Dogs with osteoarthrosis secondary to other pathologic changes had significantly increased synovial fluid antibodies to type-I and -II collagen, compared with control dogs. CONCLUSION: Increases in autoantibodies to collagen in synovial fluid are not specific for the type of joint disorder. It is unlikely that the anticollagen antibodies play an active role in the initiation of weakening of the CCL. PMID- 11108198 TI - Effects of computerized axial tomography: technical factors and scanning techniques on the accuracy of anatomical biomodels. PMID- 11108199 TI - Color image segmentation based on fuzzy rule-based reasoning applied to colonoscopic images. AB - A fuzzy color segmentation approach is developed for the analysis of colonoscopic images. The segmentation is made up of two phases: segmentation through histogram space filtering and region merging using fuzzy rule-base reasoning. The first phase involves using a scale-space filter to analyze the hue, saturation, and intensity (HSI) histograms to determine the number of classes and construct a 3-D class grid. The color image is then segmented based on the class grid. In the second phase, region merging based on applying the fuzzy rule-base is employed to guide the combining process of the segmented regions. For fuzzy reasoning, three criteria are evaluated, namely, the edge strength along the boundary, color similarity, and spatial connectivity of adjoining regions. Experimental testing of the proposed method applied on colonoscopic images was conducted, and the results are encouraging. PMID- 11108200 TI - Probabilitic fusion of hemodynamic parameter maps. AB - Hemodynamic parameter maps (HPMs) depict values of parameters of hemodynamic response, namely, gain, lag, and dispersion, in response to neuronal activation. Large intensity values of any of the HPMs indicate greater likelihoods of brain activation. We apply the probabilistic data fusion equation to combine three HPMs. The statistical map of the fused probability map derived from HPMs is compared with statistical parameter mapping (SPM) in a visual experiment. The statistical hemodynamic parameter map (SHPM) gives better contrast to distinguish activated voxels from nonactivated voxels. PMID- 11108201 TI - Combined optical and fluorescence imaging for breast cancer detection and diagnosis. AB - In this article, we present a novel approach that combines the best aspects of both endogenous optical imaging and exogenous fluorescent lifetime and yield imaging for breast cancer detection and diagnosis. Using this approach, spatial distributions of optical properties, fluorescence lifetime, and yield in tissue can be reconstructed from measured excitation and emission data at the surface of the breast tissue by regularized inverse algorithms. We show images from simulated data, as well as images from experimental data using tissue-like phantom materials in the laboratory. PMID- 11108202 TI - Bone formation into surface phosphonylated polymeric implants. AB - Through the use of two animal models, the present study demonstrates the ability of phosphonylated surfaces to bind bone. In one model, surface-treated polypropylene (PP) and polyethylene (PE) were implanted in the medial cortex of the goat tibia. In the second model, surface-treated poly(ether-ether ketone) (PEEK) and carbon fiber-reinforced PEEK (CFR-PEEK) were implanted through both cortices of the goat mandible. Selected rods of all material types were microtextured using crystallization induced microphase separation, a method for the formation of continuous, open-cell microporous surfaces in thermoplastic polymers. Microtextured and smooth rods were phosphonylated, and calcium was subsequently introduced to the phosphonylated surface by incubating the samples in a saturated solution of calcium oxide. For all substrate materials tested, phosphonylation and calcium posttreatment resulted in an increased propensity for bone binding and apposition, as measured by push out test. Microtextured PP, PE, and CFR-PEEK surfaces that were further phosphonylated and calcium treated resulted in test samples with an increased interfacial strength. PMID- 11108203 TI - Absorbable mesh aids in development of discrete, tissue-engineered constructs. AB - Absorbable mesh was investigated as a potential containment material in which to house discrete, small, tissue-engineered constructs. The mesh was fashioned into bags of varying shapes and consistent volumes. Cells were cultivated on porous, collagen beads, and the tissue constructs were placed into the bags. The mechanical integrity of the bags and feasibility of the design was tested in vitro. The bags successfully maintained their integrity as the cells developed on the collagen matrices. Furthermore, their porosity allowed access of nutrients and waste products to and from the developing tissue. Having demonstrated feasibility of processing, the next step is to optimize the cell culture specifications and materials design. PMID- 11108204 TI - Reducing condylar compression in clenching patients. AB - The two major muscle groups used during clenching activity are the masseter and temporalis muscles. EMG readings of the masseter and temporalis muscles rise significantly during times of macro-clenching. Clenching occurs when the masseter and temporalis muscles contract, pulling the mandible superiorly. The continued contraction of the masseter and temporalis muscles results in compression forces on the teeth and temporomandibular joints. Theoretical joint loading models are utilized to demonstrate the load on the TMJ due to forces generated by the masseter and temporalis muscles. This study measures the EMG readings during bilateral macro-contraction of the masseter and anterior temporalis muscles. An appliance is fabricated to disengage the posterior teeth and a second series of EMG readings are taken to record lowered EMG readings. The vector forces of the reduced EMG's recordings demonstrate reduced condylar compression during macro clenching. PMID- 11108205 TI - Correlative experimental animal and human clinical retrieval evaluations of hydroxyapatite (HA)-coated and non-coated implants in orthopaedics and dentistry. AB - Retrieval analyses disclosed in vivo dissociation of HA in orthopaedic acetabular components, but excellent bone ingrowth into intact HA coatings on dental retrievals. Initial healing and the bone interface between HA-coated and non coated implants in the posterior maxilla (Mx) and mandible (Md) was assessed in an animal model using light microscopy (LM), including confocal (CM) and Nomarski (NM) microscopy. Seventy-two implants (36 HA-coated; 36 non-coated) were placed into jaws of six dogs; half after extraction, half after 3 months healing. Animals were euthanized 3 months postimplantation. All implants osseointegrated; however, preliminary morphometry showed higher BCL for HA-coated (51%) than non coated implants (44%) in the Mx (p < 0.05). BCL for HA-coated Md implants was not significantly higher (64%) than non-coated implants (62%). Bone closely apposed both implant types; however, LM suggested a more intimate association with HA coatings. Serial sections disclosed a reddish coating on the HA, possibly analogous to oral tissue proteoglycans, which was not visible with non-coated implants. This material was continuous with similar material coating endosteum, osteoid regions, and osteocyte (Os) lacunae close to the implant. An interdigitating canaliculi network allowed communication between interfacial Os and Os deeper within the bone. Data suggest HA offers enhanced initial bone fixation in the Mx, and that adequate bone exists for non-coated implant stability in the Md. No HA dissociation was seen with implants in the animal study, which was consistent with retrieved human HA dental implants. PMID- 11108206 TI - A biomechanical evaluation of the Christensen temporomandibular joint implant. AB - The Christensen TMJ implant is often used clinically as a total joint replacement of the temporomandibular joint. The system consists of a thin fossa component and a condylar component with a polished articular head. In this study, we analyzed the surface finish and the metal structure of the components. We also measured the contact areas between the two components for different load levels. Such information may be useful in evaluating clinical performance as well as in making future improvements in the design of these implants. PMID- 11108207 TI - A 3-D FEM analysis of single and multiple screw-root dental implant fixed in a mandible. AB - Replacement of single tooth using a threaded titanium screw root coated with hydroxyapatite (HA) for faster bone apposition to implant site is common. Uncoated pure titanium is also used for osseointegration. Usually bone fixation of implant takes four to six months in either case. Quite often, a good number of teeth in a mandible or maxilla are replaced. Usually it is often said that bones appear to be well designed from the point of view of structural engineering. The "maximum-minimum law" claimed by Roux is a rational concept that states that bone provides maximum strength with a minimum of construction material. According to this proposition, stress distribution in bone will be almost uniform under a set of loading conditions. This was found to be true in the case of normal human mandibular bone as examined by some Japanese scientists. The present authors are interested in examining the stress distribution during multiple single tooth replacements using 3D-FEM technique to ascertain how the stress pattern changes with such implantation of three screws in a row in the human mandible, as we are in the process of clinical trial of hydroxyapatite coated and uncoated titanium implant. This will be of considerable interest to the dental surgeons, who prompted us to address this problem. Our results depicted the mean values of various stress in cortical and cancellous bone while applying the maximum masticatory load of 50 N in each teeth. The generated stress level is within the safe range of stress for bones. However, such screws cannot be applied to osteoporotic or other diseased mandible where bone strength may be quite low. PMID- 11108208 TI - Stress analysis of an artificial temporal mandibular joint. AB - To design a temporal mandibular joint (TMJ), the designer should pay attention to the range of movement in the joint, the strength of the joint, and the size of the implant should conform, so that it does not hamper facial configuration. As a number of designs are available, in this study we have considered one of the most common and widely used implants for analysis. The main objective of this study is to examine the stress-strain behavior at the implant and what is happening at the implant bone interface. We have also examined whether implant material can be replaced by UHMWPE (ultra high molecular weight polyethylene) instead of titanium or Co-Cr-Mo alloy. Whether the change of positions of the screw used for fixation has any effect or not, we have modeled it considering actual shape and size, then divided it into number finite elements by using a FEM package. An appropriate surgical construct was modeled and loaded and studied for different parameters. We have shown that the metallic prostheses are good from a stress-strain point of view and UHMWPE cannot be used as such. PMID- 11108209 TI - KIDSCARE: a network for children with special health care needs. AB - In December 1997, Arkansas Medicaid in cooperation with Title V and Children's Medical Services (CMS) implemented guidelines for the preauthorization of therapy services for Children with Special Health Care Needs (CSHCN). The University of Arkansas for Medical Sciences (UAMS) in partnership with Arkansas Medicaid (MCD), and Chidren's Medical Services (CMS) were given the charge of developing clinical practice guidelines for physical therapy (PT), occupational therapy (OT), and speech/language therapy (SLP) for this population based on primary diagnosis and age appropriateness. This process involved the implementation of a statewide, internet-based, clinical information support network. The objective of this article is to describe KIDSCARE, a clinical information support network, currently under development, for the determination of medical necessity and allocation of therapy services for CSHCN. PMID- 11108210 TI - A radio controller using speech for the blind. AB - This article describes a low-cost, portable real-time DSP-based speech controller system to provide radio interface control command applications for the blind. The system recognizes spoken Mandarin Chinese words on a DSP chip (TMS320C31) using a hidden Markov model. The function of the radio set, which includes a tuner, tape, and compact disc, were evaluated under both noisy and noiseless environments. Four subjects took part in the experiment and achieved 83 and 90% mean recognition rates under noisy and noiseless conditions, respectively. In addition, because this system is based on a DSP chip, it can easily be programmed to execute speaker-independent algorithms. PMID- 11108211 TI - An artificial neural network for the electrocardiographic diagnosis of left ventricular hypertrophy. AB - OBJECTIVE: A neural network was constructed to predict the presence of left ventricular hypertrophy (LVH) using both clinical information and the electrocardiogram (ECG). DESIGN AND SETTING: In this retrospective study of 317 adult male patients, clinical parameters were age and history/physical examination: normal, heart failure, LV outflow obstruction, mitral regurgitation or aortic regurgitation. Multiple ECG parameters were used. A back-propagation neural network was constructed. The network was trained on 217 patients. A test set of 100 patients was then evaluated. The network was used to predict both LV mass and LVH by the criterion of LV mass index > 132 g/m2. RESULTS: LV mass was predicted with an accuracy of 79%. In predicting LVH, the network showed 82% correct diagnosis, sensitivity 94%, and specificity 65%. Positive predictive accuracy was 81% and negative predictive accuracy was 89%. CONCLUSIONS: The neural network integrates clinical and ECG data and its resultant prediction of LVH is superior to that obtained using conventional ECG diagnostic criteria. PMID- 11108212 TI - Metal-oxide nanoparticles for the reinforcement of dental restorative resins. AB - Metal oxide nanoparticles were synthesized from tantalum ethoxide and zirconium isopropoxide and subsequently surface grafted with vinyl silane and silyl methacrylate coupling agents. The nanoparticles were then dispersed into a commercial dental resin, and the composite was photocured into rigid three-point bend and fracture toughness specimens. The optically transparent/translucent cured composites demonstrated strength, toughness, and elastic modulus inferior to the unfilled material. Therefore, modifications in surface functionalization are being made to improve coupling and reduce interparticle associations. PMID- 11108213 TI - Effect of carbon coating on the properties of gamma irradiated ultra-high molecular-weight polyethylene specimens. AB - We reported the effect of carbon coating on the changes in properties of the ultra-high-molecular-weight polyethylene standard material (Hospital for Special Surgery) alter gamma irradiation in air and storage for 2 years. The coating showed a slight improvement in crystallinity (X-ray) and tensile properties (under cyclic loading) over the uncoated and irradiated control group. The oxidation level as measured by Fourier- transform infrared spectroscopy was unaffected by coating. PMID- 11108214 TI - Relationship between fracture toughness and impact strength of acrylic bone cement. AB - This work involved determining the fracture toughness, KIc (in MPa(square root)m) (using rectangular compact tension specimens) and impact strength, IS (in kJ/m2) (Charpy type specimens) of Surgical Simplex P and three variants of Palacos R acrylic bone cements. The best fit to these results yielded a power relationship KIc = 0.795(IS)0.59. The usefulness of this relationship is detailed, especially for the purpose of performing quality control checks on bone cements. PMID- 11108215 TI - Improved orthopaedic bone cement formulations based on rubber toughening. AB - Poly(methylmethacrylate) (PMMA) bone cements have been used for the fixation of hip and knee implants since the early 1960s. Aseptic loosening, related to fracture of the PMMA, continues to be the primary mode of failure for these prostheses. Failed prostheses must be replaced causing additional expense and patient trauma. Furthermore, the average lifetime of the revised prosthesis is significantly lower than that of a primary prosthesis. Recent work by Moseley and co-workers led to the development of a promising new rubber toughened cement. It is comprised of a matrix of the traditional PMMA with dispersed rubber particles to modify mechanical properties and, in particular, improve fracture toughness. The fracture toughness of the experimental material was 167% greater than the toughness of a nontoughened control; however, the elastic modulus and compressive strength were reduced. The reductions in properties should not pose a clinical problem based on results of the implant model reported by Moseley. More serious concerns were mixing and delivery problems and high residual monomer concentrations. The formulation and chemical/mechanical characterization of new toughened acrylic formulations that have residual monomer levels equivalent to Simplex and better mixing properties are reported. PMID- 11108216 TI - RBF networks for source localization in quantitative electrophysiology. AB - The backpropagation neural network methods have been proposed recently to solve the inverse problem in quantitative electrophysiology. A major advantage of the technique is that once a neural network is trained, it no longer requires iterations or access to sophisticated computations. We propose to use RBF networks for source localization in the brain, and systematically compare their performance to those of Levenberg-Marquardt (LM) algorithms. We show the use of two types of Radial Basis Function Networks (RBF) network: a classic network with fixed number of hidden layer neurons and an improved network, Minimal Resource Allocation Network (MRAN), recently proposed by one of the authors, capable for dynamically configuring its structure so as to obtain a compact topology to match the data presented to it. PMID- 11108217 TI - Development of a frequency-dependent-type apex locator with automatic compensation. AB - Among apex locators, the frequency dependent type is more accurate and convenient to use than others. However, the accuracy of the apex locator is still influenced by the presence of various electrolytes used in root canal treatments. In this study, we developed a frequency-dependent electronic apex locator minimizing the influence of electrolytes on the measurement of root canal lengths. It was confirmed that the two frequencies of 0.5 and 10 kHz are better than the conventional ones of 0.4 and 8 kHz. The impedance ratio of the two different frequencies represents the position of the file in root canal, and the voltage difference of the two frequencies represents the status of the fluid in the root canal. As a result of compensation using the voltage differences of the two frequencies, the errors decreased significantly on average from +0.54 mm to +0.18 mm in H2O2 solution (p < 0.01), and from -0.33 mm to -0.01 mm in NaOCl solution (p < 0.01) in the tooth model experiments. PMID- 11108218 TI - Biomedical application of an electronic nose. AB - The analysis of volatiles secreted outside the human body to get information on the health status of the individuals has been proposed several times in the past. This kind of analysis is complex both from the point of view of sample collection and data interpretation when, for instance, gas chromatography is utilized. In the recent years the advent of chemical sensors and chemical sensors systems (the so-called electronic noses) opened the way to the possibility of fast and simple analysis of odors in many fields, and, recently, among them, in medicine. In this paper some examples of these applications are illustrated. The results, although preliminary, encourage in pursuing these researches that can give rise to a better comprehension of the role of smell and odor in humans and, possibly in the near future, in novel diagnostic tools. PMID- 11108219 TI - An active contour model algorithm for tracking endocardiac boundaries in echocardiographic sequences. AB - The use of active contour models to track the boundaries of anatomic structures in medical images is a technique that has attracted a great number of efforts during the last decade. Segmentation techniques based in deformable active contours were proposed first by Kass et al. Because of the problems appearing using these models, some solutions have been introduced, such as balloon force or Gradient Vector Flow force (GVF), derived from the Gradient Vector Flow vectorial field. Results obtained with these forces in the tracking endocardiac task in echocardiographic sequences were not adequate. We have designed a new external force called hybrid force, which, by combining both forces, joins the main features of each one. PMID- 11108220 TI - Efficient sampling in 3-D stereologic volumetry. AB - Present 3-D stereology techniques use fixed grid sampling where the sampling grid of test points is randomly placed in 3-D. In order to increase randomness of sampling, in this paper we propose random grid sampling scheme where grid sections are placed randomly over 2-D cross-sections. A random grid sampler has superior efficiency and accuracy compared with fixed grid sampling. Using MR head scans we demonstrate the use of random grid sampling in volumetry of structures in 3-D biomedical images. PMID- 11108221 TI - Autocorrelation function analysis of ECG signals in 20 rabbits. AB - In this experiment, we measured ECG signals of lead I, respectively, during normal and arrhythmia after 20 rabbits were anaesthetized. These signals were recorded, then the information was put into a computer and it analyzed Autocorrelation (AC) Function, and Power Spectrum. The results make us know: (1) periodicity corresponding relationship of ECG in normal rabbits can derive reflects from changes of ECG AC function changes of ECG signals in arrhythmia rabbits can derive relatively sensitive reflection from changes of ECG function (attenuating area S and the value of k) in normal rabbits. (2) The Autocorrelation Function of Electrocardio change but ECG in the normal. All these provide another useful method for the diagnosis of arrhythmia. PMID- 11108222 TI - Beyond a code of ethics for bioengineers: the role of ethics in an integrated compliance program. AB - Developing a code of ethics for biomedical engineering professionals is a very important first step in clarifying their professional obligations and in helping to establish and maintain their professional autonomy. However, it is only that- a first step. Unless ways can be found to bring the principles contained in this code to bear on the everyday decision making of these professionals, this code will have little practical influence. One effective way to bring a code of ethics to bear on decision making is to integrate it into organizational compliance programs. Such programs often have company-specific codes of ethics attached to them, and these company-specific codes can either include the principles contained in the professional code of ethics or reference the code by title. After defining what I take to be the challenge of compliance, I consider four (4) roles that codes of ethics and ethics generally can play in helping to create and sustain programs at the organizational level that integrate ethics and compliance and thereby aim to make a practical difference in the everyday decision making of bioengineering professionals. These four roles include: framing the program, grounding the standards, achieving critical distance, and creating and sustaining an ethical organizational culture. PMID- 11108223 TI - The need for a professional code of ethics in biomedical engineering: a lesson from history. PMID- 11108224 TI - Ethical considerations for biomedical scientists and engineers: issues for the rank and file. AB - Biomedical science and engineering is inextricably linked with the fields of medicine and surgery. Yet, while physicians and surgeons, nurses, and other medical professionals receive instruction in ethics during their training and must abide by certain codes of ethics during their practice, those engaged in biomedical science and engineering typically receive no formal training in ethics. In fact, the little contact that many biomedical science and engineering professionals have with ethics occurs either when they participate in government funded research or submit articles for publication in certain journals. Thus, there is a need for biomedical scientists and engineers as a group to become more aware of ethics. Moreover, recent advances in biomedical technology and the ever increasing use of new devices virtually guarantee that biomedical science and engineering will become even more important in the future. Although they are rarely in direct contact with patients, biomedical scientists and engineers must become aware of ethics in order to be able to deal with the complex ethical issues that arise from our society's increasing reliance on biomedical technology. In this brief communication, the need for ethical awareness among workers in biomedical science and engineering is discussed in terms of certain conflicts that arise in the workaday world of the biomedical scientist in a complex, modern society. It is also recognized that inasmuch as workers in the many branches of bioengineering are not regulated like their counterparts in medicine and surgery, perhaps academic institutions and professional societies are best equipped to heighten ethical awareness among workers in this important field. PMID- 11108225 TI - Managed care and the challenge of medical ethics. PMID- 11108226 TI - Wrongdoing in biomedical research: an ethical diagnosis and prescription. AB - Attention is focused on wrongdoing as a practice-specific notion to be fleshed out by reference to the ethos of a practice such as biomedical research. Wrongdoing in this sense, which is not the same thing as scientific misconduct, has not received the attention it deserves. There are two reasons for this: (1) we have a tendency to be ethically reactive and (2) we tend to be preoccupied with questions that are highly charged politically, socially, and morally. Explaining this further, two types of ethical questions are distinguished- whether-we-ought questions and how-we-ought questions. Using the Baltimore case for purposes of illustration, it is argued that failure to attend to the latter sort of questions is detrimental to the practice of biomedical research. Answering such questions requires careful attention to the ethos of the practice of biomedical research as well as action on the part of practitioners, particularly those who serve as mentors to persons entering the profession. PMID- 11108227 TI - Biomedical research: some ethical challenges. AB - Research ethics is currently becoming the focus of attention as advancements in science are more in the public spotlight. Notoriety brings attention not only to the achievements, but to some of the more pressing issues that have risen to the surface in the debate on research ethics. Such issues include: conflicts of interest and industrial funding, research misconduct, and the need of ethics training for biomedical scientists. PMID- 11108228 TI - The ethical and social dimensions of home-based telemedicine. AB - The growth of home-based telemedicine invites a discussion about the ethical implications of this technology for the traditional goals and moral ideals of medical practice. To this end, I examine the normative connections between home based telemedicine and the following: (1) health care justice, (2) quality of life for patients and family caregivers, and (3) provider-patient relationships. PMID- 11108229 TI - What is life, and what is a machine? The ontology of bioengineering. AB - In his Keynote address to the First Conference at Clemson University on Ethical Issues in Biomedical Engineering, George Bugliarello suggested that a most important ethical issue for bioengineering "is the positioning of the bio-machine interface." "Where," he asked, "should the biological organism end and the machine begin?" Central to this question of the limits of life and engineering is the more fundamental question of how life differs and how it is similar to a machine. This paper argues that until very recently, science, by its very nature, has treated life as if it were a machine, or has treated the parts of living systems as if they were machines. The distinctive feature of a machine is that its behavior is linear and hence predictable. On the other hand, living organisms may not be linear, but rather nonlinear systems. Thus, the interface between organism and machine may be conceived as the interface between nonlinear and linear systems. PMID- 11108230 TI - Using rhetorical theory in medical ethics cases. AB - In this paper I argue that rhetorical theory is a valuable tool in medical ethics cases. The case I use as an example is one in which traditional, philosophy-based medical ethics are applied. In this case the traditional ethical approach is not adequate to the task. Key issues and problems are not addressed, resulting in a problem that seems to be solved on the surface, but, when rhetorically analyzed, it's obvious that none of the issues have been resolved in any satisfactory way. By using rhetorical theory, such as that Michel Foucault uses in Power/Knowledge, we discover that the reason this case has not been solved is that the power issues have not been addressed. Using Foucault's concepts of "subjugated knowledge", "local knowledge", "situated knowledge", and "docile bodies", we can tease out the real issues that surface in this ethics case and solve them. Foucault also recommends we use theory as a "toolkit". I propose a model that is a further iteration of this idea. My model uses numerous rhetorical and literary theories, depending on the issues that need to be addressed in each individual medical ethics case. I briefly describe the various theories and include a handout of what the new model of using rhetorical theory in such cases would look like. PMID- 11108231 TI - Genetic testing of the general population: ethical and informatic concerns. AB - Whether we like it or not, genetic testing will almost certainly become routine medical practice within the next 25 years. Integrated circuit chips already exist that can perform 400 genetic tests simultaneously, thus greatly reducing the costs. At least one company is already working on a prototype for a handheld genetic tester that would allow primary care physicians to perform hundreds or thousands of genetic tests on a simple blood smear in just a few minutes. "Genetic report cards" for children are not very far off at all. The use of such widespread testing poses a variety of ethical dilemmas. One problem that has not been appreciated sufficiently, however, is the question of how to interpret the test results. Because of the ways the genes implicated in diseases are discovered and marketed, quantitative analysis of the tests can be extremely misleading. The difficulty is that we simply do not have sufficient information about variance in genetic and other factors in the general population to make accurate projections of a patient's risk, given the presence of a gene. This uncertainty is obscured, however, when we provide the patient with a numerical analysis of risk because it is well established that people tend to overestimate the information content of numerical projections. This situation is made far worse by the fact that we do not have enough adequately trained genetic counselors to handle the load that will soon be placed on them (and studies have shown that physicians are generally very poorly prepared to act as accurate sources of information on complex genetic issues). For these reasons, I argue that access to genetic testing should be treated the same way as access to new medical procedures and medications--namely, withheld from the general public until proven safe and effective in large-scale trials. This is certain to be an unpopular policy, but it seems the only way to prevent a great deal of abuse of genetic tests. PMID- 11108232 TI - Developing a code of ethics for human cloning. AB - Under what conditions might the cloning of human beings constitute an ethical practice? A tendency exists to analyze human cloning merely as a technical procedure. As with all revolutionary technological developments, however, human cloning potentially exists in a broad social context that will both shape and be shaped by the biological techniques. Although human cloning must be subjected to technical analysis that addresses fundamental ethical questions such as its safety and efficacy, questions exist that focus our attention on broader issues. Asserting that cloning inevitably leads to undesirable consequences commits the fallacy of technological determinism and untenably separates technological and ethical evaluation. Drawing from the Report of the National Bioethics Advisory Committee and Aldous Huxley's Brave New World, we offer a draft "Code of Ethics for Human Cloning" in order to stimulate discussion about the ethics of the broader ramifications of human cloning as well as its particular technological properties. PMID- 11108233 TI - Ethical issues in cloning. AB - There is great public concern with the ethics of human cloning. This paper briefly examines some of what I identify as pseudo-problems or myths associated with cloning, and some of the more substantial ethical concerns. PMID- 11108234 TI - The use of millimeter band electromagnetic waves in clinical oncology. PMID- 11108235 TI - Effect of EHF radiation on metabolism of Cyanobacteria spirulina platensis and other photosynthesizing organisms. AB - A stimulatory influence of EHF radiation on the growth and physiological activity of photosynthetic organisms is shown. Some primary responses to this action are studied. Oxygen is considered to be of great importance in the mechanism of the stimulators effect of EHF radiation. PMID- 11108236 TI - Fundamentals and prospects of passive thermoacoustic tomography. AB - The problems and possibilities of passive thermoacoustical tomography are discussed. Algorithms for reconstruction of internal temperature in the human body are proposed. These algorithms take into account heat transfer and blood circulation and the absorption factor, obtained previously. The results of reconstruction of deep temperature in the human hand in simulations with the medium with a heated object are reported. These results support the possibility of the correlation measurements of the thermal acoustic radiation. Such measurements allow the information on ultrasound absorption by the object under study to be obtained and open the way to the development of a passive acoustic tomography system using a priori information on the absorption factor. PMID- 11108237 TI - Morphological changes in skin nerves caused by electromagnetic radiation of the millimeter range. AB - The morphological changes in skin nerves of BALB/C mice after millimeter wavelength range electromagnetic exposure at a frequency of 42.25 GHz and power of 50 mW/cm2 were studied. Immediately after 15 minutes of exposure, the destruction of the cytoplasm of myelinated and unmyelinated axons was found. PMID- 11108238 TI - MM therapy in traumatology and orthopedy. PMID- 11108239 TI - Green fluorescent proteins light the way to a better understanding of the function and regulation of specific anterior pituitary cells. PMID- 11108240 TI - Localization of G protein alpha-subunits in the human fetal adrenal gland. AB - The aim of the present study was to investigate the presence and localization of the main G protein alpha-subunits in the human fetal adrenal gland during the second trimester of gestation. Immunofluorescence studies conducted on sections from frozen glands obtained immediately after therapeutic abortion indicated that the alpha s subunit of the heterotrimeric Gs protein was detected in all adrenal cell types, except for endothelial cells. The other alpha-subunits had a more specific pattern of distribution. Indeed, the alpha il-2 protein was restricted to the definitive zone, whereas alpha i3 labeling was mainly expressed in the fetal zone. The alpha q protein subunit was localized in vascular endothelial cells at the periphery of the adrenal gland and in fetal cells at the center. Finally, chromaffin cells expressed alpha s, alpha q, and alpha o1, but not alpha o2 nor alpha i. Altogether, these results indicate that the human fetal adrenal gland is not only unique in its particular morphology and expression of steroidogenic enzymes, but also by the differential expression of G protein alpha subunits. Such cell specific distribution in glands from midgestational fetuses may account for the absence or the different responses to stimuli, when compared with the adult adrenal gland. PMID- 11108241 TI - Tumor necrosis factor-alpha converting enzyme (TACE) is a growth hormone binding protein (GHBP) sheddase: the metalloprotease TACE/ADAM-17 is critical for (PMA induced) GH receptor proteolysis and GHBP generation. AB - The GH binding protein (GHBP), which exists in many vertebrates, is a circulating high affinity binding protein corresponding to the extracellular domain of the GH receptor (GHR). In humans, rabbits, and several other species, the GHBP is generated by proteolysis of the GHR and shedding of its extracellular domain. We previously showed that GHBP shedding is inducible by the phorbol ester phorbol 12 myristate,13-acetate (PMA) and inhibited by the metalloprotease inhibitor, Immunex Corp. Compound 3 (IC3). The metzincin metalloprotease, tumor necrosis factor-alpha (TNF-alpha)-converting enzyme (TACE), catalyzes the shedding of TNF alpha from its transmembrane precursor, a process that is also inhibitable by IC3. TACE may hence be a candidate for GHBP sheddase. In this study, we reconstitute fibroblasts derived from a TACE knockout mouse (Null cells) with either the rabbit (rb) GHR alone (Null/R) or rbGHR plus murine TACE (Null/R+T). Although GHR in both cells was expressed at similar abundance, dimerized normally and caused JAK2 activation in response to GH independent of TACE expression, PMA was unable to generate GHBP from Null/R cells. In contrast, PMA caused ample GHBP generation from TACE reconstituted (Null/R + T) cells, and this GHBP shedding was substantially inhibited by IC3 pretreatment. Corresponding to the induced shedding of GHBP from Null/R + T cells, PMA treatment caused a significant loss of immunoblottable GHR in Null/R+T, but not in Null/R cells. We conclude that TACE is an enzyme required for PMA-induced GHBP shedding and that PMA-induced down-regulation of GHR abundance may in significant measure be attributable to TACE-mediated GHR proteolysis. PMID- 11108242 TI - Photoperiod regulates growth, puberty and hypothalamic neuropeptide and receptor gene expression in female Siberian hamsters. AB - In seasonal mammals, both the growth and reproductive axes are regulated by photoperiod. Female Siberian hamsters were kept, for up to 12 weeks, in long-day (LD) or short-day (SD) photoperiod, from weaning at 3 weeks of age (Exp 1). LD hamsters had characteristically faster growth and higher asymptotic body weight, adiposity, and leptin gene expression in adipose tissue. Only LD females attained puberty. Gene expression in the hypothalamic arcuate nucleus for leptin receptor (OB-Rb), POMC, and melanocortin 3-receptor (MC3-R) was higher in LD but did not change from weaning levels in SD. In contrast, gene expression in the arcuate nucleus for cocaine and amphetamine-regulated transcript (CART) was higher in SD than LD, a difference that was apparent at 2 weeks post weaning. Transfer of SD females to LD at 15 weeks post weaning (Exp 2) increased body weight, leptin signal, and gene expression for POMC but failed to induce normal puberty onset or to increase gene expression for OB-Rb and MC3-R. Therefore, photoperiodic regulation of puberty may be modulated by age, by photoperiodic history, and by changes in leptin signaling and the activity of the leptin-sensitive hypothalamic melanocortin system (POMC, MC3-R). A role for CART in photoperiodic regulation of growth is suggested, because the changes in CART gene expression preceded significant divergence of growth trajectories in the opposite photoperiods. PMID- 11108243 TI - Extracellular calcium-sensing receptor expression and its potential role in regulating parathyroid hormone-related peptide secretion in human breast cancer cell lines. AB - Metastasis of breast cancer to bone occurs with advanced disease and produces substantial morbidity. Secretion of PTH-related peptide (PTHrP) from breast cancer cells is thought to play a key role in osteolytic metastases and is increased by transforming growth factor-beta (TGFbeta), which is released from resorbed bone. Elevated extracellular calcium (Ca2+(o)) also stimulates PTHrP secretion from various normal and malignant cells, an action that could potentially be mediated by the Ca2+(o)-sensing receptor (CaR) originally cloned from the parathyroid gland. Indeed, we previously showed that both normal breast ductal epithelial cells and primary breast cancers express the CaR. In this study we investigated whether the MCF-7 and MDA-MB-231 human breast cancer cell lines express the CaR and whether CaR agonists modulate PTHrP secretion. Northern blot analysis and RT-PCR revealed bona fide CaR transcripts, and immunocytochemistry and Western analysis with a specific anti-CaR antiserum demonstrated CaR protein expression in both breast cancer cell lines. Furthermore, elevated Ca2+(o) and the polycationic CaR agonists, neomycin and spermine, stimulated PTHrP secretion dose dependently, with maximal, 2.1- to 2.3-fold stimulation. In addition, pretreatment of MDA-MB-231 cells overnight with TGFbeta1 (0.2, 1, or 5 ng/ml) augmented both basal and high Ca2+-stimulated PTHrP secretion. Thus, in PTHrP secreting breast cancers metastatic to bone, the CaR could potentially participate in a vicious cycle in which PTHrP-induced bone resorption raises the levels of Ca2+(o) and TGFbeta within the bony microenvironment, which then act in concert to evoke further PTHrP release and worsening osteolysis. PMID- 11108244 TI - Leukemia inhibitory factor can substitute for nidatory estrogen and is essential to inducing a receptive uterus for implantation but is not essential for subsequent embryogenesis. AB - A stage critical in mammalian development is embryo implantation. At this point, the blastocyst establishes a close interaction with the uterine tissues, a step necessary for its continued embryonic development. In many mammalian species, including man, uterine expression of the cytokine, leukemia inhibitory factor (LIF) is coincident with the onset of implantation and in mice LIF is essential to this process. The reasons for implantation failure have not been established. Here we show in LIF-deficient mice that up to the onset of implantation, changes in uterine cell proliferation, hormone levels, blastocyst localization, as well as expression of lactoferrin and Muc-1, do not differ from wild-types. However, the uterus fails to respond to the presence of embryos or to artificial stimuli by decidualizing. In mice, implantation and decidualization are induced by nidatory estrogen. We show that uterine expression of LIF is up-regulated by estrogen and LIF can replace nidatory estrogen at inducing both implantation and decidualization in ovariectomized mice. Implantation of LIF-deficient embryos in the LIF-deficient females, with normal development to term is rescued by i.p. injection of LIF. Transient expression of LIF on D4 of pregnancy is therefore only required to induce a state of receptivity in the uterus permitting embryo implantation and decidualization. LIF is neither required by the embryo for development nor for the maintenance of pregnancy. PMID- 11108245 TI - Regulation of thyroid follicular cell function by intracellular redox-active copper. AB - Pyrrolidine dithiocarbamate (PDTC) is a metal-chelating compound that exerts prooxidant or antioxidant effects and is widely used to study redox regulation of cell function. In the present study, we investigated effects of PDTC on the function of rat thyroid follicular FRTL-5 cells. Treatment of the cells with PDTC resulted in a marked decrease in Pax-8 messenger RNA level and its DNA-binding activity. This decrease was associated with a significant reduction in thyroperoxidase (TPO) messenger RNA level. Expression of TTF-1 and thyroglobulin was not affected by PDTC. Treatment with PDTC also decreased DNA-binding activity of p53, a tumor suppressor protein, and increased cell proliferation rates. These changes were not observed by the treatment with another antioxidant, N-acetyl-L cysteine, suggesting that the metal-chelating, prooxidant property of PDTC is responsible for its effects. Indeed, the intracellular level of copper was significantly increased by PDTC. Treatment with bathocuproinedisulfonic acid, a noncell-permeable chelator of Cu1+, abrogated the copper increase by PDTC and its effects on Pax-8 and TPO expression as well as on p53 binding. Taken together, these results indicate that the intracellular level of redox-active copper is crucial for Pax-8 and TPO expression and for proliferation of thyroid follicular cells. PMID- 11108246 TI - p38 mitogen-activated protein kinase mediates tumor necrosis factor-alpha-induced apoptosis in rat fetal brown adipocytes. AB - Tumor necrosis factor-alpha (TNFalpha) induces apoptosis and cell growth inhibition in primary rat fetal brown adipocytes. Here, we examine the role played by some members of the mitogen-activated protein kinase (MAPK) superfamily. TNFalpha activates extracellular regulated kinase-1/2 (ERK1/2) and p38MAPK. Inhibition of p38MAPK by either SB203580 or SB202190 highly reduces apoptosis induced by TNFalpha, whereas ERK inhibition potentiates it. Moreover, cotransfection of an active MKK3 mutant and p38MAPK induces apoptosis. p38MAPK inhibition also prevents TNFalpha-induced cell cycle arrest, whereas MEK1 inhibition enhances this effect, which correlates with changes in proliferating cell nuclear antigen expression, but not in cyclin D1. c-Jun and activating transcription factor-1 are potential downstream effectors of p38MAPK and ERKs upon TNFalpha treatment. Thus, TNFalpha-induced c-Jun messenger RNA expression requires ERKs activation, whereas p38MAPK inhibition enhances its expression. In addition, TNFalpha-induced activating transcription factor-1 phosphorylation is extensively decreased by SB203580. However, TNFalpha-induced NF-kappaB DNA binding activity is independent of p38MAPK and ERK activation. On the other hand, C/EBP homology protein does not appear to mediate the actions of TNFalpha, because its expression is almost undetectable and even reduced by TNFalpha. Finally, although TNFalpha induces c-Jun N-terminal kinase (JNK) activation, transfection of a dominant negative of either JNK1 or JNK2 had no effect on TNFalpha-induced apoptosis. These results suggest that p38MAPK mediates TNFalpha induced apoptosis and cell cycle arrest, whereas ERKs do the opposite, and JNKs play no role in this process of apoptosis. PMID- 11108247 TI - Retinoic acids and thyroid hormone act synergistically with dexamethasone to increase growth hormone-releasing hormone receptor messenger ribonucleic acid expression. AB - The effects of all-trans-retinoic acid (RA), 9-cis-retinoic acid (9cRA), and thyroid hormone (T3) on GH-releasing hormone receptor (GHRH-R) messenger RNA (mRNA) expression were studied using ribonuclease protection assay in the fetal rat pituitary gland and in MtT/S cells, a clonal GH cell line derived from an estrogen-induced somatotropic tumor in the rat. Although RA (1 microM), 9cRA (1 microM), or T3 (1 nM) alone showed little effect on GHRH-R mRNA expression in the MtT/S cells, each of these substances was found to act synergistically with dexamethasone (DEX; 500 nM) to increase GHRH-R mRNA expression. The effects of RAs and T3 were dose dependent, with maximum effects observed at 1 microM and 1 nM, respectively. The maximum effect of RAs or T3 was not further augmented by the addition of T3 or RAs, respectively. No apparent differences were observed in this study between the actions of RA and 9cRA. The Northern analyses showed that MtT/S cells express retinoic acid receptor alpha2 mRNA and thyroid hormone receptor beta2 mRNA, and DEX did not affect the levels of these mRNAs. This suggests that the role of DEX in enabling RAs or T3 to up-regulate GHRH-R mRNA levels is not an induction of the expression of each specific receptor for RAs and T3. The similar enhancement of DEX induction of GHRH-R mRNA by RAs or T3 was also observed in the fetal rat pituitary gland in culture, suggesting that RA and/or T3 is involved in the mechanisms responsible for the developmentally regulated expression of GHRH-R mRNA. PMID- 11108248 TI - Existence of galanin in lumbosacral sympathetic ganglionic neurons that project to the quail uterine oviduct. AB - Oviposition in birds is conducted by vigorous contractions of the uterine oviduct. We recently isolated an oviposition-inducing peptide that was identified as avian galanin from mature quail oviducts. This peptide was localized in neuronal fibers terminating in muscle layers in the uterine oviduct and evoked vigorous uterine contractions through binding to receptors located in the uterus. However, no cell bodies that express avian galanin were detected in the uterus or other oviduct regions. To understand the control mechanism of avian oviposition by galanin, we identified the neurons that synthesize galanin and project to the uterus with the combination of retrograde labeling with neurobiotin and immunocytochemistry for galanin in mature Japanese quails. Retrograde labeling with neurobiotin from the uterus revealed that lumbosacral sympathetic ganglionic neurons located in the uterine side projected their axons to the uterine muscle layer. Abundant elementary granules were observed in somata of the retrogradely labeled sympathetic ganglionic neurons, suggesting that labeled neurons may function as a neurosecretory cell. Immunocytochemical analysis with the antiserum against avian galanin showed an intense immunoreaction restricted to somata of the retrograde-labeled ganglionic neurons. Preabsorbing the antiserum with avian galanin resulted in a complete absence of the immunoreaction. Competitive enzyme linked immunosorbent assay using antigalanin serum confirmed that avian galanin existed in the sympathetic ganglionic neurons. Expression of the avian galanin messenger RNA in the neurons was further verified by Northern blot analysis. In addition, both avian galanin and its messenger RNA in the neurons were highly expressed in mature birds, unlike in immature birds. These results suggest that lumbosacral sympathetic ganglionic neurons innervating the uterine muscle produce avian galanin in mature birds. Because this peptide acts directly on the uterus to evoke oviposition through a mechanism of the induction of vigorous uterine contraction, galaninergic innervation of the uterine oviduct may be essential for avian oviposition. PMID- 11108249 TI - Molecular cloning and expression of a functionally different alternative splice variant of prointerleukin-1alpha from the rat testis. AB - We report here the characterization of an alternative splice variant of prointerleukin-1alpha (proIL-1alpha), constitutively expressed by the normal adult rat testis. In addition to the classical 32K proIL-1alpha (32proIL-1alpha) messenger RNA, the testis produced a shorter variant encoding a putative protein of 24K (24proIL-1alpha). In situ hybridization demonstrated constitutive expression of the splice transcript in the seminiferous tubules. This alternative complementary DNA lacked the fifth exon, harboring the calpain cleavage site essential for generation of mature 17K IL-1alpha. This was verified by calpain treatment, producing the expected cleavage products of recombinant 32proIL 1alpha, but not of 24proIL-1alpha. Similarly, expression in COS-7 cells demonstrated processing of 32proIL-1alpha to the mature 17K form and secretion, whereas 24proIL-1alpha remained unprocessed. Both 32proIL-1alpha and 24proIL 1alpha showed a dose-dependent stimulatory effect in a thymocyte proliferation assay, although at lower potency than mature 17K IL-1alpha. In contrast, when tested on hCG-stimulated Leydig cells in vitro, a dose-dependent inhibition of testosterone production was obtained with mature 17K IL-1alpha and at a lower potency with 32proIL-1alpha, whereas 24proIL-1alpha was inactive. In conclusion, the three IL-1 bioactive proteins described here contribute to IL-1 protein heterogeneity and may serve as constitutive paracrine mediators in the testis. PMID- 11108250 TI - Chronic blockade of the melanocortin 4 receptor subtype leads to obesity independently of neuropeptide Y action, with no adverse effects on the gonadotropic and somatotropic axes. AB - Neuropeptide Y (NPY) is a powerful orexigenic factor, and alphaMSH is a melanocortin (MC) peptide that induces satiety by activating the MC4 receptor subtype. Genetic models with disruption of MC4 receptor signaling are associated with obesity. In the present study, a 7-day intracerebroventricular infusion to male rats of either the MC receptor antagonist SHU9119 or porcine NPY (10 nmol/day) was shown to strongly stimulate food and water intake and to markedly increase fat pad mass. Very high plasma leptin levels were found in NPY-treated rats (27.1 +/- 1.8 ng/ml compared with 9.9 +/- 0.9 ng/ml in SHU9119-treated animals and 2.1 +/- 0.2 ng/ml in controls). As expected, NPY infusion induced hypogonadism, characterized by an impressive decrease in seminal vesicle and prostate weights. No such effects were seen with the SHU9119 infusion. Similarly, whereas the somatotropic axis of NPY-treated rats was fully inhibited, this axis was normally activated in the obese SHU9119-treated rats. Chronic infusion of SHU9119 strikingly reduced hypothalamic gene expression for NPY (65.2 +/- 3.6% of controls), whereas gene expression for POMC was increased (170 +/- 19%). NPY infusion decreased hypothalamic gene expression for both POMC and NPY (70 +/- 9% and 75.4 +/- 9.5%, respectively). In summary, blockade of the MC4 receptor subtype by SHU9119 was able to generate an obesity syndrome with no apparent side effects on the reproductive and somatotropic axes. In this situation, it is unlikely that hyperphagia was driven by increased NPY release, because hypothalamic NPY gene expression was markedly reduced, suggesting that hyperphagia mainly resulted from loss of the satiety signal driven by MC peptides. NPY infusion produced hypogonadism and hyposomatotropism in the face of markedly elevated plasma leptin levels and an important reduction in hypothalamic POMC synthesis. In this situation NPY probably acted both by exacerbating food intake through Y receptors and by reducing the satiety signal driven by MC peptides. PMID- 11108251 TI - Allele-specific histone acetylation accompanies genomic imprinting of the insulin like growth factor II receptor gene. AB - The mouse insulin-like growth factor II receptor (Igf2r) gene encodes two reciprocally imprinted RNA transcripts: paternally imprinted Igf2r sense and maternally imprinted Igf2r antisense. Although DNA methylation has been implicated in the initiation and maintenance of genomic imprinting, acetylation of core histones has recently been appreciated as another important factor that regulates gene expression. To determine whether histone acetylation participates in the regulation of Igf2r imprinting, we examined the relative abundance of acetylated histones in interspecific mice (M. spretus x C57BL/6). Oligonucleosomes derived from liver were immunoprecipitated with acetyl-histone antiserum and were analyzed for the allelic distribution of DNA from the region of the sense and antisense Igf2r promoters. In nucleosomes associated with the Igf2r sense promoter, histone acetylation was demonstrated on the maternal allele, which is transcriptionally active. There was much less histone acetylation on the suppressed paternal allele. In nucleosomes associated with the Igf2r antisense promoter, the active paternal allele was heavily acetylated, whereas the suppressed maternal allele was underacetylated. Treatment of cultured fibroblasts with the histone deacetylase inhibitor Trichostatin A induces partial relaxation of genomic imprinting as well as decreased DNA methylation of both Igf2r sense and antisense promoters. These results demonstrate that increases in histone acetylation can lead to decreased DNA methylation, thereby modulating the regulation of the imprinted expression of Igf2r sense and antisense transcripts. PMID- 11108252 TI - Mice deficient in liver production of insulin-like growth factor I display sexual dimorphism in growth hormone-stimulated postnatal growth. AB - Insulin-like growth factor I (IGF-I) is essential for normal intrauterine and postnatal growth and development. Using the Cre/loxP-induced conditional knockout system, we have established a liver-specific IGF-I-deficient (LID) mouse model. Circulating IGF-I levels were decreased by approximately 75% without any apparent effect on their growth and development. To determine the role of extra-hepatic IGF-I in GH-induced postnatal growth, we tested the effects of GH on growth rates in these mice. Female, but not male, LID mice displayed significantly accelerated growth rates in response to daily injections of GH for 5 weeks. The GH-induced peripubertal growth in female LID mice was not affected by ovariectomy, nor did castration change the growth pattern in male LID mice. Thus, factors other than gonadal steroids mediate this sexual dimorphism. We postulate that the sexual dimorphic response to GH observed in LID mice may be related to genetically programmed differences in GH secretion patterns. PMID- 11108253 TI - Leptin deficiency induced by fasting impairs the satiety response to cholecystokinin. AB - Leptin administration potentiates the satiety response to signals such as cholecystokinin (CCK), that are released from the gut during a meal. To investigate the physiological relevance of this observation, we hypothesized that leptin deficiency, induced by fasting, attenuates the satiety response to CCK. To test this hypothesis, 48-h-fasted or fed rats were injected with i.p. saline or CCK. Fasting blunted the satiety response to 3.0 microg/kg CCK, such that 30-min food intake was suppressed by 65.1% (relative to saline-treated controls) in fasted rats vs. 85.9% in the fed state (P < 0.05). In a subsequent experiment, rats were divided into three groups: 1) vehicle/fed; 2) vehicle/fasted; and 3) leptin-replaced/fasted; and each group received 3.0 microg/kg i.p. CCK. As expected, the satiety response to CCK was attenuated by fasting in vehicle treated rats (30-min food intake: vehicle/fed, 0.3 +/- 0.1 g; vehicle/fasted, 1.7 +/- 0.4 g; P < 0.01), and this effect was prevented by leptin replacement (0.7 +/ 0.2 g, P < 0.05 vs. vehicle/fasted; P = not significant vs. vehicle/fed). To investigate whether elevated neuropeptide Y (NPY) signaling plays a role in the effect of leptin deficiency to impair the response to CCK, we measured the response to 3.0 microg/kg i.p. CCK after treatment with 7.5 microg intracerebroventricular NPY. We found that both CCK-induced satiety and its ability to increase c-Fos-like-immunoreactivity in key brainstem-feeding centers were attenuated by NPY pretreatment. We conclude that an attenuated response to meal-related satiety signals is triggered by leptin deficiency and may contribute to increased food intake. PMID- 11108254 TI - Requirement for follicle-stimulating hormone action in the formation of primordial follicles during perinatal ovarian development in the hamster. AB - Whereas FSH action is critical for the growth of preantral follicles, its role in the development of primordial follicles is controversial. The objective of the present study was to evaluate whether perinatal (fetal through early postnatal) FSH action is needed for the formation of primordial follicles, which first appear in the hamster ovary on the 7th to 8th day of postnatal life. A single dose of FSH-specific polyclonal antibody was injected into pregnant hamsters on the 12th, 13th, or 14th day of gestation and into newborn hamsters. Some of the antibody-exposed postnatal hamsters were injected with a single dose of equine CG (eCG) to check the reversibility of the antibody action. Ovaries were collected on D8pn or D12pn, and the percentage of primordial, primary, and secondary follicles was quantitated morphometrically. Ovaries of 8-day-old hamsters that were born to mothers treated with a single s.c. dose of the anti-FSH-antibody on day 12 of gestation had significantly reduced numbers of primordial follicles, compared with those treated with preimmune serum or saline (2.4% vs. 25%); however, the antibody inhibition was nearly completely reversed (approximately 18%) by a single injection of eCG on the first day of life. Delaying antibody treatment during late gestation caused a time-dependent block in granulosa cell differentiation, with a consequent proportional increase in the percentage of primordial follicles. This indicates that FSH-induction of primordial follicle development begins at a critical time of ovarian development. On the other hand, shortening the postnatal duration of eCG exposure reduced the degree of reversal, suggesting that prolonged perinatal FSH action is essential for developing the full gamut of primordial follicles. These results provide the first direct evidence that FSH action during fetal ovarian development is critical for the onset of primordial follicle formation. PMID- 11108255 TI - Lipopolysaccharide directly stimulates the intrapituitary interleukin-6 production by folliculostellate cells via specific receptors and the p38alpha mitogen-activated protein kinase/nuclear factor-kappaB pathway. AB - Bacterial lipopolysaccharide (LPS) activates the immune system and induces increases in peripheral cytokines, which, in turn, affect the endocrine system. In particular, LPS-induced cytokines stimulate the hypothalamic-pituitary-adrenal axis to increase levels of antiinflammatory-acting glucocorticoids. In the present work, we show that LPS directly stimulates interleukin (IL)-6 release by mouse pituitary folliculostellate (FS) TtT/GF tumor cells and FS cells of mouse pituitary cell cultures. The stimulatory effect of LPS was strongly enhanced in the presence of serum, suggesting that LPS is only fully active as a complex with LPS-binding protein (LBP). Both TtT/GF cells and mouse pituitaries expressed CD14, which binds the LPS/LBP complex, and Toll-like receptor type 4, which induces LPS signals. LPS increased phospoinositol turnover in TtT/GF cells and induced phosphorylation of p38alpha mitogen-activated protein kinase and the inhibitor (IkappaB) of nuclear factor-kappa B. Nuclear factor-kappa B was activated by LPS in TtT/GF cells. Functional studies demonstrated that My4 (an antibody blocking the interaction between LPS/LBP and CD14), SB203580, (a specific inhibitor of p38alpha mitogen-activated protein kinase phosphorylation), dexamethasone, and the messenger RNA translation inhibitor cycloheximide all inhibited LPS-induced IL-6 production by TtT/GF cells and mouse pituitary FS cells. LPS-induced intrapituitary IL-6 may modulate the function of anterior pituitary cells during bacterial infection/inflammation. PMID- 11108256 TI - Role of endogenous nociceptin in the regulation of arginine vasopressin release in conscious rats. AB - The effects of central administration of the opioid-like peptide nociceptin (also known as orphanin FQ) were investigated on the secretion of arginine vasopressin (AVP) in response to dehydration and hyperosmolar or hypovolemic stimulation in conscious rats. Intracerebroventricular (i.c.v.) administration of nociceptin suppressed plasma AVP concentration in a dose-dependent manner (0.1-10 microg/rat) in dehydrated rats, and the maximum effect was obtained 10 min after the administration (dehydration with 10 microg/rat nociceptin, 3.11 +/- 0.27 pg/ml vs. control, 10.32 +/- 0.96 pg/ml). The plasma AVP increase in response to either hyperosmolality [i.p. injection of hypertonic saline (HS) (600 mosml/kg)] or hypovolemia [i.p. injection of polyethylene glycol (PEG)] was also significantly blunted when nociceptin was injected i.c.v. (HS with 10 microg/rat nociceptin, 1.16 +/- 0.09 pg/ml vs. control, 1.82 +/- 0.30 pg/ml; PEG with 10 microg/rat nociceptin, 0.91 +/- 0.16 pg/ml vs. control, 2.41 +/- 0.26 pg/ml). Pretreatment with a selective opioid kappa-receptor antagonist, nor binaltorphimine (1 microg/ rat, i.c.v.) or naloxone (2.5 mg/rat, s.c. injection) did not reverse the inhibitory effects of nociceptin on AVP release. Moreover, when plasma AVP was suppressed by acute water loading, immunoneutralization of endogenous nociceptin by antinociceptin-antiserum i.c.v. significantly reversed the suppression (0.57 +/- 0.12 pg/ml vs. control, 0.25 +/- 0.04 pg/ml). These results suggest that central nociceptin is physiologically involved in the control of AVP release through an inhibitory action. PMID- 11108257 TI - Central, but not peripheral, glucose-sensing mechanisms mediate glucoprivic suppression of pulsatile luteinizing hormone secretion in the sheep. AB - Changes in glucose availability are proposed to modulate pulsatile GnRH secretion, and at least two anatomical sites, the liver and hindbrain, may serve as glucose sensors. The present study determined the relative importance of these putative glucose-sensing areas in regulating pulsatile LH secretion in the sheep. Our approach was to administer the antimetabolic glucose analog, 2-deoxy-D glucose (2DG) into either the hepatic portal vein or the fourth ventricle in gonadectomized females in which LH pulse frequency was high. In the first study, a catheter was placed in the ileocolic vein to determine the effects of local injection of 2DG into the hepatic portal system on the release of LH. After monitoring the pattern of LH secretion for 4 h, 2DG (250 mg/kg) was infused (500 microl/min) into the liver for 2 h. For comparison, animals were also given the same dose of 2DG into a jugular vein for 2 h. Administration of 2DG into either the hepatic portal or jugular vein reduced LH pulse frequency to the same extent. Infusion of the lower dose (50 mg/kg) locally into the hepatic portal vein did not affect plasma LH profiles. Collectively, these results are interpreted to indicate that the liver does not contain special glucose-sensing mechanisms for the glucoprivic suppression of LH pulses. In the second study, 2DG (5 mg/kg) was infused (50 l/min) for 30 min into the fourth ventricle or lateral ventricle. During the subsequent 4-h sampling period, pulsatile LH secretion was significantly suppressed, but there was no significant difference in LH pulse frequency between sites of infusion. Peripheral 2DG concentrations were not detectable after either fourth or lateral ventricle infusions, indicating that the 2DG had acted centrally to suppress LH pulses. Plasma cortisol concentrations increased more in animals infused with 2DG into the fourth ventricle than in those infused into the lateral ventricle, suggesting that 2DG infused into lateral ventricle is transported caudally into the fourth ventricle and acts within the area surrounding the fourth ventricle. Overall, these findings suggest that an important glucose-sensing mechanism is located circumventricularly in the fourth ventricle. Moreover, the liver does not appear to play an important role in detecting glucoprivic action of 2DG to suppress pulsatile LH secretion. PMID- 11108258 TI - Oxytocin stimulates the translocation of oxytocinase of human vascular endothelial cells via activation of oxytocin receptors. AB - Oxytocinase (OTase) degrades several small peptides such as oxytocin (OT), and thus plays important roles in fetal development and maintenance of human homeostasis during pregnancy. The physiological effects of OT are mediated via its receptor (OTR). Although the interactions between OT and OTR have studied extensively, the relationship to OTase remains to be clarified. It is known that human umbilical vascular endothelial cells express OTR messenger RNA; therefore, they were selected for examination of this question in the present study. RT-PCR experiments confirmed the existence of messenger RNA for OTase, and assessment of protein levels and activity clarified that OT increases the activity of OTase at the cell surface via binding to OTR. This stimulation appears to involve translocation of OTase from cytosolic to the cell surface in response to cellular signal transduction pathways linked to the OTR. Protein kinase C stimulation significantly increased the cell surface activity of OTase, whereas its inhibition resulted in reduction. In summary, our findings provide clear evidence that OT triggers directly OTase translocation in human umbilical vascular endothelial cells via a protein kinase C-dependent pathway coupled to OTR. PMID- 11108259 TI - Luteinizing hormone-releasing hormone (LHRH) biosynthesis and secretion in embryonic LHRH. AB - Evidence indicates that LH-releasing hormone (LHRH) neurons can exhibit neuroendocrine secretory properties before entrance into the central nervous system. In this study, we evaluated LHRH biosynthesis and secretion in embryonic LHRH neurons maintained in nasal explants. Using ELISA and calcium imaging techniques, peptide content and single neuron activities were examined. LHRH neurons maintained for 7-10 days in vitro were found to possess a similar amount of LHRH/cell as the equivalent aged LHRH cells in vivo (postnatal day 1). LHRH peptide was measured in the medium of these relatively young cultures, and 40 mM KCl stimulated a 4-fold increase in LHRH secretion. KCl enhanced medium also resulted in a significant increase in LHRH content per culture (24.5 pg vs. 32.3). A similar effect was observed after muscimol-enhanced media (32.2 pg). Both agents also stimulated a substantial rise in intracellular calcium. Pretreatment of cultures with tetrodotoxin partially blocked the affects of muscimol on both peptide content and calcium activity, but not KCl. Calcium depleted medium blocked the effects of KCl yet only attenuated the effects of muscimol. Treatment of cultures with cycloheximide blocked the effects of both muscimol and KCl. These results indicate that developing LHRH neurons are capable of synthesizing, secreting, and rapidly replenishing stores of LHRH peptide. PMID- 11108260 TI - Adrenergic regulation of mitogen-activated protein kinase in rat pinealocytes: opposing effects of protein kinase A and protein kinase G. AB - The role of adrenergic stimulation in the regulation of mitogen-activated protein kinase (MAPK) in rat pinealocytes was investigated by measuring phosphorylated MAPK using Western blot analysis and a MAPK enzymatic assay. Stimulation with the endogenous neurotransmitter, norepinephrine (NE; a mixed alpha- and beta adrenergic agonist), concentration dependently increased the phosphorylation of both p44 and p42 isoforms of MAPK. This effect of NE was blocked by PD98059 and U0126 (two inhibitors of MEK). Treatment with prazosin or propranolol significantly reduced the effect of NE on MAPK phosphorylation, suggesting the involvement of both alpha- and beta-adrenergic receptors. Investigation into the intracellular mechanisms of NE action revealed that the increase in MAPK phosphorylation was blocked by KT5823 (a protein kinase G inhibitor), but was enhanced by H89 (a protein kinase A inhibitor). Calphostin C (a protein kinase C inhibitor) and KN93 (a Ca2+/calmodulin-dependent protein kinase inhibitor) also attenuated NE-mediated MAPK activation, but to a lesser degree. Furthermore, inhibition of MAPK phosphorylation by (Bu)2cAMP was effective in reducing MAPK activation by (Bu)2cGMP, an active phorbol ester or ionomycin. These results indicate that the effect of NE on MAPK phosphorylation represents mainly the integration of two signaling mechanisms, protein kinase A and protein kinase G, each having an opposite effect on MAPK phosphorylation. PMID- 11108261 TI - A role for Akt in mediating the estrogenic functions of epidermal growth factor and insulin-like growth factor I. AB - This study examines whether the serine/threonine protein kinase, Akt, is involved in the cross-talk between epidermal growth factor (EGF) and insulin-related growth factor I (IGF-I) receptors and ER-alpha. Treatment of MCF-7 cells with either EGF or IGF-I resulted in a rapid phosphorylation of Akt and a 14- to 16 fold increase in Akt activity, respectively. Akt activation was blocked by inhibitors of phosphatidylinositol 3-kinase, but not by an inhibitor of the ribosomal protein kinase p70S6K. Stable transfection of cells with a dominant negative Akt mutant blocked the effects of EGF and IGF-I on ER-alpha expression and activity, whereas stable transfection of cells with a constitutively active Akt mutant mimicked the effects of EGF and IGF-I. In the latter cells, there was a decrease in the amount of ER-alpha protein and messenger RNA (70-80%) and an increase in the amount of progesterone receptor protein, messenger RNA (4- to 9- and by 3.5- to 7-fold, respectively) and pS2 (3- to 5-fold). Coexpression of wild type ER-alpha and the dominant negative Akt mutant in COS-1 cells also blocked the growth factor-stimulated activation of ER-alpha, but coexpression of the wild type receptor with the constitutively active Akt mutant increased ER-alpha activity. Receptor activation was blocked by an antiestrogen. Studies using mutants of ER-alpha demonstrated that Akt increased estrogen receptor activity through the amino-terminal activation function-1 (AF-1). Serines S104 S106, S118, and S167 appear to play a role in the activation of ER-alpha by Akt. PMID- 11108262 TI - Exposure of newborn male and female rats to environmental estrogens: delayed and sustained hyperprolactinemia and alterations in estrogen receptor expression. AB - Environmental estrogens (xenoestrogens) are synthetic compounds that are abundant in the environment and mimic natural estrogens. The estrogenicity of two such compounds, bisphenol A (BPA) and octylphenol (OP), during development of the neuroendocrine system was investigated. The objective was to compare the effects of neonatal exposure to BPA, OP, and diethylstilbestrol (DES), a potent synthetic estrogen, on prepubertal serum PRL levels and estrogen receptor (ER) expression in the anterior pituitary and medial basal hypothalamus. Receptor expression in the uterus and prostate, two peripheral estrogen-responsive tissues, was also examined. Newborn male and female Fischer 344 rats were s.c. injected on days 1-5 after birth with corn oil (control), BPA and OP (100 or 500 microg/day), or DES (5 microg/day). Rats were bled on days 15, 20, and 25 and on the day of death (day 30), and serum PRL was analyzed by RIA. Relative expressions of ERalpha and ERbeta were determined by RT-PCR. BPA and OP induced delayed, but progressive, increases in serum PRL levels, up to 3-fold above control levels, in both males and females. The low dose of either compound was equally or more effective as the high dose in eliciting and sustaining elevated serum PRL levels, namely hyperprolactinemia. In contrast, the DES treatment resulted in a transient rise in serum PRL levels. BPA, OP, and, to a lesser extent, DES increased the expression of both ERalpha and ERbeta in the anterior pituitary of males, but not females, whereas the hypothalamic ERs were less responsive to these compounds. DES treatment caused down-regulation of ERalpha expression in the uterus and up regulation of ERbeta in the prostate, whereas BPA or OP was without effect. In conclusion, exposure of newborn rats of either sex to environmental estrogens results in delayed and sustained hyperprolactinemia and differential alterations in ER expression in the hypothalamus and pituitary. DES appears to target the lower reproductive tract more effectively than the neuroendocrine system. PMID- 11108263 TI - Luteinizing hormone receptors are self-associated in the plasma membrane. AB - We have evaluated rat LH receptor self-association and lateral dynamics for functional and nonfunctional receptors after binding of hormone. We demonstrate, for the first time, that grouped receptors observed in electron or light microscopy represent actual receptor dimers or oligomers rather than simply the concentration of receptors within membrane microdomains. Fringe fluorescence photobleaching recovery methods showed that, after binding of either LH or human CG (hCG), functional wild-type LH receptors, expressed on 293 cells (LHR-wt cells), have mobilities that are 25% lower than those of nonfunctional LH receptors containing an arginine substitution for lysine at position 583 (LHR K583R cells). Because lateral diffusion coefficients in two dimensions depend only on the logarithm of the molecular size of the diffusing species, this result implies that functional receptors exist in substantially larger membrane complexes than do nonfunctional receptors. In single-cell measurements of fluorescence energy transfer after hormone binding, functional LH receptors were also characterized by receptor self-aggregation. Values for fluorescence resonant energy transfer efficiency were 13 +/- 2% and 17 +/- 3% between fluorophore conjugated LH or hCG, respectively, bound to receptors on LHR-wt cells. However, there was little or no energy transfer between receptors on LHR-K583R cells. These results suggest that receptor functionality involves receptor-receptor interactions and that the extent of such receptor self-association depends on whether LH or hCG binds the receptor. PMID- 11108264 TI - The melanin-concentrating hormone receptor couples to multiple G proteins to activate diverse intracellular signaling pathways. AB - The receptor for melanin-concentrating hormone (MCH) was recently identified as the orphan G protein-coupled receptor SLC-1. In this study, a CHO cell line expressing the MCH receptor (Kd = 1.3 nM; binding capacity, 3.6 pmol/mg protein) is used to assess the ability of the MCH receptor to couple to Gi, Go, and Gq proteins. The results demonstrate that MCH inhibits forskolin-stimulated cAMP production in a pertussis toxin- (PTX)-sensitive manner in CHO-MCHR cells (EC50 = 100 pM), indicating that the MCH receptor couples to one or more members of the Gi subfamily of G proteins. In addition, MCH stimulates increases in phosphoinositide metabolism (EC50 = 50 nM) and in intracellular free Ca2+ levels (EC50 = 10 nM). MCH-stimulated inositol phosphate production and increases in intracellular free Ca2+ are partially inhibited (60% and 40%, respectively) by PTX pretreatment, demonstrating that there are at least two components of each of these signaling pathways. One component is PTX sensitive and therefore mediated through a Gi/Go protein. A distinct G protein-coupled (probably Gq type) mediates the PTX-insensitive component. To distinguish Gi vs. Go coupling, MCH-stimulated mitogen-activated protein (MAP) kinase activity was examined. Gi and Go use separate signaling pathways to mediate MAP kinase activation in CHOcells. Protein kinase C (PKC) activity is essential in the Go-dependent MAP kinase signaling pathway, but is not required in the GC-dependent MAP kinase signaling pathway. MCH stimulated MAP kinase activity is decreased (50%), but not abolished, by inhibition of PKC activity or depletion of cellular PKC, indicating that MCH stimulated MAP kinase activity is mediated through both Gi- and Go-dependent signaling mechanisms. The results of this study are the first to clearly demonstrate that the MCH receptor couples to multiple G proteins to mediate several diverse intracellular signaling pathways. PMID- 11108265 TI - ADAMTS-1: A cellular disintegrin and metalloprotease with thrombospondin motifs is a target for parathyroid hormone in bone. AB - PTH stimulates bone formation in animals and humans, and the expressions of a number of genes have been implicated in the mediation of this effect. To discover new bone factors that initiate and support this phenomenon we used differential display RT-PCR and screened for genes that are selectively expressed in osteoblast-enriched femoral metaphyseal primary spongiosa of young male rats after a single s.c. injection of human PTH-(1-38) (8 microg/100 g). We show that one of the messenger RNAs that is up-regulated in bone is ADAMTS-1, a new member of the ADAM (A disintegrin and metalloprotease) gene family containing thrombospondin type I motifs. ADAMTS-1 consists of multiple domains common to ADAM family of proteins, including pro-, metalloprotease-like, and disintegrin like domains. However, unlike other ADAMs, ADAMTS-1 does not possess a transmembrane or cytoplasmic domain and is a secreted protein. Northern blot analysis confirmed that ADAMTS-1 was up-regulated in both metaphyseal (14- to 35 fold) and diaphyseal (4.2-fold) bone 1 h after PTH-(1-38) injection and returned to control levels by 24 h. We also analyzed the regulation of ADAMTS-1 in response to various PTH/PTH-related peptide (PTHrP) analogs and found that PTH-(1 31) and PTHrP-(1-34), which activate the protein kinase A (PKA) pathway, induce ADAMTS-1 expression 1 h after injection, whereas PTH-(3-34) and PTH-(7-34), which do not activate the PKA pathway, did not regulate expression. To investigate the effect of other osteotropic agents, we analyzed ADAMTS-1 expression after a single dose of PGE2 (6 mg/kg) and found that it was up-regulated 1 h after injection and returned to control levels by 6 h. In vitro ADAMTS-1 is expressed in primary osteoblasts and osteoblastic cell lines, but was not detectable in osteoclasts generated from macrophage colony-stimulating factor/receptor activator of NF-kappaB ligand/transforming growth factor-beta1-treated bone marrow cells. Treatment of UMR 106 osteosarcoma cells with PTH, PGE2, forskolin, or (Bu)2cAMP increased ADAMTS-1 expression 7-, 4-, 5-, and 5-fold, respectively. Also, in vitro treatment with 1alpha,25-dihydroxyvitamin D3 increased ADAMTS-1 expression 3-fold. Tissue distribution analysis showed that ADAMTS-1 is expressed at high levels in many tissues, including the heart, lung, liver, skeletal muscle, and kidney. Taken together, these results demonstrate that ADAMTS-1 is specifically up-regulated in bone and osteoblasts by the osteotropic agents PTH, PTHrP, and PGE2 possibly via the cAMP/PKA pathway. We speculate that the rapid and transient increase in ADAMTS-1 expression may contribute to some of the effects of PTH on bone turnover. PMID- 11108266 TI - A comparative study of hormonal regulation of three secretory proteins (prostatic secretory protein-PSP94, probasin, and seminal vesicle secretion II) in rat lateral prostate. AB - The rat dorsolateral prostate secretes several major known proteins, although their physiological and reproductive functions are largely undefined. In the present study we examined and compared the in vivo hormonal regulation of the messenger RNA (mRNA) expression of three major secretory proteins, including prostatic secretory protein of 94 amino acids (PSP94 or beta-microseminoprotein), probasin, and seminal vesicle secretion II (SVSII), in long-term castrated lateral prostates (LP) by in situ hybridization and semiquantitative RT-PCR. The protein levels of PSP94 in the castrated LPs were also examined by Western blotting. PSP94 is a small protein newly isolated from the rat prostate gland and demonstrates highly specific expression in the LP. The results of in situ hybridization showed that PSP94, probasin, and SVSII were highly expressed in the intact LP. The hybridization signals of probasin and PSP94 disappeared in the 60 day postcastrated LPs, whereas the signals of SVSII dropped sharply in the 14-day postcastrated LPs. Similar patterns of decreasing mRNA levels of the three proteins in the castrated LPs were observed by RT-PCR analysis. Their mRNA transcripts were restored to normal levels after replacement with testosterone. The results indicate that these secretory proteins are all under androgen regulation in the rat LP. Interestingly, we also observed that their degrees of sensitivity or responsiveness to androgen withdrawal are different. Their mRNA levels dropped in response to duration of castration in the following decreasing order: SVSII, PSP94, and probasin. Besides androgen [dihydrotestosterone (DHT)], we also examined the effects of glucocorticoid [dexamethasone (DEX)], progestin [medroxyprogesterone acetate (MPA)], and zinc on their gene expressions in castrated LPs. We observed that the mRNA transcripts of both PSP94 and probasin were increased after treatments with DHT, DEX, and MPA, suggesting that these two proteins could also be regulated by glucocorticoid and progestin. In contrast with probasin, PSP94 and SVSII were not induced by ZnSO4 treatment. On the other hand, SVSII expression was only increased significantly by DHT and moderately by MPA, but not by DEX, suggesting that SVSII is under strict control by androgen. PMID- 11108267 TI - Expression of cyclin-dependent kinase inhibitors in epiphyseal chondrocytes induced to terminally differentiate with thyroid hormone. AB - A growing body of evidence suggests that systemic hormones and peptide growth factors may exert their effects on cell growth and differentiation in part through regulation of the cell division cycle. We hypothesized that thyroid hormone regulates terminal differentiation of growth plate chondrocytes in part through controlling cell cycle progression at the G1/S restriction point. Our results support this hypothesis by demonstrating that treatment of epiphyseal chondrocytes with thyroid hormone under chemically defined conditions results in the arrest of DNA synthesis and the onset of terminal differentiation, indicating that thyroid hormone is one factor capable of regulating the transition between cell growth and differentiation in these cells. This terminal differentiation process is associated with induction of the cyclin/cyclin-dependent kinase inhibitors p21(cip-1 waf-1) and p27kip1, suggesting that thyroid hormone may regulate terminal differentiation in part by arresting cell cycle progression through induction of cyclin-dependent kinase inhibitors. PMID- 11108268 TI - Bone morphogenetic proteins induce gremlin, a protein that limits their activity in osteoblasts. AB - Bone morphogenetic proteins (BMP) induce the differentiation of cells of the osteoblastic lineage and enhance the function of the osteoblast. Growth factor activity is regulated by binding proteins, and we previously showed that BMPs induce noggin, a glycoprotein that binds and blocks BMP action. Recently, additional BMP antagonists, such as gremlin, have been described, but there is no information about their expression or function in osteoblasts. We tested for the expression of gremlin and studied its induction by BMPs in cultures of osteoblast enriched cells from 22-day-old fetal rat calvariae (Ob cells). BMP-2 caused a time- and dose-dependent increase in gremlin messenger RNA and polypeptide levels, as determined by Northern and Western blot analyses. The effects of BMP-2 on gremlin transcripts were independent of new protein synthesis. BMP-2 increased the rate of gremlin transcription as determined by nuclear run-on assays. Fibroblast growth factor-2 and platelet-derived growth factor BB also induced gremlin, but other hormones and growth factors had no effect. Gremlin prevented the stimulatory effects of BMP-2 on DNA, collagen, noncollagen protein synthesis, and alkaline phosphatase activity in Ob cells. In conclusion, BMPs induce gremlin transcription in Ob cells, a mechanism that probably limits BMP action in osteoblasts. PMID- 11108269 TI - Desensitization and internalization of human and xenopus gonadotropin-releasing hormone receptors expressed in alphaT4 pituitary cells using recombinant adenovirus. AB - Nonmammalian vertebrates express at least two forms of GnRH and distinct forms of GnRH receptor (GnRH-R) have coevolved with their ligands. Mammalian and nonmammalian GnRH-R have key structural differences (notably the lack of C terminal tails in mammalian GnRH-R) and comparative studies are beginning to reveal their functional relevance. However, cellular context and receptor density influence G protein-coupled receptor function and may be important variables in such work using heterologous expression systems. Here we report a comparative study using alphaT4 cells (gonadotrope progenitors that lack endogenous GnRH-R) transfected with a mammalian (human) or nonmammalian (Xenopus laevis type I) GnRH R. Because conventional transfection strategies proved inefficient, recombinant adenovirus expressing these receptors were constructed, enabling controlled and efficient GnRH-R expression. When expressed in alphaT4 cells at physiological density, these GnRH-Rs retain the pharmacology of their endogenous counterparts (as judged by ligand specificity in radioligand binding and inositol phosphate accumulation assays) but do not activate adenylyl cyclase and are not constitutively active. Moreover, the Xenopus GnRH-R rapidly desensitizes and internalizes in these cells, whereas the human GnRH-R does not, and the internalization rates are not dependent upon receptor number. These data extend studies in COS, HEK, and GH3 cells showing that other GnRH-R with C-terminal tails desensitize and internalize rapidly, whereas tail-less mammalian GnRH-R do not. Retention of these distinctions at physiological receptor density in gonadotrope lineage cells, supports the argument that the evolution of nondesensitizing mammalian GnRH-Rs is functionally relevant and related to the development of mammalian reproductive strategies. PMID- 11108270 TI - Role of leptin in peroxisome proliferator-activated receptor gamma coactivator-1 expression. AB - Peroxisome proliferator-activated receptor-gamma coactivator-1 (PGC-1), a cold induced protein expressed in brown adipose tissue (BAT), plays a role in adaptive thermogenesis by up-regulating uncoupling proteins (UCP). Here, we explore its relationship to the thermogenic actions of leptin, which also up-regulates UCPs. We find that PGC-1 messenger RNA (mRNA) is markedly reduced in BAT of obese leptin-deficient (ob/ob mice) and leptin-unresponsive (db/db mice and Zucker diabetic fatty fa/fa rats) rodents. Whereas, after cold exposure (6 C for 7 h), PGC-1 mRNA increases 2.6-fold in BAT of lean +/+ rats, it rises only 30% in fa/fa rats. Four days after induction of hyperleptinemia (>30 ng/ml) in Wistar rats, by adenovirus gene transfer, PGC-1 mRNA in BAT was 2.3-fold and UCP-1, 4-fold above controls. In isolated white adipocytes, PGC-1 mRNA increased 4.4-fold within 6 h of incubation with 20 ng/ml of leptin. We conclude that leptin action is required for normal basal and cold-stimulated PGC-1 expression in BAT in rodents and that hyperleptinemia rapidly up-regulates its expression, at least in part, by direct action. PMID- 11108271 TI - Insulin-Like growth factor I (IGF-I) and cyclic adenosine 3',5'-monophosphate regulate IGF-binding protein-3 gene expression by transcriptional and posttranscriptional mechanisms in mammary epithelial cells. AB - Insulin-like growth factor I (IGF-I) is a potent mitogen for both normal and transformed mammary epithelial cells (MEC), and IGF-binding protein-3 (IGFBP-3) potentiates IGF-I action in these cells. The synthesis of IGFBP-3 is stimulated by both IGF-I and agents that increase intracellular cAMP (e.g. forskolin) in the bovine MEC line MAC-T. In addition, the combination of IGF-I and cAMP increases IGFBP-3 messenger RNA to a greater extent than does either treatment alone. The molecular mechanisms responsible for this regulation are not known and therefore represent the focus of this study. The half-life of IGFBP-3 messenger RNA in untreated MAC-T cells was determined to be 11 h. Exposure to IGF-I or forskolin increased the half-life to 27 and 101 h, respectively. Nuclear run-on assays indicated that IGFBP-3 transcription rates were increased 3.5 +/- 0.83-fold (n = 4) in cells treated with a combination of IGF-I and forskolin. To further study this regulation, 1.1 kb of the 5'-flanking region of the IGFBP-3 promoter were fused to a promoterless reporter plasmid encoding luciferase. Transient transfection assays indicated that both IGF-I and forskolin alone produced small, but significant, increases in IGFBP-3 promoter activity of 1.57 +/- 0.12 and 1.59 +/- 0.08-fold (P < 0.01), respectively (mean +/- SE; n = 7). However, the combination of IGF-I and forskolin increased IGFBP-3 promoter activity 2.25 +/- 0.14-fold above control values (P < 0.01), suggesting that these factors activate discrete signaling pathways that act in concert to stimulate IGFBP-3 gene transcription. Deletion analysis indicated that promoter fragments containing as little as 267 bp upstream of the TATA box retained responsiveness to IGF-I and forskolin. This region contains a 200-bp sequence that is approximately 80% homologous between the murine and bovine promoters. It contains several conserved AP-2 and Sp1 consensus binding sequences that may be important for the effects of IGF-I and forskolin on IGFBP-3 promoter activity. In summary, these data indicate that IGF-I and cAMP, working through separate signaling pathways, activate both transcriptional and post-transcriptional mechanisms to stimulate IGFBP-3 synthesis in MEC. PMID- 11108272 TI - Growth hormone improves bioenergetics and decreases catecholamines in postinfarct rat hearts. AB - The aims of this study were to examine, in vivo, the effects of GH treatment on myocardial energy metabolism, function, morphology, and neurohormonal status in rats during the early postinfarct remodeling phase. Myocardial infarction (MI) was induced in male Sprague Dawley rats. Three different groups were studied: MI rats treated with saline (n = 7), MI rats treated with GH (MI + GH; n = 11; 3 mg/kg x day), and sham-operated rats (sham; n = 8). All rats were investigated with 31P magnetic resonance spectroscopy and echocardiography at 3 days after MI and 3 weeks later. After 3 weeks treatment with GH, the phosphocreatine/ATP ratio increased significantly, compared with the control group (MI = 1.69 +/- 0.09 vs. MI + GH = 2.42 +/- 0.05, P < 0.001; sham = 2.34 +/- 0.08). Treatment with GH significantly attenuated an increase in left ventricular end systolic volume and end diastolic volume. A decrease in ejection fraction was prevented in GH-treated rats (P < 0.05 vs. MI). Myocardial and plasma noradrenaline levels were significantly lower in MI rats treated with GH. These effects were accompanied by normalization of plasma brain natriuretic peptide levels (sham = 124.1 +/- 8.4; MI = 203.9 +/- 34.7; MI + GH = 118.3 +/- 8.4 ng/ml; P < 0.05 vs. MI). In conclusion, GH improves myocardial energy reserve, preserves left ventricular function, and attenuates pathologic postinfarct remodeling in the absence of induction of left ventricular hypertrophy in postinfarct rats. The marked decrease in myocardial content of noradrenaline, after GH treatment, may protect myocardium from adverse effects of catecholamines during postinfarct remodeling. PMID- 11108273 TI - Glucagon-like peptide-1 induces cell proliferation and pancreatic-duodenum homeobox-1 expression and increases endocrine cell mass in the pancreas of old, glucose-intolerant rats. AB - Glucose homeostasis in mammals is maintained by insulin secretion from the beta cells of the islets of Langerhans. Type 2 diabetes results either from primary beta-cell failure alone and/or a failure to secrete enough insulin to overcome insulin resistance. Here, we show that continuous infusion of glucagon-like peptide-1 (7-36) (GLP-1; an insulinotropic agent), to young and old animals, had effects on the beta-cell of the pancreas other than simply on the insulin secretory apparatus. Our previous studies on a rodent model of glucose intolerance, the aging Wistar rat, show that a plateau in islet size, insulin content, and beta-cell mass is reached at 13 months, despite a continuing increase in body weight. Continuous sc infusion of GLP-1 (1.5 pM/kg x min), over 5 days, resulted in normal glucose tolerance. Our current results in both young and old rats demonstrate that treatment caused an up-regulation of pancreatic duodenum homeobox-1 (PDX-1) expression in islets and total pancreas, induced pancreatic cell proliferation, and beta-cell neogenesis. The effects on levels of PDX-1 messenger RNA were abrogated by simultaneous infusion of Exendin (9-39), a specific antagonist of GLP-1. PDX-1 protein levels increased 4-fold in whole pancreata and 6-fold in islets in response to treatment. Beta-cell mass increased to 7.2 +/- 0.58 from 4.88 +/- 0.38 mg, treated vs. control, respectively, P < 0.02. Total pancreatic insulin content also increased from 0.55 +/- 0.02 to 1.32 +/- 0.11 microg/mg total pancreatic protein. Therefore, GLP-1 would seem to be a unique therapy that can stimulate pancreatic cell proliferation and beta-cell differentiation in the pancreas of rodents. PMID- 11108274 TI - The role of selenocysteine 133 in catalysis by the human type 2 iodothyronine deiodinase. AB - Human type 2 iodothyronine deiodinase (hD2) catalyzes the activation of T4 to T3. D2, like types 1 and 3 deiodinases, contains selenocysteine (Sec) in the highly conserved active center at position 133. To evaluate the contribution of Sec133 to the catalytic properties of hD2, we generated mutants in which cysteine (Cys) or alanine (Ala) replaced Sec133. The Km (T4) of Cys133D2 was 2.1 microM, strikingly higher than that of native D2 (1.4 nM). In contrast, the relative turnover number was 10-fold lower for Cys133D2, illustrating the greater potency of Se than S in supporting catalysis. The AlaD2 mutant was inactive. Studies in intact cells transiently expressing the native or Cys133D2 enzyme exhibited saturation kinetics expected from the Km as measured under in vitro conditions, indicating rapid equilibration of extracellular and intracellular T4. Blockade of the NTCP, OATP1-3, and LST-1 transporters with 10 mM sodium taurocholate did not alter the deiodination rate of T4 by Cys133D2 transiently expressed in intact cells, suggesting that intracellular transport of T4 is not rate limiting. These results illustrate that selenium plays a critical role in deiodination catalyzed by hD2. PMID- 11108275 TI - The tetrabasic KKKK(147-150) motif determines intracrine regulatory effects of PthrP 1-173 on chondrocyte PPi metabolism and matrix synthesis. AB - Expression of PTHrP is a major regulator of growth cartilage development and also becomes robust in osteoarthritic cartilage. We further defined how PTHrP 1-173, which we observed to be the preferentially expressed PTHrP isoform in normal and osteoarthritic cartilage, functions in chondrocytes. We transfected both immortalized human juvenile costal chondrocytes (TC28 cells) and rabbit articular chondrocytes with wild-type PTHrP 1-173 and mutants of putative PTHrP 1-173 endoproteolytic processing sites. Wild-type PTHrP 1-173 inhibited collagen synthesis and decreased extracellular PPi (which critically regulates hydroxyapatite deposition) by 50-80% in both chondrocytic cell types. In contrast, PTHrP 1-173 mutated at the PTHrP 147-150 motif KKKK (but not the other site-directed mutants) and increased both extracellular PPi and collagen synthesis by >50%. Synthetic PTHrP 140-173 mutated at amino acids 147-150 and also increased extracellular PPi, and wild-type 140-173 decreased extracellular PPi in permeabilized cells. The 147-nuclear localization of PTHrP. We conclude that the tetrabasic 147-150 motif functions to determine how PTHrP 1-173 regulates collagen synthesis and levels of extracellular PPi by an intracrine mechanism in chondrocytes, and it may prove useful as a therapeutic target for regulation of mineralization. PMID- 11108277 TI - Estradiol attenuates angiotensin-induced aldosterone secretion in ovariectomized rats. AB - Estrogen replacement therapy significantly reduces the risk of cardiovascular disease in postmenopausal women. Previous studies indicate that estradiol (E2) decreases angiotensin II (AT) receptor density in the adrenal and pituitary in NaCl-loaded rats. We used an in vivo model that eliminates the potentially confounding influence of ACTH to determine whether the E2-induced decrease in adrenal AT receptor expression affects aldosterone responses to angiotensin II (Ang II). Female rats were ovariectomized, treated with oil (OVX) or E2 (OVX+E2; 10 microg, s.c.) for 14 days, and fed a NaCl-deficient diet for the last 7 days to maximize adrenal AT receptor expression and responsiveness. On days 12-14 rats were treated with dexamethasone (DEX; 25 microg, i.p., every 12 h) to suppress plasma ACTH. On day 14 aldosterone secretion was measured after a 30-min infusion of Ang II (330 ng/min). Ang II infusion increased the peak plasma aldosterone levels to a lesser degree in the OVX+E2 than in the OVX rats (OVX, 1870 +/- 290 pg/ml; OVX+E2, 1010 +/- 86 pg/ml; P < 0.05). Ang II-induced ACTH and aldosterone secretion was also studied in rats that were not treated with DEX. In the absence of DEX, the peak plasma aldosterone response was also significantly decreased (OVX, 5360 +/- 1200 pg/ml; OVX+E2, 2960 +/- 570 pg/ml; P < 0.05). However, E2 also reduced the plasma ACTH response to Ang II (P < 0.05; OVX, 220 +/- 29 pg/ml; OVX+E2, 160 +/- 20 pg/ml), suggesting that reduced pituitary ACTH responsiveness to Ang II contributes to the effect of E2 on Ang II-induced aldosterone secretion. Adrenal AT1 binding studies confirmed that E2 significantly reduces adrenal AT1 receptor expression in both the presence and absence of DEX in NaCl deprived rats. These results indicate that E2-induced decreases in pituitary and adrenal AT1 receptor expression are associated with attenuated pituitary ACTH and adrenal aldosterone responses to Ang II and suggest that estrogen replacement therapy may modulate Ang II-stimulated aldosterone secretion as part of its well known cardioprotective actions. PMID- 11108276 TI - The acute and chronic stimulatory effects of endothelin-1 on glucose transport are mediated by distinct pathways in 3T3-L1 adipocytes. AB - We have recently shown that pretreatment with endothelin-1 (ET-1) for 20 min stimulates GLUT4 translocation in a PI3-kinase-dependent manner in 3T3-L1 adipocytes (Imamura, T. et al., J Biol Chem 274:33691-33695). This study presents another pathway by which ET-1 potentiates glucose transport in 3T3-L1 adipocytes. ET-1 treatment (10 nM) leads to approximately 2.5-fold stimulation of 2 deoxyglucose (2-DOG) uptake within 20 min, reaching a maximal effect of approximately 4-fold at approximately 6 h, and recovering almost to basal levels after 24 h. Insulin treatment (3 ng/ml) results in an approximately 5-fold increase in 2-DOG uptake at 1 h, and recovering to basal levels after 24 h. The ETA receptor antagonist, BQ 610, inhibited ET-1 induced glucose uptake both at 20 min and 6 h, whereas the ETB receptor antagonist, BQ 788, was without effect. Interestingly, ET-1 stimulated 2-DOG uptake at 6 h, not at 20 min, was almost completely blocked by the protein-synthesis inhibitor, cycloheximide and the RNA synthesis inhibitor, actinomycin D, suggesting that the short-term (20 min) and long-term (6 h) effects of ET-1 involve distinct mechanisms. GLUT4 translocation assay showed that 20 min, but not 6 h, exposure to ET-1 led to GLUT4 translocation to the plasma membrane. In contrast, 6 h, but not 20 min, exposure to ET-1 increased expression of the GLUT1 protein, without affecting expression of GLUT4 protein. ET-1 induced 2-DOG uptake and GLUT1 expression at 6 h were completely inhibited by the MEK inhibitor, PD 98059, and partially inhibited by the PI3-kinase inhibitor, LY 294002, and the G alpha i inhibitor, pertussis toxin. The PLC inhibitor, U 73122, was without effect. These findings suggest that ET-1 induced GLUT1 protein expression is primarily mediated via MAPK, and partially via PI3K in 3T3-L1 adipocytes. PMID- 11108278 TI - High levels of glucose stimulate angiotensinogen gene expression via the P38 mitogen-activated protein kinase pathway in rat kidney proximal tubular cells. AB - The present studies investigated whether the effect of high levels of glucose on angiotensinogen (ANG) secretion and gene expression in kidney proximal tubular cells is mediated at least in part via the activation of p38 mitogen-activated protein kinase (p38 MAPK). Rat immortalized renal proximal tubular cells (IRPTCs) were cultured in monolayer. The levels of immunoreactive rat ANG (IR-rANG) secreted into the medium and the levels of cellular ANG messenger RNA were determined by a specific RIA for rat ANG and a RT-PCR assay, respectively. Phosphorylation of cellular p38 MAPK was determined by Western blot analysis using the Phospho Plus p38 MAPK antibody kit. High levels of glucose (i.e. 25 mM) and phorbol 12-myristate 13-acetate (PMA; 10(-7) M) increased the secretion of IR rANG and cellular ANG messenger RNA as well as phosphorylation of p38 MAPK in IRPTCs. This stimulatory effect of high levels of glucose and PMA was blocked by SB 203580 (a specific inhibitor of p38 MAPK), but not by SB 202474 (a negative control of SB 203580). High levels of D-sorbitol or 2-deoxy-D-glucose (i.e. > or = 35 mM) also stimulated the phosphorylation of p38 MAPK, but did not stimulate ANG secretion or gene expression. GF 109203X (an inhibitor of protein kinase C) blocked the stimulatory effect of high levels of glucose and PMA on ANG gene expression, whereas it did not block the effect of high levels of glucose, sorbitol, or 2-deoxy-D-glucose on p38 MAPK phosphorylation in IRPTCs. These studies demonstrate that the stimulatory effect of a high level of glucose (25 mM) on ANG gene expression in IRPTCS may be mediated at least in part via activation of p38 MAPK signal transduction pathway and is protein kinase C independent. PMID- 11108279 TI - Expression and regulation of transcripts encoding two members of the NR5A nuclear receptor subfamily of orphan nuclear receptors, steroidogenic factor-1 and NR5A2, in equine ovarian cells during the ovulatory process. AB - Steroidogenic factor-1 (SF-1, NR5A1a) is a member of the NR5A nuclear receptor subfamily and has been implicated as a key transcriptional regulator of all ovarian steroidogenic genes in vitro. To establish links between the expression of SF-1 and that of the steroidogenic genes in vivo, the objectives of this study were to clone equine SF-1 and examine the regulation of its messenger RNA (mRNA) in follicular cells during human CG (hCG)-induced ovulation. The equine SF-1 primary transcript was cloned by a combination of RT-PCR techniques. Results showed that the transcript was composed of a 5'-untranslated region (UTR) of 161 bp, an open reading frame (ORF) of 1386 bp that encodes a highly-conserved 461 amino acid protein, and a 3'-UTR of 518 bp. The cloning of SF-1 also led to the unexpected and serendipitous isolation of the highly-related orphan nuclear receptor NR5A2, which was shown to include a 5'-UTR of 243 bp, an ORF of 1488 bp, and a 3'-UTR of 1358 bp. The NR5A2 ORF encodes a 495-amino acid protein that is 60% identical to SF-1, including 99%-similar DNA-binding domains. Northern blot analysis revealed that SF-1 and NR5A2 were expressed in all major steroidogenic tissues, with the exception that NR5A2 was not present in the adrenal. Interestingly, NR5A2 was found to be, by far, the major NR5A subfamily member expressed in the preovulatory follicle and the corpus luteum. Using a semiquantitative RT-PCR/Southern blotting approach, the regulation of SF-1 and NR5A2 mRNAs in vivo was studied in equine follicular cells obtained from preovulatory follicles isolated between 0 and 39 h post hCG. Results showed that the theca interna was the predominant site of SF-1 mRNA expression in the follicle, and that hCG caused a significant decrease in SF-1 levels between 12-39 h in theca interna and between 24-39 h post hCG in granulosa cells (P < 0.05). In contrast, the granulosa cell layer was the predominant, if not the sole, site of NR5A2 mRNA expression in the follicle. Importantly, NR5A2 was much more highly expressed in granulosa cells than SF-1. The administration of hCG caused a significant decrease in NR5A2 transcripts in granulosa cells at 30, 36, and 39 h post hCG (P < 0.05). Thus, this study is the first to report the concomitant regulation of SF-1 in theca interna and granulosa cells throughout the ovulation/luteinization process, and to demonstrate the novel expression and hormonal regulation of NR5A2 in ovarian cells. Based on the marked expression of NR5A2 in equine granulosa and luteal cells and on mounting evidence of a functional redundancy between SF-1 and NR5A2 in other species, it is proposed that NR5A2 may play a key role in the regulation of gonadal steroidogenic gene expression. PMID- 11108280 TI - Insulin-mediated cellular insulin resistance decreases osmotic shock-induced glucose transport in 3T3-L1 adipocytes. AB - Similar to insulin, osmotic shock treatment of 3T3-L1 adipocytes causes translocation of GLUT4 protein to the plasma membrane and an increase in glucose transport activity. In our study, we evaluated the effect of chronic insulin treatment on the osmotic shock signaling pathway leading to GLUT4 translocation and glucose uptake. We found that chronic administration of insulin to the adipocytes induced cellular resistance to osmotic shock-stimulated GLUT4 translocation and glucose transport. We found that chronic insulin treatment attenuated shock-induced Gab-1 tyrosine phosphorylation. Furthermore, chronic insulin exposure led to a marked impairment in the ability of Gab-1 to associate with p85 subunit of PI 3-kinase in response to acute shock and insulin stimulation. Cells that were chronically treated with insulin showed a 70% and a 61% decrease in Gab-1 associated PI 3-kinase activity in shock- vs. insulin treated cells, respectively. In addition, we found that chronic insulin treatment inhibited both insulin- and osmotic shock-induced membrane ruffling, indicating that two PI 3-kinase dependent effects, GLUT4 translocation and membrane ruffling are decreased in chronically insulin-treated cells. The results described above clearly demonstrate that chronic insulin treatment induces a state of cellular resistance to osmotic shock signal transduction. PMID- 11108281 TI - Interleukin-1beta induces the expression of insulin-like growth factor binding protein-1 during decidualization in the primate. AB - Since interleukin (IL)-1 can modulate fetal/maternal interactions, we hypothesized that IL-1beta is one potential embryonic cytokine that regulates the conceptus-induced decidual response in baboon stromal fibroblasts. Treatment of stromal fibroblasts with IL-1beta (10 ng/ml, 10 min) resulted in the phosphorylation of p38 mitogen-activated protein kinase and IkappaB-alpha. This suggests that IL-1beta induces multiple signaling pathways in stromal cells that result in the activation of mitogen-activated protein kinase cascade and the transcription factor NF-kappaB. After 4 h of stimulation, IL-1beta induced gene expression of cyclooxygenase-2 (COX-2) but not cyclooxygenase-1 (COX-1). PGE2 synthesis paralleled COX-2 messenger RNA expression. The addition of hormones [36 nM estradiol-17beta, 1 microM medroxyprogesterone acetate, and 100 ng/ml relaxin] to IL-1beta-treated cells induced insulin-like growth factor binding protein-1 (IGFBP-1) messenger RNA expression after 3 days of incubation. A specific COX-2 inhibitor, NS 398 (10 nM), partially inhibited IGFBP-1 protein synthesis. In contrast, the induction of IGFBP-1 by N6, 2'-O-dibutyryladenosine 3:5'-cyclic monophosphate (dbcAMP) and hormones was not affected by NS 398 treatment. Both dbcAMP and IL-1beta, in the presence of hormones, can independently induce IGFBP 1 gene expression and decidualization. However, if IL-1beta and dbcAMP were added together, IGFBP-1 expression was inhibited. These data suggest that IL-1beta can activate multiple signaling pathways that either positively or negatively regulate IGFBP-1 gene expression and decidualization. PMID- 11108283 TI - A secreted fluorescent reporter targeted to pituitary growth hormone cells in transgenic mice. AB - In stable transfection experiments in the GH-producing GC cell line, a construct containing the entire signal peptide and the first 22 residues of human GH linked in frame with enhanced green fluorescent protein (eGFP), produced brightly fluorescent cells with a granular distribution of eGFP. This eGFP reporter was then inserted into a 40-kb cosmid transgene containing the locus control region for the hGH gene and used to generate transgenic mice. Anterior pituitaries from these GH-eGFP transgenic mice showed numerous clusters of strongly fluorescent cells, which were also immunopositive for GH, and which could be isolated and enriched by fluorescence-activated cell sorting. Confocal scanning microscopy of pituitary GH cells from GH-eGFP transgenic mice showed a markedly granular appearance of fluorescence. Immunogold electron microscopy and RIA confirmed that the eGFP product was packaged in the dense cored secretory vesicles of somatotrophs and was secreted in parallel with GH in response to stimulation by GRF. Using eGFP fluorescence, it was possible to identify clusters of GH cells in acute pituitary slices and to observe spontaneous transient rises in their intracellular Ca2+ concentrations after loading with Ca2+ sensitive dyes. This transgenic approach opens the way to direct visualization of spontaneous and secretagogue-induced secretory mechanisms in identified GH cells. PMID- 11108282 TI - Parathyroid hormone-related protein down-regulates bone sialoprotein gene expression in cementoblasts: role of the protein kinase A pathway. AB - PTH-related protein (PTHrP) acts as a paracrine and/or autocrine regulator of cell proliferation, apoptosis, and differentiation and is implicated in tooth development. The current studies employed cementoblasts to determine the role(s) and mechanisms of PTHrP in regulating cementum formation. Results demonstrated that PTHrP repressed gene expression and protein synthesis of bone sialoprotein (BSP) and abolished cementoblast-mediated biomineralization in vitro. The BSP gene inhibition required protein synthesis. The PTHrP analog (1-31) and other activators of the PKA pathway (3-isobutyl-1-methylxathine (IBMX), forskolin (FSK) and Sp-Adenosine-3', 5'-cyclic monophosphorothioate (Sp-cAMPss) also down regulated BSP gene expression and blocked cementoblast-mediated biomineralization. In contrast, the PTHrP analog (7-34), a PTHrP antagonist, and the activators of the PKC pathway [phorbol 12-myristate 13-acetate (PMA) and phorbol 12, 13-dibutyrate (PDBu)] promoted BSP gene expression. In addition, the PKA pathway inhibitor (9-(2-tetrahydrofuryl) adenine (THFA) partially, but significantly reversed the PTHrP-mediated down-regulation of BSP gene expression. Furthermore, THFA alone significantly increased BSP messenger RNA (mRNA) expression in cementoblasts. In contrast, the inhibitor of the PKC pathway (GF109203X) did not reverse the PTHrP inhibitory effect on BSP gene expression. Furthermore, GF109203X alone dramatically reduced the BSP transcript levels. These data indicate that the cAMP/PKA pathway mediates the PTHrP-mediated down regulation of BSP mRNA expression in cementoblasts; and furthermore, this pathway may, through an intrinsic inhibition mechanism, regulate the basal level of BSP mRNA expression. In contrast, the activation of PKC promotes BSP gene expression. These data provide new insights into the molecular mechanisms involved in PTHrP regulation of cementogenesis. PMID- 11108284 TI - Atrial (ANP) and brain natriuretic peptide (BNP) expression after myocardial infarction in sheep: ANP is synthesized by fibroblasts infiltrating the infarct. AB - Cardiac gene expression of atrial natriuretic peptide (ANP) and that of brain natriuretic peptide (BNP) are markedly elevated after myocardial infarction. The cellular distribution and temporal responses of ANP and BNP messenger RNA (mRNA) expression were compared by in situ hybridization for 5 weeks after left coronary artery ligation in sheep. Ligation resulted in highly reproducible, transmural, left ventricular infarcts. Within the infarct, ANP mRNA appeared from 7 days after ligation, whereas BNP expression was undetectable in the infarct at any time. The cells synthesizing ANP were shown by in situ hybridization and immunocytochemistry to be fibroblasts invading the infarct. The appearance of ANP expression coincided with the transition of these cells to the myofibroblast phenotype. Treatment of cultured cardiac fibroblasts with transforming growth factor-beta (10 ng/ml) induced the expression of alpha-smooth muscle actin, characteristic of the transformation to myofibroblasts, and raised ANP concentrations in the medium. In the surviving myocardium of the left ventricle, ANP and BNP expression increased in response to ligation, BNP mRNA was particularly strong at the lateral margins of the infarct. In both left and right atria, levels of BNP mRNA increased markedly over the first 18 h, whereas levels of atrial ANP mRNA decreased over 3 days after infarction. This is the first report of ANP expression and synthesis by cardiac fibroblasts invading the fibrotic scar, suggesting that ANP may be involved in regulating fibroblast proliferation during reparative fibrosis. PMID- 11108285 TI - A small composite probasin promoter confers high levels of prostate-specific gene expression through regulation by androgens and glucocorticoids in vitro and in vivo. AB - Transient transfection studies have shown that the probasin (PB) promoter confers androgen selectivity over other steroid hormones, and transgenic animal studies have demonstrated that the PB promoter will target androgen, but not glucocorticoid, regulation in a prostate-specific manner. Previous PB promoters either targeted low levels of transgene expression or became too large to be conveniently used. The goal was to design a PB promoter that would be small, yet target high levels of prostate-specific transgene expression. Thus, a composite probasin promoter (ARR2PB) coupled to the bacterial chloramphenicol acetyltransferase reporter (ARR2PBCAT) was generated and tested in prostatic and nonprostatic cell lines and in a transgenic mouse model. In PC-3, LNCaP, and DU145 prostate cancer cell lines, the ARR2PB promoter gave basal expression and was induced in response to androgen and glucocorticoid treatment after cotransfection with the respective steroid receptor. Basal expression of ARR2PBCAT in the nonprostatic COS-1, MCF-7, ZR-75-1, and PANC-1 cell lines was very low; however, CAT activity could be induced in response to androgens and glucocorticoids when cells were cotransfected with either the AR or GR. In contrast to the transfection studies, ARR2PBCAT transgene expression remained highly specific for prostatic epithelium in transgenic mice. CAT activity decreased after castration, and could be induced by androgens and, in addition, glucocorticoids. This demonstrates that the necessary sequences required to target prostate-specific epithelial expression are contained within the composite ARR2PB minimal promoter, and that high transgene expression can now be regulated by both androgens and glucocorticoids. The ARR2PB promoter represents a novel glucocorticoid inducible promoter that can be used for the generation of transgenic mouse models and in viral gene therapy vectors for the treatment of prostate cancer in humans. PMID- 11108286 TI - Intracellular calcium and protein kinase C mediate expression of receptor activator of nuclear factor-kappaB ligand and osteoprotegerin in osteoblasts. AB - Receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) produced by osteoblasts/stromal cells are involved as positive and negative regulators in osteoclast formation. Three independent signals have been proposed to induce RANKL expression in osteoblasts/stromal cells: vitamin D receptor-, cAMP-, and gp130-mediated signals. We previously reported that intracellular calcium-elevating compounds such as ionomycin, cyclopiazonic acid, and thapsigargin induced osteoclast formation in cocultures of mouse bone marrow cells and primary osteoblasts. Increases in calcium concentration in culture medium also induced osteoclast formation in cocultures. Treatment of primary osteoblasts with these compounds or with high calcium medium stimulated the expression of both RANKL and OPG messenger RNAs (mRNAs). 1,2-Bis(o aminophenoxy)ethane-N,N,N',N'-tetraacetic acid)-tetra(acetoxymethyl)ester, an intracellular calcium chelator, suppressed both ionomycin-induced osteoclast formation in cocultures and expression of RANKL and OPG mRNAs in primary osteoblasts. Phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, also stimulated osteoclast formation in these cocultures and the expression of RANKL and OPG mRNAs in primary osteoblasts. Protein kinase C inhibitors such as calphostin and staurosporin suppressed ionomycin- and PMA induced osteoclast formation in cocultures and expression of RANKL and OPG mRNAs in primary osteoblasts. Ionomycin stimulated RANKL mRNA expression in ST2 and MC3T3-G2/PA6 cells, but not in MC3T3-E1 or NIH-3T3 cells. These effects were closely correlated with osteoclast formation in response to ionomycin in cocultures with these stromal cell lines. OPG strongly inhibited osteoclast formation induced by calcium-elevating compounds and PMA in cocultures, suggesting that RANKL expression in osteoblasts is a rate-limiting step for osteoclast induction. Forskolin, an activator of cAMP signals, also stimulated osteoclast formation in cocultures. Forskolin enhanced RANKL mRNA expression but suppressed OPG mRNA expression in primary osteoblasts. These results suggest that the calcium/protein kinase C signal in osteoblasts/stromal cells is the fourth signal for inducing RANKL mRNA expression, which, in turn, stimulates osteoclast formation. PMID- 11108287 TI - Hormonal control of gubernaculum development during testis descent: gubernaculum outgrowth in vitro requires both insulin-like factor and androgen. AB - The gubernaculum connects the gonad to the inguinoscrotal region and is involved in testis descent. It rapidly develops in the male fetus, whereas development in the female fetus is lacking. Possible factors involved in gubernaculum development are androgens, anti-Mullerian hormone (AMH), and insulin-like factor (Insl3). Sexual dimorphism in gubernaculum development correlated with the mitotic activity of cells in the gubernacular bulbs from male and female fetuses. Androgen receptor expression was restricted to the mesenchymal core of the gubernacular bulb, whereas skeletal muscle was detected in its outer layer. In an organ culture system devised to further study gubernaculum development in vitro, morphology of gubernacular explants grown in the presence of testes was comparable with that of gubernacula developed in vivo. Testicular tissue or medium containing R1881, a synthetic androgen, had a growth stimulatory effect on gubernacular explants compared with ovarian tissue or basal medium only. Moreover, Amh-/-, Amh+/-, and Insl3+/- testes stimulated the growth of gubernacular explants to the same extent as control testes. Insl3-/- testes, however, did not produce such an activity. This study reveals an essential role for both androgen and Insl3 in the gubernaculum outgrowth during transabdominal testis descent. PMID- 11108288 TI - Smad2 and 3 mediate transforming growth factor-beta1-induced inhibition of chondrocyte maturation. AB - Transforming growth factor-beta (TGF-beta) is a multifunctional regulator of a variety of cellular functions, including proliferation, differentiation, matrix synthesis, and apoptosis. In growth plate chondrocytes, TGF-beta slows the rate of maturation. Because the current paradigm of TGF-beta signaling involves Smad proteins as downstream regulators of target genes, we have characterized their role as mediators of TGF-beta effects on chondrocyte maturation. Both Smad2 and 3 translocated to the nucleus upon TGF-beta1 signaling, but not upon BMP-2 signaling. Cotransfection experiments using the TGF-beta responsive and Smad3 sensitive p3TP-Lux luciferase reporter demonstrated that wild-type Smad3 potentiated, whereas dominant negative Smad3 inhibited TGF-beta1 induced luciferase activity. To confirm the role of Smad2 and 3 as essential mediators of TGF-beta1 effects on chondrocyte maturation, we overexpressed both wild-type and dominant negative Smad2 and 3 in virally infected chondrocyte cultures. Overexpression of both wild-type Smad2 and 3 potentiated the inhibitory effect of TGF-beta on chondrocyte maturation, as determined by colx and alkaline phosphatase activity, whereas dominant negative Smad2 and 3 blocked these effects. Wild-type and dominant negative forms of Smad3 had more pronounced effects than Smad2. Our results define Smad2 and 3 as key mediators of the inhibitory effect of TGF-beta1 signaling on chondrocyte maturation. PMID- 11108289 TI - Direct effects of nerve growth factor on thecal cells from antral ovarian follicles. AB - TrkA, the nerve growth factor (NGF) tyrosine kinase receptor, is expressed not only in the nervous system, but also in nonneural cells, including discrete cellular subsets of the endocrine and immune system. In the rat ovary, trkA receptor abundance increases strikingly in thecal-interstitial cells during the hours preceding the first ovulation. Blockade of either trkA transducing capacity or NGF biological activity inhibited ovulation, suggesting a role for NGF in the ovulatory process of this species. To identify some of the processes that may be affected by trkA activation in the thecal compartment, we used purified thecal cells/thecal fibroblasts from bovine ovaries (heretofore referred to as thecal cells). Ribonuclease protection assays employing bovine-specific cRNA probes demonstrated the presence of the messenger RNAs (mRNAs) encoding NGF and its receptors, p75 NTR and trkA, in the thecal compartment of small, medium, and large antral follicles and showed that trkA mRNA is also expressed in granulosa cells. In situ hybridization and immunohistochemical examination of intact ovaries confirmed these cellular sites of NGF and trkA synthesis. TrkA mRNA, but not NGF mRNA, was lost within 48 h of placing thecal cells in culture. Thus, to study trkA-mediated actions of NGF on these cells we transiently expressed the receptor by transfection with a vector containing a full-length rat trkA complementary DNA under transcriptional control of the cytomegalovirus promoter. Because ovulation is preceded by an LH-dependent increase in androgen and progesterone production, the ability of NGF to modify the release of these steroids was determined in freshly plated cells still containing endogenous trkA receptors and in cells undergoing luteinization in culture that were transiently transfected with the trkA-encoding plasmid. NGF stimulated both androgen and progesterone release in freshly plated thecal cells, but not in luteinizing cells provided with trkA receptors. As ovulation in rodents requires an increased formation of PGE2 and has been shown to be antedated by proliferation of thecal fibroblasts, we determined the ability of NGF to affect these parameters in trkA transfected thecal cells. The neurotrophin rapidly stimulated PGE2 release and amplified the early steroidal response to hCG in trkA-expressing cells, but not in cells lacking the receptor. Likewise, NGF stimulated [3H]thymidine incorporation into trkA-containing cells, but not into cells that had lost the receptor in culture. Induction of ovulation in immature rats by gonadotropin treatment verified that an increased cell proliferation in the thecal compartment, determined by the incorporation of bromodeoxyuridine into cell nuclei, occurs 4-5 h before ovulation in this species. These results suggest that the contribution of NGF to the ovulatory process includes a stimulatory effect of the neurotrophin on steroidogenesis, PGE2 formation, and proliferative activity of thecal compartment cells. PMID- 11108290 TI - Disulfide bond mutations in follicle-stimulating hormone result in uncoupling of biological activity from intracellular behavior. AB - The crystal structure of human CG reveals that each subunit is a member of the superfamily of cystine-knot growth factors. Although the distribution of the cysteine residues in all the beta-subunits is conserved, the conformation of the human FSH dimer differs from that of the CG/LH dimers. This suggests that the function of the cystine bonded loops in the human FSHbeta-subunit may differ from that in the CGbeta-subunit. To address this issue, we deleted two disulfide bonds in the FSHbeta domain: cys 20-104 and cys 28-82, which correspond to the disulfide bonds 26-110 and 34-88, respectively, in the CGbeta-subunit. The cys 26 110 bond is associated with the "seat-belt" region and cys 34-88 is a bond in the cystine knot. Coexpression of the wild-type alpha-subunit with the FSHbeta cysteine mutants in CHO cells revealed no detectable heterodimer. The FSHbeta mutants were then incorporated into a single chain where the beta-subunit is genetically fused to the alpha-subunit. In such a model, the rate-limiting subunit assembly step is by-passed and mutations that otherwise block heterodimer formation can be evaluated in terms of biological activity. Compared with the nonmutated single chain, the single-chain 28-82 mutant is secreted more slowly and its recovery is substantially reduced, whereas secretion and recovery of the 20-104 mutant was not significantly affected. The receptor binding affinity of the cys 28-82 mutant did not differ from wild-type and binding of the cys 20-104 mutant was decreased only 2-fold. The signal transduction data parallel the binding affinities, although the maximal accumulation of cAMP is less for the cys 20-104 mutant than that seen for cys 28-82 and nonmutated single-chains variants. These data support the hypothesis that the determinants for intracellular behavior and bioactivity of the gonadotropins are not the same, and that the cystine knot is a critical determinant for the formation of a stable, assembly competent subunit. In addition, the data imply that the "seat-belt" conformation does not play a prominent role in the bioactivity of FSH. PMID- 11108291 TI - Neuroendocrine aging in the female rat: the changing relationship of hypothalamic gonadotropin-releasing hormone neurons and N-methyl-D-aspartate receptors. AB - The reproductive axis undergoes alterations during aging, resulting in acyclicity and the loss of reproductive function. In the hypothalamus, changes intrinsic to GnRH neurons may play a critical role in this process, as may changes in inputs to GnRH neurons from neurotransmitters such as glutamate. We investigated the effects of age and reproductive status on neuroendocrine glutamatergic NMDA receptors (NRs), their regulation of GnRH neurons, and their expression on GnRH neurons, in female rats. First, we quantified NR subunit messenger RNAs (mRNAs) in preoptic area-anterior hypothalamus (POA-AH) and medial basal hypothalamus (MBH), the sites of GnRH perikarya and neuroterminals, respectively. In POA-AH, NR1 mRNA levels varied little with age or reproductive status. NR2a and NR2b mRNA levels decreased significantly between cycling and acyclic rats. In MBH, NR mRNAs all increased with aging, particularly in acyclic animals. Second, we tested the effects of N-methyl-D,L-aspartate (NMA) on GnRH mRNA levels in POA-AH of aging rats. NMA elevated GnRH mRNA levels in young rats, but decreased them in middle aged rats. Third, we quantified expression of the NR1 subunit on GnRH perikarya in aging rats using double label immunocytochemistry. NR1 expression on GnRH cell bodies varied with age and reproductive status, with 30%, 19%, and 46% of GnRH somata double labeled with NR1 in young proestrous, middle-aged proestrous, and middle-aged persistent estrous rats, respectively. Thus, 1) the expression of hypothalamic NR subunit mRNAs correlates with reproductive status; 2) changes in NR subunit mRNA levels, if reflected by changes in protein levels, may result in alterations in the stoichiometry of the NR during aging, with possible physiological consequences; 3) the effects of NR activation on GnRH mRNA switches from stimulatory to inhibitory during reproductive aging; and 4) expression of the NR1 subunit on GnRH perikarya changes with reproductive status. These molecular, physiological, and cellular neuroendocrine changes are proposed to be involved in the transition to acyclicity in aging female rats. PMID- 11108292 TI - The expression of osteoprotegerin and RANK ligand and the support of osteoclast formation by stromal-osteoblast lineage cells is developmentally regulated. AB - The one or more molecular mechanisms that determine the obligatory sequence of resorption followed by formation during bone remodeling is unclear. RANK ligand (RANK-L) is an essential requirement for osteoclastogenesis, and its activity is neutralized by binding to the soluble decoy receptor, osteoprotegerin (OPG). Because both molecules are produced by osteoblast lineage cells, we studied their developmental regulation in a conditionally immortalized human marrow stromal (hMS[2-15]) cell line. These cells can simulate the complete developmental sequence from undifferentiated precursor(s) to cells with the complete osteoblast phenotype that are capable of forming mineralized nodules. During osteoblast differentiation, RANK-L messenger RNA levels decreased by 5-fold, whereas OPG messenger RNA levels increased by 7-fold, resulting in a 35-fold change in the RANK-L/OPG ratio. OPG protein also increased by 6-fold. Mouse bone marrow cells generated osteoclast-like cells in coculture with undifferentiated hMS(2-15) cells, but did not when cocultured with hMS(2-15) cells in varying stages of differentiation, unless an excess of RANK-L was added. Thus, undifferentiated marrow stromal cells with a high RANK-L/OPG ratio can initiate and support osteoclastogenesis, but after differentiation to the mature osteoblast phenotype, they cannot. We speculate that the developmental regulation of OPG and RANK-L production by stromal/osteoblast cells contributes to the coordinated sequence of osteoclast and osteoblast differentiation during the bone remodeling cycle. PMID- 11108293 TI - Female fertility is reduced in mice lacking Ca2+/calmodulin-dependent protein kinase IV. AB - Ca2+/calmodulin-dependent protein kinase IV (CaMKIV) is a serine/threonine protein kinase with limited tissue distribution. CaMKIV is highly expressed in the testis, where it is found in transcriptionally inactive elongating spermatids. We have recently generated mice deficient in CaMKIV. In the absence of CaMKIV, the exchange of sperm nuclear basic proteins in male spermatids is impaired, resulting in male infertility secondary to defective spermiogenesis. The involvement of CaMKIV in female fertility has not been addressed. Here we report that female fertility is markedly reduced in CaMKIV-deficient mice due to impaired follicular development and ovulation. CaMKIV is expressed in the ovary, where it is localized in granulosa cells. We further find that in cultured granulosa cells, CaMKIV expression and subcellular localization are hormonally regulated. As granulosa cells differentiate, CaMKIV levels decrease and the kinase translocates from the nucleus into the cytoplasm. Our results demonstrate a critical role for CaMKIV in female reproduction and point to a potential function in granulosa cell differentiation. PMID- 11108294 TI - High insulin-like growth factor 1 (IGF-1) and insulin concentrations trigger apoptosis in the mouse blastocyst via down-regulation of the IGF-1 receptor. AB - Women with polycystic ovary syndrome have significantly higher rates of pregnancy loss, as well as elevated insulin and IGF-1 levels. In this study, preimplantation embryos exposed to high concentrations of IGF-1 or insulin undergo extensive apoptosis of the ICM nuclei. Lack of BAX expression, the caspase inhibitor, zVAD, or the ceramide synthase inhibitor, fumonisin B1, prevents this event, suggesting involvement of programmed cell death effector pathways. In other systems, the IGF-1 concentration regulates IGF-1R expression and thus high concentrations lead to down-regulation of the receptor. Here, data show a decrease in IGF-1 receptor protein expression, both by confocal immunofluorescent microscopy and by Western analysis upon exposure to 130 nM IGF 1. Insulin-stimulated glucose uptake, an event regulated via the IGF-1 receptor, is decreased upon exposure to excess IGF-1, suggesting decreased function of the receptor. The data also show that, by blocking receptor signal transduction or by decreasing receptor expression, the apoptotic event can be recreated, thus strongly suggesting that the mechanism of high IGF-1 induced apoptosis is decreased downstream IGF-1 receptor signaling. This embryotoxic insult by high IGF-1 levels may be responsible for the high incidence of pregnancy loss seen in women with polycystic ovary syndrome. PMID- 11108296 TI - Central effect of ghrelin, an endogenous growth hormone secretagogue, on hypothalamic peptide gene expression. AB - Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor (GHS R), was originally purified from the rat stomach. Like the synthetic GHSs, ghrelin specifically releases GH following intravenous administration. Also consistent with the central actions of GHSs, ghrelin-immunoreactive cells were shown to be located in the hypothalamic arcuate nucleus as well as the stomach. However, the central actions of ghrelin have not been elucidated. Here, we used radioactive in situ hybridization histochemistry to examine the effects of central administration of rat ghrelin on neuropeptide genes that are expressed in hypothalamic neurons that were previously shown to express GHS-R. We found that central administration of ghrelin increased both agouti-related protein (AGRP) mRNA levels (245.8 +/- 28.3% of the saline-treated controls; p < 0.01) in the hypothalamus and food intake (5.7 +/- 0.9 g ghrelin vs. 1.9 +/- 0.5 g saline; p < 0.05). On the other hand, 1 microg of rat ghrelin central administration did not alter the episodic GH release of freely moving adult male rats. Thus, ghrelin has an alternative role in stimulating food intake via an increase of AGRP rather than the release of GH from the pituitary. PMID- 11108295 TI - In vivo effects of bisphosphonates on the osteoclast mevalonate pathway. AB - Estrogen deficiency is a leading cause of osteoporosis associated with increased osteoclastic bone resorption. In vitro studies indicate that the clinically used nitrogen-containing bisphosphonates (N-BPs) such as alendronate (ALN), risedronate (RIS) and ibandronate (IBA) suppress bone resorption via inhibition of the mevalonate pathway enzyme farnesyl diphosphate (FPP) synthase in osteoclasts (Ocs). The object of this study was to test the hypothesis that N-BPs inhibit the mevalonate pathway of Ocs in vivo. The mevalonate pathway enzyme hydroxymethyl-glutaryl-coenzyme A reductase (HMGR), is modulated by feedback inhibition from downstream metabolites. We therefore evaluated the in vivo expression of HMGR in Ocs from animals treated with BP. The N-BPs, ALN, IBA and RIS, selectively suppressed HMGR expression in up to 85% of rat tibia osteoclasts, after 48 hr treatment. Etidronate and clodronate, bisphosphonates that do not inhibit FPP synthase, were without effect. Simvastatin treatment opposed ALN reduction of HMGR expression, suggesting regulation by a metabolite(s) between mevalonate and FPP. These data provide the first in vivo evidence for N-BP effects on the mevalonate pathway in osteoclasts, and strongly support the hypothesis that N-BPs act via this mechanism. PMID- 11108297 TI - Local integration of glutamate signaling in the hypothalamic paraventricular region: regulation of glucocorticoid stress responses. AB - The present study is a test of the hypothesis that endogenous glutamatergic input to the hypothalamic paraventricular nucleus (PVN) is involved in stress-induced activation of the hypothalamic-pituitary-adrenocortical (HPA) axis. We examined whether corticosterone (CORT) responses to restraint stress could be attenuated by bilateral 50 nl microinjections of kynurenic acid (KYN, ionotropic glutamate receptor antagonist) into the medial PVN of conscious rats. Immediately following microinjection, rats were subjected to 30 min restraint, and stress-induced plasma CORT was measured at 30, 60, 120, and 180 min following the onset of restraint. KYN (50 pmol) significantly reduced cumulative CORT responses to acute restraint stress by 24%. In contrast, microinjections centered dorsal to the PVN increased CORT responses by 31%, suggestive of a disinhibition of local PVN inhibitory input. KYN injections immediately anterior, ventral, or posterior to the PVN had no effect, suggesting an absence of potential diffusion artifacts. These results provide evidence that endogenous glutamate in the PVN and surrounding peri-PVN region plays a physiologically significant role in both generating and limiting glucocorticoid stress responses in awake rats. PMID- 11108298 TI - Differential pituitary gene expression profiles associated- to aging and spontaneous tumors as revealed by rat cDNA expression array. AB - Aging of the rat pituitary is often accompanied by the occurrence of adenomas. We asked whether complementary DNA hybridization array was adapted to identify gene expression patterns linked to aging and associated spontaneous adenomas. Thus, [32P]dATP-labeled cDNAs were prepared from pituitaries of three month-old rats (Y) and tumor-bearing 20-28-month-old rats (OT). The cDNAs were hybridized to identical membrane arrays allowing to study simultaneously 588 known genes (Clontech 7738-1). Among the 79 genes detected, the GH gene was predominantly expressed in both groups. Twenty-eight genes in the OT group and 15 in the Y group were found to be expressed at a higher level. The largest differences were of about 17 fold and were observed for the galanin and glutathione S transferase genes in the Y and OT groups, respectively. Relative RT-PCR was applied to validate the OT versus Y expression pattern obtained via cDNA array hybridization. The results were consistent for 14 out the 15 genes tested. In the light of these results, differential membrane array hybridization appears suitable to identify gene expression profiles associated with pituitary aging. PMID- 11108299 TI - Increased stem cell dose, as obtained using currently available technology, may not be sufficient for engraftment of haploidentical stem cell transplants. AB - The best strategies for haploidentical stem cell transplants are not known. We used a standard myeloablative pretransplant conditioning regimen (30 mg/kg VP-16, 120 mg/kg cyclophosphamide, and 12 Gy of TBI in six fractions), an increased peripheral stem cell dose of > 10 x 10(6) CD34+ cells/kg, T cell depletion (with CD34+ cell selection and CD4/CD8 depletion steps) to < 1 x 10(5) CD3+ cells/kg and cyclosporine post transplant. Ten patients (7M/3F, median age 11 (3-33) years) with high-risk leukemia (AML in 4, MDS in 2, CML in 1 and T-ALL in 3) received a hemopoietic stem cell transplant (HSCT) from a haploidentical father or sibling. The median number of CD34+ cells was 12.9 (9.5-45.7) x 10(6) cells/kg; median number of CD3+ cells was 0.41 (0.09-1.89) x 10(5) CD3+ cells/kg. All patients initially achieved 0.5 x 10(9)/l neutrophils at a median 12 (10-21) days. Graft failure in two consecutive patients out of four on the original protocol led to a modification adding ATG pretransplant and OKT3 post transplant. Graft failure was observed in one out of six subsequent patients. Acute GVHD > or = grade II was observed in three patients. Three of 10 patients are alive in CR at > 24 and >3 (2) months after transplant. Seven patients died: four of transplant related complications and three of relapse. Increased stem cell dose (> or = 10 x 10(6) CD34+ cells/kg) as obtained using currently available technology may not be sufficient to ensure stable engraftment in patients with high-risk leukemia using standard myeloablative conditioning regimens. PMID- 11108300 TI - The influence of early transplantation, age, GVHD prevention regimen, and other factors on outcome of allogeneic transplantation for CML following BuCy. AB - Results in 164 patients who underwent allogeneic marrow transplantation following busulfan and cyclophosphamide over a 15 year period were analyzed. Age (median 37, range 14-66 years) did not significantly affect the incidence of graft-versus host disease (GVHD), but patients who received methotrexate with cyclosporine had a significantly lower incidence (P = 0.002) of chronic GVHD compared to those who received methylprednisolone with cyclosporine. Hepatic veno-occlusive disease (VOD) occurred less frequently in chronic phase patients (P = 0.002) and in those transplanted shortly after diagnosis (P = 0.001). Five year leukemia-free survival (LFS) for the entire group was 49% (95% CI 41-57%). For 102 patients who underwent transplantation in chronic phase, results were significantly improved by transplantation at a short interval following diagnosis, particularly within 3 months (P = 0.01), by the use of methotrexate and not corticosteroids for GVHD prevention (P = 0.03), and by use of HLA-identical sibling donors (P = 0.01). Age was not a significant adverse prognostic factor and transplantation was successfully performed in individuals up to age 66. Allogeneic transplantation in CML, including older patients and those with unrelated donors, can be most safely and effectively performed shortly after diagnosis. PMID- 11108301 TI - High-dose idarubicine, busulphan and melphalan as conditioning for autologous blood stem cell transplantation in multiple myeloma. A feasibility study. AB - Extensive studies have tested the clinical impact of double and triple sequential transplants as front-line therapy in MM, following the suggestion that dose escalation can overcome the marked drug resistance characteristic of this disease, but the superiority of such approaches vs one single transplant has still to be demonstrated. The aim of our study was to evaluate the feasibility and efficacy of high-dose idarubicine intensification of a standard busulphan melphalan conditioning regimen in MM. Twenty-eight patients (median age 55 years) with sensitive disease received PBSCT after high-dose idarubicine combined with busulphan and melphalan and followed by s.c. rhG-CSF until PMN recovery. The most severe toxicity was represented by oral mucositis which resolved with hemopoietic reconstitution. Overall response and CR rate were 52% and 40%, respectively. Currently, 36 patients are alive and 19 are progression-free a median of 20 months (12-36) from transplant. The 3-year projected probability of progression free survival for patients transplanted after first-line treatment is 60%. The combination of Ida/Bu/Melph appears a promising alternative regimen for PBSCT in myeloma, with low transplant-related toxicity and fast hematological recovery. Long-term follow-up and a prospective randomized study, now ongoing, will probably clarify whether an idarubicine-intensified regimen will result in superior outcomes to conventional conditioning and even be comparable to a double consecutive transplant program. PMID- 11108302 TI - Autologous stem cell transplantation (ASCT) with immunologically purged progenitor cells in patients with advanced stage follicular lymphoma after early partial or complete remission: toxicity, follow-up of minimal residual disease and survival. AB - The role of autologous stem cell transplant (ASCT) in indolent lymphomas is a controversial issue. From 1994 to 1999, we performed ASCT with immunologically purged progenitor cells in 15 patients with advanced stage follicular lymphoma (FL) after early partial or complete remission. Results of the purging strategy and follow-up of minimal residual disease after transplant were analyzed with PCR amplification of bcl-2/IgH rearrangement for the t(14;18) translocation. A comparison of transplanted patients with a group of controls was carried out to evaluate differences in progression-free survival and overall survival. Eighty percent of patients received one chemotherapy regimen before ASCT and were in first remission. All the patients received cyclophosphamide plus hyperfractionated total body irradiation as the conditioning regimen. Nine patients were transplanted with bone marrow (BM) and six with peripheral blood progenitor cells (PBPC). Engraftment was delayed in one patient transplanted with BM. Two patients died during the transplant procedure. Ten of 12 evaluable patients were PCR positive in the BM for bcl-2 rearrangement at diagnosis. Six of them (60%) were still positive after chemotherapy, and one patient was transplanted with a positive hematopoietic product after purging. With a median follow-up of 27 months, six of eight evaluable patients still remain PCR negative in the BM. With a median follow-up of 4.7 years from diagnosis, progression-free survival was 83% (95% CI: 63-100). The risk of disease progression of non transplanted patients was 19.2 times higher than that of transplanted patients (P = 0.01), but no differences were found in overall survival. Regarding patients in first remission, the risk of relapse was 12.6 times higher in non-transplanted than in transplanted patients (P = 0.04). This procedure seems to offer a good chance to achieve a PCR-negative state and prolonged freedom from recurrence. According to these results, prospective randomized trials are warranted. PMID- 11108303 TI - Is it safe to treat allogeneic stem cell transplant recipients at home during the pancytopenic phase? A pilot trial. AB - After myeloablative treatment and allogeneic stem cell transplantation (ASCT), patients are kept isolated in the hospital to prevent infections during neutropenia. To date, 22 patients have been given the choice of being treated at home. Eleven could not be treated at home, and they served as controls. Most had haematological malignancies. The donors were 12 HLA-compatible unrelated, nine HLA-identical siblings and one twin. In the home care group, three developed bacteraemia, compared to nine in the controls (P < 0.01). Patients in the home care group had fewer days of total parenteral nutrition (median 3 vs 24, P < 0.001), required fewer erythrocyte transfusions (median 4 vs 8, P = 0.01), fewer days on i.v. antibiotics (median 6 vs 13 days), and on analgesics (median 0 vs 15) than the controls (P < 0.05). Days with fever, time to engraftment, days with G-CSF and acute GVHD were the same in the two groups. Seven of 11 patients treated at home were readmitted to the ward for a median of 3 (0-7) days, due to fever or lack of a caregiver at home. Days to discharge to the out-patient clinic were faster in the group treated at home (median 20 vs 35 days, P < 0.01). Patients who were treated at home enjoyed being active and taking a walk when they felt like it. This preliminary report suggests that home care after ASCT is not only safe, but superior to isolation in the hospital. PMID- 11108304 TI - Hospital capacity and post-transplant survival after allogeneic bone marrow transplantation: analysis of data from the Japan Society for Hematopoietic Cell Transplantation. AB - The association between hospital capacity and survival after allogeneic bone marrow transplantation (allo-BMT) was examined using the dataset accumulated by the Japan Society of Hematopoietic Cell Transplantations (JSHCT). The subjects were 3134 patients who received first allo-BMTs between 1991 and 1997 reported to the JSHCT. They were divided into three groups by cumulative hospital experience of allo-BMTs: low volume (capacity) (LV; < or = 25 cases), moderate volume (capacity) (MV; 26-75 cases) and high volume (capacity) (HV; > or = 76 cases). Using a proportional hazards model, the association of hospital experience with early survival at day 100 (D100S), and overall survival (OS) were examined. For leukemia patients, leukemia-free survival (LFS) was also analyzed. When HV was defined as the reference group, the hazard ratios (HRs) of OS for all subjects were 1.10 (95% confidence interval; 0.97-1.25) for MV and 1.25 for LV (1.08 1.44). The HRs with D100S were 1.20 (0.96-1.51) for MV and 1.40 (1.08-1.80) for LV. Larger values were observed for OS and D100S in cases of leukemia. Survival after BMT from sibling donors was clearly influenced by hospital experience, but this was not the case from unrelated donors. These findings suggest that size of the transplant team should be considered in order to improve the outcome of sibling BMT in general. PMID- 11108305 TI - Significant MLR but not CTL responses against recipient antigens generated in T cells from bone marrow chimeras recovered from acute GVHD. AB - Lethally irradiated AKR mice received BMT from H-2D and minor lymphocyte stimulatory (Mls)-1 disparate B10.A mice. No GVHD signs were detected in AKR recipients of T cell-depleted BM cells (1 x 10(7)) alone ([B10.A --> AKR] T-). When B10.A splenic T cells (1 x 10(5)) were injected in addition to T cell depleted BM cells ([B10.A --> AKR] T+), overt GVHD was observed. [B10.A --> AKR] T+ chimeras recovered from the GVHD 8 weeks after BMT. In T cells from these [B10.A --> AKR] T+ chimeras, a substantial population of Mls-1a-reactive Vbeta6+ T cells was present, whereas the Vbeta6+ cells were deleted in [B10.A --> AKR] T- chimeras. T cells from [B10.A --> AKR] T+ chimeras showed considerable MLR but no CTL response against AKR cells (split tolerance). Upon stimulation with AKR stimulators or anti-CD3 MoAb, T cells from [B10.A --> AKR] T+ chimeras produced significantly more IL-4 but significantly less IFN-gamma compared with those from [B10.A --> AKR] T- chimeras or unmanipulated B10.A mice. The serum level of IgG1 in [B10.A --> AKR] T+ chimeras was also significantly higher than that in [B10.A -> AKR] T- or B10.A mice. The present findings suggest that the split tolerance observed in BMT chimeras recovered from GVHD is attributable to the Th2 dominant state. PMID- 11108306 TI - Apoptosis of keratinocytes is associated with infiltration of CD8+ lymphocytes and accumulation of Ki67 antigen. AB - In 18 allogeneic blood/marrow transplanted (BMT) cases, skin biopsies were taken for acute GVHD diagnosis. In 14 cases clinical and histopathology data were consistent with acute GVHD with toxic lesions in four cases. All skin biopsies were stained for the presence of apoptotic cells (TUNEL technique). Nine patients' skin biopsies were positive for this stain. Apoptotic cells were seen in epidermal cells of the epidermis and in hair follicles. The presence of apoptotic cells was associated with (1) advanced stage (> or = II grade) acute GVHD (P = 0.015) and (2) heavy mononuclear cell infiltration (P = 0.015), predominantly of CD8+ cells (P = 0.025). Ki67 immunoreactivity of epidermal cells was different in the epidermis having and lacking apoptotic cells. Ki67 MoAb staining of epidermal cells revealed that nuclei were enlarged and stained stronger in biopsies with than biopsies without apoptotic cells (P = 0.008). Cyclin A positive cells were in a high proportion and cdc2-positive cells in a low proportion in patients' biopsies with acute GVHD and toxic lesions. In addition cyclin A accumulation was seen in keratinocytes in GVHD biopsies. Therefore, mitosis of skin cells is impaired, likely due to the genotoxic effect of conditioning, which renders the cells susceptible to acute GVHD-associated apoptotic damage. PMID- 11108308 TI - Reproductive status in long-term bone marrow transplant survivors receiving busulfan-cyclophosphamide (120 mg/kg). AB - There are few published data on the recovery of fertility after 'little' Bu-Cy (busulfan 16 mg/kg, cyclophosphamide 120 mg/kg) conditioning for BMT. To address this, we identified 19 females aged less than 40 years at transplant and 47 males from a single centre who were alive a minimum of 2 years after BMT with little Bu Cy as conditioning and who were evaluable for testing. FSH, LH, testosterone and inhibin B levels were measured in males. Twenty-six also had semen analysis, a median of 5 years post transplant; 21 had detectable sperm, with 11 having counts >20 x 10(6)/ml. There was an association between prolonged chronic graft-versus host disease and low sperm counts. FSH and inhibin B levels correlated with sperm counts but not to the extent that they could reliably predict counts in individual patients. An additional six of seven males attempting to father children did so, a median of 3.2 years post transplant. Low testosterone levels were noted in 12% of males, most of whom had symptoms consistent with androgen deficiency. FSH, LH and oestradiol levels in the absence of hormone replacement therapy were measured in females; all remained amenorrheic with endocrine evidence of ovarian failure. These results have implications for fertility counselling and hormone replacement therapy both pre- and post BMT. PMID- 11108307 TI - Use of PEG-rHuMGDF in platelet engraftment after autologous stem cell transplantation. AB - This paper summarizes a pilot, sequential dose-escalation study of PEG-rHuMGDF in patients with advanced malignancies who had delayed platelet recovery after autologous stem cell transplantation (ASCT). Patients were randomized to receive either placebo (n = 11) or PEG-rHuMGDF at 5 (n = 9), 10 (n = 6), or 25 (n = 7) microg/kg/day by subcutaneous injection for 14 days and were monitored for 5 weeks. Across all treatment groups, eight patients had platelet recovery to > or = 20 x 10(9)/l by day 21. The proportion of patients achieving platelet recovery, the median number of days and units of platelet transfusions were similar for the placebo and the PEG-rHuMGDF groups. PEG-rHuMGDF was well tolerated at all dosages. The incidence rates of adverse events in all groups were similar. No deaths on study, no drug-related serious adverse events, and no development of neutralizing antibodies to MGDF occurred. PMID- 11108309 TI - Respiratory viral infections in primary immune deficiencies: significance and relevance to clinical outcome in a single BMT unit. AB - Respiratory viral infections are major causes of morbidity and mortality in children with SCID and other primary immunodeficiencies who require BMT. Twenty two of 73 (30%) such children were admitted with respiratory viral infections, of whom 13/22 (59%) died. All viruses were detected in nasopharyngeal aspirate (NPA). Virus was only found in BAL in those with LRTI. Eleven of 22 (50%) had paramyxovirus infections, all with severe viral pneumonitis which worsened post BMT. Five of 11 (45.5%) survived overall. All 11 received aerosolised ribavirin; five of 11 received additional inhaled immunoglobulin and corticosteroid. Three of 5 (60%) survived compared with two of six (33.3%) not thus treated. Three of 22 (13.6%) had adenoviruses; one died of disseminated disease, including pneumonia despite intravenous ribavirin. Eleven patients had rhinovirus detected; nine of 11 (82%) were asymptomatic or coryzal and survived. Two patients with additional severe lung pathologies had LRT rhinovirus and died. All patients received intravenous immunoglobulin. No treatments resulted in viral clearance without successful T cell engraftment. Respiratory viruses, particularly paramyxoviruses and adenoviruses are common, significant pathogens in these patients, significantly worsening outcome of BMT. NPA is an ideal specimen for diagnosis and monitoring of infection. Aggressive treatments may reduce viral replication and damage. Nebulised immunoglobulin and corticosteroid in LRTI may improve respiratory function and outcome. PMID- 11108310 TI - No effect of nadroparin prophylaxis in the prevention of central venous catheter (CVC)-associated thrombosis in bone marrow transplant recipients. AB - Complications of CVCs in 382 consecutive patients receiving a stem cell transplantation (SCT) were analysed. Early complications were pneumothorax (3.6%), haematothorax (0.5%), dislocation (3%) and dysfunction (3.6%). Eighty seven-associated infections (22%) were observed, leading to removal of the CVC in 26 patients. More bacteraemias were associated with double- or triple-lumen CVCs, 19% vs 5% in single lumen CVCs (P < 0.0001). Coagulase-negative staphylococci were the predominant microorganisms in 72%. A special point of investigation was CVC-associated thrombosis and the prophylactic value of nadroparin. Two consecutive regimens with nadroparin were used and compared; 7 days 2850 IE nadroparin and 10 days 5700 IE nadroparin. The incidence of CVC-associated thrombosis was 6.9% in 382 patients with 390 catheters. The incidence was 8% in patients receiving one of the prophylactic nadroparin regimens compared to 6% in a comparable control group without prophylaxis. A short course of nadroparin was unable to prevent thrombotic complications after discontinuation. PMID- 11108311 TI - Melphalan-associated pulmonary toxicity following high-dose therapy with autologous hematopoietic stem cell transplantation. AB - Melphalan can rarely cause interstitial pneumonitis and fibrosis. Although it has been reported previously in patients after conventional doses, we report four cases developing diffuse interstitial pneumonitis (DIP) after high-dose melphalan based therapy. In a 3-year period, four of 57 (7%) consecutive patients undergoing high-dose melphalan (200 mg/m2; MEL 200) were identified with DIP. Two patients who were heavily pre-treated with alkylators developed progressive respiratory failure despite high-dose steroids and eventually died. The other two patients previously treated with vincristine, adriamycin, and dexamethasone (VAD) improved dramatically on high-dose steroids with complete resolution of their pneumonitis. Melphalan should be added to the growing list of alkylators causing pulmonary toxicity. PMID- 11108312 TI - Extramedullary T cell lymphoblastic transformation of chronic myeloid leukaemia successfully treated with matched unrelated donor bone marrow transplantation. AB - Chronic myeloid leukaemia (CML) inevitably terminates in blast crisis (BC) which is of myeloid phenotype in approximately two-thirds and B-lymphoid in one-third of patients. T cell BC is rare and associated with poor prognosis. We describe the case of a 48-year-old woman with extramedullary T cell lymphoblastic transformation. After treatment with combination chemotherapy she achieved a second chronic phase and underwent an allogeneic BMT from an HLA-matched unrelated donor. At 30 months follow-up she is still in complete molecular remission and in good clinical condition. We conclude that unrelated donor BMT should be considered as a therapeutic option for patients with extramedullary BC. PMID- 11108313 TI - Philadelphia-positive T-ALL in a patient with follicular lymphoma. AB - Follicular lymphoma is a B cell malignancy prone to transformation into a large cell diffuse histology. This progression may be multi-clonal as determined by IgH rearrangement. Similar multi-clonal occurrences have been described in immunocompromised patients. However, the lymphoma cells remain predominantly of B cell type. Rarely, composite lymphomas with diffuse T cell histology have been reported arising from follicular lymphoma. The development of T cell leukaemia in a patient with a pre-existing B cell malignancy is an extremely rare event. The occurrence of T cell acute lymphoblastic leukaemia (T-ALL) following follicular lymphoma (FL) has not previously been reported. We report a case of Philadelphia positive (Ph+) T cell ALL developing in a patient who previously had FL which may give some insight into the cell of origin and the defects responsible for malignant transformation of the lymphocytes. PMID- 11108314 TI - Chronic graft-versus-host disease-related polymyositis as a cause of respiratory failure following allogeneic bone marrow transplant. AB - An unusual case of respiratory failure and dropped head syndrome as a complication of severe chronic graft-versus-host disease (GVHD)-related polymyositis is described. The patient required tracheostomy and mechanical ventilation but recovered following treatment with aggressive immunosuppression and intensive rehabilitation. The differential diagnoses of muscle weakness in the bone marrow transplant (BMT) patient and the dropped head syndrome are both discussed. To our knowledge, this is the first reported case of respiratory failure requiring mechanical ventilation occurring as a complication of GVHD related polymyositis. PMID- 11108315 TI - Umbilical cord blood transplantation from unrelated HLA-matched donor in an adult with severe aplastic anemia. AB - A 23-year-old male suffering from severe aplastic anemia (SAA) weighing 60 kg was successfully treated by unrelated allo-CBSCT (cord blood stem cell transplantation). A six-loci HLA-identical umbilical cord blood (UCB) was infused after conditioning with low-dose cyclophosphamide (CTX) and antilymphocyte globulin (ALG). The prophylaxis of GVHD consisted of CsA and MTX. The infused cord blood provided 1.89 x 10(7) nucleated cells per kg, CD34-positive cells: 0.89%. Neutrophils >0.5 x 10(9)/l were reached 10 days after transplant, and platelets greater than 50.0 x 10(9)/l at day 26. RBC and platelet transfusion independence were reached on days 15 and 18. The patient developed grade 1 skin GVHD 10 months after engraftment of the donor cells. Microsatellite DNA fingerprinting indicated a stable and persistent donor-recipient mixed chimerism, whilst the circulating red cells remain of host origin. PMID- 11108316 TI - Spectral karyotyping (SKY) refinement of a complex karyotype with t(20;21) in a Ph-positive CML patient submitted to peripheral blood stem cell transplantation. AB - A patient with a Ph-positive chronic myeloid leukaemia (CML) was submitted to allogeneic peripheral blood stem cell transplantation from an HLA-haploidentical related donor 7 years after the diagnosis. Six months later, he showed a disease relapse while cytogenetic analysis displayed a complex karyotype. To characterise the chromosomal rearrangements spectral karyotype (SKY) analysis was used. This redefined all chromosome rearrangements and revealed a t(20;21)(q11;q22). FISH analysis with a specific probe for the AML1 gene disclosed disruption of this gene which was partially translocated on to the long arm of chromosome 20. It is likely that this rearrangement, unusual for CML, was implicated in the disease evolution towards blastic crisis (BC). PMID- 11108317 TI - Autologous cord blood transplantation. A procedure with potential beyond bone marrow replacement? PMID- 11108318 TI - Aspergillus tracheobronchitis after allogeneic bone marrow transplantation. PMID- 11108319 TI - Extramedullary relapses at uncommon sites after allogeneic stem cell transplantation. PMID- 11108320 TI - Autologous bone marrow transplantation for childhood acute lymphoblastic leukemia in second remission. PMID- 11108321 TI - Frontline transplantation of autologous CD34+ selected blood cells for advanced mantle cell lymphoma: no evidence of long-term cure: a single centre experience. PMID- 11108322 TI - High-dose chemotherapy and autologous peripheral blood stem cell transplantation in lymphoma without blood product support. PMID- 11108323 TI - Stroke: epidemiology, clinical picture, and risk factors--Part I of III. AB - The aim of this review is to present the current knowledge regarding stroke. It will appear in three parts (in part II the pathogenesis, investigations, and prognosis will be presented, while part III will consist of the management and rehabilitation). In the current part (I) the definitions of the clinical picture are presented. These include: amaurosis fugax, vertebrobasilar transient ischemic attack, and stroke (with good recovery, in evolution and complete). The role of the following risk factors is discussed in detail: age, gender, ethnicity, heredity, hypertension, cigarette smoking, hyperlipidemia, diabetes mellitus, obesity, fibrinogen and clotting factors, oral contraceptives, erythrocytosis and hematocrit level, prior cerebrovascular and other diseases, physical inactivity, diet and alcohol consumption, illicit drug use, and genetic predisposition. In particular, regarding the carotid arteries, the following characteristics are analyzed: atheroma, carotid plaque echomorphology, carotid stenosis, presence of ulcer, local variations in surface deformability, pathological characteristics, and dissection. Finally the significance of the cerebral collateral circulation and the conditions predisposing to cardioembolism and to cerebral hemorrhage are presented. PMID- 11108324 TI - Coronary angioplasty and stenting in orthotopic heart transplants: a fruitful act or a futile attempt? AB - Accelerated allograft coronary artery disease remains the major cause of mortality after the first year of transplantation. Despite the extensive use of stents and angioplasty in coronary artery disease, there is a paucity of data about the efficacy of such interventions in orthotopic heart transplants. The authors herein report the outcome of those patients in their institution who had undergone percutaneous coronary artery angioplasty and stenting at a late stage of their transplantation. Within a 12-year period, 106 adult patients underwent orthotopic heart transplantation at their institution. Eight of these patients with 17 lesions underwent deployment of nine stents and eight angioplasties 8.1 +/- 3.2 years posttransplantation. There were 15 denovo and two restenotic lesions. The indications for intervention were presence of symptoms in five patients and severity of lesions in three asymptomatic patients detected on their follow-up angiogram. All patients had angiographic worsening of lesions at their follow-up angiogram. The initial procedural success for both stented and angioplastied lesions was 100%. Within a mean angiographic follow-up of 261 days, all balloon angioplastied lesions had developed restenosis, whereas within a mean period of 67 days, 50% of stented lesions had developed restenosis. On the follow up angiogram, deterioration of the nontreated segments were noted throughout the coronary arterial tree; however, the immediate proximal and distal parts of the target segments demonstrated an exaggerated hyperproliferative response as compared to other sites. The overall median time to the detection of restenosis for both stented and angioplastied lesions was 5.2 months (inner quartile 2.5-6.2 months). The authors conclude that angioplasty and stenting late in the course of transplantation is associated with a significant restenosis rate and in such patients earlier or alternative catheter-based interventions must be considered. PMID- 11108325 TI - The effect of high-protein diets on coronary blood flow. AB - Recent research has demonstrated that successful simultaneous treatment of multiple risk factors including cholesterol, triglycerides, homocysteine, lipoprotein (a) [Lp(a)], fibrinogen, antioxidants, endothelial dysfunction, inflammation, infection, and dietary factors can lead to the regression of coronary artery disease and the recovery of viable myocardium. However, preliminary work revealed that a number of individuals enrolled in the original study went on popular high-protein diets in an effort to lose weight. Despite increasing numbers of individuals following high-protein diets, little or no information is currently available regarding the effect of these diets on coronary artery disease and coronary blood flow. Twenty-six people were studied for 1 year by using myocardial perfusion imaging (MPI), echocardiography (ECHO), and serial blood work to evaluate the extent of changes in regional coronary blood flow, regional wall motion abnormalities, and several independent variables known to be important in the development and progression of coronary artery disease. Treatment was based on homocysteine, Lp (a), C-reactive protein (C-RP), triglycerides, total cholesterol, high-density lipoprotein cholesterol, low density lipoprotein cholesterol, very low-density lipoprotein cholesterol, and fibrinogen levels. Each variable was independently treated as previously reported. MPI and ECHO were performed at the beginning and end of the study for each individual. The 16 people (treatment group/TG) studied modified their dietary intake as instructed. Ten additional individuals elected a different dietary regimen consisting of a "high-protein" (high protein group/HPG) diet, which they believed would "improve" their overall health. Patients in the TG demonstrated a reduction in each of the independent variables studied with regression in both the extent and severity of coronary artery disease (CAD) as quantitatively measured by MPI. Recovery of viable myocardium was seen in 43.75% of myocardial segments in these patients, documented with both MPI and ECHO evaluations. Individuals in the HPG showed worsening of their independent variables. Most notably, fibrinogen, Lp (a), and C-RP increased by an average of 14%, 106%, and 61% respectively. Progression of the extent and severity of CAD was documented in each of the vascular territories with an overall cumulative progression of 39.7%. The differences between progression and extension of disease in the HPG and the regression of disease in the TG were statistically (p<0.001) significant. Patients following recommended treatment for each of the independent variables were able to regress both the extent and severity of their coronary artery disease (CAD), as well as improve their myocardial wall motion (function) while following the prescribed medical and dietary guidelines. However, individuals receiving the same medical treatment but following a high protein diet showed a worsening of independent risk factors, in addition to progression of CAD. These results would suggest that high-protein diets may precipitate progression of CAI) through increases in lipid deposition and inflammatory and coagulation pathways. PMID- 11108326 TI - Identificaiton of chlamydia penumoniae DNA in caroitd plaques. AB - Chlamydia pneumoniae (CP) is a bacterium that in recent years has been investigated as an etiologic agent for atherosclerosis. It is a ubiquitous microorganism that has been isolated in various regions of the vascular system and its prevalence is about 10% in the patient population. This study involved a group of 43 patients (27 men, 16 women, mean age 68 years) who underwent carotid endarterectomy. About 9.3% of the patients yielded plaques that tested positive for the DNA genome of Chlamydia pneumoniae. PMID- 11108327 TI - Hyperinsulinemia, lipoprotein (a), and Chlamydia pneumoniae antibodies--are they risk factors or serologic predictors for progression of coronary artery disease? AB - The authors studied 134 patients with unstable angina pectoris symptoms and 32 subjects without coronary artery disease (CAD) for the presence of classical risk factors such as hypercholesterolemia, smoking, and family history of CAD. In addition they analyzed plasma insulin levels, lipoprotein (a) (Lp [a]) levels, and antibody titers against Chlamydia pneumoniae. All patients had a heart catheterization. Patients with diabetes mellitus were excluded from the study. Fasting insulin, low-density lipoprotein (LDL) cholesterol and Chlamydia pneumoniae immunoglobulin G (IgG) and IgA antibody titers did not show any difference in CAD from healthy control subjects, whereas Lp(a) was increased and high-density lipoprotein (HDL) decreased in CAD patients. These data indicate that lipoprotein (a), low HDL cholesterol, and smoking, but neither hyperinsulinemia nor elevated Chlamydia pneumoniae titers, are risk factors or predictors for CAD. PMID- 11108328 TI - Diffuse and severe left ventricular dysfunction induced by epicardial coronary artery spasm. AB - Endothelial dysfunction and effectiveness of treatment of calcium antagonists are suggestive of coronary artery spasm as an underlying disorder in dilated cardiomyopathy (DCM). The aim of this study is to determine whether or not the epicardial coronary artery spasm can induce severe cardiac dysfunction like DCM. Thirty-four consecutive patients with angiographically normal coronary arteries and diffuse left ventricular hypokinesis whose causes had been unknown underwent acetylcholine provocation test and left ventricular biopsy. Eight patients were excluded according to the clinical and laboratory data and biopsy findings suggesting myocarditis or other systemic diseases. According to the results of the acetylcholine provocation test, 17 patients were finally diagnosed as having DCM, and nine patients (35% of the study patients), who had acetylcholine-induced diffuse and multivessel coronary spasm, were diagnosed as having DCM-like vasospastic angina pectoris (VSA). Clinical and cardiac catheterization data including hemodynamics and biopsy findings were similar between the two groups except that left ventricular end-systolic volume was significantly greater in DCM than in DCM-like VSA. After the acetylcholine provocation test, DCM patients received both a beta blocker and an angiotensin-converting enzyme inhibitor, and DCM-like VSA patients received antianginal drugs. In echocardiographic findings at predischarge and those after 6-month drug treatment, both DCM-lke VSA and DCM showed significant reduction in end-diastolic and end-systolic diameters and significant increase in fractional shortening and ejection fraction, whereas changes in ejection fraction and fractional shortening were significantly greater in DCM-like VSA than those in DCM. Epicardial coronary artery spasm can induce diffuse and severe left ventricular dysfunction like DCM in VSA. Although antianginal drugs markedly improve left ventricular function of these patients, only the acetylcholine provocation test can identify DCM-like VSA. PMID- 11108329 TI - Acute success rates of percutaneous transluminal coronary angioplasty in the treatment of chronic complete occlusions with use of the 0.014 magnum Meier wire. AB - The 0.014 inch magnum Meier wire was used as the primary tool for recanalization of chronic total coronary artery occlusions in 230 consecutive patients treated by a single operator over a 3-year period. Exclusive use of the magnum wire resulted in an acute success rate of 80.9% in all occlusions and 64.7% in occlusions with a duration of >6 months. The complication rate of this procedure was extremely low with only one nontransmural myocardial infarction occurring. There were no vessel perforations, no in-hospital deaths, and no need for acute surgery. After failure to recanalize with the magnum wire, various other devices (conventional stiff guidewires, jagwire, crosswire) were used resulting in only six additional successful recanalizations but also in two vessel perforations with spontaneous closure of the perforation hole. Therefore, the 0.014-inch magnum Meier recanalization wire is highly effective for recanalization of chronic coronary artery occlusions, if used as the primary tool by an experienced operator, and is associated with an extremely low complication rate. PMID- 11108330 TI - Role of activated protein C resistance in left atrial thrombogenesis in patients with mitral stenosis. AB - Activated protein C resistance (APC-R) is the most common inherited cause of clinically apparent venous thromboembolism. Previous data indicate that left atrial thrombus (LAT) formation is a common complication in mitral stenosis (MS) and a hypercoagulable state exists in these patients. The aim of this study was to invastigate the association between APC-R and LAT formation in patients with MS. Seventy-seven consecutive patients with rheumatic MS were included in this study. Transesophageal echocardiography was performed on all patients to assess the presence of any thrombus or spontaneous echo contrast (LASEC) in the left atrial cavity or appendage. Thirty four of the patients had LAT and 43 did not. Prevelance of APC-R was smiliar between the two groups of patients with and without LAT (23% vs 16%, p=0.425). LAT(+) patients had higher incidence of atrial fibrillation (AF, 74% vs 51%, p=0.046) and LASEC (71% vs 19%, p<0.001) compared to LAT(-) patients. On multivariate regression analysis, only the presence of LASEC achieved statistical significance as an independent risk factor for LAT formation (p=0.0001, odds ratio=9.589, 95% confidence interval [CI] =3.143 29.251). Because onunivariate analysis both LASEC and AF were associated with LAT, we also compared the prevelance of APC-R in the subgroups of patients who have these risk factors with and without LAT. There was a correlation between the presence of APC-R and LAT in the AF(+) subgroup of MS patients (p=0.033, odds ratio=8.167, 95% CI=1.001-72.812). However, the presence of APC-R was not associated with the increased risk of LAT in LASEC(+) patients (p=0.217, odds ratio=1.200, 95% CI=1.003-1.435). Although the presence of APC-R itself is not an independent risk factor for LAT formation in MS, it may increase the risk of LAT when present in combination with AF (as an additional risk factor) in these patients. PMID- 11108331 TI - Acute ECG changes and chest pain induced by neck motion in patients with cervical hernia--a case report. AB - We report two cases of acute cervical angina and ECG changes induced by anteflexion of the head. Cervical angina is defined as chest pain that resembles true cardiac angina but originates from cervical discopathy with nerve root compression. In these patients, Prinzmetal's angina, valvular heart disease, congenital heart disease, left ventricular aneurysm, and cardiomyopathy were excluded. After all, the patient's chest pain was reproduced by anteflexion of head, at this time, their ECGs showed nonspecific ST-T changes in the inferior and anterior leads different from the basal ECG. ECG changes returned to normal when the patient's neck moved to the neutral position. To our knowledge, these are the first cases of cervical angina associated with acute ECG changes by neck motion. PMID- 11108332 TI - Abdominal aortic aneurysm infected with Helicobacter pylori--a case report. AB - The authors present a case of an abdominal aortic aneurysm infected with Helicobacter pylori bacteria. In their literature search, the authors found no other report of the Helicobacter pylori involved in an infected abdominal aortic aneurysm. PMID- 11108333 TI - Acute reversible cardiomyopathy and thromboembolism after cisplatin and 5 fluorouracil chemotherapy--a case report. AB - Acute development of cardiomyopathy and occlusive thromboembolic events following cisplatin and 5-fluorouracil (5-FU) is rare but frequently lethal. The authors report the successful management of such an event in a 52-year-old man with squamous cell carcinoma of the soft palate. The possible pathophysiological mechanisms are discussed. PMID- 11108335 TI - Conformity assessment PMID- 11108334 TI - Coronary artery-left ventricle fistula complicating balloon angioplasty--a case report. AB - The authors describe a coronary artery fistula complicated balloon angioplasty. The proximal left anterior descending coronary artery was dilated, but a septal branch was occluded by thrombus. Angioplasty was used on the septal branch, but a pseudoaneurysm communicating with the left ventricle occurred. Follow-up angiography revealed spontaneous closure of the fistula. PMID- 11108337 TI - Transient response of quasi-isotropic fiber composite laminates to internal line sources AB - This article considers the propagation of elastic waves in an eight-ply quasi isotropic laminate arising from line sources of dislocation located at each of the seven interfaces in turn. The laminate is composed of identical layers of a fiber composite material which is modeled as a homogeneous transversely isotropic elastic continuum with the axis of transverse isotropy along the fiber direction. The line source sets up a straight crested wave traveling along the laminate in the direction normal to the load line and the elastodynamic equations within each layer are solved by taking the Laplace transform with respect to time and the Fourier transform with respect to the spatial coordinate in the direction of propagation. The resulting system of six first-order differential equations in each layer is solved to obtain the transforms of the displacement and stress components throughout the laminate. The time history of any displacement or stress component at any location may then be recovered by numerical inversion of the double transform. The graphs presented show the time history of the normal displacement of the top surface of the laminate at distances of 1 and 20 plate thicknesses from the plane of action of the sources. These graphs are for four different orientations of the line of action of the sources, namely, at angles 0, 30, 60, and 90 degrees to the fiber direction in the surface layer. The numerical inversion involves a summation over different modes of Rayleigh-Lamb waves in the laminate and results are also presented showing the contributions to the overall response from some of the individual modes. PMID- 11108338 TI - The effect of moisture on compressional and shear wave speeds in unconsolidated granular material AB - The effect of water vapor on the shear and compressional wave speeds in two different kinds of glass beads and in Ottawa sand has been measured. The nominal diameter of the glass beads was 125 microm and of the sand, 500 microm. Measurements were made as the water vapor was introduced slowly into the evacuated material. The vapor pressure isotherm for the beads made of glass with a high concentration of titanium and barium oxides was fit reasonably well by the simple Brunauer-Emmett-Teller (BET) theory. For the Ottawa sand, the BET theory fit the vapor pressure isotherm if the surface area of the grains was assumed to be three times the area calculated, assuming all of the grains were spheres with a diameter of 500 microm. In these two materials, the vapor had little effect on the wave speeds. For beads made of glass containing sodium oxide, however, the wave speeds approximately double with the introduction of water vapor, and the vapor pressure isotherm had the BET shape only if the saturated vapor pressure was assumed to be lowered by 20%. These results have been explained by assuming that a chemical reaction occurred between the lime glass and the water to form a gel. PMID- 11108339 TI - On ultrasonic guided waves in a thin anisotropic layer lying between two isotropic layers AB - In this paper, dispersion of guided waves in a three-layered sandwich plate is considered. The focus here is on a configuration consisting of a thin anisotropic layer sandwiched between two identical isotropic layers. This configuration could model, for example, a superconducting tape where the middle layer is a brittle superconductor and the surrounding layers are isotropic and ductile. An approximate dispersion relation correct to O(h) governing the guided waves is obtained by expanding the field inside the thin middle layer in powers of the small thickness h of the layer. Numerical examples are given for two specific systems with superconducting middle layers. Some characteristic features, particularly at low frequencies, are investigated. Comparison between the exact and approximate dispersion relations are made to show that the approximation works well in the frequency interval of interest. The characteristic features may be useful for ultrasonic measurements of the anisotropic elastic constants of the thin layer. PMID- 11108340 TI - Ultrasonic cavitation monitoring by acoustic noise power measurement AB - In this paper, a new tool is proposed to carry out acoustic cavitation monitoring and to have an overview of its effects in applications. After a brief review of the cavitation characterization techniques, it is shown that cavitation noise is a suitable and accurate indicator of the cavitation activity induced in a liquid. In the first part of this study, the origin of the first spectral component of the cavitation noise is discussed. The f/2 and 2 f component evolution measurement at a driving frequency around 1 MHz confirms Neppiras' ones and gives an indicator of the cavitation inception. In the second part, the cavitation noise spectrum distortion is considered as a function of the acoustic power transmitted to the liquid in order to obtain an indicator of cavitation activity. In the last part, this new tool is used to bring to the fore the hysteresis effect associated with the cavitation. An experimental correlation between cavitation noise power measurement and the sonochemical activity in an oxidization process is also presented. PMID- 11108341 TI - A turbulence spectral model for sound propagation in the atmosphere that incorporates shear and buoyancy forcings AB - A three-dimensional model for turbulent velocity fluctuations in the atmospheric boundary layer is developed and used to calculate scattering of sound. The model, which is based on von Karman's spectrum, incorporates separate contributions from shear- and buoyancy-forced turbulence. New equations are derived from the model that predict the strength and diffraction parameters for scattering of sound as a function of height from the ground and atmospheric conditions. The need is demonstrated for retaining two distinct scattering length scales, one associated with scattering strength and the other with diffraction. These length scales are height dependent and vary substantially with the relative proportions of shear and buoyancy forcing. The turbulence model predicts that for forward-scattered waves the phase variance is much larger than the log-amplitude variance, a behavior borne out by experimental data. A new method for synthesizing random fields, based on empirical orthogonal functions, is developed to accommodate the height dependence of the turbulence model. The method is applied to numerical calculations of scattering into an acoustic shadow zone, yielding good agreement with previous measurements. PMID- 11108342 TI - Evaluation of sound propagation models used in bottom volume scattering studies AB - The proper evaluation of sound propagation between sources/receivers and scatterers is important in characterizing bottom volume scattering. In this article, several sound propagation models used in bottom volume scattering studies are evaluated and their results compared to the exact solution obtained through a numerical wave number integration technique. It is found that Hines' approach [J. Acoust. Soc. Am. 88, 324-334 (1990)] works well for the two isovelocity half-space case except when the grazing angle is close to the critical angle. The far-field approximation, given by Ivakin [Sov. Phys. Acoust. 32(6), 492-496 (1986)] and Mourad and Jackson [J. Acoust. Soc. Am. 94, 344-358 (1993)], has a performance depending upon the sound speed structure in the sediment. For an isovelocity slow bottom, it agrees well with the exact solution. However, discrepancies arise for an isovelocity fast bottom or a bottom with a complex sound speed structure. In addition, the appropriateness of using the equivalent surface scattering strength as a function of grazing angle in volume scattering characterizations is studied. In conclusion, precautions need to be taken in modeling both the propagation effects and the scattering mechanisms associated with the bottom volume scattering process. PMID- 11108343 TI - A comparison of bistatic scattering from two geologically distinct abyssal hills AB - The bistatic scattering characteristics of two geologically distinct abyssal hills located on the western flank of the Mid-Atlantic Ridge, known as B' and C', are experimentally compared using data acquired with low-frequency towed-array systems at 1/2 convergence zone (approximately 33 km) stand-off. The comparison is significant because the abyssal hills span the two classes of elevated seafloor crust that cover the Mid-Atlantic Ridge. The highly lineated B' feature is representative of abyssal hills composed of outside corner crust, the most commonly occurring category, whereas the domed C' promontory is representative of the rougher, low-aspect-ratio abyssal hills composed of inside corner crust. The latter are less common and usually restricted to segment valley margins. The mean biazimuthal scattering distributions of the two abyssal hills each exhibit Lambertian behavior with comparable albedos, suggesting that the distinction between abyssal hills composed of differing crust is not significant in modeling long-range reverberation. The adverse effect of using bathymetry that undersamples seafloor projected area in scattering strength analysis is also quantified with data from the B' ridge. Specifically, the use of undersampled bathymetry can lead to significant overestimates in the strength of seafloor scattering. PMID- 11108344 TI - Penetration of acoustic waves into rippled sandy seafloors AB - The Helmholtz-Kirchhoff integral and the Kirchhoff approximation are applied to model the penetration of sound waves into rough sandy seafloors at grazing angles above and below the critical angle. As the seafloor of interest is anisotropic, emphasis is placed on simulating the response from a two-dimensional interface. The analytical development of the method is first presented, followed by numerical examples. Simulations and data acquired at sea are in very good agreement in the 2-15 kHz band [Maguer et al., J. Acoust. Soc. Am. 107, 1215-1225 (2000)]. The model predicts, in agreement with the 2-15 kHz acoustic data, the contributions due to roughness effects that dominate the evanescent wave component over most of this frequency band. Secondary effects such as coherent (Bragg) influence patterns and the loss of signal coherence with grazing angle or depth are correctly predicted. The model simulations strongly suggest that roughness of the sediment interface is most likely the cause of anomalous sound penetration into the seabed. PMID- 11108345 TI - Matched-impulse-response processing for shallow-water localization and geoacoustic inversion AB - In this paper, impulse response matching is proposed for source localization and environmental inversion. The ocean impulse response is estimated using a cross correlation procedure applied to data from the propagation of a broadband pulse in a shallow-water environment. Source localization and geoacoustic parameter estimation are then performed through time-domain correlations between the estimated impulse responses at spatially separated phones and synthetic replica impulse responses. The method is both spatially and temporally coherent. Parameter space search uses a hierarchical scheme designed to exploit the sensitivity of the acoustic field to the unknown parameters. Tested on the SWellEX-96 and synthetic data, the proposed method is shown to be more robust than conventional (linear), incoherent, broadband matched field processing. PMID- 11108346 TI - Kramers-Kronig relations applied to finite bandwidth data from suspensions of encapsulated microbubbles AB - In this work, the Kramers-Kronig (K-K) relations are applied to experimental data of resonant nature by limiting the interval of integration to the measurement spectrum. The data are from suspensions of encapsulated microbubbles (Albunex) and have the characteristics of an ultrasonic notch filter. The goal is to test the consistency of this dispersion and attenuation data with the Kramers-Kronig relations in a strict manner, without any parameters from outside the experimental bandwidth entering in to the calculations. In the course of reaching the goal, the artifacts associated with the truncation of the integrals are identified and it is shown how their impacts on the results can be minimized. The problem is first approached analytically by performing the Kramers-Kronig calculations over a restricted spectral band on a specific Hilbert transform pair (Lorentzian curves). The resulting closed-form solutions illustrate the type of artifacts that can occur due to truncation and also show that accurate results can be achieved. Next, both twice-subtracted and lower-order Kramers-Kronig relations are applied directly to the attenuation and dispersion data from the encapsulated microbubbles. Only parameters from within the experimental attenuation coefficient and phase velocity data sets are used. The twice subtracted K-K relations produced accurate estimates for both the attenuation coefficient and dispersion across all 12 data sets. Lower-order Kramers-Kronig relations also produced good results over the finite spectrum for most of the data. In 2 of the 12 cases, the twice-subtracted relations tracked the data markedly better than the lower-order predictions. These calculations demonstrate that truncation artifacts do not overwhelm the causal link between the phase velocity and the attenuation coefficient for finite bandwidth calculations. This work provides experimental evidence supporting the validity of the subtracted forms of the acoustic K-K relations between the phase velocity and attenuation coefficient. PMID- 11108347 TI - Transverse curvature of the acoustic slowness surface in crystal symmetry planes and associated phonon focusing cusps AB - Conditions are derived for the existence of focusing cusps in ballistic phonon intensity patterns for propagation directions in crystal symmetry planes. Line caustics are known to be associated with lines of vanishing Gaussian curvature (parabolic lines) on the acoustic slowness surface, while cusps are associated specifically with points where the direction of vanishing principal curvature is parallel to the parabolic line. A parabolic line meets a crystal symmetry plane sigma at a right angle, and so it is the vanishing of the slowness-surface curvature transverse to sigma that conditions the existence of a cusp. A relation for the transverse curvature is derived and analyzed. It is shown that in an arbitrary symmetry plane sigma there may be up to four pairs of inversion equivalent cuspidal points for SH (out-of-plane polarized) waves, and up to eight pairs of cuspidal points associated with the in-plane polarized (usually quasi transverse) waves. In tetragonal crystals, the symmetry planes containing the four-fold axis can have at most two pairs of cusps for the SH waves and up to six pairs of cusps for the in-plane waves. In cubic crystals, the face symmetry planes sigma cannot have cuspidal points for SH waves, as is known, while four pairs of cusps for in-plane waves exist in sigma if and only if the outer-most slowness sheet has a concave region embracing the four-fold axis. The points of vanishing transverse curvature on the slowness surface in symmetry planes of tetragonal and cubic media are identified by concise relations, facilitating their explicit analysis. PMID- 11108348 TI - On a time-domain representation of the Kramers-Kronig dispersion relations AB - The development of Kramers-Kronig dispersion relations is typically carried out in the frequency domain. An alternative approach known as the time-causal theory develops dispersion relations for media with attenuation obeying a frequency power law through analysis in the time domain [T. L. Szabo, J. Acoust. Soc. Am. 96, 491-500 (1994)]. Although both approaches predict identical dispersion relations, it is perceived that these two approaches are distinct from each other. It is shown, however, that the time-causal theory is in essence a time domain formulation of the Kramers-Kronig dispersion relations for the special case of media with attenuation obeying a frequency power law. Additionally, it is shown that time-domain representations of the Kramers-Kronig dispersion relations are available for a broader class of media than simply those with power law attenuation. The time-causal theory and the Kramers-Kronig dispersion relations can be viewed as two complementary, yet equivalent, approaches to the study of dispersion. PMID- 11108349 TI - The effect of the physical properties of the tube wall on the attenuation of sound in evaporating and condensing gas-vapor mixtures AB - An investigation of sound propagation in an air-water vapor mixture contained in a cylindrical tube with wet walls was recently completed [Hickey et al., J. Acoust. Soc. Am. 107, 1126-1130 (2000)]. A generalization to include the heat flux at the tube wall is presented here. The attenuation of sound in air-water vapor mixtures can be affected by the thermal properties of the tube wall. The controlling parameter is epsilons, which is a proportionality constant that relates the heat flux per degree Kelvin for the substrate to that of the gas mixture. For a given amount of heat, provided by expansion and rarefaction of the working fluid, different substrates will undergo different temperature excursions. These temperature swings at the boundary change the vapor pressure of the condensate and thus reduce the diffusion of vapor to and from the boundary resulting in a reduction of the attenuation. PMID- 11108350 TI - Impedance and admittance matrices of symmetric piezoelectric annular bimorphs and their applications AB - The 5 x 5 impedance and admittance matrices of a symmetric triple-layer piezoelectric annular bimorph (PAB) are presented and the PAB with the piezoelectric layers partially covering the shim layer in harmonic motion is analyzed using the matrices. The electromechanical flow vectors are defined as the mechanical displacements and rotations at the inner and outer boundaries and the electric current of the piezoelectric layer; the conjugate parameters, i.e., the electromechanical effort vectors, are accordingly defined. The impedance matrix which relates the flow vector with the effort vector is divided into three matrices, the mechanical, electrical, and electromechanical coupling impedance matrices; each matrix is represented by a block network with five ports. The resonance and the antiresonance frequencies and the effective electromechanical coupling factors of the PAB excited by the partially covering piezoelectric layers are then calculated for various boundary conditions imposed at the inner and outer surfaces. The numerical results by the impedance matrix are compared with those by the finite element methods; they are in excellent agreement with each other. It is also shown that the matrix representations can be easily applied to the piezoelectric circular bimorph (PCB) with the extended shim layer. PMID- 11108351 TI - Measured capacitance of a condenser microphone as a function of diaphragm displacement AB - This paper shows that the capacitance of a polarized condenser microphone measured with a sinusoidal electrical signal is quantitatively related to the displacement of the microphone diaphragm. In fact, the capacitance measured with an ac signal is the "input capacitance" of the microphone when it acts as a transmitter. The expressions for the diaphragm displacement and the input capacitance are derived with damping taken into account. The input capacitance differs from the static capacitance with bias by an additional term which varies with the polarization voltage and frequency. Therefore, in general, it is impossible to measure the static capacitance with bias using a low frequency ac signal without the diaphragm being blocked. In order to measure the true static capacitance with bias, a static method based on the dc voltage distribution is presented. The experimental results for the input capacitance and the static capacitance with bias, obtained with the abovementioned ac and dc methods, respectively, give a high level of confidence in the theory. The experimental results reported by some others also support the theory. PMID- 11108352 TI - A method for measuring the diaphragm tension of condenser microphones using electric admittance AB - This paper presents a method to measure the diaphragm tension of a condenser microphone where the fundamental frequency of the diaphragm is found from the input conductance of the microphone as a function of frequency. In this contact free, purely electrical approach only the diaphragm and the backplate of the microphone are used as the main system for exciting and detecting the diaphragm vibration. PMID- 11108353 TI - An acoustic/thermal model for self-heating in PMN sonar projectors AB - Dielectric hysteresis and a strong material temperature dependence uniquely couple the acoustic output and temperature of a sonar projector driven by electrostrictive Pb(Mg1/3, Nb2/3)O3 (PMN). Both the source level and the source of self-heating, i.e., dielectric hysteresis, dramatically decrease as the PMN driver heats. The final temperature delineates outstanding PMN transducers from mediocre PMN transducers, so accurate acoustic performance prediction requires accurate transducer temperature prediction. This study examined this self-heating phenomenon by combining an electro-acoustics model for a PMN flextensional transducer with a thermal finite element model. The sonar model calculated the source level and heat generation rate for the PMN driver as a function of temperature. This computed source level varied 12 dB over a 75 degrees C temperature range solely due to the temperature dependent ceramic. The heat transfer model used the computed heat rate to predict the transducer's transient thermal response. The results clearly demonstrate that the transducer reached a steady-state equilibrium temperature, where the heat generated by the PMN driver balanced the heat dissipated. While the transducer model predicted a significant temperature rise, the corresponding acoustic output still surpassed the output of an equivalent Pb(Zr,Ti)O3 (PZT) transducer by 8 dB. Good agreement with experiments made on a PMN flextensional transducer validated the model. PMID- 11108354 TI - Novel graph model and analysis method for piezoelectric thickness-drive transducers AB - This study develops a new graph model and analysis scheme for representing and analyzing a piezoelectric thickness-drive transducer. The proposed TWSF graph model, expressed via electrical and acoustical variables instead of electromechanical analogy, simplifies understanding of the physical mechanism of the transducer. The primary advantage of the graph model analogy is that performance characteristics, such as the electrical and mechanical impedance, and the force and velocity responses of the transducer system can be calculated directly using the topology scheme based on the graph model. The dynamic equations do not need to be formulated according to the analogy model for design and analysis. The proposed method also does not require computation schemes to solve the equations for obtaining the characteristics and the responses. Compared to the classical method, the present graph and analysis scheme offers a more efficient technology to obtain precise results with less analysis. Moreover, the derived results are expressed as both analytical and closed-form formulas, making them convenient to apply in analysis and design. PMID- 11108355 TI - A computational acoustic field reconstruction process based on an indirect boundary element formulation AB - The objective of the work presented in this paper is to develop a computational capability based on the indirect boundary element method (IBEM) for evaluating appropriate velocity boundary conditions on an assembly of piston type sources in order to recreate a prescribed acoustic field. Information for the acoustic pressure of the original acoustic field at certain field points constitutes the input to the developed process. The velocities on the piston type sources are computed from transfer functions evaluated between the field points where the acoustic pressure of the original field is prescribed and the velocity boundary condition on each element of the generic source. The IBEM is employed for computing the transfer functions in order to accommodate the presence of openings in the model and radiation from both sides of the piston type sources. Simulating the generic source as a thin surface that radiates from both sides eliminates the presence of irregular frequencies in the analysis. A singular value decomposition (SVD) solver is integrated with the IBEM computations in order to evaluate the velocity boundary conditions from the transfer functions. The number of field points where the acoustic pressure is defined can be considerably smaller than the number of elements where the velocity is computed. The solution that demonstrates the smallest magnitude is selected from all possible solutions. An algorithm is also developed for identifying the optimum field points where the acoustic pressure of the original field must be prescribed. The optimum field points are selected from a set of prescribed candidate points. The number of optimum points is considered smaller than the number of elements where the velocity is computed. The properties of the transformation matrix and the quality of the reconstruction depend on the location of the field points. Thus, the selection of the optimum points is based on achieving the highest possible orthogonality among the vectors that comprise the range of the transformation matrix. Several validation and application cases are presented. PMID- 11108356 TI - Dispersion and characteristic surfaces of waves in laminated composite circular cylindrical shells AB - The dispersion behaviors and characteristic surfaces of waves in a laminated composite circular cylindrical shell are investigated using a semianalytical method based on the theory of three-dimensional elasticity. The radial displacement of the shell is modeled by finite elements, while the axial and circumferential displacements are expanded as the complex exponentials. The associated characteristic equation is developed by means of the Hamilton's principle. The eigenvalues are established in terms of the Rayleigh quotient. Six characteristic wave surfaces, viz., the phase velocity, phase slowness, and phase wave surfaces, as well as the group velocity, group slowness, and group wave surfaces, are introduced to visualize the effects of anisotropy on wave propagation. Numerical examples demonstrate that the ratio of the inner radius to the thickness of the shell has a stronger influence on the frequency spectra in the circumferential wave than on that in the axial wave; that negative group velocity appears at a range of smaller wave numbers and the range varies as the wave normal and the ratio of the inner radius to the thickness of the shell; and that the characteristic wave surfaces vary with the propagation modes of waves, the ratio of the inner radius to the thickness of the shell, and the lay-ups of the laminated shells. PMID- 11108357 TI - Analysis of the acoustic scattering at variable incidences from an extra thin cylindrical shell bounded by hemispherical endcaps AB - Through an experimental approach, in this paper we investigate the acoustic wave scattering processes involved in the acoustic backscattering at variable incidences from an air-filled submerged cylindrical shell with hemispherical endcaps. Given the 1% shell thickness and the explored low frequency domain, the wave types studied are the circumferential or helical S0 wave and the helical T0 wave only. Between the axial (in the direction of the main axis of the object) and the normal incidences (normal to the main axis), two distinct angular zones can be observed depending on hemispherical or cylindrical excitation. In these zones, after a pressure wave excitation, different series of echoes on the echo wave forms are identified by their arrival times and related wave types. From results in the time domain and those obtained in the frequency domain, each acoustic response from the target corresponding to the two zones of excitation is compared with the acoustic response of canonical objects (spherical shell for axial excitation and tube for normal excitation). This analysis of the acoustic response from the target at various incidences, highlights the influence of both the endcaps and the finite length for a cylindrical shell on scattering. The study is intended to make a contribution to the knowledge of the identification of such geometrically complex objects. PMID- 11108358 TI - Acoustic radiation due to the inelastic impact of a sphere on a rectangular plate AB - Impact between two structures is an important source of noise in industry. However, it is not a well-known phenomenon from a theoretical point of view. The models available in the literature for impact noise often address only part of the phenomenon. In this paper, a more comprehensive model is developed. The acoustic radiation due to the inelastic impact of a sphere with a rectangular simply supported thin plate is formulated and validated experimentally. The impact force is calculated from Hertz's law, which has been extended to the case of plastic deformations, and the acoustic radiation in the time domain is obtained using Rayleigh's integral, the plate being discretized in rectangular elements. The model is used to show that the sound-radiation mechanism includes two separate sources: the initial deformation of the plate and the propagation of the bending waves. The model is also used to determine the contribution to impact noise of parameters that characterize the two impacting structures or the impact configuration such as plasticity, damping, materials, and impact velocity. PMID- 11108359 TI - Active control of coupled structural/acoustic intensities in a fluid-loaded elastic plate AB - In this paper, active control of structural intensity and radiated acoustic power in a coupled structure-fluid system is explored. The response of an infinite fluid-loaded plate and the resultant acoustic pressure field are calculated using a hybrid analytic/numerical approach. Two control strategies are examined: (1) the minimization of the total input power, and (2) the minimization of a weighted sum of structural intensity and the radiated acoustic power. Below coincidence, the vibration response of the plate is dominated by nondecaying waves, thus it can be reduced very efficiently by the active control strategies of minimizing the total input power or minimizing the structural intensity. Below coincidence, the acoustic radiation is caused by fast decaying structural waves thus, and active control strategies that aim to minimize the structural intensity have little impact on the radiated pressure field. Furthermore, the control strategy for minimizing the radiated acoustic power does not perform satisfactorily below coincidence since the controllers cannot be coupled to the fast decaying waves. Above coincidence, a slowly decaying structural wave dominates both the plate response and the acoustic radiation, and thus the structural intensity and the radiated acoustic power can be reduced significantly by each of the two control strategies. PMID- 11108360 TI - Reducing seat dip attenuation AB - Strategies for reducing seat dip attenuation in concert halls are considered. It is shown that the dip is established 4 ms after the direct sound from the stage arrives at the listener. Sound scattered from the seats and floor is the main cause of the dip. By controlling these very early reflections the attenuation can be reduced to below its subjective threshold. With this in mind, changes to the shape and impedance of the seats and floor are trialed using a boundary element model and a physical scale model. It is found that the seat dip effect can be rendered inaudible by introducing a 1-m pit under the seats. Smaller improvements are produced by changing the impedance of the seat squab. PMID- 11108361 TI - Image shift caused by strong lateral reflections, and its relation to inter-aural cross correlation AB - Conditions of sound fields were studied to avoid the occurrence of image shift, an acoustical phenomenon caused by extremely strong lateral reflections. Three types of image shift are defined as variations of the normal condition, in which there is only one auditory image perceived in the source direction. From a series of listening tests in electrically reproduced sound fields, each of the three types of image shift was found to have its own specific frequency range in which the image shift occurred. They were also found to relate to each of three measures which are all derived from the inter-aural cross correlation function (IACF) of binaural impulse responses of a sound field. The results of measurement made in three existing concert halls suggested acceptable ranges for the measures, in which the image in the source direction is not lost in the 500 Hz and higher frequency bands. However, the 250 Hz and lower frequency bands were found not so important in evaluating sound fields of concert halls in relation to the image shift. Finally, 0.85 is found to be the upper limit for the preferable range of [1 - IACC(E3)]. PMID- 11108362 TI - Method to resolve microphone and sample location errors in the two-microphone duct measurement method AB - Utilizing the two-microphone impedance tube method, the normal incidence acoustic absorption and acoustic impedance can be measured for a given sample. This method relies on the measured transfer function between two microphones, and the knowledge of their precise location relative to each other and the sample material. In this article, a method is proposed to accurately determine these locations. A third sensor is added at the end of the tube to simplify the measurement. First, a justification and investigation of the method is presented. Second, reference terminations are measured to evaluate the accuracy of the apparatus. Finally, comparisons are made between the new method and current methods for determining these distances and the variations are discussed. From this, conclusions are drawn with regards to the applicability and need for the new method and under which circumstances it is applicable. Results show that the method provides a reliable determination of both microphone locations, which is not possible using the current techniques. Errors due to inaccurate determinination of these parameters between methods were on the order of 3% for R and 12% for Re Z. PMID- 11108363 TI - Acoustic absorption measurement of human hair and skin within the audible frequency range. AB - Utilizing the two-microphone impedance tube method, the acoustic absorption of human skin and hair is measured in the frequency range 1-6 kHz. Various locations on a number of human subjects are measured to determine if the presence of bone or an air pocket affects the acoustic absorption of human skin. The absorption coefficient of human hair is also measured. Additional techniques are utilized to minimize errors due to sample mounting methods. Techniques are employed to minimize potential errors in sensor and sample locations. The results of these measurements are compared to relevant historical papers on similar investigations. Results for skin measurements compare well with previous work. Measured hair absorption data do not agree with previous work in the area but do coincide with expected trends, which previous works do not. PMID- 11108364 TI - Measurements of anisotropic sound propagation in glass wool AB - The attenuation coefficient and phase velocity of plane sound waves propagating in three perpendicular directions in glass wool were measured in the frequency range 50-10,000 Hz. For glass wool of mass density 14 kg/m3 at the frequency 1,000 Hz, the attenuation constant for propagation perpendicular to the glass wool sheets was 75 dB/m, and for propagation parallel with the sheets 57 dB/m. For mass density 30 kg/m3, the corresponding numbers were 140 and 100 dB/m. The measured values were compared with calculated ones taking into account the movements of the fiber skeleton. The calculations need the elastic moduli, which were measured. For density 14 kg/m3 and deformation perpendicular to the glass wool sheets, the static modulus was 2.0 kPa, and for parallel deformation 120 kPa. The corresponding numbers for mass density 30 kg/m3 were 16 and 390 kPa. PMID- 11108365 TI - Optimum near-field performance of microphone arrays subject to a far-field beampattern constraint AB - This article addresses the problem of maximizing the near-field gain of a microphone array subject to a constraint on the far-field beampattern. The problem arises when acquiring speech from a near-field talker in the presence of a strong source of interference located farther from the array. When the angles of incidence from the near-field target and the far-field interference are identical, enforcing a null constraint in the interference direction reduces array gain and robustness. This article shows how to mitigate this effect by selection of the constrained response level. A suitable selection is to force the beampattern in the interference direction to be proportional to the unconstrained beampattern. The proportionality constant can then be used to trade off interference reduction and array gain. Specific numerical examples are provided. PMID- 11108366 TI - Adaptive beamforming at very low frequencies in spatially coherent, cluttered noise environments with low signal-to-noise ratio and finite-averaging times AB - Realistic simulations with spatially coherent noise have been run in order to compare the performance of adaptive beamforming (ABF), inverse beamforming (IBF), and conventional beamforming (CBF) for the case of finite-averaging times, where the actual spatial coherence of the acoustic field, or covariance matrix, is not known a priori, but must be estimated. These estimation errors cause large errors in the ABF estimate of the directionality of the acoustic field, partly because ABF is a highly nonlinear algorithm. In addition, it is shown that ABF is fundamentally limited in its suppression capability at very low frequency (VLF), based on the sidelobe level of the conventional beampattern in the direction of the noise interferer [G. L. Mohnkern, "Effects of Errors and Limitations on Interference Suppression," NOSC Technical Document 1478, Naval Ocean Systems Center (1989)]. The simulations include a low-level plane wave signal of interest, a stronger noise plane wave interferer, and spatially random background noise. Both IBF and ABF performed significantly better than CBF, and IBF's performance was slightly better than ABF's performance. The performances of IBF and the ABF algorithm, the minimum variance distortionless response (MVDR) [A. H. Nuttall and D. W. Hyde, "Unified Approach to Optimum and Suboptimum Processing for Arrays," USL Report Number 992, Naval Underwater Systems Center, New London, CT (22 April 1969)] were recently compared independently [J. S. D. Solomon, A. J. Knight, and M. V. Greening, "Sonar Array Signal Processing for Sparse Linear Arrays," Defense Science and Technology Organization (DSTO) Technical Report (June 1999)] using measured data, with the result that IBF outperformed MVDR. This result is significant because MVDR requires orders of magnitude more processing power than IBF or CBF. PMID- 11108367 TI - The mode-coupling Liouville-Green approximation for a two-dimensional cochlear model. AB - The Liouville-Green [or Wentzel-Kramers-Brillouin (WKB)] approximation for the two-dimensional cochlear mechanics problem disagrees with the finite-difference solution in the region after the response peak. This disagreement has left doubts about the validity of the Liouville-Green approximation, and has never been satisfactorily explained. In this paper, it is shown that the Liouville-Green approximation fails to satisfy Laplace's equation. A new solution is proposed, called the mode-coupling Liouville-Green approximation, in which energy is coupled into a second wave mode, so as to obey Laplace's equation. The new approximation gives excellent quantitative agreement with the finite-difference solution. Furthermore, it may provide an explanation for a second vibration mode observed in biological cochleas. Also proposed is a high-order formulation of the stapes displacement term, which is necessary to obtain good agreement between the Liouville-Green approximation and finite-difference solutions at low frequencies. PMID- 11108368 TI - Noise-evoked otoacoustic emissions in humans. AB - Click-evoked otoacoustic emissions (CEOAEs) and acoustical responses evoked by bandlimited Gaussian noise (noise-evoked otoacoustic emissions; NEOAEs) were measured in three normal-hearing subjects. For the NEOAEs the first- and second order Wiener kernel and polynomial correlation functions up to the sixth order were calculated by cross correlating the noise stimulus and the emission response. Emission components were only found in the first-order Wiener kernel. These kernels were dependent on the stimulus level, indicative of the nonlinear response of the cochlea. For each subject the frequency contents of the CEOAE and the first-order Wiener kernel were nearly identical. In the second-order Wiener kernels and higher-order polynomial correlation functions no significant contributions were identified. Apparently, odd and even order nonlinearities cannot be detected by Wiener kernel analysis. PMID- 11108369 TI - Basilar membrane responses to broadband stimuli. AB - Basilar membrane (BM) responses to two types of broadband stimuli-clicks and Schroeder-phase complexes--were recorded at several sites at the base of the chinchilla cochlea. Recording sites (characteristic frequency, CF, in the range of 5.5-18 kHz) span the 1-4-mm basal region of the basilar membrane. BM responses to clicks consisted of undamped oscillations with instantaneous frequency that increased over time until it reached a value around CF. The time constant of this glide is CF dependent. Throughout the entire region under study, BM vibration exceeded umbo motion by up to 60 dB. Nonlinear properties of BM responses to clicks resemble those found in the more studied 8-10-kHz region. Amplitude spectra of Schroeder-phase complex stimuli, which consist of a series of sinusoidal components summed in negative (-SCHR) and positive Schroeder phase (+SCHR), are flat. The envelope of BM responses to +SCHR stimuli contains valleys, or dips, that are wider than those found in responses to the -SCHR stimuli. Hence, BM responses to the former stimuli are "peakier" than responses to the latter. Differences in response waveforms are less obvious in linear cochleae. Suppression of a near-CF tone by -SCHR stimuli was larger than that evoked by +SCHR stimuli. PMID- 11108370 TI - Effect of turgor pressure on outer hair cell motility. AB - The motor in the outer hair cell converts electrical energy directly into mechanical energy. There are two possible mechanisms for such a motor: one depends on changes in the membrane area ("area motor") and the other on changes in the stiffness ("stiffness motor"). These two mechanisms, which are not mutually exclusive, give different predictions on turgor pressure dependence of the amplitude. It was found that an increased pressure shifts the voltage dependence but does not change the amplitude of length changes. This observation is incompatible with a pure stiffness motor model. It was also confirmed that length changes are closely related to charge movements by monitoring the membrane capacitance. It can be concluded that hair cell motility is primarily based on area changes, and not changes in the elastic moduli. PMID- 11108371 TI - Effects of hearing impairment and presentation level on masking period patterns for Schroeder-phase harmonic complexes. AB - Masking period patterns (MPPs) for Schroeder-phase harmonic complexes containing equal-amplitude harmonics of a 100-Hz fundamental were determined for 5-ms tonal probes at 4,000 and 1,000 Hz. Maskers consisted of harmonics 2-50 (200-5,000 Hz bandwidth) for 4,000-Hz probes and harmonics 2-20 (200-2,000 Hz) for 1,000-Hz signals. Masked thresholds were determined for probe onsets 153, 155.5, 158, 160.5, and 163 ms following masker onset (masker duration=460 ms). Overall, results were similar for both probe frequencies. For listeners with normal hearing, MPPs for positive Schroeder-phase complexes masking 60 dB SPL probes were highly modulated and became flatter when probe level was increased to 80 dB SPL. MPPs were less modulated for listeners with sensorineural hearing loss than for normally hearing listeners at both 60 and 80 dB SPL probe levels. Thresholds in negative Schroeder-phase maskers were more similar across the two groups of listeners and across differences in probe position and probe level. The findings support an interpretation involving differences in the shape of the basilar membrane waveform generated by each masker and possible influences of nonlinear cochlear processing on these internal responses. For normally hearing listeners, 60 dB SPL probes were most difficult to detect when temporally positioned so that probe frequency and masker instantaneous frequency were closely matched. For 80 dB SPL probes and for hearing-impaired listeners, probes presented at these same positions were often more easily detected than probes at other positions. The latter result appears to involve benefit associated with in-phase addition of the probe to a portion of the masker similar to the probe in both frequency and phase. This benefit was reduced or entirely eliminated when probe phase was altered so that this in-phase addition did not occur. PMID- 11108372 TI - Frequency selectivity as a function of level and frequency measured with uniformly exciting notched noise. AB - Thresholds for detecting sinusoidal signals were measured as a function of the spectral width of a notch in a noise masker. The notch was positioned both symmetrically and asymmetrically around the signal frequency. The noise was designed to create equal excitation per ERB within its passbands (uniformly exciting noise), after allowing for the transfer function of the headphone and the middle ear. For a signal frequency of 250 Hz, the level per ERB ranged from 35 to 80 dB in 15-dB steps. For signal frequencies of 500, 1,000, 2,000, and 4,000 Hz, the level per ERB ranged from 40 to 70 dB per ERB in 15-dB steps. Auditory filter shapes were derived from the data by modeling the auditory filter as the sum of a sharply tuned tip filter and a broader tail filter. The gain of the tip filter was assumed to be a function of level. The shape of the tip filter and the gain and shape of the tail filter were assumed to be level independent. The data for all levels were fitted simultaneously. The data were fitted best when the gain of the tip filter was assumed to be a function of the signal level (as opposed to the masker level per ERB). The filter shapes showed a level dependence that qualitatively resembled the level dependence of filtering on the basilar membrane. The maximum gain of the tip filter tended to increase with increasing center frequency up to 1 kHz, but to remain roughly constant for higher frequencies. PMID- 11108373 TI - Frequency modulation detection interference produced by asynchronous and nonsimultaneous interferers. AB - The effect of asynchronous and nonsimultaneous interferers on detection of sinusoidal frequency modulation (FM) was compared with the effect of a synchronous interferer. In a two-interval, two-alternative forced-choice (2I 2AFC) adaptive procedure, listeners had to detect FM with a modulation frequency of 15 Hz, imposed on a 1-kHz sinusoidal carrier (the target). The 200-ms target was presented either alone (baseline condition), or with an interferer whose timing relative to the target was varied. The interferer was a 2.3-kHz sinusoidal carrier which was also frequency modulated at a rate of 15 Hz. Experiment one showed that thresholds for detection of FM increased significantly, both with a synchronous FM interferer, and also with asynchronous interferers (starting 200 ms before and stopping 200 ms after the target). Moreover, "gapped" interferers that were turned off during presentation of the target (presented for 200 ms before and for 200 ms after the target but not simultaneously) produced the same significant increase in thresholds as an asynchronous interferer that was not interrupted. In contrast, thresholds were not affected by the presence of a gapped unmodulated sinusoidal interferer. Experiment two showed that increasing the duration of the silent gap (centered on presentation of the target) between FM interferers from 200 to 600 ms did not abolish the interference. Thus nonsimultaneous FM interferers produced frequency modulation detection interference (FMDI) even when the silent gap between the interferers and target clearly led to the interferers and target being perceived as separate auditory objects. A possible explanation for the findings is the existence of an asymmetry in perception of steady and modulated sounds, as recently proposed by Cusack and Carlyon [Br. J. Audiol. 34.2, 112 (2000)]. Alternative explanations in terms of ringing in a hypothetical modulation filter bank and adaptation seem unlikely. PMID- 11108374 TI - Effects of relative phase and frequency spacing on the detection of three component amplitude modulation. AB - These experiments explored the effect of relative modulator phase on the detection of a three-component modulator applied to a 4,000-Hz sinusoidal carrier with a level of 70 dB SPL. The central modulator component had a frequency of 50 Hz, and the two other components had frequencies of 50+/-5, 10, 25, 40, or 45 Hz. Thus, the modulator waveform was always periodic. Each modulator component had the same modulation index, m. The relative phases of the components were chosen to give a variety of modulation waveforms differing in the ratio of maximum to minimum value (max-min) and in crest factor. In experiment 1, modulation detection thresholds were measured by varying m, using an adaptive two-interval forced-choice procedure. Thresholds were found to be independent of relative modulator phase and of the frequency spacing of the components. In experiment 2, detectability (d') of the modulation was measured for several fixed values of m. Detectability was found to be independent of relative modulator phase and of the frequency spacing of the components. The results are not consistent with the idea that modulation detection thresholds are determined by the max-min value or crest factor of the envelope. The results are consistent with a model which assumes that the stimuli are subjected to a nonlinearity, and thresholds are determined by the root-mean-square value (or the mean square value) of the ac component of the envelope, following this nonlinearity. The nonlinearity may partly reflect compression on the basilar membrane, but other nonlinearities may be involved. This model can also explain some aspects of earlier results on the sensitivity to relative modulator phase [E. A. Strickland and N. F. Viemeister, J. Acoust. Soc. Am. 99, 3638-3646 (1996)]. PMID- 11108375 TI - Failure to unlearn the precedence effect. AB - Studies of the precedence effect using two binaural clicks have shown that listeners' ability to discriminate changes in the interaural time difference (ITD) of the lagging click is much poorer than that for the leading click [e.g., Zurek, J. Acoust. Soc. Am. 67, 952-964 (1980)]. This difference is thought to reflect an auditory process that suppresses directional information from the lagging sound and attributes greater perceptual weight to directional information contained in the leading one. A report by Saberi and Perrott [J. Acoust. Soc. Am. 87, 1732-1737 (1990)] suggested that listeners can "unlearn" this suppression of the lag's directional information after training with an adaptive psychophysical procedure involving 100 reversals and extremely small step sizes. Here, an attempt was made to find a similar effect using psychophysical procedures that are more common to precedence studies. Eight subjects were rigorously trained on the precedence task using either a blocked procedure or an adaptive procedure to vary ITD. Listeners showed no sign of unlearning. After 9-31 h of participating in the task, all subjects maintained high lag just-noticeable differences (jnd's) and low single source jnd's. This failure to train away the precedence effect (as manifested in discrimination suppression) suggests that directional information contained in the lagging source is not easily accessed. Several possible explanations for the discrepancies between the present study and Saberi and Perrott's finding are discussed. PMID- 11108376 TI - Investigating perceptual features of electrode stimulation via a multidimensional scaling paradigm. AB - To achieve the most effective speech processing for individuals with cochlear implants, it is important to understand the perceptual features associated with the stimulation parameters. In general, when electrodes are stimulated in order from apex to base, the pitch of the perceived sound changes in an orderly fashion from low to high. Some deviations from this assumed order have been documented. Also, pitch is the dominant perceptual attribute of a sound when the stimuli associated with different electrodes have been accurately loudness balanced. In this study, the results of a multidimensional scaling (MDS) paradigm were compared to the results of a pitch-ranking procedure for six subjects implanted with multichannel cochlear prostheses. Results indicate that there may be multiple percepts that change with electrode location. Not surprisingly, the dominant percept is strongly correlated with pitch. The results also indicate that the structure of the second percept is consistent across subjects, although not interpretable using the data measured in this study. Furthermore, results indicate that MDS data can be used to pinpoint indiscriminable electrodes more accurately than pitch data. The results of this study may have importance for the design of the next generation of speech processors. PMID- 11108377 TI - Evaluation of feedback-reduction algorithms for hearing aids. AB - Three adaptive feedback-reduction algorithms were implemented in a laboratory based digital hearing aid system and evaluated with dynamic feedback paths and hearing-impaired subjects. The evaluation included measurements of maximum stable gain and subjective quality ratings. The continuously adapting CNN algorithm (Closed-loop processing with No probe Noise) provided the best performance: 8.5 dB of added stable gain (ASG) relative to a reference algorithm averaged over all subjects, ears, and vent conditions. Two intermittently adapting algorithms, ONO (Open-loop with Noise when Oscillation detected) and ONQ (Open-loop with Noise when Quiet detected), provided an average of 5 dB of ASG. Subjects with more severe hearing losses received greater benefits: 13 dB average ASG for the CNN algorithm and 7-8 dB average ASG for the ONO and ONQ algorithms. These values are conservative estimates of ASG because the fitting procedure produced a frequency gain characteristic that already included precautions against feedback. Speech quality ratings showed no substantial algorithm effect on pleasantness or intelligibility, although subjects informally expressed strong objections to the probe noise used by the ONO and ONQ algorithms. This objection was not reflected in the speech quality ratings because of limitations of the experimental procedure. The results clearly indicate that the CNN algorithm is the most promising choice for adaptive feedback reduction in hearing aids. PMID- 11108378 TI - Speech recognition by normal-hearing and cochlear implant listeners as a function of intensity resolution. AB - The importance of intensity resolution in terms of the number of intensity steps needed for speech recognition was assessed for normal-hearing and cochlear implant listeners. In experiment 1, the channel amplitudes extracted from a six channel continuous interleaved sampling (CIS) processor were quantized into 2, 4, 8, 16, or 32 steps. Consonant recognition was assessed for five cochlear implant listeners, using the Med-El/CIS-link device, as a function of the number of steps in the electrical dynamic range. Results showed that eight steps within the dynamic range are sufficient for reaching asymptotic performance in consonant recognition. These results suggest that amplitude resolution is not a major factor in determining consonant identification. In experiment 2, the relationship between spectral resolution (number of channels) and intensity resolution (number of steps) in normal-hearing listeners was investigated. Speech was filtered through 4-20 frequency bands, synthesized as a linear combination of sine waves with amplitudes extracted from the envelopes of the bandpassed waveforms, and then quantized into 2-32 levels to produce stimuli with varying degrees of intensity resolution. Results showed that the number of steps needed to achieve asymptotic performance was a function of the number of channels and the speech material used. For vowels, asymptotic performance was obtained with four steps, while for consonants, eight steps were needed for most channel conditions, consistent with our findings in experiment 1. For sentences processed though 4 channels, 16 steps were needed to reach asymptotic performance, while for sentences processed through 16 channels, 4 steps were needed. The results with normal-hearing listeners on sentence recognition point to an inverse relationship between spectral resolution and intensity resolution. When spectral resolution is poor (i.e., a small number of channels is available) a relatively fine intensity resolution is needed to achieve high levels of understanding. Conversely, when the intensity resolution is poor, a high degree of spectral resolution is needed to achieve asymptotic performance. The results of this study, taken together with previous findings on the effect of reduced dynamic range, suggest that the performance of cochlear implant subjects is primarily limited by the small number (four to six) of channels received, and not by the small number of intensity steps or reduced dynamic range. PMID- 11108379 TI - Air-wood coupling and the Swiss-cheese violin. AB - Some problems with the conventional formalism for describing the coupling of fluid vibrations to those of an enclosing shell are examined. An alternative ("dynamic") basis for expanding the normal modes, in which the "pure shell modes" include incompressible motion of the fluid, is proposed. This new approach is applied to Hutchins's "Swiss-cheese violin," the behavior of whose air modes for the case of a rigid shell have been calculated by Shaw. Measurements are presented of various response functions of this instrument as a function of both frequency and the number of open rib holes. These results show the predicted "mode veering" behavior, and can be easily matched to theory with the assumption of plausible parameters. PMID- 11108380 TI - Frequency dependence of ultrasonic backscattering in cancellous bone: autocorrelation model and experimental results. AB - The goal of this study is to model the frequency dependence of the ultrasonic backscatter coefficient in cancellous bone. A twofold theoretical approach has been adopted: the analytical theoretical model of Faran for spherical and cylindrical elastic scatterers, and the scattering model for weakly scattering medium in which the backscatter coefficient is related to the autocorrelation function of the propagating medium. The ultrasonic backscatter coefficient was measured in 19 bone specimens (human calcaneae) in the frequency range of 0.4-1.2 MHz. The autocorrelation function was computed from the three-dimensional (3D) microarchitecture measured using synchrotron radiation microtomography. Good agreement was found between the frequency dependence of the experimental (f3.38+/ 0.31) and autocorrelation modeled (f3.48+/-0.26) backscatter coefficients. The results based on Faran theory (cylindrical Faran model: f2.89+/-0.06 and spherical Faran model: f3.91+/-0.04) show qualitative agreement with experimental data. The good prediction obtained by modeling the backscatter coefficient using the autocorrelation function of the medium opens interesting prospects for the investigation of the influence of bone microarchitecture on ultrasonic scattering. PMID- 11108381 TI - The role of leaf structure in vibration propagation. AB - The leaf and its structural components play a key role in the propagation of short transient signals produced by insects. In this paper, it is shown how the complex structure of an apple leaf could be modeled by a much simpler one for the analysis of vibratory signal propagation. Waves were produced by impacts of small spheres and the propagation studied using two laser vibrometers, followed by a wavelets analysis. Three components of the leaf were investigated: the midvein, minor veins, and the interspaced homogeneous regions making up the leaf lamina. The loss of signal energy over the leaf lamina and across minor veins and midvein was studied. For the midvein, the loss of energy decreased from 80% at the leaf base to 40% at the apex. For minor veins, the loss of energy decreased from 70% at the leaf base to 31% at the apex. The loss in homogeneous regions was 40%. A signal decomposition into two frequency ranges, above and below 1.7 kHz, showed that the midvein acted as a low-pass filter. As energy loss was mainly a function of vein diameter and not vein type, veins smaller or equal to 0.2 mm were considered as equivalent to homogeneous regions. Hence, a model leaf reduced to the leaf lamina and veins with a diameter >0.2 mm is retained for the study of signal propagation in a leaf. PMID- 11108382 TI - Echolocation behavior of big brown bats, Eptesicus fuscus, in the field and the laboratory. AB - Echolocation signals were recorded from big brown bats, Eptesicus fuscus, flying in the field and the laboratory. In open field areas the interpulse intervals (IPI) of search signals were either around 134 ms or twice that value, 270 ms. At long IPI's the signals were of long duration (14 to 18-20 ms), narrow bandwidth, and low frequency, sweeping down to a minimum frequency (Fmin) of 22-25 kHz. At short IPI's the signals were shorter (6-13 ms), of higher frequency, and broader bandwidth. In wooded areas only short (6-11 ms) relatively broadband search signals were emitted at a higher rate (avg. IPI= 122 ms) with higher Fmin (27-30 kHz). In the laboratory the IPI was even shorter (88 ms), the duration was 3-5 ms, and the Fmin 30- 35 kHz, resembling approach phase signals of field recordings. Excluding terminal phase signals, all signals from all areas showed a negative correlation between signal duration and Fmin, i.e., the shorter the signal, the higher was Fmin. This correlation was reversed in the terminal phase of insect capture sequences, where Fmin decreased with decreasing signal duration. Overall, the signals recorded in the field were longer, with longer IPI's and greater variability in bandwidth than signals recorded in the laboratory. PMID- 11108383 TI - The single sonic muscle twitch model for the sound-production mechanism in the weakfish, Cynoscion regalis. AB - A model of the weakfish, Cynoscion regalis, sound-production mechanism based on damped driven oscillators is presented. The weakfish "purr" consists of several short pulses of sound separated by intervals of no sound. Each pulse is produced by an individual simultaneous twitch of each sonic muscle causing swimbladder oscillations that radiate sound into the surrounding water. The sonic muscles are modeled as a stretched string with a time-varying tension force. The swimbladder is modeled as a highly damped, driven oscillator undergoing radial oscillations. Although the swimbladder functions as an impedance-matching device between the sonic muscles and the surrounding fluid, its transient response to the sonic muscle pulses includes frequencies that are not a part of the spectrum of the sonic muscle excitations. Differential equations of motion for the sonic muscles and swimbladder are given and are numerically solved to produce predicted waveforms matching those of measured weakfish sounds. The model leads to better understanding of the weakfish sound-producing mechanism and the effects of environmental and physiological factors on sound production. This model may also lead to a better understanding of sound production in other species, particularly other members of Family Sciaenidae, with similar mechanisms for sound production. PMID- 11108384 TI - Variations in temporal patterns of speech production among speakers of English. PMID- 11108385 TI - Varroa jacobsoni (Acari: Varroidae) is more than one species. AB - Varroa jacobsoni was first described as a natural ectoparasitic mite of the Eastern honeybee (Apis cerana) throughout Asia. It later switched host to the Western honeybee (A. mellifera) and has now become a serious pest of that bee worldwide. The studies reported here on genotypic, phenotypic and reproductive variation among V. jacobsoni infesting A. cerana throughout Asia demonstrate that V. jacobsoni is a complex of at least two different species. In a new classification V. jacobsoni is here redefined as encompassing nine haplotypes (mites with distinct mtDNA CO-I gene sequences) that infest A. cerana in the Malaysia Indonesia region. Included is a Java haplotype, specimens of which were used to first describe V. jacobsoni at the beginning of this century. A new name, V. destructor n. sp., is given to six haplotypes that infest A. cerana on mainland Asia. Adult females of V. destructor are significantly larger and less spherical in shape than females of V. jacobsoni and they are also reproductively isolated from females of V. jacobsoni. The taxonomic positions of a further three unique haplotypes that infest A. cerana in the Philippines is uncertain and requires further study. Other studies reported here also show that only two of the 18 different haplotypes concealed within the complex of mites infesting A. cerana have become pests of A. mellifera worldwide. Both belong to V. destructor, and they are not V. jacobsoni. The most common is a Korea haplotype, so-called because it was also found parasitizing A. cerana in South Korea. It was identified on A. mellifera in Europe, the Middle East, Africa, Asia, and the Americas. Less common is a Japan/Thailand haplotype, so-called because it was also found parasitizing A. cerana in Japan and Thailand. It was identified on A. mellifera in Japan, Thailand and the Americas. Our results imply that the findings of past research on V. jacobsoni are applicable mostly to V. destructor. Our results will also influence quarantine protocols for bee mites, and may present new strategies for mite control. PMID- 11108386 TI - Colonization of vineyards by phytoseiid mites: their dispersal patterns in the plot and their fate. AB - The effect of wind and woody margins on the dispersal and population dynamics of phytoseiid mites was studied in a vine plot for a period of two years. Mites were sampled in the plot and in the surrounding vegetation (crops and natural vegetation) in order to determine phytoseiid mite abundance. The surrounding vegetation was considered to be a reservoir of phytoseiids from where the vine plot could be invaded. Directional and non-directional soil and aerial traps were placed in the plot to determine predatory mite exchange between the two areas. Colonization of the plot occurred in two stages: first, mite migration into the plot, followed by their establishment. The two-year study partially clarified the first of these two stages. Kampimodromus aberrans was the main species caught in the aerial traps. Phytoseiid mite dispersal within the vine plot seemed to be affected by both wind (direction, intensity and regularity) and phytoseiid mite density in the woody margin. However, the woody margin had a large effect only over a short distance. Some observations pointed towards an effect of other reservoir areas but it was not possible to characterize these. The population density of the phytoseiid mites in the plot increased from 1996 to 1998, but these increases are much smaller than one would expect on the basis of the number of mites migrating by air in the plot. Moreover, blocks where most mites were trapped were not the blocks where densities of phytoseiid mites on vine leaves were the largest. It therefore seems likely that not all migrants were able to develop. Their settlement pattern was not determined and this could constitute a potential research focus for the future. PMID- 11108387 TI - Species of the genus Psoroptes (Acari: Psoroptidae): a taxonomic consideration. AB - The biosystematic status of mite species belonging to the genus Psoroptes Gervais, 1841 is difficult to determine by phenotypic methods and has been subject to taxonomic revisions and ongoing debate. At present, the existence of five species, P cuniculi (Delafond, 1859), P. ovis (Hering, 1838). P. equi (Hering, 1838), P. cervinus Ward, 1915 and P. natalensis Hirst, 1919, is generally accepted. This classification is based mainly on the host species, the localization of the mites on their hosts and morphological characters of male mites. However, a critical review of the literature indicates that the features used to discriminate between the five species are not unequivocal: (a) the localization of mite populations on host animals is not completely strict, (b) the lengths of the outer opisthosomal setae of male mites, which are the main morphological features used for species discrimination, overlap between the five postulated species, and (c) host specificity cannot be deduced from results of transfer experiments. Rather, conspecificity of the members of the genus Psoroptes has to be presumed which is supported by molecular genetic analyses. On these grounds and on rules of priority P. cervinus Ward, 1915, P. cuniculi (Delafond, 1859), P. natalensis Hirst, 1919 and P. ovis (Hering, 1838) are seen as synonyms of P. equi (Hering, 1838). PMID- 11108388 TI - Arrestment response of the predatory mite Amblyseius longispinosus to Schizotetranychus nanjingensis webnests on bamboo leaves (Acari: Phytoseiidae, Tetranychidae). AB - The response of the predatory mite Amblyseius longispinosus (Acari: Phytoseiidae) to the webnest of the spider mite Schizotetranychus nanjingensis (Acari: Tetranychidae) was examined using two-choice tests in the laboratory. A. longispinosus females were found significantly more often on leaves with webnests than on leaves without webnests and were often observed searching under the webbing. Because spider mites and their eggs were removed from the webnests before experiments, predators responded to stimuli associated with webbing, mite feeding damage and other residues in the webnests. PMID- 11108389 TI - Localisation and functional studies on the 5'-nucleotidase of the cattle tick Boophilus microplus. AB - The ecto-5'-nucleotidase from the cattle tick Boophilus microplus is an unusual enzyme, hydrolysing a variety of nucleoside mono-, di- and triphosphates to release the free nucleoside. The gene has been sequenced and the recombinant protein expressed as a functional, active enzyme. Nevertheless, the function of the enzyme in the tick remains obscure. The enzyme is present throughout the life cycle, but in largest amounts in unfed larvae and adult ticks. The tissue location has been studied in adult female ticks by Western blotting, RT-PCR and immunofluorescence. All methods show the enzyme to be principally in the Malpighian tubules, though significant amounts are also present on the surface of ovaries and in detectable amounts in other tissues. This, together with the known specificity of the enzyme, suggests a role in purine salvage pathways. Sensitivity of ticks to allopurinol, an inhibitor of hypoxanthine-guanine phosphoribosyltransferase, supports the importance of purine salvage in this tick and the potential role of nucleotidase in this pathway. PMID- 11108390 TI - Biology, ecology, and management of the bulb mites of the genus Rhizoglyphus (Acari: Acaridae). AB - Bulb mites of the genus Rhizoglyphus (Claparede) (Acari: Acaridae) have been identified as pests of many crops and ornamentals in storage, in the greenhouse, and in the field. The most important hosts are species in the family Liliaceae (e.g. Allium spp.), but bulb mites will often attack other important crops such as potatoes (Solanum sp.) and carrots (Daucus carota). Despite their economic importance and broad distribution, the systematics of the genus remains in a state of confusion and is in need of a comprehensive revision. In addition, the field biology and ecology of these mites is not well understood, and methods for sampling, monitoring, and loss assessment are limited. Management of bulb mites is complicated by their short generation time, high reproductive potential, broad food niche, interactions with other pests and pathogens, and unique adaptations for dispersal. Historically, control of these acarine pests has relied on the use of synthetic miticides and insecticides, but this option is now limited due to documented resistance and withdrawal of registration of some products. Alternative control strategies, including cultural and biological control, have shown limited success, but need to be further developed and implemented. PMID- 11108391 TI - Comparison of life-history traits of the two male morphs of the bulb mite, Rhizoglyphus robini. AB - Two basic male morphs occur in several species of the family Acaridae: heteromorphic fighters, possessing a thickened and sharply terminated third pair of legs, and homeomorphic males with unmodified legs. We compared major life history traits of the two morphs in the bulb mite, Rhizoglyphus robini. We found no significant differences in development time or virility, but homeomorphic males lived 23% longer than heteromorphs. We discuss the possibility that the trade-off between longevity and adaptation for fighting maintains genetic variation for the male morph in the studied species. PMID- 11108392 TI - Cold storage of Halotydeus destructor (Acari: Penthaleidae) for use in experiments. AB - Survival of medium sized nymphal stages of redlegged earth mite Halotydeus destructor (Tucker) (mainly tritonymphs and deutonymphs) stored under low temperature (1.5 degrees C) in sealed plastic boxes remained more than 50% after 12 days of storage, with some mites surviving for up to eight weeks. Adding fresh subclover leaves into the storage box increased the survival rate of mites from 12% to 28%, 19 days after the storage started. Mites stored for two weeks at low temperatures showed feeding activity in a screening experiment similar to mites collected directly from the field. This indicated that cold storage of redlegged earth mite can be used to build up mite numbers for large screening experiments, or to extend the period of availability of mites collected from the field. However, their reproductive ability was greatly reduced after three weeks at low temperature. Thus, care should be taken when using mites for experiments measuring reproduction. The implications of low temperatures for reducing field populations of mites in midwinter are also discussed. PMID- 11108393 TI - Tullgren extraction of soil mites (Acarina): effect of refrigeration time on extraction efficiency. PMID- 11108394 TI - An epizootic of Calacarus heveae (Acari: Eriophyidae) caused by Hirsutella thompsonii on rubber trees. PMID- 11108395 TI - Chemical composition of lipophylic compounds from the body surface of unfed adult Ixodes persulcatus ticks (Acari: Ixodidae). AB - Chemical compositions of ethereal extracts of the body surface of unfed male and female Ixodes persulcatus ticks (Acari: Ixodidae) were studied by gas chromatography using mass-spectrometric detection. More than 100 different organic compounds were detected. The predominant components were saturated fatty hydrocarbons, saturated and unsaturated fatty acids, aldehydes, squalene, cholesterol and cholesterol derivatives. A number of compounds found in I. persulcatus are known as components of pheromones or constituents of dermal gland secretions in tick species of the genus Amblyomma: nonanoic acid, saturated fatty acids having from 14 to 16 carbons, and squalene. Saturated fatty aldehydes have not been reported previously as body surface components of hard ticks. Substituted phenols were not found in the extracts, although they are known as common components of sex and attraction-aggregation-attachment pheromones in Amblyomma ticks. With a few exceptions (henicosanal, 2,4-holestadiene and two unidentified cholesterol derivatives), there was no marked difference in composition of surface components between male and female I. persulcatus. The possible role of the different chemical groups in communication between I. persulcatus ticks is discussed. PMID- 11108396 TI - Attraction of immature stages of the American dog tick (Dermacentor variabilis) to 2,6-dichlorophenol. AB - To determine whether 2,6-dichlorophenol is solely a sex pheromone, the response to it by the various stages of the American dog tick, Dermacentor variabilis, were compared. In contrast to adults, 2,6-dichlorophenol was attractive to unfed nymphs and to unfed larvae. Use of this chemical also prompted the expression of a novel type of 'feeding' posture behavior in adults. The overlap in attraction to other substituted phenols plus the lack of functional value of this response for larvae and nymphs rules out the possibility that 2,6-dichlorophenol is a general attractant. However, 2,6-dichlorophenol likely plays a dual role as an attachment stimulant in the adult tick. PMID- 11108397 TI - The role of dental public health in Europe. PMID- 11108398 TI - A systematic review of public water fluoridation--a commentary. PMID- 11108399 TI - The dental needs, demands and attitudes of a group of homeless people with mental health problems. AB - OBJECTIVE: This study investigated the dental needs, demands and attitudes of a group of homeless people living in a hostel in Birmingham, many of whom had mental health problems. BASIC RESEARCH DESIGN: Seventy subjects underwent a dental examination. The clinical criteria for the examination were especially selected to be simple and cause minimal discomfort to the subject, but be reproducible and cover the wide range of conditions expected to be found. Five of the subjects were selected to take part in semi structured interviews. RESULTS: Thirty-one per cent of the subjects were found to be edentulous, with only 32% wearing dentures. The dentate subjects had a mean DMFT (+/-SE) of 15.9 (+/-7.8). High levels of dental need were found amongst the dentate subjects who had an average of 3.6 (+/-3.9) decayed teeth and 54% had one or more teeth with obvious pulpal involvement. Eighty-five per cent of the dentate subjects had some dental wear leading to exposed dentine. The periodontal condition was generally poor, 50% of dentate subjects having excessively mobile teeth. The interviews revealed a low level of perceived need and indicated that difficulties would be encountered in tailoring services to meet this client group's requirements. CONCLUSIONS: High levels of normative need were found in this group of people, however it was concluded that providing dental services to meet this need would prove difficult. PMID- 11108400 TI - Comparing the ability of different area measures of socioeconomic status to segment a population according to caries prevalence. AB - OBJECTIVE: to compare the ability of different area based indicators of socio economic status to segment a population of 5-year-old children according to caries prevalence. BASIC RESEARCH DESIGN: The study population consisted of all 5 year-old children in seven districts in the North West Region of England who were examined in whole population surveys during the 1995/6 NHS epidemiological survey. This population was segmented according to caries prevalence by market penetration analyses using Super Profiles and the ONS geodemographic classifications, Jarman and Townsend (at ward and ED level) deprivation indices, the single census variables of unemployment and percentage of households without a car, and also the school that the children attended. Lorenz curves were plotted from the outputs of the penetration analyses. MAIN OUTCOME MEASURES: Overall caries prevalence for the population was 55.3%. All the indicators provided a very similar picture. Large differences were detected between the highest and lowest market penetration rankings for each indicator, ranging from 42.5% for the ONS geodemographic classification to 31.4% for the Townsend index at enumeration district level. However, for each indicator, the fall between these two extremes was gradual. This picture was represented by the similar effectiveness scores of around 10% for each indicator which were derived from the Lorenz curves. Each indicator could identify approximately 30% of those with disease in the topmost 25% of the total population in the penetration ranking. When the analysis was restricted to those children with severe disease (dmft> or =5) a similar picture was found, however the effectiveness score increased to 19%. CONCLUSIONS: There was a remarkably consistent inequitable distribution of population disease prevalence found between deprived and affluent area types, irrespective of what measuring instrument was used. Although there was a large difference in prevalence (dmft>0 and dmft>5) found between deprived and affluent areas, there was a gradual fall between the two extremes. This gradual fall has implications for oral health improvement strategies. PMID- 11108401 TI - An economic evaluation of a publicly funded dental prevention programme in regional and rural Victoria: an extrapolated analysis. AB - OBJECTIVE: To determine the long-term cost-benefit of a community-wide, publicly funded dental prevention programme. DESIGN: A modelled economic analysis which extrapolated the effectiveness and cost-effectiveness results of a three-year comprehensive preventive dental programme conducted in a single cohort of adolescents in the non-fluoridated towns of Geelong and Ballarat, Victoria, Australia. Assumptions were made for both benefits and costs. Sensitivity analysis was undertaken to report a range of estimates of potential programme benefits. SETTING: All secondary colleges in two non-fluoridated regional centres and their surrounding rural areas. SUBJECTS: All Year 7 to 9 students; mean age range of 12.5 to 15.5 years. RESULTS: The incremental benefit-to-cost ratios under all assumptions improved with each successive year of the community-wide programme and, even with the most conservative of assumptions, the overall ten year benefit-to-cost ratio was above unity. CONCLUSION: While the analysis has inherent limitations as a result of its reliance on a range of assumptions, the findings do suggest that there are benefits to be gained from the implementation of a comprehensive dental preventive programme throughout the secondary school system in non-fluoridated centres comparable to Geelong and Ballarat. PMID- 11108402 TI - The temporal and spatial epidemiology of lip cancer in England and Wales. AB - OBJECTIVE: To examine trends in lip cancer mortality and incidence during the 20th century, and its geographical distribution, in England and Wales as a prerequisite to establishing any essential differences in the aetiology of lip and intra-oral cancer. METHOD: Age-standardised rates for lip cancer mortality by gender from 1901 to 1991 were derived from archived OPCS data. Standardised incidence rates from 1962 to 1986 were also calculated. Registrations of lip cancer between 1979 and 1983 in the 15 RHAs covering England and Wales were obtained from the same source and standardised incidence ratios (SIR) computed. RESULTS: Progressive and sustained falls in lip cancer mortality and incidence were recorded in the period covered. In males, these related particularly to cohorts born after 1856. In addition, the incidence of lip cancer in both genders was found to be raised in a band stretching from East Anglia to the South West and also in the Trent and Wales RHAs. CONCLUSIONS: The findings support an hypothesis that employment in the agriculture, forestry and fishing industries, and also pipe smoking, were the major risk factors for lip cancer in the 20th century. Further research is required to differentiate between the aetiological risk factors for lip and intra-oral cancer. PMID- 11108403 TI - Oral health status and treatment need of 11-13-year-old urban children in Tibet, China. AB - OBJECTIVE: To describe the oral health status and treatment need of 11-13-year old children in urban Tibet and to determine if there was a difference in oral health status between the Tibetan and Han children. SURVEY DESIGN AND SUBJECTS: The two largest primary schools in Lhasa, Tibet, were selected and all children aged between 11 and 13 years were surveyed. Each child was interviewed and clinically examined in the schools, using portable equipment, by one of four calibrated examiners. The examination procedures and diagnostic criteria used followed those recommended by the World Health Organization. RESULTS: 347 children (207 Tibetans and 140 Han) were surveyed. Over 90% of the children claimed to brush their teeth at least once a day. Three-quarters had never visited a dentist. Caries prevalence was 44% in the Tibetans and 24% in the Han (P<0.001). The mean DMFT scores of the Tibetans and Han were 0.8 and 0.4 respectively (P<0.001). Only 1% of the children in both ethnic groups had healthy gums (highest CPI score = 0) and about two-thirds of them had calculus. One-third of the children were in need of treatment for dental caries and most of the required treatment items were one-surface fillings but 10% of the children needed extraction. CONCLUSION: Dental caries and treatment need level of both Tibetan and Han children in Tibet was low but their periodontal health status was unsatisfactory. PMID- 11108404 TI - Changing knowledge, attitudes, and practices of Thai oral health personnel with regard to AIDS: an evaluation of an educational intervention. AB - OBJECTIVE: This study evaluated the effect of an educational intervention in improving Thai oral health personnel's (OHP) knowledge, attitudes and practices (KAP) regarding HIV/AIDS. RESEARCH DESIGN: The study used a pre-test/post-test design with study and control groups. RESULTS: of the pre-test questionnaire were used to design the intervention. Three months after the study group received the intervention, the same questionnaire was given to both groups. INTERVENTION: A three-day workshop was conducted using a variety of teaching methods: lectures, videos, role-plays, interviews with HIV infected persons, and demonstrations. SETTING: The study was conducted in rural government dental clinics in three provinces in southern Thailand. One hundred and three OHP in 23 dental clinics were in the study group while 46 OHP in II dental clinics were in the control group. OUTCOME MEASURES: The outcomes were knowledge and attitudes regarding HIV/AIDS, perception of occupational risk, willingness to treat HIV infected persons and adherence to recommended infection control procedures. RESULTS AND CONCLUSIONS: The educational intervention resulted in significant improvement in many domains of KAP in the study group, while there was little change in the control group. The post-test questionnaire showed that further improvements are needed in attitudes towards HIV/AIDS and practices regarding accidental needle stick injury. The intervention was both effective and appropriate and should be considered for national use. PMID- 11108405 TI - The oral assessment in Down syndrome questionnaire (OADS): development of an instrument to evaluate oral health problems in individuals with Down syndrome. AB - OBJECTIVE: To develop a French language instrument whose primary aim is the description of oral health status in Down syndrome (DS) individuals. METHOD: Due to the reduced intellectual capacity in DS individuals, the instrument was designed to be completed by their parents. Items were generated through a literature review plus interviews with relevant professionals and DS parents. Following pilot testing, the 31-item questionnaire was divided into seven domains (access, function, development, signs, pain, disability and a global evaluation of oral health status), and subjected to a psychometric evaluation of internal reliability, test-retest reliability and discriminant validity. Internal reliability was assessed through evaluation of Cronbach's alpha or the Kuder Richardson-20 value where appropriate. Test-retest reliability was assessed through the evaluation of the intra-class correlation coefficient (ICC) for each domain. Discriminant validity was assessed through evaluations of hypothesised differences in domain scores between different groups of DS individuals by age and between DS and non-DS individuals. The theoretical domain categorisation was empirically evaluated through the generation of inter-item correlation coefficients. RESULTS: Internal reliability coefficients ranged from 0.43 for the disability domain to 0.83 for the function domain, while the domain ICCs ranged from 0.56 to 0.77 for the signs and function domains respectively. For discriminant validity, the domains largely performed as hypothesised. Finally, inter-item correlation coefficients largely supported the proposed domain structure of the questionnaire. CONCLUSION: This investigation of the psychometric properties of a proxy. French language, assessment of oral health problems in the DS population has demonstrated an instrument with good preliminary indicators of reliability and validity. PMID- 11108406 TI - Identifying local patterns of cross boundary flow and developing socio-economic profiles for general dental practitioners. AB - OBJECTIVES: The objective of this study was to assess cross-boundary flow in general dental practices in Dudley. DESIGN: Data were collected on NHS patients attending 57 general dental practitioners in Dudley Health District. Socio economic data were attached to recognised postcodes in order to establish district of residence of patients and the socio-economic (ACORN) status of the neighbourhood in which they lived. SETTING: General dental practices in Dudley. SUBJECTS: General Dental Service patients. RESULTS: Using this methodology a profile of the patient population attending the participating practitioners was drawn up, giving an insight into patterns of dental attendance. CONCLUSION: These profiles will act as indicators of practices in which there are high levels of cross boundary flow and inform discussions on local commissioning. PMID- 11108407 TI - Assessment of muscle tone. PMID- 11108408 TI - Comprehensive geriatric assessment revisited...again. PMID- 11108409 TI - Complaints, doctors and older people. PMID- 11108410 TI - Re-thinking metabolic strategies for older people with type 2 diabetes mellitus: implications of the UK Prospective Diabetes Study and other recent studies. PMID- 11108411 TI - Adherence to recommendations of community-based comprehensive geriatric assessment programmes. AB - BACKGROUND: non-adherence to the recommendations of short-term community-based consultative comprehensive geriatric assessment programmes is a threat to the effectiveness of these programmes. OBJECTIVE: to synthesize the literature on patient and physician adherence to recommendations of community-based comprehensive geriatric assessment programmes. METHOD: I identified papers cited by an English language literature search of MEDLINE, Health Star and CINAHL databases from January 1980 to November 1999. This search was supplemented with literature identified from the reference sections of these publications. RESULTS: patient adherence rates ranged from 46 to 76%, which approximates to the rates for the consulting physician adherence (49-79%). I identified many characteristics of patient, treatment, care provider and clinical setting which influenced adherence. Understanding these factors has led to the development of adherence-enhancing strategies. However, without systematic evaluations it is difficult to evaluate the relative effectiveness of these interventions. CONCLUSION: further research which targets more representative samples and uses validated assessment tools and multiple data collection methods is needed to expand our knowledge of patterns and predictors of adherence and to evaluate the relative effectiveness of adherence-enhancing intervention strategies. PMID- 11108412 TI - Complaints concerning the hospital care of elderly patients: a 12-month study of one hospital's experience. AB - OBJECTIVE: to determine the number, instigators, nature and outcome of complaints concerning elderly patients treated at a single hospital over 1 year. DESIGN: descriptive analysis of computerized data gathered prospectively; follow-up of complaints until resolution. SETTING: large, urban, university teaching hospital in Australia. SUBJECTS: all patients aged 65 years and above whose hospital care was the subject of complaint. METHODS: analysis of computerized database of all complaints made in a single year. RESULTS: 1.44 complaints were made per 1000 occasions of service to elderly people (95% confidence intervals, 1.19 - 1.69). This was similar to the overall complaint rate of 1.32 per 1000 occasions of service for patients of all age groups (95% confidence intervals, 1.19- 1.45). However, 73% of complaints were made by advocates rather than by elderly patients themselves and 96% related to communication or treatment issues. Many complaints resulted in an explanation and/or an apology and, to date, none has resulted in litigation. CONCLUSIONS: complaints concerning older hospitalized people are as common as those concerning younger patients. Analysis of complaints provides pointers for improvements in quality of care. PMID- 11108413 TI - Complications of carotid sinus massage--a prospective series of older patients. AB - BACKGROUND: there is a causal association between carotid sinus hypersensitivity, falls and syncope in elderly subjects. Neurological complications during carotid sinus massage have been reported in case studies and two retrospective series. Our aim was prospectively to ascertain the incidence of complications occurring after carotid sinus massage performed for diagnostic purposes in a consecutive series of patients. METHODS: 1000 consecutive subjects aged 50 years or over who attended the accident and emergency department with syncope or 'unexplained' falls had carotid sinus massage. Carotid sinus massage was performed for 5 s on the right and then left sides both supine and upright (70 degrees head-up tilt) with continuous heart rate and phasic blood pressure recording. Contraindications to carotid sinus massage were the presence of a carotid bruit, recent history of stroke or myocardial infarction or previous ventricular tachyarrhythmia. RESULTS: complications occurred in nine patients immediately after cessation of carotid sinus massage. Eight had transient neurological complications possibly attributable to carotid sinus massage: visual disturbance, 'pins and needles' and sensation of finger numbness in two cases each, leg weakness in one and sensation of 'being drunk' in one. All transient complications resolved within 24 h. In one patient mild weakness of the right hand persisted. CONCLUSIONS: no subjects had cardiac complications and 1% had possible neurological symptoms, which resolved in most cases. Persistent neurological complications are uncommon, occurring in 1:1000 patients (0.1%) or 1: 3805 episodes of carotid sinus massage (0.03%). PMID- 11108414 TI - Responses to the prolonged head-up tilt followed by sublingual nitrate provocation in asymptomatic older adults. AB - BACKGROUND: prolonged head-up tilt testing and sublingual nitrate provocation are increasingly used in the diagnosis of neurocardiogenic syncope. However there are few data regarding the results of these tests in asymptomatic older subjects. OBJECTIVE: to assess the responses to the prolonged head-up tilt test followed by sublingual glyceryl trinitrate provocation in asymptomatic subjects over the age of 60 years. DESIGN: observational study. METHODS: we recruited 64 asymptomatic subjects over the age of 60 (39 men, 25 women) from two general practice lists in Nottingham and Leicester. Exclusion criteria were: history of syncope, ischaemic heart disease, cerebrovascular disease, marked aortic stenosis, carotid artery disease and being unable to stand for the duration of the test. All subjects underwent a full clinical examination, a 12-lead electrocardiogram and a 30-40 min head-up tilt test, during which we monitored the heart rate and blood pressure continuously. We ended the test prematurely if the subjects developed syncope or symptoms of presyncope associated with hypotension with or without bradycardia. If they remained asymptomatic at the end of this period, they received 400 microg of sublingual glyceryl trinitrate and monitoring continued for another 15 min. SETTINGS: two teaching hospitals in Nottingham and Leicester. RESULTS: six (9%) of the subjects had a positive response (syncope or presyncope) to the prolonged head-up tilt test prior to glyceryl trinitrate provocation. After provocation, 30 (52%) of the remaining 58 subjects had a positive response. CONCLUSION: the role of sublingual glyceryl trinitrate provocation following prolonged head-up tilt testing in the diagnosis of neurocardiogenic (vasovagal) syncope in older people is questionable, as many asymptomatic older subjects demonstrate syncopal or presyncopal symptoms. PMID- 11108415 TI - Nutritional supplementation of elderly hip fracture patients. A randomized, double-blind, placebo-controlled trial. AB - BACKGROUND: undernourishment is common in elderly hip fracture patients and has been linked to poorer recovery and increased post-operative complications. OBJECTIVE: to determine whether a nutritional supplement may (i) help elderly patients return to pre-fracture functional levels 6 months post-fracture and (ii) decrease fracture-related complications and mortality. DESIGN: a double-blind, randomized, placebo-controlled clinical trial. SETTING: a county hospital near Barcelona. SUBJECTS: 171 patients, aged 70 and older, hospitalized for hip fracture between July 1994 and July 1996. METHODS: we randomized patients to intervention (n = 85) or control (n = 86) group. Patients received a nutritional supplement containing 20 g of protein and 800 mg of calcium or placebo for 60 days. We determined functional levels by the Barthel index, the mobility index and by the use of walking aids. We performed assessments during hospitalization and at 2 and 6 months post-fracture. FINDINGS: the two groups were comparable at study entry. We observed no differences in return to functional status 6 months post-fracture (61% intervention group vs 55% in control group) nor in fracture related mortality (13% in intervention group vs 10% in control group). The intervention group suffered fewer in-hospital [odds ratio 1.88 (95% CI 1.01 - 3.53), P = 0.05] and total complications [odds ratio 1.94 (95% CI 1.02-3.7), P = 0.04] than the control group. CONCLUSION: based on our results, we cannot recommend routine nutritional supplementation of all elderly hip fracture patients. While nutritional supplementation may be useful in decreasing complications, this reduction does not result in improvement in functional recovery and nor does it decrease fracture-related mortality. Selected patients may, however, benefit from nutritional supplementation. PMID- 11108416 TI - Low-intensity physical activity benefits blood lipids and lipoproteins in older adults living at home. AB - OBJECTIVE: to examine the influence of low-intensity, habitual physical activity on blood lipids and lipoproteins and other cardiovascular risk factors in older adults living at home. DESIGN: cross-sectional observational study. PARTICIPANTS: a convenience sample of healthy, older adults (n = 155) who were mainly non Hispanic, white (96.8%), female (65.2%) and on medications for cardiometabolic related disorders (60.6%) and had an average age of 74.2 +/- 0.5 years. METHODS: we used a question from the Yale Physical Activity Survey to assess the typical number of hours per day spent in motion during the past month, collapsing responses into <5 and > or = 5 h/day. We determined blood lipids, lipoproteins and glucose with Kodak Ektachem serum oxidase assays or finger stick using the Cholestech LDX system enzymatic technique, and measured blood pressure by auscultation. Waist circumference was the indicator of abdominal fat distribution and body mass index the measure of overall adiposity. RESULTS: after adjusting for age, sex, adiposity, postprandial state, medication use and method of blood sampling, greater amounts of daily accumulated movement were associated with more favourable blood lipid-lipoprotein profiles. Subjects reporting > or = 5 h of daily movement had higher levels of high-density lipoprotein cholesterol [mean difference (95% confidence interval): 0.23 mmol/l (0.07, 0.39); P = 0.002] and a lower ratio of total to high-density lipoprotein cholesterol [average difference: -0.92 (-1.36, -0.48); P = 0.003]. They had lower levels of low-density lipoprotein cholesterol [mean difference: -0.39 mmol/l (-0.80, 0.03); P = 0.074)] and a lower ratio of triglyceride to high-density lipoprotein cholesterol [mean difference: -1.31 (-2.50, -0.12); P = 0.059]. Total cholesterol was similar in the two groups (P > 0.05). The mean blood glucose was 1.49 mmol/l lower (-2.67, 0.31) in the more active group (P = 0.02), independent of age, sex, adiposity, medication use and postprandial state. CONCLUSIONS: low-intensity, habitual physical activity is a sufficient stimulus to enhance blood lipids/lipoproteins and glucose in older adults, independent of abdominal and overall adiposity. PMID- 11108417 TI - Walking speed as a good predictor for the onset of functional dependence in a Japanese rural community population. AB - OBJECTIVE: to investigate and compare the predictive values of four physical performance measures for the onset of functional dependence in older Japanese people living at home. DESIGN: a population-based prospective cohort study. SETTING: Nangai village, Akita Prefecture, Japan. METHODS: out of the population aged 65 years and older living in Nangai (n = 940) in 1992, we measured hand grip strength, one-leg standing, and usual and maximum walking speeds in 736 subjects who were independent in the five basic activities of daily living. Their functional status was assessed each year for the subsequent 6 years. The outcome event was the onset of functional dependence, defined as a new disability in one or more of the five basic activities of daily living, or death of a subject who had shown no disability at the previous follow-up. RESULTS: even after controlling for age, sex and a number of chronic conditions, lower scores on each baseline performance measure showed increased risk for the onset of functional dependence. Maximum walking speed was most sensitive in predicting future dependence for those aged 65-74 years, while usual walking speed was most sensitive for people aged > or = 75 years. CONCLUSION: walking speed was the best physical performance measure for predicting the onset of functional dependence in a Japanese rural older population. PMID- 11108418 TI - The prevalence of diabetes mellitus and quality of diabetic care in residential and nursing homes. A postal survey. AB - OBJECTIVE: to investigate the prevalence of known diabetes mellitus in care homes and the patterns of diabetes care in these institutions. DESIGN: a postal questionnaire sent to all 98 care homes in Sheffield. RESULTS: 70 care homes (71%) returned the questionnaire, indicating that 233 (8.8%) of 2648 residents were known to have diabetes. Of these, 76 (33%) were treated with diet alone, 105 (45%) with diet plus oral medication and 52 (22%) with insulin. Only seven registered nurses (2%) in the homes had certified diabetes training. Forty-two homes (60%) did not carry out a structured, diabetes-related assessment of residents on entry and only 29 (42%) had regular review of diabetic residents by a general practitioner or practice nurse. Most homes (89%) were visited by an optician, 56 (80%) also had a regular chiropody service, although 32 (46%) of these charged their residents for this service. CONCLUSIONS: the known prevalence of diabetes is similar to that reported previously. This study highlights the need for structured care with defined standards for care-home residents with diabetes. PMID- 11108419 TI - Effect of systematic review of medication by general practitioner on drug consumption among nursing-home residents. AB - BACKGROUND: nursing-home patients usually have many medical problems and often take many drugs. They are therefore at risk from drug side effects and interactions. AIMS: to evaluate the impact of a visit by a general practitioner and a comprehensive repeat prescribing review on the consumption of inappropriate drugs in nursing homes. METHOD: two general practitioners made one comprehensive visit to four randomly selected nursing homes. In each home we discussed all patients in detail with a senior member of staff. We reviewed the prescribing record of each patient and stopped items if we considered them inappropriately prescribed or unnecessary. RESULTS: repeat prescriptions were altered in 65% of patients: 51% had an item stopped and 26% had an item changed to a cheaper alternative or the dose reduced. There was a reduction in the mean number of repeat prescriptions prescribed. CONCLUSIONS: a single visit by a general practitioner to a nursing home and a comprehensive repeat prescribing review can lead to a reduction in the number of items prescribed and to substantial savings for the health service. Further rigorous, cost-effectiveness studies are needed. PMID- 11108420 TI - Infective discitis as an uncommon but important cause of back pain in older people. AB - CASE REPORTS: two elderly patients (aged 70 and 80 years) presented with severe back pain and restriction of spinal movements. Inflammatory markers were raised and in each case computed tomography findings confirmed infective discitis. One patient improved with antibiotics but the second developed paraplegia, a recognized complication of discitis. CONCLUSION: the association of back pain, restricted spinal movements and raised inflammatory markers should act as 'red flags', alerting the clinician to the presence of serious, but potentially treatable pathology. PMID- 11108421 TI - Anticoagulation of older patients. PMID- 11108422 TI - Anticoagulation of older patients. PMID- 11108423 TI - Adherence to hip protector use in elderly people requiring domicilary care is greater in fallers than non-fallers. PMID- 11108424 TI - Respite rewards. PMID- 11108425 TI - Patent foramen ovale and survival in old age. PMID- 11108426 TI - Allocation of outpatient clinic time in geriatric medicine: a survey of responses to the Royal College of Physicians' guidelines. PMID- 11108427 TI - Does frality predispose to adverse drug reactions in older patients? PMID- 11108428 TI - Tiredness: a feature of coeliac disease. PMID- 11108429 TI - Looking back. PMID- 11108430 TI - The four cornerstones of well-being in a multicultural society. PMID- 11108432 TI - Clinical pharmacokinetics of toremifene. AB - Toremifene is a chlorinated triphenylethylene derivative of tamoxifen approved for use in the treatment of patients with metastatic breast cancer. Toremifene is well tolerated in patients, and common adverse effects of this drug include vasomotor symptoms such as hot flashes and vaginal discharge. This compound is administered to patients orally at a dose of 60 mg/day, although alternative methods of administration have been investigated. Oral bioavailability is estimated to be approximately 100%. At steady state, toremifene and its metabolites are highly protein bound (>95%). Toremifene is metabolised in the liver by cytochrome P450 enzymes, and it is eliminated primarily in the faeces following enterohepatic circulation. The half-life of toremifene is approximately 5 days, and steady state is reached by 6 weeks depending on the dose given. The pharmacokinetics of toremifene have been shown to be altered by certain liver conditions, but age and kidney function do not appear to be as significant. PMID- 11108431 TI - Pharmacogenetics: a tool for individualizing antineoplastic therapy. AB - This article reviews the clinical relevance of pharmacogenetics in cancer chemotherapy, with emphasis on drugs for which genetic differences in enzyme metabolism have been demonstrated to affect patient outcome. About 10% of children with leukaemia are intolerant to mercaptopurine (6-mercaptopurine) because of genetic defects in mercaptopurine inactivation by thiopurine S methyltransferase. However, mercaptopurine dose intensity, a critical factor for outcome in patients deficient in thiopurine S-methyltransferase, can be maintained by means of thiopurine S-methyltransferase phenotyping or genotyping. Patients with reduced fluorouracil (5-fluorouracil) catabolism are more likely to be exposed to severe toxicity. The measurement of dihydropyrimidine dehydrogenase activity in patients cannot be considered fully predictive, and the role of dihydropyrimidine dehydrogenase gene variants in this syndrome has yet to be clarified. With regard to irinotecan, patients with Gilbert's syndrome phenotype have reduced inactivation of the active topoisomerase I inhibitor 7-ethyl-10 hydroxycamptothecin (SN-38) caused by a mutation in the UDP glucuronosyltransferase 1A1 gene promoter. This subset of patients is more likely to be exposed to irinotecan toxicity and could be identified by genotyping for gene promoter variants. Finally, the experience with amonafide represents a model for dose individualization approaches that use simple phenotypic probes. PMID- 11108433 TI - Pharmacokinetics and pharmacodynamics of cephalosporins in cerebrospinal fluid. AB - Largely because of their low lipophilicity, cephalosporins poorly penetrate through the blood-brain barrier, achieving relatively low cerebrospinal fluid (CSF) concentrations. However, the minimum bactericidal concentrations (MBCs) of the extended spectrum cephalosporins for common meningeal pathogens are generally low; thus, therapeutic CSF drug concentrations several-fold greater than the MBC can be achieved with currently recommended dosage regimens. However, the effectiveness of cephalosporin therapy is unreliable in patients with meningitis caused by highly penicillin-resistant pneumococci. As in other body sites, the bactericidal activity of cephalosporins in CSF predominantly depends on the time their concentrations exceed the MBC of infecting organisms (t>MBC). Experimental studies show that, for maximal efficacy, t>MBC values greater than 90% of the dosage interval are required in meningitis. Such values are usually achieved in humans with currently recommended dosage regimens because the half-lives of cephalosporins are 2- to 3-fold longer in CSF than in serum. Several advanced generation cephalosporins have shown equal efficacy in clinical trials, but only cefotaxime, ceftriaxone and ceftazidime are currently approved for the treatment of patients with bacterial meningitis. PMID- 11108436 TI - Immunogenicity of hydrolysate formulas in children (part 1). Analysis of 202 reactions. AB - Cow's milk protein hydrolyzed formulas appeared in the 1940s with the aim of decreasing or eliminating the allergenicity of cow's milk proteins, in addition to reducing the risk of sensitization. In recent years, the so-called "hypoallergenic" formulas have been developed. The use of such hydrolyzed formulas is based on the premise that predigested proteins, when fed as amino acids and peptides, provide nutrients in a nonantigenic form. Thus, protein hydrolyzed formulas have been classified as hypoallergenic. These formulas are processed by heat and enzymatic hydrolysis, and the conformational and sequential structures are more or less changed. The formulas contain peptides of lower molecular weight than the native protein source, which are thought to be less immunogenic. Hydrolyzed formulas appear to be nutritionally adequate and infants generally gain weight until they refuse the formula because of its bad taste. However, caution should be taken when such formulas are given for prolonged periods since no data are available on nutritional assessment of infants exclusively fed hydrolyzed formulas for several months. In this paper we report and discuss more than 202 reactions to different hydrolyzed formulas, including cases of anaphylactic shock and apparent life-threatening events. The cross reactivity between different hydrolyzed formulas and cow's milk proteins, and the potential immunogenicity of such formulas are discussed. We conclude that none of the hydrolyzed formulas are nonallergenic, both for allergic children and for high-risk babies. Moreover, we suggest that double-blind placebo-controlled food challenge studies in larger cohorts of babies evaluated with well-defined and well-validated diagnostic methods may establish a more reliable prevalence of allergy to hydrolyzed formulas. PMID- 11108435 TI - Articular diffusion of meloxicam after a single oral dose: relationship to cyclo oxygenase inhibition in synovial cells. AB - OBJECTIVE: To investigate the distribution of meloxicam in the human knee joint and to compare it with the inhibition of cyclo-oxygenase (COX) activity in synovial cells. DESIGN: Prospective pharmacokinetic study and in vitro laboratory investigation. PATIENTS AND PARTICIPANTS: 42 male and female patients aged 26 to 85 years hospitalised for rheumatic disease and requiring a diagnostic and/or therapeutic knee puncture. METHODS: After a single oral dose of meloxicam 15mg, synovial fluid and blood samples were collected once per patient at various intervals after administration. Meloxicam concentrations were determined by a validated high performance liquid chromatography assay, protein binding by equilibrium dialysis, and pharmacokinetic parameters were calculated by noncompartmental analysis from the mean drug concentration-time profiles. The inhibitory effect of meloxicam on COX activity was investigated separately in unstimulated or interleukin-1beta-stimulated human synovial cells from osteoarthritic patients. RESULTS: Meloxicam was found in synovial fluid at the earliest sampling time (1 hour). Peak concentrations were reached approximately 6 hours postdose in both plasma (842 microg/L) and synovial fluid (320 microg/L). A plateau was observed after the distribution phase (6 hours), corresponding to a constant ratio of drug concentration between synovial fluid and plasma of about 0.47. This ratio was higher in patients with acute inflammation (0.58) than in those with no inflammation (0.38). Meloxicam was extensively bound to protein, mainly to serum albumin. The area under the drug concentration-time curve (AUC) in plasma was more than 2.5 times that in synovial fluid. The AUC for free meloxicam was similar in plasma and synovial fluid. The 50% inhibitory concentrations (IC50) for basal and stimulated COX activity in human synovial cells were 33.7 nmol/L (11.8 microg/L) and 2.0 nmol/L (0.70 microg/L), respectively. The free concentration of meloxicam in synovial fluid was higher than the IC50 for stimulated COX activity from 6 to 36 hours postdose. CONCLUSION: On the basis of free synovial concentrations and the IC50 for stimulated COX activity, meloxicam is expected to have a long duration of action. Inhibition of COX activity is expected to be more marked in inflamed synovium compared with non-inflamed synovium. PMID- 11108437 TI - NSAID facial angioedema in a selected pediatric atopic population. AB - Epidemiological data for drug reactions in pediatric medical literature as well as in specialized periodicals are scarce. A relationship between nonsteroidal antiinflammatory drugs (NSAIDs), facial angioedema and atopic status has been described in adults. A 10-year retrospective random review of 1,007 charts of atopic children (60.9% male) attending an allergy clinic for management of asthma and/or rhinitis was carried out. Careful attention was given to the written history of NSAID facial angioedema reactions (41 out of 1007, 4.07%) and atopy was confirmed if the patient had a family history and at least one positive skin prick test (>3 mm wheal compared to glycerosaline control) to aeroallergens. Telephone recall was performed when available. Patients were classified into four age groups as follows: a) 0-5 years old; b) 6-10 years old; c) 11-15 years old; and d) 16-21 years old. NSAID facial angioedema rates were as follows: group a 10/493 (2.0%), group b 14/361 (3.8%), group c 10/121 (8.2%), and group d 7/32 (21.8%). Aspirin was the most commonly reported NSAID, and less common were pyrazolones and ibuprofen. Of the 41 patient with chart-reported reactions, 27 (66%) could be contacted by telephone. Of these, 17 patients confirmed the facial angioedema NSAID reaction occurring once or more due to inadvertent exposure. No reactions were reported in the remaining 10 patients since no other NSAID, except acetaminophen, had been used for fever or pain. In conclusion, our data show the age dependency of these reactions and its rather frequent occurrence in such selected pediatric atopic populations. Since NSAIDs are used more frequently in younger children, exposure would not be a plausible explanation for these observations. PMID- 11108438 TI - Prick-prick with fresh foods in patients with latex allergy. AB - The quick spread of AIDS and other contagious infectious diseases has resulted in what was first voluntary, and subsequently recommended and compulsory, use of protection from contact with blood or bodily fluids. This protection has been especially widespread in the healthcare field. In the in vivo diagnosis of food allergy, it has been proven that the skin prick-prick test is sometimes more sensitive than skin prick test with commercial extracts. The aim of our study was to prove that handling fresh foods prepared for the prick-prick test with latex gloves can tamper with the results in patients with latex allergy. Statistically significant differences were found (p <0.001) between patients and controls in the prick-prick tests against the different foods after handling with latex gloves. No significant differences were found in controls for each prick-prick test for food with or without manipulation with latex gloves. Significant differences were found in the group of patients when performing prick-prick with the different foods before and after manipulation with latex gloves. We also observed that there were significant differences in prick-prick test between patients with latex sensitization and nonsensitized controls, and that the results of prick-prick test varied for each patient depending on whether or not foods had been handled with latex gloves. PMID- 11108434 TI - Pharmacokinetically guided administration of chemotherapeutic agents. AB - The current practice for the dose calculation of most anticancer agents is based on body surface area in m2, although lower interpatient variation in pharmacokinetic parameters has been reported with pharmacokinetically guided administration. As chemotherapeutic agents have a narrow therapeutic window, pharmacokinetically guided administration may lead to less toxicity and higher efficacy than administration on the basis of body surface area. Pharmacokinetically guided administration, using parameters such as area under the plasma concentration-time curve (AUC), steady-state plasma drug concentration and drug exposure time above a certain plasma concentration, has been studied for many antineoplastic agents. Assessment of pharmacokinetic profiles allows the characterisation of relationships between pharmacokinetic parameters and efficacy and toxicity. AUC appears to be more closely correlated with pharmacodynamics than does the dose per unit of body surface area. In particular, the AUC-guided administration of carboplatin has been extensively studied, based on the close relationship between the renal clearance of the drug and glomerular filtration rate. Several formulae and limited sampling models have been derived to predict the AUC of carboplatin. The relationship between AUC and pharmacodynamics has also been studied for other anticancer agents, for example fluorouracil, topotecan, etoposide, cisplatin and busulfan, but all less extensively than for carboplatin. The pharmacokinetically guided administration of these agents needs to be investigated further before the use of alternative administration formulae can become standard clinical practice. Prospective studies of pharmacokinetically guided versus surface area-based administration should be performed to validate pharmacokinetic-pharmacodynamic relationships and to facilitate optimal dosage of anticancer agents in the clinic. PMID- 11108439 TI - Effect of fluticasone propionate on soluble CD30 release in patients with severe allergic asthma. AB - Previous studies have demonstrated improvements in health-related quality of life in asthmatic patients after treatment with fluticasone propionate. CD30 is a marker of Th2 lymphocytes, which are key cells in the pathogenesis of allergic inflammation. There is also a soluble form of CD30 (sCD30) released by CD30+ cells. Since serum sCD30 levels are high in allergic patients, in our study we examined the possible role of fluticasone propionate in modulating sCD30 release in patients with severe allergic asthma. In addition, we evaluated a possible correlation between sCD30 and FEV1 in these patients. To this end two groups of subjects were enrolled: 20 healthy nonatopic control subjects (group A) and 20 atopic patients with severe bronchial asthma receiving fluticasone propionate at the total dosage of 1 mg/day for 8 weeks (group B). Serum samples were examined before and after the treatment period. sCD30 serum levels were determined by the commercial ELISA-kit (Dako). The limit of detection of the assay was 1 U/ml. Our data show that sCD30 basal serum levels were significantly (p <0.05) higher in patients of group B respect to group A subjects (8.35+/-4.88 vs. <1 IU/ml, respectively). In addition, we found that sCD30 serum levels were undetectable in patients of group B after fluticasone propionate therapy. In group B a positive correlation between serum sCD30 levels and FEV1 values before fluticasone propionate treatment was noted (Rho = -0.644, p <0.005). The fluticasone propionate inhibition of sCD30 release may partly explain how fluticasone propionate exerts its antiinflammatory activity, through the modulation of Th2 cells. PMID- 11108440 TI - House dust mites in the city of Lima, Peru. AB - Since mites are the most common house dust allergens, knowledge about the species most prevalent in a region is important for diagnostic and specific immunotherapy purposes. In order to establish the prevalence of house dust mites in different city districts, 100 house dust samples were collected from different parts of Lima. Lima is a city of tropical climate located along the coast of the Pacific Ocean. The relative air humidity is 80-90% and the various districts studied are located at altitudes ranging from 37-355 meters. The mite Blomia tropicalis was the organism most frequently detected, being present in 59% of the house dust samples. Dermatophagoides pteronyssinus occupied second place (15.9%), followed by Chortoglyphus arcuatus and Tyrophagus putrescentiae. These four mites, taken together, represented more than 90% of the mites detected. No specimen of the species Dermatophagoides farinae was detected. We conclude that B. tropicalis and D. pteronyssinus are the most common house dust mites in Lima. Considering the high prevalence of B. tropicalis in Lima and the fact that its cross-reactivity with antigens of the mites of the family Pyroglyphidae is minimal, we conclude that sensitization to this mite should be investigated separately in allergic patients living in Lima. PMID- 11108441 TI - The interrelationship between production of reactive oxygen intermediates and inflammatory cytokines by peripheral blood leukocytes in vitro from children with food allergy. AB - In this study, the activity of neutrophils measured using the chemiluminescence test was estimated in 49 children with food allergy. It was shown that average chemiluminescence values without, as well as after, stimulation by formyl methionyl-leucine phenylalanine and phorbol myristate acetate were considerably higher in children with allergy in comparison to those in the control group. The correlation between the production of reactive oxygen intermediates and interleukin (IL)-4, IL-5 and IL-10 concentration suggests that an autocrine regulatory mechanism may play a role in allergy in children. PMID- 11108442 TI - Cytofluorometric analysis of the neutrophil-specific BH2-Ag on cells from neonates to adults. AB - Monoclonal antibody (MAB) BH2C6 recognizes a plasma membrane antigen, the BH2-Ag, specifically expressed by human neutrophils. While studies with peripheral blood and bone marrow from healthy adults clearly demonstrate the absence of BH2-Ag from other cellular components except neutrophils, they also indicate that the BH2-Ag is expressed more strongly by mature than immature neutrophils. The purpose of this study was to determine the expression of the BH2-Ag by peripheral blood neutrophils from premature newborns to adults. Seventy-two donors were studied in six age groups: newborns <36 weeks of gestational age; newborns >36 weeks of gestational age; 0.5-2 years; 4-8 years; 12-17 years; >30 years. Expression of the BH2-Ag by peripheral blood neutrophils was examined by cytofluorography using MAB BH2-C6 directly labeled with fluorescein isothiocyanate (FITC). Neutrophils were reacted in parallel with FITC-MAB directed against CD11b, the alpha-chain of the CD11b/CD18 antigen (CR3). BH2-Ag is expressed by 98.3-99.6% of the neutrophils in all groups, and is absent on other blood cells, including those of very premature newborns. Statistical comparisons with respect to the mean fluorescence intensity of the FITC-MAB BH2C6 bound did not support a significant difference in the expression of BH2-Ag in any age group. CD11b expression was also detected in every individual studied and its mean fluorescence intensity correlated significantly with that of BH2Ag (p <0.001). The uniform presence of BH2Ag in every individual including a very premature infant suggests that BH2-Ag is likely to be an essential component of neutrophil development in humans. A highly significant correlation between the mean fluorescence intensity obtained with MAB BH2C6 and MAB CD11b suggests a possible interactive role of the two antigens in neutrophil development and/or function. PMID- 11108443 TI - Local nasal immunotherapy and bronchial hyperreactivity in seasonal allergic rhinitis: an observational pilot study. AB - The clinical efficacy of local nasal immunotherapy (LNIT) in patients with allergic rhinitis is amply documented. The aim of the study was to determine the effect on bronchial hyperresponsiveness (BHR) assessed at baseline and after 3 years of LNIT or pharmacological treatment alone. Forty-three randomized patients with allergic oculorhinitis were enrolled (26 positive to Graminaceae and 17 to Parietaria judaica pollens). All patients were asked whether they were willing to follow a 3-year treatment course involving preseasonal LNIT with a powder extract of Graminaceae or Parietaria pollens. Twenty-four patients (16 allergic to Graminaceae and eight to Parietaria) selected LNIT and the other 19 opted for symptomatic pharmacological treatment only. The latter was considered the control group. On the basis of positive or negative bronchial hyperresponsiveness and the LNIT, four subgroups were established and followed in open conditions, during which a record was kept of symptom scores, drug use, spirometry and methacholine test findings. After 3 years, patients treated with LNIT had a significant reduction of symptoms and drug intake. In the controls, symptoms worsened, thus requiring more drugs to control them. Bronchial hyperresponsiveness significantly improved in hyperreactive patients receiving LN7IT 10 of whom were no longer hyperreactive and one at a higher threshold. Among controls, bronchial hyperresponsiveness did not change, with the exception of three nonreactive patients who became hyperreactive, one of them with asthma. These findings confirm the efficacy of LNIT in allergic rhinitis suggesting that it might have systemic activity interfering with bronchial hyperresponsiveness and hence the onset of bronchial asthma. PMID- 11108444 TI - Candida albicans allergen immunotherapy in recurrent vaginal candidiasis. AB - Recurrent vaginal candidiasis is a worldwide problem affecting millions of women. Candida albicans is a potent allergen in some situations and it has been suggested that local hypersensitivity to this yeast can be a factor in the prolongation of the disease. A small number of studies show some benefits of allergen immunotherapy in these patients. We evaluated the efficacy of C. albicans allergen immunotherapy in women with immediate skin test positive for this fungus. We conducted a prospective cohort study of women with recurrent vaginal candidiasis referred by gynecologists to an allergist. All patients had established positive C. albicans in vaginal cultures and were unresponsive to other modes of treatment. None had diabetes or AIDS. In their allergy evaluation, they had an immediate (prick or intradermal) skin test positive to C. albicans. These women were offered the option of C. albicans allergen immunotherapy for a period of 24 months. The efficacy of therapy was evaluated by comparison of the mean incidence of episodes per year before and after immunotherapy Over a period of 33 months, 34 women were enrolled (mean age 33.5 years; range 18-57). They were treated with weekly injections of C. albicans allergenic extract over a mean (+/- SD) period of 17.4+/-7.2 months (4-24 months). Twenty-two of the women had improvement in symptoms. There were complete responses, i.e., absence of acute episodes for 2 years, in nine patients (26%) and partial responses, i.e., decrease in the number and intensity of episodes, in 13 patients (38%). Overall there was a 64% improvement in these patients (95% CI: 46.5-79.9). Eleven patients showed no improvement and one worsened. The effects were evident after 2 12 months of therapy (mean 3.5 months). The mean incidence of episodes of vaginitis per year decreased from a mean (+/- SD) of 8.5+/-2.6 to 3.6+/-4.3 (p = 0.000). The study also showed that the majority of patients were atopic (70%), had allergic rhinitis (67%) and familial history of allergy (70%). Our results suggest that in a subgroup of patients with recurrent vaginal candidiasis, C. albicans allergen immunotherapy lowers the number and intensity of episodes. PMID- 11108445 TI - Pollen mixtures used as health food may be a harmful source of allergens. AB - We describe herein one case of systemic anaphylaxis due to the ingestion of an undefined mixture of pollens, sold as a dietary supplement. The patient, who suffered from rhinoconjunctivitis due to grass pollen (with sensitization to several trees), had a severe episode of anaphylaxis immediately after eating this health food. The episode required emergency care. We attempted to study the pollen mixture responsible, but no pollen granules could be identified. We prepared a solid phase with the pollen mixture, and we observed a RAST positivity with the patient's serum and pools of sera containing specific IgE to trees. Furthermore, a RAST-inhibition assay of the patient's serum showed highly positive results with grasses, birch, alder and Compositae. Therefore, we concluded that the pollen mixture contained determinants capable of cross reacting with the patient's IgE. This case report is evidence of the possible risks due to the use of undefined herbal products by allergic patients. PMID- 11108446 TI - Selective T-cell deficiency in Turner's syndrome. AB - The case of a 29-year-old Caucasian woman with 45 X0 karyotype, known as Turner's syndrome, and a recently diagnosed selective T-cell deficiency is reported. The main clinical features of the patient were recurrent sinopulmonary infections and a negative skin test with seven common recall antigens. Laboratory findings included lymphocytopenia, highly elevated CD45RA/CD45R0 ratio, as well as reduced expression of the co-stimulatory molecules CD154, CD86, CD80 and CD28 on CD4+ cells in combination with disturbed lymphocyte transformation in vitro. Markedly decreased levels of interleukin (IL)-2R, both on lymphocyte surface as well as the soluble analog, suggest a new form of x-linked immunodeficiency associated with Turner's syndrome. PMID- 11108447 TI - Morphological, biochemical and molecular biology approaches for the diagnosis of lysosomal storage diseases. PMID- 11108448 TI - Epizootic malignant catarrhal fever in three bison herds: differences from cattle and association with ovine herpesvirus-2. AB - Three bison herds in Colorado experienced high mortality from malignant catarrhal fever (MCF). In comparison with cattle, the bison had a more rapidly progressive disease, fewer clinical signs, and milder inflammatory histologic lesions. There was consistent association with ovine herpesvirus-2 (OHV-2). Contact with sheep was not consistent. Of 17 animals in herd A, 15 died of acute MCF; 1 was slaughtered while healthy; and 1 developed clinical signs of MCF, was treated with corticosteroids and antibiotics, and died of fungal abomasitis and rhinitis after 5 months. In herds B and C, approximately 300 of 900 and 18 of 20 died of MCF following brief clinical disease. The nearest sheep were 1 mile away from herd A, but direct contact with sheep could be documented in herds B and C. Complete gross and histologic examinations were conducted on 34 animals, including all animals in herd A, and MCF was diagnosed in 31. In addition, field necropsies were performed on all dead animals in herd B and most in herd C and MCF was diagnosed on the basis of the gross lesions in most animals. Clinical signs of each animal in herd A were recorded. Illness was brief, usually 8-48 hours. Clinical signs were subtle; separation from the herd was often observed. In all 3 herds, hemorrhagic cystitis and multifocal ulceration of the alimentary tract were consistently found at necropsy. Mild lymphocytic vasculitis was present in multiple organs. Ovine herpesvirus-2 was found by polymerase chain reaction (PCR) in 71 of 105 formalin-fixed tissue specimens from 29 of 31 animals with MCF. In herd A, blood samples from 13 animals were collected at 5 time points and tested by PCR for the presence of OHV-2 viral sequences in peripheral blood leukocytes. Nine bison with a positive PCR test and 4 with negative results prior to clinical illness died of MCF. PMID- 11108449 TI - Production and evaluation criteria of specific monoclonal antibodies to the hemagglutinin of the H7N2 subtype of avian influenza virus. AB - To enhance the rapidity in diagnosing the spread of avian influenza virus (AIV) in chicken layer flocks, studies were initiated to develop more sensitive and specific immunological and molecular methods for the detection of AIV. In this study, the purification of the hemagglutinin protein (H) from field isolates of H7N2, the production of monoclonal antibodies (MAbs), and their evaluation as diagnostic reagents are reported. Hybridomas were generated by fusion of SP2/0 Ag14 myelomas and spleen cells from immunized mice. Hybridomas secreting antibodies specific for the H protein were assayed by an ELISA and cloned using limiting dilution. The MAbs produced were characterized by hemagglutination inhibition (HI), immunohistochemistry (IHC), indirect fluorescent antibody assay (IFA), Western blots, and IFA flow cytometry using various AIV subtypes (i.e., H4N2, H5N3, H7N2). Of the various MAbs assayed, 6 had consistent and reproducible results in each of the assays used. The results obtained in this investigation enhanced the usage of the MAbs to viral H protein in the surveillance of AIV in chickens. PMID- 11108450 TI - Interstitial pneumonia in feedlot cattle: concurrent lesions and lack of immunohistochemical evidence for bovine respiratory syncytial virus infection. AB - The objectives of this study were to describe the nature and distribution of microscopic lung lesions in feedlot cattle with interstitial pneumonia and to determine whether bovine respiratory syncytial virus (BRSV) antigen was present in affected lungs. Lungs with macroscopic lesions compatible with interstitial pneumonia were collected from cattle from 5 west-central Saskatchewan feedlots that had been on feed for greater than 60 days at the time of death. Interstitial pneumonia was most consistently present in dorsal portions of caudal lung lobes and in 21/28 cases (75%) had a multifocal to coalescing distribution. All 28 lungs exhibited hyaline membrane formation and some degree of type II alveolar epithelial cell hyperplasia, consistent with an acute to subacute duration. Twenty-one of 28 cases (75%) had concurrent bronchopneumonia in at least 1 lung lobe; bronchopneumonia was grossly evident in 9/28 cases (32%). Chronic bronchitis or bronchiolitis was present in at least 1 section in 12/28 (43%) of the lungs, and 25/28 (89%) had at least 1 focus of bronchiolitis fibrosa obliterans. Bronchopneumonia and bronchiolitis fibrosa obliterans were markedly less common in 10 sets of bovine lungs obtained from an abattoir. Bovine respiratory syncytial virus antigen was demonstrated using immunohistochemistry in 2/28 cases and was associated with bronchiolar epithelial necrosis that was more severe than the bronchiolar lesions in the BRSV antigen-negative cases. Interstitial pneumonia in feedlot cattle in this study was more frequently associated with suppurative bronchopneumonia and bronchiolitis fibrosa obliterans than with BRSV infection. PMID- 11108451 TI - Detection of Akabane viral antigens in spontaneous lymphohistiocytic encephalomyelitis in cattle. AB - A 5-month-old Japanese black bull calf and twenty-seven 1-27-day-old calves exhibiting neurological signs between August and October 1998 were examined. The bull calf exhibited rapid breathing, fever, hypersensitivity, and ataxia and was euthanized 4 days after the onset of symptoms. The 27 calves primarily exhibited ataxia, and 15 had arthrogryposis. Histological examination of the bull calf revealed perivascular infiltraction by mononuclear cells, diffuse to multifocal gliosis, and neuronal necrosis in the brain and spinal cord. Multiple malacic foci were found in the midbrain in 5 cases. In contrast, in the 15 calves necropsied in October, there were fewer inflammatory changes, but there was neuronal cell loss in the ventral horn and a decrease in myelinated axons in the lateral and ventral funiculi. Immunohistochemical examination using a rabbit antiserum against Akabane virus strain OBE-1 revealed a large amount of viral antigen in the degenerating neurons and glial cells of the bull calf, mainly in the spinal gray matter. Small amounts of viral antigen in swollen axons and a few glial cells were found in 5 of 27 calves. Thirteen of the 27 calves had high neutralization antibody titers against the Akabane virus, whereas there was no significant antibody titer in most of the calves necropsied during August. The present study revealed that viral antigen detection was very useful for the diagnosis of Akabane diseases in the 5-month-old bull calf that was suspected to be infected postnatally, while it had limited usefulness in the other young calves. PMID- 11108452 TI - Osteopenic disease in growing pigs: diagnostic methods using serum and urine calcium and phosphorus values, parathoromone assay, and bone analysis. AB - This research was performed to evaluate the utility of several serum and urine parameters as well as bone ash and plasma parathormone assay to diagnose and monitor diet-related osteopenia in growing pigs. Five diets were tested as follows: calcium-deficient, phosphorus-replete; moderate-deficiency of calcium and phosphorus; marked deficiency of calcium and phosphorus; calcium replete, phosphorus deficient; and vitamin D deficient. Parameters monitored included serum calcium and phosphorus as well as ratios of urine calcium to creatinine, phosphorus to creatinine, calcium to phosphorus, and percent fractional excretions of calcium and phosphorus. Plasma parathormone (PTH) levels were monitored in 2 of 3 experiments. Osteopenic bone differences at necropsy were evaluated by bone density, percent ash, ash per milliliter bone, calcium per milliliter bone, and phosphorus per milliliter bone. Marked change in urine mineral parameters, especially the calcium-to-phosphorus ratio, typically occurred within 1 to 2 days of treatment and preceded significant change in serum mineral or plasma PTH by 2 to 3 weeks. When monitored, plasma PTH levels were elevated following treatment, which confirms the hyperparathyroid state induced by the test diets. Significant differences in bone mineralization between control and treatment diets at necropsy were generally observed. The results of this study indicate that the analysis of urine minerals offers an early, noninvasive technique to investigate diet-associated osteopenic disease in growing pigs, which can be supported further by bone mineral analysis at postmortem using techniques herein described. Several urine mineral reference intervals for application to field investigations are included. Research into application of similar techniques to evaluate calcium and phosphorus homeostasis in pigs of all ages, including gestating and lactating gilts and sows, appears warranted. PMID- 11108453 TI - Ribotyping and restriction endonuclease analysis reveal a novel clone of Bordetella bronchiseptica in seals. AB - The goal of the present study was to characterize, by ribotyping and restriction endonuclease analysis (REA), 35 phocine Bordetella bronchiseptica isolates and to ascertain their relationship to one another and to isolates acquired from other host species. Thirty-four isolates were obtained in Scotland during a 10-year period encompassing the 1988 epizootic; the remaining isolate was obtained independently in Denmark. All phocine isolates had an identical Pvu II ribotype unique from the 18 ribotypes previously detected in strains from heterologous hosts. Alternative restriction enzymes, useful for subgrouping strains within Pvu II ribotypes, also failed to discriminate among isolates from seals. The exclusive occurrence of a single ribotype of B. bronchiseptica in a particular host species has not been previously observed. Similarly, REA based on either HinfI or Dde I profiles did not reveal detectable polymorphisms, although unique patterns were readily distinguished among a limited number of isolates from other host species. This is the first report demonstrating the utility of REA using frequently cutting enzymes for discrimination of B. bronchiseptica strains. These data suggest that B. bronchiseptica-induced respiratory disease in seals along the Scottish shore may be due to the circulation of a single, unique clone. PMID- 11108454 TI - In vitro susceptibility of porcine respiratory pathogens to tilmicosin. AB - Bacterial isolates obtained from swine with various clinical diseases were tested for susceptibility to tilmicosin by minimum inhibitory concentration (MIC) and Kirby-Bauer disk diffusion tests using National Committee on Clinical Laboratory Standards methodology. The tilmicosin MIC90 was < or =0.125 microg/ml for Erysiopelothrix rhusiopathiae, < or = 1 microg/ml for Haemophilus parasuis isolates, 8 microg/ml for Actinobacillus suis and Pasteurella multocida type A, 16 microg/ml for toxigenic and nontoxigenic P. multocida type D, 64 microg/ml for Bordetella bronchiseptica, and >128 microg/ml for Staphylococcus hyicus and Streptococcus suis. The results of disk diffusion testing matched well with the MIC results for each pathogen. This in vitro survey of tilmicosin activity against various swine isolates suggests that further clinical evaluation of tilmicosin in swine may be warranted for disease associated with E. rhusiopathiae, H. parasuis, and A. suis but not B. bronchiseptica, S. suis, or S. hyicus. PMID- 11108455 TI - Culture of strategically pooled bovine fecal samples as a method to screen herds for paratuberculosis. AB - Fecal samples from 733 cows in 11 dairy herds with a low prevalence of paratuberculosis were cultured for the presence of Mycobacterium avium subsp. paratuberculosis both individually and after combining (pooling) in groups of 5. The culture procedure was the modified Jorgensen method, which uses NaOH and oxalic acid for decontamination and modified Lowenstein-Jensen agar slants for cultivation. Pooling was performed by mixing fecal samples from 5 animals ordered by age, herein referred to as strategic pooling. Culture of individual fecal samples detected M. a. paratuberculosis infections in 43 of the 733 cows and 7 of 11 infected herds (herd sensitivity = 64%). Culture of pooled fecal samples detected M. a. paratuberculosis in 28 of 151 pooled samples representing 8 of the infected 11 herds (herd sensitivity = 73%). Feces of the 43 culture-positive cows was included in 32 pools: of these 32 pools, 26 were culture positive and 6 were culture negative. In addition to the 26 positive pools containing feces from cows that were found culture positive on individual fecal samples, another 2 pools were culture positive, although comprised of feces from cows with negative results after culture of individual fecal samples. From the total of 45 infected cows that were found (43 by individual fecal culture and an additional 2 by pooled fecal culture), individual fecal culture detected 43 of these 45 (96%), while pooled fecal culture detected 39 (87%). Culture of strategically pooled fecal samples using the modified Jorgensen method was equivalent in herd sensitivity to the culture of individual fecal samples and is significantly less expensive. PMID- 11108456 TI - Pathogenesis of liver lesions caused by experimental infection with Piscirickettsia salmonis in juvenile Atlantic salmon, Salmo salar L. AB - Piscirickettsia salmonis, the etiologic agent of salmonid rickettsial septicemia (SRS), or piscirickettsiosis, causes substantial economic losses to the salmon industry. The pathogenesis of the disease has not been fully characterized. The aim of this study is to describe the hepatic lesions associated with experimental P. salmonis infection in Atlantic salmon juveniles. Fish were maintained in fresh water and inoculated intraperitoneally (IP), orally, or on the gill surface with P. salmonis. A group of uninfected fish was kept as control. Liver samples from 5 fish in each inoculated group and 3 controls were collected weekly and processed for histological and immunohistochemical examination. Thickening of the liver capsule by inflammatory cells was a characteristic histologic feature of IP inoculated fish. Three weeks post-IP inoculation, 8 fish had died and 2 fish were sampled. Histological changes at this time consisted of vasculitis, presence of fibrin thrombi, vacuolated hepatocytes and focal areas of necrosis. Leukocytes containing intracytoplasmic basophilic microorganisms were seen within hepatic sinusoids. Vasculitis and intracytoplasmic vacuoles were prominent features in fish inoculated orally and on the gill surface. The presence of P. salmonis within hepatocellular vacuoles, endothelial cells, and leucocytes was confirmed by immunohistochemistry. The intracellular location of P. salmonis and the vascular damage seen in infected fish are characteristic of rickettsial infections. Histological lesions induced by experimental infection with P. salmonis using the oral and gill surface routes were similar to those observed in natural outbreaks of piscirickettsiosis. The tropism of P. salmonis for endothelial cells explains the vascular lesions observed in SRS, whereas hepatic lesions are due to ischemic necrosis and direct injury by intracytoplasmic organisms. PMID- 11108457 TI - Polymerase chain reaction for definitive identification of Yersinia ruckeri. AB - Yersinia ruckeri causes enteric red mouth (ERM) disease in salmonids. Serologic identification of Y. ruckeri is hampered by cross-reactivity with other bacterial isolates of fish origin. Oligonucleotide primers incorporating Y. ruckeri unique sequences were designed to amplify a 409 bp fragment of Y. ruckeri 16S rDNA. The primers did not amplify other genetically related Yersinia or a wide variety of other aquatic or piscine bacteria. This assay provides a rapid, definitive identification of Y. ruckeri that is not subject to the variability inherent in serologic methods. PMID- 11108458 TI - Utilization of a rate enhancement hybridization buffer system for rapid in situ hybridization for the detection of porcine circovirus in cell culture and in tissues of pigs with postweaning multisystemic wasting syndrome. AB - A rapid in situ hybridization (ISH) technique for the detection of porcine circovirus (PCV) nucleic acid in cell culture and formalin-fixed paraffin embedded tissues was developed. A fluorescein-labeled RNA probe was transcribed from a plasmid containing 530 bp of the ORF1 of a PCV isolated from a pig with postweaning multisystemic wasting syndrome (PMWS). Hybridization using standard hybridization buffer was performed at 42 C for 16 hours and was compared to hybridization using rate enhancement hybridization (REH) buffer at 67 C for 2 hours. Hybridization was detected with an alkaline phosphatase-conjugated antifluorescein antibody. In both cultured cells and tissues from pigs with PMWS, the signal intensity and number of labeled cells in sections hybridized with REH buffer were equal to those of sections hybridized with standard hybridization buffer. The total time required for ISH using the REH buffer is 7-8 hours, thus making this protocol suitable for application in routine PCV diagnosis. PMID- 11108459 TI - Nephrotoxicosis in a cat following ingestion of Asiatic hybrid lily (Lilium sp). PMID- 11108460 TI - Bovine viral diarrhea virus in New World camelids. AB - A virus known to cause multiple problems in cattle, bovine viral diarrhea virus, was isolated from 3 different cases in New World camelids. Virus isolation, immunoperoxidase staining, and fluorescent antibody staining were used to detect the virus. The herds involved were screened for antibody titers to bovine viral diarrhea and virus isolation from the buffy coat. Bovine viral diarrhea virus should be considered as a cause of death in young and old New World camelids. PMID- 11108461 TI - An autosomal recessive, lethal, neurologic disease of Gordon Setter puppies. AB - This report details clinical, necropsy, and pedigree data on an inherited, lethal, neurologic disease of young Gordon Setters. This disorder is characterized by an early age of onset, gait and postural abnormalities, progressive weakness, and recumbency by 5-6 weeks of age. Although clinically distinctive, postmortem changes in affected pups were minimal. Gross lesions were not observed. Microscopic changes were subtle and consisted of astrocyte swelling, primarily in the cerebrocortical and cerebellar white matter, and white matter tracts of the brainstem. Immunohistochemistry for glial fibrillary acidic protein revealed a marked increase in the number and staining intensity of astrocyte cytoplasmic processes in affected pups compared with age-matched controls. Neither cerebral inflammation nor neuronal necrosis was identified. Pedigree analysis of affected litters demonstrated an autosomal recessive mode of inheritance. A diagnosis of this heritable disease should be based on the early age of onset (3-4 weeks of age), characteristic clinical signs, rapid progression to recumbency by 5-6 weeks of age, identification of swollen astrocytes primarily in the cerebellar and cerebrocortical white matter and white matter tracts of the brainstem, and the exclusion of other disease processes. PMID- 11108462 TI - Adenovirus hepatitis in a boa constrictor (Boa constrictor). AB - A boa constrictor was submitted for postmortem evaluation. At necropsy, there were no substantial lesions except in the liver. Light microscopy revealed severe multifocal to coalescing coagulative necrotic hepatitis, with basophilic and eosinophilic intranuclear inclusions in hepatocytes within the necrotic foci. The histopathological findings suggested a viral hepatitis. An adenoviral infection was diagnosed by means of transmission electronic microscopy and in situ hybridization techniques. PMID- 11108463 TI - Serotyping of Mannheimia (Pasteurella) haemolytica isolates from the upper Midwest United States. AB - Mannheimia (Pasteurella) haemolytica biotype A serotype1 (A1) is the primary bacterial agent responsible for the clinical signs and pathophysiologic events in bovine pneumonic pasteurellosis. The goal of this study was to determine the prevalence of other serotypes of M. haemolytica biotype A organisms obtained from the upper Midwest diagnostic laboratories. A total of 147 M. haemolytica isolates were collected from Minnesota, South Dakota, and Michigan. Isolates were tested against M. haemolytica antisera obtained from the National Animal Disease Center, Ames, Iowa. Results indicated that M. haemolytica serotype 1 represented approximately 60%, serotype 6 represented 26%, and serotype 2 represented 7% of the total examined isolates. In addition, 7% of the isolates were serotype 9, 11, or untypable. This finding suggests that M. haemolytica serotypes other than serotype 1 can be isolated from the lung lesions of diseased cattle and seem to be capable of causing the pathologic changes observed in the lung with pneumonic pasteurellosis. PMID- 11108464 TI - Immunohistochemical diagnosis of chronic wasting disease in preclinically affected elk from a captive herd. AB - An immunohistochemical (IHC) method was used to test brain tissues from 17 elk in a captive herd in which chronic wasting disease (CWD) had previously occurred. The IHC technique detects the protease-resistant prion protein (PrP-res), which is considered a disease-specific marker for transmissible spongiform encephalopathies (TSE), regardless of the species affected. Of the 17 elk tested, 10 were positive by IHC. Only 2 of these 10 animals had shown clinical signs and histologic lesions of CWD, and an additional animal had histologic lesions only. The most consistently IHC-positive tissue was medulla oblongata, especially the obex. These results show that the PrP-res IHC test on brain tissue, specifically medulla oblongata at the obex, should be considered an essential component of any surveillance study intended to determine the incidence of CWD in captive or free ranging cervids. PMID- 11108465 TI - Comparison of virus isolation, reverse transcription-polymerase chain reaction, immunohistochemistry, and in situ hybridization for the detection of porcine reproductive and respiratory syndrome virus from naturally aborted fetuses and stillborn piglets. AB - Virus isolation, reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, and in situ hybridization methods were compared for the detection of porcine reproductive and respiratory syndrome virus (PRRSV). Seven aborted fetuses and 6 stillborn piglets naturally infected with PRRSV were used in the study. Viral antigen and viral nucleic acid were detected in macrophages and dendritic cells in the spleen, tonsil, lymph nodes, and thymus; in macrophages of liver, heart, and lung; and in endothelial cells and myocytes of the heart. Viral antigen and viral nucleic acid were most consistently detected in the spleen. Of the 13 samples, 6 were positive for PRRSV by all 4 techniques. Four (31%) samples were positive for PRRSV by RT-PCR, in situ hybridization, and virus isolation. Two (15%) samples were positive for PRRSV by virus isolation, RT PCR, and in situ hybridization. One (8%) was positive for PRRSV by virus isolation and RT-PCR. The RT-PCR identified the presence of PRRSV more frequently than the other methods. However, when only formalin-fixed tissues are submitted, immunohistochemistry and in situ hybridization would be useful methods for the detection of PRRSV antigen and nucleic acid. PMID- 11108466 TI - Algorithms for identifying Boolean networks and related biological networks based on matrix multiplication and fingerprint function. AB - Due to the recent progress of the DNA microarray technology, a large number of gene expression profile data are being produced. How to analyze gene expression data is an important topic in computational molecular biology. Several studies have been done using the Boolean network as a model of a genetic network. This paper proposes efficient algorithms for identifying Boolean networks of bounded indegree and related biological networks, where identification of a Boolean network can be formalized as a problem of identifying many Boolean functions simultaneously. For the identification of a Boolean network, an O(mnD+1) time naive algorithm and a simple O (mnD) time algorithm are known, where n denotes the number of nodes, m denotes the number of examples, and D denotes the maximum in degree. This paper presents an improved O(momega-2nD + mnD+omega-3) time Monte Carlo type randomized algorithm, where omega is the exponent of matrix multiplication (currently, omega < 2.376). The algorithm is obtained by combining fast matrix multiplication with the randomized fingerprint function for string matching. Although the algorithm and its analysis are simple, the result is nontrivial and the technique can be applied to several related problems. PMID- 11108467 TI - Algorithms for extracting structured motifs using a suffix tree with an application to promoter and regulatory site consensus identification. AB - This paper introduces two exact algorithms for extracting conserved structured motifs from a set of DNA sequences. Structured motifs may be described as an ordered collection of p > or = 1 "boxes" (each box corresponding to one part of the structured motif), p substitution rates (one for each box) and p - 1 intervals of distance (one for each pair of successive boxes in the collection). The contents of the boxes--that is, the motifs themselves--are unknown at the start of the algorithm. This is precisely what the algorithms are meant to find. A suffix tree is used for finding such motifs. The algorithms are efficient enough to be able to infer site consensi, such as, for instance, promoter sequences or regulatory sites, from a set of unaligned sequences corresponding to the noncoding regions upstream from all genes of a genome. In particular, both algorithms time complexity scales linearly with N2n where n is the average length of the sequences and N their number. An application to the identification of promoter and regulatory consensus sequences in bacterial genomes is shown. PMID- 11108468 TI - Potential energy function for continuous state models of globular proteins. AB - One of the approaches to protein structure prediction is to obtain energy functions which can recognize the native conformation of a given sequence among a zoo of conformations. The discriminations can be done by assigning the lowest energy to the native conformation, with the guarantee that the native is in the zoo. Well-adjusted functions, then, can be used in the search for other (near-) natives. Here the aim is the discrimination at relatively high resolution (RMSD difference between the native and the closest nonnative is around 1 A) by pairwise energy potentials. The potential is trained using the experimentally determined native conformation of only one protein, instead of the usual large survey over many proteins. The novel feature is that the native structure is compared to a vastly wider and more challenging array of nonnative structures found not only by the usual threading procedure, but by wide-ranging local minimization of the potential. Because of this extremely demanding search, the native is very close to the apparent global minimum of the potential function. The global minimum property holds up for one other protein having 60% sequence identity, but its performance on completely dissimilar proteins is of course much weaker. PMID- 11108469 TI - Optimal amnesic probabilistic automata or how to learn and classify proteins in linear time and space. AB - Statistical modeling of sequences is a central paradigm of machine learning that finds multiple uses in computational molecular biology and many other domains. The probabilistic automata typically built in these contexts are subtended by uniform, fixed-memory Markov models. In practice, such automata tend to be unnecessarily bulky and computationally imposing both during their synthesis and use. Recently, D. Ron, Y. Singer, and N. Tishby built much more compact, tree shaped variants of probabilistic automata under the assumption of an underlying Markov process of variable memory length. These variants, called Probabilistic Suffix Trees (PSTs) were subsequently adapted by G. Bejerano and G. Yona and applied successfully to learning and prediction of protein families. The process of learning the automaton from a given training set S of sequences requires theta(Ln2) worst-case time, where n is the total length of the sequences in S and L is the length of a longest substring of S to be considered for a candidate state in the automaton. Once the automaton is built, predicting the likelihood of a query sequence of m characters may cost time theta(m2) in the worst case. The main contribution of this paper is to introduce automata equivalent to PSTs but having the following properties: Learning the automaton, for any L, takes O (n) time. Prediction of a string of m symbols by the automaton takes O (m) time. Along the way, the paper presents an evolving learning scheme and addresses notions of empirical probability and related efficient computation, which is a by product possibly of more general interest. PMID- 11108470 TI - Contig selection in physical mapping. AB - In physical mapping, one orders a set of genetic landmarks or a library of cloned fragments of DNA according to their position in the genome. Our approach to physical mapping divides the problem into smaller and easier subproblems by partitioning the probe set into independent parts (probe contigs). For this purpose we introduce a new distance function between probes, the averaged rank distance (ARD) derived from bootstrap resampling of the raw data. The ARD measures the pairwise distances of probes within a contig and smoothes the distances of probes across different contigs. It shows distinct jumps at contig borders. This makes it appropriate for contig selection by clustering. We have designed a physical mapping algorithm that makes use of these observations and seems to be particularly well suited to the delineation of reliable contigs. We evaluated our method on data sets from two physical mapping projects. On data from the recently sequenced bacterium Xylella fastidiosa, the probe contig set produced by the new method was evaluated using the probe order derived from the sequence information. Our approach yielded a basically correct contig set. On this data we also compared our method to an approach which uses the number of supporting clones to determine contigs. Our map is much more accurate. In comparison to a physical map of Pasteurella haemolytica that was computed using simulated annealing, the newly computed map is considerably cleaner. The results of our method have already proven helpful for the design of experiments aimed at further improving the quality of a map. PMID- 11108471 TI - RNA pseudoknot prediction in energy-based models. AB - RNA molecules are sequences of nucleotides that serve as more than mere intermediaries between DNA and proteins, e.g., as catalytic molecules. Computational prediction of RNA secondary structure is among the few structure prediction problems that can be solved satisfactorily in polynomial time. Most work has been done to predict structures that do not contain pseudoknots. Allowing pseudoknots introduces modeling and computational problems. In this paper we consider the problem of predicting RNA secondary structures with pseudoknots based on free energy minimization. We first give a brief comparison of energy-based methods for predicting RNA secondary structures with pseudoknots. We then prove that the general problem of predicting RNA secondary structures containing pseudoknots is NP complete for a large class of reasonable models of pseudoknots. PMID- 11108472 TI - NOTUNG: a program for dating gene duplications and optimizing gene family trees. AB - Large scale gene duplication is a major force driving the evolution of genetic functional innovation. Whole genome duplications are widely believed to have played an important role in the evolution of the maize, yeast, and vertebrate genomes. The use of evolutionary trees to analyze the history of gene duplication and estimate duplication times provides a powerful tool for studying this process. Many studies in the molecular evolution literature have used this approach on small data sets, using analyses performed by hand. The rapid growth of genetic sequence data will soon allow similar studies on a genomic scale, but such studies will be limited unless the analysis can be automated. Even existing data sets admit alternative hypotheses that would be too tedious to consider without automation. In this paper, we describe a program called NOTUNG that facilitates large scale analysis, using both rooted and unrooted trees. When tested on trees analyzed in the literature, NOTUNG consistently yielded results that agree with the assessments in the original publications. Thus, NOTUNG provides a basic building block for inferring duplication dates from gene trees automatically and can also be used as an exploratory analysis tool for evaluating alternative hypotheses. PMID- 11108473 TI - A computational method for NMR-constrained protein threading. AB - Protein threading provides an effective method for fold recognition and backbone structure prediction. But its application is currently limited due to its level of prediction accuracy and scope of applicability. One way to significantly improve its usefulness is through the incorporation of underconstrained (or partial) NMR data. It is well known that the NMR method for protein structure determination applies only to small proteins and that its effectiveness decreases rapidly as the protein mass increases beyond about 30 kD. We present, in this paper, a computational framework for applying underconstrained NMR data (that alone are insufficient for structure determination) as constraints in protein threading and also in all-atom model construction. In this study, we consider both secondary structure assignments from chemical shifts and NOE distance restraints. Our results have shown that both secondary structure assignments and a small number of long-range NOEs can significantly improve the threading quality in both fold recognition and threading-alignment accuracy, and can possibly extend threading's scope of applicability from homologs to analogs. An accurate backbone structure generated by NMR-constrained threading can then provide a great amount of structural information, equivalent to that provided by many NMR data; and hence can help reduce the number of NMR data typically required for an accurate structure determination. This new technique can potentially accelerate current NMR structure determination processes and possibly expand NMR's capability to larger proteins. PMID- 11108474 TI - Matching simulation and experiment: a new simplified model for simulating protein folding. AB - Simulations of simplified protein folding models have provided much insight into solving the protein folding problem. We propose here a new off-lattice bead model, capable of simulating several different fold classes of small proteins. We present the sequence for an alpha/beta protein resembling the IgG-binding proteins L and G. The thermodynamics of the folding process for this model are characterized using the multiple multihistogram method combined with constant temperature Langevin simulations. The folding is shown to be highly cooperative, with chain collapse nearly accompanying folding. Two parallel folding pathways are shown to exist on the folding free energy landscape. One pathway contains an intermediate--similar to experiments on protein G, and one pathway contains no intermediates-similar to experiments on protein L. The folding kinetics are characterized by tabulating mean-first passage times, and we show that the onset of glasslike kinetics occurs at much lower temperatures than the folding temperature. This model is expected to be useful in many future contexts: investigating questions of the role of local versus nonlocal interactions in various fold classes, addressing the effect of sequence mutations affecting secondary structure propensities, and providing a computationally feasible model for studying the role of solvation forces in protein folding. PMID- 11108475 TI - A simple iterative approach to parameter optimization. AB - Various bioinformatics problems require optimizing several different properties simultaneously. For example, in the protein threading problem, a scoring function combines the values for different parameters of possible sequence-to-structure alignments into a single score to allow for unambiguous optimization. In this context, an essential question is how each property should be weighted. As the native structures are known for some sequences, a partial ordering on optimal alignments to other structures, e.g., derived from structural comparisons, may be used to adjust the weights. To resolve the arising interdependence of weights and computed solutions, we propose a heuristic approach: iterating the computation of solutions (here, threading alignments) given the weights and the estimation of optimal weights of the scoring function given these solutions via systematic calibration methods. For our application (i.e., threading), this iterative approach results in structurally meaningful weights that significantly improve performance on both the training and the test data sets. In addition, the optimized parameters show significant improvements on the recognition rate for a grossly enlarged comprehensive benchmark, a modified recognition protocol as well as modified alignment types (local instead of global and profiles instead of single sequences). These results show the general validity of the optimized weights for the given threading program and the associated scoring contributions. PMID- 11108476 TI - Universal DNA tag systems: a combinatorial design scheme. AB - Custom-designed DNA arrays offer the possibility of simultaneously monitoring thousands of hybridization reactions. These arrays show great potential for many medical and scientific applications, such as polymorphism analysis and genotyping. Relatively high costs are associated with the need to specifically design and synthesize problem-specific arrays. Recently, an alternative approach was suggested that utilizes fixed, universal arrays. This approach presents an interesting design problem-the arrays should contain as many probes as possible, while minimizing experimental errors caused by cross-hybridization. We use a simple thermodynamic model to cast this design problem in a formal mathematical framework. Employing new combinatorial ideas, we derive an efficient construction for the design problem and prove that our construction is near-optimal. PMID- 11108477 TI - Early eukaryote evolution based on mitochondrial gene order breakpoints. AB - The comparison of the gene orders in a set of genomes can be used to infer their phylogenetic relationships and to reconstruct ancestral gene orders. For three genomes this is done by solving the "median problem for breakpoints"; this solution can then be incorporated into a routine for estimating optimal gene orders for all the ancestral genomes in a fixed phylogeny. For the difficult (and most prevalent) case where the genomes contain partially different sets of genes, we present a general heuristic for the median problem for induced breakpoints. A fixed-phylogeny optimization based on this is applied in a phylogenetic study of a set of completely sequenced protist mitochondrial genomes, confirming some of the recent sequence-based groupings which have been proposed and, conversely, confirming the usefulness of the breakpoint method as a phylogenetic tool even for small genomes. PMID- 11108478 TI - The NOESY jigsaw: automated protein secondary structure and main-chain assignment from sparse, unassigned NMR data. AB - High-throughput, data-directed computational protocols for Structural Genomics (or Proteomics) are required in order to evaluate the protein products of genes for structure and function at rates comparable to current gene-sequencing technology. This paper presents the JIGSAW algorithm, a novel high-throughput, automated approach to protein structure characterization with nuclear magnetic resonance (NMR). JIGSAW applies graph algorithms and probabilistic reasoning techniques, enforcing first-principles consistency rules in order to overcome a 5 10% signal-to-noise ratio. It consists of two main components: (1) graph-based secondary structure pattern identification in unassigned heteronuclear NMR data, and (2) assignment of spectral peaks by probabilistic alignment of identified secondary structure elements against the primary sequence. Deferring assignment eliminates the bottleneck faced by traditional approaches, which begin by correlating peaks among dozens of experiments. JIGSAW utilizes only four experiments, none of which requires 13C-labeled protein, thus dramatically reducing both the amount and expense of wet lab molecular biology and the total spectrometer time. Results for three test proteins demonstrate that JIGSAW correctly identifies 79-100% of alpha-helical and 46-65% of beta-sheet NOE connectivities and correctly aligns 33-100% of secondary structure elements. JIGSAW is very fast, running in minutes on a Pentium-class Linux workstation. This approach yields quick and reasonably accurate (as opposed to the traditional slow and extremely accurate) structure calculations. It could be useful for quick structural assays to speed data to the biologist early in an investigation and could in principle be applied in an automation-like fashion to a large fraction of the proteome. PMID- 11108479 TI - Tissue classification with gene expression profiles. AB - Constantly improving gene expression profiling technologies are expected to provide understanding and insight into cancer-related cellular processes. Gene expression data is also expected to significantly aid in the development of efficient cancer diagnosis and classification platforms. In this work we examine three sets of gene expression data measured across sets of tumor(s) and normal clinical samples: The first set consists of 2,000 genes, measured in 62 epithelial colon samples (Alon et al., 1999). The second consists of approximately equal to 100,000 clones, measured in 32 ovarian samples (unpublished extension of data set described in Schummer et al. (1999)). The third set consists of approximately equal to 7,100 genes, measured in 72 bone marrow and peripheral blood samples (Golub et al, 1999). We examine the use of scoring methods, measuring separation of tissue type (e.g., tumors from normals) using individual gene expression levels. These are then coupled with high dimensional classification methods to assess the classification power of complete expression profiles. We present results of performing leave-one-out cross validation (LOOCV) experiments on the three data sets, employing nearest neighbor classifier, SVM (Cortes and Vapnik, 1995), AdaBoost (Freund and Schapire, 1997) and a novel clustering-based classification technique. As tumor samples can differ from normal samples in their cell-type composition, we also perform LOOCV experiments using appropriately modified sets of genes, attempting to eliminate the resulting bias. We demonstrate success rate of at least 90% in tumor versus normal classification, using sets of selected genes, with, as well as without, cellular-contamination-related members. These results are insensitive to the exact selection mechanism, over a certain range. PMID- 11108480 TI - Systematic and fully automated identification of protein sequence patterns. AB - We present an efficient algorithm to systematically and automatically identify patterns in protein sequence families. The procedure is based on the Splash deterministic pattern discovery algorithm and on a framework to assess the statistical significance of patterns. We demonstrate its application to the fully automated discovery of patterns in 974 PROSITE families (the complete subset of PROSITE families which are defined by patterns and contain DR records). Splash generates patterns with better specificity and undiminished sensitivity, or vice versa, in 28% of the families; identical statistics were obtained in 48% of the families, worse statistics in 15%, and mixed behavior in the remaining 9%. In about 75% of the cases, Splash patterns identify sequence sites that overlap more than 50% with the corresponding PROSITE pattern. The procedure is sufficiently rapid to enable its use for daily curation of existing motif and profile databases. Third, our results show that the statistical significance of discovered patterns correlates well with their biological significance. The trypsin subfamily of serine proteases is used to illustrate this method's ability to exhaustively discover all motifs in a family that are statistically and biologically significant. Finally, we discuss applications of sequence patterns to multiple sequence alignment and the training of more sensitive score-based motif models, akin to the procedure used by PSI-BLAST. All results are available at httpl//www.research.ibm.com/spat/. PMID- 11108481 TI - Using Bayesian networks to analyze expression data. AB - DNA hybridization arrays simultaneously measure the expression level for thousands of genes. These measurements provide a "snapshot" of transcription levels within the cell. A major challenge in computational biology is to uncover, from such measurements, gene/protein interactions and key biological features of cellular systems. In this paper, we propose a new framework for discovering interactions between genes based on multiple expression measurements. This framework builds on the use of Bayesian networks for representing statistical dependencies. A Bayesian network is a graph-based model of joint multivariate probability distributions that captures properties of conditional independence between variables. Such models are attractive for their ability to describe complex stochastic processes and because they provide a clear methodology for learning from (noisy) observations. We start by showing how Bayesian networks can describe interactions between genes. We then describe a method for recovering gene interactions from microarray data using tools for learning Bayesian networks. Finally, we demonstrate this method on the S. cerevisiae cell-cycle measurements of Spellman et al. (1998). PMID- 11108482 TI - Sequencing-by-hybridization at the information-theory bound: an optimal algorithm. AB - In a recent paper (Preparata et aL, 1999) we introduced a novel probing scheme for DNA sequencing by hybridization (SBH). The new gapped-probe scheme combines natural and universal bases in a well-defined periodic pattern. It has been shown (Preparata et al, 1999) that the performance of the gapped-probe scheme (in terms of the length of a sequence that can be uniquely reconstructed using a given size library of probes) is significantly better than the standard scheme based on oligomer probes. In this paper we present and analyze a new, more powerful, sequencing algorithm for the gapped-probe scheme. We prove that the new algorithm exploits the full potential of the SBH technology with high-confidence performance that comes within a small constant factor (about 2) of the information-theory bound. Moreover, this performance is achieved while maintaining running time linear in the target sequence length. PMID- 11108483 TI - Effect of regular physical exercise on resting nasal resistance. AB - OBJECTIVE: This study was conducted to determine (1) if long-term regular training changes resting nasal resistance in humans and (2) if the changes are related to the structural component or mucosal component of nasal resistance. METHODS: We used a case-control study to compare a group of 16 athletes to 15 sedentary people of similar age. Nasal resistance was measured by computerized head-out body plethysmograph posterior rhinometry. Physical activity was evaluated by the Baecke questionnaire. RESULTS: The p values (t-test) were very significant for the Baecke sports and total scores (p < .0001) but not for the other variables: age, untreated nasal resistances, decongested nasal resistances, and Baecke work and leisure scores. There were no significant correlations between nasal resistances and indexes of physical activity in all subjects (Pearson's correlation coefficient). The subjects with extremely low and high sports and total scores were paired and studied with the Signed test and the Wilcoxon signed rank test. No significant relationship was found between the nasal resistances and the Baecke scores. CONCLUSIONS: Resting nasal resistances in a group of endurance-trained athletes are identical to those found in a group of sedentary individuals, and this relationship stands for both the structural and mucosal components of nasal resistance. A new study of the same parameters is warranted to follow a cohort of sedentary subjects as they enroll in a physical training program. PMID- 11108484 TI - Canal-wall-down tympanoplasty with mastoidectomy for advanced cholesteatoma. AB - OBJECTIVE: The objective of this study was to review 8.5 years of the senior author's experience with canal-wall-down mastoid surgery for extensive cholesteatoma with high-grade atelectasis and severely destructed ossicles. DESIGN: A retrospective review was conducted. SETTING: The setting was a tertiary care medical centre. METHODS: Available records consulted included 104 canal-wall down mastoidectomy for advanced-stage cholesteatomas between July 1984 and December 1992. MAIN OUTCOME MEASURES: Recurrence, hearing results, and dry ear rate were analyzed. RESULTS: The recurrence rate was 4 of 104 (3.8%), and 9.6% of subjects suffered from recurrent otorrhea. Thirty-seven of 104 (35.6%) achieved the closure of air-bone gap within 20 dB. The availability of stapes suprastructure influenced the postoperative hearing level significantly (p < .001). CONCLUSION: Even in treating advanced cholesteatoma, canal-wall-down mastoidectomy provides a low recurrence rate, establishes a high dry ear rate, and preserves adequate hearing when the stapes suprastructure is available for reconstruction. PMID- 11108485 TI - Screw fixation mentoplasty. AB - A technique of transoral screw fixation mentoplasty is described for management of facial disproportion due to microgenia. One hundred and fifty-two patients followed for an average of 5.3 years revealed that screw fixation mentoplasty serves well to reduce perioperative morbidity and ensure a long-term improvement in facial balance and aesthetics. PMID- 11108486 TI - Long-term follow-up of cochlear implant users with ossified cochlea. AB - OBJECTIVE: Cochlear implantation surgery in ossified cochlea is a challenge, even for the experienced otologist. Short-term assessments of auditory perception show that implantation in partial or even extensive ossified cochlea could be achieved with varying success, but no long-term follow-up results have been published yet. DESIGN AND METHODS: This paper proposes a retrospective review of eight Nucleus cochlear implant users with ossified cochlea who have been followed on a 12- to 60-month period. MAIN OUTCOME MEASURE: Auditory performances of users are reported at each control on a scale of 100 units divided into 4 skill zones of 25 units based on Erber's proposition, that is, detection, discrimination, identification, and recognition. RESULTS: Three of the eight subjects showed some progression in their auditory performances during the follow-up. The five other subjects showed no long-term progress in their auditory performance. CONCLUSIONS: Open-set comprehension could be achieved with the insertion of 9 to 10 electrodes of the Nucleus device. Auditory performance in users with ossified cochlea seems to be influenced by the same factors as in users with patent cochlea. PMID- 11108487 TI - Surgical outcome after paediatric cochlear implantation: diminution of complications with the evolution of new surgical techniques. AB - OBJECTIVE: Cochlear implantation is a commonly performed surgical procedure with specific aspects in the paediatric population. The surgical outcome associated with this procedure in a paediatric population is analyzed. METHOD: A retrospective study was performed of all children receiving a cochlear implant at The Hospital for Sick Children from 1990 to 1998. During this period, 104 patients received a cochlear implant. RESULTS: The complications encountered were classified as major (4%), minor (3%), and device failure (2%). The surgical techniques developed to decrease complications are discussed. CONCLUSION: Paediatric cochlear implantation is a safe procedure with a low incidence of complications. Nevertheless, the surgeon is now facing new challenges with cochlear implantation performed in patients with an abnormal cochlea, who carry a higher rate of complications, and cochlear reimplantation in case of device failure. PMID- 11108488 TI - Ultrasound and a new videobronchoscopic technique to measure the subglottic diameter. AB - OBJECTIVE: The aim of this study was to evaluate ultrasound (US) and a new videobronchoscopic (VB) technique in the measurement of the subglottic lumen diameter. DESIGN: This blind prospective animal study of 62 recently sacrificed rabbits was conducted at the Montreal Children's Hospital. METHODS: Three different diameter measurements of the subglottis were assessed using B-mode US on each intact animal. The same diameter measurements, as well as a fourth, were estimated with a VB measuring technique. Finally, the subglottic area was surgically exposed allowing for direct caliper measurements of all four corresponding diameters. All three US measurements on each of the 62 rabbits and all four VB measurements on each of the 60 rabbits were compared with the gold standard corresponding caliper measurements. RESULTS: Statistical analysis revealed strikingly convergent values in subglottic lumen diameter measurements using US and calipers. The mean difference between these methods for all 186 observations was 0.11 mm. With 95% confidence, the maximum discordance was less than 0.30 mm on the smallest evaluated lumen. Convergent values were also demonstrated in subglottic lumen diameter measurements using VB and calipers. The mean difference between these methods for all 240 observations was 0.16 mm. With 95% confidence, the maximum discordance was less than 0.37 mm on the smallest evaluated lumen. Pearson's correlation coefficient supported a strong and positive relationship between US and caliper measurements as well as VB and caliper measurements. Simple linear regression model indicated that the subglottic lumen diameter could be predicted by both US and our VB technique. CONCLUSION: This work represents the first attempt to validate B-mode US and a VB technique as measuring tools for the subglottic lumen diameter. Our results have clearly shown that both methods are precise measuring modalities for this purpose. With further studies, these two objective methods of measuring the subglottic diameter could be adopted universally. The precise knowledge of this diameter could help physicians recognize and describe the severity of a narrowed lumen. Reporting results in such a standardized fashion, by either US or VB, could facilitate communication among clinicians and institutions. PMID- 11108489 TI - Vestibular impairment in peripheral sensory neuropathies. AB - OBJECTIVE: Patients with peripheral sensory neuropathies frequently complain of dizziness, mainly unsteadiness, which occasionally seems to be more severe than the neuropathy. This study is an attempt to determine whether in cases where dizziness is a prominent feature the underlying cause is, at times, a concomitant vestibular dysfunction. METHOD: The files of 50 patients suffering from peripheral sensory neuropathies were retrospectively reviewed. Patients with central nervous system disease capable of inducing vestibular dysfunction were excluded from this study. In 47 of 50 patients, bithermal caloric tests were performed as part of electronystagmographic recordings. According to the caloric responses, these patients were divided into three groups: group 1: bilaterally reduced responses, group 2: unilaterally reduced responses, and group 3: normal symmetric responses. RESULTS: In 17 of 47 patients, the caloric responses were reduced bilaterally (group 1). In 8 of 47 patients, the responses were reduced unilaterally (group 2). In 22 of 47 patients, the caloric responses were normal and symmetric (group 3). CONCLUSION: Vestibular dysfunction was encountered in 25 of 47 patients (53.2%) suffering from peripheral sensory neuropathy. One wonders whether the pathologic process affecting the peripheral sensory nerves may also affect the vestibular nerve. This seems possible in view of the fact that the vestibular nerve, a sensory nerve itself, has a histologic structure similar to the peripheral sensory nerves. PMID- 11108490 TI - MultiDimensional Voice Program analysis in children with vocal cord nodules. AB - Computer-assisted voice analysis has recently been introduced as a noninvasive approach to the management of paediatric dysphonia. The aim of this study was to determine which parameters of voice analysis distinguish vocal cord nodules from normal patterns. Following diagnosis by flexible nasolaryngoscopy, 12 male children with vocal cord nodules, aged 7 to 12, underwent voice analysis by MultiDimensional Voice Program (MDVP; Kay Elemetrics, Lincoln Park, NJ, USA). These subjects were divided into two age groups (7-9 years, n = 5; 10-12 years, n = 7) and compared to age-matched controls. Results suggest that across all age groups, subjects with vocal cord nodules had statistically significant (p < .01) elevations in absolute jitter, jitter percent, relative average perturbation, pitch period perturbation quotient, smoothed amplitude perturbation quotient, and fundamental frequency variation. Further studies are required to assess the role of MDVP in the diagnosis of other voice pathologies and the monitoring of voice therapy. PMID- 11108491 TI - Unusual complication of endotracheal intubation: retropharyngeal space abscess, mediastinitis, and empyema. PMID- 11108492 TI - Squamous cell carcinoma of the thyroglossal duct cyst: report of a new case and literature review. PMID- 11108493 TI - Coil embolization of pseudoaneurysms of the external carotid artery: case series. PMID- 11108494 TI - Prostatic adenocarcinoma metastasizing to the vocal folds. PMID- 11108495 TI - Early detection of occult thyroid cancer metastases in small cervical lymph node by genetic analysis of fine-needle aspiration specimens. PMID- 11108496 TI - Medicinal leech in head and neck reconstruction. PMID- 11108497 TI - The Driving Vengeance Questionnaire (DVQ): the development of a scale to measure deviant drivers' attitudes. AB - The Driving Vengeance Questionnaire (DVQ) was developed and administered to assess drivers' use of vengeance when faced with common driving situations. Subjects in the development of the scale were 266 male and female university students. The scale was then administered to 271 university students (both male and female) and 74 male inmates who were classified as either violent or nonviolent offenders on the basis of the amount of force used in committing the offence. A Cronbach alpha of .83 (M = 40.76, n = 310) indicated a high level of internal consistency for the DVQ. Younger drivers (18-23 years old) indicated higher levels of vengeance while driving than did older drivers (24-66 years old, M = 44.35 and 37.81, respectively). Those with less driving experience (0-6 years) expressed higher levels of vengeance while driving than more experienced drivers (6+ years, M = 42.95 and 38.81, respectively). Male drivers responded with greater vengeance to the questionnaire items than females (M = 42.07 and 39.62, respectively). The level of force used in commission of crime failed to correlate with the DVQ. A factor analysis was performed with a different sample of subjects using a slightly modified version of the DVQ to deal with the issue of whether horn honking constituted an appropriate measure of aggression. When the two relevant DVQ items were changed to read "leaning on horn" rather than mere honking, a strong, single factor of vengeance was found to characterize the scale. Suggestions were made for the use of DVQ in the screening of driving license applicants and in the study of problem drivers. PMID- 11108498 TI - Predictors of dropout among men who batter: a review of studies with implications for research and practice. AB - Identifying the characteristics of men who drop out of batterers' programs is crucial for prevention, intervention, and research. This article reviews studies of program attrition to establish a description of men who fail to complete group based batterers' interventions. Studies indicate that men who drop out are more likely to be unemployed, be unmarried and/or childless, have lower incomes, and less education than men who remain. Dropouts are also more likely to have a criminal history, to report substance abuse or related problems, and to present with particular relationship concerns or orientations. The relationship between court referral and dropout was inconsistent across studies and may vary according to socioeconomic status. Psychopathology is consistently related to dropout, but may be associated with other factors (e.g., comorbidity or referral source). Age, race, childhood exposure to violence, and battering history are all inconsistently associated with dropping out. The implications of these findings for research and program development are discussed. PMID- 11108499 TI - Measuring interference with employment and education reported by women with abusive partners: preliminary data. AB - This study examines the reliability and convergent validity of the Work/School Abuse Scale (W/SAS), a measure of the ways that abusive men interfere with women's participation in education and employment. Results indicate good reliability as measured by coefficient alpha and significant correlations with both a revised version of the Conflict Tactics Scale and the Psychological Abuse Index. The W/SAS is a useful measure of the ways in which physical force and other means of interfering with women's lives isolate them from activities that might provide income, social contacts, and a sense of accomplishment. It may also be used to examine whether changes in welfare policies affect levels of physical force and nonviolent interference in women's employment and education, as suggested by the Family Violence Option to the 1996 revisions in federal welfare policies. PMID- 11108500 TI - Labeling partner violence: when do victims differentiate among acts? AB - Domestic violence professionals have debated whether all physical assaults by partners should be labeled abuse. This study examined the use of labels such as "abuse," "victim," and "battered woman" in a sample of women (n = 78) who had sustained at least one physical assault in their current or most recent relationship. Self-labeling followed a differentiating strategy, that is, women experiencing more frequent and more severe assaults were more likely to apply labels. Lower partner income, being Black, lower relationship commitment, and having ended the relationship also were associated with increased self-labeling. Labeling of hypothetical acts followed an inclusive strategy, that is, all assaults were considered abusive. These results suggest that contextual factors influence labeling. Prevention and intervention programs may be able to increase their effectiveness by including more situational context in their messages. PMID- 11108501 TI - Interpreting and defensively responding to threat: examining appraisals and coping with acquaintance sexual aggression. AB - Resistance and prevention programming aimed at strengthening women's ability to protect themselves against acquaintance sexual aggression has lacked attention to the cognitive and emotional processes women engage in when encountering such threats. Building upon current theory related to cognitive appraisal and coping processes, this study applies a theoretical model of how women evaluate and respond to sexual aggression by male acquaintances. Two hundred and two college women who had been sexually victimized by male acquaintances responded to a questionnaire that assessed their cognitive appraisals of and emotional and behavioral responses to the incident, in addition to aggression characteristics. Path analytic regression analyses examined theorized relationships among primary and secondary appraisal and emotional response variables in addition to their collective prediction of behavioral responding. The hypothesized model accounted for significant variance in behavioral responding and indicated different patterns of appraisals, emotions, and aggression characteristics predicting women's assertive and diplomatic behavioral responses to their assaults. These findings are consistent with research and theory related to individuals' appraisal of and coping with threatening events. Theoretical and intervention implications for resistance and prevention efforts are discussed. PMID- 11108502 TI - Supreme Court examines impact of errors in detecting bias during jury selection. AB - For many attorneys (and their trial consultants) jury selection is perceived as the pivotal trial within a trial that provides them an opportunity to participate in assembling a jury that will be receptive if not predisposed to their position. For prospective jurors, jury selection is also seen as a "trial" but in the very different sense that they may see it as placing their beliefs and attitudes on trial. The trial judge may also feel as if he or she is on trial, as an erroneous ruling during jury selection can be grounds for reversal on appeal regardless of the evidence that is subsequently introduced. PMID- 11108503 TI - Acute neurologic complications in children with systemic cancer. AB - Neurologic complications are common in children with cancer, but the literature dealing with this subject is sparse. Using a symptoms and signs approach, the most common causes for requesting a neurologic evaluation for this population are reviewed. The spectrum of neurologic symptoms in children with cancer differs from adults and requires the consulting neurologist to have a thorough knowledge of childhood cancer and its effects on the nervous system. PMID- 11108504 TI - Migrating partial seizures in infancy: two new cases. AB - Two infants presented at 3 weeks and 3 months of age with intractable partial seizures. Extensive investigations failed to identify an underlying cause. There was no response to antiepileptic drug therapy and no developmental progress following the onset of the seizures. In both infants there was a distinctive pattern of seizures that arose independently from multiple regions of both hemispheres. Interictal electroencephalograms revealed multifocal epileptiform activity. The infants died aged 9 and 12 months. One underwent postmortem examination, which was normal with no hippocampal sclerosis. These infants fulfill the diagnostic criteria of the syndrome of migrating partial seizures in infancy described by Coppola and colleagues in 1995. PMID- 11108505 TI - Central conduction time of magnetic brain stimulation in attention-deficit hyperactivity disorder. AB - Twenty-seven children and adolescents aged 4 to 18 years, fulfilling the Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV) criteria for attention-deficit hyperactivity disorder, were included in this study. The diagnosis was determined by a pediatric neurologist and a psychologist examined all 27 subjects with tests that included the Wechsler Intelligence Scale for Children-Revised (WISC-R) and reading, writing, handedness, personality, and anxiety scores. Subjects with histories of epilepsy, head injury, drug abuse, or psychotic disorders were excluded. Culturally based causes or emotional disorders were also excluded. Transcranial magnetic stimulation was performed on all subjects, with recording of the motor evoked potential at the biceps brachii. Central motor conduction time was calculated by cervical root stimulation. Due to shape variability, the amplitude of the motor responses was not measured in the study. The mean value of central conduction time in the subjects was very significantly increased (P < .001) compared to that in a group of normal controls, case-matched for IQ, age, and sex. A central motor conduction time greater than 12 ms indicates abnormality. A second finding in the subjects was the significant difference of central conduction time on the side-to-side stimulation (P = .03). These findings are correlated with delay in the maturation of the corticomotoneuronal system and might provide neurophysiologic data for diagnosis. PMID- 11108507 TI - Anticonvulsant blood levels: historical review with a pediatric focus. AB - Epilepsy is heterogeneous and its treatment is often complicated by variable drug responses. Buchtal et al reported a close correlation between serum phenytoin levels, electroencephalographic findings, and clinical status in 1960. They suggested that physicians adjust dosage to attain a "therapeutic level." The concept was enthusiastically received. "Therapeutic serum levels" were proposed for most anticonvulsant drugs, and by 1975, most authorities believed that pharmacokinetic factors explained individual differences in drug response. However, Froscher found that measuring levels did not improve patient outcome. More recently, Schumacher's double-blind study found no correlation between phenytoin levels and seizure control or adverse effects. Pharmacodynamic variables (differences in drug responsiveness) are more important than pharmacokinetic factors for many drugs, especially receptor-active drugs. Pharmacokinetic variables were studied first, and led to a simplistic model. They are less significant than pharmacodynamic factors in the case of warfarin anticoagulation. Anticonvulsant levels can reveal noncompliance and pharmacokinetic differences. They say nothing about pharmacodynamics. Reports of "subtherapeutic levels" imply a need to increase dosage, but this is not supported by outcome data. We still lack evidence that specific drug levels are a valid intermediate target 40 years after Buchtal's paper. Responses to some anticonvulsants could depend primarily on pharmacokinetic factors, while pharmacodynamic factors could be supreme for others. PMID- 11108506 TI - Proton magnetic resonance spectroscopic observations of epilepsia partialis continua in children. AB - We performed magnetic resonance spectroscopy in three pediatric patients (two boys and one girl, ages 11 to 17 years) with epilepsia partialis continua. Single voxel proton magnetic resonance spectroscopy was performed on each patient. Data were acquired from voxels of 4 or 8 cm3 from the affected hemisphere and from contralateral homologous regions. The spectral peaks of several metabolites (N acetyl-aspartate, choline, creatine, and lactate) were measured. Neuropathologic findings revealed Rasmussen's syndrome in two children and gliosis in one. We observed increased lactate-to-creatine ratios and reduced N-acetyl-aspartate-to creatine ratios in the affected hemispheres in all three children with epilepsia partialis continua. These data support previous reports. The largest increase in the lactate-to-creatine ratio was detected in a patient with Rasmussen's syndrome and ongoing epilepsia partialis continua at the time of measurement. The other two patients had an increase in the lactate-to-creatine ratio and a decrease in the N-acetyl-aspartate-to-creatine ratio in the affected area. The increased lactate-to-creatine ratio was associated with recurrent focal seizures from different underlying pathologies. PMID- 11108508 TI - Continuous neostigmine infusion in post-thymectomy juvenile myasthenic crisis. AB - A 10-year-old boy with myasthenia gravis had severe post-thymectomy myasthenic crisis necessitating reintroduction of mechanical ventilation on the 5th day after thymectomy. He did not respond to therapy with oral pyridostigmine, corticosteroids, or high-dose intravenous immunoglobulin. Finally, in addition to the usual supportive care, he was treated successfully with continuous intravenous infusion of neostigmine. Continuous infusion of neostigmine was used for the first time in post-thymectomy myasthenic crisis in a child to the best of our knowledge. PMID- 11108509 TI - Progression in a case of Kearns-Sayre syndrome. AB - The quantitative relationship between deleted mitochondrial DNA and the clinical course of patients with Kearns-Sayre syndrome is poorly understood. We investigated this point using tissue samples obtained at age 10 years when the patient was diagnosed as Kearns-Sayre syndrome and at age 20 years when he died of disseminated intravascular coagulation. By long polymerase chain reaction, a shortened mitochondrial genome (8.8 kb; normal, 16.6 kb) was detected in the patient. By quantitative competitive polymerase chain reaction, the percentage of deletion-carrying mitochondrial DNA was not increased as expected and did not differ significantly by tissue type or sampling time or correlate with clinical course. Although we could not demonstrate that the amounts of wild-type mitochondrial DNA decreased with accelerating progression, it was emphasized that such a reduction of mitochondrial DNA in various tissues, including those of the central nervous system, could play a significant pathogenetic role, since only wild-type mitochondrial DNA is functional in patients with large-scale deletions of mitochondrial DNA. PMID- 11108510 TI - Endocrine disorders in two sisters affected by MELAS syndrome. AB - A variety of endocrine and metabolic defects, including hypothalamopituitary hypofunction and diabetes mellitus, has been reported in association with mitochondrial disorders. We describe two sisters affected by mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS) syndrome in whom DNA analysis showed an A-->G transition at the 3243rd nucleotide position on the transfer RNALeu(UUR) gene with 65% and 45% of mutant-type mitochondrial DNA present in the blood cells of the younger and the older sister, respectively. The younger sister had severe involvement of the central nervous system with mental retardation, epilepsia partialis continua, and strokelike episodes. Endocrine investigations showed an extensive neuroendocrine dysfunction with growth hormone deficiency, hypothalamopituitary hypothyroidism, prepubertal gonadotropin levels, and absence of any secondary sexual characteristics at the age of 12 6/12 years. The neurologically normal older sister was affected by diabetes mellitus and had normal hypothalamopituitary function. Our report confirms that the endocrine system can be affected differently by the same mitochondrial DNA mutation, depending on the heteroplasmia phenomenon. A complete endocrine evaluation must be performed in patients affected by mitochondrial disease and the existence of a mitochondrial disorder should be taken into account in patients with endocrine abnormalities, even if neuromuscular signs are lacking. PMID- 11108511 TI - G8363A mutation in the mitochondrial DNA transfer ribonucleic acidLys gene: another cause of Leigh syndrome. AB - We identified a G-->A transition at nt-8363 in the mitochondrial DNA transfer ribonucleic acidLys gene in blood and muscle from a 13-month-old girl who had clinical and neuroradiologic evidence of Leigh syndrome and died at age 27 months. The mutation was less abundant in the same tissues from the patient's mother, who developed myoclonus epilepsy with ragged red fibers (MERRF) in her late 20s. In both mother and daughter, muscle histochemistry showed ragged red and cytochrome c oxidase-negative fibers and biochemical analysis showed partial defects of multiple respiratory-chain enzymes. A maternal half-sister of the proband had died at 2.5 years of age from neuropathologically proven Leigh syndrome. The G8363A mutation, which previously had been associated with cardiomyopathy and hearing loss, MERRF, and multiple lipomas, also should be included in the differential diagnosis of maternally inherited Leigh syndrome. PMID- 11108512 TI - Purple glove syndrome caused by oral administration of phenytoin. AB - A severely handicapped boy had been treated with phenytoin and his seizures were controlled well. At 10 years of age, a pharmacy gave about 1000 mg of phenytoin instead of the prescribed 100 mg of the drug per day. Several hours after the initial administration, the patient became drowsy and his hands and feet turned dark purple with marked swelling. Four days later, his mother stopped administering the phenytoin to him and took him to hospital. After fluid therapy was started, the swelling and discoloration of both his hands and feet improved gradually and disappeared 11 days after drug discontinuation. Purple glove syndrome is defined as the edema, discoloration, and pain occurring in the distal limb where intravenous phenytoin has been administered. This might be the first report of purple glove syndrome caused by the oral administration of a large quantity of phenytoin. PMID- 11108513 TI - A 15-year follow-up of a boy with pyridoxine (vitamin B6)-dependent seizures with autism, breath holding, and severe mental retardation. AB - Pyridoxine (vitamin B6) (2q31) dependency is a rare autosomal-recessive disorder that causes a severe seizure disorder of prenatal or neonatal onset. The abnormality appears to inhibit the binding of vitamin B6 to the enzyme glutamic acid decarboxylase-1, which is needed for the biosynthesis of gamma-aminobutyric acid (GABA). Most patients with pyridoxine-dependent seizures require lifelong treatment with pyridoxine. The full range of associated symptomatology is unknown since fewer than 100 cases have been reported. A majority of cases are mentally retarded. We report a 15-year-old boy with pyridoxine-dependent seizures, nonpyridoxine-dependent seizures, severe mental retardation, autistic disorder, aerophagia, breath holding, and self-injury. This complex outcome should alert clinicians to the wide range of neuropsychiatric outcomes associated with this disorder. PMID- 11108514 TI - Moyamoya syndrome after radiation therapy for optic pathway glioma: case report. AB - We present a 4-year-old girl with neurofibromatosis-1 who developed moyamoya syndrome characterized by bilateral stenosis or occlusion of the distal internal carotid arteries and their branches, leading to the development of an abnormal vascular network. In light of a literature review, the postradiation vasculopathy of the moyamoya type and its relationship with neurofibromatosis-1 are discussed. PMID- 11108515 TI - Myasthenia gravis and associated autoimmune diseases in children. AB - Myasthenia gravis has been associated with other autoimmune disorders. We report two children with myasthenia gravis and another autoimmune disease: an 18-month old boy with ocular myasthenia gravis and Hashimoto's disease and a 14-year-old girl presenting with autoimmune polymyositis, then generalized myasthenia gravis 2 years later. The rare combinations of myasthenia gravis and Hashimoto's disease or polymyositis in children are discussed, and we also briefly review myasthenia gravis and other associated autoimmune diseases in children. PMID- 11108516 TI - New-onset tic disorder following acute hemorrhage of an arteriovenous malformation. AB - The etiology of tic disorder includes idiopathic, postencephalitic, head injury, carbon monoxide poisoning, stroke, and developmental syndromes. We report a case of new-onset complex motor and vocal tics that began after hemorrhage of an arteriovenous malformation located in the left frontal lobe. We have found no reported cases of new-onset tics related to arteriovenous malformations or hemorrhage into the frontal lobes. The patient is a 16-year-old right-hand dominant boy who presented with generalized tonic-clonic seizures. Evaluation, including magnetic resonance imaging, revealed a left frontal arteriovenous malformation, confirmed by angiogram. Following resection, there was an intraparenchymal hemorrhage of the left frontal lobe with intraventricular hemorrhage, noted most prominently in the left lateral and IIIrd ventricles, and a subdural hematoma caudal to the craniotomy. The postoperative course was complicated by hemiparesis and global aphasia. During recovery, the patient developed what was thought to be a complex partial seizure evidenced by head turning to the right with vocalization and left upper extremity clonic jerks. These were brief and occurred multiple times per day. A trial of carbamazepine was given with no improvement. It was noted that the spells occurred more frequently under stress, as when the patient was frustrated with communication. The diagnosis was changed to complex motor tics and the therapy changed to clonidine. The tics subsequently improved by 80%, although they were still present. We believe the development of complex motor tics due to frontal hemorrhage represents a unique etiology and could complicate postsurgical recovery in similar cases. PMID- 11108517 TI - Growth and development in patients with untreated clefts. AB - Motivated by the observation that individuals with clefts of the lip and palate exhibit different facial growth than those individuals without facial clefts, many studies have assessed dentofacial morphology in order to understand these differing growth patterns. This review addresses current understanding of facial growth in patients with untreated clefts, drawing broad conclusions about the state of the art, and presents a challenge for cleft care in the future. PMID- 11108519 TI - Dental treatment of predental and infant patients with clefts and craniofacial anomalies. AB - Children with cleft lip and/or palate differ in facial morphology and dentition from normal noncleft children. This paper reviews the current understanding of early dento-maxillary development in children with palatal clefts and the role of presurgical orthopedic treatment prior to primary palatal surgery. Finally, shortcomings of previous approaches to studying the outcome of treatment are presented, with a challenge to reevaluate how success is measured in the new millennium. PMID- 11108521 TI - Dental and prosthodontic care for patients with cleft or craniofacial conditions. AB - Prosthodontic treatment has a long and rich history in the care of patients with cleft lip and palate. Because of increased knowledge of craniofacial growth and development and improved surgical and orthodontic treatment, today's cleft patients receive better care and in less time. This requires less prosthetic intervention. Still, prosthetics retains an important, if somewhat diminished, place in cleft care, and the prosthodontist remains an integral member of the cleft/craniofacial habilitation team. This review presents the current state of the art in dental/prosthodontic care for patients with cleft palate and related craniofacial anomalies. PMID- 11108522 TI - Secondary cleft lip nasal reconstruction: state of the art. AB - OBJECTIVE: This paper summarizes the state of the art in secondary cleft lip nasal reconstruction, distilled from the many papers written on the subject and from the author's experience with many of those procedures over the past 25 years. METHODS: The evaluation starts with the skeletal base and the need for LeFort 1 or alveolar bone grafting is discussed. The boney dorsum is next evaluated and a "monobloc" osteotomy considered. The cartilaginous dorsum follows and a "spreader-strut" graft is entertained. The tip cartilages are approached with either an open Potter or Dibbell preferred or replacement conchal graft if the tip has been destroyed by previous surgery. The skin envelope is then adjusted using methods described by Tajima, Dibbell, and Bardach. PMID- 11108523 TI - Video-imaging assessment of nasal morphology in individuals with complete unilateral cleft lip and palate. AB - OBJECTIVE: The purpose of this study was to develop a video-imaging mathematical method to assess nostril morphology. DESIGN: This retrospective study involved two age-matched groups: 28 subjects with complete unilateral cleft lip and palate (CUCLP) and 19 noncleft controls. Nose casts were reproducibly oriented in a jig such that the casts could be rotated about the coronal axis. Video images of the nostrils were captured and then analyzed for area, perimeter, centroid, principal axis, moments about the major and minor axes (I11, I22), anisometry, bulkiness, lateral offset, internostril angle, and rotational angle. RESULTS: All parameters identified nostril asymmetry in both groups. The results of the analyses using anisometry, I11, and I22 showed that, in both groups, one nostril was rounder and one was more elliptical. This asymmetry, however, differed between the two groups, and the difference was primarily based on the degree of ellipticity of the nostrils. Maximum dimension, perimeter, lateral offset, I11, and I22 were more asymmetric in the cleft group. In the control group, the right nostril was more elliptical and had a greater perimeter, and the left-side nostril had a greater bulkiness (enfolding). CONCLUSIONS: The method developed was validated for assessment of nasal morphology in cleft and noncleft samples. Nostril morphology was asymmetric in both groups but more asymmetric in the cleft group than the control group. The dominant influence of the cleft resulted in more elliptical noncleft nostrils and greater nostril shape asymmetry in the cleft group. The validated video-imaging method can now be used to assess the efficacy of treatment on nasal morphology. PMID- 11108524 TI - Comparative study of mandibular stability after sagittal split osteotomies: biocortical versus monocortical osteosynthesis. AB - OBJECTIVE: A comparative study of clinical mandibular stability following bilateral sagittal split osteotomies by means of monocortical versus bicortical osteosynthesis was undertaken. DESIGN: This retrospective study utilized cephalometric radiographs, which were taken at 1 week and 6 months postoperatively. SETTING: The research was carried out at the Department of Plastic and Reconstructive Surgery of the Nagasaki University School of Medicine. PATIENTS: A total of 32 patients who underwent only sagittal split osteotomies and mandibular set back in our unit was included in this study. Of these patients, 17 patients were osteosynthesized monocortically, and 15 patients were osteosynthesized bicortically. MAIN OUTCOME MEASURES: Four measurements--gonial angle (GA), mandibular plane (MP), SNB, and percentage of upper face to total face height (percent upper to total face)--were completed to evaluate postoperative mandibular movement. RESULT: Statistical analyses of cephalometric measurements (GA, MP, SNB, and percent upper face to total face height) showed that monocortical fixed mandibles were more changeable postoperatively on the GA and percent upper face to total face height, but MP and SNB showed no significant differences among the groups. CONCLUSION: These findings suggested that the postoperative excessive shear force stress, produced by the compressive action of the masseter muscle, transformed the mandibular shape as the distal segment rotated clockwise and proximal segment rotated counterclockwise. Consequently, the mandible was bent at the miniplate. It was concluded that bicortical osteosynthesis was more rigid against this shearing stress than monocortical osteosynthesis. PMID- 11108525 TI - Cranial base and facial skeleton asymmetries in individuals with unilateral cleft lip and palate. AB - OBJECTIVE: Individuals with unilateral cleft lip and palate (UCLP) manifest a plethora of phenotypic characteristics, including asymmetric development of the middle and lower facial skeleton. The purpose of this study was to retrospectively investigate the development of cranial base asymmetries in patients with UCLP noted on posteroanterior cephalometric radiographs. METHODS: Thirty individuals with UCLP and 64 controls participated in this study. Medial and lateral cranial base asymmetries were analyzed on frontal cephalometric radiographs relative to three developmental stages. Furthermore, the development of horizontal and vertical lower facial asymmetry in these patients with UCLP was assessed in relation to cranial base, nasomaxillary, and dentoalveolar structures. RESULTS: Individuals with UCLP demonstrated cranial base asymmetries that did not significantly differ from individuals without cleft. In addition, lower facial asymmetry in patients with UCLP correlated with horizontal lower facial and dentoalveolar asymmetries but not with cranial base or nasomaxillary structures. CONCLUSIONS: No significant vertical cranial base asymmetries were detected in patients with UCLP. Horizontal lower facial asymmetry appeared to develop in close relation to the vertical asymmetries of mandibular fossae and dentoalveolus. PMID- 11108526 TI - Quantitative 3D maxillary arch evaluation of two different infant managements for unilateral cleft lip and palate. AB - OBJECTIVE: A two-institution retrospective study was undertaken to determine whether two different prepalatoplasty protocols quantitatively affect maxillary arch morphology in infants with complete unilateral cleft lip and palate (UCLP). DESIGN: Serial maxillary dental casts, obtained at regular intervals through the first 18 months of life from preintervention until palatoplasty were evaluated quantitatively using computer-assisted three-dimensional digitization and analysis for three populations: institution 1 (protocol 1), institution 2 (protocol 2), and unaffected individuals (neither cleft nor treatment). Sequential UCLP patients from institution 1 were matched for age and initial alveolar cleft width, sex and cleft side having been demonstrated to be nonsignificant, with UCLP patients from institution 2 and to unaffected individuals for age for the analysis. SETTING: Both treatment institutions are well-established regional interdisciplinary cleft centers. Institution 1 is located in a tertiary, academic children's hospital in a metropolis within a primarily agrarian region of the Midwest; institution 2 is a freestanding private clinic located in a small city within a primarily agrarian region of an eastern state; the unaffected population is a historic archive acquired in the 1930s. Data acquisition (model digitization) and computer processing were performed at institution 1. PATIENTS: Eighty-five casts of 28 infants from institution 1, 106 casts of 31 infants from institution 2, and 68 casts of 29 unaffected infants were analyzed. All infants had alginate impressions taken prior to intervention and at several additional 6-month intervals after that, consistent with each institution's treatment protocol. INTERVENTIONS: At institution 1, patients with UCLP underwent lip adhesion and placement of a passive alveolar molding plate at 7 weeks of age, definitive cheiloplasty at 7 months of age, and one-stage palatoplasty at 14 months of age. At institution 2, patients with UCLP underwent definitive cheiloplasty at 3 months of age, had no maxillary orthopedics, and had vomer flap hard palate repair at 12 months of age and soft palate repair at 18 months of age. MAIN OUTCOME MEASURES: The outcome measures included directly digitized (cleft segment and hemialveolar ridge lengths) and derived (alveolar base width, alveolar cleft gap, maxillary frenum-alveolar base perpendicular angle, and rates of change over time of digitized cleft segment and hemialveolar ridge lengths) features. The data were assessed by comparing simple linear regression lines and an unpaired, two-tailed t test. RESULTS: Prior to initiating therapy, there were no statistically significant differences between the two populations with clefts. However, both populations with clefts differed significantly from unaffected individuals (p < .001), with increased maxillary base widths and larger perpendicular/frenum angles. At the time of palatoplasty, the two populations with clefts had statistically significant differences between them in the maxillary base width (p < .01) and the cleft gap distance (p < .05). The base width of institution 1 did not differ significantly from that of widths of unaffected children, and that of institution 2 was significantly less, although the latter had already received first-stage palate repair. Alveolar segment growth rates were similar for the greater and lesser segments, respectively, and the left side hemialveolus of both groups. The growth rate for the noncleft side hemialveolus of institution 2 exceeded (p < .05) that of both institution 1 and unaffected patients. CONCLUSION: Two different regimens for the initial management of UCLP can significantly affect maxillary alveolar arch growth with respect to the treatment used and in comparison with unaffected controls. PMID- 11108527 TI - The origin and development of the upper lateral incisor and premaxilla in normal and cleft lip/palate monkeys induced with cyclophosphamide. AB - OBJECTIVE: Cleft lip/palate (CLP) is a common human congenital defect in which the maxillary lateral incisors are often absent, malformed, and malpositioned. The present study was designed to examine the origin of the upper primary lateral incisor relative to the medial nasal process (MNP) and maxillary process (MP) fusion area and to the premaxillary/maxillary (incisive) suture in monkeys. METHOD: Scanning electron microscopy, histology, skeletal staining, and drying techniques were used to study facial development in embryo and fetal monkey specimens. A teratogenic dose of cyclophosphamide was administered to pregnant monkeys prior to fusion of the MNP and MP and fetuses were examined for CLP. RESULTS: Formation of the anterior maxilla involved fusion of the MNP and MP at stages 14-18. At stages 18-20, the palatal portion of the MNP had formed the medial and lateral incisive mounds. By stage 22, the upper primary lateral incisor has formed within the MP, lateral to the MNP/MP fusion area and to the ossifying premaxilla. Ossification of the premaxilla begins in the MNP and subsequently spreads laterally across the MNP/MP fusion area into the MP. Accordingly, the lateral incisor undergoes a complex positional shift (mainly medial) relative to the incisive suture both prenatally and postnatally and is finally located medial to the suture. Examination of the cyclophosphamide-induced CLP fetuses showed that the lateral incisor is located lateral to the alveolar cleft and does not shift medial to the incisive suture. CONCLUSION: Understanding the origin of the lateral incisor (the tooth closest to the cleft) and the shift after its formation provides clues to high incidence of malformations and ectopia of this incisor in cleft patients. PMID- 11108528 TI - The effect of vowels on nasalance scores. AB - OBJECTIVE: Nasalance scores were compared for nine different speech stimuli with vowel content controlled. DESIGN: The nine speech stimuli included four vowels spoken in isolation and five sentences. The four vowels were /i/, /u/, /ae/, and /a/. Four of the five sentences were loaded with High Front, High Back, Low Front, or Low Back vowels, and the fifth sentence contained a mixture of vowel types. SETTING: Academic and clinical craniofacial center. SUBJECTS: The subjects were 19 children with velopharyngeal dysfunction (VPD) and 19 children without history of communication disorder. MAIN OUTCOME MEASURES: The main outcome measures were the nasalance scores associated with the nine different speech stimuli for two groups of subjects. RESULTS: For the VPD group, analysis of variance procedures revealed that nasalance scores for high-vowel sentences and the mixed-vowel sentence were significantly higher than the nasalance scores for the two low-vowel sentences. This pattern was the same for the non-VPD group except for the High Back/Low Back contrast, which was not significant. In both groups, nasalance scores for sustained vowels were significantly higher for the High Front vowel /i/ than for any other vowel, and nasalance was significantly higher for the High Back vowel /u/ than for either of the Low vowels /ae/ or /a/. There was no significant difference between Low vowels. CONCLUSION: Nasalance scores may be affected by the vowel content of the speech stimulus. This should be taken into consideration on a clinical basis and for research purposes. PMID- 11108529 TI - The load-displacement characteristics of neonatal rat cranial sutures. AB - OBJECTIVE: Recently several centers have attempted to distract the craniofacial skeleton in infants with craniosynostosis. To effectively achieve this goal, we must first understand the normal sutural response to tensile forces. The objective of this study was to determine the load-displacement characteristics of neonatal rat sutures. METHODS: Thirty cranial sutures were harvested from 1-week old Wistar rats (10 each coronal, posterior frontal, and sagittal). The width of the harvested bone-suture-bone construct was standardized to 4 mm. The specimens, kept moist, were mounted fresh and distracted at 10 microm/sec until rupture using a Vitrodyne V1000 universal tester. Standard load-displacement curves were constructed. The stiffness, defined as tensile force/change in suture length, and the ultimate stress, defined as tensile force at suture rupture/cross sectional area, were calculated. RESULTS: These sutures demonstrated classical viscoelastic behavior. During the elastic phase, they elongated approximately 1 microm for every 1 g of force (10(4) N/m). The ultimate tensile stress was approximately 4 MN/m2. The estimated mean elastic modulus was 10 megapascals. The posterior frontal sutures were significantly less stiff than the other two sutures (Kruskal Wallis nonparametric analysis of variance, p = .0023). The difference in the ultimate stress was also significant (p = .0201). CONCLUSIONS: This study provides data regarding the basic mechanical behavior of neonatal cranial sutures in a mammalian system. PMID- 11108530 TI - Congenital heart disease in a patient with microform cleft lip. AB - OBJECTIVE: Microform cleft lip (MCL), also known as congenital healed cleft lip, is a rare anomaly that is thought to be a spontaneously healed variant of the more common clefting of lip and palate. Although conventional clefting is known to be associated with congenital heart disease (CHD), MCL has not been similarly associated. We present one patient with MCL and CHD in the form of coarctation of the aorta, an association not previously described. CONCLUSION: As is done in patients with cleft lip, cleft palate, or both, patients with MCL should be carefully examined for evidence of CHD and referred to a pediatric cardiologist if deemed advisable. PMID- 11108531 TI - Harvesting iliac bone graft: decreasing the morbidity. PMID- 11108532 TI - Recommendations for quality improvement in genetic testing for cystic fibrosis. European Concerted Action on Cystic Fibrosis. AB - These recommendations for quality improvement of cystic fibrosis genetic diagnostic testing provide general guidelines for the molecular genetic testing of cystic fibrosis in patients/individuals. General strategies for testing as well as guidelines for laboratory procedures, internal and external quality assurance, and for reporting the results, including the requirements of minimal services in mutation testing, the nomenclature for describing mutations, procedures to control false-positive amplification reactions and to validate tests, and guidelines to implement a quality system in a molecular diagnostic laboratory are reviewed. PMID- 11108533 TI - Validation of a highly sensitive ICP-MS method for the determination of platinum in biofluids: application to clinical pharmacokinetic studies with oxaliplatin. AB - ELOXATIN (Oxaliplatin) is a novel platinum containing anti-cancer agent with a diaminocyclohexane carrier ligand which has been approved in several major European countries. Clinical studies have demonstrated that the compound exhibits marked activity against colorectal cancers in combination with 5-fluorouracil (5 FU). The aim of this work was to develop and validate a highly sensitive inductively coupled plasma mass spectrometry assay for the determination of oxaliplatin-derived platinum in plasma ultrafiltrate, plasma and whole blood and to apply this technique to clinical pharmacokinetic studies with oxaliplatin. Ultratrace detection of platinum in plasma ultrafiltrate was achieved using ultrasonic nebulisation combined with ICP-MS. This technique allows detection of platinum at the 0.001 microg Pt/ml level in only 100 microl of matrix. Assays in blood and plasma utilised a standard Meinhardt nebuliser and spray chamber, achieving detection limits of 0.1 microg Pt/ml in 100 and 200 microl of matrix, respectively. The assays were validated (accuracy and precision within +/- 15%) over the concentration ranges: 0.001-0.250 microg Pt/ml in plasma ultrafiltrate and 0.1-10 microg Pt/ml for plasma and whole blood. The effect of sample digestion. dilution, long term frozen storage and quantitation in the presence of 5-FU were also investigated and validated. The method was used to monitor platinum exposure following oxaliplatin administration (130 mg/m2) to cancer patients. Following a 2 h i.v. infusion, peak platinum levels declined in a triphasic manner in all blood compartments. Free platinum was detected in plasma ultrafiltrate at low levels (0.001 0.010 microg Pt/ml) for up to 3 weeks. In conclusion, a highly sensitive and specific assay has been developed for the determination of platinum in biofluids. This method enabled characterisation of the long term exposure to platinum in patients following oxaliplatin treatment. PMID- 11108534 TI - Determination of mirtazapine in tablets by UV spectrophotometric and derivative spectrophotometric methods. AB - Mirtazapine, 1, 2, 3, 4, 10, 14b-hexahydro-2-methyl-pyrazino [2, 1-a] pyrido [2, 3-c] [G.L. Stimmel, J.A. Dopheide and S.M. Stahl, Pharmacotheraphy 17(1) (1997) 10] benzazepine, is a new and well tolerated antidepressant. It blocks pre synaptic alpha2-adrenergic receptors and postsynaptic serotonin type 2 and type 3 receptors. The drug is rapid and completely absorbed after oral administration. Mirtazapine was analyzed by HPLC and gas chromatography with nitrogen-sensitive detection. In this study, mirtazapine was analyzed by using UV spectrophotometry, first and second order derivative spectrophotometry. The type of solvent, the degree of derivation, range of wavelength and n value were chosen in order to optimize the conditions. The concentration of mirtazapine in its methanolic solutions were determined between the wavelength range of 225-360 nm in the linearity range of 1-100, 2-100 and 1-120 microg ml(-1) by using the values obtained from UV. first-order derivative (n = 5, delta lambda = 17.5 nm) and second-order derivative (n = 9, delta lambda = 31.5 nm) spectrum of the substance, respectively. The developed UV Spectrophotometric, first-order and second-order derivative spectrophotometric methods were applied to a pharmaceutical preparation as tablet form. Developed UV and derivative UV spectrophotometric method in this study are accurate, sensitive, precise, reproducible and can be directly and easily applied to the pharmaceutical preparations. PMID- 11108535 TI - Kinetic study of the indomethacin synthesis and thermal decomposition reactions. AB - The kinetics of indomethacin synthesis (achieved through a new method) was studied at 80 degrees C. The reaction proceeds in four steps. In the first step, by the condensation of levulinic acid with p-chlorobenzoyl-p-methoxy-phenyl hydrazine in homogeneous acidic catalysis an intermediate is formed. In the second step, by the isomerisation of this intermediate a hydrazo compound is formed. In the third step, the isomerisation is followed by the o-benzydinic transposition of the reaction product mentioned above. In the fourth step, indomethacin is formed through a cyclization reaction. The rate constant of the indomethacin synthesis reaction was determined assuming that the cyclization reaction constitutes the rate-determining step. Spectrophotometric methods were used both in order to investigate the kinetics of the synthesis reaction and to verify the proposed mechanism. Then, a thermogravimetric study was performed on the purpose of finding out the temperature range in which indomethacin is stable. From the thermogravimetric curve kinetic parameters have been derived, using different calculation techniques. PMID- 11108536 TI - Simultaneous estimation of phenylpropanolamine HCl, guaiphenesin and diphenylpyraline HCl in syrups by LC. AB - A simple, precise and accurate HPLC method was developed for the simultaneous estimation of phenyl-propanolamine HCl, guaiphenesin and diphenylpyraline HCl in syrup. The method was carried out on a Shimpak C8 column with a mobile phase consisting of acetonitrile-triethylamine (pH adjusted to 3.5 using orthophosphoric acid; 0.5%), (35:65, v/v) at a flow rate of 1.2 ml min(-1). Detection was carried out at 210 nm. Diphenhydramine was used as internal standard. The validation of the method was also carried out. PMID- 11108537 TI - Extractive spectrophotometric methods for determination of diltiazem HCl in pharmaceutical formulations using bromothymol blue, bromophenol blue and bromocresol green. AB - Three simple and sensitive extractive spectrophotometric methods have been described for the assay of diltiazem hydrochloride either in pure form or in pharmaceutical formulations. The developed methods involve formation of coloured chloroform extractable ion-pair complexes of the drug with bromothymol blue (BTB), bromophenol blue (BPB) and bromocresol green (BCG) in acidic medium. The extracted complexes showed absorbance maxima at 415 nm for all three methods. Beer's law is obeyed in the concentration ranges 2.5-20.0, 2.5-10.0 and 2.5-12.5 microg ml(-1) with BTB, BPB and BCG, respectively. The methods have been applied to the determination of drug in commercial tablets and capsules. Results of analysis were validated statistically and through recovery studies. PMID- 11108538 TI - A quantitative and qualitative high performance liquid chromatographic determination of acetaminophen and five of its para-substituted derivatives. AB - An accurate HPLC determination of acetaminophen in tablet dosage form is reported, together with an effective separation of five of its para-substituted derivatives using an isocratic reversed-phase system. The column consisted of an octadecylsilane 10 microm stationary phase. The mobile phase was a mixture of acetonitrile and water (7:3). The retention times for acetaminophen, O acetylacetaminophen, O-ethylacetaminophen, O-(2 nitrobenzenesulphonyl)acetaminophen, O-benzylacetaminophen and O-(3,5 dinitrobenzoyl)acetaminophen were 2.83, 3.25, 3.56. 3.78, 4.37 and 6.03 min, respectively. PMID- 11108539 TI - Strategies for the improvement of an amperometric cholesterol biosensor based on electropolymerization in flow systems: use of charge-transfer mediators and platinization of the electrode. AB - Different configurations based on an amperometric biosensor with cholesterol oxidase entrapped in a polypyrrole film have been developed with a view to improving the analytical properties of this biosensor. The alternatives considered involve the simultaneous entrapment of the enzyme and a charge transfer mediator as well as previous platinization of the surface of the Pt electrode. Both artificial (a ferrocene derivative) and natural (flavin nucleotides) mediators were studied as constituents of the charge-transfer process between the enzyme and the electrode. The comparative study of these biosensors, which were prepared in situ in a continuous flow system, made it possible to determine the advantages and disadvantages of each configuration when applied to flow-injection determination of cholesterol. PMID- 11108540 TI - A colorimetric field method to assess the authenticity of drugs sold as the antimalarial artesunate. AB - Artesunate is the most widely used of the artemisinin derivatives. These drugs are being used increasingly throughout the tropical world, and are an essential component of the treatment of multi-drug resistant malaria. The recent and widespread appearance of counterfeit artesunate tablets in several countries in Southeast Asia poses a serious threat to health in this region. We have developed a simple, inexpensive colorimetric test to determine artesunate authenticity in tablets. The test is based on a reaction between an alkali decomposition product of artesunate and a diazonium salt, fast red TR (FRTR). The appearance of a yellow color indicates the presence of artesunate. The specificity of the test is dependent on the pH of the reaction. Among other antimalarials tested, (i.e. artemisinin, artemether, chloroquine, quinine, primaquine, sulfadoxine, and pyrimethamine) only artesunate produced a positive color reaction at pH 4. The assay requires only 1% of the total weight of a standard tablet containing 50 mg of artesunate and can be completed within 10 min. The method was tested on six genuine artesunate tablets and six counterfeit artesunate tablets obtained in Southeast Asia. The average amount of artesunate in the genuine tablets was determined to be 50.8 +/- 2.9 mg while the counterfeit tablets were found to contain no artesunate. PMID- 11108541 TI - Identification of photodegradation products of nilvadipine using GC-MS. AB - Nilvadipine (NV) photodegradation products have been analysed with gas chromatography-mass spectrometry (GC-MS). The photodegradation was carried out in the conditions recommended in the first version of the document issued by the International Conference on Harmonization (ICH), currently in force in the studies of photochemical stability of drugs and therapeutic substances. Methanol solutions of NV were irradiated with a high-pressure UV lamp - type HBO 200. The maximum intensity at the wavelength lambda = 365 nm was achieved by applying the interference filter and Wood's filter. Using the Reinecke salt as a chemical actinometer, apparent quantum yields of photodegradation were obtained, which after extrapolation to the zero time of irradiation gave the actual quantum yield ((phi = 7.58 x 10(-5). The structure of three nilvadipine photodegradation products was established, after mass spectra analysis of compounds registered during GC-MS carried out of irradiated nilvadipine solutions. The quantitative results of GC-MS analyses enabled to determination of the kinetic parameters of NV photodegradation, calculated from the dependence In c =f(t). Quantitatively the process was described with the calculated rate constant of decomposition (k), decomposition time of 50% of the compound (t0.5) and decomposition time of 10% of the compound (t0.1). The exposure of nilvadipine to UV light was found to lead to aromatization of the DHP ring and elimination of the HCN molecule. PMID- 11108542 TI - Solid-state studies on the hemihydrate and the anhydrous forms of flunisolide. AB - Flunisolide exists in at least two different anhydrous crystalline forms (I and II) and in a hemihydrate form with distinctly different physico-chemical properties. Modification II and the hemihydrate form are the commercial products. Form I was obtained by heating all other forms above 230 degrees C. The different crystalline forms of flunisolide were investigated by FTIR spectroscopy, X-ray powder diffractometry, differential scanning calorimetry (DSC), thermogravimetric analysis and thermomicroscopy both coupled with FTIR spectroscopy (TG-FTIR and FTIR thermomicroscopy). The three forms were easily differentiated by their IR spectra, X-ray powder diffraction patterns and thermal behaviour. Their stability was investigated under different experimental conditions to verify the tendency to solid solid transition and to study the existence range of the three forms. The relationship among the two anhydrous polymorphs and the hemihydrate form and their equilibrium solubilities in water at 20 degrees C were also investigated. PMID- 11108543 TI - Comparison of antibody binding to immobilized group specific affinity ligands in high performance monolith affinity chromatography. AB - A novel biochromatographic principle is introduced taking the quantitative analysis of affinity interactions between antibodies and immobilized group specific ligands (protein A, G, and L) as example. The name high performance monolith affinity chromatography (HPMAC) is proposed for this technique. HPMAC uses rigid, macroporous monoliths, so-called convective interaction media (CIM) disks, as stationary phase. An optimized procedure is described for the covalent immobilization of the group specific affinity ligands to such disks. The binding of polyclonal bovine IgG and a recombinant human antibody (type IgGl-kappa) to all affinity disks is discussed. An essential feature of HPMAC is its compatibility to unusually high mobile phase flow rates ( > 4 ml/min). Chromatographic experiments are thus completed within seconds without significant loss in binding capacity and retentive power. This makes HPMAC a promising tool for applications in fast process monitoring or screening. As an example for the former, the direct quantitative isolation of recombinant antibodies from serum free culture supernatant is demonstrated. PMID- 11108544 TI - A sensitive method for the measurement of the novel pet endectocide, selamectin (UK-124,114), in dog and cat plasma by chemical derivatisation and high performance liquid chromatography with fluorescence detection. AB - An analytical method has been developed for the novel pet endectocide, selamectin (USAN, UK-124, 114), in dog and cat plasma to facilitate pharmacokinetic profiling for this compound. The method involves solid phase extraction of the compound and internal standard from plasma followed by chemical derivatisation using triethylamine and trifluoroacetic anhydride. This reaction yields a highly fluorescent product and thus provides a sensitive assay. Using a sample volume of 1.0 ml for dog plasma the assay has been validated over a concentration range of 0.2-40 ng/ml. Due to smaller plasma volumes for cat plasma samples, the assay was validated over a concentration range of 1.0- 200 ng/ml using a sample volume of 0.2 ml. The analyte has been shown to be stable for 48 h at room temperature and through three freeze thaw cycles in dog plasma. The analytical method is highly specific and proved suitable for the analysis of selamectin in dog and cat plasma samples following doses of compound by parenteral and non-parenteral routes. PMID- 11108545 TI - HPTLC screening assay for urinary cotinine as biomarker of environmental tobacco smoke exposure among male adolescents. AB - For selective screening determination of urinary cotinine, i.e. (S)-1-methyl-5-(3 pyridyl)-2-pyrrolidinone, the major metabolite of nicotine, the high-performance thin-layer chromatographic (HPTLC) method have been proposed. Prior the final HPTLC analysis the procedure required a solid-phase extraction (SPE) of cotinine from collected urine samples with 1-methyl-2-pyrrolidinone as an internal standard. Densitometrical quantitation of cotinine on the chromatograms have been performed with a 16-grayscale scanner using the specialized software implemented on a desktop microcomputer. The lower detection limit of cotinine was 6 microg/l allowing the method to be applied for the measurement a concentration of this compound in urine samples collected from 35 elementary schoolboys exposed on both moderate and/or significant ETS. The mean recovery of cotinine from urine samples was 93%. The mean intra-day accuracy for the analysis of cotinine in range 6-750 microg/l. including four paralell measurements, was 2.9 %. The results of cotinine measurements by proposed SPE-HPTLC procedure were used in the pilot studies for assessment of hazard from home ETS on the health status of elementary schoolboys, especially an increased risk for infectious respiratory track diseases. PMID- 11108546 TI - Inosine and 2'-deoxyinosine and their synthetic analogues: lipophilicity in the relation to their retention in reversed-phase liquid chromatography and the stability characteristics. AB - The purines and among them inosine synthetic nucleoside derivatives and analogues belong to a group of compounds to which the attention is being paid because of their biological activities. Relationships of their various parameters are being investigated because of their effect on biological (antineoplastic, virostatic, immunosuppressive) properties. Hydrophobicity parameters expressed as the logarithm of the partition coefficient (log P) and the capacity factor k' for naturally occurring inosine, 2'-deoxyinosine, 2'-deoxyadenosine and 2' deoxyguanosine and for inosine synthetic analogues 5'-deoxyinosine, 5'-chloro-5' deoxyinosine and 2',3'-dideoxyinosine were measured. The effect of methanol percentage in the mobile phase and its pH on the retention of the studied compounds in a reversed-phase system was also examined. There was a good correlation between the lipophilicity expressed as log P and capacity factor k'. It was also determined that dissociation has a marginal effect on capacity factor k' in this group of nucleoside derivatives as the k' values were almost unchanged at various pH of the mobile phase used. The stability of the all investigated compounds was investigated in basic, neutral and acidic conditions. The values of the reaction constant k1 were calculated and effects of nucleoside structural characteristic on stability are discussed. PMID- 11108547 TI - The uterotonic activity of compounds isolated from the supercritical fluid extract of Ekebergia capensis. AB - The wood of Ekebergia capensis Sparrm. is used by the local Zulu community in KwaZulu-Natal Province, South Africa to facilitate childbirth. In this investigation, the uterotonic properties of extracts from this tree were evaluated using both pregnant and non-pregnant guinea pig uterine smooth muscle in vitro. The extracts were prepared using water modified supercritical carbon dioxide at 400 atm and 80 degrees C. As samples of these extracts displayed positive results when screened for uterotonic activity, gravity column chromatography followed by NMR spectroscopy was performed in an attempt to isolate and elucidate the structures of the compounds that were present in the extract. The extract yielded five known compounds of which only two, viz. oleanonic acid and 3-epioleanolic acid, displayed uterotonic activity. Receptor binding assays were subsequently performed with 3-epioleanolic acid to ascertain its mode of action. Bradykinin (30 ng/100 microl) and acetylcholine (1 microg/100 microl) were used as the B2 and cholinergic receptor agonists respectively with icatibant (HOE 140) (30 ng/100 microl) and atropine (60 micro/100 microl) as their corresponding antagonists. 3-epioleanolic acid was observed to mediate its effect through the cholinergic receptor. The results of this study show that two compounds from the extract of this tree possess varying degrees of agonist activity on uterine smooth muscle with minor changes in the molecular structure affecting its intrinsic activity on uterine muscle. PMID- 11108548 TI - LC-MS for the identification of oxygen heterocyclic compounds in citrus essential oils. AB - The oxygen heterocyclic compounds (coumarins, psoralens and polymethoxylated flavones) present in the nonvolatile residue of the essential oils of Mandarin, Sweet Orange, Bitter Orange, Bergamot and Grapefruit were analysed with an HPLC/API/MS system equipped with an APcI probe in positive mode. The use of hyphenated techniques, such as LC/MS provides a great information about the content and nature of constituents of natural complex matrices, such as essential oils. In this work, MS spectra were recorded at different voltages, to obtain structural information in addition to molecular weight information. The different response of the compounds identified has been also evaluated. The method allowed the confirmation of the identification of the main components of the fraction, previously reported for the different oils. MS characteristics of coumarins, psoralens and polymethoxylated flavones with different substitution patterns were determined on the basis of the response obtained with the APcI interface. Interface parameters were optimised to obtain a contemporaneous response for all the three classes of components. PMID- 11108549 TI - LC analysis of benzophenone-3: II application to determination of 'in vitro' and 'in vivo' skin penetration from solvents, coarse and submicron emulsions. AB - The aim of this study was to determine the skin penetration of benzophenone-3 in vitro and in vivo in order to investigate a possible influence of formulation. Six different vehicles, three solvents and three different emulsion types were evaluated in vitro and in vivo. Each vehicle was applied to the skin model at 2 mg cm(-2). First, histological studies on ear pigskin and human skin were evaluated. In vitro measurements were performed with static diffusion cells using pigskin at 1, 2, 4, and 8-h. In vivo, benzophenone-3 concentration in stratum corneum was evaluated by the stripping method after 30-min application on forearm of volunteers. It was shown that ear pigskin and human skin appear similar and in both experiments significant differences between vehicles were noticed. The six vehicles could be ranked in the same order of benzophenone-3 skin concentration. At 8-h, the highest concentration of benzophenone-3 in skin was obtained with propylene glycol, and O/W submicron emulsion. On the contrary. the two oily solvents. W/O emulsion and O/W coarse emulsion restrain the concentration of this UV-filter in the skin. At each time, permeability in vitro and in vivo were well correlated. Low concentrations were measured in the receptor fluid suggesting that percutaneous absorption of this UV-filter across the skin would be minimal. The in vitro and in vivo skin penetration capacity of benzophenone-3 from six vehicles was confirmed and quantified. A satisfactory relationship between binary in vitro and in vivo was established. PMID- 11108550 TI - Meiosis-associated calcium waves in ascidian oocytes are correlated with the position of the male centrosome. AB - We have used confocal microscopy to measure calcium waves and examine the distribution of tubulin in oocytes of the ascidian Ciona intestinalis during meiosis. We show that the fertilisation calcium wave in these oocytes originates in the vegetal pole. The sperm penetration site and female meiotic apparatus are found at opposite poles of the oocyte at fertilisation, confirming that C. intestinalis sperm enter in the vegetal pole of the oocyte. Following fertilisation, ascidian oocytes are characterised by repetitive calcium waves. Meiosis I-associated waves originate at the vegetal pole of the oocyte, and travel towards the animal pole. In contrast, the calcium waves during meiosis II initiate at the oocyte equator, and cross the oocyte cytoplasm perpendicular to the point of emission of the polar body. Immunolocalisation of tubulin during meiosis II reveals that the male centrosome is also located between animal and vegetal poles prior to initiation of the meiosis II-associated calcium waves, suggesting that the male centrosome influences the origin of these calcium transients. Ascidians are also characterised by an increase in sensitivity to intracellular calcium release after fertilisation. We show that this is not simply an effect of oocyte activation. The data strongly suggest a role for the male centrosome in controlling the mechanism and localisation of post fertilisation intracellular calcium waves. PMID- 11108551 TI - Behaviour of mouse primary spermatocyte nuclei after fusion to enucleated metaphase II oocytes. AB - Primary spermatocytes originating from prepubertal mouse testes were electrofused to metaphase II (MII)-stage oocytes, enucleated either by the conventional micromanipulation method or by chemical treatment with etoposide and cycloheximide. These experiments were followed by assessment of morphological changes in transferred nuclei using light microscopy, by chromosomal analyses and by screening of hybrids for the presence or absence of DNA synthesis using anti bromodeoxyuridine antibody and immunofluorescence staining of the hybrids. The results show differences between the two types of ooplasts in susceptibility to activation stimuli. However, when activated, both types of ooplasts gave rise to hybrids of similar morphology. From 35.3% to 63% of activated hybrids originating from chemically or microsurgically enucleated oocytes, respectively, contained one large pronucleus in cytoplasm, 62% or 31.6% hybrids from those two groups, respectively, possessed two smaller pronuclei and a few contained three or four pronuclei. No DNA synthesis was detected in any hybrid containing one or more pronuclei. The chromosome spreads of hybrids with premature chromosome condensation (PCC) morphology (before activation) show that most of the hybrids had a diploid (2n) number of chromosomes. The nature and regularity of the cell division cycle in the hybrids are discussed. PMID- 11108552 TI - Immunohistochemical detection of calmodulin and calmodulin-dependent protein kinase II in the mouse testis. AB - We reported previously that in mouse testis calmodulin-dependent protein phosphatase (calcineurin) is localised in the nuclei of round and elongating spermatids (Cell Tissue Res. 1995; 281: 273-81). In this study, we studied the immunohistochemical localisation of calcium/calmodulin-dependent protein kinase (CaM kinase II) using antibodies against CaM kinase IIgamma from chicken gizzard and specific antibodies raised against the amino acid sequence Ileu480-Ala493 of this enzyme, and compared it with the distribution of calmodulin. Indirect immunofluorescence was most concentrated in early spermatocytes and localised in the outermost layer of seminiferous tubules where the calmodulin level was relatively low. Measurements of immuno-gold particle densities on electron micrographs revealed that CaM kinase II is transiently increased in the nucleus of zygotene spermatocytes. These observations suggest the involvement of CaM kinase II in the meiotic chromosomal pairing process. An extremely high concentration of calmodulin in spermatogenic cells undergoing meiosis may not be directly related to activation of calmodulin-dependent kinases and phosphatases. PMID- 11108553 TI - Fine-structural cytochemical and immunocytochemical observations on nuclear bodies in the bovine 2-cell embryo. AB - Nuclear bodies occurring during the 2-cell stage of bovine embryos (obtained either by in vitro fertilisation of in vitro matured ovarian oocytes, or collection after fertilisation and cleavage in vivo) were studied using ultrastructural cytochemistry and immunocytochemistry to determine whether their occurrence may be linked with the onset of embryonic transcription. In addition, the species-specific ultrastructural features of the interchromatin structures of the 2-cell bovine embryo were displayed. Three different types of nuclear bodies were distinguished: (i) nucleolus precursor bodies (NPBs), (ii) loose bodies (LBs) and (iii) dense bodies (DBs). In order to determine their possible functional significance, we considered parallels between these three nuclear entities and interchromatin compartments reported in other cells. As detected by their preferential ribonucleoprotein staining, all types of nuclear bodies contained ribonucleoproteins. In contrast to the other types of nuclear bodies studied, NPBs contained argyrophilic proteins but in no case they did show morphological features of functional nucleoli. Both compact and vacuolated forms of NPBs were seen in both in vivo and in vitro embryos, sometimes simultaneously in the same nucleus. LBs and DBs reacted with antibodies to Sm antigen, indicating the presence of a group of nucleoplasmic, non-nucleolar small nuclear ribonucleoproteins (snRNPs). The immunoreactivity for Sm antigen was more intense and homogeneous in DBs than in LBs. DBs were seen in both categories of embryo. A possible kinship of DBs with the sphere organelle known from oocytes of different animal species or the prominent spherical inclusions of the early mouse embryo nuclei is suggested. The last type of intranuclear body, the LBs, showed a composite structure. Their granular component, occurring in clusters and displaying immunoreactivity for Sm antigen, was similar to interchromatin granules and was therefore named IG-like granules. Another component forming the LBs showed a much finer structure and a lower immunoreactivity with anti-Sm antibodies. We suggest that this amorphous component may be related to the IG associated zone. All three types of intranuclear bodies were often seen close together, suggesting their possible mutual functional relationship. From these and other observations we conclude that the intranuclear bodies in 2-cell bovine embryos correspond, with the exception of the NPB, to similar structures/compartments supposed to accumulate inactive spliceosomal components in certain phases of somatic cell nucleus functions. Accordingly, the occurrence of such nuclear bodies does not represent cytological evidence for RNA synthesis. In contrast to this, an important morphological feature revealing the status of the bovine 2-cell embryo is the vacuolisation of the NPB. PMID- 11108554 TI - Behaviour and role of an intra-acrosomal antigenic molecule, acrin 3, during mouse fertilisation in vitro. AB - In this study we examined the behaviour and role of an intra-acrosomal antigenic molecule, acrin 3, during mouse fertilisation in vitro by assessing the effect of its pertinent monoclonal antibody mMC101. Experiments were designed to assess the effect of mMC101 on sperm-zona pellucida binding, the acrosome reaction, zona pellucida penetration, sperm-egg fusion, and fertilisation in vitro. mMC101 did not affect sperm motility or primary and secondary binding to the zona pellucida, but significantly inhibited fertilisation of zona-pellucida-intact oocytes in a dose-dependent manner. In the presence of mMC101 at 100 microg/ml concentration in TYH medium, none of the oocytes developed to pronuclear stage by 5 h after co incubation of the gametes, but the pronucleus formation rate recovered to some extent (45.3%) after 8 h, indicating a delay of early embryonic development. mMC101 also delayed and significantly suppressed zona pellucida penetration by sperm. Acrin 3 dispersed and did not remain on completely acrosome-reacted sperm. Although mMC101 did not influence the zona-pellucida-induced acrosome reaction, it significantly inhibited fertilisation when acrosome-reacted sperm in the presence of mMC101 inseminated zona-pellucida-free oocytes. However, fertilisation remained unaffected when acrosome-reacted sperm in the absence of mMC101 inseminated zona-pellucida-free oocytes even in its presence. Thus, acrin 3 appears to facilitate zona pellucida penetration and is also likely to be involved in sperm-oocyte fusion by modifying the sperm plasma membrane during the acrosome reaction. PMID- 11108555 TI - Energy substrates and the completion of spontaneous meiotic maturation. AB - This study was carried out to examine how different combinations of pyruvate and glucose affect spontaneous meiotic maturation of cumulus-cell-enclosed mouse oocytes (CEO) to metaphase II (MII). Most experiments used an open system in which oocytes were cultured in 1 ml medium in plastic tubes. Initial experiments examined the dose response effects of pyruvate or glucose alone in the presence or absence of 2 mM glutamine. When medium lacked both pyruvate and glucose, more than 91% of the oocytes died in glutamine-free medium during 15 h of culture; viability was restored with the addition of glutamine, but only 11% of the CEO reached MII. In the absence of glutamine, 62-68% of oocytes completed maturation in 0.23-2.3 mM pyruvate, while 44-60% MII was observed in 0.55-27.8 mM glucose. The addition of glutamine to these cultures had a general suppressive effect on the completion of maturation. When glucose was added to pyruvate-containing cultures, the combination of 1 mM pyruvate/5.5 mM glucose was most effective in supporting maturation (about 90% MII), with little effect of glutamine. No further increase in maturation was observed when glucose was increased five-fold (to 27.8 mM). The positive effect of glucose was in part attributed to stimulation of glycolysis and increased production of pyruvate, since a reduced culture volume (8 microl), which allows the accumulation of secreted pyruvate, improved maturation in glucose-containing, but not pyruvate-containing, medium, and FSH, which stimulates glycolysis, increased progression to MII in glucose containing, but not pyruvate-containing, medium. Yet these results also suggest that glucose has a beneficial effect on maturation apart from simple provision of pyruvate. The pyruvate effect was directly on the oocyte, because denuded oocytes responded more effectively than CEO to this energy substrate. The highest percentage of MII oocytes (96-97%) occurred in microdrop cultures containing glucose but lacking glutamine. These results indicate that glutamine supports oocyte viability but is not an adequate energy source for the completion of spontaneous meiotic maturation and may be detrimental. In addition, while pyruvate and glucose alone can each support meiotic progression of CEO to MII, optimal maturation requires the provision of both substrates to the culture medium when a large volume (1 ml) is used. It is concluded that careful attention to specific energy substrate supplementation and culture volume is important to optimise spontaneous meiotic maturation in vitro. PMID- 11108556 TI - Actin filament distribution in blocked and developing pig embryos. AB - Actin filaments play an important role in cell division. The present study was designed to examine the relationship between actin filament distribution and pig embryo development. When in vivo matured and fertilised pig oocytes were cultured in TCM 199 or NCSU 23, in various proportions, 45-65% of inseminated oocytes developed to the 2- to 4-cell stages but blastocyst development was observed only in NCSU 23 (34%) or NCSU 23 containing 10% TCM 199 (7%). Supplementation of NCSU 23 medium with 20% or more TCM 199 resulted in no blastocyst formation. Examination of actin filaments indicated that microfilaments were distributed in the cortex, at the junction of blastomeres and in the perinuclear area in the embryos cultured in NCSU 23, but perinuclear actin filaments were not observed in embryos cultured in TCM 199. When 2- to 4-cell stage embryos obtained from TCM 199 were transferred to NCSU 23 medium at 36 h after in vivo fertilisation, 57% of the cleaved embryos developed to blastocysts, which was no different from the proportion obtained after culture in NCSU 23 alone (56%). In addition, when 2- to 4-cell stage embryos obtained from TCM 199 were transferred to NCSU 23, most embryos showed perinuclear actin filaments within 6h. The results indicate that the composition of the culture medium plays an important role in the polymerisation of actin filaments, which in turn influences embryo development. It is possible that pig embryo development was blocked by some components in TCM 199 which prevented actin filament polymerisation. PMID- 11108558 TI - Transurethral microwave thermotherapy. PMID- 11108557 TI - Extrafollicular mediation of oocyte maturation by radial nerve factor in starfish Pisaster ochraceus. AB - In starfish ovaries follicle cells that envelop each oocyte are thought to mediate the production of a maturation inducing substance (MIS), identified as 1 methyladenine, that induces maturation and spawning of oocytes after exposure to a gonadotropic substance secreted by the radial nerve (RNF). Studies were carried out to assess the possible role of extrafollicular cells within the ovarian wall in mediating this signal transduction process in the ovary of Pisaster ochraceus. Oocyte maturation and spawning occurred following the addition of RNF to intact ovarian tissue in vitro whereas no maturation occurred following the addition of RNF to germinal vesicle (GV) oocytes or GV oocytes surrounded by follicle cells. In contrast, oocyte maturation occurred when small ovarian wall fragments, lacking mature follicles, were incubated with GV oocytes and RNF. Neither actinomycin D nor cycloheximide altered RNF induction of oocyte maturation in the presence of the ovarian wall tissue whereas preheating (boiling water for 5 min) the tissue obliterated its response to RNF. Non-ovarian tissues failed to produce MIS in response to RNF. Results suggest that ovarian components other than the follicle cells that envelop fully grown immature oocyte are responsive to RNF and represent a significant and previously unrecognised intra-ovarian source of MIS. PMID- 11108559 TI - Microwave applicators for thermotherapy of benign prostatic hyperplasia: a primer. AB - Microwave thermotherapy for treatment of benign prostatic hyperplasia (BPH) is becoming increasingly more common. This article provides an introduction to the functional principles of microwave antennas for delivery of energy to the prostatic gland. Different antenna designs (monopole, dipole, and helical coil types) and impedance matching are discussed. PMID- 11108560 TI - Aspects on transurethral microwave thermotherapy of benign prostatic hyperplasia. AB - The underlying principle behind new minimal invasive procedures, such as microwave thermotherapy, is to coagulate the prostatic adenomatous tissue by means of heat. This article describes the action of heat on tissue and identifies areas of concern during treatment. The extent of the necrosis during treatment is governed by two physical variables: the intraprostatic temperature and the duration of the heat exposure. The prostatic blood flow is a key factor for the outcome of microwave treatment because it acts as a coolant and may effectively sink the temperature in the treatment area. Blood flow can vary substantially between patients and may change significantly during treatment. By measuring the intraprostatic temperature and varying the microwave power accordingly, it is possible to compensate for the large variations in prostatic blood flow and obtain consistent treatment. PMID- 11108561 TI - Transurethral microwave thermotherapy in the armamentarium of therapeutic modalities for benign prostatic hyperplasia. AB - Transurethral microwave thermotherapy (TUMT) has gained a firm place in the spectrum of therapeutic modalities for management of patients with lower urinary tract symptoms suggestive of bladder outflow obstruction. To achieve optimum results following TUMT, intense research focuses on appropriate patient selection, heat-tissue interactions, and modification of technical specifications. Results of TUMT are good to excellent for the majority of patients, but there is a non-negligible number of patients who respond poorly. The selection of favorable candidates for TUMT aims to improve the therapeutic results, and both clinical baseline parameters and intrinsic characteristics of the prostate (histologic composition and vasculature) may influence treatment outcome. TUMT achieves therapeutic response through coagulative necrosis of the hyperplastic tissue, but additional theories have been proposed recently, suggesting that TUMT may cause neural destruction and induce apoptosis. Individualization of the treatment is expected to offer the best results, and because the temperature achieved inside the prostate determines the actual parenchymal necrosis, thermal monitoring during treatment will permit application of microwave energy in a feedback mode. Various microwave devices differ in technical specifications (operating frequency, design of antenna, cooling system), and recently introduced software programs (high-energy protocols, heat shock strategy, short-duration protocols) aim at better efficacy, providing a more patient-friendly procedure. TUMT has survived the "test of time" that other, initially promising, modalities have failed. What remains to be determined is the maximum benefit that patients and health systems can gain from such a technique. PMID- 11108562 TI - Outcome predictors of high-energy transurethral microwave thermotherapy. AB - PURPOSE: Despite the good results of high-energy transurethral microwave thermotherapy (TUMT) for treatment of benign prostatic hyperplasia (BPH), it still is difficult to predict the response to treatment on an individual basis. In addition to clinical baseline parameters, histologic parameters seem to play a role in response variance after TUMT. High-energy TUMT has become widely accepted as a minimally invasive outpatient treatment in patients with lower urinary tract symptoms and BPH. Most patients benefit substantially from targeted microwave thermotherapy; however, little is known about optimal patient selection and the most relevant outcome parameters. MATERIALS AND METHODS: We evaluated Medline based studies published between 1989 and 2000, including 900 patients suffering from lower urinary tract symptoms due to BPH who were undergoing TUMT. We evaluated outcome predictors for TUMT, such as histopathological parameters, prostate-specific antigen, and volume. RESULTS: Histologic and clinical outcome parameters were identified. Patient-to-patient differences in stromal-to epithelial ratio of prostate tissue did affect outcomes. Poor responders to TUMT seemed to have a higher vessel density and a lower epithelial/stromal ratio. Relatively more abundant epithelial cells in the prostate tissue may lead to more favorable outcomes. Use of higher energy, patients with higher grade of obstruction, younger patient age, larger prostate volume (>25 mL), and higher prostate-specific antigen levels seemed to be associated with a better response to TUMT. CONCLUSIONS: New energy protocols could help tailor treatment to the individual needs of each patient. Nomograms based on volume, age, and pressure flow parameters could assist in making clinical recommendations and identifying treatment responders; however, the total amount of energy appeared to have a high impact on the prediction of response. PMID- 11108563 TI - Transurethral microwave thermotherapy versus transurethral resection of prostate. AB - PURPOSE: Invasiveness, delayed morbidity, and the high cost of transurethral resection of the prostate (TURP) have resulted in the proposal of transurethral microwave thermotherapy (TUMT) as an alternative and less invasive treatment. METHODS: Only a few randomized studies have evaluated the functional results of TUMT vs. TURP. Despite restricted inclusion criteria, they all concluded that TUMT is as effective as TURP in relieving subjective symptoms, but that it generally leads to less improvement in objective symptoms. RESULTS: TUMT results seem to last at least 36 months; at 1-year follow-up, the rate of repeat treatment is 3.2%. The rates of morbidity in both groups are roughly in the same range. Acute urinary retention (10-13.5%) and postoperative voiding discomfort are the main occurrences after TUMT; bleeding, retrograde ejaculation (50-80%), and urethral strictures (3.1-6.6%) occur after TURP. Cost evaluation, related only to Targis and U.S. data, favors TUMT. However, it seems likely that the overall cost of TUMT procedures may decrease if TUMT becomes a widely used technique, including fair reimbursement from private insurance companies or from social security departments in European countries. TUMT is safe and effective to treat BPH. CONCLUSIONS: Nevertheless, to date, the improvement that occurs in most variables seems to be less than that after TURP. PMID- 11108564 TI - Efficacy and safety of the new high-energy 30-minute transurethral microwave thermotherapy: results of 1-year follow-up in a multicenter study. AB - PURPOSE: To assess the efficacy and durability of a new 30-minute algorithm for high-energy transurethral microwave thermotherapy (TUMT, Prostasoft 3.5) in the treatment of men with lower urinary tract symptoms (LUTS) caused by benign prostatic hyperplasia. MATERIALS AND METHODS: A total of 167 men (mean age 67 years) with bothersome LUTS were treated with the new TUMT protocol. Evaluation included assessment of the short- and long-term objective and subjective outcome measures of this treatment. RESULTS: The treatment is well tolerated. The International Prostate Symptom Score improved from a mean of 19.2 at baseline to 7.9 at 12 months after treatment. Maximum urinary flow improved from 8.9 to 16.4 mL/s at 12 months. Mean duration of catheterization was 16.1 days. Urodynamic evaluation showed a change from the obstructed to the nonminimally obstructed zone. There were no serious complications. CONCLUSION: High-energy TUMT using the new high-dose Prostasoft 3.5 protocol appears to be a safe, effective, and durable treatment. The faster procedure improves tolerance of the treatment. Subjective and objective improvements were significant and the treatment-related morbidity low. PMID- 11108565 TI - Initial and superior experience with 30 minute TUMT. AB - Transurethral microwave thermotherapy (TUMT) is an evolving technique with different machines, protocols, intraprostatic temperatures, marketing claims, and clinical outcomes that can be confusing to the clinician. We report our initial and superior results with 30 Minute TUMT over previous treatment protocols in 16 patients. Patient discomfort and acceptance are greatly improved, with reduced analgesic requirements (11 vs. 24 mL of remifentanil), visual analogue pain scores of 0-2, and no power interruption required in any patients. All four patients in urinary retention are catheter-free 1 week after therapy. Post treatment catheterization was required in only one patient who was voiding spontaneously before the procedure. Urinary flow rates and postvoid residuals improved in all patients. Prostatic cavities were found in all patients having prostate ultrasound 3 months after TUMT. 30 Minute TUMT is not simply a shortened 30-minute TUMT treatment. Rather it is a very different TUMT with an initial power of 80 W and initial urethral cooling water of 48 degrees F/8 degrees C. Mean maximum intraprostatic temperatures achieved are 154 degrees F/68 degrees C or 43 degrees F/24 degrees C greater than previous versions of microwave thermotherapy. 30 Minute TUMT s increased cooling and shorter times result in minimal discomfort and elimination of routine catheterization, but the initial 80 W energy and avoidance of power interruption provide higher intraprostatic temperatures and prostatic cavities in almost all patients in this office-based treatment. PMID- 11108566 TI - Microwave thermotherapy in patients with chronic urinary retention. AB - PURPOSE: To evaluate the efficacy of high-energy transurethral microwave thermotherapy (TUMT) in the treatment of chronic urinary retention (CUR) due to benign prostatic hyperplasia (BPH). MATERIALS AND METHODS: In this prospective cohort study, 29 patients with CUR due to BPH underwent high-energy TUMT. Prior to treatment and during a 12-week follow-up, the following parameters were determined: quality of life (QOL) score, peak flow rate (Qmax) by uroflowmetry, and postvoid residual urine (PVR). In those with treatment failure (PVR > 150 mL or urinary retention), pressure-flow studies were performed and compared to pre TUMT urodynamics. RESULTS: Of the 29 patients, 21 (72%) regained the ability to void spontaneously at 5 weeks. The actuarial median time for spontaneous voiding to be restored was 3.5 weeks (95% confidence interval [CI] 2.9-4.8 weeks). Mean QOL score at 12 weeks post-TUMT (2.2; 95% CI 1.5-2.7) was lower than that at 1 week (4.6; 95% CI 3.9-5.8) by 51% (p < 0.0005). Further, a 55% increase in mean Qmax (p < .0005) determined by uroflowmetry was observed by 12 weeks vs. 1 week after high-energy TUMT. TUMT failed in 8 patients due to a hypocontractile detrusor. CONCLUSIONS: We concluded that high-energy TUMT is a potentially useful option for patient with CUR who are not candidates for prostatectomy. PMID- 11108567 TI - Transurethral microwave thermotherapy of the prostate without intravenous sedation: results of a single United States center using both low- and high energy protocols. TJUH TUMT Study Group. AB - PURPOSE: Previous studies have indicated that high-energy transurethral microwave thermotherapy (TUMT) requires intravenous (IV) sedation and/or narcotics for patient tolerance. This study was performed to determine tolerability, patient acceptance, and efficacy of TUMT using both low- and high-energy protocols in a single United States university setting. MATERIALS AND METHODS: Between August 11, 1997 and October 28, 1999, 210 men (mean age 64.9 +/- 9.1 years) presenting with symptomatic benign prostatic hyperplasia (BPH) received treatment with a Prostatron TUMT using either the low-energy Prostasoft 2.O or high-energy Prostasoft 2.5 software. Each patient had digital rectal examination and prostate specific antigen level consistent with BPH, American Urological Association symptom score > or = 15, and Qmax <15 mL/s. Each patient received TUMT with only ibuprofen 400 mg by mouth (PO), lorazepam 1.0 mg PO, and ketorolac 30 mg intramuscularly (IM) prior to TUMT. A few patients who were concerned about limited pain threshold received oxycodone 5 mg/acetaminophen 325 mg PO. Of 210 patients treated, 12-month efficacy data were available for analysis in 80 patients. RESULTS: Forty-eight men (mean age 65 +/- 9.2 years) received low energy 2.0 software TUMT, and 32 men (mean age 65.1 +/- 9.2 years) were treated with high-energy 2.5 software. Mean prostatic volume was 44.3 +/- 23.9 mL and 60.7 +/- 26.4 mL for the 2.0 and 2.5 groups, respectively. Mean energy delivered was 108.8 +/- 50.4 kJ and 173.1 +/- 41.1 kJ for the 2.0 and 2.5 treatment groups, respectively. International Prostate Symptom Score decreased from 23 pre-TUMT to 8 post-TUMT and 21 pre-TUMT to 10 post-TUMT at 12 months in the 2.0 and 2.5 groups, respectively. Mean peak flow rate improved 31.9% from 9.1 mL/s pre-TUMT to 12.0 mL/s post-TUMT and 45.8% from 9.6 mL/s pre-TUMT to 14.0 mL/s post-TUMT at 12 months in the 2.0 and 2.5 groups, respectively. All but two patients tolerated treatment without IV sedation. One patient experienced intolerable rectal spasm, and treatment was terminated in another patient because of poorly controlled hypertension. CONCLUSIONS: Patients can be treated safely with TUMT using either low or high energy, with almost universal patient tolerance and without the need for IV sedation or narcotics, if they premedicated effectively using a PO/IM regimen. Patients experience significant relief of symptoms whether low- or high energy TUMT is used; however, high-energy TUMT improves flow rate to a greater extent than does low-energy therapy. PMID- 11108569 TI - Water-induced thermotherapy for benign prostatic hyperplasia. AB - Water-induced thermotherapy (WIT), administered by the Thermoflex System, represents a novel minimally invasive technique for the treatment of benign prostatic hyperplasia (BPH). The Thermoflex System consists of an extracorporeal heat source and a proprietary closed-loop catheter system. Water, heated to 60 degrees C, is continuously circulated through the catheter to a treatment balloon, which conducts thermal energy to targeted prostatic tissue. The combination of heat and compression reduces the heat sink effect of the circulating blood, thus enhancing the thermal energy transfer to the compressed tissue. WIT treatment is performed using only topical urethral anesthetic, in a single 45-minute session. The 2-year follow-up data from a European multicenter study consisting of 125 patients showed an improvement in peak urine flow of 87.4% (from baseline 8.7 +/- 1.9 to 16.3 +/- 9.1 mL/s) and in the International Prostate Symptom Score (IPSS) of -54.2% (from baseline 24 +/- 5 to 11 +/- 5). Patient tolerance of WIT was rated as "excellent" or "good" in 91.8% of the procedures. WIT is efficacious, simple, and inexpensive, has few side effects, and does not need special probes to monitor prostate or rectum temperature; thus, it can be used in hospitals, outpatient clinics, and doctors' offices. PMID- 11108568 TI - Periurethral transurethral microwave thermotherapy for the treatment of benign prostatic hyperplasia: an interim 1-year safety and efficacy analysis using the thermatrx TMx-2000. AB - Transurethral microwave thermotherapy (TUMT) represents an accepted minimally invasive approach to the management of patients with benign prostatic hyperplasia (BPH). The TherMatrx TMx-2000 represents a further evolution in TUMT technique. This device uses periurethral transurethral microwave thermotherapy (P-TUMT) technology to directly target the BPH tissue adjacent to the prostatic urethra by using a catheter without a urethral-cooling surface. This article provides a technical review of the device and describes the results of a randomized, controlled multicenter study of P-TUMT for the treatment of symptomatic BPH. A discussion of the physiologic effects of P-TUMT is presented and compared to conventional TUMT. A comparison of P-TUMT to contemporary TUMT series in relation to efficacy and complications is also described. This study concludes that P-TUMT using the TherMatrx TMx-2000 device represents a minimally invasive, efficacious, and well-tolerated treatment for symptomatic BPH. PMID- 11108570 TI - Is transurethral microwave thermotherapy an alternative to medical therapy for patients with benign prostatic hyperplasia? AB - Scientific evidence supports the safety and efficacy of transurethral microwave thermotherapy (TUMT) as well as medical therapy for management of patients with benign prostatic hyperplasia (BPH). TUMT is being increasingly considered as an alternative to medical management with alpha-blockers or finasteride in patients with lower urinary tract symptoms of BPH. Enduring clinical benefits have been demonstrated after a single 1-hour microwave treatment session under topical anesthesia, and associated morbidity is low. Optimal results are obtained with the delivery of high thermal doses and accurate targeting of microwave energy. Extensive evidence from randomized clinical trials supports the safety and efficacy of both microwave treatment and medical management. Randomized clinical trial data have only recently become available directly comparing these two approaches to BPH treatment. These data indicate that greater long-term improvements in symptoms, peak urinary flow rates, and quality of life are attained with microwave treatment as compared with alpha-blockade. Furthermore, the actuarial rate of treatment failure is markedly lower in patients undergoing microwave vs. alpha-blocker treatment. However, the onset of action with alpha blocker treatment is more rapid. Limitations of alpha-blockade are side effects and lack of efficacy leading to treatment failure in some patients. Maximal effects of finasteride are modest and require a period of months to be manifested, although the side effect profile and tolerability of this agent are favorable. Neoadjuvant and adjuvant alpha-blocker therapy can accelerate symptom and flow rate improvement after microwave treatment. In contrast to medical management, microwave treatment is highly versatile, allowing patients over a broad range of baseline symptom severities and prostate sizes to be treated with a high probability of success. Compared with medical management, microwave treatment also appears to possess greater versatility, allowing patients who fall within a broad range of baseline symptom severities and prostate sizes to be treated with a high probability of success. PMID- 11108571 TI - Retrovirus-mediated gene transfer of granulocyte colony-stimulating factor receptor (G-CSFR) cDNA into MDS cells and induction of their differentiation by G CSF. AB - Myelodysplastic syndromes (MDS) are clonal disorders in which the proper differentiation of hematopoietic stem cells is impaired. There is no effective treatment for this stem cell disorder at present. In an attempt to find a new strategy that promotes the differentiation of MDS blast cells, we tried retroviral transduction of granulocyte colony-stimulating factor receptor (G CSFR) into an interleukin-3-dependent MDS cell line, MDS-L, since expression of G CSFR is known to be essential for the differentiation of myeloid progenitor cells and this expression is impaired in most MDS cells. Ectopic expression of human G CSFR cDNA in MDS-L cells gave rise to granulocytic differentiation by G-CSF stimulation. G-CSF caused the transformants expressing G-CSFR to display a morphological characteristic of mature granulocytes, upregulated CD11b on the cell surface, and improved NBT reduction activity. These results demonstrate that MDS-L cells ecopically expressing G-CSFR are induced to granulocytic differentiation upon exposure to G-CSF, and shed light on the molecular mechanisms of maturation arrest in MDS cells. PMID- 11108572 TI - Interleukin-6 and interleukin-8 levels in serum and synovial fluid of patients with osteoarthritis. AB - Concentrations of interleukin (IL)-6 and IL-8 in serum and synovial fluid obtained from patients with osteoarthritis (OA) of the knee were determined by the chemiluminescence-ELISA (CL-ELISA) method, the sensitivity of which is 100 1,000 times greater than that of the conventional ELISA method. The results were compared with those obtained from patients with rheumatoid arthritis (RA) and from healthy subjects. The mean IL-6 and IL-8 levels in synovial fluid indicated higher concentrations in RA than in OA. The IL-6 and IL-8 levels in serum were significantly higher in RA and OA relative to controls. Among OA patients in whom remarkable improvement was noted in hydrarthrosis, the synovial fluid IL-6 and IL 8 levels at the initial examination were relatively higher, and were markedly decreased after treatment with sodium hyaluronate (NaHA). Among those in whom no improvement was noted in hydrarthrosis, the synovial fluid IL-6 and IL-8 levels at the time of initial examination were relatively lower, and hydrarthrosis was not significantly improved even after treatment with NaHA. In addition, there was a tendency for the synovial fluid IL-6 and IL-8 levels to decrease as HA levels increased. Evaluation of X-ray findings revealed that the IL-6 levels in synovial fluid at the initial examination in low-grade cases tended to be significantly higher than in high-grade cases. In low-grade cases, as determined by X-ray findings, there was a significant decrease in IL-6 levels in synovial fluid after treatment with NaHA. PMID- 11108573 TI - Effect of interferon-alpha on CD20 antigen expression of B-cell chronic lymphocytic leukemia. AB - Chimeric CD20 monoclonal antibody as alternative therapy in relapsed low-grade non-Hodgkin's lymphoma (NHL) has produced responses in nearly 50% of patients. Augmenting CD20 expression on tumor cells and/or inducing its expression may increase the cell kill and effectiveness of antibody therapy. Peripheral blood lymphocytes from 19 patients with B-cell chronic lymphocytic leukemia (B-CLL) were incubated in vitro in the presence of interferon-alpha (IFN-alpha) (500 U/ml and 1,000 U/ml) for 24 and 72 hours. The effect on CD20 expression was studied by flow cytometry. The differences in the percentage positivity, the mean fluorescence intensity (MFI), and the product of percentage positivity and MFI were used to assess upregulation. There was a significant upregulation of CD20 expression on B cells seen at both concentrations after 24-hour priming (p < 0.01). B-CLL cells cultured for 72 hours in the presence of IFN-alpha also showed upregulation of CD20 expression; however, the degree of upregulation was much lower than that seen at 24 hours. There was no statistically significant increase in CD20 antigen expression on normal lymphocytes following cytokine exposure. These results suggest that IFN-alpha priming may augment the effectiveness of antibody therapy by directly upregulating CD20 antigen expression in addition to its indirect action through effector cells of the host. PMID- 11108574 TI - Use of G-CSF for granulocyte transfusion therapy. AB - Patients with neutropenia, especially neutropenia following aggressive myeloablative therapy, are at high risk for developing infectious complications caused by bacteria and opportunistic fungi. Infections remain one of the leading causes of treatment failure in patients with cancer. Thus, new and innovative therapeutic strategies are needed for management of neutropenic patients with infection. Because neutrophils represent the first line of host defense, granulocyte transfusion therapy should be a logical therapeutic approach. Although such therapy has been employed sporadically for several decades, clinical benefit has been compromised by technical problems and low granulocyte yields resulting from inadequate donor stimulation. The discovery of granulocyte colony-stimulating factor (G-CSF) as a means to elevate blood neutrophil counts when administered to normal donors has rekindled interest in granulocyte transfusion therapy. Extensive experience has been gained worldwide with G-CSF in clinical practice, and adverse events have been minimal when G-CSF has been administered to patients or healthy persons in human trials. This review focuses on the use of G-CSF in granulocyte transfusion therapy, including technical considerations of granulocyte leukapheresis and storage, donor selection and stimulation, as well as treatment results and associated risks. PMID- 11108575 TI - Ex vivo expansion of human hematopoietic progenitors and cells to support high dose chemoradiation therapy: five years of clinical experience. AB - The identification of cytokines-soluble or membrane-bound regulators of hematopoietic stem and progenitor cell survival, proliferation, and differentiation - and the definition of culture conditions that enable cell and progenitor expansion, has lead to the first clinical trials using cultured cells in addition to or in place of unmanipulated cells. The use of ex vivo expanded cells can improve several aspects of autologous and allogeneic hematopoietic cell and progenitor transplantation, such as reducing or abolishing the nadir that follows high-dose chemoradiation therapy regimens, or reducing the clinical risks associated with the use of small numbers of progenitors as in cord blood transplantation and in autologous transplantation for poor mobilizers. In addition, biological questions raised by ex vivo expansion are shared by scientists and clinicians interested in gene transfer into hematopoietic stem cells. We here review the biological problems associated with ex vivo expansion: defining efficient culture conditions, considering not only scientific and biological issues but also regulatory and commercial issues, defining appropriate surrogate endpoints that predict engraftment and superior clinical efficacy to that obtained with the use of unmanipulated grafts. We also review the results of the first clinical trials that have demonstrated the feasibilty of this approach, and have shown some of its limitations; demonstration of clinical efficacy will require more preclinical and clinical work. PMID- 11108576 TI - Compensating for our load of mutations: freezing the meltdown of small populations. AB - We have investigated the reduction of fitness caused by the fixation of new deleterious mutations in small populations within the framework of Fisher's geometrical model of adaptation. In Fisher's model, a population evolves in an n dimensional character space with an adaptive optimum at the origin. The model allows us to investigate compensatory mutations, which restore fitness losses incurred by other mutations, in a context-dependent manner. We have conducted a moment analysis of the model, supplemented by the numerical results of computer simulations. The mean reduction of fitness (i.e., expected load) scaled to one is approximately n/(n+2Ne), where Ne is the effective population size. The reciprocal relationship between the load and Ne implies that the fixation of deleterious mutations is unlikely to cause extinction when there is a broad scope for compensatory mutations, except in very small populations. Furthermore, the dependence of load on n implies that pleiotropy plays a large role in determining the extinction risk of small populations. Differences and similarities between our results and those of a previous study on the effects of Ne and n are explored. That the predictions of this model are qualitatively different from studies ignoring compensatory mutations implies that we must be cautious in predicting the evolutionary fate of small populations and that additional data on the nature of mutations is of critical importance. PMID- 11108577 TI - Taxon sampling, correlated evolution, and independent contrasts. AB - Independent contrasts are widely used to incorporate phylogenetic information into studies of continuous traits, particularly analyses of evolutionary trait correlations, but the effects of taxon sampling on these analyses have received little attention. In this paper, simulations were used to investigate the effects of taxon sampling patterns and alternative branch length assignments on the statistical performance of correlation coefficients and sign tests; "full-tree" analyses based on contrasts at all nodes and "paired-comparisons" based only on contrasts of terminal taxon pairs were also compared. The simulations showed that random samples, with respect to the traits under consideration, provide statistically robust estimates of trait correlations. However, exact significance tests are highly dependent on appropriate branch length information; equal branch lengths maintain lower Type I error than alternative topological approaches, and adjusted critical values of the independent contrast correlation coefficient are provided for use with equal branch lengths. Nonrandom samples, with respect to univariate or bivariate trait distributions, introduce discrepancies between interspecific and phylogenetically structured analyses and bias estimates of underlying evolutionary correlations. Examples of nonrandom sampling processes may include community assembly processes, convergent evolution under local adaptive pressures, selection of a nonrandom sample of species from a habitat or life-history group, or investigator bias. Correlation analyses based on species pairs comparisons, while ignoring deeper relationships, entail significant loss of statistical power and as a result provide a conservative test of trait associations. Paired comparisons in which species differ by a large amount in one trait, a method introduced in comparative plant ecology, have appropriate Type I error rates and high statistical power, but do not correctly estimate the magnitude of trait correlations. Sign tests, based on full-tree or paired comparison approaches, are highly reliable across a wide range of sampling scenarios, in terms of Type I error rates, but have very low power. These results provide guidance for selecting species and applying comparative methods to optimize the performance of statistical tests of trait associations. PMID- 11108578 TI - Dynamics of speciation and diversification in a metapopulation. AB - We develop a simple framework for modeling speciation and diversification as a continuous process of accumulation of genetic (or morphological) differences accompanied by species and subpopulation extinction and/or range expansion. This framework can be used to approach a number of questions such as species-area distribution, species-range size distribution, the rate of ecological turnover, asymmetries of range division between sister species, waiting time until speciation and extinction, the relationship between the geographic range size and the probability of speciation, the relationships between subpopulation-level parameters and metapopulation-level parameters, and the effects of taxonomic level on these rates, distributions, and parameters. We illustrate some of these applications using numerical simulations. We develop approximations describing the dependence of the number of different taxonomic units, their average range size, and the rate of their turnover on the system size, the rate of fixation of genetic (or morphological) changes in local demes, and the rate of local extinction and colonization. PMID- 11108579 TI - Quantitative genetic variation in Daphnia: temporal changes in genetic architecture. AB - Nonadditive genetic variation and genetic disequilibrium are two important factors that influence the evolutionary trajectory of natural populations. We assayed quantitative genetic variation in a temporary-pond-dwelling population of Daphnia pulex over a full season to examine the role of nonadditive genetic variation and genetic disequilibrium in determining the short-term evolutionary trajectory of a cyclic parthenogen. Quantitative traits were influenced by three factors: (1) clonal selection significantly changed the population mean phenotype during the course of the growing season; (2) sexual reproduction and recombination led to significant changes in life-history trait means and the levels of expressed genetic variation, implying the presence of substantial nonadditive genetic variation and genetic disequilibrium; and (3) Egg-bank effects were found to be an important component of the realized year-to-year change. Additionally, we examined the impact of genetic disequilibria induced by clonal selection on the genetic (co)variance structure with a common principal components model. Clonal selection caused significant changes in the (co)variance structure that were eliminated by a single bout of random mating, suggesting that a build-up of disequilibria was the primary source of changes in the (co)variance structure. The results of this study highlight the complexity of natural selection operating on populations that undergo alternating phases of sexual and asexual reproduction. PMID- 11108580 TI - Exploring the genetic basis and proximate causes of female fertility advantage in gynodioecious Thymus vulgaris. AB - In many gynodioecous species, females produce more viable seeds than hermaphrodites. Knowledge of the relative contribution of inbreeding depression in hermaphrodites and maternal sex effects to the female fertility advantage and the genetic basis of variation in female fertility advantage is central to our understanding of the evolution of gender specialization. In this study we examine the relative contribution of inbreeding and maternal sex to the female fertility advantage in gynodioecious Thymus vulgaris and quantify whether there is genetically based variation in female fertility advantage for plants from four populations. Following controlled self and outcross (sib, within-population, and between-population) pollination, females had a more than twofold fertility advantage (based on the number of germinating seeds per fruit), regardless of the population of origin and the type of pollination. Inbreeding depression on viable seed production by hermaphrodites occurred in two populations, where inbreeding had been previously detected. Biparental inbreeding depression on viable seed production occurred in three of four populations for females, but in only one population for hermaphrodites. Whereas the maternal sex effect may consistently enhance female fertility advantage, inbreeding effects may be limited to particular population contexts where inbreeding may occur. A significant family x maternal sex interaction effect on viable seed production was observed, illustrating that the extent of female fertility advantage varies significantly among families. This result is due to greater variation in hermaphrodite (relative to female) seed fertility between families. Despite this genetic variation in female fertility advantage and the highly female biased sex ratios in populations of T. vulgaris, gynodioecy is a stable polymorphism, suggesting that strong genetic and/or ecological constraints influence the stability of this polymorphism. PMID- 11108581 TI - Evolution and biogeography of the woody Hawaiian violets (Viola, Violaceae): arctic origins, herbaceous ancestry and bird dispersal. AB - Specialists studying the genus Viola have consistently allied the Hawaiian violets comprising section Nosphinium--most of which are subshrubs or treelets- with putatively primitive subshrubs in certain South American violet groups. Hawaiian violets also possess inflorescences, a floral disposition otherwise found only in other genera of the Violaceae, thus strengthening the hypothesis of a very ancient origin for the Hawaiian species. A survey of phylogenetic relationships among infrageneric groups of Viola worldwide using nuclear rDNA internal transcribed spacer (ITS) sequences revealed a dramatically different biogeographic origin for the Hawaiian violets: A monophyletic Hawaiian clade was placed in a close sister relationship with the amphi-Beringian tundra violet, V. langsdorffii s. 1., in a highly derived position. This remarkable and unforeseen relationship received strong clade support values across analyses, and monophyly of the Hawaiian lineage was further indicated by a unique 26-base-pair deletion in section Nosphinium. The high polyploid base chromosome number (n approximately equal to 40) in the Hawaiian violets relates them to Alaskan and eastern Siberian populations in the polyploid V. langsdorffii complex. More than 50 species of the 260 allochthonous birds wintering in the Hawaiian Islands are found to breed in the Arctic, occupying habitats in which individual birds might have encountered ancestral V. langsdorffii populations and served as dispersers to the central Pacific region. Acquisition of derived morphological traits (e.g., arborescence and inflorescences), significance of a confirmed Arctic origin for a component of the Hawaiian flora, and the likelihood of other "cryptic" Arctic elements in the Hawaiian flora deserving independent molecular phylogenetic corroboration are discussed. PMID- 11108582 TI - Evolution of floral display in Eichhornia paniculata (Pontederiaceae): direct and correlated responses to selection on flower size and number. AB - Trade-offs between flower size and number seem likely to influence the evolution of floral display and are an important assumption of several theoretical models. We assessed floral trade-offs by imposing two generations of selection on flower size and number in a greenhouse population of bee-pollinated Eichhornia paniculata. We established a control line and two replicate selection lines of 100 plants each for large flowers (S+), small flowers (S-), and many flowers per inflorescence (N+). We compared realized heritabilities and genetic correlations with estimates based on restricted-maximum-likelihood (REML) analysis of pedigrees. Responses to selection confirmed REML heritability estimates (flower size, h2 = 0.48; daily flower number, h2 = 0.10; total flower number, h2 = 0.23). Differences in nectar, pollen, and ovule production between S+ and S- lines supported an overall divergence in investment per flower. Both realized and REML estimates of the genetic correlation between daily and total flower number were r = 1.0. However, correlated responses to selection were inconsistent in their support of a trade-off. In both S- lines, correlated increases in flower number indicated a genetic correlation of r = -0.6 between flower size and number. In contrast, correlated responses in N+ and S+ lines were not significant, although flower size decreased in one N+ line. In addition, REML estimates of genetic correlations between flower size and number were positive, and did not differ from zero when variation in leaf area and age at first flowering were taken into account. These results likely reflect the combined effects of variation in genes controlling the resources available for flowering and genes with opposing effects on flower size and number. Our results suggest that the short-term evolution of floral display is not necessarily constrained by trade-offs between flower size and number, as is often assumed. PMID- 11108583 TI - Competition for pollination influences selection on floral traits of Ipomopsis aggregata. AB - Although rarely tested, it is often assumed that interspecific competition results in the divergence of traits related to resource use. Using a plant pollinator system as a model, I tested the prediction the presence of a competitor for pollination influences the strength and/or direction of pollinator mediated selection on floral traits. I measured phenotypic selection via female fitness on five floral traits of Ipomopsis aggregata in seven populations. Four contained only conspecifics (I only) and three also contained the competitor Castilleja linariaefolia (C + I). Directional selection via fruits/plant and conspecific pollen deposited/flower on corolla length was positive and significantly stronger in C + I populations. This difference in selection was apparently driven by interpopulation variation in the degree to which reproduction of I. aggregata was pollen limited. Consistent with expectations of interspecific competition, I. aggregata plants in C + I populations received less conspecific pollen per flower and set fewer seeds per fruit and fruits per plant than those in I only populations. Ipomopsis aggregata's corollas were also significantly longer in C + I populations, suggesting that there had been a response to a similar selective regime in past generations. Phenotypic correlations between corolla length and width, which determine the variation in I. aggregata's flower shape, were significantly weaker in C + I populations. These data suggest that competition for pollination can influence the strength of selection on and patterns of correlations among floral traits of I. aggregata. If I. aggregata populations with and without competitors for pollination are linked by gene flow, then measuring selection in competitive and noncompetitive environments maybe necessary to accurately predict how floral traits will evolve. PMID- 11108584 TI - Pollen discounting and the evolution of selfing in Arenaria uniflora (caryophyllaceae). AB - Although most models of mating system evolution assign a central role to the male transmission advantage of selfing genotypes, empirical data on the male fitness consequences of increased self-pollination are still uncommon. Here, I use measures of pollen import and export by focal plants in genotyped arrays to investigate the effects of floral morphology and pollination environment on self and outcross male function. Plants from an autogamous population of Arenaria uniflora (Caryophyllaceae) exhibit complete pollen discounting relative to closely related outcrossers, as do morphologically intermediate F1 hybrids between the two populations. However, the low cumulative male fitness of hybrids probably results from reduced pollen number or competitive ability, rather than a nonlinear relationship with floral morphology. When surrounded by selfers, plants from the outcrosser population self-fertilize at nearly the same rate as selfers (>80%), but have much lower self male fitness due to reduced fruit set. Because outcross siring success is also extremely low (<8%) in this treatment, these mate limited outcrossers are at male fitness disadvantage to both pseudocleistogamous selfers and nonlimited outcrossers. The relative male fitness of plants with different mating systems appears dependent on the ecological context, as well as on morphological trade-offs. PMID- 11108585 TI - Phylogeographic patterns and high levels of chloroplast DNA diversity in four Packera (asteraceae) species in southwestern Alberta. AB - Chloroplast DNA (cpDNA) haplotype variation is compared among alpine and prairie/montane species of Packera from a region in southwestern Alberta that straddles the boundary of Pleistocene glaciation. The phylogeny of the 15 haplotypes identified reveals the presence of two groups: one generally found in coastal and northern species and the other from species in drier habitats. The presence of both groups in all four species and most populations from southwestern Alberta is evidence of past hybridization involving species or lineages that may no longer be present in the region. With the exception of the alpine P. subnuda (phiST = 1.0), interpopulational subdivision of haplotype variation is low (phiST < 0.350), suggesting that interpopulational gene flow is high. However, based on haplotype distribution patterns, we propose that Pleistocene hybridization and incomplete lineage sorting have resulted in reduced subdivision of interpopulational variation so that gene flow may not be as high as indicated. Drift has been more important in the alpine species populations, especially P. subnuda. PMID- 11108586 TI - Evaluation of major genetic loci contributing to inbreeding depression for survival and early growth in a selfed family of Pinus taeda. AB - The magnitude of fitness effects at genetic loci causing inbreeding depression at various life stages has been an important question in plant evolution. We used genetic mapping in a selfed family of loblolly pine (Pinus taeda L.) to gain insights on inbreeding depression for early growth and viability. Two quantitative trait loci (QTLs) were identified that explain much of the phenotypic variation in height growth through age 3 and may account for more than 13% inbreeding depression in this family. One of these QTLs maps to the location of cad-nl, a lignin biosynthesis mutation. Both QTLs show evidence of overdominance, although evidence for true versus pseudo-overdominance is inconclusive. Evidence of directional dominance for height growth was noted throughout the genome, suggesting that additional loci may contribute to inbreeding depression. A chlorophyll-deficiency mutation, spf did not appear to be associated with growth effects, but had significant effects on survival through age 3. Previously identified embryonic viability loci had little or no overall effect on germination, survival, or growth. Our results challenge, at least in part, the prevailing hypothesis that inbreeding depression for growth is due to alleles of small effect. However, our data support predictions that loci affecting inbreeding depression are largely stage specific. PMID- 11108587 TI - Genotypic diversity and gene flow in brooding and spawning corals along the Great Barrier Reef, Australia. AB - Marine organisms exhibit great variation in reproductive modes, larval types, and other life-history traits that may have major evolutionary consequences. We measured local and regional patterns of genetic variation in corals along Australia's Great Barrier Reef to determine the relative contributions of sexual and asexual reproduction to recruitment and to infer levels of gene flow both locally (among adjacent sites, < 5 km apart) and regionally (among reefs separated by 500-1,200 km). We selected five common brooding species (Acropora cuneata, A. palifera, Pocillopora damicornis, Seriatopora hystrix, and Stylophora pistillata) and four broadcast spawners (Acropora hyacinthus, A. cytherea, A. millepora, and A. valida), which encompassed a wide range of larval types and potential dispersal capabilities. We found substantial genotypic diversity at local scales in six of the nine species (four brooders, two spawners). For these six, each local population displayed approximately the levels of multilocus genotypic diversity (Go) expected for outcrossed sexual reproduction (mean values of Go:Ge ranged from 0.85 to 1.02), although consistent single-locus heterozygous deficits indicate that inbreeding occurs at the scale of whole reefs. The remaining three species, the brooder S. hystrix and the spawners A. valida and A. millepora displayed significantly less multilocus genotypic diversity (Go) than was expected for outcrossed sexual reproduction (Ge) within each of several sites. Acropora valida and A. millepora showed evidence of extensive localized asexual replication: (1) a small number of multilocus (clonal) genotypes were numerically dominant within some sites (Go:Ge values were as low as 0.17 and 0.20): (2) single-locus genotype frequencies were characterized by both excesses and deficits of heterozygotes (cf. Hardy-Weinberg expectations), and (3) significant linkage disequilibria occurred. For the brooding S. hystrix Go:Ge values were also low within each of four sites (x = 0.48). However, this result most likely reflects the highly restricted dispersal of gametes or larvae, because levels of genetic variation among sites within reefs were extremely high (FSR = 0.28). For all species, we detected considerable genetic subdivision among sites within each reef (high FSR-values), and we infer that larval dispersal is surprisingly limited (i.e., Nem among sites ranging from 0.6 to 3.3 migrants per generation), even in species that have relatively long planktonic durations. Nevertheless, our estimates of allelic variation among reefs (FRT) also imply that for all four broadcast spawning species and three of the brooders, larval dispersal is sufficient to maintain moderate to high levels of gene flow along the entire Great Barrier Reef (i.e., Nem among reefs ranged from 5 to 31). In contrast, widespread populations of S. hystrix and S. pistilata (the two remaining brooders) are relatively weakly connected (Nem among reefs was 1.4 and 2.5, respectively). We conclude that most recruitment by corals is very local, particularly in brooders, but that enough propagules are widely dispersed to ensure that both broadcast spawning and brooding species form vast effectively panmictic populations on the Great Barrier Reef. PMID- 11108588 TI - Isolation by distance in equilibrium and nonequilibrium populations of four talitrid species in the Mediterranean Sea. AB - Allozymic variation at 21-23 loci was studied in 28 populations of Talitrus saltator, 23 populations of Orchestia montagui, 13 populations of O. stephenseni, and five populations of Platorchestia platensis from the Mediterranean Basin. Different levels of gene flow (Nmtheta) were detected within each species at the scale of the whole Mediterranean: O. montagui and P. platensis had low population structure, with levels of Nmtheta > or = 1, whereas the T. saltator and 0. stephenseni populations have values of Nmtheta < 1. The relationship between Nmtheta and geographic distance was analyzed to test for the presence of an isolation by distance pattern in the spatial genetic variation within each species. A model of isolation by distance is useful to describe the pattern of genetic structuring of study species at the scale of the whole Mediterranean: geographic distance explained from 28% to 70% of the variation in gene flow. In the Aegean area all species showed an island model of genetic structuring regardless of the levels of gene flow. PMID- 11108589 TI - Quantitative genetics of sexual plasticity: the environmental threshold model and genotype-by-environment interaction for phallus development in the snail Bulinus truncatus. AB - Sexual polymorphisms are model systems for analyzing the evolution of reproductive strategies. However, their plasticity and other binary traits have rarely been studied, with respect to environmental variables. A possible reason is that, although threshold models offer an adequate quantitative genetics framework for binary traits in a single environment, analyzing their plasticity requires more refined empirical and theoretical approaches. The statistical framework proposed here, based on the environmental threshold model (ETM), should partially fill this gap. This methodology is applied to an empirical dataset on a plastic sexual polymorphism, aphally, in the snail Bulinus truncatus. Aphally is characterized by the co-occurrence of regular hermaphrodites (euphallics) together with hermaphrodites deprived of the male copulatory organ (aphallics). Reaction norms were determined for 40 inbred lines, distributed at three temperatures, in a first experiment. A second experiment allowed us to rule out maternal effects. We confirmed the existence of high broad-sense heritabilities as well as a positive effect of high temperatures on aphally. However a significant genotype-by-environment interaction was detected for the first time, suggesting that sexual plasticity itself can respond to selection. A nested series of four ETM-like models was developed for estimating genetical effects on both mean aphally rate and plasticity. These models were tested using a maximum likelihood procedure and fitted to aphally data. Although no perfect fit of models to data was observed, the refined versions of ETM models conveniently reduce the analysis of complex reaction norms of binary traits into standard quantitative genetics parameters, such as genetic values and environmental variances. PMID- 11108590 TI - Reproductive isolation between divergent races of pea aphids on two hosts. II. Selection against migrants and hybrids in the parental environments. AB - Sympatric races of pea aphids on alfalfa and red clover are highly ecologically specialized and significantly reproductively isolated. Much of the restriction of gene flow between the specialized populations is due to habitat choice behavior of the winged colonizers (Via 1999). Here, we document additional pre- and postmating reproductive isolation through selection against migrants and hybrids in the parental environments. First, a group of randomly chosen genotypes from each race that were experimentally migrated between hosts had very low survival and reproduction on the alternate host relative to genotypes originating from that host (natives). Such selection against cross-host migrants forms a premating barrier to gene flow because it is likely to reduce migrant frequencies before the sexual forms are induced in the fall. Our reciprocal transplant experiment also shows that natural selection acts directly on individual migrants between the crops to favor host choice behavior: genotypes from each host suffered large losses of fitness when forced to migrate to the alternate host plant relative to the fitness they would have enjoyed had they been able to choose their native host. In a companion field study, sequential sampling throughout the summer in newly colonized fields of both alfalfa and clover revealed a decrease in the frequency of host-specific marker alleles characteristic of the alternate crop. These field data further support the hypothesis that selection disfavors migrants that cross between crops. Second, when two sets of F1 hybrids between the races were reciprocally tested on alfalfa and clover, both sets had significantly lower average fitness than the specialized parent in each of the two environments. This demographic selection against hybrids in the parental environments is a source of postmating reproductive isolation between the specialized races. Finally, significant genetic variation in fitness traits was seen among F1 hybrid genotypes from both crosses between alfalfa and clover specialists. Although this variation suggests that a generalized pea aphid could evolve, such generalists are not seen in field collections of these populations. PMID- 11108591 TI - Different mechanisms underlie phenotypic plasticity and interspecific variation for a reproductive character in drosophilids (Insecta: Diptera). AB - The insect ovary is a modular structure, the functional unit of which is the ovariole. Ovariole number is positively correlated with potential reproductive output. Among drosophilids (Insecta: Diptera), ovariole number shows both phenotypic plasticity and substantial interspecific and interpopulational variation. Here we examine the mechanistic connection between phenotypic plasticity and genetically fixed variation in ovariole number within the melanogaster species group. When a laboratory population of Drosophila melanogaster was reared under reduced food conditions, differences in ovariole number were entirely due to alterations in cell differentiation during the wandering stage at the very end of larval development. Cell growth and cell death were not affected. When these same flies were reared under a variety of temperatures, ovariole number differences arose during the latter half of the third (final) larval instar. Cell differentiation was affected, although cell number was not, and ovariole number differences were established before metamorphosis. In contrast, genetically fixed, interspecific and interpopulational variability in ovariole number was caused by alterations in the dynamics of cell differentiation and by cell number differences. Furthermore, the stages affected were different in different species and populations in the melanogaster species group, ranging from the first (D. sechellia) through the middle of the third (D. simulans and D. mauritiana) larval stage. Therefore, the mechanistic bases for plasticity-based variability are largely distinct from the mechanistic bases for interspecific and interpopulational variability. Our results suggest that phenotypic plasticity indicates evolutionary flexibility in underlying ontogenetic processes. PMID- 11108592 TI - Factors affecting the genetic load in Drosophila: synergistic epistasis and correlations among fitness components. AB - Two factors that can affect genetic load, synergistic epistasis and sexual selection, were investigated in Drosophila melanogaster. A set of five chromosomal regions containing visible recessive mutations were put together in all combinations to create a full set of 32 homozygous lines fixed for different numbers of known mutations. Two measures of fitness were made for each line: productivity (a combined measure of fecundity and egg-to-adult survivorship) and competitive male mating success. Productivity, but not male mating success, showed a pattern of strong average synergistic epistasis, such that the log fitness declined nonlinearly with increasing numbers of mutations. Synergistic epistasis is known to reduce the mutation load. Both fitness components show some positive and some negative interactions between specific sets of mutations. Furthermore, alleles with deleterious effects on productivity tend to also diminish male mating success. Given that male mating success can affect relative fitness without changing the mean productivity of a population, these additional effects would lead to lower frequencies and lower fixation rates of deleterious alleles without higher costs to the mean fitness of the population. PMID- 11108593 TI - Expression of cytoplasmic incompatibility in Drosophila simulans and its impact on infection frequencies and distribution of Wolbachia pipientis. AB - The aim of this study is to examine the expression of cytoplasmic incompatibility and investigate the distribution and population frequencies of Wolbachia pipientis strains in Drosophila simulans. Nucleotide sequence data from 16S rDNA and a Wolbachia surface protein coding sequence and cytoplasmic incompatibility assays identify four distinct Wolbachia strains: wHa, wRi, wMa, and wAu. The levels of cytoplasmic incompatibility between six lines carrying these strains of bacteria and three control lines without bacteria are characterized. Flies infected with wHa and wRi are bidirectionally incompatible, and males that carry either strain can only successfully produce normal numbers of offspring with females carrying the same bacterial strain. Males infected with wAu do not express incompatibility. Males infected with the wMa strain express intermediate incompatibility when mated to females with no bacteria and no incompatibility with females with any other Wolbachia strain. We conduct polymerase chain reaction/restriction fragment length polymorphism assays to distinguish the strain of Wolbachia and the mitochondrial haplotype to survey populations for each type and associations between them. Drosophila simulans is known to have three major mitochondrial haplotypes (siI, sill, and siIII) and two subtypes (siIIA and siIIB). All infected lines of the sil haplotype carry wHa, wNo, or both; wMa and wNo are closely related and it is not clear whether they are distinct strains or variants of the same strain. Infected lines with the silIA haplotype harbor wRi and the siIIB haplotype carries wAu. The wMa infection is found in siIII haplotype lines. The phenotypic expression of cytoplasmic incompatibility and its relation to between-population differences in frequencies of Wolbachia infection are discussed. PMID- 11108594 TI - A long-term study on seasonal changes of gametic disequilibrium between allozymes and inversions in Drosophila subobscura. AB - Seasonal variation (spring, early summer, late summer, and autumn) of gametic disequilibrium between gene arrangements (OST and O3+4) of the O chromosome and Lap, Pept-1, and Acph allozyme loci, located inside these inversions, has been recorded in a natural population of Drosophila subobscura during seven years over a 15-year period. The length of the study allowed us to investigate the temporal variation of the allozyme-inversion associations by statistical methods of time series analysis. Cyclic seasonal changes of allozyme-inversion associations for both Lap and Pept-1 are detected in the natural population. In both cases, the patterns of seasonal change are due to the seasonal change of frequency of Lap and Pept-1 allozymes occurring exclusively within the OST gene arrangement. In contrast, the allozyme frequencies at these loci within the O3+4 gene arrangement are stable along seasons. The patterns of temporal variation of allozyme inversion associations for Lap and Pept-1 in the natural population are contrasted with those previously published that correspond to gene arrangements of the O chromosome and nucleotide polymorphism at the rp49 region located inside these inversions, suggesting that natural selection is operating on these allozyme-inversion associations. PMID- 11108595 TI - Cryptic female choice in the yellow dung fly Scathophaga stercoraria (L.). AB - Both female choice and male-male competition may take place during reproduction in many species. Female choice tends to be less obvious than male-male competition and consequently has received less attention from researchers. The opportunity for cryptic female choice arises after multiple insemination. Through postcopulatory processes, a female could alter the pattern of paternity among her offspring so that it does not directly reflect the different contributions of sperm made by her mates. To be able to determine if a female alters the relative sperm contributions of her mates, the behaviors and influences of the males must therefore be first taken into account. The interest of each male is to father all the offspring, and the interest of each female is to maximize paternal quality. Female yellow dung flies have complex internal reproductive tracts that may give them considerable control over the fertilization success of stored sperm from different males. In laboratory trials to date, the last male to mate has usually been most successful. In the present study, cryptic choice occurred in Scathophaga stercoraria and the pattern of choice was consistent with previously reported results. The fertilization success of a female's second mate (P2) was substantially larger if a female was kept at constant temperature and if the second male was genetically similar to her at the phosphoglucomutase (Pgm) locus. Females from the field normally have three spermathecae, but some have four. Lines were bred to have either three or four spermathecae. Flies from the different lines were crossed to generate females with similar genetic backgrounds that had either three or four spermathecae. P2 was significantly lower for high quality females, that is, those that laid larger-than-average-clutches, with four spermathecae than for low-quality females with four spermathecae; female quality had no influence on P2 for females with three spermathecae. The results suggest that only large females may benefit from increased spermathecae number by being able to act against male interests. Females may only have three spermathecae, even though genetic variation for more is present, because selection for more spermathecae is weak. PMID- 11108596 TI - The origins of premating reproductive isolation: testing hypotheses in the grasshopper Chorthippus parallelus. AB - There are many proposed routes for the origin of premating reproductive isolation, but few systematic studies aimed at testing their relative importance. Accumulated information about the biogeographical history of the European meadow grasshopper, Chorthippus parallelus, has allowed us to make a planned series of comparisons among populations aimed at distinguishing the contributions of some of these hypotheses. We have compared the effects on assortative mating of long term isolation in glacial refugia, founder events during postglacial colonization, and sympatry with a closely related species. A likelihood-based analysis allowed us to separate effects of variation in male and female mating propensity among populations from variation in mate choice leading to assortative mating. All three effects contributed significantly to the overall variation in mating pattern in a set of 21 pairwise comparisons among seven populations. Male cuticular composition, but not other candidate signals, was significantly associated with the level of assortative mating. Of the hypotheses for the origin of reproductive isolation, only the predictions of the founder hypothesis explained a significant amount of the variation in assortative mating. This does not rule out the possiblity that there may be some other explanation. Having established the pattern of divergence, it is possible to generate hypotheses that explain our results at least as well as the founder hypothesis. However, because many such post hoc hypotheses are possible, they cannot be tested with this dataset. On this basis, our results favor the hypothesis that some aspect of the colonization process tends to accelerate divergence in mating signals leading to premating reproductive isolation. This could be accomplished through any one of several mechanisms. Colonization involves many bottlenecks as new populations are established at the edge of the range by long-distance migrants. Genetic effects may be important, but these bottlenecks may also alter the conditions under which mates are found and chosen, as suggested by Kaneshiro. At the same time, the colonizing populations may encounter novel environmental challenges. PMID- 11108597 TI - Phylogeography of the jumping spider Habronattus pugillis (araneae: salticidae): recent vicariance of sky island populations? AB - In island systems with diverging populations, the history of island formation and genealogical estimates of divergence dates can be mutually informative. In the "sky islands" of southeastern Arizona, climate-induced contraction of woodlands appears to have fragmented populations of woodland-dwelling species onto disjunct mountain ranges. Montane populations of the jumping spider, Habronattus pugillis, display striking amounts of phenotypic divergence among ranges. Paleoclimatic estimates date woodland fragmentation at approximately 10,000 years ago, suggesting that phenotypic divergence has been extraordinarily rapid in these spiders. This phylogeographic study of populations of H. pugillis attempts to clarify the species' history of isolation and divergence and to address the suitability of available paleoclimatic data for dating divergences among populations of the region's woodland-dwelling organisms. Mitochondrial sequence data of spiders from 13 mountain ranges was used to reconstruct genealogical relationships. Gene trees show that small mountain ranges tend to have populations whose sequences form monophyletic groups, whereas larger ranges do not. Paraphyly among genes from larger ranges could result from either recent migration or incomplete lineage sorting. I use phylogenetic and geographic information to test these alternatives, and conclude that incomplete lineage sorting best explains the observed paraphyly. Gene trees are concordant with some of the predictions of vegetation history generated by examination of topography. Dates estimated for divergence of populations vary from 30,000 years to more than 2 million years ago, suggesting multiple vicariance events that are older than would be inferred from paleoclimatic studies. These findings illustrate that use of any single paleontological dataset to calibrate molecular clocks can potentially greatly underestimate actual divergence times. PMID- 11108598 TI - Carotenoid limitation and mate preference evolution: a test of the indicator hypothesis in guppies (Poecilia reticulata). AB - Under the indicator models of mate choice, female preferences evolve to exploit the condition-dependence or "indicator value" of male traits, which in turn may cause these traits to evolve to elaborate extremes. If the indicator value of a male trait changes, the payoff function of the female preference for that trait should change and the preference should evolve to a new optimum. I tested this prediction in the guppy, Poecilia reticulata, a species in which the indicator value of a sexually selected male trait, carotenoid coloration, varies geographically. Carotenoid coloration is thought to be an indicator of foraging ability and health because animals must obtain carotenoid pigments from their diet. The primary dietary source of carotenoids for guppies is unicellular algae, the abundance of which varies among natural streams because of variation in forest canopy cover. Carotenoid availability limits male coloration to a greater extent in streams with greater forest canopy cover. Thus, the indicator value of male coloration covaries positively with canopy cover. To test the indicator model prediction, I measured genetic divergence in the strength of female preferences for carotenoid coloration between high- and low-carotenoid availability streams in each of three river drainages. Second-generation laboratory-born females were given a choice between full-sib males raised on three different dietary levels of carotenoids. For all six populations, male attractiveness (as determined from the responses of females to male courtship displays) increased with dietary carotenoid levels. However, the strength of female preferences differed between populations in the predicted direction in only one of three river drainages. These results fail to support a crucial prediction of the indicator model. More studies taking an interpopulation approach to studying mate preference evolution are needed before the explanatory value of the indicator models can be rigorously assessed. PMID- 11108599 TI - Divergence with gene flow in the rock-dwelling cichlids of Lake Malawi. AB - Within the past two million years, more than 450 species of haplochromine cichlids have diverged from a single common ancestor in Lake Malawi. Several factors have been implicated in the diversification of this monophyletic clade, including changes in lake level and low levels of gene flow across limited geographic scales. The objectives of this study were to determine the effect of recent lake-level fluctuations on patterns of allelic diversity in the genus Metriaclima, to describe the patterns of population structure within this genus, and to identify barriers to migration. This was accomplished through an analysis of allele frequencies at four microsatellite loci. Twelve populations spanning four species within Metriaclima were surveyed. The effect of lake-level fluctuations can be seen in the reduced genetic diversity of the most recently colonized sites; however, genetic diversity is not depressed at the species level. Low levels of population structure exist among populations, yet some gene flow persists across long stretches of inhospitable habitat. No general barrier to migration was identified. The results of this study are interpreted with respect to several speciation models. Divergence via population bottlenecks is unlikely due to the large allelic diversity observed within each species. Genetic drift and microallopatric divergence are also rejected because some gene flow does occur between adjacent populations. However, the reduced levels of gene flow between populations does suggest that minor changes in the selective environment could cause the divergence of populations. PMID- 11108600 TI - Character displacement in polyphenic tadpoles. AB - Biologists have long known that closely related species are often phenotypically different where they occur together, but are indistinguishable where they occur alone. The causes of such character displacement are controversial, however. We used polyphenic spadefoot toad tadpoles (Spea bombifrons and S. multiplicata) to test the hypothesis that character displacement evolves to minimize competition for food. We also sought to evaluate the role of phenotypic plasticity in the mediation of competitive interactions between these species. Depending on their diet, individuals of both species develop into either a small-headed omnivore morph, which feeds mostly on detritus, or a large-headed carnivore morph, which specializes on shrimp. Laboratory experiments and surveys of natural ponds revealed that the two species were more dissimilar in their tendency to produce carnivores when they occurred together than when they occurred alone. This divergence in carnivore production was expressed as both character displacement (where S. multiplicata's propensity to produce carnivores was lower in sympatry than in allopatry) and as phenotypic plasticity (where S. multiplicata facultatively enhanced carnivore production in S. bombifrons, and S. bombifrons facultatively suppressed carnivore production in S. multiplicata). In separate experiments, we established that S. bombifrons (the species for which carnivore production was enhanced) was the superior competitor for shrimp. Conversely, S. multiplicata (the species for which carnivore production was suppressed and omnivore production enhanced) was the superior competitor for detritus. These results therefore demonstrate that selection to minimize competition for food can cause character displacement. They also suggest that both character displacement and phenotypic plasticity may mediate competitive interactions between species. PMID- 11108601 TI - Differential performance among LDH-B genotypes in Rana lessonae tadpoles. AB - The European pool frog, Rana lessonae, is widely polymorphic for two common alleles (b,e) at the lactate dehydrogenase-B (LDH-B) locus. We compared fitness related larval life-history traits among LDH-B genotypes, which originated from segregation in heterozygous parents, in an artificial pond experiment where tadpoles of R. lessonae from a Swiss population were raised together with tadpoles of the hemiclonal hybrid R. esculenta at two densities. In R. lessonae, LDH-B e/e homozygotes at each density had a higher proportion of metamorphs among survivors, reached metamorphosis earlier, and were heavier at metamorphosis than b/b homozygotes; b/e heterozygotes had intermediate values. That e/e individuals were superior to b/b in both time to and mass at metamorphosis is surprising because these two life-history traits are thought to reflect a performance trade off; e/e genotypes apparently compensated for shorter time to metamorphosis by a higher growth rate. The two alleles showed the same performance ranking when combined in hybrids with a R. ridibunda allele: When R. esculenta from Swiss populations reared in the same ponds had received the e allele rather than the b allele from their R. lessonae parent, they reached metamorphosis earlier, but did not differ in mass at metamorphosis. The degree of linkage disequilibrium in the source population of the eight R. lessonae used as parents of the R. lessonae tadpoles is unknown, so we cannot exclude the possibility that the performance differences are caused by some anonymous tightly linked gene, rather than the LDH B locus, that constitutes the genomically localized target of natural selection. A causal involvement of LDH-B is plausible, nevertheless, because this enzyme takes part in the central energy-metabolizing processes and has been reported to underlie fitness differences in other animals; also, differential performance of LDH-B genotypes has been observed in R. lessonae larvae from another population. The present results suggest strong directional selection for allele e; the sum of available data, including an independent laboratory experiment, suggests that partial environment-dependent overdominance combined with balancing selection favoring e/e homozygotes under some and b/b homozygotes under other conditions may be partially responsible for the broad maintenance of the LDH-B polymorphism in R. lessonae. PMID- 11108602 TI - Experimental evidence for the adaptive evolution of growth rate in the garter snake Thamnophis elegans. AB - The western terrestrial garter snake (Thamnophis elegans) varies significantly in individual growth rates and life-history traits (maturation, reproduction, and survival) among adjacent populations in nature. This study focuses on assessing the genetic and environmental components of the substantial among-population variation in growth rates. Litters of neonates from nine populations inhabiting either mountain meadow or lakeshore habitat were reared for one year in a common garden experiment with two temperature treatments. Diet, frequency of feeding, light exposure, and daytime temperatures were identical for all individuals. The two different nighttime temperatures (20 degrees C and 25 degrees C) were chosen to mirror field differences in nighttime thermoregulatory constraints for mountain-meadow and lakeshore snakes, respectively. Temperature and source habitat interacted to affect first-year growth rate. Neonates from meadow dams grew fastest in the cooler treatment, whereas those from lakeshore dams grew fastest in the warmer treatment. The observation that naive neonates, which were gestated and raised under identical conditions, grew fastest in environments characteristic of their natal population is evidence both that there are genetic differences among populations for growth and that these differences reflect adaptation to local habitats at a very small geographic scale. In addition, significant directional selection for large birthweight was measured for neonates from all populations. These results are considered in the context of population colonization history, migration and selection, and competing models for growth rate variation. PMID- 11108603 TI - An experimental test of the thermoregulatory hypothesis for the evolution of endothermy. AB - The thermoregulatory hypothesis proposes that endothermy in mammals and birds evolved as a thermoregulatory mechanism per se and that natural selection operated directly to increase body temperature and thermal stability through increments in resting metabolic rate. We experimentally tested this hypothesis by measuring the thermoregulatory consequences of increased metabolic rate in resting lizards (Varanus exanthematicus). A large metabolic increment was induced by feeding the animals and consequent changes in metabolic rate and body temperature were monitored. Although metabolic rate tripled at 32 degrees C and quadrupled at 35 degrees C, body temperature rose only about 0.5 degrees C. The rate of decline of body temperature in a colder environment did not decrease as metabolic rate increased. Thus, increasing the visceral metabolic rate of this ectothermic lizard established neither consequential endothermy nor homeothermy. These results are inconsistent with a thermoregulatory explanation for the evolution of endothermy. PMID- 11108604 TI - Bounded hybrid superiority in an avian hybrid zone: effects of mate, diet, and habitat choice. AB - There has been considerable debate in the study of hybrid zones as to whether hybrids may be superior to parental types within the area of contact (bounded hybrid superiority). In birds, naturally occurring hybridization is relatively common, and hybridization within this group always involves mate choice. If hybrids are superior, females choosing heterospecific mates should be expected to show higher fitness under the conditions prevalent in the hybrid zone. Hybrid superiority under these circumstances would reduce reinforcement and thereby help to maintain the hybrid zone. To examine this issue, we studied reproductive performances of hybrids and parental species of gulls (Larus occidentalis and Larus glaucescens) at two colonies within a linear hybrid zone along the west coast of the United States. This hybrid zone contains predominantly gulls of intermediate phenotype. Previous studies indicated that hybrids were superior to one or both parental types, but provided no data on possible mechanisms that underlie this hybrid superiority. Using a hybrid index designed specifically for these species, we identified to phenotype more than 300 individuals associated with nests, including both individual males and females within 73 pairs in the central portion of the hybrid zone and 74 pairs in the northern portion of the hybrid zone. There was little evidence of assortative mating, and what little there was resulted solely because of pairings within intergrades. In the central hybrid zone, females paired with hybrid males produced larger clutches and hatched and fledged more chicks compared with females paired to western gull males. This was a result of heavy predation on eggs in sand habitat, where male western gulls established territories. In contrast, many hybrid males established territories in vegetated cover that was less vulnerable to predation. In the northern part of the hybrid zone, clutch size did not differ among pair categories, however, there were differences in hatching and fledging success, with females paired to hybrid males showing better success compared to females paired to glaucous-winged gull males. Hybrids showed better hatching and fledging success in the north because hybrids are more like western gulls than glaucous winged gulls in foraging behavior, taking a higher percentage of fish in their diet, which enhances chick growth and survival. This is believed to be the first documentation of bounded hybrid superiority that delineates the mechanisms that underlie hybrid superiority. PMID- 11108605 TI - The evolution of sexual dimorphism in the house finch. I. Population divergence in morphological covariance structure. AB - Patterns of genetic variation and covariation strongly affect the rate and direction of evolutionary change by limiting the amount and form of genetic variation available to natural selection. We studied evolution of morphological variance-covariance structure among seven populations of house finches (Carpodacus mexicanus) with a known phylogenetic history. We examined the relationship between within- and among-population covariance structure and, in particular, tested the concordance between hierarchical changes in morphological variance-covariance structure and phylogenetic history of this species. We found that among-population morphological divergence in either males or females did not follow the within-population covariance patterns. Hierarchical patterns of similarity in morphological covariance matrices were not congruent with a priori defined historical pattern of population divergence. Both of these results point to the lack of proportionality in morphological covariance structure of finch populations, suggesting that random drift alone is unlikely to account for observed divergence. Furthermore, drift alone cannot explain the sex differences in within- and among-population covariance patterns or sex-specific patterns of evolution of covariance structure. Our results suggest that extensive among population variation in sexual dimorphism in morphological covariance structure was produced by population differences in local selection pressures acting on each sex. PMID- 11108606 TI - Evolution at the mouse t complex: why is the t haplotype preserved as an integral unit? AB - Segregation distorters are selfish genetic elements that bias Mendelian segregation in their favor. All well-known segregation distortion systems consist of one or more "distorter" loci that act upon a "responder" locus. At the t complex of the house mouse, segregation distortion is brought about by the harmful effect of t alleles at a number of distorter loci on the wild-type variant of the responder locus. The responder and distorter alleles are closely linked by a number of inversions, thus forming a coherent t haplotype. It has been conjectured that the close integration of the various components into a "complete" t haplotype has been crucial for the evolutionary success of these selfish genetic elements. By means of a population genetical metapopulation model, we show that this intuition may be unfounded. In fact, under most circumstances an "insensitive" t haplotype retaining only the responder did invade and reach a high frequency, despite the fact that this haplotype has a strong segregation disadvantage. For certain population structures, the complete t haplotype was even competitively excluded by partial t haplotypes with lower segregation ratios. Moreover, t haplotypes carrying one or more recessive lethals only prevailed over their nonlethal counterparts if the product of local population size and migration rate (Nm) was not much smaller or larger than one. These phenomena occurred for rather realistic fitness, segregation, and recombination values. It is therefore quite puzzling that partial t haplotypes are absent from natural house mousepopulations, and that t haplotypes carrying recessive lethals prevail over nonlethal t haplotypes. PMID- 11108607 TI - The D' measure of overall gametic disequilibrium between pairs of multiallelic loci. AB - The D' coefficient is one of the most commonly used measures of the extent of gametic disequilibrium between multiallelic loci. It has been suggested that the range of the D' measure of overall disequilibrium between pairs of multiallelic loci depends on allele frequencies, except under some very restricted conditions. Nevertheless, the problem of dependence of the range of D' has not been characterized under a wide set of possible polymorphisms. Evaluation of the utility of D' as a measure of the strength of overall disequilibrium between all possible pairs of alleles at two multiallelic loci requires better knowledge of its range than is currently available. In this work, the conditions of polymorphism under which the range of D' is frequency independent are given. It is found that the range of D' is more often independent of allelic frequencies than is commonly thought. Furthermore, the range of D' undergoes only small fluctuations as a function of the polymorphisms at the loci. Numerical cases and microsatellite data from humans are used for illustration. These observations indicate that the D' coefficient is a useful tool for the estimation and comparison of the extent of overall disequilibrium across pairs of multiallelic loci. PMID- 11108608 TI - Population and subspecific genetic differentiation in the foxtail pine (Pinus balfouriana). AB - We performed an allozyme survey of genetic differentiation in Pinus balfouriana, a subalpine conifer endemic to California that is comprised of two allopatric subspecies, one in the Klamath Mountains and the other in the southern Sierra Nevada. Although the two subspecies are morphologically distinct and gene flow between them is virtually nonexistent, we observed much higher levels of differentiation among populations within a subspecies than between the two subspecies. Differentiation is particularly strong in the Klamath populations (multilocus FST = 0.242), which are small, isolated, and ecologically marginal. We attribute this strong differentiation to the mountain island effect, in which populations restricted to high elevations become isolated from each other on different mountains separated by unsuitable intervening habitat, with consequent reduced gene flow allowing populations to evolve independently. Populations of P. balfouriana in the Klamath region only exist scattered on the few highest ridges and peaks that rise above 2,000 m, which defines the lower limit of the species elevational distribution. This pattern of distribution has allowed genetic drift and allelic sorting through historical events to produce strong population-level differentiation, which was likely in place before the two subspecies were geographically separated. Because P. balfouriana occurs on both serpentine soils and nonserpentine soils in the Klamath Mountains, we tested for genetic differentiation between populations growing on serpentine versus nonserpentine soils and our results were equivocal. Our data, combined with several other studies of conifers, show that the mountain island effect can produce significant genetic differentiation in conifers whose life-history traits of widely dispersed pollen, long generation times, and high outcrossing rates would lead us to predict a more homogenous population genetic structure. PMID- 11108609 TI - Starvation resistance and adult body composition in a latitudinal cline of Drosophila melanogaster. AB - Latitudinal geographic variation in Drosophila melanogaster is pervasive. Parallel clines in traits such as body size, egg size, ovariole number, and development time have been found on several continents throughout the world. However, a cline in starvation resistance and fat content in D. melanogaster has so far been found only in India. Here we investigate starvation resistance and fat content in 10 populations from South America, in which clines in body size, egg size, and development time have previously been found. We find no evidence for a cline in starvation resistance or fat content in South America. We therefore suggest that the cline in starvation resistance in India may have evolved in response to specific climatic variation found only in India. PMID- 11108610 TI - Evaluation of selection on cliff swallows. AB - Estimates of the intensity of selection based on measurements of the living and the dead require knowledge of the fraction of the original population dying. We apply recently developed methods (Blanckenhorn et al. 1999) to estimate the intensity of selection in a population of cliff swallows. In this population the fraction of individuals dying was unknown, but certainly high. The inferred selection is very strong and impossible to achieve if the original population is assumed to have followed a normal distribution. We consider several alternative explanations for this result including measurement biases, undetected immigration, and sampling biases. Of these, sampling biases are perhaps the most likely. We conclude that the intensity of selection on the swallows was probably strong, but its absolute magnitude is unknown. PMID- 11108611 TI - Androgen receptors in cranial nerve motor nuclei of male and female rats. AB - This study compared AR proteins in four cranial nerve motor nuclei among male and female rats that were intact, gonadectomized, or gonadectomized and given TP by immunohistochemistry. AR-immunoreactive (ir) neurons were found, in descending order of abundance, in the nucleus ambiguus, hypoglossal nucleus, and the facial and trigeminal motor nuclei of both males and females of intact and gonadectomized plus TP rats. Virtually every neuron of the nucleus ambiguus was AR-ir. In contrast, AR-ir neurons were either restricted to a specific area of the hypoglossal nucleus, or randomly distributed in the facial and trigeminal motor nuclei. The predominant AR-ir site shifted from cell nuclei to the cytoplasm, depending upon the presence or absence of ligand. Sex differences in the amount and staining intensity of AR-ir neurons were discernable in all four motor nuclei of intact rats, and these differences were maintained in gonadectomized plus TP rats, with the exception of the nucleus ambiguus. The immunostaining results were complemented by results from AR binding studies. Cytosolic AR binding values for the hypoglossal and facial motor nuclei of females were only approximately 50% of those of males despite the absence of a sex difference in neuron number. These results indicate that intrinsic sex differences in AR levels and androgenic regulation of AR exist in cranial nerve motor nuclei, and that there are differences in the abundance and distribution pattern of AR responsive neurons in cranial nerve motor nuclei. These results are consistent with the idea that sex differences in AR could account for sex differences observed in nerve regeneration and neuron loss following cranial nerve injury. PMID- 11108613 TI - Contribution of neurons to habituation to mechanical stimulation in Caenorhabditis elegans. AB - In Caenorhabditis elegans, a light touch induces a locomotor response. Repeated touches, however, result in an attenuation of response, that is, habituation. Withdrawal responses elicited by anterior touch are controlled by anterior mechanosensory neurons (AVM and ALMs), and by four pairs of interneurons (AVA, AVB, AVD, and PVC) (Chalfie et al., 1985; White et al., 1986). To identify the neurons that participate in habituation, we ablated these neurons with a laser microbeam and investigated the resulting habituation of the operated animals. The animals lacking both left and right homologues AVDLR were habituated more rapidly than intact animals. We propose that chemical synapses at AVD play a critical role in the habituation of intact animals. PMID- 11108612 TI - Retinoic acid-induced changes in polysialyltransferase mRNA expression and NCAM polysialylation in human neuroblastoma cells. AB - Polysialic acid (PSA) is a dynamically regulated carbohydrate modification of the neural cell adhesion molecule NCAM, which is implicated in neural differentiation and cellular plasticity. The cloning and characterization of two polysialyltransferases, termed ST8SiaII (STX) and ST8SiaIV (PST), opened up new perspectives in the search for factors that control this unique cell surface glycosylation. In vitro and transfection approaches revealed that ST8SiaII and ST8SiaIV are independently capable of synthesizing PSA on NCAM with slightly different specificities towards the major NCAM isoforms and glycosylation sites. Their overlapping but distinct expression patterns during brain development point towards an independent transcriptional regulation. However, the factors driving their joint or distinct expression, as well as the significance of divergent expression patterns in vivo, are not yet understood. In the present study, the mRNA expression of ST8SiaII and ST8SiaIV was comparatively analyzed in neuronal differentiation of PSA-positive human neuroblastoma cell lines induced by retinoic acid (RA), phorbolester, or growth factors. Using a semiquantitative RT PCR strategy, we demonstrated a general decrease in the mRNA level of ST8SiaII upon differentiation of SH-SY5Y and LAN-5 cells. In contrast, a drastic increase of ST8SiaIV was specifically induced by RA-treatment of SH-SY5Y cells. To explore the significance of these changes, the cellular capacity to perform PSA synthesis and the degree of NCAM polysialylation were analyzed. Our data indicate that the increased expression of ST8SiaIV enables an accelerated polysialylation of NCAM, which, however, is not converted into higher amounts of PSA. PMID- 11108614 TI - Rapid bidirectional modulation of mRNA expression and export accompany long-term facilitation and depression of Aplysia synapses. AB - Serotonin (5-HT) and the neuropeptide Phe-Met-Arg-Phe-amide (FMRFa) modulate synaptic efficacy of sensory neurons (SNs) of Aplysia in opposite directions and for long duration. Both long-term responses require changes in mRNA and protein synthesis. The SN-specific neuropeptide, sensorin A, is a gene product that appears to be increased by 5-HT and decreased by FMRFa. We examined whether changes in sensorin A mRNA levels in the cell body and neurites of SNs accompany long-term facilitation and depression. Both 5-HT and FMRFa evoked rapid changes in sensorin A mRNA levels in the SN cell bodies: an increase with 5-HT and a decrease with FMRFa. Parallel changes in sensorin A mRNA levels in SN neurites were detected 2 h and 4 h later. These rapid changes in mRNA expression and net export required the presence of the appropriate target motor cell L7. The neuromodulators failed to produce changes in mRNA expression or export when SNs were cultured alone or with the inappropriate target cell L11. The changes in mRNA expression were transient because mRNA levels returned to control values 24 h after treatment, while synaptic efficacy remained altered by the respective treatments. These results indicate that two neuromodulators produce distinct, but transient, target-dependent effects on expression and export of a cell-specific mRNA that correlate with changes in synaptic plasticity. PMID- 11108615 TI - Response biases in auditory forebrain regions of female songbirds following exposure to sexually relevant variation in male song. AB - In many species of songbirds, individual variation between the songs of competing males is correlated with female behavioral preferences. The neural mechanisms of song based female preference in songbirds are not known. Working with female European starlings (Sturnus vulgaris), we used immunocytochemistry for ZENK protein to localize forebrain regions that respond to sexually relevant variation in conspecific male song. The number of ZENK-ir cells in ventral caudo-medial neostriatum [NCMv] was significantly higher in females exposed to longer songs than in those exposed to shorter songs, whereas variation in the total duration of song exposure yielded no significant differences in ZENK expression. ZENK expression in caudo-medial ventral hyperstriatum [cmHV] was uniformly high in all subjects, and did not vary significantly among the three groups. These results suggest that subregions of NCM in female starlings are tuned to variation in male song length, or to song features correlated therewith. Female starlings exhibit robust behavioral preferences for longer over shorter male songs (Gentner and Hulse; Anim Behav 59:443-458, 2000). Therefore, the results of this study strongly implicate NCM in at least a portion of the perceptual processes underlying the complex natural behavior of female choice. PMID- 11108616 TI - Altered gravitational forces affect the development of the static vestibuloocular reflex in fish (Oreochromis mossambicus). AB - Young fish (Oreochromis mossambicus) were exposed to microgravity (micro g) for 9 to 10 days during space missions STS-55 and STS-84, or to hypergravity (hg) for 9 days. Young animals (stages 11-12), which had not yet developed the roll-induced static vestibuloocular reflex (rVOR) at micro g- and hg-onset, and older ones (stages 14-16), which had already developed the rVOR, were used. For several weeks afterwards, the rVOR was recorded after termination of mug and hg. Here are the main results: (1) In the stage 11-12 fish, the rVOR gain (response angle/roll angle) measured for roll angles 15 degrees, 30 degrees, and 45 degrees was not affected by microgravity if animals were rolled from the horizontal to the inclined posture, but was increased significantly if animals were rolled in the opposite manner. The rVOR amplitude (maximal eye movement during a complete 360 degrees roll) of micro g animals increased significantly by 25% compared to 1g controls during the first postflight week, but decreased to the control level during the second postflight week. Microgravity had no effect in stage 14-16 fish on either rVOR gain or amplitude. (2) After 3g exposure, both rVOR gain and amplitude were significantly reduced for both stage 11-12 and stage 15 fish. One g readaptation was completed during the second post-3g week. Hypergravity at 2 or 2.5 g had no effect. (3) Hypergravity at all three levels tested (2g, 2.5g, and 3g) accelerated the morphological development as assessed by external morphological markers. Exposure to micro g- or 3g-periods during an early developmental period modifies the physiological properties of the neuronal network underlying the static rVOR; in susceptible developmental stages, these modifications include sensitization by microgravity and desensitization by hypergravity. PMID- 11108617 TI - Single breath-hold 3D contrast-enhanced method for assessment of cardiac function. AB - Cardiac MRI function measurements are typically performed using 2D sequences and require multiple breath-holds to image the entire heart. A single 3D acquisition using a T(1)-shortening agent has many potential advantages over techniques that acquire multiple 2D images, including more consistent contrast and precise slice coverage. However, 3D techniques currently require much longer than a single breath-hold to complete. It has been shown that for MR angiography undersampled projection reconstruction can acquire much higher resolution per unit time than Fourier imaging with acceptable artifacts. By employing a gated, undersampled projection technique, high-resolution 3D multiphase volumes of the heart can be acquired in a single breath-hold. Short repetition times result in good myocardial suppression and a temporal aperture of 60 ms. PMID- 11108618 TI - k-space weighted image contrast (KWIC) for contrast manipulation in projection reconstruction MRI. AB - A novel technique for manipulating contrast in projection reconstruction MRI is described. The method takes advantage of the fact that the central region of k space is oversampled, allowing one to choose different filters to enhance or reduce the amount that each view contributes to the central region, which dominates image contrast. The technique is implemented into a fast spin-echo (FSE) sequence, and it is shown that multiple T(2)-weighted images can be reconstructed from a single image data set. These images are shown to be nearly identical to those acquired with the Cartesian-sampled FSE sequence at different effective echo times. Further, it is demonstrated that T(2) maps can be generated from a single image data set. This technique also has the potential to be useful in dynamic contrast enhancement studies, capable of yielding a series of images at a significantly higher effective temporal resolution than what is currently possible with other methods, without sacrificing spatial resolution. PMID- 11108619 TI - Interrelations of T(1) and diffusion of water in acute cerebral ischemia of the rat. AB - Interrelation of T(1) and diffusion of water was studied in rat models of acute global and focal cerebral ischemia. Cortical T(1), as quantified with an inversion recovery method, increased by 4-7% within a few minutes of global ischemia at 4.7 and 9.4 T, but a significantly smaller change was detected at 1.5 T. The initial T(1) change occurred within seconds of cardiac arrest, much earlier than the extensive diffusion drop after 1-2 min. Thus, the initial increase in T(1) upon acute cerebral ischemia is directly caused by cessation of blood flow. In transient middle cerebral artery occlusion (MCAO), prolonged T(1) relaxation was detected within 10 min, with a subsequent increase during the course of ischemia. Spin density did not change during the first hour, showing that T(1) increase was not caused by net accumulation of water. Interestingly, partial recovery of T(1) upon release of MCAO, occurring independent of long-term tissue outcome, was observed only in concert with diffusion recovery. PMID- 11108620 TI - Macromolecule and water magnetization exchange modeling in articular cartilage. AB - Magnetization exchange effects between the matrix macromolecules (e. g., collagen and proteoglycan) and water were examined in normal, deuterated, and proteoglycan depleted articular cartilage. Relaxation results (T(2), T(1rho), and T(1)) suggested that a four-site exchange scheme provided an accurate model for articular cartilage relaxation and interspin group coupling details. Magnetization exchange within the collagen-bulk-water, proteoglycan-collagen, and collagen fibrillar water-collagen cartilage subsystems were quantified. Although collagen-bulk-water was the largest of the cartilage coupling subsystems ( approximately 90% signal) and is exploited in MRI, the rates of magnetization transfer (MT) within the latter subsystems were appreciably larger. Magnetization exchange rates for proteoglycan-collagen and collagen fibrillar water-collagen were 120 s(-1) and 4.4 s(-1), respectively. The observation of these latter two exchange subsystems suggested potential clinical MRI-MT applications in detecting molecular abnormalities associated with osteoarthritis. PMID- 11108621 TI - Water diffusion compartmentation and anisotropy at high b values in the human brain. AB - Biexponential diffusion decay is demonstrated in the human brain in vivo using b factors up to 4000 sec mm(-2). Fitting of the signal decay data yields values for the slow and fast diffusion components and volume fractions in agreement with previous studies in rat and human brain. In addition, differences in the fitted parameters are demonstrated in the white and gray matter and diffusion anisotropy is demonstrated in both the slow and fast diffusing components. Apparent anisotropy in the component fractions is discussed in terms of directionally dependent exchange rates between the compartments. The lack of a relationship between the estimated contribution to the signal of the fast and slow components and echo time appears to rule out T(2) differences in the observed water compartments. Values obtained for the fast diffusion coefficient, including differences between white and gray matter and the degree of anisotropy are compatible with the predictions of extracellular diffusion of water based on tortuosity models and the diffusion of tetramethylammonium ions in rat brain. PMID- 11108622 TI - Different magnetization transfer effects exhibited by the short and long T(2) components in human brain. AB - Magnetization transfer ratios (MTRs) were measured separately for the two T(2) components in white matter. For both binomial and off-resonance sinc MT pulses, the MTR was larger for the short T(2) component than for the long T(2) component. This differential MT effect disappeared for delays between the MT pulse and the multi-echo pulse sequence longer than 200 msec, indicating exchange between the two components. When using the sinc MT pulse, the MTR for the short T(2) component was similar for different white matter structures, whereas it varied for different white matter structures when using the binomial pulse-a phenomenon attributed to direct saturation. When the sinc pulse frequency was brought closer to resonance, MTRs in white matter and doped water phantoms increased for both components but more so for the shorter T(2) component. This behavior was consistent with a Bloch equation model of direct saturation. PMID- 11108623 TI - High-resolution MRI characterization of human thrombus using a novel fibrin targeted paramagnetic nanoparticle contrast agent. AB - In this study, the sensitivity of a novel fibrin-targeted contrast agent for fibrin detection was defined in vitro on human thrombus. The contrast agent was a lipid-encapsulated perfluorocarbon nanoparticle with numerous Gd-DTPA complexes incorporated into the outer surface. After binding to fibrin clots, scanning electron microscopy of treated clots revealed dense accumulation of nanoparticles on the clot surfaces. Fibrin clots with sizes ranging from 0.5-7.0 mm were imaged at 4.7 T with or without treatment with the targeted contrast agent. Regardless of sizes, untreated clots were not detectable by T(1)-weighted MRI, while targeted contrast agent dramatically improved the detectability of all clots. Decreases in T(1) and T(2) relaxation times (20-40%) were measured relative to the surrounding media and the control clots. These results suggest the potential for sensitive and specific detection of microthrombi that form on the intimal surfaces of unstable atherosclerotic plaque. PMID- 11108624 TI - Magnetic resonance thermometry for predicting thermal damage: an application of interstitial laser coagulation in an in vivo canine prostate model. AB - Magnetic resonance image-guidance for interstitial thermal therapy has proven to be a valuable tool in its traditional role in device localization and, more recently, in monitoring heat deposition within tissue. However, a quantitative understanding of how temperature-time exposure relates to thermal damage is crucial if the predictive value of real-time MR thermal-monitoring is to be fully realized. Results are presented on interstitial laser coagulation of two canine prostate models which are shown to provide an opportunity to evaluate three models of thermal damage based on a threshold maximum temperature, an Arrhenius damage integral, and a temperature-time product. These models were compared to the resultant lesion margin as derived from post-treatment T(1)- and T(2) weighted MR images, as well as from direct histological evaluation of the excised canine prostate. Histological evaluation shows that the thermal-injury boundary can be predicted from a threshold-maximum temperature of approximately 51 degrees C or an equivalent Arrhenius t(43) period of 200 minutes, but it is not reliably predicted using the temperature-time product. The methods described in this study are expected to have implications for the treatment of benign prostatic hyperplasia and prostate cancer with interstitial laser coagulation, which will be the focus of future human studies. PMID- 11108625 TI - Diffusion tensor MRI of the spinal cord. AB - Apparent diffusion tensor (ADT) measurements on the spinal cord using a pulsed field-gradient (PFG) multi-shot echo-planar imaging (EPI) sequence are presented. In a study of 10 healthy volunteers, the obtained rotationally invariant anisotropy information is compared to the results obtained by the rotationally dependent methods. The water diffusivity in the direction parallel to the fibers was found to be almost 2.5 times higher than the average diffusivity in directions perpendicular to the fibers and showed cylindrically symmetric anisotropy characteristics. The influence of partial volume effects and the point spread function on the measured results was evaluated, and it was the concluded that a resolution of 1 mm in the read and phase directions is required to obtain unbiased values. Possible clinical implications were demonstrated by investigating the diffusion characteristics of 10 patients suffering from narrowing of the cervical canal. The changes in the diffusion characteristics were found to be large enough to allow a robust detection of diffusion changes in the spine, even in cases in which conventional T(2) and T(1) weighted images were unable to detect any lesion. PMID- 11108626 TI - In vivo quantitative microimaging of rat spinal cord at 7T. AB - In vivo T(2), ADC, and MT properties of the GM and WM of the rat spinal cord were measured at 7T in the cervical region. The GM T(2), T(2GM) = 43.2 +/- 1.0 msec is significantly reduced compared to the WM T(2), T(2WM) = 57.0 +/- 1.6 msec. Diffusion is anisotropic for both GM and WM, with a larger ADC value along the cord axis (ADC(GM//) = 1.05 +/- 0.09 10(-9) m(2)sec(-1) and ADC(WM//) = 1.85 +/- 0.18 10(-9) m(2)sec(-1)) than perpendicular to this plane (ADC(GM)( perpendicular) approximately 0.50 * 10(-9) m(2)sec(-1) and ADC(WM)( perpendicular) approximately 0.18 * 10(-9) m(2)sec(-1)). The MT properties do not significantly differ between the WM and the GM, but allow one to distinguish the thin CSF layer from the WM. DWI with the sensitizing gradient perpendicular to the cord axis leads to the best contrast between GM and WM in the cervical region. PMID- 11108627 TI - Noninvasive assessment of cardiac ischemic injury using (87)Rb and (23)Na MR imaging, (31)P MR, and optical spectroscopy. AB - The aim of the study was to compare and analyze different noninvasive indices of cell damage in the isolated pig heart model of regional ischemia. We used (23)Na and (87)Rb MR imaging to evaluate Na(+)/K(+) balance, (31)P MR spectroscopy to measure energetics, and optical spectroscopy to assess oxymyoglobin (MbO(2)). Hearts were subjected to 120-min occlusion of the left anterior descending artery and were then reperfused for 120 min. Reperfusion resulted in an increase in (23)Na (37 +/- 18% of the posterior wall) and decrease in (87)Rb (55 +/- 15%) image intensities, partial recovery of PCr, ATP, the total phosphates, and MbO(2) in the anterior wall. The above changes are consistent with the irreversible cell damage in the anterior wall, confirmed by lack of staining with triphenyltetrazolium chloride. Changes in Na(+) and Rb(+) in the infarct area inversely correlated and their ratio is a more sensitive index of cell injury than either of them alone. PMID- 11108628 TI - Intravascular susceptibility agent effects on tissue transverse relaxation rates in vivo. AB - Since vascular architecture differs among tissues, it was hypothesized that the change in transverse relaxation rate produced by a given tissue concentration of susceptibility contrast agent also varies by tissue. This is relevant to strategies to map regional blood volume by MRI using indicator dilution techniques. R*(2) was measured in rat organs over a range of susceptibility agent concentrations at 1.5 T. Rat red blood cells loaded with dysprosium-DTPA-BMA served as an intravascular susceptibility agent. Tissue samples were frozen in vivo and dysprosium concentrations were independently measured using inductively coupled plasma atomic emission spectroscopy. The slope (k) of R*(2) vs. tissue dysprosium concentration in sec(-1) mM(-1) for myocardium was 97.1 (95% C.I. 77. 0-117.2), liver 122.6 (108.3-136.9), spleen 22.5 (8.8-36.3), kidney 68.1 (58.6 77.6), and skeletal muscle 77.9 (4.1-151.6); k was significantly different (P < 0.05) for all pairings except those with skeletal muscle. Therefore, relative values of tissue blood volume derived from dynamic images of first pass contrast effects may be in error because k is not constant for different conditions. PMID- 11108629 TI - Quantitative gadopentetate-enhanced MRI of breast tumors: testing of different analytic methods. AB - This study assessed several proposed imaging strategies and analytic methods based on gadopentetate-enhanced MRI to differentiate benign from malignant breast tumors in a blinded experimental animal study. Steady-state dynamic MRI and first pass imaging, performed with either T(1)- or T*(2)- weighted sequences, were compared. Semiquantitative and quantitative analysis methods, based on empirical measures of the data or physiological models, were subsequently applied to the imaging datasets. Comparative measures provided pathologic distinction of benign from malignant tumors, tumor grading, and histologic determination of microvascular density. Of the eight tested methods, only one, an estimate of first-pass perfusion using T *(2)-weighted imaging, showed an almost significant (P = 0.05) difference between benign and malignant tumors and correlated almost significantly (r =.3, P = 0.06) with the tumor grade. All other tests, performed either with steady-state imaging or with T(1)-weighted first-pass imaging, failed to differentiate benign from malignant tumors. In addition, they yielded poor correlations with tumor grade and microvascular density. PMID- 11108630 TI - Impact of signal-to-noise on functional MRI. AB - Functional magnetic resonance imaging (fMRI) has recently been adopted as an investigational tool in the field of neuroscience. The signal changes induced by brain activations are small ( approximately 1-2%) at 1.5T. Therefore, the signal to-noise ratio (SNR) of the time series used to calculate the functional maps is critical. In this study, the minimum SNR required to detect an expected MR signal change is determined using computer simulations for typical fMRI experimental designs. These SNR results are independent of manufacturer, site environment, field strength, coil type, or type of cognitive task used. Sensitivity maps depicting the minimum detectable signal change can be constructed. These sensitivity maps can be used as a mask of the activation map to help remove false positive activations as well as identify regions of the brain where it is not possible to confidently reject the null hypothesis due to a low SNR. PMID- 11108631 TI - Low-latency temporal filter design for real-time MRI using UNFOLD. AB - To improve real-time control of interventional procedures such as guidance of catheters, monitoring of ablation therapy, or control of dosage during drug delivery, the image acquisition and reconstruction must be high speed and have low latency (small time delay) in processing. A number of different methods have been demonstrated which increase the speed of MR acquisition by decreasing the number of sequential phase-encodes. A design and implementation of the UNFOLD method which achieves the desired low latency with a recursive temporal filter is presented. The recursive filter design is characterized for this application and compared with more commonly used moving average filters. Experimental results demonstrate low-latency UNFOLD for two applications: 1) high-speed, real-time imaging of the heart to be used in conjunction with cardiac interventional procedures; and 2) the injection of drugs into muscle tissue with contrast enhancement, i.e., monitoring needle insertion and injection of a drug with contrast enhancement properties. Proof-of-concept was demonstrated by injecting a contrast agent. In both applications the UNFOLD technique was used to double the frame rate. PMID- 11108632 TI - Coronary angiography by real-time MRI with adaptive averaging. AB - Cardiac and respiratory motion present significant challenges for MR coronary angiography, which have not been completely resolved to date by either breath holding or respiratory navigation. Adaptive averaging during real-time MRI may provide a useful alternative to these techniques. In this method, cross correlation is used to automatically identify those real-time imaging frames in which the vessel is present, and to determine the location of the vessel within each frame. This information is then used for selective averaging of frames to increase the signal-to-noise ratio and to improve visualization of the vessel. The correlation theorem was employed to raise the speed of this algorithm by up to two orders of magnitude. Segmenting data collection and reconstruction into subimages allows the extension of this technique to higher spatial resolution. Adaptive averaging provides a robust method for coronary MRI which requires no breath-holding, navigation, or ECG gating. PMID- 11108633 TI - A weighted least-squares algorithm for estimation and visualization of relative latencies in event-related functional MRI. AB - The properties of the hemodynamic latencies in functional maps have been relatively unexplored. Accurate methods of estimating hemodynamic latencies are needed to take advantage of this feature of fMRI. A fully automated, weighted least-squares (WLS) method for estimating temporal latencies is reported. Using a weighted linear model, the optimal latency and amplitude of the fMRI response can be determined for those voxels that pass a detection threshold. There is evidence from previous studies that the hemodynamic response may be time-locked to the stimulus within certain limits, less variable earlier in its evolution, and able to resolve information about relative hemodynamic timing. This information can be used to test hypotheses about the sequence and spatial distribution of neural activity. The method can be used to weight the earliest evolution of the hemodynamic response more heavily and decrease bias resulting from the hemodynamic response function. Additionally, the WLS method can control for varying response shapes across the brain and improve latency comparisons between brain regions. The WLS method was developed to study the properties of hemodynamic latencies, which may be increasingly important as event-related fMRI continues to be advanced. PMID- 11108634 TI - Design and fabrication of a three-axis edge ROU head and neck gradient coil. AB - The design, fabrication, and testing of a complete three-axis gradient coil capable of imaging the human neck is described. The analytic method of constrained current minimum inductance (CCMI) was used to position the uniform region of the gradient coil adjacent to and extending beyond the physical edge of the coil. The average gradient efficiency of the three balanced axes is 0.37 mT/m/A and the average inductance is 827 microH. With maximum amplifier current of 200A and receive signal sweep width of +/-125 kHz, the average minimum FOV using this gradient set is 7.9 cm. The completed coil has an inner diameter of 32 cm, an outer diameter of 42 cm, and a length (including cabling connections) of 80 cm. The entire coil was built in-house. The structure is actively water cooled. Heating measurements were made to characterize the thermal response of the coil under various operating conditions and it was determined that a continuous current of 100A could be passed through all three axes simultaneously without increasing the internal coil temperature by more than 23 degrees C. Eddy current measurements were made for all axes. With digital compensation, the gradient eddy current components could be adequately compensated. A large B(o) eddy current field is produced by the Gz axis that could be corrected through the use of an auxiliary B(o) compensation coil. Preliminary imaging results are shown in both phantoms and human subjects. PMID- 11108635 TI - Very short echo time proton MR spectroscopy of human brain with a standard transmit/receive surface coil. AB - A method for localized proton spectroscopy of the human brain is proposed which can be used with a standard transmit/receive planar surface coil producing an inhomogeneous RF field. Water suppression is accomplished by a train of full passage adiabatic pulses with optimized frequencies and delays, which account for variation in the water resonance frequency and the spin-lattice relaxation time. The robust method requires minimal pulse calibration and provides high-quality spectra even at very short echo times and in the absence of outer volume saturation and, therefore, is well suited for clinical in vivo spectroscopy. Performance of the method is demonstrated on a test object and on MR spectra from the human brain at 3 T. PMID- 11108636 TI - Analysis of the measurement precision of arterial lumen and wall areas using high resolution MRI. AB - High-resolution MRI may be used to monitor the progression of human carotid atherosclerosis by measuring the lumen and wall area changes over time. The purpose of this study was to analyze the precision of quantitative measurements of lumen and wall areas. Two independent MR scans near the carotid bifurcation were conducted on eight patients within 2 weeks. The error of lumen area measurement was 6. 2%, 9.2%, and 9.7% for T(1), proton density, and T(2)-weighted images, respectively, and the error of wall area measurement was 10. 8%, 10.9%, and 12.0%. The precision of area measurement correlates strongly with image quality. PMID- 11108637 TI - An optimized method for estimating intracranial volume from magnetic resonance images. AB - The accuracy and efficiency of protocols to measure intracranial volume (ICV) from volumetric magnetic resonance imaging (MRI) studies has not been formally analyzed. The ICV of 30 control participants was obtained by tracing every slice of a MRI data set on which the cranial cavity appeared, and compared with estimated ICVs calculated by progressively selecting one of every x slices (i.e., "1-in-x") as a sampling strategy. The reliability and precision of each sampling strategy was then determined. There was virtually no reduction in reliability at the 1-in-10 sampling strategy, with a reliability exceeding 0.999. ICV can be confidently traced using a 1-in-10 sampling strategy, which should result in significant time savings. PMID- 11108638 TI - Human cardiac imaging at 3 T using phased array coils. AB - Using a two-element phased array receiver coil, single breath-hold, ECG gated cardiac images of signal-to-noise ratios up to 130 and contrast-to-noise ratios exceeding 35 between myocardium and blood were recorded at 3 T. At several locations within the myocardium, T*(2) and B(0) inhomogeneity were determined. Because of shorter T*(2) times and larger B(0) inhomogeneities attributable to enhanced susceptibility effects, real-time cardiac imaging, the use of spiral scans, and echo planar imaging are expected to be considerably more difficult at 3 T. PMID- 11108639 TI - Easy improvement of signal-to-noise in RARE-sequences with low refocusing flip angles. Rapid acquisition with relaxation enhancement. AB - It is demonstrated that the signal intensity in a RARE (TSE, FS...)-sequence with low refocusing flip angle alpha can be significantly increased by setting the flip angle of the first refocusing pulse to 90 degrees +alpha/2. In addition to the gain in signal intensity, the initial signal modulations over the first few echoes are reduced compared to a CPMG-echo train with constant alpha. PMID- 11108640 TI - Genetic epidemiology with a capital "E". AB - Three characteristics of genetic epidemiology that distinguish it from its parent disciplines are a focus on population-based research, a focus on the joint effects of genes and the environment, and the incorporation of the underlying biology of the disease into its conceptual models. These principles are illustrated by a review of the genetic epidemiology of breast and ovarian cancer. Descriptive and mechanistic models for the joint effects of genes and "environmental" risk factors such as hormones and reproductive events are compared to illustrate the need to understand the biology. The contribution of population-based research to the development of the evidence for the involvement of major genes, the discovery of BRCA1 and BRCA2, and their characterization is reviewed. Interactions of major susceptibility genes, metabolic genes, and hormones are also discussed. I conclude with some suggestions for future directions for the field, the journal, and the Society, including recent bioethics initiatives. I believe that the Society should reach out more to the epidemiology community and that the journal should shift its emphasis from pure methodology to also include more substantive papers that illustrate these principles. PMID- 11108641 TI - A single, sequential, genome-wide test to identify simultaneously all promising areas in a linkage scan. AB - Inflation of type I error occurs when conducting a large number of statistical tests in genome-wide linkage scans. Stringent alpha-levels protect against the high numbers of expected false positives but at the cost of more false negatives. A more balanced tradeoff is provided by the theory of sequential analysis, which can be used in a genome scan even when the data are collected using a fixed sample design. Sequential tests allow complete, simultaneous control of both the type I and II errors of each individual test while using the smallest possible sample size for analysis. For fixed samples, the excess N "saved" can be used in a confirmatory, replication phase of the original findings. Using the theory of sequential multiple decision procedures [Bechhoffer et al., 1968], we can replace the series of individual marker tests with a new single, simultaneous genome-wide test that has multiple possible outcomes and partitions all markers into two subsets: the "signal" versus the "noise," with an a priori specifiable genome wide error rate. These tests are demonstrated for the Haseman-Elston approach, are applied to real data, and are contrasted with traditional fixed-sampling tests in Monte Carlo simulations of repeated genome-wide scans. The method allows efficient identification of the true signals in a genome scan, uses the smallest possible sample sizes, saves the excess to confirm those findings, controls both types of error, and provides one elegant solution to the debate over the best way to balance between false positives and negatives in genome scans. PMID- 11108642 TI - Use of classification trees for association studies. AB - We propose the use of classification trees for association studies. This approach is applied to a data set from Genetic Analysis Workshop 9 (GAW9), and our analysis precisely identified two disease alleles. Our purpose is to demonstrate the great potential of tree-based analyses for genetic studies, and discuss some issues that warrant further investigation. PMID- 11108643 TI - Ascertainment issues in variance components models. AB - One of the main concerns in the family studies of complex diseases is the effect that ascertainment and correction for it may have on test procedures and estimators. Elston and Sobel [1979] and Hopper and Mathews [1982] proposed two ways to correct for ascertainment in the study of quantitative trait data. For single ascertainment, using a variance components approach, we present results of simulation studies comparing estimates from these two methods for different selection criteria. We also show results from simulations when ascertained families are analyzed either at random or by correcting for ascertainment. For discordant sibpairs, we compare a variance components model that incorporates ascertainment correction with the extreme discordant sib pairs (EDSP) design proposed by Risch and Zhang [1995]. Our results show that there is minimal difference between the two methods of ascertainment correction. In the presence of effects from a large genetic background and the segregation of a rare gene, both ascertainment affected the polygenic and environmental components of variance but had rather little impact on the estimate of the linked major gene component of variance. The results also show the EDSP is slightly more powerful the variance components procedures for common alleles, and the variance components procedure is much more powerful than using EDSP when there is a rare allele segregating in the population. PMID- 11108644 TI - Analysis of longitudinal data from twins. AB - Longitudinal data from twin pairs may be used to determine how the genetic effects influencing a quantitative trait change with age. Here a model for mixed longitudinal data of Huggins and Loesch [1998] on unrelated individuals is extended to twin studies. The model is fitted using robust statistical methods and a bootstrap procedure is proposed to estimate the percentiles. The method is applied to longitudinal twin data on body mass index in male and female twin pairs aged 5-18 years. PMID- 11108645 TI - Cancer incidence for Swedish twins studied by means of bivariate frailty models. AB - Cancer incidence rates for Swedish twins born between 1928 and 1965 and who both were alive at age 30 are studied by means of bivariate frailty models. Altogether, 7,280 fraternal (DZ) and 4,699 identical (MZ) twin pairs were followed up through December 31, 1995, for cancer status. The association between cancer incidence rates was statistically greater among the MZ than among the DZ pairs and stronger between women than between men; however, the magnitude of this association is relatively small and decreases over time. The relative decrease in dependency (association) is most easily detected using shared frailty models but may also be demonstrated, at least for women, using correlated frailty models. We also demonstrate that estimates of the correlation coefficient are similar when using any correlated frailty models derived from the power variance family but that these estimates disagree regarding the age at which the dependence is most important. The relative importance of dependence across age may sometimes be more interesting than the correlation coefficient itself. The latter may usually be estimated using alternative methods. Furthermore, when estimating correlation coefficients close to the boundary of the parameter space, simulation studies indicate that the correlated inverse Gaussian frailty model is more robust than the gamma frailty model. PMID- 11108646 TI - Bias in multipoint linkage analysis arising from map misspecification. AB - Multipoint linkage analysis methods are often used in human genetic studies. Although multipoint methods increase power for a linkage analysis and will become essential if use of diallelic markers becomes widespread, the methods in use assume an accurate meiotic marker map. Unfortunately, uncertainties in estimates of between-marker meiotic distances are large. Also, sex-averaged maps are generally used, but recombination rates differ in males and females. Both these types of map misspecification can lead to lod score bias, but such bias has not previously been systematically quantified. We examine multipoint lod score bias arising from these map misspecifications, in both the presence and absence of actual linkage. We define bias as the expected difference between the lod score computed under the misspecified map and that computed under the true map. With actual linkage, any map misspecification causes negative bias in lod scores, resulting in loss of power to detect linkage. In most cases, bias is modest, only reaching clearly detectable levels when both types of misspecification are substantial. In the absence of linkage, map misspecification can cause positive or negative bias: falsely assuming a 1:1 female:male ratio always causes positive bias; using too large a distance gives a positive bias; using too small a distance gives a negative bias. This bias can inflate the false-positive rate, especially when the sample size is modest. We conclude that although current sex averaged maps are suitable for a first-pass multipoint screen, the potential for bias from map misspecification should be evaluated in following up results from such an analysis. PMID- 11108647 TI - Using unaffected child trios to test for transmission distortion. AB - The transmission disequilibrium test (TDT), an alternative to case-control analysis that is not influenced by population stratification, focuses on affected child trios (ACTs) comprising affected cases and their parents. Unaffected child trios (UCTs) have also been proposed but mainly to rule out segregation distortion. To explore situations when UCTs are preferable to detect transmission distortion, we compared the number of UCTs and ACTs needed to achieve 80% power for a wide variety of scenarios. For a given genetic model, UCT sample size declined rapidly with increasing disease prevalence, whereas ACT sample size remained constant. Furthermore, at some prevalence value (40-60% depending on model parameters), detection of transmission distortion could be accomplished with fewer UCTs than ACTs. Such high prevalence may be found in special populations (diabetes among Pima Indians), secondary conditions (renal/retinal complications in diabetes), and pharmacogenetics (responders to treatment). Also, because exposure to an additional risk factor can increase the disease prevalence in an exposed sub-group, we explored how sample size requirements vary by exposure status. Whereas power differences between exposed and unexposed ACTs could be explained solely by genetic risk ratios, sub-group-specific disease prevalence also played an important role in UCTs. Finally, we considered the impact of including 5, 10, 20, or 30% misclassified ACTs in the UCT sample and found that a 24, 58, 180, or 550% larger sample would be required. In conclusion, UCTs can detect transmission distortion more effectively than ACTs when disease prevalence reaches 40-60%, although some efficiency may be lost owing to misclassification. Moreover, focusing on particular sub-groups defined by exposure status can potentially increase power, but such gains depend heavily on the nature of the gene-exposure interaction. PMID- 11108648 TI - Familial history of metabolic disorders and the multiple metabolic syndrome: the NHLBI family heart study. AB - A case-control study was conducted to investigate the association between family history of obesity, hypertension, and diabetes and the co-occurrence of metabolic disorders associated with the multiple metabolic syndrome (MMS). Included were 1,448 African and European American men and women aged 48-71 who participated in both the third cohort examination of the Atherosclerosis Risk in Communities study, 1992-1994, and phase I of the Family Heart Study 1993-1995. The joint occurrence of hypertension, dyslipidemia, and diabetes or impaired fasting glucose in an individual determined his/her status of "affected" (MMS: n = 97), while the absence of these three metabolic disorders determined his/her status of "unaffected" ( CONTROL: n = 527). First-degree relatives provided the information to calculate family risk scores (FRSs) for the phenotypes under study: obesity, diabetes and hypertension. Although the majority of cases were obese (76.3%), family history of obesity was associated only weakly with the MMS, while family history of diabetes, or hypertension was associated significantly with the MMS (controlling for age, race, gender, and sampling group). Obesity of cases and controls modified the strength of these associations-odds ratios were 2.5(95% CI:1.1-6.1) and 2.9(95% CI:1.2-7.0) for the diabetes and hypertension FRSs in the non-obese, while in obese individuals the respective odds ratios were 1.6(95% CI:0.9-2.8) and 1.7(95% CI:0.9-3.1). These results may imply that obesity, whether familial or environmental in nature, is associated with the development of the MMS, while in non-obese individuals a family history of diabetes, hypertension, or obesity is a marker of genetic predisposition to components of the MMS. PMID- 11108649 TI - Evidence of major gene control of cortical bone loss in humans. AB - Cortical index (CI) is the ratio of the combined cortical thickness to the total diameter of the bone. It serves for the assessment of the geometric properties of bone and for indirect evaluation of bone mass. CI is a useful predictor of osteoporosis. The aim of the present study was to test the hypothesis of major gene control of CI variation in a large sample of pedigrees from Chuvashia, Russia. Complex segregation analysis revealed that the major gene model of CI inheritance is the best fitting and most parsimonious for the present data. Parameters of the genotype-gender specific dependence of CI variation on age were estimated simultaneously with other parameters in the segregation analysis. The results of analysis showed that not only the baseline level of CI but also the age at onset of the involutive bone changes (inflection point) and the rate of the CI decrease with age (slope coefficient) are under control of the same major gene. Non-major gene effects shared by pedigree members (residual familial correlations) were found to be statistically insignificant. Approximately 73% of inter-individual variation in CI was attributable to the effects explicitly included in the model. PMID- 11108650 TI - Modeling the HLA component in rheumatoid arthritis: sensitivity to DRB1 allele frequencies. AB - Rheumatoid arthritis is an inflammatory disease for which positive associations have been described with some HLA-DRB1 alleles. The associated alleles share a similar amino acid sequence in the third hypervariable region, the shared epitope, but differ at position 71 and 86. It has been suggested that HLA susceptibility to rheumatoid arthritis could be due not only to the shared epitope but could also be influenced by specific amino acids at positions 71 and 86. In this study, we investigated the role of these amino acids in rheumatoid arthritis on 203 unrelated patients. An involvement of amino acid 71 was detected but no conclusion was possible regarding amino acid 86. A study of the sensitivity of the conclusions to marker allele frequencies was performed. We showed that the results obtained for amino acid 71 are not very sensitive to allele frequencies but those obtained at position 86 are highly sensitive. This emphasizes the importance of studying the robustness of results to variations in allele frequencies before conclusions are drawn. PMID- 11108651 TI - Associations of clinical features in neurofibromatosis 1 (NF1). AB - Neurofibromatosis 1 (NF1), an autosomal dominant disease, exhibits extreme clinical variability. This variability greatly increases the burden for affected families and impairs our ability to understand the pathogenesis of NF1. Recognition of heterogeneity within a disease may provide important pathogenic insights, therefore we tested clinical data from three large sets of NF1 patients for evidence that certain common features are more likely to occur in some NF1 patients than in others. Clinical information on 4,402 patients with NF1 was obtained from three independent databases. We examined associations between pairs of clinical features in individual affected probands. We also examined associations between the occurrence of individual features in affected relatives. Associations were summarized as odds ratios with 95% confidence intervals. We found associations between several pairs of features in affected probands: intertriginous freckling and Lisch nodules, discrete neurofibromas and plexiform neurofibromas, discrete neurofibromas and Lisch nodules, plexiform neurofibromas and scoliosis, learning disability or mental retardation and seizures. We also found associations between the occurrence of Lisch nodules, macrocephaly, short stature, and learning disability or mental retardation as individual features in parents and children with NF1. Our observations suggest that, contrary to established belief, some NF1 patients are more likely than others to develop particular manifestations of the disease. Genetic factors appear to determine the development of particular phenotypic features. PMID- 11108652 TI - Racial and genetic determinants of plasma factor XIII activity. AB - Factor XIII (F XIII), a plasma transglutaminase, is essential for normal hemostasis and fibrinolysis. Plasma F XIII consists of two catalytic A (F XIIIA) and two non-catalytic B (F XIIIB) subunits. Activated F XIII is involved in the formation of fibrin gel by covalently crosslinking fibrin monomers. As the characteristics of the fibrin gel structure have been shown to be associated with the risk of coronary heart disease (CHD), F XIII activity may play a seminal role in its etiology. In this investigation, we determined plasma F XIII activity in two racial groups, including Asian Indians (n = 258) and Chinese (n = 385). Adjusted plasma F XIII activity was significantly higher in Indian men (142 vs. 110%; P<0.0001) and women (158 vs. 111%; P<0.0001) than their Chinese counterparts. As compared to Indians where the distribution of F XIII activity was almost normal, in Chinese it was skewed towards low activity. In both racial groups, bivariate and multivariate analyses showed strong correlation of F XIII activity with plasma fibrinogen and plasminogen levels. Race explained about 25% of the variation in F XIII activity even after the adjustment of significant correlates. We also determined the contribution of common genetic polymorphisms in the F XIIIA and F XIIIB genes in affecting plasma F XIII activity. Both loci showed significant and independent effects on plasma F XIII activity in Indians (F XIIIA, P< 0.01; F XIIIB, P<0.05) and Chinese (F XIIIA, P<0.0001; F XIIIB, P<0.13) in a gene dosage fashion. This study shows that both racial and genetic components play a significant role in determining plasma F XIII activity, and consequently it may affect the quantitative risk of CHD. PMID- 11108653 TI - Haseman and Elston revisited: the effects of ascertainment and residual familial correlations on power to detect linkage. PMID- 11108654 TI - Case-parents design for gene-environment interaction by Schaid. PMID- 11108655 TI - Multipoint admixture mapping. PMID- 11108656 TI - IGES resolution concerning recent allegations against James V. Neel. PMID- 11108657 TI - When is precancerous actually postcancerous? AB - Demonstration that certain rare cancer-related mutations can (i) be shared by adjoining benign and cancerous tumor regions or (ii) be present solely in a cancerous but not in an adjoining benign tumor region are data often cited in strong support of the conventional idea that benign tumor regions consist of precancerous cells. However, considering the well-documented evidence that many malignant cell types are still capable of regression through differentiation, one can envisage an alternative (or coincident) scenario whereby (i) mutations are shared by adjoining benign and cancerous tumor regions because a cancer cell with a non-differentiation-impairing mutation differentiates into a benign (postcancerous) cell or (ii) mutations are present solely in a cancerous tumor region because a cancer cell acquires a differentiation-impairing mutation that prevents its regression into a benign cell. Only with higher-resolution lineage analyses of a type not yet performed but experimentally feasible can these scenarios be distinguished. Accordingly, it is quite possible that common cancers regularly differentiate, such that a benign tumor region may actually harbor not only precancerous but also postcancerous cells. Demonstration of this phenomenon and elucidation of its mechanism could lead to novel therapeutics designed to effect reversion of the more common cancers that, when in advanced stages, are notoriously inadequately treated by current cytotoxic regimens. PMID- 11108659 TI - Inactivation of MMAC1 in bladder transitional-cell carcinoma cell lines and specimens. AB - We recently limited the location of a candidate tumor suppressor gene in invasive (T3a/b) bladder transitional-cell carcinoma (TCC) to a 2.5-cM region at chromosome 10q23.3. This region harbors the MMAC1/PTEN/TEP1 gene (referred to hereafter as MMAC1), a dual-phosphatase tumor-suppressor gene frequently inactivated in variety of malignant tumors. In the present study, we examined whether MMAC1 is a target for inactivation by mutations and deletions in bladder TCC cell lines and specimens. MMAC1 was inactivated by homozygous deletions and mutations in three (27%) of 11 bladder cancer cell lines. One cell line, UC-3, had homozygous deletions, and two other cell lines, T-24 and UC-9, had missense mutations. T-24 had also a nonsense mutation. However, none of the 33 bladder TCC specimens examined had a mutation or deletion in the coding region. These results suggest that MMAC1 is not the primary target for inactivation in bladder TCC and that another gene, in close proximity to the MMAC1 locus, within this region of frequent allelic losses, may be the target for inactivation. PMID- 11108658 TI - Activators of peroxisome proliferator-activated receptor-alpha partially inhibit mouse skin tumor promotion. AB - Several recent reports have suggested that peroxisome proliferator-activated receptors (PPARs) may be involved in the development of neoplasias in different tissue types. The present study was undertaken to determine whether PPARs play a role in skin physiology and tumorigenesis. In an initiation-promotion study, SENCAR mice treated topically with the PPARalpha ligands conjugated linoleic acid and 4-chloro-6-(2,3-xylidino)-2-pyrimidinylthioacetic acid (Wy-14643) exhibited an approximately 30% lower skin tumor yield compared with mice treated with vehicle. The PPARgamma and PPARdelta activators troglitazone and bezafibrate, respectively, exerted little, if any, inhibitory activity. PPARalpha was detected in normal and hyperplastic skin and in papillomas and carcinomas by immunohistochemistry. In addition, PPARalpha, PPARdelta/PPARbeta, and PPARgamma protein levels were analyzed by immunoblotting in normal epidermis and papillomas. Surprisingly, the levels of all three isoforms were increased significantly in tumors as opposed to normal epidermis. In primary keratinocyte cultures, protein levels of PPARalpha and, to a lesser extent, PPARgamma were markedly increased when the cells were induced to differentiate with high-calcium (0.12 mM) conditions. In addition, we observed that Wy-14643 enhanced transcriptional activity of a peroxisome proliferator-response element-driven promoter in a mouse keratinocyte cell line. These results demonstrate that keratinocytes express functional PPARalpha, that PPARalpha may play a role in differentiation, and that ligands for PPARalpha are moderately protective against skin tumor promotion. We conclude that selective PPARalpha ligands may exert their protective role against skin tumor promotion by ligand activation of PPARalpha. PMID- 11108660 TI - Loss of heterozygosity at 13q14 correlates with RB1 gene underexpression in human breast cancer. AB - Loss of heterozygosity (LOH) on the long arm of chromosome 13 is common in human breast tumors, pointing to the existence of several suppressor genes in this region. LOH at 13q14 has been implicated in alterations of retinoblastoma gene (RB1) expression. However, attempts to identify a link between the absence of retinoblastoma protein expression and LOH at the RB1 locus by means of immunohistochemical techniques have produced conflicting results. Therefore, we quantified RB1 mRNA by means of reverse transcriptase-polymerase chain reaction in a large series of human sporadic primary breast tumors. RB1 gene underexpression was observed in 28 (21.7%) of 129 breast-tumor RNAs. Allelic loss at this locus correlated with RB1 mRNA underexpression (P < 10(-7)), demonstrating a causal link. These data, based on a technique other than immunohistochemistry, confirm that RB1 is the main target of the 13q14 LOH observed in human breast cancer. We also found that RB1 underexpression correlated with Scarff-Bloom-Richardson (SBR) histopathologic grade III (P = 0.033), negative estrogen-receptor status (P = 0.026) and large tumor size (P = 0.010). The latter correlation was due mainly to a high mitotic index (one of the three components comprising SBR grade), suggesting that RB1 influences the proliferation rate of breast tumors. RB1 status (underexpression vs. normal expression) was not associated with subsequent relapse or with shorter relapse free survival. This study shows a major role of the RB1 gene in the pathogenesis of breast cancer. RB1 gene underexpression promotes breast-tumor aggressiveness and rapid tumor-cell proliferation, making RB1 an outstanding target for future gene-based breast-cancer therapy. PMID- 11108661 TI - Induced expression of dominant-negative c-jun downregulates NFkappaB and AP-1 target genes and suppresses tumor phenotype in human keratinocytes. AB - Neoplastically transformed mouse and human keratinocytes elevate transactivation of both activator protein 1 (AP-1) and nuclear factor kappaB (NFkappaB) transcription factors. The present study addresses the question of whether elevated NFkappaB in addition to elevated AP-1-dependent gene expression is necessary for maintaining the tumor cell phenotype. When a tetracycline regulatable dominant-negative c-jun (TAM67, having a truncated transactivation domain) was expressed in tumorigenic human keratinocytes, AP-1- and NFkappaB- but not p53-dependent reporter activity was inhibited by 40-60%. Tumor phenotype, as measured by anchorage-independent growth, was inhibited by 90%. Neither AP 1/NFkappaB activation nor expression of tumor phenotype was inhibited in TAM67 harboring keratinocytes under noninducing conditions. Electrophoretic mobility shift analysis showed that induction of TAM67 expression slightly increased AP-1- but reduced NFkappaB DNA-binding activity. Immunoprecipitation showed that TAM67 interacted in keratinocyte nuclei with NFkappaB p65, suggesting that inhibition of NFkappaB by TAM67 is mediated by direct protein-protein interactions, possibly producing decreased binding to DNA or inactivating p65. To analyze the putative effector genes that may be targeted by TAM67, expression of genes responsive to AP-1 or NFkappaB was measured by reverse transcriptase-polymerase chain reaction in TAM67 transfectants with or without TAM67 induction. Induction of TAM67 inhibited or reduced the expression of collagenase I, stromelysin I (AP-1 responsive), and interleukins 1 and 6 (NFkappaB responsive). These results indicate that genes controlled by NFkappaB and by AP-1 may be transformation relevant targets of TAM67 and that TAM67 may inhibit NFkappaB activation through direct interaction with NFkappaB p65. Moreover, the findings provide proof for the principle of using inducible TAM67 as a gene therapy to suppress tumor phenotype in human carcinoma cells. PMID- 11108662 TI - Role of multidrug-resistance protein 2 in glutathione S-transferase P1-1-mediated resistance to 4-nitroquinoline 1-oxide toxicities in HepG2 cells. AB - Previous studies in our laboratory have shown that the phase III efflux transporter multidrug-resistance protein (MRP)1 can act synergistically with the phase II conjugating glutathione S-transferases (GST) to confer resistance to the toxicities of some electrophilic drugs and carcinogens. To determine whether the distinct efflux transporter MRP2 could also potentiate GST-mediated protection from electrophilic toxins, we examined the effect of regulatable GSTP1-1 expression in MRP2-rich HepG2 cells on 4-nitroquinoline 1-oxide (4NQO)-induced cytotoxicity and genotoxicity (nucleic-acid adduct formation). Expression of GSTP1-1 was associated with a fourfold to tenfold protection from 4NQO-induced cytotoxicity. Inhibition of MRP2-mediated efflux activity by sulfinpyrazone or cyclosporin A completely reversed GSTP1-1-associated resistance-a result indicating that GSTP1-1-mediated cytoprotection is absolutely dependent on MRP2 efflux activity. Moreover, MRP2 efflux activity also augmented GSTP1-1-mediated protection from 4NQO-induced nucleic-acid adduct formation. We conclude that MRP2 mediated efflux of the glutathione conjugate of 4NQO and/or another toxic derivative of 4NQO is required to support GSTP1-1-associated protection from 4NQO toxicities in HepG2 cells. PMID- 11108664 TI - Lessons learned during 26 years as editor PMID- 11108663 TI - Downregulation of p53 by sustained JNK activation during apoptosis. AB - In a previous study, we prepared short-chain fatty acid (SCFA) mixtures mimicking the composition of the digested fibers from wheat bran, oat bran, pectin, and cellulose and tested the products on U4 cells, a cell-line model for normal colonocytes. These SCFA mixes induced the cyclin-dependent kinase (cdk) inhibitors p21 and p27, which bound to cdk2/cyclin E and cdk4/cyclin D1 complexes, blocking their kinase activity and arresting cell growth. SCFAs from digested fiber may control intestinal crypt height in vivo by inducing apoptosis in growth-arrested cells at the top of the crypt. In the present study, we report that SCFA mixes induced apoptosis of U4 cells and unexpectedly caused both a sustained activation of the stress-activated protein kinase c-jun N-terminal kinase 1 (JNK1) and downregulation of the tumor suppressor protein p53. JNK1 bound to p53, and the amount of JNK1-bound p53 accurately reflected the amount of total cellular p53. After activation by SCFAs, JNK1 phosphorylated its bound p53. This phosphorylation is likely to have converted p53 into an apoptotic target because p53 breakdown correlated with caspase-3 activity, was inhibited by a caspase-3 inhibitor in a dose-dependent manner, and was inhibited by transfection of dominant-negative JNK1. Because JNK1 activation was sustained in SCFA-treated U4 cells, JNK1 can bind, phosphorylate, and release p53 for proteolysis and then continue this cycle until many p53 molecules have been phosphorylated. Loss of p53 protein was likely due to proteolysis and not to transcriptional changes because a sixfold decrease in p53 protein occurred within 3-24 h of SCFA treatment, whereas p53 mRNA levels were downregulated as much only after 2-3 d. SCFA mixes targeted p53 and possibly other cellular proteins for degradation during apoptosis by causing a sustained activation of JNKs. PMID- 11108665 TI - Editorial consultants-2000 PMID- 11108666 TI - Iatrogenic coronary artery dissections extending into and involving the aortic root. AB - We set out to determine the incidence of iatrogenic coronary artery dissection extending into the aorta and to characterize the aortic lesions. We reviewed the data from 43,143 cardiac catheterizations from September 1993 through September 1999 and found 9 coronary artery-aortic dissections for an overall incidence of 0.02%. Four of these patients were undergoing treatment for acute myocardial infarction (AMI) and aortic dissection was more common than for non-AMI patients (0.19% vs. 0.01%, P < 0.0006). Histologic analysis of tissue samples from 2 cases revealed age related changes only and no evidence of predisposing pathology. Patients with limited aortic involvement were successfully managed with stenting of the coronary dissection entry point whereas aortic dissection extending up the aorta >40 mm from the coronary os required surgical intervention. PMID- 11108667 TI - Adenosine use during aortocoronary vein graft interventions reverses but does not prevent the slow-no reflow phenomenon. AB - Slow or no reflow (SNR) complicates 10-15% of cases of percutaneous intervention (PI) in saphenous vein bypass graft (SVG). To date there have been limited options for the prevention and treatment of this common and potentially serious complication. We evaluated the procedural outcome of 143 consecutive SVG interventions. We compared patients who received pre-intervention intra-graft adenosine boluses with those who did not. In addition we examined the efficacy of adenosine boluses to reverse slow-no reflow events. Angiograms were reviewed and flow graded (TIMI grade) by film readers blinded to the use of any intraprocedural drug or clinical history. Seventy patients received intragraft adenosine boluses before percutaneous intervention (APPI), 73 received no preintervention adenosine (NoAPPI). There were no significant angiographic differences between the two groups at baseline. A total of 20 patients experienced SNR. The incidence of SNR was similar in the two groups (APPI = 14.2% vs. NoAPPI = 13.6%, P = 0.9). SNR was treated with repeated, rapid boluses (24 microg each) of intra-graft adenosine. Reversal of SNR was observed in 10 of 11 patients (91%) who received high doses of adenosine (>/=5 boluses, mean 7.7 +/- 2.6) and in 3 of 9 (33%) of those who received low doses (<5 boluses, mean 1.5 +/ 1.2). Final TIMI flow was significantly better in the high dose than in the low dose group (final TIMI 2.7 +/- 0.6 vs. 2 +/- 0.8, P = 0.04). No significant untoward complications were observed during adenosine infusion. These findings suggest that SNR after PI in SVG is not prevented by pre-intervention adenosine, but it can be safely and effectively reversed by delivery of multiple, rapid and repeated boluses of 24 microg of intra-graft adenosine. PMID- 11108668 TI - The no-reflow phenomenon: a light at the end of the tunnel? PMID- 11108669 TI - Determinants of coronary blood flow following primary angioplasty for acute myocardial infarction. AB - The aim of this study was to evaluate determinants of coronary blood flow following primary angioplasty (PA) in acute myocardial infarction (AMI). The corrected TIMI (thrombolysis in myocardial infarction) frame count and the TIMI flow grade were used as indexes of coronary blood flow, and its determinants were examined in 115 consecutive AMI patients who underwent PA (pain onset 1,500 procedures a year) with an individual experience >500 procedures in either the radial approach or the percutaneous suture of the femoral artery with the Techstar/Prostar system, conducted a prospective study from January 1 to December 31, 1999. The aim of this study was to eliminate the occurrence of access site complications by using either one of two techniques that were at the operator's discretion, i.e., systematic radial approach, or percutaneous suture of the femoral artery. A total of 956 patients were included over the study period; 60.7% of these patients had percutaneous arterial closure of the femoral artery and the remaining 39.3% were treated via the radial approach; 88.7% were stented. The patients were administered a mean 9,000 IU of heparin during the procedure; 1.9% had been fibrinolyzed and Reopro was used in 5.9%. No complications were documented in the radial group. Of the 580 patients in the femoral suture group, 96.9% had femoral suture, immediately effective in 508 cases (90.4%). Only 3 patients required additional prolonged compression. One significant hematoma (0.2%) necessitating blood transfusion was reported in the femoral group. Infection at the puncture site with subsequent antibiotic treatment was reported in 2 patients (0.3%). No further access site complications were observed at one-month follow-up. After completion of the learning curve, the two techniques (radial approach and percutaneous arterial suture) permit the almost total elimination of access site complications. PMID- 11108673 TI - Beta-radiation for coronary in-stent restenosis. AB - To determine the feasibility and safety of an intracoronary beta-radiation device in preventing the recurrence of in-stent restenosis (ISR) after successful angioplasty, we studied 37 patients treated with beta-radiation (30-mm strontium 90 source) after angioplasty. The mean reference diameter was 2.9 +/- 0.5 mm, and 62% of lesions were diffuse, including four total occlusions. Beta-radiation was successfully delivered in 36 of 37 (97%) cases. Over the course of 7.1 +/- 4.5 mo follow-up, there were no myocardial infarctions and three deaths: one from preexisting malignancy, one from progressive cardiac failure, and one from sudden cardiac death. Target vessel revascularization (TVR) was performed in seven of 36 (19%) patients. Thirty patients underwent angiography at 6 mo; three (10%) experienced restenosis (diameter stenosis > 50%) at the target site, four (13%) had edge stenoses, and two (7%) had late (> 1 mo) thrombotic occlusions. Beta radiation for ISR is associated with encouragingly low rates of target lesion restenosis and TVR. Further improvements are needed to solve the limitations of the edge effect and late occlusion. PMID- 11108674 TI - Intracoronary radiation for in-stent restenosis: is the Greek alphabet important? PMID- 11108675 TI - Nonsurgical retrieval of embolized coronary stents. AB - Embolization of coronary stents before deployment is a rare but challenging complication of coronary stenting. Different methods for nonsurgical stent retrieval have been suggested. There were 20 cases (0.90%) of intracoronary stent embolization among 2,211 patients who underwent implantation of 4,066 stents. Twelve of 1,147 manually crimped stents (1.04%) and eight of 2,919 premounted stents were lost (0.27%, P < 0.01) during retraction of the delivery system, because the target lesion could not be either reached or crossed. Percutaneous retrieval was successfully carried out in 10 of 14 patients (71%) in whom retrieval was attempted. In 10 patients, stent retrieval was tried with 1.5-mm low-profile angioplasty balloon catheters (success in 7/10) and in seven cases with myocardial biopsy forceps or a gooseneck snare (success in 3/7). Three patients (15%) underwent urgent coronary artery bypass surgery after failed percutaneous retrieval, but their outcomes were fatal. In two patients, stents were compressed against the vessel wall by another stent, without compromising coronary blood flow. In two patients, a stent was lost to the periphery without clinical side effects; treatment was conservative in these cases. Embolization of stents before deployment is a rare but serious complication of coronary stenting, with hazardous potential for the patient. Manual mounting of stents is associated with a significantly higher risk of stent embolization. Stent retrieval from the coronary circulation with low-profile angioplasty balloon catheters is a readily available and technically familiar approach that has a relatively high success rate. PMID- 11108676 TI - Cutaneous oozing of lymphatic fluid after interventional cardiac catheterization in a patient with Noonan syndrome. AB - A 15-year-old girl with Noonan syndrome, intestinal lymphangiectasia and severe valvar pulmonary stenosis had an abnormal lymphangioscintigram that showed intense activity in the inguinal regions bilaterally. Cutaneous oozing of lymphatic fluid from the groin wound complicated percutaneous balloon pulmonary valvoplasty. This previously unreported complication highlights the risk of damage to abnormal lymphatic channels in patients with Noonan syndrome who undergo interventional catheterization. PMID- 11108677 TI - Initial experience using the Palmaz Corinthian stent for right ventricular outflow obstruction in infants and small children. AB - The original Palmaz balloon expandable stent has been used extensively for the treatment of vascular stenoses in older children and young adults. Placement of the Palmaz stent in infants and small children, however, is limited by stent inflexibility, large delivery sheath size, and concerns about creating fixed obstructions after the placement of small diameter stents in growing patients. New Palmaz Corinthian stents were placed through 6 French sheaths in four high risk patients with postoperative right ventricular outflow obstruction. Patients were not considered candidates for surgical repair. Median patient age and weight were 17 months (range 5-32 months) and 7.7 kg (range 4.6-11.1 kg), respectively. Median fluoroscopy time was 58.2 min (range 55.2-172 min). No complications were encountered. In each case, successful stent placement was achieved, and surgery with cardiopulmonary bypass was avoided. Palmaz Corinthian stents are more flexible, require a smaller delivery sheath, have equal or increased radial strength, and can be maximally expanded to a greater cross sectional area when compared to the original Palmaz stent. These characteristics make the Palmaz Corinthian stent a reasonable alternative for use in a select group of infants and small children who are not candidates for surgical repair of postoperative right ventricular outflow obstruction. PMID- 11108678 TI - Light at the end of the tunnel-but still dim. PMID- 11108679 TI - Exposure to ionizing radiation in children undergoing Amplatzer device placement to close atrial septal defects. AB - To evaluate exposure to ionizing radiation during Amplatzer device occlusion, a prospective study was performed to measure surface entrance radiation dose by thermoluminescent dosimetry (TLD). Between June 1998 and April 1999, dosimetry was carried out on 12 patients with Amplatzer device occlusion of atrial septal defects (n = 10) or Fontan fenestration (n =) and 12 age-matched patients who underwent diagnostic catherization. TLD chips were placed at the posterior (PA) and right lateral (LA) chest wall as well as the thyroid (TH) and gonadal (GN) regions. The Amplatzer group had a median age of 6.4 yr (2.4-12.4 yr) and a median weight of 23.7 kg (15.6-28.9 kg), which were similar (p = NS) to those of the control group, who had a median age of 7.9 yr (3.3-16.2 yr) and a median weight of 29.9 kg (10.6-58.0 kg). Device placement was successful in 11 of 12 patients; one device was removed owing to partial obstruction of the right-upper pulmonary vein. Fluoroscopy times were also similar in the Amplatzer group (23.5 +/- 2.1 min) and the control group (16.4 +/- 3.1 min; P = NS). The measured surface entrance doses of the Amplatzer group was similar (p = NS) to those of the control group in all four regions: PA (4.96 +/- 1.88 vs. 6.07 +/- 2.16 cGy), LA (5.22 +/- 1.68 vs. 3.13 +/- 1.25 cGy), TH (0.92 +/- 0.14 vs. 0.69 +/- 0.09 cGy), and GN (0.20 +/- 0.00 vs. 0.22 +/- 0.01cGy). Fluoroscopy times and measured surface entrance doses of ionizing radiation in patients undergoing Amplatzer device occlusion are similar to those in patients undergoing routine diagnostic catheterization. PMID- 11108680 TI - Steerable control of the eustachian valve during transcatheter closure of secundum atrial septal defects. AB - Over the past decade there has been increased use of transcatheter devices for closure of secundum atrial septal defects. The presence of a large eustachian valve complicating transcatheter closure has not been described. We describe four patients with prominent eustachian valves, in three of whom we employed a simple technique to obtain control of the eustachian valve during device placement using transesophageal echo guidance. PMID- 11108681 TI - Congenital agenesis of the right pulmonary artery. AB - A 44-year-old woman with recurrent pulmonary infections developed severe hemoptysis. Chest radiography revealed a hypoplastic right lung. Absence of the right pulmonary artery, a very rare congenital anomaly, was demonstrated by computed tomography and cardiac catheterization. Severe pulmonary hypertension in the contralateral lung precluded right pneumonectomy but percutaneous embolization of a large systemic arterial collateral to the right lung provided palliative relief of hemoptysis. PMID- 11108683 TI - Anomalous left anterior descending coronary artery: malignant hospital course of a not so benign anomaly. AB - Anomalous origin of the left anterior descending coronary artery from the right sinus of Valsalva has been described as a benign anomaly. Typically it takes one of two courses, intraseptal or prepulmonic (though pre-aortic and retro-aortic courses also have been described). We describe the treatment of a patient with prepulmonic route and a malignant course before revascularization. PMID- 11108682 TI - Mechanical compression of coronary artery stents: potential hazard for patients undergoing cardiopulmonary resuscitation. AB - Mechanical compression of coronary artery stents may be associated with a fatal outcome as the result of refractory myocardial ischemia. We present the history of an 83-yr-old patient, who died owing to hemorrhagic shock 3 days after stent implantation, despite immediate cardiopulmonary resuscitation (CPR). Postmortem examination showed stent compression, probably due to mechanical deformation during CPR. This complication has been reported in two other cases in the literature, suggesting that CPR may be hazardous to patients with coronary artery stents. PMID- 11108684 TI - Infected endovascular stents managed with medical therapy alone. AB - More than 400,000 endovascular stents are put in place in the United States annually. Infectious complications have been reported in fewer than one in 10,000 cases. It remains unclear whether the optimal management strategy for these patients is with medicine alone or surgery. We report two cases of endovascular stent infections that were treated successfully with antibiotics alone. PMID- 11108685 TI - Occlusive systolic bridging of circumflex artery. AB - Intramural course of coronary arteries is a well known phenomenon on autopsy. On angiography, however, it is usually seen in relation to the left anterior descending artery. Clinically this entity may be associated with myocardial ischemia, infarction or sudden cardiac death. In this case we report an unusual angiographic finding of circumflex coronary bridging. PMID- 11108686 TI - Percutaneous removal of a large mobile right atrial thrombus using a basket retrieval device. AB - This report documents the removal of a large mobile serpiginous right atrial thrombus using percutaneous intravascular techniques with fluoroscopic and transesophageal echocardiographic guidance. Technical aspects and potential complications are discussed. PMID- 11108687 TI - Broken guidewire fragment: a simplified retrieval technique. AB - We report a technique for retrieval of a broken angioplasty wire fragment from the coronary system using a more simplified technique that does not involve the use of a snare or any other retrieval tool. With the use of an additional angioplasty wire and a balloon catheter, we could safely remove the broken wire fragment from the coronary system and circulation in a very short time. PMID- 11108688 TI - Successful treatment of coronary artery perforation in an abciximab-treated patient by microcoil embolization. AB - We describe a case of type 2 coronary artery perforation in a 73-year-old man undergoing coronary artery rotablation and stenting with abciximab therapy. The coronary artery perforation was successfully treated by coil embolization with Trufill pushable coils made from platinum alloy and synthetic fibers to promote maximum thrombogenicity. PMID- 11108689 TI - Paradoxical hypertensive response to nitroglycerin during cardiac catheterization. PMID- 11108690 TI - Focal elastic obstruction of the inferior vena cava. AB - Obstruction of the supra-hepatic inferior vena cava (IVC) is a common cause of hepatic venous hypertension and the most common cause of Budd-Chiari Syndrome. Because most cases of IVC obstruction go undiagnosed until Budd-Chiari Syndrome develops, the natural history of IVC obstruction is not well defined. We report a case of a focal, elastic, non-membranous obstruction of the IVC causing hepatic venous hypertension and elevated serum transaminases in a 36-year-old man. The obstruction was successfully treated with placement of a self-expanding metallic stent with normalization of hepatic transaminases. PMID- 11108691 TI - Percutaneous transluminal laser guide wire recanalization of chronic subclavian artery occlusion in symptomatic coronary-subclavian steal syndrome. AB - Treatment of subclavian artery stenosis by percutaneous balloon angioplasty and adjunctive stent placement was shown to be safe and efficacious, but it may be limited in tight stenoses and long occlusions. We describe the case of a patient who experienced progressive angina pectoris associated with signs of cerebrovertebral insufficiency 9 yr after bypass surgery, including left internal mammary artery (LIMA) grafting to the left anterior descending coronary artery. Angiography showed reversed flow through the LIMA graft into the subclavian artery and a 4-cm occlusion beginning at the origin of the left subclavian artery, representing a rare coronary-subclavian steal syndrome. After a conventional approach failed, recanalization was performed successfully using laser guide wire angioplasty with adjunctive stent placement in a combined radial and femoral approach. PMID- 11108692 TI - ACC/AHA guidelines for the management of patients with unstable angina and non-ST segment elevation myocardial infarction: executive summary and recommendations. PMID- 11108693 TI - Designing a computer-based simulator for interventional cardiology training. AB - Interventional cardiology training traditionally involves one-on-one experience following a master-apprentice model, much as other procedural disciplines. Development of a realistic computer-based training system that includes hand-eye coordination, catheter and guide wire choices, three-dimensional anatomic representations, and an integrated learning system is desirable, in order to permit learning to occur safely, without putting patients at risk. Here we present the first report of a PC-based simulator that incorporates synthetic fluoroscopy, real-time three-dimensional interactive anatomic display, and selective right- and left-sided coronary catheterization and angiography using actual catheters. Significant learning components also are integrated into the simulator. PMID- 11108694 TI - Computerized patient simulation to train the next generation of interventional cardiologists: can virtual reality take the place of real life? PMID- 11108695 TI - Transcatheter atrial septal defect occlusion in piglets by balloon detachable devices. AB - Detachable balloon devices were applied in the occlusion of experimental ASDs in 20 piglets. The detachable balloons were made from Latex; the occluder balloon was placed on the left atrial side and required a floppy disk and counter occluder(s) support from the right atrial side (17 experiments), or a second detachable balloon from the right side (3 experiments). Full occlusion was noticed in all cases. There was one device embolization in the descending aorta (device with a regular floppy disk and no counter-occluder). The device was covered by tissue in 3 to 4 weeks with the balloon flat on the septum in approximately 2 months; it required minimal rim and no wires in the left atrium. The double balloon model was wireless. The balloon detachable device was found effective and safe in the occlusion of experimental ASDs in piglets. PMID- 11108696 TI - ISMB-2000: bioinformatics enters a new millennium PMID- 11108697 TI - Ballast: blast post-processing based on locally conserved segments. AB - MOTIVATION: Blast programs are very efficient in finding relatively strong similarities but some very distantly related sequences are given a very high Expect value and are ranked very low in Blast results. We have developed Ballast, a program to predict local maximum segments (LMSs-i.e. sequence segments conserved relatively to their flanking regions) from a single Blast database search and to highlight these divergent homologues. The TBlastN database searches can also be processed with the help of information from a joint BlastP search. RESULTS: We have applied the Ballast algorithm to BlastP searches performed with sequences belonging to well described dispersed families (aminoacyl-tRNA synthetases; helicases) against the SwissProt 38 database. We show that Ballast is able to build an appropriate conservation profile and that LMSs are predicted that are consistent with the signatures and motifs described in the literature. Furthermore, by comparing the Blast, PsiBlast and Ballast results obtained on a well defined database of structurally related sequences, we show that the LMSs provide a scoring scheme that can concentrate on top ranking distant homologues better than Blast. Using the graphical user interface available on the Web, specific LMSs may be selected to detect divergent homologues sharing the corresponding properties with the query sequence without requiring any additional database search. PMID- 11108698 TI - PHAT: a transmembrane-specific substitution matrix. Predicted hydrophobic and transmembrane. AB - MOTIVATION: Database searching algorithms for proteins use scoring matrices based on average protein properties, and thus are dominated by globular proteins. However, since transmembrane regions of a protein are in a distinctly different environment than globular proteins, one would expect generalized substitution matrices to be inappropriate for transmembrane regions. RESULTS: We present the PHAT (predicted hydrophobic and transmembrane) matrix, which significantly outperforms generalized matrices and a previously published transmembrane matrix in searches with transmembrane queries. We conclude that a better matrix can be constructed by using background frequencies characteristic of the twilight zone, where low-scoring true positives have scores indistinguishable from high-scoring false positives, rather than the amino acid frequencies of the database. The PHAT matrix may help improve the accuracy of sequence alignments and evolutionary trees of membrane proteins. PMID- 11108699 TI - Identification of novel multi-transmembrane proteins from genomic databases using quasi-periodic structural properties. AB - MOTIVATION: Identification of novel G protein-coupled receptors and other multi transmembrane proteins from genomic databases using structural features. RESULTS: Here we describe a new algorithm for identifying multi-transmembrane proteins from genomic databases with a specific application to identifying G protein coupled receptors (GPCRs) that we call quasi-periodic feature classifier (QFC). The QFC algorithm uses concise statistical variables as the 'feature space' to characterize the quasi-periodic physico-chemical properties of multi transmembrane proteins. For the case of identifying GPCRs, the variables are then used in a non-parametric linear discriminant function to separate GPCRs from non GPCRs. The algorithm runs in time linearly proportional to the number of sequences, and performance on a test dataset shows 96% positive identification of known GPCRs. The QFC algorithm also works well with short random segments of proteins and it positively identified GPCRs at a level greater than 90% even with segments as short as 100 amino acids. The primary advantage of the algorithm is that it does not directly use primary sequence patterns which may be subject to sampling bias. The utility of the new algorithm has been demonstrated by the isolation from the Drosophila genome project database of a novel class of seven transmembrane proteins which were shown to be the elusive olfactory receptor genes of Drosophila. PMID- 11108700 TI - MaxSub: an automated measure for the assessment of protein structure prediction quality. AB - MOTIVATION: Evaluating the accuracy of predicted models is critical for assessing structure prediction methods. Because this problem is not trivial, a large number of different assessment measures have been proposed by various authors, and it has already become an active subfield of research (Moult et al. (1997,1999) and CAFASP (Fischer et al. 1999) prediction experiments have demonstrated that it has been difficult to choose one single, 'best' method to be used in the evaluation. Consequently, the CASP3 evaluation was carried out using an extensive set of especially developed numerical measures, coupled with human-expert intervention. As part of our efforts towards a higher level of automation in the structure prediction field, here we investigate the suitability of a fully automated, simple, objective, quantitative and reproducible method that can be used in the automatic assessment of models in the upcoming CAFASP2 experiment. Such a method should (a) produce one single number that measures the quality of a predicted model and (b) perform similarly to human-expert evaluations. RESULTS: MaxSub is a new and independently developed method that further builds and extends some of the evaluation methods introduced at CASP3. MaxSub aims at identifying the largest subset of C(alpha) atoms of a model that superimpose 'well' over the experimental structure, and produces a single normalized score that represents the quality of the model. Because there exists no evaluation method for assessment measures of predicted models, it is not easy to evaluate how good our new measure is. Even though an exact comparison of MaxSub and the CASP3 assessment is not straightforward, here we use a test-bed extracted from the CASP3 fold-recognition models. A rough qualitative comparison of the performance of MaxSub vis-a-vis the human-expert assessment carried out at CASP3 shows that there is a good agreement for the more accurate models and for the better predicting groups. As expected, some differences were observed among the medium to poor models and groups. Overall, the top six predicting groups ranked using the fully automated MaxSub are also the top six groups ranked at CASP3. We conclude that MaxSub is a suitable method for the automatic evaluation of models. PMID- 11108701 TI - Extending the method of mathematically controlled comparison to include numerical comparisons. AB - MOTIVATION: The method of mathematically controlled comparison has been used for some time to determine which of two alternative regulatory designs is better according to specific quantitative criteria for functional effectiveness. In some cases, the results obtained using this technique are general and independent of parameter values and the answers are clear-cut. In others, the result might be general, but the demonstration is difficult and numerical results with specific parameter values can help to clarify the situation. In either case, numerical results with specific parameter values can also provide an answer to the question of how much larger the values might be. In contrast, a more ambiguous result is obtained when either of the alternatives can have the larger value for a given systemic property, depending on the specific values of the parameters. In any case, introduction of specific values for the parameters reduces the generality of the results. Therefore, we have been motivated to develop and apply statistical methods that would permit the use of numerical values for the parameters and yet retain some of the generality that makes mathematically controlled comparison so attractive. RESULTS: We illustrate this new numerical method in a step-by-step application using a very simple didactic example. We also validate the results by comparison with the corresponding results obtained using the previously developed analytical method. The analytical approach is briefly present for reference purposes, since some of the same key concepts are needed to understand the numerical method and the results are needed for comparison. The numerical method confirms the qualitative differences between the systemic behavior of alternative designs obtained from the analytical method. In addition, the numerical method allows for quantification of the differences and it provides results that are general in a statistical sense. For example, the older analytical method showed that overall feedback inhibition in an unbranched pathway makes the system more robust whereas it decreases the stability margin of the steady state. The numerical method shows that the magnitudes of these differences are not comparable. The differences in stability margins (1-2% on average) are small when compared to the differences in robustness (50-100% on average). Furthermore, the numerical method shows that the system with overall feedback responds more quickly to change than the otherwise equivalent system without overall feedback. These results suggest reasons why overall feedback inhibition is such a prevalent regulatory pattern in unbranched biosynthetic pathways. PMID- 11108702 TI - Engineering support vector machine kernels that recognize translation initiation sites. AB - MOTIVATION: In order to extract protein sequences from nucleotide sequences, it is an important step to recognize points at which regions start that code for proteins. These points are called translation initiation sites (TIS). RESULTS: The task of finding TIS can be modeled as a classification problem. We demonstrate the applicability of support vector machines for this task, and show how to incorporate prior biological knowledge by engineering an appropriate kernel function. With the described techniques the recognition performance can be improved by 26% over leading existing approaches. We provide evidence that existing related methods (e.g. ESTScan) could profit from advanced TIS recognition. PMID- 11108703 TI - An iterative method for faster sum-of-pairs multiple sequence alignment. AB - MOTIVATION: Multiple sequence alignment is an important tool in computational biology. In order to solve the task of computing multiple alignments in affordable time, the most commonly used multiple alignment methods have to use heuristics. Nevertheless, the computation of optimal multiple alignments is important in its own right, and it provides a means of evaluating heuristic approaches or serves as a subprocedure of heuristic alignment methods. RESULTS: We present an algorithm that uses the divide-and-conquer alignment approach together with recent results on search space reduction to speed up the computation of multiple sequence alignments. The method is adaptive in that depending on the time one wants to spend on the alignment, a better, up to optimal alignment can be obtained. To speed up the computation in the optimal alignment step, we apply the alpha(*) algorithm which leads to a procedure provably more efficient than previous exact algorithms. We also describe our implementation of the algorithm and present results showing the effectiveness and limitations of the procedure. PMID- 11108704 TI - BALL--rapid software prototyping in computational molecular biology. Biochemicals Algorithms Library. AB - MOTIVATION: Rapid software prototyping can significantly reduce development times in the field of computational molecular biology and molecular modeling. Biochemical Algorithms Library (BALL) is an application framework in C++ that has been specifically designed for this purpose. RESULTS: BALL provides an extensive set of data structures as well as classes for molecular mechanics, advanced solvation methods, comparison and analysis of protein structures, file import/export, and visualization. BALL has been carefully designed to be robust, easy to use, and open to extensions. Especially its extensibility which results from an object-oriented and generic programming approach distinguishes it from other software packages. BALL is well suited to serve as a public repository for reliable data structures and algorithms. We show in an example that the implementation of complex methods is greatly simplified when using the data structures and functionality provided by BALL. PMID- 11108705 TI - Pathway analysis in metabolic databases via differential metabolic display (DMD). AB - MOTIVATION: A number of metabolic databases are available electronically, some with features for querying and visualizing metabolic pathways and regulatory networks. We present a unifying, systematic approach based on PETRI nets for storing, displaying, comparing, searching and simulating such nets from a number of different sources. RESULTS: Information from each data source is extracted and compiled into a PETRI net. Such PETRI nets then allow to investigate the (differential) content in metabolic databases, to map and integrate genomic information and functional annotations, to compare sequence and metabolic databases with respect to their functional annotations, and to define, generate and search paths and pathways in nets. We present an algorithm to systematically generate all pathways satisfying additional constraints in such PETRI nets. Finally, based on the set of valid pathways, so-called differential metabolic displays (DMDs) are introduced to exhibit specific differences between biological systems, i.e. different developmental states, disease states, or different organisms, on the level of paths and pathways. DMDs will be useful for target finding and function prediction, especially in the context of the interpretation of expression data. PMID- 11108706 TI - Detection of a surface-exposed PEST like sequence in the metabotropic glutamate receptor mGluR1 alpha. PMID- 11108707 TI - Profiles from structure based sequence alignment of porins can identify beta stranded integral membrane proteins. PMID- 11108709 TI - LIAN 3.0: detecting linkage disequilibrium in multilocus data. Linkage Analysis. AB - SUMMARY: LIAN is a program to test the null hypothesis of linkage equilibrium for multilocus data. LIAN incorporates both a Monte Carlo method as well as a novel algebraic method to carry out the hypothesis test. The program further returns the genetic diversity of the sample and the pairwise distances between its members. PMID- 11108708 TI - ODNBase--a web database for antisense oligonucleotide effectiveness studies. Oligodeoxynucleotides. AB - SUMMARY: ODNBase is a database of antisense oligodeoxynucleotides targeted to mammalian mRNAs that were reported in the literature. It includes the oligo sequences tested, the measured effectiveness, the RNA that was targeted, the type of measurement assay used, the oligo concentration applied, and the reference for each oligo. It provides a searchable interface by motif content, activity level, applied concentration and RNA name. Oligo lists matching search criteria can be downloaded in a spreadsheet compatible format. PMID- 11108710 TI - Surface-dependent coagulation enzymes. Flow kinetics of factor Xa generation on live cell membranes. AB - The initial surface reactions of the extrinsic coagulation pathway on live cell membranes were examined under flow conditions. Generation of activated coagulation factor X (fXa) was measured on spherical monolayers of epithelial cells with a total surface area of 41-47 cm(2) expressing tissue factor (TF) at >25 fmol/cm(2). Concentrations of reactants and product were monitored as a function of time with radiolabeled proteins and a chromogenic substrate at resolutions of 2-8 s. At physiological concentrations of fVIIa and fX, the reaction rate was 3.05 +/- 0.75 fmol fXa/s/cm(2), independent of flux, and 10 times slower than that expected for collision-limited reactions. Rates were also independent of surface fVIIa concentrations within the range 0.6-25 fmol/cm(2). The transit time of fX activated on the reaction chamber was prolonged relative to transit times of nonreacting tracers or preformed fXa. Membrane reactions were modeled using a set of nonlinear kinetic equations and a lagged normal density curve to track the expected surface concentration of reactants for various hypothetical reaction mechanisms. The experimental results were theoretically predicted only when the models used a slow intermediate reaction step, consistent with surface diffusion. These results provide evidence that the transfer of substrate within the membrane is rate-limiting in the kinetic mechanisms leading to initiation of blood coagulation by the TF pathway. PMID- 11108711 TI - Regulation of ribosomal S6 kinase 2 by effectors of the phosphoinositide 3-kinase pathway. AB - Ribosomal S6 kinase (S6K1), through phosphorylation of the 40 S ribosomal protein S6 and regulation of 5'-terminal oligopyrimidine tract mRNAs, is an important regulator of cellular translational capacity. S6K1 has also been implicated in regulation of cell size. We have recently identified S6K2, a homolog of S6K1, which phosphorylates S6 in vitro and is regulated by the phosphatidylinositide 3 kinase (PI3-K) and mammalian target of rapamycin pathways in vivo. Here, we characterize S6K2 regulation by PI3-K signaling intermediates and compare its regulation to that of S6K1. We report that S6K2 is activated similarly to S6K1 by the PI3-K effectors phosphoinositide-dependent kinase 1, Cdc42, Rac, and protein kinase Czeta but that S6K2 is more sensitive to basal activation by myristoylated protein kinase Czeta than is S6K1. The C-terminal sequence of S6K2 is divergent from that of S6K1. We find that the S6K2 C terminus plays a greater role in S6K2 regulation than does the S6K1 C terminus by functioning as a potent inhibitor of activation by various agonists. Removal of the S6K2 C terminus results in an enzyme that is hypersensitive to agonist-dependent activation. These data suggest that S6K1 and S6K2 are similarly activated by PI3-K effectors but that sequences unique to S6K2 contribute to stronger inhibition of its kinase activity. Understanding the regulation of the two S6K homologs may provide insight into the physiological roles of these kinases. PMID- 11108712 TI - Stimulation of extracellular signal-regulated kinases and proliferation in rat osteoblastic cells by parathyroid hormone is protein kinase C-dependent. AB - Parathyroid hormone (PTH) is known to have both catabolic and anabolic effects on bone. The dual functionality of PTH may stem from its ability to activate two signal transduction mechanisms: adenylate cyclase and phospholipase C. Here, we demonstrate that continuous treatment of UMR 106-01 and primary osteoblasts with PTH peptides, which selectively activate protein kinase C, results in significant increases in DNA synthesis. Given that ERKs are involved in cellular proliferation, we examined the regulation of ERKs in UMR 106-01 and primary rat osteoblasts following PTH treatment. We demonstrate that treatment of osteoblastic cells with very low concentrations of PTH (10(-12) to 10(-11) m) is sufficient for substantial increases in ERK activity. Treatment with PTH-(1-34) (10(-8) m), PTH-(1-31), or 8-bromo-cAMP failed to stimulate ERKs, whereas treatment with phorbol 12-myristate 13-acetate, serum, or PTH peptides lacking the N-terminal amino acids stimulated activity. Furthermore, the activation of ERKs was prevented by pretreatment of osteoblastic cells with inhibitors of protein kinase C (GF 109203X) and MEK (PD 98059). Treatment of UMR cells with epidermal growth factor (EGF), but not PTH, promoted tyrosine phosphorylation of the EGF receptor. Transient transfection of UMR cells with p21(N17Ras) did not block activation of ERKs following treatment with low concentrations of PTH. Thus, activation of ERKs and proliferation by PTH is protein kinase C-dependent, but stimulation occurs independently of the EGF receptor and Ras activation. PMID- 11108713 TI - Dimer formation of octaprenyl-diphosphate synthase (IspB) is essential for chain length determination of ubiquinone. AB - Ubiquinone (Q), composed of a quinone core and an isoprenoid side chain, is a key component of the respiratory chain and is an important antioxidant. In Escherichia coli, the side chain of Q-8 is synthesized by octaprenyl-diphosphate synthase, which is encoded by an essential gene, ispB. To determine how IspB regulates the length of the isoprenoid, we constructed 15 ispB mutants and expressed them in E. coli and Saccharomyces cerevisiae. The Y38A and R321V mutants produced Q-6 and Q-7, and the Y38A/R321V double mutant produced Q-5 and Q 6, indicating that these residues are involved in the determination of chain length. E. coli cells (ispB::cat) harboring an Arg-321 mutant were temperature sensitive for growth, which indicates that Arg-321 is important for thermostability of IspB. Intriguingly, E. coli cells harboring wild-type ispB and the A79Y mutant produced mainly Q-6, although the activity of the enzyme with the A79Y mutation was completely abolished. When a heterodimer of His-tagged wild type IspB and glutathione S-transferase-tagged IspB(A79Y) was formed, the enzyme produced a shorter length isoprenoid. These results indicate that although the A79Y mutant is functionally inactive, it can regulate activity upon forming a heterodimer with wild-type IspB, and this dimer formation is important for the determination of the isoprenoid chain length. PMID- 11108714 TI - Pancreatic beta-cell damage mediated by beta-cell production of interleukin-1. A novel mechanism for virus-induced diabetes. AB - Viral infection is one environmental factor that may initiate beta-cell damage during the development of autoimmune diabetes. Formed during viral replication, double-stranded RNA (dsRNA) activates the antiviral response in infected cells. In combination, synthetic dsRNA (polyinosinic-polycytidylic acid, poly(I-C)) and interferon (IFN)-gamma stimulate inducible nitric-oxide synthase (iNOS) expression, inhibit insulin secretion, and induce islet degeneration. Interleukin 1 (IL-1) appears to mediate dsRNA + IFN-gamma-induced islet damage in a nitric oxide-dependent manner, as the interleukin-1 receptor antagonist protein prevents dsRNA + IFN-gamma-induced iNOS expression, inhibition of insulin secretion, and islet degeneration. IL-1beta is synthesized as an inactive precursor protein that requires cleavage by the IL-1beta-converting enzyme (ICE) for activation. dsRNA and IFN-gamma stimulate IL-1beta expression and ICE activation in primary beta cells, respectively. Selective ICE inhibition attenuates dsRNA + IFN-gamma induced iNOS expression by primary beta-cells. In addition, poly(I-C) + IFN-gamma induced iNOS expression and nitric oxide production by human islets are prevented by interleukin-1 receptor antagonist protein, indicating that human islets respond to dsRNA and IFN-gamma in a manner similar to rat islets. These studies provide biochemical evidence for a novel mechanism by which viral infection may initiate beta-cell damage during the development of autoimmune diabetes. The viral replicative intermediate dsRNA stimulates beta-cell production of pro-IL 1beta, and following cleavage to its mature form by IFN-gamma-activated ICE, IL-1 then initiates beta-cell damage in a nitric oxide-dependent fashion. PMID- 11108715 TI - Evaluating the signal transduction mechanism of the parathyroid hormone 1 receptor. Effect of receptor-G-protein interaction on the ligand binding mechanism and receptor conformation. AB - Ligand binding to the PTH1 receptor is described by a "two-site" model, in which the C-terminal portion of the ligand interacts with the N-terminal domain of the receptor (N interaction), and the N-terminal region of the ligand binds the juxtamembrane domain of the receptor (J interaction). Previous studies have not considered the dynamic nature of receptor conformation in ligand binding and receptor activation. In this study the ligand binding mechanism was compared for the G-protein-coupled (RG) and uncoupled (R) PTH1 receptor conformations. The two site model was confirmed by demonstration of spatially distinct binding sites for PTH(3-34) and PTH(1-14): PTH(1-14), which binds predominantly to the J domain, only partially inhibited binding of 125I-PTH(3-34); and PTH(3-34), shown to bind predominantly to the N domain, only partially inhibited PTH(1-14)-stimulated cAMP accumulation. To assess the effect of R-G coupling, ligand binding to R was measured by displacement of 125I-PTH(3-34) with 30 microM guanosine 5'-3-O (thio)triphosphate (GTPgammaS) present, and binding to RG was measured by displacement of 125I-[MAP]PTHrP(1-36) (where MAP is model amphipathic peptide), a new radioligand that binds selectively to RG. Agonists bound with higher affinity to RG than R, whereas antagonists bound similarly to these states. The J interaction was responsible for enhanced agonist binding to RG: residues 1 and 2 were required for increased PTH(1-34) affinity for RG; residue 5 of MAP-PTHrP(1 36) was a determinant of R/RG binding selectivity, and PTH(1-14) bound selectively to RG. The N interaction was insensitive to R-G coupling; PTH(3-34) binding was GTPgammaS-insensitive. Finally, several observations suggest the receptor conformation is more "closed" at RG than R. At the R state, an open conformation is suggested by the simultaneous binding of PTH(1-14) and PTH(3-34). At RG PTH(1-14) better occluded binding of 125I-PTH(3-34) and agonist ligands bound pseudo-irreversibly, suggesting a more closed conformation of this receptor state. The results extend the two-site model to take into account R and RG conformations and suggest a model for differences of receptor conformation between these states. PMID- 11108716 TI - 7-Methylxanthine methyltransferase of coffee plants. Gene isolation and enzymatic properties. AB - Caffeine is synthesized through sequential three-step methylation of xanthine derivatives at positions 7-N, 3-N, and 1-N. However, controversy exists as to the number and properties of the methyltransferases involved. Using primers designed on the basis of conserved amino acid regions of tea caffeine synthase and Arabidopsis hypothetical proteins, a particular DNA fragment was amplified from an mRNA population of coffee plants. Subsequently, this fragment was used as a probe, and four independent clones were isolated from a cDNA library derived from coffee young leaves. Upon expression in Escherichia coli, one of them was found to encode a protein possessing 7-methylxanthine methyltransferase activity and was designated as CaMXMT. It consists of 378 amino acids with a relative molecular mass of 42.7 kDa and shows similarity to tea caffeine synthase (35.8%) and salicylic acid methyltransferase (34.1%). The bacterially expressed protein exhibited an optimal pH for activity ranging between 7 and 9 and methylated almost exclusively 7-methylxanthine with low activity toward paraxanthine, indicating a strict substrate specificity regarding the 3-N position of the purine ring. K(m) values were estimated to be 50 and 12 microM for 7 methylxanthine and S-adenosyl-l-methionine, respectively. Transcripts of CaMXMT could be shown to accumulate in young leaves and stems containing buds, and green fluorescent protein fusion protein assays indicated localization in cytoplasmic fractions. The results suggest that, in coffee plants, caffeine is synthesized through three independent methylation steps from xanthosine, in which CaMXMT catalyzes the second step to produce theobromine. PMID- 11108717 TI - In fibroblasts Vegf-D expression is induced by cell-cell contact mediated by cadherin-11. AB - Vascular endothelial growth factors (VEGFs) are a highly conserved family of growth factors all angiogenic in vivo with mitogenic and chemotactic activity on endothelial cells. VEGFs are expressed in fibroblasts either in hypoxia or in response to growth factors. Here we report that, differently from the other members of the family, Vegf-D is induced by cell-cell contact. By in situ hybridization we demonstrated that noninteracting fibroblasts express low levels of Vegf-D mRNA, whereas contacting cells express high levels of Vegf-D transcripts. By immunostaining we observed that the surface protein cadherin-11 is localized at the opposite sites of interacting cell surfaces. Ca(2+) deprivation from the culture medium determined the loss of cadherin-11 from the cell surfaces and down-regulation of Vegf-D mRNA. Moreover, a cadherin-11 antisense RNA construct inhibited Vegf-D expression in confluent BALB/c fibroblasts, whereas in NIH 3T3 cells, which express low levels of cadherin-11, Vegf-D induction could be obtained by overexpression of cadherin-11. This suggests that cell interaction mediated by cadherin-11 induces the expression of the angiogenic factor Vegf-D in fibroblasts. PMID- 11108719 TI - The BRG-1 subunit of the SWI/SNF complex regulates CD44 expression. AB - Aberrant regulation of CD44, a transmembrane glycoprotein, has been implicated in the growth and metastasis of numerous tumors. Although both CD44 overexpression and loss have been implicated in tumor progression, the mechanism of CD44 down regulation in these tumor types is not known. By immunoblot and reverse transcription-polymerase chain reaction analysis we determined that a cervical carcinoma cell line, C33A, lacks CD44 expression. To determine how CD44 is down regulated in C33A cells, we utilized cell fusions of C33A cells with a CD44 expressing cell line (SAOS-2). We found that SAOS-2 fusion restored CD44 expression in C33A cells, suggesting that a trans-acting factor present in SAOS-2 cells promotes CD44 production. C33A cells are BRG-1-deficient, and we found that CD44 was absent in another BRG-1-deficient tumor cell line, indicating that loss of BRG-1 may be a general mechanism by which cells lose CD44. Reintroduction of BRG-1 into these cells restored CD44 expression. Furthermore, disruption of BRG-1 function through the use of dominant-negative BRG-1 demonstrated the requirement of BRG-1 in CD44 regulation. Finally, we show that Cyclin E overexpression resulted in the attenuation of CD44 stimulation, which is consistent with previous observations that Cyclin E can abrogate BRG-1 action. Taken together, these results suggest that BRG-1 is a critical regulator of CD44 expression, thus implicating SWI/SNF components in the regulation of cellular adhesion and metastasis. PMID- 11108718 TI - Vascular endothelial growth factor expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and E-selectin through nuclear factor-kappa B activation in endothelial cells. AB - Vascular endothelial growth factor (VEGF) induces adhesion molecules on endothelial cells during inflammation. Here we examined the mechanisms underlying VEGF-stimulated expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and E-selectin in human umbilical vein endothelial cells. VEGF (20 ng/ml) increased expression of ICAM-1, VCAM-1, and E-selectin mRNAs in a time-dependent manner. These effects were significantly suppressed by Flk-1/kinase-insert domain containing receptor (KDR) antagonist and by inhibitors of phospholipase C, nuclear factor (NF)-kappaB, sphingosine kinase, and protein kinase C, but they were not affected by inhibitors of mitogen activated protein/extracellular signal-regulated kinase kinase (MEK) 1/2 or nitric-oxide synthase. Unexpectedly, the phosphatidylinositol (PI) 3'-kinase inhibitor wortmannin enhanced both basal and VEGF-stimulated adhesion molecule expression, whereas insulin, a PI 3'-kinase activator, suppressed both basal and VEGF-stimulated expression. Gel shift analysis revealed that VEGF stimulated NF kappaB activity. This effect was inhibited by phospholipase C, NF-kappaB, or protein kinase C inhibitor. VEGF increased VCAM-1 and ICAM-1 protein levels and increased leukocyte adhesiveness in a NF-kappaB-dependent manner. These results suggest that VEGF-stimulated expression of ICAM-1, VCAM-1, and E-selectin mRNAs was mainly through NF-kappaB activation with PI 3'-kinase-mediated suppression, but was independent of nitric oxide and MEK. Thus, VEGF simultaneously activates two signal transduction pathways that have opposite functions in the induction of adhesion molecule expression. The existence of parallel inverse signaling implies that the induction of adhesion molecule expression by VEGF is very finely regulated. PMID- 11108720 TI - Ribosomal S6 kinase 2 inhibition by a potent C-terminal repressor domain is relieved by mitogen-activated protein-extracellular signal-regulated kinase kinase-regulated phosphorylation. AB - Ribosomal S6 kinase 2 (S6K2) is a recently identified serine/threonine protein kinase that phosphorylates the 40 S ribosomal protein S6 in vitro. S6K2 is highly homologous to S6K1 in the core kinase and linker regulatory domains but differs from S6K1 in the N- and C-terminal regions and is differently localized primarily to the nucleus because of a C-terminal nuclear localization signal unique to S6K2. We have recently demonstrated that S6K2 is regulated similarly to S6K1 by the mammalian target of rapamycin pathway and by multiple PI3-K pathway effectors in vivo. However, deletion of the C-terminal domain of S6K2 enhances kinase activity, whereas analogous deletion of S6K1 is inhibitory. Here, we characterize the S6K2 C-terminal motifs that confer this differential regulation. We demonstrate that the inhibitory effects of the S6K2 C-terminal domain are only partly attributable to the nuclear localization signal but that three C-terminal proline-directed potential mitogen-activated protein kinase phosphorylation sites are critical mediators of this inhibitory effect. Site-specific mutation of these sites to alanine completely desensitizes S6K2 to activating inputs, whereas mutation to aspartic acid to mimic phosphorylation results in an activated enzyme which is hypersensitive to activating inputs. Pretreatment of cells with the mitogen-activated protein-extracellular signal-regulated kinase kinase (MEK) inhibitor U0126 inhibited S6K2 activation to a greater extent than S6K1. Furthermore, S6K2 mutants with C-terminal deletion or acidic phosphorylation site mutations displayed greatly reduced U0126 sensitivity. Thus, MEK-dependent inputs to C-terminal phosphorylation sites appear to be essential for relief of S6K2 inhibition but less critical for activation of S6K1. These data suggest a mechanism by which weak PI3-K agonists can regulate S6 phosphorylation and selective translation in the presence of mitogen-activated protein kinase signaling. PMID- 11108721 TI - Tel-2 is a novel transcriptional repressor related to the Ets factor Tel/ETV-6. AB - We report here the isolation of Tel-2, a novel member of the Ets transcription factor family, with high homology to Tel/ETV-6. Tel-2 is the second mammalian member of the Tel Ets family subclass whose prototype Tel is involved in various chromosomal translocations in human cancers. Six differentially expressed alternative splice products of Tel-2 were characterized encoding different Tel-2 isoforms which either contain or lack the amino-terminal Pointed domain and also vary at the carboxyl terminus. In contrast to Tel, which is highly expressed in several different cell types and tissues, Tel-2 is only weakly expressed in a variety of tissues and cell types, including placenta, prostate, spleen, liver, and lung. Tel-2 binds to functionally relevant Ets-binding sites of several genes and only the Tel-2 isoform containing the Pointed domain and the DNA-binding domain acts as a strong repressor of transcription. The retinoic acid receptor alpha and bone morphogenetic protein-6B (BMP-6) genes are specifically repressed by Tel-2 indicating a function for Tel-2 as an inhibitor of differentiation. Due to the important involvement of Tel in human cancer and the location of Tel-2 within the MHC cluster region, Tel-2 might be involved in chromosomal translocations in human cancer as well. PMID- 11108722 TI - A PhoP/PhoQ-induced Lipase (PagL) that catalyzes 3-O-deacylation of lipid A precursors in membranes of Salmonella typhimurium. AB - Pathogenic bacteria modify the structure of the lipid A portion of their lipopolysaccharide in response to environmental changes. Some lipid A modifications are important for virulence and resistance to cationic antimicrobial peptides. The two-component system PhoP/PhoQ plays a central role in regulating lipid A modification. We now report the discovery of a PhoP/PhoQ activated gene (pagL) in Salmonella typhimurium, encoding a deacylase that removes the R-3-hydroxymyristate moiety attached at position 3 of certain lipid A precursors. The deacylase gene (pagL) was identified by assaying for loss of deacylase activity in extracts of 14 random TnphoA::pag insertion mutants. The pagL gene encodes a protein of 185 amino acid residues unique to S. typhimurium and closely related organisms such as Salmonella typhi. Heterologous expression of pagL in Escherichia coli on plasmid pWLP21 results in loss of the R-3 hydroxymyristate moiety at position 3 in approximately 90% of the lipid A molecules but does not inhibit cell growth. PagL is synthesized with a 20-amino acid N-terminal signal peptide and is localized mainly in the outer membrane, as judged by assays of separated S. typhimurium membranes and by SDS-polyacrylamide gel analysis of membranes from E. coli cells that overexpress PagL. The function of PagL is unknown, given that S. typhimurium mutants lacking pagL display no obvious phenotypes, but PagL might nevertheless play a role in pathogenesis if it serves to modulate the cytokine response of an infected animal host. PMID- 11108723 TI - An olive oil-rich diet results in higher concentrations of LDL cholesterol and a higher number of LDL subfraction particles than rapeseed oil and sunflower oil diets. AB - We investigated the effect of olive oil, rapeseed oil, and sunflower oil on blood lipids and lipoproteins including number and lipid composition of lipoprotein subclasses. Eighteen young, healthy men participated in a double-blinded randomized cross-over study (3-week intervention period) with 50 g of oil per 10 MJ incorporated into a constant diet. Plasma cholesterol, triacylglycerol, apolipoprotein B, and very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), and low density lipoprotein (LDL) cholesterol concentrations were 10;-20% higher after consumption of the olive oil diet compared with the rapeseed oil and sunflower oil diets [analysis of variance (ANOVA), P < 0.05]. The size of IDL, VLDL, and LDL subfractions did not differ between the diets, whereas a significantly higher number (apolipoprotein B concentration) and lipid content of the larger and medium-sized LDL subfractions were observed after the olive oil diet compared with the rapeseed oil and sunflower oil diets (ANOVA, P < 0.05). Total HDL cholesterol concentration did not differ significantly, but HDL(2a) cholesterol was higher after olive oil and rapeseed oil compared with sunflower oil (ANOVA, P < 0.05).In conclusion, rapeseed oil and sunflower oil had more favorable effects on blood lipids and plasma apolipoproteins as well as on the number and lipid content of LDL subfractions compared with olive oil. Some of the differences may be attributed to differences in the squalene and phytosterol contents of the oils. PMID- 11108724 TI - Effects of overexpression of the amino-terminal fragment of apolipoprotein B on apolipoprotein B and lipoprotein production. AB - In vitro studies have shown that the binding site for microsomal triglyceride transfer protein (MTP) is within the first 17% of apoB (apoB-17). Expression of apoB-48 in McArdle cells decreases endogenous lipoprotein production; however, overexpression of human apoB in transgenic mice does not decrease endogenous mouse apoB expression. To assess this inconsistency, adenoviruses expressing human apoB-17 (AdB17) or apoB-17-beta (which contains apoB-17 plus a small lipid binding beta-sheet region of apoB, AdB-17beta) were produced. Hepatoma cells were infected with AdB17 or AdB17-beta with AdLacZ, an adenovirus expressing beta galactosidase, as a control. Overexpression of apoB-17 and apoB-17-beta in hepatoma cells to levels 2- to 3-fold greater than that of endogenous apoB did not alter endogenous apoB production. This was also true in the presence of oleic acid and N-acetyl-leucyl-leucyl-norleucinal. High levels of apoB-17 or beta galactosidase expression reduced apoB-100 production; however, control protein production was also reduced. To assess the effects of apoB-17 expression in vivo, mice of three different strains were injected with AdB17. Two days after injection, plasma apoB-17 was approximately 24 times the amount of endogenous apoB in the C57BL/6 mice, 2 times the apoB-100 in human apoB transgenic mice, and 4 times the apoB-48 in apoE knockout mice. Overexpression of apoB-17 did not decrease apoB-100 or apoB-48 concentrations in mouse plasma as assessed by Western blot analysis. These results demonstrate that although the apoB-17 binds to MTP in vitro, it does not alter endogenous apoB expression in mice or in hepatoma cells. PMID- 11108725 TI - Identification of peroxisomal targeting signals in cholesterol biosynthetic enzymes. AA-CoA thiolase, hmg-coa synthase, MPPD, and FPP synthase. AB - At least three different subcellular compartments, including peroxisomes, are involved in cholesterol synthesis. The peroxisomal targeting signals for phosphomevalonate kinase and isopentenyl diphosphate isomerase have been identified. In the current study we identify the peroxisomal targeting signals required for four other enzymes of the cholesterol biosynthetic pathway: acetoacetyl-CoA (AA-CoA) thiolase, 3-hydroxy-3-methylglutaryl-coenzyme A (HMG CoA) synthase, mevalonate diphosphate decarboxylase (MPPD), and farnesyl diphosphate (FPP) synthase. Data are presented that demonstrate that mitochondrial AA-CoA thiolase contains both a mitochondrial targeting signal at the amino terminus and a peroxisomal targeting signal (PTS-1) at the carboxy terminus. We also analyze a new variation of PTS-2 sequences required to target HMG-CoA synthase and MPPD to peroxisomes. In addition, we show that FPP synthase import into peroxisomes is dependent on the PTS-2 receptor and identify at the amino terminus of the protein a 20-amino acid region that is required for the peroxisomal localization of the enzyme. These data provide further support for the conclusion that peroxisomes play a critical role in cholesterol biosynthesis. PMID- 11108726 TI - Genetic control of HDL levels and composition in an interspecific mouse cross (CAST/Ei x C57BL/6J). AB - Strain CAST/Ei (CAST) mice exhibit unusually low levels of high density lipoproteins (HDL) as compared with most other strains of mice, including C57BL/6J (B6). This appears to be due in part to a functional deficiency of lecithin:cholesterol acyltransferase (LCAT). LCAT mRNA expression in CAST mice is normal, but the mice exhibit several characteristics consistent with functional deficiency. First, the activity and mass of LCAT in plasma and in HDL of CAST mice were reduced significantly. Second, the HDL of CAST mice were relatively poor in phospholipids and cholesteryl esters, but rich in free cholesterol and apolipoprotein A-I (apoA-I). Third, the adrenals of CAST mice were depleted of cholesteryl esters, a phenotype similar to that observed in LCAT- and acyl CoA:cholesterol acyltransferase-deficient mice. Fourth, in common with LCAT deficient mice, CAST mice contained triglyceride-rich lipoproteins with "panhandle"-like protrusions. To examine the genetic bases of these differences, we studied HDL lipid levels in an intercross between strain CAST and the common laboratory strain B6 on a low fat, chow diet as well as a high fat, atherogenic diet. HDL levels exhibited complex inheritance, as 12 quantitative trait loci with significant or suggestive likelihood of observed data scores were identified. Several of the loci occurred over plausible candidate genes and these were investigated. The results indicate that the functional LCAT deficiency is unlikely to be due to variations of the LCAT gene. Our results suggest that novel genes are likely to be important in the control of HDL metabolism, and they provide evidence of genetic factors influencing the interaction of LCAT with HDL. PMID- 11108727 TI - A simple and highly sensitive radioenzymatic assay for lysophosphatidic acid quantification. AB - The objective of the present work was to develop a simple and sensitive radioenzymatic assay to quantify lysophosphatidic acid (LPA). For that, a recombinant rat LPA acid acyltransferase (LPAAT) produced in Escherichia coli was used. In the presence of [(14)C]oleoyl-CoA, LPAAT selectively catalyzes the transformation of LPA and alkyl-LPA into [(14)C]phosphatidic acid. Acylation of LPA was complete and linear from 0 to 200 pmol with a minimal detection of 0.2 pmol. This method was used to quantify LPA in butanol-extracted lipids from bovine sera, as well as from human and mouse plasma. This radioenzymatic assay represents a new, simple, and highly sensitive method to quantify LPA in various biological fluids. PMID- 11108728 TI - Involvement of caveolin-1 in cholesterol enrichment of high density lipoprotein during its assembly by apolipoprotein and THP-1 cells. AB - High density lipoprotein (HDL) is assembled by interaction of apolipoprotein A-I with human monocytic leukemia cell line THP-1 by removing cellular cholesterol and phospholipid. Although the HDL formed with undifferentiated THP-1 cells contained only phosphatidylcholine and almost no cholesterol, the cells differentiated with phorbol 12-myristate 13-acetate (PMA) generated HDL enriched in cholesterol. The extent of cholesterol enrichment related to the cellular cholesterol level in the differentiated cells, but only weakly in the undifferentiated cells. In contrast, the differentiation had no influence on the diffusion-mediated cellular cholesterol efflux. The undifferentiated cells expressed the messages of ATP-binding cassette transporter 1 and caveolin-1, at low levels, and the PMA-induced differentiation resulted in substantial expression of both messages. Caveolin-1 protein expression was also highly induced by the PMA treatment of THP-1 cells. When the cells were treated with the antisense DNA of caveolin-1 and differentiated, both caveolin-1 synthesis and cholesterol incorporation into the HDL were reduced in parallel to generate the cholesterol-poor HDL. We concluded that caveolin-1 is involved in enrichment with cholesterol of the HDL generated by the apolipoprotein-cell interaction. This function is independent of the assembly of HDL particles with cellular phospholipid and of nonspecific, diffusion-mediated efflux of cellular cholesterol. PMID- 11108729 TI - Increased postprandial fatty acid trapping in subcutaneous adipose tissue in obese women. AB - The objective of this study was to test the hypothesis that increased fatty acid trapping by subcutaneous adipose tissue might contribute to the development and/or maintenance of obesity. To do so, venoarterial (V-A) gradients across subcutaneous adipose tissue for triglycerides, glycerol, nonesterified fatty acid (NEFA), and acylation-stimulating protein (ASP) were determined in eight lean females [body mass index (BMI), 22.2 +/- 0.6] and eight obese females (BMI, 34.4 +/- 3.4). Plasma insulin was also measured at intervals throughout this period. Fasting plasma triglyceride was significantly higher in the obese group and postprandial triglyceride was also significantly delayed. In contrast, both triglyceride clearance and fatty acid uptake by subcutaneous adipose tissue were significantly greater in the obese group compared with the lean group. Fasting insulin did not differ between the groups, but postprandial insulin values were significantly higher in the obese group. The pattern of ASP release from subcutaneous adipose tissue also appeared to differ in that it was significantly greater in the early postprandial period (0;-90 min) in the obese group versus the lean group and this correlated with greater triglyceride clearance during this period. Moreover, there were strong, positive correlations between BMI and the V-A gradient for fasting ASP, the 0- to 90-min area under the curve (AUC) for ASP V-A gradient fasting insulin, and the 0- to 90-min AUC for fatty acid incorporation into adipose tissue. Taken together, these data demonstrate that fatty acid trapping by adipose tissue can be increased even when overall plasma triglyceride clearance is delayed. The postprandial pattern of insulin, in particular, was altered in the obese, although it is certainly possible that differences in ASP release or response could also contribute to increased fatty acid trapping in the obese. The data, therefore, suggest that increased fatty acid trapping by adipose tissue may be a feature of some forms of obesity. PMID- 11108730 TI - Modulation of hepatic lipoprotein synthesis and secretion by taxifolin, a plant flavonoid. AB - In the present study, the effects of taxifolin, a plant flavonoid, on lipid, apolipoprotein B (apoB), and apolipoprotein A-I (apoA-I) synthesis and secretion were determined in HepG2 cells. Pretreatment of cells with (+/-)-taxifolin led to an inhibition of cholesterol synthesis in a dose- and time-dependent manner, with an 86 +/- 3% inhibition at 200 microM observed within 24 h. As to the mechanism underlying this inhibitory effect, taxifolin was shown to inhibit the activity of HMG-CoA reductase by 47 +/- 7%. In addition, cellular cholesterol esterification, and triacylglycerol and phospholipid syntheses, were also significantly suppressed in the presence of taxifolin. ApoA-I and apoB synthesis and secretion were then studied by pulse-chase experiments. ApoA-I secretion was found to increase by 36 +/- 10%. In contrast, an average reduction of 61 +/- 8% in labeled apoB in the medium was apparent with taxifolin. This effect on secretion appeared not to be exerted at the transcriptional level. Rather, the effect on apoB secretion was found to be exerted in the early stages of apoB degradation and to be sensitive to dithiothreitol (DTT) and insensitive to N-acetyl-leucyl-leucyl norleucinal, suggesting a proteolytic pathway involving a DTT-sensitive protease. Fractionation of secreted apoB revealed a slight shift in the distribution of secreted apoB-containing lipoproteins. Cholesteryl ester, rather than triacylglycerol, was shown to be the lipid that primarily regulated apoB secretion. In summary, our data suggest that taxifolin decreases hepatic lipid synthesis with a concomitant decrease and increase in apoB and apoA-I secretion, respectively. PMID- 11108731 TI - Long-term effects of cis and trans monounsaturated (18:1) and saturated (16:0) fatty acids on the synthesis and secretion of apolipoprotein A-I- and apolipoprotein B-containing lipoproteins in HepG2 cells. AB - The objective of this study was to compare the long-term effects of oleic (cis 18:1), elaidic (trans 18:1), and palmitic (16:0) acids on hepatic lipoprotein production, using HepG2 cells as an experimental model. The net accumulation in the medium of apolipoprotein A-I (apoA-I) was not significantly altered by fatty acids, whereas that of apoB was increased with oleic and elaidic acids. Oleic acid, and to a lesser extent elaidic and palmitic acids, increased the mass of triglycerides in the medium and the incorporation of [(3)H]glycerol into secreted triglycerides. The incorporation of [(14)C]acetate into cellular and secreted total cholesterol was stimulated by 96% and 83%, respectively, with elaidic acid but was not significantly modified by oleic or palmitic acid. Relative to oleic acid, the secretion of (14)C-labeled phospholipids and triglycerides was decreased 28% to 31% with elaidic and palmitic acids whereas that of free cholesterol and cholesteryl esters was enhanced 93% and 73%, respectively, with elaidic acid but remained unchanged with palmitic acid. Compared with oleic acid, elaidic acid stimulated the secretion of very low density lipoprotein cholesterol (VLDL-Chol), low density lipoprotein cholesterol (LDL-Chol), and high density lipoprotein cholesterol (HDL-Chol) by 43%, 70%, and 34%, respectively, whereas palmitic acid decreased VLDL-Chol but had no significant effect on LDL-Chol and HDL-Chol. The ratios of total cholesterol to HDL-Chol were 3.17, 3.60, and 3.25 with oleic, elaidic, and palmitic acids, respectively; the corresponding ratios of LDL-Chol to HDL-Chol were 0.87, 1.10, and 0.93, respectively. Compared with oleic and palmitic acids, the LDL and HDL particles secreted in the presence of elaidic acid contained higher levels of free cholesterol and cholesteryl esters and a lower content of phospholipids. The phospholipid-to-total cholesterol ratios of HDL were 1.05, 0.40, and 0.76 with oleic, elaidic, and palmitic acids, respectively. Our results indicate that in comparison with cis monounsaturated and saturated fatty acids, trans fatty acids have more adverse effects on the concentration and composition of lipoproteins secreted by HepG2 cells. PMID- 11108732 TI - Differential expression of ACAT1 and ACAT2 among cells within liver, intestine, kidney, and adrenal of nonhuman primates. AB - Two closely related enzymes with more than 50% sequence identity have been identified that catalyze the esterification of cholesterol using acyl-CoA substrates, namely acyl-CoA:cholesterol acyltransferase 1 (ACAT1) and ACAT2. Both are membrane-spanning proteins believed to reside in the endoplasmic reticulum of cells. ACAT2 has been hypothesized to be associated with lipoprotein particle secretion whereas ACAT1 is ubiquitous and may serve a more general role in cellular cholesterol homeostasis. We have prepared and affinity purified rabbit polyclonal antibodies unique to either ACAT enzyme to identify their cellular localization in liver and intestine, the two main lipoprotein-secreting tissues of the body, and for comparison, kidney and adrenal. In the liver, ACAT2 was identified in the rough endoplasmic reticulum of essentially all hepatocytes whereas ACAT1 was confined to cells lining the intercellular spaces among hepatocytes in a pattern typical of Kupffer cells. In the intestine, ACAT2 signal was strongly present in the apical third of the mucosal cells, whereas ACAT1 staining was diffuse throughout the mucosal cell, but with strong signal in goblet cells, Paneth cells, and villus macrophages. In the kidney, ACAT1 immunostaining was specific for the distal tubules and podocytes within the glomerulus. In the adrenal, ACAT1 signal was strongly present in the cells of the cortex, and absent from other adrenal cell types. No ACAT2 signal was identified in the kidney or adrenal. We conclude that only the cells of the liver and intestine that secrete apolipoprotein B-containing lipoproteins contain ACAT2, whereas ACAT1 is present in numerous other cell types. The data clearly suggest separate functions for these two closely related enzymes, with ACAT2 being most closely associated with plasma cholesterol levels. PMID- 11108733 TI - Effects of an Ala54Thr polymorphism in the intestinal fatty acid-binding protein on responses to dietary fat in humans. AB - A polymorphism in FABP2 that results in an alanine-to-threonine substitution at amino acid 54 of the intestinal fatty acid-binding protein (IFABP) is associated with insulin resistance in Pima Indians. In vitro, the threonine form (Thr54) has a higher binding affinity for long-chain fatty acids than does the alanine form (Ala54). We tested whether this polymorphism affected metabolic responses to dietary fat, in vivo. Eighteen healthy Pima Indians, half homozygous for the Thr54 form of IFABP and half homozygous for the Ala54 form, were studied. The groups were matched for sex, age, and body mass index. Plasma triglyceride, nonesterified fatty acid (NEFA), glucose, and insulin responses were measured after a mixed meal (35% of daily energy requirements, 50 g of fat) and after a high fat challenge (1362 kcal, 129 g of fat). NEFA concentrations were approximately 15% higher after the mixed meal and peaked earlier and were approximately 20% higher at 7 h in response to the high fat test meal in Thr54 homozygotes compared with Ala54 homozygotes. Insulin responses to the test meals tended to be higher in Thr54 homozygotes, but glucose and triglyceride responses were not different.The results of this study suggest that the Thr54 form of IFABP is associated with higher and prolonged NEFA responses to dietary fat in vivo. Higher NEFA concentrations may contribute to insulin resistance and hyperinsulinemia in individuals with this allele. PMID- 11108734 TI - Regulation by vitamin E of the scavenger receptor BI in rat liver and HepG2 cells. AB - The scavenger receptor class B type I (SR-BI) mediates the selective uptake of cholesterol and cholesteryl ester (CE) from high density lipoprotein (HDL) into cells. The high expression in liver and steroidogenic tissues is compatible with a role of SR-BI in reverse cholesterol transport and steroid hormone synthesis. Ways of regulation thus far described include induction by trophic hormones via cAMP-activated protein kinase A (PKA) and the effects of cellular and plasma cholesterol. Here we show that vitamin E (vitE) has a major effect on the expression of SR-BI in rat liver and in a human hepatoma-derived cell line, HepG2. Feeding rats a vitE-depleted diet resulted in an 11-fold increase in the SR-BI protein level in liver tissue. This effect was readily reversed by feeding a vitE-enriched chow. In HepG2 cells, the expression of the human SR-BI homolog was reduced when the vitE content was increased by incubating the cells with vitE loaded HDL or with phosphatidylcholine/vitE vesicles. The downregulation of human SR-BI (hSR-BI) was accompanied by a reduced level of protein kinase C (PKC) in the particulate cell fraction, and PKC inhibition decreased the expression of hSR BI and the uptake of vitE and cholesterol from HDL. Our results are consistent with the view that the cellular level of vitE exerts a tight control over the expression of SR-BI. Furthermore, the inhibitory effect of vitE on PKC seems to be involved in the signaling pathway. PMID- 11108735 TI - Oxidized LDL induces the expression of ALBP/aP2 mRNA and protein in human THP-1 macrophages. AB - The adipocyte lipid-binding protein (ALBP/aP2) belongs to a multigene family of fatty acid and retinoid transport proteins. This protein is abundantly expressed in the cytoplasm and nuclear region of adipocytes and is postulated to serve as a lipid shuttle, solubilizing hydrophobic fatty acids and delivering them to the appropriate metabolic system for utilization. This report demonstrates that human cholesterol-loaded THP-1 macrophages express ALBP/aP2 and that its expression can be stimulated by oxidized low density lipoprotein (oxLDL). The increase in mRNA expression was paralleled by a similar increase in ALBP/aP2 protein. The increase in ALBP/aP2 mRNA and protein in oxLDL-stimulated THP-1 macrophages is concentration and time dependent and is inhibited by treatment of the cells with an antioxidant inhibitor of nuclear factor-kappaB (NF-kappaB), pyrrolidine dithiocarbamate (PDTC), and with protein kinase C (PKC) inhibitors bisindolylmaleimide I and Ro-31-8220. These results suggest that activation of both NF-kappaB and PKC signaling pathways is necessary for oxLDL-induced ALBP/aP2 gene expression in THP-1 macrophages and that the upregulation of the fatty acid carrier may be a necessary event in foam cell formation. PMID- 11108736 TI - Lecithin:retinol acyltransferase from mouse and rat liver. CDNA cloning and liver specific regulation by dietary vitamin a and retinoic acid. AB - Lecithin:retinol acyltransferase (LRAT), present in microsomes, catalyzes the transfer of the sn-1 fatty acid of phosphatidylcholine to retinol bound to a cellular retinol-binding protein. In the present study we have cloned mouse and rat liver LRAT cDNA and tested the hypothesis that LRAT mRNA, like LRAT activity, is regulated physiologically in a liver-specific manner. The nucleotide sequences of mouse and rat liver LRAT cDNA each encode a 231-amino acid protein with 94% similarity between these species, and approximately 80% similarity to a cDNA for LRAT from human retinal pigment epithelium. Expression of rat LRAT cDNA in HEK293T cells resulted in functional retinol esterification and storage. RNA from several rat tissues hybridized with liver LRAT cDNA. However, LRAT mRNA was virtually absent from the liver of vitamin A-deficient animals, while being unaffected in intestine and testis. LRAT mRNA was rapidly induced by retinoic acid (RA) in liver of vitamin A-deficient mice and rats (P < 0.01). LRAT mRNA and enzymatic activity were well correlated in the same livers of rats treated with exogenous RA (r = 0.895, P < 0.0001), and in a dietary study that encompassed a broad range of vitamin A exposure (r = 0.799, P < 0.0001). Liver total retinol of <100 nmol/g was associated with low LRAT expression (<33% of control). We propose that RA, derived exogenously or from metabolism, serves as an important signal of vitamin A status. The constitutive expression of liver LRAT during retinoid sufficiency would serve to divert retinol into storage pools, while the curtailment of LRAT expression in retinoid deficiency would maintain retinol for secretion and delivery to peripheral tissues. PMID- 11108737 TI - Effect of apolipoprotein A-IV genotype and dietary fat on cholesterol absorption in humans. AB - We investigated the effect of the A-IV-2 allele, which encodes a Q360H substitution in apolipoprotein (apo) A-IV, and dietary fat on cholesterol absorption in humans. In three separate studies we compared fractional intestinal cholesterol absorption between groups of subjects heterozygous for the A-IV-2 allele (1/2) and homozygous for the common allele (1/1) receiving high cholesterol ( approximately 800 mg/day) diets with different fatty acid compositions. All subjects had the apoE 3/3 genotype. There was no difference in cholesterol absorption between the two genotype groups receiving a high saturated fat diet (33% of total energy as fat; 18% saturated, 3% polyunsaturated, 12% monounsaturated) or a low fat diet (22% of total energy as fat; 7% saturated, 7% polyunsaturated, 8% monounsaturated) diet. However, on a high polyunsaturated fat diet (32% of total energy as fat; 7% saturated, 13% polyunsaturated, 12% monounsaturated) mean fractional cholesterol absorption was 56. 7% +/- 1.9 in 1/1 subjects versus 47.5% +/- 2.1 in 1/2 subjects (P = 0.004). A post hoc analysis of the effect of the apoA-IV T347S polymorphism across all diets revealed a Q360H x T347S interaction on cholesterol absorption, and suggested that the A-IV-2 allele lowers cholesterol only in subjects with the 347 T/T genotype. We conclude that a complex interaction between apoA-IV genotype and dietary fatty acid composition modulates fractional intestinal cholesterol absorption in humans. PMID- 11108738 TI - Cholesterol, bile acid, and lipoprotein metabolism in two strains of hamster, one resistant, the other sensitive (LPN) to sucrose-induced cholelithiasis. AB - A comprehensive study of cholesterol, bile acid, and lipoprotein metabolism was undertaken in two strains of hamster that differed markedly in their response to a sucrose-rich/low fat diet. Under basal conditions, hamsters from the LPN strain differed from Janvier hamsters by a lower cholesterolemia, a higher postprandial insulinemia, a more active cholesterogenesis in both liver [3- to 4-fold higher 3 hydroxy 3-methylglutaryl coenzyme A reductase (HMG-CoAR) activity and mRNA] and small intestine, and a lower hepatic acyl-coenzyme A:cholesterol acyltransferase activity. Cholesterol saturation indices in the gallbladder bile were similar for both strains, but the lipid concentration was 2-fold higher in LPN than in Janvier hamsters. LPN hamsters had a lower capacity to transform cholesterol into bile acids, shown by the smaller fraction of endogenous cholesterol converted into bile acids prior to fecal excretion (0.34 vs. 0.77). In LPN hamsters, the activities of cholesterol 7alpha-hydroxylase (C7OHase) and sterol 27-hydroxylase (S27OHase), the two rate-limiting enzymes of bile acid synthesis, were disproportionably lower (by 2-fold) to that of HMG-CoAR. When fed a sucrose-rich diet, plasma lipids increased, dietary cholesterol absorption improved, hepatic activities of HMG-CoA reductase, C7Ohase, and S27OHase were reduced, and intestinal S27OHase was inhibited in both strains. Despite a similar increase in the biliary hydrophobicity index due to the bile acid enrichment in chenodeoxycholic acid and derivatives, only LPN hamsters had an increased lithogenic index and developed cholesterol gallstones (75% incidence), whereas Janvier hamsters formed pigment gallstones (79% incidence). These studies indicate that LPN hamsters have a genetic predisposition to sucrose-induced cholesterol gallstone formation related to differences in cholesterol and bile acid metabolism. PMID- 11108739 TI - LDL receptor deficiency unmasks altered VLDL triglyceride metabolism in VLDL receptor transgenic and knockout mice. AB - The very low density lipoprotein receptor (VLDLR) has been proposed to play a role in the delivery of fatty acids to peripheral tissues. However, despite reduced adipose tissue mass in VLDLR-deficient (VLDLR(-)(/-)) mice, this has been difficult to substantiate. In the present study, VLDLR-deficient and VLDLR overexpressing (PVL) mice were cross-bred onto a low density lipoprotein receptor knockout (LDLR(-)(/-)) background to study the VLDLR under conditions of relatively high serum VLDL and triglyceride levels. Absence of the VLDLR resulted in a significant increase in serum triglyceride levels (1.9-fold) when mice were fed a high fat diet. In contrast, overexpression of the VLDLR resulted in a significant decrease in serum triglyceride levels (2.0-fold) under similar conditions. When kept on a chow diet, a period of prolonged fasting revealed a significant increase in serum triglyceride levels in VLDLR(-)(/-); LDLR(-)(/-) mice (2.3-fold) as compared with LDLR(-)(/-) controls. This could not be attributed to altered apolipoprotein B and VLDL triglyceride production rates. Furthermore, no major differences in nascent VLDL triglyceride content were found between VLDLR(-)(/-); LDLR(-)(/-) mice and LDLR(-)(/-) controls. However, the triglyceride content of circulating VLDL of VLDLR(-)(/-); LDLR(-)(/-) mice (63%) was relatively high as compared with LDLR(-)(/-) controls (49%). These observations suggest that the VLDLR affects peripheral uptake of VLDL triglycerides. In conclusion, under conditions of LDLR deficiency in combination with high fat feeding or prolonged fasting, the effect of the VLDLR on VLDL triglyceride metabolism was revealed. PMID- 11108740 TI - Transgenic studies of fatty acid oxidation gene expression in nonobese diabetic mice. AB - Type 1 diabetes mellitus is a devastating disorder affecting both glucose and lipid metabolism. Using the nonobese diabetic (NOD) mouse model, we found that diabetic mice had a liver-specific increase in steady state mRNA levels for enzymes involved in oxidation of fatty acids. Increased mRNA abundance was observed in very long-chain acyl-CoA dehydrogenase, long-chain acyl-CoA dehydrogenase (LCAD), medium-chain acyl-CoA dehydrogenase (MCAD), carnitine palmitoyltransferase I (CPT-1a), and the gluconeogenic enzyme phosphoenolpyruvate carboxykinase, whereas short-chain acyl-CoA dehydrogenase mRNA remained unchanged. In contrast, minimal elevations in LCAD and CPT-1a mRNA were observed in hearts of diabetic mice with no significant differences found for the other enzymes. We developed NOD mice with transgenes containing regulatory elements of human MCAD gene controlling a reporter gene to determine if the increase in MCAD gene expression occurred via the well-characterized nuclear receptor response element (NRRE-1). These results demonstrated that the transgene containing the NRRE-1 and adjacent 5' sequences had elevated liver expression in diabetic mice compared with prediabetic or normal control mice. Surprisingly, the transgene that contains NRRE-1 with adjacent 3' sequences and the transgene with the NRRE-1 deleted showed minimal response to the fulminant diabetic condition.Collectively, these results indicate that in type 1 diabetes there exists an excessive and liver-specific activation of fatty acid oxidation gene expression. Using human MCAD as a prototype gene, we have shown that this increased expression is mediated at the transcriptional level but does not occur via the well characterized NRRE-1 site responsible for baseline expression in normal mice. PMID- 11108741 TI - Epidermal sphingomyelins are precursors for selected stratum corneum ceramides. AB - Epidermal ceramides (Cer) comprise a heterogeneous family of seven species, including two unique omega-hydroxylated Cer, that are key components of the stratum corneum (SC) intercellular lamellar membranes responsible for the epidermal permeability barrier. Although both glucosylceramide (GlcCer) and the phospho-sphingolipid sphingomyelin (SM) are potential precursors of SC Cer, based on reported chemical structures of epidermal GlcCer and SC Cer, it is assumed that all major subfractions of SC Cer are generated from lamellar body-derived GlcCer. Yet, we and others have shown that SM-derived Cer are required for normal barrier homeostasis. Moreover, two pools of SM, one from plasma membrane, the other from lamellar body-derived contents, are potentially available for Cer production. To clarify the role of SM as a potential precursor of bulk or specific SC Cer, we compared Cer moieties in epidermal SM, Cer generated from epidermal SM by sphingomyelinase treatment, Cer within SC, and Cer that persist in Gaucher SC, where GlcCer cannot generate Cer due to an absence of beta glucocerebrosidase. Using gas chromatography-mass spectrometry, fast atom bombardment-mass spectrometry, and nuclear magnetic resonance for Cer characterization, epidermal SM comprise three major subfractions with distinctive amide-linked (N-acyl) fatty acid (FA) compositions: that is, either long-chain FA (SM-1; C(22;-26)), short-chain FA (SM-2; primarily C(16)), and short-chain alpha hydroxy FA (SM-3; C(16;-18)). In contrast, only trace quantities of omega-hydroxy FA were present. For each SM subfraction, the sphingoid base was either sphingosine or sphinganine, but phytosphingosine was not detected. Comparison of these SM with corresponding sphingomyelinase-generated epidermal Cer and SC Cer revealed that the Cer moieties of SM-1 and SM-3 are equivalent to Cer 2 (NS) and Cer 5 (AS), respectively. Moreover, both Cer 2 and Cer 5 occurred in Gaucher SC, whereas other Cer subfractions did not occur. These results indicate that two epidermal SM, that is, SM-1 and SM-3, are important precursors of two corresponding Cer in mammalian SC, that is, Cer 2 and Cer 5, but other Cer species, including the omega-hydroxy Cer species, do not derive from SM. PMID- 11108742 TI - A new sandwich enzyme immunoassay for measurement of plasma pre-beta1-HDL levels. AB - Pre-beta1-HDL, a putative discoid-shaped high density lipoprotein (HDL) of approximately 67-kDa mass that migrates with pre-beta mobility in agarose gel electrophoresis, contains apolipoprotein A-I (apoA-I), phospholipids, and unesterified cholesterol. It participates in the retrieval of cholesterol from peripheral tissues. In this study we established a new sandwich enzyme immunoassay (EIA) for measuring plasma pre-beta1-HDL using mouse anti-human pre beta1-HDL monoclonal antibody (MAb 55201) and goat anti-human apoA-I polyclonal antibody. MAb 55201 reacted with apoA-I in lipoprotein [A-I] with molecular mass less than 67 kDa, and with pre-beta1-HDL separated by nondenaturing two dimensional electrophoresis, whereas it did not react with apoA-I in alpha-HDL. Pre-beta1-HDL levels measured by this method declined when incubated at 37 degrees C for 2 h, whereas this decrease was not observed in the presence of 2 mM lecithin:cholesterol acyltransferase inhibitor 5,5'-dithiobis (2-nitrobenzoic acid). To clarify the clinical significance of measuring pre-beta1-HDL by this method, 47 hyperlipidemic subjects [male/female 22/25; age 55 +/- 14 years; body mass index 25 +/- 4.5 kg/m(2); total cholesterol (TC) 245 +/- 64 mg/dl; triglyceride (TG) 232 +/- 280 mg/dl; HDL cholesterol (HDL-C) 51 +/- 23 mg/dl] and 25 volunteers (male/female 15/10; age 36 +/- 9.3 years; body mass index 23 +/- 3.5 kg/m(2); TC 183 +/- 28 mg/dl; TG 80 +/- 34 mg/dl; HDL-C 62 +/- 15 mg/dl) were involved. Plasma pre-beta1-HDL levels were significantly higher in hyperlipidemic subjects than in volunteers (39.3 +/- 10.1 vs. 22.5 +/- 7.5 mg/ml, P < 0.001) whereas plasma apoA-I levels did not differ (144.2 +/- 28.4 vs. 145.3 +/- 16.3 mg/dl). These results indicate that this sandwich EIA method specifically recognizes apoA-I associated with pre-beta1-HDL. PMID- 11108743 TI - Methanolysis of sphingomyelin. Toward an epimerization-free methodology for the preparation of D-erythro-sphingosylphosphocholine. AB - It is well known that acid hydrolysis of natural sphingomyelin in aqueous methanol or 1-butanol at refluxing temperature is accompanied by epimerization at the C-3 position of the long-chain base. An improved procedure for the hydrolysis of commercially available, naturally occurring sphingomyelin is described. Prolonged exposure (3;-4 days) of sphingomyelin to freshly prepared 0.5 M anhydrous methanolic hydrogen chloride (generated by trapping the gas evolved from the reaction of concentrated sulfuric acid with solid sodium chloride in anhydrous methanol) at 50 degrees C resulted in cleavage of the amide side chain. The extent of epimerization of the allylic alcohol stereocenter was quantified by integration of the C-5 signal of the (13)C nuclear magnetic resonance spectrum of lysosphingomyelin. The method described here is superior to the traditional acid hydrolysis methods because it provides the product as a approximately 10:1 ratio of d-erythro/l-threo epimers; in contrast, a ratio of approximately 1. 3:1 was obtained by the previous methods. We also report that use of dichloromethane as a cosolvent with N,N-dimethylformamide in the reaction of lysosphingomyelin with an activated fatty acid reduced the time required for completion of the N-acylation reaction. PMID- 11108744 TI - In vivo evidence of a role for hepatic lipase in human apoB-containing lipoprotein metabolism, independent of its lipolytic activity. AB - Hepatic lipase (HL) is a key player in lipoprotein metabolism by modulating, through its lipolytic activity, the triglyceride (TG) and phospholipid content of apolipoprotein B (apoB)-containing lipoproteins and of high density lipoproteins (HDL), thereby affecting their size and density. A new and separate role has been suggested for HL in cellular lipoprotein metabolism, in which it serves as a ligand promoting cellular uptake of apoB-containing remnant lipoproteins and HDL. We tested the hypothesis that HL has both a lipolytic and a nonlipolytic role in human lipoprotein metabolism, by measuring lipid plasma concentrations, lipoprotein density distribution by density gradient ultracentrifugation, and lipoprotein composition, in three subjects with HL deficiency: two of the patients (S-1 and S-3) were characterized as having neither plasma HL activity nor detectable HL protein; the third subject (S-2) had no plasma HL activity but a detectable amount (35.5 ng/ml) of HL protein. All HL-deficient subjects showed a severalfold increase in lipoprotein TG content across the lipoprotein density spectrum [very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), low density lipoprotein (LDL), and HDL] as compared with control subjects. They also had remarkably more buoyant LDL particles (LDL-R(f) = 0.342;-0.394) as compared with the control subjects (LDL-R(f) = 0.303). Subjects S-1 and S-3 (no HL activity or protein) presented with a distinct increase in cholesterol and apoB levels in the IDL and VLDL density range as compared with patient S-2, with detectable HL protein, and the control subjects. This study provides evidence in humans that HL indeed plays an important role in lipoprotein metabolism independent of its enzymatic activity: in particular, inactive HL protein appears to affect VLDL and IDL particle concentration, whereas HL enzymatic activity seems to influence VLDL-, IDL-, LDL-, and HDL-TG content and their physical properties. PMID- 11108745 TI - Dynamic cerebral autoregulation is unaffected by aging. AB - BACKGROUND AND PURPOSE: Normal aging is associated with marked changes in the cardiovascular and cerebrovascular systems. Although cerebral autoregulation (CA) is impaired in certain disease states, the effect of age per se on dynamic CA in humans is unknown and the focus of this study. METHODS: Twenty-seven young subjects (/=55 years), matched for sex and systolic blood pressure (BP), underwent measurement of cerebral blood flow velocity by transcranial Doppler ultrasound and noninvasive beat-to-beat arterial BP measurement during induced and spontaneous dynamic BP stimuli. A standard dynamic autoregulatory index (ARI) was derived for each spontaneous and induced dynamic BP stimulus to include the step response, as well as cardiac baroreceptor sensitivity (BRS), for the 2 groups. RESULTS: The mean age of the young group was 29+/-5 years, and that of the older group was 68+/-5 years. Cardiac BRS was reduced in the older group (8. 6+/-4.5 versus 16.9+/-8.8 ms/mm Hg; P:<0.0001). However, no age-related differences were demonstrated in step response plots or in ARI values for any pressor or depressor dynamic BP stimulus (P:=0. 62), with mean ARI values for all stimuli combined being 4.9+/-1.8 for the young group and 5.0+/-2.3 for the older group. CONCLUSIONS: Although increasing age is associated with a decrease in cardiac BRS, dynamic CA, as assessed by step response analysis as well as cerebral blood flow responses to transient and induced BP stimuli, is unaffected by aging. PMID- 11108746 TI - Failure to demonstrate peri-infarct depolarizations by repetitive MR diffusion imaging in acute human stroke. AB - BACKGROUND AND PURPOSE: Peri-infarct depolarizations (PIDs) have been demonstrated with diffusion-weighted MRI (DWI) in experimental stroke and are regarded as an important mechanism of ischemic injury. We tested the hypothesis that PIDs are of relevance for the early enlargement of human brain infarcts. METHODS: Ten stroke patients were investigated by repetitive imaging of the apparent diffusion coefficient (ADC) in the acute phase (7 patients) or subacute phase (3 patients) of developing cortical infarction. In each patient, 20 ADC maps were obtained from serially measured echo-planar DWI (interval of 45 seconds). Data analysis focused on the potential spatial and temporal ADC changes, including structured qualitative analysis, calculation of subtraction images, serial analysis of regions of interest positioned in the infarct core and border, and calculation of hemispheric lesion areas, depending on various ADC thresholds ranging between 0 and 800 microm(2)/s. RESULTS: Data analysis was unable to disclose any time-dependent changes in ADC that would resemble PID. In ischemic regions, the ADC reduction significantly progressed from the infarct border (555+/-96 microm(2)/s) to the infarct core (431+/-104 microm(2)/s, P:<0.01). CONCLUSIONS: By using an MRI protocol with high temporal resolution and elaborated postprocessing, we were unable to demonstrate a pattern of diffusion changes that would be indicative of PID in human stroke. Experimental infarction and human stroke may differ in the detectability of PID. PMID- 11108747 TI - Impairment of cerebrovascular reactivity by methionine-induced hyperhomocysteinemia and amelioration by quinapril treatment. AB - BACKGROUND AND PURPOSE: Human studies have shown that methionine-induced hyperhomocysteinemia impairs brachial artery endothelial function via decreasing nitric oxide activity. However, the effect of homocysteine on cerebrovascular reactivity (CVR), which has been reported to be nitric oxide related in experimental and animal studies, remains unclear in humans. Inhibition of angiotensin-converting enzyme may improve nitric oxide-mediated cerebral as well as peripheral endothelial function. The aim of the present study was to investigate the effect of methionine-induced hyperhomocysteinemia on CVR before and after treatment with quinapril, an angiotensin-converting enzyme inhibitor, in healthy adults. METHODS: Plasma homocysteine and CVR were measured at baseline and 4 hours after methionine load (0.1 g/kg body wt) before and after quinapril treatment (10 mg/d for 1 week) in both younger and older groups. CVR was assessed by transcranial Doppler ultrasonography, measuring the percent increase of flow velocity in the middle cerebral artery after brief carotid compression (expressed as transient hyperemic response ratio [THRR]). RESULTS: Homocysteine levels were significantly increased after methionine load either before or after quinapril treatment in both groups. Before quinapril treatment, postmethionine THRR was preserved in younger adults (24.2+/-5.3% versus 23.8+/-6.3% at baseline, P:=0.73) and decreased in older adults (12.9+/-2.2% versus 21.8+/-4.0% at baseline, P:<0.001). After quinapril treatment, postmethionine THRR was preserved in both groups (24.5+/-5.9% versus 24.0+/-5.0% at baseline, P:=0.42 in younger adults; 20.4+/-3.9% versus 21.3+/-3.3% at baseline, P:=0.35 in older adults). CONCLUSIONS: Our study suggests that methionine-induced hyperhomocysteinemia may be causally associated with impairment of CVR in older normal subjects. PMID- 11108748 TI - Effect of intravenous recombinant tissue plasminogen activator on ischemic stroke lesion size measured by computed tomography. NINDS; The National Institute of Neurological Disorders and Stroke (NINDS) rt-PA Stroke Study Group. AB - Background and Purpose-When given within 3 hours of symptom onset, recombinant tissue plasminogen activator (rtPA) improves outcome 3 months after ischemic stroke. Prespecified secondary end points of the National Institute of Neurological Disorders and Stroke (NINDS) rt-PA Stroke Trial were CT lesion volumes in the 2 treatment groups (tPA and placebo) at 24 hours, 7 to 10 days, and 3 months after stroke. METHODS: -The trial included 2 independent studies, part I and part II, with identical methods of data collection. Before part I, uniform standards were established for CT scanning. CT images were obtained at baseline, 24 hours, 7 to 10 days, and 3 months after stroke onset and were reviewed centrally by reviewers blinded to treatment group and clinical findings. Since the individual studies were not powered to test for lesion volume differences, data from both parts of the trial were combined for all analyses. The primary analysis was conducted with the use of an intention-to-treat algorithm (including patients who died or were lost to follow-up). Measured lesion volume (excluding deaths and those lost to follow-up) was used as a secondary outcome in an exploratory analysis. RESULTS: -After tPA treatment, there was a trend toward a reduction in 3-month median lesion volume in the tPA group: 15 cm(3) (interquartile range, 2 to 87) compared with 24 cm(3) (interquartile range, 4 to 101) in the placebo group (P:=0.06, log model) with a reduction of 11% in cumulative lesion volume, computed with Smirnov's D statistic. After exclusion of deaths and those lost to follow-up, similar trends toward positive treatment effects were seen at all time points. CONCLUSIONS: -The direction of the effect of tPA on CT lesion volume at all time points was consistent with the observed clinical effects at 3 months. CT lesion volume may not be as sensitive a measure of treatment effect as clinical evaluation, at least as used in this study. An intention-to-treat analysis for the radiographic end point in this acute ischemic stroke clinical trial is a less biased approach to account for missing radiographic data than an analysis that uses only measured radiological data. PMID- 11108749 TI - Intravenous tissue plasminogen activator for acute ischemic stroke: A Canadian hospital's experience. AB - BACKGROUND AND PURPOSE: In the United States, tissue plasminogen activator (tPA) was approved for treatment of acute ischemic stroke in 1996. Its use has only recently been approved in Canada. We sought to evaluate the safety, feasibility, and efficacy of treatment in a Canadian hospital setting. METHODS: A combined retrospective and prospective review is presented of 46 consecutive patients treated with intravenous tPA at our hospital with a treatment protocol similar to that of the National Institute of Neurological Disorders and Stroke (NINDS) trial. RESULTS: Symptomatic intracranial hemorrhage at 36 hours occurred in 1 patient (2.2%). The median time to treat was 165 minutes, with a median "door-to needle" time of 84 minutes. Compared with patients presenting initially at our hospital, patients transferred from another institution for tPA therapy were treated closer to the 3-hour time window (mean 173 versus 148 minutes, P:<0.001) but had a shorter door-to-needle time (43 versus 102 minutes, P:<0.001). For every 10 minutes closer to the 3-hour time window that any patient arrived at the hospital, 7 minutes was saved in the door-to-needle time (correlation coefficient 0.9, P:<0.001). Patient outcome did not differ from that in the NINDS trial (P:>0.75). CONCLUSIONS: Our safety and patient outcome data compare favorably with NINDS and Phase IV data. Although a 3-hour treatment window was feasible, the median door-to-needle time lengthened as more treatment time was available and the door-to-needle time was beyond recommended standards. This review has prompted changes in our community to improve treatment efficiency. PMID- 11108750 TI - Cardiac arrhythmias and stroke: increased risk in men with high frequency of atrial ectopic beats. AB - BACKGROUND AND PURPOSE: With the exception of atrial fibrillation (AF), little scientific attention has been given the associations between cardiac arrhythmias and incidence of stroke. We sought to study whether atrial and ventricular arrhythmias assessed during a 24-hour ambulatory ECG registration are associated with incidence of stroke. METHODS: The population-based cohort "Men Born in 1914" was examined with 24-hour ambulatory ECG registrations at 68 years of age. Four hundred two men without previous myocardial infarction or stroke were included, and 236 of them had hypertension (>/=160/95 mm Hg or treatment). Fourteen-year rates of stroke (fatal and nonfatal) and all-cause mortality were updated from national and regional registers. Frequent or complex ventricular arrhythmias was defined as Lown class 2 to 5. A high frequency of atrial ectopic beats (AEB) was defined as the fifth quintile (ie, >/=218 AEB per 24 hours). RESULTS: Fifty-eight men suffered a first stroke during the follow-up. Stroke rates (per 1000 person years) among men with AF (n=14), with frequent AEB (n=77), and without AF or frequent AEB (n=311) were 34.5, 19.5, and 11.6, respectively. The corresponding values among men with hypertension were 40.7, 32.3, and 14.7, respectively. Frequent AEB (compared with absence of AF and frequent AEB) was significantly associated with stroke among all men (relative risk=1.9; 95% CI, 1.02 to 3.4; P:=0.04) and among hypertensive men (relative risk=2.5; 95% CI, 1.3 to 4.8; P:=0.009) after adjustments for potential confounders. The increased stroke rates among men with Lown class 2 to 5 did not reach statistical significance. CONCLUSIONS: A high frequency of AEB is associated with an increased incidence of stroke. PMID- 11108751 TI - Chlamydia pneumoniae does not influence atherosclerotic plaque behavior in patients with established carotid artery stenosis. AB - BACKGROUND AND PURPOSE: Research for infectious agents in the etiology of atherosclerosis has identified Chlamydia pneumoniae as a possible candidate. While there is evidence of an association between presence of this microorganism and atherosclerosis, it is unclear whether infection has a genuinely etiologic role in this disease, whether its presence influences clinical outcomes, and, if so, at which stages of disease this occurs. We have approached this issue in patients with advanced carotid artery atherosclerosis using molecular biological detection methods and clinically relevant indicators of pathology in carotid artery atheroma to determine whether the presence of C pneumoniae correlates with plaque instability. METHODS: C pneumoniae was detected with the use of a sensitive nested polymerase chain reaction. Preoperative embolization and preoperative infarcts were recorded with the use of transcranial Doppler insonation of the middle cerebral artery and cerebral CT, respectively. RESULTS: C pneumoniae DNA was detected in 25.5% of a cohort of 98 symptomatic patients. There was no significant difference in plaque stability as measured by embolization rates between the chlamydial-positive and -negative specimens. There was also no correlation between the number of ipsilateral hemispheric infarcts in the territory of the middle cerebral artery and chlamydial status. CONCLUSIONS: This study confirms that C pneumoniae is a common finding in atherosclerotic plaques of the carotid artery but suggests that the presence of the infectious organism has little detectable impact on plaque instability when measured by clinically significant markers. This raises important questions for the rationale of antibiotic therapy in atherosclerosis. PMID- 11108752 TI - High proinsulin levels precede first-ever stroke in a nondiabetic population. AB - BACKGROUND AND PURPOSE: Diabetic subjects have a 3- to 6-fold increased risk for stroke compared with nondiabetic subjects, and hyperinsulinemia shows strong and consistent associations with a cluster of cardiovascular risk factors. Methods separating proinsulin from (true) insulin have demonstrated proinsulin to be more strongly associated with cardiovascular disease than insulin. The present study evaluates the associations between first-ever stroke, proinsulin, and insulin. METHODS: In this incident case-referent study of a nondiabetic population, 94 cases of first-ever stroke (59 men and 35 women) were individually age- and sex matched to 178 referents. Blood sampling was collected before the stroke event. Proinsulin and insulin were measured with highly sensitive 2-site sandwich enzyme linked immunosorbent assays. RESULTS: In the study population, high proinsulin concentration more than tripled the risk for first-ever stroke after adjustments for total cholesterol, systolic blood pressure, smoking, body mass index, and insulin, with an odds ratio of 3.4 (95% CI, 1.4 to 8.4). In women the risk was even more pronounced, with an odds ratio of 13.7 (95% CI, 1.3 to 146). Synergy was found between proinsulin and systolic blood pressure. In women, synergy was also found between proinsulin and diastolic blood pressure as well as between insulin and both blood pressures. CONCLUSIONS: High levels of proinsulin may predict later occurrence of first-ever stroke in a nondiabetic population. PMID- 11108753 TI - The quest for early predictors of stroke evolution: can TCD be a guiding light? AB - BACKGROUND AND PURPOSE: The present study aimed at evaluating the prognostic value of transcranial Doppler ultrasonography (TCD) in the acute phase of ischemic stroke, when major therapeutic decisions must be made. METHODS: Seventy three patients with a first-ever ischemic hemispheric stroke underwent neurological assessment according to the Unified Neurological Stroke Scale, clinical subgrouping according to the criteria of Bamford, CT scan, cervical duplex sonography, and TCD, all within 12 hours from stroke onset. TCD was repeated on days 2 and 7. Patients were followed for 90 days, during which we calculated the fatality rate and then assessed clinical outcome. RESULTS: Emergency TCD revealed middle cerebral artery (MCA) no-flow in 24 cases and MCA asymmetry in 30 subjects. Serial TCD showed early (<24 hours) MCA recanalization in 6 patients. After 90 days, no patient with MCA occlusion at admission was autonomous, while 17 of 19 patients (89.5%) with a normal baseline TCD were independent. The fatality rate at 3 months was 21% but was 46% in patients with MCA occlusion and 61% in patients without signs of early MCA recanalization. Total anterior circulation infarct and abnormal TCD were significantly correlated (P:<0.001) with higher mortality rate and worse outcome (Barthel Index score /=90 mm Hg and/or SBP >/=140 mm Hg and/or were taking antihypertensive medication; and the moderate group consisted of all other subjects. RESULTS: The outer and inner CCA diameters for the high group were significantly (P:<0.05) enlarged in comparison with those for the moderate and normal groups in all age/sex groups of both sexes after adjustment for body mass index, pack-years of smoking, alcohol consumption, and total serum cholesterol. Multiple regression analysis showed that age, body mass index, SBP, pack-years of smoking, alcohol consumption, and IMT were positively and significantly (P:<0.005) related to both outer and inner CCA diameters in both sexes except for between alcohol consumption and outer CCA diameter in women and showed that only serum total cholesterol level was negatively and significantly (P:<0.01) related to inner CCA diameter in both sexes. CONCLUSIONS: Our present study showed that the outer and inner CCA diameters correlated with conventional cardiovascular risk factors, including high blood pressure and IMT. These findings suggest that the outer and inner CCA diameters may be a useful indicator of carotid atherosclerosis, particularly in relation to high blood pressure. PMID- 11108757 TI - Anticoagulant patient information material is written at high readability levels. AB - BACKGROUND: Warfarin therapy requires frequent monitoring and dose adjustment. Elderly patients with atrial fibrillation, prior stroke, and lower literacy skills may have difficulty reading brochures that explain dosing instructions, procedures to follow, and the risks and benefits of anticoagulants. In general, it is recommended that brochures be written at or below the 6th-grade level. We determined the readability of patient information material being offered to patients receiving anticoagulants. METHODS AND RESULTS: We used the SMOG grade formula to measure readability of written patient materials. We obtained 50 brochures commonly used in anticoagulation management units from industry and health advocacy groups. Patient information was related to atrial fibrillation (16%, n=8), warfarin (44%, n=22), low-molecular-weight heparins (12%, n=6), or other related topics (28%, n=14). The mean readability was found to be grade 10.7 (95% CI 10.1 to 11.2); none had a readability score at the 6th-grade level or below, 12% of the brochures had readability scores at the 7th- to 8th-grade levels (n=6), 74% at the 9th- to 12th-grade levels (n=37), and 14% at higher than 12th-grade level (n=7). The readability grade level was similar for brochures produced by industry or health advocacy groups (P:=0.9) but higher for information obtained from the Internet (12.2+/-1.3 grades) compared with other sources (10.3+/-2.1 grades; P:=0.01). CONCLUSIONS: Patient education materials related to the use of anticoagulants are written at grade levels beyond the comprehension of most patients. Low-literacy brochures are needed for patients on anticoagulants. PMID- 11108758 TI - Endothelin-1 in subarachnoid hemorrhage: An acute-phase reactant produced by cerebrospinal fluid leukocytes. AB - BACKGROUND AND PURPOSE: The most potent vasoconstrictor known, endothelin-1, is currently considered to mediate cerebral vasospasm in subarachnoid hemorrhage (SAH), which can cause delayed cerebral ischemia. In our study, we performed clinical and in vitro experiments to investigate the origin and the mechanisms of the secretion of endothelin-1 in SAH. METHODS: Endothelin-1 and markers of inflammatory host response (interleukin [IL]-1ss, IL-6, and tumor necrosis factor alpha) were comparatively quantified in the cerebrospinal fluid (CSF) of SAH patients and control subjects, and concentrations were related to clinical characteristics. Furthermore, mononuclear leukocytes isolated from the CSF of SAH patients and control subjects were analyzed regarding their mRNA expression of endothelin-1 and inflammatory cytokines. Finally, complementary in vitro experiments were performed to investigate whether coincubation of blood and CSF can trigger leukocytic mRNA expression and release of these factors. RESULTS: Activated mononuclear leukocytes in the CSF of SAH patients synthesize and release endothelin-1 in parallel with known acute-phase reactants (IL-1ss, IL-6, and tumor necrosis factor-alpha). Complementary in vitro experiments not only further confirmed this leukocytic origin of endothelin-1 but also showed that aging and subsequent hemolysis of blood is sufficient to induce such endothelin-1 production. CONCLUSIONS: The demonstration that endothelin-1 is produced by activated CSF mononuclear leukocytes suggests that subarachnoid inflammation may represent a therapeutic target to prevent vasospasm and delayed cerebral ischemia after SAH. PMID- 11108759 TI - Perimesencephalic hemorrhage and CT angiography: A decision analysis. AB - BACKGROUND AND PURPOSE: The method of choice for detecting or excluding a vertebrobasilar aneurysm still is a matter of debate in patients with a characteristically perimesencephalic pattern of subarachnoid hemorrhage (SAH) on CT. We used decision analysis to compare possible diagnostic strategies in these patients. METHODS: A decision analytic model was developed to evaluate the effect of 4 different diagnostic strategies following a perimesencephalic pattern of SAH on CT: 1, no further investigation; 2, digital subtraction angiography (DSA) by catheter; 3, CT angiography as initial modality, not followed by DSA if negative; and 4, CT angiography as initial modality, followed by DSA. We used a 4% prevalence of a vertebrobasilar aneurysm given a perimesencephalic pattern of hemorrhage, a 97% sensitivity and specificity of CT angiography, and a 99.5% sensitivity and 100% specificity of DSA. In a prospectively collected series, the complication rate from DSA in patients with a perimesencephalic pattern of hemorrhage was 2.6%. We calculated the expected utility of each of the 4 diagnostic options and used sensitivity analyses to examine the influence of the plausible ranges of the various estimates used. RESULTS: The expected utilities were 99.09 for CT angiography only, 98.96 for no further investigation, 98.22 for DSA, and 96.34 for CT angiography plus DSA. The results of the sensitivity analysis indicate that over a wide range of assumptions, CT angiography only is the most beneficial option. Only when the complication rate of catheter angiography is <0.2% is DSA the preferred strategy. CONCLUSIONS: Our decision analysis shows that in patients with a perimesencephalic pattern of hemorrhage on CT, CT angiography only is the best diagnostic strategy. DSA can be omitted in patients with a perimesencephalic pattern of hemorrhage and a negative CT angiogram. PMID- 11108760 TI - Does the application of constraint-induced movement therapy during acute rehabilitation reduce arm impairment after ischemic stroke? AB - BACKGROUND AND PURPOSE: Motor dysfunction after unilateral deafferentation in primates can be overcome by restraining the unaffected limb. We asked whether a constraint-induced movement (CIM) program could be implemented within 2 weeks after stroke and whether CIM is more effective than traditional upper-extremity (UE) therapies during this period. METHODS: Twenty-three persons were enrolled in a pilot randomized, controlled trial that compared CIM with traditional therapies. A blinded observer rated the primary end point, the Action Research Arm Test (ARA). Inclusion criteria were the following: ischemic stroke within 14 days, persistent hemiparesis, evidence of preserved cognitive function, and presence of a protective motor response. Differences between the groups were compared by using Student's t tests, ANCOVA, and Mann-Whitney U: tests. RESULTS: Twenty subjects completed the 14-day treatment. Two adverse outcomes, a recurrent stroke and a death, occurred in the traditional group; 1 CIM subject met rehabilitation goals and was discharged before completing 14 inpatient days. The CIM treatment group had significantly higher scores on total ARA and pinch subscale scores (P:<0.05). Differences in the mean ARA grip, grasp, and gross movement subscale scores did not reach statistical significance. UE activities of daily living performance was not significantly different between groups, and no subject withdrew because of pain or frustration. CONCLUSIONS: A clinical trial of CIM therapy during acute rehabilitation is feasible. CIM was associated with less arm impairment at the end of treatment. Long-term studies are needed to determine whether CIM early after stroke is superior to traditional therapies. PMID- 11108761 TI - Benefit of an extended stroke unit service with early supported discharge: A randomized, controlled trial. AB - BACKGROUND AND PURPOSE: Several trials have shown that stroke unit care improves outcome for stroke patients. The aim of the present trial was to evaluate the effects of an extended stroke unit service (ESUS), with early supported discharge, cooperation with the primary healthcare system, and more emphasis on rehabilitation at home as essential elements. METHODS: In a randomized, controlled trial, 160 patients with acute stroke were allocated to the ESUS and 160 to the ordinary stroke unit service (OSUS). The primary outcome was the proportion of patients who were independent as assessed by the modified Rankin Scale (RS) (RS /=95=independent in ADL) after 26 weeks. Secondary outcomes were RS and BI scores after 6 weeks; the proportion of patients at home, in institutions, and deceased after 6 and 26 weeks; and the length of stay in institutions. RESULTS: After 26 weeks, 65.0% in the ESUS versus 51.9% in the OSUS group showed global independence (RS /=95) (P:=0.056). The odds ratios for independence (ESUS versus OSUS) were as follows: RS, 1.72 (95% CI, 1.10 to 2.70); BI, 1.54 (95% CI, 0.99 to 2.39). At 6 weeks, 54.4% of the ESUS group and 45. 6% of the OSUS group were independent according to RS (P:=0.118), and 56.3% versus 48.8% were independent according to BI (P:=0.179). The proportion of patients at home after 6 weeks was 74.4% for ESUS and 55.6% for OSUS (P:=0.0004), and the proportion in institutions was 23.1% versus 40.0%, respectively (P:=0.001). After 26 weeks, 78. 8% in the ESUS group versus 73.1% in the OSUS were at home (P:=0. 239), while 13.1% versus 17.5% were in institutions (P:=0.277). The mortality in the 2 groups did not differ. Average lengths of stay in an institution were 18.6 days in the ESUS and 31.1 days in the OSUS group (P:=0.0324). CONCLUSIONS: An ESUS with early supported discharge seems to improve functional outcome and to reduce the length of stay in institutions compared with traditional stroke unit care. PMID- 11108762 TI - Quality of life among stroke survivors evaluated 1 year after stroke: experience of a stroke unit. AB - BACKGROUND AND PURPOSE: We sought to study overall and domain-specific quality of life in stroke survivors 1 year after stroke and to identify variables that could predict quality of life after stroke. METHODS: We followed up for 1 year a cohort of 118 patients consecutively admitted to our stroke unit at San Carlos University Hospital in Madrid, Spain. The final series at 1-year follow-up consisted of 90 survivors (41 women and 49 men; mean age, 68 years; range, 32 to 90 years). A cross-sectional, descriptive design was developed. Patients completed a questionnaire that included socioeconomic variables, Hamilton Rating Scale for Depression, Sickness Impact Profile (SIP), Short Form 36, Frenchay Index, Barthel Index, Rankin Scale, and Scandinavian Stroke Scale. Independent variables were sex, age, functional status, motor impairment, and depression. We developed an ANOVA model for statistical analysis. RESULTS: We interviewed 79 patients with ischemic and 11 with hemorrhagic stroke. Thirty-eight percent of patients scored in the depressed range. Variables related to depression were status as a housewife, female sex, inability to work because of disability, and diminished social activity (P:<0.0001). Mean total SIP (24.3), SIP psychosocial dimension (27.5), and SIP physical dimension (21.2) were correlated with disability, female sex, motor impairment, and depression (P:<0.0001). CONCLUSIONS: Functional status and depression were identified as predictors of quality of life. Patients independent in their activities of daily living suffered from a deterioration of the psychosocial dimension of the SIP. PMID- 11108763 TI - Magnetic resonance techniques for the identification of patients with symptomatic carotid artery occlusion at high risk of cerebral ischemic events. AB - BACKGROUND AND PURPOSE: We sought to assess whether MRI, MR angiography, or (1)H MR spectroscopy can be used to identify patients with symptomatic carotid artery occlusion (CAO) who are at high risk of recurrent ipsilateral cerebral ischemic events. METHODS: In 115 consecutive patients with transient or moderately disabling symptoms of cerebral or retinal ischemia and ipsilateral CAO, we studied the prognostic value of (1) presence of a border-zone infarct; (2) quantitative flow in the middle cerebral artery (MCA) ipsilateral to the CAO; and (3) metabolic ratios in the centrum semiovale ipsilateral to the CAO. RESULTS: Presence of a border-zone infarct and the rate of flow in the MCA did not have a significant relationship with recurrence of cerebral ischemic events. Patients with a low N:-acetyl aspartate (NAA)/choline ratio had an annual risk of recurrent, ipsilateral, cerebral ischemic events of 16.0% (95% CI, 9.5 to 27.0), whereas this risk was 4.2% (95% CI, 2.2 to 8.0) in those with a normal NAA/choline ratio (hazard ratio, 0. 43; 95% CI, 0.19 to 1.00). Patients who on entry had had only retinal symptoms had on average a higher NAA/choline ratio (mean difference, 0.25; 95% CI, 0.13 to 0.37) and a lower risk of recurrent cerebral ischemic events (odds ratio, 0.0; 95% CI, 0.0 to 0.6) than those with cerebral ischemic symptoms. CONCLUSIONS: NAA/choline ratio measured by (1)H MRS, but not the presence of a border-zone infarct or the amount of flow in the MCA, can identify patients with symptomatic CAO who are at risk of future ipsilateral cerebral ischemic events. PMID- 11108764 TI - N-acetylaspartate distribution in proton spectroscopic images of ischemic stroke: relationship to infarct appearance on T2-weighted magnetic resonance imaging. AB - BACKGROUND AND PURPOSE: It is generally considered that tissue that appears abnormal on T2 MRI is already infarcted and that any penumbra lies outside the T2 visible lesion. We investigated the distribution of infarcted tissue using proton spectroscopic MRI. METHODS: In patients with symptoms of acute hemispheric ischemic stroke, imaged within a maximum of 3 days of stroke, we explored the distribution of N:-acetylaspartate (NAA), a marker of intact neurons, within and around the abnormal (hyperintense) areas on T2-weighted MR images, using proton spectroscopic MRI. RESULTS: In 11 patients, imaged 24 to 72 hours after stroke onset, there was little evidence of damaged neurons (reduced NAA) beyond the margins of hyperintensity on the T2 image. However, within the abnormal T2 area, there were statistically significant differences in the amount of NAA (ie, the proportion of intact neurons) between areas that were obviously abnormal on T2 (very hyperintense) and those that were only slightly abnormal (slightly hyperintense). CONCLUSIONS: The extent and degree of hyperintensity of the T2 visible lesion directly reflect the amount of neuronal damage; lack of a T2 visible lesion would suggest predominantly intact neurons at the time of imaging. We hypothesize that once tissue damage has reached a critical (probably irreversible) level, the T2 image quickly becomes abnormal without any significant time lag between the pathological staging of the infarct and its visualization on T2. Further testing in a larger study with information on blood flow levels would be required to confirm this. PMID- 11108765 TI - Serum ferritin and C282Y mutation of the hemochromatosis gene as predictors of asymptomatic carotid atherosclerosis in a community population. AB - BACKGROUND AND PURPOSE: Serum ferritin and heterozygosity for the C282Y mutation of the hemochromatosis gene have both been associated with an increased risk of cardiovascular events. The purpose of the study was to test whether either is a risk predictor for asymptomatic carotid atherosclerosis. METHODS: We assessed carotid intima-media wall thickness (IMT) and focal plaque formation by high resolution B-mode ultrasound, conventional risk factors, serum ferritin levels, and the C282Y mutation of the hemochromatosis gene in a randomly selected community population of 1098 subjects (545 women and 553 men) aged 27 to 77 years. RESULTS: After adjustment for conventional risk factors, serum ferritin was not associated with carotid mean IMT. Women with ferritin values over the first quartile (>34 microg/L) had an adjusted odds ratio of 2.1 (95% CI, 1. 3 to 3.4; P:=0.0016) for carotid plaque compared with the first quartile. Ferritin was not associated with carotid plaque in men. Subjects who were heterozygous for the C282Y mutation constituted 11. 4% of the population, and there was no independent association of this genotype with either carotid IMT or focal plaque formation. CONCLUSIONS: We conclude that in our community population, C282Y genotype status was not a risk predictor for either carotid mean IMT or plaque formation. Serum ferritin values in women were independently associated with carotid plaque. PMID- 11108766 TI - Preoperative MRA flow quantification in CEA patients: flow differences between patients who develop cerebral ischemia and patients who do not develop cerebral ischemia during cross-clamping of the carotid artery. AB - BACKGROUND AND PURPOSE: We sought to investigate whether preoperative volume flow in the internal carotid arteries (ICAs), the basilar artery (BA), and the middle cerebral arteries (MCAs) and collateral flow via the circle of Willis differ between patients who do and patients who do not develop cerebral ischemia during clamping of the carotid artery in carotid endarterectomy (CEA). METHODS: Quantitative volume flow in the ICAs, BA, and MCAs and directional flow in the circle of Willis were measured preoperatively with 2-dimensional phase-contrast MR angiography in 86 CEA patients. During the operation, electroencephalographic (EEG) recordings were obtained that were monitored by a clinical neurophysiologist. Reference volume flow values were assessed in 24 control subjects. RESULTS: In patients with an ICA stenosis without contralateral ICA occlusion (n=62), of whom 16% developed ischemic EEG changes during clamping, preoperative flow in the clamped ICA was significantly higher in patients with cerebral ischemia than in patients without (mean, 278 versus 160 mL/min; P:<0.05). Flow in the contralateral ICA (156 versus 273 mL/min; P:<0.01), flow in the BA (116 versus 165 mL/min; P:<0.05), and presence of collateral flow via the circle of Willis to the clamped ICA (0% versus 37%; P:<0.05) were significantly lower. MCA flow did not differ significantly between groups. Additionally, in patients with an ICA stenosis and a contralateral ICA occlusion (n=24), of whom 42% developed cerebral ischemia, preoperative flow in the clamped ICA was significantly higher in patients with cerebral ischemia than in patients without (309 versus 239 mL/min; P:<0.05). BA flow, MCA flow, and presence of willisian collateral flow (0% versus 14%) did not differ significantly between groups. CONCLUSIONS: Preoperative volume flow in the clamped ICA is significantly higher in CEA patients with ischemic EEG changes during clamping than in CEA patients without such changes. The latter patients probably have better developed collateral pathways preoperatively. PMID- 11108767 TI - Stent angioplasty for cervical carotid artery stenosis in high-risk symptomatic NASCET-ineligible patients. AB - BACKGROUND AND PURPOSE: Although the North American Symptomatic Carotid Endarterectomy Trial (NASCET) has shown carotid endarterectomy (CEA) to be protective compared with medical therapy alone, its stringent eligibility criteria excluded patients with severe medical, angiographic, and neurological risk factors. We sought to determine the safety and efficacy of stent angioplasty in this high-risk subset for whom the perioperative morbidity and mortality of surgery are elevated. METHODS: Twenty-eight consecutive symptomatic NASCET ineligible patients (10 female; median age, 72.2 years) underwent microcatheter based carotid stent angioplasty. Half of the patients had sustained a previous stroke. Classification of surgical risk by Sundt criteria yielded no patients in grade 1, 3 patients in grade 2 (10.7%), 8 in grade 3 (28.6%), and 17 (60.7%) in grade 4. Stratification of stroke risk for medical therapy according to the European Carotid Surgery Trial (ECST) 5-point score showed 8 patients with a score of 3 (28.6%), 12 with 4 (42.8%), and 8 with 5 (28.6%). Follow-up was obtained in all patients at a median of 14 months. RESULTS: The procedure was technically successful in all cases (100%), with immediate stenosis reduction from a mean of 80.3% to 2.7%. There were no periprocedural deaths, 1 major stroke (3.6%), no minor strokes, and 3 transient ischemic attacks (10.7%). In-hospital complications included 2 nonfatal myocardial infarctions, 1 case of acute renal failure, and 1 groin hematoma requiring transfusion. There were 5 deaths during the follow-up period, all beyond 30 days after the procedure: 3 from cardiac causes, 1 from lung cancer, and 1 following unrelated surgery. The patient with major stroke died at 7.8 months during rehabilitation. No surviving patients had further strokes, and all except 1 (95.5%) remained asymptomatic. Anatomic follow up in 20 patients showed occlusion in 2 (10%) (1 symptomatic, 1 asymptomatic) and intimal hyperplasia in 3 asymptomatic patients (15%). CONCLUSIONS: The clinical results and sustained freedom from symptoms and stroke during the short available follow-up period suggest that stent angioplasty may be useful in the treatment of symptomatic cervical carotid stenosis in high-risk patients despite a notable incidence of restenosis. PMID- 11108768 TI - Metalloproteinase inhibition reduces thrombolytic (tissue plasminogen activator) induced hemorrhage after thromboembolic stroke. AB - BACKGROUND AND PURPOSE: A potentially dangerous side effect associated with tissue plasminogen activator (tPA) use is cerebral hemorrhage. We have focused on developing drugs that could be administered with tPA to reduce the rate of hemorrhage. Since recent studies suggest that various matrix metalloproteinases (MMPs) are important in tumor necrosis factor-alpha production and membrane and vessel remodeling after ischemia, we investigated whether MMP inhibition affected the rate of hemorrhage and infarct production in the absence or presence of tPA treatment. METHODS: We occluded the middle cerebral artery of New Zealand White rabbits with radiolabeled blood clots. Five minutes after embolization, we administered either the MMP inhibitor BB-94 (30 mg/kg SC) or its vehicle. Additional groups received BB-94 or vehicle in combination with tPA, administered 60 minutes after embolization (3.3 mg/kg tPA). After 48 hours, the rabbits were killed and brains were removed, immersion fixed for 1 week in 4% paraformaldehyde, and then cut into 5-mm coronal sections that were examined for the presence of hemorrhage, infarcts, and recanalization. RESULTS: Hemorrhage after embolic stroke was detected in 24% of the control group. tPA induced macroscopically visible hemorrhage in 77% of the tPA-treated group. The rabbits treated with BB-94 had an 18% incidence of hemorrhage (P:>0.05 compared with control). However, when the combination of BB-94 and tPA was administered to rabbits, there was only a 41% incidence of hemorrhage (compared with 77% in the tPA group; P:<0. 05). Both tPA and BB-94/tPA were similarly effective at lysing clots, at 49% and 35% (P:<0.05), respectively, compared with the 5% rate of lysis in the control group. There was a trend for a reduction in the number of infarcts, but it did not reach statistical significance. CONCLUSIONS: Our data suggest that MMP inhibition attenuates mechanisms involved in tPA-induced hemorrhage. This novel form of combination therapy may show promise as a treatment strategy for acute stroke. PMID- 11108769 TI - LY353381.HCl, a selective estrogen receptor modulator, and experimental stroke. AB - BACKGROUND AND PURPOSE: The impact of postmenopausal estrogen replacement therapy on stroke prevention and stroke severity remains controversial. Previously we have shown that cerebral tissue infarction volume sustained after middle cerebral artery (MCA) occlusion is smaller in female than in male animals. This protection is lost after ovariectomy but is restored by 17ss-estradiol replacement. However, the therapeutic range for estradiol is suboptimal, since only doses resulting in a narrow range of plasma levels are protective in brain. The present study tested the hypothesis that a benzothiophene analogue and selective estrogen receptor modulator, LY353381.HCl (LY), reduces tissue infarction after MCA occlusion in estrogen-deficient, ovariectomized female rats. METHODS: Ovariectomized female Wistar rats received LY 10 mg/kg (n=16) or an equivalent volume of vehicle (n=14) by gavage for 5 to 8 days. Subsequently, each animal was anesthetized with halothane (1.2%) and treated with 2 hours of MCA occlusion by the intraluminal filament technique and 22 hours of recovery. Infarction volumes in the cerebral cortex and caudoputamen were determined by 2, 3,5-triphenyltetrazolium chloride staining and digital image analysis. End-ischemic regional cerebral blood flow (CBF) was measured in separate animal cohorts by quantitative [(14)C]iodoantipyrine autoradiography. RESULTS: Caudoputamen infarction was reduced by LY treatment (49+/-6% versus 64+/-4% of ipsilateral caudoputamen in LY and vehicle groups, respectively; P:<0.05). Cerebral cortical infarction was not different in the LY compared with vehicle group (7+/-3% versus 13+/-4% of ipsilateral cerebral cortex, respectively). Intra-ischemic blood pressure, arterial blood gases, and temporalis muscle temperature were controlled and equivalent between treatment groups. Averaged laser-Doppler flow during MCA occlusion was 36+/-3% of baseline in the LY group versus 29%+/-2% in the vehicle group. However, end-ischemic CBF or blood flow distribution within the MCA territory was not altered by LY treatment. Cortical or caudoputamen tissue volumes with end-ischemic CBF <20 mL/100 g per minute were similar in both groups. CONCLUSIONS: We conclude that LY confers neuroprotection from focal cerebral ischemia in caudoputamen in ovariectomized female rats. The mechanism of protection is not linked to preservation of ischemic cerebral blood flow, as determined by end-occlusion quantitative autoradiography. PMID- 11108770 TI - Editorial comment PMID- 11108771 TI - Postischemic cerebrovascular E-selectin expression mediates tissue injury in murine stroke. AB - BACKGROUND AND PURPOSE: Although the deleterious role of several proinflammatory mediators, including P-selectin, in reperfused stroke is well established, the role of E-selectin has not been fully characterized. METHODS: E-selectin mRNA expression was studied at 4, 10, and 24 hours after reperfusion with reverse transcription and polymerase chain reaction in mice (n=18) subjected to transient intraluminal middle cerebral artery occlusion (MCAO). Mice received intravenous injection with anti-E-selectin monoclonal antibody (10, 35, or 50 microg), nonimmune IgG, or vehicle immediately before MCAO and 90 minutes later (n=85). Others received anti-E-selectin antibody 3 or 6 hours after MCAO (n=32). Myeloperoxidase activity was measured in sham-operated mice and after 10 hours of reperfusion in saline-, nonimmune IgG-, or anti-E-selectin IgG-treated cohorts (n=17). Serial cerebral blood flow was measured with laser-Doppler flowmetry, and outcomes were assessed by neurological deficits and infarct volumes with the use of planimetric analysis of triphenyltetrazolium chloride-stained sections. RESULTS: Upregulated E-selectin expression occurred in the ischemic cerebral vasculature within 4 hours of reperfusion and persisted for 24 hours. Anti-E selectin antibody increased ischemic cortical cerebral blood flow up to 2.6-fold (P:<0.05). In addition to dose-dependent reductions in neurological deficits (P:<0.05), mortality, and infarct volumes (P:<0.01 for 35 and 50 microg), anti-E selectin treatment reduced cerebral neutrophil accumulation (P:<0.05) and was neuroprotective even if delayed until 3 hours after ischemia (P:<0. 05). CONCLUSIONS: These findings establish a functional role for E-selectin in the pathogenesis of tissue injury after cerebral ischemia and reperfusion and suggest that E-selectin blockade may be clinically useful in the treatment of reperfused stroke. PMID- 11108772 TI - A modified transorbital baboon model of reperfused stroke. AB - BACKGROUND AND PURPOSE: Although pathophysiological studies of focal cerebral ischemia in nonhuman primates can provide important information not obtainable in rodent models, primate experimentation is limited by considerations of cost, availability, effort, and ethics. A reproducible and quantitative model that minimizes the number of animals necessary to detect differences between treatment groups is therefore crucial. METHODS: Eight male baboons (weight, 22+/-2 kg) underwent left transorbital craniectomy followed by 1 hour of temporary ipsilateral internal carotid artery occlusion at the level of the anterior choroidal artery together with bilateral temporary occlusion of both anterior cerebral arteries (A1) proximal to the anterior communicating artery. A tightly controlled nitrous oxide-narcotic anesthetic allowed for intraoperative motor evoked potential confirmation of middle cerebral artery (MCA) territory ischemia. Animals survived to 72 hours or 10 days if successfully self-caring. Outcomes were assessed with a 100-point neurological grading system, and infarct volume was quantified by planimetric analysis of both MRI and triphenyltetrazolium chloride-stained sections. RESULTS: Infarction volumes (on T2-weighted images) were 32+/-7% (mean+/-SEM) of the ipsilateral hemisphere, and neurological scores averaged 29+/-9. All animals demonstrated evidence of hemispheric infarction, with damage evident in both cortical and subcortical regions in the MCA vascular territory. Histologically determined infarction volumes differed by <3% and correlated with absolute neurological scores (r=0.9, P:=0.003). CONCLUSIONS: Transorbital temporary occlusion of the entire anterior cerebral circulation with strict control of physiological parameters can reliably produce reperfused MCA territory infarction. The magnitude of the resultant infarct with little interanimal variability diminishes the potential number of animals required to distinguish between 2 treatment regimens. The anatomic distribution of the infarct and associated functional deficits offer comparability to human hemispheric strokes. PMID- 11108773 TI - Restricted dissociated sensory loss in a patient with a lateral medullary syndrome: A clinical-MRI study. AB - BACKGROUND: Various sensory syndromes in lateral medullary infarctions are described. A small variation in the location of a lesion may lead to very different clinical features, owing to the complex anatomy of the medulla oblongata. MRI may identify the location and extent of the ischemic lesions, allowing a clear clinical-anatomical correlation. CASE DESCRIPTION: We describe a man with an ischemic lesion in the right portion of the lower medulla that presented a contralateral impairment of spinothalamic sensory modalities and an ipsilateral impairment of lemniscal modalities with a restricted distribution (left forearm and hand, right hand and fingers, respectively). The restricted and dissociated sensory abnormalities represent the only permanent neurological consequence of that lesion. CONCLUSIONS: The atypical sensory syndrome may be explained by the involvement of the medial portion of spinothalamic tract and the lateral portion of archiform fibers at the level of the lemniscal decussation. PMID- 11108774 TI - Diagnostic testing for coagulopathies in patients with ischemic stroke. AB - BACKGROUND: Hypercoagulable states are a recognized, albeit uncommon, etiology of ischemic stroke. It is unclear how often the results of specialized coagulation tests affect management. Using data compiled from a systematic review of available studies, we employed quantitative methodology to assess the diagnostic yield of coagulation tests for identification of coagulopathies in ischemic stroke patients. SUMMARY OF REVIEW: We performed a MEDLINE search to identify controlled studies published during 1966-1999 that reported the prevalence of deficiencies of protein C, protein S, antithrombin III, plasminogen, activated protein C resistance (APCR)/factor V Leiden mutation (FVL), anticardiolipin antibodies (ACL), or lupus anticoagulant (LA) in patients with ischemic stroke. The cumulative prevalence rates (pretest probabilities) and positive likelihood ratios for all studies and for those including only patients aged Ala replacement at position 33). Our data identified zYa as a receptor that can bind ligands specific for Y1, Y2, and Y4 receptors, while zYb and zYc were more Y1-like. All peptides with internal deletions bound to the zYa receptor with affinities similar to that of intact pNPY. Neither the Y1-selective antagonists BIBP3226 and SR120819A nor the Y2 selective BIIE0246 bound to any of the zebrafish receptors, although the amino acids identified as important for BIBP3226 binding were almost completely conserved. These results may prove helpful in molecular modeling of the three dimensional receptor structure. PMID- 11108797 TI - Down-regulation of bcr-abl and bcl-x(L) expression in a leukemia cell line and its doxorubicin-resistant variant by topoisomerase II inhibitors. AB - K562 cells are usually resistant to apoptosis induction, probably because of the expression of bcr-abl, the hybrid gene characteristic of the Philadelphia chromosome (t 9;22). However, we have previously shown that amsacrine and, to a lesser extent, doxorubicin, could induce apoptosis in the doxorubicin-resistant variant of this cell line. In order to elucidate the role of bcr-abl in triggering apoptosis, we investigated the effect of the topoisomerase II inhibitors doxorubicin, amsacrine, and etoposide on the expression of several genes that may be related to apoptosis induction in both cell lines. This was done using a technique of reverse transcription-polymerase chain reaction coupled with HPLC of the amplified fragments to obtain semiquantitative evaluations. We showed that amsacrine, at pharmacologically relevant concentrations, was able to decrease the expression of bcr-abl down to 20% of the basal value in the doxorubicin-resistant variant only, whereas doxorubicin and etoposide were unable to do so. No effect of these drugs was seen on the expression of the normal abl gene. In addition, there was an effect of amsacrine on the expression of bcl-x(L) in the resistant cell line only, but at concentrations higher than the IC(50) of this drug. Our results emphasize the role of bcr-abl in protecting cells from apoptosis and the possible involvement of specific topoisomerase II inhibitors in overcoming resistance to apoptosis. PMID- 11108798 TI - Inactivation of human arylamine N-acetyltransferase 1 by the hydroxylamine of p aminobenzoic acid. AB - Human N-acetyltransferase 1 (NAT1) is a widely distributed enzyme that catalyses the acetylation of arylamine and hydrazine drugs as well as several known carcinogens, and so its levels in the body may have toxicological importance with regard to drug toxicity and cancer risk. Recently, we showed that p-aminobenzoic acid (PABA) was able to down-regulate human NAT1 in cultured cells, but the exact mechanism by which PABA acts remains unclear. In the present study, we investigated the possibility that PABA-induced down-regulation involves its metabolism to N-OH-PABA, since N-OH-AAF functions as an irreversible inhibitor of hamster and rat NAT1. We show here that N-OH-PABA irreversibly inactivates human NAT1 both in cultured cells and cell cytosols in a time- and concentration dependent manner. Maximal inactivation in cultured cells occurred within 4 hr of treatment, with a concentration of 30 microM reducing activity by 60 +/- 7%. Dialysis studies showed that inactivation was irreversible, and cofactor (acetyl coenzyme A) but not substrate (PABA) completely protected against inactivation, indicating involvement of the cofactor-binding site. In agreement with these data, kinetic studies revealed a 4-fold increase in cofactor K(m), but no change in substrate K(m) for N-OH-PABA-treated cytosols compared to control. We conclude that N-OH-PABA decreases NAT1 activity by a direct interaction with the enzyme and appears to be a result of covalent modification at the cofactor-binding site. This is in contrast to our findings for PABA, which appears to reduce NAT1 activity by down-regulating the enzyme, leading to a decrease in NAT1 protein content. PMID- 11108799 TI - Down-modulation through protein kinase C-alpha of lipopolysaccharide-induced expression of membrane CD14 in mouse bone marrow granulocytes. AB - We have previously shown that stimulation of mouse bone marrow granulocytes (BMC) by lipopolysaccharide (LPS) induces the expression of CD14. We found here that phorbol 12-myristate 13-acetate (PMA) blocks this LPS effect. The aim of this study was to investigate the mechanism by which PMA can block the LPS signaling pathway in BMC. The unmodified binding of a radiolabeled LPS in PMA-treated cells indicated that the PMA effect was not the consequence of a shedding or an internalization of the LPS receptor, but was rather due to a biochemical event that follows the interaction of LPS with its receptor. The observations that a selective activator of protein kinase C (PKC)-alpha (sapintoxin D) mimics the PMA effect, whereas a selective PKC-alpha inhibitor (Ro-320432) antagonizes this effect, suggest a regulatory role of PKC-alpha in the LPS signaling pathway in mouse BMC. PMID- 11108800 TI - Pharmacodynamic approach to study the gene transfer process employing non-viral vectors. AB - In the present work we set out to apply pharmacodynamic concepts derived from dose-response curves (Potency and Efficacy) to characterize the gene transfer efficiency of a vector:DNA complex. We employed two widely used vectors, the cationic lipid DOTAP (N,N, N-trimethyl 1-2-3-bis (1-oxo-9-octa-decenyl)oxy-(Z, Z) 1-propanaminium methyl sulfate) and the cationic polymer PEI (polyethylenimine, 800 kDa) to transfect several constructions of the green fluorescent protein cDNA. The analysis of dose-response curves indicated that in all cases the goodness-of-fit was > 0.99. Potency is a measure that provides information on gene activity per amount of DNA. Efficacy is a measure of maximum gene expression achievable using a specific vector:DNA complex, and depends on both the intrinsic efficacy of the gene (evaluated using different vectors to transfer the same gene construct) and on vector efficacy in DNA delivery (evaluated using a single vector to deliver different gene constructs). The results suggest that Potency and Efficacy are objective parameters for describing and comparing the goodness of vectors, as well as the intrinsic efficacy of a given gene construct. Furthermore, they are useful tools that may contribute to a better understanding of the mechanistic gene transfer process of each vector. PMID- 11108801 TI - Reversible and potent uncoupling of hog gastric (H(+)+K(+))-ATPase by prodigiosins. AB - Prodigiosin, prodigiosin 25-C, and metacycloprodigiosin all strongly inhibited the acidification activity of (H(+)+K(+))-ATPase on membrane vesicles from hog gastric mucosa (IC(50) = 32 to 103 pmol/mg protein). But, the prodigiosins, unlike omeprazole, showed little inhibitory effect on K(+)-dependent ATPase (K(+) ATPase) activity, although at higher concentrations they inhibited K(+)-ATPase activity with an IC(50) of 1.5 to 3.0 microM. Furthermore, the inhibitory effect of the prodigiosins was rapid and completely reversible unlike that of omeprazole, and the mode of inhibition was non-competitive with respect to ATP. Hog gastric (H(+)+K(+))-ATPase itself showed an absolute requirement of halide (effectively, chloride) for acidification activity. Prodigiosins also showed a chloride requirement for inhibition of vesicular acidification, and quickly reversed the acidification of vesicular pH to neutrality even in the presence of N, N'-dicyclohexylcarbodiimide (DCCD), showing their ionophoric nature of acidification inhibitory activity. In fact, tributyltin chloride (TBT, an OH( )/Cl(-) exchange ionophore) also inhibited vesicular acidification, but it inhibited K(+)-ATPase activity too. Finally, the prodigiosins inhibited the acid secretion from parietal cells isolated from rabbit gastric mucosa. These results suggest that prodigiosins are potent reversible uncouplers of (H(+)+K(+))-ATPase that inhibit gastric acid secretion. PMID- 11108802 TI - Anticancer activity evaluation of the solanum glycoalkaloid solamargine. Triggering apoptosis in human hepatoma cells. AB - Solamargine, an herbal and molluscicidal medicine derived from Solanum incanum, is a steroidal alkaloid glycoside. To characterize the anticancer mechanism of solamargine on human hepatoma cells (Hep3B), changes of cell morphology, DNA content, and gene expression of cells after solamargine treatment were studied. The appearance in solamargine-treated cells of chromatin condensation, DNA fragmentation, and a sub-G(1) peak in a DNA histogram suggests that solamargine induces cell death by apoptosis. The maximum number of dead Hep3B cells was detected within 2 hr of incubation with constant concentrations of solamargine, and no further cell death was observed after an extended incubation with solamargine, indicating that the action of solamargine was irreversible. To determine the susceptibility of cell phases to solamargine-mediated apoptosis, Hep3B cells were synchronized at defined cell cycles by cyclosporin A, colchicine, and genistein, followed by solamargine treatment. The IC(50) values of solamargine for control, G(0)/G(1)-, M-, and G(2)/M-synchronized Hep3B cells were 5.0, > 10, 3.7, and 3.1 microg/mL, implying that cells in the G(2)/M phases are relatively susceptible to solamargine-mediated apoptosis. In addition, a parallel up-regulation of tumor necrosis factor receptor (TNFR)-I and -II on Hep3B cells was detected after solamargine treatment, and the solamargine mediated cytotoxicity could be neutralized with either TNFR-I or -II specific antibody. Therefore, these results reveal that the actions of TNFR-I and -II on Hep3B cells may be independent, and both are involved in the mechanism of solamargine-mediated apoptosis. PMID- 11108803 TI - Involvement of the extracellular signal-regulated protein kinase (ERK) pathway in the induction of apoptosis by cadmium chloride in CCRF-CEM cells. AB - When CCRF-CEM cells were incubated with 5-40 microM CdCl(2,) apoptosis was observed most clearly at 10 microM. Prior to the development of apoptosis, mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK, were activated with different sensitivity to CdCl(2) exposure. ERK and p38 MAPK were phosphorylated with incubation of 1 microM CdCl(2,) but higher than 20 microM CdCl(2) was required for the clear phosphorylation of JNK. In the time-course study, ERK and p38 MAPK were phosphorylated earlier than JNK after CdCl(2) exposure. The in vitro activities of MAPKs also increased in response to CdCl(2) exposure. Pretreatment with an intracellular Ca(2+) chelator, 1, 2-bis(o aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl) ester (BAPTA/AM), suppressed almost completely CdCl(2)-induced phosphorylation of JNK and p38 MAPK, but not ERK phosphorylation, indicating that the activation of JNK and p38 MAPK depends on the intracellular Ca(2+) but that of ERK does not. On the other hand, treatment with a MAPK/ERK kinase (MEK) inhibitor, U0126 (1,4-diamino 2,3-dicyano-1,4-bis[2-aminophenylthio]butadiene ), suppressed CdCl(2)-induced ERK activation and the apoptosis as well. The inhibition of p38 MAPK activity with SB203580 (4-[4-fluorophenyl]-2-[4-methylsulfinylphenyl]-5-[4-pyridyl]1H- imidaz ole) did not protect cells from apoptosis. The present results showed that the activation of ERK, JNK, and p38 MAPK is differently regulated in CCRF-CEM cells exposed to CdCl(2,) and that the ERK pathway seems to be responsible for the induction of apoptosis by CdCl(2) exposure in this human T cell line. PMID- 11108804 TI - Change in caspase-3-like protease in the liver and plasma during rat liver regeneration following partial hepatectomy. AB - Recent studies have shown that many factors orchestrate liver regeneration after a two-thirds partial hepatectomy (PH). However, the termination mechanism in liver regeneration has not been thoroughly studied. In this paper, we report that the activity of liver caspase-3-like protease, which is specifically activated in apoptosis, increases 18, 36, and 48 hr after PH during maximal hepatocyte proliferative activity. This is the first study that shows the activation of an apoptosis-executing enzyme during physiological liver regeneration. These results suggest that apoptosis is induced in each surge of DNA synthesis as the termination mechanism. When phenoxybenzamine, an alpha-blocker that has been reported to inhibit DNA synthesis during liver regeneration, was injected 8 hr after PH, the caspase-3-like activity in the liver peaked at 15 hr after PH and the enzyme activity also increased in plasma at 18 and 24 hr after PH in sharp contrast to the case of normal regeneration. These results indicate that extensive apoptosis is caused by phenoxybenzamine and that the secondary necrosis of apoptotic cells results in the increase of caspase-3-like protease activity in the plasma. PMID- 11108805 TI - Induction of cell cycle-dependent cytotoxicity and apoptosis by new heterodinucleoside phosphate dimers of 5-fluorodeoxyuridine in PC-3 human prostate cancer cells. AB - Fluorodeoxyuridine (5-FdUrd) is an antineoplastic agent with clinical activity against different types of solid tumours. To enhance the effectiveness of this drug, we have synthesised new heterodinucleoside phosphate dimers of 5-FdUrd. These dimers were compared to 5-FdUrd for their cytotoxic effect and the cell cycle dependence of cytotoxicity, as well as for their capacity to induce apoptosis and inhibit thymidylate synthetase (TS) in androgen-independent human PC-3 prostate tumour cells. Incubation of the cells with the dimers N(4) palmitoyl-2'-deoxycytidylyl-(3'-->5')-5-fluoro-2'-deoxyuri din e (dCpam-5-FdUrd) and 2'-deoxy-5-flourouridylyl-(3'-->5')-2'-deoxy-5-fluoro-N(4)-octa decylc ytidine (5-FdUrd-5-FdC18) resulted in a marked cytotoxicity with IC(50) values of 4 microM, similar to 5-FdUrd. In contrast to 5-FdUrd, 100% toxicity was achieved with concentrations of 100-200 microM 5-FdUrd-5-FdC18. Flow cytometric analysis revealed an increase in the cell population in S-phase after treatment with 5 FdUrd, 5-FdUrd-5-FdC18, and dCpam-5-FdUrd from 36 to 63%, 50%, and 77%, respectively. dCpam-5-FdUrd was more potent than 5-FdUrd in arresting the cell cycle. Significant S-phase arrest was indicated by a decreased proportion of cells in G1- and G2/M-phases. Cell cycle arrest and inhibition of cell proliferation were followed by apoptosis, as shown by a 6- to 8-fold increased binding of Apo2.7 antibody, a 9- to 11-fold increase in caspase-3 activity, DNA fragmentation, and by cell morphology showing the appearance of apoptotic bodies. Importantly, 5-FdUrd-5-FdC18 increased the number of apoptotic cells to 160% compared to 5-FdUrd under the same conditions. As with 5-FdUrd, the two dimers also inhibited TS in a time- and concentration-dependent manner, although requiring 100-fold higher concentrations. In conclusion, dCpam-5-FdUrd and 5 FdUrd-5-FdC18 exert stronger cytotoxicity and induce more S-phase arrest and apoptosis than does 5-FdUrd in PC-3 cells, suggesting their potential role in the treatment of human prostate cancer. PMID- 11108806 TI - Rho(0) tumor cells: a model for studying whether mitochondria are targets for rhodamine 123, doxorubicin, and other drugs. AB - A human osteosarcoma cell line devoid of mitochondrial DNA (rho(0)) and its wild type parental cell counterpart (wt) are presented as a model to investigate drug targeting. By virtue of the absence of mitochondrial DNA, rho(0) cells cannot perform electron transport or oxidative phosphorylation. Since most of the drugs studied are transported by the efflux pumping systems controlled by the MDR1 and MRP1 genes, both cell lines were examined for the expression of these genes, and it was found that no MDR1 and only low amounts of MRP1 were expressed. Growth inhibition experiments indicated that doxorubicin (Dox), vinblastine, and paclitaxel were equitoxic in these cell lines. On the other hand, the IC(50) for rhodamine 123 (Rho 123) in rho(0) cells was 50 times higher than in wt cells. This result correlates with a lower accumulation of Rho 123 in rho(0) cells as measured by fluorescence microscopy and flow cytometry (3 times less than in wt cells). In contrast, when stained with Dox, both cell types accumulated similar amounts. Surprisingly, in these non-P-glycoprotein expressing cells, verapamil increased both Dox and Rho 123 retention. Overall, these data suggest that: (i) functional mitochondria do not appear to be targets for the growth inhibitory activities of Dox, paclitaxel, or vinblastine; (ii) for lipophilic cations like Rho 123, however, normal functioning mitochondria and maintenance of a normal mitochondrial membrane potential (Deltapsi(mt)) appear to play a critical role in the intracellular accumulation and subsequent cytotoxicities of these compounds; and (iii) verapamil increases drug accumulation in non-P-glycoprotein expressing cell lines, most likely by direct action on Deltapsi(mt) for Rho 123 and safranin O, and on heretofore unidentified plasma membrane transporters, as well as via interaction with low levels of MRP1, for Dox. These results should be considered when Rho 123 and verapamil are used to detect P-glycoprotein. PMID- 11108807 TI - Interaction of guanine phosphonomethoxyalkyl derivatives with GMP kinase isoenzymes. AB - Substrate activity and inhibitory potency of guanine phosphonomethoxyalkyl derivatives towards GMP kinase isoenzymes from L1210 cells were studied. 9-[2 (Phosphonomethoxy)ethyl]guanine (PMEG) and the (R)- and (S)-enantiomers of both 9 [3-hydroxy-2-(phosphonomethoxy)propyl]guanine (HPMPG) and 9-[2 (phosphonomethoxy)propyl]guanine (PMPG) were phosphorylated to the first step. Kinetic data showed that (R)-PMPG was a good substrate with a relative phosphorylation efficacy of 12% compared with the natural substrate GMP, whereas PMEG was a poor substrate with a relative phosphorylation efficacy of 1.1%. The structurally related 2,6-diaminopurine analogues 9-[2-(phosphonomethoxy)ethyl]-2, 6-diaminopurine (PMEDAP) and (R)- and (S)-9-[2-(phosphonomethoxy)propyl]-2,6 diaminopurine (PMPDAP) were not phosphorylated by any of the GMP kinase isoenzymes tested. The inhibitory activities of the individual compounds on GMP kinase isoenzymes decreased in the following order: (S)-HPMPG > (R)-PMPG > PMEG > (R)-HPMPG > (S)-PMPG > PMEDAP = (R)-PMPDAP = (S)-PMPDAP; each compound exerted a different type of inhibition. PMID- 11108808 TI - Lack of glutathione conjugation to adriamycin in human breast cancer MCF-7/DOX cells. Inhibition of glutathione S-transferase p1-1 by glutathione conjugates from anthracyclines. AB - One of the proposed mechanisms for multidrug resistance relies on the ability of resistant tumor cells to efficiently promote glutathione S-transferase (GST) catalyzed GSH conjugation of the antitumor drug. This type of conjugation, observed in several families of drugs, has never been documented satisfactorily for anthracyclines. Adriamycin-resistant human breast cancer MCF-7/DOX cells, presenting a comparable GSH concentration, but a 14-fold increase of the GST P1-1 activity relative to the sensitive MCF-7 cells, have been treated with adriamycin in the presence of verapamil, an inhibitor of the 170 P-glycoprotein (P-gp) drug transport protein, and scrutinized for any production of GSH-adriamycin conjugates. HPLC analysis of cell content and culture broths have shown unequivocally that no GSH conjugates are present either inside the cell or in the culture broth. The only anthracycline present inside the cells after 24 hr of incubation was > 98% pure adriamycin. Confocal laser scanning microscopic observation showed that in MCF-7/DOX cells adriamycin was localized mostly in the Golgi apparatus rather than in the nucleus, the preferred site of accumulation for sensitive MCF-7 cells. These findings rule out GSH conjugation or any other significant biochemical transformation as the basis for resistance to adriamycin and as a ground for the anomalous localization of the drug in the cell. Adriamycin, daunomycin, and menogaril did not undergo meaningful conjugation to GSH in the presence of GST P1-1 at pH 7.2. Indeed, their synthetic C(7)-aglycon GSH conjugates exerted a strong inhibitory effect on GST P1-1, with K(i) at 25 degrees in the 1-2 microM range, scarcely dependent on their stereochemistry at C(7). PMID- 11108809 TI - Metabolism of 4'-thio-beta-D-arabinofuranosylcytosine in CEM cells. AB - Because of the excellent in vivo activity of 4'-thio-beta-D arabinofuranosylcytosine (T-araC) against a variety of human solid tumors, we have studied its metabolism in CEM cells to determine how the biochemical pharmacology of this compound differs from that of beta-D arabinofuranosylcytosine (araC). Although there were many quantitative differences in the metabolism of T-araC and araC, the basic mechanism of action of T-araC was similar to that of araC: it was phosphorylated to T-araC-5' triphosphate (T-araCTP) and inhibited DNA synthesis. The major differences between these two compounds were: (i) T-araC was phosphorylated to active metabolites at 1% the rate of araC; (ii) T-araCTP was 10- to 20-fold more potent as an inhibitor of DNA synthesis than was the 5'-triphosphate of araC (araCTP); (iii) the half-life of T-araCTP was twice that of araCTP; (iv) the catalytic efficiency of T-araC with cytidine deaminase was 10% that of araC; and (v) the 5' monophosphate of araC was a better substrate for deoxycytidine 5'-monophosphate deaminase than was the 5'-monophosphate of T-araC. Of these differences in the metabolism of these two compounds, we propose that the prolonged retention of T araCTP is a major factor contributing to the activity of T-araC against solid tumors. The data in this study represent another example of how relatively small structural changes in nucleoside analogs can profoundly affect the biochemical activity. PMID- 11108810 TI - Electron paramagnetic resonance spectrometry evidence for bioreduction of tirapazamine to oxidising free radicals under anaerobic conditions. AB - Tirapazamine (SR 4233) is a bioreductive antitumour drug in Phase III clinical trial which is activated in hypoxic tumour regions to generate a cytotoxic species. Electron paramagnetic resonance (EPR) spectrometry was used to investigate directly the formation of free radicals as the result of tirapazamine reduction by NADPH-supplemented liver microsomes. Under anaerobic conditions, the tirapazamine nitroxide free radical EPR signal was not evident over a range of rat or human liver microsomal protein (1-5 mg) concentrations. However, in combination with 1,1',5, 5'-dimethylpyrolline-1-N-oxide (DMPO), a spin trap for short-lived free radicals, tirapazamine resulted in formation of a 1:1:1:1:1:1 spectrum with hyperfine splitting A(N) = 15.8 G A(H) = 22.3 G consistent with generation of DMPO-R, a carbon-centered radical adduct. Addition of DMSO increased the signal intensity of the carbon-centred radical by at least twofold. The hyperfine splitting constants associated with DMPO-R could be indicative of a tirapazamine carbon-centred radical per se or, more likely, carbon radicals from endogenous materials (or DMSO) in the biological matrix as a result of oxidative attack by the tirapazamine primary radical. Formation of DMPO-OH, the hydroxyl radical spin adduct, by tirapazamine in the absence of air indicates that liberation of a hydroxyl radical may be a consequence of tirapazamine bioreduction under anaerobic conditions. The reactivity of tirapazamine free radicals with endogenous microsomal substances to generate reactive carbon centred radicals indicates that tirapazamine may disrupt a wide range of cellular activities. PMID- 11108811 TI - Structure-specific control of differentiation and apoptosis of human promyelocytic leukemia (HL-60) cells by A-ring diastereomers of 2-methyl 1alpha,25-dihydroxyvitamin D(3) and its 20-epimer. AB - 1alpha,25-dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) has been shown to modulate not only proliferation and differentiation but also apoptosis of malignant cells, indicating that it would be useful for the treatment of hyperproliferative diseases such as cancer and psoriasis. Little information is available concerning structural motifs of the 1alpha,25(OH)(2)D(3) molecule responsible for modulation of differentiation and apoptosis. We synthesized all possible A-ring diastereomers of the 2-methyl-1alpha,25(OH)(2)D(3) and its 20-epimer and evaluated their biological activities in human promyelocytic leukemia (HL-60) cells. Surprisingly, the potent analogues could be clearly divided into two groups: (i) those bearing the 1alpha- and 3beta-hydroxyl groups on the A-ring were potent inducers of differentiation and growth inhibitors of HL-60 cells and (ii) those bearing the 1beta-hydroxyl group together with either 3alpha- or 3beta hydroxyl groups on the A-ring were potent stimulators of apoptosis in these cells. We have clearly identified for the first time the structural motifs on the basis of the stereochemistry of both hydroxyl groups at positions 1 and 3 of the A-ring of the 1alpha,25(OH)(2)D(3) molecule responsible for the induction of differentiation and apoptosis of HL-60 cells. These findings provide useful information not only for structure-function studies of 1alpha,25(OH)(2)D(3) analogues but also for the development of therapeutic agents for the treatment of leukemia and other cancers. PMID- 11108812 TI - Effect of antibody against integrin alpha4 on bleomycin-induced pulmonary fibrosis in mice. AB - Integrins are a family of transmembrane glycoproteins that can interact with components of the extracellular matrix. The alpha4beta1 and alpha4beta7 integrins are heterodimeric leukocyte cell surface molecules critical to their cell and matrix adhesive interactions. Evidence for a central role for the alpha4 integrins in leukocyte pathophysiology in the lung is well documented. In this study, we tested the hypothesis that neutralizing antibody for integrin alpha4 (PS2) may reduce bleomycin (BL)-induced lung fibrosis in vivo. Male C57BL/6 mice were injected intratracheally with saline (SA) or BL (0.08 U/mouse) followed by intraperitoneal injection of SA, isotype control antibody (1E6), or PS2 (100 microg) three times a week. Twenty-one days after the intratracheal instillation, mice were killed for bronchoalveolar lavage (BAL), biochemical, histopathological, and immunohistological analyses. Treatment with PS2 significantly reduced BL-induced increases in lung lipid peroxidation and hydroxyproline content. Lung histopathology also showed reduced fibrotic lesions in the BL-treated lungs by treatment with PS2. BL-treated mouse lungs also showed induction of cells with the myofibroblast phenotype, as indicated by the increased expression of alpha-smooth muscle actin (alphaSMA), whereas treatment with PS2 minimized the BL-induced alphaSMA expression. Furthermore, treatment with PS2 reduced the BL-induced increase in the BAL total cell number, and attenuated the BL-induced increase in the BAL protein level. It is concluded that integrin alpha4 may play an important role in BL-induced pulmonary fibrosis, and the use of anti-alpha4 antibody offers therapeutic antifibrotic potential in vivo. PMID- 11108813 TI - Ca(2+)-channel blockers and nucleoside triphosphate diphosphohydrolase (NTPDase) influence of diltiazem, nifedipine, and verapamil. AB - The nucleoside triphosphate diphosphohydrolases (NTPDase; EC 3.6.1. 5) are a family of ectonucleotidases associated with vascular endothelial and smooth muscle cells. These ectonucleotidases are involved in the control of vascular tone by regulating the level of circulating ATP. Ca(2+)-channel blocking agents are currently used for the treatment of hypertension. Considering the external localization of the NTPDase catalytic site and its Ca(2+) requirement for enzyme activity, a possible interference of calcium antagonists (nifedipine, verapamil HCl, and diltiazem-HCl and some of its metabolites) could be anticipated. To test that hypothesis, an NTPDase-enriched particulate fraction was used. Our results show that verapamil, diltiazem, and its metabolites all produced a concentration dependent inhibition of NTPDase, at concentrations greater or equal to 0.1 mM with verapamil and to 0.5 mM with diltiazem and its metabolites, whereas no significant effect was observed with nifedipine. Kinetic studies, carried out to define the mode of action of these drugs, showed a mixed type of inhibition. Based on their respective K(i) values (in parentheses, in mM), inhibitory potencies of these molecules were in the following order: desacetyl-N desmethyldiltiazem (M(2)-HCl; 0.6) > verapamil (0.76) > N-desmethyldiltiazem (M(A;) 0.9) > diltiazem (2.4) > desacetyl-O-desmethyldiltiazem (M(4)-HCl; 3.5) > desacetyl N, O-desmethyldiltiazem (M(6)-HCl; 3.9). Hence, these calcium antagonists can be considered as weak NTPDase inhibitors. Moreover, based on these K(i) values and the range of concentrations found in the blood, NTPDase would not be inhibited significantly in vivo. PMID- 11108814 TI - Characterization and inhibition by a wide range of xenobiotics of organic anion excretion by primary human hepatocytes. AB - Organic anion secretion by human hepatocytes was characterized using primary liver parenchymal cell cultures and the anionic fluorescent dye carboxy-2',7' dichlorofluorescein (CF). Probenecid, a well-known common blocker of the membrane transport process for anions, was shown to increase CF accumulation in primary human hepatocytes by inhibiting cellular CF efflux in a dose-dependent manner, thereby establishing the presence of an efflux system for organic anions in cultured hepatocytes. Outwardly directed transport of CF from hepatocytes was found to be temperature-dependent; it was not altered by changes in the ionic composition of the incubation medium used in efflux experiments. In addition to probenecid, various structurally and functionally unrelated xenobiotics such as glibenclamide, rifampicin, vinblastine, MK-571, indomethacin, and cyclosporin A were shown to inhibit secretion of CF by primary human hepatocytes, thus suggesting that organic anion excretion by human liver may be impaired by various drugs. Northern blot and Western blot analyses of the expression of multidrug resistance proteins (MRP), such as MRP1 and MRP2, which are known to mediate cellular outwardly directed transport of organic anions indicated that MRP2 was present at substantial levels in cultured human hepatocytes as well as in their in vivo counterparts, whereas MRP1 expression was only barely detectable. These results therefore suggest that MRP2, unlike MRP1, may contribute to the organic anion efflux system displayed by primary human hepatocytes and inhibited by a wide range of xenobiotics. PMID- 11108815 TI - Increased sensitivity of neonate atrial myocytes to adenosine A1 receptor stimulation in regulation of the L-type Ca2+ current. AB - Adenosine has cardioprotective effects against ischemia, and newborn hearts show high resistance to ischemia. The effects of purinoceptor stimulation by adenosine and ATP on the L-type Ca2+ current (ICa) were examined in atrial cells from neonate and adult rabbits. ICa was measured by the membrane-perforated patch method. Adenosine inhibited the isoproterenol-stimulated ICa more potently in neonate cells than in adult cells. The high sensitivity of neonate myocytes to adenosine was accompanied not only by an increased maximum response but also by a lower IC50 concentration. ATP also inhibited isoproterenol-stimulated ICa. The effect of ATP on neonate cells was stronger than that on adult cells at high concentrations (greater than or = 100 microM). The effect of adenosine was antagonized by an A1 adenosine receptor antagonist, 1,3-dipropyl-8 cyclopentylxanthine (DPCPX). DPCPX or an ecto-5'-nucleosidase inhibitor (alpha,beta-methylene-ADP) blocked most (approximately 60%) of the effect of ATP (30 microM), and co-addition of DPCPX and suramin (P2 receptor blocker) abolished the effect of ATP. Suramin alone did not reduce the effect of ATP significantly in neonate cells. Both the effects of adenosine and ATP were eliminated by pre treatment with pertussis toxin or by superfusion with forskolin plus 3-isobutyl-1 methylxanthine (IBMX). Inhibitors of the nitric oxide-cyclic GMP pathway did not affect the adenosine inhibition of ICa. In summary, neonatal myocardial cells are highly sensitive to adenosine A1 receptor stimulation. ATP stimulates both the adenosine A1 and P2 receptors. Adenosine A1 receptor stimulation, as a result of hydrolysis of ATP, predominantly mediates the effect of ATP, and the role of P2 receptors in the ATP inhibition of ICa is relatively small in neonate cells. The high sensitivity to adenosine may contribute to the ischemic tolerance of newborn hearts. PMID- 11108816 TI - Changes in adipose tissue hormone-sensitive lipase activity and cAMP during ethanol withdrawal. AB - The time course of the effects of ethanol withdrawal on brown and white adipose tissue hormone-sensitive lipase, cAMP production, and phosphodiesterase have been investigated after chronic drinking or liquid diet schedules. Chronic drinking significantly reduced brown adipose tissue hormone-sensitive lipase activity and cAMP levels from control. During withdrawal, there was a rebound increase to 200% control, peaking 9 h into withdrawal. White adipose tissue hormone-sensitive lipase activity and cAMP accumulation were significantly raised by both treatment schedules. Ethanol liquid diet produced a significant fall in adipose tissue hormone-sensitive lipase activity and cAMP accumulation. In brown fat, there was a rebound increase in hormone-sensitive lipase activity and cAMP; in white fat, no rebound was observed. In brown fat, the reductions in hormone-sensitive lipase activity and cAMP accumulation after chronic drinking coincided with an increase in phosphodiesterase activity. In white fat, the rise in cAMP and hormone sensitive lipase activation coincided with a decrease in phosphodiesterase activity. We conclude that the effects of chronic ethanol on hormone-sensitive lipase activity are cAMP-dependent and mediated via alterations in phosphodiesterase activity. PMID- 11108817 TI - Coupling between agonist and chloride ionophore sites of the GABA(A) receptor: agonist/antagonist efficacy of 4-PIOL. AB - Eight gamma-aminobutyric acid (GABA) mimetics were tested on their ability to differentiate native GABA(A) receptor subtypes present in various rat brain regions. In rat brain cryostat sections, little regional variations by the agonistic actions of muscimol, thiomuscimol, 4,5,6,7-tetrahydroisoazolo(5,4 c)pyridin-3-ol, piperidine-4-sulphonic acid, taurine and beta-alanine on [35S]t butylbicyclophosphorothionate ([35S]TBPS) binding to GABA(A) receptor channels were found. They were very similar to those found for GABA itself and indicated no direct correlation with single subunit distributions for any of these compounds. Only the low-efficacy GABA mimetic 5-(4-piperidyl)isoxazol-3-ol (4 PIOL) acted like a weak partial agonist or antagonist depending on the brain area. As the cerebellar granule cell layer was relatively insensitive to both modes of action, we tested 4-PIOL in recombinant alpha1beta2gamma2 (widespread major subtype) and alpha6beta2gamma2 (cerebellar granule cell restricted) receptors where it had different effects on GABA-modulated [35S]TBPS binding and on electrophysiological responses. 4-PIOL may thus serve as a potential lead for receptor subtype selective compounds. PMID- 11108818 TI - Changes in intracellular Ca2+ by activation of P2 receptors in submucosal neurons in short-term cultures. AB - Electrophysiological and Ca2+ microfluorimetric techniques were used to characterize the pharmacological profile of the P2 receptors expressed in submucosal neurons and the changes in intracellular Ca2+ associated with activation of these receptors. ATP caused a fast and slow membrane depolarizations during intracellular recordings. ATP induced a rapid inward current during whole-cell experiments. Receptors mediating the inward current and fast depolarization have the same pharmacological profile and these ATP responses were more sensitive to pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid than Basilen BlueE-3G, and potentiated by suramin. The slow depolarization was not blocked by these P2 receptor antagonists, pertussis toxin, or KT5720 (protein kinase A inhibitor). N-ethylmaleimide or protein kinase C inhibitors (staurosporine and calphostin) blocked this depolarization. ATP induced complex multi-phasic Ca2+ transients in most neurons, classified as fast, slow, or mixed fast/slow responses. In conclusion, the fast and slow Ca2+ responses were mediated by respective activation of P2X and P2Y receptors and were associated with fast and slow depolarizations, respectively. PMID- 11108819 TI - Characterisation of an endogenous bombesin receptor in CHO/DG44 cells. AB - Bombesin and its receptors have been shown to have a role regulating circadian rhythms in the hamster suprachiasmatic and dorsal raphe nuclei and have been implicated in the regulation of sleep. We have identified and characterised a bombesin receptor endogenously expressed in a Chinese hamster ovary cell line (CHO/DG44). Using a range of bombesin-like peptides, we demonstrate that this receptor displays bombesin BB2 receptor-like pharmacology. We also show that this receptor signals through inositol-[1,4,5]-trisphosphate and protein kinase C and thus provides a useful model system to aid in the interpretation of hamster suprachiasmatic nucleus studies of mammalian circadian rhythm. PMID- 11108820 TI - Methamphetamine decreases mouse striatal dopamine transporter activity: roles of hyperthermia and dopamine. AB - Multiple methamphetamine administrations rapidly decrease rat striatal dopamine transporter activity. To determine the species specificity of this phenomenon, the present studies examined effects of this stimulant on the dopamine transporter in mice. As in rats, multiple methamphetamine injections rapidly reduced striatal dopamine transporter activity; a decrease that was partially reversed 24 h later. Moreover, methamphetamine decreased binding of the dopamine transporter ligand, WIN35428, but to a lesser degree than the change in dopamine transporter function. These decreases did not appear to result from residual methamphetamine introduced by the original drug treatment. As in rats, hyperthermia contributed to this phenomenon. Unlike in rats, a role for dopamine was not observed in mice as dopamine depletion, resulting from alpha-methyl-p tyrosine pretreatment, did not prevent this decrease. In addition, unlike in rats, pretreatment with either a dopamine D1 or D2 receptor antagonist (SCH23390 or eticlopride, respectively) did not attenuate the methamphetamine-induced reduction in dopamine uptake. These findings demonstrate both similarities and differences in the acute effects of methamphetamine on dopamine transporter function in mice and rats, and suggest the mouse as an additional model for assessing the acute effects of methamphetamine on the dopamine transporter. PMID- 11108821 TI - Effects of calcitonin gene-related peptide and amylin on human osteoblast-like cells proliferation. AB - Expression of mRNA for calcitonin gene-related peptide (CGRP) and CGRP receptor has been detected in osteoblasts indicating that CGRP could play a role in bone metabolism. In the present study, we evaluated the effect of CGRP on primary culture of human osteoblast-like cells proliferation. The peptide was able to stimulate [3H]thymidine incorporation in human osteoblast-like cells with a maximal effect at 10(-8) M. The proliferating activity of CGRP was not inhibited by the two antagonists, CGRP-(8-37) or amylin-(8-37), whereas amylin fragment antagonized the proliferating activity of amylin. In human osteoblast-like cells CGRP, but not amylin, was able to stimulate adenylyl cyclase activity and this effect was completely antagonized only by CGRP-(8-37) and not by amylin-(8-37). These data suggest that the CGRP induced stimulation of cAMP is not involved in the peptide proliferating effect in human osteoblast-like cells and that in this cell population there are receptor subtypes for CGRP, distinct from that of amylin. PMID- 11108822 TI - Possible mechanisms of action in melatonin reversal of morphine tolerance and dependence in mice. AB - In our earlier study, we reported the ability of melatonin to reverse the development of morphine tolerance and dependence in mice. In the present study, we attempted to analyse the possible involvement of putative melatonin receptors, central and peripheral benzodiazepine receptors and the nitric oxide (NO) system in the mechanism of melatonin reversal of morphine tolerance and dependence in mice. Co-administration of L-N(G)-nitro arginine methyl ester (L-NAME) or melatonin with morphine during the induction phase (days 1-9) delayed the development of tolerance to the anti-nociceptive action of morphine and also reversed naloxone precipitated withdrawal jumpings. L-arginine administration during the induction phase enhanced the development of tolerance to the anti nociceptive effect of morphine but had no effect on the naloxone-precipitated withdrawal response. During the expression phase (day 10), acute administration of melatonin or L-NAME reversed, whereas L-arginine facilitated, naloxone precipitated withdrawal jumping in morphine-tolerant mice, but none of these drugs affected the nociceptive threshold in morphine-tolerant mice. Further, co administration of melatonin or L-NAME with L-arginine during the induction phase antagonized later the effects on the development of morphine tolerance. Also, prior administration of melatonin or L-NAME reversed the L-arginine potentiation of naloxone-precipitated withdrawal jumping in morphine tolerant mice. Among the antagonists for putative melatonin receptors studied, neither luzindole (melatonin MT1 and MT2 receptor antagonist) nor prazosin (melatonin MT3 receptor antagonist) antagonized the melatonin reversal of morphine tolerance and dependence. 1-(2-Chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxam ide (PK11195), a peripheral but not central benzodiazepine receptor antagonist, flumazenil, partially antagonized the melatonin reversal of naloxone-precipitated withdrawal jumping in morphine-dependent mice, but had no effect on the reversal of morphine tolerance induced by melatonin. Overall, the present observations suggest that the melatonin-induced reversal of morphine tolerance and dependence may involve its ability to suppress nitric oxide synthase (NOS) activity. Further, the melatonin-induced reversal of morphine tolerance and dependence is not mediated through its actions via putative melatonin receptors. The agonistic activity of melatonin towards peripheral benzodiazepine receptors may partially contribute to the suppression of morphine dependence but not to the reversal of tolerance to the analgesic activity of morphine. PMID- 11108823 TI - Modulation of extracellular neurotransmitter levels in the nucleus accumbens by a taurine uptake inhibitor. AB - Using in vivo microdialysis, we examined the effect of local perfusion of the taurine uptake inhibitor guanidinoethyl sulfonate on extracellular levels of various neurotransmitters in the rat nucleus accumbens. Guanidinoethyl sulfonate (500 microM-50 mM) produced a concentration-dependent increase in extracellular taurine levels. While 500 microM and 5 mM concentrations of guanidinoethyl sulfonate were largely without effect, 50 mM guanidinoethyl sulfonate produced a significant decrease in extracellular levels of aspartate, glutamate and glycine, with no effect on extracellular dopamine levels. These results indicate that guanidinoethyl sulfonate can modulate extracellular amino acid levels in the nucleus accumbens. PMID- 11108824 TI - 5-HT4 receptors do not mediate the antidepressant-like behavioral effects of fluoxetine in a modified forced swim test. AB - The receptors responsible for mediating the antidepressant effects of selective serotonin reuptake inhibitors are largely unknown. The role of the 5-HT4 receptor in mediating the antidepressant-like effects of fluoxetine in a modified rat forced swim test was examined. Fluoxetine (20 mg/kg) decreased immobility and increased swimming, a pattern shown to represent its actions on the serotonergic system. The selective 5-HT4 receptor antagonist, SB 204070A [8-amino-7-chloro-(N butyl-4-piperidyl)methylbenzo-1, 4-dioxan-5-carboxylate hydrochloride] (0.1-3 mg/kg), failed to change any of the active behaviors in the test compared with saline-treated animals. Upon combination, SB 204070A (3 mg/kg) failed to alter the effects of fluoxetine effects in the test. These data therefore suggest that activation of postsynaptic 5-HT4 receptors, subsequent to reuptake inhibition by fluoxetine, is not necessary for its antidepressant-like behavioral effects in this test. PMID- 11108825 TI - In vitro studies on L-771,688 (SNAP 6383), a new potent and selective alpha1A adrenoceptor antagonist. AB - L-771,688 (SNAP 6383, methyl(4S)-4-(3, 4-difluorophenyl)-6-[(methyloxy)methyl]-2 oxo-3-[(?3-[4-(2-pyridin yl)-1-piperidinyl]propyl?amino)carbonyl]-1,2,3, 4 tetrahydro-5-pyrimidine carboxylate) had high affinity (Ki less than or = 1 nM) for [3H]prazosin binding to cloned human, rat and dog alpha1A-adrenoceptors and high selectivity (>500-fold) over alpha1B and alpha1D-adrenoceptors. [3H]Prazosin / (+/-)-beta-[125I]-4-hydroxy-phenyl)-ethyl-aminomethylteralone ([125I]HEAT) binding studies in human and animal tissues known to contain alpha1A and non alpha1A-adrenoceptors further demonstrated the potency and alpha1A-subtype selectivity of L-771,688. [3H]L-771,688 binding studies at the cloned human alpha1A-adrenoceptors and in rat tissues indicated that specific [3H]L-771,688 binding was saturable and of high affinity (Kd=43-90 pM) and represented binding to the pharmacologically relevant alpha1A-adrenoceptors. L-771,688 antagonized norepinephrine-induced inositol-phosphate responses in cloned human alpha1A adrenoceptors, as well as phenylephrine or A-61603 (N-[5-4,5-dihydro-1H-imidazol 2yl)-2-hydroxy-5,6,7, 8-terahydro-naphthlen-1-yl] methanesulfonamide hydrobromide) induced contraction in isolated rat, dog and human prostate, human and monkey bladder neck and rat caudal artery with apparent Kb values of 0.02 0.28 nM. In contrast, the contraction of rat aorta induced by norepinephrine was resistant to L-771,688. These data indicate that L-771,688 is a highly selective alpha1A-adrenoceptor antagonist. PMID- 11108826 TI - Vascular permeability in allergic conjunctivitis in mice lacking histamine H1 receptors. AB - To clarify the role of histamine H1 receptors in allergic conjunctivitis, changes in vascular permeability of the conjunctiva were measured in histamine H1 receptor deficient mice. Wild-type mice showed a significant increase in vascular permeability of the conjunctiva induced by histamine. However, no such increase was found in histamine H1 receptor deficient mice. On the other hand, no differences were observed between wild-type and histamine H1 receptor deficient mice in response to serotonin. A significant increase in vascular permeability was observed in actively sensitized wild-type mice, whereas no increase was observed in histamine H1 receptor deficient mice. Similar findings were noted in passively sensitized animals. Histamine contents of the conjunctiva were significantly decreased by topical application of antigen in both wild-type and histamine H1 receptor deficient mice after active sensitization with antigen. These findings suggested that vascular permeability in the conjunctiva in allergic conjunctivitis is entirely regulated through histamine H1 receptor. PMID- 11108827 TI - Adenosine A1 receptor blockade reverses dysmotility induced by ischemia reperfusion in rat colon. AB - This study was designed to assess whether adenosine A1 receptor antagonists [(R) 1-[(E)-3-(2-phenylpyrazolo[1,5-a]pyridin-3-yl) acryloyl]-piperidin-2-yl acetic acid (FK352) and 8-cyclopentyl-1,3-dipropylxanthine (DPCPX)] reverse dysmotility induced by ischemia-reperfusion in the rat colon. The gene of adenosine A1 receptor was expressed in the colon. Clamping (30 min) of the colonic marginal vessels was followed by reperfusion, and the propulsive colonic motility was evaluated. Propulsion was significantly slowed by ischemia-reperfusion, while FK352 and DPCPX abolished this delay. In contrast, the non-selective adenosine receptor antagonist, 8-phenyltheophylline, failed to affect the dysmotility. Thus, adenosine A1 receptor antagonists have potent therapeutic potential against ischemia-reperfusion-induced dysmotility in the colon. PMID- 11108828 TI - Inhibitory effect of a novel quinolinone derivative, TA-270, on asthmatic inflammatory responses in sensitized guinea pigs. AB - TA-270 (4-hydroxy-1-methyl-3-octyloxy-7-sinapinoylamino-2(1H)- quinolinone), a novel quinolinone derivative, was designed as an antioxidant to scavenge reactive oxygen species. Here, we investigated the effects of TA-270, in comparison with several antiasthmatic drugs, on asthmatic responses as induced by ovalbumin in sensitized guinea pigs. When orally administered 1 h before and 3 h after the antigen challenge, TA-270 at 10 mg/kg and higher doses significantly inhibited both immediate and late responses in airway resistance induced by the antigen. The inhibitory effects were comparable to or superior, at least under the present experimental conditions, to those of several clinically used antiasthmatic drugs. Furthermore, TA-270, in a dose-dependent manner, reduced accumulation of pulmonary inflammatory cells, especially eosinophils, and significantly reversed the airway hyperresponsiveness to acetylcholine 24 h after the antigen challenge. These results suggest that TA-270 may be of therapeutic use for bronchial asthma. PMID- 11108829 TI - A novel anti-rheumatic drug suppresses tumor necrosis factor-alpha and augments interleukin-10 in adjuvant arthritic rats. AB - Tumor necrosis factor-alpha (TNF-alpha) plays an important role in the pathology of rheumatoid arthritis. When N-[1-(4-?[4-(pyrimidin-2-yl)piperazin-1 yl]methyl?phenyl)cycloprop yl] acetamide (Y-39041) (3-30 mg/kg) was orally administered to rats with established arthritis from day 15 to day 20, hindpaw volume was significantly reduced. This inhibitory activity of Y-39041 was kept up after administration was stopped. On day 17 Y-39041 suppressed lipopolysaccharide induced TNF-alpha and interleukin-6 production in serum at doses of 3-30 mg/kg, and augmented interleukin-10 production at doses of 10 and 30 mg/kg. The finding that Y-39041 suppresses TNF-alpha and interleukin-6 production and augments interleukin-10 production could be beneficial in the therapy of chronic inflammatory diseases. PMID- 11108831 TI - Does europe exist? PMID- 11108830 TI - Trichloroethanol inhibits ATP-induced membrane currents in cultured HEK 293-hP2X3 cells. AB - Membrane currents in response to the application of alpha, beta-methylene ATP (alpha,beta-meATP) were recorded by the whole-cell patch-clamp technique in human embryonic kidney 293 cells transfected with the human P2X3 receptor (HEK 293 hP2X3 cells). Trichloroethanol, the biologically active metabolite of chloral hydrate, but not ethanol itself concentration-dependently and reversibly inhibited the current responses. It was concluded that the reported analgesic effect of chloral hydrate may be due to the interruption of pain transmission in dorsal root ganglia expressing P2X3 receptors. PMID- 11108832 TI - Protein sorting in the Golgi apparatus: a consequence of maturation and triggered sorting. AB - To explain how resident proteins distribute in peak-like patterns at various positions in the secretory pathway, Glick and co-workers postulated that resident proteins comprise different populations (termed kin populations) and that these compete with each other for entering retrograde transport carriers [Glick et al. (1997) FEBS Lett. 414, 177-181]. Using modelling and computer simulation, they could demonstrate that differences in competitiveness sufficed to generate overlapping but distinct peak-like steady state distributions of the different kin populations across the Golgi stack. In this study, we have tested the robustness of the competition model and find that over-expression or changes in the number of kin populations affect their overall steady state distributions. To increase the robustness of the system, we have introduced a milieu-induced trigger for recycling. This allows for a decrease in the coupling between kin populations permitting both over-expression as well as changes in the number of kin populations. We have also extended the model to include a Golgi to endoplasmic reticulum (ER) recycling pathway and find that only a small amount of resident proteins may recycle at any time without upsetting their observed distributions in the Golgi stack. The biological relevance of a trigger-induced sorting mechanism and ER recycling is discussed. PMID- 11108833 TI - Hydrogen peroxide in the human body. AB - Hydrogen peroxide (H(2)O(2)) is widely regarded as a cytotoxic agent whose levels must be minimized by the action of antioxidant defence enzymes. In fact, H(2)O(2) is poorly reactive in the absence of transition metal ions. Exposure of certain human tissues to H(2)O(2) may be greater than is commonly supposed: substantial amounts of H(2)O(2) can be present in beverages commonly drunk (especially instant coffee), in freshly voided human urine, and in exhaled air. Levels of H(2)O(2) in the human body may be controlled not only by catabolism but also by excretion, and H(2)O(2) could play a role in the regulation of renal function and as an antibacterial agent in the urine. Urinary H(2)O(2) levels are influenced by diet, but under certain conditions might be a valuable biomarker of 'oxidative stress'. PMID- 11108834 TI - Preferential interaction of loach DNA polymerase delta with DNA duplexes containing single-stranded gaps. AB - We studied the interaction of DNA polymerase delta (pol delta) purified from the eggs of the teleost fish Misgurnus fossilis (loach) with DNA duplexes containing single-stranded gaps of 1-13 nucleotides (nt). In the absence of processivity factors (PCNA, RF-C, and ATP), pol delta elongated primers on single-stranded DNA templates in a distributive manner. However, the enzyme was capable of processive synthesis by filling gaps of 5-9 nt in DNA duplexes. These data suggest that, upon filling a small gap, pol delta interacts with the 5'-terminus downstream of the gap as well as with the 3'-terminus of the primer. Interaction of pol delta with the proximal 5'-terminus restricting the gap was confirmed by electrophoretic mobility shift assay. Analysis of the enzyme binding to DNA duplexes containing gaps of various sizes showed a much higher affinity of pol delta for duplexes with gaps of about 10 nt than for DNA substrates with primers annealed to single-stranded templates. The most efficient pol delta binding was observed in experiments with DNA substrates containing unpaired 3'-tails in primers. The data obtained suggest that DNA molecules with small gaps and single stranded tails may serve as substrates for direct action of pol delta in the course of DNA repair. PMID- 11108835 TI - Antioxidant actions of ovothiol-derived 4-mercaptoimidazoles: glutathione peroxidase activity and protection against peroxynitrite-induced damage. AB - 4-Mercaptoimidazoles derived from the naturally occurring antioxidants, ovothiols, were tested for their glutathione peroxidase-like (GSH Px-like) activity and protection against peroxynitrite-induced damage. All the thiol compounds displayed similar significant GSH Px-like activities, which are however weaker than that of the reference compound, ebselen. The inhibitions of the peroxynitrite-dependent oxidation of Evans blue dye and dihydrorhodamine 123 showed that the thiol compounds substituted on position 5 of the imidazole ring were nearly as effective as ebselen while the C-2 substituted ones were less effective. Both assays corroborate the large superiority of mercaptoimidazoles over glutathione as inhibitors of peroxynitrite-dependent oxidation. PMID- 11108836 TI - Activation of p38 mitogen-activated protein kinase is crucial in osteoclastogenesis induced by tumor necrosis factor. AB - Tumor necrosis factor (TNF) induces osteoclast differentiation from bone marrow cells in the presence of macrophage colony-stimulating factor. Treatment of bone marrow cells with SB203580 but not PD98059 inhibited TNF-induced osteoclast differentiation. In RAW264 cells which differentiate into osteoclast-like multinucleated cells by TNF treatment alone, activation of p38 mitogen-activated protein (MAP) kinase induced by murine TNF was comparable to and independent of the receptor activator of necrosis factor-kappaB ligand. Moreover, the number of multinucleated osteoclasts induced by TNF in bone marrow cell cultures derived from p38 MAP kinase gene deficient mice was significantly less than that from control mice. These results indicate that the p38 MAP kinase pathway plays a crucial role in TNF-mediated osteoclast differentiation. PMID- 11108837 TI - Electroporation-mediated gene transfer in free-swimming embryonic Xenopus laevis. AB - Xenopus laevis are a rich resource for vertebrate embryology and cell biology. Transplantation and transgenesis have provided much information about the developmental mechanisms of embryogenesis and molecule function, however existing methods have faced limitations regarding either the precise localization of gene expression or flexibility in the timing of gene transfer. Here we have found that electroporation of tailbud (stage 29/30) embryos is a rapid and efficient method of combining cell-specific expression with variation in temporal delivery. At the low voltages required for electroporation, embryos resumed normal swimming behavior and development. We conclude that electroporation has wide experimental application to Xenopus developmental and cell biology. PMID- 11108838 TI - Identification of the Y985 and Y1077 motifs as SOCS3 recruitment sites in the murine leptin receptor. AB - The leptin system provides a link between adipose mass and the central nervous system. The appetite suppressing effects of leptin are impaired in most obese patients and some mutant mice strains. Herein we describe how suppressor of cytokine signalling 3 (SOCS3), a potential mediator of this leptin resistance is recruited into the activated murine leptin receptor complex. Using a functional assay based on inhibition of leptin mediated reporter induction, and using phosphopeptide affinity chromatography we show binding of SOCS3 to the highly conserved phosphorylated Tyr-985 and Tyr-1077 motifs within the mouse leptin receptor. PMID- 11108839 TI - The T cell antigen receptor activates phosphatidylinositol 3-kinase-regulated serine kinases protein kinase B and ribosomal S6 kinase 1. AB - The present study has explored T cell antigen receptor-regulated serine kinases in human T cells. The results identify two phosphatidylinositol 3-kinase (PI3K) controlled serine kinases operating downstream of the T cell receptor (TCR) in primary T cells: (i) protein kinase B whose activation regulates the phosphorylation of glycogen synthase kinase 3 and (ii) ribosomal S6 kinase 1, a kinase with a critical role in the regulation of protein synthesis and cell growth. T cells express two isoforms of S6k1: a 70 kDa cytoplasmic kinase and an 85 kDa isoform that has a classic nuclear localisation. TCR ligation triggers a parallel engagement of both the 70 and 85 kDa isoforms of S6k1 in a response that requires PI3K function. PMID- 11108840 TI - The common C-terminal sequences of substance P and neurokinin A contact the same region of the NK-1 receptor. AB - Although neurokinin A (NKA), a tachykinin peptide with sequence homology to substance P (SP), is a weak competitor of radiolabeled SP binding to the NK-1 receptor (NK-1R), more recent direct binding studies using radiolabeled NKA have demonstrated an unexpected high-affinity interaction with this receptor. To document the site of interaction between NKA and the NK-1R, we have used a photoreactive analogue of NKA containing p-benzoyl-L-phenylalanine (Bpa) substituted in position 7 of the peptide. Peptide mapping studies of the receptor photolabeled by (125)I-iodohistidyl(1)-Bpa(7)NKA have established that the site of photoinsertion is located within a segment of the receptor extending from residues 178 to 190 (VVCMIEWPEHPNR). We have previously shown that (125)I-BH Bpa(8)SP, a photoreactive analogue of SP, covalently attaches to M(181) within this same receptor sequence. Importantly, both of these peptides ((125)I iodohistidyl(1)-Bpa(7)NKA and (125)I-BH-Bpa(8)SP) have the photoreactive amino acid in an equivalent position within the conserved tachykinin carboxyl-terminal tail. In this report, we also show that site-directed mutagenesis of M(181) to A(181) in the NK-1R results in a complete loss of photolabeling of both peptides to this receptor site, indicating that the equivalent position of SP and NKA, when bound to the NK-1R, contact the same residue. PMID- 11108841 TI - Main ethanol metabolizing alcohol dehydrogenases (ADH I and ADH IV): biochemical functions and the physiological manifestation. AB - The range of the biochemical reactions which can be catalyzed by ADH I and ADH IV is extremely wide. The most characterized functions of these enzymes are protection against excess endogenous acetaldehyde, products of lipid peroxidation, exogenous alcohols and some xenobiotics. It was found also that ADH I and ADH IV are important members of the enzyme system synthesizing retinoic acid (especially during embryogenesis). They can oxidize some steroids and participate in bioamine and prostaglandin metabolism but so far the extent of their contribution to the latter processes is under discussion. Recent data suggest a correlation between the activity of ADH I in some organs and fine physiological processes including behavior regulation and craving for alcohol in albino rats. PMID- 11108842 TI - Isolation, reconstitution and functional characterisation of the Rhodobacter sphaeroides photoactive yellow protein. AB - We report the isolation, functional reconstitution and photophysical characterisation of Rhodobacter sphaeroides photoactive yellow protein (PYP), of which the gene was recently cloned. Reconstitution of the his-tagged purified apo protein with 4-hydroxy-cinnamic acid yields the characteristic blue absorbance at 446 nm, but surprisingly also an absorbance peak at 360 nm. This additional peak is not caused by binding of a second chromophore, as confirmed with mass spectroscopy. Moreover, reconstitution with the 'locked' analogue 7-hydroxy coumarin-3-carboxylic acid yields only a single absorbance peak at 441 nm. The 446 nm and 360 nm species are part of a temperature- and pH-dependent equilibrium. Photoactivation of the protein leads to formation of a blue-shifted intermediate as in other PYPs, with a 100-fold increased groundstate recovery rate (k(pB-->pG)=500 s(-1)) compared to E-PYP. PMID- 11108843 TI - The high affinity ATP binding site modulates the SecA-precursor interaction. AB - SecA is the central component of the protein-translocation machinery of Escherichia coli. It is able to interact with the precursor protein, the chaperone SecB, the integral membrane protein complex SecYEG, acidic phospholipids and its own mRNA. We studied the interaction between prePhoE and SecA by using a site-specific photocrosslinking strategy. We found that SecA is able to interact with both the signal sequence and the mature domain of prePhoE. Furthermore, this interaction was dependent on the type of nucleotide bound. SecA in the ADP-bound conformation was unable to crosslink with the precursor, whereas the ATP-bound conformation was active in precursor crosslinking. The SecA precursor interaction was maintained in the presence of E. coli phospholipids but was loosened by the presence of phosphatidylglycerol bilayers. Examining SecA ATP binding site mutants demonstrated that ATP hydrolysis at the N-terminal high affinity binding site is responsible for the changed interaction with the preprotein. PMID- 11108844 TI - Cholecystokinin octapeptide CCK-8 and carbachol reduce [(32)P]orthophosphate labeling of phosphatidylcholine without modifying phospholipase D activity in rat pancreatic acini. AB - We have studied phospholipase D activation in [(32)P]orthophosphoric acid prelabeled rat pancreatic acini by measuring the formation of (32)P phosphatidylalcohols as stimulated in the presence of ethanol or butanol. A small but significant and time-dependent basal accumulation of [(32)P]phosphatidylethanol and [(32)P]phosphatidylbutanol was detected, which was further stimulated by phorbol myristate acetate, orthovanadate and pervanadate. However, the secretagogues cholecystokinin octapeptide and carbachol did not enhance basal accumulation of (32)P-phosphatidylalcohol, yet they decreased [(32)P]phosphatidylcholine content and stimulated the generation of [(32)P]phosphatidic acid. Our results stress the need to examine the transphosphatidylation reaction as well as agonist effects on the synthesis of phosphatidylcholine in order to assess unambiguously phospholipase D activity. PMID- 11108845 TI - The semaphorin 3A receptor may directly regulate the activity of small GTPases. AB - The axon guidance signal semaphorin 3A induces the rapid collapse of growth cones by activating a receptor complex that contains neuropilin-1 as the ligand-binding and a plexin as the signal-transducing subunit. Here we show that plexins bind Rho-like GTPases and may directly regulate their activity. The cytoplasmic domain of plexins shows sequence similarity to GTPase activating proteins (GAPs) and mutation of two arginines that correspond to the catalytic residues of Ras GAPs inactivates plexin-A1. Our data suggest that plexins may be integral membrane proteins with an intrinsic GAP activity that is essential for their ability to induce growth cone collapse. PMID- 11108846 TI - Characterization of wheat DP, a heterodimerization partner of the plant E2F transcription factor which stimulates E2F-DNA binding. AB - Recent studies suggest that the G1/S transition in plants depends on the activity of E2F transcription factors. In animal cells, E2Fs interact with DP proteins, whose identification in plants has been elusive, so far. Here we show that although an E2F-containing DNA-binding activity can be detected in plant cell extracts, purified E2F protein binds poorly to DNA. In a yeast two-hybrid screening, using wheat E2F as a bait, we have isolated a cDNA clone encoding a wheat DP (TmDP) protein. TmDP is expressed ubiquitously and exhibits a domain organization similar to animal DPs. Contrary to the specificity observed for the plant RBR/E2F interaction, human and plant E2F and DP proteins can interact in a heterologous manner. Purified TmDP protein stimulates E2F-DNA complex formation. PMID- 11108848 TI - Structural complexity in mussel beds: the fractal geometry of surface topography. AB - Seafloor topographic complexity is ecologically important because it provides habitat structure and alters boundary-layer flow over the bottom. Despite its importance, there is little agreement on how to define and measure surface complexity. The purpose of this investigation was to utilize fractal geometry of vertical cross-section profiles to characterize the surface topography of the soft-bottom mussel bed (Mytilus edulis L.) at Bob's Cove, ME, USA. Mussels there have been shown previously to have spatially ordered fractal characteristics in the horizontal plane. Two hypotheses were tested. The first was that the bed surface is fractal over the spatial scale of 1.44-200 mm, with fractal dimension less than or equal to 1.26, the value for the Koch curve, our model for bed profiles. The second was that bed surface topography (i.e., in vertical profile) is less complex than the mussel bed spatial pattern (i.e., aerial view in the horizontal plane). Both hypotheses were supported. Cross-sections of plaster casts of the bed produced 88 surface profiles, all of which were fractal over the entire spatial scale of more two orders of magnitude employed in the analysis. Fractal dimension values (D) for individual profiles ranged from 1.031 to 1.310. Fractal dimensions of entire casts ranged up to mean (1.242+/-0.046) and median (1.251) values similar to 1.26, the theoretical value of the Koch curve. The bed surface was less complex than the bed spatial pattern because every profile had D<1.36, the smallest value previously obtained from aerial views of the bed. The investigation demonstrated for the first time that surface topography of a soft bottom mussel bed was fractal at a spatial scale relevant to hydrodynamic processes and habitat structure important for benthic organisms. The technique of using cross-section profiles from casts of the bed surface avoided possible underestimates of fractal dimension that can result from other profiling methods reported in the literature. The results demonstrate that fractal dimension can be useful in the analysis of habitat space and water flow over any irregular seafloor surface because it incorporates the size, shape, and scale of roughness elements into a simple, numerical metric. PMID- 11108847 TI - Characterization of two distinct DP-related genes from Arabidopsis thaliana. AB - E2F/DP complexes play a pivotal role in the regulation of the G1/S transition in animals. Recently, plant E2F homologs have been cloned, but DP-related sequences have not been identified so far. Here we report that Arabidopsis thaliana contains at least two different DP-related genes, AtDPa and AtDPb. They exhibit an overall domain organization similar to that of their animal counterparts, although phylogenetic analysis demonstrated that they form a separate subgroup. AtDPs efficiently heterodimerize in vitro with the Arabidopsis E2F-related proteins, AtE2Fa and AtE2Fb through their dimerization domains. AtDPa and AtE2Fa are predominantly produced in actively dividing cells with highest transcript levels in early S phase cells. PMID- 11108849 TI - Evaluating the impact of predation by fish on the assemblage structure of fishes associated with seagrass (Heterozostera tasmanica) (Martens ex Ascherson) den Hartog, and unvegetated sand habitats. AB - The role of fish predation in structuring assemblages of fish over unvegetated sand and seagrass was examined using enclosure and exclusion cages to manipulate the abundance of predatory fish from November 1998 to January 1999. In our exclusion experiment, piscivorous fish were excluded from patches of unvegetated sand and seagrass to measure how they altered abundances of small fishes, i.e., fish <10 cm in length. Habitats from which piscivorous fish were excluded contained more small fish than those with partial cages, which in turn contained more fish than uncaged areas. These patterns were consistent between unvegetated sand and seagrass areas, although the relative differences between predator treatments varied with habitat. Overall, small fish were more abundant in unvegetated sand than seagrass. Atherinids and syngnathids were the numerically dominant families of small fish and varied in complex ways amongst habitats and cage treatments. The abundance of atherinids varied inconsistently between cage treatments through time. Only during the final two sampling times did the abundance of atherinids vary significantly across cage treatments. Syngnathids were strongly associated with seagrass and were significantly more abundant in caged than uncaged habitats. In our enclosure experiment, five individuals of a single species of transient piscivorous fish, Western Australian salmon (Arripidae: Arripis truttacea Cuvier), were enclosed in cages to provide an estimate of the potential for this species to impact on small fish. The abundance of small fish varied significantly between cage treatments. Small fish were more abundant in enclosure cages and exclusion cages than uncaged areas; however, there was no difference in the abundance of small fish in enclosure cages and partial cages, and no difference between exclusion cages and partial cages. These patterns were consistent amongst habitats. Atherinids and syngnathids were again the numerically dominant families of small fish; atherinids varied more with cage structure while syngnathids did not vary statistically between cages, blocks (locations within which a single replicate of each cage treatment was applied) or habitats. Dietary analysis of caged A. truttacea demonstrated the potential for this species to influence the assemblage structure of small fish through predation - atherinids were consumed more frequently in unvegetated sand than seagrass, and syngnathids were consumed only in seagrass, where they are most abundant. Observations of significant cage or predation effects depended strongly on the time at which sampling was undertaken. In the case of the atherinids, no predation or cage effects were observed during the first two sampling times, but cage effects and predation effects strongly influenced abundances of fish during the third and fourth sampling times, respectively. Our study suggests that transient piscivorous fish may be important in structuring assemblages of small fish in seagrass and unvegetated sand, and seagrass beds may provide a refuge to fishes. But the importance of habitat complexity and predation, in relation to the potentially confounding effects of cage structure, depends strongly on the time at which treatments are sampled, and the periodicity and multiplicity of sampling should be considered in future predation studies. PMID- 11108850 TI - Effects of nearness to reef and exposure to sea-swell on estimates of relative abundance of Jasus verreauxi (H. Milne Edwards, 1851) recruits on collectors. AB - The hypothesis tested was that catches of J. verreauxi recruits (numbers of pueruli to early juveniles) on sea-weed-type collectors were affected by the nearness of the collectors to vegetated reef and relative exposure of the collectors to sea-swell. Numbers of recruits on collectors set far from reef were greater than on collectors near reef. Collectors far from reef may be the first, preferred type of habitat encountered by recruits as they move across the Continental Shelf. Alternatively, fewer recruits may inhabit collectors set near reef because of the nearness of natural, vegetated reef. A cost-benefit analysis determined that the optimal sampling strategy for surveys of the abundance of recruits at one location and time, would be three replicate collectors at each of three sites. Sampling once with this strategy at one location would give a standard error of around 20% of the mean. This information was used in developing a uniform and optimal methodology for full surveys to monitor the relative abundance of J. verreauxi recruits along the coast of New South Wales. PMID- 11108851 TI - Genetic evidence for limited dispersal in the coastal California killifish, Fundulus parvipinnis. AB - The California killifish, Fundulus parvipinnis, is a marine species that lives in salt marshes, estuaries and wetlands along the California and Baja California coasts. In order to estimate levels of dispersal between different coastal habitats over its range, we have studied six populations using morphological and genetic markers. Lateral line scale and vertebrae counts showed significant differences between individuals collected north of Punta Eugenia and south of Punta Eugenia. Morphological differences across Punta Eugenia were accompanied by large genetic differences at the mitochondrial control region (5.8%). Gene flow was in general very reduced over the range of the species (pairwise average F(st)=0.70, Nm=0.30), with a strong break at Punta Eugenia (F(st)=0.95, Nm=0.03). Such limited interchanges between coastal habitats have important theoretical and conservation implications. PMID- 11108852 TI - Diurnal changes in pore water sulfide concentrations in the seagrass Thalassia testudinum beds: the effects of seagrasses on sulfide dynamics. AB - The dynamics of the seagrass-sulfide interaction were examined in relation to diel changes in sediment pore water sulfide concentrations in Thalassia testudinum beds and adjacent bare areas in Corpus Christi Bay and lower Laguna Madre, Texas, USA, during July 1996. Pore water sulfide concentrations in seagrass beds were significantly higher than in adjacent bare areas and showed strong diurnal variations; levels significantly decreased during mid-day at shallow sediment depths (0-10 cm) containing high below-ground tissue biomass and surface area. In contrast, diurnal variations in sediment sulfide concentrations were absent in adjacent bare patches, and at deeper (>10 cm) sediment depths characterized by low below-ground plant biomass or when the grasses were experimentally shaded. These observations suggest that the mid-day depressions in sulfide levels are linked to the transport of photosynthetically produced oxygen to seagrass below-ground tissues that fuels sediment sulfide oxidation. Lower sulfide concentrations in bare areas are likely a result of low sulfate reduction rates due to low organic matter available for remineralization. Further, high reoxidation rates due to rapid exchange between anoxic pore water and oxic overlying water are probably stimulated in bare areas by higher current velocity on the sediment surface than in seagrass beds. The dynamics of pore water sulfides in seagrass beds suggest no toxic sulfide intrusion into below-ground tissues during photosynthetic periods and demonstrate that the sediment chemical environment is considerably modified by seagrasses. The reduced sediment sulfide levels in seagrass beds during photosynthetic periods will enhance seagrass production through reduced sulfide toxicity to seagrasses and sediment microorganisms related to the nutrient cycling. PMID- 11108853 TI - Upper thermal tolerances of the beachflea Orchestia gammarellus (Pallas) (Crustacea: Amphipoda: Talitridae) associated with hot springs in Iceland. AB - The upper thermal tolerance (CT(max)) of beachfleas Orchestia gammarellus (Pallas) collected from a number of different locations in Iceland was determined. Differences were recorded between field populations associated with thermal springs and those from non-thermal sites. A number of reciprocal acclimation experiments (where animals from thermal and non-thermal sites were acclimated to the measured ambient temperatures of thermal (17 and 22 degrees C) and non-thermal (11 degrees C) sites) were performed. Differences between at least one thermal population and a non-thermal population were maintained following this reciprocal acclimation, supporting the hypothesis that population differences were due to non-reversible genetic differences and not local acclimatisation. Animals from one thermal site (Reykjanes) had a mean CT(max)=37.1+/-0.5 degrees C when acclimated at 11 degrees C and 38.6+/-0.3 degrees C when acclimated at 22 degrees C, whereas animals from a non-thermal site (Hvassahraun) had CT(max) values of 35.9+/-0.5 and 37.9+/-0.3 degrees C, respectively. In other cases, differences are best explained by local acclimatisation. Results are discussed in relation to ambient local conditions and the degree of isolation of the different populations. PMID- 11108854 TI - Burrowing abilities and swash behavior of three crabs, Emerita analoga Stimpson, Blepharipoda occidentalis Randall, and Lepidopa californica Efford (Anomura, Hippoidea), of exposed sandy beaches. AB - To investigate factors related to the distribution of intertidal species, and specific predictions of the swash exclusion hypothesis for exposed sandy beaches, we compared the burrowing abilities and swash behavior of three species of anomuran crabs in the superfamily Hippoidea (Emerita analoga, Blepharipoda occidentalis and Lepidopa californica) which commonly inhabit the intertidal and shallow subtidal zones of beaches along the California coast. Burrowing times in the laboratory increased significantly with crab size for all species in five sediment grain sizes ranging from fine sand to gravel (0.15 to 3.24 mm). For each species, burrowing times differed significantly among sand grain sizes, ranging from 0.3 to 21.5 s. Burrowing times for the hippid crab, E. analoga, were relatively constant across sediment types, while those of the albuneid crabs, B. occidentalis and L. californica, were rapid in fine to medium sands, and much slower in coarser sediments. Our results indicate that E. analoga is a substrate generalist while L. californica and B. occidentalis are substrate sensitive. Pre burrowing times and behavior, distance moved, and burrowing times differed among the species in the swash zone. Combined times of preburrowing and burrowing were shorter than the swash period (6 s) for most E. analoga individuals. Fifty percent of the individuals of L. californica reached the substrate and burrowed in the swash period, while no individuals of B. occidentalis burrowed in that time. Pre-burrowing behavior and time may be valuable in explaining spatial and temporal patterns in the distribution of hippoid crabs on California beaches. Our results support predictions of the swash exclusion hypothesis concerning the burrowing and locomotory abilities of sandy beach macrofauna. The substrate generalist characteristics, and unique orientation and swimming abilities of the hippid crab, E. analoga, in intertidal swash may help explain the success of this species and its congeners, and have important implications for understanding patterns of macrofauna community structure on exposed sandy beaches in California and other regions. PMID- 11108855 TI - Measurement of ingestion rate of Hydrobia ulvae (Pennant) on intertidal epipelic microalgae: the effect of mud snail density. AB - The individual mean ingestion rate of Hydrobia ulvae was measured experimentally in controlled microcosms, in the dark to avoid primary production during measurement and at constant temperature. The experimental design was based on the addition of prelabelled epipelic microalgae to microcosms in a constant proportion with unlabelled diatoms, and in such a way that algal food availability was not a limiting factor within the range of tested densities (0.3 to 4.1 snails cm(-2)). Results show that the individual mean ingestion rate decreased significantly from 26.6+/-1.1 ng Chl a snail(-1) h(-1) to 22.4+/-1.0 ng Chl a snail(-1) h(-1) between 0.7 and 3 snails cm(-2). We hypothesize that this sharp decrease (the threshold density was between 1.4 and 2.5 snails cm(-2)) may account for a density-dependent effect. We have tested this hypothesis by using a simple random walk model including basic behavioural processes such as a break in feeding activity when two individuals contact each other. The model represents quantitatively well the threshold effect, suggesting that behavioural processes have to be taken into account for estimating a global feeding activity of H. ulvae populations. PMID- 11108856 TI - Fatty acid compositions of gonadal material and diets of the sea urchin, Psammechinus miliaris: trophic and nutritional implications. AB - The fatty acid compositions of gonadal material was examined for the sea urchin Psammechinus miliaris (Gmelin) held in aquaria and fed either salmon feed pellets or the macroalga, Laminaria saccharina for 18 months. Gonadal material was also examined from P. miliaris collected from four field sites, including commercial scallop lines encrusted with the mussel, Mytilus edulis, sea cages stocked with Atlantic salmon Salmo salar and two intertidal sea-loch sites, characterised by either a fine mud or a macroalgal substratum. The fatty acid compositions of known and potential dietary material was examined. The proportions of certain fatty acids in the gonads of P. miliaris were significantly affected by diet type and location. Docosahexaenoic acid (DHA) 22:6 n-3 was significantly higher in the gonads of the sea urchins fed salmon feed in aquaria and collected from the salmon cages and scallop lines than in the gonads of the sea urchins fed L. saccharina in aquaria and collected from the intertidal sea loch sites. The salmon feed and the mussel tissue also contained a high proportion of this fatty acid. Stearidonic acid 18:4 n-3 and arachidonic acid 20:4 n-6, however, were found in significantly higher proportions than DHA in the gonads of the sea urchins fed L. saccharina and collected from the two intertidal sea-loch sites. L. saccharina was also found to contain high proportions of stearidonic and arachidonic acid. The gonads of the sea urchins collected from the intertidal site, characterised by a mud substratum, and from the scallop lines were found to contain a lower 18:1 n-9/18:1 n-7 ratio and a higher proportion of branched and odd-chained fatty acids, signifying a high dietary bacterial input, than the sea urchins held in the aquaria and collected from the salmon cage. 20:2 and 22:2 non methylene-interrupted dienoic fatty acids (NMIDs) were found in P. miliaris fed diets lacking these fatty acids suggesting de novo biosynthesis. These results, therefore, suggest that the proportions/ratios of certain fatty acids in the gonads of P. miliaris could be used to give an indication of the predominant diet type of this species in the wild. PMID- 11108858 TI - Progesterone receptor coactivators. AB - Progesterone action is mediated by intracellular progesterone receptors that regulate target gene transcription. Recently, numerous proteins termed coactivators have been identified that are recruited by the liganded progesterone receptor and enhance receptor-dependent transactivation. Coactivators are a diverse group of molecules that bring multiple structural and enzymatic functions to the promoter. The existence of coactivators represents yet another level of regulation for progesterone receptor activation. PMID- 11108859 TI - Progesterone action and responses in the alphaERKO mouse. AB - Ovarian steroids have important inter-related roles in many systems and processes required for mammalian reproduction. The female reproductive tract, ovaries, and mammary glands are all targets for both estrogen and progesterone. In addition, the actions of these hormones are intertwined in that, for example, progesterone attenuates the proliferative effect of estrogen in the uterus, whereas estrogen also induces the progesterone receptor (PR) mRNA and protein, thus enhancing progesterone actions. The generation of mice that lacks the progesterone receptor (PRKO) or the estrogen receptoralpha (alphaERKO) has provided numerous insights into the interacting roles of these hormones. The mammary glands of the PRKO mice develop with full epithelial ducts that lack side branching and lobular alveolar structures, whereas the alphaERKO mice develop only an epithelial rudiment. This indicates that estrogen is important for ductal morphogenesis, whereas progesterone is required for ductal branching and alveolar development. Both the alphaERKO and PRKO mice are also anovulatory, but exhibit different causal pathologies. The alphaERKO ovary seems to possess follicles up to the preantral stage and shows a polycystic phenotype as a result of chronic hyperstimulation by LH. The PRKO follicles seem to develop to an ovulatory stage, but are unable to rupture, indicating a role for progesterone in ovulation. The uteri of these two strains seem to develop normally; however, the function and hormone responses are abnormal in each. Because estrogen is known to induce PRs in the uterus, the progesterone responsiveness of the alphaERKO uterus was characterized. PR mRNA was detected but was not up-regulated by estrogen in the alphaERKO tissue. PRs are present in the alphaERKO tissue at 60% of the level in wild-type tissue and show a similar amount of A and B isoforms when measured by R5020 binding and detected by Western blotting. The PRs were able to mediate induction of two progesterone-responsive uterine genes: calcitonin and amphiregulin. The alphaERKO uterine tissue was also able to undergo a decidual reaction in response to hormonal and intraluminal treatments to mimic implantation; however, unlike normal wild-type uteri, this response was estrogen independent in the alphaERKO uterine tissue. PMID- 11108860 TI - Ovulation: a multi-gene, multi-step process. AB - The luteinizing hormone (LH) surge initiates a cascade of proteolytic events that control ovulation. One of the genes induced by LH is the progesterone receptor (PR). Because mice with a mutant PR gene (PRKO) fail to ovulate and are infertile, we have used them as a model in which to determine PR target genes that might mediate the ovulatory process. The matrix metalloproteinases (MMPs: MMP2, MMP9, and MMP13) appear to be expressed in ovaries of PRKO mice in a manner similar to that in their wild-type littermates. However, the expression of two other types of proteases, cathepsin L (a member of the papain family) and ADAMTS 1 (A Disintegrin And Metalloproteinase with Thrombospondin-like motifs), are selectively induced in granulosa cells of preovulatory follicles by the LH surge. Maximal levels of these proteases are observed at 12-16 h after an LH surge, the time of ovulation. Furthermore, mRNAs encoding cathepsin L and ADAMTS-1 are reduced in the PRKO mice compared to their wild-type littermates. These novel observations indicate that these two proteases regulate some key step(s) controlling ovulation. PMID- 11108861 TI - Progesterone receptors in reproduction: functional impact of the A and B isoforms. AB - Progesterone (P) is a key regulator of female reproductive activity. The effects of P are mediated by two progesterone receptor (PR) proteins, termed A and B, that arise from a single gene and act as ligand-activated transcription factors to regulate the expression of reproductive target genes. Null mutation of the PR gene in mice (PRKO) leads to pleiotropic reproductive abnormalities. This paper will review the reproductive functions of PRs delineated using the PRKO mouse. Further, we will summarize the structure and functional properties of PRs and discuss how functional differences between the PR-A and PR-B isoforms are likely to impact on the overall physiological role of the receptor in reproductive systems. PMID- 11108862 TI - Nuclear receptor conformation, coregulators, and tamoxifen-resistant breast cancer. AB - The development of tamoxifen resistance and consequent disease progression are common occurrences in breast cancers, often despite the continuing expression of estrogen receptors (ER). Tamoxifen is a mixed antagonist, having both agonist and antagonist properties. We have suggested that the development of tamoxifen resistance is associated with an increase in its agonist-like properties, resulting in loss of antagonist effects or even inappropriate tumor stimulation. Nuclear receptor function is influenced by a family of transcriptional coregulators, that either enhance or suppress transcriptional activity. Using a mixed antagonist-biased two-hybrid screening strategy, we identified two such proteins: the human homolog of the nuclear receptor corepressor, N-CoR, and a novel coactivator, L7/SPA (Switch Protein for Antagonists). In transcriptional studies, N-CoR suppressed the agonist properties of tamoxifen and RU486, and L7/SPA increased agonist effects. We speculated that the relative levels of these coactivators and corepressors may determine the balance of agonist and antagonist properties of mixed antagonists, such as tamoxifen. Using quantitative RT-PCR, we, therefore, measured the levels of transcripts encoding these coregulators, as well as the corepressor SMRT, and the coactivator SRC-1, in a small cohort of tamoxifen-resistant and sensitive breast tumors. The results suggest that tumor sensitivity to mixed antagonists may be governed by a complex set of transcription factors, which we are only now beginning to understand. PMID- 11108863 TI - Effects of progesterone on uterine leiomyoma growth and apoptosis. AB - Uterine leiomyomas appear during the reproductive years and regress after menopause, indicating the ovarian steroid-dependent growth potential. Recently we have found that the use of levonorgestrel-releasing intrauterine system (IUS) is effective in the long-term contraception and management of menorrhagic women with uterine myomas because of a striking reduction in menorrhagia. These clinical experiences prompted us to characterize the effects of progestin on the proliferation and apoptosis of leiomyoma cells cultured in vitro. As epidermal growth factor (EGF) has been shown to mediate estrogen action and play a crucial role in regulating leiomyoma growth, we also investigated the effects of sex steroids on EGF and EGF receptor (EGF-R) expression in leiomyoma cells. In cultures of leiomyoma cells, the addition of either E(2) (10 ng/ml) or P(4) (100 ng/ml) resulted in an increase in proliferating cell nuclear antigen (PCNA) expression in the cells; whereas in cultures of normal myometrial cells, the addition of E(2) augmented PCNA expression in the cells, but P(4) did not. Immunoblot analysis revealed that leiomyoma cells contained immunoreactive EGF and that P(4) treatment resulted in an increase in EGF expression in the cells. In contrast, E(2) treatment augmented EGF-R expression in cultured leiomyoma cells, but P(4) did not. These results indicate that P(4) up-regulates the expression of PCNA and EGF in leiomyoma cells, whereas E(2) up-regulates the expression of PCNA and EGF-R in those cells. It is, therefore, conceivable that P(4) and E(2) act in combination to stimulate the proliferative potential of leiomyoma cells through the induction of EGF and EGF-R expression. We also found that Bcl-2 protein, an apoptosis-inhibiting gene product, was abundantly expressed in leiomyoma relative to that in normal myometrium, suggesting that the abundant expression of Bcl-2 protein in leiomyoma cells may be one of the molecular bases for the enhanced growth of leiomyoma relative to that of normal myometrium in the uterus. Furthermore, Bcl-2 protein expression in leiomyoma cells was up-regulated by P(4), but down-regulated by E(2). Therefore, it seems likely that P(4) may also participate in leiomyoma growth through the induction of Bcl-2 protein in leiomyoma cells. PMID- 11108864 TI - Hormonal regulation of apoptosis in breast cells and tissues. AB - Few studies have referred to the implication of apoptotic processes following hormonal treatment. No data are available on the effects of progesterone in breast cells. In order to gain insights on the effects of the gonadal steroids and antiestrogens in breast cells, we have carried out studies on apoptosis in different breast materials. We have developed a model of normal breast cells in cultures that remain hormone-dependent. On these cells and in some hormone dependent breast cancer cell lines (T-47-D, ZR75-1, MCF-7) we have observed an antiapoptotic effect of estradiol (E(2)) and a potent proapoptotic effect of some antiestrogens. Progestins were also proapoptotic in normal as well as in hormone dependent breast cancer cells. In order to understand the mechanisms of these hormones on apoptosis, we studied the bcl-2 family proteins. We demonstrated that E(2) increased the antiapoptotic proteins, bcl-2 and bclx(L), whereas, the progestins drastically decreased bcl-2 expression and weakly bclx(L) levels. We investigated the mechanisms by which E(2) increased bcl-2 expression. Our results using quantitative RT-PCR showed that E(2) increased bcl-2 mRNA levels at 48 h of treatment via a transcriptional mechanism. None of the hormone treatments altered the proapoptotic protein levels, bax and bak. We also studied the in vivo expression of bcl-2 and other members of its family in biopsies of normal breast tissues according to the menstrual cycle. Bcl-2 displayed a strong cyclical variation and seemed to be the most hormone-dependent member of the family. PMID- 11108865 TI - Ovarian steroids in endometrial angiogenesis. AB - Angiogenesis, the sprouting of new blood vessels from pre-existing ones, is fundamental for human endometrial development and differentiation, which are necessary for implantation. This vascular process is supposed to be mainly mediated by the vascular endothelial growth factor (VEGF), also named vascular permeability factor (VPF). We report here the expression and modulation of VEGF and its receptors, Flk-1/KDR and Flt-1, in the functionalis throughout the menstrual cycle. Using immunocytochemistry, VEGF is localized in glandular epithelial cells and in the surrounding stroma, as well as in capillaries and spiral arterioles. The localization of VEGF on the endothelium correlates with the presence of Flt-1 and Flk-1/KDR receptors on vascular structures, including capillary strands that have not yet formed a lumen and that have been previously described in tumors as angiogenic capillaries. The strongest immunoreactivity for both VEGF and Flk-1/KDR receptor on endothelial cells is detected in the proliferative and midsecretory phases. Enhanced expression of VEGF and its Flk-1 receptors on narrow capillary strands during the proliferative phase may account for the rapid capillary growth associated with endometrial regeneration from the residual basal layer following menstrual shedding of the functionalis. The vascular expression of Flt-1 is more important in the secretory than in the proliferative phase, associated with a high microvascular density and an increase in vascular permeability in the implantation period. Consistently with these in vivo observations, the treatment of isolated endometrial stromal cells with estradiol (E(2)), or E(2) + progesterone, significantly increased VEGF mRNA over the control value in a dose-dependent manner. These results demonstrate that the expression of VEGF and its receptors is cyclically modulated by ovarian steroids, and that this endothelial growth factor acts on the endothelium in a paracrine fashion to control endometrial angiogenesis and permeability. PMID- 11108866 TI - Progesterone as a neuroactive neurosteroid, with special reference to the effect of progesterone on myelination. AB - Some steroids are synthesized within the central and peripheral nervous system, mostly by glial cells. These are known as neurosteroids. In the brain, certain neurosteroids have been shown to act directly on membrane receptors for neurotransmitters. For example, progesterone inhibits the neuronal nicotinic acetylcholine receptor, whereas its 3alpha,5alpha-reduced metabolite 3alpha, 5alpha-tetrahydroprogesterone (allopregnanolone) activates the type A gamma aminobutyric acid receptor complex. Besides these effects, neurosteroids also regulate important glial functions, such as the synthesis of myelin proteins. Thus, in cultures of glial cells prepared from neonatal rat brain, progesterone increases the number of oligodendrocytes expressing the myelin basic protein (MBP) and the 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase). An important role for neurosteroids in myelin repair has been demonstrated in the rodent sciatic nerve, where progesterone and its direct precursor pregnenolone are synthesized by Schwann cells. After cryolesion of the male mouse sciatic nerve, blocking the local synthesis or action of progesterone impairs remyelination of the regenerating axons, whereas administration of progesterone to the lesion site promotes the formation of new myelin sheaths. PMID- 11108867 TI - Neuroendocrine effects of progesterone. AB - Progesterone (P) is secreted by the corpus luteum under the control of gonadotropin releasing hormone (GnRH)/luteinizing hormone (LH). Progesterone (P) is essential for reproduction because: (1) it induces in the endometrium the transcription of specific genes involved in the implantation of the blastocyst, (2) it modulates GnRH/LH secretion by decreasing GnRH pulse frequency, which in turn enriches the gonadotroph cells in FSH and avoids a second LH surge. Using the ewe as a model, we investigated the immediate GnRH and LH responses to acute changes of circulating P levels. Our results show that P changes cause dramatic modifications in GnRH pulse frequency: P removal induces an acceleration of the pulse generator, while P administration slows the pulse frequency. LH secretion was modified in parallel to the changes in GnRH. Other experiments proved that these neuroendocrine effects of P are mediated by P itself, not by its hydroxylated metabolites, and occur at the level of P receptors. Finally, these effects require priming by estradiol. Additionally, in the final stage of the follicular phase, P plays a role in the triggering of the LH surge. This has been shown in rodents, non-human primates, and in women. Such a phenomenon is not observed in ewes, although in these species luteal P modulates the amplitude of the estradiol-induced LH surge. PMID- 11108868 TI - Nomegestrol acetate and vascular reactivity: nonhuman primate experiments. AB - Prevention of coronary artery disease has been recognized as a major benefit of estrogen replacement therapy (ERT) in postmenopausal women. However, endometrial hyperplasia induced by unopposed ERT has raised important safety concerns. Progesterone or synthetic progestins have been used in combined hormone replacement therapy (HRT) to prevent endometrial cancer risk. Therefore, a major concern has been to ensure that the vascular beneficial effects of estrogens are not opposed when combined with progestins. Nomegestrol acetate (NOMAC) is an orally active progestin widely prescribed for HRT. Its vascular effects were evaluated in two models of coronary vascular reactivity in primates: 1) the paradoxical vasoconstriction to acetylcholine (Ach) coronary infusion after 5 months of mildly atherogenic diet in ovariectomized (OVX) Cynomolgus monkeys and 2) the pharmacologically evoked coronary vasospasm in the OVX Rhesus monkey. In the first model, after 3 months of continuous oral administration in the diet at 0.1 mg/kg/day, E2 prevented the paradoxical response to Ach, alone as well as combined with 0.25 mg/kg/day NOMAC, whereas NOMAC counteracted the endometrial stimulation. In the second model, after one artificial cycle consisting of 28 days of E2 subcutaneous (s.c.) implant and of daily oral gavage with 1 mg/kg/day of NOMAC for the last 14 days, no vasospasm (0 of 11 tested animals) occurred when the complete challenge protocol, including serotonin and the thromboxane agonist U46619, was administered to OVX Rhesus monkeys. In the balanced crossover design, identical artificial cycles with medroxyprogesterone acetate (MPA) at the same dose resulted in 7 vasospasms in 12 animals. In parallel, effective progestative activity was demonstrated by a secretory pattern in endometrial sections obtained at the end of the cycle. In these two nonhuman primate cardiovascular models, NOMAC did not have the negating effects observed with MPA. PMID- 11108869 TI - Nestorone: a progestin with a unique pharmacological profile. AB - Nestorone(R) (Nestorone 16-methylene-17alpha-acetoxy-19-norpregn-4-ene-3,20 dione), formerly referred to as ST 1435, is a potent progestin when given parenterally via sustained release formulations. The pharmacological profile of Nestorone was compared with that of levonorgestrel and 3-keto-desogestrel by steroid receptor binding studies and by in vivo bioassays in rats and rabbits. 3 Keto-desogestrel showed the highest binding affinity to progesterone receptors (PR) followed by Nestorone, levonorgestrel, and progesterone. The binding affinity of Nestorone to androgen receptors (AR) was 500- to 600-fold less than that of testosterone. However, both levonorgestrel and 3-keto-desogestrel showed significant binding (40 to 70% of testosterone) to AR. None of the progestins bound to estrogen receptors (ER). The progestational activity of Nestorone, levonorgestrel, and progesterone was compared using McPhail index in immature rabbits and pregnancy maintenance and ovulation inhibition tests in rats after subcutaneous (s.c.) administration. In all three tests, Nestorone was the most potent progestin. The progestational activity of Nestorone was also compared after oral and s.c. administration in rabbits. The potency of Nestorone was over 100-fold higher upon s.c. administration than via the oral route. The androgenic activity of progestins, based on the stimulation of ventral prostate (androgenic target) and levator ani (anabolic target) growth in castrated immature rats, showed good correlation with their binding affinity to AR. Nestorone showed no androgenic or anabolic activity. Nestorone did not bind to sex hormone binding globulin (SHBG), whereas both levonorgestrel and 3-keto-desogestrel showed significant binding to SHBG. The estrogenic/antiestrogenic activity of Nestorone was investigated in immature ovariectomized rats. In contrast to estradiol and levonorgestrel, Nestorone showed no uterotropic activity in ovariectomized rats. Despite significant binding to glucocorticoid receptors (GR), Nestorone showed no glucocorticoid activity in vivo. It is concluded that a strong progestational activity, combined with lack of androgenic, estrogenic, and glucocorticoid-like activities, confer special advantages to Nestorone for use in contraception and hormone replacement therapy. PMID- 11108870 TI - In vitro characterization of trimegestone: a new potent and selective progestin. AB - Trimegestone (TMG) is a novel 19-norpregnane progestin under development for hormone replacement therapy and oral contraception. The objective of the current study was to characterize the potency and steroid receptor selectivity of TMG in several in vitro assays and to compare its activity to that of medroxyprogesterone acetate (MPA). TMG and MPA had a similar competitive binding affinity for human and rabbit progesterone receptor (PR). However, TMG had a significantly higher affinity for rat PR (IC(50) = 3.3 nM) than MPA (IC(50) = 53.3 nM). In T47D cells, both compounds increased alkaline phosphatase activity and cell proliferation with comparable potencies (EC(50s) of 0.1 nM and of 0.02 nM, respectively). To further characterize the progestational activity and steroid receptor selectivity, we established an immortalized human endometrial stromal cell line (HESC-T). This cell line lacks endogenous estrogen receptor (ER) and PR but does have functional glucocorticoid receptors (GR). When ER is transiently expressed in the cells, 17beta-estradiol (E(2)) induces PR, allowing the study of PR-regulated genes. In HESC-T cells expressing exogenous ER, and therefore PR, both TMG and MPA increased HRE-tk-luciferase activity tenfold with an EC(50) of 0.2 nM. In HESC-T cells without exogenous ER, and therefore no PR, TMG did not induce HRE-tk-luciferase activity, whereas MPA induced the reporter activity with an EC(50) of about 10 nM. This MPA-induced reporter activity is believed to be mediated through GR. The steroid receptor selectivity of TMG was further evaluated using the HRE-tk-luciferase assay in the human lung carcinoma cell line A549, which contains GR but no PR. In these cells TMG had no effect on luciferase activity, whereas MPA increased the reporter activity in a dose dependent manner with an EC(50) of approximately 30 nM. Furthermore, HRE-tk luciferase assay in mouse fibroblast cell line L929, which expresses androgen receptor (AR) but no PR, showed that TMG had weak antiandrogenic activity whereas MPA had androgenic activity. In summary, data from several in vitro assays demonstrate that TMG is a potent progestin with a better receptor selectivity profile than MPA. PMID- 11108871 TI - Pharmacokinetics of the progesterone-containing vaginal tablet and its use in assisted reproduction. AB - Natural progesterone, which is devoid of androgenic activity, is widely used in assisted reproduction for luteal and pregnancy support. The vaginal route has become the most established way to deliver natural progesterone because it is easily administered, avoids liver first-pass metabolism, and has no systemic side effects. The vagina has a large potential for absorption, and through the 'uterine first-pass effect' vaginal administration results in higher uterine progesterone concentrations. We have investigated the pharmacokinetics of natural progesterone in the form of a vaginal tablet. A single dose of 100 mg resulted in a mean C(max) of 31.53 +/- 9.15 nmol/l with a T(max) of 6.92 +/- 3.12 h. The terminal half-life was 16.39 +/- 5.25 h. The pharmacokinetic data are discussed in relation to dose, age, and estrogen priming. Single-dose pharmacokinetics of 100 mg of progesterone vaginal tablets and gelatin capsules were evaluated over 24 h. Results indicated a similar mean T(max) of 6.92 +/- 3.12 and 6.23 +/- 6.57 h, respectively. However, a significantly higher C(max) was achieved by the vaginal tablet (31.95 +/- 9.15 and 23.85 +/- 9.57 nmol/l, respectively, P < 0.05). Continuous use of vaginal progesterone did not influence the hormonal, liver, or lipid profiles evaluated. There was no case of endometrial hyperplasia. The vaginal tablet was found to be well-tolerated, safe, and easily administered. In conclusion, progesterone-containing vaginal tablets have good pharmacokinetic properties and should be used for progesterone supplementation in IVF. PMID- 11108872 TI - Progestins and cardiovascular risk markers. AB - Several risks are attributed to progestins as a class-effect; however, the progestins used in hormone replacement therapy (HRT) have varying pharmacologic properties and do not induce the same side effects. Natural progesterone (P) and some of its derivatives, such as the 19-norprogesterones, do not exert any androgenic effect and, hence, have no negative effect on the lipids. On the other hand, the 19-nortestosterone derivatives and even some 17-hydroxyprogesterones have a partial androgenic effect, which may explain some of the negative effects observed on surrogate markers of cardiovascular risk. The relevance of the lipid changes induced by sex steroids has been questioned, and studies in the female cynomolgous monkey have not shown a direct relationship to atherosclerosis. Results suggest that estrogens (E) have antiatherogenic effects and that P does not reverse the beneficial effect of estradiol. Also, sex hormones modulate the vasomotor response of the main arteries. E preserves the normal endothelium mediated dilation of coronary arteries, and P does not reverse this potential cardioprotective mechanism. In the same animal model, the addition of cyclic or continuous medroxyprogesterone acetate (MPA) to E inhibited vasodilatation by 50%, while nomegestrol acetate did not diminish the E-induced vasodilatation. Not all progestins act similarly on vasomotion or affect cardiovascular risk factors in the same way. Progestins, such as MPA or norethisterone acetate (NETA), exert a partial detrimental effect on the beneficial actions of estrogens with regard to lipid changes, atheroma development, or vasomotion. In contrast, progesterone itself does not have this inhibitory effect on lipid changes and vascular reactivity in animal models or on exercise-induced myocardial ischemia in humans. Nonandrogenic molecules of P itself and of derivatives, such as 19 norprogesterones, would appear neutral on the vessels. Several ongoing randomized controlled trials of HRT are focusing on primary or secondary prevention of coronary heart disease. Unfortunately, most of these large trials have selected the same HRT regimen for their study design. Further studies with other treatment regimens are thus needed and should consider the various steroids used in different countries. PMID- 11108873 TI - Progestins and menopause: epidemiological studies of risks of endometrial and breast cancer. AB - Estrogen replacement therapy (ERT) increases a woman's risk of developing endometrial cancer approximately 120% for each 5 years of use. ERT increases a woman's risk of developing breast cancer approximately 10% for each 5 years of use. To reduce the greatly increased endometrial cancer risk, progestins have been added to ERT (estrogen-progestin replacement therapy; EPRT) for between 5 and 15 days (usually 7 or 10 days) per month in a sequential fashion (sequential EPRT; SEPRT) or with each dose of ERT (continuous-combined EPRT; CEPRT). We conducted two large case-control studies in postmenopausal women in Los Angeles to evaluate the effects of these changes on endometrial and breast cancer risks. As expected CEPRT was not associated with any increased risk of endometrial cancer. SEPRT with the progestin being given for 10 days per month also did not increase endometrial cancer risk. SEPRT with the progestin being given for 7 days per month did increase endometrial cancer risk with only a relatively slight reduction in risk compared to ERT effectively proportional to the reduction in the number of days of unopposed estrogen. The sharp contrast between the effects of 7 days and 10 days of progestin in SEPRT suggests that the extent of endometrial sloughing or of 'terminal' differentiation at the completion of the progestin phase may play a critical role in determining endometrial cancer risk. This may provide an explanation of why endometrial cancer risk increases so sharply with age in young women even in countries where obesity-associated anovulation is very uncommon; extended periods of unopposed estrogen is not an explanation but less than 10 days of an 'adequate' progesterone level may be. EPRT significantly increased the risk of breast cancer. EPRT was associated with an approximately 24% increase in risk for each 5 years of use; the effect was some 212-fold greater than the effect of ERT, which we had previously predicted on theoretical grounds. This effect could also be predicted from the results on mammographic densities seen in the PEPI randomized trial of different forms of hormone replacement therapy (HRT). In the PEPI trial EPRT increased mammographic densities to a much greater extent than ERT. Progestins need to be given to protect the endometrium. They need to be delivered to the endometrium in a manner that will have the least effect on the breast. This can be carried out by using a vaginal or direct endometrial route of administration. The vaginal route will provide adequate endometrial progestin levels with low blood levels so that the effects of the progestin on the breast should be small; with the direct endometrial route the blood progestin levels are even lower, and the effects of the progestin on the breast will be effectively zero. If this is unacceptable to a woman, then giving progestins by mouth (or transdermally) for 10 days every 3 to 4 months should provide satisfactory protection of the endometrium when used with standard-dose conjugated estrogen (CE). This regimen has much less effect on the breast than monthly SEPRT or CEPRT. Two clinical trials of 10 mg per day of MPA for 14 days every 3 months and 0.625 mg/day of CE have been published. Both studies suggest that this approach may be satisfactory in that the extent of hyperplasia was minimal. More studies of this approach are urgently needed. PMID- 11108874 TI - Endometrial vascular changes and bleeding disturbances with long-acting progestins. AB - Unscheduled disturbances of bleeding with long-acting progestogen-only methods continue to be a major social inconvenience, and the most common reason for premature discontinuation of use. The patterns of bleeding with different methods are now well characterised, but in spite of a substantial amount of recent research we still do not clearly understand the actual mechanisms underlying these disturbances. The major changes occur locally within the endometrium, although these are influenced substantially by different dosages and regimens of contraceptive steroids. Hysteroscopic studies have demonstrated major changes in the extent of superficial endometrial vascularization, disturbances in superficial vascular morphology, and increased endometrial vascular and epithelial fragility in progestogen-only users. There may also be alterations in endometrial perfusion and vasomotion. Laboratory studies of cellular and molecular mechanisms have shown changes in vascular basement membranes, pericytes, migratory leukocytes, cellular architecture, cell adhesion molecules, and intercellular matrix breakdown systems, such as matrix metalloproteinases. It appears that continuous exposure to progestogen, in the presence of low to moderate levels of estrogen, results in an endometrium that histologically demonstrates 'suppressed secretory' changes, associated with disturbed angiogenesis, and a tendency to release molecules capable of causing focal endometrial epithelial and endothelial damage and bleeding. PMID- 11108875 TI - Progesterone and progestins: applications in gynecology. AB - Achievements obtained in infertility treatments over the past two decades have sparked interest in optimizing progesterone administration. Although progesterone is absorbed orally when ingested in micronized form, bioavailability is poor because of extensive liver metabolism. This explains why full predecidual transformation of the endometrium cannot be achieved with oral progesterone and is therefore ineffective for luteal support in in vitro fertilization (IVF). Progesterone administered non-orally can duplicate the endometrial changes normally seen in the menstrual cycle in women whose ovaries are inactive. Similar results have been reported with intramuscular (i.m.) injections and vaginal administration, although tissue levels are higher in the latter case. The recent development of a controlled and sustained release vaginal progesterone gel, Crinone(R) 8%, has made the vaginal route clinically practical by limiting the number of necessary applications to 1 per day. This regimen has been found at least as effective as intramuscular (i.m.) injections in women whose ovaries are inactive (donor egg IVF) and for luteal support in regular IVF. Hence, painful daily i.m. injections of progesterone in oil become unnecessary. The possibility of reducing the number of daily applications of vaginal progesterone to 1 per day, made possible by the sustained release gel Crinone, has opened new possibilities for long-term treatments, as in hormone replacement therapy (HRT). The low incidence of systemic side effects with use of the vaginal progesterone gel used for HRT in amenorrheic women, contrasts with findings related to use of synthetic progestins. Preliminary data suggest that vaginal progesterone can be instrumental in enhancing the notoriously poor long-term compliance of HRT. PMID- 11108876 TI - Progestin implants. AB - Progestin implants for contraception are highly effective, safe, and the most convenient choice for many women. Progestin implants currently on the market, preparing for launch, or under investigation are reviewed here. Their basic galenic and pharmacokinetic features, as well as their contraceptive effectiveness, are described. The first progestin-only contraceptive implant placed on the market was Norplant, a multiunit system. Since then, several single and double-rod implants have been developed, each using one of four different progestins: levonorgestrel, etonogestrel, Nestorone and nomegestrol acetate. Jadelle is similar to Norplant but consists of only two, rather than six, Silastic rods to simplify insertion and removal; nevertheless, levonorgesterel serum levels are identical, and performance is the same for both systems. The single implant systems reviewed here are: Implanon with a 3-year duration; Nestorone implants for breast feeding and non-breast feeding women lasting up to 2 years; and Uniplant, which is effective for 1 year. The advantages and disadvantages of progestin implants, the importance of counseling for increasing user satisfaction, and the future outlook for this contraceptive method are also discussed. PMID- 11108877 TI - Progestin-only contraceptive rings. AB - Several progestin-only long acting contraceptives are currently available in the form of implants or injectables. Vaginal rings are another contraceptive option in the final stages of development. These steroid-containing polymer rings are placed in the vagina, providing relatively constant drug release, thus allowing for lower effective doses. Vaginal rings have the advantage of being user controlled and non-provider dependent, and their use is non-coital related. The first clinical study with medroxyprogesterone acetate vaginal rings was published in 1970. Since then numerous clinical trials testing different steroids and doses have followed. A large Phase III multicenter clinical trial with a levonorgestrel ring, releasing 20 microg/day, was coordinated and sponsored by WHO. The cumulative one-year pregnancy rate was 4. 5%. The principal reasons for discontinuation were menstrual disturbances (17.2%), followed by frequent expulsion of the ring (7. 1%), and vaginal symptoms (6.0%). The finding of erythematous lesions in the vagina in some women has led to the development of a more flexible device. Collaboration with industry should facilitate the manufacture of a redesigned levonorgestrel ring with a higher release rate. The Population Council is also developing a vaginal ring containing Nestorone for 6 months of continuous use. Ovulation inhibition was achieved in over 97% of the segments studied, with rings releasing either 50, 75, or 100 microg/day. No pregnancies occurred in women using the low-dose ring, while one pregnancy each occurred in the intermediate- and high-dose ring groups for a 6-month cumulative pregnancy rate of 0.0, 1.9, and 2.1%. Bleeding irregularities were common. Nestorone is orally inactive; therefore this ring is also excellent for use in lactating women. PMID- 11108878 TI - The levonorgestrel intrauterine system in contraception. AB - The levonorgestrel intrauterine system (LNG IUS) releases 20 microg/24 h of levonorgestrel from a polymer cylinder mounted on a T-shaped frame and covered with a release rate-controlling membrane. It is approved for 5-year use. The most outstanding features of LNG IUS are its high contraceptive efficacy and reduction of menstrual blood flow. No single mode of action can account for its contraceptive efficacy. The endometrium becomes thin and inactive, and the cervical mucus turns scanty and viscous. Although ovulation may be disturbed to some degree, estradiol production continues normally. The Pearl index for LNG IUS from large clinical trials is 0.1. Extrauterine pregnancies occur in 1 in 5000 users per year. Both the volume of menstrual blood loss and the number of bleeding days are reduced. During the first year of use, 20% of women become amenorrheic. There is an initial increase in the mean number of bleeding and spotting days, but in 3 to 6 months the number of bleeding and spotting days is the same as observed in copper IUD-users. The variation between individuals is wide and unpredictable. There are also additional health benefits secondary to the inactivation of the endometrium: increased hemoglobin, decreased dysmenorrhea, a possible decrease in pelvic inflammatory disease. LNG IUS may also decrease the growth of fibroids. LNG IUS is well accepted by users, with typical annual continuation rates above 80% in clinical studies. PMID- 11108879 TI - The levonorgestrel intrauterine system: therapeutic aspects. AB - Use of the levonorgestrel-releasing intrauterine system (LNG IUS) is associated with a strong reduction in the number of days of bleeding and menstrual blood loss. This effect is based on the marked local action of the intrauterine release of levonorgestrel (LNG) on the endometrium. In suppressed endometrium the production of many highly active compounds ceases. On the other hand, LNG stimulates the synthesis of some regulatory proteins in the endometrium. Reduction of menstrual blood loss results in improvement of the body iron balance and in an increase in hemoglobin concentration. The LNG IUS has been used in the prevention and treatment of iron deficiency anemia. Many studies have demonstrated that the LNG IUS is effective in the treatment of menorrhagia. Reduction of excessive blood loss is seen as soon as the first menstruation after insertion, and at 1 year the reduction is more than 90%. The therapeutic effect is maintained for more than 5 years after first placement of the LNG IUS in the uterine cavity. Correct insertion is essential, and complications and side effects are rare; fertility is preserved, and invasive procedures such as endometrial ablation or hysterectomy and hospitalization are avoided. The third major indication for therapeutic use is in protection of the endometrium in estrogen replacement therapy during peri- and postmenopausal years. A fundal position of the system in the uterine cavity results in reduction of bleeding, and an increasing number of women have no bleeding at all during use of the IUS. Acceptance and continuation of use of the LNG IUS in hormone replacement therapy (HRT) have been high. PMID- 11108880 TI - Vaginal rings for contraception in lactating women. AB - Contraceptive methods for breastfeeding women should be safe for the mother and infant and should not interfere with lactation. Progestin-only methods meet these conditions and can be used from the sixth week postpartum. Because all progestins are excreted in milk, those that are insufficiently active by the oral route are preferable to avoid any possible effect on the baby. These steroids, however, must be administered to the mother by a non-oral route. Initially, progesterone was administered subdermally to test this concept. Subsequently, a progesterone vaginal ring was developed to be used continuously for 3 to 4 months and replaced with a new device, as needed, until weaning. Clinical trials have shown a high contraceptive efficacy (over 98.5%) and safety. The gross continuation rate of this method is approximately 40% at 12 months of use, with use-related problems being the main reason for discontinuation (26.8%). Currently, a Nestorone vaginal ring is under development, delivering 50 microg of Nestorone per day. It may be used continuously for up to one year, even if weaning occurs earlier. Both of these progestin-only rings prolong lactational amenorrhea to 10 to 12 months, which represents a health benefit and convenience for many women. The registration of the progesterone vaginal ring, developed as a contraceptive method to be used exclusively during lactation, has been approved in Chile and Peru. The fact that it is a user-controlled long-term contraceptive that delivers a natural hormone makes it an attractive option for many women. PMID- 11108881 TI - The return of the pharmaceutical industry to the market of contraception. AB - In the 1980s and 1990s, the litigious climate in the US had a catastrophic effect on sales of many major contraceptives. Although oral contraceptives escaped controversy, the intrauterine device (IUD) and Norplant(R) were two targets of damaging litigation. The IUD was withdrawn from the market in 1985. Since 1994 when the attacks began against Norplant, its US sales have dramatically declined, even though no fault has been found in the method or its development. In general, pharmaceutical companies were extremely hesitant to develop new contraceptives during this period. The bleak outlook, however, began to shift in the late 1990s, as fertility rates began to decrease worldwide and contraceptive users increased. By 2025, 2500 million women will comprise the customer base for contraception. Global pharmaceutical companies are now participating in expanding markets overseas and have launched and continue to develop a range of new long-term reversible, and highly effective, contraceptive products outside the traditional oral contraceptive field. Two new contraceptives on the way to the US market are: Mirena, a levonorgestrel-releasing intrauterine system manufactured by Schering Leiras; and Implanon, a single implant system manufactured by Organon of the Netherlands. Other birth control methods soon to be launched include: emergency contraceptives, the contraceptive patch, monthly contraceptive injections, mifepristone for medical abortion, and modified oral contraceptives. PMID- 11108882 TI - Endocrine pharmacological characterization of progesterone antagonists and progesterone receptor modulators with respect to PR-agonistic and antagonistic activity. AB - Studies were conducted to assess progesterone antagonists (PAs) and progesterone receptor modulators (PRMs) with respect to PR agonistic and antagonistic activities in vivo. These properties are not always adequately reflected in transactivation in vitro models. Studies were performed in pregnant rats, estrogen-primed rabbits (McPhail -Test), and cycling and pregnant guinea pigs. Tested compounds included mifepristone (RU486), onapristone, J867, J956, J1042, and ZK137316. J-compounds induced sub-maximum endometrial transformation and, paradoxically, inhibited effects of progesterone in rabbits. Mifepristone, onapristone, and ZK137316 behaved as 'pure' antagonists in this species. Inhibition of uterine PGF(2alpha) secretion and inhibition of luteolysis in cycling guinea pigs were more sensitive parameters of PR-agonistic and antagonistic properties. 'Pure' PAs inhibited uterine PGF(2alpha) secretion and luteal regression completely. The PR agonist R5020 reversed both effects which demonstrates a PR mediation. Agonistic PRMs (J-substances and mifepristone) showed no or blunted antiluteolytic effects compared to the 'pure' PR antagonist onapristone. When tested in pregnant guinea pigs for their labor-inducing potential, PR agonistic PRMs had much reduced or abolished abortifacient activity compared to mifepristone (mifepristone > J956 > J867/J912 > J1042). However, in cycling animals, superior antiovulatory and antiproliferative properties of the J substances were seen. Antiovulatory effects of 'pure' and agonistic PRMs are probably due to different mechanisms. The relevance of rodent studies for antiovulatory and uterine antiproliferative effects for the human is still uncertain. The non-abortifacient PRM J1042 induced stromal compaction and inhibition of endometrial proliferation in monkeys, but this effect was not stronger than that of the 'purer' PAs. 'Pure' PAs are important pharmacological tools analogous to PRKO models to study the role of PR in the menstrual cycle and in pregnancy. PMID- 11108883 TI - Nonsteroidal progestins and antiprogestins related to flutamide. AB - From the dual progestin/antiandrogenic properties of certain synthetic steroids (e.g. cyproterone acetate), it was apparent that the progesterone (P) and androgen (A) receptors must have some common ligand binding features. The nonsteroidal antiandrogen (aA) hydroxyflutamide was therefore considered a possible starting point for medicinal chemistry aimed at antiprogestin (aP) activity. Various modifications to the side chain and aryl ring substituents of flutamide yielded both P and aP activity, but always coupled with varying degrees of A or aA activity. Mineralocorticoid activity was present in some structures, but glucocorticoid and antiglucorticoid activities were not detected. Species (rat, rabbit and monkey) and chiral differences presented formidable difficulties in developing simple structure activity patterns, and low ( < 1%) in vitro uterine receptor binding belied in vivo potency of some aPs. One of the most active aPs, ZM172406, the R enantiomer of ZM150271, N-(3-chloro-4-cyanophenyl) 3,3, 3-trifluoro-2-hydroxy-2-methylpropanamide, had comparable oral potency to mifepristone in rats and monkeys. The racemate ZM150271 was an effective abortifacient during early pregnancy in pigtailed monkeys (3 x 10 mg/kg) but less effective in cynomolgus monkeys. One of the most active progestins (Pn), ZM182345, N-(4-nitro-3-trifluoromethylphenyl)-4-phenyl-2-hydroxy-2-trifluoromet hyl-pentanamide, was at least as potent as P in rats and rabbits but also possessed A activity. PMID- 11108885 TI - Antiproliferative effects of progesterone antagonists and progesterone receptor modulators on the endometrium. AB - Progesterone antagonists (PAs, antiprogestins) can modulate estrogenic effects in various estrogen-dependent tissues. These modulatory effects are complex and depend on species, tissue, type of compound, dose, and duration of treatment. In non-human primates, PAs, including mifepristone, ZK 137 316 and ZK 230 211, inhibit endometrial proliferation and induce amenorrhea. When administered chronically at relatively low doses, these compounds block the mitotic activity of endometrial epithelium and induce stromal compaction in a dose-dependent manner in both spayed and intact monkeys at high estradiol concentrations. These effects were accompanied by an atrophy of spiral arteries. The antiproliferative effects were endometrium-specific, since the estrogenic effects in the oviduct and vagina were not inhibited by PAs. Similar endometrial antiproliferative effects were also found after treatment with the progesterone receptor modulator (PRM), mesoprogestin J1042. The endometrial antiproliferative effects of PAs, particularly within the endometrial glands, were also observed in spayed rabbits. In spayed rats, however, the PAs did not inhibit, but rather enhanced, various estrogen responses, including endometrial proliferation, pointing to species specific differences. In conclusion, our studies indicate that both pure PAs and PRMs selectively inhibit estrogen-dependent endometrial proliferation in the primate endometrium without affecting estrogenic response in other estrogen dependent tissues or inducing unscheduled bleeding. Our studies indicate that the spiral arteries, which are unique to the primate endometrium, are the primary targets that are damaged or inhibited by PAs and PRMs. The damage to these unique vessels may underlay the paradoxical, endometrium-specific, antiproliferative effects of these compounds. Hence, the properties of PAs and PRMs (mesoprogestins) open up new applications in gynecological therapy and hormone replacement therapy. PMID- 11108884 TI - Preclinical experience with two selective progesterone receptor modulators on breast and endometrium. AB - Org 31710 and Org 33628 are two highly selective progesterone receptor modulators (PRMs) with respect to their anti-progestational and anti-glucocorticoid activity. The compounds have been studied both in vitro and in vivo. Org 33628 has approximately four times stronger anti-progestational activity in vitro than does Org 31710, and in rats it is about 15 times more potent in the pregnancy interruption test. Two main indications for the use of PRMs are breast cancer and fertility regulation. The effects of both Org 31710 and Org 33628 were tested in relevant models for these indications. The effects of the two compounds on breast tumor development were assessed and in rats using the DMBA model. Their potency in menses induction was tested in monkeys on a 4-day regimen in the luteal phase, and after a single dose at day 21 of the normal cycle, and under a continuous progestin treatment using desogestrel. The compounds were also tested alone in a continuous low-dose regimen. The effects on follicular development and ovulation were determined by measuring estradiol and progesterone levels. Cycle control was monitored by daily vaginal swabs. In the DMBA model, Org 31710 at oral doses of 0.8, 2.0, and 5.0 mg/kg showed a clear dose-related reduction in tumor load. With the two highest doses, an even lower tumor load was seen after a 3-week treatment period compared to the tumor load at the start of treatment. Org 33628 showed a similar efficacy as Org 31710 at a dose of 2.0 mg/kg. RU 486 after oral treatment was two times less potent in this model than Org 31710 and Org 33628. The efficacy of menses induction using the 4-day regimen is dependent on the time of administration relative to the progesterone peak in the luteal phase. The highest efficacy is achieved in the descending part of the peak, at which a 100% success rate is found with a dose of 1 mg/kg of either Org 31710 or Org 33628. In Cynomolgus monkeys, at a single dose of 15 mg/kg of Org 31710 or Org 33628 in the luteal phase, menses induction was achieved only in 60% of the treatment cycles. Surprisingly menses induction can be achieved with a single dose that is about a ten-times lower when the monkeys are treated continuously with desogestrel. Cycle control is better at low than at high doses of antiprogestin in combination with daily dosing of 4 microg/kg desogestrel. Despite the difference in receptor affinity, no difference between Org 31710 and Org 33628 was found in menses induction. In the continuous low-dose (1 mg/kg) regimen with the PRMs, follicular development occurs normally while ovulation is inhibited. Ovulation is resumed shortly after stopping treatment, and a normal menses occurs after the first progesterone peak. Both compounds may be interesting options for the prevention and treatment of breast cancer and for fertility control. PMID- 11108886 TI - Role of progesterone on peri-implantation stage endometrium-embryo interaction in the primate. AB - Progesterone secretion during the luteal phase influences oviductal and endometrial functions which are essential for embryo viability and implantation in a number of species including primates. Luteal phase estrogen is not essential for progesterone-dependent endometrial receptivity towards implantation and pregnancy in the rhesus monkey and in the human. However, synchronous development of embryo and endometrium is an essential prerequisite for evolutive implantation. Progesterone helps to maintain synchronous development of preimplantation embryo through its action on maternal uterus. The anti-nidatory action of mifepristone, a potent progesterone receptor modulator (PRM) with pronounced antiprogestagenic activity, is known to be associated with desynchronization of endometrium along with repression of glandular secretory differentiation and vascular maturation. Thus, it is likely that early luteal phase administration of mifepristone affects paracrine action of the secretory stage endometrium on the preimplantation stage embryo, and thereby inhibits embryonic development and viability. We shall examine this hypothesis using the rhesus monkey as a primate model. PMID- 11108888 TI - The progesterone antagonist onapristone retards the advanced endometrial transformation after gonadotropin stimulation in rabbits. AB - Ovarian stimulation with gonadotropins (GN) during human in vitro fertilization and embryo transfer (IVF/ET) therapy alters the ovarian steroid output, especially that of progesterone. As a consequence, endometrial transformation is advanced, and embryo implantation is hampered. This study used the rabbit model to determine if the application of the progesterone antagonist (PA) onapristone (ONA) could retard endometrial development after GN-stimulation. Rabbits were GN stimulated twice daily with 5 IU FSH and 5 IU LH on 3 consecutive days with a) hMG (n = 10) or b) with a mixture of recombinant FSH and LH (n = 10). The animals were then mated, and hCG was injected i.v. to ensure ovulation. This day is designated as day 0 post coitum (d 0 p.c.). On day 2 p.c., five animals of each group were treated with 20 mg ONA/kg body weight and five with vehicle for control. On d 5 p.c. endometrial transformation was analyzed by morphology, uteroglobin (Ugl)-mRNA expression, and proliferation. Embryos were flushed from the uteri. Their number and morphology was evaluated. The endometrium of GN stimulated control animals demonstrated very long endometrial glands and narrow stromal septa. Ugl-mRNA expression was restricted to the cells at the bottom of the gland. 17.0 +/- 4.6% (mean +/- SD) of glandular cells and 6.0 +/- 5.3% of luminal epithelial cells proliferated. In ONA-treated animals, endometrial glands were significantly shorter, and the pattern of arborization was less pronounced. Endometrial gland cells and luminal epithelial cells expressed Ugl-mRNA. Furthermore, glandular and luminal cells proliferated with high intensity (38.6 +/- 6.8% and 36.4 +/- 9.3%, respectively). These results indicate that the status of endometrial differentiation was retarded after ONA-treatment. Nevertheless, the embryos of these ONA-treated animals were well developed. In conclusion, after GN-stimulation, ONA treatment retarded the advanced endometrial transformation in rabbits. Therefore, postovulatory administration of a PA might be a possible strategy to modulate the advanced endometrial development in IVF cycles. PMID- 11108887 TI - Progestins, progesterone receptor modulators, and progesterone antagonists change VEGF release of endometrial cells in culture. AB - The influences of the synthetic progestin, medroxyprogesterone acetate (MPA), the progesterone receptor modulator J867, and the antagonist ZK137316 were studied in vitro on isolated endometrial epithelial cells, as well as endometrial fibroblasts. We evaluated the expression of estrogen receptor alpha (ER) and the progesterone receptor (PR) by RT-PCR. ER and PR were strongly expressed in the fibroblasts and epithelial cells under treatment with 10(-8) M 17beta-estradiol (E(2)). Treatment with 10(-6) M J867 or ZK137316 upregulated the PR expression as did E(2), in contrast to treatment with 10(-6) M MPA, which caused a downregulation of PR in epithelial cells, but not in fibroblasts. In addition, the vascular endothelial growth factor (VEGF) release into the cell culture medium was analyzed by a VEGF-ELISA. VEGF which plays an important role in angiogenesis, is regulated by steroid hormones as well as hypoxia. E(2) stimulates VEGF release into the medium in both cell types. MPA reduces VEGF release significantly in the fibroblast cell culture, but increases it in the epithelial cell culture. ZK137316, in the presence or absence of E(2), reduces VEGF release in fibroblast cell culture. J867 increases the VEGF production in fibroblasts only in the presence of E(2). Both compounds show no significant effects, compared to E(2), in epithelial cell culture. The different results for the epithelial cells and fibroblasts indicate that the pharmacological effects of PR modulators (PRMs) and progesterone antagonists (PAs) may be cell specific and depend on the presence or absence of partial progestagenic agonistic activities. This observation opens up new perspectives for various clinical applications. PMID- 11108889 TI - Endometrial contraception: modulation of molecular determinants of uterine receptivity. AB - Modulation of endometrial receptivity is a promising approach for fertility regulation since it allows a contraceptive to act specifically at the endometrium. This was corroborated by our previous observations that treatment with low doses of a pure progesterone antagonist (PA, antiprogestin), onapristone (ZK 98299), in bonnet monkeys inhibited fertility by selectively retarding endometrial development, without affecting the hypophyseal-hypothalamic function. In the present study, further investigations, undertaken to analyze the molecular repertoire of a nonreceptive primate endometrium, determined expression of: steroid hormone receptors, i.e. progesterone receptor (PR) and estrogen receptor (ER); cytokines, i.e. leukemia inhibitory factor (LIF): transforming growth factor beta (TGFbeta) and its receptor (TGFbetaR); and cell adhesion molecules, i.e. integrins (alpha(v)beta(3), alpha(1)beta(1)). These studies were conducted during the different phases of the normal menstrual cycle and following treatment with different doses of onapristone (2.5 mg, 5 mg, or 10 mg every third day for one cycle) in bonnet monkeys. The molecules were analysed collectively to explore the possibility of a correlation between expression of these markers and endometrial receptivity and to investigate whether there exists a regulatory link between expression of these molecules under in vivo conditions. Three types of expression patterns of endometrial factors were observed during the peri implantation period following onapristone treatment: 1) LIF, alpha(v)beta(3), and alpha(1)beta(1) showed significant (P < 0.02) down regulation in glandular epithelium of endometria in animals treated with all three doses of onapristone as compared to the control group. This was indicative of their critical role in the progesterone-driven cascade leading to implantation. 2) PR, TGFbeta, and TGFbetaR remained unaffected in the endometria from 2.5 mg treated animals and showed down regulation in animals treated with 5 and 10 mg onapristone as compared to the control group, thereby suggesting that the expression of these markers may not truely reflect endometrial receptivity per se. However, their facilitatory role in preparing the endometrium for implantation can not be ruled out since continued perturbation in the expression of these molecules may affect endometrial growth, remodelling, and differentiation, which in turn may render the endometrium nonreceptive; 3) ER remained unaltered in endometria of animals rendered infertile with 2.5, 5, and 10 mg onapristone. This observation indirectly suggests that onapristone-induced endometrial changes are mediated via some specific mechanisms. The present study clearly demonstrates that endometrial non-receptivity induced at low doses of onapristone is associated with changes in the expression pattern of specific molecular markers. However, no direct correlation was observed between in vivo expression of TGFbeta, LIF, and integrins, thereby lending support to the concept that there exists redundancy or multiple pathways which regulate implantation events. PMID- 11108890 TI - Occurrence, regulation, and significance of progesterone receptors in human meningioma. AB - The abundant expression of progesterone receptors (PR) in human meningiomas is well established. It is unknown, however, how PR expression is regulated, especially since estrogen receptors (ER) are virtually absent in these tumors. At the mRNA level, ER splice variants occur in meningioma but these appear not to be involved in the apparently autonomous PR expression. In an earlier study, because other ER-inducible proteins were either not expressed at all (pS2) or were expressed at a very low level compared to their expression in breast cancer (Cathepsin-D), the authors have postulated that the autonomous PR expression in meningioma is PR promoter-related rather than ER-related and have studied PR expression in cultured meningioma cells. PR levels appeared to decrease rapidly in vitro in monolayer as well as in three dimensional spheroid cultures. Culture conditions thus are not yet sufficient for the quantitative evaluation of PR expression. To evaluate whether PR deterioration is associated with cell turnover (meningiomas grow much faster in vitro than in vivo), the relationship between expression of the apoptotic proteins Bcl-2 and Bax and PR expression was investigated. Bcl-2 expression was found to be highest in meningioma with low PR levels, and in breast cancer tissue with high PR levels. Bax expression was not related to PR expression in any of the two tissues. Given the potential benefit of antiprogestin treatment and the occurrence in meningiomas of a protein capable of binding to the estrogen-responsive element, the expression of PR in meningioma remains a fascinating phenomenon which requires further investigation. PMID- 11108891 TI - Pregnancy termination. AB - During pregnancy, the antiprogestin mifepristone will induce uterine contractions, increase the sensitivity of the myometrium to prostaglandin, and ripen the cervix. These effects indicate that mifepristone can be used for termination of pregnancy. The clinical experience has shown that mifepristone is sufficiently effective for this purpose only if combined with a suitable prostaglandin, e.g. gemeprost or misoprostol. The combined treatment has been used for termination of early pregnancy (up to 63 days of amenorrhea) and for termination of second trimester pregnancy. During early pregnancy, the recommended dose of mifepristone is 600 mg (although 200 mg seems sufficient), followed 36-48 h later by 0.4-0.8 mg misoprostol administered either orally or vaginally, or vaginal administration of 1.0 mg gemeprost. For termination of second trimester pregnancy, the treatment with mifepristone is most commonly combined with 1.0 mg gemeprost repeated at 3-6-h intervals. The combined treatment is as effective and safe during early pregnancy as is the alternative vacuum aspiration and is also equally acceptable if the woman is allowed to choose the method she prefers. During the second trimester, the pretreatment will significantly reduce the duration of labor, dose of prostaglandin, and the frequency of side effects. PMID- 11108892 TI - Progesterone receptor modulators and progesterone antagonists in women's health. AB - Both progesterone receptor modulators (PRMs) as well as pure progesterone antagonists (PAs) have numerous proven and potential therapeutic applications in female health care. Mifepristone, a PRM with only marginal agonistic activity, together with a prostaglandin can terminate pregnancies of less than 9 weeks duration; mifepristone is also used in the preparation of women at later gestational stages whose pregnancies are terminated with prostaglandins or surgery. Mifepristone causes expulsion of the uterine contents following intrauterine fetal death and promotes dilation of the non-pregnant primigravid uterus. It is also effective in the treatment of missed abortion. Together with methotrexate, mifepristone can be used in the medical treatment of ectopic pregnancy. Both PAs and PRMs display antiproliferative effects on the endometrium. Because of this, they have application in the treatment of endometriosis, an estrogen-dependent condition. They may also be utilized to reduce myoma size, acting as both a PA and antiproliferative agent. Unlike GnRH agonists, long-term use in endometriosis and myoma is not associated with loss of bone and hypoestrogenism. PRMs may also be useful in IVF programs to prevent a premature LH surge and to delay the emergence of the implantation window. Some PRMs have potential use as hormone replacement therapy in women during menopause or in those with dysfunctional uterine bleeding. PMID- 11108893 TI - The use of progesterone antagonists and progesterone receptor modulators in contraception. AB - Progesterone antagonists (PAs) and progesterone receptor modulators (PRMs) have contraceptive potential by suppressing follicular development, delaying the surge of luteinizing hormone (LH), retarding endometrial maturation, and promoting endometrial bleeding. Mifepristone, in daily doses of 2-10 mg, blocks the LH surge and ovulation. Many of the studies were conducted in women not at risk of pregnancy, and thus the contraceptive efficacy is not yet known. Nevertheless, there is evidence that daily doses of 2 or 5 mg of mifepristone have contraceptive potential. Because of anovulation, there may be an unopposed estrogen effect on the endometrium, although this risk may be mitigated by the noncompetitive anti-estrogenic activity exhibited by both PAs and PRMs. Low doses of PAs and PRMs, which do not affect ovulation, retard endometrial maturation, indicating that the endometrium is exquisitely sensitive to these compounds. This raises the prospect of endometrial contraception, i.e. prevention of endometrial maturation without disturbing ovulation or producing alterations in bleeding patterns. This approach works well in monkeys but was not found to be very promising when given to women not using contraception. On the other hand, 200 mg mifepristone administered 48 h after the LH surge, which has minimal or no effect on ovulation and bleeding patterns, is an effective contraceptive; yet, it is not a practical approach to contraception. Late luteal phase administration of mifepristone produces menstrual bleeding. However, when mifepristone was administered every month at the end of the cycle either alone or together with prostaglandins, it was not very effective in preventing pregnancy. In contrast, a mifepristone-prostaglandin combination has been shown to be a very effective treatment for occasional menstrual regulation, with vaginal bleeding induced in 98% of pregnant women, with menses delay of 11 days or less. Mifepristone is an excellent agent for emergency contraception when used within 120 h of unprotected intercourse. It is also possible that PAs and PRMs may be used to reduce the occurrence of bleeding irregularities induced by progestin-only contraceptive methods. Both classes of progesterone receptor ligands may also have contraceptive efficacy by having a pharmacological effect on the embryo or altering tubal transport or other aspects of tubal physiology. PMID- 11108894 TI - Progesterone antagonists and progesterone receptor modulators in the treatment of breast cancer. AB - Progesterone antagonists (PAs) (antiprogestins) or progesterone receptor modulators (PRMs) form an interesting category of new hormonal agents in the treatment of breast cancer. In vitro, antiproliferative effects of different PAs are mainly observed in estrogen-stimulated growth of PR-positive tumor cell lines. Both progestin agonist/antagonist actions on mammary tumor cells are dependent on the type of cell line, culture medium and concentrations of the PAs used, and type of biologic response measured. In various experimental animal tumor models, different PAs showed a greater antitumor activity than tamoxifen or high-dose progestins. Most interestingly, combination treatment of different PAs (mifepristone, ORG 31710, onapristone) or PRMs with different antiestrogens (tamoxifen, droloxifen, ICI 164384) or with an aromatase inhibitor (atemestane) showed greater antitumor efficacy than treatment with each single type of drug alone. These additive antiproliferative effects were demonstrated in various experimental in vitro and in vivo models. In some studies, these effects were accompanied by additive effects on several cell biologic parameters. In pretreated postmenopausal patients with metastatic breast cancer, objective responses have been observed in 10-12%, and stable disease in 42-46% of the patients; in previously untreated patients objective response rates of 11 and 56% have been reported. The clinical development of onapristone was stopped because of liver toxicity. At the present time, apart from development of new pure potent PAs, clinical investigation of combined therapy of PAs with antiestrogens are urgently needed. PMID- 11108895 TI - Alterations in sex steroids and gonadotropins in post-menopausal women subsequent to long-term mifepristone administration. AB - Long-term administration of progesterone antagonists (PAs) and progesterone receptor modulators (PRMs) has been proposed as a novel hormonal therapy for various hormone dependent maladies. We studied the long-term endocrine effects of mifepristone on the kinetics of estradiol (E(2)) and its precursors, and on gonadotropin levels in five postmenopausal women treated for unresectable meningioma with mifepristone [200 mg/day] for at least 15 months. Serum samples were analyzed for LH, FSH and SHBG with fluoroimmunoassay; androstenedione (A), testosterone (T), estrone (E(1)) and E(2) were measured with radioimmunoassay (RIA). Serum levels of mifepristone were measured using both RIA and high performance-liquid chromatography (HPLC). Serum levels (mean +/- SD) of LH and FSH were suppressed from pretreatment values of 32 +/- 16 and 65 +/- 30 IU/l to 13 +/- 7 and 33 +/- 16 IU/l at 6 months (P < 0.05), respectively. Serum (mean +/- SD) A, T, E(1), and E(2) were increased from initial values of 6.9 +/- 0.9 nmol/l, 1.2 +/- 0.3 nmol/l, 77 +/- 25 pmol/l, and 29 +/- 14 pmol/l to 6 month values of 13.1 +/- 5.6 nmol/l, 1.8 +/- 0.6 nmol/l, 178 +/- 60 pmol/l, and 45 +/- 22 pmol/l (n.s.). The correlation coefficients between the levels of A, T, E(1), and E(2) were statistically significant, whereas the ratios of T/A, E(1)/A, E(2)/E(1), and E(2)/T remained unchanged. The levels of SHBG remained stable, and ranged from 48 +/- 10 to 65 +/- 9 nmol/l (mean +/- SD). Thus, prolonged mifepristone treatment marginally increased the serum levels of A, T, E(1) and E(2). These effects of mifepristone are likely due to its antiglucocorticoid effect and thus increased secretion of adrenal A. Serum levels of LH and FSH declined. The serum levels of gonadotropins and those of T, E(1) and E(2) were inversely, yet significantly, correlated. Therefore the decrease in LH and FSH might reflect the slightly increased levels of T, E(1) and E(2). However, the lack of change in SHBG and the low E(2) levels suggest that enhanced systemic estrogen effects are unlikely during long-term mifepristone treatment. PMID- 11108896 TI - The Jerusalem declaration on progesterone receptor modulators. PMID- 11108897 TI - Very low platelet counts in post-transfusion purpura falsely diagnosed as heparin induced thrombocytopenia. Report of four cases and review of literature. AB - Differential diagnosis between post-transfusion purpura (PTP) and heparin-induced thrombocytopenia (HIT) can be difficult in the initial stages of thrombocytopenia, as the early clinical presentations are often similar. Four patients are described who were suspected clinically of suffering from HIT. All four patients had recent blood transfusions and platelet alloantibodies, thus the diagnosis of PTP was made. One lethal gastrointestinal and one retroperitoneal hemorrhage developed in two of the four patients. Unusually, one patient was male and two different platelet alloantibodies were present in his serum; in another patient platelet alloantibodies and HIT-antibodies were detectable. To arrive at the right diagnosis as quickly as possible is vitally important since treatment, which has to be initiated promptly, is very different for the two syndromes. Thus, we suggest that in patients where HIT is suspected, additional information should be sought. If features consistent with PTP (such as a recent blood transfusion or a marked drop in platelet count to below 15 Gpt/L) are present, we recommend parallel testing for platelet alloantibodies to rule out PTP. PMID- 11108898 TI - Prevalence of three prothrombotic polymorphisms. Factor V G1691A, factor II G20210A and methylenetetrahydrofolate reductase (MTHFR) C 677T in Argentina. On behalf of the Grupo Cooperativo Argentino de Hemostasia y Trombosis. PMID- 11108899 TI - No effect of dietary trans isomers of alpha-linolenic acid on platelet aggregation and haemostatic factors in european healthy men. The TRANSLinE study. AB - The aim of this study was to investigate the effect of trans alpha-linolenic acid on platelet aggregation and blood haemostasis. A randomized, double blind dietary intervention trial was carried out with healthy male volunteers (n=88) in three European centers. After a 6-week washout period where subjects avoided foods containing all trans fats, subjects either continued for 6 weeks with a low trans diet or a diet where trans alpha-linolenic acid provided 0.6% of energy (supplied as oil, margarine, cheese, muffins, and biscuits). At the end of the washout period the intake of trans polyunsaturated fats was 58+/-115 mg/day; this increased in patients on the high trans diet by +1344+/-328 mg/day, compared with +10+/-67 mg/day in patients on the low trans diet (p<0.01). The change in trans alpha-linolenic acid in plasma cholesteryl esters was 0.26+/-0. 20 on the high trans and 0.00+/-0.07% of fatty acids on the low trans diet (p<0.001). No effect of the high trans diet was observed on platelet aggregation: collagen EC(50) high trans 157+/-100, low trans 152+/-90 ng/mL (NS); U44619 EC(50) high trans 81+/-61, low trans 59+/-27 nM (NS). The high trans diet did not affect platelet thromboxane production, fibrinogen levels, factor VII, activated factor VIIa, or plasminogen activator inhibitor activity. There were no center-specific differences in response to the high trans diet. A relatively high amount of trans alpha-linolenic acid for 6 weeks does not increase the risk of coronary heart disease by promoting platelet aggregation and blood coagulation. PMID- 11108900 TI - The role of prothrombotic mutations in patients with Buerger's disease. AB - Thromboangiitis obliterans (TAO), or Buerger's disease, is a segmental occlusive inflammatory disorder of the arteries and veins, and etiopathogenesis is still obscure. In the present study we investigated the prevalence of prothrombin 20210 G-->A, factor V 1691 G-->A (Factor V Leiden), and factor V 4070 A-->G (His 1299 Arg) mutations, found to be associated with increased risk for vascular thrombosis, in 36 patients with TAO. We performed a case-control study of these mutations. The odds ratio for prothrombin 20210 A allele compared with G allele was 7.98 (95% confidence intervals 2. 45-25.93). Only this prothrombotic genetic factor was associated with the risk of TAO (p=0.032). In conclusion, carrying the prothrombin 20210 G-->A may be an important prothrombotic risk factor of TAO. This genetic predisposition must be screened in these patients routinely, and clinical importance must be supported by further investigations. PMID- 11108901 TI - Successful intravenous interferon-beta treatment of chronic hepatitis C in a patient with Bernard-Soulier syndrome. PMID- 11108903 TI - Influences of fixatives on flow cytometric measurements of platelet P-selectin expression and fibrinogen binding. AB - Sample fixation is an important issue in flow cytometric platelet assays. However, previous reports were less than consistent regarding the influence of sample fixation on the assays. We evaluated the effects of formaldehyde and paraformaldehyde fixation on platelet P-selectin expression and fibrinogen binding using whole-blood flow cytometry and a Coulter EPICS XL-MCL cytometer. Fluorescent-labeled whole-blood samples were diluted with HEPES-buffered saline or fixed with formaldehyde (0.2, 0.5, and 1. 0%) or paraformaldehyde (0.5, 1.0, and 2.0%). Platelet P-selectin expression was 1.1+/-0.3% and 39.6+/-13.7% in unfixed resting and 10(-5) M ADP stimulated samples, respectively. Resting P selectin expression was not significantly altered by 0.2 or 0.5% formaldehyde fixation, but was slightly decreased by 1.0% formaldehyde fixation or PFA fixation. Formaldehyde fixation caused small increases of P-selectin expression in ADP-stimulated samples. Compared to platelet fibrinogen binding of unfixed resting (4.5+/-2.1%) and ADP-stimulated (56.7+/-22.6%) samples, formaldehyde or paraformaldehyde fixation had no significant influence on resting samples, but mildly increased fibrinogen binding in stimulated samples. Unfixed samples were stable for 2 h. Fixed samples were generally stable for at least 6 h, but not thereafter. Thus, formaldehyde and paraformaldehyde have mild but complex influences on platelet P-selectin expression and fibrinogen binding measurements. To evaluate the stabilities of unfixed and fixed samples, samples were analyzed after different durations (0, 1, 2, 4, 6, 12, and 24 h) of storage at 4 degrees C in the dark. The results suggest that sample manipulation without fixation may be used when the samples are analyzed within 2 h, and that fixation with 0.5-1.0% formaldehyde or paraformaldehyde seems to be preferable when sample analysis is delayed. Effects of fixation should be carefully evaluated when establishing flow cytometric platelet assays in every laboratory. PMID- 11108902 TI - Cardiovascular risk factors in vegetarians. Normalization of hyperhomocysteinemia with vitamin B(12) and reduction of platelet aggregation with n-3 fatty acids. AB - Hyperhomocysteinemia in association with vitamin B(12) deficiency, and increased platelet aggregation, probably due to dietary lack of n-3 fatty acids, constitute cardiovascular risk factors frequently observed in vegetarians. We tested if administration of vitamin B(12) normalizes the concentration of total plasma homocysteine, and if intake of eicosapentaenoic (20:5n-3) and docosahexaenoic (22:6n-3) fatty acids modulates platelet function in a population of lactoovovegetarians. One week after a single intramuscular injection of cyanocobalamin (10000 microg) in 18 individuals, serum vitamin B(12) increased from 149+/-63 pg/mL to 532+/-204 pg/mL (p<0.0001) and total tHcy dropped from 12.4+/-4.7 to 7.9+/-3.1 micromol/L (p<0. 0001). Ten of fourteen of these vegetarians completed an 8-week supplementation with 700 mg/day of each eicosapentaenoic and docosahexaenoic acids. Increased incorporation of these fatty acids into plasma lipids was observed in all of them, together with a significant reduction in maximum percentage or slope of platelet aggregation with all the agonists tested (ADP, epinephrin, collagen, arachidonic acid). No significant change in bleeding time was observed after n-3 fatty acid trial. Supplementation with vitamin B(12) and n-3 fatty acids corrects hyperhomocysteinemia and reduces platelet reactivity to agonists in vegetarians. Whether this supplementation improves the already reduced cardiovascular morbidity and mortality associated with vegetarian diet has yet to be demonstrated. PMID- 11108904 TI - Thrombogenic potential of contrast media in an experimental model of laser induced thrombosis. AB - Controversy still exists about the pro- or antithrombotic side effects of contrast media used in daily medical practice. Recent reports have shown that various contrast media, including ionic compounds, have deleterious prothrombotic actions. A new evaluation of these adverse side effects is reported here, with the study of the dose-effect relationship. Two ionic (ioxaglate and diatrizoate) and two non-ionic contrast media (iopamidol and iohexol) were studied. Experiments were done on 22 groups of 5 Wistar male rats each, using a Laser Argon-induced thrombosis model in mesenteric microvessels. Three parameters were studied: the number of laser beams needed to induce platelet thrombus formation, the number of emboli, and the duration of embolization. Platelet count and platelet aggregation also were determined. Iopamidol and iohexol induced a significant rise in both the number of emboli and the duration of embolization in mesenteric microvessels at doses up to 1 mL/kg. Ioxaglate and diatrizoate also significantly increased these parameters at doses up to 2 mL/kg. All the products tested decreased platelet count, inducing a -17 to -30% variation from control values. Diatrizoate and ioxaglate inhibited platelet aggregation, while iopamidol and Iohexol behaved as activators. All non-ionic, and to a lesser extent, all ionic contrast media demonstrated prothrombotic properties. PMID- 11108905 TI - Cloning and characterization of the murine antithrombin gene. PMID- 11108906 TI - Anti-thrombotic efficacies of enoxaparin, dalteparin, and unfractionated heparin in venous thrombo-embolism. AB - BACKGROUND: Few data exist by which the anti-thrombotic efficacy of different anticoagulants may be compared. We used a radiolabeled antibody specific for polymerizing fibrin to compare the in vivo anti-thrombotic potencies of different systemic anticoagulants (enoxaparin, dalteparin, and unfractionated heparin). METHODS AND RESULTS: Deep venous thrombi (DVTs) were induced in dogs' femoral veins. The dogs were then treated with one of the following subcutaneous regimens: enoxaparin 100 units/kg (1.0 mg/kg) every 12 hours (n=4), dalteparin 200 units/kg every 24 hours (n=4), or unfractionated heparin 240 units/kg every 8 hours with dose adjustment via aPTT (n=3). 111Indium-labeled anti-fibrin antibodies, specific for propagating thrombi, were given intravenously and nuclear scans of the legs were taken over the following 24 hours. Thrombus propagation was estimated by the ratio of gamma emissions from the legs containing DVTs divided by the emissions from the contralateral "control" legs. DVTs accumulated labeled anti-fibrin antibodies at the same rates in both the enoxaparin group and the dalteparin group (gamma emissions 171+/-6% and 168+/-36% of control by 24 hours, respectively). DVTs in the adjusted dose unfractionated heparin group tended to accumulate antibodies at a slower rate (129+/-19% of control by 24 hours). CONCLUSIONS: Enoxaparin and dalteparin inhibited propagation of pre-formed thrombi to the same degree. Subcutaneous unfractionated heparin, adjusted every 8 hours by aPTT, tended to suppress ongoing thrombosis more than either LMWH. PMID- 11108907 TI - In vitro and in vivo properties of bicyclic lactam inhibitors: a novel class of low molecular weight peptidomimetic thrombin inhibitors. AB - We have developed potent and selective thrombin inhibitors with a novel non peptidic structure. A bicyclic lactam was used as the scaffold on which various P1 and P3 motifs were substituted. Herein, we report the in vitro and in vivo properties of four representatives of this novel class of inhibitors. Their Ki values were less than 10 nM, they inhibited equally both free and clot-bound thrombin, and they displayed high level of specificity for thrombin over other serine proteases (trypsin, factor Xa, activated Protein C, and plasmin). They prolonged the clotting time of human plasma to twice the control value in coagulation assays (TT, APTT, and PT) at a concentration below 3 microM. Their anticoagulant activities using rat plasma were similar to, although slightly weaker, than with human plasma. Furthermore, they inhibited thrombin-induced platelet aggregation (human and rat) at concentrations close to their Ki values for thrombin. These molecules demonstrated similar dose response antithrombotic efficacy in rat arterial and venous thrombosis models when given as i.v. bolus followed by infusion. Antithrombotic efficacy of 85% and greater was observed at a dose of 5-7 microM/kg/hour in each model. Bicyclic lactam inhibitor 3, at a dose which caused a complete inhibition of visible thrombus formation in the venous and arterial models of thrombosis, showed a 1.9-2.1 and a 4.0-4.8-fold shift in APTT and TT, respectively. Unfortunately, the bicyclic lactam inhibitors exhibited low oral bioavailability in rats. Therefore, this novel class of bicyclic lactam thrombin inhibitor has the potential to be promising intravenous antithrombotic agents for the treatment of arterial as well as venous thrombosis and warrants further investigation. PMID- 11108908 TI - Down-regulation of murine tissue factor pathway inhibitor mRNA by endotoxin and tumor necrosis factor-alpha in vitro and in vivo. AB - Tissue factor pathway inhibitor (TFPI) is the protease inhibitor that regulates the extrinsic coagulation pathway initiated by the factor VIIa/TF complex. In this study, we first investigated tissue distribution of TFPI mRNA in the mouse and found that TFPI mRNA expression level was by far the highest in the lung, followed by the heart, adrenal, and adipose tissue. Since little has been known concerning the regulation of TFPI gene expression in vivo, we further analyzed the changes in the TFPI mRNA level in murine tissues after intraperitoneal injection of lipopolysaccharide (LPS), tumor necrosis factor-alpha (TNF-alpha), and interleukin-1 (IL-1). LPS and TNF-alpha dramatically decreased TFPI mRNA expression in four tissues examined (e.g., lung, heart, kidney, and adipose tissue), whereas the suppressive effect of IL-1 on TFPI mRNA was limited. The down-regulation of TFPI mRNA expression by LPS and TNF-alpha was also observed in cultured mouse endothelial cells and in cardiomyocyte cell lines. The decreased TFPI mRNA expression by LPS and TNF-alpha in tissues and in the specific cell types may contribute to an increase in the local procoagulant potential, resulting in the thrombotic tendency under septic and/or inflammatory conditions. PMID- 11108910 TI - F. Mseeh, R.F. Colman and R.W. colman-inactivation of platelet PDE2 by an affinity label. 8- PMID- 11108909 TI - Modulation of polymorphonuclear leukocytes function by nitric oxide. AB - Recognition of the endothelium-derived relaxation factor as nitric oxide (NO) gave rise to an impression that NO was synthesised only by the endothelial lining of the vessel wall. Later it was found that NO is synthesized constitutively by the enzyme nitric oxide synthase (NOS) in various cells. However, inflammatory cytokines can induce NOS (known as inducible NOS [iNOS]) activity in all the somatic cells. Blood cells, such as eosinophils, platelets, neutrophils, monocytes, and macrophages, also synthesize NO. Among them, polymorphonuclear leukocytes (PMNs) constitute an important proportion and are also the major participants in a number of pathological conditions with suggestive involvement of NO. PMNs can synthesize NO at rates similar to endothelial cells, thus suggesting the importance of PMN-derived NO in various physiological and pathological conditions. Most of the studies so far focus on the peripheral PMNs, while studies on PMNs after emigration are limited, thus warranting systematic studies on PMNs from both sources. The role of the endothelial NOS (eNOS) and functions of NO derived from the endothelial cells has been studied extensively. However, understanding of the PMNs NOS and its regulatory role in their function is unraveling. The present review summarizes the modulatory role of NO on PMNs functions and points out the discrepancies relating to presence of NOS in PMNs. This information will be helpful in understanding the importance of NO in physiological and pathological conditions associated with PMNs. PMID- 11108912 TI - Executive summary PMID- 11108914 TI - 2. Multifactorial diseases PMID- 11108911 TI - Risk estimation for multifactorial diseases. A report of the International Commission on Radiological Protection. AB - This report reviews data on naturally-occurring multifactorial diseases and develops a mathematical model to predict the impact of radiation-induced mutations on the frequencies of these diseases in the population. It provides an outline of the aetiological features and examples of multifactorial diseases, interpreted to arise as a result of the joint action of genetic and environmental factors. Examples include common congenital abnormalities (such as neural tube defects, cardiovascular malformations, cleft lip+/-palate etc.) and chronic diseases (such as coronary heart disease, essential hypertension, diabetes mellitus etc.). These diseases are not readily explained on the basis of simple mendelian patterns of inheritance. The report considers the concepts and models used to explain the inheritance patterns of multifactorial diseases with particular emphasis on the multifactorial threshold model (MTM) of disease liability. The MTM is useful for predicting risk to relatives of those affected from information on their population frequencies. In these predictions, the heritability (h(2)) provides a measure of the relative importance of transmissible genetic effects in the overall phenotypic variation. Conceptual differences between mendelian and multifactorial diseases are discussed. The genetic basis of a multifactorial disease is that a genetically susceptible individual may or may not develop the disease depending on the interaction of a number of risk factors, both genetic and environmental. Three chronic multifactorial disease entities are reviewed in depth, viz. diabetes mellitus, essential hypertension, and coronary heart disease. The report considers briefly mechanistic population genetic models developed to explain polygenic variation. The basic conclusion is that the concepts of liability and threshold (underlying the MTM model) and that of mutation-selection balance (from population genetic models) together provide a basis for developing a model for assessing the impact of radiation-induced mutations on the frequencies of multifactorial diseases in the population.The mutation component (MC) of genetic diseases quantifies the responsiveness of the genetic component of a disease to an increase in mutation rate (e.g. after radiation exposure). This report integrates the concepts of liability and threshold (from the MTM model) and of mutation-selection equilibrium (from mechanistic population genetic models) into the 'Finite Locus Threshold Model' (FLTM) for estimating MC for multifactorial diseases and the relationship between MC and h(2) of these diseases. Computer simulation studies illustrate the effects of one-time or a permanent increase in mutation rate on MC for multifactorial diseases.Finally, the report addresses the estimation of the radiation risk of multifactorial diseases. A formal revision of the estimates of risk of multifactorial diseases (and also of mendelian diseases) contained in the 1990 Recommendations of ICRP, Publication 60, must await the results of studies currently underway. While future genetic risk estimates are likely to be lower than those in current use, until the new ones become available, those provided in Publication 60 may be regarded as being adequate for use in radiological protection- they are unlikely to underestimate risk. PMID- 11108915 TI - 3. Models Of inheritance of multifactorial diseases PMID- 11108916 TI - 4. General attributes and epidemiological aspects of chronic multifactorial diseases PMID- 11108917 TI - 5. Diabetes mellitus PMID- 11108918 TI - 6. Essential hypertension PMID- 11108919 TI - 7. Coronary heart disease PMID- 11108920 TI - 8. Mechanistic models on the maintenance of multifactorial traits in the population PMID- 11108922 TI - 10. The concept of mutation component of genetic diseases and its application for estimating the responsiveness of mendelian diseases to an increase in mutation rate PMID- 11108921 TI - 9. General conclusions from studies of multifactorial diseases and their relevance for risk estimation PMID- 11108923 TI - 11. A finite-locus threshold model for estimating the mutation component of multifactorial diseases PMID- 11108924 TI - 12. Relevance for radiological protection PMID- 11108927 TI - Preface PMID- 11108925 TI - Pregnancy and medical radiation. AB - Thousands of pregnant patients and radiation workers are exposed to ionising radiation each year. Lack of knowledge is responsible for great anxiety and probably unnecessary termination of pregnancies. For many patients, the exposure is appropriate, while for others the exposure may be inappropriate, placing the unborn child at increased risk. Prenatal doses from most properly done diagnostic procedures present no measurably increased risk of prenatal death, malformation, or impairment of mental development over the background incidence of these entities. Higher doses, such as those involved in therapeutic procedures, can result in significant fetal harm. The pregnant patient or worker has a right to know the magnitude and type of potential radiation effects that might result from in utero exposure. Almost always, if a diagnostic radiology examination is medically indicated, the risk to the mother of not doing the procedure is greater than is the risk of potential harm to the fetus. Most nuclear medicine procedures do not cause large fetal doses. However, some radiopharmaceuticals that are used in nuclear medicine can pose significant fetal risks. It is important to ascertain whether a female patient is pregnant prior to radiotherapy. In pregnant patients, cancers that are remote from the pelvis usually can be heated with radiotherapy. This however requires careful planning. Cancers in the pelvis cannot be adequately treated during pregnancy without severe or lethal consequences for the fetus. The basis for the control of the occupational exposure of women who are not pregnant is the same as that for men. However, if a woman is, or may be, pregnant, additional controls have to be considered to protect the unborn child. In many countries, radiation exposure of pregnant females in biomedical research is not specifically prohibited. However, their involvement in such research is very rare and should be discouraged. Termination of pregnancy is an individual decision affected by many factors. Fetal doses below 100 mGy should not be considered a reason for terminating a pregnancy. At fetal doses above this level, informed decisions should be made based upon individual circumstances. PMID- 11108928 TI - B cells and antibodies in CNS demyelinating disease. AB - There is much evidence to implicate B cells, plasma cells, and their products in the pathogenesis of MS. Despite unequivocal evidence that the animal model for MS, EAE, is initiated by myelin-specific T cells, there is accumulating evidence of a role for B cells, plasma cells, and their products in EAE pathogenesis. The role(s) played by B cells, plasma cells, and antibodies in CNS inflammatory demyelinating diseases are likely to be multifactorial and complex, involving distinct and perhaps opposing roles for B cells versus antibody. PMID- 11108929 TI - Rapid entry and downregulation of T cells in the central nervous system during the reinduction of experimental autoimmune encephalomyelitis. AB - We investigated the mechanisms whereby a previous attack of experimental autoimmune encephalomyelitis (EAE) modifies a subsequent attack in the Lewis rat. Active immunization with myelin basic protein (MBP) and complete Freund's adjuvant 28 days after the passive transfer of MBP-sensitized spleen cells induced a second episode of EAE, which occurred earlier than in naive control animals, but was less severe overall. The pattern of neurological signs was also different in rechallenged rats, which had less severe tail and hindlimb weakness but more severe forelimb weakness. In rechallenged rats, inflammation was more severe in the cervical spinal cord, cerebellum, brainstem and cerebrum, but less severe in the lumbar spinal cord, than in controls. The early onset of EAE in rechallenged rats was explained by a memory T cell response to MBP(72-89) in the draining lymph node and spleen, and by the enhanced entry of T cells into the central nervous system (CNS). However, the number of alphabeta T cells in the spinal cord of rechallenged rats declined faster than in controls, especially in the lumbosacral cord, where the number of Vbeta8.2(+) T cells and the frequency of T cells reactive to MBP(72-89) rapidly decreased, indicating rapid downregulation of the immune response in the previously inflamed spinal cord. Apoptosis of inflammatory cells in the CNS was increased in the rechallenged rats and is likely to contribute to this downregulation. Furthermore, during the disease course the generation of encephalitogenic T cells in the peripheral lymphoid organs was limited compared with controls. Thus, a previous attack of EAE modifies a subsequent attack through the interaction of the following processes: a memory T cell response to MBP; facilitated T cell entry into the CNS; downregulation of the immune response in the CNS, including increased apoptosis of inflammatory cells; and a limited generation of encephalitogenic T cells in the peripheral lymphoid organs. PMID- 11108930 TI - Resveratrol inhibits interleukin-6 production in cortical mixed glial cells under hypoxia/hypoglycemia followed by reoxygenation. AB - Reactive oxygen intermediates (ROIs) are important mediators of a variety of pathological processes, including inflammation and ischemia/reperfusion injury. Cytokines and chemokines are detected at mRNA level in human and animal ischemic brains. This suggests that hypoxia/reoxygenation may induce cytokine production through generation of ROIs. In this study, we investigated the cytokine induction and inhibition by antioxidants in rat cortical mixed glial cells exposed to in vitro ischemia-like insults (hypoxia plus glucose deprivation). The results showed that interleukin-6 (IL-6) mRNA and protein, but not tumor necrosis factor alpha (TNF-alpha) or interleukin-1beta (IL-1beta), were induced during hypoxia/hypoglycemia followed by reoxygenation in the mixed glial cells. The accumulation of IL-6 mRNA was induced as early as 15 min after hypoxia/hypoglycemia and its level was further increased after subsequent reoxygenation. Among the antioxidants studied, only resveratrol suppressed IL-6 gene expression and protein secretion in mixed glial cultures under hypoxia/hypoglycemia followed by reoxygenation. These findings suggest that resveratrol might be useful in treating ischemic-induced inflammatory processes in stroke. PMID- 11108931 TI - Analysis of neural stem cells by flow cytometry: cellular differentiation modifies patterns of MHC expression. AB - Neural stem cells are currently considered very hopeful candidates for cell replacement therapy in neurodegenerative pathologies such as Parkinson's disease. Here we show that different cell types derived from neurospheres amplified in vitro can be identified by FACS analysis relying solely on physical parameters (FSC/SSC) or autofluorescence. Additionally, after treatment with a panel of inflammatory cytokines, neurospheres and their differentiated progeny were shown to express MHC antigens which could potentially cause transplant rejection. Astrocytes expressed the highest levels of MHC. Hence removing such cells prior to transplantation could potentially optimise transplant survival. PMID- 11108932 TI - Neonatal dexamethasone treatment induces long-lasting changes in T-cell receptor vbeta repertoire in rats. AB - Glucocorticoids are frequently administered for the prevention of chronic lung disease in infants with respiratory distress syndrome. However, neonatal treatment may have consequences for immune functioning in the long-term. Here we demonstrate that neonatal glucocorticoid treatment has long-lasting effects on mRNA expression of several Vbeta genes within the CD4 and CD8 T cell subset in rats. Changes in the peripheral T cell Vbeta repertoire may be a consequence of altered intrathymic selection events in which corticosterone plays an important role. Indeed, here we show that neonatal glucocorticoid treatment affects corticosterone production by thymic epithelial cells during neonatal life. In conclusion, changes in T cell Vbeta repertoire after neonatal glucocorticoid treatment may contribute to altered immune reactivity in later life. PMID- 11108933 TI - Augmented death in immunostimulated astrocytes deprived of glucose: inhibition by an iron porphyrin FeTMPyP. AB - The present study shows that under glucose-deprived conditions immunostimulated astrocytes rapidly undergo death due to their increased susceptibility to endogenously produced peroxynitrite. Fe(III)tetrakis(N-methyl-4' pyridyl)porphyrin (FeTMPyP), but not the structurally related compounds ZnTMPyP and H(2)TMPyP, prevented the death in glucose-deprived immunostimulated astrocytes. Consistently, FeTMPyP, not ZnTMPyP and H(2)TMPyP, completely blocked the elevation of nitrotyrosine immunoreactivity (a marker of peroxynitrite) and the depolarization of the mitochondrial transmembrane potential in glucose deprived immunostimulated astrocytes. The present data suggest that peroxynitrite may be associated with glial cell death during metabolic deterioration in the cerebral ischemic penumbra. PMID- 11108934 TI - Polyclonal immunoglobulins (IVIg) modulate nitric oxide production and microglial functions in vitro via Fc receptors. AB - Controlled trials in multiple sclerosis (MS) and case reports in acute demyelinating encephalomyelitis (ADEM) have shown that intravenous immunoglobulins (IVIg) are of therapeutic benefit in central nervous system (CNS) inflammatory diseases. Studies in experimental autoimmune encephalomyelitis (EAE) have suggested these effects are mediated by modulation of the cytokine network and T cell responses. However, there are no data on the influence of IVIg on the local immune reaction in the CNS, the site of inflammation in EAE. We have therefore studied the effect of IVIg on cultured rat microglia, the main immune cell in the CNS. IVIg increased nitric oxide (NO) production in a dose-dependent manner in microglia stimulated with IFNgamma. The increase was only marginal in LPS-treated cells, and no effect was seen in untreated microglia or after stimulation with TNFalpha or PMA. This enhancement of NO production depended on the Fc portion of IVIg and could be abrogated by the pharmacological inhibition of Syk and phosphatidylinositol 3-kinase, two enzymes involved in the signalling cascade of Fc receptors. TNFalpha secretion was dose-dependently stimulated by IVIg in both untreated microglia and after stimulation with LPS or IFNgamma. Again, this effect was mediated through the Fc portion. Finally, we showed that Fc receptor-mediated phagocytosis was inhibited by IVIg, presumably by blockade of the Fc receptor. These different effects may protect oligodendrocytes from antibody mediated phagocytosis and on the other hand could terminate the immune reaction by induction of apoptosis in infiltrating T cells via NO and TNFalpha. We propose that IVIg, in addition to known effects on the peripheral immune system, may also modulate the local immune reaction in CNS inflammatory disease. PMID- 11108935 TI - Macrophages express neurotrophins and neurotrophin receptors. Regulation of nitric oxide production by NT-3. AB - In this study we examined the expression of neurotrophins and their receptors in mouse macrophages and the effects of the neurotrophins on nitric oxide secretion. Macrophages expressed TrkB and TrkC but not BDNF, NT-3 or NT-4. LPS induced up regulation of TrkB and TrkC and of BDNF and NT-3 expression. Treatment of macrophages with NT-3 increased the secretion of nitric oxide in LPS-treated macrophages and this increase was blocked by K252a, a Trk kinase inhibitor. In contrast, BDNF and NT-4 had no significant effects on the induction of nitric oxide. Our results suggest that NT-3 play important roles in the function of macrophages during inflammatory responses and in tissue repair. PMID- 11108936 TI - VIP and PACAP inhibit Fas ligand-mediated bystander lysis by CD4(+) T cells. AB - FasL/Fas-mediated lysis represents the major cytotoxic mechanism for CD4(+) effectors, with important consequences for immune cell homeostasis. Upon stimulation by specific antigen-presenting cells (APCs), CD4(+) effectors can lyse the cognate APCs (direct targets) and neighboring innocent bystanders. Previously we showed that the neuropeptides VIP and PACAP prevent FasL expression and activation-induced cell death in T cells. In this study we investigated the effects of VIP and PACAP on FasL expression and subsequent direct and bystander lysis by CD4(+) effectors generated in vivo. VIP/PACAP inhibit FasL expression in allogeneic effectors, and reduce Fas-mediated cytotoxicity against specific allotargets and syngeneic bystanders. VIP/PACAP also inhibit FasL expression in antigen-specific CD4(+) effectors, and reduce their cytotoxic activity against both the stimulatory APC, and syngeneic or allogeneic bystanders. Since bystander lysis is involved in the pathogenesis of several autoimmune and inflammatory diseases, the identification of regulatory factors that limit this process is highly significant. PMID- 11108937 TI - delta-opioid receptors inhibit neurogenic intestinal secretion evoked by mast cell degranulation and type I hypersensitivity. AB - Histamine and the mast cell degranulator, compound 48/80 produced elevations in short-circuit current, an electrical measure of active anion secretion, across porcine ileal mucosa sheets mounted in Ussing chambers. Luminally-applied beta lactoglobulin produced similar effects in mucosal sheets from cow's milk sensitized pigs. Their secretory effects were attenuated by blockers of H(1) histamine receptors, neuronal conduction or epithelial Na(+)/K(+)/Cl(-) cotransport. The delta-opioid agonist [D-Pen(2), D-Pen(5)]enkephalin suppressed mucosal responses to these substances in a naltrindole-reversible manner. Furthermore, submucosal mast cells and delta-opioid receptor-immunoreactive nerve fibers were observed in close juxtaposition. Intestinal neural pathways linking immediate hypersensitivity to secretory host defense appear to express inhibitory delta-opioid receptors. PMID- 11108938 TI - Mice lacking myeloperoxidase are more susceptible to experimental autoimmune encephalomyelitis. AB - EAE is a demyelinating disease which serves as an animal model for multiple sclerosis (MS). Myeloperoxidase (MPO) has been implicated in MS through its presence in invading macrophages, and by association of a -463G/A promoter polymorphism with increased risk. Also, MPO at 17q23.1 is within a region identified in genome scans as a MS susceptibility locus. We here examine the incidence of EAE in MPO knockout (KO) mice. MPO is detected in invading macrophages in the CNS of wild-type mice, yet unexpectedly, MPO-KO mice have significantly increased incidence of EAE: Ninety percent of MPO-KO mice developed complete hind limb paralysis as compared to 33% of wildtype (WT) littermates (P<0.0001). This is the first evidence that MPO plays a significant role in EAE, consistent with its postulated role in MS. PMID- 11108939 TI - Rat RT1.B-transfected fibroblast lines process and present myelin antigens and activate T cells to induce experimental autoimmune encephalomyelitis. AB - The genes encoding the Lewis rat RT1.B molecule (MHC Class II I-A equivalent) were transfected and expressed in mouse DAP.3 fibroblast cells together with the gene encoding the mouse ICAM-1 molecule. Both molecules were stably expressed on the cell surface of DAP.3 cells under longterm culture conditions. The RT1.B/mICAM-1 transfectants presented antigen in a specific manner to a RT1. B restricted rat T cell hybridoma specific for the 69-89 peptide of myelin basic protein (BP). In addition, the transfectants were able to present antigen to a BP69-89-specific rat T cell line. Presentation to a RT1.D (MHC Class II I-E equivalent)-restricted BP87-99-specific T cell line was minimal. Production of the Th1 cytokine IFN-gamma by BP69-89-specific T cells when stimulated by RT1.B/mICAM-1 transfectants correlated very well with proliferation to specific antigen. Moreover, RT1.B-transfected DAP.3 cells sufficiently stimulated BP69-89 specific T cells such that they were able to transfer experimental autoimmune encephalomyelitis (EAE) to Lewis rat recipients. Thus, the RT1.B molecule is functionally expressed on the surface of transfected Dap.3 fibroblasts and is capable of MHC Class II-restricted, antigen-specific presentation to rat T cells. PMID- 11108940 TI - Effect of a cyclic heptapeptide based on the human CD4 domain 1 CC' loop region on murine experimental allergic encephalomyelitis: inhibition of both primary and secondary responses. AB - The 802-2 peptide, designed from the conserved D1-CC' loop region of human CD4, can disrupt CD4(+) T cell activation in both human and murine systems. Here, 802 2 was investigated for efficacy in acute murine experimental allergic encephalomyelitis (EAE) models, and was found to significantly reduce the severity of disease when administered either before or after the onset of symptoms. 802-2 treatment during PLP139-151 induction of EAE rendered the mice more resistant to subsequent rechallenge with antigen, and was also efficacious when initially administered during a secondary EAE response. T cells from 802-2 treated mice proliferated poorly to in vitro restimulation with PLP139-151 and exhibited decreased frequencies of IL-2, IL-4, and IFN-gamma producing cells, but were still able to respond to third-party antigens. These combined results suggest the potential therapeutic value of 802-2 for inhibition of CD4(+) T cell neuroimmunological responses. PMID- 11108941 TI - IL-10 and IFN-gamma in Guillain-Barre syndrome. Network Members of the Swedish Epidemiological Study Group. AB - Guillain-Barre syndrome (GBS) is an acute inflammatory disease affecting myelin and axons of the peripheral nervous system (PNS). GBS is considered to be caused by breakdown of tolerance to autoantigens of the PNS. The involvement of cytokines in GBS and in relation to treatment with high dose intravenous immunoglobulin (IvIg) is incompletely known. We studied the temporal profiles of IL-10 and IFN-gamma-secreting blood mononuclear cells (MNC) over the course of GBS, using enzyme-linked immunospot (ELISPOT) assays. Pretreatment levels of blood MNC spontaneously secreting IL-10 were higher in the acute phase of GBS than in control patients with aseptic meningitis, other neurological diseases, diabetic neuropathy and healthy subjects. Levels of IFN-gamma-secreting blood MNC were not increased over the course of GBS. Patients treated with IvIg had lower numbers of IL-10-secreting MNC compared to untreated patients. High levels of IL 10-secreting MNC correlated with serum anti-ganglioside IgM antibody levels, and with neurophysiological signs of axonal damage. The present data suggests that IFN-gamma is not involved in GBS pathogenesis, and IL-10 being up-regulated in the early phase of GBS and associated with axonal damage, may have a pathogenetic role in GBS. PMID- 11108942 TI - Relationship between serum levels of IL-10, MRI activity and interferon beta-1a therapy in patients with relapsing remitting MS. AB - BACKGROUND: The purposes of this study were to: (1) compare monthly serum IL-10 in patients with relapsing remitting (RR) multiple sclerosis (MS) and healthy controls, (2) determine the relationship between IL-10 and MRI activity and (3) determine the effect of interferon beta-1a (IFNB-1a) treatment on IL-10 levels. RESULTS: Median serum IL-10 levels were lower in untreated RRMS (185.5 pg/ml) compared to controls (438.5 pg/ml) (P=0.19). Serum levels of IL-10 did not appear to predict the appearance of new gadolinium-enhancing (Gd+) lesions concurrently or 1 month thereafter. However, IL-10 levels were more likely to be elevated the month during which Gd+ lesions resolved (OR=3.14, P=0.01). Median IL-10 levels were lower during IFNB-1a treatment (P=0.01). CONCLUSIONS: These observations suggest a relationship between serum IL-10 levels and resolution of abnormal vascular permeability in new lesions. PMID- 11108943 TI - Cloning and mutation analysis of the human IL-18 promoter: a possible role of polymorphisms in expression regulation. AB - Interleukin-18 is a proinflammatory cytokine, which strongly induces IFN-gamma production. We have cloned the human IL-18 promoter and screened for possible polymorphisms. Three single nucleotide polymorphisms were detected in the promoter and two polymorphisms in the 5'-nontranslated region of the gene. Three combinations of these polymorphisms were observed in the Swedish population. All IL-18 promoter alleles were found to have clear promoter activity when inserted into a luciferase reporter vector. There were no significant differences in promoter activity between alleles without stimulation, but after stimulation with PMA/ionomicin one of the alleles had clearly lower activity than the other (P<0.01). Measurement of IL-18 and IFN-gamma production in 48 multiple sclerosis patients by RT-PCR showed slightly higher expression of IL-18 in individuals homozygous for the most frequent haplotype. Two IL-18 promoter polymorphisms were analyzed by sequence specific PCR in 208 multiple sclerosis patients and 139 healthy controls, however, no significant differences were found. In summary, our data indicate that common IL-18 promoter polymorphisms influence the expression of IL-18 and potentially also of IFN-gamma. PMID- 11108944 TI - Immunomodulatory effects of interferon beta-1a in multiple sclerosis. AB - Several studies have established a role for interferon beta (IFNbeta) as a treatment for relapsing-remitting multiple sclerosis (MS). IFNbeta has been reported to decrease the relapse rate, relapse severity, progression of disability and development of new brain lesions. Its mechanisms of action, however, remain unclear. We hypothesize that immunomodulatory effects of IFNbeta may underlie its clinical efficacy. We used intracellular cytokine flow cytometry to analyze the effects of IFNbeta-1a on expression of the anti-inflammatory cytokine, IL-10, and its effects on major co-stimulatory molecules in MS patients. We found that peripheral blood mononuclear cells (PBMC) produced more IL-10 following in vitro or in vivo treatment with IFNbeta-1a. The primary cellular sources of IL-10 were monocytes and CD4(+) T lymphocytes. IL-10 production in response to IFNbeta-1a was increased in unseparated PBMC compared to purified lymphocyte cultures, indicating that interaction between monocytes and lymphocytes may influence IL-10 production in response to IFNbeta-1a. Using flow cytometry, we monitored the ex vivo expression of two major co-stimulatory pairs-B7/CD28 and CD40/CD40L-before and after intramuscular IFNbeta-1a treatment of MS patients. IFNbeta-1a lowered the expression of B7.1 on circulating B cells and increased B7.2 expression on monocytes. CD40 expression on B cells was down regulated, but CD40 on monocytes was up-regulated by IFNbeta-1a treatment. These data suggest that co-stimulatory molecules are modulated by IFNbeta, providing a possible mechanism for its in vivo immune regulatory effects. PMID- 11108945 TI - True epithelial hyperplasia in the thymus of early-onset myasthenia gravis patients: implications for immunopathogenesis. AB - The early-onset myasthenia gravis (EOMG) thymus shows characteristic medullary epithelial bands (MEB), greatly expanded perivascular infiltrates and fenestrations of the intervening basement membranes. We now compare epithelial expression of epidermal growth factor receptor (EGFR) and many integrins in EOMG and control samples. The main differences are striking/consistent thickening (in MEB) of what is normally a monolayer of perivascular epithelium, with focal protrusion into the infiltrates. This evidently hyperplastic epithelial subpopulation also strongly expresses EGFR and certain integrins. We suggest that its enhanced interactions with the locally increased extracellular matrix protein deposits may play an important role in autosensitization. PMID- 11108946 TI - Regulation of chemokine receptor CCR5 and production of RANTES and MIP-1alpha by interferon-beta. AB - Trafficking of inflammatory T cells into the brain is associated with interactions of certain chemokines with their receptors, which plays an important role in the pathogenesis of multiple sclerosis (MS). We examined whether interferon-beta (IFN-beta) had the ability to regulate the production of chemokines and the expression of their receptors in T cells derived from patients with MS. It was demonstrated for the first time that in vitro exposure of T cells to IFN-beta-1a selectively inhibited mRNA expression for RANTES and MIP-1alpha and their receptor CCR5. T cell surface expression of CCR5 was significantly reduced in MS patients treated with IFN-beta, correlating with decreased T cell transmigration toward RANTES and MIP-1alpha. The study provides new evidence suggesting that IFN-beta treatment impairs chemokine-induced T cell trafficking by reducing the production of RANTES and MIP-1alpha and the expression of their receptors CCR5. PMID- 11108947 TI - Features of sensory ataxic neuropathy associated with anti-GD1b IgM antibody. AB - Some reports have called sensory ataxic neuropathy (SAN) associated with IgM antibody against b-series gangliosides a chronic form of Miller Fisher syndrome (MFS), but this has yet to be established. We examined five patients with SAN and eight patients with IgG anti-GQ1b-positive MFS. Only one patient with SAN complained of diplopia, whose ocular movement was not limited. The other four patients had neither diplopia nor limitation of ocular movement. All the SAN patients had severe deep sense impairment, whereas one patient with MFS showed only mild vibratory sense impairment. All sera from the SAN patients had remarkably high IgM antibody titers to the b-series gangliosides GD3, GD2, GD1b, GT1b, GQ1b, GQ1b alpha, fucosyl-GD1b, and alpha galactosyl [alpha fucosyl] GD1b. An absorption study confirmed that the anti-GQ1b antibodies cross-reacted with GD3, GD2, GD1b, and GT1b. In contrast, only two samples from the MFS patients had IgG antibody to GD3, and no sample reacted with GD2, GD1b, or GT1b. SAN has different clinical or serological features from MFS, and therefore is not a chronic form of it. PMID- 11108948 TI - Induction of preprotachykinin-I and neurokinin-1 by adrenocorticotropin and prolactin. Implication for neuroendocrine-immune-hematopoietic axis. AB - We studied the complex interactions within the neuroendocrine-immune hematopoietic axis by determining a possible link among ACTH, PRL, PPT-I and the receptors for its peptides, NK-1 and NK-2. Indeed, ACTH and PRL induced the expression of PPT-I and NK-1 in human bone marrow stroma with no effect on NK-2. Consistent with a role for PPT-I in regulating the development of myeloid and erythroid progenitors, we found that ACTH and PRL, through NK-1 stimulated the proliferation of both types of progenitors. Induction of PPT-I was regulated at the transcriptional and post-transcriptional levels. The results showed that ACTH and PRL stimulated the proliferation of bone marrow progenitors, partly through PPT-I and NK-1 induction. PMID- 11108949 TI - Monocytes in multiple sclerosis: phenotype and cytokine profile. AB - Multiple sclerosis (MS) is an inflammatory demyelinating disease characterised by immune abnormalities in the central nervous system (CNS) as well as systemically. Activated, blood-borne monocytes are abundant in MS lesions, the properties of circulating monocytes are incompletely known. To delineate phenotype and levels of cytokine secreting monocytes in MS patients' blood, ELISPOT assays were used for detection and enumeration of monocytes secreting the cytokines IL-6, IL-12, TNF-alpha and IL-10. In parallel, the expression by monocytes of co-stimulatory molecules (CD40, CD80, CD86), major histocompatibility complex molecules (HLA ABC, HLA-DR) and Fcgamma receptors (CD16, CD64) was examined by flow cytometry. Levels of blood monocytes secreting IL-6 and IL-12 were higher in patients with untreated MS and other neurological diseases (OND) compared to healthy controls, while levels of monocytes secreting TNF-alpha and IL-10 did not differ between groups. MS patients' blood monocytes also displayed elevated mean fluorescence intensity for the co-stimulatory molecule CD86, and MS patients with longer disease duration (>10 years) and higher disease severity (EDSS >3) had higher percentages of CD80 expressing monocytes compared to patients with short duration or lower severity. In conclusion, monocyte aberrations occur in MS and may change over the disease course. PMID- 11108950 TI - Preface. PMID- 11108951 TI - A survey of the sixth European symposium on calcium-binding proteins. Calcium: its flow, its proteins and genomes. PMID- 11108952 TI - Fibrillin-1, a calcium binding protein of extracellular matrix. AB - Fibrillin-1 is a large extracellular matrix glycoprotein which assembles to form 10-12 nm microfibrils in extracellular matrix. Mutations in the human fibrillin-1 gene (FBN-1) cause the connective tissue disease Marfan syndrome and related disorders, which are characterised by defects in the skeletal, cardiovascular and ocular systems of the body. Fibrillin-1 has a striking modular organisation which is dominated by multiple tandem repeats of the calcium binding epidermal growth factor-like (cbEGF) domain. This review focuses on recent studies which have investigated the structural and functional role of calcium binding to cbEGF domains in fibrillin-1 and 10-12 nm microfibrils. PMID- 11108953 TI - The regulation of integrin function by Ca(2+). AB - Integrins are metalloproteins whose receptor function is dependent on the interplay between Mg(2+) and Ca(2+). Although the specificity of the putative divalent cation binding sites has been poorly understood, some issues are becoming clearer and this review will focus on the more recent information. The MIDAS motif is a unique Mg(2+)/Mn(2+) binding site located in the integrin alpha subunit I domain. Divalent cation bound at this site has a structural role in coordinating the binding of ligand to the I domain containing integrins. The I like domain of the integrin beta subunit also has a MIDAS-like motif but much less is known about its cation binding preferences. The N-terminal region of the integrin alpha subunit has been modelled as a beta-propeller, containing three or four 'EF hand' type divalent cation binding motifs for which the function is ill defined. It seems certain that most integrins have a high affinity Ca(2+) site which is critical for alphabeta heterodimer formation, but the location of this site is unknown. Finally intracellular Ca(2+) fluxes activate the Ca(2+) requiring enzyme, calpain, which regulates cluster formation of leucocyte integrins. PMID- 11108954 TI - The biology of the receptor for advanced glycation end products and its ligands. AB - Receptor for advanced glycation end products (RAGE) is a multiligand member of the immunoglobulin superfamily of cell surface molecules whose repertoire of ligands includes advanced glycation end products (AGEs), amyloid fibrils, amphoterins and S100/calgranulins. The overlapping distribution of these ligands and cells overexpressing RAGE results in sustained receptor expression which is magnified via the apparent capacity of ligands to upregulate the receptor. We hypothesize that RAGE-ligand interaction is a propagation factor in a range of chronic disorders, based on the enhanced accumulation of the ligands in diseased tissues. For example, increased levels of AGEs in diabetes and renal insufficiency, amyloid fibrils in Alzheimer's disease brain, amphoterin in tumors and S100/calgranulins at sites of inflammation have been identified. The engagement of RAGE by its ligands can be considered the 'first hit' in a two stage model, in which the second phase of cellular perturbation is mediated by superimposed accumulation of modified lipoproteins (in atherosclerosis), invading bacterial pathogens, ischemic stress and other factors. Taken together, these 'two hits' eventuate in a cellular response with a propensity towards tissue destruction rather than resolution of the offending pathogenic stimulus. Experimental data are cited regarding this hypothesis, though further studies will be required, especially with selective low molecular weight inhibitors of RAGE and RAGE knockout mice, to obtain additional proof in support of our concept. PMID- 11108955 TI - Regulation of G protein-coupled receptor kinase subtypes by calcium sensor proteins. AB - G protein-coupled receptor homologous desensitization is intrinsically related to the function of a class of S/T kinases named G protein-coupled receptor kinases (GRK). The GRK family is composed of six cloned members, named GRK1 to 6. Studies from different laboratories have demonstrated that different calcium sensor proteins (CSP) can selectively regulate the activity of GRK subtypes. In the presence of calcium, rhodopsin kinase (GRK1) is inhibited by the photoreceptor specific CSP recoverin through direct binding. Several other recoverin homologues (including NCS 1, VILIP 1 and hippocalcin) are also able to inhibit GRK1. The ubiquitous calcium-binding protein calmodulin (CaM) can inhibit GRK5 with a high affinity (IC(50)=40-50 nM). A direct interaction between GRK5 and Ca(2+)/CaM was documented and this binding does not influence the catalytic activity of the kinase, but rather reduced GRK5 binding to the membrane. These studies suggest that CSP act as functional analogues in mediating the regulation of different GRK subtypes by Ca(2+). This mechanism is, however, highly selective with respect to the GRK subtypes: while GRK1, but not GRK2 and GRK5, is regulated by recoverin and other NCS, GRK4, 5 and 6, that belong to the GRK4 subfamily, are potently inhibited by CaM, which had little or no effect on members of other GRK subfamilies. PMID- 11108956 TI - Intracellular Ca(2+) release mechanisms: multiple pathways having multiple functions within the same cell type? AB - The elevation of the cytosolic and nuclear Ca(2+) concentration is a fundamental signal transduction mechanism in almost all eukaryotic cells. Interestingly, three Ca(2+)-mobilising second messengers, D-myo-inositol 1,4,5-trisphosphate (InsP(3)), cyclic adenosine diphosphoribose (cADPR), and nicotinic acid adenine dinucleotide phosphate (NAADP(+)) were identified in a phylogenetically wide range of different organisms. Moreover, in an as yet very limited number of cell types, sea urchin eggs, mouse pancreatic acinar cells, and human Jurkat T lymphocytes, all three Ca(2+)-mobilising ligands have been shown to be involved in the generation of Ca(2+) signals. This situation raises the question why during evolution all three messengers have been conserved in the same cell type. From a theoretical point of view the following points may be considered: (i) redundant mechanisms ensuring intact Ca(2+) signalling even if one system does not work, (ii) the need for subcellularly localised Ca(2+) elevations to obtain a certain physiological response of the cell, and (iii) tight control of a physiological response of the cell by a temporal sequence of Ca(2+) signalling events. These theoretical considerations are compared to the current knowledge regarding the three messengers in sea urchin eggs, mouse pancreatic acinar cells, and human Jurkat T lymphocytes. PMID- 11108957 TI - Hierarchical organization of calcium signals in hepatocytes: from experiments to models. AB - The proper working of the liver largely depends on the fine tuning of the level of cytosolic Ca(2+) in hepatocytes. Thanks to the development of imaging techniques, our understanding of the spatio-temporal organization of intracellular Ca(2+) in this - and other - cell types has much improved. Many of these signals are mediated by a rise in the level of inositol 1,4,5-trisphosphate (InsP(3)), a second messenger which can activate the release of Ca(2+) from the endoplasmic reticulum. Besides the now well-known hepatic Ca(2+) oscillations induced by hormonal stimulation, intra- and intercellular Ca(2+) waves have also been observed. More recently, subcellular Ca(2+) increases associated with the coordinated opening of a few Ca(2+) channels have been reported. Given the complexity of the regulations involved in the generation of such processes and the variety of time and length scales necessary to describe those phenomena, theoretical models have been largely used to gain a precise and quantitative understanding of the dynamics of intracellular Ca(2+). Here, we review the various aspects of the spatio-temporal organization of cytosolic Ca(2+) in hepatocytes from the dual point of view provided by experiments and modeling. We first focus on the description and the mechanism of intracellular Ca(2+) oscillations and waves. Second, we investigate in which manner these repetitive Ca(2+) increases are coordinated among a set of hepatocytes coupled by gap junctions, a phenomenon known as 'intercellular Ca(2+) waves'. Finally, we focus on the so-called elementary Ca(2+) signals induced by low InsP(3) concentrations, leading to Ca(2+) rises having a spatial extent of a few microns. Although these small-scale events have been mainly studied in other cell types, we theoretically infer general properties of these localized intracellular Ca(2+) rises that could also apply to hepatocytes. PMID- 11108958 TI - The ALG-2/AIP-complex, a modulator at the interface between cell proliferation and cell death? A hypothesis. AB - During the development of an organism cell proliferation, differentiation and cell death are tightly balanced, and are controlled by a number of different regulators. Alterations in this balance are often observed in a variety of human diseases. The role of Ca(2+) as one of the key regulators of the cell is discussed with respect to two recently discovered proteins, ALG-2 and AIP, of which the former is a Ca(2+)-binding protein, and the latter is substrate to various kinases. The two proteins interact with each other in a Ca(2+)-dependent manner, and the role of the complex ALG-2/AIP as a possible modulator at the interface between cell proliferation and cell death is discussed. PMID- 11108959 TI - The DREAM-DRE interaction: key nucleotides and dominant negative mutants. AB - Transcriptional repressor DREAM, an EF-hand containing calcium-binding protein, blocks basal expression of target genes through specific interaction with DRE sites in the DNA. The sequence GTCA forms the central core of the DRE site, whereas flanking nucleotides contribute notably to the affinity for DREAM. Release of binding of DREAM from the DRE results in derepression, a process that is regulated by Ca(2+). Change of two amino acids within an EF-hand in DREAM blocks Ca(2+)-induced derepression and results in potent dominant negative mutants of endogenous DREAM. PMID- 11108960 TI - The annexinopathies: a new category of diseases. AB - The annexins are a family of highly homologous phospholipid binding proteins, which share a four-domain structure, with one member of the family - annexin VI - having a duplication consisting of eight domains. Thus far, ten annexins have been described in mammals. Although the biological functions of the annexins have not been definitively established, two human diseases involving annexin abnormalities ('annexinopathies') have been identified as of the time of writing. Overexpression of annexin II occurs in the leukocytes of a subset of patients having a hemorrhagic form of acute promyelocytic leukemia. Underexpression of annexin V occurs on placental trophoblasts in the antiphospholipid syndrome and in preeclampsia. Also, an animal model has been described in which annexin VII is underexpressed and is associated with disease, but the relevance of this animal model to human disease is not yet understood. Future research is likely to elucidate additional 'annexinopathies'. PMID- 11108961 TI - Modes of annexin-membrane interactions analyzed by employing chimeric annexin proteins. AB - Annexin II is a member of the annexin family of Ca(2+)- and phospholipid-binding proteins which is particularly enriched on early endosomal membranes and has been implicated in participating in endocytic events. In contrast to other endosomal annexins the association of annexin II with its target membrane can occur in the absence of Ca(2+) in a manner depending on the unique N-terminal domain of the protein. However, endosome binding of annexin II does not require formation of a protein complex with the intracellular ligand S100A10 (p11) as an annexin II mutant protein (PM AnxII) incapable of interacting with p11 is still present on endosomal membranes. Fusion of the N-terminal sequence of this PM AnxII (residues 1-27) to the conserved protein core of annexin I transfers the capability of Ca(2+)-independent membrane binding to the otherwise Ca(2+)-sensitive annexin I. These results underscore the importance of the N-terminal sequence of annexin II for the Ca(2+)-independent endosome association and argue for a direct interaction of this sequence with an endosomal membrane receptor. PMID- 11108962 TI - S100 protein-annexin interactions: a model of the (Anx2-p11)(2) heterotetramer complex. AB - The (Anx2)(2)(p11)(2) heterotetramer has been implicated in endo- and exocytosis in vivo and in liposome aggregation in vitro. Here we report on the modelling of the heterotetramer complex using docking algorithms. Two types of models are generated-heterotetramer and heterooctamer. On the basis of the agreement between the calculated (X-ray) electron density and the observed projected density from cryo-electron micrographs on the one hand, and calculated energy criteria on the other hand, the heterotetramer models are proposed as the most probable, and one of them is selected as the best model. Analysis of this model at an atomic level suggests that the interaction between the Anx2 core and p11 has an electrostatic character, being stabilised primarily through charged residues. PMID- 11108963 TI - S100A1 and S100B interactions with annexins. AB - Members of the annexin protein family interact with members of the S100 protein family thereby forming heterotetramers in which an S100 homodimer crossbridges two copies of the pertinent annexin. Previous work has shown that S100A1 and S100B bind annexin VI in a Ca(2+)-dependent manner and that annexin VI, but not annexin V, blocks the inhibitory effect of S100A1 and S100B on intermediate filament assembly. We show here that both halves of annexin VI (i.e., the N terminal half or annexin VI-a and the C-terminal half or annexin VI-b) bind individual S100s on unique sites and that annexin VI-b, but not annexin VI-a, blocks the ability of S100A1 and S100B to inhibit intermediate filament assembly. We also show that the C-terminal extension of S100A1 (and, by analogy, S100B), that was previously demonstrated to be critical for S100A1 and S100B binding to several target proteins including intermediate filament subunits, is not part of the S100 surface implicated in the recognition of annexin VI, annexin VI-a, or annexin VI-b. Evaluation of functional properties with a liposome stability and a calcium influx assay reveals the ability of both S100 proteins to permeabilize the membrane bilayer in a similar fashion like annexins. When tested in combinations with different annexin proteins both S100 proteins mostly lead to a decrease in the calcium influx activity although not all annexin/S100 combinations behave in the same manner. Latter observation supports the hypothesis that the S100-annexin interactions differ mechanistically depending on the particular protein partners. PMID- 11108964 TI - Transcriptional regulation of S100A1 and expression during mouse heart development. AB - S100A1, a member of the large EF-hand family of Ca(2+)-binding proteins, is mainly expressed in the mammalian heart. To assess the underlying mechanisms for cell- and tissue-specific expression we isolated and characterized the mouse S100A1 gene. The gene displays a high degree of homology to the human and rat genes, especially in the exonic sequences. In its promoter region and the first intron, we identified regulatory elements characteristic for cardiac and slow skeletal muscle restricted genes. Transfection assays with luciferase constructs containing different parts of the S100A1 gene demonstrated the active expression in primary mouse cardiomyocytes and that its 5'-upstream region containing a putative cardiac enhancer showed a greatly increased activity. Furthermore, we investigated the expression of the S100A1 mRNA during embryonic mouse development, using in situ hybridization. S100A1 transcripts were first detected in the primitive heart at embryonic day (E) 8, with equal levels in the atrium and ventricle. During development up to E17.5 we detected a shift in the S100A1 expression pattern with lower levels in atrial and high levels in ventricular myocardium. The regulatory elements identified in the mouse S100A1 promoter correspond well with the observed expression pattern and suggest that S100A1 has an important function during heart muscle development. PMID- 11108965 TI - Calcium-dependent translocation of S100A11 requires tubulin filaments. AB - Protein translocation between different subcellular compartments might play a significant role in various signal transduction pathways. The S100 family is comprised of the multifunctional, small, acidic proteins, some of which translocate in the form of vesicle-like structures upon increase in intracellular Ca(2+) levels. Previously, cells were fixed before and after calcium activation in order to examine the possible relocation of S100 proteins. In this study, we were able to track the real-time translocation. We compared the localization of endogenous S100A11 to that of the S100A11-green fluorescent protein. The application of thapsigargin, an agent increasing intracellular Ca(2+) levels, resulted in the relocation of the S100A11. In contrast, addition of EGTA, which specifically binds Ca(2+), either inhibited the ongoing process of translocation or prevented its induction. Since translocation was not affected by treatment with brefeldin A, it appears that S100A11 relocates in an endoplasmic reticulum Golgi-independent pathway. Furthermore, the depolymerization of actin filaments by amlexanox did not affect the capacity of S100A11 to translocate. However, the time course treatment with demecolcine, which depolymerizes tubulin filaments, resulted in cease of translocation, suggesting that the tubulin network is required for this process. PMID- 11108966 TI - Ca(2+)-binding proteins in the retina: from discovery to etiology of human disease(1). AB - Examination of the role of Ca(2+)-binding proteins (CaBPs) in mammalian retinal neurons has yielded new insights into the function of these proteins in normal and pathological states. In the last 8 years, studies on guanylate cyclase (GC) regulation by three GC-activating proteins (GCAP1-3) led to several breakthroughs, among them the recent biochemical analysis of GCAP1(Y99) mutants associated with autosomal dominant cone dystrophy. Perturbation of Ca(2+) homeostasis controlled by mutant GCAP1 in photoreceptor cells may result ultimately in degeneration of these cells. Here, detailed analysis of biochemical properties of GCAP1(P50L), which causes a milder form of autosomal dominant cone dystrophy than constitutive active Y99C mutation, showed that the P50L mutation resulted in a decrease of Ca(2+)-binding, without changes in the GC activity profile of the mutant GCAP1. In contrast to this biochemically well-defined regulatory mechanism that involves GCAPs, understanding of other processes in the retina that are regulated by Ca(2+) is at a rudimentary stage. Recently, we have identified five homologous genes encoding CaBPs that are expressed in the mammalian retina. Several members of this subfamily are also present in other tissues. In contrast to GCAPs, the function of this subfamily of calmodulin (CaM) like CaBPs is poorly understood. CaBPs are closely related to CaM and in biochemical assays CaBPs substitute for CaM in stimulation of CaM-dependent kinase II, and calcineurin, a protein phosphatase. These results suggest that CaM like CaBPs have evolved into diverse subfamilies that control fundamental processes in cells where they are expressed. PMID- 11108967 TI - Metastasis-associated protein Mts1 (S100A4) inhibits CK2-mediated phosphorylation and self-assembly of the heavy chain of nonmuscle myosin. AB - A role for EF-hand calcium-binding protein Mts1 (S100A4) in the phosphorylation and the assembly of myosin filaments was studied. The nonmuscle myosin molecules form bipolar filaments, which interact with actin filaments to produce a contractile force. Phosphorylation of the myosin plays a regulatory role in the myosin assembly. In the presence of calcium, Mts1 binds at the C-terminal end of the myosin heavy chain close to the site of phosphorylation by protein kinase CK2 (Ser1944). In the present study, we have shown that interaction of Mts1 with the human platelet myosin or C-terminal fragment of the myosin heavy chain inhibits phosphorylation of the myosin heavy chain by protein kinase CK2 in vitro. Mts1 might also bind directly the beta subunit of protein kinase CK2, thereby modifying the enzyme activity. Our results indicate that myosin oligomers were disassembled in the presence of Mts1. The short C-terminal fragment of the myosin heavy chain was totally soluble in the presence of an equimolar amount of Mts1 at low ionic conditions (50 mM NaCl). Depolymerization was found to be calcium dependent and could be blocked by EGTA. Our data suggest that Mts1 can increase myosin solubility and therefore suppress its assembly. PMID- 11108968 TI - S100A6, a calcium- and zinc-binding protein, is overexpressed in SOD1 mutant mice, a model for amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterised by selective degeneration of motoneurones. Familial ALS is an age-dependent autosomal dominant disorder in which mutations in the homodimeric enzyme Cu/Zn superoxide dismutase 1 (SOD1) is linked to the disease. An animal model for this disease is a transgenic mouse expressing the mutated human SOD1(G93A) gene. Recent electrophysiological data emphasised that the striking selective vulnerability of motoneurones might be due to their differential calcium buffering capacities. Therefore we have investigated, using immunohistochemistry, the expression of different calcium binding proteins in brainstem and spinal cord from normal and SOD1 mutated mice. Among the 13 calcium-binding proteins screened, only one, S100A6, a homodimeric calcium-binding protein able to bind four Zn(2+), appeared to be highly expressed in the SOD1 mutated mice. In brainstem, reactive astrocytes, but not motoneurones, from several regions, including nerve 12 root, were highly S100A6-positive. Hypoglossal nucleus was negative for S100A6. In dorsal root, reactive astrocytes from both white matter and anterior horn were highly reactive. If overexpression of S100A6 is specific for ALS, it will be a valuable diagnostic marker for this disease. PMID- 11108969 TI - Mechanism of calcium oscillations in migrating human astrocytoma cells. AB - Numerous studies show that intracellular calcium controls the migration rate of different mobile cell types. We studied migrating astrocytoma cells from two human cell lines, U-87MG and A172, in order to clarify the mechanisms by which calcium potentially influences cell migration. Using the wound-healing model to assay migration, we showed that four distinct components of migration could be distinguished: (i) a Ca(2+)/serum-dependent process; (ii) a Ca(2+) dependent/serum-independent process; (iii) a Ca(2+)/serum-independent process; (iv) a Ca(2+)-independent/serum-dependent process. In U-87MG cells which lack a Ca(2+)-dependent/serum-independent component, we found that intracellular Ca(2+) oscillations are involved in Ca(2+)-dependent migration. Removing extracellular Ca(2+) greatly decreased the frequency of migration-associated Ca(2+) oscillations. Furthermore, non-selective inhibition of Ca(2+) channels by heavy metals such as Cd(2+) or La(3+) almost completely abolished changes in intracellular Ca(2+) observed during migration, indicating an essential role for Ca(2+) channels in the generation of these Ca(2+) oscillations. However, specific blockers of voltage-gated Ca(2+) channels, including nitrendipine, omega conotoxin GVIA, omega-conotoxin MVIIC or low concentrations of Ni(2+) were without effect on Ca(2+) oscillations. We examined the role of internal Ca(2+) stores, showing that thapsigargin-sensitive Ca(2+) stores and InsP(3) receptors are involved in Ca(2+) oscillations, unlike ryanodine-sensitive Ca(2+) stores. Detailed analysis of the spatio-temporal aspect of the Ca(2+) oscillations revealed the existence of Ca(2+) waves initiated at the leading cell edge which propagate throughout the cell. Previously, we have shown that the frequency of Ca(2+) oscillations was reduced in the presence of inhibitory antibodies directed against beta3 integrin subunits. A simple model of a Ca(2+) oscillator is proposed, which may explain how the generation of Ca(2+) oscillations is linked to cell migration. PMID- 11108970 TI - Use of microdialysis in drug delivery studies PMID- 11108971 TI - Methodological issues in microdialysis sampling for pharmacokinetic studies. AB - Microdialysis is an in vivo technique that permits monitoring of local concentrations of drugs and metabolites at specific sites in the body. Microdialysis has several characteristics, which makes it an attractive tool for pharmacokinetic research. About a decade ago the microdialysis technique entered the field of pharmacokinetic research, in the brain, and later also in peripheral tissues and blood. Within this period much has been learned on the proper use of this technique. Today, it has outgrown its child diseases and its potentials and limitations have become more or less well defined. As microdialysis is a delicate technique for which experimental factors appear to be critical with respect to the validity of the experimental outcomes, several factors should be considered. These include the probe; the perfusion solution; post-surgery interval in relation to surgical trauma, tissue integrity and repeated experiments; the analysis of microdialysate samples; and the quantification of microdialysate data. Provided that experimental conditions are optimized to give valid and quantitative results, microdialysis can provide numerous data points from a relatively small number of individual animals to determine detailed pharmacokinetic information. An example of one of the added values of this technique compared with other in vivo pharmacokinetic techniques, is that microdialysis reflects free concentrations in tissues and plasma. This gives the opportunity to assess information on drug transport equilibration across membranes such as the blood-brain barrier, which already has provided new insights. With the progress of analytical methodology, especially with respect to low volume/low concentration measurements and simultaneous measurement of multiple compounds, the applications and importance of the microdialysis technique in pharmacokinetic research will continue to increase. PMID- 11108972 TI - On mathematical models of microdialysis: geometry, steady-state models, recovery and probe radius. AB - Commonly used methods for microdialysis recovery measurement are reviewed and the zero flow and no net flux methods are suggested as the most robust in practice. Six different mathematical models of microdialysis assumptions are investigated and compared for varying dialysis probe radius. One transmitter (dopamine), three metabolites (DOPAC, HVA and 5HIAA) and two drugs (caffeine and theophylline) were studied. Histology and functional response to a drug were measured. Deficiencies were demonstrated for several of the models, the one best explaining experimental data includes both passive diffusion and active tissue regulation in a cylindrical symmetric geometry. The recovery decreased with decreasing probe radius but smaller probes caused less tissue injury. It is concluded that a mathematical model of microdialysis must include diffusional and physiological processes in order to accurately account for experimentally observed phenomena. The experiments also demonstrated that, for small brain nuclei, the size of the nucleus may influence the recovery. PMID- 11108973 TI - Analytical considerations for microdialysis sampling. AB - Adaptations in microdialysis probe designs have made it possible to obtain samples from the extracellular fluid of a variety of tissues with high temporal resolution. The resulting small volume samples, often with low concentration of the analyte(s) of interest, present a particular challenge to the analytical system. Rapid separations can be coupled on-line with microdialysis to provide near real-time data. By combining microdialysis sampling with a liquid chromatographic or capillary electrophoretic separation and a highly sensitive detection method, a separation-based sensor can be developed. Such sensors have been applied to the investigation of drug entities as well as to study endogenous analytes. PMID- 11108974 TI - Microdialysis in peripheral tissues. AB - The objective of this review is to survey the recent literature regarding the applications of microdialysis in pharmacokinetic studies and facilitating many other studies in peripheral tissues such as muscle, subcutaneous adipose tissue, heart, lung, etc. It has been reported extensively that microdialysis is a useful technique for monitoring free concentrations of compounds in extracellular fluid (ECF), and it is gaining popularity in pharmacokinetic and pharmacodynamic studies, both in experimental animals and humans. The first part of this review discusses the use of microdialysis technique for ECF sampling in peripheral tissues in animal studies. The second part of the review describes the use of microdialysis for ECF sampling in peripheral tissues in human studies. Microdialysis has been applied extensively to measure both endogenous and exogenous compounds in ECF. Of particular benefit is the fact that microdialysis measures the unbound concentrations in the peripheral tissue fluid which have been shown to be responsible for the pharmacological effects. With the increasing number of applications of microdialysis, it is obvious that this method will have an important place in studying drug pharmacokinetics and pharmacodynamics. PMID- 11108975 TI - Blood microdialysis in pharmacokinetic and drug metabolism studies. AB - Microdialysis is a sampling technique allowing measurement of endogenous and exogenous substances in the extracellular fluid surrounding the probe. In vivo microdialysis sampling offers several advantages over conventional methods of studying the pharmacokinetics and metabolism of xenobiotics, both in experimental animals and humans. In the first part of this review article various practical aspects related to blood microdialysis will be discussed, such as: probe design, surgical implantation techniques, methods to determine the in vivo relative recovery of the analyte of interest by the probe, special analytical considerations related to small volume microdialysate samples, and pharmacokinetic calculations based on microdialysis data. In the second part of this review a few selected applications of in vivo microdialysis sampling to investigate pharmacokinetic processes are briefly discussed: determination of in vivo plasma protein binding in small laboratory animals, distribution of drugs across the blood-brain barrier, the use of microdialysis sampling to study biliary excretion and enterohepatic cycling, blood microdialysis sampling in man and in the mouse, and in vivo drug metabolism studies. PMID- 11108976 TI - Microdialysis and drug delivery to the eye. AB - The eye presents unique challenges in both the development of tools for elucidating drug disposition as well as for the development of modes of drug delivery for treatment of ocular diseases. In this paper, we present a discussion of the anatomical and physiological characteristics and limitations present in the eye for microdialysis sampling of endogenous substrates and xenobiotics. To date, over twenty papers describing microdialysis approaches for assessment of ocular drug delivery and endogenous substrate characterization have been published. Although the majority of papers describe sampling of vitreous humor, recent efforts have been directed towards ocular anterior segment sampling using microdialysis. With this approach, an appreciable reduction in animal use has been realized. In addition, simultaneous examination of administered drug and endogenous substrates modulated by the drug is possible with this approach, facilitating construction of ocular pharmacokinetic/pharmacodynamic relationships through use of relevant surrogate markers. PMID- 11108977 TI - Application of microdialysis to characterize drug disposition in tumors. AB - Microdialysis is an in vivo sampling technique that was initially developed to measure endogenous substances in the field of neurotransmitter research. In the past decade, microdialysis has been increasingly applied to study the pharmacokinetics and drug metabolism in the blood and various tissues of both animals and humans. This paper describes the general aspects of this in vivo sampling technique followed by the survey of the recent papers regarding the application of microdialysis to characterize anticancer drug disposition in solid tumors. It can be concluded that microdialysis is a very suitable method to obtain drug concentration-time profiles in the interstitial fluid of solid tumors as well as of other variety of tissues. PMID- 11108978 TI - Microdialysis in clinical drug delivery studies. AB - The introduction of in vivo microdialysis (MD) to clinical pharmacological studies has opened the opportunity to obtain previously inaccessible information about the drug distribution process to the clinically relevant target site. The aim of this review is to provide a comprehensive overview of the current literature about MD in drug delivery studies from a clinical perspective. In particular the application of MD in clinical--antimicrobial, oncological and transdermal--and neurological research will be described and the scope of MD in pharmacokinetic-pharmacodynamic (PK-PD) studies will be discussed. It is concluded that MD has a great potential for both academic and industrial research, and may become the method of choice for drug distribution studies in humans. PMID- 11108979 TI - Microdialysis in mice for drug delivery research. AB - Intracerebral microdialysis was first performed in the mouse at the end of the 1980s. Most microdialysis studies on mice were confined to neuropharmacology and changes in neurotransmitter concentrations up to 1995, although pharmacological studies were done on other tissues like the skin, kidney and implanted tumors. The use of microdialysis in mice for pharmacokinetic and drug delivery studies owes much to the recent availability of genetically engineered mice, such as mice in which the genes encoding multiple drug resistance have been knocked out. The quantitative microdialysis of blood and various tissue fluids of the mouse is now feasible and the recent development of specific microdialysis devices for use in mice should facilitate its use in these small animals. This review covers the technical aspects of microdialysis in the mouse and includes references to many of the published studies on pharmacokinetics and drug delivery. PMID- 11108980 TI - The use of microdialysis in CNS drug delivery studies. Pharmacokinetic perspectives and results with analgesics and antiepileptics. AB - Microdialysis can give simultaneous information on unbound drug concentration time profiles in brain extracellular fluid (ECF) and blood, separating the information on blood-brain barrier (BBB) processes from confounding factors such as binding to brain tissue or proteins in blood. This makes microdialysis suitable for studies on CNS drug delivery. It is possible to quantify influx and efflux processes at the BBB in vivo, and to relate brain ECF concentrations to central drug action. The half-life in brain ECF vs. the half-life in blood gives information on rate-limiting steps in drug delivery and elimination from the CNS. Examples are given on microdialysis studies of analgesic and antiepileptic drugs. PMID- 11108981 TI - Microdialysis in the study of drug transporters in the CNS. AB - Quantitative microdialysis in the central nervous system (CNS) has recently provided evidence for the existence of transporters as they relate to the brain distribution of a variety of drugs. Support for the existence of drug transporters in the blood-brain barrier (or in the blood-CSF barrier) comes from investigations that have found: unbound drug concentrations in brain fluids that are lower than corresponding levels in plasma; saturability of transport clearances across the blood-brain barrier and; the regulation of transport by putative inhibitors. Additional confirmatory evidence for the existence of active transport or carrier-mediated processes has also been derived from models that relate observed drug levels in the CNS with those in plasma or blood. The conclusion that reduced drug levels in brain fluids generally indicate the existence of active efflux transport is questioned. In the case of relatively polar compounds with modest blood-brain barrier permeability, lower unbound concentrations in brain may be a consequence of dilution by turnover of brain fluids. This review summarizes recent reports (grouped by class of compounds) where investigators have used microdialysis to examine the distribution of therapeutic agents to the CNS, and have reached conclusions regarding the functional presence of drug transporters in the brain. PMID- 11108982 TI - Confocal microscopic analysis reveals sprouting of primary afferent fibres in rat dorsal horn after spinal cord injury. AB - Following high thoracic spinal cord transection (SCT) in rats, abnormal changes in arterial pressure in response to sensory stimulation (autonomic dysreflexia) are correlated with changes in neural circuitry in the injured spinal cord. Anterograde transport of wheat germ agglutinin conjugated to Texas Red (WGATR) and confocal microscopy were used to characterize the increased arbourization of Adelta and Abeta fibre populations in laminae III-V of the dorsal horn. In cord injured animals, significantly greater areas of WGATR-labeled fibres were found in the deeper laminae of the dorsal horn than in control rats. This increased area likely reflects sprouting of the Adelta, Abeta, and possibly C fibre populations. The time course of sprouting matches the onset of autonomic dysreflexia, indicating a possible functional correlation between the two phenomena. PMID- 11108983 TI - CP 55,940 protects against ischemia-induced electroencephalographic flattening and hyperlocomotion in Mongolian gerbils. AB - The effect of CP 55,940, on electroencephalographic (EEG) spectral power decrease and hyperlocomotion induced by transient global ischemia in gerbils, was investigated. Animals were treated with CP 55,940 (4 mg/kg intraperitoneal (i.p.)) 5 min after bilateral carotid occlusion or SR 141716A (3 mg/kg i.p.) 5 min before or with both. Mean total and relative spectral power was evaluated for 1 h before (basal) and 1 and 24 h, 3 and 7 days after ischemia. Spontaneous locomotor activity was evaluated at the same times. CP 55,940 antagonized the reduction in mean total spectral power and the hyperlocomotion induced by ischemia, in comparison with vehicle group, starting from 24 h and lasting 7 days (P<0.001). Pretreatment with SR 141716A completely blocked the neuroprotective effect of CP 55,940. These findings suggest a potential therapeutic role of cannabinoids in cerebral ischemia. PMID- 11108984 TI - Ciliary neurotrophic factor increases muscle fiber number in the developing levator ani muscle of female rats. AB - In addition to its well characterized neurotrophic properties, ciliary neurotrophic factor (CNTF) also has myotrophic effects in several experimental paradigms. We have previously observed that the volume of the levator ani (LA) muscle is increased in female rats treated with CNTF during the perinatal period. In order to determine the cellular basis for the effect of CNTF on LA muscle volume, female rat pups were given daily perineal injections of CNTF or a control solution from postnatal day 1 through 6. Mean cross-sectional area of LA muscle fibers and LA fiber number were assessed on postnatal day 7. CNTF treatment increased LA muscle fiber number more than 300% while having no effect on LA fiber size. We conclude that CNTF prevents muscle fiber degeneration and/or increases myogenesis in the developing LA muscle. PMID- 11108986 TI - The effects of ascorbic acid on dopamine-induced death of PC12 cells are dependent on exposure kinetics. AB - The role of ascorbic acid on dopamine (DA) oxidation-mediated cytotoxicity was studied using the PC12 cell line. DA cytotoxicity was slightly attenuated by ascorbic acid, whereas the cytotoxicity of 6-hydroxydopamine (6-OHDA), a DA oxidation product, was markedly potentiated. To elucidate the relationship between the ascorbic acid effect and the degree of DA oxidation, ascorbic acid was added in a time-dependent fashion after DA treatment. We found greater cell death the later ascorbic acid was applied. Treatment of cells with glutathione alleviated DA- and 6-OHDA-induced cell death, while L-buthionine sulfoximine potentiated DA and 6-OHDA cytotoxicity. Ascorbic acid combined with glutathione eliminated the toxicity of DA and 6-OHDA. These results suggest that the interaction between DA and ascorbic acid is dependent upon the degree of DA oxidation and glutathione availability. PMID- 11108985 TI - Dopamine D(2)-like receptor binding in the brain of male Japanese quail (Coturnix japonica). AB - Japanese quail (Coturnix japonica) have been used extensively to study appetitive behaviors. However, little is known about the appetitive-relevant neurochemical systems in this species. The present investigation examined the distribution of D(2)-like dopamine receptors in the quail brain. [(3)H]Spiperone was incubated in brain tissue homogenates and non-specific binding was defined using (-) sulpiride. Scatchard analysis of whole brain without cerebellum and forebrain alone indicated approximate K(d)'s of 0.08 and 0.04 nM, respectively. In addition, the preferential D(3) agonist (+/-)-2-dipropylamino-7-hydroxy-1,2,3, 4 tetrahydronaphthalene hydrobromide (7-OH-DPAT) did not displace [(3)H]spiperone binding in quail forebrain. Finally, regional analysis showed that the highest densities of D(2)-like receptors were located in the forebrain. Overall, these results indicate that there is some conservation of dopaminergic mechanisms between aves and mammals. Thus, Japanese quail may be useful for examining the neuropharmacological mechanisms of dopaminergic stimulant drugs that work via D(2)-like receptor activation. PMID- 11108987 TI - Prenatal choline supplementation alters hippocampal N-methyl-D-aspartate receptor mediated neurotransmission in adult rats. AB - Manipulation of dietary choline levels in pregnant rats has been shown to result in enduring alterations in memory and hippocampal function of the offspring, but the mechanisms underlying these effects remain unclear. Hippocampal slices were prepared from adult rats that were offspring of dams fed control, choline supplemented, or choline deficient diets on days 12-17 of gestation. N-methyl-D aspartate (NMDA) receptor-mediated population excitatory postsynaptic potentials (pEPSPs) were pharmacologically isolated and evoked using electrical stimulus pulses applied to s. radiatum of area CA1. Evoked NMDA receptor-mediated pEPSPs were enhanced in slices from prenatally choline supplemented relative to controls in both male and female rats. The greatest differences occurred at the low end of the input-output curve, among responses that were less than 60% of maximal. These results are discussed in the context of previous behavioral and electrophysiological studies. PMID- 11108988 TI - Lethal forebrain ischemia stimulates sphingomyelin hydrolysis and ceramide generation in the gerbil hippocampus. AB - Ceramide, a hydrolyzed product of sphingomyelin, is reported to play an important role in apoptosis. In this study, we measured the sphingomyelin and ceramide levels in the hippocampus of the gerbil after transient forebrain ischemia for 5 min (lethal) or 2 min (sublethal). The aim was to examine alterations in the sphingomyelin cycle during delayed neuronal death, which we considered could be due to apoptosis. Sphingolipids were separated on high-performance thin-layer chromatography plates and analyzed by gas-liquid chromatography. At 30 min and 24 h after lethal ischemia, sphingomyelin levels were decreased and ceramide levels were increased compared with control levels. No significant changes were observed after sublethal ischemia. These results suggest that the sphingomyelin cycle may have a role in neuronal death. PMID- 11108989 TI - Confocal quality imaging of afferent neurons from semi-thin sections of Drosophila ganglia. AB - The aim of this study was to develop protocols for computer imaging of the thoraco-abdominal ganglion of Drosophila from serial semi-thin sections, in which specific neurons were stained and related to neuropilar structures. The central projections of a subset of transgenically labelled sensory neurons were revealed by immunohistochemistry, while Nomarski optics were used to show motor neuron targets in the neuropil. Digital photomicrographs of each section were aligned and the resultant image stacks rendered into three-dimensional (3D) images that can be rotated in real time. The result is a detailed, in-depth visualization of labelled neurons at a resolution comparable with that in confocal reconstructions, which also allows investigation of their relationships with other components of the neuropil. PMID- 11108990 TI - Accelerated functional recovery and neuroprotection by agmatine after spinal cord ischemia in rats. AB - Treatment with agmatine, decarboxylated arginine, proved to be non-toxic and to exert neuroprotective effects in several models of neurotoxic and ischemic brain and spinal cord injuries. Here we sought to find out whether agmatine treatment would also prove beneficial in a rat spinal cord ischemia model (balloon occlusion of the abdominal aorta bellow the branching point of the left subclavian artery for 5 min). Agmatine was injected (100 mg/kg, i.p. ) 5 min after beginning of re-perfusion and again once daily for the next 3 post operative days. Motor performance ('combined motor score') was recorded for up to 17 days post-operative and motoneuron cell counts (in representative spinal cord sections) performed on the 17th post-operative day. Agmatine treatment was found to accelerate recovery of motor deficits and to prevent the loss of motoneurons in the spinal cord after transient ischemia. Together, the present and previous findings demonstrate that agmatine is an efficacious neuroprotective agent and that this naturally occurring non-toxic compound should be tried for therapeutic use after neurotrauma and in neurodegenerative diseases. PMID- 11108991 TI - An N-methyl-D-aspartate antagonist, MK-801, preferentially reduces electroconvulsive shock-induced phosphorylation of p38 mitogen-activated protein kinase in the rat hippocampus. AB - Electroconvulsive shock (ECS) activates the mitogen-activated protein kinase (MAPK) family in the rat hippocampus, but the signaling pathways for this activation are not well understood. We investigated whether N-methyl-D-aspartate (NMDA) receptor mediated signaling is involved in the phosphorylation-activation of the MAPK family. The NMDA receptor antagonist, MK-801, dose-dependently reduced ECS-induced phosphorylation of p38 and its upstream kinase MKK6 up to 1 mg/kg. MK-801 also reduced the phosphorylation of ERK1/2 and MEK1, but only at high dosage, 2 mg/kg. Moreover, the reduction in the phosphorylation of p38 and MKK6 was greater than that of ERK1/2 and MEK1. Our results suggest that ECS activates p38 and ERK1/2 partly through an NMDA receptor-mediated signaling system in the rat hippocampus and that NMDA receptor mediated signaling is more responsible for the activation of the MKK6-p38 pathway than the MEK1-ERK pathway. PMID- 11108992 TI - Directional rectification of gap junctional voltage gating between dieters cells in the inner ear of guinea pig. AB - Deiters cells (DCs) are the cochlear supporting cells in inner ear and contain multiple gap junction connexin genes, which when mutated can induce hearing loss. In the present study, the gap junctions between DCs were investigated by a double voltage clamp technique. Besides asymmetric responses to the polarities of transjunctional voltage (V(j)) and transmembrane potential (V(m)), the channels were also sensitive to which cell side was stimulated in a cell pair, i.e. voltage gating had directional dependence. The direction-dependent voltage gating could result in asymmetric current flow between the cells and influenced K(+) passage. Multiple connexins may constitute non-homotypic channels with directional dependence of voltage gating to mediate functional gap junction pathways in the cochlea. This may explain how a single connexin mutation can produce hearing loss. PMID- 11108993 TI - Stimulation of the mesencephalic locomotor region inhibits the discharge of neurons in the superficial laminae of the dorsal horn of cats. AB - In decerebrate cats, we found that stimulation of the mesencephalic locomotor region (MLR) attenuated the responses of neurons in the superficial laminae of the dorsal horn to thin fiber muscle afferent input. The attenuation appeared to be more effective for group III afferent input than for group IV. These findings may shed light on the interaction between central command, (i.e. the MLR) and the muscle reflex, mechanisms which both contribute to the cardiovascular responses to exercise. PMID- 11108994 TI - Reduced production of lactate during hypoxia and reoxygenation in astrocytes isolated from stroke-prone spontaneously hypertensive rats. AB - Lactate production and expressions of monocarboxylate transporter 1 (MCT1) and lactate dehydrogenase (LDH) mRNA after hypoxia and reoxygenation (H/R) were examined by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) using astrocytes in culture isolated from stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar Kyoto rats (WKY). The basal production of lactate in SHRSP was the same as that observed in WKY. In contrast the lactate levels in SHRSP at 1 and 6 h of reoxygenation after hypoxia were significantly lower than those observed in WKY. In addition LDH and MCT1 mRNA expressions in SHRSP were significantly less strong compared with those in WKY during H/R. These findings indicate that decreased production and slow transport of lactate in SHRSP astrocytes are involved in neuronal energy depletion and possibly encourage neuronal damage, although hereditary weakness of cortical neurons is also related to cell death during H/R. PMID- 11108995 TI - The N-methyl-D-aspartate antagonistic and opioid components of d-methadone antinociception in the rat spinal cord. AB - The d-enantiomer of the opioid methadone is a weak opioid with low micromolar affinity to the N-methyl-D-aspartate (NMDA) receptor. We have investigated the antinociception and in vivo NMDA antagonism after systemic administration of d methadone in the rat spinal cord. d-Methadone caused antinociception in the Randall-Selitto model of inflammatory pain and inhibited the responses of hindlimb single motor units to noxious electrical and mechanical stimulation (ED(50) 6.6, 6.8 and 7.2 mg/kg intravenous (i.v.), respectively); the wind-up of these responses was only inhibited at the dose almost completely abolishing the baseline responses. d-Methadone inhibited the activity of spinal dorsal horn neurones evoked by both iontophoretic NMDA and (R, S)-alpha-amino-3-hydroxy-5 methyl-4-isoxazole propionic acid (AMPA, ED(50) 5.7 and 8.2 mg/kg i.v., respectively). After pre-treatment with naloxone, d-methadone was unable to inhibit nociception in the Randall-Selitto model, the NMDA- or AMPA-evoked neuronal activity or the motoneurone wind-up. Thus, in the antinociceptive dose range, the NMDA antagonism does not appear to contribute to the mechanism of d methadone antinociception. PMID- 11108996 TI - Nociceptin is a potent inhibitor of N-type Ca(2+) channels in rat sympathetic ganglion neurons. AB - Nociceptin, an endogenous agonist of the opioid receptor-like(1) (ORL(1)) receptor, is implicated in a wide range of physiological functions including cardiovascular control. However, the effect of nociceptin on peripheral sympathetic ganglion neurons has not been studied. Whole-cell voltage clamp was used to study Ca(2+) currents on freshly dissociated sympathetic superior cervical ganglion neurons from juvenile rats. Nociceptin (1 microM) caused a fast inhibition of the peak currents by 69+/-3% in all neurons. Strong positive prepulses counteracted the inhibition of the peak current by 64% and no effect of nociceptin was observed when the cells were pre-incubated with Pertussis toxin. The inhibition was reversible and dose-dependent with an EC(50) of 508+/-50 pM. Blockade of N-type channels by 1 microM omega-conotoxin GVIA reduced the peak currents by 83+/-1% and abolished the action of nociceptin. Naloxone could not prevent the inhibition by nociceptin and [D-Ala(2), N-Me-Phe(4), Gly(5)-ol] enkephalin (DAMGO) only depressed a small proportion of the current in 1/7 neurons. These data suggests that nociceptin inhibits transmitter release from sympathetic neurons by a selective blockade of N-type channels, which may be of importance for its depressive effect on the cardiovascular system. PMID- 11108997 TI - Transient up-regulation of gamma-aminobutyric acid(A) receptor binding by lubeluzole after neocortical specify lesion in rats. AB - The effect of the neuroprotective drug lubeluzole on cortical receptor binding was investigated in animals with photothrombotic ischemic lesions. Control animals were treated with the inactive stereoisomer of the drug. Lubeluzole was applied intravenously as a single bolus (0.31 mg/kg) followed by a 1-h infusion of 0.31 mg/kg. Lubeluzole selectively increased gamma-amino-butyric acid(A) (GABA(A)) receptor binding but had no significant and/or consistent effects on N methyl-D-aspartate (NMDA), (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4 propionic acid (AMPA), and kainate receptors. Lubeluzole caused a significant up regulation of GABA(A) receptor binding in the lesioned area as well as in unimpaired cortical areas of both hemispheres. This effect appeared in the hours following the lesion and peaked at 24 h. Our findings suggest that reduced cortical excitability brought about by increased binding capacities of GABA(A) receptors may contribute to the neuroprotective effect of lubeluzole. PMID- 11108998 TI - Cloning and expression of MP13 gene from rat hippocampus, a new factor related to guanosine triphosphate regulation. AB - C-Fos and the Fos-related antigens (FRA) are induced by various stimuli. A novel 35-37 kDa FRA was induced much longer after the treatment using kainic acid (KA) and may be very important for neuronal survival after brain damage. To identify this long-term FRA, we have constructed a cDNA library derived from hippocampus after KA treatment and screened it with an antibody highly conserved M-peptide region of FRAs. One gene, MP13, was cloned with a 1662 bp open reading frame and coded for a 554-amino acid protein. MP13 has a leucine zipper region, a glutamine repeat region, and has high similarity to the activator of the small guanosine triphosphate (GTP)ase Rab5. Gel retardation analysis revealed that MP13 functions as a GTP regulation related factor. PMID- 11108999 TI - Recruitment plasticity of neuromuscular compartments in exercised tibialis anterior using echo-planar magnetic resonance imaging in humans. AB - We investigated the recruitment plasticity of the superficial tibialis anterior (TA-s) and deep tibialis anterior (TA-d) regions of neuromuscular compartments (NMCs) in the m. tibialis anterior (TA) during exercise using echo-planar imaging (EPI). Six healthy men performed dorsiflexion exercise at 60% of maximum voluntary contraction at a frequency of 10 contractions/min inside the magnetic resonance imaging. Transaxial EPIs of the right leg were acquired every 6 s at rest (0.5 min), during exercise (2.5 min) and recovery (5 min). In TA-s, significantly higher signal intensities (SIs) were shown than those in TA-d from immediately after starting the exercise to recovery. It has been demonstrated that SI reflects the degree of recruitment in the activated muscle, thus our result suggest that preferential firing of motor neurons in the superficial region of the NMC occurs during exercise in human TA muscle. PMID- 11109000 TI - Spatial memory and learning deficits after experimental pneumococcal meningitis in mice. AB - Survivors of bacterial meningitis frequently suffer from long-term sequelae, particularly from learning and memory deficits. For this reason, spatial memory and learning was studied in a mouse model of ceftriaxone-treated Streptococcus pneumoniae meningitis. Persistent deficits of spatial learning despite normal motor function were observed in mice infected with 10(4) colony-forming units (CFU) in 25 microl of saline into the right forebrain in comparison to mice treated with an equal amount of saline. Survivors of meningitis performed significantly worse in memorizing a hidden platform in a Morris water maze. After 2 weeks, the difference between post-meningitis and control mice diminished. Yet, when the platform was moved after 180 days, learning of the new location was still strongly impaired in mice surviving meningitis. PMID- 11109001 TI - 5-Hydroxytryptamine(2A) and 5-hydroxytryptamine(1B) receptors are differently affected by the monoamine oxidase A-deficiency in the Tg8 transgenic mouse. AB - In brainstem-spinal cord preparations of neonatal control C3H and transgenic Tg8 mice where deletion of the gene encoding monoamine oxidase-A results in serotonin (5-hydroxytryptamine (HT)) excess, whole cell recordings of identified phrenic motoneurons (Phr Mns) were performed to study the modulation of their activity by 5-HT. In C3H mice, a dual effect was observed: (i) a facilitation via 5-HT(2A) receptors and (ii) a decrease of the transmission of the central inspiratory drive via 5-HT(1B) receptors. In Tg8 mice, the 5-HT(2A)-mediated facilitation was present but the 5-HT(1B)-mediated decrease was lacking. Therefore, the conservation of the 5-HT(2A) response vs. the loss of the 5-HT(1B) one suggest that the two types of receptors respond differently to 5-HT level changes. PMID- 11109002 TI - Hypobaric hypoxia induces fos and neuronal nitric oxide synthase expression in the paraventricular and supraoptic nucleus in rats. AB - This study examined the effects of high altitude exposure on neurons in the paraventricular nucleus (PVN) and supraoptic nucleus (SON) of the hypothalamus in adult and neonatal rats. In adult control rats, occasional Fos-like immunoreactive neurons were localized in both the hypothalamic nuclei. A marked increase in Fos positive cells was induced at 1-4 h following altitude exposure but it was reduced to levels comparable to the controls at 24 h. The expression of neuronal nitric oxide synthase (nNOS) immunoreactivity in the PVN and SON followed a similar temporal pattern. The nNOS immunoreactivity, which was constitutively expressed in the hypothalamic neurons in the control rats, was noticeably augmented at 1-4 h, but it was comparable to the controls at 24 h following altitude exposure. In postnatal rats, Fos expression was not detected in the hypothalamic neurons of the controls. Induction of Fos expression was observed in some neurons at 1-4 h following altitude exposure but it was diminished at 24 h. There was no noticeable change in nNOS expression in both the control and altitude exposed postnatal rats; in both instances, it was barely detectable. It is concluded that both the PVN and SON of the adult rats are activated at high altitude exposure and that they may be involved in the regulation of neuroendocrine, cardiovascular and respiratory functions in hypobaric hypoxia. This study has also shown the differential response of the hypothalamus neurons between the two age groups to the hypoxic insult. Our results suggest that the adult neurons are probably more sensitive to the reduced oxygen levels in hypobaric hypoxia, as reflected by the upregulated NOS expression in this age group but not in the postnatal rats. PMID- 11109003 TI - The influence of pallidal deep brain stimulation on striatal dopaminergic metabolism in the rat. AB - Deep brain stimulation of the globus pallidus internus has been recently shown to alleviate parkinsonian symptoms and levodopa-induced dyskinesias. However, its exact mechanisms of action are unclear. Pallidal neurones are connected via various pathways to the dopaminergic nigrostriatal system. In the present study we investigated the hypothesis that deep brain stimulation of the entopeduncular nucleus (corresponds to the human internal pallidum) affects striatal dopaminergic metabolism in naive and 6-hydroxydopamine (6-OHDA) lesioned rats using microdialysis. Our results show that stimulation of the entopeduncular nucleus does not significantly affect striatal dopamine metabolism (of dopamine, 3, 4-dihdroxyphenylacetic acid and homovanillic acid) in naive and 6-OHDA lesioned animals. They contrast with our previous observations that deep brain stimulation of the subthalamic nucleus increases striatal dopamine metabolism suggesting differential effects of these nuclei on striatal dopamine metabolism. PMID- 11109004 TI - Mediation of the cardiovascular response to spinal gamma-aminobutyric acid(B) receptor stimulation by adenosine A(1) receptors in anesthetized rats. AB - Cardiovascular inhibitory effects induced by intrathecal (i.t.) administration of adenosine A(1) receptor agonist and its modulation by gamma-aminobutyric acid(B) (GABA(B)) receptor was suggested by our previous report. In this experiment, we examined the mediation of cardiovascular effects of GABA(B) receptor stimulation by adenosine A(1) and A(2) in the spinal cord. I.t. administration of GABA(B) receptor agonist, baclofen (30, 60 and 100 nmol) produced a dose dependent decrease of blood pressure and heart rate. Pretreatment with adenosine A(1) receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (50 nmol), attenuated the depressor and bradycardiac effects of baclofen (100 nmol), but not with adenosine A(2) receptor antagonist, 3, 7-dimethyl-1-propargylxanthine (25 nmol). These results suggest that GABA(B) receptors in the spinal cord play an inhibitory role in the central cardiovascular regulation and that the depressor and bradycardiac actions are mediated by adenosine A(1) receptors. PMID- 11109005 TI - Roles of glutamate receptor subtypes in the development of vestibular compensation after unilateral labyrinthectomy in the guinea pig. AB - We investigated the roles of ionotropic glutamate receptor subtypes in the development and recovery of spontaneous nystagmus (SN) after unilateral labyrinthectomy (UL) in guinea pigs. When administered at 3 h after UL, N-methyl D-aspartate (NMDA) and kainate (KA), which are NMDA and non-NMDA receptor agonists, respectively, increased the frequency of SN. The effect of KA was more potent than that of NMDA. In contrast to these agonists, MK-801 and CNQX decreased the frequency of SN. Although the administration of KA at 48 h after UL increased the frequency of SN, it did not exhibit any effects at 72 h after UL. MK-801 caused a recurrence of SN following administration at 48 and 72 h after UL. Neither NMDA nor CNQX exhibited any effects after administration at 48 or 72 h after UL. A newly synthesized compound, NC-1200, which has inhibitory action on the glutamate response, decreased the frequency of SN in a dose-dependent manner following administration at 3 h after UL, but did not exhibit any effects when administered at 48 and 72 h after UL. From these results, it was found that NMDA and non-NMDA receptors play important roles in the development of SN after UL, and that the NMDA receptor contributes to the development of ocular motor compensation. PMID- 11109006 TI - A pore-lining glutamic acid in the rat olfactory cyclic nucleotide-gated channel controls external spermine block. AB - Spermine is a potent, voltage-dependent blocker of the olfactory cyclic nucleotide-gated channel from both the intracellular and extracellular sides. However, its sites of action are unknown. This study investigated the external spermine binding site in the rat CNCalpha3 subunit. Neutralization of a glutamic acid residue (E342Q) in the P-loop region eliminated voltage-dependence of block by externally applied spermine. The charge-conservative E342D mutation had little effect on spermine block. Thus, E342 forms the binding site for externally applied spermine. However, spermine remained a potent voltage-independent blocker of the E342Q mutant channel, suggesting that the mutation either created a novel binding site outside the membrane electrical field or that it dramatically changed the properties of the existing pore site. PMID- 11109007 TI - Mutation analysis of the N-methyl-D-aspartate receptor NR1 subunit gene (GRIN1) in schizophrenia. AB - Dysfunction of N-methyl-D-aspartate (NMDA) type ionotropic glutamate receptors has been implicated in the etiology of schizophrenia based on psychotomimetic properties of the antagonist phencyclidine (PCP) and observation that mice expressing low levels of the N-methyl-D-aspartate receptor NR1 subunit exhibit behavioral alterations that may be ameliorated by neuroleptic drugs. Based on the hypothesis that some schizophrenic patients have functionally deficient mutation(s) of the gene encoding N-methyl-D-aspartate receptor NR1 subunit (GRIN1), we screened 48 Japanese patients with schizophrenia for mutations in the coding region of the GRIN1 gene. Four variants, IVS2-22T>C, IVS2-12G>A, IVS4 34C>T, and 1719G/A (Pro516Pro), were identified. No non-synonymous mutation was detected. No significant association was suggested by case-control comparisons. Results indicate that genomic variations of the GRIN1 gene are not likely to be involved substantially in the etiology of schizophrenia. PMID- 11109008 TI - Pure autonomic failure in association with human alpha-synucleinopathy. AB - We studied an autopsy case with pure autonomic failure, using anti-alpha synuclein antibody. Until now there has been no report about the immunohistochemical properties of alpha-synuclein in pure autonomic failure. In conventional stainings, both pre- and post-ganglionic lesions of the sympathetic and parasympathetic nervous systems were found. Lewy bodies and Lewy neurites were abundant especially in the sympathetic nervous system. These inclusions were immunoreactive to anti-alpha-synuclein antibody. The intensity of alpha-synuclein immunoreactivity was stronger in the halos than in the cores of the Lewy bodies. The edges of the swollen neurites had strong immunoreactivity. The substantia nigra was well preserved, and no cortical Lewy bodies were seen. These findings indicate that pure autonomic failure is one of the Lewy body type alpha synucleinopathies, such as Parkinson's disease and dementia with Lewy bodies, targeting the peripheral autonomic nervous system. PMID- 11109009 TI - Association of apolipoprotein E polymorphism to clinical heterogeneity of multiple sclerosis. AB - In this study we investigated the distribution of apolipoprotein E (APO E) genotypes in sporadic multiple sclerosis (MS) cases and in normal controls. Later onset of chronic progressive MS was observed in patients carrying the epsilon2 allele, whereas APO E alleles were found at similar frequency in MS and in the control population. These findings indicate that clinical heterogeneity, but probably not susceptibility to the disease, is associated to APO E genotypes. PMID- 11109010 TI - Four year prospective evaluation of nosocomial bacteremia: epidemiology, microbiology, and patient outcome. AB - A prospective study of all patients with clinically significant nosocomial bacteremia at one institution from 1994 to 1997 was performed to: (1) describe the epidemiology and microbiology of nosocomial bacteremias; (2) determine the crude mortality associated with such infections; and (3) identify independent predictors of mortality. Four hundred four episodes of bacteremia occurred in 322 patients; the crude in-hospital mortality was 31%. Coagulase-negative staphylococci, Staphylococcus aureus, and enterococci were the leading pathogens, and intravascular catheters were the most frequently identified source. The highest mortality occurred in patients with candidemia (67%). Independent predictors of mortality included evidence of shock at the time of infection, acquisition of bacteremia in an intensive care unit, a "Do Not Attempt Resuscitation" order, and the presence of certain comorbid conditions (e.g., malignancy, HIV infection). Because many of these infections may be preventable, education of health care providers and strict adherence to established infection control practices are critical. PMID- 11109011 TI - Characterization of Burkholderia pseudomallei isolated in Thailand and Malaysia. AB - A total of 35 Burkholderia pseudomallei isolates from Thailand (16 clinical and eight soil isolates) and Malaysia (seven animal, two isolate each from clinical and soil) were investigated by their antimicrobial resistance, plasmid profiles and were typed by randomly amplified polymorphic DNA analysis. All isolates were found to be resistant to six or more of the 12 antimicrobial agents tested. Only two small plasmids of 1.8 and 2.4 megadalton were detected in two clinical isolates from Thailand. RAPD analysis with primer GEN2-60-09 resulted in the identification of 35 RAPD-types among the 35 isolates. The constructed dendrogram differentiated the 35 isolates into two main clusters and a single isolate. The wide genetic biodiversity among the 35 isolates indicate that RAPD-PCR can be a useful method to differentiate unrelated B. pseudomallei in epidemiological investigation. PMID- 11109012 TI - "Room temperature" use of CHROMagar Candida. AB - The color of colonies of 9 Candida species was examined on the chromogenic medium CHROMagar Candida incubated for 24-72 h at 25 degrees C, 30 degrees C or 37 degrees C. Colors and colony forms characteristic of C. albicans, C. dubliniensis, C. krusei and C. tropicalis were formed most rapidly and with the deepest hues at 37 degrees C. After 48 h incubation at 25 degrees C, 9 of 48 C. albicans isolates gave pink colonies instead of the green colonies characteristic for the species, and the blue-purple colony color characteristic of C. tropicalis isolates was not formed until 48 h at 25 degrees C. Incubation of the chromogenic medium at temperatures below 30 degrees C cannot be recommended for reliable presumptive identification of Candida spp., and pink colonies of C. glabrata would not be reliably distinguished from pink colonies formed by other species under any of the incubation conditions used. PMID- 11109013 TI - The use of Monte Carlo simulation to examine pharmacodynamic variance of drugs: fluoroquinolone pharmacodynamics against Streptococcus pneumoniae. AB - BACKGROUND: For fluoroquinolones, AUC:MIC ratios correlate with maximal bacterial eradication in in vitro models of infection and favorable cure rates in humans with respiratory tract infection. Inter-subject pharmacokinetic and MIC variability may impact the probability of attaining optimal AUC:MIC ratios and hence favorable clinical outcome. METHODS: Monte Carlo simulation was utilized to estimate the probability of attaining AUC:MIC ratios of 30, 40, 50, 60, 70, 80, 90, 100, 110 and 120 using AUC values from patients treated with either gatifloxacin or levofloxacin and microbiologic activity against S. pneumoniae observed in 1997 SENTRY Antimicrobial Surveillance Program. RESULTS: The probability curves for 5000 patient simulations were plotted. The median AUC:MIC ratios were 120 for gatifloxacin and 50.5 for levofloxacin. The probability of attaining AUC:MIC ratios of 30, 50, 70 and 100 for gatifloxacin were 94%, 86%, 78% and 62%, and for levofloxacin were 80%, 51%, 31% and 17%, respectively. CONCLUSION: Gatifloxacin has a higher probability of achieving target AUC:MIC ratios than levofloxacin. Monte Carlo simulation, using patient-based AUC and MIC distributions, may have implications for selection of optimal antibiotics for the empiric treatment of infections. Moreover, Monte Carlo simulation may have utility in the determination of MIC breakpoints. PMID- 11109014 TI - In vivo development of decreased susceptibility to vancomycin in clinical isolates of methicillin-resistant Staphylococcus aureus. AB - To investigate the possibility of in vivo development of decreased vancomycin susceptibility, the vancomycin susceptibilities of 12 methicillin-resistant Staphylococcus aureus (MRSA) isolates serially recovered from six patients with vancomycin therapy were tested by standard MIC determination method and population analysis. While all of the MRSA isolates were susceptible to vancomycin (MICs, 1-2 microg/ml) by standard method, population analysis showed the upward shifts indicating decreased vancomycin susceptibility among serial isolates from two patients. These bacteria with decreased vancomycin susceptibility could be selected by using vancomycin selection of pre-therapy isolates under laboratory conditions. Furthermore, the reversion phenomenon of decreased vancomycin susceptibility was confirmed after 20 serial passages of the post-therapy isolates on drug-free agar. These data suggest that in vivo isolates may develop decreased vancomycin susceptibility that is not of such magnitude to cross a breakpoint threshold. This resistance may be unstable, and appears to result from a selective or inducible process that occurs in MRSA clinical strains during vancomycin therapy. PMID- 11109015 TI - Feculent meningitis: polymicrobial meningitis in colorectal surgery. AB - Polymicrobial anaerobic meningitis is a rare event secondary to a contiguous infection in the head or neck. Anaerobic meningitis due to a meningo-intestinal fistula is a rare event with only two cases reported in the literature. We describe a new case of adult polymicrobial anaerobic meningitis after colorectal surgery and radiotherapy and review the previous two cases. PMID- 11109016 TI - Group A streptococcal appendicitis in a patient with AIDS. AB - A man with AIDS developed appendicitis and bacteremia caused by Group A streptococcus, neither of which is considered an opportunistic infection. Group A streptococcus is rarely implicated in appendicitis in children and has not previously been reported in an adult. Immunodeficiency might have predisposed the patient to this unusual infection. PMID- 11109017 TI - Cutaneous infection due to Mycobacterium kansasii. AB - A patient presented with chronic leg ulcers after a mowing accident. He received several courses of antibiotics for presumed cellulitis, underwent surgical debridement, and was treated empirically with cyclosporin for presumed pyoderma gangrenosum, all without improvement. Cultures from prior debridement revealed Mycobacterium kansasii, and he was successfully treated with triple antituberculous regimen. Cutaneous infections due to this slow growing Mycobacterium are rare and may resemble cellulitis or sporotrichosis. Mycobacterium kansasii should be included in the differential diagnosis of skin infections with an indolent course and lack of response to antibiotics. PMID- 11109018 TI - In vitro activity of GAR-936 against vancomycin-resistant enterococci, methicillin-resistant Staphylococcus aureus and penicillin-resistant Streptococcus pneumoniae. AB - We report the activity of the new glycylcycline antimicrobial agent GAR-936 against 37 clinical isolates of vancomycin-resistant enterococci (including organisms carrying the vanA, vanB, vanC-1, and vanC-2/3 genes), 26 clinical isolates of methicillin-resistant S. aureus and 30 clinical isolates of high level penicillin-resistant S. pneumoniae. All isolates of vancomycin-resistant enterococci, methicillin-resistant S. aureus, and penicillin-resistant S. pneumoniae were inhibited by < or = 1, < or = 2, or < or = 0.25 microg/ml of GAR 936, respectively. Time kill experiments using vancomycin-resistant enterococci did not demonstrate synergy or antagonism between 2 microg/ml of GAR-936 and 0.25 microg/ml of quinupristin/dalfopristin. PMID- 11109019 TI - Anti-anaerobic activity of levofloxacin alone and combined with clindamycin and metronidazole. AB - Microdilution MICs of levofloxacin against twelve anaerobes ranged between 0.5 8.0 microg/ml and those of clindamycin and metronidazole between 0.008-2.0 and 0.25->16.0 microg/ml, respectively. Combination of levofloxacin with clindamycin and/or metronidazole in time-kill tests led to synergy at levofloxacin concentrations at or below the MIC in 7/12 strains. PMID- 11109020 TI - A new diagnostic problem: isolation of Escherichia coli O157:H7 strains with aberrant biochemical properties. AB - Over a five-year period (1995-1999) the Microbial Diseases Laboratory received 34 strains of E. coli O157:H7 each with a single aberrant biochemical property. In addition, 27 O157 strains with negative or delayed motility were noted during the same time period. These observations suggest that there may be an increased likelihood to misdiagnose O157:H7 infections using commercial systems in the future due to increasing phenotypic variability. PMID- 11109021 TI - Antimicrobial susceptibility of Abiotrophia adiacens and Abiotrophia defectiva. AB - The susceptibilities of 27 Abiotrophia adiacens (proposed reclassification Granulicatella adiacens comb.nov., Collins & Lawson, 2000) and 12 Abiotrophia defectiva isolates were tested by microdilution in pyridoxal hydrochloride and lysed horse blood supplemented Mueller-Hinton broth. According to NCCLS interpretative criteria for Streptococcus spp. not Streptococcus pneumoniae, the susceptibilities of A. adiacens and A. defectiva were, respectively: penicillin, 55% and 8%; amoxicillin, 81% and 92%; ceftriaxone, 63% and 83%; meropenem, 96% and 100%; and 100% for both species with clindamycin, rifampin, levofloxacin, ofloxacin, quinupristin/dalfopristin, and vancomycin. PMID- 11109022 TI - The recovery from alcohol problems over the life course: the Lundby longitudinal study, Sweden. AB - The aim of this paper is to 'look behind the statistics' and to study alcohol related problems over time and to portray the recovery (treated and untreated) from these problems over the life course in a subset of 41 males, defined as DSM IV alcoholics from the Lundby general population project. The subjects were interviewed with semi-structured interviews by a psychiatrist. There was only minimal attrition. An analysis was made of the distribution of alcohol-related problems among the current alcoholic group compared to the recovered one at follow-up. Due to the limited number of subjects the calculations were mainly descriptive. The dominant problems were medical (64% vs. 25%), mental (45% vs. 17%), family (27% vs. 0%) and work (18% vs. 8%). The overall functioning assessed by the GAF scale implied a change from continuous to transient symptoms and problems in the recovered group. The subjects' attributions of reasons for recovery were social stabilization (84%), treatment (58%), family and peer pressure (58%) and medical complications (33%). It is argued that the qualitative findings might be used to complement the large statistical investigations in order to understand how to link alcohol-related problems and social consequences to the life course studies. PMID- 11109023 TI - Differential responsiveness to alcohol odor in human neonates: effects of maternal consumption during gestation. AB - Previous human and animal studies have shown that specific memories arise during prenatal life as a function of fetal processing of chemosensory stimuli present in the amniotic fluid. Furthermore, the animal literature indicates that fetal exposure to alcohol modifies subsequent neonatal and infantile responsiveness toward the sensory attributes of the drug. The main goal of the present study was to analyze whether differential maternal alcohol consumption during pregnancy affects neonatal reactivity to ethanol odor. To achieve this goal, an initial experiment examined how healthy human neonates respond to olfactory stimuli. In this first experiment, newborns (postpartum age: 24-48 h) were evaluated in terms of motor responsiveness elicited by repetitive stimulation with either alcohol or lemon odors. Infants exhibited a marked increase in duration of overall body and head and facial activity when the odorants were first presented. In four successive trials, behavioral responsiveness progressively declined until reaching baseline values. The introduction of a novel odorant served to dishabituate the motor responses under analysis. In the second experiment newborn's reactivity to the mentioned odorants was evaluated as a function of maternal self-reported patterns of consumption of alcohol during gestation. Mothers were classified as frequent or infrequent drinkers. Babies born to frequent drinkers exhibited heightened reactivity toward ethanol odor when compared to newborns delivered by infrequent drinkers. No differences emerged when comparing the responses of both groups of infants elicited by a non-ethanol stimulus such as lemon. The results coupled with prior human and animal research suggest the possibility of intrauterine experience with alcohol odor that yields a sensory memory likely to be retrieved immediately after birth. PMID- 11109024 TI - Conditioned place aversion to the "hangover" phase of acute ethanol administration in the rat. AB - The purpose of this study was to examine ethanol's delayed effects (termed hangover) using conditioned place testing. Four groups of rats received a single pairing of a distinctive environment (tactile and visual) 10 h after injection with ethanol (0, 2, 3, 4 g/kg, i.p. ) or saline in a counterbalanced design. Rats receiving 3 and 4 g/kg ethanol showed a conditioned place aversion to ethanol hangover. Conditioning 10 h after 0 or 2 g/kg ethanol did not produce a significant place preference or aversion. The results suggest that the hangover following an acute injection of high doses of ethanol (3-4 g/kg) produces a significant and dose-related conditioned place aversion in the rat. PMID- 11109025 TI - Scanning of five chromosomes for alcohol consumption loci. AB - In our present genetic study to map Quantitative Trait Loci (QTLs) for alcohol related behaviors, we used 44 B6.C and 36 B6.I inbred congenic Recombinant QTL Introgression (RQI) mouse strains of the b5i7 series carrying genes of BALB/cJ (C) or CXBI (I) origin on C57BL/6ByJ (B6) genetic background. Ethyl alcohol consumption (EAC) was measured in adult males, and chromosomes 1, 2, 3, 9, and 15 were scanned with polymorphic microsatellite markers. In the B6.C set of strains, multiple regression analysis yielded a model with three microsatellite markers, which explained 32% of the genetic variance (p=0.0006). The two markers with the highest significance levels in the model, D1Mit167 and D2Mit74, have been mapped to chromosome regions close to the gene opioid receptor kappa 1 (chr. 1) and opioid receptor kappa 3 (chr. 2), respectively. The results of this gene-mapping study suggest that genetic polymorphisms in kappa opioid receptors may contribute to genetic predisposition to voluntary alcohol-drinking behavior. PMID- 11109026 TI - Cellular immunity after ethanol exposure and burn injury: dose and time dependence. AB - Acute ethanol exposure prior to burn injury increases the immune dysfunction seen with burn alone, which has been partially attributed to increased circulating and splenic macrophage production of interleukin-6 (IL-6). The current studies examined the effect dose and timing of ethanol exposure prior to burn on cellular immunity. Mice with high (300 mg/dl) circulating levels of ethanol at the time of burn demonstrated further suppression of the delayed type hypersensitivity (DTH) and splenocyte proliferative responses in comparison to mice with moderate (100 mg/dl) ethanol levels. Interestingly, the increase in macrophage IL-6 secretion seen at the moderate dose was not augmented at the high dose; however, the circulating IL-6 levels did reveal a further increase at the high ethanol dose. There were no alterations in splenocyte subset populations and/or apoptosis at the moderate vs. the high ethanol dose. Moderate ethanol exposure 24 h, in comparison to 30 min, before injury resulted in similar decreases in the DTH. These results suggest that the dose-dependent effects of ethanol on immunity following burn injury are not the result of splenic macrophage IL-6 production as shown at the moderate dose and that the immune suppressive effects of ethanol in this model persist after it is cleared from the circulation. PMID- 11109027 TI - Quality of life measures and outcome in alcohol-dependent men and women. AB - A sample of 82 (41 men 41 women) DSM IV alcohol-dependent inpatients admitted for detoxification was studied at baseline and followed-up 12 weeks thereafter. The following questionnaires were administered 4-5 days after admission for detoxification: Socio-demographic information, Severity of Alcohol Dependence Questionnaire (SADQ), Alcohol Problems Questionnaire (APQ), Rotterdam Symptoms Checklist (RSCL), Life Situation Survey (LSS), Beck Depression Inventory (BDI), General Health Questionnaire (GHQ 12), and Nottingham Health Profile (NHP). All indices other than socio-demographic data, the SADQ, and APQ were administered at 12-week follow-up. After controlling for confounding factors at baseline, women were more likely to be in a higher social class, prescribed anti-depressants during the previous 12 months, drink fewer units of alcohol in a typical week, and have a higher level of psychiatric caseness scores (GHQ-12). A total of 80 subjects (97%) were successfully followed-up. Difference between gender did not significantly impact upon any of the 12-week outcome measures. There was no significant difference in the study relapse rates or time taken to relapse between men and women. The only significant total sample change was a reduction in the amount of alcohol consumed in a typical week. This was significantly related to changes in the following Quality of Life (QoL) measures, NHP emotional reaction sub-scores, LSS, and BDI scores. PMID- 11109028 TI - Augmentation of corticosterone release by means of a caffeine-ethanol interaction in rats. AB - We examined whether the acute treatment with caffeine delivered before an ethanol injection would augment plasma corticosterone (CORT) levels. The effect of caffeine on blood ethanol levels was also assessed. After 10 days of acclimatization to the colony room conditions, male Wistar rats were injected with either caffeine (5 mg/kg, ip) or saline 30 min before the delivery of ethanol (0.8 g/kg, ip) or saline, respectively. Trunk blood was then collected at 15 and 30 min after the ethanol injection for determination of plasma CORT and blood ethanol levels. CORT was measured with the use of radioimmunoassay, and blood ethanol levels were determined with the use of gas chromatography. The results showed that although caffeine and ethanol delivered singly failed to augment plasma CORT levels, the combination of both drugs produced elevations in plasma CORT levels at 15 and 30 min. These findings were found to be unrelated to changes in ethanol metabolism as caffeine failed to alter blood ethanol levels within the period tested. It was argued that the present elevations in plasma CORT levels observed in animals administered caffeine and ethanol may play a role in the caffeine-induced elevations in ethanol drinking observed elsewhere. PMID- 11109029 TI - Screening for hypercholesterolaemia versus case finding for familial hypercholesterolaemia: a systematic review and cost-effectiveness analysis. AB - BACKGROUND: In the majority of people with familial hypercholesterolaemia (FH) the disorder is caused by a mutation of the low-density lipoprotein receptor gene that impairs its proper function, resulting in very high levels of plasma cholesterol. Such levels result in early and severe atherosclerosis, and hence substantial excess mortality from coronary heart disease. Most people with FH are undiagnosed or only diagnosed after their first coronary event, but early detection and treatment with hydroxymethylglutaryl-coenzyme (HMG CoA) reductase inhibitors (statins) can reduce morbidity and mortality. The prevalence of FH in the UK population is estimated to be 1 in 500, which means that approximately 110,000 people are affected. OBJECTIVES: To evaluate whether screening for FH is appropriate. To determine which system of screening is most acceptable and cost effective. To assess the deleterious psychosocial effects of genetic and clinical screening for an asymptomatic treatable inherited condition. To assess whether the risks of screening outweigh potential benefits. METHODS: DATA SOURCES: Relevant papers were identified through a search of the electronic databases. Additional papers referenced in the search material were identified and collected. Known researchers in the field were contacted and asked to supply information on unpublished or ongoing studies. INCLUSION/EXCLUSION CRITERIA: SCREENING AND TREATMENT: The review included studies of the mortality and morbidity associated with FH, the effectiveness and cost of treatment (ignoring pre-statin therapies in adults), and of the effectiveness or cost of possible screening strategies for FH. PSYCHOSOCIAL EFFECTS OF SCREENING: The search for papers on the psychological and social effects of screening for a treatable inherited condition was limited to the last 5 years because recent developments in genetic testing have changed the nature and implications of such screening tests. Papers focusing on genetic testing for FH and breast cancer were included. Papers relating to the risk of coronary heart disease with similarly modifiable outcome (non-FH) were also included. DATA EXTRACTION AND ASSESSMENT OF VALIDITY: A data assessment tool was designed to assess the quality and validity of the papers which reported primary data for the social and psychological effects of screening. Available guidelines for systematically reviewing papers concentrated on quantitative methods, and were of limited relevance. An algorithm was developed which could be used for both the qualitative and quantitative literature. MODELLING METHODS: A model was constructed to investigate the relative cost and effectiveness of various forms of population screening (universal or opportunistic) and case-finding screening (screening relatives of known FH cases). All strategies involved a two-stage process: first, identifying those people with cholesterol levels sufficiently elevated to be compatible with a diagnosis of FH, and then either making the diagnosis based on clinical signs and a family history of coronary disease or carrying out genetic tests. Cost effectiveness has been measured in terms of incremental cost per year of life gained. RESULTS: MODELLING COST-EFFECTIVENESS: FH is a life-threatening condition with a long presymptomatic state. Diagnostic tests are reasonably reliable and acceptable, and treatment with statins substantially improves prognosis. Therefore, it is appropriate to consider systematic screening for this condition. Case finding amongst relatives of FH cases was the most cost-effective strategy, and universal systematic screening the least cost-effective. However, when targeted at young people (16 year olds) universal screening was also cost effective. Screening patients admitted to hospital with premature myocardial infarction was also relatively cost-effective. Screening is least cost-effective in men aged over 35 years, because the gains in life expectancy are small. (ABSTRACT TRUNCA PMID- 11109030 TI - A randomised controlled trial of prehospital intravenous fluid replacement therapy in serious trauma. AB - RESULTS: In total 1309 patients were entered in the study: 699 (53.4%) were treated by paramedics operating protocol A and 610 (46.6%) were treated by paramedics operating protocol B. The randomisation worked well and there were no significant differences between treatment groups in incident characteristics, ambulance performance times, or patient or injury characteristics, apart from slightly more moderate or severe head injuries in the protocol A group (25.3% versus 20.3%). Protocol compliance was poor, with only 31% of protocol A patients receiving prehospital fluids and only 80% of protocol B patients not given fluids. The estimated odds ratio for being given prehospital fluids when treated by protocol A compared to protocol B was 2.09 (95% confidence interval (CI), 1.53 to 2.81). MORTALITY: There were 73 deaths within 6 months in the 699 patients in the protocol A group (10.4%), and 60/610 (9.8%) in the protocol B group. Thus the crude odds ratio for deaths when managed by protocol A was 1.07 (95% CI, 0.73 to 1.54). Excluding 26 patients whose cause of death may not have been trauma related, the odds ratio was 1.04 (95% CI, 0.69 to 1.55). Excluding 17 patients who may have been dead on arrival of the ambulance at the scene the odds ratio was 1.04 (95% CI, 0.70 to 1.53). Adjustment for age, injury severity and whether the patient was unconscious at the scene did not significantly alter these odds ratios. COMPLICATIONS: A total of 106 patients were identified from hospital notes as having at least one of eight major complications (adult respiratory distress syndrome, sepsis, acute renal failure, coagulopathy, wound infection, pneumonia, fat embolism or pulmonary embolism). The proportions with recorded complications were similar in the two groups: 60/699 (8.5%) in the protocol A group versus 46/610 (7.5%) in the protocol B group. HEALTH STATUS: A total of 878 questionnaires were sent to patients, and 559 (64%) usable replies were received. The response rate was similar in the two groups (62.9% versus 64.6%). In all eight dimensions of the SF-36 health status measure patients who had been managed by paramedics operating protocol A reported better average health than did patients in the protocol B group. However, none of the differences were at a level considered clinically important and only for one of the eight dimensions was the difference statistically significant. COMPOSITE OUTCOMES: No significant differences in outcome were found between the two protocol groups in terms of patients who either died or had serious complications, nor for patients who either died or had known poor health. SUBGROUPS: Subgroups of patients were defined on eight characteristics (ambulance service area, whether a doctor was on scene, paramedic-patient contact time, injury severity, whether taken to theatre for emergency surgery, type of injuries, type of area, and whether the patient was treated before or after protocol cross-over). There was no evidence of any difference in mortality rates or composite outcomes between any subgroups, or between protocols within any subgroup. Time to A&E department The analysis suggests that patients given fluids spent 12-13 minutes longer at the accident scene than did patients not given fluids. However, because only one-quarter of patients were given fluids, and the specific protocol used made little difference to this, average on-scene times were largely unaffected by protocols. COSTS: In the prehospital and immediate-care phase (including A&E treatment), the mean costs of the protocol A and protocol B groups were ?419 and ?416, respectively. This small difference reflects two small and offsetting effects of protocol B: reduced on-scene time (p = 0.08) and increased use of blood in the A&E department (p = 0.03). There were no other statistically significant differences in costs, with the mean total costs being ?2706 and ?2678 in the protocol A and protocol B groups, respectively (p = 0.52). (ABSTRACT TRUNCA PMID- 11109031 TI - Intravascular ultrasound-guided interventions in coronary artery disease: a systematic literature review, with decision-analytic modelling, of outcomes and cost-effectiveness. AB - BACKGROUND: Intravascular ultrasound (IVUS) is the generic name for any ultrasound technology used in vivo within the blood vessels. More specifically, intracoronary ultrasound enables imaging of the coronary arteries from within the lumen. This review concentrates on the role of intracoronary ultrasound as an adjunct to interventional cardiology. OBJECTIVES: (1) To identify the literature on IVUS for guiding coronary interventions, and to synthesise evidence about outcomes compared with outcomes when IVUS guidance has not been used. (2) To use this evidence, together with other information about costs and outcomes, to model the cost effectiveness of IVUS guidance. (3) To synthesise the evidence on the reproducibility of measurements of cross-sectional area made using IVUS. METHODS: DATA SOURCES: (1) Electronic searches of MEDLINE, EMBASE, Science Citation Index, Index to Scientific and Technical Proceedings, Engineering Compendex, Engineering Page One, Cochrane Library, Inside (British Library), 1990-98. (2) Contacting experts and centres of expertise, 1990-99. (3) Internet search, 1990 99. METHODS: STUDY SELECTION: Studies of IVUS-guided coronary interventions performed on humans were included in the review. Non-English language studies were also included when they covered IVUS-guided stenting or angioplasty. Control evidence regarding outcomes without IVUS guidance was sought only from randomised controlled trials (RCTs). Studies investigating the reproducibility of measurements of cross-sectional area were included only if the results were expressed in terms of the mean and standard deviation of paired differences. METHODS: DATA EXTRACTION: Checklists that covered study details, patient characteristics and results were completed independently by three reviewers. Consensus was reached on any disagreements. Local data were gathered on the costs of IVUS-guided stenting. METHODS: DATA SYNTHESIS: Overall event rates were calculated by pooling patient results from the included studies. A decision analytic model was used to combine information from the literature with cost estimates, in order to predict cost-effectiveness in terms of cost per restenosis event avoided by the use of IVUS guidance. The analysis was performed from the perspective of the healthcare provider. Sensitivity analysis was undertaken. A simple extrapolation was made to long-term outcome so that cost-utility (using quality-adjusted life years (QALYs)) could be estimated. The minimum detectable change in cross-sectional area was estimated from the reproducibility results. RESULTS: Only one study on IVUS-guided angioplasty satisfied the inclusion criteria, and there were no studies on IVUS-guided atherectomy or other IVUS guided interventions that satisfied the inclusion criteria. Of the 15 articles on IVUS-guided stenting that satisfied the inclusion criteria, seven presented data on outcomes at 6 months post-intervention. The angiographic restenosis rate was 16 +/- 1%. This compared with 24 +/- 2% derived from five articles on stenting without IVUS guidance. Data for follow-up periods longer than 6 months were presented in only two studies. Data from a total of five studies were included in the decision-analytic model. The cost per restenosis event avoided was 1545 pound sterling. After extrapolation to long-term outcome, the calculated cost per QALY was 6438 pound sterling. The baseline QALY gain was only 0.03 years. Sensitivity analysis resulted in large differences between the best- and worst-case scenarios, for example, from a saving of 5000 pound sterling to a cost of 24,000 pound sterling restenosis event avoided. The smallest changes in cross-sectional area that could be measured were 1.6 mm2 by a single observer and 1.9 mm2 by different observers. CONCLUSIONS: Implications for healthcare: The evidence available is too weak for there to be any reliable implications for clinical practice. (ABSTRACT TRUNCATED) PMID- 11109032 TI - Progress but not yet there. PMID- 11109033 TI - A balanced reinvestment in health. PMID- 11109034 TI - Chimeric technologies. PMID- 11109035 TI - Results of the BRAT study--a pilot study investigating the possible significance of ASA nonresponsiveness on the benefits and risks of ASA on thrombosis in patients undergoing coronary artery bypass surgery. AB - BACKGROUND: Several studies suggest that acetylsalicylic acid (ASA) is less effective in preventing thrombotic events in ASA nonresponder patients. If so, the thrombotic event rate in ASA nonresponders should be higher than in ASA responders. OBJECTIVE: To conduct a prospective, multicentre observational pilot study to determine the thrombotic event rates in ASA responders and nonresponders. PATIENTS AND METHODS: Patients undergoing nonurgent coronary artery bypass grafting (CABG) who were prescribed 325 mg ASA/day were recruited. Patients were classified as an ASA responder or nonresponder based on the ASA effect (or lack thereof) on their bleeding times. All thrombotic events that occurred in the two years following CABG were recorded. These data were stored in a blinded fashion until the last patient follow-up, and then adjudicated by a validation committee. RESULTS: A total of 289 patients recruited at three sites completed the two-year follow-up. Of these patients, 45.3% were classified as ASA responders and 54.7% were classified as ASA nonresponders. Of ASA responders, 6.9% had thrombotic events compared with 9.5% of the ASA nonresponders, but this difference was not significant (P=0.526). CONCLUSIONS: While ASA responder or nonresponder status did not appear to affect the thrombotic event rate in patients undergoing nonurgent CABG, the possibility that ASA responder or nonresponder status affects the thrombotic event rate in more acutely ill CABG patients cannot be excluded. PMID- 11109036 TI - Thebesian sinusoids: forgotten collaterals to papillary muscles. AB - BACKGROUND: Papillary muscles of the left ventricle are prone to ischemic damage but seldom rupture, perhaps because they are protected by transendocardial diffusion and thebesian sinusoids as well as by arteries. Sinusoids are primitive vessels that precede coronary arteries as nutrient suppliers to all parts of the embryonic heart. OBJECTIVES: To determine whether sinusoids penetrate to the tips of papillary muscles. MATERIALS AND METHODS: Adult hearts with and without evidence of acute and chronic ischemic lesions were selected. Papillary muscle sinusoids were perfused by black ink from the apex of the left ventricle. Papillary muscles and the adjacent ventricular free walls were then excised and fixed before gross and microscopic examination for inked sinusoids. Some fixed specimens were cleared through methyl salicylate and examined stereoscopically. RESULTS: The meshwork of trabeculae and, importantly, intertrabecular spaces lining the apex of the left ventricle continue into the deeply undercut bases of papillary muscles. In most hearts, sinusoids can be shown to extend from the undercut bases to the tips of human papillary muscles, especially in ischemic hearts. The compact myocardium of the adjacent ventricular free wall contains few sinusoids. CONCLUSIONS: Sinusoids may supplement normal arterial flow to papillary muscles and mitigate the effects of coronary occlusion. The authors hypothesize that the apex of the human left ventricle is adapted for sinusoidal flow to papillary muscles. PMID- 11109037 TI - Clips versus suture technique: is there a difference? AB - INTRODUCTION: Coronary artery bypass grafting (CABG) is one of the most common procedures performed today, and wound complications are a major source of morbidity and cost. OBJECTIVE: To determine whether there is any difference in wound outcome (including cost in a Canadian context) between a subcuticular suture technique and skin stapling technique for closure of sternal and leg incisions in CABG patients. PATIENTS AND METHODS: One hundred and sixty-two patients undergoing CABG were prospectively, randomly placed to have their sternal and leg incisions closed with either a subcuticular suture technique or with a skin clip. Data were obtained through chart review, in-hospital assessments and follow-up visits. Nonblinded assessments were made regarding wound leakage, inflammation, infection, necrosis, swelling, dehiscence and cosmesis. Each of the parameters was graded on a scale from 1 to 4. The cost was evaluated in Canadian dollars. RESULTS: There were trends toward increased rates of in-hospital sternal (P=0.09) and leg (P=0.17) incision inflammation when the wounds were closed with skin clips. There was a significantly greater (P=0.05) rate of sternal wound infection with clips, as well as a tendency (P=0.15) toward a greater rate of mediastinitis at follow-up assessment. Cosmetic outcome was similar for both groups. The cost incurred was significantly greater when skin clips were used for closure. There was a greater than threefold difference, which translates to a greater than $10,000 difference over one year. CONCLUSIONS: Closure with a subcuticular technique achieves better outcomes than the use of skin clips. When factoring in the increased cost incurred by using clips, as well as other intangible factors such as surgical skill acquisition, subcuticular suture closure appears to be a favourable method of wound closure in CABG patients compared with the use of skin stapling techniques. PMID- 11109038 TI - Differential morphometric and ultrastructural remodelling in the left atrium and left ventricle in rapid ventricular pacing-induced heart failure. AB - BACKGROUND: Heart failure induced by rapid ventricular pacing (RVP) is associated with left atrial (LA) but not left ventricular (LV) hypertrophy. OBJECTIVE: To determine whether differences in wall tension correlate with the differential ultrastructural remodelling in the LA and LV chambers, changes in ultrastructure, systolic function and wall tension (an index of wall stress) were compared in dogs after RVP (n=7) and with no RVP (n=9). RESULTS: Compared with dogs with no RVP (controls), dogs with RVP had increased collagen volume fraction (5.3% versus 8.3%), myocyte cross-sectional area (245 versus 366 microm(2)) and hydroxyproline (222 versus 323 microg/mg protein) in the LA (all P<0.05), but not in the LV. The increase in systolic wall tension produced by RVP was greater in the LA (five versus 43 units, P<0.0004) than in the LV (227 versus 290 units, P<0.01) chambers and correlated closely with the collagen volume fraction (r=0.87), which in turn correlated with myocyte cross-sectional area (r=0.98). In the left atrium, wall tension correlated with wall stress (r=0.99). CONCLUSIONS: The results suggest that differential wall tension may provide the stimulus for differential ultrastructural remodelling (with more hypertrophy and collagen) between LA and LV chambers in RVP-induced cardiomyopathy. PMID- 11109039 TI - Guidelines for the early management of acute coronary syndromes: focus on antithrombotic and antiplatelet therapy. AB - Despite important advances in the management of non-ST segment elevation acute coronary syndromes, adverse outcomes remain common. The recent introduction of low molecular weight heparins and platelet glycoprotein IIb/IIIa receptor inhibitors is an opportunity to make a further impact on the mortality and morbidity of this common condition. Optimal use of these agents will likely result from the recognition of patients at a higher risk of an adverse outcome who are most likely to benefit. At the same time, identification of lower risk patients will avoid the use of unnecessary and potentially harmful medications. Guidelines for the application of these new agents were developed at three meetings of a multidisciplinary group of health professionals. Evidence for risk stratification of patients with non-ST segment acute coronary syndromes and the results of clinical trials for the individual antithrombotic and antiplatelet agents were reviewed. A practical algorithm for patient management is presented, which uses observations available to the physician in the first few hours after the onset of chest pain. PMID- 11109040 TI - Pulmonary effects of low dose amiodarone: a review of the risks and recommendations for surveillance. AB - Previous studies have reported an incidence of amiodarone-induced pulmonary toxicity (AIPT) of 5% to 10% with high doses of amiodarone (greater than 400 mg daily). A lower rate of 1.6% is recorded from combined placebo controlled, double blind trials involving 3439 patients receiving daily amiodarone doses of 400 mg or less. Although the rate of diagnosis of AIPT appears to be lower than previously reported, it is still considerable, and its consequences are potentially fatal if undiagnosed. Before amiodarone is initiated, baseline chest x-ray (CXR) and pulmonary function tests should be performed. Although follow-up surveillance with CXR at three- to six-month intervals has been recommended, pulmonary toxicity can develop rapidly, and radiographic abnormalities may not precede clinical toxicity. Repeat lung function testing should be reserved for patients who develop new symptoms or CXR findings. Patient self-reporting of symptoms and regular clinical evaluation are likely the easiest and most useful strategies for prompt detection of AIPT. PMID- 11109041 TI - The Robert E Beamish Award. PMID- 11109042 TI - High-dose chemotherapy in breast cancer -- the perils of history uncontrolled. AB - Breast cancer remains a common and devastating disease that affects approximately 180,000 women and results in more than 43,000 deaths annually in the United States. Although only 10% of patients have overt metastatic disease at the time of diagnosis, as many as one third of those who present with lymph node-negative disease and half of those who present with lymph node-positive disease eventually develop metastatic breast cancer. With few exceptions, metastatic breast cancer is largely incurable, and the median duration of survival remains 18 to 24 months. Over the past 3 decades, both laboratory and clinical efforts to increase survival have focused on dose intensity in chemotherapy regimens. PMID- 11109043 TI - Gender verification no more? PMID- 11109044 TI - Addressing obesity in medical practice: is weight loss medically beneficial? PMID- 11109046 TI - Risk factors for osteoporosis: A review. AB - Skeletal fragility and falls are the 2 most potent factors leading to osteoporotic fractures. The aim of this article is to review factors associated with women's risk of developing skeletal fragility and subsequent osteoporosis. Many factors have been implicated, but the evidence for some is unsubstantial. Low premenopausal bone mineral density (BMD), a decrease in BMD, and an increase in bone fragility -- which occur as a result of both aging and the menopause -- are major determinants of subsequent risk for osteoporotic fracture. In addition, low body mass index (BMI), low calcium intake, low physical activity, and smoking can affect BMD. The relative importance of the effects these physical and lifestyle factors have on BMD in midlife women is not fully established. The impact of gynecologic history (parity, lactation, oral contraceptive use, age of menarche) on BMD is uncertain. PMID- 11109047 TI - Postmenopausal hormone replacement therapy and breast cancer. AB - The concern that postmenopausal hormone replacement therapy (HRT) may cause cancer of the breast has generated much research in epidemiology, endocrinology, and tumor cell biology. The recognition that naturally occurring 17beta-estradiol is a weak genotoxic and mutagenic carcinogen provides a plausible background for the association of breast cancer with HRT. However, because of the small anticipated effect and several confounding factors, the epidemiology of this association is complex. The consensus at this writing is that long-term HRT (>10 years) is associated with an increased risk of breast cancer, which, on average, is equivalent to the risk associated with delaying menopause for the same period of time. The particular risk depends on the duration and probably the dose to which the individual woman is exposed, as well as on a number of predisposing environmental and genetic factors. One clinical implication of the data reviewed here is that the dosage of HRT chosen should be the lowest that produces the desired effect. The use of HRT in women with a history of breast cancer is also addressed. Low-dose estrogen together with a selective estrogen receptor modulator to protect the breast may be a treatment option for women with severe symptoms of estrogen deficiency. PMID- 11109049 TI - Life satisfaction, symptoms, and the menopausal transition. AB - OBJECTIVE: The aims of this study were to examine the relation between life satisfaction and the menopausal transition, identify factors predictive or associated with life satisfaction, and determine the relation between life satisfaction and other health outcomes. RESEARCH DESIGN AND METHODS: This is a prospective population-based study of 438 middle-aged Australian-born women followed for 6 years after baseline measures. Retention rate at 6 years was 90% (n = 395). Two self-reported measures of life satisfaction (Life Satisfaction Index-Z scale [LSI-Z] and Satisfaction with Life Scale [SWLS]) were used in year 6. Positive and negative affect scales and questions about satisfaction with work and daily living were also used. Sociodemographic variables were measured at baseline, and attitudes toward menopause and aging were documented at years 2 and 5, respectively. Other explanatory variables, including symptoms, health, stress, life events, sexual functioning, and lifestyle were measured in year 6. RESULTS: Women overwhelmingly endorsed positive responses to life satisfaction questions. The LSI-Z and the SWLS were highly correlated with each other (r = 0.70), with the mood scales, and with responses to questions about satisfaction with work and daily living. The LSI-Z and SWLS were not related to menopausal status, hormone levels (follicle-stimulating hormone, estradiol), age, body mass index, hot flushes, hormone replacement therapy, sexual interest, employment status, type of profession, children at home, alcohol, chronic conditions, surgery, premenstrual complaints, life events (major or secondary), and social support. Stepwise multiple regression found that life satisfaction was predicted by earlier attitudes and was positively associated with feelings for partner and exercise and negatively associated with daily hassles, interpersonal stress, dysphoric symptoms, and current smoking. CONCLUSIONS: Life satisfaction was closely related to mood, predicted by earlier attitudes, and affected by relationship to partner, stress, and lifestyle. Life satisfaction was unrelated to menopause status, hormone levels, or hormone replacement therapy. PMID- 11109052 TI - The approval of mifepristone (RU486) in the United States: What's wrong with this picture? PMID- 11109054 TI - Prevalence of asthma symptoms in Latin America: the International Study of Asthma and Allergies in Childhood (ISAAC). AB - The prevalence of respiratory symptoms indicative of asthma in children from Latin America has been largely ignored. As part of the International Study of Asthma and Allergies in Childhood (ISAAC), 17 centers in 9 different Latin American countries participated in the study, and data from 52,549 written questionnaires (WQ) in children aged 13-14 years and from 36,264 WQ in 6-7 year olds are described here. In children aged 13-14 years, the prevalence of asthma ever ranged from 5.5-28%, and the prevalence of wheezing in the last 12 months from 6.6-27%. In children aged 6-7 years, the prevalence of asthma ever ranged from 4.1-26.9%, and the prevalence of wheezing in the last 12 months ranged from 8.6-32.1%. The lower prevalence in centers with higher levels of atmospheric pollution suggests that chronic inhalation of polluted air in children does not contribute to asthma. Furthermore, the high figures for asthma in a region with a high level of gastrointestinal parasite infestation, and a high burden of acute respiratory infections occurring early in life, suggest that these factors, considered as protective in other regions, do not have the same effect in this region. The present study indicates that the prevalence of asthma and related symptoms in Latin America is as high and variable as described in industrialized or developed regions of the world. PMID- 11109055 TI - Treatment of sleep-disordered breathing in children with myelomeningocele. AB - The prevalence of moderate to severe sleep-disordered breathing (SDB) in patients with myelomeningocele may be as high as 20%, but little information is available regarding treatment of these patients. To assess the efficacy and complications of treatments for these children, we collected data on 73 patients from seven pediatric sleep laboratories. Obstructive sleep apnea (OSA, n = 30) and central apnea (n = 25) occurred more frequently than central hypoventilation (n = 12). We also describe a sleep-exacerbated restrictive lung disease type of SDB in 6 patients who had hypoxemia during sleep without apnea or central hypoventilation. For each type of SDB, effective treatments were identified in a stepwise process, moving towards more complex and invasive therapies. For OSA, adenotonsillectomy was often ineffective (10/14), whereas nasal continuous positive airway pressure (CPAP) was usually successful (18/21). For central apnea, methylxanthines and/or supplemental oxygen proved sufficient in 2 of 9 and 3 of 6, respectively, but noninvasive positive pressure ventilation was required in 7 children. For central hypoventilation, supplemental oxygen (alone or with methylxanthines), noninvasive positive pressure ventilation, and tracheostomy with positive pressure ventilation were effective in 3, 2, and 2 patients, respectively. Sleep exacerbated restrictive lung disease always required supplemental oxygen treatment, but in 2 cases also required noninvasive positive pressure ventilation; nutritional and orthopedic procedures also were helpful. Posterior fossa decompression was used for the first three types of SDB, but data were insufficient to delineate specific recommendations for or against its use. In summary, evaluation by an experienced, multidisciplinary team can establish an effective treatment regime for a child with myelomeningocele and SDB. PMID- 11109056 TI - Quality of life in patients awaiting lung transplant: cystic fibrosis versus other end-stage lung diseases. AB - The symptoms associated with chronic lung disease can impair quality of life and psychosocial functioning. The purpose of the present study was to provide a thorough baseline assessment of quality of life in patients with end-stage lung disease and being evaluated for transplant; and to assess potential differences in quality of life between patients with cystic fibrosis (CF) and those with other types of end-stage lung disease (e.g., chronic obstructive pulmonary disease (COPD), interstitial pulmonary fibrosis (IPF)). We evaluated 58 patients with CF and 52 patients with other types of end-stage lung disease who were recruited for this study during an assessment of their candidacy for lung transplant. Subjects completed a battery of questionnaires that assessed demographic factors (including work and educational status), the presence of psychological distress (anxiety and depression), availability of social support, coping styles, and physical functioning. Despite significant impairment in physical functioning in the areas of recreation, household activities, sleep, and ambulation, other indices of life quality suggested good adaptation in the majority of patients. Also, quality of life differed for patients with CF and for those with other types of end-stage lung disease. Patients with CF were more likely to be working, had lower levels of anxiety and higher levels of social support, and used more functional coping strategies than did patients with other end-stage lung disease. These results highlight the fact that patients with different types of lung disease may require different psychosocial services as they await transplant. These findings also raise the question of whether there is a difference in quality of life after transplant between patients with CF and those with other types of lung disease. PMID- 11109057 TI - Self-hypnosis for patients with cystic fibrosis. AB - This report documents the utility of self-hypnosis in patients with cystic fibrosis (CF). Sixty-three patients 7 years of age or older were offered the opportunity to be taught self-hypnosis by their pulmonologist. Forty-nine agreed to learn it. Patients generally were taught hypnosis in one or two sessions. The outcome of hypnotherapy was determined by patients' answers to open-ended questions regarding their subjective evaluation of the efficacy of hypnosis. The average age of the 49 patients who were taught and used self-hypnosis was 18.1 years (range, 7-49 years). Many of the patients used hypnosis for more than one purpose, including relaxation (61% of patients), relief of pain associated with medical procedures (31%), headache relief (16%), changing the taste of medications to make the flavor more palatable (10%), and control of other symptoms associated with CF (18%). The patients successfully utilized self hypnosis 86% of the time. No symptoms worsened following hypnotherapy. Sixteen patients chose to practice hypnosis on their own for a half year or longer. In conclusion, with the use of self-hypnosis, patients with CF can quickly learn to enhance their control over discomforts associated with therapy and their disease. Consideration should be given to making instruction in self-hypnosis available to patients with CF. PMID- 11109058 TI - Clara-cell secretory protein in preterm infants' tracheal aspirates correlates with maturity and increases in infection. AB - Clara-cell secretory protein (CCSP), produced primarily by Clara cells in the conducting airways, is the most abundant soluble protein in pulmonary lavage fluid. CCSP is thought to be an immunosuppressive or anti-inflammatory protein with protective functions in the respiratory tract against exaggerated inflammatory reactions. CCSP was measured in 98 tracheoalveolar fluid (TAF) samples from 24 preterm infants (gestational age, 27.9 +/- 2.3 weeks, birth weight 1,020 +/- 305 g) with respiratory distress syndrome during the first 2 postnatal weeks. The ratio of urea-N in serum and in TAF was used to correct for dilution of TAF samples. Concentration of CCSP in TAF when corrected for dilution increased from 3.6 +/- 11 microg/mL on day 1 to 29.6 +/- 6.9 microg/mL on day 14. CCSP correlated with gestational age. A negative correlation was found between CCSP and inspiratory oxygen concentration, and a positive correlation between CCSP and both arterial pH and base excess during the first 2 postnatal weeks. Infants with clinical and laboratory signs of infection had higher CCSP than noninfected infants, and a negative correlation was found between CCSP and leukocyte count during the first 2 postnatal weeks (all P < 0.05). We suggest that pulmonary CCSP correlates with both gestational and postnatal age, and increases in response to infection in infants with respiratory distress during the early postnatal period. PMID- 11109059 TI - Accumulation of CO(2) in reservoir devices during simulated neonatal mechanical ventilation. AB - Aerosolized albuterol is frequently administered to mechanically ventilated neonates by metered dose inhaler (MDI) and a reservoir device. These reservoirs are often placed between the Y-piece and endotracheal tube, thereby creating mechanical dead space and increasing the risk of rebreathing carbon dioxide (CO(2)). The objectives of this study were: 1) to quantify CO(2) accumulation in two commonly used reservoirs (ACE(R), Aerochamber(R)-MV) and a bidirectional nonreservoir actuator (Airlife(R) Minispacer) during mechanical ventilation of a neonatal lung model; and 2) to determine the effect of tidal volume (V(T)) on CO(2) accumulation. We hypothesized that the accumulation of CO(2) in these devices is clinically insignificant at the small tidal volumes used in mechanically ventilated premature neonates. The model was constructed to simulate CO(2) exhalation by a ventilated neonate and consisted of a neonatal ventilator circuit (rate = 40/min; peak inspiratory pressure (PIP) = 20 cm H(2)0) attached to a reservoir/actuator and neonatal test lung. The ventilator delivered inspiratory gas (room air) to the test lung, which was vented into the atmosphere by a small adjustable leak. Expiration was simulated by manually ventilating 7.1% CO(2) (partial pressure of CO(2) (PCO(2)) = 48 mm Hg) back through the model. Accumulation of CO(2) within the reservoir/actuator was measured using an end tidal CO(2) monitor. Each 4-min experiment was conducted at three V(T) (7.5 mL, 15 mL, and 25 mL), and the median PCO(2) was calculated in 0.5-min increments. For V(T) = 7.5 mL, CO(2) accumulated slowly in the ACE(R) and Minispacer(R) and reached a maximum at 4.0 min (PCO(2) = 2.3 mm Hg and 7.3 mm Hg, respectively). In contrast, the Aerochamber(R)-MV rapidly reached a PCO(2) of 9.5-10.0 mm Hg by 1 1. 5 min. A similar trend occurred with V(T) = 15 mL; however, higher partial pressures (approximately 10-12 mm Hg) were achieved with all devices. At V(T) = 25 mL, PCO(2) rose rapidly with the ACE(R), Aerochamber(R)-MV, and Minispacer(R), reaching peaks of 17.2, 12.3, and 20.3 mm Hg, respectively (P < 0.05). In conclusion, accumulation of CO(2) in reservoir/actuator depends on V(T) as well as the chamber design and internal volume. Due to the short duration of use when administering drugs via MDI, accumulation of CO(2) in these devices is not likely to be clinically relevant for the majority of ventilated newborns. PMID- 11109060 TI - Sweat analysis proficiency testing for cystic fibrosis. AB - The purpose of this report is to describe the College of American Pathologists sweat testing (SW) proficiency testing program for cystic fibrosis, to evaluate its impact on test performance, and to describe the current practice of sweat testing in North America. The study analyzed participant summary reports of the SW survey from 1994-1998 (SW 94-98) and Proficiency Testing Exception Summary reports from 1996-1998. The data collected from SW 94-98 allowed for the assessment of trends and/or changes in sweat testing practices. The data collected from SW-A 1998 provided a profile of current practices in sweat testing. While the overall performance on the SW survey is encouraging, the program has identified areas of concern. The number of poorly performing laboratories are few in number, yet if the reported results had been patient specimens, the clinical implications would have been significant. The SW survey is meeting its goal of providing feedback to institutions on their performance of sweat analysis and providing educational materials on the total testing process in an effort to improve the quality of sweat testing. Significant changes in practice have occurred in many institutions performing sweat tests, and a greater awareness of analyte identification has resulted. PMID- 11109061 TI - Sinonasal disease in cystic fibrosis: clinical characteristics, diagnosis, and management. AB - Cystic fibrosis is an autosomal recessive genetic disorder that causes dysfunction of exocrine glands, and has several clinical manifestations. Among those, sinonasal involvement is almost universal, with or without chronic sinusitis and/or nasal polyposis. This review will detail the pathophysiologic changes of the sinonasal mucosa, and the clinical manifestations, diagnosis, and treatment. Developmental anatomic abnormalities, which are identified radiologically, will also be demonstrated. Medical management is the first treatment for patients with cystic fibrosis, but effective treatment of sinonasal disease in cystic fibrosis relies heavily on surgery. In the past, nasal polyposis was the main indication for surgery, and consisted mostly of polypectomy alone. This procedure was associated with a high recurrence rate. The development of functional endoscopic sinus surgery has contributed to decreasing the morbidity of sinonasal surgery and the recurrence of nasal polyposis in cystic fibrosis. The evolution of the surgical techniques will be discussed and a review of the literature will be provided. PMID- 11109062 TI - Chiari type I malformation in children and adolescents with cystic fibrosis. AB - Chiari type I malformation is characterized by herniation of the cerebellar tonsils through the foramen magnum. An association between Chiari type I malformation and cystic fibrosis (CF) has not previously been established. We report on five children and adolescents with CF in whom Chiari type I malformations were diagnosed. Three patients were 17-18 years old at time of diagnosis, one was 3 years old, and one was 10 months of age. All patients were followed at the Cystic Fibrosis Center at St. Christopher's Hospital for Children and were diagnosed with the malformations between June 1988 and June 1997. Over this same period, 400 CF patients 18 years or younger were followed routinely. All patients had the diagnosis of Chiari type I confirmed by brain-stem MRI. Neurologic findings included swallowing dysfunction, syncopal episodes, numbness of extremities, recurrent vomiting, and headaches. No two patients had the same presenting neurologic findings. Our data suggest that Chiari type I malformation is more common in CF than in the general population. The possibility of Chiari type I malformation should be included in the differential diagnosis of unexplained neurologic complaints in patients with CF. PMID- 11109063 TI - Life-threatening hemothorax in a child following intrapleural administration of urokinase. PMID- 11109064 TI - Nineteenth annual conference on sleep disorders in infancy and childhood annenberg center for health sciences rancho mirage, california, january 18-20, 2001 PMID- 11109065 TI - Salzburg Spectacle. Russia Joins. PMID- 11109066 TI - Malpractice Insurance. PMID- 11109067 TI - Meetings, Journal and Societies. PMID- 11109068 TI - Diabetes and bariatric surgery. PMID- 11109069 TI - Obesity Surgery and Laparoscopic Technique. PMID- 11109070 TI - Obituary: Daniel A.K. Roncari, MD. PMID- 11109072 TI - Obesity Surgery Including Laparoscopy and Allied Care. PMID- 11109071 TI - The NIH Consensus Development Conference Revisited. PMID- 11109073 TI - Allopathic versus 'holistic' medicine. PMID- 11109075 TI - Recent developments in the management of testicular germ cell tumors. AB - Testicular germ cell tumors are now generally regarded as a highly curable cancer in the majority of cases, even when patients present with advance disease. However that does not imply that researchers and clinicians have become complacent in their efforts to improve our understanding and the treatment outcome of this disease. This is an overview of recent developments in the management of testis cancer. PMID- 11109076 TI - Prostate needle biopsy infection after four or six dose ciprofloxacin. AB - OBJECTIVE: This retrospective analysis is to determine rates of clinical infection after prostate needle biopsy with four versus six doses of ciprofloxacin to previous literature. MATERIALS AND METHODS: Two groups were treated with pre and post biopsy 500 mg of ciprofloxacin twice daily by either six doses (n=337) or four doses (n=288) with the first dose given 24 or 12 hours prior to the procedure respectively. RESULTS: Six (0.96%) of the 625 patients had symptomatic urinary tract infections with a positive urinalysis and/or culture. One (0.3%) infection occurred among patients receiving six doses of ciprofloxacin, and five infections (1.7%), were identified among four dose patients. Two febrile episodes occurred in the four dose group, one requiring hospitalization. CONCLUSION: A low infection rate associated with prophylactic regimens. Six doses of ciprofloxacin appears more effective than four doses in reducing the clinical and febrile infection rate following ultrasound guided biopsy of the prostate. No obvious financial benefit was observed. PMID- 11109077 TI - Reliability analysis of first and second generation PSA assays. AB - PURPOSE: To assess the reliability of first and second generation PSA assays. MATERIALS AND METHODS: In the present investigation we sought to compare pretreatment serum PSA levels determined by a first (IMx) and a second (IMMULITE) generation PSA assays to determine whether there were differences. Sera from 545 men were investigated in the range > 0 - 5330 microg/L, and prostatic histology was known, based on either transrectal ultrasound (TRUS), guided systematic needle biopsies, or transurethral resection or prostatectomy. RESULTS: Over the entire range there was an excellent correlation (r > 0.97) between the IMx and the IMMULITE PSA assays. When analyzed according to histology, there was an equivalent slope in the PSA ranges for patients with benign prostatic hyperplasia compared with prostate cancer patients. The area under the ROC curve for the IMx for the total PSA range was 0.7860, and for the IMMULITE assay the area under the ROC curve was 0.7810, a striking resemblance and not different significantly (p=0.87). CONCLUSION: For the majority of men, the first (IMx) and second (IMMULITE) generation PSA assays are equivalent. Small differences between both assays will not be of clinical significance for most men, but should be considered when comparing results of different assays in sequential determinations for a specific man. PMID- 11109078 TI - Vitamin B12 deficiency and incontinence in older people. AB - OBJECTIVES: To investigate the relationship between urinary incontinence and Vitamin B12 deficiency in community-living older people using standard serum cobalamin levels as well as the metabolites methylmalonic acid (MMA) and total homocysteine (HCYS). DESIGN: A prospective cross-sectional study of community living older people. PARTICIPANTS: Independent and cognitively normal adults (mean age=72 years, range=65-89 years) living in the community. One hundred and nineteen volunteers were recruited at group meetings, activity groups, and through the use of posters. MEASUREMENTS: Information on urinary continence was provided by the participant during a systematic medical history, and was defined as any amount of uncontrolled leakage of urine. Vitamin B12, methylmalonic acid and homocysteine levels were determined in all subjects. MAIN RESULTS: We found no significant differences between the continent and incontinent group in regard to their serum Vitamin B12 (OR=1.34, CI: 0.39-4.58, p=.424), methylmalonic acid levels (OR=0.71, CI: 0.24-2.10, p=.386), or total homocysteine levels (CI: 0.29 4.54, p=.535). CONCLUSIONS: In our study continence was not significantly affected by the B12 status of the subject; neither serum B12 levels or the metabolites MMA and HCYS (reflecting B12 function) were significantly different in the continent group versus the incontinent one. PMID- 11109079 TI - A para-testicular primitive neuroectodermal tumor in an adult: a case report and literature review. AB - OBJECTIVE: The authors describe the salient clinical, radiologic and histopathologic features of an extremely rare para-testicular primitive neuroectodermal tumor in a 25 year-old man. INTERVENTION: Excisional biopsy of the tumor en bloc was performed. Adjuvant VAdriaC-based chemotherapy (Vincristine, Doxorubicin, and Cyclophosphamide) was given post-operatively. MAIN OUTCOME MEASURES: Histopathologic examination and immunohistochemical studies were performed on formaldehyde-fixed, paraffin-embedded tumor tissue. RESULTS: Histologic examination showed an undifferentiated small cell tumor. The tumor cells stained positively with MIC-2, a marker specific for primitive neuroectodermal tumors. The patient is 12 months post surgery and has completed adjuvant chemotherapy with no evidence of recurrent disease. CONCLUSIONS: This highly unusual, peripheral primitive neuroectodermal tumor should be considered in the differential diagnosis of undifferentiated small cell neoplasms of the genitourinary system in adults, from the kidney to the testicle. We present a patient with a PNET treated based on a Ewing's family of tumors protocol. PMID- 11109080 TI - Palliative chemoirradiation for transitional carcinoma of the bladder: a case report. AB - Palliation of locally advanced transitional carcinoma of the bladder is a major problem that has not been well investigated. Recent reports from the curative literature suggest improved tumor response rates using platinum based chemoirradiation over radiotherapy alone. Because of the poor symptomatic responses seen with conventional palliative radiotherapy, we have been treating selected cases of locally advanced bladder carcinoma palliatively with low dose chemoirradiation. We present a case of an elderly female treated this way with an excellent clinical response. PMID- 11109081 TI - Urinary tract infection: from basic science to clinical application. PMID- 11109082 TI - Innate defences and resistance to gram negative mucosal infection. PMID- 11109083 TI - Pathogenicity islands of uropathogenic E. coli and evolution of virulence. PMID- 11109084 TI - Iron transport in Escherichia coli. Crystal structure of FhuA, an outer membrane iron and antibiotic transporter. PMID- 11109085 TI - The cytotoxic necrotizing factor 1 (CNF1) from uropathogenic Escherichia coli. PMID- 11109086 TI - Genetic characterization of the uropathogenic E. coli strain 536--a subtractive hybridization analysis. PMID- 11109087 TI - Analysis of the hemolysin determinants of the uropathogenic Escherichia coli strain 536. PMID- 11109088 TI - Functional variability of type 1 fimbriae of Escherichia coli. PMID- 11109089 TI - HPI of high-virulent Yersinia is found in E. coli strains causing urinary tract infection. Structural, functional aspects, and distribution. PMID- 11109090 TI - Aggregation substance of Enterococcus faecalis: a multifunctional adhesin. PMID- 11109091 TI - A role for the sigma S subunit of RNA polymerase in the regulation of bacterial virulence. PMID- 11109092 TI - Transcriptional organisation and regulation of E. coli group 2 capsule expression. PMID- 11109093 TI - Molecular basis of catheter associated infections by staphylococci. PMID- 11109094 TI - Control mechanisms in the Pap-pili system. PMID- 11109095 TI - Transcriptional analysis of the sfa and pap determinants of uropathogenic Escherichia coli strains. PMID- 11109096 TI - Structural and functional studies of the fimbrial adhesin gene regulator PapB from uropathogenic Escherichia coli. PMID- 11109097 TI - Interaction of the nucleoid-associated proteins Hha and H-NS to modulate expression of the hemolysin operon in Escherichia coli. PMID- 11109098 TI - Use of the OmpS-display--system to localize the receptor-binding region in the PapG adhesin of uropathogenic Escherichia coli. PMID- 11109099 TI - The role of the AirS two-component system in uropathogenic Escherichia coli. PMID- 11109100 TI - Examination of regulatory cross-talk between the decay accelerating factor binding fimbrial/afimbrial adhesins and type I fimbriae. AB - The revised sequence of DaaA means that the protein is 98% identical to the afimbrial adhesin regulator AFAA-III. While PapB repressed FimB OFF to ON switching in strain AAEC370A, this was not the case for DaaA. PapB, but not DaaA, reduced the level of expression of type 1 fimbriae, in static LB media. Attention is now focused on the amino terminus of the PapB protein as the possible domain involved in cross talk with type 1 fimbriae. PMID- 11109101 TI - Modulation of the polysaccharide intercellular adhesin (PIA) expression in biofilm forming Staphylococcus epidermidis. Analysis of genetic mechanisms. PMID- 11109102 TI - Induction of Staphylococcus epidermidis biofilm formation by environmental factors: the possible involvement of the alternative transcription factor sigB. PMID- 11109103 TI - Expression of virulence genes in Candida albicans. PMID- 11109104 TI - Effect of spontaneous and induced mutations on outer membrane proteins and lipopolysaccharides of Proteus penneri strain 357. PMID- 11109105 TI - Role of bacterial lectins in urinary tract infections. Molecular mechanisms for diversification of bacterial surface lectins. PMID- 11109106 TI - Adherence of enteric bacteria onto the mammalian extracellular matrix. Test-tube artefact or a virulence function? PMID- 11109107 TI - Interrelationship between virulence properties of uropathogenic E. coli and blood group phenotype of patients with chronic urinary tract infection. PMID- 11109108 TI - Glycolipid receptors of F1C fimbrial adhesin of uropathogenic Escherichia coli. PMID- 11109109 TI - Thin aggregative fimbriae on urinary Escherichia coli isolates. PMID- 11109110 TI - Type 1 pili of Citrobacter freundii mediate invasion into host cells. PMID- 11109111 TI - Serotypes, siderophore synthesis, and serum resistance of uropathogenic Klebsiella isolates. PMID- 11109112 TI - Epitope specificity of polyclonal rabbit antisera against Proteus vulgaris O antigens. PMID- 11109113 TI - Virulence factors of Escherichia coli isolated from urine of diabetic women with asymptomatic bacteriuria. PMID- 11109115 TI - Characterisation and adherence mechanisms of Escherichia coli strains causing infections in patients with a reconstructed bladder. PMID- 11109114 TI - Cytokine secretion is impaired in women with diabetes mellitus. PMID- 11109116 TI - Immunodominant proteins in human sepsis caused by methicillin resistant Staphylococcus aureus. PMID- 11109117 TI - Modern concept of antibiotic therapy of urinary tract infections. PMID- 11109118 TI - The role of anaerobic bacteria in prostatitis. PMID- 11109119 TI - Urinary tract infection in dogs. Analysis of 419 urocultures carried out in Portugal. PMID- 11109120 TI - Detection of virulence factors in uropathogenic Escherichia coli isolated from humans, dogs and cats in Portugal. PMID- 11109121 TI - Asymptomatic bacteriuria can be considered a diabetic complication in women with diabetes mellitus. PMID- 11109122 TI - Determination of genetic diversity of Proteus penneri strains using rep-PCR. PMID- 11109123 TI - Use of randomly amplified polymorphic DNA (RAPD) analysis for identification of Proteus penneri. PMID- 11109124 TI - Overview on the clinical studies with Urostim immunostimulator against urogenital infections. PMID- 11109125 TI - Porcine postweaning diarrhea isolates of Escherichia coli with uropathogenic characters. PMID- 11109127 TI - New serogroups of the genus Proteus consisting of Proteus penneri strains only. Determination of some LPS epitopes responsible for specificity. PMID- 11109126 TI - Virulence markers of human uropathogenic Escherichia coli strains isolated in Hungary. PMID- 11109128 TI - The role of Chlamydia trachomatis in asymptomatic and symptomatic urogenital infections. PMID- 11109129 TI - Psychotherapy in the twenty-first century. PMID- 11109130 TI - Thoughts about "On the millennium". PMID- 11109131 TI - Making a future. PMID- 11109132 TI - Introduction: psychotherapy with terminally ill patients. PMID- 11109133 TI - Cancer and the experience of meaning: a group psychotherapy program for people with cancer. AB - Cancer illness affects people in many ways, physical, financial, and existential. In this paper, we describe a proposed group intervention for individuals with advanced disease who want help finding a sense of meaning at this critical juncture in their lives. This intervention has a brief, semi-structured format, and is informed by the work of Viktor Frankl, empirical findings in the area of meaning and trauma, and the empirical findings of other group interventions for cancer patients. Individual sessions focus on different aspects of meaning, including responsibility to others, creativity, transcendence, and ascertaining one's values and priorities. Having goals on which to focus and feeling like part of a larger whole are critically important to the ability to find meaning and cope with terminal illness. Such goals may be generated by a number of sources, including connectedness with others, or a sense of the temporal continuity of one's own life despite the disruption posed by severe illness. Didactic discussions and experiential exercises help to facilitate exploration of these various elements in group members' lives. The finite structure of the intervention may also highlight these issues, as people who are faced with similar issues work together in a limited time frame in order to accomplish the goals they set out for themselves. PMID- 11109134 TI - The house that's on fire: meaning-centered psychotherapy pilot group for cancer patients. AB - People with advanced cancer face an existential crisis in addition to their physical suffering. The principles of a new group therapy intervention (MCGP) were introduced in another paper in this issue. This paper is a report of some of the themes and issues that arose during the first pilot group. PMID- 11109135 TI - An unfortunate family: terminal illness and the altering of the attachment bond. AB - This article emphasizes the value of a therapeutic presence in terminal death situations. An unusual clinical case illustrates the point. The presenting issue was aggression between an adult son and father. It soon became apparent, however, that this son was profoundly enmeshed with his mother. During the course of family therapy the father died, and then the mother was diagnosed with cancer. The therapist changed therapeutic goals and attended to the relationship between mother and son. He made home visits to the mother and the surviving son. She seemed to die peacefully and the son moved on with his life, which prior to the death of his mother would not have appeared likely because of his profound dependence on her. Resolving pathological attachment is not easy. Attachment theory refers to the affectional bonds or attachments, initially between child and parent and later between adult and adult. It is suggested that terminal illness and death can, with help of a therapist, alter this attachment bond, even though the literature on adult dependency and attachment does not refer to death in this manner. PMID- 11109136 TI - The whisper of death: psychotherapy with a dying Vietnam veteran. AB - Psychotherapy with a dying Vietnam veteran is described. In spite of a severe heart condition and HIV-positive diagnosis, the outspoken and provocative patient reverts to heroin and cocaine use early in the treatment. This causes a heart attack and interruption of treatment. The therapist maintains empathy, a solid bond is forged, and the patient returns, but under constrained circumstances. A turning point is reached, both in the treatment and in this final phase of the patient's life, with major life improvement ensuing. At death the patient leaves word to thank the therapist. This case exemplifies how the approach of death lends urgency to positive forces appropriate to life's final developmental stage, and how end-of-life therapy bolsters those forces. PMID- 11109137 TI - Zazen and psychotherapeutic presence. AB - Zen meditation, or zazen, has attracted the interest of many psychotherapists. The teachings and practices of the Soto Zen tradition are understood as encouraging important areas of both psychological and spiritual development. Zen, like the relational psychoanalytic theories, encourages its practitioners to become aware of the fundamentally distorted aspects of an overly individualistic view of human experience. As a spiritual practice, zazen increases the practitioner's tolerance and appreciation of the Wholeness that Buddhists refer to as Emptiness. As a psychological practice, it helps us to be more flexibly and intimately present with our patients. An effective therapeutic process, even of the most secular type, will often contain elements of the meditative process of zazen, and failure to actualize this in psychotherapy can have a negative impact on our ability to understand and help our patients. PMID- 11109138 TI - Clouds and silver linings: training experiences of psychodynamically oriented mental health trainees. AB - This paper discusses the experiences of today's psychodynamically oriented mental health trainees. Recent changes in the training environment, such as the increase in managed care, rise in use of psychotropic medication, the waning popularity of psychodynamic thinking, and reduced funding for psychotherapy training, in general, have all affected current trainees' professional development. In particular, trainees struggle with problems of demoralization, professional isolation, and reduced financial opportunities. Advantages that current trainees experience, as well as suggestions for training directors and trainees, will also be discussed. PMID- 11109139 TI - Healing madness and despair through meeting. AB - An existential perspective on human suffering suggests that the psychoanalytic view of the unconscious poses a difficult dilemma in regard to self-examination: On the one hand, not to pursue introspection may leave one as a person driven by untoward instinctual urges; on the other hand, to vigorously self-reflect, may result in inexorable despair in regard to the limitations and finitude of mortal existence. The author contends that the reason for the dilemma is rooted in Western psychology--a perspective that regards the human being as an encapsulated consciousness, set separately and competitively apart from other objects in the cosmos. This orientation is shown to pose difficulties in treating madness and despair. By the use of clinical material, the author shows how Martin Buber's notion of authentic dialogue can be useful in efforts to bridge the separation between the sufferer and the healer. PMID- 11109140 TI - Psychotherapy with substance abusers: integration of psychodynamic and cognitive behavioral approaches. PMID- 11109141 TI - Are the margins clear? PMID- 11109142 TI - What's eating you? Amblyomma ticks (Amblyomma americanum). PMID- 11109144 TI - Photo quiz. Staphylococcal scalded skin syndrome. PMID- 11109143 TI - Lymphocytoma cutis: cases linked with Lyme disease. PMID- 11109145 TI - Air bag injury and the dermatologist. AB - Most new car models have driver-side air bags and many also have passenger-side and side-impact air bags. Air bags are known to be dangerous to small children and may cause death, fractures, and cerebral spinal injury. However, the cutaneous manifestations of air bag injury are less well known. Additional potential air bag injuries include retinal damage and high-frequency hearing loss. The following case report illustrates significant burns from a low-impact air bag injury and reviews the pertinent literature. PMID- 11109146 TI - Double lip: an unusual presentation. AB - Double lip is a term used to describe a deformity of the upper or lower lip and consists of an accessory fold of redundant mucous membrane inside the vermilion border. Double lip is an uncommon congenital anomaly and is usually diagnosed and treated by a dentist or an oral surgeon. We present an unusual case of double lip -unusual because it was diagnosed and treated by a dermatologist and because of the histopathologic findings. PMID- 11109147 TI - Multiple syringomas on the abdomen, thighs, and groin. AB - Syringomas are benign adnexal tumors that occur most commonly in women. They typically present as soft, flesh-colored to slightly yellow papules on the lower eyelids. We present an unusual case of a healthy 33-year-old male with multiple, reddish brown syringomas located on the lower abdomen, thighs, and groin. Although these lesions can result in significant cosmetic disfigurement, treatment options are limited and generally disappointing. PMID- 11109148 TI - A case of a hard inguinal nodule. AB - Dermatofibrosarcoma protuberans (DFSP) is an uncommon tumor with its onset typically in the second to fifth decades of life. It most commonly presents on the trunk, and recent cytogenetic studies suggest a neural origin. A case presentation and review of the recent literature on the diagnosis, differential diagnosis, and treatment of DFSP is presented. PMID- 11109149 TI - Gangrene of the fingertips after bleomycin and methotrexate. AB - The increased use of cytostatic drugs, which are sometimes used in combination chemotherapy, may result in new and unusual cutaneous side effects. We describe a 57-year-old man with acral erythrocyanosis progressing to acute digital ischemia and gangrene that developed after combined chemotherapy (bleomycin and methotrexate) used to treat a metastatic squamous cell carcinoma of the hypopharynx. A leukocytoclastic vasculitis was found in both the acute phase and in the amputated fingertips. This supports the well-reported potential of bleomycin to trigger acral vascular toxicity. PMID- 11109150 TI - Tuberculous gumma (cutaneous metastatic tuberculous abscess) with underlying lymphoma. AB - Cutaneous tuberculosis is an infrequent first sign of disseminated tuberculosis. We describe a patient with 2 cutaneous ulcerations that grew Mycobacterium tuberculosis. Despite an initial response to antimycobacterial therapy, the fever relapsed. After several months, biopsy of a single cervical lymph node showed a T cell-rich B cell lymphoma. Our patient had metastatic tuberculous abscesses (tuberculous gummas), which are secondary to disseminated tuberculosis, and an underlying occult lymphoma, both believed to be sequentially presenting as a fever of unknown origin. PMID- 11109151 TI - Treatment of acne with oral contraceptives: criteria for pill selection. AB - Combination oral contraceptives (OCs) (those that contain estrogen and progestin) are widely used in the treatment of acne because they modify an excessively androgenic hormonal environment and can decrease lesions. Dermatologists' knowledge of the most appropriate OC may be hampered by an incomplete understanding of these agents, misleading promotion, and confusion surrounding the new generation of OCs. Despite reports attributing significance to the degree of androgenicity of the progestin components of OCs, in vitro and animal bioassays of androgenicity have little clinical relevance. Because all of today's low-dose combination OCs are estrogen dominant, they are equally beneficial in women with androgenic conditions such as acne. Use of the OC containing the lowest dose of each hormone, consistent with the patient's needs, can enhance compliance by preventing or limiting common early-cycle side effects (e.g., nausea/vomiting, breast tenderness, weight gain, headache), while providing acne improvement. PMID- 11109152 TI - Purpuric irritant contact dermatitis induced by Agave americana. AB - The sap of Agave americana, a popular ornamental plant, may cause irritant contact dermatitis. This rare eruption is typically vesiculopapular; however, a new purpuric variant with evidence of leukocytoclastic vasculitis has recently been reported. We report an additional case of a purpuric eruption associated with severe constitutional symptoms further supporting a possible vasculitic component. Both cases resulted from direct exposure to sap propelled by a chainsaw. We speculate that oxalic acid crystals, which are recognized systemic toxins, are embedded in the skin with resulting oxalism, which may result in vascular damage. PMID- 11109153 TI - Response of confluent and reticulate papillomatosis of Gougerot and Carteaud to topical tretinoin. AB - Confluent and reticulate papillomatosis (CRP) of Gougerot and Carteaud is a rare cutaneous disorder characterized by persistent, usually asymptomatic, dark papules and plaques centrally located on the back, intermammary, and epigastric areas. The eruption spreads out peripherally into a fading reticulated pattern. The pathogenesis is poorly understood, but there are several theories. Many different treatments, with varying success rates, have been attempted. We present 3 patients with CRP who had excellent results in the areas treated with topical tretinoin. The only difficulty with therapy is applying the tretinoin to the back, which sometimes necessitates a second person. However, if this situation can be overcome, topical tretinoin provides an effective, safe alternative to systemic therapies. Response to tretinoin provides support that CRP is a disorder of keratinization. Finally, the fact that 2 of the patients were brothers may support the idea that CRP has a hereditary influence. PMID- 11109154 TI - Of pills and pillows: pseudopigmentation in a patient taking amiodarone. AB - Blue discoloration of the skin can alarm patients and physicians alike. Blue coloring may, however, have a trivial and easily correctable cause, as is shown in the following case. PMID- 11109155 TI - An unusual case. PMID- 11109156 TI - The effect of antibacterial soap with 1.5% triclocarban on Staphylococcus aureus in patients with atopic dermatitis. AB - This double-blind study determined whether daily bathing with an antibacterial soap would reduce the number of Staphylococcus aureus on the skin and result in clinical improvement of atopic dermatitis. For 9 weeks, 50 patients with moderately severe atopic dermatitis bathed daily with either an antimicrobial soap containing 1.5% triclocarban or the placebo soap. They also used a nonmedicated moisturizer and 0.025% triamcinolone acetonide cream as needed, but the availability of the corticosteroid cream was discontinued after 6 weeks. The antimicrobial soap regimen caused significantly greater improvement in the severity and extent of skin lesions than the placebo soap regimen, which correlated with reductions both in S aureus in patients with positive cultures at baseline and in total aerobic organisms. Outcome measures included reductions in S aureus, total aerobic organisms, and dermatologic assessments. Overall, daily bathing with an antibacterial soap was well tolerated, provided clinical improvement, and reduced levels of skin microorganisms. PMID- 11109157 TI - Histopathology of genetically defined endometrial precancers. AB - Endometrial precancers are monoclonal, benign neoplasms prone to malignant transformation. A type collection (deposited at www.endometrium.org) of confirmed precancers has been identified by their monoclonal growth and continuity of acquired genetic markers that occur between premalignant and malignant phases of tumorigenesis. Computerized morphometry of these premalignant lesions, designated endometrial intraepithelial neoplasia (EIN), has disclosed new architectural criteria and revised cytologic criteria for their diagnosis. EIN lesions originate focally and expand in size over time, in keeping with a proliferative monoclonal origin. They are characterized by closely packed glands (volume percentage stroma < 55%) with cytology that is clearly demarcated from that of the adjacent field. A minimum homogeneous field of cytologically demarcated glands is required to accurately assess the architecture diagnostic of EIN, and morphometry-diagnosed lesions with a largest diameter of at least 1 to 2 mm have previously been shown to predict the relevant clinical outcome of concurrent or future endometrial adenocarcinoma. Nonphysiologic loss of the PTEN protein, a tumor suppressor gene mutated in many endometrioid adenocarcinomas, is seen in individual glands of endometrium exposed to unopposed estrogens and in packed clusters of EIN glands. The isolated PTEN-free glands in anovulatory endometria may be the earliest stage of endometrial tumorigenesis, but they are not readily distinguishable by routine histology, nor do they have a defined natural history. PMID- 11109158 TI - The molecular biology of endometrial tumorigenesis: does it have a message? PMID- 11109159 TI - Endometrial hyperplasia: is it time for a quantum leap to a new classification? PMID- 11109160 TI - Endocervicosis involving the uterine cervix: a report of four cases of a benign process that may be confused with deeply invasive endocervical adenocarcinoma. AB - Four cases of endocervicosis that involved the outer cervical wall and paracervical connective tissue are reported; in one case there was also transmural involvement of the urinary bladder. A diagnosis of cervical adenocarcinoma was an initial concern of the referring pathologist in three cases. The patients were from 29 to 45 years of age; there was a history of cesarean section in two cases. Two patients presented with pelvic pain, one with dysmenorrhea, and one with symptoms related to an ovarian tumor. In three cases, a gross abnormality of the outer aspect of the cervix was noted at the time of hysterectomy and in the fourth at the time of macroscopic pathologic examination. The anterior wall of the cervix in each case was involved by a firm rubbery mass, 1 to 2.5 cm in maximal dimension, with cysts seen on sectioning in two. Microscopic examination disclosed a dominant population of glands of variable size and shape, including cystically dilated glands, lined by mucinous endocervical-type epithelium that ranged from columnar to flattened. All the glands had lining cells with bland cytologic features with absent to rare mitotic figures. A periglandular stromal reaction, present in two cases, was related to mucin extravasation. A cuff of endometriotic stroma was present around rare glands in one case. The appearance of the lesion was similar to that of endocervicosis of the urinary bladder, and as in that site, raised concern for adenocarcinoma, specifically for the minimal deviation (adenoma malignum) type of cervical adenocarcinoma. Awareness of the distinctive features of endocervicosis in this site, including its dominant location in the outer portion of the cervix and paracervical connective tissue and the typical presence of an uninvolved zone of cervical wall between the endocervicosis and the eutopic endocervical glands, facilitate the correct diagnosis. PMID- 11109161 TI - Cyclin D1 expression in high-grade endometrial carcinomas--association with histologic subtype. AB - Endometrial endometrioid adenocarcinoma (EC) and serous carcinoma (ESC) are associated with different epidemiologic risk factors, precursor lesions, morphology, and survival outcomes. They also possess distinct molecular profiles. We investigated the expression of cyclin D1, a member of the G1 cyclin family that regulates the G1/S transition in the cell cycle, and estrogen and progesterone receptors (ERs and PRs, respectively) in a group of ECs and ESCs matched for histological grade. We also sought to correlate the expression of cyclin D1 with ER and PR because cyclin D1 has been reported to stimulate transcription of ER- and PR-regulated genes (1,2). We hypothesize that cyclin D1 expression covaries with histologic subtype and is related to the expression of ER and PR. Twenty ESCs and 21 ECs were examined histologically and evaluated immunohistochemically for cyclin D1, ER, and PR using commercially available monoclonal antibodies in archival, formalin-fixed, and paraffin-embedded tissue. Three ESCs (15%) and 10 ECs (48%) expressed cyclin D1 (p = 0.02). Twelve ESCs (60%) and 16 ECs (76%) expressed ER, which is not significantly different. ER positive ECs were significantly more likely to express cyclin D1 compared with ER positive ESCs (p = 0.03), but a relationship between cyclin D1 and ER expression in EC was not found. We also did not find a significant relationship between cyclin D1 and PR expression. Therefore, cyclin D1 expression in poorly differentiated endometrial carcinomas is associated with endometrioid histology. This is consistent with pathobiologic divergence in poorly differentiated endometrial carcinomas. PMID- 11109162 TI - Analysis of estrogen receptor alpha and beta in endometrial carcinomas: correlation with ER beta and clinicopathologic findings in 45 cases. AB - Estrogens play important roles in the pathogenesis of the great majority of endometrial endometrioid adenocarcinoma. Recently, a novel estrogen receptor (ER), ER beta, has been characterized, but little is known about the status of ER beta in endometrial carcinoma. We therefore examined expression of both ER alpha and ER beta in 45 cases of endometrioid endometrial adenocarcinoma using mRNA in situ hybridization, reverse transcription and polymerase chain reaction (RT-PCR), and immunohistochemistry. We also correlated the findings with various clinicopathologic parameters in these cases to examine their possible biologic significance. Accumulation of mRNA hybridization signals for both ER alpha and ER beta was detected predominantly in the cytoplasm of carcinoma cells, and to a lesser extent in some stromal cells. ER beta mRNA was detected in 16/45 cases (35.6%), and ER alpha mRNA hybridization signals were detected in 36/45 cases (80.0%). Among the 16 ER beta positive cases, 15 cases also had ER alpha mRNA hybridization signals. In the cases that expressed both ER alpha and ER beta, ER alpha mRNA hybridization signals were more widely distributed than ER beta mRNA. In 21 cases, carcinoma cells had ER alpha mRNA hybridization signals but not ER beta mRNA. There was a statistically significant positive correlation between the results of mRNA in situ hybridization and semiquantitative RT-PCR or immunohistochemistry for both ER alpha and ER beta. There were no significant correlations between ER beta mRNA expression and PR labeling index, Ki67 LI, age, or histologic grade. The results from our study indicate that ER beta is coexpressed with ER alpha, and that the estrogenic effects occur predominantly through ER alpha in endometrial carcinomas. PMID- 11109163 TI - Pathologic features of uterine leiomyomas following uterine artery embolization. AB - Bilateral uterine artery embolization has recently been employed as an alternative to operational treatment of uterine leiomyomas. The pathologic features induced by uterine artery embolization have not been previously described in detail. Usually patients experience symptomatic improvement with a reduction in size of the leiomyomas. This report describes the pathologic features in a series of 10 uterine leiomyomas where tissue was available for histologic examination following uterine artery embolization. Characteristic histologic features within the leiomyomas included massive necrosis, sometimes with dystrophic calcification, vascular thrombosis, and intravascular foreign material that elicited a histiocytic and foreign-body giant cell reaction. In some cases, intravascular foreign material was present elsewhere in the myometrium, the cervix, or paraovarian region. In occasional cases, there were foci of myometrial necrosis and microabscess formation beyond the confines of the leiomyomas. Foci of extrauterine inflammation were also occasionally identified. Histopathologists should be aware of these findings because the use of uterine artery embolization will possibly become more widespread in the future. PMID- 11109164 TI - Grading of ovarian carcinomas. AB - Grading of ovarian carcinomas can have important implications for therapeutic decisions, in particular in International Federation of Gynecology and Obstetrics (FIGO) stage I. However, there are no universally accepted grading guidelines for this type of cancer. We applied the grading system suggested by Shimizu et al. (1) to a series of ovarian carcinomas from a single institution to evaluate its prognostic significance in relation to other predictive factors. One hundred ninety-two cases of ovarian carcinomas were studied, including all major histologic types. The histologic slides were evaluated with regard to architecture (glandular = 1 point, papillary = 2 points, solid = 3 points), nuclear pleomorphism (nuclear variability < or = 2:1 = 1 point, intermediate nuclei = 2 points, nuclear variability > or = 4:1 = 3 points), and mitotic activity per 10 high-power fields using objective 40x, ocular 10x/20 (0-7 mitoses = 1 point, 8-18 mitoses = 2 points, > or = 9 mitoses = 3 points). Carcinomas with a total score of 3-5 points were designated as grade I, 6-7 points were designated as grade II, and 8-9 points were designated as grade III. Kaplan-Meier curves showed the following 5-year survival rates: grade I (n = 42) 88%, grade II (n = 98) 60%, grade III (n = 52) 38% (p < 0.0001); stage I 90%, stage II 60%, stage III 38%, stage IV 10% (p < 0.0001); residual disease < or = 2 cm 67% and > or = 2 cm 27% (p < 0.0001). Multivariate Cox analysis revealed that grade (p < 0.0002), as well as nuclear pleomorphism alone (p < 0.0001), both provided statistically significant independent prognostic information. Our observations showed that the grading system used can be easily applied to all histologic types of ovarian carcinomas yielding prognostically relevant information and can be incorporated into routine diagnostic practice. PMID- 11109165 TI - Tumor angiogenesis and thymidine phosphorylase expression in ovarian carcinomas including serous surface papillary adenocarcinoma of the peritoneum. AB - To clarify biological and clinical significance of tumor angiogenesis in the development of ovarian carcinoma, we investigated the relationship between tumor vascularity, the expression of thymidine phosphorylase (dThdPase), which is an angiogenic factor identical to platelet-derived endothelial cell growth factor (PD-ECGF), and patient outcome in ovarian carcinoma, including serous surface papillary carcinoma (SSPC). Primary tumor specimens (stages I-IV) from 54 patients were examined. Intratumoral microvessel density (IMVD) and dThdPase expression were evaluated immunohistochemically using anti-CD34 and anti-dThdPase antibodies, and results were correlated with clinicopathologic parameters and prognosis. IMVD for the 54 tumors ranged from 22.5 to 120.7 (number/0.73686 mm2/field). Twenty-three tumors were positive, and 31 tumors were negative for dThdPase expression. IMVD positively correlated with the expression of dThdPase (p < 0.01), tumor size, and peritoneal metastases (p < 0.05). However, there was no statistical correlation between IMVD, dThdPase expression, and clinical outcome. Of the 54 patients examined, 30 were diagnosed with International Federation of Gynecology and Obstetrics (FIGO) stage III or IV primary ovarian carcinoma, and 9 were diagnosed with SSPC. There were no significant differences between the two groups with respect to clinicopathologic features, IMVD, dThdPase expression, or patient outcome. In conclusion, angiogenic activity may be necessary for the growth of metastatic implants in ovarian carcinoma and SSPC. PMID- 11109166 TI - Tumor proliferation in squamous cell carcinoma of the vulva. AB - Tumor proliferation is of important prognostic significance for several neoplasms. The very few previous studies on this parameter in vulvar carcinoma have shown contradictory results. The aim of this study was to determine the prognostic significance of tumor proliferation in vulvar carcinoma. Paraffin embedded tissue of 74 squamous cell carcinomas of the vulva was immunostained for MIB-1, detecting Ki-67, and analyzed for staining patterns and the percentage of positive cells. There were three general staining patterns: a diffuse distribution (diffuse type), a localized staining at the infiltrating tumor border (infiltrating type), and a localized staining in basal parts of infiltrating tumor cell aggregates (basal type). The percentage of positive cells was not correlated with morphologic or clinical parameters, nor was it correlated with disease-free and overall survival. MIB-1 staining types were correlated with tumor type and grading. Tumors of diffuse and infiltrating type seemed to have more frequent lymph node metastasis (p = 0.053) and shorter disease-free survival (p = 0.076). In these tumors, overall survival time was reduced significantly (p = 0.02). In multivariate analysis, MIB-1 staining types were the most important factor for overall survival with an odds ratio of 4.73. In conclusion, distribution and not the percentage of proliferating cells is of prognostic significance in squamous cell carcinoma of the vulva. PMID- 11109167 TI - Inverted follicular keratosis of the vulvar skin: a lesion that can be confused with squamous cell carcinoma. AB - Although seborrheic keratoses of the vulva are described in textbooks, to our knowledge, inverted follicular keratosis has not been reported. A 27-year-old woman underwent an excisional biopsy for a small lesion of the left labium majus. Squamous cell carcinoma was considered in the clinical differential diagnosis. The initial pathologic diagnosis suggested squamous cell carcinoma in situ, and the consultation diagnosis was superficially invasive squamous cell carcinoma. On pathologic examination, a symmetrical, endophytic, epithelial tumor was observed consisting of a proliferation of basaloid cells with many areas of reactive squamous cells showing numerous squamous eddies, focal reactive nuclear atypia, and occasional mitotic figures. After the pathologic diagnosis of inverted follicular keratosis was made, a history of close perineal shaving and total body tanning was obtained. Because inverted follicular keratosis is postulated to be related to follicular injury, it is likely that the trauma of close shaving is a significant etiologic factor. There is less evidence that ultraviolet ray exposure is of etiologic importance. PMID- 11109168 TI - Benign persistent papular acantholytic and dyskeratotic eruption of the vulva: a case report. AB - We report a case of 44-year-old woman with persistent pruritic papules on the left and right labium majus of the vulva. Histopathologic examination of the vulvar biopsy specimen revealed a suprabasal separation of the epidermis with acantholysis and dyskeratosis. PMID- 11109169 TI - Concomitant adenoma and hybrid carcinoma of salivary gland type arising in Bartholin's gland. AB - We report a unique case of a salivary gland type of "hybrid carcinoma" arising within a Bartholin's gland adenoma. The tumor was characterized by large areas of an epithelial-myoepithelial carcinoma similar to that of the salivary gland with a peripheral infiltrative pattern of an adenoid cystic carcinoma (ACC). PMID- 11109170 TI - Epithelioid trophoblastic tumor metastatic to the vagina: an immunohistochemical and ultrastructural study. AB - We describe an epithelioid trophoblastic tumor (ETT) metastatic to the vagina in a 30-year-old Japanese woman. A polypoid tumor in the vaginal orifice was composed of nests of intermediate trophoblastic cells that showed a striking epithelioid appearance. In the hysterectomy specimen, a tumor infiltrated through the myometrium and showed histologic findings similar to those of the vaginal tumor. The tumor cells were positive for cytokeratin, inhibin-alpha, and melanoma cell adhesion molecule (Mel-CAM, CD146) but were only focally positive for human placental lactogen. Electron microscopic examination revealed bundles of well developed, intermediate-type filaments surrounding the nuclei. PMID- 11109171 TI - Identity testing in cervical carcinoma in case of suspected mix-up. AB - The histopathologic diagnosis is the cornerstone of modern oncology. But mix-ups of specimens can occur at any stage. The resection of a 1.2 cm polypoid cervical mass in a 25-year-old woman showed a poorly differentiated adenocarcinoma prospectively staged as T1b1 (International Federation of Gynecology and Obstetrics IB1). Even after complete embedding and serial sectioning of the whole cervix of the hysterectomy specimen after radical hysterectomy, only adenocarcinoma in situ, but no invasive tumor, was seen. To exclude a mix-up of the specimens, identity testing of the paraffin-embedded material was performed by microsatellite analysis. For both materials, we established identical results after testing the microsatellite loci HumTH01, HumVWA, HumFGA, HumACTBP2, HumF13B, and HumD8S1132. The resulting probability of identity came to 99.9999%, excluding a mix-up of the specimens. Archival paraffin-embedded specimens can be used to establish identity and can prevent the wrong patient from having major surgery. PMID- 11109172 TI - Primary ovarian dysgerminoma in a patient with a germline BRCA1 mutation. AB - Germline mutations in the BRCA1 tumor suppressor gene are associated with increased risk for the development of ovarian cancer. All such cancers thus far reported have been of the epithelial histologic type. We identified an ovarian dysgerminoma in a 16-year-old woman (proband) with a family history of ovarian cancer during a review of histopathologic characteristics of ovarian cancers from women enrolled in the Gilda Radner Familial Ovarian Cancer Registry. Mutation analysis of DNA from this patient's peripheral blood leukocytes revealed a germline BRCA1 mutation (3312insG). The mutation was also present in the mother with breast cancer, a maternal aunt and a distant cousin with ovarian cancer, and a maternal grandfather and an uncle with skin cancer. The development of the proband's dysgerminoma may be unrelated to her germline BRCA1 mutation. Alternatively, such dysgerminomas may be caused by BRCA1 mutations, but occur so infrequently compared with epithelial cancers that they are seldom identified. Analysis of a larger series of ovarian germ cell tumors may resolve this question. PMID- 11109173 TI - Mitotic arrest of endometrial epithelium after paclitaxel therapy for breast cancer. AB - We report the histopathologic findings in endometrial curettings from a 31-year old woman with dysfunctional uterine bleeding who had received paclitaxel therapy for breast carcinoma. Paclitaxel, a member of the taxane family of antineoplastic agents that is used in the treatment of breast carcinoma, ovarian carcinoma, and non-small cell lung carcinoma, acts by the simultaneous promotion of tubulin assembly into microtubules and inhibition of microtubule disassembly. The curettings in this case showed fragmented menstrual phase endometrium with striking numbers of mitotic figures. Cell divisions were arrested in metaphase. Glandular epithelial cells showed strong immunoreactivity for bcl-2 and MIB-1. We attribute this marked morphologic effect to paclitaxel-induced mitotic arrest of the endometrium. PMID- 11109174 TI - Serous tumor of low malignant potential of the fallopian tube. AB - Although serous tumors of low malignant potential (STLMP) of the ovary are relatively common, similar tumors of the fallopian tube are exceedingly rare. We report the case of an STLMP of the fallopian tube in a 31-year-old woman and review the literature. PMID- 11109175 TI - Inequality in the social consequences of illness: how well do people with long term illness fare in the British and Swedish labor markets? AB - The demand for unskilled labor has collapsed across industrialized societies, including Britain and Sweden, and rates of unemployment and economic inactivity have increased. The result is a reduction in total employment, primarily among men. These trends could be expected to hit particularly hard those people with chronic illness. The study tests two opposing hypotheses: (1) the increasingly flexible, deregulated labor market in Britain would result in an increased number of new jobs, and thus better employment opportunities for unskilled workers, including those with chronic illness; (2) the more regulated labor market in Sweden, with the associated health and social policies, would provide greater opportunities for jobs and job security for workers with chronic illness. Analysis of data on men from the British General Household Survey and the Swedish Survey of Living Conditions, 1979-1995, showed that employment rates were higher and rates of unemployment and economic inactivity were lower in Sweden than in Britain, and the differences in these rates across socioeconomic groups and between those with and without chronic illness were smaller in Sweden. The results support the hypothesis that active labor market policies and employment protection may increase the opportunities for people with chronic illness to remain in work. PMID- 11109176 TI - Job insecurity and health. AB - As employers respond to new competitive pressures of global capitalism through layoffs and the casualization of labor, job insecurity affects a growing number of workers. It appears to harm mental health, but less is known about its effects on physical health and health behaviors and the mechanisms through which it may act. The prevailing individual-centered conceptualization of job insecurity as the perception of a threat to job continuity precludes systematic investigation of the social patterning of its health effects. Analysis of data from a 1994 Canadian national probability sample of adults determined that high levels of job insecurity lowered self-rated health and increased distress and the use of medications, but had no impact on heavy drinking. The findings support one possible mechanism of action whereby job insecurity reduces feelings of control over one's environment and opportunities for positive self-evaluation; these psychological experiences, in turn, have deleterious health consequences. There is little evidence of social patterning of this relationship by gender, education, household income, age, marital status, and social support at work. PMID- 11109177 TI - Health effects of job insecurity among employees in the Swiss general population. AB - This study looks at the health consequences of the social distress caused by perceived levels of job insecurity. Through interviews with full- and part-time employees drawn from a random sample (N = 2,024) of the Swiss general population, the authors measured prevalence rates of ten self-reported indicators of health and health-related behavior according to three levels of perceived job insecurity (low, middle, high), and estimated odds ratios using logistic regression adjusted for relevant respondent characteristics. The results show that the psychosocial stress induced by job insecurity (fear of unemployment) has a negative effect on these health indicators. Fear of unemployment had a stronger unfavorable effect on health for highly educated employees than for the less educated. The authors make some recommendations for raising awareness about the health effects of job insecurity and taking these effects into account in policies and legislation affecting the labor market and work environment. PMID- 11109178 TI - Agricultural "killing fields": the poisoning of Costa Rican banana workers. AB - The poisoning of Costa Rican banana workers by multinational corporations' excessive use of pesticides is not a local issue; it is embedded in a dominant ideology expressed by the phenomenon of globalization. This ideology seeps into every aspect of our social institutions--economic, political, and legal. The practice of this ideological perspective is evident in the industrialization of global agriculture and the shift from "developmentalism"--liberal welfarism, industrialization, and urbanization--to a dominant, undemocratic, global financial elite with "economism" and a neoliberal political agenda overriding the nation-state polis. A specific effect is to transform the agricultural workers of developing countries, such as Costa Rican banana workers, into politically superfluous flesh-and-blood human beings. PMID- 11109179 TI - Strontium-90 in deciduous teeth as a factor in early childhood cancer. AB - Strontium-90 concentrations in deciduous (baby) teeth of 515 children born mainly after the end of worldwide atmospheric nuclear bomb tests in 1980 are found to equal the concentrations in children born during atmospheric tests in the late 1950s. Recent Sr-90 concentrations in the New York-New Jersey-Long Island metropolitan area have exceeded the expected downward trend seen in both baby teeth and adult bone after the 1963 ban on atmospheric testing. Sharp rises and declines are also seen in Miami, Florida. In Suffolk County, Long Island, Sr-90 concentrations in baby teeth were significantly correlated with cancer incidence for children 0 to 4 years of age. A similar correlation of childhood malignancies with the rise and decline of Sr-90 in deciduous teeth occurred during the peak years of fallout in the 1950s and 1960s. Independent support for the relation between nuclear releases and childhood cancer is provided by a significant correlation with total alpha and beta activities in local surface water in Suffolk County. These results strongly support a major role of nuclear reactor releases in the increase of cancer and other immune-system-related disorders in young American children since the early 1980s. PMID- 11109180 TI - Primary care, income inequality, and self-rated health in the United States: a mixed-level analysis. AB - Using the 1996 Community Tracking Study household survey, the authors examined whether income inequality and primary care, measured at the state level, predict individual morbidity as measured by self-rated health status, while adjusting for potentially confounding individual variables. Their results indicate that distributions of income and primary care within states are significantly associated with individuals' self-rated health; that there is a gradient effect of income inequality on self-rated health; and that individuals living in states with a higher ratio of primary care physician to population are more likely to report good health than those living in states with a lower such ratio. From a policy perspective, improvement in individuals' health is likely to require a multi-pronged approach that addresses individual socioeconomic determinants of health, social and economic policies that affect income distribution, and a strengthening of the primary care aspects of health services. PMID- 11109181 TI - Home or hospital? An evaluation of the costs, preferences, and outcomes of domiciliary chemotherapy. AB - This study compares the costs and outcomes of domiciliary and hospital-based chemotherapy, using a prospective randomized cross-over design. Eighty-seven eligible patients were recruited from oncology services at two metropolitan hospitals in Sydney, Australia. Forty patients completed study evaluation requirements, having two months of chemotherapy in each location (home and hospital). The domiciliary service was staffed by hospital-based oncology nurses. Marginal costs of domiciliary treatment over hospital treatment were estimated from the health service perspective. Home-based care was more expensive, largely due to extra nurse time. About half of the eligible patients (n = 87) and 73 percent of the evaluated patients (n = 40) preferred domiciliary care. Most evaluated patients and their informal carers were satisfied with the medical care provided, regardless of location. Patient needs were well met in either location, and no differences were found in quality of life. At current throughput rates, providing chemotherapy in the home was more expensive than providing it in hospital. However, if the demand for chemotherapy were to exceed ward capacity by up to 50 percent, moving chemotherapy into the home could provide a less costly strategy for the expansion of a chemotherapy service without compromising patient outcomes. PMID- 11109182 TI - Happy with health care? AB - The American Association of Health Plans (the main HMO trade association), in making the case against patients' rights legislation, points to polling data that show Americans are basically satisfied with managed care plans. Although large majorities, including those with HMOs, do say they are "satisfied" with their health care plans, HMO members are less satisfied than members of other types of plans. And if we look beyond personal-satisfaction ratings, we find plenty of evidence for public concern about HMOs in particular and the health care system in general. Americans are supportive of HMO regulation, and despite their willingness to say they are "satisfied" with their health care plans, they harbor a lot of worries about the future--treatment that could be denied them, costs that could ruin them, and loss of coverage. The public sees the need for major change not just in HMOs but in the health care system as a whole. As HMO lobbyists scramble for new arguments against legislation, they will likely persist in misrepresenting and misusing polling data to make their case. PMID- 11109183 TI - The influences of drug companies' advertising programs on physicians. AB - This study investigates the influences of drug companies' advertising programs on physicians. Of the 446 physicians interviewed, 53.9 percent were visited by pharmaceutical company representatives at least once a day, and 43.5 percent spent 15 minutes or more per day on these visits. With respect to the information delivered by the pharmaceutical company representatives, 67.7 percent of physicians thought it was not reliable, and 62.8 percent reported that it had no effect on their prescription writing. The promotional gifts had little effect on prescriptions for 43.9 percent of physicians, and 80.3 percent reported that these gifts were distributed unequally among doctors according to the drugs they prescribed. Only 23.5 percent of physicians supported the prohibition of promotion programs; 90.6 percent of physicians agreed that drugs are too expensive, and 82.9 percent agreed on the presence of overprescription. The authors evaluate these results and provide some suggestions for improving the sources of information for drug prescribing. PMID- 11109184 TI - Pharmaceutical expenditure in Spain: cost and control. AB - In recent years, the Spanish government has been battling to keep pharmaceutical expenditures under control. Its measures include control of prices, introduction of a "negative list" of drugs no longer reimbursed, increased cost-sharing, and introduction of overall budgets for pharmaceutical expenditures. Although the average prices of old pharmaceutical products declined by 39 percent over the last 15 years and consumption in value increased by only 10 percent, real pharmaceutical expenditures in Spain increased by 264 percent over that period. The main reason for the continuing rise in these expenditures and the failure of cost-containment measures is the introduction of new, more expensive drugs, which often fail to offer any real therapeutic advantages over products already on the market. This situation is exacerbated by a lack of effective demand-side measures such as budgets for doctors and lack of a generics market. PMID- 11109185 TI - Managing medical technology: lessons for the United States from Quebec and France. AB - Important modifications to technology assessment, diffusion, adoption, and utilization must take place if the United States is to better employ medical technology and save resources so as to assure access for the uninsured and underinsured. The United States can learn from other health systems that are more successful in achieving these goals. The author selects for comparison the health systems of France and Quebec. The discussion focuses on the differences between the three systems in the management of medical technology on a range of policy relevant dimensions, including health system structure, attitudes about planning versus market competition, government regulation, the balance between decentralization and centralization, the needs of the individual and those of the society, linkages between technology assessment and policy-making, and the importance of medical technology assessment for medical practice. Seven specific recommendations are made for better managing medical technology in the United States, drawing on what can be observed from the experiences of Quebec and France. PMID- 11109186 TI - A review of data on the health sector of the United States. AB - This report provides data on the state of U.S. health care at the start of the new century. It reveals increasing numbers of uninsured and underinsured Americans; increasing costs for health insurance, health care services, and medicines; and increasing inequalities in health and in access to health care. The author also provides data on the current state of the pharmaceutical and health service industries, including Medicaid and Medicare HMOs. The results of some opinion polls on health care, conducted among physicians and the general public, are also summarized. PMID- 11109187 TI - [Is a totally non-invasive assessment of the hemodynamic profile possible in patients with chronic heart failure?]. AB - Relevant hemodynamic information can be obtained by a comprehensive Doppler echocardiographic examination in patients with various cardiac diseases. The assessment of left heart hemodynamics by Doppler echocardiography has been addressed by several investigators. The feasibility and the accuracy of methods for the estimation of left ventricular filling pressure and cardiac output have been validated by comparative right heart catheterization. Studies have shown that Doppler echocardiography can allow the measurement of pulmonary artery pressures from the pressure gradients across the tricuspid and pulmonary valves. The possibility of completely characterizing cardiac hemodynamics noninvasively has recently been documented: in patients with acute myocardial infarction, automated cardiac output measurement along with the assessment of left ventricular filling by Doppler echocardiography may be used for the identification of hemodynamic subsets. Although Doppler echocardiography can provide noninvasive measures of hemodynamic indices, its value has been disputed since the technique is patient-dependent, time-consuming and requires meticulous acquisition and interpretation by skilled operators. The use of contrast agents may improve the accessibility of both right-sided and left-sided Doppler signals, potentially increasing the number of patients to whom the noninvasive hemodynamic assessment could be applied. PMID- 11109188 TI - [Echocardiographic follow-up of patients with heart failure. Which parameters should be measured? How often?]. AB - The echocardiographic evaluation keeps a relevant place in the evaluation of patients with heart failure and left ventricular systolic dysfunction, not only for its contribution to the diagnosis, prognostic stratification and comprehension of pathogenetic mechanisms, but also for the analysis of the evolution of the disease and the response to optimal medical therapy. On the other hand, the role of echocardiography in the follow-up of patients with diastolic dysfunction is still unclear. In patients with heart failure and left ventricular systolic dysfunction the analysis of changes in left ventricular function and dimension during follow-up is particularly relevant to recognize the potential benefit of optimal medical therapy with ACE-inhibitors and beta blockers and their prognostic significance. The echo-Doppler hemodynamic evaluation is also of clinical and prognostic value particularly for the recognition of the persistence or (re)appearance of restrictive filling pattern during follow-up. Moreover, in patients with persistent severe left ventricular systolic dysfunction, the evaluation of right ventricular function may allow for the identification of a subset of patients at high risk for cardiovascular events. A practical flow-chart of echocardiographic assessment of patients with heart failure and left ventricular systolic dysfunction includes the following steps: 1) after 3 to 6 months on optimal therapy, to detect the persistence of restrictive filling pattern, if present at diagnosis; 2) after 12 to 24 months, to analyze the response of left ventricular function and dimension to optimal medical treatment; 3) serial examinations, according to the stage of the disease or to the episodes of worsening heart failure, to identify echocardiographic indicators of disease progression, such as worsening of left ventricular and/or right ventricular function or (re)appearance of restrictive filling pattern. The changes in these parameters seem to have a relevant prognostic significance to define the risk profile of patients with heart failure and left ventricular systolic dysfunction. PMID- 11109189 TI - [Doppler echocardiography after heart transplantation]. AB - Acute rejection, graft atherosclerosis and infections are the major causes of morbility and mortality after heart transplantation. Therefore clinical follow-up after heart transplantation is usually focused on the early diagnosis and treatment of these complications. The early recognition of acute rejection episodes in cardiac allograft recipients remains a major challenge during follow up after heart transplantation. The standard method used to identify rejection is endomyocardial biopsy, but this technique shows some important inherent limitations. Even if noninvasive methods are generally considered inaccurate in the early diagnosis of acute rejection, echocardiography is routinely used in the surveillance of heart transplant recipients to monitor allograft function. Furthermore it may be used to screen significant acute rejection episodes, because it can identify several morphologic and functional changes that are related to acute rejection: increase in left ventricular myocardial thickness, changes in ultrasonic texture, pericardial effusion, systolic and diastolic dysfunction. In this setting it is very useful to perform serial echocardiographic examinations, because the changes of echocardiographic and Doppler parameters in comparison with previous examination in each individual patient can be accurate in detecting acute rejection and evaluating the response after immunosuppressive treatment. The development of coronary artery disease (CAD) is the major factor limiting long-term survival. Serial coronary artery angiography remains the only effective diagnostic tool for detecting CAD, while noninvasive tests commonly used in atherosclerotic heart disease are considered inaccurate, due to their low sensitivity. However several studies reported a high accuracy of dobutamine stress echocardiography in the diagnostic and prognostic assessment of CAD. Furthermore, rest wall motion abnormalities and left ventricular systolic dysfunction can identify patients with significant CAD and worse prognosis. In conclusion, echocardiography cannot supplant invasive methods for monitoring acute rejection and CAD, but it is a useful tool for surveillance after heart transplantation. PMID- 11109190 TI - [Open access echocardiography in patients with suspected or confirmed heart failure]. AB - Cardiac failure is a disease still characterized by high morbidity and mortality. The use of clinical criteria is not yet considered sufficient for the diagnosis of this disease by main scientific associations. Echocardiography can give important information not only for diagnosis, but also for prognosis and management of the disease. As a growing demand for echocardiography is expected in the near future scientific community should be ready to face this difficult challenge. In fact it will be necessary to implement and organize structures in which this technique will be readily and easily available, in order to facilitate the diagnosis and allow "personalized" management and follow-up in these patients. PMID- 11109191 TI - [Revisiting the clinical examination of the patient with heart failure in the light of data obtained with Doppler echocardiography]. AB - Echocardiography plays a central role in the diagnosis and assessment of heart failure as it allows us to discover missed valvular or congenital heart disease and distinguishes the different pathophysiological forms of cardiomyopathies. Echocardiographic examination shows the presence, the severity and the mechanism of mitral regurgitation. It identifies the pathophysiology of heart failure differentiating systolic from diastolic dysfunction. The different patterns of Doppler transmitral flow curves are correlated with left ventricular filling pressures and with functional capacity of the patient. The assessment of functional impairment of the patient is based not only on the measurement of left ventricular volumes and ejection fraction, but rather on an integrated analysis of several clinical and echocardiographic features including right ventricular volumes and ejection fraction. Prognostic information can be obtained from the echocardiographic examination: a short (< 125 ms) deceleration time of early filling in transmitral flow curve and its persistence despite optimized medical treatment identify patients at very high risk of subsequent death and/or hospitalization. Similarly, pulmonary hypertension measured by Doppler echocardiography is a predictor of poor outcome. Since Doppler echocardiographic assessment of hemodynamic variables in patients with advanced heart failure is accurate and reproducible, this non-invasive methodology may assist with monitoring and optimizing medical therapy in such patients. The symptoms of heart failure as well as the clinical and radiographic findings are neither sensitive nor highly specific to diagnosis. Furthermore clinical findings are not reliable for estimating hemodynamics of the individual patient. At present, the echocardiogram is the most useful and widespread tool used in heart failure patients, with regard to diagnosis, assessment and hemodynamic characterization. Due to the complex nature of the heart failure syndrome, for an exhaustive assessment of the patient, echocardiographic data should be integrated with clinical and laboratory findings. PMID- 11109192 TI - [Heart failure in childhood]. AB - Heart failure (HF) in pediatric patients shows many differences from adult HF because of biochemical and ultrastructural peculiarity due to contractility modifications for different loading conditions, often resulting from congenital heart diseases. The majority of heart diseases in pediatric patients with symptoms and signs of HF are due to severe heart diseases (such as ventricular septal defect, severe aortic stenosis, aortic coarctation syndrome). There are several reasons for early HF in the natural evolution of congenital and acquired heart diseases. They are of hemodynamic nature (pressure and volume overloading, reduced ventricular compliance), or due to systolic ventricular dysfunction. In order to optimize the medical and/or surgical treatment in these conditions an immediate etiological and pathophysiological evaluation should be done. In this regard the echocardiographic technique allows to reach a rapid and accurate diagnosis and a good therapeutic view in a non-invasive and easily to repeat way. PMID- 11109193 TI - [Role of echocardiography in the treatment of heart failure with permanent electric stimulation]. AB - Permanent cardiac pacing has been proposed for the treatment of atrioventricular and intraventricular conduction defects and related hemodynamic alterations which may worsen the performance of the failing heart. The initial positive results of right sided atrioventricular synchronous pacing have not been confirmed in later studies involving a larger number of patients with different clinical characteristics. The reason of these conflicting results may be related to the poor understanding of the complex interaction between the adopted pacing mode and the different type, grade and hemodynamic significance of conduction defects. The negative hemodynamic effects of the altered sequence and synchrony of ventricular activation during right sided pacing may outweigh the benefits of an optimal atrioventricular synchrony. Biventricular stimulation has been proposed to improve the electromechanical activation of the left ventricle in patients with left bundle branch block. Ongoing prospective studies are evaluating the potential benefits of biventricular stimulation versus alternative treatments. Although there are no standard indications to cardiac pacing in heart failure it seems that this therapeutic tool may be of value in selected patients with conduction defects which unfavorably affect the cardiac function and that are amenable to be corrected by an appropriate pacing modality. Doppler echocardiography, in its different applications, emerges has a key technique for the selection of patients who may benefit from permanent pacing and for the selection of the best pacing modality. Doppler echocardiography criteria may also be useful in the selection of homogeneous groups of patients to be enrolled in prospective studies aimed at assessing the potential benefits of permanent pacing versus alternative treatments. PMID- 11109194 TI - [Hemodynamic and coronary angiographic performance at a center with medium-low activity: organization model, activity indicators, and costs]. AB - BACKGROUND: In order to evaluate the cost-effectiveness of coronary angiography performed in a low volume Center, we examined our 1-year activity. METHODS: The organizational model of the multipurpose cardiac catheterization laboratory is described. In this type of facility both coronary angiographic and electrophysiological studies are performed. To evaluate the laboratory performance we examined the utilization level, the appropriateness of the studies, the complication rates and the number of studies that had to be repeated because of inadequate data or image quality. The costs were calculated for the in house laboratory setting (the actual scenario) and for the 25 km distant laboratory setting (the historical scenario). RESULTS: The laboratory caseload of coronary angiography was 342 studies, 46% of the overall laboratory activity; 175 patients (51%) underwent non-pharmacological therapy, 129 patients (38%) were treated with medical therapy; the percentage of patients with normal coronary arteries was 11%. Two patients (0.58%) had vascular complications, 1 patient (0.29%) developed an acute myocardial infarction 2 hours after coronary angiography without any evidence of angiographic modifications at the repeated study. In no patient the study had to be repeated because of inadequate data or image quality. The mean cost of a coronary angiography was Lit. 512,000 (265 Euro) for the actual scenario; it would have been Lit. 694,000 (359 Euro) for the historical scenario, with Lit. 182,000 (94 Euro) saved. CONCLUSIONS: These findings are consistent with the accepted criteria of good laboratory performance and cost-effectiveness. Thus coronary angiography can be performed effectively and efficiently in a low volume Center. PMID- 11109195 TI - [Assessment of plasma lipid profile in acute coronary syndromes. The use of fibrinolytics and heparin do not affect it significantly]. AB - BACKGROUND: Neurohormonal and metabolic responses to acute ischemic events associated with thrombolysis and heparin induce substantial changes in the lipid profile (acute phase response). The aim of this study was to assess changes in total cholesterol and triglycerides in patients with acute coronary syndrome, admitted to our Intensive Coronary Care Unit (ICCU). METHODS: The study included 1051 consecutive patients, 316 with unstable angina, 583 with Q wave acute myocardial infarction (AMI) and 152 with non-Q wave AMI. Total cholesterol and triglycerides were measured in all patients at time 0 (admission), at time 1 (the morning following admission), at time 2 (the morning after discontinuation of heparin treatment). RESULTS: The mean value of total cholesterol at admission was 235, 210 and 197 mg% at admission, time 1 and time 2, respectively. Triglyceride levels were 234, 178 and 189 mg%, respectively. In the subgroup of thrombolized AMI the reduction in total cholesterol at time 1 and time 2 resulted similar in comparison with non-thrombolized AMI (p = NS). The decrease in triglycerides showed a similar pattern in the different subgroups. Comparison was also done according to sex, age, and complications. CONCLUSIONS: These data confirm that mean total cholesterol and triglycerides at admission are sharply higher than values considered normal in the literature. Within 24 hours of admission there is a 10.7% drop in total cholesterol which increases to 16.2% after a few days (mean 3.4 days). Total cholesterol determination upon admission in patients with acute coronary syndromes is necessary in order to know the true concomitant lipid profile during the precipitating ischemic events. The decision of initiating early therapy with statins would then appear more justified. PMID- 11109196 TI - [Assessment of cardiotoxicity of high dose cyclophosphamide with electrocardiographic, echocardiographic, and troponin I monitoring in patients with breast tumors]. AB - BACKGROUND: High-dose cyclophosphamide is the nucleus for virtually all high-dose chemotherapy protocols. Non-hematologic dose-limiting toxicity is represented by acute cardiomyopathy, even fatal in a minority of patients. The pathophysiology of such a cardiotoxicity is still poorly understood. Postmortem studies revealed hemorrhagic myocardial cell death, endothelial damage, and interstitial edema. Recently troponins, in particular troponin I, have been found to represent uniquely sensitive and specific markers of myocyte membrane integrity, thus to increase in response to myocardial cell damage in different clinical settings. METHODS: We performed a multiparametric monitoring in 16 consecutive breast cancer patients undergoing cyclophosphamide, by means of serial ECGs, cardiac enzymes determinations (creatine phosphokinase, MB mass and troponin I) through 0 to 72 hours, and echocardiography at baseline and after 48 hours. RESULTS: Neither overt cardiac failure nor enzyme elevation were recorded. Serial ECGs revealed a reduction in QRS voltage and/or ST segment abnormalities in 6 cases. Echocardiography showed an increase in left ventricular diastolic and/or systolic diameters and volumes in 4 cases but without any decrease in fractional shortening and ejection fraction under normal values: in 2 of them abnormalities of diastolic function (E/A mitral Doppler ratio, isovolumic relaxation time and deceleration time) were also recorded. CONCLUSIONS: Our protocol of cyclophosphamide administration did not cause cardiac toxicity by myocardial cell damage, as analyzed by troponin I levels, thus suggesting that myocyte membrane injury is not the first mechanism of it. ECG (i.e. QRS voltages) and echo-Doppler (i.e. E/A ratio) monitoring lead to hypothesize that endothelial injury and interstitial edema with subsequent reduction in left ventricular diastolic compliance may be the first signs of cardiac dysfunction in this clinical setting. PMID- 11109197 TI - [Monitoring of ischemic changes of the QRS complex with a new system of computerized electrocardiography]. AB - BACKGROUND: Evoked ischemia induces morphological modifications in QRS complex that can be detected by a new electrocardiographic computerized system that we have developed (ventricular ischemic site analysis-VISA). The aim of this study was to evaluate, by continuous recording, the ischemia evolution during stress or pharmacological test and to correlate the modifications of QRS with the shift of the ST-T segment. METHODS: According to our method, the vectorial differences between rest QRS and stress QRS are recorded as deformations of the QR and/or RS segments. Sixty patients were studied: 44 had inducible ischemia during single photon emission computed tomography 99mTc-sestamibi test; 33 of these underwent ergometric test and 11 pharmacological test. RESULTS: The VISA test was positive in 91% of patients against the 61% with standard ECG. Ischemic deformations of the QRS appeared at 62.1% of stress duration, while the ST-T shift appeared at 81.4% of the test. During pharmacological stress the ST-T segment did not change. CONCLUSIONS: Our method is more sensitive than standard ECG in identifying inducible ischemia. Ischemic deformations of the QRS complex occur earlier than the repolarization modifications and they may be present also in their absence. The higher sensitivity of the VISA test depends on the fact that QRS ischemic modifications are caused both by the lesion current and a conduction delay in the ischemic zone. PMID- 11109198 TI - [Against free market in health]. AB - Universalism is a basic philosophical concept peculiar to the Italian National Health System and as such guarantees free health care for everyone in any public medical structure. On the contrary, liberalism suggests that the medical care system should be based on privately managed health care structures and governed by the laws of the market. In a liberalist society, health care totally depends on each individual's economic possibilities. Money rules the life and health of everyone. The author believes that this fact is ethically unacceptable and incoherent with the culture and social theories of European justice. At present, the revival of liberalism is threatening the universalistic foundation of the Italian National Health System. It is the moral duty of physicians working within any public medical structure, particularly ANMCO associates, to preserve and defend the system. ANMCO should utilize the prestige gained in the past years from its techno-scientific initiatives in a "politically" significant manner to maintain universalism as a philosophical, ethical and juridical principle of the Italian National Health System in the interest of the whole Italian population. PMID- 11109199 TI - [Left atrium rupture after non-penetrating injury to the back]. AB - Survival after cardiac rupture associated with blunt thoracic trauma is very uncommon. In these patients successful management demands a high index of suspicion of cardiac injury. A case of a 24-year-old woman who presented unconscious and shocked in the emergency room after motorcycle trauma strictly limited to her back is reported. Rib and sternal fractures were absent; the typical signs of cardiac tamponade were not found. Therefore the suspicion of cardiac chamber rupture was not immediate and the cardiologist was consulted after several diagnostic exams. Transthoracic echocardiography showed a pericardial effusion with clots and initial cardiac tamponade. The patient was transferred to the operating room and a large hemopericardium was disclosed. Two lacerations were noticed: the first pericardial, near the inferior vena cava, and the second one in the posterior wall of the left atrium. It is possible that the associated pericardial tear and pericardial clots could have contributed to survival. After surgical repair, carried out during cardiopulmonary bypass, the recovery was quick and complete. This case report confirms the possibility of heart chamber rupture after blunt chest trauma even in the absence of obvious thoracic lesion and it shows that the presentation could be very insidious without a "classic" clinical picture of cardiac tamponade. In front of an unexplained shock after nonpenetrating thoracic trauma, a rupture of the heart chambers should be suspected and echocardiography is mandatory. In the emergency room environment pericardiocentesis should be performed only with a quickly available cardiac surgery or in the presence of overwhelming hemodynamic failure. PMID- 11109200 TI - [Biventricular heart pacing and left ventricular heart pacing]. AB - This report describes the case of a 60-year-old patient, affected by alcoholic hypokinetic dilated cardiomyopathy, drug refractory, without surgical indication for ischemic and valvular diseases, implanted with a biventricular pacemaker. The implant was followed by a rapid clinical improvement which allowed the patient's discharge in satisfactory conditions and with strongly reduced diuretic therapy. Ventricular pacing became only left due to increased right ventricular threshold. As a consequence a remarkable decrease in cardiocirculatory compensation was observed, with a new hospitalization due to worsening dyspnea and edema. The instrumental evaluation showed a worsening of the parameters linked to interventricular delay, particularly the interventricular septum activation delay and the reduction in its kinesis. An increase in the ventricular stimulation amplitude led again to a complete capture in both ventricles, with an improvement of interventricular synchronization parameters and septal kinesis. This fact turned into a rapid recovery of satisfactory cardiocirculatory compensation with subsequent patient's discharge. PMID- 11109201 TI - [Relationship of symptom-onset-to-balloon time and door-to-balloon time with mortality in patients undergoing angioplasty for acute myocardial infarction]. PMID- 11109202 TI - [Outcome at 1 year after an invasive compared with a non-invasive strategy in unstable coronary-artery disease: the FRISC II invasive randomised trial]. PMID- 11109203 TI - [Nontraditional risk factors for coronary heart disease incidence among persons with diabetes: the Atherosclerosis Risk in Communities (ARIC) Study]. PMID- 11109204 TI - [Cardiac-specific troponin I levels and risk of coronary artery disease and graft failure following heart transplantation]. PMID- 11109205 TI - [Primary prevention of coronary heart disease in women through diet and lifestyle]. PMID- 11109206 TI - [Telecardiology and local emergency systems]. PMID- 11109207 TI - [Primary coronary angioplasty: an optimal weapon still in search of the best target?]. PMID- 11109208 TI - [Massive pulmonary embolism and contraindications of thrombolysis: role of mechanic thrombectomy]. PMID- 11109209 TI - Fluoride intake and prevalence of dental fluorosis: trends in fluoride intake with special attention to infants. AB - BACKGROUND: Although the predominant beneficial effect of fluoride occurs locally in the mouth, the adverse effect, dental fluorosis, occurs by the systemic route. The caries attack rate in industrialized countries, including the United States and Canada, has decreased dramatically over the past 40 years. However, the prevalence of dental fluorosis in the United States has increased during the last 30 years both in communities with fluoridated water and in communities with nonfluoridated water. Dental fluorosis is closely associated with fluoride intake during the period of tooth development. METHODS: We reviewed the major changes in infant feeding practices that have occurred since 1930 and the changes in fluoride intakes by infants and young children associated with changes in feeding practices. RESULTS AND CONCLUSIONS: Based on this review, we conclude that fluoride intakes of infants and children have shown a rather steady increase since 1930, are likely to continue to increase, and will be associated with further increase in the prevalence of enamel fluorosis unless intervention measures are instituted. RECOMMENDATIONS: We believe the most important measures that should be undertaken are (1) use, when feasible, of water low in fluoride for dilution of infant formulas; (2) adult supervision of toothbrushing by children younger than 5 years of age; and (3) changes in recommendations for administration of fluoride supplements so that such supplements are not given to infants and more stringent criteria are applied for administration to children. PMID- 11109210 TI - Water consumption and nursing characteristics of infants by race and ethnicity. AB - OBJECTIVES: The purpose of this project was to determine racial/ethnic differences in water consumption levels and nursing habits of children younger than 2 years old. METHODS: Data from the 1994-96 Continuing Survey of Food intakes by individuals (CSFII) were used for these analyses. Water consumption and breast-feeding data on 946 children younger than 2 years old were used. RESULTS: For black non-Hispanic children younger than 2 years old (n = 121), 5.3 percent of the children were currently being breast fed. This percentage was less than that seen in other racial/ethnic groups. For white non-Hispanic children (n = 620), this percentage was 10.8 percent; for Hispanic children (n = 146), 12.2 percent; for "other" children, 18.5 percent (n = 59). Black non-Hispanic children had the highest total water consumption (128.6 ml/kg/day) among all groups, white non-Hispanic had the lowest (113.2 ml/kg/day). These differences were not statistically significant in multivariate regression modeling. Black non-Hispanic children also drank more tap water (21.3 ml/kg/day) than white non-Hispanic children (12.7 ml/kg/day) and Hispanic children (14.9 ml/kg/day). The difference was statistically significant in multivariate regression modeling. CONCLUSIONS: The differences in breast feeding and water consumption observed among black children younger than 2 years of age could be a factor in the observed higher levels of fluorosis in black children compared to other children. PMID- 11109211 TI - The prevalence of dental caries and fluorosis in Japanese communities with up to 1.4 ppm of naturally occurring fluoride. AB - OBJECTIVES: The purpose of this study was to determine the relationship between the concentration of fluoride in drinking water and the prevalence of dental caries and fluorosis in seven Japanese communities with different concentrations of fluoride occurring naturally in the drinking water. METHODS: A total of 1,060 10- to 12-year-old lifetime residents were examined to determine the prevalence of dental caries and fluorosis in communities with trace amounts to 1.4 ppm fluoride in the drinking water in 1987. Systemic fluorides (drops or tablets) have never been available in Japan and the market share of fluoride-containing toothpaste was 12 percent at the time of the study. RESULTS: The prevalence of dental caries was inversely related and the prevalence of fluorosis was directly related to the concentration of fluoride in the drinking water. The mean DMFS in the communities with 0.8 to 1.4 ppm fluoride was 53.9 percent to 62.4 percent lower than that in communities with negligible amounts of fluoride. Multivariate analysis showed that water fluoride level was the strongest factor influencing DMFS scores. The prevalence of fluorosis ranged from 1.7 percent to 15.4 percent, and the increase in fluorosis with increasing fluoride exposure was limited entirely to the milder forms. CONCLUSIONS: The findings of this study conducted in 1987 in Japan parallel those reported by Dean et al. in the early 1940s. PMID- 11109212 TI - How do routines of daily activities and flexibility of daily activities affect tooth-cleaning behavior? AB - OBJECTIVES: The aims of this study were: (1) to investigate the level of routines and flexibility of people's daily activity and to identify how tooth cleaning fits into these activities; and (2) to evaluate the impact of different levels of routines and flexibility in daily living on pattern (frequency of tooth cleaning), structure (range of items used in tooth cleaning), performance (relative effectiveness of tooth cleaning) and the outcome of performance (gingival bleeding on probing) in tooth cleaning. METHODS: A convenience sample of 471 Brazilians aged 24 to 44 years was selected from factories, offices, banks, shops, and hospitals. Behavioral, socioeconomic, and clinical data were collected through structured interviews. Dental plaque and gingival bleeding were assessed by clinical examination. Data were analyzed by means of logistic regression. RESULTS: A highly significant relationship was observed between routines of daily activities and tooth-cleaning pattern (OR = 2.3; 95% CI = 1.34, 3.92) after adjusting for age, sex, marital status, and socioeconomic status. No significant associations were observed between routines of daily activities and gingival bleeding. A significant association was observed between tooth-cleaning frequency (OR = 1.6; 95% CI = 1.07, 2.49), performance (OR = 2.7; 95% CI = 1.77, 4.14), outcome (OR = 2.3; 95% CI = 1.31, 3.18), and flexibility of daily activities. Those who had more flexibility of daily activities had lower gingival bleeding scores. CONCLUSION: People who have a less routinized and more flexible day have higher tooth-cleaning frequency than those who have a less flexible and more routinized day. In this study, those with a more flexible day also cleaned their teeth more effectively than those who had a less flexible day, and had reduced gingival bleeding. PMID- 11109213 TI - Rampant early childhood dental decay: an example from Italy. AB - OBJECTIVES: This study sought to estimate the prevalence and related prediction factors for dental caries in 3- to 5-year-old children in Rome, Italy. METHODS: From a sample of 2,025 children, 1,494 (73.8%) were included in the analysis. Children with at least two primary maxillary incisors showing evidence of caries experience were considered affected by rampant early childhood dental decay (RECDD). Behavioral and socioeconomic variables, mutans streptococci counts, diet, and nutritional status were investigated for their association with RECDD using regression analysis. RESULTS: The prevalence of any caries was 27.3 percent, and was 7.6 percent for RECDD. Among all children, mean dft and dt scores per person were 1.1 (SD = 2.4) and 0.9 (SD = 2.3), respectively; among those classified as having RECDD, scores were 6.9 (SD = 4.2) and 6.7 (SD = 4.3), respectively. Children with RECDD had 56 percent of all the decayed teeth in the sample. Low and medium social classes, use of a baby bottle filled with sweetened beverages, high salivary mutans streptococcal levels, and malnutrition were directly associated with RECDD; milk and yogurt consumption and low Plaque Index scores were inversely associated with the condition. CONCLUSIONS: The high prevalence of RECDD suggests that the implementation of preventive programs should be a priority for dental public health. Because of its high prevalence among children as young as 3 years of age, preventive measures targeted toward pregnant women and toddlers should be developed and tested, while kindergarten students could be used for monitoring RECDD prevalence and for detection of communities at risk. PMID- 11109214 TI - Low birth weight and dental fluorosis: is there an association? AB - OBJECTIVE: The association between low birth weight and dental fluorosis was explored in a cross-sectional study to explain the higher prevalence of dental fluorosis among African-American children. METHODS: Birth weight data on 960 children were obtained from the New York State Birth Registry. Data on race, fluoride exposure, sociodemographic characteristics, and dental fluorosis were available from a cross-sectional study conducted in Newburgh and Kingston. Associations among birth weight, race, and fluoride exposure from fluoridated water, regular use of supplements, brushing before the age of 2 years, and subject-level dental fluorosis were explored using logistic regression procedures. RESULTS: The regression coefficients for the main effects and the two way interaction effects associated with low birth weight, race, and fluoride exposure were not statistically significant. Even after controlling for low birth weight and fluoride exposure, African-American children had a statistically significant higher odds ratio (OR = 2.0). An analysis of the data limited to mandibular permanent first molars showed similar effects, except for evidence of effect modification in low birth weight children exposed to fluoride supplements. CONCLUSIONS: Lower birth weight did not explain the higher prevalence of dental fluorosis observed among African-American children. PMID- 11109215 TI - Access and use of specific dental services in HIV disease. AB - OBJECTIVES: This study examined factors associated with the use of specific dental services by persons with HIV disease. METHODS: The data were derived from 1,588 adults who participated in a series of up to six interviews as part of the AIDS Cost and Service Utilization Surveys. Use of dental services such as examinations, x-rays, cleaning, fillings, extractions, root canals, crown and bridge or dentures, and periodontal procedures were evaluated using logistic regression and generalized estimating equations were applied. RESULTS: Multivariate analyses showed that medical insurance, an education beyond high school, income higher than $1,300 per month, high ambulatory visits, and receipt of psychological counseling were generally associated with higher service use. Blacks, those with an inpatient admission, and CD4+ cell counts less than 500 cells/microL were significantly less likely to use most types of dental services. CONCLUSIONS: The study concludes that disparities exist in the use of several dental services similar to those seen in the general population. PMID- 11109216 TI - Health care savings from microbiological caries risk screening of toddlers: a cost estimation model. AB - OBJECTIVES: Modeling new biomedical technologies and determining their expected cost is necessary before initiating formal clinical trials. This paper estimates an economic model for the potential cost impact of microbiological screening of toddlers for caries risk compared to the traditional method of managing pediatric caries. METHODS: Potential cost savings were calculated based on screening test properties (sensitivity and specificity) derived from a population of 1,180 children aged 1 to 3 years with a caries prevalence of 15 percent. An algorithm was then developed to allocate prevalent and anticipate incident caries, treatment effectiveness assumptions, and existing regional treatment costs. RESULTS: The cost analysis model conservatively predicts savings of 7.3 percent from screening and early intervention. Cumulative dental treatment costs for a child at age 4 years are $367.90 if the child has been screened and $396.70 otherwise. The model further predicts that cost savings increase significantly as caries prevalence increases. CONCLUSIONS: Microbiologic risk assessment for pediatric caries may be an example of a preventive public health screening technique that results in both clinical benefits and cost savings. If the model is validated by randomized clinical trials, microbiologic screening could be used by pediatric primary care providers to identify toddlers who require early referral to dentists for further risk assessment and early caries management. PMID- 11109217 TI - Perceived impact of oral health conditions among minority adolescents. AB - OBJECTIVES: This study assessed the perceived impact of oral health conditions, and the relationship of two measures of self-reported outcome, the RAND SF-36 and the Oral Health Impact Profile (OHIP), to clinical indicators of oral health among inner-city adolescents. METHODS: A convenience sample of 93 minority adolescents completed the RAND SF-36 and the OHIP and 76 of them completed a clinical dental examination assessing DMFS. RESULTS: Participants averaged 14.4 (SD = 1.2) years old; 52 percent were female; and 86 percent were African American. The mean DMFS was 8.8 (SD = 6.3). Participants reported relatively poor general health on the SF-36 as well as poor oral health on the OHIP. None of the SF-36 subscales were significantly related to DMFS. OHIP subscales were consistently related to DMFS--those with worse oral health reported more impacts. With the exception of the bodily pain subscale of the SF-36, the SF-36 and OHIP subscales were significantly correlated with Pearson's correlations ranging from .21 to -.57 (P < .05). CONCLUSIONS: Although the SF-36 and the OHIP were correlated, the OHIP appears to be more highly associated with the impacts of oral health conditions than the SF-36 among inner-city adolescents who reported low general and oral health quality of life. PMID- 11109218 TI - Creating partnerships for improving oral health of low-income children. PMID- 11109219 TI - Dietary determinants of dental caries and dietary recommendations for preschool children. AB - OBJECTIVES: The purpose of this review, commissioned by the Administration for Children and Families, the Health Resources and Services Administration, the Health Care Financing Administration, and the Department of Agriculture's Food and Nutrition Service, was to update the evidence of the dietary factors that affect dental caries, and subsequently formulate dietary recommendations for preschool children based on principles of cariology. METHODS: Literature on the dental caries process, dietary factors affecting dental caries initiation and progression, and nutrition education and counseling were reviewed and synthesized. Dietary guidelines for children at various ages were then constructed based on the review. RESULTS: Dental caries in preschool children is due to a combination of factors, including colonization of teeth with cariogenic bacteria, type of foods and frequency of exposure of these foods to the cariogenic bacteria, and susceptible teeth. Caries risk is greatest if sugars are consumed at high frequency and are in a form that is retained in the mouth for long periods. Sucrose is the most cariogenic sugar because it can form glucan that enables firm bacterial adhesion to teeth and limits diffusion of acid and buffers in the plaque. There is emerging interest in the effects of tooth development and its role in the future dental caries risk of the child. CONCLUSIONS: Nutrition education and counseling for the purposes of reducing caries in children is aimed at teaching parents the importance of reducing high frequency exposures to obvious and hidden sugars. Guidelines include: avoiding frequent consumption of juice or other sugar-containing drinks in the bottle or sippy cup, discouraging the behavior of a child sleeping with a bottle, promoting noncariogenic foods for snacks, fostering eating patterns consistent with the Food Guide Pyramid, limiting cariogenic foods to mealtimes, rapidly clearing cariogenic foods from the child's oral cavity either by toothbrushing or by consumption of protective foods, and restricting sugar-containing snacks that are slowly eaten (e.g., candy, cough drops, lollipops, suckers). Along with nutritional factors, a comprehensive approach to preventing dental caries in preschool children must include improved general dietary habits, good oral hygiene, appropriate use of fluorides, and access to preventive and restorative dental care. PMID- 11109220 TI - Caries risk assessment and prevention: strategies for Head Start, Early Head Start, and WIC. AB - OBJECTIVES: This review updates the evidence regarding caries risk assessment for infants, toddlers, and preschool children and formulates recommendations for preventive strategies for WIC, Head Start, and Early Head Start. METHODS: Literature on caries risk assessment and preventive strategies for infants, toddlers, and preschool children were reviewed and synthesized. Recommendations for WIC, Head Start, and Early Head Start were made based on the review. RESULTS: Individual caries risk for children in WIC, Head Start, and Early Head Start should be based on: (1) previous caries experience, (2) precavity lesions, (3) visible plaque, and (4) perceived risk by examiners. Recommended preventive strategies for WIC and Head Start populations include: (1) daily toothbrushing in Head Start centers using fluoridated toothpaste; (2) fluoride varnish application to children enrolled in WIC, Head Start, and Early Head Start; (3) use of chlorhexidine gels and varnishes (following FDA approval); and (4) increased use of sealants on children with precavity pit and fissure lesions. CONCLUSIONS: Early screening, risk assessment, and preventive programs in WIC, Head Start, and Early Head Start populations hold a great deal of promise for preventing dental decay in high-risk children. PMID- 11109221 TI - Access to dental care for Head Start enrollees. AB - OBJECTIVES: The objectives of this review are to characterize the oral health and dental access of Head Start children, describe barriers to their care, advance strategies to address those barriers, and consider how Head Start Performance Standards can be utilized to maximize oral health and access to dental care. METHODS: Published, programmatic, and solicited data describing oral health status and dental service utilization are reviewed together with reports of conferences exploring access barriers. Head Start performance measures for child health and development services, child health and safety, family partnerships, and community partnerships are individually evaluated for their potential to improve oral health. RESULTS: Head Start children, like all low-income children, enjoy the highest rates of dental coverage (because of Medicaid and the State Child Health Insurance Program), yet these children also experience the highest rates of tooth decay, the most unmet dental care needs, the highest rates of dental pain, and the fewest dental visits. Getting children the dental care they need is problematic because of: multiple barriers associated with public and private dental delivery systems, Medicaid program funding and administration, dental workforce sufficiency and distribution, and issues of culture and communication that stand between parents, children, and caregivers. CONCLUSIONS: To move beyond screening and to access necessary dental care requires integration between medical and dental care, recognition and elimination of barriers to care, an understanding of dental provider types and their capacities, a formally structured referral process, and regular monitoring to ensure that complete care is obtained. Action steps are suggested that can maximize the effectiveness of Head Start Performance Standards. Head Start holds tremendous potential to actively develop and implement policies that can markedly improve both access to needed dental services and the oral health status of young disadvantaged children. PMID- 11109222 TI - A psychoanalytic study of integrity and "good character". AB - When we assess a patient for analysis or reflect on our ongoing work as analysts or teachers of analysis, it seems helpful to consider the complex issues of truthfulness and courage within the self (i.e., integrity) and what it means to have "good character." PMID- 11109224 TI - The psychobiology of late childhood: significance for psychoanalytic developmental theory and clinical practice. PMID- 11109223 TI - Mirror neurons, procedural learning, and the positive new experience: a developmental systems self psychology approach. AB - In summary, we are impressed with the existence of a mirror neuron system in the prefrontal cortex that serves as part of a complex neural network, including afferent and efferent connections to the limbic system, in particular the amygdala, in addition to the premotor and motor cortex. We think it is possible to arrive at an integration that postulates the mirror neuron system and its many types of associated multimodal neurons as contributing significantly to implicit procedural learning, a process that underlies a range of complex nonconscious, unconscious, preconscious and conscious cognitive activities, from playing musical instruments to character formation and traumatic configurations. This type of brain circuitry may establish an external coherence with developmental systems self psychology which implies that positive new experience is meliorative and that the intentional revival of old-old traumatic relational configurations might enhance maladaptive procedural patterns that would lead to the opposite of the intended beneficial change. When analysts revive traumatic transference patterns for the purpose of clarification and interpretation, they may fail to appreciate that such traumatic transference patterns make interpretation ineffective because, as we have stated above, the patient lacks self-reflection under such traumatic conditions. The continued plasticity and immediacy of the mirror neuron system can contribute to positive new experiences that promote the formation of new, adaptive, implicit-procedural patterns. Perhaps this broadened repertoire in the patient of ways of understanding interrelational events through the psychoanalytic process allows the less adaptive patterns ultimately to become vestigial and the newer, more adaptive patterns to emerge as dominant. Finally, as we have stated, we believe that the intentional transferential revival of trauma (i.e., the old-old relational configuration) may not contribute to therapeutic benefit. In contrast, the revival of trauma in the old-new configuration (i.e., in the presence of a helpful other who can reduce anxiety and foster eventual positive new experience) can be beneficial, as trauma research has demonstrated. This is the process that promotes new implicit procedural learning, new-new relational configurations, and a richer understanding of the self narrative. PMID- 11109225 TI - Changes in patients' self-representation over the course of psychotherapy. PMID- 11109226 TI - Affect theory and the neurobiology of affective dysregulation. PMID- 11109227 TI - Tango and its meaning for a culture. PMID- 11109228 TI - Instincts and their physiologies--a clinician's perspective. PMID- 11109229 TI - The couch and the cushion: integrating Zen and psychoanalysis. PMID- 11109230 TI - The "voice of self-respect": women and anger. PMID- 11109231 TI - Childism as vestiges of infanticide. PMID- 11109232 TI - Guidelines for treatment of PTSD. PMID- 11109233 TI - A controlled study of imagery rehearsal for chronic nightmares in sexual assault survivors with PTSD: a preliminary report. AB - Imagery-rehearsal therapy for chronic nightmares was assessed in a randomized, controlled study of sexual assault survivors with posttraumatic stress disorder (PTSD). Nightmares, sleep quality, and PTSD were assessed at baseline for 169 women, who were randomized into two groups: treatment (n = 87) and wait-list control (n = 82). Treatment consisted of two 3-hr sessions and one 1-hr session conducted over 5 weeks. Of 169 participants, 91 women (Treatment, n = 43, Control, n = 48) completed a 3-month follow-up and 78 did not. At follow-up, nightmare frequency and PTSD severity decreased and sleep quality improved in the treatment group with small to minimal changes in the control group. Treatment effects were moderate to high (Cohen's d ranged from 0.57 to 1.26). Notwithstanding the large dropout rate, imagery-rehearsal therapy is an effective treatment for chronic nightmares in sexual assault survivors with PTSD and is associated with improvement in sleep quality and decreases in PTSD severity. PMID- 11109234 TI - Predictors of emotional numbing, revisited: a replication and extension. AB - Litz et al. (1997), theorizing that emotional numbing (EN) is the result of emotional depletion caused by chronic hyperarousal, demonstrated that a cluster of hyperarousal symptoms was a robust predictor of EN symptoms. In the present study, these findings were replicated and extended in two multiple regression analyses of data from a large, multisite investigation (T. M. Keane et al., 1998) of psychophysiological responding by male combat veterans. The arousal (D) cluster of symptoms was again the most robust predictor of EN symptoms, whereas physiological indices of arousal and reactivity accounted for negligible amounts of variance in both regression equations. These findings underscore the possible link between disturbances related to arousal and the capacity of traumatized individuals to express and experience pleasant feelings. PMID- 11109235 TI - Health problems among Latin-American and middle-eastern refugees in The Netherlands: relations with violence exposure and ongoing sociopsychological strain. AB - In two studies (n = 480; n = 156), the health problems (somatic, psychological, and migration-related complaints) of refugees were examined, in relation to violence, demographic, and asylum variables (ongoing sociopsychological strain). High frequencies for torture events and a substantial number of medical complaints were reported, but few cases of diagnosable Posttraumatic Stress Disorder (PTSD) were identified (Study I: 6%; Study II: 11%). Not only reported violence, but also the current social situation contributed to the experiencing of ongoing health complaints. Refugees attributed their somatic and psychological complaints to illness (48%) and to torture (29%) and psychological complaints, in particular, to worries related to the postmigration situation (40%). Paying attention only to health complaints and to past violent experiences is too limited an approach in responding to the needs of refugees. PMID- 11109236 TI - PTSD severity and health perceptions in female victims of sexual assault. AB - In women with chronic posttraumatic stress disorder (PTSD), poor physical health may be related to their PTSD symptoms through an underlying negative affect or distress that accompanies the disorder, through the PTSD symptoms in general, or specifically through the chronic hyperarousal present in the disorder. The current study examined the relative contribution of these factors to reported physical symptoms in female victims of sexual assault. Seventy-six women with chronic PTSD were assessed, using measures of stressful life events, psychological difficulties, and perceived health. Negative life events, anger, depression, and PTSD severity were all related to self-reported physical symptoms; however, PTSD severity predicted self-reported physical symptoms beyond these other variables. Contrary to our hypothesis, the reexperiencing cluster of PTSD, and not the hyperarousal cluster, was related to self-reported physical symptoms. PMID- 11109237 TI - Trauma severity and initial reactions as precipitating factors for posttraumatic stress symptoms and chronic dissociation in former political prisoners. AB - This study explores the relationships among trauma severity, initial trauma reactions, posttraumatic stress disorder (PTSD) symptoms, and dissociation in a group of 98 former East-German political prisoners. Trauma severity and initial reactions were assessed using a persecution checklist and a coping process questionnaire. PTSD symptoms were assessed through a structured clinical interview. Chronic dissociation was evaluated using the Dissociative Experiences Scale. The two assumptions of the study were confirmed by structural equation modeling: (1) Lifetime PTSD symptoms were predominantly predicted by initial reactions to trauma and (2) chronic dissociation was predominantly predicted by trauma severity. The first finding is discussed in relation to the participants' initial trauma processing. The second finding is discussed especially in the context of trauma duration and dissociation-prone coping attempts. Limitations concerning the long-term interval between traumatization and data collection are discussed. PMID- 11109238 TI - Prevalence, characteristics, and long-term sequelae of natural disaster exposure in the general population. AB - A sample of 935 participants from the general population completed a mail-out questionnaire containing the Trauma Symptom Inventory (J. Briere, 1995) and the Traumatic Events Survey (D. M. Elliott, 1992). The lifetime self-reported prevalence of natural disasters in this sample was 22%. Although time from the last disaster to involvement in the study was an average of 13 years, previous disaster was associated with significantly higher scores on 6 of 10 symptom scales. Disaster characteristics (especially the presence of physical injury, fear of death, and property loss) were better predictors of symptomatology than was disaster type. Disaster exposure continued to predict symptomatology after controlling for interpersonal violence history, although interpersonal violence accounted for more overall symptom variance. PMID- 11109239 TI - Predictors of psychological distress following serious injury. AB - Posttraumatic psychological distress was assessed in 109 survivors of serious physical injury during acute hospitalization and at 3 months postdischarge. Participants had an average of 4.4 injuries, with a mean injury severity score of 15.5, denoting moderate to severe injuries. Using the Impact of Event Scale (IES), the mean total IES score in-hospital was 22.5 and at 3 months postdischarge was 30.6. Approximately 32% of individuals experienced high levels of distress in-hospital, and this increased to 49% at 3 months postdischarge. The regression model that best explained the variance in posttraumatic psychological distress at 3 months postdischarge included greater psychological distress during hospitalization, a positive drug/alcohol screen on hospital admission, younger age, and the lack of anticipating problems returning to normal life activities. These findings suggest that factors present during acute hospitalization may be used to identify individuals at risk for increased psychological distress, several months following serious physical injury. PMID- 11109240 TI - Mental health service utilization by victims of crime. AB - Records of 318 adult and 608 child victims of crime, eligible for Crime Victims Compensation (CVC) in Washington State, were examined. Demographic, crime, and mental health information was collected. A majority of child victims had experienced sexual assault (88%); adults were victims of both sexual (38%) and physical assault (40%). The median number of mental health sessions used by children was 23 sessions, at an average cost of $975 per case to the CVC program. Adults used a median of 15 mental health sessions at a cost of $905. Patterns of treatment utilization were associated with some demographic, crime, and psychological variables. Sexual assault and PTSD diagnosis were associated with greater use for both children and adults. Therapist variables were unrelated to use. Findings are discussed in light of concerns about coverage of mental health services. PMID- 11109241 TI - An assessment of gender differences in the perception of benefit resulting from the loss of a child. AB - The current study focused on a sample of adults (N = 67) experiencing bereavement following the loss of a child. The Post Traumatic Growth Inventory (PTGI) was used to assess whether bereaved parents were able to perceive benefit from their trauma, and whether there were gender differences in perception of benefit. The impact of the following variables on the PTGI was also assessed: the nature and length of time since the loss, and the age and marital status of the bereaved. The results indicated that bereaved parents do perceive benefit from their loss. However, there was poor evidence to suggest perception of benefit along gender lines. Results also indicated a potential relation between greater perception of benefit and those bereaved through illness, and more perception of benefit for the longer the time elapsed since the bereavement. Lastly, there was a tendency for younger individuals and married respondents to obtain higher scores on the PTGI. PMID- 11109242 TI - Psychometric properties of the Stanford Acute Stress Reaction Questionnaire (SASRQ): a valid and reliable measure of acute stress. AB - A reliable and valid measure is needed for assessing the psychological symptoms experienced in the aftermath of a traumatic event. Previous research suggests that trauma victims typically experience dissociative, anxiety and other symptoms, during or shortly after a traumatic event. Although some of these symptoms may protect the trauma victim from pain, they may also lead to acute stress, posttraumatic stress, or other disorders. The Stanford Acute Stress Reaction Questionnaire (SASRQ) was developed to evaluate anxiety and dissociation symptoms in the aftermath of traumatic events, following DSM-IV criteria for acute stress disorder. We present data from multiple datasets and analyses supporting the reliability and construct, convergent, discriminant, and predictive validity of the SASRQ. PMID- 11109243 TI - Hostility and functional health status in women veterans with and without posttraumatic stress disorder: a preliminary study. AB - This study investigated hostility and functional health status in 90 women veterans with and without PTSD. Compared to women without PTSD, women veterans with PTSD reported significantly higher levels of hostility. Minority status was associated with increased hostility. Compared to a national sample, hostility scores for women with PTSD were greater by a factor of 1.5 PTSD diagnosis was also associated with poorer functioning on all SF-36 Health Survey scales. Controlling for age and education, hostility was related to all SF-36 Health Survey scales in the women with PTSD. PMID- 11109244 TI - Physicochemical characterization of a new crystal form and improvements in the pharmaceutical properties of the poorly water-soluble antiosteoporosis drug 3,9 bis(N,N-dimethylcarbamoy-loxy)-5H-benzofuro[3,2-c]quinoline-6-one (KCA-098) by solid dispersion with hydroxypropylcellulose. AB - The present study was undertaken to improve the oral absorption of KCA-098, an antiosteoporosis drug. In this study, the form 2 of KCA-098 was used as a desirable crystal form for pharmaceutical formation among three kinds of crystal forms, 1, 2, and 3. Solid dispersions of KCA-098 with hydroxypropylcellulose (HPC) or poly(vinylpyrrolidone) (PVP) were prepared by the solvent method. The physicopharmaceutical properties of the solid dispersions were characterized by powder x-ray diffraction, FTIR spectroscopy, and differential scanning calorimetry (DSC). The powder x-ray diffractograms suggest that KCA-098 in the HPC-SL solid dispersion existed in a partial crystalline state as a new crystal form that could be produced by recrystallization from the solvent. Dissolution from the solid dispersions was markedly enhanced in comparison with that of the drug alone. The dissolution enhancement was observed to be greater for the solid dispersion with HPC-SL than for that with PVP. The KCA-098/HPC-SL (1:2) solid dispersion capsule showed a 3.5-fold increase in the initial concentration and 2.5-fold increase in initial concentration of dissolved drug after 60 min, compared with the values for a physical mixture of KCA-098 (form 2)/lactose (1:2). The in vivo absorption of the drug was investigated after oral administration of KCA-098 or its solid dispersion. The area under the plasma concentration curve of KCA-098 after oral administration of the KCA-098/HPC-SL (1:2) solid dispersion capsule was three-fold greater than that for the drug itself. PMID- 11109245 TI - Topical liposomal gel of tretinoin for the treatment of acne: research and clinical implications. AB - An attempt was made to pharmaceutically develop a topical liposomal tretinoin (TRE) gel and clinically evaluate the developed formulation for the treatment of acne in patients. Liposomes of TRE were prepared using the lipid film hydration technique and the entrapment efficiency of TRE in liposomes was optimized to 79.96%. The drug retention in liposomes and in liposomal TRE gel (Carbopol 934 gel base) studied at three storage conditions indicated maximum drug retention at refrigeration temperature. For liposomal TRE gel, reduced drug leakage was observed as compared to that of liposomes at all three storage conditions. Diffusion studies of plain TRE gel and liposomal TRE gel suggested prolongation (3.4 times reduction in flux value) of drug diffusion and almost two-fold increase in skin drug retention after liposomal encapsulation of drug. A comparative double-blind clinical study of the developed liposomal TRE gel, carried out on 30 acne patients over a period of 3 months, demonstrated significant enhancement (about 1.5-fold) in drug efficacy. More remarkable improvement was observed in the treatment of comedones, where the mean percent reduction in lesions increased from 62.36% for plain TRE gel to 94.17% for liposomal TRE gel. Erythema and irritation associated with the use of plain TRE gel was reduced considerably with the use of liposomal TRE gel. The findings of this investigation therefore underscore potential utility of commercialization of liposomal TRE gel in the treatment of acne. PMID- 11109246 TI - Drug release kinetics from polymeric films containing propranolol hydrochloride for transdermal use. AB - Polymeric films containing propranolol hydrochloride (PPN) were formulated and evaluated with a view to select a suitable formulation for the development of transdermal drug delivery systems. Films containing different ratios of ethyl cellulose (EC), poly(vinylpyrrolidone) (PVP), and PPN were prepared by mercury substrate method. In vitro drug release and skin permeation studies were conducted using paddle over disk and modified Franz diffusion cell, respectively. The drug release profiles from the polymeric film indicated that the drug content in the film decreased at an apparent first-order rate, whereas the quantity of drug release was proportional to the square root of time. The release rate of PPN increased linearly with increasing drug concentration and PVP fraction in the film, but was found to be independent of film thickness. The increase in release rate may be due to leaching of hydrophilic fraction of the film former, which resulted in the formation of pores. It was also observed that the release of drug from the films followed the diffusion-controlled model at low drug concentration. A burst effect was observed initially, however, at high drug loading level, which may be due to rapid dissolution of the surface drug followed by the diffusion of the drug through the polymer network in the film. The in vitro skin permeation profiles displayed increased flux values with increase of initial drug concentration in the film, and also with the PVP content. From this study, it is concluded that the films composed of EC/PVP/PPN, 9:1:3, 8:2:2, and 8:2:3, should be selected for the development of transdermal drug delivery systems using a suitable adhesive layer and backing membrane for potential therapeutic applications. PMID- 11109247 TI - Freeze-drying and lyopreservation of diblock and triblock poly(lactic acid) poly(ethylene oxide) (PLA-PEO) copolymer nanoparticles. AB - In this study, the formulation and process parameters that determine successful production and long-term stability of freeze-dried poly(lactic acid) (PLA) nanoparticles with "hairy-like" poly(ethylene oxide) (PEO) surfaces were investigated. Nanoparticles with grafted (covalently bound) PEO coatings were produced by the salting-out method from blends of PLA and PLA-PEO diblock or triblock copolymers. PLA nanoparticles with physically adsorbed PEO were also produced. The redispersibility of the nanoparticles after freeze-drying under various conditions was assessed. The surface of the nanoparticles was characterized and classified in terms of "brush" and "loop" conformations. Upon freeze-drying, it appeared that the presence of PEO at the nanoparticle surface could severely impair the redispersibility of the particles, especially in the PEO-grafted systems. This effect was shown to be related to the amount and molecular weight of PEO in the various formulations. In most cases, particle aggregation was prevented by use of trehalose as lyoprotective agent. Increasing the concentration of particles in the suspension to be freeze-dried was shown to induce much less damage to the nanoparticles, and freezing the suspension at a very low temperature (-196 degrees C) was found to further improve the lyoprotective effect. Most of the lyoprotected nanoparticles remained stable for at least 12 weeks at 4 and -25 degrees C. The production and preservation of freeze-dried PLA-PEO diblock and triblock copolymer nanoparticles is feasible under optimized lyoprotective conditions. PMID- 11109248 TI - The effect of compression and decompression speed on the mechanical strength of compacts. AB - The purpose of this work was to investigate the effect of punch speed on the compaction properties of pharmaceutical powders; in particular, to separate out differences between the effect of the compression and decompression events. Tablets were prepared using an integrated compaction research system. Various "sawtooth" punch profiles were followed to compare the effects of different punch speeds on the crushing strength of the resulting tablets. The loading and unloading speeds were varied independently of one another. In general, when the compression speed was equal to the decompression speed, the tablet crushing strength was observed to decrease as the punch velocity increased. When the compression speed was greater than or less than the decompression speed, the results varied, depending on the material undergoing compaction. Reduction of the unloading speed from 300 to 10 mm/sec for pregelatinized starch and microcrystalline cellulose produced a significant increase in crushing strength, whereas no significant increase in crushing strength was observed until the loading speed was reduced to 10 mm/sec. Reduction of the unloading speed had a similar effect on the direct compression (DC) ibuprofen, however, even greater improvement in the crushing strength was observed when the loading speed was reduced. No improvement in the DC acetaminophen tablets was observed when the unloading speed was reduced, however, a significant increase in crushing strength was produced when the rate of loading was reduced. This work showed that the strength of tablets can be improved and some tableting problems such as capping can be minimized or prevented by modifying the rates of loading/unloading. PMID- 11109249 TI - The influence of varying precompaction and main compaction profile parameters on the mechanical strength of compacts. AB - The purpose of this work was to investigate how altering the method of force application could be beneficial to tablet production in order to increase tablet strength and eliminate or minimize the incidence of capping and lamination. An integrated compaction research system (i.e., compaction simulator) was used throughout this study. Compaction profiles containing a single compaction event and a double (pre- and main) compaction event were created. The ratio and magnitude of the pre- and main compaction pressures were varied and the time interval between the pre- and main compaction events was altered to determine the effects on the crushing strengths and capping tendency of the final compacts. In all cases, for a given pressure, the double compaction event produced stronger tablets than the single compaction event. When the ratio and magnitude of the pre and main compaction pressures were varied, the results differed depending on the material undergoing compaction. Dicalcium phosphate/microcrystalline cellulose and pregelatinized starch tablets had no significant difference in crushing strength values regardless of whether the precompaction pressure was less than or greater than the main compaction pressure. However, both direct compression (DC) acetaminophen and DC ibuprofen were found to have increased crushing strengths and decreased capping/lamination when the precompaction pressure was less than the main compaction pressure. When the time interval between the pre- and main compaction events was varied from 30 to 500 msec, no significant difference in the crushing strength or capping/lamination tendency was observed. It was concluded that the effect of altering the ratio and magnitude of the pre- and main compaction pressures varied from one material to another, suggesting that the profiles should be tailored individually for the specific material undergoing compaction. PMID- 11109250 TI - Isolation technology for research and development applications: from concept to production. AB - A prototype parenteral manufacturing facility based on isolation technology was designed, constructed, and commissioned at Warner-Lambert Co., Morris Plains, NJ, with emphasis on its application to research and development (R&D) settings. The facility contains closed isolators for holding, transferring, and manufacturing sterile products. Vaporized hydrogen peroxide (VHP) was used for sanitization of the isolators. Various factors were evaluated to ensure complete distribution of VHP inside the isolators. VHP sanitization validation of the isolators was performed using chemical and biological indicators, and by swab testing the inside surfaces of the isolators. On the basis of these studies, operating conditions for routine VHP sanitization of the various isolators were established. Performance qualification of the manufacturing facility was conducted via media fills, which demonstrated sterile integrity of the manufacturing process. The media fills revealed certain deficiencies in handling procedures of filled product, which were subsequently corrected. The Warner Lambert isolation technology-based parenteral facility proved to be a reliable and cost-effective alternative to standard clean room technology. The facility is ideally suited for manufacturing small batches. Closed isolator technology has its limitations when used for production-size batches involving automated processing. PMID- 11109251 TI - Influence of emulsion droplet surface charge on indomethacin ocular tissue distribution. AB - The aim of this study was to compare the corneal penetration of indomethacin from Indocollyre [a marketed hydro-poly(ethylene glycol) (PEG) ocular solution] to that of a negatively and a positively charged submicron emulsion. Male albino rabbits were separated randomly into three groups and each group (N = 15) was treated with either one drop of radiolabeled 0.1% Indocollyre, or 0.1% indomethacin positively or negatively charged submicron emulsion, respectively. The rabbits were sacrificed at selected time points and the eyes were enucleated. The eyes were dissected into the different tissues: cornea, conjunctiva, aqueous humor, iris, lens, vitreous, sclera, and retina. The samples were weighed before radioactivity counting. Regardless of the preparation instilled, the highest concentration of indomethacin was achieved in the cornea followed by conjunctiva, sclera retina, and aqueous humor. However, the positively charged emulsion provided significantly higher drug levels than the control solution and negatively charged emulsion only in the aqueous humor and sclera-retina. Furthermore, the spreading coefficient of the positively charged emulsion on cornea is four times higher than that of the negatively charged emulsion. It was therefore deduced that the positively charged submicron emulsions have better wettability properties on the cornea compared to either saline or the negatively charged emulsion. The positive charge may prolong the residence time of the drop on the epithelial layer of the cornea and thus enable better drug penetration through the cornea to the internal tissues of the eye, as confirmed by the animal studies. PMID- 11109252 TI - Effect of sodium dodecyl sulfate on iontophoresis of hydrocortisone across hairless mouse skin. AB - The purpose of the present work was to study the effect of sodium dodecyl sulfate (SDS), an anionic surfactant, on the iontophoretic transport of a neutral drug hydrocortisone (HC) across hairless mouse skin. The transport studies were conducted using Side-Bi-Side diffusion apparatus and drug concentration in the receptor cell was analyzed using reversed-phase HPLC. A theoretical model was described, tested, and found to agree well with experimental data (R2 = 0.9766). Anodal iontophoresis significantly enhanced the transport of HC compared to cathodal iontophoresis and passive diffusion, suggesting that the transport of the neutral solute occurs via the electro-osmotic flow. The effect of SDS on the transport of HC was highly concentration-dependent and driving mode-dependent. Below the critical micelle concentration (cmc), increasing the concentration of SDS increased both the passive and the iontophoretic fluxes of HC, but the increase was most significant with anodal iontophoresis. Above the cmc, passive transport of HC continued to increase with an increase in the SDS concentration. The transport after anodal iontophoresis, however, reached a plateau and then leveled off. Further increase in SDS concentration decreased flux, suggesting that the transport of micellar-solubilized drug is retarded by anodal iontophoresis, possibly due to electrostatic attraction. PMID- 11109253 TI - In situ liposomal preparation containing amphotericin B: related toxicity and tissue disposition studies. AB - The purpose of this research was to develop a novel hydroalcoholic method for the preparation of liposome entrapping inclusion complex of amphotericin B (AmB) with a view to obtaining reduced toxicity and superior tissue distribution of AmB in vivo. The method involves initial preparation of AmB-hydroxypropyl-beta cyclodextrin (AmB-HPBCD) intercalated proliposome which is subsequently converted into a liposome dispersion by single dilution method. The AmB-liposome (L-AmB) derived from proliposome exhibited a superior entrapment efficiency (94.8 +/- 0.7%) compared to liposomes prepared by employing a conventional solvent-based technique (89.7 +/- 2.9%). The dose that was lethal to 50% of subjects (LD50) (mg/kg) value of AmB contained in stabilized proliposome-based liposomes was 18.6 +/- 0.25, whereas that in conventional solvent-based liposomes was 7.8 +/- 0.25. Incorporation of AmB-HPBCD into hydroalcoholic liposomes enhanced antifungal activity in experimental aspergillosis in Balb/c mice in vivo: 80% survival was recorded after 7 days of therapy and the fungal load in lung was reduced. The results clearly demonstrate that preferential uptake of L-AmB entrapped inclusion complex (AmB-HPBCD) by the reticulo endothelial system correlated with diminished levels of AmB in infected (p > 0.05) and noninfected (p > 0.001) kidney after 24 hr compared to that observed with conventional solvent-based L-AmB. Therefore, proliposome-based liposome entrapping inclusion complex of AmB with HPBCD offered an improved therapeutic efficacy by lowering toxicity as well as by altering the tissue distribution pattern. PMID- 11109254 TI - Effect of peptide loading and surfactant concentration on the characteristics of physically crosslinked hydrogel. AB - The purpose of this study was to investigate the effect of varying drug load and concentration of a surfactant (sodium lauryl sulfate [SLS]) on the release characteristics of a model peptide (bovine serum albumin [BSA]), and study the net effects of the swelling properties of the hydrogel matrix [poly(vinyl alcohol) (PVA)]. The PVA hydrogel was prepared by a freeze-thaw process in the absence of a chemical crosslinking agent. The effect of protein loading on drug release was examined at three levels (0.65, 1.3, and 2%), whereas the effect of SLS was studied at four levels (0, 0.07, 0.13, and 0.26%). The baseline time for reaching equilibrium swelling was 48 hr for the hydrogel containing 0.65% BSA, and the equilibrium swelling time decreased significantly as the protein load was increased to 2%. The net effect of increased BSA concentrations resulted in faster BSA dissolution from the hydrogel matrix. The equilibrium-swelling ratio decreased from 21 to 10% when SLS was added to the PVA solution, which resulted in a reduction in the extent of equilibrium swelling; however, the time to reach equilibrium swelling was increased. The investigation provided a mechanistic basis toward the development of a hydrogel formulation by altering the concentration of two fundamental components, i.e., drug and surfactant, within the delivery system. PMID- 11109255 TI - Degradation of a fluoropyridinyl drug in capsule formulation: degradant identification, proposed degradation mechanism, and formulation optimization. AB - The purpose of this paper was to investigate the degradation chemistry of a fluoropyridinyl drug candidate in capsule formulation and to optimize the formulation based on a proposed degradation mechanism. Small developmental batches of capsules were made by tituration of drug substance and excipients using a mortar and pestle, followed by manual encapsulation. Degradants were identified by LC-MS/MS and LC-photodiode array detector (PDA) and were monitored by LC-ultraviolet detector (UVD) during stability studies. It was found that the drug could undergo a nucleophilic substitution reaction in which hydroxyl groups replace the fluorine substituents on the pyridine rings. The initial degradation rate is independent of the drug concentration but dependent on the temperature, the pH of the microenvironment, and the excipient type. On the basis of these experimental results, a nucleophilic substitution reaction mechanism for the degradation was proposed and a successful capsule formulation was developed. PMID- 11109256 TI - Freeze-dried inclusion complexes of tolfenamic acid with beta-cyclodextrins. PMID- 11109257 TI - In vitro determination of disintegration time of quick-dissolve tablets using a new method. PMID- 11109258 TI - General equations for in situ salt screening of multibasic drugs in multiprotic acids. PMID- 11109259 TI - Preparation, characterization, and in vitro evaluation of 1- and 4-month controlled release orntide PLA and PLGA microspheres. AB - PURPOSE: To prepare, characterize and evaluate in vitro sustained delivery formulations for a novel LHRH antagonist, Orntide acetate, using biodegradable microspheres (ms). METHODS: Poly(d,l-lactide) (PLA) and poly(d,l-lactide-co glycolide) (PLGA) were characterized for molecular weight (Mw, Mn) using gel permeation chromatography (GPC) and content of free end carboxyl groups (acid number, AN) by a titration method. 1- and 4-month Orntide ms were prepared by a dispersion/solvent extraction/evaporation process and characterized for drug content (HPLC), bulk density (tapping method), particle size (laser diffraction method), surface morphology (scanning electron microscopy, SEM), and structural integrity of encapsulated peptide by Fourier Transform Matrix Assisted Laser Desorption mass spectrometry (FT-MALDI). Peptide binding to PLA and PLGA and non specific adsorption to blank ms was studied in 0.1 M phosphate buffer pH 7.4 (PB) and 0.1 M acetate buffer pH 4.0 (AB). In vitro release of peptide was assessed in PB and AB. RESULTS: Mw for the PLGA copolymers varied from 10,777 to 31,281 Da and was 9,489 Da for PLA. AN was between 4.60 and 15.1 for the hydrophilic resomers and 0.72 for the hydrophobic 50:50 PLGA copolymer. Spherical ms (3.9 mu to 14 mu in diameter) with mostly nonporous surface and varying degree of internal porosity were prepared. FT-MALDI mass spectra of the extracted peptide showed that the encapsulation process did not alter its chemical structure. Peptide binding to PLGA and PLA and non-specific adsorption to blank PLGA ms were dependent upon pH and were markedly higher in PB than in AB. The initial in vitro release in PB varied from 0.5 to 26%/24 h but due to substantial binding of the peptide to the polymeric matrix the long-term release in PB could not be determined. Application of a dialysis method allowed for a more accurate determination of in vitro release and a good total drug recovery. CONCLUSIONS: Orntide acetate was successfully incorporated into PLA and PLGA ms and the 1- and 4-month in vitro release profiles were achieved by polymer selection and optimization of the manufacturing parameters. PMID- 11109260 TI - Ultrasound and twin-twin transfusion syndrome. AB - This article examines twin-twin transfusion syndrome, a condition in which identical twins share a single placenta with abnormal anastomoses. The syndrome causes unequal blood flow between the twins that, if untreated, is likely to result in the death of one or both. Ultrasound is used to diagnose the syndrome and guide treatment, which may consist of aggressive amniocentesis, endoscopic laser surgery or selective feticide. PMID- 11109261 TI - Seated mammography for older patients. AB - Standing for a mammography exam is difficult for some elderly patients due to dizziness, loss of strength or other age-related changes. This article reports on a pilot study that examined seated mammography as an alternative for older women. Results indicate that seated and standing mammography produced images of comparable quality and patients were equally satisfied with the procedures. The author advocates further investigation with a larger cohort of patients. PMID- 11109262 TI - Principles of cardiac catheterization. AB - Cardiac catheterization, the insertion of catheters into the heart to measure pressures, obtain images and facilitate treatment, is a relatively new procedure and has evolved quickly into a critical diagnostic and therapeutic tool. This article discusses the history of cardiac catheterization, indications and contraindications for its use, catheterization equipment and procedures, patient follow-up and possible complications. PMID- 11109263 TI - Extracorporeal shock wave lithotripsy. AB - Urinary tract stones have plagued mankind since ancient times. Until recently, this painful condition was corrected surgically, sometimes with serious complications. This article discusses extracorporeal shock wave lithotripsy (ESWL), a noninvasive treatment that uses shock waves to fragment stones so they can be flushed out of the body through urination. The author describes how urinary tract stones form, their signs and symptoms, different types of ESWL equipment, how the treatment is performed and the recurrence and prevention of urinary tract stones. PMID- 11109264 TI - Developing an online program. PMID- 11109265 TI - Contrast-enhanced electron beam CT. PMID- 11109266 TI - Clinical experience requirements. PMID- 11109268 TI - Sinus series. PMID- 11109267 TI - A postsurgical puzzle. PMID- 11109269 TI - Positioning nursing research to contribute to the scientific mission of the Pediatric Oncology Cooperative Group. PMID- 11109270 TI - Pediatric oncology nursing in cooperative group clinical trials comes of age. AB - OBJECTIVES: To provide a review of the nursing committees in both the Children's Cancer Group and the Pediatric Oncology Group and discuss intergroup nursing research collaboration in preparation for the merger of these cooperative clinical trials groups. DATA SOURCES: Review articles, reports, and newsletters from cooperative clinical trials groups. CONCLUSIONS: Nurses have established a vital presence in the pediatric cancer cooperative groups over the past 20 years through education, clinical practice, and collaborative research. IMPLICATIONS FOR NURSING PRACTICE: With the unification of the pediatric cooperative groups into the single Children's Oncology Group, the time is uniquely right to build a program of nursing research allied with pediatric oncology cooperative group clinical trials. PMID- 11109271 TI - Fatigue in children and adolescents with cancer: evolution of a program of study. AB - OBJECTIVES: To describe the development of a research program focused on cancer related fatigue in children and adolescents and the resulting definition and model. DATA SOURCES: Research studies, review articles, and clinical examples. CONCLUSIONS: Fatigue has been identified by children and adolescents who are receiving treatment for cancer as one of the most distressing treatment-related symptoms they experience, yet fatigue is rarely assessed by health professionals and infrequently reported by patients or their parents. IMPLICATIONS FOR NURSING PRACTICE: An improved understanding of the contributory and alleviating factors that cause fatigue in this patient population will provide them with greater comfort during treatment for cancer. PMID- 11109272 TI - Cognitive consequences and central nervous system injury following treatment for childhood leukemia. AB - OBJECTIVES: To determine the relationship between membrane damage and intellectual and academic abilities in children with acute lymphoblastic leukemia (ALL) and pilot test a math intervention for children with ALL who were affected. DATA SOURCES: Research studies and review articles. CONCLUSIONS: Despite the prophylactic central nervous system (CNS) treatment for long-term disease-free survival, many children with ALL subsequently experience declines in intellectual and academic skills. IMPLICATIONS FOR NURSING PRACTICE: Improving academic abilities in children who have received CNS treatment is of high priority and may have longlasting implications on quality of life. PMID- 11109273 TI - The PRO-SELF Program: a self-care intervention program for patients receiving cancer treatment. AB - OBJECTIVES: To describe the development, testing, and refinement of the PRO-SELF Program as used in randomized clinical trials. DATA SOURCES: Research studies and articles. CONCLUSIONS: The PRO-SELF Program has made an important contribution in enhancing patients' self-care and reducing morbidity. It has the potential to contribute to self-care abilities of children and adolescents who have cancer. IMPLICATIONS FOR NURSING PRACTICE: It is imperative that patients and their families have the essential information, skills, and support to carry out effective self-care symptom management. PMID- 11109274 TI - Fostering coping by adolescents with newly diagnosed cancer. AB - OBJECTIVES: To review a research program about adolescents' coping with life threatening illness and nurses' role in fostering the adolescents' coping efforts. DATA SOURCES: Research studies, review articles, and book chapters. CONCLUSIONS: Adolescents who have cancer are at risk of negative health outcomes. Promoting positive coping during the developmental period of adolescence helps adolescents to assume greater responsibility for their health actions. IMPLICATIONS FOR NURSING PRACTICE: Certain behaviors initiated by health care providers on hopefulness in adolescents assists the ill adolescents in articulating their hopes and realising or redefining those hopes. PMID- 11109275 TI - [Gastric ulceration in horses: etiology, diagnosis, and therapy: a review]. AB - Since it has become possible to make an ante-mortem diagnosis of gastric ulceration in horses by means of endoscopy, interest in the presence and treatment of this syndrome has increased. Several endoscopic surveys have indicated that the frequency of gastric ulceration in Thoroughbreds in training is fairly high. Less is known about other breeds and horses that are kept under different (training) conditions. The equine stomach is covered by two different kinds of mucous membranes: squamous and glandular. These two areas differ from one another in the incidence and aetiology of ulceration and, therefore, the therapeutic approach is not similar. Clinical signs that typically are associated with gastric ulceration include recurrent colic, poor appetite, weight loss, and, in foals, diarrhoea. Often symptoms are less obvious (such as poor performance), or not even noticeable. Treatment is possible with, for example, H2-antagonists or proton pump inhibitors. Management measurements are important in preventing ulcer recurrence. PMID- 11109276 TI - [Individual behavioral characteristics of pigs and their impact on production]. AB - Two studies were carried out with pigs to determine the relationship between back test results and production parameters and between back test results and other factors. In the first study, 823 piglets were tested with the back test at 10 and 17 days of age. Production parameters such as average daily weight gain and lean meat percentage were determined. In the second study, the back test was performed on 566 piglets at 3, 10, and 17 days of age. The number of escape attempts in the back test (back test score) of the mother was known for 364 piglets. Parameters concerning the health of the sow and piglets were recorded, as well as the sow's reaction to piglet removal for testing. The relationships between production parameters and back test scores of the animals were calculated, as well as the influence of birth weight, sex (all males were castrated), parents, and health parameters on back test scores. Back test scores were fairly consistent over successive tests for each piglet. Males had higher back test scores than females, and piglets from sows with low back test scores also had low scores. Finally, a higher back test score was correlated with a higher lean meat percentage and a better carcass grading at slaughter; no relation with daily weight gain was found. It is concluded that there are individual differences in the way pigs cope with a stressful situation, as measured with the back test, and that this coping behaviour is consistent. A positive correlation exists between back test scores at a young age and lean meat percentage at slaughter. The response to stress, and hence back test scores, is assumed to be inheritable. PMID- 11109277 TI - [Diet food and ectoparasites play an important role in dermatology]. PMID- 11109278 TI - [Proposal to restart infectious bovine rhinotracheitis control]. PMID- 11109279 TI - [BVD control: about persistent shedders and "quiet" carriers]. PMID- 11109280 TI - [Complaints about small animal insurer NHZ insurance administration]. PMID- 11109282 TI - [Risks of the veterinary profession]. PMID- 11109283 TI - [Dendritic cells and their use in the therapy of neoplastic diseases]. AB - Dendritic cells (DC) constitute a heterogeneous leukocyte population. Their main function is to capture and process antigens (Ag) and present them to immunocompetent cells. Heterogeneity of DC is reflected at several levels. Myeloid and lymphoid lineage of the DC can be distinguished according to the precursor cell they originate from. The functional differentiation is of the great importance. DC can induce either specific immune reaction or tolerance to certain Ag. It depends upon the microenvironment where the processing of Ag takes place. Phenotypic and functional differences between the subtypes of DC are being extensively investigated for the purpose of their use in the immunotherapy of various diseases, tumors in particular. It appears that one of the causes of the specific anti-tumor immunity failure is the insufficient function of DC in vivo in patients with malignant diseases. Recent technology advances has enabled to generate and cultivate DC in sufficient amounts in vitro from their precursors. Coculturing of DC with the tumor Ag in the presence of cytokine mixture leads to the efficient Ag presentation and to the generation of specific cytotoxic lymphocytes capable of killing tumor cells. Subsequent application of these tumor Ag pulsed DC to the laboratory animals, and to the patients in first clinical studies, can induce regression of malignant disease. On the other side the ability of DC to induce tolerance to certain Ag is the subject of investigation in the field of immunotherapy of hypersensitivity states induced either by outer Ag (allergy) or inner Ag (autoimmune diseases). In this review we summarize source and ontogeny of DC, their morphology, phenotype, function and different ways of their generation in vitro. We emphasize the use of DC in the clinical practice aimed at the immunotherapy of tumor diseases. PMID- 11109284 TI - [Chronic renal allograft rejection. Part 2. Therapeutic possibilities] . AB - Chronic rejection represents an important cause of renal allograft loss in the long term follow up. New insights into etiopathogenesis of the chronic rejection offer possibilities for experimental therapy. Novel immunosuppressants, such as mycophenolic acid, tacrolimus or rapamycin as well as lipid lowering drugs, angiotensin-converting enzyme inhibitors or AT-1 receptor blockers, may reduce of effects the risk factors on the progression of chronic rejection. In the future, gene therapy may offer additional possibilities to prevent chronic rejection. This review deals with possibilities of prevention of the chronic renal allograft rejection based on experimental evidences and current therapeutic concepts and puts these options into a rational perspective. PMID- 11109285 TI - [Adult height of patients after long-term treatment of idiopathic growth hormone deficiency. (Czech Registry 1991-1998)]. AB - BACKGROUND: Final height was evaluated in patients from the Czech Register with idiopathic growth hormone deficiency treated with GH. METHODS AND RESULTS: 23 patients had the isolated growth hormone deficiency (group 1), and 37 suffered from multiple pituitary hormones deficiencies (group 2). The patients from group 1 and 2 were given growth hormone for 6.7 +/- 2.2 years and 9.6 +/- 3.0 years, respectively. The patients with isolated growth hormone deficiency reached final height -1.4 +/- 1.3 SDS, those with multiple hormone deficiencies were taller ( 0.7 +/- 1.5 SDS). The height gain equalled +2.5 +/- 0.6 SDS and +3.4 +/- 1.1 SDS in group 1 and 2, respectively. 70% of the patients in group 1 and 93% of group 2 reached final heights within the target limits (+/- 2 SDS). CONCLUSION: The final height positively correlated with the target height (mid-parental height) and height at the onset of puberty (group 1 and 2). There was a negative correlation between the final height and the chronological age at the beginning of growth hormone therapy in group 1. PMID- 11109286 TI - [Treatment of iodine deficiency in pregnant women in the Teplice District]. AB - BACKGROUND: Sufficient supply of the population with iodine becomes again a world problem. Particularly the iodine deficiency during pregnancy may have serious negative effects on the foetal development. Estimation of the level of iodine supply of pregnant women became therefore the aim of the study. Investigation was done in the district Teplice. METHODS AND RESULTS: The cohort was formed by 348 pregnant women (age 17 to 45 years), who received during the pregnancy iodine supplementation in the daily dose of 130 respective 100 micrograms (Materna, Jod 100). Control group consisted of 231 women (age 18 to 35 years). Average ioduria of pregnant women at the beginning of gestation (not affected by supplementation) was 127 micrograms I/l, median 115 micrograms I/l of urine. Ioduria of non pregnant women of the control group was 100 micrograms I/l median 84 micrograms I/l of urine. Low ioduria (below 100 micrograms I/l) was found in 40% of pregnant and 62% fertile women of the control group. Supplementation resulted in the statistically significant increase of ioduria in the 20th week (average value was 186 micrograms I/l, median 156 micrograms I/l); low values had found only 20% of women. The fourth day after delivery, ioduria was statistically lower than in the 20th week (121 micrograms I/l, median 99 micrograms I/l), however, 76% of women finished with supplementation after the childbirth. Values of ioduria were still higher in supplemented than in non-supplemented women. CONCLUSIONS: Iodine intake in the group of pregnant and non-pregnant women was not optimal. Even during the recommended iodine supplementation (100 and 130 micrograms), 20% of women had lower values of ioduria. Health education, which first of all involves the risk groups, is therefore highly recommended. PMID- 11109287 TI - [Development of abdominal and gluteal circumference (waist and hips) in adults]. AB - BACKGROUND: Abdominal and gluteal (waist and hip) circumferences are accepted as indicators of central fat distribution and abdominal (waist) circumference is generally regarded as a simple anthropometric measure of visceral fat distribution. METHODS AND RESULTS: In the years 1987-1988 there was carried out an anthropometric survey on the region of former Czechoslovakia. Crossover study was performed in 16,400 of adult men and women aged from 20 to more than 70 years on the basis of the three level random sample. Except for many other anthropometric measures, abdominal and gluteal (waist and hip) circumferences were assessed according to Martin (M62/1; M64/1). The results show that gluteal (hip) circumference is longer than a abdominal (waist) circumference in adulthood similarly as in childhood. Abdominal circumference is larger in men than in women in agreement with sexual dimorphism. Abdominal circumference is not significantly different in Czech and Slovak men and Czech and Slovak women. Nevertheless, abdominal circumference in Slovak women increases to maximum at the beginning of their fifth decade and then decreases concurrently with lowering of BMI. In Czech women increase in abdominal circumference lasts till seventh decade. This difference can result from the earlier involution in Slovak women. Maximal intersexual difference is found at the beginning of the third decade (21-25 years) and at the end of follow-up in both populations. CONCLUSIONS: Gluteal (hip) circumference is not significantly different in men and women to the half of the forth decade in both Czech and Slovak populations. In following decades it is markedly longer in women probably due to the higher accumulation of subcutaneous fat. In the second part of seventh decade in Czech population and about five years earlier in the Slovak group the average circumference in men decreases. This change can be caused by the age involution of adipose tissue. The examined data were constructed into percentile nets of the abdominal and gluteal circumferences for both men and women and Czech and Slovak populations. PMID- 11109288 TI - [Importance of the Genome Project]. AB - Methodological and technological development of the molecular biology during the last decades of the twenties century has shifted our interest from the particular to synthetic problems. Studies has been undertaken both at the level of structure of the individual genetic information and on the level of its function and mutual relation within a single cell or in a multicellular organism. This crucial period culminated in the human genome analysis project, which is now celebrating its first major success. Parallel to it came the analysis of genomes of many other, though usually much thinner organisms. PMID- 11109289 TI - [Resistance of cells in hematopoietic malignancies to cytostatic agents. Part 1]. AB - Resistance to cytotoxic drugs is a serious drawback in the treatment of patients with tumours, both of the haemopoietic and non-haemopoietic origin. The cytotoxic effect of drugs on the malignant cells manifests as the process of apoptosis. In the resistant malignant cells, apoptosis becomes prevented by several mechanisms. The multidrug resistance (MDR) is one of the principal mechanisms when the cancer cells develop resistance to multiple chemically and functionally unrelated cytotoxic compounds. The decrease of the cytotoxic drug concentration at the molecular target site may come from activation of some efflux membrane systems participating in the transport of cytotoxic drugs out of the cell (e.g. Pgp, MDR, and LRP). Another mechanism of resistance is the increased enzymatic detoxification of the drug by glutathion-S-transferase system. Changes in the molecular target of the cytotoxic drug such as topoisomerase molecule can be also responsible for the resistance. At least two additional mechanisms of resistance of tumour cells were identified. Resistance can come from either deregulation of proapoptic mechanisms in tumour cells or by increased activity of reparation processes which control the damaged molecule of DNA. Several methods that detect the cause of resistance in distinct cell populations have been developed. The great effort is now focused on both the detection of mechanisms of the resistance and on the clinical procedures of overcoming the resistance to cytotoxic drugs. PMID- 11109290 TI - [Clinical spectrum of ventricular tachycardias]. AB - Substantial information has been accumulated concerning the mechanisms, electrocardiographic patterns and electrophysiologic characteristics of various ventricular tachycardias. As a result, several distinct clinical entities of ventricular tachycardia can be identified, each of them with different manifestation, prognostic importance and management strategies. Among monomorphic tachycardias, the sustained ventricular tachycardia after myocardial infarction is the most frequent form, accounting for more than 70% of all ventricular tachycardias. Monomorphic ventricular tachycardias are less frequently associated with dilated or hypertrophic cardiomyopathy, arrhythmogenic right ventricular dysplasia and other diseases. Distinct entity that should not go unrecognised is the bundle branch re-entry and/or interfascicular re-entry. Finally, there is a heterogeneous groups of idiopathic ventricular tachycardias occurring in patients without hay structural heart disease. Polymorphic ventricular tachycardias are often associated with either congenital or acquired QT interval prolongation, the so called torsade de pointes arrhythmias. In addition, some polymorphic ventricular tachycardias without QT interval prolongation may be triggered by ischaemia and/or can occur in patients with previous myocardial infarction. The review describes all the most important varieties of monomorphic and polymorphic ventricular tachycardias together with the treatment options. PMID- 11109292 TI - [Trends in publishing of Czech and Slovak medical literature]. AB - Czech and Slovak medical literature (articles in journals and proceedings, book reviews, monographs) is registered in the Bibliographia medica cechoslovaca basis generated by the National Medical Library, Prague. Quantitative trends in the publishing of this literature in the period 1978-1999 are outlined. PMID- 11109293 TI - [A view from the other side of the equator (health care and medicine in Australia)]. PMID- 11109291 TI - [Methods in molecular cytogenetics for determination of prognostically important chromosome changes in patients with chronic lymphatic leukemias]. AB - BACKGROUND: We applied classical cytogenetics, FISH and CGH to investigate prognostic important chromosomal changes, deletions of 17p, 11q and trisomy of chromosome 12 in 90 B-CLL patients at the time of diagnosis. METHODS AND RESULTS: Using classical cytogenetics the chromosomal changes were detected in 17 (18%) patients. Trisomy 12 was found in three patients, deletion 11q in two patients and deletion 17p in four patients. The employment of FISH and CGH revealed chromosomal changes in 52 (58%) patients, the trisomy of chromosome 12 was detected in 12 (13%) patients, the deletions of 11q and deletions of 17p in 10 patients (11%). Statistically significant correlation between higher disease activity and the stage of disease and the presence of deletion 11q and 17p was found. The trisomy of chromosome 12 was found in patients with abnormal markers and in patients with higher stage of the disease. CONCLUSIONS: According to our results, the majority of chromosomal abnormalities in B-CLL may escape detection when classical cytogenetics is the only diagnostic technique used. It stresses the importance of employing additional techniques including FISH and CGH at the time of diagnosis. PMID- 11109294 TI - 23rd annual meeting of the Canadian College of Neuropsychopharmacology and Joint Meeting with the Association francaise de psychiatrie biologique and the Association marocain de psychiatrie biologique. Marrakesh, Morocco, Apr. 10-12, 2000, and Casablanca, Morocco, Apr. 12-14, 2000 PMID- 11109295 TI - Chronobiology and mood disorders: background and introduction. PMID- 11109296 TI - Influence of sleep-wake and circadian rhythm disturbances in psychiatric disorders. AB - Recent evidence shows that the temporal alignment between the sleep-wake cycle and the circadian pacemaker affects self-assessment of mood in healthy subjects. Despite the differences in affective state between healthy subjects and patients with psychiatric disorders, these results have implications for analyzing diurnal variation of mood in unipolar and bipolar affective disorders and sleep disturbances in other major psychiatric conditions such as chronic schizophrenia. In a good proportion of patients with depression, mood often improves over the course of the day; an extension of waking often has an antidepressant effect. Sleep deprivation has been described as a treatment for depression for more than 30 years, and approximately 50% to 60% of patients with depression respond to this approach, especially those patients who report that their mood improves over the course of the day. The mechanisms by which sleep deprivation exerts its antidepressant effects are still controversial, but a reduction in rapid eye movement sleep (REM sleep), sleep pressure and slow-wave sleep (SWS), or a circadian phase disturbance, have been proposed. Although several studies support each of these hypotheses, none is sufficient to explain all observations reported to date. Unfortunately, the disturbed sleep-wake cycle or behavioural activities of depressed patients often explain several of the abnormalities reported in the diurnal rhythms of these patients. Thus, protocols that specifically manipulate the sleep-wake cycle to unmask the expression of the endogenous circadian pacemaker are greatly needed. In chronic schizophrenia, significant disturbances in sleep continuity, REM sleep, and SWS have been consistently reported. These disturbances are different from those observed in depression, especially with regard to REM sleep. Circadian phase abnormalities in schizophrenic patients have also been reported. Future research is expected to clarify the nature of these abnormalities. PMID- 11109297 TI - Premenstrual syndrome and premenstrual dysphoric disorder: guidelines for management. AB - The inclusion of research diagnostic criteria for premenstrual dysphoric disorder (PMDD) in the Diagnostic and Statistical Manual of Mental Disorders, 4th edition, recognizes the fact that some women have extremely distressing emotional and behavioural symptoms premenstrually. PMDD can be differentiated from premenstrual syndrome (PMS), which presents with milder physical symptoms, headache, and more minor mood changes. In addition, PMDD can be differentiated from premenstrual magnification of physical or psychological symptoms of a concurrent psychiatric or medical disorder. As many as 75% of women with regular menstrual cycles experience some symptoms of PMS, according to epidemiologic surveys. PMDD is much less common; it affects only 3% to 8% of women in this group. The etiology of PMDD is largely unknown, but the current consensus is that normal ovarian function (rather than hormone imbalance) is the cyclical trigger for PMDD-related biochemical events within the central nervous system and other target organs. The serotonergic system is in a close reciprocal relation with the gonadal hormones and has been identified as the most plausible target for interventions. Thus, beyond conservative treatment options such as lifestyle and stress management, other non-antidepressant treatments, or the more extreme intervneitons that eliminate ovulation altogether, selective serotonin reuptake inhibitors (SSRIs) are emerging as the most effective treatment option. Results from several randomized, placebo-controlled trials in women with PMDD have clearly demonstrated that SSRIs have excellent efficacy and minimal side effects. More recently, several preliminary studies indicate that intermittent (premenstrual only) treatment with selective SSRIs is equally effective in these women and, thus, may offer an attractive treatment option for a disorder that is itself intermittent. PMID- 11109298 TI - Pathophysiology of seasonal affective disorder: a review. AB - The study of the pathophysiology of seasonal affective disorder (SAD, also known as winter depression) has historically been intimately linked to investigations into the mechanisms of action of light therapy. This paper reviews the studies on the pathophysiology of SAD with emphasis on circadian, neurotransmitter, and genetic hypotheses. There is substantial evidence for circadian phase shift and serotonergic hypotheses, but conflicting results may indicate that SAD is a biologically heterogeneous condition. Recent progress in defining the molecular mechanisms of the human circadian clock and retinal phototransduction of light will provide important new directions for future studies of the etiology and pathophysiology of SAD. PMID- 11109301 TI - Proton magnetic resonance spectroscopy (1H-MRS) of the cerebellum in men with schizophrenia. AB - OBJECTIVE: To investigate whether there are cerebellar vermis abnormalities in schizophrenia. DESIGN: Prospective imaging study with proton magnetic resonance spectroscopy (1H-MRS). SETTING: Schizophrenia clinic at a large urban hospital. PATIENTS AND CONTROLS: Twelve right-handed male patients with schizophrenia, and 12 control subjects with no psychiatric history. INTERVENTIONS: MRS data were acquired from a 2.0 x 2.0 x 2.0 cm volume of interest that included the entire cerebellar vermis. OUTCOME MEASURES: Spectral peak arising from N-acetylaspartate (NAA), phosphocreatine/creatine (Cr) and choline (Cho). RESULTS: There were no significant differences between the patients with schizophrenia and the controls in cerebellar vermis ratios of NAA to Cr (p = 0.71) or Cho to Cr (p = 0.50). CONCLUSIONS: This study does not support earlier structural studies that found abnormalities of the cerebellar vermis in schizophrenia, although it does support reported neurochemical studies. It does not rule out cerebellar involvement in schizophrenia through mechanisms such as aberrant circuitry. Larger in vivo structural/neurochemical and functional imaging studies in other parts of the cerebellum are needed. PMID- 11109300 TI - Psychopharmacotherapy of anorexia nervosa, bulimia nervosa and binge-eating disorder. AB - Pharmacotherapy for anorexia nervosa is considered to be of limited efficacy. However, many studies suffer methodological limitations, and the utility of newer drugs in the treatment of anorexia has not been examined yet. Although there have been more fruitful investigations on the efficacy of medication in the management of bulimia nervosa, there are still many unresolved issues regarding the optimal management of partial remission during the acute treatment phase and the intensity and duration of pharmacotherapy to achieve optimal prophylaxis. Selective serotonin reuptake inhibitors (SSRIs) control the binge urges in binge eating disorder, but more trials are required to investigate the utility of SSRIs and other agents in maintenance treatment. We review the current status of psychopharmacotherapy for anorexia nervosa, bulimia nervosa and binge-eating disorder and evaluate the merits of newer agents in the treatment of these disorders. PMID- 11109302 TI - What is the best approach to management of acute depression in bipolar disorder? PMID- 11109299 TI - Neurochemical and metabolic aspects of antidepressants: an overview. AB - Antidepressants, in addition to being effective therapeutic agents for depression, have also proved to be multifaceted drugs useful for treating a number of other psychiatric and neurologic disorders. Despite the widespread use of these drugs, much remains to be understood about their mechanisms of action and other important aspects, such as their metabolism and potential interactions with other drugs. This article reviews research conducted in the authors' laboratories on various aspects of antidepressants, including trace amines and antidepressants, gamma-aminobutyric acid and antidepressants, drug metabolism, development and application of rapid, sensitive assay procedures for antidepressants and their metabolites; and drug development based on analogues of the antidepressants phenelzine and tranylcypromine. The significance of this work to future drug development is also discussed. PMID- 11109303 TI - E-mail communications with patients. AB - Technological advances have opened the door for the use of e-mail as a viable alternative for communications between physicians and patients. The purpose of this article is to offer recommendations to physicians who are considering communicating with patients via e-mail. PMID- 11109304 TI - Missouri: 29th in nation for quality health care. Results of a national study of fee-for-service Medicare. PMID- 11109305 TI - Operative experiences with thoracoabdominal aortic aneurysms. AB - During a 27-year period, resection of 690 aneurysms of the descending thoracic and/or abdominal aorta were performed. Thirty (4.3%) were thoracoabdominal aneurysms. Although the series of thoracoabdominal aneurysms is small, there was continued improvement in protection of the abdominal viscera and spinal cord from ischemic injury. Operative survivors experienced good late (68% at 5 yrs.) survival. Each of the last 12 pts. in the series survived the operation and 9 are still alive. The surgical results justify a more aggressive stance regarding resection of the thoracoabdominal aneurysms. PMID- 11109306 TI - [The effect of cocarboxylase treatment on erythrocyte transketolase and blood thiamine in patients with end stage renal disease undergoing maintenance hemodialysis]. AB - The depressed ETKA in ESRD patients is supposed to be caused and/or aggravated by several factors among which the diminished content of thiamine in blood and/or disturbances of thiamine utilization seem to play the major role. This role stems from the fact that thiamine acts as the cofactor of transketolase. In order to check the therapeutic significance of this relationship we introduced the thiamine pyrophosphoric acid ester chloride (Cocarboxylasum-CC) administration in 25 patients (mHD + CC). Immediately after each HD performance CC was given i.v. during 12 weeks in a doses of 5 mg/kg b.w., 3 times a week. The blood for ETKA value, free and total thiamine in plasma and erythrocytes, as well as, the total protein and albumins/globulins index investigation was drawn before, after 6 and 12 weeks of CC administration, and 3 months after cessation of this therapy. In 10 patients, on maintenance HD nontreated by CC (mHD), the blood was drawn at the same time intervals. Normal values we obtained from 15 healthy volunteers. For ETKA evaluation photocolorimetric method was used, thiamine content in blood was estimated by fluorimetric method. At the beginning of the study the mean value of ETKA, in two examined groups, was found statistically decreased (p < 0.01) when compared with normals. Mean values of thiamine in plasma and erythrocytes were lower but did not differ significantly from those in normals. After 6 weeks of CC administration ETKA value increased, but only after 12 weeks it increased significantly (p < 0.01), reaching normal value. On the other hand, striking increase in plasma thiamine and erythrocyte thiamine levels was observed after 6 weeks of CC administration already (p < 0.01). Three months after cessation of CC administration significant decrease in ETKA value and thiamine level in blood was observed (p < 0.01). ETKA returned to lower value than in normals even in the presence of still high thiamine levels in blood. In mHD patients nontreated by CC the ETKA value and thiamine levels in blood did not change significantly during all periods of study. The nutritional status assessed by total protein and albumins/globulins index did not change in both groups through the study. We conclude, the administration of high doses of CC to ESRD patients on maintenance hemodialysis HD was successful in terms of increasing ETKA value and thiamine levels in blood without any side effects. Thus, supplementation with large doses of CC deserves further study because it promises to be another adjunct in the treatment of potential thiamine deficiency and metabolic disturbances in the course of dialysotherapy. PMID- 11109307 TI - [Tissue factor and tissue factor pathway inhibitor in peritonitis treated with dialysis]. AB - TFPI-tissue factor pathway inhibitor appears to play a primary role in regulating TF-induced coagulation. During CAPD procoagulant and anticoagulant activities of the mesothelium are balanced under normal conditions. The aim of the work was to assess TF and TFPI concentrations during peritonitis in CAPD patients. The study was performed in 9 CAPD subjects with peritonitis and 14 clinically stable CAPD patients. TF, total, free and truncated TFPI, thrombomodulin concentrations were measured in plasma and dialysate; C-reactive protein and tumor necrosis factor TNF alpha were assayed in serum. In 8 patients with S. aureus peritonitis TF and TFPI were not found in dialysate, but were detectable in a case with E. coli peritonitis. C-reactive protein and TNF alpha significantly elevated at the beginning of peritonitis, fell sharply after the therapy. Further studies are needed to establish whether the kind of bacterial peritonitis (Gram-positive or negative) may affect TF and TFPI in plasma and dialysate in CAPD patients. PMID- 11109308 TI - [Value of magnesium and calcium in serum and hair of children and adolescents with neurologic diseases]. AB - A group of children with neurological diseases was examined, in which in 70-95% (according to age) of children the contents of magnesium in hair and in 39-44% in serum were lowered. The most frequent neurological signs in the relevant group of children were described. A case of a child with hypomagnesemia in whom had been falsely diagnosed and positive therapeutic effects epilepsy were obtained after supplementation with magnesium preparations (Asmag, Magne B6) were presented. PMID- 11109309 TI - [Risk factors for surgical treatment of supratentorial cerebral arterio-venous angiomas]. AB - The authors present the influence of the selected risk factors in surgical treatment of supratentorial cerebral arterio-venous angiomas. The material represents 38 patients treated in the Neurosurgical Department Collegium Medicum of the Jagiellonian University in the last decade. In 28 of the patients the first sign of angioma was subarachnoid haemorrhage (SAH) and the remaining 10 were revealed with seizures. The assessment of the patients when admitted to the Department was graded using the WFNS scale. The diagnosis was established based on clinical symptoms, the results of computerized tomography and cerebral angiography. The extent of the SAH was graded by the Fisher's scale. The detected angiomas were classified according to the Spetzler scale. All patients were operated on using the microsurgical technique. After occluding the feeding arteries, a selective excision was made into the angiomas and the pathological cerebral tissue. The radical of the operation was controlled by cerebral angiography. The results of the surgical treatment were graded by the modified "outcome" Spetzler scale. The following risk factors were analysed: age, WFNS grade, Fisher's grade and Spetzler's grade. The statistical calculations were made using the chi square test and the Mann-Whitney U-test (p < 0.05). In 7 of the patients the results were very good (18.4%), in 12 the results were good (31.6%), and in 14 the results were bad (36.8%). Five of the patients died (13.2%). There was no statistically significant relationship between the risk factors and the results of treatment of supratentorial cerebral arterio-venous angiomas. In the percentage analysis, the best results were obtained from young patients without SAH and with superficial pattern of venous drainage. PMID- 11109310 TI - [Incidence of atrial fibrillation in paced patients with complete atrioventricular block]. AB - The aim of the study was to evaluate the incidence of chronic AF in patients paced due to complete AV block. The study group consisted of 130 pts (70 F, 60 M), mean age 68.6 +/- 14.3 y in whom pacemakers were implanted in the years 1990 1998 due to III degree AV block. There were 76 pts with VVI, 24 DDD and 30 VDD modes of pacing. Follow-up period was mean. 47.6 +/- 25.9 m (7-110). Chronic AF developed in 25 pts (19.2%). In the VVI group the incidence of AF was significantly higher than in DDD and VDD groups--30.3% vs. 4.17% and 3.33% respectively, p < 0.01. Ventriculo-atrial conduction (VAC) was found in 19 pts (14.6%), but did not correlate with the development of AF. Stroke occurred in 2 pts with VVI pacing and chronic AF. In conclusion, in pts with complete AV block VDD and DDD pacing significantly reduce the incidence of chronic AF as compared to VVI. PMID- 11109311 TI - [Evaluation of early cardiovascular involvement in patients with systemic sclerosis]. AB - The aim of the study was to evaluate cardiac function in patients with systemic sclerosis by means of noninvasive methods in order to detect early dysfunction of cardiovascular system. MATERIAL AND METHODS: We studied a group of 19 patients (15 women, 4 men, aged 17-74 yrs, av. 51 +/- 11) with systemic sclerosis comparing the results with a group of 23 healthy volunteers (17 women, 6 men aged 21-69 yrs, av 53 +/- 15). All the patients with SSc were taking corticosteroids, immunosuppressants or vasodilators at the time of the study. In all the patients we performed 24-hour Holter monitoring for the evaluation of arrhythmias, conduction disturbances, ischaemia, heart rate variability (HRV) and late potentials (LP). The following parameters of HRV in time domain were analyzed: SDNN, SDANN, SDNNI, rMSSD, pNN50. Standard ECG was performed to assess QT interval (QT, QTc, QTd). In all the patients the echocardiography examination was performed (M-Mode, 2-D, Doppler echocardiography). The morphology of heart structures and haemodynamic function were analyzed. RESULTS: In patients with SSc Holter monitoring revealed tendency to tachycardia. The mean heart rate was 81 +/ 11 vs. 71 +/- 9 in controls. Conduction disturbances were observed in 3 pts. In 6 pts we found significant ventricular arrhythmia. Silent ischaemia episodes were detected in 6 pts. Concerning HRV analysis the significantly lower values were detected in pts with SSc vs. controls: SDNN 123 vs 170; SDNNI 51 vs 76; SDANN 110 vs 152; rMSSD 29.6 vs 54; pNN50 6.1 vs 21. Late potentials were present in one patient with SSc vs none in the control group. The mean values QT-371, QTc-419, QTd-40- did not exceed the ranges of normal values. No signs of systolic cardiac dysfunction were detected, while in 6 pts we recognized left ventricle diastolic dysfunction. Valvular lesions were observed in 8 pts, but only in 2 pts they were hemodynamically important. CONCLUSIONS: 24-hour Holter monitoring and ECHO examination are valuable methods, which allow to detect early dysfunction of cardiovascular system in patients with systemic scleroderma presenting no apparent cardiac impairment symptoms. PMID- 11109312 TI - Modified dexamethasone and gonadotropin-releasing hormone agonist (Dx-GnRHa) test in the evaluation of androgen source(s) in hirsute women. AB - INTRODUCTION: Polycystic ovary syndrome (PCOS) and congenital adrenal hyperplasia (COH) are heterogeneous disorders, in which excess of androgens may be caused by improper function of ovaries and/or adrenals. In many cases an overlap between ovarian and adrenal type of functional hyperandrogenism has been observed. The relationship between adrenal and ovarian metabolism in hyperandrogenic women is not totally known and etiologic diagnosis of female hyperandrogenism is often difficult. The aim of the present study was to evaluate the usefulness of combined Dexamethasone-Triptoreline testing in distinguishing ovarian and adrenal type of functional hyperandrogenism, and checking if the test could be shortened in order to economise it. MATERIALS AND METHODS: We have examined 57 women with androgen excess divided into two groups: ovarian (n = 42) and adrenal (n = 15) and 20 women with idiopathic hirsutism. There was also one patient suffering from Morris syndrome taken under examination just for curiosity. The blood for hormonal assay was taken in baseline conditions at 8.00 a.m. for LH, FSH, PRL, cortisol, T, DHEAS, 17OHP, E2. Dx was given for 4 days 0.5 mg p.o. every 6 hours. 8 hours after the last Dx administration, the blood was taken for 17OHP and T. Immediately after that Triptorelin 100 mg was given s.c. Then the blood was collected every 4 hours during 24 hours for 17OHP estimation. RESULTS: Decrease in T levels (from 1.65 +/- 0.52 to 0.73 +/- 0.25 ng/ml) after Dexamethasone administration was observed in adrenal group, which indicates adrenal glands as a source of excessive androgen production. No significant differences were seen in ovarian group. But in women from ovarian group supranormal 17OHP response after Triptoreline administration was seen: (ng/ml): at 8.00 am-0.68 +/- 0.44, 12.00- 1.21 +/- 0.7*, 16.00--1.71 +/- 1.19*, 20.00--2.39 +/- 1.81*, 24.00--3.41 +/- 2.64*, 4.00--3.91 +/- 2.82*, 8.00--6.06 +/- 2.43* (*p < 0.01, **p < 0.001). Such a response is typical for women with well defined PCOS and other forms of functional ovarian hyperandrogenism and indicates ovary as a source of androgens. Significant differences were also noticed in idiopathic group: 8.00--0.31 +/- 0.09, 12.00--0.38 +/- 0.17, 16.00--1.41 +/- 0.62*, 20.00--1.52 +/- 0.97*, 24.00- 1.89 +/- 0.83*, 4.00--2.17 +/- 0.83*, 8.00--1.83 +/- 0.71** (*p < 0.01). 17OHP levels did not change significantly during the whole test in adrenal group: 8.00- 1.83 +/- 1.24, 12.00--1.91 +/- 1.37, 16.00--1.95 +/- 0.86, 20.00--2.19 +/- 0.93, 24.00--2.63 +/- 1.58, 4.00--2.56 +/- 1.78, 8.00--237 +/- 0.94. But patients from this group had exaggerated 17OHP response to ACTH (from 4.32 +/- 1.31 to 15.34 +/ 4.1 ng/ml). In patient suffering from Morris syndrome, after Triptoreline, serum 17OHP levels reminded on the same level as they were before drug administration. CONCLUSIONS: Combined Dx-Triptorelin test can be very useful to distinguish ovarian and adrenal type of functional hyperandrogenism. The number of times of blood collection for 17OHP can be reduced to 4 times a day (during 24 hours): at 8.00, 20.00, 24.00, 8.00. PMID- 11109313 TI - [Influence of hormone replacement therapy on the quality of life in postmenopausal women with hypertension]. AB - AIM: The effect of hormone replacement therapy (HRT) on the quality of life in women with hypertension is still not clear. Thus, the aim of the study was to assess the effect of hormone replacement therapy on quality of life in postmenopausal women with essential hypertension by using a battery of standardized questionnaires. MATERIAL AND METHODS: The study population consisted of 53 women (mean age 50.9 +/- 6.3 years) with mild and moderate essential hypertension (mean duration 6.4 +/- 6.4 years). The postmenopausal status was defined as the absence of menstrual blood loss during > 6 months and blood estradiol concentration < 50 pg/ml, accompanied by follicle-stimulating hormone (FSH) levels > 21 U/I. Twenty seven women were blindly randomised to transdermal hormone replacement therapy (HRT) and received 17-beta-estradiol and noretisterone acetate, TTS. Twenty six women were randomly selected as controls. The subjects were evaluated at baseline (after 2 weeks' wash-out from hypotensive drug period) and after three months of HRT using self-administered standardized quality of life questionnaires: the Psychological General Well-being Index (PGWB) and the Subjective Symptoms Assessment Profile (SSA-P). RESULTS: No differences were found in blood pressure values, heart rate, body mass index and distribution of body fat tissue between women receiving HRT and controls at baseline and after 3 months of follow-up. There were no significant differences in the baseline total PGWB score as well as in its subscale between two groups. Similarly, the frequency and intensity of subjective symptoms assessed by SSA-profile were the same in both groups at baseline. After 3 months, a significant improvement in PGWB total score was observed in women receiving HRT. This effect was due to improvement in anxiety, positive well-being and vitality. Moreover, emotional distress, symptoms of flushing, sweating and trembling hands also diminished and sexual capacity improved in women treated with HRT. CONCLUSION: A three-month hormone replacement therapy in hypertensive postmenopausal women slightly improves the general well-being, seems to decrease emotional tension, increase sexual capacity and markedly relieves some vasomotor symptoms. PMID- 11109314 TI - [Knowledge about hypertension and blood pressure level]. AB - BACKGROUND: The improvement of efficacy of the hypertension prevention and patient education largely depend on determination of the hypertension risk factors and increase in knowledge about hypertension. The aim of this study was to assess correlation between the environmental factors and knowledge about hypertension and occasionally measured blood pressure values. MATERIALS AND METHODS: The studied group consisted of 485 subjects, who voluntarily participated in the street-based hypertension screening program in Cracow (summer 1997). All subjects were asked to fill out a questionnaire concerning their health behaviours (i.e. stress, smoking and drinking habits) and a test to evaluate their knowledge about hypertension and its risk factors. The blood pressure measurement were taken using semiautomatic device (Digital Blood Pressure UA-702), in the sitting position, after a rest minimum five minutes. The study group consisted of persons with negative history of hypertension (n = 440), and untreated hypertensive patients (n = 45). In order to assess the influence of different factors on blood pressure level, the subjects were divided into two groups according to the presence or absence of a particular risk factor. In the statistical analysis Student's t-test, chi 2 and linear regression analysis with adjustment for possible confuses were used. All values were presented as mean +/- SD. RESULTS: The mean age was 37.1 +/- 17.8 years. Participants were well educated (75.9% had finished college or high school), and there were more women than men in the study group (57.5% vs 42.5%). 24.1% of participants had blood pressure values exceeding 140/90 mmHg. Multiple linear regression demonstrated that age, body mass index and knowledge about hypertension significantly influenced the level of systolic blood pressure; while only body mass index was among the factors determining diastolic blood pressure. CONCLUSIONS: This study confirms the influence of age, male gender, body mass index, alcohol consumption, stress and the snoring on the blood pressure level. The association between the knowledge about hypertension risk factors or hypertension and occasionally measured blood pressure values has been demonstrated. PMID- 11109315 TI - [The role of statins in prevention of ischemic stroke]. AB - Although associations between cholesterol and coronary heart disease (CHD) are well accepted, the association between cholesterol and stroke remains unclear. Epidemiological studies suggest lack of apparent correlation between cholesterol and cerebrovascular events, however meta-analyses of secondary prevention trials tested statins (HMG-CoA reductase inhibitors) efficacy in reducing cholesterol revealed a powerful statistically significant effect to reduce stroke as well as CHD (32%). Mechanism for stroke reduction can be connected with nonlipid mechanism of statins action: modifying endothelial function and inflammatory responses, plaque stabilisation and inhibition of plaque progression and thrombus formation in the intracranial and extracranial carotid arteries. Stroke events may be also reduced partially as a consequence of CHD reduction. PMID- 11109316 TI - [How efficacy of antianginal drugs should be evaluated]. AB - This article studies the different methods of antianginal drug evaluation in the long-term treatment of ischemic heart disease. Subjective and objective methods are discussed with their limitations. The authors review serial ECG stress testing, this being the most frequently used method of antianginal pharmacotherapy evaluation. Safety, selected topics related to bioethics and statistical analysis in the clinical evaluation of drugs are discussed. Meta analyses and clinical multicenter mega-trials, which are currently gaining in popularity and are likely to play an important role in the complex process of therapeutic decision making in the future, are discussed in the final section of this paper. PMID- 11109317 TI - [Sexual dysfunction during treatment of depression]. AB - Sexual dysfunction in depressed patients is an important clinical problem. Its etiology is complex, however pharmacotherapy is one of the most important factors. It is important for therapists to consider effects of antidepressants on sexual functions in planning treatment strategy. PMID- 11109318 TI - [Microbiological threat from buildings and rooms and its influence on human health (sick building syndrome)]. AB - In buildings we can observe many different strains of bacteria, over 400 species of mould fungi, many strains of fungus causing the rotting of wood and wood like materials, many species of algae, aphids, and other types of growths and seed plants and also over 30 types of mites especially those seen in house dust. Buildings, especially their interiors have a very specific microclimate. Within it areas of so called ecological lows are formed in which conditions for settlement, growth and reproduction of these organisms take place. A building, which is a hazard to the health of its residents, is called a "sick building" from the term "sick building syndrome". The incidence and development of some types of mould fungus is associated with the production of very toxic metabolites which are called secondary metabolites i.e. mycotoxins. Long term human, especially in relation to children, contact with the species producing the most potent mycotoxins like aflatoxin--Apergillus flavus, ochratoxins--Aspergillus ochraceus, rubratoxins--Penicillium rubrum or strachybotrytoxins--Strachybotrys chartarum may even be the cause of death. Mould fungus or just mould is a saprophytic fungus derived from many different systemic groups (Mucor, Aspergillus, Penicillium, Fusarinum). Fungi can produce lethal mycotoxins such as: alternariol, aflatoxins, gliotoxins, ochratoxins, nivalenol, cytinine, dicumarol, rugulosine, trichoviridine and about 200 more which considering their mutagenicity are potentially dangerous to humans, animals, flora and microorganisms. Research which was begun by Prof. Julian Aleksandrowicz and Prof. Boleslaw Smyk in 1970 and 1971 showed that the so called "leukaemia houses" of leukaemia victims had an abundance of toxinogenic fungus in them, particularly the most potent fungus which turned out to be Aspergillus flavus. Toxinogenic funguses are abundant in many living spaces and cellars in older and also in new housing. Mycotoxins have been shown to be very toxic and harmful and it is no wonder that many inhabitants of these living spaces are constantly ill, mainly upper respiratory tract infections, lethargy, constant headaches, nausea and a general ill feeling. Inhabiting these living spaces for a considerable period may lead to cancer. PMID- 11109319 TI - [Tick spirochetosis--Lyme borreliosis]. AB - Lyme boreliosis is currently the most common tick-borne infection. It may cause various clinical symptoms depending on organ localization and duration of the infection. The disease may be symptomless, subclinical or with full clinical manifestation. Usually three clinical stages may be distinguished. In stage I erythema migrans and flu-like symptoms usually develop. In stage II, connected with the infection spreading with blood and lymph, beside joint pains, neuroboreliosis appears, sometimes the disease involves other organs such as heart, eyes, testicles, joints. Stage III, chronic in its character, usually develops in patients who had previously reported joint and neurological complaints. Encephalopathy and fibromyalgia accompany joint involvement. Diagnostics of Lyme borreliosis is based on clinical evaluation and laboratory test including culture of the bacteria obtained from biopsies and serological tests. There are no established standards of the treatment--some examples of the therapy are presented in the paper. The disease if not treated has a progressive course in most causes, however in some patients it can resolve spontaneously even with no treatment. PMID- 11109320 TI - [Use of bioimpedance spectroscopy techniques for monitoring fluid balance in patients with end stage renal failure]. AB - Adequate body hydration is considered the key element of fluid management in critically ill patients including group with end stage renal failure. Bioimpedance technique is widely used as a non-invasive, simple and accurate method to measure body composition. The purpose of the paper was to prescribe the using of single, and multifrequency bioimpedance spectroscopy technique for estimation of fluid balance in end-stage renal patients. We also discussed some measurements and data modelling problems, including postural change effect, and intercompartmental fluid shift during dynamic monitoring of fluid balance. PMID- 11109321 TI - [Is it possible to recognize complicated heart sarcoidosis?]. AB - Authors present 3 cases of pulmonary sarcoidosis coexisting with disturbances in ecg. Cardiac scintigraphy showed segmental disorders of perfusion in those patients. We believe that morphological diagnosis of sarcoidosis and coexistent disturbances in ecg are indications to perform cardiac perfusion scintigraphy. Changes in cardiac perfusion can also suggest the presence of sarcoid granulomas in the heart muscle. PMID- 11109322 TI - [Meningitis with acute respiratory distress syndrome in adults and jaundice--case report]. AB - The case of leptospirosis with adult respiratory distress syndrome (ARDS) and severe jaundice in young abattoir worker has been presented. Institution of invasive therapeutic methods: continuous mandatory ventilation (CMV) with positive end-expiratory pressure (PEEP) and plasmapheresis, together with antibioticotherapy resulted in complete recovery. We discuss some characteristic features of the leptospirosis, which with the data from epidemiological history can lead to early diagnosis of this forgotten disease. PMID- 11109323 TI - [A severe adverse event after vaccine for diphtheria, tetanus, pertussis and poliomyelitis (DTP + polio) in a 4.5 month old infant]. AB - Neurological complications of immunisation are rare. We report the case of neuroallergic reaction 5 days after receiving the second course of DTP + Polio vaccine in a 4.5 month old girl. The pathological signs developed 48 hours after the vaccination. On admission the infant demonstrated a persistent fever (39 degrees C), resistant to Paracetamol, unusual crying, irritability, consciousness disorder, and opistotonus. Physical examination showed generalised mild, macular rash and redness and swelling at the injection site. The neurological assessment revealed weakness of upper and lower limbs, meningeal signs such as stiff neck, Kernigs and Brudzinski signs. There were no pathological changes in the CSF. The diagnosis was established as the neuro-allergic Adverse Event Following DPT + Polio Immunisation. After 10 day treatment with Dexamethasone and Phenobarbital the infant was discharged with no neurological changes. A psychomotorical development is good. PMID- 11109324 TI - [Diagnosis and therapy of thoracic and abdominal injuries in the Czech Republic]. PMID- 11109326 TI - [Mediastinal tumor--a combined cervical and thoracic surgical approach]. AB - In the submitted case-history the authors describe a tumor which developed paravertebrally in the upper three intercostal spaces. During its removal a not very well known surgical procedure was used permitting subluxation of the clavicle. PMID- 11109325 TI - [Repair of traumatic lesions in peripheral nerves--initial experience with laser anastomosis]. AB - Experience with conventional repair of peripheral nerves clearly indicates that there is still room for improvement, mainly in so far as functional results are concerned. It appears that the present microtechnique has reached its limitation given by physical and technological means, materials and methodology. The use of CO2 laser was tried on rat sciatic nerve. An equal number of nerves were severed surgically and sutured, using microsuture technique in one group and the same number of nerves were repaired using the CO2 laser technique. For the setting of the parameters, optimal for our purpose of reuniting the tissues, we have utilised morphological and electrophysiological qualitative and quantitative characteristics. The results were assessed by comparing both functionally and morphologically, the functional comparison being made electrophysiologically and the morphological comparison microscopically and morphometrically. The comparison shows a larger degree of scarring and constriction of the nerves anastomosed by classical microsuture than that of the laser technique. Action potentials going through the anastomosis have been noticed in both groups without significant difference in quality of CMAP, with the exception the latency. PMID- 11109327 TI - [Large volume tumors with uncertain biological activity located in the pleural cavity]. AB - The authors discuss specific problems of an exact diagnosis and indication for surgical treatment in patients with large tumours in the pleural space. Our series includes even some rare tumours of varied histogenesis. The biological activity of those tumours is frequently uncertain and causes an unpredictable prognosis. The prognostic conclusion is based on a bioptic diagnosis, established by histological examination. Complete extirpation is essential. PMID- 11109328 TI - [Streptococcus pneumoniae--an infectious agent in coxitis]. AB - Based on their own observation the authors describe pneumococcal coxitis in a young man. With regard to the atypical clinical picture of the disease the authors draw attention to the necessity to consider in the differential diagnosis of pain in the inguinal and coxal area also infectious arthritis. This concerns in the first place physicians in the first line of contact. PMID- 11109329 TI - [Testing of implants used in the correction of abdominal wall defects in hernia surgery]. AB - The aim of this study is experimental testing of various kinds of meshes. They are used for correction of abdominal wall defects during operations of hernias (especially by the laparoscopic technique). The following implants were tested: Goretex, Prolen, Mersilen and Vicryl. Each of them represents one group of artificial materials (nonabsorbable, monofilaments multifilament, absorbable) which are usually used. Semiabsorbable materials were not tested because they are not usually used. The objects of testing were firstly constant quality of implants lasting in the time and secondly the reaction of tissues after implantation of those materials. PMID- 11109330 TI - [Desmoid]. AB - The author presents the case-record of a patient with familial adenomatous polyposis and a subsequent large desmoid of the abdominal wall, a review of the symptomatology, and treatment. PMID- 11109331 TI - [The Jurasz surgical procedure in an unusual situation]. AB - The authors present an account on the use of Jurasz' operation for drainage of an abscess of the omental bursa after a piercing injury of the abdomen within a relatively short interval after development of this complication. PMID- 11109332 TI - [Late postoperative colonic stenosis caused by the biofragmentable anastomosis ring (BAR)]. AB - The authors focused their attention on the prevalence of late postoperative stenoses of anastomoses following resection of the colon. In 42 patients the anastomosis was implemented by means of a Valtrac ring--i.e. by compressive biofragmentable seamless technique. Within an interval of 6 months after surgery the majority of these patients were subjected to colonoscopic examination with direct visualization of the intestinal anastomosis. The authors did not record any late postoperative stenoses of the BAR anastomosis of the colon. Moreover on colonoscopic examination in the majority of patients the original intestinal anastomosis was only difficult to detect. PMID- 11109333 TI - [Ruptured abdominal aortic aneurysms]. AB - The authors present the results of a retrospective study concerning 96 patients with an abdominal aortic aneurysm rupture who underwent surgery in the 2nd Department of Surgery in Brno in the time period from 1989 to 1999. In 30 cases (31.2%) there was an isolated retroperitoneal rupture, 66 patients (68.8%) also had haemoperitoneum. The mean age of the patients was 74 years and the male/female ratio was 76 (79.2%): 20 (20.8%). Twelve patients (40%) did not survive the retroperitoneal rupture, and 34 patients (51.5%) did not survive the intraabdominal rupture. Total mortality in the study group was 47.9%. The type of the rupture and the size of the haematoma that is in correlation with the time of diagnosis and operation, are among the most decisive prognostic factors. Patients with a small haematoma show higher survival rates. In large retroperitoneal haematomas accompanied by a haemorrhagic shock, the mortality was about 40% in the presented study group, in intraabdominal ruptures the mortality rate increases to values over 50%. The results are in accordance with foreign literature data, and the authors agree with the opinion that haemodynamically unstable patients with suspected abdominal aortic aneurysm rupture should be, after necessary preparations, urgently operated without complementary examinations. PMID- 11109334 TI - [Treatment of occlusions of deep femoral artery prostheses with a venous patch: the Bartos method of reoperation]. AB - The author deals with the approach to a thrombosis of an aorto-femoral prothesis. The principle of the operation, described by the author as Bartos reoperation involves retrograde desobliteration of the prothesis and open desobliteration of the a. profunda femoris with a venous patch. It is implemented from one incision in the groin. The procedure is simple and feasible even in the majority of risk patients. PMID- 11109335 TI - [Use of the internal jugular vein as the outflow tract in arteriovenous shunts for hemodialysis] . AB - In the presence of a long stenosis or an occlusion in the axillo-subclavian venous tract, the vena jugularis interna can be useful as an outflow tract in arteriovenous shunts for haemodialysis. The use of this vein was indicated for primary shunts with vascular grafts and also for treatment of venous hypertension of the upper limb. In both these indications a 6 mm ePTFE (GORE) prosthesis was used. Regular follow-up by Doppler ultrasound was useful for diagnosis of a developing of stenosis in these shunts. With early treatment of these stenoses the long-term function of these shunts could be ensured. PMID- 11109336 TI - [Endoscopic stricturotomy--possible treatment of stenoses of colorectal anastomoses based on a stapling technique]. AB - The authors present the possibility of using colonoscopic stricturotomy by means of a papillotomy commonly used for sphinctertomy of the Papilla Vateri in order to influence a stenosis of a colorectal anastomosis implemented by the stapler technique. The operation proper and postoperative course were without any complications. PMID- 11109337 TI - [Adenocarcinoma of the appendix--case report]. AB - The authors describe a case with a rare diagnosis of adenocarcinoma of the appendix in an adult female patient. The patient was indicated for surgical revision on account of acute appendicitis with a peroperative finding suspect of malignity of the vermiform appendix. A radical resection of the ileocoecal area was performed along with part of the ascendant colon and appropriate mesocolon as a primary operation. According to the postoperative histological evaluation of the resected portion the diagnosis of adenocarcinoma of the appendix was confirmed. PMID- 11109338 TI - Bowel habits--normal or functional? PMID- 11109339 TI - Normal bowel habits and prevalence of functional bowel disorders in Singaporean adults--findings from a community based study in Bishan. Community Medicine GI Study Group. AB - BACKGROUND/AIM OF STUDY: Data on the epidemiology of bowel frequency and functional bowel disorders in the East are limited. The aims of this study were to determine the most common bowel frequency and the prevalence of functional bowel disorders in Singaporean adults. METHODS: A cross sectional study, using a reliable and valid questionnaire was carried out in a random sample of residents aged 16 years and above in Bishan, 68% responded (n = 271). RESULTS: The most common (59.0 +/- 6.5%) bowel frequency was once a day with 96.8 +/- 5.6% of individuals having bowel frequency between 3 times/week and 3 times/day. The prevalences of irritable bowel syndrome, chronic constipation and chronic diarrhoea were 3.2 +/- 2.3%, 7.3 +/- 3.5% and 6.9 +/- 3.4%, respectively. Women were found to have a lower bowel frequency (p < 0.001) and a higher prevalence of chronic constipation (11.3 +/- 6.0% vs. 3.6 + 3.5%, p < 0.05) than men. CONCLUSIONS: Normal bowel frequency may be defined as bowel movements between 3 times per week and three times per day. The prevalence of irritable bowel syndrome in the general population of Singapore was low compared with those reported in the West. PMID- 11109340 TI - Relation between morbidity and current treatment in patients who present with acute asthma to polyclinics. AB - BACKGROUND: It has been suggested that resources for asthma intervention should be focused mainly on patients in the community who experience a high burden of disease. These are who patients who have acute exacerbations which require urgent treatment. AIM: To assess the morbidity and identify deficiencies in the treatment of patients who present for urgent treatment of acute exacerbations to primary care clinics. PATIENTS: Adult patients who received urgent treatment for acute exacerbation of bronchial asthma SETTING: 4 primary care polyclinics. METHODS: A cross-sectional survey of consecutive patients which related regular preventive treatment to current asthma activity. Poor asthma control was defined as step 2 or higher (American National Asthma Education and Prevention Program, report II, 1997) or > or = 2 emergency room visits in 6 months. RESULTS: There were 116 patients of whom 53% were women. The mean (SD) age was 45(15) years and duration of current exacerbation 3 (3) days. The acute symptoms were successfully treated in 93% of patients. Quick relief medication was used regularly in 91% and inhaled corticosteroids (ICS) in 55%. Oral salbutamol was prescribed in 14% of patients. The asthma was poorly controlled in 54%. In the poorly controlled group 33% were not on regular ICS treatment and 64% were not receiving "add on" medication. CONCLUSIONS: Patients treated for acute asthma in primary care clinics: (1) were older and had less acutely severe exacerbations than those who presented to emergency rooms, (2) over half had poorly controlled asthma and (3) a third of patients with poor asthma control were inadequately treated. PMID- 11109342 TI - Relation between grip strength and hand bone mineral density in healthy women aged 30-70. AB - AIM OF STUDY: To determine the site-specific relationship between grip strength and hand bone mineral density (BMD) measured with dual energy x-ray absorptiometry (DXA) in healthy women. The correlation of hand BMD and BMD at axial sites has also been assessed. METHOD: Twenty-nine healthy housewives, aged 30-70, were included in the study. Women were grouped according to their menopausal status (12 premenopausal and 17 postmenopausal). Grip strength of the dominant hand was measured using a Jamar dynamometer. BMD of the antero-posterior spine, femoral neck, trochanter, and Ward's triangle were measured with DXA. For the hand BMD measurements, the analysis software, which was modified from the software of small animals and developed for hand BMD measurements, was used. RESULTS: Hand BMD moderately correlated with grip strength in postmenopausal women. There was no significant correlation between grip strength and hand BMD in premenopausal women. Significant positive correlations were determined between the hand BMD and BMD at axial sites. CONCLUSION: Grip strength may be an independent indicator of hand BMD in postmenopausal women, and also a site specific relationship. Hand BMD measurements may indirectly reflect the BMD of axial sites especially in postmenopausal women. PMID- 11109341 TI - Evaluation of new WHO diagnostic criteria for diabetes on the prevalence of abnormal glucose tolerance in a heterogeneous Nepali population--the implications of measuring glycated hemoglobin. AB - AIM OF THE STUDY: The present study was designed to (a) evaluate the implications of revised WHO diagnostic criteria on prevalence of abnormal glucose tolerance, (b) compare glycated hemoglobin level amongst healthy, impaired glucose tolerance (IGT), impaired fasting glucose (IFG) and diabetic subjects and (c) evaluate the assay of glycated hemoglobin in screening IGT, IFG from normal subjects. METHODOLOGY: Hospital based, cross-sectional study. Plasma glucose and glycated hemoglobin (gHb) were estimated by glucose oxidase and affinity chromatography method respectively. RESULTS: The crude prevalence of IFG, IGT and diabetes were 9%, 18% and 5.29% respectively with no significant difference between Mongol and non-Mongol population. Newly introduced IFG group falsely incorporate 12% diabetic subjects and fails to detect 83% IGT subjects as impaired glucose metabolism. The gHb level is raised in IGT and diabetic group but not in IFG group. CONCLUSION: The assay of gHb may be used to screen abnormal glucose tolerance but paired estimation of fasting glucose increases the reliability of diagnosis. The level of gHb in mild carbohydrate intolerance mostly depend on the level of rise in post prandial glucose (where the variation is wide, as in IGT) but not on the narrow variance in fasting plasma glucose level as found in IFG. PMID- 11109343 TI - Vision screening of 4-year-old children in Singapore. AB - AIM OF THE STUDY: To evaluate the feasibility of an improved visual acuity screening program for Singapore 4-year-old preschool children and to draw up an appropriate referral criteria as well as evaluating the rates and outcomes of these referrals. METHOD: A total of 450 children aged 4 to 4 1/2 years, who attended 3 polyclinics of the Family Health Service (FHS) for their 4-year-old Developmental Health Screening during the study period from 1/4/1997 to 30/6/1997 were recruited for the study. Children who were tested with Snellen (or Sloan) visual acuity chart resulting in visual acuity of 6/9 or worse, or failed to pass the 3 mm medium plate at 30 cm distance (300 seconds of arc) in the Frisby Stereotest, or were found to have strabismus, or were untestable in either visual acuity test or stereotest were offered referral to ophthalmologists in the hospitals for specialist assessment. RESULT: 82.7% of the 450 children were successfully screened with Snellen (or Sloan) chart while 91.6% were successfully screened with Frisby Stereotest. In all, 180 children were evaluated by ophthalmologists. Majority of the children were referred because of their abnormal visual acuity test while only 2 children were referred for failing stereotest alone. Among the 180 children referred, 63 (35.0%) were found to have refractive errors for which spectacles were prescribed. Eight children had amblyopia and 2 children had strabismus which were not detected at the polyclinic screening. The untestable children evaluated had significantly higher abnormality rate (37.5%) than that of children who had 6/9 vision (8.8%) therefore they should be offered referral for further evaluation. There was high "refused referral" rate of 39.0%. Parents of children who were untestable or had 6/9 vision were found to be more likely to refuse offer of referral. If these two groups of children were excluded, the "refused referral" rate dropped to 13.3%. When the referral criteria for visual acuity was reset at 6/12 instead of 6/9, the referral rate dropped from 39.6% to a more manageable 26.7% and the positive predictive value improved from 35.4% to 48.3% and none of the children with amblyopia were missed being screened-out. CONCLUSION: The study confirmed the feasibility of doing visual acuity screening at 4 to 41/2 year-old. The referral criteria for abnormal visual acuity should be set at 6/12. The efficacy of adding Frisby stereotest needs further evaluation. PMID- 11109344 TI - Bone marrow and peripheral blood changes in non-Hodgkin's lymphoma. AB - BACKGROUND: 47 patients with non-Hodgkin's lymphoma (NHL) were studied retrospectively to determine their marrow and blood changes at diagnosis. METHODS: The blood counts, blood films, marrow smears, trephine and tissue biopsies of patients at diagnosis were reviewed. The scheme proposed by the International Lymphoma Study Group (REAL) was utilised for lymphoma subclassification. RESULTS AND CONCLUSION: 21.3% had lymphoblastic lymphoma, 21.3% had peripheral T-cell lymphoma (unspecified), 29.8% had diffuse large B cell NHL, 10.6% had Burkitt's lymphoma and 17% had others. The incidences of anaemia, one or more abnormal counts, lymphocytopenia, increased marrow reticulin and marrow eosinophilia at diagnosis were 66%, 85.1%, 41.3%, 40.9% and 44.7% respectively. Marrow involvement was present in 46.8% of the patients, with diffuse infiltration noted in 71.4% of these cases. Abnormal counts and anaemia were common in all the NHL subtypes. In lymphoblastic lymphoma, the common haematological abnormalities were peripheral atypical lymphocytes and diffuse marrow involvement. In peripheral T-cell lymphoma (unspecified), common features were peripheral lymphocytopenia, increased marrow reticulin and eosinophilia. In diffuse large B-cell NHL, peripheral lymphocytopenia, peripheral myeloid precursors and/or nucleated red cells and marrow involvement were common. In Burkitt's lymphoma, diffuse marrow involvement and eosinophilia were common. No significant differences were noted between most of the haematological parameters of B and T-NHLs. In comparison with other reports, we recorded higher overall incidences of anaemia and diffuse marrow involvement, and a lower incidence of marrow infiltration in peripheral T-cell lymphoma (unspecified). PMID- 11109345 TI - Percutaneous epididymal sperm aspiration in a man with congenital bilateral absence of the vas deferens undergoing an assisted reproduction program. AB - Surgical retrieval of spermatozoa for in-vitro fertilisation (IVF) programs for severely oligospermic men has been in use for several years now. In the recent 2 to 3 years, clinicians have begun to move towards non-surgical methods of retrieving sperm in certain selected groups of men. Percutaneous epididymal sperm aspiration (PESA) has had good results in terms of number of sperm obtained, as well as the fertilisation and pregnancy rates. The first reported use of such a technique in Singapore is described. PMID- 11109346 TI - Myelodysplastic syndrome with monosomy 7 and pulmonary aspergillosis. AB - A 43-year-old man with no past history presented with symptoms of fever, cough and dyspnoea arising from invasive pulmonary aspergillosis and was found to have myelodysplastic syndrome with monosomy 7. Before initiation of chemotherapy, he deteriorated rapidly, developing multi-organ failure requiring mechanical ventilation, and he eventually succumbed despite amphotericin B treatment. The importance of monosomy 7 in determining immune function in patients with myelodysplastic syndrome is emphasised. PMID- 11109347 TI - Porphyria cutanea tarda. AB - Porphyrias are a group of rare metabolic disorders in which excessive quantities of porphyrins, or their precursors, are produced. They are due to specific enzyme deficiencies resulting in abnormalities in the control of the porphyrin-haem metabolic pathway. Porphyria cutanea tarda (PCT) is the most common of all the porphyrias. However this condition is rarely seen in our Asian countries. We describe a patient with PCT who presented clinically with blistering eruptions over the sun-exposed areas. Coral pink fluorescence of uroporphyrins in an acidified urine specimen is diagnostic. Definitive treatment involves the use of low-dose chloroquine and interval venesection. PMID- 11109348 TI - Diabetic retinopathy and serum lipids. AB - The association between serum lipid levels and diabetic retinopathy has been investigated in many studies. Some studies show a positive relationship between serum cholesterol and low-density lipoprotein levels and retinal hard exudation. Other studies show serum triglyceride levels as being important in the progression of retinopathy. Certain other studies show no relationship between serum lipid levels and diabetic retinopathy. We review the literature on this subject and illustrate this report with an example of a diabetic with severe diabetic maculopathy and high serum lipid levels. PMID- 11109349 TI - The changing pattern of cytomegalovirus retinitis in human immunodeficiency virus disease. AB - There have been profound changes in the pattern of cytomegalovirus (CMV) retinitis over the last two decades. The epidemiology and behaviour of CMV retinitis has been significantly altered by Acquired Immune Deficiency Syndrome (AIDS). It was uncommon prior to the AIDS epidemic, but soon became the most common retinal infection in AIDS patients. In the past several years, highly active anti-retroviral treatment (HAART) has achieved a dramatic improvement in the prognosis for patients infected with human immunodeficiency virus (HIV). As a result, HIV patients are living longer and have a reduced risk of CMV retinitis. Some patients with CMV retinitis who respond to HAART develop a transient symptomatic vitritis while others undergo no reactivation of their retinitis despite having no specific anti-CMV therapy. This pattern is likely to undergo further change as the treatment of HIV and CMV disease continues to improve. PMID- 11109350 TI - Clinics in diagnostic imaging (50). Leiomyosarcoma of bladder. AB - A 39-year-old man presented with intermittent painless macroscopic haematuria. He was subsequently diagnosed to have a high grade leiomyosarcoma of the bladder. He underwent total cystoprostatectomy and radiotherapy but died of metastatic disease five months after the initial diagnosis. Aspects of the imaging and diagnostic pathway of painless haematuria are discussed. The management of leiomyosarcomas in the urinary system is highlighted. PMID- 11109351 TI - [Openness when it comes to medical errors]. PMID- 11109352 TI - [Is the age of general practice over?]. PMID- 11109353 TI - [Evidence-based medicine or clinical pragmatism?]. PMID- 11109354 TI - [Nursing homes]. PMID- 11109355 TI - [Mental impairment in nursing home residents]. AB - BACKGROUND: Nursing homes are the corner stones in long-term institutional care for frail elderly people. Expansion and strengthening of community-based services during the last decade has enabled frail elderly who earlier would have moved into a nursing home to stay in their own home or in service apartments. MATERIAL AND METHODS: In a cross-sectional, anonymised study, changes in nursing homes in the city of Bergen between 1985 and 1996/97 were analysed with regard to prevalence of mentally impaired residents. The following items were assessed by nurses who were in daily contact with the residents: mental capacity by means of Clinical Dementia Rating (CDR), activities of daily living (ADL), and behaviour. The study population consisted of 1,296 residents from 21 institutions in 1985 and 1,141 residents from 16 institutions i 1996/97. RESULTS: The percentage of mentally impaired residents increased significantly, from 75 to 82, and the percentage of residents who were in need of extended nursing care rose from 87 to 91. The median duration of stay did not change significantly, but was reduced for residents with slight mental impairments (CDR 1), from 871 to 721 days. INTERPRETATION: Today, predominately mentally impaired and physically disabled elderly are cared for in general nursing homes. These are patients in need of continuous medical care and supervision, and do not fit into an ideology which "demedicalizes" severe medical conditions in old people. Both nursing homes and sheltered livings are needed. PMID- 11109356 TI - [Are old patients able to get up from the floor?]. AB - BACKGROUND: Inability to get up from the floor after an accidental fall is a frightening and potentially dangerous experience. Especially elderly individuals living alone are exposed to this. MATERIAL AND METHODS: In a geriatric five-day ward 206 patients living alone were tested for their ability to rise from the floor. This was carried out under safe conditions in a training room. Those who failed were offered instruction and training given by a physiotherapist, up to five sessions. The training programme is described. RESULTS: 154 patients (75%) were able to get up unaided within two minutes. The time needed varied considerably. Of the 52 who failed 39 accepted training, and 22 finally managed to get up independently. INTERPRETATION: Teaching elderly patients living alone how to get up from the floor is often successful and will probably reduce the risk of being unable to get up for an extended period of time. PMID- 11109357 TI - [Benzodiazepines--quality assurance of prescriptions in own general practice]. AB - BACKGROUND: Even if benzodiazepines are recommended for short-term use, they are prescribed for years. Doctors often feel uncomfortable about this. MATERIAL AND METHODS: In this audit the prescription of benzodiazepines by one general practitioner was studied on the basis of the medical records, a mailed questionnaire to patients, and special benzodiazepine consultations which included diagnostic assessment. 17 long-term benzodiazepine users were given a follow-up which included general information, personal advice and alternative treatment aimed at discontinuing their use of benzodiazepines. RESULTS: 76 out of 754 patients had received prescriptions for a benzodiazepine during the previous three years. The questionnaire was completed by 63 patients. The results revealed that the treatment had originally been initiated by a general practitioner in 78% of the cases, most commonly for anxiety (40%), sleeping difficulties (23%), or depression (10%). Their doctor had previously suggested benzodiazepine withdrawal in 46% of the patients. One third were long-term users. 46% of these had used this medication more than ten years; 90% suffered from chronic illness. After 17 months, 47% of the patients had stopped using benzodiazepines while another 29% used significantly less than before. INTERPRETATION: A broad and individually tailored intervention towards patients in general practice may be a useful tool in reducing long-term use of benzodiazepines. PMID- 11109358 TI - [Legionella pneumonia--important differential diagnosis in pneumonia after travelling abroad]. AB - BACKGROUND: Legionella pneumophila is a rare cause of pneumonia in Norway. From 1992 to 1999, only 27 cases were reported to the Norwegian Surveillance System for Communicable Diseases. MATERIAL AND METHODS: Five cases diagnosed at the Akershus Central Hospital over the last three years are presented. RESULTS: All patients acquired their infection while travelling abroad, mainly in the Mediterranean area, and all fell ill within fourteen days of returning home. The course was serious with marked hypoxaemia in three of the patients, and one patient died. Confusion and altered mental state were prominent features of the clinical presentation. INTERPRETATION: Patients returning from visits abroad and presenting with a pneumonia within to weeks after arriving in Norway, should be investigated for Legionella as a causative agent unless there is a prompt response to empiric penicillin therapy. PMID- 11109359 TI - [What does affect the general practitioners' choice of contract and plans to relocate?]. AB - BACKGROUND: The aim of this study was to examine general practitioners' choice of contract and location in Norway. GPs can choose between two types of contract: a contract by which they are paid a salary, and a contract by which they are paid on a fee-for-service basis plus a fixed grant. METHOD: The data were collected by a questionnaire sent to a representative sample of GPs in Norway (N = 1,639). RESULTS: Salaried physicians and contract physicians show different characteristics. Salaried physicians tend to be younger than contract physicians and to prefer leisure to higher income. Most salaried physicians were located in rural areas. The following tendencies were observed with respect to location: GPs wanted to move from rural to central areas. Physicians who reported that their workload was too high, wanted to move to an area where the workload was lower. Physicians who reported that they had too few patients did not want to move. Physicians who were often on duty to provide emergency services wanted to move. INTERPRETATION: According to standard market theory, physicians are expected to move to areas where demand is high when demand in their own areas falls. Our results indicate that public regulation is necessary in order to obtain an optimal distribution of physicians. PMID- 11109360 TI - [Beta1 integrins and edema formation in acute inflammation--new therapeutic possibilities?]. AB - BACKGROUND: The role of the intestitium (the extracellular and extravascular tissue) with regard to transcapillary fluid balance and control of interstitial fluid volume has normally been considered to be a "passive controller". MATERIAL AND METHODS: This review is based upon literature collected through the authors' own studies and through Medline searches. RESULTS: Recent studies, however, indicate that the connective tissue cells can actively modulate physical properties of the interstitial matrix, so that it becomes an "active" participant in transcapillary fluid exchange and thereby interstitial fluid homeostasis. The beta 1-integrin system seems to provide a common pathway by which the cells can raise and lower interstitial fluid pressure and thereby regulate the tissue fluid volume. INTERPRETATION: Experiments in which a new anti-inflammatory agent (alpha trinositol), platelet-derived growth factor (PDGF), and a prostagladin F2 alpha analog (latanoprost) modulate interstitial fluid pressure and oedema generation in acute inflammation, suggest that the extracellular matrix can be a target for pharmacological intervention during inflammatory processes. PMID- 11109361 TI - [Metotopic craniosynostoses]. AB - BACKGROUND: Premature closure of the metopic suture leads to inhibited growth of the frontal bones, producing a keel-shaped forehead (trigonocephaly). Simple metopic synostosis is usually sporadic. Trigonocephalic patients account for 8 16% of the referrals to craniofacial centers, with a marked male predominance. Intracranial pressure (ICP) may be increased, whereas shunt-dependent hydrocephalus is infrequent. Infrequently, patients have intra- or extracerebral anomalies; one third have varying degrees of neuropsychological problems. The treatment is primarily surgical. MATERIAL AND METHODS: We present two patients who during infancy developed increasingly keel-shaped foreheads, retruded orbital rims, increased biparietal diameter and close-set eyes (hypothelorism). Both had raised ICP, but normal psychomotoric development. They were operated using radical fronto-orbital surgical remodelling. RESULTS: Recovery was uneventful. Three months post-operatively, they had pleasing cosmetic results with no symptoms of increased ICP. INTERPRETATION: Where metopic craniosynostosis is suspected, the infant should be examined clinically with palpation of fontanelles and sutures, evaluated with respect to the shape and development of the facial skeleton, as well as by X-ray of the skull sutures. Radical fronto-orbital surgical remodelling gives a stable correction of the craniofacial deformity and generally a satisfactory cosmetic result. PMID- 11109362 TI - [Distraction osteogenesis--a new therapeutic principle in complex craniofacial synostosis]. AB - BACKGROUND: Surgical treatment of syndromal craniofacial synostosis consists of combined neurosurgical and maxillofacial reconstructions of the neurocranium and midface. Patients often need several operations if they are to achieve a functionally and cosmetically good result. This is, in part, due to limitations in the amount of "acute" correction possible peroperatively with conventional surgical technique. MATERIAL AND METHODS: In recent years, a new method called distraction osteogenesis has been developed, by which the skeletal elements are gradually advanced. The method is based upon a process whereby the reparative callus formed between the surfaces of two bone segments is subjected to traction. This leads to new bone formation parallel to the vector of distraction. RESULTS: Distraction osteogenesis allows for greater mobilizations, hence the promise of fewer reoperations. INTERPRETATION: The method seems to be a valuable treatment modality in syndromal craniofacial synostosis, where major fronto-orbital, midface or maxillary reconstructions are often called for. PMID- 11109363 TI - [Pyoderma gangrenosum--possible diagnosis in therapy-resistant wounds]. AB - Pyoderma gangrenosum is a chronic ulcerative inflammatory skin disease. The condition is often associated with inflammatory bowel disease. We report three patients with pyoderma gangrenosum successfully treated with cyclosporin. There were difficulties in establishing the diagnosis, and antibiotics, surgical excisions and different wound dressings had been used without effect. PMID- 11109364 TI - [The district practitioner (1836-1984)]. AB - In 1836, a system of district physicians was established in Norway. In addition to providing individual medical services, the district physicians supervised other health care providers and looked after vulnerable groups of patients. They were also chairmen of the local boards of health, and were supposed to engage in anything that might improve local health conditions. The first district physicians were given responsibility for enormous areas, and would rarely get in close contact with the people in their districts. When the districts were divided into smaller units, they came closer to the local people, but they had to struggle hard for recognition. Over the years, however, they succeeded by means of health information and increasingly effective medical remedies. In the late 1950s, local health services were in a deep crisis, but towards the end of the 1960s new and radical political trends initiated a revitalisation. Primary health care became increasingly popular among young doctors, and a large number of new positions were established. However, in 1984, at the height of this boom, the government chose to close down the district physician service. PMID- 11109366 TI - [Physical activity--a crucial factor in the prevention of cardiovascular diseases]. AB - BACKGROUND: Cardiovascular disease is a major health problem. Metabolic syndrome and type 2 diabetes increase the risk of developing cardiovascular disease. MATERIAL AND METHODS: The articles for this review were found by MEDLINE search. RESULTS: Regular physical activity or cardiorespiratory fitness reduces the risk of cardiovascular disease mortality in general and of coronary heart disease in particular. Existing data are convincing regarding the relationship between physical activity and cardiovascular disease, but not conclusive regarding stroke. Regular physical activity or cardiorespiratory fitness is beneficially associated with lipids, blood pressure, insulin sensitivity, obesity, haemostasis and endothelial function. An increased level of physical activity has the potential to improve these factors simultaneously. Regular physical activity decreases the risk of developing type 2 diabetes. Increased physical activity might have a beneficial effect on metabolic control in those who have type 2 diabetes. INTERPRETATION: Physical activity should be strongly emphasised in the prevention of cardiovascular disease and type 2 diabetes. PMID- 11109365 TI - [Adverse effects of antiepileptic agents]. PMID- 11109367 TI - [Physical activity and bodily pain in children]. AB - BACKGROUND: Musculoskeletal complaints are frequently reported not only by adults, but also by children. Sedentary lifestyle has been suggested as a possible cause. MATERIAL AND METHODS: 569 pupils in primary school in a Norwegian municipality, aged 10-15 years, answered a questionnaire on bodily pain, self esteem, body image and physical activity. RESULTS: We found a tendency that the least and the most physically active children reported most complaints. The two groups reported complaints from different regions. INTERPRETATION: Whether complaints in childhood develop into adult chronic pain conditions, and whether increased activity among the most physically passive children can hinder such a development are still unanswered questions. PMID- 11109368 TI - [Should medical errors be avoided at any price?]. PMID- 11109369 TI - [Incidents in health services--prevention and management]. PMID- 11109370 TI - [From Botswana to Tanzania]. PMID- 11109371 TI - [Polycystic lipomembranous osteodysplasia]. PMID- 11109372 TI - [Non-practical guidelines?]. PMID- 11109373 TI - [Cesarean section?]. PMID- 11109374 TI - [R minus]. PMID- 11109375 TI - [Diagnostic sensitivity, diagnostic specificity and predictive value of positive tests]. PMID- 11109376 TI - [Professional independency and sponsoring]. PMID- 11109377 TI - [Tidsskriftet and priorities by the Medical Society]. PMID- 11109378 TI - [The family practice system and students]. PMID- 11109379 TI - [Ownership and administration forms of Norwegian hospitals]. PMID- 11109380 TI - [Social medicine--a working tool for the practitioner]. PMID- 11109381 TI - [Can physicians be leaders?]. PMID- 11109382 TI - [The best health care in the world?]. PMID- 11109383 TI - [Hospital capacity and waiting time for treatment--is there a connection?]. AB - BACKGROUND: The association between hospital capacity and waiting time for treatment is uncertain. MATERIAL AND METHODS: Waiting times for patients on waiting lists for inpatient treatment in 1998 were analysed to disclose possible associations with the hospitals' treatment resources, i.e., general costs, number of beds, doctors or nurses in relation to the population of its catchment area, and the relation between acute and elective admissions. Waiting times were calculated from the National Patient Register, which collects information on hospital stays. Resource data and data on acute admissions were taken from the SAMDATA publications for 1998. RESULTS: Median waiting time varied from 50 to 300 days among the hospitals. Statistical regression models were, however, unable to explain the variation in waiting time on the basis of any variable related to hospital resources or acute admissions that may influence the capacity for elective admissions. INTERPRETATION: To avoid breaches of the guarantees for patients guaranteed a maximum of three months on the waiting list, the median waiting time should be below 12-15 days. This goal may, however, be much too ambitious in view of the fact that the median waiting time for patients with mammary or colon cancer is about 30 days. PMID- 11109384 TI - [The public's expectations of the health services]. AB - BACKGROUND: The challenge of finding ways of allocating public health resources is much debated. Many argue that the public should play a major role in deciding what services should be delivered and paid for. The aim of this study was to collect information on the public opinion on various health policy issues. MATERIAL AND METHODS: A representative sample of 1,342 Norwegians was interviewed in 1998 about their attitudes towards various health policy issues. RESULTS: The results showed that Norwegians have great expectations of the national health services. The majority wants immediate access, free choice, and minimal out-of pocket payments. Factor analysis yielded four latent variables in the response pattern: economic rationing, market-orientation, access and out-of-pockets payment. Women were less in favour of economic rationing, less market-oriented and wanted more influence than men. Free access to services grew more important by age. Politically conservative voters were most in favour of market orientation. INTERPRETATION: To involve the public in priority issues is hard, as their expectations are extensive and contradicting. However, it is most important to involve them in order to establish the understanding that public health services cannot supply everything free of charge to everyone. PMID- 11109385 TI - [Head of department--manager or physician?]. AB - BACKGROUND: In Norway, as in other countries, questions regarding medical leadership in hospital departments are much discussed. The purpose of this study was to determine how much time medical heads of hospital departments spend on various leadership tasks. MATERIAL AND METHODS: Information was collected by a questionnaire survey in 1996. RESULTS: 567 out of 657 (86%) completed the questionnaire. 71% shared the departmental leadership with a nurse, and 48% of these were content with such co-leadership. Nearly all the respondents were clinically active. 49% of heads of large departments used more than half their working hours on administration, compared with 7% of heads of small departments. INTERPRETATION: Selection criteria for heads of hospital departments should be adjusted to the work they actually do. Clinical competence is of importance for all heads of clinical departments; the importance of administrative competence varies with the size of the department. PMID- 11109386 TI - [Head of department--selected and trained as a leader?]. AB - BACKGROUND: The importance of recruitment and leadership training for heads of hospital departments has been discussed for several years. We wanted to study how these heads were recruited, and to what extent they were trained for their leadership tasks. MATERIAL AND METHODS: Information was collected by a questionnaire; 567 out of 657 (86%) responded. RESULTS: 37% of the respondents had been interviewed before being appointed. In the interviews, most of them had been asked about leadership experience and training, a larger proportion among the heads of large departments (more than 99 employees) than small (less than 30 employees). 20% of the heads of small departments and 43% of heads of large departments had undergone leadership training at university level. 25% had spent more than one week per year during the last three years on additional training in leadership and administration compared to 81% on continuing medical education. INTERPRETATION: The results indicate that employers place great importance on leadership training, whereas the heads of department appear to place more importance on continuing medical education. PMID- 11109387 TI - [Is quality assurance only an empty phrase?]. AB - BACKGROUND: Norwegian hospitals and their leaders are required by law to engage in quality assurance. We wanted to study to what extent the heads of hospital departments were actually engaged in such activities. MATERIAL AND METHODS: Data were collected by questionnaires sent to heads of hospital departments in Norway (n = 657), of whom 567 (86%) responded. RESULTS: Only 23% of those interviewed prior to their appointment had been asked about experience in quality assurance, less than 30% had written instructions for their work, and only about 40% received regular follow-up from the hospital administration. The majority registered complaints and mistakes, and was engaged in teaching quality assurance. 58% of the heads of small departments and 73% of those of large departments reported that quality in general suffered because of the demands for higher clinical productivity. INTERPRETATION: Most heads of hospital departments in Norway are engaged in quality assurance work, but the study indicates that hospital administration attaches little importance to this type of work. PMID- 11109388 TI - [Smoking, self-experienced health and social integration among adolescents]. AB - BACKGROUND AND OBJECTIVE: In spite of increased preventive efforts against smoking in the Norwegian population, a substantial number of adolescents starts smoking. This study was meant to increase our understanding of smoking behaviour among adolescents by examining the relationship between subjective and emotional health and social integration on the one hand and smoking on the other. METHOD: The study was based on data from a cross-sectional survey of 828 students in upper secondary schools (91% of all students) in Forde carried out in 1997 (age group 16-19). Univariate and multivariate analyses were used to examine the relationship between smoking and relevant predictor variables. RESULTS: Self assessed health, emotional and psychosomatic health were lower among smokers than non-smokers. Female students were also worse off concerning these health complaints. Smoking prevalence was three times higher among students attending vocational secondary schools compared to students attending college proparatory programmes. Smokers reported better social integration and intimacy with friends than did non-smokers. They also reported greater problems with intimacy with parents. CONCLUSION: In preventive work among adolescents, we have to be aware of an important ambiguity: Smoking is associated with reduced subjective and emotional health, but also with better social integration and intimacy with friends. PMID- 11109389 TI - [The placebo phenomenon--how can it be understood?]. AB - BACKGROUND: While previously being considered "medicine without effect", it is now well documented that placebo works. The understanding of the placebo phenomenon should start with the placebo concept and its historical use related to scientific medicine and the biomedical paradigm. MATERIAL AND METHODS: Theoretical analysis. RESULTS: While scientific medicine is about causality in a domain of substances (energy), placebo works in a mental domain (information). When it works, it is as something meaningful, and not as something causal or true in an empirical sense. INTERPRETATION: Empirical studies on placebo suggest that the body cannot be sufficiently understood as a closed biological system; it should be understood as a system webbed in communication with its cultural context. Even if the paradigmatic consequences of this are challenging, the practical implications amount to no more than to give the art of medicine the status it deserves--not as an alternative to scientific medicine, bus as a necessary part of optimal treatment. PMID- 11109390 TI - [Manpower resources and patient treatment at a regional hospital]. AB - BACKGROUND: Significant increases in hospital budgets over the last few years have not resulted in a comparable increase in patient volume. MATERIAL AND METHODS: The increase in hospital admissions, surgical procedures and outpatient care from 1995 to 1999 at a large regional hospital was compared to the increase in hospital staff numbers over date same period. RESULTS: While admissions were up 6.3%, surgical procedures 3.3% and outpatient consultations 6.7%, the overall increase in staff numbers was 12.0%. The number of physicians was up by 30.6%, that of nurses by 13.1%. Secretarial/administrative and technical staff numbers also increased significantly more than the patient population. The data showed that each group of health care workers had a significant overall decrease in productivity, defined as consultations/examinations per head. However, several departments broke this trend; they showed increases in productivity, i.e. that the increase in patient-related activities greatly increased the increase in staff numbers. INTERPRETATION: Further analysis of how departments that manage to increase productivity are organised and how they plan their activities could give information that would be of value to other departments and units. Hospital productivity and effectiveness are probably more related to management at the patient level that to superior top management or to the ownership structure of the hospital. PMID- 11109391 TI - [Observational units--same good service to lower costs?]. AB - BACKGROUND: With increased demand for hospital services and limited resources in the health sector, modes of organisation of services which give the same health outcome, but at lower cost, are of interest. Observation units in hospitals imply rapid diagnostic procedures and/or brief, but adequate treatment. We looked at the evidence for whether stays in observational units provide similar health outcome and reduce the admissions costs for hospitals compared to standard inpatient stays. MATERIAL AND METHODS: We systematically evaluated four randomized controlled studies that have compared the outcome for chest pain and asthma patients admitted to either observational units or directly to ordinary hospital units. The outcomes in focus were health effects measured as mortality and/or complications, length of stay and direct hospital costs. RESULTS: The studies indicate that for the conditions included, there are no health outcome differences between patients treated in observational units on one hand or in ordinary hospital units on the other. Stays in observational units do not seem to imply increased risk of complication. Furthermore, the studies show a reduction in length of stay and hospital costs associated with the use of observational units. INTERPRETATION: The main reason for the reduction in length of stay and cost is quicker diagnosis in observational units than in the ordinary hospital units. The preconditions for these results are clear by defined criteria for selection of patients to the different units and well-defined protocols for making diagnoses and treating patients. If many patients after staying in observational units are transferred to ordinary hospital units or are discharged and then readmitted, the economic benefits of observational units could be undermined. PMID- 11109392 TI - [On comparison of hospital performance]. AB - BACKGROUND: The motivation to identify the causes of rising health care cost and variations across providers has intensified in all industrialized countries. These countries have an ongoing debate on efficiency and effectiveness in hospital production. In this debate, national and international comparative studies are important. MATERIAL AND METHODS: There are very few international comparative studies that include Norwegian hospitals. Actually we know very little about how Norwegian hospitals are performing compared to others. This paper gives an introduction to comparative studies and to the DEA model which is often used in such studies and also a multilevel model which is not so common. RESULTS: A short review is given of a comparative study of Norwegian and North American hospitals. I also discuss the feasibility of comparative studies of hospitals from the Nordic countries, with references to several comparative studies performed in these countries. INTERPRETATION: Comparative studies are often closely linked to national health politics, policy making and reforms; thus the outcome of such studies is important for the hospitals included. This makes such studies a sensitive field of research. It is important to be aware of the strength and weaknesses of comparative studies and acknowledge their importance beyond the development of new knowledge. PMID- 11109393 TI - [Folling's disease]. AB - Norway is a small country and we have few examples of medical scientists that has discovered and cultivated unknown territory. One very good example is the man who discovered the first link between metabolic disease and brain development. Asbjorn Folling was born in 1888 and discovered "his disease" (phenylketonuria = PKU) in 1934. This article gives a description of his life, discovery and work. PMID- 11109394 TI - [Weight increase as adverse effect of drugs]. PMID- 11109395 TI - [Children and motor competence]. AB - Recently, the topic of motor competence has figured prominently in the media. The claims made are many, but the research that support the statements is seldom cited. The aim of this review article is to address that deficiency by documenting what is really known about the motor competence of children. Motor competence not only allows children to carry out everyday practical tasks, but it is also an important determinant of their level of self-esteem and of their popularity and status in their peer group. While many studies have shown a significant correlation between motor problems and other problems in the social sphere, it has been difficult to establish causal relationships with any degree of confidence, as there appear to be several interactions which need to be taken into account. Research has shown that 6-10% of Norwegian children in the 7 to 10 year age group have a motor competence well below the norm. It is unusual for motor problems to simply disappear over time. In the absence of intervention the syndrome is likely to continue to manifest itself. More recent research points to some of the circularity in this causal network, children with motor problems having been shown to be less physically active than their peers. In a larger health perspective this in itself can have very serious consequences for the child. PMID- 11109396 TI - [Should people with epilepsy exercise?]. AB - Many people with epilepsy, especially those with uncontrolled seizures, live a sedentary life and have low physical fitness. Regular physical exercise may have a moderate seizure preventive effect in 30-40% of the patient population, while in about 10%, strenuous exercise may provoke seizures. Among those prone to exercise-induced seizures there is a preponderance of patients with very poor physical fitness and symptomatic epilepsy. The underlying mechanisms of interaction between epilepsy and physical exercise are mainly unknown. As a general rule, people with epilepsy should be stimulated to participate in recreational and athletic activities. However, as this patient group is highly heterogeneous, counselling on this topic should be individualised and take into account both seizure type and frequency. A distinction may be drawn between three categories: activities with no restrictions, activities with some restrictions, and activities that are not advisable. PMID- 11109397 TI - [Physical activity for mental health]. AB - BACKGROUND: About 50% of the population will be affected by a mental disorder during their lifetime; the most common forms are mood and anxiety disorders and abuse of or dependence on drugs or alcohol. The standard forms of therapy are medication and various forms of psychotherapy. The cost of treating disease is escalating, and the health care system will never be able to meet the need for treatment in this large group of patients. Hence, development of effective self help strategies is important. MATERIAL AND METHODS: In this paper, the scientific basis for promoting exercise as treatment for mental disorders is evaluated on the basis of a review of the literature. RESULTS: Beneficial psychological effects of exercise are best documented for mild to moderate forms of unipolar depression and chronic fatigue syndrome; in these disorders, exercise is an alternative to traditional forms of treatment. A therapeutic effect may also be achieved in panic and generalised anxiety disorder, schizophrenia, conversion and somatoform pain disorder, and alcohol abuse and dependence. INTERPRETATION: Beneficial effects of exercise are well documented. A simple and inexpensive approach like exercise is helpful and might be important for public health. PMID- 11109398 TI - [Social medicine--tracing the lost discipline]. PMID- 11109399 TI - [Give to the ceasar what belongs to him....]. PMID- 11109400 TI - [District practitioner--the fall and the end of an institution]. PMID- 11109401 TI - [Licence for elderly physicians]. PMID- 11109402 TI - [Health services for the aged]. PMID- 11109404 TI - Editorial pet peeves and the colonized title. PMID- 11109403 TI - [Successful introduction of a drug]. PMID- 11109405 TI - Health status and resources of rural homeless women and children. Iowa Homeless Research Team. AB - The purpose of this research is to describe the health status and health resources for homeless women and children in a Midwestern rural community. A group of 31 rural homeless women in a shelter participated in the study by answering questions on the Rural Homeless Interview developed by the investigators. The findings revealed higher than expected rates of illness, accidents, and adverse life events, with the incidence of substance abuse and mental illness being comparable to data from other homeless populations. The data on children were limited by lack of knowledge on the part of their mothers. Some mothers reported that their children were in foster care, had been adopted, or were being cared for by others. The inability to access health and dental care was reported by half of the participants. PMID- 11109406 TI - A causal model of depression in early adolescents. AB - The purpose of this study was to test the extent to which a causal model developed from a theoretical formulation of depression was consistent with data obtained from early adolescents, age 12 to 14. In this cross-sectional correlational design, the final sample consisted of 225 adolescents who responded to a demographic data sheet and instruments measuring depression, self-esteem, state anxiety, and perceived stress in classrooms. The causal model was tested via the LISREL 7 program, using a maximum likelihood structural equation model. The results yielded a chi-square (1, N = 225) = .71, p = .401, indicating a good fit of the model to the data. Perceived stress had the strongest direct, indirect, and total effect on depression in early adolescents. Contrary to expectation, self-esteem did not have a direct effect on depression, and girls did not report higher levels of depression than did boys. PMID- 11109407 TI - The effect of video-based interventions on self-care. AB - Caregivers for dependent, elderly community dwellers are frequently themselves at risk for illnesses exacerbated by stress. This study evaluates a videotape scenario intervention designed to increase self-care behaviors in three groups of caregivers randomly assigned to a control or to one of two experimental groups. Within the 6- to 8-week trial, attitudes toward self-care became more positive in the experimental groups, and all groups reported endorsement of behavioral changes related to self-care. PMID- 11109408 TI - Bearing illness and injury. AB - This qualitative study examined how individuals with catastrophic illness and injury managed their personal and social world. The 28 males and females had endured their chronic conditions from 3 to 25 years prior to the study. Participants were individually interviewed. Responses were analyzed using grounded theory methods. Individuals with catastrophic illness and injury experienced three phases in bearing their situation: finding out, facing reality, and managing reality. Individuals did not progress through stages as has been argued by stage theorists. Rather, the phases flowed together and were reexperienced continuously. Individuals employed three strategies--protecting, modifying, and boosting--in all of the phases to help them endure their circumstances. PMID- 11109409 TI - Systemic oxygen-free radical production in iron-loaded mice. AB - Although iron is an essential element for normal cell metabolism, in excess quantities it is highly cytotoxic and lethal. In fact, acute iron poisoning is a leading cause of overdose mortality in young children. Hereditary hemochromatosis, a disorder of iron metabolism, is currently the most prevalent genetic disorder in the world, which results in organ failure and premature mortality. Hence, an enhanced understanding of its pathogenesis is critical for providing safe and effective nursing care to affected individuals and their families. Although the exact mechanism of iron's toxicity is not known, it was hypothesized that chronic iron loading would result in increased tissue (heart, liver, and spleen) concentrations of iron and increased free radical production in a murine model (n = 20). Our results show that chronic iron loading results in highly significant dose-dependent increases in tissue concentrations of iron and systemic free radical generation (p < 0.001). PMID- 11109410 TI - The evolution of nursing education in a postindependence context--Ghana from 1957 to 1970. AB - Development of nursing education in Ghana between 1957 and 1970 is characterized by dynamic change and growth. Published manuscripts, personal interviews, and letters were used to analyze evolution of nursing education during this period. Following independence in 1957, developments in nursing education continued to be strongly influenced by external organizations and their designated experts. Policies, such as the local training of nurses and Africanization, provided impetus for nurses to further their education to assume senior positions in nursing education and administration. Emphasis on training nurses to work in a hospital-based curative health system, which had been the legacy of colonialism, gradually shifted to a broad-based education that prepared nurses to work in a variety of settings. Changes in nursing education occurred within an economic climate that presented ongoing impediments, yet the vision of the first generation of Ghanaian nurse leaders facilitated the tremendous progress seen during this period. PMID- 11109411 TI - Information exchange. PMID- 11109412 TI - [Long-term outcome after coronary emergency intervention]. AB - BACKGROUND AND OBJECTIVE: Coronary angioplasty (CAG) has become an acceptable method of treating an acute coronary syndrome (myocardial infarction [MI] or unstable angina [UA]). It was the aim of this study to determine whether the results of such emergency treatment differed from those after elective CAG. PATIENTS AND METHODS: Results of emergency CAG in 581 patients (aged 60 +/- 11 years; 77% males) admitted to the authors' hospital between July 1994 and December 1996 were compared with those of elective CAG in 2,460 patients (aged 61 +/- 10, admitted during the same period. Follow-up information was obtained after 22.4 +/- 11 months in 93.2% of the patients by examination, written answers to annual questionnaires, data being collected on survival, repeat cardia catheterizations, other interventions, aorto-coronary bypass, occurrence of myocardial infarction, the patients' general state and drugs received. RESULTS: 19 of 517 patients (3.7%) of the group who had undergone elective CAG had died during the follow-up period, compared with 107 of 2436 of the emergency cohort (4.4%; not significant). There were also no significant differences regarding repeat cardiac catheterization, interventions, coronary bypass or re-admission. The proportion of subsequent emergency CAG among all CAGs was 16.8% in the emergency cohort, 8.8% after elective angiography (p < 0.001). CONCLUSION: Coronary angiography performed in patients with an acute coronary syndrome has no prognostic significance regarding mortality and morbidity after the acute phase of the disease. PMID- 11109413 TI - [Colonoscopy in patients over 80 years of age. Indications, methods and results]. AB - BACKGROUND AND OBJECTIVE: Diseases of the colon, especially cancer, are age specific and thus occur more frequently in an aging population. Early diagnosis is prognostically decisive. Indications for coloscopy are often delayed in the elderly. It was the aim of this study to determine advantages and disadvantages of coloscopy in those aged over 80 years. PATIENTS AND METHODS: A total of 157 coloscopies in patients aged over 80 years (average 83 years, oldest patient 94 years) were retrospectively analysed (63 males, 94 females) regarding indications, results and complications, with special reference to cancer of the colon. The main indication for coloscopy in this cohort was occult faecal blood or non-acute rectal bleeding (27%), which together were important indicators of endoscopically significant findings (in 76%). Not only blood in the stool but manifold other signs can point to colorectal carcinoma. CONCLUSION: Even in patients aged over 80 years coloscopy is a safe and well-tolerated investigation. Its results can be improved by thorough pre-investigational intestinal preparation so that repeat examination is unnecessary. The high incidence of significant findings and the easy accessibility to the site of colon cancer should provide an earlier and more common indication of total coloscopy in this age group. PMID- 11109414 TI - [Eosinophilic myocarditis in Churg-Strauss syndrome. A rare cause of left heart decompensation with lung edema]. AB - HISTORY: A 50-year-old woman was admitted because of marked dyspnoea at rest and signs of left heart failure with pulmonary oedema. 9 years ago, the diagnostic constellation of bronchial asthma, polyneuropathy, pericardial effusion and eosinophilia had indicated Churg-Strauss syndrome. Since then she had remained symptom-free under maintenance doses of azathioprine (for 2 years) and gradually reduced doses of steroids. INVESTIGATIONS: Chest X-ray showed signs of pulmonary congestion and cardiomegaly, echocardiography demonstrating enlargement of the left heart with marked impairment of ventricular function, and both revealed pericardial effusion. The electrocardiogram showed complete absence of R waves and ST elevation in leads V1-V5. Coronary angiography excluded coronary artery disease. Myocardial biopsy contained signs of active but no longer acute myocarditis with eosinophilic tissue infiltration and microgranulomas. White blood cell count was normal, but there was marked eosinophilia (39%). IgE was elevated (601 kIU/l). DIAGNOSIS, TREATMENT AND COURSE: In view of the good therapeutic effects 9 years ago, this relapse of Churg-Strauss syndrome with eosinophilic myocarditis was again treated with azathioprine and steroids. In addition, diuretics, digitalis and ACE-inhibitors successfully treated the heart failure. In the course of treatment the signs of inflammation, including the eosinophilia, regressed or became normal. CONCLUSION: After a 10-year remission without complication of a Churg-Strauss syndrome the onset of cardiac signs is the decisive long-term prognostic factor. PMID- 11109415 TI - [Acute pancreatitis--adverse effect of 5-aminosalicylic acid (mesalazine) in various galenic dosage forms]. AB - HISTORY AND ADMISSION FINDINGS: A 34-year-old woman, being treated with oral mesalazine for colitis, developed upper abdominal pain radiating to the back one week after the drug was started, but the symptoms quickly regressed after mesalazine had been discontinued. The patient was admitted about 15 months later because of watery diarrhoea and lower abdominal pain. Physical examination revealed dehydration and pain on pressure over the lower abdomen. INVESTIGATIONS: Coloscopy demonstrated Crohn's disease of the colon, more marked on the left. Laboratory tests indicated inflammatory disease. TREATMENT AND COURSE: During treatment with oral prednisone and mesalazine enema the diarrhoea and laboratory parameters of inflammatory disease regressed. But 10 days after the start of treatment the patient complained of upper abdominal pain radiating to the back. Amylase and lipase levels were, respectively, four and twelve times normal. Ultrasound demonstrated an enlarged head of the pancreas, while endoscopy of the oesophagus, stomach and duodenum was unremarkable. 3 days after prednisone and mesalazine had been discontinued the symptoms had regressed and laboratory tests were normal. The symptoms did not recur when prednisone was administered again. CONCLUSION: Mesalazine may cause an acute pancreatitis when given either orally or by rectal infusion. PMID- 11109416 TI - [Dissection of extracranial arteries supplying the brain]. PMID- 11109417 TI - [Lipoprotein(a): epidemiology and therapeutic approaches]. PMID- 11109418 TI - [Liability for suicide during inpatient hospital treatment. Verdict of the federal court 20 June 2000]. PMID- 11109419 TI - [Detection of HFE polymorphism in German patients with hereditary hemochromatosis]. PMID- 11109420 TI - [Detection of HFE polymorphism in German patients with hereditary hemochromatosis]. PMID- 11109421 TI - [Cognition disorders and dementia--new end-organ damage in hypertension]. PMID- 11109422 TI - [Effectiveness of felodipine in hypertensive patients with mild cerebral cognition disorders in a randomized double-blind study]. AB - BACKGROUND AND OBJECTIVE: Cognitive impairment occurs more frequently in hypertensives than in normotensive individuals. Early signs of cognitive impairment are predictors of dementia in late life. Felodipine is capable of almost normalizing plasma viscosity, which is elevated in most of hypertensive patients, thus improving microcirculation. The aim of this study was to evaluate whether this hemorheologic property of felodipine in addition to its blood pressure lowering effect can improve cognitive performance in hypertensive patients. PATIENTS AND METHODS: Randomized, double-blind comparison between felodipine 10 mg and hydrochlorothiazide 50 mg amiloride 5 mg (HCT/amiloride) in patients 50-70 years of age with impaired cognitive function (c.l. test 1-2 points) and with resting blood pressure values of diastolic > 95 and < or = 115 mmHg and/or systolic > 160 and < or = 210 mmHg. Blood pressure measurements and evaluation of total short term storage capacity were done at the beginning and after 12 weeks of treatment. RESULTS: 31 patients (14 felodipine and 17 HCT/amiloride) were included in the per protocol analysis. Blood pressure values at the beginning and after 12 weeks of treatment were (mmHg): for felodipine systolic 168 +/- 4 and 150 +/- 6 (p < 0.01), diastolic 108 +/- 3 and 88 +/- 4 (p < 0.001). For amiloride/HCT systolic 173 +/- 8 and 150 +/- 10 (p < 0.01), diastolic 105 +/- 5 and 88 +/- 5 (p < 0.001). Short term storage capacity improved by 15 +/- 6 bits during felodipine treatment (p < 0.001) and by 9 +/- 9 bits during amiloride/HCT treatment (p < 0.05). Thus cognitive improvement was superior by 67% in the felodipine group compared to amiloride/HCT (p < 0.05). CONCLUSION: In this study a pronounced improvement of mental performance occurred in patients treated with felodipine. Since the cognitive gain was significantly superior to amiloride/HCT treatment there must be an additional blood pressure independent effect of felodipine, such as enhancing microcirculation. Whether these properties possibly counteract the development of dementia in hypertensives has to be evaluated in long term studies in more patients. PMID- 11109423 TI - [Mucoviscidosis screening of newborn infants in the Dresden district. Results from 1 June 1996 to 31 March 2000]. AB - PROBLEM: Cystic fibrosis (CF) is the most common congenital defect of metabolism in Europe. Therefore we tried to detect this disease as early as possible before clinical symptoms occur. Thus early diagnosis can be the basis for an early start of treatment. PATIENTS AND METHODS: As part of a complete neonatal screening program in our region we investigated the concentration of immuno-reactive trypsin (IRT) in dried blood samples. Genomic investigation of the same blood sample for the most common CF mutations was performed when a critical value of IRT was exceeded. RESULTS: From 6/1996 until 3/2000 (46 months) we investigated the blood of 49,926 newborn children. Due to a high IRT value (> 70 ng/ml) in 579 cases, a genomic investigation was performed. In 38 children we detected one of the three most frequent CF mutations (delta F508, G551D, R553X) by polymerase chain reaction (PCR). The sweat test (pilocarpine iontophoresis) confirmed cystic fibrosis in 8 newborns. Four times a homocygocity for the mutation delta F508 was found and four times a compound heterocygocity (one time delta F508/R553X und three times delta F508/others). Only three of these eight CF patients already had clinical symptoms of the disease at this time, only in one case had this diagnosis been considered. An additional newborn with meconium ileus had been diagnosed as cystic fibrosis before performing the screening. Up to now we have not found any case of false negative testing. CONCLUSION: We found this procedure of neonatal testing practicable and therefore recommend its continuation. The genomic test should include the search for additional CF mutations. As alternative method a second IRT-test should be considered at the end of the first month of life for those children with initial IRT-concentrations above 150 ng/ml and without evidence of the three tested CFTR-mutations. PMID- 11109424 TI - [Etiology of initially unexplained confusion of excitability in deadly nightshade poisoning with suicidal intent. Symptoms, differential diagnosis, toxicology and physostigmine therapy of anticholinergic syndrome]. AB - HISTORY AND ADMISSION FINDINGS: After a walk in a wood a 55-year-old teacher was admitted to the emergency unit of a university hospital because of somnolence and excitability. Her rectal temperature was 37.8 degrees C, she had sinus tachycardia (rate of 130/min) but no other significant findings. INVESTIGATIONS: With the exception of C-reactive protein (10 mg/dl), MCV (101 fl), MCH (34 pg) and arterial blood gases (pH 7.483, pCO2 35.5 mmHg, base excess 5.1 mmp/l) laboratory tests were within normal limits. Qualitative screening of serum for benzodiazepines, barbiturates and antidepressives was negative. Neurological examination, including lumbar puncture and cranial computed tomography were noncontributory. TREATMENT AND COURSE: 10 hours after admission the patient developed signs of an anticholinergic syndrome with mydriasis, dry mouth, tachycardia, hot skin and an atonic bladder. Physostigmine 2 mg completely reversed the neurological and mental symptoms. After gas chromatography, mass spectrometry of a urine sample showed an atropine molecular fragment with a molecular weight of 271. At intervals of 3 to 5 hours the recurrence of confusion and excitability required 4 further i.v. injection of physostigmine. The patient subsequently became accessible to psychiatric examination and reported that during the walk she had swallowed 8-10 berries of deadly nightshade with suicidal intent. CONCLUSION: In case of excitability and confusion as well as somnolence or coma of uncertain aetiology an anticholinergic syndrome caused by ingestion of atropine-containing plants or psychoactive drugs (phenothiazines, butyrophenones, tri- or tetracyclic antidepressants) should be included in the differential diagnosis. If there are suggestive clinical findings (tachycardia, somnolence, coma or threatened respiratory arrest, physostigmine should be given if there are no contraindications. PMID- 11109425 TI - [Cholinergic syndrome with unconsciousness in amanita poisoning]. AB - HISTORY AND ADMISSION FINDINGS: A 41-year-old patient was found in his flat in a state of coma. After emergency treatment his vital signs were stable and he was transferred to an acute hospital with possible cannabis intoxication. The patient, a hobby gardener, was previously well and had an adversion to the use of any chemical substances. The main symptom showed a cholinergic syndrome with deep coma. We assumed plant ingestion because of the clinical picture and history. INVESTIGATIONS: The laboratory results were within normal limits apart from a slight rise of the serum creatinine kinase level. The electrocardiogram showed a bradycardia. A drug-screening could not be performed. TREATMENT AND COURSE: The differential diagnosis of plant alkaloids or mushroom toxins were considered due to possible plant ingestion and a cholinergic syndrome. Later the toadstool (Amanita muscaria) was found. After treatment oft the cholinergic syndrome with high doses of atropine primary poison elimination was performed. 24 hours later the patient awoke from his coma. Visual hallucinations persisted for a few days. No organic damage due to the intoxication was found. CONCLUSION: Toxic mushroom ingestion can produce a variety of clinical pictures. Most commonly an anticholinergic syndrome is found, but this was not the case in this patient. The effect of the poison depends on the amount and the preparation, so that no reliable outcome prediction can be made. The drug "poisonous mushroom" is legal and hallucinogenic substances are trendy. As a result clinical signs like those described here will have to be expected in the future. PMID- 11109426 TI - [The role of diagnostic imaging in neuromuscular diseases]. PMID- 11109427 TI - [Nuclear magnetic resonance tomography in multiple sclerosis]. PMID- 11109428 TI - [Testing for blood alcohol level within the scope of social medicine expert assessment]. PMID- 11109429 TI - [Testing for blood alcohol level within the scope of social medicine expert assessment]. PMID- 11109430 TI - [Pacemaker defect]. PMID- 11109431 TI - [Periumbilical pain]. PMID- 11109432 TI - [Retinal vascularization in sarcoidosis]. AB - OBJECTIVES: To describe the characteristics of retinal vasculitis related to sarcoidosis, and to study the radio-clinical findings of sarcoidosis in case of retinal vasculitis. METHODS: We performed a retrospective study on 33 cases of retinal vasculitis associated with sarcoidosis. Our patients had a complete ophthalmological examination and a systematic fluorescein angiography was performed. Recorded data included clinical, biological and radiological evaluation of sarcoidosis. RESULTS: Retinal vasculitis were and remained asymptomatic in almost a third of the patients (n = 10). In 11 patients (33%) a decreased visual acuity of more than 3 lines was observed. Periphlebitis was observed in all cases, and 75.7% were of non ischemic forms. Thirty patients out of 33 (90.9%) were treated with systemic steroids in whom 17 were treated for ophthalmological reasons. The presence of anterior or posterior uveitis, more than one attack of vasculitis, and significant hyalitis were statistically associated with visual acuity decrease. Ninety percent of patients regained their initial visual acuity, only three patients at the end of the medical follow-up remained with a visual acuity below or equal to 2/10. Thirty one patients (91%) presented with mediastinal and pulmonary signs concordant with sarcoidosis on chest x-rays. Extrapulmonary localisations were common in case of retinal vasculitis (91% of patients). CONCLUSION: Retinal vasculitis associated with sarcoidosis are often asymptomatic. In most cases, there are bilateral, non ischaemic periphlebitis. They seem to have a relatively better prognosis than in other etiologies of vasculitis and they are part of an evolutive multisystemic sarcoidosis. Most of the time treatment with systemic steroids was indicated for ophthalmological or extra ophthalmological reasons. PMID- 11109433 TI - [Good clinical practice in using antibiotics in the hospital. Current status in 207 public and private hospitals in 1999]. AB - OBJECTIVES: The purpose of this study was to map activities developed in hospitals to monitor antibiotic usage and evaluate implementation of French guidelines for good clinical practice on use of antibiotics in the hospital setting. METHODS: A questionnaire was mailed to the head of the pharmacy of 300 French hospitals. The questionnaire targeted methods developed to monitor antibiotic usage (antibiotic committees, local recommendations, types of prescription and dispensing, surveillance, information and evaluation activities). RESULTS: The response rate was 69% (207 answers). A local committee supervised antibiotic usage in 49% of the hospitals (nosocomial, drug or antibiotic committees). Local recommendations existed in 120 hospitals (59%) and 42% of the hospitals had a validation process before dispensing drug in accordance with the recommendations. Antibiotic prescription was nominal in 65% of the hospitals and specific monitoring was carried out in 42% of them. Antibiotic consumption was monitored in 80% of the hospitals and resistance was monitored in 53%. Twelve percent of the hospitals used an electronic network to share information on prescription and bacteriological results. Regular internal training existed in 20% of the hospitals and evaluation methods (medical audits, impact measures) in 14%. DISCUSSION: Careful monitoring of antibiotics is implemented in most hospitals. Strict application of guidelines, definition and implementation of indicators, and evaluation methods must be improved. Implementation of better hospital monitoring of antibiotics requires: i) a local consensus to limit the antibiotics available and guidelines to adapt to local infections; ii) dissemination of guidelines and training for prescribers; iii) implementation of a dispensing system to check the validity of prescriptions according to local guidelines; iv) implementation of indicators to monitor bacterial resistance and the volume of antibiotics used. PMID- 11109434 TI - [Simultaneous onset of rheumatoid polyarthritis and type 1 diabetes]. AB - BACKGROUND: Rheumatoid arthritis and insulin-dependent diabetes mellitus are both autoimmune disorders of unknown etiology. We report the case of a patient who developed the two diseases simultaneously. CASE REPORT: A 64-year-old man with no remarkable medical history developed insulin-dependent diabetes disclosed by ketoacidosis that occurred 3 weeks after onset of a bilateral symmetrical polyarthritic syndrome characteristic of rheumatoid arthritis. DISCUSSION: These two disorders share common susceptibility of subjects with MHC class II molecules HLA DRB1*04. Immunological studies have also shown a common Th1 type cytokine secretion pattern in both diseases. Epidemiological studies have not however clearly demonstrated a link between them. PMID- 11109435 TI - [Collapsing glomerulopathy and cytomegalovirus, what are the links?]. AB - BACKGROUND: Collapsing glomerulopathy is a form of focal and segmental glomerulosclerosis which occurs preferentially in black people. It causes severe nephrotic syndrome and quickly progresses towards end-stage renal failure. CASE REPORT: We report the case of a 16-year-old black girl from Guadeloupe who was admitted for tetanus and edema in 1996. She had hypoparathyroidism, renal failure and a nephrotic syndrome as well as cytomegalovirus infection. Renal biopsy showed collapsing glomerulopathy. The renal function improved on glucocorticoid and ganciclovir therapy and her serum creatinine stabilized around 250 mumol/l for two years. DISCUSSION: Collapsing nephropathy is the cellular type of focal and segmental glomerulosclerosis. The main etiology is the human immunodeficiency virus. A viral infection may be involved in its pathogenesis. Other viruses could be linked with this nephropathy. This case report relates a case associated with a cytomegalovirus viruria. The clinical course might be related with the antiviral treatment. PMID- 11109436 TI - [Complete regression of a somatotropin-secreting adenoma with lanreotide monotherapy]. PMID- 11109437 TI - [Pulmonary cryptococcoma in a patient infected by HIV virus]. PMID- 11109438 TI - [Hospital in tune with handicapped persons. A plea for hospital accessibility!]. PMID- 11109439 TI - [Clinical benefits of hypocholesteremic drug treatments]. PMID- 11109440 TI - [Aphthae or painful ulcers induced by nicorandil]. AB - A RECOGNIZED CAUSE OF BUCCAL APHTHOSIS: Nicorandil is the leading drug in a new pharmacology class of well tolerated anti-angina products with vasodilating action. The first cases of buccal aphthosis or ulcerations induced by nicorandil were reported in 1996. Among drugs inducing buccal aphthosis or ulcerations, the largest body of information available on reported cases concern nicorandil. CHARACTERISTIC LESIONS: Forty cases of painful buccal lesions induced by nicorandil have been recorded. Among the 8 publications in the literature, 7 were reported by French groups. The prevalence of this adverse effect is an estimated 5%. Patient age varies from 60 to 90 years with an even gender distribution. A history of aphthosis is noted in 23% of the cases. The lesions vary in size from 0.5 cm to 3 cm and in number from 1 to 10, generally localized on the inner aspect of the cheeks or on the tongue. The lesions develop for about 3 to 36 before diagnosis and the delay to onset of signs after initiating nicorandil treatment is 15 days to 24 months (generally 2 months). Lesions basically develop after high-dose treatments and, for a few cases, after increasing the dosage. Cure is obtained in all cases after 1 to 12 weeks. OPEN QUESTIONS: The probability that nicorandil is the cause of these ulcerations or aphthae is very high. Certain terms do however need to be defined with precision. Are we talking about aphthae or ulcerations? The term painful ulceration would appear to be preferable because it includes both ulcerations and aphthae in cases lacking further information. The wide variability in the geographical distribution could likely be linked to underreporting. The notion of a dose effect suggests a toxic mechanism that could develop in predisposed populations (ethnic origin...) and would be directed against buccal targets (salivary glands...) via the active drug or its metabolites or via drug interactions. This new adverse effect of nicorandil is quite spectacular and very painful. However, no life-threatening event has been observed. PMID- 11109441 TI - [Value of stereotaxic biopsy in diagnosis of subclinical breast lesions]. AB - ALTERNATIVE TO SURGERY: New stereotactic guided breast biopsy procedures may constitute a major issue for the diagnosis of non-palpable breast lesions detected at mammography by eliminating the need for surgery in many women with benign breast disease. INDICATIONS: Vacuum-assisted core biopsies provide more complete sampling than the conventional 14-gauge stereo-tactic core biopsies, reducing the number of unsatisfactory biopsies. The more invasive advanced breast biopsy device obtains an intact lesion in its entirety for histological assessment. Currently, there is no definite strategy delineating the precise indications for the diagnosis of screening detected abnormalities. PERSPECTIVES: Because of the increase of the diagnostic armamentarium, care of women with non palpable breast lesions should be multidisciplinary, involving radiologist, surgeons and histologists and rigorous medical and economic evaluation of diagnostic strategies involving these new health technologies should be pursued. PMID- 11109442 TI - [Rapidly progressing pulmonary nodules reveal Wegener disease]. PMID- 11109443 TI - [Thomas Mann and radiology]. PMID- 11109444 TI - [The imaging diagnosis of diabetes mellitus]. PMID- 11109445 TI - [A magnetic resonance study of 39 children with different causes of short stature]. AB - PURPOSE: To investigate the relationship between Magnetic Resonance (MR) findings and the presence of isolated growth hormone deficiency (IGHD) or multiple pituitary hormone deficiency (MPHD). MATERIAL AND METHODS: 39 children, 27 boys and 12 girls (mean age 9.6 years) were enrolled. The growth hormone deficiency and MPHD were defined by standard laboratory methods. MR scans of the hypothalamo hypophyseal tract were carried out on all patients before the therapy. Perinatal histories were reviewed. RESULTS: The pituitary anterior lobe was considered small in 13/39 (33%). Twenty-six of 39 (66%) patients did not show any abnormalities of the hypothalamohypophyseal tract. An ectopic neurohypophysis was found in 8/39 patients (20%). Only 2 patients among those with an ectopic neurohypophysis showed a small anterior pituitary lobe. Thirty-three of 39 (84.6%) patients showed IGHD while 6/39 (15.4%) patients showed MPHD. Twenty eight of 39 (71%) patients had a severe deficiency while 11/39 (28%) patients had a moderate deficiency. CONCLUSION: Our study confirms the usefulness of MRI in the diagnostic and therapeutic approach to short stature in children; in fact 48% of patients in our series showed MR findings reasonably related to hormonal deficiency. PMID- 11109446 TI - [The assessment of the impact of a double reading by expert readers in a mass mammographic study]. AB - PURPOSE: To evaluate the role of double reading of screening mammograms by expert radiologists. MATERIAL AND METHODS: We analyzed the results of independent readings of a proficiency test of screening mammography (140 cases, 32 cancers) performed by four expert radiologists. Double reading was simulated by matching the four original readings in 6 possible combinations. The impact of double reading over single reading was evaluated in terms of increased sensitivity and increased recall rate. RESULTS: Of 32 carcinomas 22, 6, or 4 were identified by 4, 3, or 2 readers, respectively. Of 108 cases negative for cancer a recall for further investigations was suggested by 4, 3, 2, 1 or no reader(s) in 3, 3, 9, 14, or 79 cases, respectively. Inter-reader diagnostic repeatability was good (k = 0.65). Single readers achieved an average sensitivity of 89% (range 87.5-90.6%) and an average recall rate of 12.2% (range 7.4-16.6%). Simulated double reading achieved an average increase in sensitivity of 8.8% (range 6.2-10.95%) and an average increase of recall rate of 6.2% (range 3.8-8.3%). CONCLUSIONS: Even though the relative increase of recall rate is relevant (+53.2%), the corresponding gain in sensitivity justifies the use of double reading, which was confirmed to be worthwhile also when expert radiologists are involved. This study confirms the opportunity of adopting double reading as a routine procedure in mammographic screening. PMID- 11109447 TI - [The radiological diagnostic and clinical approach to the patient with stab and cut wounds of the chest. The authors' personal experience]. AB - PURPOSE: To report our personal experience with the clinical and radiological diagnostic approach to stab and cut wounds of the thoracic cage and its content, a type of injury whose diagnosis and treatment, as well as the surgical approach, vary case by case. CT of deep penetrating wounds permits correct assessment of severe changes such as pneumothorax, hemothorax and pneumomediastinum. MATERIAL AND METHODS: In the last three years we examined 57 patients (48 men and 9 women; mean age 34 years, range 16-54): chest radiography was performed in 51 of them, with orthogonal projections in the standing and sitting positions. Chest CT was performed in emergency with i.v. contrast agent injection, with scans from the midneck to the diaphragm insertion to study border regions. Thoracostomy with pleural drainage was performed in 35 patients with pneumothorax and hemothorax while thoracotomy was performed in 8 patients, namely 4 with injury to the diaphragm, 2 to the heart, 1 with tear of the main bronchial artery and 1 of the aortic arch. RESULTS: The most frequent symptoms we found were chest pain (100% of cases) and dyspnea (84%); laboratory data showed anemia and decreased hematocrit levels in 28 cases. Chest radiography was negative in 14 cases. The patients were then examined with CT to exclude radiographic underestimation of minimal pneumothorax, small lacerocontusive or hemorrhagic foci and hemothorax, which were observed in 4, 2 and one cases, respectively, and where radiography was actually negative for traumatic changes. Chest radiography was positive in 43 cases: the most frequent finding was pneumothorax, with 37 cases (86%)--8 of them associated with hemothorax and 5 with pneumomediastinum. Lacero-hemorrhagic foci of lung parenchyma were found in 5 cases and single pulmonary hematoma from punch crossing was seen in 1 case. DISCUSSION AND CONCLUSIONS: CT was an accurate tool and had higher sensitivity than chest radiography in detecting and detailing pneumothorax, pneumomediastinum and lacero-hemorrhagic foci, as well as in quantifying hemothorax. Chest radiography had 12% false negatives and therefore we decided to perform CT in all the patients with penetrating wounds to prevent radiographic underestimation. Given the low rate of false negatives (7/57 cases) CT might appear superfluous but since in 2 of these 7 cases we had massive pneumothorax and pneumomediastinum associated with neck emphysema we suggest its use to prevent complications, clinical failures and medicolegal problems. CT permits correct assessment of penetrating stab and cut wounds of the chest and efficient and targeted treatment, which can be conservative, with thoracostomy with pleural drainage, or surgical. PMID- 11109448 TI - [Spiral computed tomography in the assessment of vascular lesions of the pelvis due to blunt trauma]. AB - PURPOSE: We investigated the role of Helical Computed Tomography (CT) in the evaluation of low or high flow vascular injuries in patients with blunt pelvic trauma. MATERIAL AND METHODS: From May 1998 to December 1999, forty-nine patients (32 men and 17 women, ranging in age 14-59 years) with acute symptoms from blunt pelvic trauma were submitted to Computed Tomography (CT). A conventional radiography of the pelvis had been performed in all cases. CT was performed with a helical unit (thickness 8 mm, reconstruction interval 8 mm, pitch 1.5) after intravenous contrast agent (150-180 mL) rapid infusion (4-5 mL/s, 60 s acquisition delay from bolus starting) and using a power injector. A second spiral acquisition was performed in all cases from the iliac roofs to the inferior border of the pubic symphysis. Vascular hemorrhage was considered as low flow when the hematoma appeared as a focal homogeneous density area and as high flow when associated with contrast agent extravasation. Moreover, traumatic assessment included evaluation of the hematoma, of the leakage site and of the involved vessel. RESULTS: Radiologic examination of the pelvis revealed fractures in 35/49 patients (71.4%). Helical CT allowed us to identify low flow hemorrhage in 37 patients, affected with hematomas from fracture of the iliac wing or of the sacrum (14 cases), tear of the pelvic (3 cases) or extrapelvic (4 cases) muscular structures, or injury of the venous plexus (20 cases). In four patients two vascular injuries were detected. High flow hemorrhage was seen in 12 patients, who had Helical CT findings of contrast agent extravasation along the common iliac vein (3 cases), external iliac artery (3 cases), internal iliac artery (4 cases), internal pudendal artery (1 case), obturator artery (1 case), inferior epigastric artery (2 cases), superior gluteal artery (2 cases), inferior gluteal artery (1 case), cremasteric artery (1 case). In 6 patients with high flow hemorrhage, two vascular injuries were shown. In all these patients, an extraperitoneal hematoma was associated with the contrast agent extravasation. DISCUSSION AND CONCLUSIONS: Fractures of the pelvic ring generally result from severe trauma. Management of these injuries must include not only treatment of the skeletal trauma but also of the associated shock and complications. Major blood loss usually occurs as a result of bleeding from the branches of the internal iliac artery. With respect to pelvic plain radiography, CT provides superior detailing of fractures, position of fracture fragments and extent of diastasis of the sacroiliac joints and pubic symphysis. Moreover CT provides diagnostic information regarding the presence or absence of pelvic bleeding and can identify the site of bleeding. In our experience, Helical CT allows us to distinguish high flow hemorrhage, where vascular injuries must be treated first, from low flow hemorrhage which can be managed differently. PMID- 11109449 TI - [The imaging quality of the ureteral intramural tract in the adult male. A comparison between suprapubic and transrectal echography]. AB - PURPOSE: US is usually considered of little help in studying the ureters because it only shows proximal and distal tracts. The distal ureter is also difficult to study with most of the other imaging methods. Transrectal probes improve depiction of the intramural and pelvic tracts of the ureter, as well as image quality, dramatically providing good results in a short time. The first and, to our knowledge, the only report on this procedure has been Holm's (1994), where the author reported good results though in a small series. For years we have used transrectal US, performed for prostate studies, also to examine intramural ureters and have obtained good results in a short time. Thus we decided to compare the transrectal with the transabdominal approach. MATERIAL AND METHODS: We examined 92 randomly selected male patients with various urologic conditions but no ureter involvement using 3.5 MHz convex and 5.0 MHz biplane transrectal probes. Image quality was rated on an arbitrary scale (0-4) and then submitted to statistical analysis. RESULTS: Transrectal images were clearly superior to transabdominal ones (p = 0.012) and particularly, ureters were depicted in 93% versus 60% of cases, respectively. The amount of urine in the bladder appears to play a major role (the greater the amount the worse the image quality), while ureter depiction is independent of body habit and bladder squeezing. CONCLUSIONS: Our results are clearly superior to Holm's. Also, considering that the transrectal examination requires a short time, is cost-effective and little invasive, we believe that this method can play an extremely important role in lower ureter studies. PMID- 11109450 TI - [An evaluation of orthotopic ileal neobladders by dynamic digital urography]. AB - PURPOSE: To report on the diagnostic capabilities of dynamic digital urography in the evaluation of orthotopic ileal neobladders. MATERIAL AND METHODS: Ten male patients (aged 61.3 +/- 14.7 years) with orthotopic ileal neobladder (4 Studer, 4 Camey type II with spiralized ileal segment and 2 classic Camey type II) underwent dynamic digital urography using an angiographic unit (Philips DVI/ARC A). Eight patients were asymptomatic and two presented mild nycturia. All patients should have undergone follow-up conventional urography, which was replaced, for the purposes of our study, by dynamic digital urography. The intestinal cleansing regimen usual for the double contrast barium enema was used in all the patients. Ninety mL of a nonionic iodinated contrast agent were administered i.v. with an automatic injector. After precontrast mask images, two postcontrast sequences were acquired 15 s and 30 min after the injection (each made of 20 images acquired every 10 seconds). Postprocessing consisted of digital image subtraction and videorecording. RESULTS: Renal pelvis and calyceal systems were well visualized in 18/20 excretory systems. Eighteen of 20 ureters were visualized completely up to the ureteral jet, and two were only partially visualized. Calyceal, pyelic and ureteral enlargement with ureteral kinking were observed in a patient with classic Camey type II. High motility was seen in 11 ureters, moderate in 5 and mild in 4. The orthotopic ileal neobladder was well opacified in 8/10 patients. Effective peristalsis of the afferent ileal segment was seen in all the Studer type neobladders. Ureteral reflux was not observed in any patient. DISCUSSION: After orthotopic ileal neobladder reconstruction, the most frequent complications include urinary leakage, intestinal obstruction, venous thrombosis, stenosis of the neobladder anastomoses, incontinence, cancer recurrence, stone formation. In the follow-up, many diagnostic tools are used: intravenous urography, retrograde cystography, urodynamic studies, transabdominal and transrectal ultrasonography. With dynamic digital urography the nephrographic evaluation was possible in all patients, as well as the evaluation of calyceal, pyelic and ureteral opacification and even ureteral peristalsis. Moreover, this diagnostic tool allows the morphofunctional dynamic study of the ureteral neobladder anastomoses and of the orthotopic ileal neobladder, even evaluating the residual peristalsis of the detubularized ileal segment. A major drawback of the new method is the relatively high radiation dose given to the patient. A limitation of the study is the selection of a population of completely or nearly asymptomatic patients. CONCLUSIONS: Dynamic digital urography provides useful morphologic and functional information in the follow-up of patients with orthotopic ileal neobladder and could replace conventional urography in symptomatic patients. PMID- 11109451 TI - [The treatment of iatrogenic pneumothorax with small-gauge catheters. The author's personal experience in 30 cases]. AB - PURPOSE: Pneumothorax (PNX) is the most frequent complication in patients who have undergone lung biopsy. If PNX is asymptomatic and < 30%, it does not require treatment, while if it is > 30% and the patient is symptomatic treatment is needed. As a rule surgery is required and patients are hospitalized and undergo intrathoracic drainage with positioning of a large gauge catheter--i.e. over 15 French (F). In the last 10 years radiologists have begun treating PNX with much smaller catheters (7-10 F). We report the execution technique using 6.3 F catheters and the results obtained in 30 patients with symptomatic iatrogenic PNX and/or iatrogenic PNX > 30%. MATERIAL AND METHODS: All the patients underwent CT guided lung biopsy. Immediately after the procedure some follow-up scans were performed and a further expiratory radiograph with the patient in upright position was carried out after at least 2 hours. If an asymptomatic PNX < 30% was found the patient was discharged and submitted to radiographic follow-up the following morning and every 24 hours thereafter for 2 days. If there was a symptomatic PNX and/or a PNX > 30% an intrathoracic drainage catheter was positioned. Under fluoroscopic or CT guidance we positioned a 5.7 F intrathoracic pig-tail catheter at a point corresponding to the 3rd or 4th intercostal space on the midclavear line. After manual suction of intrathoracic air we connected the catheter to a Hemlick valve and repeated the chest radiograph 4 hours later. If the PNX had not reformed the patient was discharged and submitted to radiographic follow-up every 24 hours for 3-5 days. On the contrary if the PNX had reformed, or if pain and/or dyspnea symptoms or signs persisted, the catheter was connected to a continuous-suction system and the patient rehospitalized for about 6 days. Oximetry was performed in all patients before biopsy, on PNX diagnosis, and after pulmonary re-expansion. RESULTS: All the cases were resolved and 9 patients were followed-up in the outpatients department. Drainage had to be repeated in 2 patients only and the 5.7 F catheters replaced with an 8 F and a 10 F catheters. Oximetric data were always correlated with the presence/absence of PNX. In particular, in PNX > 30% we found over 10% reduction relative to prebiopsy values. This datum was corrected and came to meet the prebiopsy value as soon as the lung was re-expanded. No significant changes were seen in PNX < 30%. CONCLUSIONS: Small gauge catheters provide the following advantages: the procedure presents a low risk of complications, is easy to carry out and much better tolerated by the patient; also in some cases the cost is lower because no hospitalization is required. The close correlation of oximetric values with the presence/absence of PNX < 30% could be considered to decrease follow-up radiographic examinations. Finally the possibility of treating iatrogenic PNX using radiological techniques further promotes the acceptability of lung biopsy by colleagues from other branches of medicine. PMID- 11109452 TI - [The percutaneous treatment of uterine fibromas by means of transcatheter arterial embolization]. AB - INTRODUCTION: We report our preliminary experience with arterial embolization of uterine fibroids in seven women, focusing on the technical aspects of the procedure and the clinical and morphological results during the follow-up. MATERIAL AND METHODS: February to December 1999 seven women (mean age 47) underwent transcatheter arterial embolization of both uterine arteries as a permanent treatment for fibroids. We included in this study single or multiple, bleeding and/or large fibroids, symptomatic on compression, contraindicated for myomectomy because of high surgical or anesthesiologic risks or myomata in which myomectomy could probably be converted into hysterectomy. Fibroids enlarging the uterus to the size of 25 weeks' pregnancy or more, pedunculated myomata or small submucous fibroids--smaller than 5 cm--were excluded. Uterine arterial embolization was performed bilaterally, till a total blockage of flow, by injecting permanent embolization material: polyvinyl-alcohol (PVA) particles of increasing size from 150 to 500 mu and in varying amounts from 10 to 24 mL, depending on fibroid size and degree of vascularization. RESULTS: A technical success was achieved in all cases and no late complications were seen. At 6-month clinical follow-up all compressive symptoms had disappeared; regular menses had returned in 57% of patients, milder hyper-dysmenorrhea was present in 28% compared to pretreatment symptoms; only in one case (14%) was permanent amenorrhea observed. The 3-month and 6-month US follow-up studies showed an average 40.7% (range 10-50%) and 51% (range 25-83%) reduction in the fibroid size, respectively. All the small myomata (about 2 cm in size) were unidentifiable at 6-month US follow-up. In no cases did new fibroids appear. DISCUSSION: Surgery is the traditional treatment for symptomatic uterine fibroids (hysterectomy, myomectomy). More recently, hormone therapy and operative endoscopy (laparoscopy and hysteroscopy) have been introduced as alternatives, together with uterine embolization previously applied preoperatively in extensive bleeding neoplasms or to control post-partum hemorrhage. Transcatheter embolization of the uterine arteries feeding large fibroids is a minimally invasive technique which could be safely used as an alternative to surgery, and a valuable in the definitive treatment of symptomatic, large or multiple, intramural or submucosal fibroids. In agreement with literature findings, in the present series symptoms resolved completely in over 85% of cases after embolization, with an average reduction in fibroid size over 50% at 6-month follow-up in large fibroids, whereas smaller size myomata were no longer detectable at US and no new fibroids had formed. CONCLUSIONS: Our preliminary experience confirms that arterial embolization is a promising alternative to surgery in the definitive treatment of fibroids, thanks to its high efficacy and safety, also reducing patient hospitalization and costs. PMID- 11109453 TI - [Antineoplastic perfusion with percutaneous stop-flow control in the treatment of advanced pelvic malignant neoplasms]. AB - PURPOSE: The object of our study was to apply percutaneous stop-flow technique to advanced pelvic cancer in order to evaluate its feasibility, standardize the procedure and obtain preliminary results. MATERIAL AND METHODS: April to December 1997 we submitted ten patients with advanced pelvic cancer to percutaneous stop flow technique. Seven patients had a pelvic recurrence from carcinoma of the rectum, two patients had inoperable recto-sigmoid cancer, and another one had a local recurrence of ovarian cancer. All treatments were performed under general anesthesia. A stop-flow balloon catheter was placed via a transfemoral arterial and venous access above the aortocaval bifurcation and below the emergence of renal arteries and veins. The pelvic district was isolated by filling the balloon catheters and pneumatic cuffs at the thigh, and the antineoplastic agents (cisplatinum, 80 mg/m2 followed by mitomycin C, 30 mg/m2) were sequentially infused by means of an extracorporeal circuit. Blood flow was interrupted for a maximum of 20 min to limit tissue damage, especially of the anal sphincter. Hemofiltration was run during the last 3 min of stop-flow and in the following minutes, achieving at least 5 liters of ultrafiltration. Morphological response was evaluated by CT or MR scan performed prior to and 40 days after the treatment. RESULTS: Over a 2-year follow-up 2 of our 10 patients are alive and 8/10 have died (median survival 9.6 months). Death followed tumor progression in 6 cases; one patient died during the procedure and another one after 7 days, both secondary to pulmonary embolism. Complications included intra-arterial rupture of the balloon in one case and a large inguinal hematoma 10 days after the treatment, requiring hospitalization. No patient showed positive morphological response; two patients only showed stable disease. CONCLUSIONS: This trial supports the feasibility of using the percutaneous stop-flow procedure in an angiography room setting; the stop-flow technique failed to permit the effective control of the tumors. PMID- 11109454 TI - [Intramuscular metastases from a melanoma diagnosed by magnetic resonance. A case report]. PMID- 11109455 TI - [Hemorrhagic encephalopathy due to cyclosporin 1 year after heart transplantation. A case report]. PMID- 11109456 TI - [Isolated bilateral neurofibromas of the brachial plexus in neurofibromatosis type 1. The magnetic resonance aspects in a case]. PMID- 11109457 TI - [Wegener's granulomatosis: the neuroradiological findings in a case of atypical clinical onset]. PMID- 11109458 TI - [Bronchial carcinoid studied by spiral computed tomography with 3-dimensional reconstructions and virtual endoscopy. A case]. PMID- 11109459 TI - [An aneurysm and pseudoaneurysm of the splenic artery. 2 cases. Their treatment by transcatheter embolization and a review of the literature]. PMID- 11109460 TI - [An aneurysm of the abdominal aorta associated with a periaortic lymphoma. A case report]. PMID- 11109461 TI - [A monotypic variant of hepatic angiomyolipoma completely composed of perivascular epithelioid cells. A case]. PMID- 11109462 TI - [An isolated complete fracture of pancreas due to skiing trauma. Integrated imaging in a case]. PMID- 11109463 TI - [Intrahepatic neurofibromatosis in childhood. 2 cases]. PMID- 11109464 TI - [From tubers to the fine lace of the lung]. PMID- 11109465 TI - [Aortic stenosis and intestinal blood loss from angiodysplasia: valve replacement is a therapeutic option]. AB - Three patients, men aged 67, 68 en 61, had for many years suffered from recurrent iron deficient anaemia and were found to have an aortic stenosis. Coloscopy identified angiodysplastic lesions, and we suspected additional lesions in the small intestine. Despite surgical and endoscopic treatment, gastrointestinal bleeding recurred. Replacement of the stenosed aortic valve with a bioprosthesis led to a recovery in the haemoglobin levels of all three patients. Angiodysplasias are a common source of gastrointestinal bleeding with an invisible origin. Data from the literature suggest an association between aortic stenosis and angiodysplasia although epidemiological evidence is lacking. PMID- 11109466 TI - [Disability claim review by non-physicians to meet the legal requirement of Dutch work disability certification laws]. AB - The number of assessments of incapacity for work has increased to such an extent in the Netherlands (450,000 per year, of which 150,000 are first assessments) that experiments are now being undertaken to employ non-physicians for these assessments. There is no shortage of insurance physicians however: the number available is underestimated as many of these have resigned their regular jobs to perform the same activities on a freelance or second basis. A problem is certainly that these physicians are hindered by an exceedingly bureaucratic system in which e.g. patients are summoned by the administrative section to be reassessed who have already resumed working. Non-physicians may assist in the assessments, but in most cases there is a relation between psychic, somatic and social factors, which requires the generalist physician's eye. The main problem is that government regulations lead to a steady growth in the number of (re)assessments required. A possible but onorthodox solution is proposed which involves medical certification of incapacity for work. This may discourage long term absenteeism and at the same time it may draw the physician's attention to the existence of an industrial health care problem. PMID- 11109467 TI - [Diagnosis and treatment of tuberculous lymphadenitis of the neck]. AB - The incidence of extrapulmonary tuberculosis is rising. The patients are predominantly immigrants and HIV-infected persons. Tuberculous lymphadenitis of the neck is the most common presentation, in the Netherlands about 200 patients a year. Fine needle aspiration with auramine/Ziehl-Neelsen stain investigation, culture and cytological examination is the diagnostic procedure of choice. If this fails to be conclusive excision biopsy is the next step. If the diagnosis is suspected on clinical, epidemiological, laboratory and bacteriological (presence of acid-fast bacilli) or cytological/histological grounds, treatment is always started, awaiting culture results. The principles for treatment are the same as for pulmonary tuberculosis. On the basis of the available literature it can be proposed to shorten the duration of treatment from 9 to 6 months. PMID- 11109468 TI - [Roaming through methodology. XXVII. Problems with longitudinal studies: confounding of the relationship between age, time of measurement and birth cohort]. AB - Nowadays, age-related research questions are investigated using longitudinal study designs where measurements are repeated over time in the same subjects. Cross-sectional studies are used less often for this purpose, as they provide measurements at only one point in time thus making it more difficult to establish relationships between variables and outcomes. However, longitudinal measurements also have drawbacks, even in investigations that, for example, are aimed at age related changes in physical and psychosocial characteristics during growth and development in young people. In the study of age-related trends, several types of confounding may occur: birth cohort effects, time-of-measurement effects, and test-and-learning effects. If control measurements are carefully collected, for example, in a multiple longitudinal design, or by not repeatedly measuring a control group, the confounding may be quantified and the study results corrected accordingly. PMID- 11109469 TI - [Clinical thinking and decision making in practice. A patient with anal cancer and hypercalcemia]. AB - A 57-year-old male patient, recently known with an anal carcinoma with inguinal lymph node involvement, was admitted because of anorexia, nausea, vomiting and constipation. On physical examination the patient was dehydrated, and a systolic murmur, grade III/VI, punctum maximum apex cordis, was heard. Serum calcium was raised (4.50 mmol/l), as was the serum creatinine (328 mumol/l). Both values had been normal 14 days before admission. Serum parathormone was suppressed. A bone scan did not reveal evident lesions in the skeleton. FDG-PET scan showed uptake of the tracer into the bone marrow. A bone biopsy showed metastasis of a squamous cell carcinoma. Shortly after that the patient died. Hypercalcaemia is associated with cancer. Colorectal/anal carcinomas have a low incidence of hypercalcaemia. The prognosis of patients with cancer associated with hypercalcaemia is poor. PMID- 11109470 TI - [Diagnostic image (12). Ramsay Hunt syndrome]. AB - A 17-year-old woman developed a unilateral facial paralysis with vesicles in the left ear and in the throat. This combination is known as Ramsay Hunt syndrome. PMID- 11109471 TI - [Cesar therapy is temporarily more effective in patients with chronic low back pain than the standard treatment by family practitioner: randomized, controlled and blinded clinical trial with 1 year follow-up]. AB - OBJECTIVE: To determine the effectiveness of a special form of exercise therapy ('Cesar therapy') on self reported recovery and improvement of posture amongst patients with chronic aspecific lower back pain. DESIGN: Prospective randomized controlled and blinded investigation. METHOD: After informed consent had been obtained, patients with chronic aspecific lower back pain were given, on a randomized basis, either an exercise therapy (experimental group, n = 112) or a standard treatment by their general practitioner (control group, n = 110). Outcome measures were self reported recovery of back pain and improvement of posture (thoracic and lumbar spine, pelvis). Self reported recovery was determined by means of a dichotomized 7-point scale (questionnaire). Posture was measured qualitatively by a panel of 11 Cesar therapists (blinded) and quantitatively by an optical-electronic posture recording system (Vicon). Measurements were taken at baseline (pre-randomization) and at 3, 6 and 12 months after randomization. RESULTS: Three months after randomization, patients who were treated according to Cesar therapy, reported an improvement in their back symptoms (80%) significantly more often than the control group (47%). In both groups, however, only small improvements in posture were found. The judgement of the Cesar panel exhibited a significant difference between the two groups, with respect to the spine, in favour of Cesar therapy. Differences between the groups were still present 6 months after randomization, but could no longer be detected at 12 months after randomization. CONCLUSION: Cesar therapy was significantly more effective than standard treatment among patients with chronic lower back pain for a period of 6 months after randomization. PMID- 11109472 TI - [Clinical presentation, treatment, and follow-up of 32 patients with a primary intracranial germinoma, registered during the previous 15 years in the Dutch Pathological-Anatomical National Automated Archive (PALGA)]. AB - OBJECTIVE: Evaluation of clinical presentation, treatment and follow-up of patients with intracranial germinoma in the Netherlands. DESIGN: Retrospective. METHOD: The case histories of 32 patients with histologically verified intracranial germinoma, registered in the period 1983-1999 in the Pathological Anatomical Nationwide Automated Archive (PALGA), were studied. Fifty of the 59 registered patients were found of whom 6 had no germinoma. Informed consent was obtained from 32 of the 44 patients with respect to studying their medical records for age, symptoms at presentation, diagnostic investigations, presence of tumour markers, treatment and follow up. RESULTS: The patient group consisted of 23 men and 9 women aged 6 to 35.6 years (mean: 17.3) and was subdivided with respect to their tumour localization. In patients with pineal localization symptoms of increased intracranial pressure and eye movement disorders were most prominent, whereas in patients with suprasellar localization endocrine disorders prevailed. Thirty-one patients were treated with radiotherapy, one with combined radiotherapy and chemotherapy and one surgically. Twenty-six patients had remained disease free after a median follow-up of 39 months (range: 0-144 months). One patient developed an intracranial embryonal carcinoma and another a testis seminoma. Two patients died because of recurrences. Two other patients died of causes not directly related to the germinoma. CONCLUSION: At the time of this study 84% of all patients treated with radiotherapy were disease-free. Although the percentage patients who had recovered after treatment (surgical and radiotherapy) was high, many patients either already had or subsequently developed neurological and endocrinological deficiencies. PMID- 11109473 TI - [Primary liver cell carcinoma as complication of alpha 1-antitrypsin deficiency in a 67 year old man]. AB - A 67-year-old male patient with pulmonary emphysema was diagnosed with liver cirrhosis. Further investigations revealed an alpha 1-antitrypsin deficiency caused by a PiZZ mutation. The liver cirrhosis was complicated by the development of a hepatocellular carcinoma. The patient died from the consequences of mesentary vein thrombosis. The protease inhibitor alpha 1-antitrypsin controls the tissue damaging effects of proteases which are produced by granulocytes. In the case of alpha 1-antitrypsin deficiency, progressive damage of the lung tissue occurs, resulting in emphysema. The accumulation of abnormal alpha 1-antitrypsin in hepatocytes can result in cirrhosis, with an increased chance of carcinoma. The deficiency is caused by a mutation in the Pi-gene on chromosome number 14. Although treatment options are at present limited, making an early diagnosis has important implications for the prognosis and intended management with respect to the prevention of complications, both in the patient as well as in first degree relatives (children and siblings). PMID- 11109474 TI - [Fifty years of plastic surgery in Netherlands III. Structural surgery of genitalia]. PMID- 11109475 TI - [Acute and chronic low back pain: results of systematic reviews]. PMID- 11109476 TI - [Acute and chronic low back pain: results of systematic reviews]. PMID- 11109477 TI - Methodological issues and challenges for a feminist community psychology: an introduction to a special issue. PMID- 11109478 TI - Feminist approaches to social science: epistemological and methodological tenets. AB - This paper is a primer for community psychologists on feminist research. Much like the field of community psychology, feminist scholarship is defined by its values and process. Informed by the political ideologies of the 1970s women's movement (liberal, radical, socialist feminism, and womanism), feminist scholars reinterpreted classic concepts in philosophy of science to create feminist epistemologies and methodologies. Feminist epistemologies, such as feminist empiricism, standpoint theory, and postmodernism, recognize women's lived experiences as legitimate sources of knowledge. Feminist methodologies attempt to eradicate sexist bias in research and find ways to capture women's voices that are consistent with feminist ideals. Practically, the process of feminist research is characterized by four primary features: (1) expanding methodologies to include both quantitative and qualitative methods, (2) connecting women for group-level data collection, (3) reducing the hierarchical relationship between researchers and their participants to facilitate trust and disclosure, and (4) recognizing and reflecting upon the emotionality of women's lives. Recommendations for how community psychologists can integrate feminist scholarship into their practice are discussed. PMID- 11109479 TI - Searching for feminism: an analysis of community psychology literature relevant to women's concerns. AB - Articles published in both the American Journal of Community Psychology and Journal of Community Psychology, from their inception in 1973 through 1997, were content analyzed for women relevance, diversity, feminism, and historical change. Overall, 9.8% of the articles reviewed (N = 2,178) were considered women relevant, 4% recognized diversity among women, and 3% were considered feminist. There was an average yearly increase in women-relevant and feminist articles from 7.3 pre-1990 to 11.2 post-1990, and 1.6 pre-1990 to 4.6 post-1990, respectively. Overall, mental health and motherhood were the most addressed content areas. Among feminist articles, gender roles and violence against women were most salient. Race and SES were the most noted issues of diversity in both women relevant and feminist articles. While an increase in feminist publications by both journals is promising, stereotypes of women and other oppressed groups continue to be perpetuated. PMID- 11109480 TI - Speaking for ourselves: feminist methods and community psychology. AB - Although feminist and community psychology share a number of epistemological and methodological perspectives that guide their respective theories and research practices, it has been argued that community psychology has not fully integrated a feminist perspective into the discipline. This paper examines how community psychology and feminist research methods might combine to help us better understand women's experiences without essentializing or universalizing those experiences. The authors offer a series of suggested directions for feminist research that may also prove promising for community psychology. Particular attention is paid to feminist social constructionist approaches insofar as they address the complex relationship between epistemology and methodology. PMID- 11109481 TI - Feminist and community psychology ethics in research with homeless women. AB - This paper presents a feminist and community psychology analysis of ethical concerns that can arise throughout the process of doing research with women who are homeless. The unique contexts of the lives of women who are homeless demand that researchers redefine traditional ethical constructs such as consent, privacy, harm, and bias. Research that fails to do this may perpetuate the stereotyping, marginalization, stigmatization, and victimization homeless women face. Feminist and community research ethics must go beyond the avoidance of harm to an active investment in the well-being of marginalized individuals and communities. Using feminist and community psychology ethics, this paper addresses some common problems in research with women who are homeless, and argues for the transformation of research from a tool for the advancement of science into a strategy for the empowerment of homeless women and their communities. PMID- 11109482 TI - Subverting culture: promoting HIV/AIDS prevention among Puerto Rican and Dominican women. AB - This article discusses the challenges faced by researchers and interventionists when attempting to promote change in social norms and normative beliefs that promote HIV/AIDS risk-related behaviors among Puerto Rican and Dominican women. The article focuses on the role of culture in HIV/AIDS prevention with women by analyzing the sociohistorical context of some cultural beliefs and by illustrating the tension between risk-related and protective cultural beliefs in research conducted by the authors with women in both New York and Puerto Rico. The authors propose that promoting changes in sex-related social norms and normative beliefs might be constructed as a subversive act and present the challenge this analysis poses for community psychology. They conclude that this conceptualization might be construed as subversive because rather than idealizing culture, it promotes changes that respect diversity within the culture and foster participation in the development of new cultural values, beliefs and norms. PMID- 11109483 TI - Stories of relative privilege: power and social change in feminist community psychology. AB - Stories about community work in New Zealand and Scotland are presented to describe and reflect on issues central to feminist community psychology. Organizing a lesbian festival, Ingrid Huygens describes feminist processes used to equalize resources across Maori (indigenous) and Pakeha (white) groups. Heather Hamerton presents her experiences as a researcher using collective memory work to reflect on adolescent experiences related to gender, ethnicity, and class. Sharon Cahill chronicles dilemmas and insights from focus groups about anger with women living in public housing in Scotland. Each story chronicles experiences related to oppression and privilege, and describes the author's emotions and reflections. Individually and collectively, the stories illustrate the potential offered by narrative methods and participatory processes for challenging inequalities and encouraging social justice. PMID- 11109484 TI - The use of anonymous DNA markers in assessing worldwide relatedness in the yeast species Pichia kluyveri Bedford and Kudrjavzev. AB - Pichia kluyveri, a sexual ascomycetous yeast from cactus necroses and acidic fruit, is divided into three varieties. We used physiological, RAPD, and AFLP data to compare 46 P. kluyveri strains collected worldwide to investigate relationships among varieties. Physiology did not place all strains into described varieties. Although the combined AFLP and RAPD data produced a single most parsimonious tree, separate analysis of AFLP and RAPD data resulted in significantly different trees (by the partition homogeneity test). We then compared the distribution of strains per band to an expected distribution. This suggested we could separate both the AFLP and RAPD datasets into bands from rapidly and slowly changing DNA regions. When only bands from slowly changing regions (from each dataset) were included in the analysis, both the RAPD and AFLP datasets supported a single tree. This second tree did not differ significantly from the cladogram based on all of the DNA data, which we accepted as the best estimate of the phylogeny of these yeast strains. Based on this phylogeny, we were able to demonstrate the strong influence of geography on the population structure of this yeast, confirm the monophyly of one variety, question the utility of maintaining another variety, and demonstrate that the physiological differences used to separate the varieties did not do so in all cases. PMID- 11109485 TI - Heat shock protein 80 of Neurospora crassa: sequence analysis of the gene and expression during the asexual phase. AB - Heat shock protein 80 (Hsp80) of Neurospora crassa, a member of the stress-90 protein family, is a cytosolic molecular chaperone that interacts directly with Hsp70 to form a hetero-oligomeric complex. The complete nucleotide sequence of the gene encoding this protein, along with the 5'- and 3'-flanking DNA, is reported. The coding sequence is interrupted by two introns, 61 and 30 nucleotides, respectively, in length. The deduced amino acid sequence corresponds to a 695-residue polypeptide with a calculated molecular mass of 78,894 Da and an average pI of 4.94. Primer extension experiments demonstrated two transcription start sites, a major and a minor one. No sequence motifs resembling the standard eukaryotic heat shock elements were evident in the putative promoter region. Immunoblot analysis showed Hsp80 protein to be present in the mature, dormant conidia, while the hsp80 transcripts were not detected. Both the transcripts and the protein were present in the germinating conidia in the absence of externally applied stress. PMID- 11109486 TI - Transplantation and subsequent behavior of mitochondria in cells of Phytophthora. AB - Mitochondria isolated from streptomycin-resistant (S(r)) protoplasts of Phytophthora parasitica were transferred into chloramphenicol-resistant (Cpr) protoplasts of P. parasitica or Phytophthora capsici with an average successful rate of 1.7 x 10(-4), using a selective medium containing streptomycin. No colonies appeared when self-fusion products of donor mitochondria or recipient protoplasts were exposed to the selective medium. Mitochondria isolated from Cpr protoplasts of P. capsici were also transferred into S(r) protoplasts of P. parasitica with a similar success rate using a selective medium containing chloramphenicol. Zoospores produced by the Cpr + S(r) intraspecific mitochondrial hybrid gave rise to S(r) and Cpr + S(r) cultures. The second generation zoospores produced by S(r) and Cpr + S(r) cultures also gave rise to S(r) and Cpr + S(r) cultures, suggesting the possible occurrence of fusion between some of the Cpr mitochondria and S(r) mitochondria, and the displacement of non-fused Cpr mitochondria in the receptor protoplast by the donor S(r) mitochondria. Zoospores produced by the interspecific mitochondrial hybrid gave rise to Cpr, S(r), Cpr + S(r), and Cps + Ss cultures. The second generation zoospores produced by Cpr + S(r) or S(r) cultures also gave rise to the same four types of cultures, suggesting the existence of residual antibiotic-sensitive mitochondria (Cps + Ss) in the parental isolates and the random distribution of Cpr, S(r), and Cps + Ss mitochondria during asexual reproduction. Results suggest that the phenotype of antibiotic resistance/sensitivity was the end result of the interactions among the three types of mitochondria. PMID- 11109487 TI - Purification of the NADP+:F420 oxidoreductase of Methanosphaera stadtmanae. AB - Methanosphaera stadtmanae (DSM 3091) is a methanogen that requires H2 and CH3OH for methanogenesis. The organism does not possess an F420-dependent hydrogenase and only low levels of F420. It does however possess NADP+:F420 oxidoreductase activity. The NADP+:F420 oxidoreductase, the enzyme which catalyses the electron transfer between NADP+ and F420 in this organism, was purified and characterized. NAD+, NADH, FMN, and FAD could not be used as electron acceptors. Optimal pH for F420 reduction was 6.0, and 8.5 for NADP+ reduction. During the purification process, it was noted that precipitation with (NH4)2SO4 increased total activity 16-fold but reduced the stability of the enzyme. However, recombination of cell free extracts with resuspended 65-90% (NH4)2SO4 pellet returned activity to near cell-free extract levels. Neither high salt or protease inhibitors were effective in stabilizing the activity of the partially purified enzyme. The purified enzyme from M. stadtmanae possessed a molecular weight of 148 kDa as determined by gel filtration chromatography and native-PAGE, consisting of alpha, beta, and gamma subunits of 60, 50, and 45 kDa, respectively, using SDS-PAGE. The Km values were 370 microM for NADP+, 142 microM for NADPH, 62.5 microM for F420, and 7.7 microM for F420H2. These values were different from the Km values observed in the cell free extract. PMID- 11109488 TI - Subtilisins of Bacillus spp. hydrolyze keratin and allow growth on feathers. AB - Keratinase is a serine protease produced by Bacillus licheniformis PWD-1 that effectively degrades keratin and confers the ability to grow on feathers to a protease-deficient B. subtilis strain. Studies presented herein demonstrate that B. licheniformis Carlsberg strain NCIMB 6816, which produces the well characterized serine protease subtilisin Carlsberg, also degrades and grows on feathers. The PWD-1 and Carlsberg strains showed a similar time-course of enzyme production, and the purified serine proteases have similar enzymatic properties on insoluble azokeratin and soluble FITC-casein. Kinetic analysis of both enzymes demonstrated that they have high specificity for aromatic and hydrophobic amino acids in the P1 substrate position, although keratinase discriminates more than subtilisin Carlsberg against charged residues at this site. Nucleotide sequence analysis of the serine protease genes from B. licheniformis strains PWD-1, Carlsberg NCIMB 6816, ATCC 12759, and NCIMB 10689 showed that the kerA-encoded protease of PWD-1 differs from the others only by having V222, rather than A222, near the active site serine S220. Further, high-level expression of subE-encoded subtilisin from B. subtilis (78% similar to subtilisin Carlsberg) also confers growth on feathers on a protease-deficient B. subtilis strain. While strain PWD-1 and the kerA protease efficiently degrade keratin, keratin hydrolysis and growth on feathers is a property that can be conferred by appropriate expression of the major subtilisins, including the industrially produced enzymes. PMID- 11109489 TI - A survey of 16S rRNA and amoA genes related to autotrophic ammonia-oxidizing bacteria of the beta-subdivision of the class proteobacteria in contaminated groundwater. AB - In this study, we investigated the size and structure of autotrophic ammonia oxidizer (AAO) communities in the groundwater of a contamination plume originating from a mill-tailings disposal site. The site has high levels of dissolved N from anthropogenic sources, and exhibited wide variations in the concentrations of NO3- and NH3 + NH4+. Community structures were examined by PCR DGGE targeting 16S rDNA with band excision and sequence analysis, and by analysis of amoA fragment clone libraries. AAO population sizes were estimated by competitive PCR targeting the gene amoA, and correlated significantly with nitrate concentration. Most samples revealed novel diversity in AAO 16S rDNA and amoA gene sequences. Both 16S rDNA and amoA analyses suggested that all samples were dominated by Nitrosomonas sp., Nitrosospira sp. being detected in only 3 of 15 samples. This study indicated numerical dominance of Nitrosomonas over Nitrosospira in groundwater, and suggests that groundwater ammonia oxidizers are more similar to those dominating freshwater sediments than bulk soil. PMID- 11109490 TI - Effects of mesophilic and thermophilic composts on suppression of Fusarium root and stem rot of greenhouse cucumber. AB - Three composts were tested for their ability to suppress root and stem rot caused by the soil borne fungal pathogen Fusarium oxysporum f. sp. radicis-cucumerinum (FORC) on cucumber. Two of the composts were prepared from separated dairy solids either by windrow (WDS) or vermicomposting (VMC) while the third, obtained from International Bio-Recovery (IBR), was prepared from vegetable refuse using aerobic digestion. Three sets of potting mixes were prepared by mixing the composts with sawdust at varying ratios, and seeded with cucumber cv. Corona. After 14 days of growth in the greenhouse, inoculum of FORC (20 mL of 5 x 10(6) micro-conidia per mL) was applied to each pot at three different times (14, 21, and 35 days). In unamended inoculated pots, the pathogen caused stunted growth and reduced flowers. Amendment of WDS in the potting mix suppressed these symptoms, while VMC and IBR had no effect. All three composts reduced the FORC colony forming units (cfu) at the end of the experiment (10 weeks). There was a large increase of fluorescent bacteria near the vicinity of roots particularly in WDS amended potting mixes. When water extracts of the composts were plated onto acidified potato dextrose agar (APDA), only IBR contained a potent thermostable inhibitor to FORC. This inhibitor was removed by activated charcoal but was not partitioned into petroleum ether at acid, basic, or neutral pH. Inhibition of FORC by IBR was not due to electrical conductivity or trace elements in the compost. Contrasting effectiveness of the WDS and VMC made from the same waste suggests that composting method can influence the disease suppression properties of the finished compost. PMID- 11109491 TI - Isolation and characterization of highly thermophilic xylanolytic Thermus thermophilus strains from hot composts. AB - This is the first detailed report of xylanolytic activity in Thermus strains. Two highly thermophilic xylanolytic bacteria, very closely related to non-xylanolytic T. thermophilus strains, have been isolated from the hottest zones of compost piles. Strain X6 was investigated in more detail. The growth rate (optical density monitoring) on xylan was 0.404.h-1 at 75 degrees C. Maximal growth temperature was 81 degrees C. Xylanase activity was mainly cell-bound, but was solubilized into the medium by sonication. It was induced by xylan or xylose in the culture medium. The temperature and pH optima of the xylanases were determined to be around 100 degrees C and pH 6, respectively. Xylanase activity was fairly thermostable; only 39% of activity was lost after an incubation period of 48 h at 90 degrees C in the absence of substrate. Xylanolytic T. thermophilus strains could contribute to the degradation of hemicellulose during the thermogenic phase of industrial composting. PMID- 11109492 TI - Natural incidence of endophytic bacteria in pea cultivars under field conditions. AB - Pea plants grown in the field were used to study the natural incidence of endophytic bacteria in the stem. Eleven pea cultivars at the flowering stage were screened for the presence of endophytic bacteria using a printing technique with surface disinfested stem cross-sections on 5% Trypticase Soy Agar (TSA). Five cultivars showed colonization. Cultivar Twiggy showed the highest and most consistent colonization and was further investigated. Stems of cv. Twiggy at the pod stage were analyzed for endophytic bacterial types and populations. Cross sections of surface disinfested stems were printed on 5% TSA. Endophytic bacterial populations decreased from the lower to the upper part of the stem. One section from the third and the fourth internode was surface disinfested, homogenized, and spiral plated on the media 5% TSA, R2A, and SC (Davis et al. 1980). Over a series of 30 samples, 5% TSA gave significantly better recovery of bacterial endophytes compared with R2A and SC media. For most stems, populations ranged from 10(4) to 10(5) CFU/g except in one of the field blocks in which endophyte populations were uniformly higher. Comparison of colony counts by spiral plating and printing showed a positive correlation. The most frequently recovered bacterial types were Pantoea agglomerans and Pseudomonas fluorescens. Less frequently isolated were Pseudomonas viridiflava and Bacillus megaterium. PMID- 11109493 TI - A factorial design analysis of chitin production by Cunninghamella elegans. AB - Chitin production by Cunninghamella elegans (IFM 46109) was studied with a two level full factorial design, varying time of cultivation and the concentration of D-glucose, L-asparagine, and thiamine in the culture medium. The material extracted was characterized by infrared and NMR spectroscopy. The highest chitin yield, 28.8%, was comparable with the highest in the literature and was obtained with a medium containing 60 g.L-1 of glucose, 3 g.L-1 of asparagine, and 0.008 mg.L-1 of thiamine. Increasing the time of cultivation from 24 h to 72 h did not affect chitin production. The three factors showed significant positive effects on chitin production, without interactions between them. PMID- 11109494 TI - Nutritional requirements of Brettanomyces bruxellensis: growth and physiology in batch and chemostat cultures. AB - The nutritional requirements of Brettanomyces bruxellensis have been investigated. Batch culture and chemostat pulse techniques were used to identify growth-limiting nutrients. The study included determination of the essential components of the culture medium and quantification of the effects of the components. Among the components tested, ammonium sulfate and yeast extract had a significant effect on glucose consumption, growth, and ethanol production. However, if the ammonium sulfate concentration is above 2 g/L, an inhibitory effect on B. bruxellensis growth is observed. The yeast extract appears to be the most important and significant component for growth. The maximum amount of synthesized biomass is proportional to the concentration of yeast extract added to the culture broth (in the tested range). Magnesium and phosphate ions are probably not essential for B. bruxellensis. These ions appear to be supplied in sufficient amounts by the yeast extract in the culture medium. Brettanomyces bruxellensis appears to have very low nutritional requirements for growth. PMID- 11109495 TI - Combined effects of Brassica napus seed meal and Trichoderma harzianum on two soilborne plant pathogens. AB - The effects of soil amendment with rapeseed meal from Brassica napus cv. 'Dwarf Essex' (high glucosinolate concentrations) and 'Stonewall' (low glucosinolate concentrations) on the biological control activity of Trichoderma harzianum towards Sclerotinia sclerotiorum and Aphanomyces euteiches were evaluated. Trichoderma harzianum added to soil reduced myceliogenic germination of S. sclerotiorum by 94%, but did not affect carpogenic germination. In contrast, 100% reduction in carpogenic germination was observed in soil amended with Dwarf Essex meal, along with a 33% reduction in myceliogenic germination. With Stonewall meal as soil amendment, carpogenic germination was reduced by 44% and myceliogenic germination was not affected. Both Dwarf Essex and Stonewall meals inhibited colonization of sclerotia in soil by T. harzianum, from 100% to 0% and 8%, respectively, so that biocontrol activity of T. harzianum was reduced in the presence of either meal. Aphanomyces euteiches root rot of pea was significantly reduced by T. harzianum alone (100%), by amendment with Dwarf Essex meal alone (77%), and by T. harzianum in combination with Dwarf Essex meal (100%). Amendment with Stonewall meal alone did not control root rot, and combination of Stonewall meal with T. harzianum reduced the biocontrol efficacy of T. harzianum. PMID- 11109496 TI - Use of lycorine and DAPI staining in Saccharomyces cerevisiae to differentiate between rho0 and rho- cells in a cce1/delta cce1 nuclear background. AB - In the yeast Saccharomyces cerevisiae, mutants are viable with large deletions (rho-), or even complete loss of the mitochondrial genome (rho0). One class of rho- mutants, which is called hypersuppressive, is characterised by a high transmission of the mutated mitochondrial genome to the diploid progeny when mated to a wild-type (rho+) haploid. The nuclear gene CCE1 encodes a cruciform cutting endonuclease, which is located in the mitochondrion and is responsible for the highly biased transmission of the hypersuppressive rho- genome. CCE1 is a Holliday junction specific endonuclease that resolves recombination intermediates in mitochondrial DNA. The cleavage activity shows a strong preference for cutting after a 5'-CT dinucleotide. In the absence of the CCE1 gene product, the mitochondrial genomes remain interconnected and have difficulty segregating to the daughter cells. As a consequence, there is an increase in the fraction of daughter cells that are rho0. In this paper we demonstrate the usefulness of lycorine, together with staining by 4',6-diamidino-2-phenylindole (DAPI), to assay for the mitotic stability of a variety of mitochondrial genomes. We have found that rho+ and rho- strains that contain CT sequences produce a large fraction of rho0 progeny in the absence of CCE1 activity. Only those rho- mitochondrial genomes lacking the CT recognition sequence are unaffected by the cce1 allele. PMID- 11109497 TI - Genomic relatedness of Staphylococcus aureus phages of the International Typing Set and detection of serogroup A, B, and F prophages in lysogenic strains. AB - On the basis of HindIII-restriction digest analysis of genomic DNAs, the S. aureus bacteriophages of the International Typing Set were divided into five clusters designated as A, F, Ba, Bb, and Bc. The clusters A and F include all the phages of serogroups A and F and correspond to species 3A and 77 proposed by Ackermann and DuBow (1987). On the other hand, the phages of serogroup B were divided into three clusters designated as Ba, Bb, and Bc that differ significantly each from the other in their restriction patterns. The clusters Ba and Bb may represent two separate species, while the cluster Bc may include more than one phage species. For each of the phage serogroups A, B, and F, common HindIII-restriction fragments of phage 3A (1700 bp), of 53 (4060 bp), and of 77 (8300 bp) were used for the preparation of probes specific to the phages of serogroups A, B, and F. These probes were very effective, making it possible to detect up to three different prophages in a given lysogenic strain at the same time. Restriction enzyme maps of phages 3A, 53, and 77, each representing a different serogroup, were constructed. The restriction maps of phage 3A and that of phage 77 are linear, whereas that of phage 53 is circular and exhibits a circular permutation. DNAs of the phages of serogroups A and F have cohesive ends. On each restriction map, the sites corresponding to specific probes are indicated. The size of intact genomic DNA of all phages estimated by PFGE varies within the range of 41.5-46.2 kb. PMID- 11109498 TI - Glucosidase inhibitor from Umbilicaria esculenta. AB - Inhibitory activity toward beta-glucosidase was detected in extracts of the lichen, Umbilicaria esculenta. The extract showed strong inhibition of disaccharide hydrolytic enzymes of mold and mammalian origin, but weak or no inhibition of polysaccharide hydrolytic enzymes except glucoamylase and laminarinase. The inhibitor in the extract was very stable, retaining more than 95% of its activity when treated with heat, acid, alkali, and some hydrolytic enzymes. Purified inhibitor was identified as 1-deoxynojirimycin (1,5-dideoxy-1,5 immino-D-glucitol) by NMR spectrometry, which was known to be produced by Streptomyces sp. and the plant Morus sp. Extracts from Parmelia austrosinensis, Parmelia praesorediosa, and an unidentified lichen species, showed glucosidase inhibitory activities similar to U. esculenta. PMID- 11109499 TI - Genome size of Eperythrozoon suis and hybridization with 16S rRNA gene. AB - The genome size of Eperythrozoon suis, an unculturable haemotropic mycoplasma, was estimated using pulsed-field gel electrophoresis (PFGE). Gamma irradiation was used to introduce one (on the average) double-strand break in the E. suis Illinois chromosome. Restriction enzymes that cut infrequently were also used to analyze genome size. The size estimate for the full-length genome was 745 kilobases (kb), whereas the size estimates based on the summation of restriction fragments ranged from 730 to 770 kb. The 16S rRNA gene was located on the 120-kb MluI fragment, 128-kb NruI fragment, 25-kb SacII fragment, and 217-kb SalI fragment by Southern blotting. PMID- 11109500 TI - Use of lactose to induce expression of soluble NifA protein domains of Herbaspirillum seropedicae in Escherichia coli. AB - Overexpression and purification are procedures used to allow functional and structural characterization of proteins. Many overexpressed proteins are partially or completely insoluble, and can not be easily purified. The NifA protein is an enhancer-binding protein involved in activating the expression of nif and some fix genes. The NifA protein from many organisms is usually insoluble when over-expressed, and therefore difficult to work with in vitro. In this work we have overexpressed the central + C-terminal and the central domains of the Herbaspirrilum seropedicae NifA protein in an Escherichia coli background. Expression was induced with either IPTG or lactose. The data showed that induction with lactose promoted a significantly higher percentage of these proteins in the soluble fraction than with IPTG. This probably reflects a slower kinetics of induction by lactose. PMID- 11109501 TI - Haptoglobin: function and polymorphism. AB - Haptoglobin is an acute phase protein capable of binding haemoglobin, thus preventing iron loss and renal damage. Haptoglobin also acts as an antioxidant, has antibacterial activity and plays a role in modulating many aspects of the acute phase response. There are 3 major haptoglobin phenotypes--Hp(1-1), Hp(2-1) and Hp(2-2). Possession of a particular phenotype has been associated with a variety of common disorders (e.g. cardiovascular disease, autoimmune disorders, malignancy), a fact which can only be explained by the idea that possession of a particular phenotype offers some protection against the development of these disorders. Knowledge of phenotype could therefore aid in the prognosis of disease and allow treatment to be better tailored to suit an individuals' needs. PMID- 11109502 TI - Pediatric reference values of estradiol, testosterone, lutropin, follitropin and prolactin. AB - Reference ranges for lutropin, follitropin, estradiol, testosterone, and prolactin were established by electrochemiluminescence immunoassays (Elecsys 2010, Roche Diagnostics) in 299 children of eight different age groups. The estradiol concentrations did not differ between the genders but dropped significantly in the first few months of life. The testosterone concentrations were higher in male than in female children, whereas the concentrations of follitropin were higher in female than in male children. The concentrations of lutropin were higher in male children (months 1-12) and higher in pubertal girls (years 13-20) in comparison to the corresponding age and gender. PMID- 11109503 TI - A miniaturised semiautomated system for the identification of Yersinia species within the genus Yersinia. AB - Commercially available identification systems based on biochemical reactions of bacteria are not suited for typing the species of the genus Yersinia (Y.) or the biovars (BV) of the species Y. enterocolitica. This failure is caused by the limited number of biochemical reactions applied, resulting in the absence of important discriminatory key reactions. The MICRONAUT identification system (Merlin, Bornheim-Hersel) makes use of dried substrates/enzymes reactions in the wells of a 96-well microtitration plate, reading of the results by a scanner device and typing of the isolate by the calculation of probabilities according to a data base. For this study a special identification panel was designed on which 38 substrates and enzyme reactions were configurated including 20 reactions for the identification of the species of the genus and the Y. enterocolitica biovars. The database was calculated using the results obtained from a total of 250 Yersinia strains of the eleven species of the genus. Reevaluation of the results of these strains revealed an overall sensitivity of 98%, as only four strains were not identified satisfactorily. Considering also questionable results the sensitivity was still 85%. The system was also used to identify Y. pestis isolates, but in this case reading was done visually. The printouts usually cite species designation, identification quality and probabilities. The sealing of the plates in an aluminium bag guarantees long life and long lasting quality. However, an evaluation of the system with a considerable number of strains has to be done in a next step. The 'Yersinia identification set' can replace time consuming tube testing in the future and is a big step forward towards a sensitive identification of Yersinia isolates in the routine laboratory. PMID- 11109504 TI - Falsely increased HbA1c values by HPLC and falsely decreased values by immunoassay lead to identification of Hb Okayama and help in the management of a diabetic patient. AB - A falsely increased HbA1c value of 47.1% was determined by cation exchange chromatography during a routine HbA1c measurement of the blood sample from a 63 year-old diabetic male patient living in Hamburg, Germany. In former determinations by immunological assays falsely decreased values in the range of 5.0 to 5.5% were obtained. The sample was inconspicuous in alkaline hemoglobin electrophoresis. But acid hemoglobin electrophoresis confirmed the falsely increased value. These facts let us consider the existence of a heterozygous "silent hemoglobin variant", such as hemoglobin Okayama, with an amino acid substitution in one of the first four amino acids of the beta chain, representing the epitope of common immunoassays. DNA analyses confirmed this presumption and we found the heterozygous mutation hemoglobin Okayama [beta 2 (NA 2) His (CAC)- >Gln (CAA)]. Knowing that an immunological assay only detects about half of the present HbA1c, the obtained values can be used for therapeutic management of this diabetic patient. PMID- 11109505 TI - Abrupt versus gradual withdrawal of vitamin K antagonist treatment in patients with venous thromboembolic disease: assessment of hypercoagulability and clinical outcome. AB - BACKGROUND: It is yet unclear whether vitamin K antagonist treatment should be stopped abruptly or gradually after an episode of venous thromboembolism. The mode of withdrawal might influence a potential development of a hypercoagulable state, which could influence the risk for recurrent disease. METHODS: We prospectively studied 37 consecutive patients in whom acenocoumarol was discontinued either abrupt (18) or gradually (19) (2/3 and 1/3 of the initial dose for one week). Blood sampling was performed at various time points up to 18 days after complete withdrawal and was analysed for INR, prothrombin fragment F1 + 2 and D-dimer. All patients were clinically followed-up for the assessment of the association between hypercoagulability and occurrence of disease such as recurrent venous thromboembolism or malignancy. RESULTS: An approximately fourfold increase was observed (median increase from 0.3 to 1.3 nmol/l) in the F1 + 2 levels after both abrupt and gradual withdrawal and in the D-dimer concentrations in the abrupt withdrawal group (0.10 to 0.44 mg/l), while those in whom acenocoumarol was discontinued gradually showed a less pronounced increase of the D-dimer levels (0.11 to 0.29 mg/L) (not significant). During follow-up one recurrent venous thromboembolic event occurred in each group, and a diagnosis of cancer was made four times. All these patients had the highest D-dimer concentrations measured in the entire study group. CONCLUSIONS: This study indicates the potential for a hypercoagulable state after acenocoumarol discontinuation, which was not prevented by tapering the acenocoumarol dose. D dimer, measured 2 to 3 weeks after acenocoumarol withdrawal, might be an important tool to identify patients at risk for recurrent venous thromboembolism and/or for the presence of an underlying malignancy. PMID- 11109506 TI - Specific detection of plasmid bearing Yersinia isolates by PCR. AB - A total of 210 isolates belonging to 9 different species of the genus Yersinia (Y.) was investigated with three different PCR assays targeting two plasmoidal genes, the Yersinia adhesin gene (yadA) and the V-antigen gene. The yadA PCR assay described in 1995 by Blais and Phillipe, targeting a Y. enterocolitica specific gene region and a newly designed assay targeting the gene region functionally responsible for autoagglutination, were compared. Both assays identified the same Y. enterocolitica strains. To exclude the possibility that false negative results were obtained due to mutations that had occurred in parallel in both gene regions, a third PCR assay by Neubauer et al. (2000) targeting a conserved region of the V-antigen gene was used as control. Again, DNA of the same Y. enterocolitica strains was amplified. In contrast to the yadA PCR assay described by Blais and Phillipe, the newly established yadA and the V antigen PCR assays amplified DNA from Y. pseudotuberculosis strains. Therefore, by using the PCR technique as a molecular tool spontaneous mutations could be excluded as the cause of anomalous reactions in PCR assays targeting genes of the Yersinia virulence plasmid. Based on these results, it can be assumed that all presumptive pathogenic Yersinia isolates can be identified on the basis of PCR analysis. These molecular assays may also produce fewer false positive reactions in comparison to phenotypic tests such as the autoagglutination test which depend heavily on the handler's experience. It has to be stressed that the PCR assays used in this study have not been evaluated for routine use. Therefore, standardization of the PCR methodology including sample preparation, primer target sequences and PCR reagents is needed for the reliable and safe diagnosis of pathogenic Yersinia spp. in future. PMID- 11109507 TI - Do enzymatic and Jaffe creatinine assays produce comparable results for pre and post dialysis specimens from patients with chronic renal failure? PMID- 11109508 TI - Current diagnostic techniques in genital herpes: their role in controlling the epidemic. AB - Genital herpes continues to be a public health problem in both developed and developing countries. Laboratory confirmation of clinical diagnosis is important, particularly as there are other conditions which present similarly to genital herpes, while atypical presentations of genital herpes also occur. Herpes simplex virus type 2 (HSV-2) is the most common cause of genital herpes globally, although HSV-1 genital herpes infections occur also. Type-specific serology has overcome the technical problems posed by earlier cross-reactive HSV serological assays, hence HSV-2 specific antibodies can be identified using both in-house and commercial assays. All HSV-2 serological assays require an appropriately thorough validation, and here an understanding of the natural history of genital HSV infection is important. Validated HSV-2 specific antibody assays have featured in seroepidemiological studies which have emphasised the largely asymptomatic nature of this infection. Subsequent seroepidemiological studies have included a sexual lifestyle questionnaire to identify risk factors for genital HSV-2 infection. This has given rise to serological screening proposals which, it is argued, may arrest the spread of genital herpes in the general population. However, counter arguments to such proposals are important to consider. As regards diagnosis and management of genital herpes in individual patients, situations have been identified where type-specific serology may be of benefit. PCR has become the method of laboratory diagnosis for HSV encephalitis over the past decade, but its role in the diagnosis of genital herpes has been addressed only in recent years. Evaluation of HSV PCR on specimens from genital herpes cases has shown PCR to be more sensitive than virus culture, the traditional "gold standard" of HSV identification. However, questions remain regarding the acceptance of HSV into routine diagnostic settings, particularly concerning sample preparation, although automation and the ability to include diagnosis of other genital infections in a multiplex PCR is an advantage. Such developments should enhance the role of PCR in genital herpes diagnosis and ultimately reduce costs relative to traditional methods such as culture and HSV antigen detection. Finally, the use of type specific serology and HSV PCR in genital herpes research is noted. PMID- 11109509 TI - An overview of assays for serum HBV DNA. PMID- 11109510 TI - Butylaminiperindopril decreases transforming growth factor-beta 1 messenger RNA production in lungs of C57Bl6 mice after low-dose whole-body irradiation. AB - Transforming growth factor (TGF)-beta is believed to play a key role in the development of many autoimmune and malignant diseases, such as radiation and drug induced organ disease. The aim of the present study was to determine messenger RNA (mRNA) production of TGF-beta 1 in the lungs of C57Bl6 mice after low-dose whole-body irradiation. Control (irradiated) and irradiated angiotensin converting enzyme (ACE) inhibitor-treated animals were simultaneously examined. The ACE inhibitor group received butylaminiperindopril for 9 days after irradiation (7 Gy) at a daily dose of 0.1 mg/kg per rectum. On day 9 all mice were sacrificed and the production of mRNA TGF-beta 1 in lung tissue was determined semiquantitatively using reverse transcriptase polymerase chain reaction. In butylaminiperindopril-treated mice, a decrease in transcript of TGF beta 1 (to 59% in comparison with controls) was observed. PMID- 11109511 TI - Effects of cilnidipine on lipids, lipoproteins and fibrinolytic system in hypertensive patients. AB - Sixteen Japanese patients of both sexes aged 46-78 years with essential hypertension were studied at the cardiac clinic of the Department of Cardiology, Shizuoka General Hospital, Shizuoka, Japan. Serum lipids, lipoproteins, plasma fibrinolytic parameters, renin and noradrenaline were determined before and after 3 months of cilnidipine treatment. Systolic and diastolic blood pressures and heart rate were reduced while renin and noradrenaline levels remained unchanged after cilnidipine treatment. Total cholesterol and tissue plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1) and t-PA-PAI-1 complex were reduced. Changes in the other lipids, lipoproteins and fibrinolytic parameters were not significant after cilnidipine treatment. A negative correlation was found between low-density lipoprotein cholesterol and t-PA antigen levels after cilnidipine treatment. In conclusion, cilnidipine was effective for the treatment of hypertension and did not cause reflex tachycardia in Japanese patients. Cilnidipine treatment produced a beneficial lipid profile (decrease in total cholesterol), but did not show a consistent effect on fibrinolytic parameters in hypertensive patients. The metabolic interaction between beneficial lipid changes and fibrinolysis will be of value to better our understanding of the antiatherogenic effects of cilnidipine treatment in hypertensive patients. PMID- 11109512 TI - Comparison of the effects of felodipine and cilazapril on exercise performance in patients with mild to moderate hypertension. A crossover study. AB - This double-blind crossover study was designed to compare the effects of felodipine and cilazapril on exercise performance in hypertensive patients. After a 2-week placebo run-in period, 40 patients with mild to moderate hypertension were randomized into two parallel groups to receive either felodipine (10 mg) or cilazapril (5 mg) for 4 weeks. After another 2-week washout period, treatments were then crossed over for a further 4-week study period. All patients were given an extensive rest and exercise evaluation at the end of the placebo period. Extensive rest and exercise evaluations were repeated after a 4-week treatment period and again after the second washout period and after the second 4-week treatment period. Before each exercise test, epinephrine, norepinephrine and dopamine plasma levels and plasma renin activity were measured. Two groups were similar at baseline for systolic and diastolic blood pressure and heart rate as well as for laboratory and hormonal variables and duration of exercise test. At the end of treatment diastolic blood pressure was significantly reduced in the felodipine group (p = 0.019). Duration of exercise test was longer than at baseline (p = 0.031) in the felodipine group. Plasma dopamine levels were significantly increased in the cilazapril group. Plasma renin activity significantly increased in the felodipine group. In conclusion, our data show that the two drugs have the same effectiveness in resting conditions but that felodipine is more effective in lowering maximum exercise diastolic blood pressure and in improving exercise time with an double product increase (not significant); it has no statistically significant effect on maximal exercise systolic blood pressure. PMID- 11109513 TI - Evaluation of the effects of zafirlukast 40 mg b.i.d. in addition to preexisting therapy of high-dose inhaled steroids on symptomatic patients with reversible respiratory obstruction: preliminary data. AB - Clinical evidence of the efficacy of zafirlukast is available for mild-to moderate asthma, but further evidence is needed on its use in the more severe forms, uncontrolled by traditional therapy. The aim of this study was to evaluate the efficacy of zafirlukast 40 mg b.i.d. as an add-on to high dose inhaled corticosteroids (HDICs). A secondary aim was to assess the drug's safety and the patients' evaluation of oral therapy and compliance. Twenty-two asthmatic patients (13 females, aged 12-70 years) taking short-acting beta 2-agonists as needed and HDICs as maintenance therapy, entered a 2-week screening phase. At the end, 18 patients with baseline forced expiratory volume (FEV)1 = 50-80% of predicted, a reversibility of > or = 15% and a > or = 10 weekly total of daytime symptom score, were given zafirlukast 40 mg b.i.d. and were reevaluated every 4 weeks for 12 weeks. Adverse events, withdrawals and changes in hematology were recorded. A diary and a questionnaire were used for subjective patient assessment. Sixteen patients (nine females, mean age 40) completed the study. Mean FEV1, (absolute value and percentage vs. predicted) was significantly higher (p < 0.05) vs. baseline after zafirlukast, with a mean improvement of 0.2 l/sec (2.4 vs. 2.2) or 6% (76% vs. 70%) of predicted. Similarly, a significant (p < 0.001) increase in peak expiratory flow was observed (6 vs. 5.5 l/sec). There was no statistically significant variation in forced expiratory flow 25-75. Analysis of the diaries showed a statistically significant reduction (p < 0.05) of morning symptoms (0.73 vs. 4.6) and of daytime symptoms score (2.3 vs. 5.3). There was no evidence of improvement in night time symptoms and use of beta 2-agonists. Three patients experienced adverse events and four suffered a single exacerbation not requiring hospital admission. Subjective evaluation was positive: 75% reported an improvement, 25% found no change and 40% particularly appreciated the oral therapy. In conclusion, treatment with zafirlukast (40 mg b.i.d.) showed a significant improvement in function parameters and symptoms. Zafirlukast was well tolerated and accepted by patients. Further research is needed, as the small number of patients does not allow definitive conclusions to be drawn. PMID- 11109514 TI - [Pharmacological activity of the new adamantane derivative--potential antiparkinson preparation during subchronic administration]. AB - The effect of a new potential antiparkinsonian drug, N-(2 adamantyl)hexamethyleneimine hydrochloride (A-7), upon some behavioral parameters was studied in rats after subchronic 14-day treatment. Also studied was the effect of abrupt termination of a prolonged (22-day) drug administration. The subchronic treatment with A-7 at a dose of 10, 20, or 40 mg/kg did not produce significant changes in coordination, explorative behavior, muscle tone, and aggression level. At the same time, the 14-day treatment with A-7 at a dose of 10 mg/kg increased the pain threshold; A-7 at a dose of 40 mg/kg (but not 10 or 20 mg/kg) increased the spontaneous motor activity. Upon abrupt termination of the drug administration after a prolonged treatment sensitivity of the test animals with respect to the MFTP neurotoxin as compared to that observed in the non-drug control group. PMID- 11109515 TI - [Subtypes and neuronal localization of muscarinic receptors in rat cerebral hemispheres]. AB - The subtypes of pre- and postsynaptic muscarinic receptors in rat cerebral hemispheres were determined using a new approach based on the radioligand analysis. PMID- 11109516 TI - [Effect of diazepam on the metabolic and morphometric parameters of neurons in the dorsal hippocampus in normal and stressed rats]. AB - Diazepam (0.5 mg/kg) decreases the swimming-stress-induced activation of the pyramidal neurons in the dorsal hippocamp. This is manifested by a decrease in the stress-induced glycogen consumption, an increase in the RNA content, and (less pronounced) increased in the nucleocytoplasmic ratio. PMID- 11109517 TI - [Effect of bromantane on the rat neurologic status in two month course]. AB - The oral administration of bromantan for two months on a toxic dose level produced a sex-dependent psychodysleptic action upon rats: the effective (30 mg/kg), intermediate (150 mg/kg), and toxic (600 mg/kg) doses reduced the motor activity in males, while not affecting (or increasing) this activity in females. The effective dose stimulated, and the toxic dose suppressed, the research activity and increased the number of grooming episodes, while ambiguously influencing the emotional state of rats. In the initial stage of treatment, bromantan causes hypothermia; in the second month, this effect is replaced by slight hyperthermia. Prolonged administration of a large dose of bromantan oppressed food uptake and slightly increased drink uptake. Upon the bromantan treatment, the body weight increased in females and decreased in males. The bromantan treatment course increased the muscle strength of rats; the operant activity was optimized during the first month of the course. The general physiological and behavioral characteristics of animals restored within two months after termination of the treatment course. During this period, the test animals exhibited no significant behavioral symptoms indicative of the drug dependence. The two-month treatment did not lead to the development of tolerance with respect to the optimizing drug action upon the physical and operant capacity. PMID- 11109518 TI - [Comparative study of hemodynamic effects of antidepressants (tetrindole and desipramine) during immobilization stress in hypertensive rats]. AB - Tetrindole (a tetracyclic antidepressant, reversible MAO inhibitor of the A type) and desipramine (a tricyclic antidepressant, nonselective inhibitor of the reverse catecholamine trapping) produce opposite hemodynamic effects in stroke prone hypertensive rats (SHR-SP): tetrindole reduced, whereas desipramine increased, the heart rate and arterial pressure. Under the acute immobilization stress conditions in SHR-SP, tetrindole inhibited development of the post stressor tachycardia, while desipramine did not change the heart rate and increased the arterial pressure. PMID- 11109519 TI - [Effects of chronic administration of moxonidine in hypertensive rats SHR-SP]. AB - Study of the hemodynamic response (arterial pressure and heart rate) by radio- and telemetric techniques in stroke-prone hypertensive rats (SHR-SP) aged 6-7 months showed that a chronic oral administration of moxonidine at a daily dose of 2 and 10 mg/kg) leads to hypotension (7 +/- 3 and 21 +/- 5%, respectively) and bradycardia (5 +/- 1 and 14 +/- 5%, respectively). The moxonidine administration at a greater dose violated the circadian heart rate profile, reduced the motor activity of rats by 34 +/- 15%, and showed a drug dependence syndrome in the heart rate (but not in the arterial pressure). It is concluded that moxonidine administration at large doses may give rise to side effects. PMID- 11109520 TI - [Cardioprotective effect of glycyram in myocardial damage induced by isadrine]. AB - Experiments on rats with isadrine induced myocardial disorder showed that glycyrram reduces the extent of myocardial inflammatory edema, inhibits an increase in the level of marker enzymes (creatine kinase, lactate dehydrogenade, asparatate aminotransferase) in the blood serum, prevents a decrease in the myocardial glycogen and an increase in the total lipids, inhibits increase in the lipid peroxidation and decrease in the antioxidant activity in the blood, and improves the ECG characteristics. These data suggest that glycyrram is a promising agent for the treatment of cardiac disorders accompanied by inflammatory and necrotic changes in myocardium, including myocarditis and myocardial infarction. PMID- 11109521 TI - [The use of delta-receptor antagonist DuP 734 for correction of the contractile diastolic cardiac dysfunction in reperfusion injury and oxidative stress]. AB - Intraperitoneal administration of the sigma-receptor antagonist DuP 734 (1 mg/kg) 15 min before heart excision produces a decrease in the reperfusion heart contractility and prevented from the reperfusion induced cardiac cell lesion upon global ischemia of the isolated perfused rat heart. At the same time, a preliminary treatment with DuP 734 potentiated the reperfusion induced suppression of the cardiac pump function, while affecting neither the cardiac contractility dysfunction nor the cardiac cell injury during the oxidative stress. It is concluded that DuP 734 is not effective in preventing the myocardial stunning. The cardio-protector effect of DuP 734 during reperfusion is not related to inhibition of the free radical cell damage. PMID- 11109522 TI - [Effect of the dry aspen bark extract on the gastric secretory function]. AB - Dry aspen (Populus tremula L.) bark extract exhibits antiulcerogenic activity as demonstrated by the results of experiments using the models of gastric ulcers in rats according to H. Shay. The cytoprotective action of the dry aspen bark extract was also demonstrated in experiments on dogs with fistula according to Basov. PMID- 11109523 TI - [Effects of bemethyl, ethomersol, and yakton on the liver regeneration after partial hepatectomy]. AB - It is experimentally demonstrated for the first time that the new drugs bemithyl, etomerzol, and yakton are capable of accelerating the process of liver regeneration following partial hepatectomy. The drugs produce a hasty gain in the mass of liver, increase in the content of nucleic acids and glycogen, and improve the functional state, as manifested by a decrease in the blood bilirubin and a reduction in the hexenal sleep duration. Bemithyl, etomerzol, and yakton produce a positive effect upon the liver morphology and the intracellular regeneration process. The repair activity of the new drugs exceeds that of a combination of the well-known regeneration stimulants riboxin and potassium orotate, representing derivatives of purine and pyrimidine bases. PMID- 11109524 TI - [Experimental study of hemostimulating properties of the tablet form of recombinant human erythropoetin]. AB - Specific biological activity of the domestic preparation of recombinant human erythropoietin (RHEPO) in an original pill form was experimentally studied in vitro and in vivo. Administration of the RHEPO parent substance at a concentration of 0.5 10 U/ml significantly accelerated the growth of erythroid colony-forming cells (CFU-E) in the culture of intact nonadherent myelokaryocytes. The erythropoiesis-stimulating properties of the new RHEPO preparation are comparable with those of the well-known injection preparations. The drug action is related to increasing the erythroid cell content in the bone marrow and the reticulocyte and erythrocyte counts in the peripheral blood. PMID- 11109525 TI - [Anti-aggregant activity of the blood vessel wall and its regulation by GABAergic agents in hypokinesia]. AB - Restricted motor activity on the background of platelet hyperaggregation in the whole blood and plasma leads to inhibition of the antiaggregant activity of the blood vessel wall, thus creating prerequisites for the development of thromboembolic disturbances. Under these conditions, GABA and piracetam produce a pronounced protective action with respect to the platelet-vessel wall interactions. PMID- 11109526 TI - [Effect of oral antidiabetic agents on fructose biosynthesis]. AB - The oral antidiabetic drugs (glyformin, glyclaside, glycvidon, glybenclamide) at a concentration of 10-196 mM affect the synthesis of fructosamine in an incubation medium containing 40 mM of glucose and 5% of human serum albumin. All these drugs decrease the fructosamine yield measured in the medium on the 4th day of incubation at 37-198 degrees C, while the effect of glycvidon is pronounced even on the 7th day. The introduction of glyformin, glycvidon, and glybenclamide at an amount close to the maximum daily dose over a period of one month reduces the level of fructosamine in the blood of rats with experimental diabetes mellitus, while not affecting the level of glucose in the blood of test animals. Thus, the oral antidiabetic drugs reduce the level of fructosamine--an agent known to modify the protein structure, thus favoring the development of complications in the course of diabetes under permanent hyperglycemia conditions. PMID- 11109527 TI - [Effect of the cholinesterase reactivator dipyroxime in various models of delayed hypersensitivity during acute intoxication by acrylonitrile]. AB - Delayed type hypersensitivity (DTH) response with respect to sheep erythrocytes was studied on various models in CBA mice against the background of acute intoxication with nitrile acrylic acid (NAA). The effect of dipiroxime on the DTH response under these conditions was determined and the relationship of these reactions with the activity of alpha-naphthylbutyratesterase in splenic cells and popliteal lymph nodes was assessed. Dipiroxime partly recovered DTH in various experimental series (except for the reaction of suppressor cell transfer) by restoring the alpha-naphthylbutyratesterase activity in cells of the lymphoid organs studied. PMID- 11109528 TI - [Immunomodulating effects of vitamin K preparations and its potentiation by riboxine in acute cold stress]. AB - The vitamin K preparations phylloquinone, menadione, and vikasol produce an immunomodulant effect under acute cold-induced stress conditions. A combination of phylloquinone, menadione and riboxin provides an effective means of correcting the immunological reactivity and antioxidant state under the conditions of immersion cooling. The immunomodulant effect of naphthoquinones and riboxin is correlated with their antioxidant activty. PMID- 11109529 TI - [Effect of 2-methyl-4-amino-6-hydroxypyrimidine on the functional state of the body exposed to extreme conditions]. AB - The experiments on mice showed that 2-methyl-4-amino-hydroxypyrimidine (MAHP) exhibits a pronounced immunostimulant action upon the animal organism under extremal conditions. The effect of MAHP is related to the preferential activation of immunocompetent cells responsible for the immunity. PMID- 11109530 TI - [Effect of ammonium succinate on pharmacological effects of acetylsalicylic acid]. AB - Ammonia succinate potentiates the main pharmacological properties and reduces the toxic effects (ulcerogenic action and general toxicity) of acetylsalicylic acid. The new preparation astam, representing a combination of acetylsalicylic acid with ammonia succinate in a 2:1 ratio, is proposed. Astam exhibits antiexudative, capillary-reinforcing, antiproliferative, pain-relieving, antipyretic, antiaggregant, and antioxidant properties. In addition, the drug inhibits the development of structural-metabolic disorders in the case of chronic immune inflammation of joints and various internal organs. PMID- 11109531 TI - [Medicinal plant preparations used as adjuvant therapeutics in experimental oncology]. AB - Experiments on mice inoculated with metastasing Lewis lung carcinoma showed that the antitumor and antimetastatic effects of cyclophosphan (cyclophosphamide) are potentiated by the extracts of phytopreparations based on Baikal scullcap (Scutellaria baikalensis), rhodiola (Rhodiola rosea), common licorice (Glycyrrhiza glabra), and their principal acting components--baikalin, paratyrosol, and glycyrram. PMID- 11109532 TI - [Effects of rumalon and arteparon correcting metabolism in the connective tissue on the functional state of the liver in toxic hepatitis]. AB - Effects of the connective tissue metabolism correctors rumalon (a glycosaminoglycane peptide complex) and arteparon (glucosamine sulfate) on the cholepoietic and absorption-excretion functions of liver were studied on a model of toxic liver damage. Rumalon exhibits pronounced anticholelithiasis and cholagogic properties and improves the absorption-excretion liver function. Arteparon produced no cholagogic action, inhibited the elimination of bromosulfalein, and showed no anticholelithiasis activity. The difference in the pharmacological effects of two drugs is related to their structural distinctions and the differing characteristics of aglycons and anions entering into their compositions. PMID- 11109533 TI - [Effect of the intravenous laser blood irradiation on efficacy of drug preparations]. AB - The effect of the intravenous laser blood (ELB) treatment on the sensitivity of blood components with respect to drugs was studied in patients with nonspecific reactive hepatitis and chronic hepatitis. An ELB course reduced the functional activity of thrombocytes in the presence of fibrinogen and adrenaline hydrochloride (collagen inductors), which must be taken into account when these drugs are used as hemostatic agents. PMID- 11109534 TI - [Therapeutic effect of hepatoprotectors in experimental Reye syndrome]. AB - Maxar and legalon--hepatoprotectors containing polyphenols--exhibit a therapeutic effect with respect to the experimental Reye's syndrome induced in rats by intraperitoneal injections of 4-pentenoic acid. Maxar restores the activity of enzymes of the hepatic origin, normalizes the content of bilirubin, glucose, phenols, and malonic aldehyde in the blood serum, stimulates the production of ketone bodies and ammonia detoxication, and improves the histologic structures of liver and cortex. Legalon also decreases the structural-metabolic disorders accompanying the Reye's syndrome, but to as lower ewxtent. PMID- 11109535 TI - [Effects of pikamilon on the brain water-electrolyte balance and in models of responses to the noise and vibration exposure, and their combination with motion sickness]. AB - Electric impedance measurements on awake rabbits with electrodes implanted into cortex, thalamus, and hypothalamus showed that modeling of the noise and general (wide-band) vibration, as well as their combination with sea-sickness action, leads to disturbances in the aqueous-electrolyte balance in brain. This is manifested by extracellular and (less pronounced) intracellular edema development. Picamilon administration (10 mg/kg, i.v.) completely prevented development of the vibration-induced intracellular edema and markedly reduced the development of damage in all other cases studied. PMID- 11109536 TI - [Drug side effect or negative placebo effect?]. PMID- 11109537 TI - Establishment and characteristics of four sub-strains of F344 rats reared on various low protein and low energy diets. AB - Four sub-strains, reared by sib-mating and having for their origin the F344/DuCrj strain of rats, were established by feeding with different levels of low protein and low energy diets, and their characteristics investigated. The amounts of crude protein (CP) and digestible energy (DE) in the four diets were 17.6%-3.0 kcal, 10.5%-2.5 kcal, 8.4%-2.0 kcal, and 10.5%-2.5 kcal, respectively, and the four sub-strains established here were provisionally designated as F344/Tig1, F344/Tig2, F344/Tig3 and F344/Tig4, respectively. Intakes of nitrogen-corrected metabolizable energy (MEn) did not differ, and a large intake of digestible crude protein (DCP) was observed in F344/Tig1 rats. The body weight of rats provided with lower-nutrient diets showed a tendency to decrease until the F2 generation, but no change among the generations was seen subsequently, and the same compiled differences in protein content were maintained. Similar transitions were observed in the lifetime rearing test. Though F344/Tig3 rats, which were reared on minimum nutrients, showed a tendency to delayed puberty, we were easily able to breed four pairs in every generation using procedures similar to those used for other strains of rats. There were no differences among the F344/Tig1 to -3 strains of rats in body length, digestive tract length, or organ weight per body weight, and all the rats had a normal range of biochemical values. But the F344/Tig4 showed a high glutamic-oxaloacetic transaminase (GOT), and a tendency to decreased liver function and a shorter lifespan. These sub-strains of F344 rats clarified differences in fatty acid compositions, such as docosahexaenoic acid (DHA) in serum, liver and the brain. The rats were intended to be useful animal models for the study of nutritional environments and their influence on the memory and learning. PMID- 11109538 TI - Effects of aging on bone mineral content and bone biomarkers in female cynomolgus monkeys. AB - Age-related changes in bone mineral content and bone biomarkers were assessed over the complete lifespan of female cynomolgus monkeys. The bone mass of the lumbar spine increased linearly from birth to about 2.5 years of age, and this increase gradually slowed thereafter until a peak bone mass was achieved at 9 years of age. The bone mass stabilized after 9 years of age, showing no sign of further reduction with age. In contrast with the significant increase in bone mass before 2.5 years of age, significant decreases occurred in the serum concentrations of the following bone formation markers: intact osteocalcin, bone specific alkaline phosphatase and amino-terminal propeptide of type I procollagen, but the serum concentration of carboxy-terminal propeptide of type I procollagen did not change significantly throughout the entire lifespan. Concerning the bone resorption markers, the levels of tartrate-resistant acid phosphatase fluctuated throughout the entire lifespan. The skeleton of an aging female monkey undergoes changes similar to those observed in senescent humans, but did not undergo the menopausal changes seen in women. The use of female cynomolgus monkeys to model human skeletal interventions should therefore be undertaken with consideration of the similarities and differences between cynomolgus monkeys and humans. PMID- 11109539 TI - Functional natural killer T cells in experimental mouse strains, including NK1.1- strains. AB - Natural killer T (NKT) cells are a newly discovered subset of lymphocytes. It appears that this subset has potential as important regulators of immune responses. But because there are relatively few NKT cells in lymphoid organs and because of technical difficulties in detecting NKT cells in most mouse strains, the roles of NKT cells have not been fully identified and little attention has been paid to the roles of NKT cells in immunological experiments in which NK1.1- strains were used. To examine the existence of functional NKT cells in various strains of experimental mice, including NK1.1- strains, we utilized alpha galactosylceramide (KRN7000) which is thought to react specifically with NKT cells. Indeed, we could confirm that early cytokine (IL-4 and IFN-gamma) secretion at 2 h after the injection of KRN7000 was dependent on NKT cells. With this in vivo system, we have successfully detected the presence of functional NKT cells in various mouse strains, including AKR/N, BALB/c, C3H/HeJ, C3H/HeN, C57BL/6, C.B-17, CBA/N, NC, NOD, SJL, W/Wv, aly/aly and aly/+. Notable increases of serum IL-4 were detected in W/Wv and aly/+ strains, and defective response of IFN-gamma in SJL mice and that of IL-4 in NOD mice were observed. This is the first report to show the functional significance of NKT cells in cytokine secretion in various mouse strains in response to a ligand for the T cell receptor of NKT cells. PMID- 11109540 TI - Teratologic studies on rat perinates and offspring from dams treated with ethylnitrosourea (ENU). AB - Ethylnitrosourea (ENU), a well known DNA alkylating agent, induces anomalies in the central nervous system (CNS), craniofacial tissues, limbs and male reproductive organs. Recently we clarified that excess cell death caused by apoptosis occurred in these organs and tissues of rat fetuses from dams treated with ENU at day 13 of gestation (GD13). In this study, we examined fetuses at GD21 and offspring at 10 weeks of age after ENU administration to pregnant rats at GD13 in order to clarify the relationship between ENU-induced apoptosis in the fetal tissues and teratogenicity of ENU. Severe intrauterine growth retardation was observed in the ENU group, and the body weight of the offspring in the ENU group was significantly lower than that of the control group throughout the experiment. In addition, a high incidence of microencephaly, ectrodactyly and curved caudal vertebrae was observed in the offspring from dams treated with ENU at GD13. Judging from the results of our previous and present studies, it was strongly suggested that ENU-induced apoptosis in rat fetal tissues may play an important role in the induction of anomalies in the corresponding tissues. PMID- 11109541 TI - Generation of a polymorphic marker linked to thymoma susceptibility gene of rat 1 by genetically-directed representational difference analysis. AB - BUF/Mna (BUF) is a rat strain susceptible to spontaneous development of thymomas. We have previously shown that the thymoma susceptibility is controlled principally by a dominant susceptibility gene located on chromosome 7, thymoma susceptibility gene of rat 1 (Tsr1). To generate genetic markers tightly linked to Tsr1, we performed genetically directed representational difference analysis (GDRDA) with three combinations of the tester and driver DNAs. From 124 ?ACI/NMs x (BUF x ACI/NMs) F1? backcross rats, 12 rats with the ACI/BUF genotype in the Tsr1 region (A/B rats) and 13 rats with the ACI/ACI genotype in the region (A/A rats) were selected, and their DNAs were pooled, respectively. Three kinds of tester DNAs, i) inbred BUF, ii) (BUF x ACI)F1, and iii) the pool from the A/B rats, were subtracted by the driver DNA prepared from the pool of the A/A rats. The three combinations yielded one, two, and one polymorphic marker(s), respectively. One marker, D7Ncc28, was isolated commonly by the three combinations of subtraction, and another marker, D11Ncc12 was isolated only by the second combination. Linkage analysis demonstrated that D7Ncc28 was located in the 8.3 cM region where Tsr1 has been mapped. The three combinations of subtraction were shown to be almost equally capable of isolating polymorphic markers in a specific chromosomal region. PMID- 11109542 TI - Localization and age-related changes in cytochrome P450 expression in APA hamster livers. AB - To establish the baseline data, age-related changes and the regional expression of the hepatic P450 isozymes in Syrian hamsters of the APA strain at 3, 6, 12, 18 months old were examined by immunological techniques. Immunohistochemical analysis of liver serial sections revealed that the midzonal and perivenous regions (zones 2 and 3, respectively) were stained with the anti-rat CYP1A1/2, 2B1/2 and 2E1 antibodies. These three antibodies most intensely stained the hepatocytes around the central vein. An anti-rat CYP3A2 staining section had a staining pattern with equally intense reactions in zones 2 and 3. On the other hand, CYP2C6, 2C11 and 4A1 were distributed diffusely throughout the hepatic acinus. There was no age-related difference in the expression pattern of any of the P450 isozymes examined. Total P450 content had a peak at 6 months of age and decreased to 60% of that level thereafter. Western-blot analysis revealed that the peak expressions of the isozymes detected with anti-rat CYP1A1/2, 2C6, 2E1 and 3A2 antibodies were observed in 6-month-old hamsters and declined in older ones. The CYP2B and 2C11 content reached the maximum at the age of 6 months and maintained almost the same level thereafter. The CYP4A level did not change from 3 to 6 months, and then declined to about 40% of the younger level at 12 and 18 months of age. These results suggest that the hepatic P450 isozymes of APA hamsters have region-specific expressions and most isozymes have their peaks of expression at 6 months of age, which differs from the patterns for rat P450. PMID- 11109543 TI - Marking behavior is innate and not learned in the Mongolian gerbil. AB - We studied whether marking behavior in Mongolian gerbils would be innate or learned behavior. The marking behavior was defined as "animals rubbing their abdominal scent glands on small protruding objects". Between 21 and 90 days of age, Mongolian gerbils, which were kept under such conditions that they would be unable to learn this behavior, were observed at intervals of 5-15 days to find out if there were signs of the behavior or not. Six male and four female Mongolian gerbils were used for observing. Neonate Mongolian gerbils during the age of 3 to 28 days were fostered by ICR mother mice. Weaning Mongolian gerbils were then individually kept away from the others. Marking behavior was observed in 2 out of 6 males at 50 days of age and 2 of 4 females at 60 days and the mean frequency of the marking behavior for 10 min was 3.5 in the males and 5.0 in the females. These results suggest that marking behavior was innate and not learned behavior in Mongolian gerbils. PMID- 11109544 TI - Hematological characteristics of rats spontaneously developing eosinophilia. AB - Hematological and genetic characteristics of newly found eosinophilic rats were studied. Hematologically, high blood eosinophil counts started at 6 weeks of age. Almost all 10-week-old rats had eosinophilia with individual counts above 500/microliter and 5 to 100 times the normal level. Proliferating eosinophils had normal morphology. An increase in lymphocyte counts was observed at 5 weeks of age, one week earlier than the onset of eosinophilosis. In bone marrow, proliferation of eosinophils was also observed at 8 weeks of age and thereafter progressed, suggesting a role in the pathogenesis of eosinophilia in this rat. The results of genetic cross experiments revealed the disease to be hereditary. The spontaneously eosinophilic rat therefore warrants attention as a model for studying the underlying mechanisms of human and animal eosinophilia. PMID- 11109545 TI - Distribution of body weight, blood insulin and lipid levels in the SMXA recombinant inbred strains and the QTL analysis. AB - In the SMXA recombinant inbred (RI) strains, we measured body weight, blood insulin and lipid (triglyceride, total cholesterol and phospholipid) levels in each strain. In the five traits, mean values of substrains varied remarkably and showed a continuous spectrum of distribution, suggesting control by multiple genes at distinct loci for each trait. We also screened for quantitative trait loci (QTLs) involved in the five traits. Suggestive QTLs for body weight (Chromosomes 1 and 6), insulin (Chromosomes 1, 3, 10 and 17), triglyceride (Chromosomes 4 and 11) and phospholipid (Chromosome 18) levels were detected. The SMXA RI strains are unique tools for analyzing genetic factors that influence body weight, blood insulin and lipids levels. PMID- 11109546 TI - Squamous cell carcinoma of the oral cavity in an infant cynomolgus monkey. AB - Squamous cell carcinoma was observed in the oral cavity in a one-year-old male cynomolgus monkey. Histopathologically, the tumor consisted of various shaped cells and its assemblies infiltrated into the surrounding connective tissues. Although no obvious metaplastic keratinized cancer pearls were found in the tumor cells, the intercellular bridges were observed. Immunohistochemically, tumor cells were stained with anti-keratin, but not with anti-vimentin. On virological examinations, no papilloma virus antigen or Epstein-Barr Virus small mRNA could not be detected. Under the electron microscope, incomplete tonofibrils and desmosomes in the cytoplasm and microvillus of the cell membrane were observed, suggesting a malignancy or low differentiation of the tumor cells in the present case. This is the first case of squamous cell carcinoma observed in very young macaques, to our knowledge. PMID- 11109547 TI - Generation of transgenic rats with YACs and BACs: preparation procedures and integrity of microinjected DNA. AB - The aim of the present study was to investigate differences in the methods for preparing a large DNA fragment to be used for making transgenic rats from the standpoint of transgenic production efficiency and integrity of the introduced gene. In yeast artificial chromosome (YAC) transgenesis, three methods for preparing DNA for microinjection were compared: amplification of YAC in yeast (AMP), amplification of YAC in yeast and removal of the amplification element (AMP/RE), and no amplification of the YAC in yeast (AMP-). Production efficiency per microinjected ovum with DNA by the AMP method was four times higher than that by the AMP/RE and AMP-. Based on these results, we favor the AMP method in spite of the thymidine kinase gene-induced male sterility. In bacterial artificial chromosome (BAC) transgenesis, linear DNA fragments for microinjection prepared by three kinds of purification procedures were compared: Not I digestion and CsCl gradient ultra-centrifugation (Prep. 1), CsCl gradient ultra-centrifugation, Not I digestion, gel electrophoresis, and beta-agarase digestion (Prep. 2), and CsCl gradient ultra-centrifugation, Not I digestion, pulse field gel electrophoresis, and beta-agarase digestion (Prep. 3). Although the efficiency of producing transgenic rats was similar with all these three DNA preparations, integration of the intact DNA fragment only occurred with the Prep. 3 procedure. We therefore favor the Prep. 3 method for preparing BAC DNA fragments. These results indicate that the method used to prepare a large DNA fragment such as YAC and BAC DNAs is important in order to produce transgenic rats with an intact transgene. PMID- 11109548 TI - Digging behavior of ddY mouse. AB - In the present studies, the behavior of ddY mice digging wood chips was carefully observed. When mice were individually placed on new 5 cm-thick wood chips, their behavior was found to be the same irrespective of their age or sex. The behavior was not prevented by non-noxious 5 black steel rods which were used to measure digging ability, and was not related to habituation or learning. But moist or dirty chips remarkably weakened digging ability. These findings strongly suggest that the digging behavior is a natural and instinctive one, but not an expression of anxiety as previously reported. PMID- 11109550 TI - Replication of enterotropic and polytropic murine coronaviruses in cultured cell lines of mouse origin. AB - To understand the virus-cell interactions that occur during murine coronavirus infection, six murine cell lines (A3-1M, B16, CMT-93, DBT, IC-21 and J774A.1) were inoculated with eight murine coronaviruses, including prototype strains of both polytropic and enterotropic biotypes, and new isolates. All virus strains produced a cytopathic effect (CPE) with cell-to-cell fusion in B16, DBT, IC-21 and J774A.1 cells. The CPE was induced most rapidly in IC-21 cells and was visible microscopically in all cell lines tested. In contrast, the coronaviruses produced little CPE in A3-1M and CMT-93 cells. Although most virus-infected cells, except KQ3E-infected A3-1M, CMT-93 and J774A.1 cells, produced progeny viruses in the supernatants when assayed by plaque formation on DBT cells, the kinetics of viral replication were dependent on both the cell line and virus strain; replication of prototype strains was higher than that of new isolates. There was no significant difference in replication of enterotropic and polytropic strains. B16 cells supported the highest level of viral replication. To determine the sensitivity of the cell lines to murine coronaviruses, the 50% tissue culture infectious dose of the coronaviruses was determined with B16, DBT, IC-21 and J774A.1 cells, and compared to that with DBT cells. The results indicate that IC 21 cells were the most sensitive to murine coronaviruses. These data suggest that B16 and IC-21 cells are suitable for large-scale preparation and isolation of murine coronaviruses, respectively. PMID- 11109549 TI - Effects of chronic administration of sibutramine on body weight, food intake and motor activity in neonatally monosodium glutamate-treated obese female rats: relationship of antiobesity effect with monoamines. AB - When the hypothalamic ventromedial nucleus and arcuate nucleus were destroyed in rats by treatment with monosodium glutamate in the neonatal stage, increase in the Lee index (body weight 1/3/body length) and in retroperitoneal fat as well as decreases in spontaneous motor activity, food consumption and growth hormone secretion function associated with hypothalamic low body length obesity (monosodium glutamate-treated obesity; MSG-OB) were observed as these rats grew. Treatment with sibutramine at 3 and 10 mg/kg p.o. once a day continuously for 14 days improved these parameters, and the degree of improvement was dose related. The plasma lipid values in MSG-OB rats, which were the same as those in normal rats, were decreased by consecutive administration of sibutramine. Levels of hypothalamic monoamines (MAs) such as norepinephrine, 5-HT (serotonin) and dopamine and their metabolites DOPAC, HVA and 5-HIAA were decreased in MSG-OB rats, and further decrease in them, though slight, was observed with consecutive daily administration of sibutramine, probably as a result of the feedback attributable to an increase in MA in synapses caused by inhibition of MA uptake by sibutramine. These results suggest that sibutramine can activate the MA nervous system by MA uptake inhibition in regions of the brain such as the lateral hypothalamic area and the paraventricular nucleus, which control food intake and sympathetic nerve activity, and the nigrostriatal area related to the extrapyramidal motor system, and thereby exhibit anti-obesity effects in the MSG OB rat. PMID- 11109551 TI - Aortic atheromatous lesions developed in APA hamsters with streptozotocin induced diabetes: a new animal model for diabetic atherosclerosis. 1. Histopathological studies. AB - To develop an adequate animal model for atherosclerosis in large vessels of patients with diabetes, i.e. diabetic macroangiopathy, we induced diabetes in APA hamsters with a single injection of streptozotocin (SZ) and examined the aorta histopathologically and immunohistochemically. As a result, hyperglycemia and hyperlipidemia were continuously observed for 26 weeks after the SZ injection (WAI) in APA hamsters. Fatty streaks characterized by a subendothelial accumulation of many foam cells were observed, limited to the aortic arches as early as 6 WAI. In addition to larger fatty streaks developing with the duration of diabetes, fibrous plaques and plaques containing calcium deposits or cholesterol clefts developed at 26 WAI. These lesions are generally similar to the atheromatous lesions developed in humans. Moreover, depositions of apolipoprotein E and advanced glycation end-products immunohistochemically detected in the lesions were very similar to those found in humans. The diabetic APA hamster is therefore considered to be a useful model for studying the formation of atheromatous lesions in diabetic patients. PMID- 11109552 TI - APA hamster model for diabetic atherosclerosis. 2. Analysis of lipids and lipoproteins. AB - Syrian hamsters of the APA strain (APA hamsters) have recently been shown to have atheromatous lesions in the aortic arches under diabetic condition induced by a single injection of streptozotocin (SZ). In that model, fatty streaks, which are the initial lesions of atherogenesis, develop by 6 weeks after the injection (WAI). In this study, we evaluated plasma lipid concentrations and lipoprotein profiles in diabetic APA hamsters at 6 WAI to reveal the early stage of atherogenesis clinicopathologically. As a result, by biochemical analysis, hyperglycemic APA hamsters showed signs of hypercholesterolemia and hypertriglyceridemia. Low-density lipoprotein (LDL) cholesterol significantly increased, but high-density lipoprotein (HDL) cholesterol significantly decreased. Agarose gel electrophoresis showed an obvious increase in the fractions of chylomicron, LDL and abnormal lipoprotein. Plasma LDL in diabetic animals was in a state more susceptible to oxidization. In addition, a significant increase in glycated LDL was also found in the diabetic animals by enzyme linked immunosorbent assay (ELISA). Moreover, lipid peroxidation product (4-hydroxynonenal (4 HNE))-adducted proteins and advanced glycation end-products (AGE) were immunohistochemically detected in the foam cells of the fatty streaks. These results revealed that diabetic APA hamsters had hyperlipidemia characterized by increases in chylomicron, LDL and abnormal lipoprotein, and suggested that oxidized LDL and/or glycated LDL might be actively uptaken by macrophages and play an important role in the initial stage of atherogenesis. PMID- 11109553 TI - Ocular fundus abnormalities detected by fluorescein and indocyanine green angiography in the Royal College of Surgeons dystrophic rat. AB - The ocular fundi of the Royal College of Surgeons (RCS) dystrophic rats were examined by conventional fundus photography, fluorescein angiography (FA) and indocyanine green angiography (IA). In the fundus, a reddish colored background was observed in the RCS dystrophic rats at 3 weeks of age. At 9 weeks of age, the background had changed to pale in color. In FA, the RCS dystrophic rats at 3 week of age demonstrated background fluorescence with homogeneous brightness. Fluorescent dye leakage was observed in the late phase of the postinjection period at 9 weeks of age. In IA, the RCS dystrophic rats at 3 weeks of age had background fluorescence with homogeneous brightness, and at 5 weeks of age, spots of hyperfluorescence were scattered over the dark background. At 7 weeks of age, numerous delimited, irregular round spots of hyperfluorescence appeared over the dark background. Such hyperfluorescent lesions had further increased in number and size in the RCS dystrophic rats at 9 weeks of age. In this way, ocular findings related to abnormalities in the retinal pigment epithelium and choroid in the RCS dystrophic rat were demonstrated by fundus photography, fluorescein angiography and indocyanine green angiography. PMID- 11109554 TI - Influence of age on duodenal brush border membrane and specific activities of brush border membrane enzymes in Wistar rats. AB - To examine age-related changes in the morphology of intestinal brush border membrane (BBM; microvilli) and specific activities of intestinal BBM enzymes including alkaline phosphatase (ALP), gamma-glutamyl transpeptidase (gamma-GT), and disacchridase, four groups of Wistar rats were sacrificed at 2.5 wk, 5 wk, 5 mon and 23 mon. In an electron microscopic examination, morphologically a less dense BBM structure in the duodenum of rats aged 23 mon was observed than that of rats aged 5 mon. Specific activity of ALP in the duodenum from 5-mon-old rats was significantly higher than from rats aged 2.5 wk and 23 mon. The mucosal tissues from 5-wk-old rats had significantly higher specific activity of gamma-GT than did tissues from the other ages. In sucrase and maltase specific activities, 5 mon-old rats had higher activities of these enzymes than other age groups, especially 2.5-wk- and 23-mon-old rats. There was also a significant effect of site on intestinal BBM enzyme activities in post-weanling rats. Regional gradients of ALP and gamma-GT along the entire small intestine (duodenum > jejunum > ileum) were remarkable. Disaccharidase activities peaked in the jejunum and declined toward both the duodenum and ileum. Taken together the result obtained here suggested that 5-mon-old rats had the most elevated intestinal function. This result also strongly indicated that the structure of the intestinal BBM and development of intestinal BBM enzymes in Wistar rate were markedly influenced by age during the postnatal period. PMID- 11109555 TI - Ultrasonic isolation calls in genetically high- and low-emotional rat pups. AB - The present study was undertaken to investigate the relationship between the emotionality and the modulation of ultrasonic vocalizations in Tsukuba High Emotional (THE) and Tsukuba Low Emotional (TLE) strain rat pups 3-18 days old. The THE pups, while isolated from their dam and littermates and placed in a cold environment, emitted ultrasonic isolation calls at a high rate until day 15. In contrast, ultrasound production was at a consistently low level throughout the test period in the TLE pups. The ultrasonic isolation calls of THE pups were attenuated to the same level as those of the TLE pups after administration of diazepam (1 mg/kg, s.c.), a benzodiazepine receptor agonist, at 6 and 12 days of age. These findings suggest that the high emotionality of the rat pups was reflected largely by the emission of ultrasounds in response to isolation distress rather than the number of the benzodiazepine receptors in the brain that might play a role in physiologic mediation of the rat pup isolation call. PMID- 11109556 TI - A parallel comparison of age-related peripheral nerve changes in three different strains of mice. AB - Strain-specific differences contributing to spontaneous age-related peripheral nerve changes were examined in three different strains of 100-week-old female mice housed under the same conditions over the same period: inbred C57BL and C3H strains, and the hybrid B6C3F1 strain. A lower incidence of obesity and significantly lower body weight, grasping power of fore- and hind-limbs, blood lipid level, tail-flick latency and motor nerve conduction velocity were observed in C57BL mice; significantly lower body temperature, blood glucose and HbA1c levels were observed in C3H mice. Histological examination conducted on isolated sciatic nerves and brachial plexuses revealed peripheral nerve lesions, characterized by axonal degeneration and remyelination, in all strains. Although the extent of histopathologic change in nerve fibers was similar in quality to those observed in all three mouse strains, the incidence and severity of nerve lesions in B6C3F1 and C3H mice were significantly greater than those observed in C57BL mice. PMID- 11109557 TI - Comparison of the effects of two fixatives for immunolocalization of testosterone in the testes of the cynomolgus monkey, mouse and rat. AB - We compared the effect of two fixatives, Bouin's fixative and neutralized buffered 4% formaldehyde (10% formalin), for immunolocalization of testosterone in the testes of cynomolgus monkeys, mice and rats. In the samples fixed with Bouin's fixative, immunoreactive testosterone was detected as intense deposits in the cytoplasm of Leydig cells of monkeys and mice. Immunoreactive testosterone was detected not only in Leydig cells of rats but also moderately shown within tubules. Immunoreactive testosterone could not be detected in the testes of monkeys, mice or rats fixed with neutralized buffered formalin because of the poor morphology caused by the fixative. It is concluded that Bouin's fixative is a suitable fixative for immunolocalization of testosterone in the testes of cynomolgus monkeys, mice and rats. PMID- 11109558 TI - Inhibitory effect of apple pectin and culture condensate of Bifidobacterium longum on colorectal tumors induced by 1,2-dimethylhydrazine in transgenic mice harboring human prototype c-Ha-ras genes. AB - The number and tumor score of colorectal tumors induced by 1,2-dymethylhydrazine in transgenic (Tg) mice carrying human c-Ha-ras genes were significantly reduced by ingestion of apple pectin (AP) or a culture condensate of Bifidobacterium longum (MB) when compared with a control diet. There was no statistical difference in the incidence of colorectal tumors in Tg mice between the AP or MB diet and the control diet. This study demonstrated that Tg mice are a useful tool for screening inhibition of colorectal tumors by functional foods. PMID- 11109559 TI - Detection and elimination of contaminating microorganisms in transplantable tumors and cell lines. AB - As a quarantine of biological materials, we tested 96 transplantable tumors and cell lines for contamination with microorganisms in a mouse antibody production (MAP) test, enzymatic assay and microbiological culture. Contamination with lactic dehydrogenase elevating virus (LDV), mycoplasmas and Pasteurella pneumotropica was detected. A considerable difference in the contamination rate was observed between in vivo- and in vitro- propagated tumors. LDV in the tumors could be eliminated by both in vitro subculture and subpassage in nude rats. Mycoplasmas were eliminated by means of the mycoplasma-removal agent and P. pneumotropica by subpassage in mice. These results suggest that there is still a high risk of contamination in transplantable tumors and emphasizes the importance of adequate microbiological quality control. PMID- 11109560 TI - Running hamster: a new type of mutant characterized as speedy, rotative running without motor and balance deficits. AB - Running hamster OYC (RHO) is a mutant spontaneously arisen in a closed colony of Armenian Hamsters and has a characteristic of speedy, rotative running behavior. There are several reports which describe such mutants as abnormal walking behavior in rodents due to ataxia or balance deficits. In this report, we describe results of several motor and balance tests in order to clarify whether or not RHO mutants have such disorders. RHO mutants showed no sign of either motor or auditory deficits, and scarcely had balance deficits tested. The animals only had slight head tilting at the starting of running. These results suggest that RHO is a really new type of mutant which has not previously been reported. PMID- 11109561 TI - Exposure to airborne fungi, MVOC and mycotoxins in biowaste-handling facilities. AB - Health impacts due to fungi in indoor air can only be estimated reliably, if both fungal propagules and fungal secondary metabolites are qualified and quantified. In the present study, the fungal species composition in a compost facility is compared to the spectrum of microbial metabolites in the air with regard to the physiological properties of different fungal species. A number of relevant fungi was tested for the production of both volatile and non-volatile metabolites on different substrata. The profiles of mycotoxins and microbial volatile organic compounds (MVOC) turned out to be specific for certain species in pure culture. Consequently, the fungi may have different toxicological health impacts, though information on the relevance of microbial volatiles is still limited. PMID- 11109562 TI - Species-specific profiles of mycotoxins produced in cultures and associated with conidia of airborne fungi derived from biowaste. AB - The potential to produce mycotoxins and non-volatile secondary metabolites was investigated for approximately 250 freshly isolated fungal strains. Among the eleven most relevant species, viz. Aspergillus flavus, A. fumigatus, A. niger, A. parasiticus, A. versicolor, Emericella nidulans, Paecilomyces variotii, Penicillium brevicompactum, P. clavigerum, P. crustosum, and P. polonicum, a wide range of metabolites partly of toxicological relevance was identified. Several unknown metabolites were found for the less frequent species, which were primarily investigated for chemotaxonomic delimitation from closely related species. The spectra of metabolites in conidial extracts and culture extracts (containing also mycelium and medium) were compared for a limited number of relevant fungi. Some mycotoxins, such as sterigmatocystin in Emericella nidulans, were not present in the conidial extracts, though produced by most strains. Fumigaclavine C, tryptoquivaline, and trypacidin, characteristic for A. fumigatus, were found in conidial extracts, but highly toxic compounds such as gliotoxin and fumitremorgens were not present. Finally, compounds such as cyclopenol, cyclopenin, and penitrem A being characteristic for certain penicillia, were found in conidial extracts and are therefore assumed to occur in native bioaerosols. PMID- 11109564 TI - Scanning electron microscopical investigations of broncho-alveolar casts after intratracheal asbestos fibre instillation. AB - The evaluation of the toxicity of mineral fibres has been tried to achieve in experimental animal models. However, the appearance of fibres in the pleural space could not be explained satisfactorily. Histomorphological examinations showed that intratracheal instillation of asbestos fibres leads to parabronchial and intraalveolar granulomatous tissue reactions and bronchial epithelial regenerations. For further elucidation of the pathogenesis of lung cancer and of mesothelioma the localisation and transport of inhaled fibres is of high interest. Thus, a three dimensional visualization of the structure of rat lungs before and after intratracheal instillation of UICC crocidolite fibres was performed by plastic casts to follow the way of asbestos fibres in the lung tissues and the pleura. The casts allowed to demonstrate airway structures with imprints of epithelial cells and blood vessels of normal and treated animals by scanning electron microscopy. Instilled asbestos fibres transformed bronchial structures and resulted in cystic deformations of the pleural surface. The penetration of single fibres through bronchial trunks and the visceral pleura could be shown for the first time in a three-dimensional topography of the affected tissue. Now, there is support for similar results of histomorphological examinations indicating the possibility that asbestos fibres could penetrate the pleura and migrate into the pleural space. The question if the migration of fibres is a mechanical movement or an active transport is still under discussion. PMID- 11109563 TI - GIS-supported investigation of a nosocomial Salmonella outbreak. AB - Within an outbreak at a university hospital 102 persons (44 patients, 26 nursery school children and one relative as well as 31 employees) have been diagnosed to be infected by Salmonella enteritidis. Ninety-nine persons complied with the "primary case"-definition. The source of infection could not be detected in retrospect by hygienic-microbiological methods due to missing food samples. But GIS (Geographical Information System)-supported epidemiological investigation and analysis of food production showed that most likely vanilla pudding had been the vehicle of infection. Contamination of the pudding could be put down to the fact that its production took place in direct spatial and temporal association with the preparation of turkey. Probably further infections caused by this primary source were avoided by immediate measures. The making out of an HACCP-concept as well as structural and technical short-term redevelopment measures proved to be decisive factors to decrease the risk of further infections. From these experiences, some recommendations could be derived for the investigation of food borne outbreaks in hospitals. PMID- 11109565 TI - PCP in the blood plasma: current exposure of the population in Germany, based on data obtained in 1998. AB - The fungicidal substance pentachlorophenol (PCP) had been used commonly for wood protection and leather impregnation in Germany until 1989, when this substance was prohibited by law. Hence, the body burden in the general population in Germany has been steadily declining. The reference values (95th percentiles) in blood plasma decreased from 20 micrograms PCP/l in 1991 to 12 micrograms/l in 1996. In 1998 the current exposure in a large residential population was investigated. 623 persons with an average age of 34.6 years (0-62 years) were investigated. For all of them there was neither evidence of occupational contact with PCP nor of the presence of PCP in the residential indoor environment. The mean PCP concentration in the plasma samples was 2.4 +/- 3.9 micrograms/l, the median 1.7 micrograms/l, and the 95th percentile 6.1 micrograms/l, the maximum value was 59.3 micrograms/l. In children and adolescents higher median and 95th percentiles were obtained than in adults (median 2.5 vs. 1.5 micrograms/l and 95th percentile 7.7 vs. 5.9 micrograms/l). All values above 20 micrograms/l were checked again individually: relationships with the level of PCP in household dust could not be detected. In one family leather clothing containing PCP, however, was found to be the cause of unusually high PCP values in the blood. Our investigations confirm a trend observed in recent years: exposure to PCP in the population in Germany decreases steadily and leads to an actualized reference value of 6.1 micrograms PCP/l plasma. In individual cases, however, greatly increased PCP levels in blood can still occur today, for example due to leather clothing treated with PCP. PMID- 11109566 TI - A cytogenetic study on the teaching staff of a polluted school with a questionable increased incidence of malignancies. AB - Exposure to pollutants, in particular polychlorinated biphenyls (PCB), was established at a school built in 1966. Because of a statistically conspicuous increased frequency of breast cancer observed in the teachers of the school this study was performed to ascertain whether the teachers in the polluted school have an increased level of micronucleated cells (MN) or sister chromatid exchanges (SCE) as an expression of a raised cytogenetic risk. Teachers in a directly adjacent school served as one control group and those from a school about 30 km away as a second one. Each teacher had to answer a questionnaire and after venous blood samples had been taken, the number of MN and SCE in peripheral lymphocytes were determined. For the teachers in the polluted school, in addition, the length of stay in the building during the last month and year was recorded. Thereby no correlation with the number of MN and SCE was proven. In comparison with the two control groups, neither the number of MN nor SCE was increased in the teachers of the polluted school. Even if their predictive value for cancer risk assessment is disputed, MN and SCE have a high rating as standard procedures in the proof of an exposure to genotoxic agents. This study thus does not provide any evidence that, for the teachers in the polluted school, a relevant exposure to genotoxic agents exists. PMID- 11109567 TI - Nosocomial outbreak of vancomycin-resistant Enterococcus faecium at a German university pediatric hospital. AB - Nosocomial Infections caused by vancomycin-resistant enterococci (VRE) are an emerging threat to critically ill patients. At the University Hospital Eppendorf, VRE were isolated from 38 patients between August 1993 and April 1997, of whom 32 were hospitalized at the Department of Pediatrics. Pulsed-field gel electrophoresis revealed that 26 Enterococcus faecium isolates from patients of the Department of Pediatrics were identical or closely related, and that isolates from three additional patients of the same department were possibly related. All of these isolates were of vanA genotype. They were resistant to glycopeptides, ampicillin, ciprofloxacin, clindamycin, and erythromycin. Most isolates displayed high-level resistance to gentamicin, but all remained susceptible to quinupristin/dalfopristin. Implementation of stringent hand disinfection and environmental disinfection policies, as well as measures for patient isolation contained this first outbreak of VRE at a German Children's hospital, which emphasizes the importance of hygienic measures for the control of nosocomial spread of these organisms. PMID- 11109568 TI - Epidemiologic methods for the prevention of nosocomial infections. AB - There is an ongoing controversy in Europe about the benefits and limitations of epidemiologic methods for the prevention and control of hospital-acquired infections. Hospital epidemiology, aimed at measuring the necessity, or effect, of preventive strategies for nosocomial infection control, is still an unknown field in many European institutions. The conceptual framework presented here is not intended as a complete review of modern hospital epidemiology, but should be considered rather a viewpoint which tries to bridge the gap between microbiology based hospital hygiene and hospital epidemiology in Europe. The explanatory power and limitations of descriptive, analytical and interventional epidemiology are described. Based on the assumption that nosocomial infections have causal and preventive factors that can be identified through systematic investigation, epidemiologic methods add important knowledge to reduce hospital-acquired infections. PMID- 11109569 TI - A multiplex-PCR method to detect enterohemorrhagic (EHEC) and enteropathogenic (EPEC) Escherichia coli in artificially contaminated foods. AB - In view of the considerable public health risk of the enteropathogenic (EPEC) and enterohemorrhagic (EHEC) Escherichia coli, a multiplex-PCR based method was used for the amplification of the slt genes and of the eaeA gene. Three pairs of primers, different from the oligonucleotides previously used by other authors, were exploited for the amplification. Different E. coli serotypes were tested with the optimized protocol. Fifty three artificially contaminated samples and sixty naturally contaminated samples were processed with the multiplex-PCR. All the artificially contaminated samples gave positive results independently of the number of cells inoculated. On the contrary, the naturally contaminated samples were all negative. The results obtained from this experiment demonstrated that this protocol could be used for monitoring the spread of these organisms in food. PMID- 11109570 TI - Influence of sequential cultivation on virulence of Legionella pneumophila and Staphylococcus aureus. AB - Virulence of Legionella pneumophila strain Monza 3 and Staphylococcus aureus strain Smith diffuse was investigated after sequential cultures on nutrient media. L. pneumophila lost its ability to multiply within Acanthamoeba polyphaga after 50 passages on Legionella selective agar, while S. aureus maintained its pathogenicity in the mouse peritonitis model after 100 sequential cultures on tryptic soy agar. These results demonstrate high preservation of virulence in staphylococci in contrast to legionellae. Differentiation of virulent and avirulent phenotypes of legionellae by the protozoal model may be helpful for detecting sources of infection in water hygiene. PMID- 11109571 TI - Isolation of Vibrio alginolyticus from seawater aquaria. AB - The seawater bacterium Vibrio alginolyticus was detected in 5 of 20 water samples from seawater aquaria (from 3 of 5 units) and also from the surface of diseased stony corals. A total of 45 isolates were differentiated biochemically, of which 13 isolates (29%) proved to be V. alginolyticus. All those strains produced the virulence factors caseinase and lipase, 11 strains amylase and gelatinase. 7 strains showed lecithinase activity and 2 strains produced hemolysins. All examined strains showed a marked toxicity to vero cells proven by the MTT bioassay, but no toxicity to plant cells with the saline alga Asteromonas gracilis as model. The isolates were mostly resistant to beta-lactam antibiotics, macrolides and lincomycin. However, they proved to be susceptible to aminoglycoside- and polypeptide-antibiotics as well as to tetracyclines, chloramphenicol, florfenicol, enrofloxacin and sulfamethoxazol-trimethoprim. The possible participation of this bacterium in the bleaching and dying of stony corals is mentioned as well as its role as human pathogen. PMID- 11109572 TI - Screening of the municipal water system of La Plata, Argentina, for human intestinal parasites. AB - The La Plata River, though severely contaminated by intestinal parasites through the discharge of tons of crude fecal material from a main sewage channel, nevertheless provides drinking water to two-thirds of La Plata, Argentina, after conventional purification at a processing plant. With intestinal parasitosis being endemic here, we investigated the importance of this water in transmitting such pathogens to the city's populace by means of standard methodology for sample acquisition and processing involving filter-concentration of waterborne particulates. Of 14 tap-water samples collected from the distribution network, 12 pertained to four zones (A-D) within the city center; while the remaining 2 were obtained near the processing plant, 15 kilometers outside the city. Although parasites were found within the samples derived from the four urban zones, none were detected in the specimens obtained near the plant. The four downtown areas differed from each other as to the quantity and nature of the parasites present in their water: whereas zones A and B registered similar lower levels of contaminants, C and D exhibited higher values significantly different from the former two and from each other. Given an average parasite count/l citywide of 0.38 and a probability of encountering a parasite within 11 of water of 0.32, the municipal network is seen to contribute to the transmission of intestinal parasites. A routine system of water-quality control is therefore needed throughout the city along with the establishment of infrastructures for locating and eliminating peripheral sources of contamination. PMID- 11109573 TI - Playing with reality: III. The persistence of dual psychic reality in borderline patients. AB - This paper is a contribution towards understanding the difficulties of severely borderline patients as they are uncovered within the psychoanalytic process. The authors aim to extend a model from previous papers, bringing a developmental perspective to bear on self-representation and self-organisation. This model rests on an understanding of the child's experience of psychic reality in both normal and neurotic people. The authors explore the relevance of these developmental ideas in thinking about severe borderline and related disturbances seen in adult patients, from the point of view of both understanding the pathology and considering technique. Illustrations are given from a case that helped to prompt the formulation of these states in terms of persistent, pervasive distortions of the experience of psychic reality. There is then an attempt to elucidate the difficulties of borderline patients in tolerating both separation and intimacy, consideration of the way in which bodily experiences may be used as an equivalent of mental experience or contact, and a discussion of the impact of disturbances in the experience of psychic reality on the analytic process. PMID- 11109574 TI - Becoming a subject: some theoretical and clinical issues. AB - The author presents a cluster of theoretical and clinical thoughts and some clinical material concerned with the issue of subjectivity in psychoanalysis. For convenience, he has organised these thoughts around the notion of 'becoming a subject'. This entails a process of recovery, or discovery, of unconscious subjective elements. There is a discussion of what we mean by the term 'subject', with reference to various strands of philosophical and psychoanalytical thinking. It is suggested that becoming a subject involves a shift towards a 'subjective position'. This refers to how being a subject involves some capacity to take up different positions without becoming fixed in any frozen state of being. In order to be fully in touch with another person, in a truly subjective position, one begins to grasp the other's point of view; the other is seen as other, as a person or a subject, in a context, orientated to others and being affected by others. A subjective position involves allowing experiences of the other to interpenetrate oneself, so that they make an impact. In order to clarify these ideas, clinical material from the analysis of a bulimic patient is presented to illustrate some of the difficulties for this patient in becoming a subject and allowing the analyst to make an impact on her. PMID- 11109575 TI - Freud comes to Palestine. A study of psychoanalysis in a cultural context. AB - The authors describe the founding of Hashomer Hatzair as a radical Zionist scouting movement in Eastern Europe between 1913 and 1919, a little known episode in the rich history of Freud's impact upon this century. As refugees in Vienna, the young adherents of the movement experienced enormous personal and collective turmoil. Desperate to construct new, viable identities, these intellectually vibrant young men and women were drawn to Freud as part of their project of self creation. Beginning in the 1920s, as members of Hashomer Hatzair settled in agriculturally based collectives known as kibbutzim, the educational leadership of the movement argued that psychoanalytically informed education was the key to raising children free of bourgeois neuroses. They established strong ties with European analysts, translated and published psychoanalytic texts, insisted that educators be analysed or, at least, psychoanalytically informed, and built a complex educational system founded on their particular understanding of Freudian insights. For them, psychoanalysis was also seen as a general prophylactic guaranteeing the mental hygiene of the community as a whole. The authors examine the complex relationship between Hashomer Hatzair and psychoanalysis. In particular, they ask why these young adults were so drawn to Freud and what their particular reading of the psychoanalytic texts was, and demonstrate how these young pioneers created a 'usable Freud' as part of their project of designing and building a utopian society. PMID- 11109576 TI - Autistic devices in small children in mourning. AB - The authors set out to show what happens to two small girl-patients who, starting from similar events (mother's pregnancy and birth of a sibling), developed traumatic mourning that could not be managed in the way that is usual for normal or neurotic children. Autistic devices were generated in these patients in order to protect their mental economy. They consider that this way of reacting is not justified either by the quality or by the intensity of the external stimuli and that these patients may have had a predisposition. To operate with this mechanism, the patients had to distort their mental apparatus and their own self in such a way that expressions such as language were affected in both of them: verbal language disappeared in one of them and in the other fulfilled functions other than communication. Atrophies and breaks in development to impede perception and meaning of the facts from becoming loose were also generated, strengthening the denial. The analyst's protective inclusion and his sustained effort to understand what was happening, to maintain communication and a receptive and understanding attitude, plus interpretation, made it possible for both patients to rely on a new device, constituted by the analyst, who helped to lessen the hard protective layer and make sense organs and self more permeable, processing the conflict so that more normal development could resume. PMID- 11109577 TI - Varieties of long-term outcome among patients in psychoanalysis and long-term psychotherapy. A review of findings in the Stockholm Outcome of Psychoanalysis and Psychotherapy Project (STOPP). AB - This paper reports the main findings of a large-scale study of subsidized psychoanalysis and long-term psychotherapy. More than 400 people in various phases, before, during and after subsidized psychoanalysis or long-term psychodynamic psychotherapy, were followed up for a period of three years with personal interviews, questionnaires and official statistics. Our analyses revealed progressive improvement the longer patients were in treatment- impressively strong among patients in psychoanalysis--on self-rating measures of symptom distress and morale. Improvement, however, was equally weak in both groups on a self-rating measure of social relations. Dosage factors (treatment duration and session frequency in combination) partly accounted for the outcome differences between those referred to psychoanalysis and those referred to long term psychotherapy. Attitudes and ideals among therapists and analysts concerning the goals and means of psychotherapy were also associated with patient outcome, although in rather complex ways. A significant part of the outcome differences between patients in psychoanalysis and in psychotherapy could be explained by the adoption, in a large group of therapists, of orthodox psychoanalytic attitudes that seemed to be counterproductive in the practice of psychotherapy but not in psychoanalysis. It is suggested that this effect may be a negative transfer of the psychoanalytic stance into psychotherapeutic practice and that this may be especially pronounced when the attitudes are not backed up by psychoanalytic training. PMID- 11109578 TI - Who was Rumpelstiltskin? AB - The author describes the way in which Rumpelstiltskin has perplexed and enthralled readers since the brothers Grimm recovered the tale from the realm of folklore. In standard translation, the story now lives in a fixed literary form and, consequently, in the imagination of every child who has ever heard or read the story--virtually every person in the English-speaking world. Therefore, deeply rooted in childhood experiences, Rumpelstiltskin can be expected to appear in analysis, and he does. The compelling central character is the title figure, Rumpelstiltskin, whose name and actions tell us who he is and what he was intended to 'mean'--especially to his contemplated audience. The original narrators of and listeners to this tale were female visitors to the evening spinning chamber (Spinnstube), where women gathered and told tales to amuse themselves to ward off sleep while they spun. The butt of this story is male impotence and bluster, and the key to the story's meaning arises from matching the etymological roots of the central character's name with his actions as they appear philologically and psychoanalytically. PMID- 11109579 TI - A father's abdication: Lear's retreat from 'aesthetic conflict'. AB - The author explores the potential contribution of Shakespeare's 'King Lear' to psychoanalytic thinking, linking a reading of the play focused on the emotional tensions inherent in the parental function of endowing ('heriting') the next generation with the developmental struggle characterised by Donald Meltzer as the 'aesthetic conflict'. Following Meltzer's definition of passion as the 'consortium' of Bion's emotional links, love, hate and the urge to know (L, H and K), the author develops an understanding of 'aesthetic conflict' linked with the tension inherent in that constellation. It is suggested that L and H split off from each other and from K become attempts to possess and control, while K split off from L and H becomes an attack on dreaded emotional links, oscillating between attempting to ignore them and attempting to overcome them. The author suggests an affinity between Bion's K link and what in 'King Lear' is pictured as a capacity to 'see feelingly' in the context of the struggle to give the object its freedom. This way of characterising 'aesthetic conflict' is linked with a fresh look at weaning as a lifelong developmental process, which in turn leads to a reconsideration of the psychoanalytic models of the dynamics of mourning. PMID- 11109580 TI - Bion and binocular vision. AB - The author describes his contact with Bion over a twenty-year period, from Bion's supervision of his control case in London in 1960 to the period from 1968 to 1978 when they were both working in Los Angeles. He outlines Bion's views on the use of 'instinct' and intuition in patient observation, the depressive position in patient and analyst, and memory and desire as impediments to knowledge of 'ultimate reality'. Some case material is presented, illustrating how Bion's ideas, particularly concerning attacks on linking, informed the course of the treatment. The author then discusses Freud's, Klein's and Bion's approaches to the problem of resistance, Bion's expansion of some of Klein's ideas, his definitions of psychosis and his formulation concerning thoughts that develop before thinking. The author then argues how essential it is for the analyst to differentiate between primitive projections from the patient that are pre-verbal attempts to communicate a state of mind and those that are an expression of hostility or control. He then discusses the importance of understanding idealizing projections and differentiating these from a healthy positive transference. He concludes by characterising Bion's way of working in terms of his humility, his courage and, fundamentally, his use of his intuitive binocular mind. PMID- 11109581 TI - On Slochower's 'Erotic complications'. PMID- 11109582 TI - On Feldman's 'Some views on the manifestation of the death instinct in clinical work'. PMID- 11109583 TI - On Feldman's 'Some views on the manifestation of the death instinct in clinical work'. PMID- 11109584 TI - Other views: a discussion on a memoir of early childhood loss. PMID- 11109585 TI - Charles Rycroft (1914-1998). PMID- 11109586 TI - 'The central phobic position: a new formulation of the free association method' by Andre Green. PMID- 11109587 TI - [Vaccination. Meeting "Vaccinations 2000" Compiegne, June 22 2000]. PMID- 11109588 TI - [Atherosclerosis. XIIth International Symposium on Atherosclerosis Stockholm, 25 29 June 2000]. PMID- 11109589 TI - [The return of vitamin deficiencies]. PMID- 11109590 TI - [Aspirin and venous thromboembolism: some indications, but no certitude]. PMID- 11109591 TI - [Anemia caused by vitamin B 12 deficiency in subjects aged over 75 years: new hypotheses. A study of 20 cases]. AB - PURPOSE: New hypotheses have recently been developed on vitamin B12 deficiency and the frequently observed occurrence in the elderly subject of food cobalamin malabsorption, i.e., the non-dissociation of B12 and its carrier protein (ND B12), and the possibility of rectifying this imbalance by oral crystalline B12 supplementation. The aim of this study was therefore to confirm these hypotheses in a series of patients aged over 75 years with anemia due to B12 deficiency. METHODS: A retrospective study was carried out over a 5-year period on patients aged over 75 years presenting with megaloblastic anemia (hemoglobin [Hb] < 12 g/dL) and vitamin B12/cobalamin deficiency (B12 < 160 pg/mL). RESULTS: Twenty cases were analyzed. The average age of the patient population was 82.5 +/- 6 years, and the F/M sex ratio was 1:2. Mean Hb levels were 7.9 +/- 2.4 g/dL, mean serum B12 levels were 83 +/- 24 pg/mL, and mean homocysteinemic levels were 35 +/ 27 mumol/L. The diagnosis was as follows: food cobalamin malabsorption/ND B12 (n = 10), Biermer's disease/pernicious anemia (n = 5), malabsorption due to pancreatic insufficiency (n = 1), and low dietary B12 levels (n = 1). Disorders associated with ND B12 were: atrophic gastritis and Helicobacter pylori infection (n = 6), antacid or biguanide intake (n = 3), alcohol abuse (n = 2), or idiopathic syndrome (n = 2). In the patients who were followed up (n = 10), i.m. (n = 5) or oral (n = 5) administration of crystalline B12 resulted in the correction of hematological abnormalities. CONCLUSION: In the elderly subject, food cobalamin/ND B12 malabsorption appears to be the main cause of B12 deficiency, and is frequently associated with atrophic gastritis. In these cases, administration of oral crystalline B12 may be an efficient means of treating this disorder. PMID- 11109592 TI - [Cryptococcus neoformans infection in hematologic malignancies]. AB - PURPOSE: Cryptococcus is an opportunistic infection that affects immunodepressed patients and is a classical complication of AIDS-stage HIV infection. The aim of this study was to investigate Cryptococcus neoformans infections in patients with hematological malignancies. METHODS: Six cases have been described of cryptococcosis detected in Nantes, France over the past 10 years in patients with hematological malignancies. RESULTS: This infection has been found particularly in the context of lymphoproliferative disorders (chronic lymphocytic leukemia, Waldenstrom macroglobulinemia, Hodgkin's disease, and non-Hodgkin's lymphoma), and also following cytotoxic therapy. In four cases, the patients were treated with fludarabine, which rapidly caused long-duration marked lymphocytopenia, notably in CD4 cells. Cell-mediated immunity plays a major role in systemic defense against C. neoformans. It therefore seems that fludarabine favors the spread of cryptococcal infections. CONCLUSION: In the context of lymphoproliferative syndromes treated with cytotoxic drugs, in particular fludarabine, it appears important to take into account the possible presence of cryptococcal infection in the presence of respiratory, neurological or cutaneous disorders, so that a correct diagnosis can be made and the appropriate treatment administered. PMID- 11109593 TI - [Hereditary hemochromatosis]. AB - INTRODUCTION: Hereditary hemochromatosis is a fairly common disease in the Caucasian population, with a prevalence estimated at between 1.5 to 3/1,000 inhabitants. Over the past few years, its symptomatology has altered; at present, its clinical aspect with diabetes mellitus, cirrhosis, and darker skin pigmentation only constitutes 10% of new cases of this disease. CURRENT KNOWLEDGE AND KEY POINTS: In 1996, the discovery of the C282Y mutation in the HFE gene radically altered the diagnostic approach to hereditary hemochromatosis. At present, any patient admitted with an isolated case of asthenia, or with arthralgia or hypertransaminasemia should be examined via transferrin-saturation testing: if the transferrin saturation coefficient is > 45%, then the presence of the C282Y mutation should be investigated to confirm the diagnosis of hemochromatosis. A liver biopsy is no longer necessary to establish the diagnosis, but this is still useful in cases of possible cirrhosis, which is the main risk factor for hepatocellular carcinoma. Phlebotomy remains the sole recommended treatment, and should be undertaken in a case-specific manner. Family screening should be carried out for all first-degree relatives for every new case that is diagnosed. FUTURE PROSPECTS AND PROJECTS: The discovery of the HFE gene has permitted hereditary hemochromatosis to be easily differentiated from other forms of hepatic iron overload including a new syndrome, dysmotabolic hepatosiderosis. Casos of homozygotic C282Y without hepatic iron overload have been described, but the clinical outcome of some of these cases requires further study, and adds to the controversy on whether systematic population screening should be made available. PMID- 11109594 TI - [Olfactory disorders due to medications: analysis and review of the literature]. AB - INTRODUCTION: Olfactory disorders caused by medicinal drug intake are an uncommon occurrence. However, such an etiology should be systematically taken into account and investigated, as olfactory loss may be reversible once the particular treatment has been stopped. CURRENT KNOWLEDGE AND KEY POINTS: An analysis of the literature shows that of the large number of drugs that are apparently responsible for olfactory disorders, this adverse side effect has in fact only been observed in animal study populations, and no clinical case report has been made on the subject. The real toxicity to man is therefore only hypothetical. Of the 150,000 cases recorded in the pharmacovigilance centers in France, only 68 have reported olfactory complications (0.05% of cases), and only 22% of the medical files on this disorder reach a satisfactory level of plausibility. Cardiovascular drugs are mainly implicated in the development of olfactory disorders. Of these, certain drugs in particular should be mentioned: conversion enzyme (ACE) inhibitors which are responsible for taste disorders, some betablockers, and a calcium antagonist (a dihydropyridine derivative). Olfactory disorders have also been reported following administration of drugs used in anesthesia, in cancerology, endocrinology (carbimazole), in immunology (interferon), in the treatment of infectious diseases (ciprofloxacine, dioxycycline, terbinafine), and in rheumatology (D-penicillamine). FUTURE PROSPECTS AND PROJECTS: It is frequently difficult to establish a direct relationship between drug exposure and an olfactory disorder, and it is often not easy to determine with any certainty the causative role of the drug in the development of this disorder. PMID- 11109595 TI - [Non-steroidal anti-inflammatory agents with selective inhibitory activity on cyclooxygenase-2. Interest and future prospects]. AB - INTRODUCTION: Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit the production of primary prostanoids by blocking the access of arachidonic acid to the active site of the cyclooxygenases (COXs). Because the prostanoids produced by COX-1 appear to play a physiological role (protection of the gastric mucosa, platelet aggregation, vascular homeostasis, maintenance of renal sodium-water balance) while those produced by COX-2 seem mainly to intervene in the inflammatory response and in certain processes associated with cell proliferation, the hypothesis has been put forward that the NSAIDs that are selective COX-2 inhibitors should theoretically be capable of maintaining NSAID therapeutic properties but also have fewer adverse side effects due to the maintenance of prostaglandin production at normal physiological levels. CURRENT KNOWLEDGE AND KEY POINTS: The hypothesis of COX isoenzyme selectivity has led to a proposed classification for COX inhibitors: 1) COX-1 selective inhibitors (low dosage aspirin); 2) COX non-selective inhibitors (the majority of classified NSAIDs, which when administered over the long term, e.g., in cases of rheumatoid arthritis, cause duodenal ulcers in 20% of cases and gastric hemorrhage in 1-4% of cases/year); 3) COX-2 preferential inhibitors (meloxicam and nimesulide, which have fewer gastric side effects than standard NSAIDs, but which are not risk-free at high doses); 4) COX-2 selective inhibitors (celecoxib and rofecoxib). Preliminary clinical studies have shown that COX-2 selective inhibitors are as efficient as standard NSAIDs and have fewer adverse digestive side effects, thereby confirming the interest of this proposed classification. In the UK, the aforementioned studies have led to the commercialization of rofecoxib for the treatment of pain and osteoarthritis, while celecoxib has been introduced in medical practice in the USA and other countries for the treatment of rheumatoid arthritis and osteoarthritis. FUTURE PROSPECTS AND PROJECTS: Various epidemiological and laboratory studies have indicated that NSAIDs may be able to reduce the risk of cancer (colorectal cancer in particular) and Alzheimer's disease due to their inhibitory activity on COXs, especially COX-2. The therapeutic contribution of COX-2 specific inhibitors has to be more fully evaluated, particularly as these agents could delay the healing of duodenal ulcers and interfere with several COX-2-induced physiological functions. It is therefore suggested that until further information becomes available, this new class of NSAIDs should be used with caution in certain patient populations. PMID- 11109596 TI - [Acute neuromyocarditis secondary to diet-induced beriberi. Case report]. AB - INTRODUCTION: Thiamine deficiency can be determined by various clinical signs; some of these symptoms may be acute, and require an urgent diagnosis. In countries such as ours with a high standard of living, this disorder is more commonly observed in cases of severe alcoholism, and cases of diet-associated thiamine deficiency are rare, and therefore not easily recognized. The metabolic disorders resulting from vitamin B1 deficiency are responsible for the well-known central or peripheral neurological symptoms, and also for the less common and often more acute cardiovascular reactions. Immediate thiamine/vitamin B1 supplementation is of major importance. The rapid reversal of symptoms following this treatment is often considered as a diagnostic index. EXEGESIS: In this study, an original case of diet-associated thiamine deficiency has been reported, with clinical symptoms including myocarditis and subacute peripheral nerve involvement in a young adult. The disorder was the result of a thiamine deficiency in the diet, which was exclusively based on milled rice. CONCLUSION: The present report is interesting both as regards its clinical aspects and its etiology, and it emphasizes the importance in the differential diagnosis of a given case of taking the possibility of diet-related thiamine deficiency into account, although this is an uncommon etiology in developed countries. PMID- 11109597 TI - [Pericardial extravasation as an indicator of congestive heart failure due to thiamine deficiency in a young adult]. AB - INTRODUCTION: Thiamine (B1) deficiency is one of the classical causes of congestive heart failure. Although in the western world and in other developed regions this disorder is generally associated with chronic alcoholism, it may also only occur as a result of a deficient diet. EXEGESIS: A young patient was admitted for electrocardiographic examination, and pericardial extravasation was recorded. The etiological assessment showed a case of congestive heart failure due to thiamine (B1) deficiency. A hemodynamic examination and investigation of thiamine blood levels confirmed this diagnosis. The patient's health status improved following thiamine administration, with complete and rapid regression of symptoms of congestive heart failure. CONCLUSION: Although cardiomyopathic beriberi is infrequent, it should systematically be taken into account in the etiology of congestive heart failure. The present study also notes that a dietary thiamine deficiency is uncommon, but should nevertheless be considered when other symptoms of denutrition are present. PMID- 11109598 TI - [Association of Kaposi sarcoma--multiple myeloma. A new case]. AB - INTRODUCTION: Kaposi's disease is an angiogenic multifocal cancer process that has several forms, namely Mediterranean, African, HIV-associated, and secondary to a preexisting immunodepressive state (hematological disorder, corticosteroid therapy, immunodepressive treatment). Whatever its form, Kaposi's sarcoma is probably associated with a chronic viral human herpes type 8 infection (HHV8). EXEGESIS: This virus has been implicated in the pathogenesis of multiple myeloma (17 cases recorded to date). In the present study, a further case of Kaposi's sarcoma associated with multiple myeloma has been reported. However, Epstein-Barr virus, cytomegalovirus, hepatitis B and C, HIV and HHV8 serologies were negative. Radiotherapy on the lower limbs was initiated. CONCLUSION: It is concluded that HHV8 does not appear to play a pathogenic role in cases of multiple myeloma, given the rarity of the association between Kaposi's sarcoma/multiple myeloma/HHV8. PMID- 11109599 TI - [A case of cervical calcinosis]. PMID- 11109600 TI - [Scurvy, a still current diagnosis]. PMID- 11109601 TI - [Analgesic effect of a diphosphonate perfusion during the acute phase of vertebral compression in the elderly subject]. PMID- 11109602 TI - [Giant cell hepatitis in the adult associated with HIV and hepatitis C virus co infection]. PMID- 11109603 TI - [Diagnosis of cerebrospinal fluid leakage at the base of the skull]. AB - There are still several problems surrounding the diagnosis of cerebrospinal fluid leak. Currently the method of choice for cerebrospinal fluid detection is qualitative determination of beta-2-transferrin. Faster and more efficient methods (beta-trace) are under clinical investigation. The major problem is localisation of the site of leakage. Combination of several radiological methods increases the rate of correct diagnosis. In surgery the use of intrathecal sodium fluorescein improves visualisation of the site of leakage and thus increases the chances of secure and stable closure of the cerebrospinal fluid fistula. PMID- 11109604 TI - [Cause of death and autopsy findings in patients of the Swiss HIV Cohort Study (SHCS)]. AB - The Swiss HIV Cohort Study (SHCS) is a prospective cohort study of HIV-infected adolescents and adults seen at 7 outpatient clinics (Swiss University Hospitals in Basle, Berne, Geneva, Lausanne, Zurich, the St. Gall Cantonal Hospital and the Civico Hospital in Lugano). The SHCS serves as an infrastructure for different research projects and includes about 70% of all patients with advanced disease in Switzerland. From April 1984 to November 1995 3120 HIV-infected patients of the SHCS died. Autopsies were performed in 314 of these patients. The aim of our study is to analyse autopsy findings as well as causes of death in those 314 HIV infected patients. An HIV-related cause of death was found in 271 (86%) of the patients, 12 patients (4%) died of a drug overdose, and 3 (1%) of the patients committed suicide. 28 (9%) died either from an HIV unrelated or unidentified cause. The five most frequent causes of death were: bacterial pneumonia (52 patients, 17%), Pneumocystis carinii pneumonia (40 patients, 13%), lymphoma (34 patients, 11%), cytomegalovirus infection (33 patients, 11%), and toxoplasmosis (30 patients, 10%). During our study marked progress occurred in treating HIV infected patients and preventing opportunistic infections. These improvements have further changed the natural course of acquired immunodeficiency syndrome. They are reflected in the falling rate of Pneumocystis carinii pneumonia and toxoplasmosis, as well as an increase in lymphoma as a cause of death over the period of our study. PMID- 11109605 TI - [Neonatal alloimmune thrombocytopenia due to a rare anti-HPA-1b antibody]. AB - Foetal and neonatal alloimmune thrombocytopenia is caused by transplacental transfer of antibodies directed against platelet antigens and affects approximately 1 in 1000-2500 neonates. Clinically relevant complications are the intracranial haemorrhages that occur in 10-20% of cases. 20 platelet antigen systems are currently known. However, immunisation is most frequently seen against two of these (HPA-1a and HPA-5b). Treatment options include transfusion of compatible or, if these are not available while urgently needed, random donor platelets, intravenous immunoglobulin, and steroids. We report on a case of neonatal alloimmune thrombocytopenia due to an anti-HPA-1b antibody in the third pregnancy of a 31-year-old Caucasoid woman. The infant was treated with repeated maternal and random donor platelet transfusions and with a single dose of intravenous immunoglobulin. PMID- 11109606 TI - [CO2-laser therapy of stigmatizing cutaneous lesions in tuberous sclerosis (Bourneville-Pringle) and in neurofibromatosis 1 (von Recklinghausen)]. AB - Tuberous sclerosis (Bourneville-Pringle) and type I neurofibromatosis (von Recklinghausen) are familial multiple tumour syndromes. Both entities have in common that the cutaneous manifestations can stigmatize the carriers and considerably reduce the quality of life. CO2-laser surgery (vaporisation) consistently yields good to excellent aesthetic results in the treatment of adenoma sebaceum in tuberous sclerosis. Patients with type I neurofibromatosis who carry several hundreds neurofibromas usually benefit from a numerical reduction in their skin lesions, despite the less predictable aesthetic result, ranging from excellent to fair. The present study reports on the treatment outcome of CO2-laser surgery in 8 patients with adenoma sebaceum and 8 with neurofibromatosis. PMID- 11109607 TI - [ACE inhibitor-induced intestinal angioedema]. PMID- 11109608 TI - [Concerning: Stettler C, Mueller B, Diem P. What you always wanted to know about HbA1c. Schweiz Med Wochenschr 2000;130:993-1005]. PMID- 11109609 TI - [Medico-social problems of chronic articular and spinal diseases]. PMID- 11109610 TI - [In vivo and vitro effects of interferon-alpha 2b on functional activity of T lymphocytes from patients with rheumatoid arthritis]. AB - AIM: To investigate the effect of recombinant alpha 2b-interferon (r alpha 2b IFN) on functional capacity of peripheral blood (PB) T cells in rheumatoid arthritis (RA) patients and the relationship between functional characteristics of T lymphocytes and the disease activity. MATERIALS AND METHODS: PB mononuclear cells (PBMC) were separated by Ficoll-Verografine++ gradient centrifugation from 24 healthy donors (HD) and 75 RA patients 19 of which were treated with r alpha 2b-IFN (realdiron, Biofa, Lithuania) in the dosage 1 million IU i.m. each other day for 20 days, 10 injections a course. Cell surface markers (CD3, CD4, CD8) and adhesion molecules (CD18, CD54, CD2) were analyzed using specific monoclonal antibodies (MoAbs) and flow cytometry on the PBMC, freshly isolated and treated for 72 hours with medium alone, PHA, r alpha 2b-IFN and their combination. The proliferative response of PBMC to MoAbs for CD3, PHA and r alpha 2b-IFN were assessed by 3H-thymidine incorporation. The percentage of spontaneous and inducing apoptosis was quantified by flow cytometry using propidium iodide staining. RESULTS: The expression of CD18 was lower on RA PB lymphocytes compared to HD PB lymphocytes (p < 0.05). After stimulation of PBMC in both RA patients and HD with PHA, percentages of CD2+, CD3+, CD4+, CD18+ cells significantly diminished (p < 0.05), whereas the percentages of CD54+ and CD18+ (p < 0.05) cells increased. We have found three types of RA PB lymphocytes response to complex factors in vitro: 1) the presence of the proliferative response to T mitogens but not to r alpha 2b-IFN (56% of the patients); 2) the presence of the increased proliferative response to T-mitogens and r alpha 2b-IFN (17% of the patients); 3) the absence of the proliferative response to T-mitogens and r alpha 2b-IFN (27% of the patients). PBMC of HD demonstrate only the first type of the response. R2 alpha b-IFN demonstrated own mitogenic effect and increased mitogen induced proliferation in PBMC cultures with a high proliferative response to T mitogens. The levels of spontaneous and inducing apoptosis were increased in RA PB lymphocytes compared to HD. After stimulation with PHA, RA PB lymphocytes preferentially underwent apoptosis whereas cells of HD proliferated. High disease activity correlated positively with an increase of a proliferative response to mitogens and apoptosis and a decrease in the percentage of lymphocytes, expressed adhesion molecules. The treatment with r alpha 2b-IFN induces changes in T-cell response to mitogens similarly to those after incubation with r alpha 2b-IFN in vitro before treatment. CONCLUSION: Functional capacity of RA PB lymphocytes relates to the disease activity. Inhibitory or stimulatory effects of r alpha 2b IFN depend on functional activity of RA lymphocytes. Using the test with alpha 2b IFN incubation, we may predict changes of apoptosis and proliferation levels caused by different agents in RA lymphocytes after treatment with r alpha 2b-IFN. PMID- 11109611 TI - [Alpha-2 macroglobulin in serum and synovial fluid of patients with rheumatoid arthritis and osteoarthrosis]. AB - AIM: To study concentrations of alpha-2-macroglobulin (A-2-MG) in blood serum and synovial fluid (SF) in patients with rheumatoid arthritis and formulation of basic clinical, laboratory and x-ray criteria of RA activity. MATERIALS AND METHODS: A-2-MG levels were measured in the serum and SF from 151 RA patients and in the serum of 20 patients with osteoarthrosis (OA) and 62 healthy donors. The serum concentration for RA patients was 166 +/- 65.3 mg%, for OA patients--175.26 +/- 36.99 mg% and for healthy donors--177.772 +/- 50 mg%. Mean concentration of SF A-2-MG in RA patients was 98.77 +/- 82.43 mg%. CONCLUSION: Changes in the concentration of A-2-MG are unrelated to inflammation activity in the joints of RA and OA patients. Serum and synovial concentration of this protein corresponds to changes in the concentration of IgM and rheumatoid factors in RA patients. PMID- 11109612 TI - [Immunological aspects of early stage rheumatoid arthritis diagnosis]. AB - AIM: To study immune status of patients with rheumatoid arthritis (RA) to improve immunodiagnosis at early stage of the disease. MATERIALS AND METHODS: Immunological examination covered 28 patients with rheumatoid arthritis (RA) aged 16 to 72 years. The duration of RA varied from 1.5 months to 1.5 years. Lymphocyte population and T-lymphocyte subpopulation were measured using monoclonal antibodies. Serum Ig were measured in Reafarm plates. RESULTS: Patients with stage II articular function insufficiency (AFI) demonstrated a significant lowering of the absolute number of lymphocytes, natural killers, elevated concentration of IgA compared to patients with less severe AFI. Patients with systemic symptoms had significantly decreased percentage of T-lymphocytes vs patients with isolated articular syndrome. Natural killers' levels were elevated in all the patients in early RA. A significant rise in the percentage of B lymphocytes and serum IgG concentrations were also seen. In T-lymphopenia, relative amount of T-helpers and T-suppressors was significantly elevated while the ratio T-helpers/T-suppressors was reduced. CONCLUSION: Changes found in the immune status allows diagnosis of early RA, characterize immune disorders, help to select adequate immunomodulating therapy supporting function of the suppressor cells. PMID- 11109613 TI - [Long-term drug therapy of rheumatoid arthritis and remission]. AB - AIM: To evaluate the duration of rheumatoid arthritis (RA) remission with respect to different drug treatments. MATERIALS AND METHODS: Remission duration observed at 20-year follow-up of 442 RA patients living in the south of Estonia has been reviewed. Also, the data are provided on the disease onset, articular status, systemic lesions, RA activity and progression, the latest exacerbation and previous remission, standard laboratory indices, humoral immunity, examination of the biopsy of the articular tissues and subcutaneous fat for amyloid. RESULTS: According to the retrospective analysis, slow-progressive RA course occurs primarily in remissions longer than 5 years and less frequently in remissions lasting from 1 to 5 years. No matter what the drug was used, 14% of the patients have failed the treatment. 3% of the patients were in remission longer than 5 years. Short-term remissions (1-3 months) were induced in 13%, stable ones (3 months-1 year) in 39% of the cases. These remissions were observed in early RA, more frequently in patients with initial arthritis of the small joints. Remissions from 1 to 5 years were registered in 31% of the patients. CONCLUSION: RA remissions up to 1 year represent temporary clinical improvement and do not inhibit progression of the rheumatoid process. Consideration of association between RA clinical course and remission duration helps to recognize groups of RA risk and to timely change treatment policy. PMID- 11109614 TI - [Approaches to prediction of gastropathy development risk due to non-steroidal antiinflammatory drugs]. AB - AIM: Examination of gastric secretion in rheumatoid arthritis (RA) patients and its response to sodium diclophenak and indomethacine. MATERIALS AND METHODS: 46 RA patients entered the study. All of them have been long on nonsteroid antiinflammatory drugs (NAID). Two groups of patients were analysed: group 1- free of NAID gastropathy, group 2--with NAID gastropathy. pH was followed by Gastroscan system. RESULTS: The analysis of 24-h pH-grams in both groups when NAID were not taken showed that in NAID gastropathy acid-forming gastric function is not affected while the alkalizing function markedly declines. Indomethacine is more aggressive to upper gastrointestinal tract mucosa than sodium diclophenak. CONCLUSION: Dynamic intragastric pH-metry can prognosticate the risk of NAID gastropathy. PMID- 11109615 TI - [Treatment with megadoses of dexaven (dexamethasone) versus methypred (6 methylprednisolone) of patients with rheumatoid arthritis]. AB - AIM: To compare clinical effectiveness and tolerance of methylprednisolone (methypred) and dexamethasone (dexaven) in patients with rheumatoid arthritis (RA), to estimate side effects and complications rate. MATERIALS AND METHODS: The trial included 31 patients with seropositive RA (27 females, 4 males) stage II and III. Dexaven pulse-therapy was given to 16 patients in a dose 2 mg/kg for 3 days, 15 patients received methypred in a classic dose 1000 mg for 3 days. Clinical response was assessed on day 1, 7 and 30 after the treatment. RESULTS: Both drugs significantly reduced severity of arthralgia, morning joint stiffness, number of inflamed joints, the disease activity diminished 2-3-fold. Side effects were minimal. CONCLUSION: Dexaven (dexamethasone) is a drug of choice in pulse therapy of RA. It is not inferior to routine treatment with methylprednisolone (methypred). PMID- 11109616 TI - [Effects of low-intensity infrared impulse laser therapy on inflammation activity markers in patients with rheumatoid arthritis]. AB - AIM: To evaluate effects of low-intensity infrared impulse laser therapy (IRILT) on concentration of immunity activation [not readable: see text] (soluble receptors of TNF-alpha and neopterin) and indicator of the inflammation activity (concentration of C-reactive protein) in patients with rheumatoid arthritis (RA). MATERIALS AND METHODS: Enzyme immunoassay, radioimmunoassay, enzyme immunoassay and radial immunodiffusion were used to measure soluble receptors of TNF-alpha, neopterin and C-reactive protein in 38 females with verified RA receiving IRILT or sham procedures. RESULTS: IRILT induced lowering of neopterin, TNF-alpha soluble receptors (p < 0.01) and C-reactive protein (p < 0.01). CONCLUSION: The findings give pathogenetical grounds for IRILT use in RA as this treatment suppresses functional activity of macrophages which serve the main source of neopterin and the receptors synthesis. PMID- 11109617 TI - [Leiden, G20210A mutations in prothrombin gene and antiphospholipid antibodies in systemic lupus erythematosus and antiphospholipid syndrome]. AB - AIM: To estimate incidence rate of Leiden mutation in factor V gene, prothrombin gene mutation responsible for replacement of G for A in position 20210 (G20210A) of its 3'-end noncoding part in patients with systemic lupus erytheamtosus (SLE) and antiphospholipid syndrome (APS) and their relationships with antiphospholipid antibodies (aPL): lupus anticoagulant (LA) and anticardiolipin antibodies (aCL). MATERIALS AND METHODS: The trial included 33 patients (2 males and 31 females) aged 20-62 years (mean age 32.7 +/- 9.8 years). 29 patients suffered from SLE, 17 of them had also APS (criteria by G. R. V Hudhes). 3 patients had primary APS, 1 female had hemorrhagic vasculitis. IgG and IgM-aCL were detected by enzyme immunoassay. In revealing mutation, DNA was used isolated from peripheral blood by standard methods based on polymerase chain reaction (PCR). Primary screening of Leiden mutation was made according to Bertina et al. with Mnl I restrictase. The mutation was confirmed by an original technique with allele-specific primers. G20210A mutation in the prothrombin gene was determined with restrictase Taq I after introduction of artificial restriction site in PCR product. The patients were divided into 2 groups. Group 1 incorporated 12 patients with SLE without APS. Group 2--21 patients with APS (17 with SLE + APS, 3 with primary APS and 1 with hemorrhagic vasculitis). RESULTS: Patients of group 1 had neither thrombotic complications nor Leiden mutation. Two patients of group 2 had heterozygous Leiden mutation. Both females were aPL-positive and had previously recurrent thrombophlebitis. One of them had also recurrent disorders of cerebral circulation. None of the examinees had any G20210A mutations in the prothrombin gene. CONCLUSION: Detection of genetic defects in APS provide arguments in the dispute about the necessity, duration and choice of anticoagulant therapy. If patients with APL syndrome appeared to have besides aPL also genetic defects in coagulation, this will enable identification of patients at high risk of thrombosis which need permanent administration of anticoagulants among which only low-molecular ones or glycosaminoglycanes are indicated. Mutation in gene of factor V is absolute contraindication to phenilin and quamarines. PMID- 11109618 TI - [Intrarenal hemodynamics in patients with systemic rheumatic diseases]. AB - AIM: To study intrarenal hemodynamics (IRHD) in patients with systemic lupus erythematosus (SLE) and sclerodermia systematical (SS). MATERIALS AND METHODS: IRHD was studied in 30(28 females and 2 males) and 25(22 females and 3 males) patients with SLE and SS, respectively, and mean duration of the disease 5.6 +/- 0.5 and 7.5 +/- 1.2 years, respectively. Lupus nephritis was diagnosed in 21(70%), sclerodermic nephropathy in 10(40%) patients. Patients with chronic renal failure were not included. IRHD was investigated using protein loading which provided the index of functional renal reserve (FRR). RESULTS: Impaired IRHD was detected in 21(70%) patients with SLE. 19 patients had lupus nephritis. FRR depletion was found also in 2 patients free of lupus nephritis symptoms. 10 of them (8 with active nephritis and 2 with intact kidneys) had arterial hypertension. FRR depletion was found also in 12(48%) of SS patients. 7 of them had sclerodermic renal lesions. CONCLUSION: Impaired IRHD was encountered frequently both in SLE and SS patients and can develop in the absence of renal affection symptoms. PMID- 11109619 TI - [Dynamic interphasic tensimetry of blood and urine in systemic lupus erythemotosus]. AB - AIM: To study dynamic surface tension (DST) of blood and urine in SLE patients. MATERIALS AND METHODS: Clinical, immunological examinations, DST of blood and urine using computer tensiometers were made in 67 SLE patients aged 15 to 62 years. RESULTS: Changes in interphasic tensiograms in males and females were characterized regarding the course, activity of the pathological process and the presence of certain clinical symptoms. Effective therapy return to normal DST of the biological fluids. Correlations between interphasic tensiograms of blood, urine and levels of proteins, lipids, non-protein and inorganic surfactants and surface-inactive substances were established as well as relations of DST with biochemical composition of the biological fluids. CONCLUSION: The above diagnostic method may be useful for differential diagnosis, assessment of the clinical course, activity of the disease, control over effectiveness of the treatment. PMID- 11109620 TI - [Interferon therapy effects on activation of T-lymphocytes in patients with systemic lupus erythematosus]. AB - AIM: To elucidate the effects of combined therapy with glucocorticosteroids (GCS) and recombinant human interferon alpha or gamma (IFN) on proliferative responses of T-lymphocytes activated by various surface molecules (CD3 and CD2) in patients with SLE. MATERIALS AND METHODS: A 3-month trial entered 3 groups 15 patients each with verified SLE by APA criteria (1982). Patients of group 1, 2 and 3 received IFN-alpha (realdiron, Biofa, Lithuania) in a single dose 3 million IU i.m., IFN-gamma (inflagen, Biofa, Lithuania) in a single dose 3 million IU i.m. and cyclophosphamide in a dose 200 mg i.m. once a week, respectively. T lymphocyte proliferative response was assessed to stimuli of two types: CD3 dependent (classic activation) and CD2-dependent (alternative activation). The analysis was made by inclusion of 3H-thymidine after 72-hour incubation of peripheral blood mononuclear cells with various stimuli. The response was assessed before the treatment, on the treatment day 20 and after the treatment. The blood from 27 donors was also examined. Flow cytometry estimated the percentage of the cells expressing molecules CD3, CD4, CD8. RESULTS: The effect is found of alpha and gamma INF on functional capacity of T-lymphocytes and on the number of cells expressing surface molecules CD3 and CD4. Realdiron produced two-phase reaction to a proliferative response to mitogenic stimuli by CD3 dependent activation pathway: the initial rise then lowering. CD2-dependent way of T-cell activation is associated with weakening of responses to all combinations of stimuli with participation of autologous red cells. This group of patients to the end of the therapy exhibited a significant decrease in the number of cells expressing CD3 and CD4 (p < 0.05 and p < 0.001, respectively). Inflagen enhanced CD3-dependent activation of T cells and normalized the response to all types of the alternative stimuli. This group demonstrated an increase in the number of cells expressing CD3 and CD4 (p < 0.01 and p < 0.05, respectively). The changes in the number of CD8+ cells in both the groups were statistically insignificant. The controls had T-cell responses reduced by both activation pathways. CONCLUSION: Preparations of both alpha and gamma interferon have a multidirectional influence on functional potential and phenotype of T-lymphocytes of SLE patients. PMID- 11109621 TI - [Spondylarthropathies and rheumatoid arthritis in some Finno-Ugrian populations in Russia]. AB - AIM: Determination of the prevalence of spondylarthropathies (SAP) and rheumatoid arthritis (RA) among Finno-Ugrian population of Russia. MATERIALS AND METHODS: A one stage expedition trial was made of representative samples of Mordovian and Mari populations including 1312 citizens aged over 14 years. RESULTS: Incidence rate of SAP among Mordovian and Mari examinees was 0.7 and 0.2%, respectively, ankylosing spondylitis being most frequent finding. RA occurred in 2.5 and 0.6%, respectively. All the RA cases were females without family history of this disease. CONCLUSION: SAP and RA prevalence among Mordovian and Mari populations was high but their course is more favorable than among other population of Russia. PMID- 11109622 TI - [Differential diagnosis of chronic urogenic arthritis]. AB - AIM: To investigate features of chronic urogenic arthritis (CUA) and its differences with psoriatic arthritis (PA) and ankylosing spondylarthritis (AS) with joint lesion. MATERIALS AND METHODS: CUA, AS, PA were diagnosed according to S. M. Sidelnikova et al., S. van der Linden and Agababova, respectively, in 94 patients. The disease ran for more than 3 years. Articular syndrome was examined in CUA, AS and PA. RESULTS: Articular syndrome in CUA remains for the most part monoarticular with affection of the joints of the low extremities. AS has clinical and x-ray signs of sacroileitis, affection of the spinal column and hip joints. PA runs with multiple arthritic lesions of the hand and foot joints. CONCLUSIONS: Chronic infection persists in all the CUA and PA patients. Its exacerbation coincides with arthritis aggravations. Only in CUA there is a chronological connection between acute or aggravated chronic urogenital infection and initial symptoms of joint disease. PMID- 11109623 TI - [Hyperaggregation syndrome in patients with osteolytic psoriatic arthritis]. AB - AIM: To study blood rheology in patients with psoriatic arthritis (PA) having local or advanced osteolysis. MATERIALS AND METHODS: The trial included 16 patients with significant PA and clinical and x-ray symptoms of joint surface osteolysis. Kinetics of red cell aggregation and disaggregation was studied in specially designed erythroagregometer. RESULTS: The assessment of red cell aggregation indicated the presence of hyperaggregation syndrome in the majority of PA patients with associated osteolysis. CONCLUSION: Hyperaggregation may contribute to bone destruction. Its onset can be reviewed as a risk factor for serious destructive changes in the joints. PMID- 11109624 TI - [Mechanisms of steroid osteoporosis development in patients with hormone dependent bronchial asthma]. AB - AIM: To examine mechanisms of steroid osteoporosis development in patients with hormone-dependent bronchial asthma (BA). MATERIALS AND METHODS: The trial included 73 patients with hormone-dependent BA, 19 BA patients who have never taken oral steroid hormones and 15 healthy subjects. RESULTS: By computed tomography of the lumbar spine, bone tissue density in steroid-dependent BA was significantly decreased vs BA patients who were not treated with steroids. Osteopenia was discovered in 61.9% of the examinees who have received oral glucocorticoids. Development of osteopenia in this case was not dependent of the steroids dose and duration of the treatment. Patients with hormone-dependent BA vs healthy subjects had higher 24-h urine calcium excretion, normal levels of parathyroid hormones, calcitonin, low concentrations of 25-oxyvitamin D3, osteocalcin in the serum. Three subgroups of patients with osteopenia were identified according to the statistics: with prevalent resorption of bone tissues (24.1%), with low concentrations of vitamin D3 and osteocalcin (52.6%), with highly active resorption in combination with low vitamin D3 and weak function of osteoblasts (23.3%). CONCLUSION: Mechanisms of steroid osteoporosis development are heterogeneous. This should be considered in its treatment. PMID- 11109625 TI - [Gastrointestinal bleeding induced by nonsteroidal antiinflammatory drugs]. PMID- 11109627 TI - [In Process Citation] PMID- 11109626 TI - [Osteoarthrosis]. PMID- 11109628 TI - [New fields of cyclosporin A (sandimmun) application in rheumatology: treatment of systemic lupus erythematosus]. PMID- 11109629 TI - [Tick-borne borreliosis (Lyme disease). Ecology, clinical picture and etiology]. PMID- 11109630 TI - [Rheumatoid arthritis in the light of psychological problems]. PMID- 11109631 TI - [Perspectives of diuretics as first-line therapy in management of arterial hypertension]. PMID- 11109632 TI - [Genetic diversity of viruses. Consequences for screening and prevention]. AB - The evolution of viruses contributes to their diversification, whether it be a result of their own replication, or host-pressure dependent. Certain viral types, groups or subtypes are therefore found in certain regions of the world or in certain populations. The development of blood screening reagents is nearly always based on viral antigens or viral sequences derived from 'prototype' strains or antibodies raised against these prototype strains. Therefore in situations where an individual is infected by a viral strain that is genetically and antigenically distantly related to the prototype strain used in the development of the test, screening failure may occur. In the present article, this has been illustrated via 3 models, the human immunodeficiency virus (HIV), the hepatitis B virus (HBV), and the B19 parvovirus. Viral diversity also has a negative effect on the prevention of blood-transmitted viral infections. The example provided concerns vaccination failure and/or seroprophylaxis against hepatitis B. PMID- 11109633 TI - [Evaluation of spontaneous peri-transfusional screening for HVC and HIV at the Rouen University Hospital Center]. AB - INTRODUCTION: Since October 1996, French hospitals have been instructed to introduce screening for hepatitis C virus (HCV) and human immunodeficiency virus (HIV) in all patients before and 3 months after each blood transfusion. The aim of this study was to assess the degree to which this recommendation had been taken into account in a university hospital via a pre- and post transfusion screening comparison. PATIENTS AND METHODS: A retrospective study on the use or non-use of screening tests for HCV and HIV was carried out in 2 groups of 150 randomly selected patients who had received blood transfusions in 1996 and in 1998. RESULTS: The coverage by pre-transfusion screening tests for HCV and HIV varied from 23% in 1996 to 20% in 1998 (not significant). The post-transfusion screening tests were performed by the hospital in 6% of the cases in 1996 and in 3% of the cases in 1998 involving blood transfusion. CONCLUSION: This study suggests that in the majority of patients, screening (particularly post transfusion screening) for HCV and HIV was not carried out, and that over the 2 year period considered no noticeable improvement was observed. However, these results only concerned one hospital in which no specific screening program had been introduced. It is therefore possible that these findings are not representative of the situation in other hospitals; further studies would be useful in this regard. PMID- 11109634 TI - [Peripheral stem cell collection, search for predictive factors: a multicenter study. Working Group on Transfusion and Therapeutic Techniques of the French Blood Transfusion Society ]. AB - AIMS: A multicentric study involving 12 centers was made to investigate the results of peripheral stem cell collection carried out between 1996 and 1997 from 655 patients with hemopathic syndromes or malignant tumors, The aim of this investigation was to determine the predictive factors for transplant quality, and to thereby optimize collection procedures. PATIENTS AND METHODS: Information sheets were completed for 1,346 cytapheretic sessions, i.e., 655 grafts. The samples were taken after induction chemotherapy and exposure to hematopoeitic colony-stimulating growth factors (except the LMCs). Each graft was defined as being of good or bad quality depending on the number of CD34+ cells that it contained. Based on the data available in the literature, a workgroup consensus was reached that a level of CD34+ cells +/- 2.10(6)/kg recipient body weight constituted a good transplant criterion. The 2 subgroups (good graft versus lower quality graft) were compared by univariate analysis followed by discriminant multivariate analysis. RESULTS: It was established that a number of parameters were significantly linked to the criterion of collection quality; however, 3 predictive factors emerged from the multivariate analysis--the level of circulating CD34+ cells; the number of cytaphereses; the number of blood volumes treated. CONCLUSION: It was concluded that the level of circulating CD34+ cells seems to be an essential aspect in predicting the quality of the transplant and the number of cytaphereses required to obtain a sufficiently rich collection. Moreover, it also appears that at least 2 blood volumes should be treated to optimize the results. PMID- 11109636 TI - Decision aides in immunohematology: detection of anti-erythrocyte antibodies. AB - Detection and identification of irregular red-cell antibody in the serum or plasma of a patient is of prime importance for the prevention of hemolytic transfusion reactions and the biological supervision of the hemolytic disease of the foetus or the newborn. Practice in these tests is replete with complex biological problems. Using problem solving strategies, we discuss the recognition and resolution of the most frequent difficulties encountered in red cell antibody identification. PMID- 11109635 TI - [Molecular basis and structure-activity relationships of the Duffy blood group antigens: chemokine and Plasmodium vivax receptors]. AB - Duffy blood group antigens are of major interest in clinical medicine as they are not only involved in blood transfusion risks and occasionally in neonatal hemolytic disease, but also in the invasion of red blood cells by the hemoparasitic Plasmodium vivax. The FY locus maps to chromosome 1q22-q23, and is composed of 4 alleles: FY*A and FY*B (coding for the Fya and Fyb antigens, respectively), FY*X and FY*Fy. The Duffy antigens are carried by a 336 amino-acid glycoprotein named the Duffy Antigen/Receptor for Chemokines (DARC) that can bind with high affinity selected members of the CXC and CC classes of chemokines. Today, the genetic bases of the Duffy system have been characterized. The identification of the polymorphisms associated with the 4 alleles FY*A, FY*B, FY*Fy and FY*X has led to the development of a complete genotyping of the Duffy system by PCR, which increases the safety and lessens the risk of blood transfusion, and is useful in determining feto-maternal incompatibilities and in genetic filiation analyses. DARC is not solely expressed in erythroid cells: the same polypeptide isoform is found on the surface of endothelial cells of post capillary venules throughout the body and also on the surface of Purkinje cells in the cerebellum, although it is encoded by different RNA messengers in each case, i.e., 1.35 and 7.5 kb, respectively. The preliminary analyses of receptor ligand interaction have shown the existence of a chemokine-binding pocket defined by the close proximity of the first and fourth transmembrane domains of the DARC protein, and also by the importance of the N-terminal extracellular region for the binding of Plasmodium vivax merozoites. PMID- 11109637 TI - Dialysis techniques in the critically-ill patient. PMID- 11109638 TI - Presymptomatic testing for genetic diseases: new frontiers, new dilemmas. PMID- 11109639 TI - Prevalence and risk factors for colonisation with gram-negative bacteria in an intensive care unit. AB - OBJECTIVE: To investigate prevalence and determine risk factors for colonisation with Gram-negative bacteria in ICU patients. DESIGN: Prospective, surveillance study. SETTING: 26-bed surgical and paediatric ICU. PATIENTS: 159 patients- whereof 22 infants--admitted to the surgical/paediatric ICU over a two-month period. INTERVENTION: In all patients routine microbiological monitoring was performed by thrice weekly oral swabs, urine sampling and, additionally, tracheal aspirates in patients on mechanical ventilation (MV) and by anal swabs once weekly. RESULTS: Population characteristics: Mean age of the adult population was 51.1 +/- 17.6 year. Mean age of the paediatric population was 6.3 +/- 5.3 year. The mean APACHE II-score was 18 +/- 9.1. The mean PRISM-score was 9.7 +/- 5.4. The mean ICU stay was 7.5 +/- 11.4 days. 43.4 percent of patients received mechanical ventilation (MV). The mean number of mechanical ventilation days was 11.1 +/- 14.7 days. 32.1% of patients experienced colonisation with Gram-negative bacteria. Prevalence of colonisation increased with length of ICU stay. The probability of colonisation was 24% after an ICU stay of 3 days (= median ICU stay). Time to colonisation was not different between the controlled sites (p > 0.05). 47% of colonizations were due to multiresistant strains. Higher APACHE II scores and MV were associated with a higher prevalence of colonisation (p < 0.01). The ICU mortality was 8% among adult and 4% among paediatric patients. CONCLUSION: Patients with high APACHE II-scores, on mechanical ventilation and with an ICU stay of more than 3 days are most at risk for colonisation with Gram negative bacteria. These patients should be cared with the optimal precautions in the prevention of colonisation and infection. PMID- 11109640 TI - [Cost effectiveness of vaccination against pneumococcal bacteremia in the elderly: the results in Belgium]. AB - BACKGROUND: Several studies have shown that pneumococcal vaccination of older persons would be cost-effective in preventing pneumococcal pneumonia, but evidence of clinical protection for this condition is uncertain. Given much better evidence of vaccination effectiveness against invasive disease, studies showing that vaccination is cost-effective in preventing invasive disease alone could provide strong support for public policies to vaccinate older persons. METHODS: We examined the cost-effectiveness of preventing invasive pneumococcal infection by vaccination with the 23-valent pneumococcal polysaccharide vaccine of persons > or = 65 years in age in Belgium. The direct medical costs expressed per quality adjusted life year (QALYs) of a cohort of vaccinated persons was compared with the costs per QALY in a cohort of persons who are not vaccinated. RESULTS: Preventing invasive pneumococcal infections by vaccinating elderly persons clearly benefits people's health. By vaccinating 10,000 persons over 65 years of age, approximately eight QALYs can be gained compared with no vaccination. Achieving these health benefits however requires additional costs,: 30,000 ECU per QALY gained. The cost-effectiveness ratio is slightly better (i.e. 25,000 ECU per QALY) for the age group 65-75 years, and slightly worse (i.e. 35,000 ECU per QALY) for the age group 75-84 years. It increases sharply to 77,000 ECU per QALY for the persons over 85 years of age. An extensive one dimensional sensitivity analysis did not greatly affect these results. If vaccination is also clinically effective in preventing pneumococcal pneumonia, vaccinating all elderly persons is cost saving. CONCLUSION: Using empirical epidemiological data, pneumococcal vaccination to prevent invasive pneumococcal disease is acceptably to moderately cost-effective in Belgium. On the basis of our findings, we believe public health authorities should consider policies for encouraging pneumococcal vaccination for all persons > or = 65 years in age. PMID- 11109642 TI - Hodgkin's disease in a patient with Von Hippel-Lindau disease. A case report. AB - Von Hippel-Lindau (VHL) disease is an autosomal dominant inherited disorder with a predisposition to develop a wide variety of lesions: retinal, cerebellar, spinal and medullar hemangioblastomas, renal cell carcinomas, phaeochromocytomas, and renal, pancreatic and epididymal cysts are the most frequent manifestations of the disease. The prevalence of VHL disease has been estimated to be 1 per 36,000 persons. We report the case of a 68-year-old woman with Von Hipple-Lindau disease who developed high fever with pulmonary and hepatic lesions proven to be Hodgkin's disease on biopsy. To our knowledge, this is the first report of Hodgkin's disease in a patient with Von Hippel-Lindau. PMID- 11109641 TI - Physician's involvement in the smoking cessation process of their patients. Results of a 1998 survey among 4,643 Belgian physicians. AB - This report relates to the 1,667 responses to a selfadministered mail-back questionnaire sent by BELTA to a sample of 4,643 physicians (17.3% current smokers) who are in professional contact with patients (response rate: 35.9%). Links between active smoking and disease are considered as well-demonstrated by 98.8% physicians and for passive smoking by 85.3%, for foetal consequences of smoking during pregnancy by 96.4%. Nicotine dependence is admitted by 83.3%. Interaction of smoking with drug metabolism is insufficiently known. Modulation of the specific approach of smoking cessation, according to the various stages of the cessation cycle, to the level of nicotine dependence and to the psychological status of the smoker is not sufficiently perceived by the physicians. Patient's smoking status is systematically determined by less than half the physicians, of whom nearly 90% claim to inform their smoking patients on smoking-related risks, and 84.2% to tackle the problem of cessation. The intervention is mostly limited to a firm advice, completed by nicotine replacement for a maximum of 50% of smokers (especially gum and patch). Referral to specialized structures is unfrequent (between 10 and 20%). Follow up after cessation is clearly deficient. In this retrospective study of their activity patterns, physicians' reports may reflect their intentions rather than their actual practices. We conclude that smoking issues and cessation techniques should be more intensively taught both at graduate and postgraduate levels, in order to obtain a more active behaviour of health professionals against smoking. PMID- 11109643 TI - Penicillin or vancomycin plus ceftazidime in febrile neutropenic patients after stem cell transplantation. AB - The response of gram-positive cocci to third generation cephalosporin therapy in febrile neutropenic patients is not optimal. We evaluated the safety and efficacy of ceftazidime plus penicillin (C + P) and compared it to ceftazidime plus vancomycin (C + V) in febrile neutropenic patients. The study includes 64 patients admitted to the Department of Haematology. Thirty-six patients were treated with C + V and 28 patients with C + P. Control of infection was observed in 78% of the patients in the C + V group and in 57% of the patients in C + P group (p = 0.5). Infection was the cause of the death of 1 patient in each group. We conclude that the combination of C + P has the same activity as the combination of C + V in febrile neutropenic patients. The morbidity and mortality were identical in both groups but cost effectiveness was in favour of the C + P group. PMID- 11109644 TI - [Sarcoidosis]. PMID- 11109645 TI - [The treatment and prognosis of sarcoidosis. Apropos 53]. AB - OBJECTIVE: We retrospectively studied 53 cases of sarcoidosis which have been diagnosed at our service during de last 14 years. The criteria for starting therapy with corticosteroids and recurrences were analyzed. METHOD: The patients mean (SD) age was 42 (15) years old (range 16-76) and the majority were female (72%). 15 patients (28.3%) received corticosteroids. The only differences with respect to patients not treated were the presence of respiratory symptoms (47% vs 18%; p = 0.04) and the abnormality of spirometric parameters (DLCO/VA p = 0.01; CV p = 0.002). RESULTS: 17 (32%) patients recurred. 14 (82%) of them required corticosteroids. This percentage was significantly greater than that of patients treated at first episode (82% vs 28%, p = 0.0002). All patients improved with treatment. The only difference with respect to patients without recurrences were to be treated at the first episode (53% vs 17%, p = 0.007). PMID- 11109646 TI - [Why evidence-based medicine? 20 years of meta-analysis]. AB - BACKGROUND: Meta-analysis, described within evidence-based medicine, has become a frequent issue in recent medical literature. An exhaustive search of reported meta-analysis from any medical specialty is described. MATERIAL AND METHODS: Search of papers included in Medline or Embase between 1973-1998. A study of intra and inter-reviewers liability about selection and classification have been performed. A descriptive analysis of the reported papers (frequency tables and graphics) is described, including differences of mean of reported meta-analysis papers by medical specialty and year. RESULTS: 1,518 papers were selected and classified. Most frequently found (45.91%) were: methodology (15.7%), psychiatry (11.79%), cardiology (10.01%) and oncology (8.36%). Inter personal agreement was 0.93 in selecting papers and 0.72 in classifying them. Between 1977-1987 overall mean of reported studies of meta-analysis (1.67 + 4.10) was significatively inferior to the 1988-1998 (49.54 + 56.55) (p < 0.001). Global number of meta analysis was positively correlated (p < 0.05) with the number of studies about fundamentals and methodology during the study period. CONCLUSIONS: The method used to identify meta-analysis reports can be considered to be adequate; however, the agreement in classifying them in medical specialties was inferior. A progressive increase in the number of reported meta-analysis since 1977 can be demonstrated. Specialties with a greater number of meta-analysis published in the literature were: psychiatry, oncology and cardiology. Diffusion of knowledge about fundamentals and methodology of meta-analysis seems to have drawn and increase in performing and reporting this kind of analysis. PMID- 11109647 TI - [The effects of enalapril on exercise capacity and right ventricular function in patients with chronic cor pulmonale]. AB - BACKGROUND: Chronic cor pulmonale (CPC) in patients with chronic obstructive pulmonary disease (COPD) is a predictor of mortality. ACE inhibitors (ACEIs) improve the symptoms and reduce mortality of left ventricular or congestive failure, however their long term use in patients with CPC has not been tested. The aim of this study is to assess the effect on exercise tolerance and cardiorespiratory function of long term administration of enalapril in patients with CPC. METHODS: Placebo-controlled, double blind, randomized study. 28 patients (24 men and 4 women, mean age of 68.11 +/- 7.78, range 51-79) with CPC and without exacerbation of their respiratory symptoms at baseline were double blind randomised to receive enalapril or placebo for 6 months, added to their previous therapy. Respiratory function test, exercise tolerance ("six minutes walking test") and isotopic ventriculography were performed at baseline and at the end of the study. RESULTS: At baseline there were no differences in FEV1, FVC, FEV1/FVC and exercise tolerance. Both placebo and enalapril were well tolerated. At the end of the study, patients taking enalapril increased their exercise tolerance an 8.9% (19 m) vs 4.7% (33 m) in the placebo group (p 0.44; 95 percent confidence interval, -41.10 to 91.99). RVEF improved a 6.4% with enalapril but worsened a 7.09% in placebo group (p 0.15; 95 percent confidence interval, -12.87 to 2.10). CONCLUSIONS: Long term administration of enalapril do not improve neither exercise tolerance, nor right ventricular (RV) function, although given in increasingly doses is well tolerated and might prevent further worsening in RV systolic function. PMID- 11109648 TI - [The prognostic markers of survival and progression in HIV-1 infection. A study of CD4+ lymphocytes, antigen p24 and viral load during 3 years in a cohort of 251 patients]. AB - BACKGROUND: Prospective study of survival and AIDS or death progression in a cohort of 251 HIV infected patients whose seroconversion time is unknown, with 1 main objective: To analyse CD4+ lymphocytes count, p24 antigen plasmatic levels and viral load as surrogate markers. PATIENTS AND METHODS: 251 patients were included, most of them undergoing antiretroviral therapy, followed consecutively in the HIV/AIDS Unity of Internal Medicine Service of the Hospital Universitario Arnau de Vilanova in Lleida. We made clinical and analytical baseline studies and every 3 months thereafter. In relation to CD4+ lymphocytes, 3 groups were established: group I, 500 or more cells/mL; group II, 200-499 cells/mL and group III, < 200 cells/mL. In the same way, with p24 antigen we established 3 group: group I, < 20 pg/mL, group II, 20-39 pg/mL, group III 40 or more pg/mL. We studied survival in relation to baseline levels and stability, the latter being understood as persistent levels in the initial group, or better, over 3 year period. Survival analysis was made by Kaplan-Meier estimation. Relative risk was calculated by Cox's proportional hazards model. RESULTS: During the 36 months of follow-up 53 patients died. AIDS progression risk or death was 4.8 times higher for the p24 antigen > = 40 pg/mL group than for the p24 antigen < 20 pg/mL one; patients with p24 antigen between 20-39 pg/mL relative risk was 2.5 times higher than those included in p24 antigen < 20 pg/mL group. These results emphasize that if we take into account the p24 antigen stability during these 36 months. In relation to progression study, 36 patients progressed. AIDS progression risk or death for p24 antigen > = 40 pg/mL group was 7.69 times higher in relation to that with p24 antigen levels between 20-39 pg/mL. The bivariable study shows that CD4 lymphocytes counts and p24 antigen level have quite an independent value. The comparison with viral load by PCR determination makes manifest discrepancy, difficult to explain. CONCLUSIONS: p24 antigen plasma level is a good survival and AIDS progress or death surrogate markers in HIV infected patients, and it is useful for 3 years or more. An isolated value < 20 pg/mL and, furthermore, the stability in successive controls under this concentration is a sign of good prognosis. Its value is emphasized with CD4+ lymphocytes count. It seem necessary that more comparative studies with viral load are required. PMID- 11109649 TI - [A case of pulmonary sarcoidosis induced by interferon alfa treatment in a female patient with hepatitis C]. AB - In spite of the generalized use of alpha interferon to treat hepatitis C there are very few cases in the literature on pulmonary sarcoidosis due to this therapy. A new case with this diagnosis is reported and the possible pathogenic mechanisms driving alpha interferon to sarcoid histologic changes are discussed. PMID- 11109650 TI - [Mucinous adenocarcinoma of the appendix associated with ovarian tumors and pseudomyxoma peritonei. The difficulty in differential diagnosis]. AB - Mucinous adenocarcinoma of the appendix is rare. If there is a concomitant ovarian tumor to determine the primary might be difficult. Histological features are not always determinant, but there are some macroscopic findings that may suggest an origin in the appendix. We report a case of synchronous tumors in appendix and ovaries with pseudomyxoma peritonei. The patient presented with mass sensation in the right lower quadrant, asthenia, anorexia and weight loss. Abdominal ultrasound and CT scan showed a tumor involving cecum, appendix, terminal ileum and pelvis. Findings on colonoscopy and biopsies were inconclusive. At laparotomy, the tumor compressed appendix, cecum and ascendant colon, terminal ileum, ovaries and peritoneum. Histopathological analysis demonstrated a well-differentiated mucinous adenocarcinoma of appendiceal origin with metastasis in ovaries and peritoneum (pseudomyxoma peritonei). PMID- 11109651 TI - [Lobar emphysema due to bronchial atresia associated with a bronchogenic cyst]. AB - We report a case of bronchial atresia in the apical segment of right down lobule associated to subcarinal bronchogenic cyst in an eighteen years old patient who consults by long time bronchitis. The thorax X-ray shows an image suggested of mucoid impaction. The respiratory function tests are not essential for the diagnostic. The fiber bronchoscopy is normal and the diagnostic is established by mean image technics (thoracic HR-TC and MR). PMID- 11109652 TI - [Subacute thyroiditis associated with positive antibodies to the Epstein-Barr virus]. AB - Subacute thyroiditis is an inflammatory disorder of the thyroid caused probably by viruses. It is clinically characterized by the presence of anterior cervical pain and/or painful goiter, and rarely as fever of unknown origin or as prolonged fever syndrome. We report a case of a 49-year-old female admitted to the hospital because of fever during last month, leukocytosis and accelerated erythrocyte sedimentation rate. Following observation, slight tenderness over the thyroid gland and signs of hyperthyroidism occurred. After the laboratory studies, low thyroidal radioactive iodine uptake and fine-needle aspiration cytology (FNAC) of thyroid, she was diagnosed of subacute thyroiditis with hyperthyroidism. We believe that the etiologic agent was the Epstein-Barr virus because heterophile and Epstein-Barr virus-specific antibodies were positive. The patient was treated with acetaminophen (1.500 mg/day) with prompt and complete resolution of the clinical and laboratory abnormalities. There has been no recurrence of the disease during a 1 year follow-up. PMID- 11109653 TI - [Pulmonary fibrosis. The fibrogenic factors and therapeutic possibilities]. PMID- 11109654 TI - [Prognosis, evidence-based medicine and patient preferences]. PMID- 11109655 TI - [A "lupus-like" syndrome as the form of presentation of pulmonary adenocarcinoma]. PMID- 11109656 TI - [A proliferative syndrome and venous thrombosis]. PMID- 11109657 TI - [Reflex sympathetic dystrophy and primary biliary cirrhosis]. PMID- 11109658 TI - [Partial reflex sympathetic dystrophy in its radial form]. PMID- 11109659 TI - [Acute pancreatitis with a fatal evolution due to antimonials in patients with visceral leishmaniasis and HIV infection]. PMID- 11109660 TI - [Pericardial tamponade as the first manifestation of Hashimoto's disease]. PMID- 11109661 TI - [Renal infarct as the form of presentation of mitral stenosis]. PMID- 11109662 TI - [A decrease in height in women with non-insulin-dependent diabetes mellitus (NIDDM)]. PMID- 11109663 TI - [Truncus communis-type congenital cardiopathy: survival up to 53]. PMID- 11109664 TI - [The internal medicine service and the palliative care unit]. PMID- 11109665 TI - [Major surgery in thoracic injuries]. AB - Chest injuries have a high and steadily increasing incidence in western countries, but only some of the most common problems they create require an emergency thoracotomy or surgical video thoracoscopy. Flail chest, persistent pneumothorax, massive haemothorax, mediastinal emphysema, cardiac tamponade and intrathoracic foreign bodies can be identified as major surgical problems. Some of such patients (i.e. those with flail chest or foreign bodies) would be immediately candidates for major intervention. Other require fast but diagnostic procedures, because the choice of a therapy is dependent upon a precise identification of the damage. Injuries of trachea and primary bronchi, oesophagus, diaphragma, vena cava, great lung vessels, heart and aorta may represent important surgical emergencies; some leading rapidly to death. Fortunately, major surgical procedures are not really frequent in the management of thoracic traumas. Only 42 (3.5%) of nearly 2,000 patients with non-penetrating thoracic injuries had a thoracotomy or an surgical video thoracoscopy. The figure is far different for penetrating wounds; in fact 12 patients (41%) of 29 underwent mayor surgery. PMID- 11109666 TI - [The surgery of tumors and "limited situations"]. AB - The term "limit" applied to cancer surgery, denotes the ideological moment beyond which one cannot and should not propose any aggressive treatment. Such limits may concern the operability of a patient and may be represented by some general characteristics independent of the patient's current disease status (e.g. very old age, poor performance status, poor cardiac, respiratory, renal hepatic or mental conditions). They may concern the neoplastic involvement of the organ affected by the tumor: if undertaken, surgery should guarantee a reasonable duration of life, and a quality of life that makes it worth living. Other factors to be taken in consideration are the possibility the tumor spread to local or distant sites, as well as certain extreme conditions such as cancer, cachexia, liver/kidney failure, irreversible septic-toxic shock, ect. Moreover, there may be limits related to the structural conditions of the establishment where the operation is to be carried out (facilities, equipment, pharmacological supplies, medical and paramedical personnel) and to the social environment and the economic situation of the patient, in view of the assistance required following surgery. Lastly, a severe assessment of one's own fitness to perform any specific task should be part of the daily preparation of any surgeon. PMID- 11109667 TI - [The dehiscence of colorectal anastomoses: the risk factors]. AB - OBJECTIVE: To evaluate the results of emergency and elective colorectal resective surgery; to identify general and local factors that influence the anastomotic leak rate. MATERIAL AND METHOD: 200 selected consecutive patients (115 males and 85 females, medium age 50.6 years, range 16-87) underwent resective colorectal surgery between 1990 to 1997. 154 (77.0%) were operated in elective surgery and 46 (23.0%) in urgency, for carcinoma, diverticular disease, mesenteric infarction, chronic intestinal disease, dolicosigma, anastomotic leakage, familiar polyposis or lesions by firearm. The operations consisted in 58 right colectomy, 28 left colectomy, 6 resection of the transverse and 29 of the sigmoid colon, 40 anterior resection, 12 total colectomy, 19 closing of colostomy, 6 by passes. Anastomoses were performed in 88 cases by manual and in 110 by mechanical sutures. RESULTS: We observed 12 (6%) anastomotic leakages. Mortality rate was 1.0%. 13%.0 of these patients were underwent before to emergency and 3.9% to elective surgery; 5.7% by manual and 6.4% by mechanical suture. Diagnosis of leakage was made by clinical features, blood vessel examinations and abdominal TC scan. DISCUSSION: The risk factors of anastomotic leakage are general or local. Chronic obstructive pulmonary disease, perioperative transfusion, level of serum albumin, use of corticosteroid in the first group and sepsis, bowel obstruction, anastomotic level and tension and poor blood supply in the second, appear the most important causative factors in the development of anastomotic leaks. CONCLUSION: The incidence of dehiscence in colo-rectal surgery was seen significatively lower when anastomoses were performed in ideal circumstances than in the presence of one or more unfavorable factors. Healing remains a process depending more on the patient than on any aspect of the surgical technique. PMID- 11109668 TI - [Palliative treatments in obstructive jaundice due to periampullary neoplasms]. AB - OBJECTIVE: The evaluation of the palliative procedures, surgical and endoscopical or radiological, in the treatment of patients affected by obstructive jaundice resulting from periampullary tumors. SUBJECTS: Patients with jaundice by periampullary tumors undergoing to surgery or to endoscopical or radiological procedures from january 1987 to april 1998. RESULTS: Jaundice has come down in all patients after surgery. Mortality after surgery was 5.2% (5.9% in geriatric patients); morbidity 15.8% (17.6% in geriatric patients) survival 10.4 months after surgery (8.8 months in geriatric patients) versus 3.1 months after non surgical procedures. DISCUSSION: In all patients periampullary tumors are more frequent than hepatic hilum and common bile duct tumors. We have performed surgical and not surgical palliative procedures more frequently than curative resection (DCP), especially in geriatric patients (94.5% versus 79%). In geriatric patients we have chosen, between bile-digestive by-passes, the cholecysto-jejunal anastomosis because it is easier and faster to carry out than choledochojejunal anastomosis with the same results as well as from literature data. We have always performed a gastroenteric anastomosis in association with palliative surgical procedures to prevent or to solve a duodenal obstruction. This additional treatment didn't show an increasing of mortality and morbidity as well as from literature data. CONCLUSIONS: We have reserved the palliative non surgical procedures only to high surgical risk patients. In the other cases we have chosen palliative surgery for better long-term results and quality of life in the general series patients as well as in geriatric patients. PMID- 11109669 TI - [Hemorrhagic emergency due to esophagogastric varices in the portal hypertension patient]. AB - High gastrointestinal hemorrhage represents the more frequent (12-71.1%) and heavy complication of hepatic cirrhosis and correlates to portal hypertension; it is weighed by global mortality which sways from 30 to 50%. High gastrointestinal hemorrhage gives, therefore, a serious of diagnostic and therapeutic problems not easy to guide for at least 3 reasons: numerous causes of bleeding; hepatic failure; the marigold possible therapies. Aim of this work is to clarify some diagnostic and therapeutic features about high gastrointestinal hemorrhage in cirrhotic patient, because such eventuality often presents dramatic aspects, which endangers the patient's life. Our experience shows a casuistry referred to the period of time which goes from 1987 to 1998 and that comprehend 143 examined patients: 91 of them have been submitted to medical treatment (endoscopic sclerotherapy, glupressin e/o somatostatin); in 52 cases it has been possible to realize a surgical treatment, different from the elective therapy (33 pz) and emergency therapy. Immediate hemostatic effect obtained in both the conditions, has been satisfying with best results at a distance of three years and five years given by devascularization. As matter stands our preference of the devascularization surgical treatment, it seems appropriate to pay attention to the operation of mesocaval anastomosis which, either in our very brief experience (3 cases) or by international literature, seems to offer encouraging results. PMID- 11109670 TI - [Traumatic retroperitoneal lesions]. AB - The presence of lesions on the retroperitoneum generally worsens the prognosis in traumatic pathology; it implies more attention and skills from both the medical and surgical aspect. All type of trauma, blunt or open, may involve retroperitoneal structures and organs; specifically there may be lesions on the great vessels, pancreas, duodenum, oesophagus and genitourinary apparatus. Mortality is high, compared to abdominal traumatic lesions confined within the peritoneal sac. Treatment of single or associated lesions requires a multidisciplinary approach, as the surgical repair implies a specific knowledge and experience on different organs, whose habitual pathology lies on the hands of more surgical specialists. Lesions of great vessels are immediately life threatening; moreover the choice to "open" a patient for a retroperitoneal hematoma has to be taken upon a careful estimation. It could be better in more than a situation leave such hematoma in its place, specially in the iliac region, waiting for the spontaneous resolution of the hemorrhagic source and of the hematoma itself. The involvement of oesophagus, duodenum or pancreas determines instead a poorer prognosis at a distance. In conclusion retroperitoneal traumatic lesions are among the most challenging and serious emergencies, and necessitate a maximum of attention and expertise by the surgical team involved. Only with a careful judgement about the tactics and the procedures to carry on it is possible to obtain valid results, which often means to safe the patients life. PMID- 11109671 TI - [The emergency treatment of aneurysms of the supra-aortic trunks and of the internal carotid]. AB - In the last 10 years, four patients with aneurysms of the supraaortic trunks and the internal carotid artery have been operated in emergency at Department of Vascular Surgery of University of Bari. The first case was a mycotic aneurysm of the carotid bifurcation, the second an aneurysm of internal carotid artery interesting the retropharynx, the third a post-operatory pseudoaneurysm of common carotid in a patient affected by Takayasu's disease and the last an atherosclerotic aneurysm of the innominate artery. Clinical picture suggested an immediate surgical treatment in all patients. Different procedure were used depending on size and localization of each lesion. Results in all cases have been satisfactory. The aneurysms of supraaortic trunks and of internal carotid artery are very rare; therefore when clinical picture is dramatic or quickly worsening, emergency treatment is mandatory in order to reduce mortality risk and minimize complications. PMID- 11109672 TI - Ductal carcinoma in situ (DCIS) of the breast: thirty-two consecutive cases under 50 yrs detected by mammography: treatment and results. AB - Thirty-two consecutive cases of ductal carcinoma in situ of the breast in women under fifty are presented. Diagnostic procedure, pathological aspects, treatment and outcome are reported. All the patients had their cancer diagnosed by mammography. The tumor was marked by stereotactic or ultrasound guided localization. Lumpectomy without axillary node dissection was the surgical treatment of thirty lesions with good cosmetic results; radiation therapy was advised in all of these cases. In two cases mastectomy with immediate reconstruction was performed because of the multifocality of the cancer, none of the patients experienced local or distant recurrence. This experience emphasizes the importance of mammographic screening for women 40 years of age, in fact this approach allowed the diagnosis of a large number of DCIS. A correct definition of the problem and a multidisciplinary therapeutical approach is warranted to prevent the high local recurrence rate reported in the past. PMID- 11109673 TI - [Biliary ileus: a review of the literature and report of a clinical case treated by minilaparotomy]. AB - OBJECTIVE: To evaluate the diagnostic and therapeutic approach to gallstone ileus. MATERIAL AND METHODS: A gallstone ileus report, diagnosed by integrated radiological approach, treated by minilaparotomy plus enterolithotomy and followed up for two years. RESULTS: Traditional radiologic findings, ultrasonography and computed tomography showed both the stone in the bowel lumen and the cholecystoduodenal fistula with air in the biliary tract. The enterolithotomy alone worked out the obstruction and no complication was observed. DISCUSSION: Integrated radiologic examinations are indicated in defining nature, site and seriousness of gallstone ileus. In most cases surgical treatment is mandatory to work out both the bowel occlusion and bilioenteric illness even though it is often wiser to perform an operation of enterolithotomy alone owing to the general and local conditions of the patient. After this surgical approach, bilioenteric fistula disappears in most cases with no residual lithiasis; in the cases in which fistula and/or lithiasis persist, a decisive surgical operation can be performed in a second step. PMID- 11109674 TI - Curative resection for colorectal cancer in the elderly. Prognostic factors and five-year follow-up. AB - The purpose of this perspective study was to evaluate which prognostic factors predict long-term survival and disease-free survival (DFS) of elderly patients (> or = 65 years) who underwent surgery for colorectal carcinoma. Between January 1992 and December 1998, 196 colorectal cancer patients > or = 65 years (114 M; 82 F; mean age: 75 years; range: 65-92) underwent surgery. One hundred forty-five (74%) of them underwent curative surgery and emergency surgery was more common in patients > or = 75 years of age than among those younger than 75 years (39% vs 23%; p = 0.01). The overall peroperative mortality rate was 3% (n = 6). The median length of hospital stay was 18 days (range: 3-86 days). By univariate analysis, intraoperative bleeding (> or = 500 cc; p = 0.002), length of surgery (> or = 240 min.; p = 0.004), and rectal cancer (p = 0.0001) were associated with complications. By multivariate analysis, only rectal cancer (p = 0.002) was associated with complications. The overall 1, 3-, and 5-year survival rate and DFS rate were 97%, 82%, 74%, and 86%, 64% and 60% respectively. Using multivariate analysis only tumour stage (p < 0.0001) and peroperative blood transfusions (> or = 500 cc; p = 0.006) were associated with outcome. Treatment decisions in elderly patients with colorectal carcinoma should not be influenced by the chronologic age of the patient. PMID- 11109675 TI - [The surgical treatment of liver metastases from colorectal carcinoma: our experience]. AB - Data from twelve patients who had hepatic resections for colorectal liver metastases were retrospectively analyzed to determine: 1) whether the use of the ultrasonic surgical dissector and the Argon laser can significantly simplify major hepatic resections and decrease both perioperative blood loss and postoperative morbidity and mortality, and 2) whether an adequate patients selection for surgery can effectively determine an improvement in recurrence rate. We performed 4 bisegmentectomies (2 of V and VI; 2 of VI and VII); 1 trisegmentectomy (V, VI, VII); 2 left lobectomies; 1 right hepatectomy and 4 wedge resections, using both the ultrasonic surgical dissector to fractionate and aspirate the hepatic parenchyma and to clear major vascular and biliary structures and the Argon laser for the coagulation of minor vascular and biliary vessels. The resected metastases averaged 5.5 cm (range: 1.5-7.5); blood transfusion requirements were significantly reduced from previous reports, averaging only 1.25 units (range: 0.3); the average operative time was 238 minutes (110 to 420 minutes). There were no operative deaths, operative morbidity rate was 16.6. The results indicate that the ultrasonic surgical dissector and the Argon laser have made a significant contribution to our marked decrease in the average blood loss and transfusion requirement. The long-term results seems to be improved by an adequate patients selection. PMID- 11109676 TI - [Zenker's diverticulum. Apropos a case]. AB - The authors report a case of Zenker's diverticulum in a patient 72 years old who underwent surgery. The pharyngoesophageal function was investigated before and after cricopharyngeal myotomy and diverticulopexy, with oesophageal manometry. Preoperative manometry showed an incomplete relaxation of the upper oesophageal sphincter and increased of pharyngeal pressure. This diverticulum has a pulsion pathogenesis and in this case is not associated with gastroesophageal reflux. It is important to check whether an associated oesophageal pathology exist once Zenker's diverticulum has been diagnosed: X-ray examination of oesophagus and stomach are capable of identifying the presence of diverticulum as well as other pathological association. In the case showed the clinical manifestation are represented by: cervical dysphagia, sensation of foreign body while eating due to the accumulation of ingested food in the diverticulum, and noisy deglutition. The surgical treatment in this case consist of diverticulopexy with cricopharyngeal myotomy. This case is treated with diverticulopexy for two reason: because is not very big and to reduce post-operative period. In conclusion the authors shows the importance of this surgery for not very large sized pouches, and emphasise the importance of manometric and radiographic control in pre and post-operative period. This kind of surgery reduce post-operative complication and the period to stay in bed. PMID- 11109677 TI - Thyroid paraganglioma: report of a case and review of the literature. AB - A case of thyroid paraganglioma is reported. Immunohistochemically the tumor showed negativity for thyroglobulin and calcitonin and positivity for chromogranin A and S-100. Protein-positive sustentacular cells were demonstrated. The authors discuss the previous literature on these tumors. PMID- 11109678 TI - [The treatment of laparocele by means of a reticulo-laminar prosthesis (Composix Mesh). A technical note]. AB - In this paper, the authors present a modified Rives' technique for the treatment of incisional hernias. This technique requires the use of a nonabsorbable prosthesis (Composix Mesh) made of a double-knit layer of monofilament polypropylene bonded with a single layer of low-porosity e-PTFE inserted behind the rectus muscles and fixed by metallic clips. The outer side (polypropylene) encourages complete host tissue incorporation to reduce recurrences, the inner side (e-PTFE) minimizes tissue attachment and, therefore, visceral complications. PMID- 11109679 TI - [An intramural hematoma of the duodenum]. AB - Intramural hematoma of the duodenum is a rare event which is usually associated with trauma. Because of the rarity of this problem, there has been little conformity of opinions as to diagnosis and treatment of this disease. The authors report on a case of intramural hematoma of the duodenum post-traumatically occurred in a young woman. Etiopathogenesis, diagnosis and treatment of hematoma of the duodenum are thoroughly examined in the present study. Plain abdominal radiography, oral barium study, ultrasound examination, CT and RNM are diagnostic tools in this disease. It appears that most patients with intramural hematoma of the duodenum would respond well to conservative management; surgery should be reserved for those cases that remain obstructed over seven days or have evidence of peritonitis. However surgery is mandatory in cases of uncertain diagnosis. The evacuation of hematoma is considered the most effective and safest surgical treatment. PMID- 11109680 TI - The more things change... PMID- 11109681 TI - UN human rights committees and public health. PMID- 11109682 TI - Trends in coronary heart disease--has the socio-economic differential changed? AB - OBJECTIVE: To compare both trends in rates of coronary heart disease and levels of coronary risk factors between different socio-economic groups. METHODS: Rates of coronary events for men and women aged 25 to 69 years were estimated from a population-based register in the Lower Hunter Region of New South Wales from 1985 to 1993. Risk factor levels were estimated for men and women aged 35 to 64 years from three surveys of risk factors conducted in 1983, 1988/89 and 1994 in the same study population. RESULTS: There was a decline in major coronary events from 1985 to 1993. The greatest decline was for fatal coronary events, which fell by between 4.3% and 9.1% per year. Trends in event rates were similar for all socio economic groups, except for trends in non-fatal definite myocardial infarction among women. Women from the areas with high socio-economic status tended to have a greater reduction in non-fatal definite myocardial infarction compared with women from low socio-economic areas. Trends in risk-factor levels were similar except the prevalence of cigarette smoking among women from the lowest quintile of socio-economic status did not decline. CONCLUSION: Prevention strategies seem to have had a beneficial impact on this population, resulting in similar declines in rates of coronary events for all socio-economic groups. However, event rates are still high, suggesting a further reduction in mortality and morbidity is possible. This could be achieved by further reductions in smoking, cholesterol and blood pressure, especially among people from areas with low socio-economic status. PMID- 11109683 TI - Access and equity in the provision of general practitioner services for women in Australia. AB - OBJECTIVE: To assess geographical equity in the availability, accessibility and out-of-pocket costs of general practitioner (GP) services for women in Australia. METHOD: Data on general practice consultations during 1995 and 1996 for women aged 18-23 years (n = 5,260), 45-50 years (n = 7,898) and 70-75 years (n = 6,542) in the Australian Longitudinal Study on Women's Health were obtained from the Health Insurance Commission. A sub-study of 4,577 participants provided data on access to health services. RESULTS: Older women were more likely to have no out of-pocket costs for their GP consultations, but in all age groups, the proportion was lower in rural areas than in urban areas (older age: 60% rural areas, 76% capital cities; mid-age: 24% rural areas, 40% capital cities; young age: 35% rural areas, 52% capital cities). Among mid-aged women, the median out-of-pocket cost per consultation ranged from $2.11 in capital cities to $6.48 in remote areas. Women living in rural and remote areas gave lower ratings for the availability, accessibility and affordability of health services than women living in urban areas. CONCLUSIONS: This study has shown a striking gradient in financial and nonfinancial barriers to health care associated with area of residence. IMPLICATIONS: The geographical imbalance in the supply and distribution of GP services in Australia has long been recognised but inequities in the affordability of services must also be addressed. Longitudinal survey data and Health Insurance Commission data provide a means to evaluate policies designed to improve access to health services in rural and remote areas. PMID- 11109684 TI - Improved sun protection behaviour in children after two years of the Kidskin intervention. AB - OBJECTIVE: To evaluate a school-based intervention in terms of reducing children's sun exposure and improving their use of sun protection measures. METHODS: 'Kidskin' is a five-year, school-based intervention study in Perth, Western Australia, of a cohort of children who were five or six years old in 1995. The study involves three groups: control, 'moderate' and 'high' intervention. Children in the control schools received the standard health curriculum; those in the intervention schools received a multicomponent intervention, including a specially designed curriculum. Children in the high intervention group also received program materials over the summer holidays and were offered sun-protective swimwear at a low cost. After two years, parents completed a questionnaire about their child's sun-related behavior. RESULTS: Children in the intervention groups--especially the 'high' group--were reported to have had less sun exposure. This involved covering the back more often, spending more time in the shade when outdoors and wearing a style of swimsuit that covered the trunk. There was also evidence that children in the intervention groups spent less time outdoors in the middle of the day. There was little difference between groups in the wearing of hats or sunscreen. CONCLUSIONS: Our school-based intervention improved children's sun protection, but had little effect on specific behaviours that have already been vigorously promoted. IMPLICATIONS: School-based prevention campaigns would benefit from focusing on sun protection using clothing and shade, and reducing sun exposure in the middle of the day. There may be little potential to improve hat and sunscreen use. PMID- 11109685 TI - Do forecasts of UV indexes influence people's outdoor behaviour? AB - OBJECTIVE: To investigate Australian adults' awareness of the ultraviolet (UV) indexes forecast in the media, and whether these UV forecasts influence their behaviour in the sun. METHODS: A self-administered questionnaire was used on two occasions in 1997 to ask about knowledge of UV indexes shown in the media and about possible influence on outdoor behaviour. SETTING AND PARTICIPANTS: Participants were 977 residents (423 men; 554 women) of Nambour originally randomly selected in 1986 from the electoral roll, who have been followed up subsequently. RESULTS: The majority of people--92% of men and 86% of women- reported having seen or heard the UV indexes forecast during summer. Of these, significantly fewer men (107; 28%) than women (209; 46%) reported that their outdoor behaviour was influenced by knowledge of the forecast (p = 0.001). Neither age nor skin type, nor history of sunburns or skin cancer, affected knowledge of UV forecasts or their influence on behaviour. CONCLUSIONS AND IMPLICATIONS: Although most people are aware of the forecasts of UV indexes in the media, the majority do not take them into account in their outdoor behaviour. Compared with women, men were more aware of, but less influenced by, forecasts of UV indexes. Better communication of the implications of the UV indexes is needed, particularly to men, if they are to adapt their outdoor behaviour to improve their sun protection. PMID- 11109686 TI - A study of the patterns and correlates of substance use among adolescents applying for drug treatment. AB - OBJECTIVE: To inform planners by providing a psychosocial and drug-use profile of adolescents who have applied for a drug-treatment program. METHOD: The setting was a residential drug-treatment program in Sydney for adolescents from NSW and the ACT. The design was a descriptive study of consecutive program applicants over 18 months. Study participants were 14-18 years, 53% were male. Most assessments were telephone interviews. The instrument incorporated the Opiate Treatment Index, Adolescent Drug Abuse Diagnosis, Severity of Dependence Scale and Symptom Checklist 90-Revised (SCL-90-R). RESULTS: Study participants tended to be poly-substance users, mostly using cannabis, heroin and/or alcohol. Heavy use in terms of frequency and amounts of use were reported, e.g. 50% of the sample used heroin daily and the mean number of standard drinks consumed on the last day of drinking was 18. High levels of problems in the areas of social functioning, criminal activity, psychological distress, physical health, HIV risk and substance dependence were reported. For example, most participants were unemployed and 88% had committed a crime in the previous month. Higher rates of some problems were identified among females, heroin users and benzodiazepine users. CONCLUSIONS: The sample reported a high level of involvement in substance use and associated problems. The profile suggested that improvements might be difficult to achieve and to maintain. IMPLICATIONS: A comprehensive, intensive, longer-term drug-treatment program to address the number and extent of substance related problems for such adolescents is recommended. PMID- 11109687 TI - The THC content of cannabis in Australia: evidence and implications. AB - OBJECTIVE: To examine evidence on three claims that: 1) the THC content of Australian cannabis plants has increased up to 30 times; 2) problems experienced by cannabis users have increased in Australia in recent years; and 3) an increase in THC content is the most likely explanation of any increase in cannabis-related problems. METHODS: These claims were assessed by examining data: 1) on THC potency in Australia, the United States and New Zealand; 2) on cannabis-related problems; and 3) from the 1998 National Drug Strategy Household Survey on patterns of cannabis use. RESULTS: 1) Published data do not show a 30-fold increase in THC potency of cannabis but show a more modest increase in the US. 2) There is suggestive evidence of an increase in cannabis-related problems among people seeking treatment for alcohol and drug problems, juvenile offenders and young adults with psychosis. 3) There are two other more plausible explanations for these reportedly higher rates of cannabis-related problems among adolescents and young adults: (i) more potent forms of cannabis ('heads') are more widely used; and (ii) cannabis users are initiating cannabis at an earlier age, thereby increasing the prevalence of harmful patterns of use. CONCLUSIONS: There has probably been a modest increase in the THC content of cannabis, but changing patterns of cannabis use have probably made a larger contribution to any increase in rates of cannabis-related problems among young Australian adults. IMPLICATIONS: Better data on the THC content of cannabis, the extent of cannabis related problems and the ability of users to titrate the dose of cannabis would contribute to more informed debate. PMID- 11109688 TI - Evaluating aboriginal empowerment programs: the case of Family WellBeing. AB - OBJECTIVE: To evaluate the effectiveness of a Family WellBeing empowerment course. METHOD: A range of methods were used, including: theory-driven analysis of literature and project documentation; participant observation; and analysis of course participants' personal narratives against set empowerment criteria. RESULTS: Participation in the Family WellBeing course resulted in high levels of personal empowerment. The course enhanced participants' sense of self-worth, resilience, ability to reflect on root causes of problems and problem-solving ability, as well as belief in the mutability of the social environment. They were able to bring about modest, but significant, improvements in their general sense of wellbeing and those of the people around them in ways that were previously impossible. There was no evidence of organisational and community empowerment, such as stronger social networks and systems-level changes. DISCUSSION: The effectiveness of the Family WellBeing course shows the importance of resourcing Aboriginal people to develop their own programs that address trauma and other issues resulting from settler colonisation. CONCLUSIONS: The study highlights three lessons for the use of empowerment interventions to improve health conditions, particularly among socially disadvantaged groups: 1) A need to adopt an ecological approach that simultaneously addresses empowerment at multiple settings or levels. 2) A need to ensure that such programs reach a critical mass of the target group. 3) Policy-makers and practitioners need to take a longer term approach to empowerment interventions, including properly resourced longitudinal studies to document and enhance the evidence base for such interventions. PMID- 11109689 TI - Indigenous women's perceptions of breast cancer diagnosis and treatment in Queensland. AB - OBJECTIVE: To identify social, structural and personal factors among indigenous women in Queensland associated with the detection of breast cancer, and the treatment and post-treatment care and support of cancer. METHODS: Qualitative research including interviews, case studies and focus group discussions were conducted, among Aboriginal women and service providers in urban, rural and remote areas of Queensland over nine months in 1998/99. RESULTS: A range of factors were identified as influencing women's willingness to perform BSE, receive screening mammograms, and receive and complete treatment compared to the non-indigenous population. Personal history of health services, provision of information about mammography, the cost of treatment and care, and availability of personal support, all influenced women's willingness to access services and maintain treatment. Indigenous women in Queensland experience various barriers to effective and appropriate detection, treatment and care of breast cancer. CONCLUSION: Barriers to diagnosis, treatment and care can be addressed by increasing women's awareness of breast cancer and the benefits of preventative health behaviour, and improving the quality and appropriateness of health care and counselling services for Indigenous women and their families. IMPLICATIONS: Indigenous women's knowledge and practice relating to the early diagnosis and prevention of breast cancer may improve through outreach work with women, to encourage their confidence in preventative health. Women's commitment to preventive health will also be enhanced by improved quality and access to health care, and improved relationships between practitioners and patients. PMID- 11109690 TI - Comparison of estimates of population levels of physical activity using two measures. AB - OBJECTIVE: To compare estimates of population levels of 'adequate activity' for health benefit in different age and sex groups using two different measures- kilocalories (kcals) and Mets.mins. METHODS: 10,464 mid-age women (47-52 years) from the second survey of the Australian Longitudinal Study on Women's Health (ALSWH, 1998) and 2,500 men and women (18-75 years) from the 1997 Active Australia national survey, answered questions about physical activity. Kcals and Mets.mins were calculated from self-reported time spent in walking, moderate and vigorous activity, and self-reported body weight. 'Adequate activity' was defined as a minimum of 800 kcals or 600 Mets.mins. RESULTS: There were differences in the estimates of 'adequate activity' using the two methods among women participants in both surveys, but not among the male participants in the Active Australia survey. A significant proportion of the women in both surveys (6.4% of the ALSWH women and 8.5% of the Active Australia women, mean weight 60 kg) were classified as 'inactive' when the kcals method was used despite reporting levels of activity commensurate with good health. Fewer than 1% (mean weight 105 kg) were classified as 'active' using kcals when reporting lower than recommended levels of activity. Agreement between the two methods was better among men; only 3% were misclassified because of low or very high weight. CONCLUSIONS: The Mets.mins method of estimating 'adequate' activity assesses physical activity independently of body weight and is recommended for use in future population surveys, as it is less likely to under-estimate the prevalence of physical activity in women. IMPLICATIONS: Women and men aged 45-59 and women aged > 60 should be the target of specific health promotion strategies to increase population levels of physical activity. PMID- 11109691 TI - Effectiveness of the National Death Index for establishing the vital status of older women in the Australian Longitudinal Study on Women's Health. AB - OBJECTIVE: To assess the effectiveness of the National Death Index (NDI) in identifying participants in the oldest cohort of the Australian Longitudinal Study on Women's Health (ALSWH) who had died between 1996 and 1998. METHODS: Identifying information for each woman was matched with the NDI using a probabilistic algorithm and clerical review. Differences in full name, date of birth, State of residence and date of last contact were used to assess the probability of a true match. RESULTS: NDI identified 410 matches of death records for 409 women; 386 were categorised as true matches and 23 were doubtful matches. Responses to the follow-up survey confirmed that for six of the doubtful matches the women had died, 16 were alive and the vital status of one woman remained unconfirmed at 30 June 1998. Twelve deaths, known to have occurred before July 1998, were not identified through NDI. The sensitivity of the NDI for identifying known deaths was 95%. Detailed identifying information, particularly the middle name, was important for accurate identification of the vital status. CONCLUSIONS: Using surname, all given names, gender, date of birth, State of residence and age at last contact as matching variables, the NDI was an effective tool for identifying women who had died. IMPLICATIONS: Routinely collected mortality data in the NDI are useful for the practice of epidemiology. PMID- 11109692 TI - Development of a client-generated health outcome measure for community nursing. AB - OBJECTIVE: To develop a client-generated outcome measure for use in community nursing. METHOD: Participants for the study were identified from the case load of community health nurses, from a nursing home service and from residents of a retirement village. All participants had a diagnosis of venous leg ulcer (VLU) and/or type 2 diabetes. Preliminary development of the measure involved focus groups of community clients and health professionals, and pilot testing of an existing quality of life (QoL) measure, the Patient-Generated Index. The resulting Client-Generated Index was tested for reliability and validity. RESULTS: The Pearson's correlation coefficient between administration of the CGI at T1 and T2 was 0.526 (n = 51; p = 0.0001). The CGI correlated significantly with four of eight dimensions of the SF-36, and with pain as a clinical marker for VLU r = 0.54 (p = 0.001). Overall, participants with VLU reported a lower QoL (mean CGI score 2.8) compared to those with diabetes (mean CGI score 4.1). CONCLUSIONS: The CGI was developed to measure outcomes in community health settings. Some measures of its reliability and validity are demonstrated and further research is needed to validate the instrument using other client groups. IMPLICATIONS: If routine assessment and evaluation is to contribute to measures of outcome, the instruments need to be concise and acceptable to health care providers. The CGI has all these properties. PMID- 11109693 TI - Home safety assessment in the prevention of falls among older people. AB - OBJECTIVE: Home safety assessment was examined as part of a randomised trial of falls prevention interventions among older community dwellers. METHOD: Falls prevention strategies, including education and awareness-raising, exercise, home modifications and medical assessment, were trialed with 252 members of the National Seniors Association. Falls outcomes were monitored using a daily calendar diary during intervention and follow-up periods. RESULTS: The home assessment group was significantly more likely to modify their home environment than the controls (p < 0.0001). Participants, regardless of group allocation, reported a significant reduction in concern about falling (p < 0.0001). During the intervention, the home assessment group had lower incidence rates for falls and injuries than the control group, although differences were not significant. The lowered rates were sustained post-intervention. CONCLUSIONS: While the effect on falls incidence of a home safety intervention on its own could not be demonstrated, other benefits, including improved confidence attributable to awareness of such falls prevention measures, were recorded. IMPLICATIONS: The null effects of home modifications on falls prevention in this study may indicate that the program is more appropriate for the frail aged. PMID- 11109694 TI - Preference to have used a medically supervised injecting centre among injecting drug users in Kings Cross, Sydney. AB - OBJECTIVE: To assess factors affecting preference to have last injected in a medically supervised injecting centre (MSIC) among injecting drug users (IDUs) attending a needle syringe program (NSP) in Kings Cross, Sydney. METHODS: All NSP attenders over a two-day period in August 1999 were asked where they last injected, whether they injected alone and if they would have preferred to use an MSIC. This was in addition to the routine data collected, which included age, gender and last drug injected. RESULTS: Among the 178 respondents, 52 (29%) last injected in a public place and 77 (44%) last injected alone. Seventy-one per cent of all respondents would have preferred to use an MSIC. Of those who injected in public, 83% would have preferred to use an MSIC compared to 66% of those who injected in private, which was significant (p = 0.03). Age, gender, last drug injected and having injected alone did not affect preference to use an MSIC. CONCLUSIONS: Respondents' high preference to use an MSIC suggests that it may well achieve its public order and public health objectives. IMPLICATIONS: An MSIC may significantly shift current patterns of illicit drug use in Kings Cross, the community impact of which should be monitored and managed. PMID- 11109695 TI - Perception of social value predicts participation in school-based research. AB - OBJECTIVE: To investigate factors affecting the participation of schools in a serosurvey. METHODS: A telephone interview was conducted with a representative of 80 schools (response rate 92%). The schools had been randomly selected to participate in a seroprevalence survey evaluating a measles vaccination campaign of Victorian school-aged children in 1998. RESULTS: Univariate analysis suggested that responses to the interview were not influenced by school level (primary/secondary), geographic location, funding source or participation in the seroprevalence survey. There was, however, a strong association of participation in the seroprevalence survey with the perception of value to students and the value to the community. Factor analysis identified two issues: the societal value and practical issues, which explained most of the variance in participation (pseudo R2 = 0.84). CONCLUSION: The perception of the benefits of the study strongly influenced the decision by school representatives to participate in the seroprevalence survey. IMPLICATIONS: Recruitment of schools for health-related research may be improved if the number of research projects in schools is monitored and possibly restricted, and the social value of the research is emphasised. PMID- 11109696 TI - Substance use by indigenous and non-indigenous primary school students. AB - OBJECTIVE: Recent Australian research with adolescents aged 13 to 17 years has found that Indigenous youth are more likely than non-Indigenous adolescents to smoke tobacco and cannabis, although they may be less likely to use alcohol. The objective of this study was to examine whether this pattern exists among younger children. METHOD: A school-based, self-report survey was conducted in primary schools that had high proportions of Aboriginal and Torres Strait Islander children. Four schools were located in metropolitan Brisbane and three in Far North Queensland (sample n = 507 students: 270 girls, 237 boys, aged 9-13 years). RESULTS: Significant numbers of these children had started to experiment with recreational drugs. Twenty-two per cent had attempted to smoke at least one cigarette, 14% smoked in the preceding year, while 3% had smoked more than 10 cigarettes in their lives. Thirty-eight per cent had had at least one drink of alcohol, while 6% had smoked marijuana at least once. There was no significant association between Indigenous/non-Indigenous background and risk of smoking tobacco or marijuana, while Indigenous children were less likely than non Indigenous children to report experience with alcohol. CONCLUSIONS: Contrary to data from secondary school students, Indigenous youth in primary schools were not more likely than non-Indigenous children to have experimented with tobacco or marijuana, or to be frequent tobacco smokers. It appears therefore that the excessive uptake of drug use among Indigenous youth occurs in the early stages of secondary school. This finding underlines the importance of preventive education in primary schools, especially for Indigenous children who have a high risk of making the transition to drug use in adolescence. PMID- 11109697 TI - Should public health experts listen to the people or tell them what to think and do? Yes and no, more or less. AB - This article refers to Margo Kingston's analysis of Pauline Hanson's doomed election campaign in 1998 (Kingston M. Off the Rails: The Pauline Hanson Trip. Sydney: Allen and Unwin; 1999) for insights that may resolve the Chapman/Mooney debate. Should we listen to Pauline Hanson or tell her and her One Nation colleagues what to think and do? Yes, and no, more or less. If more Australian citizens were given access to the material, social, educational and other privileges that Margo Kingston has enjoyed we wouldn't have a debate. PMID- 11109698 TI - The effect of an introductory letter on participation rates using telephone recruitment. PMID- 11109699 TI - Does the ANZJPH take women seriously? PMID- 11109701 TI - Rank-based tests for interactions in a two-way design when there are ties. AB - Among the non-parametric approaches to defining and testing the hypothesis of no interactions in a two-way design, the method by Patel and Hoel is appealing because it is a function of the ranks and therefore invariant under monotonic increasing transformations of the data, and it does not require the assumption of no main effects for the approach to be valid. Recently, Wilcox compared three different methods for implementing their method, each method differing in how the appropriate standard error is estimated. In terms of Type I errors, only one of the methods gave satisfactory results in simulations, but it assumes there are no tied values. Cliff has considered the problem of tied values in detail, and his results are easily extended to the problem considered by Wilcox. This paper describes this extension and compares it to the method recommended by Wilcox. PMID- 11109700 TI - Geographic inequalities in mortality: comments on an article by Wilkinson et al. PMID- 11109702 TI - Standard errors of the principal component loadings for unstandardized and standardized variables. AB - The asymptotic standard errors of the estimates of the principal component loadings for standardized variables are derived under the assumption of multivariate normality. The standard errors are obtained for the usual unrotated case where a loading matrix is orthogonal and for the cases with orthogonally or obliquely rotated components. The corresponding standard errors for unstandardized variables and the asymptotic correlations among the estimators of the parameters for the unstandardized and standardized variables are simultaneously derived, together with the standard errors for the standardized variables. Results of a simulation illustrate the accuracy of the theoretical asymptotic standard errors and correlations. PMID- 11109703 TI - Testing treatment effects in repeated measures designs: trimmed means and bootstrapping. AB - Non-normality and covariance heterogeneity between groups affect the validity of the traditional repeated measures methods of analysis, particularly when group sizes are unequal. A non-pooled Welch-type statistic (WJ) and the Huynh Improved General Approximation (IGA) test generally have been found to be effective in controlling rates of Type I error in unbalanced non-spherical repeated measures designs even though data are non-normal in form and covariance matrices are heterogeneous. However, under some conditions of departure from multisample sphericity and multivariate normality their rates of Type I error have been found to be elevated. Westfall and Young's results suggest that Type I error control could be improved by combining bootstrap methods with methods based on trimmed means. Accordingly, in our investigation we examined four methods for testing for main and interaction effects in a between- by within-subjects repeated measures design: (a) the IGA and WJ tests with least squares estimators based on theoretically determined critical values; (b) the IGA and WJ tests with least squares estimators based on empirically determined critical values; (c) the IGA and WJ tests with robust estimators based on theoretically determined critical values; and (d) the IGA and WJ tests with robust estimators based on empirically determined critical values. We found that the IGA tests were always robust to assumption violations whether based on least squares or robust estimators or whether critical values were obtained through theoretical or empirical methods. The WJ procedure, however, occasionally resulted in liberal rates of error when based on least squares estimators but always proved robust when applied with robust estimators. Neither approach particularly benefited from adopting bootstrapped critical values. Recommendations are provided to researchers regarding when each approach is best. PMID- 11109704 TI - Exact finite-sample significance and confidence regions for goodness-of-fit statistics in one-way multinomials. AB - As multinomial processing tree models become more popular in psychology, appropriate methods for statistical inference also become more necessary. Conventional methods are all based on the asymptotic chi-square approximation to the exact multinomial test, but the accuracy of this approximation has been shown to be poor in the usual small-sample situation. This paper describes an efficient algorithm that allows the exact multinomial test to be applied without incurring prohibitive computational costs. The algorithm is well suited to addressing three different aspects of statistical inference in one-way multinomials: (i) evaluating the exact significance of a multinomial test; (ii) determining significance at a given preset level; and (iii) enumerating exact confidence regions. Examples are given that illustrate how this algorithm accomplishes each of these tasks, and an analysis of its computational cost in some small-sample situations is also provided. PMID- 11109705 TI - Statistical analysis of nonlinear structural equation models with continuous and polytomous data. AB - A general nonlinear structural equation model with mixed continuous and polytomous variables is analysed. A Bayesian approach is proposed to estimate simultaneously the thresholds, the structural parameters and the latent variables. To solve the computational difficulties involved in the posterior analysis, a hybrid Markov chain Monte Carlo method that combines the Gibbs sampler and the Metropolis-Hasting algorithm is implemented to produce the Bayesian solution. Statistical inferences, which involve estimation of parameters and their standard errors, residuals and outliers analyses, and goodness-of-fit statistics for testing the posited model, are discussed. The proposed procedure is illustrated by a simulation study and a real example. PMID- 11109706 TI - Optimal scaling of a longitudinal choice variable with time-varying representation of individuals. AB - A new method of homogeneity analysis is proposed for analysing the repeated measurements of n individuals' choices among m categories. In this method, time varying scores are assigned to each individual, while time-invariant ones are assigned to each category. For obtaining such scores, a requirement of homogeneity for time-varying individual scores is introduced in addition to the individual-category homogeneity requirement underlying the existing homogeneity analysis. The loss function based on these two requirements is minimized under a suitable normalization condition. This minimization is explicitly attained through eigen-value decomposition, and the graphical representation of the resulting scores allows us to easily find the longitudinal changes in individuals and the relationships among categories. PMID- 11109707 TI - A local influence approach to identifying multiple multivariate outliers. AB - We make use of Cook's local influence approach and its recent modification by Poon and Poon to develop measures for detecting multivariate outliers. The motivation and the foundation of the theory are geometrical and are different from classical approaches; however, whilst the proposed measure exhibits a form similar to those in the literature, it still has a considerable advantage in having transformed the classical measures to the unit interval. The new approach unifies outlier identification measures using geometrical concepts. It involves no distributional assumption or large-sample properties, and allows the flexibility of identifying outliers with respect to different metrics. The approach therefore provides a valid reason for using the various measures in complicated situations, such as in non-normal cases and in small-sample problems. PMID- 11109708 TI - Testing Thurstonian Case V ranking models using posterior predictive checks. AB - This paper presents the results of a Monte Carlo study which investigates the validity of the method of posterior predictive checks (PPC) for testing the fit of a Thurstonian Case V ranking model. The PPC method is employed as an alternative to standard goodness-of-fit tests which are of limited use even when the number of items to be ranked is small. Several test quantities are formed to assess the fit of the Case V ranking model to data for various sample sizes and for two types of violations of the Case V assumptions: heterogeneous stimulus variances and rankers from different populations. The study concludes that the PPC method is useful in detecting local and global misfits of a Thurstonian Case V model, even when the ranking data are sparse. PMID- 11109709 TI - Time-dependent Poisson counter models of response latency in simple judgment. AB - An important class of sequential-sampling models for response time (RT) assumes that evidence for competing response alternatives accrues in parallel and that a response is made when the evidence total for a particular response exceeds a criterion. One member of this class of models is the Poisson counter model, in which evidence accrues in unit increments and the waiting time between increments is exponentially distributed. This paper generalizes the counter model to allow the Poisson event rate to vary with time. General expressions are obtained for the RT distributions for the two- and the m-alternative cases. Closed-form expressions are obtained for response probabilities under a proportional-rates assumption and for mean RT under conditions in which the integrated event rate increases as an arbitrary power of time. An application in the area of early vision is described, in which the Poisson event rates are proportional to the outputs of sustained and transient channels. PMID- 11109710 TI - Confidence intervals for hidden Markov model parameters. AB - Three methods for computing confidence intervals (CIs) of hidden Markov model parameters are compared in the context of 'long' time series, T > 100, namely likelihood profiling, bootstrapping and CIs based on a finite-differences approximation to the Hessian. First it is shown that with 'long' time series computing the exact Hessian is not feasible. In simulation studies quadratic and cubic interpolation polynomials for the likelihood profiles are compared. Likelihood profiling and bootstrapping produce similar CIs, whereas the CIs from the finite-differences approximation of the Hessian are mostly too small. PMID- 11109711 TI - Statistical software for microcomputers: SigmaPlot 2000 and SigmaStat2 AB - I cannot see any advantages for SigmaStat. SigmaPlot does indeed have many excellent features and any psychologist could feel proud of many of the graphs it produces (not the box plots). As to the competition, JMP-IN produces a similar rage of graphs (and a rotating 3D plot for factor analysis), but has much less flexibility about appearance in terms of colours, fills and other features of graph elements. It can also be difficult to make different graphs the same size appear neatly on the page. STATVIEW produces less graph forms, and does not perform the non-linear regression, but has similar excellent control of the colour, form and sizing of different graph elements. Both STATIVEW and JMP-IN have well implemented 'by variable' facilities and produce graphs well linked to their associated statistical analyses. SigmaPlot wins on the flexibility of its error bars. However, EXCEL and other spreadsheets are also well worth considering, as they produce the same range of graphics and are equally flexible over error bars. An experimenter would have to be very sure that the slight advantages in flexibility of presentation from SigmaPlot outweighed the hassle of having to totally re-organize their data. PMID- 11109712 TI - Direct blood pressure monitoring. AB - Veterinary care has continued to advance by implementing more of the equipment and techniques that are commonly used in human medicine. This includes the placement of arterial catheters and pulmonary artery catheters and continuous monitoring of arterial pressure, central venous pressure, and pulmonary artery pressure. This article describes the technique for placement of appropriate catheters, the equipment that is needed, and the waveforms that are obtained when measuring direct arterial pressures, central venous pressures, and pulmonary arterial pressures. PMID- 11109713 TI - Colloid osmometry. AB - Complications related to intravascular fluid resuscitation and maintenance can become life-threatening. Overhydration and underhydration can lead to significant perfusion abnormalities and can delay or prevent recovery. A working knowledge of transcapillary fluid dynamics gives the veterinarian the basis for evaluating the cause of alterations that make up Starling's forces. By combining this knowledge with patient assessment and laboratory diagnostics, the veterinarian can make a logical decision about the best treatment to correct the Starling's force alteration. Colloid osmometry is a particularly useful tool for assessing a patient's colloid osmotic pressure. It allows the veterinarian to distinguish between reduced oncotic pressure and increased hydrostatic pressure as a cause for intravascular fluid loss or edema formation. PMID- 11109714 TI - Peritoneal dialysis in emergency and critical care medicine. AB - Peritoneal dialysis is a technique that has been used to treat acute renal failure in humans since 1923. Peritoneal dialysis is used in people to manage acute and chronic renal failure, as well as to remove dialyzable toxins (ethylene glycol, barbiturates, and ethanol), reduce severe metabolic disturbances, and for the treatment of peritonitis, pancreatitis, uroabdomen, hypothermia, and fluid overload. In veterinary medicine, acute renal failure is the prevailing indication for dialysis. This report will discuss the pathophysiology of peritoneal dialysis, indications, and contraindications. Catheter selection and placement will be reviewed. Types of dialysate solution will be discussed and the protocol established for instituting peritoneal dialysis. The report will conclude with a discussion of potential complications and methods to minimize them. PMID- 11109715 TI - Hemodialysis. AB - Hemodialysis is a therapeutic procedure that uses the extracorporeal circulation of a patient's blood to ameliorate the azotemia, fluid, electrolyte, and acid base abnormalities characteristic of the uremic syndrome. Hemodialysis is principally used for the management of acute and chronic renal failure that is refractory to conventional medical therapy. Additional applications include acute intoxications (e.g., ethylene glycol poisoning) and preoperative conditioning of renal transplant recipients. Hemodialysis is a technically demanding procedure that requires an extensive array of sophisticated delivery equipment and specifically trained and dedicated staff to perform, monitor, and ensure the integrity and safety of the procedure in critically ill patients. The advent of neonatal dialysis delivery equipment has ensured that hemodialysis is a feasible, efficacious, safe, and indispensable therapy for dogs and cats with life threatening renal failure. Increased awareness and acceptance of hemodialysis as an effective renal replacement therapy coupled with increased owner demand guarantee a bright future for hemodialysis as a viable therapeutic modality. PMID- 11109716 TI - Strategies for mechanical ventilation. AB - With the advancement of veterinary critical care medicine, an increasing number of veterinary patients are being supported with positive-pressure ventilation. Animals with potentially reversible ventilatory failure (PaCO2 > 60 mmHg) caused by neuromuscular disease or pulmonary parenchymal disease or with pulmonary parenchymal disease causing hypoxemia (PaO2 < 60) despite supplemental oxygen are candidates for ventilatory support. The equation of motion for the respiratory system is defined and is used to describe the potential interactions between the patient and the ventilator. Commonly used modes of ventilation are described in terms of control and phase variables. The intent of this report is to aid clinicians in choosing an optimal ventilatory strategy for each patient that will best achieve the desired physiologic goals with minimal detrimental side effects. PMID- 11109717 TI - Weaning from mechanical ventilation. AB - Patients that require positive pressure ventilation to maintain sufficient alveolar ventilation or pulmonary gas exchange may eventually reach a point in the course of their care wherein mechanical ventilation is no longer necessary. This process of transferring the work of breathing from the ventilator back to the patient is referred to as ventilator weaning. The term "ventilator weaning" may be used to refer to all methods by which this transfer of workload may be accomplished. In many patients, particularly those with short-lasting or readily correctable causes of respiratory insufficiency (e.g., general anesthesia), the discontinuation of positive pressure ventilation may be easily achieved. Indeed, in patients awakening from general anesthesia, the axiom "awake enough to blink, awake enough to breath" may prove to be a sufficient guideline. However, in those patients requiring long-term mechanical ventilatory support, the process can prove to be both frustrating and exceptionally challenging. It is of crucial importance to identify those patients that may be successfully weaned because of both the financial impact of prolonged intensive care unit hospitalization and the risks imposed on the patient by the process of positive pressure ventilation. To be able to predict which patients may be ready to be weaned from the ventilator requires an understanding of the balance between the work of breathing (ventilatory load) and the ability of the patient's respiratory pump to meet those needs (ventilatory capacity). The management of patients experiencing difficulty during the weaning process requires that the clinician recognize imbalances between ventilatory load and capacity and to correct these imbalances once identified. PMID- 11109718 TI - Transvenous cardiac pacing. AB - Transvenous pacing therapy is a life-saving technique for patients with clinically significant bradyarrhythmias. For most symptomatic bradyarrhythmias in small animals, there is no effective substitute for cardiac pacing. The methods employed for pacemaker placement, although potentially time-consuming, are not technically difficult. This article discusses the indications, techniques, clinical decision-making, and potential complications associated with temporary and permanent transvenous cardiac pacing. PMID- 11109719 TI - Epidural analgesia in veterinary critical care. AB - Epidural analgesia has minimal systemic effects and is a useful technique for relieving pain in critical care patients. Before administration, patients must be thoroughly assessed to identify any preexisting conditions that preclude the safe use of this technique. Analgesia can be achieved by administration of local anesthetics, opioids, alpha 2 agonists, or a combination of these analgesic agents. Concurrent administration of more than one drug allows the synergistic interaction of these agents and generally improves the level of analgesia achieved, lengthens the duration of action, and lowers the dose of each drug required to achieve analgesia. Complications of epidural techniques are infrequent and include both iatrogenic and idiopathic problems, most of which have no permanent sequelae. This review provides a detailed description of the epidural analgesia technique and lists multiple sources of specialized supplies necessary for either single injection or epidural catheter placement. It also provides direction for monitoring the critical care patient with an epidural catheter. PMID- 11109720 TI - [Human immunodeficiency virus infection and "Recommendations 2000". A step forward on the line of long-term therapeutic strategies. PMID- 11109721 TI - [Temporal evolution of tuberculosis and human immunodeficiency virus infection in a population cared for in a hospital in Malaga]. AB - BACKGROUND: We have studied the incidence of tuberculosis++ disease, HIV infection and their association during a period of 6 years using samples analysed in a third level hospital laboratory. MATERIAL AND METHOD: 21,242 samples for mycobacteria and 63,425 for HIV antibodies were analysed between 1993 and 1998. The protocol used for mycobacteria consisted of Lowenstein-Jensen, hemoculture, biochemical tests for identification and DNA probe. The diagnosis of HIV was performed using screening with mix EIA HIV 1 + 2, confirmed with Western-blot. Spearman coefficient correlation was used for study of tendency. RESULTS: 1,613 samples (7.5%) positive for mycobacteria from 566 patients (98 females and 428 males) aging between 31 and 40 years (46.9%) were detected. The highest incidence for tuberculosis was observed in 1995 (49.2 x 10(5) and was followed by a decreasing linear tendency. Mycobacterium tuberculosis (96%) and M. bovis (2.7%) were most frequent agents. 2,295 samples (3.6%) showed anti-HIV (406 females and 1,889 males). 54.1% of the infected patients belonged to the age interval 21 to 40 years. The highest incidence was observed in 1994 and was followed by a significant decrease (p < 0.05). The percentage of patients co-infected with tuberculosis-HIV was 39.7% with a maximum in 1995 followed by a linear decrease. CONCLUSIONS: The incidence of both infections and co-infection was very high in the first triennium and was followed by a progressive decrease. The decrease of HIV preceded tuberculosis. The result suggest a possible epidemiological correlation between both infections. PMID- 11109722 TI - [Resistance to penicillin and other antimicrobials in 301 clinical isolates of Streptococcus pneumoniae]. AB - BACKGROUND: The aim of this study was to assess the susceptibility to penicillin of Streptococcus pneumoniae clinical strains and to analyze the association between penicillin resistance and cefotaxime and cefixime activity in S. pneumoniae isolates with decreased sensitivity to penicillin. METHODS: 301 S. pneumoniae clinical strains were isolated from patients during 1995-1996. Susceptibility to penicillin, cefotaxime, cefepime, erythromycin, chloramphenicol, tetracycline, cotrimoxazole and ciprofloxacin were studied. RESULTS: 38.2% isolates were penicillin-susceptible and 61.8% were penicillin resistant; 20.6% showed high-level resistance. Resistance rates to erythromycin, chloramphenicol, tetracycline, cotrimoxazole and ciprofloxacin were, respectively, 30.9, 30.2, 40.9, 66.4, and 13.3% overall, and 54.8, 54.8, 61.3, and 93.5% in the 62 strains with high-level resistance to penicillin. Strains resistant to cefotaxime and cefepime were 13.9 and 14.9%, respectively. MIC50 and MIC90 for cefotaxime and cefepime in penicillin-resistant strains were 0.5 and 1 mg/ml. CONCLUSIONS: A high proportion of S. pneumoniae isolates showed resistance to penicillin, in agreement with other Spanish reports. Moreover, resistance to penicillin was significantly associated (p < 0.001) with resistance to erythromycin, chloramphenicol, tetracycline and cotrimoxazole, but not with ciprofloxacin. MIC50 and MIC90 for cefotaxime and cefepime were similar, and lower than those for penicillin in penicillin-resistant pneumococci strains. PMID- 11109723 TI - [Onychomycoses caused by non-dermatophytic filamentous fungi in Cadiz]. AB - BACKGROUND: The incidence and clinical significance of other than dermatophytes fungi or moulds causing onychomycosis is unknown, because they may be colonising organisms rather than pathogen. This report presents the results of a study conducted between 1997 and 1999 to determine the incidence and aetiology of onychomycosis by non-dermatophytic filamentous fungi in the population of Cadiz (Spain). PATIENTS AND METHODS: Diagnosis of onychomycosis was performed by direct microscopic examination, culture and, some times, by histologic examination, on samples from 610 patients with clinical suspected fungal nail infections. RESULTS: Among 196 (32%) cases of ungual mycosis detected, 29 (15%) of them were caused by non-dermatophytic filamentous fungi, presenting positive direct microscopy and repeated cultures. Superficial and distal onychomycosis were the most frequent clinical types. Twenty two patients had onychomycosis of toenails. The highest incidence was found in women and subjects over the age 40. Scopulariopsis spp. (n = 11), Aspergillus spp. (n = 6), Alternaria spp. (n = 5) and Fusarium spp. (n = 4) were the most common fungi. Occasionally, Acremonium spp. and Scedosporium spp. were isolated. CONCLUSION: The incidence of onychomycosis caused by opportunistic fungi is not well known. For their diagnosis, it is important to select correctly the appropriate site for specimen collection, as well as direct microscopy and fungal cultures. The incidence of onychomycosis is high in Cadiz (Spain), being higher in women and older people. Predispondent factors are not always identified in the patients. Toenails were infected more than fingernails in both sexes. The results of our study suggest that Scopulariopsis spp. is an important agent of onychomycosis. Epidemiological investigations should be performed in every country in order to determine the fungal species responsible of onychomycosis. PMID- 11109724 TI - [Haemophilus influenzae infections in children less than 5 years of age in the community of Murcia during the 1992-1999 period]. AB - BACKGROUND: Invasive disease due to Haemophilus influenzae has changed substantially since the introduction of the conjugated vaccine. This report studies the incidence and the clinical-epidemiological characteristics of invasive H. influenzae disease in children under five years of age during 1992-94 (before vaccination), 1995-97 (voluntary vaccination) and 1998-99 (obligatory vaccination). PATIENTS AND METHODS: The study was performed by reviewing the clinical histories of 39 patients with H. influenzae isolates from sterile samples, according to microbiology data. The reference population's subgroup of the study, 40,322 children under 5, comprises 60% of the total in our community. RESULTS: The overall incidence of invasive H. influenzae disease was 12.1/100,000 children under 5 and 15.7, 12.4 and 6.2 for the following periods respectively: before vaccination, during voluntary vaccination, and obligatory vaccination. All cases except one of them, were produced by H. influenzae type b and they were seen in children under 3. Meningitis accounted for more than half of the cases (51.3%). Fever was the most frequent sign (38 of 39 cases). Epiglottitis was the cause of the highest average hospital stay (20.8 days). All the patients were treated with cefotaxime, but half were also administered other antibiotics. Sequelae were seen in 7 cases, with no deaths. CONCLUSIONS: The incidence of invasive H. influenzae disease was drastically reduced and no cases were seen in the last year. However, one vaccinated patient suffered a bacteremic etmoiditis. This case and the possibility of infection due to non-b serotypes requires ongoing surveillance for these infections. PMID- 11109725 TI - [Recommendation of GESIDA (AIDS Study Group)/National Plan on AIDS with respect to the anti-retroviral treatment in adult patients infected with the human immunodeficiency virus in the year 2000 (I)]. AB - OBJECTIVE: To update the recommendations for antiretroviral therapy in adult HIV infected persons according to the new scientific advances and the existence of new antiretroviral drugs in the last two years. METHODS: The antiretroviral therapy recommendations have been condensed by a panel of experts from the Spanish AIDS Study Group (Grupo de Estudio de sida-GESIDA) of the Spanish Infectious Diseases and Clinical Microbiology Society (SEIMC) and from the Clinical Advisory Panel of the Secretariat of the Spanish National Plan on AIDS (SPNS) of the Ministry of Health. Three levels of evidence have been established depending if the data came from randomised and controlled studies, from cohort or case-control studies or from descriptive studies and expert opinions. For that purpose we have reviewed the advances in HIV pathophysiology and results of efficacy (clinical, virologic and immunologic) and security (toxicity) from clinical trials involving antiretroviral therapy lasting at least 12 months, from cohort studies and pharmacokinetic and security data of antiretroviral drugs, presented in international conferences or published in biomedical journals in the last two years. In each situation we have established either to recommend or to consider or not recommend antiretroviral therapy. RESULTS: Nowadays, antiretroviral therapy consisting of at least three drugs constitutes the election therapy for chronic HIV infection, since it delays clinical progression, increases significantly the survival and diminishes hospital admissions and associated costs. The decision to start antiretroviral therapy must be based upon three elements: presence or absence of symptoms, plasma viral load and CD4+ cells counts. Thus, in asymptomatic cases with a high CD4+ cells count (> 500/microL) and low viral load (< 10,000 copies/ml by branched DNA [bDNA] or < 20,000 copies/ml by reverse-transcription polymerase chain reaction [RT-PCR] or nucleic acid sequence based amplification [NASBA]) we recommend to delay antiretroviral therapy. In symptomatic patients we recommend to start it, and in asymptomatic patients, we could recommend or consider antiretroviral therapy initiation depending on the risk of progression, established by the viral load and the CD4+ cells count. In any case, if therapy is started, the objective must be to reach an undetectable viral load (< 50 copies/ml). The adherence to antiretroviral therapy plays a key role for its initial moment and for the duration of the antiviral response, antiretroviral therapy can achieve a restoration of cellular immunity in the advanced patients. There are few therapeutic options in failing patients due to cross-resistance. Resistance studies can be useful in this setting. The toxicity is a new and limiting factor of antiretroviral therapy which requires to look for new therapeutic options. Antiretroviral therapy criteria for acute infection, pregnancy, post-exposure prophylaxis and when to use resistance testing are discussed. CONCLUSIONS: In this moment, there is a more conservative attitude towards starting antiretroviral therapy than in previous recommendations in which a virus eradication was considered. On the other hand, the high number of disposable drugs, the more sensitive monitorization methods (plasma viral load) and the possibility of performing resistance studies make therapeutic strategies more dynamic and individualised for each patient and situation. In any case, it is mandatory to ensure a perfect adherence to antiretroviral therapy from the patients. PMID- 11109726 TI - [Meningococcus carriers: an enigma at the end of the 20th Century]. PMID- 11109727 TI - [Exfoliative erythromatous eruption in an 8-year-old girl]. PMID- 11109728 TI - [Anaphylactic reaction and epigastralgia in a patient with an antecedent of pulmonary hydatidosis. PMID- 11109729 TI - [Catheter preservation at 4 degrees C. Influence on quantitative cultures]. PMID- 11109730 TI - [Thrombophlebitis of the innominate vein in a patient with a pacemaker]. PMID- 11109731 TI - [Kingella kingae in a 10-year-old girl]. PMID- 11109732 TI - [Levofloxacin and tuberculosis of renal localization]. PMID- 11109733 TI - [Disseminated infection with Mycobacterium avium complex in immunocompetent patients]. PMID- 11109734 TI - [Escherichia coli O157:H7 in patients infected with the human immunodeficiency virus: presentation of a case]. PMID- 11109735 TI - [Vulvovaginal candidiasis resistant to azoles]. PMID- 11109736 TI - [Urinary infection caused by Enterococcus faecalis with a mucoid phenotype: an unusual morphotype]. PMID- 11109737 TI - [Questions and answers about joint prosthesis infections]. PMID- 11109738 TI - [Comparative epidemiologic study of caries in German and Hungarian adolescents]. AB - The aim of the study was to assess the trend of caries prevalence in German and Hungarian population sample. The authors examined 15-16-year-old Hungarian adolescents in Debrecen and Budapest in 1997 (451 persons), then re-examined them in 1999 (377 persons). These data were compared to those of 15-16 and 17-18-year old adolescents examined in the Eastern part of Germany (Plauen) in 1991 (668 persons) and in 1995 (586 persons). The results showed significant caries reduction in Germany. The DMFT values of 17-18-year-old adolescents were nearly half in Germany (x = 5.2) compared to those of Hungary (x = 9.0). This fact suggests the necessity of introduction of effective preventive programs including salt fluoridation in Hungary. PMID- 11109739 TI - [Adsorption of human serum proteins to titanium dioxide]. AB - In order to find an in vitro biochemical model for investigation of osseointegration in terms of dental implantology, the aim of the present study was to analyse the adsorption of human serum proteins to titanium dioxide. Titanium dioxide powder was suspended in human serum. After incubation and centrifugation the TiO2 with the adsorbed proteins were washed with distilled water, ethylendiaminetetraacetic acid (EDTA) supplemented with ammonium hydrogencarbonate (NH4HCO3) solution, and sodium dodecylsulphate (SDS), after centrifugation the supernatant fluid was collected and SDS polyacrylamide gel and native (Biomidi) gel electrophoresis were conducted to determine the type of the adsorbed proteins. Our results show, that albumin was adsorbed to TiO2, but it could be easily removed. The adsorption of a 94 kDa protein was much stronger than the other proteins. The method seems to be useful in the investigation of the protein adsorbing ability of differently treated titanium implant surfaces. PMID- 11109740 TI - [Cantilever bridges--a cross-sectional study]. AB - Life expectancy and functional performance of the cantilever bridges is determined by the length of the cantilever pontic(s) and by the number and localisation of the abutment teeth. No overall epidemiological survey covering this topic was found in the literature. 529 distally cantilevered bridges were evaluated by the authors. There were 310 upper and lower cases. The following parameters were recorded: length of the cantilever part (number of units), the relationship of the cantilevered pontics and the bridge, and the position of the fixed restoration in the dental arch. Previously the cantilevered bridge design was not accepted in Hungary but in spite of this fact a large number of distally cantilevered bridges were made in the country. Most of the examined cantilever bridges are properly designed: two abutments are supporting one pontic (342 cases, 65.56%). However some of the solutions are too risky: one abutment, one cantilevered pontic (110 cases, 20.45%), or two abutments, two cantilevered pontics (53 cases, 9.85%). PMID- 11109741 TI - Occupational skin diseases among the nurses in the region of Lodz. AB - We examined 223 nurses from the Lodz region, referred to the Nofer Institute of Occupational Medicine in 1995-99 because of suspected occupational dermatoses. The diagnosis of contact allergy was based on the positive results of patch tests, and immediate allergy to common allergens and latex on the results of prick tests, as well as on the determinations of specific IgE antibodies. Contact allergy was diagnosed in 66.4% of nurses. The most frequent sensitisers were quaternary ammonium compounds (benzalkonium) (23.8%), nickel (21.5%) and formaldehyde (20.6%); allergy was somewhat less frequently caused by thimerosal (14.3%), fragrances (12.1%), glutaraldehyde (10.8%), cobalt (9.9%), thiurams (6.7%) or glyoxal (4.9%). Allergy to rubber was diagnosed in 40 patients (17.9%), including 25 cases of immediate allergy to latex, 8 cases of contact dermatitis from thiuram rubber curing accelerators, and 7 cases of mixed allergy. It has been concluded that the contact with disinfectants (40.8% of allergic nurses), metals (30.9%) and rubber (17.9%) was the most frequent source of allergy in this group of medical personnel. PMID- 11109742 TI - Effects of exposure to tobacco smoke in pregnancies complicated by oligohydramnios and premature rupture of the membranes. I. Concentration of Cd and Pb in blood and Zn, Cu, Cd and Pb in amniotic fluid. AB - To assess the exposure to tobacco smoke in pregnant women with oligohydramnios, idiopathic or caused by premature rupture of the membranes (PROM), cotinine concentrations were measured, using enzyme-like immunosorbent assay (ELISA). In women with idiopathic oligohydramnios (22-31 weeks of gestation), serum cotinine concentration was 1010 +/- 445 micrograms/L which provides evidence that women of this group were heavy smokers. In these women, significantly higher Cd concentrations in blood and amniotic fluid were found as compared to other pregnant women. A positive correlation between Cd concentrations in blood and amniotic fluid was observed (PROM r = 0.784; p < 0.001; idiopathic oligohydramnios r = 0.7118; p < 0.02). In oligohydramnios cases of both types, Cd concentration in amniotic fluid was over two times and Pb concentration ten times lower than blood concentrations of these metals, whereas amniotic fluid Zn concentration was two times lower than that found earlier in women with normal pregnancy. In the group of women with idiopathic oligohydramnios who were mostly exposed to tobacco smoke, a considerably larger number of still births and new borns with CNS disorders than in PROM cases, were observed. Zn deficiency at increased exposure to Cd and Pb could play a significant role in etiology of these abnormalities. A positive correlation was found between Zn and Cu concentrations (r = 0.862; p < 0.05) in PROM cases which indicates regular transport of trace metals to the fetal ovum. The condition of infants born to this group of women was much better, and prematurity was the only complication of pregnancy. PMID- 11109743 TI - Film badges for personal dosimetry of roentgen radiation. AB - The paper describes the calibration procedure for and data obtained from routine X-ray dosimetry service in Poland. In 1998 the number of readouts amounted to 152,000. The dosimetry concerned 31,281 workers in 3,113 enterprises. The annual collective dose was assessed to be 16.7 man.Sv. The analysis of the calibration quality is discussed as well. Only 2.9% of all points lay outside the quality curve while the number of the acceptable outliers was 5%. PMID- 11109744 TI - Does potassium dichromate induce apoptosis in lymphocytes? AB - The aim of this work was to investigate the possibility of apoptosis in the human peripheral blood lymphocytes after treatment with potassium dichromate (K2Cr2O7), a potential occupational carcinogenic and mutagenic agent. Lymphocytes were stimulated by phytohemagglutinin, cultured for 72 h and incubated with either 0.2 mM or 0.4 mM K2Cr2O7 for the last 24 h or 48 h of culture. The condensation and margination of chromatin with emerging 'half-moon' structure, characteristic of apoptosis were observed. Phosphatidylserine displaced from the inner to outer side of the cellular membrane in 54% of cells after a 48-h incubation with 0.4 mM K2Cr2O7 (annexin-V+/PI-); 39% of these cells were of late apoptotic--secondary necrotic form (annexin-V+/PI+). Following the agarose gel electrophoresis of DNA, a 'ladder pattern' typical of apoptosis, was found. The results of the present study demonstrate that K2Cr2O7 can induce in the human peripheral blood lymphocytes changes similar to apoptotic ones. PMID- 11109745 TI - Long-term effects of acute exposure to chlorphenvinphos on behavioural responsiveness to amphetamine and scopolamine in rats. AB - We investigated the effect of acute exposure to chlorphenvinphos (CVP) (2-chloro 1 (2,4-dichlorophenyl)-vinyl-diethyl-phosphate), an organophosphate anticholinesterase, on the amphetamine- and scopolamine-induced open-field locomotion in Wistar rats. CVP was administered at a single i.p. dose of 1.0 mg/kg (1/10 of LD50). In part of the rats cholinesterase (ChE) was determined. Three hours after CVP injection, the ChE activity decreased by about 27%. It returned to the preinjection level within 14 days after the exposure. In the behavioural part of the experiment, the animals were challenged with 1.0 mg/kg amphetamine or 0.75 mg/kg scopolamine three weeks after CVP exposure, i.e. after a period of time sufficient for cholinesterase recovery. It has been found that in the CVP-exposed rats, the behavioural responses to amphetamine or scopolamine challenge (the increase in locomotor activity) was significantly reduced compared to the controls. This suggests that acute exposure to CVP produced an increase in cholinergic activity which persisted long after ChE activity had returned to normal. PMID- 11109746 TI - Subchronic inhalation toxicity of 1,2,3-trimethylbenzene (hemimellitene) in rats. AB - Toxic effects of exposure to 1,2,3-trimethylbenzene (hemimellitene) in the condition of subchronic inhalation experiment were examined. Rats were exposed to vapours of hemimellitene at concentrations of 123 mg/m3, 492 mg/m3 and 1230 mg/m3, 6 h/day, 5 days/week for 3 months. After termination of a 3-month inhalation, animals were necropsied. Blood samples were obtained and selected organs were weighed and prepared for histological examinations. Subchronic inhalation exposure to hemimellitene resulted in an overall, low systemic toxicity. There were no changes in body weight gain and food consumption. At a concentration of 1230 mg/m3, the increase in relative liver weight was observed in male rats. It was accompanied by slight increase in sorbitol dehydrogenase activity. The increase in alkaline phosphatase activity was found in females only. Some disturbances in haematological parameters, characterised by the decrease in red blood cells and slight increase in white blood cells, segmented neutrophils and lymphocytes were observed in rats at high exposure concentration of 1230 mg/m3. The pulmonary lesions as well as the increased number of goblet cells and interstitial lung parenchyma infiltration were noted in male and female rats from the highest exposure groups. PMID- 11109747 TI - The role of reactive oxygen species in the development of malignancies. AB - Numerous studies on animal models and cell cultures confirm the role of free radicals/reactive oxygen species and oxidative damages in the process of the initiation and promotion of neoplastic diseases. Also epidemiological studies attribute an important role to disturbances in the pro- and antioxidative potential in the development of malignancies and highlight the essential function of antioxidants to protect the cells from the attack of free oxygen radicals. The current literature data on the parameters of the anti- and prooxidant status in patients with malignant diseases and their role in pathogenesis of the disease is reviewed in this article. PMID- 11109748 TI - Occupational health from the perspective of future accession of Poland to the European Union. Brief Summary Report of a Workshop organized by the Nofer Institute of Occupational Medicine Lodz, Poland, 30 June-1 July, 2000. PMID- 11109749 TI - Polish bibliography of occupational medicine, 1999. Part 1. PMID- 11109750 TI - Non-small cell lung cancer(NSCLC)--spectrum, management and prospects. PMID- 11109751 TI - Non-small cell lung cancer: disease spectrum and management in a tertiary care hospital. AB - OBJECTIVES: To review the spectrum of presentation and the surgical management of Non Small Cell Lung Cancer (NSCLC) and the role of various diagnostic modalities in predicting the post-operative stage and the correlation of the post-operative stage with the chances of recurrence. SETTINGS: The Aga Khan University Hospital, Karachi--a tertiary care referral center in Paksitan. METHODS: This is a retrospective study of medical records of all the patients who were managed at the Aga Khan University Hospital (AKUH) between 1988 and 1998. The patients with a diagnosis of lung cancer were identified using the ICD-9-CM coding system and the data was analyzed for patients with NSCLC only. RESULTS: A total of 773 patients were admitted with a diagnosis of lung cancer at AKUH over 10 years period. Out of these 21 (2.71%) underwent staging mediastinoscopy and 20 (2.58%) patients underwent exploratory thoracotomy and biopsy without any resection, as the disease was found to be unresectable. Only 18 (2.32%) patients underwent surgical resection. There were 15 males and 3 females and the mean age was 53 years. Mean duration of symptoms was 12 months and cough and haemoptysis were the main presenting symptoms. Most of the tumors were located on the right side. CT scan and mediastinoscopy were mainly used to stage the disease. Complete surgical resection including en-bloc resection of adjacent structures was attempted, when possible. Median follow up was 24 months and the recurrence rate was 39%. There was no significant correlation between post-op stage and recurrence. CONCLUSION: It is concluded that most of the patients present at the advanced stage and resection is possible only in a small number of patients. The size of primary tumor and local extension should not contra-indicate surgery in patients with negative mediastinal nodes and without distant metastasis as it can be performed safely. All pulmonary lesion in the adults must be thoroughly investigated as early diagnosis and complete resection is the only key to cure and long term survival. PMID- 11109752 TI - A comparative study of positive pressure ventilation via laryngeal mask airway and endotracheal tube. AB - OBJECTIVE: To investigate the use of Laryngeal Mask Airway (LMA) and its comparison with the endotracheal tube for positive pressure ventilation. SETTING: A tertiary care teaching hospital. METHODS: Fifty adult ASA I and II patients undergoing peripheral limb surgery were randomly allocated to 2 groups for LMA or endotracheal tube insertion. A standardized anaesthetic technique was used. The groups were then compared regarding haemodynamic changes on insertion as well as removal of LMA and ETT and any complications that occurred were noted. RESULTS: The haemodynamic response to insertion was significantly attenuated (p < 0.05) in LMA group as compared to ETT group. The cardiovascular response to extubation was not significantly different between the groups. A higher incidence of coughing and mild hypoxaemia at extubation was noted in ETT group as compared to LMA group (p < 0.05) and blood was detected in 4 cases after LMA removal. CONCLUSION: It is concluded that the use of LMA during positive pressure ventilation is safe in selected cases. There is an attenuated haemodynamic response to insertion of LMA as compared to endotracheal tube which will be beneficial in certain patients e.g., those with ischaemic heart disease, vascular disease and hypertensives. PMID- 11109753 TI - Relationship of skin ulcers and physical deformity with employment status and compliance with health promotion in leprosy. AB - OBJECTIVE: To determine if recurrence of ulcers and physical deformity in leprosy is associated with employment status and compliance with health promotion advice. METHODS: Between April-August 1992, a cohort of 55 consecutive leprosy patients admitted with skin ulcers were studied for ulcer recurrence, physical deformity (taking into account neuromuscular and skeletal damage), employment status, compliance with health promotion advice and knowledge of the disease. RESULTS: High grade physical deformity was present in 34/55 (62%) patients while recurrent ulceration occurred in 40/55 (75%) patients. With regard to employment status, the odds of high grade physical deformity were significantly higher for street traders and unemployed compared to semiskilled and skilled workers (odds ratio 4.2, 95% confidence interval 1.01-19.8, p = 0.03). There was a trend of higher odds of recurrence of ulcers for street traders and unemployed compared to semiskilled and skilled workers (odds ratio 2.3, 95% confidence interval 0.5-9.4, p = 0.2). With regard to health promotion, there was poor compliance with advice about protective footwear and care of insensitive extremities. Level of knowledge about the disease and its prevention was also inadequate. CONCLUSION: Physical deformity was associated with lack of reasonable employment among leprosy patients. There was poor compliance with preventative advice. Health promotion strategies should be directed toward targeted health education and prevention of physical deformities. PMID- 11109754 TI - Risk factors for stunting and wasting at age six, twelve and twenty-four months for squatter children of Karachi, Pakistan. AB - OBJECTIVE: A high proportion of stunting and wasting in children under-five has been reported from developing countries. This paper presents the nutritional status of a two year cohort of urban squatter children in Karachi, Pakistan and assesses risk factors for wasting and stunting at the reference ages of six, twelve and twenty-four months. METHODS: A birth cohort of 738 children were visited at specific intervals by trained nurses to collect information on anthropometric measurements, feeding practices and intercurrent illnesses. Socioeconomic and demographic information included water and sanitation facilities, availability of electricity, type of house construction material and average monthly income. Information about the mother's reproductive history was also obtained. RESULTS: At two years the proportion of stunting and wasting was 41.8% and 10.6% respectively. Intrauterine growth retarded children had a higher risk of stunting and wasting at all reference ages as compared to children who were appropriate for gestational age. In the logistic regression models, intrauterine growth retardation was the only significant risk factor that remained in all models at each reference age. CONCLUSION: The consistent association of IUGR for stunting and wasting adds to the growing body of evidence that by improving maternal health we will ultimately break the vicious cycle of malnourishment and improve the health and well-being of future generations. We suggest interventions to improve the nutritional status of Pakistani urban children living in squatter settlements focused on mothers and children. PMID- 11109755 TI - DNA ploidy analysis of borderline epithelial ovarian tumours. AB - OBJECTIVE: Borderline epithelial ovarian tumours not uncommonly pose a great difficulty to surgical pathologists as morphologically they may show very similar features as those of malignant epithelial tumours except invasion. However it is important to separate these from their invasive counterparts because of their superior prognosis. Recently, attention has been focussed on the prognostic value of flow cytometric analysis of DNA ploidy in borderline epithelial ovarian tumours. The purpose of this study is to investigate whether flow cytometric analysis of cellular DNA content acts as a useful adjunct to the histopathological diagnosis of borderline malignancy. MATERIALS AND METHODS: Fifteen histologically confirmed borderline serous epithelial tumours of the ovary were selected. Samples were analyzed on a FACScan flow cytometer using the software MODFIT. A total of 10,000 nuclei were counted each time. RESULTS: The mean CV for the 15 cases was 3.67 (Range 2.4-5.0). In the DNA histograms a diploid sample was defined as one that had a single Go/Gl peak. An aneuploid tumour was defined as one that displayed an additional distinct peak. All 15 cases of borderline serous epithelial tumours showed a diploid stemline with DNA index between 0.9-1.10. CONCLUSION: This study suggests that aneuploidy if ever demonstrated in histologically confirmed borderline tumours should prompt extensive sampling of the tumour and a close follow up. PMID- 11109756 TI - Exposer rate of hepatitis E (IgG) in a selected population of children and adults in Karachi. AB - AIMS: To see exposure rate of hepatitis E (IgG) in 100 apparently healthy children and adults. SUBJECTS AND METHODS: Sera from 100 healthy children aged 1 day to 10 years and 100 healthy adults aged 18-45 years were analysed for exposure to hepatitis E (IgG) using ELISA. RESULTS: Two samples from children were excluded from the study due to improper storage leaving 98 samples for analysis. Of 98 sera from children 19.4% were positive for IgG indicating previous exposure to hepatitis E. The exposure rate increased with age and was 10% in children below 1 year of age, 14% at 2 years, 19% at 3 years and 28% at 10 years. In adults overall exposure was 16%. There was no predominance of either sex in both the groups and all individuals belonged to middle to lower socioeconomic strata. CONCLUSION: An exposure of 19% in children and 16% in adults indicates high faecal contamination of drinking water and re-addressing of the issue of use of boiled water on individual level, supply of potable water on the government level and a need to produce a vaccine on international level. PMID- 11109757 TI - Clinical protocols: introduction to a useful strategy in clinical practice. PMID- 11109758 TI - Congenital leukaemia presenting as bilateral renal masses. PMID- 11109759 TI - Aplastic anemia in a patient with factor IX deficiency. PMID- 11109760 TI - Endocrine cell dysplasia (nesidioblastosis): a relatively unrecognized entity in Pakistan? PMID- 11109761 TI - Clinical spectrum of systemic lupus erythematosus at the Aga Khan University Hospital. AB - BACKGROUND: Systemic lupus erythematosus is a disease of unknown etiology, which at onest may involve only one organ system or be multisystemic. The aim of our study is to determine the clinical presentation of SLE patients presenting to AKUH to establish whether guidelines laid down about this disease are in agreement with our experience. METHODS: A retrospective log review was carried out at AKUH, based on data obtained from 165 files of individuals admitted to the hospital over a period of 12 years with a confirmed diagnosis of SLE. RESULTS: From the sample size of 165, 143 (86.7%) were females and 22 (13.3%) males. The mean age of diagnosis was 30.9 years. Frequency of symptomatology was observed to be in the following order: systemic 78.8%, musculoskeletal 63% and hematological 60.6%. Oninvestigation ANA levels were positive in 112 patients. CONCLUSION: Our result lead us to conclude that the classification set forth by the American Rheumatological Association is applicable to patients presenting with SLE in our setting. PMID- 11109762 TI - [Serial measurement of anti-interleukin-8 IgG autoantibody in cerebrospinal fluid of infants with bacterial meningitis]. AB - We serially measured concentrations of interleukin (IL)-8 and anti-IL-8 IgG autoantibody in cerebrospinal fluid of infants with bacterial meningitis, and also measured these concentrations in cerebrospinal fluid obtained from infants without meningitis on admission. We have reported that the IL-8 concentration in cerebrospinal fluid of infants with purulent meningitis rapidly decreases after the initiation of therapy. Thus, in the present study, the IL-8 concentration in infants with purulent meningitis only before the initiation of therapy was significantly higher compared with that in infants without meningitis. However, the concentration of anti-IL-8 IgG autoantibody was still high after the initiation of therapy. The concentration of anti-IL-8 IgG autoantibody was significantly higher compared with that in infants without meningitis until the 15th day after the initiation of therapy. The time lag between the decrease of IL 8 and anti-IL-8 IgG autoantibody demonstrated in the present study could be used to indicate the past presence of a large amount of IL-8, even if the IL-8 concentration was already low. PMID- 11109763 TI - [Drug sensitivity, conjugative R plasmids and plasmid profiles of Salmonella isolated from humans with infectious enteritis]. AB - Using 92 Salmonella strains isolated from patients suspected of having infectious diseases of the intestinal tract who visited 13 hospitals in Japan during the six years between 1991 and 1996, we investigated the drug susceptibility, prevalence of conjugative R plasmid, and the plasmid profiles. 1) Of the bacterial isolates tested, 52.2% showed drug-resistance. Regarding the drug-resistance patterns, 70.8% of the isolates were resistant to a single drug, while 29.2% were multi drug-resistant. 2) Dividing the resistance patterns by the serotypes, among Salmonella Enteritidis isolates, single-drug resistance to SM was the most frequent, being detected in 27 isolates. Single-drug resistance to NA and two drug resistance to SM/TC were the second-most frequent, each being detected in isolates. Among Salmonella Hadar isolates, four isolates showed two-drug resistance to SM/TC, and one isolate showed single-drug resistance to TC. Among Salmonella Typhimurium isolates, one isolate each showed three-drug resistance to ABPC/CER/KM and KM/TC/CP. Among Salmonella Agona isolates, one isolate each showed two-drug resistance to SM/TC and single-drug resistance to SM. Among Salmonella Derby isolates, two isolates showed single-drug resistance to SM. 3) The prevalence of conjugative R plasmid was investigated in 48 drug-resistant isolates, and six isolates (12.5%) contained the plasmid. 4) The prevalence of the plasmid was investigated in 29 drug-resistant S. Enteritidis isolates, and 22 isolates (75.9%) contained the plasmid. These isolated were classified by the plasmid profiles into types H1 to H7. 5) Regarding the plasmid profiles of the S. Enteritidis isolates, a position corresponding to 60 Kbp was the most frequently detected in 90.5%. PMID- 11109764 TI - [Survey on Toxocara cati in domestic cats]. AB - A total of 542 domestic cats aged from 1 month to 20 years were investigated for Toxocara cati (T. cati) by means of fecal examination. The cats were classified in the groups according to their kept environment (indoor or outdoor) and age (1 6 month, 7 month-1 year, 2-3 year, 4-5 year and over 6 year). The indoor kept cats were major in all ages, and the friendly relationship was suggested between domestic cats and humans. The parasitic rates of T. cati a higher percentage was noted in the groups of indoor kept cats of 1-6 month (27.1%) than that of outdoor kept cats of 1-6 month (17.9%), 7 month-1 year (18.5%) and 4-5 year (14.3%). The high parasitism of T. cati in indoor kept 1-6 month cats, which are may be closely connected to humans, is an important problem from the public health of view, because of the direct source of visceral larva migraine. The parasitism of T. cati in outdoor kept cats is also a serious matter as the origin of environmental contamination. It suggests that a positive sanitary instruction against an owner seems necessary for prevention of visceral larva migraine in humans. PMID- 11109765 TI - [Comparison between cytomegalovirus hepatitis and Epstein-Barr virus hepatitis in healthy adults]. AB - In order to determine the factors responsible for the differentiation of cytomegalovirus (CMV) hepatitis and Epstein-Barr virus (EBV) hepatitis in previously healthy adults, the clinical features and laboratory data of both types of hepatitis were retrospectively analyzed. CMV hepatitis showed a tendency to increase in our department. In comparison with EBV hepatitis, CMV hepatitis occurred in significantly older hosts than EBV hepatitis. We found that lymphadenopathy, cough and sore throat was more common in EBV hepatitis than in CMV hepatitis. The number of peripheral white blood cell count and atypical lymphocytes, and serum GOT, GPT, LDH and CRP levels of CMV and EBV hepatitis showed no significant differences. PMID- 11109766 TI - [Species and serovar-distribution, and drug-resistance of Shigella strains isolated from imported and domestic cases during 1995-1999 in Tokyo]. AB - A total of 290 Shigella strains consisting of 180 imported strains and 110 domestic strains isolated during 1995-1999 in Tokyo were examined regarding their species and serovar-distribution and their drug-resistance. In both groups, S. sonnei (70.0% in the imported strains, 80.9% in the domestic strains) was found to be the most prevalent species, followed by S. flexneri (20.0% in the imported strains, 19.1% in the domestic strains). S. dysenteriae and S. boydii were only isolated in the imported cases. Among the S. flexneri serovar, 1b, 2a, 6, 2b, and 3a were predominant in the imported strains, whereas 1b and 2a were predominant in the domestic strains. Provisional new serovar Shigella strains were isolated from 11 imported cases and 2 domestic cases. The drug-resistance test using 9 drugs (CP, TC, SM, KM, ABPC, ST, NA, FOM, and NFLX) showed that 92.2% of the imported strains and 94.5% of the domestic strains were resistant to some of the drugs tested. Drug-resistance patterns of the resistant strains varied in 25 types. Among those, a triple drug-resistance type with TC.SM.ST was found as the most frequent pattern in both groups. None of the strains were resistant to NFLX. PMID- 11109767 TI - [Evaluation of influenza virus type A detection kit]. AB - We evaluated a new Influenza Virus Type A Detection Kit (InfluA-AD "Seiken") and compared it to virus isolation and RT-PCR using throat swab specimens. Our results were as follows: 1) From 13 specimens in which Influenza virus type A was isolated, InfluA-AD "Seiken" detected 12 while RT-PCR detected 9. 2) From 7 specimens in which Influenza virus type B was isolated, InfluA-AD "Seiken" detected 0 while RT-PCR detected 1. 3) From 2 specimens in which Adeno virus was isolated, InfluA-AD "Seiken" detected 0 and RT-PCR detected 0. 4) From 28 specimens in which no virus was isolated, InfluA-AD "Seiken" detected 6 while RT PCR detected 8. InfluA-AD "Seiken" showed a sensitivity of 92.3% and specificity of 83.8% when compared to virus isolation. We conclude that the new test will be useful for the diagnosis of Influenza virus type A infection particularly in big commercial laboratories. PMID- 11109768 TI - [Influence of macrolide therapy on microorganism of chronic respiratory tract infections]. PMID- 11109769 TI - [Verotoxin producing ability of verotoxin-producing Escherichia coli strains isolated from fecal specimens of healthy persons is lower than that of patients]. PMID- 11109770 TI - [A clinical study in the collagen disease patients developed pulmonary tuberculosis during corticosteroid administration]. AB - The study subjects consisted of 14 pulmonary tuberculosis (PTB) patients with collagen disease. They are under corticosteroid treatment and the mean age is 56.4 years. The length of time from the development of collagen disease to the development of PTB averaged 4.1 years. The breakdown of collagen disease are SLE (6 patients), MCTD (3 patients), PN (2 patients), and PSS, PM, Sjogren syndrome (1 case, each). Thirteen cases were bacilli positive by the sputum examination on admission to our hospital. Chest X-ray findings on admission revealed cavitation in 3 cases and non-cavitation in 11 cases, of which 5 cases had miliary tuberculosis. Corticosteroid preparation had been administered to all of the 14 cases for more than one year. The mean dose of corticosteroid preparation administered when PTB developed was 13.9 mg (prednisolone) and it was more than 20 mg in 8 cases. The median duration from the start of the respiratory symptoms to diagnosis was 39.2 days. The delay in the discovery exceeding 1 month were seen in 9 cases. In the cases of collagen disease, when the disease course extends over a long period of time, and even when the dose of corticosteroid preparations are decreased, there is a need to be note on the risk of developing PTB. There are many non-cavitary cases with sputum smear positive. The fact suggested that an appropriate diagnosis is need so that the discovery of PTB should not be delayed. PMID- 11109771 TI - [Detection of rifampin-resistant Mycobacterium tuberculosis by line probe assay (LiPA)]. AB - A recently described reverse hybridization-based line probe assay is used for the rapid detection of the mutations in the rpoB genes of rifampin-resistant Mycobacterium tuberculosis and for the identification of the M. tuberculosis complex. A multicenter study that included 5 laboratories was performed to evaluate the line probe assay in comparison with the in vitro susceptibility test. A total of 406 mycobacteria isolates which were composed of 103 rifampin resistant and 230 rifampin-susceptible M. tuberculosis isolates, and 73 mycobacteria other than tubercle bacilli (MOTT), were subjected to this study. All 333 M. tuberculosis isolates were discriminated correctly from MOTT bacilli by a line probe assay. Concordance rates with sequencing results for five wild type probes (S probes) and four specific mutations (R probes) for detecting the mutations in the rpoB genes were both 100%. The overall concordance rate with the in vitro susceptibility testing results was 98.5% (328 of 333 isolates). These results indicate that a line probe assay kit may be useful for the rapid diagnosis of rifampin-resistant tuberculosis. PMID- 11109772 TI - [Comparison of chest CT findings between suspected and definite cases of primary pulmonary M. avium complex infection]. AB - It is very difficult to treat pulmonary infection with MAC, because we have few effective drugs against this organism. In this situation, an early diagnosis and treatment are very important to manage this disease. We evaluated chest CT scans of the primary pulmonary MAC infection which had no underlying lung diseases and no immunocompromised diseases such as HIV infection. We defined suspected cases of pulmonary MAC infection as cases in which abnormal features of chest CT scans were recognized but frequency of detection of organisms of MAC did not fulfil the diagnostic criteria for atypical mycobacteriosis according to Japanese Mycobacteriosis Research Group of the National Chest Hospitals. CT scans of suspected cases were compared with the definite cases. Results obtained were as follows: 1. In classification by CT scans of primary pulmonary MAC infection, the proportion of localized type and diffuse type was the same both in suspected and definite cases. In localized type, more tuberculosis-like pattern was seen in definite cases. 2. In suspected cases, characteristic features of CT scans of primary pulmonary MAC infection were recognized in the same frequency as in definite cases. 3. In pulmonary tuberculosis-like type, definite cases showed more cavitary lesions than suspected cases. These results showed that a careful long term follow-up of suspected cases with frequent bacteriological tests of sputum and chest CT scannings was important for early diagnosis of primary pulmonary MAC infection. PMID- 11109773 TI - [A case of tuberculous aneurysm of the aorta]. AB - We reported a rare case of tuberculous aneurysm of the aorta managed successfully with urgent surgical therapy. A 35-year-old woman was admitted to our hospital complaining of fatigue and hemoptysis. Laboratory tests showed severe anemia, slight liver dysfunction, elevated level of C-reactive protein, and negative syphilis serologies. The chest roentgenogram revealed widening of right upper mediastinum, two nodular shadows in right middle lobe, and left-sided infiltration shadow with pleural effusion. The pleural effusion was bloody and its level of adenosine deaminase was normal. Culture of pleural effusion specimen remained negative. A computed tomography scans of the chest revealed an aortic aneurysm on the aortic hiatus. Rapid increase in pleural effusion was followed by hemothorax a few hours later. After operation, she received antituberculosis therapy. Histopathologically, the resected lung showed inflammatory process including granulation of giant cells and epithelioid cells. The specimens of the aortic aneurysm revealed rupture of whole layer of aortic wall and inflammatory cell infiltrations. These findings suggested that the case to be a tuberculous aneurysm of the aorta. Therefore, we diagnosed the case as the rupture of tuberculous aneurysm of the aorta. PMID- 11109774 TI - [Immunology in the 20th century--progress made in research on infectious and immunological diseases]. AB - The new era of the modern medicine was opened 100 years ago by Robert Koch and Louis Pasteur who demonstrated that various infectious diseases were caused by their respective microbes. Koch discovered Mycobacterium tuberculosis, the causative agent of tuberculosis. The first breakthrough in the modern medicine to combat against infectious diseases was the discovery of anti-diphtheria toxin antibody by E.A. von Behring and S. Kitasato. The concept of immunity--immune from disease--has thus been established. The immune response between antigen and antibody sometimes provides the host with a harmful effect. The concept of allergy was introduced by Richet and later by Prausnitz and Kustner. Why the same immune response leads to the different outcome, immunity or allergy had not been made clear until the discovery of IgE by Drs. Kimishige and Teruko Ishizaka in 1968: The IgG antibody plays a role in immunity whereas IgE antibody is involved in allergy. Tuberculin skin reaction which is well known as the diagnostic tool for mycobacterial infection was studied by M. Chase in 1945 demonstrating that it was able to be transferred to the healthy individual by immune cells but not by antibody. The immune response is now categorized into two; soluble immunity- immediate type allergy and cell-mediated immunity--delayed type allergy. The rapid progress in the molecular biology in the past decades has also accelerated the progress in immunology, several of which include discovery of two types of lymphocytes; T and B cells; concept of two T cells; Th1 and Th2 cells; and the discovery of cytokines which regulate immune cell responses. The mechanism of the immune response is now understood at the gene level. Several immunological diseases can now be successfully treated by controlling the levels of cytokines involved. For example, refractory rheumatoid arthritis is now under control by the administration of recombinant soluble TNF receptor molecules to the patients. The complete human genome sequence is currently under investigation. We can now envisage the advent of the days when every disease can be diagnosed and intervened at the gene level. PMID- 11109775 TI - [Prospect of chemotherapy in the 21st century]. AB - "Golden Era in Chemotherapy" has begun with the discovery of penicillin in the early 1940's and lasted for two decades during which many antibiotics were discovered. However, the once-believed bright prospect that every infectious disease could be eliminated on the earth by the discovery of antibiotics had to be canceled owing to the emerging of drug-resistant microbes. It was indeed a rat race. We are now at the point when we have to seek another way to combat infectious diseases: One possible way might be not to eradicate the microbes but to coexist with them so long as they do no harm to the human hosts. The first step of infection with pathogens to the host is the adherence of the microbes to the surface of host cells. Therefore, the method how to inhibit this adhesion of microbes to the host cells may provide a new tool to prevent the development of infectious diseases without elimination of microbes from the host. This is just an example of strategy by which humans and pathogens coexist at peace and should be taken into consideration for the development of new-type antibiotics or "anti infective drugs" in the 21st century. The analysis of genome sequences has been accelerated recently for various pathogenic bacteria one by one. New targets in the pathogenic microbes for the development of new antibiotics can, therefore, be determined from the genetic point of view. The discovery of antibiotics has indeed been the history of collection of innumerable species and/or strains of bacteria from the soils to search for the biologically active anti-pathogenic agents. The current progress in the technology of molecular genetics, however, will certainly make it possible to search for active molecules by DNA technology; bacterial DNA but not whole microorganisms from the soil is to be transformed into the conventional bacteria and searched for active molecules with combat against pathogens. PMID- 11109776 TI - [Riddles in human tuberculous infection]. AB - Tuberculosis is indeed an infectious disease caused by Mycobacterium tuberculosis. However, only a small percentage of individuals infected develops overt disease, tuberculosis whereas the infected bacilli persist alive years long within the vast majority of persons infected but remained healthy. There are several riddles or enigmas in the natural history of M. tuberculosis infection in humans. Some of them are as follows: 1. What is the virulence of M. tuberculosis? 2. How does M. tuberculosis persist dormant within the host? 3. What determines the development of disease from remaining healthy after infection with M. tuberculosis? 4. What is the mechanism of "endogenous reactivation" of dormant M. tuberculosis within the host? 5. Can we expect more potent anti-TB vaccine than BCG in near future? Most of these issues cited above remain unsolved. What is urgently needed today to answer correctly to these questions is the production of appropriate animal model of tuberculosis infection which mimics human tuberculosis. Murine TB does not reflect human TB at all. What characterizes the mycobacterial organism is its armour-plated unique cell wall structure which is rich in lipid and carbohydrate. Cord factor or trehalose dimycolate (TDM), the main component of cell wall, has once been regarded as the virulence factor of mycobacteria. Cord factor is responsible for the pathogenesis of TB and cachexia or even death of the patients infected. However, cord factor in itself is not toxic but exerts its detrimental effect to the host through the excessive stimulation of the host's immune system to produce abundant varied cytokines including TNF-alpha. How to evade this embarrassing effect of mycobacterial cell wall component on the host immune system seems very important for the future development of better TB vaccine than the currently used BCG. PMID- 11109777 TI - [Tuberculosis control of urban areas in Japan]. AB - The rates of tuberculosis remain high in urban areas. The declining speed of tuberculosis incidence rate in urban areas has been slower than other areas. Efforts and resources to tuberculosis control must be concentrated on urban locations to eradicate tuberculosis in Japan. 1. Tuberculosis control in a public health center of urban area: Teru OGURA and Chiyo INOGUCHI (Toshima City, Ikebukuro Public Health Center, Tokyo Metropolitan) A wide range of TB control measures is implemented by public health centers, such as a patient registration, home-visit guidance, contact examination in urban areas. Directors of every health center have the direct responsibility for tuberculosis control measures in their jurisdiction. Ikebukuro is urban areas where there are many offices, shopping and amusement facilities. Urban people is often on the move looking for job, so public health centers are often not easy to carry out contact examinations as planned. In recent years, homelessness has been recognized as a growing urban social problem. Their incidence of tuberculosis is high. Special TB control program must be carried out in urban areas. 2. Tuberculosis Control in Tokyo Metropolitan: Kazumasa MATSUKI (Department of Infectious Diseases and Tuberculosis, Bureau of Public Health, Tokyo Metropolitan) There has been a steady decline in the TB wards. The beds for TB patients are running short and even smear positive TB cases cannot be put in a hospital without waiting several days. Staffs of an urban emergency department must protect tuberculosis infection by environmental controls of emergency room. Tokyo Metropolitan government supports the engineering improvements of emergency room to hospitals. Directly observed therapy for tuberculosis patients at a district has been implemented to complete their therapy. On DOT, a trained health worker observes the patient take anti-TB medication. 3. Usefulness of Molecular Epidemiologic approach on Tuberculosis Control: Atsushi HASE (Osaka City Institute Laboratory of Health and Environment) DNA fingerprinting establishes the genetic relatedness of Mycobacterium tuberculosis isolates and has become a powerful tool in tuberculosis epidemiology. To use DNA fingerprinting to assess the efficacy of current tuberculosis infection-control practices. Combining conventional epidemiologic techniques with DNA fingerprinting of M. tuberculosis can improve the understanding of how tuberculosis is transmitted. Patients were assigned to clusters based on mycobacterial isolates with identical DNA fingerprints. Clusters were assumed to have arisen from recent transmission. We analyzed M. tuberculosis isolates from patients reported to the tuberculosis registry by RFLP techniques. These results were interpreted along with demographic data. Patients infected with the same strains were identified according to their RFLP patterns, and patients with identical patterns were grouped in clusters. RFLP patterns of high incidence districts have more variations than other areas. This suggests that the source of tuberculosis infection are quite diverse and complicated. Tuberculosis patients may accumulate to high incidence districts from other places after infection. 4. Structure of High Incidence of Tuberculosis and Control Plan in Osaka City: Yoichi TATSUMI (Bureau of Infection Control, Osaka City Office) The case notification rate in Osaka City is the highest in Japan. That of all TB cases and smear positive TB cases was 1573 and 216 per 100,000 population in 1997 at Airin District in Osaka City. The main reason for this highest incidence rate is that there are many homeless people and it is a mobile population. Most of residents are daily laborers. They come from all over Japan and stay there, mainly in rented rooms, to look for jobs. Thousands of homeless people also live in tents on streets or in parks. We are making to new strategic plan to intensify tuberculosis control measures throughout the city. Osaka city government h PMID- 11109778 TI - [Tuberculous infection and prevention]. AB - In Japan, the number of tuberculosis infected patients has been increasing again, especially in the elderly. The incidence of newly registered tuberculosis patients who are over 70 years is 34%. The outbreak of tuberculosis also has been increasing, and has become a serious social problem. The elderly have a high risk of developing tuberculosis because of their immunosuppressed condition due to underlying disease and aging. Elderly active tuberculous people also confer a risk of tuberculosis infection to the non-tuberculosis infected young generation. In this symposium, we discussed about 1) the tuberculosis outbreak related to the strategy for tuberculous prevention, 2) the health examination to detect tuberculous people in the middle-aged and elderly, 3) the nosocomial transmission of tuberculosis in the hospital, 4) the nutritional damage and immunosuppressive state in elderly people related to developing active tuberculosis in latent tuberculous infection, and 5) the organ transplantation and tuberculosis focusing on living related liver transplantation. Seven cases of tuberculosis outbreak were reported in Osaka from 1989 to 1998, 2 cases in the hospital and 5 in others. Tuberculous infection index (maximum sputum Gaffky score multiplied by the number of months of persisted cough) was 8 and 15 in the hospital, 3, 0, 84, 14, and 27 in others. Three cases (43%) were observed in persons with less than 10 of this index. It is essential to evaluate carefully for tuberculosis outbreak in extraordinary examination, if the tuberculous infectious index is low. There are various immunosuppressive patients with malignancy and other underlying disease in the hospital, so we have to pay careful attention for tuberculous outbreak when there is an active tuberculous patient. In these seven cases of extraordinary examination for tuberculous epidemic, only one (0.4%) of 241 cases who received isoniazide prophylactic therapy developed active tuberculosis. Isoniazide prophylactic therapy was an important strategy for the prevention of tuberculous outbreak. (Isamu TAKAMATSU, Osaka Prefectural Habikino Hospital, Osaka). It is essential to prevent tuberculosis in the elderly to achieve tuberculosis control in public health. The health examination was an important strategy for the detection of tuberculosis in the middle-aged and elderly. The incidence of tuberculosis detection is only 9.3% by the health examination at this age. However, the health examination has well detected active tuberculosis patients in elderly (34.5%), and also 16.8% in smear positive cases with pulmonary cavitary formation. The early detection of persons with active tuberculosis is essential, and further discussion regarding cost-performance and accuracy of the health examination for tuberculosis should also be essential. Prophylactic therapy of isoniazide also might be considered for the high risk middle-aged and elderly people with underlying diseases. (Masako OMORI et al., Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association, Tokyo). The cases of nosocomial transmission of tuberculosis in the hospital have been increasing. In younger persons, the incidence of tuberculosis infected nurses and doctors is relatively higher than healthy control. The transmission of tuberculosis from elder active tuberculosis patients to healthy non-tuberculosis infected medical workers has been well recognized. It is very essential to follow guidelines for the prevention of tuberculous transmission in the hospital from the Japan Tuberculosis Society including routine tuberculin skin test for fresh medical workers. Primary education for tuberculosis in medical and nursing school is also an important strategy. Further discussion might be essential that BCG could prevent tuberculosis transmission in tuberculin skin test negative adults. (ABSTRACT TRUNCATED) PMID- 11109779 TI - [Industrial medicine on the threshold of XXI century]. AB - In XXI century priority problems of industrial hygiene, besides practical solutions concerning health care for workers and general population, would be further fundamental researches on mechanisms underlying human response to occupational factors, their regulation, diagnosis of premorbid conditions, rehabilitation measures. PMID- 11109780 TI - [Occupational diseases of sensorimotor system in builders]. AB - The study covered influence of occupational factors on health state in 2 groups of workers engaged into facing (59 plasterers and 74 house painters) and matched in age, sex and length of service. Work of plasterers and house painters appeared to encounter risk of occupational sensomotor system disorders specified not only by type and hardness, but also prevalent direction of local muscular load. PMID- 11109781 TI - [Cerebral control over locomotor function in occupational hand disease caused by local muscular overstrain]. AB - Predominant orientation of local muscular load to proximal or distal portions of shoulders and upper limbs considerably influences features of changes in brain functions and blood supply. Characteristic for plasterers, localization of myogeloses in upper part of trapezium muscles causes not only impaired vertebrobasillar circulation, but also venous cerebral stasis and caudal medullar dysfunction. Clinical symptoms are vestibular disorders, depressed functions of hearing, chronic venous cerebral congestion, posture-tonic disorders. Local loads on distal portions of limbs have other mechanisms of cerebral and peripheral disorders. PMID- 11109782 TI - [Fibrogenicity of dust emitted by highly aluminiferous refractories]. AB - The authors proved variable clinical and pathogenetic features of disease in workers under long occupational exposure to dust of kaolin and its baking products. Therefore, special experimental studies should cover kaolin and mullite as primary and final product in entire technologic sequence of high-alumina refractories production. Natural refractory clay and mullite dust were administered to rats intratracheally during chronic experiments. Lungs of the rats exposed to mullite dust demonstrated reliable changes of lipid content in 1 month and those of hydroxyproline content--in 3 months. When exposed to intratracheal administration of kaolin and mullite dust, respiratory system develops typical changes--slow development of benign diffuse and sclerotic form of pneumoconiosis that is associated or, more frequently, preceded by chronic dust bronchitis. PMID- 11109783 TI - [Clinical features of respiratory diseases caused by mullite dust]. AB - The authors evaluate clinical peculiarities of chronic bronchitis, silicotuberculosis and pneumoconiosis (mullitosis) in workers exposed to highly aluminiferous clay. PMID- 11109784 TI - [Cytogenetic disorders in workers of coal-tar chemical industry]. AB - The authors studied frequencies of chromosomal aberration (CA) and sister chromatid exchanges (SCE) in workers of coal-tar chemical plant (34 individuals). The studies revealed significant increase of CA frequency (7.97 +/- 0.63%), when compared with reference groups 3.37 +/- 0.39% and 3.64 +/- 0.37%. PMID- 11109785 TI - [Social problems of mandatory medical insurance]. AB - Efficacious system of social insurance and social security is a system of economic relations, where every member is provided with guarantee of certain living standard necessary for development and utilization of the member's abilities and which can aid in case if some of the abilities are lost. PMID- 11109786 TI - [Aspects of combined action of lead, copper and zinc]. AB - Character and intensity of metabolic, functional and morphologic changes in subchronic combined action of lead and zinc, especially lead and copper result from their antagonism. Based on experimental data, the authors recommend that, considering industries with "lead-copper" or "lead-zinc" combinations in air, sanitary surveillance should use the same MACs as for the isolated metals. PMID- 11109787 TI - [Microcirculation system status in chronic pesticide intoxication]. PMID- 11109788 TI - [Prevalence of peptic ulcer in sailors and fishermen of the North Sea basin]. PMID- 11109789 TI - [New information on chemical toxicity]. PMID- 11109790 TI - [Human fundamental rights and reproductive health in international documents]. PMID- 11109791 TI - Preoperative assessment of trigeminal neuralgia and hemifacial spasm using constructive interference in steady state-three-dimensional Fourier transformation magnetic resonance imaging. AB - Results of microvascular decompression (MVD) for trigeminal neuralgia (TN) and hemifacial spasm (HFS) may be improved by accurate preoperative assessment of neurovascular relationships at the root entry/exit zone (REZ). Constructive interference in steady state (CISS)-three-dimensional Fourier transformation (3DFT) magnetic resonance (MR) imaging was evaluated for visualizing the neurovascular relationships at the REZ. Fourteen patients with TN and eight patients with HFS underwent MR imaging using CISS-3DFT and 3D fast inflow with steady-state precession (FISP) sequences. Axial images of the cerebellopontine angle (CPA) obtained by the two sequences were reviewed to assess the neurovascular relationships at the REZ of the trigeminal and facial nerves. Eleven patients subsequently underwent MVD. Preoperative MR imaging findings were related to surgical observations and results. CISS MR imaging provided excellent contrast between the cranial nerves, small vessels, and cerebrospinal fluid (CSF) in the CPA. CISS was significantly better than FISP for delineating anatomic detail in the CPA (trigeminal and facial nerves, petrosal vein) and abnormal neurovascular relationships responsible for TN and HFS (vascular contact and deformity at the REZ). Preoperative CISS MR imaging demonstrated precisely the neurovascular relationships at the REZ and identified the offending artery in all seven patients with TN undergoing MVD. CISS MR imaging has high resolution and excellent contrast between cranial nerves, small vessels, and CSF, so can precisely and accurately delineate normal and abnormal neurovascular relationships at the REZ in the CPA, and is a valuable preoperative examination for MVD. PMID- 11109792 TI - Cerebral oximetry for the detection of cerebral ischemia during temporary carotid artery occlusion. AB - The near-infrared spectroscopy cerebral oximeter was assessed as a monitoring device for detecting and/or predicting cerebral ischemia during carotid endarterectomy (CEA) and the balloon occlusion test in 24 patients, 12 males and 12 females aged 28 to 77 years (mean 59.9 years). Tolerance testing of complete internal carotid artery (ICA) occlusion by balloon inflation for 20 minutes was performed in nine patients (cerebral aneurysm 6, neck tumor 3) and CEA was performed in 15 patients. The probe of the cerebral oximeter was placed on the forehead of the affected side and regional cerebral oxygen saturation (rSO2) was monitored continuously during all procedures. Stump pressure was measured just after ICA occlusion. Collateral circulation detected by digital subtraction angiography was classified into three groups: good, moderate, or poor. Stump pressure was 41-90 mmHg (mean 61.3 mmHg) in the good collateral circulation group, 40-43 mmHg (41.5 mmHg) in the moderate group, and 14-30 mmHg (23.8 mmHg) in the poor group. Change in rSO2 after ICA occlusion was +3.5(-)-4.2% (mean 1.6%) in the good collateral circulation group, -1.2(-)-6.6% (-3.2%) in the moderate group, and -2.4(-)-10.2% (-6.6%) in the poor group. Changes in rSO2 were significantly different between the good and poor collateral circulation groups (p < 0.01). A greater than 5% fall in rSO2 was observed in 0 of 15 patients in the good collateral circulation group, one of five in the moderate group, and three of four in the poor group. The cerebral oximeter is a useful, real-time, non-invasive method to measure brain oxygenation during CEA, skull base surgery, or other procedures which need to evaluate brain ischemia. A fall of greater than 10% from the rSO2 baseline value is dangerous, but less than 5% is safe. PMID- 11109793 TI - Increased expression of deoxyribonucleic acid methyltransferase gene in human astrocytic tumors. AB - The relationship between the grade of astrocytic tumor and the expression of deoxyribonucleic acid methyltransferase (DNA-MTase) gene was examined. The levels of DNA-MTase messenger ribonucleic acid (mRNA) were measured by semiquantitative reverse transcriptase-polymerase chain reaction in surgical specimens from 12 astrocytic tumors (4 astrocytomas, 6 anaplastic astrocytomas, and 2 glioblastomas) and two normal brain tissues, and in four glioma cell lines. Compared to normal brain tissues, the levels of DNA-MTase mRNA were increased by 16- to 55-fold in low grade astrocytomas, and significantly increased by 200- to 4500-fold in high grade astrocytomas (anaplastic astrocytomas and glioblastomas) and more than 4500-fold in glioma cell lines. In situ hybridization with paraffin embedded surgical specimens of human astrocytic tumors showed DNA-MTase mRNA was abundantly expressed in high grade astrocytomas. The detection of increased DNA MTase expression in astrocytic tumor indicates involvement in the tumorigenesis and suggests that blocking of this change with specific inhibitors may offer new therapeutic strategies for malignant astrocytic tumors. PMID- 11109794 TI - Infratentorial arteriovenous malformation associated with persistent primitive trigeminal artery--case report. AB - A 58-year-old male presented with a rare association of an infratentorial arteriovenous malformation (AVM) and ipsilateral persistent primitive trigeminal artery (PPTA) manifesting as sudden onset of headache and vomiting. Computed tomography revealed subarachnoid hemorrhage, and digital subtraction angiography demonstrated an infratentorial AVM mainly fed by the left superior cerebellar artery via the left PPTA. The patient refused radical treatment for the AVM, and was conservatively treated. The adjacent AVM may have been important in the preservation of the PPTA, as blood flow into the infratentorial AVM via the PPTA and the hemodynamic stress to the PPTA could have disturbed the spontaneous closure of the PPTA. PMID- 11109795 TI - Cavernous sinus hemangioma treated with gamma knife radiosurgery: usefulness of SPECT for diagnosis--case report. AB - A 79-year-old female presented with cavernous sinus hemangioma manifesting as double vision due to right oculomotor and trochlear nerve pareses. Computed tomography and magnetic resonance imaging revealed bony erosion and a right cavernous sinus tumor with "tail sign" after contrast medium administration. Thallium-201 (201Tl) single photon emission computed tomography (SPECT) showed low uptake within the tumor, and technetium-99m-human serum albumin diethylenetriaminepenta-acetic acid SPECT disclosed high uptake within the tumor. 201Tl SPECT usually shows very high uptake in meningiomas and malignant tumors, so the tumor was considered to be an unrelated benign tumor. The patient underwent partial resection of the tumor. Histological examination of the specimen confirmed cavernous hemangioma. The oculomotor nerve paresis partially improved. Gamma knife radiosurgery was carried out 4 months after the operation. The tumor markedly shrank with full recovery of extraocular movement 6 months after radiosurgery. SPECT is useful for distinguishing cavernous sinus hemangiomas from other cavernous tumors. Radiosurgery should be performed after partial resection or biopsy for cavernous sinus hemangiomas and may be the initial treatment for patients with small cavernous sinus hemangioma if the diagnosis can be established based on neuroimaging. PMID- 11109796 TI - Gliosarcoma associated with a huge cyst--case report. AB - A 55-year-old female presented with a unique case of gliosarcoma with a huge cystic component manifesting as loss of consciousness, left-sided hemiparesis, and anisocoria. Computed tomography demonstrated a large cyst in the right frontal lobe, and enhancement of the mural nodule after administration of contrast medium. Emergent operation was performed. Xanthochromic fluid was aspirated, and the tumor was resected. The histological diagnosis was gliosarcoma based on the presence of gliomatous and sarcomatous components. She underwent a second operation because of tumor regrowth 3 weeks after the first operation. The postoperative course was satisfactory during radiation therapy with 60 Gy and chemotherapy. The diagnosis of gliosarcoma was difficult to make preoperatively because of the neuroradiological findings similar to low-grade gliomas. Gliosarcoma should be included in the differential diagnosis of huge cystic tumors. PMID- 11109797 TI - Brain metastasis of epithelioid sarcoma--case report. AB - A 20-year-old-female first presented with an epithelioid sarcoma of the right thumb, and the right thumb was amputated. Five years later, a metastasis was found in the right lower lung and a partial lobectomy was performed. Three years later, computed tomography showed a metastatic brain tumor in the left frontal lobe, which was removed surgically. Adjuvant radiotherapy and chemotherapy were given after all operations. Histological examination showed all resected tumors were epithelioid sarcoma. She has maintained a good activity of daily living level as an outpatient for 2 years, although subcutaneous metastases and bronchial lymph node metastases have been observed. Such intensive treatment of slowly growing tumors often prolongs survival time, even in patients with multiple metastases. PMID- 11109798 TI - Preoperative embolization of upper cervical cord hemangioblastoma concomitant with venous congestion--case report. AB - A 16-year-old male presented with a large, solid hemangioblastoma located in the upper cervical cord manifesting as hyperactive reflexes, subtle weakness, and diminished position sense in all extremities. Neuroimaging studies indicated venous congestion due to arteriovenous shunt through the tumor. Preoperative embolization was accomplished without morbidity, and resulted in marked devascularization of the tumor and elimination of an early filling vein. Four days after embolization, the tumor was totally excised without excessive intraoperative bleeding. His neurological deficits gradually improved after surgery. Preoperative embolization is a valuable adjunct to surgical excision of large intramedullary hemangioblastomas, especially those associated with arteriovenous shunt, as cord dysfunction related to venous congestion and the risk of torrential intraoperative bleeding are reduced. PMID- 11109799 TI - Migration of the shunt tube after lumboperitoneal shunt--two case reports. AB - A 60-year-old male and a 36-year-old female suffered shunt migration after lumboperitoneal shunt procedures, upward into the spinal subarachnoid space and downward into the abdominal cavity, respectively. Defects of the fixation devices in the shunt system are considered the main cause in both cases. Upward migration of the lumbar tube in the subarachnoid space is extremely rare. We suppose that raised abdominal pressure is related to this unusual complication. PMID- 11109800 TI - Prevention of cerebrospinal fluid leakage and delayed loss of preserved hearing after vestibular schwannoma removal: reconstruction of the internal auditory canal in the suboccipital transmeatal approach--technical note. AB - The suboccipital transmeatal approach uses packing of a muscle or fat graft into the internal auditory canal (IAC) to prevent postoperative cerebrospinal fluid (CSF) leakage. However, preserved hearing after removal of vestibular schwannomas may decline over time because of the progressive constriction of cochlear vascular supply due to scarring of the IAC. We propose a surgical technique for IAC reconstruction, which separates the preserved cochlear nerve and vasculature from the graft, and regains the CSF space in the IAC. Prior to the drilling of the posterior wall of the IAC, the dura mater of the petrous bone forming the posterior wall of the IAC is harvested for IAC reconstruction. After completion of tumor removal, a "roof" of the IAC is reconstructed using the dura mater, and a muscle or fat graft soaked with fibrin glue is placed on the "roof" of the IAC. The IAC was reconstructed using this technique in 26 consecutive patients with vestibular schwannomas who underwent tumor removal via the suboccipital transmeatal approach. Postoperative magnetic resonance imaging confirmed the regained CSF space in the IAC. No delayed hearing loss occurred in four patients with preserved hearing. No CSF leakage occurred after surgery. This new technique of IAC reconstruction may prevent delayed hearing loss as well as postoperative CSF leakage after removal of vestibular schwannomas via the suboccipital transmeatal approach. PMID- 11109801 TI - [Evaluation of CYFRA 21-1 and ProGRP in serum and bronchoalveolar lavage fluid of patients with benign lung disease]. AB - CYFRA 21-1 and ProGRP have recently been established as new tumor markers for lung cancer. However, there are few reports evaluating concentrations in their bronchoalveolar lavage (BAL) fluid. In this study, we examined 81 patients with benign lung disease. The mean values of CYFRA 21-1 in the BAL fluid of each lung disease were as follows: bronchiolitis obliterans organizing pneumonia (BOOP), 3.9 +/- 2.1 ng/ml (positive rate 50%); collagen vascular disease associated interstitial pneumonia (CVD-IP), 10.7 +/- 15.7 ng/ml (positive rate 50%); diffuse panbronchiolitis (DPB), 4.2 +/- 6.4 ng/ml (positive rate 29%); idiopathic pulmonary fibrosis (IPF), 1.5 +/- 2.1 ng/ml (positive rate 17%); pulmonary infiltration with eosinophilia, 6.3 +/- 7.1 ng/ml (positive rate 44%); sarcoidosis, 4.6 +/- 6.2 ng/ml (positive rate 27%); and healthy volunteer (HV), 0.6 +/- 0.6 ng/ml; and total, 4.4 +/- 5.6 ng/ml (positive rate 32%). The mean values of ProGRP in the BAL fluid were as follows: DPB, 5.0 +/- 7.6 pg/ml (positive rate 0%); IPF, 6.4 +/- 10.6 pg/ml (positive rate 0%); HV, 12.4 +/- 8.3 pg/ml; and total, 5.6 +/- 8.7 pg/ml (positive rate 0%). These results indicate that the two tumor markers have no disease specificity in benign lung disease. PMID- 11109802 TI - [Low body mass index and exercise capacity in patients with chronic obstructive pulmonary disease (COPD)]. AB - This study examined retrospectively the relationships between body weight and exercise capacity in patients with chronic obstructive pulmonary disease (COPD). Seventeen patients with a %FEV1 less than 55% (mean +/- SD 36% +/- 8.8%) and minimum body weights of the body mass index (BMI) less than 20 (17.3 +/- 1.7) performed incremental exercise testing using a treadmill. Seventeen %FEV1-matched control patients with normal body weights were selected. There were no significant differences in the patients' characteristics or their pulmonary function tests (including vital capacity, carbon monoxide diffusing capacity, and arterial blood gases). Low BMI patients Is this the weaning of (67.8 +/- 6.3 years old) were younger than the control patients (73.1 +/- 8.5 years old), but the difference was not statistically significant. The exercise capacities of low BMI patients were significantly superior to those of the control patients (316.5 +/- 171.5 seconds vs 204.1 +/- 116.3 seconds, p = 0.038) and total walking distance without statistical significance (194.9 +/- 117.0 m vs 125.7 +/- 98.0 m, p = 0.071). Also, low BMI patients achieved higher maximal minute ventilation volume during exercise than the controls. The major factor limiting exercise in patients with low BMI was ventilation. Moderately low body weight may not be a risk factor in Japanese COPD patients. PMID- 11109803 TI - [A clinicopathological study of pulmonary cryptococcosis--chest CT and pathologic correlation]. AB - We reviewed the clinicopathological features in 12 patients (7 males and 5 females; mean age 54 yr) with pulmonary cryptococcosis. Eleven of the patients were asymptomatic and the disease was detected by chest radiograph abnormalities. The underlying systemic disease had been diagnosed as diabetes mellitus in two. Chest CT scans showed a solitary nodule in 9 of the 12 patients, multiple nodules in 2, and infiltration in 1. The nodular diameter was less than 2 cm in 10 of the 12. All nodules were located in the subpleural region. On the chest CT, cavitary nodules, scattered nodules, or both, and spiculated nodules were difficult to distinguish from pulmonary tuberculosis and primary lung cancer, respectively. According to McDonnell's pathological classification of pulmonary cryptococcosis, the resected 8 lungs revealed peripheral pulmonary granuloma in 5 and granulomatous pneumonia in 3. It is important to perform a pathological examination for the diagnosis of pulmonary cryptococcosis to avoid misdiagnosis as lung cancer or pulmonary tuberculosis. PMID- 11109804 TI - [Evaluation of fluorine-18-fluorodeoxyglucose whole body positron emission tomography imaging in the clinical diagnosis of lung cancer]. AB - In recent years the use of positron emission tomography (PET) with fluorine-18 fluorodeoxyglucose (FDG) has become a valuable tool in the detection of a variety of tumors including lung cancer. To determine its role in the diagnosis of patients with suspected lung cancer, we compared the results of FDG-PET with those of the other scintigraphic imaging techniques (67Ga-planar image, 201Tl SPECT and 99mTc-bone scintigraphy) used worldwide in patients with lung cancer. The analysis group consists of 178 patients, 159 malignant pulmonary diseases and 19 benign pulmonary diseases. FDG-PET was performed in 65 patients (51 malignant pulmonary diseases, 14 benign pulmonary diseases). FDG-PET had a sensitivity, specificity and accuracy of 98.0%, 78.6% and 93.8%, respectively, in detecting malignant pulmonary nodules. In N staging, sensitivity, specificity and accuracy were 66.7%, 81.3% and 76.0%, respectively. In M staging, the accuracy was 100%. Thus, FDG-PET imaging was more accurate than the other types of scintigraphic imaging. In our observations, whole-body 18FDG-PET images improved diagnostic accuracy in the evaluation of lung lesions and the staging of lung cancer. PMID- 11109805 TI - [PL granule-induced pneumonia requiring mechanical ventilation]. AB - PL granule is one of the most common forms of composite medicine for colds in Japan, including acetaminophen. We report a case of PL granule-induced pneumonia that required mechanical ventilation. A 72-year old man who had been prescribed amiodarone for more than one year before for arrhythmogenic right ventricular dystrophy, repeatedly took PL granules at one-week intervals because of sneezing and fever. He then underwent acute respiratory failure. He needed mechanical ventilation for severe hypoxemia, but recovered with glucocorticoid pulse therapy. Because the blastoid transformation test using his peripheral blood lymphocytes was positive on stimulation with PL granules, but negative on amiodarone stimulation, he was diagnosed as having drug (PL granule)-induced pneumonia. Only one case of PL granule-induced pneumonia and seven of acetaminophen-induced pneumonia have been reported. None of these patients needed mechanical ventilation. PL granule-induced pneumonia should be treated cautiously because of possible acute exacerbation. PMID- 11109806 TI - [A case of interferon beta-induced pneumonia]. AB - A 58-year-old man had been treated with one intravenous injection of 120 mg of nimustine hydrochloride (ACNU), ten thrice-weekly doses of 3,000,000 U of interferon beta, and brain irradiation for cerebral glioblastoma. One month later he had fever, appetite loss, a productive cough and dyspnea. Chest radiography and CT showed diffuse, nonsegmental ground glass opacity in both lung fields. Hypoxemia and lung shadows were exacerbated day by day. Bronchoalveolar lavage revealed an increases in the total cell count and the percentages of lymphocytes and neutrophils, and a decrease of the CD 4/8 ratio. Interferon beta therapy was stopped, and steroid pulse therapy and prednisolone 40 mg administration were initiated. The symptoms, hypoxemia and lung shadows quickly improved. Reported cases of interferon beta-induced pneumonia are rare. PMID- 11109807 TI - [Rapid development of multiple meniscal signs complicating invasive pulmonary aspergillosis in a patient receiving chemotherapy for acute leukemia]. AB - This report describes the rapid development of multiple meniscal signs complicating invasive pulmonary aspergillosis in a 53-year-old man receiving chemotherapy for acute leukemia. While undergoing first induction therapy for AML, he developed chest pain, and multiple bilateral infiltrations were seen in chest roentgenograms. Administration of antibiotics, antifungal agents, steroid pulse therapy and G-CSF was begun. Pulmonary cavities with meniscal signs developed. The next day, pneumothorax and hemothorax were noted. Although drainage and mechanical ventilation were performed, the patient died after massive hemoptysis. Invasive pulmonary aspergillosis was diagnosed at autopsy. PMID- 11109808 TI - [Three cases of Mycobacterium avium lung disease in an iron foundry]. AB - We report three cases of M. avium lung disease, all occurring in the same iron foundry over a 5-year period. Case 1: A 38-year-old man was admitted to our hospital because of abnormal chest x-ray shadows observed during a routine checkup. M. avium complex was isolated from the sputum and a CT scan showed multiple nodular shadows with cavities. Case 2: A 63-year-old man presented with dyspnea. Chest CT showed nodular shadows in both upper lobes. M. avium complex was isolated from his sputum. Case 3: A 62-year-old man was hospitalized for treatment of diabetes mellitus. A nodular cavitary shadow* OK?* in the right upper lobe was observed in the radiograph. CT scanning demonstrated a nodular shadow with thick pleural indentation. M. avium was isolated from the sputum and gastric juices. There were similarities in the radiographic findings in all cases: nodular shadows with cavities in the upper lung fields. These findings differed from those in another M. avium infection in the lung which occurred in a middle-aged woman. We suspected inhalation of an aerosol contaminated by M. avium complex. There have hitherto been no reports of group infections in healthy persons. We suspected the same environmental source for the infection. PMID- 11109809 TI - [A case report of pulmonary nocardiosis successfully treated with a combination of sulfamethoxazole-trimethoprim (ST) and sparfloxacin]. AB - We encountered a case of pulmonary nocardiosis that responded dramatically to combined ST and sparfloxacin treatment. A 55-year-old woman presented with fever, cough and yellowish sputum. She had been under treatment with oral prednisolone (15 mg per day) since July 1997 after a diagnosis of Evans syndrome. A high fever of 39.8 degrees C was noted on January 30, 1998. The patient was hospitalized for bloody sputum, bilateral hypochondriac pain and evidence of infiltrative opacities in the left lower lobe on chest radiography. Bacterial pneumonia was suspected, and she was treated with piperacillin, but her clinical symptoms did not improve. Sputum culture and serologic examination failed to lead to a definitive diagnosis. Nocardia farcinica was isolated by culturing tissue obtained by CT-guided transcutaneous pulmonary biopsy, leading to a diagnosis of pulmonary nocardiosis. The results of an MIC test for antimicrobial agents led to treatment with a combination of ST and sparfloxacin, and the clinical symptoms improved. These clinical observations suggest that, when pneumonia is diagnosed in patients who have been receiving oral steroids for a prolonged period, pulmonary nocardiosis should be considered in the differential diagnosis to enable selection of appropriate antimicrobial agents. PMID- 11109810 TI - [Development of a case of Mycobacterium avium complex disease from right pleural effusion]. AB - Moist pleurisy in patients with Mycobacterium avium Complex (MAC) is rarer than tuberculosis. We encountered an extremely rare case of MAC disease in a 75-year old man who initially had only right pleural effusion. Gaffky VII was detected in the pleural effusion, and Mycobacterium avium was identified by culture and PCR. Although administration of antitubercular agents (RFP, INH, EB, and SM) + CAM and thoracic lavage were repeated, the Gaffky persisted strongly. Accordingly, pulmonary decortication and filling of the cavity with an omental flap were performed as surgical treatments. However, fistulas were formed between the remaining empyema cavity and the surgical wounds. Fenestration was also carried out. Postoperatively, centriacinar abnormalities appeared on computed tomography (CT). It has been reported that MAC disease begins with centriacinar abnormalities and the incidence of the lymphatic developmental pattern was low. Tuberculosis (the idiopathic pleuritis type) is considered to be caused this pattern from the primary infection focus. Therefore, the onset of unilateral effusion is extremely rare in patient with MAC disease, suggesting that the lymphatic developmental pattern occurs less frequently in patients with MAC disease. Furthermore, in this case, we speculated that centriacinar abnormalities were the MAC infection foci and could be detected by CT due to surgical invasion. PMID- 11109811 TI - [Pulmonary suppuration due to a mixed infection of anaerobic bacterium and Actinomyces]. AB - We report a case of pulmonary suppuration due to a mixed infection of anaerobic bacterium and Actimomyces. A 49-year-old man was admitted to our hospital because of hemoptysis on March 30, 1999. A chest X-ray film showed a localized shadow in the right middle lobe, and a tumor shadow was recognized on chest CT. The anaerobic bacterium were isolated from specimens collected bronchofiberscopically. A diagnosis of pulmonary suppuration due to anaerobic bacterium was made, and treatment with sulbactam/ampicillin, followed by imipenem/cilastatin, was initiated. Although his clinical symptoms and laboratory data improved rapidly following this treatment, the abnormal finding on the chest radiographs remained, with only slight improvement. Accordingly, surgical resection of part of the right middle lobe was performed on June 29 using a video associated thoracic surgery technique. Actinomycosis was recognized by pathological examination of the resected lesion. A revised diagnosis of pulmonary suppuration due to a mixed infection of anaerobic bacterium and Actinomyces was made. Anaerobic bacterium in the oral cavity are recognized as significant pathogens in pulmonary suppuration. In the present case, we considered anaerobic bacterium and Actinomyces aspirated from the oral cavity into the lung to have caused the pulmonary suppuration. PMID- 11109812 TI - [Three cases of primary pulmonary malignant lymphoma]. AB - Primary pulmonary malignant lymphoma is a rare disease that is thought to belong to a category of malignant lymphomas arising from mucosa- or bronchus-associated lymphoid tissue (MALT or BALT). We encountered 3 cases of primary pulmonary malignant lymphoma, Case 1: In a 51-year-old male, an abnormal shadow was detected in chest radiography in the right S9 after an operation for thyroid carcinoma. A right lower lobectomy was performed. The diagnosis was malignant lymphoma (marginal zone B-cell lymphoma). Immunohistochemical staining for IgM gave a positive result. Case 2: Multiple nodular shadows were noted in both lungs of a 55-year-old man after a bout of pneumonia. Video-assisted thoracoscopic surgery was performed, and the diagnosis was malignant lymphoma (marginal zone B cell lymphoma). Gene analysis revealed rearrangement of a heavy chain gene. Case 3: An abnormal shadow was seen in the chest radiograph of a 60-year-old man. He was treated by right upper and middle lobectomy. The diagnosis was Hodgkin's disease, nodular sclerosing type. Chemotherapy was given after surgery and the patient is now alive without recurrence. As the pulmonary malignant lymphoma was difficult to diagnosepreoperatively, it was necessary to resect the mass for diagnostic purposes. The prognosis of a resected solitary lesion in the lobe was good. Therefore lobectomy was performed as the treatment of choice. Systemic chemotherapy is performed for the diffuse type of pulmonary lymphoma. PMID- 11109813 TI - [A case of resected squamous cell carcinoma of the lung complicated with sarcoidosis]. AB - A 64-year-old man with uveitis was admitted to our hospital for detailed investigation of an abnormal shadow on his chest X-ray. Chest radiography and computed tomography of the chest showed mediastinal lymphadenopathy and a tumor shadow in the left hilum. Transbronchial tumor biopsy revealed squamous cell carcinoma. Left upper lobectomy and drainage of bilateral hilar and mediastinal lymph nodes were performed. Histopathological examination revealed the coexistence of squamous cell carcinoma with many non-caseating epithelioid cell granulomas in all hilar and mediastinal drainage lymph nodes, but no metastasis. Non-caseating epithelioid cell granulomas were also seen in the interstitium and alveolar spaces. Coexistence of sarcoidosis and lung cancer in the same patient is not common, and only 29 cases, including ours, have been reported. This case also provides the concept that surgical tumor resection should be considered even if bilateral mediastinal lymphadenopathy is found in a case of lung cancer complicated with sarcoidosis. PMID- 11109814 TI - [A case of intrathoracic schwannoma derived from the left vagus nerve]. AB - We report a case of schwannoma asymptomatic for 9 years, derived from the left vagus nerve in the middle mediastinum. This spindle-shaped tumor caused paralysis of the left recurrent nerve as an initial clinical manifestation with cough. T2 weighted magnetic resonance imaging (MRI) showed a neurogenic tumor with a characteristic target appearance and with constituents of different intensities: mucinous material in the peripheral zone and solitary tissue in the central zone. But, this different intensity is not directly reflected by the histopathologic features of Antoni types A and B. These findings suggest that MRI is useful for determining the parent nerve of a neurogenic tumor and is helpful for the diagnosis. PMID- 11109815 TI - [The study on the effect of the transurethral catheter in pressure-flow study- the comparison with the urination in uroflowmetry]. AB - OBJECTIVE: Pressure-flow study (PFS) is an excellent method for detecting the bladder outlet obstruction. However, PFS with the transurethral catheter has the disadvantage of unphysiological urination during testing. In the present study, we compared the urination in PFS with that in uroflowmetry (UFM) and analyzed the effect of the transurethral catheter. MATERIALS AND METHODS: The subjects were 43 men who underwent PFS at Yamato Takada Municipal Hospital. PFS and UFM were compared with respect to four parameters, i.e., Qmax, the residual urine volume, the region on Liverpool nomogram, and the flow curve pattern (Jorgensen's classification). RESULTS: For PFS using a transurethral catheter, (1) Qmax was decreased by a mean of 2.6 ml/s and the residual urine volume was increased by a mean of 26.3 ml compared with UFM, and (2) the Liverpool nomogram and the flow curve pattern showed more impairment of urination with PFS than UFM. CONCLUSION: It is suggested that PFS using a transurethral catheter may not reflect physiological urination. PMID- 11109816 TI - [The brachytherapy with low dose-rate iridium for prostate cancer]. AB - PURPOSE: Brachytherapy as an option for the treatment of prostate cancer has been commonly performed in USA. As the permanent seeding of the radioactive materials is strictly restricted by the law in Japan, brachytherapy must be performed by the temporary implant. This treatment has been performed at a few facilities in Japan mostly using high dose-rate iridium. Only our facility has been using low dose-rate iridium (LDR-Ir) for prostate cancer. This study evaluates the clinical results of the treatment. PATIENTS AND METHODS: Since December 1997 to December 1999, 26 patients with histologically diagnosed as prostate cancer (Stage B, 92%; Stage C, 8%) underwent brachytherapy. Twenty-two patients received brachytherapy alone, three were treated with a combination of brachytherapy and external beam radiotherapy (ERT) and one was treated with a combination of brachytherapy and neoadjuvant endocrine therapy. Patients ranged in age from 61 to 84 (median 76) years old. Treatment was initiated with perineal needle placement. From 10 to 14 needles were placed through the holes on the template which was fixed to the stabilizer of the transrectal ultrasound probe. After the needle placement. CT scan was performed to draw distribution curves for the treatment planning. LDR-Ir wires were introduced to the sheath and indwelled during the time calculated from dosimetry. Peripheral dose was 70 Gy for the monotherapy of brachytherapy. For the combination therapy, 40 Gy was given by brachytherapy and 36 Gy with ERT afterwards. LDR-Ir wires were removed after completion of the radiation and patients were followed with serum PSA level and annual biopsy. RESULTS: During 2 to 26 (median 12) months follow-up, 8 out of 9 patients with initial PSA level above 20 ng/ml showed PSA failure. All 13 patients with initial PSA level lower than 20 ng/ml were free from PSA failure. Eight out of 11 patients with Gleason's score 7 or higher showed PSA failure, and all 14 patients (including three patients with combined therapy) with Gleason's score 6 or less were free from PSA failure. Annual biopsy was performed in 8 patients, and 4 patients histologically revealed tumor free and 4 patients (two of them showed PSA failure) were tumor positive. No major complication was observed, however, some minor side-effect as irritability was seen in 65% of the patients. CONCLUSIONS: The results showed that brachytherapy with LDR-Ir was an acceptable treatment as long as the patients were selected strictly with PSA level and Gleason's score. A good candidate for this treatment is the patient whose PSA level is lower than 20 ng/ml and Gleason's score is 6 or less. The treatment is effective and safe, but further observation is necessary to reach the conclusion. PMID- 11109817 TI - [Treatment of metastatic seminoma by chemotherapy.: an experience]. AB - PURPOSE: To describe the outcome of chemotherapy using cisplatin-based regimen, and experimental combination with carboplatin and ifosfamide to treat advanced seminoma. METHODS: From 1981 to Jan. 1999, 15 patients with Stage IIA, IIB, IIIA or IIIC metastatic seminoma and one patient with lung disease, who suffered a relapse of his primary mediastinal lesion were treated. Three of these patients had relapsed, following surveillance for Stage I testicular cancer, and another had received prophylactic radiotherapy to the retroperitoneal lymph nodes in advance. The first patient's regimen consisted of cisplatin and cyclophosphamide. Since 1983, cases have been treated with the same regimen as that used to treat non-seminomatous germ-cell tumors; cisplatin/vinblastine/bleomycin (PVB); cisplatin/vinblastine/actinomycin D/cyclophosphamide/bleomycin (VAB-6); cisplatin/etoposide/bleomycin (BEP). From 1993, six patients with non-bulky metastatic seminoma participated in a trial involving 3 courses of carboplatin (400 mg/m2) and ifosfamide (2,000 mg/m2, 3 days). RESULTS: Of the entire group, 10 patients (62.5%) achieved a CR after chemotherapy alone. Four cases who received radiation, following chemotherapy, produced CR. Surgical resection of residual tumors were performed on 2 patients. Resected tumors were fibrous and no evidence of malignancy. All those individuals who participated in this study, are alive and disease-free today, from 11 months to 18 years. Carboplatin and ifosfamide demonstrated only mild toxicity, during a 4-week cycle, with subjects being treated on an outpatient basis. CONCLUSION: As expected, the type of chemotherapy we used, to treat non-seminomatous germ-cell tumors proved to be highly effective for seminomatous types, as well. Carboplatin and ifosfamide performed well and safe, in the treatment of non-bulky metastatic seminoma. Comparative studies of long-term treatment results and QOL, using either radiotherapy or low-toxicity chemotherapy for Stage IIA disease should be undertaken. PMID- 11109818 TI - [Clinicopathological study of the testicular microlithiasis]. AB - PURPOSE: Testicular microlithiasis (TM) is a relatively rare condition characterized by calcific concref1p4 within the seminiferous tubules. Little has been reported on the incidence or the clinical implication of TM among Japanese. To address the problem, we evaluated pathologic specimens from biopsies and orchiectomies, of testes with various conditions. MATERIALS AND METHODS: Pathologic specimens of the testes of 200 cases, 56 from orchiectomy and 144 from testicular biopsy, were investigated. RESULTS: The pathological diagnosis of TM was confirmed in seven (3.5%) cases, four of which were associated with germ cell tumors and the other three were obtained from testicular biopsies performed for examination of infertile men. Of the 41 patients with germ cell tumors, four (9.8%) were found to have TM, and another three (2.5%) were identified among 122 patients with infertility. The prevalence of TM is significantly higher in specimen with germ cell tumors than those without germ cell tumors (p < 0.05). CONCLUSIONS: Although TM is rarely encountered, this condition is relatively often accompanied by testicular malignancy. Further investigation would be fundamental to ascertain the relationship between TM and testicular malignancy. PMID- 11109819 TI - [Granulocyte colony-stimulating factor producing undifferentiated carcinoma of urinary bladder: a case report]. AB - A case of bladder cancer producing granulocyte colony-stimulating factor (G-CSF) is reported. A 94-year-old woman with a progressive, grade 3 undifferentiated carcinoma, showed marked leukocytosis (maximum 29,780/mm3) with an elevated G-CSF (420 pg/ml). Immunohistochemical examination with monoclonal antibody specific for G-CSF revealed positive staining. Further examination for epidermal growth factor receptor (EGF-R) and p53 were both positive and Ki67 index were 40.7%. These data suggested that this tumor had extremely aggressive growing nature, as the biological character is this. PMID- 11109820 TI - [A case of Churg-Strauss syndrome which cheaf complain was gross hematuria]. AB - We report a case of Churg-Strauss syndrome complaining of gross hematuria. A 74 year-old man was admitted to our hospital for further examination of gross hematuria. Abdominal CT and retrograde pyelography revealed left renal pelvic tumor, and left nephrectomy was performed. There was no tumor, but submucous hemorrhage was seen in the renal pelvis. The histopathologic diagnosis was allergic granulomatous angitis. The administration of prednisolone was done, but he suddenly died of acute heart failure after 1 month postoperatively. PMID- 11109821 TI - [Secondary leukemia following ultra high-dose chemotherapy with peripheral blood stem cell autotransplantation for refractory testicular cancer]. AB - Secondary leukemia following chemotherapy or radiotherapy for mediastinal germ cell tumors in a well-described entity. It also may occur in patients with testicular germ cell tumors. We report a case of secondary leukemia occurring in a 31-year-old man who received ultra high-dose chemotherapy with peripheral blood stem cell autotransplantation (PBSCT) for a refractory testicular cancer (pathology; Seminoma, Embryonal carcinoma, Yolk sac tumor, Choriocarcinoma) with IIIB2 under Japanese classification, poor-risk group under Indiana classification. The initial levels of serum LDH, AFP and beta-HCG were high at 959 IU/l, 1,452 ng/ml and 800 ng/ml. He received total 11 cycles of systemic chemotherapy (2 cycles of PVB regimen, 4 cycles of PEB regimen, 3 cycles of VIP regimen and 2 cycles of ultra high-dose chemotherapy with PBSCT for pulmonary and para-aortic lymph node metastasis following his initial orchiectomy. The total amount of etoposide (VP-16), cisplatin (CDDP), carboplatin (CBDCA) and ifosfamide (IFM), this patient received was 7,225 mg/m2, 1,510 mg/m2 1,750 mg/m2, and 50.5 g. He has survived with CR of disease. Severe and persistent pancytopenia developed 25 months after his initial orchiectomy. Bone marrow examination showed AML (M2 with eosinophilia) under French-America-British (FAB) classification. Therefore, he was diagnosed as secondary leukemia following high-dose chemotherapy. He received total 6 cycles of systematic chemotherapy for the secondary leukemia in the internal department. He is planing to have bone marrow transplantation. To our knowledge, this is the first reported case in the literature relevant to secondary leukemia following ultra high-dose chemotherapy with PBSCT in testicular tumor in Japan. PMID- 11109822 TI - [Influence of spontaneous otoacoustic emission (SOAE) on transiently evoked otoacoustic emission (TEOAE)]. AB - The literature has reports on the influence of spontaneous otoacoustic emission (SOAE) on transiently evoked otoacoustic emission (TEOAE), but most do not take factors such as age, gender, and hearing level into consideration. We focused on these conditions. Subjects were 78 women with normal hearing aged 19 to 24 years (mean = 21.4). All had pure tone thresholds of 15 dB HL or better at 1 kHz, 2 kHz, and 4 kHz. ILO88 was used to record TEOAE and SOAE. Echo power (EP) and reproducibility (Repro) were compared between groups with and without SOAE. No significant audiometric difference was seen between groups. Total echo power (TEP) and whole reproducibility (WR) were significantly greater in the group having SOAE, consistent with previous reports (p < 0.01). EP and Repro classified by frequency bands were also significantly greater in the group having SOAE at 1 kHz to 4 kHz. Subjects were divided based on the number of SOAE and the above parameters compared. We found that as the SOAE number increased, EP and Repro increased. In conclusion, the existence of SOAE influences TEOAE parameters and must be taken into account during clinical testing. PMID- 11109823 TI - [Histopathological study of the temporal bones in patients with primary carcinomas of the ear]. AB - The most common symptoms of patients with carcinomas of the middle ear or mastoid are otorrhea, facial paralysis, and hearing loss, including a sensorineural element and vertigo. The latter two symptoms are indicators of inner ear damage. However, few reports have been made concerning the histopathological changes that occur in the inner ear in the presence of a tumor. The present study was performed to determine the pattern of tumor invasion in the inner ear and the histopathological changes that occur in the inner ear in cases of ear carcinomas. Temporal bone sections from five patients (age: #39-73 years; 3 males and 2 females) who died from a primary carcinoma of the ear were studied histologically. The following features were examined: 1) localization of the tumor in the temporal bone, 2) pattern of tumor invasion in the inner ear, 3) pathological changes in the inner ear, including the cochlea, vestibule and semicircular canals. Tumor cells were still present in the temporal bone sections of all the patients except one, even though the patients had received various treatments for the carcinoma, including radiation therapy, surgery and chemotherapy. Marked inflammatory and necrotic changes were observed in cases where the tumor had invaded the external auditory canal, middle ear cleft, internal auditory canal, and in some cases the inner ear. In cases where the tumor invaded the inner ear via the internal auditory canal rather than directly from the middle ear, the otic capsule is thought to have acted as a barrier against tumor invasion. In addition, marked degenerative changes throughout the entire inner ear structures were noted. These changes may have arisen from an attenuated blood supply to the inner ear as a result of pressure from the tumor in the internal auditory canal, tumor infiltration of the labyrinthine artery. PMID- 11109824 TI - [Distribution of intraglandular metastatic foci in the contralateral lobe of papillary thyroid carcinoma]. AB - The distribution of metastatic foci in the opposite lobe has not been studied in detail despite of several reports on the high incidence of contralateral metastasis. Whether foci spread to the upper one-third of the contralateral lobe influences the choice of total or subtotal thyroidectomy. Metastasis was studied in 66 patients 11 men and 55 women aged 24-73 years (mean; 51.3), undergoing primary total thyroidectomy from 1988 to 1996. Serial blocks of the opposite lobe, approximately 5 mm thick were sliced and stained by hematoxylin-eosin. Metastases were found in 44 patients (67%). Based on the size of the primary focus, these patients were divided into group A (smaller than one-third of the lobe) and group B (greater than that of group A). The average size of the primary focus was 21 mm in group A and 36 mm in group B. The contralateral metastatic rate was 64% (14/22) in group A and 68% (30/44) in group B. The distribution of metastatic foci in the opposite lobe was studied in 44 positive patients. The spread to the upper one third occurred in 61% (27/44); 29% (4/14) in group A and 76% (23/30) in group B a significant difference (p < .003). We thus concluded that the larger the primary focus, the wider the spread of metastatic foci to the opposite lobe. PMID- 11109825 TI - [Globus pharyngeus and bone mineral density]. AB - While globus pharyngeus is a common disorder, the cause of this anomaly remains unclear. Gastroesophageal reflux, cricopharyngeal spasm, and many other etiologies have been considered as possible causes. Some researchers believe that the disorder is probably multifactorial in origin. Patients with globus pharyngeus are usually female, and the majority of them are menopausal. These middle aged females often have a reduced bone mineral density. This study was undertaken to determine whether males and females with globus pharyngeus have a reduced bone mineral density. We studied 12 men and 17 women with globus pharyngeus who came to Tokai University, Isehara Kyodo Hospital and the Hagino E.N.T. Clinic between February 1992 and February 1994 and compared them to a control group of 12 males and 15 females. Each patient met the criteria for 'primary globus pharyngeus' (Bradley 1987) as determined by endoscopy and none of the patients showed any signs of inflammation, tumors, or gastroesophageal reflux. The second midcarpal bone mineral density of each subject was measured using a computed X-ray densitometer and analyzed using microdensitometry. Compared with control group, patients with globus pharyngeus had a lower bone mineral density in their second midcarpal bone. The sigma GS/D was significantly lower in the globus pharyngeus patients than in the control group (p < 0.01) and significantly lower in the female patients than in the female control group (p < 0.01). The GSmax was significantly lower in the patients than in the control group (p < 0.01) and significantly lower in the female patients than in the female control group (p < 0.01). The GSmin was significantly lower in the patients than in the control group (p < 0.01), significantly lower in the female patients than in the female control group (p < 0.01), and significantly lower in the male patients than in the male control group (p < 0.05). In conclusion, globus pharyngeus appears to be related to a decrease in bone mineral density. PMID- 11109826 TI - [Reliability of electrogustatory threshold--electrogustometry using CNV]. AB - CNV (contingent negative variation), an event-related potential, is induced when an oncoming imperative stimulus (S2) is anticipated after recognition of a warning stimulus (S1). With this in mind, we assessed the usefulness of electrogustometry using CNV as an objective gustatory test with an electric gustatory stimulus as a warning stimulus, S1. A gustatory test was performed on a total of 100 sides of the tongue in 50 subjects. Excluding no response cases on 5 sides in 5 subjects, the CNV threshold was successfully determined on 95 sides in 50 subjects. When the CNV threshold values obtained and the conventionally measured subjective threshold values were compared, a very high correlation was demonstrated between the two thresholds, with a correlation coefficient of 0.961. To assess the reliability of the CNV threshold, three additional measurements on 5 sides in 5 subjects with normal subjective threshold values were performed on different days, and the CNV threshold value variation among the three measurements in all 5 subjects never exceeded 13 microA. Separately, subjective threshold and CNV threshold values were compared in 4 patients with facial palsy (hence with gustatory abnormality) who consented to a follow-up examination. The difference between the subjective threshold and CNV threshold values in all 4 patients never exceeded 10 microA. These findings demonstrate that the reliability of the CNV threshold is satisfactory. Next, a gustatory imitation experiment was performed to assess the usefulness of electrogustometry using CNV as a means of detecting gustatory abnormality malingering. The experiment was performed on a total of 6 subjects: 3 physicians with knowledge of CNV (knowledge group) and 3 others with no knowledge of CNV (no-knowledge group). The differences between the imitated threshold and the subjective threshold values in all of these subjects never exceeded 10 microA, and the difference between imitated threshold and CNV threshold values never exceeded 13 microA. Comparison between the knowledge group and the no-knowledge groups showed that the imitated threshold values were very close to the subjective threshold and CNV threshold values. Based on the above results, it was concluded that gustometry using CNV is useful and can serve well as a method of objective evaluation of gustatory sensation. The results also suggested that it will be useful in diagnosing malingering. PMID- 11109827 TI - [Resistance to Fas-mediated apoptosis in head and neck squamous cell carcinoma cells]. AB - Various types of carcinoma cells have been shown to escape immune recognition by constitutive resistance to Fas receptor (Fas)-mediated apoptosis. The purpose of this study was to examine the sensitivity to Fas-mediated apoptosis of human head and neck squamous cell carcinoma (HNSCC) cells. We applied an anti-Fas monoclonal antibody (CH11) to HNSCC cell lines and monitored their cell death. All three HNSCC cell lines examined expressed Fas protein but not Fas-ligand mRNA. CH11 did not induce cell death (CH11-resistant) in any of the three HNSCC cell lines examined. Treatment with actinomycin D (ActD) converted the phenotypes of the CH11-resistant HNSCC cell lines from CH11-resistant to CH11-sensitive, suggesting that the resistance to CH11-induced apoptosis was dependent on RNA synthesis. Western blot analysis did not show any differences in expression of Bcl-2 between the non-treated and ActD-treated HNSCC cell lines. Expression of Bcl-XL, on the other hand, was greatly reduced in the ActD-treated HNSCC cell lines, implying that Fas signaling in the CH11-resistant HNSCC cell lines might be regulated by an Bcl-XL-inhibitable step. PMID- 11109828 TI - [Effect of paclitaxel on cisplatin resistant head and neck squamous multicellular tumor spheroids]. AB - Paclitaxel has been reported to be effective for the treatment of CDDP resistant tumors. Thus, the efficacy of paclitaxel on CDDP resistant HEp-2 and KB head and neck squamous carcinoma cell lines was evaluated in monolayer and multicellular tumor spheroids (MTS). Cell lines with a tenfold resistance to CDDP were used in this study (Tanaka, K. et al, Keio J Med, 70. 1993). MTS were developed using the liquid overlay culture technique. After exposure to graded concentrations of drugs and different exposure time, the cells were subjected to a clonogenic assay. The effect of paclitaxel on both monolayer and MTS was dependent on the drug concentration and related to the exposure time. For HEp-2 MTS, 10(-7) M/L of paclitaxel resulted in a cell death rate of approximately 90% in both parent and resistant cells. For KB MTS, the cells were more resistant to paclitaxel than the HEp-2 cells, and a 72 hour exposure time was needed to achieve a cell death rate of approximately 90%. These data suggest that paclitaxel may be effective for treating CDDP resistant head and neck cancer. PMID- 11109829 TI - [Flexibility in the adaptation of the vestibulo-ocular reflex to modified visual inputs in humans]. AB - The vestibulo-ocular reflex (VOR) serves to stabilize images on the retina. To maintain appropriate performance and minimize image slippage throughout life, the VOR is subject to long-term adaptive regulation in response to visual input. Adaptive changes in VOR gain (eye velocity/head velocity) can be evoked either by fitting subjects with magnifying, miniaturizing, or reversing spectacles during normal behavior or by moving a large visual field in or out of phase relative to the subject's head movement. These changes exhibit frequency-selectivity. Here, we examine the flexibility of VOR gains by causing VOR in similar directions to undergo different behavioral gain changes. Nine healthy adults, ranging in age from 24 to 38 (mean 28.5) with no history of neurotological symptoms participated in the study. All subjects demonstrated clinically normal functioning on a screening battery of tests that included combined neurologic and otologic physical examinations. Horizontal and vertical eye positions were recorded by bitemporal DC coupled electrooculography (EOG). The subject sat in a rotating chair. The axis of rotation of the body was always earth-vertical, the interaural axis crossing the axis of rotation of the chair. The head was positioned at 20 degrees down in all experiments and was stabilized in this position using a chin rest. The chair was 78 cm in diameter and was shielded by a half-cylindrical optokinetic screen positioned in front of the subjects. Random dot patterns were projected onto this screen. During per- and post-adaptation periods, goggles were fitted to ensure that the subject was in complete darkness and the chair was rotated sinusoidally. The amplitude of the rotating chair was 30 degrees and 60 degrees. Frequencies of rotation were 0.1 Hz, 0.2 Hz, 0.3 Hz and 0.4 Hz for amplitudes of 30 degrees and 0.1 Hz, 0.2 Hz, and 0.3 Hz for amplitudes of 60 degrees. To induce VOR adaptation, the retinal slippage velocity caused by the visual input of a large field was changed for short-term; the change was produced by a combination of sinusoidal head rotation and random dot patterns. During each adaptation session, the frequencies of sinusoidal head rotation were either 0.1 Hz or 0.3 Hz and the amplitude was 30 degrees. The random dot patterns were synchronized with the sinusoidal head rotation in the same direction to make the retinal slippage zero (x0 experiment) and in the opposite direction to double the retinal slippage (x2 experiment). Therefore, a total of four adaptation protocols were tested. The subjects were asked to fix their eyes on a single dot by looking straight ahead in the x0 experiment and to follow the dot in the random dot pattern in the x2 experiment. Each adaptation session lasted for 30 minutes. Each subject participated in couple of adaptation experiments everyday. The average VOR gain and phase lag were calculated using a Fourier transformation. Out of all the subjects who participated in the x2 adaptation experiment at 0.3 Hz with an amplitude of 30 degrees, seven subjects showed a steady increase in VOR gain during a couple of the trials. One out of the remaining two subjects showed a decrease in VOR gain in all three trials. Another subject showed an increase in VOR gain during three trials and a decrease in two trials. In the x2 adaptation experiment with a range of 30 degrees at 0.3 Hz (peak velocity: 28 degrees/s), the percent change in gain (post-pre/pre) was 133% at the same stimulation and 100% at 0.4 Hz (peak velocity: 37 degrees/s). The percent change in gain was 65% for amplitudes of 60 degrees at 0.1 Hz (peak velocity: 18 degrees/s) and 64% for amplitudes 60 degrees at 0.2 Hz (peak velocity: 37 degrees/s). In the x1 adaptation experiment (30 degrees at 0.3 Hz), the percent change in gain was -62% for the same conditions and -50% for amplitude of 60 degrees at 0.1 Hz and -30% for amplitudes of 60 degrees at 0.2 Hz. No change, in VOR gain was observed at the other frequencies. The PMID- 11109830 TI - [Dysphagia: mechanism, treatment and rehabilitation]. PMID- 11109831 TI - [Hemodynamic stresses and blood vessel]. PMID- 11109832 TI - [Medical cost in the elderly]. PMID- 11109834 TI - [Aging--senescence--dementia]. PMID- 11109833 TI - [The current status and perspectives in Alzheimer's disease research]. PMID- 11109835 TI - [Medical decision making and clinical epidemiology]. PMID- 11109836 TI - [Visions and perspectives for the nursing care in the 21st century--supporting our aged society]. PMID- 11109837 TI - [To spread and make use of guidelines for hypertension in the elderly--1999 revised version]. PMID- 11109838 TI - [Hematopoietic stem cell transplantation and aging]. PMID- 11109839 TI - [Effect of blood pressure control on diabetic nephropathy]. PMID- 11109840 TI - [Self-determination of the elderly at the terminal stage of life and their family]. PMID- 11109841 TI - [The terminal care in the elderly: a position statement from the Japan Geriatric Society--a draft of JGS Ethics Committee]. PMID- 11109842 TI - [Quality of life at the end of life--from the viewpoint of feeding]. PMID- 11109843 TI - [What is the "terminal" of the elderly patients]. PMID- 11109844 TI - [Exercise training improves activities of daily living in elderly patients with congestive heart failure]. AB - Exercise training which is one of the multidisciplinary interventions for elderly patients with congestive heart failure, plays an important role for improving the quality of life and reducing the re-admission rate of these patients. We assessed the validity of exercise training for the improvement of patient's skeletal muscle functions and activities of daily living along with monitoring cardiac functions. Exercise training programs were performed in 12 patients with congestive heart failure (New York Heart Association class III or IV), including 5 with valvular disease, 4 with dilated cardiomyopathy and 3 with ischemic cardiomyopathy (mean 79 +/- 9 years). All patients were admitted because of exacerbation of congestive heart failure and were treated conventionally. The exercise training program was started after stabilization of their cardiac condition. The medication was not changed during the training period. After exercise training programs, the cardio-thoracic ratio decreased from 63.8 +/- 7.9% to 60.1 +/- 6.9% (p < 0.01), ejection fraction on echocardiography increased from 47.4 +/- 18.2% to 56.0 +/- 17.5% (p < 0.01), and brain natriuretic peptide decreased from 404.8 +/- 267.5 pg/ml to 313.6 +/- 239.5 pg/ml (p < 0.05). The quadriceps muscle power increased from 0.77 +/- 0.36 Nm/kg to 0.97 +/- 0.41 Nm/kg (p < 0.01). The maximum walking distance on flat surface increased from 149 +/- 164 m to 456 +/- 394 m (p < 0.05). In most patients, the activities of daily living, especially mobility, improved. Appropriate exercise training for the elderly patients with congestive heart failure improves activities of daily living and also reduces the amount of required care by the patients. PMID- 11109845 TI - [Why do medical students not have a geriatrics textbook?]. AB - To study the opinions of medical students on textbooks of geriatrics, we made a questionnaire for 5th-year medical students. All students had their own textbook (s) of internal medicine, and nearly half of the students had textbook (s) of physical examination. However, only 3.6% of students had their own textbook of geriatrics. The reasons for not having a textbook of geriatrics were as follows: 1) no understanding which textbook is better (59.1%), 2) no need to have a textbook (15.5%), 3) no appropriate textbooks (4.1%), and 4) others (17.6%). The most important point for choosing a textbook was as follows: 1) information from friends or seniors (38.9%), 2) looking by oneself (47.7%), 3) recommendation of a teacher (9.8%), and 4) others (3.6%). PMID- 11109846 TI - [Effects of calcium channel blockers on nocturnal polyuria in the elderly]. AB - Factors reducing the quality of life (QOL) related to micturition and their treatment were studied based on frequency volume chart for 52 consecutive patients with hypertension (the administration of calcium channel blockers) and 28 controls without hypertension, aged 50 years and over (mean age 68.6, range 50 85) without lower urinary tract dysfunction. The micturition records for two days and the QOL index related to urinary symptoms were reviewed. The factors reducing the QOL were increase of the voided volume and frequency during the night and the ratio of night-time voided volume to 24-hour urine output, which significantly increased in the controls (p < 0.05, respectively). Especially, the voiding frequency during the night and the ratio of night-time voided volume increased significantly in the controls aged 70 years and over (p < 0.01, p < 0.05, respectively). Overall, it was suggested that the administration of calcium channel blockers for control of blood pressure might be effective to reduce the nocturnal polyuria in the elderly. PMID- 11109847 TI - [The comparison of blood pressure of community-dwelling elderly subjects in Okinawa and Shikoku]. AB - In Japan hypertension is frequent, but the prevalence of hypertension in Okinawa has been known to be lower than in other areas in Japan. Now it has been almost 30 years since Okinawa reverted to Japan. So we investigated to know whether the prevalence of hypertension was still lower today or not. We compared the differences of prevalence of hypertension and blood pressure (BP) levels between the two community-dwelling elderly subjects aged 75 years or more: 305 in Ie in Okinawa (M:F = 107:198, mean age: 81 years old), and 99 in Omogo in Ehime, Shikoku (M:F = 45:54, mean age: 81 years old). We visited the homes of the elderly and measured their BP twice in a sitting position and asked them whether they were taking medicine for hypertension or not. According to the 1999 revised guidelines on the treatment of hypertension in the elderly by the Japanese Society of Geriatrics, we defined hypertension as systolic BP (SBP) > or = 160 mmHg or diastolic BP (DBP) > or = 90 mmHg or taking medicine for hypertension, and normotension as SBP < 160 mmHg and DBP < 90 mmHg and not taking medicine for hypertension. Hypertension rates were 52% in Ie, and 59% in Omogo, indicating no significant difference. However, in Ie only 54% of the elderly with hypertension were taking medicine for hypertension, as opposed to 74% in Omogo. These results suggest the possibility that in Okinawa hypertension is thought to be less important than in other districts in Japan. PMID- 11109848 TI - [A case of pneumonia caused by Mycobacterium avium complex successfully treated with clarithromycin]. AB - We reported on a 79-year-old woman with pneumonia caused by Mycobacterium avium complex (MAC). She was admitted with fever, general fatigue, and cough. A chest X ray film showed infiltrative shadows in the right lung field. In spite of administration of conventional antibiotics, the infiltrative shadows enlarged. A chest CT scan revealed areas of consolidation and ground glass opacities. Bronchoalveolar lavage (BAL) examination revealed an increased number of lymphocytes. Transbronchial lung biopsy revealed many granulomatous regions with giant cells. Mycobacterium intracellulare was found in the culture of BAL fluid and identified by PCR. Treatment was started with rifampicin, ethambutol, and clarithromycin. However, rifampicin and ethambutol were soon discontinued because of severe anorexia. Her symptoms and the radiographic appearance markedly improved following treatment of clarithromycin alone. Subsequently small doses of rifampicin and ethambutol were restarted because her general condition was much improved. These findings suggest that clarithromycin is an effective and tolerable agent for elderly patients with MAC. PMID- 11109849 TI - [The relationship between disability status and scores of comprehensive geriatric assessment]. PMID- 11109850 TI - A general axisymmetric contact mechanics model for layered surfaces, with particular reference to artificial hip joint replacements. AB - A general axisymmetric contact mechanics model for layered surfaces is considered in this study, with particular reference to artificial hip joint replacements. The indenting surface, which represents the femoral head, was modelled as an elastic solid with or without coating, while the other contacting surface, which represents the acetabular cup, was modelled as a two-layered solid. It is shown that this model is applicable to current total hip joint prostheses employing ultra-high molecular weight polyethylene (UHMWPE) acetabular cups against metallic, metallic with coating or ceramic femoral heads as well as metal-on metal combinations. The effect of cement is also investigated for these prostheses using this model. The use of a metallic bearing surface bonded to a UHMWPE substrate for acetabular cups is particularly examined for metal-on-metal hip joint replacements. Both the contact radius and the contact pressure distribution are predicted for examples of these total hip joint replacements, under typical conditions. Application of contact mechanics to the design of artificial hip joint replacements employing various material combinations is discussed. PMID- 11109851 TI - Long-term results for Kinemax and Kinematic knee bearings on a six-station knee wear simulator. AB - A long-term wear test was performed on Kinemax and Kinematic (Howmedica Inc.) knee bearings on the Durham six-station knee wear simulator. The bearings were subjected to flexion/extension of 65-0 degrees, anterior-posterior translation of between 4.5 and 8.5 mm and a maximum axial load of 3 kN. Passive abduction/adduction and internal/external rotation were also permitted, however, two of the stations had a linkage system which produced +/- 5 degrees active internal/external rotation. The bearings were tested at 37 degrees C in a 30 per cent bovine serum solution and the test was run to 5.6 x 10(6) cycles. The bearings from stations 2 and 3, and stations 4 and 5 were swapped during the test to investigate the effects of interstation variability. The average wear rate and standard error was 3.00 +/- 0.98 mg/10(6) cycles (range 1.33-4.72 mg/10(6) cycles) for the Kinemax bearings and 3.78 +/- 1.04 mg/10(6) cycles (range 1.87 4.89 mg/10(6) cycles) for the Kinematic bearings. There were no significant differences in wear rates between the different bearing designs, the addition of active internal/external rotation or a change of stations. However, the wear tracks were different for the two types of bearings and with active internal/external rotation. The wear rates and factors were generally lower than previously published in vitro wear results; however, this may have been due to a difference in the axial loads and lubricants used. The appearance of the wear tracks with active internal/external rotation was comparable with those seen on explanted knee bearings. PMID- 11109852 TI - A device for improved reduction of tibial fractures treated with external fixation. AB - A widely used method of treatment for unstable tibial shaft fractures is unilateral external fixation. The majority of fixators act as three distinct devices: an intra-operative reduction device, a device to maintain fracture alignment during healing and an aid to healing by allowing movement at the fracture site. Conventional operative techniques require the surgeon to manipulate a number of degrees of freedom at once, making reduction of the fracture difficult, and results in the fixator being out of alignment with the long axis of the bone. An operative method has been developed that separates reduction and fixation. A dedicated device has been designed to improve the per operative control of fracture fragments during fracture reduction. The device has been used in clinical trials for the reduction of 22 diaphyseal tibial fractures. Compared with previous operative techniques there has been a saving of 53 per cent in fracture reduction time and an overall saving of 10 per cent in operating time. Fracture alignment has been improved compared with reductions achieved with a fixator which potentially improves healing and lowers the rate of malunion. In each case the fixator has been applied in alignment with the bone, improving dynamization and reducing the likelihood of malunion due to fixator cam slippage. PMID- 11109853 TI - External ring fixators: an overview. AB - External fixation is widely used in the fixation of fractures and limb deformities. The mechanical characteristics of a specific external fixator are major factors in determining the biomechanical environment at a fracture/osteotomy site and, hence, affect the healing process. Although the optimal biomechanical environment for healing of a fracture or an osteotomy is unknown, a specific range of interfragmentary motion exists which promotes healing. It is therefore desirable that the mechanics of an external fixator can be manipulated to enable the surgeon to control the range of interfragmentary motion. The characteristics of an external fixator are defined by a large number of variables. Therefore, to gain control over the degree of interfragmentary motion, an understanding of the effect of each variable and how it interacts with the others to determine the overall characteristics of the device is required. For the past two decades, individual components and whole-frame configurations have been studied in depth. This article provides a summary of previous work concerning the mechanics of external ring fixators and how they affect the biomechanical environment at the fracture/osteotomy site. PMID- 11109854 TI - Radiographic assessment of the cement mantle thickness of the femoral stem in total hip replacement: a case study of 112 consecutive implants. AB - The results are reported of a radiographic study of cement mantle thickness in 112 consecutive primary hip replacements. Measurements were made by three observers of the apparent cement thickness medially and laterally using standard anterior-posterior radiographs. The average cement thickness was 3.2 mm, which is 1.2 mm greater than the size difference between the broach and the prosthesis, and was in the range 2-5 mm in 67 per cent of all measurement points. This has significance for the design of instrumentation to prepare the femoral cavity to give a defined cement mantle thickness. There was a greater cement mantle thickness proximally than distally. In 95 cases it was possible to determine the orientation of the stem within the cement mantle, which showed an even distribution between varus and valgus orientation; 49 per cent were within 1 degree of neutral and only one case was more than 5 degrees from neutral. PMID- 11109855 TI - Design of spinous process hooks for flexible fixation of the lumbar spine. AB - A prototype flexible fixation system for the lumbar spine was subjected to tensile testing to failure and cyclic tensile testing in order to determine any regions of weakness. The system consisted of a spinous process hook and two laminar hooks made of stainless steel (316L). Each laminar hook was attached to the spinous process hook by a loop of polyester braid secured by a crimped metal sleeve. In five tensile tests, the system failed by irreversible deformation of the spinous process hook at 2.5 +/- 0.3 kN (mean +/- standard deviation). In three cyclic tests, in which the applied tension varied sinusoidally between 0.04 and 0.4 kN at a frequency of 5 Hz, failure occurred after less than 400,000 loading cycles. This occurred as a result of fatigue crack initiation and propagation in the spinous process hook. A finite element model showed a stress concentration in the region where the crack occurred, which raised the applied stress above the tensile fatigue strength of this stainless steel. The spinous process hook was redesigned for manufacture in a titanium alloy (Ti-6AI-4V ELI) to minimize artefacts in magnetic resonance imaging. Further finite element models showed no unacceptable stress concentrations. PMID- 11109856 TI - Mechanical testing of a flexible fixation device for the lumbar spine. AB - The aim of this study was to test mechanically a new flexible fixation system for the lumbar spine. This device incorporates loops of polyester braid which are secured by a crimped titanium sleeve. In tensile tests, all loops failed, by slippage through the crimped sleeve, at 434 +/- 25 N (mean +/- standard deviation from five loops) for a single crimp and 415 +/- 15 N (from five loops) for two crimps. The intact system was then tested according to the ASTM standard. In a static test, all five specimens failed by slippage of the braid through the sleeve. Initial slippage occurred between 600 and 700 N, but the mean maximum load sustained was 1090 +/- 140 N. Dynamic tests were performed on ten constructs at a frequency of 5 Hz, under a range of loading conditions. The maximum load applied in any of the tests was 825 N. Two constructs did not complete the required 5 x 10(6) test cycles because of fracture of their spinous process hooks. However, other tests, under the same conditions, showed no signs of failure. Fracture occurred as a result of fretting damage from the recommended stainless steel roll pins. PMID- 11109858 TI - Parametric optimization for tumour identification: bioheat equation using ANOVA and the Taguchi method. AB - Breast cancer is the number one killer disease among women. It is known that early detection of a tumour ensures better prognosis and higher survival rate. In this paper an intelligent, inexpensive and non-invasive diagnostic tool is developed for aiding breast cancer detection objectively. This tool is based on thermographic scanning of the breast surface in conjunction with numerical simulation of the breast using the bioheat equation. The medical applications of thermographic scanning make use of the skin temperature as an indication of an underlying pathological process. The thermal pattern over a breast tumour reflects the vascular reaction to the abnormality. Hence an abnormal temperature pattern may be an indicator of an underlying tumour. Seven important parameters are identified and analysis of variance (ANOVA) is performed using a 2n design (n = number of parameters, 7). The effect and importance of the various parameters are analysed. Based on the above 2(7) design, the Taguchi method is used to optimize the parameters in order to ensure the signal from the tumour maximized compared with the noise from the other factors. The model predicts that the ideal setting for capturing the signal from the tumour is when the patient is at basal metabolic activity with a correspondingly lower subcutaneous perfusion in a low temperature environment. PMID- 11109857 TI - Time-dependent mechanical behaviour of the periodontal ligament. AB - The process of tooth displacement in response to orthodontic forces is thought to be induced by the stresses and strains in the periodontium. The mechanical force on the tooth is transmitted to the alveolar bone through a layer of soft connective tissue, the periodontal ligament. Stress and/or strain distribution in this layer must be derived from mathematical models, such as the finite element method, because it cannot be measured directly in a non-destructive way. The material behaviour of the constituent tissues is required as an input for such a model. The purpose of this study was to determine the time-dependent mechanical behaviour of the periodontal ligament due to orthodontic loading of a tooth. Therefore, in vivo experiments were performed on beagle dogs. The experimental configuration was simulated in a finite element model to estimate the poroelastic material properties for the periodontal ligament. The experiments showed a two step response: an instantaneous displacement of 14.10 +/- 3.21 microns within 4 s and a more gradual (creep) displacement reaching a maximum of 60.00 +/- 9.92 microns after 5 h. This response fitted excellently in the finite element model when 21 per cent of the ligament volume was assigned a permeability of 1.0 x 10( 14) m4/N s, the remaining 97 per cent was assigned a permeability of 2.5 x 10( 17) m4/N s. A tissue elastic modulus of 0.015 +/- 0.001 MPa was estimated. Our results indicate that fluid compartments within the periodontal ligament play an important role in the transmission and damping of forces acting on teeth. PMID- 11109859 TI - Influence of gelatin and bovine serum lubricants on ultra-high molecular weight polyethylene wear debris generated in in vitro simulations. AB - Ultra-high molecular weight polyethylene (UHMWPE) wear debris induced osteolysis has a major role in the late aseptic loosening and ultimate failure of total hip replacements (THR). Clinically relevant in vitro simulations of wear are essential to predict the osteolytic potential of bearing surfaces in artificial hip joints. Newborn calf or bovine serum has been accepted as a boundary lubricant for such in vitro tests, but its biological stability has been questioned. This study compared the wear factors, number of wear particles and levels of microbial contamination produced in bovine serum and a gelatin-based lubricant. The wear factors produced by the two lubricants were not significantly different, however the wear debris morphology produced was substantially different. The bovine serum became contaminated with micro-organisms within 28 h, whereas the protein-based lubricant remained uncontaminated. The results showed that bovine serum was not a stable boundary lubricant. They also showed that although the wear factors for the two solutions were not significantly different, the protein-based lubricant was not a suitable alternative to bovine serum because the wear debris produced was not clinically relevant. PMID- 11109860 TI - The design of a finger wear simulator and preliminary results. AB - A dual-cycle finger wear simulator has been designed, manufactured and commissioned. The simulator interspersed dynamic flexion-extension motion under light load with a heavier static 'pinch' load to a test prosthesis immersed in a lubricant heated to 37 degrees C. A validation test was undertaken on a size 2 Swanson prosthesis, leading to prosthesis failure in less than 1 million cycles. A second test was carried out on a Durham metacarpophalangeal prosthesis. After 4.8 million cycles a total wear factor for the joint of 0.60 x 10(-6) mm3/N m was calculated, with no cracks or damage visible. Both test results compare well with earlier tests undertaken on the Stokoe finger wear simulator. PMID- 11109861 TI - Assessment of the non-linear behaviour of plastic ankle foot orthoses by the finite element method. AB - The stiffness characteristics of plastic ankle foot orthoses (AFOs) are studied through finite element modelling and stress analysis. Particular attention is given to the modelling and prediction of non-linear AFO behaviour, which has been frequently observed in previous experimental studies but not fully addressed analytically. Both large deformation effects and material non-linearity are included in the formulation and their individual influence on results assessed. The finite element program is subsequently applied to the simulation of a series of tests designed to investigate the relation between AFO trimline location and stiffness for moderate and large rotations. Through careful consideration and identification of key modelling parameters, the developed finite element solution proves to be a reliable and effective alternative means of assessing variations of a typical plastic AFO design so that particular patient requirements could be met, in the long term. PMID- 11109862 TI - Review of arm motion analyses. AB - Interest in arm movements has increased tremendously in recent years. This interest has been motivated by different goals: the desire for a more scientific approach to replacement or support of the joints of the upper limb, the need for input to biomechanical computer models, and the clinical interest in comparing normal movements with pathological movements. The availability of commercial marker-tracking systems has facilitated achieving these goals. However, the complex nature of arm movements and the lack of standardized movements raises many challenges. In comparison with gait analysis, few arm motion analyses have been conducted. The purpose of this review is to aid researchers and clinicians interested in conducting an arm motion study in choosing the appropriate methodology. This is accomplished both by describing the methods used in past investigations and by highlighting important findings. Due to the variety of research goals, there is sometimes more than one appropriate method and the choice is left to the reader. Nevertheless, since it is extremely desirable to record and express the data in a standardized way, standardization proposals are described. This review, which focuses on methodology rather than results, addresses the following topics: motivations and tasks studied, tracking methods, the shoulder complex, joint centres and rotation axes, marker positions, coordinate system definitions, terminology and rotations, accuracy, and presentation methods. PMID- 11109863 TI - Hierarchical coding in the perception and memory of spatial layouts. AB - Two experiments were performed to investigate the organization of spatial information in perception and memory. Participants were confronted with map-like configurations of objects which were grouped by color (Experiment 1) or shape (Experiment 2) so as to induce cognitive clustering. Two tasks were administered: speeded verification of spatial relations between objects and unspeeded estimation of the Euclidean distance between object pairs. In both experiments, verification times, but not distance estimations, were affected by group membership. Spatial relations of objects belonging to the same color or shape group were verified faster than those of objects from different groups, even if the spatial distance was identical. These results did not depend on whether judgments were based on perceptually available or memorized information, suggesting that perceptual, not memory processes were responsible for the formation of cognitive clusters. PMID- 11109864 TI - Mixing incompatible mapped location-relevant trials with location-irrelevant trials: effects of stimulus mode on the reverse Simon effect. AB - When location-relevant trials with an incompatible spatial stimulus-response mapping are mixed with location-irrelevant trials, responses on the latter trials are faster when stimulus and response locations do not correspond than when they do. Experiments 1 and 2 showed that this reverse "Simon effect" also occurs when the location information is presented verbally or symbolically on both location relevant and location-irrelevant trials. The reversal was absent, however, in conditions of Experiments 1-3 in which the mode of presentation was different on the location-relevant trials than on the location-irrelevant trials. Experiment 4 demonstrated that differences in physical characteristics between the location relevant and location-irrelevant stimuli were not sufficient to eliminate the reverse Simon effect. These findings imply that the short-term associations between stimulus location information and responses defined for the location relevant task are relatively mode specific. PMID- 11109865 TI - Vertical versus horizontal spatial compatibility: right-left prevalence with bimanual responses. AB - For two-choice tasks in which stimulus and response locations vary along horizontal and vertical dimensions, the spatial compatibility effect is often stronger on the horizontal than vertical dimension. Umilta and Nicoletti [(1990) Spatial stimulus-response compatibility (pp. 89-116). Amsterdam: North-Holland] attributed this right-left prevalence effect to an inability to code vertical location when horizontal codes are present simultaneously. Hommel [(1996) Perception & Psychophysics, 43, 102-110] suggested instead that it reflects a voluntary strategy. This study reports four experiments that examine this issue. Experiment 1 was a conceptual replication of Hommel's Experiment 1, with responses made on a numeric key-pad and subjects instructed in terms of the vertical or horizontal dimension. The results replicated Hommel's findings that showed a right-left advantage with horizontal instructions; however, with vertical instructions, we found a benefit of vertical compatibility alone that he did not. This benefit for vertical compatibility alone was eliminated in Experiment 2 using a varied practice schedule similar to that used by Hommel. Experiment 3 showed right-left prevalence and a benefit of vertical compatibility alone, even with varied practice and vertical instructions, when subjects responded on perpendicularly arranged hand-grips. These benefits were eliminated in Experiment 4 using Hommel's method of urging subjects to respond only in terms of the instructed dimension. With bimanual responses, right-left prevalence is a robust phenomenon that is evident when comparing across vertical and horizontal instructions and, when the right-left distinction is relatively salient, within the vertical instructions condition alone. PMID- 11109866 TI - Spatial memory averaging, the landmark attraction effect, and representational gravity. AB - The effect of a large stationary landmark on memory for the location of a stationary target was examined. Memory for a stationary target was displaced toward the landmark, and targets that were larger, further from, or above the landmark exhibited greater magnitudes of displacement. Displacement was generally larger when the landmark vanished prior to judgment than when the landmark was visible during judgment. Memory for stationary targets offset from the major vertical or horizontal cardinal axis of the landmark was also displaced toward that cardinal axis. The data support the hypotheses that spatial memory averaging of the locations of a target and landmark occurs, and that this averaging may be combined with representational gravity in determining the remembered position of a stationary target. PMID- 11109867 TI - Modulation of inhibition of return by type and number of dynamic changes of the cue. AB - Inhibition of return (IOR) is a response delay when the target is preceded by an irrelevant stimulus (cue) at the same location. In a previous study, we investigated the separate and joint effects on IOR of cue onset and offset. IOR was much greater when cue onset was followed by cue offset (on-off cue) than when the cue was a single event (on or off cues). The aim of the present study was to test whether the greater IOR with an on-off cue is due to the presence of two cue events. Three experiments were conducted. In Experiment 1 we replicated, with a different delay between cue onset and offset, the finding that IOR is greater with an on-off cue than with a single cue event. In Experiments 2 and 3, we used cues formed by two events. In Experiment 2, an on-off cue was compared with an off-on cue, whereas in Experiment 3 an on-on cue was compared with an off-off cue. Results showed that the magnitude of IOR did not simply depend on the number of cue events occurring before the target. IOR was greater with two different events than with two identical events and greater when was preceded by an off event than an on-event. Therefore, IOR was greatest with an on-off cue, which likely also benefited from a gap effect. Possible mechanisms underlying IOR were discussed. PMID- 11109868 TI - Effects of spatial and symbolic precues on localization performance. AB - One of the fundamental properties of spatial vision is the ability to localize objects in space. According to a recent proposal, accurate localization performance involves the operation of two systems: the attention system and the eye movement system. Upon stimulus presentation, attention is shifted to the target area: this provides coarse location information. Subsequently, a saccadic eye movement is executed: this provides fine location information. In this study we tested predictions derived from this model concerning the effects of precue information on localization performance. In a series of five experiments we manipulated duration of precue (71, 400, and 1,000 ms) and type of precue (spatial versus symbolic). Results showed that very short duration (i.e., 71 ms) spatial precues improved localization performance whereas very short duration symbolic precues did not. In contrast, the 1,000 ms duration precue condition showed similar amounts of precuing benefit for the spatial and symbolic precues. This pattern of differential precuing effects corroborated the two-process model of localization performance. PMID- 11109869 TI - Polypharmacy and non-compliance in the hypertensive elderly patient. AB - BACKGROUND: Elderly patients are major consumers of prescription and nonprescription medications and the proper use of these agents can lead to more cost-effective strategies in reaching optimal health. The use of medications for the treatment of multiple co-morbid conditions in a single patient increases the risks of polypharmacy and non-compliance and raises the burden on the health care system and society. OBJECTIVES: To analyse the extent and nature of total and anti hypertensive, polypharmacy the most prevalent groups of drugs, the compliance rate and the costs related to polypharmacy in the hypertensive elderly. METHODS: A descriptive cross-sectional study was made of sixty-nine patients 65 years of age or older on follow-up for arterial hypertension at a central hospital in Lisbon. The study protocol consisted of a questionnaire performed by the physician. We calculated the monthly amount of individual prescription expense, after cost reduction from social insurance. RESULTS: The patients used an average of 4.4 prescribed medications with a corresponding average of 6.4 pills per day. Drug use was greater in women than men. Hypertensive therapy involved a mean of 2 drugs. Antiplatelet drugs, coronary vasodilators, bezodiazepines, glucose regulators and hypolipidemic agents were the other major groups of drugs. Non-compliance was identified in only 14% of the patients. The average of individual prescription expenses was 5,076 Portuguese escudos (PTE) per month, of which 2,226 PTE was the average cost of antihypertensive agents. CONCLUSIONS: With this study we were able to show the extent of poly-pharmacotherapy in a population of hypertensive elderly patients. We found a high rate of compliance, although the costs were frequently high. PMID- 11109870 TI - Inhibition of the carotid baroreflex by urinary bladder distension. AB - OBJECTIVE: The cardiovascular responses to urinary bladder distension include a rise in arterial blood pressure, an increase in vascular resistance and heart rate, which are abolished by sectioning the pelvic and hypogastric nerves. There are indications in the literature that inputs from the arterial baroreceptors and urinary bladder converge at the same autonomic efferent pathways. Recent data provided evidence of a partial inhibitory action of the carotid baroreflex on the reflex cardiovascular responses evoked by bladder distension. In this study, we investigate the influence of urinary bladder distention upon cardiovascular responses elicited by carotid baroreceptor stimulation. METHODS: Wistar Rats were anaesthetised with alpha-choloralose (80 mg/kg, i.p.) supplemented as necessary. The carotid sinus was stimulated randomly with different volumes of saline (0.5 2.0 ml). The ureters were ligated and the bladder was cannulated. Arterial blood pressure (BP), ECG, heart rate (HR), and bladder pressure (BLP) were monitored. Baroreceptor stimulation was undertaken on two levels of BLP (2 and 20 cmH2O). The cardiovascular response to baroreceptor activation was assessed as the maximal variation of BP, HR and R-R interval for different volumes of saline injected. RESULTS: Our results showed that in all tests baroreceptor stimulation evoked a decrease in BP and HR but the magnitude of these responses was significantly different for each volume of saline and for two levels of BLP. The effectiveness and reproducibility of responses to these stimuli were confirmed by injecting a fixed volume of saline into the carotid sinus of two animals at two levels of BLP. In five animals, the variation in BP increased as the magnitude of the stimulus to the baroreceptor increased. However, the response was reduced with increased BLP. The fitted regression line was calculated for all the tests (n = 82; s = 8.2; Yint = 0.62 for BLP = 2 cmH2O; s = 9.5; Yint = 2.9; r = 0.69 for BLP = 20 cmH2O). Similar effects were observed and similar patterns of regression lines were obtained with regard to HR changes (n = 48; s = 13.3; Yint = 2.7; r = 0.82 for BLP = 2 cmH2O; s = 15.8; Yint = 0.89; r = 0.76 for BLP = 20 cmH2O) and interval changes (n = 48; s = 7.1; Yint = 4.3; r = 0.78 for BLP = 2 cmH2O; s = 8.5; Yint = 0.2; r = 0.76 for BLP = 20 cm H2O). CONCLUSIONS: These data suggest that the cardiovascular responses evoked by carotid baroreceptor stimulation are significantly different when the urinary bladder is empty or full with the response diminishing as the urinary bladder is distended. PMID- 11109871 TI - Quality of life in patients undergoing coronary revascularization. AB - INTRODUCTION: This study was aimed at the evaluation of the impact of coronary revascularization surgery on the quality of life (QOL) and identification of some variables which may contribute to influence the patients' own perception of their health condition. METHODOLOGY: We prospectively studied 150 consecutive patients subjected to isolated coronary bypass surgery during a 3-month period, with evaluation of their perception of QOL before surgery and 6 months thereafter. The measurement instruments used were the MOS Health Survey (SF-36) and the Nottingham Health Profile (NHP). Additionally, a questionnaire for identification of lifestyles was introduced. RESULTS: The majority of the patients were male (94%), above 50 years of age (81%), with a low educational level (65%), married (90%) and pensioners (44%). About one third (38%) had marked physical limitations (CCS class III/IV), with comorbidity (80%), previous myocardial infarction (49%) and 3-vessel coronary disease (68%). There was no operative mortality, but 29% had postoperative complications, albeit minor in the majority. Admission time was less than 8 days in 88.6% of the cases. Surgery proved beneficial in improving QOL, with better perception after surgery in all dimensions of both measurement instruments (p < 0.001). A higher level of education was related to a better perception of the energy dimension, married patients had a better improvement in the dimensions of physical pain and social isolation. More severe preoperative angina determined less favourable scores in the dimensions of mental health, social function and vitality; comorbidity had a negative impact on vitality and physical mobility; and the number of risk factors and postoperative complications had a negative impact on the dimensions of energy and emotional reactions, and social isolation and physical function, respectively. Six months after surgery, 62% of the patients who were still working before surgery had resumed their professional activity, but about 20% had retired; the majority had adopted healthier lifestyles with a decrease in tobacco and alcohol consumption and a more balanced diet. CONCLUSIONS: Coronary revascularization substantially improves QOL, with a significant impact on the clinical variables of the psycho social dimensions. PMID- 11109872 TI - [Evidence-based cardiology in the Revista Portuguesa de Cardiologia]. PMID- 11109873 TI - [Scientific evidence-based cardiology: the principles and practice]. AB - Every clinical cardiologist, no matter what the clinical work (invasive or non invasive), has to face several problems concerning clinical knowledge and medical information in everyday practice. Diagnostic and therapeutic advances in cardiology are occurring at an increasing pace and every cardiologist responsible for patient care in hospitals, clinics or emergency rooms needs to be up-to-date in order to provide the best possible medical care. On the other hand, society is increasingly making doctors accountable for the provision of good quality care in a cost-effective way. In the context of scarce resources, this situation requires a rigorous and rational approach by the individual cardiologist. These apparent contradictions can be solved by practicing evidence-based cardiology (EBC). This review introduces the concept of EBC, its principles, practice, and methodological steps: 1) formulation of a structured clinical activity; 2) scientific evidence; 3) critical appraisal of this evidence using explicit methods; and 4) synthesis and practical application of this evidence. In this sense, EBC arises from the patient, and after the best possible scientific evidence is selected, it is applied to the individual case. EBC allows the individual cardiologist to keep up with the medical literature while improving reading habits and the form in which relevant clinical information is selected. It also increases confidence in the clinical decisions, reducing practice variation as well as improving doctor-patient communication. Lastly EBC can be used as a powerful tool for pre, post and continuous medical education. PMID- 11109874 TI - [Rheumatic fever--a review of cases]. AB - OBJECTIVE: To analyse clinical presentation of rheumatic fever (RF), with special emphasis on cardiac involvement, electrocardiographic and echocardiographic findings and the outcome of the cases referred to Maria Pia Children's Hospital from January 1990 to September 1999. METHODS: We retrospectively analyzed the clinical files of all cases referred to pediatric cardiology clinics with the suspicion of acute RF (Group 1) or with rheumatic valvular disease and heart failure (Group 2). In group 1 we studied the following: age and sex distribution, year of diagnosis, presence of Jones criteria treatment and outcome. In group 2 we analysed provenance, age of initial onset of RF, age of cardiology referral, treatment and outcome. RESULTS: Thirteen cases were identified, 8 in groups 1 and 5 in group 2. Group 1 included 3 girls and 2 boys, mean age of 10 years. The diagnosis of RF was based in the presence of 2 major and 1 minor manifestation (4/8), 1 major and 2 minor manifestations (1/8) and chorea in 3 cases associated with clinical carditis in one and subclinical carditis in another. Colour Doppler echocardiography showed pathological mitral regurgitation jet in 6 cases, associated with aortic regurgitation in 2 and dilatation of left ventricle in 3. All were treated with penicillin associated with anti-inflammatory drugs in 5 and haloperidol in 3. Group 2 included 3 girls and 2 boys, mean age 9.56 years. Four were from African countries (Angola and Guinea), and one came from the north of Portugal. The elapsed time between the initial acute attack and cardiology referral varied from 5 months to 3 years. All presented severe mitral insufficiency associated with aortic and/or tricuspid valve lesions, and heart failure. All five underwent valve surgery. The secondary prophylaxis was recommended in every patient. There was a recurrence in a child who had interrupted chemoprophylaxis. The patients from African countries were lost for follow-up. CONCLUSIONS: RF still remains a problem in present times, with serious cardiac sequela in African countries. Colour Doppler echocardiography is a valuable tool for the detection of pathological valvular regurgitation and subclinical carditis if strict criteria are used. The need for appropriate treatment of streptococcal pharyngitis and secondary prophylaxis is emphasized. PMID- 11109875 TI - Acute myocardial infarction due to occlusion of the left main. PMID- 11109876 TI - Renovascular hypertension due to angiodysplasia. PMID- 11109877 TI - [Trends in coronary artery bypass surgery results: a recent, 9-year study]. PMID- 11109878 TI - Brief history of otology in the Bordeaux School. PMID- 11109879 TI - The cost of running a multidisciplinary head and neck oncology service--an audit. AB - It seems well established that a modern approach to the treatment of head and neck cancer should involve a multidisciplinary team. However the "cost" of providing such a service in the United Kingdom has not as yet been investigated. We present the results of a detailed, prospective audit which has identified and quantified the "cost" of input from all members of the team involved in the in patient care of these patients. We also discuss some of the resourcing issues, which arise from this work. PMID- 11109880 TI - Evaluation of the primary tumour and the metastatic lymph node with or without extracapsular spread by means of argyrophillic nucleolar regions (AgNOR). AB - Though TNM staging is the most popular clinical system for defining the prognosis of Head and Neck Squamous Cell Carcinoma (HNSCC), different clinical trials of the tumours with the same TNM stage have caused doubts about this system. Extra Capsular Spread (ECS) in a metastatic lymph node is one of the recently defined prognostic factors in HNSCC. The hypothesis of this study was to assess the relationship between the primary tumour of laryngeal and oral cavity squamous cell carcinoma and the metastatic lymph nodes with and without ECS. The argyrophillic Nucleolar Organiser Regions (AgNOR) count was used as an index of the grade of malignancy of the neoplastic tissue. As a result, significantly higher AgNOR counts were obtained in the metastatic lymph nodes with ECS than in the primary tumours, while the lowest AgNOR counts were found in the metastatic lymph nodes without ECS. However, when primary tumours of the metastatic lymph nodes with ECS and the primary tumours of the metastatic lymph nodes without ECS were compared, no significant difference was found. PMID- 11109881 TI - [Papillary carcinoma from the thyroglossal duct cyst: review of the literature and report of a case]. AB - Cancer of a thyroglossal duct cyst is very rare. Clinical presentation is identical that of a benign cyst, and the diagnosis is histopathological. We report the case of 36 year-old woman treated for papillary carcinoma of a thyroglossal duct cyst. The treatment was surgical excision according Sistrunk method. There was no distant spread. After 22 months follow up, the patient is alive, with no signs of recurrence. PMID- 11109882 TI - [Essential parathyroid cysts: a misleading lesion]. AB - Though a rare lesion, non-functioning parathyroid cyst is of clinical significance because it usually mimics a thyroid nodule. The cyst can be ectopic in location and therefore constitutes a differential diagnosis to a bronchial or thymic cyst. Two recent cases of non-functioning parathyroid assay are reported. Needle puncture with estimation of the level of parathyroid hormone in the aspirate allows the diagnosis to be made before surgery. PMID- 11109883 TI - [Parathyroid carcinoma: diagnosis and treatment]. AB - Pre operative and per-operative suspicion, and recognition of parathyroid carcinoma are essential in the effective approach and the prognosis of this type of tumor. On the basis of our experience over 20 years, helped by the literature we review our 179 cases of primary hyperparathyroidism out of which 5 were malignant tumors and the 174 others were benign (adenomas and hyperplasias). We conclude that patients with primary hyperparathyroidism, reflected by high levels of serum calcium (75%) and elevated levels of parathormone (100%) are suspect of carrying a parathyroid malignancy particularly when these levels are profoundly abnormal. Furthermore, a severe clinical presentation featuring, palpable cervical mass, renal symptoms in 30% to 60% of all, bone disease in 90%, is suspect of malignancy. The operative findings such as important size of tumour, local seeding, local invasion are also significant features of malignancy. All this data gathered by comparing carcinomas to adenomas suggest and confirm what many reviews on the subject have reported, early recognition of the parathyroid carcinoma is of primal importance, because aggressive initial surgical management should be applied, as total extirpation of the malignancy is the only treatment allowing maximal survival rates. PMID- 11109884 TI - [Adenoid cystic carcinoma (CAC) of the naso-sinonasal cavities: report of 5 cases, review of the literature]. AB - Adenoid cystic carcinoma is a rare tumour which mainly affects the major and accessory salivary glands. We report 5 new cases affecting the nose and sinuses, and based on these review the literature to determine how this disorder can be better treated, given the significant morbidity and mortality of the condition. PMID- 11109885 TI - [Stapedotomy and anatomical variations of the facial nerve]. AB - Among the 595 stapedotomies performed between 1992 and 1999 (surgeon R. Hausler), there were 40 cases (6.7%) where the facial nerve had an abnormal course. In 32, a partial nerve prolapse over the oval window was noted with (6 cases, 1 being a duplicated nerve around the oval window) or without (26 cases) dehiscence in the long bony canal. In 8 cases, there was a total prolapse of the nerve over the oval window, with 2 special cases: facial nerve having an inferior course over the oval window and the promontory; facial nerve being widely spread over the oval window and the promontory. Concomitant anomalies of the stapes were seen and several patients had dysmorphic syndromes with conductive hearing loss since early childhood. Stapedotomy was performed in 39 patients. In the 32 cases of partial nerve prolapse, a small piston (0.4 mm) was placed in the lower part of the oval window which was sometimes enlarged towards the promontory, except when the nerve was duplicated: the prosthesis was placed into the footplate between the nerve branches. In the 8 patients with total facial nerve prolapse, the prosthesis was either placed directly in a burr hole into the promontory just below the oval window (6 cases), or, when the nerve ran over the promontory and over the oval window, the prosthesis was placed above the oval window at the site where the facial nerve is usually located (1 case). In the case where the nerve was spread widely over the oval window and the promontory, no prosthesis was placed. In the 39 patients where a stapedotomy was performed, the average hearing level gain ranged from -15 dB to 40 dB (average: 18 dB) at 0.5, 1, 2 and 4 kHz. The average residual air-bone gap was < 30 dB in 36 patients (92.3%), < 20 dB in 30 (77%) and < 10 dB in 16 (41%). A post-operative additional hearing loss > or = 10 dB occurred in 3 cases (10, 12.5 and 12.5 dB). There were no cases of post operative deafness or facial palsy. This analysis shows that in many cases with an aberrant course of the facial nerve, stapedotomy using adequate and sometimes non-conventional techniques can give post-operative hearing improvement. PMID- 11109886 TI - [Abnormal trajectory of the internal carotid artery in the middle ear. Report of a case]. AB - The authors report a case with an aberrant course of the internal carotid artery within the middle ear--a rare abnormality with only about fifty cases so far reported in the literature. The combination of pulsatile tinnitus and a retro tympanic mass calls for a precise anatomical investigation of the temporal bone before any surgery is undertaken. The role of the various complementary investigations is discussed. A high resolution CT scan is the key investigation, and is sufficient to give the diagnosis in the majority of cases. Angio-MRI currently affords the possibility of confirming this vascular malpositioning in a non-invasive way. Arteriography should be used only when there is remaining doubt about the possibility of a glomus tumour, or before treatment by clamping. The most logical course to adopt would seem to us to be to abstain from treatment once this diagnosis has been made, given the risk of major haemorrhage and the potential neurological complications. PMID- 11109887 TI - [Cochlear implantation in the adult and the child. Results of the Marseille experience between 1991 and 1999. AB - The rehabilitation of patients with profound bilateral deafness has been notable in the 1990s for the use of cochlear implantation. In 1991 a cochlear implantation programme was inaugurated by the ENT team at the Timone University Hospital, in partnership with the Public Assistance for Marseille Hospitals, allowing funding for six implants each year. This article presents the main results of this programme. Between March 1991 and November 1999, 94 patients with profound bilateral deafness were assessed. Forty patients (29 adults and 11 children) were thought to be unsuitable. To date 27 adults (4 with prelingual deafness) and 22 children have been rehabilitated using a cochlear implant. Patient selection was carried out by a multidisciplinary team. It has been possible to quantify the degree of deafness and the excitability of the auditory nerve in the adult, the cochlear permeability, and the motivation and personality of the patients. Postimplant evaluation has allowed us to study the various factors resulting from wearing an implant. In adults, testing was carried out using open lists. In children, education methods were added to perceptive, expressive and behavioural performance to arrive at a profile. The results are presented and discussed in comparison with those found in the literature. PMID- 11109888 TI - [Facial paralysis: treatment with an acyclovir-methylprednisolone combination, preliminary results]. AB - RATIONALE: Many actual data suspect a viral etiology to Bell's palsy. Herpes viridae are more and more incriminated. On these basis, we have studied the efficacity of parenteral association of 30 mg/Kg/j of Aciclovir and 1 mg/Kg/j of Methylprednisolone in the treatment of Bell's palsy with less than 12 days of evolution. METHOD: The evaluation concern 53 patients. We also evaluated the functional motor result (using House and Brackmann staging) and search prognostic factors in clinic and paraclinic data. A viral investigation have been made in most of the cases. RESULTS: Only one of our patients treated with Aciclovir keeps some sequeles (stage III of House and Brackmann classification). We didn't found any deleterious effect. Statistically, the electromyography is the only one test with a prognostic value. But we feel that delay between the starting and the treatment of the palsy is important. Viral tests show sometime abnormal Ig against the Herpes viridae group. CONCLUSIONS: These promising results are lightened with a review of the literature. A multicentric investigation is already in place for a stronger statistic effect. PMID- 11109889 TI - [Vestibular neurotomy by a retrosigmoid approach: functional results. Evaluation through a triple clinical approach (otoneurologic, orthoptic and physical medicine)]. AB - Vestibular neurotomy remains the surgical procedure of choice in the management of peripheral vertigo resistant to the usual medical treatment, for patients with preserved hearing. Meniere's disease generally sums up most of the surgical indications. The authors report a personnel series of patients who underwent vestibular neurotomy during the last five years, from January to December 1998. The aim of this study was to assess with a 6 months to 3.5 years follow-up, postoperative complications and functional results, particularly the improvement of dizziness, residual unsteadiness and its impact on quality of life, and the recovery of socio-professional and physical activities. Of a total of 41 patients, 21 were evaluated by both an otoneurologic and physical medicine approach, to analyze the main sensory components of equilibrium (vestibular, proprioceptive, cervical, visual). In most of the cases, the results show disappearance of vertigo which was the main complaint. However, disabling residual instability is common in those patients who was present both a cervical pathology and binocular visual impairment or a defect in convergence, often unrecognised. The authors emphasise the necessity of a cervical examination and orthoptic investigations in addition to the vestibular assessment. This multi disciplinary approach allows better identification of the different factors (vestibular, cervical, orthoptic) and lead to specific rehabilitation which can permit the patient with residual unsteadiness to return to work and lifestyle, the real measure of success of vestibular neurotomy. PMID- 11109890 TI - [The physician and death]. PMID- 11109891 TI - [Sexual development of the child and the onset of gender identity]. AB - In view of the numerous questions raised by the recently disclosed pedophilia, sexual abuses and prostitution of children, it appeared worthwhile to review the recent studies dealing with the child sexual development and the onset of the gender identity. These studies consist partly in retrospective analysis of pathological situations and partly in the analysis of normal development. It appears that the social and emotional surroundings of the child, provided that they were continuous and devoid of ambiguity, play a prominent role in this process. At present, numerous authors consider that gender identity is achieved at about 2 years of age when the child has a harmonious general development. The family physician and the paediatrician are on the first line to detect potential problems provided that they were aware of the main determinants and symptoms involved. PMID- 11109892 TI - [Clinical approach to inflammatory syndromes in the aged]. AB - Primary care physicians have often to provide care to elderly patients presenting with non specific general complaints such as anorexia, weight loss and fatigue associated with biological inflammatory tests (increased erythrocyte sedimentation rate, increased CRP, anemia of inflammatory origin). In elderly patients, inflammatory diseases of unknown origin are most often related to an infectious illness (particularly bacterial endocarditis or tuberculosis), a systemic autoimmune disorder (temporal arteritis, polymyalgia rheumatica or ANCA positive vasculitis) or a neoplastic process. A methodological clinical approach is discussed and the most valuable complementary tests are proposed. PMID- 11109893 TI - [Reconstructive surgical techniques after palliative tumor excision]. AB - Palliative care is more and more of concern in medical information. If there is no exact definition about the real place of surgery in this concept, the rule of reconstructive surgery is even less clear. The goal of this paper is to try to define, on one hand the general characteristics of palliative surgery, and on the other hand, to underline the role of reconstructive surgery in four specific indications: the head and neck area, the thorax, the pelvis and the extremities. The resection surgery, associated with flap coverage, may seems an excessive treatment in such palliative indications. Nevertheless, the results presented here tend to demonstrate that, after careful patient selection a better quality of life could be obtained with an acceptable surgery-induced morbidity (regarding preoperative disability). More than technical aspect, it is by a different approach of the problem that this type of surgery is to be distinguished. PMID- 11109894 TI - [Prevention of cardiovascular diseases and patient compliance: limiting factors and proposals]. AB - Despite epidemiological evidence in favour of cardiovascular prevention, many surveys show that only a minority of patients with coronary heart disease are receiving hypolipidemic therapy. The compliance is the neglected issue in preventive cardiology, for many reasons. The patients are reluctant to change their long lasting nutritional and smoking habits. The physicians are more motivated by acute care and are not trained in preventive medicine. Moreover there is too often a lack of interface between specialists and GP. To improve compliance, it is necessary to give clear instructions and tailored regimens. The treatment must be monitored and the follow up organised. Last but not least, the reimbursement of hypolipidemic drugs should be adapted to Evidence Based Medicine. PMID- 11109895 TI - [Unsuspected hepatic rupture with peritoneal extravasation of contrast media revealed by computed tomography]. AB - Computed tomography (CT) plays a major role as the initial diagnostic tool in adult blunt abdominal trauma. Indications of CT are hemodynamical instability, abdominal defense and high-speed deceleration. We present the case of a 22 year old man involved in a motor-vehicle accident admitted in our emergency department with headache but without clinical signs of intraabdominal trauma. The abdominal CT was asked because of the high-speed deceleration and because the patient had to come to the CT room for a head CT. The abdominal multislice helical CT performed with contrast bolus injection showed a massive intraperitoneal extravasation from a hepatic laceration and avulsion. The patient died during operation despite immediate management of the abdominal traumatic lesions detected by CT. PMID- 11109896 TI - [Glitazones (thiazolidinedione)]. AB - Insulin resistance is the major defect in type 2 diabetes. Troglitazone is the first of a new class of drugs, the thiazolidinediones (TZD) with insulin sensitising actions. The TZD activate PPAR gamma (Peroxisome Proliferator Activated Receptor gamma). In clinical trials, the TZD decrease plasma glucose, plasma insulin and Hb A1C. Moreover they are synergistic with the glucose lowering drugs (biguanides and sulfonylureas) and with insulin therapy. The TZD also decrease triglycerides and increase HDL cholesterol, while reducing LDL oxidation. Few side effects have been reported but concerns persist about their safety. Hepatic dysfunction is seen in about 2% of the patients receiving troglitazone, leading sometimes to liver failure. This potentially lethal side effect appears to be less frequent with the second generation TZD, rosiglitazone and pioglitazone. Drug interactions, fluid retention, induction of colon polyps are other potential unwanted effects. This new class of drugs could play an important role in diabetes therapy, if clinical trials prove their long term efficacy and safety. PMID- 11109897 TI - [Multiple birth]. PMID- 11109898 TI - [Implications of early discharge of the newborn from hospital]. PMID- 11109899 TI - [Foundation of the Free University of Brussels Institute of Mucoviscidosis]. PMID- 11109900 TI - [Nutritional aspects of mucoviscidosis]. PMID- 11109901 TI - [Care of the child with severe asthma]. PMID- 11109902 TI - [Paul A. Bastenie (1906-1985)]. PMID- 11109903 TI - [The Bone and Joint Decade 2000-2010]. PMID- 11109904 TI - [AA, AL and A beta 2M amyloidoses as seen by the rheumatologist]. PMID- 11109905 TI - [Current treatment of AL amyloidosis]. AB - AL amyloidosis is a plasma cell disorder characterized by the tissue accumulation of immunoglobulin light chains. The prognosis of AL amyloidosis is poor, with a median survival inferior to two years. Amyloid deposits, responsible for the clinical manifestations of this disease, can regress at least partially after the suppression of the production of amyloid precursors. Therefore, current treatment strategies in AL amyloidosis aim at reducing the plasma cell clones, using chemotherapy regimens applied in multiple myeloma. A combination of melphalan and prednisone is the standard therapy of AL amyloidosis, but its results remain disappointing. Intensive chemotherapy regimens, with high-dose melphalan followed by peripheral blood stem cell transplantation, lead to increased response rates and improved survival, but are complicated by severe short term morbidity and mortality. These regimens offer the best clinical perspectives for selected patients, until the introduction of innovative agents capable of interfering with amyloid fibril formation. PMID- 11109906 TI - [Takayasu's arteritis--case report and review of the literature]. AB - A 26 year old Albanian woman presented with intermittent claudication of upper limbs in association with alleviation of radial pulses, reduction of arterial pressures, bilateral axillary bruits and subocclusive lesions of proximal part of both humeral arteries on arteriography and MRI. Takayasu's arteritis was diagnosed according to ACR criteria. A treatment of prednisone was started together with methotrexate. The response was favourable and symptoms like claudication and malaise vanished. Takayasu's arteritis is a vasculitis which affects large vessels such as aorta and its main branches. This disease involves mainly premenopausal women; it is very rare in Europe. Diagnosis is lying on clinical features and arteriography results. Treatment of choice is corticosteroids, and immunosuppressors; sometimes a surgical procedure is necessary if stenosis is fixed. PMID- 11109907 TI - [Treatment of gout]. PMID- 11109908 TI - [Prevention and treatment of corticosteroid-induced osteoporosis]. AB - Osteoporosis and fractures are frequent and severe consequences of long-term treatment with glucocorticoids. Trabecular bone is mainly affected with a decrease of bone formation and an increase of bone resorption. Prevention and treatment of corticosteroid-induced osteoporosis is based upon general measures such as calcium and vitamin D supplementation, adequate+ protein intake, regular physical exercise, hormonal replacement therapy and upon specific means like therapies used in primary osteoporosis. Bisphosphonates which are potent bone resorption inhibitors have been shown to increase bone mineral density and to decrease fracture rate. Therefore they appear as first choice in the prevention as well as in the treatment of corticosteroid-induced osteoporosis. PMID- 11109909 TI - [Treatment of coxarthrosis with fitted total hip prostheses replacement]. AB - The treatment of hip osteoarthritis with total hip arthroplasty has continuously evolved since it was first introduced in the sixties. The problem of aseptic loosening of the cemented prostheses, mainly in young active patients, has stimulated two different types of research: on one side the improvement of cementing techniques and on the other side the development of cementless osteoinegrable implants. We discuss the problems of these cementless hip prostheses. Recently published anatomic and biomechanic studies have led to the development of personalized custom femoral stems for each patient. The conception technique and first clinical results are described. PMID- 11109910 TI - [Metatarsalgias]. PMID- 11109911 TI - ["My child walks on his tiptoes"]. AB - Toe walking is a symptom that occurs frequently in the normal childhood population. The management of children who walk in a symmetrical and permanent way on the toes is presented. In most cases no etiology of toe walking is found. Idiopathic Toe Walking (ITW) is considered a diagnosis of exclusion and is employed only when all other possibilities have been eliminated with a meticulous clinical examination and various investigations. If any etiology is found, the treatment should be first non operative whereas heel cord lengthening is the treatment of choice in ITW. This surgical treatment should be limited to children with functional impairement and is ideally carried out as late as possible. PMID- 11109912 TI - ["Round back" in children and adolescents]. AB - The hyper-kyphosis or "the postural round back" is one of the most common complaints in orthopedic practice. In the majority of cases, the thoracic kyphosis are painless and flexible. The vertebral bodies are normal on radiograms. This is "the kyphotic attitude" or postural round-back. A medical treatment is not the necessary rule. Life hygiene, sports and simple supervision are needed. Nevertheless, there are still pathological fixed kyphosis, induced in the majority of cases by Scheuermann's disease. The other possible etiologies (congenital, paralytic, post-traumatic, Pott's disease, postradiation, or metabolic origin) are a lot rare ones and will be excluded by clinical examination and imaging studies. The structural hyper-kyphosis require treatment. We will approach successively steps of the diagnosis and treatment of the hyper kyphosis of the adolescent. PMID- 11109913 TI - [Public health research personnel and health and prevention program personnel: an impossible marriage? The example of research in adolescent health]. PMID- 11109914 TI - [Diagnosis and treatment of ascites: recommendations for clinical practice. "RCP Ascites" workgroup]. PMID- 11109915 TI - [What is your diagnosis? Mandibular dental fistula. Syn: dentogenic fistula, cutaneous dental sinus]. PMID- 11109916 TI - [Therapy concept in differentiated thyroid gland carcinoma--results of 25 years with 257 patients]. AB - BACKGROUND: Differentiated thyroid carcinoma is a unique tumour in that a low risk-patient population with a tumour-related death rate of near 0% can be found. Yet in these patients the risk and risk factors of curable recurrences must be considered. The question therefore arises, whether in defined subgroups of low risk-patients a reduced extent of treatment (hemithyroidectomy, total thyroidectomy without 131I ablation) may result in cure without recurrence, including low morbidity of treatment and reduced costs. STUDY DESIGN: In a consecutive series of 257 patients suffering from papillary (146) or follicular (111) carcinomas operated by one surgeon over a period of 25 years, a reduced extent of treatment was carried out essentially in subgroups of minimally invasive follicular carcinoma, namely in pT1,2-tumors of young patients, or in those with capsular invasion alone, and in pT1,2 N0 papillary tumors, representing a subgroup of TNM stage I and II tumors. For N-staging selective lymphadenectomy was carried out at the beginning of the study, but elective lymphadenectomy was used since 1996 for papillary carcinoma. Follow-up was 1-25 (mean = 8) years. All excised tumors were examined by one pathologist. RESULTS: 167 (approximately 2/3) of the patients presented with a single nodule, whereas in 1/3 a concomitant benign nodular goiter or an immunothyropathy was found. The percentage of grossly invasive follicular carcinoma decreased during the 25 year period from 41 to 10% of all patients (p < 0.0005), whereas the percentage of papillary cancer rise from 35% to 66% (p < 0.005). Hemithyroidectomy, total thyroidectomy without, and with 131I ablation respectively, were performed in 32%, 24%, and 44% of patients. In papillary carcinoma N1-status was found in 21 (23%)/ 92 patients with selective, and in 18/54 patients (33%) with elective lymphadenectomy, respectively (n.s.). One (0.4%) patient died postoperatively. Permanent hypoparathyroidism occurred in 1.9% (3% for total thyroidectomy), permanent recurrent nerve lesion in 1.6% of patients (1% of nerves at risk). PAPILLARY CARCINOMA: No tumour-related death and no serious recurrence occurred in the low risk-group (TNM stages I and II) (n = 112 (84%)), including a T4-, N1 , M1-status in 9%, 20% and 3% of patients, respectively. 4/112 patients (3.6%) developed a recurrence (3 nodal, 1 contralateral, following 131I ablation in 2 instances). Only one (1%) instance of a nodal recurrence occurred in N0-tumors (n = 97). In TNM stages III and IV (high risk) patients (with T4-, N1-, M1-status in 79%, 58%, and 8% respectively), residual and recurrent disease occurred in 7 (33%) patients, leading to 6 (29%) tumor-related deaths. Follicular carcinoma: One 74-year old patient (1.6%) died from minimally invasive follicular carcinoma (n = 54 (51%); mean age 48 years). In 44 (81%) patients treatment did not include remnant ablation. 7 (13%) patients died from widely invasive follicular carcinoma (n = 53 (49%); mean age 64 years) and 3 (6%) further patients are alive with a serious recurrence. No curable recurrence was observed in follicular carcinoma. CONCLUSIONS: The decrease in goiter endemicity during the last decades in Switzerland paralleled a decrease in the incidence of grossly invasive follicular carcinoma over the 25 year period of the study. Following selective treatment, low risk TNM stage I and II-patients with papillary carcinoma had a tumor-related death rate of 0% and a low (3.6%) recurrence rate. N1-status represents a risk factor for nodal recurrence (even with remnant ablation). Elective vs. selective lymphadenectomy lead to slight stage migration but it presented no advantage in terms of recurrence and death which were rare events. No death occurred in the subgroups of minimally invasive follicular carcinoma of young (< 45) patients (41%) and in the patients without vascular invasion (28%), even without remnant ablation in most instances. No curable recurrence occurred in foll PMID- 11109917 TI - [Results of neurorehabilitation. An outcome study 20 months after stroke]. AB - 65 stroke survivors who were discharged home after completing an in-patient rehabilitation program were evaluated at home 20 months post-stroke by physiotherapists. 59 patients (91%) still lived in the community. Functional abilities remained stable with only 11% deteriorating and 25% improving in basal activities of daily living (BADL). 58% of patients needed assistance for at least one BADL and 46% showed signs of impaired cognition. Falls occurred in more than half of patients. Rehospitalisation was common (31%). Aside from living partners, care was provided by relatives in 58% and by home services in 46%. 25% of patients attended day care. Nearly half of patients still received rehabilitative therapy, especially if marked initial deficit was present. In conclusion, 20 months post stroke the majority of survivors who have completed rehabilitation successfully experience persistent limitations but remain in a stable functional status. This seems to be true for more severely disabled patients, too, if rehabilitative therapies, home services and day care are consequently provided. PMID- 11109918 TI - [Sports-induced macrohematuria]. PMID- 11109919 TI - [Why even a discussion about rationing?]. PMID- 11109920 TI - [Legal questions on rationing]. PMID- 11109921 TI - [Problems in rationing]. PMID- 11109922 TI - [From the viewpoint of the general practitioner]. PMID- 11109923 TI - [Rationing in surgery and high-tech medicine]. AB - There has been much debate about cost-cutting in health care. In this context, the term rationing means that acceptable services and measures with proven benefit are subject to limited allocation (or complete refusal thereof) due to a shortage of resources. After an introductory discussion about the "terminology" of rationing, achievements in premium health care will be critically elucidated in this regard. Chapter 2 focuses on the question of whether age is a valid criterion for the rationing of surgical care. Chapter 3 critically analyses the "societal solution" to the rationing problem. The way to solve society's tendency to become insatiable (pleonexia) is the same as the way to solve the shortage of resources, i.e. by "circumspection" or "moderation" on the part of each individual (sophrosyne). PMID- 11109924 TI - [Differential diagnostic considerations in CK-MB level increase]. AB - We present two patients who were hospitalized due to thrombo-embolic disease. Both patients had an increase in total creatine kinase activity with the creatine kinase MB fraction value exceeding the total creatine kinase activity. We determined that the high values for creatine kinase MB fraction in the immunoinhibition assay were due to the existence of macro creatine kinase type I in one patient and a highly elevated creatinine kinase BB fraction in the other patient. The patient with macro CK type I had ulcerative colitis and the other patient with elevated CK BB fraction was diagnosed with prostatic carcinoma. PMID- 11109925 TI - [Nicotine and smoking cessation]. PMID- 11109926 TI - 25th Congress of the International Society of Urology. Singapore, 29 October-2 November 2000. Abstracts. PMID- 11109927 TI - American Society of Hematology 42nd annual meeting, Part 1. December 1-5, 2000. San Francisco, California, USA. Abstracts. PMID- 11109928 TI - American Society of Hematology 42nd annual meeting, Part 2. December 1-5, 2000. San Francisco, California, USA. Abstracts. PMID- 11109929 TI - Management of unstable angina. Guidelines--2000. The National Heart Foundation of Australia, The Cardiac Society of Australia and New Zealand. PMID- 11109930 TI - RSNA 2000. Radiological Society of North America 86th annual meeting. November 26 December 1, 2000. Chicago, Illinois, USA. Abstracts. PMID- 11109931 TI - 14th International Bone Densitometry Workshop. September 3-7, 2000. Warnemunde, Germany. Abstracts. PMID- 11109932 TI - Proceedings of the British Pharmacological Society Meeting. 12-14 July 2000. Abstracts. PMID- 11109934 TI - 6th European Congress of Pharmaceutical Sciences, EUFEPS 2000. September 16-19, 2000. Budapest, Hungary. Abstracts. PMID- 11109933 TI - 5th Three-Country Symposium on Biological Psychiatry. 5-8 October 2000, Vienna, Austria. Abstracts. PMID- 11109935 TI - Society of General Internal Medicine 23rd annual meeting. Boston, Massachusetts, USA. May 4-6, 2000. Abstracts. PMID- 11109936 TI - 3rd International Congress on Phytomedicine. October 11-13, 2000. Munich, Germany. Abstracts. PMID- 11109937 TI - Methods to evaluate services. PMID- 11109938 TI - Complex service evaluation. AB - Services represent the practical manifestation of the synthesis of research knowledge and real world factors. In order to develop the evaluation of complex services, there needs to be a consensus about what a complex service is. I suggest that it is a system for the supplying of a public need. Whilst there is strong academic and policy support for a systems-based approach, there is only limited understanding in the clinical and managerial community and limited skills within the health and social care research community on systems methodologies. Evaluation of complex systems will probably need an integration of existing evaluative methods with a soft systems approach. PMID- 11109939 TI - Evaluating action research. PMID- 11109940 TI - [Promotion of breast feeding: it's the role of pediatricians...]. PMID- 11109941 TI - [Does neonatal screening of cystic fibrosis affect outcome? Comparative study of two cohorts in Britanny and Loire-Atlantique with follow-up after ten years]. AB - Neonatal screening for cystic fibrosis was started in Brittany in 1989 but not in the adjacent department of Loire-Atlantique. This study compares the outcome from the children of both populations nine years after the beginning of the screening. Those children were seen in different centers but with the same following guidelines. POPULATION AND METHODS: All children with cystic fibrosis born between 01/01/89 and 31/12/97 in Brittany and the Loire-Atlantique, excluding the meconium ileus, were compared for their initial characteristics and their outcome after nine years of follow-up. RESULTS: There was no significant difference between both populations for sex ratio, gestational age, birth biometry, percentage of homozygotes delta F508, and mean age of children. Age at diagnosis was lower in Brittany (37 vs 372 days, P < 10(-7)), as was the delay for starting pancreatic supplementation (1.5 vs 14.3 months, P < 10(-7)). Percentage of children hospitalized at least once was higher in Loire-Atlantique (84.4 vs 40.3%, P < 10(-4)). There was no significant difference for colonization with Pseudomonas aeruginosa. Z-scores for weight and height were better in Brittany, as were Shwachman's and Brasfield's scores. CONCLUSION: The homogeneity of both populations and their follow-up points out that even if the numbers of children are small and the study is retrospective, some benefits of neonatal screening appear, which are already found in other countries where it is partly practiced. This leads us recommend its general use in our populations, which should be associated with the follow-up of the screened children in cystic fibrosis centers to achieve the most of its benefits. PMID- 11109942 TI - [Severe malaria in children in intensive care. National survey 1990-1995]. AB - BACKGROUND: Severe malaria is a frequent complication of Plasmodium falciparum infections. More than one million children die of malaria each year. MATERIAL AND METHODS: A French survey was carried out on 15 cases admitted to pediatric intensive care units between 1990 and 1995. The aim of this work was to evaluate the occurrence, mortality, morbidity and treatment of severe malaria in French intensive care units. RESULTS: All cases were imported from Africa except one case of airport malaria. Diagnosis of many of these cases was delayed. All cases were treated with quinine, and five children received a loading dose. One child died and one has neurological sequelae. DISCUSSION: Despite improvement in management, the prognosis of severe malaria remains poor. With reference to the literature, we propose management of severe malaria, emphasizing the necessity of a rapid effect with a loading dose of quinine. PMID- 11109943 TI - [Gravely ill newborns (excluding greatly premature) in 1986 and 1995 in three deparments of Pays de la Loire]. AB - AIM: Evaluation of the progress in the treatment of seriously ill newborn infants under hospital care over the nine-year period from 1986 to 1995 in three departments of the Pays de la Loire region in France. POPULATION AND METHODS: The category of seriously ill newborn infants included only infants born after at least 32 weeks of gestation and having a Cullen severity score higher than 2. The two groups of seriously ill newborn infants at risk in 1986 and 1995 were first compared to a control group of healthy newborn babies delivered during the same years. The two groups were then compared with each other. RESULTS: In 1986 and 1995, the numbers of births were respectively 32,876 and 29,446, and the numbers of seriously ill newborn infants under hospital care were respectively 307 and 245. However, between 1986 and 1995 the risk factors as well as the causes of morbidity had changed. The average period of hospitalization decreased by five days. The mortality rate dropped from 0.16% to 0.09% (P < 0.05) and the number of serious complications decreased from 0.07% to 0.03% (P < 0.05). CONCLUSION: The improvement in the care of seriously ill newborn infants between the two reference periods, 1986 and 1995, may be attributed not only to technical progress but also to a better organization of the perinatal care in our region. PMID- 11109944 TI - [Sternocleidomastroid inflammatory pseudotumors of muscle in children]. AB - PATIENTS AND METHODS: The files of 12 children presenting with a sternocleidomastoid tumor of infancy at the Timone Children's Hospital in Marseille between 1990 and 1999 were retrospectively studied. All of them underwent physical and ultrasonographic examination. RESULTS: The mass was firm, within the lower two-thirds of the sternocleidomastoid muscle. Ultrasonographic examination showed a soft tissue mass independent of the great vessels. Biopsy revealed a benign fibrous lesion. Eleven children were treated with stretching exercises and a surgical therapy was performed for one in whom physical therapy was unsuccessful. CONCLUSION: The sternocleidomastoid tumor of infancy is a lateral neck mass affecting children between the ages of two and four weeks. Most of the time it spontaneously resolves in four to eight months. It may be associated with congenital muscular torticollis and requires physical therapy. Surgery should be reserved for children who have failed with this treatment. PMID- 11109945 TI - [Personal sports training in the management of obese boys aged 12 to 16 years]. AB - OBJECTIVE: Estimation of both physical and psychological effects of an adapted physical training on children undergoing an obesity treatment. MATERIAL AND METHODS: The survey was carried out on 36 obese boys (ages = 12-16 years) who stayed in the medical center for at least four months. Eighteen of them were trained with the SELF method (the SELF-training is global, progressive, adapted to each boy, controlled and takes place within a ten-week period with five sessions a fortnight, each session lasting 30 to 40 minutes). The parameters that were studied concerned auxology, breathing function exploration, aerobic and anaerobic capacities, muscle strength and psychomotor qualities; the subjective effects of the training were estimated with a questionnaire about life quality, and the hand test. At inclusion the results were reported to a standard kind of population. At the end of the training the results of the 18 boys that were trained were compared to those of the 18 controls. RESULTS: Compared to a standard population, the obese children' aerobic capacity is diminished for the maximum power but is identical in absolute value for the VO2 max; their anaerobic capacities, muscle strength and psychomotor capacities are lower and their psyche is affected by the disease. After a three-month training period and after comparison with the 'control' group, there can be noticed a significant improvement in the psychomotor capacities, a major tendency for the improvement of the aerobic capacities and very positive effects on the psyche. CONCLUSION: SELF-training in association with dietetics appears to be very useful in the therapeutic care of obese children. For the follow-up at home it would need to be registered within the domain of physiotherapy. PMID- 11109946 TI - [Perianal streptococcal dermatitis]. AB - BACKGROUND: Pediatric perianal streptococcal dermatitis (PSD) is a well-defined clinical entity. However, its highly uniform presentation remains surprisingly unrecognized by many practitioners 33 years after its first description. CASE REPORT: A seven-year-old girl had a three-week history of perianal and vulva redness with well-defined margins. Functional symptoms associated perirectal tenderness and pain during defecation, which was responsible for constipation. At onset she also presented with a sore throat, which resolved spontaneously, and she had been complaining for a few days about a perioral impetigo. She received mycostatin unsuccessfully for an alleged candidiasis. Positive cultures for group A beta-hemolytic streptococci from both perirectal and perioral swabs confirmed the diagnosis of PSD. Therapy with amoxicillin (50 mg/kg/d) was prescribed for ten days. Perianal lesions were cleared by day 2. CONCLUSION: Since PSD can masquerade as candidiasis, psoriasis, seborrheic dermatitis, inflammatory bowel disease or even sexual abuse, it remains an underdiagnosed entity. This situation leads to delayed diagnosis and treatment which in turn might increase the frequency of secondary complications related to streptococcal infections (i.e., post-streptococcal acute nephritis and rheumatism, guttate psoriasis, etc.). PMID- 11109947 TI - [Meningitis and interferon-alpha in the cereprospinal fluid. What diagnostic and therapeutic approach? A case report]. AB - BACKGROUND: In meningitis without germs, the existence of an inflammatory syndrome leads toward a bacterial etiology while the detection of interferon alpha (IFN-alpha) in the cerebrospinal fluid (CSF) argues for a viral meningitis. The coexistence of the inflammatory syndrome and the presence of IFN-alpha in the CSF makes this differentiation difficult. The reported case yields the picture and begs the question on the diagnostic approach and the required therapeutic attitude. CASE REPORT: A six-week-old infant, exclusively breast-fed, was hospitalized for fever. The examination showed an important inflammatory syndrome and meningeal attempt with a cellularity at 94/mm3 with 53% polymorphonuclear neutrophils, contrasting with normal proteinorrhachia and glycorrhachia. The IFN alpha in the CSF was present at 4 UI/mL while the bacteriological culture and the viral search by PCR were negative. The clinical and biological worsening within the first 36 hours, in spite of the parenteral dispensation of a triple antibiotic therapy (amoxicillin, ceftriaxone, netilmicin), then a favorable clinical and biological response after adjunction of vancomycin, led toward a pneumococcal meningitis with reduced sensitivity to beta-lactams. The maternal antibiotic therapy by amoxicillin and its presence in the maternal milk favored the hypothesis of a decapitated bacterial meningitis. CONCLUSION: In the presence of a meningitis without germs, the coexistence of a sizable inflammatory syndrome and the detection of IFN-alpha in the CSF must be considered as an unusual phenomenon and motivate the pursuit of antibiotic therapy until viral identification. PMID- 11109948 TI - [Use of interferon in a case of hepatic hemangioma]. AB - Hepatic hemangioendothelioma can have a fatal outcome. After failure of classic therapies, and when surgery or arterial embolization are not possible, those benign tumours may benefit from interferon alpha treatment. CASE REPORT: We report a case of a 14-month-old infant who presented with hepatic hemangiomatosis and cardiac failure. The disease can neither be controlled by steroids nor by radiotherapy associated with digitalo-diuretic treatment. Due to the important vascular volume of the tumour, neither surgical care nor arterial embolization were possible. Thus this infant was treated for ten months by interferon alpha. The evolution was favourable: cardiac failure, calcifications and volume of the angiomatosis were regressive. CONCLUSION: Interferon alpha treatment helps to control cardiac failure and the course of hepatic hemangioma in childhood. PMID- 11109949 TI - [X-linked lymphoproliferative syndrome (XLP syndrome): from clinic to gene]. AB - X-linked lymphoproliferative syndrome (XLP, also known as Duncan's disease) is characterised by an extreme sensitivity to Epstein Barr virus (EBV), resulting in a complex phenotype manifested by severe or fatal mononucleosis, acquired hyogammaglobulinemia and malignant lymphoma. The gene responsible for XLP has recently been identified by a positional cloning and a functional cloning approach and encodes a small cytoplasmic protein involved in signal transduction of T and NK cells. The identification of the XLP gene will permit direct diagnosis of XLP in families with a single affected male. Recent progress in immunobiology and genetics of this primary immunodeficiency disease are presented. PMID- 11109950 TI - [Medical informatics (hospital information systems) in neonatology: reality and insufficiencies]. AB - The generalized implementation in France of hospital information systems (HIS) is often considered by the medical practitioners as a useless constraint. Nevertheless, they are now largely used by the administrative authorities for their economical evaluation of medical care. In neonatology HIS is applied to the hospitalized sick neonates as well as to the healthy newborn infants during their maternity hospital stay with their mother following birth. This paper focuses on the practical aspects and difficulties of the current French HIS in neonatology. PMID- 11109951 TI - [Physician or seer? Very early diagnosis and prognosis. The example of William's syndrome]. AB - Recent major breakthroughs in the field of genetics have allowed very early diagnosis of many genetic diseases. This is usually considered as beneficial progress. But is it that simple? Which information is given to the parents? How useful is it? And for whom? How will it influence the child's development? The case of three children with Williams' syndrome and the current knowledge on the cognitive and behavioural phenotype of this syndrome provide food for thought on these questions. Answers can be neither unique nor final. The purpose is not to deny present knowledge or tomorrow's discoveries, but rather to take time to think about the consequences of a very early diagnosis announcement, particularly the anticipation of the handicap that it carries. PMID- 11109952 TI - [Ethics and best practice in the consulting management of children with cleft lip and palate, and their parents]. AB - Surgical treatment is only one part of the management of the child with cleft lip and palate. This paper exclusively focuses on other important aspects of this management. This includes the information and psychological supports of the parents, whether the cleft lip is diagnosed prenatally or at birth, the practical aspects of the consultation within the team of the different specialists involved (surgeon, anesthetist, dentist, orthodontist, speech therapist, otorhinolaryngologist, geneticist, child psychologist). The school teacher must also be concerned at the beginning of the first school year. PMID- 11109953 TI - [Inguinal hernia in pediatric practice]. AB - Inguinal hernia is a most common pathology in paediatric practice. Diagnosis is usually easy and rarely requires paraclinic exams (sonography). The main risk is strangulation, which can be complicated by testicular or ovarian atrophy. This justifies early surgical treatment. Operative mortality rate is almost nil. There is a risk of recurrence and of occurrence of contralateral hernia. PMID- 11109954 TI - [Evaluation after 6 years: the pediatric subsection of the French National University Committee]. PMID- 11109955 TI - [Maternal-fetal infection with Gardnerella vaginalis]. PMID- 11109956 TI - [Is prevention fo maternal-fetal infection by antibiotic prophylaxis worth the risk?]. PMID- 11109957 TI - Comparative analysis of free-radical lipid peroxidation rate in poikilothermal vertebrates. PMID- 11109958 TI - The structure of the active center of beta-peptide membrane-bound methane monooxygenase (pMMO) from Methylococcus capsulatus bath. PMID- 11109959 TI - Characteristics of changes in the erythrocyte resistance to peroxidation hemolysis in white rats intoxicated with halothane and the effects of sodium thiosulfate under these conditions. PMID- 11109960 TI - Structural and temporal links between the components of humoral immunity. PMID- 11109962 TI - Hemolymph cholinesterase of the Pacific gastropod Neptunea eulimata: substrate inhibitor analysis. PMID- 11109961 TI - Digestion of oxidatively modified proteins by proteolytic enzymes from neutrophils of mice of different ages. PMID- 11109963 TI - Variability of the protein component of atmospheric aerosol above large forests in southwestern Siberia. PMID- 11109964 TI - Ribosomal protein TL5 of T. thermophilus is incorporated in the E. coli 50S ribosomal subunit. PMID- 11109965 TI - Specific features of albumin denaturation in physiological preparations exposed to thermal and radiation factors. PMID- 11109966 TI - Transcription factor STAT1 is bound to proteasomes in A-431 cells. PMID- 11109967 TI - Cytotoxic activities of conjugates of human transferrin and A subunits of plant toxins both in vitro and in vivo. PMID- 11109968 TI - Effect of T-helper 1 cytokines on secretion of T-helper 2 cytokines by term trophoblast cells in culture. AB - A successful pregnancy has been postulated to be the result of a discrete balance between T-helper 1 (Th1) and T-helper 2 (Th2) type cytokines involved in growth and development of the conceptus. The aim of the present study was to examine the effect of Th1 cytokines (interleukin-1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF alpha)) on the release of Th2 cytokines including IL-6 and IL-10 by trophoblast cells obtained from term placenta. Trophoblast cells isolated by enzymatic disaggregation and Percoll gradient fractionation were cultured in supplemented medium alone or with varying concentrations of the selected recombinant cytokines. After 48 h of incubation, samples of the culture supernatant were analyzed for the Th2 cytokines IL-6 and IL-10 using specific ELISA assays. Both IL-1 beta and TNF alpha had no effect on the cell number and viability as determined by MTT assay. IL-1 beta significantly stimulated trophoblast release of IL-6 in a dose-dependent manner (3.3-, 5.5-, 10.3- and 22.4-fold higher compared to the control at 10, 50, 100, 500 U IL-1 beta/ml respectively, p < 0.05). TNF alpha also stimulated release of IL-6 by these cells. However, the stimulation at lower concentrations was not very high and a significant (p < 0.05) stimulation was observed only at higher concentrations (1.1-, 1.3-, 2.6- and 5.9-fold higher at 500, 1000, 1500, 2000 U TNF alpha/ml respectively). In contrast, neither IL-1 beta or TNF alpha exerted any significant effect on IL-10 release by term trophoblast cells (p > 0.05). The results of this study provide evidence that production of Th2 cytokines might be under the control of different regulatory pathways. PMID- 11109969 TI - The use of a starting dose of recombinant follicle stimulating hormone for controlled ovarian hyperstimulation: a randomized pilot study. AB - The aim of this study was to compare the costs and effects of two different controlled ovarian hyperstimulation treatments: a starting dose of recombinant follicle stimulating hormone (FSH) followed by highly purified urinary FSH; or highly purified urinary FSH alone. Forty-six infertile patients, after being given luteal gonadotropin-releasing hormone (GnRH) agonist, were randomly assigned to the two stimulation protocols. During the ovarian stimulation regimen the patients underwent transvaginal ultrasonographic evaluation of follicular number and size. The retrieved oocytes were classified on the basis of the criteria of Acosta and colleagues. To study the impact of embryo quality on implantation, the embryos were graded morphologically before replacement. Pregnancy rates were ascertained and the costs of the two different protocols were analyzed. The number of days of FSH stimulation and the cost of gonadotropin treatment were similar in both groups. The number of follicles > 17 mm in size, the number of collected oocytes, and pregnancy rate per cycle were significantly higher in the group partially treated with recombinant gonadotropin. We conclude from these results that the use of recombinant FSH in the early phase of controlled ovarian hyperstimulation leads to significant improvements in pregnancy rate per cycle without increasing the costs of treatment. PMID- 11109970 TI - Human chorionic gonadotropin and intravaginal natural progesterone are equally effective for luteal phase support in IVF. AB - This prospective randomized study compared human chorionic gonadotropin (hCG) and micronized transvaginal progesterone for luteal support in 310 in vitro fertilization (IVF) patients treated with leuprolide acetate and gonadotropins in a long protocol, and showing normal ovarian response. Both treatment groups were homogeneous for age, BMI, stimulation treatment and ovarian response. Pregnancy rates per embryo transfer were not significantly different (33.1% for the hCG group versus 38.7% for the progesterone group). For IVF patients with a normal response to stimulation under pituitary suppression, the use of hCG or progesterone for luteal support does not seem to have any effect on pregnancy rate. The choice of luteal treatment must balance medical hazard and patient convenience, as both therapeutic regimens seem equally effective. PMID- 11109971 TI - Effects of low day 3 luteinizing hormone levels on in vitro fertilization treatment outcome. AB - In a previous study we demonstrated that women with day 3 luteinizing hormone (LH) values < 3 IU/l subjected to controlled ovarian hyperstimulation without pituitary desensitization responded with a lower number of follicles > 15 mm compared to women with a higher basal LH level. The aim of this study was to determine whether in patients with day 3 LH levels < 3 IU/l a further reduction of serum LH concentration by gonadotropin-releasing hormone (GnRH) analog impairs follicular response to follicle stimulating hormone (FSH) and treatment outcome in in vitro fertilization (IVF) cycles. For this purpose we retrospectively studied 249 consecutive women subjected to standard IVF treatment employing pituitary desensitization with buserelin and follicular stimulation with urinary highly purified FSH. The patients were divided into two groups according to their day 3 LH value. The first group (group A) showed day 3 LH levels < 3 IU/l and the second (group B) had day 3 LH levels > 3 IU/l. Group A and B patients did not show statistically significant differences in the ovarian response to FSH, nor in IVF treatment outcome, showing that in FSH treated GnRH analog suppressed cycles, the ovarian responsiveness and IVF outcome do not differ according to basal LH values. However, the high dosage of FSH we employed in group A and B patients could account, at least in part, for this result. Indeed, comparative evaluations with unsuppressed cycles (our previous study) strongly suggest that a reduced ovarian responsiveness to gonadotropins in patients with day 3 LH values < 3 IU/l should be considered in clinical practice. PMID- 11109972 TI - Impaired insulin action on granulosa-lutein cells in women with polycystic ovary syndrome and insulin resistance. AB - We studied the in vitro response to insulin of granulosa-lutein cells derived from patients with polycystic ovary syndrome (PCOS) and clinically defined insulin resistance. Insulin sensitivity was assessed by continuous infusion of glucose with model assessment test (CIGMA). Insulin resistant (PCOS-IR; n = 8), noninsulin resistant (PCOS-NIR; n = 9) patients with PCOS, and women with tubal factor infertility (TF; n = 8) underwent controlled ovarian stimulation with long term gonadotropin-releasing hormone (GnRH) agonist, recombinant follicle stimulating hormone (FSH), and in vitro fertilization. Primary cultures of granulosa-lutein cells were incubated with insulin (10, 100, 500 ng/ml) and/or luteinizing hormone (LH) (10, 100 ng/ml) in the presence of low density lipoprotein (100 micrograms/ml). The progesterone and lactate accumulation were measured in the culture medium. LH potently stimulated the progesterone secretion in all groups. Insulin alone had no effect on progesterone release in any of the groups, but stimulated lactate formation in the PCOS-NIR and TF groups. Insulin augmented the effect of LH on progesterone secretion selectively in the PCOS-NIR group. The expression of the insulin receptor was determined by Western blotting in separate cultures of granulosa-lutein cells, and showed receptor down regulation in the PCOS-IR patients. We infer that the in vitro effect of insulin on progesterone and lactate release by granulosa-lutein cells is impaired in insulin resistant PCOS patients. PMID- 11109973 TI - Hormonal and clinical effects of chronic gonadotropin-releasing hormone agonist treatment in polycystic ovary syndrome. AB - The aim of the study was to evaluate the hormonal (focusing on the urinary steroid profile) and clinical effects of chronic gonadotropin-releasing hormone (GnRH) agonist treatment in patients with polycystic ovary syndrome (PCOS) suffering from hirsutism. A long-acting GnRH agonist was administered for 6 months in eight PCOS patients. Hormonal effects were measured by determining serum luteinizing hormone (LH), follicle stimulating hormone (FSH), prolactin, testosterone and estradiol concentrations, and by profiling urinary steroids using capillary gas chromatography of 24-hour urine samples. To evaluate 5 alpha reductase enzyme activity, the ratios of androsterone to etiocholanolone and 5 alpha-tetrahydrocortisol to tetrahydrocortisol were calculated in urine samples. The ratio of androgen to cortisol metabolites was also determined before, and 3 and 6 months after therapy. LH and estradiol levels were suppressed significantly after the first injection and testosterone after the second injection of the GnRH agonist. Thus, serum testosterone was normalized. Ratios of urinary steroids reflecting 5 alpha-reductase enzyme activity (androsterone to etiocholanolone and 5 alpha-tetrahydrocortisol to tetrahydrocortisol) and the ratio of androgen to cortisol metabolites decreased significantly after 3 months of treatment. Degree of hirsutism, assessed by Ferriman-Gallwey score, diminished after 6 months, but not significantly. In conclusion, our data show that long-acting GnRH agonist treatment of PCOS patients is effective in reducing serum and urinary androgen levels, but it is not accompanied by an effective reduction in hirsutism during a 6-month treatment period. A longer or a combined treatment would be needed to achieve significant improvement in hirsutism. Gas chromatographic profiling of urinary steroids and the use of specific ratios of the excreted metabolites seems to be a sensitive tool both in the diagnosis of PCOS and in monitoring ovarian suppression. PMID- 11109974 TI - Six-month oral dehydroepiandrosterone supplementation in early and late postmenopause. AB - The adrenal production of the delta 5-androgens, dehydroepiandrosterone (DHEA) and its sulfate ester dehydroepiandrosterone sulfate (DHEAS), declines linearly with aging. The evidence that DHEA or DHEAS administration may alleviate some of the problems related to aging has opened new perspectives for clinical research. The present study aims to investigate the effects of a 6-month DHEA supplementation in early and late postmenopausal women, with normal or overweight body mass index (BMI), on the level of circulating steroids, sex hormone binding globulin (SHBG), beta-endorphin and gonadotropins, and on the adrenal gland response to dexamethasone suppression and adrenocorticotropic hormone (ACTH) stimulation. Early postmenopausal women (50-55 years) both normal weight (BMI 20 24, n = 9) and overweight (BMI 26-30, n = 9) and late postmenopausal women (60-65 years) both of normal weight and overweight, were treated with oral DHEA (50 mg/day). Circulating DHEA, DHEAS, 17-OH pregnenolone, progesterone, 17-OH progesterone, allopregnenolone, androstenedione, testosterone, dihydrotestosterone, estrone, estradiol, SHBG, cortisol, luteinizing hormone, follicle stimulating hormone and beta-endorphin levels were evaluated monthly and a Kupperman score was performed. The product/precursor ratios of adrenal steroid levels were used to assess the relative activities of the adrenal cortex enzymes. Before and after 3 and 6 months of therapy, each women underwent an ACTH stimulating test (10 micrograms i.v. in bolus) after dexamethasone administration (0.5 mg p.o.) to evaluate the response of cortisol, DHEA, DHEAS, androstenedione, 17-OH pregnenolone, allopregnanolone, progesterone and 17-OH progesterone. The between-group differences observed before treatment disappeared during DHEA administration. Levels of 17-OH pregnenolone remained constant during the 6 months. Levels of DHEA, DHEAS, androstenedione, testosterone and dihydrotestosterone increased progressively from the first month of treatment. Levels of estradiol and estrone significantly increased after the first/second month of treatment. Levels of SHBG significantly decreased from the second month of treatment only in overweight late postmenopausal women, while the other groups showed constant levels. Progesterone levels remained constant in all groups, while 17-OH progesterone levels showed a slight but significant increase in all groups. Allopregnanolone and plasma beta-endorphin levels increased progressively and significantly in the four groups, reaching values three times higher than baseline. Levels of cortisol and gonadotropins progressively decreased in all groups. The product/precursor ratios of adrenal steroid levels at the sixth month were used to assess the relative activities of the adrenal cortex enzymes and were compared to those found before therapy. The 17,20-desmolase, sulfatase and/or sulfotransferase, 17,20-lyase and 5 alpha-reductase activities significantly increased, while the 3 beta-hydroxysteroid-oxidoreductase activity did not vary. On the contrary, the 11-hydroxylase and/or 21-hydroxylase activities showed a significant decrease after 6 months of treatment. In basal conditions, dexamethasone significantly suppressed all the adrenal steroids and this suppression was greater after 3 and 6 months of treatment for DHEA, DHEAS and allopregnanolone, while it remained unchanged for other steroids. Before treatment, ACTH stimulus induced a significant response in all parameters; after the treatment, it prompted a greater response in delta 5- and delta 4-androgens, progesterone and 17-OH progesterone, while cortisol responded less in both younger and older normal-weight women. The endometrial thickness did not show significant modifications in any of the groups of postmenopausal women during the 6 months of treatment. Treatment with DHEA was associated with a progressive improvement of the Kupperman score in all groups, with major effects on the vasomotor symptoms in PMID- 11109975 TI - Bone mineral density in women with idiopathic hirsutism. AB - Idiopathic hirsutism is relatively uncommon, affecting approximately 6% of hirsute women. In the present study we compared the bone mineral density (BMD) of women with idiopathic hirsutism with controls. A group of 20 women diagnosed with idiopathic hirsutism was evaluated with respect to BMD and the findings were compared to those of a control group consisting of 10 normal women. Hirsutism was graded according to the Ferriman-Gallwey score and threshold was considered to be a score more than 4. There was no statistically significant difference with respect to patients' mean age, BMI and body fat composition. Serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), androstenedione, testosterone, free testosterone, 17 alpha-hydroxyprogesterone, dehydroepiandrosterone sulfate (DHEAS), sex hormone binding globulin (SHBG) and estradiol were assessed in both groups and no statistically significant differences were found. There was no statistically significant difference with respect to bone turnover--which was evaluated by determining the serum levels of parathyroid hormone, calcium, alkaline phosphatase and osteocalcin as well as the urinary secretion of calcium and hydroxyproline, corrected for the creatinine values--between the two groups. Statistical analysis was performed using the t test for unpaired data to compare age, BMD and biochemical data, and Wilcoxon's rank test was used to compare physical activity and calcium intake. Statistical significance was defined as p < 0.05. The BMD at the level of the 2nd to 4th lumbar vertebrae (L2-L4) and the total BMD were higher in women with idiopathic hirsutism compared to those in the control group, suggesting a possible direct effect of androgens on the osseous tissue of hirsute women. PMID- 11109976 TI - The effect of combined oral contraception with or without spironolactone on bone mineral density of hyperandrogenic women. AB - We studied the effect of treatment with combined oral contraception (COC) with or without spironolactone on the bone mineral density (BMD) of hyperandrogenic women. A group of 22 women (group 1) was treated with ethinylestradiol plus desogestrel for 21 days each month for 12 months, while another group of 21 patients (group 2) was treated with ethinylestradiol and desogestrel for 21 days each month plus spironolactone daily for 12 months. There was no statistically significant difference with respect to mean age, body mass index (BMI) and BMD between the two groups of patients before the treatment. Serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), androstenedione, total testosterone, dehydroepiandrosterone sulfate (DHEAS), sex hormone binding globulin (SHBG), prolactin and estradiol were assessed in both groups and no statistically significant difference was found before treatment. Nor was there any statistically significant difference in bone turnover between the two groups. Statistical analysis was performed using the Student's t test for unpaired data to compare age, BMD and biochemical data, and statistical significance was defined as p < 0.05. The BMD before and after 12 months of treatment showed no statistically significant difference between the patients of group 1 and those of group 2, suggesting that both ethinylestradiol plus desogestrel, and ethinylestradiol and desogestrel plus spironolactone daily for 12 months at the given doses do not affect the BMD of the treated women, while the addition of spironolactone improves the efficacy of hirsutism treatment. PMID- 11109977 TI - Serum osteocalcin and urinary crosslaps are suitable markers of bone turnover in response to short-term hormone replacement therapy. AB - In this study we evaluated the effect of short-term hormone replacement therapy (HRT) on bone formation (serum osteocalcin) and resorption markers (urinary type I collagen peptides (crosslaps), urinary total free pyridinoline (TPYRI) and urinary free deoxypyridinoline (DPYRI)) as well as female sex hormones (serum estradiol, follicle stimulating hormone (FSH) and luteinizing hormone (LH)) in a group of early postmenopausal women with severe estrogen deficiency. The 46 healthy postmenopausal women with serum estradiol levels < 10 ng/l were subsequently divided into two groups, according to their compliance with HRT. In the group taking HRT significant changes from baseline values could be observed in estradiol, FSH, urinary crosslaps and serum osteocalcin levels after 6 months, whereas no changes could be observed in LH, TPYRI and DPYRI from baseline values. In the group which refused HRT all values were increased relative to baseline values, indicating increased bone turnover. Serum osteocalcin and urinary crosslaps were significantly decreased in women taking HRT in comparison to the group refusing HRT. After 6 months the treated patients showed a decrease in urinary crosslaps of 42% (SD 12%) and in serum osteocalcin of 24% (SD 6%) in comparison with baseline values. In patients who refused HRT, urinary crosslaps were increased by 43% (SD 20%) and serum osteocalcin levels decreased by 2% (SD 9%) compared to baseline values. In postmenopausal women suffering from severe estrogen deficiency (estradiol < 10 ng/l) serum osteocalcin and urinary crosslaps are significantly increased, indicating a clear correlation between estrogen deficiency and an increase in bone resorption as well as bone formation. The recommended HRT dose was sufficient to reduce the rate of bone turnover to premenopausal values. Serum osteocalcin and urinary crosslaps are suitable candidates not only for the assessment of a high postmenopausal bone turnover, but also for monitoring the response to and for verifying the actual intake of HRT or other antiresorptive treatment. PMID- 11109978 TI - Comparison of the biochemical effects of testosterone and estrogen on bone markers in surgically menopausal women. AB - Twenty-five women with a previous total abdominal hysterectomy with bilateral salpingo-oophorectomy (TAH BSO) were given estradiol 50 mg implants at baseline, followed at 16 weeks with the combination of estradiol 50 mg and testosterone 100 mg. Blood samples were taken at 8-weekly intervals over 32 weeks. Serum levels of estradiol, testosterone, sex hormone binding globulin (SHBG) and agents involved in skeletal growth (growth hormone (GH), insulin-like growth factor 1 (IGF-1), carboxy terminal pro-peptide of type 1 pro-collagen (PICP; a bone formation marker) and cross-linked carboxy terminal telopeptide (ICTP; a marker of bone resorption)) were measured. Serum PICP levels increased significantly after estradiol alone (p = 0.0032) but the addition of testosterone had no significant effects on bone markers GH and IGF-1. These biochemical changes confirm previous studies, which found that the addition of testosterone did not augment the effect of estradiol implants on bone mineral density. Although physiological hormone replacement therapy in oophorectomized women would include replacement of both estradiol and testosterone, this may not to be necessary for prevention of osteoporosis where adequate serum estradiol levels are reached. PMID- 11109979 TI - Altered sensitivity to anti-endometriosis medicines in an adenomyosis patient with thyroid dysfunction. AB - We report the rare case of a patient with adenomyosis who showed thyroid dysfunction of unknown etiology during treatment for endometriosis. She responded to nafarelin acetate and danazol when she was euthyroid but not to buserelin acetate when she was hyperthyroid. This suggests that the response to anti endometriotic medicine in our patient was altered by impaired thyroid function and that this could occur in other such patients. PMID- 11109981 TI - The Tri-Joint Congress 2000. Canadian Association of Occupational Therapists, Canadian Physiotherapy Association, Canadian Association of Speech-Language Pathologists and Audiologists. Toronto, Canada, May 24-27, 2000. Abstracts. PMID- 11109980 TI - Polycystic ovary syndrome: an endocrine and metabolic disease. AB - Polycystic ovary syndrome (PCOS) is a common disorder occurring during female reproductive years, characterized by a number of heterogeneous clinical and biochemical features. Clinical presentation is characterized by hirsutism, acne, androgen-dependent alopecia, menstrual dysfunction, infertility and ultrasonographically-documented cystic ovaries. Tonic elevation of luteinizing hormone (LH) secretion is a regular feature of PCOS. Abnormal secretion of estrogen and high serum levels of free testosterone are also present. In addition to these endocrine abnormalities, specific metabolic alterations, such as hyperinsulinemia and insulin resistance are more frequent. Metabolic derangements associated with PCOS may predispose the patient to a range of diseases with attendant morbidity and mortality risks, so it is important to consider the syndrome in terms of both endocrine and metabolic aspects, achieving a correct hormone equilibrium and preventing metabolic alterations. PMID- 11109982 TI - 7th Annual conference of the International Society for Quality of Life Research. 29-31 October 2000. Vancouver, Canada. Abstracts. PMID- 11109983 TI - [Bibliographic databases. Medline via PubMed]. PMID- 11109984 TI - Innervation of the pigeon oviduct: correlation of NADPH diaphorase with acetylcholinesterase, tyrosine hydroxylase, and neuropeptides. AB - The motility of the avian oviduct is controlled by hormones and neurons, but little is microscopically known about a neural network in the oviduct. The present study was investigated to determine the distribution of nitric oxide synthesizing neurons in the oviduct of the pigeon by histochemistry for nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d). The NADPH-d reaction was seen in the neurons and fibers. NADPH-d neurons were mainly distributed around the arterioles of the intermuscular tissue in the upper oviduct (infundibulum, magnum, and isthmus); in addition, NADPH-d neurons were also seen in the smooth muscle layers and lamina propria in the lower oviduct (uterus and vagina). NADPH-d neurons were found singly or in small groups of two eight cell bodies. The number of NADPH-d neurons was smallest in the infundibulum, gradually increased toward the vagina. NADPH-d was also shown to be strongly positive in many neurons in the ganglia of the vaginal adventitia. Bundles of NADPH-d fibers ran in the smooth muscle layer, surrounded blood vessels, or connected with small groups of NADPH-d neurons by forming strands. Thin fibers branched from these bundles and constituted a finer network in the smooth muscle layer and lamina propria. Acetylcholinesterase staining in neurons and fibers showed a similar pattern of NADPH-d distribution in the oviduct. By double staining, 70 approximately 77% of neurons showed colocalization of NADPH-d and acetylcholinesterase in the uterus and vagina. Tyrosine hydroxylase immunoreactivity stained only nerve fibers and were distributed largely around blood vessels in the oviduct. Nerve fibers immunoreactive for calcitonin-gene related peptide, galanin, methionine-enkephalin, substance P, or vasoactive intestinal peptide were found sparsely in the oviduct. These results demonstrate that nitrergic neurons make up a large subpopulation of intrinsic neurons that are closely associated with a cholinergic system in the pigeon oviduct, thus suggesting that nitric oxide and acetylcholine could be used to modify the relaxation of the avian oviduct. PMID- 11109985 TI - Reduced hindbrain and enteric neuronal response to intestinal oleate in rats maintained on high-fat diet. AB - Rats maintained on a high-fat diet (HF) reduce their food intake less in response to intestinal infusion of oleic acid than rats maintained on a low-fat diet (LF). Inhibition of gastric emptying by intestinal infusion of oleate also is attenuated in rats fed a high-fat diet. It is well documented that intestinal oleate reduces food intake and inhibits gastric emptying via vagal sensory neurons. In addition, activation of intrinsic myenteric neurons participates in oleate-induced changes in gastrointestinal motility. To determine whether diminished behavioral and gastric reflex responses to intestinal oleate infusion is accompanied by reduced vagal sensory and myenteric neuronal activation, we examined expression of Fos-like immunoreactivity (Fos-li) in the dorsal hindbrains and the small intestinal enteric plexuses of rats maintained on HF or LF, following, intraintestinal infusion of oleate (0.06 kcal/ml) or the oligosaccharide, maltotriose (0.26 kcal/ml). Following oleate infusion there was a dramatic increase in the number of Fos-li nuclei in the NTS and AP of LF rats but not in HF rats. There also were significantly more Fos-li neuronal nuclei in the upper small intestinal submucosal and myenteric plexuses of the LF rats than the HF rats. In contrast to the effects of oleate infusion, maltotriose infusion significantly and similarly increased Fos-li nuclei in the hindbrains of both LF and HF rats. The results indicate that adaptation to high-fat diet selectively reduces vagal and enteric neuronal sensitivity to intestinal oleate and suggests that reduced sensitivity to the satiation and gastric inhibitory effects of oleate in high-fat fed rats may be mediated by a selective reduction in the neuronal response to intestinal stimulation by fatty acid. PMID- 11109986 TI - Respiratory activity in neonatal rats. AB - In neonatal animals in vitro preparations have been employed widely to study the central control of respiration. These preparations have limitations in that reflex afferent inputs and kinesiological studies cannot be performed. Here, we describe an alternative in situ experimental model for studying both peripheral and central control of the respiratory system in neonatal rats. Using technology based on adult mammals, we introduce an intra-arterially perfused working heart brainstem preparation (WHBP) that permits studies on eupnoeic respiration in neonatal rats from within a few hours of birth. Using this preparation we demonstrate a three-phase respiratory rhythm as revealed by the activity in phrenic and recurrent laryngeal motor nerves, the respiratory modulation of laryngeal resistance and the firing patterns of respiratory neurones recorded from the ventrolateral medulla. We conclude that the neonatal rat WHBP is an in situ preparation because it produces a respiratory rhythm similar to that of adult in vivo mammal preparations but distinct from in vitro preparations. PMID- 11109987 TI - Differential distribution of ionic channels and muscarinic receptors at the cat tracheal neuromuscular junction. AB - The ionic and pharmacological properties of atropine-sensitive excitatory junction potentials (EJPs) were investigated by the double sucrose gap and microelectrode membrane potential recording methods, and compared with those of inward currents evoked by carbachol (I(CCh)), in cat tracheal smooth muscle. A single and repetitive field stimulation (10-30 V, 50 micros, 20 Hz) evoked atropine-inhibitable EJPs and associated twitch-like contractions. Reduction in external Na+ concentration strongly, but in the external Cl- concentration, decreased the EJP amplitude after 5 min superfusion, although prolonged exposure to low Cl- solutions attenuated the EJPs modestly. Chloride channel blockers such as 9-AC and niflumic acid (each 100 microM), at concentrations high enough to inhibit I(CCh) almost completely, failed to abolish the EJPs. Pirenzepine, AF DX116 and 4-DAMP all effectively inhibited the EJPs at their concentrations to block respective muscarinic receptor subtypes relatively specifically, while depletion of internal stores by 10 mM caffeine and/or 3 microM ryanodine caused only a partial decrease in the EJP amplitude. These properties are considerably different from those of I(CCh) which is activated exclusively through activation of the M3 receptor/IP3-mediated Ca2+ release pathway and reflects mostly a Ca2+ dependent Cl- current, and suggests the differential distribution of muscarinic receptors and ionic channels inside and outside of the cholinergic neuromuscular junction of this muscle. PMID- 11109988 TI - Three types of putative presympathetic neurons in the rostral ventrolateral medulla studied with rat brainstem-spinal cord preparation. AB - To study the electrophysiological properties of presympathetic neurons in the rostral ventrolateral medulla (RVLM), intracellular recordings were performed by the whole-cell patch-clamp technique. We utilized the neonatal rat brainstem spinal cord preparation, in which the sympathetic neuronal network is thought to be preserved, unlike in slice preparation. In response to stimulation in the ipsilateral Th2 spinal segment including intermediolateral cell column (IML), 33 of 151 non-respiratory RVLM neurons showed antidromic action potentials with a constant latency of 45 ms, and can be considered as presympathetic neurons. We classified and characterized the RVLM presympathetic neurons into three types: 'regularly firing neurons (n=7)', which showed ramp depolarization and frequent action potentials (4.2+/-0.9 spikes/s) with rare excitatory postsynaptic potentials (EPSPs); 'irregularly firing neurons (n=21)', which exhibited many EPSPs that modulated the firing rate; and 'silent-type neurons (n=5)', which discharged action potentials only during current-induced depolarization. Lucifer Yellow staining showed that the irregularly firing neurons were significantly larger and had more dendrites than the regularly firing neurons. All regularly firing neurons retained their discharges during low-Ca2+ -high-Mg2+ superfusion that blocks synaptic input, whereas the discharges in 11 of 16 irregularly firing neurons were abolished, suggesting that the regularly firing neurons discharged independently of synaptic input. Seven of 31 RVLM neurons were hyperpolarized by stimulation of vagal afferent nerves. In summary, three types of RVLM presympathetic neurons were characterized by the patch-clamp technique in the brainstem-spinal cord preparation, in which the connection was preserved from vagal afferent to the Th2 spinal segment through the RVLM. Since antidromic action potentials were demonstrated by stimulation in the Th2 spinal segment in 33 neurons of all three types, all types of RVLM neurons constitute a part of the sympathetic neuronal network. PMID- 11109989 TI - Sympathetic and parasympathetic interaction in vascular and secretory control of salivary glands in anaesthetised dogs. AB - The present study was undertaken to examine sympathetic-parasympathetic interactions in the regulation of salivary gland function, with special reference to the possible role of the sympathetic cotransmitter neuropeptide Y (NPY). In dogs anaesthetised with pentobarbitone, electrical stimulation of the parasympathetic nerve to the submandibular gland evoked an increase in glandular blood flow and salivary secretion. Sympathetic nerve stimulation evoked a significant prolonged attenuation of vasodilator and secretory responses to subsequent parasympathetic stimulation. This attenuation was not significantly altered by alpha- and beta-adrenoceptor blockade. Systemic administration of the sympathetic cotransmitter, NPY, mimicked the effect of the sympathetic stimulation by significantly attenuating vasodilatation and salivary secretion. The NPY Y1 receptor agonist, [Leu31, Pro34]NPY and the specific NPY Y2 receptor agonist N-acetyl[Leu28, Leu31]NPY 24-36 both significantly attenuated the vasodilatation and salivary secretion evoked by stimulation of the parasympathetic nerve. The NPY Y1 receptor antagonist, GR231118 significantly antagonised the attenuation of vasodilatation caused by both sympathetic stimulation and the NPY Y1 receptor agonist. GR231118 also inhibited the pressor response of NPY. Intra-arterial injection of methacholine and stimulation of the parasympathetic nerve both caused local vasodilatation in the gland which was significantly attenuated by pretreatment with sympathetic stimulation or the NPY Y1 agonist. The NPY Y2-specific agonist did not attenuate methacholine-induced vasodilatation but did attenuate vasodilatation evoked by parasympathetic stimulation. The results indicate that NPY as a sympathetic cotransmitter may have a role in the regulation of vascular secretory function of salivary glands. PMID- 11109990 TI - Activation of GABA receptors in the NTS of awake rats reduces the gain of baroreflex bradycardia. AB - In the present study we evaluated the effects of bilateral microinjection of muscimol (a GABA(A) receptor agonist) and baclofen (a GABA(B) receptor agonist) into the lateral commissural nucleus tractus solitarii (NTS) of awake rats on the gain of the baroreflex (BG) activated by a short duration (10-15 s) infusion of phenylephrine (Phe, 2.5 microg/0.05 ml, i.v.). Microinjection of muscimol (50 pmol/50 nl, n=8) into the NTS produced a significant increase in baseline mean arterial pressure ((MAP) 122+/-6 vs. 101+/-2 mmHg), no changes in baseline heart rate (HR) and a reduction in BG (-1.59+/-0. 1 vs. -0.69+/-0.1 beats/mmHg). Microinjection of baclofen (6.25 pmol/50 nl, n=6) into the NTS also produced a significant increase in baseline MAP (138+/-5 vs. 103+/-2 mmHg), no changes in baseline HR and a reduction in BG (-1.54+/-0.3 vs. -0.53+/-0.2 beats/mmHg). Considering that the reduction in BG could be secondary to the increase in MAP in response to microinjection of muscimol (n=6) or baclofen (n=7) into the NTS, in these two groups of rats we brought the MAP back to baseline by infusion of sodium nitroprusside (NP, 3.0 microg/0.05 ml, i.v.). Under these conditions, we verified that the BG remained significantly reduced after muscimol (-1.49+/-0.2 vs. -0.35+/-0.2 beats/mmHg) and after baclofen (-1.72+/-0.2 vs. -0.33+/-0.2 beats/mmHg) when compared to control. Reflex tachycardia was observed during the normalization of MAP by NP infusion and, in order to prevent the autonomic imbalance from affecting BG, we used another group of rats treated with atenolol (5 mg/kg, i.v.), a beta1 receptor antagonist. In rats previously treated with atenolol and submitted to NP infusion, we verified that BG remained reduced after microinjection of muscimol or baclofen into the NTS. The data show that activation of GABA(A) and GABA(B) receptors, independently of the changes in the baseline MAP or HR, inhibited the neurons of the NTS involved in the parasympathetic component of the baroreflex. PMID- 11109991 TI - Electrical properties and synaptic potentials of rabbit pancreatic neurons. AB - Pancreatic ganglia receive innervation from a wide variety of extrinsic nerves and supply the predominant innervation to pancreatic acini, islets, and ducts. This study used intracellular recordings to investigate the electrical properties and synaptic potentials of rabbit pancreatic neurons. Neurons had a mean resting membrane potential of -54+/-0.4 mV and generated action potentials with a mean overshoot of 10+/-0.4 mV and a mean after-spike hyperpolarization (ASH) of 11+/ 0.5 mV with duration of 210+/-19 ms. Action potentials exhibited a high threshold (-15+/-1 mV) for intracellular stimulation and a phasic firing pattern was observed in response to prolonged depolarizing currents. Stimulation of attached nerve bundles evoked multiple fast excitatory postsynaptic potentials (fEPSPs) which were abolished by hexamethonium in 75% of neurons, while a non-cholinergic fEPSP was observed in 25% of the neurons. Repetitive stimulation (3-30 Hz) evoked muscarinic slow EPSPs with a mean amplitude of 8+/-2 mV and duration of 5+/-1 s in a small subset (21%) of neurons. Exogenous muscarine evoked a mean slow depolarization of 10+/-1 mV amplitude in 22% of neurons tested. Following repetitive nerve stimulation non-cholinergic late, slow EPSPs with a mean amplitude of 4.3+/-0.4 mV were recorded in 32% of neurons. Nicotinic transmission was subject to inhibition mediated by presynaptic muscarinic receptors at low (0.5 Hz) stimulus frequencies in 80% of neurons. At higher frequencies (> or =1 Hz), either facilitation or depression of nicotinic transmission was observed depending on the ganglion studied. A population (9%) of neurons exhibited spontaneous, low-amplitude pacemaker-like potentials. Spontaneous fEPSPs and action potentials were also observed and these occasionally occurred in rhythmically timed bursts. Thus, distinct subpopulations of pancreatic neurons could be identified on the basis of both their intrinsic electrical properties and the receptors mediating and/or modulating synaptic transmission. These neurons function as critical sites of integration for synaptic input from extrinsic pancreatic nerves and thereby determine the postganglionic firing patterns presented to the pancreatic exocrine and endocrine secretory cells. PMID- 11109993 TI - Morphological study of cultured cardiac ganglionic neurons from different postnatal stages of rats. AB - This study sought to establish a culture model of cardiac ganglia (CG) neurons of the Sprague-Dawley (SD) rat which could by used to study the distinct characteristics of CG neurons. After culturing, the morphology and immunocytochemistry of CG neurons obtained on different days after birth were compared. Samples of CG neurons were taken from the posterior atrial wall of rats aged 7, 14, 21 and 40 postnatal days (designated as P7, P14, P21 and P40, respectively). During 3-6 days of culture, the morphological changes of the cultured neurons were monitored using a light microscope. Immunocytochemical staining of the neurofilaments (NF-L, -M and -H) was performed to identify the CG neurons and the changes in morphology. The differences in size of the CG soma of each culture were compared by morphometry. Frozen sections of CG neurons were used as the in vivo control of the above experiments. The results showed that the rate of growth in size of the CG soma was highest in the P7 group, and was slower after weaning (21 days after birth). Cultured neurons were categorized into unipolar-like (Type I), multipolar-like (Type II), and bipolar-like (Type III) based on their morphological characteristics. In NF immuocytochemical staining, there were strong responses to NF-H and NF-M in all cultures, but not to NF-L. More specifically, responses to NF-H were mainly observed in perikaryons and neurites, whereas the responses to NF-M were mainly in perikaryons. The present study has established a culture system for cardiac ganglia neurons of SD rats. Our results show that the intracardiac neurons were still developing in their somata and the processes and that various responses to different antibodies of NF for CG neurons occurred in different postnatal stages in rats. PMID- 11109992 TI - Acute injection of 17beta-estradiol enhances cardiovascular reflexes and autonomic tone in ovariectomized female rats. AB - Among the many benefits of long-term hormone replacement therapy to postmenopausal women is a significant reduction in risk for and progression of cardiovascular disease. However, long-term estrogen replacement therapy has been associated with several undesirable, and likely dose-dependent, side-effects. There is some evidence to suggest that the dose of estrogen which confers optimal beneficial effects on the cardiovascular system is much lower than that which is currently prescribed for postmenopausal women. The following experiments were conducted to determine the dose-response relationship of acutely administered estrogen on autonomic tone and reflex control of heart rate in ovariectomized Sprague-Dawley female rats. Rats were anaesthetized with sodium thiobutabarbital (100 mg/kg) and instrumented to record blood pressure, heart rate and efferent parasympathetic and sympathetic nerve activities. The sensitivity of the cardiac baroreflex was tested using intravenous injection of either phenylephrine hydrochloride (0.025-0.1 mg/kg) or sodium nitroprusside (0.0025-0.01 mg/kg). Intravenous injection of estrogen produced dose-dependent increases in the magnitude of the baroreflex sensitivity and parasympathetic tone while reducing sympathetic tone with a maximal effect observed at 1 x 10(-3) mg/kg. Prior administration of the selective estrogen receptor antagonist, ICI 182,780 blocked the estrogen-induced changes in baroreflex sensitivity and autonomic tone. These results demonstrate that acutely administered, low-dose estrogen has beneficial effects on autonomic tone and cardiovascular reflexes. PMID- 11109994 TI - Nervous control of blood flow microkinetics in the infrared organs of pit vipers. AB - The pit organ of pit vipers contains a membrane which serves as an infrared retina, processing infrared information by the degree to which the temperature of trigeminal nerve receptors (terminal nerve masses) is raised. The receptors are arranged in a monolayer array within the pit membrane and irrigated by a capillary network which both supplies energy to the terminal nerve masses and serves as a heat exchange mechanism. This mechanism maintains the receptors at a stable temperature level to increase or decrease their sensitivity and to reduce to a minimum the afterimage effect of a moving stimulus. We used a Doppler laser blood flow meter to measure the local changes in blood flow in response to a point heat source (a small soldering iron) and to direct stimuli (red and infrared lasers). Resection of any one of the trigeminal A-delta fiber trunks innervating the pit membrane abolished blood flow response in the area innervated, but resection of the main trunk between the primary neurons and the medulla left the response intact. In addition to the A-delta fibers the pit membrane contains autonomic and sensory C-fiber innervation, but preganglionic resection of parasympathetic neurons, and chemical blocking of postganglionic fibers with atropine and capsaicin had no influence on the blood flow changes. Therefore, on the basis of the rapid response time and the similarity of the blood flow curves to electrophysiological recordings from the receptors, we surmised that all blood flow changes were due to a vasomotor reaction, modulated by the terminal nerve masses directly, resulting in a change in local heat capacity that cools the stimulated receptors back to a basal temperature. PMID- 11109995 TI - Renal nerves and endothelins interaction in the control of renal excretory function in conscious Long-Evans rats. AB - The role of renal nerves and endothelins, acting at ET(A) receptors, in the regulation of renal excretory function was investigated in male Long-Evans rats. Catheters were placed in the femoral vein for fluid and drug infusion, in the femoral artery for blood pressure recording as well as in the bladder for urine collection. Infusion of 16.4 nmol/kg/min of the ET(A) receptor antagonist BQ-123 for 50 min was performed in freely moving, intact and renal denervated rats. As a result of BQ- 123 infusion, urine flow rate diminished (P < 0.02) and Uosm increased (P < 0.05) in the intact rats, but not in the renal denervated rats. Bilateral renal denervation itself as well as ET(A) receptor inhibition in both intact and renal denervated rats did not change the mean arterial pressure, heart rate, or the excretion of sodium, potassium and chloride. The data obtained suggest an interrelationship between renal nerves and endothelin-A receptors in the regulation of renal water excretion. PMID- 11109996 TI - NADPH-diaphorase reveals presumptive sympathetic primary afferents in the developing human spinal cord. AB - Numerous studies have elucidated two visceral afferent pathways in the spinal cord of mammals, the lateral collateral pathway (LCP) and the medial collateral pathway (MCP). The present study utilized NADPH-diaphorase histochemistry to visualize afferent pathways in the developing human thoracolumbar spinal cord. Diaphorase-positive fiber bundles, strikingly similar to the previously defined LCP and MCP, were observed coursing along the lateral and medial aspects of the dorsal horn to the base of the dorsal horn, the intermediate gray, and/or the dorsal commissure. Furthermore, some axons forming the MCP crossed in the dorsal commissure to the contralateral side of the spinal cord. In addition, axons projecting in the LCP often appeared to terminate within clusters of diaphorase labeled sympathetic preganglionic neurons, supporting the concept that monosynaptic connections may exist between primary afferents and autonomic motor neurons. Thus, nitric oxide may be involved in both afferent and efferent neurons in reflex pathways of the human sympathetic nervous system. PMID- 11109997 TI - Canonical correlation and chi-square: relationships and interpretation. AB - A 2 x 2 chi-square can be computed from a phi coefficient, which is the Pearson correlation between two binomial variables. Similarly, chi-square for larger contingency tables can be computed from canonical correlation coefficients. The authors address the following series of issues involving this relationship: (a) how to represent a contingency table in terms of a correlation matrix involving r - 1 row and c - 1 column dummy predictors; (b) how to compute chi-square from canonical correlations solved from this matrix; (c) how to compute loadings for the omitted row and column variables; and (d) the possible interpretive advantage of describing canonical relationships that comprise chi-square, together with some examples. The proposed procedures integrate chi-square analysis of contingency tables with general correlational theory and serve as an introduction to some recent methods of analysis more widely known by sociologists. PMID- 11109998 TI - Statistical significance levels of nonparametric tests biased by heterogeneous variances of treatment groups. AB - The statistical significance levels of the Wilcoxon-Mann-Whitney test and the Kruskal-Wallis test are substantially biased by heterogeneous variances of treatment groups--even when sample sizes are equal. Under these conditions, the Type I error probabilities of the nonparametric tests, performed at the .01, .05, and .10 significance levels, increase by as much as 40%-50% in many cases and sometimes as much as 300%. The bias increases systematically as the ratio of standard deviations of treatment groups increases and remains fairly constant for various sample sizes. There is no indication that Type I error probabilities approach the significance level asymptotically as sample size increases. PMID- 11109999 TI - Ipsilateral loudness adaptation over multiple intensity levels. AB - Could monaural loudness adaptation be a simple artifact of psychophysical contrast? From adaptation data based on the Ipsilateral Comparison Paradigm (ICP), A. J. Dange, J. S. Warm, E. M. Weiler, and W. N. Dember (1993) concluded that loudness adaptation was not an artifact of psychophysical contrast, but their conclusion was dependent on results from one intensity. This study, involving multiple intensities, re-examined the issue of contrast versus adaptation and generally supported the conclusions of Dange et al. The results also showed an unexpected asymmetry of adaptation based on the direction of the referent modulation used with the ICP technique. Some implications are discussed. PMID- 11110000 TI - The relationship between cognitive ability and positive and negative priming in identity and spatial priming tasks. AB - The authors examined whether stimulus activation and inhibition in the identity priming task are related to the temporal lobe, and whether these processes in the spatial priming task are related to the parietal lobe. Forty participants performed spatial and identity positive and negative priming tasks, the Vandenberg Mental Rotation task, and the Digit Span task. Both men and women showed significant positive and negative priming in the identity and spatial tasks with no gender difference. The magnitude of identity positive priming was predicted by the Digit Span task, and the magnitude of spatial positive priming was predicted by the mental rotation task. Only women showed a correlation between spatial ability and spatial negative priming. The results are partially consistent with the dorsal-ventral model of cognitive inhibition. PMID- 11110001 TI - Recall of television commercials as a function of viewing context: the impact of program-commercial congruity on commercial messages. AB - The effect of the congruity between the involvement types of advertising commercial and a television program on the effectiveness of the commercial was studied. Participants (N = 103) viewed either a cognitive or an affective commercial for a product, which was embedded in either a cognitive or an affective television program. The results showed that the effects of the congruence influence the impact on memory. Free recall and cued recall were significantly influenced by the program-commercial congruity. Free recall and cued recall were significantly higher for the cognitively involving commercial in the cognitively involving program context than in the affectively involving program context. Similarly, free recall and cued recall were significantly higher for the affectively involving commercial in the affectively involving program context than in the cognitively involving program context. PMID- 11110002 TI - Intuitive covariation assessment of the illusory correlation. AB - Most of the research concerned with the illusory correlation is modeled after the seminal work of D. L. Hamilton and R. K. Gifford (1976). However, S. A. Haslam and C. McGarty (1994) have voiced concerns over the dependent measures used within this paradigm. Therefore, in this study, the authors tested a new dependent variable that has high face validity. This measure was modeled after the work of J. R. McGahan and R. Wight (1989) and consisted of a set of propositional statements representing either the illusory correlation, the contingency opposite the illusory correlation, or the noncontingency. A second purpose of this study was to validate other studies that have used dependent measures modeled after the work of J. R. McGahan and R. Wight (1989). Demonstrating that this measure can be used to detect a well-documented phenomenon (i.e., the illusory correlation) would strengthen the results and conclusions from other studies. To this end, results from 2 experiments indicate that this measure does provide a valid alternative to those measures that are commonly used in illusory correlation studies. The results thereby give credence to other studies that have used similar dependent measures. PMID- 11110003 TI - Locomotor analysis of the taiep rat. AB - Locomotor activity (tremor, ataxia, immobility, epilepsy, and paralysis) in the taiep rat, which suffers from a myelin deficient disorder, has not been previously documented. This study used walking track analysis of footprints to analyze locomotor activity in the taiep rat in comparison to normal, age-matched controls. The results confirmed differences between normal and taiep rats in terms of stride length, step length, and stride width. In addition, we found significant interactions between age and condition for stride and step length. The results suggest that locomotor analysis is a sensitive indicator of myelin deficiency. The results are discussed in terms of the underlying myelin deficiency and possible treatment regimens. PMID- 11110004 TI - Departures from sensible play in computer blackjack. AB - Gambling has been viewed as irrational, and even though blackjack offers rational strategies (i.e., Basic [E. Thorp, 1966] and card counting), people exhibit departures from rationality (e.g., "Never Bust" strategies). To determine whether departures from rational behavior reflect ignorance or fatigue, university students were provided with on-line Basic advice while playing a simplified computer blackjack. Although the on-line advice initially affected the totals these players sat on, it was eventually discarded for higher risk strategies. Irrational play did not reflect ignorance or fatigue and was not necessarily conservative. Real fluctuations of odds in blackjack may lead to situations in which Basic is not perceived by players as effective. Because Basic is not a personalized strategy, it seems less likely to be maintained in the face of losses. Players were more optimistic that they might win when utilizing their personalized strategies. PMID- 11110005 TI - Pictures, words, and sounds: from which format are we best able to reason? AB - The effect of presentation format on reasoning was studied with a sentence verification task. Background information was presented in single-format and combined conditions that included pictured, printed, or spoken versions of the stimulus items. In Experiment 1, a test sentence appeared together with the background at varied stimulus onset asynchronies, to study how format influences the acquisition of the stimulus information. In Experiments 2 and 3, however, the test sentence followed the presentation of the background, to test the effect of format on memory. Reaction time responses to the test sentences showed a consistent picture advantage. However, when participants responded to materials stored in memory, both pictured and spoken formats provided quicker responses in comparison to printed words, and the format difference was smaller than when materials were readily available on the screen. Multimedia presentations, when compared with single-format conditions, did not provide additional benefits. PMID- 11110006 TI - Abrupt and brief discontinuation of antidepressant treatment: effects on cognitive function and psychomotor performance. AB - The abrupt discontinuation of antidepressants can result in a syndrome of adverse events, including somatic, mood and psychomotor reactions. This study examined the effects of discontinuing and resuming antidepressant treatment with four selective serotonin reuptake inhibitors (SSRIs) on cognitive and psychomotor function. Eighty-seven patients receiving maintenance therapy with fluoxetine, sertraline, paroxetine or citalopram had their treatment interrupted for 4-7 days using double-blind placebo. Assessments of aspects of cognitive and psychomotor performance, mood and symptoms were carried out at each visit. Following interruption of treatment, significant differences between the groups emerged. Paroxetine treated patients experienced significantly more cognitive failures (P = 0.007), poorer quality of sleep (P = 0.016), and an increase in depressive symptoms, as rated both subjectively, using the Zung scale (P = 0.006) and by the clinician, using the Montgomery-Asberg Depression Rating Scale (P = 0.0003) and Clinical Global Impression (P = 0.0003), compared to some or all of the other drugs. All changes were reversed on reinstatement of treatment. Abrupt discontinuation of treatment with paroxetine leads to deterioration in various aspects of health and functioning, which may be related to the antidepressant discontinuation syndrome. These effects are not evident in patients receiving fluoxetine, sertraline and citalopram, suggesting they are not an SSRI class phenomenon. PMID- 11110007 TI - Disabilities and quality of life in pure and comorbid generalized anxiety disorder and major depression in a national survey. AB - Using a nationally representative sample, this study examines the disease specific impairments of DSM-IV generalized anxiety disorder (GAD) by comparing them to the impairments associated with major depressive disorder (MDD). Results are based on 4181 respondents between the ages of 18-65 years who were interviewed with the 12-month version of the Munich-Composite International Diagnostic Interview as part of the German National Health Interview and Examination Survey-Mental Health Supplement (GHS). After controlling for age, gender, and other psychopathology, 'pure' current GAD without MDD (n = 33), pure MDD (n = 344) and comorbid GAD and MDD (n = 40) were each associated with high impairment as defined by poor self-perceived health, at least 3 days limited or impaired in the past month, and low quality of life scores [from the Short Form 36 Health Survey (SF-36)]. Quality of life scores on several of the SF-36 scales were significantly lower for respondents with pure GAD as compared to respondents with pure MDD. Overall, the results show that DSM-IV GAD is associated with high impairment even after controlling for other psychopathology. The impairment outcomes for GAD were comparable in size to those for MDD. These findings underline the significance of this disorder from a clinical and social perspective and provide support for the independent diagnostic status of GAD. PMID- 11110008 TI - An assessment of selective serotonin reuptake inhibitor discontinuation symptoms with citalopram. AB - The selective serotonin reuptake inhibitors (SSRIs) have recently been associated with a variety of somatic and psychiatric symptoms upon abrupt drug discontinuation. These symptoms have been variously termed SSRI withdrawal, or SSRI discontinuation syndrome. Although all of the available SSRIs have been reported to cause discontinuation symptoms, some appear to have a greater propensity to cause these adverse events than others. Data from a previously completed placebo-controlled, double-blind study designed to assess citalopram in depression relapse prevention were analysed to assess patients for the emergence of discontinuation effects following randomization to placebo after 8 weeks of active drug treatment. Side-effects that occurred during the first 2 weeks following randomization to active drug (n = 150) or placebo (n = 72) were measured using the UKU unwanted side-effect list. The proportion of patients that experienced one or more events over the 2-week period following randomization was similar in the two groups, and there was no association between citalopram dose prior to randomization and the reporting of symptoms. Most of the events that did occur were mild in intensity and none resulted in discontinuation from the study. Events occurring at a higher frequency in the placebo group were most associated with the central nervous system (CNS). These events may reflect a re-emergence of depressive symptoms, since only 14.8% of patients randomized to placebo who did not relapse experienced CNS events, a low symptom incidence that was non significant (P = 0.562) compared to patients continuing treatment (10.9%). Therefore, this assessment suggests that any symptoms associated with rapid discontinuation of citalopram are mild and transient, and emphasizes the significant role re-emerging depression and / or anxiety may play in the assessment and identification of SSRI discontinuation symptoms. PMID- 11110009 TI - The efficacy of sertraline in panic disorder: combined results from two fixed dose studies. AB - Data from two fixed-dose studies of sertraline in panic disorder were pooled in order to provide sufficient power for the analysis of treatment response in clinically relevant subgroups. Male and non-fertile female patients meeting DSM III-R criteria for moderate-to-severe panic disorder with or without agoraphobia completed a 1-2 week placebo run-in period, and then were randomized to 12 weeks of double-blind treatment with either placebo, or one of three fixed daily doses of sertraline (50 mg, 100 mg, or 200 mg). Eighty-two patients were treated with placebo and 240 patients were treated with one of three doses of sertraline. All three sertraline doses produced significant efficacy compared to placebo, with no consistent evidence of a dose-response effect. For the subset of patients with subsyndromic depression at baseline [baseline Hamilton Depression Rating scale (HAM-D > 12 and < or = 21], sertraline yielded a significantly higher panic-free rate than did placebo (P = 0.021), again, by a conservative endpoint (Last Observation Carried Forward method, LOCF) analysis. Sertraline was well-tolerated at all dose levels, with no significant between-dose differences in patients discontinuing due to adverse events. The presence of mild-to-moderate subsyndromic levels of depression did not reduce the anti-panic efficacy of sertraline. PMID- 11110010 TI - Effects of risperidone on affective symptoms in patients with schizophrenia. AB - The effects of risperidone on affective symptoms were determined by an analysis of pooled data from six double-blind trials of risperidone versus haloperidol in 1254 patients with chronic schizophrenia. Symptoms indicating mania were assessed by the Positive and Negative Syndrome Scale (PANSS) excitement and grandiosity items and by the excited cluster (excitement, hostility, uncooperativeness, and poor impulse control); anxious / depressive symptoms were assessed by the PANSS anxious / depressive cluster (somatic concern, anxiety, guilt feelings, and depression). Mean change scores from baseline to endpoint were compared in patients receiving risperidone, haloperidol or placebo by analysis of variance with factors for trial and baseline score included in the model. In all patients, change scores on excitement and grandiosity items and excited and anxious / depressive clusters were significantly greater for risperidone than for haloperidol or placebo. Dropouts due to inefficacy were less frequent with risperidone (5 of 59; 8%) than with haloperidol (7 of 38; 18%) or placebo (8 of 10; 80%). In patients with anxious / depressive symptoms at baseline (anxiety / depression cluster score > or = the median), anxiety / depression scores decreased significantly more with risperidone than with haloperidol, and symptom reduction occurred faster with risperidone. These results are consistent with previous reports and suggest that risperidone is more efficacious than haloperidol for affective symptoms in patients with schizophrenia. PMID- 11110011 TI - Reboxetine in the treatment of bulimia nervosa: a report of seven cases. AB - Controlled trials in patients with bulimia nervosa have demonstrated efficacy of antidepressant medications with serotonergic function (e.g. fluoxetine) as well as noradrenergic function (e.g. desipramine). Seven outpatients with bulimia nervosa according to DSM-IV criteria were treated openly with 8 mg of reboxetine, a selective noradrenaline reuptake inhibitor (NRI) over a 12-week period. The patients were assessed with the Structured Clinical Interview for DSM, Clinical Global Impression, 17-item Hamilton Depression Rating scale (HAM-D), Eating Disorder Inventory, Eating Disorders Questionnaire, daily self-ratings of eating behaviour, and the UKU side-effect rating scale. Three patients dropped out prematurely, one after 6 weeks and two after 4 weeks of reboxetine treatment. The reasons for premature attrition were rapid remission in one patient after 2 weeks and constipation, which led to an increase in episodes of laxative intake in two patients. In the total group, the monthly binge eating frequency showed a reduction of 73% and the frequency of vomiting episodes per month decreased by 67%. Furthermore, there was a concomitant decrease of depression ratings (HAM-D: from 12.2-6.1). Reboxetine seems to be an option for the treatment of bulimia nervosa. PMID- 11110012 TI - Insomnia related to biperiden withdrawal in two schizophrenic patients. AB - It is not uncommon for patients who are receiving antipsychotic medication to be given anticholinergic agents, such as biperiden, despite the relative absence of neurological side-effects. Two cases of schizophrenia are reported in which insomnia developed after biperiden withdrawal or reduction. The insomnia continued until biperiden treatment was reinstated, despite the fact that the patients did not exhibit signs or report symptoms indicative of antipsychotic drug-induced neurological side-effects. The occurrence of insomnia following the withdrawal of biperiden or reduction in the dose has not been previously reported. One potential explanation for the insomnia is cholinergic rebound following the withdrawal of biperiden. PMID- 11110013 TI - Serotonin and anxiety: current models. AB - Evidence for the importance of the serotonin system in anxiety disorders has increased substantially in recent years. Although preclinical research has provided an important source of hypotheses, clinical work on the value of selective serotonin reuptake inhibitors (SSRIs) in these conditions has been particularly persuasive. In this paper, a number of models of serotonin in the anxiety disorders are reviewed, and the clinical advantages of the SSRIs in anxiety disorders are emphasized. Models of serotonin in anxiety disorders include: 1) a 'see-saw model'; 2) an 'amygdala model'; and 3) a 'basal ganglia model'. While any simplistic schema in this complex area must fail, these various models have some heuristic value for the clinician. From a clinical perspective, the availability of the SSRIs has proved a significant step forward in the treatment of the anxiety disorders such as panic disorder, obsessive-compulsive disorder, post-traumatic stress disorder, and social anxiety disorder (social phobia); certainly these agents offer several advantages over previously existing pharmacotherapeutic alternatives such as the benzodiazepines and tricyclic antidepressants. PMID- 11110014 TI - Pharmacotherapy of obsessive-compulsive disorder: experience with the selective serotonin reuptake inhibitors. AB - Since the introduction of the selective serotonin reuptake inhibitors (SSRIs) a decade ago, they have become first-line agents in the treatment of obsessive compulsive disorder (OCD). Numerous clinical trials have confirmed their efficacy, and established their superior risk-benefit ratio in comparison with clomipramine, a non-selective serotonin reuptake inhibitor. Relatively higher doses and longer duration of treatment may be necessary to effect a response in OCD, with long-term treatment being required to maintain therapeutic gains. Despite the advances represented by the SSRIs, treatment resistance remains a problem. While no one solution exists, various strategies, including pharmacotherapy augmentation, look promising. PMID- 11110015 TI - The efficacy of the selective serotonin reuptake inhibitors for social anxiety disorder (social phobia): a meta-analysis of randomized controlled trials. AB - This article reviews all available studies on the use of selective serotonin reuptake inhibitors (SSRIs) for the pharmacotherapy of social anxiety disorder. Using the search methods laid out by the Cochrane Collaboration, 25 published reports of SSRI effectiveness for social anxiety disorder were identified, of which eight were randomized, double-blind, placebo-controlled trials (RCTs). The odds ratios of responder status ('much improved' or 'very much improved' on the Clinical Global Impression Scale) for SSRI versus placebo varied between 2.1 and 26.2. In no RCT was the lower confidence limit less than 1. The number needed to treat varied from 1.6 to 4.2. The number of patients who responded to drug was approximately twice the number who responded to placebo. Comparing the change in mean Liebowitz Social Anxiety Scale score in patients treated with drug versus those treated with placebo, the effect sizes of the RCTs varied from 0.3 to 2.2. In four RCTs the effect size was 'large', in one 'moderate' and in two 'small'. Furthermore, response rates and effect sizes for SSRIs were larger than those seen in trials of the reversible monoamine oxidase inhibitors (RIMAs). It may be concluded with a high degree of confidence that SSRI treatment for social anxiety disorder is effective, both in reducing total levels of social anxiety and in improving patients' overall clinical condition. PMID- 11110016 TI - Selective serotonin reuptake inhibitors in the treatment of panic disorder and agoraphobia. AB - This review article summarizes comparator-controlled, short-term studies with currently available selective serotonin reuptake inhibitors (SSRIs) in the treatment of panic disorder and agoraphobia. Fluvoxamine, fluoxetine, paroxetine, sertraline and citalopram have all been proven to be superior to pill-placebo in the treatment of panic disorder, agoraphobia and associated symptoms such as depression. Direct comparisons with other antidepressants, benzodiazepines, cognitive-behavioural therapies or combinations of SSRIs with psychotherapeutic interventions are scarce. The majority of studies have reported on fluvoxamine whereas, to date, sertraline and citalopram have been compared only with placebo. Meta-analyses have suggested that combining an antidepressant with exposure in vivo produces the greatest treatment gains. Since this procedure is already commonly used in everyday clinical practice, it is recommended that future research in the treatment of panic disorder be directed towards the investigation of a combination of SSRIs with exposure therapy. PMID- 11110017 TI - Selective serotonin reuptake inhibitors in the treatment of post-traumatic stress disorder: a meta-analysis of randomized controlled trials. AB - Advances in the neurobiology of post-traumatic stress disorder (PTSD) and the availability of modern psychotropics have led to renewed interest in the pharmacotherapy of this disorder. In this paper we focus on trials of the selective serotonin reuptake inhibitors (SSRIs) in PTSD. Studies of the pharmacotherapy of PTSD were identified using methods developed by the Cochrane collaboration. Although a range of open trials of different SSRIs in PTSD show promise, there are few controlled pharmacotherapy studies in this disorder. Nevertheless, pharmacotherapy for PTSD appears to have reasonably robust effects, with odds ratios for responder status, defined as 'much improved' or 'very much improved' on the Clinical Global Impression Scale (CGI), on drug versus placebo varying from 2.2 to 5.6 in randomized controlled trials of different agents. The SSRIs appear both safe and effective for this indication. Additional research with these agents is necessary to clarify many questions, including predictors of response, duration of treatment, comparison with other agents, and integration with psychotherapy. In the interim, however, the SSRIs can be recommended as a first-line medication for the treatment of PTSD. PMID- 11110018 TI - Selective serotonin reuptake inhibitors in mixed anxiety-depression. AB - The overlap between the depressive and anxiety disorders is extremely common. The introduction of the selective serotonin reuptake inhibitors (SSRIs) has, more than any other development, bridged the gap in terms of efficacy in both sets of disorders. A substantial body of data exists suggesting that the available SSRIs have substantial efficacy in anxiety symptoms co-occurring with depression. The clear utility of the SSRIs in disorders classified apart from depression is also established. Whilst panic disorder is the best studied, evidence on the efficacy of the SSRIs in disorders that previously did not attract much pharmacotherapeutic interest, such as social anxiety disorder and post-traumatic stress disorder is accumulating. PMID- 11110019 TI - Selective serotonin reuptake inhibitors in the treatment of paediatric anxiety disorders: a review. AB - Anxiety disorders (obsessive-compulsive disorder, social phobia/selective mutism, panic disorder, separation anxiety, generalized anxiety disorder, simple phobia and post-traumatic stress disorder) are the most prevalent psychiatric disorders in children and adolescents. The selective serotonin reuptake inhibitors (SSRIs)- citalopram, fluoxetine, fluvoxamine, paroxetine and sertraline--have demonstrated efficacy in treating anxiety disorders in adults. Although less information is available on the use of these agents in the paediatric population, research into the SSRIs for childhood anxiety disorders is increasing. This article reviews current literature, including case reports as well as open and controlled trials, on the effectiveness and tolerability of the SSRIs in the paediatric population. It also discusses developmental differences in children that should be considered in the utilisation of the SSRIs in paediatric patients. PMID- 11110020 TI - Determination of benzo[a]pyrene and other polycyclic aromatic hydrocarbons (PAHs) at trace levels in human tissues. AB - A sensitive and rugged gas chromatographic-mass spectrometric (GC-MS) method was developed for determining 11 polycyclic aromatic hydrocarbons (PAHs) in 4-g specimens of human lung, breast, and liver tissue. The method quantitation limit (MQL) was 0.01-0.02 ng/g for benzo[a]pyrene (BaP) and six other five- and six ring PAHs. The MQL was higher for four-ring PAHs because of their presence at trace levels in method blanks. The average MQLs for pyrene and chrysene were 0.05 and 0.03 ng/g, respectively. The method was applied to 200 human tissue specimens (89 lung, 68 breast, and 43 liver) obtained from patients during surgery. Quality control results demonstrated average recoveries of 80% or better from reagent controls spiked at the 0.2-ng level and average recoveries from tissue fortified at the 0.25-ng/g level of 66-95%. The precision of the method was determined from duplicate analyses of specimens (16-38% RSD) and from duplicate GC-MS analysis of tissue extracts (8-17%RSD). Benzo[a]pyrene was detected at measurable levels in 87% of the lung specimens. This method makes possible the measurement of ambient levels of PAHs in small samples of human tissue such as those obtained by biopsy. PMID- 11110021 TI - Determination of dialkyl phosphates in human urine using gas chromatography-mass spectrometry. AB - Organophosphates are used as pesticides in agriculture and pest control. They are metabolized to dialkylphosphates, which are excreted in urine. Determination of these metabolites is useful for assessing human exposure to organophosphates. This publication describes a new, reliable, and very sensitive analytical procedure for quantitating dimethylphosphate (DMP), diethylphosphate (DEP), O,O dimethylthiophosphate (DMTP), O,O-diethylthiophosphate (DETP), O,O dimethyldithiophosphate (DMDTP), and O,O-diethyldithiophosphate (DEDTP) in human urine. The analytes are extracted from acidified urine into a mixture of diethylether and acetonitrile. Dibutylphosphate serves as internal standard. Derivatization is performed using pentafluorobenzylbromide at 40 degrees C overnight. After further liquid-liquid extraction, analysis is carried out by gas chromatography-mass spectrometry. The limits of detection are 5 microg/L urine for DMP and 1 microg/L for the other five metabolites. Using the new procedure, 54 spot urine samples from persons in the general population in Germany were analyzed. Nearly every sample contained DMP, DEP, and DMTP, and the median values (95th percentiles) of the concentrations were 30 microg/L (105 microg/L), 4 microg/L (21 microg/L), and 22 microg/L (174 microg/L), respectively. DETP and DMDTP were found in lower concentrations. PMID- 11110022 TI - Analysis of six anticonvulsant drugs using solid-phase extraction, deuterated internal standards, and gas chromatography-mass spectrometry. AB - A rapid method for simultaneously determining the anticonvulsant drugs carbamazepine, ethosuximide, phenobarbitone, phenytoin, primidone, and valproic acid is described. Blank plus single-point calibration gives reliable quantitation from therapeutic to high fatal concentrations, except for ethosuximide, for which it gives semiquantitative results. Whole blood and liver tissue samples containing deuterated internal standards were extracted using Bond Elut Certify columns. Butyl derivatives were formed using n-iodobutane and TMAH under mild conditions and were extracted into ethyl acetate as a cleanup step. Recoveries were greater than 50%, except for valproic acid (42%). Sample preparation time was less than 2 h, and the GC run time was less than 20 min per injection. At least two ion pairs formed by electron impact ionization were monitored for each drug. Intraday CVs were less than 6.28% (4.20%) and interday CVs less than 14.1% (for midtherapeutic concentrations in blood [liver], except for ethosuximide). Linearity was observed from subtherapeutic to high fatal levels for all drugs. This method has been applied to forensic cases and has significantly reduced analytical time while improving case-work quality. Results of a case study involving anticonvulsant drugs are given. PMID- 11110023 TI - Determination of (+)- and (-)-bromoisovalerylurea in sera of overdosed subjects. AB - Bromoisovalerylurea (bromisovalum) is a sedative-hypnotic given orally as a racemic mixture of optical isomers (i.e., (+)- and (-)-enantiomer) and frequently taken in overdose in order to commit suicide. Sera from 16 overdosed subjects were analyzed for each enantiomer by high-performance liquid chromatography on chiral stationary phases. The (+)-enantiomer concentration was lower than the (-) enantiomer concentration in all specimens, that is, the ratio of the (+) enantiomer to the total concentration ranged from about 50% to 0%. The ratio of the (+)-enantiomer was continuously decreasing in each subject. The data indicate that the drug in gastrointestinal tract was absorbed into blood nonstereoselectively and that the drug in blood was eliminated stereoselectively. The enantioselective determination of this drug will give useful information on absorption and elimination. PMID- 11110024 TI - Extraction and cleanup methods for analysis of phenolic and neutral organohalogens in plasma. AB - A method for the analysis of potential endocrine-disrupting compounds, such as phenolic halogenated compounds (e.g., chlorinated and brominated phenols) and hydroxylated PCBs, in blood plasma is presented. Neutral halogenated compounds, specifically brominated diphenyl ethers and PCBs, are also included in the evaluation. An efficient denaturation and extraction step is described, and three methods for lipid removal are evaluated. The latter includes a nondestructive method based on high-resolution gel permeation chromatography (HR-GPC), a newly developed silica gel/sulfuric acid column, and lipid removal by sulfuric acid treatment. Recoveries, based on gas chromatography with an electron capture detector (GC-ECD), were between 70 and 90% for most of the studied compounds. The recoveries of phenolic compounds were generally slightly lower than those of the neutral compounds. The sulfuric acid treatment and silica gel/sulfuric acid column gave the highest yields for acid stable compounds, although a few target compounds were lost during that treatment and all compounds were recovered with the HR-GPC method. PMID- 11110025 TI - A highly sensitive spectrophotometric method with solid-phase extraction for the determination of methylmercury in human hair. AB - Methylmercury is the most toxic among the mercury species. In order to provide a quick method to screen samples for methylmercury, a highly sensitive spectrophotometric method was established. The method was based on formation of the red complex of methylmercury with thio-Michler's ketone, which can be extracted with n-butanol. The organic layer was determined at a maximum absorption wavelength of 564 nm. The Lambert-Beer law was obeyed for methylmercury from 1.00 x 10(-7) mol/L to 2.00 x 10(-5) mol/L with a correlation coefficient of 0.9992. The detection limit was 3 x 10(-8) mol/L. The method was used to determine methylmercury in human hair. The recovery was from 94% to 102%, and the relative standard deviation was 2.5%. The results agreed with those obtained by gas chromatography with electron capture detection. PMID- 11110026 TI - Investigation of nitrite adulteration on the immunoassay and GC-MS analysis of cannabinoids in urine specimens. AB - Nitrite ion has been identified as the active ingredient of two commercial adulterants that could cause discrepant results between the immunoassay screening and gas chromatographic-mass spectrometric (GC-MS) confirmation of 11-nor-delta9 tetrahydrocannabinol-9-carboxylic acid (THCCOOH) in urine. Procedures to chemically convert the nitrite ion at the beginning of sample preparation for GC MS analysis may not overcome all nitrite adulteration cases because portions of the THCCOOH might have been lost between the time of sample collection and the time of analysis. This study was conducted to further investigate the influence of both urine sample matrix and the duration of nitrite exposure on nitrite interference of THCCOOH detection. Forty clinical "THC-positive samples" that had been screened and confirmed positive for the presence of THCCOOH were spiked with 0.15M or 0.3M of nitrite. The levels of THCCOOH at various time intervals after nitrite spiking were monitored by instrument-based cannabinoids immunoassays (Syva EMIT d.a.u. and/or Roche Abuscreen ONLINE assays) and by an onsite THC immunoassay (Roche ONTRAK TESTSTIK). Results from this report demonstrate that the two outstanding "urine specimen factors" that dictated the effectiveness of the nitrite adulteration were urinary pH and the original drug concentration before nitrite spiking. Significant decreases in the immunoassay results could be observed within 4 h of nitrite treatment in the majority of samples with acidic urinary pH values. Regardless of their original concentration of THCCOOH (GC-MS ranging from 33 to 488 ng/mL), all of the 20 samples that had acidic pH values gave negative immunoassay results 1 day after nitrite adulteration. In contrast, the immunoassay results of samples with neutral or basic pH values were less affected by nitrite exposure in the same studies. Approximately two-thirds of the samples with pH values greater than 7.0 remained immunoassay-positive 3 days after nitrite spiking. Nevertheless, some of the adulterated urine that showed no change in immunoassay results might exhibit significant decrease in GC-MS recoveries even with bisulfite treatment, collaborating with the observations that a portion of samples screened positive with THC immunoassay in the laboratory could fail to confirm with GC-MS analysis. The decrease or loss of immunoassay detectable cannabinoid cross-reactives in acidic "THC-positive samples" can be attenuated by chemically increasing the pH value of the samples to the basic pH range. PMID- 11110027 TI - GC-MS analysis of the total delta9-THC content of both drug- and fiber-type cannabis seeds. AB - A GC-MS method was performed to determine the total delta9-THC content in both drug- and fiber-type cannabis seeds. Drug-type seeds were found to contain much higher levels of delta9-THC (35.6-124 microg/g) than fiber (hemp) seeds (0-12 microg/g). The majority of delta9-THC was found to be located on the surface of the seeds. Approximately 90% of the total delta9-THC was removed by a simple, quick wash with chloroform. Washed drug-type seeds contained less than 10 microg/g. Separation of the seeds into the kernel and testa showed that the bulk of delta9-THC is located in the testa, mainly on the outside. The kernels of drug and fiber-type cannabis seeds contained less than 2 and 0.5 microg delta9-THC/g seeds, respectively. Fluctuations in the delta9-THC content of different replicates of the same type of seeds could be the result of the degree of contamination on the outside of the seeds. PMID- 11110028 TI - Automated in-tube solid-phase microextraction coupled with liquid chromatography electrospray ionization mass spectrometry for the determination of selected benzodiazepines. AB - A simple, rapid, and sensitive method, which allowed us to simultaneously determine seven benzodiazepines (diazepam, nordiazepam, temazepam, oxazepam, 7 aminoflunitrazepam, N-desmethylflunitrazepam, and clonazepam) in buffer solution and in urine and serum samples, was investigated by automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS). In-tube SPME, in which the analytes were extracted from the sample directly into an open tubular capillary column by repeated draw/eject cycles of sample solution, is an extraction technique for organic compounds in aqueous samples. The separation of benzodiazepines was carried out under ion-suppressed reversed-phase conditions by using methanol/50mM ammonium acetate in water (60:40) as a mobile phase with a Supelco LC-18 column. The optimal extraction condition was 10 draw/eject cycles of 30 mL of sample in 100mM Tris-HCl (pH 8.5) at a flow rate of 0.3 mL/min using a piece of 60-cm length Supelco-Q plot capillary column as the extraction capillary. The quantitative study was explored by operating in selected-ion monitoring (SIM) mode. The calibration curves were linear in the range from 0.5 ng/mL or 2 ng/mL to 500 ng/mL. The detection limits were from 0.02 ng/mL to 2 ng/mL. At the optimized capillary and fragmentor voltages, the characteristic ions for each compound clearly showed up in the spectra and it is possible to use the LC-MS to identify these compounds. The method was applied to the analysis of biological samples without interfering peaks. However, the recoveries for some of the compounds in serum samples need to be further improved. PMID- 11110029 TI - New fluorescence polarization immunoassays for analysis of barbiturates and benzodiazepines in serum and urine: performance characteristics. AB - The performance of the new fluorescence polarization immunoassay reagents Cassette COBAS INTEGRA Serum Benzodiazepines assay (SBENZ) and Cassette Serum Barbiturates assay (SBARB) was evaluated as compared to other immunoassays (Abbott TDx Serum Benzodiazepines, Abbott TDx Urine Benzodiazepines, Behring EMIT Serum Benzodiazepines, Abbott ADx Serum Barbiturates, Behring EMIT Serum Barbiturates, and the COBAS INTEGRA Barbiturates (BARB) urine assay) and gas chromatography-mass spectrometry (GC-MS). Recoveries of nordiazepam and secobarbital using the SBENZ and SBARB assays, respectively, were equivalent for serum, plasma, and urine. Cross-reactivities of structurally related benzodiazepines, barbiturates, and their metabolites were very similar in serum and urine for the SBENZ and SBARB assays. Precision was within 5.4% for SBENZ serum and within 11% from 10 to 100 ng/mL for urine. Precision was within 5% for SBARB serum and within 7% from 136 to 277 ng/mL for the urine application. The standard curves for SBENZ and SBARB were stable for at least 16 weeks with the reagents stored open on the COBAS INTEGRA analyzer. Clinical comparison of the SBENZ serum assay indicated an increased pickup rate, as confirmed by GC-MS, compared to TDx and EMIT. The diagnostic sensitivities of the SBENZ serum application, TDx, and EMIT versus GC-MS were 100%, 89%, and 36%, respectively. The diagnostic specificities were 71%, 79%, and 100%, respectively. The diagnostic sensitivities of the SBENZ urine application and TDx versus GC-MS were 100% and the diagnostic specificities were 88%. The increased positive pick-up of the SBENZ assay compared to the other immunoassays is most probably due to the difference in the limit of detection (LOD) and the increased cross-reactivity for the low-dose benzodiazepines. Clinical comparison of the SBARB serum assay indicated an increased positive pick-up rate, as confirmed by GC-MS. The diagnostic sensitivities of the SBARB serum application, ADx, and EMIT versus GC MS were 96%, 65%, and 35%, respectively. The diagnostic specificities were all 100%. The diagnostic sensitivities for the SBARB urine application and BARB versus GC-MS were all 100%, and the diagnostic specificities were all 91%. The SBENZ and SBARB kits demonstrated increased sensitivity for the detection of benzodiazepines and barbiturates in both serum and urine compared to the other immunoassays. PMID- 11110030 TI - Units for sweat patch results. PMID- 11110031 TI - Aqueous humor nitric oxide levels differ in patients with different types of glaucoma. AB - Nitric oxide (NO) has effects on the regulation of aqueous humor dynamics, but the exact mechanism is not yet established. To investigate the possible roles of NO in glaucoma, we determined NO levels in aqueous humor and plasma in glaucoma patients and a control group, cataract patients. The study is an open trial with purposed sampling. One hundred fifty-two patients, including 87 glaucoma patients and 65 cataract patients from two medical centers, were recruited. NO levels in the samples were measured by a chemiluminescence assay. We found that, although the mean aqueous humor NO level (mean +/- SEM) was higher in the glaucoma patients than in the cataract patients (39.7 +/- 1.5 microM vs. 35.5 +/- 1.3 microM, p < 0.05), NO levels varied significantly in different types of glaucoma. The juvenile glaucoma patients had the lowest mean NO level (8.4 +/- 0.9 microM), while the acute angle-closure glaucoma and neovascular glaucoma patients had the highest mean NO levels (64.8 +/- 7.6 microM, 67.3 +/- 8.2 microM). In comparison, the mean plasma NO level in the glaucoma patients was not statistically different from that in the cataract patients (14.1 +/- 1.2 microM vs. 13.9 +/- 1.1 microM, p = 0.91). Our data may provide information for applying NO-mimicking nitrovasodilators in the treatment of glaucoma. PMID- 11110032 TI - Inhibition of interleukin-1-induced uveitis and corneal fibroblast proliferation by interleukin-1 blockers. AB - Interleukin-1 (IL-1) is known to trigger induction of inducible nitric oxide synthase (iNOS) to persistently mass produce nitric oxide (NO) to induce various diseases such as cancer, inflammation, Alzheimer's disease and eye diseases, including uveitis, retinopathy, age-related macular degeneration, glaucoma and myopia. Therefore, IL-1 blockers could become an important class of drugs to fight numerous diseases. Among the many compounds studied so far, 1-methyl hydrazino analogs are among the most promising agents to be developed. A minor structural change of 1-methyl hydrazino group into 1-methyl thiosemicarbazido group enhanced their anti-inflammatory activity but reduced their antiproliferation activity on corneal fibroblast cell growth. PMID- 11110033 TI - Enhancement of the mydriatic response to tropicamide by bioadhesive polymers. AB - The aim of this work was to evaluate how the addition of mucoadhesive polymers to aqueous solutions affects the ocular response of tropicamide (0.2%; w/v). The polymer solutions tested were carboxymethylcellulose sodium salt (CMC-Na; 1%; w/v), hyaluronic acid sodium salt (HA-Na; 0.1%; w/v) and polyacrylic acid (PAA; 0.2%; w/v). Polymeric solutions were compared to a nonviscous formulation (AS). In vitro mucoadhesion measurements were expressed as a percentage of the adhesion force mucin-mucin, considering this one as 100% mucoadhesion. The values ofmucoadhesion obtained were 172%, 127%, 103% and 87.6% for formulations with CMC, PAA, HA and AS, respectively. The mydriatic response of tropicamide was determined in adult male New Zealand rabbits, weighing 1.7-2 Kg, by pupil diameter measurements at different times after instillation. The area under the mydriatic response-time curve (AUC 0-6 hr) was interpreted as an indication of the bioavailability of tropicamide in each vehicle. The AUC 0-6 hr was related to the in vitro mucoadhesion for each formulation. Tropicamide solutions with CMC-Na and PAA resulted in mucoadhesion and AUC 0-6 hr values approximately 1.9 and 1.4 times higher than AS. Although the solution with HA-Na was less mucoadhesive than PAA, the hyaluronic acid solution resulted in a higher AUC mydriasis/time value. Formulations with HA-Na and PAA presented values of surface tension close to that observed in the lacrimal fluid, with the Imax (maximum pupil diameter) being higher than for CMC-Na and AS. Greater than 90% of the mydriatic effect disappeared 4.5 hr after instillation for PAA and AS. Nevertheless, the mydriatic effect remained up to 5.5 hr for HA-Na and CMC-Na. HA-Na solution enhanced the bioavailability oftropicamide, presenting a value of mucoadhesion similar to the reference mucin-mucin. PMID- 11110034 TI - Role of catalase in pre- and postjunctional responses of mammalian irides to hydrogen peroxide. AB - In the present study, we examined the effect of inhibition of catalase with 3 aminotriazole (3-AT) on hydrogen peroxide (H2O2)-induced enhancement of sympathetic neurotransmission in bovine irides and on the inhibitory effect of this oxidant on norepinephrine (NE) release from human irides, in vitro. Furthermore, we investigated the effect of 3-AT on H2O2-induced attenuation of contractile responses to carbachol in the bovine isolated irides. Isolated mammalian irides were prepared for studies of [3H]NE release using the superfusion method and for contractile studies using isolated organ baths. At concentrations less than 100 microM, H2O2 had no significant effect on field stimulated [3H]NE release from bovine or human irides. In bovine irides, 3-AT caused significant (P < 0.001) leftward shifts of concentration-response curves to H2O2 (10-300 microM). 3-AT also increased H2O2-induced attenuation of evoked [3H]NE release from human isolated irides. Low concentrations of H2O2 (< 100 microM) had no effect on carbachol contractions. However, 3-AT unmasked an inhibitory effect of low concentrations of H2O2 (3-100 microM) on carbachol induced contractions. We conclude that inhibition of catalase causes both pre- and postjunctional responses of isolated mammalian irides to be more susceptible to oxidative stress induced by H2O2. PMID- 11110035 TI - Comparison of tear sampling techniques for pharmacokinetics analysis: ofloxacin concentrations in rabbit tears after sampling with schirmer tear strips, capillary tubes, or surgical sponges. AB - This study compared the precision and accuracy of 4 tear sampling methods. In vivo, albino rabbits were treated with single bilateral eye drops ofofloxacin 0.3% solution, 3 hr after which tear samples were collected using capillary tubes (CT), surgical sponges (SS), or tear strips for 15 sec (15sTS) or 60 sec (60sTS). In vitro, CT, SS, and tear strips were spiked with known volumes of ofloxacin solution in order to assess the bioanalytical accuracy of each technique. Ofloxacin levels were quantified by HPLC in all samples. Results showed that tear volumes and ofloxacin masses sampled in vivo depended on sampling method. Tear volume followed the rank order 60sTS > 15sTS > SS > CT. The volume collected by 60sTS exceeded precorneal tear volume. Ofloxacin mass followed the order 60sTS approximately 15sTS > SS > CT. Tear concentrations (mean +/- SD; N = 12) were 3.28 +/- 3.76 microg/g for 60sTS, 10.3 +/- 10.0 microg/g for 15sTS, 9.75 +/- 8.04 microg/g for SS, and 5.83 +/- 3.35 microg/g for CT. In vitro, SS, 15sTS, and 60sTS yielded accuracies of 103-107% and coefficients of variation (CV) < 9%. CT was only 85% accurate with a CV of 31%, indicating incomplete extraction during analysis. We concluded from this study that: 1) rabbit tear sampling by SS was rapid, easy, accurate, precise, and easily analyzable; 2) sampling by CT or 15sTS was accurate, but may require aggressive extraction (for CT) or be affected by tear flow rate (for 15sTS); and 3) tear sampling by 60sTS underestimated actual tear concentrations. PMID- 11110036 TI - Acetylcholine-induced relaxation in rat ocular vasculature. AB - Endothelial cell function is often evaluated by the assessment ofendothelium dependent relaxation to acetylcholine. The purpose of this study was to characterize the acetylcholine-induced vasodilator responses in the intact ocular vasculature tree in isolated perfused rat eyes. At a fixed perfusion flow rate (5 microl/min), the resultant perfusion pressure reflected the total vascular resistance of the perfused vasculature. The baseline vascular resistance, and hence perfusion pressure, was manipulated by perfusion with high concentrations of potassium Krebs (60, 90, or 124 mM). The response to bolus administration of acetylcholine (10(-8) to 10(-4) M) was then assessed over a range of perfusion pressures. Acetylcholine induced a dose-dependent relaxation response which was larger in magnitude with increased levels of induced vascular tone. The relationship between the size of the acetylcholine-induced response and the baseline perfusion pressure was essentially linear with a correlation coefficient of 0.86 at 10(-5) M and of 0.88 at 10(-4) M acetylcholine. Our results suggest that, in the isolated perfused rat eye, the initial vascular tone has to be considered when evaluating endothelial cell function by acetylcholine-induced relaxation. PMID- 11110037 TI - The short-term effect of pentoxifylline on rabbit choroidal blood flow. AB - This study evaluates the effect of different doses ofpentoxifylline on rabbit choroidal blood flow (CBF). Sixteen albino rabbits of both sexes, weighing 2.0 3.0 kg, were randomly separated into four groups. The first group of rabbits received 2 ml normal saline injection through the ear vein. They served as the control group (group n). Three different doses of pentoxifylline, 1 mg/kg, 5 mg/kg, and 10 mg/kg, were injected intravenously to groups p1, p5 and p10, respectively. By means of a laser Doppler flowmeter, the blood cell flux (PF), velocity, and the concentration of moving blood cells (CMBC) were recorded simultaneously. The laser probe was advanced through the pars plana and positioned near the retinal surface. The mean arterial pressure was recorded at the same time. There was a significant increase in PF at 1, 5 and 10 min in group p10 rabbits compared with the control group (p=0.0005, 0.0416, and 0.0087, respectively). The velocity increased at 5 min in group p5 rabbits (p=0.0082) and at 1, 5 and 10 min in group p10 rabbits (p=0.0188, 0.0080, and 0.0207, respectively) as compared with the controls. The CMBC decreased after injection of pentoxifylline and reached statistical significance at 5 and 10 min in group p5 rabbits (p=0.0019 and 0.0046, respectively) and at 5 min in group p10 rabbits (p=0.0447). These results show that larger doses of pentoxifylline (10 mg/kg) increased the CBF of rabbits. This effect was achieved primarily by an increase in blood cell velocity. PMID- 11110038 TI - Concentration of vascular endothelial growth factor in the subretinal fluid of retinal detachment. AB - The purpose of this study was to investigate the release of vascular endothelial growth factor (VEGF) within the subretinal fluid in eyes with retinal detachment. Subretinal fluid was collected from patients with retinal detachment undergoing surgery for scleral buckling. Serum samples were also collected. The concentration of VEGF in the subretinal fluid and serum was investigated by enzyme-linked immunospecific assay. The average concentration of VEGF in serum samples was 168 +/- 153 pg/ml (mean +/- standard deviation). It was lower than the VEGF concentration in the subretinal fluid (485 +/- 570 pg/ml) in the same 18 patients with retinal detachment (p < 0.05). The average concentration of VEGF was 355 +/- 373 pg/ml in 31 samples of the subretinal fluid collected from simple rhegmatogenous retinal detachment. It was lower than the average concentration of 901 +/- 385 pg/ml in 8 samples of the subretinal fluid from retinal detachment with proliferative vitreoretinopathy (p < 0.05). The results suggest that the relative retinal ischemia in detached retina increases the release of VEGF into the subretinal space. Also, the concentration of VEGF within the subretinal fluid in proliferative vitreoretinopathy was higher than in simple rhegmatogenous retinal detachment. PMID- 11110040 TI - The ocular pharmacokinetics of ketanserin and its metabolite, ketanserinol, in albino rabbits. AB - Ketanserin, a hypotensive drug with 5-HT2 receptor antagonism, when administered by topical infusion of a 0.25% w/v solution by corneal and scleral applications, was found to lower intraocular pressure with four times more activity than its metabolite, ketanserinol. Drug and metabolite were measured periodically in the corneal epithelium, corneal stroma/endothelium, aqueous humor, iris/ciliary body, conjunctiva, sclera and lens during the infusion period (0-120 min) and the postinfusion period (120-240 min) using a fluorometric reversed-phase HPLC assay developed and verified for the research. The infusion results showed that drug entered the eye by both the corneal and scleral routes. Lateral diffusion occurred between the conjunctiva and corneal epithelium. Drug and metabolite were also detected in the untreated fellow eyes, suggesting contralateral systemic redistribution. In vitro metabolism was studied and found to occur in the corneal epithelium, iris/ciliary body and bulbar and palpebral conjunctiva but not in the corneal stroma/endothelium, aqueous humor and sclera. From noncompartmental analysis, zero-order infusion rate constants, first-order absorption constants, mean residence time, volumes of distribution at steady state (Vss) and clearance (Cle) were obtained using equations specific to the topical infusion method. Vss and Cle of aqueous humor (0.972 ml and 13.55 microl/min) were greater than aqueous humor volume (0.311 ml) and turnover rate (4.7 microl/min). PMID- 11110039 TI - Acarbose partially inhibits microvascular retinopathy in the Zucker Diabetic Fatty rat (ZDF/Gmi-fa). AB - We compared quantitative capillary retinopathic changes between non-insulin dependent diabetic Zucker Diabetic Fatty (ZDF) rats and heterozygous nondiabetic Zucker controls and evaluated the effect of an orally administered glucosidase inhibitor, acarbose, on retinopathy in these animals. Four groups of eight rats were analyzed: treated and untreated ZDF and treated and untreated Zuckers. Retinal capillary basement membrane thickening and retinal capillary cell density were determined from transmission electron microscopy and trypsin digestion preparations. ZDF rats had thicker basement membranes (p<0.0001) and more cells per unit capillary length (p=0.0003) compared to Zuckers. Acarbose treatment significantly reduced basement membrane thickening in the treated ZDF rats (p=0.001), but the effects on cell density showed only a favorable trend. Acarbose treatment has an ameliorative effect on the development of microvascular retinopathy in the ZDF rat, probably due to lessening of hyperglycemia. PMID- 11110041 TI - Teratocarcinosarcoma of the nasal cavity. Report of a case showing favorable prognosis. AB - We report a very rare case of teratocarcinosarcoma of the nasal cavity showing a favorable prognosis. The patient was a 66-year-old man with a mass completely obstructing the right nasal cavity. Subsequently, extirpation of the mass and Denker-Watsuji operation were performed, and the patient was treated with a combination of radiation therapy and chemotherapy. Neither recurrence nor distant metastasis was observed during follow-up lasting 30 months. Histologic examination of the resected mass revealed several tissue elements including columnar and squamous epithelia with atypia, smooth muscle cells with rare mitotic activity, and neuroectodermal tissue. The glandular epithelium and smooth muscle cells were reminiscent of a primitive intestinal organoid structure, suggestive of teratomatous tumorigenesis. Our case and a review of the literature indicate that the absence of invasiveness to the stroma or surrounding tissue is closely related to a favorable prognosis. PMID- 11110042 TI - Fibronectin-binding proteins secreted by Mycobacterium avium. AB - Mycobacterium avium is an intracellular pathogen and a major opportunistic infectious agent observed in patients with acquired immune deficiency syndrome (AIDS). Fibronectin is an extracellular matrix protein and is a virulence factor for several extracellular pathogenic bacteria binding to mucosal surfaces. We investigated the fibronectin (FN)-binding proteins in the culture filtrate of M. avium by two-dimensional electrophoresis (2DE). Proteins in Sauton medium of M. avium after 3 weeks were separated by 2DE. The proteins were blotted onto polyvinylidene difluoride membrane and incubated with FN. FN-binding proteins were detected by Western blotting using anti-FN antibody. FN bound to five spots (33 kDa, 32 kDa, 31 kDa, 30 kDa and 25 kDa). N-terminal amino acids of these were determined. The 33 kDa spot corresponded to antigen 85 (Ag 85) C. The 32 and 31 kDa spots were either Ag 85 A or Ag 85 B. The 30 kDa spot corresponded to Ag 85 B of M. avium. The 25 kDa spot corresponded to MPA51 (M. avium MPB51). Thus, FN bound exclusively to the Ag 85 complex and MPA51. PMID- 11110043 TI - Characterisation of isolates of Staphylococcus aureus from acute, chronic and subclinical mastitis in cows in Norway. AB - Eighty-six Staphylococcus aureus isolates from cases of bovine mastitis were characterised biochemically and with respect to serotype, multilocus enzyme electrophoresis genotypes, antibiotic sensitivity, and production of enterotoxins A through D (SEA-D) and toxic shock syndrome toxin-1 (TSST-1). The samples were obtained from 81 different cows from 79 Norwegian dairy herds in 10 different counties in southern Norway. There was an equal representation of isolates from cases of acute, chronic and subclinical mastitis. Multilocus enzyme electrophoresis using 13 genetic loci showed that 69 of 86 isolates had the same electrophoretic type. This common electrophoretic type comprised isolates that differed in the expression of other phenotypical characteristics studied. Fifty eight percent of the isolates produced one or more enterotoxins, predominantly a combination of SEC and TSST-1. Capsular serotyping revealed that 95% of the isolates belonged to serotype 8. No correlation was found between the factors studied and the clinical classification of mastitis. It appears that the majority of S. aureus isolates recovered from cases of bovine mastitis in Norway are genetically closely related and express common phenotypical characteristics. PMID- 11110044 TI - Changed expression of leukocyte adhesion molecules and increased production of reactive oxygen species caused by Streptococcus pyogenes in human whole blood. AB - To elucidate the innate immune responses to group A streptococci (GAS) important in the pathophysiology of sepsis, flow cytometric techniques were applied to study the effects of live and heat-inactivated GAS, including their particulate and soluble components, on the expression of leukocyte adhesion molecules CD11b (Mac-1) and CD62L (L-selectin), and leukocyte production of reactive oxygen species (ROS) in human whole blood. GAS caused marked time- and concentration dependent increases in CD11b and ROS, while CD62L was downregulated. Live and heat-inactivated GAS induced similar changes in leukocyte adhesion molecules, whereas ROS production induced by heat-inactivated GAS (and its particulate fraction) was 4 (2.5)-fold higher than with live GAS. Leukocyte nitric oxide production (24 h) was not enhanced. Although GAS proved a more potent inducer of ROS production, leukocyte responses to GAS were similar to those reported for lipolysaccharides, indicating that Gram-positive and Gram-negative bacteria activate common pathways in the inflammatory response. High ROS production may contribute to tissue damage caused by GAS. PMID- 11110045 TI - Sudden unexpected death as a consequence of indinavir-induced nephropathy. A case report. AB - A 60-year-old male had tested in 1986, at age 46, positive for human immunodeficiency virus (HIV). In mid-1996 he was started on a protease inhibitor regimen, which included indinavir, lamivudine and stavudine, and remained on this therapy until his death. In April 1999 he was hospitalized after a fainting episode. Although examination focusing on cardiac disease did not disclose any remarkable findings, he died suddenly one week after being discharged from hospital. At autopsy the kidneys were enlarged, with a total weight of 500 g, patchy pale gray and pinkish. Microscopy showed leukocytic cell casts in many of the tubules and collecting ducts. In many of these casts there were clefts left by crystals. In the interstitium, both in the cortex and the medulla, there was focal inflammation and fibrosis. Death was attributed to sudden cardiac dysfunction, probably ventricular fibrillation as a consequence of severe nephropathy with electrolyte disturbances. It is likely that kidney damage developed secondary to the indinavir treatment as indinavir can cause not only nephrolithiasis but also crystal-induced acute renal failure. PMID- 11110046 TI - Chlamydia trachomatis seropositivity is associated both with stillbirth and preterm delivery. AB - The cause of stillbirth and preterm delivery is often unknown. We studied the prevalence of Chlamydia trachomatis antibodies in mothers with stillbirth and preterm labor. Serum specimens from 72 mothers with stillbirth after the 21st gestational week, and from 48 mothers with preterm delivery between gestational weeks 23 and 29, both from the greater Helsinki area, and cord blood from 96 consecutive liveborn deliveries at the Department of Obstetrics and Gynecology, the University of Helsinki, were studied for antibodies to C. trachomatis immunotypes CJHI, GFK and BED by microimmunofluorescence test. The prevalence of C. trachomatis antibodies was highest, 33.3%, in mothers with stillbirth, 18.8% in mothers with preterm delivery, and 10.4% in cord blood. The IgM seropositivity rate was high among mothers with preterm delivery (8.3%). We conclude that C. trachomatis IgG antibodies are frequently detected in sera from mothers with stillbirth, suggesting past infection, while mothers with preterm delivery often have serum IgM antibodies, suggesting of acute infection. PMID- 11110047 TI - Rapid development in vitro and in vivo of resistance to ceftazidime in biofilm growing Pseudomonas aeruginosa due to chromosomal beta-lactamase. AB - The aim of this study was to examine the development of resistance of biofilm growing P. aeruginosa during treatment with ceftazidime. Biofilms were established in vitro using a modified Robbins device (MRD) and in vivo in the rat model of chronic lung infection. Three P. aeruginosa strains isolated from the lungs of cystic fibrosis (CF) patients (MICceftazidime-basal/induced beta lactamase activity: PAO 579= 0.8 mg/l-19/550 milliunits, 19676A=50 mg/l-38/957 milliunits, 17107B=100 mg/l-504/947 milliunits) were studied. After 1 or 2 weeks of continuous or intermittent (4 h/day) administration of ceftazidime to biofilms established in MDR, a statistically significant development of resistance to ceftazidime in PAO 579 or 19676A bacterial populations occurred. When ceftazidime was administered 4 h/day (200 mg/l) for 2 weeks, the frequency of resistant 19676A having MIC>25 mg/l was 4.4 10(-1) compared to 6.0-10(-5) in the control biofilm. The same trend was observed after continuous administration of ceftazidime. MICceftazidime of the more resistant variants was increased 500-fold for PAO 579 and 8-fold for 19676A, and the specific basal beta-lactamase activities from 19 to 1,400 units for PAO 579 and from 38 to 10,000 units for 19676A. Chronic P. aeruginosa lung infection was established with alginate embedded PAO 579, 19676A or 17107B in 146 Lewis rats which were treated with ceftazidime 4 g/kg intraperitoneally twice a day for 1 week. A statistically significant development of resistance was observed for all three strains. The MICceftazidime of the more resistant variants was increased 15-fold for PAO 579, 8-fold for 19676A, and 4-fold for 17107B, and the specific basal 3-lactamase activity from 19 to 100 units for PAO 579, from 38 to 1,300 units for 19676A, and from 500 to 1,300 units for 17107B. It was shown that, during treatment with ceftazidime, biofilm-growing P. aeruginosa had the capacity to develop resistance due to the production of chromosomal beta-lactamase. PMID- 11110048 TI - Enhancement of lymphocyte proliferation induced by interleukin-12 and anti interleukin-10 in HIV-infected patients during highly active antiretroviral therapy. AB - Based on the potentially important role of IL-10 and IL-12 in the pathogenesis of HIV infection, we have examined the effect of highly active antiretroviral therapy (HAART) on the production of these two cytokines, and whether addition of IL-12 or anti-IL-10 in vitro could improve the proliferative response in peripheral blood mononuclear cells (PBMC) from HIV-infected patients during such therapy. Our findings are: (i) After initiating HAART there were no significant changes in PHA- or MAC-PPD-stimulated IL-10 and IL-12 levels in PBMC supernatants in the patient group as a whole. (ii) However, while a decline in IL-10 synthesis was shown in patients with high baseline MAC-PPD- and PHA-stimulated IL-10 levels, IL-10 increased in patients with lower baseline levels. A similar pattern was seen for MAC-PPD-stimulated IL-12 levels. (iii) Exogenously added IL-12 and anti-IL-10 markedly and additively improved MAC-PPD-stimulated PBMC proliferation in vitro. Thus, a loss of cell-mediated immune response exists in HIV-infected patients also during apparently successful HAART and this can be significantly improved by addition of IL-12 and anti-IL-10, at least in vitro. These results suggest that further exploration of both IL-10 and IL-12 as targets for immunomodulating therapy in HIV-infected patients in addition to HAART might be important. PMID- 11110049 TI - V3 sequence analysis and biological characterization of HIV-1 isolates from asymptomatic and early symptomatic Tanzanian individuals. AB - The aim of this study was to determine HIV-1 V3 sequences, in vitro biological characteristics and co-receptor usage of virus isolates from Tanzania. Virus was isolated from 14 of 17 samples investigated. Four of the isolates induced syncytia in MT-2 cells and used the CXCR4 co-receptor, while the remaining 10 isolates used the CCR5 co-receptor characteristic of non-MT-2 tropic viruses. One of the four MT-2 tropic isolates also used the CCR5 and CCR3 co-receptors. Proviral DNA was detected in all 14 isolates and PCR products were subjected to DNA sequencing. Unambiguous V3 amino acid sequences were obtained from 11 amplificates. Phylogenetic analysis indicated that these sequences were divergent and clustered in HIV-1 subtypes A, C or D. Sequences from the viruses that induced syncytia in MT-2 cells presented characteristic V3 phenotype-associated amino acids. Results of co-receptor analysis are in concordance with the isolate phenotype as determined by replication and induction of syncytia in MT-2 cells. The considerable diversity illustrated by a limited number of isolates from Tanzania is in accordance with reports from other regions of Africa. PMID- 11110050 TI - Extrapleural solitary fibrous tumor: clinicopathologic study of 17 cases and molecular analysis of the p53 pathway. AB - Solitary fibrous tumor (SFT) occurring at various extrapleural sites is sometimes difficult to diagnose because of its histologic variability. Although a solitary fibrous tumor is usually a slow-growing tumor with favorable prognosis, a small number of malignant cases have been reported. In the present study, we examined the clinical behavior, histologic, immunohistochemical and molecular features of 17 cases of extrapleural SFT. Four tumors were located in the pelvic cavity, two in the nasal cavity, two were confined to the pulmonary parenchyma, and there was one each in the meninges, kidney, mediastinum, retroperitoneum, temporal region, neck, groin, buttock and thigh. Histologically, all the tumors were characterized by the presence of areas consisting of a proliferation of bland spindle cells with variable amounts of thick, often hyalinized or keloid-like intercellular collagen bundles. Highly cellular areas were observed in three tumors, frequent mitoses in two, and cellular pleomorphism and tumor necrosis in one each. All 17 tumors showed immunoreactivity to CD34 and 15 (88%) to bcl-2 protein. The labeling indices of p53, mdm2 protein and Ki-67 were generally low. PCR-SSCP and a subsequent sequence analysis of the p53 gene disclosed point mutation at codon 161 in exon 5 in one of the 13 cases analyzed. According to follow-up information, none of the patients had developed local recurrence or distant metastasis. Our results suggest that most extrapleural SFTs behave in a benign fashion even in a higher histologic grade group, and it is difficult to predict their clinical outcome. Complete surgical excision in order to obtain clear margins and long-term follow-up is advisable for patients with an extrapleural SFT. PMID- 11110051 TI - A new DIG-PCR-EIA method for the detection of Chlamydia pneumoniae DNA in clinical samples. AB - Chlamydia pneumoniae, a common respiratory pathogen, may also play a role in the pathogenesis of other chronic conditions. For accurate detection of infected persons and verification of results obtained by other PCR methods, a DIG-PCR-EIA method was evaluated. In the DIG-PCR-EIA, a 437 bp DNA sequence was amplified and hybridized with a newly synthesized 229 bp biotin-labeled probe. The end product was detected by an enzyme immunoassay. The sensitivity of DIG-PCR-EIA was compared with Southern blot hybridization and one-step HR/HL PCR, which was the routine method used. DNA was detected to the level of 20 elementary bodies of DIG EIA-PCR compared to less than 2 by Southern blot, and 200 by HR/HL PCR. Thus a 100-fold increase in sensitivity could be expected by DIG-EIA-PCR compared to the routine method. Throat swabs and adenoid tissue from 22 children with otitis and middle ear secretions from 29 children, as well as throat swabs from 179 blood donors, were analyzed with DIG-EIA-PCR, HL/HR PCR and nested touchdown PCR. 32% of the ear secretions were positive by DIG-EIA-PCR as compared to 5% by the other two methods. Three adenoid tissue samples were positive by all methods applied. Among the child and adult throat samples, 18% and 32%, respectively, were positive by DIG-EIA-PCR and 5% and 10% by HR/HLPCR. The results indicate the suitability of DIG-PCR-EIA for verification of results of HR/HL PCR. DIG-PCR-EIA has a potential for increased sensitivity and adaptation for automation. It should be further evaluated using various types of tissue specimens and DNA extraction methods. PMID- 11110052 TI - Prognostic significance of biologic markers in node-negative breast cancer patients: a prospective study. AB - It is generally thought that future advances in the treatment and cure of breast cancer patients will be made possible through a deeper understanding of tumor biology and an improved capability to define the prognosis of each single patient. This will lead to the formulation of new, more selective, and patient tailored therapies. It is therefore important, when studying potential prognostic factors, to follow methodologic requirements and guidelines which involve the carrying out of prospective studies as confirmatory steps. Repeatedly or recently investigated prognostic markers (tumor size, menopausal status, ER, PgR, 3H thymidine labeling index, c-erbB-2 and p27 expression) were evaluated on a series of 286 prospectively recruited node negative breast cancer patients who underwent loco-regional treatment alone and were closely followed. The individual and relative prognostic contribution of each variable with respect to other factors, as well as their ability to identify node negative patients at risk, were assessed by univariate and multivariate analysis. At a five-year follow-up, only tumor size (p = 0.021) and TLI (p = 0.016) individually proved to be significant prognostic indicators of relapse-free survival. Conversely, p27 expression was not related to RFS and c-erbB-2 expression appeared to have only a short-term effect on patient prognosis. TLI and tumor size, tested in multivariate analysis along with ER and menopausal status, maintained their independent prognostic relevance. The study, performed on a large series of node-negative patients given loco-regional treatment alone, for the first time prospectively recruited, showed the prognostic relevance of TLI and its independence from other clinico pathologic and biologic factors over a five-year period. PMID- 11110053 TI - Apolipoprotein E polymorphism and breast carcinoma: correlation with cell proliferation indices and clinical outcome. AB - There is preliminary evidence that polymorphism of apolipoprotein E (apoE, protein; APOE, gene), one of the key regulatory proteins in cholesterol metabolism, influences the pathobiology of carcinoma of the colon, prostate and breast and also primary tumours of the brain. This study was designed to determine whether APOE polymorphism is related to variation in the rate of tumour cell proliferation and clinical outcome in carcinoma of the breast. One hundred and eleven infiltrating ductal carcinomas, for which follow up data were available, were included in the study. Estrogen and progesterone receptor status (ER, PR) cell proliferation index (MIB- 1) and APOE genotypes were determined from paraffin-embedded tissue by standard methods. Positive correlations were found between grade and tumour size, grade and presence of metastasis, grade and MIB-1 expression, as well as between ER and PR. Survival correlated inversely with tumour size and the presence of positive lymph nodes. Both steroid receptors correlated inversely with MIB- 1 expression. PR positive status also correlated inversely with high histological grade and presence of lymph node metastases. APOE allele frequencies resembled those of the general population. No significant associations were found between possession of either APOE epsilon2 or epsilon4 alleles and the parameters investigated. Although there is evidence to suggest that APOE epsilon4 may predispose to the development of carcinoma of the breast our data do not support the hypothesis that APOE genotype influences the rate of tumour cell proliferation or the clinical course. PMID- 11110054 TI - Effects of Bcl-2 modulation with G3139 antisense oligonucleotide on human breast cancer cells are independent of inherent Bcl-2 protein expression. AB - We have investigated the effects of transient Bcl-2 down-regulation induced by the Bcl-2 antisense oligodeoxynucleotide (ODN) G3139 (Genta Incorporated) in high Bcl-2 protein expressing, estrogen receptor (ER) positive MCF-7 and low Bcl-2 expressing, ER negative MDA435/LCC6 human breast cancer cells. Treatment with Bcl 2 antisense ODN in vitro caused > 80% reduction of Bcl-2 protein levels in a sequence specific manner for both cell lines. Maximum mRNA reduction was achieved within 24 h of the first antisense ODN exposure whereas full protein down regulation required antisense exposure over 48 h. This Bcl-2 reduction was associated with 80-95% loss of viable cells compared to untreated cells. Similar cytotoxic effects were observed in both cell lines despite a nine-fold intrinsic difference in Bcl-2 protein expression suggesting that the relative degree of down-regulation of Bcl-2 is more important than the absolute reduction. Cell death associated with G3139 exposure exhibited properties indicative of apoptosis such as mitochondrial membrane depolarization and caspase activation. Combined treatment with G3139 and cytotoxic agents resulted in additive cytotoxicity in both cell lines. However, under most conditions studied, the direct cytotoxic activity of G3139 antisense was not synergistic with the cytotoxic agents. These results suggest that while Bcl-2 clearly constitutes an attractive therapeutic target due to its role in regulating apoptosis in breast cancer cells, additional mechanisms are important in the control of apoptosis arising from exposure to anticancer agents in vitro. PMID- 11110055 TI - Unique features of breast cancer in Taiwan. AB - Between April 1990 and December 1997, 811 consecutive patients with 830 newly diagnosed breast cancers having their primary treatments in our institution were included in this study. Sixty three percent of breast cancer patients were premenopausal. The early-onset breast cancer (age < or = 40) composed 29.3% of all patients. The five-year survival rate of all patients was 80.4% (95% confidence interval [CI], 76.2-84.6%). The five-year overall survival rate for stage 0 was 95.7% (95% CI, 87.3-100%), stage I, 93.9% (95% CI, 88.9-98.9%), stage II, 88.5% (95% CI, 82.0-95.1%), stage III, 65.0% (95% CI, 54.0-75.9%), and stage IV, 18.5% (95% CI, 3.4-33.7%). Multivariate analysis of primary operable breast cancer revealed that axillary lymph node involvement, high nuclear grade and early-onset breast cancer (age < or = 40) were poor prognostic factors. The early onset breast cancer had a more aggressive clinical behavior than that of the older age group, their five-year disease-free survival rates for stage I, stage II and stage III diseases being only 64.7%, 66.5%, and 43.3%, respectively. In these patients the only meaningful prognostic factor was extensive axillary lymph node metastasis (> or = 10). In summary, breast cancer patients in Taiwan tend to be younger than their counterpart in western countries. The early-onset breast cancer had poorer prognostic features for all stages comparing to the older age group. Standard pathologic factors are not good predictors of their outcome. For these patients new biologic markers need to be sought to distinguish between high and low risk and the treatment strategy for them should be guided by the aggressive characteristics of the disease. PMID- 11110056 TI - Serum lipid levels during and after adjuvant toremifene or tamoxifen therapy for breast cancer. AB - Tamoxifen decreases serum cholesterol (S-cholesterol) level about 10% and low density lipoprotein cholesterol (S-LDL) 15-20%, but in most studies it has increased serum triglyceride levels and had little effect on serum high-density cholesterol (S-HDL). The effect of another antiestrogen, toremifene, on the serum lipid profile has not been completely studied. We monitored serum lipid levels longitudinally in 141 axillary node-positive postmenopausal breast cancer patients who received randomly either 40 mg toremifene or 20 mg tamoxifen as adjuvant therapy for 36 months, and in 34 postmenopausal women who received no adjuvant systemic therapy after surgery for axillary node-negative breast cancer. No significant differences were found between the drugs in their effects on S cholesterol, LDL, HDL, or triglyceride levels, or on the cholesterol-to-HDL or LDL-to-HDL ratios. For both drugs the S-cholesterol and S-LDL absolute lowering effect was the greater the higher the pretreatment level. For a patient with a median pretreatment value, toremifene decreased S-cholesterol 6% and tamoxifen 13%, and S-LDL decreased by 13% and 23%, respectively, at 6 months of therapy. Six months after stopping three-year antiestrogen therapy S- cholesterol and S LDL levels had returned to the pretreatment levels. In conclusion, we found no major differences between 40 mg toremifene and 20 mg of tamoxifen in their effect on the serum lipid levels, which return to the pretreatment levels within 6 months after cessation of therapy. PMID- 11110057 TI - Treatment of advanced, refractory breast cancer with alternating docetaxel and epirubicin/cyclophosphamide plus human granulocyte colony-stimulating factor. AB - PURPOSE: A phase II study was performed to investigate the efficacy and tolerance of alternating docetaxel and epirubicin/cyclophosphamide plus recombinant human granulocyte colony-stimulating factor (G-CSF) in patients with advanced breast cancer who failed previous non-anthracycline/taxane-containing palliative chemotherapy. PATIENTS AND METHODS: Between November 96 and June 98, a total of 45 patients participated in this trial. Chemotherapy consisted of docetaxel 100 mg/m2 given as a 1-h infusion on day 1, and epirubicin 100 mg/m2 plus cyclophosphamide 800 mg/m2 both administered on day 21. G-CSF 5 microg/kg/day was given subcutaneously from days 22-28 during each cycle. Treatment courses were repeated every 42 days for a total of three courses unless prior evidence of progressive disease. RESULTS: The overall response rate was 57.8% (95% confidence interval, 42.1-72.3%), including seven complete (15.5%) and 19 partial remissions (42.3%); nine patients (20%) had stabilization of disease and 10 (22.3%) progressed. The median time to treatment failure was 7.0 months (range 1.5-26.0), and the median overall survival time 15.0 months (range 2.0-37.0+) with 12 patients (27%) currently still alive with metastatic disease. Myelosuppression was commonly observed with WHO grade 3/4 neutropenia in 20 patients (44%) complicated by septicemia in five (11%). Severe nonhematologic toxicity included stomatitis in five patients (11%), skin and peripheral neurotoxicity each in one patient; alopecia was seen in all 45 patients with complete hair loss in 26 (58%). CONCLUSIONS: Our data suggest that alternating docetaxel and epirubicin/cyclo-phosphamide plus G-CSF is an effective and tolerable second-line combination regimen for the treatment of advanced breast cancer. PMID- 11110058 TI - Biphasic effect of medroxyprogesterone-acetate (MPA) treatment on proliferation and cyclin D1 gene transcription in T47D breast cancer cells. AB - While progesterone is a known differentiation-inducing factor in the human endometrium, for the breast epithelium both proliferation-inducing and inhibiting effects have been described. Cyclin D1, which is required for cell cycle progression in G1 and has been shown to play an important role in the pathogenesis of breast cancer has been implicated as a possible mediator of such effects. In the present study we thus investigated the effects of the progestin agonist MPA (medroxy-progesterone acetate) on proliferation of T47D breast cancer cells. In parallel experiments, the regulation of the human cyclin D1 promoter as well as cyclin D1 protein levels under the influence of MPA were studied. Our results show an increase of proliferative activity in T47D cells after 24 and 48 h of MPA treatment followed by inhibition of proliferation after 72 h. In Western blot analysis an increased expression level of cyclin D1 protein can be observed after 24h of MPA stimulation, while at 72h the protein levels are barely detectable. Transient transfection experiments with a luciferase reporter plasmid containing the human cyclin D1 promoter showed an induction of the promoter after 24 and 36h of MPA treatment followed by a reduction in promoter activity. In conclusion, our results confirm the existence of a biphasic response of T47D cell proliferation in response to MPA treatment, consisting of stimulation of proliferation followed by inhibition, and further implicate cyclin D1 as a mediator of these effects, since the cyclin D1 promoter shows a similar biphasic response in this context. PMID- 11110059 TI - Treatment with the pure antiestrogen faslodex (ICI 182780) induces tumor necrosis factor receptor 1 (TNFR1) expression in MCF-7 breast cancer cells. AB - Apoptosis induction by the pure antiestrogen faslodex, also known as ICI 182780 (ICI), is associated with an effective down-regulation of Bcl-2 expression in the human breast cancer cell line MCF-7. Recent observations point out that beside members of the Bcl-2 family also the TNFR1 signaling pathway may be involved in apoptosis induction by antiestrogens. In this report we have analyzed the expression of members of the TNFR1 signaling pathway during the apoptotic process induced by the pure antiestrogen faslodex and by tamoxifen (Tam) in MCF-7 breast cancer cells. Treatment with 10(-7) M ICI or 10(-7) M Tam leads to a time dependent increase of TNFR1 and TRADD steady-state mRNA levels in MCF-7 cells. In contrast, Bcl-2 expression was strongly decreased following administration of ICI but only weakly after administration of Tam. Western blot analysis and studies by the use of fluorescence microscopy and flow cytometry revealed a time dependent induction of TNFR1 protein and cell surface expression in MCF-7 cells in response to treatment with ICI. To investigate if TNFR1 is functionally involved in apoptosis induction by antiestrogens, we tested whether TNFR1 blocking antibodies can counteract the growth inhibitory action of Tam and ICI. Coincubation of MCF-7 cells with antiestrogens (ICI or Tam) and blocking TNFR1 antibodies lead to an increase in cell viability. Our results provide evidence for a cross talk between the TNFR1 signaling pathway and antiestrogens during the process of apoptosis induction in MCF-7 breast cancer cells. The superiority of the pure antiestrogen ICI to induce apoptosis in MCF-7 cells may result from its capability to modulate the induction of apoptosis via Bcl-2 as well as TNF-associated signal transduction pathways. PMID- 11110060 TI - Female smokers have increased postoperative narcotic requirements. AB - This study investigated the influence of tobacco use on postoperative narcotic requirements of female patients following pelvic surgery. The history of tobacco use was taken by telephone survey, and the amount of postoperative narcotic used was obtained from a retrospective review of the patients' hospital charts. Postoperative narcotic use for patients who never smoked was 10.9 mg/12 hr (n = 83, S.E. = 0.5), for former smokers was 13.0 mg/12 hr (n = 33, S.E. = 0.8) and for current smokers was 13.1 mg/12 hr (n = 53, S.E. = 0.7). Patients who never smoked used significantly less narcotic than former smokers (p = .02) or current smokers (p = .007). There was no difference between current and former smokers. Patients who have smoked required more narcotic for postoperative pain control. This effect was equally strong for former as for current smokers. PMID- 11110061 TI - The association between cigarette smoking and drug abuse in the United States. AB - Cigarette smoking has been identified as an independent risk factor for many human diseases. However, the association between cigarette smoking and illegal drug use has not been thoroughly investigated. We have analyzed the 1994 National Household Survey on Drug Abuse to clarify whether cigarette smoking has any effect on the initiation of illegal drug use. Data from 17,809 respondents completing the 1994 "new" (1994-B) questionnaire were analyzed. Logistic regression analyses were performed with the use of statistical package SUDAAN, taking into consideration the multistage sampling design. The results show that those who had smoked cigarettes were far more likely to use cocaine (OR = 7.5; 95% CI: 5.7-9.9), heroin (OR = 16.0; 95% CI: 6.8-37.9), crack (OR = 13.9; 95% CI: 7.9-24.5) and marijuana (OR = 7.3; 95% CI: 6.2-8.7). The associations are consistent across age-strata and remain after adjusting for race and gender. This study suggests that cigarette smoking may be a gateway drug to illegal drug use. PMID- 11110062 TI - Selected secondhand smoke research: summary and comment. AB - Secondhand smoke is one of the more controversial public health issues. It is controversial because laws regulating secondhand smoke create conflict between the rights of smokers and non-smokers. The results of secondhand smoke research frequently focus on risk factors in four areas: heart disease, cancer, respiratory disorders, and middle ear discase. While many studies have found hazards in each of these four areas, there is some disagreement regarding the degree and extent of the threat posed by these hazards. Future research should discover more risks associated with secondhand smoke and suggest appropriate educational, medical, legal, and environmental remedies for this problem. Then society can establish prevention programs and enact laws which protect non smokers, but at the same time infringe as little as possible on the rights of others. PMID- 11110063 TI - A case for addressing cigarette use in methadone and other opioid treatment programs. AB - Most persons attending drug treatment smoke cigarettes. They will eventually experience predictable, but devastatingly high, tobacco-related mortality. Recent studies indicate that many clients are ready to quit smoking and that quitting does not threaten progress made in treatment. Methadone and other opioid treatment providers are in an excellent position to address tobacco use among their clients. The present paper describes the prevalence of smoking among methadone clients, reviews promising interventions, and describes how programs can implement a systematic approach to smoking cessation that includes creating a cue system for identifying smokers, providing brief on-site intervention, and arranging follow-up or more intensive treatment. PMID- 11110064 TI - Ultrarapid opioid detoxification of two children with congenital heart disease. AB - The cases of two children with congenital heart disease and severe opioid dependency who underwent ultrarapid opioid detoxification are presented. This technique entails rapid opioid reversal with the opioid antagonist naloxone while under general anesthesia. The procedures were not technically difficult to perform and both children were successfully detoxified at the end of the procedure. In the weeks following the procedure, the first child exhibited accelerated neurodevelopment. Ultrarapid opioid detoxification is possible in children and may have a neurodevelopmental advantage over a prolonged wean. PMID- 11110065 TI - Plasma naltrexone during opioid detoxification. AB - Orogastric naltrexone is used for opioid detoxification, but it is not known how gastric absorption affects plasma concentrations of the drug. We measured plasma naltrexone during orogastric naltrexone, given in repeated doses of 12.5 mg, 25 mg, 50 mg and 50 mg. Plasma naltrexone was measured after each naltrexone dose. The increase in plasma naltrexone was highly variable between patients during orogastric administration. Adequate detoxification was questioned in 4 of 10 patients because plasma naltrexone did not increase above 5 ng/ml. There was a negative correlation between plasma naltrexone and the presence of withdrawal symptoms on the day after the procedure (r = -0.78, P < 0.05). These results show that the increase in plasma naltrexone is variable during orogastric administration and this may impair successful detoxification. PMID- 11110066 TI - Influence of psychiatric comorbidity on HIV risk behaviors: changes during drug abuse treatment. AB - This study evaluated whether psychiatric comorbidity is related to change in HIV high risk behaviors during outpatient drug abuse treatment. Participants were opioid abusers entering methadone treatment. Psychiatric and substance use diagnoses were determined at intake. Information on HIV high risk drug use and sexual behaviors, psychosocial functioning, and urine toxicology was assessed at intake and at month six. Subjects were divided into those with versus without a lifetime comorbid non-substance use psychiatric disorder. The comorbid group reported more injection equipment sharing, lower rates of condom use, and higher rates of alcohol use at intake and follow-up. Overall injection drug use behavior decreased over the follow-up period for both groups, however. Methadone treatment had a beneficial effect on HIV risk behaviors, and though some risk behaviors improved signiticantly for both groups, comorbid subjects continued to have higher rates of HIV risk factors than noncomorbid subjects. PMID- 11110067 TI - Selective guide to current reference sources on topics discussed in this issue. PMID- 11110068 TI - ASAM-AMBHA joint statement on credentialing and privileging. American Society of Addiction Medicine and American Managed Behavioral Healthcare Association. PMID- 11110069 TI - Public policy statement on opioid antagonist agent detoxification under sedation or anesthesia (OADUSA). American Society of Addiction Medicine (ASAM). PMID- 11110070 TI - Improving the imaging report in patients with acute intracerebral hemorrhage. PMID- 11110071 TI - Diffusion-weighted MRI in cystic or necrotic intracranial lesions. AB - Our purpose was to investigate the signal intensities of cystic or necrotic intracranial lesions on diffusion-weighted MRI (DWI) and measure their apparent diffusion coefficients (ADC). We examined 39 cystic or necrotic intracranial lesions in 33 consecutive patients: five malignant gliomas, seven metastases, two other necrotic tumours, a haemangioblastoma, three epidermoids, an arachnoid cyst, seven pyogenic abscesses, 12 cases of cysticercosis and one of radiation necrosis. DWI was performed on a 1.5 T unit using a single-shot echo-planar spin echo pulse sequence with b 1,000 s/mm2. The signal intensity of the cystic or necrotic portion on DWI was classified by visual assessment as markedly low (as low as cerebrospinal fluid), slightly lower than, isointense with, and slightly or markedly higher than normal brain parenchyma. ADC were calculated in 31 lesions using a linear estimation method with measurements from b of 0 and 1,000 s/ mm2. The cystic or necrotic portions of all neoplasms (other than two metastases) gave slightly or markedly low signal, with ADC of more than 2.60 x 10(-3) mm2/s. Two metastases in two patients showed marked high signal, with ADC of 0.50 x 10(-3) mm2/s and 1.23 x 10(-3) mm2/s, respectively. Epidermoids showed slight or marked high signal, with ADC of less than 1.03 x 10(-3) mm2/s. The arachnoid cyst gave markedly low signal, with ADC of 3.00 x 10(-3) mm2/s. All abscesses showed marked high signal, with ADC below 0.95 x 10(-3) mm2/s. The cases of cysticercosis showed variable signal intensity; markedly low in five, slightly low in three and markedly high in four. PMID- 11110072 TI - Detectability and detection rate of acute cerebral hemisphere infarcts on CT and diffusion-weighted MRI. AB - Our purpose was to compare the detectability and detection rate of acute ischaemic cerebral hemisphere infarcts on CT and diffusion-weighted MRI (DWI). We investigated 32 consecutive patients with acute hemisphere stroke with unenhanced CT and DWI within 6 h of stroke onset. The interval between CT and DWI ranged from 15 to 180 min (mean 60 min). Infarct detectability on CT and DWI was determined by comparing the initial CT, DWI and later reference images in a consensus reading of five independent examiners. The "true" detection rate was assessed by analysing all single readings. Two patients had intracerebral haematomas on DWI and CT and were excluded. There were 27 patients with ischaemic infarcts; all were visible on DWI and proven by follow-up. DWI was negative in three patients without a final diagnosis of infarct (100% sensitivity, 100% specificity, chi2 = 30, P < 0.0001). Ischaemic infarcts were visible on 15 and not seen on 12 CT studies (55 % sensitivity, 100% specificity, chi2 = 1.48, P = 0.224). With regard to the single readings (30 examinations x 5 examiners = 150 readings), 63 CT readings were true positive and 72 false negative (sensitivity 47 %, specificity 86%, chi2 = 2.88, P = 0.089). Of the DWI readings 128 were true positive and 7 false negative (sensitivity 95%, specificity 87 %, chi2 = 70.67, P < 0.0001). Interobserver agreement was substantial for CT (chi = 0.72, 95 % confidence interval, 0.6-0.84) and DWI (chi = 0.82, 95 % confidence interval, 0.46-1). Taken together, detectability and detection rate of acute (< 6 h) hemisphere infarcts are significantly higher with DWI than with CT. PMID- 11110073 TI - Diffusion-weighted MRI suggests the coexistence of cytotoxic and vasogenic oedema in a case of deep cerebral venous thrombosis. AB - We report a 20-year-old woman who suffered headaches before presenting with a state of fluctuating vigilance. MRI showed diffuse high signal in the basal ganglia bilaterally on diffusion- and T2-weighted images, which had areas of both low and high apparent diffusion coefficient, presumed to correspond to cytotoxic and vasogenic oedema. MR venography showed no flow in the deep cerebral veins or straight sinus. Heparin was given, with clinical recovery. On follow-up MRI, the appearances became normal. PMID- 11110074 TI - Bilateral thalamic glioma: case report. AB - We report a 63-year-old man who had a rare bilateral thalamic glioma. He complained of difficulty with calculations and had mental deterioration. T1 weighted images revealed bilateral thalamic swelling with homogeneous low signal and no contrast enhancement. The tumour, showing decrease of N-acetylaspartate and the presence of lactate on magnetic resonance spectroscopy, was diagnosed as an astrocytoma by stereotactic biopsy. PMID- 11110075 TI - Leptomeningeal myelomatosis mimicking a subdural haematoma. AB - We report a rare appearance at presentation of meningeal myelomatosis without bone involvement, in the form of an extraaxial mass of mixed density, resembling a chronic subdural haematoma. PMID- 11110076 TI - Primary central nervous system immunocytoma: MRI and spectroscopy. AB - We report on a young woman with a primary cerebral immunocytoma. Most primary cerebral nervous system lymphomas (PCNSL) are highly malignant undifferentiated B cell tumours, there are few data on the clinical course, MRI and spectroscopy findings of this rare PCNSL subtype. MRI revealed a radially enhancing tumour with mild perifocal oedema. MR spectroscopy indicated low cell turnover. Slow clinical progression, no significant changes with treatment, and imaging findings were consistent with a low-grade malignant tumour. PMID- 11110077 TI - Traumatic brachiocephalic pseudoaneurysm presenting with delayed stroke: case report. AB - We report a traumatic pseudoaneurysm of the internal carotid artery bifurcation and subclavian artery with recurrent strokes events in a 19-year-old man. He was admitted with an acute left hemiparesis. His history revealed a similar episode 1 year and a major car accident 3 years previously. Contrast enhanced MR angiography confirmed colour Doppler sonographic findings of a carotid and subclavian artery pseudoaneurysm presumably resulting from seat-belt trauma. The pseudoaneurysm, containing thrombus, was thought to be the source of artery-to artery embolism. PMID- 11110078 TI - An arachnoid granulation in the straight sinus. AB - We report CT, MRI and angiographic findings of an arachnoid granulation in the straight sinus in a young man. Its density and signal intensity were isodense and isointense with cerebrospinal fluid on CT and MRI, respectively. The lesion appeared as a filling defect on MR venography and conventional angiography. PMID- 11110079 TI - MRI and 18F-fluorodeoxyglucose positron emission tomography in hemimegalencephaly. AB - We report hemimegalencephaly in a 44-year-old woman with mental retardation, epilepsy and a mild hemiparesis. In addition to typical findings on MRI, 2-deoxy 2[18F]fluorodeoxyglucose positron-emission tomography (PET) demonstrated glucose hypometabolism of the affected hemisphere. The results of PET have been coregistered with morphological information from the MRI studies by image fusion. PMID- 11110080 TI - MRI in tick-borne encephalitis. AB - The tick-borne encephalitis (TBE) virus gives rise to epidemic encephalitis. Mild forms usually manifest as influenza-like episodes or are clinically silent. MRI is usually normal in TBE. We describe severe TBE in a patient who presented with fever and altered mental status after a tick bite and a specific antibody response to TBE. MRI revealed pronounced signal abnormalities in the basal ganglia and thalamus, without contrast enhancement. These findings coincide well with neuropathological studies of severe nerve cell degeneration with inflammatory cell infiltrates, neuronophagia and reactive astrocytosis in the deep grey matter. We review the literature and discuss the relevant differential diagnosis. PMID- 11110081 TI - Visibility of the central canal on MRI. AB - The central canal of the spinal cord is present at birth and becomes progressively obliterated. Cadaver studies have shown that it may persist partially or completely. To our knowledge, this entity has not been described on MRI. We reviewed 794 MRI studies of the spinal cord, and found 12 patients (aged 14 to 65 years) who had an intramedullary cavity. The cavity was at the junction of the ventral 1/3 and dorsal 2/3 of the spinal cord, except at the level of the lumbar enlargement, where it was central. It was filiform in most cases, although sometimes fusiform (3 to 4 mm in diameter), and had regular contours. The cavity were thoracic in 69 % of cases. The clinical features were totally unrelated to the image, and there were no anatomical factors (Chiari malformation, dysraphism) predisposing to syringomyelia. The images were perfectly compatible with a persistent central canal, which we interpret as a variant of normal anatomy. Therefore it is important to regard these findings as normal, to avoid unnecessary treatment and follow-up. PMID- 11110082 TI - A computerised relational database for auditing endovascular treatment of patients with intracranial aneurysms. AB - We describe the development and design of a database for auditing patients with intracranial aneurysms and their endovascular treatment. The database has been used since 1992. Our department's version now contains records of over 800 patients and well over 1,000 aneurysms. The advantages of a relational database for this type of audit are discussed. Copies of the software can be obtained free of charge from the authors. PMID- 11110083 TI - Transarterial embolisation of complex cavernous sinus dural arteriovenous fistulae with low-concentration cyanoacrylate. AB - We report the effectiveness of low-concentration n-butyl-2-cyanoacrylate (NBCA) Lipiodol-tungsten mixture (10-15 %) in the management of patients with aggressive or recurrent complex cavernous dural arteriovenous fistulae (CSDAVF). We treated five patients with complex CSDAVF with a low concentration of an NBCA-Lipiodol tungsten mixture after catheterisation of the feeding arteries arising from the external carotid artery. Three had a recurrent CSDAVF after transarterial particulate embolisation. Three refused transvenous treatment or could not be treated in this way; two patients had also feeding dural branches of the internal carotid artery. All patients had complete resolution of symptoms and signs within a month of the procedure. No definite neurological complication was found during follow-up ranging from 12 to 36 months. Transarterial embolisation with low concentration cyanoacrylate appears to be an effective alternative management of aggressive or recurrent CSDAVF. PMID- 11110084 TI - Multiple dural arteriovenous fistulae involving the cavernous and sphenoparietal sinuses. AB - A 72-year-old woman who presented with a unilateral oculomotor nerve palsy was shown to have a very rare condition: multiple dural arteriovenous fistulae (DAVF) involving the cavernous and sphenoparietal sinuses. The sphenoparietal DAVF was cured completely by transarterial embolisation. Symptomatic relief was accomplished by this procedure. The cavernous sinus DAVF progressed to acquire cortical venous drainage, and was obliterated completely by transvenous embolisation. PMID- 11110085 TI - Transfer of internal carotid artery thrombus to the external carotid artery using a balloon microcatheter. AB - We report a case of acute occlusion of the left internal carotid artery successfully treated by withdrawing a large clot into the external carotid artery using a balloon microcatheter and urokinase. This technique may not only reduce the amount of thrombolytic agent required but also lower the incidence of haemorrhagic transformation. PMID- 11110086 TI - De novo development of presumed cavernomas following resolution of E. Coli subdural empyemas. AB - Cavernomas fall within the group of angiographically occult lesions and may be found in up to 4 % of the population [1]. They may occur at any age, and with the advent of MRI incidental cavernomas are increasingly identified. The pathogenesis is uncertain. Familial cases are well recognised with a reported prevalence of 10 15 % [2-3]. The incidence of new lesions has been reported at 0.4 lesions per patient per year in cases with familial cavernomas [4]. Presumed cavernomas have been documented following radiation for malignancy [5-6], and stereotactic cerebral biopsy [7]. There have been no previously documented cases of de novo genesis of cavernomas following bacterial meningitis and subdural empyemas. PMID- 11110087 TI - Wernicke's encephalopathy induced by total parenteral nutrition in patient with acute leukaemia: unusual involvement of caudate nuclei and cerebral cortex on MRI. AB - We report a 13-year-old girl with leukaemia and Wernicke's encephalopathy induced by total parenteral nutrition. MRI showed unusual bilateral lesions of the caudate nuclei and cerebral cortex, as well as typical lesions surrounding the third ventricle and aqueduct. After intravenous thiamine, the patient improved, and the abnormalities on MRI disappeared. PMID- 11110088 TI - Organization, management and operation of contemporary academic mass spectrometry service facilities. AB - The rapid evolution of mass spectrometry in the past 15 years has moved mass spectrometry facilities from the traditional model in which instruments were located in and used for a single department's samples to a distributed model servicing entire universities. In this paper we describe two such shared instrument facilities that have evolved from a base in a single department to facilities that service a broad clientele. The Purdue University Campus-wide Mass Spectrometry Center (CWMSC) is a decentralized facility with multiple sites on campus. The CWMSC is a limited-access facility in which samples are run by service facility personnel in close cooperation with investigators. The Vanderbilt University Mass Spectrometry Research Center (VU-MSRC) is a centralized facility in the medical school that provides services to the university at large. The VU-MSRC is an open-access facility in which users are expected to prepare and analyze their own samples under the guidance of a trained operator. Perhaps the most significant benefit achieved by these models has been the minimization of academic barriers and the resultant intellectual cross fertilization that has greatly enriched research at institutions where this approach has been adopted. The advantages and limitations of both models are discussed in terms of the traditional academic paradigm of service, research and education. PMID- 11110089 TI - Studies on the dephosphorylation of phosphotyrosine-containing peptides during post-source decay in matrix-assisted laser desorption/ionization. AB - Phosphorylation of tyrosine residues in proteins is a common regulatory mechanism, although it accounts for less than 1% of the total O-phosphate content in proteins. Whereas aromatic phosphorylation sites can be identified by a number of different analytical techniques, sequence analysis of phosphotyrosine containing proteins at the low picomole or even femtomole level is still a challenging task. This paper describes the post-source decay in matrix-assisted laser desorption/ionization mass spectrometry of phosphotyrosine-containing model peptides by comparing their fragmentation behavior with sequence-homologous unphosphorylated peptides. Whereas the parent ions showed significant losses of HPO3, all phosphorylated fragment ions of the b- and y-series displayed only minor dephosphorylated signals, which often were not detectable. Surprisingly, one of the studied phosphotyrosine-containing sequences displayed, in addition to the [M + H - 80]+ ion, a more abundant [M + H - 98]+ ion, which could be explained by elimination of phosphoric acid. This dephosphorylation pattern was very similar to the patterns obtained for phosphoserine- and phosphothreonine containing peptides. Because the dephosphorylation pattern of the parent ion is often used to identify modified amino acids in peptides, we investigated possible dephosphorylation mechanisms in detail. Therefore, we substituted single trifunctional amino acid residues and incorporated deuterated phosphotyrosine residues. After excluding direct elimination of phosphoric acid from tyrosine, we could show that the obtained loss of H3PO4 depends on aspartic acid and arginine residues. Most likely the HPO3 group is transferred to aspartic acid followed by cleavage of phosphoric acid forming a succinimide. On the other hand, arginine appears to induce the H3PO4 loss by protonation of phosphotyrosine leaving a phenyl cation. PMID- 11110090 TI - Collision-induced fragmentation of underivatized N-linked carbohydrates ionized by electrospray. AB - The electrospray mass spectra and collision-induced fragmentation of neutral N linked glycans obtained from glycoproteins were examined with a Q-TOF mass spectrometer. The glycans were ionized most effectively as adducts of alkali metals, with lithium providing the most abundant signal and caesium the least. Singly charged ions generally gave higher ion currents than doubly charged ions. Addition of formic acid could be used to produce [M + H]+ ions, but these ions were always accompanied by abundant cone-voltage fragments. The energy required for collision-induced fragmentation was found to increase in a linear manner as a function of mass with the [M + Na]+ ions requiring about four times as much energy as the [M + H]+ ions for complete fragmentation of the molecular ions. Fragmentation of the [M + H]+ ions gave predominantly B- and Y-type glycosidic fragments whereas the [M + Na]+ and [M + Li]+ ions produced a number of additional fragments including those derived from cross-ring cleavages. Little fragmentation was observed from the [M + K]+ and [M + Rb]+ ions and the only fragment to be observed from the [M + Cs]+ ion was Cs+. The [M + Na]+ and [M + Li]+ ions from all the N-linked glycans gave abundant fragments resulting from loss of the terminal GlcNAc moiety and prominent, though weaker, ions as the result of 0,2A and 2,4A cross-ring cleavages of this residue. Most other ions were the result of successive additional losses of residues from the non-reducing terminus. This pattern was particularly prominent with glycans containing several non-reducing GlcNAc residues where successive losses of 203 u were observed. Many of the ions in the low-mass range were products of several different fragmentation routes but still provided structural information. Possibly of most diagnostic importance was an ion formed by loss of 221 u (GlcNAc molecule) from an ion that had lost the 3-antenna and the chitobiose core. This latter ion, although coincident in mass with some other 'internal' fragments, often provided additional information on the composition of the antennae. Other ions defining antenna composition were weak cross-ring fragments produced from the core branching mannose residue. Glycans containing Gal-GlcNAc residues showed successive losses of this moiety, particularly from the B-type fragments resulting from loss of the reducing-terminal GlcNAc residue. The [M + Na]+ and [M + Li]+ ions from high-mannose and hybrid glycans gave a series of ions of composition (Man)nNa/Li+ where n = 1 to the total number of glycans in the molecule, allowing these sugars to be distinguished from the more highly processed complex glycans. Other ions in the spectra of the high-mannose glycans were diagnostic of chain branching but insufficient information was available to determine their mode of formation. PMID- 11110092 TI - Analysis of polar hydrophilic aromatic sulfonates in waste water treatment plants by CE/MS and LC/MS. AB - The present work describes the development and optimization of a capillary (zone) electrophoresis/mass spectrometric (CE/MS) analysis method for polar hydrophilic aromatic sulfonates (ASs). The compounds were detected by negative ion electrospray ionization (NIESI) and selected ion monitoring (SIM). In comparison with CE/UV, for CE/MS a lower-concentration volatile ammonium acetate buffer (5 mM) without organic modifier and a higher separation voltage were better suited for separation. Sensitivity of CE/MS was slightly better than for CF/UV, with the limit of detection (LOD) ranging between 0.1 and 0.4 mg l(-1). For verification of the CE/MS results, ASs were also analysed by ion-pair liquid chromatography/diode array UV detection coupled in series with electrospray mass spectrometry (IPC/DAD/ESI-MS). Real water samples of different waste water treatment plants (WWTPs) in Catalonia (NE Spain) were extracted by solid-phase extraction (SPE) with LiChrolut EN and analysed with CE/MS and LC/MS. ASs were found in influent and effluent water samples of the WWTPs in the microg l(-1) concentration range. LC/MS offered a higher separation efficiency and sensitivity than CE/MS. Therefore with LC/MS more compounds could be identified in the WWTPs. The persistency of the ASs was distinct: some compounds were well degraded during the water treatment process, while others were quite persistent. PMID- 11110096 TI - Current literature in mass spectrometry. PMID- 11110093 TI - Study on fragmentation behavior of 5/7/6-type taxoids by tandem mass spectrometry. AB - The mass spectrometric behaviors of seven compounds, namely four 4beta(20),5 oxetane 5/7/6-type taxoids (i.e. taxayuntin A, taxayuntin B, taxayuntin and taxayuntin C) and three 4(20)-methylene 5/7/6-type taxoids (i.e. brevifoliol, taxchinin A and 7-acetyl-10-deacetyl-7-debenzoylbrevifoliol) have been investigated by the positive ion FAB-MS/MS technique. The fragmentation has been correlated with the types and positions of substituents of these compounds. It has been found that with the OH group at the C-10 position, taxayuntin A, taxayuntin B and 7-acetyl-10-deacetyl-7-debenzoylbrevifoliol are dominated by the loss of H2O, while with the BzO group at the C-10 position, taxayuntin, taxayuntin C, brevifoliol and taxchinin A preferentially eliminate the BzO group. In addition, C-2 is an active site, and neutral loss from the C-2 position readily occurs. The four 4beta(20),5-oxetane 5/7/6-type taxoids produce the terminal product ion with a stable conjugated system at m/z 311, while the 4(20) methylene 5/7/6-type taxoids brevifoliol and 7-acetyl-10-deacetyl-7 debenzoylbrevifoliol produce this ion at m/z 237, and taxchinin A at m/z 253. Interestingly, characteristic fragment ions involving the loss of a 118 u group were observed for taxayuntin, and a possible fragmentation mechanism is given. The major fragmentation pathways and mechanisms of ion formation for the compounds are proposed on the basis of CID spectra and accurate mass measurements. The results of this paper will be helpful for structural analysis of analogs. PMID- 11110097 TI - The binding and distribution of albendazole and its principal metabolites in Giardia duodenalis. AB - Trophozoites of the protozoan parasite Giardia duodenalis were exposed to various albendazole concentrations for 4 h, washed, fixed and incubated with antibodies raised against albendazole and its two major metabolites albendazole sulphoxide and albendazole sulphone. Tubulin antibodies were also used. A peroxidase- or FITC-conjugated secondary antibody was used to detect the primary antibody with transmission electron microscopy or confocal laser scanning microscopy, respectively. Albendazole, a benzimidazole compound, was detected in the mid dorsal region of trophozoites, albendazole sulphoxide in the posterior-dorsal region and albendazole sulphone in clusters above the median bodies. Tubulin was recognised in the ventral disk. This is the first indication that G. duodenalis may be capable of metabolising albendazole and the potential path of the metabolised drug traced within the trophozoite. Fluorescence measurements revealed that albendazole sulphoxide binding decreased and albendazole sulphone binding increased with exposure of the trophozoites to increasing albendazole concentration. This indicates that if albendazole was being metabolised by trophozoites, it occurred to a greater extent following exposure to higher albendazole concentrations. PMID- 11110098 TI - Characterization of urinary metabolites of testosterone, methyltestosterone, mibolerone and boldebone in greyhound dogs. AB - Androgenic steroids are used in female greyhound dogs to prevent the onset of estrus; moreover, these steroids also have potent anabolic activity. As anabolic steroids increase muscle mass and aggression in animals, the excessive use of these agents in racing greyhounds gives an unfair performance advantage to treated dogs. The biotransformation of most anabolic steroids has not been determined in greyhound dogs. The objective of the present study was to identify the urinary metabolites of testosterone, methyltestosterone, mibolerone, and boldenone in greyhound dogs. These steroids were administered orally (1 mg/kg) to either male or female greyhound dogs and urine samples were collected pre administration and at 2, 4, 8, 12, 24, 72, and 96 h post-administration. Urine extracts were analyzed by high-performance liquid chromatography/mass spectrometry (HPLC/MS) to identify major metabolites and to determine their urinary excretion profiles. Major urinary metabolites, primarily glucuronide, conjugated and free, were detected for the selected steroids. Sulfate conjugation did not appear to be a major pathway for steroid metabolism and excretion in the greyhound dog. Phase I biotransformation was also evaluated using greyhound dog liver microsomes from untreated dogs. The identification of several in vivo steroid metabolites generated in this study will be useful in detecting these steroids in urine samples submitted for drug screening. PMID- 11110099 TI - Metabolism of prostaglandin F2alpha during fasting in prepubertal gilts. AB - The study was conducted to investigate a possible mechanism behind earlier observations of fasting-induced increases of blood concentrations of prostaglandin (PG) F2alpha metabolite (P-PG) in gilts. Six animals were fasted for 28 h, then administered i.v. PGF2alpha (500 ng/kg body weight). Blood samples were withdrawn at 1, 3, 5, 7, 10, 15, 20, 30, 40, 60 min, 2 and 3 h after the injection. A control group followed an identical protocol, except that they were fed during the corresponding 28 h-period. P-PG increased as previously observed during the 28 h of fasting. The P-PG increase in terms of area under the concentration time curve (AUC) following injection was significantly larger in the fasted than in the non-fasted gilts. In the fasted animals, the mean P-PG maximum concentration (Cmax) was 7145 pmol/L, the corresponding value for the non fasted animals was 4566 pmol/L. PGs are metabolised through beta-oxidation in the liver. The results of this study imply that reduced 15-ketodihydro-PGF2alpha breakdown in the liver might contribute to the fasting-induced increases in P-PG. PMID- 11110100 TI - Pharmacokinetics of ketoprofen enantiomers after intravenous administration of racemate in camels: effect of gender. AB - The pharmacokinetics of ketoprofen (KP) enantiomers were studied in ten female and eight male camels after a single intravenous dose (2.0 mg/kg) of racemic KP. A high performance liquid chromatographic (HPLC) method was developed for the quantitation of the R- and S-enantiomers without derivatization of the samples using a S,S-Whelk-01 chiral stationary phase column. The data collected (median and range) were as follows: the areas under the curve to infinity (AUC) (microg/mL per h) were 22.4 (13.5-29.7) and 19.8 (13.8-22.1) for R- and S-KP, respectively, in female camels while the corresponding values in male camels were 16.0 (12.9-22.4) and 14.4 (11.0-19.3). In both sexes, the AUC for the R enantiomer was significantly larger than that of the S-enantiomer. Total body clearances (Cl(t)) were 44.6 (33.7-74.1) and 50.6 (45.2-72.4) mL/kg per h for R- and S-KP, respectively, in female camels and were 62.8 (44.6-77.8) and 69.6 (51.8 91.1) mL/kg per h for R- and S-KP, respectively, in male camels. In both sexes of camels, the Cl(t) values for R-KP were significantly lower than its corresponding antipode. The steady-state volumes of distribution (Vss) were 97.9 (82.8-147.2) and 102.0 (90.1-169.0) mL/kg for R- and S-KP, respectively, in female camels and were significantly different from each other, while the respective values in male camels were 151.5 (105.3-222.3) and 154.0 (114.7-229.0) mL/kg but were not significantly different from each other. The volumes of distribution (area) followed a similar pattern, where the values for R- and S-KP in female camels were 118.5 (95.6-195.2) and 137.6 (115.8-236.2) mL/kg, respectively, and the respective values in male camels were 215.6 (119.1-270.1) and 229.1 (143.3-277.4) mL/kg. The elimination half-lives (t1/2beta) were 1.88 (1.42-2.34) h and 1.83 (1.67-2.26) h for R- and S-KP, respectively, in female camels and were significantly different from each other, while the corresponding values in male camels were 2.11 (1.50-4.20) and 2.33 (1.52-3.83) h for R and S-KP, respectively, but were not significantly different from each other. The mean residence time followed a similar pattern. All pharmacokinetic parameters for R- and S-KP in female camels were significantly different from their corresponding values in male camels. The extent of protein binding for R- and S-KP was evaluated in vitro by ultrafiltration. The extents of protein binding for R- and S-KP were not significantly different from each other when each enantiomer was supplemented separately. However, when the enantiomers were supplemented together, protein binding of R-KP was significantly higher than that of S-KP in female but not in male camels. PMID- 11110101 TI - Comparative disposition of tripelennamine in horses and camels after intravenous administration. AB - The pharmacokinetics of tripelennamine (T) was compared in horses (n = 6) and camels (n = 5) following intravenous (i.v.) administration of a dose of 0.5 mg/kg body weight. Furthermore, the metabolism and urinary detection time was studied in camels. The data obtained (median and range in brackets) in camels and horses, respectively, were as follows: the terminal elimination half-lives were 2.39 (1.91-6.54) and 2.08 (1.31-5.65) h, total body clearances were 0.97 (0.82-1.42) and 0.84 (0.64-1.17)L/h/kg. The volumes of distribution at steady state were 2.87 (1.59-6.67) and 1.69 (1.18-3.50) L/kg, the volumes of the central compartment of the two compartment pharmacokinetic model were 1.75 (0.68-2.27) and 1.06 (0.91 2.20) L/kg. There was no significant difference (Mann-Whitney) in any parameter between camels and horses. The extent of protein binding (mean +/- SEM) 73.6 + 8.5 and 83.4 +/- 3.6% for horses and camels, respectively, was not significantly statistically different (t-test). Three metabolites of T were identified in urine samples of camels. The first one resulted from N-depyridination of T, with a molecular ion of m/z 178, and was exclusively eliminated in conjugate form. This metabolite was not detected after 6 h of T administration. The second metabolite, resulted from pyridine ring hydroxylation, had a molecular ion of m/z 271, and was also exclusively eliminated in conjugate form. This metabolite could be detected in urine sample for up to 12 h after T administration. The third metabolite has a suspected molecular ion of m/z 285, was eliminated exclusively in conjugate form and could be detected for up to 24 h following T administration. T itself could be detected for up to 27 h after i.v. administration, with about 90% of eliminated T being in the conjugated form. PMID- 11110102 TI - The impact of acute phase response on the plasma clearance of antipyrine, theophylline, phenytoin and nifedipine in rabbits. AB - The impact of acute phase response (APR) on the plasma clearances of antipyrine, theophylline, phenytoin and nifedipine was studied using 50 female rabbits. APR was induced by a bolus intramuscular injection of Escherichia coli lipopolysaccharide (LPS, 50 microg/kg). No abnormal findings, other than an increase in rectal body temperature and the plasma concentration of interleukin-6 (IL-6), were observed in the LPS-treated animals. Twenty-four hours after LPS injection, the pharmacokinetic parameters of the four drugs were obtained following intravenous administrations of antipyrine (7 mg/kg), theophylline (5 mg/kg), phenytoin (10 mg/kg) and nifedipine (1 mg/kg). Total body clearances of antipyrine, theophylline, phenytoin and nifedipine in LPS-treated rabbits decreased, and terminal elimination half-life and the mean residence time of these drugs increased compared with those in the control rabbits. The apparent volume of distribution for phenytoin and nifedipine increased after the LPS injection, although the binding percentage of the drugs with plasma protein did not change. These results suggested that APR appears to decrease the plasma clearances of these drugs in rabbits, which may be due to the suppression of the activity of cytochrome P450 enzymes. PMID- 11110103 TI - Withdrawal time estimation of veterinary drugs: extending the range of statistical methods. AB - In order to use a drug in a food producing animal, evidence has to be provided that after a certain withdrawal time, drug residues in tissues, such as muscle meat, fat, liver, kidney etc., are below a given maximum residue limit (MRL), for a majority of animals. Several statistical methods, both regression based and nonparametric based methods, have been proposed, each relying on different sets of assumptions, which may or may not hold for the specific data situation. The purpose of this paper is to enrich the range of methods, i.e. to provide approaches for situations where current methods are inappropriate. Bayesian methods, using Markov chain Monte Carlo, are proposed to derive inference on the parameters of interest. PMID- 11110104 TI - Single-dose pharmacokinetics of flumequine in cod (Gadus morhua) and goldsinny wrasse (Ctenolabrus rupestris). AB - Knowledge of the pharmacokinetic properties of drugs to combat bacterial infections in cod (Gadus morhua) and wrasse (Ctenolabrus rupestris) is limited. One antimicrobial agent likely to be effective is flumequine. The aim of this study was to investigate the pharmacokinetic properties of flumequine in these two species. Flumequine was administered intravenously to cod (G. morhua) at a dose of 5 mg/kg bodyweight and wrasse (C. rupestris) at a dose of 10 mg/kg. Flumequine was also administered orally to both species at a dose of 10 mg/kg body weight, and as a bath treatment at a dose of 10 mg/L water for 2 h. Identical experimental designs were used otherwise. The study was performed in seawater with a salinity of 3.2% and a temperature of 8.0 +/- 0.2 degrees C (cod) and 14.5 +/- 0.4 degrees C (wrasse). Pharmacokinetic modelling of the data showed that flumequine had quite different pharmacokinetic properties in cod and wrasse. Following intravenous administration, the volumes of distribution at steady-state (Vss) were 2.41 L/kg (cod) and 2.15 L/kg (wrasse). Total body clearances (Cl) were 0.024 L/hxkg (cod) and 0.14 L/hxkg (wrasse) and the elimination half-lives (t1/2lambda z) were calculated to be 75 h (cod) and 31 h (wrasse). Mean residence times (MRT) were 99 h (cod) and 16 h (wrasse). Following oral administration, the t1/2 lambda z were 74 h (cod) and 41 h (wrasse). Maximal plasma concentrations (tmax) were 3.5 mg/L (cod) and 1.7 mg/L (wrasse), and were observed 24 h post administration in cod and 1 h post-administration in wrasse. The oral bioavailabilities (F) were calculated to be 65% (cod) and 41% (wrasse). Following bath administration, maximal plasma concentrations were 0.13 mg/L (cod) and 0.09 mg/L (wrasse), and were observed immediately after the end of the bath. PMID- 11110105 TI - Single-dose pharmacokinetics of flumequine in the eel (Anguilla anguilla) after intravascular, oral and bath administration. AB - Knowledge of the pharmacokinetic properties of drugs to combat bacterial infections in the European eel (Anguilla anguilla) is limited. One antimicrobial agent likely to be effective is flumequine. The aim of this study was to investigate the pharmacokinetic properties of flumequine in European eels in fresh water. Flumequine was administered to eels (Anguilla anguilla) intravenously (i.v.) and orally (p.o.) at a dose of 10 mg/kg body weight, and as a bath treatment at a dose of 10 mg/L water for 2 h. The study was performed in fresh water with a temperature of 23 + 0.3 degrees C, pH 7.15. Identical experimental designs were used. Two additional bath treatments were also performed, one in which the pH in the water was lowered by approximately 1 unit to 6.07 (dose: 10 mg/L) and one at a dose of 40 mg/L for 2 h in a full-scale treatment. Following i.v. administration, the volume of distribution at steady state was 3.4 L/kg. Total body clearance was 0.012 L/h per kg and the elimination half-life (t1/2lambda z) was calculated to be 314 h. Mean residence time was 283 h. Following oral administration, the t1/2lambda z was 208 h. Maximal plasma concentration (Cmax) was 9.3 mg/L, at 7 h after administration (Cmax). The oral bioavailability (F) was calculated to be 85%. Following bath administration in 10 mg/L for 2 h, maximal plasma concentration was 2.1 mg/L, observed immediately after the end of the bath. The 'bioavailability' in eel following a 2-h bath treatment was 19.8%. Reducing the pH in the bath to 6.07 produced a maximal plasma concentration of 5.5 mg/L, observed immediately after the end of the bath. The 'bioavailability' was increased to 41% by the lowering of the pH. A similar effect was observed in a full-scale treatment (1 kg eels/L water). The CO2 produced by the eel lowered the pH and increased 'bioavailability' to 35%. PMID- 11110106 TI - The effect of raloxifene on coronary arteries in aged ovariectomized ewes. AB - BACKGROUND: Ovariectomized sheep are a useful model of postmenopausal osteoporosis and other postmenopausal conditions. Estrogen may have a protective effect on the coronary arteries in postmenopausal women. The effects of raloxifene, a selective estrogen receptor modulator, on coronary arteries in aged ovariectomized ewes was investigated. METHODS AND RESULTS: Forty eight aged ewes were randomly assigned to undergo sham surgery (Sham, n = 7), ovariectomy (OVX, n = 10), ovariectomy with estradiol supplementation (OVXE, n = 8), ovariectomy with raloxifene supplementation, 0.02 mg/kg per day (RAL1, n = 10), or ovariectomy with raloxifene supplementation, 0.10 mg/kg per day (RAL2, n = 13). Contrast coronary angiography was performed 6 months after intervention. Diameters of the right main and left anterior descending coronary arteries in the RAL1, RAL2 and Sham groups were not different from each other, but were significantly greater than the OVX and OVXE groups. Intracoronary nitroglycerin did not affect the relationships of the diameters in any group. There were no differences in vascular remodeling between the groups. CONCLUSIONS: The results indicate that raloxifene in this sheep model allows greater dilation of coronary arteries than estrogen. Raloxifene may provide a significant protective functional effect on coronary arteries in postmenopausal heart disease. PMID- 11110107 TI - Effects of danofloxacin and tilmicosin on circulating neutrophils in beef heifers. PMID- 11110108 TI - Subcutaneous bioavailability of levamisole in goats. PMID- 11110109 TI - Functional impairment in COPD patients: the impact of anxiety and depression. AB - The authors examined the relationship between functional status and comorbid anxiety and depression and the relationship between utilization of health care resources and psychopathology in elderly patients with chronic obstructive pulmonary disease (COPD). Elderly male veterans (N = 43) with COPD completed anxiety, depression, and functional status measures. The authors constructed regression models to explore the contribution of COPD severity, medical burden, depression, and anxiety to the dependent variables of functional impairment and health care utilization. Anxiety and depression contributed significantly to the overall variance in functional status of COPD patients, over and above medical burden and COPD severity, as measured by the 8 scales of the Medical Outcomes Study (MOS) 36-item Short Form Health Survey. Surprisingly, medical burden and COPD severity did not contribute significantly to overall variance in functional status. Few patients were receiving any treatment for anxiety or depression. PMID- 11110110 TI - Depression in Korean immigrants with hepatitis B and related liver diseases. AB - The authors evaluated 50 Korean immigrants who had chronic viral hepatitis or who were healthy carriers for the hepatitis B virus in terms of the relationships between their depression scores, psychosocial stressors, social support, and biological markers of dysfunction. All participants completed a questionnaire, describing their worries and concerns, and the shortform of the Beck Depression Inventory (BDI-sf). Hepatic transaminases, albumin levels, and prothrombin times were measured during routine clinic follow-up visits and were abstracted from the medical record. Values recorded within 3 months before and within 3 months after the psychiatric interview were correlated with BDI scores. BDI-sf total scores were significantly associated with transaminase elevations (P<0.001) both before and after BDI-sf administration. BDI scores were not associated with other measures of liver dysfunction or other medical causes of depression. Patients with higher BDI-sf total scores had more psychosocial stressors (P = 0.008) and lower Global Assessment of Functioning (GAF) scores (P = 0.000). PMID- 11110111 TI - Consultation-liaison psychiatrists' management of somatoform disorders. AB - The authors studied interventions recommended by consultation-liaison (C-L) psychiatrists when they diagnosed somatoform disorder prospectively in a cohort of 4,401 consecutive inpatients referred to the C-L psychiatry service of a general teaching hospital, using standardized MICRO-CARES methodology. A DSM-III R somatoform disorder was diagnosed in 2.9%, somatoform pain disorder in 1.4%, conversion disorder in 0.7%, hypochondriasis or somatization disorder undifferentiated/not otherwise specified in 0.6%, and somatization disorder in 0.2%. In 3.4%, somatoform disorder was considered a differential diagnosis. Psychiatric comorbidity included mood disorder (39%), personality disorder (37%), and psychoactive substance use disorder (19%). Recommendations were made about antidepressants in 40% of the patients, anxiolytics in 18%, sedatives in 18%, and antipsychotics in 10%. Psychiatrists recommended the following: more laboratory tests for 14%; additional medical/surgical consultations for 11%; an increase in the vigor of medical treatment for 13%; and psychological treatment for 76%; also they stressed an earlier discharge of 16%. Psychiatrists were more likely to request a prolongation of inpatient stay for patients with comorbid somatoform, mood, anxiety, and personality disorder. Differences in characteristics and treatment of the subgroups tended to be consistent with their constructs and comorbid psychiatric diagnoses. PMID- 11110112 TI - The relationship between pain and depression in a trial using paroxetine in sufferers of chronic low back pain. AB - Previous studies have shown a positive association between pain and depression, though evidence supporting a direct link between these two variables is less robust. Using a placebo-controlled trial, the authors examined the analgesic and antidepressant efficacy of paroxetine (20 mg) in chronic low back pain sufferers. The authors examined the associations among pain, depression, disability, and illness attitudes. Paroxetine showed no effects on pain or depression compared with placebo; however, subjects randomized to paroxetine were more likely to reduce concomitant analgesic medication. The cross-sectional association of depression and pain at baseline (r = 0.2, P = 0.02) was weaker than the association between depression and disability (r = 0.3, P = 0.004). Similarly, the association of change in depression scores with change in pain (r = 0.25, P = 0.016) was weaker than change between depression and disability (r = 0.49, P<0.0005). Whereas the relationship between pain and depression became nonsignificant when disability and illness attitudes were controlled, the relationship between depression and disability remained highly significant when pain and illness attitudes were controlled. These data are consistent with the association between pain and depression being wholly modulated by disability and illness attitudes, with no direct relationship between pain and depression. PMID- 11110113 TI - Prognostic awareness and the terminally ill. AB - The authors rated patients who were in advanced stages of cancer and in their final few weeks of life on their level of awareness of their medical prognosis (N = 200, mean age = 71.0 years). The authors measured prognostic awareness with a semistructured interview, dividing patients into those acknowledging No Awareness, Partial Awareness, and Complete Awareness. The authors also administered a semistructured interview for depressive disorders, along with an assessment of various demographic and social support measures. Nineteen patients (9.5%) denied awareness of both their terminal prognosis and foreshortened life expectancy. Thirty-four patients (17%) were placed in the partial awareness category, with the remaining 147 patients (73.5%) reporting complete awareness. Depression was nearly three times greater among patients who did not acknowledge their prognosis, as compared with those who demonstrated partial or complete acknowledgment (chi2 = 7.094), P = 0.029). In addition to depression, male patients, older patients, and those having "intense social contact" were associated with lower ratings of prognostic awareness. Dying patients differ in respect to their capacity to acknowledge their prognosis. Prognostic disavowal is most likely to arise in patients with underlying psychological distress and emotional turmoil. PMID- 11110114 TI - Timing of psychiatric consultations: the impact of social vulnerability and level of psychiatric dysfunction. AB - The authors examined the timing of patient referrals to a psychiatric consultation-liaison service in relation to the patient's social vulnerability and level of psychiatric dysfunction. One hundred consecutive patients were assessed with the INTERMED, a method to document biopsychosocial and health care related aspects of disease. Although 30% of patients were referred within the first day of admission, 19% of requests for referrals were made after 2 weeks. Late referral was associated with high social vulnerability and early referral with severe psychiatric dysfunction. The authors illustrate the disadvantages of a psychiatric liaison model focusing on psychopathology alone and demonstrate the need for an integrated patient assessment in the general hospital, focusing on detecting frail elderly patients. PMID- 11110115 TI - Psychiatric symptoms and medical utilization in primary care patients. AB - In two studies, the authors evaluated the impact of psychiatric disorders on medical care utilization in a primary care setting. In the first study, 526 consecutive patients in a teaching hospital primary care practice completed the 18-item RAND Mental Health Inventory to identify clinically significant depression and/or anxiety and a questionnaire about the use of psychiatric treatment and psychoactive medications. The medical utilization of those patients defined as depressed and/ or anxious was compared with those defined as not depressed and/or anxious. Patients identified as depressed and/or anxious reported significantly increased medical utilization, but this was not confirmed by the hospital's computerized record system. In the second study, the authors analyzed medical care utilization for the years before and after the first outpatient psychiatry appointment of a sample of 91 patients referred from the same primary care practice to the hospital's outpatient psychiatry clinic over a 1-year period. In both studies there was not a statistically significant difference in medical utilization among those patients receiving psychiatric treatment. The findings demonstrate the difficulties in examining cost offset in a primary care population and raise questions about it as a realistic outcome measure of the effect of psychiatric treatment. PMID- 11110116 TI - Prevalence of psychotic symptoms in delirium. AB - Psychosis in delirium has been an underresearched area. The authors retrospectively examined the prevalence of psychotic symptoms and possible associated factors in the records of 227 consecutive hospitalized patients. These patients had been diagnosed with delirium, according to the DSM-IV criteria and referred to a psychiatry consult service. The authors compared patients, with or without psychosis, on demographic variables, medical and psychiatric history; number of medications, etiology of delirium, and cognitive state. The prevalence of psychotic symptoms was 42.7% (n = 97) with 27% of patients (n = 61) having visual hallucinations, 12.4% (n = 28) having auditory hallucinations, 2.7% (n = 6) having tactile hallucinations, and 25.6% (n = 58) having delusions. The presence of visual hallucinations, but not delusions or auditory hallucinations, was significantly associated with more active medical diagnoses and multiple etiologies causing the delirium. Psychotic symptoms are not uncommon in delirium, but specific psychotic symptoms may have different factors contributing to their development. Visual hallucinations appear to be associated with a greater number of active medical disorders, but other factors associated with the development of psychotic symptoms in delirium are currently unknown. PMID- 11110117 TI - The evaluation of maternal competency. AB - There is limited discussion of maternal competency in the consultation-liaison psychiatry literature. As awareness and reporting of child abuse is increasing, maternal ability to care for newborns is more often called into question. Maternal risk factors for harm and neglect have been identified, and positive signs of maternal ability have also been recognized as important to appraise. Specific domains in the maternal competency exam should be assessed by the psychiatrist, nursing staff, social work staff and pediatrician. The competency exam by the psychiatrist requires a sensitive and nonjudgmental inquiry into maternal behavior and thoughts. The authors present a case study of an inpatient maternal competency consultation that illustrates some of the dilemmas encountered. Future directions should include more involvement by psychiatrists in preventive efforts and interventions that focus on pregnant women at risk in prenatal clinics and in the community. PMID- 11110118 TI - A pregnant woman's fear of her baby. PMID- 11110119 TI - Buspirone as an antidote to venlafaxine-induced bruxism. PMID- 11110120 TI - Clarithromycin--a precipitant for acute psychotic stress. PMID- 11110121 TI - Clozapine and tuberculosis. PMID- 11110123 TI - Dynamical mechanism for sharp orientation tuning in an integrate-and-fire model of a cortical hypercolumn. AB - Orientation tuning in a ring of pulse-coupled integrate-and-fire (IF) neurons is analyzed in terms of spontaneous pattern formation. It is shown how the ring bifurcates from a synchronous state to a non-phase-locked state whose spike trains are characterized by clustered but irregular fluctuations of the interspike intervals (ISIs). The separation of these clusters in phase space results in a localized peak of activity as measured by the time-averaged firing rate of the neurons. This generates a sharp orientation tuning curve that can lock to a slowly rotating, weakly tuned external stimulus. Under certain conditions, the peak can slowly rotate even to a fixed external stimulus. The ring also exhibits hysteresis due to the subcritical nature of the bifurcation to sharp orientation tuning. Such behavior is shown to be consistent with a corresponding analog version of the IF model in the limit of slow synaptic interactions. For fast synapses, the deterministic fluctuations of the ISIs associated with the tuning curve can support a coefficient of variation of order unity. PMID- 11110122 TI - Consultation-liaison psychiatry drug--drug interactions update. PMID- 11110124 TI - Statistical signs of common inhibitory feedback with delay. AB - Cross-correlation histograms (CCHs) sometimes exhibit an isolated central peak flanked by two troughs. What can cause this pattern? The absence of CCH satellite peak makes an oscillatory common input doubtful. It is here shown using a simple counting model that a common inhibitory feedback with delay can account for this pattern. PMID- 11110125 TI - On the computational power of winner-take-all. AB - This article initiates a rigorous theoretical analysis of the computational power of circuits that employ modules for computing winner-take-all. Computational models that involve competitive stages have so far been neglected in computational complexity theory, although they are widely used in computational brain models, artificial neural networks, and analog VLSI. Our theoretical analysis shows that winner-take-all is a surprisingly powerful computational module in comparison with threshold gates (also referred to as McCulloch-Pitts neurons) and sigmoidal gates. We prove an optimal quadratic lower bound for computing winner-take-all in any feedforward circuit consisting of threshold gates. In addition we show that arbitrary continuous functions can be approximated by circuits employing a single soft winner-take-all gate as their only nonlinear operation. Our theoretical analysis also provides answers to two basic questions raised by neurophysiologists in view of the well-known asymmetry between excitatory and inhibitory connections in cortical circuits: how much computational power of neural networks is lost if only positive weights are employed in weighted sums and how much adaptive capability is lost if only the positive weights are subject to plasticity. PMID- 11110126 TI - Reclassification as supervised clustering. AB - In some branches of science, such as molecular biology, classes may be defined but not completely trusted. Sometimes posterior analysis proves them to be partially incorrect. Despite its relevance, this phenomenon has not received much attention within the neural computation community. We define reclassification as the task of redefining some given classes by maximum likelihood learning in a model that contains both supervised and unsupervised information. This approach leads to supervised clustering with an additional complexity penalizing term on the number of new classes. As a proof of concept, a simple reclassification algorithm is designed and applied to a data set of gene sequences. To test the performance of the algorithm, two of the original classes are merged. The algorithm is capable of unraveling the original three-class hidden structure, in contrast to the unsupervised version (K-means); moreover, it predicts the subdivision of one of the original classes into two different ones. PMID- 11110127 TI - A model of invariant object recognition in the visual system: learning rules, activation functions, lateral inhibition, and information-based performance measures. AB - VisNet2 is a model to investigate some aspects of invariant visual object recognition in the primate visual system. It is a four-layer feedforward network with convergence to each part of a layer from a small region of the preceding layer, with competition between the neurons within a layer and with a trace learning rule to help it learn transform invariance. The trace rule is a modified Hebbian rule, which modifies synaptic weights according to both the current firing rates and the firing rates to recently seen stimuli. This enables neurons to learn to respond similarly to the gradually transforming inputs it receives, which over the short term are likely to be about the same object, given the statistics of normal visual inputs. First, we introduce for VisNet2 both single neuron and multiple-neuron information-theoretic measures of its ability to respond to transformed stimuli. Second, using these measures, we show that quantitatively resetting the trace between stimuli is not necessary for good performance. Third, it is shown that the sigmoid activation functions used in VisNet2, which allow the sparseness of the representation to be controlled, allow good performance when using sparse distributed representations. Fourth, it is shown that VisNet2 operates well with medium-range lateral inhibition with a radius in the same order of size as the region of the preceding layer from which neurons receive inputs. Fifth, in an investigation of different learning rules for learning transform invariance, it is shown that VisNet2 operates better with a trace rule that incorporates in the trace only activity from the preceding presentations of a given stimulus, with no contribution to the trace from the current presentation, and that this is related to temporal difference learning. PMID- 11110128 TI - An analysis of orientation and ocular dominance patterns in the visual cortex of cats and ferrets. AB - We report an analysis of orientation and ocular dominance maps that were recorded optically from area 17 of cats and ferrets. Similar to a recent study performed in primates (Obermayer & Blasdel, 1997), we find that 80% (for cats and ferrets) of orientation singularities that are nearest neighbors have opposite sign and that the spatial distribution of singularities deviates from a random distribution of points, because the average distances between nearest neighbors are significantly larger than expected for a random distribution. Orientation maps of normally raised cats and ferrets show approximately the same typical wavelength; however, the density of singularities is higher in ferrets than in cats. Also, we find the well-known overrepresentation of cardinal versus oblique orientations in young ferrets (Chapman & Bonhoeffer, 1998; Coppola, White, Fitzpatrick, & Purves, 1998) but only a weak, not quite significant overrepresentation of cardinal orientations in cats, as has been reported previously (Bonhoeffer & Grinvald, 1993). Orientation and ocular dominance slabs in cats exhibit a tendency of being orthogonal to each other (Hubener, Shoham, Grinvald, & Bonhoeffer, 1997), albeit less pronounced, as has been reported for primates (Obermayer & Blasdel, 1993). In chronic recordings from single animals, a decrease of the singularity density and an increase of the ocular dominance wavelength with age but no change of the orientation wavelengths were found. Orientation maps are compared with two pattern models for orientation preference maps: bandpass-filtered white noise and the field analogy model. Bandpass filtered white noise predicts sign correlations between orientation singularities, but the correlations are significantly stronger (87% opposite sign pairs) than what we have found in the data. Also, bandpass-filtered noise predicts a deviation of the spatial distribution of singularities from a random dot pattern. The field analogy model can account for the structure of certain local patches but not for the whole orientation map. Differences between the predictions of the field analogy model and experimental data are smaller than what has been reported for primates (Obermayer & Blasdel, 1997), which can be explained by the smaller size of the imaged areas in cats and ferrets. PMID- 11110129 TI - Improvements to the sensitivity of gravitational clustering for multiple neuron recordings. AB - We outline two improvements to the technique of gravitational clustering for detection of neuronal synchrony, which are capable of improving the method's detection of weak synchrony with limited data. The advantages of the enhancements are illustrated using data with known levels of synchrony and different interspike interval distributions. The novel simulation method described can easily generate such test data. An important dependence of the sensitivity of gravitational clustering to the interspike interval distribution of the analysed spike trains is described. PMID- 11110130 TI - Neural coding: higher-order temporal patterns in the neurostatistics of cell assemblies. AB - Recent advances in the technology of multiunit recordings make it possible to test Hebb's hypothesis that neurons do not function in isolation but are organized in assemblies. This has created the need for statistical approaches to detecting the presence of spatiotemporal patterns of more than two neurons in neuron spike train data. We mention three possible measures for the presence of higher-order patterns of neural activation--coefficients of log-linear models, connected cumulants, and redundancies--and present arguments in favor of the coefficients of log-linear models. We present test statistics for detecting the presence of higher-order interactions in spike train data by parameterizing these interactions in terms of coefficients of log-linear models. We also present a Bayesian approach for inferring the existence or absence of interactions and estimating their strength. The two methods, the frequentist and the Bayesian one, are shown to be consistent in the sense that interactions that are detected by either method also tend to be detected by the other. A heuristic for the analysis of temporal patterns is also proposed. Finally, a Bayesian test is presented that establishes stochastic differences between recorded segments of data. The methods are applied to experimental data and synthetic data drawn from our statistical models. Our experimental data are drawn from multiunit recordings in the prefrontal cortex of behaving monkeys, the somatosensory cortex of anesthetized rats, and multiunit recordings in the visual cortex of behaving monkeys. PMID- 11110131 TI - Gaussian processes for classification: mean-field algorithms. AB - We derive a mean-field algorithm for binary classification with gaussian processes that is based on the TAP approach originally proposed in statistical physics of disordered systems. The theory also yields an approximate leave-one out estimator for the generalization error, which is computed with no extra computational cost. We show that from the TAP approach, it is possible to derive both a simpler "naive" mean-field theory and support vector machines (SVMs) as limiting cases. For both mean-field algorithms and support vector machines, simulation results for three small benchmark data sets are presented. They show that one may get state-of-the-art performance by using the leave-one-out estimator for model selection and the built-in leave-one-out estimators are extremely precise when compared to the exact leave-one-out estimate. The second result is taken as strong support for the internal consistency of the mean-field approach. PMID- 11110132 TI - The Bayesian evidence scheme for regularizing probability-density estimating neural networks. AB - Training probability-density estimating neural networks with the expectation maximization (EM) algorithm aims to maximize the likelihood of the training set and therefore leads to overfitting for sparse data. In this article, a regularization method for mixture models with generalized linear kernel centers is proposed, which adopts the Bayesian evidence approach and optimizes the hyperparameters of the prior by type II maximum likelihood. This includes a marginalization over the parameters, which is done by Laplace approximation and requires the derivation of the Hessian of the log-likelihood function. The incorporation of this approach into the standard training scheme leads to a modified form of the EM algorithm, which includes a regularization term and adapts the hyperparameters on-line after each EM cycle. The article presents applications of this scheme to classification problems, the prediction of stochastic time series, and latent space models. PMID- 11110133 TI - A Bayesian committee machine. AB - The Bayesian committee machine (BCM) is a novel approach to combining estimators that were trained on different data sets. Although the BCM can be applied to the combination of any kind of estimators, the main foci are gaussian process regression and related systems such as regularization networks and smoothing splines for which the degrees of freedom increase with the number of training data. Somewhat surprisingly, we find that the performance of the BCM improves if several test points are queried at the same time and is optimal if the number of test points is at least as large as the degrees of freedom of the estimator. The BCM also provides a new solution for on-line learning with potential applications to data mining. We apply the BCM to systems with fixed basis functions and discuss its relationship to gaussian process regression. Finally, we show how the ideas behind the BCM can be applied in a non-Bayesian setting to extend the input dependent combination of estimators. PMID- 11110134 TI - Anterior versus posterior plating in cervical corpectomy. AB - This is a prospective study comparing anterior and posterior plating in cervical corpectomy. Each group comprised 30 patients who were candidates for corpectomy. In the first group, anterior plating was done using Orosco-type titanium plates. In the second group, lateral mass plating was done. In all cases, titanium cages were used to replace the removed vertebral body or bodies. The mean follow-up was 12.68 months (SD 3.85 months). Pseudarthrosis was not encountered in either group. Posterior plating was better than anterior plating in terms of the stability of the construct and problems related to the hardware. Screw breakage was encountered in seven patients with anterior plating (23.33%). This complication was not seen in the group with posterior plating. The difference between the groups was statistically significant (chi-square = 7.92, p = 0.004). Screw loosening was encountered in 2 patients in the group with anterior plating and in only 1 patient in the group with posterior plating. The difference between the incidence of screw loosening in both groups was not statistically significant (chi-square = 0.35, p = 0.5). Sinking-in of the cage was encountered in 7 cases with anterior plating and in only 3 cases with posterior plating. However, the difference between the groups was not statistically significant (chi-square = 1.92, p = 0.16). PMID- 11110135 TI - Rotationplasty--surgical treatment modality after failed limb salvage procedure. AB - Twelve patients aged between 10.9 and 64.7 (mean 28.5) years with a malignant tumour of the knee region underwent a rotationplasty after failed primary limb salvage procedure. The reasons for failure which finally lead to surgery were recurrent infection in 10 patients and local recurrence of the tumour in 2. The number of operations before the rotationplasty was performed was between 2 and 23 (mean 6.7). According to the primary tumour site, 9 patients underwent a rotationplasty type A1, 3 patients type A2, and 1 patient type BII. In 9 patients the rotationplasty was successful, but 3 patients finally had to undergo amputation. Intraoperative preservation of the vessels was difficult in these 3 patients due to infection and oedema of the arteries or massive fibrous tissues after the previous surgery. After rotationplasty, 3 of 9 patients had to undergo additional surgery because of thrombosis, pseudarthrosis and infection (n = 5, range 1-2). The mean follow-up after rotationplasty was 34.9 (range 13-65) months. The mean functional status according to the MSTS criteria in patients after rotationplasty scored 21.3 of 30 points. In the group of amputees, the score was 19 (range 16-22). Concerning the health-related quality-of-life, the mean score in physical functioning was 76.3 in the group with a rotationplasty versus 50.0 in the group of amputees. Patients with a rotationplasty reached a higher score of global health status (77.1 vs 58.3). Based on the present results we are convinced that rotationplasty can be recommended as a treatment option after a failed limb salvage procedure. PMID- 11110136 TI - Ultrasonically driven instruments in the transfemoral approach--an aid to preservation of bone stock and reduction of implant length. AB - The transfemoral approach to the femur with implantation of the Wagner SL stem provides a means of dealing with the difficult revision problems of extensive endosteolysis and with peri-prosthetic fractures. This surgical approach has been modified such that with the aid of an ultrasonically driven cement removal instrument (OSCAR) the length of the osteotomy is reduced, preserving bone stock, and it is possible to implant a shorter prosthesis in approximately 60% of cases. PMID- 11110137 TI - Grading of functional results of elbow joint arthrolysis after fracture treatment. AB - In the treatment of posttraumatic contracture of the elbow joint, arthrolysis is a proven procedure. We used a stepwise operative approach starting laterally and including an additional medial and dorsal incision if needed. A total of 91 patients with arthrolysis of the elbow could be followed-up on average 44 months (range 9-102 months) joint after operative (58, 63.7%) and non-operative (33, 36.3%) fracture treatment. The mean preoperative range of motion (ROM) in flexion/extension was 49 degrees (SD +/- 38 degrees), while in pronation/supination it was 89 degrees (SD +/- 66 degrees). Postoperatively, the ROM was on average 94 degrees (SD +/- 27 degrees) in flexion/extension and 129 degrees (SD +/- 52 degrees) in pronation/supination. Using our own grading system, it became evident that most patients had a functional benefit from the procedure, although the quality of the improvement differed. For example, postoperatively 59.3% of the patients were grade I (> or = 90 degrees) in flexion/extension compared with 16.5% preoperatively. Although the rest also showed improvements, their functional benefit was less. The earlier the release of the joints was performed, the better was the functional outcome (p < 0.05). The importance of an intensive early rehabilitation programme is emphasised while indications for this procedure should only be seen in compliant patients. PMID- 11110138 TI - Histological and microbiological findings in non-infected and infected revision arthroplasty tissues. The OSIRIS Collaborative Study Group. Oxford Skeletal Infection Research and Intervention Service. AB - An assessment of clinical and laboratory findings is generally required to distinguish between septic and aseptic loosening of a hip implant. In order to evaluate the diagnostic utility of histological and microbiological investigative techniques to differentiate between these two conditions, we analysed their results in 617 patients with hip implant loosening. Histology and microbiology study confirmed the clinical diagnosis of septic loosening in approximately 98% and 89%. respectively. The clinical diagnosis of aseptic loosening was confirmed by histology in 99% of cases. In all but 2 of 81 cases of septic loosening, in which an organism was isolated on microbiological culture, the histological diagnosis of septic loosening was made on the basis of the degree of the acute inflammatory infiltrate (i.e. the presence of 1 or more neutrophil polymorphs per high power field (x 400) on average after examination of at least 10 high power fields) in periprosthetic tissues. In 10 patients for whom there was a strong clinical suspicion of septic loosening but no organisms were isolated on microbiological culture, the histological findings, using the above criteria, were in keeping with the clinical diagnosis of septic loosening. As almost 11% of cases of septic loosening would not have been diagnosed by microbiological investigation alone, our findings indicate that histological examination of periprosthetic tissues should form part of the investigative protocol to distinguish between aseptic and septic loosening. PMID- 11110139 TI - Effects of single-dose versus fractionated irradiation on the suppression of heterotopic bone formation--an animal model-based follow-up study in rats. AB - The histological and enzymatic effects of single-dose irradiation of 7 Gray (Gy) versus fractionated irradiation of 5 x 2 Gy on the suppression of heterotopic ossification were examined over a period of 60 days in adult male Wistar rats (n = 57). The standardized osteogenesis model system in rats 19, 10, 11, 16, 19] was used for this purpose. The course of developing ossifications was documented quantitatively and qualitatively by means of quantitative computed tomography/osteodensitometry and digital luminescence radiography. Assessment of the activities of the enzymes alkaline and acid phosphatase throughout the experiment as well as characterization of the isoenzyme of alkaline phosphatase (AP) in connection with histological observations displayed a metaplasia of the ingrowing connective tissue into bone-typical cells during osteoinduction. Thus, the increase of AP is the first sign of a functional transformation of mesenchymal stem cells into chondroid bone cells. The increase in the acid phosphatase level with a maximum of activity between the 15th and 30th day (according to the respective treatment group) is highly suggestive of a remodeling process paralleling incipient chondroclast and osteoclast activity. In the animal groups undergoing irradiation, the above-mentioned increase of enzymes occurred after a delay. Furthermore, the maximum values observed were lower than those in the group not undergoing irradiation. Both findings were more manifest in the animal group which underwent 5 x 2 Gy of radiation than in the group which underwent single-dose irradiation of 7 Gy. Radiation suppresses matrix-induced osteogenesis. The histological and enzymatic course of this process was unchanged in the animals which did not undergo irradiation. However, it was quantitatively reduced and accompanied by a retardation of osteogenesis. Both effects were again reduced with fractionated irradiation of 5 x 2 Gy, which is theoretically dose equivalent to a 1 x 7 Gy application. Histological examinations revealed damage to the migratory, proliferating mesenchymal stem cell population by irradiation doses which had relatively small effects on preosteoblasts, osteoblasts, chondroblasts and other specialized cell forms. Therefore, it may be concluded that the smaller degree of heterotopic ossification in the irradiated groups was due to damage of and a decrease in the number of mesenchymal stem cells at the implant site. Our results stress the necessity of instituting postoperative irradiation therapy as early as possible to prevent heterotopic ossification. In view of experimentally proven better effects, fractionated irradiation has to be preferred to a dose-equivalent single-dose radiation, especially considering the fewer side-effects noted with fractionated irradiation. PMID- 11110140 TI - Inheritance of the accessory navicular bone. AB - The accessory navicular bone is one of the most symptomatic bones of the foot. Although it has been reported to be present in various members of the same family, there is a lack of knowledge about its inheritance in the literature. We examined three families and suggest that it has an autosomal dominant trait with incomplete penetrance. PMID- 11110141 TI - The pelvic external fixation: the mid-term results of 41 patients treated with a newly designed fixator. AB - The purpose of this study was to evaluate the clinical and radiological results of unstable pelvic fractures treated with a new external fixation device. Between May 1992 and May 1998, 43 patients with unstable pelvic fractures were treated with a new anterior pelvic external fixator. Two died, and therefore 41 patients' results were evaluated. There were 29 men and 12 women, and their average age was 34 years (range 12-70 years). Traffic accidents accounted for 34 injuries. Three patients fell from a height, 3 were injured in industrial accidents, and 1 was hit by a train. According to the Tile classification, there were 24 type B pelvic injuries and 17 type C. Associated injuries were observed in 21 patients. A considerable reduction of the pelvic pain was noted after application of the fixator in all patients. Excessive blood transfusion was not required in any patient. The average follow-up was 24 months (range 12-50 months). Clinical results at final evaluation were good according to the criteria of Matta and Saucedo in 34 patients and poor in 7. In conclusion, the new pelvic external fixator is effective, safe, and easy to apply in the treatment of unstable pelvic fractures. The fixator can be used alone in patients with type B pelvic injuries such as open book and lateral compression. However, it does not provide sufficient stability for severely displaced type C injuries when applied alone. Nevertheless, it may be helpful for fixing type C injuries like a posterior iliac fracture without dislocation of the sacroiliac joint. PMID- 11110142 TI - Influence of trypsin on the biological bonding of cartilaginous surface to bone in rabbits. AB - Biological healing between cartilage and bone is difficult, because their histological structure and physiological function are completely different, and the healing ability of cartilage is limited. A full-thickness articular cartilage defect was created in the femoral groove of 48 Japanese white rabbits. The defects were filled with a piece of articular cartilage from the contralateral patella with its superficial zone facing down to the defect. Before grafting, a piece of articular cartilage was treated with 0.05% trypsin or phosphate-buffered saline (PBS) for 5 min. The healing process was histologically observed at 2, 4, 8, and 16 weeks after operation. We found that the incorporation process of cartilage implant treated with trypsin was much better than that of PBS pretreated cartilage. Proliferation and reorganization of chondrocytes in the cartilage implant and subchondral bone ingrowth were observed in the trypsin pretreated group. Although the observation period in our study was not very long, our results suggest that trypsin pretreatment might be one of the most promising methods to facilitate the repair process between cartilage and bone. PMID- 11110143 TI - The 'mirrored' Bennett fracture of the base of the fifth metacarpal. AB - Fractures of the base of the metacarpals are usually treated conservatively. The intra-articular fracture of the base of the first metacarpal ('Bennett fracture') is an exception to this rule because inadequate repositioning and fixation of the dislocated radial fragment lead to permanent deformity of the joint and subsequent degenerative joint disease. The dislocated intra-articular fracture of the base of the fifth metacarpal is similar to a Bennett fracture in many aspects. Repositioning of this 'mirrored' Bennett fracture cannot be guaranteed by a plaster cast. Inadequate repositioning will lead to pain, reduced strength and early degenerative joint disease. We present six patients with dislocated intra-articular fractures of the base of the fifth metacarpal to illustrate the necessity of surgical reduction and fixation. PMID- 11110144 TI - Destructive spondyloarthropathy mimicking spondylitis in long-term hemodialysis patients. AB - A 63-year-old man with end-stage renal disease (ESRD) who had been undergoing hemodialysis for 18 years suffered persistent neck pain, progressive quadriparesis, and a deteriorating ataxic gait during the 6 months before admission. A sudden onset of aggravating quadriparesis and an inability to ambulate occurred during his trip to Sydney, Australia, 1 week before this admission. Vertebral tuberculosis osteomyelitis of the C5/6 segment was considered and treated in a hospital there. Findings from cervical magnetic resonance imaging (MRI; low signal intensity on both T1- and T2-weighted images) were diagnostic of destructive spondyloarthropathy (DSA) and distinguishable from spinal osteomyelitis preoperatively. Amyloid masses, mainly composed of B-2 microglobulin, filled in disc and paradiscal ligaments, with adjacent end-plate destruction by cytokine-mediated reactive inflammation, and appeared to be mostly related to the pathogenesis of DSA. The cervical spine, especially C5/6, is the most common site of DSA. Spinal instability and neurologic compression cause the clinical symptoms and signs. Adequate decompression and successful cervical fusion ensure the best therapeutic results. PMID- 11110145 TI - Isolated palmar dislocation of the distal radioulnar joint in a football player. AB - Palmar dislocation of the distal radioulnar joint without concomitant fracture of the radius or ulna is an uncommon injury. We report one case in a college football player. This case was unusual in that open reduction was required 2 days after the injury because of an unsuccessful closed reduction. PMID- 11110146 TI - A rare case of enthesopathy of the bicipital tuberositas of the radius. AB - We present a case of a patient with complaints in both elbows, due to a prominent bicipital tuberosity with an enlarged bursa, in which operative resection was successful. PMID- 11110147 TI - Hypoplasia of bilateral humeral trochlea associated with bilateral ulnar nerve palsy. AB - We document a case of bilateral ulnar nerve palsy that developed in an 27-year old Japanese man who had bilateral hypoplasia of the humeral trochlea. Surgical management produced good results regarding the ulnar nerve palsy. The pathogenesis of the nerve paresis in this particular condition is discussed. There have been no reports outside Japan. Whether this deformity occurs only in persons of Japanese extraction or is simply overlooked by foreign surgeons is an interesting question. PMID- 11110148 TI - Childhood periosteal chondroma. AB - We report here three fully documented cases of periosteal chondroma diagnosed and treated in our hospital. There have been few references since Lichtenstein first described this condition as a separate tumour, and none of them concerned children under 10 years old. deSantos accurately describes the radiological features of the tumour. All of our patients were under 10 years old at the moment of the appearance of the lesion, which was always detected in the long bones of the upper limbs. The study included a previous histological examination in two cases and a follow-up in all of them. From this study we learned that invasive diagnosis procedures are unnecessary as we could follow the behaviour and evolution of this cartilaginous benign tumour for a long period of time. PMID- 11110149 TI - Prevention of lid retraction after lower lid blepharoplasties: an overview. AB - This overview article covers the techniques that aim to prevent lid retraction after lower blepharoplasties. After a brief review of applied anatomy (anterior, middle, posterior lamella), the causes of blepharochalasis and of postoperative lid retraction are addressed. Clinical examinations are described that can detect a candidate at risk. Preventive measures are described, dealing with patient positioning, approaches (transcutaneous-transconjunctival-combination of both), incision types, flap dissection (skin-skin-muscle-dermal flaps), muscle suspension techniques (muscle-muscle, muscle-periosteum), horizontal wedge excisions, lateral (tendon and tarsal) and medial canthal procedures, CO2 laser skin resurfacing and combinations. These techniques are described and critically appraised. PMID- 11110150 TI - Time series analysis of births of children with orofacial clefts. AB - Observation of the dates when children with orofacial clefts were born, gives the impression that they were born in clusters, between which there were intervals of different duration. An analysis was therefore undertaken of the period between 1972 to 1997 when 1112 such children were born. For diagnostic checking of stationarity of the time series the Box-Ljung test was used. Since it was found that the time series was stationary, analysis of autocorrelation function was performed. The results confirmed the impression that children with clefts are born in clusters, between which there are gaps of different duration. This suggests that the birth of a child with a cleft is not a random and independent event. PMID- 11110151 TI - Obstructive sleep apnoea: multiple comparisons of cephalometric variables of obese and non-obese patients. AB - BACKGROUND: Pathogenesis of obstructive sleep apnoea (OSA) is complex and not yet fully understood. Several factors contribute to OSA severity. Obesity is believed to play an important role. Nevertheless, not all OSA patients are obese. Therefore, the different features that cause nocturnal upper airway obstruction in obese and non-obese OSA patients could be expected. PURPOSE: To investigate the different components of cervico-craniofacial skeletal and upper airway soft tissue morphology among obese OSA, non-obese OSA patients and the controls. PATIENTS: One hundred male OSA patients were classified into two groups on the basis of body mass index (BMI) as obese (BMI > or = 30 kg/m2) and non-obese (BMI < 30 kg/m2). Consequently, 57 obese and 43 non-obese OSA patients were examined and compared with a control group of 36 healthy males. STUDY DESIGN: A comprehensive cephalometric analysis of cervico-craniofacial skeletal and upper airway soft tissue morphology was performed. Sixty-eight cephalometric variables were compared among the three groups by one way analysis of variance with Bonferroni's test. RESULTS: Both OSA groups had aberrations of cervico craniofacial skeletal as well as upper airway soft tissue morphology when compared with the controls. These anatomic deviations were confined to cervico craniofacial skeletal structures in the non-obese OSA patients, whereas the obese OSA patients had more abnormalities in the upper airway soft tissue morphology, head posture and position of the hyoid bone. CONCLUSION: The findings imply that there should be different treatment regimens for the two subgroups of OSA patients. Cephalometric analysis together with various considerations of BMI is highly recommended as one of the most important tools in diagnosis and treatment planning for OSA patients. PMID- 11110152 TI - Single-stage CO2 laser assisted uvuloplasty for treatment of snoring and mild obstructive sleep apnoea. AB - PURPOSE: The purpose of this study was to describe a single-stage laser assisted uvuloplasty (uvulectomy) and to determine its effectiveness in treatment of snoring and mild obstructive sleep apnoea (OSA). PATIENTS AND METHODS: All patients treated with laser assisted uvuloplasty in a 49-month period for snoring and/or mild OSA were studied. Frequency of snoring before and after surgery, loudness of snoring and postoperative discomfort were investigated. Patients were asked to evaluate change in daytime energy, sleep habits, missed days of work and also overall satisfaction following laser assisted uvuloplasty. RESULTS: Thirty patients underwent a single-stage laser assisted uvuloplasty. A preoperative diagnosis of OSA was established in 19 patients, the remaining 11 patients were treated for snoring. There were no complications and only one patient required an additional stage. A questionnaire was completed by 18 patients (10 patients diagnosed with sleep apnoea, and eight patients with snoring only). Preoperatively the frequency of snoring averaged 9.3 cm on a visual analogue scale. Postoperatively there were 12 patients with either none or very minimal snoring and six patients who had an average score of 3.2. Loudness of snoring also decreased from an average of 5.4 to 2.5 cm. Postoperative discomfort averaged 1.1 cm. Improvement in sleep was noted by 16 patients and improved daytime energy was noted by 17 patients. Eleven patients reported that they missed at least one day of work postoperatively with an average of 3 days missed. Patient satisfaction was reported by 17 patients with only one stating that he was unsatisfied with the procedure. CONCLUSIONS: Laser-assisted uvuloplasty (uvulectomy) is an effective surgical procedure for treatment of snoring and some types of OSA. A single-stage procedure appears to be effective and may further decrease the morbidity associated with this disease. PMID- 11110153 TI - Incidence of os japonicum in Anatolian dry skulls and plain cranium radiographs of modern Anatolian population. AB - INTRODUCTION: The zygoma may sometimes be divided into two parts by either a horizontal or a vertical suture. Such a bipartite bone has been given the name of os japonicum as it has been mostly observed in Japanese. MATERIAL: In this study 1266 zygomatic bones in 633 Anatolian dry skulls and 1348 zygomatic bones in 674 plain cranium radiographs of adult patients have been examined. RESULTS: Os japonicum was present in 2.2% of female and 1.7% of male subjects. All of the 24 multipartite bones observed in the study were bipartite except one. In addition, of 690 female zygomatic bones examined radiologically 15 (2.2%), and 658 male bones 12 (1.8%) were bipartite or tripartite, a total of 674 plain cranium radiographs. CONCLUSION: The results were compared with those of other populations. As a result evaluation of both dry skulls and plain radiographs as a single sample, our results were again closely similar to Main Island Japan and other East Asia groups. PMID- 11110154 TI - Growing skull fracture of ethmoid: a report of two cases. AB - We describe a rare sequel of ethmoid fracture--a growing skull fracture associated with cerebrospinal fluid rhinorrhoea following trauma sustained in adult life. The natural history of its development, diagnosis, and the results of surgery are discussed. The literature is reviewed with regard to aetiology, incidence, imaging characteristics and management of this rare post-traumatic complication. PMID- 11110155 TI - Reconstruction of orbital floor fractures using bioactive glass. AB - INTRODUCTION: The management of orbital floor fractures continues to be debated. Bioactive glasses and glass-ceramics are in the new group of materials developed for the repair of bone defects which are beyond any innate healing capacity due to their size. OBJECTIVE: We compared the use of alloplastic implants (bioactive glass) with conventional autogenous grafts (cartilage--plus or minus Iyophilized dura) for the repair of orbital floor defects after trauma. MATERIAL AND METHODS: Twenty-eight patients having orbital floor fractures with persistent diplopia, enophthalmos, and/or infraorbital nerve paraesthesia were operated on from 1991 to 1995 at Turku University Central Hospital. Reconstruction was either with bioactive glass (S93P4) or autogenous cartilage implants. RESULTS: Postoperative tomograms in the 28 patients showed adequate maintenance of orbital and maxillary sinus volume without any evidence of resorption in either group. None of 14 patients in the study group had any evidence of dystopia or complications relating to implants follow-up. One had infraorbital nerve paraesthesia and another had entropion postoperatively. Among the 14 control subjects there were three cases of persistent diplopia, two of infraorbital nerve paraesthesia and one of enophthalmos. CONCLUSION: Bioactive glass implants are well-tolerated and seem to be a promising repair material for orbital floor fractures. Their use leads to less morbidity as no donor site operation is needed. Also it provides favourable healing as it is bioactive, biocompatible and causes new bone formation. PMID- 11110156 TI - Treatment of complex mandibular fractures using titanium mesh. AB - INTRODUCTION: The treatment of complex fractures with miniplates is often difficult and unsatisfactory. Such fractures include extremely atrophic mandibles, discontinuity defects, and marked comminuted fractures. AIM: With this paper we want to call to mind once more that titanium mesh is useful for the treatment of such fractures and its use can reduce the number of complications. PATIENTS: Between January 1996 and December 1998 we treated with titanium mesh 17 patients with fractures of extremely atrophic mandibles, mandibular discontinuity defects or comminuted fractures. RESULTS: Union occurred without complication in 70% of fractures treated with titanium mesh. In 20% there were minor complications such as postoperative haematoma. In only one case did infection occur, a more severe complication. CONCLUSION: Because of its geometry and the excellent physical and biomechanical properties, titanium mesh helps to achieve better stabilization of complex mandibular fractures than conventional miniplates do. Complications such as infection and non union can largely be avoided and bony continuity of the mandible can be restored. PMID- 11110157 TI - Surgical technique for the treatment of high-flow arteriovenous malformations of the mandible. AB - The high-flow intraosseous arteriovenous malformation is a problematic vascular lesion which may affect bone and the dentition. Variable clinical presentations of this anomaly have resulted in a gamut of treatment modalities being reported ranging from simple curettage, resection, radiotherapy, sclerosing injections, and various forms of embolization, to immediate replantation of the resected segments. Embolization techniques alone have not been universally successful and have often resulted in rapid development of collaterals from surrounding vessels. Definitive treatment has usually involved complete surgical resection (when feasible) either alone, or in combination with other modalities such as embolization. Jaw resection, however, is deforming and leaves a defect often requiring subsequent reconstruction of the hard and soft tissues and replacement of any teeth lost with the resected segment. We report a surgical technique to treat mandibular arteriovenous malformations, which permits ligation of the feeding vessels and provides access allowing for complete removal of the intraosseous lesion. At the same time it not only prevents facial deformity by preserving the mandibular bone and oral soft tissue, but also, and more importantly, may preserve the dentition as well. PMID- 11110158 TI - Retropharyngeal abscess secondary to penetrating foreign bodies. AB - A retrospective study was conducted on patients with upper aerodigestive tract foreign bodies requiring operative intervention over a 12-year period to aid in the recognition and management of foreign body associated complications. Oesophagoscopies were performed for the removal of foreign bodies in 37 patients, age one to 82 years with a male to female ratio of 1.2:1. Retropharyngeal abscesses accounted for eight of 11 foreign body-associated complications. Fish bones were the cause in six cases, chicken bone and a pen refill in one case each. An abscess was already present at the time of initial procedure in six cases and developed in two cases after successful removal of the foreign body. A high level of suspicion for a retropharyngeal abscess should be maintained in cases with perforation, and in patients with immunodeficiency. PMID- 11110159 TI - Blood viscosity and hematological changes during prolonged submergence in normoxic water of northern and southern musk turtles (Sternotherus odoratus). AB - Musk turtles (Sternotherus odoratus) can survive at least 150 days of submergence in normoxic water at 3 degreesC, during which time there are large increases in packed cell volume (PCV). We investigated the effects of submergence in normoxic water at 3 degreesC on the blood viscosity of musk turtles from northern (Massachusetts) and southern (Alabama) locales. Blood was collected from air breathing turtles and after 20, 50, 100, and 150 days of submergence in normoxic water at 3 degreesC. Hematological responses to submergence were similar in the two groups, therefore the results were combined. Packed cell volume increased steadily above that of controls after 20, 50, 100, and 150 days of submergence. Hemoglobin concentration also progressively increased above that of controls after 20, 50, and 100 days of submergence but declined to near control values after 150 days. Blood viscosity increased with increasing PCV; however, blood viscosity of musk turtles appears less affected by PCV than is blood viscosity of mammalian species. As such, musk turtles appear to be able to maintain adequate blood flow to tissues while increasing the oxygen carrying capacity of the blood during prolonged submergence. However, after 150 days submergence, oxygen delivery should decrease due to a reduced oxygen carrying capacity of the blood and an increased resistance to blood flow, which may limit the length of time these turtles can remain viable during hibernation. PMID- 11110161 TI - Mitogen-activated protein kinases and anoxia tolerance in turtles. AB - The response of two vertebrate mitogen-activated protein kinase (MAPK) family members, the extracellular signal-regulated kinases (ERKs) and c-Jun NH2-terminal kinases (JNKs), to anoxia exposure in vivo was examined in organs (liver, heart, kidney, brain, spleen) of the anoxia-tolerant adult turtle, Trachemys scripta elegans. ERKs activities rose during anoxia only in spleen (a 2.8-fold increase). JNK activity showed a significant increase only in liver (4-fold increase) after 5 hr of anoxic submergence but declined thereafter. Levels of the transcription factor c-Fos were strongly suppressed in liver, heart, and kidney of anoxia exposed turtles, whereas levels increased 2-fold in anoxic brain. The effect of anoxia on c-Myc was organ-specific and variable with 2- and 1.5-fold increases in protein expression in kidney and brain, respectively, and a 60% decrease in anoxic spleen. These results for an anoxia-tolerant animal suggest the potential importance of the MAPKs and of the immediate-early genes (c-fos, c-myc) in mediating adaptive responses to oxygen deprivation. PMID- 11110160 TI - Characterization of retinal oil droplets in diurnal geckos (reptilia, gekkonidae). AB - Retinal oil droplets have been documented in the retinae of four genera of diurnal geckos (Phelsuma, Gonatodes, Quedenfeldtia, and Pristurus), while other large diurnal genera (Sphaerodactylus and Lygodactylus) lack oil droplets. Where they occur, droplets are found only in the minor members of double cones of type B of the extrafoveal and peripheral regions, whereas in the foveal cones droplets could not be detected. Oil droplets in gecko retinae have neither an internal structure nor an own membrane; this is typical for oil droplets of amphibians and sauropsids. The droplets are nonfluorescent and definitely transparent. Thus, they do not function as filters modifying the spectral composition of the light reaching the outer segments. The constant relationship between the diameters of the oil droplets and those of the outer segment bases might still suggest an optical function of the droplets (e.g., as microlenses focusing light on the outer segments). However, as the ecologically very similar genera Phelsuma, Gonatodes, and Lygodactylus differ in the presence or absence of oil droplets, this potential function seems to be not of physiological significance. PMID- 11110162 TI - Incorporation and metabolism of cortisol in oocytes of tilapia (Oreochromis mossambicus). AB - The entry and metabolism of 3H-cortisol in oocytes were investigated using isolated follicles of the tilapia (Oreochromis mossambicus) in order to examine the mechanisms of incorporation of maternal hormones into oocytes. The composition of 3H-labeled steroids in the oocyte was analyzed by high-performance liquid chromatography. A significant amount of cortisol was converted to cortisone and an unidentified molecule by the follicular layer. The contents of 3H-cortisol and 3H-cortisone in the oocyte reached an equilibrium level within 12 hr, whereas the content of the unidentified metabolite continued to increase for 36 hr. The total content of the incorporated cortisol and its metabolites was proportional to cortisol in the medium over the concentration range of 5 ng/ml to 5 microg/ml. The amounts of cortisone and the unidentified molecule increased proportionally when the concentration of cortisol in the medium was lower than 500 ng/ml, whereas they reached a plateau when the concentration of cortisol exceeded 500 ng/ml. Cortisol entry was reversible, because 90% of cortisol and cortisone in the oocyte was lost within 18 hr when the medium was changed to that without 3H-cortisol. On the other hand, 50% of the unidentified molecule was preserved at the end of the incubation. In conclusion, the entry of cortisol into the oocyte was considered to be nonspecific and due probably to simple diffusion. However, a considerable amount of cortisol (50-70%) was specifically converted to cortisone and another unidentified molecule during passage through the follicular layer. PMID- 11110163 TI - Effect of induced triploidy on fin regeneration of juvenile rainbow trout, Oncorhynchus mykiss. AB - Triploidy was induced in the rainbow trout in order to evaluate whether the altered numbers and sizes of triploid cells could modify fin regeneration. Amputation of one lobe of the tail fin of diploid and triploid juveniles resulted in regeneration for experimentals and controls. Nevertheless, both rate and frequency of regeneration in triploids were significantly increased as compared with diploids, a fact that can be attributed to the increased nuclear and cellular volume in a wide range of tissues, whereas the cell numbers were reduced. These data suggest that a great deal of interesting and important research could be done using triploid animals as experimental models for studying the regeneration of appendages. PMID- 11110164 TI - Involvement of glycolytic metabolism in developmental inhibition of rat two-cell embryos by phosphate. AB - To elucidate the mechanism by which phosphate induces developmental inhibition of rat 2-cell embryos, we examined the mutual effects of glucose and other glycolytic and non-glycolytic sugars, the non-metabolizable glucose analogue, and glycolytic inhibitors on the inhibitory effect of phosphate. In the absence of glucose, 30-49% of embryos treated with 10-500 microM phosphate were able to develop to morula and blastocysts. On the other hand, in the presence of 5 mM glucose, 10 microM phosphate decreased the developmental rate of 2-cell embryos to the 4-cell stage and completely inhibited the development beyond the 4-cell stage. In contrast, glucose showed no influence on development in phosphate-free medium. Similarly to glucose, the other glycolytic sugars fructose (5 mM) and mannose (5 mM) enhanced the inhibitory effect of 10 microM phosphate but had no influence in the absence of phosphate. In contrast, the non-glycolytic sugar and non-metabolizable glucose analogue N-acetylglucosamine and 3-O-methylglucose (3-O MGlc), respectively, did not enhance the effects of phosphate. 2-Deoxyglucose (2DGlc), another glucose analogue that is non-metabolizable but is converted by hexokinase to 2DGlc 6-phosphate, at concentrations as low as 0.1 mM completely inhibited cell cycle progression of 2-cell embryos cultured in glucose-free (Glc( )) medium with 10 microM phosphate. In contrast, in the absence of phosphate, 2DGlc at the same concentration allowed 55% of 2-cell embryos to develop to morula and blastocyst stages. Addition of an inhibitor of enolase in glycolysis, sodium fluoride (NaF), at 1 mM to the Glc(-) medium also enhanced the inhibitory effects of 10 microM phosphate, whereas 1 mM NaF in the absence of phosphate showed no inhibitory effects on the development of 2-cell embryos to morula and blastocyst stages. From these results, disturbance of glycolysis is a critical reason for the developmental inhibition caused by phosphate in early rat embryos in culture. PMID- 11110165 TI - Hatching of an estuarine crab, Sesarma haematocheir: from disappearance of the inner (E3) layer to rupture of the egg case. AB - Hatching of decapod crustaceans is characterized by the sudden rupture of the egg case. This study focused on the following two issues regarding the hatching mechanism of the estuarine terrestrial crab Sesarma haematocheir: (1) dissolution of the egg case, and (2) the site where the egg case breaks. The egg case comprises three layers: the outer two (E1 and E2) layers and the inner (E3) thin layer (0.2 microm in thickness). The outer layers showed no morphological changes upon hatching, but the inner layer (E3) was markedly digested. The digestion of this layer would enable the embryo to absorb ambient water via reverse peristalsis of the intestine, resulting in an increase of the volume. The egg case always ruptured perpendicular to the longitudinal axis of the embryo. In addition, breakage of the egg case occurred at the dorsal thorax of the embryo. The three major organs positioned at this area were (1) a sharp projection (dorsal spine), (2) an assemblage of muscles, and (3) a pair of secretory glands, each of which was about 30 microm in diameter. The dorsal projection is soft before hatching, and it is clear that the egg case does not break with the posterior expansion of this projection. The rupture instead appears to be caused by the expansion of the muscles arranged perpendicular to the body axis. In addition, some (unknown) factor might weaken the egg case just before hatching. The secretory glands may be a kind of rosette gland, but the role that this gland plays at hatching is not known. As a duct comes out from the center and enters the dorsal projection, some active substance may be released at the tip of this projection. However, immunochemical studies are not consistent with this substance being an ovigerous hair stripping substance (OHSS). PMID- 11110166 TI - Richard Rosner Awards for the best papers by Fellows in Forensic Psychiatry or Forensic Psychology. PMID- 11110167 TI - Assessing the risk of recidivism in physicians with histories of sexual misconduct. AB - Physicians who engage in sexual conduct with patients usually cause serious harm and have a high rate of recidivism. Although offending physicians may lose their privilege to practice, they have the right to appeal for restoration of the license. Yet medical licensing board members do not currently have any clear standards by which to predict whether a given physician is likely to abuse again. Using New York as a paradigm, this paper offers practical, clinically based guidelines for assessing the risk of restoring an offending physician's license. These guidelines are derived from psychoanalytic theories of character, the insights of therapists who have worked with abusive physicians, and the psychiatric model of assessing dangerousness. Recognizing character patterns and psychological vulnerabilities of physicians with histories of sexual misconduct will help board members identify those who are at high risk of abusing again if their licenses are restored. PMID- 11110168 TI - Elderly sexual offenders admitted to a maximum-security forensic hospital. AB - The purpose of the study was to determine the clinical and demographic characteristics of elderly sexual offenders at a maximum-security forensic hospital. Charts of male elderly patients charged with sexual offenses were reviewed to obtain clinical and demographic data. The majority of sexual offenders had mood or psychotic disorders. Almost one third had a history of violent or assaultive behavior. 57% had significant medical history. A history of violence and sexual assaultive behavior may be risk factors for future sexual offenses. PMID- 11110169 TI - Clinical and demographic characteristics of elderly offenders at a maximum security forensic hospital. AB - The purpose of the study was to determine the clinical and demographic characteristics of the male elderly offenders admitted to a maximum-security forensic hospital. Charts of male elderly patients were reviewed to obtain clinical and demographic data. Seventy-seven percent of geriatric felons were involved in violent crime, 41% of which had psychotic symptoms. Forty-five percent of offenders with a history of head trauma/neurologic disorder were charged with violent offenses. Fifty-nine percent had previous psychiatric hospitalization. Most elderly male offenders involved in violent crimes had primary psychotic and mood disorders, cognitive impairment, and a history of head trauma/neurologic disorder. The small number of subjects precludes clear conclusions and needs further study. PMID- 11110170 TI - The state of electroconvulsive therapy in Texas. Part I: reported data on 41,660 ECT treatments in 5971 patients. AB - The Texas Legislature in 1993 mandated a quarterly reporting requirement for hospitals and physicians performing electroconvulsive therapy (ECT) in the state (United States Government hospitals were excluded). The Texas Department of Mental Health and Mental Retardation (TDMHMR) was designated as the agency responsible for collecting and maintaining the data. This paper reviews the ECT data from 16 quarterly reports (09/01/93 through 08/31/97). The reports contained data on 41,660 ECT treatments in approximately 5971 patients. The results of this study support the proposition that ECT is an extremely safe and effective treatment for those individuals suffering from a serious mental illness. In Texas, ethnic groups other than non-Hispanic Anglo-Americans appear to be underserved in regards to ECT. Those patients without appropriate insurance or adequate personal finds are also underserved as a result of the few county and state hospitals performing ECT and the relatively small number of patients treated with ECT at those hospitals. Recommendations are suggested to improve the quality of the database and in informing the public as to the safety and efficacy of this valuable treatment modality. What, at first, was seen as an unwarranted legislative foray into the practice of medicine, has, in the end, become a source of valuable data supporting the use of ECT as an important treatment modality. PMID- 11110171 TI - The state of electroconvulsive therapy in Texas. Part 2: contact with physicians, hospitals, medical liability insurance companies, and manufacturers of stimulus generating equipment. AB - Since mid-1993, all ECT treatments performed in the state of Texas (except for United States government hospitals) must be reported every quarter to the Texas Department of Mental Health and Mental Retardation (TXMHMR) on a data collection form provided by the Department. Part 1 of this paper reviewed that data. This paper reviews the responses to questionnaires and contacts made with physicians, hospitals, medical liability insurance companies, and manufacturers of stimulus generating devices regarding their experience with ECT in Texas. Questionnaires were sent to physicians and hospitals that had not performed ECT during the final two quarters of the review period. Medical liability insurance companies and the manufacturers of the stimulus generating equipment used in ECT were contacted regarding their experience with liability claims. The results indicate that medical liability in regards to the performance of ECT is extremely low. Physicians and hospitals that stopped performing ECT did so for reasons other than medical liability. PMID- 11110172 TI - Characteristics of mentally retarded criminal offenders in Northern Taiwan. AB - The characteristics and criminal behavior in mentally retarded individuals remain largely unstudied. This retrospective study sought to establish a set of reference of criminal behavior characteristics in an ethnic Chinese mentally retarded group. Data were collected from forensic psychiatric evaluation of 32 mentally retarded offenders. Of the 32 offenders, only four (12.5%) cases were female. Mean age at the time of the offenses was 31. By IQ testing, 23 (71.9%) of the group fell into the mild mental retardation range, seven (21.9%) into the moderate mental retardation range, and two (6.2%) into the severe mental retardation range. Nineteen (59.3%) of the group also suffered from additional mental disorder. Eight (25%) had definite neurological deficit. Fourteen (43.8%) were repeat offenders. A total of 24 (75%) of the offenders had committed crimes against property, with 13 having committed petty theft. Furthermore, the pattern of offending shows differences from that of the general population or other mental disorders. The property offenses, especially petty theft and arson, were frequently seen. There was no noteworthy above average frequency of sexual offenses. PMID- 11110174 TI - Comparing the additive composition of smokeless gunpowder and its handgun-fired residues. AB - Detecting the use of handguns via the determination of the organic additives in smokeless gunpowder residues (OGSR) presents a promising alternative to primer metal residue analysis. Compositional analysis of the gunpowder additives nitroglycerin, diphenylamine, and ethyl centralite provides information that can associate residue samples with unfired gunpowder. We evaluated the composition of seven reloading smokeless gunpowders, both in bulk and as single particles, by ultrasonic solvent extraction/capillary electrophoresis. Handgun-fired residues obtained from three common weapon calibers loaded with the known reloading powders were compared with the unfired powders. In general, the composition of the residues was similar to that found in the unfired powders. For double-base powders, comparing the ratio of the propellant (P) to the total amount of stabilizer (S) for both residue and gunpowder samples proved to be a useful measurement for identification. This P/S ratio demonstrated that the additives in the residues did not greatly change relative to the unfired powder, providing a useful indicator to aid in forensic powder and residue evaluation. PMID- 11110173 TI - Detection of sequence variation in the HVII region of the human mitochondrial genome in 689 individuals using immobilized sequence-specific oligonucleotide probes. AB - We have developed a rapid, immobilized probe-based assay for the detection of sequence variation in the hypervariable segment II (HVII) of the mitochondrial DNA (mtDNA) control region. Using a panel of 17 sequence-specific oligonucleotide (SSO) probes immobilized on nylon membrane strips, we typed 689 individuals from four population groups. The genetic diversity value for each population was calculated from the frequency data, and the frequencies of distinct "mitotypes" in each group were determined. We performed DNA sequence analysis of 129 samples to characterize the sequences associated with "blanks" (absence of probe signals) and weak probe signals. Out of 689 samples, we observed five heteroplasmic samples (excluding the variable C-stretch beginning at position 303) using the immobilized SSO probe panel. The SSO probe strips were used for the analysis of shed hairs and bloodstains from several criminal cases in Sweden, one of which is described here. We conclude that this mtDNA typing system is useful for human identification and significantly decreases casework turnaround time. PMID- 11110175 TI - Diimide-enhanced fingerprint detection with photoluminescent CdS/dendrimer nanocomposites. AB - The chemical development of latent fingerprints by nanocomposites that involve photoluminescent cadmium sulfide nanoparticle aggregates with Starburst dendrimer is demonstrated. The dendrimer bonds to fingerprint residue via its terminal functional groups. When these are amino groups (generation 4 dendrimer), the binding is enhanced by fingerprint pre-treatment with diimide. The diimide converts carboxylic acid moieties of the fingerprint residue to esters that then react with the dendrimer amino groups to form amide linkages. The cadmium sulfide/generation 4 dendrimer development of fingerprints is enhanced by elevated temperature also. Finally, fingerprint development with carboxylate functionalized cadmium sulfide/generation 3.5 dendrimer nanocomposites is examined. Here, diimide treatment of the dendrimer itself aids the subsequent fingerprint labeling, which involves amino acid of the figerprint residue. Nanocomposite fingerprint detection is compatible with time-resolved imaging for background fluorescence elimination. PMID- 11110176 TI - Improved method for shooting distance estimation. Part III. Bullet holes in cadavers. AB - An improved method to estimate firing distance on human body surfaces is described. The novel part of the method includes a chemical test in addition to the traditional visual and microscopic examinations of the gunshot wounds. This chemical test consists of a transfer of the gunpowder residues from the area of a gunshot wound to an adhesive lifter; the residues are then visualized as total nitrite after alkaline hydrolysis by the Modified Griess Test (MGT). When cadavers are in an advanced stage of decomposition or when gunshot wounds are in hairy areas, the information obtained by this chemical test can be crucial for shooting distance evaluation. In other cases it may improve the accuracy of the examination. In some cases, the results obtained by this test may assist in the discrimination between entrance and exit gunshot wounds. PMID- 11110177 TI - Time since discharge of rifles. AB - The estimation of time since the latest discharge of rifles has been achieved by the SPME sampling technique and the GC-TEA analytical system. An unidentified compound, designated as the TEA2 compound, was detected in all the rifles investigated. The same compound was observed in shotguns and spent cartridges in our previous work. This compound escapes rapidly from the inside of rifle barrels, but can still be detected there one to two months after the shooting. The decrease of the TEA2 peak with time after shooting is non-exponential, and the curve-fitting procedure proposed for the estimation of time since discharge of shotguns can be applied also for rifles. PMID- 11110179 TI - Ballistic characterization of the Remington Premier Copper Solid sabot shotgun slug. AB - We evaluated the impact and penetration characteristics of the Remington Copper Solid sabot shotgun slug with standardized ballistic tests and used this information to predict tissue wounding patterns. This unique ammunition, first distributed in 1993, is composed of a solid copper, hollow-point slug with longitudinal slots cut into the nose. The slug is fitted into a hard plastic sabot with 8 finger-like projections and loaded into a shotgun shell with two plastic wads separating it from the underlying gunpowder charge. The ammunition was fired through a 12-gage shotgun using a rifled barrel, a smooth-bore barrel with rifled choke, and a smooth-bore barrel with a smooth modified choke into targets consisting of poster board and 10% ballistic gelatin at a variety of distances. The copper slug and plastic sabot created single 8-fingered asterisk shaped defects in the poster board when fired at distances of less than 7 to 9 ft (approximately 2 to 3 m). All three barrel types performed similarly. At greater distances, the sabot impacted the targets separately from the slugs and created variably shaped defects that reflected base-first, nose-first, and side-first impacts. Increasing muzzle-to-target distances generally increased the impact distances between the slug and sabot. There was no predictable relationship between the sabot and slug impact points for any of the three barrel types. With each barrel tested, the wads created separate defects from the slug at distances greater than 5 ft (1.5 m). The distances between the slug and the wad impact points increased with increasing muzzle-to-target distances up to 40 ft (12 m), after which the wads generally no longer struck the targets. The slug created atypical defects at distances between 7 and 150 ft (approximately 2 to 45 m), probably due to yawing or tumbling. When the slug impacted the gelatin block in a nose-first orientation, the slotted nose portion tended to fragment and radially deposit pieces in the gelatin that were visible on radiographs. When the slug struck the gelatin target in a side-first orientation, it passed through the gelatin intact. The slug, sabot, and wads of this unique projectile separate and create independent impact points in a stereotypical manner independent of barrel type. This pattern of separation allows estimates to be made of ranges of fire. Wounds created in human tissues by this ammunition would likely have similar asterisk-shaped configurations, and nose fragments may be deposited in tissues and seen radiographically. Rectangular wounds created by the tumbling or yawing slug might be mistaken for intermediate target wounds. PMID- 11110178 TI - Suicide: a ten-year retrospective study. AB - Suicide is a complex phenomenon associated with psychological, biological, and social factors, claiming approximately 30,000 lives each year in the United States. We retrospectively reviewed all cases referred to the Medical Examiners' Office/Forensic Pathology Section at the Medical University of South Carolina from January 1988 to December 1997. The cases of suicide totaled 678. All of the cases were analyzed as to age/race/sex, method of suicide, time of year, and toxicological results. Files were also reviewed to determine if the victim left behind a suicide note. The ages ranged from 12 to 94 years; males comprised 79.5% of the victims, and whites 78.3%. The male to female and white to black ratios were both 3.9:1. The most common methods were gunshot wounds, accounting for 64.6% of the cases. No correlation existed with time of year, and the number of cases was not increased around major holidays. The group of victims 65 years and older and the pediatric group under the age of 18 were also examined separately. PMID- 11110180 TI - Comparison of patterns of decomposition in a hanging carcass and a carcass in contact with soil in a xerophytic habitat on the Island of Oahu, Hawaii. AB - Decomposition studies were conducted to determine differences in rates and patterns of decomposition of carcasses hanging and exposed on the surface of the soil. These studies were conducted between 17 October and 17 December 1997 inside of Diamond Head Crater on the island of Oahu, Hawaii. The animal model was the domestic pig. Sus scrofa. The rate of biomass removal from the hanging carcass was significantly slower than that observed for the control carcass during the bloat and decay stages of decomposition. Internal temperatures for the control carcass were elevated above the ambient air temperatures during the earlier stages of decomposition (bloated and decay), while those recorded for the hanging carcass approximated the ambient air temperatures. There was a greater diversity of arthropod species recorded and numbers of individuals observed were higher for the control carcass. A significant site of arthropod activity was observed on the surface of the soil immediately under the hanging carcass and this became the primary site of arthropod activity as decomposition progressed. PMID- 11110181 TI - Asphyxial deaths and petechiae: a review. AB - Conjunctival and facial petechiae, although nonspecific findings, are considered hallmarks of asphyxial deaths. Consensus in the literature suggests that their pathogenesis is related to the combined effects of increased cephalic venous pressure and hypoxic damage to endothelial cells. Despite the common knowledge that they are neither predictable findings in all asphyxial deaths nor rare in natural, nonasphyxial deaths, the belief persists that petechiae are corroborative evidence of asphyxia. We suggest that a clear, physiologically based understanding of the pathogenesis of petechiae of the head is critical for their appropriate interpretation. We present a review of the literature and the basis of our conclusion that conjunctival and facial petechiae are the product of purely mechanical vascular phenomena, unrelated to asphyxia or hypoxia. PMID- 11110182 TI - The multidisciplinary approach to dealing with families: a model for medical examiners. AB - Medical examiners function primarily to determine cause and manner of death, and to document illness and injuries. Equally important, however, is the role of providing family members with the initial tools to work through the grief process. By initiating contact with the family, facilitating access to other appropriate services and by making provisions for adequate viewing and meeting facilities, we fulfill these ethical duties and carry out our responsibilities as physicians. We hope to fill some of the void that exists in the forensic medical literature with this presentation and generate awareness for the neglected role of grief intervention. PMID- 11110183 TI - Algal colonization of submerged carcasses in a mid-order woodland stream. AB - One of the primary goals of forensic pathology is the determination of time of death. In aquatic systems, one method to do this is to analyze the colonization of a corpse by algae. Algal communities typically follow a serial colonization pattern, therefore the taxa present at any given time may provide clues about postmortem submersion time. This study was undertaken to examine the algal colonization on rat carcasses in a medium-order woodland stream. Two habitats were studied: a low flow pool and a high flow riffle, with rats being removed from each site every 3 to 6 days over 31 days. The diversity of colonizing taxa increased at both sites as the study progressed, and after 17 days similar taxa were present (Sorensen's similarity index >60%) in each site. Some taxa, such as desmids (Chlorophyta), tended to increase in diversity throughout the study, making them possible indicators of submersion time. Diatoms were the most abundant taxa found in each site and accounted for 63 of the 92 total taxa identified. Due to their ubiquitous presence in nearly all streams, we suggest that diatoms may be the key organisms for the study of postmortem submersion in lotic systems. PMID- 11110184 TI - Separation of visibly-excited fluorescent components in fingerprint residue by thin-layer chromatography. AB - The use of lasers for the detection of fingermarks is widespread in the forensic field. Despite this, and the fact that many studies have been conducted into the composition of fingermark residue, the components responsible for the inherent visible fluorescence remain unidentified. Traditionally compositional studies have been performed on sweat, sebum, or skin surface washes, none of which are truly representative of the situation when a fingerprint is deposited on a surface. In this paper thin-layer chromatography (TLC) has been performed on sebum-rich fingermarks laid directly onto TLC plates and an argon ion laser used to visualize the separated components. It has been found to be a robust and reproducible method for studying the fluorescent components in fingermark residue and is considered to be more realistic than other methods of sample preparation as it eliminates the chances of extraneous matter being extracted from the skin surface. Investigations into the nature of the separated compounds have also been made and the results are reported. PMID- 11110185 TI - A new visibly-excited fluorescent component in latent fingerprint residue induced by gaseous electrical discharge. AB - A technique that exposes fingerprint residue to a gaseous electrical discharge in nitrogen followed by treatment with ammonium hydrogen carbonate vapors to produce fluorescence is investigated. Particular attention is made to fluorescence observed via laser illumination at 514 nm. Insight into the nature of the fluorescent components is achieved through the use of thin-layer chromatography (TLC) of fingerprint residue. Results reported indicate the fluorescence observed is from previously non-fluorescent fractions of the fingerprint residue, and TLC results point towards lipid derivatives as a possible source of the fluorescence. PMID- 11110186 TI - A method for collection of gunshot residues from skin and other surfaces. AB - A method of collecting gunshot residues from the skin of persons who have been injured by firearms has been developed. The method uses a commercially available, adhesive, transparent plastic film. This method is also useful for collecting gunshot residues from other objects, such as leather. The shooting distance is later estimated by ocular, microscopic or IR examination in combination with various chemographic tests. PMID- 11110187 TI - The accelerated polyvinyl-alcohol method for GSR collection--PVAL 2.0. AB - The polyvinyl-alcohol collection method (PVAL) is used in forensic practice to gather topographical information about gunshot residues (GSR) from the hands to decide if the subject has made use of firearms. The results allow a distinction between suicide and homicide. The only inconvenience of PVAL was that the procedure took about 60 min because three layers of liquid PVAL had to be applied and dried. Therefore, the collection method was only applied to corpses. The improved and accelerated PVAL 2.0 uses a sandwich technique. Cotton gauze for stabilization is moistened with a 10% PVAL solution. A solid film of PVAL (Solublon) is spread on the cotton mesh. The gauze is then modeled to the hand and dried with a hair dryer. After removing the cotton gauze, the traces are embedded in the water-soluble PVAL. The procedure does not take more than 15 min. The results demonstrate the qualities and advantages of PVAL: topographical distribution of GSR, highest gain of GSR, sampling of all other traces like blood, backspatter etc., and humidity does not reduce the gain. In addition, with the new PVAL 2.0 dislocation of GSR or contamination are excluded. PVAL 2.0 can also be applied on live suspects. PMID- 11110188 TI - A systematic and quantitative analysis of PCR template contamination. AB - A quantitative and systematic analysis is provided for ubiquitously present template DNA interfering with the quantification of human DNA by PCR. Two sources contributing to DNA background were identified. The first one is interpreted as DNA present in chemicals and on equipment and the second as caused by operator handling. The amounts were equivalent to 2.5 and 8.9 pg per mL of sample, and the estimated frequencies of contamination were 65 and 35%, respectively, resulting in an effective limit of detection of 17.4 pg/mL. Below this level--named effective laboratory background--a result could not be considered as authentic. Knowledge of these parameters is important for laboratories that analyze minute amounts of human DNA by PCR for purposes such as quantification, typing, and sequencing. PMID- 11110189 TI - Paint and tape: collection and storage of microtraces of paint in adhesive tape. AB - The collection and preservation of microtrace evidence with the aid of an adhesive tape is a method of choice in forensic science. This technique is rapid and easy and allows the concentration of microtraces on a carrier, which facilitates further investigations in the laboratory. Adhesive tapes are currently used to secure microtraces of fibers and glass, but hardly for traces of automotive paint and other lacquers for fear of interference with the analysis of binders. A collection of automotive paint consisting of original and repair lacquers collected by tape has been evaluated. After various times of storage within the tape, these samples were compared with untreated references by microscope FT-IR and microspectrophotometry (MSP). Another set of paints was collected in 1984, stored within the tape until 1995, and examined the same way. About 170 layers of lacquer with various types of binder were examined. With the exception of one clear lacquer no difference between treated samples and references was detected. This small difference observed could be correlated to the exposure to xylene (extractant) and was not caused by the storage within the adhesive tape. PMID- 11110190 TI - Restoration tactics for seriously corroded Cu and Cu-alloy firearms evidence. AB - A cleaning procedure for seriously weathered or corroded bullets and cartridge casings was developed and implemented for evidence specimens from a multiple homicide. The restoration protocol entailed successive treatments with increasingly aggressive chemical solvents and cleaning solutions while monitoring the progress of the method by optical microscopy. Treatment of worst-case, Cu alloy jacketed bullets and casings resulted in reconditioned specimens that subsequently underwent successful firearms examinations. PMID- 11110191 TI - Distribution of HLA-DQA1, polymarker, CSF1PO, vWA, TH01, TPOX, D16S539, D7S820, D13S317, and D5S818 alleles in East Bengali and West Punjabi populations from Indo-Pak Subcontinent. AB - Blood samples were collected from 115 individuals residing in the Pakistani state of West Punjab and 81 Bengali individuals residing in the state of East Bengal, India. These samples were analyzed for the loci HLA-DQA1, PM (LDLR, GYPA, HBGG, D7S8, and GC) and eight short tandem repeats: CSF1PO, TPOX, THO1, vWA, D16S539, D7S820, D13S317, and D5S818. Departures from Hardy-Weinberg (HWE) were observed in Punjabi population at LDLR, THO1, D13S317, D5S818, and D16S539 and at CSF1PO and THO1 in Bengali population. PMID- 11110192 TI - Northeast Italy population data using multiplex PCR (HUMCD4, HUMTH01, HUMTPOX, and HUMCSF1P0) loci. AB - Allele and genotype frequencies for four short tandem repeat (STR) loci (HUMCD4, HUMTH01, HUMTPOX, and HUMCSF1P0) were determined in 100 unrelated individuals from Veneto (Northeast Italy). After a multiplex polymerase chain reaction (PCR) amplification, semi-automatic DNA profiling was performed using an A.L.F.express DNA Sequencer. Conditions were optimized for the PCR co-amplification of these four STR loci and the quadruplex PCR was performed on various forensic DNA samples such as whole blood, blood-stains, teeth, and saliva from Caucasians living in the Northeast Italy. The distribution of the genotype frequencies showed no significant deviation from Hardy-Weinberg expectations in the sampled population. The combined Power of Discrimination (PD) of the quadruplex was 0.9999. PMID- 11110193 TI - Screening of amphetamine/methamphetamine and their derivatives in urine using FPIA and Triage 8 and the Scope and limits of a subsequent identification by means of the REMEDi HS system. AB - This study describes screening and identifying amphetamines, methamphetamines, and their derivatives in urine using immunochemical (Triage, FPIA) and chromatographic techniques (REMEDi HS). Amphetamines, methamphetamines, MDMA (3,4 methylenedioxymethamphetamine), MDA (3,4-methylenedioxyamphetamine), MDE (3,4 methylenedioxyethyl-amphetainine), MBDB (N-methyl-1-(3,4-methylenedioxyphenyl)-2 butanamine), BDB (3,4-(methylenedioxyphenyl)-2-butanamine), PMA (4 methoxyamphetamine), DOM (2,5-dimethyloxy-4-methylamphetamine), DOB (4-bromo-2,5 dimethyloxyamphetamine), amphetaminil, pholedrine, fenfluramine, and amfepramone were subjected to a comparative study. For this, the substances were analyzed to determine their specific threshold concentration for a positive detection in the Triage test and their limit of detection and positive threshold concentration for the FPIA test and the results compared. Furthermore, the capabilities of a more detailed analysis with the REMEDi system were studied. This HPLC system was able to produce information on the single drugs and main metabolites found in the sample with the danger of false-positive or false-negative screening results greatly minimized. PMID- 11110194 TI - Paternity-testing on paraffin-embedded abortion tissue: preparation of fetal cells may be indispensable. AB - In a case of sexual abuse, a paternity test was performed on paraffin embedded abortion material. STR typing was successful only after isolating fetal tissue from the abortion-material and separately extracting DNA from the excised fetal cells. Examination with five STRs led to a paternity index of 332, confirming the abuse that had resulted in pregnancy. PMID- 11110195 TI - Metastatic calcification of the cardiac conduction system with heart block: an under-reported entity in chronic renal failure patients. AB - Systemic metastatic calcification is a common complication of chronic renal failure. Cardiac involvement is particularly ominous, especially when the cardiac conduction system is affected. Conduction defects, arrhythmias, and sudden death have all been reported with conduction system calcification; however, these are relatively under-reported or unrecognized causes of cardiac morbidity and mortality. We describe a 40-year-old man with Von Hippel-Lindau disease who had been maintained on hemodialysis for two years following bilateral nephrectomies for renal cell carcinoma. The patient presented with symptomatic complete heart block that had progressed from Mobitz type I atrioventricular block. Two months later, while being internally paced, the patient died unexpectedly after a complicated hospital admission. Postmortem revealed extensive vascular, myocardial, and conduction system calcification. Conduction system calcification may cause sudden death in chronic renal failure patients during hospital admission, or unexpectedly while the patient is in the community. Knowledge of this condition is necessary to detect it, as the conduction system is not routinely examined. A routine abbreviated conduction system examination is warranted for patients with systemic metastatic calcification, especially if they have sudden death or a known history of heart block. PMID- 11110196 TI - Detection and correction of a migration anomaly on a 310 genetic analyzer. AB - During STR analysis on the 310 Genetic Analyzer, retarded migration of GS500ROX size standards and alleles in some samples was observed. The contribution of reagents, capillary and performance optimized polymer POP 4 to the observed anomaly was experimentally eliminated. Variation in electrophoresis temperature between 55 degrees C and 65 degrees C did not alter the rate of migration of GX500ROX size standard and sample alleles. An eroded connector for the cathode mounted on the heat plate assembly caused the abnormal migration. Hence, it is important to verify the mobility of all fragments in the size standard for each sample to avoid any erroneous allele calls by the automated data analysis software. PMID- 11110197 TI - Sequence polymorphisms of the mitochondrial DNA control region in 100 German Caucasians. PMID- 11110198 TI - Allelic distribution for two short tandem repeat loci: D10S2325 and D20S161. PMID- 11110199 TI - Allele frequency distributions for three STR loci in the Han and Thai populations. PMID- 11110200 TI - STR data for the AMPFlSTR profiler plus loci among four predominant populations of Eastern India. PMID- 11110201 TI - Analytical methods for ignitable liquid residues. PMID- 11110202 TI - A DNA-based approach to the identification of insect species used for postmortem interval estimation and partial sequencing of the cytochrome oxydase b subunit gene I: a tool for the identification of European species of blow flies for postmortem interval estimation. PMID- 11110203 TI - Minimal standards for the performance and interpretation of toxicology tests in legal proceedings and on the commentary of Kidwell DA, Smith FP. J Forensic Sci 2000;45(1):237-239. PMID- 11110204 TI - Promega Corporation reveals primer sequences in its testing kits. PMID- 11110205 TI - Ecological risks of diazinon from agricultural use in the Sacramento-San Joaquin River Basins, California. AB - A probabilistic risk assessment was conducted to evaluate the likelihood and ecological significance of potential toxic effects of diazinon in the Sacramento San Joaquin system. Diazinon, an organophosphorus insecticide, is used in the Sacramento-San Joaquin River Basin as a dormant spray on almonds and other tree crops, as well as for other agricultural and urban applications. Diazinon and other pesticides have been detected in the Sacramento and San Joaquin Rivers and their tributaries. Diazinon exposure was characterized based on monitoring programs conducted in 1991-94. Diazinon effects were characterized using laboratory toxicity data for 63 species, supplemented by results from field mesocosm and microcosm studies. The assessment addressed the possibility that reductions in invertebrate populations could lead to impacts on species of fish that feed on those invertebrates. The risk assessment concluded that fish in these rivers are not at risk from the direct effects of diazinon in the water. Invertebrates are at greater risk, especially in agriculturally dominated streams and drainage channels during January and February. Cladocerans--including Daphnia magna and Ceriodaphnia dubia, two common bioassay species--are especially sensitive to diazinon and other organophosphates and are likely to be subject to acute toxic effects in some locations at some times. Any ecological damage that may occur, however, is brief and limited to cladocerans. None of the fish species of concern depend on cladocerans as critical components of their diet. Invertebrates that are not affected by observed concentrations of diazinon (copepods, mysids, amphipods, rotifers, and insects) are preferred foods for fish in the Sacramento-San Joaquin system. PMID- 11110206 TI - Generating probabilistic spatially-explicit individual and population exposure estimates for ecological risk assessments. AB - Exposure to chemical contaminants in various media must be estimated when performing ecological risk assessments. Exposure estimates are often based on the 95th-percentile upper confidence limit on the mean concentration of all samples, calculated without regard to critical ecological and spatial information about the relative relationship of receptors, their habitats, and contaminants. This practice produces exposure estimates that are potentially unrepresentative of the ecology of the receptor. This article proposes a habitat area and quality conditioned exposure estimator, E[HQ], that requires consideration of these relationships. It describes a spatially explicit ecological exposure model to facilitate calculation of E[HQ]. The model provides (1) a flexible platform for investigating the effect of changes in habitat area, habitat quality, foraging area, and population size on exposure estimates, and (2) a tool for calculating E[HQ] for use in actual risk assessments. The inner loop of a Visual Basic program randomly walks a receptor over a multicelled landscape--each cell of which contains values for cell area, habitat area, habitat quality, and concentration--accumulating an exposure estimate until the total area foraged is less than or equal to a given foraging area. An outer loop then steps through foraging areas of increasing size. This program is iterated by Monte Carlo software, with the number of iterations representing the population size. Results indicate that (1) any single estimator may over- or underestimate exposure, depending on foraging strategy and spatial relationships of habitat and contamination, and (2) changes in exposure estimates in response to changes in foraging and habitat area are not linear. PMID- 11110207 TI - Efficient sensitivity/uncertainty analysis using the combined stochastic response surface method and automated differentiation: application to environmental and biological systems. AB - Estimation of uncertainties associated with model predictions is an important component of the application of environmental and biological models. "Traditional" methods for propagating uncertainty, such as standard Monte Carlo and Latin Hypercube Sampling, however, often require performing a prohibitive number of model simulations, especially for complex, computationally intensive models. Here, a computationally efficient method for uncertainty propagation, the Stochastic Response Surface Method (SRSM) is coupled with another method, the Automatic Differentiation of FORTRAN (ADIFOR). The SRSM is based on series expansions of model inputs and outputs in terms of a set of "well-behaved" standard random variables. The ADIFOR method is used to transform the model code into one that calculates the derivatives of the model outputs with respect to inputs or transformed inputs. The calculated model outputs and the derivatives at a set of sample points are used to approximate the unknown coefficients in the series expansions of outputs. A framework for the coupling of the SRSM and ADIFOR is developed and presented here. Two case studies are presented, involving (1) a physiologically based pharmacokinetic model for perchloroethylene for humans, and (2) an atmospheric photochemical model, the Reactive Plume Model. The results obtained agree closely with those of traditional Monte Carlo and Latin hypercube sampling methods, while reducing the required number of model simulations by about two orders of magnitude. PMID- 11110208 TI - Variations in concepts of "susceptibility" in risk assessment. AB - The Food Quality Protection Act and the 1996 amendments to the Safe Drinking Water Act are two of the most recent examples of legislation calling for protection of susceptible subpopulations. As regulatory deadlines draw nearer, controversies in scientific and policy arenas increase about incorporating susceptibility in risk assessment. The previously accepted working definition of "susceptibility" has already been called into question. Part of the controversy results from different disciplines conceiving of susceptibility in different ways. Understanding the conceptual differences embodied within definitions can provide a basis on which a revised working definition may be developed across disciplines. The purposes of this article are to describe the varying definitions of susceptibility, discuss the differing concepts incorporated in the definitions, and recommend ways in which susceptibility may be defined and framed to meet current risk assessment needs. The present analysis of definitions from the fields of ecology, biology, engineering, medicine, epidemiology, and toxicology revealed different emphases that relate to the underlying perspectives and methods of each field. It is likely that susceptibility will need to be formally defined for public policy purposes, but until that time, the use of more informal communication and decision-making processes is suggested to develop and utilize a new working consensus on the definition of susceptibility. PMID- 11110209 TI - Exposure assessment of heavy metals resulting from farmland application of wastewater sludge in Tianjin, China: the examination of two existing national standards for soil and farmland-used sludge. AB - Land application is one of the major methods of managing municipal sludge in China. The sludge is used for fertilizing and conditioning soil, but due to the high concentration of heavy metals and other chemicals that it contains, improper use of sludge will lead to the contamination of farmland soil. To provide guidance on the application of sludge in China, the Control Standards for Pollutants in Sludge for Agricultural Use (CSPSAU) were enacted, and implemented in 1985. Afterwards, the National Environment Quality Standards for Soil (NEQSS) were also formulated and put into effect in 1996. In this article, these two national standards were examined by means of exposure assessment. The main exposure pathway to humans that was considered was dietary intake of crops grown on the sludge-applied farmland. Five major types of agricultural crops (rice, wheat, tuber roots, vegetables, and fruits) and three groups of exposure population (the urban individual group, the rural sludge-applying individual group, and the rural sludge nonapplying individual group) were assessed. This case study in Tianjin, China, shows the necessity of reexamining the national standards of the CSPSAU and the NEQSS in the context of risk assessment. More comprehensive surveys and monitoring programs assessing heavy metals contained in farmland soils and crop tissues will be necessary for examining the risks to human health. PMID- 11110210 TI - Daily soil ingestion estimates for children at a Superfund site. AB - Ingestion of contaminated soil by children may result in significant exposure to toxic substances at contaminated sites. Estimates of such exposure are based on extrapolation of short-term-exposure estimates to longer time periods. This article provides daily estimates of soil ingestion on 64 children between the ages of 1 and 4 residing at a Superfund site; these values are employed to estimate the distribution of 7-day average soil ingestion exposures (mean, 31 mg/day; median, 17 mg/day) at a contaminated site over different time periods. Best linear unbiased predictors of the 95th-percentile of soil ingestion over 7 days, 30 days, 90 days, and 365 days are 133 mg/day, 112 mg/day, 108 mg/day and 106 mg/day, respectively. Variance components estimates (excluding titanium and outliers, based on Tukey's far-out criteria) are given for soil ingestion between subjects (59 mg/day)2, between days on a subject (95 mg/day)2, and for uncertainty on a subject-day (132 mg/day)2. These results expand knowledge of potential exposure to contaminants among young children from soil ingestion at contaminated sites. They also provide basic distributions that serve as a starting point for use in Monte Carlo risk assessments. PMID- 11110211 TI - Assessment of airborne exposure to trihalomethanes from tap water in residential showers and baths. AB - This study evaluates airborne concentrations of common trihalomethane (THM) compounds in bathrooms during showering and bathing in homes supplied with chlorinated tap water. Three homes in an urban area were selected, each having three bedrooms, a full bath, and approximately 1,000 square feet of living area. THMs were concurrently measured in tap water and air in the shower/bath enclosure and the bathroom vanity area using Summa canisters. Chloroform (TCM), bromodichloromethane (BDCM), and chlorodibromomethane (CDBM) were quantified using U.S. Environmental Protection Agency (EPA) Method TO-14. Air samples were collected prior to, during, and after the water-use event for 16 shower and 7 bath events. Flow rate and temperature were measured, but not controlled. The increase in average airborne concentration (+/- standard error) during showers (expressed as microg/m3 in shower enclosure or bathroom air per microg/L in water) was 3.3+/-0.4 for TCM, 1.8+/-0.3 for BDCM, and 0.5+/-0.1 for CDBM (n = 12), and during baths was 1.2+/-0.4 for TCM, 0.59+/-0.21 for BDCM, and 0.15+/ 0.05 for CDBM (n = 4). The relative contribution of each chemical to the airborne concentrations was consistent for all shower and bath events, with apparent release of TCM > BDCM > CDBM. The results are therefore consistent with their relative concentration in tap water and their vapor pressures. When the shower findings for TCM are normalized for water concentration, flow rate, shower volume, and duration, the average exposure concentrations in these urban residences are about 30% lower than those reported by other investigators using EPA analytical methods. This difference is likely attributable primarily to greater air exchange rates in residential shower/bath stalls compared to more "airtight" laboratory shower chambers. This appears to be the first field study to thoroughly evaluate THM exposures from residential showers and baths, and can be used to validate previously published models of tap water volatile chemical transfer to indoor air. PMID- 11110212 TI - Are the members of a paving crew uniformly exposed to bitumen fume, organic vapor, and benzo(a)pyrene? AB - The goal of this investigation was to assess if and when a crew of paving workers is uniformly exposed to bitumen fume, organic vapor, and benzo(a)pyrene. Data on paving workers with up to six repeated exposure measurements were extracted from a database of exposure measurements developed within a study of the European asphalt industry (N = 591). The uniformity of exposures to bitumen fume, organic vapor, and benzo(a)pyrene was evaluated while grouping individuals by job title, primary tasks, crew membership, and use of coal tar (discontinued in Western Europe). The estimated ranges within which 95% of individual mean exposures were expected to fall ((BW)R0.95) were used to assess exposure uniformity. Variance components were estimated by constructing mixed-effects models, with grouping variables as fixed effects and worker identity as random effect. The influence of duration of the sampling survey on estimates of exposure variability for a crew was also examined. There was a substantial variability in exposures between paving crews, as well as persons holding the same job or doing the same task, but each crew was uniformly exposed to bitumen fume and benzo(a)pyrene ((BW)R0.95 2 and 1, respectively). Workers within the same crew engaged in paving with coal tar-containing binders were not, however, uniformly exposed to benzo(a)pyrene. Also, organic vapor exposures were not uniform among the members of a paving crew ((BW)R0.95 = 15). Sampling campaigns of up to 7 months had little impact on the estimates of within- and between-worker variability. These findings should assist investigators studying paving operations in optimizing their sampling, exposure assessment, and risk evaluation protocols. The results support the notion that only empirically determined predictors of exposure can yield optimal grouping, unlike a priori grouping strategies based on general descriptors such as jobs title or tasks performed. PMID- 11110213 TI - Risk/risk trade-offs in pesticide regulation: an exploratory analysis of the public health effects of a ban on organophosphate and carbamate pesticides. AB - Efforts to reduce pesticide-related risks to consumers and farmworkers often neglect the possibility that measures to reduce the target risk may introduce or enhance countervailing risks. These may arise from substitute pesticides or pest control practices, from increased levels of pests or pest-related hazards, from increased levels of toxic natural pesticides in plants, from increased costs and decreased consumption of health-enhancing fruits and vegetables, or from direct income effects on consumers and farmers. The effect of the countervailing risks may partially or completely offset the reduction in the target risk. A risk-trade off analysis was conducted of a potential ban on the use of organophosphate and carbamate (OP/Carbamate) insecticides in U.S. agriculture. Although this scenario is extreme, it has the analytic virtue of dispensing with the infinite number of "next-best" OP/Carbamates that might be substituted for specific combinations of crops and pests should only selected uses be banned. The analysis relies on detailed descriptions of the alternative pesticides and pest-control measures that would be used for each of 14 major crops. The effects of pest-control cost changes on prices and consumption and effects on consumer and producer incomes are projected using a general-equilibrium economic model. Several countervailing risks that may be significant were found, including acute toxicity to farmworkers from substitute pesticides, cancer and noncancer risks from substitute pesticides, and mortality induced by changes in disposable income. Other countervailing risks are more difficult to estimate or weigh. Potential increases in natural plant pesticides following an OP/Carbamate ban are discussed but data are lacking to quantify the effects. Changes in diet following the ban have both positive and negative effects, and the ultimate change is difficult to estimate. Although a net risk cannot be estimated, several approaches were illustrated that would be useful in risk-trade-off analyses. Key factors complicating comprehensive analysis of risk/risk trade-offs for pesticides were also identified, including data gaps and shortcomings of current risk assessment methods. PMID- 11110214 TI - Risk perception in the U.K. oil and gas production industry: are expert loss prevention managers' perceptions different from those of members of the public? AB - This article investigates potential differences in risk perception between experts (loss-prevention managers in the U.K. oil and gas production industry) and nonexperts (managers and students). Extant research on expert versus nonexpert perceptions of risk is reviewed, followed by the present study concerning risk perceptions of seven pen-picture scenarios involving the occurrence of hazardous events in the U.K. oil and gas production industry. In contrast to many of the earlier studies of expert versus nonexpert perceptions of risk, the present analysis concludes that experts did not judge the overall riskiness of the portrayed hazardous events as less risky than the nonexperts. Nevertheless, the experts believe more strongly than our nonexperts that the risks portrayed in the scenarios pose little threat to future generations, are more precisely known, and are relatively controllable. Use of multiple regression analysis to help uncover the basis of overall riskiness assessments for expert and lay respondents was inconclusive, however. Finally, little evidence was found that nonexperts were any more heterogeneous in their risk perceptions than experts. It may be that the nature of the risks assessed in the present study may account for the general lack of clear expert versus nonexpert differences in overall perceptions of the riskiness of hazardous events in the North Sea. Earlier findings of strong expert versus nonexpert differences in risk perception assessed hazards of major public concern. It is inferred that using such extreme hazards may have resulted in an exaggerated view of differences in expert versus public (nonexpert) perception of risk. PMID- 11110215 TI - Public perceptions of health risks from polluted coastal bathing waters: a mixed methodological analysis using cultural theory. AB - This article explores public perceptions of, and attitudes toward, possible health risks from polluted coastal bathing waters in the United Kingdom. Cultural theory is applied in the present analysis, using a mixed methodology of quantitative analysis from interviews and qualitative interpretation of focus group discussions to provide insights into how different cultural solidarities view a number of issues. These include risks to health; attitudes toward regulation; public consultation and information provision; and trust, blame, and accountability applied to different stakeholders in the bathing-water-quality debate. The results show that individuals' standpoints can be represented on a number of dimensions, consistent with cultural theory, including perceptions of power and authority, beliefs in the efficacy of collective action, and acceptance or rejection of incremental change as opposed to radical solutions. The discussion focuses both on methodological and substantive issues related to the use of cultural theory as a research tool, and on policy recommendations arising from this research. PMID- 11110216 TI - Do people actually pursue risk elimination in environmental risk management? AB - This study examines whether people pursue the total elimination of environmental risks even if the risk reduction occurs incrementally and requires steadily increasing amounts of money to achieve. Participants' willingness to pay (WTP) was measured for various levels of reduction in the risk of cancer caused by dioxin. Results showed that: (1) people were willing to pay more for an initial reduction in risk than for subsequent reductions, (2) the WTP for the final risk decrement-which achieved total risk elimination-was higher than the WTP for either of the two previous decrements but barely half of that for the first reduction, and (3) the manner in which the questions were framed did not affect participants' responses. These results suggest that the public will not always try to pursue perfect safety at the cost of a large sum of money, but rather may seek a decreasing expenditure as the risk level is reduced. PMID- 11110217 TI - Perception of hazards: the role of social trust and knowledge AB - Recent research indicates that social trust of those who manage a hazard is strongly correlated to judgments about the hazard's risk and benefits. The present study investigates the more specific question of "For which hazards is this?" It was postulated that when an individual lacks knowledge about a hazard, social trust of authorities managing the hazard determines perceived risks and benefits. On the other hand, when an individual has personal knowledge about a hazard and therefore does not need to rely on managing authorities, social trust is unrelated to judged risks and benefits. Participants (N = 91) assessed risks, benefits, and trust in managing authorities and personal knowledge associated with 25 hazardous technologies and activities. As expected, strong correlations between social trust and judged risks and benefits were observed for hazards about which people did not possess much knowledge. No significant correlations between social trust and judged risks and benefits were found for hazards about which people were knowledgeable. Results suggest that the lay public relies on social trust when making judgments of risks and benefits when personal knowledge about a hazard is lacking. Replicating findings of other studies, the present study also found negative correlations between perceived risks and perceived benefits. When social trust was controlled for, correlations between perceived risks and benefits diminished. Implications of the results for risk management are discussed. PMID- 11110218 TI - Protection motivation and risk communication AB - The purpose of this study was to explore the utility of protection motivation theory (PMT) in the context of mass media reports about a hazard. Content elements of a hazard's severity, likelihood of occurring, and the effectiveness of preventive actions were systematically varied in a news story about a fabricated risk: exposure to fluorescent lighting lowering academic performance. Results of this experiment (N = 206) suggest that providing information about the severity of a hazard's consequences produces greater information seeking. In addition, information about levels of risk, severity, and efficacy combined jointly to produce greater rates of willingness to take actions designed to avoid the hazard. Results are seen as providing general support for PMT and are discussed within the broader framework of information seeking and heuristic and systematic information processing. PMID- 11110219 TI - A comparison of organic and conventional fresh produce buyers in the Boston area. AB - Food safety concerns and the demand for organically grown produce have increased significantly in the United States over the last decade. Key differences in lifestyle characteristics, food safety attitudes and beliefs, perceived food safety risks, and valuation of health risk reductions between organic and conventional food buyers remain largely unknown, however. To better characterize how buyers of organic fresh produce differ from their conventional counterparts, over 700 food shoppers were sampled from ten major retail stores in the Boston area. Survey results show that self-reported organic buyers are more likely than conventional buyers to engage in a variety of health-promoting and environmentally friendly behaviors. Organic buyers are less trusting of federal food safety agencies than are conventional buyers, and perceive greater benefits associated with organically grown produce than do their conventional counterparts. Further, organic buyers have significantly higher risk perceptions than do conventional buyers for food safety hazards associated with conventionally grown produce. Compared to conventional buyers, organic produce buyers also perceive significant risk reductions associated with switching to organically grown produce and are willing to pay a higher price to reduce perceived food safety risks. Few sociodemographic differences between buyer types were observed, possibly due to how organic and conventional food stores were matched. Survey findings highlight the need for greater public education about a range of food safety issues and farming practices to ensure that consumers are making informed decisions in the marketplace. PMID- 11110220 TI - From breeder reactors to butterflies: risk, culture, and biotechnology. AB - Social theories of risk suggest that a combination of scientific and cultural perspectives converge to influence risk perception. This article first surveys sociological perspectives suggesting that risks from modern technological development have become predominant concerns in the social consciousness. Particular attention is given to those theses describing how social elements work to create perception of risks in relation to new technologies. The themes that emerge from this survey are then related to contemporary debates concerning biotechnology. Specific attention is given to recent controversies regarding genetically modified crops, and parallels are drawn between debates over nuclear power and biotechnology. A procedural ethic for public discourse and decision making over the diffusion of genetically modified foods is offered. Ethical and social theories are linked with the hope that by recognizing the social dimensions of debates over new technologies a broader framework for conducting risk analysis may emerge. PMID- 11110222 TI - Why did they lie? Socio-economic bias in reporting menarcheal age. AB - PRIMARY OBJECTIVE: To show that socio-economically dependent cultural bias distorts results of the status quo method of estimating age at first menstruation. METHODS: Questionnaires asking for menstrual status and the recalled age at menarche were distributed to approximately 1000 Javanese girls who attended junior and senior high schools in Malang. Age of participants ranged from 11.98 to 18.89 years. Probit analysis was applied to the status quo data while average menarcheal age recalled by girls > or =16 year old was also calculated. t-test, F-test, ANOVA and chi2 tests were applied to test significance of differences between groups. MAIN OUTCOMES AND RESULTS: Girls of low occupational status fathers (Group 1) had a probit average menarcheal age of 13.99 years (SD = 1.33, n = 524). Girls of medium occupational status fathers (Group 2), had a lower probit average menarcheal age (13.06 SD = 1.38, n = 315). Girls of fathers with the highest occupational score (Group 3) rarely reported premenarcheal status (less than 10% in all age groups studied) and the probit analysis of their reports yielded an unbelievable average of 9.61 (SD = 3.41, n = 157) years. Group 3 girls tend to report their menarcheal status incorrectly, probably due to a 'fashion' of appearing mature or to 'contagion' during filling out of a questionnaire. The recalled average menarcheal age of Group 3 females questioned when they were aged 16 years and above, yields an average that seems to be more reliable (12.74 years, SD = 1.41, n = 7), because at a fully postmenarcheal age there is no need to enhance one's status by falsely claiming maturity. PMID- 11110221 TI - Modelling health-related performance indices. AB - PRIMARY OBJECTIVE: The purpose of the present review is to critically examine the various models used to describe health-related performance indices (e.g. grip strength, lung function, arterial blood pressure, anaerobic and aerobic power, etc.). MAIN OUTCOMES AND RESULTS: The majority of health-related performance indices are known to increase proportionally with body size and decline with age, although some, for example arterial blood pressure and cholesterol, increase with both age and excessive body weight. Historically, the confounding effects of body size and age have been controlled using multiple linear or polynomial regression models assuming a constant additive error variance. Based on maximum likelihood goodness-of-fit criterion, an alternative class of allometric models with a multiplicative error term arc shown to describe such proportional relationships more adequately in both cross-sectional and longitudinal studies. CONCLUSIONS: When modelling health-related performance indices, multiplicative, allometric models incorporate the concept of a proportional association as an integral part of its model form. Such models guarantee realistic values of health-related response variables both within and beyond the range of observations, ensuring biologically sound estimates of the response variables for subjects of all ages. After a logarithmic transformation, the models can be fitted using ordinary linear regression that will naturally help to overcome the heteroscedasticity observed in such data. However, if the logarithmic transformation fails to provide a constant error variance, an alternative approach is to model the error variance itself using log-linear regression. For longitudinal data, the use of multilevel modelling provides an alternative method of modelling error variation observed in health-related performance indices. PMID- 11110223 TI - Energy expenditure, physical exertion and time allocation among Huli-speaking people in the Papua New Guinea Highlands. AB - OBJECTIVES: The study aimed to (1) elucidate the energetic adaptations of Huli people by comparing the three sub-populations in relation to their diversified natural and socioeconomic environment, based on energy expenditure and time allocation data; and (2) assess the applicability of a new index of physical exertion levels of activities (physical exertion index: PEI). RESEARCH DESIGN AND METHODS: Married males and females (n = 43) were selected, including 14 from a hilly village, 13 from a flat swampy village and 16 migrants to a town. Continuous heart rate (HR) monitoring, in conjunction with minute-by-minute observation of activities, was undertaken. Total energy expenditure was assessed by flex-HR method and physical activity level (PAL) was calculated as multiples of basal metabolic rate. Observed activities were divided into 15 categories and the PEI was calculated for each: PEI = (mean HR of a categorized activity)/(flex HR) x 100. RESULTS: No significant difference was found in PAL among the three sub-populations: 1.77, 1.92 and 1.81 for men and 1.78, 1.98 and 1.66 for women in hilly and flat villages, and a town, respectively. The comparison of the two village groups revealed that hilly terrain did not affect substantially the physical intensity of agricultural activities. On the other hand, the town migrants were engaged in sedentary paid jobs and these were less energy consuming than agricultural work. However, the different energy costs between paid jobs and agricultural work were offset by longer work time in the town group than in the two village groups (251 vs 50 or 70 min in males; 258 vs 152 or 138 min in females), resulting in similar PAL in the three groups. CONCLUSIONS: Despite contrasting natural and social environments, no significant difference was found in daily physical activity level among the three sub-populations (two villages and a town) of Huli-speaking people in Papua New Guinea Highlands. PMID- 11110224 TI - The distribution and geographical origin of some French surnames. AB - The geographical distribution of the principal form of a French surname and its close variants (in spelling or pronunciation) has been analysed based on the French births registers, respectively from 1891 to 1915 and from 1916 to 1940, and on telephone directories for 1975. The spatial analysis of the individuals with the principal form of the surname suggests the existence of a positive spatial autocorrelation and three major areas of concentration in central France, the Loire country and Northern France. Within these three main areas of concentration, in 1975 most of the telephone subscribers with the principal surname are listed in rural communes with fewer than 3000 inhabitants, suggesting that the geographical pattern is not recent. Moreover, between the end of the 19th century and 1975 this pattern remains fairly constant. The geographical distribution of the holders of a 'variant' form of the surname shows the existence of a positive correlation between the absolute frequency of the principal surname and that of the variants, but only one area of concentration appears in Loire country. By analogy with the theory of centres of origin in genetics and linguistics, it is suggested that this unique area of concentration could be the region in which the surname originated. If that hypothesis proves to be correct, two main consequences follow: large migration movements occurred before the general rural exodus of the last century, and there has been no major, recent admixture of populations. PMID- 11110225 TI - Elements of the surname structure of Austria. AB - The isonymy structure of Austria was studied using the surname distributions in 1081002 private telephone users selected from about 4000000 registered in a 1996 commercial CD-ROM, which contains all Austrian users. The sample was distributed in 120 towns representing an approximately uniform distribution over the country. The number of different surnames found in the whole analysis was 140766. Lasker's distance, the negative value of the logarithm of isonymy between localities, was found to be linearly and significantly correlated with the log of geographic distance, with r = 0.565 +/- 0.011. A dendrogram was built with the matrix of isonymy distance, using the Unweighted Pair-Group Method using Arithmetic averages, UPGMA. It separates the Austrian towns in five main clusters, one along the central portion of the country, another one which occupies the northern region of central Austria; then comes a third cluster at the north-eastern part, a fourth cluster in the western region, and finally a small cluster towards the border with Slovenia. Within each, small subclusters with specific geographic distributions could be delimited. The main clusters correspond fairly well to the classic regions of Austria. The results were compared with those obtained in similar analyses of Switzerland, Germany, Italy and Venezuela. From the present analysis, isolation by distance emerges clearly, and it is stronger than in Germany but smaller than that observed in Italy, Switzerland and Venezuela. The random component of inbreeding estimated from isonymy, at the level of resolution used here, indicates that the inbreeding level in Austria is rather uniform. PMID- 11110226 TI - Allometric relationship between body size and peak VO2 relative to age at menarche. AB - The present study examined allometric coefficients relating peak VO2 and body size relative to the time of menarche. Peak oxygen uptake (peak VO2) during exercise on a bicycle ergometer, stature and body mass were measured at annual intervals in a mixed-longitudinal sample of 40 active girls from 11 to 14 years of age. The girls were interviewed about their menarcheal status at each examination. The data were treated relative to the time before and after menarche: 2 years before (n = 18), 1 year before (n = 26), during the year of menarche (+/- 6 months, n = 32), 1 year after (n = 35) and 2 years after menarche (n = 22). Allometric coefficients were calculated for each of the five menarcheal groups based on logarithmic transformations of peak VO2 and body mass and peak VO2 and stature. The major axis of VO2 and body mass or stature (log transformed) was also calculated. This is the most appropriate slope for comparison with theoretical allometry coefficients. Mean peak VO2 increases from 2.1 +/- 0.19 L 2 years before menarche to 2.3 +/- 0.26 L 2 years after menarche. The slope of the major axis for body mass is always higher (0.508-0.926) than that for the allometric coefficient (0.323-0.591) in each of the menarcheal groups. The major axis slope and allometric coefficient are lowest between body mass and peak VO2 during the year of menarche. The slope of the major axis is below the theoretical allometric coefficients assuming geometric or elastic similarity, 2/3 or 3/4, before and at menarche and increases after menarche. Although the differences are not statistically significant, the results suggest that the relationship between body mass and peak VO2 at menarche is lower compared with relationships before and after this maturational landmark. Allometric coefficients for stature relative to peak VO2 show a similar pattern. PMID- 11110227 TI - The genetics of hand-clasping: a review and a familial study. PMID- 11110228 TI - Rosiglitazone in the treatment of type 2 diabetes mellitus: a critical review. AB - OBJECTIVE: This article reviews the pharmacology, pharmacokinetics, clinical efficacy, adverse effects, drug interactions, and dosing of rosiglitazone, the second thiazolidinedione approved for the treatment of type 2 diabetes mellitus. METHODS: Background information for this article was obtained from searches of MEDLINE , Iowa Drug Information Service, and International Pharmaceutical Abstracts, as well as from data on file with the manufacturer of rosiglitazone. RESULTS: Rosiglitazone is indicated for use alone or in combination with metformin or sulfonylureas for the maintenance of glycemic control in patients with type 2 diabetes mellitus. Rather than stimulation of insulin secretion, rosiglitazone's primary mechanism of action is sensitization of tissues to insulin through activation of the peroxisome proliferator-activated receptor gamma and increasing expression of the glucose transporter-4 receptor. Rosiglitazone is administered orally, is absorbed almost completely, and is 99.8% bound to plasma proteins. The majority of a dose is metabolized by the cytochrome P-450 2C8 isozyme, with the inactive metabolites excreted primarily in the urine. Four to 8 mg/d of rosiglitazone given alone or in combination with metformin, sulfonylureas, or insulin has produced reductions in baseline fasting plasma glucose and glycosylated hemoglobin in studies of up to 1 year's duration. Common adverse effects (occurring in > or = 5.0% of patients) include upper respiratory tract infection, injury, and headache. Edema, weight gain, and increased low density lipoprotein cholesterol concentrations have also been observed. It is recommended that rosiglitazone be avoided in patients with alanine aminotransferase levels >2.5 times normal. No clinically relevant drug interactions have been documented with rosiglitazone to date. The initial starting daily dose of rosiglitazone is 4 mg in single or divided doses, without regard to meals, to a maximum of 8 mg. CONCLUSIONS: No direct comparative trials of the efficacy and safety of rosiglitazone versus those of the other available thiazolidinedione, pioglitazone, have yet been performed. The role of rosiglitazone as a single agent and in combination with other antidiabetic agents remains to be clarified as additional comparative and long-term data become available. PMID- 11110229 TI - Clinical experience with pantoprazole in gastroesophageal reflux disease. AB - BACKGROUND: Pantoprazole is a new proton pump inhibitor indicated for the treatment of erosive esophagitis associated with gastroesophageal reflux disease (GERD) and is available in both oral and intravenous (IV) formulations. OBJECTIVE: This paper reviews the pharmacologic properties of pantoprazole and summarizes the findings from clinical studies of this drug. METHODS: This review was compiled from the published literature, abstracts from clinical trials, and data on file with the manufacturer of pantoprazole. RESULTS: Pantoprazole selectively accumulates in the acidic environment of gastric parietal cells and acts at the final step of acid secretion by binding 2 key cysteine residues of the proton pump involved in gastric acid production. The bioavailability of pantoprazole is not altered by concomitant administration of food or antacids or with repeated dosing. Both oral and IV formulations of pantoprazole exhibit linear pharmacokinetics. Several clinical trials have proved pantoprazole superior to histamine-2-receptor antagonists (H2RAs) in reducing acid secretion and elevating gastric pH levels. Pantoprazole has been shown to be more effective than ranitidine (P < 0.05), famotidine (P < 0.001), and nizatidine (P < 0.05), and at least as effective as omeprazole, in healing erosive esophagitis and relieving associated symptoms of GERD, including regurgitation. Pantoprazole is also more effective than the H2RA nizatidine for the treatment of nighttime heartburn (P < 0.05). Studies have shown pantoprazole to be well tolerated; adverse events, including headache, diarrhea, flatulence, abdominal pain, eructation, nausea, and rash, occurred in < or = 6% of patients. The oral and IV formulations of pantoprazole are equally potent in inhibiting gastric acid secretion; thus, switching between formulations requires no dosage adjustments. Special patient populations, including the elderly and patients with renal or mild to moderate hepatic impairment, can take pantoprazole without an adjustment in dosage. CONCLUSIONS: Because of its unique pharmacokinetic properties, mechanism of action, and reduced potential for producing cytochrome P-450-based drug interactions, pantoprazole in both oral and IV formulations is effective over a full 24 hours and is well tolerated in a variety of patient types. PMID- 11110230 TI - Effects of losartan and candesartan monotherapy and losartan/hydrochlorothiazide combination therapy in patients with mild to moderate hypertension. Losartan Trial Investigators. AB - OBJECTIVE: The goal of this multicenter, double-blind, randomized, parallel-group study was to compare the effects of losartan potassium (hereafter referred to as losartan), candesartan cilexitil (hereafter referred to as candesartan), and losartan/hydrochlorothiazide (HCTZ) in patients with mild to moderate hypertension (sitting diastolic blood pressure [SiDBP] 95-115 mm Hg). METHODS: A total of 1161 patients were randomized in a 2:2:1 ratio to 12 weeks of treatment with losartan 50 mg QD, possibly titrated to 100 mg QD (n = 461); candesartan 8 mg QD, possibly titrated to 16 mg QD (n = 468); or losartan 50 mg QD, possibly titrated to losartan 50 mg plus HCTZ 12.5 mg QD (n = 232). At 6 weeks, the regimens of patients not reaching a goal SiDBP <90 mm Hg were titrated as described, whereas patients achieving this goal continued with low-dose monotherapy. The single primary end point at 12 weeks tested the equivalence of the 2 monotherapy regimens, predefined as a maximum between-treatment difference in the mean change from baseline trough SiDBP of 2.5 mm Hg. RESULTS: At 12 weeks, changes in SiDBP/sitting systolic blood pressure (SiSBP) of -12.4/-14.4 mm Hg with losartan 50 mg/100 mg and -13.1/-15.8 mm Hg with candesartan 8 mg/16 mg demonstrated equivalence between the 2 monotherapy regimens (95% CI for difference in SiDBP, -1.6 to 0.2). At 12 weeks, the losartan 50 mg/50 mg plus HCTZ 12.5 mg regimen had reduced SiDBP/SiSBP significantly more (-14.3/-18.0 mm Hg) than either the candesartan 8 mg/16 mg (SiDBP, P = 0.045; SiSBP, P = 0.017) or losartan 50 mg/100 mg regimen (SiDBP and SiSBP, P = 0.001). During the last 6 weeks, patients whose regimen had been titrated to losartan 50 mg plus HCTZ 12.5 mg (n = 114) showed a greater reduction in SiDBP/SiSBP (-14.5/ -18.7 mm Hg) than did those whose regimen had been titrated to either losartan 100 mg (-10.5/-12.3 mm Hg; n = 211) or candesartan 16 mg (-11.5/-13.2 mm Hg; n = 206), representing a clinically meaningful > or = 2.5-mm Hg) difference. All 3 treatments were well tolerated, with few patients experiencing drug-related adverse events (6.9% losartan 50 mg/100 mg, 7.5% candesartan 8 mg/16 mg, 3.0% losartan 50 mg/ 50 mg plus HCTZ 12.5 mg). Candesartan 8 mg/16 mg increased serum uric acid levels (0.13 mg/dL; 95% CI, 0.04 to 0.23), whereas losartan 50 mg/100 mg decreased them (-0.14 mg/dL; 95% CI, -0.24 to -0.04), and losartan 50 mg/50 mg plus HCTZ 12.5 mg left them unchanged (0.06 mg/dL; 95% CI, -0.07 to 0.20). CONCLUSIONS: Losartan 50 mg/100 mg and candesartan 8 mg/16 mg were comparable treatments in terms of blood pressure reduction. After titration, losartan 50 mg plus HCTZ 12.5 mg was superior to either candesartan 16 mg or losartan 100 mg in reducing hypertension. Losartan, but not candesartan, lowered serum uric acid levels and attenuated the expected increase in uric acid levels with HCTZ 12.5 mg. PMID- 11110231 TI - Patients' acceptance of a switch from dorzolamide to brinzolamide for the treatment of glaucoma in a clinical practice setting. AB - BACKGROUND: The first topically active carbonic anhydrase inhibitor, dorzolamide, was developed to circumvent the adverse systemic effects of oral carbonic anhydrase inhibitors. However, its use has been associated with ocular discomfort. OBJECTIVE: The present study examined the acceptability of brinzolamide, as measured by patients' ratings and stated preferences, in patients with glaucoma previously treated with dorzolamide in the clinical practice setting. METHODS: This was a prospective, open-label, noncomparative study conducted shortly after the approval of brinzolamide. Ophthalmologists in private practice in the continental United States were asked to select patients currently using dorzolamide as their sole or combination therapy for glaucoma. Patients underwent a screening assessment in which they were asked to rate their ocular comfort with dorzolamide on a scale from 1 to 6. Brinzolamide was then substituted for dorzolamide, and patients returned for a follow-up visit approximately 1 to 3 months later. At this visit, patients were asked about ocular comfort, their preferred medication, and whether they thought ocular comfort influenced their adherence to treatment. Intraocular pressure (IOP) was measured at both visits. RESULTS: Valid visit dates (ie, both baseline and follow up dates) were available for 447 of 501 patients from 68 of 73 sites (range, 1-40 patients per site). Because not all measurements were available for all patients at each visit, the sample size varied for each measurement. Demographic data were not available. The switch to brinzolamide resulted in a mean decrease in IOP of approximately 0.8 mm Hg (P < 0.001, paired t test). Sixty-nine percent of patients (274/397) reported an improvement of > or =1 grade in their comfort rating with brinzolamide versus dorzolamide. The mean (+/- SD) improvement in comfort rating was 1.43 +/- 1.48 grades (P < 0.001, Wilcoxon rank sum test). When patients were asked whether their adherence to treatment was affected by the occurrence of burning and stinging, 43% (173/399) answered affirmatively. Fifty nine percent (251/424) preferred brinzolamide to dorzolamide. At the end of the study, based on patient preference, physician judgment, and other factors, 73% of responding patients (301/410) continued with brinzolamide therapy. CONCLUSIONS: In this study, the switch from dorzolamide to brinzolamide resulted in overall improvements in comfort and ocular hypotensive efficacy. However, studies using a more rigorous randomized, controlled, crossover design are needed to support these observations. PMID- 11110232 TI - A multicenter, randomized, double-blind study of the antihypertensive efficacy and tolerability of irbesartan in patients aged > or = 65 years with mild to moderate hypertension. AB - BACKGROUND: Blockade of the renin-angiotensin-aldosterone system (RAAS) is the preferred mechanism of action for controlling hypertension in select groups of patients, including those with diabetic nephropathy and heart failure. Currently, 2 classes of drugs work by blocking the RAAS, albeit by differing mechanisms: angiotensin-converting enzyme (ACE) inhibitors and angiotensin II angiotensin type 1 receptor blockers (ARBs). OBJECTIVE: The goal of this study was to assess the comparative efficacy and tolerability of the ARB irbesartan and the ACE inhibitor enalapril in patients > or = 65 years of age with mild to moderate hypertension (sitting diastolic blood pressure [DBP], 95 to 110 mm Hg). METHODS: Elderly (> or = 65 years of age) patients were recruited from 26 Canadian study centers for a randomized, double-blind, 8-week clinical trial. Exclusion criteria included sitting DBP >110 mm Hg or sitting systolic blood pressure (SBP) >200 mm Hg, angina pectoris, myocardial infarction, cardiac procedure, stroke, or transient ischemic attack within 6 months of randomization, as well as other preexisting or present severe medical or psychologic conditions. Patients were randomly assigned to receive a single daily dose of irbesartan 150 mg (n = 70) or enalapril 10 mg (n = 71) with treatment doses of study drugs doubled at week 4 for sitting DBP > or = 90 mm Hg. Reductions from baseline blood pressure measurements at trough (24 +/- 3 hours after the last dose of medication) were assessed for sitting DBP and sitting SBP. Comparative tolerability to study drugs was also assessed. RESULTS: The intent-to-treat analysis demonstrated similar reductions at week 8 in both DBP and SBP for both groups. For the primary efficacy analysis of sitting DBP, there was a mean reduction from baseline of 9.6 mm Hg and 9.8 mm Hg for the irbesartan and enalapril groups, respectively (P = 0.93). The mean reduction from baseline in sitting SBP was 10.1 mm Hg and 11.6 mm Hg for the irbesartan and enalapril groups, respectively (P = 0.54). Normalization rates (sitting DBP <90 mm Hg) at week 8 did not differ between groups (52.9% in the irbesartan group and 54.9% in the enalapril group; P = 0.81). No statistical difference existed between the 2 groups with respect to serious adverse events or discontinuations due to adverse events. Irbesartan was associated with a significantly lower incidence of cough than was enalapril (4.3% vs 15.5%, respectively; P = 0.046). CONCLUSIONS: Irbesartan is an effective and well-tolerated antihypertensive for elderly patients with mild to moderate hypertension. This study establishes that irbesartan has better tolerability than enalapril with respect to cough and suggests that irbesartan is as effective at lowering blood pressure but better tolerated than an ACE inhibitor in hypertensive patients > or = 65 years of age. PMID- 11110233 TI - A comparison of tazarotene 0.1% gel once daily plus mometasone furoate 0.1% cream once daily versus calcipotriene 0.005% ointment twice daily in the treatment of plaque psoriasis. AB - BACKGROUND: Both tazarotene (a retinoid prodrug) and calcipotriene (a synthetic analog of vitamin D3) are effective in the treatment of plaque psoriasis, but no reports in the literature directly compare the efficacy and tolerability of these 2 drugs. Tazarotene is commonly used in conjunction with a topical corticosteroid. In this study, tazarotene was used with mometasone furoate (a synthetic corticosteroid), and the 2-drug regimen was compared with calcipotriene monotherapy. OBJECTIVE: This study was conducted to compare the efficacy and tolerability of tazarotene 0.1% gel once daily plus mometasone furoate 0.1% cream once daily with those of calcipotriene 0.005% ointment twice daily in the treatment of plaque psoriasis. METHODS: In this multicenter, investigator blinded, parallel-group study, adult patients with chronic, stable plaque psoriasis affecting 5% to 20% of their body surface area were randomly allocated to receive up to 8 weeks of treatment with either tazarotene 0.1% gel once daily (in the evening) plus mometasone furoate 0.1% cream once daily (in the morning) or calcipotriene 0.005% ointment twice daily. Patients were assessed at baseline and at weeks 2, 4, and 8 of treatment. Patients who demonstrated complete clearance of plaque psoriasis after 2 or 4 weeks of treatment and those whose psoriasis had improved > or = 50% after 8 weeks of treatment entered a 12-week posttreatment follow-up phase during which they applied only moisturizer. Patients were reassessed after 4, 8, and 12 weeks of posttreatment follow-up. Physician-rated measures of efficacy included global improvement, plaque elevation, scaling, erythema, and percentage of body surface area involvement. Patient-rated assessments included efficacy of study treatment compared with previous therapies, comfort of treated skin, outlook for long-term control of psoriasis, and overall impression of treatment. RESULTS: Of 120 patients with moderate to severe psoriasis enrolled from 3 centers, 106 (88%) completed the study. No significant differences in baseline clinical variables were observed between the 2 groups. Twenty-seven patients (45%) in the tazarotene plus cortico steroid group achieved marked improvement (> or = 75% global improvement) after 2 weeks of treatment compared with 15 patients (26%) in the calcipotriene group (P < or = 0.05). Between-group comparisons of the percentage of patients achieving complete or almost complete clearance (> or = 90% global improvement) did not reach statistical significance at any time point. When compared with the calcipotriene regimen, the tazarotene plus corticosteroid regimen resulted in significantly greater efficacy on trunk lesions in reducing plaque elevation (at the end of treatment and at week 4 of the posttreatment phase, P < or = 0.05), scaling (week 4 of treatment and week 4 of the posttreatment phase, P < or = 0.05), erythema (week 4 of treatment and at the end of treatment, P < or = 0.05), and percentage of body surface area involvement (weeks 2 and 4 of treatment, P < or = 0.01). In addition, the tazarotene plus corticosteroid regimen was significantly more effective in reducing the percentage of body surface area involvement in upper limb lesions (weeks 2 [P < or = 0.05] and 4 [P < or = 0.01] of treatment). Forty-two of 55 patients (76%) in the tazarotene plus corticosteroid group rated their medication as more or much more effective than previous therapies compared with 30 of 52 patients (58%) in the calcipotriene group (P < or = 0.05). Although adverse events (burning, pruritus, irritation, and erythema) occurred in a significantly greater proportion of patients who received tazarotene plus corticosteroid than in those who received calcipotriene (P < or = 0.05), 47 of 55 patients (85%) in both groups rated the comfort of their treated skin as "somewhat comfortable" or better and both groups had similar discontinuation rates due to treatment-related adverse events (3% and 5%, respectively). CONCL PMID- 11110234 TI - Retrospective analysis of the safety profile of oral and intravenous ciprofloxacin in a geriatric population. AB - BACKGROUND: Ciprofloxacin is a broad-spectrum fluoroquinolone antibiotic used in the treatment of a wide range of mild to moderate gram-positive and gram-negative infections. Although extensive information is available on the safety profile of ciprofloxacin in adults, few published data exist regarding the tolerability and toxicity of this drug in patients aged > or = 65 years. OBJECTIVES: The objectives of this retrospective analysis were to compare the safety profile of ciprofloxacin in patients aged > or = 65 years versus patients aged <65 years and to compare the adverse-event profile of ciprofloxacin with that of other comparator antimicrobial agents used in clinical trials. METHODS: We retrospectively reviewed 23 prospective, controlled anti-infective clinical trials in the US Bayer ciprofloxacin database that included patients aged > or = 65 years. These trials comprised the submission file of the original and supplemental New Drug Application for ciprofloxacin. The incidence of treatment emergent and drug-related adverse events was assessed. RESULTS: Of the 6863 patients in these 23 clinical trials, 3579 received ciprofloxacin therapy and 3284 received comparator antimicrobial agents. Of the ciprofloxacin-treated patients, 898 (25.1%) were aged > or = 65 years; 887 (27.0%) of the patients who received comparator antimicrobial agents were aged > or = 65 years. Among ciprofloxacin-treated patients, drug-related adverse events were reported more often in those aged <65 years (24.0%) compared with those aged > or = 65 years (17.9%). The incidence of drug-related adverse events in the comparator group was also higher in those aged <65 years (25.1%) than in those aged > or = 65 years (16.8%). Premature discontinuation due to any adverse event was reported in 3.9% (105 of 2681) and 3.7% (33 of 898) of ciprofloxacin-treated patients aged <65 years and > or = 65 years, respectively. Corresponding rates for the comparator antimicrobial group were 3.9% (93 of 2397) and 3.8% (34 of 887), respectively, for patients aged <65 years and > or = 65 years. The most common drug-related adverse events reported for ciprofloxacin-treated patients aged <65 years and > or = 65 years were digestive system-related (18.1% and 11.4%, respectively) and central nervous system-related events (6.6% and 4.9%, respectively). CONCLUSIONS: This retrospective analysis suggests that there is no clinically important difference in the safety profile of ciprofloxacin in patients aged > or = 65 years versus patients aged <65 years. PMID- 11110235 TI - The efficacy of naftidrofuryl in patients with vascular or mixed dementia. PMID- 11110236 TI - Functional response of Euseius finlandicus and Amblyseius andersoni to Panonychus ulmi on apple and peach leaves in the laboratory. AB - The functional response of adult females of the predatory mites Euseius (Amblyseius) finlandicus and Amblyseius andersoni to larvae and adult females of the fruit tree red spider mite Panonychus ulmi was determined on apple and peach leaf disks in the laboratory at 25 degrees C and 16:8 (L:D). For adult females of P. ulmi the predation efficiency of E. finlandicus was higher on peach than on apple, whereas that of A. andersoni was higher on apple than on peach. Efficiency of predation on larvae of P. ulmi by either predator did not differ significantly between apple and peach. On both plants, A. andersoni had a higher predation rate than E. finlandicus on larvae of P. ulmi. It is concluded that in the laboratory the host plant has a substantial effect on predation efficiency of A. andersoni and E. finlandicus when they preyed on adults but not when they preyed on larvae of P. ulmi. PMID- 11110237 TI - The effect of non-pathogenic phylloplane fungi on life-history traits of urticae (Acari: Tetranychidae). AB - The direct effect of three common non-pathogenic phylloplane fungi on the life history traits of Tetranychus urticae Koch was investigated on intact leaves of the garden bean Phaseolus vulgaris L. (var. Tendergreen Improved), under conditions of low and high water-deficit stress. The survival rate of T. urticae was always reduced by an increase in water-deficit stress, whereas the effect of the fungi depended in part on the watering regime. On two of the three fungi tested, Alternaria alternata (Fr.) Keissler and Epicoccum nigrum Link, the mites showed a significant increase in net reproduction compared to those reared under control (i.e. no fungi added) conditions, independent of water-deficit stress, resulting in a higher intrinsic rate of increase. The third fungus tested, Cladosporium cladosporioides (Fres.) de Vries, had a similar response with respect to net reproduction when reared under low water-deficit stress but not when reared under a high water-deficit stress. There was, however, no evidence of an interactive effect with water-deficit stress and presence or absence of fungus affecting the net reproduction of the mites, for any of the three fungal species tested. An interactive effect between fungal species and water-deficit stress was observed for the intrinsic rate of increase of the mites. It is suspected that the fungi are acting as a source of an otherwise limiting resource and the differences observed between the fungal treatments is due to differences in consumption, based on the different sizes of their conidia. PMID- 11110238 TI - Disaggregation of aggregated platelets by apyrase from the tick, Ornithodoros savignyi (Acari: Argasidae). AB - Apyrase, secreted by ticks during feeding, is a platelet aggregation inhibitor that functions as a regulator of the host's hemostatic system. This present study concerns the disaggregation effect of salivary gland apyrase from the tick Ornithodoros savignyi. Secondarily aggregated platelets, disaggregated by apyrase, exhibited a reversal of shape from a spherical (aggregated) form to a discoid form, reminiscent of reversible aggregation at low ADP concentrations in citrated platelet-rich plasma. However, they showed a dilatory open canaliculary system and an absence of granules indicating disaggregation after degranulation had taken place. In contrast, disaggregation by the fibrin(ogen)olytic enzyme, plasmin, showed that platelets degranulated, but retained a spherical form with numerous extended pseudopods. While thrombin had no effect on aggregation or clotting of platelets disaggregated with plasmin, it did activate those platelets disaggregated with apyrase and clotted the plasma. This is the first study to describe the disaggregating effects of tick derived apyrase on aggregated platelets. It also shows that apyrase can disaggregate platelets even after secondary aggregation and degranulation of platelets has taken place. Platelet aggregation is one of the main barriers encountered by ticks during feeding and counteraction of this process by ticks is an important factor for successful feeding. PMID- 11110239 TI - Does geographic range affect the attractant-aggregation-attachment pheromone of the tropical bont tick, amblyomma variegatum? AB - The tropical bont tick, Amblyomma variegatum, transmits heartwater in sub-Saharan Africa and in the Caribbean. This species has a broad geographic distribution, ranging from Madagascar and other islands in the Indian Ocean through most of sub Saharan Africa, to several islands in the eastern Caribbean Sea. Blood fed male A. variegatum secrete an attraction-aggregation-attachment (AAA) pheromone which, combined with CO2, excites host finding and formation of feeding clusters of these ticks. However, it is not known whether the composition of the pheromone varies throughout A. variegatum's geographic range. Extracts of fed male ticks were examined for phenols and volatile organic acids by gas chromatography and mass spectrometry to determine whether differences occur in the pheromone components of populations of this species across the geographic range (Guadeloupe, Zimbabwe, Zambia and Rwanda). No significant difference in the chemical composition of the pheromone in relation to geographic range was found. No significant differences in rates of attachment in response to native versus foreign extracts were found in on-host attachment tests comparing ticks from two countries. Guadeloupe (Caribbean) and Zimbabwe (African). This finding was confirmed in more detailed studies with ticks from Guadeloupe and four African countries (Kenya, Rwanda, Zambia and Zimbabwe). On-host attachment assays from these countries did not detect consistent differences in response to extracts from different locations. In an olfactometer bioassay, females were not consistently more attracted to extracts from their native locality than from any of the foreign localities. We conclude that despite the widespread distribution of A. variegatum over both hemispheres, no significant differences in pheromone composition or biological responses to male tick pheromone secretions occur. PMID- 11110240 TI - Feeding patterns of immature stages of Hyalomma truncatum and Hyalomma marginatum rufipes on different hosts. AB - In this study we examine the feeding patterns of immature stages of Hyalomma truncatum and Hyalomma marginatum rufipes ticks on different hosts. Larvae of H. truncatum developed through a three-host pattern on two species of field mice, Rhabdomys pumilio and Lemniscomys rosalia. On guinea-pigs, both Hyalomma species followed a mixed two-host and three-host pattern, with the latter route being preferred, since more than 70% of the fully fed larvae dropped off from their hosts. H. truncatum was a two-host tick on rabbits. Larvae of H. marginatum rufipes did not prefer R. pumilio and L. rosalia as hosts. On guinea-pigs, H. marginatum rufipes immatures showed a mixed two-host and three-host pattern with a bias towards the three-host life cycle, since approximately 58% of the fully fed larvae dropped off. On rabbits, H. marginatum rufipes was exclusively a two host tick. Mean engorgement weights and blood quantities ingested by H. truncatum nymphs that developed through a three-host pattern on mice were significantly higher (p < 0.0001) than for those that developed through a two-host pattern on guinea-pigs and rabbits. For H. marginatum rufipes, there were no significant differences (p > 0.05) between engorgement weights of nymphs that developed through two-host and three-host patterns. However, there were significant differences (p < 0.0001) in blood quantities ingested by nymphs of this tick species following feeding on different hosts. PMID- 11110242 TI - Comparison of five algorithms for the detection of ventricular fibrillation from the surface ECG. AB - The introduction and widening application of automatic external defibrillators (AEDs) present very strong requirements for external ECG signal analysis. Highly accuratc discrimination between shockable and non-shockable rhythms is required, with sensitivity and specificity aimed to approach the maximum values of 100%. We undertook an assessment of the performance of five detection algorithms, selected from among several others for their good published results. Test signals were 71 8 s ECG episodes on sinus rhythm and 90 8 s episodes on ventricular fibrillation, which were taken from the well known ECG-signal databases of the American Heart Association (AHA) and the Massachusetts Institute of Technology Beth Israel Hospital (MIT-BIH-'cudb' and 'vfdb' files). The purpose of this study is to assess the accuracy of the algorithms with signals other than those used for their development. An expected reduction of the sensitivity and specificity was found. The results could be used for further assessment, e.g. of noise and artefact sensitivity, for comparison with newly developed algorithms, etc. PMID- 11110241 TI - The measurement of impedance to assess myocardial contractility and rhythm stability. AB - By introducing an alternating current through the ventricular cavity via a multipolar catheter the potential difference measured between adjacent electrodes can be used to assess ventricular volume. This is possible by the measurement of intracardiac impedance. The data obtained can be used to construct pressure volume loops allowing the continuous real-time assessment of myocardial contractility. Clinical applications of this research tool are now becoming apparent. Further derivations of impedance measurement have been incorporated into commercially available permanent pacing systems, allowing effective heart rate modulation. PMID- 11110243 TI - Spectral components of heart rate variability determined by wavelet analysis. AB - Spectral components of heart rate variability (HRV) are determined in the time frequency domain using a wavelet transform. Based on the finer estimation of low frequency content enabled by the logarithmic resolution of the wavelet transform, corrections of spectral intervals, already defined by Fourier and model based methods, are proposed. The characteristic peaks between 0.0095 and 0.6 Hz are traced in time and four spectral intervals are defined, I (0.0095-0.021 Hz), II (0.021-0.052 Hz), III (0.052-0.145 Hz) and IV (0.145-0.6 Hz), within which peaks are located for all subjects included. These intervals are shown to be invariant regardless of the age and the state of the system. We also show that the frequency and power of the spectral components are related to age, AMI and particularly to type II diabetes mellitus. PMID- 11110244 TI - Image correlation method for measuring blood flow velocity in microcirculation: correlation 'window' simulation and in vivo image analysis. AB - To elucidate the function of the microcirculation system, it is very important to know the blood flow velocity and its distribution in the microvessels. We have developed an automated system for measuring blood flow velocity in microcirculation by image correlation. The 'window' in the image correlation method is equivalent to the sensors in various other measurement methods. We performed simulations with virtual blood flow images consisting of random dots before measuring actual ones, and examined the optimum window shape and size. We found that by reducing the size of a circular window to the size of erythrocytes we could measure in vivo blood flow images with high accuracy. We recorded them with a high-speed video camera system at high temporal resolution, and measured the velocity in microvessels of normal Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHR). SHR had higher blood velocity than WKY even though the vessel diameters were the same. Using this method to measure the blood flow velocity profile at the bent corner of SHR's arteriole at the heart systole, we found that erythrocytes flow faster at the inner side of the bend, so the vessel wall was exposed locally to higher shear stress in the hypertensive condition. PMID- 11110245 TI - Intracranial pressure and clinical status: assessment of two intracranial pressure transducers. AB - The aim of this study was to determine the in vivo accuracy and reliability of intracranial pressure (ICP) measurement using the Codman MicroSensor by comparison with the Camino ICP transducer and associated clinical and radiological information. Paired ICP readings were recorded every minute in 17 patients. A total of 56,790 validated paired measurements were obtained over a wide range of ICP values (- 16 mm Hg to 114 mm Hg). Recording periods ranged from 3 hours to 6 days (median 41 hours). In 11 patients the MicroSensor and Camino readings were in good agreement. Paired readings were within 10 mmHg for 99% of the recording time and both readings were compatible with clinical intracranial events at all times (in one case it was not possible to verify the clinical information). In six patients large differences occurred between transducer readings (> 10 mm Hg apart for 41% of the recording period). In one case, either reading could have been compatible with intracranial clinical events. In two cases, although both readings were comparable, Camino readings were more consistent with clinical and radiological findings. In three cases, the MicroSensor readings were inconsistent with the clinical condition of the patients whereas the Camino readings were compatible. These results suggest that, during routine clinical use in our department, the MicroSensor provided misleading information in 18% of our patients and thus is not sufficiently reliable for routine use in the detection of adverse clinical events. PMID- 11110246 TI - Tissue oxygen saturation measured by near infrared spectrophotometry correlates with arterial oxygen saturation during induced oxygenation changes in neonates. AB - The aim of this study was to compare quantitatively the changes in tissue oxygen saturation (TOS), determined by two algorithms (TOSc and TOSa) based on near infrared spectrophotometry, to the changes in arterial oxygen saturation (SaO2) measured by pulse oximetry. TOSc is an algorithm derived by the manufacturer (Critikon) based on a modified Beer-Lambert law; TOSa, our own algorithm, uses the diffusion approximation of light transport for the semi-infinite boundary condition. Slow changes of more than 3% in SaO2 were carried out in 20 mechanically ventilated neonates by altering the inspired oxygen fraction. For each change the regression lines of TOSc versus SaO2, TOSa versus SaO2 and TOSc versus TOSa were calculatcd. For each infant the mcan slope, intercept and r2 of these lines were determined. In 18 preterm infants we obtained median 9.5 (range one to 13) measurements corresponding to a total of 166 measurements. The mean SaO2 was 91.6 (SD 2.3)%, TOSc was 64.7 (SD 7.2)% and TOSa was 71.4 (SD 11.0)%. Changes in TOSc and TOSa were strongly correlated to changes in SaO2 (r2 = 0.86 and r2 = 0.87). TOSc considerably but systematically underestimated the size of the change: delta TOSc = 0.49 delta SaO2. TOSa quantified changes reasonably correctly: delta TOSa = 0.90 delta SaO2. Changes in TOSc and TOSa were highly correlated (r2 = 0.98). These results are promising, but the large inter individual variation requires further work. PMID- 11110247 TI - An evaluation of the characteristics and performance of neonatal phototherapy equipment. AB - This paper reviews the current state of knowledge and practice in neonatal phototherapy, and assesses methods of evaluating the characteristics and performance of different equipment. Artificial lighting (usually fluorescent) has been used for the past 30 years in the treatment of neonatal jaundice. Widely differing light outputs and spectra are used, making comparison and evaluation difficult for clinicians. Manufacturers of neonatal phototherapy equipment have no standard for assessing the performance of their equipment, and information that is supplied is at best confusing or deceptive to the users. Best practice is usually based on empirical data from equipment in use, but there is wide agreement that present phototherapy is sub-optimal, i.e. does not achieve maximum rate of bilirubin clearance for minimum therapeutic dose. Several studies in the last ten years have emphasized the importance of both the wavelength and intensity of light for optimal phototherapy. These are discussed and a technique is proposed for normalizing the output of different systems to make comparison easier and to enable optimal treatments to be designed. PMID- 11110248 TI - The spatio-chromatic sensitivity of the human visual system. AB - The response of the human visual system depends on a multitude of image features, such as the wavelength (colour) of the visual stimulus and its spatial frequency content. Hence we need to take into account the spatial and chromatic sensitivity as well as spatio-chromatic interactions to properly characterize visual sensitivity. In this paper we report two experiments that further characterize the spatio-chromatic sensitivity of the human visual system for stationary stimuli, namely the detection of small visual orientation differences and the detection of blur. In both cases we find that the visual system is equally sensitive to red-green and to black-white modulations for a wide range of spatial parameters. Furthermore, the contrast dependence for red green and black white modulations is identical, suggesting that the same mechanism mediates both types of stimulus. Our results are in accordance with the hypothesis that both tasks are mediated by the parvocellular as opposed to the magnocellular pathway. PMID- 11110249 TI - A quality control procedure for force platforms. AB - A force platform is a technical method of quantitatively assessing balance indirectly. The use of force platforms in physiotherapy departments has become more prominent over the last few years. However, the main drawback in the use of force platforms is the lack of comprehensive calibration procedures, which casts doubt on the results obtained with these systems. Existing calibration tests are limited to testing the spatial accuracy of the force platform. This paper describes a comprehensive quality control test procedure which was developed. It is proposed that the developed quality control test procedure could be used to test all types of force platform and it includes a description of how the tests should be carried out, the frequency with which they should be carried out and the expected performance for each of the tests as recommended for the most part by the Association Francaise de Posturologie. PMID- 11110250 TI - Development of a novel method for assessing balance: the quantitative posturography system. AB - Balance is the ability to maintain equilibrium while sitting or standing. There are a number of different methods which are used to assess balance: technical methods such as sway magnetometry, ataxia meter and force platforms, and clinical methods such as the functional reach test, Berg balance test and fall risk index. The most frequently used technical method is the force platform. There are two types of force platform, a static and a dynamic force platform, of which the dynamic force platform has been found to be more sensitive to detect impaired balance. The quantitative posturography system (QPS) described in this paper is a type of dynamic force platform; however, it has a simpler design than the currently available dynamic force platforms and can match the subject's sway exactly for tilting in the anterio-posterior and medio-lateral directions with its novel design. This paper describes the novel design of the QPS and its calibration. PMID- 11110251 TI - Continuous monitoring of single-sweat-gland activity. AB - A conductivity measurement system using a small ion-free-solution perfusion chamber has been developed to monitor single sweat-gland activity (SSGA) continuously at the skin surface. The chamber has a small open space of 0.2 mm2 at the bottom and has a transparent window. Single sweat pores were visualized by the starch/iodine method and the chamber was attached onto a single sweat pore using a magnifying lens affixed at the window. Silver electrodes were installed inside the chamber, and, by perfusing ion-free solution through the chamber at a constant flow rate, the conductivity of the solution was measured at the inlet and the outlet of the chamber. Continuous SSGA was monitored at the palm, finger tip and chest skin surface when the subjects were seated in a resting position and under stresses such as hand grasping with a dynamometer and performing mental arithmetic. Different types of response were observed from different sweat pores. The response time of this system was less than 0.15 s. The present results reveal that continuous sweat activity can be monitored even from a single sweat gland. PMID- 11110252 TI - DXA body composition studies are not affected by extracellular water measurements using stable sodium bromide dilution. AB - Body composition studies using dual energy x-ray absorptiometry (DXA) are being increasingly reported in the literature. When DXA body composition measurements are combined with body water studies, stable bromide is often administered to measure extracellular water. Bromine attenuates x-rays significantly more than soft tissue and so could affect DXA body composition analysis. DXA scans were performed on 26 adults (12 F, 14 M) before and after the intravenous injection of 3 g sodium bromide (NaBr). No significant differences were noted pre- and post NaBr infusion for whole-body fat mass, fat-free soft tissue mass and bone mineral content. These findings were supported by a simple mathematical analysis of the likely effect of the sodium bromide infusion. This showed that when 3 g NaBr was introduced into the body, the effect on fat mass estimates was expected to be marginally less than the precision of the DXA technique. PMID- 11110253 TI - Computers in imaging and health care: now and in the future. AB - Early picture archiving and communication systems (PACS) were characterized by the use of very expensive hardware devices, cumbersome display stations, duplication of database content, lack of interfaces to other clinical information systems, and immaturity in their understanding of the folder manager concepts and workflow reengineering. They were implemented historically at large academic medical centers by biomedical engineers and imaging informaticists. PACS were nonstandard, home-grown projects with mixed clinical acceptance. However, they clearly showed the great potential for PACS and filmless medical imaging. Filmless radiology is a reality today. The advent of efficient softcopy display of images provides a means for dealing with the ever-increasing number of studies and number of images per study. Computer power has increased, and archival storage cost has decreased to the extent that the economics of PACS is justifiable with respect to film. Network bandwidths have increased to allow large studies of many megabytes to arrive at display stations within seconds of examination completion. PACS vendors have recognized the need for efficient workflow and have built systems with intelligence in the management of patient data. Close integration with the hospital information system (HIS)-radiology information system (RIS) is critical for system functionality. Successful implementation of PACS requires integration or interoperation with hospital and radiology information systems. Besides the economic advantages, secure rapid access to all clinical information on patients, including imaging studies, anytime and anywhere, enhances the quality of patient care, although it is difficult to quantify. Medical image management systems are maturing, providing access outside of the radiology department to images and clinical information throughout the hospital or the enterprise via the Internet. Small and medium sized community hospitals, private practices, and outpatient centers in rural areas will begin realizing the benefits of PACS already realized by the large tertiary care academic medical centers and research institutions. Hand-held devices and the Worldwide Web are going to change the way people communicate and do business. The impact on health care will be huge, including radiology. Computer-aided diagnosis, decision support tools, virtual imaging, and guidance systems will transform our practice as value-added applications utilizing the technologies pushed by PACS development efforts. Outcomes data and the electronic medical record (EMR) will drive our interactions with referring physicians and we expect the radiologist to become the informaticist, a new version of the medical management consultant. PMID- 11110254 TI - The reality of picture archiving and communication systems (PACS): a survey. AB - Toward the end of 1997 vendors were succeeding in installing picture, archiving and communication systems (PACS) in larger numbers. It was hard to separate fact from fiction at times. This survey was undertaken by 2 members of the academic community to confirm what was operational, how well the installed systems worked, and what they were doing. Fax questionnaires were sent to nearly 1,000 facilities worldwide to identify PACS of any size in clinical operation on a date certain, February 1, 1998. A total of 177 PACS were identified. Facilities furnished responses during the first survey period from May 1 to November 1, 1998. A second survey, done in February and March of 2000, sought long-term follow-up data. Many systems operated 5 or more types of modalities. Computed tomography (CT) was the most common modality type at 83%, but the distribution of the rest held surprises. There also was an unexpectedly large use of soft copy reading and filmless operation in 1998. Clear trends toward increased use of computed radiography and digital radiography and a significant expansion on the percentage of all of a facility's examinations on the PACS were seen over the 2 years. Satisfaction with original PACS vendors was relatively high. Eighty-nine percent remained with their original vendor. Only 10 sites reported they changed vendors, and 4 facilities said they abandoned their system. The users reported their expectations of the PACS had been met in 81% of cases. Some (65%) declared their systems were cost effective. The most striking response was that 97% of the users would recommend PACS to others. PMID- 11110255 TI - Iterative interim techniques for reduced costs and a better mammography workstation: an opinion. AB - The promise of digital mammography has focused considerable resources on the challenges of mammography workstations, but the risk of wasted time and resources in such efforts is very high. Although final testing of the workstation's image quality and ergonomics is common, a number of interim evaluation and refinement techniques can be applied throughout the design and development process. The use of such techniques holds potential not only to save time and money but also to produce a superior workstation. PMID- 11110256 TI - Significant savings in radiologic report turnaround time after implementation of a complete picture archiving and communication system (PACS). AB - One of the important advantages of the picture archiving and communication system (PACS) is the time saved in comparison with the conventional system. A group of 100 radiologic studies done in a conventional radiology department is compared with another group of the same number done in a completely filmless PACS department to assess the difference in the radiologist report turnaround time. There was a statistically significant (P < .00001) decrease in the median imaging to-dictation time (IDT) of the PACS group (3 hours and 40 minutes) in comparison with the pre-PACS group (25 hours and 19 minutes). This can be attributed to the fact that PACS eliminates all the workload associated with hard copy films, thus, improving the department's efficiency and decreasing the number of lost films. PMID- 11110257 TI - Wavelet compression on detection of brain lesions with magnetic resonance imaging. AB - The purpose of this report is to assess clinically acceptable compression ratios on the detection of brain lesions at magnetic resonance imaging (MRI). Four consecutive T2-weighted and the corresponding T1-weighted images obtained in 20 patients were studied for 109 anatomic sites including 50 with lesions and 59 without lesions. The images were obtained on a 1.5-T MR unit with a pixel size of 0.9 to 1.2 x 0.47 mm and a section thickness of 5 mm. The image data were compressed by wavelet-based algorithm at ratios of 20:1, 40:1, and 60:1. Three radiologists reviewed these images on an interactive workstation and rated the presence or absence of a lesion with a 50 point scale for each anatomic site. The authors also evaluated the influence of pixel size on the quality of image compression. At receiver operating characteristic (ROC) analysis, no statistically significant difference was detected at a compression ratio of 20:1. A significant difference was observed with 40:1 compressed images for one reader (P = .023), and with 60:1 for all readers (P = .001 to .012). A root mean squared error (RMSE) was higher in 0.94- x 0.94-mm pixel size images than in 0.94- x 0.47 mm pixel size images at any compression ratio, indicating compression tolerance is lower for the larger pixel size images. The RMSE, subjective image quality, and error images of 10:1 compressed 0.94- x 0.94-mm pixel size images were comparable with those of 20:1 compressed 0.94- x 0.47-mm pixel size images. Wavelet compression can be acceptable clinically at ratios as high as 20:1 for brain MR images when a pixel size at image acquisition is around 1.0 x 0.5 mm, and as high as 10:1 for those with a pixel size around 1.0 x 1.0 mm. PMID- 11110258 TI - Breast imaging using an amorphous silicon-based full-field digital mammographic system: stability of a clinical prototype. AB - An amorphous silicon-based full-breast imager for digital mammography was evaluated for detector stability over a period of 1 year. This imager uses a structured CsI:TI scintillator coupled to an amorphous silicon layer with a 100 micron pixel pitch and read out by special purpose electronics. The stability of the system was characterized using the following quantifiable metrics: conversion factor (mean number of electrons generated per incident x-ray), presampling modulation transfer function (MTF), detector linearity and sensitivity, detector signal-to-noise ratio (SNR), and American College of Radiology (ACR) accreditation phantom scores. Qualitative metrics such as flat field uniformity, geometric distortion, and Society of Motion Picture and Television Engineers (SMPTE) test pattern image quality were also used to study the stability of the system. Observations made over this 1-year period indicated that the maximum variation from the average of the measurements were less than 0.5% for conversion factor, 3% for presampling MTF over all spatial frequencies, 5% for signal response, linearity and sensitivity, 12% for SNR over seven locations for all 3 target-filter combinations, and 0% for ACR accreditation phantom scores. ACR mammographic accreditation phantom images indicated the ability to resolve 5 fibers, 4 speck groups, and 5 masses at a mean glandular dose of 1.23 mGy. The SMPTE pattern image quality test for the display monitors used for image viewing indicated ability to discern all contrast steps and ability to distinguish line pair images at the center and corners of the image. No bleeding effects were observed in the image. Flat field uniformity for all 3 target-filter combinations displayed no artifacts such as gridlines, bad detector rows or columns, horizontal or vertical streaks, or bad pixels. Wire mesh screen images indicated uniform resolution and no geometric distortion. PMID- 11110259 TI - Are both creatinine and urea clearances necessary as indices of small solute clearance adequacy in peritoneal dialysis? PMID- 11110260 TI - Postoperative adhesion formation and the use of adhesion preventing techniques in cardiac and general surgery. AB - The formation of postoperative adhesions is an inevitable sequel to surgical intervention, and, as part of the healing process, they are often beneficial. Nevertheless, the presence of adhesions may impose postoperative and reoperative surgical problems. An overview of some of the attempts to overcome such problems is presented, and the research surrounding them is discussed. PMID- 11110261 TI - Novel control system for blood glucose using a model predictive method. AB - We developed a novel blood glucose control system, using a model predictive method, to achieve optimal control of the blood glucose level in severely diabetic or pancreatectomized patients. This system is designed to predict glucose level changes in advance, considering delayed response time and the administered doses of insulin. This method is also designed to calculate the most appropriate insulin infusion rate by considering differences in individual response to insulin. In this study, we compared our system with a conventional proportional and differential controller (PD controller) to determine whether the new system could regulate the glucose level efficiently in pancreatectomized dogs. The model predictive control method resulted in a significant reduction of mean insulin infusion rate compared with the conventional PD controller (0.71 mU/kg per min vs. 1.81 mU/kg per min, p = 0.0005), when the glucose level in both methods reached the planned target level (100 mg/dl). The new system also tended to have a reduced mean glucose infusion rate for compensating for overshooting of the glucose level compared with the PD controller (0.7 mg/kg per min vs. 1.1 mg/kg per min, p = 0.16). These results indicate that the new system should be a useful tool for regulating the glucose level in severely diabetic patients. PMID- 11110262 TI - A prodrug approach for delivery of t-PA: construction of the cationic t-PA prodrug by a recombinant method and preliminary in vitro evaluation of the construct. AB - Previously, we reported a novel prodrug approach, that could lead to targeted thrombolysis without the risk of bleeding. The approach consists of a protein conjugate made of two components: a fibrin targeting antibody (Ab) linked to an anionic heparin, and a plasminogen activator (PA) derivatized with cationic species. These two components are linked by means of an electrostatic interaction. Because the cationic species are small, the modified PA would retain its thrombolytic activity. However, this activity would be inhibited after binding to the counterpart due to the blockage of the PA active site by the appended macromolecules. Because protamine is a clinical antagonist to heparin, it can be used in humans to dissociate the modified PA from its counterpart. Thus, the approach would permit the administration of a fibrin targeting but inactive thrombolytic drug (thereby alleviating the bleeding risk by avoiding systemic generation of plasmin), and subsequently a triggered release of the active drug to the fibrin deposit. In our previous work, we demonstrated the feasibility of the approach by producing a positively charged PA by means of chemical conjugation of a cationic CRRRRRRR peptide with urokinase. In this study, we further extended our work and produced a similar cationic t-PA by means of a recombinant DNA approach; i.e., by fusion of a poly(Arg)7 peptide to the kringle-1 domain of t-PA. Results obtained from the restriction enzyme analysis and the Western blot yielded full identification of this recombinant protein. This recombinant poly(Arg)7-modified-t-PA protein conjugate (termed "rmt-PA" hereafter) completely retained the fibrinolytic activity of the original recombinant, unmodified t-PA (termed "rt-PA" hereafter), as measured by the chromogenic assay and fibrin agar lysis assay. The prodrug and triggered release features of the proposed approach were confirmed by partial inhibition of the plasminogen activating activity of this protein by heparin, and the partial reversal of such inhibition by protamine. PMID- 11110263 TI - Simulated lipoprotein transport in the wall of branched arteries. AB - Study of arterial blood flow dynamics improves our understanding of the development of cardiovascular diseases such as atherosclerosis. The transport and accumulation of macromolecules in the arterial wall can be influenced by local fluid mechanics. We used numeric simulations to investigate such transport in a T junction model. Presumably an in vitro experiment would consist of gel segments inserted in the walls of a mechanical flow T-junction model near branch points where separation and recirculation zones are expected. The transport of low density lipoprotein (LDL) was investigated theoretically at these sites in a two dimensional numeric T-branch model. In the numeric model, the hydraulic conductivity of the porous gel wall segments was varied for a fixed species diffusivity to provide simulations with wall transmural Peclet numbers ranging from 0.3 to 30. Steady state flow patterns in the lumen of the two dimensional T branch were simulated at Reynolds numbers of 250 and 500, using the software package FIDAP 7.61 to implement the finite element method. The simulations demonstrated that wall Peclet numbers greater than 1.0 were needed to achieve species concentration gradients within the wall that varied in the axial direction, thereby reflecting the influence of disturbed flow and pressure patterns in the lumen. As expected, the transmural concentration gradients were steeper when convection predominated. Blood flow in the lumen can influence the distribution of macromolecules in the arterial wall and needs to be investigated for the relevance to atherosclerosis. PMID- 11110264 TI - A novel hemoglobin-adenosine-glutathione based blood substitute: evaluation of its effects on human blood ex vivo. AB - Chemically modified hemoglobin (Hb) solutions are under current investigation as potential red cell substitutes. Researchers at Texas Tech University have developed a novel free Hb based blood substitute product. This blood substitute is composed of purified bovine Hb cross-linked intramolecularly with o-adenosine 5'-triphosphate and intermolecularly with o-adenosine, and conjugated with reduced glutathione (GSH). In this study, we compared the effects of our novel blood substitute and unmodified (U) Hb, by using allogenic plasma as the control, on human blood components: red blood cells (RBCs), platelets, monocytes (Mo), and low-density lipoproteins (LDLs). The pro-oxidant potential of both Hb solutions on RBCs was examined by the measurement of osmotic and mechanical fragility, conjugated dienes (CD), lipid hydroperoxides (LOOH), thiobarbituric acid reactants (TBAR-S), isoprostanes (8-iso PGF2alpha) and intracellular GSH. The oxidative modification of LDLs was assessed by CD, LOOH, and TBAR-S, and the degree of apolipoprotein (apo) B cross-linking. The effects of Hb on platelets have been studied by monitoring their responses to the aggregation agonists: collagen, ADP, epinephrine, and arachidonic acid. Monocytes were cultured with Hb solutions or plasma and tested for TNF-alpha and IL-1beta release, then examined by electron microscopy. Results indicate that native UHb initiates oxidative stress of many blood components and aggravates inflammatory responses of Mo. It also caused an increase in RBC osmotic and mechanical fragility (p < 0.001). While the level of GSH was slightly changed, the lipid peroxidation of RBC increased (p < 0.001). UHb was found to be a stimulator of 8-iso PGF2alpha synthesis, a potent modulator of LDLs, and an effective potentiator of agonist induced platelet aggregation. Contrarily, our novel blood substitute did not seem to induce oxidative stress nor to increase Mo inflammatory reactions. The osmotic and mechanical fragility of RBCs was similar to that of the control. Such modified Hb failed to alter LDLs, increase the production of 8-iso PGF2alpha, but markedly inhibited platelet aggregation. The effect of this novel blood substitute can be linked with the cytoprotective and anti-inflammatory properties of adenosine, which is used as a cross-linker and surface modifier, and a modification procedure that lowers the hemoglobin pro-oxidant potential. PMID- 11110265 TI - Formation of occlusive platelet aggregates in whole blood caused by low concentrations of ADP. AB - Minute concentrations of ADP are released when platelets are exposed to shear stress during extracorporeal flow. However, based on current methods, these low concentrations have not been shown to have a significant impact on platelet function. We report here the formation of rigid microaggregates (MA) in response to low concentrations of ADP. A newly developed light scattering whole blood aggregometer (LSWBA) was used to detect an aggregation dose response to ADP (0-2 microM) in heparinized (1.5 U/ml) human blood. Although the LSWBA showed that ADP induced MA were reversible, evidence provided by constant pressure filtration (50 mm Hg) suggested that aggregates existed as rigid particles in the blood for up to 6 minutes. The possible implications of these findings to extracorporeal circulation are discussed. PMID- 11110266 TI - Effect of shear stress on platelet adhesion to expanded polytetrafluoroethylene, a silicone sheet, and an endothelial cell monolayer. AB - We visualized in real-time platelets adhering to the surface of three representative biomaterials, by using an apparatus consisting of a modified cone and plate rheometer combined with an upright epifluorescence microscope under two shear flows (0.1 and 5.0 dyne/cm2). The materials were expanded polytetrafluoroethylene (ePTFE), silicone sheet, and a monolayer of bovine endothelial cells (ECs) formed on glass, all of which are opaque materials used for artificial blood vessels and medical devices. According to quantitative analysis, the monolayer of ECs formed on glass had better blood compatibility than did either the ePTFE or the silicone sheet under shear flow conditions. Under a shear flow condition of 0.1 dyne/cm2, platelet adhesion was silicone sheet > ePTFE. In contrast, under a shear flow condition of 5.0 dyne/cm2, ePTFE > silicone sheet. These results indicate that the intensity of shear stress could modify the order of hemocompatibility of the materials. Therefore, direct observation of platelet adhesion under shear flow conditions is indispensable for testing and screening biomaterials and for providing a precise quantitative evaluation of platelet adhesion. PMID- 11110267 TI - Specific removal of anti-A and anti-B antibodies by using modified dialysis filters. AB - Removal of anti-A and anti-B blood group antibodies from human blood has been shown to facilitate cross-matched kidney transplantation by preventing hyperacute rejection. Patients in these studies had anti-A and anti-B antibodies removed by using plasmapheresis, followed by immunoadsorption onto packed bead columns. We conducted a study to assess the feasibility of selectively removing anti-A and anti-B antibodies directly from blood by using modified dialysis filters. An anti A and anti-B specific antigen was covalently attached to the lumenal surfaces of hollow fibers within selected commercial dialysis modules. The filters were able to reduce the anti-A and anti-B titers of 300 ml of blood to 2 or below. A low molecular weight fraction of our antigen system was found to have no antibody binding capacity. The standard antigen was purified by removal of the low molecular weight fraction and a dialysis filter was modified by using the purified antigen. This filter displayed a six-fold higher capacity than a dialysis filter modified with the same mass of standard antigen. We conclude that selective blood group antibody removal by antigen modified dialysis filters is feasible and may be a simpler system than plasmapheresis followed by immunoadsorption. PMID- 11110268 TI - Normothermic blood perfusion of isolated rabbit kidneys. II. In vitro evaluation of renal function followed by orthotopic transplantation. AB - This study describes the use of a blood perfusion apparatus to assess the renal function of isolated kidneys. Eight fresh kidneys were obtained from healthy rabbits and perfused with blood at 36 degrees C for 2 hours. Rabbit blood was drawn and diluted to a hematocrit of 25%. The kidneys were evaluated for their capacity to support life in an autograft model. Blood and urine samples were taken at regular time intervals during kidney perfusion. Serum creatinine was measured in surviving rabbits after transplantation. Over the course of the perfusion, arterial pressure was maintained at 87.2 +/- 5.5 mm Hg. The renal blood flow (3.7 +/- 1.0 ml/min per g) and urine output (0.11 +/- 0.04 ml/min per g) were continuously monitored. Glomerular filtration rate (0.29 +/- 0.02 ml/min per g) and fractional reabsorption (FR) of sodium and glucose indicated appreciable tubular function (FR(Na) = 67.9 +/- 8.5%, FR(Glu) = 91.2 +/- 5.8%). Protein was excluded from urine at 99.8% +/- 0.1%. After transplantation, the peak creatinine was 6.8 +/- 3.2 mg/dl at 1.90 +/- 0.92 days for the seven surviving rabbits and was above 16 mg/dl for the only rabbit that died 4 days after operation. The level of free hemoglobin generated at the end of the perfusion (2.6% +/- 2.8%) was correlated with the postoperative peak creatinine (r2 = 0.84). Perfusion of seven additional kidneys by using the roller pump lead to a final hemolysis of only 0.34 +/- 0.14%. Kidneys transplanted after 2 hours of blood perfusion were able to resume normal function and support life. Hemolysis was a measurable stress factor causing delayed function of the kidney after transplantation. Introduction of a roller pump significantly reduced the hemolysis. PMID- 11110269 TI - Nitric oxide added to the sweep gas infusion reduces local clotting formation in adult blood oxygenators. AB - Nitric oxide (NO) is an inhibitor of platelet aggregation. We analyzed the effect of direct infusion of NO into adult blood oxygenators on local clot formation. Nonheparinized calves in a control group (n = 3) and NO group (n = 4) were connected to a jugulocarotid cardiopulmonary bypass (CPB; centrifugal pump) for 6 hours. The venous line and pumphead were heparin coated, whereas the oxygenator, the heat exchanger, and the arterial line were not. A total of 80 ppm of NO was mixed with the sweep gas infusion in the NO group. The pressure gradient through the oxygenator (deltaP.Ox.) was monitored, and its evolution was compared between groups. Oxygenators membranes were analyzed and photographed, allowing for calculation of the percentage of surface area covered with clots by using a computer image analysis program. The deltaP.Ox. reached a plateau of 193 +/- 26% of the basal value in the NO group after 120 minutes, whereas a similar plateau of 202 +/- 22% was reached after only 20 minutes in the control group (p < 0.05). The surface area of the oxygenator covered with clots was significantly reduced in the NO group (0.54 +/- 0.41%) compared with the control group (5.78 +/ 3.80%, p < 0.05). However, general coagulation parameters were not modified by local NO administration. The activated coagulation time remained stable between 110 and 150 seconds in both groups (p = not significant [ns]), and there were no differences in hematocrit, thrombin time, partial thromboplastin time, or fibrinogen between groups during the 6 hours of CPB. Thus, the mixed infusion of a continuous low dose of NO into adult oxygenators during prolonged CPB prevented local clot formation, whereas the general coagulation pattern remained unchanged. PMID- 11110270 TI - Heparin bonding of the extracorporeal circuit reduces thrombosis during prolonged lung assist in goats. AB - This study investigated whether an artificial membrane lung of nonmicroporous polyolefin hollow fibers bonded with heparin could prolong venoarterial extracorporeal lung assist (ECLA) with low dose systemic heparin in goats. We compared heparin bonded circuits (Carmeda Bioactive Surface, "HB" group, n = 5) with non heparin bonded circuits ("NHB" group, n = 5) in venoarterial ECLA (V-A ECLA) for 7 days. Activated coagulation time (ACT) was maintained at approximately 130 sec by systemic infusion of small doses of heparin in the HB group, and at 200-230 sec in the NHB group. Thrombus formation was assessed by visual examination of the circuit, and possible cerebral embolization of thrombi was observed from behavioral abnormalities of the animals. The mean heparin dose given during ECLA was 20.4 +/- 3.6 U/kg per hr in HB, and 50.9 +/- 14.2 U/kg per hr in NHB, significantly less in HB than NHB (p < 0.01). Blood gas changes across the oxygenator, bypass flow rate, platelet aggregation activity, platelet counts, fibrin monomer (FM) test, and antithrombin-III (AT-III) activity did not differ between the two groups. In HB, thrombi were fewer and no abnormal neurologic symptoms were observed during ECLA. Numerous thrombi were observed in all oxygenators with NHB. One NHB goat developed convulsions and cerebral hemorrhage on the 6th day of ECLA. Nonmicroporous polyolefin hollow fibers can be bonded with heparin. An artificial membrane lung constructed of these fibers showed good anticoagulation by decreased thrombus formation with a small dose of infused heparin. PMID- 11110271 TI - Scaffold precoating with human autologous extracellular matrix for improved cell attachment in cardiovascular tissue engineering. AB - Cell attachment to a scaffold is a precondition for the development of bioengineered valves and vascular substitutes. This attachment is generally facilitated by the use of precoating factors, but some can cause toxic or immunologic side effects. Autologous extracellular matrix (ECM) is used as a precoating factor in our study. Ascending aortic tissue was cultured to obtain human myofibroblasts. Autologous ECM was extracted from the same aortic tissue. Poly(glycolic acid) (PGA) scaffolds were precoated with autologous ECM, human serum, or poly-L-lysine; the control group was pretreated with phosphate buffered saline (PBS). Myofibroblasts were seeded onto each scaffold, and the cell attachment was assayed and compared. Compared with the control group, precoating with human serum, poly-L-lysine, and ECM increased number of attached cells by 24%, 53%, and 48%, respectively. Differences between precoating groups were significant (p < 0.01), except for ECM versus poly-L-lysine. Scanning electron microscopy also demonstrated the high degree of cell attachment to the PGA fibers on scaffolds precoated with ECM and poly-L-lysine. Precoating polymeric scaffold with autologous human extracellular matrix is a very effective method of improving cell attachment in cardiovascular tissue engineering without the potential risk of immunologic reactions. PMID- 11110272 TI - Intrathoracic esophageal replacement with a collagen sponge--silicone double layer tube: evaluation of omental-pedicle wrapping and prolonged placement of an inner stent. AB - In a previous study, we replaced a 5 cm gap created in the canine intrathoracic esophagus with an artificial esophagus. However, although newly formed esophageal tissue subsequently bridged the gap, mild stenosis occurred, and this seemed to be caused by inadequate regeneration of the skeletal muscle. In the present study, we evaluated whether omental pedicle wrapping (OMPx) of the prosthesis could promote tissue regeneration and whether prolonged retention of the silicone tube within the prosthesis could prevent stenosis. A gap was created in 14 dogs, and the defect was repaired by our prosthesis. OMPx was performed in 5 of the 14 dogs (OMPx group) but not in the rest (control group). The silicone tube was retained for 4 weeks in the control group and for 8 weeks in the OMPx group. All of the dogs in the control group survived for more than 3 months, except for those that were killed. Four dogs in the OMPx group died within 3 months, one caused by perforation at 7 months. Only the thin epithelial and submucosal layer regenerated in the OMPx group. OMPx is not effective for promoting tissue regeneration, and prolonged retention of the silicone tube interrupts epithelial regeneration. PMID- 11110273 TI - Cytokine mediated endothelial activation during and after normothermic cardiopulmonary bypass: heparin-bonded versus non heparin-bonded circuits. AB - Studies evaluating cytokine production under normothermic cardiopulmonary bypass (CPB) are limited. We evaluated cytokine production, levels of thrombomodulin (TM), and soluble endothelium-derived adhesion molecules (ICAM-1) under normothermic CPB with and without heparin-bonded circuits. Nine patients treated with non heparin-bonded circuits (control group), and seven patients treated with heparin-bonded circuits (heparin group) were the subjects. Granulocyte elastase (G-E), and interleukin (IL) -6 and IL-8 were chosen as proinflammatory mediators, and TM and ICAM-1 served as indicators for endothelial damage. Blood samples were obtained before CPB, 30 minutes after initiation of CPB, at the termination of CPB, and 2 and 24 hours after CPB. G-E values in the heparin group were lower than those in the control group after 30 minutes of CPB. A G-E surge occurred at the end of CPB, and IL-6 and IL-8 surges were observed 2 hours after CPB in both groups. TM and ICAM-1 values, which were reduced at the initiation of CPB, returned to initial levels 2 hours after CPB, and exceeded them 24 hours after CPB compared with preCPB levels. Both groups showed similar changes. We conclude that there are no significant differences in serial G-E, IL-6, IL-8, TM, or ICAM 1 levels between the heparin and control groups during or after normothermic CPB for 2 to 3 hours. PMID- 11110274 TI - Ovalis TAH: development and in vitro testing of a new electromechanical energy converter for a total artificial heart. AB - A new electromechanical energy converting system has been developed to yield an efficient and durable orthotopic total artificial heart (TAH). The energy converter we developed transforms the unidirectional rotational motion of the motor into a longitudinal forward-reverse movement of an internal geared oval, linked directly to pusher plates on both sides. To ensure a permanent positive connection between the drive gear and the internally geared wheel, a ball bearing runs inside an oval shaped guide track. Motor, gear unit, and conical pusher plates are seated between alternately ejecting and filling ventricles. The unidirectional motion of the brushless DC motor affords easier motor control, reduces energy demand, and ensures longer life of the motor when compared with a bidirectional motion system. In vitro testing has been performed on a mock circulation loop. The overall system efficiency of the TAH Ovalis was 27-39% (mean, 36%) for the pump output range of 2-7 L/min. The maximum output of 7 L/min can be obtained with a pump rate of 130 min(-1) and an afterload pressure of 140 mm Hg. For an average sized human with a mean cardiac output of 6 L/min at a mean aortic pressure of 120 mm Hg, 5 watts of input power would be required. The size of the prototype is 560 cm3, the weight is 950 g. Our first in vitro studies demonstrated the excellent efficiency and pump performance of this new electromechanical energy converter. The results prove the feasibility of this new concept's use as an energy converter for a total artificial heart. PMID- 11110275 TI - Analysis of the interventricular pressure waveform in the moving-actuator total artificial heart. AB - Right and left filling pressures are important parameters in the automatic control of a total artificial heart (TAH) within normal physiologic ranges. Our TAH is composed of a moving actuator, right and left ventricles, and an interventricular space (IVS) enclosed by a semirigid housing. During operation of the TAH, the IVS volume is changed dynamically by the difference between the ejection volume of one ventricle and the inflow volume of the other. We measured the interventricular pressure (IVP) waveform by using a pressure sensor and analyzed the relationship between the IVP and the preload condition. From in vitro and in vivo experiments, we found that the measured filling pressures were linearly related to the negative peak value of the IVP. Additionally, we found that we could use the time interval from actuator start to the positive peak value of the IVP (outflow valve opening) as a useful parameter to estimate the blood filling volume of the diastole ventricle. PMID- 11110276 TI - Assessment of synchronized direct mechanical ventricular actuation in a canine model of left ventricular dysfunction. AB - Direct mechanical ventricular actuation (DMVA) is an experimental procedure that provides biventricular cardiac assistance by intracorporeal pneumatic compression of the heart. The advantages this technique has over other assist devices are biventricular assistance, no direct blood contact, pulsatile blood flow, and rapid, less complicated application. Prior studies of nonsynchronized DMVA support have demonstrated that a subject can be maintained for up to 7 days. The purpose of this study was to determine the acute hemodynamic effects of cardiac synchronized, partial DMVA support in a canine model (RVP) of left ventricular (LV) dysfunction. The study consisted of rapidly pacing seven dogs for 4 weeks to create LV dysfunction. At the conclusion of the pacing period, the DMVA device was positioned around the heart by means of a median sternotomy. The animals were then imaged in a 1.5 T whole body high speed clinical MR system, with simultaneous LV pressure recording. Left ventricular pressure-volume (PV) loops of the nonassisted and DMVA assisted heart were generated and demonstrated that DMVA assist shifted the loops leftward. In addition, assist significantly improved pressure dependent LV systolic parameters (left ventricular peak pressure and dp/dt max, p < 0.05), with no diastolic impairment. This study demonstrates that DMVA can provide synchronized partial assist, resulting in a decrease in the workload of the native heart, thus having a potential application for heart failure patients. PMID- 11110277 TI - Mixed venous oxygen saturation as a promising parameter for physiologic control of total artificial heart. AB - Mixed venous oxygen saturation (SvO2) has been proposed as one of the suitable parameters for physiologic control of a total artificial heart (TAH). To establish the practical application of SvO2, we investigated the response of cardiac output (CO) and SvO2 to step-loaded exercise. A normal calf was surgically equipped with an ultrasonic flowmeter probe and an oximetry catheter in the pulmonary artery to measure CO and SvO2, respectively. Three stage step treadmill exercise tests (1, 2, and 4 km/h) were performed three times. While CO increased from 8.9 L/min at preexercise level to 9.7, 10.2 and 11.4 L/min at 1, 2, and 4 km/h, respectively, SvO2 decreased from 59.6% to 56.8, 55.3, and 52.2%, respectively. There existed a linear correlation between the magnitude of changes in CO and SvO2. CO and SvO2 exhibited a similar course of change, expressing an inverted exponential curve. The time constant of SvO2 was from 19 to 35 seconds, whereas that of CO was from 21 to 39 seconds. We conclude that SvO2 changes in close association with CO during exercise and has good potential to be a parameter for physiologic control of a TAH, by reflecting the recipient's CO demand without conspicuous time delay. PMID- 11110278 TI - Adjunctive antibiotic/anticoagulant lock therapy in the treatment of bacteremia associated with the use of a subcutaneously implanted hemodialysis access device. AB - To improve vascular access for hemodialysis, a new device (Dialock Hemodialysis Access System, Biolink Corporation, Middleboro, MA) has been developed. Implanted subcutaneously, the device is accessed by percutaneous puncture. Attached to the device are two catheters that are implanted into the superior vena cava or right atrium. Clinical results thus far have been promising. However, use of this device is not free from infectious complications. In the present pilot study, 25 maintenance hemodialysis patients were implanted with 26 Dialock devices. The incidence of bacteremia was 2.9/1,000 catheter days. In 14 episodes of bacteremia in 8 patients the infection was successfully treated with a combination of systemic antibiotic treatment and adjunctive antibiotic/anticoagulant lock therapy. The lock therapy entailed the instillation of both an antibiotic and an anticoagulant into the device. We believe that the antibiotic/anticoagulant lock technique is an effective, adjunctive therapeutic modality in the treatment of infections related to the use of indwelling vascular access devices. PMID- 11110279 TI - Tunneled hemodialysis catheter survival: comparison of radiologic and surgical implantation. AB - Cuffed, tunneled hemodialysis catheters (caths) are often implanted in the operating rooms (OR) by surgeons or by interventional radiologists in radiology suites (RS). Comparative outcome studies between OR and RS placed caths are few and tend to favor the specialty of the authors. In this longitudinal study, we monitored cath survival in patients while awaiting maturation of their fistulae, and compared outcomes between OR and RS placement. A total of 95 caths were placed in 50 patients between July 1996 and July 1999. Radiologically placed caths had a shorter primary patency duration than OR placed caths (80 +/- 40 days vs. 100 +/- 31 days, p = 0.04) and a lower primary patency rate at 120 days than OR placed caths (42% vs. 67%, p = 0.04). Cumulative infection rate per 1,000 catheter days was higher in RS than OR cases (3.8 Vs 2.2, p = 0.09), whereas mean sepsis free duration was shorter in RS than OR (60 +/- 45 days vs. 88 +/- 40 days, p = 0.02). The risk of infection was 1.7 times greater in RS than OR cases (chi-square = 6.4, p = 0.01). The RS placed caths also had a higher rate of primary nonfunction (31% vs. 8.3%, p = 0.04) and bleeding complications (42% vs. 17%, p = 0.04), but significantly shorter procedure scheduling time than OR cases (1.1 +/- 0.3 days vs. 2.5 +/- 0.6 days, p < 0.0001). In conclusion, radiologically placed caths seem to have higher rates of infection, bleeding, and functional failure but shorter scheduling time than surgically placed caths. Discussions are under way to improve the survival of RS placed caths at our affiliated hospitals. PMID- 11110280 TI - Quality of care differences by ownership in United States renal dialysis facilities. AB - Recent studies of patient outcomes in the United States have shown mixed conclusions regarding the effect of for-profit ownership on treatment quality. To test whether outcome quality differs across ownership types for patients being treated for end stage renal disease, a cross-sectional study was performed using data from 180,913 end stage renal disease patients receiving center based hemodialysis in 1996. Results indicated that patients in for-profit renal dialysis facilities had slightly higher mortality during the study period than patients in not-for-profit facilities, after controlling for patient case mix and market type. For profit ownership seems to affect not only treatment inputs, according to previous research, but also patient outcomes, as indicated by the results here. PMID- 11110281 TI - Testing of a 50 cc stroke volume completely implantable artificial heart: expanding chronic mechanical circulatory support to women, adolescents, and small stature men. AB - The development of a completely implanted total artificial heart at our institution has progressed to successful in vivo and in vitro testing of a device that is nearing clinical testing. This system consists of a 70 cc stroke volume pump originally designed to be used in men of average stature. Implantation of this system remains limited by patient size; hence, many women and adolescent patients will likely be precluded from support because of their smaller stature. A system similar in design, but with a 50 cc stroke volume pump has been developed. The first in vivo study of this device has been undertaken. A calf was supported for 33 days. The animal was extubated and ambulatory within the first 6 hours of implantation, and remained healthy until the thirty-third postoperative day when it suffered an embolic neurologic event. The pump and operating system worked flawlessly throughout the period of support. Further in vivo and in vitro testing will be undertaken. Development of a scaled down total artificial heart system expands this type of circulatory support to those critically ill patients previously deemed poor candidates because of their smaller body habitus. PMID- 11110282 TI - Extracorporeal adsorption as a new approach to treatment of botulism. AB - Botulism is a paralytic disease caused by a toxin produced by the bacterium Clostridium botulinum. Outbreaks of the illness take place with a mortality rate of 10%, and the potential terrorist use of the toxin has become a serious concern. The current treatment includes administration of antitoxin, which can cause serious allergic reactions. Recently, we have successfully treated a 64 year old woman with the illness with IMMUSORBA TR350 (Asahi Medical, Tokyo, Japan), an extracorporeal adsorptive column containing polyvinylalcohol tryptophan as an adsorptive agent, which has been widely used in Japan to treat myasthenia gravis and Guillain-Barre syndrome. Initially, the patient developed ocular muscle weakness and a variant of the Guillain-Barre syndrome was suspected. After extracorporeal treatment, her neurologic symptoms remarkably improved. After a series of treatments, botulinum toxin type B was isolated in the food she had eaten, establishing the diagnosis. An in vitro study revealed that the adsorptive column removed botulinum toxin to a significant extent. Our recent findings suggest that treatment with the adsorptive column TR350 can be a feasible option for botulism, which is a rare but potentially lethal disease. PMID- 11110284 TI - Long-term anticoagulation with recombinant hirudin in a patient on left ventricular assist device support. AB - We report on a 24 year old man with a Novacor left ventricular assist device (LVAD) who underwent long-term treatment with intravenous recombinant hirudin due to a heparin induced thrombocytopenia (HIT II) after suffering from an ischemic stroke. PMID- 11110283 TI - Effects of transmyocardial laser revascularization by using a prototype pulsed CO2 laser on contractility and perfusion of chronically ischemic myocardium in a porcine model. AB - The purpose of this study was to test a new prototype pulsed CO2 laser to be used for transmyocardial laser revascularization (TMR). We wanted to determine whether it can reduce thermal damage and mitigate induced ischemia with improvement in contractile reserve of the heart as evidenced by contrast echocardiography at rest and under dobutamine stress. TMR is an emerging surgical strategy for treatment of myocardial ischemia not amenable to conventional percutaneous or surgical revascularization. Eleven pigs underwent ameroid occluder placement at the origin of the circumflex coronary artery. Six weeks later, occlusion of the circumflex coronary artery was documented. TMR was then carried out on 10 pigs by using a prototype pulsed CO2 laser that delivered 8-12 joules in 1.5 ms with a spot size of 1 mm. Six weeks after TMR, the pigs were restudied. The animals developed significant ischemia after 6 weeks of ameroid occlusion, at rest (p = 0.01) and at peak stress (p = 0.004). Wall motion for the ischemic segments improved significantly 6 weeks after TMR at peak stress (p = 0.02). TMR results in an improvement in wall motion in our model of chronic ischemia and improves wall motion score index more during induced stress than at rest. PMID- 11110285 TI - Medical device associated infections. PMID- 11110286 TI - Physiology of wound healing and surgical wound care. AB - Wound healing is a systemic process, which occurs stepwise and involves the stages of hemostasis, inflammation, and repair. Hemostasis with fibrin formation creates a protective wound scab. The scab provides a surface beneath which cell migration and movement of the wound edges can occur. Inflammation brings nutrients to the area of the wound, removes debris and bacteria, and provides chemical stimuli for wound repair. Repair begins immediately after wounding and proceeds rapidly through the processes of epithelialization, fibroplasia, and capillary proliferation into the healing area. Different tissues have their own normal rates of growth during the process of healing. The optimal rate of healing is approached when factors advantageous to healing are present and factors having the ability to disturb or retard the healing processes are controlled or absent. These factors are discussed. PMID- 11110287 TI - Dialysis access related infections. AB - Infection is a common cause of morbidity and mortality in end-stage renal disease patients. Unintentional pathogens are introduced into an immunocompromised host during hemodialysis and peritoneal dialysis by means of the access (arteriovenous fistula, arteriovenous graft, central venous catheter, or peritoneal dialysis catheter). Gram positive organisms are most common with Staphylococcus aureus and coagulase negative Staphylococcus predominating. Preventive measures are mandatory and the key to decreasing infection rates. PMID- 11110288 TI - Staphylococcus aureus infections in dialysis patients: focus on prevention. AB - Staphylococcus aureus infections are a major cause of morbidity and hospitalization in dialysis patients. The risk of infection relates to the type of access. Patients with acute hemodialysis (HD) catheters are at the greatest risk of S. aureus bacteremia, followed by tunneled HD catheters, and grafts. Patients with a fistula have a rate similar to that of peritoneal (PD) patients. In PD patients, however, S. aureus is the second most common cause of peritonitis, is often associated with a catheter infection, and frequently requires catheter removal for resolution. S. aureus infections in dialysis patients are much more common in nasal carriers. S. aureus moves from the nasal reservoir to the hands and skin, and from there to infect the access. Therefore, prevention of infection can be aimed at treating the carriage or in applying antibiotics at the catheter exit site, thus preventing colonization and subsequent infection of the catheter. For HD patients with a permanent access (either fistula or graft), intranasal mupirocin, twice a day for 5 days followed by a once weekly application, is effective in reducing the risk of S. aureus bacteremia. Cost analysis indicates that treating all patients would result in more cost savings than treating just carriers. For patients with acute HD catheters, exit site mupirocin applied as part of routine care during each HD treatment, reduces the risk of S. aureus exit site infection and bacteremia. For PD patients, S. aureus infections can be diminished by using mupirocin at the exit site as part of daily exit site care. Prophylaxis against S. aureus is under utilized in dialysis patients and, if implemented, could lower the rate of these serious infections. PMID- 11110289 TI - Polytetrafluoroethylene hemoaccess site infections. AB - The advent of permanent hemodialysis access has made possible the use of chronic hemodialysis in patients with end-stage renal disease. Although autogenous arteriovenous fistulae remain the conduit of choice, their construction is not always feasible. Prosthetic grafts made of polytetrafluoroethylene (PTFE) are typically the second-line choice for hemoaccess. However, these grafts suffer from decreased rates of patency and an increased number of complications. Although thrombosis is the most common complication, infection of PTFE grafts accounts for a significant number of hospitalizations and uses a large amount of healthcare resources. In this monograph, we address infectious complications of PTFE hemoaccess grafts and present a review of the recent literature. PMID- 11110290 TI - Infection and thrombosis in total artificial heart technology: past and future challenges--a historical review. AB - On the basis of animal testing and a single clinical implant during the 1960s, development of the total artificial heart (TAH) began in earnest in the 1970s. The goal was to produce a pump that could treat biventricular heart failure or any other condition that necessitated removal of the patient's native heart. The early TAHs were pneumatically powered, with externalized drivelines. After undergoing in vivo evaluation in hundreds of sheep and calves at several centers (mainly the Utah Heart Institute), these pumps were implanted in humans, initially for permanent cardiac replacement and later for bridging to transplantation. In both the in vivo experimental setting and the clinical setting, infection and thrombosis were problematic, infection being encountered much more frequently than thrombosis in clinical cases. To minimize these problems, four research groups, funded by NIH, began in 1988 to develop permanent, transcutaneously powered, totally implantable, electromechanical TAHs. For the first time, TAH technology was able to minimize infection and thrombosis, as confirmed by current in vivo studies. These new TAHs will undergo preclinical, pre-IDE studies this year and clinical trials in the near future. This article briefly reviews the evolution of TAH technology, with an emphasis on the prevention and management of infection and thrombosis. PMID- 11110291 TI - Infectious complications associated with ventricular assist systems. AB - Infectious complications during support with a ventricular assist system (VAS) can cause severe morbidity and mortality, affecting nearly one-half of all VAS recipients. Because of the lack of a uniform definition of infection, the incidence of this complication is hard to determine accurately. It is approximately 50% for patients being supported by an implantable VAS as a bridge to heart transplantation and 28% for patients supported by an external, short term VAS. Infections can be classified according to the involvement or noninvolvement of the implanted device and according to the severity of the infection. Severe infections involving the implanted device may preclude heart transplantation for some patients, but numerous patients with milder infections have undergone successful transplantation. Numerous factors predispose VAS patients to infection. Postoperative bleeding necessitating re-operation is an important contributing factor. Endotracheal tubes, intravascular catheters, and other indwelling tubes necessary for the care of postsurgical patients are also common routes of contamination. Control of infection may be improved with new VAS designs, antibiotic impregnated drivelines, and innovative therapies such as antibiotic beads. The next generation of VASs should be inherently less susceptible to infection because of their smaller size, reduced thrombogenicity, and better flow characteristics. In addition to more effective antibiotics, improved VAS designs that incorporate transcutaneous energy transmission systems may reduce infectious complications and allow safe, long-term VAS support. PMID- 11110292 TI - Fungal infections in ventricular assist devices. AB - Device infections in patients supported by a mechanical circulatory support system remain an important problem, particularly as we enter the era of permanent device implantation. This article focuses on fungal infections that occur in patients with ventricular assist devices. The nature of fungal, especially Candida species, colonization and infection in severely ill, hospitalized patients will be described. Information regarding the effect of the artificial surface-blood interface on the immune system's ability to combat fungal organisms will also be presented. Basic aspects of the fungal-host interaction serve as the foundation for a discussion of clinical management protocols for preventing and treating fungal infections in patients supported by a ventricular assist device. PMID- 11110293 TI - Infection in patients after implantation of an orthopedic device. AB - During the last several decades, the use of appropriate antibiotics has significantly improved our ability to prevent and treat infection that occurs after implantation of an orthopedic device. Despite improved prevention and treatment of this condition, patients who develop an infection secondary to implantation of an orthopedic device face increased mortality, morbidity, and/or delayed recovery. The presence of an orthopedic device significantly reduces the number of bacteria required to produce colonization and decreases the ability of the body's own defense mechanism and antibiotics to resolve this condition. Efforts devoted to prevention of infection are much more effective than those spent treating the condition once it has developed. Pretreatment of patients with antibiotics and the use of ultra clean surgical rooms have been shown effective. Prevention will become increasingly important as antibiotic resistant strains of bacteria become more prevalent and the number of arthroplasty procedures performed also increases. PMID- 11110294 TI - Role of biofilms in antimicrobial resistance. AB - Biofilms are formed by a spectrum of microorganisms, including pathogens, and provide a means for these organisms to protect themselves against antimicrobial agents. Several mechanisms have been proposed to explain this phenomenon of resistance within biofilms, including delayed penetration of the antimicrobial into the biofilm extracellular matrix, slowing of growth rate of organisms within the biofilm, or other physiologic changes brought about by interaction of the organisms with a surface. The practical implications of biofilm formation are that alternative control strategies must be devised both for testing the susceptibility of the organisms within the biofilm and treating the established biofilm to alter its structure. A number of testing protocols have been developed. Effective treatment strategies will incorporate antimicrobials or other agents that have been demonstrated to penetrate and kill biofilm organisms, or treatments that disrupt or target specific components of the biofilm matrix. A better understanding of the role of biofilms in infection and how in vivo biofilms respond to selected treatments requires more study. PMID- 11110295 TI - Activation and control of complement, inflammation, and infection associated with the use of biomedical polymers. AB - It is generally acknowledged that artificial biomaterials are much less immunologically active than transplants or tissue derived biomaterials. However, activation of both the coagulation cascade and the complement system is a common occurrence when human blood is exposed to biomaterial surfaces during extracorporeal procedures, such as renal hemodialysis or cardiopulmonary bypass. Both individual and collective activation of these cascades often produce local and systemic effects. A number of complement activation products function as the mediators of inflammation. They serve as ligands for specific receptors on polymorphonuclear leukocytes, monocytes, macrophages, mast cells, and other cells. Such an interaction leads to induction of cellular responses in adhered cells, including release of oxidative products, lysosomal enzymes, or both, which often contribute to a number of pathologic conditions. Most pathogens invading the human body are attacked by the immune system directly following entry, especially when they are in contact with blood. However, bacteria and parasites have developed a large number of specific strategies to overcome immune defense among others by avoiding either recognition or eradication by complement. In this aspect, of concern are several microorganisms responsible for formation of antibiotic resistant biofilms on biomaterial surfaces, namely Staphylococcus epidermidis, Staphylococcus aureus, and Pseudomonas aeruginosa. PMID- 11110296 TI - Enhanced risk of infection with device-associated thrombi. AB - Thrombosis and infection are two well-recognized risks with prosthetic devices that contact blood. Many of the currently used biomaterials may present an attractive surface for thrombus development as well as bacterial adhesion and colonization. Clinical experience with vegetative endocarditis patients has suggested that thrombosis may lead to enhanced risk of infection, and the possibility that adherent bacteria may enhance the risk of thrombosis has been noted by several investigators. To investigate the correlation between thrombosis and infection, a series of tests were conducted to assess the affinity of pathogen with surfaces in the absence and presence of blood components. Coronary stents were used as a model device to attract thrombi in a recirculating loop in vitro. Fresh heparinized blood was used to investigate thrombus development and bacterial interaction. (111)Indium-labeled Staphylococcus epidermidis and (111)Indium-labeled platelets were utilized to quantify bacterial interaction with thrombi under various test conditions. Anticoagulants, antiplatelet agents, and antibiotics were utilized in attempts to selectively influence bacteria, platelets, or thrombosis. The results suggest that under appropriate conditions, bacteria may preferentially adhere to platelet rich thrombus. These observations also suggest that by reducing the risk of thrombosis, the risk of device associated infection may also be reduced. PMID- 11110297 TI - Prion disease and medical devices. AB - Prions are novel proteinaceous-infectious agents that have been implicated in transmissible spongiform encephalopathies. It is now generally accepted that noninfectious prion proteins are normally produced by the host and may undergo a conformational change to an abnormal, pathologic form, which appears to be responsible for disease symptoms. Many methods of decontamination and sterilization are claimed to be ineffective against prion proteins. Incidences of iatrogenic transmission of prions due to medical devices have been reported, and the recommended clinical practices when handling suspected cases are reviewed. Recent results with a peracetic acid based sterilant indicate that it may be a safe and effective means of prion inactivation on medical devices. PMID- 11110298 TI - Bovine spongiform encephalopathy: an overview. AB - Bovine spongiform encephalopathy (BSE), widely known as "mad cow disease," is a chronic, degenerative disease affecting the central nervous system of cattle. Worldwide, there have been more than 180,000 cases since the disease was first diagnosed in 1986 in Great Britain. Bovine spongiform encephalopathy has had a substantial impact on the livestock industry in the United Kingdom. The disease has also been confirmed in native-born cattle in Belgium, Denmark, France, Ireland, Luxembourg, Liechtenstein, The Netherlands, Northern Ireland, Portugal, and Switzerland. However, over 95% of all BSE cases have occurred in the United Kingdom. Bovine spongiform encephalopathy is not known to exist in the United States. PMID- 11110299 TI - Can infections be imaged in implanted devices? AB - Approximately 30,000 to 60,000 patients per year in the United States are candidates for heart transplants, mechanical assist devices, or both. These procedures, devices, and the associated short- and long-term care required are expensive and command significant utilization of health care resources. Because device related infection and thrombosis are potentially devastating complications of implanted device utilization, early diagnosis of infection, thrombosis, or both, would be helpful for initiation of early therapy to prevent untoward clinical events. Therefore, the development of a robust imaging technology for identification of infection could be cost effective if early assessment, diagnosis, and therapy of infected devices led to improvements in morbidity and mortality. PMID- 11110300 TI - Hemodynamic effect of iodinated high-viscosity contrast medium in the rat kidney: a diffusion-weighted MRI feasibility study. AB - RATIONALE AND OBJECTIVES: To assess the abilities of dynamic diffusion-weighted MRI to demonstrate the effects in vivo of a high-viscosity iodinated contrast agent on medullary and cortical blood flow in the rat kidney. METHODS: Dynamic diffusion-weighted, echoplanar MR images obtained from five b-value single-shot acquisitions and their isotropic apparent diffusion coefficient maps were obtained from nine rats anesthetized by pentobarbital sedation, before and after intravenous injection of a high-viscosity, dimeric iso-osmolar iodinated contrast medium (iodixanol), and compared with those obtained from four control rats that received saline. RESULTS: The mean baseline apparent diffusion coefficient values were 1.64 +/- 0.05 x 10(-3) mm2/s for the cortex and 1.75 +/- 0.06 x 10(-3) mm2/s for the medulla. In the iodixanol group, a significant decrease in renal diffusion was observed at 12 minutes and lasted at least until 24 minutes. The decrease in diffusion occurred earlier for the cortex and lasted less than for the medulla. There was no significant modification in diffusion over time in the control group. CONCLUSIONS: This preliminary experience in rats shows that dynamic diffusion-weighted MRI can be used to study noninvasively the in vivo renal hemodynamic response after injection of iodinated contrast. PMID- 11110301 TI - Arteriography and portal venography on routine follow-up after orthotopic liver transplantation. AB - RATIONALE AND OBJECTIVES: To describe the findings of routinely performed angiographic examinations in patients at discharge 2 months after orthotopic liver transplantation (OLT) and at follow-up 1 year later. METHODS: The findings of 315 angiographic examinations performed in 190 patients 2 months and 1 year after OLT were reviewed, and the changes at the anastomotic site of the hepatic artery and portal vein were analyzed. RESULTS: Routine angiography 2 months and 1 year after OLT demonstrated a normal anastomosis or low-grade stenosis in 82% and 84% of the patients (hepatic artery) and in 88% and 84% (portal vein), respectively. High-grade stenosis occurred in 9% and 5% of the patients (hepatic artery) and in 3% and 5% (portal vein). Hepatic artery occlusion and portal vein occlusion were observed in two and seven patients and in one and three patients, respectively. In 76% of patients, the anastomotic site of the hepatic artery did not change significantly. In eight patients, a normal anastomosis or a low- or medium-grade stenosis developed into high-grade stenosis or occlusion. Conversely, in nine patients, medium- or high-grade stenosis developed into a normal anastomosis or a low-grade stenosis. In all eight patients who initially had a high-grade stenosis, the hepatic artery proved to be patent at 1 year. In 98% of patients, the anastomotic site of the portal vein did not change significantly. In one patient who initially had a normal anastomosis, occlusion was found at I year. CONCLUSIONS: In most patients, routine angiography 2 months and 1 year after OLT demonstrated normal findings or a low-grade stenotic anastomosis of the hepatic artery and portal vein. Significant changes occurred mainly at the anastomotic site of the hepatic artery and could not be predicted by previous angiograms. PMID- 11110302 TI - Influence of bubble size distribution on the echogenicity of ultrasound contrast agents: a study of SonoVue. AB - RATIONALE AND OBJECTIVES: To study the relative contributions of different bubble size classes to SonoVue's echogenicity in fundamental acoustic imaging modes. SonoVue is a contrast agent, previously known as BR1, with a bubble size distribution extending from approximately 0.7 to 10 microm. METHODS: A model for the acoustic response of SonoVue was determined and validated for a set of experimental data. This model was used to simulate the acoustic response of a standard batch of SonoVue as the sum of responses of non-overlapping bubble size classes. RESULTS: The simulation was first validated for a standard SonoVue bubble size distribution. When this distribution was considered as five size classes with equal numbers of bubbles, it was found that bubbles smaller than 2 microm accounted for 60% of the total number but contained only 5% of the total gas volume. The simulation results indicated marked differences in the acoustic contributions from these classes, with 80% of the acoustic efficacy provided by bubbles 3 to 9 microm in diameter. The study also compared bubble distributions in number, surface, and volume, with the distribution computed in terms of acoustic efficacy. CONCLUSIONS: This study shows why bubble volume is a much better indicator of SonoVue's efficacy than is bubble count. A low threshold in diameter was found for SonoVue microbubbles at approximately 2 microm, under which size bubbles do not contribute appreciably to the echogenicity at medical ultrasound frequencies. PMID- 11110303 TI - A new type of artificial urinary calculi: in vitro study by spiral CT. AB - RATIONALE AND OBJECTIVES: Artificial urinary calculi similar to natural stones have long been sought in urologic research. In an experimental study, the authors assessed the CT characteristics of a new type of artificially produced urinary calculus [BON(N)-STONE]. METHODS: Six different types of urinary calculi (uric acid, struvite, cystine, calcium oxalate, brushite, and apatite) were produced by a coating technique in which several layers of a suspension of pure substance were applied around a core and dried. A total of 60 stones (10 per group) were studied by spiral CT at two energy levels (100 and 120 kV, 250 mA) with 1-mm slice thickness. RESULTS: All calculi showed a small hyperdense core surrounded by a homogeneous matrix and a slightly hyperdense outer rim. From the least to the most dense, the stone types were uric acid, struvite, cystine, calcium oxalate, brushite, and apatite. Absolute CT values at 100 and 120 kV could differentiate between all groups of stones at a significance level of P < 0.001 or better. Attenuation values were in a comparable range to reported values for natural stones, with the exception of uric acid and struvite, which were notably lower. CONCLUSION: These artificially produced urinary calculi showed properties similar to those of natural stones. Thus, this seems to be a promising stone model for further investigations. PMID- 11110304 TI - Motor cortex brain activity induced by 1-Hz transcranial magnetic stimulation is similar in location and level to that for volitional movement. AB - RATIONALE AND OBJECTIVES: The relatively high temporal and spatial resolution of functional MR imaging was used to compare the blood oxygenation level dependent (BOLD) response associated with movement induced by transcranial magnetic stimulation (TMS) with that for a similar movement executed volitionally (VOL). METHODS: Seven healthy adults were studied in a 1.5-T MR scanner. One hertz TMS at 110% of motor threshold was applied over the motor cortex for the thumb in 21 pulse trains in alternation with VOL every 63 seconds and interleaved with functional MR imaging. RESULTS: BOLD increases in motor cortex associated with TMS and VOL movement were similar (2%-3%). Mean separation of their centers of activity was 3.7 + 1.9 mm (mean displacement: left/right = 0.3 +/- 4.1 mm; superior/inferior = 0.7 +/- 1.9 mm). There was no indication of supraphysiological brain activity. CONCLUSIONS: Motor cortex BOLD response associated with thumb movement induced by 1-Hz TMS at 110% motor threshold is similar in both location and level to that caused by a similar movement executed volitionally. PMID- 11110305 TI - Hemostatic effect of glue-lipiodol mixture plugged in the needle tract after renal biopsies in a high-risk, anticoagulated rabbit model. AB - RATIONALE AND OBJECTIVES: To investigate the hemostatic effect of a glue-lipiodol mixture plugged immediately into the needle tract after renal biopsy of high risk, anticoagulated rabbits by use of a large-core gun biopsy needle and the subsequent pathological changes of the biopsy tract. METHODS: Twenty-five rabbits weighing 2 kg were divided into five groups (five rabbits each) according to time of sacrifice at 0, 2, 4, 6, and 8 days after biopsy. After anesthesia was induced, both kidneys were exposed and a bolus of 100 U/kg heparin was administered intravenously. Blood sampling was done twice, once before and once after heparinization, and activated partial thromboplastin times were measured for each. Then renal biopsies were performed in 25 pairs of kidneys by using an 18-gauge automated biopsy gun. The glue-lipiodol mixture was not injected into the first biopsy sites (control). Immediately after the second biopsies were done, 0.5 mL of the glue-lipiodol mixture at a 1:3 ratio was plugged into the needle tract through the outer cannula while withdrawing it slowly. Bleeding times of the two biopsies of each pair of kidneys were measured. All rabbits were humanely killed at their previously scheduled times, and histopathological findings were evaluated for the presence of inflammation, necrosis, foreign body reaction, and fibrosis around the biopsy tract. All parameters were classified into four categories according to the degree of severity, from 0 to + + +. RESULTS: Twenty-five cases (100%) of the controls bled after the biopsies were performed. Four of them (16%) showed immediate, massive, pulsatile bleeding. In contrast, only 11 of 25 cases (44%) bled at the second biopsy (plugged) sites and only two of them (8%) showed pulsatile bleeding. Mean bleeding times were 228 seconds in controls and 26 seconds in the glue-lipiodol mixture-plugged sites (P < 0.000). Histopathological examination of the needle tract in the plugged group revealed mild inflammation in the 0- to 2-day groups and moderate inflammation and mild necrosis in the 4- and 6-day groups. In the 8-day group, inflammation was diminished and only mild fibrosis was noted. There was no foreign body reaction in any of the specimens of the plugged group. CONCLUSIONS: We conclude that a glue-lipiodol mixture, when used as a plug material for the renal biopsy tract, is an efficacious method of bleeding control that is associated with a low incidence of pathological alterations in an anticoagulated rabbit model, thus demonstrating its future potential for clinical application. PMID- 11110306 TI - Arteriovenous malformations: assessment of gliotic and ischemic changes with fluid-attenuated inversion-recovery MRI. AB - RATIONALE AND OBJECTIVES: To evaluate the diagnostic potential of fluid attenuated inversion-recovery (FLAIR) MRI in the assessment of patients with cerebral arteriovenous malformations (AVMs) and to correlate the MR findings with clinical symptoms, in particular, perilesional gliosis and ischemic changes. METHODS: Forty-five patients with cerebral AVMs were examined with FLAIR and conventional T1- and T2-weighted MRI by using identical slice parameters. Images were assessed in a two-reader study for detection and delineation of gliotic and ischemic tissue. Also, the extent of the flow void phenomenon and image artifacts were evaluated. RESULTS: FLAIR MRI was rated superior to the conventional T2 weighted fast spin-echo imaging in the assessment of intralesional and perilesional gliosis. The superior delineation was a result of the suppression of cerebrospinal fluid, mild T1 weighting, and the more pronounced flow void phenomenon. There was no significant correlation between the extent of gliosis and the clinical symptoms. However, larger AVMs had more extensive signal changes. CONCLUSIONS: FLAIR is a valuable MRI technique to assess gliotic and ischemic changes in or close to cerebral AVMs. Because gliotic and ischemic changes are common findings and are known to be associated with epilepsy, in the assessment of these patients FLAIR is clinically useful and may guide decisions about treatment-for instance, the extent of surgical resection of the potential epileptogenic focus. PMID- 11110307 TI - Preoperative and postoperative assessment of laparoscopic inguinal hernia repair by dynamic MRI. AB - RATIONALE AND OBJECTIVES: To determine the value of dynamic MRI for seroma detection, hernia recurrence, and mesh placement in patients after laparoscopic inguinal hernia repair. METHODS: Thirteen inguinal hernias in 10 consecutive patients were evaluated before and after surgery by using an MRI protocol consisting of coronal T1-weighted (fast field echo) and T2-weighted (turbo spin echo) images and two sequences obtained during straining (turbo field echo gradient technique). All patients underwent a transabdominal preperitoneal laparoscopic inguinal hernia repair. MRI scans were reviewed for the presence of postoperative fluid collections, recurrent hernia, and mesh localization. RESULTS: In all patients, an inguinal hernia was identified on the preoperative MRI and was absent on the postoperative MRI. In all patients treated laparoscopically, the mesh and its position were clearly identified. Three small fluid collections were found on the postoperative MRI scans. CONCLUSIONS: Dynamic MRI can demonstrate small, postoperative fluid collections and a sufficient hernioplasty by showing the proper position of the mesh and the absence of a hernia. PMID- 11110308 TI - LASIK advertising 2000--are we putting our wallets in front of our wisdom? PMID- 11110309 TI - Cut edges and surface characteristics produced by different microkeratomes. AB - PURPOSE: To determine the cutting characteristics of seven different microkeratomes and to compare the cut edges and surface characteristics of the corneas with respect to different keratome parameters, ie, blade oscillation frequencies and keratome speed. METHODS: Lamellar keratectomies were performed using each microkeratome on eight freshly enucleated porcine corneas. The freshly cut corneal bed was then examined using scanning electron microscopy. A scoring system was used to evaluate the serration of the cut edge and the regularity of the corneal wound bed. RESULTS: Serrated cut edges were produced by the Microtech microkeratome, the Automatic Corneal Shaper, the Draeger rotor keratome, and the Schwind microkeratome. The other tested cutting devices generally yielded a smooth cut edge. Smooth and regular wound surfaces were obtained with the Schwind microkeratome, the Automatic Corneal Shaper, the Microtech microkeratome, and the MKM set. The specimens cut with the Schwind microkeratome showed particularly regular surface characteristics. CONCLUSION: The relationship between keratome propulsion speed, blade oscillation frequency, and blade material appears crucial for the quality of the microkeratome cut. Our results favor a low advancement/oscillation ratio among automated microkeratomes. PMID- 11110310 TI - Laser in situ keratomileusis for correction of high astigmatism after penetrating keratoplasty. AB - PURPOSE: To evaluate the safety and efficacy of laser in situ keratomileusis (LASIK) for correction of high astigmatism after penetrating keratoplasty, and to assess the refractive results and predictability of the procedure. METHODS: LASIK was performed on 19 patients (19 eyes) with high astigmatism after penetrating keratoplasty, using the Chiron Automated Corneal Shaper and the Chiron-Technolas Keracor 116 excimer laser. The amount of preoperative refractive astigmatism ranged from 6.50 to 14.50 D (mean, 9.21 +/- 1.95 D) and the spherical component of manifest refraction ranged from -7.00 to +1.25 D (mean, -2.14 +/- 2.11 D). All patients completed a minimum follow-up of 12 months. RESULTS: Refraction was stable after 3 months. At 1 year after LASIK, the amount of refractive astigmatism was reduced to a mean of 1.09 +/- 0.33 D (range, 0.50 to 1.75 D), with 57.9% of the eyes within +/- 1.00 D of refractive astigmatism. The mean percent reduction of astigmatism was 87.9 +/- 3.7%. The postoperative spherical component of manifest refraction ranged from -1.00 to +1.75 D with a mean of +0.43 +/- 0.82 D. Vector analysis showed that the mean amount of axis deviation was 1.1 +/- 1.3 degrees and the mean percent correction of preoperative astigmatism was 92.6 +/- 8.4%. There were no intraoperative complications. Spectacle-corrected visual acuity was not reduced in any eye, and improved by 2 or more lines in 42.1% of eyes after LASIK. CONCLUSION: LASIK with the Chiron Technolas Keracor 116 excimer laser was effective for correction of both astigmatism and myopia after penetrating keratoplasty. The procedure proved to be safe and gave fairly predictable and stable refractive results. PMID- 11110311 TI - Comparison of photorefractive keratectomy and laser in situ keratomileusis for myopia of -6 D or less using the Nidek EC-5000 laser. AB - PURPOSE: We compared the efficacy, predictability, and safety of photorefractive keratectomy (PRK) and laser in situ keratomileusis (LASIK) for the surgical correction of low and moderate myopia. METHODS: A retrospective study was performed to evaluate uncorrected and spectacle-corrected visual acuity, and manifest refraction 1 year after PRK or LASIK. All procedures were done using an automatic microkeratome (Chiron Ophthalmic) and the Nidek EC-5000 excimer laser. RESULTS: PRK was performed in 75 eyes of 45 patients and LASIK in 133 eyes of 77 patients. Mean age for PRK patients was 32.8 years (range, 18 to 52 yr) and LASIK patients was 29.6 years (range, 18 to 49 yr). Mean preoperative spherical equivalent refraction for PRK patients was -3.28 D (range, -1.00 to -6.00 D) and LASIK, -3.86 D (range, -1.00 to -6.00 D). One year after surgery, mean spherical equivalent refraction for Group 1 (baseline, -1.00 to -3.00 D) PRK eyes was -0.18 +/- 0.61 D (range, -1.50 to +0.75 D) and for LASIK eyes, -0.08 +/- 0.61 D (range, -1.50 to +1.62 D), with no statistically significant difference. For Group 2 eyes (baseline, -3.25 to -6.00 D), mean spherical equivalent refraction for PRK eyes was -0.44 +/- 0.87 D (range, -2.00 to +2.12 D) and for LASIK eyes, -0.09 +/- 0.83 D (range, -1.50 to +1.75 D), with no statistically significant difference. The antilogarithm of the mean UCVA (antilogUCVA) in Group 1 for PRK was 0.79 +/- 0.21 (20/25) and for LASIK was 0.87 +/- 0.19 (20/23), with no statistically significant difference. The antilogUCVA in Group 2 for PRK eyes was 0.70 +/- 0.24 (20/28) and for LASIK eyes was 0.83 +/- 0.18 (20/24), with a statistically significant difference (0.7 vs. 0.83, P < .005). The percentage of eyes with a postoperative UCVA >20/40 in Group 1 for PRK was 91.5% (38 eyes) and for LASIK was 95% (50 eyes) (no statistically significant difference), and in Group 2 for PRK eyes, it was 82% (27 eyes) and 97.5% (78 eyes) for LASIK (statistically significant difference, P < .05). CONCLUSION: PRK and LASIK with the Nidek EC 5000 excimer laser are effective and safe for correcting low to moderate myopia, but LASIK eyes showed better results for moderate myopia in terms of uncorrected visual acuity. PMID- 11110312 TI - Laser in situ keratomileusis for myopia and hyperopia using the Lasersight 200 laser in 300 consecutive eyes. AB - PURPOSE: To evaluate effectiveness, safety, predictability, and short-term stability of laser in situ keratomileusis (LASIK) using the LaserSight Compac-200 Mini excimer laser with software version 9.0, for all refractive errors. METHODS: One hundred fifty consecutive patients (300 eyes) that received bilateral LASIK for myopia, hyperopia, and astigmatism were studied prospectively. A new 9.0 software version applying a modified nomogram that takes advantage of bilateral surgery was used. Follow-up at 6 months was available for 267 eyes (89%). RESULTS: Six months postoperatively, 131 eyes (96.32%) in the low to moderate myopia group (-1.00 to -5.99 D; n=136) had a spherical equivalent refraction within +/-1.00 D, and 123 eyes (90.44%) were within +/-0.50 D of emmetropia. In the high to extreme myopia group (-6.00 to -25.00 D; n=114), 97 eyes (87.08%) had a spherical equivalent refraction within +/-1.00 D and 78 eyes (68.42%) were within +/-0.50 D of emmetropia. In the hyperopia group (+1.00 to +6.00 D; n=50), 44 eyes (88%) had a postoperative spherical equivalent refraction within +/-1.00 D, and 31 eyes (62%) were within +/-0.50 D of emmetropia. Mean change in spherical equivalent refraction at 6 months was less than -0.50 D in the low to high myopia groups and -1.16 +/- 0.55 D in the extreme myopia group. At 6 months follow-up, uncorrected visual acuity was 20/20 or better in 73 eyes (54%) in the low to moderate myopia groups and 21 eyes (18%) in the high to extreme myopia groups. In the hyperopia group at 6 months follow-up, uncorrected visual acuity was 20/20 or better in 31 eyes (62%) and 20/40 or better in 41 eyes (82%). Only two eyes had a temporary loss of two or more lines of spectacle-corrected visual acuity due to corneal folds that were surgically treated. Six months after LASIK, no eye had lost any lines of best spectacle-corrected visual acuity in this series. CONCLUSIONS: Our modified LASIK nomogram with the 9.0 software of the LaserSight 200 excimer laser (with a larger and smoother ablation pattern) resulted in safe and effective outcomes for the treatment of low to high myopia, astigmatism, and hyperopia. PMID- 11110313 TI - Prospective study of photorefractive keratectomy for hyperopia using an axicon lens and erodible mask. AB - PURPOSE: To evaluate prospectively the long-term safety, efficacy, and visual performance following photorefractive keratectomy (PRK) for hyperopia using an erodible mask and axicon lens system. METHODS: Eighteen eyes of 9 patients with a mean preoperative spherical equivalent refraction of +2.26 +/- 0.82 D (range, +1.13 to +4.00 D) underwent PRK with the Summit Apex Plus excimer laser following manual scraping of the epithelium. Eyes were prospectively evaluated 1, 3, 6, 9, 12, 18, and 24 months following the procedure. Primary outcome variables included cycloplegic refraction and uncorrected visual acuity (UCVA). Visual performance was determined by contrast sensitivity measurements under scotopic (21 lux) and photopic (324 lux) conditions and best spectacle-corrected visual acuity (BSCVA) under scotopic, photopic, and glare conditions. RESULTS: For 18 eyes, 98.2% of the mean preoperative spherical equivalent refraction was corrected to +0.04 +/- 0.87 D (range, -1.38 to +2.00 D) at 24 months after PRK. Twelve eyes (67%) were within +/-0.50 D of attempted correction and 15 eyes (83%) were within +/-1.00 D. Stability within +/-0.50 D was achieved after 6 months. Two eyes (11%) experienced almost complete regression of the refractive effect. There was no statistically significant decrease in contrast sensitivity under scotopic or photopic conditions. (P > .05). Best spectacle-corrected visual acuity showed progressive improvement in the early postoperative period. By 24 months, 0 eyes (0%) lost 2 or more lines of BSCVA under scotopic and photopic conditions and 1 eye (5.5%) lost 2 or more lines under glare conditions. Fourteen eyes (78%) had grade 1 to 3 anterior stromal haze at 24 months which was characteristically mid peripheral and did not adversely affect visual performance. CONCLUSION: Photorefractive keratectomy with the the Summit Apex Plus excimer laser for low to moderate hyperopia resulted in an effective reduction of hyperopia without compromising long-term visual performance. Stability and recovery of distance uncorrected and best spectacle-corrected visual acuity took approximately 6 months. PMID- 11110314 TI - Photorefractive keratectomy in ophthalmic residents. AB - PURPOSE: To find out how ophthalmologists themselves experience the correction of myopia after photorefractive keratectomy. Visuomotor functions were of special interest. METHODS: Four ophthalmology residents and one medical engineer underwent photorefractive keratectomy for myopia. Objective measurements including refraction, corneal topography, perimetry, contrast sensitivity, pattern visual evoked potentials, in vivo confocal microscopy, and a car driving simulator test were performed preoperatively, postoperatively, and at 6 months. Subjective evaluation was reported. RESULTS: Performing ophthalmological examinations and microsurgery without spectacles was easier postoperatively and was appreciated by the four ophthalmology residents. Minimal haze formation, good accuracy, and normal performance in the car driving simulator were also observed. Visual fields, contrast sensitivity, and pattern visual evoked potentials did not show changes. Negative observations included postoperative pain for 2 to 4 days, dry eye symptoms, a period of anisometropia between operations, and hypersensitivity of the lids. CONCLUSIONS: The four ophthalmic residents were satisfied with the outcome of their refractive surgery. Low to moderate myopic correction did not affect the objective measurements of high and low contrast sensitivity, pattern visual evoked potentials, or simulated car driving in dark illumination. PMID- 11110315 TI - Early rhegmatogenous retinal detachment following laser in situ keratomileusis for high myopia. AB - PURPOSE: Four eyes had early rhegmatogenous retinal detachment within 3 months of laser in situ keratomileusis (LASIK) for correction of high myopia using the microkeratome, Clear Corneal Molder. METHODS: In two eyes, retinal detachment resulted from horseshoe tears, one occurring in an otherwise normal region of the retina and the other at the margin of an area of lattice degeneration detected during preoperative examination. The first eye was treated with retinopexy using a 287 encircling scleral exoplant, drainage of subretinal fluid, and laser photocoagulation by indirect ophthalmoscopy. The other eye was treated with pneumatic retinopexy and cryotherapy. In the other eyes, retinal detachment was the result of giant tears with no evidence of prior retinal degeneration. These eyes were treated with pars plana vitrectomy, fluid-gas exchange with 15% perfluoropropane (C3F8), endolaser photocoagulation, and a 42 encircling scleral exoplant. RESULTS: After treatment, the first two eyes achieved spectacle corrected visual acuity of 20/40. In the last two eyes, final spectacle-corrected visual acuity was 20/400 in one eye and light perception in the other. CONCLUSIONS: Although no cause-effect relationship between LASIK and retinal detachment can be stated, these cases suggest that LASIK may be associated with retinal detachment, particularly in highly myopic eyes. Further studies are necessary to determine high-risk patient characteristics. PMID- 11110316 TI - Late onset diffuse lamellar keratitis associated with an epithelial defect in six eyes. AB - PURPOSE: To report six cases of late onset diffuse lamellar keratitis associated with epithelial defects 2 to 12 months following uncomplicated laser in situ keratomileusis (LASIK). METHODS: Retrospective case series. RESULTS: The interface inflammation and epithelial defects were treated aggressively with topical corticosteroids and topical antibiotics with complete resolution over 1 to 2 weeks. There were no complications or loss of best spectacle-corrected visual acuity. These cases illustrate new understanding in the etiology of diffuse lamellar keratitis following lamellar surgery. CONCLUSION: Late onset diffuse lamellar keratitis in association with epithelial defects may occur following LASIK. Treatment with topical antibiotics and topical corticosteroids may result in uncomplicated, complete resolution. PMID- 11110317 TI - Diffuse lamellar keratitis induced by trauma 6 months after laser in situ keratomileusis. AB - This case report illustrates an unusual presentation of diffuse lamellar keratitis triggered by a foreign body striking the eye 6 months after laser in situ keratomileusis (LASIK). The etiology of diffuse lamellar keratitis is unclear. The infiltrate within the plane of the flap after removal of the foreign body supports the theory that diffuse lamellar keratitis is an inflammatory reaction. PMID- 11110318 TI - PRK as a possible risk factor in cataractogenesis. PMID- 11110319 TI - Case report of ointment in the anterior chamber after radial keratotomy. PMID- 11110320 TI - Corneal iron pigmentation after LASIK for hyperopia. PMID- 11110321 TI - Measuring corneal refractive power after LASIK. PMID- 11110322 TI - Standard graphs for reporting refractive surgery. PMID- 11110323 TI - Come on in, the hot tub's fine! PMID- 11110324 TI - Maternal surgery. PMID- 11110325 TI - Counseling of Norplant users. PMID- 11110326 TI - Caring for women living with ovarian cancer: recommendations for advanced practice nurses. AB - This article summarizes advice for advanced practice nurses (APNs) that grew out of research with women living with premenopausal ovarian cancer. We claim that the process of diagnosis and being told, battle metaphors, treatment expectations, the patient's sense of normalcy, her sense of being heard, her ability to make sense of her new world, her inability to have children, issues of sexuality, and the irrelevance of most support groups are important considerations in the treatment of such women. The APN's major role in caring for these women is understanding the experience as it informs the APN's practice and serving as advocates for the women. PMID- 11110327 TI - The nurse's role in misoprostol induction: a proposed protocol. AB - The use of misoprostol (PGE 1) for induction of labor has been studied in the United States since 1993. The current medical literature contains studies of this non-FDA-approved indication for misoprostol. The data reveal that misoprostol is as effective or more effective in inducing labor as are oxytocin and prostaglandin E2 (PGE 2), with less cost. Nurses need to understand the physiology of prostaglandins and management of misoprostol in labor. Nurses often must assess the safety of mother and fetus during a misoprostol induction without a protocol based on research findings. This article presents a sample protocol. PMID- 11110328 TI - Effect of less frequent bathing of preterm infants on skin flora and pathogen colonization. AB - OBJECTIVE: To determine if less frequent bathing alters colony count or type of organism in skin flora of preterm infants. DESIGN: Descriptive, repeated measures study. SETTING: A regional neonatal intensive-care unit. PARTICIPANTS: Forty-five preterm infants, 31 weeks mean gestational age (SD +/- 1.6 weeks) and 17 days mean postnatal age (SD +/- 3.7 days). INTERVENTIONS: Before the study, all infants received a bath every other day. On Day 1 of the study, a routine sponge bath was given, then no further bathing was performed for 4 days. MAIN OUTCOME MEASURE: Serial axillary skin cultures to identify the number of colony forming units (CFU) and type of organism were obtained within 30 minutes of the bath on Day 1 and at the same time on Days 2, 3, and 4. RESULTS: Normal skin flora CFU count, predominantly coagulase-negative staphylococci, increased within 48 hours after bathing compared to values 30 minutes after bathing. There were no differences in normal skin flora CFU on Days 2, 3, and 4. Pathogens were identified in 12 infants for at least one time point during the study. Significantly fewer pathogens were found in the cultures over time, despite longer interval since bathing, and no infant developed symptoms of infection during the study period. CONCLUSION: Findings from this study suggest that the frequency of bathing of preterm infants can be reduced without increasing the risk of infection. PMID- 11110329 TI - Sleep disturbances during pregnancy. AB - OBJECTIVE: To assess sleep patterns and prevalence of sleep disturbances during pregnancy. DESIGN: Cross-sectional design; prospective questionnaire. SETTING: Outpatient, private obstetric clinic. PARTICIPANTS: 127 consecutive patients, with women evaluated at one of four points in pregnancy, 8-12 weeks (n = 37), 18 22 weeks (n = 28), 25-28 weeks (n = 24), and 35-38 weeks (n = 38). MAIN OUTCOME MEASURE: Questionnaire of sleep habits and sleep disturbances. RESULTS: A large percentage of the women experienced sleep disturbances during pregnancy, These problems included frequent night wakings, difficulty falling asleep, and symptoms of sleep apnea. Few differences in sleep patterns were found across pregnancy, although women were found to sleep more and nap more by the end of pregnancy. CONCLUSION: Sleep disturbances are common during pregnancy, especially late in pregnancy. PMID- 11110330 TI - Maternal-fetal attachment in twin pregnancies. AB - OBJECTIVE: To describe maternal attachment to twin fetuses and determine whether such attachment differs between the twins. DESIGN: A descriptive, correlational, nonexperimental design was used to collect data on a convenience sample of high risk and low-risk women pregnant with twins. SETTING: Questionnaires were distributed or mailed by presidents of parents of multiples clubs to interested participants. Participants mailed the questionnaires back to the researcher via stamped return envelope. PARTICIPANTS: During a 14-month period, 214 eligible women from 41 states and the District of Columbia completed the survey instruments. MAIN OUTCOME MEASURE: The Prenatal Attachment Inventory. RESULTS: Pregnant women reported greater attachment to Twin B than to Twin A. CONCLUSION: Pregnant women may be more attached to Twin B (typically the nonpresenting fetus) because this twin may be easier to see and touch. While the mean attachment scores between twins differed, the size of the difference was small. Further study is needed to determine the predictive value and clinical significance of this finding for future mother-infant relationships. PMID- 11110331 TI - Bacterial vaginosis and Chlamydia trachomatis among pregnant abused and nonabused Hispanic women. AB - OBJECTIVE: To compare the prevalence of bacterial vaginosis (BV) and Chlamydia trachomatis (CT) among abused and nonabused pregnant Hispanic women. DESIGN: Retrospective audit of charts of 233 pregnant, abused Hispanic women and 468 pregnant, nonabused Hispanic women. SETTING: Three urban prenatal clinics of a public health department in the southwestern United States. SAMPLE: The medical records of 701 pregnant Hispanic women. MAIN OUTCOME MEASURE: Diagnosis of BV and/or CT among abused and nonabused pregnant women. RESULTS: Combined prevalence of BV and CT was significantly higher for abused women (z score = 2.55; df = 138; p < .05). There was no significant difference between abused and nonabused women for CT alone (z score = .96; df = 33; p < .05); however, prevalence of BV was significantly higher for abused women (z score = 1.99; df = 104; p < .05). CONCLUSION: In this sample of pregnant Hispanic women prevalence of BV was significantly higher in those who had been abused, indicating the need for targeted screening of all abused pregnant women for BV. PMID- 11110332 TI - Mothers' ideas about their role in feeding their high-risk infants. AB - OBJECTIVE: To describe how mothers of preterm infants who are learning to nipple feed view their own and their infant's role in the feeding process. DESIGN: Descriptive, comparative study. SETTING: Two neonatal intensive-care units (NICU) in the Midwest. PARTICIPANTS: A convenience sample of 22 mothers of very-low birth-weight infants. MAIN OUTCOME MEASURES: Interviews were rated for mothers' level of thinking about their co-regulatory role in their infants' feeding on a 6 point scale. The higher the score the more flexible, contingent, adaptive, and reflective the mother's thinking is regarding her infant's and her own behavior during feeding. High and low scores were compared using paired t tests. RESULTS: The co-regulation scores ranged from 1 to 6, with a mean score of 3.3 (SD = 1.4). Mothers scoring higher on the co-regulation measure were significantly older and their infants were younger gestationally at birth. Their infants tended toward having been in the NICU for a longer period of time and spending more days on oxygen. CONCLUSIONS: Understanding feeding from the parents' perspective can guide clinicians as they support the development of parents' feeding skills. Intervention, rather than beginning with how to feed, may need to begin with how to take the infant's perspective, how to explore infant behavior that is novel, and ways of viewing the process of feeding as co-regulated. PMID- 11110333 TI - Variation in feminine hygiene practices as a function of age. AB - OBJECTIVE: To examine variation in feminine hygiene practices as a function of increasing age. DESIGN: A nonexperimental, descriptive research design with study sample stratified by age. PARTICIPANTS: Women over 18 years of age. Of 713 women who completed and returned the feminine hygiene practices questionnaire, 180 were younger than 41 years, 171 were 41-48 years, 184 were 49-57 years, and 178 were 58 years or older. All participants were members of a California professional home economics organization. MAIN OUTCOME MEASURES: A 40-item feminine hygiene practices questionnaire dealing with body cleansing practices, feminine products usage, and both general and specific menses collection and protection practices. RESULTS: Significant age-related differences were found in several areas of body cleansing and feminine products usage, including sponge bathing and use of feminine and deodorant spray, wet wipes, and panty liners. In all age groups, 20 30% of women reported douching, even in the face of continued reports of the dangers in this practice. In addition, reduced numbers of women (n = 245) in all age groups reported washing hands before using tampons or pads, although more (n = 314) reported washing their hands afterward. CONCLUSIONS: Continuing education about proper feminine hygiene practices, especially regarding douching and handwashing before and after genitourinary contact, will be important across all age groups. PMID- 11110334 TI - Pregnancy and breast cancer. AB - Pregnancy-associated breast cancer (PABC) and pregnancy subsequent to breast cancer are two areas of concern facing women of childbearing age. The current approach to the management of PABC is to treat the cancer with some modification because of the pregnancy. The clinical management of both PABC and pregnancy occurring after breast cancer in young survivors, with emphasis on issues in clinical decision making, clinical management, and client education and support, are addressed. PMID- 11110335 TI - Tomatoes, Pap smears, and tea? Adopting behaviors that may prevent reproductive cancers and improve health. AB - Women who adopt healthy lifestyles and practice preventive healthy behaviors can reduce the risks of some cancers and other diseases, such as heart disease and sexually transmitted infections. Risk factors associated with the most common gynecologic cancers and breast cancer are identified, diagnostic and screening alternatives are described, and preventive health practices and information to include when counseling women about making healthy choices are suggested. PMID- 11110336 TI - Helping women and their families cope with the impact of gynecologic cancer. AB - The impact of gynecologic cancer on the woman and her family depends on psychosocial factors and cancer-specific factors. Family assessments determine how the family is adapting to the woman's illness. Nursing interventions that families say are helpful include providing physical care, providing information, and giving support. Strategies used by families to cope with the stress and emotional strain of caregiving include taking time for themselves, maintaining a sense of humor, and focusing on the present. PMID- 11110337 TI - The evolution of the nurse practitioner role in the neonatal intensive care unit. AB - The role of the Neonatal Nurse Practitioner has been evolving since the early 1970s. The original concept and design was born out of individual hospital's needs for highly professional and skilled personnel at the bedside. Thus, initial programs were hospital-based and granted certificates. Over the past 20 years, a gradual shift toward graduate degrees and standardization of programs has been seen. The role and responsibilities of the neonatal nurse practitioner have also expanded over that time period. From their strictly clinical beginnings, neonatal nurse practitioners now contribute to research, nursing and medical education, and administration. This article looks at the neonatal nurse practitioner role and the impact that education and legislation have had on its evolution. PMID- 11110338 TI - Single umbilical artery is associated with an increased incidence of structural and chromosomal anomalies and growth restriction. AB - The objective to characterize neonatal outcome associated with ultrasonographic identification of a single umbilical artery. Pregnancies diagnosed with single umbilical artery antenatally were identified. All prenatal/antenatal and pediatric records were reviewed for maternal demographics, associated anomalies, karyotypic analysis, pregnancy complications, and neonatal outcome. Twenty-seven pregnancies complicated by fetal single umbilical artery were identified. Of the 27 pregnancies, 5 (18.5%) underwent pregnancy termination and 1 (3.7%) experienced fetal demise. Of the 21 liveborn infants, 4 (19%) died within the first year of life. Sixty-seven percent of fetuses had an associated structural anomaly. Sixteen of the 27 pregnancies underwent amniocentesis and 7 of these were chromosomally abnormal. All of the karyotypically abnormal fetuses had a structural defect in addition to the single umbilical artery. Of the six fetuses without any associated structural or chromosomal anomalies, three (50%) demonstrated growth restriction. Single umbilical artery is relatively rare finding. When a single umbilical artery is identified, a vigilant search for associated anomalies should be undertaken. Pregnancies identified as having fetuses with associated structural anomalies should be offered amniocentesis. Pregnancies with isolated single umbilical artery should be carefully monitored for evidence of fetal growth restriction. PMID- 11110339 TI - Comparison of clinical versus ultrasound estimation of fetal weight. AB - A prospective study was conducted of the accuracy of fetal weight estimation comparing clinical and ultrasound estimation with actual birth weight in 200 Iranian pregnant women. The results showed that there was statistically significant difference between clinical estimate of fetal weight and actual birth weight, as well as between ultrasound estimation and actual birth weight. The mean error of clinical estimation was 101 g, or 32 g/kg, for a 3.2% error. The error of ultrasound in the same population was 141 g, or 45 g/kg, for a 4.5% error. Analysis of these data revealed statistically significant difference between the mean error of clinical estimation and ultrasound estimation of fetal weight. Our study indicates that the mean clinical estimate of fetal weight is equal to ultrasound for the estimation of fetal weight in our population. This has important implications for developing countries where there is a lack of technologically advanced ultrasound machines capable of performing sophisticated functions like fetal weight estimation. PMID- 11110340 TI - Effect of antibiotic therapy in preterm premature rupture of the membranes on neonatal mortality and morbidity. AB - The objective of this study was to evaluate the effect of prophylactic antibiotic treatment on prenatal and neonatal outcome after preterm premature rupture of the membranes (P-PROM). The study population consisted of 172 pregnant women admitted to the Rabin Medical Center in Israel with P-PROM at 23-34 gestational weeks. The patients were divided into two groups by mode of expectant management prophylactic ampicillin and erythromycin (study group, n = 121) or observation only (control group, n = 51) and compared for neonatal outcome. There were no significant differences between the groups in age, gravidity, parity, or gestational age at admission. Significantly more women in the study group had an interval of more than 24 hours from onset of P-PROM to delivery (98 women, 81%) than the control group (32 women, 63%) (p = 0.001). The neonatal mortality rate was significantly lower in the study group (6.6%) than the control group (1 7.6%) (p = 0.03). The lower neonatal mortality rate associated with prophylactic antibiotic treatment in P-PROM has important clinical implications for the effectiveness of this type of management. PMID- 11110341 TI - Frequent presence of Helicobacter pylori genome in the saliva of patients with hyperemesis gravidarum. AB - Recently, possible involvement of Helicobacter pylori (H. pylori) in hyperemesis gravidarum have been reported based on serological studies and the therapeutic effects of antibiotics. In this study, we examined for the presence of H. pylori genome [by (PCR) of saliva] in combination with serological techniques. Thirty four patients with hyperemesis and 29 normal pregnant subjects were examined for serum anti-H. pylori IgG antibodies and PCR of saliva. By serum antibody test, 16 of 34 hyperemesis patients (47.5%) were positive for anti-H. pylori IgG antibody, while 6 of 29 control subjects (20.6%) were positive (chi2 p < 0.0005). PCR revealed positive H. pylori genome in 21 cases out of 34 hyperemesis (61.8%, 14 of 16 patients positive for H. pylori antibody and 7 of H. pylori-antibody negative 18 patients) and 8 of 29 control subjects (27.6%) (chi2 p < 0.000001). We suggest chronic infection of H. pylori as one of the important factors on the pathogenesis of hyperemesis gravidarum even though it may not be the single cause of the disorder. PMID- 11110342 TI - Transient renal tubular acidosis in pregnancy. AB - Renal tubular acidosis in pregnancy is a very rare disorder. Most cases are either inherited or secondary to maternal disease or ingestion of toxic chemicals. We report a 22-year-old woman, previously healthy, who presented at 27 weeks of gestation with preterm labor. Investigation revealed renal tubular acidosis with no obvious etiology. Labor was stopped with various tocolytic drugs and her electrolyte imbalance was corrected. She was delivered at 36 weeks, by cesarean for a nonreassuring fetal heart tracing, of an appropriate-for gestational-age infant weighing 2905 g. Evaluation 3 and 6 months postpartum revealed gradual, but complete resolution of the acidosis and electrolyte abnormality. The infant is now 7 months old, is growing normally with normal electrolytes, and with no evidence of acidosis. PMID- 11110343 TI - Congenital varicella syndrome: a rare case of central nervous system involvement without dermatological features. AB - An unusual case of congenital varicella syndrome with significant central nervous system involvement, but without dermatological features at birth is described. The mother contracted chicken pox at 15 weeks' gestation. Congenital varicella syndrome involves multiple systems, but rarely without skin lesions identifiable at birth. Although varicella infection in pregnant women is an uncommon complication, the fetal embryopathy that may result can be devastating. Antenatal diagnosis of fetal embryopathy during the first 20 weeks of pregnancy should be established by amniocentesis or cordocentesis when a mother presents in the first trimester with chicken pox, and appropriate risk counselling provided. PMID- 11110344 TI - Continuous measurement of core body temperature in preterm infants. AB - We tested a transcutaneous core temperature sensor using a method that relies on the principle of zero heat flow. We tested the hypothesis that transcutaneous and rectal temperatures would track within 0.3 degrees C of each other for >90% of the time. A thermistor was placed between the infant's abdomen or back and the incubator's or radiant warmer's mattress, or within the axilla, attached to the skin with a foam adhesive disk insulator. Thirty preterm infants were either placed on their abdomens or backs in a convective incubator or under a radiant warmer, and continuous transcutaneous and rectal temperatures were measured for 1 hour. There were no significant differences between abdominal and core temperatures or between axillary and core temperatures measured in double-walled convective incubators or in radiant warmers. The rectal-abdominal temperature difference was significantly less than the rectal-axillary difference (p < 0.02) in convective incubators, but not when the infant was placed prone under radiant warmers (p = 0.27). Transcutaneous thermometry is reliable for monitoring core body temperature as indicated by rectal temperature in stable preterm infants in a convective incubator. PMID- 11110345 TI - Instrumental delivery of the fetal head at the time of elective repeat cesarean: a randomized pilot study. AB - We sought to ascertain whether the routine use of instruments, forceps or vacuum, at the time of elective repeat cesarean delivery, permits a delivery that is as safe for mother and infant and as easy for mother and physician as traditional manual delivery of the fetal head. In this prospective study 44 women undergoing elective repeat cesarean were randomized to deliver by Vacuum (V), Forceps (F), or by Manual (M) means. Groups were compared with regard to demographic variables and maternal and neonatal outcomes. Deliveries were timed from entry into the uterus until full delivery of the infant. Maternal pain scores were assessed using a 10-cm visual analog scale. There were no differences in demographic variables except that the M group had fewer women with up to two cesareans. A large percentage of women in each group were delivered with the randomized instrument. Use of the V did not demonstrate fewer extensions of the uterine incision or lesser amounts of blood loss as measured by serial hemoglobin determinations. There was a trend for the F group to require a longer period of time for delivery (p = 0.061). Women in the V group reported significantly lower pain scores (p = 0.015). There were no serious neonatal injuries. The routine use of instruments at the time of elective repeat cesarean delivery appears safe and effective. PMID- 11110346 TI - Low-dose continuous indomethacin in early days of age reduce the incidence of symptomatic patent ductus arteriosus without adverse effects. AB - We conducted a comparative study to evaluate whether the low-dose continuous indomethacin therapy in the early days of age would reduce incidence of the symptomatic patent ductus arteriosus (PDA) without the adverse effects. Thirty seven infants were in the historical comparison group, and 39 infants were given low-dose indomethacin continuously (0.004 mg/kg/h) from 6-12 postnatal hours until the recognition of closing PDA. Low-dose continuous indomethacin significantly decreased the incidence of symptomatic PDA at 5 days of age (p < 0.01) as compared with the historical comparison group. There was no episode of decreasing urinary output and necrotizing enterocolitis in the indomethacin group. We conclude that the low-dose continuous indomethacin therapy results in decreasing the incidence of symptomatic PDA without significant adverse reactions. PMID- 11110347 TI - Stroke in blacks: a call for a unified expansion of perspective. PMID- 11110348 TI - Black/white differences in symptoms and health satisfaction reported by older hemodialysis patients. AB - Although Black end-stage renal disease (ESRD) patients on dialysis report better functioning and well-being than do White patients, little is known about the association of race with disease symptoms and treatment side effects. Interviews were conducted with 183 older Black and 125 older White in-center hemodialysis (HD) patients in Georgia. Patients were identified in a stratified (by race and sex) random sample of patients aged 60+ years selected from the ESRD Network census of all patients in that age category. Self-assessed disease symptoms and/or side effects of treatment, disability days, and health satisfaction were measured. Data were analyzed via logistic or linear regression, controlling for the effects of patients' gender, age, months on dialysis, primary diagnosis of diabetes, cardiovascular co-morbidity, HD treatment time, and usual interdialytic weight gain. Older Whites, compared to older Blacks, were at increased risk for reporting nausea, sexual dysfunction, recent bed disability days, fatigue, greater HD recovery time, and health dissatisfaction. The relation of these complaints to dialysis adequacy and patients' nutritional status merits continued study. PMID- 11110349 TI - Treating hyperlipidemia in African Americans. AB - The purpose of this paper is to provide information concerning the treatment of hyperlipidemia in African Americans. There is a similar prevalence of hypercholesterolemia in Blacks and Whites, but Blacks have been neglected in most pharmacological studies of hyperlipidemia. Intimal atherosclerotic involvement of the aorta and coronary artery occurs in young Black and White males. Epidemiological studies suggest there is a higher mortality rate due to coronary disease in Blacks vs Whites in the United States. Thirty-four percent of Blacks require a fasting lipid profile and 9% of Black adults aged 20 years or older would require lipid-lowering therapy. However, fewer Blacks (26%) than Whites (47%) are aware of their hypercholesterolemia. Studies with lovastatin and pravastatin show efficacy in the treatment of hypercholesterolemia. The Antihypertensive and Lipid Lowering Treatment to Prevent Heart Attack Trial (ALLHAT) is designed to determine all-cause mortality for subjects receiving pravastatin vs a control group receiving "usual care." Randomized, blinded, prospective trials are needed to assess the impact of hyperlipidemia as a risk factor, and the impact of its reduction in African Americans. PMID- 11110350 TI - Lower hypertension prevalence in first-generation African immigrants compared to US-born African Americans. AB - Both genetic and environmental factors have been hypothesized to explain the higher prevalence of hypertension in US African Americans compared to populations still residing in western Africa. Studies of first-generation immigrants can help to identify risk factors for increased chronic disease expression in the developed world. Since we could identify no prior studies of hypertension in African immigrants to the United States, we conducted a cross-sectional survey of African-born and US-born African-American health professionals to compare the two groups for the prevalence of hypertension (blood pressure > or = 140/90 mm Hg or use of antihypertensive medication) and risk factors for hypertension (body mass index, lifestyle factors, and psychosocial variables hypothesized to relate to hypertension). Subjects were registered pharmacists and nurses recruited by mail. For the 182 individuals who completed study measurements (95 US-born and 87 African-born), the unadjusted odds ratio for hypertension associated with birthplace was 2.16 (95% CI = 1.12, 3.98). After adjustment for body mass index and age, the OR for birthplace was 1.92 (95% CI = 0.92, 4.00). No lifestyle or psychosocial variables were associated with hypertension prevalence. We conclude that there is a lower prevalence of hypertension in first-generation African immigrants that cannot be readily explained by the environmental effects measured in this study. Larger scale studies with African immigrants could advance understanding of the causes of the increased hypertension prevalence in US-born African Americans. PMID- 11110351 TI - Probability of coronary artery disease among patients attending primary health care centers (PHCCs) in Southwest Saudi Arabia. AB - In order to predict the magnitude of coronary artery disease (CAD) among Saudi patients attending the primary health care centers (PHCCs), the probability of coronary artery disease (PCAD) was estimated for 696 subjects (50.4% males and 49.5% females), aged 30-70 years, by a computer model based on the Framingham Heart Study. The results indicated a high percentage of CAD risk factors, namely: hypercholesterolemia (31%); diabetes mellitus (30%); hypertension (13.8%); family history of CAD (6%); and obesity (45%). The estimated PCAD rates within 5, 10 and 12 years were 4.31%, 9.88%, and 12.25%, respectively. PCAD was significantly higher among males (P=.0001) and those with two or more CAD risk factors (P=.0001). A repeated measure analysis showed that there was a consistent relation between the overall PCAD rates at 5, 10 and 12 years, and a relationship between PCAD rates and other risk factors (eg, males had a higher risk of CAD than females). Likewise, individuals with two or more risk factors had a higher PCAD. The present study predicts an incremental scale of developing CAD in the Saudi population, which may lead to an upward trend on CAD mortality similar to that of the WHO MONICA populations of most countries in 1950 and early 1960s. The concept of preventive cardiology aiming at decreasing the prevalence of CAD risk factors should be implemented in all regions of Saudi Arabia in order to avoid the predicted upward trend of CAD mortality and to get a favorable decline of CAD risk factors and mortality similar to that in the United States and Australia. PMID- 11110352 TI - The urinary levels of catecholamines, aldosterone and cortisol in hypertensive East Africans: a pilot study. AB - This pilot project studies the prevalence of hypertension among unique social groups in Kenya, as well as the hormonal profiles accompanying the hypertensive and normotensive states in these populations. The purpose of this report is to enlarge and improve upon the statistical data currently available concerning the prevalence, etiology and prognosis of hypertensive disease in this region. In this study, the urinary concentrations of three vasoactive metabolites were measured in hypertensive and normotensive outpatients. The excretion values for the metabolites were ultimately tabulated as the quantity excreted per milligram of creatinine. The results demonstrate that the subjects with elevated blood pressures (>140/90 mm Hg) excreted double the concentrations (ng/mg creatinine) of cortisol and aldosterone excreted by normotensives. There were no apparent differences in urinary catecholamines between hypertensives and normotensives. PMID- 11110353 TI - Prevalence and factors associated with high blood pressure in Korean Americans. AB - This study, conducted from October 1998 to February 1999, included a cross sectional epidemiological survey administered to Korean Americans (KAs) living in Maryland (N = 761). One third (32%) of participants had high blood pressure (HPB: SBP > or = 140 mm Hg and/or DBP > or = 90 mm Hg or were on hypertension medication). HBP was more common among males (35%) than females (30%) and those who were aged 50 years or older (53%) compared to those who were less than 50 years old (12%). The prevalence of HBP in Maryland KAs was found to be much higher than in other Americans (24%) and in their counterparts in Korea (22% overall: 29% in males, 11% in females). Only 40% of the hypertensive KAs were taking HBP medication, and 74% of those did not have controlled HBPs. Further, multivariate logistic analyses were conducted to estimate the relative biobehavioral risk factors related to hypertension. Several significant risk factors were identified, including family history of HBP, gender, level of education, level of acculturation, regular exercise, and being overweight. Findings from this study indicate that culturally relevant approaches to prevention and treatment of HBP are urgently needed to address the HBP problem in Korean Americans. PMID- 11110354 TI - Correlation between the internalization theme of racial identity attitude survey B and systolic blood pressure. AB - OBJECTIVES: This brief report describes our first attempts at defining relationships between racial identity attitudes as measured by the Racial Identity Attitudes Survey-B (RIAS-B) and cardiovascular responses during the presentation of stressful stimuli. DESIGN: We manipulated verbal tasks to study reactivity and derived correlation coefficients to define relationships between racial identity attitudes and cardiovascular responses. METHOD: The participants (N = 17) were African-American students participating in a summer program to enhance minority careers (N = 15) or enrolled in introductory psychology. All students held a dominant Internalization theme for racial identity as assessed by the RIAS-B. After the initial baseline, the participants responded to three verbal tasks incorporating the description of a frustrating event with racial overtones (HIAFF), a description of the furniture in their house (LOAFF) and serial digit multiplication (MATH). RESULTS: The data indicate significant correlations between the Internalization score on the RIAS-B and the systolic blood pressure (SBP) for tasks as grouped by content (HIAFF r = .60; MATH r = .55) or by sequence (TASK1 r = .53, TASK2 r = .50). There were no significant correlations between Internalization score and pulse rate or diastolic blood pressure (DBP). Students had a high incidence of a positive family history for cardiovascular diseases (70%). CONCLUSIONS: The data suggest that as identity accumulation increases, so does reactivity. Internalization and the related self esteem that this theme might engender does not attenuate the cardiovascular stress response in African-American college students with a prevalence of a positive family history for cardiovascular disease. PMID- 11110355 TI - Use of major therapeutic procedures: are Hispanics treated differently than non Hispanic Whites. AB - OBJECTIVES: This study examines whether Hispanics and non-Hispanic White patients, hospitalized with one of a wide range of conditions, receive major therapeutic procedures at the same rates. METHODS: The study examined hospital stays of Hispanic and non-Hispanic White adults using 1993 discharge abstract data from the Healthcare Cost and Utilization Project State Inpatient Database for California, Florida and New York, states containing half the Hispanic population in the country. Logistic regression modeling was used to identify the effect of ethnicity on the likelihood of receiving a major therapeutic procedure separately for 63 conditions, controlling for age, gender, disease severity, health insurance, income level of patient's community, and hospital characteristics. RESULTS: Hispanics are less likely than non-Hispanic Whites to receive major therapeutic procedures for 38% of the 63 conditions examined and more likely for 6% of the conditions. CONCLUSIONS: This study identified many conditions with apparent variations in treatment based on patient ethnicity. Future studies should examine reasons for disparities between ethnic groups, why these disparities occur for some conditions and not others, and appropriateness of procedures received. PMID- 11110356 TI - Mammography screening in single older African-American women: a study of related factors. AB - OBJECTIVE: Using baseline data from an intervention study, we examined cognitive, psychological, social and medical care factors in relation to the use of a mammogram in the preceding year among single African-American women aged 65 and older. METHODS: Study subjects were 325 African-American women aged 65 and older who were divorced, widowed, separated or never-married, and lived in ten public housing complexes in Nashville, Tennessee. In-person interviews were conducted to collect information on breast screening behavior, knowledge and attitude, social network and activities, emotional and psychological symptoms and signs, and medical care use. RESULTS: Compared with those who had not had a mammogram in the preceding year, women who had had a mammogram in the preceding year were three times more likely to have a regular doctor (95% confidence interval [CI] 1.4-5.0) and about six times more likely to have a doctor's recommendation for a mammogram (95%CI 3.4-11.1). In addition, they were more likely to: (a) have attended a meeting on breast health or received educational materials on breast cancer; (b) agree that a woman needs a mammogram even though she has no breast problem; (c) agree that a woman can have breast cancer without having symptoms; (d) have living children and grandchildren; and (e) attend social activities more frequently. CONCLUSIONS: While access to regular medical care and receiving a physician's recommendation are strongly associated with mammography among these older, single African-American women, education on breast health and social networks also appear to be influential. PMID- 11110357 TI - Clinical features of spontaneous intracerebral hemorrhage in Hispanics and non Hispanic Whites in New Mexico: a community study. AB - OBJECTIVES: To compare clinical features of spontaneous intracerebral hemorrhage (ICH) between Hispanics and non-Hispanic Whites in Albuquerque, New Mexico, occurring during the year 1993. DESIGN: Analysis of data collected during a study of ICH incidence in a community. METHODS: Review medical records with multiple relevant cerebrovascular diagnostic discharge codes in all 9 acute care hospitals in Albuquerque, and in the records of the State Medical Examiner. All suspected cases were verified by cerebral computed tomography or autopsy. Vascular risk factors, test results, and acute outcome were abstracted. RESULTS: There were 38 Hispanics and 46 non-Hispanic Whites with ICH. We found no statistically significant differences between these two ethnic groups in the prevalence of hypertension or diabetes mellitus, in hematoma volume, in seasonal fluctuation of ICH incidence, or in acute mortality. However, based on our sample size, only large differences in the prevalence of risk factors between these two ethnic groups could be detected with statistical significance (>20-30 percentage points). There was a trend toward a higher proportion of subcortical ICH (centered in the basal ganglia, brainstem, or cerebellum) in Hispanics (82%) than in non-Hispanic Whites (62%, P=.09). CONCLUSIONS: The higher proportion of subcortical ICH among the Hispanics suggests that chronic hypertension may play a greater role as a risk factor for ICH in this ethnic group than in non-Hispanic Whites in New Mexico. Our findings should be confirmed by larger community studies. PMID- 11110358 TI - Race, clinical factors and pre-term birth in a low-income urban setting. AB - While infant mortality rates have declined for both White and African-American populations, the perennial two-fold excess in risk for African Americans remains unchanged, and indeed, may have increased since 1985. One potential explanation for the excess risk in African Americans might be racial differences in maternal clinical risk factors, such as prior pregnancy history and pregnancy complications. This paper examines the contributions of such clinical indicators to racial differences in pre-term delivery in a study sample of urban, low-income women, aged 18 to 43 years. METHODS: Study participants were enrolled during their first prenatal care visit at one of four hospital-based, prenatal care clinics in Baltimore City. Medical history and pregnancy outcome data were abstracted from clinical records. Multiple logistic regression models were used to assess the independent relationship between race and pre-term birth, after controlling for clinical factors. RESULTS: Without adjustment for clinical risk factors, African-American women were 1.8 times more likely than White women to have a pre-term birth outcome (95% confidence interval 1.20-2.78). After statistical adjustment for the clinical variables, however, the association between race and pre-term birth was diminished (OR = 1.64, 95% confidence interval: 0.99-2.72). Moreover, the associations between certain clinical risks and pre-term birth were stronger for African-American than White women. CONCLUSION: These results suggest that attention to clinical risk factors among African-American women may be an important avenue for reducing Black/White racial disparities in pre-term birth. PMID- 11110359 TI - Changes in US health care access in the 90s: race and income differences from the CARDIA Study. Coronary Artery Risk Development in Young Adults. AB - OBJECTIVE: Health care financing is changing rapidly in the United States. We investigated whether and how health care access is changing concurrently with changes in financing, with special attention to a minority population. METHODS: We examined a longitudinal biracial (half African-American, half White) urban cohort of 3,565 individuals, aged 25-37 years old, in 1992-93 and again in 1995 96. We measured access by self-reported (1) health insurance status, (2) regular source of medical care, and (3) lack of care due to financial problems. RESULTS: In 1992-93, 30.3% of the cohort experienced at least one access barrier, with a decline to 26.8% in 1995-96 (P<.005). However, access improved more for Whites than for African Americans; and access improved for higher, but not for lower, income groups (7% improvement for high income, vs 2% deterioration for lower income, P<.01). In addition, there was an 11% to 19% absolute increase in individuals making co-payments for health care utilization across all race/sex groups, with African Americans having markedly higher proportions of cost sharing. African-American, low income, and unemployed individuals reported more acute care, but fewer outpatient visits. Income and employment explained racial differences. CONCLUSION: While access has improved or stabilized for higher income groups, there is a widening gap according to income, accompanied by an acute care pattern for low income groups that may be both inadequate and cost inefficient. PMID- 11110360 TI - Low birth-weight infants: the continuing ethnic disparity and the interaction of biology and environment. AB - African-American infants weigh on average 200-300 grams less at birth than do European-American infants, leading to a two-fold higher rate of low birth-weight (LBW) infants. This birth weight disparity has not changed significantly over the past 95 years. Numerous research studies have been undertaken to elucidate this disparity. While various factors have been found to be associated with increased risk for having a LBW infant, including maternal anthropometrics, health and age, prenatal care, and socioeconomic status, none have been found to entirely and adequately explain the continued birth-weight differential. Researchers have concluded that there is something different in the environment and/or genetics of African-American women compared to European-American women, but are at a loss to clearly define the factor other than to say it must relate to the racism suffered by African-American women leading to more stress during pregnancy. While racism is probably an additional factor, one genetic/environmental variable, which has been overlooked, is the interaction of heavy pigmentation with degree of latitude. Heavy pigmentation blocks ultraviolet B (UVB) radiation. At high latitudes, such as in the US region, inadequate exposure to UVB radiation prevents the conversion of the prohormone to the hormonal form of vitamin D. The resulting low levels of serum vitamin D in the pregnant woman disrupt calcium homeostasis leading to intrauterine growth retardation, premature labor, and hypertension: all risk factors for LBW infants. PMID- 11110361 TI - Obesity and the African-American woman: a cultural tolerance of fatness or other neglected factors. AB - This article takes a critical look at the much-touted cultural tolerance explanation of obesity among African-American women with the hope of stimulating a re-examination of its relative merits. After reviewing evidence that runs contrary to the cultural tolerance explanation, the focus turns to an examination of some of the explanation's inherent risks and limitations. Among those discussed is that an overstated cultural tolerance explanation seems to have had a tendency of polarizing thinking in this area of public health, in that if a population group is not pre-occupied with the ideal of thinness, there has been a proclivity to assume they are indifferent about their weight. A corollary risk and limitation also discussed is that factors, which adversely affect the less extreme weight reduction behavior of African-American women but are outside the cultural realm, have tended to be neglected. The factor identified that has been most conspicuously neglected in recent years, but which can profoundly affect the weight management behavior of African-American women on many different levels, is the enduring socioeconomic disparity compared with European-American women. PMID- 11110362 TI - Radiologic differentiation between bacterial and viral lower respiratory infection in children: a systematic literature review. AB - A systematic literature review was performed to quantify the accuracy of chest radiography in differentiating bacterial from viral lower respiratory infection in children. Relevant studies were identified in a systematic literature search and were included in the review according to predetermined criteria. Five of 13 relevant identified studies met the inclusion criteria. No clinically useful degree of accuracy was demonstrated, but great caution is needed in interpreting the findings because of the suboptimal nature of the reference standards, even in included studies. It is recommended that future surveys of the microbial etiology of pneumonia that employ a credible reference standard (such as lung aspiration) be used as opportunities to perform studies of diagnostic accuracy. PMID- 11110363 TI - Altered sleeping arrangements in pediatric patients with epilepsy. AB - Parental fears concerning seizure occurrence may be associated with behavioral changes within the home environment. One possible change involves sleeping arrangements. Questionnaires concerning demographics, medical history, and sleeping arrangements were completed by parents of 179 children with epilepsy and by parents of 155 children with diabetes for comparison purposes. Based on parental response, 40 (22%) children with epilepsy changed to less independent sleeping arrangements. Logistic regression suggested that parental concern over seizure occurrence was highly associated with this change (p=<0.001). In contrast, 13 (8%) of the children with diabetes changed to a less independent sleep pattern. Results suggest changes in sleeping arrangements may alert the pediatrician to possible parental anxiety that may need to be addressed. PMID- 11110364 TI - Effects of lead counseling for children with lead levels > or = 20 microgram/dL: impact on parental knowledge, attitudes, and behavior. AB - The purpose of this study was to assess parental knowledge, attitudes, and behavior concerning lead reduction counseling. Of 108 children with confirmed venous lead levels > or = 20 microg/dL, 75 (69%) of the parents were interviewed by telephone 6-9 months later. The majority of parents recalled being given specific lead reduction strategies. Knowledge of cleaning interventions was associated with parents who could state the lead level, who perceived a benefit from knowing it was elevated, and whose children were referred to a specialty lead clinic. Recall of nutritional interventions was associated with parents who could state the lead level and whose children were referred to a specialty lead clinic. Although 79% of parents thought that it was beneficial to know their child's lead level was elevated, only 65% reported implementing lead reduction strategies. The majority of parents recalled receiving lead reduction counseling but reported low compliance with lead reduction strategies. Further research is needed to determine the causes of the discrepancy between knowledge and making the behavior changes necessary to comply with lead reduction interventions. PMID- 11110365 TI - Thyroid function in very-low-birth-weight infants with intraventricular hemorrhage. AB - The objective of this investigation was to study the natural course of thyroid function in infants with intraventricular hemorrhage (IVH). A cohort of infants < 1,500 grams birth weight, n=247, were included in the analysis. Total T4 and thyrotropin from newborn screening during the 1st week of life (Test 1) and from repeat screening at 2-4 weeks postnatal age (Test 2) were compared in infants with IVH (n=43) and a group of infants without IVH. Fifty-nine percent of infants still had transient hypothyroxinemia at the time of Test 2. After multivariate analysis, infants with IVH had an increased odds of having a T4 < or = 6 microg/dL on Test 1 (OR 2.8, 95% CI 1.2-6.5), but at the time of Test 2 IVH was not associated with an increased odds of having a low T4. Only gestational age (OR 1.6, 95% CI 1.1-2.5) remained associated with an increased odds of having an extremely low T4 (< or = 4 microg/dL) at this time. Transient hypothyroxinemia remains common at 2-4 weeks of age in preterm infants. IVH is not independently associated with having a low T4 at this time. PMID- 11110366 TI - Sex, drugs, rock 'n' roll revisited. PMID- 11110367 TI - Methylene-blue-induced hyperbilirubinemia and phototoxicity in a neonate. PMID- 11110368 TI - Early pediatric neurodevelopmental profile of children with autistic spectrum disorders. PMID- 11110369 TI - Laboratory evaluation of children with autistic spectrum disorders: a guide for primary care pediatricians. PMID- 11110370 TI - Recurrent supraventricular tachycardia as a complication of nebulized albuterol treatment. PMID- 11110371 TI - Effect of telephone follow-up on frequency of health maintenance visits among children attending free immunization clinics: a randomized, controlled trial. PMID- 11110372 TI - Replacement of the aortic root in patients with Marfan syndrome. PMID- 11110373 TI - Factors associated with age and operation for children with congenital heart disease. PMID- 11110374 TI - BSP--a retired test revisited. PMID- 11110375 TI - Evaluation of the bromosulfophthalein 30-minute retention test for the diagnosis of hepatic disease in dogs. AB - The purpose of this study was to evaluate efficacy of bromosulfophthalein (BSP) retention testing in dogs with and without histopathologically confirmed hepatobiliary disease. Medical records of 150 dogs with hepatobiliary disease having both a BSP test and hepatic biopsy were retrieved. Histopathologic slides of liver tissue were reviewed, and dogs were classified according to 1 of 11 histopathologic categories. Twenty-five clinically normal random-source dogs were used as controls for hepatic biopsy and BSP testing. No dogs suffered adverse effects due to BSP administration. BSP retention was significantly (P < .05) higher in hospitalized (13.9%) than control (3.2%) dogs, but the test could not distinguish between hospitalized dogs with different types of hepatobiliary disease. Sensitivity, specificity, and predictive values of BSP retention as a test for hepatic disease were calculated. Using 5.0% as a cutoff for normal BSP retention resulted in a specificity of 88% and a sensitivity of 76%. Using 6.0% as a cutoff for normal BSP retention resulted in a specificity of 100% and a sensitivity of 70%. Dogs of this study having BSP retention of >6% had at least an 86% chance of having an abnormal liver. We concluded that continued use of BSP retention testing is warranted as a noninvasive diagnostic test for liver disease in dogs. PMID- 11110376 TI - Passive transfer of colostral immunoglobulins in calves. AB - Passive transfer of colostral immunoglobulins has long been accepted as imperative to optimal calf health. Many factors, including timing of colostrum ingestion, the method and volume of colostrum administration, the immunoglobulin concentration of the colostrum ingested, and the age of the dam have been implicated in affecting the optimization of absorption. The practice of colostrum pooling, the breed and presence of the dam, and the presence of respiratory acidosis in the calf also may affect passive transfer. Various tests have been reported to accurately measure passive transfer status in neonatal calves. The radial immunodiffusion and the enzyme-linked immunosorbent assay (ELISA) are the only tests that directly measure serum IgG concentration. All other available tests including serum total solids by refractometry, sodium sulfite turbidity test, zinc sulfate turbidity test, serum gamma-glutamyl transferase activity, and whole blood glutaraldehyde gelation estimate serum IgG concentration based on concentration of total globulins or other proteins whose passive transfer is statistically associated with that of IgG. This paper presents a comprehensive review of the literature of passive transfer in calves including factors that affect passive transfer status, testing modalities, effects of failure of passive transfer on baseline mortality, consequences of failure of passive transfer, and some treatment options. Many previously accepted truisms regarding passive transfer in calves should be rejected based on the results of recent research. PMID- 11110377 TI - Postoperative radiotherapy for canine soft tissue sarcoma. AB - Thirty-five dogs with 37 soft tissue sarcoma tumors that were incompletely excised and treated with radiotherapy in the postoperative, adjuvant setting were reviewed. Variables evaluated included age, sex, tumor site, tumor histology, total tumor radiation dose, radiotherapy field size. time to recurrence, and survival. The majority of tumors were fibrosarcomas and hemangiopericytomas, but small numbers of other tumor types were also represented. Total tumor radiation dose ranged from 42 to 57 Gy given in 3- to 4.2-Gy daily fractions on a Monday through Friday schedule. Overall median survival was 1,851 days. Median time to local recurrence was greater than 798 days. Soft-tissue sarcoma tumors at oral sites had a statistically significant lower median survival (540 days) as compared to other tumor sites (2,270 days). Radiotherapy may be a useful adjuvant therapy for incompletely excised soft-tissue sarcomas with a reasonable expectation for long-term patient survival. PMID- 11110378 TI - Identification of matrix metalloproteinases in canine cutaneous mast cell tumors. AB - Presence of matrix metalloproteinases has been associated with tumor invasion and metastasis in human neoplasia. The presence of matrix metalloproteinase 2 and matrix metalloproteinase 9 was determined in canine mast cell tumor tissue and normal stromal tissue from 24 dogs with spontaneously occurring cutaneous mast cell tumors. Seventeen of the mast cell tumors were of histologic grade 2, and 7 were of histologic grade 3. Gelatin zymography and computer assisted densitometry image analysis were used to quantify matrix metalloproteinase concentration. Bands from canine tissues migrated in the same location as human proenzyme and active enzyme matrix metalloproteinase 2 and matrix metalloproteinase 9 standards. A semiquantitative value for each patient sample was obtained by comparing the optical assessment density of each unknown band to the optical density of the human standard. The presence of matrix metalloproteinase 2 and matrix metalloproteinase 9 in histologic grade 2 mast cell tumors and histologic grade 3 mast cell tumors was compared, as was presence of matrix metalloproteinases in tumor and stromal tissue. There was dramatically more proenzyme matrix metalloproteinase 9 activity in histologic grade 3 mast cell tumors when compared to grade 2 tumors (P = .03). There was also dramatically more active enzyme matrix metalloproteinase 2 and active enzyme matrix metalloproteinase 9 activity in tumor tissue compared to stromal tissue (P = .02, P < .0001). This study demonstrates that the proenzyme and active enzyme forms of matrix metalloproteinase 2 and matrix metalloproteinase 9 are present in canine mast cell tumors. This appears to be related to the degree of histologic malignancy, although histologic grade 1 tumors were not evaluated. PMID- 11110380 TI - Prevalence of bovine herpesvirus-4 infection in cats in Central Michigan. AB - The gammaherpesvirus bovine herpesvirus-4 (BHV-4) has been isolated from a wide variety of animals, including lions and domestic cats. Although BHV-4 antibodies have been detected in normal cats and cats with urinary disorders, the epidemiology and pathogenic role of BHV-4 in cats is unknown. The purpose of this study was to determine the prevalence of BHV-4 antibodies and viral nucleic acid in a population of free-roaming cats. Plasma and peripheral blood leukocyte samples were collected from 52 male and 52 female free-roaming cats impounded at a regional animal control facility in Central Michigan. Plasma concentrations of BHV-4 antibodies were measured with an indirect fluorescent antibody test. Peripheral blood leukocyte DNA was isolated, and a 2-stage polymerase chain reaction with heminested primers delineating a conserved portion of the BHV-4 glycoprotein B gene homologue was used to amplify BHV-4-specific DNA sequences. BHV-4 antibodies were detected in 38 (73%) male and 23 (44%) female cats. Seropositive cats were significantly more likely to be male than female (odds ratio = 3.22; P = .007). Cell-associated viremia was detected in 17 (33%) male and 11 (21%) female cats. Of the 61 seropositive cats, 23 (38%) had a detectable viremia; only 5 (12%) seronegative cats had detectable viremia. Seropositive cats were significantly more likely to be viremic than seronegative cats (OR = 4.30: P = .009). Our results suggest that BHV-4 infection may be more widespread in certain cat populations than previously reported. Furthermore, many cats seropositive for BHV-4 antibodies have a concurrent cell-associated viremia. PMID- 11110379 TI - Prognostic significance of serum alkaline phosphatase activity in canine appendicular osteosarcoma. AB - Sixty-one dogs with appendicular osteosarcoma were treated with amputation and chemotherapy of cisplatin and doxorubicin. Serum samples were obtained before and after treatment for determination of total alkaline phosphatase (TALP) activity as well as the activities of the constituent bone (BALP), liver (LALP), and corticosteroid-induced (CALP) isoenzymes. The relationship between alkaline phosphatase activities and survival was examined by Cox proportional hazards regression analysis and Kaplan-Meier log rank analysis. Mean activity of TALP, BALP, and LALP decreased significantly after treatment (P < .001). TALP and LALP activities before treatment were significantly correlated with survival (P = .006 and .001, respectively). The correlation between BALP activity before treatment and survival approached significance (P = .054). CALP activity and TALP, BALP, and LALP activities after treatment were not significantly correlated with survival. Dogs with normal pretreatment TALP and BALP activities survived significantly longer than dogs with increased pretreatment activities (P = .001 and .003, respectively). Median survival times for dogs with normal or increased TALP activities before treatment were 12.5 and 5.5 months, respectively; and median survival times for dogs with normal or increased BALP activities before treatment were 16.6 and 9.5 months, respectively. In the design of future clinical trials involving dogs with osteosarcoma, consideration should be given to stratifying the randomization according to alkaline phosphatase activity. In addition, alkaline phosphatase activity should be a factor considered by clinicians attempting to tailor the aggressiveness of adjuvant chemotherapy to the needs of individual patients or owners. PMID- 11110381 TI - The clinical and metabolic effects of rapid weight loss in obese pet cats and the influence of supplemental oral L-carnitine. AB - The efficacy, safety, and metabolic consequences of rapid weight loss in privately owned obese cats by means of a canned weight-reduction diet and the influence of orally administered L-carnitine on rate of weight loss, routine clinical evaluations, hepatic ultrasonography, plasma amino acid profiles, and carnitine analytes were evaluated. A double-blinded placebo-controlled design was used with cats randomly divided into 2 groups: Group 1 (n = 14) received L carnitine (250 mg PO q24h) in aqueous solution and group 2 (n = 10) received an identical-appearing water placebo. Median obesity (body condition scores and percentage ideal body weight) in each group was 25%. Caloric intake was restricted to 60% of maintenance energy requirements (60 kcal/kg) for targeted ideal weight. The reducing formula was readily accepted by all cats. Significant weight loss was achieved by week 18 in each group without adverse effects (group 1 = 23.7%, group 2 = 19.6%). Cats receiving carnitine lost weight at a significantly faster rate (P < .05). Significant increases in carnitine values developed in each group (P < .02). However, significantly higher concentrations of all carnitine moieties and a greater percentage of acetylcarnitine developed in cats of group 1 (P < .01). The dietary formula and described reducing strategy can safely achieve a 20% weight reduction within 18 weeks in obese cats. An aqueous solution of L-carnitine (250 mg PO q12h) was at least partially absorbed, was nontoxic, and significantly increased plasma carnitine analyte concentrations as well as rate of weight loss. PMID- 11110382 TI - Spirocerca lupi esophageal granulomas in 7 dogs: resolution after treatment with doramectin. AB - Seven dogs with Spirocerca lupi esophageal granulomas were identified based on the site of involvement (ie, distal esophagus) and characteristic endoscopic appearance. Six dogs presented with signs of esophageal disease and 1 dog was asymptomatic. Ova were only identified in the feces of 2 dogs. On thoracic radiographs, 4 dogs had evidence of a caudodorsal mediastinal mass, and 2 of these dogs had spondylitis of midthoracic vertebrae. On endoscopy, single esophageal nodules were observed in 5 dogs, 1 dog had 3 nodules, and 1 dog had 6 nodules. All 7 dogs were treated with doramectin at a dosage of 200 microg/kg SC at 14-day intervals for 3 treatments. Dogs had physical and endoscopic examinations at 2, 4, and 6 weeks after treatment. By 6 weeks, clinical signs had resolved in 6 dogs (1 dog was asymptomatic), and the esophageal nodules had completely resolved in 4 dogs, and incompletely resolved in 3 dogs. Two dogs with incomplete resolution were treated again with doramectin at 500 microg/kg PO daily for an additional 6 weeks. Complete resolution of the esophageal nodules was confirmed by endoscopy in all dogs. Nodules had resolved in 4 dogs by 6 weeks, in 2 dogs by 12 weeks (after 6 weeks additional daily oral therapy), and in 1 dog 22 months after the initial 200 microg/kg treatment regimen. No dog experienced adverse effects to the drug, and all symptomatic dogs have been free of disease for periods ranging from 3 to 4 years. PMID- 11110383 TI - Serum-effusion albumin gradient in dogs with transudative abdominal effusion. AB - A prospective clinical study in dogs with transudative abdominal effusions examined the clinical usefulness of the serum albumin-effusion albumin (SA-EA) gradient. In humans, the SA-EA gradient facilitates classification of abdominal effusion, with a gradient > or = 1.1 indicating the presence of portal hypertension. Gradient values proved useful for predicting therapeutic response to sodium restriction and diuresis in humans. Of 49 dogs evaluated, 25 had hepatobiliary disease (group 1) and 24 had other nonhepatobiliary conditions (group 2). Portal hypertension was clinically suspected in 24 of 25 dogs in group 1 and in 15 of 24 dogs in group 2. A broad range of SA-EA gradients was found. A gradient > or = 1.1 was found in 22 of 25 (88.0%) dogs with liver disease and in 14 of 24 (58.3%) dogs with other disorders. The median SA-EA gradient was higher in group 1 than in group 2, with values of 1.4 (range, 0.7-3.1) and 1.1 (range, 0.3-2.6), respectively (P < .04). Considerable overlapping of SA-EA gradients occurred between groups and among dogs with diverse conditions such that gradient values could not distinguish dogs with hepatobiliary disease from dogs with other conditions. The overall diagnostic accuracy of the SA-EA gradient in predicting portal hypertension in dogs with and without hepatobiliary disease (69.4%) exceeded that of hypoalbuminemia (57.1%). These findings suggest that portal hypertension is a predominant force in formation of transudative abdominal effusion in dogs with hepatobiliary disease and in dogs with other disorders. Whether the SA-EA gradient can be used to guide therapeutic mobilization of effusion in dogs remains to be proved. PMID- 11110384 TI - Idiopathic hypercalcemia in cats. AB - Unexplained hypercalcemia has been increasingly recognized in cats since 1990. In some instances, hypercalcemia has been associated with calcium oxalate urolithiasis, and some affected cats have been fed acidifying diets. We studied the laboratory findings, clinical course, and treatment of 20 cats with idiopathic hypercalcemia. Eight (40%) of the cats were longhaired and all 14 cats for which adequate dietary history was available had been fed acidifying diets. Clinical signs included vomiting (6 cats), weight loss (4 cats), dysuria (4 cats), anorexia (3 cats), and inappropriate urinations (3 cats). Hypercalcemia was mild to moderate in severity. and serum parathyroid hormone concentrations were normal or low. Serum concentrations of phosphorus, parathyroid hormone related peptide, 25-hydroxycholecalciferol, and calcitriol were within the reference range in most cats. Diseases commonly associated with hypercalcemia (eg, neoplasia, primary hyperparathyroidism) were not identified despite thorough medical evaluations and long-term clinical follow-up. Azotemia either did not develop (10 cats) or developed after the onset of hypercalcemia (3 cats), suggesting that renal failure was not the cause of hypercalcemia in affected cats. Seven of 20 cats (35%) had urolithiasis, and in 2 cats uroliths were composed of calcium oxalate. Subtotal parathyroidectomy in 2 cats and dietary modification in 11 cats did not result in resolution of hypercalcemia. Treatment with prednisone resulted in complete resolution of hypercalcemia in 4 cats. PMID- 11110385 TI - Serum feline trypsin-like immunoreactivity in cats with exocrine pancreatic insufficiency. AB - Exocrine pancreatic insufficiency is thought to occur rarely in cats. This assumption has been made based on the lack of a specific test for this disease in the cat. Clinical data from the 1st 20 cats with serum feline trypsin-like immunoreactivity (fTLI) concentrations < or = 8 microg/L are presented. In 17 of these 20 cats compelling evidence for a diagnosis of exocrine pancreatic insufficiency (EPI) was present and in the remaining 3 supportive evidence for a diagnosis of EPI was available. The conclusion was made that serum fTLI concentration is a specific test for EPI in the cat. PMID- 11110386 TI - Surgical and medical treatment of an Arabian filly with proliferative enteropathy caused by Lawsonia intracellularis. PMID- 11110388 TI - deLahunta honored with Kirk Award. Winner known for his legendary work ethic. PMID- 11110389 TI - Gene therapy for cancer. AB - The basic strategies for gene therapy that have been explored include immunogene therapy such as cytokine gene transfer, selective prodrug activation, so-called suicide genes, transfer of a tumor suppressor gene, and inhibition of activated oncogenes by antisense mechanisms. Many therapeutic protocols have so far been registered in the office of the Recombinant DNA Advisory Committee, NIH, as of June 1999. Regarding the basic research in gene therapy, we planned a clinical application of gene therapy for an esophageal cancer patient at our institution. We review herein the present status of gene therapy for cancer overseas, and describe a protocol of clinical trials for gene therapy for esophageal cancer at our institution. PMID- 11110390 TI - Videothoracoscopic surgery for thoracic neurogenic tumors: a 7-year experience. AB - While videothoracoscopic surgery has rapidly become accepted as an effective method of performing minimally invasive surgery, the advantages and feasibility of using this surgical technique for the treatment of neurogenic tumors of the thorax are not yet well defined. Between August 1992 and May 1999, 15 solitary thoracic neurogenic tumors were surgically excised using videothoracoscopic surgery in our hospital. The patients comprised six women and nine men, with a mean age of 38.1 years. The mean tumor size was 3.5 cm, with a range of 1.5-6.5 cm and included 12 schwannomas, 2 ganglioneuromas, and 1 neurofibroma. Among the 15 patients, 4 were treated using videothoracoscopic surgery plus minithoracotomy. The only complication associated with videothoracoscopic surgery was hoarseness which developed in one patient. Our experience indicates that videothoracoscopic surgery is a useful alternative to facilitate the excision of small thoracic neurogenic tumors. PMID- 11110391 TI - The significance of surgery for bulky N2 small-cell lung cancer: a clinical and in vitro analysis of long-term survivors. AB - Until recently, stage III small-cell lung cancer (SCLC) has not been considered an indication for surgical treatment, however, stable subculture has become possible in about 80% of the patients with SCLC in our hospital. Therefore, we have been performing surgery for all patients with cStage I-III resectable tumors and giving combined-modality therapy based on the sensitivity assay in vitro since April 1982. In the present study, we reviewed 30 consecutive patients with cStage III and pStage III SCLC, including 27 with N2 disease who underwent surgery between 1982 and 1992, 7 of whom (23%) survived disease-free for over 5 years. We examined the cell characteristics in vitro and the actual treatment of five patients with bulky N2 lesions, four of whom were long-term survivors and one of whom died 11 months after surgery, as controls for comparison with the long-term survivors. Based on the results of the cell characteristics in vitro, SCLC was determined to be a heterogeneous tumor. Thus, surgical excision of the primary tumor with adjuvant therapy might be necessary to achieve long-term survival and maintain good performance status, and to improve quality of life in patients with bulky N2 SCLC by eliminating drug-resistant tumor cells within the primary tumor. PMID- 11110392 TI - Deterioration of visceral perfusion caused by intra-abdominal hypertension in pigs ventilated with positive end-expiratory pressure. AB - Experimental studies and clinical experience suggest that the combination of positive end-expiratory pressure (PEEP) ventilation and intra-abdominal hypertension might alter splanchnic hemodynamics to a significantly greater degree than the effect of either of them alone. Therefore, we assessed the intestinal and hepatic hemodynamics in two steps of PEEP ventilation, adding tense pneumoperitoneum in a pig model. The hepatic artery, portal vein, and superior mesenteric artery blood flow, as well as the hepatic and intestinal mucosal microcirculation, and the hepatic pO2 and intestinal mucosal pH, were assessed before, then with 5 cmH2O and 10 cmH2O PEEP alone, and in combination with a 12-mmHg pneumoperitoneum, in ten domestic pigs. Statistical analysis of the hepatic and intestinal measurements revealed a significant decrease (P = 0.001) in all parameters in relation to the baseline, during the 5-cmH2O and 10 mmH2O PEEP ventilation period. The addition of 12 mmHg intra-abdominal pressure led to an extreme deterioration in all parameters (P = 0.001), in relation to both the baseline and the 10-cmH2O PEEP measurement. These findings demonstrate that PEEP and intra-abdominal hypertension act cumulatively on the abdominal viscera, producing conditions of extremely low hypoperfusion and ischemia. PMID- 11110393 TI - Three-dimensional simulation of coronary artery bypass grafting with the use of computational fluid dynamics. AB - In search of an optimal anastomosis conformation in coronary artery bypass grafting surgery and flow visualization, three-dimensional simulation of the anastomosis has been developed with the use of computational fluid dynamics. To simulate the surgery, a Y-figure model with proximal stenosis was developed in three cases according to angles ranging from 10 degrees to 30 degrees. The boundary condition of velocity and flow of the model were given based on the average velocity and flow of coronary artery measured intraoperatively. Using this information, the fluid dynamics of three models were calculated by commercial computation fluid dynamics code. The convergent results showed the least recirculating jet in the 10 degrees anastomosis without back flow into the graft. The total energy loss in the field was most affected in 30 degrees anastomosis. The wall shear stress profile showed maximum values at the heel region where recirculating jet takes place. This simulation suggests that the more acute is the angle of anastomosis, the smaller is the energy loss. Based on this phenomenon, surgery should therefore be designed to obtain as acute an angle as possible in light of the fluid dynamics. PMID- 11110394 TI - Injury to the gastric mucosa and cellular dynamics in a rat model of duodenogastric reflux: the possible significance of gastrin induction and a heat shock protein. AB - Injury to the gastric mucosa caused by duodenogastric reflux (DGR) is often encountered after gastrectomy or truncal vagotomy (V) with pyloroplasty. This study was designed to investigate the histological features of the gastric mucosa under such conditions. A rat model of DGR and DGR+V was established and the thickness of the oxyntic mucosa was measured. Cellular dynamics in the presence of injury to the gastric mucosa caused by DGR were investigated by the immunohistochemical staining of bromodeoxyuridine (BrdU) and heat shock protein 70 (HSP70). The relationship between persistent hypergastrinemia and mucosal injury was also studied. Duodenogastric reflux activated the intracellular induction of HSP70 in our rat model of DGR. Hypergastrinemia was noted in the V group. Compared with values from the DGR group, the numbers of BrdU-labeled cells increased, the glandular proliferation zone expanded, and the thickness of the oxyntic mucosa was significantly higher in the DGR+V group. Compared with the DGR group, there was greater induction of HSP in the DGR+V group during the acute stage. This finding suggests that denervation of the gastric mucosa and hypergastrinemia after vagotomy may be associated with the expression of HSP. PMID- 11110395 TI - Thyroid metastasis from gastric carcinoma: report of a case. AB - Thyroid metastasis from gastric carcinoma is rare. In this report, we describe the case of a patient in whom a rapidly growing thyroid metastasis was found 3 months after an operation for gastric carcinoma. PMID- 11110396 TI - Persistent hypocalcemia with elevated parathyroid hormone levels after long-term primary hyperparathyroidism: report of a case. AB - We report herein the case of a 48-year-old man with long-term persistent primary hyperparathyroidism (pHPT) despite undergoing a parathyroidectomy in 1976, followed by a reoperation in 1978, for whom resection of a parathyroid adenoma in the upper mediastinum was eventually performed. His postoperative course was complicated by recurrent hypocalcemia refractory to oral calcium substitution and significantly elevated levels of parathyroid hormone (PTH). The radiological findings are presented, and we discuss the possible reasons for the coincidence of severe hypocalcemia with increased PTH levels in association with pHPT. PMID- 11110397 TI - Neurilemmoma arising in the brachial plexus in association with breast cancer: report of case. AB - A 48-year-old woman without von Recklinghausen's disease presented to our department for investigation of a left breast lump and a lump in the left axilla. A presumptive diagnosis of left breast cancer with axillary lymph node involvement was made based on the findings of physical examination and needle biopsy of the left breast lump. However, on exploration we discovered that the axillary lump was a neurogenic tumor arising in the brachial plexus. Enucleation of the neurogenic tumor, which was subsequently histologically confirmed as neurilemmoma, as well as a modified radical mastectomy, was performed. Postoperatively, the patient experienced slight numbness in her left second and third fingers, but this symptom improved within 5 months. When last seen 21 months after her operation, there were no signs of recurrence of either the breast cancer or the neurilemmoma. PMID- 11110398 TI - Cervico-mediastinal bronchogenic cyst occurring in the prenatal period: report of a case. AB - We experienced a case of cervico-mediastinal bronchogenic cyst in which a cervical cystic mass was detected by prenatal ultrasonography. On prenatal ultrasound, a unilocular, well-defined and hypoechoic mass was detected in the fetal neck. The baby was born by a normal vaginal delivery at 40 weeks of gestation, and had no respiratory distress. Radiological investigations demonstrated a cyst in the cervico-mediastinal region, which displaced the trachea to the left. At the age of 32 days, an elective resection was easily performed through a right inferior collar incision after first aspirating the contents of the cyst. Prenatal sonography showing abnormal findings is effective for identifying cysts in the perinatal period and allows for the timely resection of such cysts before respiratory distress occurs. PMID- 11110399 TI - Video-assisted thoracoscopic management of an anterior mediastinal teratoma: report of a case. AB - Video-assisted thoracoscopic surgery (VATS) is a newly developed technique, the advantages of which mainly benefit the patient. Its feasibility and indications have been described widely, but to the best of our knowledge no report of the successful VATS management of a large mediastinal teratoma has ever been documented. When such a tumor is encountered, conversion to thoracotomy would usually be carried out. We report our experience of removing a large teratoma, 8 x 7 x 11 cm in size, from the anterior mediastinum, employing VATS and utilizing only four small intercostal incisions without spreading the ribs. PMID- 11110400 TI - Aortic dissections complicating open cardiac surgery: report of three cases. AB - Between June 1991 and February 1999, three patients suffered ascending aortic dissection as a complication of cardiopulmonary bypass operations with aortic cannulation at our hospital. The dissection occurred during the operation in two of the three patients and several months after the operation in one. Among a total of 2207 cardiac operations performed during this period, the incidence of perioperative ascending aortic dissection was 0.14%. In addition to visual inspection and palpation, either epicardial or transesophageal echocardiography proved extremely useful for establishing an intraoperative diagnosis of ascending aortic dissection as a complication of open cardiac operation. One of the three patients underwent closed plication but subsequently died of vital organ ischemia. In this case, failure of reapproximation of the injured intima by closed plication might have led to extension of the dissection. Despite prolonged cardiopulmonary bypass and myocardial ischemic time, graft replacement of the ascending aorta was successfully carried out in the other two patients. Thus, we believe that graft replacement of the ascending aorta should be performed for patients with extensive aortic dissection complicating an open cardiac operation. PMID- 11110401 TI - Cystic adventitial disease of the popliteal artery: elongation into the media of the popliteal artery and communication with the knee joint capsule: report of a case. AB - Cystic disease of the popliteal artery is a rare disorder in which most cases involve the formation of an adventitial cyst that disturbs the popliteal artery blood flow. We present herein the case of a patient presenting with popliteal artery occlusion due to compression by a cyst which formed at the media of the popliteal artery. The onset occurred during a baseball game in which he played catcher. Preoperative magnetic resonance imaging demonstrated a communication of the cyst with the adjacent knee joint. This unusual case could provide important clues to help identify the pathogenesis of this disease. PMID- 11110402 TI - Primary gastric lymphoma with spontaneous perforation: report of a case. AB - Primary gastric lymphoma with spontaneous perforation is rare. We report herein the case of a 53-year-old-man who was admitted to our hospital with severe epigastralgia. Emergency endoscopic examination showed a perforated gastric tumor in the lower body of the greater curvature, and a distal subtotal gastrectomy with lymph node dissection was performed. The resected tumor measured 10.0 x 8.0 cm and was associated with an area of ulceration, 8.0 x 6.0 cm in size, and perforation, 1.0 x 0.5 cm in size. Pathological examination confirmed a diagnosis of B-cell malignant lymphoma of the diffuse, medium-sized cell type. According to the Ann Arbor and Naqvi classifications, the lymphoma was stage II and stage III, respectively. Postoperative adjuvant chemotherapy comprising cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP) was given. The patient is currently well with no signs of recurrence, 2 years 4 months after his operation. PMID- 11110403 TI - Nonoperative treatment of reperforated duodenal ulcer: report of three cases. AB - During the period between 1983 and 1998, a total of 58 patients were admitted to the surgical department of Akita Red Cross Hospital with acute duodenal perforation. Of these 58 patients, 16 were treated operatively and 42 were treated nonoperatively. Among the 38 men and 4 women who received nonoperative treatment, 3 developed reperforation. The incidence of reperforation was 7.1% and the mean average interval from the initial treatment until reperforation was 3.5 years. Endoscopic biopsy and/or serum anti-H. pylori IgG measurement revealed Helicobacter pylori infection in all three patients. No serious complications developed during the nonoperative treatment of reperforation in these three patients, and their recovery was uneventful. The hospital stay ranged from 10 to 18 days, with a mean stay of 12 days after the first perforation and from 14 to 18 days, with a mean stay of 15.6 days after the reperforation. Nonoperative treatment proved successful as a life-saving procedure for reperforation of a duodenal ulcer in all three patients. PMID- 11110404 TI - Long-term survival achieved by resection of metastases in the liver and lung in a patient with recurrent colonic cancer: report of a case. AB - A 58-year-old man who underwent a potentially curative resection of cancer of the sigmoid colon at another hospital was subsequently followed up at our hospital. A lateral segmentectomy was performed for a solitary hepatic metastasis, and partial resection of right S1 was later carried out for a pulmonary metastasis. Another pulmonary metastasis was found 6 years after his third operation and to minimize the area to be resected, bronchial arterial infusion chemotherapy was performed twice. A 51% reduction in the size of the tumor was achieved, so a right upper lobectomy and wedge resection of the bronchus were performed. The patient remains alive 14 years after the initial resection of colonic cancer. This case is considered noteworthy because it demonstrates the potential effectiveness of local adjuvant chemotherapy and the possibility of extended survival in a patient who has undergone resection of both hepatic and pulmonary metastases from colonic cancer. PMID- 11110405 TI - Successful treatment of recurrent liver metastases from gastric cancer by repeated hepatic resections: report of a case. AB - We describe herein the case of a patient in whom recurrent liver metastases from gastric cancer were successfully treated by performing repeated hepatic resections. A 63-year-old man underwent a total gastrectomy with regional lymph node dissection for an advanced gastric cancer on November 17, 1992, the pathological findings of which confirmed a diagnosis of well-differentiated tubular adenocarcinoma, ss, INFalpha, ly1, v0, n1(+). Follow-up computer tomography (CT) and ultrasonography scans done 7 months after the gastrectomy revealed a metastasis in the liver S5, and a partial resection of S5 was performed on July 5, 1993. Subsequently, on November 17, 1994, an anterior segmentectomy of the liver was performed for a liver metastasis in the liver S8, then on August 11, 1998, a partial resection of the liver S6 was performed for a metastasis in the liver S6. The pathological findings of each liver specimen resected were compatible with metastatic adenocarcinoma from the primary gastric cancer. The liver tumors were expansive-growing tumors with capsules and massive necrosis. The patient is currently well with no evidence of recurrence on repeat CT scans, 6 years 6 months since-the initial gastrectomy, and 5 years 10 months since the first hepatic resection. PMID- 11110406 TI - Intra-abdominal gallstone spillage detected during umbilical trocar site hernia repair after laparoscopic cholecystectomy: report of a case. AB - It is well known that spilled gallstones may occur as a complication during laparoscopic cholecystectomy. We present herein the case of a patient in whom intra-abdominal gallstone spillage was found during repair of an incisional hernia at the umbilical trocar port site 3 months after a laparoscopic cholecystectomy. We describe this case and present a review of the recent literature to stress the importance of preventing stone spillage and retrieving any stones that are spilled into the abdomen. PMID- 11110407 TI - Rapid progression of intrahepatic cholangiocarcinoma after drainage of large cystic lesions: report of a case. AB - We report herein a case of cholangiocarcinoma with large cystic lesions. Computed tomography (CT) demonstrated large cysts in segment IV and the paracaval portion of the caudate lobe, and a solid tumor in the anterior segment of the right lobe of the liver which was contiguous to the cyst in the paracaval portion of the caudate lobe. The large cysts were diagnostically misleading and a liver abscess was suspected. Thus, percutaneous transhepatic drainage of the cyst was performed. The fluid in the cyst was negative bacteriologically, but malignant cells were detected. A CT scan done 2 weeks after drainage of the cyst showed progression of the solid tumor with intrahepatic metastasis and replacement of the cystic lesions by the solid tumor. Following percutaneous transhepatic portal embolization, a right hepatic trisegmentectomy with caudate lobectomy was performed. Pathological examination confirmed cholangiocellular carcinoma. These results indicate that drainage of the cystic lesion induced the tumor progression in the liver. Therefore, the possibility of cholangiocarcinoma with a large cystic lesion should be borne in mind when considering the differential diagnosis of a cystic lesion in the liver, and appropriate surgical therapy should be carefully selected. PMID- 11110408 TI - A novel navel presenting as an umbilical polyp and urachal sinus associated with an epigastric cleft: a clue to anterior midline fusion defects. AB - Umbilical and periumbilical disorders may present with a diverse group of anomalies and reflect the developmental embryological events they result from. A rare occurrence in a newborn of an umbilicus with an umbilical polyp together with an urachal sinus associated with a supraumbilical abnormal skin area known as epigastric cleft is reported herein, to help to elucidate embryological steps of anterior midline fusion defects and urachal remnants. PMID- 11110409 TI - Mediastinal irrigation with superoxidized water after open-heart surgery: the safety and pitfalls of cardiovascular surgical application. AB - Postoperative mediastinal infection after open-heart surgery via median sternotomy is a devastating complication. In this paper, we describe a simple method of irrigating the mediastinum using superoxidized water to prevent perioperative contamination. After mediastinal hemostasis was done, warm superoxidized water of more than 21 was uninterruptedly irrigated for 5 min immediately prior to sternal closure. We have used this method in 25 patients undergoing cardiac surgery, and noted significant perioperative electrocardiographic changes, including ST elevation, without hemodynamic compromise in 15 of these patients. PMID- 11110410 TI - Treatment of severe hypercholesterolemia in apolipoprotein E-deficient mice by intramuscular injection of plasmid DNA. AB - We report on systemic delivery and long-term biological effects of apolipoprotein E (apoE) obtained by intramuscular (i.m.) plasmid DNA injection. ApoE plays an important role in lipoprotein catabolism and apoE knock-out mice develop severe hypercholesterolemia and diffuse atherosclerosis. We have injected apoE-deficient mice with 80 microg of a plasmid vector (pCMV-E3) encoding the human apoE3 cDNA under the control of the CMV promoter-enhancer in both posterior legs. Local expression of the transgene was demonstrated throughout 16 weeks. Human apoE3 recombinant protein reached 0.6 ng/ml serum level. After i.m. injection of pCMV E3 expression vector the mean serum cholesterol concentrations decreased from 439 +/- 57 mg/dl to 253 +/- 99 mg/dl (P < 0.05) 2 weeks after injection and persisted at a significantly reduced level throughout the 16 weeks observation period (P < 0.005). Serum cholesterol was unaffected and reached an absolute level of 636 +/- 67 mg/dl in control groups. Finally, injection of pCMV-E3 into apoE-deficient mice resulted in a redistribution of cholesterol content between lipoprotein fractions, with a marked decrease in VLDL, IDL and LDL cholesterol content and an increase in HDL cholesterol. These results demonstrate that severe hypercholesterolemia in apoE-deficient mice can be effectively reversed by i.m. DNA injection, and indicate that this approach could represent a useful tool to correct several hyperlipidemic conditions resulting in atherosclerosis. PMID- 11110411 TI - Glucose-stimulated and self-limiting insulin production by glucose 6-phosphatase promoter driven insulin expression in hepatoma cells. AB - The liver is an attractive target organ for insulin gene expression in type 1 diabetes as it contains appropriate cellular mechanisms of regulated gene expression in response to blood glucose and insulin. We hypothesize that insulin production regulated by both glucose and insulin may be achieved using the promoter of the glucose 6-phosphatase gene (G6Pase), the expression of which in the liver is induced by glucose and suppressed by insulin. Recombinant adenoviral vectors expressing the reporter gene CAT or insulin under transcriptional direction of the G6Pase promoter were constructed. Glucose-stimulated as well as self-limiting insulin production was achieved in vector-transduced hepatoma cells in which expression of the insulin gene was controlled by the G6Pase promoter. While insulin strongly inhibited the G6Pase promoter activity under low glucose conditions, its inhibitory capacity was attenuated when glucose levels were elevated. At the physiologic glucose level of 5.5 mM glucose, vector-transduced hepatoma cells produced a self-limited level of insulin at approximately 0.2-0.3 ng/ml, which is within the range of fasting levels of insulin in normal animals. These results indicate that the G6Pase promoter possesses desirable features and may be developed for regulated hepatic insulin gene expression in type 1 diabetes. PMID- 11110412 TI - Airway gene transfer in mouse nasal-airways: importance of identification of epithelial type for assessment of gene transfer. AB - Mouse nasal airways are often used for the assessment of both reporter and cystic fibrosis transmembrane conductance regulator (CFTR) gene transfer to respiratory epithelia. However, the mouse nasal cavity is lined by both olfactory (OE) and respiratory epithelium (RE). Previous gene transfer studies have suggested that OE may be more efficiently transduced by adenoviral vectors than RE. However, to provide data pertinent to CFTR gene transfer in humans, measurements of CFTR function in mice by transepithelial potential difference (TPD) should be directed towards respiratory rather than olfactory epithelium. We report a new technique to mark the position of the TPD sensing cannula tip in the mouse nasal cavity that permitted us to correlate TPD measurements with epithelial cell type. Using this technique, we found TPD values did not discriminate between respiratory and olfactory epithelia. We next assessed relationships between anatomic regions accessed by the TPD cannula and epithelial type. The frequently used insertion depth of approximately 5 mm from the nose tip predominantly recorded the TPD from anterior dorsal olfactory epithelium. Measurement of the TPD of respiratory epithelium in our study was maximized by insertion of the TPD cannula probe to 2.5 mm depth. Because TPD measurements are not sensitive to epithelial type, adequate control of position and TPD catheter insertion depth are required to ensure accurate estimation of CFTR gene transfer into the target RE in the mouse nasal cavity. PMID- 11110413 TI - A preclinical model of hepatocyte gene transfer: the in vivo, in situ perfused rat liver. AB - Delivering retroviruses targeted to hepatocytes in vivo involves the injection of retroviruses directly into the portal vein. The aim of this work was to establish a clinically relevant system for retrovirus-mediated gene transfer in a new model of in vivo, in situ perfused rat liver and to study the transgene expression. At 24 h after partial hepatectomy, the liver was completely excluded from the splanchnic circulation using an extracorporeal shunt. Two independent normothermal, oxygenated perfusion systems were used. First, liver perfusion was carried out with a recirculating system (1 h). Culture supernatant containing retroviruses (1.5 x 10(8) ffu/ml, beta-galactosidase gene) was used as perfusate. Then the liver perfusion was maintained for more 30 min in a single liver passage system using culture medium without retroviruses as perfusate. High hepatocyte transduction rates (up to 34.4%) were obtained. PCR analysis showed no provirus in extrahepatic organs. Viral titrations performed simultaneously (inflow and outflow liver lines) showed that after 1 h of perfusion (up to 30 successive liver passages) retroviruses were still detected in the liver outflow perfusate (up to 2.0 x 10(7) ffu/ml). Washing the liver for 30 min dramatically decreased the leakage of retroviruses in the outflow. In order to be of clinical use, the injection of retroviruses targeted to hepatocytes in vivo should be done while the liver is completely excluded from the splanchnic circulation to avoid any extrahepatic retrovirus diffusion. PMID- 11110414 TI - A blood-tumor barrier limits gene transfer to experimental liver cancer: the effect of vasoactive compounds. AB - We have evaluated gene transfer efficiency to tumor nodules in diethylnitrosoamine (DENA)-induced hepatocellular carcinoma (HCC) in rats using adenoviral vectors administered by three different routes: intraportal, intra arterial and intratumoral injection. Our results showed that intraportal infusion could not transduce tumor nodules greater than 1 mm in diameter while the intra arterial route allowed transduction of nodules up to 2-5 mm in diameter. Tumors greater than this size were resistant to transduction by intravascular route, but could be transduced by direct intratumoral injection, indicating that the obstacle preventing gene transfer to tumor cells was mainly at the level of tumor vasculature and not at the level of neoplastic cells. We have studied the extracellular matrix in tumoral lesions to assess whether nodules with different size and histological pattern have different profiles in relation to transduction efficacy. Immunohistochemical detection showed a high expression of fibronectin (FN), laminin (LN) and alpha-smooth muscle actin (alpha-SMA) in those large HCC, which were resistant to adenoviral infection. Intra-arterial infusion of vasoactive compounds (histamine, angiotensin II or nitric oxide donor nitroglycerin) before vector administration enhanced gene transfer to tumor nodules that were poorly transduced without pre-treatment. Nitroglycerin was active to enhance transduction of large tumors with trabecular or pseudoglandular histological pattern, which were impermeable to adenoviral vectors even after histamine or angiotensin treatments. Our data indicate the presence of a physical barrier between blood and neoplastic cells, which prevents transduction of the tumor by vectors given by the intravascular route. The thickness and impermeability of the barrier increases as the tumor nodule grows. Vasoactive compounds may be of value in gene therapy of liver cancer by increasing transduction efficiency by intravascularly administered vectors. PMID- 11110415 TI - Ultrasound enhancement of cationic lipid-mediated gene transfer to primary tumors following systemic administration. AB - The impact of a localized application of ultrasound on gene transfer to primary tumors following systemic administration of cationic lipid based transfection complexes was investigated. We have previously shown that systemic administration of DOTMA (N-[(1-(2-3-dioleyloxy) propyl)]-N-N-N-trimethylammonium chloride):cholesterol-based transfection complexes to tumor-bearing mice resulted in expression in the tumor and other tissues, primarily the lungs. Application of ultrasound to the tumor before or after the injection resulted in a significant increase in gene transfer to the tumor with no increase observed in other tissues. The magnitude of increased expression ranged from three- to 270-fold depending upon the DNA dose. The following parameters were optimized for maximal increase: duration of ultrasound application, the time interval between plasmid injection and sonoporation, and plasmid dose. A combination of plasmid quantitation and fluorescence microscopy showed that ultrasound increased tumor uptake of the plasmid and that uptake was limited to the tumor vasculature. Using an IL- 12 expression plasmid, the combination of a single plasmid dose (10 microg) and ultrasound treatment produced significantly higher levels of IL-12 in tumor. This increased expression was sufficient to inhibit tumor growth compared with the control conditions. These data demonstrate the potential application of sonoporation as an effective method for enhancing the expression of systemically administered genes in tumor endothelium for cancer gene therapy. PMID- 11110416 TI - Immunotherapy of spontaneous type 1 diabetes in nonobese diabetic mice by systemic interleukin-4 treatment employing adenovirus vector-mediated gene transfer. AB - We have previously shown that systemic injection of multiple low doses of recombinant murine interleukin-4 (mIL-4) can prevent type 1 diabetes (T1D) in nonobese diabetic (NOD) mice by activating regulatory T helper (Th) 2 cells in vivo. Here, we have developed a gene transfer approach to the prevention of T1D by testing the therapeutic potential of an adenovirus gene transfer vector engineered to express mIL-4. We found that only two systemic injections of a recombinant adenovirus type 5 vector-expressing mIL-4 (Ad5mIL-4) reduces destructive insulitis and protects NOD mice from the onset of diabetes by eliciting intrapancreatic Th2 cell responses. Host immune responses against the adenovirus vector were detectable; however, the levels of antibody production were insufficient to preclude Ad5mIL-4 treatment as a possible therapeutic agent against T1D. Thus, adenovirus-mediated delivery of IL-4 provides protection of NOD mice from T1D and represents a clinically viable therapeutic approach. PMID- 11110417 TI - Decoy against nuclear factor-kappa B attenuates myocardial cell infiltration and arterial neointimal formation in murine cardiac allografts. AB - Acute rejection and graft arteriopathy in cardiac transplantation limit the long term survival of recipients; these processes are enhanced by several cytokines and adhesion molecules. Nuclear factor-kappa B (NFkappaB) is critical in the transcription of multiple genes involved in inflammation and cell proliferation. To test the hypothesis that NFkappaB decoy can attenuate acute rejection and arteriopathy, we performed single intraluminal delivery of NFkappaB decoy into murine cardiac allografts using a hemagglutinating virus of Japan (HVJ) artificial viral envelope (AVE)-liposome method. No decoy or scrambled decoy transfer was performed for control. Hearts were heterotopically transplanted from BALB/c to C3H/He mice (major mismatch group) and from DBA/2 to B10.D2 mice (minor mismatch group). Nontreated or scrambled decoy transfected allografts of the major mismatch group were acutely rejected, while NFkappaB decoy prolonged their survival. While severe cell infiltration and intimal thickening with enhancement of inflammatory factors were observed in untreated or scrambled decoy-treated allografts of minor mismatch group at day 28, NFkappaB decoy attenuated these changes. We conclude that NFkappaB is critically involved in the development of acute as well as chronic rejection of the transplanted hearts. NFkappaB decoy attenuates both acute rejection and graft arteriopathy by blocking the activation of several genes. PMID- 11110418 TI - Immune response induced by retrovirus-mediated HSV-tk/GCV pharmacogene therapy in patients with glioblastoma multiforme. AB - This study was conducted to investigate immunological components of somatic gene therapy for primary glioblastoma multiforme (GBM) in adults. It involved 13 patients treated by surgical resection of tumor with subsequent radiation therapy. Seven of them received additional herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) gene therapy by direct intracerebral injection of retrovirus (RV) vector producing cells (VPC) during tumor surgery and subsequent systemic administration of GCV. Peripheral blood for FACS immunophenotyping, isolation of peripheral mononuclear cells (PMNC), and serum ELISA assays for IL 12 and soluble Fas ligand (sFasL) was collected daily during the first 4 post operative weeks. Tumor specimens were obtained at primary or recurrent surgery and at autopsy. Tumors from gene therapy patients showed varying degrees of peritumoral necrosis around the former tumor resection cavity. Numbers of tumor infiltrating lymphocytes found weeks after gene therapy were not significantly increased compared with primary tumors. Mitotic tumor cells were sparse close to the VPC injection sites, but abundant in brain areas somewhat distant from these sites. Serum ELISA revealed significantly increased sFasL and IL-12 levels in the gene therapy group compared with controls. Immunophenotyping of PMNC did not show a significant activation of T cells or NK cells during gene therapy. Interferon gamma secretion was evaluated by ELISPOT assays employing PMNC cocultivated with autologous tumor cells. It demonstrated an antitumor immune response in the gene therapy group, but not in the control group. These findings support the concept of in vivo induction of a systemic immune response by local intracerebral HSV tk/GCV pharmacogene therapy for primary human GBM. PMID- 11110419 TI - Genetic immunization against cervical carcinoma: induction of cytotoxic T lymphocyte activity with a recombinant alphavirus vector expressing human papillomavirus type 16 E6 and E7. AB - Infection of genital epithelial cells with human papillomavirus (HPV) types 16 and 18 is closely associated with the development of cervical carcinoma. The transforming potential of these high-risk HPVs depends on the expression of the E6 and E7 early viral gene products. Since the expression of E6 and E7 is selectively maintained in premalignant and malignant cervical lesions these proteins are attractive candidates for immunotherapeutic and prophylactic strategies. This report describes the construction, characterization and the in vivo immunotherapeutic potential of recombinant Semliki Forest virus (SFV) expressing the HPV16 E6 and E7 proteins (SFV-E6E7). Western blot analysis and immunofluorescence staining demonstrated expression of E6 and E7 in BHK cells infected with SFV-E6E7. Immunization of mice with SFV-E6E7 resulted in an efficient in vivo priming of HPV-specific CTL activity. The induced CTL lysed murine tumor cells transformed with the HPV16 genome and EL4 cells loaded with an immunodominant class I-binding HPV E7 peptide. CTLs could reproducibly be induced by immunization with three injections of as few as 10(5) infectious units of SFV E6E7. Protection from tumor challenge was studied using the tumor cell line TC-1. Immunization with 5 x 10(6) SFV-E6E7 particles protected 40% of the mice from tumor challenge. These results indicate that E6E7 expression by the efficient and safe recombinant SFV system represents a promising strategy for immunotherapy or immunoprophylaxis of cervical carcinoma. PMID- 11110420 TI - Stabilized plasmid-lipid particles for systemic gene therapy. AB - The structure of 'stabilized plasmid-lipid particles' (SPLP) and their properties as systemic gene therapy vectors has been investigated. We show that SPLP can be visualized employing cryo-electron microscopy to be homogeneous particles of diameter 72 +/- 5 nm consisting of a lipid bilayer surrounding a core of plasmid DNA. It is also shown that SPLP exhibit long circulation lifetimes (circulation half-life >6 h) following intravenous (i.v.) injection in a murine tumor model resulting in accumulation of up to 3% of the total injected dose and concomitant reporter gene expression at a distal (hind flank) tumor site. In contrast, i v. injection of naked plasmid DNA or plasmid DNA-cationic liposome complexes did not result in significant plasmid delivery to the tumor site or gene expression at that site. Furthermore, it is shown that high doses of SPLP corresponding to 175 microg plasmid per mouse are nontoxic as assayed by monitoring serum enzyme levels, whereas i.v. injection of complexes give rise to significant toxicity at dose levels above 20 microg plasmid per mouse. It is concluded that SPLP exhibit properties consistent with potential utility as a nontoxic systemic gene therapy vector. PMID- 11110421 TI - Retroviral vector design studies toward hematopoietic stem cell gene therapy for mucopolysaccharidosis type I. AB - To optimize a gene transfer system for hematopoietic stem cell gene therapy of patients with mucopolysaccharidosis (MPS) type I, 10 retroviral vectors were constructed to express the human alpha-L-iduronidase (IDUA) cDNA. These vectors were designed to evaluate the potential effects of specific promoters, the addition of selectable markers, and the use of multiple promoters versus an internal ribosome entry site for expression of IDUA and selectable maker genes. The effect of vector design was investigated in primary patient fibroblasts (F(MPS)) or murine fibroblast cell lines; while overall comparison of transgene expression was determined in patients' peripheral blood lymphocytes (PBL(MPS)) and CD34+ progenitors (PBPC(MPS)). We observed that the human PGK promoter introduced the highest IDUA activity per 1% relative transgene frequency in F(MPS). Use of the same promoter to separately regulate both the therapeutic gene and a drug-resistance gene resulted in decreased expression of the unselected gene. Co-selection using bicistronic vectors not only increased the number of transductants, but also elevated transgene expression under selective pressure in transgene-positive progenitors. Bicistronic vector LP1CD overcame down-regulation and practically introduced the highest IDUA level in unselected PBL(MPS) and an intermediate level in PBPC(MPS). These studies provide a better understanding of factors contributing to efficient gene expression in hematopoietic cells. PMID- 11110422 TI - Effects of physical training on the recovery of the autonomic nervous activity during exercise after coronary artery bypass grafting: effects of physical training after CABG. AB - Analysis of heart rate variability (HRV) can identify patients at risk of sudden cardiac death after myocardial infarction. The present study examined the effect of 2 weeks of supervised aerobic exercise training on the recovery of the autonomic nervous activity, exercise capacity, and cardiac output (CO) after coronary artery bypass grafting (CABG). Twenty-eight patients were randomly divided into the training group or the control group and performed exercise tests at 1 week, 3 weeks, 3 months, 6 months and 1 year after CABG. The HRV was measured, and the high-frequency component of HRV was used as an index of parasympathetic nerve activity (PNA); the plasma norepinephrine concentration (NE) was used as an index of sympathetic nervous activity. Cardiac output was also measured. In the training group, peak VO2, peak CO and PNA during exercise had improved at 3 weeks, but there was no improvement in these indices in the control group. NE decreased 1 week after CABG in both groups. These results indicate that physical training soon after CABG improves not only the exercise capacity, but also PNA. PMID- 11110423 TI - Efficacy of mitral valve replacement for patients with mitral regurgitation and a dilated left ventricle. AB - Mitral regurgitation is a significant complication of end-stage cardiomyopathy, and its existence predicts poor survival. In general, it is thought that mitral valve replacement (MVR) alone is ineffective; however, there are few detailed reports of the clinical course of patients who have undergone MVR. Five patients with mitral regurgitation whose preoperative left ventricular end-systolic volume index was more than 100 ml/m2 were studied. Although their prognosis late after MVR became poor, none of them died within 30 days of the operation. Postoperative cardiac catheterization was performed 6.3+/-1.1 months after surgery; the end diastolic volume had reduced (before: 193+/-26 ml/m2; after: 166+/-34 ml/m2, p<0.05), but the end-systolic volume had not (before: 110+/-7 ml/m2; after: 112+/ 32 ml/m2). The end-systolic wall stress was substantially elevated preoperatively (238+/-29 kdyne/cm2) and tended to increase after surgery (295+/-96 kdyne/cm2). All the patients were able to return to work at some stage postoperatively (their New York Heart Association functional class improved to I or II), but 3 of the 5 patients died suddenly of heart failure at 3.3+/-1.6 years after surgery and the New York Heart Association functional class of the others worsened to III again. Mitral valve surgery, including MVR, can manage severe end-stage heart disease with mitral regurgitation. PMID- 11110424 TI - Possible role of chronic infection with Chlamydia pneumoniae in Japanese patients with acute myocardial infarction. AB - Chlamydia pneumoniae, a common human respiratory pathogen, has been implicated in the pathogenesis of coronary heart diseases (CHD) in several seroepidemiological studies. The present case-control study investigated the relation between serologic evidence of C. pneumoniae infection and CHD in a Japanese population. Two groups of cases were enrolled: 26 patients with acute myocardial infarction (AMI) and 46 patients with effort angina pectoris (e-AP). Their data were compared with 58 age-matched healthy controls and also compared with 53 patients with vasospastic angina (VSA) as pathological control subjects. Anti-C. pneumoniae specific IgA and IgG antibody titers were measured by enzyme-linked immunosorbent assay (ELISA). The mean indices of IgG-type antibody in AMI and e AP were not significantly different from those in either the normal controls or VSA group. On the other hand, the mean indices of IgA-type antibody in AMI were significantly higher than in the normal controls (1.39+/-0.83 in AMI vs 0.84+/ 0.58 in controls, p<0.001) and VSA (1.39+/-0.83 in AMI vs 1.05+/-0.61 in VSA, p<0.05) group. However, the differences in the IgA titers in the e-AP group compared with the normal controls did not reach a significant level. The odds ratio associated with the seropositivity of IgA for AMI against the normal controls was 3.89 (95% confidence interval (CI): 1.16-13.10) and that against VSA was 6.90 (95% CI: 1.73-27.52) after adjustment for risk factors for CHD and/or age, sex and smoking status. In 6 patients the elevated IgA titers were sustained even at 3 months after the episode of AMI. These results suggest that seropositivity for IgA-type antibody against C. pneumoniae may be a significant risk factor for the development of AMI. The possible mechanisms include chronic inflammation in the coronary artery due to persistent C. pneumoniae infection. PMID- 11110425 TI - Risk factors that discriminate 'high- risk' from 'low-risk' Japanese patients with coronary artery disease. AB - Although various risk factors have been implicated in the progression of coronary artery disease (CAD), coronary risk factors specifically related to the long-term prognosis for high-risk CAD have not been determined. The study enrolled 311 consecutive Japanese patients with CAD who underwent diagnostic coronary arteriography and divided them into 2 groups: (i) 135 high-risk patients with either impaired left ventricular function (ejection fraction <50%) or multivessel disease and (ii) 176 low-risk patients with normal left ventricular function and 0- or 1-vessel disease. The prevalence of risk factors including age, gender, smoking, hypertension, diabetes mellitus (DM), obesity and lipid variables were compared between the 2 groups. The prevalence of DM, a serum high-density lipoprotein (HDL)-cholesterol level below 35 mg/dl and a serum lipoprotein (Lp) (a) level above 25 mg/dl was significantly higher in the high-risk group as compared with the low-risk group. Multiple logistic regression analysis demonstrated that DM (odds ratio (OR): 1.72, 95% confidence intervals (CI): 1.02 2.92, p<0.05), a low HDL-cholesterol level (OR: 2.49, 95% CI: 1.49-4.17, p<0.001) and a high Lp(a) level (OR: 1.68, 95% CI: 1.02-2.76, p<0.05) were all independent risk factors for high-risk CAD. However, if the patients with 0-vessel disease were excluded from the low-risk group, a low HDL-cholesterol level was found to be the only independent predictor for high-risk CAD (OR: 2.07, 95% CI: 1.15-3.70, p<0.05). Among both men and smokers in this population, a higher Lp(a) level was found to be a significant predictor for high-risk CAD. A low serum level of HDL cholesterol, a high serum level of Lp(a) and DM were significant predictors of high-risk in patients with CAD. Among patients with a significant coronary stenosis or left ventricular dysfunction, a low serum level of HDL-cholesterol was the only significant predictor for high-risk CAD. PMID- 11110426 TI - Increased plasma antigen levels of monocyte chemoattractant protein-1 in patients with restenosis after percutaneous transluminal coronary angioplasty. AB - Monocyte chemoattractant protein-1 (MCP-1) plays an important role in the progression of atherosclerosis in coronary arteries. To examine whether or not plasma antigen levels of MCP-1 are related to restenosis after percutaneous transluminal coronary angioplasty (PTCA), the plasma antigen levels of MCP-1 were measured by enzyme-linked immunosorbent assay (pg/ml) before, 24 and 48 h, and 3 months after elective PTCA for stable exertional angina performed between June 1997 and March 1998. Restenosis was defined as recurrence of stenosis greater than 50% of the diameter in the dilated segment at 3-month follow-up angiography. There were no differences in plasma MCP-1 antigen levels before and at 24 h after PTCA between restenosis (R; n=27) and no-restenosis (N; n=43) groups (R vs N: 633+/-35 vs 589+/-34, and 669+/-41 vs 575+/-36 pg/ml before and at 24 h after PTCA, respectively), but plasma MCP-1 antigen levels were higher at 48 h and 3 months after PTCA in the R than in N group (R vs N: 678+/-41 vs 558+/-35, and 735+/-35 vs 571+/-32 pg/ml at 48 h and 3 months after PTCA, respectively). These data suggest that the MCP-1 production and macrophage accumulation in the balloon injured site is partially associated with restenosis after PTCA. PMID- 11110427 TI - Ventricular arrhythmias with left bundle branch block pattern and inferior axis: assessment of their mechanisms on the basis of response to ATP, nicorandil and verapamil. AB - The present study investigated the mechanism of ventricular arrhythmias showing left bundle branch block (LBBB) pattern with an inferior axis. The effects of 3 drugs, adenosine triphosphate (ATP), nicorandil and verapamil, were evaluated in 17 patients. ATP suppressed the arrhythmias in 14 patients and nicorandil suppressed them in 8 of those 14. Verapamil suppressed 5 of the 6 ATP-nicorandil sensitive arrhythmias. Four patients with ATP- or nicorandil-sensitive arrhythmias were not sensitive to verapamil. On the other hand, 3 of the ATP insensitive arrhythmias were sensitive to neither nicorandil nor verapamil. The QT intervals and QTc were shortened by nicorandil in 5 of the 6 patients who were sensitive to all 3 drugs. One mechanism of suppression by nicorandil could be related to less Ca++ entering the myocardium, which would decrease the duration of the action potential as indicated by the shortened QT intervals. The results suggest that the mechanism of some ventricular arrhythmias is related to triggered activity. Arrhythmias that are sensitive to ATP or nicorandil, but not to verapamil, may be caused by abnormal automaticity. On the other hand, arrhythmias that are insensitive to all 3 drugs might be related to reentry. The features of ventricular arrhythmias with LBBB pattern and inferior axis differ and therefore the causative mechanisms are not the same. PMID- 11110428 TI - Descending aorta wall volume and coronary artery disease: a comparative study using enhanced computed tomography of the chest and coronary angiography. AB - The study examined the association between aortic wall volume (AWV) detected by enhanced computed tomography and coronary artery atherosclerosis observed on angiography. In 180 cases, AWV was measured as the total wall volume of a 7-cm portion of the descending thoracic aorta distal from the tracheal bifurcation. Coronary artery atherosclerosis was angiographically quantified by both Gensini score, in terms of the severity of coronary artery stenosis, and Extent score, in terms of the severity of coronary artery involvement. Mean AWV values between the patients with significant coronary artery stenosis and those without significant stenosis were 9.83+/-4.04 cm3 and 8.09+/-2.39 cm3, respectively (p<0.001). AWV was a significantly independent variable for significant coronary artery disease (p=0.0097) and an Extent score > or = 60 (p=0.0092). Calcification of AWV, however, was not associated with coronary atherosclerosis. The quantification of aortic atherosclerosis was useful for diagnosing coronary artery disease. PMID- 11110429 TI - Effect of long-term exercise training on blood viscosity during endurance exercise at an anaerobic threshold intensity. AB - Blood viscosity (etaB) is low in athletes, but the effect of exercise training on etaB during endurance exercise at an anaerobic threshold (AT) intensity in non athletes is not well known, although it is known that exercise training sometimes induces the hyperviscosity syndrome. Fourteen subjects were recruited and divided into 2 groups: those who trained at an AT intensity for 30 min/day, 3 times weekly for 1 year (Group T, n=8), and sedentary subjects (Group C, n=6). The test protocol consisted of a single 30-min treadmill exercise at each individual's AT intensity, which was determined in advance. The etaB, plasma viscosity (etaP), and hematocrit were measured just before and at the end of the treadmill exercise. The subjects were not allowed to drink any water before exercise. In the Group C subjects, the hematocrit and etaP increased significantly and the etaB tended to increase. However, in the Group T subjects, the hematocrit and etaP did not increase and the etaB decreased significantly. These data indicate that long-term exercise training attenuates the increase in blood viscosity during exercise. PMID- 11110430 TI - Impaired heart rate response during incremental exercise in patients with acute myocardial infarction and after coronary artery bypass grafting: evaluation of coefficients with Karvonen's formula. AB - Heart rate (HR) response during exercise in patients with ischemic heart disease was evaluated, and the appropriateness of Karvonen's method for determining rehabilitation exercise target HR was investigated. The study group comprised 24 patients with acute myocardial infarction (AMI) and 37 patients who had undergone coronary artery bypass grafting (CABG). Cardiopulmonary exercise testing (CPX) was performed with a cycle ergometer and changes in HR (deltaHR)/changes in work rate (deltaWR) and interval changes of the coefficient of Karvonen's formula were evaluated. In the AMI group and the CABG group, deltaHR/deltaWR were significantly lower than those of age-matched control subjects (p<0.01). Karvonen's coefficients ranged from 0.37 to 0.54 when calculated from actual peak HR and 0.21 to 0.32 calculated from the predicted peak HR. An impaired HR response was found in patients with AMI and those who had had CABG up to 6 months previously. Because the Karvonen's coefficient values, which ranged from 0.6 to 0.8, were elevated for these patients, and considering the data from the CPX, increased exercise is recommended for such cases. PMID- 11110431 TI - Relationship between plasma oxidized low-density lipoprotein and the coronary vasomotor response to acetylcholine in patients with coronary artery disease. AB - The present study examined the relation of plasma oxidized low-density lipoprotein (LDL) levels to plasma LDL cholesterol levels and the impairment of endothelium-dependent coronary vasorelaxation in patients with coronary artery disease (CAD). In the first study, the relationship between plasma levels of oxidized LDL and LDL cholesterol were investigated in 88 patients with CAD. In the second study, the changes in the diameter of the left anterior descending (LAD) and the left circumflex (LCX) coronary arteries were measured after intracoronary administration of acetylcholine (15 microg) and isosorbide dinitrate (2.5 mg) in 15 patients with CAD. Plasma oxidized LDL levels were determined with a sandwich enzyme-linked immunosorbent assay. Plasma oxidized LDL levels did not correlate with plasma LDL cholesterol levels (r=-0.03, p=NS). The % diameter changes (mean+/-SEM) in the LAD and LCX after intracoronary acetylcholine were -8.3+/-3.5% and -10+/-4.2%, respectively. The % diameter changes in the LAD and LCX after intracoronary isosorbide dinitrate were 23+/ 4.8% and 23+/-5.1%, respectively. The % diameter changes in the LAD and LCX inversely correlated with plasma oxidized LDL levels after intracoronary acetylcholine (LAD: r=-0.55, p=0.03; LCX: r=-0.59, p=0.02), but were not after intracoronary isosorbide dinitrate. Plasma LDL cholesterol, triglyceride, and high-density lipoprotein cholesterol levels did not correlate with the coronary vasoreaction to acetylcholine. In conclusion, plasma oxidized LDL levels do not correlate with plasma LDL-cholesterol levels and are related to impairment of endothelium-dependent coronary vasodilation in patients with CAD. PMID- 11110432 TI - Alterations in the inotropic responses to forskolin and Ca2+ and reduced gene expressions of Ca2+-signaling proteins induced by chronic volume overload in rabbits. AB - Volume overload results in eccentric cardiac hypertrophy, but it is still unknown how this mechanical overload modulates the inotropic response to exogenous Ca2+ or adenylyl cyclase stimulation. Inotropic responsiveness in vivo and the levels of gene expression of Ca2+ signaling proteins were studied in rabbit hearts hypertrophied as a result of volume overload at 4 and 12 weeks after arteriovenous shunt formation. In sham-operated control rabbits, left ventricular (LV)+dP/dt was augmented in response to graded doses of CaCl2. Dose-related changes of LV+dP/dt to CaCl2 were attenuated significantly in shunt rabbits with volume overload. Forskolin dose-dependently augmented LV+dP/dt in sham rabbits, which was also attenuated significantly in rabbits with volume overload. The mRNA levels of dihydropyridine receptor, Na+/Ca2+ exchanger, sarcoplasmic reticulum Ca2+-ATPase, and ryanodine receptor decreased significantly at 4 and 12 weeks in the volume-overload rabbits compared with the sham rabbits, but the mRNA levels of phospholamban and calsequestrin remained unchanged. Chronic volume overload alters contractile responsiveness to Ca2+ or adenylyl cyclase stimulation, and downregulation of steady state mRNA levels of Ca2+ signaling proteins might be, at least in part, related to this pathologic process. PMID- 11110433 TI - Effect of diadenosine tetraphosphate (AP4A) on coronary arterial microvessels in the beating canine heart. AB - Diadenosine tetraphosphate (AP4A) can be released from activated platelets and the present study examined its effect on coronary arterial microvessels. The role of purinoceptors in the coronary microcirculation in vivo was also investigated. In open chest dogs, coronary arterioles were observed using a microscope with a floating objective. In Protocol 1, AP4A (1, 10, 100 and 1,000 micromol/L) was superfused onto the heart surface before and during the superfusion of 10 micromol/L of 8-phenyltheophylline (8-PT), a P1 purinoceptor blocker. In Protocol 2, AP4A (0.1, 1, 10, and 100 nmol x kg(-1) x min(-1)) was infused into the left anterior descending coronary artery before and during the superfusion of 10 micromol/L of 8-PT. In addition to 8-PT, 30 micromol/L of pyridoxalphosphate-6 azophenyl 2',4'-disulphonic acid (PPADS), a P2X purinoceptor blocker in Protocol 3, or 300 micromol/L of N(omega)-nitro-L-arginine (LNNA) in Protocol 4, was continuously superfused, and 4 doses of AP4A were cumulatively superfused as in Protocol 1. In Protocol 5, 10 micromol/L of alpha,beta-methylene ATP, an agonist of P2X purinoceptors, was superfused for 60 min. Superfused AP4A dilated arterioles in a dose-dependent manner. The magnitude of dilatation was greater in smaller arterioles (small vessel < or = 150 microm: 24.5+/-2.2% vs large vessel > 150 microm: 10.6+/-1.5% at a dose of 1,000 micromol/L, p<0.001). On the other hand, intraluminally applied AP4A also dilated arterioles, but no size dependency was shown. In the presence of 8-PT, vasodilatory responses to superfused and intraluminally applied AP4A were attenuated and the lower doses of AP4A constricted arterioles. This vasoconstrictor effect was not affected by PPADS. The vasodilatory effect of the higher doses of AP4A was almost abolished in the presence of LNNA. Alpha,beta-methylene ATP had no effect on coronary microvascular diameters. AP4A has bidirectional effects on coronary arterial microvessels: vasodilatory effects mediated by P1 purinoceptors and NO, which might be mediated by P2Y purinoceptors, and a vasoconstrictor effect, which is not mediated by P2X purinoceptors. PMID- 11110434 TI - Quantitative evaluation of a directly depolarized area induced by high-output pacing on the cardiac muscle. AB - Quantitative information is needed on the directly depolarized area (DDA) induced by high-output energy during a precise mapping procedure to detect the origin of a tachycardia. In the present study, a DDA caused by high-output energy was quantitatively evaluated in the exposed canine heart. In 8 dogs, the right atrial and ventricular surfaces were exposed through a right thoracotomy and pacing with various outputs was delivered from the epicardial surface. A comb-shaped 16 polar electrode array and/or a 224 polar mat electrode array were used for recording the epicardial electrograms. The local activation time was measured at each electrode site, and the relationship of the distance between the electrode location from the pacing site and the local activation time was plotted and fitted to a primary regression line. The intercept of the regression line on the horizontal axis was defined as the radius of the 'DDA' and this was evaluated at each pacing output. The radius of the DDA was 0.6+/-0.1 mm with a 2 V and 3.8+/ 0.2 mm with a 10 V output when it was evaluated in a direction perpendicular to the fiber orientation of the pectinate muscle, 0.8+/-0.1 mm with a 2 V and 4.1+/ 0.3 mm with a 10 V output in a direction parallel to the pectinate muscle fiber orientation, and 0.9+/-0.3 mm with a 2 V and 3.6+/-0.5 mm with a 10 V output in the right ventricle. The DDA extended according to the increase in stimulation outputs at all sites, and there was no significant difference in the pacing site or the direction of the stimulation propagation. The DDA caused by high-output energy is a purely physical phenomenon that depends only on stimulation output and tissue resistance. The diameter of the DDA exceeded 4 mm (ie, the size of a standard tip electrode for catheter ablation) when pacing was delivered with an output greater than 6 V. PMID- 11110435 TI - Limited balloon expansion through the struts of the Palmaz-Schatz and NIR stents compared with the Multi-Link stent. AB - After placing a stent in the main vessel of a bifurcation lesion, it is sometimes necessary to perform further balloon inflation in order to treat an ostial lesion in a side branch. The stent struts may prevent full balloon expansion at the ostium of a side branch, resulting in residual ostial stenosis. The degree of completeness of balloon inflation may vary significantly depending on the stent design and structure. A model of a bifurcation lesion with an angle of 45 degrees was created from acrylic resin. The diameters of the main vessel and the side branch were both 3.5 mm. Deployment of the Palmaz-Schatz stent (n=5), NIR stent (n=5) or Multi-Link stent (n=5) was performed in the main vessel with a 3.5-mm balloon catheter inflated to 12 atm. A 3.5-mm balloon catheter was then inflated to 12 atm through the stent struts of the main vessel and into the ostium of the side branch. The degree of completeness of balloon inflation (% balloon expansion) was calculated as (smallest diameter of balloon catheter/reference diameter of balloon catheter) x 100%. The minimal lumen diameter (MLD) and cross sectional area (CSA) at the ostium of the side branch created with the stent struts were also measured. Limited balloon expansion through the struts was observed with the Palmaz-Schatz stent and the NIR stent, but almost full balloon expansion was observed with the Multi-Link stent (% balloon expansion: Palmaz Schatz stent 80%, NIR stent 60%, Multi-Link stent 94%, p<0.01). The MLD and CSA of the dilated struts, representing the ostium of the side branch, of the Palmaz Schatz stent (2.2+/-0.1 mm, 4.5+/-0.3 mm2) and the NIR stent (1.8+/-0.1 mm, 3.1+/ 0.3 mm2) were significantly smaller compared with those of the Multi-Link stent (3.0+/-0.2 mm, 8.4+/-0.6 mm2) (p<0.01). The struts of the Palmaz-Schatz stent and the NIR stent deployed in the main vessel of a bifurcation prevent full expansion of a balloon catheter inflated at the side branch ostium. In contrast, almost full balloon expansion through the struts of the Multi-Link stent is achieved. PMID- 11110436 TI - A peculiar form of focal atrial tachycardia mimicking atypical atrial flutter. AB - A 55-year-old man was referred because of congestive heart failure and atrial flutter. A 12-lead electrocardiogram (ECG) showed positive P waves in leads II, III, and aVF with a continuously undulating pattern that lacked an isoelectric baseline. Tachycardia was diagnosed as atypical atrial flutter based on classical criteria. An electrophysiological study and catheter ablation using an electroanatomical system revealed the mechanism of the tachycardia to be focal atrial tachycardia originating from the left atrial roof. This case indicates that focal atrial tachycardia may present as atypical atrial flutter on the surface ECG. PMID- 11110437 TI - Metastatic cardiac papillary carcinoma originating from the thyroid in both ventricles with a mobile right ventricular pedunculated tumor. AB - A 62-year-old man with a history of surgical therapy for papillary thyroid carcinoma was admitted because of chest pain, dyspnea on effort, pretibial edema, and slight fever. An electrocardiogram showed ST segment elevation in the precordial leads and low voltage in the limb leads. A large solid mass was demonstrated in both ventricles, with pericardial effusion, by echocardiography, thoracic computed tomography scan, transesophageal echocardiography, and angiography. A punch biopsy of the tumor revealed metastatic papillary carcinoma. During radiation therapy, the patient suddenly died of ventricular fibrillation. At autopsy, the tumor occupied almost the entire right ventricular cavity, expanding toward the main trunk of the pulmonary artery with a mobile peduncle and it had infiltrated the left ventricular wall through the interventricular septum. Microscopic examination confirmed metastatic papillary thyroid carcinoma. Only 2 other cases of cardiac metastases of papillary thyroid carcinoma have been reported and this case is the first report of metastases in both ventricles with a mobile right ventricular pedunculated tumor. PMID- 11110438 TI - Cardiogenic shock triggered by verapamil and atenolol: a case report of therapeutic experience with intravenous calcium. AB - Cardiogenic shock developed in a 72-year-old Japanese woman during combination therapy with verapamil and atenolol for recurrent supraventricular arrhythmia. She had coronary atherosclerosis, liver cirrhosis and bradycardia-tachycardia syndrome. Despite of the high-dose catecholamines and counterpulsation, she progressively deteriorated. Bolus administration of intravenous calcium chloride (CaCl2) immediately resolved her hemodynamic collapse. PMID- 11110440 TI - Qualitative blood flow differentiation: depiction of a left to right cardiac shunt across a ventricular septal defect using electron-beam computed tomography. AB - Three-dimensional imaging using electron-beam computed tomography (EBCT) has been used to assess static anatomical information in heart disease. With volume rendering, differences in objects can be distinguished through selection of the shape of opacity and color curves for CT values. If there is a difference between the CT values for arterial and venous blood, differences in opacity and color between them can be set. In a newborn baby with a left to right cardiac shunt across the ventricular septal defect (VSD), EBCT could depict arterial blood crossing the VSD into the right ventricle. PMID- 11110439 TI - Successful management of intractable coronary spasm with a coronary stent. AB - Although the long-term survival of patients suffering from coronary spasm is usually excellent, serious complications can develop, such as disabling pain, myocardial infarction, ventricular tachyarrhythmias, atrioventricular block and sudden cardiac death. A 40-year-old man who had intractable chest pain from coronary artery spasm suffered ventricular fibrillation and an acute anterior myocardial infarction upon first admission. The patient underwent a coronary angiogram, which revealed a spontaneous focal spasm at the proximal left anterior descending coronary artery (LAD). He was treated by the combination of nitrate and calcium channel blocker, but continued to complain of severe chest pain despite intensive medical therapy and he had to be treated in the emergency room 5 times during an 8-month follow-up period. An ergonovine coronary angiogram was performed and an intracoronary ultrasound examination, which revealed a focal spasm at the same site of the proximal LAD with a small amount of localized eccentric atheromatous plaque. A coronary artery stent was placed in the proximal LAD and his symptoms resolved. A follow-up coronary angiogram was performed 3 years after stenting and the stent remained patent without any in-stent restenosis or spasm. PMID- 11110441 TI - Agreement opens doors to practice in Great Britain. PMID- 11110442 TI - Lawsuit settlement draws ire of research community: congress delays agreement to regulate research on rats, mice, birds. PMID- 11110443 TI - Providing guidance on antimicrobial use. PMID- 11110444 TI - A look at la medicina veterinaria Cubana. PMID- 11110445 TI - Veterinary Medical Assistance Teams rise and shine. PMID- 11110446 TI - Review of the accreditation process for Western University of Health Sciences. PMID- 11110447 TI - Golden rules to nurture nephrologic logic. PMID- 11110448 TI - What is your diagnosis? Severe pleural effusion and pulmonary atelectasis. PMID- 11110449 TI - Anesthesia case of the month. PMID- 11110450 TI - One-step placement of a percutaneous nonendoscopic low-profile gastrostomy port in cats. PMID- 11110451 TI - Some thoughts on veterinary medicine and surgery practice acts. PMID- 11110452 TI - Analysis of seroconversion to porcine circovirus 2 among veterinarians from the United States and Canada. PMID- 11110453 TI - Rabies preexposure vaccination among veterinarians and at-risk staff. AB - OBJECTIVE: To measure rabies preexposure vaccination rate and identify factors potentially associated with lack of vaccination among veterinarians and at-risk staff. DESIGN: Cross-sectional survey. STUDY POPULATION: At-risk veterinary medical association (VMA) members, their staff members, and animal shelter and wildlife rehabilitation center personnel located in a California county. PROCEDURE: A questionnaire was mailed to VMA members and managers of animal shelters and wildlife rehabilitation centers. Respondents were requested to provide data on vaccination history and potential factors associated with vaccination status for themselves and their at-risk staff members. Vaccinated and unvaccinated individuals were compared by use of univariate and logistic regression analyses to identify factors associated with vaccination status. RESULTS: Fifty-eight percent (79/137) of persons who received questionnaires responded; 74 were eligible for the study. Respondents provided data for 47.6% (219/460) of their staff members. The vaccination rate was greater among respondents (85.1 %) than among their staff members (17.5%). Among staff members, age and duration of employment were significantly associated with vaccination status. CONCLUSIONS AND CLINICAL RELEVANCE: A large proportion of at-risk staff members working in veterinary clinics, animal shelters, and wildlife rehabilitation centers in the study area did not receive rabies preexposure vaccination per the Centers for Disease Control and Prevention's published recommendations of the Advisory Committee on Immunization Practices (ACIP). The cost of the preexposure vaccine series may be a barrier, particularly for young employees who are commonly short-term, part-time, or volunteer workers. Efforts are needed to increase awareness of the ACIP recommendations and to increase access to vaccination through agencies such as public health clinics. PMID- 11110454 TI - Survey of anesthesia techniques and concerns in private veterinary practice. PMID- 11110455 TI - Long-term outcome of gonadectomy performed at an early age or traditional age in cats. AB - OBJECTIVE: To determine long-term results and complications of gonadectomy performed at an early age (prepubertal) or at the traditional age in cats. DESIGN: Cohort study. ANIMALS: 263 cats from animal shelters. PROCEDURE: Cats that underwent gonadectomy were allotted to 2 groups on the basis of estimated age at surgery (traditional age, > or = 24 weeks old; prepubertal, < 24 weeks old). Adoptive owner information was obtained from shelter records, and telephone interviews were conducted with owners to determine physical or behavioral problems observed in the cats after adoption. Follow-up information was obtained from attending veterinarians for cats with complex problems or when owners were uncertain regarding the exact nature of their cat's problem. RESULTS: Compared with traditional-age gonadectomy, prepubertal gonadectomy did not result in an increased incidence of infectious disease, behavioral problems, or problems associated with any body system during a median follow-up period of 37 months. Additionally, the rate of retention in the original adoptive household was the same for cats that underwent prepubertal gonadectomy as those that underwent traditional-age gonadectomy. CONCLUSIONS AND CLINICAL RELEVANCE: Prepubertal gonadectomy may be performed safely in cats without concern for increased incidence of physical or behavioral problems for at least a 3-year period after gonadectomy. PMID- 11110456 TI - Evaluation of efficacy of selamectin and fipronil against Ctenocephalides felis in cats. AB - OBJECTIVE: To evaluate efficacy of monthly administration of selamectin and fipronil against Ctenocephalides felis in cats. DESIGN: Randomized controlled trial. ANIMALS: 36 healthy cats. PROCEDURE: Cats known to be free of fleas were infested with 100 unfed adult fleas on days -28 and -21. On days 0, 30, 60, 90, and 120, sixteen cats (8 pairs/treatment group) were treated by topical administration of selamectin (6 mg/kg [2.7 mg/lb] of body weight) or fipronil (7.5 mg/kg [3.4 mg/lb]). Four control cats (2 pairs) were not treated. On day -6 and every 2 weeks after initial treatment, comb counts were performed to detect fleas. Flea counts were recorded, and fleas (< or =50) that had been removed were replaced onto the cat. On day 89, fleas were not replaced. On day 91 and every 7 days until the end of the study (day 150), cats were challenged with 20 adult fleas. Flea counts were compared between and within treatments. RESULTS: 14 days after treatment, geometric mean flea counts were reduced by 71.2% by fipronil treatment and 35.3% by selamectin treatment. Both treatments resulted in 97 to 98% reduction in flea counts on day 29 and 99.8 to 100% reduction from day 44 to the end of the study. CONCLUSIONS AND CLINICAL RELEVANCE: Selamectin is as effective as fipronil in treating infestation in cats housed for 3 months in a flea-infested environment under conditions known to support the flea life cycle and in protecting against subsequent weekly challenges with C felis for an additional 2 months. PMID- 11110457 TI - Evaluation of efficacy of selamectin, fipronil, and imidacloprid against Ctenocephalides felis in dogs. AB - OBJECTIVE: To evaluate efficacy of monthly administration of selamectin, fipronil, and imidacloprid against Ctenocephalides felis in dogs. DESIGN: Randomized controlled trial. ANIMALS: 44 healthy dogs. PROCEDURE: Dogs known to be free of fleas were infested with 100 unfed adult fleas on days -28 and -21. On days 0, 30, 60, 90, and 120, dogs (12/group) were treated by topical administration of selamectin (6 mg/kg [2.7 mg/lb] of body weight), fipronil (7.5 mg/kg [3.4 mg/lb]), or imidacloprid (10 mg/kg [4.5 mg/lb]); 8 untreated dogs were used as controls. On day -6 and every 2 weeks after initial treatment, comb counts of viable adult fleas were made, and fleas (< or =50/dog) were replaced onto the dog from which they were removed. On day 89, fleas were not replaced. On day 91 and every 7 days until the end of the study, dogs were challenged with 20 adult fleas. RESULTS: 14 days after initial treatment, geometric mean flea counts were reduced by 97.5 to 99.1 % for all treatments, compared with pretreatment counts on day -6. Selamectin, fipronil, and imidacloprid reduced geometric mean flea counts by 99.7 to 100% from day 29 to the end of the study. CONCLUSIONS AND CLINICAL RELEVANCE: Selamectin is as effective as fipronil and imidacloprid in reducing C felis infestation in dogs housed for 3 months in a flea-infested environment under conditions known to support the flea life cycle, and in protecting against subsequent weekly challenges with C felis for an additional 2 months. PMID- 11110458 TI - Effect of citrate concentration on coagulation test results in dogs. AB - OBJECTIVE: To determine the effect of citrate concentration (3.2 vs 3.8%) on coagulation tests in dogs. DESIGN: Original study. ANIMALS: 30 clinically healthy dogs and 12 dogs with hereditary hemostatic disorders. PROCEDURE: Blood was collected from all dogs directly into collection tubes containing 3.2 or 3.8% buffered citrate. Prothrombin time (PT), activated partial thromboplastin time (aPTT), and fibrinogen concentration were measured by use of 3 clot-detection assay systems (2 mechanical and 1 photo-optic). Factor VIII and factor IX coagulant activities (FVIII:C and FIX:C, respectively) were determined by use of a manual tilt-tube method and a mechanical clot-detection device. RESULTS: Significant differences were not detected in median PT, fibrinogen concentration, FVIII:C, or FIX:C between 3.2 and 3.8% citrate for any assay system. A significant prolongation in aPTT for 3.2% citrate, compared with 3.8% citrate, was found in 1 mechanical system. CONCLUSIONS AND CLINICAL RELEVANCE: Citrate concentration does not significantly affect results of most coagulation assays, regardless of assay system. The aPTT was mildly influenced by the citrate concentration, although this was animal-, instrument-, and reagent-dependent. The choice of 3.2 or 3.8% citrate as an anticoagulant for coagulation tests has minimal influence on assay results in healthy dogs or dogs with hereditary hemostatic disorders. PMID- 11110459 TI - Evaluation of the effect of limited food consumption on radiographic evidence of osteoarthritis in dogs. AB - OBJECTIVE: To determine prevalence of radiographic evidence of osteoarthritis in 4 diarthrodial joints of dogs with restricted feed intake, compared with dogs without restricted feed intake. DESIGN: Paired feeding study. ANIMALS: 48 Labrador Retrievers. PROCEDURE: Dogs in litters from 7 dams and 2 sires were paired by sex and weight within litters and randomly assigned to a control-fed group or a limit-fed group that received 25% less food than the control-fed group. Radiographic evaluation of prevalence and severity of osteoarthritis in the hip, shoulder, elbow, and stifle joints was performed when dogs were 8 years of age. RESULTS: Radiographic evidence of osteoarthritis that affected multiple joints was significantly more common in the control-fed group than in the limit fed group. Prevalence of lesions in the hip joint was 15/22 in the control-fed group and 3/21 in the limit-fed group. Prevalence of lesions in the shoulder joint was 19/22 in the control-fed group and 12/21 in the limit-fed group; lesions in this joint were generally mild. Severity, but not prevalence, of osteoarthritis in the elbow joint was greater in the control-fed group than in the limit-fed group. CONCLUSIONS AND CLINICAL RELEVANCE: Prevalence and severity of osteoarthritis in several joints was less in dogs with long-term reduced food intake, compared with control dogs. Food intake is an environmental factor that may have a profound effect on development of osteoarthritis in dogs. PMID- 11110460 TI - Use of ciliogenesis in the diagnosis of primary ciliary dyskinesia in a dog. AB - Primary ciliary dyskinesia is a congenital condition that may cause chronic rhinitis and bronchopneumonia. Primary ciliary dyskinesia may be diagnosed by induction of ciliogenesis by use of in vitro cell culture. Induction of ciliogenesis allows for differentiation between primary and secondary ciliary dyskinesia. PMID- 11110461 TI - Visceral leishmaniasis in a dog from Maryland. AB - Visceral leishmaniasis is a zoonotic disease that can be transmitted from dogs to humans by a sand-fly vector. Endemic cases of visceral leishmaniasis among dogs in Oklahoma, Texas, and Ohio have been reported. Recent reports of visceral leishmaniasis in Foxhounds in the eastern coastal states has raised new concerns about the importance of this disease in the United States. PMID- 11110462 TI - Career racing performance in Thoroughbreds treated with prosthetic laryngoplasty for laryngeal neuropathy: 52 cases (1981-1989). AB - OBJECTIVE: To compare racing performance before and after prosthetic laryngoplasty for treatment of laryngeal neuropathy in inexperienced and experienced Thoroughbred racehorses. DESIGN: Retrospective study. ANIMALS: 52 Thoroughbred racehorses treated with prosthetic laryngoplasty for laryngeal neuropathy. PROCEDURE: Lifetime race records were analyzed by use of a verified regression model. Individual race records and hospital records were also reviewed. RESULTS: Experienced horses had a decline in performance, as measured by performance index, earnings percentage, and mean prediction error, during the 6-month period before prosthetic laryngoplasty. Performance improved after surgery, relative to performance in 1 to 4 races immediately before surgery, but did not attain previous baseline values for performance index and earnings percentage, although racing speed was restored to baseline values. Factors associated with failure to attain baseline levels of performance included other racing-related injuries and disorders, major complications of surgery, and age. Individually, however, many horses had long and successful careers after surgery. Performance of inexperienced horses after surgery was at least equal to that of experienced horses. CONCLUSIONS AND CLINICAL RELEVANCE: In addition to warning clients of the complications associated with prosthetic laryngoplasty, it may be prudent to provide a guarded prognosis for full restoration of racing performance in older horses, unless they are especially talented and are free of other racing related problems. PMID- 11110463 TI - Hepatic effects of halothane and isoflurane anesthesia in goats. AB - OBJECTIVE: To determine hepatic effects of halothane and isoflurane anesthesia in young healthy goats. DESIGN: Randomized prospective clinical trial. ANIMALS: 24 healthy 9-month-old female goats. PROCEDURE: Goats were sedated with xylazine hydrochloride and ketamine hydrochloride and anesthetized with halothane (n = 12) or isoflurane (12) while undergoing tendon surgery. End-tidal halothane and isoflurane concentrations were maintained at 0.9 and 1.2 times the minimal alveolar concentrations, respectively, and ventilation was controlled. Venous blood samples were collected approximately 15 minutes after xylazine was administered and 24 and 48 hours after anesthesia, and serum aspartate aminotransferase (AST), sorbitol dehydrogenase (SDH), alkaline phosphatase (ALP), and gamma-glutamyltransferase (GGT) activities and bilirubin concentration were measured. Goats were euthanatized 25 or 62 days after anesthesia, and postmortem liver specimens were submitted for histologic examination. RESULTS: All goats recovered from anesthesia and survived until euthanasia. Serum SDH, GGT, and ALP activities and bilirubin concentration did not increase after anesthesia, but serum AST activity was significantly increased. However, serum hepatic enzyme activities were within reference limits at all times in all except 1 goat in which serum AST activity was high 24 and 48 hours after anesthesia. This goat had been anesthetized with halothane and had the longest duration of anesthesia. No clinically important abnormalities were seen on histologic examination of liver specimens. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that use of halothane or isoflurane for anesthesia in young healthy goats is unlikely to cause hepatic injury. PMID- 11110464 TI - Hyperglycemia, hypernatremia, and hyperosmolarity in 6 neonatal llamas and alpacas. AB - Neonatal camelids can develop hyperglycemia, hypernatremia, and hyperosmolarity in response to a combination of stress and inadequate water intake. Clinical signs of this syndrome include a fine head tremor, ataxia, and a base-wide stance of the hind limbs, but biochemical analyses are necessary to confirm the diagnosis. Camelids appear to be susceptible to this syndrome because of a poor insulin response to hyperglycemia; hypernatremia results from free water loss associated with glucose diuresis. Water loss associated with glucose diuresis may necessitate a higher rate of fluid administration in camelids with this syndrome than is typically used for treatment of hypernatremia in calves. PMID- 11110465 TI - Strain-induced symmetry breakdown in HOLZ lines from L1(2) titanium tri aluminides. AB - Energy filtered CBED patterns of the Cr-stabilized L1(2) phase of a titanium tri aluminide alloy reveal deficit higher order Laue zone (HOLZ) lines in the zeroth order diffraction disk, for which the expected 4-mm symmetry is significantly broken in the <001> projection. This apparent break of symmetry may be explained by the presence of lattice strains of order 10(-3). Effects of strain-induced lattice distortions on HOLZ line symmetries are calculated by an introduction of rhombohedral, tetragonal or monoclinic distortions to the cubic unit cell. It is shown how tensile and shear components of strain affect the overall HOLZ line symmetries in different ways. PMID- 11110466 TI - A reconstruction method of surface morphology with genetic algorithms in the scanning electron microscope. AB - A reconstruction method of surface morphology with genetic algorithms from a line scan in the scanning electron microscope (SEM) has been developed. This method is an application of genetic algorithms, which are search algorithms based on the mechanics of natural selection and genetics, to surface morphology in SEM. To reconstruct the microstructures of a specimen surface on a substrate, the system requires only a line scan taken by a conventional SEM. To deal with statistical fluctuations in the grey level of each pixel on the experimental line scan, the reduced chi-square distribution is used as the objective function and a scheme for minimizing the number of vertices in the reconstructed surface profile is adopted. The resolution in the Z-direction tends to be statistical and independent of the angle of the specimen surface. PMID- 11110467 TI - Quantitative intensity measurement of equal thickness fringes in Si and MgO crystal images with an energy-filtering transmission electron microscope using an imaging plate. AB - Quantitative measurement of intensity profiles of equal thickness fringes has been carried out in Si and MgO crystal images with an energy-filtering transmission electron microscope using an imaging plate. The crystals have a 90 degrees wedge-shape with [110] surfaces for Si and with [100] surfaces for MgO, and are observed under the exact axial incidence of a 200 keV electron beam along the [100] axis for Si and along the [110] axis for MgO. The intensities are measured in bright field and 022 and 040 dark field images for Si, and in bright field and 111, 002, 220, 113, 222, and 004 dark field images for MgO, with and without an energy slit having +/- 5 eV energy width for incident electrons. The intensity profiles obtained from the images are presented as standard experimental data for calculation of electron diffraction intensities. A few simulation programs for high-resolution transmission electron microscopy are checked by comparing the calculated diffraction intensities with the experimental data. The complex potential suitable for matching the data is discussed. PMID- 11110468 TI - Application of SEM-modified X-ray microscope to entomology and histology, and effects of X-ray coherence in imaging. AB - A projection X-ray microscope has been constructed by modifying a scanning electron microscope (HITACHI S-2500CX). Characteristic internal structures and their changes in an aphid, a fly, an ant, a water bear and a beetle are observed first, non-destructively, by X-ray microscope. Stereo-pair X-ray micrographs of Golgi-stained sections of porcine liver show a network of bile canaliculi and stellate cells. The very clear micrographs presented indicate the usefulness of the microscope in studies of entomology and histology. In some micrographs, very thin whiskers or hairs are visible, which suggests that a phase-contrast effect is present in the imaging. X-ray micrographs of an ant hair taken at various distances between the X-ray source and the specimen, D(s), indicate that the image enhancement due to the phase contrast increases with increasing D(s). Degree of the lateral coherence of X-rays of wave length 0.1 nm is theoretically estimated near the hair, with the result that the coherence increases from about zero (almost complete incoherence) at D(s) = 0.1 mm to about unity (almost complete coherence) at D(s) = 6 mm, in agreement with the observation. PMID- 11110470 TI - The microstructural analysis of SiC nanorods by high-resolution electron microscopy. AB - Beta-SiC nanorods have been synthesized by the reaction of SiO and carbon nano capsules. For the synthesis of SiC nanorods, it was examined that the reaction temperature and the ratio of SiO to carbon nanocapsules are important and the most appropriate temperature and ratio are around 1380 degreesC and 5:2, respectively. The synthesized SiC nanorods were characterized by high-resolution electron microscopy. Most of the SiC nanorods are straight and have the diameter of 30-150 nm while the SiC tips of the SiC nanorods have the size 100-400 nm. The SiC nanorods have many stacking faults normal to the [111] direction. Each SiC nanorod has one kind of preferential axis direction, which is either parallel or normal to the [111] direction. Based on the microstructural analysis of the SiC nanorods, a possible growth mechanism of the SiC nanorods is proposed. PMID- 11110469 TI - Localization of chromophore absorption signals in TEM with an improved prism mirror-prism filter. AB - A corrected prism-mirror-prism electron energy filter with curved entrance and exit faces was designed and incorporated into a Zeiss EM902 transmission electron microscope. The installation of the new filter required little modification to the existing microscope geometry and electronics. The filter had an energy resolution of 1.1 eV over the full image plane (acceptance half angle 10 mradian). The improved energy resolution was applied in studies of the low electron energy loss region that includes molecular orbital excitations or chromophore energy absorptions. Chromophore signal behaviour under electron irradiation was characterized using embedded crystals of hematin and of the dye mercury orange. Images of these crystals confirmed the expected decrease in signal intensity on shifting the selected energy loss from the region of molecular orbital excitations (less than approximately 5 eV) to higher energy losses. Electron irradiation-induced fading of the chromophore signal from hematin and mercury orange yielded similar 1/e dose values of 1.1 x 10(5) e(-) nm(-2) and 1.4 x 10(5) e(-) nm(-2) respectively. In a cellular context, chromophore signals were obtained from energy-filtered images of RIF-1 cell sections containing the photosensitizer chlorin e6 and from sections of BS-C-1 cells with cytoskeletal labelling via FITC-conjugated antibodies. The respective signals were extracted using a dose-dependent method or a shift in selected energy. Chromophore signal distributions were in agreement with fluorescence light microscopic images, but provided detail at higher spatial resolution. PMID- 11110471 TI - Beam alignment and related problems of spherical aberration corrected high resolution TEM images. AB - For spherical aberration corrected transmission electron microscopes recently developed, the Scherzer resolution in proportion to C(s) 1/4 lambda 3/4 is still inevitably limited by the influence of some other electron optical factors, such as chromatic aberration coefficient of objective lens (C(c)), beam divergence and alignment, incident electron energy spread (deltaE), and the instability of accelerating voltage (deltaV) and of lens current (deltaI). Depending on the image resolution guaranteed by C(s) correction, the defocus spread caused by C(c), deltaV, deltaI and deltaE would need to be reduced. The effect of beam alignment as a phase shift in contrast transfer function is also studied for the C(s) corrected microscopes with different values of C(s), defocus and spatial frequency. There is a relationship between the phase shift and defocus that allows us to find a series of 'alignment-free' focal conditions. A triangular relation among C(s), defocus and lattice spacing is established for proper image contrast and 'alignment-free' imaging with the C(s) corrected microscope. PMID- 11110472 TI - Analytical high-resolution TEM study of supported gold catalysts: orientation relationship between Au particles and TiO2 supports. AB - An analytical high-resolution TEM study was carried out for gold deposited on TiO2 (anatase and rutile) to obtain a structural insight into the unique catalytic properties of the system. Preferred orientations between Au particles and the TiO2 support were often observed and a model for the epitaxial orientation between the Au particles and anatase TiO2 was proposed. These results were consistent with the results calculated by the coincidence of reciprocal lattice point (CRLP) method. PMID- 11110473 TI - Polysomes of eukaryotic cells observed by electron microscopy. AB - Shapes of polysomes of eukaryotic cells were determined by surface spreading of cells. We examined unicellular protozoan Tetrahymena cells, rabbit reticulocytes, and cultured MDCK, BHK and HeLa cells. Polysomes had ring-, 8- and caterpillar shaped forms, indicating that eukaryotic cells contain fundamentally circular polysomes. Isolated and partially purified polysomes from Tetrahymena cells had features of polysomes similar to those of spread cells. These findings indicate that circular polysomes are not an effect of the spreading, but are actual features of the cells. PMID- 11110474 TI - The effects of diethylcarbamazine on the morphology and ultrastructure of foetal mouse cerebellar neurons in culture. AB - This paper examines morphological and ultrastructural changes in foetal mouse cerebellar neurons treated with a culture medium containing diethylcarbamazine (DEC), a widely used anti-parasitic drug known to interfere with vesicle transfer to and from the Golgi apparatus. The effects of DEC were drastic, with the complete disorganization of the Golgi apparatus and the appearance of large vacuoles within the cytoplasm. Quantitative data comparing normal and DEC-treated neurons in culture demonstrated significant differences between the length of Golgi cisternae and numbers of clear vesicles and secretory vesicles. The evidence therefore suggests that the various synthetic and secretory activities involving these organelles are severely impaired. PMID- 11110475 TI - Scanning electron microscopic changes in the morphology of rabbit pulmonary tissue biopsied following ischemia and reperfusion: a window of opportunity? AB - Reperfusion is known to cause tissue damage in ischemic pulmonary tissue. We investigated the time frame of this occurrence by examining electron microscopic changes in lung tissue. Isolated, perfused, and ventilated rabbit lungs (and heart) were placed en bloc in a 37 degrees C chamber and perfused through the pulmonary artery at 15 mm Hg pressure with oxygenated Krebs-Henseleit buffer, pH 7.4, 70 ml min(-1), for 20 min and the pulmonary pump and ventilator were stopped. The resultant ischemic state was maintained for 2 h, and reperfusion resumed with the same buffer. The lungs of four groups of rabbits (n = 5 per group) were each subjected to 30 min, 1, 2, and 4 h of reperfusion respectively. Upon completion, lungs were biopsied for scanning electron microscopy. Ischemic damage including the loss of lung architecture, and edema were seen. Reperfusion restored some of the tissue anatomy and the return to normalcy increased up to 1 h of reperfusion after which the damage increased with time. Results suggest that damage due to ischemia alone may be reversible. Initial recovery is due to the re establishment of circulation. However, with time, the damage seen may be due to free radicals and with 4 h of reperfusion, cell death may have occurred. PMID- 11110476 TI - An ultrastructural study on the effect of streptozotocin on the islets of Langerhans in mice. AB - The ultrastructural changes in the morphology of the islets of Langerhans in response to streptozotocin were studied in the mice pancreas. Male white albino CSI mice were given a single intravenous injection of 75 mg kg(-1) body weight streptozotocin, and were sacrificed at different time intervals up to 48 h following the treatment. Their pancreases were excised and randomly processed for electron microscopic examination. Hyperglycaemia and glucosuria were detected 8 h after treatment, became remarkably high at 24 h and persisted then after. Light and electron microscopic examination of the islets of Langerhans from treated mice revealed an early chromatin aggregation and cytoplasmic vesiculation in the central B cells during the first 2 h of treatment. Nuclear shrinkage and pyknosis with swelling of mitochondria and endoplasmic reticulum were evident 8 h later, and lysis of B cells occurred 12 h after treatment. The morphology of A and D cells at the margin of the islets and in between B cell debris looked perfectly unaltered. Macrophage infiltration among lytic B cells was seen 24 h after drug administration, which contained clear and large phagocytic vacuoles. The necrobiotic and phagocytic figures disappeared from the pancreatic sections of 48 h treated mice, and the islets were smaller in size and consisted entirely of intact A and D cells with occasional degranulated B cells. No features of apoptosis were ever recorded, and the exocrine pancreatic tissue was protected from the effect of streptozotocin. In conclusion, the present study illustrates the sequence of morphological changes that occurs in the islets of Langerhans of mice after streptozotocin administration. It also confirms that streptozotocin at a high single dose in mice produces a specific necrosis of B cells with no evidence of apoptotic figures as another mechanism of cell death. PMID- 11110477 TI - High-resolution freeze-etching replica images of the disk and the plasma membrane surfaces in purified bovine rod outer segments. AB - Molecular organization of the photoreceptor disk membrane was revealed by the freeze-deep etching replica method using purified and successively rinsed bovine rod outer segment (ROS). Various membrane particles with different shape and sizes were found on cytoplasmic surface (PS face) as well as on both P and E fracture faces, which are presumed to be peripheral membrane proteins such as transducin, phosphodiesterase, guanylate cyclase and so on. Membrane particles seen on PS face were catalogued in size. The histogram on their number and size showed that they were classified at least into two major groups, the group of particles about 50 nm2 in size and the group of particles about 115 nm2 in size. The distribution density of the 115 nm2 particle was 1200 microm(-2) in native state, but it decreased to 125 microm(-2) after washing with hypotonic buffer solution. Namely, the group of the 115 nm2-particle seems to be mainly composed of peripheral membrane proteins. Rinsing with the sucrose free buffer at the final step of the purification procedure enabled us to observe three types of filaments localized in ROS (filaments connecting disk to disk at the margin, filaments connecting disk to the plasma membrane, filaments associated with PS face of disk membrane); and also to find characteristic domains with crystal arrangement of particles on the external surface (ES face) of ROS plasma membrane. PMID- 11110478 TI - Structural change of bovine retinal cGMP phosphodiesterase by release of its gamma subunit: direct imaging by improved low angle rotary shadowing. AB - Cyclic GMP phosphodiesterase (PDE), a key enzyme for phototransduction, contains two catalytic subunits, Palpha and Pbeta, and two identical regulatory subunits, Pgammas. Neither the structure of the subunits of PDE nor their changes in structure during PDE regulation have been known. Here, improved low angle rotary shadowing was applied to depict the three-dimensional structure of bovine PDE (Palphabetagammagamma) and its changes by Pgamma release. Palphabetagammagamma and Palphabetagamma were isolated from photoreceptor membranes after treatment with a hydrolysis-resistant GTP analogue, and Palphabeta was prepared from Palphabetagammagamma tryptic digestion. Images of Palphabetagammagamma consisted of two crooked strands. These two strands faced each other to make a ring shape, but this ring structure was bent at the centre line between the two strands. In Palphabetagamma, one of these strands changed its shape toward reducing the central space of the ring structure. This ring appeared to be more bent at the centre line. In Palphabeta, both strands changed their shape such that the ring structure appeared to be a twisted quasi ring shape. These observations suggest that in Palphabetagammagamma each Pgamma is complexed with a catalytic subunit, and that the shapes of Palpha and Pbeta are drastically changed by the Pgamma release. These shape changes are no doubt crucial for various PDE regulations, such as activation of cGMP hydrolysis by Palphabeta, interaction of Palphabeta with GARP2 and a GARP2-like protein and cGMP binding to non-catalytic sites on Palphabeta. PMID- 11110479 TI - Changes in the blood biochemical and haematological profile of neonatal calves with age. AB - Fourteen calves were used to investigate the changes from birth to 83 days of age in the concentrations of serum albumin, alkaline phosphatase, beta hydroxybutyrate, plasma cortisol, serum creatine kinase, creatinine, iron, plasma fibrinogen, serum gamma-glutamyl transferase, plasma glucose, haptoglobin, serum non-esterified fatty acids, total protein, transferrin, triglycerides, urea and gamma globulin; the haematological variables measured were: basophils, eosinophils, haematocrit, haemoglobin, lymphocytes, mean cell haemoglobin, mean cell haemoglobin concentration, mean cell volume, monocytes, band neutrophils, neutrophils, platelets, red blood cells and white blood cells. The changes are presented as a series of graphs and the values are discussed in relation to the published reference ranges for adult cattle. Two populations of calves were identified which gave rise to a bimodal distribution for some of the variables. Differences in haematocrit, haemoglobin and red blood cell counts were apparent at birth, with raised values for these measurements being associated with an increased white blood cell and neutrophil count between three and 27 days of age. PMID- 11110480 TI - Canine distemper virus neutralising antibodies in vaccinated dogs. AB - The associations between the levels of canine distemper virus neutralising antibodies and vaccination history, age and gender were investigated in a cross sectional study of a sample of 4627 dogs from the Finnish urban dog population. Dogs vaccinated with either Canlan (Langford Laboratories) or Dohyvac (Solvay Animal Health) or with both, had significantly lower titres than those vaccinated with Candur (Behringwerke), Duramune (Fort Dodge Laboratories) or Nobivac (Intervet), or with combinations including at least one of these. The vaccines were classified as having low and high immunogenicity on the basis of the geometric mean titre achieved by the vaccine when compared with the geometric mean titre of the entire dataset. The low geometric mean titre of Canlan, Dohyvac and their combination groups resulted from the large proportion of dogs without detectable titres, especially dogs under one year of age, rather than from uniformly low titres. An age-stratified comparison of vaccine usage and titres showed that the division of the vaccines into low and high immunogenicity vaccines was apparent in the dogs less than two years of age but not in the older dogs. The first vaccination with the high immunogenicity vaccines resulted in a higher proportion of dogs with detectable antibodies than even repeated vaccination with the low immunogenicity vaccines. Neither the time elapsed since the most recent vaccination nor the gender of the dogs was associated with the titre of antibody. PMID- 11110481 TI - Phenylephrine eyedrops as a diagnostic test in equine grass sickness. AB - The effect of an ocular administration of the alpha-1 adrenergic agonist phenylephrine was studied in 23 cases of grass sickness and 12 control horses. In the horses with grass sickness there was a significantly greater mean increase in the size of the palpebral fissure, as measured by the change in the angle of the eyelashes to the head observed from a frontal view. PMID- 11110482 TI - Prevalence of porcine epidemic diarrhoea virus and transmissible gastroenteritis virus infection in Korean pigs. PMID- 11110483 TI - Severe osteochondrosis in two 10-month-old beef calves. PMID- 11110484 TI - Developmental anomalies in aborted and stillborn calves in The Netherlands. PMID- 11110485 TI - Early neutering of dogs. PMID- 11110486 TI - Surviving in mixed practice. PMID- 11110487 TI - Regulation--the way forward for veterinary nurses? PMID- 11110488 TI - Controls on animal feeds and medicines: achieving the right balance. PMID- 11110489 TI - Costs and benefits of an expanding referral service. PMID- 11110490 TI - Use of an antibiotic footbath in the treatment of bovine digital dermatitis. AB - One hundred and eleven dairy cows with digital dermatitis, on six commercial farms, were used to test the efficacy of a footbath containing erythromycin for treating the condition. Four days after 55 of the cows had walked through the footbath after two successive milkings their lesions were significantly less active and painful than those of the 56 untreated cows. These 56 cows were then treated in the same way and both groups were re-examined seven days later. There were no significant differences between the clinical signs observed in the two groups, and the benefits of the treatment had persisted for the 11 days of the trial. PMID- 11110491 TI - Investigation of romifidine and detomidine for the clinical sedation of horses. AB - The effects of two intravenous doses of romifidine (80 and 120 microg/kg) and one dose of detomidine (20 microg/kg) were compared in a blinded study in 30 horses requiring to be sedated for routine dental treatment. Several physiological parameters were assessed before and for two hours after the administration of the drugs, and the horses' teeth were rasped 30 minutes after they were administered. Romifidine produced a dose-dependent effect on most parameters. Detomidine at 20 microg/kg was similar to romifidine at 120 microg/kg in the magnitude of its sedative effects, but was similar to romifidine at 80 pg/kg in its duration. There were no significant differences between the three treatments in terms of the clinical procedure score. PMID- 11110492 TI - Fractures of the central tarsal bone in eight racing greyhounds. AB - The case records of eight greyhounds that had undergone a surgical repair of either a type III or type IV fractured central tarsal bone were reviewed. In some cases difficulties were encountered which resulted in less than ideal fixation methods being used, but the fractures healed satisfactorily by intertarsal ankylosis and were radiographically complete after four to six months. For a successful outcome it was not necessary to preserve functional intertarsal joints by interfragmentary reconstruction and compression. The absence of dorsal slab fixation, poor dorsal screw positioning, or loss of the dorsal fragment, had no influence on the results. It took from six months to one year for the dogs to return to racing. PMID- 11110493 TI - Fibriscess, not abscess, resulting from a localised inflammatory response to infection in reptiles and birds. PMID- 11110494 TI - Onion poisoning in a herd of dairy cattle. PMID- 11110495 TI - Veterinary procedures for falcons re-entering the wild. PMID- 11110496 TI - Isolation of Mycoplasma mycoides small colony type from contagious caprine pleuropneumonia in India. PMID- 11110498 TI - Advisers for RCVS certificate courses. PMID- 11110497 TI - Moving backwards on resistance. PMID- 11110499 TI - Attaching and effacing E coli in rabbits. PMID- 11110500 TI - Descriptive epidemiology short reports in Diabetic Medicine--an opportunity to present data with the population as the unit of variation. PMID- 11110501 TI - Mydriasis and glaucoma: exploding the myth. A systematic review. AB - AIMS: To investigate the risk of inducing acute glaucoma following mydriasis. METHODS: Systematic review of published research 1933-1999. RESULTS: The risk of inducing acute glaucoma following mydriasis with tropicamide alone is close to zero, no case being identified. The risk with long-acting or combined agents is between 1 in 3,380 and 1 in 20,000. The presence of chronic glaucoma constitutes no additional risk. CONCLUSIONS: Mydriasis with tropicamide alone is safe even in people with chronic glaucoma. It should be advised in all patients when thorough retinal examination is indicated. PMID- 11110502 TI - The Child Health Questionnaire in children with diabetes: cross-sectional survey of parent and adolescent-reported functional health status. AB - AIMS: To study parent and adolescent-reported physical, psychosocial and family wellbeing in children aged 5-18 years with diabetes. METHODS: SUBJECTS: 5-18 year-olds attending a diabetes clinic at a tertiary children's hospital. MEASURES: (1) Child Health Questionnaire (CHQ) PF-50, a functional heath status measure for children aged 5-18 years (parents); (2) CHQ CF-80, a similar self report measure (adolescents aged 12-18 years); (3) 11 study-designed questions related to diabetes-specific concerns (parents); (4) global ratings of physical and psychosocial health (clinicians); (5) HbA1c level (all subjects). CHQ data were compared with Australian normative data collected six months earlier. RESULTS: Reports were obtained from 128 parents and 71 adolescents (90 and 92% response). The CHQ demonstrated good psychometric properties in this sample of children with diabetes. Parents reported children with diabetes to have generally poorer health than children in the normative sample, especially on psychosocial and parent/family scales. Psychosocial health was markedly lower in 5-11-year olds with HbA1c > 8.8%, but not in 12-18-year-olds. Presence of diabetes-related symptoms and concerns correlated with lower physical and psychosocial functioning. Parents and clinicians concurred in their ratings of health for 12 18-year-olds but not 5-11-year-olds. Adolescents reported their own health similarly to adolescents in the normative sample. CONCLUSIONS: Parents report children aged 5-18 years with diabetes to have poorer health than children in the normative sample across all domains. Clinicians may underrate the impact of diabetes for younger children, with possible therapeutic implications. In providing an overall description of health, instruments like the CHQ may add another dimension to the care of children with diabetes and can feasibly be used within clinical settings. PMID- 11110503 TI - Does dietary protein intake correlate with markers suggestive of early diabetic nephropathy in children and adolescents with Type 1 diabetes mellitus? AB - AIMS: To examine the relationship between dietary protein intake and possible early markers of diabetic nephropathy (creatinine clearance (CrCI), kidney volume and albumin excretion rate (AER)). METHODS: One hundred and forty-five subjects with diabetes for 5-10 years, divided into three pubertal groups, participated. Kidney volume was measured by ultrasound, and serum creatinine and HbA1c were assayed. Two or three 24-h urine collections were obtained for albumin, creatinine and urea excretion rates. Dietary protein intake was estimated from urinary urea nitrogen excretion rate. Glomerular filtration rate was estimated by creatinine clearance. RESULTS: Mean protein intake was 1.22 +/- 0.48 g x kg(-1) x day(-1) Protein intake was significantly higher in males than females (P < 0.0001) and highest in prepubertal compared to mid-pubertal and post-pubertal subjects (P < 0.001). In multiple regression analysis, protein intake was positively associated with CrCl (P < 0.0001), and male sex (P < 0.0001) and negatively associated with body surface area (P = 0.0013) and age (P = 0.01). Kidney volume and AER were not related to dietary protein intake. CONCLUSIONS: This cross-sectional study failed to show a significant relationship between dietary protein intake and markers of early nephropathy, other than CrCl. However, a longitudinal, prospective study is required to definitively assess the role of protein intake in the evolution of diabetic nephropathy. PMID- 11110504 TI - Glucagon-like peptide (GLP)-1 and leptin concentrations in obese patients with Type 2 diabetes mellitus. AB - AIMS: To assess differences in circulating leptin and glucagon-like peptide (GLP) 1 concentrations before and after an oral glucose load, in euglycaemic and isoinsulinaemic conditions, between obese patients with and without Type 2 diabetes mellitus. METHODS: Ten male obese (body mass index (BMI) > 30 kg/m2) patients with Type 2 diabetes and 20 matched non-diabetic subjects were studied. Leptin, GLP-1(7-36)amide and GLP-1(7-37) concentrations were measured 0, 30, 60, and 90 min after a 50-g oral glucose load administered 90 min after the beginning of a euglycaemic hyperinsulinaemic clamp. RESULTS: GLP-1(7-36)amide concentrations before the glucose load were significantly lower in diabetic patients than in controls (median (quartiles): 50.5 (44.7-53.2) vs. 128.7(100 172.5) pg/ml; P < 0.01), while no difference was observed in baseline GLP-1(7 37). In non-diabetic subjects, GLP-1(7-36)amide and GLP-1(7-37) concentrations increased significantly after the oral glucose load, while no glucose-induced increase in GLP-1 concentration was observed in diabetic patients. GLP-1(7 36)amide at 30, 60, and 90 min, and GLP-1(7-37) at 30 min, of the glucose challenge, were significantly lower in diabetic patients. Leptin concentrations were not significantly different in diabetic patients when compared to non diabetic subjects, and they did not change after the oral glucose load. DISCUSSION: Leptin concentrations are not significantly modified in obese Type 2 diabetic patients. GLP-1(7-36)amide baseline concentrations are reduced in Type 2 diabetes; moreover, diabetic subjects show an impaired response of GLP-1 to oral glucose in euglycaemic, isoinsulinaemic conditions. This impairment, which is not the result of differences in glycaemia or insulinaemia during assessment, could contribute to the pathogenesis of hyperglycaemia in Type 2 diabetes mellitus. PMID- 11110505 TI - Fasting plasma glucose as a screening test for gestational diabetes in a multi ethnic, high-risk population. AB - AIMS: Screening every pregnant woman for gestational diabetes mellitus (GDM), as widely recommended for high-incidence populations, strains the healthcare system excessively. This study investigated the value of fasting plasma glucose (FPG) as an alternative to the more cumbersome oral glucose tolerance test (OGTT). METHODS: One thousand six hundred and forty-four pregnant women in a multi ethnic, high-risk population were evaluated by the FPG as a screening test among two principal subgroups, i.e. women (n = 1276) at risk for GDM on clinical grounds and those women (n = 368) with a positive post 50-g, 1-h plasma glucose challenge test (GCT). Two threshold values for FPG 'ruled in or ruled out' a GDM diagnosis, which was confirmed by the 3-h, 100-g OGTT, using Carpenter's modified criteria as the 'gold standard'. RESULTS: In the women with a positive clinical history, at an optimal cut-off value of FPG < 4.4 mmol/l to rule out GDM; a sensitivity of 94.7% was achieved, 21 (1.6%) women being false negatives. Using a FPG > or = 5.3 mmol/l to rule in GDM; the specificity was 94.0% with 53 (4.2%) women being classified as false positives. FPG would have eliminated need for the OGTT in 50.9% pregnant women (misclassification rate 5.8%). In the positive GCT group, using similar cut-offs for FPG, a sensitivity of 96.6% and specificity of 90.8% was achieved with a potential to avoid 51.6% OGTTs (misclassification rate 7.3%). The positive predictive value of the GCT was 31.8% compared to 80.2% for FPG at 5.3 mmol/l. CONCLUSIONS: While previously neglected as a screening test for GDM, in selected high-risk populations the FPG offers a potentially simple, practical algorithm to screen for GDM by being cost-effective and patient friendly. A wider application should be explored. PMID- 11110506 TI - Predictors of mortality and end-stage diabetic complications in patients with Type 1 diabetes mellitus on intensified insulin therapy. AB - AIMS: To assess predictors of mortality and end-stage diabetic complications in patients with Type 1 diabetes mellitus on intensified insulin therapy. METHODS: A cohort of 3,674 patients (insulin treatment before age 31) who had participated in a 5-day in-patient group treatment and teaching programme for intensification of insulin therapy between 9/1978 and 12/1994 were reassessed after 10 +/- 3 (mean +/- SD) years. RESULTS: Vital status and data on blindness, amputations, and renal replacement therapy were documented for 97% patients; 7% patients had died, 1.3% had become blind, 2% had amputations and 4.6% started renal replacement therapy. Using the Cox proportional hazards model, the following risk factors of mortality as assessed at baseline were identified: nephropathy (at least macroproteinuria), hazard ratio 3.8 (95% confidence interval 2.6-5.6); smoking, 1.9 (1.4-2.6); diabetes duration, 1.5 (1.2-1.8) for a difference of 10 years; serum cholesterol, 1.1 (1.0-1.2) for a difference of 1 mmol/l; lower social status, 1.4 (1.1-1.8) for a difference of 1 out of 3 levels; age, 1.3 (1.1 1.6) for a difference of 10 years; male sex, 1.4 (1.1-1.9); systolic blood pressure, 1.1 (1-1.2) for a difference of 10 mmHg. For the combined endpoint - blindness or amputations or renal replacement therapy - predictors were: nephropathy, foot complications, HbA1c, smoking, cholesterol, systolic blood pressure, retinopathy, hypertension, and social status. CONCLUSION: In Type 1 diabetic patients who start intensified insulin therapy, nephropathy remains the strongest predictor of mortality and end-stage complications. Glycosylated haemoglobin is a risk factor of end-stage complications but not of mortality. Conventional risk factors comparable to the general population, particularly smoking become operative as predictors of both mortality and end-stage complications. PMID- 11110507 TI - Impact of the Xba1-polymorphism of the human muscle glycogen synthase gene on parameters of the insulin resistance syndrome in a Danish twin population. AB - AIMS: To establish the impact on the insulin resistance syndrome of the intron 14 Xba1-polymorphism in human muscle glycogen synthase (GYS1). METHODS: Parameters related to the insulin resistance syndrome were measured in 244 monozygotic twins and 322 dizygotic twins with or without impaired glucose tolerance. In addition a standard oral glucose tolerance test (OGTT) was performed. The twins were genotyped for Xba1-polymorphism in GYS1 intron 14. RESULTS: The allele frequency of Xba1 non-cutters (A1) was 0.95 and of cutters (A2) was 0.05. Of the 566 twins examined, 90.0% had the genotype A1A1 and the remainder had the genotype A1A2. No A2A2-genotypes were detected. In 11 genotypic discordant dizygotic twin pairs the insulin resistance was significantly increased in the twins carrying the A1A2 genotype regardless of sex (HOMA index 1.81 (A1A1) vs. 2.57 (A1A2), P < 0.05). Diastolic blood pressure was increased in female carriers of the A2-allele with impaired glucose tolerance or Type 2 diabetes mellitus (79 +/- 1 vs. 94 +/- 4 mmHg, P < 0.01). Apart from a marginal increased waist-to-hip ratio, no other elements of the insulin resistance syndrome were associated with the polymorphism. CONCLUSIONS: The Xba1-polymorphism of the human muscle glycogen synthase gene is correlated to insulin resistance and to diastolic blood pressure. The polymorphism does not involve any known transcription factor or any structural change in GYS1, and these correlations are therefore most probably caused by linkage to other functional polymorphisms in GYS1 or other gene polymorphisms on chromosome 19. PMID- 11110508 TI - The prevalence of diabetes, association with cardiovascular risk factors and implications of diagnostic criteria (ADA 1997 and WHO 1998) in a 1996 community based population study in Hong Kong Chinese. Hong Kong Cardiovascular Risk Factor Steering Committee. American Diabetes Association. AB - AIMS: While the American Diabetes Association (ADA) 1997 diagnostic criteria advocate the use of fasting plasma glucose only, the World Health Organization (WHO) criteria retain the use of the standard oral glucose tolerance test (OGTT). The present study evaluated the relative merit of the respective diagnostic criteria in Chinese. METHODS: Data collected for the Hong Kong Cardiovascular Risk Factor Prevalence Study was analysed. This was a representative population based study, conducted from 1995 to 1996 among 2,900 Chinese subjects aged 25-74 years using a 75-g OGTT. RESULTS: The prevalence of diabetes (known plus unknown) was 6.2% (95% confidence interval 5.3-7.1%), 9.2% (8.1-10.3%), and 9.8% (8.7 10.9%) based on ADA 1997, WHO 1985 and WHO 1998 criteria, respectively, with a very high prevalence in older subjects. The 2,451 subjects classified as normal under ADA 1997 criteria were heterogenous: 15.3% had impaired glucose tolerance; 2.1% had diabetes under WHO 1998 criteria. These latter two smaller groups had cardiovascular risk profiles comparable to that found among the impaired fasting glucose subjects (under ADA), but worse than that among the concordant normal glucose tolerance subjects. CONCLUSIONS: The ADA criteria underestimate both diabetes prevalence and cardiovascular risk in this population. Hence fasting glucose alone is an inadequate approach and OGTT should be retained to identify at-risk individuals in both clinical diagnosis and epidemiological studies. PMID- 11110510 TI - Prevalence of pernicious anaemia in patients with Type 1 diabetes mellitus and autoimmune thyroid disease. AB - AIMS: To determine the prevalence of pernicious anaemia in patients with Type 1 diabetes mellitus and autoimmune thyroid disease. METHODS: A randomly selected asymptomatic group of 63 patients with Type 1 diabetes who also had autoimmune thyroid disease was studied. Blood samples were taken and assayed for serum B12. Those subjects with serum B12 concentrations below the reference range had a further blood sample taken for determination of intrinsic factor antibody. RESULTS: One patient had been diagnosed previously to have pernicious anaemia. Three patients had low serum B12 concentration and positive intrinsic factor antibody, confirming the diagnosis of pernicious anaemia. The prevalence of pernicious anaemia in this population with Type 1 diabetes and concomitant autoimmune thyroid disease was 6.3%. In female patients the prevalence of pernicious anaemia was 8.5%. CONCLUSIONS: Patients who have both Type 1 diabetes mellitus and autoimmune thyroid disease are at risk of developing pernicious anaemia. PMID- 11110509 TI - Type 2 diabetes mellitus, impaired glucose tolerance and associated factors in a rural Palestinian village. AB - AIMS: To investigate the prevalence of Type 2 diabetes mellitus and impaired glucose tolerance (IGT) and to identify risk factors associated with diabetes in a rural Palestinian village. METHODS: A cross-sectional, population-based study investigating 500 adults aged 30-65 years (response rate 85%) determined the diabetes status using the oral glucose tolerance test (OGTT). A standard questionnaire, a simple clinical examination and laboratory tests assessed blood lipids, blood pressure, waist-to-hip ratio (WHR), body mass index (BMI) and other risk factors for diabetes RESULTS: The prevalence of Type 2 diabetes was 9.8% (95% confidence interval 7.3-12.3) and IGT 8.6% (6.1-11.1), while the prevalence standardized to the European population was 11.6% (8.8-14.4) for Type 2 diabetes and 10.3% (7.6-13.0) for IGT. Age, positive family history, high triglycerides level, and high WHR were significantly associated with Type 2 diabetes. CONCLUSIONS: Of the factors associated with diabetes, WHR and triglycerides levels are potentially modifiable, and should be addressed by preventive health activities. The high prevalence of Type 2 diabetes mellitus and its potential increase as a result of the ageing of the Palestinian population constitutes a major public health problem. PMID- 11110511 TI - Management of asymptomatic hypoglycaemia in children and adolescents with Type 1 diabetes mellitus. PMID- 11110512 TI - Capturing the prevalence of diagnosed diabetes. PMID- 11110513 TI - Alcohol septal ablation in the management of obstructive hypertrophic cardiomyopathy. PMID- 11110514 TI - Diagnosing coronary artery disease in patients with hypertension: a resolved dilemma? AB - Patients with hypertension frequently complain of chest pain and exhibit ischemic like ST segment changes on exercise electrocardiogram. However, the specificity of such changes for predicting significant coronary artery disease is very low, since these patients often exhibit a normal coronary angiogram. Several alternative non-invasive tests have been proposed and, recently, the relative performance of the available techniques has been systematically assessed. The purpose of this article is to review the relevant literature on the diagnostic tests employed in the clinical setting. Recent evidence suggests that stress echocardiography yields a better diagnostic accuracy than perfusion scintigraphy in identifying significant epicardial coronary artery disease in patients with hypertension. The low specificity of myocardial scintigraphy probably relates to the fact that this method traces perfusion abnormalities, not necessarily caused by epicardial coronary artery disease, possibly due to microvascular disease, and not axiomatically causing obvious wall motion abnormalities. PMID- 11110515 TI - Role of growth hormone in chronic heart failure: therapeutic implications. AB - Chronic heart failure is a multi-etiological cardiovascular disorder with high prevalence and poor prognosis. Medical treatment of dilated cardiomyopathy is aimed at alleviating heart failure symptoms. Diuretics, angiotensin-converting enzyme (ACE) inhibitors and very recently, beta-blockers have been shown to have favorable effects on symptoms, exercise capacity and mortality. Growth hormone (GH) and insulin-like growth factor (IGF)-1 are involved in several physiological processes such as the control of muscle mass and function, body composition and regulation of nutrient metabolism. The role of GH and IGF-1 as modulators of myocardial structure and function is well established. Receptors for both GH and IGF-1 are expressed by cardiac myocytes; therefore, GH may act directly on the heart or via the induction of local or systemic IGF-1, while IGF-1 may act by endocrine, paracrine or autocrine mechanisms. Patients with acromegaly have an increased propensity to develop ventricular hypertrophy and cardiovascular diseases; impaired cardiac efficiency can also be observed in patients with GH deficiency. Animal models of pressure and volume overload have demonstrated up regulation of cardiac IGF-1 production and expression of GH and IGF-1 receptors, implying that the local regulation of these factors is influenced by hemodynamic changes. Moreover, experimental studies suggest that GH and IGF-1 have stimulatory effects on myocardial contractility, possibly mediated by changes in intracellular calcium handling. Heart failure is due to ventricular dilation with inadequate wall thickening that leads to impaired cardiac performance; therefore, based on previous evidence we would expect beneficial effects from the use of GH in these patients. Several papers have highlighted the positive influence of GH in the regulation of heart development and performance. In patients with GH deficiency, GH administration dramatically improves cardiac function. In small open studies, acute and chronic GH treatment has demonstrated beneficial effects in patients with heart failure due to ischemic or idiopathic cardiomyopathy. Recently, two randomized, placebo-controlled studies did not show any significant GH-mediated improvement in cardiac performance in patients with dilated cardiomyopathy, despite significant increases in IGF-1. Acquired GH resistance might be an important feature of severe heart failure and explain the diverse responses to GH therapy observed in different patients. Whether GH treatment will finally find a place in the treatment of heart failure, and with which modalities, remains to be established. PMID- 11110516 TI - Coronary stenting beyond standard indications. Immediate and follow-up results. AB - BACKGROUND: Coronary stent has become an accepted treatment modality for selected indications. However, the literature shows diverse results when indications for coronary stenting are different from those tested in large randomized trials. The purpose of this study was to determine immediate and follow-up clinical and angiographic results in patients treated with coronary stenting for indications not specifically tested in large randomized trials. METHODS: Coronary stents were implanted in a total of 2060 lesions (1757 patients) in seven groups with expanded indications: left main coronary lesions, calcified lesions, small vessels (< 3 mm in size), small vessels with diffuse disease, large vessels with diffuse disease, and bifurcation lesions treated with stents in both branches or with one stent implanted only in the major branch. Stents were implanted using high balloon pressure for final inflation and in most cases with intravascular ultrasound. Clinical follow-up was achieved in 96% of patients at a mean time of 12+/-7 months. RESULTS: Primary success (range 89-96%) and acute complications (range 5.7-13%) were comparable in all groups. At follow-up, the mortality rate was highest in the group of left main stenting (12.5%) but 20% of these patients had coronary stenting on non-elective basis. The restenosis rates ranged between 16-43%. The restenosis rate was highest in the group of bifurcation lesions with stent implantation in both vessels leading to a major adverse cardiac event (MACE) rate of 62% in this group. However, the survival rate at 1 and 2 years in the overall study group was 97 and 96%, and the event free survival was 76 and 74%, respectively. The procedure-related predictors of MACE were: final intravascular ultrasound result, use of stents with non-slotted tube morphology, final stent percent stenosis, and vessel size. CONCLUSIONS: Coronary stenting beyond standard indications is feasible, with acceptable primary success and complication rates. However, the overall MACE rates were relatively high (34 62%), in particular for the indication of bifurcation lesions with stents implanted in both vessels. PMID- 11110517 TI - Twenty-five-year cardiovascular disease incidence among middle-aged men. Disease burden, time shape, predictors, risk probabilities. AB - BACKGROUND: This analysis aims at describing the 25-year experience in cardiovascular incidence in a sample of middle-aged men in rural Italy. METHODS: A total of 1712 men aged 40-59 years were examined in 1960 in two rural Italian cohorts belonging to the Seven Countries Study. Major risk factors were measured and a monitoring system for incidence of cardiovascular events was in operation for the next 25 years centered on the first event occurring within each subgroup of cardiovascular diseases. Data were modeled by the Weibull distribution incorporated in the accelerated failure time model. RESULTS: Among 1695 cardiovascular disease-free men at entry, 51.3% experienced at least one cardiovascular event, the first being a coronary heart disease (35.6%), a cerebrovascular disease (17.8%) or a peripheral artery disease (13.0%). Among all first events the most common was intermittent claudication (17.6%), followed by angina pectoris (16.6%). A proportion of 13% of all initial episodes was rapidly fatal, whereas 38 % of subjects developing a cardiovascular event died within the 25-year deadline. Age, cigarette smoking, diabetes, corneal arcus, serum cholesterol, systolic blood pressure, and vital capacity fed into the accelerated failure time model showed a statistically significant association with cardiovascular disease incidence (inverse for vital capacity). The same model provided the shape of hazard during the 25 years with major events continuing to appear in an increasing exponential fashion, while soft criteria events were less likely to appear as a first event in the late phase of follow-up. CONCLUSIONS: In middle-aged men 25-year incidence of cardiovascular diseases is high and more relevant compared to that of traditional hard criteria coronary heart disease that in the past has attracted almost exclusive interest. PMID- 11110518 TI - Early spontaneous recovery of left ventricular function in patients with myocarditis. AB - The natural history of myocarditis is varied. We describe 6 out of a cohort of 15 consecutive patients with histopathologic evidence of myocarditis who showed a remarkable early symptomatic and spontaneous recovery of left ventricular systolic function. The left ventricular ejection fraction increased to > or = 50% at discharge, and this improvement was maintained at late follow-up. The other 9 patients, despite clinical improvement, were not thought to have spontaneous recovery. Neither clinical severity of the illness (NYHA functional class) nor left ventricular ejection fraction at presentation demonstrated any difference in the two groups. By contrast, a smaller left ventricular internal diameter at end diastole and a smaller left atrial dimension as determined by transthoracic echocardiography were predictive of spontaneous recovery. Firstly, we confirm that the natural history of myocarditis is indeed varied with the possibility of early spontaneous recovery; secondly we suggest that left ventricular internal diameter at end-diastole and left atrial dimension may have prognostic implications in this disease. PMID- 11110519 TI - Spontaneous early clinical remission of myocarditis without immunosuppressive treatment. PMID- 11110520 TI - Mechanisms of myocardial ischemia in a patient with left main coronary artery atresia. AB - This report describes the different clinical and instrumental manifestations of coronary ischemia in a patient with left main coronary artery atresia. Exercise test and thallium-201 perfusion scintigraphy during isometric exercise test were negative for angina and electrocardiographic changes. Conversely, dipyridamole infusion caused severe angina, marked ST-segment changes and diffuse thallium-201 uptake abnormalities. This peculiar anatomical condition offers the opportunity of high-lighting the role played by the microcirculation in determining myocardial ischemia. PMID- 11110521 TI - Aortic valve repair for traumatic aortic insufficiency. AB - Prolapse of a commissural portion of the aortic valve due to partial intimal tear following a blunt chest trauma is a rare condition. Aortic valve repair is a technically demanding operation and the presence of aortic incompetence due to leaflet prolapse often leads to aortic valve replacement. We report the case of a patient with aortic insufficiency due to commissural disruption following a road traffic accident, and in whom aortic valve repair was performed. PMID- 11110522 TI - Left main coronary artery atresia without induced ischemia. PMID- 11110523 TI - Multiple cardiac rhabdomyomas. PMID- 11110524 TI - Further explanations for the formation of syringomyelia: back to the drawing table. PMID- 11110525 TI - Is there room for MR imaging in the assessment of hereditary motor and sensory neuropathies? PMID- 11110526 TI - Diffusing into the future. PMID- 11110527 TI - The status of status: seizures are bad for your brain's health. PMID- 11110528 TI - CSF flow measurement in syringomyelia. AB - BACKGROUND AND PURPOSE: CSF circulation has been reported to represent a major factor in the pathophysiology of syringomyelia. Our purpose was to determine the CSF flow patterns in spinal cord cysts and in the subararachnoid space in patients with syringomyelia associated with Chiari I malformation and to evaluate the modifications of the flow resulting from surgery. METHODS: Eighteen patients with syringomyelia were examined with a 3D Fourier encoding velocity imaging technique. A prospectively gated 2D axial sequence with velocity encoding in the craniocaudal direction in the cervical region was set at a velocity of +/- 10 cm/s. Velocity measurements were performed in the larger portion of the cysts and, at the same cervical level, in the pericystic subarachnoid spaces. All patients underwent a surgical procedure involving dural opening followed by duroplasty. Pre- and postoperative velocity measurements of all patients were taken, with a mean follow-up of 10.2 months. We compared the velocity measurements with the morphology of the cysts and with the clinical data. Spinal subarachnoid spaces of 19 healthy subjects were also studied using the same technique. RESULTS: A pulsatile flow was observed in syrinx cavities and in the pericystic subarachnoid spaces (PCSS). Preoperative maximum systolic cyst velocities were higher than were diastolic velocities. A systolic velocity peak was well defined in all cases, first in the cyst and then in the PCSS. Higher systolic and diastolic cyst velocities are observed in large cysts and in patients with a poor clinical status. After surgery, a decrease in cyst volume (evaluated on the basis of the extension of the cyst and the compression of the PCSS) was observed in 13 patients. In the postoperative course, we noticed a decrease of systolic and diastolic cyst velocities and a parallel increase of systolic PCSS velocities. Diastolic cyst velocities correlated with the preoperative clinical status of the patients and, after surgery, in patients with a satisfactory foraminal enlargement evaluated on the basis of the visibility of the cisterna magna. CONCLUSION: CSF flow measurement constitutes a direct evaluation for the follow-up of patients with syringomyelic cysts. Diastolic and systolic cyst velocities can assist in the evaluation of the efficacy of surgery. PMID- 11110529 TI - MR imaging of the cauda equina in hereditary motor sensory neuropathies: correlations with sural nerve biopsy. AB - BACKGROUND AND PURPOSE: Although spinal root abnormalities are known to occur, spinal MR examination is seldom performed in hereditary motor and sensory neuropathies (HMSN). The following work was undertaken to assess the MR imaging spectrum of lumbosacral spinal nerve root abnormalities and determine whether intradural nerve root involvement could be related to any biopsy feature. METHODS: Ten consecutive patients (eight male, two female; age range, 28-65 yrs) with Charcot-Marie-Tooth (CMT) (type I = 5, type II = 2) and Dejerine-Sottas disease (DSD) (n = 3) underwent a contrast-enhanced lumbosacral MR examination. Sural nerve biopsy was performed in all patients. Atypical clinical features were present in two patients. The MR scans of each patient were reviewed for possible causes of myeloradiculopathy, spinal nerve root and ganglia dimensions, signal change, and abnormal enhancement. RESULTS: In the seven patients with CMT, abnormal MR findings were intradural nerve root hypertrophy (n = 2), signal abnormalities (n = 2), and enhancement (n = 3). Two of three patients with DSD had the abnormal MR finding of intradural nerve root enhancement. In both patients with atypical clinical features, MR imaging showed nerve root hypertrophy and enhancement. Both findings were related to an increased number of onion bulbs at sural nerve biopsy. Inflammatory infiltrates were not observed in any patients. CONCLUSION: In patients with HMSN enhancement of intradural spinal nerve roots, whether or not associated with marked thickening, may be found on lumbosacral MR examinations. Spinal nerve root thickening may be responsible for atypical symptoms, and its visibility on MR images represents a useful adjunct to diagnosis. Lumbosacral spinal nerve root abnormalities were related to an extremely high number of onion bulbs (indicating active demyelination) at sural nerve biopsy. Nerve root enhancement does not seem to be related to inflammatory infiltrates. PMID- 11110530 TI - Diffusion-weighted MR imaging of the normal human spinal cord in vivo. AB - BACKGROUND AND PURPOSE: Diffusion-weighted imaging is a robust technique for evaluation of a variety of neurologic diseases affecting the brain, and might also have applications in the spinal cord. The purpose of this study was to determine the feasibility of obtaining in vivo diffusion-weighted images of the human spinal cord, to calculate normal apparent diffusion coefficient (ADC) values, and to assess cord anisotropy. METHODS: Fifteen healthy volunteers were imaged using a multi-shot, navigator-corrected, spin-echo, echo-planar pulse sequence. Axial images of the cervical spinal cord were obtained with diffusion gradients applied along three orthogonal axes (6 b values each), and ADC values were calculated for white and gray matter. RESULTS: With the diffusion gradients perpendicular to the orientation of the white matter tracts, spinal cord white matter was hyperintense to central gray matter at all b values. This was also the case at low b values with the diffusion gradients parallel to the white matter tracts; however, at higher b values, the relative signal intensity of gray and white matter reversed. With the diffusion gradients perpendicular to spinal cord, mean ADC values ranged from 0.40 to 0.57 x 10(-3) mm2/s for white and gray matter. With the diffusion gradients parallel to the white matter tracts, calculated ADC values were significantly higher. There was a statistically significant difference between the ADCs of white versus gray matter with all three gradient directions. Strong diffusional anisotropy was observed in spinal cord white matter. CONCLUSION: Small field-of-view diffusion-weighted images of the human spinal cord can be acquired in vivo with reasonable scan times. Diffusion within spinal cord white matter is highly anisotropic. PMID- 11110531 TI - Value of bone scan imaging in predicting pain relief from percutaneous vertebroplasty in osteoporotic vertebral fractures. AB - BACKGROUND AND PURPOSE: Patient selection for percutaneous vertebroplasty is often complicated by the presence of multiple fractures or non-localizing pain. Our purpose was to determine whether increased activity revealed by bone scan imaging is predictive of a positive clinical response to percutaneous vertebroplasty. METHODS: A retrospective chart review conducted at our institution yielded 28 vertebroplasty treatment sessions that had been performed after obtaining bone scan imaging for painful, osteoporotic compression fractures in 27 patients. Thirty-five compression fractures were treated during these 28 treatment sessions. In all cases, increased activity was revealed by bone scan imaging before treatment with vertebroplasty. Positive outcome was defined as subjective decrease in pain severity and/or increased level of patient mobility. RESULTS: Subjective pain relief was noted in 26 (93%) of 28 treatment sessions. In 14 (100%) of 14 cases with quantifiable pain levels, pain improved at least 3 points on a 10-point scale (range of improvement, 3-10 points; mean improvement, 7.4 points). Among the remaining 14 treatment sessions in which patients were unable or unwilling to quantify pain severity, the pain relief was described as complete or excellent pain relief in 11 (78%) of 14 cases. In 14 (100%) of 14 cases for which semiquantitative assessment of mobility was available, mobility improved at least one level (5-point graded scale; range of improvement, 1-4 points; mean improvement, 1.7 points). CONCLUSIONS: Increased activity revealed by bone scan imaging is highly predictive of positive clinical response to percutaneous vertebroplasty. PMID- 11110532 TI - Diffusion tensor MR imaging of the brain: effect of diffusion weighting on trace and anisotropy measurements. AB - BACKGROUND AND PURPOSE: In human brain, the relationship between MR signal and b value is complicated by cerebral perfusion, restricted diffusion, anisotropy, cellular membrane permeability, and active cellular transport of water molecules. Our purpose was to evaluate the effect of the number and strength of diffusion sensitizing gradients on measured isotropic apparent diffusion coefficients (ADCi), fractional anisotropy (FA), and their respective SD in different anatomic locations of the brain. METHODS: Quantitative apparent diffusion coefficients and diffusion anisotropy brain maps were obtained from 10 healthy volunteers by using six different levels of diffusion weighting (b0 = 0, bl = 160, b2 = 320, b3 = 480, b4 = 640, and b5 = 800 s/mm2), applied sequentially in six different directions (Gxx, Gyy, Gzz, Gxy, Gxz, Gyz) and coupled to a single-shot spin-echo echo-planar (2,045/115 [TR/TE]) MR imaging technique. ADCi, FA, eigenvalues (lambda1, lambda2, lamdba3)1 of the principal eigenvectors, and their respective SD were measured from seven different anatomic locations in the brain. Repeated measures analysis of variance was used to evaluate for the existence of significant differences in the average and SD of the calculated ADCi and FA as a function of the number and strength of b values. When a difference existed, the Bonferroni t method was used for paired comparisons of the groups. RESULTS: The measured ADCi was affected by the number and strength of b values (P < .05). The SD of the ADCi was affected by the strength (P < .05) but not the number of b values (P > .05). The measured FA was unaffected by the number and strength of b values (P > .05). The SD was affected by the number and strength of b values (P < .05). CONCLUSION: The number and strength of b values do influence measures of diffusion and anisotropy. Attention to the choice of diffusion sensitization parameters is important in decisions regarding clinical feasibility (acquisition time) and normative measures. PMID- 11110533 TI - High-b-value diffusion-weighted MR imaging of suspected brain infarction. AB - BACKGROUND AND PURPOSE: Recent technological advances in MR instrumentation allow acquisition of whole-brain diffusion-weighted MR scans to be obtained with b values greater than 1,000. Our purpose was to determine whether high-b-value diffusion-weighted MR imaging improved contrast and detection of signal changes in acute and chronic brain infarction. METHODS: We prospectively evaluated the MR scans of 30 subjects with a history of possible brain infarction on a 1.5-T MR imager with 40 mT/meter gradients (slew rate 150 T/m/s) by use of the following single-shot echo-planar diffusion-weighted MR sequences: 1) 7,999/ 71.4/1 (TR/TE/excitations, b = 1,000; 2) 999/ 88.1/3, b = 2,500; and 3) 7,999/ 92.1/4, b = 3,000. Diffusion-weighted MR imaging was performed in three orthogonal directions during all sequences. All subjects were scanned with fast fluid attenuated inversion recovery (FLAIR) (10,006/145/2,200/1 [TR/TE/TI/excitations]) and fast spin-echo T2-weighted (3,650/95/3 [TR/TE/excitations], echo train length, 8). The diagnosis of brain infarction was established by clinical criteria. RESULTS: Twenty women and 10 men with a mean age of 67.7 years were enrolled in the study. One subject was excluded owing to poor image quality. Twelve of 29 subjects had a clinical diagnosis of acute infarction. All 12 had lesions that were hyperintense on diffusion-weighted images at all three b values; five were cortical and seven subcortical. There was increased contrast of all lesions on high-b-value scans (b = 2,500 and 3,000). Lesions that were hypointense on diffusion-weighted images were identified and evaluated at the three different b values. At b = 1,000, there were 19 hypointense lesions, whereas at b = 2,500 and 3,000 there were 48 and 55 lesions, respectively. On FLAIR and T2-weighted images, these low-signal lesions were predominantly chronic, subcortical, ischemic lesions and lacunar infarcts, but four chronic cortical infarcts, one porencephalic cyst, and one primary brain tumor were also found. Low-signal lesions were also noted to have increased contrast on high-b value diffusion-weighted scans. CONCLUSION: High-b-value diffusion-weighted MR imaging (b = 2,500 or b = 3,000) had no impact on diagnosis of acute infarction. High-b-value diffusion-weighted MR imaging (b = 2,500) combined with diffusion weighted MR imaging at b = 1,000 improves tissue characterization by increasing the spectrum of observed imaging abnormalities in patients with suspected brain infarction. PMID- 11110534 TI - High-b-value diffusion-weighted MR imaging of adult brain: image contrast and apparent diffusion coefficient map features. AB - BACKGROUND AND PURPOSE: Recent improvements in MR gradient technology allow significant increases in diffusion weighting without prohibitive signal-to-noise degradation. The purpose of our investigation was to establish normative references for the signal intensity characteristics and apparent diffusion coefficient values of the adult brain at high b values. METHODS: Fifty adults underwent diffusion-weighted single-shot spin-echo echo-planar MR imaging. Isotropic diffusion-weighted images were obtained with b values of 0, 1,000, 2,000, 2,500, 3,000, and 3,500 s/mm2. Qualitative assessments were made in multiple regions of interest in gray and white matter. Three apparent diffusion coefficient maps were generated for each of six patients with a 2-point technique at a b value of 0 and at b values of 1,000, 2,000, and 3,000 s/mm2. RESULTS: Increasing b values result in a progressive decrease in the gray to white matter signal intensity ratio. Isointensity between gray and white matter results at b values between 1,000 and 2,000 s/mm2. At b values greater than 2,000, the gray white pattern reverses relative to the usual b value of 1,000. Apparent diffusion coefficient values were shown to decrease with increasing b values. CONCLUSION: Attention to the reversal of gray-white contrast and the dependence of apparent diffusion coefficient on the b value are important in avoiding erroneous assignment of pathologic abnormalities to normal regions. This study provides the normative data for future diffusion investigations performed at high b values. PMID- 11110535 TI - Selective neuronal necrosis associated with status epilepticus: MR findings. AB - We present the MR imaging findings in an autopsy-proven case of selective neuronal necrosis involving the entire left cerebral hemispheric cortex, left thalamus, and contralateral cerebellum following a period of status epilepticus. Imaging findings include diffusion abnormality on diffusion-weighted images and increased intensity on T2-weighted images in the above-mentioned regions of the brain. PMID- 11110536 TI - Rapid alterations in diffusion-weighted images with anatomic correlates in a rodent model of status epilepticus. AB - BACKGROUND AND PURPOSE: Diffusion-weighted MR imaging has emerged as a noninvasive tool for the detection of regional neuronal damage. We hypothesize that changes in diffusion-weighted images will correlate with pathophysiologic alterations caused by pilocarpine-induced status epilepticus. METHODS: MR images of brain tissues were examined in vivo by use of T2- and diffusion-weighted imaging at 3, 6, 12, and 24 hours after pilocarpine-induced seizures. Histologic verification of neuronal damage was also performed after imaging to assess the extent and the time course of neuronal cell death. RESULTS: The piriform cortex, amygdala, and retrosplenial (and somatosensory) cortex displayed significant apparent diffusion coefficient (ADC) decreases 12 hours after seizure initiation. In contrast, an ADC rise of 19% was observed in the hippocampus 24 hours after seizure induction. Histologic data from the piriform cortex and amygdala confirmed severe neuronal loss, whereas hippocampal damage was much less pronounced at 12 hours. Interestingly, very little histologic damage was seen in the retrosplenial cortex. CONCLUSION: This study capitalized on diffusion weighted imaging as a sensitive technique for the early identification of seizure induced neuronal damage and differentiation of regional severity of these alterations. Hippocampal neuropathology is slower and longer in duration (approximately 7 days), while the piriform cortex and amygdala exhibit very rapid neurodegenerative alterations (approximately 24 hours) after pilocarpine-induced status epilepticus. These histologic changes are reflected in opposing ADC values within these regions. PMID- 11110537 TI - Magnetization transfer ratio is unable to lateralize epileptic foci in patients with temporal lobe epilepsy. AB - BACKGROUND AND PURPOSE: A preliminary report suggested that magnetization transfer ratio (MTR) was useful to lateralize epileptic foci in patients with refractory temporal lobe epilepsy (TLE). We attempted to confirm this finding in a larger group by investigating the relationship between MTR of mesial temporal structures and seizure lateralization in patients with refractory TLE. METHODS: We compared the MTR of amygdalae and hippocampi of 10 patients with unilateral TLE to values obtained from 10 healthy control participants. RESULTS: Three of 10 patients with TLE had MTR values that were 2 SD below the normal mean; the MTR abnormality was concordant with electroclinical lateralization in only one of the three. CONCLUSION: We conclude that MTR measurements of amygdalae and hippocampi are not useful for lateralization of TLE. PMID- 11110538 TI - Dementia, quantitative neuroimaging, and apolipoprotein E genotype. AB - BACKGROUND AND PURPOSE: Quantitative MR imaging differences in an elderly population of subjects with various clinical disorders (including dementia, particularly Alzheimer's disease and vascular dementia) and disorders of mild cognitive impairment were examined. Potential quantitative MR differences were assessed by presence or absence of the apolipoprotein E (APOE) epsilon4 allele and by level of cognitive deficit. METHODS: One hundred eighty subjects with a diagnosis of dementia or other clinical disorders were identified from an eligible population of 5,677 elderly individuals. Age, duration of disease, and head size (where appropriate) were considered as covariates. APOE genotype was determined by polymerase chain reaction using buccal material. Axial and coronal intermediate- and T2-weighted MR images were quantified using a multispectral segmentation algorithm. Cognitive status was assessed by means of a modified Mini Mental Status Examination. RESULTS: All types of dementing illness showed significant volume reductions in the majority of structures examined, particularly in the total brain, hippocampus, and white and gray matter, and increased CSF and ventricular volumes. Subjects with mild cognitive impairment showed fewer atrophic changes but were still distinguishable from the 24 control subjects. Presence of an epsilon4 allele was associated with smaller hippocampal volume in subjects with Alzheimer's disease and vascular dementia within just 1 year of disease onset. For other analyses, atrophy related to the presence of the epsilon4 allele disappeared after controlling for age and length of disease. CONCLUSION: The effects of the epsilon4 allele on brain morphology may be subtly expressed early in the development of dementia, but do not specifically affect cerebral atrophy thereafter. Cognitive impairment is associated with atrophy irrespective of diagnosis and presence of epsilon4. PMID- 11110539 TI - Functional MR imaging in Alzheimer's disease during memory encoding. AB - BACKGROUND AND PURPOSE: We applied functional MR imaging with a learning task in healthy elderly volunteers and in patients with Alzheimer's disease to study brain activation during memory performance. The purpose was to determine the feasibility of functional MR imaging during a learning task in healthy elderly volunteers and in patients with Alzheimer's disease and to test our hypothesis that brain activation is decreased in the medial temporal lobe (MTL) memory system in patients with Alzheimer's disease compared with control volunteers. METHODS: In 12 patients with mild to moderate forms of Alzheimer's disease and 10 elderly control volunteers, activation of the MTL memory system was studied. We used two learning tasks that required the encoding of new information into memory. After the functional MR imaging experiment, participants were tested for recognition of the encoded objects. RESULTS: In the elderly control volunteers, activation during memory encoding was observed in medial and lateral temporal lobe structures (fusiform, parietal and occipital parts, and hippocampal formation) and in the frontal cortex, as reported previously in studies of young control volunteers. Focusing on the MTL, we observed that activation was significantly decreased in patients with Alzheimer's disease compared with control volunteers in the left hippocampus and parahippocampal gyrus bilaterally during the first encoding task but not during the second (P < .05, uncorrected). CONCLUSION: Functional MR imaging with a learning task seems feasible in elderly volunteers and in patients with Alzheimer's disease. The measured functional signal decrease in MTL areas warrants further exploration of the (early) diagnostic usefulness of functional MR imaging in cases of Alzheimer's disease and other dementias. PMID- 11110540 TI - Corpus callosum infarcts with atypical clinical and radiologic presentations. AB - Infarcts of the corpus callosum have not been well documented in the radiologic literature. We present five cases that were unusual in either their clinical or radiologic presentation or both. Biopsies were performed in three of the five cases, and in time, all lesions evolved in a pattern consistent with infarct. Recognition of the varied clinical and radiologic presentation of infarcts of the corpus callosum will obviate the need for biopsy in most patients. PMID- 11110541 TI - Using 80 kVp versus 120 kVp in perfusion CT measurement of regional cerebral blood flow. AB - Perfusion CT studies of regional cerebral blood flow (rCBF), involving sequential acquisition of cerebral CT sections during IV contrast material administration, have classically been reported to be achieved at 120 kVp. We hypothesized that using 80 kVp should result in the same image quality while significantly lowering the patient's radiation dose, and we evaluated this assumption. In five patients undergoing cerebral CT survey, one section level was imaged at 120 kVp and 80 kVp, before and after IV administration of iodinated contrast material. These four cerebral CT sections obtained in each patient were analyzed with special interest to contrast, noise, and radiation dose. Contrast enhancement at 80 kVp is significantly increased (P < .001), as well as contrast between gray matter and white matter after contrast enhancement (P < .001). Mean noise at 80 kVp is not statistically different (P = .042). Finally, performance of perfusion CT studies at 80 kVp, keeping mAs constant, lowers the radiation dose by a factor of 2.8. We, thus, conclude that 80 kVp acquisition of perfusion CT studies of rCBF will result in increased contrast enhancement and should improve rCBF analysis, with a reduced patient's irradiation. PMID- 11110542 TI - A comparison of magnetization transfer ratio, magnetization transfer rate, and the native relaxation time of water protons related to relapsing-remitting multiple sclerosis. AB - BACKGROUND AND PURPOSE: Magnetization transfer (MT) imaging and measurements of the magnetization transfer ratio (MTR) have extended our capability to depict and characterize pathologic changes associated with multiple sclerosis (MS). We wanted to investigate whether the analysis of other MT parameters, such as magnetization transfer rate (k(for)) and relative measure of water content (T1(free)), adds insight into MS-related tissue changes. METHODS: Quantitative MT imaging by use of phase acquisition of composite echoes was performed in nine patients with clinically definite relapsing-remitting MS and eight healthy control subjects on a 1.5-T MR system. We analyzed a total of 360 regions of interest and compared control white matter with various types of lesions and normal-appearing white matter in MS. RESULTS: We found a strong correlation between the MTR and k(for), but this relation was non-linear. A slight but significant reduction of the MTR in normal-appearing white matter of patients with MS was attributable to a reduced transfer rate only, whereas a lower MTR was associated with both a reduction of k(for) and an increase of T1(free) in regions of dirty white matter. Moreover, areas such as edema and T1-isointense lesions had a similar MTR but could be differentiated on the basis of Tl(free). CONCLUSION: Estimates of k(for) and T1(free) appear to complement MTR measurements for the understanding of MT changes that occur with different types of MS abnormalities in the brain. PMID- 11110543 TI - Brain arteriovenous malformations: assessment with dynamic MR digital subtraction angiography. AB - BACKGROUND AND PURPOSE: Conventional catheter angiography (CCA) is the current reference standard for the diagnosis, assessment, and management of pial brain arteriovenous malformations (AVMs). The purpose of this study was to develop an MR angiographic technique that produces dynamic images comparable to those provided by CCA and to apply the technique to the investigation of pial brain AVMs. METHODS: Twenty patients with brain AVMs referred for stereotactic radiosurgery were recruited. All patients had CCA performed on a 1.5-T superconducting system. Sixty images were obtained at a rate of one image per second. Slices were orientated to produce Towne, lateral, and anteroposterior projections. A set of mask images was taken and then a series during the passage of a bolus of contrast material. MR examinations were assessed independently by neuroradiologists blinded to the conventional catheter angiographic findings. RESULTS: The nidus of the AVMs was depicted in 19 of the 20 patients, and correlation with CCA was excellent for measurements of maximum diameter. Venous drainage was correctly assessed in 18 of 19 cases. CONCLUSION: MR digital subtraction angiography shows promise as a noninvasive, dynamic angiographic tool for planning stereotactic radiosurgery of AVMs already delineated by catheter angiography. At present, it suffers from temporal and spatial resolution, which impede the assessment of some brain AVMs. PMID- 11110544 TI - Contrast-enhanced MR angiography of intracranial giant aneurysms. AB - BACKGROUND AND PURPOSE: Intravoxel phase dispersion and flow saturation often prevent adequate depiction of intracranial giant aneurysms on 3D time-of-flight (3D-TOF) MR angiography (MRA). Additional diagnostic difficulties may arise from T1 contamination artifact of an associated blood clot. Our aim was to assess whether contrast-enhanced MRA could improve the evaluation of giant aneurysms and to compare two different types of contrast-enhanced MRA. METHODS: We studied 11 aneurysms in 10 patients (age range, 31-77 years) with giant aneurysms of the anterior (n = 9) and posterior (n = 2) cerebral circulation by comparing 3D-TOF, first-pass dynamic contrast-enhanced MRA, and steady-state contrast-enhanced 3D TOF sequences. Additional comparison with digital subtraction angiography (DSA) was performed in eight aneurysms. RESULTS: In nine of 11 aneurysms, 3D-TOF did not adequately show the lumen and exiting vessels. Contrast-enhanced 3D-TOF and dynamic contrast-enhanced MRA showed the aneurysm sac and exiting vessels in all of these cases. Dynamic contrast-enhanced MRA showed a better intravascular contrast than did contrast-enhanced 3D-TOF, which led to better delineation of the aneurysms. T1 contamination artifact from intra- or extraluminal blood clot was evident on the 3D-TOF images in four cases. The artifact was less marked on the contrast-enhanced 3D-TOF image and was completely eliminated on the dynamic contrast-enhanced MRA image by subtraction of precontrast images. The diagnostic information provided by dynamic contrast-enhanced MRA was comparable to that provided by DSA. CONCLUSION: Precontrast 3D-TOF is inadequate for the assessment of giant cerebral aneurysms. Both contrast-enhanced 3D-TOF and dynamic contrast enhanced MRA reliably show the aneurysm sac and connected vessels. Dynamic MRA provides a superior contrast between flow and background and eliminates T1 contamination artifact. It should therefore be considered as the MRA sequence of choice. PMID- 11110545 TI - Delayed reconfiguration of a Guglielmi detachable coil mass associated with late occlusion of an adjacent aneurysm and parent vessel. AB - We present a case of asymptomatic, progressive, late occlusion of the left superior cerebellar artery (SCA) and an aneurysm arising from the junction of the SCA and basilar artery after embolization of an adjacent aneurysm arising between the left posterior cerebral artery and the left SCA. The delayed occlusion was associated with reconfiguration of the Guglielmi detachable coils at the neck of the treated aneurysm. PMID- 11110546 TI - Intracranial hemorrhage after stenting and angioplasty of extracranial carotid stenosis. AB - BACKGROUND AND PURPOSE: The transluminal angioplasty and stenting procedure has been recently advocated as a potential alternative to surgical endarterectomy for the treatment of severe extracranial carotid stenosis. This study assesses the incidence and significance of intracranial hemorrhage occurring after this procedure. METHODS: We retrospectively reviewed 104 carotid arteries (96 internal, two external, and six common) in 90 patients (63 male; mean age, 69.4 years; range, 48-88 years) who underwent primary stenting and angioplasty by use of Wallstents (103/104) at three centers between January 1996 and January 1999. Seventy-five (83%) patients were referred by neurosurgery departments. Seventy one (68%) arteries were symptomatic; the mean stenosis percentage was 85% (range, 40-99%). RESULTS: Four intraparenchymal hemorrhages occurred, representing 4.4% of patients and 3.8% of vessels, after angioplasty and stent placement. The mean preoperative stenosis percentage was 95% (range, 90-99%). One hemorrhage occurred immediately after stent placement, whereas the three other hemorrhages occurred in a delayed fashion (mean, 2.8 days). The mean hematoma size was 4.8 cm (range, 2-8 cm). Three patients had associated subarachnoid or intraventricular bleeding; the fourth had associated subdural hemorrhage. Three hemorrhages were fatal; the fourth experienced two seizures only. No acute neurologic symptoms were present prior to hemorrhages, and there was no postprocedural hypertension in these patients. All had been receiving antiplatelet agents as well as intraprocedural IV heparin. CONCLUSION: Intracranial hemorrhage can occur after carotid angioplasty and stenting. We speculate that this represents cerebral hyperperfusion injury. The 3.8% incidence of cerebral hemorrhage observed is approximately sixfold greater than that reported post endarterectomy (0.6%) (95% CI, 0.2-8.7%). This is not statistically significant in this small study group. This trend may reflect patient selection, different anticoagulation protocols, and/or study population size. Additional data are needed to determine the safety and efficiency of carotid stenting as a treatment for carotid stenosis. PMID- 11110547 TI - Superselective continuous arterial infusion chemotherapy through the superficial temporal artery for oral cavity tumors. AB - BACKGROUND AND PURPOSE: High-dose intraarterial chemotherapy with repeated one shot infusion may be useful for treating head and neck tumors. We evaluated the efficacy of superselective continuous arterial infusion chemotherapy administered via a coaxial catheter system and compared the results with those of subselective catheterization for treatment of oral cavity tumors. METHODS: Forty-nine consecutive patients with tumors of the oral cavity (clinical stage I, 12 cases; stage II, 19 cases; stage III, six cases; stage IV, 12 cases) were treated by arterial infusion chemotherapy. After a guiding catheter was advanced into the superficial temporal artery, superselective catheterization was performed using a coaxial system microcatheter. Superselective catheterization was accomplished in 34 cases, and was unsuccessful in 15, owing to difficulties in performing catheterization or to multiple feeding arteries. In the latter cases, the tip of the catheter was placed near the origin of the feeding arteries (subselective catheterization). RESULTS: Thirty (88%) of 34 patients had a complete response to superselective arterial infusion chemotherapy and two (6%) had a partial response. Twelve (80%) of 15 patients had a complete response to subselective arterial infusion chemotherapy and three (20%) had a partial response. Local recurrence was more frequent after subselective treatment (13%) than after superselective (6%) treatment. CONCLUSION: Superselective continuous arterial infusion chemotherapy may be suitable for local control of oral cavity tumors, with a low rate of recurrence. PMID- 11110548 TI - Angiographic anatomy of the laterocavernous sinus. AB - BACKGROUND AND PURPOSE: The laterocavernous sinus (LCS) has recently been recognized as one of the major drainage pathways of the superficial middle cerebral vein (SMCV). Our purpose was to investigate the drainage pattern of the SMCV, with special emphasis on the angiographic anatomy of the LCS. METHODS: The drainage pathways of the SMCV were evaluated prospectively on 100 selective carotid angiograms obtained in 65 consecutive patients. RESULTS: The SMCV was absent in 19% of cases. A classic termination into the cavernous sinus (CS) was found in 20%, a paracavernous sinus in 39%, and an LCS in 22%. The LCS drained toward the pterygoid plexus (27%), the superior petrosal sinus (18%), the posterior aspect of the CS (32%), or a combination of these pathways (23%). A complete absence of connection between the LCS and CS was observed in 63.5% of the patients. CONCLUSION: The LCS is a laterosellar venous space that is anatomically and angiographically distinct from the CS. Secondary small anastomoses between the LCS and CS may make it difficult to differentiate the two structures. Appreciation of the course and connection pattern of the LCS is important, particularly when planning an endovascular approach to treatment of lesions in the region of the CS. PMID- 11110549 TI - The choline/creatine ratio in five benign neoplasms: comparison with squamous cell carcinoma by use of in vitro MR spectroscopy. AB - BACKGROUND AND PURPOSE: The choline (Cho)/creatine (Cr) ratio has been shown to be a reliable proton MR spectroscopy metabolic marker for differentiating squamous cell carcinoma (SCCA) from normal muscle in the upper aerodigestive tract. However, it is unclear whether the Cho/Cr ratio can be used to differentiate a malignant tumor from a benign neoplasm in the extracranial head and neck. Our purpose was to determine whether the Cho/Cr ratio can be used to differentiate benign from malignant tumors in this region. METHODS: In vitro one dimensional proton MR spectroscopy (2,000/136,272 [TR/TE]) was performed at 11 T on tissue specimens obtained from glomus tumors (n = 3), inverting papilloma (n = 1), and schwannoma (n = 1). Cho/Cr area ratios were calculated and compared with similar, previously reported in vitro (11 T) findings and with samples of SCCA and normal muscle. RESULTS: The Cho/Cr ratio was elevated in relation to muscle in all benign tumors at TE = 136 (glomus tumors = 4.52, inverting papilloma = 3.85, schwannoma = 2.2) and at TE = 272 (glomus tumors = 8.01, inverting papilloma = 2.1, schwannoma = 4.28). The average Cho/Cr ratio for benign lesions was 3.92 (TE = 136) and 6.11 (TE = 272). The Cho/Cr ratio was significantly higher in benign tumors than in both SCCA and muscle. The average Cho/Cr ratio for muscle at TEs of 136 and 272 was 1.16 and 1.31, respectively, whereas for SCCA the average Cho/Cr ratio at TEs of 136 and 272 was 1.67 and 2.45, respectively. CONCLUSION: In our small group, the Cho/Cr ratio was significantly higher in benign tumors than in muscle and SCCA of the extracranial head and neck. PMID- 11110550 TI - Echomorphologic and histopathologic characteristics of unstable carotid plaques. AB - BACKGROUND AND PURPOSE: Our hypothesis was that the carotid plaques associated with retinal and cerebrovascular symptomatology and asymptomatic presentation may be differ from each other. The aim of this study was to identify the sonographic and histopathologic characteristics of plaques that corresponded to these three clinical manifestations. METHODS: The echo process involved duplex preoperative imaging of 71 plaques (67 patients, 21 plaques were associated with retinal, 25 with cerebrovascular symptoms, and 25 were asymptomatic), which was performed in a longitudinal fashion. Appropriate frames were captured and digitized via S video signal in a computer and digitized sonograms were normalized by two echo anatomic reference points: the gray scale median (GSM) of the blood and that of the adventitia. The GSM of the plaques was evaluated to distinguish dark (low GSM) from bright (high-GSM) plaques. Subsequent to endarterectomy, the plaques were sectioned transversely, and a slice at the level of the largest plaque area was examined for the relative size of necrotic core and presence of calcification and hemorrhage. RESULTS: Retinal symptomatology was associated with a hypoechoic plaque appearance (median GSM: 0), asymptomatic status with a hyperechoic plaque appearance (median GSM: 34), and cerebrovascular symptomatology with an intermediate plaque appearance (median GSM: 16) (P = .001). The histopathologic characteristics did not disclose differences between the three clinical groups. The hypoechoic plaque appearance was associated only with the presence of hemorrhage (median GSM for the hemorrhagic plaques, 6, and for the non hemorrhagic ones, 20 [P = .04]). The relative necrotic core size and the presence of calcification did not show any echomorphologic predilection. CONCLUSION: Our results showed that distinct echomorphologic characteristics of plaques were associated with retinal and cerebrovascular symptomatology and asymptomatic status. Histopathologically, only the presence of hemorrhage proved to have an echomorphologic predilection. PMID- 11110551 TI - Intraosseous neurilemmoma of the mandible. AB - We report a rare case of intraosseous neurilemmoma of the mandible, with an emphasis on radiographic findings. The tumor, located mainly in the premolar region, presented as an expansive, unilocular, well-defined, radiolucent lesion on plain radiography. No dilatation of the mandibular canal was identified. MR imaging helped to identify the solid nature of the tumor. A biopsy was necessary to make the final diagnosis because of the relatively nonspecific nature of the lesion. PMID- 11110552 TI - Pleomorphic adenoma of the nasal septum: MR features. AB - We report the MR imaging features of a pleomorphic adenoma of the nasal septum. To our knowledge, whereas the CT findings of pleomorphic adenomas of the nasal septum have been reported, the MR features of this rare entity have not been reported in the English-language literature. PMID- 11110553 TI - Three-dimensional CT maximum intensity projections of the calvaria: a new approach for diagnosis of craniosynostosis and fractures. AB - Three-dimensional CT maximum intensity projection (MIP) can depict suture patency, extent of synostosis (ie, complete versus incomplete bone bridging), fracture extent and conspicuity, and 3D calvarial deformity as a single set of projections in children with suspected craniosynostosis or skull fracture. Three dimensional CT MIP may provide, in only eight views, all the required information to make the diagnosis of craniosynostosis and calvarial fracture extent currently requiring the combined information of 3D CT shaded surface displays and 2D axial CT images (a total of 58 views), and in some cases complementary skull radiographs. Three-dimensional MIP can be added to calvarial helical (spiral) CT imaging with only 5 minutes of additional postprocessing time. PMID- 11110555 TI - Thirty-eighth annual meeting of the American Society of Neuroradiology. PMID- 11110554 TI - Assessment of the deep gray nuclei in holoprosencephaly. AB - BACKGROUND AND PURPOSE: Although holoprosencephaly has been known for many years, few detailed analyses have been performed in a large series of patients to outline the range of morphology in this disorder, particularly regarding the deep gray nuclear structures. We reviewed a large patient cohort to elucidate the combinations of morphologic aberrations of the deep gray nuclei and to correlate those findings with recent discoveries in embryology and developmental neurogenetics. METHODS: A retrospective review of the imaging records of 57 patients (43 MR studies and 14 high-quality CT studies) to categorize the spectrum of deep gray nuclear malformations. The hypothalami, caudate nuclei, lentiform nuclei, thalami, and mesencephalon were graded as to their degree of noncleavage. Spatial orientation was also evaluated, as was the relationship of the basal ganglia to the diencephalic structures and mesencephalon. The extent of noncleavage of the various nuclei was then assessed for statistical association. RESULTS: In every study on which it could be accurately assessed, we found some degree of hypothalamic noncleavage. Noncleavage was also common in the caudate nuclei (96%), lentiform nuclei (85%), and thalami (67%). Complete and partial noncleavage were more common in the caudate nuclei than in the lentiform nuclei. The degree of thalamic noncleavage was uniformly less than that in the caudate and lentiform nuclei. Abnormalities in alignment of the long axis of the thalamus were seen in 71% of cases, and were associated with degree of thalamic noncleavage; 27% of patients had some degree of mesencephalic noncleavage. CONCLUSION: The hypothalamus and caudate nuclei are the most severely affected structures in holoprosencephaly, and the mesencephalic structures are more commonly involved than previously thought in this "prosencephalic disorder." These findings suggest the lack of induction of the most rostral aspects of the embryonic floor plate as the cause of this disorder. PMID- 11110556 TI - 2000 ASNR Gold Medal Award. American Society of Neuroradiology. PMID- 11110557 TI - 2000 ASNR presidential address. PMID- 11110558 TI - Moyamoya syndrome in cocaine-dependent patients. PMID- 11110559 TI - Anaphylaxis after steroid and local anesthetic injections. PMID- 11110560 TI - The substantia nigra is also involved in Japanese encephalitis. PMID- 11110561 TI - Publication ethics. PMID- 11110562 TI - 10-year experience with extracorporeal shockwave lithotripsy in the state of Colorado. AB - PURPOSE: To evaluate trends in the utilization of extracorporeal shockwave lithotripsy (SWL) and the potential need for medical prophylaxis of urolithaisis in the state of Colorado. MATERIALS AND METHODS: We examined patient and stone characteristics of individuals undergoing SWL for renal or upper-ureteral stones over a 10-year period (1987-1996) at the Kidney Stone Center of the Rocky Mountains. There were no significant changes in the in-state physician referral patterns nor SWL treatment criteria over this time interval. All patients were treated on the Dornier HM3 lithotripter. From September 1999 to December 1999, 198 consecutive patients undergoing SWL filled out a 10-point questionnaire regarding their interest in medical prophylaxis of urolithiasis. RESULTS: The number of patients from Colorado rose 32.5%: from 15.7 per 100,000 population in 1987 to 20.8 per 100,000 in 1996. Patient demographics such as sex, race, age, and history of nephrolithiasis did not change. Furthermore, there were no significant changes in the treated stone size or stone location. The overall increase in treatment numbers was attributable equally to increases in the number of upper ureteral and renal stones. Of the 198 patients questioned, 114 (58%) were recurrent stone formers, but only 52 (45%) of these had been offered a metabolic evaluation. CONCLUSIONS: Over the 10 years since the introduction of WSL in Colorado, there has been a gradual increase in its utilization. This higher utilization is probably multifactorial. Patients undergoing SWL have a strong desire to prevent future stone episodes and are very interested in medical prophylaxis of their stone disease. PMID- 11110563 TI - Controversial cases in endourology. PMID- 11110564 TI - An endourologic approach to complete ureteropelvic junction and ureteral strictures. AB - BACKGROUND AND PURPOSE: Complete stricture of the ureteropelvic junction (UPJ), ureter, or both represents a secondary upper tract obstruction and is a challenge for surgical management. The endourologic repair of these complete strictures remains controversial because of the many unsatisfactory results in the literature. The aim of this study was to achieve recanalization of the ureter or the UPJ using endourologic techniques to prove durable success of this technique. PATIENTS AND METHODS: We present data on the 21 patients with complete UPJ or ureteral strictures treated over 5-year period. The length of the obliterated portion of the ureter or UPJ ranged from 0.3 to 1.7 cm. The stricture was at the UPJ level in 12 patients (57%), in the upper ureter in 3, and in the lower ureter in 4. The technique was a combined approach, with antegrade introduction of the guidewire and retrograde cold-knife incision in the majority of the cases. In five cases, the incision was carried out in the reverse direction with a guidewire introduced retrograde up to the stricture level. An originally designed 6F to 7F polyethylene double-J stent with a movable 12F to 16F silicon sheath or percutaneous tube was placed at the completion of the procedure. RESULTS: The follow-up period ranged from 6 to 48 months. Recanalization was achieved in 17 patients (81%), of whom 14 became symptom free. Other surgical outcomes necessitated open surgical intervention (pyeloplasty, nephrectomy) in two patients. One patient developed a clinically significant recurrent urinary tract infection and deterioration of kidney function. Thus, the overall success rate of the endourologic management of the complete UPJ and ureteral strictures was 67% in our series. CONCLUSION: Endourologic management with retrograde or antegrade pyeloureterotomy can be successful in patients with short (up to 1.0-cm) obliterative strictures who are without extensive hydronephrosis and with preserved renal function. PMID- 11110565 TI - Successful management of lower-pole moiety ureteropelvic junction obstruction in a partially duplicated collecting system using minimally invasive retrograde endoscopic techniques. AB - Although the true incidence of ureteropelvic junction (UPJ) obstruction in the lower-pole moiety of an incompletely duplicated renal collecting system remains elusive, the description of this entity in the published literature is exceedingly rare. To our knowledge, we report the first case of this entity managed successfully by ureteroscopic holmium laser incision of the stenotic UPJ segment. This case underscores the utility of minimally invasive techniques in the management of selected cases of UPJ obstruction associated with a partially duplicated collecting system. PMID- 11110567 TI - Nephrostomy tube after percutaneous nephrolithotomy: large-bore or pigtail catheter? AB - BACKGROUND AND PURPOSE: A nephrostomy tube is an integral part of any percutaneous renal surgery. Commonly, a nephrostomy tube that is 2F to 3F smaller than the percutaneous tract is used after percutaneous nephrolithotomy (PCNL). In our experience, quite a few patients have pain at the nephrostomy tube site, and many patients complain of a prolonged urinary leak after tube removal when a large nephrostomy tube is used. This prospective study was planned to document whether these symptoms could be attributed to the size of the nephrostomy tube and whether a small pigtail catheter could reduce these problems without increasing complications. PATIENTS AND METHODS: Forty well-matched patients in whom a one-stage PCNL was done for calculus disease were studied prospectively. Alternate patients had a 28F nephrostomy tube or a 9F pigtail catheter placed at the end of the procedure. Patients were observed for the duration of hematuria, number of analgesic injections needed, and the duration of urinary leak after tube removal. RESULTS: The groups were comparable in the amount and duration of hematuria after PCNL. There was a statistically significant difference in the analgesic need and the duration of urinary leak after tube removal, both of which were less in patients having a pigtail catheter. CONCLUSIONS: A pigtail catheter nephrostomy tube after PCNL reduces the hospital stay by reducing the duration of the urinary leak. The postoperative course is smooth, as patient has less pain and needs less analgesic support. There is no statistically significant increase in the postoperative bleeding secondary to use of a pigtail catheter. Second-look nephroscopy was easy in the one patient with a pigtail nephrostomy catheter who needed the procedure. PMID- 11110566 TI - Retrograde endopyelotomy using an original home-made diathermy probe. AB - PURPOSE: To describe an easily home-made diathermy probe for the performance of retrograde endopyelotomies using a small-caliber rigid ureteroscope. METHODS: The diathermy probe is easily built by putting a rigid guidewire through a 5F ureteral catheter, the distal tip being naked and slightly bent. The other end is linked to the electric generator. Among a total of 24 retrograde endopyelotomies performed to treat ureteropelvic junction obstruction in the last 7 years, five were done using this device. RESULTS: Surgery with the probe took an average of 30 minutes. There were no complications, and, as of today, all cases are successes. CONCLUSION: This device allows the performance of retrograde endopyelotomy using a small-caliber ureteroscope. Long-term results will presumably match the good results obtained using the larger-caliber ureteroscopes with the classic cold knife. PMID- 11110568 TI - Antegrade ureteral stent placement: positioning without use of a retraction string. AB - PURPOSE: To evaluate a simple method of antegrade ureteral stent insertion allowing optimal positioning of the stent without the use of a retraction string. PATIENTS AND METHODS: Seventeen stents were placed in sixteen patients with ureteral obstruction. Materials included a long vascular introducer sheath and radiopaque markers on the tips of both the sheath and the pusher catheter. For optimal positioning of the proximal pigtail in the renal pelvis, the distal end of the sheath was used to hold a large portion of the pigtail in the extended state prior to its deployment. RESULTS: All stent placements were successful. In one case, the tip of the proximal pigtail was caught in a lower-pole calix. In another case, repeat stent placement was necessary because of recurrent stricture several months after removal of the first stent. All stents functioned properly, as demonstrated by follow-up nephrostography 2 or 3 days after each procedure. CONCLUSION: The insertion method we describe is simple, easy to perform, and fast and avoids the risks associated with the use of a retraction string. PMID- 11110569 TI - Liposome-coated metal stents: an in vitro evaluation of controlled-release modality in the ureter. AB - PURPOSE: In vitro preparation of liposome-covered metal stents and loading of liposomal drug formulations that will slowly release the drug in the vicinity of the stent. MATERIALS AND METHODS: Polytetrafluoroethylene-coated stents were used. Large multilamellar (MLV) liposomes (phosphatidylcholine:cholesterol 1:1 mol/mol), empty or entrapping the corticosteroid anti-inflammatory drug, dexamethasone, were prepared by the thin-film hydration method and applied to pieces of stent using a simple and mild evaporation technique. Initially, a freeze-drying method for applying liposomes to stents was also evaluated, but it failed to produce stents that efficiently retain liposomal lipid when incubated in an aqueous environment. The presence of liposomes on the stent surface was confirmed by scanning electron microscopy. RESULTS: After analyzing the release of liposomal lipid (using a phospholipid assay) and liposomal drug (by a modified dexamethasone high-pressure liquid chromatography method) in an in vitro system developed to simulate in vivo conditions, it was found that 39.11+/-6.8% of the lipid and 50.84+/-5.48% of the drug was released from the stent pieces during 48 hours of incubation in the presence of artificial urine. The amount of dexamethasone released from stents during their application procedure was found to be negligible in an in vitro dry run. CONCLUSION: The use of stent-associated liposomal drug formulations as slow-release depots could be an efficient method of treating the untoward event of ureteral stent obstruction. PMID- 11110570 TI - Vesicourethral anastomosis during laparoscopic radical prostatectomy: the running suture method. AB - Vesicourethral reconstruction is the most critical and time-consuming step of laparoscopic radical prostatectomy. We describe the use of two hemicircumferential running sutures that has significantly simplified the procedure in our last 30 patients. The vesicourethral reconstruction took 31 minutes on average. Six months postoperatively, 84% of the patients were fully continent, and no bladder neck stenosis had occurred. The economy of intracorporeal suturing provided by this novel method, together with geometric factors such as the optimal position of the trocars, contributes to the improvement of ergonomy, allowing the surgeon to decrease operating times. PMID- 11110571 TI - Open laparoscopic access using a radially dilating trocar: experience and indications in 50 consecutive cases. AB - BACKGROUND: Laparoscopy can be performed using needle access for initial insufflation or open access. PATIENTS AND METHODS: A technique for open laparoscopic access to the abdomen using a radially dilating cannula was used in 52 operations in 50 patients. Indications included age <2 years, severe kyphosis, and creation of an umbilical stoma. RESULTS: There was one case of minor leakage of carbon dioxide that did not affect the procedure being performed. There were two cases of preperitoneal placement, which were recognized immediately; in both, peritoneal access was easily obtained. CONCLUSION: Open laparoscopic access is safely and easily performed with a radially dilating trocar. This is the preferred technique at our institution for patients who meet the criteria for open access. PMID- 11110572 TI - Holmium laser resection v transurethral resection of the prostate: results of a randomized trial with 2 years of follow-up. AB - BACKGROUND AND PURPOSE: The holmium laser (2140 nm) can be used to ablate, resect, and enucleate the enlarged prostate. The 2-year results of a randomized trial comparing holmium laser resection of the prostate (HoLRP) and transurethral resection (TURP) are presented. PATIENTS AND METHODS: The 120 patients were randomized to either TURP (N = 59) or HoLRP (N = 61). The patients were reviewed at 1, 3, 6, 12, 18, and 24 months postoperatively. Eighty six (72%) of the patients were available for review at the 2-year mark. RESULTS: At 2 years, there was no significant difference between the two groups in AUA Symptom Score, peak flow rate (Qmax) value, or quality of life score. Adverse events, including reoperations, incontinence, and loss of erectile potency, were also similar. CONCLUSIONS: The HoLRP and TURP procedures result in similar clinical outcomes at 2 years. PMID- 11110573 TI - Correlation between central zone perfusion defects on gadolinium-enhanced MRI and intraprostatic temperatures during transurethral microwave thermotherapy. AB - BACKGROUND AND PURPOSE: The likelihood of success of thermoablation of prostatic hyperplasia depends on delivering an optimal thermal dose, but data on the temperatures achieved with these methods are few. We sought to develop a noninvasive method for monitoring intraprostatic heat distribution. PATIENTS AND METHODS: Thirteen patients ranging from 50 to 76 (mean 61.3+/-8.1) years were enrolled in this study, all of whom had evidence of obstruction by uroflowmetry and pressure-flow studies. The mean total volume of the gland was 40.3+/-13.1 cc, while the mean adenoma volume was 20.4+/-10.1 cc, as estimated by preoperative transrectal ultrasonography. All the patients were treated with the Urologix Targis device for at least 45 minutes. Continuous temperature mapping was performed during the therapy using spatially dispersed thermosensors at 16 prostatic sites. The patients were evaluated 5 to 12 days postoperatively with MRI of the prostate utilizing a pelvic phased-array coil at 1.5 T. RESULTS: Postprocedure MRI demonstrated a mean perfusion defect of 28.1+/-2.1% and 63.6+/ 34% of the total gland and transition zone volumes, respectively. The mean anteroposterior (AP) and transverse diameters of the perfusion defects, as measured on the MRI images, were 29.2+/-5.2 mm and 32.7+/-5.9 mm, respectively. The maximum mean peak temperatures were 66.8+/-13 degrees C and were recorded at 4 mm from the urethra. No temperatures higher than 45 degrees C were recorded beyond 15 mm on either side of the urethra in the AP direction and beyond 16 mm on either side of the urethra in the transverse diameter. This perfusion defect was persistent for 27.7+/-5.2 mm in the superoinferior diameter, which is equivalent to the length of the antenna (28 mm). CONCLUSION: Perfusion defect diameters as measured by postprocedure MRI accurately represent the prostatic tissues exposed to temperatures of > or =45 degrees C for 45 minutes or more. So, MRI provides an accurate, noninvasive method for screening the effective heat pattern generated in the prostate during transurethral microwave thermotherapy. PMID- 11110574 TI - Controlling indoor allergens. PMID- 11110575 TI - Soluble CD30 and CD23 in cord blood are not related to atopy in early childhood. AB - Atopic disease, including atopic dermatitis (AD), is associated with a T-helper 2 (Th2)-dependent immune response. The cytokine receptor CD30 appears to be preferentially expressed on, and its soluble form (sCD30) released by, Th2 cells. Therefore, sCD30 may be a potential marker for atopic disorders. The aim of this study was to test the hypothesis that the sCD30 level in cord blood could be used to predict the development of atopy or AD in early childhood. In a case-control study, levels of sCD30, as well as soluble low-affinity immunoglobulin E (IgE) receptor (sCD23), interleukin-4 (IL-4) and IgE, were measured in cord blood in 35 children who subsequently developed allergic sensitization and AD before the age of three, and the results were compared to those of 35 matched children without a history of atopy. There was no difference in cord blood levels of sCD30 between controls (32.5 U/ml; 19.7-80.1) and children with subsequent atopy and AD (32.2 U/ml; 22-75.9) (median; quartiles). The concentration of sCD30 showed no relation to the levels of total IgE, sCD23 or IL-4. Levels of sCD23 were similar in children with subsequent atopy (60.2 U/ml; 44.5-76.8) and controls (55.2 U/ml; 38.3-72.5), whereas IL-4 was detectable in 10 of the atopic children and in only two of the controls (p <0.05). In conclusion, cord blood levels of sCD30 or sCD23 do not seem to be related to the subsequent development of atopy or AD at the age of three. PMID- 11110576 TI - Wheezing in early life and asthma at school age: predictors of symptom persistence. AB - Early childhood wheezing is associated with asthma later in life. However, the high spontaneous recovery rate and the lack of firm predictors for persistence of wheezing complicates the development of evidence-based guidelines for long-term management of wheezy infants and toddlers. Our aim was to define variables that could be used to identify wheezy individuals younger than 3 years of age who would continue to be symptomatic at school age. The method used was a questionnaire-based cross-sectional survey of 2,027 randomly chosen, 6-13-year old school children. Altogether 1,829 (90%) questionnaires were returned. Emergency medical care had been sought for 186 (10.2%) children for wheezing during the first 3 years of life, and only 17.2% of these children had received similar emergency treatment during the 12 months preceding the survey. The total proportion of children with current asthma at school age was 11.4%. A logistic regression analysis indicated that for the early wheezers, a family history of asthma, an itchy rash or food allergy, and exposure to tobacco smoke at home before the age of 3 years, were all independently associated with symptom persistence until school age. Among all wheezy children younger than 3 years, those who have a history of food allergy, itchy rash, asthma occurrence in a sibling or parent, or are exposed to tobacco smoke during the first years of life are at highest risk for symptom persistence until school age. PMID- 11110577 TI - Airway nitric oxide in infants with acute wheezy bronchitis. AB - Concentrations of nitric oxide (NO) in exhaled air are increased in children and adults with asthma, and NO measurements are used as a non-invasive marker to monitor airway inflammation in these patients. To define the role of NO in infants with acute wheezy bronchitis, we measured nasal and end-tidal NO concentrations in 17 infants with acute virus-associated wheezy bronchitis, in 22 term infants without respiratory disease, and in nine premature infants. Nasal NO measurements were performed with an olive placed in the infant's nose; end-tidal NO concentrations were assessed during tidal breathing through a snuggly fitting face mask. Both end-tidal NO concentrations and nasal NO concentrations were reduced in infants with acute wheezy bronchitis. There were no differences in NO concentrations between term infants and premature infants. Measurements by both techniques were highly reproducible, as assessed by repeated measurements three times daily on three consecutive days in eight premature infants. Reduced airway NO concentrations in infants with virus-associated acute wheezy bronchitis are in contrast to findings in adults where both upper and lower airway NO levels are increased in patients with asthma. Whether this reflects a different inflammatory reaction to upper airway infections in acutely wheezy infants or pathophysiologic differences in airway response remains to be determined. PMID- 11110578 TI - Acute childhood asthma in Finland: a retrospective review of hospital admissions from 1976 to 1995. AB - The prevalence of childhood asthma has increased markedly in many Western societies during recent decades. We wanted to study whether the incidence and severity of childhood asthma in Finland had changed during the time-period 1976 95. Hospital admission rates from 1976 to 1995 were obtained from the National Hospital Discharge Register and the individual intensive care unit (ICU) registers of the five university hospitals in Finland. The number and length of treatment periods for childhood asthma in all Finnish hospitals and at the ICUs of the five university hospitals were analyzed. The number of children receiving special reimbursement for asthma medication costs was obtained from the central register of the Social Insurance Institution. The data showed that during the time-period investigated, hospital admissions as a result of asthma had increased by 2.8-fold, but the mean length of hospital stay had more than halved (from 7.3 to 2.6 days). The increase in hospital admissions showed greatest significance in the 0-4-year age-group among both sexes (p <0.001). In contrast, a significant reduction in hospital admissions was found among the 10-14-year age-group (p <0.001). No discernible change in admission to ICUs was seen. During the same time-period, the number of children receiving special reimbursement for asthma medication costs increased 7.5-fold. Hence, a major increase has occurred in the number of children diagnosed with asthma that has not been paralleled by a proportionate increase in the number of hospital admissions. While the prevalence of mild and moderate asthma has increased, the occurrence of severe asthma has remained essentially unchanged. PMID- 11110579 TI - Intervention models on psycho-social health in families with an asthmatic child. AB - Despite an increase in the prevalence of asthma during the last few decades, the need for hospital treatment of children with asthma has become less. One reason for this is that children and their parents are now more involved in the treatment of the disease, and responsibility has been shifted from the medical care system to the family. This new responsibility may cause increased psycho social tension within the family. We conducted a pilot study on three limited methods of intervention to find the best way to help families in this respect. All three methods (individual family meetings, family group meetings, and evaluation of the child's environment in school) reduced the psychosocial burden of having a child with asthma. This indicates that families should be supported by being given the opportunity to participate in meetings to discuss the disease or to have the environment in the child's school evaluated, in addition to receiving regular medical care. PMID- 11110580 TI - Diagnosis of food allergy in Finland: survey of pediatric practices. AB - Food-related symptoms are common in the first years of life, and food allergy should be diagnosed using an elimination challenge test. We surveyed Finnish hospital-based pediatricians using a self-completion questionnaire to ascertain the current clinical practice: 24 of the 25 pediatricians (representing 24 of 25 hospitals) so approached gave evaluable responses. Food allergies were diagnosed using a clinical elimination challenge test in patients with suspected allergy to cow's milk or cereals (wheat, rye, barley, oats). Of the 24 departments, four reported that they performed challenge in all patients before diagnosis was confirmed, and 14 performed challenge in most patients before diagnosis was confirmed. The duration of the challenge varied from 0.5 to 7 days (median 4 days). A 1-week challenge was used in eight hospitals. The double-blind placebo controlled challenge was used in seven of the hospitals, and in none routinely. Altogether, 16 of the respondents agreed that there is a need to establish clinical guidelines for the diagnosis of food allergy. In conclusion, despite a long tradition of medical education on the subject of food allergy, practices vary for its diagnosis. There is therefore a requirement for appropriate clinical guidelines. PMID- 11110581 TI - Anti-centromere antibodies as a marker of Raynaud's phenomenon in pediatric rheumatologic diseases. AB - To examine the possible relationship between anti-centromere antibodies (ACA) and pediatric rheumatologic diseases, we investigated the presence of ACA (using enzyme immunoassay) in the sera of 45 children and adolescents with such diseases and compared the results with a group of 42 age- and gender-matched healthy subjects. ACA were present ( > or =10 U/ml) in three out of five patients (60%) with scleroderma (SCD), in seven out of 16 (43.8%) patients with systemic lupus erythematosus (SLE), in two out of five patients (40%) with mixed connective tissue disease (MCTD), in one out of four patients (25%) with dermatomyositis (DMS), and in two out of 14 patients (14.3%) with juvenile rheumatoid arthritis (JRA). ACA were also detected in a single patient with anti-phospholipid syndrome (APL) who had digital gangrene and hemiparesis, as well as in two healthy subjects. ACA positivity was related to the presence of Raynaud's phenomenon in the studied sample, as 86% of patients suffering from the phenomenon were ACA positive. ACA positivity was associated with older age, high blood pressure and high erythrocyte sedimentation rate (ESR) values, and lower hemoglobin and weight and height percentile values. It was also higher among anti-nuclear antibody positive subjects. Raynaud's phenomenon and ACA positivity shared almost the same clinical and laboratory associations in the studied patients. Thus, ACA are probably among the markers of Raynaud's phenomenon in pediatric rheumatologic diseases. Their value as predictors of future development of the phenomenon needs further evaluation. PMID- 11110582 TI - Nasal peak inspiratory flow through Turbuhaler in children with symptomatic rhinitis and in healthy children. AB - Topical treatment of allergic or vasomotor rhinitis is possible by means of pressurized metered dose inhalers, aqueous spray, or dry powder inhalers. In children, little is known about nasal drug delivery by dry powder inhalation. The airflow through the device is critical for the drug release and a sufficient nasal inspiratory flow is needed for intranasal drug delivery from a dry powder inhaler. In order to investigate from what age children with allergic or vasomotor rhinitis can reliably use such a device, device-dependent nasal peak inspiratory flow (DnPIF) was measured. The maximal DnPIF was measured in children aged 4-13 years making use of a dry powder inhaler (Turbuhaler) connected to a spirometer (Vitalograph). In the clinically relevant context, instructions from the doctor and one week's use of a Turbuhaler at home were found to be sufficient to obtain a good inhalation technique and were shown to improve DnPIF at least as effectively as visual feedback training at the clinic. Children with rhinitis, as well as healthy children from the age of 6 years, were able to generate a DnPIF sufficient to obtain a reliable nasal delivery of a dry powder drug dose. DnPIF values correlated with age. Consequently, a recommendation to use a nasal Turbuhaler from the age of 6 for topical drug delivery in the treatment of allergic or vasomotor rhinitis seems reasonable. PMID- 11110583 TI - Snail anaphylaxis during house dust mite immunotherapy. AB - This study reports a 12-year-old girl who developed an anaphylactic reaction following snail ingestion during house dust mite (HDM) immunotherapy treatment. Radioallergosorbent (RAST) inhibition showed cross-reactivity between the two allergens, leading to consideration of HDM as the sensitizing agent. Children undergoing HDM immunotherapy treatment should be aware of the potential risks of hypersensitivity reactions to invertebrate foods. PMID- 11110584 TI - The Mediterranean diet revisited--towards resolving the (French) paradox. PMID- 11110585 TI - Neurological potassium channelopathies. AB - Potassium channel dysfunction has been implicated in a variety of genetic and acquired neurological disorders that are collectively referred to as the potassium channelopathies. These include acquired neuromyotonia, episodic ataxia type-1, hereditary deafness syndromes, benign familial neonatal convulsions and hypokalaemic periodic paralysis. Insight into potassium channel structure and function is crucial to understanding the pathophysiology of these conditions. This article describes potassium channel structure and function and then outlines what is known about the immunology and genetics of the neurological potassium channelopathies. PMID- 11110586 TI - Long-term follow-up of patients presenting to adult nephrologists with chronic pyelonephritis and 'normal' renal function. AB - We studied the natural history, and therefore prognosis, of patients with chronic pyelonephritis presenting to adult nephrologists with a plasma or serum creatinine <90 mmol/l. From the Newcastle chronic pyelonephritis database, 255 patients with radiologically-proven disease were reviewed. Median follow-up was 95 months (95%CI 82. 3-109.3). Plasma creatinine was < or =90 micromol/l (P(Cr)< or =90 group) at presentation in 138. At presentation, hypertension, bilateral disease and proteinuria were less frequent in the P(Cr)< or =90 group (hypertension 19% vs. 32%, p<0.05; bilateral disease 25% vs. 70%, p<0.001; proteinuria 18% vs. 60%, p<0.001). With the exception of two patients, the renal prognosis of this group was excellent. Patients over the age of 18 years presenting to adult nephrologists with a diagnosis of chronic pyelonephritis and a creatinine < or =90 micromol/l can be reassured that the chances of developing end-stage renal failure in the future are very small. Most could be referred back to their general practitioner for long-term follow-up. PMID- 11110587 TI - Can clinical assessment of chest pain be made more therapeutic? AB - We describe the referral and management of consecutive patients attending a cardiac service with the presenting complaint of chest pain. Of 610 consecutive new referrals to five Oxford cardiac clinics over 12 weeks, 202 had chest pain as the presenting complaint: 91 (45%) angina, 101 (50%) non-cardiac chest pain, 8 (4%) both and 2 (1%) uncertain diagnosis. Information in clinic letters was sometimes ambiguous and contradictory and suggested a lack of precise information to patients. Patients with non-cardiac chest pain often had long histories, including considerable previous use of services and specialist investigations. There were delays in referral and assessment of patients. There are opportunities for simple changes in assessment procedures which might have substantial advantages for outcome and resource: (i) more detailed referral information from general practitioners, with an explicit statement of the reasons for referral; (ii) minor modifications to augment the assessment by provision of unambiguous information to patients and primary care at discharge. PMID- 11110588 TI - Fetal and maternal outcomes in Indo-Asian compared to caucasian women with diabetes in pregnancy. AB - Maternal and fetal complications are increased when pregnancy is complicated by diabetes, and this may be further influenced by racial and cultural differences. We examined fetal and maternal outcomes in Indo-Asian and Caucasian women attending the same antenatal diabetes service to see if there were any differences. Women with diabetes mellitus (type 1, type 2 and gestationally acquired disease) complicating pregnancy, registered at the combined diabetes/antenatal clinic of this University teaching hospital over the period 1990-1998 were included. Fetal outcomes examined were miscarriage <24 weeks, stillbirths, neonatal deaths up to 28 days of life, perinatal mortality, congenital malformations and size for gestational age. Maternal outcomes examined were rates of caesarean section and vaginal deliveries, and number of pre-term deliveries <37 completed weeks of gestation. Outcomes for Indo-Asian and Caucasian women were similar, with a take-home baby rate of 96% and 92%, respectively. There was no perinatal mortality in Indo-Asian women, who were more likely to have a vaginal delivery and less likely to have a baby large for gestational age. Pregnancies complicated by type 2 diabetes in both groups pose the greatest threat to a successful pregnancy outcome. Indo-Asian and Caucasian women attending the same antenatal diabetes service have comparable outcomes. Attendance for pre-pregnancy care needs to be encouraged to combat the high early pregnancy loss and congenital malformation rate identified, particularly in those with type 2 disease, irrespective of ethnicity. PMID- 11110589 TI - Tumour necrosis factor alpha in the diagnostic assessment of pleural effusion. AB - We investigated the role of tumour necrosis factor-alpha (TNF) in the evaluation of pleural effusion aetiology. Using a commercially-available ELISA kit, concentrations of TNF were measured in the serum and pleural fluid of patients with malignant effusions (n=19), uncomplicated parapneumonic effusions (n=13), and exudative (n=13) and transudative (n=13) effusions due to congestive heart failure (CHFex and CHFtr, respectively). Serum TNF did not differ significantly between the four groups (p>0.05). In the group with malignancy, pleural fluid TNF was significantly higher than in the other groups (p<0.001), which were not significant different from each other (p>0.05). However, a considerable overlap between all four groups was found. Pleural fluid TNF was significantly higher than serum TNF in the malignant and the uncomplicated parapneumonic groups (p<0.001), and there was a significant positive correlation between serum TNF and pleural fluid TNF in the group with uncomplicated parapneumonic effusion (r=0.7, p<0.005), in the group with CHFex (r=0.54, p<0.01), and in the group with CHFtr (r=0. 8, p<0.005), but not in the group with malignancy. Pleural fluid TNF:serum TNF (TNF ratio) was significantly higher in the malignancy group than in the other groups (p<0.001); no significant difference was found between the other three groups (p>0.05). At an optimal cut-off point of 2.0 for TNF ratio, determined by ROC analysis for discrimination between malignant and non-malignant groups, sensitivity was 84%, specificity 90%, and total accuracy 88% (p<0. 0001). TNF ratio might be helpful in the diagnostic assessment of exudative pleural effusion. PMID- 11110590 TI - Immune cytopenias as the presenting finding in primary Sjogren's syndrome. AB - A diagnostic delay of several years in primary Sjogren's syndrome is common, even in patients who present with sicca symptoms. It is much more likely in cases with prominent symptomatic extraglandular involvement. We report on three such patients who presented as Coomb's positive haemolytic anaemia, systemic symptoms with agranulocytosis and gingival bleeding due to immune thrombocytopenia, to alert clinicians to the fact that primary Sjogren's syndrome may present as clinically significant immune-mediated cytopenia in the absence of sicca symptoms. Sjogren's syndrome, a common autoimmune disorder, should be considered in the differential diagnosis of apparently 'idiopathic' cytopenias and actively sought by directed history, Schirmer test and autoantibody screening. PMID- 11110591 TI - Duty and the beast: animal experimentation and neglected interests. PMID- 11110592 TI - Status epilepticus treated with a muscle relaxant: the first success. AB - In 1958, an 11-year-old girl with status epilepticus was given the current treatments which failed to control the convulsions. In order to stop the fits, protect the airway, prevent hypoxia and hyperpyrexia, intermittent positive pressure ventilation (IPPV) and complete muscle paralysis with d-tubocurarine was used for a total of 6 h. The girl made a complete recovery, the first patient to do so using this plan of action. PMID- 11110593 TI - Why I volunteer for the american cancer society PMID- 11110594 TI - Pediatric oncology: regulatory initiatives. AB - The majority of children with cancer receive therapy as participants in clinical research protocols coordinated by national pediatric cooperative groups. One of the highest priorities of these groups is the development of novel therapies. Due to differences in the biology of pediatric and adult tumors and in physiology between adults and children, it is usually necessary to evaluate the safety and effectiveness of new drugs separately in adult and pediatric populations. To stimulate the development of new therapies for pediatric indications and encourage the submission of clinical data to support pediatric product labeling, the U.S. Food and Drug Administration has undertaken two initiatives. In 1998 the FDA issued a regulation (The 1998 Final Pediatric Rule) that mandated if a drug or biological is under review for a claim and the disease exists in both pediatric and adult populations, pediatric studies must be performed. Section 111 of the FDA Modernization Act of 1997 states that if a drug product has exclusivity based on a patent or marketing license, the exclusivity can be extended by six months for the submission of pediatric data to the FDA. The incentive applies to both approved drugs and those under development. These initiatives are intended to enhance access to new anticancer therapies and promote labeling for pediatric oncology indications. PMID- 11110595 TI - Childhood cancers: hepatoblastoma. AB - Hepatoblastoma is the most common primary liver tumor in children, accounting for just over 1% of pediatric cancers. The etiology is unknown, but it has been associated with Beckwith-Weidemann syndrome, familial adenomatosis polypi, and low birth weight. The primary treatment is surgical resection, however, chemotherapy plays an important role by increasing the number of tumors that are resectable. The prognosis for patients with resectable tumors is fairly good, however, the outcome for those with nonresectable or recurrent disease is poor. PMID- 11110596 TI - Induction of differentiation and apoptosis- a possible strategy in the treatment of adult acute myelogenous leukemia. AB - A differentiation block with accumulation of immature myeloid cells characterizes acute myelogenous leukemia (AML). However, native AML cells often show some morphological signs of differentiation that allow a classification into different subsets, and further differentiation may be induced by exposure to various soluble mediators, e.g., all trans-retinoic acid (ATRA) and several cytokines. Combination therapy with ATRA and chemotherapy should now be regarded as the standard treatment for the acute promyelocytic leukemia variant of AML. Several agents can induce leukemic cell differentiation for other AML subtypes, although these effects differ between patients. Differentiation may then be associated with induction of apoptosis, and differentiation-inducing therapy may therefore become useful in combination with intensive chemotherapy to increase the susceptibility of AML blasts to drug-induced apoptosis. However, it should be emphasized that differentiation and apoptosis can occur as separate events with different regulation in AML cells, and future studies in AML should therefore focus on: A) the identification of new agents with more predictable effects on differentiation and apoptosis; B) the use of clinical and laboratory parameters to define new subsets of AML patients in which differentiation/apoptosis induction has a predictable and beneficial effect, and C) further characterization of how AML blast sensitivity to drug-induced apoptosis is modulated by differentiation induction. PMID- 11110597 TI - Management of bone metastases. AB - Metastatic bone disease develops as a result of the many interactions between tumor cells and bone cells. This leads to disruption of normal bone metabolism, with the increased osteoclast activity seen in most, if not all, tumor types providing a rational target for treatment. The clinical course of metastatic bone disease in multiple myeloma, breast and prostate cancers is relatively long, with patients experiencing sequential skeletal complications over a period of several years. These include bone pain, fractures, hypercalcemia, and spinal cord compression, all of which may profoundly impair a patient's quality of life. External beam radiotherapy and systemic endocrine and cytotoxic treatments are the mainstay of treatment in advanced cancers. However, it is now clear that the bisphosphonates provide an additional treatment strategy, which reduces both the symptoms and complications of bone involvement. Additionally, new specific molecules such as osteoprotogerin have been developed that are based on our improved understanding of the cellular signaling mechanisms involved in cancer induced bone disease. These potent molecules are now entering clinical trials. Ongoing research is aimed at trying to define the optimum route, dose, schedule and type of bisphosphonate in metastatic bone disease and its use in the prevention and treatment of osteoporosis in cancer patients. In vitro suggestions of direct anticancer activity and some promising clinical data in early breast cancer have resulted in considerable interest in the possible adjuvant use of bisphosphonates to inhibit the development of bone metastases. PMID- 11110598 TI - The initial results in muscle-invading bladder cancer of RTOG 95-06: phase I/II trial of transurethral surgery plus radiation therapy with concurrent cisplatin and 5-fluorouracil followed by selective bladder preservation or cystectomy depending on the initial response. AB - PURPOSE: To assess the safety, tolerance, and efficacy of transurethral surgery plus concomitant cisplatin, 5-fluorouracil (5-FU), and radiation therapy in conjunction with selective bladder preservation in patients with muscle-invading bladder cancer. Patients and Methods. Thirty-four eligible patients with clinical stage T2-T4a, Nx M0 bladder cancer without hydronephrosis were entered into a protocol aimed at selective bladder preservation. Treatment began with as complete a transurethral resection as possible followed by induction chemoradiation. This consisted of cisplatin 15 mg/m(2) i.v. and 5-fluorouracil (5 FU) 400 mg/m(2) i.v. in the mornings on d 1, 2, 3, 15, 16, and 17. On d 1, 3, 15, and 17, radiation was given immediately following the chemotherapy using twice-a day 3 Gy per fraction cores to the pelvis for a total radiation dose of 24 Gy. Response was evaluated by cystoscopy, cytology, and rebiopsy four weeks later. Patients with a complete response received consolidation therapy with the same drugs and doses on d 1, 2, 3, 15, 16, and 17 combined with twice-daily radiation therapy to the bladder and bladder tumor volume of 2.5 Gy per fraction for a total consolidation dose of 20 Gy and a total induction plus consolidation dose to the bladder and bladder tumor of 44 Gy. Patients who did not achieve a complete response were advised to undergo prompt cystectomy, as were those with a subsequent invasive recurrence. The median follow up is 29 months. RESULTS: Of the 34 eligible patients, 26 had a visibly complete transurethral resection. One patient did not complete induction treatment due to acute hematologic toxicity. After induction treatment, 22 (67%) of the 33 patients had no tumor detectable on urine cytology or rebiopsy. Of the 11 patients who still had detectable tumor, six underwent radical cystectomy and five underwent consolidation chemoradiation (one because of refusal to have the recommended cystectomy and four because the treating institutions erroneously assigned them to receive consolidation chemoradiation rather than cystectomy). No patient has required a cystectomy for radiation toxicity. Six patients have died of bladder cancer. The actuarial overall survival at three years is 83%. The probability of surviving with an intact bladder is 66% at three years. A total of seven patients (21%) developed grade 3 or grade 4 hematologic toxicity in conjunction with this treatment. CONCLUSION: This aggressive protocol comprising local surgery plus concurrent 5 FU, cisplatin, and high-dose hypofractionated radiation has been associated with moderately severe hematologic toxicity. Longer follow-up will be necessary to assess efficacy. Both the 67% complete response rate to induction therapy and the 66% three-year survival with an intact bladder are encouraging. PMID- 11110599 TI - Malignancy in neurofibromatosis type 1. AB - Neurofibromatosis type 1 (NF1) represents a major risk factor for development of malignancy, particularly malignant peripheral nerve sheath tumors (MPNST), optic gliomas, other gliomas, and leukemias. The oncologist will see NF1 patients referred for treatment of malignancy, and should be alert to the possibility of undiagnosed NF1 among patients with cancer. Brain tumors tend to have a more indolent course in NF1 than in the general population, and hence are best managed conservatively. MPNST, in contrast, do not respond to standard chemotherapy or radiation therapy. The most effective treatment of MPNST appears to be early diagnosis and surgery, but early diagnosis is hampered by frequent occurrence within preexisting large tumors, making new growth or change difficult to detect. New insights into pathogenesis now offer hope of development of specific methods of treatment with reduced toxicity and more precise molecular targeting. There is an urgent need, however, to develop methods to measure tumor growth and monitor outcomes, develop preclinical drug screening systems, and further explore the pathogenesis of the disorder to determine whether mechanisms other than Ras regulation may be important in pathogenesis. PMID- 11110600 TI - Nonmyeloablative allogeneic stem cell transplantation: early promise and limitations. AB - Allogeneic stem cell transplantation is used to treat a variety of malignant and nonmalignant hematologic diseases. Conventionally, high-dose chemoradiotherapy based preparative regimens were considered essential both for tumor eradication and facilitation of donor stem cell engraftment. It is now apparent that an immune-mediated graft-versus-tumor effect has a pivotal role in the curative potential of allogeneic stem cell transplantation. This has prompted the development of less toxic, nonmyeloablative but profoundly immunosuppressive preparative regimens, often fludarabine- or radiation-based. Full donor engraftment can be achieved; however, a significant number of patients achieve a mixed chimeric state. Mixed hematopoietic chimerism provides a platform for the use of adoptive immunotherapy using donor lymphocyte infusions to maximize the immune-mediated antitumor effect, but the optimal usage has yet to be determined. Immediate procedure-related mortality with nonmyeloablative regimens has been low, but graft-versus-host disease remains a major clinical concern and treatment challenge. Major tumor responses have been seen in many hematologic malignancies primarily including patients with highly chemorefractory disease. Follow-up data have been short and additional time is needed to determine the efficacy and toxicities of this immunotherapy. This approach has potential for widespread clinical application including HLA mismatched and matched unrelated donor transplantation, exploration of a graft-versus-solid tumor effect, and correction of phenotypic expression in nonmalignant disorders. PMID- 11110601 TI - Biological concepts of prolonged neoadjuvant treatment plus GM-CSF in locally advanced tumors. AB - Local treatment with surgery and radiotherapy gives unsatisfactory results in patients with locally advanced cancer. In many cases distant metastases appear shortly after the removal of the primary tumor. Selecting breast cancer as a model for locally advanced disease, we are extrapolating our findings to other solid tumors. Neoadjuvant chemotherapy has improved survival of these patients by downstaging the primary tumors allowing local treatment and early elimination of distant micrometastases. We recently reported in this journal on a study of 42 patients with locally advanced breast cancer (LABC) who received prolonged neoadjuvant chemotherapy of doxorubicin, cyclophosphamide, and GM-CSF prior to surgery and postoperative radiotherapy. These results were promising and prompted us to initiate an international randomized phase III study in which either six neoadjuvant cycles or three neoadjuvant cycles plus three adjuvant cycles are being compared. In LABC patients treated with six neoadjuvant chemoimmunotherapy cycles, we observed a significant rise in the dendritic cell content of the axillary tumor-draining lymph nodes after therapy, associated with an encouraging disease free survival and overall survival. We hypothesize that the prolonged presence of draining lymph nodes in combination with the repeated tumor antigen release, dendritic cell recruitment, and activation may account for the observed increased survival of LABC patients. Based on our findings and the results of preclinical studies, we hypothesize that it is more effective to administer chemotherapy in an extended neoadjuvant regimen, taking advantage of the concurrent biological and immunological processes in the primary tumor and its draining lymph nodes. PMID- 11110602 TI - Uncovering functionally relevant signaling pathways using microarray-based expression profiling. AB - The introduction of microarray technology to the scientific and medical communities has fundamentally altered the way in which we now address basic biomedical questions. Microarrays technology facilitates a more complete and inclusive experimental approach where alterations in the transcript level of entire genomes can be simultaneously assayed in response to a variety of stimuli. Conceptually different approaches to the development of microarray technology have resulted in the generation of two different array formats: oligonucleotide arrays and cDNA arrays. The application of microarray and related technologies to identify specific targets of defined genes that have clearly been implicated in cancer progression requires a specific experimental approach. The objective of this approach is to define changes in transcriptional profile that occur in response to modulating the expression level of the gene to be studied. The resulting altered expression profile can then be viewed as a blueprint by which that gene effects its cellular function. We have used oligonucleotide array-based expression profiling in collaboration with Affymetrix to identify downstream transcriptional targets of the BRCA1 tumor-suppressor gene as a means of defining its function. BRCA1 has been implicated in at least three functional pathways, namely, mediating the cellular response to DNA damage, as a cell cycle checkpoint protein and in the regulation of transcription. The physiological significance of these properties and their implications for the function of BRCA1 as a tumor suppressor gene remain to be defined. PMID- 11110603 TI - The molecular perspective: the ribosome. PMID- 11110604 TI - The cell cycle. PMID- 11110605 TI - Sharing new approaches to translational research in non-small cell lung cancer. PMID- 11110606 TI - Prophylactic cranial irradiation in small-cell lung cancer: is it still controversial or is it a no-brainer? PMID- 11110607 TI - Intravenous pantoprazole: a new tool for acutely ill patients who require acid suppression. AB - Until now, oral proton pump inhibitors have not been available as parenteral therapy in the acute care setting. Pantoprazole is the first parenteral proton pump inhibitor to become available in Canada. This agent is superior to the parenteral histamine 2 receptor antagonists with respect to acid suppressive effects and is not associated with tolerance development. Another advantage over the histamine 2 receptor antagonists is that pantoprazole does not require dosage adjustment in patients with renal impairment. Dosage adjustments are also not required for elderly patients or those with hepatic impairment when the drug is used at the usual dose for a limited period of time. Contrary to intravenous cimetidine and ranitidine, which have negative inotropic and chronotropic effects, intravenous pantoprazole is well tolerated and has no significant effect on heart rate, contractility or blood pressure. The lack of drug interactions for this agent also simplifies its use, especially in patients who may require multiple drugs during hospitalization. Parenteral pantoprazole is effective in the treatment of reflux esophagitis. It is also promising for the treatment of upper gastrointestinal bleeding and in the perioperative care of patients with Zollinger-Ellison syndrome, but further research in these areas is necessary. Once the patient is able to tolerate oral medications, parenteral therapy can be easily converted to oral therapy using an oral dose that was equivalent to the parenteral dose (ie, 40 mg given intravenously is equivalent to 40 mg given orally). PMID- 11110608 TI - Muscle cramps: a 'complication' of cirrhosis. AB - Muscle cramps are a common complaint in clinical practice. They are associated with various metabolic, endocrine, neurological and electrolyte abnormalities. A variety of hypotheses have been generated to explain the cause of muscle cramping, yet none has been able to support a consistent pathophysiological mechanism. Muscle cramps are painful, involuntary contractions of skeletal muscle. They occur frequently in individuals with cirrhosis, regardless of the etiology, and are thought to be a symptom of cirrhotic-stage liver disease. The pathophysiology of these cramps remains elusive; hence, a specific therapy has not been identified. Many therapeutic approaches have been offered, yet their efficacy, safety and mechanism of action remain poorly defined. This review defines muscle cramps and illuminates its prevalence in the cirrhotic individual. Current theories relating to the pathogenesis of muscle cramps are reviewed, and an overview of the various pharmacological agents that have had therapeutic success for this distressing and frustrating symptom is provided. PMID- 11110609 TI - Diagnosis and treatment of gastroesophageal reflux disease in infants and children. AB - Gastroesophageal reflux is a frequent, nonspecific phenomenon in infants and children. The recommended approach for infants with uncomplicated regurgitation is the reassurance of the parents about the physiological nature of excessive regurgitation, and if necessary, completed with dietary recommendations for formula-fed infants. If, despite these efforts, the symptoms persist, the administration of prokinetics such as cisapride is recommended before investigations such as esophageal pH monitoring are begun. Cisapride is the drug of choice because it has the best efficacy and safety profile. In infants and children presenting with symptoms that suggest esophagitis, endoscopy of the upper gastrointestinal tract is recommended. If there is severe esophagitis, acid suppression with H2 receptor antagonists or proton pomp inhibitors is recommended, eventually in combination with prokinetics. In life-threatening situations, or in patients who are resistant to or dependent on acid suppressive medication, a surgical procedure such as laparoscopic Nissen should be considered. Esophageal pH monitoring is recommended to document gastro- esophageal reflux disease in children presenting with unusual presentations such as chronic respiratory disease. Treatment consists of prokinetics and/or acid suppressive drugs, and surgery should be considered in many of these patients. PMID- 11110610 TI - Management of Barrett's esophagus. AB - There have been major recent advances in the understanding of the pathogenesis and epidemiology of Barrett's esophagus and adenocarcinoma of the esophagus. The advent of potent acid suppression with proton pump inhibitors and safe, minimally invasive antireflux procedures has made alleviating symptoms and eliminating peptic complications achievable goals for the vast majority of patients. Endoscopic surveillance of Barrett's esophagus is considered the standard of care and is widely used in clinical practice. Neither medical nor surgical antireflux procedures, however, result in the regression of Barrett's esophagus in any consistent manner. Thermal and chemical endoscopic ablation techniques show promise in both the management of high grade dysplasia and the reversal of Barrett's esophagus, but these techniques are still of unproven benefit, and can be costly and risky. Therefore, prospective and controlled studies with long term follow-up are needed before incorporating ablative techniques into routine clinical practice. Management of high grade dysplasia remains controversial. Alternative management strategies include surveillance, resection or ablation, tailored to the individual patient and the available expertise. Targets for future research include defining appropriate surveillance intervals; finding biological markers that identify patients at higher risk of progressing to cancer; defining the cancer risk and the appropriate management of patients with short segment Barrett's esophagus; understanding the natural history of dysplasia and comparing alternatives for the management of high grade dysplasia; and studying whether surgical management can delay or prevent the progression to dysplasia and adenocarcinoma. PMID- 11110611 TI - Primary gastric lymphoma of MALT: considerations of pathogenesis, diagnosis and therapy. AB - Primary gastric lymphoma of mucosa-associated lymphoid tissue (MALT) is a distinct entity with its own histological classification. Epidemiological, histomorphological, molecular biological and experimental data clearly underline that infection of the gastric mucosa by Helicobacter pylori plays an important role in both the development and progression of MALT lymphoma. Considering the histological grade of malignancy and dissemination (stage) of the disease as decisive prognostic factors, and therapeutic determinants, endoscopic bioptical diagnosis and endoscopic ultrasound are of particular importance. In cases of localized (stage 1), low grade lymphoma, eradication of H pylori offers a promising and fascinating therapeutic option. Surgical resection, radiotherapy or chemotherapy, and their combination, have proven to be effective treatment modalities. There is a need to clarify whether operative or conservative therapeutic strategies should be favoured in the future. PMID- 11110612 TI - Nutrition and gastric cancer. AB - The reasons for the worldwide decline in stomach cancer incidence and mortality rates are not fully understood, but dietary changes are clearly implicated. While the possible mechanisms of gastric carcinogenesis and the impact of Helicobacter pylori eradication remain open to debate, at least two practical recommendations to increase fruit and vegetable intake, and to reduce consumption of salt - are already supported by epidemiological evidence. These dietary recommendations may also be beneficial in the prevention of other cancers and chronic diseases. Promising evidence of a favourable effect of certain vitamins, such as vitamin C and E and beta-carotene, and minerals, such as selenium, justifies additional investigation. PMID- 11110613 TI - Pregnancy and the biliary tract. AB - Pregnancy induces many physiological changes, some of which may have pathological results. In population studies, gallstones were found in 6.5% to 8.4% of nulliparous women, and in 18.4% to 19.3% of women with two to three or more pregnancies. In women followed throughout pregnancy, neoformation of gallstones was documented in 3% to 8.1% depending on the population. Some 20% to 30% of these gallstones redissolve postpartum. The frequency of biliary colic during pregnancy is controversial, and the recommended therapeutic approach during pregnancy is conservative. When essential, invasive procedures are relatively well tolerated, preferably during the second trimester. Biliary sludge disappears postpartum in the great majority. Gallstones and sludge are most likely caused by biliary stasis, prolonged intestinal transit and increased cholesterol saturation of bile, which were all demonstrated to occur during pregnancy. PMID- 11110614 TI - Thrombocytopenia in liver disease. AB - Moderate thrombocytopenia is a frequent finding in cirrhosis of the liver and well tolerated in most instances. The pathophysiology of thrombocytopenia in liver disease has long been associated with the concept of hypersplenism, where portal hypertension was thought to cause pooling and sequestration of all corpuscular elements of the blood, predominantly thrombocytes in the enlarged spleen. The concept of hypersplenism was never proven beyond any doubt but was widely accepted for the lack of alternative explanations. With the discovery of the lineage-specific cytokine thrombopoietin (TPO) the missing link between hepatocellular function and thrombopoiesis was found. TPO is predominantly produced by the liver and constitutively expressed by hepatocytes. TPO production in humans is dependent on functional liver cell mass and is reduced when liver cell mass is severely damaged. This leads to reduced thrombopoiesis in the bone marrow and consequently to thrombocytopenia in the peripheral blood of patients with advanced-stage liver disease. With recombinant TPOs in development, patients with liver disease and TPO seem to be the ideal target population for this drug. Once the efficacy of thrombopoietin in patients with liver disease is proven, a potent yet safe drug may be available to treat cirrhotic patients undergoing invasive or surgical procedures, during bleeding episodes or when undergoing therapy with myelosuppressive drugs such as interferon-alpha. PMID- 11110615 TI - Neonatal cholestasis: a red alert for the jaundiced newborn. AB - Neonatal jaundice may indicate cholestasis rather than a benign, physiological condition. Any four-week-old newborn with persistent jaundice should have a fractionated bilirubin screen to determine whether the hyperbilirubinemia is unconjugated. Conjugated hyperbilirubinemia, a hallmark of neonatal cholestasis, is pathological and requires further investigation. These infants need prompt diagnosis, early intervention and careful follow-up to ensure continued growth and development. Recent progress in the physiology of bile flow is reviewed, and the evaluation and management of neonatal cholestasis are summarized. Further advances in delineating the cellular and molecular processes that regulate bile acid metabolism in both health and disease will lead to a greater understanding of the conditions causing neonatal cholestasis. Unravelling the etiopathogenesis of these neonatal cholestatic disorders will allow the development of novel diagnostic and therapeutic interventions that ultimately will effectuate the prognosis for these young patients. PMID- 11110616 TI - Clinical biology and potential use of thrombopoietin. AB - The discovery of platelet growth factors raised expectations that an effective method for abrogating thrombocytopenia would soon be available in the clinic. The cytokines initially described were pleiotropic in nature, and stimulation of platelet production was generally modest. However, one of these agents, interleukin-11, was successfully shown to reduce the incidence of severe thrombocytopenia in patients receiving dose-intensive chemotherapy, and has now received approval from the United States Food and Drug Administration for this purpose. Initial clinical trials of thrombopoietin, the central regulator of megakaryocytopoiesis and thrombopoiesis, and its analogues showed these agents to be the most potent stimulators of thrombopoiesis and to be associated with few adverse effects. They have also been shown to enhance platelet recovery after chemotherapy, but early results from trials investigating their ability to prevent severe thrombocytopenia associated with the treatment of leukemia and bone marrow transplantation have been disappointing. In addition, subcutaneous administration of one of these agents, megakaryocyte growth and development factor, has been shown to induce the formation of antibodies that neutralize native thrombopoietin and cause thrombocytopenia. Platelet growth factors remain promising therapeutic agents; however, there are a number of obstacles to overcome before they find general use in the clinic. PMID- 11110617 TI - Clinical experience with artificial liver support systems. AB - Fulminant hepatic failure is a devastating disease that, despite recent therapeutic advances, continues to be associated with high morbidity and mortality. Orthotopic liver transplantation has emerged as the sole modality of treatment that significantly improves survival. However, the critical shortage of donors precludes timely transplantation for all patients. Consequently, almost half of all patients with fulminant hepatic failure die before a graft becomes available. This has generated interest in developing a system that would support patients until either native liver regeneration occurs or an optimal donor liver can be found. Investigators have used biological, artificial and bioartificial techniques in an attempt to improve survival in liver failure. This article reviews the history, the current state of the art and future directions of artificial liver support. PMID- 11110618 TI - Nutrition of liver transplant patients. AB - Good cooperation between the hepatologist, surgeon and anesthesiologist is required to determine the appropriate perioperative nutritional management for the liver transplant patient. For preoperative risk stratification, nutritional assessment according to resting energy expenditure by indirect calorimetry, and body cell mass by bioelectrical impedence analysis, may be superior to anthropometric parameters. When considering impaired glucose tolerance in the early postoperative period, requirements of energy intake and macronutrients are no different from those established in major abdominal surgery. Preference should be made to use the enteral route whenever possible. Fat emulsions containing medium- and long-chain triglycerides have neither a negative impact on reticulo endothelial system recovery of the graft, nor any obvious metabolic advantages. There is no evidence for the routine use of branched-chain amino acids. Even in the case of good graft function, long term dietary evaluation and counselling may be useful. Impaired glucose tolerance, hyperlipidemia and hypercholesterolemia should be considered carefully. The role of preoperative nutritional therapy using oral supplements and the value of immune-enhancing substrates should be evaluated with special regard to a decrease in postoperative septic complications and for possible impact on immune tolerance after transplantation. PMID- 11110619 TI - Iron and liver diseases. AB - A mild to moderate iron excess is found in patients with liver diseases apparently unrelated to genetic hemochromatosis. Iron appears to affect the natural history of hepatitis C virus-related chronic liver diseases, alcoholic liver disease and nonalcoholic steatohepatitis by leading to a more severe fibrosis and thus aiding the evolution to cirrhosis. A higher frequency of mutations of the HFE gene, the gene responsible for hereditary hemochromatosis, is found in patients with liver diseases and increased liver iron than in normal patients. Patients with excess iron are potentially at a higher risk of developing hepatocellular carcinoma. Iron depletion therapy could interfere with fibrosis development and possibly reduce the risk of liver cancer occurrence. PMID- 11110620 TI - Medical management of chronic cholestatic liver diseases. AB - The purpose of the present review is to discuss the diagnosis and management of cholestatic liver diseases. Differential diagnoses to consider are described, including causes of extrahepatic biliary obstruction such as gallstones, strictures, extrabiliary malignancies and pancreatitis. In addition, diseases that cause intrahepatic cholestasis such as primary biliary cirrhosis, primary sclerosing cholangitis, hepatocellular diseases and a variety of miscellaneous causes including drugs that may cause cholestasis are discussed. Primary biliary cirrhosis and primary sclerosing cholangitis are reviewed in detail, and management options are identified. The prognosis of patients with these diseases is discussed, and the Mayo Mathematical Models in Cholestatic Liver Disease for both primary biliary cirrhosis and primary sclerosing cholangitis are provided. Finally, management options for the complications of cholestasis are provided. PMID- 11110621 TI - Hepatocellular bile salt transport: lessons from cholestasis. AB - Hepatic uptake and excretion of bile salts and several nonbile salt organic anions (eg, bilirubin) are mediated by a distinct set of polarized transport systems at the basolateral and apical plasma membrane domains of hepatocytes and bile duct epithelial cells (cholangiocytes). With the increasing availability of molecular probes for these transporters, evidence now exists that decreased or even absent expression of hepatobiliary transport proteins in hepatocytes or cholangiocytes may explain impaired transport function that results in hyperbilirubinemia and cholestasis. This review summarizes the molecular defects in hepatocellular membrane transporters that are associated with hereditary and acquired forms of cholestatic liver disease. PMID- 11110622 TI - Intensive care management of patients with acute liver failure with emphasis on systemic hemodynamic instability and cerebral edema: a critical appraisal of pathophysiology. AB - Acute liver failure (ALF) is a devastating disease leading to multiorgan dysfunction. The most dramatic impact of ALF is on the brain, as hepatic encephalopathy and intracranial hypertension (IH) develop. IH is associated with systemic hemodynamic instability, alterations in the regulation of cerebral blood flow and the development of cerebral edema. This review focuses on the pathophysiology of IH with special emphasis on cerebral blood flow and the development of cerebral edema. Based on these considerations, both traditional and new treatments for the management of IH in the future are discussed. PMID- 11110623 TI - Update on peripheral arterial vasodilation, ascites and hepatorenal syndrome in cirrhosis. AB - In cirrhosis of the liver, according to the peripheral arterial vasodilation hypothesis, relative underfilling of the arterial tree triggers a neurohumoral response (activation of renin-angiotensin-aldosterone system, sympathetic nervous system, nonosmotic release of vasopressin) aimed at restoring circulatory integrity by promoting renal sodium and water retention. Evidence has accumulated for a major role of increased vascular production of nitric oxide as the primary cause of arterial vasodilation in cirrhosis. Ascites is a common complication in cirrhosis. Treatment of ascites consists of a low salt diet with diuretics, and paracentesis together with plasma volume expanders in diuretic-resistant patients. Progression of cirrhosis may result in hepatorenal syndrome, a state of functional renal failure that carries an ominous prognosis. Orthotopic liver transplantation has remained the only curative treatment for patients with advanced liver disease; other modalities such as transjugular intrahepatic portosystemic shunt or vasopressin analogues may serve as a bridge to transplantation. Another complication of decompensated cirrhosis is spontaneous bacterial peritonitis, the incidence of which can be reduced by primary or secondary antibiotic prophylaxis by using orally active antibiotics. PMID- 11110624 TI - Are TIPS tops in the treatment of portal hypertension? A review on the use and misuse of transjugular intrahepatic portosystemic shunts. AB - Complications of portal hypertension are the Achilles heel of end-stage liver disease. Although initially developed in the 1960s, transjugular intrahepatic portosystemic shunts (TIPS) have recently gained popularity for decompressing the portal vein in patients with portal hypertension. The main indications for TIPS are the treatment of variceal hemorrhage unresponsive to endoscopic treatment and refractory ascites. Although several other applications for TIPS have been reported, they have not been tested in controlled trials. TIPS are not appropriate as initial therapy for variceal hemorrhage and ascites. Due to the virtually universal development of TIPS stenosis in the majority of patients, careful monitoring of stent patency is required. Several complications of TIPS are recognized, some of which are potentially fatal. Consequently, careful patient selection for TIPS is of paramount importance. Until further clinical trials become available, TIPS should be considered as a therapeutic option for the treatment of refractory variceal hemorrhage and refractory ascites in selected patients. PMID- 11110625 TI - Sex-related liver injury due to alcohol involves activation of Kupffer cells by endotoxin. AB - Females have a greater susceptibility to ethanol-induced liver injury than males. Females who drink ethanol regularly and have been overweight for 10 years or more are at greater risk for both hepatitis and cirrhosis than males, and females develop ethanol-induced liver injury more rapidly and with less ethanol than males. Female rats on an enteral ethanol protocol exhibit injury more quickly than males and have widespread fatty changes over a larger portion of the liver lobule. Moreover, levels of plasma endotoxin, intracellular adhesion molecule-1, free radical adducts, infiltrating neutrophils and nuclear factor kappa B are doubled in female rat livers compared with male rat livers after enteral ethanol treatment. Additionally, estrogen treatment in vivo increases the sensitivity of hepatic macrophages or Kupffer cells to endotoxin. Evidence has been presented that Kupffer cells are pivotal in the development of ethanol-induced liver injury. Destroying Kupffer cells with gadolinium chloride or decreasing bacterial endotoxin by sterilizing the gut with antibiotics inhibits early inflammation due to ethanol. Similar results have been obtained with anti-tumour necrosis factor alpha antibody. These data pointed to the hypothesis that ethanol-induced liver injury involves elevations in circulating endotoxin concentrations leading to activation of Kupffer cells, which causes a hypoxia-reoxygenation injury. This theory has been tested using pimonidazole, a 2-nitroimidazole marker, to quantify hypoxia in downstream, pericentral regions of the hepatic lobule. After chronic enteral ethanol treatment, pimonidazole binding doubles. Enteral ethanol also increases free radicals detected with electron spin resonance. Radical adducts, with coupling constants such as alpha-hydroxyethyl radical, have been shown to arise from ethanol. Importantly, hypoxia and radical production detected in bile are also decreased by the destruction of Kupffer cells with gadolinium chloride. These data support the hypothesis that Kupffer cells contribute to the vital sex differences in liver injury caused by ethanol. PMID- 11110626 TI - The role of surgery in the management of acute pancreatitis. AB - The clinical course of an episode of acute pancreatitis varies from a mild, transitory form to a severe necrotizing form characterized by multisystem organ failure and mortality in 20% to 40% of cases. Mild pancreatitis does not need specialized treatment, and surgery is necessary only to treat underlying mechanical factors such as gallstones or tumours at the papilla of Vater. On the other hand, patients with severe necrotizing pancreatitis need to be identified as early as possible after the onset of symptoms to start intensive care treatment. In this subgroup of patients, approximately 15% to 20% of all patients with acute pancreatitis, stratification according to infection status is crucial. Patients with infected necrosis must undergo surgical intervention, which consists of an organ-preserving necrosectomy followed by postoperative lavage and/or drainage to evacuate necrotic debris, which appears during the further course of the condition. Primary intensive care treatment, including antibiotic treatment, delays the need for surgery in most patients until the third or fourth week after the onset of symptoms. At that time, necrosectomy is technically easier to perform and the bleeding risk is reduced, compared with necrosectomy earlier in the disease course. In patients with sterile necrosis, the available data strongly support a conservative approach (ie, intensive care unit treatment). Surgery is rarely necessary in these patients. PMID- 11110627 TI - Epigastric fullness. AB - Epigastric fullness may be caused by a disordered gastric motor function, resulting in delayed gastric emptying, but may also be caused by rapid emptying, leading to a distention of the proximal small intestine. A rational diagnostic approach to a patient complaining of epigastric fullness is needed to reveal the underlying disorder or disease and to enable an adequate, targeted therapy. The clinical impression based on symptoms is unreliable and cannot distinguish function disorders and benign disease from severe conditions. PMID- 11110628 TI - Nutritional and pharmacological enhancement of gut-associated lymphoid tissue. AB - There has been an explosion of research in the field of nutrition over the past quarter century. Clinical studies have demonstrated the effectiveness of providing nutrition by the enteral route in reducing septic morbidity in critically ill patients. These improved outcomes have been substantiated by animal models that show that enteral nutrition decreases gut permeability while maintaining the gut-associated lymphoid tissue (GALT) in mucosal immunity. Evidence points to the important immunological role of the gut in the maintenance of mucosal immunity at both intestinal and extraintestinal sites. The preservation of this mucosal immunity by enteral nutrition is consistent with the lower morbidity seen in severely injured patients who receive nutrition via the gastrointestinal tract. For patients who are unable to be fed by the enteral route and who require parenteral nutrition, several supplements show promise in enhancing the mucosal immune system defenses. The nutritional and pharmacological tactics that may enhance the GALT and thereby maintain mucosal immunity are examined. PMID- 11110629 TI - The artificial anal sphincter. AB - The artificial anal sphincter as treatment for end stage anal incontinence was first described in 1987. Published series concern a total of 42 patients, with a success rate of approximately 80%. Infection has been the most serious complication, but a number of technical complications related to the device have also occurred and required revisional procedures in 40% to 60% of the patients. The artificial anal sphincter may be used for the same indications as dynamic graciloplasty except in patients with a previously irradiated or severely scarred perineum. PMID- 11110630 TI - Constipation: a physiological approach. AB - The first step in managing a patient with constipation is to understand the precise nature of the complaint. Is the onset recent? What are the frequency and form of the stools, and how much effort is required to defecate? Is constipation steady or alternating as in irritable bowel syndrome? Are there structural, metabolic or pharmacological confounders? Is the patient depressed? Has dietary fibre been tried at a sufficient dose? What are the patient's understanding and beliefs about the symptoms? Has there been sufficient and appropriate investigation? Armed with the answers to these questions, physicians can help most patients through lifestyle, dietary and pharmacological adjustments, along with supplementary fibre. Some patients may require regular doses of an osmotic laxative. Those few that fail these measures should have their transit time estimated while on a high fibre diet; if it is normal, further testing is unlikely to help. The above efforts should be re-emphasized, and reassurance should be offered. Some patients may require a psychological assessment. If transit time is prolonged and the patient may benefit from surgery for colonic inertia or biofeedback for anismus, then colon and anorectal function should be assessed. The decision to perform further tests should be made carefully, and unrealistic expectations should be discouraged. Before surgery is offered, the patient should have the benefit of receiving an expert opinion. Biofeedback helps some patients with isolated anorectal dysfunction. PMID- 11110631 TI - New imaging techniques for the evaluation of gastrointestinal diseases. AB - This article provides an overview of recently developed, noninvasive imaging modalities for the evaluation of gastrointestinal disease processes. The advent of spiral computed tomography, magnetic resonance cholangiopancreatography and conventional magnetic resonance imaging has facilitated the noninvasive assessment of pancreaticobiliary disease. Magnetic resonance cholangiopancreatography provides projectional images of the biliary tree and pancreatic duct, similar to those achieved by direct cholangiography, without the need to administer contrast medium. Spiral computed tomographic colonography provides virtual colonoscopic images of the colonic mucosa, allowing the detection of polyps without the risk associated with colonoscopy. PMID- 11110632 TI - [Chronic lymphoid leukemia complicated with ascites]. AB - Ascites are common in malignancies of the gastrointestinal tract but are rarely associated with chronic lymphoid leukemia. An unusual progression of chronic lymphoid leukemia, rapidly complicated with ascites, is presented, followed by a discussion of the causes and types of ascites found in solid and blood malignancies. PMID- 11110633 TI - Evaluation of a digital camera for acquiring radiographic images for telemedicine applications. AB - Many rural sites cannot afford a digitizer to digitize radiographic films and transmit them via a telemedicine network for review by a radiology specialist. This project tested the feasibility of using a consumer digital still camera to photograph radiographic images and transmit them via a telemedicine network to a consulting hub site. In this study, the feasibility of using a digital camera to photograph plain film radiographs of 40 bone trauma cases from a rural health center in Arizona was tested. The cases were transmitted to the Arizona Telemedicine Program hub site using a private asynchronous transfer mode network based on T1 carriers. Two orthopedic surgeons and two radiologists reviewed the cases on a color monitor and the original film images. The readers also rated image quality. There were no significant differences in diagnostic accuracy between conventional film and telemedicine reading. Diagnostic agreement between film and monitor viewing was quite high, as was agreement in confidence ratings. Image quality was generally rated as excellent to good in both viewing conditions. Cases that did not correlate well were judged to have poor image quality, or diagnoses were based on photographs that had part of the diagnostic region of interest cropped off. It was determined that a digital still camera can be used effectively in many cases to photograph radiographic images for transmission and viewing via a telemedicine network, as long as adequate views, zoomed in regions of interest, and good quality original films are used in the acquisition process. PMID- 11110634 TI - The physiologic cipher at altitude: telemedicine and real-time monitoring of climbers on Mount Everest. AB - Advanced wearable biosensors for vital-signs monitoring (physiologic cipher) are available to improve quality of healthcare in hospital, nursing home, and remote environments. The objective of this study was to determine reliability of vital signs monitoring systems in extreme environments. Three climbers were monitored 24 hours while climbing through Khumbu Icefall. Data were transmitted to Everest Base Camp (elevation 17,800 feet) and retransmitted to Yale University via telemedicine. Main outcome measures (location, heart rate, skin temperature, core body temperature, and activity level) all correlated through time-stamped identification. Two of three location devices functioned 100% of the time, and one device failed after initial acquisition of location 75% of the time. Vital signs monitors functioned from 95%-100% of the time, with the exception of one climber whose heart-rate monitor functioned 78% of the time. Due to architecture of automatic polling and data acquisition of biosensors, no climber was ever without a full set of data for more than 25 minutes. Climbers were monitored continuously in real-time from Mount Everest to Yale University for more than 45 minutes. Heart rate varied from 76 to 164 beats per minute, skin temperature varied from 5 to 10 degrees C, and core body temperature varied only 1-3 degrees C. No direct correlation was observed among heart rate, activity level, and body temperature, though numerous periods suggested intense and arduous activity. Field testing in the extreme environment of Mount Everest demonstrated an ability to track in real time both vital signs and position of climbers. However, these systems must be more reliable and robust. As technology transitions to commercial products, benefits of remote monitoring will become available for routine healthcare purposes. PMID- 11110635 TI - Telemedicine at the top of the world: the 1998 and 1999 Everest extreme expeditions. AB - The National Aeronautics and Space Administration (NASA) initially established a Commercial Space Center (CSC) in the Department of Surgery at Yale University School of Medicine to further develop and evaluate technologies in information systems, telecommunications applied to medicine, and physiologic sensors. The CSC is known as the Medical Informatics and Technology Applications Consortium (MITAC). The overall purpose for this NASA program is to leverage technology, innovation, and resources from industry and academia through collaborative partnerships. The Yale-NASA CSC/MITAC organized the Everest Extreme Expeditions (E3) for the spring Himalayan climbing seasons in the years 1998 and 1999. The primary mission was to deliver advanced medical support with global telemedicine capabilities to one of the world's most remote and hostile settings--Mount Everest. The purpose was both humanitarian (providing medical support) and scientific (conducting medical and technology research). The Yale team provided medical care for the Everest Base Camp community; conducted validation experiments for several types of advanced medical technologies in this remote, hostile environment; and performed real-time monitoring of selected climbers, while also assessing the basic science of altitude physiology. Additionally, the teams conducted outreach medical care to the citizens of Nepal and provided several educational forums for a variety of medical and nonmedical personnel- including school-age children. As part of the project's mission, the E3 medical teams at both Nepal and New Haven were on a 24-hour emergency call system to deliver medical care in the event of a crisis. Unlike most of the teams at Everest, the mission of E3 was not to climb the 29,028-foot mountain the Nepalese call Sagarmatha ("Sky Head"). The mountain served as an extreme testing ground for telemedicine. The lessons learned from this testbed are reviewed here and further clarify the abilities to provide better health care in remote and extreme environments--which for some may even be their home environment during/after a medical illness. PMID- 11110636 TI - Virtual reality in telemedicine. AB - Virtual reality (VR) can be considered as the leading edge of a general evolution of present communication interfaces involving the television, computer, and telephone. The main characteristic of this evolution is the full immersion of the human sensorimotor channels into a vivid and global communication experience. Because telemedicine principally focuses on transmitting medical information, VR has the potential to enhance this function. Particularly, VR can be used in telemedicine as an advanced communication interface, which enables a more intuitive mode of interacting with information, and as a flexible environment that enhances the feeling of physical presence during the interaction. In this article, the state of the art in VR-based telemedicine applications is described. This technology is now used in remote or augmented surgery as well as surgical training, which are critically dependent on eye-hand coordination. Recently, however, different researchers have tried to use virtual environments in medical visualization and for assessment and rehabilitation in neuropsychology. This article also discusses technological, ergonomical, and human factor issues, and specific guidelines are presented for expanding the use of VR in telemedicine. PMID- 11110637 TI - An industry, clinical, and academic telehealth partnership venture: progress, goals achieved, and lessons learned. AB - The goal of this project was to develop and test a multi-application telehealth workstation with an interface seamless to the end-users. This project required collaboration among the private sector, a clinical regional referral setting, a clinical tertiary receiving center, and a university academic unit. The project applied the usability testing methodology to design and test the workstation: (1) the developmental phase focused on planning, prototype workstation development, and end-user trials in the private sector research and development environment, and (2) the operational testing phase moved the workstation into the clinical setting. The latter included training of end-users and addressing policy and ethical issues. In addition, the partners documented the goals achieved and lessons learned. The project resulted in the refinement of the workstation for clinical applications. The unique, diverse, and at times complementary goals and lessons learned by each partner are noted. Collaboration around a shared goal was a key element in achieving the refined workstation. Collaborators reported that the lessons learned will aid them as they pursue telehealth activities. PMID- 11110638 TI - Telepathology networking in VISN-12 of the Veterans Health Administration. AB - The Veterans Integrated Service Network (VISN)-12, headquartered in Chicago, has implemented a telepathology network between the eight VISN-12 hospital laboratories and Loyola University Medical School linked by an economical, high speed wide-area network (WAN). Implementation of the WAN has reduced monthly telecommunications costs in VISN-12 by approximately 67%. In addition to telepathology, the WAN enables real-time teleradiology (general, computer tomography, and ultrasound), telefluoroscopy, telenuclear medicine imaging, telepsychiatry, and other forms of teleconsultation. Current applications of telepathology in VISN-12 include: primary diagnosis and consultation in surgical pathology, interpretation of serum protein electrophoresis and immunofixation gels, provision of support for consolidated microbiology laboratories, review of problematic peripheral blood smears, and distance learning. We have learned a variety of lessons from telepathology. The enthusiasm and technical skill of providers are essential for success. As well, frequent communication and rapid technical support are necessary. Finally, in a supportive environment, telepathology is a tool that can help bring together clinical laboratories with shared missions and goals. PMID- 11110639 TI - Implementing store-and-forward telemedicine: organizational issues. AB - This article documents a study of an organization's cultural readiness for successful implementation of a store-and-forward telemedicine system in a military health care environment. The study focused on the organization's cultural attributes that reflect its learning propensity and thereby its capability to adapt effectively and utilize the new technology. Results suggest that the organization did not possess the most favorable attributes for the utilization of a new technology, and the utilization of the new system was significantly lower than expected during the first 6 months of implementation. PMID- 11110640 TI - Long-distance education in radiology via a clinical telemedicine system. AB - The objectives of the present study are to examine quality of learning, improved comprehension, and the preferred method of learning with teleradiology. Thirteen pairs of medical students transmitted 2,133 radiological series of skeletal injuries telemedically from a community hospital to the main emergency department. Students marked their diagnosis, quality of learning, improvement in comprehension, and preferred method of learning, each marked on a Likert scale. The quality of learning of the 13 sessions marked by the pairs of the students was ranked as excellent = 6, good = 16, fair = 4, unsatisfactory = 0, poor = 0. This was in contrast with the radiological series, which was marked as excellent = 55% or poor or unsatisfactory = 27.5%, most of the poor marks, as explained by the students, being awarded to transmissions in the second half of what had been very long sessions. The pre-tuition comprehension improved from 73 series marked as poor, and 1,037 series marked as unsure. The preferred method of tuition was teleradiology only in 21.8%, teleradiology teaching enhanced by clinical features, lectures and personal study in 62%. Teleradiological sessions needed to be reduced to an hour. Teleradiological teaching is a good method as long as the session lasts no more than 60 minutes, is interrupted by a 10-minute coffee break, and is supplemented by clinical features, lectures, and personal study. PMID- 11110641 TI - Canadian experiences in telehealth: equalizing access to quality care. AB - The Canadian Conference "TExpo'98: Interactive Health" focused on four telehealth themes: community needs, Canadian experiences, industry perspectives, and access/security/interoperability issues. Health and socioeconomic needs have been the driving force behind telehealth initiatives; telelearning is one of the major Canadian initiatives. To encourage Canadian telehealth initiatives, the federal government is building a national health infrastructure. One element in this framework is concerned with empowering the public, strengthening health care services, and ensuring accountability. Technological advancements and innovative partnerships among health communities, government, users, professional bodies, and industry are critical to continued growth. Key issues including access, evaluation, implementation, privacy, confidentiality, security, and interoperability are of universal concern to participants. Research that examines the benefits and costs of telehealth is needed. PMID- 11110642 TI - Altered levels of hypothalamic-pituitary-adrenocortical axis hormones in baboons and mice during the course of infection with Schistosoma mansoni. AB - Baboons with primary or secondary exposure to Schistosoma mansoni were compared with each other over a 12-week infection period and with baseline values obtained from uninfected baboons with respect to serum levels of the hypothalamic pituitary-adrenal (HPA) axis hormones-corticotropin-releasing hormone, adrenocorticotropic hormone, dehydroepiandrosterone sulfate (DHEA-S), and cortisol. Baboons with primary infections, when worm recovery and oviposition rates were high and hepatic schistosome egg granulomas were large, had decreasing levels of these hormones as infection progressed, compared with both uninfected and reexposed baboons. The most reduced hormone level was that of DHEA-S. Reduction of DHEA-S and cortisol levels also occurred in primary murine infections. Reexposed baboons with low worm recovery and oviposition rates and small (modulated) hepatic granulomas showed the opposite pattern: HPA axis hormone levels were maintained at, or exceeded, the baseline values of uninfected baboons. These results suggest that HPA axis hormones may play a role in regulating the establishment, maturation, and oviposition of schistosomes and the progression of schistosomiasis. PMID- 11110643 TI - The free radical scavenger alpha-phenyl-tert-butyl nitrone aggravates hippocampal apoptosis and learning deficits in experimental pneumococcal meningitis. AB - The effect of adjuvant therapy with the radical scavenger alpha-phenyl-tert-butyl nitrone (PBN; 100 mg/kg given intraperitoneally every 8 h for 5 days) on brain injury and learning function was evaluated in an infant rat model of pneumococcal meningitis. Meningitis led to cortical necrotic injury (median, 3.97% [range, 0% 38.9%] of the cortex), which was reduced to a median of 0% (range, 0%-30.9%) of the cortex (P<.001) by PBN. However, neuronal apoptosis in the hippocampal dentate gyrus was increased by PBN, compared with that by saline (median score, 1.15 [range, 0.04-1.73] vs. 0.31 [range, 0-0.92]; P<.001). Learning function 3 weeks after cured infection, as assessed by the Morris water maze, was decreased, compared with that in uninfected control animals (P<.001). Parallel to the increase in hippocampal apoptosis, PBN further impaired learning in infected animals, compared with that in saline-treated animals (P<.02). These results contrast with those of an earlier study, in which PBN reduced cortical and hippocampal neuronal injury in group B streptococcal meningitis. Thus, in pneumococcal meningitis, antioxidant therapy with PBN aggravates hippocampal injury and learning deficits. PMID- 11110644 TI - Potency of a genetically detoxified mucosal adjuvant derived from the heat-labile enterotoxin of Escherichia coli (LTK63) is not adversely affected by the presence of preexisting immunity to the adjuvant. AB - The objective of the current studies was to evaluate whether the potency of a genetically detoxified mucosal adjuvant, derived from heat-labile enterotoxin of Escherichia coli (LTK63), was adversely affected by preexisting immunity. Studies of mice and pigs have involved consecutive intranasal immunization with LTK63 and 2 different vaccines (influenza virus hemagglutinin and a protein-polysaccharide conjugate of Neisseria meningitidis group C). The antibody responses to the vaccines plus LTK63 in naive animals were compared with the responses achieved in animals that previously had been immunized with the alternate vaccine plus LTK63. The data showed that the responses of both animal models to intranasal immunization were not adversely affected by the presence of preexisting immunity to the LTK63 adjuvant. PMID- 11110645 TI - Immunization against influenza: comparison of various topical and parenteral regimens containing inactivated and/or live attenuated vaccines in healthy adults. AB - Methods for enhancing immune responses to influenza were explored in 2 double blind, placebo-controlled trials. Intranasal (inl) immunization with monovalent, live attenuated, cold-adapted recombinant (CR) or inactivated influenza virus (MIV) vaccine and intramuscular (im) immunization with MIV were evaluated in various combinations. Healthy susceptible adults were assigned randomly to receive 10(7.1) TCID(50) of CR (A/H1N1 or A/H3N2), homologous MIV (15 microg), or placebo inl and placebo or homologous MIV im (6 groups in each study). Serum antibody responses were greatest in groups given im vaccine (with or without inl vaccine). A 2-fold increase in nasal wash antibody response frequencies was seen in groups given combined inl (CR or MIV) and im vaccine, compared with subjects given a single im (MIV) or inl (CR or MIV) vaccine. Combined inl and im immunization is a promising approach for enhancing both local and systemic immune responses against influenza. PMID- 11110646 TI - JC virus genotypes in France: molecular epidemiology and potential significance for progressive multifocal leukoencephalopathy. AB - JC virus (JCV) induces progressive multifocal leukoencephalopathy (PML), especially in human immunodeficiency virus (HIV)-infected patients. Although JCV genotypes have primarily been associated with geographic patterns, a distinctive neuropathogenicity was recently attributed to genotype 2. A multicenter study was conducted to describe the distribution of JCV genotypes in France and to investigate correlations between genotypes and PML. Genotypes were determined by sequencing 494 bp in the VP1 capsid gene. Peripheral JCV was studied in 65 urine samples from 43 HIV-infected patients and from 22 control subjects. Genotypes 1, 4, 2, and 3 were detected in 52.3%, 30.8%, 12.3%, and 4.6% of the samples, respectively. In 56 brain or cerebrospinal fluid samples, PML-associated JCV of genotypes 1, 2, 4, and 3 was found in 66%, 19.7%, 8.9%, and 5.4%, respectively. Infection with JCV genotypes 1 or 2 was correlated with PML (odds ratio, 3.29). On the other hand, infection with JCV genotype 4 could represent a lower risk for PML. PMID- 11110648 TI - Protection of Aotus monkeys by Plasmodium falciparum EBA-175 region II DNA prime protein boost immunization regimen. AB - Aotus monkeys received 4 doses of Plasmodium falciparum EBA-175 region II vaccine as plasmid DNA (Dv-Dv) or recombinant protein in adjuvant (Pv-Pv) or as 3 doses of DNA and 1 dose of protein (Dv-Pv). After 3 doses, antibody titers were approximately 10(4) in DNA-immunized monkeys and 10(6) in protein-immunized monkeys. A fourth dose did not significantly boost antibody responses in the Dv Dv only or Pv-Pv only groups, but titers were boosted to approximately 10(6) in monkeys in the Dv-Pv group. Four weeks after the last immunization, the animals were challenged with 10(4) P. falciparum-parasitized erythrocytes. Peak levels of parasitemia were lower in the 16 monkeys that received region II-containing plasmids or proteins than in the 16 controls (geometric mean: 194,178 and 410,110 parasites/microL, respectively; P=.013, Student's t test). Three of 4 monkeys in the Dv-Pv group did not require treatment. These data demonstrate that immunization with EBA-175 region II induces a significant antiparasite effect in vivo. PMID- 11110647 TI - Intervention with Shiga toxin (Stx) antibody after infection by Stx-producing Escherichia coli. AB - Shiga toxins (Stxs) produced by Escherichia coli (STEC) cause systemic vascular damage, manifested as hemolytic uremic syndrome in humans and as edema disease in pigs. Edema disease, a naturally occurring disease of pigs, was used to determine whether Stx antibodies, administered after infection and after the onset of Stx production, could prevent the systemic vascular damage and clinical disease caused by Stxs. A total of 119 STEC-infected pigs were treated with low, medium, or high doses of Stx antibody or with placebo. After inoculation with STEC, antibodies or placebo was injected intraperitoneally at 2 days postinoculation (DPI; low dose) or 4 DPI (medium and high doses). Edema disease was prevented with the low- and high-dose Stx antibody treatments administered at 2 and 4 DPI, respectively. High-dose antibody treatment also reduced the incidence and extent of vascular lesions. The degree of protection depended on the dose of antibody and the time of administration. PMID- 11110650 TI - The Evolution of Candida Species and Fluconazole Susceptibility among Oral and Vaginal Isolates Recovered from Human Immunodeficiency Virus (HIV)-Seropositive and At-Risk HIV-Seronegative Women. AB - Antifungal agents can effectively treat mucosal candidiasis; however, their use can lead to colonization with less susceptible species and to resistance among normally susceptible strains. Oral and vaginal Candida isolates obtained at 3 points over 2 years from human immunodeficiency virus (HIV)-seropositive and at risk HIV-seronegative women were identified by species and were evaluated for in vitro fluconazole susceptibility. Prevalence of non-C. albicans strains increased over time, and these strains were more likely among women reporting current antifungal use. Among C. albicans isolates, resistance was rare, with no evidence for progressive reduction in susceptibility over time. Among non-C. albicans isolates, reduced susceptibility occurred frequently and increased with time. HIV seropositive women were more likely to have non-C. albicans isolates with reduced susceptibility as were women reporting current antifungal use. This evolution and selection of mucosa-colonizing Candida species with reduced susceptibility could play a critical early role in the development of antifungal resistance among C. albicans isolates responsible for refractory candidiasis. PMID- 11110649 TI - Serum samples from infants vaccinated with a pneumococcal conjugate vaccine, PncT, protect mice against invasive infection caused by Streptococcus pneumoniae serotypes 6A and 6B. AB - Streptococcus pneumoniae serogroup 6 is an important cause of respiratory tract disease worldwide. Vaccination with 6B polysaccharide induces antibody response to the cross-reacting serotype 6A, but the protective capacity of 6A antibodies induced in infants remains unknown. In this study, passive immunization with serum samples obtained from infants vaccinated with an octavalent polysaccharide protein conjugate vaccine, PncT, protected mice against bacteremia and/or lung infection caused by intranasal challenge with serotypes 6B and 6A. Protective infant serum samples had significantly higher serotype-specific IgG levels and opsonic activity than did nonprotective serum samples. The protective level to either serotype was approximately 1 microg of specific IgG antibodies injected per mouse (corresponding to approximately 0.3 microg/mL). The protection was strongly related to opsonophagocytic antibody levels measured in vitro. These results demonstrate that PncT induces antibodies in infants that protect mice against invasive disease caused by the homologous serotype and by the cross reacting serotype 6A. PMID- 11110652 TI - Differentiation of human cytomegalovirus genotypes in immunocompromised patients on the basis of UL4 gene polymorphisms. AB - The variety of clinical manifestations of human cytomegalovirus infection probably results from both viral and host factors. Because genetic markers that span the viral genome are needed to identify such viral factors, polymorphisms at the UL4 gene locus were analyzed. DNA sequence analyses revealed 4 UL4-based genotypes, 2 of which were closely related but distinguishable by an uncommon polymorphism that results in overexpression of gpUL4. Similarities in the spectra of polymorphisms detected in various sets of samples reveal that all UL4 types infect a diversity of organs in different patient groups and in different geographic locales. Simultaneous infection by >1 UL4 type is common in AIDS patients. Data from sequencing analyses and from a rapid and simple UL4 typing assay did not detect linkage between UL4 and glycoprotein B types, which suggests that UL4 genotyping should be useful for studies that attempt to identify cytomegalovirus genes involved in disease pathogenesis. PMID- 11110651 TI - Immune mediators in cerebrospinal fluid during cryptococcosis are influenced by meningeal involvement and human immunodeficiency virus serostatus. AB - Pro- and anti-inflammatory mediators (tumor necrosis factor [TNF]-alpha, interleukin [IL]-6, IL-8, IL-10, and soluble TNF receptor II [sTNFR] II) were measured in cerebrospinal fluid (CSF) before treatment (day 0), and after 2 weeks and 3 months of antifungal therapy in 51 human immunodeficiency virus (HIV) positive and 7 HIV-negative patients with culture-confirmed cryptococcosis. On day 0, all mediator concentrations, except IL-10 in HIV-positive patients, were higher in patients with meningeal, rather than extrameningeal cryptococcosis or in control subjects (P<.05). For meningitis patients, all mediator levels, except sTNFR II, were higher in HIV-negative than HIV-positive patients (P<.05). Day 0 CSF IL-8 levels were higher in HIV-positive patients receiving antiretroviral therapy than in untreated persons (P<.02). Day 0 sTNFR II levels were higher in HIV-positive survivors at 3 months, and elevated levels were sustained in HIV positive patients with meningitis. Overall, these data support the idea that inflammatory responses are crucial to the eradication of cryptococcal infections in the central nervous system. PMID- 11110653 TI - Subduction and slab detachment in the Mediterranean-Carpathian region. AB - Seismic tomography models of the three-dimensional upper mantle velocity structure of the Mediterranean-Carpathian region provide a better understanding of the lithospheric processes governing its geodynamical evolution. Slab detachment, in particular lateral migration of this process along the plate boundary, is a key element in the lithospheric dynamics of the region during the last 20 to 30 million years. It strongly affects arc and trench migration, and causes along-strike variations in vertical motions, stress fields, and magmatism. In a terminal-stage subduction zone, involving collision and suturing, slab detachment is the natural last stage in the gravitational settling of subducted lithosphere. PMID- 11110654 TI - Sedimentary rocks of early Mars. AB - Layered and massive outcrops on Mars, some as thick as 4 kilometers, display the geomorphic attributes and stratigraphic relations of sedimentary rock. Repeated beds in some locations imply a dynamic depositional environment during early martian history. Subaerial (such as eolian, impact, and volcaniclastic) and subaqueous processes may have contributed to the formation of the layers. Affinity for impact craters suggests dominance of lacustrine deposition; alternatively, the materials were deposited in a dry, subaerial setting in which atmospheric density, and variations thereof mimic a subaqueous depositional environment. The source regions and transport paths for the materials are not preserved. PMID- 11110655 TI - Planar hexacoordinate carbon: a viable possibility. AB - The viability of molecules with planar hexacoordinate carbon atoms is demonstrated by density-functional theory (DFT) calculations for CB6(2-), a CB6H2 isomer, and three C3B4 minima. All of these species have six pi electrons and are aromatic. Although other C3B4 isomers are lower in energy, the activation barriers for the rearrangements of the three planar carbon C3B4 minima into more stable isomers are appreciable, and experimental observation should be possible. High-level ab initio calculations confirm the DFT results. The planar hexacoordination in these species does not violate the octet rule because six partial bonds to the central carbons are involved. PMID- 11110656 TI - Geodynamic evidence for a chemically depleted continental tectosphere. AB - The tectosphere, namely the portions of Earth's mantle lying below cratons, has a thermochemical structure that differs from average suboceanic mantle. The tectosphere is thought to be depleted in its basaltic components and to have an intrinsic buoyancy that balances the mass increase associated with its colder temperature relative to suboceanic mantle. Inversions of a large set of geodynamic data related to mantle convection, using tomography-based mantle flow models, indicate that the tectosphere is chemically depleted and relatively cold to 250 kilometers depth below Earth's surface. The approximate equilibrium between thermal and chemical buoyancy contributes to cratonic stability over geological time. PMID- 11110657 TI - Teleconvection: remotely driven thermal convection in rotating stratified spherical layers. AB - We report the discovery of a convective phenomenon found to occur in a rotating spherical system in which an inner convectively unstable fluid layer is bounded by a corotating outer convectively stable fluid layer. Although convection is thermally driven in the unstable interior, the resulting convective motions concentrate primarily in the stable outer region. This phenomenon, which we term teleconvection, suggests that fluid motions observed at the "surface" of a planet (such as Jupiter's alternating zonal winds) may be driven by an energy source located deep inside the planet. PMID- 11110658 TI - Rapid changes in the hydrologic cycle of the tropical Atlantic during the last glacial. AB - Sedimentary time series of color reflectance and major element chemistry from the anoxic Cariaco Basin off the coast of northern Venezuela record large and abrupt shifts in the hydrologic cycle of the tropical Atlantic during the past 90,000 years. Marine productivity maxima and increased precipitation and riverine discharge from northern South America are closely linked to interstadial (warm) climate events of marine isotope stage 3, as recorded in Greenland ice cores. Increased precipitation at this latitude during interstadials suggests the potential for greater moisture export from the Atlantic to Pacific, which could have affected the salinity balance of the Atlantic and increased thermohaline heat transport to high northern latitudes. This supports the notion that tropical feedbacks played an important role in modulating global climate during the last glacial period. PMID- 11110659 TI - Synchronous radiocarbon and climate shifts during the last deglaciation. AB - Radiocarbon data from the Cariaco Basin provide calibration of the carbon-14 time scale across the period of deglaciation (15,000 to 10, 000 years ago) with resolution available previously only from Holocene tree rings. Reconstructed changes in atmospheric carbon-14 are larger than previously thought, with the largest change occurring simultaneously with the sudden climatic cooling of the Younger Dryas event. Carbon-14 and published beryllium-10 data together suggest that concurrent climate and carbon-14 changes were predominantly the result of abrupt shifts in deep ocean ventilation. PMID- 11110660 TI - A primitive enantiornithine bird and the origin of feathers. AB - A fossil enantiornithine bird, Protopteryx fengningensis gen. et sp. nov., was collected from the Early Cretaceous Yixian Formation of Northern China. It provides fossil evidence of a triosseal canal in early birds. The manus and the alular digit are long, as in Archaeopteryx and Confuciusornis, but are relatively short in other enantiornithines. The alula or bastard wing is attached to an unreduced alular digit. The two central tail feathers are scalelike without branching. This type of feather may suggest that modern feathers evolved through the following stages: (i) elongated scale, (ii) central shaft, (iii) barbs, and finally (iv) barbules and barbicel. PMID- 11110661 TI - Glucose-dependent insulin release from genetically engineered K cells. AB - Genetic engineering of non-beta cells to release insulin upon feeding could be a therapeutic modality for patients with diabetes. A tumor-derived K-cell line was induced to produce human insulin by providing the cells with the human insulin gene linked to the 5'-regulatory region of the gene encoding glucose-dependent insulinotropic polypeptide (GIP). Mice expressing this transgene produced human insulin specifically in gut K cells. This insulin protected the mice from developing diabetes and maintained glucose tolerance after destruction of the native insulin-producing beta cells. PMID- 11110662 TI - Response to RAG-mediated VDJ cleavage by NBS1 and gamma-H2AX. AB - Genetic disorders affecting cellular responses to DNA damage are characterized by high rates of translocations involving antigen receptor loci and increased susceptibility to lymphoid malignancies. We report that the Nijmegen breakage syndrome protein (NBS1) and histone gamma-H2AX, which associate with irradiation induced DNA double-strand breaks (DSBs), are also found at sites of VDJ (variable, diversity, joining) recombination-induced DSBs. In developing thymocytes, NBS1 and gamma-H2AX form nuclear foci that colocalize with the T cell receptor alpha locus in response to recombination activating gene (RAG) protein mediated VDJ cleavage. Our results suggest that surveillance of T cell receptor recombination intermediates by NBS1 and gamma-H2AX may be important for preventing oncogenic translocations. PMID- 11110663 TI - Identification of synergistic signals initiating inner ear development. AB - Tissue manipulation experiments in amphibians more than 50 years ago showed that induction of the inner ear requires two signals: a mesodermal signal followed by a neural signal. However, the molecules mediating this process have remained elusive. We present evidence for mesodermal initiation of otic development in higher vertebrates and show that the mesoderm can direct terminal differentiation of the inner ear in rostral ectoderm. Furthermore, we demonstrate the synergistic interactions of the extracellular polypeptide ligands FGF-19 and Wnt-8c as mediators of mesodermal and neural signals, respectively, initiating inner ear development. PMID- 11110664 TI - The contribution of noise to contrast invariance of orientation tuning in cat visual cortex. AB - Feedforward models of visual cortex appear to be inconsistent with a well-known property of cortical cells: contrast invariance of orientation tuning. The models' fixed threshold broadens orientation tuning as contrast increases, whereas in real cells tuning width is invariant with contrast. We have compared the orientation tuning of spike and membrane potential responses in single cells. Both are contrast invariant, yet a threshold-linear relation applied to the membrane potential accurately predicts the orientation tuning of spike responses. The key to this apparent paradox lies in the noisiness of the membrane potential. Responses that are subthreshold on average are still capable of generating spikes on individual trials. Unlike the iceberg effect, contrast invariance remains intact even as threshold narrows orientation selectivity. Noise may, by extension, smooth the average relation between membrane potential and spike rate throughout the brain. PMID- 11110665 TI - Efficient initiation of HCV RNA replication in cell culture. AB - Hepatitis C virus (HCV) infection is a global health problem affecting an estimated 170 million individuals worldwide. We report the identification of multiple independent adaptive mutations that cluster in the HCV nonstructural protein NS5A and confer increased replicative ability in vitro. Among these adaptive mutations were a single amino acid substitution that allowed HCV RNA replication in 10% of transfected hepatoma cells and a deletion of 47 amino acids encompassing the interferon (IFN) sensitivity determining region (ISDR). Independent of the ISDR, IFN-alpha rapidly inhibited HCV RNA replication in vitro. This work establishes a robust, cell-based system for genetic and functional analyses of HCV replication. PMID- 11110666 TI - Multigenerational cortical inheritance of the Rax2 protein in orienting polarity and division in yeast. AB - Diploid yeast cells repeatedly polarize and bud from their poles, probably because of highly stable marks of unknown composition. Here, Rax2, a membrane protein, was shown to behave as such a mark. The Rax2 protein itself was inherited immutably at the cell cortex for multiple generations, and Rax2 was shown to have a half-life exceeding several generations. The persistent inheritance of cortical protein markers would provide a means to couple a cell's history to the future development of a precise morphogenetic form. PMID- 11110667 TI - Rescue of photoreceptor degeneration in rhodopsin-null Drosophila mutants by activated Rac1. AB - Rhodopsin is essential for photoreceptor morphogenesis; photoreceptors lacking rhodopsin degenerate in humans, mice, and Drosophila. Here we report that transgenic expression of a dominant-active Drosophila Rho guanosine triphosphatase, Drac1, rescued photoreceptor morphogenesis in rhodopsin-null mutants; expression of dominant-negative Drac1 resulted in a phenotype similar to that seen in rhodopsin-null mutants. Drac1 was localized in a specialization of the photoreceptor cortical actin cytoskeleton, which was lost in rhodopsin-null mutants. Thus, rhodopsin appears to organize the actin cytoskeleton through Drac1, contributing a structural support essential for photoreceptor morphogenesis. PMID- 11110668 TI - Identification of alloreactive T-cell epitopes on the Rhesus D protein. AB - Although considerable effort has been devoted to characterizing alloantibodies specific for the Rhesus D (RhD) blood group antigen, virtually nothing is known about the helper response that drives their production. Therefore, the aim of this study was to map alloreactive T-cell epitopes on the RhD protein. Peripheral blood mononuclear cells (PBMCs) were obtained from 22 RhD-negative volunteers in whom anti-D alloantibodies had developed after deliberate immunization or RhD incompatible pregnancy. The PBMCs were stimulated with a panel of up to 68 overlapping synthetic 15-mer peptides spanning the complete sequence of the RhD protein. One or more peptides elicited proliferative responses by PBMCs from all 22 of the alloimmune volunteers but from only 2 of 8 alloantibody-negative control donors. Proliferation of PBMCs from the alloimmune donors was mediated by major histocompatibility complex class II-restricted T cells expressing the CD45RO marker of previous activation or memory. The number of peptides that induced proliferative responses was unrelated to either the frequency of, or time since, exposure to RhD-positive red blood cells, but it correlated strongly (R(s) = 0.75; P <.003) with the level of anti-D antibodies in deliberately immunized donors. The patterns of stimulatory peptides varied among alloimmune volunteers, but particular sequences were commonly recognized, with 4 peptides each eliciting a response in more than 50% of these donors. Identification of such peptides containing dominant alloreactive helper epitopes is the first step in the development of improved or new approaches to preventing hemolytic disease of the newborn that are based on modulating the T-cell response to the RhD protein. PMID- 11110669 TI - Iron homeostasis: new tales from the crypt. AB - The enterocyte is a highly specialized cell of the duodenal epithelium that coordinates iron uptake and transport into the body. Until recently, the molecular mechanisms underlying iron absorption and iron homeostasis have remained a mystery. This review focuses on the proteins and regulatory mechanisms known to be present in the enterocyte precursor cell and in the mature enterocyte. The recent cloning of a basolateral iron transporter and investigations into its regulation provide new insights into possible mechanisms for iron transport and homeostasis. The roles of proteins such as iron regulatory proteins, the hereditary hemochromatosis protein (HFE)-transferrin receptor complex, and hephaestin in regulating this transporter and in regulating iron transport across the intestinal epithelium are discussed. A speculative, but testable, model for the maintenance of iron homeostasis, which incorporates the changes in the iron-related proteins associated with the life cycle of the enterocyte as it journeys from the crypt to the tip of the villous is proposed. PMID- 11110670 TI - The human basophil: a new appreciation of its role in immune responses. PMID- 11110671 TI - I-309 binds to and activates endothelial cell functions and acts as an angiogenic molecule in vivo. AB - Several chemokines have been shown to act as angiogenic molecules or to modulate the activity of growth factors such as fibroblast growth factor 2 (FGF-2) and vascular endothelial growth factor (VEGF). The detection of the CC chemokine receptor (CCR) 8 message in human umbilical vein endothelial cells (HUVECs) by reverse transcription- polymerase chain reaction (RT-PCR) and RNase protection assay (RPA), prompted us to investigate the potential role exerted by the CC chemokine I-309, a known ligand of such receptor, in both in vitro and in vivo angiogenesis assays. We show here that I-309 binds to endothelial cells, stimulates chemotaxis and invasion of these cells, and enhances HUVEC differentiation into capillary-like structures in an in vitro Matrigel assay. Furthermore, I-309 is an inducer of angiogenesis in vivo in both the rabbit cornea and the chick chorioallantoic membrane assay (CAM). PMID- 11110672 TI - Functional expression of CCR1, CCR3, CCR4, and CXCR4 chemokine receptors on human platelets. AB - Platelets are known to contain platelet factor 4 and beta-thromboglobulin, alpha chemokines containing the CXC motif, but recent studies extended the range to the beta-family characterized by the CC motif, including RANTES and Gro-alpha. There is also evidence for expression of chemokine receptors CCR4 and CXCR4 in platelets. This study shows that platelets have functional CCR1, CCR3, CCR4, and CXCR4 chemokine receptors. Polymerase chain reaction detected chemokine receptor messenger RNA in platelet RNA. CCR1, CCR3, and especially CCR4 gave strong signals; CXCR1 and CXCR4 were weakly positive. Flow cytometry with specific antibodies showed the presence of a clear signal for CXCR4 and weak signals for CCR1 and CCR3, whereas CXCR1, CXCR2, CXCR3, and CCR5 were all negative. Immunoprecipitation and Western blotting with polyclonal antibodies to cytoplasmic peptides clearly showed the presence of CCR1 and CCR4 in platelets in amounts comparable to monocytes and CCR4 transfected cells, respectively. Chemokines specific for these receptors, including monocyte chemotactic protein 1, macrophage inflammatory peptide 1alpha, eotaxin, RANTES, TARC, macrophage derived chemokine, and stromal cell-derived factor 1, activate platelets to give Ca(++) signals, aggregation, and release of granule contents. Platelet aggregation was dependent on release of adenosine diphosphate (ADP) and its interaction with platelet ADP receptors. Part, but not all, of the Ca(++) signal was due to ADP release feeding back to its receptors. Platelet activation also involved heparan or chondroitin sulfate associated with the platelet surface and was inhibited by cleavage of these glycosaminoglycans or by heparin or low molecular weight heparin. These platelet receptors may be involved in inflammatory or allergic responses or in platelet activation in human immunodeficiency virus infection. PMID- 11110673 TI - Characterization of Epstein-Barr virus-infected B cells in patients with posttransplantation lymphoproliferative disease: disappearance after rituximab therapy does not predict clinical response. AB - Post-transplantation lymphoproliferative disease (PTLD) is associated with Epstein-Barr virus (EBV). Quantitative and qualitative differences in EBV in peripheral blood mononuclear cells (PBMCs) of PTLD patients and healthy controls were characterized. A quantitative competitive polymerase chain reaction (QC-PCR) technique confirmed previous reports that EBV load in PBMCs is increased in patients with PTLD in comparison with healthy seropositive controls (18 539 vs 335 per 10(6) PBMCs, P =.0002). The average frequency of EBV-infected cells was also increased (271 vs 9 per 10(6) PBMCs, P =.008). The distribution in numbers of viral genome copies per cell was assessed by means of QC-PCR at dilutions of PBMCs. There was no difference between PTLD patients and healthy controls. Similarly, no differences in the patterns of viral gene expression were detected between patients and controls. Finally, the impact of therapy on viral load was analyzed. Patients with a past history of PTLD who were disease-free (after chemotherapy or withdrawal of immunosuppression) at the time of testing showed viral loads that overlapped with those of healthy seropositive controls. Patients treated with rituximab showed an almost immediate and dramatic decline in viral loads. This decline occurred even in patients whose PTLD progressed during therapy. These results suggest that the increased EBV load in PBMCs of PTLD patients can be accounted for by an increase in the number of infected B cells in the blood. However, in terms of viral copy number per cell and pattern of viral gene expression, these B cells are similar to those found in healthy controls. Disappearance of viral load with rituximab therapy confirms the localization of viral genomes in PBMCs to B cells. However, the lack of relationship between the change in viral load and clinical response highlights the difference between EBV infected PBMCs and neoplastic cells in PTLD. PMID- 11110674 TI - Association of complementation group and mutation type with clinical outcome in fanconi anemia. European Fanconi Anemia Research Group. AB - Fanconi anemia (FA) is a clinically and genetically heterogeneous disorder. Clinical care is complicated by variable age at onset and severity of hematologic symptoms. Recent advances in the molecular biology of FA have allowed us to investigate the relationship between FA genotype and the nature and severity of the clinical phenotype. Two hundred forty-five patients from all 7 known complementation groups (FA-A to FA-G) were studied. Mutations were detected in one of the cloned FANC genes in 169 patients; in the remainder the complementation group was assigned by cell fusion or Western blotting. A range of qualitative and quantitative clinical parameters was compared for each complementation group and for different classes of mutation. Significant phenotypic differences were found. FA-G patients had more severe cytopenia and a higher incidence of leukemia. Somatic abnormalities were less prevalent in FA-C, but more common in the rare groups FA-D, FA-E, and FA-F. In FA-A, patients homozygous for null mutations had an earlier onset of anemia and a higher incidence of leukemia than those with mutations producing an altered protein. In FA-C, there was a later age of onset of aplastic anemia and fewer somatic abnormalities in patients with the 322delG mutation, but there were more somatic abnormalities in patients with IVS4 + 4A --> T. This study indicates that FA patients with mutations in the FANCG gene and patients homozygous for null mutations in FANCA are high-risk groups with a poor hematologic outcome and should be considered as candidates both for frequent monitoring and early therapeutic intervention. (Blood. 2000;96:4064-4070) PMID- 11110675 TI - Molecular characterization of a case of atransferrinemia. AB - Hereditary atransferrinemia is a rare but instructive disorder that has previously been reported in only 8 patients in 6 families. It is characterized by microcytic anemia and by iron loading, and can be treated effectively by plasma infusions. We now report the first case known in the United States. We determined the sequences flanking the exons of the human transferrin gene and sequenced all of the exons and some of the flanking regions of the patient's DNA and that of her parents. The patient's DNA revealed a 10-base pair (bp) deletion, followed by a 9-bp insertion of a duplicated sequence. There was also a G-->C transversion at complementary DNA (cDNA) nt 1429, predicting that a proline was substituted for the alanine in amino acid position 477 (Ala 477 Pro). The latter mutation occurs at an evolutionarily highly conserved site; 704 control alleles were screened and this point mutation was not found. Each of the patient's transferrin genes contains one mutation, ie, the patient is a compound heterozygote for these mutations, because one was found in each of her parents. In addition to these mutations, which we regard to be causative in the patient's atransferrinemia, a silent polymorphism at cDNA 1572 G-->C was found in exon 13 as well as 2 previously unreported polymorphisms at IVS8 + 62 c-->t and IVS14-4 c-->a. The mutation in nt 1572 and that in intron 8 were common in the general population; the intron 14 mutation is rare. PMID- 11110676 TI - Karyotypic analysis predicts outcome of preremission and postremission therapy in adult acute myeloid leukemia: a Southwest Oncology Group/Eastern Cooperative Oncology Group Study. AB - The associations of cytogenetics with complete remission (CR) rates, overall survival (OS), and outcomes after CR were studied in 609 previously untreated AML patients younger than 56 years old in a clinical trial comparing 3 intensive postremission therapies: intensive chemotherapy, autologous transplantation (ABMT), or allogeneic bone marrow transplantation (alloBMT) from matched related donors. Patients were categorized into favorable, intermediate, unfavorable, and unknown cytogenetic risk groups based on pretreatment karyotypes. CR rates varied significantly (P <.0001) among the 4 groups: favorable, 84% (95% confidence interval [CI], 77%-90%); intermediate, 76% (CI, 71%-81%); unfavorable, 55% (CI, 48%-63%); and unknown, 54% (CI, 33%-74%). There was similar significant heterogeneity of OS (P <.0001), with the estimated relative risk of death from any cause being 1.50 (CI, 1.10-2.05), 3. 33 (CI, 2.43-4.55), and 2.66 (CI, 1.59 4.45) for the intermediate, unfavorable, and unknown risk groups, respectively, compared with the favorable group. In multivariate analyses, the effects of cytogenetic risk status on CR rate and OS could not be explained by other patient or disease characteristics. Among postremission patients, survival from CR varied significantly among favorable, intermediate, and unfavorable groups (P =.0003), with significant evidence of interaction (P =.017) between the effects of treatment and cytogenetic risk status on survival. Patients with favorable cytogenetics did significantly better following ABMT and alloBMT than with chemotherapy alone, whereas patients with unfavorable cytogenetics did better with alloBMT. Cytogenetic risk status is a significant factor in predicting response of AML patients to therapy; however, to tighten treatment correlates within genetically defined AML subsets, a significantly larger leukemia cytogenetic database is warranted. PMID- 11110677 TI - Evolving characteristics of AIDS-related lymphoma. AB - Over time, the epidemiologic and demographic characteristics of AIDS have changed in the United States, while the use of highly active antiretroviral therapy has changed the natural history of the disease. The goal of the study was to ascertain any changes in the epidemiologic, immunologic, pathologic, or clinical characteristics of AIDS-related lymphoma (ARL) over the course of the AIDS epidemic. Records of 369 patients with ARL diagnosed or treated at a single institution from 1982 through 1998 were reviewed. Single institutional data were compared to population-based data from the County of Los Angeles. Significant changes in the demographic profile of patients with newly diagnosed ARL have occurred, with the later time intervals associated with a higher prevalence in women (P =.25), in Latino/Hispanic individuals (P <.0001), and in those who acquired human immunodeficiency virus (HIV) heterosexually (P =.01). These changes were similar in both countywide, population-based analyses and in those from the single institution. The median CD4(+) lymphocyte count at lymphoma diagnosis has decreased significantly over the years, from 177/dL in the earliest time period (1982-1986), to 53/dL in the last time period from 1995 to 1998 (P =.0006). The pathologic spectrum of disease has also changed, with a decrease in the prevalence of small noncleaved lymphoma (P =.0005) and an increase in diffuse large cell lymphoma (P <.0001). Despite changes in the use of antiretroviral or chemotherapy regimens, the median survival has not significantly changed. PMID- 11110678 TI - Serum level of soluble urokinase-type plasminogen activator receptor is a strong and independent predictor of survival in human immunodeficiency virus infection. AB - Human immunodeficiency virus-1 (HIV-1) infection has been shown to result in up regulation of the urokinase-type plasminogen activator receptor (uPAR/CD87) on leukocytes in vitro and in vivo. The objective of this study was to investigate whether this up-regulation is paralleled by higher serum levels of soluble uPAR (suPAR) in patients with advanced HIV-1 disease and whether the serum level of suPAR is predictive of clinical outcome. Using an enzyme-linked immunosorbent assay, the level of suPAR was measured retrospectively in serum samples from 314 patients with HIV-1 infection. By Kaplan-Meier and Cox regression analyses, the serum suPAR levels were correlated to survival with AIDS-related death as the end point. High levels of serum suPAR (greater than median) were associated with poor overall survival, and Kaplan-Meier analysis on patients stratified by suPAR level demonstrated a continuous increase in mortality rates with higher suPAR levels. After adjustment for accepted prognostic markers-including Centers for Disease Control and Prevention-defined clinical stages, CD4 counts, viral load, beta2 microglobulin, and age-the prognostic strength of suPAR remained highly significant, indicating that the serum suPAR level is a novel, strong, and independent predictor of survival in HIV-1 infection. This report is the first to demonstrate an important association between the plasminogen activator system and disease progression in HIV-1 infection. PMID- 11110679 TI - Engraftment and survival after unrelated-donor bone marrow transplantation: a report from the national marrow donor program. AB - We analyzed engraftment of unrelated-donor (URD) bone marrow in 5246 patients who received transplants facilitated by the National Marrow Donor Program between August 1991 and June 1999. Among patients surviving at least 28 days, 4% had primary graft failure (failure to achieve an absolute neutrophil count > 5 x 10(8)/L before death or second stem-cell infusion). Multivariate logistic regression analysis showed that engraftment was associated with marrow matched at HLA-A, HLA-B, and DRB1; higher cell dose; younger recipient; male recipient; and recipient from a non-African American ethnic group. More rapid myeloid engraftment was associated with marrow serologically matched at HLA-A and HLA-B, DRB1 match, higher cell dose (in non-T-cell-depleted cases), younger recipient, recipient seronegativity for cytomegalovirus (CMV), male donor, no methotrexate for graft-versus-host disease prophylaxis, and transplantation done in more recent years. A platelet count higher than 50 x 10(9)/L was achieved by 47% of patients by day 100. Conditional on survival to day 100, survival at 3 years was 61% in those with platelet engraftment at day 30, 58% in those with engraftment between day 30 and day 100, and 33% in those without engraftment at day 100 (P <.0001). Factors favoring platelet engraftment were higher cell dose, DRB1 allele match, recipient seronegativity for CMV, HLA-A and HLA-B serologically matched donor, and male donor. Secondary graft failure occurred in 10% of patients achieving initial engraftment, and 18% of those patients are alive. These data demonstrate that quality of engraftment is an important predictor of survival after URD bone marrow transplantation. PMID- 11110680 TI - The human immunodeficiency virus type-1 central DNA flap is a crucial determinant for lentiviral vector nuclear import and gene transduction of human hematopoietic stem cells. AB - Gene transfer in human hematopoietic stem cells (HSCs) has great potential for both gene therapy and the understanding of hematopoiesis. As HSCs have extensive proliferative capacities, stable gene transfer should include genomic integration of the transgene. Lentiviral vectors are now preferred to oncoretroviral vectors especially because they integrate in nondividing cells such as HSCs, thereby avoiding the use of prolonged cytokine stimulation. Human immunodeficiency virus type-1 (HIV-1) has evolved a complex reverse transcription strategy including a central strand displacement event controlled in cis by the central polypurine tract (cPPT) and the central termination sequence (CTS). This creates, at the center of HIV-1 linear DNA molecules, a 99-nucleotide-long plus-strand overlap, the DNA flap, which acts as a cis-determinant of HIV-1 genome nuclear import. The reinsertion of the DNA flap sequence in an HIV-derived lentiviral vector promotes a striking increase of gene transduction efficiency in human CD34(+) hematopoietic cells, and the complementation of the nuclear import defect present in the parental vector accounts for this result. In a short ex vivo protocol, the flap-containing vector allows efficient transduction of the whole hierarchy of human HSCs including both slow-dividing or nondividing HSCs that have multiple lymphoid and myeloid potentials and primitive cells with long-term engraftment ability in nonobese diabetic/severe combined immunodeficiency mice (NOD/SCID). PMID- 11110681 TI - Stable in vivo expression of glucose-6-phosphate dehydrogenase (G6PD) and rescue of G6PD deficiency in stem cells by gene transfer. AB - Many mutations of the housekeeping gene encoding glucose-6-phosphate dehydrogenase (G6PD) cause G6PD deficiency in humans. Some underlie severe forms of chronic nonspherocytic hemolytic anemia (CNSHA) for which there is no definitive treatment. By using retroviral vectors pseudotyped with the vesicular stomatitis virus G glycoprotein that harbor the human G6PD (hG6PD) complementary DNA, stable and lifelong expression of hG6PD was obtained in all the hematopoietic tissues of 16 primary bone marrow transplant (BMT) recipient mice and 14 secondary BMT recipients. These findings demonstrate the integration of a functional gene in totipotent stem cells. The average total G6PD in peripheral blood cells of these transplanted mice, measured as enzyme activity, was twice that of untransplanted control mice. This allowed the inference that the amount of G6PD produced by the transduced gene must be therapeutically effective. With the same vectors both the cloning efficiency and the ability to form embryoid bodies were restored in embryonic stem cells, in which the G6PD gene had been inactivated by targeted homologous recombination, thus effectively rescuing their defective phenotype. Finally, expression of normal human G6PD in hG6PD-deficient primary hematopoietic cells and in human hematopoietic cells engrafted in nonobese diabetic/severe combined immunodeficient mice was obtained. This approach could cure severe CNSHA caused by G6PD deficiency. PMID- 11110682 TI - Evidence that ceramide mediates the ability of tumor necrosis factor to modulate primitive human hematopoietic cell fates. AB - In this study, it is shown that short-term exposure of normal human marrow CD34(+)CD38(-) cells to low concentrations of tumor necrosis factor (TNF) in the presence of 100 ng/mL Flt3 ligand and Steel factor and 20 ng/mL interleukin-3 (IL 3), IL-6, and granulocyte colony-stimulating factor, in either bulk or single cell serum-free cultures, markedly reduces their ability subsequently to generate colony-forming cells (CFCs) in 6-week stromal cell-containing long-term cultures without affecting their viability, mitogenic response, or short-term ability to produce CFCs. A similar differential effect on the functional attributes of CD34(+)CD38(-) cells was seen when C2- or C6-ceramide, but not dihydro-C2 ceramide (an inactive analog of ceramide), was substituted for TNF. The addition of D-erythro-MAPP (a specific inhibitor of intracellular ceramide degradation) enhanced the ability of TNF to selectively eliminate long-term culture-initiating cell (LTC-IC) activity. These findings indicate that TNF can directly modulate the ability of CD34(+)CD38(-) cells to maintain their LTC-IC function at doses below those required to initiate apoptosis, cell cycle arrest, or both, and they suggest that this may be mediated by the TNF-induced generation of intracellular ceramide. Identification of a signaling intermediate that can influence primitive hematopoietic cell fate decisions offers a new approach to the investigation of signaling mechanisms in normal stem cell populations and to how these may be altered in leukemic cells. PMID- 11110683 TI - Primitive hematopoietic stem cell function in vivo is uniquely high in the CXB-12 mouse strain. AB - Bone marrow cells (BMCs) from CXB-12/HiaJ (CXB-12) mice had 14 times the total long-term repopulating ability found in the best of 11 other CXB recombinant inbred (RI) lines. BMCs from each RI line donor were mixed with genetically marked standard competitor BMCs from the BALB/cByxC57BL/6 F1 (CByB6F1) hybrid, the mice used to produce the RI lines, and the mixtures repopulated lethally irradiated CByB6F1 recipients. Percentages of donor-type erythrocytes and lymphocytes measured the actual long-term repopulating functions of the donor RI lines relative to the standard competitor. CXB-12 BMCs repopulated better after 3 or 6 months than after 1 month, suggesting that the most primitive precursors were involved. Compared to CByB6F1 standard competitor cells, CXB-12 cells repopulated 3 to 12 times as well, with their advantage increasing when higher doses of cells were transplanted, probably because of hybrid resistance of the recipient against low doses. This was far better than expected, because F1 cells normally function 2 to 3 times as well as cells from an inbred strain. In competitive dilution, the advantage resulted from 2 factors: more precursor cells and more function per precursor. In the model that best fit the data, CXB-12 donors had 2.4 times the concentration of hematopoietic stem cells (HSCs) as the CByB6F1 standard, and each HSC repopulated 1.4 times as well. CXB-12 mice did not have elevated erythrocyte and lymphocyte numbers in blood and marrow and did not have unusually elevated concentrations of colony-forming unit spleen, cobblestone colonies, and long-term colony-initiating cells in marrow. PMID- 11110684 TI - WNT signaling modulates the diversification of hematopoietic cells. AB - WNT proteins compose a family of secreted signaling molecules that regulate cell fate and behavior. The possible influence of WNTs on hematopoietic cell fate was examined. Both hematopoietic progenitor cell (HPC)-enriched embryonic avian bone marrow cells and the quail mesodermal stem cell line QCE6 were used for these studies. Under optimized conditions, the bone marrow and QCE6 cells behaved identically and developed into red blood cells (RBCs), monocytes, macrophages, granulocytes, and thrombocytes. This broad range of blood cell phenotypes exhibited by QCE6 cells was dependent on their active expression of WNT11. However, when QCE6 cells were prevented from producing WNT11-by expression of a stably transfected WNT11 antisense transgene-the cultures were dominated by highly vacuolated macrophages. RBCs were absent from these cultures, and the presence of monocytes was greatly diminished. Exposure of these WNT11 antisense cells to soluble WNT11 or WNT5a restored the broad range of blood cell phenotypes exhibited by parental QCE6 cells. Overexpression of WNT protein in QCE6 cells further increased the prevalence of RBCs and monocytes and greatly diminished the appearance of macrophages. Accordingly, treatment of HPC-enriched bone marrow cultures with soluble WNT11 or WNT5a inhibited macrophage formation. Instead, monocytes and RBCs were the prevalent cells displayed by WNT-treated bone marrow cultures. Together, these data indicate that WNTs may play a major role in regulating hematopoietic cell fate. PMID- 11110685 TI - Stromal-derived factor 1 and thrombopoietin regulate distinct aspects of human megakaryopoiesis. AB - The role of the chemokine binding stromal-derived factor 1 (SDF-1) in normal human megakaryopoiesis at the cellular and molecular levels and its comparison with that of thrombopoietin (TPO) have not been determined. In this study it was found that SDF-1, unlike TPO, does not stimulate alpha(IIb)beta(3)(+) cell proliferation or differentiation or have an antiapoptotic effect. However, it does induce chemotaxis, trans-Matrigel migration, and secretion of matrix metalloproteinase 9 (MMP-9) and vascular endothelial growth factor (VEGF) by these cells, and both SDF-1 and TPO increase the adhesion of alpha(IIb)beta(3)(+) cells to fibrinogen and vitronectin. Investigating the intracellular signaling pathways induced by SDF-1 and TPO revealed some overlapping patterns of protein phosphorylation/activation (mitogen-activated protein kinase [MAPK] p42/44, MAPK p38, and AKT [protein kinase B]) and some that were distinct for TPO (eg, JAK STAT) and for SDF-1 (eg, NF-kappa B). It was also found that though inhibition of phosphatidyl-inositol 3-kinase (PI-3K) by LY294002 in alpha(IIb)beta(3)(+) cells induced apoptosis and inhibited chemotaxis adhesion and the secretion of MMP-9 and VEGF, the inhibition of MAPK p42/44 (by the MEK inhibitor U0126) had no effect on the survival, proliferation, and migration of these cells. Hence, it is suggested that the proliferative effect of TPO is more related to activation of the JAK-STAT pathway (unique to TPO), and the PI-3K-AKT axis is differentially involved in TPO- and SDF-1-dependent signaling. Accordingly, PI-3K is involved in TPO-mediated inhibition of apoptosis, TPO- and SDF-1-regulated adhesion to fibrinogen and vitronectin, and SDF-1-mediated migration. This study expands the understanding of the role of SDF-1 and TPO in normal human megakaryopoiesis and indicates the molecular basis of the observed differences in cellular responses. (Blood. 2000;96:4142-4151) PMID- 11110686 TI - Stromal-derived factor 1-induced megakaryocyte migration and platelet production is dependent on matrix metalloproteinases. AB - Despite the discovery of thrombopoietin (TPO) and its contribution to megakaryocytopoiesis, the exact mechanisms and sites of platelet production are unknown. It has been shown that mature megakaryocytes (MKs) functionally express the stromal-derived factor 1 (SDF-1) receptor, CXCR4. SDF-1-induced migration of mature MKs through endothelial cell layers results in increased platelet production. Because the migration of polyploid MKs from the bone marrow microenvironment requires remodeling of the perivascular extracellular matrix, it was hypothesized that mature polyploid MKs may express matrix metalloproteinases (MMPs), facilitating their exit into the bone marrow extravascular space. In this report, it is demonstrated that SDF-1 induces the expression and release of gelatinase B (MMP-9) by purified mature polyploid human MKs and an adeno-CXCR4 infected megakaryocytic cell line. Neutralizing antibody to MMP-9, but not MMP-2, blocked SDF-1-induced migration of MKs through reconstituted basement membrane, suggesting that expression of MMP-9 is critical for MK migration. Incubation of mature MKs with a synthetic MMP inhibitor, 5-phenyl-1,10-phenanthrolene, resulted in the inhibition of platelet formation, suggesting that the expression of MMPs is not only critical for megakaryocyte migration but also for subsequent platelet release. Confirming these results, adeno-SDF-1 injection into normal mice resulted in increased platelet counts, a process that could be blocked by a synthetic MMP inhibitor. These results suggest mobilization of MKs involves sequential expression and activation of chemokine receptors such as CXCR4, MMP-9, followed by transendothelial migration. MMP inhibitors may have potential use in the treatment of thrombotic and myeloproliferative disorders. (Blood. 2000;96:4152-4159) PMID- 11110687 TI - During ontogeny primitive (CD34(+)CD38(-)) hematopoietic cells show altered expression of a subset of genes associated with early cytokine and differentiation responses of their adult counterparts. AB - Comparison of gene expression profiles in closely related subpopulations of primitive hematopoietic cells offers a powerful first step to elucidating the molecular basis of their different biologic properties. Here we present the results of a comparative quantitative analysis of transcript levels for various growth factor receptors, ligands, and transcription factor genes in CD34(+)CD38( ) and CD34(+)CD38(+) cells purified from first trimester human fetal liver, cord blood, and adult bone marrow (BM). In addition, adult BM CD34(+)CD38(-) cells were examined after short-term exposure to various growth factors in vitro. Transcripts for 19 of the 24 genes analyzed were detected in unmanipulated adult BM CD34(+)CD38(-) cells. Moreover, the levels of transforming growth factor beta (TGF-beta), gp130, c-fos, and c-jun transcripts in these cells were consistently and significantly different (higher) than in all other populations analyzed, including phenotypically similar but biologically different cells from fetal or neonatal sources, as well as adult BM CD34(+) cells still in G(0) after 2 days of growth factor stimulation. We have thus identified a subset of early response genes whose expression in primitive human hematopoietic cells is differently regulated during ontogeny and in a fashion that is recapitulated in growth factor stimulated adult BM CD34(+)CD38(-) cells, before their cell cycle progression and independent of their subsequent differentiation response. These findings suggest a progressive alteration in the physiology of primitive hematopoietic cells during development such that these cells initially display a partially "activated" state, which is not maximally repressed until after birth. (Blood. 2000;96:4160-4168) PMID- 11110688 TI - The alpha(IIb)beta(3) integrin and GPIb-V-IX complex identify distinct stages in the maturation of CD34(+) cord blood cells to megakaryocytes. AB - Megakaryocytopoiesis is a complex multistep process involving cell division, endoreplication, and maturation and resulting in the release of platelets into the blood circulation. Megakaryocytes (MK) progressively express lineage restricted proteins, some of which play essential roles in platelet physiology. Glycoprotein (GP)Ib-V-IX (CD42) and GPIIb (CD41) are examples of MK-specific proteins having receptor properties essential for platelet adhesion and aggregation. This study defined the progressive expression of the GPIb-V-IX complex during in vitro MK maturation and compared it to that of GPIIb, an early MK marker. Human cord blood CD34(+) progenitor cells were cultured in the presence of cytokines inducing megakaryocytic differentiation. GPIb-V-IX expression appeared at day 3 of culture and was strictly dependent on MK cytokine induction, whereas GPIIb was already present in immature CD34(+) cells. Analysis by flow cytometry and of the messenger RNA level both showed that GPV appeared 1 day later than GPIb-IX. Microscopy studies confirmed the late appearance of GPV, which was principally localized in the cytoplasm when GPIb-IX was found on the cell surface, suggesting a delayed program of GPV synthesis and trafficking. Cell sorting studies revealed that the CD41(+)GPV(+) population contained 4N and 8N cells at day 7, and was less effective than CD41(+)GPV(-) cells in generating burst-forming units of erythrocytes or MK colonies. This study shows that the subunits of the GPIb-V-IX complex represent unique surface markers of MK maturation. The genes coding for GPIb-IX and GPV are useful tools to study megakaryocytopoiesis and for tissue-specific or conditional expression in mature MK and platelets. (Blood. 2000;96:4169-4177) PMID- 11110689 TI - Zebrafish homolog of the leukemia gene CBFB: its expression during embryogenesis and its relationship to scl and gata-1 in hematopoiesis. AB - Mammalian CBFB encodes a transcription factor (CBF beta) that in combination with CBF alpha 2 binds to specific DNA sequences and regulates expression of a number of hematopoietic genes. CBFB is associated with human leukemias through a chromosome 16 inversion and is essential for definitive hematopoiesis during mouse embryo development. We have isolated a zebrafish cbfb complementary DNA (cDNA) clone from a zebrafish kidney cDNA library. This cbfb is highly homologous to human and mouse CBFB/Cbfb genes at both the DNA and protein level. In biochemical analyses, cbfbeta binds to human CBF alpha 2 and enhances its DNA binding. During zebrafish development, cbfb is expressed in the lateral plate mesoderm at tail bud stage and in the intermediate cell mass (ICM, the location of embryonic hematopoiesis) between the 21- to 26-somite stages. The cbfb is also expressed in Rohon-Beard cells, cranial nerve ganglia, hindbrain, retina, branchial arches, jaw, and fin buds. Expression of cbfb is decreased or absent in the ICM and Rohon-Beard cells in some hematopoietic mutants and is unaffected in others. We have also analyzed the expression of scl and gata-1 in the same hematopoietic mutants to ascertain the relative order of these transcription factors to cbfb in zebrafish hematopoiesis. Our results indicate that cbfb is expressed in early hematopoietic progenitors and that its expression pattern in the hematopoietic mutants is similar to that of scl. (Blood. 2000;96:4178-4184) PMID- 11110690 TI - Human hematopoietic stem cells stimulated to proliferate in vitro lose engraftment potential during their S/G(2)/M transit and do not reenter G(0). AB - An understanding of mechanisms regulating hematopoietic stem cell engraftment is of pivotal importance to the clinical use of cultured and genetically modified transplants. Human cord blood (CB) cells with lymphomyeloid repopulating activity in NOD/SCID mice were recently shown to undergo multiple self-renewal divisions within 6 days in serum-free cultures containing Flt3-ligand, Steel factor, interleukin 3 (IL-3), IL-6, and granulocyte colony-stimulating factor. The present study shows that, on the fifth day, the transplantable stem cell activity is restricted to the G(1) fraction, even though both colony-forming cells (CFCs) and long-term culture-initiating cells (LTC-ICs) in the same cultures are approximately equally distributed between G(0)/G(1) and S/G(2)/M. Interestingly, the G(0) cells defined by their low levels of Hoechst 33342 and Pyronin Y staining, and reduced Ki67 and cyclin D expression (representing 21% of the cultured CB population) include some mature erythroid CFCs but very few primitive CFCs, LTC-ICs, or repopulating cells. Although these findings suggest a cell cycle-associated change in in vivo stem cell homing, the cultured G(0)/G(1) and S/G(2)/M CD34(+) CB cells exhibited no differences in levels of expression of VLA 4, VLA-5, or CXCR-4. Moreover, further incubation of these cells for 1 day in the presence of a concentration of transforming growth factor beta(1) that increased the G(0)/G(1) fraction did not enhance detection of repopulating cells. The demonstration of a cell cycle-associated mechanism that selectively silences the transplantability of proliferating human hematopoietic stem cells poses both challenges and opportunities for the future improvement of ex vivo-manipulated grafts. (Blood. 2000;96:4185-4193) PMID- 11110691 TI - Characterization and functional analysis of laminin isoforms in human bone marrow. AB - Laminins are a family of disulfide-linked heterotrimeric proteins consisting of 3 different subunits termed alpha, beta, and gamma chains. Combinations of 11 characterized laminin subunits (alpha 1-alpha 5, beta 1-beta 3, and gamma 1-gamma 3) generate at least 12 laminin isoforms, which can serve different functions. Although expression of laminin in the hematopoietic microenvironment has been known for many years, the nature of the laminin isoforms present in the human bone marrow is poorly characterized. The present study attempts to clarify this issue. Reverse transcriptase-polymerase chain reaction analysis of human bone marrow stromal cells suggested the expression of many laminin isoforms in the marrow. Northern blot and immunoblot analysis, however, showed that laminin-8/9 and laminin-10/11 are the most abundant laminin isoforms synthesized by human bone marrow stromal cells. Other isoforms, if present, certainly play a minor role in the hematopoietic microenvironment. Functionally, laminin-10/11 preparations showed strong adhesive interactions with human CD34(+) cell lines. Antibodies against the beta 1 integrin subunit inhibited these interactions. Other laminin isoforms, especially laminin-1 and laminin-2/4, showed only weak or no adhesive interactions with the hematopoietic cell lines tested, explaining former negative results. In addition to its adhesion-mediating properties, laminin-10/11 preparations also showed a mitogenic activity for human hematopoietic progenitor cells. Taken together, these data suggest that laminin in the bone marrow plays a hitherto unexpected important function in the development of hematopoietic progenitor cells. (Blood. 2000;96:4194-4203) PMID- 11110692 TI - Interferon-gamma-induced apoptotic responses of Fanconi anemia group C hematopoietic progenitor cells involve caspase 8-dependent activation of caspase 3 family members. AB - Hematopoietic progenitor cells (HPC) from mice nullizygous at the Fanconi anemia (FA) group C locus and children with Fanconi anemia group C (FA-C) are hypersensitive to interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha. This hypersensitivity results, in part, from the capacity of these cytokines to prime the fas pathway. Because fas-mediated programmed cell death in many cells involves sequential activation of specific caspases, we tested the hypothesis that programmed cell death in FA HPC involves the ordered activation of specific caspase molecules. Lysates from lymphoblasts treated with both agonistic anti-fas antibody and IFN-gamma contained activated caspase 3 family members (caspases 3, 6, and 7), as well as caspase 8, whereas activation of caspases 1, 2, 4, 9, and 10 was not detected. The apoptotic effects of fas agonists in IFN-gamma-treated human and murine FA-C cells were blocked when pretreated with inhibitors (ac-DEVD cho, CP-DEVD-cho, Z-DEVD-FMK) of the caspase 3 protease. Inhibitors (ac-YVAD-cho, CP-YVAD-cho, Z-YVAD-FMK) of caspase 1 did not block apoptosis or caspase 3 activation. Treatment of FA cells with the fluoromethyl ketone tetrapeptide caspase 8 inhibitor (ac-IETD-FMK) did suppress caspase 3 activation. A 4-fold greater fraction of IFN-induced FA-C cells expressed caspase 3 than FA-C cells complemented by retroviral-mediated transfer of FANCC. Therefore fas-induced apoptosis in Fanconi anemia cells of the C type involves the activation of caspase 8, which controls activation of caspase 3 family members and one direct or indirect function of the FANCC protein is to suppress apoptotic responses to IFN-gamma upstream of caspase 3 activation. (Blood. 2000;96:4204-4211) PMID- 11110693 TI - Plasminogen activator inhibitor-1 deficiency protects against atherosclerosis progression in the mouse carotid artery. AB - Dissolution of the fibrin blood clot is regulated in large part by plasminogen activator inhibitor-1 (PAI-1). Elevated levels of plasma PAI-1 may be an important risk factor for atherosclerotic vascular disease and are associated with premature myocardial infarction. The role of the endogenous plasminogen activation system in limiting thrombus formation following atherosclerotic plaque disruption is unknown. This study found that genetic deficiency for PAI-1, the primary physiologic regulator of tissue-type plasminogen activator (tPA), prolonged the time to occlusive thrombosis following photochemical injury to carotid atherosclerotic plaque in apolipoprotein E-deficient (apoE(-/-)) mice. However, anatomic analysis revealed a striking difference in the extent of atherosclerosis at the carotid artery bifurcation between apoE(-/-) mice and mice doubly deficient for apoE and PAI-1 (PAI-1(-/-)/apoE(-/-)). Consistent with a previous report, PAI-1(+/+)/apoE(-/-)and PAI-1(-/-)/apoE(-/-) mice developed similar atherosclerosis in the aortic arch. The marked protection from atherosclerosis progression at the carotid bifurcation conferred by PAI-1 deficiency suggests a critical role for PAI-1 in the pathogenesis of atherosclerosis at sites of turbulent flow, potentially through the inhibition of fibrin clearance. Consistent with this hypothesis, intense fibrinogen/fibrin staining was observed in atherosclerotic lesions at the carotid bifurcation compared to the aortic arch. These observations identify significant differences in the pathogenesis of atherosclerosis at varying sites in the vascular tree and suggest a previously unappreciated role for the plasminogen activation system in atherosclerosis progression at sites of turbulent flow. (Blood. 2000;96:4212 4215) PMID- 11110694 TI - Vascular endothelial growth factor-stimulated endothelial cells promote adhesion and activation of platelets. AB - Coagulation abnormalities, including an increased platelet turnover, are frequently found in patients with cancer. Because platelets secrete angiogenic factors on activation, this study tested the hypothesis that platelets contribute to angiogenesis. Stimulation with vascular endothelial growth factor (VEGF, 25 ng/mL) of human umbilical vein endothelial cells (HUVECs) promoted adhesion of nonactivated platelets 2.5-fold. In contrast, stimulation of HUVECs with basic fibroblast growth factor (bFGF) did not promote platelet adhesion. By blocking tissue factor (TF) activity, platelet adhesion was prevented and antibodies against fibrin(ogen) and the platelet-specific integrin, alpha(IIb)beta(3), inhibited platelet adhesion for 70% to 90%. These results indicate that VEGF induced platelet adhesion to endothelial cells is dependent on activation of TF. The involvement of fibrin(ogen) and the alpha(IIb)beta(3) integrin, which exposes a high-affinity binding site for fibrin(ogen) on platelet activation, indicates that these adhering platelets are activated. This was supported by the finding that the activity of thrombin, a product of TF-activated coagulation and a potent platelet activator, was required for platelet adhesion. Finally, platelets at physiologic concentrations stimulated proliferation of HUVECs, indicative of proangiogenic activity in vivo. These results support the hypothesis that platelets contribute to tumor-induced angiogenesis. In addition, they may explain the clinical observation of an increased platelet turnover in cancer patients. Platelets may also play an important role in other angiogenesis-dependent diseases in which VEGF is involved, such as diabetes and autoimmune diseases. (Blood. 2000;96:4216-4221) PMID- 11110695 TI - Spontaneous thrombosis in mice carrying the factor V Leiden mutation. AB - A polymorphism in coagulation factor V, factor V Leiden (FVL), is the major known genetic risk factor for thrombosis in humans. Approximately 10% of mutation carriers experience clinically significant thrombosis in their lifetime. In a small subset of patients, thrombosis is associated with coinheritance of other prothrombotic gene mutations. However, the potential contribution of additional genetic risk factors in the majority of patients remains unknown. To gain insight into the molecular basis for the variable expressivity of FVL, mice were generated carrying the homologous mutation (R504Q [single-letter amino acid codes]) inserted into the endogenous murine Fv gene. Adult heterozygous (FvQ/+) and homozygous (FvQ/Q) mice are viable and fertile and exhibit normal survival. Compared with wild-type mice, adult FvQ/Q mice demonstrate a marked increase in spontaneous tissue fibrin deposition. No differences in fetal development or survival are observed among FvQ/Q, FvQ/+ or control littermates on the C57BL/6J genetic background. In contrast, on a mixed 129Sv-C57BL/6J genetic background, FvQ/Q mice develop disseminated intravascular thrombosis in the perinatal period, resulting in significant mortality shortly after birth. These results may explain the high degree of conservation of the R504/R506 activated protein C cleavage site within FV among mammalian species and suggest an important contribution of other genetic factors to the thrombosis associated with FVL in humans. (Blood. 2000;96:4222-4226) PMID- 11110697 TI - Human CLP36, a PDZ-domain and LIM-domain protein, binds to alpha-actinin-1 and associates with actin filaments and stress fibers in activated platelets and endothelial cells. AB - A 38-kd protein that associates with F-actin structures in activated platelets and endothelial cells was purified, cloned, and characterized. The protein contains an N-terminal PDZ motif, a large intervening sequence, and a C-terminal LIM domain and was identified as the human homolog of rat CLP36. The study showed that CLP36 associates with actin filaments and stress fibers that are formed during shape change and spreading of platelets and during migration and contraction of endothelial cells. CLP36 binds to alpha-actinin-1 as shown by coimmunoprecipitation, pull-down experiments, yeast 2-hybrid analysis, and blot overlay assays and colocalizes with alpha-actinin-1 along endothelial actin stress fibers. In contrast to alpha-actinin-1, CLP36 was absent from focal adhesions in both activated platelets and endothelial cells. The N-terminal part of CLP36 containing the PDZ domain and the intervening region, but not the LIM domain, targeted enhanced green fluorescent protein fusion proteins to stress fibers in endothelial cells. Yeast 2-hybrid analysis demonstrated that the intervening sequence, but not the PDZ or the LIM domain of CLP36, binds to the spectrinlike repeats 2 and 3 of alpha-actinin-1. The study further shows that CLP36 binds to alpha-actinin in resting platelets and translocates as a CLP36/alpha-actinin complex to the newly formed actin cytoskeleton in activated platelets. The results indicate that CLP36 binds via alpha-actinin-1 to actin filaments and stress fibers in activated human platelets and endothelial cells. The study suggests that CLP36 may direct alpha-actinin-1 to specific actin structures and at this position might modulate the function of alpha-actinin-1. (Blood. 2000;96:4236-4245) PMID- 11110696 TI - Defects in the cappuccino (cno) gene on mouse chromosome 5 and human 4p cause Hermansky-Pudlak syndrome by an AP-3-independent mechanism. AB - Defects in a triad of organelles (melanosomes, platelet granules, and lysosomes) result in albinism, prolonged bleeding, and lysosome abnormalities in Hermansky Pudlak syndrome (HPS). Defects in HPS1, a protein of unknown function, and in components of the AP-3 complex cause some, but not all, cases of HPS in humans. There have been 15 inherited models of HPS described in the mouse, underscoring its marked genetic heterogeneity. Here we characterize a new spontaneous mutation in the mouse, cappuccino (cno), that maps to mouse chromosome 5 in a region conserved with human 4p15-p16. Melanosomes of cno/cno mice are immature and dramatically decreased in number in the eye and skin, resulting in severe oculocutaneous albinism. Platelet dense body contents (adenosine triphosphate, serotonin) are markedly deficient, leading to defective aggregation and prolonged bleeding. Lysosomal enzyme concentrations are significantly elevated in the kidney and liver. Genetic, immunofluorescence microscopy, and lysosomal protein trafficking studies indicate that the AP-3 complex is intact in cno/cno mice. It was concluded that the cappuccino gene encodes a product involved in an AP-3 independent mechanism critical to the biogenesis of lysosome-related organelles. (Blood. 2000;96:4227-4235) PMID- 11110698 TI - Fyn and Lyn phosphorylate the Fc receptor gamma chain downstream of glycoprotein VI in murine platelets, and Lyn regulates a novel feedback pathway. AB - Activation of platelets by collagen is mediated by the complex glycoprotein VI (GPVI)/Fc receptor gamma (FcR gamma chain). In the current study, the role of 2 Src family kinases, Fyn and Lyn, in GPVI signaling has been examined using murine platelets deficient in one or both kinases. In the fyn(-/-) platelets, tyrosine phosphorylation of FcR gamma chain, phopholipase C (PLC) activity, aggregation, and secretion are reduced, though the time of onset of response is unchanged. In the lyn(-/-) platelets, there is a delay of up to 30 seconds in the onset of tyrosine phosphorylation and functional responses, followed by recovery of phosphorylation and potentiation of aggregation and alpha-granule secretion. Tyrosine phosphorylation and aggregation in response to stimulation by collagen related peptide is further attenuated and delayed in fyn(-/-)lyn(-/-) double mutant platelets, and potentiation is not seen. This study provides the first genetic evidence that Fyn and Lyn mediate FcR immune receptor tyrosine-based activation motif phosphorylation and PLC gamma 2 activation after the ligation of GPVI. Lyn plays an additional role in inhibiting platelet activation through an uncharacterized inhibitory pathway. (Blood. 2000;96:4246-4253) PMID- 11110699 TI - Thrombosis and shock induced by activating antiplatelet antibodies in human Fc gamma RIIA transgenic mice: the interplay among antibody, spleen, and Fc receptor. AB - Transgenic mouse lines were created that express Fc gamma RIIA on platelets and macrophages at human physiologic levels, and they were used to explore the consequences in vivo of activating antiplatelet antibodies. Anti-CD9 antibody activated platelets of Fc gamma RIIA transgenic (tg) mice and, following injection in vivo, caused more rapid severe thrombocytopenia than nonactivating antiplatelet antibody. Anti-CD9 injected into Fc gamma RIIA tg crossed with FcR gamma-chain knockout (gamma-KO) mice caused thrombosis and shock in all mice, and death in 16 of 18 mice. The shock depended on platelet Fc receptor density and antibody dose. On histologic examination, the lung vasculature of anti-CD9 treated Fc gamma RIIA tg x gamma-KO mice contained extensive platelet-fibrin thrombi. Thrombosis and shock in Fc gamma RIIA tg mice in the context of the FcR gamma-chain knockout suggested the importance of the interplay of intravascular platelet activation and splenic clearance. Reduction of splenic clearance surgically (splenectomy) or functionally (monoclonal antibody treatment) also facilitated anti-CD9-mediated shock in Fc gamma RIIA tg mice. The spleen, which clears nonactivating antibody-coated platelets leading to thrombocytopenia, appears to play a protective role in the thrombosis and shock observed with activating antiplatelet antibody. The data indicate that antibodies, which activate platelets in an Fc gamma RIIA-dependent manner, can lead to thrombosis, shock, and death. Furthermore, antibody titer, platelet Fc receptor density, and splenic clearance are likely important determinants of the outcome. (Blood. 2000;96:4254-4260) PMID- 11110701 TI - Antiadhesive function of 130-kd glycoform of CD43 expressed in CD4 T-lymphocyte clones and transfectant cell lines. AB - Conflicting findings regarding proadhesion and antiadhesion in cell-to-cell interactions were previously reported for CD43. We examined possible differences in the role of the 130-kd glycoform and the 115-kd glycoform of CD43 in cellular adhesion in vitro. We generated a monoclonal antibody (MFT3) that discriminates between helper and nonhelper murine T-cell clones. Characterization of MFT3 with use of biochemical analysis and complementary DNA (cDNA) transfection experiments showed that it is specific for the 130-kd glycoform of CD43. T-cell clones that expressed the 130-kd CD43 glycoform showed decreased homocytic aggregation and decreased adhesion to spleen cells, B-lymphoma cell lines, and fibroblastic cell lines compared with T-cell clones negative for the 130-kd glycoform. Expression of core 2 beta-1, 6-N-acetylglucosaminyltransferase (C2GnT) cDNA together with CD43 cDNA resulted in expression of both the 130-kd CD43 glycoform and the 115-kd CD43 glycoform in fibroblastic cell lines. Using these cell lines, we showed that the 130-kd glycoform but not the 115-kd glycoform of CD43 has an antiadhesive function in cellular interactions. Our findings suggest that the antiadhesive function of CD43 is primarily carried out by the 130-kd glycoform. (Blood. 2000;96:4267-4275) PMID- 11110700 TI - Polymorphonuclear leukocyte activation and hemostasis in patients with essential thrombocythemia and polycythemia vera. AB - Thrombohemorrhagic complications are a major cause of morbidity and mortality in patients with essential thrombocythemia (ET) and polycythemia vera (PV). The pathogenesis of these complications is not completely clarified. Several studies have described abnormalities of red blood cells and platelets in these patients. However, no studies are available on changes in the polymorphonuclear leukocytes (PMNs), which can play an important role in the activation of the hemostatic system. In patients with ET (n = 37) and PV (n = 34), a series of PMN activation parameters (PMN membrane CD11b and leukocyte alkaline phosphatase [LAP] antigen expression, cellular elastase content, plasma elastase, and myeloperoxidase levels) was evaluated simultaneously with the levels of plasma markers of endothelial damage (thrombomodulin and von Willebrand factor antigen) and hypercoagulation (thrombin-antithrombin complex, prothrombin fragment 1 + 2, and D-dimer). The results show the occurrence of PMN activation in both groups of patients compared with a control group of healthy subjects. An increase in CD11b and LAP expression by PMN membrane was observed, together with a significant increase in cellular elastase content, plasma elastase, and myeloperoxidase levels. In addition, patients had high plasma levels of endothelial and hypercoagulation markers compared with controls. For the first time, these data show that in ET and PV, 2 hematologic conditions that place patients at increased risk for thrombosis, an in vivo leukocyte activation occurs and is associated with laboratory signs of endothelium and coagulation system activation. (Blood. 2000;96:4261-4266) PMID- 11110702 TI - Ligation of E-cadherin on in vitro-generated immature Langerhans-type dendritic cells inhibits their maturation. AB - Epithelial tissues of various organs contain immature Langerhans cell (LC)-type dendritic cells, which play key roles in immunity. LCs reside for long time periods at an immature stage in epithelia before migrating to T-cell-rich areas of regional lymph nodes to become mature interdigitating dendritic cells (DCs). LCs express the epithelial adhesion molecule E-cadherin and undergo homophilic E cadherin adhesion with surrounding epithelial cells. Using a defined serum-free differentiation model of human CD34(+) hematopoietic progenitor cells, it was demonstrated that LCs generated in vitro in the presence of transforming growth factor beta1 (TGF-beta1) express high levels of E-cadherin and form large homotypic cell clusters. Homotypic LC clustering can be inhibited by the addition of anti-E- cadherin monoclonal antibodies (mAbs). Loss of E-cadherin adhesion of LCs by mechanical cluster disaggregation correlates with the rapid up-regulation of CD86, neo-expression of CD83, and diminished CD1a cell surface expression by LCs-specific phenotypic features of mature DCs. Antibody ligation of E-cadherin on the surfaces of immature LCs after mechanical cluster disruption strongly reduces the percentages of mature DCs. The addition of mAbs to the adhesion molecules LFA-1 or CD31 to parallel cultures similarly inhibits homotypic LC cluster formation, but, in contrast to anti-E-cadherin, these mAbs fail to inhibit DC maturation. Thus, E-cadherin engagement on immature LCs specifically inhibits the acquisition of mature DC features. E-cadherin-mediated LC maturation suppression may represent a constitutive active epithelial mechanism that prevents the uncontrolled maturation of immature LCs. (Blood. 2000;96:4276-4284) PMID- 11110703 TI - Interferon alpha prevents spontaneous apoptosis of clonal Th2 cells associated with chronic hypereosinophilia. AB - A recent study identified a clonal expansion of CD3(-)CD4(+)cells secreting Th2 type cytokines in 4 patients with chronic hypereosinophilia. Because interferon alpha (IFN-alpha) is used in the therapy of the idiopathic hypereosinophilic syndrome, the effects of this cytokine on the survival of clonal Th2 cells isolated from the blood of 2 patients were determined. First, these cells displayed a high rate of spontaneous apoptosis on culture in cytokine-free medium and were also sensitive to Fas-mediated apoptosis induced by soluble Fas ligand. Addition of IFN-alpha or interleukin-2 (IL-2) to culture medium resulted in significant protection against spontaneous but not Fas-induced apoptosis. Although spontaneous apoptosis of the clonal Th2 cells was clearly associated with down-regulation of both bcl-2 and bcl-x(L) levels, IFN-alpha had no significant effect on the expression of these antiapoptotic proteins, whereas addition of IL-2 resulted in higher levels of bcl-2. On the other hand, IFN-alpha decreased the numbers of cells with disrupted mitochondrial transmembrane potential both during spontaneous apoptosis and after exposure to protoporphyrin IX. Thus, IFN-alpha might promote the survival of clonal Th2 cells, an effect that could be relevant to the therapeutic approach for patients with chronic hypereosinophilia caused by clonal expansion of Th2-type cells. (Blood. 2000;96:4285-4292) PMID- 11110705 TI - The long pentraxin PTX3 binds to apoptotic cells and regulates their clearance by antigen-presenting dendritic cells. AB - Pentraxins are acute-phase proteins produced in vivo during inflammatory reactions. Classical short pentraxins, C-reactive protein, and serum amyloid P component are generated in the liver in response to interleukin (IL)-6. The long pentraxin PTX3 is produced in tissues under the control of primary proinflammatory signals, such as lipopolysaccharide, IL-1 beta, and tumor necrosis factor-alpha, which also promote maturation of dendritic cells (DCs). Cell death commonly occurs during inflammatory reactions. In this study, it is shown that PTX3 specifically binds to dying cells. The binding was dose dependent and saturable. Recognition was restricted to extranuclear membrane domains and to a chronological window after UV irradiation or after CD95 cross-linking-induced or spontaneous cell death in vitro. PTX3 bound to necrotic cells to a lesser extent. Human DCs failed to internalize dying cells in the presence of PTX3, while they took up normally soluble or inert particulate substrates. These results suggest that PTX3 sequesters cell remnants from antigen-presenting cells, possibly contributing to preventing the onset of autoimmune reactions in inflamed tissues. (Blood. 2000;96:4300-4306) PMID- 11110704 TI - Immunologic dynamics in hemophiliac patients infected with hepatitis C virus and human immunodeficiency virus: influence of antiretroviral therapy. AB - Infection with hepatitis C virus (HCV) or human immunodeficiency virus (HIV) or both is common in hemophiliac patients due to putative transmission through clotting factor concentrates. Recently, highly active antiretroviral therapy (HAART) has been found to markedly improve viremia and immunologic parameters in patients infected with HIV. This report considers interactions between these viral infections, the immune system, and antiretroviral therapy. A total of 130 male hemophiliac patients were grouped according to type of viremia (HCV, HIV, both, or neither). Along with 30 healthy men age-matched to viremic patients, these groups were compared with respect to viral load and immunologic parameters. Thirty-five patients treated as above for HIV were serially followed up. HCV infection was associated with reduced peripheral B-cell and CD4(+)-cell counts and with increased serum IgG and IgM levels, whereas HIV infection was associated with reduced peripheral CD4(+)-cell counts and increased serum IgG and IgA levels. In patients with both viruses, HCV and HIV RNA load correlated inversely with peripheral B-cell and CD4(+)-cell counts, respectively. HAART reduced levels of both viruses in the blood. Of the 25 patients with both viruses, HAART eliminated HCV in 2. In conclusion, immunologic dynamics differed between hemophiliac patients infected with HCV, HIV, or both. The relative dynamics of HCV viral load, peripheral B-cell count, and serum IgM level were similar to those of HIV viral load, CD4(+)-cell count, and serum IgA. (Blood. 2000;96:4293 4299) PMID- 11110706 TI - Differential effects of CD30 activation in anaplastic large cell lymphoma and Hodgkin disease cells. AB - CD30 is a member of the tumor necrosis factor (TNF) receptor superfamily that is expressed on activated lymphocytes, as well as on neoplastic cells of Hodgkin disease (HD) and anaplastic large cell lymphoma (ALCL). A number of reports have shown that, depending on cellular context, CD30 signaling can exert a variety of effects, ranging from cell death to cellular proliferation. In the present study this disparity was examined, using a number of ALCL- and HD-derived cell lines. Activation of CD30 led to the induction of apoptotic death of ALCL cells, along with the selective reduction of TNF receptor-associated factor 2 and impairment in the ability of these cells to activate the pro-survival transcription factor nuclear factor kappa B (NF-kappa B). In contrast, HD cells, which constitutively express NF-kappa B, were not susceptible to CD30-induced apoptosis but could be sensitized following ectopic overexpression of a superdominant I kappa B. These studies suggest that NF-kappa B plays a determining role in the sensitivity or resistance of lymphoma cells to CD30-induced apoptosis, which may have important consequences in the clinical treatment of CD30-positive neoplasia. (Blood. 2000;96:4307-4312) PMID- 11110707 TI - Interferon alpha down-regulates telomerase reverse transcriptase and telomerase activity in human malignant and nonmalignant hematopoietic cells. AB - Recently, the derepressed expression of the catalytic subunit of telomerase, human telomerase reverse transcriptase (hTERT), the enzyme that elongates telomeres, has been implicated as an important step in the immortalization process. The exact regulation of hTERT expression, which is the rate-limiting factor for telomerase activity, is at present unclear. As transformed cells seem to be dependent on a constitutive telomerase activity, the availability of inhibitors would potentially be of great value in antineoplastic therapy. Interferons (IFNs) have been successfully used in the treatment of several forms of malignancies, but the underlying molecular mechanisms responsible for the antitumor activity are poorly defined. In this study we have investigated the effects of IFNs on hTERT expression and telomerase activity. We found that IFN alpha rapidly (commonly within 4 hours) and significantly down-regulates the expression of hTERT and telomerase activity in a number of human malignant hematopoietic cell lines, primary leukemic cells from patients with acute leukemia as well as T-lymphocytes from healthy donors. This effect of IFN-alpha did not seem to depend on IFN-alpha-mediated cell growth arrest or alterations in c-myc expression. The finding that IFN induces a repression of hTERT and a decrease in telomerase activity suggests a novel mechanism that may play a significant role in the antitumor action of IFN. (Blood. 2000;96:4313-4318) PMID- 11110709 TI - BAL is a novel risk-related gene in diffuse large B-cell lymphomas that enhances cellular migration. AB - Clinical risk factor models such as the International Prognostic Index are used to identify diffuse large B-cell lymphoma (DLB-CL) patients with different risks of death from their diseases. To elucidate the molecular bases for these observed clinical differences in outcome, differential display was used to identify a novel gene, termed BAL (B-aggressive lymphoma), which is expressed at significantly higher levels in fatal high-risk DLB-CLs than in cured low-risk tumors. The major BAL complementary DNA encodes a previously uncharacterized 88 kd nuclear protein with a duplicated N-terminal domain homologous to the nonhistone portion of histone-macroH2A and a C-terminal alpha-helical region with 2 short coiled-coil domains. Of note, the BAL N-terminus and secondary structure resemble those of a recently identified human protein, KIAA1268. In addition, both BAL and KIAA1268 map to chromosome 3q21, further suggesting that these genes belong to a newly identified family. BAL is expressed at increased levels in DLB CL cell lines with an activated peripheral B cell, rather than a germinal center B cell, phenotype. This observation and the characteristic dissemination of high risk DLB-CLs prompted studies regarding the role of BAL in B-cell migration. In classical transwell assays, stable BAL-overexpressing B-cell lymphoma transfectants had significantly higher rates of migration than vector-only transfectants, indicating that the risk-related BAL gene promotes malignant B cell migration. (Blood. 2000;96:4328-4334) PMID- 11110708 TI - Nucleophosmin-anaplastic lymphoma kinase associated with anaplastic large-cell lymphoma activates the phosphatidylinositol 3-kinase/Akt antiapoptotic signaling pathway. AB - More than half of anaplastic large-cell lymphomas (ALCLs) have a chromosomal translocation t(2;5) that leads to the expression of a hybrid protein composed of the nucleolar phosphoprotein nucleophosmin (NPM) and the anaplastic lymphoma kinase (ALK) that exhibits an unregulated tyrosine kinase activity. We have previously identified PLC-gamma as a crucial downstream signaling molecule of NPM ALK that contributes to its mitogenic potential. Here, we show that NPM-ALK recruits the C-terminal SH2 domain of the phosphatidylinositol 3-kinase (PI 3kinase) p85 subunit. PI 3-kinase assays revealed that the kinase is activated by NPM-ALK in vivo, in turn activating PKB/Akt in NPM-ALK-expressing cells. The use of 2 specific PI 3-kinase inhibitors, wortmannin and LY294002, demonstrated the requirement of PI 3-kinase for the growth of NPM-ALK-transformed cell lines, as well as a cell line established from a patient with ALCL. Primary murine bone marrow retrovirally transduced with NPM-ALK showed a transformed phenotype that was reversible on treatment with PI 3-kinase inhibitors. Flow cytometric analysis revealed that wortmannin-treated NPM-ALK-transformed cell lines underwent apoptosis. Furthermore, apoptosis induced by overexpression of the proapoptotic molecule Bad could be partially blocked by the overexpression of NPM-ALK. Thus, NPM-ALK activates the antiapoptotic PI 3-kinase/Akt pathway, which likely contributes to the molecular pathogenesis of ALCL. (Blood. 2000;96:4319-4327) PMID- 11110710 TI - An adherent condition is required for formation of multinuclear osteoclasts in the presence of macrophage colony-stimulating factor and receptor activator of nuclear factor kappa B ligand. AB - Identification of receptor activator of nuclear factor-kappaB (RANK) and RANK ligand (RANKL) has provided new insights into the osteoclast differentiation pathway. Osteoclast precursor cells were isolated using monoclonal antibodies against c-Fms and RANK, and the effect of adherence on the in vitro differentiation and proliferation of these cells was examined in 2 different types of stromal-cell-free culture systems: a semisolid culture medium (a nonadherent system) and a liquid culture medium (an adherent system). Osteoclast precursor cells were not able to differentiate into mature osteoclasts efficiently in the semisolid culture system. Trimerized RANKL enhanced osteoclast differentiation in semisolid cultures, but not to the extent seen when cells were allowed to adhere to plastic. Initial precursor cells were capable of differentiating into macrophages or osteoclasts. Once these cells were transferred to adherent conditions, striking differentiation was induced. Multinuclear cells were observed even after they had displayed phagocytic activity, which suggests that cell adhesion plays an important role in the differentiation of osteoclast precursor cells. Integrins, especially the arginine glycine-aspartic acid (RGD)-recognizing integrins alpha(v) and beta(3), were needed for osteoclast-committed precursor cells to proliferate in order to form multinuclear osteoclasts, and the increase in cell density affected the formation of multinuclear cells. A model of osteoclast differentiation with 2 stages of precursor development is proposed: (1) a first stage, in which precursor cells are bipotential and capable of anchorage-independent growth, and (2) a second stage, in which the further proliferation and differentiation of osteoclast committed precursor cells is anchorage-dependent. (Blood. 2000;96:4335-4343) PMID- 11110711 TI - Similar pattern of thymic-dependent T-cell reconstitution in infants with severe combined immunodeficiency after human leukocyte antigen (HLA)-identical and HLA nonidentical stem cell transplantation. AB - Donor T cells after stem cell transplantation reconstitute by 2 different pathways: by expansion from grafted, mature T cells and by intrathymic maturation from progenitor cells. This study characterized thymic-dependent reconstitution of CD4(+) T cells following different transplant modalities in patients with severe combined immunodeficiency (SCID). Three groups of patients were studied: one group after transplantation from human leukocyte antigen (HLA)-identical siblings with unmanipulated grafts without conditioning, a second group after transplantation from HLA-nonidentical parents with T-cell-depleted grafts without preconditioning, and a third group with prior conditioning. Reconstitution of the T-cell compartment was monitored by determining the expression of CD45 isoforms by developing CD4(+) cells in the peripheral blood and in discriminating expanded (CD45RO(+)) and newly generated (CD45RA(+)) T cells. Concomitantly, changes in the size of the thymus were evaluated sequentially by ultrasonography. Reconstitution of CD4(+)CD45RA(+) cells was delayed in all patients for several months, including patients after HLA-identical transplantation, and was always paralleled by normalization of the size of the thymus. No engraftment of donor progenitor cells was observed, as studied in one patient transplanted without conditioning. CD4(+)CD45RO(+) cells were detected early after transplantation only in patients given unmanipulated grafts. The study showed that thymic dependent T-cell maturation in these patients with SCID runs an autonomous course, independent of graft manipulation, of major HLA disparities, and of whether conditioning is used or not. In addition, thymic maturation may not require engraftment of donor-derived CD34(+) cells in the marrow. (Blood. 2000;96:4344-4349) PMID- 11110713 TI - Frequency and kinetics of polyclonal and clonal B cells in the peripheral blood of patients being treated for multiple myeloma. AB - Recent studies concerning the numbers of circulating clonal B cells in patients with multiple myeloma (MM) have reported conflicting data regarding the exact level and phenotype of clonal B cells and their response to treatment. In this report we document that the peripheral blood tumor burden at presentation was reduced by induction therapy to a low level, regardless of the initial tumor burden. However, the residual clonal compartment persisted before and after transplant. The level of clonal cells showed no correlation with CD19(+) cell levels. In a single patient with MM, high numbers of phenotypically aberrant clonal cells with altered CD19 expression were identified. (Blood. 2000;96:4357 4359) PMID- 11110712 TI - Keratinocyte growth factor facilitates alloengraftment and ameliorates graft versus-host disease in mice by a mechanism independent of repair of conditioning induced tissue injury. AB - We have previously shown that pretreatment of mice with keratinocyte growth factor (KGF), an epithelial tissue repair factor, can ameliorate graft-versus host disease (GVHD) after intensive chemoradiotherapeutic conditioning and allogeneic bone marrow transplantation (BMT). To determine whether this effect was dependent on a KGF-mediated mechanism affecting repair of conditioning induced epithelial cell injury, we studied GVHD in the absence of conditioning using BALB/c severe combined immune-deficient (SCID) recipients given C57BL/6 T cells. KGF (5 mg/kg per day, subcutaneously) given either before or after T-cell transfer enhanced body weights and extended survival. KGF-treated recipients had elevated serum levels of the Th2 cytokine interleukin 13 (IL-13) on day 6 after T cell transfer concomitant with reduced levels of the inflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interferon gamma (IFN-gamma). A 3-day KGF pretreatment also depressed the secondary in vitro mixed lymphocyte response (MLR) of C57BL/6 splenocytes taken 7 days after in vivo alloimmunization with irradiated BALB/c spleen cells. To determine whether KGF would inhibit host antidonor-mediated BM rejection, pan-T-cell-depleted BALB/c BM cells were infused into sublethally irradiated C57BL/6 mice and administered KGF either before or before and after BMT. Surprisingly, all KGF schedules tested actually resulted in enhanced alloengraftment. The presence of KGF receptor on donor antihost alloreactive T cells could not be detected by binding studies with radiolabeled KGF, reverse transcriptase-polymerase chain reaction, and Western blotting. Therefore, the mechanism of action of KGF on inhibiting T-cell-mediated immune effects may not be due to a direct effect of KGF on T cells. These studies demonstrate that KGF, by mechanisms independent of repair of conditioning-induced injury, has great potential as an anti-GVHD therapeutic agent with the added benefit of inhibiting the rejection of pan-T-cell-depleted donor BM allografts. (Blood. 2000;96:4350-4356) PMID- 11110714 TI - t(3;11) translocation in treatment-related acute myeloid leukemia fuses MLL with the GMPS (GUANOSINE 5' MONOPHOSPHATE SYNTHETASE) gene. AB - The partner gene of MLL was identified in a patient with treatment-related acute myeloid leukemia in which the karyotype suggested t(3;11)(q25;q23). Prior therapy included the DNA topoisomerase II inhibitors, teniposide and doxorubicin. Southern blot analysis indicated that the MLL gene was involved in the translocation. cDNA panhandle polymerase chain reaction (PCR) was used, which does not require partner gene-specific primers, to identify the chimeric transcript. Reverse-transcription of first-strand cDNAs with oligonucleotides containing known MLL sequence at the 5' ends and random hexamers at the 3' ends generated templates with an intra-strand loop for PCR. In-frame fusions of either MLL exon 7 or exon 8 with the GMPS (GUANOSINE 5'-MONOPHOSPHATE SYNTHETASE) gene from chromosome band 3q24 were detected. The fusion transcript was alternatively spliced. Guanosine monophosphate synthetase is essential for de novo purine synthesis. GMPS is the first partner gene of MLL on chromosome 3q and the first gene of this type in leukemia-associated translocations. (Blood. 2000;96:4360 4362) PMID- 11110715 TI - Familial-skewed X-chromosome inactivation as a predisposing factor for late-onset X-linked sideroblastic anemia in carrier females. AB - X-linked sideroblastic anemia (XLSA) is caused by mutations in the erythroid specific 5-aminolevulinic acid synthase (ALAS2) gene. An elderly woman who presented with an acquired sideroblastic anemia is studied. Molecular analysis revealed that she was heterozygous for a missense mutation in the ALAS2 gene, but she expressed only the mutated gene in reticulocytes. Her 2 daughters and a granddaughter were heterozygous for this mutation, had normal hemoglobin levels, and expressed the normal ALAS2 gene in reticulocytes. A grandson with a previous diagnosis of thalassemia intermedia was found to be hemizygous for the ALAS2 mutation. Treatment with pyridoxine completely corrected the anemia both in the proband and her grandson. All women who were analyzed in this family showed skewed X-chromosome inactivation in leukocytes, which indicated a hereditary condition associated with unbalanced lyonization. Because the preferentially active X chromosome carried the mutant ALAS2 allele, acquired skewing in the elderly likely worsened the genetic condition and abolished the normal ALAS2 allele expression in the proband. (Blood. 2000;96:4363-4365) PMID- 11110716 TI - Abnormalities of primitive myeloid progenitor cells expressing granulocyte colony stimulating factor receptor in patients with severe congenital neutropenia. AB - To define the basis for faulty granulopoiesis in patients with severe congenital neutropenia (SCN), the expression of granulocyte colony-stimulating factor receptor (G-CSFR) in primitive myeloid progenitor cells and their responsiveness to hematopoietic factors were studied. Flow cytometric analysis of bone marrow cells based on the expression of CD34, Kit receptor, and G-CSFR demonstrated a reduced frequency of CD34(+)/Kit(+)/ G-CSFR(+) cells in patients with SCN. The granulocyte-macrophage colony formation of CD34(+)/Kit(+)/G-CSFR(+) cells in patients was markedly decreased in response to G-CSF alone and to the combination of stem cell factor, the ligand for flk2/flt3, and IL-3 with or without G-CSF in serum-deprived semisolid culture. In contrast, no difference in the responsiveness of CD34(+)/Kit(+)/G-CSFR(-) cells was noted between patients with SCN and subjects without SCN. These results demonstrate that the presence of qualitative and quantitative abnormalities of primitive myeloid progenitor cells expressing G-CSFR may play an important role in the impairment of granulopoiesis in patients with SCN. (Blood. 2000;96:4366-4369) PMID- 11110717 TI - A new mutation in the HNF4 binding region of the factor VII promoter in a patient with severe factor VII deficiency. AB - Investigation of the molecular basis of a severe factor VII (fVII) deficiency revealed compound heterozygosity in the fVII gene. On the paternal allele the patient had 3 structural gene abnormalities frequently associated with fVII deficiency. A new mutation, a C to T transition at position -55 relative to the translational start site, was found on the maternal allele. The study demonstrates that this mutation partially impeded binding of the transcriptional activator, hepatic nuclear factor 4, to the fVII promoter while greatly reducing reporter gene expression in hepatic cells. (Blood. 2000;96:4370-4372) PMID- 11110718 TI - Unbalanced X-chromosome inactivation with a novel FVIII gene mutation resulting in severe hemophilia A in a female. AB - This report is of a 14-month-old girl affected with severe hemophilia A. Both her parents had normal values for factor VIII activity, and von Willebrand disease type 2N was excluded. Karyotype analysis demonstrated no obvious alteration, and BclI Southern blot did not reveal F8 gene inversions. Direct sequencing of F8 gene exons revealed a frameshift-stop mutation (Q565delC/ter566) in the heterozygous state in the proposita only. F8 gene polymorphism analysis indicated that the mutation must have occurred de novo in the paternal germline. Furthermore, analysis of the pattern of X chromosome methylation at the human androgen receptor gene locus demonstrated a skewed inactivation of the derived maternal X chromosome from the lymphocytes of the proband's DNA. Thus, the severe hemophilia A in the proposita results from a de novo F8 gene frameshift-stop mutation on the paternally derived X chromosome, associated with a nonrandom pattern of inactivation of the maternally derived X chromosome. (Blood. 2000;96:4373-4375) PMID- 11110719 TI - Non-specific effects of vaccines in developing countries. We need evidence about the effect of vaccines on mortality from all causes. PMID- 11110720 TI - Does physical activity prevent cancer? Evidence suggests protection against colon cancer and probably breast cancer. PMID- 11110721 TI - Age related macular degeneration. New hope for a common problem comes from photodynamic therapy. PMID- 11110722 TI - Stem cell research. The Uk government should sanction carefully regulated research. PMID- 11110723 TI - Quality improvement reports: a new kind of article. They should allow authors to describe improvement projects so others can learn. PMID- 11110724 TI - Consultant suspended for research fraud. PMID- 11110726 TI - In brief PMID- 11110728 TI - Statins could be useful in organ transplantation PMID- 11110727 TI - Antenatal testing reduces vertical transmission of HIV. PMID- 11110729 TI - Milburn to monitor implementation of NICE guidance. National Institute for Clinical Excellence. PMID- 11110730 TI - Prescription fraud costs Wales pound 15m a year. PMID- 11110731 TI - Netherlands gives more protection to doctors in euthanasia cases. PMID- 11110733 TI - GMC decides on a new structure PMID- 11110732 TI - UK defines action to tackle health in developing countries. PMID- 11110734 TI - Routine vaccinations and child survival: follow up study in Guinea-Bissau, West Africa. AB - OBJECTIVE: To examine the association between routine childhood vaccinations and survival among infants in Guinea-Bissau. DESIGN: Follow up study. PARTICIPANTS: 15 351 women and their children born during 1990 and 1996. SETTING: Rural Guinea Bissau. MAIN OUTCOME MEASURES: Infant mortality over six months (between age 0-6 months and 7-13 months for BCG, diphtheria, tetanus, and pertussis, and polio vaccines and between 7-13 months and 14-20 months for measles vaccine). RESULTS: Mortality was lower in the group vaccinated with any vaccine compared with those not vaccinated, the mortality ratio being 0.74 (95% confidence interval 0.53 to 1.03). After cluster, age, and other vaccines were adjusted for, BCG was associated with significantly lower mortality (0.55 (0.36 to 0.85)). However, recipients of one dose of diphtheria, tetanus, and pertussis or polio vaccines had higher mortality than children who had received none of these vaccines (1.84 (1.10 to 3.10) for diphtheria, tetanus, and pertussis). Recipients of measles vaccine had a mortality ratio of 0.48 (0.27 to 0.87). When deaths from measles were excluded from the analysis the mortality ratio was 0.51 (0.28 to 0.95). Estimates were unchanged by controls for background factors. CONCLUSIONS: These trends are unlikely to be explained exclusively by selection biases since different vaccines were associated with opposite tendencies. Measles and BCG vaccines may have beneficial effects in addition to protection against measles and tuberculosis. PMID- 11110736 TI - Renal function-and how to assess it. PMID- 11110735 TI - Randomised controlled trial of dual blockade of renin-angiotensin system in patients with hypertension, microalbuminuria, and non-insulin dependent diabetes: the candesartan and lisinopril microalbuminuria (CALM) study. AB - OBJECTIVES: To assess and compare the effects of candesartan or lisinopril, or both, on blood pressure and urinary albumin excretion in patients with microalbuminuria, hypertension, and type 2 diabetes. DESIGN: Prospective, randomised, parallel group, double blind study with four week placebo run in period and 12 weeks' monotherapy with candesartan or lisinopril followed by 12 weeks' monotherapy or combination treatment. SETTING: Tertiary hospitals and primary care centres in four countries (37 centres). PARTICIPANTS: 199 patients aged 30-75 years. INTERVENTIONS: Candesartan 16 mg once daily, lisinopril 20 mg once daily. MAIN OUTCOME MEASURES: Blood pressure and urinary albumin:creatinine ratio. RESULTS: At 12 weeks mean (95% confidence interval) reductions in diastolic blood pressure were 9.5 mm Hg (7.7 mm Hg to 11.2 mm Hg, P<0.001) and 9.7 mm Hg (7.9 mm Hg to 11.5 mm Hg, P<0.001), respectively, and in urinary albumin:creatinine ratio were 30% (15% to 42%, P<0.001) and 46% (35% to 56%, P<0.001) for candesartan and lisinopril, respectively. At 24 weeks the mean reduction in diastolic blood pressure with combination treatment (16.3 mm Hg, 13.6 mm Hg to 18.9 mm Hg, P<0. 001) was significantly greater than that with candesartan (10.4 mm Hg, 7.7 mm Hg to 13.1 mm Hg, P<0.001) or lisinopril (mean 10.7 mm Hg, 8.0 mm Hg to 13.5 mm Hg, P<0.001). Furthermore, the reduction in urinary albumin:creatinine ratio with combination treatment (50%, 36% to 61%, P<0.001) was greater than with candesartan (24%, 0% to 43%, P=0.05) and lisinopril (39%, 20% to 54%, P<0.001). All treatments were generally well tolerated. CONCLUSION: Candesartan 16 mg once daily is as effective as lisinopril 20 mg once daily in reducing blood pressure and microalbuminuria in hypertensive patients with type 2 diabetes. Combination treatment is well tolerated and more effective in reducing blood pressure. PMID- 11110737 TI - Efficacy and safety of galantamine in patients with mild to moderate Alzheimer's disease: multicentre randomised controlled trial. Galantamine International-1 Study Group. AB - OBJECTIVE: To evaluate the efficacy and safety of galantamine in the treatment of Alzheimer's disease. DESIGN: Randomised, double blind, parallel group, placebo controlled trial. SETTING: 86 outpatient clinics in Europe and Canada. PARTICIPANTS: 653 patients with mild to moderate Alzheimer's disease. INTERVENTION: Patients randomly assigned to galantamine had their daily dose escalated over three to four weeks to maintenance doses of 24 or 32 mg. MAIN OUTCOME MEASURES: Scores on the 11 item cognitive subscale of the Alzheimer's disease assessment scale, the clinician's interview based impression of change plus caregiver input, and the disability assessment for dementia scale. The effect of apolipoprotein E4 genotype on reponse to treatment was also assessed. RESULTS: At six months, patients who received galantamine had a significantly better outcome on the 11 item cognitive subscale of the Alzheimer's disease assessment scale than patients in the placebo group (mean treatment effect 2.9 points for lower dose and 3.1 for higher dose, intention to treat analysis, P<0.001 for both doses). Galantamine was more effective than placebo on the clinician's interview based impression of change plus caregiver input (P<0.05 for both doses v placebo). At six months, patients in the higher dose galantamine group had significantly better scores on the disability assessment for dementia scale than patients in the placebo group (mean treatment effect 3.4 points, P<0.05). Apolipoprotein E genotype had no effect on the efficacy of galantamine. 80% (525) of patients completed the study. CONCLUSION: Galantamine is effective and well tolerated in Alzheimer's disease. As galantamine slowed the decline of functional ability as well as cognition, its effects are likely to be clinically relevant. PMID- 11110738 TI - Recurrent generalised urticaria at insulin injection sites. PMID- 11110740 TI - Putting women in control PMID- 11110742 TI - It's like standing on a deserted platform PMID- 11110739 TI - Problem solving treatment and group psychoeducation for depression: multicentre randomised controlled trial. Outcomes of Depression International Network (ODIN) Group. AB - OBJECTIVES: To determine the acceptability of two psychological interventions for depressed adults in the community and their effect on caseness, symptoms, and subjective function. DESIGN: A pragmatic multicentre randomised controlled trial, stratified by centre. SETTING: Nine urban and rural communities in Finland, Republic of Ireland, Norway, Spain, and the United Kingdom. PARTICIPANTS: 452 participants aged 18 to 65, identified through a community survey with depressive or adjustment disorders according to the international classification of diseases, 10th revision or Diagnostic and Statistical Manual of Mental Disorders, fourth edition. INTERVENTIONS: Six individual sessions of problem solving treatment (n=128), eight group sessions of the course on prevention of depression (n=108), and controls (n=189). MAIN OUTCOME MEASURES: Completion rates for each intervention, diagnosis of depression, and depressive symptoms and subjective function. RESULTS: 63% of participants assigned to problem solving and 44% assigned to prevention of depression completed their intervention. The proportion of problem solving participants depressed at six months was 17% less than that for controls, giving a number needed to treat of 6; the mean difference in Beck depression inventory score was -2. 63 (95% confidence interval -4.95 to -0.32), and there were significant improvements in SF-36 scores. For depression prevention, the difference in proportions of depressed participants was 14% (number needed to treat of 7); the mean difference in Beck depression inventory score was -1.50 (-4.16 to 1.17), and there were significant improvements in SF-36 scores. Such differences were not observed at 12 months. Neither specific diagnosis nor treatment with antidepressants affected outcome. CONCLUSIONS: When offered to adults with depressive disorders in the community, problem solving treatment was more acceptable than the course on prevention of depression. Both interventions reduced caseness and improved subjective function. PMID- 11110741 TI - Prevention of ischaemic stroke. PMID- 11110743 TI - Effect of multifaceted intervention promoting early switch from intravenous to oral acetaminophen for postoperative pain: controlled, prospective, before and after study. AB - PROBLEM: Need to improve the efficiency of postoperative pain management by early switching from intravenous to oral acetaminophen. DESIGN: Implementation of local guidelines aimed at improving nurses' and doctors' behaviour. A controlled, prospective, before and after study evaluated its impact on appropriateness and costs. BACKGROUND AND SETTING: Orthopaedic surgery department (intervention) and all other surgical departments (control) of a university hospital. Five anaesthetists and 30 nurses of orthopaedic department participated in study. KEY MEASURES FOR IMPROVEMENT: Reducing number of acetaminophen injections per patient, reducing consumption of acetaminophen injections; cost savings over a one year period. STRATEGIES FOR IMPROVEMENT: Multifaceted intervention included a local consensus process, short educational presentation, poster displayed in all nurses' offices, and feedback of practices six months after implementation of guidelines. EFFECTS OF CHANGE: Mean number of acetaminophen injections per patient decreased from 6.81 before intervention to 2.36 six months after. Monthly consumption of acetaminophen injections per 100 patients decreased by 320.9 (95% confidence interval 192.4 to 449.4) in intervention department and remained unchanged in control departments. Annual cost reduction was projected to be pound 15,100. LESSONS LEARNT: Simple and locally implemented guidelines can improve practices and cut costs. Educational interventions can improve professionals' behaviour when they are based on actual working practices, use interactive techniques such as discussion groups, and are associated with other effective implementation strategies. PMID- 11110744 TI - Results of genetic testing: when confidentiality conflicts with a duty to warn relatives. PMID- 11110745 TI - Obituaries PMID- 11110746 TI - Fergie and parents' diet dilemmas. PMID- 11110747 TI - Drugs for Alzheimer's disease PMID- 11110748 TI - Charity spice PMID- 11110749 TI - Auntz PMID- 11110750 TI - So many places for learning PMID- 11110751 TI - Vaccines may have non-specific effects in developing countries PMID- 11110753 TI - Galantamine slows functional decline in Alzheimer's disease PMID- 11110752 TI - Diabetic patients with hyper- tension benefit from dual blockade of renin angiotensin system PMID- 11110754 TI - Psychological interventions can reduce depressive disorders in community settings PMID- 11110755 TI - Drug administration can be improved by simple guidelines discussed by all staff affected PMID- 11110756 TI - It is difficult to apply a moral rule for strangers to family obligations PMID- 11110757 TI - Mountain rescue medicine PMID- 11110758 TI - Acknowledgment to reviewer PMID- 11110759 TI - The oxidative paradox: another piece in the puzzle. PMID- 11110760 TI - Platelets: unindicted coconspirators in inflammatory tissue injury. PMID- 11110761 TI - The cellular actions of beta-adrenergic receptor agonists: looking beyond cAMP. PMID- 11110762 TI - Repolarization alternans: toward a unifying theory of reentrant arrhythmia induction. PMID- 11110763 TI - G proteins and heart failure: is Galphaq a novel target for heart failure? PMID- 11110764 TI - Integrative analysis of calcium cycling in cardiac muscle. AB - The control of intracellular calcium is central to regulation of contractile force in cardiac muscle. This review illustrates how analysis of the control of calcium requires an integrated approach in which several systems are considered. Thus, the calcium content of the sarcoplasmic reticulum (SR) is a major determinant of the amount of Ca(2+) released from the SR and the amplitude of the Ca(2+) transient. The amplitude of the transient, in turn, controls Ca(2+) fluxes across the sarcolemma and thence SR content. This control of SR content influences the response to maneuvers that modify, for example, the properties of the SR Ca(2+) release channel or ryanodine receptor. Specifically, modulation of the open probability of the ryanodine receptor produces only transient effects on the Ca(2+) transient as a result of changes of SR content. These interactions between various Ca(2+) fluxes are modified by the Ca(2+) buffering properties of the cell. Finally, we predict that, under some conditions, the above interactions can result in instability (such as alternans) rather than ordered control of contractility. PMID- 11110765 TI - Regulation of cardiac L-type calcium channels by protein kinase A and protein kinase C. AB - Voltage-dependent L-type Ca(2+) channels are multisubunit transmembrane proteins, which allow the influx of Ca(2+) (I:(Ca)) essential for normal excitability and excitation-contraction coupling in cardiac myocytes. A variety of different receptors and signaling pathways provide dynamic regulation of I:(Ca) in the intact heart. The present review focuses on recent evidence describing the molecular details of regulation of L-type Ca(2+) channels by protein kinase A (PKA) and protein kinase C (PKC) pathways. Multiple G protein-coupled receptors act through cAMP/PKA pathways to regulate L-type channels. ss-Adrenergic receptor stimulation results in a marked increase in I:(Ca), which is mediated by a cAMP/PKA pathway. Growing evidence points to an important role of localized signaling complexes involved in the PKA-mediated regulation of I:(Ca), including A-kinase anchor proteins and binding of phosphatase PP2a to the carboxyl terminus of the alpha(1C) (Ca(v)1.2) subunit. Both alpha(1C) and ss(2a) subunits of the channel are substrates for PKA in vivo. The regulation of L-type Ca(2+) channels by Gq-linked receptors and associated PKC activation is complex, with both stimulation and inhibition of I:(Ca) being observed. The amino terminus of the alpha(1C) subunit is critically involved in PKC regulation. Crosstalk between PKA and PKC pathways occurs in the modulation of I:(Ca). Ultimately, precise regulation of I:(Ca) is needed for normal cardiac function, and alterations in these regulatory pathways may prove important in heart disease. PMID- 11110766 TI - Ventricular fibrillation: how do we stop the waves from breaking? AB - Combined experimental and theoretical developments have demonstrated that in addition to preexisting electrophysiological heterogeneities, cardiac electrical restitution properties contribute to breakup of reentrant wavefronts during cardiac fibrillation. Developing therapies that favorably alter electrical restitution properties have promise as a new paradigm for preventing fibrillation. PMID- 11110767 TI - Role of endothelium-derived nitric oxide in the regulation of cardiac oxygen metabolism: implications in health and disease. AB - Endothelium-derived NO is considered to be primarily an important determinant of vascular tone and platelet activity; however, the modulation of myocardial metabolism by NO may be one of its most important roles. This modulation may be critical for the regulation of tissue metabolism. Several physiological processes act in concert to make endothelial NO synthase-derived NO potentially important in the regulation of mitochondrial respiration in cardiac tissue, including (1) the nature of the capillary network in the myocardium, (2) the diffusion distance for NO, (3) the low toxicity of NO at physiological (nanomolar) concentrations, (4) the fact that low PO(2) in tissue facilitates the action of NO on cytochrome oxidase, and (5) the formation of oxygen free radicals. A decrease in NO production is involved in the pathophysiological modifications that occur in heart failure and diabetes, disease states associated with altered cardiac metabolism that contributes to the evolution of the disease process. In contrast, several drugs (eg, angiotensin-converting enzyme inhibitors, amlodipine, and statins) can restore or maintain endogenous production of NO by endothelial cells, and this mechanism may explain part of their therapeutic efficiency. Thus, the purpose of this review is to critically evaluate the role of NO in the control of mitochondrial respiration, with special emphasis on its effect on cardiac metabolism. PMID- 11110768 TI - Current perspectives on the use of gene therapy for hypertension. AB - Systemic hypertension is a pathophysiological state that is manifested as high blood pressure and is a major risk factor for stroke, ischemic heart disease, peripheral vascular disease, and progressive renal damage. Pulmonary hypertension occurs in 3 distinct forms: primary pulmonary hypertension, pulmonary hypertension of the newborn, or secondary pulmonary hypertension attributable to a variety of lung and cardiovascular diseases. This review discusses the use of gene therapy in the control of systemic and pulmonary hypertension. Overexpression of vasodilator genes as well as antisense knockdown of vasoconstrictor genes has been successfully used in animal models of both forms of hypertension. Furthermore, the use of viral vectors to deliver these constructs has achieved long-term control of hypertension. The successful establishment of gene therapy techniques in the animal models of hypertension coupled with the anticipated advances in the genetic aspects of this disease would make it highly feasible to attempt gene delivery in the control of human hypertension. PMID- 11110770 TI - Diabetes-induced vascular hypertrophy is accompanied by activation of Na(+)-H(+) exchange and prevented by Na(+)-H(+) exchange inhibition. AB - Vascular disease often involves vessel hypertrophy with underlying cellular hypertrophy or hyperplasia. Experimental diabetes stimulates hypertrophy of the rat mesenteric vasculature, and we investigated the hypothesis that this hypertrophy is associated with activation of Na(+)-H(+) exchange (NHE) activity. We measured the NHE activity in isolated, intact blood vessels from control and streptozotocin-induced diabetic adult rats using concurrent myography and fluorescence spectroscopy. The role of inhibiting NHE activity in preventing the development of the mesenteric hypertrophy in streptozotocin-diabetic rats was investigated by administration of cariporide (100 mg/kg body weight per day in 3 doses by gavage) after induction of diabetes and subsequently determining vessel weight and structure. The weight of the mesenteric vasculature was not increased 1 week after streptozotocin treatment but was significantly increased by an average of 56% at 3 weeks. NHE activity in mesenteric arteries showed an enhanced maximal velocity (V:(max)) in diabetic vessels at 1 and 3 weeks (0.246+/-0.006 and 0. 238+/-0.007 versus 0.198+/-0.007 pH U/min) with no change in the apparent K:(m). Moreover, NHE-1 mRNA in mesenteric arterioles at 3 weeks after streptozotocin treatment was increased by >60% (55.8+/-6. 4 versus 91.3+/-12.3 fg). Administration of cariporide significantly reduced mesenteric vascular weight, the wall/lumen ratio, and mesenteric extracellular matrix accumulation in the diabetic animals. Our study shows that diabetes in vivo correlates with elevated NHE activity and mRNA in the mesenteric vasculature and furthermore that inhibition of this system prevents the hypertrophic response. These data suggest that NHE may be a target for therapeutic modulation of vascular changes in diabetes. PMID- 11110769 TI - Myocardial cell death in human diabetes. AB - The renin-angiotensin system is upregulated with diabetes, and this may contribute to the development of a dilated myopathy. Angiotensin II (Ang II) locally may lead to oxidative damage, activating cardiac cell death. Moreover, diabetes and hypertension could synergistically impair myocardial structure and function. Therefore, apoptosis and necrosis were measured in ventricular myocardial biopsies obtained from diabetic and diabetic-hypertensive patients. Accumulation of a marker of oxidative stress, nitrotyrosine, and Ang II labeling were evaluated quantitatively. The diabetic heart showed cardiac hypertrophy, cavitary dilation, and depressed ventricular performance. These alterations were more severe with diabetes and hypertension. Diabetes was characterized by an 85 fold, 61-fold, and 26-fold increase in apoptosis of myocytes, endothelial cells, and fibroblasts, respectively. Apoptosis in cardiac cells did not increase additionally with diabetes and hypertension. Diabetes increased necrosis by 4 fold in myocytes, 9-fold in endothelial cells, and 6-fold in fibroblasts. However, diabetes and hypertension increased necrosis by 7-fold in myocytes and 18-fold in endothelial cells. Similarly, Ang II labeling in myocytes and endothelial cells increased more with diabetes and hypertension than with diabetes alone. Nitrotyrosine localization in cardiac cells followed a comparable pattern. In spite of the difference in the number of nitrotyrosine-positive cells with diabetes and with diabetes and hypertension, apoptosis and necrosis of myocytes, endothelial cells, and fibroblasts were detected only in cells containing this modified amino acid. In conclusion, local increases in Ang II with diabetes and with diabetes and hypertension may enhance oxidative damage, activating cardiac cell apoptosis and necrosis. PMID- 11110771 TI - A role for platelets and endothelial selectins in tumor necrosis factor-alpha induced leukocyte recruitment in the brain microvasculature. AB - The mechanisms mediating leukocyte recruitment into the cerebral nervous system during inflammation are still poorly understood. The objective of this study was to investigate the leukocyte recruitment in the brain microcirculation by intravital microscopy. Superfusion of the brain with artificial cerebrospinal fluid did not induce leukocyte rolling or adhesion. However, intraperitoneal tumor necrosis factor-alpha (TNF-alpha) caused marked leukocyte rolling and adhesion in the brain microcirculation. Histology revealed that the recruitment was primarily of neutrophils. Both E- and P-selectin were required for TNF-alpha induced leukocyte recruitment, as rolling was reduced after treatment with either anti-E- or anti-P-selectin antibody and eliminated in E- or P-selectin-deficient mice. A significant increase in brain P- and E-selectin expression was seen after TNF-alpha treatment, but both were an order of magnitude less than in any other tissue. We observed significant platelet paving of TNF-alpha-stimulated endothelium and found that anti-platelet antibody reduced leukocyte rolling and adhesion, as did acetylsalicylic acid (aspirin). However, depletion of platelets did not reduce cerebral P-selectin expression. Moreover, chimeric mice lacking P selectin on endothelium but not platelets had significantly decreased P-selectin expression and reduced leukocyte recruitment in the brain. This suggests a role for endothelial P-selectin in cerebral leukocyte recruitment. In conclusion, TNF alpha-induced neutrophil recruitment into the brain requires both endothelial E selectin and P-selectin as well as platelets, but platelet P-selectin was not a major contributor to this process. PMID- 11110772 TI - Prolonged hypercapnia-evoked cerebral hyperemia via K(+) channel- and prostaglandin E(2)-dependent endothelial nitric oxide synthase induction. AB - Mechanisms for secondary sustained increase in cerebral blood flow (CBF) during prolonged hypercapnia are unknown. We show that induction of endothelial NO synthase (eNOS) by an increase in prostaglandins (PGs) contributes to the secondary CBF increase during hypercapnic acidosis. Ventilation of pigs with 6% CO(2) (PaCO(2 approximately)65 mm Hg; pH approximately 7.2) caused a approximately 2.5-fold increase in CBF at 30 minutes, which declined to basal values at 3 hours and gradually rose again at 6 and 8 hours; the latter increase was associated with PG elevation, nitrite formation, eNOS mRNA expression, and in situ NO synthase (NOS) reactivity (NADPH-diaphorase staining). Subjecting free floating brain sections to acidotic conditions increased eNOS expression, the time course of which was similar to that of CBF increase. Treatment of pigs with the cyclooxygenase inhibitor diclofenac or the NOS inhibitor Nomega-nitro-L arginine blunted the initial rise and prevented the secondary CBF increase during hypercapnic acidosis; neuronal NOS blockers 1-(2-trifluoromethylphenyl) imidazole and 3-bromo-7-nitroindazole were ineffective. Diclofenac abolished the hypercapnia-induced rise in cerebrovascular nitrite production, eNOS mRNA expression, and NADPH-diaphorase reactivity. Acidosis (pH approximately 7.15, PCO(2 approximately )40 mm Hg; 6 hours) produced similar increases in prostaglandin E(2) (PGE(2)) and eNOS mRNA levels in isolated brain microvessels and in NADPH-diaphorase reactivity of brain microvasculature; these changes were prevented by diclofenac, by the receptor-operated Ca(2+) channel blocker SK&F96365, and by the K(ATP) channel blocker glybenclamide. Acidosis increased Ca(2+) transients in brain endothelial cells, which were blocked by glybenclamide and SK&F96365 but not by diclofenac. Increased PG-related eNOS mRNA and NO dependent vasorelaxation to substance P was detected as well in rat brain exposed to 6 hours of hypercapnia. PGE(2) was the only major prostanoid that modulated brain eNOS expression during acidosis. Thus, in prolonged hypercapnic acidosis, the secondary CBF rise is closely associated with induction of eNOS expression; this seems to be mediated by PGE(2) generated by a K(ATP) and Ca(2+) channel dependent process. PMID- 11110773 TI - Role of structural barriers in the mechanism of alternans-induced reentry. AB - Previously, using an animal model of T-wave alternans in structurally normal myocardium, we demonstrated that repolarization can alternate with opposite phase between neighboring myocytes (ie, discordant alternans), causing spatial dispersions of repolarization that form the substrate for functional block and reentrant ventricular fibrillation (VF). However, the mechanisms responsible for cellular discordant alternans and its electrocardiographic manifestation (ie, T wave alternans) in patients with structural heart disease are unknown. We hypothesize that electrotonic uncoupling between neighboring regions of cells by a structural barrier (SB) is a mechanism for discordant alternans. Using voltage sensitive dyes, ventricular action potentials were recorded from 26 Langendorff perfused guinea pig hearts in the absence (ie, control) and presence of an insulating SB produced by an epicardial laser lesion. Quantitative analysis of magnitude and phase of cellular alternans revealed that in controls, action potential duration alternated in phase at all ventricular sites above a critical heart rate (269+/-17 bpm), ie, concordant alternans. Also, above a faster critical heart rate threshold (335+/-24 bpm), action potential duration alternated with opposite phase between sites, ie, discordant alternans. In contrast, only discordant but not concordant alternans was observed in 80% of hearts with the SB, and discordant alternans always occurred at a significantly slower heart rate (by 68+/-28 bpm) compared with controls. Therefore, the SB had a major effect on the alternans-heart rate relation, which served to facilitate the development of discordant alternans. Whether a SB was present or not, discordant alternans produced considerable increases (by approximately 170%) in the maximum spatial gradient of repolarization, which in turn formed the substrate for unidirectional block and reentry. However, by providing a structural anchor for stable reentry, discordant alternans in the presence of a SB led most often to sustained monomorphic ventricular tachycardia rather than to VF, whereas in the absence of a SB discordant alternans caused VF. SBs facilitate development of discordant alternans between cells with different ionic properties by electrotonically uncoupling neighboring regions of myocardium. This may explain why arrhythmia-prone patients with structural heart disease exhibit T wave alternans at lower heart rates. These data also suggest a singular mechanism by which T-wave alternans forms a substrate for initiation of both VF and sustained monomorphic ventricular tachycardia. PMID- 11110774 TI - Hypoxia increases the sensitivity of the L-type Ca(2+) current to beta-adrenergic receptor stimulation via a C2 region-containing protein kinase C isoform. AB - The effects of hypoxia on the L-type Ca(2+) current (I:(Ca-L)) in the absence and presence of the ss-adrenergic receptor agonist isoproterenol (Iso) were examined. Exposing guinea pig ventricular myocytes to hypoxia alone resulted in a reversible inhibition of basal I:(Ca-L). When cells were exposed to Iso in the presence of hypoxia, the K:(0.5) for activation of I:(Ca-L) by Iso was significantly decreased from 5.3+/-0.7 to 1.6+/-0.1 nmol/L. The membrane impermeant thiol-specific oxidizing compound 5, 5'-dithio-bis(2-nitrobenzoic acid) (DTNB) attenuated the inhibition of basal I:(Ca-L) by hypoxia 81.3+/-9.4% but had no effect on the increase in sensitivity of I:(Ca-L) to Iso. In addition, DTT mimicked the effects of hypoxia on basal I:(Ca-L) and the increase in sensitivity to Iso. Neither the inhibitors of guanylate cyclase LY-83583 or methylene blue nor the NO synthase inhibitor N:(G)-monomethyl-L-arginine monoacetate had any effect on the basal inhibition of I:(Ca-L) or the decrease in K:(0.5) for activation of I:(Ca-L) by Iso during hypoxia. However, the protein kinase C (PKC) inhibitors bisindolylmaleimide I and Go 7874 significantly attenuated the increase in sensitivity of I:(Ca-L) to Iso. More specifically, the response was attenuated when cells were dialyzed with a peptide inhibitor of the C2 region-containing classical PKC isoforms. The same effect was not observed with the PKCepsilon peptide inhibitor. These results suggest that hypoxia regulates I:(Ca-L) through the following 2 distinct mechanisms: direct inhibition of basal I:(Ca-L) and an indirect effect on the sensitivity of the channel to ss adrenergic receptor stimulation that is mediated through a classical PKC isoform. PMID- 11110775 TI - The beta(2)-adrenergic receptor delivers an antiapoptotic signal to cardiac myocytes through G(i)-dependent coupling to phosphatidylinositol 3'-kinase. AB - Recent studies have shown that chronic beta-adrenergic receptor (beta-AR) stimulation alters cardiac myocyte survival in a receptor subtype-specific manner. We examined the effect of selective beta(1)- and beta(2)-AR subtype stimulation on apoptosis induced by hypoxia or H(2)O(2) in rat neonatal cardiac myocytes. Although neither beta(1)- nor beta(2)-AR stimulation had any significant effect on the basal level of apoptosis, selective beta(2)-AR stimulation protected myocytes from apoptosis. beta(2)-AR stimulation markedly increased mitogen-activated protein kinase/extracellular signal-regulated protein kinase (MAPK/ERK) activation as well as phosphatidylinositol-3'-kinase (PI-3K) activity and Akt/protein kinase B phosphorylation. beta(1)-AR stimulation also markedly increased MAPK/ERK activation but only minimally activated PI-3K and Akt. Pretreatment with pertussis toxin blocked beta(2)-AR-mediated protection from apoptosis as well as the beta(2)-AR-stimulated changes in MAPK/ERK, PI-3K, and Akt/protein kinase B. The selective PI-3K inhibitor, LY 294002, also blocked beta(2)-AR-mediated protection, whereas inhibition of MAPK/ERK activation at an inhibitor concentration that blocked agonist-induced activation but not the basal level of activation had no effect on beta(2)-AR-mediated protection. These findings demonstrate that beta(2)-ARs activate a PI-3K-dependent, pertussis toxin sensitive signaling pathway in cardiac myocytes that is required for protection from apoptosis-inducing stimuli often associated with ischemic stress. PMID- 11110776 TI - Cardiomyocyte apoptosis induced by Galphaq signaling is mediated by permeability transition pore formation and activation of the mitochondrial death pathway. AB - Expression of the wild-type alpha subunit of Gq stimulates phospholipase C and induces hypertrophy in cardiomyocytes. Addition of Gq-coupled receptor agonists additionally activates phospholipase C, as does expression of a constitutively active mutant form of Galphaq. Under these conditions, hypertrophy is rapidly succeeded by apoptotic cellular and molecular changes, including myofilament disorganization, loss of mitochondrial membrane potential, alterations in Bcl-2 family protein levels, DNA fragmentation, increased caspase activity ( approximately 4-fold), cytochrome c redistribution, and nuclear chromatin condensation in approximately 12% of the cells. We used various interventions to define the molecular relationships between these events and identify potential sites at which these features of apoptosis could be rescued. Treatment with caspase inhibitors prevented DNA fragmentation and promoted myocyte survival; however, cytochrome c release and loss of mitochondrial membrane potential still occurred. In contrast, treatment with bongkrekic acid, an inhibitor of the mitochondrial permeability transition pore, not only prevented DNA fragmentation and reduced nuclear chromatin condensation but also preserved mitochondrial membrane potential and limited cytochrome c redistribution to only approximately 2% of cells. These data demonstrate the central role of mitochondrial membrane potential in initiation of caspase activation and downstream apoptotic events and suggest that preservation of mitochondrial integrity is crucial for prolonging the life and function of cardiomyocytes exposed to pathological levels of stress. PMID- 11110777 TI - Shear stress-dependent regulation of the human beta-tubulin folding cofactor D gene. AB - The flowing blood generates shear stress at the endothelial cell surface. The endothelial cells modify their phenotype by alterations in gene expression in response to different levels of fluid shear stress. To identify genes involved in this process, human umbilical vein endothelial cells were exposed to laminar shear stress (venous or arterial levels) in a cone-and-plate apparatus for 24 hours. Using the method of RNA arbitrarily primed polymerase chain reaction, we cloned a polymerase chain reaction fragment representing an mRNA species downregulated by arterial compared with venous shear stress (shear stress downregulated gene-1, SSD-1). According to Northern blot analysis, corresponding SSD-1 cDNA clones revealed a similar, time-dependent downregulation after 24 hours of arterial shear stress compared with venous shear stress or static controls. Three SSD-1 mRNA species of 2.8, 4.1, and 4.6 kb were expressed in a tissue-specific manner. The encoded amino acid sequence of the human endothelial SSD-1 isoform (4.1-kb mRNA species) revealed 80.4% identity and 90.9% homology to the bovine ss-tubulin folding cofactor D (tfcD) gene. Downregulation of tfcD mRNA expression by shear stress was defined at the level of transcription by nuclear run-on assays. The tfcD protein was downregulated by arterial shear stress. The shear stress-dependent downregulation of tfcD mRNA and protein was attenuated by the NO synthase inhibitor Nomega-nitro-L-arginine methyl ester. Furthermore, the NO donor DETA-NO downregulated tfcD mRNA. Because tfcD was shown to be a microtubule-destabilizing protein, our data suggest a shear stress-dependent regulation of the microtubular dynamics in human endothelial cells. PMID- 11110778 TI - Role of NAD(P)H oxidase in angiotensin II-induced JAK/STAT signaling and cytokine induction. AB - Inflammatory processes involve both synthesis of inflammatory cytokines, such as interleukin-6 (IL-6), and the activation of their distinct signaling pathways, eg, the janus kinases (JAKs) and signal transducers and activators of transcription (STAT). Superoxide (O(2)(-)) anions activate this signaling cascade, and the vasoconstrictor angiotensin II (Ang II) enhances the formation of O(2)(-) anions via the NAD(P)H oxidase system in rat aortic smooth muscle cells. Ang II activates the JAK/STAT cascade via its type 1 (AT(1)) receptor and induces synthesis and release of IL-6. Therefore, we investigated the role of O(2)(-) anions generated by the NAD(P)H oxidase system on the Ang II activation of the JAK/STAT cascade and its impact on IL-6 synthesis. Ang II stimulation of rat aortic smooth muscle cells induced a rapid increase in O(2)(-) anions determined by laser fluoroscopy, which can be abolished by DPI, a flavoprotein inhibitor. Ang II-induced phosphorylation of JAK2, STAT1alpha/ss, STAT3, and IL-6 synthesis can be abolished by DPI, as determined by immunoprecipitations and Northern blot analysis. Electroporation of neutralizing antisera targeted against p47(phox), a NAD(P)H oxidase subunit, abolished Ang II-induced JAK/STAT activation and IL-6 synthesis. Inhibition of JAK2 by its inhibitor AG490 (10 micromol/L) blocked not only JAK2 activation but also IL-6 synthesis. These results suggest that stimulation of the JAK/STAT cascade by Ang II requires O(2)( ) anions generated by the NAD(P)H oxidase system, and O(2)(-) anion-dependent activation of the JAK/STAT cascade seems to be additionally involved in Ang II induced IL-6 synthesis. Thus, Ang II-induced inflammatory effects seem to require O(2)(-) anions generated by the NAD(P)H oxidase system. PMID- 11110779 TI - De novo expression of macrophage migration inhibitory factor in atherogenesis in rabbits. AB - Macrophage migration inhibitory factor (MIF) has been shown to play an important role in macrophage-mediated diseases. We investigate the potential role of MIF in atherogenesis using a hypercholesterolemic rabbit model. New Zealand White rabbits fed with a 2% cholesterol diet developed hypercholesterolemia and early fatty streaks at 1 month. The lesions became advanced at 3 months and were associated with de novo MIF expression by vascular endothelial cells (VECs) and smooth muscle cells (SMCs), as demonstrated by immunohistochemistry, reverse transcriptase-polymerase chain reaction, and in situ hybridization. By contrast, there was no increase in MIF levels in rabbits fed a normal diet. In early atherogenesis, marked upregulation of MIF mRNA and protein by VECs and some intimal cells were closely associated with CD68(+) monocyte adhesion onto and subsequent migration into subendothelial space. Of significance, the accumulation of macrophages was exclusively localized to areas of strong MIF expression, which may be associated with the macrophage-rich fatty streak lesion formation. Upregulation of MIF by SMCs is transient during atherogenesis. Importantly, strong MIF expression by activated macrophages may be responsible for the development of foam cell-rich lesions. Finally, the ability of MIF to induce intercellular adhesion molecule-1 expression by VECs implicates its pathogenic role in atherogenesis. In conclusion, the present study provides the first demonstration that MIF is markedly upregulated during atherogenesis. Upregulation of MIF by VECs and SMCs may play a role in macrophage adhesion, transendothelial migration, accumulation, and, importantly, transformation into foam cells. Furthermore, strong MIF expression by macrophages may both initiate and amplify the atherogenesis process. PMID- 11110780 TI - Independent signals control expression of the calcineurin inhibitory proteins MCIP1 and MCIP2 in striated muscles. AB - Calcineurin, a calcium/calmodulin-regulated protein phosphatase, modulates gene expression in cardiac and skeletal muscles during development and in remodeling responses such as cardiac hypertrophy that are evoked by environmental stresses or disease. Recently, we identified two genes encoding proteins (MCIP1 and MCIP2) that are enriched in striated muscles and that interact with calcineurin to inhibit its enzymatic activity. In the present study, we show that expression of MCIP1 is regulated by calcineurin activity in hearts of mice with cardiac hypertrophy, as well as in cultured skeletal myotubes. In contrast, expression of MCIP2 in the heart is not altered by activated calcineurin but responds to thyroid hormone, which has no effect on MCIP1. A approximately 900-bp intragenic segment located between exons 3 and 4 of the MCIP1 gene functions as an alternative promoter that responds to calcineurin. This region includes a dense cluster of 15 consensus binding sites for NF-AT transcription factors. Because MCIP proteins can inhibit calcineurin, these results suggest that MCIP1 participates in a negative feedback circuit to diminish potentially deleterious effects of unrestrained calcineurin activity in cardiac and skeletal myocytes. Inhibitory effects of MCIP2 on calcineurin activity may be pertinent to gene switching events driven by thyroid hormone in striated muscles. The full text of this article is available at http://www. circresaha.org. PMID- 11110781 TI - Activation of Rho is required for ligand-independent oncogenic signaling by a mutant epidermal growth factor receptor. AB - Mutations in the epidermal growth factor receptor have been identified in several human tumor types, including gliomas. These receptor mutants have deletions in their extracellular ligand-binding domains and are, therefore, no longer regulated by ligand, resulting in constitutive activation of the receptor kinase. These mutants have been proposed to transduce oncogenic signals via ligand independent signaling pathways. Avian viral homologues of these oncogenic epidermal growth factor receptors exhibit structurally homologous deletions and form tumors in chickens. One such mutant, S3v-ErbB, transforms fibroblasts in vitro, and transformation has been correlated with the formation of a novel tyrosine phosphoprotein complex. V-ErbB-mediated complex formation and transformation have been shown to occur independently of Ras activation. The major aims of this study are to further characterize this ligand-independent v ErbB oncogenic signaling pathway. Here we show that both v-ErbB-mediated phosphoprotein complex formation and transformation are inhibited by a dominant negative mutant of Rho. This inhibition is specific for dominant negative Rho; dominant negative mutants of Rac and Cdc42 have no effect on transformation or on tyrosine phosphorylation of the phosphoprotein complex. Based on these observations, we propose that S3v-ErbB stimulates a Rho-dependent tyrosine kinase, resulting in complex formation and ultimately oncogenic transformation. PMID- 11110782 TI - Antiproliferative effects of insulin-like growth factor-binding protein-3 in mesenchymal chondrogenic cell line RCJ3.1C5.18. relationship to differentiation stage. AB - Chondrogenesis results from a complex equilibrium between chondrocyte proliferation and differentiation. Insulin-like growth factors (IGFs) have a crucial role in chondrogenesis, but their mechanisms of action are not well defined. IGF-binding protein-3 (IGFBP-3) is the major carrier for circulating IGFs in postnatal life, and has been shown to have IGF-independent effects on proliferation of several cancer cell lines. In this study, we have evaluated the IGF-independent and -dependent effects of IGFBP-3 on chondrocyte proliferation and the relationship of these effects with chondrocyte differentiation stage. We used the RCJ3.1C5.18 nontransformed mesenchymal chondrogenic cell line, which, over 2 weeks of culture, progresses through the differentiation pathway exhibited by chondrocytes in the growth plate. We demonstrated that IGFBP-3 inhibited, in a dose-dependent manner (1-30 nm), the proliferation of chondroprogenitors and early differentiated chondrocytes, stimulated by des-(1-3)-IGF-I and longR(3)-IGF I (IGF-I analogs with reduced affinity for IGFBP-3), and by insulin and IGF-I. In terminally differentiated chondrocytes, IGFBP-3 retained the ability to inhibit cell proliferation stimulated by IGF-I, but had no effect on cell growth stimulated by insulin, or des-(1-3)-IGF-I or longR(3)IGF-I. By monolayer affinity cross-linking, we demonstrated a specific IGFBP-3-associated cell-membrane protein of approximately 20 kDa. We determined that IGFBP-3 has an antiproliferative effect on chondrocytes and, that this effect is related to the differentiation process. In chondroprogenitors and early differentiated chondrocytes, antiproliferative effect of IGFBP-3 is mainly IGF-independent, whereas, following terminal differentiation this effect is IGF-dependent. PMID- 11110783 TI - N-unsubstituted glucosamine in heparan sulfate of recycling glypican-1 from suramin-treated and nitrite-deprived endothelial cells. mapping of nitric oxide/nitrite-susceptible glucosamine residues to clustered sites near the core protein. AB - We have analyzed the content of N-unsubstituted glucosamine in heparan sulfate from glypican-1 synthesized by endothelial cells during inhibition of (a) intracellular progression by brefeldin A, (b) heparan sulfate degradation by suramin, and/or (c) endogenous nitrite formation. Glypican-1 from brefeldin A treated cells carried heparan sulfate chains that were extensively degraded by nitrous acid at pH 3.9, indicating the presence of glucosamines with free amino groups. Chains with such residues were rare in glypican-1 isolated from unperturbed cells and from cells treated with suramin and, surprisingly, when nitrite-deprived. However, when nitrite-deprived cells were simultaneously treated with suramin, such glucosamine residues were more prevalent. To locate these residues, chains were first cleaved at linkages to sulfated l-iduronic acid by heparin lyase and released fragments were separated from core protein carrying heparan sulfate stubs. These stubs were then cleaved off at sites linking N substituted glucosamines to d-glucuronic acid. These fragments were extensively degraded by nitrous acid at pH 3.9. When purified proteoglycan isolated from brefeldin A-treated cells was incubated with intact cells, endoheparanase catalyzed degradation generated a core protein with heparan sulfate stubs that were similarly sensitive to nitrous acid. We conclude that there is a concentration of N-unsubstituted glucosamines to the reducing side of the endoheparanase cleavage site in the transition region between unmodified and modified chain segments near the linkage region to the protein. Both sites as well as the heparin lyase-sensitive sites seem to be in close proximity to one another. PMID- 11110784 TI - Novel Intracellular SbV reducing activity correlates with antimony susceptibility in Leishmania donovani. AB - The standard treatment of human visceral leishmaniasis involves the use of pentavalent antimony (Sb(V)). Its mechanism of action is unknown because of the limited information available about intracellular antimony metabolism and about the genes that regulate these processes. Herein, flow injection-inductively coupled plasma mass spectrometry (ICP-MS), flow injection hydride generation ICP MS, and ion chromatography ICP-MS were used to measure antimony accumulation and intracellular metabolism in the human protozoan parasite Leishmania donovani. Amastigotes (the intracellular form) and promastigotes (the extracellular form) accumulate Sb(V) and Sb(III) via separate transport systems. Stage-specific intracellular Sb(V) reducing activity was apparent in amastigotes, which reduced the negligibly toxic Sb(V) to highly toxic Sb(III). This amastigote-specific reducing activity was deficient in the Pentostam-resistant mutant L. donovani Ld1S.20. These data indicate that parasite susceptibility to Sb(V) correlates with its level of Sb(V) reducing activity. Also, in promastigotes of both wild type L. donovani and the Pentostam-resistant mutant L. donovani Ld1S.20, Sb(V) inhibited the toxicity of Sb(III) but not of As(III). Both Sb(V) and Sb(III) were toxic to wild-type amastigotes. However, as observed in promastigotes, in mutant amastigotes Sb(V) inhibits Sb(III) but not As(III) activity. Anion exchange chromatography showed that intracellular antimony metabolism occurred in both promastigotes and amastigotes. These data demonstrate that the interaction between the two antimony oxidation states occurs intracellularly, within the parasite. The results also indicate that Sb(V) anti-leishmanial activity is dependent on its reduction to Sb(III). The mechanism of this novel intracellular Sb(V) reduction has yet to be identified, and it may or may not be enzymatic. This is the first description of intracellular Sb(V) reducing activity in Leishmania as well as in any prokaryotic or eukaryotic cell. PMID- 11110785 TI - The ligands of CXC chemokine receptor 3, I-TAC, Mig, and IP10, are natural antagonists for CCR3. AB - Th1 and Th2 lymphocytes express a different repertoire of chemokine receptors (CCRs). CXCR3, the receptor for I-TAC (interferon-inducible T cell alpha chemoattractant), Mig (monokine induced by gamma-interferon), and IP10 (interferon-inducible protein 10), is expressed preferentially on Th1 cells, whereas CCR3, the receptor for eotaxin and several other CC chemokines, is characteristic of Th2 cells. While studying responses that are mediated by these two receptors, we found that the agonists for CXCR3 act as antagonists for CCR3. I-TAC, Mig, and IP10 compete for the binding of eotaxin to CCR3-bearing cells and inhibit migration and Ca(2+) changes induced in such cells by stimulation with eotaxin, eotaxin-2, MCP-2 (monocyte chemottractant protein-2), MCP-3, MCP-4, and RANTES (regulated on activation normal T cell expressed and secreted). A hybrid chemokine generated by substituting the first eight NH(2)-terminal residues of eotaxin with those of I-TAC bound CCR3 with higher affinity than eotaxin or I-TAC (3- and 10-fold, respectively). The hybrid was 5-fold more potent than I-TAC as an inhibitor of eotaxin activity and was effective at concentrations as low as 5 nm. None of the antagonists described induced the internalization of CCR3, indicating that they lack agonistic effects and thus qualify as pure antagonists. These results suggest that chemokines that attract Th1 cells via CXCR3 can concomitantly block the migration of Th2 cells in response to CCR3 ligands, thus enhancing the polarization of T cell recruitment. PMID- 11110786 TI - Cystic fibrosis transmembrane conductance regulator facilitates ATP release by stimulating a separate ATP release channel for autocrine control of cell volume regulation. AB - These studies provide evidence that cystic fibrosis transmembrane conductance regulator (CFTR) potentiates and accelerates regulatory volume decrease (RVD) following hypotonic challenge by an autocrine mechanism involving ATP release and signaling. In wild-type CFTR-expressing cells, CFTR augments constitutive ATP release and enhances ATP release stimulated by hypotonic challenge. CFTR itself does not appear to conduct ATP. Instead, ATP is released by a separate channel, whose activity is potentiated by CFTR. Blockade of ATP release by ion channel blocking drugs, gadolinium chloride (Gd(3+)) and 4,4'-diisothiocyanatostilbene 2,2'disulfonic acid (DIDS), attenuated the effects of CFTR on acceleration and potentiation of RVD. These results support a key role for extracellular ATP and autocrine and paracrine purinergic signaling in the regulation of membrane ion permeability and suggest that CFTR potentiates ATP release by stimulating a separate ATP channel to strengthen autocrine control of cell volume regulation. PMID- 11110787 TI - Differential binding of cAMP-dependent protein kinase regulatory subunit isoforms Ialpha and IIbeta to the catalytic subunit. AB - Limited trypsin digestion of type I cAMP-dependent protein kinase holoenzyme results in a proteolytic-resistant Delta(1-72) regulatory subunit core, indicating that interaction between the regulatory and catalytic subunits extends beyond the autoinhibitory site in the R subunit at the NH(2) terminus. Sequence alignment of the two R subunit isoforms, RI and RII, reveals a significantly sequence diversity at this specific region. To determine whether this sequence diversity is functionally important for interaction with the catalytic subunit, specific mutations, R133A and D328A, are introduced into sites adjacent to the active site cleft in the catalytic subunit. While replacing Arg(133) with Ala decreases binding affinity for RII, interaction between the catalytic subunit and RI is not affected. In contrast, mutant C(D328A) showed a decrease in affinity for binding RI while maintaining similar affinities for RII as compared with the wild-type catalytic subunit. These results suggest that sequence immediately NH(2)-terminal to the consensus inhibition site in RI and RII interacts with different sites at the proximal region of the active site cleft in the catalytic subunit. These isoform-specific differences would dictate a significantly different domain organization in the type I and type II holoenzymes. PMID- 11110788 TI - Fidelity of nucleotide insertion at 8-oxo-7,8-dihydroguanine by mammalian DNA polymerase delta. Steady-state and pre-steady-state kinetic analysis. AB - Nucleotide insertion opposite 8-oxo-7,8-dihydroguanine (8-oxoG) by fetal calf thymus DNA polymerase delta (pol delta) was examined by steady-state and pre steady-state rapid quench kinetic analyses. In steady-state reactions with the accessory protein proliferating cell nuclear antigen (PCNA), pol delta preferred to incorporate dCTP opposite 8-oxoG with an efficiency of incorporation an order of magnitude lower than incorporation into unmodified DNA (mainly due to an increased K(m)). Pre-steady-state kinetic analysis of incorporation opposite 8 oxoG showed biphasic kinetics for incorporation of either dCTP or dATP, with rates similar to dCTP incorporation opposite G, large phosphorothioate effects (>100), and oligonucleotide dissociation apparently rate-limiting in the steady state. Although pol delta preferred to incorporate dCTP (14% misincorporation of dATP) the extension past the A:8-oxoG mispair predominated. The presence of PCNA was found to be a more essential factor for nucleotide incorporation opposite 8 oxoG adducts than unmodified DNA, increased pre-steady-state rates of nucleotide incorporation by >2 orders of magnitude, and was essential for nucleotide extension beyond 8-oxoG. pol delta replication fidelity at 8-oxoG depends upon contributions from K(m), K(d)(dNTP), and rates of phosphodiester bond formation, and PCNA is an important accessory protein for incorporation and extension at 8 oxoG adducts. PMID- 11110789 TI - Unwinding of a DNA triple helix by the Werner and Bloom syndrome helicases. AB - Bloom syndrome and Werner syndrome are genome instability disorders, which result from mutations in two different genes encoding helicases. Both enzymes are members of the RecQ family of helicases, have a 3' --> 5' polarity, and require a 3' single strand tail. In addition to their activity in unwinding duplex substrates, recent studies show that the two enzymes are able to unwind G2 and G4 tetraplexes, prompting speculation that failure to resolve these structures in Bloom syndrome and Werner syndrome cells may contribute to genome instability. The triple helix is another alternate DNA structure that can be formed by sequences that are widely distributed throughout the human genome. Here we show that purified Bloom and Werner helicases can unwind a DNA triple helix. The reactions are dependent on nucleoside triphosphate hydrolysis and require a free 3' tail attached to the third strand. The two enzymes unwound triplexes without requirement for a duplex extension that would form a fork at the junction of the tail and the triplex. In contrast, a duplex formed by the third strand and a complement to the triplex region was a poor substrate for both enzymes. However, the same duplex was readily unwound when a noncomplementary 5' tail was added to form a forked structure. It seems likely that structural features of the triplex mimic those of a fork and thus support efficient unwinding by the two helicases. PMID- 11110790 TI - Integrin-independent tyrosine phosphorylation of p125(fak) in human platelets stimulated by collagen. AB - Collagen fibers or a glycoprotein VI-specific collagen-related peptide (CRP-XL) stimulated tyrosine phosphorylation of the focal adhesion kinase, p125(fak) (FAK), in human platelets. An integrin alpha(2)beta(1)-specific triple-helical peptide ligand, containing the sequence GFOGER (single-letter nomenclature, O = Hyp) was without effect. Antibodies to the alpha(2) and beta(1) integrin subunits did not inhibit platelet FAK tyrosine phosphorylation caused by either collagen fibers or CRP-XL. Tyrosine phosphorylation of FAK caused by CRP-XL or thrombin, but not that caused by collagen fibers, was partially inhibited by GR144053F, an antagonist of integrin alpha(IIb)beta(3). The intracellular Ca(2+) chelator, BAPTA, and the protein kinase C inhibitor, Ro31-8220, were each highly effective inhibitors of the FAK tyrosine phosphorylation caused by collagen or CRP-XL. These data suggest that, in human platelets, 1) occupation or clustering of the integrin alpha(2)beta(1) is neither sufficient nor necessary for activation of FAK, 2) the fibrinogen receptor alpha(IIb)beta(3) is not required for activation of FAK by collagen fibers, and 3) both intracellular Ca(2+) and protein kinase C activity are essential intermediaries of FAK activation. PMID- 11110791 TI - Three novel components of the human exosome. AB - The yeast exosome is a complex of 3' --> 5' exoribonucleases. Sequence analysis identified putative human homologues for exosome components, although several were found only as expressed sequence tags. Here we report the cloning of full length cDNAs, which encode putative human homologues of the Rrp40p, Rrp41p, and Rrp46p components of the exosome. Recombinant proteins were expressed and used to raise rabbit antisera. In Western blotting experiments, these decorated HeLa cell proteins of the predicted sizes. All three human proteins were enriched in the HeLa cells nucleus and nucleolus, but were also clearly detected in the cytoplasm. Size exclusion chromatography revealed that hRrp40p, hRrp41p, and hRrp46p were present in a large complex. This cofractionated with the human homologues of other exosome components, hRrp4p and PM/Scl-100. Anti-PM/Scl positive patient sera coimmunoprecipitated hRrp40p, hRrp41p, and hRrp46p demonstrating their physical association. The immunoprecipitated complex exhibited 3' --> 5' exoribonuclease activity in vitro. hRrp41p was expressed in yeast and shown to suppress the lethality of genetic depletion of yeast Rrp41p. We conclude that hRrp40p, hRrp41p, and hRrp46p represent novel components of the human exosome complex. PMID- 11110792 TI - Control of Drosophila paramyosin/miniparamyosin gene expression. Differential regulatory mechanisms for muscle-specific transcription. AB - To define the transcriptional mechanisms contributing to stage- and tissue specific expression of muscle genes, we performed transgenic analysis of Drosophila paramyosin gene regulation. This gene has two promoters, one for paramyosin and one for miniparamyosin, which are active in partially overlapping domains. Regions between -0.9 and -1.7 kilobases upstream of each initiation site contribute to the temporal and spatial expression patterns. By comparing the Drosophila melanogaster and Drosophila virilis promoters, conserved binding sites were found for known myogenic factors, including one MEF2 site and three E boxes. In contrast with previous data, our experiments with the paramyosin promoter indicate that the MEF2 site is essential but not sufficient for proper paramyosin gene transcription. Mutations in the three E boxes, on the other hand, do not produce any effect in embryonic/larval muscles. Thus MEF2 site- and E box-binding proteins can play different roles in the regulation of different muscle-specific genes. For the miniparamyosin promoters, several conserved sequences were shown to correspond to functionally important regions. Our data further show that the two promoters work independently. Even when both promoters are active in the same muscle fiber, the transcription driven by one of the promoters is not affected by transcription driven by the other. PMID- 11110793 TI - Hrs interacts with sorting nexin 1 and regulates degradation of epidermal growth factor receptor. AB - Hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) is a mammalian homologue of yeast vacuolar protein sorting (Vps) protein Vps27p; however, the role of Hrs in lysosomal trafficking is unclear. Here, we report that Hrs interacts with sorting nexin 1 (SNX1), a recently identified mammalian homologue of yeast Vps5p that recognizes the lysosomal targeting code of epidermal growth factor receptor (EGFR) and participates in lysosomal trafficking of the receptor. Biochemical analyses demonstrate that Hrs and SNX1 are ubiquitous proteins that exist in both cytosolic and membrane-associated pools, and that the association of Hrs and SNX occurs on cellular membranes but not in the cytosol. Furthermore, endogenous SNX1 and Hrs form a approximately 550-kDa complex that excludes EGFR. Immunofluorescence and subcellular fractionation studies show that Hrs and SNX1 colocalize on early endosomes. By using deletion analysis, we have mapped the binding domains of Hrs and SNX1 that mediate their association. Overexpression of Hrs or its SNX1-binding domain inhibits ligand-induced degradation of EGFR, but does not affect either constitutive or ligand-induced receptor-mediated endocytosis. These results suggest that Hrs may regulate lysosomal trafficking through its interaction with SNX1. PMID- 11110794 TI - Competitive binding of bismuth to transferrin and albumin in aqueous solution and in blood plasma. AB - Several bismuth compounds are currently used as antiulcer drugs, but their mechanism of action is not well established. Proteins are thought to be target sites. In this work we establish that the competitive binding of Bi(3+) to the blood serum proteins albumin and transferrin, as isolated proteins and in blood plasma, can be monitored via observation of (1)H and (13)C NMR resonances of isotopically labeled [epsilon-(13)C]Met transferrin. We show that Met(132) in the I132M recombinant N-lobe transferrin mutant is a sensitive indicator of N-lobe metal binding. Bi(3+) binds to the specific Fe(3+) sites of transferrin and the observed shifts of Met resonances suggest that Bi(3+) induces similar conformational changes in the N-lobe of transferrin in aqueous solution and plasma. Bi(3+) binding to albumin is nonspecific and Cys(34) is not a major binding site, which is surprising because Bi(3+) has a high affinity for thiolate sulfur. This illustrates that the potential target sites for metals (in this case Bi(3+)) in proteins depend not only on their presence but also on their accessibility. Bi(3+) binds to transferrin in preference to albumin both in aqueous solution and in blood plasma. PMID- 11110795 TI - Nuclear localization of yeast Nfs1p is required for cell survival. AB - Saccharomyces cerevisiae Nfs1p is mainly found in the mitochondrial matrix and has been shown to participate in iron-sulfur cluster assembly. We show here that Nfs1p contains a potential nuclear localization signal, RRRPR, in its mature part. When this sequence was mutated to RRGSR, the mutant protein could not restore cell growth under chromosomal NFS1-depleted conditions. However, this mutation did not affect the function of Nfs1p in biogenesis of mitochondrial iron sulfur proteins. The growth defect of the mutant was complemented by simultaneous expression of the mature Nfs1p, which contains the intact nuclear localization signal but lacks its mitochondrial-targeting presequence. These results suggest that a fraction of Nfs1p is localized in the nucleus and is essential for cell viability. PMID- 11110797 TI - Fragile T-stem in disease-associated human mitochondrial tRNA sensitizes structure to local and distant mutations. AB - Mutations in human mitochondrial isoleucine tRNA (hs mt tRNA(Ile)) are associated with cardiomyopathy and opthalmoplegia. A recent study showed that opthalmoplegia related mutations gave rise to severe decreases in aminoacylation efficiencies and that the defective mutant tRNAs were effective inhibitors of aminoacylation of the wild-type substrate. The results suggested that the effectiveness of the mutations was due in large part to an inherently fragile mitochondrial tRNA structure. Here, we investigate mutant tRNAs associated with cardiomyopathy, and a series of rationally designed second-site substitutions introduced into both opthalmoplegia- and cardiomyopathy-related mutant tRNAs. A source of structural fragility was uncovered. An inherently unstable T-stem appears susceptible to misalignments. This susceptibility sensitizes both domains of the L-shaped tRNA structure to base substitutions that are deleterious. Thus, the fragile T-stem makes the structure of this human mitochondrial tRNA particularly vulnerable to local and distant mutations. PMID- 11110796 TI - Induction of nitric-oxide synthase and activation of NF-kappaB by interleukin-12 p40 in microglial cells. AB - Interleukin-12 (IL-12) is composed of two different subunits, p40 and p35. Expression of p40 mRNA but not that of p35 mRNA in excessive amount in the central nervous system of patients with multiple sclerosis (MS) suggests that IL 12 p40 may have a role in the pathogenesis of the disease. However, the mode of action of p40 is completely unknown. Because nitric oxide produced from the induction of nitric-oxide synthase (iNOS) also plays a vital role in the pathophysiology of MS, the present study was undertaken to explore the role of p40 in the induction of NO production and the expression of iNOS in microglia. Both IL-12 and p40(2), the p40 homodimer, dose-dependently induced the production of NO in BV-2 microglial cells. This induction of NO production was accompanied by an induction of iNOS protein and mRNA. Induction of NO production by the expression of mouse p40 cDNA but not that of the mouse p35 cDNA suggests that the p40 but not the p35 subunit of IL-12 is involved in the expression of iNOS. In addition to BV-2 glial cells, p40(2) also induced the production of NO in mouse primary microglia and peritoneal macrophages. However, both IL-12 and p40(2) were unable to induce the production of NO in mouse primary astrocytes. Because activation of NF-kappaB is important for the expression of iNOS, we investigated the effect of p40(2) on the activation of NF-kappaB. Induction of the DNA binding as well as the transcriptional activity of NF-kappaB by p40(2) and inhibition of p40(2)-induced expression of iNOS by SN50, a cell-permeable peptide carrying the nuclear localization sequence of p50 NF-kappaB, but not by SN50M, a nonfunctional peptide mutant, suggests that p40(2) induces the expression of iNOS through the activation of NF-kappaB. This study delineates a novel role of IL-12 p40 in inducing the expression of iNOS in microglial cells, which may participate in the pathogenesis of neuroinflammatory diseases. PMID- 11110798 TI - The Caenorhabditis elegans unc-32 gene encodes alternative forms of a vacuolar ATPase a subunit. AB - Eukaryotes possess multiple isoforms of the a subunit of the V(0) complex of vacuolar-type H(+)-ATPases (V-ATPases). Mutations in the V-ATPase a3 isoform have recently been shown to result in osteopetrosis, a fatal disease in humans, but no function has yet been ascribed to other isoforms. In Caenorhabditis elegans, the unc-32 mutant was originally isolated on the basis of its movement defect. We have isolated four new mutant alleles, the strongest of which is embryonic lethal. We show here that unc-32 corresponds to one of the four genes encoding a V-ATPase a subunit in the nematode, and we present their expression patterns and a molecular analysis of the gene family. unc-32 gives rise via alternative splicing to at least six transcripts. In the uncoordinated alleles, the transcript unc-32 B is affected, suggesting that it encodes an isoform that is targeted to synaptic vesicles of cholinergic neurons, where it would control neurotransmitter uptake or release. Other isoforms expressed widely during embryogenesis are mutated in the lethal alleles and would be involved in other acidic organelles. Our results indicate that V-ATPase a subunit genes are highly regulated and have tissue-specific function. PMID- 11110799 TI - On a potential global role for vitamin K-dependent gamma-carboxylation in animal systems. Evidence for a gamma-glutamyl carboxylase in Drosophila. AB - The vitamin K-dependent gamma-carboxylation of glutamate to gamma carboxyglutamate was originally well characterized in the mammalian blood clotting cascade. gamma-Carboxyglutamate has also been found in a number of other mammalian proteins and in neuropeptides from the venoms of marine snails belonging to the genus Conus, suggesting wider prevalence of gamma-carboxylation. We demonstrate that an open reading frame from a Drosophila melanogaster cDNA clone encodes a protein with vitamin K-dependent gamma-carboxylase activity. The open reading frame, 670 amino acids in length, is truncated at the C-terminal end compared with mammalian gamma-carboxylase, which is 758 amino acids. The mammalian gene has 14 introns; in Drosophila there are two much shorter introns but in positions precisely homologous to two of the mammalian introns. In addition, a deletion of 6 nucleotides is observed when cDNA and genomic sequences are compared. In situ hybridization to fixed embryos indicated ubiquitous presence of carboxylase mRNA throughout embryogenesis. Northern blot analysis revealed increased mRNA levels in 12-24-h embryos. The continued presence of carboxylase mRNA suggests that it plays an important role during embryogenesis. Although the model substrate FLEEL is carboxylated by the enzyme, a substrate containing the propeptide of a Conus carboxylase substrate, conantokin G, is poorly carboxylated. Its occurrence in vertebrates, molluscan systems (i.e. Conus), and Drosophila and the apparently strong homology between the three systems suggest that this is a highly conserved and widely distributed post translational modification in biological systems. PMID- 11110800 TI - Direct demonstration that homotetrameric chaperone SecB undergoes a dynamic dimer tetramer equilibrium. AB - We have shown here that the cytosolic bacterial chaperone SecB is a structural dimer of dimers that undergoes a dynamic equilibrium between dimer and tetramer in the native state. We demonstrated this equilibrium by mixing two tetrameric species of SecB that can be distinguished by size. We showed that the homotetrameric species exchanged dimers, because when the mixture was analyzed both by size exclusion chromatography and native polyacrylamide gel electrophoresis a third hybrid tetrameric species was detected. Furthermore, treatment of SecB with 5,5'-dithiobis-(2-nitrobenzoic acid), which modifies the sulfhydryl group on cysteines, caused irreversible dissociation to a dimer indicating that cysteine must be involved in the stabilizing interactions at the dimer interface. It is clear that the two dimer-dimer interfaces of the SecB tetramer are differentially stable. Dissociation at one interface allows for a dynamic dimer-tetramer equilibrium. Because only dimers were exchanged it is clear that the other interface between dimers is significantly more stable, otherwise oligomers should have formed with a random distribution of monomers. PMID- 11110801 TI - Identification of a phosphorylation site in the hinge region of the human progesterone receptor and additional amino-terminal phosphorylation sites. AB - We have previously reported the identification of seven in vivo phosphorylation sites in the amino-terminal region of the human progesterone receptor (PR). From our previous in vivo studies, it was evident that several phosphopeptides remained unidentified. In particular, we wished to determine whether human PR contains a phosphorylation site in the hinge region, as do other steroid receptors including chicken PR, human androgen receptor, and mouse estrogen receptor. Previously, problematic trypsin cleavage sites hampered our ability to detect phosphorylation sites in large incomplete tryptic peptides. Using a combination of mass spectrometry and in vitro phosphorylation, we have identified six previously unidentified phosphorylation sites in human PR. Using nanoelectrospray ionization mass spectrometry, we have identified two new in vivo phosphorylation sites, Ser(20) and Ser(676), in baculovirus-expressed human PR. Ser(676) is analogous to the hinge site identified in other steroid receptors. Additionally, precursor ion scans identified another phosphopeptide that contains Ser(130)-Pro(131), a likely candidate for phosphorylation. In vitro phosphorylation of PR with Cdk2 has revealed five additional in vitro Cdk2 phosphorylation sites: Ser(25), Ser(213), Thr(430), Ser(554), and Ser(676). At least two of these, Ser(213) and Ser(676), are authentic in vivo sites. We confirmed the presence of the Cdk2-phosphorylated peptide containing Ser(213) in PR from in vivo labeled T47D cells, indicating that this is an in vivo site. Our combined studies indicate that most, if not all, of the Ser-Pro motifs in human PR are sites for phosphorylation. Taken together, these data indicate that the phosphorylation of PR is highly complex, with at least 14 phosphorylation sites. PMID- 11110802 TI - Relative contribution by GABA or glycine to Cl(-)-mediated synaptic transmission on rat hypoglossal motoneurons in vitro. AB - The relative contribution by GABA and glycine to synaptic transmission of motoneurons was investigated using an hypoglossus nucleus slice preparation from neonatal rats. Spontaneous, miniature, or electrically evoked postsynaptic currents (sPSCs, mPSCs, ePSCs, respectively) mediated by glycine or GABA were recorded under whole cell voltage clamp after blocking excitatory glutamatergic transmission with kynurenic acid. The overall majority of Cl(-)-mediated sPSCs was glycinergic, while only one-third was GABAergic; 70 +/- 10% of mPSCs were glycinergic while 22 +/- 8% were GABAergic. Tetrodotoxin (TTX) application dramatically reduced the frequency (and slightly the amplitude) of GABAergic events without changing frequency or amplitude of glycinergic sPSCs. These results indicate that, unlike spontaneous GABAergic transmission, glycine mediated neurotransmission was essentially independent of network activity. There was a consistent difference in the kinetics of GABAergic and glycinergic responses as GABAergic events had significantly slower rise and decay times than glycinergic ones. Such a difference was always present whenever sPSCs, mPSCs, or ePSCs were measured. Finally, GABAergic and glycinergic mPSCs were differentially modulated by activation of glutamate metabotropic receptors (mGluRs), which are abundant in the hypoglossus nucleus. In fact, the broad-spectrum mGluR agonist (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (50 microM), which in control solution increased the frequency of both GABAergic and glycinergic sPSCs, enhanced the frequency of glycinergic mPSCs only. These results indicate that on brain stem motoneurons, Cl(-)-mediated synaptic transmission is mainly due to glycine rather than GABA and that GABAergic and glycinergic events differ in terms of kinetics and pharmacological sensitivity to mGluR activation or TTX. PMID- 11110803 TI - Task-dependent modulation of the sensorimotor transformation for smooth pursuit eye movements. AB - To investigate the transformation of retinal image velocity into smooth pursuit eye velocity, eye movements were measured in the presence of two moving targets. In the first experiment, the targets were identical in all respects except for direction of motion, and the monkey was not cued to attend to either target. In this experiment, smooth pursuit eye velocity elicited by two targets was the vector average of the response evoked by each target alone. In subsequent experiments, we examined the effects of stimulus and task parameters on the selectivity of pursuit. When the targets were made different colors and monkeys were cued for the color of the rewarded target, their pursuit eye movements were biased in the direction of the rewarded target, but still showed a substantial influence of the nonrewarded target (distractor). It did not matter whether the same target color was used for an entire session or whether the color was randomized from trial to trial. Reducing uncertainty about the axis of motion of the rewarded target also had little effect. However, the pattern of image motion appeared to have a substantial effect; radial image motion favored averaging, and winner-take-all pursuit was found only with nonradial image motion. We conclude that the sensorimotor interface for pursuit uses a flexible decision rule that can vary continuously from vector averaging to winner-take-all. We present a simple recurrent network model that reflects this range of behavior. The model has allowed us to identify three computational elements (selection bias, competitive inhibition, and response normalization) that should be taken into consideration in future models of smooth pursuit. PMID- 11110804 TI - Genetic and pharmacological demonstration of differential recruitment of cAMP dependent protein kinases by synaptic activity. AB - cAMP-dependent protein kinase (PKA) is believed to play a critical role in the expression of long-lasting forms of hippocampal long-term potentiation (LTP). Can distinct patterns of synaptic activity induce forms of LTP that require different isoforms of PKA? To address this question, we used transgenic mice that have genetically reduced hippocampal PKA activity, and a specific pharmacological inhibitor of PKA, Rp-cAMPS. Transgenic mice [R(AB) mice] that express an inhibitory form of a particular type of regulatory subunit of PKA (type-Ialpha) showed significantly reduced LTP in area CA1 of hippocampal slices as compared with slices from wild-type mice. This impairment of LTP expression was evident when LTP was induced by applying repeated, temporally spaced stimulation (4 1-s bursts of 100-Hz applied once every 5 min). In contrast, LTP induced by applying just 60 pulses in a theta-burst pattern was normal in slices from R(AB) mice as compared with slices from wild-type mice. We found that Rp-cAMPS blocked the expression of LTP induced by both spaced tetra-burst and compressed theta-burst stimulation in hippocampal slices of wild-type and R(AB) mice, respectively. Since Rp-cAMPS is a PKA inhibitor that is not selective for any particular isoform of PKA and these R(AB) mice show reduced hippocampal PKA activity resulting from genetic manipulation of a single isoform of PKA regulatory subunit, our data support the idea that distinct patterns of synaptic activity can produce different forms of LTP that significantly engage different isoforms of PKA. In particular, theta-burst LTP significantly recruits isoforms of PKA containing regulatory subunits other than the mutant RIalpha subunit, whereas tetra-burst LTP requires PKA isoforms containing the mutant RIalpha subunit. Thus, altering both the total amount of imposed synaptic activity and the temporal spacing between bursts of imposed activity may subtly modulate the PKA dependence of hippocampal LTP by engaging distinct isoforms of PKA. In a broader context, our findings suggest that synaptic plasticity in the mammalian brain might be importantly regulated by activity-dependent recruitment of different isoforms of key signal transduction molecules. PMID- 11110805 TI - Freshly isolated astrocytes from rat hippocampus show two distinct current patterns and different [K(+)](o) uptake capabilities. AB - Whether astrocytes predominantly express ohmic K(+) channels in vivo, and how expression of different K(+) channels affects [K(+)](o) homeostasis in the CNS have been long-standing questions for how astrocytes function. In the present study, we have addressed some of these questions in glial fibrillary acidic protein [GFAP(+)], freshly isolated astrocytes (FIAs) from CA1 and CA3 regions of P7-15 rat hippocampus. As isolated, these astrocytes were uncoupled allowing a higher resolution of electrophysiological study. FIAs showed two distinct ion current profiles, with neither showing a purely linear I-V relationship. One population of astrocytes had a combined expression of outward potassium currents (I(Ka), I(Kd)) and inward sodium currents (I(Na)). We term these outwardly rectifying astrocytes (ORA). Another population of astrocytes is characterized by a relatively symmetric potassium current pattern, comprising outward I(Kdr), I(Ka), and abundant inward potassium currents (I(Kin)), and a larger membrane capacitance (C(m)) and more negative resting membrane potential (RMP) than ORAs. We term these variably rectifying astrocytes (VRA). The I(Kin) in 70% of the VRAs was essentially insensitive to Cs(+), while I(Kin) in the remaining 30% of VRAs was sensitive. The I(Ka) of VRAs was most sensitive to 4-aminopyridine (4-AP), while I(Kdr) of ORAs was more sensitive to tetraethylammonium (TEA). ORAs and VRAs occurred approximately equally in FIAs isolated from the CA1 region (52% ORAs versus 48% VRAs), but ORAs were enriched in FIAs isolated from the CA3 region (71% ORAs versus 29% VRAs), suggesting an anatomical segregation of these two types of astrocytes within the hippocampus. VRAs, but not ORAs, showed robust inward currents in response to an increase in extracellular K(+) from 5 to 10 mM. As VRAs showed a similar current pattern and other passive membrane properties (e.g., RMP, R(in)) to "passive astrocytes"in situ (i.e., these showing linear I-V curves), such passive astrocytes possibly represent VRAs influenced by extensive gap-junction coupling in situ. Thus, our data suggest that, at least in CA1 and CA3 regions from P7-15 rats, there are two classes of GFAP(+) astrocytes which possess different K(+) currents. Only VRAs seem suited to uptake of extracellular K(+) via I(Kin) channels at physiological membrane potentials and increases of [K(+)](o). ORAs show abundant outward potassium currents with more depolarized RMP. Thus VRAs and ORAs may cooperate in vivo for uptake and release of K(+), respectively. PMID- 11110806 TI - Roles of high-voltage-activated calcium channel subtypes in a vertebrate spinal locomotor network. AB - Lamprey spinal cord neurons possess N-, L-, and P/Q-type high-voltage-activated (HVA) calcium channels. We have analyzed the role of the different HVA calcium channels subtypes in the overall functioning of the spinal locomotor network by monitoring the influence of their specific agonists and antagonists on synaptic transmission and on N-methyl-D-aspartate (NMDA)-elicited fictive locomotion. The N-type calcium channel blocker omega-conotoxin GVIA (omega-CgTx) depressed synaptic transmission from excitatory and inhibitory interneurons. Blocking L type and P/Q-type calcium channels with nimodipine and omega-agatoxin, respectively, did not affect synaptic transmission. Application of omega-CgTx initially decreased the frequency of the locomotor rhythm, increased the burst duration, and subsequently increased the coefficient of variation and disrupted the motor pattern. These effects were accompanied by a depression of the synaptic drive between neurons in the locomotor network. Blockade of L-type channels by nimodipine also decreased the frequency and increased the duration of the locomotor bursts. Conversely, potentiation of L-type channels increased the frequency of the locomotor activity and decreased the duration of the ventral root bursts. In contrast to blockade of N-type channels, blockade or potentiation of L-type calcium channels had no effect on the stability of the locomotor pattern. The P/Q-type calcium channel blocker omega-agatoxin IVA had little effect on the locomotor frequency or burst duration. The results indicate that rhythm generation in the spinal locomotor network of the lamprey relies on calcium influx through L-type and N-type calcium channels. PMID- 11110808 TI - Ryanodine-sensitive stores regulate the excitability of AH neurons in the myenteric plexus of guinea-pig ileum. AB - Myenteric afterhyperpolarizing (AH) neurons are primary afferent neurons within the gastrointestinal tract. Stimulation of the intestinal mucosa evokes action potentials (AP) that are followed by a slow afterhyperpolarization (AHP(slow)) in the soma. The role of intracellular Ca(2+) ([Ca(2+)](i)) and ryanodine-sensitive Ca(2+) stores in modulating the electrical activity of myenteric AH neurons was investigated by recording membrane potential and bis-fura-2 fluorescence from 34 AH neurons. Mean resting [Ca(2+)](i) was approximately 200 nM. Depolarizing current pulses that elicited APs evoked AHP(slow) and an increase in [Ca(2+)](i), with similar time courses. The amplitudes and durations of AHP(slow) and the Ca(2+) transient were proportional to the number of evoked APs, with each AP increasing [Ca(2+)](i) by approximately 50 nM. Ryanodine (10 microM) significantly reduced both the amplitude and duration (by 60%) of the evoked Ca(2+) transient and AHP(slow) over the range of APs tested (1-15). Calcium induced calcium release (CICR) was graded and proportional to the number of APs, with each AP triggering a rise in [Ca(2+)](i) of approximately 30 nM Ca(2+) via CICR. This indicates that CICR amplifies Ca(2+) influx. Similar changes in [Ca(2+)](i) and AHP(slow) were evoked by two APs in control and six APs in ryanodine. Thus, the magnitude of the change in bulk [Ca(2+)](i) and not the source of the Ca(2+) is the determinant of the magnitude of AHP(slow). Furthermore, lowering of free [Ca(2+)](i), either by reducing extracellular Ca(2+) or injecting high concentrations of Ca(2+) buffer, induced depolarization, increased excitability, and abolition of AHP(slow). In addition, activation of synaptic input to AH neurons elicited a slow excitatory postsynaptic potential (sEPSP) that was completely blocked in ryanodine. These results demonstrate the importance of [Ca(2+)](i) and CICR in sensory processing in AH neurons. Activity dependent CICR may be a mechanism to grade the output of AH neurons according to the intensity of sensory input. PMID- 11110807 TI - BK-Type K(Ca) channels in two parasympathetic cell types: differences in kinetic properties and developmental expression. AB - The intrinsic electrical properties of identified choroid and ciliary neurons of the chick ciliary ganglion were examined by patch-clamp recording methods. These neurons are derived from a common pool of mesencephalic neural crest precursor cells but innervate different target tissues and have markedly different action potential waveforms and intrinsic patterns of repetitive spike discharge. Therefore it is important to determine whether these cell types express different types of plasma membrane ionic channels, and to ascertain the developmental stages at which these cell types begin to diverge. This study has focused on large-conductance Ca(2+)-activated K(+) channels (K(Ca)), which are known to regulate spike waveform and repetitive firing in many cell types. Both ciliary ganglion cell types, identified on the basis of size and somatostatin immunoreactivity, express a robust macroscopic K(Ca) carried by a kinetically homogeneous population of large-conductance (BK-type) K(Ca) channels. However, the kinetic properties of these channels are different in the two cell types. Steady-state fluctuation analyses of macroscopic K(Ca) produced power spectra that could be fitted with a single Lorentzian curve in both cell types. However, the resulting corner frequency was significantly lower in choroid neurons than in ciliary neurons, suggesting that the underlying K(Ca) channels have a longer mean open-time in choroid neurons. Consistent with fluctuation analyses, significantly slower gating of K(Ca) channels in choroid neurons was also observed during macroscopic activation and deactivation at membrane potentials positive to -30 mV. Differences in the kinetic properties of K(Ca) channels could also be observed directly in single-channel recordings from identified embryonic day 13 choroid and ciliary neurons. The mean open-time of large-conductance K(Ca) channels was significantly greater in choroid neurons than in ciliary neurons in excised inside-out patches. The developmental expression of functional K(Ca) channels appears to be regulated differently in the two cell types. Although both cell types acquire functional K(Ca) at the same developmental stages (embryonic days 9-13), functional expression of these channels in ciliary neurons requires target-derived trophic factors. In contrast, expression of functional K(Ca) channels proceeds normally in choroid neurons developing in vitro in the absence of target-derived trophic factors. Consistent with this, extracts of ciliary neuron target tissues (striated muscle of the iris/ciliary body) contain K(Ca) stimulatory activity. However, K(Ca) stimulatory activity cannot be detected in extracts of the smooth muscle targets of choroid neurons. PMID- 11110809 TI - Processing of kinetically defined boundaries in areas V1 and V2 of the macaque monkey. AB - We recorded responses in 107 cells in the primary visual area V1 and 113 cells in the extrastriate visual area V2 while presenting a kinetically defined edge or a luminance contrast edge. Cells meeting statistical criteria for responsiveness and orientation selectivity were classified as selective for the orientation of the kinetic edge if the preferred orientation for a kinetic boundary stimulus remained essentially the same even when the directions of the two motion components defining that boundary were changed by 90 degrees. In area V2, 13 of the 113 cells met all three requirements, whereas in V1, only 4 cells met the criteria of 107 that were tested, and even these demonstrated relatively weak selectivity. Correlation analysis showed that V1 and V2 populations differed greatly (P < 1.0 x 10(-6), Student's t-test) in their selectively for specific orientations of kinetic edge stimuli. Neurons in V2 that were selective for the orientation of a kinetic boundary were further distinguished from their counterparts in V1 in displaying a strong, sharply tuned response to a luminance edge of the same orientation. We concluded that selectivity for the orientation of kinetically defined boundaries first emerges in area V2 rather than in primary visual cortex. An analysis of response onset latencies in V2 revealed that cells selective for the orientation of the motion-defined boundary responded about 40 ms more slowly, on average, to the kinetic edge stimulus than to a luminance edge. In nonselective cells, that is, those presumably responding only to the local motion in the stimulus, this difference was only about 20 ms. Response latencies for the luminance edge were indistinguishable in KE-selective and nonselective neurons. We infer that while responses to luminance edges or local motion are indigenous to V2, KE-selective responses may involve feedback entering the ventral stream at a point downstream with respect to V2. PMID- 11110810 TI - Dopamine increases excitability of pyramidal neurons in primate prefrontal cortex. AB - Dopaminergic modulation of neuronal networks in the dorsolateral prefrontal cortex (PFC) is believed to play an important role in information processing during working memory tasks in both humans and nonhuman primates. To understand the basic cellular mechanisms that underlie these actions of dopamine (DA), we have investigated the influence of DA on the cellular properties of layer 3 pyramidal cells in area 46 of the macaque monkey PFC. Intracellular voltage recordings were obtained with sharp and whole cell patch-clamp electrodes in a PFC brain-slice preparation. All of the recorded neurons in layer 3 (n = 86) exhibited regular spiking firing properties consistent with those of pyramidal neurons. We found that DA had no significant effects on resting membrane potential or input resistance of these cells. However DA, at concentrations as low as 0.5 microM, increased the excitability of PFC cells in response to depolarizing current steps injected at the soma. Enhanced excitability was associated with a hyperpolarizing shift in action potential threshold and a decreased first interspike interval. These effects required activation of D1-like but not D2-like receptors since they were inhibited by the D1 receptor antagonist SCH23390 (3 microM) but not significantly altered by the D2 antagonist sulpiride (2.5 microM). These results show, for the first time, that DA modulates the activity of layer 3 pyramidal neurons in area 46 of monkey dorsolateral PFC in vitro. Furthermore the results suggest that, by means of these effects alone, DA modulation would generally enhance the response of PFC pyramidal neurons to excitatory currents that reach the action potential initiation site. PMID- 11110811 TI - Serotonergic and peptidergic modulation of the buccal mass protractor muscle (I2) in aplysia. AB - Plasticity of Aplysia feeding has largely been measured by noting changes in radula protraction. On the basis of previous work, it has been suggested that peripheral modulation may contribute to behavioral plasticity. However, peripheral plasticity has not been demonstrated in the neuromuscular systems that participate in radula protraction. Therefore in this study we investigated whether contractions of a major radula protraction muscle (I2) are subject to modulation. We demonstrate, first, that an increase in the firing frequency of the cholinergic I2 motoneurons will increase the amplitude of the resulting muscle contraction but will not modulate its relaxation rate. We show, second, that neuronal processes on the I2 muscle are immunoreactive to myomodulin (MM), RFamide, and serotonin (5-HT), but not to small cardioactive peptide (SCP) or buccalin. The I2 motoneurons B31, B32, B61, and B62 are not immunoreactive to RFamide, 5-HT, SCP, or buccalin. However, all four cells are MM immunoreactive and are capable of synthesizing MMa. Third, we show that the bioactivity of the different modulators is somewhat different; while the MMs (i.e., MMa and MMb) and 5-HT increase I2 muscle relaxation rate, and potentiate muscle contraction amplitude, MMa, at high concentrations, depresses muscle contractions. Fourth, our data suggest that cAMP at least partially mediates effects of modulators on contraction amplitude and relaxation rate. PMID- 11110812 TI - Properties of rhythmic activity generated by the isolated spinal cord of the neonatal mouse. AB - We examined the ability of the isolated lumbosacral spinal cord of the neonatal mouse (P0-7) to generate rhythmic motor activity under several different conditions. In the absence of electrical or pharmacological stimulation, we recorded several patterns of spontaneous ventral root depolarization and discharge. Spontaneous, alternating discharge between contralateral ventral roots could occur two to three times over a 10-min interval. We also observed other patterns, including left-right synchrony and rhythmic activity restricted to one side of the cord. Trains of stimuli delivered to the lumbar/coccygeal dorsal roots or the sural nerve reliably evoked episodes of rhythmic activity. During these evoked episodes, rhythmic ventral root discharges could occur on one side of the cord or could alternate from side to side. Bath application of a combination of N-methyl-D,L-aspartate (NMA), serotonin, and dopamine produced rhythmic activity that could last for several hours. Under these conditions, the discharge recorded from the left and right L(1)-L(3) ventral roots alternated. In the L(4)-L(5) segments, the discharge had two peaks in each cycle, coincident with discharge of the ipsilateral and contralateral L(1)-L(3) roots. The L(6) ventral root discharge alternated with that recorded from the ipsilateral L(1) L(3) roots. We established that the drug-induced rhythm was locomotor-like by recording an alternating pattern of discharge between ipsilateral flexor and extensor hindlimb muscle nerves. In addition, by recording simultaneously from ventral roots and muscle nerves, we established that ankle flexor discharge was in phase with ipsilateral L(1)/L(2) ventral root discharge, while extensor discharge was in phase with ipsilateral L(6) ventral root discharge. Rhythmic patterns of ventral root discharge were preserved following mid-sagittal section of the spinal cord, demonstrating that reciprocal inhibitory connections between the left and right sides of the cord are not essential for rhythmogenesis in the neonatal mouse cord. Blocking N-methyl-D-aspartate receptors, in both the intact and the hemisected preparation, revealed that these receptors contribute to but are not essential for rhythmogenesis. In contrast, the rhythm was abolished following blockade of kainate/AMPA receptors with 6-cyano-7-nitroquinoxalene-2,3 dione. These findings demonstrate that the isolated mouse spinal cord can produce a variety of coordinated activities, including locomotor-like activity. The ability to study these behaviors under a variety of different conditions offers promise for future studies of rhythmogenic mechanisms in this preparation. PMID- 11110813 TI - Differential changes of potassium currents in CA1 pyramidal neurons after transient forebrain ischemia. AB - CA1 pyramidal neurons are highly vulnerable to transient cerebral ischemia. In vivo studies have shown that the excitability of CA1 neurons progressively decreased following reperfusion. To reveal the mechanisms underlying the postischemic excitability change, total potassium current, transient potassium current, and delayed rectifier potassium current in CA1 neurons were studied in hippocampal slices prepared before ischemia and at different time points following reperfusion. Consistent with previous in vivo studies, the excitability of CA1 neurons decreased in brain slices prepared at 14 h following transient forebrain ischemia. The amplitude of total potassium current in CA1 neurons increased approximately 30% following reperfusion. The steady-state activation curve of total potassium current progressively shifted in the hyperpolarizing direction with a transient recovery at 18 h after ischemia. For transient potassium current, the amplitude was transiently increased approximately 30% at approximately 12 h after reperfusion and returned to control levels at later time points. The steady-state activation curve also shifted approximately 20 mV in the hyperpolarizing direction, and the time constant of removal of inactivation markedly increased at 12 h after reperfusion. For delayed rectifier potassium current, the amplitude significantly increased and the steady-state activation curve shifted in the hyperpolarizing direction at 36 h after reperfusion. No significant change in inactivation kinetics was observed in the above potassium currents following reperfusion. The present study demonstrates the differential changes of potassium currents in CA1 neurons after reperfusion. The increase of transient potassium current in the early phase of reperfusion may be responsible for the decrease of excitability, while the increase of delayed rectifier potassium current in the late phase of reperfusion may be associated with the postischemic cell death. PMID- 11110814 TI - Effects of repeated cold stress on activity of hypothalamic neurons in rats during performance of operant licking task. AB - The present study investigated the effects of repeated cold stress on single neuron activity in the lateral hypothalamic area (LHA) and medial hypothalamic area (MHA) of behaving rats. The rats were trained to lick a protruding spout in response to one of several cue-tone stimuli (CTSs) to ingest water, or amino acid, NaCl or glucose solution. Following this training, the rats were raised under either stressed (repeated temperature changes between -3 and 24 degrees C) or control (24 degrees C) condition for 2 mo. During this period, neuronal activity was recorded in the LHA and MHA. For rats raised under the stressed condition, mean spontaneous firing rate of LHA neurons was significantly greater than for rats under the control condition. More LHA neurons in the stressed rats responded, with an accompanying decrease in activity (inhibitory response), to CTSs than in the control rats. During extinction learning, some LHA neurons enhanced or reversed the responses to CTSs in the stressed rats, whereas no LHA neurons showed such response changes in the control rats. In contrast to the effects of the stressed condition on LHA neuron activity, mean spontaneous firing rate of MHA neurons in the stressed rats was significantly smaller than in the control rats. Fewer MHA neurons in the stressed rats responded to CTSs and/or ingestion of sapid solutions. The preceding results suggested that repeated cold stress produces a specific pattern of changes in spontaneous activity and responses to sensory stimuli in LHA and MHA neurons; this could underlie the behavioral changes induced by repeated cold stress such as hyperphagia and hyper reactivity to sensory stimuli. PMID- 11110815 TI - Discharge pattern of renal sympathetic nerve activity in the conscious rat: spectral analysis of integrated activity. AB - We investigated the periodic characteristics of bursting discharge in renal sympathetic nerve activity (RSNA) in conscious rats. Employing a discrete fast Fourier transform algorithm, a power spectrum analysis was used to quantify periodicities present in rectified and integrated RSNA whose signal-to-noise ratio in the recordings was greater than six. In conscious rats with intact baroreceptors, RSNA was characterized by four frequency components occurring at about 0.5, 1.5, 6, and 12 Hz, which corresponded to the low-frequency fluctuation of heart rate, respiration, and frequency of heart beat, and its harmonics, respectively. After intravenous infusion of sodium nitroprusside (SNP) to elicit reflex increases in RSNA and heart rate, the power for the component at 6 Hz followed the changes in heart beat frequency and was significantly increased, while those for the three other components were attenuated or experienced no change. In sino-aortic denervated (SAD) conscious rats, all four components were abolished, and the power spectrum was well fitted by a flat or Lorentzian curve, suggesting an almost random pattern. Only a respiratory-related component, which suggested common central modulation, appeared sporadically for short periods but was absent for the most part. Therefore most of this component together with the low-frequency component was also likely due to the baroreceptor-dependent peripheral modulation. The activity was sorted in 15 subgroups on the basis of spike amplitudes in the RSNA. Each subgroup showed frequency characteristics similar to the whole nerve activity. These results suggest that all periodicity in the RSNA of conscious rats with intact baroreceptors is caused by the baroreceptor input. PMID- 11110816 TI - NMDA receptor-dependent plasticity of granule cell spiking in the dentate gyrus of normal and epileptic rats. AB - Because granule cells in the dentate gyrus provide a major synaptic input to pyramidal neurons in the CA3 region of the hippocampus, spike generation by granule cells is likely to have a significant role in hippocampal information processing. Granule cells normally fire in a single-spike mode even when inhibition is blocked and provide single-spike output to CA3 when afferent activity converging into the entorhinal cortex from neocortex, brainstem, and other limbic regions increases. The effects of enhancement of N-methyl-D aspartate (NMDA) receptor-dependent excitatory synaptic transmission and reduction in gamma-aminobutyric acid-A (GABA(A)) receptor-dependent inhibition on spike generation were examined in granule cells of the dentate gyrus. In contrast to the single-spike mode observed in normal bathing conditions, perforant path stimulation in Mg(2+)-free bathing conditions evoked graded burst discharges in granule cells which increased in duration, amplitude, and number of spikes as a function of stimulus intensity. After burst discharges were evoked during transient exposure to bathing conditions that relieve the Mg(2+) block of the NMDA receptor, there was a marked increase in the NMDA receptor-dependent component of the EPSP, but no significant increase in the non-NMDA receptor dependent component of the EPSP in normal bathing medium. Supramaximal perforant path stimulation still evoked only a single spike, but granule cell spike generation was immediately converted from a single-spike firing mode to a graded burst discharge mode when inhibition was then reduced. The induction of graded burst discharges in Mg(2+)-free conditions and the expression of burst discharges evoked in normal bathing medium with subsequent disinhibition were both blocked by DL-2-amino-4-phosphonovaleric acid (APV) and were therefore NMDA receptor dependent, in contrast to long-term potentiation (LTP) in the perforant path, which is induced by NMDA receptors and is also expressed by alpha-amino-3-hydroxy 5-methyl-4-isoxazoleproprionate (AMPA) receptors. The graded burst discharge mode was also observed in granule cells when inhibition was reduced after a single epileptic afterdischarge, which enhances the NMDA receptor-dependent component of evoked synaptic response, and in the dentate gyrus reorganized by mossy fiber sprouting in kindled and kainic acid-treated rats. NMDA receptor-dependent plasticity of granule cell spike generation, which can be distinguished from LTP and induces long-term susceptibility to epileptic burst discharge under conditions of reduced inhibition, could modify information processing in the hippocampus and promote epileptic synchronization by increasing excitatory input into CA3. PMID- 11110817 TI - Postural control in the lamprey: A study with a neuro-mechanical model. AB - The swimming lamprey normally maintains the dorsal-side-up orientation due to activity of the postural control system driven by vestibular organs. Commands for postural corrections are transmitted from the brain stem to the spinal cord mainly by the reticulospinal (RS) pathways. As shown in previous studies, RS neurons are activated by contralateral roll tilt, they exhibit a strong dynamic response, but much weaker static response. Here we test a hypothesis that decoding of these commands in the spinal cord is based on the subtraction of signals in the left and right RS pathways. In this study, we used a neuro mechanical model. An intact lamprey was mounted on a platform that restrained its postural activity but allowed lateral locomotor undulations to occur. The activity in the left and right RS pathways was recorded by implanted electrodes. These natural biological signals were then used to control an electrical motor rotating the animal around its longitudinal axis toward the stronger signal. It was found that this "hybrid" system automatically stabilized a normal orientation of the lamprey in the gravitational field. The system compensated for large postural disturbances (lateral tilt up to +/-180 degrees ) due to wide angular zones of the gravitational sensitivity of RS neurons. In the nonswimming lamprey, activity of RS neurons and their vestibular responses were considerably reduced, and the system was not able to stabilize the normal orientation. However, the balance could be restored by imposing small oscillations on the lamprey, which elicited additional activation of the vestibular organs. This finding indicates that head oscillations caused by locomotor movements may contribute to postural stabilization. In addition to postural stabilization, the neuro-mechanical model reproduced a number of postural effects characteristic of the lamprey: 1) unilateral eye illumination elicited a lateral tilt ("dorsal light response") due to a shift of the equilibrium point in the vestibular-driven postural network; 2) removal of one labyrinth resulted in a loss of postural control due to an induced left-right asymmetry in the vestibulo-reticulospinal reflexes, which 3) could be compensated for by asymmetrical visual input. The main conclusion of the present study is that natural supraspinal commands for postural corrections in the roll plane can be effectively decoded on the basis of subtraction of the effects of signals delivered by the left and right RS pathways. Possible mechanisms for this transformation are discussed. PMID- 11110818 TI - Voltage-dependent ion channels in CAD cells: A catecholaminergic neuronal line that exhibits inducible differentiation. AB - Cell lines derived from tumors engineered in the CNS offer promise as models of specific neuronal cell types. CAD cells are an unusual subclone of a murine cell line derived from tyrosine hydroxylase (TH) driven tumorigenesis, which undergoes reversible morphological differentiation on serum deprivation. Using single-cell electrophysiology we have examined the properties of ion channels expressed in CAD cells. Despite relatively low resting potentials, CAD cells can be induced to fire robust action potentials when mildly artificially hyperpolarized. Correspondingly, voltage-dependent sodium and potassium currents were elicited under voltage clamp. Sodium currents are TTX sensitive and exhibit conventional activation and inactivation properties. The potassium currents reflected two pharmacologically distinguishable populations of delayed rectifier type channels while no transient A-type channels were observed. Using barium as a charge carrier, we observed an inactivating current that was completely blocked by nimodipine and thus associated with L-type calcium channels. On differentiation, three changes in functional channel expression occurred; a 4-fold decrease in sodium current density, a 1.5-fold increase in potassium current density, and the induction of a small noninactivating barium current component. The neuronal morphology, excitability properties, and changes in channel function with differentiation make CAD cells an attractive model for study of catecholaminergic neurons. PMID- 11110819 TI - Biophysical characterization of rat caudal hypothalamic neurons: calcium channel contribution to excitability. AB - Neurons in the caudal hypothalamus (CH) are responsible for the modulation of various processes including respiratory and cardiovascular output. Previous results from this and other laboratories have demonstrated in vivo that these neurons have firing rhythms matched to the respiratory and cardiovascular cycles. The goal of the present study was to characterize the biophysical properties of neurons in the CH with particular emphasis in those properties responsible for rhythmic firing behavior. Whole cell, patch-clamped CH neurons displayed a resting membrane potential of -58.0 +/- 1.1 mV and an input resistance of 319.3 +/- 16.6 MOmega when recorded in current-clamp mode in an in vitro brain slice preparation. A large proportion of these neurons displayed postinhibitory rebound (PIR) that was dependent on the duration and magnitude of hyperpolarizing current as well as the resting membrane potential of the cell. Furthermore these neurons discharged tonically in response to a depolarizing current pulse at a depolarized resting membrane potential (more positive than -65 mV) but switched to a rapid burst of firing to the same stimulus when the resting membrane potential was lowered. The PIR observed in these neurons was calcium dependent as demonstrated by the ability to block its amplitude by perfusion of Ca(2+)-free bath solution or by application of Ni(2+) (0.3-0.5 mM) or nifedipine (10 microM). These properties suggest that low-voltage-activated (LVA) calcium current is involved in the PIR and bursting firing of these CH neurons. In addition, high-voltage activated calcium responses were detected after blockade of outward potassium current or in Ba(2+)-replacement solution. In addition, almost all of the CH neurons studied showed spike frequency adaptation that was decreased following Ca(2+) removal, indicating the involvement of Ca(2+)-dependent K(+) current (I(K,Ca)) in these cells. In conclusion, CH neurons have at least two different types of calcium currents that contribute to their excitability; the dominant current is the LVA or T-type. This LVA current appears to play a significant role in the bursting characteristics that may underlie the rhythmic firing of CH neurons. PMID- 11110820 TI - Context compensation in the vestibuloocular reflex during active head rotations. AB - The vestibuloocular reflex (VOR) needs to modulate its gain depending on target distance to prevent retinal slip during head movements. We investigated gain modulation (context compensation) for binocular gaze stabilization in human subjects during voluntary yaw and pitch head rotations. Movements of each eye were recorded, both when attempting to maintain gaze on a small visual target at straight-ahead in a darkened room and after its disappearance (remembered target). In the analysis, we relied on a binocular coordinate system yielding a version and a vergence component. We examined how frequency and target distance, approached here by using vergence angle, affected the gain and phase of the version component of the VOR and compared the results to the requirements for ideal performance. Linear regression analysis on the version gain-vergence relationship yielded a slope representing the influence of target proximity and an intercept corresponding to the response at zero vergence ("default gain"). The slope of the fitted relationship, divided by the geometrically required slope, provided a measure for the quality of version context compensation ("context gain"). In both yaw and pitch experiments, we found default version gains close to one even for the remembered target condition, indicating that the active VOR for far targets is already close to ideal without visual support. In near target experiments, the presence of visual feedback yielded near unity context gains, indicating close to optimal performance (retinal slip <0.4 degrees /s). For remembered targets, the context gain deteriorated but was still superior to performance in corresponding passive studies reported in the literature. In general, context compensation in the remembered target paradigm was better for vertical than for horizontal head rotations. The phase delay of version eye velocity relative to head velocity was small (approximately 2 degrees) for both horizontal and vertical head movements. Analysis of the vergence data from the near target experiments showed that context compensation took into account that the two eyes require slightly different VORs. In the DISCUSSION, comparison of the present default VOR gains and context gains with data from earlier passive studies has led us to propose a limited role for efference copies during self generated movements. We also discuss how our analysis can provide a framework for evaluating two different hypotheses for the generation of binocular VOR eye movements. PMID- 11110821 TI - Synaptic connections from multiple subfields contribute to granule cell hyperexcitability in hippocampal slice cultures. AB - Limbic status epilepticus and preparation of hippocampal slice cultures both produce cell loss and denervation. This commonality led us to hypothesize that morphological and physiological alterations in hippocampal slice cultures may be similar to those observed in human limbic epilepsy and animal models. To test this hypothesis, we performed electrophysiological and morphological analyses in long-term (postnatal day 11; 40-60 days in vitro) organotypic hippocampal slice cultures. Electrophysiological analyses of dentate granule cell excitability revealed that granule cells in slice cultures were hyperexcitable compared with acute slices from normal rats. In physiological buffer, spontaneous electrographic granule cell seizures were seen in 22% of cultures; in the presence of a GABA(A) receptor antagonist, seizures were documented in 75% of cultures. Hilar stimulation evoked postsynaptic potentials (PSPs) and multiple population spikes in the granule cell layer, which were eliminated by glutamate receptor antagonists, demonstrating the requirement for excitatory synaptic transmission. By contrast, under identical recording conditions, acute hippocampal slices isolated from normal rats exhibited a lack of seizures, and hilar stimulation evoked an isolated population spike without PSPs. To examine the possibility that newly formed excitatory synaptic connections to the dentate gyrus contribute to granule cell hyperexcitability in slice cultures, anatomical labeling and electrophysiological recordings following knife cuts were performed. Anatomical labeling of individual dentate granule, CA3 and CA1 pyramidal cells with neurobiotin illustrated the presence of axonal projections that may provide reciprocal excitatory synaptic connections among these regions and contribute to granule cell hyperexcitability. Knife cuts severing connections between CA1 and the dentate gyrus/CA3c region reduced but did not abolish hilar-evoked excitatory PSPs, suggesting the presence of newly formed, functional synaptic connections to the granule cells from CA1 and CA3 as well as from neurons intrinsic to the dentate gyrus. Many of the electrophysiological and morphological abnormalities reported here for long-term hippocampal slice cultures bear striking similarities to both human and in vivo models, making this in vitro model a simple, powerful system to begin to elucidate the molecular and cellular mechanisms underlying synaptic rearrangements and epileptogenesis. PMID- 11110822 TI - Functional expression of L-, N-, P/Q-, and R-type calcium channels in the human NT2-N cell line. AB - The biophysical and pharmacological properties of voltage-gated calcium channel currents in the human teratocarcinoma cell line NT2-N were studied using the whole cell patch-clamp technique. When held at -80 mV, barium currents (I(Ba)s) were evoked by voltage commands to above -35 mV that peaked at +5 mV. When holding potentials were reduced to -20 mV or 5 mM barium was substituted for 5 mM calcium, there was a reduction in peak currents and a right shift in the current voltage curve. A steady-state inactivation curve for I(Ba) was fit with a Boltzmann curve (V(1/2) = -43.3 mV; slope = -17.7 mV). Maximal current amplitude increased from 1-wk (232 pA) to 9-wk (1025 pA) postdifferentiation. Whole cell I(Ba)s were partially blocked by specific channel blockers to a similar extent in 1- to 3-wk and 7- to 9-wk postdifferentiation NT2-N cells: 10 microM nifedipine (19 vs. 25%), 10 microM conotoxin GVIA (27 vs. 25%), 10 microM conotoxin MVIIC (15 vs. 16%), and 1.75 microM SNX-482 (31 vs. 33%). Currents were completely blocked by 300 microM cadmium. In the presence of nifedipine, GVIA, and MVIIC, approximately 35% of current remained, which was reduced further by SNX-482 (7 14% of current remained), consistent with functional expression of L-, N-, and P/Q-calcium channel types and one or more R-type channel. The presence of multiple calcium currents in this human neuronal-type cell line provides a potentially useful model for study of the regulation, expression and cellular function of human derived calcium channel currents; in particular the R-type current(s). PMID- 11110823 TI - Changes in the responses of Purkinje cells in the floccular complex of monkeys after motor learning in smooth pursuit eye movements. AB - We followed simple- and complex-spike firing of Purkinje cells (PCs) in the floccular complex of the cerebellum through learned modifications of the pursuit eye movements of two monkeys. Learning was induced by double steps of target speed in which initially stationary targets move at a "learning" speed for 100 ms and then change to either a higher or lower speed in the same direction. In randomly interleaved control trials, targets moved at the learning speed in the opposite direction. When the learning direction was the ON direction for simple spike responses, learning was associated with statistically significant changes in simple-spike firing for 10 of 32 PCs. Of the 10 PCs that showed significant expressions of learning, 8 showed changes in simple-spike output in the expected direction: increased or decreased firing when eye acceleration increased or decreased through learning. There were no statistically significant changes in simple-spike responses or eye acceleration during pursuit in the control direction. When the learning direction was in the OFF direction for simple-spike responses, none of 15 PCs showed significant correlates of learning. Although changes in simple-spike firing were recorded in only a subset of PCs, analysis of the population response showed that the same relationship between population firing and eye acceleration obtained before and after learning. Thus learning is associated with changes that render the modified population response appropriate to drive the changed behavior. To analyze complex-spike firing during learning we correlated complex-spike firing in the second, third, and fourth 100 ms after the onset of target motion with the retinal image motion in the previous 100 ms. Data were largely consistent with previous evidence that image motion drives complex spikes with a direction selectivity opposite that for simple spikes. Comparison of complex-spike responses at different times after the onset of control and learning target motions in the learning direction implied that complex spikes could guide learning during decreases but not increases in eye acceleration. Learning caused increases or decreases in the sensitivity of complex spikes to image motion in parallel with changes in eye acceleration. Complex-spike responses were similar in all PCs, including many in which learning did not modify simple-spike responses. Our data do not disprove current theories of cerebellar learning but suggest that these theories would have to be modified to account for simple- and complex-spike firing of floccular Purkinje cells reported here. PMID- 11110824 TI - Regeneration of cerebral-buccal interneurons and recovery of ingestion buccal motor programs in Aplysia after CNS lesions. AB - In the sea slug Aplysia, rhythmic biting is eliminated after bilateral cerebral buccal connective (CBC) crushes and recovers within 14 days postlesion (dpl). The ability of cerebral-buccal interneuron-2 (CBI-2) to elicit ingestion buccal motor programs (iBMPs; i.e., fictive rhythmic ingestion) and to regenerate synaptic connections with target buccal neurons was assessed with intracellular recordings and dye injections. Isolated central ganglia were obtained from control animals and from lesioned animals at selected times after bilateral CBC crushes. Within 3 wk postlesion, transected CBI-2 axons sprouted at least 10 fine neurites confined to the core of the CBC that projected across the crush site toward the buccal ganglia. When fired with depolarizing current steps, CBI-2 was not observed to elicit iBMPs in preparations until 14 dpl. Thereafter a progressive enhancement in CBI-2's ability to elicit iBMPs was observed with time postlesion. By 40 dpl, CBI-2-elicited iBMPs were indistinguishable from those of controls. CBI-2 regenerated monosynaptic connections with appropriate buccal premotor- and motorneurons by 14 dpl, and the strength of these connections increased with time postlesion. Dramatic frequency facilitation was exhibited by the regenerating CBI 2 buccal synapses; for instance, at early postlesion times, no observable excitatory postsynaptic potentials (EPSPs) were obtained with 1- Hz stimulation of CBI-2, while at 7 Hz, a dramatic increase in EPSP amplitude was obtained with successive spikes. The present study shows that the time course of axonal and synaptic regeneration by command-like interneuron CBI-2 is correlated with the recovery of ingestion buccal motor programs elicited by CBI-2. These results parallel our previous findings of functional neural regeneration in the feeding system and suggest that functional neural regeneration is at least in part mediated by regeneration of specific synaptic pathways. PMID- 11110825 TI - Developmental regulation of calcium channel-mediated currents in retinal glial (Muller) cells. AB - Whole cell voltage-clamp recordings of freshly isolated cells were used to study changes in the currents through voltage-gated Ca(2+) channels during the postnatal development of immature radial glial cells into Muller cells of the rabbit retina. Using Ba(2+) or Ca(2+) ions as charge carriers, currents through transient low-voltage-activated (LVA) Ca(2+) channels were recorded in cells from early postnatal stages, with an activation threshold at -60 mV and a peak current at -25 mV. To increase the amplitude of currents through Ca(2+) channels, Na(+) ions were used as the main charge carriers, and currents were recorded in divalent cation-free bath solutions. Currents through transient LVA Ca(2+) channels were found in all radial glial cells from retinae between postnatal days 2 and 37. The currents activated at potentials positive to -80 mV and displayed a maximum at -40 mV. The amplitude of LVA currents increased during the first postnatal week; after postnatal day 6, the amplitude remained virtually constant. The density of LVA currents was highest at early postnatal days (days 2-5: 13 pA/pF) and decreased to a stable, moderate level within the first three postnatal weeks (3 pA/pF). A significant expression of currents through sustained, high voltage-activated Ca(2+) channels was found after the third postnatal week in approximately 25% of the investigated cells. The early and sole expression of transient currents at high-density may suggest that LVA Ca(2+) channels are involved in early developmental processes of rabbit Muller cells. PMID- 11110826 TI - Visual and tactile information about object-curvature control fingertip forces and grasp kinematics in human dexterous manipulation. AB - Most objects that we manipulate have curved surfaces. We have analyzed how subjects during a prototypical manipulatory task use visual and tactile sensory information for adapting fingertip actions to changes in object curvature. Subjects grasped an elongated object at one end using a precision grip and lifted it while instructed to keep it level. The principal load of the grasp was tangential torque due to the location of the center of mass of the object in relation to the horizontal grip axis joining the centers of the opposing grasp surfaces. The curvature strongly influenced the grip forces required to prevent rotational slips. Likewise the curvature influenced the rotational yield of the grasp that developed under the tangential torque load due to the viscoelastic properties of the fingertip pulps. Subjects scaled the grip forces parametrically with object curvature for grasp stability. Moreover in a curvature-dependent manner, subjects twisted the grasp around the grip axis by a radial flexion of the wrist to keep the desired object orientation despite the rotational yield. To adapt these fingertip actions to object curvature, subjects could use both vision and tactile sensibility integrated with predictive control. During combined blindfolding and digital anesthesia, however, the motor output failed to predict the consequences of the prevailing curvature. Subjects used vision to identify the curvature for efficient feedforward retrieval of grip force requirements before executing the motor commands. Digital anesthesia caused little impairment of grip force control when subjects had vision available, but the adaptation of the twist became delayed. Visual cues about the form of the grasp surface obtained before contact was used to scale the grip force, whereas the scaling of the twist depended on visual cues related to object movement. Thus subjects apparently relied on different visuomotor mechanisms for adaptation of grip force and grasp kinematics. In contrast, blindfolded subjects used tactile cues about the prevailing curvature obtained after contact with the object for feedforward adaptation of both grip force and twist. We conclude that humans use both vision and tactile sensibility for feedforward parametric adaptation of grip forces and grasp kinematics to object curvature. Normal control of the twist action, however, requires digital afferent input, and different visuomotor mechanisms support the control of the grasp twist and the grip force. This differential use of vision may have a bearing to the two-stream model of human visual processing. PMID- 11110827 TI - Groups II and III metabotropic glutamate receptors differentially modulate brief and prolonged nociception in primate STT cells. AB - The heterogeneous family of G-protein-coupled metabotropic glutamate receptors (mGluRs) provides excitatory and inhibitory controls of synaptic transmission and neuronal excitability in the nervous system. Eight mGluR subtypes have been cloned and are classified in three subgroups. Group I mGluRs can stimulate phosphoinositide hydrolysis and activate protein kinase C whereas group II (mGluR2 and 3) and group III (mGluR4, 6, 7, and 8) mGluRs share the ability to inhibit cAMP formation. The present study examined the roles of groups II and III mGluRs in the processing of brief nociceptive information and capsaicin-induced central sensitization of primate spinothalamic tract (STT) cells in vivo. In 11 anesthetized male monkeys (Macaca fascicularis), extracellular recordings were made from 21 STT cells in the lumbar dorsal horn. Responses to brief (15 s) cutaneous stimuli of innocuous (brush), marginally and distinctly noxious (press and pinch, respectively) intensity were recorded before, during, and after the infusion of group II and group III mGluR agonists into the dorsal horn by microdialysis. Different concentrations were applied for at least 20 min each (at 5 microliter/min) to obtain cumulative concentration-response relationships. Values in this paper refer to the drug concentrations in the microdialysis fibers; actual concentrations in the tissue are about three orders of magnitude lower. The agonists were also applied at 10-25 min after intradermal capsaicin injection. The group II agonists (2S,1'S,2'S)-2-(carboxycyclopropyl)glycine (LCCG1, 1 microM-10 mM, n = 6) and (-)-2-oxa-4-aminobicyclo[3.1.0]hexane-4, 6 dicarboxylate (LY379268; 1 microM-10 mM, n = 6) had no significant effects on the responses to brief cutaneous mechanical stimuli (brush, press, pinch) or on ongoing background activity. In contrast, the group III agonist L(+)-2-amino-4 phosphonobutyric acid (LAP4, 0. 1 microM-10 mM, n = 6) inhibited the responses to cutaneous mechanical stimuli in a concentration-dependent manner, having a stronger effect on brush responses than on responses to press and pinch. LAP4 did not change background discharges significantly. Intradermal injections of capsaicin increased ongoing background activity and sensitized the STT cells to cutaneous mechanical stimuli (ongoing activity > brush > press > pinch). When given as posttreatment, the group II agonists LCCG1 (100 microM, n = 5) and LY379268 (100 microM, n = 6) and the group III agonist LAP4 (100 microM, n = 6) reversed the capsaicin-induced sensitization. After washout of the agonists, the central sensitization resumed. Our data suggest that, while activation of both group II and group III mGluRs can reverse capsaicin-induced central sensitization, it is the actions of group II mGluRs in particular that undergo significant functional changes during central sensitization because they modulate responses of sensitized STT cells but have no effect under control conditions. PMID- 11110828 TI - Synaptic efficacy and the transmission of complex firing patterns between neurons. AB - In central neurons, the summation of inputs from presynaptic cells combined with the unreliability of synaptic transmission produces incessant variations of the membrane potential termed synaptic noise (SN). These fluctuations, which depend on both the unpredictable timing of afferent activities and quantal variations of postsynaptic potentials, have defied conventional analysis. We show here that, when applied to SN recorded from the Mauthner (M) cell of teleosts, a simple method of nonlinear analysis reveals previously undetected features of this signal including hidden periodic components. The phase relationship between these components is compatible with the notion that the temporal organization of events comprising this noise is deterministic rather than random and that it is generated by presynaptic interneurons behaving as coupled periodic oscillators. Furthermore a model of the presynaptic network shows how SN is shaped both by activities in incoming inputs and by the distribution of their synaptic weights expressed as mean quantal contents of the activated synapses. In confirmation we found experimentally that long-term tetanic potentiation (LTP), which selectively increases some of these synaptic weights, permits oscillating temporal patterns to be transmitted more effectively to the postsynaptic cell. Thus the probabilistic nature of transmitter release, which governs the strength of synapses, may be critical for the transfer of complex timing information within neuronal assemblies. PMID- 11110829 TI - Learning- and expectation-related changes in the human brain during motor learning. AB - We have studied a simple form of motor learning in the human brain so as to isolate activity related to motor learning and the prediction of sensory events. Whole-brain, event-related functional magnetic resonance imaging (fMRI) was used to record activity during classical discriminative delay eyeblink conditioning. Auditory conditioned stimulus (CS+) trials were presented either with a corneal airpuff unconditioned stimulus (US, paired), or without a US (unpaired). Auditory CS- trials were never reinforced with a US. Trials were presented pseudorandomly, 66 times each. The subjects gradually produced conditioned responses to CS+ trials, while increasingly differentiating between CS+ and CS- trials. The increasing difference between hemodynamic responses for unpaired CS+ and for CS- trials evolved slowly during conditioning in the ipsilateral cerebellar cortex (Crus I/Lobule HVI), contralateral motor cortex and hippocampus. To localize changes that were related to sensory prediction, we compared trials on which the expected airpuff US failed to occur (Unpaired CS+) with trials on which it occurred as expected (Paired CS+). Error-related signals in the contralateral cerebellum and somatosensory cortex were seen to increase during learning as the sensory prediction became stronger. The changes seen in the ipsilateral cerebellar cortex may be due either to the violations of sensory predictions, or to learning-related increases in the excitability of cerebellar neurons to presentations of the CS+. PMID- 11110830 TI - Comparison of odor receptive field plasticity in the rat olfactory bulb and anterior piriform cortex. AB - Recent work in the anterior piriform cortex (aPCX) has demonstrated that cortical odor receptive fields are highly dynamic, showing rapid changes of both firing rate and temporal patterning within relatively few inhalations of an odor, despite relatively maintained, patterned input from olfactory bulb mitral/tufted cells. The present experiment examined the precision (odor-specificity) of this receptive field plasticity and compared it with the primary cortical afferent, olfactory bulb mitral/tufted cells. Adult Long-Evans hooded rats, urethan anesthetized and freely breathing, were used for single-unit recording from mitral/tufted and aPCX layer II/III neurons. Partial mapping of receptive fields to alkane odors (pentane, heptane, and nonane) was performed before and immediately after habituation (50-s exposure) to one of the alkanes. The results demonstrated that odor habituation of aPCX responses was odor specific, with minimal cross-habituation between alkanes differing by as few as two carbons. Mitral/tufted cells, however, showed strong cross-habituation within the odor set with the most profound cross effects to carbon chains shorter than the habituating stimulus. The results suggest that although mitral/tufted cells and aPCX neurons have roughly similar odor receptive fields, aPCX neurons have significantly better odor discrimination within their receptive field. The results have important implications for understanding the underlying bases of receptive fields in olfactory system neurons and the mechanisms of odor discrimination and memory. PMID- 11110831 TI - Convergence of multimodal sensory input onto higher-level neurons of the crayfish olfactory pathway. AB - Intracellular electrophysiological studies of lateral protocerebral interneurons (LPIs) in the crayfish Procambarus clarkii have revealed convergence of multimodal sensory information onto these higher-level cells of the crustacean central olfactory pathway. Antennular stimulation by odors or electrical shocks generates excitatory-inhibitory sequences in some LPIs as does electrical or hydrodynamic stimulation of the antennae. Photic stimulation of the ipsilateral compound eye generates excitatory responses in LPIs, usually in the form of trains of impulse bursts that are timed to the peaks of the spontaneous oscillatory activity that characterizes these neurons. Focal electrical stimulation of the olfactory lobe, the termination point of antennular afferent input, or the accessory lobe, where higher-level visual and tactile inputs converge, also generates brief excitation and a delayed, prolonged inhibition in LPIs. Both phases of this activity are thought to be transmitted to the lateral protocerebrum via deutocerebral projection neurons, which have extensive dendritic arborizations in the olfactory lobe and the accessory lobe. The excitatory pathway is thought to synapse directly with target LPIs, whereas the inhibitory pathway is probably indirect and mediated through GABAergic interneurons within the lateral protocerebrum. There is evidence that both presynaptic and postsynaptic inhibition suppress activity in LPIs. Preliminary observations suggest that a small cluster of neurons adjacent to the hemi ellipsoid body are inhibitory to LPI activity. Multimodal inhibitory and excitatory modulation of LPI activity may play a part in the contextual identification of odors in the crayfish olfactory system. PMID- 11110832 TI - Peptide-induced Ca(2+) movements in a tonic insect muscle: effects of proctolin and periviscerokinin-2. AB - Although most of the characterized insect neuropeptides have been detected by their actions on muscle contractions, not much is known about the mechanisms underlying excitation-contraction coupling. Thus we initiated a pharmacological study on the myotropic action of the peptides periviscerokinin-2 (PVK-2) and proctolin on the hyperneural muscle of the cockroach Periplaneta americana. Both peptides required extracellular Ca(2+) to induce muscle contraction, and a blockage of sarcolemmal Ca(2+) channels by Mn(2+) or La(3+) inhibited myotropic effects. The peptides were able to induce contractions in dependence on the extracellular Ca(2+) concentration in muscles depolarized with high K(+) saline. A reduction of extracellular Na(+), K(+), or Cl(-) did not effect peptide action. Nifedipine, an L-type Ca(2+)-channel blocker, partially blocked the response to both peptides but to a much lesser extent than contractions evoked by elevated K(+). Using calcium imaging with fluo-3, we show that proctolin induces an increase of the intracellular Ca(2+) concentration. In calcium-free saline, no increase of the intracellular Ca(2+) concentration could be detected. The inhibiting effect of ryanodine, thapsigargin, and TMB-8 on peptide-induced contractions suggests that Ca(2+) release from the sarcoplasmic reticulum plays a major role during peptide-induced contractions. Preliminary experiments suggest that the peptides do not employ cyclic nucleotides as second messengers, but may activate protein kinase C. Our results indicate that the peptides induce Ca(2+) influx by an activation or modulation of dihydropyridine-sensitive and voltage independent sarcolemmal Ca(2+) channels. Ca(2+)-induced Ca(2+) release from intracellular stores, but not inositol trisphosphate-induced Ca(2+) release, seems to account for most of the observed increase in intracellular Ca(2+). Additionally, both peptides were able to potentiate glutamate-induced contractions at threshold concentrations. PMID- 11110833 TI - Context-specific adaptation of the vertical vestibuloocular reflex with regard to gravity. AB - We determined whether head position with regard to gravity is an important context for angular vestibuloocular reflex (aVOR) gain adaptation. Vertical aVOR gains were adapted with monkeys upright or on side by rotating the animals about an interaural axis in phase or out of phase with the visual surround for 4 h. When aVOR gains were adapted with monkeys upright, gain changes were symmetrical when tested in either on-side position (23 +/- 7%; mean +/- SD). After on-side adaptation, however, gain changes were always larger when animals were tested in the same on-side position in which they were adapted. Gain changes were 43 +/- 16% with ipsilateral side down and 9 +/- 8% with contralateral side down. The context-specific effects of head position on vertical aVOR gain were the same whether the gain was increased or decreased. The data indicate that vertical aVOR gain changes are stored in the context of the head orientation in which changes were induced. This association could be an important context for expressing the adapted state of the aVOR gain during vertical head movement. PMID- 11110834 TI - Tracking the hemodynamic responses to reward and punishment in the striatum. AB - Research suggests that the basal ganglia complex is a major component of the neural circuitry that mediates reward-related processing. However, human studies have not yet characterized the response of the basal ganglia to an isolated reward, as has been done in animals. We developed an event-related functional magnetic resonance imaging paradigm to identify brain areas that are activated after presentation of a reward. Subjects guessed whether the value of a card was higher or lower than the number 5, with monetary rewards as an incentive for correct guesses. They received reward, punishment, or neutral feedback on different trials. Regions in the dorsal and ventral striatum were activated by the paradigm, showing differential responses to reward and punishment. Activation was sustained following a reward feedback, but decreased below baseline following a punishment feedback. PMID- 11110835 TI - Central versus peripheral origin of vestibuloocular reflex recovery following semicircular canal plugging in rhesus monkeys. AB - We have previously shown that there is a slowly progressing, frequency-specific recovery of the gain and phase of the horizontal vestibuloocular reflex (VOR) in rhesus monkeys following plugging of the lateral semicircular canals. The adapted VOR response exhibited both dynamic and spatial characteristics that were distinctly different from responses in intact animals. To discriminate between adaptation or recovery of central versus peripheral origin, we have tested the recovered vestibuloocular responses in three rhesus monkeys in which either one or both coplanar pairs of vertical semicircular canals had been plugged previously by occluding the remaining semicircular canals in a second plugging operation. We measured the spatial tuning of the VOR in two or three different mutually orthogonal planes in response to sinusoidal oscillations (1.1 Hz, +/-5 degrees, +/-35 degrees /s) over a period of 2-3 and 12-14 mo after each operation. Apart from a significant recovery of the torsional/vertical VOR following the first operation we found that these recovered responses were preserved following the second operation, whereas the responses from the newly operated semicircular canals disappeared acutely as expected. In the follow-up period of up to 3 mo after the second operation, responses from the last operated canals showed recovery in two of three animals, whereas the previously recovered responses persisted. The results suggest that VOR recovery following plugging may depend on a regained residual sensitivity of the plugged semicircular canals to angular head acceleration. PMID- 11110836 TI - Brief and short-term corticofugal modulation of subcortical auditory responses in the big brown bat, Eptesicus fuscus. AB - Recent studies show that the auditory corticofugal system modulates and improves ongoing signal processing and reorganizes frequency map according to auditory experience in the central nucleus of bat inferior colliculus. However, whether all corticofugally affected collicular neurons are involved in both types of modulation has not been determined. In this study, we demonstrate that one group (51%) of collicular neurons participates only in corticofugal modulation of ongoing signal processing, while a second group (49%) of collicular neurons participates in both modulation of ongoing signal processing and in reorganization of the auditory system. PMID- 11110837 TI - Seizures induce simultaneous GABAergic and glutamatergic transmission in the dentate gyrus-CA3 system. AB - Monosynaptic and polysynaptic responses of CA3 pyramidal cells (PC) to stimulation of the dentate gyrus (DG) are normally blocked by glutamate receptor antagonists (GluRAs). However, after kindled seizures, GluRAs block the monosynaptic excitatory postsynaptic potential (EPSP) and isolate a monosynaptic inhibitory postsynaptic potential (IPSP), suggesting that mossy fibers release GABA. However, kindling epilepsy induces neuronal sprouting, which can underlie this fast inhibitory response. To explore this possibility, the synaptic responses of PC to DG stimulation were analyzed in kindled epileptic rats, with and without seizures, and in nonepileptic rats, immediately after a single pentylenetetrazol (PTZ)-induced seizure, in which sprouting is unlikely to have occurred. Excitatory and inhibitory synaptic responses of PC to DG stimulation were blocked by GluRAs in control cells and in cells from kindled nonseizing rats, confirming that inhibitory potentials are disynaptically mediated. However, a fast IPSP could be evoked in kindled epileptic rats and in nonepileptic rats after a single PTZ-induced seizure. The same response was induced after rekindling the epileptic nonseizing rats. This IPSP has an onset latency that parallels that of the control EPSP and is not altered under low Ca(2+) medium or halothane perfusion. In addition, it was reversibly depressed by L(+)-2-amino-4 phosphonobutyric acid (L-AP4), which is known to inhibit transmitter release from mossy fibers. These results demonstrate that seizures, and not the synaptic rearrangement due to an underlying epileptic state, induce the emergence of fast inhibition in the DG-CA3 system, and suggest that the mossy fibers underlie this plastic change. PMID- 11110838 TI - Joaquin Cravioto (1922-1998). PMID- 11110839 TI - The deltaF508 mutation in the cystic fibrosis transmembrane conductance regulator alters control of essential fatty acid utilization in epithelial cells. AB - Essential fatty acid (EFA) incorporation into phospholipid is influenced by chloride channels, suggesting that the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) may regulate aspects of EFA metabolism. The objective of this study was to determine whether the DeltaF508 mutation in the CFTR lowers 18:2(n-6) levels in phospholipid. Control cells, CF cells and CF cells transfected with the "normal" CFTR gene or the DeltaF508 CFTR gene were cultured for 3-5 d and used to determine [1-(14)C]18:2(n-6) incorporation into cell lipids. CF cells exhibited low 18:2(n-6) levels in phospholipid, reduced [1 (14)C]18:2(n-6) incorporation into phospholipid (50% of control) and greater [1 (14)C]18:2(n-6) incorporation into the triacylglycerol fraction (400% of control; P: < 0.05). Kinetic modeling of time course data for [1-(14)C]18:2(n-6) incorporation revealed a loss of metabolic control over the intracellular partitioning of 18:2(n-6) between phospholipid and triacylglycerol pools in CF cells. Expression of the normal CFTR gene in transfected CF cells increased chloride efflux and the incorporation of [1-(14)C]18:2(n-6) into phospholipid and triacylglycerol fractions. The increased incorporation of [1-(14)C]18:2(n-6) into phospholipid was attributed to significantly increased incorporation of [1 (14)C]18:2(n-6) into phosphatidylcholine and phosphatidylinositol. In CF cells expressing the defective DeltaF508 CFTR gene, conversion of [1-(14)C]18:2(n-6) to 20:4(n-6) by desaturation-chain elongation was 1.8-fold greater (P: < 0.05) than observed for CF cells transfected with the normal gene. The observations suggest that CF results in a defect in the utilization of 18:2(n-6), which is attributed in part to the defective CFTR. PMID- 11110840 TI - Ascorbate deficiency impairs the muscarinic-cholinergic and ss-adrenergic receptor signaling systems in the guinea pig submandibular salivary gland. AB - Ascorbic acid is preferentially concentrated in the hypothalamus, pituitary and adrenal glands. Its level in the acini of salivary glands is relatively high. We therefore hypothesized that ascorbate may have a role in salivary gland function. Ascorbate-deficient guinea pigs had lower stimulated whole salivary flow rates than well-fed, age-matched controls (P: < 0.005). Total salivary protein concentration was also markedly (P: < 0.005) reduced in the deficient guinea pigs. SDS-PAGE and densitometric quantification of protein bands confirmed significant reduction in specific salivary proteins (e.g., amylase, proline-rich proteins) in the saliva samples of malnourished guinea pigs. Some protein bands not seen in control saliva were detected in the saliva of malnourished guinea pigs. Ascorbate deficiency also produced a significant (P: < 0.005) reduction in the ss-adrenergic receptor density (subtype 1; 95 +/- 19 fmol/mg protein compared with 179 +/- 27 fmol/mg protein for the controls). No significant difference was observed between the two groups with respect to the ss-adrenergic receptor subtype 2. Additionally, ascorbate-deficient guinea pigs had significantly lower muscarinic-cholinergic receptor densities (50 +/- 5 vs. 74 +/- 8 fmol/mg protein for controls). Our data support the conclusion that diminished membrane receptors might impair the capacity of the transmembrane signaling system, resulting in salivary gland hypofunction in ascorbate-deficient guinea pigs. Without implying extrapolation of our findings in experimental animals to humans, it is perhaps relevant that many conditions often associated with salivary gland hypofunction in humans (e.g., smoking or drug ingestion) deplete cellular ascorbate. PMID- 11110841 TI - Maternal hypertriglyceridemia during late pregnancy does not affect the increase in circulating triglycerides caused by the long-term consumption of a sucrose rich diet by rats. AB - Feeding a sucrose-rich diet (SRD) during pregnancy enhances maternal hypertriglyceridemia. The goal of this study was to investigate whether this effect is modified when pregnancy is initiated in rats at different times during feeding of a SRD (63 g sucrose/100 g). One group of rats was fed the SRD; another group received the same diet except that the sucrose was replaced by an equal amount of cornstarch. At different times during the feeding of the diets, i.e., 5, 45 or 90 d, half of the rats were mated; after serial tail blood collections, rats were studied at d 20 of pregnancy. Virgin rats fed the same diets were always studied in parallel. Plasma triglycerides increased progressively in virgin rats fed the SRD from d 1 to 35, declined thereafter up to d 50, increased again to attain the highest level at d 65-70, partially declined at d 100 and increased again at d 110. During late pregnancy, rats fed the control diet (CD) always had greater plasma triglyceride concentrations than virgin rats, whereas triglyceride levels did not differ between pregnant and virgin rats fed the SRD. These intergroup differences were similar to those seen for plasma VLDL triglycerides. The liver triglyceride concentration in virgin rats fed the SRD was always significantly higher than that of rats fed the CD, whereas it did not differ in pregnant rats fed the SRD for either 25 or 65 d from those fed the CD. However, in those fed the SRD for 110 d, values were higher than in either pregnant or virgin rats fed the CD. We propose that the known capability of the liver to enhance triglyceride secretion during pregnancy protects dams from developing a fatty liver when fed a SRD for short periods of time, although not for long-term treatments. PMID- 11110842 TI - Long-term high protein intake does not increase oxidative stress in rats. AB - The maximum dietary protein intake that does not cause adverse effects in a healthy population is uncertain. We tested whether a high protein intake enhances oxidative stress. Adult rats were adapted to different casein-based diets containing either an adequate (13.8%; AP), medium (25.7%; MP), or high (51.3%; HP) level of crude protein; a fourth group received a HP diet but no RRR-alpha tocopherol acetate (HP-toc). After 15 wk of feeding, plasma protein carbonyl concentration, liver lipid peroxide levels [thiobarbituric acid-reacting substances (TBARS)], reduced glutathione (GSH) status and leucine kinetics ([1 (13)C]leucine) were measured. Higher concentrations of protein carbonyls and TBARS were found in rats fed the AP and the HP-toc diets compared with those fed the MP and HP diets (P: < 0.05). GSH concentrations in plasma did not differ but total blood GSH concentrations were significantly (P: < 0.05) lower in rats fed the HP-toc diet compared with those fed the AP, MP and HP diets. Liver GSH concentrations were significantly (P: < 0.01) lower in rats fed the AP diet compared with the other groups. Rates of postabsorptive leucine oxidation (LeuOX) and flux (Q(Leu)) were positively correlated with the dietary protein level (for AP, MP, and HP, respectively: LeuOX, 74.9 +/- 28.5, 109 +/- 35.2, 142.3 +/- 38.4 micromol/(kg. h); Q(Leu), 425 +/- 102, 483 +/- 82, 505 +/- 80 micromol/(kg. h). Only HP-toc resulted in a significantly greater protein breakdown (PB(Leu)) and Q(Leu). No difference was seen in nonoxidative leucine disposal. Long-term intake of high protein diets did not increase variables of oxidative stress, in contrast to our initial hypothesis. An unexpected finding was that adequate protein feeding (AP) may in fact induce oxidative stress. PMID- 11110843 TI - Bacterial dissemination and metabolic changes in rats induced by endotoxemia following intestinal E. coli overgrowth are reduced by ornithine alpha ketoglutarate administration. AB - The efficacy of ornithine alpha-ketoglutarate (OKG) in preventing bacterial translocation and dissemination, metabolic disorders and changes in mucosal enzyme activities was assessed in a model of bacterial translocation in rats. Antibiotic decontamination was performed 4 d before intragastric inoculation with an Escherichia coli strain (10(10) bacteria/kg body). Two days later, the rats were given either a lipopolysaccharide (LPS) 0127:B8 or a saline injection and were deprived of food for 24 h. Enteral nutrition, [Osmolite, 880 kJ/(kg. d)] supplemented with either OKG (LPS + OKG) or glycine (Saline + Gly or LPS + Gly), was then given for 2 d. Urinary total nitrogen losses and 3-methylhistidine excretion were determined daily. On killing at d 3, bacterial translocation to the mesenteric lymph nodes (MLN) and dissemination to the spleen and liver were evaluated, jejunal mucosa enzyme activities were assayed and tissue free amino acids in muscles were measured. Endotoxin induced translocation from the gut lumen to the MLN in all groups, whereas dissemination occurred only in LPS treated rats. OKG significantly reduced dissemination of the bacteria in the spleen. 3-Methylhistidine excretion was greater in the LPS + Gly group (+25%, P: < 0.05) than in either the LPS + OKG or Saline + Gly group. The group fed the OKG enriched diet had higher muscular glutamine, ornithine and arginine concentrations than did the Gly-supplemented groups (P: < 0.05). Intestinal sucrase and aminopeptidase activities were higher in the LPS + OKG group than in the LPS + Gly group (-30%, P: < 0.05). OKG supplementation limits bacterial dissemination and metabolic changes after injury in rats and thus may be useful in the prevention of gut-derived sepsis in critically ill patients. PMID- 11110844 TI - Dietary selenium and arsenic affect DNA methylation in vitro in Caco-2 cells and in vivo in rat liver and colon. AB - Selenium is an essential trace element for human health, and it has received considerable attention for its possible role as an anticarcinogenic agent. The purpose of the present study was to determine whether changes in the amount and the chemical form of selenium would affect DNA methylation and whether this effect would be modified by arsenic. Caco-2 cells, a human colon cancer cell line, were exposed to 0, 1 or 2 micromol supplemental selenite/L and 0, 1 or 2 micromol supplemental arsenite/L for 7 d. DNA isolated from Caco-2 cells not treated with selenite was significantly (P: < 0. 0001) hypomethylated compared with that from cells treated with 1 or 2 micromol selenite/L. DNA isolated from Caco-2 cells not treated with arsenite was significantly (P: < 0.0001) hypomethylated compared with DNA isolated from cells treated with 1 or 2 micromol arsenite/L. In addition, methylation of the p53 promoter region of Caco-2 cells decreased when cells were cultured in the absence of selenite and in the absence of arsenite. Sixty weanling male Fischer 344 rats were fed a torula yeast-based diet supplemented with 0, 0.1 or 2 mg selenium/kg diet as either selenite or selenomethionine in the presence or absence of 5 mg arsenic/kg diet as arsenite for 6 wk. Similar to the results with Caco-2 cells, rats fed selenium-deficient diets had significantly (P: < 0.0001) hypomethylated liver and colon DNA compared with rats fed 0.1 or 2.0 microg selenium/g diets as either selenite or selenomethionine. Thus, alterations in DNA methylation may be a potential mechanism, whereby deficient dietary selenium increases liver and colon tumorigenesis. PMID- 11110845 TI - Dietary restriction of single essential amino acids reduces plasma insulin-like growth factor-I (IGF-I) but does not affect plasma IGF-binding protein-1 in rats. AB - The effects of dietary restriction of a single essential amino acid (EAA) on insulin-like growth factor-I (IGF-I) and IGF-binding protein (IGFBP)-1 were investigated in rats. Rats were fed experimental diets containing amino acid (AA) mixtures in which the concentrations of all EAA were at levels recommended by the National Research Council (control), in which a single EAA was restricted to 20% of that of the control diets (Leu(-), Lys(-), Met(-) or Thr(-)), or in which the diet was devoid of amino acids (AA(-)). To eliminate the effect of differences in energy intake, rats were fed the mean amount of food as consumed by the AA(-) group on the previous day. Growth was significantly retarded in rats fed diets restricted in just one EAA compared with that of rats fed the control diet, and further growth retardation was observed in rats fed the AA(-) diet. On the other hand, the plasma IGF-I concentrations in the groups with a single EAA restriction or in the AA(-) group were 66% (P: < 0. 05) and 50% (P: < 0.05) of that of the control group, respectively. The effect of any single EAA restriction was not significantly different from that of total AA deprivation. The plasma IGFBP-1 concentration in the control group did not differ from that of rats fed diets with the single EAA restrictions except for methionine restriction, but it was approximately 6-fold greater in the AA(-) group. Differences in plasma IGFBP-1 concentration under these conditions could be explained by differences in hepatic IGFBP-1 mRNA contents. Based on these results, we conclude that restriction of single EAA does not affect IGFBP-1 synthesis in vivo, although the deprivation of a single EAA has been reported to increase IGFBP-1 production in hepatocyte cultures. Our results also indicated that a single EAA restriction decreased IGF I production but did not affect IGFBP-1 production. The present study suggests that not only plasma IGF-I, but also IGFBP-1, affects the magnitude of growth retardation in vivo. PMID- 11110846 TI - Copper deficiency induces hepatic fatty acid synthase gene transcription in rats by increasing the nuclear content of mature sterol regulatory element binding protein 1. AB - Dietary copper (Cu) deficiency results in an accelerated rate of hepatic fatty acid synthase gene transcription and an enhanced rate of hepatic lipid synthesis. Because the nuclear transcription factor sterol regulatory element binding protein-1 (SREBP-1) is a strong enhancer of fatty acid synthase promoter activity, it was hypothesized that Cu deficiency induces fatty acid synthase gene transcription by increasing the nuclear localization of mature SREBP-1. Male weanling rats were pair-fed a Cu-adequate (6.0 mg/kg) or Cu-deficient (0.6 mg/kg) diet (AIN-93) for 28 d. DNase I hypersensitivity site mapping of the hepatic fatty acid synthase gene revealed the presence of four major hypersensitivity sites located at -8700 to -8600, -7300 to -6900, -600 to -400 and -100 to +50. Although Cu deficiency did not change the hypersensitivity site pattern or intensity, in vitro footprinting of the region between -100 and +50 indicated that Cu deficiency enhanced DNA protein interactions within this region. The sequence between -68 and -58 contains the DNA recognition sequence for SREBP-1 and upstream stimulatory element-1 (USF-1). Western blot analysis revealed that the dietary Cu deficiency increased the hepatic nuclear content of mature SREBP-1 by 150% (P: < 0.05), and it concomitantly decreased the membrane content of precursor SREBP-1 by 45% (P: < 0.05). Changes in the hepatic distribution of SREBP-1 associated with Cu deficiency were not accompanied by changes in SREBP-1 mRNA. The nuclear content of USF-1 was unaffected by dietary Cu status. The hepatic increase in mature SREBP-1 of Cu-deficient rats was accompanied by a 400% increase and an 80% decrease in the abundance of fatty acid synthase and cholesterol 7-alpha hydroxylase mRNA, respectively. hepatic These data indicate that a Cu deficiency stimulates hepatic lipogenic gene expression by increasing the hepatic translocation of mature SREBP-1. PMID- 11110847 TI - Bcl-2 is not reduced in the death of MCF-7 cells at low genistein concentration. AB - Soy consumption has been associated with a lower incidence of breast cancer in Southeast Asia. Among the phytochemicals in soy, genistein has been suggested to be chemopreventive. Because genistein is an estrogen-receptor (ER) agonist, the chemopreventive mechanism has been attributed to its ability to compete with estrogen for receptor binding. In this study, we used an ER-positive cell line to investigate the effects of different genistein concentrations on the apoptotic response. The threshold concentration at which a significant number of cells underwent apoptosis was titrated to be 25 micromol/L. At or above this concentration, c-jun N-terminus kinase was activated and Bax and Bcl-2 expression were both elevated. The elevated Bcl-2 protein might neutralize the proapoptotic effect of Bax. Therefore, the mechanism of genistein-induced apoptosis at this concentration might rely largely on the stress pathway rather than the pathway mediated by the Bcl-2 family of proteins. PMID- 11110848 TI - Indole-3-carbinol is a negative regulator of estrogen receptor-alpha signaling in human tumor cells. AB - Estrogen, via its binding to the estrogen receptor (ER), plays an important role in breast cancer cell proliferation and tumor development. Indole-3-carbinol (I3C), a compound occurring naturally in cruciferous vegetables, exhibits a potent antitumor activity via its regulation of estrogen activity and metabolism. This study was designed to determine the effect of I3C on the potential to inhibit the ER-alpha. Using a reporter gene driven by the estrogen receptor, I3C (10-125 micromol/L) significantly repressed the 17ss-estradiol (E2)-activated ER alpha signaling in a dose-dependent manner. I3C and breast cancer susceptibility gene 1 (BRCA1) synergistically inhibited transcriptional activity of ER-alpha. Moreover, I3C down-regulated the expression of the estrogen-responsive genes, pS2 and cathepsin-D, and up-regulated BRCA1. The inhibitory effects of I3C did not contribute to its cytotoxic effects because these activities were observed at less than toxic concentrations. These results further suggest that antitumor activities of I3C are associated not only with its regulation of estrogen activity and metabolism, but also its modulation of ER transcription activity. PMID- 11110849 TI - Vitamin E supplementation improves cell-mediated immunity and oxidative stress of Asian men and women. AB - Vitamin E is an efficient antioxidant and a modulator of the immune system. Although racial differences in both baseline vitamin E level and immunologic subsets are known, no reliable data exist for the Asian population. Furthermore, the extent of the effect of alpha-tocopherol in protecting lymphocyte cells against oxidative stress and its association with cell-mediated immunity have not been elucidated. This study was undertaken to investigate the immunologic and antioxidant effects of vitamin E in healthy ethnic Chinese men and women. Volunteers < 35 y old (n = 26) were supplemented with 233 mg/d dl-alpha tocopherol for 28 d. The in vitro proliferative response to phytohemagglutinin (PHA) or lipopolysaccharide (LPS) of T-lymphocytes was determined in the study group before and after vitamin E supplementation. Cell-mediated immunity subsets and hydrogen peroxide production in T-lymphocytes were investigated by flow cytometry. The oxidant-antioxidant balance in plasma and urine was studied by spectrophotometric and gas chromatography-mass selective detection methods. The antioxidant properties of vitamin E were established (P: < 0.01) by the elevation of plasma vitamin E, together with depression in both plasma malondialdehyde and urinary DNA adduct 8-hydroxy-2'-deoxyguanosine after supplementation. Our data suggest a specific requirement for vitamin E in total-T and T-helper cell proliferation. We present the first evidence of the beneficial effects of supplemental vitamin E in healthy Chinese individuals on cell-mediated immunity and oxidative stress. PMID- 11110850 TI - Transgalactooligosaccharides stimulate calcium absorption in postmenopausal women. AB - The aim of this study was to investigate whether a product rich in transgalactooligosaccharides (TOS, Elix'or) stimulates true Ca absorption in postmenopausal women. The study was a double-blind, randomized crossover study, consisting of two 9-d treatment periods separated by a 19-d washout period. During the treatment periods, 12 subjects drank 200 mL yogurt drink twice (at breakfast and lunch) containing either TOS (20 g/d) or the reference substance, sucrose. On d 8 of each treatment period, (44)Ca and (48)Ca were administered orally and intravenously, respectively. Before and during the 36 h after isotope administration, urine was collected and the ratios of isotopes present were measured by inductively coupled plasma mass spectrometry (ICP-MS). From the isotope enrichments, true calcium absorption was calculated. TOS increased true calcium absorption 16%, from (mean +/- SD) 20.6 +/- 7.0% during the reference treatment to 23.9 +/- 6.9% during the TOS treatment (P: = 0.04, one-sided). In conclusion, in this study in postmenopausal women, greater Ca absorption was observed after consumption of a product rich in TOS (Elix'or) compared with the reference treatment. This increase in Ca absorption was likely due solely to TOS. The increased Ca absorption was not accompanied by increased urinary Ca excretion, meaning that TOS also may indirectly increase the uptake of Ca by bones and/or inhibit bone resorption. PMID- 11110851 TI - Conjugated linoleic acid reduces body fat mass in overweight and obese humans. AB - Conjugated linoleic acid (CLA) has been shown to reduce body fat mass (BFM) in animals. To investigate the dose-response relationships of conjugated linoleic acid with regard to BFM in humans, a randomized, double-blind study including 60 overweight or obese volunteers (body mass index 25-35 kg/m(2)) was performed. The subjects were divided into five groups receiving placebo (9 g olive oil), 1.7, 3.4, 5.1 or 6.8 g conjugated linoleic acid per day for 12 wk, respectively. Dual energy X-ray absorptiometry was used to measure body composition [measurements at wk 0 (baseline), 6 and 12]. Of the 60 subjects, 47 completed the study. Eight subjects withdrew from the study due to adverse events; however, no differences among treatment groups were found regarding adverse events. Repeated-measures analysis showed that a significantly higher reduction in BFM was found in the conjugated linoleic acid groups compared with the placebo group (P: = 0.03). The reduction of body fat within the groups was significant for the 3.4 and 6.8 g CLA groups (P: = 0.05 and P: = 0.02, respectively). No significant differences among the groups were observed in lean body mass, body mass index, blood safety variables or blood lipids. The data suggest that conjugated linoleic acid may reduce BFM in humans and that no additional effect on BFM is achieved with doses > 3.4 g CLA/d. PMID- 11110852 TI - Urinary excretion of folate catabolites responds to changes in folate intake more slowly than plasma folate and homocysteine concentrations and lymphocyte DNA methylation in postmenopausal women. AB - Folate turnover involves urinary excretion, fecal excretion, and catabolism that involves cleavage of the C9-N10 bond to yield pterins and para aminobenzoylglutamate (pABG). Little is known about the relationship between the function of folate pools and their rates of catabolism. We report here an investigation of excretion of urinary pABG and its primary excretory form, para acetamidobenzoylglutamate (ApABG) in samples collected during a previously published study of postmenopausal women. Ten women (49-63 y) were fed a low folate diet (56 microg/d) supplemented with folic acid to yield total folate intakes of 195 microg/d (d 1-5), 56 microg/d (d 6-41), 111 microg/d (d 42-69), 286 microg/d (d 70-80) and 516 microg/d (d 81-91). This caused changes in plasma folate, plasma homocysteine and global methylation of lymphocyte DNA. For each subject, a 7-d pooled urine sample was collected over d 1-7, 36-42, 64-70 and 85 91. ApABG constituted >85% of total catabolite excretion, and folate intake did not significantly influence ApABG or pABG excretion. The molar ratio of total catabolite excretion/folate intake varied significantly, with ratios of 1.0 +/- 0.17 (d 1-7), 3.0 +/- 0.55 (d 36-42), 1.1 +/- 0.18 (d 64-70) and 0. 33 +/- 0.054 (d 85-91). These observations indicate that the rate of folate catabolite excretion is related mainly to masses of slow-turnover folate pools governed by long-term folate intake. Folate pools functioning in some forms of methyl group metabolism respond to dietary changes in folate intake much more rapidly. PMID- 11110853 TI - Poor micronutrient status of active pulmonary tuberculosis patients in Indonesia. AB - Malnutrition is observed frequently in patients with pulmonary tuberculosis (TB), but their nutritional status, especially of micronutrients, is still poorly documented. The objective of this study was to investigate the nutritional status of patients with active TB compared with that of healthy controls in Jakarta, Indonesia. In a case-control study, 41 out-patients aged 15-55 y with untreated active pulmonary TB were compared with 41 healthy controls selected from neighbors of the patients and matched for age and sex. Cases had clinical and radiographic abnormalities consistent with pulmonary TB and at least two sputum specimens showing acid-fast bacilli. Anthropometric and micronutrient status data were collected. Compared with the controls, TB patients had significantly lower body mass index, skinfold thicknesses (triceps, biceps, subscapular, suprailiac), mid-upper arm circumference, proportion of fat, and concentrations of serum albumin, blood hemoglobin, plasma retinol and plasma zinc, whereas plasma zinc protoporphyrin concentration, as a measure of free erythrocyte protoporphyrin concentration, was greater. When patients and controls were subdivided on the basis of nutritional status, concentrations of serum albumin, blood hemoglobin, and zinc and retinol in plasma were lower in malnourished TB patients than in well-nourished healthy controls, well-nourished TB patients and malnourished healthy controls. In conclusion, the nutritional status of patients with active pulmonary TB was poor compared with healthy subjects, i.e., significantly more patients were anemic and more had low plasma concentrations of retinol and zinc. Low concentrations of hemoglobin, and of retinol and zinc in plasma were more pronounced in malnourished TB patients. PMID- 11110854 TI - Dietary phytate reduction improves zinc absorption in Malawian children recovering from tuberculosis but not in well children. AB - High dietary phytate content that compromises zinc nutriture is thought to be a major problem among children of the developing world. Zinc stable isotope techniques permit the quantitative assessment of the effect of phytate reduction on zinc homeostasis. We tested the hypothesis that zinc absorption would be increased in Malawian children fed a reduced-phytate corn-plus-soy diet compared with a standard high phytate diet. Twenty-three children hospitalized in Blantyre, Malawi, were enrolled. Children were selected from those recovering from tuberculosis and from well children (those with minor injuries, those awaiting elective surgery or healthy siblings). Children received a diet of corn plus-soy porridge (either low phytate or high phytate) for a period of 3-7 d and then participated in a zinc stable isotope study. The study included the administration of oral and intravenous zinc stable isotopes and 7-d collections of urine and stool. The diet was maintained throughout the duration of specimen collection. Zinc isotopic enrichments in urine and stool were measured, and zinc fractional absorption, total zinc absorption, endogenous fecal zinc, net zinc retention and size of the exchangeable zinc pool were calculated. Among the 14 children recovering from tuberculosis, dietary phytate reduction resulted in higher fractional absorption (0.41 +/- 0.14 versus 0.24 +/- 0.09, mean +/- SD, P: < 0.05) and total zinc absorption (169 +/- 55 versus 100 +/- 46 microg/(kg. d), P: < 0.05). No effect of phytate reduction was seen in the well children (n = 9). Phytate reduction did not decrease the absolute endogenous fecal zinc, but it did decrease it relative to total absorbed zinc. These preliminary results indicate that phytate reduction may be beneficial in improving zinc nutriture in groups with increased zinc requirements who consume a cereal-based diet. PMID- 11110855 TI - Overweight and underweight coexist within households in Brazil, China and Russia. AB - The possibility that underweight and overweight coexist within households and understanding such an occurrence have not been studied sufficiently. In fact, underweight and overweight are thought of as resulting from very different environmental, behavioral and individual risk factors. This study identified households in which overweight and underweight coexist and explored household level associations such as urban residence and income. Using three large national surveys from Brazil, China and Russia, the prevalence of such households ranged from 8% in China and Russia to 11% in Brazil. Even more important from the public health perspective is the finding that these under/over households accounted for a high proportion of all households with an underweight member in China (23%), Brazil (45%), and Russia (58%). The prevalence of the underweight/overweight household was highest in the urban environment in all three countries. There was no clear pattern in the prevalence of the underweight/overweight household type by income. Multivariable logistic regression was used to test the significance of the association of household type with urban residence and income while controlling for household size and household demographics by gender. Further analysis was done to consider the age relationships within the underweight/overweight pair. The underweight child coexisting with an overweight nonelderly adult was the predominant pair combination in all three countries. These findings illustrate the need for public health programs that are able to address underweight and overweight simultaneously. PMID- 11110856 TI - Maternal perception of the onset of lactation is a valid, public health indicator of lactogenesis stage II. AB - Test weighing is the "gold standard" for documenting lactogenesis stage II. However, this method is impractical for use in population studies. Maternal perception of the timing of the onset of lactation may be a useful proxy for lactogenesis stage II. This study seeks to validate maternal perception of the onset of lactation as a marker of lactogenesis stage II. Women (n = 60) were recruited after cesarean delivery. Beginning at 24 h postpartum (pp), the onset of lactation was assessed 3 times daily by both test weighing and maternal perception. Delayed onset of lactation was defined as follows: 1) milk transfer < 9.2 g/feeding at 60 h pp and 2) maternal perception >/= 72 h pp. Misclassification analyses were conducted. Multivariate logistic regression, bivariate analyses and Cox survival analyses were used to evaluate the determinants and consequences of delayed onset of lactation, using both definitions. The sensitivity and specificity of delayed maternal perception as an indicator of delayed lactogenesis were 71.4 and 79.3%, respectively. Four risk factors for low milk transfer were significant (P: < 0.05) or nearly significant (P: 0.05) absorption rate during small intestinal passage. The majority of the absorbed genistin appeared vascularly as genistein (4.4%), in addition to minor amounts of unchanged genistin (2.1%) and genistein glucuronide (1.5%). In the luminal effluent, a considerable increase of genistein (338%) as well as daidzein (190%) as cleavage products of the glucosides and malonyl-glucosides was observed. The distribution of daidzein compounds in the small intestine was not different from that of genistein compounds (P: > 0.05), except for the blood vessels, which had extremely low total amounts. Sulfate derivatives of genistein and daidzein compounds were not detectable. An effect of tofu ingredients was observed on absorption rate of genistin, on glucuronidation and on distribution of genistein glucuronide in the intestine. PMID- 11110863 TI - Polyunsaturated (n-3) fatty acids susceptible to peroxidation are increased in plasma and tissue lipids of rats fed docosahexaenoic acid-containing oils. AB - Docosahexaenoic acid [DHA, 22:6(n-3)], a major component of membrane phospholipids in brain and retina, is profoundly susceptible to oxidative stress in vitro. The extent of this peroxidation in organs when DHA is ingested in mammals, however, is not well elucidated. We investigated the effect of dietary DHA-containing oils (DHA 7.0-7.1 mol/100 mol total fatty acids), in the form of triacylglycerols (TG), ethyl esters (EE) and phospholipids (PL), on tissue lipid metabolism and lipid peroxidation in rats. Groups of Sprague-Dawley rats were fed semipurified diets containing 15 g/100 g test oils and were compared with those fed 80% palm oil and 20% soybean oil as the control (unsupplemented group) for 3 wk. The DHA oil diets markedly increased (P: < 0.05) the levels of DHA in the plasma, liver and kidney, 1.5-1.9, 2.5-3.8 and 2.2-2.5 times the control values, respectively, whereas there was a concomitant reduction (P: < 0.05) in arachidonic acid. All forms of DHA oil caused lower TG concentrations in plasma (P: < 0.05) and liver (P: < 0.05), but had no effect in kidney. The DHA oil-fed rats had greater phospholipid hydroperoxide accumulations in plasma (191-192% of control rats), liver (170-230%) and kidney (250-340%), whereas the alpha tocopherol level was reduced concomitantly (21-73% of control rats). Consistent with these results, rats fed DHA-containing oils had more thiobarbituric reactive substances in these organs than the controls. Thus, high incorporation of (n-3) fatty acids (mainly DHA) into plasma and tissue lipids due to DHA-containing oil ingestion may undesirably affect tissues by enhancing susceptibility of membranes to lipid peroxidation and by disrupting the antioxidant system. PMID- 11110864 TI - Abdominal fat deposition and fatty acid synthesis are lower and beta-oxidation is higher in broiler chickens fed diets containing unsaturated rather than saturated fat. AB - We evaluated the effects of dietary fat type on fat metabolism and deposition in broiler chickens. Birds were fed diets containing either 8 g dietary saturated (beef tallow) or polyunsaturated fat (sunflower oil)/100 g for 32 d. The abdominal fat deposition of chickens fed the sunflower oil-enriched diet was significantly lower than that of chickens fed the tallow-enriched diet (2.63 +/- 0.47 versus 3.03 +/- 0.44 g/100 g live wt.; P = 0.033). The specific activities of heart carnitine palmitoyltransferase I and L-3-hydroxyacyl-CoA dehydrogenase were higher (P < or = 0.03) in chickens fed the sunflower oil-enriched diets, indicating a greater rate of beta-oxidation. Liver fatty acid synthetase activity was lower (P = 0.01) in chickens fed the sunflower oil-enriched diet, suggesting reduced hepatic lipogenesis in this group. Postprandial plasma triglyceride levels were significantly lower (P < 0.05) in birds fed the sunflower oil enriched diet, indicating a higher rate of dietary lipid clearance from the bloodstream to tissues. In conclusion, the lower fat deposition observed in broilers fed sunflower oil-enriched diets appears to be the net result of an increased rate of lipid catabolism and lower rate of fatty acid synthesis despite higher dietary fat absorption. PMID- 11110865 TI - Vitamin B-12 deficiency and hyperhomocysteinemia are partly ameliorated by cobalt and nickel supplementation in pigs. AB - Vitamin B-12 deficiency and hyperhomocysteinemia alter the metabolism of trace elements. This study tested the hypothesis that there is a reverse relationship in which diets high in iron, copper, nickel and cobalt would influence vitamin B 12 deficiency outcomes including hyperhomocysteinemia. Piglets (German Landrace x Pietrain) were assigned to six groups of 8 and fed one of the following diets for 166 d: a vitamin B-12-adequate and folate-fortified diet (30 microg/kg vitamin B 12 and 0.5 mg/kg folate) with normal trace element concentrations or one of five vitamin B-12-free, folate nonsupplemented diets (0.36 mg/kg), with either normal trace element concentrations or high concentrations of iron (300 mg/kg), copper (30 mg/kg), cobalt (1 mg/kg) or nickel (6 mg/kg). Feed intake and weight gain did not differ significantly among the groups. Vitamin B-12-deficient pigs developed diminished serum and liver concentrations of vitamin B-12 and folate, an accumulation of iron in the liver and hyperhomocysteinemia. The magnitude of changes differed among vitamin B-12-deficient groups. Vitamin B-12-deficient pigs fed 6 mg/kg nickel had distinctly higher vitamin B-12 concentrations in liver and serum and 45% lower serum concentration of homocysteine than the corresponding deficiency group fed 1 mg/kg nickel; iron concentration in liver was completely normalized. Vitamin B-12-deficient pigs fed 1 mg/kg cobalt had 47% lower homocysteine concentrations in serum than the vitamin B-12-deficient group fed 0.13 mg/kg cobalt, but the vitamin B-12 status was unaffected. Supplementation of iron and copper did not affect these variables. The dietary manipulations had no detrimental effects on variables symptomatic of oxidative stress. The findings indicate a collaborative relationship between vitamin B-12 metabolism and the trace elements nickel and cobalt. PMID- 11110866 TI - In vivo rates of skeletal muscle protein synthesis in rats are decreased by acute ethanol treatment but are not ameliorated by supplemental alpha-tocopherol. AB - Some studies have shown that reductions in tissue protein synthesis, under a variety of cytotoxic conditions, are ameliorated by alpha-tocopherol (ATC) supplementation. We have also shown evidence of increased oxidative stress and reduced protein synthesis rates in alcohol-exposed muscle. Serum levels of ATC fall and rates of muscle protein synthesis are reduced in patients with alcoholic myopathy. We therefore tested the hypothesis that treatment with ATC could ameliorate the ethanol-induced changes in muscle protein synthesis, a contributory event in the pathogenesis of alcoholic muscle disease. Studies were carried out on gastrocnemius (Type II fiber-predominant and usually considered representative of the musculature as a whole), soleus (Type I fiber-predominant) and plantaris (Type II fiber-predominant) muscles. For comparative purposes, we also investigated the liver. Young male Wistar rats (90 g body weight) were injected intraperitoneally (i.p.) daily with ATC (30 mg/kg body weight) in Intralipid fat emulsion (0.1 mL/100 g body, i.p.) for 5 d. Controls were similarly injected with the Intralipid vehicle alone. After ATC supplementation, rats were given ethanol (75 mmol/kg body weight, i.p., 2.5 h) or saline (0.15 mol/L NaCl, i. p.). Fractional rates of tissue protein synthesis (i.e., the percentage of the tissue protein pool renewed each day, k(s), %/d) and RNA activities [i.e., the amount of protein synthesis each day per unit RNA, k(RNA), mg protein/d/mg RNA)] were then measured. Supplementation increased ATC concentrations in plasma, gastrocnemius and liver. There was no effect of ATC supplementation alone on k(s) in any of the tissues. ATC supplementation in the absence of alcohol increased k(RNA) in the plantaris muscle. In nonsupplemented groups, acute ethanol treatment reduced skeletal muscle (soleus, plantaris and gastrocnemius) k(s). Hepatic k(s) was not altered by ethanol, although ATC concentrations in this tissue increased due to ethanol. However, none of the changes in muscle k(s) or k(RNA) due to ethanol were significantly affected by ATC supplementation. In conclusion, ATC supplementation does not appear beneficial in ameliorating acute alcohol toxicity in skeletal muscle as defined by reductions in protein synthesis. PMID- 11110867 TI - Secretion of alpha-tocopherol in VLDL is decreased by dietary protein insufficiency in young growing rats. AB - The concentrations of alpha-tocopherol in plasma and most peripheral tissues were shown previously to be low in young growing rats fed a low protein diet. To examine the secretion rates of VLDL alpha-tocopherol and triglycerides, and lipoprotein lipase activity, weanling rats were fed a low protein (LP, 8 g/100 g lactalbumin) or a normal protein (NP, 20 g/100 g lactalbumin) diet for 6 wk. The absolute secretion rate of VLDL triglyceride (micromol/h) of the LP group was not significantly different from that of the NP group (P: > 0.05), but was significantly higher (P: < 0.05) when expressed relative to body weight [micromol/(h. kg)]. The secretion rates of VLDL alpha-tocopherol were significantly lower (P: < 0.05) in the LP group than in the NP group. The activities of hepatic lipase, lipoprotein lipase and total heparin-releasable lipase in plasma of the LP group were only 50-60% those of the NP group (P: < 0.05). The results demonstrated that the secretion rate of VLDL alpha-tocopherol and activities of lipases in postheparin plasma were significantly lower in rats fed a low protein diet. Thus, the redistribution of alpha-tocopherol from liver to peripheral tissues appears to have been impaired by dietary protein insufficiency. PMID- 11110868 TI - Excess dietary methionine markedly increases the vitamin B-6 requirement of young chicks. AB - A soy-protein isolate diet that contained essentially no bioavailable vitamin B-6 was used to establish the quantitative effect of excess dietary methionine on the vitamin B-6 requirement of young chicks. When made adequate in vitamin B-6, chicks fed the basal diet required 2 g/kg supplemental DL-methionine to achieve maximal growth, and 10 g/kg additional DL-methionine (total = 12 g/kg) was found to be a tolerable excess level that would not depress voluntary food intake or growth rate. When chicks were fed seven graded doses of supplemental pyridoxine (PN) in diets that contained either adequate (2 g/kg) or excess (12 g/kg) methionine, the vitamin B-6 requirement for maximal growth was found to increase (P: < 0.01) from 0.73 to 1.05 mg/kg, a 44% increase, when 10 g/kg excess methionine was present in the diet. Indeed, this level of excess dietary methionine depressed (P: < 0.01) growth at all PN dose levels < or =1 mg/kg, but not at PN doses of 1.2 or 1.4 mg/kg. Because dietary intakes of both vitamin B-6 and methionine can affect plasma homocysteine levels, dietary methionine (and protein) intake should be considered important factors in setting safe and adequate requirement levels for vitamin B-6. PMID- 11110869 TI - Lysosomal enzyme activities are decreased in the retina and their circadian rhythms are different from those in the pineal gland of rats fed an alpha linolenic acid-restricted diet. AB - The retinal rod outer segment (ROS) is shed and digested daily by phagosomes in retinal pigment epithelial (RPE) cells. We previously observed significantly fewer large phagosomes in rats fed an alpha-linolenic acid (ALNA)-deficient diet. Rats fed a safflower oil diet (ALNA-restricted) or a perilla oil diet (ALNA sufficient) through two generations were adapted to a 24-h cycle with light from 0700 to 1900 h. They were killed at 0500, 0900, 1300 and 1700 h to determine the activities of four lysosomal enzymes in retina, including beta-glucosidase, beta glucuronidase, hexosaminidase and acid phosphatase. The enzyme activities at 0500 h were the lowest and then increased gradually until 1700 h, exhibiting similar circadian rhythms in the two dietary groups. However, the activities at each time point were significantly lower in the safflower group. In the pineal gland, the activities were maximum at 1300 h, except for beta-glucosidase, and were not different between groups. These diets had qualitatively similar but quantitatively different effects on the fatty acid compositions of the retina and the pineal gland. These results indicate that decreased amplitudes in electroretinogram and altered size distribution of phagosomes, as induced by a restricted intake of ALNA, are associated with decreased lysosomal enzyme activities in the retina but not in the pineal gland. PMID- 11110870 TI - More Americans are eating "5 a day" but intakes of dark green and cruciferous vegetables remain low. AB - Epidemiological investigations repeatedly show that the regular consumption of dark green and cruciferous vegetables, tomatoes and citrus fruits in particular is related to reduced cancer risk. We used the 1994-1996 Continuing Survey of Food Intakes by Individuals to examine the types of fruits and vegetables consumed by Americans. The analytic sample population, which consisted of 4806 men and women (25-75 y old) who completed two nonconsecutive 24-h recalls, consumed 3.6 +/- 2.3 servings of vegetables and 1.6 +/- 2.0 servings of fruit daily. Iceberg lettuce, tomatoes, French fried potatoes, bananas and orange juice were the most commonly consumed fruits and vegetables, accounting for nearly 30% of all fruits and vegetables consumed. The most popular items, lettuce and tomatoes, were consumed by 39-42% of the sample population during the reporting period. Fewer respondents (16-24%) consumed French fried potatoes, bananas or orange juice. Only 3% of the sample consumed broccoli during the reporting period. White potato consumption averaged 1.1 servings daily, with French fried potatoes representing 0.4 serving. Tomato product consumption averaged 0.5 serving daily, dark green vegetable consumption averaged 0.2 serving daily and citrus, berries or melon consumption amounted to nearly 0.8 serving daily. These data indicate that Americans are consuming more fruits and vegetables but that dark green and cruciferous vegetable intake is low. PMID- 11110871 TI - Dietary patterns are associated with body mass index in multiethnic women. AB - This cross-sectional study investigated the relationship between dietary patterns and body mass index among 514 women with different ethnic backgrounds who completed a validated food-frequency questionnaire. An exploratory factor analysis with orthogonal rotation started with 23 food items and resulted in four factors that accounted for 93% of the total variance. Confirmatory factor analysis with the 16 items that had factor loadings of at least 0.60 validated the four dietary patterns. The most significant dietary pattern, "meat," was characterized by high intake of processed and red meats, fish, poultry, eggs, fats and oils, and condiments. The "vegetable" pattern loaded high on different vegetables, whereas the third pattern named "bean" was high in legumes, tofu and soy protein. The major components of the "cold foods" pattern were fruit, fruit juice and cold breakfast cereals. Although the "meat" pattern was predominant among Hawaiians and the "bean" pattern very common among Chinese and Japanese women, factors two and four were not related to ethnicity. After adjustment for daily energy intake, the "meat" pattern was positively associated with body mass index (r = 0.17, P: = 0.0001), whereas the other three patterns showed negative relationships to body mass index (r = -0.076, P: = 0.084, r = -0.13, P: = 0.003, and r = -0.13, P: = 0.003) for vegetables, beans and cold foods, respectively. The associations were similar in direction and magnitude for all ethnic groups. The study results support the ideas that choosing the right foods may be important in weight control and that food-based dietary patterns may be useful in dietary counseling. PMID- 11110872 TI - Serum total homocysteine concentration is related to self-reported heart attack or stroke history among men and women in the NHANES III. AB - High circulating total homocysteine (tHcy) concentration, which is influenced by folate and vitamin B-12 status, is a suspected cause of cardiovascular events. This relation has been investigated in both case-control and prospective studies but has not been evaluated for different sex x age subgroups of the general U.S. population. We used data on adult (i.e., aged > or =40 y) male (n = 1097) and female (n = 1107) participants in the third National Health and Nutrition Examination Survey, excluding diabetics and those supplemented with estrogen, vitamins or minerals, to evaluate the association between serum tHcy concentration and self-report of heart attack or stroke. After adjustment for age, race-ethnicity, smoking, blood pressure, blood pressure medication, body mass index and serum concentrations of creatinine and cholesterol, past events were reported 2.4 (95% confidence interval 1.0-5.5) times as often by men with tHcy concentration of >12 micromol/L as by men with lower values. The odds ratio for women was 2.6 (95% confidence interval 1.1-6.6) after adjustment for the same factors plus menopausal status. A stronger relation in men aged < or =60 y compared with older men may help reconcile conflicting results of earlier studies. PMID- 11110873 TI - Description of the long-term lipogenic effects of dietary carbohydrates in male Fischer 344 rats. AB - The introduction of high fructose corn syrup as a substitute sweetener for sucrose in the mid-1970s has contributed to a general increase in fructose consumption in the U.S. diet. Although several previous investigations suggested that dietary fructose increases serum triglyceride concentration and body fat, these studies have, in general, evaluated this effect in young rats fed the experimental diets for a relatively short period of the life span of the animals. Moreover, these investigations did not control for the possible effects that increased adiposity due to fructose feeding may have on serum triglyceride concentration. The purpose of the current investigation was to describe the long term effects of specific dietary carbohydrates on serum lipid concentrations and body composition. To this end, we measured serum triglyceride, total cholesterol and HDL cholesterol concentrations and body composition of rats aged 9, 18 and 26 mo that had free access to or were restricted to 60% of free access intake of one of five diets that varied in carbohydrate source (cornstarch, sucrose, glucose, fructose or equimolar fructose plus glucose) starting at 3 mo of age. Dietary fructose significantly increased serum triglyceride concentration across the life span in rats that had free access to food or were calorie restricted. The source of dietary carbohydrate did not have a significant effect on body composition, total cholesterol or the distribution of the cholesterol fractions. These data suggest that dietary fructose per se and not the interaction between fructose and the energy content of the diet increases serum triglyceride concentration in rats. PMID- 11110874 TI - A nucleoside-nucleotide mixture may reduce memory deterioration in old senescence accelerated mice. AB - We investigated the effects of a mixture of dietary nucleosides and nucleotides (NS + NT) on memory in 1- and 7-mo-old senescence-accelerated mice (SAM). Memory retention was studied with passive avoidance (step-through) and active avoidance (shuttle) tests. For 14 wk, mice in the control groups were fed a 20 g of casein/100 g diet, whereas the NS + NT groups were fed this diet supplemented with a 0.5 g of NS + NT mixture/100 g. All mice were killed at wk 14, and we studied the brain histopathology. Lipofuscin, monovacuoles and multiple vacuoles of various brain regions were measured. Body weight, food intake and ambulatory activity did not differ between the control and NS + NT groups. In old mice, the time of passive avoidance was significantly higher in the NS + NT group than in the control group at d 1 and 7 (P: < 0.05). However, such an effect of NS + NT was not observed in young mice. In the active avoidance test, the incidence of successful avoidance in old mice was higher in the NS + NT group than in the control group at d 1 and 2 (P: < 0.05). The percentages of specific brain cells containing lipofuscin were lower in NS + NT groups than in the control groups in both young and old mice (P: < 0.05). The number of monovacuoles and multiple vacuoles in specific brain regions tended to be lower (P: = 0.1-0.25) in NS + NT than in control groups, with significant differences in the microvacuoles of the middle cortex of young mice and in the multiple vacuoles in the hind cortex of old mice (P: < 0. 05). These results suggest that increased dietary NS + NT may be associated with decreases in the age-induced deterioration of brain morphology and certain memory tasks. PMID- 11110875 TI - Multivitamin/mineral supplementation improves plasma B-vitamin status and homocysteine concentration in healthy older adults consuming a folate-fortified diet. AB - Elevated homocysteine has been identified as an independent risk factor for cardiovascular and cerebrovascular disease. Although multivitamin use has been associated with low plasma homocysteine concentrations in several observational studies, no clinical trials have been conducted using multivitamin/mineral supplements to lower homocysteine. We determined whether a multivitamin/mineral supplement formulated at about 100% Daily Value will further lower homocysteine concentration and improve B-vitamin status in healthy older adults already consuming a diet fortified with folic acid. In this randomized, double-blind, placebo-controlled trial, 80 free-living men and women aged 50-87 y with total plasma homocysteine concentrations of > or =8 micromol/L received either a multivitamin/mineral supplement or placebo for 56 d while consuming their usual diet. After the 8-wk treatment, subjects taking the supplement had significantly higher B-vitamin status and lower homocysteine concentration than controls (P: < 0.01). Plasma folate, pyridoxal phosphate (PLP) and vitamin B-12 concentrations were increased 41.6, 36.5 and 13.8%, respectively, in the supplemented group, whereas no changes were observed in the placebo group. The mean homocysteine concentration decreased 9.6% in the supplemented group (P: < 0.001) and was unaffected in the placebo group. There were no significant changes in dietary intake during the intervention. Multivitamin/mineral supplementation can improve B-vitamin status and reduce plasma homocysteine concentration in older adults already consuming a folate-fortified diet. PMID- 11110876 TI - A rodent model of protein turnover used to design an experiment for measuring the rates of channeling, recycling and protein synthesis. AB - We described previously a mechanistic model of whole-body protein turnover in rodents. Channeling was defined as the flow of amino acids from the extracellular compartment to aminoacyl tRNA and protein synthesis. Recycling was defined as the flow of amino acids from protein degradation to aminoacyl tRNA (protein synthesis) without mixing with the intracellular pool of amino acids. In this paper, the model is applied to tissues and whole body and is used to develop an experimental protocol for estimating protein fractional synthesis rate, recycling and channeling. Channeling, recycling and protein synthesis must be estimated simultaneously because changes in specific radioactivities over time are highly dependent on the rate of protein synthesis. Injection-specific radioactivities, body weights and experimental variation were used with the model to generate data at different rates of recycling and channeling. The data generated were then used to determine the best time points and experimental method to estimate percentages of recycling, channeling and protein synthesis rate by the iterative Method of Maximum Likelihood. Specific radioactivity at each time point was based on simulated data from three rodents at each of six time points. Predicted protein synthesis rates were within 5%/d of observed rates for all methods. Predicted rates of recycling and channeling were generally within 15% of observed rates except recycling in muscle at high channeling and high recycling. Standard deviations of the predictions of percentages of channeling and recycling were between 0.148 and 44.5% for the pulse dose method, 0.0655 and 197% for the continuous infusion method and 0.351 and 962% for the flooding dose method. The experimental design that yields the best estimates of channeling, recycling and protein synthesis is the pulse dose. Changes in amino acid specific radioactivities in the extracellular, aminoacyl tRNA and protein pools were greatest and should be measured at 2, 6, 10, 40, 70 and 100 min in the pulse method. PMID- 11110877 TI - Creating a new paradigm in nutrition research within the National Cancer Institute. AB - Almost two decades after Doll and Peto (1981) provided evidence that one third of cancer deaths are related to diet, it remains unclear which dietary components may be key in cancer prevention. Although the complexity of the diet can become overwhelming, the National Cancer Institute (NCI) of the National Institutes of Health (NIH) has remained steadfast in its commitment to defining the roles that diet and nutrition have in the development of cancer and has provided increased research and training support to assist in unraveling this interrelationship. Evidence for this sustained commitment is highlighted by a fourfold increase in NCI expenditures for nutrition research and training from 1983 to 1998; this substantial increase reflects a trend that is occurring in some universities and the private sector. More than one third of the nutrition-related NCI research is funded by the Division of Cancer Prevention. Supported investigations cover the gamut from basic mechanisms of action of dietary constituents, methodology development, human metabolic studies, clinical trials of dietary modification and the chemopreventive potential of individual nutrients to population-based studies. PMID- 11110881 TI - Secretory, endocrine and autocrine/paracrine function of the adipocyte. AB - Obesity is a major public health problem in Western countries, and >55% of adult Americans are overweight or obese. A major contributor to the epidemic of obesity is the current environment, which is characterized by increased availability of high energy foods and decreased physical activity. Several studies also demonstrated that genetic susceptibility contributes to obesity in some populations. Obesity research has focused primarily on the role of the hypothalamus in neuroendocrine regulation of food intake. However, a growing number of studies support a potential contribution of adipose tissue, via its newly discovered secretory function, to the pathogenesis of obesity and co-morbid conditions including cardiovascular disease, diabetes and hypertension. This paper will review the role of four factors secreted by adipose tissue (leptin, agouti, angiotensin II and prostaglandins) and their functions in the regulation of energy balance and whole-body homeostasis. Several other peptide and nonpeptide substances are secreted from adipose tissue; their function and regulation have been documented extensively. PMID- 11110883 TI - Hormonal signaling and transcriptional control of adipocyte differentiation. AB - Recent advances regarding the biology of adipose tissue have identified the adipocyte as an important mediator in many physiologic and pathologic processes regarding energy metabolism. Consideration for a central role of adipose tissue in the development of obesity, cardiovascular disease and noninsulin-dependent diabetes mellitus has resulted in new incentives toward understanding the complexities of adipocyte differentiation. Current knowledge of this process includes a cascade of transcriptional events that culminate in the expression of peroxisome proliferator-activated receptor-gamma (PPARgamma) and CCAAT/enhancer binding protein-alpha (C/EBPalpha). These prominent adipogenic transcription factors have been shown to regulate, directly or indirectly, the gene expression necessary for the development of the mature adipocyte. Hormonal and nutritional signaling that impinges on these trans-acting factors provides a molecular link between lipids and lipid-related compounds and the gene expression important for glucose and lipid homeostasis. Knowledge concerning the transcriptional events mediating adipocyte differentiation provides a basis for understanding the physiologic processes associated with adipose tissue as well as for the development of therapeutic interventions in obesity and its related disorders. PMID- 11110885 TI - Adipocyte differentiation and gene expression. AB - The major function of adipocytes is to store triacylglycerol in periods of energy excess and to mobilize this energy during times of deprivation. The short-term control of these lipogenic and lipolytic processes is carefully modulated by hormonal signals from the bloodstream, which provide an inventory of the body's metabolic state. Long-term changes in fat storage needs are accomplished by altering both the size and number of fat cells within the body because terminally differentiated adipocytes cannot divide. Alterations in the number of fat cells within the body must be accomplished by the differentiation of preadipocytes, which act as the renewable source of adipocytes. Our understanding of the events that occur during preadipocyte differentiation has advanced considerably in the last few years and has relied mainly on the use of tissue culture models of adipogenesis. This article will discuss the various models used for studying the preadipocyte differentiation process, with the mouse 3T3-L1 cell culture line described in detail. We focus on those genetic events that link effectors to induction of adipocyte gene expression. PMID- 11110887 TI - Regulation of leptin production in humans. AB - Serum levels of the adipocyte hormone leptin are increased in proportion to body fat stores as a result of increased production in enlarged fat cells from obese subjects. In vitro studies indicate that insulin and glucocorticoids work directly on adipose tissue to upregulate in a synergistic manner leptin mRNA levels and rates of leptin secretion in human adipose tissue over the long term. Thus, the increased leptin expression observed in obesity could result from the chronic hyperinsulinemia and increased cortisol turnover. Superimposed upon the long-term regulation, nutritional status can influence serum leptin over the short term, independent of adiposity. Fasting leads to a gradual decline in serum leptin that is probably attributable to the decline in insulin and the ability of catecholamines to decrease leptin expression, as observed in both in vivo and in vitro studies. In addition, increases in serum leptin occur approximately 4-7 h after meals. Increasing evidence indicates that insulin, in concert with permissive effects of cortisol, can increase serum leptin over this time frame and likely contributes to meal-induced increases in serum leptin. Further research is required to elucidate the cellular and molecular mechanisms underlying short- and long-term nutritional and hormonal regulation of leptin production and secretion. PMID- 11110889 TI - Genetic studies of brown adipocyte induction. AB - We seek to discover an effective method for utilizing thermogenesis to reduce the caloric load in obese individuals. Experimental evidence indicates that nonshivering thermogenesis is the most effective cellular and biochemical mechanism known for reducing excessive adiposity. In this presentation, we describe our experiments aimed at understanding how nonshivering thermogenesis can be induced. In addition, these experiments have led to a genetic approach for the identification of variant genes that coordinate the expression of pathways of gene transcription that are associated with brown adipocyte induction. PMID- 11110892 TI - Evolutionary annotation of the genome. PMID- 11110893 TI - A case for evolutionary genomics and the comprehensive examination of sequence biodiversity. AB - Comparative analysis is one of the most powerful methods available for understanding the diverse and complex systems found in biology, but it is often limited by a lack of comprehensive taxonomic sampling. Despite the recent development of powerful genome technologies capable of producing sequence data in large quantities (witness the recently completed first draft of the human genome), there has been relatively little change in how evolutionary studies are conducted. The application of genomic methods to evolutionary biology is a challenge, in part because gene segments from different organisms are manipulated separately, requiring individual purification, cloning, and sequencing. We suggest that a feasible approach to collecting genome-scale data sets for evolutionary biology (i.e., evolutionary genomics) may consist of combination of DNA samples prior to cloning and sequencing, followed by computational reconstruction of the original sequences. This approach will allow the full benefit of automated protocols developed by genome projects to be realized; taxon sampling levels can easily increase to thousands for targeted genomes and genomic regions. Sequence diversity at this level will dramatically improve the quality and accuracy of phylogenetic inference, as well as the accuracy and resolution of comparative evolutionary studies. In particular, it will be possible to make accurate estimates of normal evolution in the context of constant structural and functional constraints (i.e., site-specific substitution probabilities), along with accurate estimates of changes in evolutionary patterns, including pairwise coevolution between sites, adaptive bursts, and changes in selective constraints. These estimates can then be used to understand and predict the effects of protein structure and function on sequence evolution and to predict unknown details of protein structure, function, and functional divergence. In order to demonstrate the practicality of these ideas and the potential benefit for functional genomic analysis, we describe a pilot project we are conducting to simultaneously sequence large numbers of vertebrate mitochondrial genomes. PMID- 11110894 TI - Testing the "proto-splice sites" model of intron origin: evidence from analysis of intron phase correlations. AB - A few nucleotide sites of nuclear exons that flank introns are often conserved. A hypothesis has suggested that these sites, called "proto-splice sites," are remnants of recognition signals for the insertion of introns in the early evolution of eukaryotic genes. This notion of proto-splice sites has been an important basis for the insertional theory of introns. This hypothesis predicts that the distribution of proto-splice sites would determine the distribution of intron phases, because the positions of introns are just a subset of the proto splice sites. We previously tested this prediction by examining the proportions of the phases of proto-splice sites, revealing nothing in these proportion distributions similar to observed proportions of intron phases. Here, we provide a second independent test of the proto-splice site hypothesis, with regard to its prediction that the proto-splice sites would mimic intron phase correlations, using a CDS database we created from GenBank. We tested four hypothetical proto splice sites G / G, AG / G, AG / GT, and C/AAG / R. Interestingly, while G / G and AG / GT site phase distributions are not consistent with actual introns, we observed that AG / G and C/AAG / R sites have a symmetric phase excess. However, the patterns of the excess are quite different from the actual intron phase distribution. In addition, particular amino acid repeats in proteins were found to partially contribute to the excess of symmetry at these two types of sites. The phase associations of all four sites are significantly different from those of intron phases. Furthermore, a general model of intron insertion into proto splice sites was simulated by Monte Carlo simulation to investigate the probability that the random insertion of introns into AG / G and C/AAG / R sites could generate the observed intron phase distribution. The simulation showed that (1) no observed correlation of intron phases was statistically consistent with the phase distribution of proto-splice sites in the simulated virtual genes; (2) most conservatively, no simulation in 10,000 Monte Carlo experiments gave a pattern with an excess of symmetric (1, 1) exons larger than those of (0, 0) and (2, 2), a major statistical feature of intron phase distribution that is consistent with the directly observed cases of exon shuffling. Thus, these results reject the null hypothesis that introns are randomly inserted into preexisting proto-splice sites, as suggested by the insertional theory of introns. PMID- 11110895 TI - Slow rate of evolution in the mitochondrial control region of gulls (Aves: Laridae). AB - We sequenced part of the mitochondrial control region and the cytochrome b gene in 72 specimens from 32 gull species (Laridae, Larini) and 2 outgroup representatives (terns: Laridae, Sternini). Our control region segment spanned the conserved central domain II and the usually hypervariable 3' domain III. Apart from some heteroplasmy at the 3' end of the control region, domain III was not more variable than domain II or the cytochrome b gene. Furthermore, variation in the tempo of evolution of domain III was apparent between phyletic species groups. The lack of variation of the gull control region could not be explained by an increase in the proportion of conserved sequences in these birds, and the gull control region showed an organization similar to those of other avian control regions studied to date. A novel invariant direct repeat was identified in domain II of gulls, and in domain III, two to three inverted, sometimes imperfect, repeats are able to form a significantly stable stem-and-loop structure. These putative secondary structures have not been reported before, and a comparison between species groups showed that they are more stable in the group with the more conserved control region. The unusually slow rate of evolution of control region part III of the gulls could thus be partly explained by the existence of secondary structures in domain III of these species. PMID- 11110896 TI - Reconstructing genealogies of serial samples under the assumption of a molecular clock using serial-sample UPGMA. AB - Reconstruction of evolutionary relationships from noncontemporaneous molecular samples provides a new challenge for phylogenetic reconstruction methods. With recent biotechnological advances there has been an increase in molecular sequencing throughput, and the potential to obtain serial samples of sequences from populations, including rapidly evolving pathogens, is fast being realized. A new method called the serial-sample unweighted pair grouping method with arithmetic means (sUPGMA) is presented that reconstructs a genealogy or phylogeny of sequences sampled serially in time using a matrix of pairwise distances. The resulting tree depicts the terminal lineages of each sample ending at a different level consistent with the sample's temporal order. Since sUPGMA is a variant of UPGMA, it will perform best when sequences have evolved at a constant rate (i.e., according to a molecular clock). On simulated data, this new method performs better than standard cluster analysis under a variety of longitudinal sampling strategies. Serial-sample UPGMA is particularly useful for analysis of longitudinal samples of viruses and bacteria, as well as ancient DNA samples, with the minimal requirement that samples of sequences be ordered in time. PMID- 11110897 TI - An ancient transpecific polymorphism shows extreme divergence in a multitrait cline in an intertidal snail (Nucella lapillus (L.)). AB - Clines in intraspecific genetic variation are frequently associated with an environmental transition. Here, divergence among nucleotide sequences of two nuclear loci, cytosolic and mitochondrial malate dehydrogenase (cMDH and mMDH, respectively), is described, in a multitrait cline over a distance of ca. 3 km where shell phenotype, allozyme, mitochondrial DNA haplotype, and centric fusion (Robertsonian translocations) frequencies covary with temperature and humidity and change abruptly in a continuous population of the dog-whelk (Nucella lapillus), a common intertidal snail of the north temperate Atlantic. Protein electrophoresis has already shown two alleles of mMDH varying from fixation of one allele to near fixation of the other, whereas cMDH appears to be monomorphic. The results of this study show a striking disparity in nucleotide sequence divergence among alleles at the two loci, with extreme molecular differentiation in one of them. Four alleles of cMDH were found to have nucleotide and amino acid sequence divergences of 0.4% and 0.3%, respectively. In contrast, the two mMDH cDNA alleles differed by 23% and 20% at the nucleotide and amino acid levels, respectively. Analysis of a 91-bp partial nucleotide sequence of mMDH from Nucella freycineti, the closest relative of N. lapillus, revealed two similar alleles and indicated that the divergence in mMDH in N. lapillus represents an ancient transpecific polymorphism in these Nucella. Together with earlier studies on variation in N. lapillus, it is argued that the polymorphism in mMDH and the clines in N. lapillus represent the presence of two persistent coadapted gene complexes, multitrait coevolving genetic solutions to environmental variation, which may presently enable this snail to exploit a diverse environment successfully. PMID- 11110898 TI - Rooting a phylogeny with homologous genes on opposite sex chromosomes (gametologs): a case study using avian CHD. AB - We describe a previously unrecognized form of gene homology using the term "gametology," which we define as homology arising through lack of recombination and subsequent differentiation of sex chromosomes. We demonstrate use of gametologous genes to root each other in phylogenetic analyses of sex-specific avian Chromo-helicase-DNA binding gene (CHD) sequences. Phylogenetic analyses of a set of neognath bird sequences yield monophyletic groups for CHD-W and CHD-Z gametologs, as well as congruent relationships between these two clades and between them and current views of avian taxonomy. Phylogenetic analyses including paleognath bird CHD sequences and rooting with crocodilian CHD sequences, suggest an early divergence for paleognath CHD within the avian CHD clade. Based on our CHD analyses calibrated with avian fossil dates, we estimate the divergence between CHD-W and CHD-Z at 123 MYA, suggesting an early differentiation of sex chromosomes that predates most extant avian orders. In agreement with the notion of male-driven evolution, we find a faster rate of change in male-linked CHD-Z sequences. PMID- 11110899 TI - Diversity in organization and the origin of gene orders in the mitochondrial DNA molecules of the genus Saccharomyces. AB - Sequencing of the Saccharomyces cerevisiae nuclear and mitochondrial genomes provided a new background for studies on the evolution of the genomes. In this study, mitochondrial genomes of a number of Saccharomyces yeasts were mapped by restriction enzyme analysis, the orders of the genes were determined, and two of the genes were sequenced. The genome organization, i.e., the size, presence of intergenic sequences, and gene order, as well as polymorphism within the coding regions, indicate that Saccharomyces mtDNA molecules are dynamic structures and have undergone numerous changes during their evolution. Since the separation and sexual isolation of different yeast lineages, the coding parts have been accumulating point mutations, presumably in a linear manner with the passage of time. However, the accumulation of other changes may not have been a simple function of time. Larger mtDNA molecules belonging to Saccharomyces sensu stricto yeasts have acquired extensive intergenic sequences, including guanosine-cytosine rich clusters, and apparently have rearranged the gene order at higher rates than smaller mtDNAs belonging to the Saccharomyces sensu lato yeasts. While within the sensu stricto group transposition has been a predominant mechanism for the creation of novel gene orders, the sensu lato yeasts could have used both transposition- and inversion-based mechanisms. PMID- 11110900 TI - Effects of sequence alignment and structural domains of ribosomal DNA on phylogeny reconstruction for the protozoan family sarcocystidae. AB - Finding correct species relationships using phylogeny reconstruction based on molecular data is dependent on several empirical and technical factors. These include the choice of DNA sequence from which phylogeny is to be inferred, the establishment of character homology within a sequence alignment, and the phylogeny algorithm used. Nevertheless, sequencing and phylogeny tools provide a way of testing certain hypotheses regarding the relationship among the organisms for which phenotypic characters demonstrate conflicting evolutionary information. The protozoan family Sarcocystidae is one such group for which molecular data have been applied phylogenetically to resolve questionable relationships. However, analyses carried out to date, particularly based on small-subunit ribosomal DNA, have not resolved all of the relationships within this family. Analysis of more than one gene is necessary in order to obtain a robust species signal, and some DNA sequences may not be appropriate in terms of their phylogenetic information content. With this in mind, we tested the informativeness of our chosen molecule, the large-subunit ribosomal DNA (lsu rDNA), by using subdivisions of the sequence in phylogenetic analysis through PAUP, fastDNAml, and neighbor joining. The segments of sequence applied correspond to areas of higher nucleotide variation in a secondary-structure alignment involving 21 taxa. We found that subdivision of the entire lsu rDNA is inappropriate for phylogenetic analysis of the Sarcocystidae. There are limited informative nucleotide sites in the lsu rDNA for certain clades, such as the one encompassing the subfamily Toxoplasmatinae. Consequently, the removal of any segment of the alignment compromises the final tree topology. We also tested the effect of using two different alignment procedures (CLUSTAL W and the structure alignment using DCSE) and three different tree-building methods on the final tree topology. This work shows that congruence between different methods in the formation of clades may be a feature of robust topology; however, a sequence alignment based on primary structure may not be comparing homologous nucleotides even though the expected topology is obtained. Our results support previous findings showing the paraphyly of the current genera Sarcocystis and Hammondia and again bring to question the relationships of Sarcocystis muris, Isospora felis, and Neospora caninum. In addition, results based on phylogenetic analysis of the structure alignment suggest that Sarcocystis zamani and Sarcocystis singaporensis, which have reptilian definitive hosts, are monophyletic with Sarcocystis species using mammalian definitive hosts if the genus Frenkelia is synonymized with Sarcocystis. PMID- 11110901 TI - Assessing an unknown evolutionary process: effect of increasing site-specific knowledge through taxon addition. AB - Assessment of the evolutionary process is crucial for understanding the effect of protein structure and function on sequence evolution and for many other analyses in molecular evolution. Here, we used simulations to study how taxon sampling affects accuracy of parameter estimation and topological inference in the absence of branch length asymmetry. With maximum-likelihood analysis, we find that adding taxa dramatically improves both support for the evolutionary model and accurate assessment of its parameters when compared with increasing the sequence length. Using a method we call "doppelganger trees," we distinguish the contributions of two sources of improved topological inference: greater knowledge about internal nodes and greater knowledge of site-specific rate parameters. Surprisingly, highly significant support for the correct general model does not lead directly to improved topological inference. Instead, substantial improvement occurs only with accurate assessment of the evolutionary process at individual sites. Although these results are based on a simplified model of the evolutionary process, they indicate that in general, assuming processes are not independent and identically distributed among sites, more extensive sampling of taxonomic biodiversity will greatly improve analytical results in many current sequence data sets with moderate sequence lengths. PMID- 11110902 TI - A power analysis of microsatellite-based statistics for inferring past population growth. AB - We present results concerning the power to detect past population growth using three microsatellite-based statistics available in the current literature: (1) that based on between-locus variability, (2) that based on the shape of allele size distribution, and (3) that based on the imbalance between variance and heterozygosity at a locus. The analysis is based on the single-step stepwise mutation model. The power of the statistics is evaluated for constant, as well as variable, mutation rates across loci. The latter case is important, since it is a standard procedure to pool data collected at a number of loci, and mutation rates at microsatellite loci are known to be different. Our analysis indicates that the statistic based on the imbalance between allele size variance and heterozygosity at a locus has the highest power for detection of population growth, particularly when mutation rates vary across loci. PMID- 11110903 TI - Molecular phylogeny of osteoglossoids: a new model for Gondwanian origin and plate tectonic transportation of the Asian arowana. AB - One of the traditional enigmas in freshwater zoogeography has been the evolutionary origin of Scleropages formosus inhabiting Southeast Asia (the Asian arowana), which is a species threatened with extinction among the highly freshwater-adapted fishes from the order Osteoglossiformes. Dispersalists have hypothesized that it originated from the recent (the Miocene or later) transmarine dispersal of morphologically quite similar Australasian arowanas across Wallace's Line, but this hypothesis has been questioned due to their remarkable adaptation to freshwater. We determined the complete nucleotide sequences of two mitochondrial protein genes from 12 osteoglossiform species, including all members of the suborder Osteoglossoidei, with which robust molecular phylogeny was constructed and divergence times were estimated. In agreement with previous morphology-based phylogenetic studies, our molecular phylogeny suggested that the osteoglossiforms diverged from a basal position of the teleostean lineage, that heterotidines (the Nile arowana and the pirarucu) form a sister group of osteoglossines (arowanas in South America, Australasia, and Southeast Asia), and that the Asian arowana is more closely related to Australasian arowanas than to South American ones. However, molecular distances between the Asian and Australasian arowanas were much larger than expected from the fact that they are classified within the same genus. By using the molecular clock of bony fishes, tested for its good performance for rather deep divergences and calibrated using some reasonable assumptions, the divergence between the Asian and Australasian arowanas was estimated to date back to the early Cretaceous. Based on the molecular and geological evidence, we propose a new model whereby the Asian arowana vicariantly diverged from the Australasian arowanas in the eastern margin of Gondwanaland and migrated into Eurasia on the Indian subcontinent or smaller continental blocks. This study also implicates the relatively long absence of osteoglossiform fossil records from the Mesozoic. PMID- 11110904 TI - A simple method for classifying genes and a bootstrap test for classifications. AB - A new simple method for classifying genes is proposed based on Klastorin's method. This method classifies genes into monophyletic groups which are made distinct from each other by evolutionary changes. The method is applicable as long as the phylogenetic tree of genes is obtained. There is a fast algorithm for obtaining the classification. A bootstrap test of a classification is also presented. As an example, we classified opsin genes. The classification obtained by this method is the same as the previous classification based on the function of opsins. PMID- 11110905 TI - Multigene phylogeny of land plants with special reference to bryophytes and the earliest land plants. AB - A widely held view of land plant relationships places liverworts as the first branch of the land plant tree, whereas some molecular analyses and a cladistic study of morphological characters indicate that hornworts are the earliest land plants. To help resolve this conflict, we used parsimony and likelihood methods to analyze a 6, 095-character data set composed of four genes (chloroplast rbcL and small-subunit rDNA from all three plant genomes) from all major land plant lineages. In all analyses, significant support was obtained for the monophyly of vascular plants, lycophytes, ferns (including PSILOTUM: and EQUISETUM:), seed plants, and angiosperms. Relationships among the three bryophyte lineages were unresolved in parsimony analyses in which all positions were included and weighted equally. However, in parsimony and likelihood analyses in which rbcL third-codon-position transitions were either excluded or downweighted (due to apparent saturation), hornworts were placed as sister to all other land plants, with mosses and liverworts jointly forming the second deepest lineage. Decay analyses and Kishino-Hasegawa tests of the third-position-excluded data set showed significant support for the hornwort-basal topology over several alternative topologies, including the commonly cited liverwort-basal topology. Among the four genes used, mitochondrial small-subunit rDNA showed the lowest homoplasy and alone recovered essentially the same topology as the multigene tree. This molecular phylogeny presents new opportunities to assess paleontological evidence and morphological innovations that occurred during the early evolution of terrestrial plants. PMID- 11110906 TI - A phylogenetic tree of the Wnt genes based on all available full-length sequences, including five from the cephalochordate amphioxus. AB - The WNT: gene family is large, and new members are still being discovered. We constructed a parsimony tree for the WNT: family based on all 82 of the full length sequences currently available. The inclusion of sequences from the cephalochordate amphioxus is especially useful in comprehensive gene trees, because the amphioxus genes in each subfamily often mark the base of the vertebrate diversification. We thus isolated full-length cDNAs of five amphioxus WNT: genes (AmphiWnt1, AmphiWnt4, AmphiWnt7, AmphiWnt8, and AmphiWnt11) for addition to the overall WNT: family tree. The analysis combined amino acid and nucleotide sequences (excluding third codon positions), taking into account 97% of the available data for each sequence. This combinatorial method had the advantage of generating a single most-parsimonious tree that was trichotomy-free. The reliability of the nodes was assessed by both jackknifing and Bremer support (decay index). A regression analysis revealed that branch length was strongly correlated with branch support, and possible reasons for this pattern are discussed. The tree topology suggested that in amphioxus, at least an AmphiWnt5 and an AmphiWnt10 have yet to be discovered. PMID- 11110907 TI - Evidence for diet effects on the composition of silk proteins produced by spiders. AB - Silks are highly expressed, secreted proteins that represent a substantial metabolic cost to the insects and spiders that produce them. Female spiders in the superfamily Araneoidea (the orb-spinning spiders and their close relatives) spin six different kinds of silk (three fibroins and three fibrous protein glues) that differ in amino acid content and protein structure. In addition to this diversity in silks produced by different glands, we found that individual spiders of the same species can spin dragline silks (drawn from the spider's ampullate gland) that vary in content as well. Freely foraging ARGIOPE: argentata (Araneae: Araneoidea), collected from 13 Caribbean islands, produced dragline silk that showed an inverse relationship between the amount of serine and glycine they contained. X-ray microdiffraction of the silks localized these differences to the amorphous regions of the protein that are thought to lend silks their elasticity. The crystalline regions of the proteins, which lend silks their strength, were unaffected. Laboratory experiments with ARGIOPE: keyserlingi suggested that variation in silk composition reflects the type of prey the spiders were fed but not the total amount of prey they received. Hence, it may be that the amino acid content (and perhaps the mechanical properties) of dragline silk spun by ARGIOPE: directly reflect the spiders' diet. The ability to vary silk composition and, possibly, function is particularly important for organisms that disperse broadly, such as Argiope, and that occupy diverse habitats with diverse populations of prey. PMID- 11110908 TI - Wilkinson support calculated with exact probabilities: an example using Floricaula/LEAFY amino acid sequences that compares three hypotheses involving gene gain/loss in seed plants. AB - This paper describes a method for quantifying the extent to which a character supports a hypothesized monophyletic group. The basic idea was first proposed by Wilkinson in 1998; hence, we call it Wilkinson support. A character provides Wilkinson support if it could have changed state on the branch leading to the hypothesized monophyletic group without requiring any extra steps in an evolutionary tree. We describe a method to determine the exact probability that a character would provide Wilkinson support for a random group of the same size as the hypothesized monophyletic group. A character's weight is defined as the negative natural log of this probability. The sum over all characters of these weights in a data set is a measure of total weighted support. We exemplify this method using 30 Floricaula/LEAFY amino acid sequences. One copy of this gene occurs in angiosperms, but two copies occur in the other four seed plant groups. Angiosperms could have been primitively single-copy or could have lost either of the two paralogs. These possibilities correspond to three hypotheses of monophyly. We use total weighted Wilkinson support to evaluate these three hypotheses, and all three are shown to be significantly different from random as individual hypothesized monophyletic groups. Comparing these three hypotheses for total weighted support reveals that one has much more support than do the other two. This hypothesis favors the "mostly-male" theory of flowering-plant origins. PMID- 11110909 TI - Microsatellite size homoplasy, SSCP, and population structure: a case study in the freshwater snail Bulinus truncatus. AB - The extent of microsatellite size homoplasy, as well as its effect on several population genetics statistics, was investigated in natural populations using the single-strand conformation polymorphism (SSCP) method. The analysis was conducted using 240 individuals from 13 populations of the freshwater snail Bulinus truncatus at a GT(n)CT(m) compound microsatellite locus. We showed that SSCP can be used to uncover, at least partly, size homoplasy in the core sequence of this category of loci. Eight conformers (SSCP variants) were detected among the three size variants (electromorphs). Sequencing revealed that each conformer corresponded to a different combination of repeats in the GT(n) and CT(m) arrays. Part of this additional variability was detected within populations, resulting in a substantial increase in gene diversity in four populations. Additional variability also changed the values of parameters used to analyze population differentiation among populations: pairwise tests of differentiation were significant much more often with conformers than with electromorphs. On the other hand, pairwise estimates of F(st) were either smaller or larger with conformers than with electromorphs, depending on whether or not electromorphs were shared among populations. However, estimates of F(st) (or analogs) over all populations were very similar, ranging between 0.66 and 0.75. Our results were consistent with the theoretical prediction that homoplasy should not always lead to stronger population structure. Finally, conformer sequences and electromorph size distribution suggested that single-point and/or stepwise mutations occurring simultaneously in the different repeated arrays of compound microsatellites produce sequence variation without size variation and hence generate more size homoplasy than expected under a simple stepwise mutation model. PMID- 11110910 TI - Purifying selection detected in the plastid gene matK and flanking ribozyme regions within a group II intron of nonphotosynthetic plants. AB - In a striking contrast, matK is one of the most rapidly evolving plastid genes and also one of the few plastid genes to be retained in all nonphotosynthetic plants examined to date. DNA sequences of this region were obtained from photosynthetic and nonphotosynthetic plants of Orobanchaceae and their relatives. The resulting plastid DNA phylogeny was congruent with that recently obtained from analyses of rps2 and provided much better resolution. This phylogeny was then used to examine the relative degrees of evolutionary constraint of both the matK gene and the non-protein-coding regions that flank it inside the trnK intron. The method of subtree contrasts was introduced to compare levels of constraint. matK has evolved with a low but significant level of constraint on its amino acid sequence in both photosynthetic and nonphotosynthetic plants. Constraint is greater in photosynthetic than in nonphotosynthetic plants of this group. Domain X, thought to contain the active site of the protein, is not significantly more constrained than the rest of the protein. The portions of the flanking regions that are thought to form paired stem structures also show constraint, but in this case, there is no significant difference in degree of constraint between photosynthetic and nonphotosynthetic plants. PMID- 11110911 TI - Nucleotide variation at the yellow gene region is not reduced in Drosophila subobscura: a study in relation to chromosomal polymorphism. AB - In contrast to Drosophila melanogaster and Drosophila simulans, the yellow (y) gene region of Drosophila subobscura is not located in a region with a strong reduction in recombination. In addition, this gene maps very close to the breakpoints of different inversions that segregate as polymorphic in natural populations of D. subobscura. Therefore, levels of variation at the y gene region in this species relative to those found in D. melanogaster and D. simulans may be affected not only by the change in the recombinational environment, but also by the presence of inversion polymorphism. To further investigate these aspects, an approximately 5.4-kb region of the A (=X) chromosome including the y gene was sequenced in 25 lines of D. subobscura and in the closely related species Drosophila madeirensis and Drosophila guanche. The D. subobscura lines studied differed in their A-chromosomal arrangements, A(st), A(2), and A(1). Unlike in D. melanogaster and D. simulans, levels of variation at the y gene region of D. subobscura are not reduced relative to those found at other genomic regions in the same species (rp49, Acp70A, and Acph-1). This result supports the effect of the change in the recombinational environment of a particular gene on the level of neutral variation. In addition, nucleotide variation is affected by chromosomal polymorphism. A strong genetic differentiation is detected between the A(1) arrangement and either A(st) or A(2), but not between A(st) and A(2). This result is consistent with the location of the y gene relative to the breakpoints of inversions A(1) and A(2). In addition, the pattern of nucleotide polymorphism in A(st)+A(2) and A(1) seems to point out that variation at the y gene region within these chromosomal classes is in the phase transient to equilibrium. The estimated ages of these arrangements assuming a star genealogy indicate that their origin cannot predate the D. madeirensis split. Therefore, the present results are consistent with a chromosomal phylogeny where Am(1), which is an arrangement present in D. madeirensis but absent in current populations of D. subobscura, would be the ancestral arrangement. PMID- 11110912 TI - Evolution of two-component signal transduction. AB - Two-component signal transduction (TCST) systems are the principal means for coordinating responses to environmental changes in bacteria as well as some plants, fungi, protozoa, and archaea. These systems typically consist of a receptor histidine kinase, which reacts to an extracellular signal by phosphorylating a cytoplasmic response regulator, causing a change in cellular behavior. Although several model systems, including sporulation and chemotaxis, have been extensively studied, the evolutionary relationships between specific TCST systems are not well understood, and the ancestry of the signal transduction components is unclear. Phylogenetic trees of TCST components from 14 complete and 6 partial genomes, containing 183 histidine kinases and 220 response regulators, were constructed using distance methods. The trees showed extensive congruence in the positions of 11 recognizable phylogenetic clusters. Eukaryotic sequences were found almost exclusively in one cluster, which also showed the greatest extent of domain variability in its component proteins, and archaeal sequences mainly formed species-specific clusters. Three clusters in different parts of the kinase tree contained proteins with serine-phosphorylating activity. All kinases were found to be monophyletic with respect to other members of their superfamily, such as type II topoisomerases and Hsp90. Structural analysis further revealed significant similarity to the ATP-binding domain of eukaryotic protein kinases. TCST systems are of bacterial origin and radiated into archaea and eukaryotes by lateral gene transfer. Their components show extensive coevolution, suggesting that recombination has not been a major factor in their differentiation. Although histidine kinase activity is prevalent, serine kinases have evolved multiple times independently within this family, accompanied by a loss of the cognate response regulator(s). The structural and functional similarity between TCST kinases and eukaryotic protein kinases raises the possibility of a distant evolutionary relationship. PMID- 11110913 TI - Widespread occurrence of spliceosomal introns in the rDNA genes of ascomycetes. AB - Spliceosomal (pre-mRNA) introns have previously been found in eukaryotic protein coding genes, in the small nuclear RNAs of some fungi, and in the small- and large-subunit ribosomal DNA genes of a limited number of ascomycetes. How the majority of these introns originate remains an open question because few proven cases of recent and pervasive intron origin have been documented. We report here the widespread occurrence of spliceosomal introns (69 introns at 27 different sites) in the small- and large-subunit nuclear-encoded rDNA of lichen-forming and free-living members of the Ascomycota. Our analyses suggest that these spliceosomal introns are of relatively recent origin, i.e., within the Euascomycetes, and have arisen through aberrant reverse-splicing (in trans) of free pre-mRNA introns into rRNAs. The spliceosome itself, and not an external agent (e.g., transposable elements, group II introns), may have given rise to these introns. A nonrandom sequence pattern was found at sites flanking the rRNA spliceosomal introns. This pattern (AG-intron-G) closely resembles the proto splice site (MAG-intron-R) postulated for intron insertions in pre-mRNA genes. The clustered positions of spliceosomal introns on secondary structures suggest that particular rRNA regions are preferred sites for insertion through reverse splicing. PMID- 11110914 TI - Inferring lifestyle from gene expression patterns. PMID- 11110915 TI - Rapid evaluation of the phylogenetic congruence of sequence data using likelihood ratio tests. PMID- 11110917 TI - Book reviewers - A note of thanks PMID- 11110916 TI - Manuscript reviewers - A note of thanks PMID- 11110918 TI - Abstractors of current literature - A note of thanks PMID- 11110919 TI - Research trainee prizes from the 2000 RSNA scientific assembly and annual meeting PMID- 11110920 TI - Medical physics: some recollections in diagnostic X-ray imaging and therapeutic radiology. AB - Medical physics has changed dramatically since 1895. There was a period of slow evolutionary change during the first 70 years after Roentgen's discovery of x rays. With the advent of the computer, however, both diagnostic and therapeutic radiology have undergone rapid growth and changes. Technologic advances such as computed tomography and magnetic resonance imaging in diagnostic imaging and three-dimensional treatment planning systems, stereotactic radiosurgery, and intensity modulated radiation therapy in radiation oncology have resulted in substantial changes in medical physics. These advances have improved diagnostic imaging and radiation therapy while expanding the need for better educated and experienced medical physics staff. PMID- 11110921 TI - Health care rationing: every physician's dilemma. PMID- 11110922 TI - CT characterization of adrenal masses: the time has come. PMID- 11110923 TI - Tumor ablation with radio-frequency energy. AB - Tumor ablation by using radio-frequency energy has begun to receive increased attention as an effective minimally invasive approach for the treatment of patients with a variety of primary and secondary malignant neoplasms. To date, these techniques have been used to treat tumors located in the brain, musculoskeletal system, thyroid and parathyroid glands, pancreas, kidney, lung, and breast; however, liver tumor ablation has received the greatest attention and has been the subject of a large number of published reports. In this article, the authors review the technical developments and early laboratory results obtained with radio-frequency ablation techniques, describe some of the early clinical applications of these techniques, and conclude with a discussion of challenges and opportunities for the future. PMID- 11110924 TI - Pulmonary infections in immunocompromised hosts: the importance of correlating the conventional radiologic appearance with the clinical setting. AB - The lung is one of the most frequently involved organs in a variety of complications in the immunocompromised host. Among the pulmonary complications that occur in this kind of patient, infection is the most common and is associated with high morbidity and mortality. Although chest radiography and computed tomography (CT) are essential diagnostic tools, radiologists often have difficulty in establishing the correct diagnosis on the basis of radiologic findings alone. The reasons are that the immunocompromised host is potentially susceptible to infection from many different microorganisms and that radiologic findings are seldom specific for the detection of a particular pathogen. Experience has shown that a particular clinical setting predisposes patients to infection by particular pathogens. The setting comprises (a) the specific epidemiologic or environmental exposure, (b) the type of underlying immune defect, (c) the duration and severity of immune compromise, and (d) the progression rate and pattern of the radiologic abnormality. Correlating the radiologic appearance with the clinical setting can expedite diagnosis and appropriate therapy. In this review, the authors describe the clinical settings that are helpful in choosing the radiologic approach to treatment of the immunocompromised host who presents with suspected pulmonary infection. PMID- 11110925 TI - MR imaging-guided percutaneous cryotherapy of liver tumors: initial experience. AB - PURPOSE: To describe the cryoablation of liver tumors by using a percutaneous approach and intraprocedural magnetic resonance (MR) imaging monitoring and to assess the feasibility and safety of the procedure. MATERIALS AND METHODS: Fifteen hepatic tumors (mean diameter, 2.9 cm) in 12 patients were treated (18 total cryoablations). Fourteen were metastases and one was a hemangioma; all were proved at biopsy. By using a 0.5-T open MR imaging system, cryoneedles were placed and lesions ablated by using real-time monitoring. Clinical signs and symptoms were assessed and laboratory tests performed. Intraprocedural depictions of iceballs were compared with contrast material-enhanced MR imaging-based estimates of cryonecrosis that were obtained 24 hours after cryoablation. RESULTS: MR imaging-guided percutaneous cryotherapy resulted in no serious complications and no clinically important changes in serum liver enzymes or creatinine or myoglobin levels. Intraprocedural MR imaging demonstrated iceballs as sharply marginated regions of signal loss that expanded and engulfed tumors. The maximal iceball size was 4.9 x 2.2 x 2.2 cm with the use of one cryoneedle and 6.0 x 5.6 x 4.9 cm with three cryoneedles. Intraprocedural iceball depictions correlated well with postprocedural cryonecrosis estimates. CONCLUSION: MR imaging-guided percutaneous cryotherapy of liver tumors is feasible and safe. MR imaging can be used to estimate cryotherapy effects and guide therapy intraprocedurally. PMID- 11110926 TI - Radio-frequency ablation of renal cell carcinoma: early clinical experience. AB - PURPOSE: To report the authors' early experience with radio-frequency (RF) ablation of renal cell carcinoma. MATERIALS AND METHODS: Twenty-four percutaneous RF ablation treatments for nine tumors were performed in eight patients with renal cell carcinoma. Indications included coexistent morbidity, previous surgery, or solitary kidney in patients with a life expectancy shorter than 10 years. Smaller (3 cm) and/or central lesions (n = 6) were treated with cluster or multiple electrodes. Patients returned for a second treatment when follow-up imaging depicted tumor enhancement. Follow-up imaging was performed at 1 and 3 months and then at 6-month intervals, with a mean follow up of 10.3 months. Seven patients were alive at least 6 months after their initial treatment. RESULTS: All five exophytic tumors were free of enhancement. One of three central tumors was free of enhancement. One tumor had both central and exophytic components and was free of enhancement. Three tumors were 3 cm or smaller and free of enhancement. Of the six tumors larger than 3 cm, four were free of enhancement. CONCLUSION: Percutaneous RF ablation is a promising treatment for select patients with renal cell carcinoma. The ultimate role of this modality will continue to evolve and warrants further study. PMID- 11110927 TI - Thyroid tissue: US-guided percutaneous interstitial laser ablation-a feasibility study. AB - PURPOSE: To evaluate percutaneous interstitial laser photocoagulation (ILP) as a palliative treatment of recurrent thyroid carcinoma untreatable with surgery or radioiodine administration. MATERIALS AND METHODS: By using 18 resected thyroid glands, the volume and histologic pattern of ILP-induced thyroid damage were assessed. In vivo treatment feasibility was evaluated by using a low-energy laser in two volunteers before thyroidectomy for huge autonomously functioning nodules. With ultrasonographic (US) monitoring, a 21-gauge spinal needle was inserted into the thyroid nodules. A 300-microm quartz fiberoptic guide was inserted through the needle lumen, and the fiber tip was placed in direct contact with the tissue. Laser irradiation was performed with a 1.064-nm Nd:YAG laser in surgically resected glands, which were treated with 2, 3, 5, or 7 W. RESULTS: Tissue ablation was well-defined histologically, and its area was related to laser irradiation parameters (range, 0-26 mm). No correlation was found between US images and the actual extent of laser-induced lesions. Large colloid or fluid collections did not permit regular heat diffusion within the tissue. In vivo low energy ILP was performed without technical difficulties or complications. CONCLUSION: ILP induces well-defined tissue ablation correlated with energy parameters in thyroid glands devoid of cystic areas. ILP could be a therapeutic tool for highly selected problems in thyroid tumor treatment. PMID- 11110928 TI - Hydrodynamic thrombectomy system versus pulse-spray thrombolysis for thrombosed hemodialysis grafts: a multicenter prospective randomized comparison. AB - PURPOSE: To evaluate the safety and efficacy of a hydrodynamic thrombectomy system in a prospective, multicenter randomized comparison with pulse-spray thrombolysis in hemodialysis grafts. MATERIALS AND METHODS: Nine centers enrolled 120 adult patients with recently (500 cases) that participated in a randomized, double blinded study of intracoronary RT completed a questionnaire that included demographics and experience, regulatory issues, scheduling and interaction with patients, time commitment, involvement of the radiation oncologist, and ideas for overcoming hurdles. RESULTS: Licensing was perceived as a substantial hurdle; Nuclear Regulatory Commission approval took more than 5 months at five of 12 sites. At two higher-volume sites, 10-20 procedures were performed per week; 75% of these radiation oncologists did not see the patient prior to the procedure and were not involved in obtaining informed consent. The mean time spent per case was 30-90 minutes; however, there were major concerns about case scheduling (<50% had any input in case scheduling) and after-hours coverage. Radiation oncologists performed fluoroscopy and cineangiography at most centers (92% and 83%); they also performed intracoronary contrast material injections (67%), interpreted intravascular ultrasonographic images (42%), and repositioned the intracoronary RT catheter (33%). CONCLUSION: The authors identify several issues that need to be addressed before intracoronary RT becomes a part of widespread clinical practice. Close collaboration between cardiologists and radiation oncologists at various levels is required to ensure that patients derive maximal benefit from this new technology. PMID- 11110935 TI - Multimodality MR imaging assessment of myocardial viability: combination of first pass and late contrast enhancement to wall motion dynamics and comparison with FDG PET-initial experience. AB - PURPOSE: To combine three magnetic resonance (MR) imaging modalities-dobutamine stress cine, first pass, and late contrast material-enhanced T1-weighted imaging and to compare the results with 2-[fluorine 18]fluoro-2-deoxy-D-glucose (FDG) positron emission tomography (PET) in the assessment of unviable myocardium in coronary artery disease. MATERIALS AND METHODS: Ten patients with multivessel coronary artery disease underwent MR imaging before and 6 months after bypass surgery. Left ventricular cine MR imaging was performed at rest and during dobutamine infusion. Inversion-recovery gradient-echo images were obtained to study myocardial contrast enhancement at first pass and 5 minutes later. FDG PET was performed with orally administered acipimox before surgery. RESULTS: With dobutamine cine MR imaging, unviable myocardium was detected with a sensitivity of 79% and a specificity of 93%; postoperative wall thickening was the standard. First-pass analysis increased these values to 97% and 96%; analysis of late enhancement with T1-weighted imaging, to 62% and 98%. FDG PET had a sensitivity of 81% and a specificity of 86%. CONCLUSION: The combination of first-pass enhancement analysis and wall motion assessment with stress significantly increases the specificity of MR imaging in the detection of unviable sectors. PMID- 11110936 TI - Case 33: a burst fracture of the first lumbar vetebral body. PMID- 11110937 TI - Case 29: gastric trichobezoar and subphrenic abscess. PMID- 11110938 TI - Pretransplantation surveillance for possible hepatocellular carcinoma in patients with cirrhosis: epidemiology and CT-based tumor detection rate in 430 cases with surgical pathologic correlation. AB - PURPOSE: To determine the prevalence of clinically unsuspected hepatocellular carcinoma (HCC) with advanced cirrhosis and assess the sensitivity of helical computed tomographic (CT) surveillance for tumor detection in these patients. MATERIALS AND METHODS: Prospective direct correlation of CT findings with explanted liver specimen findings was performed in 430 transplant recipients with cirrhosis. The prevalence of clinically unsuspected HCC according to liver disease cause was evaluated. Serum alpha-fetoprotein (AFP) values in patients with and those without tumor were recorded. Prospective and retrospective CT tumor detection was evaluated with respect to CT technique and time from CT to transplantation. RESULTS: HCC was found in 59 (14%) of 430 transplant recipients without suspicion of tumor before referral for transplantation. HCC was most prevalent with hepatitis B (27%) and hepatitis C (22%). Serum AFP values were not sensitive for detection of most small tumors. With triphasic helical CT, the prospective and retrospective rates of identifying patients with tumor were 59% and 68%, respectively; the prospective and retrospective tumor nodule detection rates were 37% and 44%, respectively. Tumor detection rates were highest with CT performed within 67 days before transplantation. CONCLUSION: Clinically unsuspected HCC is most prevalent with cirrhosis secondary to hepatitis B or C, and, when evaluated at CT, is best detected with triphasic contrast material enhanced helical imaging performed within 67 days before transplantation. PMID- 11110939 TI - Nontumorous hepatic arterial-portal venous shunts: MR imaging findings. AB - PURPOSE: To determine the magnetic resonance (MR) imaging findings of small nontumorous hepatic arterial-portal venous (arterioportal) shunts in the liver. MATERIALS AND METHODS: MR images in 25 patients with 38 small nontumorous arterioportal shunts verified with surgery or follow-up imaging were included in this study. The causes of arterioportal shunts were iatrogenic causes in 11 patients and/or cirrhotic changes in the remaining patients. Nonenhanced T1- and T2-weighted images and multiphase contrast material-enhanced dynamic images were retrospectively reviewed and compared with conventional hepatic arteriograms to determine the MR characteristics related to the focal hemodynamic changes. RESULTS: On arterial-dominant-phase dynamic MR images, 29 (76%) of the 38 arteriographically suggested nontumorous arterioportal shunts displayed abnormal findings distinguished against the surrounding hepatic parenchyma, including wedge-shaped (n = 14), nodular (n = 9), or irregularly outlined (n = 6) areas of focal contrast enhancement. The signal intensity on nonenhanced T1- and T2 weighted images of the corresponding areas appeared unremarkable except for three wedge-shaped high-signal-intensity areas (three [8%] of 38) on T2-weighted images accompanied by prolonged contrast enhancement. Most (24 [83%] of 29) areas of abnormal signal intensity were located at the periphery of the liver parenchyma. CONCLUSION: A small nontumorous arterioportal shunt should be considered one of the causes of focal parenchymal hyperperfusion abnormalities on contrast-enhanced dynamic MR images of the liver in the absence of abnormal signal intensity on static MR images. PMID- 11110940 TI - Intraductal papillary mucinous tumors of the pancreas: helical CT with histopathologic correlation. AB - PURPOSE: To evaluate the accuracy of preoperative computed tomography (CT) in predicting the location and type of ductal involvement and malignant transformation in intraductal papillary mucinous (IPM) pancreatic tumors and to determine the predictive factors for malignancy at CT. MATERIALS AND METHODS: The helical CT scans obtained in 36 operated on patients with a diagnosis of IPM pancreatic tumor were retrospectively assessed. CT-histopathologic correlation was then performed. RESULTS: The final diagnoses of IPM tumor were combined type (n = 26) and branch duct type (n = 10) lesions. Histologic analysis revealed adenocarcinoma (n = 9), hyperplasia (n = 8), low-grade dysplasia (n = 12), and high-grade dysplasia (n = 7). The lesions were located mainly in the head or uncinate process (n = 20) or were diffuse or multifocal (n = 12). In 12 patients (13 cases), CT-histopathologic correlation was poor, including that in the evaluation of ductal involvement (n = 7), evaluation of lesion location (n = 2), and diagnosis of malignant transformation (n = 4). The most specific predictive signs of malignancy were presence of diabetes and, at CT, a solid mass, main pancreatic duct dilatation greater than 10 mm, diffuse or multifocal involvement, and attenuating or calcified intraluminal content. CONCLUSION: The main causes of poor CT-histopathologic correlation were related to evaluation of main pancreatic duct involvement and diagnosis of malignant transformation. The association between diabetes and specific CT criteria was highly suggestive of malignancy. PMID- 11110941 TI - Utility of intravenously administered contrast material at CT colonography. AB - PURPOSE: To determine if intravenously administered contrast material improves overall reader confidence in the assessment of the colon, large-bowel wall conspicuity, and diagnostic accuracy in the detection of colorectal polyps and cancers at computed tomographic (CT) colonography. MATERIALS AND METHODS: Two hundred patients underwent CT colonography in both supine and prone positions. A five-point scale was used to assess the effect of contrast enhancement on overall reader confidence and bowel wall conspicuity. Eighty-one patients underwent CT colonography with complete colonoscopic or surgical correlation; diagnostic accuracy was compared in 48 patients who received contrast material and 33 who did not. RESULTS: Bowel preparation was ideal in 38 (19%) of 200 patients. Enhanced prone CT images had significantly better scores for reader confidence (4.9 +/- 0.1 vs 4.6 +/- 0.1, P: <.005) and bowel wall conspicuity (4.6 +/- 0.2 vs 4.2 +/- 0.2, P: <.005) compared with those of nonenhanced prone images despite no significant difference in bowel distention (3.8 +/- 0.2 vs 3.9 +/- 0. 1, P: =.8). Enhancement significantly improved the ability to depict medium (6-9-mm) polyps (75% vs 58%, P: <.05). Three large (10-19-mm) polyps were detected only with contrast enhancement; two remained submerged despite dual positioning. CONCLUSION: The use of intravenously administered contrast material significantly improved reader confidence in the assessment of bowel wall conspicuity and the ability of CT colonography to depict medium polyps in suboptimally prepared colons. PMID- 11110942 TI - Evaluation of JPEG and wavelet compression of body CT images for direct digital teleradiologic transmission. AB - PURPOSE: To determine acceptable levels of JPEG (Joint Photographic Experts Group) and wavelet compression for teleradiologic transmission of body computed tomographic (CT) images. MATERIALS AND METHODS: A digital test pattern (Society of Motion Picture and Television Engineers, 512 x 512 matrix) was transmitted after JPEG or wavelet compression by using point-to-point and Web-based teleradiology, respectively. Lossless, 10:1 lossy, and 20:1 lossy ratios were tested. Images were evaluated for high- and low-contrast resolution, sensitivity to small signal differences, and misregistration artifacts. Three independent observers who were blinded to the compression scheme evaluated these image quality measures in 20 clinical cases with similar levels of compression. RESULTS: High-contrast resolution was not diminished with any tested level of JPEG or wavelet compression. With JPEG compression, low-contrast resolution was not lost with 10:1 lossy compression but was lost at 3% modulation with 20:1 lossy compression. With wavelet compression, there was loss of 1% modulation with 10:1 lossy compression and loss of 5% modulation with 20:1 lossy compression. Sensitivity to small signal differences (5% and 95% of the maximal signal) diminished only with 20:1 lossy wavelet compression. With 10:1 lossy compression, misregistration artifacts were mild and were equivalent with JPEG and wavelet compression. Qualitative clinical findings supported these findings. CONCLUSION: Lossy 10:1 compression is suitable for on-call electronic transmission of body CT images as long as original images are subsequently reviewed. PMID- 11110943 TI - Improved US visualization of the pancreatic tail with simethicone, water, and patient rotation. AB - PURPOSE: To evaluate the effect of degassed water, simethicone, and patient rotation on ultrasonographic (US) visualization of the pancreatic tail. MATERIALS AND METHODS: Seventy patients in whom visualization of the pancreatic tail was poor at US were reevaluated in the upright position after ingesting 2 cups (500 mL) of water with 80 mg of simethicone followed by rotating three times on the examination table. In a few patients, the right posterior oblique position was used. Pancreatic tail visualization and disbursement of gastric gas were evaluated. Seventy patients who received 500 mL of distilled water only served as control subjects. RESULTS: Pancreatic tail visualization in patients versus control subjects was complete in 55 (79%) versus five (7%) of 70 patients and control subjects, partial in 10 (14%) versus 38 (54%), and not improved in five (7%) versus 27 (39%). The effect on diminishing gastric air was closely correlated with the degree of improved visualization in most patients. All patients tolerated the procedure well, with no side effects. The technique added a mean of 8 versus 5 minutes to the examination in patients versus control subjects. The full acoustic window effect of the simethicone-water mixture lasted approximately 10 minutes. CONCLUSION: The simethicone-water-rotation technique is simple, safe, inexpensive, and effective for improving pancreatic tail visualization in ambulatory patients and is superior to the use of water alone. PMID- 11110944 TI - Doppler US in patients with crohn disease: vessel density in the diseased bowel reflects disease activity. AB - PURPOSE: To determine if neovascularization associated with Crohn disease, as detected with Doppler ultrasonography (US), reflects clinical disease activity. MATERIALS AND METHODS: A devised measurement, vessel density, was estimated with color Doppler US. Patients with Crohn disease underwent clinical and laboratory assessment in which the Crohn disease activity index was measured; patients underwent abdominal US the same week. Color Doppler US was performed by using a 7.5-10.0- or 8.0-12.0-MHz transducer, the lowest possible pulse repetition frequency without aliasing, a low wall filter, and high Doppler gain settings. The length and thickness of the affected loops were measured, and the number of color Doppler signals per square centimeter in the bowel loop was counted. Pulsed Doppler US was used to confirm that the signals originated from arteries or veins and not from movement artifacts. RESULTS: Ninety-two patients (aged 7-20 years; mean, 14.85 years; 44 female, 48 male) underwent 119 examinations; 85 were performed in patients with active disease. Affected loops were thicker (10.6 vs 4. 6 mm; P: <.001) and had a higher vessel density with disease (69 of 119 examinations) than during remission (two of 34 examinations; P: <.001). CONCLUSION: Vessel density in affected bowel loops, as estimated with Doppler US, and bowel wall thickness (>5 mm) reflect disease activity in patients with Crohn disease. PMID- 11110945 TI - Nonenhanced helical CT and US in the emergency evaluation of patients with renal colic: prospective comparison. AB - PURPOSE: To compare nonenhanced helical computed tomography (CT) with ultrasonography (US) for the depiction of urolithiasis. MATERIALS AND METHODS: During 9 months, 45 patients (mean age, 44 years; mean weight, 92.5 kg) prospectively underwent both nonenhanced helical CT (5-mm collimation; pitch of 1.5) and US of the kidneys, ureters, and bladder. US evaluation included a careful search for ureteral calculi. Presence of calculi and obstruction and incidental diagnoses were recorded. Clinical, surgical, and/or imaging follow-up data were obtained in all patients. The McNemar test was used to compare groups. RESULTS: Diagnoses included 23 ureteral calculi and one each of renal cell carcinoma, appendicitis, ureteropelvic junction obstruction, renal subcapsular hematoma, cholelithiasis, medullary calcinosis, and myelolipoma. CT depicted 22 of 23 ureteral calculi (sensitivity, 96%). US depicted 14 of 23 ureteral calculi (sensitivity, 61%). Differences in sensitivity were statistically significant (P: =.02). Specificity for each technique was 100%. When modalities were compared for the detection of any clinically relevant abnormality (eg, unilateral hydronephrosis and/or urolithiasis in patients with an obstructing calculus), sensitivities of US and CT increased to 92% and 100%, respectively. One case of appendicitis was missed at US, whereas medullary calcinosis and myelolipoma were missed at CT. CONCLUSION: Nonenhanced CT has a higher sensitivity for the detection of ureteral calculi compared with US. PMID- 11110946 TI - Characterization of indeterminate (lipid-poor) adrenal masses: use of washout characteristics at contrast-enhanced CT. AB - PURPOSE: To determine whether computed tomographic (CT) scans and attenuation measurements on contrast material-enhanced and nonenhanced CT scans could be used to characterize adrenal masses, in particular, to characterize these lesions by using adrenal washout characteristics at contrast-enhanced CT. MATERIALS AND METHODS: Eighty-six patients (49 men, 37 women; age range, 29-86 years; mean age, 72 years) with 101 adrenal lesions depicted at contrast-enhanced CT underwent delayed (mean, 9 minutes) enhanced scanning. Seventy-eight patients also underwent nonenhanced CT. Mean diameter of the benign lesions was 2.1 cm (range, 1.0-4.2 cm); mean diameter of the malignant lesions was 2.3 cm (range, 1.0-4.1 cm). Region-of-interest measurements were obtained at nonenhanced, dynamic enhanced, and delayed enhanced CT and were used to calculate a relative percentage washout as follows: 1 - (Hounsfield unit measurement on delayed image / Hounsfield unit measurement on dynamic image) x 100%. RESULTS: Ninety-nine of 101 lesions were correctly characterized as benign or malignant with a relative percentage washout threshold of 50% on delayed scans; benign lesions demonstrated more than 50% washout; and malignant lesions, less than 50% washout. Two benign lesions demonstrating less than 50% washout were characterized as benign by using conventional CT. CONCLUSION: Calculation of relative percentage washout on dynamic and delayed enhanced CT scans may lead to a highly specific test for adrenal lesion characterization, reduce the need for, and possibly obviate, follow-up imaging or biopsy. PMID- 11110947 TI - US characterization of ovarian masses: a meta-analysis. AB - PURPOSE: To compare the effectiveness of current ultrasonographic (US) techniques for characterizing ovarian masses. MATERIALS AND METHODS: Through a MEDLINE literature search, articles with imaging-histopathologic correlation and data that allowed calculation of contingency tables were identified. Results of morphologic assessment, Doppler US, color Doppler flow imaging, and combined techniques were compared. RESULTS: Among 89 data sets from 46 included studies (5,159 subjects), 35 sets used morphologic information, 36 measured Doppler US indexes, 10 assessed tumor vascularity with color Doppler flow imaging, and eight used combined techniques. Summary receiver operating characteristic curves revealed significantly higher performance for combined techniques than for morphologic information (P: =.003), Doppler US indexes (P: =.003), or color Doppler flow imaging alone (P: =.001). The Q* point (and 95% CI) for combined techniques was 0.92 (0.87, 0.96) versus 0. 85 (0.83, 0.88) for morphology, 0.82 (0.78, 0.86) for Doppler US, and 0.73 (0.58, 0.87) for color Doppler flow imaging. Morphologic assessment showed a trend toward better performance than color Doppler flow imaging (P: =.09) or Doppler US indexes (P: =.07). Doppler US index results were better in earlier studies (P: =.005). CONCLUSION: Combined US techniques and a diagnostic algorithm perform significantly better than morphologic assessment, color Doppler flow imaging, or Doppler US indexes alone in characterizing ovarian masses. PMID- 11110948 TI - Captopril MR renography in a swine model: toward a comprehensive evaluation of renal arterial stenosis. AB - PURPOSE: To test the feasibility of captopril magnetic resonance (MR) renography and to validate the technique in an animal model of renal arterial stenosis. MATERIALS AND METHODS: Seven pigs with induced renal arterial stenosis were studied. MR renography was performed with a T1-weighted approach by using three dimensional fast imaging with steady-state precession, or FISP, sequences after administration of a bolus of 0.1 mmol of gadopentetate dimeglumine per kilogram of body weight. Captopril was administered to improve the specificity. RESULTS: The results demonstrate that differences in renographic curves and indices are observed only if an anatomically substantial stenosis, typically a diameter reduction of more than 70%, is present and captopril is administered. CONCLUSION: In this preliminary experience in an animal model, captopril MR renography provided data consistent with expectations based on conventional renographic results. PMID- 11110949 TI - Nephrotoxic nephritis and obstructive nephropathy: evaluation with MR imaging enhanced with ultrasmall superparamagnetic iron oxide-preliminary findings in a rat model. AB - PURPOSE: To evaluate the role of magnetic resonance (MR) imaging enhanced with ultrasmall superparamagnetic iron oxide (USPIO) in the evaluation and differentiation of different types of nephropathies. MATERIALS AND METHODS: Two experimental rat models of nephropathies were studied: a model of nephrotoxic nephritis induced by means of intravenous injection of sheep anti-rat glomerular basement membrane serum (n = 43) and a model of obstructive nephropathy (n = 6). Imaging sessions were performed with a spectrometer operating at 4.7 T with fast low-angle shot, or FLASH, sequences. Signal intensity was measured in each kidney compartment before and 24 hours after intravenous injection of USPIO (90 micromol of iron per kilogram of body weight). MR findings were compared with histologic data and urine protein levels. RESULTS: In the nephrotoxic nephritis model 24 hours after injection of USPIO, a significant signal intensity decrease (P: <.05) was present only in the cortex where the glomerular lesions were located. In the obstructive nephropathy model, the signal intensity decrease (P: <.05) was located in all kidney compartments in response to diffuse interstitial lesions. The decrease in signal intensity was due to iron uptake by either macrophages or mesangial cells gaining endocytic activity and was correlated, in the nephrotoxic nephritis model, to the degree of proteinuria. CONCLUSION: Twenty-four-hour delayed USPIO-enhanced MR imaging may help identify and differentiate various types of nephropathies. PMID- 11110950 TI - Percutaneous tumor ablation: increased coagulation by combining radio-frequency ablation and ethanol instillation in a rat breast tumor model. AB - PURPOSE: To determine if percutaneously applied radio frequency (RF) combined with percutaneous ethanol instillation (PEI) can increase the extent of ablation in rat breast tumors. MATERIALS AND METHODS: R3230 mammary adenocarcinoma was implanted bilaterally in the mammary fat pads of 18 female rats. The tumor nodules measured 1. 2-1.5 cm. Eight tumors each were treated with (a) conventional, monopolar RF (96 mA +/- 28; 70 degrees C for 5 minutes); (b) PEI (250 microL of ethanol infused over 1 minute); (c) combined therapy of PEI immediately followed by RF ablation; or (d) combined therapy of RF ablation immediately followed by PEI. Four tumors were not treated and served as controls. Histopathologic examination included staining for mitochondrial enzyme activity. Resultant coagulation necrosis was compared between treatment groups. RESULTS: Coagulation necrosis was observed only within treated tumors. Tumors treated with RF alone had 6.7 mm +/- 0.6 of coagulation surrounding the electrode, and those treated with PEI alone had 6.4 mm +/- 0.6 of coagulation around the instillation needle (not significant). Significantly increased coagulation of 10.1 mm +/- 0.9 (P: <.001) was observed with the combined therapy of PEI followed by RF. RF followed by PEI did not increase coagulation (6.4 mm +/- 0.8 around the needle; not significant). CONCLUSION: PEI followed by RF ablation therapy increases the extent of induced coagulation necrosis in rat breast tumors, as compared with either therapy alone. PMID- 11110951 TI - Mammography in 53,803 women from the New Hampshire mammography network. AB - PURPOSE: To describe measures of mammography performance in a geographically defined population and evaluate the interpreter's use of the Breast Imaging Reporting and Data System (BI-RADS). MATERIALS AND METHODS: Mammographic data from 47,651 screening and 6,152 diagnostic examinations from November 1, 1996, to October 31, 1997, were linked to 1,572 pathologic results. Mammographic outcomes were based on BI-RADS assessments and recommendations reported by the interpreting radiologist. The consistency of BI-RADS recommendations was evaluated. RESULTS: Screening mammography had a sensitivity of 72.4% (95% CI: 66.4%, 78.4%), specificity of 97.3% (95% CI: 97.25%, 97.4%), and positive predictive value of 10.6% (95% CI: 9.1%, 12.2%). Diagnostic mammography had higher sensitivity, 78.1% (95% CI: 71.9%, 84.3%); lower specificity, 89.3% (95% CI: 88.5%, 90.1%); and better positive predictive value, 17.1% (95% CI: 14.5%, 19.8%). The cancer detection rate with screening mammography was 3.3 per 1,000 women, with a biopsy yield of 22.4%, whereas the interval cancer rate was 1. 2 per 1,000. Nearly 80% of screening-detected invasive malignancies were node negative. The recall rate for screening mammography was 8. 3%. Ultrasonography was used in 3.5% of screening and 17.5% of diagnostic examinations. BI-RADS recommendations were generally consistent, except for probably benign assessments. CONCLUSION: The sensitivity of screening mammography in this population-based sample is lower than expected, although other performance indicators are commendable. BI-RADS "probably benign" assessments are commonly misused. PMID- 11110952 TI - Breast masses with peripheral rim enhancement on dynamic contrast-enhanced MR images: correlation of MR findings with histologic features and expression of growth factors. AB - PURPOSE: To investigate the histologic bases of rim enhancement of breast masses demonstrated on dynamic contrast material-enhanced magnetic resonance (MR) images. MATERIALS AND METHODS: Dynamic MR images of breast lesions (invasive carcinoma, n = 29; other, n = 6) in 35 women were reviewed. In each patient, subtraction images of the dynamic contrast-enhanced study were obtained, and early and delayed rim enhancement and delayed internal enhancement were evaluated. The MR findings were correlated with the ratio of microvessel density of the peripheral to the central portion of the lesion, fibrosis, and other histologic features, including expression of vascular endothelial growth factor (VEGF) and transforming growth factor ss1. RESULTS: Early rim enhancement was observed in 29% and delayed rim enhancement was noted in 60% of all patients. Small cancer nests, a high ratio of peripheral-to-central microvessel density, peripheral VEGF expression, and a low ratio of peripheral-to-central fibrosis were correlated with early rim enhancement. Delayed rim enhancement was correlated with a high degree of fibrosis and inflammatory changes. Delayed internal enhancement was correlated with a high degree of fibrosis. CONCLUSION: Rim enhancement of breast lesions at MR imaging is due to a combination of angiogenesis, distribution and degree of fibrosis, expression pattern of VEGF, and various histologic features. PMID- 11110953 TI - US of mammographically detected clustered microcalcifications. AB - PURPOSE: To determine whether ultrasonography (US) can depict breast masses associated with mammographically detected clustered microcalcifications and whether the visibility at US is different between benign and malignant lesions. MATERIALS AND METHODS: Ninety-four patients with 100 mammographically detected microcalcification clusters prospectively underwent US with a 10- or 12-MHz transducer before mammographically guided presurgical hook-wire localization. The visibility of breast masses at US was correlated with histologic and mammographic findings. RESULTS: Surgical biopsy revealed 62 benign lesions, 30 intraductal cancers, and eight invasive cancers. At US, breast masses associated with microcalcifications were seen in 45 (45%) of 100 cases. US depicted more breast masses associated with malignant (31 [82%] of 38) than with benign (14 [23%] of 62) microcalcifications (P: <.001). In malignant microcalcification clusters larger than 10 mm, US depicted associated breast masses in all 25 cases. There was no statistically significant difference in shape and distribution of calcific particles, as well as in breast composition, at mammography between US visible and invisible groups. CONCLUSION: Given a known mammographic location, US with a high-frequency transducer can depict breast masses associated with malignant microcalcifications, particularly clusters larger than 10 mm. US can be used to visualize large clusters of microcalcifications that have a very high suspicion of malignancy. PMID- 11110954 TI - Changes in calcaneal trabecular bone structure after heart transplantation: an MR imaging study. AB - PURPOSE: To use high-spatial-resolution magnetic resonance (MR) imaging to analyze the trabecular bone structure of the calcaneus in patients before and after heart transplantation and to compare this technique with bone mineral density (BMD) measurement in predicting therapy-induced bone loss and vertebral fracture status. MATERIALS AND METHODS: High-spatial-resolution 1.5-T MR imaging of the calcaneus was performed in 40 men 11-120 months after heart transplantation, in 11 men before heart transplantation, and in 10 age-matched male volunteers. Sagittal and transverse T1-weighted spin-echo images with a voxel size of 0.195 x 0.195 x 1.000 mm were obtained, and structure measurements analogous to bone histomorphometric values were calculated. In addition, the BMD of the lumbar spine was determined in the transplant recipients pre- and postoperatively by using quantitative computed tomography, and vertebral fracture status was assessed. RESULTS: Significant differences in structure and BMD measurements were found between patients before and after heart transplantation (P <. 05). In 17 (42%) of 40 transplant recipients, vertebral fractures were found. Although structure measurements were significantly different between patients with and those without fractures (P <.05), BMDs were not. Correlations between time after transplantation and some structure measurements were moderately significant (P <. 05), but such correlations with BMD measurements were not. CONCLUSION: MR imaging-derived structure measurements in the calcaneus are useful for monitoring bone changes after heart transplantation and assessing vertebral fracture status. PMID- 11110955 TI - Reactive bone marrow changes in infectious spondylitis: quantitative assessment with MR imaging. AB - PURPOSE: To evaluate diffuse, reactive bone marrow changes in unaffected vertebrae on magnetic resonance (MR) images in patients with proved infectious spondylitis. MATERIALS AND METHODS: Percentage signal intensity increase of the unaffected bone marrow on contrast material-enhanced MR images (percentage enhancement) was calculated retrospectively in 22 cases of infectious spondylitis and 86 cases without bone marrow disease. Multiple regression analysis and Student t test statistics were performed. RESULTS: Multiple regression analysis showed a significant influence of age and the presence of spondylitis on the values of percentage enhancement (P: <.001). For those aged 35 years or younger, the mean percentage enhancement was 43.2% +/- 4.0 for patients with infectious spondylitis (n = 3) and was 26.4% +/- 8.6 for the control group (n = 23). For those older than 35 years, the mean percentage enhancement was 28.2% +/- 12.2 for patients with infectious spondylitis (n = 19) and 17.5% +/- 7.9 (P: <.001) for the control group (n = 63). Six (27%) of 22 patients with infectious spondylitis showed abnormal percentage enhancement values in unaffected bone marrow when the upper limit of the normal value was 2 SDs above the mean of the control group. CONCLUSION: On MR images, reactive bone marrow changes can be found in unaffected vertebrae in patients with infectious spondylitis. The signal intensity changes and increased percentage enhancement associated with this disease are similar to those of myeloproliferative and diffuse neoplastic disorders and bone marrow stimulation in hemolytic anemia. PMID- 11110956 TI - Cerebrotendinous xanthomatosis: the spectrum of imaging findings and the correlation with neuropathologic findings. AB - PURPOSE: To describe imaging findings and their neuropathologic correlate in patients with cerebrotendinous xanthomatosis (CTX). MATERIALS AND METHODS: Computed tomographic (CT) and magnetic resonance (MR) images in 24 patients with symptoms (mean age at time of imaging, 37 years; mean disease duration, 18 years) were reviewed for site and frequency of brain, spinal cord, and Achilles tendon involvement. Two patients died, and imaging findings were compared with postmortem neuropathologic findings. RESULTS: Apart from nonspecific supratentorial atrophy and deep white matter changes, more typical hyperintense lesions were seen on T2-weighted images in the dentate nucleus (in 79% of patients), globus pallidus, substantia nigra, and inferior olive and extended into adjacent white matter as disease progressed. In these locations, lipid crystal clefts and perivascular macrophages, neuronal loss, demyelination, fibrosis, and reactive astrocytosis were found at microscopic examination. Hypointensity was sometimes found on T2-weighted images in the dentate nucleus and was related to deposition of hemosiderin and calcifications. CT depicted fewer lesions; all had low attenuation, except for the calcifications. Spinal cord MR imaging revealed increased signal intensity in the lateral and dorsal columns on T2-weighted images. Achilles tendon xanthomas displayed intermediate signal intensity on T1- and T2-weighted images. CONCLUSION: The typical pattern of MR imaging findings reflects the classic histopathologic findings and should prompt the diagnosis of CTX. PMID- 11110957 TI - MR-Intracranial pressure (ICP): a method to measure intracranial elastance and pressure noninvasively by means of MR imaging: baboon and human study. AB - PURPOSE: To develop a noninvasive method for intracranial elastance and intracranial pressure (ICP) measurement. MATERIALS AND METHODS: Intracranial volume and pressure changes were calculated from magnetic resonance (MR) imaging measurements of cerebrospinal fluid (CSF) and blood flow. The volume change was calculated from the net transcranial CSF and blood volumetric flow rates. The change in pressure was derived from the change in the CSF pressure gradient calculated from CSF velocity. An elastance index was derived from the ratio of pressure to volume change. The reproducibility of the elastance index measurement was established from four to five measurements in five healthy volunteers. The elastance index was measured and compared with invasive ICP measurements in five patients with an intraventricular catheter at MR imaging. False-positive and false-negative rates were established by using 25 measurements in eight healthy volunteers and six in four patients with chronically elevated ICP. RESULTS: The mean of the fractional SD of the elastance index in humans was 19.6%. The elastance index in the five patients with intraventricular catheters correlated well with the invasively measured ICP (R:(2) = 0.965; P: <.005). MR imaging derived ICPs in the eight healthy volunteers were 4.2-12.4 mm Hg, all within normal range. Measurements in three of the four patients with chronically elevated ICP were 20.5-34.0 mm Hg, substantially higher than the normal limit. CONCLUSION: MR imaging-derived elastance index correlates with ICP over a wide range of ICP values. The sensitivity of the technique allows differentiation between normal and elevated ICP. PMID- 11110958 TI - Combined perfusion- and diffusion-weighted MR imaging in acute ischemic stroke during the 1st week: a longitudinal study. AB - PURPOSE: To compare findings with different magnetic resonance (MR) perfusion maps in acute ischemic stroke. MATERIALS AND METHODS: Combined diffusion-weighted (DW) and perfusion-weighted (PW) MR imaging was performed in 49 patients with acute (<24 hours) stroke, on the 1st and 2nd days and 1 week after stroke. Volumes of hypoperfused tissue on maps of relative cerebral blood volume (rCBV), relative cerebral blood flow (rCBF), and mean transit time (MTT) were compared with the volume of infarcted tissue at DW imaging. RESULTS: The mean infarct volume increased from 41 to 65 cm(3) between the 1st and 2nd days (P: <.001; n = 49). On the 1st day, all perfusion maps on average showed hypoperfusion lesions larger than the infarct at DW imaging (P: <.001; n = 49). MTT maps showed significantly (P: <.001) larger hypoperfusion lesions than did rCBF maps, which showed significantly (P: <.001) larger hypoperfusion lesions than did rCBV maps. The sizes of the initial perfusion-diffusion mismatches correlated significantly with the extent of infarct growth (0.479 < r < 0.657; P: C variant in the 3'-flanking region of AT(1)R gene with essential hypertension in Tibetans]. AB - OBJECTIVE: To investigate whether CA-repeat polymorphism and A1166 --> C variant in the 3n-flanking region of AT(1)R gene are in association with the genetic susceptibily to essential hypertension (EH) in Tibetans. METHODS: A case-control study was carried out. Sibpair analysis and family linkage analysis were conducted. The CA-repeat polymorphism of AT &(1) R gene was identified by polymerase chain reaction(PCR) with fluorescence labeled dCTP as substrate and by semi-automatic sequence technology. The A1166 -->C variant was detected by PCR RFLP. RESULTS: Association of AT&(1)R gene locus with EH was confirmed through the case-control study in well-characterized group of 113 Tibetan EH patients and 131 normotensives(chi(2)=26.44, P<0.001). A closer examination of this gene locus found 11 alleles from Tibetan population; allele A7 (138 bp) was more frequent in both the patients and the controls. Allele A8(140 bp) was in strong positive association with genetic susceptibility to EH in Tibetans. Frequency of allele A8 was 20.5% in EH and 7.3% in normotensives. The difference of allele frequencies between the groups was significant (chi(2)=9.64, P=0.002, OR=3.46, 95% CI 1.44 8.51). Affected sibpair analysis showed chi(2)=3.85, P=0.025; family linkage analysis gave Lod score of 0.80. No association between A1166 --> C variant in AT(1)R gene and EH in Tibetans was observed (P>0.05). CONCLUSION: The result suggests that CA-repeat polymorphism of AT(1)R gene be in association with EH in Tibetans, which implicates that AT(1)R gene may be in linkage disequilibrium with the causative genes of EH. PMID- 11110973 TI - [Chromosome localization of the dentinogenesis imperfecta type II locus]. AB - OBJECTIVE: To scrutinize the linkage between dentinogenesis imperfecta type II and chromosome 4q21 in a Tianjin-Tanggu family. METHODS: Blood samples were collected from 13 members of the family. DNA was analyzed with 4 short tandem repeat polymorphisms markers UGATA62A11, DSP(P), SPP1 and D4S1563 Y using fluorescence-based PCR. The linkage between four markers on chromosome 4q21 and dentinogenesis imperfecta type II was tested by Lod score analysis. RESULTS: GATA62A11 and DSP(P) suggested linkage and yielded a Lod score of 1.63 at theta =0, and 1.68 at theta =0 by means of the MLINK software, respectively. Genotype and haplotype were acquired. CONCLUSION: The disease gene of the dentinogenesis imperfecta type II family is located on chromosome 4q. The result will be helpful for the further identification of the dentinogenesis imperfecta type II gene. PMID- 11110974 TI - [Analysis of the interaction of the polymorphisms of presenilin-1 gene and ApoE gene in Alzheimerns disease]. AB - OBJECTIVE: To explore the interaction of the polymorphism of 3' -terminal intron of the eighth exon of presenilin-1(PS-1) gene with the polymorphism of apolipoprotein E(ApoE) gene in the pathogenesis of sporadic Alzheimerns disease(SAD). METHODS: The polymorphisms of 3'-terminal intron of the eighth exon of presenilin-1 gene and ApoE gene were detected by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique in 75 patients with SAD and 73 normal controls. RESULTS: The polymorphisms of PS-1 gene and ApoE gene were associated obviously with SAD. The onset of SAD was negatively associated with allele 2 of PS-1 gene and E(2) allele of ApoE gene,and it was positively associated with allele 1 of PS-1 gene. The onset of SAD was not associated with E(3) allele and E(4) allele of ApoE gene. CONCLUSION: These results indicated that allele 2 of PS-1 gene and E(2) allele of ApoE gene might protect people from SAD, that allele 1 of PS-1 gene might increase the risk of the onset of SAD, and that the onset of SAD was not associated with the interaction of the polymorphisms of PS-1 gene and ApoE gene. PMID- 11110975 TI - [In vitro study on transduction of human O(6)-methylguanine-DNA-methyltransferase cDNA into human umbilical cord blood CD34(+) cells]. AB - OBJECTIVE: To explore human umbilical cord blood hematopoietic progenitor cells transduced with human O(6)-methylguanine-DNA-methyltransferase (MGMT) gene increase resistance to 1,3-Bis(2-Chloroethyl)-1-Nitrosourea(BCNU). METHODS: The present authors obtained a full length cDNA fragment encoding the human MGMT from a patient with cholelithiasis liver tissue by RT-PCR method and confirmed by DNA sequencing. The fragment was cloned into pGEM-T vector and further subcloned into G1Na retrovirus vector. Then the G1Na-MGMT was transfected into the packaging cell lines GP+E86 and PA317 by LipofectAMINE method; using the medium containing BCNU for cloning selection and ping-ponging supernatant infection between ecotropic producer clone and amphotropic producer clone, the authors obtained high titer amphotropic PA317 producer clone with the highest titer up to 1.6x10(6) CFU/ml. Cord blood CD34(+) cell were transfected repeatedly with supernatant of retrovirus containing human MGMT cDNA under stimulation of hemopoietic growth factors. RESULTS: PCR, RT-PCR, Southern blot, Northern blot, Western blot and MTT analyses showed that MGMT gene had been integrated into the genomic DNA of cord blood CD34(+) cells and expressed efficiently in the transfected cells. The transgene recipient cells conferred 4 folds stronger resistance to BCNU than that of the non-transduced. CONCLUSION: The retrovirus vector-mediated transfer of MGMT drug resistance gene into human cord blood CD34(+) cells and expression could confer the resistance of transgene cells to BCNU toxicity. PMID- 11110976 TI - [Study on the relationship of alteration and expression of p16 gene to pancreatic carcinoma]. AB - OBJECTIVE: To directly investigate the effect of genetic alteration(homozygous deletion and point mutation) and expression of p16 gene on pancreatic carcinomas. METHODS: Thirty-five cases were analyzed for genetic alteration and expression of p16 gene by polymerase chain reaction(PCR), single strand conformation polymorphism(SSCP), DNA sequencing and immunohistochemical method. RESULTS: The analysis of pancreatic carcinoma for p16 gene revealed alteration in 19 of 35 cases, among which 12 pancreatic carcinomas had 522 bp homozygous deletion at least and 7 cases had two point mutations at the same site. One of them is 126th codon GTC --> AAT (Val126Asn); the other is 127th codon GCA --> GCG (A127A). The 3D (three-dimensional) structure of P16 protein determined by computer techniques according to PDB indicated that Val126Asn influenced the space structure of P16 protein and affected the function of P16 protein. Twelve cases revealed no P16 protein and 9 cases showed low level expression of P16 protein. CONCLUSION: The alteration of p16 gene and abnormal expression of P16 protein are significantly correlated with the biological behavior and clinical staging of pancreatic carcinoma and may hence be helpful to prognostication PMID- 11110977 TI - [Study on a downstream signal molecule of human CASK/LIN-2]. AB - OBJECTIVE: To elucidate the function of human CASK/LIN-2, a novel member of membrane-associated guanylate kinase homologs family (MAGUK), by using the yeast two-hybrid system(LexA) to screen the protein interacting with the guanylate kinase-like domain(GK) of hCASK and identify possible protein that might involve downstream signal transduction of hCASK. METHODS: PCR strategy was used to amplify GK domain cDNA fragment of hCASK. The DNA was subcloned into pLexA to construct bait vector pLexA-GK. The pLexA-GK was transformed into yeast host strain EGY48; through testing the bait protein and a repression assay, it was demonstrated that the bait protein did not have transcription activation for leu and lacZ reporter genes, however it could enter nucleus, bind LexA operator. Human fetus brain cDNA library plasmids were transformed into EGY48 containing pLexA-GK and p8op-lacZ, and screened on plates of Gal/Raf his- trp- ura- leu- and Gal/Raf his- trp- ura-X-gal. The clones with galactose dependent leu+ and lacZ+ were isolated; their library plasmids were rescued by means of transforming E.coli KC8. These library plasmids were transformed into the yeast again to re screen. The remaining positive clones were analyzed by PCR and restriction endonuclease. Finally two kinds of target DNA fragments were obtained. RESULTS: The specificity testing indicated that the two target proteins could specially bind GK domain of hCASK. After DNA sequencing and BLASTn analysis on NCBI, it was shown that the two target DNA fragments, 732 and 683 bp, had high sequence similarity with human inhibitors of differentiation 1(Id1) mRNA, with identities of 97%(630/645) and 98%(656/666), respectively. CONCLUSION: GK domain of hCASK could specially interact with Id1 in yeast. Id1 might be a downstream signal molecule of hCASK, which might involve in regulation of cell differentiation. PMID- 11110978 TI - [The association and linkage analysis between the FcgammaR II a-131 and system lupus erythematosus]. AB - OBJECTIVE: To shed light on the relationship between FcgammaR II a-131 and systemic lupus erythematosus(SLE) in southern Chinese Han population. METHODS: A population-based and family-based study was carried out. FcgammaR II a-131 of each subject was measured by using PCR-allele specific oligonucleotide hybridization(ASO) method. RESULTS: (1) The distribution of FcgammaR II A-131 genotype in cases is significantly different from that in controls (P<0.05). So is the frequency of FcgammaR II aR-131 allele (P < 0.01) which suggests that subjects who have R131 allele tend to be more susceptible to SLE. The subjects with R/R131 homozygous genotype have a higher risk of suffering from SLE. (2) The distribution of FcgammaR II a-131 varies in different races, with identical distribution type among Chinese and Japanese. (3) The results of family-based association analysis and transmitted/disequilibrium test(TDT) suggest that there is not any linkage evidence between FcgammaR II a-131 and SLE. Possibly, the sample size was too small to get positive result. CONCLUSION: This study suggests that FcgammaR II a-131 is a major factor predisposing to the development of SLE in southern Chinese Han population. PMID- 11110979 TI - [Is Gly460Trp variant of alpha-adducin associated with essential hypertension in the Hans of Chinese population]. AB - OBJECTIVE: To determine whether the variant of adducin alpha subunit (alpha adducin) is in association with essential hypertension in the Hans of Chinese population. METHODS: One hundred and eighty-three patients with essential hypertension and 129 normotensive subjects were included in this study. Genomic DNA was isolated from 3 ml of whole blood. Polymerase chain reaction, single strand conformational polymorphism and DNA sequencing analysis techniques were used to detect the variant in the exon 10 of alpha-adducin gene. The clinical data of height, weight and blood pressure were also measured in all subjects. RESULTS: A G-->T substitution at nucleotide 614 in exon 10 of alpha-adducin gene which resulted in the Gly460Trp variant was found in the Hans. The distribution of Gly/Gly, Gly/Trp and Trp/Trp genotypes was coincident with Hardy-Weinberg equilibrium and there were no statistical differences in the three genotypes and Trp allele frequencies between hypertensive and normotensive subjects (P>0.05). The variant of alpha-adducin genotypes was not associated with any significant differences in age, body mass index and blood pressure. CONCLUSION: A G-->T substitution at nucleotide 614 in exon 10 of alpha-adducin gene is found, but there is lack of the association between the variant of alpha-adducin gene and essential hypertension in the Hans of Chinese population. PMID- 11110980 TI - [Genetic polymorphism of 10 STR loci on chromosome 9 and its forensic application in Chinese population]. AB - OBJECTIVE: To have a thorough knowledge of the genetic polymorphism of 10 short tandem repeats loci of chromosome 9 in Chinese population and its application in forensic science. METHODS: The 9 STR tetrameric loci and one trimeric tandem repeat locus were chosen from Genome Databank. Eighty-three EDTA-blood samples were collected from the unrelated family in Baiyin city, Gansu province. DNA was extracted by Chelex method and amplified by the polymerase chain reaction(PCR). The PCR products were analyzed by PAGE electrophoresis. RESULTS: The polymorphisms of all 10 STR loci have been obtained in Chinese Han population. The 10 STR loci follow the Mendel ns law. CONCLUSION: The data of all 10 STR loci is beneficial to understanding the population genetics of them in Chinese Han population. PMID- 11110981 TI - [The study on hereditary polymorphism of thiopurine S-methyltransferasein Chinese Han population of Shanghai area]. AB - OBJECTIVE: To gain an insight into the hereditary polymorphism of thiopurine methyltransferase(TPMT) activity in Chinese Han population of Shanghai. METHODS: The present authors measured the erythrocyte TPMT activity in 320 healthy Chinese volunteers and 51 children with acute lymphoblastic leukemia(ALL) by means of radiochemical assay. RESULTS: The TPMT activity levels ranged from 4. 32 to 32.33 U/ml pRBCs, with a mean value of (16.64+/-4.50) U/ml pRBCs, male (16.78+/-4.96) U/ml pRBCs, female (16.52+/-4.44) U/ml pRBCs. Eight point one percent of the sample had low activity. The TPMT activity levels for subjects who were <12 years, 13-18 years, 19-45 years and >45 years old were (16.52+/-4.31) U/ml pRBCs, (16. 71+/-4.24) U/ml pRBCs, (16.28+/-5.21) U/ml pRBCs and (17.11+/-3.98) U/ml pRBCs, respectively; the TPMT activity levels for healthy volunteers and patients with ALL were (16.65+/-4.72) U/ml pRBCs and (16.52+/-4.47) U/ml pRBCs, respectively. CONCLUSION: There were no differences of TPMT activity in gender, age, and between healthy volunteers and patients with ALL. PMID- 11110982 TI - [Studies on the polymorphism of MICA gene in four Chinese populations]. AB - OBJECTIVE: To understand with greater clearness the genetic polymorphism of (GCT)n repeat of MHC class I chain-related gene A (MICA)in some Chinese populations and provide preliminary genetic evidence for the independent origin of Chinese Baima Tibetan (BMT). METHODS: ACD-blood or saliva specimens of 411 unrelated individuals from four Chinese populations were collected. A primer pair spanning exon 5 of MICA gene was used to amplify the GCT region. Alleles were detected by PCR and denaturing PAGE. Comparison of the allelic distributions among the four populations was carried out. RESULTS: Five previously reported alleles have been observed in all the four populations, but the allelic distributions are different from one another. The most frequent allele is the A5 in all the four populations (0.325 in BMT, 0.345 in Tibetans, 0.390 in Chengdu Hans and 0.319 in Qiangs). A5.1 allele is the second most frequent allele in Chengdu Hans (0.230) and in Qiangs (0.293), while the second most frequent alleles for BMT and Tibetans are A4 (0.254) and A9 (0.272) respectively. The distribution of alleles in BMT is significantly different from that in the other three populations. CONCLUSION: Alleles of MICA gene exon 5 are conservative in all populations studied so far. The results suggest that genetically BMT might be an independent ethnic population. PMID- 11110983 TI - [Methods for microdissection-PCR-silver stain technique in paraffin-embedded tissue sections]. AB - OBJECTIVE: To use the microdissection-PCR-silver stain technique in the paraffin- embedded tissue sections and analyze the microsatellite instability in colorectal cancer. METHODS: By microdissection technique, the lymphocytes and mesenchymal cells, normal mucous epithelial cells, displasia, adenoma, and carcinoma cells were recovered from paraffin-embedded tissue sections. The cells were digested by protinase K; the cell lysates were used as PCR template and the PCR products were analysed by denatured polyacrylamide gel electrophoresis and silver stain. Four microsatellite loci, TGF-betaRII(A)(10), hMSH(2)(A)(26)-Bat-26, hMSH(3)(A)(8), hMSH(6)(C)(8), were analyzed. RESULTS: In all 28 specimens, hMSH(3)(A)(8) and hMSH(6)(C) 8) loci were amplified successfully. TGF-betaRII(A)10), Bat-26 had consistent amplification in 23/28, 22/28 specimens, respectively. And Bat-26 microsatellite instability was found in carcinoma cells in 5 specimens, and in adenoma cells in one of the 5 specimens. CONCLUSION: Microdissection-PCR silver stain technique can be used in paraffin-embedded tissue sections with satisfactory results. This method can be employed in analyzing microsatellite instability in colorectal carcinoma. PMID- 11110984 TI - [ABO genotyping by PCR-direct sequencing method]. AB - OBJECTIVE: To analyze the sequence difference between human A, B, and O alleles and establish the method of ABO genotyping by PCR direct sequencing. METHODS: PCR direct sequencing technique was used to analyze two regions of cDNA from A transferase gene, 233-433 and 660-788. RESULTS: Two nucleotide substitutions at 258th and 297th were found in 233-433 region, and a nucleotide substitution at 700th was found in 660-788 region. At 258th, the nucleotide was guanine in A and B alleles, and adenine in O allele. At 297th, the nucleotide was adenine in A allele, and guanine in B allele. As this position, O allele was subdivided into two types, O(A) and O(G). At 700th, the nucleotide was guanine in A and O alleles, and adenine in B allele. Therefore, 8 genotypes, AA, AO(A), AB, BB, BO(G), O(A) O(A), O(G) O(G) and O(A) O(G), could be clearly determined by only analyzing the 233-433 region. The other two genotypes, AO(G) and BO(A), could be further distinguished by analyzing the 660-788 region. CONCLUSION: The technique of PCR-direct sequencing provides an effective and new method for ABO genotyping further. PMID- 11110985 TI - [The application of competitive RT-PCR to detect minimal residual disease in patients with 8;21 translocation]. AB - OBJECTIVE: To construct and evaluate the competitive quantitative RT-PCR method for detecting the fusion gene of AML1-ETO in the patients of t(8;21)AML(acute myeloid leukemia). METHODS: The authors used the method of splicing by overlapping extension (SOE) to obtain the competitive DNA fragment, with which they set up the competitive quantitative RT-PCR assay to detect AML1-ETO chimeric genes in the patients of t(8;21) AML. RESULTS: The competitive DNA fragment has been obtained; the sensitive competitive RT-PCR method is developed, which allows the quantitation of the number of AML1-ETO transcriptions in t(8;21)AML patients at different phases. CONCLUSION: The competitive quantitative RT-PCR method, based on SOE, is simple and convenient; t(8;21) AML patients in different survival conditions express AML1-ETO gene quite differently. PMID- 11110986 TI - [Progress in the studies on the molecular genetics of schizophrenia]. AB - Although population genetic studies have long confirmed the genetic vulnerability of schizophrenia,ongoing advances in molecular genetic technology and biostatistic analysis are only now making it possible to search for the susceptibility gene of the disease. This article reviewed some of the recent findings in this area: (1) The heritability of schizophrenia is estimated around 60%-80%. The phenotype differentiation is based on standard diagnostic scales and symptom rating scales. (2) The two main approaches to finding the genes that influence the disorder are now genomic scan and candidate gene detection. Affected sib-pair (ASP) method and transmission disequilibrium test(TDT) are considered promising analyses. (3) The positive candidate regions with some independent replicable reports concentrated on 6p, 22q and 8p. Positive findings of candidate gene research involved 5-HT2A receptor, DRD3, NT-3, etc. Further directions to identify the susceptibility genes include: Applying more precise instruments to define clinical phenotype of the disease. Application of proper biological markers such as electrophysiologic parameters and brain imaging will be a prospective approach. Using larger sample to increase statistic power and developing more powerful statistic analysis, and performing advanced molecular genetic technique such as DNA pooling, DNA chips, genomic mismatch scanning (GMS), representational difference analysis(RDA), comparative genomic hybridization(CGH) and two-dimensional DNA typing methods will also facilitate this research area to greater perspective. PMID- 11110987 TI - Marriage in eating disorders comparisons between patients and spouses and changes over the course of treatment. AB - OBJECTIVES: This study examines one aspect of the marital satisfaction, intimacy, of couples where one member has an eating disorder (ED), before and after intensive day hospital treatment for ED. METHODS: Subjects were consecutive patients (n=22) attending a day hospital program for EDs and their spouses. The Waring Intimacy Questionnaire (WIQ) was administered to the couples at admission to and discharge from the treatment program. RESULTS: Patients showed less favorable ratings of the marriage at admission and discharge, compared to spouses, but patient's ratings improved significantly over the course of treatment. Patients generally improved in terms of their ED symptoms during the treatment. Spousal ratings showed satisfactory ratings of intimacy at the start of treatment and did not change over the course of treatment. CONCLUSION: The self-reported marital dissatisfaction of patients with an ED is at least partially alleviated after symptomatic treatment of the ED. Treatment of the ED in one member does not diminish marital satisfaction in the other member of the couple. The long-term prognosis for these couples remains unknown. PMID- 11110988 TI - Epidemiological evidence for a relationship between life events, coping style, and personality factors in the development of breast cancer. AB - OBJECTIVE: Review empirical evidence for a relationship between psychosocial factors and breast cancer development. METHODS: Standardised quality assessment criteria were utilised to assess the evidence of psychosocial predictors of breast cancer development in the following domains: (a) stressful life events, (b) coping style, (c) social support, and (d) emotional and personality factors. RESULTS: Few well-designed studies report any association between life events and breast cancer, the exception being two small studies using the Life Events and Difficulties Schedule (LEDS) reporting an association between severely threatening events and breast cancer risk. Seven studies show anger repression or alexithymia are predictors, the strongest evidence suggesting younger women are at increased risk. There is no evidence that social support, chronic anxiety, or depression affects breast cancer development. With the exception of rationality/anti-emotionality, personality factors do not predict breast cancer risk. CONCLUSION: The evidence for a relationship between psychosocial factors and breast cancer is weak. The strongest predictors are emotional repression and severe life events. Future research would benefit from theoretical grounding and greater methodological rigour. Recommendations are given. PMID- 11110989 TI - Anemia and macrocytosis in the prediction of serum folate and vitamin B12 status, and treatment outcome in major depression. AB - BACKGROUND: Folate and B12 deficiencies may result in macrocytic anemia, and are common in major depression; hypofolatemia may result in poorer antidepressant response. We wished to determine whether anemia or macrocytosis predict hypofolatemia, low B12, or refractoriness to antidepressants. METHODS: After obtaining serum folate, B12, and hematological indices, 213 depressed adults were treated with fluoxetine 20 mg/day. Amelioration of depressive symptoms was measured. RESULTS: Neither macrocytosis nor anemia predicted low serum folate/B12, or antidepressant refractoriness. Among 39 patients with hypofolatemia, none had macrocytosis; 28% had low HCT; 41% had low RBC. Among 25 patients with low B12, none had macrocytosis; 24% had low HCT; 28% had low RBC. Among non-responders, 3% had macrocytosis; 24% had low HCT; 25% had low RBC. CONCLUSION: Anemia and macrocytosis should not be used to predict folate or B12 deficiencies, or refractoriness to antidepressants. Measurement of folate and B12 should be considered when evaluating treatment refractoriness. PMID- 11110990 TI - Body image and body change methods in adolescent boys. Role of parents, friends and the media. AB - OBJECTIVE: This study examines sociocultural influences affecting both body image and body change methods in adolescent boys. METHODS: Twenty boys in grade 7 (aged 12-13) and twenty boys in grade 9 (aged 14-15) were individually interviewed. The influence of parents, siblings, friends and the media on both body image and body change methods was evaluated. RESULTS: For approximately a third of the boys, parents, siblings, friends and the media were perceived to have at least some influence over boys' feelings about their bodies and body change methods. In particular, feedback from mothers and female friends were viewed as having a positive impact on boys' body image whereas feed-back from fathers and male friends was viewed as more important in influencing body change methods. The media was also viewed as contributing to boys' body satisfaction but it was seen to encourage greater exercise to alter body size and shape. CONCLUSION: The differences and similarities between the sociocultural messages received by males and females are discussed. The implication of these findings in fostering better health among adolescent males are explored. PMID- 11110991 TI - Predictors of eating disorder scores in children ages 6 through 14: a longitudinal study. AB - The objective of this study was to identify variables that predict higher eating disorder scores in non-clinical boys and girls ages 6 through 14. Two hundred sixteen children participated and were tested annually for 3 years. A TV-video procedure was used to measure the accuracy of body size judgments. Variables examined included demographic, familial, sociocultural, social, esteem, and clinical variables. Predictors of higher eating disorder scores for both sexes included height and weight, children's perceptions of parental concerns about their body size, low body esteem, and depression. For girls only, a larger perceived body size and smaller idealized body size were also predictors. Teasing was a predictor for boys only. An analysis of longitudinal changes suggests that low body esteem becomes a significant factor around age 9, depression emerges as a predictor at age 10, and body size judgments in perceived and ideal sizes at ages 11 and 12. Changes over 2 years in individuals' weight and height, teasing, body dissatisfaction, and eating disorder scores were also found to predict higher eating disorder scores. PMID- 11110992 TI - An international study exploring levels of postpartum depressive symptomatology. AB - OBJECTIVE: Differences in postpartum depressive symptomatology (PPDS) among an international sample of 892 women from nine countries representing five continents were explored. METHOD: Edinburgh Postnatal Depression Scale (EPDS) and Beck Depression Inventory (BDI) were used to assess PPDS among a convenience sample that completed the two questionnaires twice, yielding a total of four sets of scores per subject. Women sampled were primiparae with no obstetrical complications, and had a healthy baby. Depression history and therapy were ruled out as exclusion criteria. RESULTS: Mean scores for EPDS and BDI varied across sites at both time points (P value<.001). European and Australian women had the lowest levels of PPDS, USA women fell at the midpoint, and women from Asia and South America had the highest depressive symptom scores. The moderate concordance between the EPDS and BDI suggested that the measures have complementary uses for screening and assessment. CONCLUSION: Utility of EPDS and BDI for yielding profiles of postpartum women's depressive symptomatology was demonstrated. Further research to validate depressive symptom measures with diverse international populations is indicated. PMID- 11110993 TI - Patterns of referral in patients with medically unexplained motor symptoms. AB - OBJECTIVE: To investigate the pattern and reasons for referrals in 64 patients with a stable diagnosis of motor conversion symptoms who had been assessed at the National Hospital for Neurology and Neurosurgery (NHNN). METHOD: Patients were interviewed on average 6 years after their original admission to the NHNN. Hospital notes and GP records were consulted. RESULTS: Psychiatrists at the NHNN saw 75% of patients. Treatment was initiated in 60% of these. During the 6-year follow up, many patients continued to be referred to neurologists and other specialists, but subsequent psychiatric referral was rare. Many changed their GP after discharge from the NHNN and a disproportionate number of re-referrals was made by GPs who had known their patients for less than 6 months. Psychological attribution of symptoms was rare and did not appear to be related to the pattern of referrals. CONCLUSION: The pattern of care of these patients was inconsistent and many felt dissatisfied with the treatment they received. This led to further referrals, unnecessary use of valuable resources and unnecessary exposure to iatrogenic damage. Further studies should aim to assist GPs and other clinicians in deciding when referral is likely to be beneficial. PMID- 11110994 TI - Adolescent cocaine and injection stress effects on the estrous cycle. AB - Chronic cocaine exposure during critical periods of development induces short- and long-term effects. During the pubertal period, the hypothalamic-pituitary gonadal (HPG) axis undergoes many dynamic changes. The present study investigated whether chronic periadolescent cocaine alters reproductive maturity in the rat. Sixty female Long-Evans hooded rats were randomly assigned to one of three conditions (20 mg cocaine/kg/day, saline injected and uninjected), for dosing from postnatal day 21 (P21) through P60. Several indicators of reproductive maturation and functioning were assessed during and following treatment. Cocaine exposure had no effect on the onset of puberty or on the date of first ovulation. The number of proestrus-estrus transitions was significantly lower in cocaine exposed females compared to uninjected females, but not compared to saline injected controls. This reduction was observed during exposure to cocaine, as well as after the cessation of injections. During the dosing period, cocaine exposed rats also exhibited a greater number of cycles that had no clear P-E transition than did UN subjects; this effect disappeared once injections stopped. These alterations suggest immediate, and possibly persisting, alterations in the control of ovulation after chronic cocaine exposure throughout adolescence. Interestingly, during the injection period, the saline-injected females had a significantly greater number of diestrus days compared to uninjected and cocaine injected animals, as well as a lower proportion of regular 4- and 5-day cycles. These differences disappeared once injections stopped. These results suggest a stress-induced irregularity of the estrous cycle, possibly attenuated by cocaine and recoverable after exposure. The present findings indicate that the HPG axis is susceptible to short-term, and possibly to long-term, alterations induced by cocaine exposure throughout the adolescent period. PMID- 11110995 TI - Exercise influences spatial learning in the radial arm maze. AB - Previous studies indicate that the hippocampus is active during exercise, and that neurotrophin expression, receptor density, and survival of dentate gyrus granule cells in the hippocampus can be modified by moderate voluntary exercise. The present study was designed to test the consequences of voluntary exercise on a hippocampal-related behavior. Exercising and control rats were tested on the standard and delayed nonmatch-to-position (DNMTP) version of the eight-arm radial maze, both of which are sensitive to hippocampal damage. Voluntarily exercising rats ran in running wheels attached to their home cage for 7 weeks prior to and throughout testing, and took 30% fewer trials to acquire criterion performance than sedentary controls. Both groups spent the same average time per arm. Once the eight-arm maze had been learned to criterion, group differences were not apparent. Exercise can facilitate acquisition of a hippocampal-related spatial learning task, but does not affect performance following acquisition. Further work will be necessary to link these effects to hippocampal-related variables shown to be influenced by exercise. PMID- 11110996 TI - The effect of dehydroepiandrosterone on Zucker rats selected for fat food preference. AB - When allowed to select between macronutrients in a 1-h-a-day meal paradigm, Zucker rats consume 20-80% of their total caloric intake as fat. If they receive an intraperitoneal injection of DHEA 2 h before such a test meal, they consume fewer total calories. The magnitude of this effect on each macronutrient depends upon the animal's initial preference for fat; the higher the initial fat preference, the more profound is the decrease in caloric intake and the more pronounced the effect on fat consumption. Doses as low as 25 mg DHEA/kg body weight are effective. Lean Zucker rats that prefer to consume a high-fat diet have higher epinephrine and dopamine levels in select regions of the hypothalamus known to control food intake. Administration of DHEA to such animals 2 h before decapitation reduces the content of norepinephrine and these monoamines to levels that mimic the values found in the low-fat-preferring animals. It is hypothesized that exogenous DHEA causes the acute release of norepinephrine, epinephrine, and dopamine in select regions of the hypothalamus, and this release causes a decrease in food intake, particularly fat. PMID- 11110997 TI - Time structure of behavioral patterns related to feed pecking in chicks. AB - Nonrandom time patterns of pecking acts by 16 chicks were detected using the software Theme during three videotaped pecking sessions (M, C, and A). At 15 days of age pecking session, M (mash) was recorded when chicks ate a mash diet. Pecking session C (change) at 16 or 17 days of age was recorded immediately after the change of the diet to pellets presented either as regular cylinders (P) to eight chicks, or as semiovoid (Po) to eight other chicks. Pecking session A (adapted) was recorded 5 or 6 days after adaptation to P and Po. Successful (consumatory) pecks were 72%, 52%, and 61% of all pecks for sessions M, C, and A, respectively. The head of the chicks remained in a steady position between two consecutive pecks for a longer period during C (65% of the time) than M and A (54%). During C, the pecking rate was less for P (0.54 pecks/s) than for Po (0.79 pecks/s). Two consistent time patterns involving four acts frequently observed were: head rotation (or exploratory peck)-->head in steady position-->consumatory peck-->head in steady position with jaw movements. Time intervals within a pattern were stable throughout sessions. However, the proportion of synchronized (included in a pattern) vs. nonsynchronized (not included in a pattern) acts decreased immediately after the change of feed form (session C). These results suggest that pecking at feed is composed of two distinct sets of acts: consistently organized patterns little affected by the form of the pecked particles and nonsynchronized acts that may be involved in sensory information. PMID- 11110998 TI - Increased alcohol intake and behavioral disinhibition in rats with ventral striatal neuron loss. AB - A previous study of ours reported excessive alcohol intake, enhanced defensive aggressiveness (hyperreactivity towards the experimenter), impulsive behavior, and reduced cortical serotonin levels in rats following extensive basal forebrain axon-sparing lesions involving the septal area and the ventral striatum. This constellation of signs resembles that seen clinically in "Dionysian" alcoholics. The present investigation aimed at examining the effect of ibotenic acid lesions restricted to the septal area or the ventral striatum on this behavioral profile. Experiment 1 indicated that medium-sized lesions (induced by infusing 0.35 microl ibotenic acid in each hemisphere) encompassing the septal area or the ventral striatum elicited a qualitatively similar behavioral profile. Both lesion types markedly enhanced the intake of 6% ethanol, and both groups were significantly more hyperreactive towards the experimenter. A brief doorbell signal elicited significantly more fleeing in rats with basal forebrain lesions, and licking from an electrified waterspout in the punished drinking test caused lesser suppression of locomotor activity than normal. Both groups also showed significant deficits in food hoarding. Histological examination revealed that the posterior portion of the ventral striatal lesion typically overlapped with the anterior portion of the septal lesion. Experiment 2 avoided this neuropathological overlap, and examined groups bearing small discrete lesions (induced by infusing 0.15 microl ibotenic acid in each hemisphere) restricted to either the accumbens part of the ventral striatum or the dorsal septal area. Lesions to the nucleus accumbens were associated with an increase in home-cage alcohol drinking, no hyperreactivity towards the experimenter, potentiation of fleeing at the expense of freezing in response to a sudden auditory signal, and disinhibited behavior in the punished drinking test with increased punished responding and reduced behavioral suppression. Rats with small septal lesions showed a weak enhancement of defensive aggression, but no other behavioral alterations. Our results suggest that ventral striatal neuron loss gives rise to excessive alcohol drinking and enhanced impulsivity. PMID- 11110999 TI - Effects of sensory stimulation and post-ingestive consequences on satiation. AB - This study investigated the influence of sensory stimulation, with and without post-ingestive consequences, on satiation by varying the form of a preload and the timing of a mixed meal presented after the preload. Twenty-four, normal weight, non-dieting, college-aged women were randomized to different preload groups: water preload (Water), sip-and-spit energy-dense preload (Taste), or energy-dense preload (Taste/kcal). Volume of fluid consumed prior to the meal was controlled. All participants had sessions in which a meal was provided immediately (0 min) or 30 min after the preload. Results showed equal suppression of intake for participants receiving sensory stimulation from an energy-dense preload (Taste and Taste/kcal groups) in comparison to a water preload (Water group). No effect of time from preload to food consumption was found; the suppression of intake was similar whether the meal immediately followed the preload or was 30 min after the preload. These findings suggest that sensory aspects of food can influence satiation, and in the conditions of this study, had a larger influence on satiation than post-ingestive consequences. PMID- 11111000 TI - Acoustic startle and open-field behavior in mice bred for magnitude of swim analgesia. AB - Acoustic startle response (ASR) and open-field activity was examined in the 46th generation of mice that have been selectively bred for high analgesia (HA) and for low analgesia (LA) induced by 3-min swimming in 20 degrees C water. These lines were earlier found to differ in brain opioid receptor density and in the expression of opioid-mediated phenomena, as analgesic sensitivity to opiates and reversibility of swim stress-induced analgesia (SSIA) by naloxone. For comparison, a randomly bred control (C) line was used. To measure the amplitude of ASR, the mice were exposed to 110-dB acoustic stimuli in a Coulbourn apparatus. In saline-injected mice, the ASR force was found significantly lower in the LA than in the HA, as well in the C line, but did not differ between the two last lines. Naltrexone hydrochloride (10 mg/kg IP 30 min before ASR testing) augmented the startle in the opioid receptor-dense HA line, but had no effect in the opioid receptor-deficient LA line, as well in the C line; therefore, the ASR magnitude in naltrexone-injected HA mice was significantly higher compared to the C line. HA mice displayed less activity in an open-field test; that is, they remained immobile longer in the center of the field, and thereafter performed less ambulation and less rearing against the wall compared to the LA line. Naltrexone failed to modify the open-field activity in any line. The results confirm that the pattern of ASR depends on the genetic makeup of the animals. The higher amplitude of ASR, taken together with the lower open-field activity of HA mice, can be interpreted in terms of higher anxiety level, compared to the LA line. It is suggested that the higher ASR in HA mice relies on a nonopioid mechanism, which is tonically inhibited by the opioid system. PMID- 11111001 TI - Sucrose intake as a function of its cost and the cost of chow. AB - The avid consumption of pure carbohydrate solutions, which often results in a distortion of nutrient balance, is generally presumed to be driven by their taste. In the first of two experiments, we examined the effect of consumption cost on rats' intake of three concentrations of sucrose solution (8%, 16%, and 32%) when a nutritionally complete chow was concurrently freely available. In the second experiment, we examined the intake of 24% sucrose solution and chow as the consumption costs of both were varied. Increasing the cost of sucrose resulted in a reduction in the percent calories taken from sucrose; the steepness of the decline in intake with price was inversely related to the sucrose concentration and to the cost of chow. Chow calories were substituted for relatively expensive sucrose calories. An increase in the cost of chow resulted in a reduction in the percent of calories taken from chow and a protein-poor diet. The cost of sucrose did not affect the slope of the chow intake curve, presumably because, despite its sweet taste, sucrose was not a substitute for the protein, fat, and micronutrients in chow. Total caloric intake was conserved in all cases.Thus, the avid consumption of sucrose solution is curtailed when it is costly; but the degree of change in intake with cost depends on the cost of an alternative food. These results suggest that diet selection involves a comparison not only of the taste and post-ingestive consequences of available foods, but also of the cost of calories and nutrients in the foods. Selection appears to be guided first by caloric requirements and the relative cost of calories, then by nutrient requirements and the relative cost of nutrients, and finally by taste. PMID- 11111002 TI - Thymic stress in artificially reared and maternally reared rat pups. AB - It is well documented that the hypothalamic-pituitary-adrenal (HPA) axis influences immunological responses to stress. Maternal factors have been shown to be necessary for appropriate modulation of the HPA axis in the developing rat. The purpose of this study was to determine whether artificially reared (AR) infant rat pups (a procedure whereby infant rats are gastrostomized and reared independently of maternal factors) have an altered function of the HPA axis in terms of thymocyte apoptosis (programmed cell death) and other indices of thymic stress. AR and maternally reared (MR) Long-Evans rat pups were randomized to control, fasted, stressed, and fasted+stressed treatment groups, as well as an unhandled, MR naive group that served as a baseline control. AR rat pups were significantly heavier than MR (p<0.001). AR rat pups had significantly lighter thymuses than did the MR pups (p<0.001) and fasted pups had significantly lighter thymuses than unfasted pups, regardless of whether they were in the AR or MR condition (p<0.005). AR pups had significantly lower thymic cell numbers and a greater percent of necrotic cells than did MR pups. There were no significant effects of rearing condition on the percent of apoptotic thymocytes. The thymocyte alterations observed in this study between the two rearing conditions suggest that AR reduces thymic weight and cell numbers, which may have consequences for the development of adult cellular immunity. PMID- 11111003 TI - Gustatory responsiveness to food-associated sugars and acids in pigtail macaques, Macaca nemestrina. AB - Taste-preference thresholds for five food-associated sugars and acids, respectively, as well as relative sweet-taste preferences were assessed in six pigtail macaques using two-bottle choice tests of brief duration (1 min). In experiment 1, the animals were found to significantly prefer concentrations as low as 10 mM maltose and sucrose, 20 mM fructose and glucose, and 30 mM lactose over tap water. In experiment 2, the monkeys were given a choice between all binary combinations of the same five saccharides presented in equimolar concentrations of 50, 100, 200, and 400 mM. Preferences for individual sugars were stable across the concentrations tested and indicate the following order of relative effectiveness: maltose>sucrose>glucose> or =fructose> or =lactose. In experiment 3, Macaca nemestrina was found to significantly discriminate concentrations as low as 5 mM malic acid, 10 mM ascorbic acid, 20 mM citric acid and acetic acid, and 0.5 mM tannic acid from the alternative stimulus. With the latter substance, the monkeys rejected all suprathreshold concentrations tested, whereas with the former four substances, the animals showed an inverted U-shaped function of preference. The results showed pigtail macaques to be the first primate species tested so far whose taste-preference threshold for maltose is as low as that for sucrose, and which - similar to rodents - prefers maltose over equimolar concentrations of sucrose and other saccharides. Further, unlike most other primates, pigtail macaques do not generally reject acidic tastants but show a substance- and concentration-dependent change in their behavioral response that may range from rejection to preference. The results support the assumption that the gustatory responsiveness of M. nemestrina to food-associated sugars and acidic tastants might reflect an evolutionary adaptation to its dietary habits. PMID- 11111004 TI - The nitric oxide pathway is an important modulator of stress-induced fever in rats. AB - Psychological stress evokes a number of physiological responses, including a rise in body temperature (T(b)), which has been suggested to be the result of an elevation in the thermoregulatory set point, i.e., a fever. This response seems to share similar mechanisms with infectious fever. A growing number of studies have provided evidence that nitric oxide (NO) has a modulatory role in infectious fever, but no report exists about the participation of NO in stress fever. Thus, the present study aimed to verify the hypothesis that NO modulates stress fever by using restraint stress as a model. To this end, we tested the effects of the non-specific NO synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester (L NAME) or its inactive enantiomer N(G)-nitro-D-arginine methyl ester (D-NAME) on colonic T(b) of restrained or unrestrained rats. A rapid increase in T(b) was observed when animals were submitted to restraint. Intravenous (i.v.) injection of L-NAME at a dose (10 mg/kg) that caused no change in T(b) when administered alone significantly attenuated the elevation in T(b) elicited by stress, indicating that the NO pathway may mediate stress fever. Moreover, intracerebroventricular (i.c.v.) L-NAME (250 microg/microl) caused a rise in T(b) of euthermic animals and enhanced stress fever, supporting that NO in the central nervous system (CNS) leads to a reduction in T(b) and, therefore, this is unlikely to be the site where NO may mediate stress fever. Taken together, these data indicate that the NO pathway plays an important role in modulating restraint stress-induced fever in rats. PMID- 11111005 TI - Immune alterations in three mouse strains following 2-deoxy-D-glucose administration. AB - Using 2-deoxy-D-glucose (2-DG)-induced stress, our laboratory has developed studies to define stress effects on immune responses. Here, we report effects of increasing doses of 2-DG on the immune response of BALB/c, C57BL/6 and BDF(1) mice 2 h after three injections of 0 to 2000 mg/kg of 2-DG. Female 4- to 5-week old mice were euthanized and blood and spleens were collected. A suspension of partially purified mature T splenocytes was obtained by negative selection using J11.d2 antibodies. Glucose and corticosterone levels were measured in the plasma of each mouse. Splenocyte and mature T splenocyte suspensions were tested in in vitro proliferation assays with or without concanavalin A. Splenocytes were analyzed for the following cell-surface markers: CD3, TCR alpha/beta, CD4, CD8 and major histocompatibility complex (MHC) Class II. Significant increases in blood glucose levels were observed in C57BL/6 and BALB/c strains with the highest 2-DG dose (p<0.05). Corticosterone levels were higher in BDF(1) mice and C57BL/6 mice following the administration of 1000 and 2000 mg/kg of 2-DG, respectively (p<0.01). In vitro proliferation of mature T splenocytes in the presence of concanavalin A was decreased in BDF(1) (p<0.05) but not in BALB/c and C57BL/6 mice. In addition, in BDF(1) mice the decrease was highly correlated with an increase of CD3+ and TCR alpha/beta+ cells in the spleen. These results demonstrated high variability in the response of different mouse strains to 2-DG induced stress. PMID- 11111006 TI - Comparison of methyl anthranilate and denatonium benzoate as aversants for learning in chicks. AB - Methyl anthranilate (MeA) has been widely used as a taste aversant for domestic chicks in the one-trial passive avoidance learning (PAL) task. However, MeA has a strong smell that may be aversive to chicks. Therefore, odourless denatonium benzoate (DB) has been suggested as an alternative taste aversant in PAL. The present study was designed to compare the efficacy of MeA and DB as aversants in the one-trial PAL task. In this task, young chicks peck a visually conspicuous bead coated with a taste aversant and in a single trial learn to avoid a similar, but uncoated bead at subsequent presentation. In Experiment 1, chicks were trained using a silver-coloured bead coated with 100% MeA, 0.5% DB or distilled water. After 3 h, MeA-trained, but not DB-trained chicks, exhibited significantly higher avoidance of the test bead than water-trained chicks. In Experiment 2, three pre-training presentations of an uncoated red bead preceded training with the silver bead. MeA-trained chicks showed significantly higher avoidance of the test bead than water-trained chicks. The numbers of water- and DB-trained chicks that avoided pecking the test bead were low and not significantly different from each other. However, DB-trained chicks exhibited significantly longer latencies to peck the test bead than water-trained chicks, indicating that they had retained some memory of the task. Thus, 0.5% DB is a weaker aversant than MeA and it does not induce high levels of learning in the one-trial PAL task. However, DB may prove useful for investigating weakly reinforced learning. PMID- 11111007 TI - Impaired neural regulation of insulin secretion related to the leptin receptor gene mutation in Wistar fatty rats. AB - The Wistar fatty (WF) rat is a model of obese Type 2 diabetes mellitus (DM). These rats were bred by crossing Zucker fatty (ZF) and Wistar-Kyoto (WKY) rats. A homo-allelic leptin receptor gene mutation has been reported in ZF rats. We report here how these genetic factors contribute to plasma insulin regulation. The fasting plasma insulin levels were higher in WKY and Wistar lean (WL) rats than in Zucker lean (ZL) rats (p<0.05). The levels in WF and ZF rats were higher than in their respective lean littermates, WL and ZL rats (p<0.05). After intragastric glucose load, the plasma insulin increase was reduced upon pretreatment by intracerebroventricular (i. c.v.) methylatropine (an antagonist of the cholinergic receptor) injection in WL rats (p<0.05) but not in WF rats. Plasma glucagon-like peptide-1 (GLP-1) response to intragastric glucose load was not affected by methylatropine. After selective hepatic-vagotomy, plasma insulin levels increased in wild-type ZL rats (p<0.05). This increase was not observed in heterozygote ZL rats. Surprisingly, this response of plasma insulin was not shown in wild-type WL and WKY rats. ZF and WF rats did show a prominent decrease in insulin response (p<0.05). These results indicate that the genetic factor in ZF rats is associated with impaired vagal nerve-mediated control of insulin secretion. The genetic factor in WKY rats may diminish sensitivity to the vagal information of insulin release and contribute to insulin resistance. Therefore, we conclude that the presence of both genetic factors in a homo-allelic state is important to the development of DM in WF rats. PMID- 11111008 TI - Attenuation of the anorectic effects of cholecystokinin and bombesin by the specific amylin antagonist AC 253. AB - Previous studies provided evidence for an interaction between the satiety effects of cholecystokinin (CCK), bombesin (BBS), and amylin. Amylin released in response to CCK (or BBS) was supposed to mediate part of CCK's (or BBS's) anorectic effect since the amylin and calcitonin gene-related peptide (CGRP) antagonist CGRP 8-37 attenuated their anorectic action. Due to the low specificity of CGRP 8-37 for amylin vs. CGRP binding sites, the aim of the present study was to test whether the specific amylin antagonist AC 253 also influenced the anorectic effects of CCK and BBS. Injections took place at dark onset in 24-h food-deprived rats. At a dose that attenuated the anorectic effect of amylin (5 microg/kg), the amylin antagonist AC 253 (500 microg/kg) significantly attenuated the anorectic effects of CCK and BBS (0.5 microg/kg). It can therefore be concluded that amylin, rather than CGRP, mediates part of the anorectic effects of CCK and BBS. PMID- 11111009 TI - The vomeronasal organ is involved in discrimination of individual odors by males but not by females in golden hamsters. AB - The vomeronasal organ (VNO) has a wide variety of functions in terrestrial vertebrates, some of which involve responses to classical pheromones whereas others do not. We examined the role of the VNO in discrimination of individual differences in odors of male and female golden hamsters using a habituation paradigm. Removal of the VNO resulted in elimination of the ability of male hamsters to discriminate between some individually distinctive odors (e.g., flank gland secretion), but not others (e.g., urine). In females, such lesions had no effect. The type of test trial also influenced the results; in test trials employing a single, novel odor, removal of the VNO in males did have an effect but in test trials in which both the novel and the familiar odor were presented, VNO removal had no effect. It is concluded that (a) there is a sex difference in the role of the VNO in the discrimination of individual odors, (b) the role of the VNO in discrimination of individual odors varies from odor to odor, and (c) deficits due to VNO removal are more readily observed in more difficult tasks. PMID- 11111010 TI - Effect of maternal separation on feeding behavior of rats in later life. AB - Effects of maternal separation on feeding behavior, particularly on rebound hyperphagia, in adult rats were examined. Time-restricted scheduled feeding (2 h per day for 6 days), was given at the age of 3, 6, 9 or 12 weeks in rats that were maternal separated from postnatal days (PD) 1-21 and control rats. Following the time-restricted scheduled feeding, rats were fed freely for 24 h (rebound hyperphagia). Body weight, daily normal food consumption and food consumption during time-restricted scheduled feeding and rebound hyperphagia were measured. Body weight of 3-week-old maternally separated rats were less than those of control rats. There was no significant difference in normal daily food consumption. Food consumption during rebound hyperphagia was significantly increased in 6- to 9-week-old female maternally separated rats, but there was no difference observed in males. Postnatal maternal separation enhanced rebound hyperphagia of female rats in later life. These results indicate that postnatal maternal separation made rats more vulnerable to the development of abnormal feeding behavior in response to food restriction in later life. PMID- 11111011 TI - Fos expression in female hamsters after various stimuli associated with mating. AB - Detection of the expression of c-fos mRNA or its protein product, Fos, has been used to indicate differences in neuronal response to exogenous stimuli. Factors contributing to differences in Fos expression as a result of various stimuli associated with mating have been extensively studied in the female rat. Less is known about the factors that contribute to Fos expression in female hamsters. Female hamsters differ from female rats in several aspects of sexual behavior; therefore, it seems likely that Fos expression may also differ. The purpose of this study was to determine which factors associated with mating selectively affect Fos expression in the female hamster. Animals were ovariectomized, hormone treated, and then exposed to several behavioral conditions. Fos expression in several brain areas was then assessed via immunocytochemistry (ICC). As has been found by others, mating increases Fos immunoreactivity in a number of brain regions. Specifically, vaginal-cervical stimulation (VCS) was determined to be the salient factor contributing to Fos expression in the preoptic area (POA) and bed nucleus of the stria terminalis (BNST) of ovariectomized hormone primed female hamsters that received a mating interaction. PMID- 11111012 TI - The psychosocial impact of psoriasis: physical severity, quality of life, and stigmatization. AB - Men (N=58) and women (N=43) living with psoriasis completed questionnaires assessing quality of life and feelings of stigmatization. Physician ratings of disease severity were used in conjunction with these variables to account for psychosocial impact. Results showed that ratings of severity were poor predictors of quality of life and stigmatization. Demographic variables (e.g., sex and education) were also generally poor predictors of psychosocial outcome. It is concluded that attempts to understand the psychological impact of psoriasis in terms of current measures of disease severity and demographic characteristics will be limited. PMID- 11111013 TI - Central bombesin inhibits food intake and the orexigenic effect of neuropeptide Y in the neonatal chick. AB - It is well known that central injection of bombesin (BN) suppresses feeding in mammalian and avian species, but the anorexigenic effect of central BN are still open with special reference to the chick. The dose response (0, 0.1 and 0.5 microg) of intracerebroventricular (ICV) injection of BN was examined in Experiment 1. ICV injection of BN inhibited food intake in a dose-dependent manner. Experiment 2 was done to determine whether BN interacts with the orexigenic effect of neuropeptide Y (NPY) in the neonatal chick. Central administration of NPY (2.5 microg) greatly enhanced food intake, but co-injection of BN (0.5 microg) suppressed food intake. The dose response of NPY (2.5 microg) co-injected with three levels of BN (0, 0.1 and 0.5 microg) was examined in Experiment 3. ICV injection of BN attenuated the hyperphagia by NPY in a dose related fashion. It is suggested that central BN may interact with NPY for the regulation of feeding in the neonatal chick. PMID- 11111014 TI - Degradation and glycemic effects of His(7)-glucitol glucagon-like peptide-1(7 36)amide in obese diabetic ob/ob mice. AB - Glucagon-like peptide-1(7-36)amide (tGLP-1) has attracted considerable potential as a possible therapeutic agent for type 2 diabetes. However, tGLP-1 is rapidly inactivated in vivo by the exopeptidase dipeptidyl peptidase IV (DPP IV), thereby terminating its insulin releasing activity. The present study has examined the ability of a novel analogue, His(7)-glucitol tGLP-1 to resist plasma degradation and enhance the insulin-releasing and antihyperglycemic activity of the peptide in 20-25-week-old obese diabetic ob/ob mice. Degradation of native tGLP-1 by incubation at 37 degrees C with obese mouse plasma was clearly evident after 3 h (35% intact). After 6 h, more than 87% of tGLP-1 was converted to GLP-1(9 36)amide and two further N-terminal fragments, GLP-1(7-28) and GLP-1(9-28). In contrast, His(7)-glucitol tGLP-1 was completely resistant to N-terminal degradation. The formation of GLP-1(9-36)amide from native tGLP-1 was almost totally abolished by addition of diprotin A, a specific inhibitor of DPP IV. Effects of tGLP-1 and His(7)-glucitol tGLP-1 were examined in overnight fasted obese mice following i.p. injection of either peptide (30 nmol/kg) together with glucose (18 mmol/kg) or in association with feeding. Plasma glucose was significantly lower and insulin response greater following administration of His(7)-glucitol tGLP-1 as compared to glucose alone. Native tGLP-1 lacked antidiabetic effects under the conditions employed, and neither peptide influenced the glucose-lowering action of exogenous insulin (50 units/kg). Twice daily s.c. injection of ob/ob mice with His(7)-glucitol tGLP-1 (10 nmol/kg) for 7 days reduced fasting hyperglycemia and greatly augmented the plasma insulin response to the peptides given in association with feeding. These data demonstrate that His(7)-glucitol tGLP-1 displays resistance to plasma DPP IV degradation and exhibits antihyperglycemic activity and substantially enhanced insulin-releasing action in a commonly used animal model of type 2 diabetes. PMID- 11111015 TI - Influence of secretin and L-NAME on vascular permeability in the coronary circulation of intact and diabetic rats. AB - The permeability in the intact and diabetic rat coronary circulation after administration of secretin (3.0 micromol/kg i.v.), an inhibitor of NOS (nitric oxide synthase), and L-NAME (N(G)-nitro-L-arginine-methyl ester hydrochloride) (1 mg/kg i.v.), and both substances given together, were studied. To measure protein extravasation Evans blue dye was used as a marker of vascular permeability. The vascular permeability of the left ventricle did not differ in intact and diabetic rats. In the diabetes state increased permeability of atria was observed. Administration of secretin did not influence the coronary vascular permeability in either the intact or the diabetic rats. L-NAME increased the atria permeability and did not change left ventricle permeability. In diabetes, injection of L-NAME caused a decrease in the permeability in both the atria and left ventricle. In intact rats secretin diminished the L-NAME effect in the atria. In diabetic rats co-administration of secretin+L-NAME increased the permeability of the atria and left ventricle, but L-NAME administered alone decreased them. Secretin modified the effect of L-NAME on coronary permeability in intact and diabetic rats. PMID- 11111017 TI - Direct action of growth hormone-releasing hormone agonist JI-38 on normal human fibroblasts: evidence from studies on cell proliferation and c-myc proto-oncogene expression. AB - Growth hormone-releasing hormone (GHRH) is secreted by the hypothalamus and stimulates the release of growth hormone from the pituitary. Recent studies also indicate that in addition to its neuroendocrine function, GHRH may play a direct role in the proliferation of cancer cells, acting as growth factor for various human tumors. In the present study we investigated the effects of JI-38, an agonistic analog of GHRH, on the rate of proliferation of normal human diploid dermal fibroblasts (NHF) cultured in vitro. The effects of JI-38 on the levels of mRNA for c-myc proto-oncogene were also tested. Exposure to 10(-7) M JI-38 stimulated the rate of proliferation of early passage NHF by about 100%. Exposure of NHF cells to 10(-8)-5x10(-6) M JI-38 for 24 h resulted in about 0.5-3.5 fold increase in the levels of mRNA for c-myc proto-oncogene. The ability of JI-38 to stimulate the proliferation of NHF cells was abolished in cells cultured at late passage. Continuous exposure to 10(-7) M JI-38, over 6-7 passages (15-20 population doublings), progressively reduced the rate of proliferation of NHF compared with cells exposed to medium alone, indicating that GHRH agonist acted as a growth inhibitor. Our results suggest that at certain developmental stages, GHRH may act on various tissues, stimulating cell proliferation. PMID- 11111016 TI - Serum pancreastatin levels predict response to hepatic artery chemoembolization and somatostatin analogue therapy in metastatic neuroendocrine tumors. AB - INTRODUCTION: Neuroendocrine tumors often metastasize to the liver and present with disabling hormonal symptoms. Hepatic artery chemoembolization (HACE) combined with somatostatin therapy, pre-embolization, peri-embolization and post embolization, at doses to control symptoms, is an aggressive approach that can relieve hormonal symptoms with minimal morbidity and mortality. METHODS: Chemoembolization was performed using 30 mg of adriamycin, 50 mg of mitomycin, 12 ml of hexabrix, 10 ml of ethiodol, and 360-500-microm particles. Pancreastatin, a split product of chromogranin A, was measured pre-HACE and post-HACE in all patients. RESULTS: Forty-three chemoebolization procedures were performed in 34 symptomatic patients from December 1995 to August 1999. Seventeen patients had intestinal primaries (50%), seven had pancreatic primaries (20%), five had bronchial primaries (15%), and five had unknown primaries (15%). Systemic pancreastatin levels were improved or stable in 31 patients (78%). Symptoms were improved in these 31 patients (78%). Systemic serotonin levels were improved or stable in 24 patients (60%). Radiographic improvement or stability was seen in 18 patients (45%). Procedural related morbidity included pain, fevers, nausea, vomiting, and transient elevations of liver function studies in 75-100% of patients. There was one procedural related mortality (2%). Less than 20% improvement in pancreastatin levels from baseline was associated with death in five of five patients (100%). This was not observed with serotonin levels. CONCLUSION: Measurement of serum pancreastatin levels is an easy and useful method to predict success in patients who undergo HACE plus somatostatin therapy for metastatic neuroendocrine tumors to the liver. This therapeutic approach is effective in relieving symptoms in 78% of patients, with minimal major morbidity or mortality. PMID- 11111018 TI - Transgenic mouse models of angiotensin receptor subtype function in the cardiovascular system. AB - Angiotensin II mediates is biological actions via different subtypes of G protein coupled receptors, termed AT(1) and AT(2) receptors. In rodents, two AT(1) receptors have been identified, AT(1A) and AT(1B), whereas in humans a single AT(1) receptor exists. Recently, a number of transgenic animal models have been generated which overexpress or lack functional angiotensin II receptor subtypes. This review focuses on the physiological significance of angiotensin II receptor subtype diversity in the cardiovascular system. In the mouse, AT(1A) receptors are the major regulators of cardiovascular homeostasis by determining vascular tone and natriuresis. In addition, AT(1A) receptors mediate growth-stimulating signals in vascular and cardiac myocytes. AT(1B) receptors participate in blood pressure regulation, and their functions become apparent when the AT(1A) receptor gene is deleted. Deletion of the mouse gene for the AT(2) receptor subtype led to hypersensitivity to pressor and antinatriuretic effects of angiotensin II in vivo, suggesting that the AT(2) receptor subtype counteracts some of the biological effects of AT(1) receptor signalling. PMID- 11111019 TI - Metabolism of glucagon by dipeptidyl peptidase IV (CD26). AB - Glucagon is a 29-amino acid polypeptide released from pancreatic islet alpha cells that acts to maintain euglycemia by stimulating hepatic glycogenolysis and gluconeogenesis. Despite its importance, there remains controversy about the mechanisms responsible for glucagon clearance in the body. In the current study, enzymatic metabolism of glucagon was assessed using sensitive mass spectrometric techniques to identify the molecular products. Incubation of glucagon with purified porcine dipeptidyl peptidase IV (DP IV) yielded sequential production of glucagon(3-29) and glucagon(5-29). In human serum, degradation to glucagon(3-29) was rapidly followed by N-terminal cyclization of glucagon, preventing further DP IV-mediated hydrolysis. Bioassay of glucagon, following incubation with purified DP IV or normal rat serum demonstrated a significant loss of hyperglycemic activity, while a similar incubation in DP IV-deficient rat serum did not show any loss of glucagon bioactivity. Degradation, monitored by mass spectrometry and bioassay, was blocked by the specific DP IV inhibitor, isoleucyl thiazolidine. These results identify DP IV as a primary enzyme involved in the degradation and inactivation of glucagon. These findings have important implications for the determination of glucagon levels in human plasma. PMID- 11111020 TI - Chronic hypoxia induced down-regulation of angiotensinogen expression in rat epididymis. AB - The presence of an intrinsic renin-angiotensin system (RAS) in the rat epididymis has been previously established by showing the expression of several key RAS components, and in particular angiotensinogen, the indispensable element for the intracellular generation of angiotensin II. In this study, the possible involvement of this local epididymal RAS in the testicular effects of chronic hypoxia was investigated. Semi-quantitative reverse-transcription polymerase chain reaction (RT-PCR), Western blotting and by in situ hybridization histochemistry of the rat epididymis were used to show changes in localization and expression of angiotensinogen. Results from RT-PCR analysis demonstrated that chronic hypoxia caused a marked decrease (60%) in the expression of angiotensinogen mRNA, when compared with that in the normoxic epididymis. Western blot analysis demonstrated a less decrease (35%) in the expression of angiotensinogen protein. In situ hybridization histochemistry showed that the reduced angiotensinogen mRNA in chronic hypoxia was specifically localized to the epididymal epithelium from the cauda, corpus and caput regions of the epididymis; a distribution similar to that of normoxic rats. It was concluded that chronic hypoxia decreases the transcriptional and translational expression of angiotensinogen, and thus local formation of angiotensin II, in the rat epididymis. PMID- 11111021 TI - Erratum to Abstracts, Neuropeptides 2000. PMID- 11111022 TI - Effect of lipoteichoic acid on the uptake of Streptococcus pyogenes by HEp-2 cells. AB - Lipoteichoic acid (LTA) is thought to play a role in the interactions between Streptococcus pyogenes and host cells. We have examined the effect of exogenous LTA on the adherence and entry of S. pyogenes JRS4 strain into HEp-2 epithelial cells. LTA markedly inhibited bacterial entry in a concentration-dependent manner, up to 250 microg ml(-1). In contrast, LTA had only a slight inhibitory effect on adherence. LTA also inhibited the entry but not adherence of Salmonella typhimurium strain into HEp-2 cells. Binding experiments showed a dose-dependent binding of LTA to cells up to 10 microg ml(-1). Confocal laser microscopy imaging and analysis revealed that LTA was internalized by the epithelial cells and colocalized with F-actin. These results might imply that, following binding, exogenous LTA enters HEp-2 cells and exerts a cytotoxic effect that interferes with bacterial internalization. A possible target for LTA activity might be the actin cytoskeleton, which is known to be essential for bacterial uptake. PMID- 11111024 TI - Rapid gene inactivation in Pseudomonas aeruginosa. AB - A rapid and efficient method to inactivate genes in Pseudomonas aeruginosa has been developed. It is based on pKnockout vectors which carry either a gentamicin or a streptomycin/spectinomycin resistance cassette allowing for selection in P. aeruginosa where these vectors do not replicate. A PCR fragment of the gene of interest carrying 5'- and 3'-truncations is cloned into a pKnockout vector, mobilized into P. aeruginosa, and subsequently integrated into the chromosomal copy of the target gene. The orientation of the fragment determines whether (i) the target gene is disrupted without blocking the transcription of downstream genes or (ii) the insertion exerts a polar effect thereby leading to inactivation of a whole operon. PMID- 11111023 TI - Production of green fluorescent protein by the methylotrophic bacterium methylobacterium extorquens. AB - The production of green fluorescent protein (GFP) in Methylobacterium extorquens was studied by creating four different constructs using pJB3KmD, pRK310 and pVK101 vectors, as well as pLac and soluble methane monooxygenase (sMMO) promoters. Plasmids were introduced into the cells by electroporation. Expression of GFP by selected clones was evaluated by growing cells in complex or defined media. The use of pRK310 as an expression vector containing the lacZ promoter resulted in a 100-fold increase of GFP production when compared to cells containing the pLac-GFP-pJB3KmD construct. Higher production of GFP was observed also in cells containing pLac-GFP-pRK310 and pmmoX-GFP-pVK101 constructs. While the transcriptional regulation of the smmo gene in Methylosinus trichosporium OB3b is known to be copper-dependent, expression of GFP by M. extorquens clones harboring pmmoX-promoters was not strongly controlled by the presence of copper in the medium. The production of GFP was generally constant throughout the growth of M. extorquens carrying the pLac-GFP-pRK310 construct. GFP yields varied between 850 and 1000 microg of GFP g biomass(-1). However, the yield of GFP in cells carrying pmmoX-GFP-pVK101 was somewhat reduced after the mid-exponential phase of growth. PMID- 11111025 TI - Negative fungal chemotropism to toxic metals. AB - Hyphal growth responses of Geotrichum candidum, Gliocladium roseum, Humicola grisea and Trichoderma viride to Cu and Cd were studied using a simple tessellated agar tile system. Negative chemotropic behaviour of hyphae, which included curling and growth away from metal-containing domains, occurred in all species and with both metals. Both toxic metal and sucrose concentrations in the medium modulated the magnitude of the negative chemotropic effects observed. In general, greater concentrations of metals led to a higher level of negative chemotropism in response to Cu and Cd, which could be reduced with increasing concentrations of sucrose in the medium. This suggests that resource availability affects the ability of these fungi to grow into metal-laden domains. PMID- 11111026 TI - Identification of secreted proteins of the cyanobacterium Synechocystis sp. strain PCC 6803. AB - We investigated the spectrum of secreted proteins in the cyanobacterium Synechocystis, and identified these proteins by amino-terminal sequencing. In total, seven sequences have been determined that corresponded to the proteins Sll0044, Sll1694, Sll1891, Slr0924, Slr0841, Slr0168, and Slr1855. The protein Sll1694 of 18 kDa that formed one of two major bands on SDS-PAGE was identified as cyanobacterial pilin, PilA. The amino-terminal sequence of another protein that formed a second major band was blocked. The analysis of the data revealed that five of seven proteins had distinct putative leader sequences for secretion. PMID- 11111027 TI - Cloning and heterologous expression of a sulfur oxygenase/reductase gene from the thermoacidophilic archaeon Acidianus sp. S5 in Escherichia coli. AB - A thermoacidophilic, obligately chemolithotrophic, facultatively aerobic archaebacterium, Acidianus sp. S5, was isolated from acidothermal springs in southwest China. The sulfur oxygenase/reductase (SOR) gene of Acidianus sp. S5 was cloned and expressed in Escherichia coli. Several primers were designed and successfully applied for detection and cloning of the sor gene. A 3.7-kb EcoRI fragment containing the sor gene and three neighboring open reading frames was sequenced. Sequence analysis indicated that the sor gene of Acidianus sp. S5 showed 81% identity to the sor gene of Acidianus ambivalens. E. coli cells carrying the sor gene on pBV220SOR were able to overproduce SOR upon a temperature shift from 30 to 42 degrees C. SOR produced in E. coli catalyzes the oxidation of elemental sulfur and concomitant production of sulfite, thiosulfate and hydrogen sulfide. The recombinant enzyme exhibits the same catalytic properties as the one from Acidianus S5. PMID- 11111028 TI - Gene disruption analysis of DppA isolated as a periplasmic molecular chaperone like protein for folding of dimethyl sulfoxide reductase in Rhodobacter sphaeroides f. sp. denitrificans. AB - The effect of inactivation of DppA, a dipeptide transport protein identified as a periplasmic molecular chaperone-like protein, on the formation of active dimethyl sulfoxide reductase (DMSOR) was examined in Rhodobacter sphaeroides f. sp. denitrificans. All of the dppA-disrupted mutants produced a normal level of native form of DMSOR and grew by DMSO respiration, indicating that the loss of DppA protein alone had no effect on the formation of active DMSOR. The periplasmic fraction of the dppA-disrupted mutant also had the activity to prevent aggregation of acid-unfolded DMSOR. Two proteins, DctP and BztA, were further identified as the proteins with the activity. Their activities, however, were much lower than that of DppA. These results suggest that several substrate binding proteins might be implicated in the folding of unfolded DMSOR in the periplasm. PMID- 11111029 TI - The pyrroloquinoline quinone synthesis genes of Gluconobacter oxydans. AB - A Tn5-induced glucose dehydrogenase (GDH) deficient mutant of Gluconobacter oxydans IFO 3293 was characterised. DNA sequencing showed that the insertion site occurred in an open reading frame with homology to the pqqE gene. It was shown that acid production could be restored by addition of the coenzyme pyrroloquinoline quinone (PQQ) to the medium. The pqq cluster of G. oxydans ATCC 9937 was cloned and sequenced. It has five genes pqqA-E. The cluster could complement the Tn5-induced mutation in IFO 3293. Pulsed-field gel electrophoresis suggested that the pqq genes are not closely linked to the ribF gene that produces the riboflavin cofactor for the gluconic acid dehydrogenase. PMID- 11111030 TI - Enzymatic versus spontaneous S-methyl thioester synthesis in Geotrichum candidum. AB - The synthesis of short chain S-methyl thioesters was investigated in Geotrichum candidum strain GcG. The results indicated the involvement of an enzymatic reaction in this microorganism that led to the synthesis of S-methyl thioacetate (MTA) when methanethiol and acetyl-CoA were used as substrates. MTA was generated from these substrates by enzymatic or spontaneous reactions, whose relative importance depended largely on pH and temperature. For longer chain acyl-CoA compounds (C3 to C6), thioester synthesis was primarily spontaneous. Short chain fatty acid activation by a CoA residue probably is a prerequisite for the synthesis of S-methyl thioesters. PMID- 11111031 TI - Multiplex PCR-based detection and identification of Leuconostoc species. AB - A multiplex polymerase chain reaction (PCR) assay has been developed for rapid and reliable identification of Leuconostoc species, by using species-specific primers targeted to the genes encoding 16S rRNA. This assay can detect and differentiate Leuconostoc species from mixed populations in natural sources as well as from pure cultures, within 3 h. This assay system consists of a total of 10 primers, two primers from each target species, and comprises two multiplex PCR reactions: one reaction for Leuconostoc carnosum, Leuconostoc citreum and Leuconostoc mesenteroides, and another reaction for Leuconostoc gelidum and Leuconostoc lactis. This multiplex PCR assay was used to identify 31 Leuconostoc strains isolated from kimchi, a fermented-cabbage product, and the results showed perfect correlation with the results of a polyphasic method, including 16S rDNA sequencing and DNA-DNA hybridization. In addition, this assay enables simultaneous detection of the above-mentioned Leuconostoc species when chromosomal DNA from these Leuconostoc species was mixed. Thus, these results suggest that this multiplex PCR is a rapid and reliable method for identification of Leuconostoc species in pure cultures or in mixed populations. PMID- 11111032 TI - Taxol from Tubercularia sp. strain TF5, an endophytic fungus of Taxus mairei. AB - The diterpenoid taxol is an important anticancer agent used widely in the clinic. The purpose of this work was to identify a taxol-producing endophytic fungus (strain TF5) isolated from Taxus mairei and study its anticancer activities. Strain TF5 was identified as a Tubercularia sp. according to the morphology of the fungal culture, the mechanism of spore production and the characteristics of the spores. Strain TF5 produced taxol, when grown in potato dextrose liquid medium and analyzed by thin layer chromatography, high performance liquid chromatography, ultraviolet and mass spectrometry. The fungal taxol, which was isolated from the organic extract of the TF5 culture, had strong cytotoxic activity towards KB and P388 cancer cells in vitro, tested by the MTT assay. Observed with immunofluorescence and electron microscopy, the fungal taxol enhanced microtubule stability and bundling in culture cells and induced tubulin polymerization in vitro similar to the authentic taxol. PMID- 11111033 TI - PCR differentiation of Saccharomyces cerevisiae from Saccharomyces bayanus/Saccharomyces pastorianus using specific primers. AB - The aim of the present study was to design species-specific primers capable of distinguishing between Saccharomyces cerevisiae, Saccharomyces bayanus/Saccharomyces pastorianus. The 5'-specific primers were designed from the ITS-1 region (between positions 150 and 182 from the 3'-SSU end) and the 3' specific primers were located in the LSU gene (positions 560-590 from the 5'-end of this gene). These primers were tested with different collections and wild strains of these species and the results showed that the primers were capable of distinguishing between S. cerevisiae strains and S. bayanus/S. pastorianus. Not enough sequence differences were found between S. bayanus and S. pastorianus to design specific primers for these species using this region. This method offers an effective tool for a quick differentiation of the Saccharomyces strains of the most common species involved in industrial processes. PMID- 11111034 TI - Isolation and piezoresponse of the rpoA gene encoding the RNA polymerase alpha subunit from the deep-sea piezophilic bacterium Shewanella violacea. AB - The rpoA gene encoding the alpha subunit of RNA polymerase from the deep-sea piezophilic bacterium Shewanella violacea DSS12 was cloned and sequenced. The rpoA gene was found to encode a polypeptide consisting of 329 amino acids with a molecular mass of 36238 Da. S. violacea alpha protein was expressed in a ts Escherichia coli mutant, to confirm whether the rpoA gene is functional. It complemented this mutation, indicating a chimeric RNA polymerase is assembled at the non-permissive temperature. Recombinant alpha protein was overexpressed using an expression plasmid harboring the rpoA gene and purified to near homogeneity. Primer extension analysis revealed that two transcriptional initiation sites are recognized by sigma(70) RNA polymerase. It also indicated that pressure response (piezoresponse) in the alpha operon occurred at the transcriptional level, suggesting some positive regulators may interact with the transcriptional apparatus and regulate the expression of the operon at different pressure conditions. PMID- 11111035 TI - Dynamic accumulation and degradation of poly(3-hydroxyalkanoate)s in living cells of Azotobacter vinelandii UWD characterized by (13)C NMR. AB - The synthesis and degradation of poly(3-hydroxybutyrate) (PHB) and poly(3 hydroxybutyrate-co-hydroxyvalerate) (P(HB-co-HV)) by Azotobacter vinelandii UWD were investigated using natural abundance solution (13)C nuclear magnetic resonance (NMR) in vivo in shake flask culture and in fermenter culture. The synthesis and the degradation of poly(3-hydroxyalkanoate)s (PHA) monomers hydroxybutyrate (HB) and hydroxyvalerate (HV) had different rates. The amount of HB and HV increased dramatically in the initial degradation stage. The results suggest that the intracellular PHA of strain UWD was the subject of dynamic metabolic processing. (13)C NMR in vivo analysis provided a rapid, easy, accurate, non-destructive method to obtain valuable information on the metabolism of PHA. PMID- 11111036 TI - Combined molecular ecological and confocal laser scanning microscopic analysis of peat bog methanogen populations. AB - Confocal laser scanning microscopy, using fluorescently labelled oligonucleotide probes targeting the 16S rRNA of different physiological groups of methanogens, was used to identify which methanogenic genera were present and to describe their in situ spatial locations in samples taken at different depths from blanket peat bog cores. Total bacterial DNA was also extracted and purified from the samples and used as template for amplification of 16S rRNA and regions of methyl CoM reductase-encoding genes using the polymerase chain reaction, as well as for oligonucleotide hybridisation experiments. These techniques, used in concert, demonstrated that methanogens of several physiological groups were present in highest numbers in the mid regions of 25 cm deep peat cores. Some discrepancies were apparent in the findings of the microscopic and molecular methods, though these may be partially accounted for by the different sensitivities of the techniques employed. The combined approaches used in this study gave an insight into the diversity and distribution of methanogens in peat environments not possible using molecular ecological methods alone. PMID- 11111037 TI - The family of light-harvesting-related proteins (LHCs, ELIPs, HLIPs): was the harvesting of light their primary function? AB - Light-harvesting complex proteins (LHCs) and early light-induced proteins (ELIPs) are essential pigment-binding components of the thylakoid membrane and are encoded by one of the largest and most complex higher plant gene families. The functional diversification of these proteins corresponded to the transition from extrinsic (phycobilisome-based) to intrinsic (LHC-based) light-harvesting antenna systems during the evolution of chloroplasts from cyanobacteria, yet the functional basis of this diversification has been elusive. Here, we propose that the original function of LHCs and ELIPs was not to collect light and to transfer its energy content to the reaction centers but to disperse the absorbed energy of light in the form of heat or fluorescence. These energy-dispersing proteins are believed to have originated in cyanobacteria as one-helix, highly light-inducible proteins (HLIPs) that later acquired four helices through two successive gene duplication steps. We suggest that the ELIPs arose first in this succession, with a primary function in energy dispersion for protection of photosynthetic pigments from photo-oxidation. We consider the LHC I and II families as more recent and very successful evolutionary additions to this family that ultimately attained a new function, thereby replacing the ancestral extrinsic light-harvesting system. Our model accounts for the non-photochemical quenching role recently shown for higher plant psbS proteins. PMID- 11111038 TI - Efficient insertional mutagenesis in Streptococcus thermophilus. AB - Bacteria have always been considered ideal organisms for genetic analysis. While this is true for some model organisms, like Escherichia coli, Bacillus subtilis and, more recently, Lactococcus lactis, genetic analysis of other organisms is often prevented by lack of valuable tools, like vectors, transposons and methods for transformation, gene inactivation and random insertional mutagenesis. This is the case of the moderately thermophilic bacterium Streptococcus thermophilus, an organism that, in spite of its widespread use for food fermentations, is only poorly characterized. We report here an insertional mutagenesis system that allows efficient random mutagenesis, easy characterization of the interrupted genes and construction of stable null mutations. This may become a powerful S. thermophilus tool for both genetic analysis and construction of 'food-grade' mutants of this biotechnologically relevant microorganism. PMID- 11111039 TI - Differential DNA binding and transcription modulation by three T-box proteins, T, TBX1 and TBX2. AB - T-box genes encode a family of phylogenetically conserved DNA-binding proteins that regulate gene expression during embryogenesis. While the developmental importance of many T-box genes has been well documented, little is known about how family members differ in their DNA binding properties and ability to modulate transcription. Here we show that although TBX1, TBX2 and the Xenopus T protein (Xbra) share only 50-60% identity within their DNA-binding domains they can bind the same DNA sequence in vitro. However, the proteins differ in three important respects. While TBX1 protein binds a palindromic T oligonucleotide as a dimer, as had been previously reported for Xbra, TBX2 appears to bind the same DNA sequence as a monomer. Also, T protein/DNA complexes are stabilized in vitro by the addition of specific antibodies, whereas TBX2/DNA complexes are not stabilized by antibodies. Most importantly, TBX2 represses while Xbra activates transcription of the same chimeric reporter plasmid. TBX1, although capable of binding to the chimeric promoter, has no effect on transcription. Thus, while the DNA binding domains of T-box proteins share substantial homology, these proteins differ in both their DNA binding and transcriptional modulation properties. These results suggest that the various T-box proteins, while highly conserved, likely use different mechanisms to modulate transcription and may have different targets in vivo. PMID- 11111040 TI - Cloning and characterization of the genes encoding the ankyrin repeat and SOCS box-containing proteins Asb-1, Asb-2, Asb-3 and Asb-4. AB - Members of the suppressor of cytokine signalling (SOCS) family of proteins have been shown to inhibit cytokine signalling via direct interactions with JAK kinases or activated cytokine receptors. In addition to their novel amino terminal regions and SH2 domains that mediate these interactions, the SOCS proteins also contain carboxy-terminal regions of homology called the SOCS box. The SOCS box serves to couple SOCS proteins and their binding partners with the elongin B and C complex, possibly targeting them for degradation. Several other families of proteins also contain SOCS boxes but differ from the SOCS proteins in the type of domain or motif they contain upstream of the SOCS box. We report here the cloning, characterization, mapping and expression analysis of four members of the ankyrin repeat and SOCS box-containing (Asb) protein family. PMID- 11111041 TI - Characterization of the mouse Smad1 gene and its expression pattern in adult mouse tissues. AB - Smad1 belongs to a family of receptor-activated proteins which mediate signals from TGF-beta superfamily ligands, including TGF-beta and BMPs. Although much is known about the biochemistry of Smad1 signal transduction, the role of Smad1 in vivo is still unclear. Here we present the first description of the genomic structure of the mouse Smad1 gene and the characterization of its expression pattern in adult mouse tissues by immunohistochemistry. The Smad1 gene contains 7 exons and spans >42 kb of genomic DNA. Its coding region is contained within 6 exons and all introns, except intron 1, follow the GT/AG rule. Immunohistochemical analysis shows that Smad1 is widely expressed in adult mouse tissues, with a varying degree of nuclear localization in different cell types, suggesting a regulated function for this protein. This study assigns all of the exon-intron boundaries of the mouse Smad1 gene and provides the basis for assessing the functional significance of this gene using targeted gene manipulation in the mouse. PMID- 11111042 TI - Genetic and biochemical characterization of a protein phosphatase with dual substrate specificity in Streptomyces coelicolor A3(2). AB - A gene encoding a protein phosphatase (SppA) with a phosphoesterase motif, which was predicted by the genome project of the Gram-positive bacterium Streptomyces coelicolor A3(2), was cloned by PCR in pET32a(+) and expressed in Escherichia coli. SppA fused to thioredoxin (TRX-SppA) showed distinct heat-stable phosphatase activity toward p-nitrophenyl phosphate with optimal pH 8.0 and optimal temperature 55 degrees C. Mn2+ greatly enhanced enzyme activity, as is found with other protein Ser/Thr phosphatases. TRX-SppA was not inhibited by sodium orthovanadate or okadaic acid, both of which are known to be specific inhibitors of protein phosphatases. TRX-SppA showed phosphatase activity toward not only phosphoThr (pThr) and pTyr but also oligopeptides containing pSer, pThr, and pTyr, indicating that SppA is a protein phosphatase with dual substrate specificity. Disruption of the chromosomal sppA gene resulted in severe impairment of vegetative growth. All of these observations show that SppA, a protein phosphatase with dual specificity, plays an important, but not essential, role in vegetative growth of S. coelicolor A3(2). The presence of a single copy of sppA in all the 13 Streptomyces species examined, as determined by Southern hybridization, suggests a common role of SppA in general in Streptomyces species. PMID- 11111043 TI - Modulation of early growth response (EGR) transcription factor-dependent gene expression by using recombinant adenovirus. AB - Early growth response (EGR) transcription factors link initial cytoplasmic events to long-term alterations of cellular gene expression and are induced by various stimuli. To test their roles in cell physiology, we constructed adenoviral recombinants encoding NGFI-A binding protein 2 (NAB2, a repressor of EGR1, EGR2, and EGR3), EGR1, NAB-insensitive EGR1(I293F) (EGR1*), EGR2, and the NAB-binding, repressive domain 1 (R1) of EGR1. These viruses regulated EGR-dependent expression of GFP and luciferase reporter genes in heterologous expression assays. Infection of a myoblast cell line with EGR1 and EGR1* adenovirus induced the endogenous gene for platelet-derived growth factor A chain (PDGF-A). In addition, in neuroblastoma cells, the two novel EGR1 target genes EGR3 and NAB2 were identified by using adenoviral transfer of EGR1 and EGR1*. Our results demonstrate that recombinant adenovirus is useful to regulate heterologous and endogenous EGR target gene expression, and suggest that EGR transcription factors can autoregulate themselves. PMID- 11111044 TI - Cloning and analysis of the untranslated regions of the Xenopus laevis Connexin30 mRNA. AB - The full-length 5' and 3' untranslated regions (UTRs) of Xenopus laevis Connexin30 (Cx30) mRNA were cloned and sequenced. The Cx30 messenger contains a 148 nt 5' UTR and a 480 nt 3' UTR. Four different constructs were made to enable the analysis of the role of the Cx30 UTRs in translation efficiency and in protein localization in the early Xenopus embryo. Transcripts encoded the Green Fluorescent Protein (GFP) reporter and contained the 5' and 3' UTR of either Cx30 or globin. In vivo analyses after injection of the transcripts into one cell stage Xenopus embryos showed that the Cx30 3' UTR enables very efficient translation. The 5' UTR was slightly inhibitory compared with the globin 5' UTR. The localization of the produced GFP was analyzed. GFP was ubiquitously expressed in all parts of the embryo. Based on this observation we conclude that neither the 5' UTR nor the 3' UTR confers specific localization of the translation of the Cx30 mRNA in the embryo. PMID- 11111045 TI - Identification of upstream stimulatory factor as an activator of the human dipeptidyl peptidase IV gene in Caco-2 cells. AB - The 5' upstream region (-448/-443) of the human dipeptidyl peptidase IV gene promoter containing a consensus E-box (CACGTG) was shown to bind upstream stimulatory factor using nuclear extracts from mouse (3T3) fibroblasts and the human intestinal and hepatic epithelial cell lines Caco-2 and HepG2. Supershift analysis with specific antibodies to USF-1 and USF-2 indicates that USF-1 is the primary isoform binding to the E-box in nuclear extracts from these cells. Using cell culture, transient cotransfection of USF expression vectors with dipeptidyl peptidase IV promoter constructs revealed that both USF-1 and USF-2 caused an approximately tenfold increase in reporter gene expression in Caco-2 cells. Mutant forms of USF-1 and -2 lacking the DNA binding or transcriptional activation domains were unable to stimulate reporter gene expression. Mutation of the E-box prevented binding of USF, although stimulation of reporter gene expression by cotransfection with USF was reduced by only 50%. By using a series of deletion constructs in cotransfection experiments, a second possible site of USF interaction with the dipeptidyl peptidase IV promoter was localized to the 119/-88 region. PMID- 11111046 TI - Cloning of a human tRNA isopentenyl transferase. AB - A cDNA of human origin is shown to encode a tRNA isopentenyl transferase (E.C. 2.5.1.8). Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA. The deduced amino acid sequence contains a number of regions conserved in known tRNA isopentenyl transferases. The similarity to the S. cerevisiae MOD5 protein is 53%, and to the Escherichia coli MiaA protein 47%. The human sequence was found to contain a single C2H2 Zn-finger-like motif, which was detected also in the MOD5 protein, and several putative tRNA transferases located by BLAST searches, but not in prokaryotic homologues. PMID- 11111047 TI - Stability of DNA repeats in Escherichia coli dam mutant strains indicates a Dam methylation-dependent DNA deletion process. AB - In this study we report on the stabilization of short direct repetitive DNA elements. We arranged a 20 bp SK-primer element in a direct repeat manner within the cloning vector pBluescript KS (+). This resulted in an array of 27 direct repeats consisting of 24 bp units. We show that this plasmid could only be propagated without deletion of repeats in dam mutant Escherichia coli hosts, whereas all efforts to use strains that were defective in the methylation dependent restriction system and the recA- or the mismatch repair-dependent deletion system failed. The deletions always affected whole repeat units and not parts of them, leading to an unpredictable reduction of the unit number down to a range of between 12 and two during propagation. We conclude that a Dam methylation-dependent, but recA- and mismatch repair-independent, deletion mechanism caused the DNA rearrangements without an obvious involvement of the known methylated-DNA restriction systems. PMID- 11111048 TI - Genetic structure and functional implication of the fcb gene cluster for hydrolytic dechlorination of 4-chlorobenzoate from Pseudomonas sp. DJ-12. AB - The fcb gene cluster responsible for the hydrolytic dechlorination of 4 chlorobenzoate (4CBA) was cloned from the chromosomal DNA of Pseudomonas sp. DJ 12, and its nucleotide sequence analyzed. The gene cluster was organized in the order fcbB-fcbA-fcbT1-fcbT2-cbT3-fcbC, which is different from that reported in other bacteria. A promoter-like sequence (-35 and -10 region) is located upstream of the fcbB gene and putative ribosome-binding sequences were found upstream of the respective orfs. A stem-loop transcription terminator structure is found downstream of fcbC. This suggests that the six orfs are transcribed into a polycistronic mRNA. The FcbA, FcbB, and FcbC enzymes for dechlorination of 4CBA have a relationship in common with the enzymes involved in fatty acid metabolism on the basis of their deduced amino acid sequences. The proteins encoded by fcbT1, fcbT2, and fcbT3 show similarity to those encoded by dctP, dctQ, and dctM of Rhodobacter capsulatus respectively, which encode transporter proteins for C4 dicarboxylate. It is likely, therefore, that these proteins of DJ-12 play a role in transport of 4CBA into the cell. PMID- 11111049 TI - Identification of two putative novel folate receptor genes in humans and mouse. AB - Utilizing a 'database mining' strategy to detect novel folate receptors (FR), we identified two potential novel members in the mouse and human. The mouse gene (Folbp3) was sequenced and found to predict a 28.2 kDa protein that consists of 244 amino acids that is highly expressed in both the thymus and spleen, suggesting a potential role in the immune system. The human gene (FR-delta) is mapped to chromosome 11q14, and predicts a 27.7 kDa protein that is comprised of 241 amino acids. However, expression of the human gene was not detected in 59 samples from both adult and embryonic tissue sources, suggesting a highly restricted spatial/temporal expression pattern, an alternatively spliced variant or an additional FR pseudogene. Using T31 mouse radiation hybrid mapping, Folbp3 was mapped to a region on mouse chromosome 9 that is syntenic to human chromosome 19p13. As the chromosomal locations of Folbp1 murine and Folbp2 genes were previously unknown, we utilized the same approach and mapped both genes to a region of mouse chromosome 7 that is syntenic to the human FR loci on chromosome 11q13. PMID- 11111050 TI - Complete nucleotide sequence of the prophage VT1-Sakai carrying the Shiga toxin 1 genes of the enterohemorrhagic Escherichia coli O157:H7 strain derived from the Sakai outbreak. AB - Shiga toxins 1 and 2 (Stx1 and Stx2) are encoded by prophages lysogenized in enterohemorrhagic Escherichia coli (EHEC) O157:H7 strains. Lytic growth of the phage particles carrying the stx1 genes (stx1A and stx1B) of the EHEC O157:H7 strain RIMD 0509952, which was derived from the Sakai outbreak in 1996 in Japan, was induced after treatment with mitomycin C, but the plaque formation of the phage was not detected. We have determined the complete nucleotide sequence of the prophage VT1-Sakai. The integration site of the prophage was identified within the yehV gene at 47.7 min on the chromosome. The stx1 genes were downstream of the Q gene in the prophage genome, suggesting that their expression was regulated by the Q protein, the regulator of the late gene expression of the phage, which is similar to that of the stx1 or stx2 genes carried by the lambdoid phages reported previously. The sequences of the N gene and its recognition sites, nutL and nutR, were not homologous to those of the phages carrying the stx genes thus far reported, but they were very similar to those of bacteriophage phi21. The sequences of the repressor proteins, CI and Cro, that regulate expression of the early genes had low similarities with those of the known repressors of other phages, and their operator sequences were different from any sequence reported. These data suggest that multiple genetic recombination among bacteriophages with different immunities took place to generate the prophage VT1 Sakai. Comparison between the sequences of VT1-Sakai and lambda suggests that the ancestor of VT1-Sakai was produced by illegitimate excision, like lambda gal and bio phages. PMID- 11111051 TI - Identification of an alternatively spliced form of the Tat interactive protein (Tip60), Tip60(beta). AB - Tip60 was originally isolated as a Tat interactive protein. It was subsequently shown that Tip60 had histone acetyltransferase (HAT) activity. In studies to understand gene-expression regulation that might involve HAT activity, we PCR amplified Tip60 from a human heart marathon-ready cDNA library. As a result, we identified an alternatively spliced form of Tip60, Tip60beta (we refer to the previously cloned Tip60 as Tip60alpha). Tip60beta cDNA is slightly smaller than Tip60alpha, and sequencing indicates that there is a deletion of 156 bp in the coding region of the gene. The predicted Tip60beta protein therefore lacks 52 amino acids when compared with Tip60alpha. The Tip60alpha gene is encoded by 14 exons, and Tip60beta is an alternatively spliced form resulting from the exclusion of exon 5 during the splicing process. Exon 5 encodes a proline-rich region that is known to be important for protein-protein interaction. Tip60beta is expressed in a variety of human tissues and cell lines, and the protein is present in both the nucleus and cytoplasm in contrast to Tip60alpha, which is entirely nuclear. The results suggest that Tip60beta may have functions additional to those of Tip60alpha in cells and tissues. PMID- 11111052 TI - Light-modulated NADP-malate dehydrogenases from mossfern and green algae: insights into evolution of the enzyme's regulation. AB - Chloroplast NADP-dependent malate dehydrogenase is one of the best-studied light regulated enzymes. In C3 plants, NADP-MDH is a part of the 'malate valve' that controls the export of reducing equivalents in the form of malate to the cytosol. NADP-MDH is completely inactive in the dark and is activated in the light with reduced thioredoxin. Compared with its permanently active NAD-linked counterparts, NADP-MDH exhibits N- and C-terminal sequence extensions, each bearing one regulatory disulphide. Upon reduction of the C-terminal disulphide, the enzyme active site becomes accessible for the substrate. Reduction of the N terminal disulphide promotes a conformational change advantageous for catalysis. To trace the evolutionary development of this intricate regulation mechanism, we isolated cDNA clones for NADP-MDH from the mossfern Selaginella and from two unicellular green algae. While the NADP-MDH sequence from Selaginella demonstrates the classic cysteine pattern of the higher plant enzyme, the sequences from the green algae are devoid of the N-terminal regulatory disulphide. Phylogenetic analysis of new sequences and of those available in the databases led to the conclusion that the chloroplast NADP-MDH and the cytosolic NAD-dependent form arose via duplication of an ancestral eubacterial gene, which preceded the separation of plant and animal lineages. Redox-sensitive NADP-MDH activity was detected only in the 'green' plant lineage starting from the primitive prasinophytic algae but not in cyanobacteria, Cyanophora paradoxa, red algae and diatoms. The latter organisms therefore appear to utilize mechanisms other than the light-regulated 'malate valve' to remove from plastids excessive electrons produced by photosynthesis. PMID- 11111053 TI - Differential mRNA fingerprinting by preferential amplification of coding sequences. AB - The need for rapid identification of differentially expressed genes will persist even after the complete human genomic sequence becomes available. The most popular method for identifying differentially expressed genes acquires expressed sequence tags (ESTs) from the extreme 3' non-coding end of mRNAs. Such ESTs have limitations for downstream applications. We have developed a method, termed preferential amplification of coding sequences (PACS), that was applied to identify differentially expressed coding sequence tags (dCSTs) between osteoblasts and osteosarcoma cells. PACS was achieved by PCR with a set of primers to anchor at sequences complementary to AUG sequences in mRNAs and another set of primers to anchor at a PCR-amplifiable distance from AUG sequences. An initial screen identified 103 candidate dCSTs after screening approximately 15% of the expressed genes between the two cell types. Of these sequences, 27 represent CSTs of known genes and two are from 3'-ESTs of known mRNAs. Thus, PACS identified CSTs approximately 13.5 times more often than it identified 3' ESTs, attesting to the objective of the method. Since many of the dCSTs represent known genes, their identity and potential relevance to osteosarcoma could be immediately hypothesized. Differential expression of many of the dCSTs was further demonstrated by northern blotting or RT-PCR. Since PACS is not dependent on the existence of a poly A tail on an mRNA, it should have application to identify dCSTs for both prokaryotic and eukaryotic organisms. Additionally, PACS should aid in the identification of cell-specific or tissue specific genes and bidirectional acquisition of cDNA sequence enabling rapid retrieval of full-length cDNA sequence of novel genes. PMID- 11111054 TI - Human RNA-specific adenosine deaminase (ADAR1) gene specifies transcripts that initiate from a constitutively active alternative promoter. AB - The human ADAR1 gene specifies two size forms of RNA-specific adenosine deaminase, an interferon (IFN) inducible approximately 150 kDa protein and a constitutively expressed N-terminally truncated approximately 110 kDa protein, encoded by transcripts with alternative exon 1 structures that initiate from different promoters. We have now identified a new class of ADAR1 transcripts, with alternative 5'-structures and a deduced coding capacity for the approximately 110 kDa protein. Nuclease protection and 5'-rapid amplification of cDNA ends (5'-RACE) revealed five major ADAR1 transcriptional start sites that mapped within the previously identified and unusually large (approximately 1.6 kb) exon 2. These transcripts were observed with RNA from human amnion U cells and placenta tissue. Their abundance was not affected by IFN-alpha treatment of U cells in culture. Transfection analysis identified a functional promoter within human genomic DNA that mapped to the proximal exon 2 region of the ADAR1 gene. Promoter activity was not affected by IFN. These results suggest that transcripts encoding the constitutively expressed approximately 110 kDa form of the ADAR1 editing enzyme are initiated from multiple promoters, including one within exon 2, that collectively contribute to the high basal level of deaminase activity observed in nuclei of mammalian cells. PMID- 11111055 TI - Design, expression and functional characterization of a synthetic gene encoding the Chlamydia trachomatis major outer membrane protein. AB - A synthetic gene coding for the Chlamydia trachomatis serovar L2 major outer membrane protein (MOMP) was designed, constructed and expressed in Escherichia coli. The native amino acid sequence was reverse translated and the resulting nucleotide combinations manipulated in order to evenly distribute 25 unique restriction sites along the length of the gene while retaining the native amino acid sequence. The synthetic gene was cloned into a T7 promoter-controlled plasmid (pET-3a) and the expressed product was analyzed to assess antigenicity, cellular localization and function. Monoclonal antibodies specific for native MOMP reacted to the expressed product by immunoblot. Outer membrane fractionation confirmed that the processed protein was located in the outer membrane. MOMP expressed in E. coli and present in the outer membrane was shown to function as a general diffusion porin. This system provides the means to produce readily modifiable MOMP either in purified form or as a membrane-associated protein, and so facilitate the investigation of its functional, structural and antigenic properties. PMID- 11111056 TI - Two differentially spliced forms of topoisomerase IIalpha and beta mRNAs are conserved between birds and humans. AB - Screening of chicken cDNA libraries has identified four distinct forms of topoisomerase IIalpha and beta cDNAs. Two of these, designated topo IIalpha-1 and topo IIbeta-1, were previously deposited in the database. The other two, topo IIalpha-2 and topo IIbeta-2, are novel variants that appear to be conserved between chicken and human. Topo IIalpha-2 encodes a protein with an additional 35 amino acids inserted after K321 of the chicken topo IIalpha-1 protein sequence. Topo IIbeta-2 encodes a protein missing 86 amino acids following V27 in the topo IIbeta-1 protein sequence. We have also detected several alternatively spliced forms of human topo IIalpha. One of these, topo IIalpha-3, appears to correspond to chicken topo IIalpha-2. The other two are novel. The existence of these alternatively spliced forms in mature cytoplasmic RNA was confirmed by RT-PCR in several cell lines. Interestingly, these alternatively spliced forms carry sites for post-translational modification, suggesting that they may be subject to differential regulation from the canonical forms. These results suggest that cells express a more complex repertoire of topo II isoforms than previously thought, raising the possibility that different forms of topo II may fulfil specialized functions in chromosome dynamics. PMID- 11111057 TI - Albicidin antibiotic and phytotoxin biosynthesis in Xanthomonas albilineans requires a phosphopantetheinyl transferase gene. AB - Xanthomonas albilineans produces a family of highly potent antibiotics and phytotoxins called albicidins, which are key pathogenesis factors in the systemic development of leaf scald disease of sugarcane. A gene (xabA) required for albicidin biosynthesis encodes a peptide of 278 amino acids, including the signature sequence motifs for phosphopantetheinyl transferases (PPTases) that activate polyketide and non-ribosomal peptide synthetases. The Escherichia coli entD gene, which encodes a PPTase involved in biosynthesis of enterobactin (a siderophore), restored biosynthesis of albicidin (a DNA replication inhibitor) in X. albilineans Tox- LS156 (xabA::Tn5). We conclude that XabA is a PPTase required for post-translational activation of synthetases in the albicidin biosynthetic pathway. This is the first antibiotic or toxin biosynthesis gene characterized from the genus Xanthomonas, and the first demonstration that antibiotic synthetases in the Pseudomonadaceae, like those in the Enterobacteriaceae and in Gram-positive bacteria, can require activation by a PPTase. Coupled with the recent demonstration of a separate albicidin biosynthetic gene cluster, the results indicate the possibility for overproduction of albicidins,which allows better understanding and application of these potent inhibitors of prokaryote DNA replication. PMID- 11111058 TI - Friedreich's ataxia and iron metabolism. AB - The possible causes of abnormal iron metabolism in patients with Friedreich's ataxia are considered. Reduced expression of a frataxin homologue in yeast is associated with mitochondrial iron accumulation at the expense of cytosolic iron, and the same phenomenon can be demonstrated in these patients. A decrease in cytosolic iron causes the expression of a high-affinity iron-uptake protein, and therefore Friedreich's ataxia can be considered to be a disease of abnormal intracellular iron distribution. Friedreich's ataxia is of autosomal recessive inheritance, and the gene associated with it has been mapped to chromosome 9. This encodes the protein frataxin which regulates mitochondrial iron transport. The commonest mutation causing this disorder is an expanded GAA repeat in the gene for this protein. Different point mutations may account for some of the variations in the phenotypic features that are often found, and these variations are discussed. These findings have raised therapeutic possibilities in a condition for which previously there was no specific treatment. There are intracellular enzymes which are very sensitive to injury by oxygen-free radicals. Treatment has therefore been tried with ibebenone which acts as a free-radical scavenger, with some evidence of improvement. Iron chelating agents, such as deferoxamine, have also been given, but the finding of normal serum iron and ferritin casts doubt on the rationale of this. However the finding that the accumulation of iron in the mitochondria of the cells in patients with this form of ataxia will cause oxidative stress and cell death, gives hope for more effective treatment in the future, possibly with gene therapy. PMID- 11111059 TI - Neurotuberculosis among Filipino children: an 11 years experience at the Philippine Children's Medical Center. AB - Tuberculous meningitis (TBM) remains the most common form of neurotuberculosis in children. Four hundred and five cases of tuberculous meningitis (ages 3-156 months) seen at the Philippine Children's Medical Center (PCMC) from 1987 to 1998 were reviewed. Inclusion criteria include clinical and laboratory profile of TBM with pertinent evidence on imaging such as computed tomography and/or cranial sonography or histologic evidence of TBM. Nearly half of the cases were below age 2. The most common neurologic findings were altered sensorium, neck rigidity, motor and cranial deficits. The mortality rate was 16%. The neuropathologic findings in 31 autopsied cases were basal exudates in 100%, hydrocephalus in 71%, caseation necrosis in 68%, and 35% with infarcts. The most important determinant of outcome is the stage of illness at which the diagnosis is made and appropriate treatment is given. Although computed tomography was more definitive, cranial sonography was a very useful diagnostic tool considering the frequent occurrence below age 2. A short course (6 months) anti-tuberculous therapy for neurotuberculosis was shown to be adequate; shunting of cases with hydrocephalus did not show definite benefit. PMID- 11111060 TI - Schizencephaly: clinical and imaging features in 30 infantile cases. AB - Schizencephaly is an uncommon structural disorder of cerebral cortical development, characterized by congenital clefts spanning the cerebral hemispheres from the pial surface to the lateral ventricles and lined by cortical gray matter. Either an antenatal environmental incident or a genetic origin could be responsible for this lesion which occurs between the third and fourth month of gestation. We report the clinical and cranial imaging features of 30 children, of whom 15 had unilateral and 15 had bilateral lesions. Their ages at the time of the first presentation ranged from 1 month to 10 years. They were thoroughly studied from clinical, epileptical, imaging and electroencephalographic (EEG) viewpoints. Five patients were investigated by cranial computed tomography (CT), eight by cranial magnetic resonance (MR) imaging, and 17 by both methods. The clinical features consisted of mild hemiparesis in 17 cases (57%), 12/17 were related to a unilateral phenotype (80% of all unilateral forms) and 5/17 to a bilateral phenotype. A tetraparesis was present in nine cases, all of which were due to a bilateral cleft. Bilateral forms were significantly associated with tetraparesis, whereas unilateral forms were associated with hemiparesis. Mental retardation was observed in 17 cases (57%), and was observed significantly more often in bilateral clefts (80%). When both hemispheres are involved, an absence of reorganization of the brain function between the two hemispheres leads to severe mental deficits, in addition to the cerebral anomaly itself. Eleven patients had seizures (seven from unilateral and three from bilateral forms). The degree of malformation was not related to the severity of epilepsy. Migration disorders, such as dysplasia or heterotopia, were observed in 30% of cases and are also important etiopathogenetic factors. The septum pellucidum was absent in 13 cases (43%), with septo-optical dysplasia in two cases. Corpus callosum dysgenesis was noted in 30% of cases. Four cases of mega cisterna magna were noted. Although familial cases and environmental factors have been previously reported, schizencephaly appears to be, in the majority of cases, sporadic. PMID- 11111061 TI - Unprovoked seizures after complex febrile convulsions. AB - Children with complex febrile convulsions bear a higher risk of developing epilepsy than children with simple febrile convulsions. Complex febrile convulsions are defined by the presence of prolonged seizures, partial seizures and multiple seizures occurring during the same day. The aim of this study is to delineate the relative significance of each of the three criteria defining complex febrile convulsions. Fifty-seven out of 477 children (12%) admitted for febrile convulsions had complex febrile convulsions and normal neurological examination. Follow-up was available for 48 (84%) of them. Thirteen of these 48 (27%) had epilepsy at follow-up. The mean age of seizure onset among the patients with subsequent afebrile seizures was significantly lower than the rest (10.8 months versus 16.8 months). The patients with partial febrile convulsions showed a trend toward a higher risk (45%) of developing epilepsy than the patients with multiple febrile convulsions (21%). PMID- 11111062 TI - MRI spectrum of cortical malformations in tuberous sclerosis complex. AB - The diagnostic and prognostic value of magnetic resonance imaging in the tuberous sclerosis complex has increasingly been recognized. In this paper, we review the presumed pathogenesis of the cerebral dysgenesis seen in this condition in the light of magnetic resonance imaging features of selected patients. In addition to typical findings related to tubers, we show and discuss varied cortical malformations (from simple localized cortical dysplasia to transmantle dysplasia and schizencephaly) similar to those seen in sporadic cerebral dysgenesis. These cases support the hypothesis that the tuberous sclerosis complex focally affects the radial glial-neuronal complex as a basic unit for brain development. Abnormal stem cells would create dysplastic glia and neurons that fail to differentiate, proliferate, migrate and form a normally organized cortex. PMID- 11111063 TI - Siblings of Schwartz-Jampel syndrome with abnormal muscle computed tomographic findings. AB - Schwartz-Jampel syndrome (SJS) is a disorder characterized by myotonia, joint contractures, skeletal abnormalities, facial dysmorphism and growth retardation. We present two boys of ages 4 and 8 years with SJS. Their clinical, electromyographic and histopathological findings were similar to those described, except for computed tomography (CT) images that revealed diffuse high attenuation in sternocleidomastoid muscles and low attenuation in the paraspinal, quadriceps, sartorius, soleus and gastrocnemius muscles. This is the first report describing abnormal muscle CT findings associated with SJS. Additional studies of muscle CT might help to improve understanding of the pathogenesis of SJS. PMID- 11111064 TI - Mental retardation with rare fragile site expressed at 2q11. AB - Several rare autosomal folate sensitive fragile sites were reported in individuals with mental retardation, neurological abnormalities, and multiple congenital malformations. Only three of them: fra(11)(q22.3), fra(X)(q27.3) and fra(X)(q28), are known to be associated with mental retardation and phenotypic abnormalities. A possible association of the other rare fragile sites with idiopathic mental retardation is still being discussed. Here, a girl who has a fragile site at 2q11 with minor congenital anomalies and mental retardation is presented. This case has recalled the question of idiopathic mental retardation that might be the clinical expression of rare FSFS. Fragility was observed at 2q11 with a frequency of 3% in her cells along with a partial endoreduplication at 2 q11-->qter. PMID- 11111065 TI - Medical and economic effects of twin gestations. AB - OBJECTIVE: To determine the incidence and trends of twinning in the United States and to review the medical and economic effects of twin versus singleton gestations. METHODS: Pertinent and recent studies on twin gestations were obtained through a MEDLINE database search of the English language between December 1987 and December 1999. Data from the 1995-1996 National Center for Health Statistics were also used to compare gestational age at delivery, fetal growth restriction, and perinatal mortality for twin and singleton gestations. Studies that have evaluated perinatal risks in relation to advanced reproductive technology also were reviewed and summarized. The economic implications of twinning from a societal perspective and infant quality of life issues of twins compared with singleton gestations are reviewed. RESULTS: Due to delayed childbearing and increased use of reproductive technologies, the incidence of twin gestations in the United States has been increasing. Twin pregnancies have a higher risk of complications, including pregnancy-induced hypertension, anemia, antepartum and postpartum hemorrhage, and maternal mortality. In addition, twin infants are more likely to deliver preterm, have low birth weight and greater perinatal mortality rates. These outcomes influence health care costs and quality of life for both parents and children. CONCLUSIONS: Women carrying twin fetuses are at increased risk for perinatal and obstetric complications. The increased perinatal risks that accompany twin fetuses may be partly due to the increasing use of advanced reproductive technologies. The economic burdens, as well as the potential for decreased quality of life among twins, needs careful evaluation. PMID- 11111066 TI - Magnesium sulfate and fetal plasma concentrations of epinephrine, norepinephrine, and vasopressin in response to acute hypoxemia in goats. AB - OBJECTIVE: We measured fetal plasma concentrations of epinephrine, norepinephrine, and vasopressin during acute hypoxemia in goats and tested whether hypermagnesemia altered these endocrine responses. METHODS: Five chronically catheterized goat fetuses at 124-129 days' gestation were used. After 4 hours of infusion (magnesium or vehicle as controls), 30 minutes of hypoxemia was induced by infusing nitrogen gas through a maternal tracheal catheter. Fetal plasma concentrations of epinephrine, norepinephrine, and vasopressin were measured before and during hypoxemia. Both magnesium sulfate and vehicle infusions were performed in each animal. Repeated-measures analysis of variance (ANOVA) and two-way ANOVA with post hoc test were used to determine statistical significance. RESULTS: During hypoxemia, fetal PO(2) decreased significantly from 30 to 14 mmHg with no significant changes in fetal pH or PCO(2) in both groups. Fetal heart rate was reduced significantly by hypoxemia, but to a lesser extent in the magnesium group (change in decrease in fetal heart rate: 41 beats per minute [bpm] in controls versus 26 bpm with magnesium). Mean blood pressure did not change significantly during hypoxemia in both groups. Fetal plasma concentrations of epinephrine, norepinephrine, and vasopressin significantly increased from the prehypoxemic values both with magnesium and in controls. There were no significant differences in these hormone concentrations between magnesium and the controls. CONCLUSION: Magnesium sulfate had no effect on fetal plasma concentrations of vasopressin, epinephrine, and norepinephrine during acute hypoxemia. PMID- 11111067 TI - Hepatocyte growth factor concentration in maternal and umbilical cord blood samples and expression in fetal liver. AB - OBJECTIVE: To determine whether human placenta secretes hepatocyte growth factor (HGF) and could influence fetal liver development. METHODS: Expression of HGF and c-met mRNA in paired samples of first- and second-trimester fetal liver and placenta was compared using a quantitative ribonuclease protection assay. Serum HGF concentration in 30 samples of paired umbilical and maternal blood from term pregnancies was evaluated using an enzyme-linked immunosorbent assay. RESULTS: HGF and c-met mRNA were expressed at similar levels in liver and placenta, with expression increasing from 9 to 16 weeks' gestation. Median serum HGF values were 1.4 ng/mL (maternal venous), 1.2 ng/mL (cord venous), and 1.3 ng/mL (cord arterial). The maternal venous HGF levels were significantly higher than fetal venous levels (P =.02). CONCLUSIONS: This study does not support the hypothesis that the placenta secretes HGF, because maternal serum levels were higher than fetal and there was no significant difference between umbilical arterial and venous samples. Fetal liver expresses abundant HGF mRNA during the first and second trimester and expression increases in line with receptor (c-met) expression, suggesting that hepatic growth and development are independent of placental HGF. PMID- 11111068 TI - Plasma prostaglandin E(2) metabolite levels during labor induction with a sustained-release prostaglandin E(2) vaginal insert. AB - OBJECTIVE: To measure prostaglandin E(2) levels during labor induction with a sustained-release vaginal polymer insert (prostaglandin E(2) insert) and to determine whether Bishop score change correlated with tachysystole. METHODS: Twelve primiparas and 12 multiparas were treated with a 0.3 mg per hour sustained release polymer vaginal prostaglandin E(2) insert for up to 24 hours. Bishop score was assessed at start and end of therapy, and serum samples were collected at 4-hour intervals. Prostaglandin E(2) metabolite (PGEM) levels were measured by specific enzyme immunoassay. RESULTS: Exposure averaged 13.5 +/- 7.2 hours. Four patients (16.7%, three nulliparas) had tachysystole. Mean PGEM levels increased from 187 +/- 42 pg/mL at baseline to 548 +/- 110 pg/mL at 12 hours (P <.05) and remained relatively stable thereafter. Nulliparas with Bishop score changes of four points or more had the highest increase, with average peak levels of 985 +/- 109 pg/mL, compared with 452 +/- 58 pg/mL for all others (P <.001). Patients with tachysystole had higher 4-hour (P <.01) and overall (P <.04) increases in PGEM level. Removal of the insert led to an average decrease of 335 pg/mL in PGEM levels (P <.01). The decrease correlated with the PGEM level measured before removal (r =.94, P <.0001) and the maximum PGEM increase from baseline (r =.94, P <.0001). The mean mixed venous cord PGEM level was 409 +/- 375 pg/mL. CONCLUSION: Administration of the prostaglandin E(2) insert led to a sustained increase in circulating PGEM levels in women who had labor induction. Peak PGEM levels correlated with Bishop score improvement. Rapid increases in prostaglandin E(2) levels might cause tachysystole. PMID- 11111069 TI - Dopamine and morphine stimulate nitric oxide release in human endometrial glandular epithelial cells. AB - OBJECTIVE: Previous studies have shown that human endometrial glandular epithelial cells contain endothelial nitric oxide synthase indicating that the endometrium might produce nitric oxide (NO). We conducted this study to identify stimuli that can activate a transient NO release from endometrial glandular epithelial cells because NO is an important intracellular and intercellular signal transduction pathway in reproductive cycle. METHODS: Endometrial glandular epithelial cells, free of endothelial cells, were isolated from human endometrial specimens and maintained viable in RPMI 1640 medium with 2% fetal bovine serum for 2-4 days. Nitric oxide release from the glandular cells in response to stimuli was monitored continuously amperometrically. RESULTS: Among the substances examined, we found that dopamine and morphine stimulated a transient surge of NO production that was dose-dependent, whereas estrogen, progesterone, or relaxin (RLX) had no short-term effect on NO release. Cells treated with RLX or dopamine for 4 days enhanced the dopamine-induced NO release fourfold to sixfold, with the peak of the NO surge shifting from 35 to 15 seconds. CONCLUSION: Endometrial glandular cells were capable of producing NO. Dopamine and morphine were potent stimuli for a transient surge of NO release from endometrial glandular cells. Furthermore, prolonged exposure to dopamine or RLX enhanced the sensitivity of NO release in endometrial glands. PMID- 11111070 TI - Effect of hypoxia on stimulatory effect of TGF-beta 1 on MMP-2 and MMP-9 activities in mouse fibroblasts. AB - OBJECTIVE: Because chronic low-grade hypoxia has been implicated in the pathogenesis of fibrosis and postoperative adhesion formation, we hypothesized that hypoxia may modulate the effect of transforming growth factor-beta 1 (TGF beta 1) on matrix metalloproteinase (MMP)-2 and MMP-9 at both transcriptional and translational levels. METHODS: Mouse fibroblasts were placed in a hypoxic environment with or without 1 ng/mL TGF-beta 1 for varying periods of time. Zymography was performed on cell supernatants collected after each treatment. Gelatinolytic bands corresponding to MMP-2 and MMP-9 were quantiated by densitometry. Multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) was also performed for MMP-2 and MMP-9 on total RNA extracted from cells after each treatment. Analysis of PCR-amplified products was performed by 2% agarose gel followed by ethidium bromide staining of DNA bands. Scanning densometry was used to determine the ratio of intensity of each band relative to beta-actin. RESULTS: Hypoxia resulted in a 64% decrease in MMP-9 activity and 80% decrease in MMP-9 mRNA level but did not affect MMP-2 mRNA level or activity. TGF-beta 1 treatment resulted in 180% and 50% increases in MMP-2 and MMP-9 activities, respectively. Increases of 37.5% and 40% in MMP-2 and MMP-9 mRNA levels, respectively, were seen. However, under hypoxic conditions, TGF-beta1 resulted in a 160% increase and 45% decrease in MMP-2 and MMP-9 activities and a 37.5% increase and 71% decrease in MMP-2 and MMP-9 mRNA levels, respectively. CONCLUSION: Hypoxia suppresses the stimulatory effect of TGF-beta1 on the activity of MMP-9 but not MMP-2. This may suggest an important role for MMP-9 under hypoxic conditions in the pathogenesis of tissue fibrosis and postoperative adhesion formation. PMID- 11111071 TI - Constitutive androstane receptor expression in the rat cervix during gestation. AB - OBJECTIVE: To investigate expression of the constitutive androstane receptor (CAR) in the pregnant rat cervix. METHODS: Rat uterine tissue was obtained on gestational days 12, 16, 20, 21, and 22 (the day of parturition), and postpartum day 1. In addition, liver, lung, kidney, heart, and skeletal muscle tissue were obtained. Expression of the two known CAR isoforms was evaluated using reverse transcriptase-polymerase chain reaction. RESULTS: These studies confirmed CAR expression in the liver; however, CAR was not demonstrated in the myometrium or cervical tissue. CONCLUSIONS: The currently described CAR1 and CAR2 isoforms are not expressed in rat uterine tissue; therefore, they do not appear to participate in parturition in the pregnant rat. PMID- 11111073 TI - Cholesterol in the year 2000. AB - Cholesterol research was one of the key areas of scientific investigation in the 20th century. Little was known about the structure of cholesterol until the pioneering research of A. Windaus and H. Wieland in the first part of the century. The structure of cholesterol was completely elucidated in 1932. With the development of isotopic tracers in the 1930s studies on cholesterol biosynthesis were initiated. In 1942 K. Bloch and D. Rittenberg showed that deuterium-labeled acetate was incorporated into the ring structure and side chain of cholesterol. Another important discovery from Bloch's laboratory was that squalene was a precursor of cholesterol. In 1956, the main elements of the biosynthetic pathway became known when isopentenyl pyrophosphate was discovered as a precursor. In 1966, J. Cornforth and G. Popjak predicted that there were 16234 possible stereochemical pathways by which mevalonate could be converted into squalene. They subsequently showed which of these pathways was correct. In the 1970s and 1980s K. Bloch was able to provide intriguing evidence for an evolutionary advantage of cholesterol over lanosterol or some of the intermediates in the conversion of lanosterol to cholesterol. The last quarter of the 20th century was when M. Brown and J. Goldstein showed that the low density lipoprotein receptor was a key regulator of cholesterol homeostasis. They have also demonstrated that cholesterol balance in the cell is transcriptionally regulated via the sterol regulatory element binding protein. In the later part of the 20th century drugs were developed that effectively lower plasma cholesterol and lessen the risk of atherosclerosis and cardiovascular disease. PMID- 11111072 TI - Overexpression of testisin, a serine protease expressed by testicular germ cells, in epithelial ovarian tumor cells. AB - OBJECTIVE: In a continued effort to identify and characterize secreted proteases that are overexpressed in ovarian carcinomas, we discovered the testisin protease as such a candidate. When this discovery was originally made, no data existed in the literature or in the GenBank database that identified such a gene. Our main objective was to determine whether this gene was overexpressed exclusively in ovarian tumor tissues compared with normal ovary and whether it was expressed in any other normal tissues. METHODS: mRNA was isolated and cDNA was prepared from 34 ovarian tumors (four adenomas, three low malignant potential tumors, and 27 carcinomas) and seven normal ovaries. The testisin mRNA expression level relative to internal control, beta-tubulin, was determined by Northern blot analysis and semiquantitative polymerase chain reaction (PCR). RESULTS: Northern blot hybridization showed that the testisin transcript was abundant in ovarian carcinoma but was not detected in normal ovary. On examination of Northern blots from normal fetal and adult tissues, only adult testis showed abundant transcripts of testisin. Semiquantitative PCR examination showed that the testisin mRNA levels in ovarian tumors of low malignant potential and in ovarian carcinomas were significantly higher than in normal ovaries (P <.01). Testisin mRNA level in ovarian carcinomas was also significantly higher than in ovarian adenomas (P <.05). Testisin overexpression rates in advanced stage (stage 2 or 3) diseases were significantly higher than that in early stage diseases (stage 1) in ovarian carcinoma samples (P <.05). CONCLUSIONS: The induction of the testisin transcript might contribute to the development, progression, and invasive or metastatic capacity of ovarian carcinomas. PMID- 11111074 TI - The structure of the catalytic portion of human HMG-CoA reductase. AB - In higher plants, fungi, and animals isoprenoids are derived from the mevalonate pathway. The carboxylic acid mevalonate is formed from acetyl-CoA and acetoacetyl CoA via the intermediate 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA). The four-electron reduction of HMG-CoA to mevalonate, which utilizes two molecules of NADPH, is the committed step in the biosynthesis of isoprenoids. This reaction is catalyzed by HMG-CoA reductase (HMGR). The activity of HMGR is controlled through synthesis, degradation and phosphorylation. The human enzyme has also been targeted successfully by drugs, known as statins, in the clinical treatment of high serum cholesterol levels. The crystal structure of the catalytic portion of HMGR has been determined recently with bound reaction substrates and products. The structure illustrates how HMG-CoA and NADPH are recognized and suggests a catalytic mechanism. Catalytic portions of human HMGR form tight tetramers, explaining the influence of the enzyme's oligomeric state on the activity and suggesting a mechanism for cholesterol sensing. PMID- 11111075 TI - Biochemical and genetic aspects of mevalonate kinase and its deficiency. AB - Mevalonate kinase (MK) is an essential enzyme in the mevalonate pathway which produces numerous cellular isoprenoids. The enzyme has been characterized both at the biochemical and the molecular level in a variety of organisms. Despite the fact that mevalonate kinase is not the rate-limiting enzyme in isoprenoid biosynthesis, its activity is subject to feedback regulation by the branch-point intermediates geranyldiphosphate, farnesyldiphosphate and geranylgeranyldiphosphate. Recently, the importance of mevalonate kinase was demonstrated by the identification of its deficiency as the biochemical and molecular cause of the inherited human disorders mevalonic aciduria and hyperimmunoglobulinemia D and periodic fever syndrome. The pathophysiology of these disorders is not yet understood, but eventually will give insight into the in vivo role of mevalonate kinase and isoprenoid biosynthesis with respect to the acute phase response and fever. The subcellular localization of mevalonate kinase is still a matter of debate. The enzyme could be localized predominantly in the cytosol, or in peroxisomes, or it is associated differentially with peroxisomes. Here we review the biochemical and molecular properties of MK, and discuss its biological significance, the regulation of its enzyme activity and finally its subcellular localization. PMID- 11111076 TI - Isoprenyl diphosphate synthases. AB - Isoprenyl diphosphate synthases catalyze consecutive condensations of isopentenyl diphosphates with allylic primer substrates to form linear backbones for all isoprenoid compounds including cholesterol. These synthases are classified according to the final chain length of their end products and the stereochemistry of the newly formed double bonds. Mutagenesis and X-ray crystallography data have uncovered the basic catalytic and chain length determination mechanisms of E isoprenyl diphosphate synthases and shed light on their possible evolutionary course. Although much less is known about the Z-isoprenyl diphosphate synthase family, successful cloning and subsequent crystallizations in the near future will no doubt bring more insight as researchers begin to unravel the essential components and precise reaction mechanisms of this cellular machinery. PMID- 11111077 TI - Structure and regulation of mammalian squalene synthase. AB - Mammalian squalene synthase (SQS) catalyzes the first reaction of the branch of the isoprenoid metabolic pathway committed specifically to sterol biosynthesis. SQS produces squalene in an unusual two-step reaction in which two molecules of farnesyl diphosphate are condensed head-to-head. Recent studies have advanced understanding of the reaction mechanism, the functional domains of the enzyme, and transcriptional regulation of the gene. Site-directed mutagenesis has identified conserved Asp, Tyr, and Phe residues that are essential for SQS activity. The Asp residues are hypothesized to be required for substrate binding; the Tyr and Phe residues may stabilize carbocation reaction intermediates. The elucidation of SQS crystal structure will most likely direct future research on the relationship between enzyme structure and function. SQS activity, protein, and mRNA levels are regulated by cholesterol status and by the cytokines TNF alpha and IL-1beta. Activation of the SQS promoter in response to cholesterol deficit is mediated by sterol regulatory element binding proteins SREBP-1a and SREBP-2. The precise contributions made by individual SREBPs and accessory transcription factors to SQS transcriptional control, and the mechanisms underlying cytokine regulation of SQS are major foci of current research. PMID- 11111078 TI - Sterol methyl transferase: enzymology and inhibition. AB - Sterol C-methylations catalyzed by the (S)-adenosyl-L-methionine: Delta(24) sterol methyl transferase (SMT) have provided the focus for study of electrophilic alkylations, a reaction type of functional importance in C-C bond formation of natural products. SMTs occur generally in nature, but do not occur in animal systems, suggesting that the difference in sterol synthetic pathways can be exploited therapeutically and in insect-plant interactions. The SMT genes from several plants and fungi have been cloned, sequenced and expressed in bacteria or yeast and bioengineered into tobacco or tomato plants. These enzymes share significant amino acid sequence similarity in the putative sterol and AdoMet binding sites. Investigations of the molecular recognition of sterol fitness and studies with stereospecifically labeled substrates as well as various sterol analogs assayed with native or mutant SMTs from fungi and plants have been carried out recently in our own and other laboratories. These analyses have led to an active-site model, referred to as the 'steric-electric plug' model, which is consistent with a non-covalent mechanism involving the intermediacy of a 24beta-methyl (or ethyl) sterol bound to the ternary complex. Despite the seeming differences between fungal and plant SMT activities the recent data indicate that a distinct SMT or family of SMTs exist in these organisms which bind and transform sterols according to a similar mechanistic plan. Vascular plants have been found to express different complements of C(1)/C(2)-activities in the form of at least three SMT isoforms. This enzyme multiplicity can be a target of regulatory control to affect phytosterol homeostasis in transgenic plants. The state of our current understanding of SMT enzymology and inhibition is presented. PMID- 11111079 TI - Peroxisomal protein targeting and identification of peroxisomal targeting signals in cholesterol biosynthetic enzymes. AB - At least three different subcellular compartments, including peroxisomes, are involved in cholesterol synthesis. Recently, it has been demonstrated that peroxisomes contain a number of enzymes involved in cholesterol biogenesis that previously were considered to be cytosolic or located in the endoplasmic reticulum. Peroxisomes have been shown to contain acetoacetyl-CoA thiolase, HMG CoA synthase, HMG-CoA reductase, mevalonate kinase, phosphomevalonate kinase, phosphomevalonate decarboxylase, isopentenyl diphosphate isomerase and FPP synthase. Moreover, the activities of these enzymes are also significantly decreased in liver tissue and fibroblast cells obtained from patients with peroxisomal deficiency diseases. In addition, the cholesterol biosynthetic capacity is severely impaired in cultured skin fibroblasts obtained from patients with peroxisomal deficiency diseases. These findings support the proposal that peroxisomes play an essential role in isoprenoid biosynthesis. This paper presents a review of peroxisomal protein targeting and of recent studies demonstrating the localization of cholesterol biosynthetic enzymes in peroxisomes and the identification of peroxisomal targeting signals in these proteins. PMID- 11111080 TI - Regulation of gene expression by SREBP and SCAP. AB - Sterol regulatory element binding proteins (SREBPs) function as transcription factors that activate specific genes involved in cholesterol synthesis, endocytosis of low density lipoproteins, the synthesis of both saturated and unsaturated fatty acids and glucose metabolism. As such, these proteins provide a link between lipid and carbohydrate metabolism. There are three SREBPs, SREBP-1a, SREBP-1c and SREBP-2, that are encoded by two genes. SREBPs are synthesized as 125 kDa precursor proteins that are localized to the endoplasmic reticulum. The precursor is transported to the Golgi by a chaperone protein (SREBP-cleavage activating protein) and then cleaved by two proteases to release the mature, transcriptionally active 68 kDa amino terminal domain. Recent studies have shown that formation of mature SREBP is controlled at multiple levels in response to changes in the levels of oxysterols, insulin/glucose and polyunsaturated fatty acids. These recent findings have important clinical implications relevant to hyperlipidemia and diabetes and are the topic of this review. PMID- 11111081 TI - Sterols and gene expression: control of affluence. AB - Intracellular and extracellular cholesterol levels are tightly maintained within a narrow concentration range by an intricate transcriptional control mechanism. Excess cholesterol can be converted into oxysterols, signaling molecules, which modulate the activity of a number of transcription factors, as to limit accumulation of excess of cholesterol. Two key regulatory pathways are affected by oxysterols. The first pathway involves the uptake and de novo synthesis of cholesterol and is controlled by the family of sterol response element binding proteins, whose activity is regulated by a sterol-dependent feedback mechanism. The second pathway, which only recently has become a topic of interest, involves the activation by a feedforward mechanism of cholesterol utilization for either bile acid or steroid hormone synthesis by oxysterol-activated nuclear receptors, such as liver X receptor and steroidogenic factor-1. Furthermore, biosynthesis and enterohepatic reabsorption of bile acids are regulated by the farnesol X receptor, a receptor activated by bile acids. Both the feedback inhibition of cholesterol uptake and production and the stimulation of cholesterol utilization will ultimately result in a lowering of the intracellular cholesterol concentration and allow for a fine-tuned regulation of the cholesterol concentration. PMID- 11111082 TI - Oxysterol biosynthetic enzymes. AB - Oxysterols, herein defined as derivatives of cholesterol with a hydroxyl group on the side chain, play several roles in lipid metabolism. Members of this class regulate the expression of genes that participate in both sterol and fat metabolism, serve as substrates for the synthesis of bile acids, and are intermediates in the transfer of sterols from the periphery to the liver. Three abundant naturally occurring oxysterols are 24-hydroxycholesterol, 25 hydroxycholesterol, and 27-hydroxycholesterol. The cholesterol hydroxylase enzymes that synthesize each of these have been isolated over the last several years and their study has produced insight into the biology of oxysterols. This article focuses on the properties of these enzymes. PMID- 11111083 TI - Biologic role(s) of the 25(R),26-hydroxycholesterol metabolic pathway. AB - Expression of the gene coding for the synthesis of 25(R), 26-hydroxycholesterol in many tissues and the finding that this sterol can be the sole pathway for the production of bile acids have led to a renewed interest in this metabolic pathway. A further impetus for exploring the normal biologic roles that are served by expression of the CYP27A1 gene is the knowledge that mutations in humans are associated with accelerated atherosclerosis and with severe neurologic impairment. The molecular mechanisms governing these phenotypic expressions are not known but in light of the traditional role of steroids as ligands for receptors that regulate gene expression it seems likely that the intermediates in this pathway modulate a number of enzymatic activities that remain to be elucidated. PMID- 11111084 TI - Mammalian acyl-CoA:cholesterol acyltransferases. AB - Cholesterol, the chief sterol found in vertebrates, exists both as a free sterol and as a component of cholesterol esters, which are synthesized by acyl CoA:cholesterol acyltransferase (ACAT) enzymes. Considerable knowledge concerning cholesterol ester metabolism has accumulated during the past century. However, rapid advances have occurred in the past 7 years since the cloning of an ACAT gene, including the discovery that two ACATs function in mammalian biology. A clearer picture of the functions of ACAT enzymes in cellular cholesterol metabolism and physiologic processes is now emerging. These insights may have relevance for the development of ACAT inhibitors for treating hypercholesterolemia or atherosclerosis in humans. PMID- 11111085 TI - Conservation of eukaryotic sterol homeostasis: new insights from studies in budding yeast. AB - The model eukaryote Saccharomyces cerevisiae (budding yeast) has provided significant insight into sterol homeostasis. The study of sterol metabolism in a genetically amenable model organism such as yeast is likely to have an even greater impact and relevance to human disease with the advent of the complete human genome sequence. In addition to definition of the sterol biosynthetic pathway, almost to completion, the remarkable conservation of other components of sterol homeostasis are described in this review. PMID- 11111086 TI - Cholesterol and phospholipid metabolism in macrophages. AB - Cholesterol-loaded macrophages are present at all stages of atherogenesis, and recent in vivo data indicate that these cells play important roles in both early lesion development and late lesion complications. To understand how these cells promote atherogenesis, it is critical that we understand how lesional macrophages interact with subendothelial lipoproteins, the consequences of this interaction, and the impact of subsequent intracellular metabolic events. In the arterial wall, macrophages likely interact with both soluble and matrix-retained lipoproteins, and a new challenge is to understand how certain consequences of these two processes might differ. Initially, the major intracellular metabolic route of the lipoprotein-derived cholesterol is esterification to fatty acids, but macrophages in advanced atherosclerotic lesions progressively accumulate large amounts of unesterified, or free, cholesterol (FC). In cultured macrophages, excess FC accumulation stimulates phospholipid biosynthesis, which is an adaptive response to protect the macrophage from FC-induced cytotoxicity. This phospholipid response eventually decreases with continued FC loading, leading to a series of cellular death reactions involving both death receptor induced signaling and mitochondrial dysfunction. Because macrophage death in advanced lesions is thought to promote plaque instability, these intracellular processes involving cholesterol, phospholipid, and death pathways may play a critical role in the acute clinical manifestations of advanced atherosclerotic lesions. PMID- 11111087 TI - Steroidogenic acute regulatory protein (StAR) and the intramitochondrial translocation of cholesterol. AB - The steroidogenic acute regulatory (StAR) protein regulates the rate limiting step in steroidogenesis, the transport of cholesterol from the outer to the inner mitochondrial membrane. Insight into the structure and function of StAR was attained through molecular genetic studies of congenital lipoid adrenal hyperplasia, a rare disease caused by mutations in the StAR gene. Subsequent functional analysis defined two major domains within the StAR protein, the N terminal mitochondrial targeting sequence and the C-terminus, which promotes the translocation of cholesterol between the two mitochondrial membranes. Two models of StAR's mechanism of action, (1) stimulation of cholesterol desorption from the outer mitochondrial membrane and (2) an intermembrane shuttle hypothesis, are discussed with respect to the known biochemical and biophysical events associated with the process of steroidogenesis and the structure of StAR. StAR gene expression is regulated primarily at the transcriptional level, and the roles of transcription factors that govern basal and cAMP-dependent StAR expression including SF-1, C/EBP beta, Sp1 and GATA-4 are reviewed. PMID- 11111088 TI - Cholesterol modification of proteins. AB - The demonstration over 30 years ago that inhibitors of cholesterol biosynthesis disrupt animal development suggested an intriguing connection between fundamental cellular metabolic processes and the more global processes of embryonic tissue patterning. Adding a new dimension to this relationship is the more recent finding that the Hedgehog family of tissue patterning factors are covalently modified by cholesterol. Here we review the mechanism of the Hedgehog autoprocessing reaction that results in this modification, and compare this reaction to that undergone by other autoprocessing proteins. We also discuss the biological consequences of cholesterol modification, in particular the use of cholesterol as a molecular handle in the spatial deployment of the protein signal in developing tissues. Finally, the developmental consequences of chemical and genetic disruption of cholesterol homeostasis are summarized, along with the potential importance of cholesterol-rich lipid rafts in production of and response to the Hh signal. PMID- 11111089 TI - Functional aspects of polyisoprenoid protein substituents: roles in protein protein interaction and trafficking. AB - There are now numerous examples of post-translational modification with geranylgeranyl or farnesyl substituents. Once thought of as solely a mechanism for association of proteins with membranes, other functional aspects of protein prenylation have come to be appreciated. Although, in almost all instances, such proteins are membrane associated, they are often found to also engage in protein protein interactions. In some instances, such interactions are critical aspects of prenylated protein trafficking. In this review, the role of prenylation in mediating protein-protein interactions will be considered. The hypothesis will be developed that such interactions occur through recognition of the prenyl group and a second domain, on the prenylated protein, by a heterodimeric protein partner. PMID- 11111090 TI - Cholesterol and caveolae: structural and functional relationships. AB - Caveolae are free cholesterol (FC)- and sphingolipid-rich surface microdomains abundant in most peripheral cells. Caveolin, a FC binding protein, is a major structural element of these domains. Caveolae serve as portals to regulate cellular FC homeostasis, possibly via their association with ancillary proteins including scavenger receptor B1. The FC content of caveolae regulates the transmission of both extracellular receptor-mediated and endogenous signal transduction via changes in the composition of caveolin-associated complexes of signaling intermediates. By controlling surface FC content, reporting membrane changes by signal transduction to the nucleus, and regulating signal traffic in response to extracellular stimuli, caveolae exert a multifaceted influence on cell physiology including growth and cell division, adhesion, and hormonal response. Cell surface lipid 'rafts' may assume many of the functions of caveolae in cells with low levels of caveolin. PMID- 11111091 TI - Cholesterol and hepatic lipoprotein assembly and secretion. AB - The assembly and secretion of apo B100 containing lipoproteins (i.e., VLDL) by the liver and cholesterol metabolism are interrelated on several different levels and for several different physiologic reasons. Firstly, hepatic VLDL is the major precursor for LDL, which in the human is the major vehicle responsible for transporting cholesterol to peripheral tissues. Secondly, cholesterol is supplied to many tissues by a specific uptake of LDL via LDL receptor, which is expressed in a regulated manner by most mammalian tissues. Thirdly, the rate of hepatic cholesterol biosynthesis and metabolism to bile acids correlates with production of VLDL. This apparent coordinate expression of cholesterol biosynthetic/catabolic enzymes and hepatic VLDL assembly/secretion are mediated at least in part through the sterol response element binding protein (SREBP) transcription factor family. Their gene targets include a plethora of enzymes that regulate glycolysis, energy production, lipogenesis and cholesterol catabolism. Studies of hepatoma cells overexpressing CYP7A1, the rate-limiting enzyme controlling bile acid synthesis, show that as a result of increased mature SREBP1, there is a coordinate induction of lipogenesis and the assembly and secretion of VLDL. These and additional studies show that the bile acid synthetic pathway and the VLDL assembly/secretion pathway are coordinately linked through SREBP-dependent transcription. Based on studies showing that within the liver acinus, the expression of CYP7A1 is mainly in the pericentral region while HMG CoA reductase is mainly periportal, we propose that a 'metabolic zonal segregation' plays an important role in coordinate regulation of cholesterol and VLDL metabolism. This putative 'metabolic zonal segregation' may provide segregation of metabolic functions which may be mutually antagonistic. For example, there may be physiologic states in which the bile acid synthetic pathway may compete with the VLDL assembly/secretion pathway for a limited amount of cholesterol. Metabolic antagonism (e.g., competition for cholesterol) may be avoided via inducing SREBP-mediated transcription. Adaptation of catabolic hepatocytes to accommodate the expression of VLDL assembly/secretion may occur in response to activation of SREBP-mediated transcription. Support for these is discussed. PMID- 11111092 TI - Release of cellular cholesterol: molecular mechanism for cholesterol homeostasis in cells and in the body. AB - Most mammalian somatic cells are unable to catabolize cholesterol and therefore need to export it in order to maintain sterol homeostasis. This mechanism may also function to reduce excessively accumulated cholesterol, which would thereby contribute to prevention or cure of the initial stage of atherosclerotic vascular lesion. High-density lipoprotein (HDL) has been believed to play a main role in this reaction based on epidemiological evidence and in vitro experimental data. At least two independent mechanisms are identified for this reaction. One is non specific diffusion-mediated cholesterol 'efflux' from cell surface. Cholesterol molecules desorbed from cells can be trapped by various extracellular acceptors including various lipoproteins and albumin, and extracellular cholesterol esterification mainly on HDL may provide a driving force for the net removal of cell cholesterol by maintaining a cholesterol gradient between lipoprotein surface and cell membrane. The other is apolipoprotein-mediated process to generate new HDL by removing cellular phospholipid and cholesterol. The reaction is initiated by the interaction of lipid-free or lipid-poor helical apolipoproteins with cellular surface resulting in assembly of HDL particles with cellular phospholipid and incorporation of cellular cholesterol into the HDL being formed. Thus, HDL has dual functions as an active cholesterol acceptor in the diffusion-mediated pathway and as an apolipoprotein carrier for the HDL assembly reaction. The impairment of the apolipoprotein-mediated reaction was found in Tangier disease and other familial HDL deficiencies to strongly suggest that this is a main mechanism to produce plasma HDL. The causative mutations for this defect was identified in ATP binding cassette transporter protein A1, as a significant step for further understanding of the reaction and cholesterol homeostasis. PMID- 11111093 TI - Lecithin cholesterol acyltransferase. AB - Cholesterol transport in circulation and its removal from tissues depends on the activity of lecithin cholesterol acyltransferase (LCAT). LCAT is a soluble enzyme that converts cholesterol and phosphatidylcholines (lecithins) to cholesteryl esters and lyso-phosphatidylcholines on the surface of high-density lipoproteins. This review presents key background information and recent research advances on the structure of human LCAT, its reactions and substrates, and the expression of the LCAT gene. While the three-dimensional structure of LCAT is not yet known, a partial model now exists that facilitates the study of structure-function relationships of the native enzyme, and of natural and engineered mutants. The LCAT reaction on lipoproteins consists of several steps, starting with enzyme binding to the lipoprotein/lipid surface, followed by activation of LCAT by apolipoproteins, binding of lipid substrates and the catalytic steps giving rise to the lipid products. Quantitative data are presented on the kinetic and equilibrium constants of some of the LCAT reaction steps. Finally, overexpression of the human LCAT gene in mice and rabbits has been used to examine the physiologic role of LCAT in vivo and its protective effect against diet induced atherosclerosis. PMID- 11111094 TI - Molecular biology and pathophysiological aspects of plasma cholesteryl ester transfer protein. AB - Plasma cholesteryl ester transfer protein (CETP) facilitates the transfer of cholesteryl ester (CE) from high density lipoprotein (HDL) to apolipoprotein B containing lipoproteins. Since CETP regulates the plasma levels of HDL cholesterol and the size of HDL particles, CETP is considered to be a key protein in reverse cholesterol transport, a protective system against atherosclerosis. CETP, as well as plasma phospholipid transfer protein, belongs to members of the lipid transfer/lipopolysaccharide-binding protein (LBP) gene family, which also includes the lipopolysaccharide-binding protein (LBP) and bactericidal/permeability-increasing protein. Although these four proteins possess different physiological functions, they share marked biochemical and structural similarities. The importance of plasma CETP in lipoprotein metabolism was demonstrated by the discovery of CETP-deficient subjects with a marked hyperalphalipoproteinemia (HALP). Two common mutations in the CETP gene, intron 14 splicing defect and exon 15 missense mutation (D442G), have been identified in Japanese HALP patients with CETP deficiency. The deficiency of CETP causes various abnormalities in the concentration, composition, and functions of both HDL and low density lipoprotein. Although the pathophysiological significance of CETP in terms of atherosclerosis has been controversial, the in vitro experiments showed that large CE-rich HDL particles in CETP deficiency are defective in cholesterol efflux. Epidemiological studies in Japanese-Americans and in the Omagari area where HALP subjects with the intron 14 splicing defect of CETP gene are markedly frequent, have shown an increased incidence of coronary atherosclerosis in CETP-deficient patients. The current review will focus on the recent findings on the molecular biology and pathophysiological aspects of plasma CETP, a key protein in reverse cholesterol transport. PMID- 11111095 TI - Cellular and physiological roles of SR-BI, a lipoprotein receptor which mediates selective lipid uptake. AB - High-density lipoproteins (HDL) play an important role in protection against atherosclerosis by mediating reverse cholesterol transport - the transport of excess cholesterol from peripheral tissues to the liver for disposal. SR-BI is a cell surface receptor for HDL and other lipoproteins (LDL and VLDL) and mediates the selective uptake of lipoprotein cholesterol by cells. Overexpression or genetic ablation of SR-BI in mice revealed that it plays an important role in HDL metabolism and reverse cholesterol transport and protects against atherosclerosis in mouse models of the disease. If it plays a similar role in humans then it may be an attractive target for therapeutic intervention. We will review some of the recent advances in the understanding of SR-BI's physiological role and cellular function in lipoprotein metabolism. PMID- 11111096 TI - From cholesterol transport to signal transduction: low density lipoprotein receptor, very low density lipoprotein receptor, and apolipoprotein E receptor-2. AB - The discovery of an ever growing number of low density lipoprotein (LDL) receptor gene family members has triggered research into many different directions. Here we first summarize the results of classical studies on the role of the LDL receptor in cholesterol transport, the structure/function relationships delineated with the help of LDL receptor mutations in familial hypercholesterolemia, and the elegant way in which cells regulate cholesterol at the transcriptional level. The second part deals with a multifunctional, structurally very close relative, the very low density lipoprotein (VLDL) receptor. While it is involved in lipoprotein transport in certain tissues and species, detailed studies on its function have generated new knowledge about the growing spectrum of ligands and about exciting and unexpected aspects of receptor biology. In particular, these investigations have elucidated the roles of LDL receptor gene family members in ligand-mediated signal transduction. In the third part of this review article, we provide first insight into the roles of the VLDL receptor and of another small relative, the so-called apolipoprotein E receptor 2, in such signaling processes. These findings suggest that to date, only the tip of an iceberg has been uncovered. PMID- 11111097 TI - Cholesterol and atherosclerosis. PMID- 11111098 TI - Dietary cholesterol and atherosclerosis. AB - The perceived relationship between dietary cholesterol, plasma cholesterol and atherosclerosis is based on three lines of evidence: animal feeding studies, epidemiological surveys, and clinical trials. Over the past quarter century studies investigating the relationship between dietary cholesterol and atherosclerosis have raised questions regarding the contribution of dietary cholesterol to heart disease risk and the validity of dietary cholesterol restrictions based on these lines of evidence. Animal feeding studies have shown that for most species large doses of cholesterol are necessary to induce hypercholesterolemia and atherosclerosis, while for other species even small cholesterol intakes induce hypercholesterolemia. The species-to-species variability in the plasma cholesterol response to dietary cholesterol, and the distinctly different plasma lipoprotein profiles of most animal models make extrapolation of the data from animal feeding studies to human health extremely complicated and difficult to interpret. Epidemiological surveys often report positive relationships between cholesterol intakes and cardiovascular disease based on simple regression analyses; however, when multiple regression analyses account for the colinearity of dietary cholesterol and saturated fat calories, there is a null relationship between dietary cholesterol and coronary heart disease morbidity and mortality. An additional complication of epidemiological survey data is that dietary patterns high in animal products are often low in grains, fruits and vegetables which can contribute to increased risk of atherosclerosis. Clinical feeding studies show that a 100 mg/day change in dietary cholesterol will on average change the plasma total cholesterol level by 2.2-2.5 mg/dl, with a 1.9 mg/dl change in low density lipoprotein (LDL) cholesterol and a 0.4 mg/dl change in high density lipoprotein (HDL) cholesterol. Data indicate that dietary cholesterol has little effect on the plasma LDL:HDL ratio. Analysis of the available epidemiological and clinical data indicates that for the general population, dietary cholesterol makes no significant contribution to atherosclerosis and risk of cardiovascular disease. PMID- 11111099 TI - Tangier disease and ABCA1. AB - Tangier disease is an autosomal recessive genetic disorder characterized by a severe high-density lipoprotein (HDL) deficiency, sterol deposition in tissue macrophages, and prevalent atherosclerosis. Mutations in the ATP binding cassette transporter ABCA1 cause Tangier disease and other familial HDL deficiencies. ABCA1 controls a cellular pathway that secretes cholesterol and phospholipids to lipid-poor apolipoproteins. This implies that an inability of newly synthesized apolipoproteins to acquire cellular lipids by the ABCA1 pathway leads to their rapid degradation and an over-accumulation of cholesterol in macrophages. Thus, ABCA1 plays a critical role in modulating flux of tissue cholesterol and phospholipids into the reverse cholesterol transport pathway, making it an important therapeutic target for clearing excess cholesterol from macrophages and preventing atherosclerosis. PMID- 11111100 TI - Niemann-Pick type C: a disorder of cellular cholesterol trafficking. PMID- 11111101 TI - Biochemical and genetic aspects of 7-dehydrocholesterol reductase and Smith-Lemli Opitz syndrome. AB - In recent years, several inherited human disorders caused by defects in cholesterol biosynthesis have been identified. These are characterized by malformations, multiple congenital anomalies, mental and growth retardation and/or skeletal and skin abnormalities indicating a pivotal role of cholesterol in morphogenesis and embryonic development. The first recognized and most common of these developmental disorders is Smith-Lemli-Opitz syndrome, an autosomal recessive trait caused by mutations in the DHCR7 gene resulting in a deficiency of the encoded sterol Delta(7)-reductase, alternatively called 7 dehydrocholesterol reductase (EC 1.3.1.21). This enzyme catalyzes the final step in cholesterol biosynthesis, which is the reduction of the Delta(7) double bond of 7-dehydrocholesterol to produce cholesterol. PMID- 11111102 TI - X-Linked dominant disorders of cholesterol biosynthesis in man and mouse. AB - The X-linked dominant male-lethal mouse mutations tattered and bare patches are homologous to human X-linked dominant chondrodysplasia punctata and CHILD syndrome, rare human skeletal dysplasias. These disorders also affect the skin and can cause cataracts and microphthalmia in surviving, affected heterozygous females. They have recently been shown to result from mutations in genes encoding enzymes involved in sequential steps in the conversion of lanosterol to cholesterol. This review will summarize clinical features of the disorders and describe recent biochemical and molecular investigations that have resulted in the elucidation of the involved genes and their metabolic pathway. Finally, speculations about possible mechanisms of pathogenesis will be provided. PMID- 11111103 TI - A systematic review to evaluate the effectiveness of interventions to promote the initiation of breastfeeding. AB - BACKGROUND: Human breastmilk provides complete nutrition for infants and helps protect against certain childhood diseases. Despite this, rates of initiation of breastfeeding in the UK remain low relative to other countries. In 'Our healthier nation' action report, the government has highlighted the promotion of breastfeeding in order to assist improvements in health and to reduce the health inequalities of mothers and children in the UK. OBJECTIVES: The primary aim of this systematic review was to evaluate existing evidence to identify which promotion programmes are effective at increasing the number of women who start to breastfeed. In addition, the review aimed to assess the impact of such programmes on the duration and/or exclusivity of breastfeeding and the intermediate and process outcomes. Where the strength and quality of the evidence permitted, the review aimed to identify implications for practice within the UK and priority areas for future research. METHODS: DATA SOURCES: A range of electronic databases were searched from inception to November 1998, several relevant journals were hand-searched, and references of retrieved papers were examined. Relevant experts, organisations and lay groups were contacted to help identify further published or unpublished material. Additionally, an expert panel was consulted. SELECTION CRITERIA: Four types of criteria were used to select eligible studies for this review: STUDY DESIGN - randomised controlled trials (RCTs), non-RCTs with concurrent controls, and before-after studies (cohort or cross-sectional). PARTICIPANTS - pregnant women, mothers in the immediate postpartum period before the first breastfeed, any participant linked to pregnant women or new mothers, or any participant who may breastfeed in the future, or be linked to a breastfeeding woman in the future. INTERVENTIONS - any type of intervention designed to promote the uptake of breastfeeding was included; control groups could receive an alternative breastfeeding promotion programme or standard care. OUTCOMES - the primary outcome was initiation of breastfeeding; secondary outcomes (duration and exclusivity of breastfeeding) were included if initiation was reported in the same study; intermediate and process outcomes were also included, and need not necessarily be associated with reported initiation rates. DATA EXTRACTION AND VALIDITY ASSESSMENT: Data were extracted into structured tables. All included studies were checked against a comprehensive methodological checklist. Different checklists were used for RCTs, non-RCTs and before-after studies. Data extraction and validity assessment were independently checked by a second reviewer. DATA SYNTHESIS: The studies were grouped according to intervention type, and were combined using a narrative synthesis. For individual RCTs and non-RCTs reporting initiation of breastfeeding, relative risks with associated 95% confidence intervals were estimated, with calculations performed on an intention-to-treat basis where possible. Pooling of relative risks was considered inappropriate owing to the lack of similarity across the studies. RESULTS: A total of 59 studies met the selection criteria, comprising 14 RCTs, 16 non-RCTs and 29 before-after studies. Interventions were grouped into the following categories: health education, health sector initiatives (HSI) - general, HSI - Baby Friendly Hospital Initiative (BFHI), HSI - training of health professionals, HSI - US Department of Agriculture's Special Supplemental Nutrition Program for Women, Infants, and Children (WIC), HSI - social support from health professionals, peer support, media campaigns, multifaceted interventions. In many cases, studies were dissimilar in terms of the type of intervention(s), participants and the definitions of outcomes. Methodological problems of some studies also limited interpretation of findings. (ABSTRACT TRUNCATED) PMID- 11111104 TI - World congress of gastroenterology 2005 canadian bid federation PMID- 11111105 TI - Implications of antibiotic resistance in the management of Helicobacter pylori infection. PMID- 11111106 TI - Acid-induced esophageal shortening in humans: a cause of hiatus hernia? AB - BACKGROUND: Hiatus hernia and gastroesophageal reflux disease commonly coexist, and there is pathophysiological evidence that the presence of a hiatus hernia contributes to abnormal acid reflux. However, the cause of hiatus hernia remains unclear. In an animal model, it has been shown that acute acid injury to the esophagus results in esophageal shortening, raising the possibility that reflux esophagitis per se can contribute to the formation of hiatus hernia by inducing esophageal shortening. AIM: To determine whether luminal acid produces esophageal shortening in humans. METHODS: Twelve volunteers were each studied on two occasions, one week apart, in a double-blind, crossover trial. The location of the lower esophageal sphincter (LES), as well as the LES resting pressure and axial length were determined at baseline and then again after 20 min of either acid or saline perfusion. RESULTS: Acid perfusion did not induce significant changes in resting LES pressure but resulted in proximal migration of the LES (ie, esophageal shortening) by an average of 0.5 cm, with the largest proximal migration being 1.8 cm. In contrast, saline perfusion resulted in slight distal migration of the LES (ie, esophageal lengthening). CONCLUSIONS: Intraluminal acid perfusion causes longitudinal axis shortening of the esophagus and suggests that gastroesophageal acid reflux may contribute to the cause of hiatus hernia. PMID- 11111107 TI - Outcomes following liver transplantation for patients with alcohol- versus nonalcohol-induced liver disease. AB - OBJECTIVE: To document and compare the outcomes of adult patients who received liver transplants for alcohol- and nonalcohol-induced liver diseases who attended a liver transplantation follow-up clinic in an urban, nontransplantation centre at a time when no formal alcohol abuse program for transplant candidates and/or recipients was offered. PATIENTS AND METHODS: The study population comprised 10 alcoholic patients and 48 nonalcoholic patients followed for an average of 41 months (range five to 79 months) and 46 months (range two to 116 months), respectively. Primary outcome variables included rates of recidivism, duration of abstinence after transplantation and compliance with post-transplant medical follow-up visits. Time to discharge after transplantation, episodes of graft rejection, liver and renal biochemical abnormalities, diabetes, hypertension, sepsis, strictures, complications unrelated to transplantation and changes in psychosocial status were secondary outcome variables. RESULTS: Significant differences were found with respect to a higher incidence of recidivism (50% for alcoholic patients compared with 2% for nonalcoholic patients, P<0.0001), a shorter period of abstinence after transplantation (14.7+/-17.2 months for alcoholic patients compared with 26.3+/-23.0 months for nonalcoholic patients, P<0.05) and more missed office visits (2.7+/-3.5 for alcoholic patients compared with 1.0+/-1.9 for nonalcoholic patients, P=0.05) in the alcoholic group. The alcoholic group also had a lower incidence of rejection episodes (10% for alcoholic patients compared with 44% for nonalcoholic patients, P<0.05) but higher rates of post-transplantation diabetes (40% for alcoholic patients compared with 2% for nonalcoholic patients, P<0.05), more nontransplantation related complications (20% for alcoholic patients compared with 0% for nonalcoholic patients, P<0.05), and higher serum creatinine but lower bilirubin and cyclosporine A levels (P<0.05, respectively). Marital separations were also more common in the alcoholic group (20% for alcoholic patients compared with 0% for nonalcoholic patients, P<0.05). CONCLUSIONS: In the absence of formal alcohol abuse programs, the post-transplantation outcome in alcoholic patients generally does not compare well with that of patients who undergo transplantation for nonalcohol-related liver diseases. PMID- 11111108 TI - Meeting review--Helicobacter pylori: basic mechanisms to clinical cure 2000. AB - The meeting Helicobacter pylori: Basic Mechanisms to Clinical Cure 2000, held in Bermuda from March 26 to 29, 2000, gathered physicians and scientists from all corners of the world. State-of-the-art reviews and the most recent developments in the field were presented. This article summarizes the highlights of this meeting, including important scientific and clinical developments. PMID- 11111109 TI - Implications of antibiotic resistance in the management of Helicobacter pylori infection: Canadian Helicobacter Study Group. AB - Eradication of Helicobacter pylori from the gastric and duodenal mucosa is an important clinical goal in the treatment of infected patients with peptic ulcer disease and other H pylori-associated conditions. Although several oral drug combination regimens are associated with eradication rates of approximately 85% in controlled trials, the success rate in patients infected with a resistant strain of H pylori is closer to 75%. Resistance to metronidazole and clarithromycin, which are common components of combination treatment regimens, is of greatest concern. Reported rates of H pylori resistance to various antibiotics vary considerably. In Canada, the data documenting H pylori susceptibility are limited but suggest that resistance to these antibiotics varies geographically and within specific treatment groups. Although susceptibility testing is not a prerequisite for initial treatment of individual patients infected with H pylori, formal efforts to identify and monitor both the causes and prevalence of antibiotic resistance across Canada are a much needed step in the ongoing management of this important infection. Recommended treatment regimens may be useful, even for treating apparently resistant H pylori strains. However, it is important to understand the mechanisms of the development of resistant strains to manage patients with treatment failure better. PMID- 11111110 TI - Antibiotic susceptibility and resistance testing: an overview. AB - The results of in vitro antibiotic susceptibility testing can predict the clinical response to treatment and guide the selection of antibiotics. The minimum inhibitory concentration (MIC) of an organism is the lowest concentration of an antibiotic that will inhibit its growth. Bacteria are classified as sensitive, intermediate or resistant based on breakpoint MIC values that are arbitrarily defined and reflect the achievable levels of the antibiotic, the distribution of MICs for the organism and their correlation with clinical outcome. Broth dilution, agar dilution and gradient diffusion (the 'E test'), where twofold serial dilutions of antibiotic are incorporated into tubes of broth, agar plates or on a paper strip, respectively, are different methods to measure the MIC of an organism. The disk diffusion method defines an organism as sensitive or resistant based on the extent of its growth around an antibiotic containing disk. MIC values are influenced by several laboratory factors. To ensure reproducible results, the laboratory must closely follow methods developed by the National Committee for Clinical Laboratory Standards, which defines standard growth media, incubation temperature and environment, the inoculum and quality control parameters. PMID- 11111111 TI - Epidemiology of the antibiotic resistance of Helicobacter pylori in Canada. AB - BACKGROUND: The rate of Helicobacter pylori resistance to antibiotics determines the cure rate of treatment regimens containing such antibiotics. AIMS: To review the literature to determine the rates of H pylori resistance to metronidazole and clarithromycin in Canada, and whether these rates vary in different regions of Canada. METHODS: The literature was reviewed extensively for the prevalence of antibiotic-resistant H pylori in Canada by searching MEDLINE from January 1980 to May 1999, as well as abstracts of the American Gastroenterology Association Digestive Disease Week, Canadian Digestive Disease Week and The European H pylori Study Group Meetings from January 1995 to May 1999. RESULTS: Eleven studies that estimated H pylori resistance to metronidazole resistance and nine that estimated resistance to clarithromycin in Canada were identified. Rates of resistance for metronidazole and clarithromycin varied from 11% to 48% and 0% to 12%, respectively. Studies that obtained their estimates using the E-test and those that did not clearly exclude patients who had undergone previous attempts at H pylori eradication had higher estimates of resistance, accounting for this variability in results. CONCLUSIONS: The prevalence of primary H pylori resistance in Canada appears to be 18% to 22% for metronidazole and less than 4% for clarithromycin. These rates appear to be consistent across the different regions studied in Canada, but many regions have not been studied. PMID- 11111112 TI - Effects of in vitro antibiotic resistance on treatment: bismuth-containing regimens. AB - Bismuth compounds remain useful for Helicobacter pylori eradication therapy. These include colloidal bismuth subcitrate (CBS), bismuth subsalicylate (BSS) and, most recently, ranitidine bismuth citrate (RBC). CBS appears to prevent the development of imidazole resistance when coadministered with nitroimidazoles. Traditional triple therapy with bismuth, metronidazole and tetracycline or amoxicillin (BMT/A) only partially overcomes metronidazole resistance. However, the addition of a PPI to bismuth triple therapy largely overcomes established metronidazole resistance if treatment is given for at least one week or more. When RBC rather than PPI is used with clarithromycin, this dual regimen appears to be more effective in preventing the development of secondary clarithromycin resistance. The triple combination of RBC, metronidazole and clarithromycin appears to be effective against metronidazole resistant strains of H pylori. Thus, overall, there is some evidence that bismuth compounds may prevent the development of antibiotic resistance and that existing antibiotic resistance may at least be partially overcome in vitro and in vivo. With the growing emergence of H pylori resistance to metronidazole and clarithromycin, further research to clarify the role of bismuth compounds is required. PMID- 11111113 TI - Blood transfusion requirements after liver biopsy. AB - It is common practice to type and screen the blood before performing a percutaneous liver biopsy. Many practitioners think that this is unnecessary but do not have a reason to change their practice. The requirements for transfusion after biopsy were determined in a consecutive sample of cases at a tertiary care teaching hospital with the use of health record review and the anecdotal recall of gastroenterologists and others performing biopsies. One of 266 liver biopsies required a transfusion after biopsy over a two-year period. One other case of hemorrhage with a fatal outcome was recalled by several individuals. It is concluded that the incidence of significant hemorrhage after liver biopsy is low, and that it may not be necessary to type and screen the blood of low risk patients before biopsy. PMID- 11111114 TI - Sensitivity to interferon-alpha and interferon-stimulated gene factor 3 binding activity in human chronic myelogenous leukemia cell line KT-1. AB - The mechanism of responsiveness of chronic myelogenous leukemia (CML) cells to interferon (IFN)-alpha was examined by using two subclones of CML cell line KT-1 which exhibited significantly different sensitivities to the antiproliferative and apoptosis-inducing effects of IFN-alpha. IFN-stimulated gene factor 3 (ISGF3) formation by IFN-alpha was reduced in the IFN-alpha-resistant subclone compared to the IFN-alpha-sensitive subclone. We conclude that the level of ISGF3 formation is responsible for the difference in IFN-alpha responses between these subclones. PMID- 11111115 TI - Consequences of total and subtotal myeloperoxidase deficiency: risk or benefit ? AB - A group of 100 totally or subtotally myeloperoxidase (MPO)-deficient individuals was compared to a reference population of 118 probands selected at random. Data for a protective effect of the deficiency against cardiovascular damage are presented. On the other hand, a significantly higher occurrence of severe infections and chronic inflammatory processes was noted among the deficient patients. An increased incidence of cancer among the MPO-deficient individuals was not demonstrated. PMID- 11111116 TI - The inflammation meter: novel technology to detect the presence of infection/inflammation in patients without leukocytosis but with increased leukocyte adhesiveness/aggregation. AB - OBJECTIVES: To reveal the presence of infection/inflammation in patients with relatively normal white blood cell count (WBCC) by using the leukocyte adhesiveness/aggregation test (LAAT). METHODS: The LAAT was performed by using a simple slide test and image analysis (Inflamet), the WBCC, by an electronic cell analyzer, C-reactive protein, by Laser nephelometry and CD11b/CD18 by whole blood flow cytometry. RESULTS: Forty out of a cohort of 121 patients with nonviral acute febrile illness had a WBCC within normal limits. The intensity of the inflammatory response in these individuals as judged by either C-reactive protein, or fibrinogen concentrations, erythrocyte sedimentation or polymorphonuclear leukocyte CD11b/CD18 expression was similar to that observed in patients with a leukocytic response. Our present finding that 63% out of the group with documented infection/inflammation and no leukocytosis had a significantly increased LAAT suggest that the lack of leukocytosis is in part a pseudoleukopenia, or is associated with some degree of uncompensated tissue leukostasis. CONCLUSIONS: The lack of a leukocytic response in a patient with nonviral infection/inflammation is by no means a sign of a less inflammatory response. The increased state of leukocyte adhesiveness/aggregation might help to disclose the presence of inflammation in these individuals. PMID- 11111117 TI - Oral contraceptives can cause falsely low vitamin B(12) levels. AB - Serum vitamin B(12) radioimmunoassays may give falsely low results in patients with folate deficiency, multiple myeloma, megadose of vitamin C and following radioisotope organ scan. We evaluated 10 consecutive healthy women on oral contraceptives (OC) who had falsely low vitamin B(12) levels, as reflected by normal urine methylmalonic acid and plasma homocysteine. After 1-month cessation of OCs, vitamin B(12) returned to the normal range in all women. Transcobalamin I (TCI) blood level was decreased in 60% of patients. OCs may cause temporary low vitamin B(12) blood levels of no clinical significance that can be associated with low TCI levels PMID- 11111118 TI - Comparative genomic hybridization study of de novo myeloid neoplasia. AB - Comparative genomic hybridization (CGH) was used to detect chromosomal imbalances in 20 patients with a diagnosis of myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). The results obtained were compared with G-banding analysis. This last methodology showed 50% of cases with clonal abnormalities whereas CGH detected 70% of cases with copy number changes. Gains were more frequent than losses and constituted 66% of total changes detected. The most common gains included chromosomes 21 and chromosome region 18p for AML and chromosome 17 and region 1p33p35 for MDS. Losses represent 34% of changes and the regions involved were 5q31q32, 7q22, 7p12 and 13q21q22. CGH revealed additional chromosome imbalances in 12 of 20 cases (60%) which were not detected by traditional cytogenetic studies, demonstrating complex karyotype in 50% (6/12). Combination of CGH and G-banding provides an efficient method to identify critical regions present in the malignant clone, which is of great value in the prognosis and outcome of myeloid neoplasias. PMID- 11111119 TI - Premarital screening of beta-thalassemia trait in the province of Denizli, Turkey. AB - A premarital screening program aiming at reducing the incidence of thalassemia major was started under the auspices of the Regional Health Administration in 1995 in the city of Denizli in the Aegean region of Turkey. In this report we assessed the 4-year results of the screening program. All couples who applied for marriage procedures were screened for beta-thalassemia trait by automatic red cell indices and Hb A(2) determination. The couples at risk were counseled and offered prenatal diagnosis and termination of pregnancy in case of an affected fetus. From October 1995 to August 1999, a total of 19,804 subjects (9,902 couples) were recruited for this study. The prevalence of beta-thalassemia trait with increased Hb A(2) was found to be 2.6% (514/19,804). In addition to the thalassemia trait, 22 patients (0.11%) had sickle trait. In 15 of the 9,902 couples, both partners were found to be carriers of the beta-thalassemia trait. After genetic counseling, 2 of the 15 planned carrier marriages were canceled. Seven couples declared that they do not want to have a child at present. Prenatal diagnosis was sought by 6 couples. One fetus was found to be normal, 4 had thalassemia minor and 1 had thalassemia major; this pregnancy was terminated by elective abortion. This study indicated that premarital screening is a very useful tool for detecting carrier couples and an effective way of controlling thalassemia major. PMID- 11111120 TI - Granulocytic sarcoma with breast and skin presentation: a report of a case successfully treated by local radiation and systemic chemotherapy. AB - Granulocytic sarcoma (GS) is a rare extramedullary tumor composed of immature myeloid cells. It is usually associated with leukemia or other myeloproliferative disorders, but can also occur without overt hematologic disease, i.e. in patients with a normal bone marrow and no history of acute myelogenous leukemia. This primary extramedullary lesion may indeed represent a diagnostic and therapeutic dilemma for both the hematopathologist and oncologist. We describe a case of GS diagnosed in a nonleukemic patient and review the literature regarding the pathologic features and treatment of this condition. PMID- 11111121 TI - The fitness-for-work evaluation of a young patient with essential thrombocythemia. AB - We describe a case of a military cadet, a candidate for submarine service, who was diagnosed with essential thrombocythemia (ET). The estimated risk in the literature for major clinical complications in ET patients is about 5-20%. Our report concerns a young asymptomatic patient where the estimated risk is probably much lower. We think that the activity in the marine corps, which at times is involved with some degree of dehydration, might increase the risk for thromboembolic events. The possibility of a major thromboembolic complication occurring at sea could complicate and endanger the patient's life, especially when an evacuation to a hospital is required. A submarine officer has a responsibility to other people's life who depend on his skills and ability. In case of a major complication which can cause sudden incapacitation, the consequences can be catastrophic to many. Opposed to the considerations to disqualify the young cadet stands the fact that his training was expensive, and that it would be unfortunate to disqualify a very talented young person. We thus recommended to clear the cadet as a marine officer in teaching jobs, in sites were medical assistance is available in a short period of time. In our opinion he should be disqualified from serving in combat jobs where there is a greater risk for him and for the men under his command. PMID- 11111122 TI - IgD multiple myeloma preceding the development of extensive extramedullary disease without medullary involvement. AB - We present a unique case of IgD multiple myeloma (MM) preceding the development of extensive extramedullary disease without medullary involvement. A 63-year-old man was diagnosed with IgD-lambda MM when he developed anemia. After 3 months of chemotherapy, he was in complete remission as evidenced by the disappearance of bone marrow (BM) plasmacytosis, monoclonal IgD protein in his serum, and Bence Jones proteinuria. Six months after diagnosis, his disease took an unusual course with the development of plasmacytomas in the skin, without medullary involvement. He then received chemotherapy, resulting in the complete disappearance of the subcutaneous plasmacytomas. Two years after the initial diagnosis, his disease took an aggressive clinical course with retroperitoneal relapse, leading to the patient's death within 1 month. The two separate episodes of extramedullary disease were associated with elevated serum lactic dehydrogenase levels and the absence of plasma cells in the BM. This case provides evidence of two separate transformations of the original malignant MM clone. PMID- 11111124 TI - Hydroxyurea as a cause of drug fever. AB - We report on a patient with essential thrombocythemia treated with hydroxyurea who became febrile 3 weeks after the treatment was started. After drug withdrawal, the fever resolved but after rechallenge there was recurrence of the fever. Although hydroxyurea-induced fever is rare, this drug must be added to the list of drugs that produce fever and the physicians should be aware of this possibility. PMID- 11111123 TI - Sudden bilateral deafness from hyperleukocytosis in chronic myeloid leukemia. AB - Sudden-onset bilateral deafness as a clinical manifestation of hyperleukocytosis in chronic myeloid leukemia (CML) is a rare occurrence. We found only 27 clinical descriptions in 16 published papers. In this work, the authors present a review on deafness in CML and describe a new case with prominent hyperleukocytosis, where the neurological findings suggest slowing of the circulation through small blood vessels in the brainstem as the cause of deafness. The evolution was good after treatment. To our knowledge, this is the second case documented with electrical auditory brainstem-evoked potentials and the first with magnetic resonance imaging. PMID- 11111125 TI - Consider 'hyperviscosity syndrome' in unexplained breathlessness. AB - A 58-year-old man presenting with increasing shortness of breath over several months is reported here. All investigations were repeatedly normal until he suddenly and rapidly deteriorated, warranting admission, and a diagnosis was made of early multiple myeloma associated with the hyperviscosity syndrome. The plasma viscosity was extremely high compared to the low concentration of paraprotein present, and the possible mechanisms are discussed. Although uncommon, the hyperviscosity syndrome is an important cause of dyspnoea which is readily reversible, and should be considered when investigations of cardiac and pulmonary function are normal. PMID- 11111126 TI - CD3+, CD4-, CD8-, TCR alpha beta-, TCR gamma delta+ granular lymphocyte proliferative disorder without lymphocytosis and clinical symptoms. AB - Granular lymphocyte-proliferative disorder is characterized by a proliferation of large granular lymphocytes (LGLs). It is often associated with neutropenia, rheumatoid arthritis (RA), and pure red cell aplasia (PRCA). Phenotypic analysis has demonstrated that in most cases, the LGLs show a clonal rearrangement of the TCR alpha beta rearrangement. We are reporting a patient with TCR gamma delta LGL proliferation without clinical findings and lymphocytosis. The patient showed an expansion of the CD3+, CD16+, CD56+, and CD57+ LGL populations which involved coexpression of TCR gamma delta with TCR J gamma and J delta 1 gene rearrangement. Autoimmune manifestations, including RA and PRCA, have not appeared and the results of laboratory examinations have not changed for 1 year after the diagnosis. PMID- 11111127 TI - IGF-1 and its binding proteins IGFBP-1 and 3 as nutritional markers in prepubertal children. AB - OBJECTIVE: To assess the validity of the use of IGF-1, IGFBP-1 and IGFBP-3 as biochemical markers of nutritional status in prepubertal healthy children. DESIGN: Cross-sectional survey. SETTING: Healthy children from the Madrid area. PARTICIPANTS: Prepubertal children (aged 7-10 years) with a body mass index (BMI) above the 90th percentile (n = 25) and below the 10th percentile (n = 31) were selected from 2,559 included in the CAENPE study (Food Intake and Nutritional Status in Schoolchildren from Madrid). RESULTS: Overweight children were found to have higher serum levels of IGF-1 (306+/-162.2 vs. 209+/-71.2 ng/ml, p<0.001) and IGFBP-3 (3.3+/-1.0 vs. 2.9+/- 0.5 mg/l, p<0.01) and lower serum levels of IGFBP-1 (4.3+/-3.9 vs. 13.8+/-7.4 microg/l, p<0.01). There was a positive correlation with BMI, for IGF-1 and IGFBP-3, and negative correlation for IGFBP-1. No differences in albumin and transferrin concentrations were observed between both groups of children. CONCLUSION: IGF-1, IGFBP-1 and IGFBP-3 clearly classify over- and underweight prepubertal children, showing a good correlation with BMI. They can be used as biochemical markers of caloric nutritional status in this population. PMID- 11111128 TI - Electrolytes, water, RNA, total creatine and calculated resting membrane potential in muscle tissue from pregnant women. AB - BACKGROUND: The nutritional situation of the fetus and pregnant woman is important for human health, but knowledge of how nutrition affects maternal metabolism and physiology during pregnancy is limited. Such knowledge is important, for example in body composition studies, when information about lean tissue composition is needed. Muscle, a main part of lean tissue, changes its composition in response to age and sex, but the effect of pregnancy on this composition is unknown. METHODS: Muscle samples from 11 pregnant and 16 nonpregnant women were analyzed for water, electrolytes, total creatine, alkali soluble protein (ASP), DNA and RNA. Plasma was analyzed for electrolytes. The amount of extracellular and intracellular water as well as the resting membrane potential (RMP) in muscle were calculated. RESULTS: Pregnant women had lower plasma concentrations of potassium and sodium but higher muscle concentrations of sodium and water (total and extracellular) than nonpregnant women. RMP was more negative in pregnant than in nonpregnant women. Total creatine in muscle (per kilogram ASP) was increased during pregnancy. The muscle content of RNA (per kilogram DNA) was lower in gestational week 18 than in nonpregnant controls. CONCLUSION: Pregnancy influences muscle composition in several ways that are relevant for an increased understanding of interactions between nutrition and reproduction. PMID- 11111129 TI - Vitamin A deficiency in mice enhances the colonic level of purine enzyme activity. AB - BACKGROUND: Vitamin A is an important nutritional factor that regulates normal growth and functions of epithelial cells of the gastrointestinal tract. OBJECTIVE: The objective of this study was to examine the role of vitamin A on the histological and biochemical changes in the colon of mice. METHODS: To address this issue, vitamin A deficiency (VAD) was developed in mice by placing them on a VAD diet from weaning up to 120-170 days. Infiltration of inflammatory cells in the colon was determined histologically. Activities of adenosine deaminase, adenylate deaminase, purine nucleoside phosphorylase and myeloperoxidase were determined. RESULTS: VAD in mice induced a significant increase in the number of mast cells per 100 crypts. There was also an abundance of other connective tissue cells such as plasma cells, lymphocytes and neutrophils around the crypts in the lamina propria. The colonic activity of adenosine deaminase and adenylate deaminase was increased due to VAD, whereas purine nucleoside phosphorylase activity remained unchanged. Immunohistochemical analysis showed an increased expression of adenosine deaminase in VAD mice colon. The increase in myeloperoxidase activity was not statistically significant. CONCLUSIONS: VAD causes upregulation of purine enzyme, which together with an increased number of inflammatory cells might exacerbate colonic injuries in VAD condition. PMID- 11111130 TI - Bioavailability of n-3 polyunsaturated fatty acids (PUFA) in foods enriched with microencapsulated fish oil. AB - OBJECTIVE: Incorporation of fish oil into food products provides a means of increasing n-3 fatty acid intake, particularly in populations where fish consumption remains low. The aim of the present study was to evaluate the bioavailability of n-3 PUFA in microencapsulated fish-oil-enriched foods compared with an equal amount of n-3 PUFAs contained in fish oil capsules. METHODS: Twenty five healthy female volunteers were randomly assigned to one of two groups for the 4-week intervention: one group received 0.9 g of n-3 PUFA/day as fish oil capsule (capsule group), while the second group (food group) received an equal amount of n-3 PUFA/day from enriched foods. Baseline and post-intervention samples were analysed for platelet fatty acid composition. RESULTS: There was no significant difference in the change in platelet arachidonic acid (AA), eicosapentaenoic acid (EPA), or docosahexaenoic acid (DHA) between the two groups following the intervention. CONCLUSIONS: The results indicate that n-3 PUFA from microencapsulated fish-oil-enriched foods are as bioavailable as n-3 PUFA in a capsule. Fortification of foods with microencapsulated fish oil, therefore, offers an effective way of increasing n-3 PUFA intakes and status in line with current dietary recommendations. PMID- 11111131 TI - Bioavailability of folate from processed spinach in humans. Effect of food matrix and interaction with carotenoids. AB - The effect of the food matrix and dietary fibre on the bioavailability of folate is not known. In a controlled, 3-week dietary intervention study, 28 men and 42 women were divided into six groups to receive either a control diet (n = 10), or the control diet plus 20 g/MJ per day (n = 12 per group) of whole-leaf spinach, minced spinach, liquefied spinach, or liquefied spinach to which dietary fibre (10 g/kg wet weight) was added. The sixth group received the control diet plus a synthetic carotenoid supplement with similar amounts of beta-carotene and lutein as found in spinach. A significantly higher plasma folate response was found for the pooled spinach groups than for the control group. Among the spinach groups no significant differences were detected. However, the plasma folate response of the pooled minced and liquefied spinach groups was greater than that of the whole leaf spinach group (p = 0.03). Re-addition of dietary fibre to the liquefied spinach to compensate for the fibre broken down during liquefaction did not reduce the plasma folate response. The consumption of the carotenoid supplement did not have an effect on plasma folate concentrations compared with the control group. The food matrix in which the folate is entrapped plays a role in folate bioavailability. PMID- 11111132 TI - Lower absorption of cholesteryl oleate in rats supplemented with Areca catechu L. extract. AB - Areca catechu L. extracts I and II, prepared using two different solvent systems, exhibited strong inhibitory activities against pancreatic cholesterol esterase (pCEase) in vitro. To determine their cholesterol-lowering effects, these two extracts were investigated by analyzing plasma lipid levels, intestinal enzyme activities, and the absorption of cholesteryl oleate. For 6 days, male rats were fed a diet containing cholesteryl oleate (0.5 g/100 g of body weight) either with or without the Areca nut extract supplements. The supplementation of the two Areca nut extracts significantly lowered the concentrations of plasma cholesterol by 13. 4 and 11.7% and plasma triglycerides by 35.0 and 36.9%, respectively, compared with the pre-experimental values. However, when the cholesteryl oleate diet was fed without any Areca nut extract in high-cholesterol control, the plasma cholesterol and triglyceride concentrations significantly increased by 13.6 and 15.9%, respectively, compared with the pre-experimental values. After 6 days of treatment, the intestinal pCEase activities were significantly lower in the groups supplemented with the Areca nut extracts (37.8 and 26.5%) than in the group with no extract supplement (83.2%). The supplements also significantly elevated the excretion of [1,2(n)-(3)H]cholesteryl oleate administered orally, when determined by the large intestinal contents, 930.5 Bq/day (Areca I) and 1,766.3 Bq/day (Areca II) vs. 98.1 Bq/day (high-cholesteryl oleate (CO) control). The inhibition of pCEase activity with the supplementation of the Areca nut extracts could account for the decrease in [1,2(n)-(3)H]cholesteryl oleate absorption that resulted in decreased radioactivity in blood. PMID- 11111133 TI - Dietary habits and cardiovascular risk in the Spanish population: the DRECE study (II) micronutrient intake. Dieta y Riesgo de Enfermedades Cardiovasculares en Espana. AB - OBJECTIVE: To evaluate the dietary micronutrient intake in the adult Spanish population participating in the DRECE study. METHODS: The cross-sectional study was performed in two stages in 1991 and 1996 in 43 primary care clinics. One thousand two hundred people 'with cardiovascular risk' and 600 'without risk' answered a food frequency questionnaire. RESULTS: Significant increases in vitamin C, retinol, lycopenes, beta-cryptoxanthin and vitamin E intakes were found. Vitamin A, alpha-carotenoid and lutein intakes decreased. Vitamin B(12), B(6) and folic acid intakes increased in people with cardiovascular risk, whereas only the last two increased in the control group. Nearly 100% of the people consumed the recommended dietary allowances for vitamins B(12) and B(6) and >70% for folic acid. Calcium, iron, and zinc intake increased in both groups, but magnesium and selenium intake increased only in people at risk. Vitamin A, B(1) and zinc intakes have decreased, and >50% of the people do not consume the recommended dietary allowance. CONCLUSION: Antioxidant vitamins and vitamin B(12), B(6) and folic acid intakes seem to be adequate in the adult Spanish population, no significant differences appear regarding their cardiovascular risk status. Vitamin A, B(1) and zinc intakes are not appropriate. PMID- 11111134 TI - The leg depressor and levator muscles in the squat lobster Munida quadrispina (Galatheidae) and the crayfish Procambarus clarkii (Astacidae) have multiple heads with potentially different functions. AB - The proximal leg muscles of decapod crustaceans, controlling movements at the first two joints, are anatomically more complex than the better-studied distal leg muscles. Despite extensive research on their involvement in diverse behaviors, no complete descriptions of the anatomy and innervation of these muscles for any species have been published. We describe the anatomy and innervation of the depressor muscle in the second leg of the squat lobster Munida quadrispina and compare its anatomy with that of its homologue in the crayfish Procambarus clarkii and its antagonist, the levator, in both species. Of the six anatomically distinct heads comprising M. quadrispina's depressor muscle, one arises in the coxa (coxal head) and five are bi-articular (cross two joints), arising from widely dispersed sites on the thoracic endophragmal skeleton (dorsal, sternal, caudal, ventral-rostral, ventral-caudal heads). The heads' widely divergent force vectors are accommodated by the depressor apodeme's bifurcation at a thin flexible point. In total, eighteen neurons with central somata were backfilled from nerve branches to the heads. The common inhibitor and at least one neuron of unknown function with rostro-lateral soma and extremely sparse neurites innervate all heads. The sixteen excitatory motoneurons' somata are clustered in two locations, five rostral and eleven caudal to the neuropil. Rostral motoneurons innervate the two ventral heads (rostral and caudal). Their integrating segments lie rostral to those of the caudal group motoneurons and are straight or 'Y'-shaped, the latter longer and larger in diameter. Both morphological types have one prominent medial neurite that crosses the midline and could allow direct interaction between bilateral pairs of rostral motoneurons. The caudal motoneurons provide partially shared innervation to the remaining four heads. Six provide exclusive innervation, one to the caudal head, two to the sternal head, and three to the bi-articular dorsal head and uni articular coxal head which share innervation. Of the remaining five caudal motoneurons, two are shared by the caudal head and the dorsal-coxal pair of heads and three are shared by the caudal and sternal heads. P. clarkii has two depressor muscles; the rostral depressor has a single head morphologically similar to the ventral rostral head in M. quadrispina; the caudal depressor muscle has four heads (dorsal, coxal, caudal, sternal) that insert on the large caudal depressor apodeme. The overall organization of depressor muscle heads in P. clarkii and M. quadrispina is similar, taking into consideration the different internal and external thoracic anatomies and the different resting stances, horizontal and tilted, respectively. As in other species, M. quadrispina and P. clarkii have two levator muscles, each with an apodeme of complex structure attributable to the levators' role in leg autotomy. The caudal levator arises in the coxa; it has two heads in M. quadrispina, but only one in P. clarkii. In both species the rostral levator has three heads all arising within the thorax. Divergent force vectors and partially independent innervation of different heads composing complex musculature at single joints constitute an anatomical level of organization that neural mechanisms must accommodate to produce adaptive movements. PMID- 11111135 TI - Colocalization of neuronal NO synthase with urotensins I and II in the caudal neurosecretory neurons and the urophysis of the teleost oreochromis niloticus: a gold immunoelectron microscopic study. AB - The intracellular distribution of neuronal nitric oxide synthase (nNOS) was studied in the caudal neurosecretory system of a teleost, Oreochromis niloticus (Cichlids), by means of post-embedding immunogold labeling with a polyclonal antibody directed against nNOS of human origin. Ultrastructural examination demonstrated that neuronal NOS-like molecules are distributed within the Dahlgren cell perikarya, the neurosecretory axons, and the urophysial axon terminals. In the neurosecretory somata, gold particles for nNOS were mainly cytosolic, whereas in the neurosecretory axons and axon terminals they were associated with the membrane and/or the dense core of neurosecretory granules. Double immunogold labelings for nNOS/urotensin I (UI) and nNOS/urotensin II (UII) demonstrated that nNOS-like molecules are colocalized with UI and/or UII in the neurosecretory granules contained within the urophysial terminals. The present findings suggest that both a soluble cytosolic and a particulate neuronal NOS are expressed in the caudal neurosecretory neurons. They confirm previous biochemical data on the same species. PMID- 11111136 TI - Exploitation of an ancient escape circuit by an avian predator: prey sensitivity to model predator display in the field. AB - Certain insectivorous birds, such as the painted redstart (Myioborus pictus), undertake flush pursuit--a characteristic display that elicits an escape reaction by an insect, which the bird then chases in the air and eats. This account describes experiments showing that flush pursuit uses visual displays, which are likely to exploit an ancient neural circuit in dipteran insects, the visual systems of which are well documented as detecting looming stimuli and triggering an escape responses. Using models that decompose components of the redstart display, specific elements of the display were analyzed for their contribution in triggering visually induced escape behavior by dipterous insects. Elements tested were pivoting body movements, patterning on the spread tail and wings, and visual contrast of model redstarts against pale and dark backgrounds. We show that contrasting patterns within the plumage are crucial to foraging success, as is contrast of the bird against a background. Visual motion also significantly contributes to the successful flushing. In contrast, unpatterned models and patterned models that do not contrast with the background are less successful in eliciting escape responses of flies. Natural visual stimuli provided by Myioborus pictus are similar to those known to trigger looming and time-to-collision neurons in the escape circuits of flies and other insects, such as orthopterans. We propose that the tuning properties of these neural pathways might have contributed to the evolution of foraging displays in flush-pursuing birds. PMID- 11111137 TI - Eye design and color signaling in a stomatopod crustacean Gonodactylus smithii. AB - Many species of stomatopod crustaceans have multiple spectral classes of photoreceptors in their retinas. Behavioral evidence also indicates that stomatopods are capable of discriminating objects by their spectral differences alone. Most animals use only two to four different types of photoreceptors in their color vision systems, typically with broad sensitivity functions, but the stomatopods apparently include eight or more narrowband photoreceptor classes for color recognition. It is also known that stomatopods use several colored body regions in social interactions. To examine why stomatopods may be so 'concerned' with color, we measured the absorption spectra of visual pigments and intrarhabdomal filters, and the reflectance spectra from different parts of the bodies of several individuals of the gonodactyloid stomatopod species, Gonodactylus smithii. We then applied a model of multiple dichromatic channels for color encoding to examine whether the finely tuned color vision was specifically co-evolved with their complex color signals. Although the eye design of stomatopods seems suitable for detecting color signals of their own, the detection of color signals from other animals, such as reef fishes, can be enhanced as well. Color vision in G. smithii is therefore not exclusively adapted to detect its own color signals, but the spectral tuning of some photoreceptors (e.g. midband Rows 2 and 3) enhances the contrast of certain color signals to a large enough degree to make co-evolution between color vision and these rather specific color signals likely. PMID- 11111138 TI - Cardiac involvement in primary myopathies. AB - Simultaneous or temporarily staggered affection of both the skeletal as well as the cardiac muscle (cardiac involvement, CI) is a frequent finding in primary myopathies (MPs). CI leads to impulse generation defects, impulse conduction defects, thickened myocardium, left ventriculalr hypertrabeculation, dilatation of the cardiac cavities, secondary valve insufficiency, reduction of coronary vasodilative reserve, intracardial thrombus formation, and heart failure with systolic and diastolic dysfunction. CI has been found in Duchenne muscular dystrophy (MD), Becker MD, Emery-Dreifuss MD, facioscapulohumeral MD, sarcoglycanopathies, myotubular congenital MD, myotonic dystrophies type 1 and 2, proximal myotonic myopathy, myoadenylate deaminase deficiency, glycogenosis type II, III, IV, VII and IX, carnitine deficiency, mitochondriopathy, desmin MP, nemaline MP, central core disease, multicore MP, congenital fiber-type disproportion MP, Barth syndrome, McLeod syndrome and Bethlem MP. Patients with any of the above-mentioned myopathies should be cardiologically investigated as soon as their diagnosis is established, since sufficient cardiac therapy improves CI in MPs and since management of these patients is influenced by the degree of CI. PMID- 11111139 TI - Effects of spinal cord stimulation and coronary artery bypass grafting on myocardial ischemia and heart rate variability: further results from the ESBY study. AB - In the present study, 24-hour ECG recordings were analyzed from the Electrical Stimulation versus Coronary Artery Bypass Surgery (ESBY) Study where spinal cord stimulation was compared to CABG (coronary artery bypass grafting) in selected patients with severe angina pectoris. During the monitoring period, the spinal cord stimulation was discontinued to evaluate possible long-term effects of this treatment. The number of ischemic episodes and the duration of ischemia decreased in the CABG group at the follow-up when compared to spinal cord stimulation (p<0.05). In spite of this, the number of anginal attacks decreased (p<0.0001) in both groups. The fact that the anginal symptoms decreased in the spinal cord stimulation group in spite of discontinued stimulation and lack of effects on ischemic ST changes could indicate a long-term primary analgesic effect of this treatment in addition to the well-documented acute anti-ischemic effect. PMID- 11111140 TI - Activated polymorphonuclear leukocytes and monocytes in the peripheral blood of patients with ischemic heart and brain conditions correspond to the presence of multiple risk factors for atherothrombosis. AB - OBJECTIVE: Risk factors like hypertension, diabetes mellitus, dyslipidemia and smoking contribute to the pathogenesis of atherothrombosis. We investigated whether the multiplicity of risk factors for atherothrombosis is associated with leukocyte activation. METHODS: We examined the availability of CD11b/CD18 antigen on the surface of peripheral blood polymorphonuclear leukocytes and monocytes in patients with acute ischemic heart and brain conditions. RESULTS: There was a highly significant (p<0.00001) increment in the availability of the CD11b/CD18 antigen on the surface of the polymorphonuclear leukocytes in patients with multiple (2 or more) vascular risk factors [mean fluorescence intensity (MFI) +/- SD, 210+/-102] as opposed to individuals with none or 1 risk factor for atherothrombosis (MFI 159+/-73). Similar results were observed on the monocytes: 309+/-151 and 235+/-97, respectively (p<0. 00001). CONCLUSION: The multiplicity of risk factors for atherothrombosis is associated with the up-regulation of CD11b/CD18 antigen on the surface of peripheral blood polymorphonuclear leukocytes and monocytes, suggesting the presence of an increased inflammatory response and leukocyte activation in these individuals. PMID- 11111141 TI - The impact of risk factors for atherosclerosis on the vasomotor effects of inhibition of nitric oxide synthesis in patients with normal angiograms. AB - We assessed the impact of systematic risk factors on the vasomotor effects of inhibition of nitric oxide synthesis. N(G)-monomethyl-L-arginine (LNMMA) was infused intracoronarily at 4, 8 and 16 micromol/min followed by intracoronary bolus administration of 250 microg nitroglycerin. Computerized angiography was used to assess the changes in the diameter of coronary segments. During the LNMMA infusions there was no significant difference in LNMMA response between smokers and non-smokers (-5.5+/-0.8 and -6.6+/-0.6%, respectively) or between hypertensives and normotensives (-6.4+/-1.1 and -6.1+/-0.6%, respectively), but the response was less in hypercholesterolaemic patients (-4.5+/-0.7 vs. -8.0+/ 0.6%, p<0.05). Thus, the reduced nitric oxide activity is related to hypercholesterolaemia but not to smoking and hypertension. PMID- 11111142 TI - Treatment benefit in the enhanced external counterpulsation consortium. AB - The present study utilized a cohort of 2,289 consecutive patients enrolled in the Enhanced External Counterpulsation (EECP) Consortium to evaluate whether results of university studies showing EECP safety and effectiveness in treating angina can be generalized. EECP was found to be safe and well tolerated with a 4.0% rate of adverse experiences. Angina class improved in 74% of patients with limiting angina (Canadian Cardiovascular Society, CCS, functional class II-IV), with patients most impaired at baseline demonstrating the greatest improvement (39.5% of patients in CCS III and IV improved 2 or more classes). Efficacy was independent of provider setting or experience, women responded as well as men, and although younger patients demonstrated a greater likelihood of improvement, EECP was effective in patients ranging from 19 to 97 years. Extending the benefit of EECP treatment to a wider range of patients may be indicated based on these findings. PMID- 11111143 TI - Effects of a slow-release nifedipine on 24-hour ambulatory blood pressure and ischemic changes on 24-hour ambulatory electrocardiogram in patients with severe coronary artery disease. AB - Calcium antagonists have long been used as first-line drugs for hypertension and angina. However, deleterious effects have also been reported in patients treated with calcium antagonists. Thus, we evaluated the effect of a slow-release twice daily formulation of nifedipine in 10 patients with severe coronary artery disease. Twenty-four-hour ambulatory electrocardiography (AECG) and blood pressure monitoring (ABPM) were performed simultaneously to detect any association between ischemic episodes on the ECG and changes in blood pressure (BP) and heart rate with and without nifedipine. Increased oxygen demand due to an increased systolic BP and heart rate was associated with ischemic episodes without nifedipine, while those with nifedipine were accompanied by a fall in diastolic BP and a rapid increase in heart rate. This slow-release twice-daily formulation of nifedipine may induce myocardial ischemia through a heart-rate increase and a decrease in coronary blood flow due to lower diastolic BP in patients with severe coronary artery disease. A once-daily formulation of nifedipine might be of great value for such patients. PMID- 11111144 TI - Tachycardias in infants, children and adolescents: safety and effectiveness of radiofrequency catheter ablation. AB - Radiofrequency catheter ablations provide an effective control of a variety of supraventricular and ventricular tachycardias in adults. This study was designed to evaluate the efficacy and safety of radiofrequency catheter ablations in infants, children and adolescents. Ninty-three ablations were performed in 84 patients ranging from 5 months to 18 years of age. All but 1 patient were successfully treated (98.8%). Two patients required 1 and 2 additional attempts to achieve success. Tachyarrhythmic episodes recurred in 4 patients within 1-5 months after ablation, which were successfully treated by repeating the intervention. Significant complications occurred in 3 of the ablations (3.2%). Although radiofrequency ablations are very effective and safe in pediatric patients, indications should be restricted in patients younger than 4 years because of a higher risk of possibly life-threatening complications. PMID- 11111145 TI - Effects of intracoronary tissue-type plasminogen activator treatment in kawasaki disease and acute myocardial infarction. AB - We retrospectively studied 3 patients with Kawasaki disease (KD) and acute myocardial infarction (AMI) who were treated with intracoronary administration of tissue-type plasminogen activator (t-PA). Two-dimensional echocardiogram on the next day of the treatment revealed reduction of thrombus and improvement of the cardiac function in all 3 patients. However, a 12-month-old patient treated with 200,000 U/kg of t-PA at 48 h after the onset of AMI died of recurrent myocardial infarction. The other 2 patients treated with 400,000 and 800,000 U/kg, respectively, showed clear, though not prompt, improvement in clinical symptoms and laboratory data. The intracoronary thrombolytic therapy using high-dose t-PA appears effective in treating AMI associated with KD. PMID- 11111146 TI - Early reperfusion assessment and repeated thrombolysis in acute myocardial infarction estimated by repeated standard electrocardiography. A randomised, double-blind, placebo-controlled pilot study. AB - Thrombolytic therapy with streptokinase (SK) in acute myocardial infarction (AMI) does not result in early reperfusion in approximately 25% of patients. We hypothesized that early repeated thrombolysis with rt-PA in patients with early failed reperfusion would result in myocardial reperfusion. Fifty-nine AMI patients with a symptom delay of <6 h, treated with SK were included. ECG was taken on admission and after 90 and 180 min. An ST recovery of > or =25% at 90 min was interpreted as successful reperfusion. Sixteen patients had failed reperfusion at 90 min and were randomized to repeated thrombolysis with rt-PA or placebo. At 180 min from SK start, ST recovery was higher in the placebo group than in the rt-PA group (71 vs. 40%, p = 0.05). No serious bleeding complications were observed. Due to the limited sample size it was not possible to draw prominent conclusions. PMID- 11111147 TI - Formulas for dynamic exercise heart rate and blood pressure. AB - Formulas are proposed for the ranges of normal heart rate and blood pressure during dynamic exercise. They apply to normotensive men and women approximately 20-50 years old. The numerical values of the parameters may have to be modified for other populations, partial populations or other age groups. PMID- 11111148 TI - Fatal dilated cardiomyopathy associated with a mitochondrial DNA deletion. AB - A 27-year-old man was admitted to hospital because of severe cardiac failure. Investigation revealed dilated cardiomyopathy with a left ventricular ejection fraction of 15-20%. During adolescence the patient had been investigated for growth retardation and he also had progressive external ophthalmoplegia. There had been no symptoms of cardiac disease until 2 weeks before admittance. An endomyocardial biopsy showed cardiomyocytes deficient in cytochrome c oxidase (COX) in a mosaic pattern. A skeletal muscle biopsy showed mitochondrial myopathy with COX-deficient ragged-red fibers. Molecular genetic analysis revealed a heteroplasmic, 3.8-kb, mitochondrial DNA (mtDNA) deletion in heart and muscle. PCR-based quantification of the proportion of mtDNA with deletion showed 47% mutated mtDNA in the myocardial biopsy and 68% in muscle. In spite of treatment, the condition deteriorated and the patient died 5 days after admittance. This case demonstrates that mtDNA deletions may occasionally be the cause of severe dilated cardiomyopathy, and that morphological and molecular genetic diagnosis may be obtained by endomyocardial biopsy. PMID- 11111149 TI - Isolated left ventricular abnormal trabeculation is a cardiac manifestation of neuromuscular disorders. AB - OBJECTIVES: Isolated left ventricular abnormal trabeculation (ILVAT) is defined as >3 coarse trabeculations of the left ventricular wall, apically to the papillary muscles, in hearts without congenital malformations. The aims of the study were to assess by echocardiography the prevalence of ILVAT, look for cardiac findings in ILVAT and find out if ILVAT is associated with neuromuscular disorders. DESIGN AND RESULTS: In the course of 3 years, ILVAT was found in 15/17, 648 patients (0.08%). Twelve patients had heart failure, all had ECG abnormalities. All patients had neuromuscular disorders. A definite diagnosis could be established in 10 patients (metabolic myopathy, n = 9; Becker's muscular dystrophy, n = 1). The remaining 5 patients refused further neurological investigations. CONCLUSIONS: ILVAT is a rare disorder associated with ECG abnormalities, heart failure and neuromuscular disorders, which is a reason why both cardiologists and neurologists should be involved. PMID- 11111150 TI - Role of mitochondria in neuronal apoptosis. AB - Apoptosis is a controlled form of cell death that participates in the demise of neuronal cells during development, neurodegenerative disorders and exposure to neurotoxic agents. In recent years, the mitochondria have emerged as being pivotal in controlling apoptosis. They house a number of apoptogenic molecules that are released into the cytoplasm at the onset of apoptosis. These include cytochrome c, apoptosis-inducing factor and various caspases. Mitochondria also play an important role in intracellular Ca(2+) regulation, which is crucial to excitotoxic neurodegeneration. Alterations in energy (ATP) production by mitochondria (due to hypoxia or mutations in genes encoding mitochondrial proteins of the electron transport chain) can induce apoptosis in neurons or increase their sensitivity to apoptosis. PMID- 11111151 TI - Nitric-oxide-induced inhibition of mitochondrial complexes following aglycaemic hypoxia in neonatal cortical rat brain slices. AB - The effect of aglycaemic hypoxia (AH) on the activity of the mitochondrial respiratory chain complexes was measured in superfused neonatal cortical brain slices. After 30 min AH, there were no significant changes in the activities of complex I, II-III and IV or citrate synthase compared to controls. Following 30 min AH and a 30-min reperfusion period (with oxygen and glucose), the activities of complex II-III and complex IV were significantly reduced (by 25 and 17%, respectively). These reductions in enzyme activity were not abrogated by removing external calcium prior to and throughout AH, but could be reversed by the presence of the nitric oxide (NO) synthase inhibitor N omega-nitro-L-arginine during these periods. These data suggest that NO or an NO-derived species is involved in the decreases in mitochondrial enzyme activities observed after AH PMID- 11111153 TI - Biochemical characterization of the mitochondrial permeability transition in isolated forebrain mitochondria. AB - Induction of the mitochondrial permeability transition (PT) has been proposed to contribute to neuronal cell death. Nearly all studies of the biochemistry of PT induction, however, have been conducted in isolated liver mitochondria. To better understand PT induction in brain mitochondria, we used Ficoll gradients to purify nonsynaptosomal mitochondria from the forebrains of male Fischer 344 rats. Incubation of these mitochondria with Ca(2+) was associated with a loss of absorbance. Inorganic phosphate enhanced this loss of absorbance, and the PT inhibitor cyclosporin A reduced it, especially in conjunction with ADP. These findings suggest that Ca(2+)-mediated loss of absorbance resulted from PT induction. Na(+), which enhances mitochondrial Ca(2+) efflux, but stimulates mitochondrial free radical production, had no effect on PT induction. These data confirm the existence of tissue-specific differences in the nature of PT induction. PMID- 11111152 TI - Brain mitochondrial responses to postischemic reperfusion: a role for calcium and hydrogen peroxide? AB - During early recirculation following global brain ischemia, mitochondria are exposed to markedly elevated Ca(2+) concentrations and a short-lived production of reactive oxygen species, including hydrogen peroxide (H(2)O(2)). A brief increase in mitochondrial Ca(2+) and a subsequent increase in mitochondrial glutathione content have been observed. In the present study, we have confirmed the increase in mitochondrial glutathione in a rat model of global forebrain ischemia. This change was not inhibited by treatment of the rats with FK506, contrasting with our previous finding that cyclosporin A partially blocked the increase. These results suggest that induction of the mitochondrial permeability transition may be necessary for the increase in glutathione content in these organelles. To further investigate possible mitochondrial responses during early postischemic reperfusion, mitochondria isolated from normal brain were exposed to Ca(2+) and H(2)O(2), under conditions similar to those in intact cells. Respiratory activity was substantially modified when the mitochondria were exposed to Ca(2+) and H(2)O(2) together. Two distinct and largely noninteracting mechanisms apparently accounted for the responses to these agents. The effects of Ca(2+), but not H(2)O(2), were inhibited by cyclosporin A, again implicating the permeability transition in some of the mitochondrial changes. PMID- 11111154 TI - Microglial cells in culture express a prominent glutathione system for the defense against reactive oxygen species. AB - To obtain information on the glutathione metabolism of microglial cells, the content of glutathione and activities of enzymes involved in the defense against peroxides were determined for microglia-rich cultures from rat brain. These cultures contain approximately 90% microglia cells as determined by immunocytochemical staining for glial markers, by the phagocytosis activity of the cells and by the production of superoxide after stimulation of the cells with phorbolester. For these cultures, a glutathione content of 41.2 +/- 11.2 nmol/mg protein and a specific activity of glutathione reductase of 15.2 +/- 3.2 nmol/(min x mg protein) were determined. These values are significantly higher than those found for astroglial or neuronal cultures. In addition, with 68.7 +/- 23.5 nmol/(min x mg protein), the specific activity of glutathione peroxidase in microglial cultures was 78% higher than in cultured neurons. The specific catalase activity of microglial cultures was less than 40% that of astroglial or neuronal cultures. Microglial cultures contain only marginal amounts of oxidized glutathione. However, on application of oxidative stress by incubation of microglial cultures with hydrogen peroxide or with the superoxide-producing hypoxanthine/xanthine oxidase system, cellular glutathione was rapidly oxidized. These results demonstrate that microglial cells have a prominent glutathione system, which is likely to reflect the necessity for self-protection against reactive oxygen species when produced by these or surrounding brain cells. PMID- 11111155 TI - Transport of L-citrulline in neural cell cultures. AB - Uptake of L-[(14)C]citrulline was studied in cell culture models of the main neural cell populations, in astroglia-rich primary cultures derived from neonatal rat brain, in rat glioma cells C6-BU-1, in cells of the murine microglial clone N11 and in the glioma x neuroblastoma hybrid cell line 108CC15 with neuronal properties. For comparison, cells of the peripheral macrophage cell line RAW 264.7 were also investigated. A saturable component of uptake was found in all cases with K(M) values between 0.4 and 3.4 mM and V(max) values between 15 and 35 nmol.min(-1).(mg protein)(-1). A nonsaturable component dominated uptake at high concentrations of extracellular citrulline. Rates of uptake of L-citrulline were not affected when Na(+) or Cl(-) were omitted from the incubation medium or in the presence of depolarizing concentrations of K(+). Saturable uptake of citrulline was strongly inhibited by an excess of histidine or beta-2 aminobicyclo-(2.2.1)-heptane-2-carboxylic acid; excess amounts of arginine, creatine, glutamate, cysteic acid or N-methyl-alpha-aminoisobutyric acid did not reduce citrulline uptake. Preincubation of the cells with bacterial lipopolysaccharide and interferon gamma did not stimulate transport of citrulline. The results suggest that at physiological concentrations citrulline is taken up by neural cells with the help of transport system L for large neutral amino acids. Therefore, in the brain, effective utilization of extracellular citrulline as part of an intercellular trafficking of intermediates of an NO/citrulline cycle depends on the concentrations of all neutral amino acids present. PMID- 11111156 TI - Simple models of threshold curves in the expression of inborn errors of metabolism: application to some experimental observations. AB - The expression of an enzymatic deficiency in a metabolic network can present a biochemical threshold. This threshold can be characterised thus: (1) a low activity of the enzyme can sustain a normal flux, but (2) a minute further decrease of its activity makes the flux collapse. We give simple mathematical models displaying such a behaviour, and we apply the models to some examples of oxidative phosphorylation dependency on respiratory chain complex deficiency. PMID- 11111157 TI - Lysine-rich histone (H1) is a lysyl substrate of tissue transglutaminase: possible involvement of transglutaminase in the formation of nuclear aggregates in (CAG)(n)/Q(n) expansion diseases. AB - Histone H1, which contains about 27% lysine, is an excellent lysyl donor substrate of Ca(2+)-activated guinea pig liver tissue transglutaminase as judged by rapid fluorescence enhancement in the presence of the glutaminyl-donor substrate 1-N-(carbobenzoxy-L-glutaminylglycyl)-5-N-(5'N'N'-dimethylamino naphth alenesulfonyl) diamidopentane. Sodium dodecyl sulfate gel electrophoresis of a 30 min reaction mixture revealed the presence of fluorescent high-M(r) aggregates, which are also formed when histone H1 is incubated solely with activated tissue transglutaminase. Aggregate formation is even more pronounced when histone H1 is incubated with activated tissue transglutaminase and dimethylcasein (glutaminyl donor only). The findings suggest not only that histone H1 is an especially good lysyl substrate of tissue transglutaminase, but that it is also a glutaminyl substrate. Histone H1 is a good lysyl substrate of transglutaminase purified from Streptoverticillium mobaraense, suggesting that the ability of histone H1 to act as a transglutaminase lysyl substrate is widespread. In agreement with previous studies, it was found that human beta-endorphin is a moderately good substrate of tissue transglutaminase. At least 8 neurodegenerative diseases, including Huntington's disease, are caused by (CAG)(n) expansions in the genome and by an expansion of the corresponding polyglutamine domain within the expressed, mutated protein. Polyglutamine domains are excellent substrates of liver and brain transglutaminases. A hallmark of many of the (CAG)(n)/polyglutamine expansion diseases is the presence of polyglutamine-containing aggregates within the cytosol and nuclei of affected neurons. Transglutaminase activity occurs in both of these compartments in human brain. In future studies, it will be important to determine whether transglutaminases play a role in (1) cross-linking of histone H1 to glutaminyl donors (including polyglutamine domains) in nuclear chromatin, (2) the formation of nuclear aggregates in (CAG)(n)/polyglutamine expansion diseases, (3) DNA laddering and cell death in neurodegenerative diseases and (4) depletion of neuropeptides in vulnerable regions of Huntington's disease brain. PMID- 11111158 TI - Non-invasive methods for studying brain energy metabolism: what they show and what it means. AB - This review summarises the ways in which magnetic resonance spectroscopy (MRS) and related methods can be used as windows on brain energy metabolism in vivo. (31)P-MRS can measure acute changes in non-oxidative ATP synthesis in transient states, and at steady state reflects the balance of ATP demand and mitochondrial function. (13)C-MRS labelling methods can measure a variety of carbon fluxes. The few (31)P- and (13)C-MRS studies of the response to functional activation suggest quite large increases in oxidative metabolism. Functional magnetic resonance imaging measures the hyperoxygenation that results from increase in cerebral blood flow in excess of glucose oxidation, attenuated somewhat by a smaller increase in oxygen consumption. Previous positron emission tomography studies disagree on the size of activation response. These are powerful but demanding techniques, valuable in understanding both normal physiology and pathophysiology. However, discrepancies remain to be reconciled, and this will require increasing sophistication of both techniques and analytical models. PMID- 11111159 TI - (13)C MR spectroscopy study of lactate as substrate for rat brain. AB - In order to address the question whether lactate in blood can serve as a precursor for cerebral metabolites, fully awake rats were injected intravenously with [U-(13)C]lactate or [U-(13)C]glucose followed 15 min later by decapitation. Incorporation of label from [U-(13)C]glucose was seen mainly in glutamate, GABA, glutamine, aspartate, alanine and lactate. More label was found in glutamate than glutamine, underscoring the predominantly neuronal metabolism of pyruvate from [U (13)C]glucose. It was estimated that the neuronal metabolism of acetyl CoA from glucose accounts for at least 66% and the glial for no more than 34% of the total glucose consumption. When [U-(13)C]lactate was the precursor, label incorporation was similar to that observed from [U-(13)C]glucose, but much reduced. Plasma analysis revealed the presence of approximately equal amounts of [1,2,3-(13)C]- and [1,2-(13)C]glucose, showing gluconeogenesis from [U-(13)C]lactate. It was thus possible that the labeling seen in the cerebral amino acids originated from labeled glucose, not [U-(13)C]lactate. However, the presence of significantly more label in [U-(13)C]- than in [2,3-(13)C]alanine demonstrated that [U (13)C]lactate did indeed cross the blood-brain barrier, and was metabolized further in the brain. Furthermore, contributions from pyruvate carboxylase (glial enzyme) were detectable in glutamine, glutamate and GABA, and were comparatively more pronounced in the glucose group. This indicated that relatively more pyruvate from lactate than glucose was metabolized in neurons. Surprisingly, the same amount of lactate was synthesized via the tricarboxylic acid cycle in both groups, indicating transfer of neurotransmitters from the neuronal to the astrocytic compartment, as previous studies have shown that this lactate is synthesized primarily in astrocytes. Taking into consideration that astrocytes take up glutamate more avidly than GABA, it is conceivable that neuronal lactate metabolism was more prominent in glutamatergic neurons. PMID- 11111160 TI - Brain creatine kinase and creatine transporter proteins in normal and creatine treated rabbit pups. AB - Systemic creatine (Cr) supplementation increases brain phosphocreatine (PCr) and prevents hypoxic seizures in 15-day-old rabbits. Between 5 and 30 days of age during normal development, rabbit gray matter mitochondrial creatine kinase (Mi CK) increases 400% while cytosolic CK (BB-CK) increases 60%. In white matter, both isoenzymes show smaller, similar increases (40%) during this period. The Cr transporter protein decreases 60% between 5 and 15 days in both regions. In vivo CK rate constants measured by (31)P nuclear magnetic resonance increase 30% between 10 and 20 days, and then fall 50% between 20 and 30 days in predominantly gray matter slices. Similar maturational changes are seen in predominantly white matter slices. Injecting Cr at 15 days does not significantly change BB-CK or Mi CK isoenzymes or the in vivo CK reaction rate constants. Thus, the largest change in the CK system associated with suppression of hypoxic seizures in Cr-treated rabbits is increased PCr in gray and white matter. PMID- 11111161 TI - Effect of acute insulin-induced hypoglycemia on fetal versus adult brain fuel utilization, assessed by (13)C MRS isotopomer analysis of [U-(13)C]glucose metabolites. AB - Tight glycemic control during diabetic pregnancy has been shown to significantly reduce the occurrence of congenital malformations and other effects of maternal diabetes on the offspring. However, intensive insulin therapy often causes recurring acute maternal hypoglycemia, which has been found to be harmful to the developing fetus, although the mechanisms involved are not clear. The aim of our work was to study the effect of acute insulin-induced maternal hypoglycemia on glucose metabolism in the fetal brain. To this end, near-term pregnant New Zealand rabbits were rendered hypoglycemic, and [U-(13)C]glucose was infused into maternal circulation. The metabolic fate of the (13)C-labeled glucose was then studied in fetal brain extracts by (13)C NMR isotopomer analysis, together with conventional biochemical assays of glucose and lactate levels in both plasma and brain. For comparison [U-(13)C]glucose was also administered to insulin-induced hypoglycemic young adult rabbits. Our results showed that while plasma glucose levels were significantly reduced (approximately 70%) relative to controls, no changes in cerebral glucose levels could be detected. Lactate levels were found to be significantly decreased in hypoglycemic fetal plasma and brain. No differences in lactate levels between control and hypoglycemic young rabbit plasma and brain were observed. These differences were attributed to the utilization of lactate as an energy substrate in the fetal brain, but not in the adult brain. Higher relative (13)C enrichments of most glucose metabolites, except lactate, in the hypoglycemic fetal and young rabbit brains, observed by (13)C NMR, stem from reduced endogenous plasma glucose pools, thereby diluting the labeled glucose to a lower extent. The relative glucose (or glucose-derived lactate) flux via the pyruvate carboxylase and pyruvate dehydrogenase pathways (PC/PDH ratio) was not altered under hypoglycemic conditions in the fetal brain for both glutamine and glutamate, but significantly increased in the adult brain for both glutamine and glutamate. The presented data indicate the ability of the fetal brain to maintain energy metabolism during acute hypoglycemia, via lactate utilization. The increase in the adult PC/PDH ratio was suggested by us to stem from increased PC activity, in order to replenish TCA cycle intermediates. PMID- 11111163 TI - Effects of ammonia exposition on glioma cells: changes in cell volume and organic osmolytes studied by diffusion-weighted and high-resolution NMR spectroscopy. AB - NH(4)Cl (10 mM) caused a sustained increase in the cell volume in immobilized, perfused F98 glioma cells to approx. 125% of control after 3 h, as measured by diffusion-weighted (1)H NMR spectroscopy. Concomitantly, the glutamine (Gln) concentration increased by 130%, accompanied by a marked decrease in cytosolic osmolytes, i.e. myo-inositol and taurine, determined from (1)H NMR spectra of PCA extracts. Inhibition of Gln synthetase partially prevented the increase in water content. While losses of organic osmolytes are also observed under hypotonic conditions, the rapid cell swelling is followed by the regulatory cell volume decrease (RVD), and is accompanied by decreased cytosolic Gln. We suggest that the rise in intracellular osmolarity, which is attributed to NH(4)Cl metabolism to Gln, but also to alanine (Ala), is not compensated by the release of other osmolytes, and causes cell swelling without RVD. PMID- 11111162 TI - Metabolism of 3-(13)C-malate in primary cultures of mouse astrocytes. AB - Malate, specifically labeled with carbon 13 on C(3), was synthesized by chemical means and used to study malate metabolism by primary cultures of mouse cortical astrocytes. 3-(13)C-Malate in combination with glucose as well as 3-(13)C-malate alone were used as substrates; the effect of 3-nitropropionic acid, an inhibitor of succinate dehydrogenase and fumarase was also examined. The consumption of malate was only 0.26 micromol/mg of protein, approx. 25-fold lower than the consumption of glucose. Besides lactate, glutamine and fumarate were the two major metabolites released to the medium. Very low and similar levels of isotopic enrichment were detected on C(2) and C(3) of lactate; glutamine was labeled on C(2) and C(3) to a similar extent as well and labeling on C(4) was only detected when glucose was not added. These labeling studies suggest that cytosolic malic enzyme is not active in primary astrocytes and support the occurrence of pyruvate recycling in astrocytes. PMID- 11111164 TI - Infant nutrition--brain development--disease in later life. An introduction. PMID- 11111165 TI - The role of dietary n-6 and n-3 fatty acids in the developing brain. AB - The dietary requirements for essential fatty acids and the possibility of a specific role for the polyunsaturated fatty acid docosahexaenoic acid (DHA) is one of the most controversial areas in infant nutrition. DHA is found in unusually high concentrations in the brain and is selectively accumulated during fetal and infant brain growth. DHA can be synthesised through a complex series of chain elongation-desaturation reactions from alpha-linolenic acid, but the efficiency of this process in young infants is not clear. Clinical studies on the potential benefits to neural development of dietary DHA have yielded conflicting results. Recent studies have provided evidence that plasma DHA is available to developing brain and that DHA is involved in dopamine and serotonin metabolism. These findings should guide clinical studies to more sensitive measures of the functional roles of dietary n-3 fatty acids and to clinical conditions where n-3 fatty acids may have benefit. PMID- 11111166 TI - Diet, lipids and brain development. AB - Brain development is a sequential anatomical process characterised by specific well-defined stages of growth and maturation. One of the fundamental and necessary events in the normal development of the central nervous system in vertebrates is the formation of a myelin sheath. It is becoming more evident that this process is influenced by dietary lipids. A number of findings have indicated that the administration of a diet deficient in essential fatty acids during development causes hypomyelination in the rat brain. Our studies have shown that lipids can also play a role in accelerating myelinogenesis in the brain of rats whose mothers had been fed, during pregnancy and lactation, a lipid fraction extracted from yeast grown on n-alkanes. Further studies have shown that accelerated myelinogenesis is connected to a precocious appearance of behavioural reflexes. Thus, the use of particular lipids in human nutrition must be carefully screened for possible effects on brain development. PMID- 11111167 TI - Maternal high n-6 polyunsaturated fatty acid intake during pregnancy increases voluntary alcohol intake and hypothalamic estrogen receptor alpha and beta levels among female offspring. AB - Identification of nongenetic biological factors that predispose to alcohol abuse is central to attempts to prevent alcoholism. Since an exposure to estradiol in utero increases voluntary alcohol intake in adulthood, we investigated whether an increase in pregnancy estradiol levels, caused by feeding pregnant mice a high fat corn oil diet, also influences voluntary alcohol intake among female offspring. In addition, the effect on estrogen receptor alpha (ER-alpha) and ER beta protein levels in the brain using Western blot assay, was determined. Pregnant CD-1 mice were kept on a high n-6 polyunsaturated fatty acid (PUFA; 43% calories from corn oil) or low n-6 PUFA diet (16% calories from corn oil) throughout gestation, and switched to a Purina laboratory chow after the pups were born. When 4 months of age, the female offspring were given a choice between 5% alcohol and tap water. The offspring of high n-6 PUFA mothers voluntarily consumed more alcohol than the offspring of low n-6 PUFA mothers. ER-alpha and ER beta protein levels in the hypothalamus were 1.5- and 2-fold higher, respectively, in the female offspring of high n-6 PUFA mothers than in the low n 6 PUFA offspring. No significant changes in the protein levels of ER-alpha and ER beta were seen in the frontal brain. Our findings indicate that a maternal exposure to a high n-6 PUFA diet during pregnancy increases alcohol intake among female offspring. This behavioral change, together with previously observed increase in aggressiveness and reduction in depressive-like behavior in these offspring, may be linked to an increase in the hypothalamic ER-alpha and ER-beta levels. PMID- 11111168 TI - Effect of docosahexaenoic acid content of maternal diet on auditory brainstem conduction times in rat pups. AB - Previous studies of dietary docosahexaenoic acid (DHA; 22:6n-3) effects on neurodevelopment have focused mainly on effects on the visual system; these studies may be confounded by effects on the retina rather than on neural pathways. Auditory brainstem conduction times (ABCTs) provide an alternate measure of central neural development. We conducted a dose-response study in which ABCTs were measured in pups whose dams were fed diets containing one of three levels of DHA (2, 4 or 6% of total fatty acids) from a single cell oil. Diets were fed during pregnancy and lactation, and pups were randomly cross fostered on postnatal day 3 to minimize litter effects. ABCTs showed a dose response effect, with higher levels of dietary DHA being associated with longer conduction times on postnatal day 31 (p < 0.05). Higher dietary DHA was reflected in pup cerebrums collected on postnatal days 3 and 31, and levels of arachidonic acid (AA, 20:4n-6) were inversely related to levels of DHA. This study demonstrated that the auditory brainstem response is sensitive for identifying effects of diet on neurodevelopment, and that supplementing the maternal diet with high levels of DHA may negatively impact development of the central auditory system of offspring. PMID- 11111169 TI - Dual effect of laparoscopy on cell-mediated immunity. AB - Laparoscopic influence on cell-mediated immunity and tumour evolution is controversial. The objective of the present study was to assess tumour growth and immune patterns after laparoscopy on an experimental study. Lewis rats, bearing an intrapancreatic ductal carcinoma randomly underwent one of the following 2 hour procedures: anaesthesia, laparotomy or CO(2) pneumoperitoneum. Cell-mediated immunity was investigated through determination of serum IL1beta concentrations by ELISA and TNFalpha, IL6 and iNOS gene transcriptions in blood white cells and peritoneal cells by RT-PCR 1 day after operation. Tumour growth and spread patterns were assessed on anatomopathological examination 2 weeks after surgery. Tumour growth and spread were unaffected no matter what procedure was applied, but port-site seeding occurred in half of the cases undergoing laparoscopy. No significant change in acute-phase protein response, represented by IL1beta serum concentration, was found after laparoscopy. TNFalpha, IL6 and inducible NO synthase gene transcriptions were enhanced in blood white cells and depressed in peritoneal immune cells after laparoscopy. In our experimental conditions, cell mediated immune response to CO(2) pneumoperitoneum seems to be a good systemic immune activation and a less acute peritoneal immune response as opposed to control laparoscopy. This early impairment of peritoneal macrophage immune activity, observed after a long-lasting CO(2) pneumoperitoneum, might be responsible for the high rate of port site recurrence. PMID- 11111170 TI - A novel auxiliary partial orthotopic liver transplantation model in rats. AB - BACKGROUND: Although auxiliary partial orthotopic liver transplantation (APOLT) has become a well-accepted procedure recently, a practical experiment model in APOLT using small animals has yet to be developed. METHODS: Male Lewis rats were used for both donors and recipients. An auxiliary partial graft was obtained by ex vivo resection of the donor right and caudate lobes, and was transplanted orthotopically into the recipient after resection of the recipient medial and left hepatic lobes. Portal vein and hepatic duct reconstructions were by the cuff technique, and supra- and intrahepatic vena cava were sutured continuously. Operative outcomes, serum chemistry, liver tissue blood flow, angiographic and histopathological findings were then examined. Conventional orthotopic liver transplantation (OLT) procedures were also undertaken as a control. RESULTS: One day, 1-week and 1-month survival rate of APOLT group was 100, 85 and 85%, respectively. AST in the APOLT group on the 1st postoperative day was significantly higher than in the OLT group. No significant differences were recognized in serum albumin and total bilirubin levels between the two groups. Although the portogram of an APOLT rat showed slight narrowing at the cuff anastomosis site, both the graft and the native liver were opacified similarly. The liver tissue blood flow on the 5th postoperative day in the native liver and the graft returned to as high as 95 and 74% of the values on laparotomy, respectively. Histological examinations of the auxiliary graft 1 month after transplantation showed mild ductular proliferation and mononuclear cell infiltration around the portal triads. CONCLUSION: This novel APOLT model in rats allows practical and reproducible results, and may be of value in the basic study of APOLT procedures. PMID- 11111171 TI - Prolongation of cardiac allograft survival by selective injection of donor liver leukocytes in non-immunosuppressed rats. AB - Liver grafts are spontaneously accepted in several animal combinations and are able to induce acceptance of another organ originating from the same donor, which would be rejected when transplanted alone. However, the exact mechanism of this unique tolerance induction capability remains unclear. The aim of our study was to investigate the ability of nonparenchymal liver cells to induce tolerance when they were separated from their parenchymal environment. In the murine combination we used (BN --> LEW), heart transplants were constantly tolerated after combined liver plus heart grafting, but rejected when transplanted alone. Nonparenchymal liver cells were isolated from BN rat livers by enzymatic digestion and injected, at different times, to LEW rats, which were recipients of BN heart transplants. The average number of mononuclear cells obtained after isolation was 20 x 10(6)/5 g of rat liver. Immediate trypan-blue exclusion test showed more than 95% of viable cells. Phenotypic studies showed a predominant (47%) lymphocyte population, 7% were monocytes and 46% were cellular debris. Among the lymphocyte population, the majority of cells were bearing the NKR-P1 receptor and about 30% CD3 receptors. Inoculation of nonparenchymal liver cells 7 and 30 days prior to heart transplantation significantly prolonged graft survival compared to controls (14.6 and 12.7 vs. 8.1 days; p = 0.0008 and 0.0059, respectively), whereas simultaneous injection (day 0) had no effect. Injection of donor splenocytes or nonparenchymal liver cells from a third party, at any time, had no effect on rejection. These results provide some more evidence about the specific role of liver lymphocytes in allogenic unresponsiveness. They also suggest that the hepatic parenchymal environment is necessary for the optimal development of this phenomenon. PMID- 11111173 TI - Intra- and post-operative assessment of renal cortical perfusion by laser Doppler flowmetry in renal transplantation in the rat. AB - Renal blood flow (RBF) in the period immediately following transplantation has an important prognostic value. Here we report for the first time on the use of laser Doppler flowmetry (LDF) for the measurement of renal cortical perfusion (RCP) during all the important steps in renal transplantation. Left orthotopic kidney transplantation was performed in Lewis rats (n = 14) after 2 h of cold ischaemia and preservation in EuroCollins solution. Under baseline conditions, RCP in the donor and recipient kidneys were similar with a coefficient of variability of 11 and 12%, respectively. There was a progressive increase in RCP during the first 60 min after transplantation with a return to normal values 2 weeks later. In conclusion, LDF provides a rapid and continuous measure of RCP without interference to the operative site and may prove a useful tool for the measurement of RBF during kidney transplantation. PMID- 11111172 TI - Effect of a novel immunosuppressant, FTY720, on allograft survival after renal transplant in rats. AB - FTY720 was developed by chemical modification of ISP-1 which was purified from culture filtrates of an ascomycete, Isaria sinclairii. We evaluated the effect of FTY720 on allograft survival using a rat renal transplantation model in which Wistar King Aptekman Hokkaido rats (WKAH, RT1(K)) served as the organ donor and Lewis rats (LEW, RT1(l)) as the recipient. WKAH renal allografts were acutely rejected by the untreated LEW recipients at a mean graft survival +/- SD of 7.2 +/- 0.4 days (n = 5). Consecutive oral administration of FTY720 following transplantation significantly prolonged allograft survival in a dose-dependent manner over the range of 0. 05-3 mg/kg/day. The mean allograft survival of the recipients treated with FTY720 at a doses of 0.05, 0.1, 0.5, 1, and 3 mg/kg/day was 12.2 +/- 3.3 (n = 5, p < 0.05, vs. untreated host), 11.2 +/- 2.4 (n = 5, p < 0.05, vs. untreated host), 13.6 +/- 0.9 (n = 5, p < 0.01, vs. untreated host), 14.6 +/- 1.7 (n = 5, p < 0.01, vs. untreated host) and 20.2 +/- 0.8 days (n = 5, p < 0.01, vs. untreated host). In the recipients treated with FTY720 (3 mg/kg/day), the number of peripheral blood lymphocytes significantly decreased. From the results of the flow cytometric study, FTY720 significantly diminished the percentage of interleukin-2 receptor (IL-2R)-positive cells in the allografts (6.34 +/- 0.81% in the untreated recipients vs. 3.10 +/- 0.86% in the recipients treated with FTY720, p < 0.05). As to the CD4/CD8 ratio of splenic cells and graft infiltrate, there was no significant difference between the untreated hosts and the recipients treated with FTY720. In conclusion, FTY720 significantly extended rat renal allograft survival and the immunosuppressive effects of FTY720 may be due to a reduction in not only the number of peripheral lymphocytes but also the percentage of IL-2R-positive cells in the allografts. PMID- 11111174 TI - Effects of dibutyryl cyclic adenosine monophosphate on the ultrastructure of endothelial cells in rat lungs cold preserved for 15 hours. AB - BACKGROUND: Dibutyryl cyclic adenosine monophosphate (db-cAMP) has been shown to protect vascular endothelial cells by increasing the level of intracellular cAMP, and we have previously reported its effectiveness in lung preservation. Here, the effects of db-cAMP in lung preservation were ultrastructurally investigated, and the ultrastructural changes before reperfusion were correlated with pulmonary function after reperfusion. METHODS: The lungs of 17 Lewis rats were flushed with perfusate and prostaglandin E(1), and were then divided into three groups. In the fresh group (n = 6), the lungs were flushed with extracellular-type trehalose containing (ET-K) solution and were reperfused immediately. In the control group (n = 6) and db-cAMP group (n = 5), the lungs were flushed with ET-K solution and ET-K solution plus db-cAMP (2 mM), respectively, and were reperfused after cold preservation at 4 degrees C for 15 h. Before reperfusion, tissue was sampled and ultrastructurally analyzed by transmission electron microscopy. RESULTS: In the endothelial cells of pulmonary arterioles, the incidence of protrusion was significantly lower in the fresh and db-cAMP groups than in the control group (p < 0.05). The incidence of detachment and microvillus formation were significantly lower in the fresh and db-cAMP groups than in the control group (p < 0.01). The ultrastructure of the alveoli did not allow separation of the control and db-cAMP groups. The shunt fraction and wet to dry weight ratio of the lung tissue after reperfusion were significantly lower in the fresh and db-cAMP groups than in the control group (p < 0.01). Positive correlations were found between the incidence of these ultrastructural changes in the endothelial cells of the pulmonary arterioles and pulmonary function after reperfusion. CONCLUSION: These findings suggest that db-cAMP might attenuate the lung injury caused by cold preservation and ischemia-reperfusion, partly by suppressing the acceleration of the structural changes in the endothelial cells in the pulmonary arterioles. PMID- 11111175 TI - Preconditioning with short cycles improves ischemic tolerance in rat fast- and slow-twitch skeletal muscle. AB - AIM: The aim of this study was to investigate whether the efficacy of ischemic preconditioning (IP) in rat skeletal muscle depends on the duration of the preconditioning cycles. METHODS: Rats were divided into four groups (n = 10 each). The right hindlimb of rats in group A were subjected to 2.5 h of tourniquet ischemia followed by 2 h of reperfusion (I-R). Thereafter, muscular function was analyzed in vitro and high-energy phosphates (HEP) were determined by HPLC. Before I-R, right hindlimbs of rats in groups B-D subjected to IP with three cycles each consisting of 2.5, 5 or 10 min of ischemia followed by reperfusion for the same duration. RESULTS: Postischemic function of the extensor muscle was significantly improved with all three preconditioning protocols. Postischemic function of the soleus muscle was only improved by IP with three cycles of 5 min of ischemia and 5 min of reperfusion. Postischemic HEP tissue levels were not influenced by IP. CONCLUSION: This study shows for the first time that IP increases ischemic tolerance not only of fast-twitch but also of slow twitch skeletal muscle. The efficacy of IP seems to be less dependent on the duration of the single preconditioning cycle than on the number of cycles performed. Three cycles each of 2.5, 5 or 10 min ischemia and reperfusion significantly improved postischemic skeletal muscle function. Tissue levels of HEPs, however, were not influenced by IP indicating that preservation of HEPs does not play a major role in the effects of IP on rodent skeletal muscle. PMID- 11111176 TI - Sulindac inhibits growth of rat colon carcinoma by inducing apoptosis. AB - Sulindac possesses an inhibitory effect on colorectal cancer development. Rat colon cancer cells, ACL-15, inoculated subcutaneously in F344 rats were used. Sulindac was administered at 8 mg/kg twice daily for 7 consecutive days. Sulindac group and control group were compared regarding tumor volume and body weight. At sacrifice, the tumors were collected and examined for tumor-infiltrating lymphocytes, apoptotic index, and microvessel density. The tumor volume in the sulindac group was significantly smaller than that in the control group. Body weight, microvessel density, and tumor-infiltrating lymphocyte score were not significantly different between the two groups. The apoptotic index was significantly higher in the sulindac group than in the control group. Sulindac inhibited tumor growth by inducing apoptosis. These findings may be helpful in designing new treatment strategies in colorectal cancer patients. PMID- 11111177 TI - Effect of sigmoid colon distension on the rectosigmoid junction. Description of the rectosigmoid junction tightening reflex and its clinical implications. AB - PURPOSE: The sigmoid colon (SC) is the site of stool storage. The stools accumulate in the SC until, at a certain volume, the mechanoreceptors in the SC wall are stimulated, evoking the sigmoidorectal junction inhibitory reflex with a resulting SC contraction, rectosigmoid junction (RSJ) relaxation and passage of the stools to the rectum. However, the RSJ status during stool accumulation in the SC has been scarcely addressed in the literature. The current study investigated this point. METHODS: A balloon-ended tube was introduced into the SC of 21 healthy volunteers [mean age (+/- SD) 36.8 +/- 10.3 years; 15 men and 6 women]. The pressures in the SC and RSJ were measured by means of a perfused tube, at rest and during balloon inflation with carbon dioxide at two rates: slow (3 ml/min) and rapid (150 ml/min). The tests were repeated after individual anesthetization of the SC and RSJ. RESULTS: During slow SC distension up to 80 ml included, the RSJ pressure progressively increased while the SC exhibited no pressure response (p > 0.05). At a distending volume of 100 ml, the pressure in the SC rose (p < 0. 01) and declined in the RSJ (p < 0.05), and the balloon was dispelled to the rectum. Rapid SC distension up to 40 ml included, effected no SC pressure response (p > 0.05) while the RSJ showed progressive pressure elevation. At 60 ml distension, the SC recorded a pressure rise (p < 0.001) and the RSJ a pressure decrease (p < 0. 05); the balloon was dispelled to the rectum. The pressure in the RSJ did not respond to distension of the anesthetized SC. CONCLUSION: The study has shown that, during accumulation of stools in the SC, leakage to the rectum seems to be prevented by a reflex action which we call 'rectosigmoid junction tightening reflex'. This reflex probably acts to control both storage and emptying of the SC contents. Reflex dysfunction might lead to defecation disorders. We suggest that the RSJ tightening reflex be included as an investigative tool in the diagnosis of defecation disorders. PMID- 11111178 TI - A new model for the induction of tumours in the forestomach of rats by N-methyl-N nitrosourea. AB - BACKGROUND AND AIM: Experimental carcinogenesis models provide a useful tool in the study of the aetiopathogenesis and treatment of gastric cancer. We developed a model based on the administration of N-methyl-N-nitrosourea (NMU) in Wistar rats for the induction of maximal yield of gastric carcinomas with a short latency period, and being exclusively localized at the gastric level. METHODS: A gastric antiperistaltic fistula was performed in 90 Wistar rats classified into eight different groups. Fifteen days after surgery 5, 10, 15 or 20 mg of NMU/100 g were administered through the fistula once a week for a 3- to 5-week period. Before the administration of NMU, a pyloric blockade was made in order to obtain a temporary isolation of the stomach. At 20 weeks, animals were sacrificed and organs were removed for histological study. RESULTS: All rats treated with 15 mg NMU/100 g once a week for 5 weeks, after pyloric blockade maintained for 1 h, developed well-differentiated carcinomas in the forestomach. Carcinomas were multiple in 11% of cases and appeared with papillomatous lesions in 33% of rats. No tumours were observed in any other organs. In the other groups, no gastric carcinomas were diagnosed. CONCLUSION: The high incidence of carcinomas in the forestomach, the absence of tumours in other organs and the short latency period represent valuable criteria for the use of our model in chemotherapeutic investigations, as well as in the study of cancer evolution without interferences caused by tumour development in other organs. PMID- 11111179 TI - Are the elemental diets beneficial for colonic anastomoses? PMID- 11111180 TI - Andropause: hormone replacement therapy in the ageing male. PMID- 11111181 TI - Extracorporal shock wave therapy in the treatment of Peyronie's disease. First results of a case-controlled approach. AB - OBJECTIVE: To test whether extracorporal shock wave therapy (ESWT) has an effect in the treatment of Peyronie's disease. METHODS: 22 patients with Peyronie's disease and previous unsuccessful oral drug therapy were treated with ESWT in a prospective design with a follow-up of at least 3 months; 23 age-matched patients without previous therapy received oral placebo drug for 6 months daily as control. The standard follow-up included palpation, ultrasound, autophotography and evaluation of symptomatology based on a symptom score. The shock waves were applied under ultrasound guidance using the 'Storz Minilith SL1' lithotripter. RESULTS: The results show a significant decrease in penile curvature in the patients treated with ESWT. Concerning the decrease in pain, subjective improvement and improvement in the quality of sexual intercourse, there was no significant difference to the case-control group. The inhomogeneity of the 2 groups may influence these results due to the questionable varying natural history. CONCLUSIONS: A prospective, controlled multicenter study with standardized parameters (concerning technique and patients) is urgently required to test the effect of ESWT. PMID- 11111182 TI - Clinical experience with intraurethral alprostadil (MUSE) in the treatment of men with erectile dysfunction. A retrospective study. Medicated urethral system for erection. AB - OBJECTIVE: The Food and Drug Administration (USA) approved the transurethral administration of prostaglandin (alprostadil in January 1997), which had an efficacy of approximately 50% in clinical trials. We studied its effectiveness in clinical practice. METHODS: Patient and partner education was followed by an initial office trial of a medicated urethral system for erection (MUSE) after other medical risk factors were corrected during a 2- to 4-month period. The initial titration dose of alprostadil was usually 125 or 250 microg. Further titration, if needed, was instituted by the patient at home. Success was determined as the satisfactory completion of sexual intercourse in more than 66% of attempts, with a minimum of two being required. RESULTS: Two hundred and seventy patients entered the trials, and follow-up information was available in 229 (85%). The overall success rate was 56%. The dose required was 500 microg in 49.2% and 1,000 microg in 42.2%. Of the 44% in whom treatment failed, 61.4% did so because of lack of efficacy and 38.6% because of side effects (genital pain or urethral bleeding). Minor urogenital symptoms, which did not interfere with treatment, occurred in an additional 40% of patients. CONCLUSIONS: The efficacy of transurethral administration of alprostadil (56%) is higher than the initial published clinical trial data and higher than recent reported clinical experiences, although higher doses were required in our study. Men over 50 years of age, having an organic cause for erectile dysfunction, had better responses. Patient and partner education is important for successful treatment, and the in office initial titration is an integral part of this success. Prior correction of medical risk factors may enhance the success rate. PMID- 11111183 TI - Endoscopic bladder neck suspension revisited: long-term results of Stamey and Gittes procedures. AB - OBJECTIVE: To evaluate the long-term results of Stamey's and Gittes' procedures for genuine stress incontinence. METHODS: 72 needle procedures (34 Stamey; 38 Gittes) performed by a single surgeon between 1988 and 1994 were retrospectively reviewed. All patients had genuine stress incontinence on preoperative video urodynamics. Review was at 3 months and thereafter clinically determined. Update information was gained by a patient satisfaction questionnaire. RESULTS: Data were available for 9 years for the Stamey group (mean 8.4 years) and 6 years for the Gittes' (mean 5.3 years). At 3 months, 93% were dry. There was a gradual attrition with 38% of the Stamey and 14% of the Gittes patients remaining dry or improved at 5 years. At 9 years, only 28% of the Stamey patients maintained their improvement. 26% of the original cohort underwent a second procedure. All patients who had repeat needle operations have failed. 48 questionnaires (67%) were returned. Only 25% of patients expressed satisfaction with their operation. CONCLUSION: Early success rates with endoscopic bladder neck suspension are replaced by long-term failures. The durability is poor with an ongoing recurrent incontinence rate. Repeat procedures are not worthwhile. Gittes' procedure appears to have an earlier failure rate compared to Stamey's operation. PMID- 11111184 TI - Reproducibility of cystometry in overactive detrusor. AB - OBJECTIVE: To examine the reproducibility of cystometry in the overactive detrusor. METHODS: The study sample involved 30 patients of the placebo arm in double-blind clinical trials for an overactive detrusor. They had demonstrated detrusor overactivity and underwent the second cystometry after 2-4 weeks. Nonparametric tests for paired data were used to examine the reproducibility of four variables: volume at first desire to void, volume at first involuntary contraction, cystometric capacity, and the maximum pressure of involuntary contraction. Percent change and within-subject standard deviation were calculated to assess intraindividual variability. RESULTS: The second test results showed a significant and systematic change for the better. Volume variables increased by 10-13% (p<0.01), involuntary contraction was not elicited in 3 cases (10%), and the maximum contraction pressure decreased by 18% in the remaining cases. Intraindividual variability was not small. Seventeen patients (57%) demonstrated > or = 25% change in one or more variables, and the 95% confidence interval of cystometric capacity, for example, was calculated as (x -57, x +57), where x is a test result. No specific patients' demographics were found related to variability. CONCLUSION: Repeat cystometry in the overactive detrusor is not highly reproducible and may be subject to a systematic effect for the 'better'. Whether this is due to the placebo effect or the learning effect could not be determined. PMID- 11111185 TI - Comparison of the Bard Trak test with voided urine cytology in the diagnosis and follow-up of bladder tumors. AB - OBJECTIVES: To compare the results of the BTA Trak test with voided urine cytology (VUC) in the diagnosis and follow-up of bladder tumors. PATIENTS AND METHODS: Urine samples were obtained from 53 patients with bladder tumor (77 samples) and 53 patients treated for bladder tumor with no evidence of disease on the basis of cystoscopic evaluation (88 samples). Urine samples were collected prior to cystoscopy. The BTA assay was performed by the BTA Trak test according to the manufacturer's recommendations. A value >14 U/ml was considered abnormal. RESULTS: There was a statistically significant increase in median BTA value with increasing stage of tumor: 11.9, 57.9 and 391.0 U/ml respectively for stages pTa, pT1 and pT2/3 (p<0.0001, Kruskal-Wallis test). There was also a correlation between increasing grade and median BTA values measured at 6.9, 13.1 and 235.0 U/ml in grades 1, 2 and 3 tumors respectively (p<0.0001, Kruskall-Wallis test). The overall sensitivity of the BTA Trak test was 58.4% compared to 46.7% for VUC, a difference of 11.7%, which was statistically significant (McNemar test, p<0.005). The sensitivity of both tests combined was 63.6%. The specificity of the VUC (94.3%) was significantly higher than that of the BTA Traktrade mark (75.0%) (p<0.005, McNemar test). The accuracy of the Bard Trak test (67.3%) was similar to that of VUC (66.9%). CONCLUSION: The BTA Trak test is more sensitive than urinary cytology in the detection of bladder tumors but the improvement involved is insufficient to consider decreasing the frequency of endoscopic examinations in the follow-up of superficial bladder tumor. PMID- 11111186 TI - p53 immunohistochemistry as a prognostic marker in bladder cancer. Playground for urology scientists? AB - OBJECTIVES: Anomalies of the p53 tumor suppressor gene have been reported for a variety of different tumor types. Also in urothelial cancer accumulation of the P53 protein has been linked with an unfavorable prognosis of the patients. Despite the growing number of publications confusion remains because key questions regarding p53 accumulation in bladder cancer are still unanswered. The objective of this manuscript was to review all published literature on the association of p53 accumulation and prognosis of patients with bladder cancer. Furthermore, putative reasons for the conflicting results should be defined as a basis for future research. METHODS: The entry criteria for the analysis were met by 43 trials comprising 3,764 patients out of 138 publications found through Medline search. RESULTS: Comparison between the trials yielded considerable differences obviously due to technical aspects, e.g. the selection of the antibody and the use of different cut-off values, study design and patient selection. CONCLUSIONS: From this analysis it becomes evident that further retrospective investigations will not contribute to the solution of the problem and thus are obsolete. There is an obvious need for standardization of the assay procedure and the assessment of the specimens as well as for the initiation of a prospective multicenter trial to provide definite answers. PMID- 11111187 TI - Bacillus Calmette-Guerin perfusion therapy for the treatment of transitional cell carcinoma in situ of the upper urinary tract. AB - OBJECTIVES: The aim of this study is to evaluate the efficacy and safety of intrarenal bacillus Calmette-Guerin (BCG) instillation as a treatment for transitional cell carcinoma in situ (CIS) of the upper urinary tract. METHODS: Diagnostic criteria of upper urinary tract CIS were (1) positive urinary cytology, (2) negative multiple random biopsy of the bladder and prostatic urethra, (3) negative radiographic findings in the upper urinary tract and (4) two serial positive cytologies in selective ipsilateral urine sampling from the pyeloureteral system. Eleven patients diagnosed as having upper urinary tract CIS were enrolled in this study. Thus, 11 renal units were treated with BCG instillation. After placing a 6-french Double-J stent, BCG (80 mg) in 40 ml saline was instilled into the bladder weekly, 6 times in total as one course. RESULTS: At the end of one course, 9 cases showed negative urinary cytology. Among these 9 cases, 2 showed recurrence in the upper urinary tract after 4 months and 8 months of disease-free interval, respectively. These 2 cases have received an additional course of BCG instillation, but the urinary cytology did not normalize. Mean recurrence-free time was 19.6 months. Of the other 7 cases who responded to the first course of instillation, 6 cases were alive with no evidence of the disease. The remaining patient died of rectal cancer with no evidence of transitional cell carcinoma (TCC). Of the 2 cases who showed positive urinary cytology even after the first course, 1 underwent nephroureterectomy. The other case was diagnosed as having malignant lymphoma 3 months after the end of this instillation therapy, and he died of malignant lymphoma. As side effects, 8 cases (72.7%) showed bladder irritability, and 4 presented fever higher than 38 degrees C. However, no patient needed antitubercular treatment. CONCLUSION: As for the short-term response, BCG instillation for the treatment of upper urinary tract CIS is considered to be effective and safe. Longer follow-up and further experience with this treatment are required. PMID- 11111188 TI - 4-Year follow-up results of a European prospective randomized study on neoadjuvant hormonal therapy prior to radical prostatectomy in T2-3N0M0 prostate cancer. European Study Group on Neoadjuvant Treatment of Prostate Cancer. AB - OBJECTIVES: To evaluate the long-term effects of 3-month neoadjuvant hormonal treatment in patients treated by radical prostatectomy for locally confined prostate cancer. METHODS: We report the results of 402 patients (220 with a clinical T2 tumor and 182 with a clinical T3 tumor) of whom 192 randomly received neoadjuvant total androgen deprivation using a LHRH analogue (goserelin) plus flutamide for a period of 3 months and 210 underwent radical prostatectomy only. RESULTS: 'Clinical downstaging' was seen in 30% of cases in the neoadjuvantly treated group (NEO). 'Pathological downstaging' occurred in 7 and 15% of cases in the direct radical prostatectomy (DP) group and the NEO group, respectively (p<0.01). In patients with clinical T2 as well as in patients with clinical T3 tumors, a significant difference in the number of positive margins was shown in favor of the NEO group (cT2, p<0.01; cT3, p = 0.01). This advantage, although there was a trend in favor of the NEO group, specifically in cT2 tumors, did not translate in a significantly better PSA progression rate (p = 0.18). However, when evaluating the local control rate in cT2 tumors, we observed local recurrence in 3 of 102 (3%) patients in the NEO group versus 12 of 114 (11%) patients in the DP group. The difference is statistically significant (p = 0.03). In the cT3 group, this difference was not statistically significant (NEO group: 15 of 87 (17%), and DP group: 21 of 95 (22%) patients; p = 0.41). CONCLUSIONS: In this study, the clinical revelance of pathological downstaging and the lower percentage of patients with positive margins in the neoadjuvantly treated group with a clinical T2 tumor is not confirmed by a lower PSA progression rate. However, this study indicates that there may be a trend that this advantage in favor of the NEO group directly translates into a better local control rate in clinical T2 tumors. Better local control in cT2 tumors is only going to be of relevance if subsequently you can show that there is a better survival for these patients. Unfortunately, this article reports a study which is not yet mature enough to show relevant information. Presently, neoadjuvant therapy should not be given outside clinical research settings. PMID- 11111189 TI - Nondetected tumor (pT0) after prolonged, neoadjuvant treatment of localized prostatic carcinoma. AB - OBJECTIVES: 135 patients with stage T1-3N0M0 prostatic carcinoma were submitted to prolonged PSA-monitored neoadjuvant endocrine treatment (PPNET). The rate of pT0 reports was three times higher (15%) than after the standard 3-month therapy (5%). The present work was done to elucidate the initial characteristics of these tumors, to see if additional workup of these prostatectomy specimens is able to detect tumor vestiges and, if so, to describe their morphology. METHODS: The original clinical and histopathological data of 20 pT0 cases were reviewed and an additional histopathological workup of the prostatectomy specimens was done. RESULTS: The majority of patients had initially small (9 patients cT1, 8 patients cT2, 3 patients cT3) and well-differentiated tumors (18 patients Gleason score <7). Microscopic assessment of 4,503 slides revealed very small tumor remnants (mean volume 0.2 ml) in 13 of the 20 prostatectomy specimens. Severe tumor regression was seen in 3 cases, slight to moderate regression in 10 cases. CONCLUSIONS: A pT0 report following detailed routine histopathological workup has to be regarded as a maximal therapeutic effect, but not as tumor elimination. PPNET clearly increases the rate of pT0 reports, implicating that the conventional 3 months of pretreatment does not exploit the possibilities of neoadjuvant therapy. PMID- 11111191 TI - Fatherhood in testicular cancer patients with carcinoma in situ in the contralateral testicle. AB - We report paternity by assisted fertilization in 3 highly oligospermic patients with stage-I unilateral testicular germ cell tumor and contralateral carcinoma in situ. If such patients plan future paternity, surveillance is the optimal treatment allowing maximal recovery of spermatogenesis after orchiectomy. Multiple semen cryopreservations should be done during the first post-orchiectomy year. PMID- 11111190 TI - Gastrointestinal presentation of germ cell malignancy. AB - OBJECTIVES: To summarize monoinstitutional experience with gastrointestinal (GI) presentations of germ cell malignancy and to review recent medical literature on this issue. METHODS: Retrospective review of 5 cases with advanced germ cell malignancy (testicular 2 and retroperitoneal 3) and involvement of the upper GI tract and a comparison with published observation. RESULTS: In 4 patients the duodenum and in 1 patient the distal part of the esophagus were involved in germ cell malignancy. In 3 patients grade 3 or grade 4 anemia represented the principle initial symptom. Ulceration of the upper GI tract was in 1 case complicated by an aortoduodenal fistula with rupture of the aorta. This patient and 2 other cases needed emergency surgery due to GI hemorrhage before and/or during the initial phase of chemotherapy. Our observations compare well with the literature, showing the need of multimodality therapy of these complications. CONCLUSION: In young males with a malignant tumor in the upper GI tract, the diagnosis of germ cell malignancy should be considered. Treatment of this condition requires a multimodality approach, not rarely including emergency surgery. Though these patients often belong to a poor-prognosis group, our results and the literature review show that long-term survival is possible using modern treatment principles. In particular, the risk of GI hemorrhage, during the initial phase of therapy, should not be overseen. PMID- 11111192 TI - Prognostic significance of microvascular invasion in localized renal cell carcinoma. AB - OBJECTIVES: The treatment of localized and even advanced renal cell carcinoma (RCC) is radical nephrectomy. However, 30% of these patients progress after radical nephrectomy. Prognostic factors are needed in order to determine the course of disease in patients undergoing radical nephrectomy. The aim of this study is to study the prognostic significance of microvascular invasion (MVI) in patients who had undergone radical nephrectomy for localized RCC. METHODS: Between June 1989 and February 1999, pathologic sections of the specimens from 41 patients without metastases, nodal involvement or macroscopic venous involvement were investigated for MVI. RESULTS: MVI was observed in 17% of the patients. MVI was related to the grade of the tumor and tumor size (p = 0.032, p = 0.017). In sarcomatoid-type RCC, MVI was more common than in other histologic types (p = 0.003). After a median follow-up of 48 months, the progression rate was 29% in patients with MVI and 17% without MVI (p = 0.001). Median progression time was 3 months in those with MVI and 41 months with no MVI (p = 0.01). The survival rate decreased from 85 to 70% in patients with MVI during a median follow-up of 48 months (p = 0.031). In multivariate analysis, MVI was not found to be an independent prognostic factor. CONCLUSION: Although MVI is closely related to progression and prognosis, in multivariate analysis it was not found to be an independent prognostic factor in localized RCC. We conclude that MVI should also be evaluated together with tumor grade in predicting the prognosis of patients with localized RCC. PMID- 11111193 TI - Abdomino-perineal repair of recurrent and complex bladder neck-prostatic urethra contractures. AB - OBJECTIVE: To evaluate the effectiveness of abdomino-perineal repair in treating complex and recurrent bladder neck-prostatic urethra contractures. METHODS: The study included 6 patients retrospectively. Their ages ranged from 66 to 75 years (67.83+/-8.13, mean + SD). All presented with a long history of voiding difficulty and urinary incontinence. Prior to definitive treatment all the patients had been subjected to multiple unsuccessful transurethral resections of bladder neck-prostatic urethra contractures. They were all subjected to diagnostic work-up including retrograde and voiding urethrogram, urethroscopy and urodynamics. Finally all the patients underwent abdomino-perineal excision of the stenotic area and end to end anastomosis. All of them received simultaneously an artificial urinary sphincter insertion for concurrent sphincteric deficiency and three were subjected simultaneously to clam type ileocystoplasty for intractable detrusor instability. The follow-up ranged from 8 to 56 months (24.42+/-19.64, mean + SD). RESULTS: The actual surgical time ranged from 3.5 to 5.5 h (4.1+0.4, mean + SD). Five patients void satisfactorily remaining unobstructed through the entire follow-up period, while one has to empty his augmented bladder by means of clean intermittent self catheterization. Five patients are continent and one significantly improved. Major complications due to infection-erosion of the prosthetic material were encountered in one patient. CONCLUSIONS: Abdomino perineal repair is an effective surgical procedure for the management of recurrent and complex bladder neck-prostatic urethra contractures, although it is time consuming and requires in the majority of cases combined techniques in order to achieve optimal results. PMID- 11111194 TI - Ureterocystoplasty in bilaterally functional kidneys. AB - OBJECTIVE: Traditional augmentation cystoplasty using gastrointestinal segments is known to be associated with metabolic abnormalities and alterations in the bladder causing potential carcinogenesis. In this respect alternative techniques have been searched preferably lined by urothelium. We performed ureterocystoplasty in 7 patients with a diagnosis of neurogenic bladder and investigated the clinical and functional aspects. PATIENTS AND METHODS: Between 1995 and 1999, ureterocystoplasty was performed using both ureters in 4 male and 3 female children with bilaterally functional kidneys. Patients' ages varied between 1 and 7 (mean 4.7) years. Before the operation all the children were incontinent, had a small-capacity noncompliant bladder, and high-grade (IV-V, International Classification System) reflux on voiding cystouretrography (VCU). Technetium-99m DTPA renal scintigraphy was also performed in all children to evaluate renal function before and after the operation. RESULTS: Before the operation the mean end-filling intravesical pressure was 45.6 (35-60) cm H(2)O which decreased to 18.9 cm H(2)O 3 months postoperatively. The mean bladder capacity 3 months after ureterocystoplasty was found to be 279.3 (250-330) ml. All the children were continent and VCU showed the absence of reflux. There was mild to moderate improvement in renal function after surgery in both kidneys on technetium-99m DTPA renal scintigraphy. Three (43%) patients could void spontaneously with abdominal straining, whereas 4 (57%) children could empty their bladders by clean intermittent catheterization. A double-J stent was inserted in 1 (14%) patient because of a rise in serum creatinine after the removal of the ureteral catheter. Patients were followed for a mean period of 30 (8-50) months and all the children remained continent. The bladder capacity and end-filling pressure measurements were also stable. CONCLUSION: Ureterocystoplasty was found to be an effective method for bladder augmentation in bilaterally functional kidneys with an acceptable complication rate PMID- 11111195 TI - Causes and treatment of residual urine volume after orthotopic bladder replacement in women. AB - OBJECTIVES: Intact innervation of the female urethra is conditional for normal urination. In the past, urethrectomy was performed as part of cystectomy. After intense anatomical studies of the female pelvis, urethral-function-sparing cystectomy was developed. METHODS: Our clinical group consists of 41 female patients who were operated from 1993 to 1998 for bladder cancer, utilizing cystectomy with orthotopic bladder replacement. RESULTS: In 28 patients, complete daytime continence was restored and in 13 patients, daytime continence was socially satisfactory (1-2 pads were used due to mild stress incontinence). The drawback of orthotopic replacements in females is the frequent development of serious residual volume, which was seen in one third of the 41 patients. The functional results of orthotopic neobladders and therapy of residual urine volume were documented using urodynamic studies. CONCLUSIONS: Postvoiding residual volume may be caused by isolated dysfunction of the urethra and can be treated with clean intermittent self-catheterization or with alpha-blockers, which improve evacuation of the neobladder. PMID- 11111196 TI - Arterial blood pressure following different types of urinary stone therapy. Presented at the 8th European Symposium on Urolithiasis, Parma, Italy, 1999. AB - OBJECTIVE: Several studies reported increased blood pressure (BP) values following extracorporeal shock wave lithotripsy (ESWL) treatment of renal stones. It is unclear, however, whether this is due to ESWL, since nephrolithiasis itself increases the relative risk of developing hypertension. Therefore we prospectively studied the BPs of stone patients undergoing different types of treatment. METHODS: 252 stone patients (63% males, 37% females, median age 44.3, range 11.7-86.4 years) participated. 168 suffered from uretral stones: 50 underwent ESWL; 40 ureteroscopy, and 78 patients passed stones spontaneously (SP). 84 had renal stones: 60 underwent ESWL; 8 percutaneous nephrolithotomy/open surgery, and 16 no treatment. Systolic (SBP) and diastolic (DBP) BP were measured according to Riva-Rocci prior to, immediately after, and 3, 6, 12, 18 and 24 months after stone therapy. RESULTS: Immediately after SP, SBP decreases, whereas after active stone treatment increases (highest after ESWL) in SBP were seen. DBP was unchanged. During the further follow-up, a gradual increase in BP was observed in all groups. At 24 months in all groups, regardless of the stone location and type of treatment, SBP and DBP were significantly higher than the pretreatment levels (p = 0.000). There was no a difference between renal and ureteral stones, or between the ESWL treatment and the other groups. CONCLUSION: Renal stone disease itself rather than the type of treatment significantly increases SBP and DBP during a follow-up period of 24 months. The underlying mechanisms remain to be elucidated. PMID- 11111197 TI - Surgical treatment of chordee without hypospadias. AB - OBJECTIVE: The aim of this study was to retrospectively assess the efficacy of the surgical techniques commonly used in three types of chordee without hypospadias (Devine and Horton classification). METHODS: Twenty-six patients, ranging in age from 3 to 14 years, had chordee without hypospadias and underwent Nesbit dorsal plication (9 cases out of 12, type III), associated extensive mobilization of the urethra (10 cases out of 10, type II), and vascularized neourethra (3 cases out of 4, type I). RESULTS: Eleven subjects undergoing follow up investigation for the milder forms of chordee were satisfied with the outcome achieved; there was no impediment of any kind in the sex lives of 5 adult subjects. Of the 11 patients undergoing follow-up after more complex surgery for Devine and Horton type I-II chordee without hypospadias, a residual abnormal curvature was present in 4 subjects. These patients were submitted to one or more reoperations with outcomes defined as satisfactory. CONCLUSION: To avoid the risks of persistent chordee, the authors suggest more radical and accurate operations for the treatment of type I and II pseudohypospadias. PMID- 11111198 TI - Paramedian extraperitoneal approach for combined nephrectomy and augmentation ureterocystoplasty in children with neurovesical dysfunction. AB - OBJECTIVES: We describe and evaluate our approach to combined nephrectomy and augmentation ureterocystoplasty using a single paramedian extraperitoneal incision. PATIENTS AND METHODS: Three patients with neurogenic bladders (2 posterior urethral valves and 1 myelodysplasia) underwent nephrectomy and augmentation ureterocystoplasty. The mean age of the patients was 4.6+/-1.5 years. The indications for the procedure included control of urinary incontinence or preservation and stabilization of renal function. RESULTS: The integrity of the peritoneal cavity was easily preserved throughout the procedure using a paramedian incision. No complications were encountered in these patients. Early postoperative resumption of normal diet and activity was noted in all patients. CONCLUSION: The paramedian extraperitoneal approach through a single incision provides the advantages seen with other extraperitoneal techniques combining two incisions. The single paramedian incision has the potential to save on operative time and obviates the need to change the patient's position on the operating table while under anesthesia. Furthermore, the paramedian extraperitoneal approach provides the reconstructive surgeon with the chance to convert the procedure into a transperitoneal technique to incorporate bowel segments in order to complement ureterocystoplasty. PMID- 11111199 TI - Impaired bone metabolism following augmentation cystoplasties in growing rats. AB - OBJECTIVE: The aim of this study was to evaluate the possible risk of impaired bone metabolism following augmentation cystoplasties with different gastrointestinal segments. METHOD: 60 young rats underwent augmentation cystoplasties using gastric, ileal or sigma segments, or sham operations. An additional group undergoing sigma-cystoplasty received the bisphosphonate ibandronate to inhibit osteoclast-mediated bone resorption. Bone mass in the lumbar spine and tibia was analyzed monthly by in vivo densitometry. Bone turnover was assessed monthly using current bone metabolism markers for a period of 16 weeks. Bone ashing and serum analyses of the osteotropic hormones parathyroid hormone (PTH), and 25-OH vitamin D3 were performed at study conclusion. RESULTS: Following ileocystoplasty, reduced bone mineral density (BMD) was seen throughout the observation period; this was pronounced in the trabecular bone. The decline in BMD was associated with decreased serum 25-OH vitamin D3 levels. Following sigmacystoplasty, bone calcium content was significantly decreased; this could be prevented by ibandronate. No skeletal changes occurred in the gastrocystoplasty group. Serum pH was not altered in any group, and markers of bone resorption indicated normal bone resorption rates. CONCLUSION: There is a significant correlation between impaired bone metabolism and the type of segment used for bladder augmentation. While the use of the ileum (and probably the colon too) causes osteopenia, gastrocystoplasties seem to have little influence on bone turnover. PMID- 11111200 TI - Monoclonality of asynchronous bilateral lymphoma of the testis. AB - OBJECTIVES: Lymphoma is the most frequent testicular malignancy in men over 60 years of age. Even though patients present initially with localized disease, the high incidence of bilateral involvement, synchronous or not, and early systemic dissemination are characteristic of these neoplasms. Sometimes the interval between tumor involvement of both testes is long. The question is raised whether either the patient has a predisposition to present new clones of transformed lymphocytes, or the same disease using the same pathway from a systemic reservoir infiltrates the contralateral testis. METHOD: Polymerase chain reaction and DNA sequencing were used to detect immunoglobulin heavy chain (IgH) rearrangement in paraffin-embedded specimens from asynchronous tumors affecting the right and left testis of a 85-year-old man with an interval period of 13 months. RESULTS: Both tumors showed the same IgH rearrangement. CONCLUSIONS: The lymphoma affecting the left and right testis derived from the same clone. It makes a strong case that lymphoma of the testis is the first manifestation of a systemic disease and should be treated aggressively early at the beginning of the disease. PMID- 11111201 TI - Paraffinoma of the external genitalia after autoinjection of vaseline. AB - Five cases of sclerosing lipogranulomas of the external genitalia after previous injection of vaseline are presented. The public should be informed about the disastrous effects of paraffin injections and the importance of a radical but organ-preserving surgical treatment. PMID- 11111202 TI - Bladder replacement and urinary diversion PMID- 11111203 TI - Randomised controlled trials versus real life practice in BPH. Introduction and concluding remarks. PMID- 11111204 TI - Invasive and minimally invasive treatment modalities for lower urinary tract symptoms: what are the relevant differences in randomised controlled trials? AB - OBJECTIVES: This manuscript reviews the outcomes of invasive and minimally invasive treatments of lower urinary tract symptoms due to prostatic enlargement. METHODS: The MEDLINE database was searched for Medical Subject headings and text words including prostatic hyperplasia, treatment, surgery, thermal treatments, thermotherapy, laser, TUNA and vaportrode. Data from both randomised and non randomised controlled trials were considered. RESULTS: All invasive treatments produce significant changes of all subjective and objective outcome parameters. The best clinical outcome has been reported for open prostatectomy followed by transurethral resection of the prostate. Complications of the different invasive techniques were difficult to analyse because of the heterogeneity of categories among different papers and lack of standard criteria. The major attraction of all minimally invasive treatment options is the low risk of bleeding requiring blood transfusions. Retrograde ejaculation was one of the most frequently reported complications for all invasive techniques. Some of the so-called less invasive treatment options appeared to be associated with a rather high incidence of minor complications somehow contradicting their minimally invasiveness. Re-treatment rate observed in patients receiving various minimally invasive treatments was always higher than following standard treatment options such as transurethral resection. CONCLUSIONS: Open prostatectomy and transurethral resection of the prostate outperform all minimally invasive treatment modalities as regards efficacy and durability of outcome. The lack of standard criteria to evaluate complications and side effects makes treatment comparisons difficult. Endorsement of the clinical research criteria proposed by the last WHO-sponsored International Consultation on BPH is strongly recommended to improve the clinical value of randomised and non-randomised controlled trials. More information is needed on long-term complications and cost-effectiveness of minimally invasive treatment modalities. PMID- 11111205 TI - Medical treatment modalities for lower urinary tract symptoms: what are the relevant differences in randomised controlled trials? AB - OBJECTIVES: Relevant differences in efficacy and tolerability will be reviewed among medical treatment modalities for lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO). Available data on the long-term effectiveness of these treatments will be also discussed. METHODS: Information reviewed here comes mainly from published scientific articles and abstracts describing direct comparative trials among alpha1-adrenoceptor antagonists, finasteride, and/or phytotherapy. RESULTS: Direct comparative trials demonstrated alpha1-adrenoceptor antagonists to be more effective than finasteride in improving symptoms and increasing urinary flow. Moreover, finasteride did not perform better than placebo in those studies that included a placebo arm. While finasteride treatment appears more beneficial in patients with an enlarged prostate volume (>50 ml), the efficacy of alpha1-adrenoceptor antagonists is not related to prostate size. To study the efficacy of plant extracts, adequately performed placebo-controlled and direct comparative trials are needed. Medical treatment modalities generally have a low incidence of adverse events. Regarding long-term effectiveness of medical treatment, the few available data show that finasteride can reduce the risk of acute urinary retention (AUR) and surgery. Short-term, direct comparative studies suggest that, like finasteride, alpha1 adrenoceptor antagonists have a comparable positive effect on disease progression. CONCLUSION: More comparative information is needed on the long-term efficacy, tolerability, and effectiveness of medical treatments for LUTS. Information on disease progression (i.e., long-term complications related to BPO) and treatment outcomes (i.e., switch to other therapy or surgery) is necessary because such information directly effects a treatment's cost-effectiveness. PMID- 11111206 TI - Alpha1-blocker therapy for lower urinary tract symptoms suggestive of benign prostatic obstruction: what are the relevant differences in randomised controlled trials? AB - Randomised controlled trials (RCTs) provide the best available external evidence for the use of alpha1-blockers in treating lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO). Placebo-controlled and actively controlled RCTs evidenced the efficacy of alpha1-blockers in augmenting urine flow, relieving symptoms, reducing bother and improving quality of life in patients with LUTS. This improvement involves both filling (irritative) and voiding (obstructive) symptoms, occurs promptly and is well-maintained over time. Treatment benefit is independent of prostate size and baseline prostate specific antigen (PSA). There is no evidence of relevant differences between the different alpha1-blockers in this regard and all alpha1-blockers can be accepted as appropriately efficacious at the presently recommended doses. The best available external evidence for relevant differential properties of alpha1-blockers relates to their clinical selectivity in terms of the absence/presence of ancillary cardiovascular, i.e. anti-hypertensive effects. Anti-hypertensive alpha1-blockers (terazosin and doxazosin in particular) are more likely to cause dizziness and other cardiovascular untoward effects, eventually leading to premature treatment discontinuation. Alfuzosin (although primarily developed as an anti-hypertensive agent) and tamsulosin in contrast, are better tolerated; the former nevertheless carries a more distinct risk of symptomatic impairment of blood pressure control. Although indirect comparisons between different studies suggest a higher risk of retrograde ejaculation with tamsulosin, this hypothesis failed to be confirmed in direct comparative RCTs. PMID- 11111207 TI - Treatment satisfaction of patients with lower urinary tract symptoms: randomised controlled trials vs. real life practice. AB - Randomised controlled trials (RCTs) are an important scientific tool to determine the efficacy and tolerability of a given treatment relative to placebo or other treatment forms. However, due to strict inclusion and exclusion criteria the patient populations in RCTs may not be fully representative for those routinely consulting the physician. Moreover, participation in a formal study puts physician and patient in a situation where they may react different than in real life. In contrast real life practice (RLP) studies cannot determine treatment efficacy or tolerability in absolute terms since they typically do not include a control group and are purely observational. On the other hand, they tend to be more representative for real treatment outcomes. Thus, RCTs have high internal but less external validity whereas RLP studies have less internal and greater external validity. Hence, RCTs and RLP studies should not be considered as mutually exclusive but rather as complementing each other. Specific advantages and disadvantages of RCTs and RLP studies will be discussed using published evidence for the treatment of lower urinary tract symptoms suggestive of benign prostatic obstruction with alpha1-adrenoceptor antagonists and other treatments. PMID- 11111208 TI - How do symptoms indicative of BPH progress in real life practice? The UK experience. AB - OBJECTIVE: Lower urinary tract symptoms (LUTS) are usually, but not exclusively associated with benign prostatic hyperplasia (BPH). Using a population identified from the UK General Practice Research Database (GPRD), we describe the changes in the management of LUTS/BPH and assess the effectiveness of medical therapy between 1992 and 1998. METHODS: 61,364 men with LUTS/BPH and without a record of prostatic cancer were identified on the database. 14,195 were treated with an alpha1-blocker or finasteride. Treatment failure was defined as prostatic surgery, catheterisation or a switch in medical therapy. RESULTS: LUTS/BPH incidence increased linearly from the age of 45 to 85 years (r2 = 0.992) and prevalence increased from 3.5% to 35% for men in their late 40s and 80s respectively. Prostatectomy rates increased linearly from the age of 50 to 80 years (r2 = 0.984). Between 1992 and 1998, total treated-patient time had increased 3-fold, patients have been medically treated earlier and have increasingly been prescribed the LUTS/BPH-specific treatments finasteride, tamsulosin and alfuzosin in comparison to older treatments (indoramin, prazosin). In parallel, there has been a progressive increase in the interval between first diagnosis and prostatic surgery, and this interval is significantly longer for medically treated patients than those receiving no medical therapy. The intervals between the start and failure of medical therapy were significantly shorter for patients receiving indoramin and prazosin than for those receiving specific LUTS/BPH treatments. CONCLUSIONS: Between 1992 and 1998 there has been a significant lengthening of the period between first diagnosis of LUTS/BPH and surgery. This postponement of surgery is associated with earlier treatment and the increased use of specific LUTS/BPH treatments that appear more effective than older products in delaying treatment failure. PMID- 11111209 TI - How are lower urinary tract symptoms managed in real life practice? The French experience. AB - OBJECTIVES: We tried to outline the management of LUTS suggestive of BPO in real life practice in France. In addition, preliminary results regarding switch rates of different initial treatment approaches in primary care are given in order to assess the effectiveness of therapies. METHODS: IMS and GERS market data of drugs for LUTS suggestive of BPO in France are presented. In addition, data from the THALES database for 1997 and 1998 containing the patient medical records of 620 French GPs were analysed with special emphasis on the initial management by GPs and switch rate to another treatment option. RESULTS: The prevalence and incidence of LUTS suggestive of BPO is high and more than half of the French LUTS patients receive a medical treatment from their GP. alpha1-Adrenoceptor antagonists are prescribed most in the management of LUTS suggestive of BPO in France and their use is still increasing. Finasteride is prescribed much less whereas the use of phytotherapy is, although decreasing, still rather high. The switch rate of tamsulosin as initial treatment to another treatment option is low (14.4%), compared to other therapies like terazosin (26.5%), reflecting its favourable benefit/risk ratio in real life practice. CONCLUSIONS: In France, the prescription rate of alpha1-adrenoceptor antagonists, and especially tamsulosin, is high and still growing. Further information is however needed to study the impact of initial treatment choice on the progression of LUTS suggestive of BPO. PMID- 11111210 TI - Increased nuchal translucency in a case of long-chain 3-hydroxyacyl- coenzyme A dehydrogenase deficiency. AB - We present a case where the embryo showed an increased nuchal edema and a metabolic disorder. At 31 weeks of gestation the fetus developed a cardiomegaly and a hydrops. In this case, a long-chain 3-hydroxacyl-coenzyme A dehydrogenase deficiency (LCHAD deficiency) was confirmed by biochemical investigations in cultured chorionic villus cells and by DNA analysis. This metabolic disease causes a reduced production of mitochondrial trifunctional proteins and is a very rare autosomal-recessive disease. PMID- 11111211 TI - Adenosine triphosphate for cardioversion of supraventricular tachycardia in two hydropic fetuses. AB - OBJECTIVE: We performed a retrospective study to check the effectiveness of adenosine triphosphate (Striadyne) for cardioversion of fetal supraventricular tachycardia (SVT) and to evaluate neonatal outcome after prenatal treatment of severe SVT with fetal hydrops. METHODS: Two hydropic fetuses with SVT were treated with Striadyne injection into the umbilical vein, as an additional treatment to the digoxin given intravenously to the mother. Both fetuses were in severe condition, with ultrasound, Doppler and laboratory signs of fetal distress and congestive heart failure. RESULTS: Sinus rhythm was obtained in both cases for different periods of time, without side effects of Striadyne. The children survived. There were severe cardiac and neurologic problems after delivery. CONCLUSIONS: Striadyne was an effective drug in converting SVT to the sinus rhythm in hydropic fetuses. Digoxin was useless in these fetuses in spite of the therapeutic level which was obtained in both mothers. We suppose that fetal SVT causing fetal hydrops could be the reason of brain damage, and intensive antiarrhythmic treatment seemed to be necessary. PMID- 11111212 TI - Aneuploidy and isolated mild ventriculomegaly. Attributable risk for isolated fetal marker. AB - BACKGROUND: Does the prenatal ascertainment of isolated mild ventriculomegaly increase the a priori risk for aneuploidy when isolated or not associated with advanced maternal age? Does isolated mild ventriculomegaly increase the risk for pediatric developmental delay? METHODS: The Wayne State University (WSU) Reproductive Genetics abnormal case data base and the Madigan Army Medical Center (MAMC) experience were reviewed to compare the rates of aneuploidy for cases with fetal ventriculomegaly. Cases were classified by maternal age and associated sonographic markers of aneuploidy. Aneuploidy rates were compared between the isolated ventriculomegaly, ventriculomegaly with advanced maternal age (AMA), and ventriculomegaly associated with multiple anomalies. Rates of aneuploidy were compared to identify association. RESULTS: A total of 118 cases with ventriculomegaly were identified for comparison. Ninety-four cases were identified in the WSU cohort; 46 demonstrated isolated ventriculomegaly alone, and aneuploidy was present in 3/25 (12%) with invasive fetal testing, 0/24 (0%) cases in the MAMC cohort demonstrated aneuploidy. Isolated mild ventriculomegaly cases at MAMC were identified for further tests. DISCUSSION: Although the two study populations vary in age and risk distributions, the attributable risk for isolated mild ventriculomegaly poses a counseling conundrum due to the neurodevelopmental implication of this minor dysmorphism more so than its association with aneuploidy. PMID- 11111213 TI - Prenatal diagnosis of smith-magenis syndrome (del 17p11.2). AB - Smith-Magenis syndrome is associated with a microdeletion of the short arm of chromosome 17 with phenotypic abnormalities including dysmorphic facies, self injurious behavior, mental and neurologic disturbances, and congenital cardiac defects. The majority of patients present in mid-childhood or adulthood. We describe a fetus in which the diagnosis of Smith-Magenis syndrome was made at 16 weeks of gestation following amniocentesis for increased risk for Down syndrome detected by second-trimester maternal serum screening. Ultrasound evaluation revealed multiple fetal anomalies. The pregnancy was terminated at 20 weeks of gestation. Post-mortem findings included dysmorphic facial features, tetralogy of Fallot, a thymic duct remnant, pancreatic islet cell hyperplasia, and abnormal lung fissuring. This represents the second case of prenatally diagnosed Smith Magenis syndrome. Molecular genetic techniques in the diagnosis of the Smith Magenis syndrome and other small deletions are becoming an important tool in the genetic evaluation of ultrasound abnormalities. PMID- 11111214 TI - Parental decision-making differences between patients in two healthcare systems for choroid plexus cysts. AB - OBJECTIVE: We evaluated the medical-sociological implications of parental perception of risk and decision-making choices for prenatally ascertained choroid plexus cysts (CPCs) between two obstetric populations with similar clinical situations. METHODS: The Wayne State University (WSU) Reproductive Genetics database and the Madigan Army Medical Center (MAMC) experience were reviewed to compare the rates of aneuploidy and invasive testing for cases with CPC. Aneuploidy rates were compared between those with isolated CPC, CPC with advanced maternal age (AMA), and CPC associated with multiple anomalies. RESULTS: 186 cases were identified in the WSU cohort, of whom 27 (15%) declined invasive fetal testing. In the remaining 159 cases, aneuploidy was present in 2/132 (1.5%) isolated CPCs, 3/11 (27%) CPCs with AMA, and 15/16 (93%) CPCs with multiple anomalies. 107 cases were identified in the MAMC cohort, of whom 99 (92%) declined invasive fetal testing. No cases of aneuploidy were found in the 3/12 AMA cases or 5/95 non-AMA cases who underwent amniocentesis. CONCLUSIONS: The 2 cases of aneuploidy with isolated CPC cannot be ignored, and provide an estimated attributable risk of at least 0.8%, a higher risk than 38 years of age. However, the parental sociologic context may be as important as the genetic-prognostic risk for decision-making. PMID- 11111215 TI - Fetal obstructive uropathy in trisomy syndromes. AB - Fetal obstructive uropathy has seldom been described in trisomy syndromes, and its relationship to these syndromes remains unclear. Five trisomic male fetuses, four with trisomy 18 and one with trisomy 21, were identified out of 110 fetuses evaluated for fetal obstructive uropathy. We performed detailed examination on the urinary tracts of four of these fetuses, three with trisomy 18 and one with trisomy 21, following termination in the second trimester. All four had a markedly distended urinary bladder (megacystis), abdominal wall distension, and a small, poorly developed urethra thoughout its full length. All four also had poor development of the prostate with virtual absence of glandular development, as compared to age-matched controls. Posterior urethral valves were not identified in any case. Three of the fetuses (two with trisomy 18 and one with trisomy 21) had unilateral or bilateral hydroureters, and resulting renal tubulocystic or glomerulocystic change. Review of this database reveals an unexpectedly high frequency of trisomies, particularly trisomy 18, suggesting that the relationship may not be coincidental. Abnormal prostate development may be causally related to fetal obstructive uropathies and may be an under-recognized trait in trisomy syndromes. Karyotypic analysis of all fetuses with obstructive uropathy is important since in utero surgical intervention may be contraindicated in cases of fetal aneuploidy. PMID- 11111216 TI - Frequency of prenatal diagnosis of birth defects in Houston, Galveston and the Lower Rio Grande Valley, Texas 1995. AB - BACKGROUND: Estimates of the proportion of birth defects diagnosed before birth exist for only a few types of birth defects and for a few geographic regions in the United States. This population-based study examines rates of prenatal diagnosis for previously unstudied birth defects in a new geographic region. METHODS: Active surveillance of 23 categories of birth defects among 111,902 infants born in 77 birthing hospitals in Texas in 1995 identified 852 infants or fetuses with major birth defects. Surveillance was conducted by the Texas Birth Defects Monitoring Program of the Texas Department of Health. Two regions were covered, the Houston/Galveston metropolitan area as well as the Lower Rio Grande Valley of Texas. Rates of prenatal diagnosis were evaluated for 23 different types of birth defects, using proportions and 95% confidence intervals. RESULTS: One third of the 852 infants or fetuses with birth defects were prenatally diagnosed. Diagnosis rates varied greatly depending on the type of birth defects and were lower among infants born to Black and Hispanic women. More than 60% of anencephaly, encephalocele, gastroschisis and trisomies 13 and 18 were diagnosed antenatally. Many of the fetuses that were electively terminated had birth defects or combinations of birth defects that were potentially lethal. Prevalence rates for birth defects generally do not include fetuses that die or are electively terminated before 20 weeks of gestation. Thus, 36% of anencephaly, 21% of omphalocele, 15% of encephalocele and between 7 and 10% of spina bifida, hydrocephaly, renal agenesis, and trisomies 13, 18, and 21 were not included in our published rates. CONCLUSIONS: Published rates for specific types of birth defects are spuriously low. This should be considered when investigating alleged clusters and comparing rates of birth defects across geographic areas. Since many elective abortions are for lethal or potentially lethal birth defects, a major effect of prenatal diagnosis is the resultant decrease in infant mortality attributable to birth defects. PMID- 11111217 TI - The resolution of fetal hydrops using combined maternal digoxin and dexamethasone therapy in a case of isolated complete heart block at 30 weeks gestation. AB - The development of hydrops fetalis in cases of isolated complete heart block is associated with a very poor prognosis. Various pharmacological strategies have been proposed, involving both direct fetal access and transplacental therapy, with inconsistent results in small numbers of subjects. The optimal antenatal management will remain uncertain until multicentre controlled trials are organised. We report the complete resolution of fetal hydrops at 30 weeks of gestation using combination of maternal digoxin and dexamethasone therapy, despite persistence of the complete heart block. A Caesarean section was performed at 37 weeks of gestation due to evidence of fetal intrauterine growth restriction. The baby girl is now 8 months of age and remains well, with a heart rate of 45-50 beats per minute on no medication and without pacing. PMID- 11111218 TI - Percutaneous laser ablation of fetal congenital cystic adenomatoid malformation: too little, too late? AB - OBJECTIVE: Congenital cystic adenomatoid malformation, type III (CCAM III) lesions are large, bulky tumors which can cause mediastinal shift, prevent normal pulmonary growth, and compress the esophagus, thus leading to complications of nonimmune hydrops, pulmonary hypoplasia and polyhydramnios. Because the mortality rate of untreated fetuses with CCAM and hydrops is high, early delivery or intrauterine resection of the enlarged pulmonary lobe (lobectomy) is indicated; however, open fetal resection of CCAM at less than 30 weeks is associated with perioperative mortality that approaches 40%, as well as the usual maternal and fetal morbidity of open fetal surgery. As an alternative, percutaneous laser ablation of a CCAM III lesion with hydrops was attempted. METHODS: A 30-year-old G3 P1011 with CCAM III in the left fetal hemithorax developed mediastinal shift, hydrops and polyhydramnios at 23 weeks' gestation. After pregnancy termination and open fetal resection were declined, an 18-gauge needle was placed into the fetal tumor percutaneously under real-time ultrasonographic guidance, using sterile technique with light sedation. A cleaved 400-microm Nd:YAG laser fiber was passed through the needle lumen, and using a power setting of 15 W, a total of 2,943 J of laser energy was delivered in pulses of 1.0 s at 0.2-second intervals over two sessions one week apart. RESULTS: Although tumor size decreased, the hydrops worsened and fetal death occurred. CONCLUSIONS: The fetus with CCAM complicated by hydrops is already so compromised by the advanced state of the disease that insufficient time is available for necrotic tissue reabsorption after minimally invasive therapy with laser energy. Until earlier markers for intervention are determined, percutaneous laser debulking of CCAM is unlikely to be successful. PMID- 11111219 TI - Ultrafast scan magnetic resonance in prenatal diagnosis. AB - OBJECTIVE: To determine whether magnetic resonance (MR) can give additional information in prenatal diagnosis of congenital anomalies, when the ultrasound (US) analysis is not conclusive. METHODS: Ultrafast MR scanning examined 39 pregnant women with 41 fetuses in whom US was suspicious of fetal congenital abnormalities. Two techniques were used namely (1) HASTE inversion recovery sequence and (2) FISP 2D. RESULTS: Thirty-nine patients with 41 fetuses were referred for MR because of an equivocal US with regard to brain, spine, skeletal and miscellaneous anomalies. In 1 twin pregnancy, 1 co-twin has not been examined with MRI because of its demise. In 22 of them, additional information was obtained by MR. In 9 the MR was confirmative with the US examination. Four were false negative, comparing with the postnatal diagnosis. Three failed because of maternal claustrophobia and in 2 a diagnosis could not be made. From the 40 fetuses in this study, 38 were examined postnatally by MR, US, plain X-ray or autopsy was performed to confirm the prenatal diagnosis. CONCLUSION: The use of MRI in obstetrics has been limited, until recently. With fast MRI sequences it is not necessary to sedate the fetus. It is advisable in cases where US is equivocal concerning congenital anomalies of the fetus to use MR with fast or ultrafast scan technique, especially when the central nervous system is concerned. PMID- 11111220 TI - Sonographic measurement of the fetal iliac angle as a marker for trisomy 21. Concerning the article by Grange et al. PMID- 11111221 TI - The roots of geriatric medicine: care of the aged in Byzantine times (324-1453 AD). AB - BACKGROUND: The search for the roots of geriatric medicine, which has been considered a relatively new branch. OBJECTIVE: The purpose of the study is the research of the original Byzantine medical texts and the contemporary historical sources so as to bring to light knowledge about ancient medical care. METHODS: The medical texts of Byzantine physicians were studied and analysed, as well as the Histories and Chronicles of their contemporary writers, so as to locate the extracts in the texts concerning geriatric care from the scientific point of view and that of the 'vox populi' which the historians and chroniclers express. RESULTS: The problems of old age occupied physicians from earliest Byzantine times. They had dealt with the characteristics, symptoms and accompanying diseases of the aged and endeavoured to confront all the medical problems faced by the elderly, providing a special healthy regimen for the third age and taking steps for the prevention of diseases of this age group and their treatment. Parallel to this, the research of contemporary historical texts proves the concern of all society for the special problems of the aged and the significant impact of scientific geriatric medicine on the population. CONCLUSION: The study and analysis of the original medical and historical texts of the Byzantine period (324-1453 AD), written in Greek language, prove that the roots of medical care of old age could be traced from ancient Greek and Byzantine medicine. PMID- 11111222 TI - Glutathione-S-transferase in the ageing rat brain cerebrum and the effect of chlorpromazine. AB - Lipid peroxidation increases during ageing and has been implicated in the pathogenesis of degenerative processes associated with ageing. Despite the importance of the enzyme glutathione-S-transferase (GST) in the biotransformation and detoxification of lipid peroxidation products, there have been extremely limited studies of GST in the ageing brain. The drug chlorpromazine is known to have an activating influence on the activities of certain antioxidant enzymes (glutathione peroxidase, superoxide dismutase, etc.) and it also has anti-lipid peroxidative and anti-lipofuscin influences. Therefore, information about the age related changes in brain GST and the effect of chlorpromazine on it in the ageing brain will be of further interest. We have, therefore, studied the effect of age on the activity of GST in the whole homogenate and cytosol fractions from the cerebral hemispheres of rats aged 1, 2, 3, 6, 12, 18 and 24 months. The effect of chlorpromazine treatment (10 mg/kg i.p. on alternate days for 6 months) was examined in the whole homogenate and cytosol fraction from the cerebral hemispheres of 6-, 12-, 18- and 24-month-old rats. The results showed that the values for GST specific activities in the cytosol fraction from all the age groups were higher then those in the whole homogenate; and the pattern of age changes in the whole homogenate differed from that in the cytosol. In the cytosol fraction the enzyme activity showed several phases of alterations: a progressive increase at 3 months of age, followed by a steady level up to 12 months of age; this phase was followed by a fall in the activity (at 18 months of age) then turned to a gradual increase. In the whole homogenate there were two phases of alterations: a progressive increase up to 12 months of age, which then turned to a somewhat gradual decrease with ageing. Thus, the decline in GST activity during ageing was evident in both the whole homogenate and cytosol. The results from chlorpromazine experiments showed that the drug elevated the GST activity in 12-, 18-, and 24-month-old animals but not in the 6-month-old animals. The drug's effects were most profound in 12-month-old animals. In conclusion, this study demonstrated an impairment of brain GST activity during ageing and the results further showed that the drug chlorpromazine attenuated the age-related impairment in the enzyme activity. The enhancement of the GST status of the ageing brain following chlorpromazine treatment is indicative of an additional antioxidative property of this drug. PMID- 11111223 TI - Senile tremor. What is the prevalence and severity of tremor in older adults? AB - BACKGROUND/SETTING: It is well recognized that mild tremor is common among older adults, but the prevalence and clinical characteristics of this tremor have not been studied in detail. OBJECTIVES: To examine a cohort of normal older adults to: (1) ascertain the prevalence of mild test-detectible tremor; (2) quantify the severity and functional impact of this tremor, and (3) determine whether age, gender and concomitant illness predict the severity of tremor. PARTICIPANTS: 76 normal older adults >55 years of age (mean age = 73.7 years). DESIGN: Healthy older adults were identified in a community-based case-control study of essential tremor in Washington Heights-Inwood, New York. All subjects underwent a medical interview and a videotaped neurological examination. The examination included six tests: arm extension, pouring water, drinking water, using a spoon, finger-to nose movements, and drawing spirals with each arm. Two neurologists rated the severity of tremor using a 0 to +3 clinical rating scale and a total tremor score (TTS) was calculated (range = 0-36). Forward stepwise linear regression was used to determine the association between TTS and other variables. RESULTS: Virtually all (75 or 98.7%) showed signs of tremor (TTS > 0.5). The mean TTS = 6.3 (range = 0-14.5), corresponding to a tremor that was either mild or intermittent. Twenty eight of 76 (36.8%) received tremor ratings of +2 (clearly oscillatory tremor of moderate amplitude and usually present) during at least one of the six tests; a tremor rating of +2 was 2.1 times more likely to occur in the nondominant than in the dominant hand. Those who were aged 57-74 years had a lower mean TTS (5.8) than those aged 75-93 (6.8), but this was not significant. Only 2 patients (2.6%) answered 'yes' to the question 'do you have uncontrollable shaking in your hands?' None was taking medication to treat tremor. Gender, ethnicity, concomitant illness (diabetes, arthritis, heart disease), and medications were not associated with a higher TTS. CONCLUSION: Mild but test-detectible tremor was present in almost all normal older adults, and in one-third this tremor attained a moderate amplitude during at least one activity. Characterization of this tremor would be of value to practitioners who care for older adults. PMID- 11111224 TI - Endometrial cancer in elderly women: A histologic and steroid receptor study. AB - BACKGROUND: Information on steroid receptor content in endometrial tissue of aging women is limited and somewhat controversial. The high incidence of endometrial cancer (EC) and the implication of hormone replacement therapy (HRT) in this group prompted the investigation of steroid receptors and endometrial cancer histology in the elderly. OBJECTIVE: Review of histologic characteristics correlated with estrogen and progesterone receptors (ER and PR) status in EC in women over 75 years of age in order to determine the prevalence of a more aggressive endometrial neoplasm arising in late postmenopausal atrophic endometrium of elderly patients. METHODS: Histologic slides and deeper sections stained immunohistologically for ER/PR from 54 cases of EC in women aged 75-95 years were reviewed. The histologic characteristics and degree of differentiation were correlated with the steroid receptor status, evaluated on a scale of 0-3. Benign endometrial tissue from women of the same group was used as controls. RESULTS: The 57.4% endometrioid adenocarcinomas were mostly moderately and poorly differentiated. The nonendometrioid carcinomas were anaplastic, papillary, clear cell, squamous cell, mixed mullerian and nongestational choriocarcinoma. The staining intensity for ER/PR decreased with the degree of dedifferentiation being weak or absent in nonendometrioid tumors. CONCLUSION: Elderly patients have less differentiated EC displaying histologically nonendometrioid patterns ('alienation') with no differential loss of receptors in cancer. ER/PR are partly preserved in endometrioid tumors and controls. We conclude that differential loss of receptor capacity is not a factor in pathogenesis of this age-related cancer. PMID- 11111225 TI - Acute phase markers are associated with reduced plasma lipid levels in a population of hospitalized elderly patients. AB - BACKGROUND: Several epidemiological studies have documented the presence of a 'J' or 'U' association between total cholesterol levels and total mortality. Not only the mechanism underlying the association between increased mortality and low total cholesterol values is not completely clear, but the relationship itself also appears to be complex in the elderly. OBJECTIVE: The aim of the study was to evaluate the possible association between some biohumoral markers of the acute phase, comorbidity, disability, and reduced levels of some lipoprotein parameters in a sample of hospitalized elderly subjects. METHODS: 341 patients over 65 years of age (185 males, 156 females; mean age 76.2 years), consecutively admitted to our department from 1994 to 1995, were studied. Acute phase was defined as the simultaneous presence of: (1) increased alpha2-plasma protein on electrophoresis (>12%); (2) high fibrinogen concentration (>450 mg/dl), and (3) increased blood sedimentation rate (>15 and >20 mm 1 h in males and females, respectively). RESULTS: The prevalence of signs of acute phase was higher in males and in the youngest patients, but did not change with the level of comorbidity. Patients with signs of acute phase were characterized by lower total, low-density lipoprotein (LDL)- and high-density lipoprotein (HDL)-cholesterol levels compared to subjects without signs of acute phase; this difference was significant even after adjustment for indicators of comorbidity, disability, and nutritional status. Multivariate logistic regression analysis evidenced that the simultaneous presence of these three markers of acute phase was independently associated with low levels of total cholesterol [odds ratio (OR) 2.1, 95% confidence interval (CI) 1.1 - 3.9], and HDL-cholesterol (OR 2.3, 95% CI 1.2 - 4.2), considered as the sex-specific first quintile. CONCLUSION: The findings of this study demonstrate an independent association between acute phase markers and low levels of total and HDL-cholesterol, suggesting that recognized or subclinical diseases in elderly patients may determine a reduction in these plasma lipids. Low level of total and HDL-cholesterol should be considered as possible clinical markers of an underlying state of acute phase rather than a sign of malnutrition. Given the high prevalence of chronic diseases in the elderly, epidemiological studies addressing the lipid profile in this age group should take into account the possible confounding effect of the presence of signs of acute phase. PMID- 11111226 TI - Risk factors and incidence of postoperative delirium in elderly Chinese patients. AB - OBJECTIVES: To investigate the incidence of postoperative delirium among elderly patients and to examine the interrelationship between basic vulnerability and precipitating factors for delirium. DESIGN AND METHODS: This was a prospective cohort study. Data were collected in a tertiary medical center in Taipei, Taiwan. From the 1st to the 5th postoperative day, nurses assessed patients using a confusion-screening tool. Patients with signs of delirium were closely examined for changes in behavior and cognitive status and vital signs, and laboratory data were collected to further validate the organic etiology of delirium. Patients were finally diagnosed according to the DSM-IV criteria in consensus meetings. SUBJECTS: Seven hundred and one elderly patients, that were admitted consecutively for elective orthopedic or urologic surgery, were enrolled in this study. All subjects met the following criteria: (1) 65 years of age or older; (2) able to communicate orally in Chinese, and (3) not unconscious, delirious, deaf, or aphasic upon admission. RESULTS: The overall incidence of delirium among these subjects was 5.1%. Logistic regression analysis identified that older age and preexisting cognitive impairment were vulnerability factors, and that the use of psychoactive drugs was a precipitating factor for delirium. Patients with both basic vulnerability and the precipitating factor had a 56-fold increased probability of delirium (0.28 vs. 0.005 in comparison with those who did not exhibit these factors). CONCLUSION: Few risk factors of postoperative delirium in the older Chinese sample were identified. The only modifiable risk factor appears to be the use of psychoactive drugs. PMID- 11111227 TI - Acute confusional states in patients undergoing hip surgery. a prospective observation study. AB - BACKGROUND: There is general agreement that acute confusional state (ACS) is common among elderly patients admitted to hospital, although exact figures are difficult to obtain. The objective of the current study was to investigate the onset of ACS during hospital stay and to isolate possible predisposing, facilitating and precipitating factors associated with the onset of ACS. METHODS: Non-confused patients, greater than or = 65 years of age, undergoing orthopedic hip surgery, were consecutively included in the study (n = 225). Of these, 149 patients were operated on because of acute hip fracture and 76 underwent elective hip-replacement surgery. ACS was diagnosed by the DSM-IV criteria for delirium. Structured observations of ACS onset were performed every 2nd to every 4th hour during the patients' entire hospital stay. A protocol was used to document the observations on sleep, activities, well being and behavior. The Mini-Mental State Examination was used to measure cognitive functioning. RESULTS: Of 225 patients 20% were diagnosed with ACS. The incidence of ACS was 24.3% in the group of hip fractured patients and 11.7% in the hip-replacement surgery patients. The onset of ACS was postoperative (mean 24 +/- 21 h after surgery) in all but 8 patients. The duration of ACS among recovered patients was generally less than 48 h (mean 42 +/- 43 h). Predisposing factors were older age, cognitive impairment and pre existing cerebrovascular or other brain diseases. Facilitating factors related to ACS were associated with communication and social isolation, e.g. impaired hearing and sight, reticence and passivity. One precipitating factor, besides surgery, may be the use of psychopharmacological drugs. CONCLUSION: The incidence of ACS was 20% among hip surgery patients. Older age and social isolation were factors associated with ACS. Increased attention and interaction with older patients could be of value in avoiding ACS during hospitalization. PMID- 11111228 TI - Measuring human age by estimating lifetime caloric consumption. AB - BACKGROUND: This study was prompted by a desire to measure human age by a means other than chronological or biological age. The purpose of the study was to use the total lifetime caloric intake in men as an index of aging. METHOD: The study was carried out by developing an algorithm to approximate the lifetime caloric intake. The total caloric intake is expressed such that one calorie-age year is defined as the total calories expended by an individual during his/her 20th year. RESULTS: The results indicated that (1) the caloric intake of men progressively decreases with advancing age, that (2) the caloric-age is usually less than chronological age, and that (3) the caloric-age algorithm is a simple although crude method to assess lifetime caloric intake. CONCLUSION: The most important conclusion is that a person can get a better appreciation of the importance of avoiding excess caloric intake if human age is measured in terms of lifetime caloric intake. PMID- 11111229 TI - Visual analogue scales for pain assessment in Alzheimer's disease. AB - BACKGROUND: In earlier studies, pain assessment in patients with Alzheimer's disease (AD) was conducted by interview, for which reliability is questionable considering the decline in expressive and receptive language abilities in AD. As similar language problems occur in young children, the reliability of pain assessment in this latter population is increased by employing visual analogue scales. OBJECTIVE: By employing visual analogue scales, the current study investigated whether (1) nondemented elderly persons and AD patients comprehend the purpose of the scales and (2) AD patients, compared to nondemented elderly persons, report suffering less pain intensity and pain affect. METHODS: Three visual analogue scales, i.e. the Colored Analogue Scale (CAS), the Faces Pain Scale (FPS), and the Facial Affective Scale (FAS) were administered to patients in an early and midstage of AD and to nondemented elderly persons. RESULTS: The results show that the percentage of subjects who comprehended the CAS, FAS and FPS was for the nondemented elderly persons 100, 75 and 100%, respectively, for the early AD group 100, 50 and 60%, respectively, and for the midstage AD group 80, 20 and 30%, respectively. Furthermore, elderly persons without dementia reported experiencing more intense pain and pain affect than the early and midstage AD group. Interestingly, the early and midstage AD patients did not differ in reporting pain affect. CONCLUSION: Visual analogue scales may improve pain assessment in those AD patients who fully comprehend the meaning of the scales. As only the minority of midstage AD patients understood the purpose of the FAS and FPS, the search for tools, particularly to assess pain affect in this population, must continue. PMID- 11111230 TI - Benefits and complications of regular blood transfusion in patients with beta thalassaemia major. AB - Early and regular blood transfusion therapy in patients with homozygous beta thalassaemia decreases the complications of severe anaemia and prolongs survival. In the long term, however, the beneficial effects of transfusions are limited by the organ damage resulting from iron overload, a consequence of the body's limited capacity to excrete iron, and by the complications of infection with blood-borne agents. Transfusion regimens for beta-thalassaemia have changed substantially during the past four decades. In current protocols, pre-transfusion haemoglobin concentration should not exceed 95 g/l. This allows adequate control of anaemia, with a relatively low rate of iron accumulation. Although iron chelation therapy has successfully improved survival free from cardiac disease, thalassaemic patients continuously present new clinical challenges. In fact, the vast majority of them suffer from post-transfusion chronic hepatitis C, which is expected to significantly contribute to morbidity in the forthcoming years. Furthermore, recent studies demonstrated that thalassaemics are at high risk of acquiring several blood-borne viruses. The potential role of these multiple infections in inducing clinical disease is still uncertain, and needs to be thoroughly clarified in future surveys. PMID- 11111231 TI - Hepatitis C virus: routes of infection and genotypes in a cohort of anti-HCV positive French blood donors. AB - BACKGROUND: We evaluated and analysed risk factors of HCV-infected blood donors according to HCV genotypes in order to improve the transfusion policy and safety of blood supply. MATERIALS AND METHODS: HCV-RNA was analysed in sera from 518 anti-HCV-positive blood donors, who were invited to medical consultation and interview as to risk factors by means of an extensive questionnaire. HCV genotyping was done on all samples positive for HCV-RNA. RESULTS: Of the 518 sera, 399 (77%) were HCV-RNA positive, and 394 of 399 HCV genotypes were identified. Major genotypes were 1b (34.3%), 3a (24%), 1a (19.5%) and 2 (11.4%). Of the donors, 289 (55.8%) were interviewed regarding their risk behaviour: 27% were former intravenous drug users (IVDUs), 26% had been transfused, 8% had a history of invasive diagnostic procedures, and 13% a history of surgery. Among the 224 interviewed donors, genotypes 1a and 3a were mainly associated with IVDU (51 and 45% respectively) and genotype 1b, with transfusion and nosocomial infections (40 and 25%, respectively). CONCLUSION: In this population of anti-HCV positive blood donors, nosocomial infection may be a route of HCV spread, but the main risk factor remains IVDU, particularly in young men. The transfusion policy will improve if predonation interviews of such young men are done with a specific and sensitive questionnaire. PMID- 11111232 TI - Long-term hematopoietic engraftment after autologous peripheral blood progenitor cell transplantation in pediatric patients: effect of the CD34+ cell dose. AB - BACKGROUND AND OBJECTIVES: We analyzed the relationship between long-term hematopoietic recovery and the number of CD34+ cells infused in order to determine the optimal dose of CD34+ cells for rapid and stable engraftment. PATIENTS AND METHODS: Between November 1993 and December 1998, 96 consecutive autologous transplantations were performed in 92 pediatric patients with different malignancies. Peripheral blood progenitor cells (PBPC) were mobilized by G-CSF alone (12 microg/kg/day s.c., Neupogen((R)); Amgen, Thousand Oaks, Calif., USA) and collected using a Cobe Spectra blood cell separator (Cobe, Denver, Colo., USA) through a central venous catheter with double lumen. The CD34+ cell contents of apheresis products were assessed by means of flow cytometric analysis using an Epics Elite flow cytometer (Coulter, USA). RESULTS: The median number of CD34+ cells infused was 3.2 x 10(6)/kg (range 0.17-44.4). The median times for short-term engraftment (neutrophil count >0.5 x 10(9)/l and platelet count >20 x 10(9)/l) was 9 (range: 7-16) and 13 days (range: 7-91), respectively. The median times for long-term engraftment (platelet count >50 x 10(9)/l and >100 x 10(9)/l) was 21 (range: 10-249) and 45 days (range: 12-288). When the infused CD34+ cell dose was >/=5 x 10(6)/kg (median 7.99, range 5.01 44.4), there was a statistically significant increase in the rate of short- and long-term hematopoietic recovery compared to patients transplanted with a lower number of CD34+ cells (p < 0.0001). The earlier recovery in the high CD34+ cell group resulted in less transfusional support, fewer days on intravenous antibiotics and shorter hospitalization. CONCLUSIONS: This study confirms that G CSF-mobilized PBPC provide rapid short- and long-term hematopoietic engraftment in pediatric patients undergoing autologous transplantation if a CD34+ cell dose >/=5.0 x 10(6)/kg is infused. As this PBPC dose seems to have clinical and potentially economic implications, it should be considered the optimal dose for apheresis. PMID- 11111233 TI - Effects of leucocyte depletion on rheologic properties of human CPDA-1 blood. AB - BACKGROUND AND OBJECTIVES: Leucocyte depletion improves the quality of stored blood units. We have studied its role on blood viscosity. MATERIALS AND METHODS: Viscosity of CPDA-1 blood units was measured in a Couette viscometer at shear rates of 94.5 and 0.1 s(-1) prior to and following filtration with the Leukotrap((R)) A1 system on day 0 and after 21 days at +4 degrees C. RESULTS: On day 0, high but not low shear viscosity was significantly decreased. The red blood cell morphology was unaffected. On day 21, blood viscosity was increased similarly for unfiltered and filtered samples at both shear rates. The echinocytosis observed after storage correlated with the increase in viscosity. CONCLUSION: Leucocyte depletion is associated with a decrease in high shear viscosity. This effect is, however, completely lost after 21 days. PMID- 11111234 TI - Influence of methylene blue photoinactivation treatment on coagulation factors from fresh frozen plasma, cryoprecipitates and cryosupernatants. AB - OBJECTIVE: To study the influence of virus photoinactivation with methylene blue (MB) on the coagulation factors of fresh frozen plasma (FFP) and the corresponding cryoprecipitates and cryosupernatants derived from it. MATERIALS AND METHODS: The photoinactivation procedure of the German Red Cross (Springe) was applied using Biomat (Grifols, Spain). Twenty isogroup pools of three plasma units were made from 60 U of FFP. The pools were split into three bags. One of them was photoinactivated, and pre- and postinactivation samples (MB-plasma) were obtained. The second bag was treated in the same way, followed by the preparation of MB-cryoprecipitate and MB-cryosupernatant. The third bag was not photoinactivated, and was processed in the same way to obtain control cryoprecipitate and cryosupernatant. The prothrombin time and activated partial thromboplastin time were analysed, as well as fibrinogen, factors (F) II, V, VII, VIII, IX, XI and XIII, antithrombin III, von Willebrand (vW) F:RCo, vWF:Ag and the multimeric structure of vWF. RESULTS: In plasma, the proteins most sensitive to photoinactivation were fibrinogen, FV, FVIII, FIX and FXI (24, 32, 28, 23 and 27% loss, respectively). In the MB-cryoprecipitate, the losses were higher for FVIII (23%), moderate for fibrinogen, FXIII and vWF:RCo (18, 14 and 13%, respectively) and minimal (only 3%) for vWF:Ag. In MB-cryosupernatants, the losses were higher for FV (26%) and moderate for fibrinogen (16%), FIX (18%) and FXI (19%), as well as for FII and FXIII (15%). The multimeric structure of vWF was not modified in MB-plasma or in MB-cryoprecipitates. The supernatants (both MB treated as well as controls) showed an absence of multimers of very high and high molecular weight. CONCLUSIONS: The quantitative and qualitative conservation of coagulation factors achieved in MB-plasma-derived products suggest that they are useful for the global replacement of coagulation factors and for deficiencies in FV and FXI. In countries lacking the economic resources to obtain virally inactivated concentrates, MB-cryoprecipitates could be useful in von Willebrand's disease and fibrinogen and FXIII deficiencies. MB-cryosupernatants could be employed in thrombotic thrombocytopenic purpura, in the correction of total or partial deficiencies of prothrombin complex factors and in specific deficiencies of FV and FXI. PMID- 11111235 TI - Evaluation of solvent/detergent-treated plasma in patients with a prolonged prothrombin time. AB - OBJECTIVES: To compare the laboratory and clinical outcome of patients who received solvent/detergent-treated plasma (SDP) and fresh-frozen plasma (FFP). METHODS: A randomized, double-blinded study to assess the ability of SDP and FFP to reduce a prolonged prothrombin time (PT) to T in exon 3 of GYPA, is associated with the Or+ phenotype. We report here the first case of hemolytic disease of the newborn (HDN) caused by anti-Or, and expand the information on the nature of the Or determinant. MATERIALS AND METHODS: A woman, gravida 4, para 0, delivered a baby whose red blood cells (RBCs) were positive (2+) on the direct antiglobulin test (DAT). The mother's serum, an eluate made from the baby's RBCs and the RBCs of the baby's father were investigated. Exon 3 of GYPA, extracted from the father's genomic DNA, was amplified and sequenced. RESULTS: The mother's serum reacted at room temperature, 37 degrees C and on the indirect antiglobulin test with RBCs from the baby's father. The father's RBCs were M+N+S-s+Or+. The antibody in the mother's serum and in the baby's eluate was identified as anti Or. The serum did not react with the father's RBCs treated with trypsin (180,000 U/ml), but did react with his alpha-chymotrypsin-treated RBCs. Amplification and sequencing of DNA from the father revealed a single point mutation, 204C --> T, in GYPA exon 3. At birth, the baby had clinical symptoms of HDN and was transfused with 36 ml of packed RBCs and received phototherapy for eight days. At week 11, the baby's M+N+S+s+Or+ RBCs were negative on the DAT. CONCLUSION: This is the first case of HDN caused by anti-Or. The observed point mutation, 204C --> T, confirms that of a previous report and predicts a change of Arg (Or-) to Trp (Or+) at amino acid 31. PMID- 11111239 TI - Classification of standard alleles of the MN blood group system. AB - BACKGROUND AND OBJECTIVES: We had classified the M alleles of the MN blood group system into two subtypes, M(G) (standard M) and M(T), based on a G/T substitution in intron 1 of the glycophorin A (GPA) gene. This study provides further study on nucleotide sequences of M(G), M(T) and N alleles to profile the new allele, M(T). MATERIALS AND METHODS: M(G), M(T) and N alleles of the GPA gene were amplified using GPA gene-specific primers to avoid co-amplification of the genes of glycophorins B and E. Then the 5'-flanking region, exons 1-7 and introns 1-4 of the alleles were analyzed by DNA sequencing. RESULTS: There were 17 nucleotide substitutions and deletions between M(G) (standard M) and N alleles. Ten of the M(T) nucleotides were M(G)-type but the other 7 were N-type. M(T) allele also showed one base change and one deletion that were observed in neither the M(G) nor the N allele. Moreover, we found nucleotide substitutions within each allele, allowing further classification of the alleles. CONCLUSION: By the sequence data of M(G), M(T) and N alleles, the three alleles could be further classified into M101 and M102, M201 and M202, and N101 and N102, respectively. PMID- 11111240 TI - [Ophthalmic pathology at a Lebanese clinic: the example of Sidon]. AB - Ophthalmic consultations account for 15% of all activity at the medical department of the Caritas Center at Sidon. Over a period of 26 months, 1,000 ophthalmic diagnoses were made at this center, with some patients diagnosed with up to three conditions (for example, presbyopia, with cataracts and dryness of the eye). Two major symptoms, redness and pain, were the cause of most of the consultations. Ametropia (37.8%) and conjunctivitis (33.2%) were the most frequent diagnoses. Cheap reading glasses are sold at the center, to reduce the cost of optical treatment, and they are sometimes bought purely for the frame. We found no cases of xerophthalmia due to vitamin A deficiency or of florid trachoma, indicating that eye health is adequate in the region studied. The frequency of certain diseases causing blindness (cataracts, glaucoma, retinopathies and neuropathies) is certainly underestimated. Retinopathies were of diabetic origin and complicated in 70% of cases (stage IV or higher) because diabetologists do not systematically send patients to ophthalmologists. They often wait for loss of vision to become evident, by which time it is often too late for effective treatment to be given. For many reasons, the elderly do not consult as frequently as young patients. The feasibility of a mobile consultation service should be studied. It seems to be very important to initiate national mass campaigns for: - health education in the field of ophthalmology; - screening and treatment of diseases causing irreversible blindness; - recovery of old glasses for the use of poor patients. PMID- 11111241 TI - [Complications of illegal induced abortions at Bamako (Mali) between December 1997 and November 1998]. AB - We carried out a descriptive retrospective study over a period of one year, in the Gynecology and Obstetrics Referrals Department of Gabriel-Toure Hospital at Bamako. We analyzed 1,081 files of patients presenting at the department with complications of spontaneous or induced abortion. We identified and studied 189 cases of illegal induced abortions. The patients were young (mean age 21.8 years), of a low socioeconomic level and most (71.4%) had no living child. The gestational age of the fetus was less than 12 weeks for 19.5% of the women and between 13 and 16 weeks for 47.6%. The abortions were carried out by traditional practitioners (3.7%), general practitioners (9%), trainee nurses (10.5%) and state nurses and midwives (57.1%). In more than half the cases (71.4%), the abortion took place at the home of the practitioner. Several methods of abortion were used (e.g. curettage, uterine probes). In 44.4% of the cases, the woman refused to comment on the reasons for the abortion. The other women cited mostly academic reasons (20.63% of cases) and fear of their parents (13.22%). The main reasons for consultation were bleeding (51.3%), hyperthermia (35.4%), pain (9. 52%) and neurological problems (3.1%). Three types of complication were identified: bleeding (47%), infectious complication (33.3%) and drug poisoning (4.2%). The rate of maternal mortality was 10% and the management of the patients required surgical (from curettage to hysterectomy) and medical treatment, with a mean stay in hospital of 10 days. PMID- 11111242 TI - [Health risks study in a pottery environment in Morocco]. AB - Morocco is famous for its potteries, the largest of which are located at Rabat, Safi, Marrakech and Fes. This cross-sectional, descriptive epidemiological survey was carried out over an eight-month period, from January to August 1997. The study population consisted of 290 male workers from 36 workshops. The study involved a social and medical survey (a questionnaire and medical examination for all workers, with biological assessment for a representative sample of 95 craftsmen) and an analysis of the working conditions in which atmospheric pollution at the pottery was evaluated. Atmospheric concentrations of zinc, copper, iron, chrome and lead were determined. Only lead levels were found to be significantly high and were analyzed on three occasions. This study demonstrates poor working conditions and a lack of respect for the regulations concerning specific prevention measures and the health rules applicable to establishments where personnel are routinely exposed to the risk of lead poisoning. Various pathological conditions were observed, with the following prevalences: skeletal muscle 67.6%, dermatological 8.3%, digestive 58%, respiratory 28% and neurological 35.5%. Several nonspecific, often minor, clinical signs were recorded for most of the potters but the toxicological analysis confirmed lead contamination in 74% of the exposed subjects (plasma lead concentration, CPU, ALAU). The potteries of Morocco are not subject to any protection. Special technical and medical surveillance should be introduced and the laws concerning exposure to lead should be applied. PMID- 11111243 TI - [Peritoneal hydatidosis: a study of 25 cases in Morocco]. AB - We carried out a retrospective study of 25 cases of peritoneal hydatidosis. The incidence of this disease was 6.9%, the sex ratio of the patients was about 2/1 and the mean age of the patients was 31.8 years. Peritoneal echinococcal disease was most frequently secondary to the rupture or splitting of hydatid cysts in the liver (84% of cases) or, more rarely, in the spleen (4% of cases). The principal symptoms were unusual abdominal pain and abdominal masses. Ultrasound scan is the radiological method of choice for investigation and for assessing the number of hydatid cysts in the abdomen. It was used in 20 cases in this series and led to diagnosis of the disease in 95% of these cases. The sensitivity of CT scan for topographical diagnosis was about 90%. Serological tests were negative for the five remaining patients. Surgical management depends on the location and number of hydatid cysts and on the general state of the patient. Total cyst removal was performed in ten patients, pericystectomy in nine cases and marsupialization in six cases in which the cysts were located in the Douglas cul-de-sac. None of the patients was treated with albendazole. None of the patients died and the morbidity rate was 20%, due mainly to the hepatic location of the cysts. We observed one case of small bowel occlusion due to a missed daughter vesicle, two abscesses of the residual cavity, one case of pleurisy and one case of unexplained febrile syndrome. No recurrence was observed over a follow-up period of five years. PMID- 11111244 TI - [Study of the protein profile of the Adele tribe of Togo]. AB - Plasma proteins provide precise information about the physiological status of an individual. In this study, we compared the plasma protein profiles of 168 individuals from the Adele ethnic group, from an isolated rural area of Togo, with those of 159 individuals from an urban population from the capital, Lome. The Adele villages are located in the Atakora mountains. The subjects were volunteers, all apparently healthy and aged between 18 and 65 years. We separated serum proteins by electrophoresis and identified proteins specific for nutritional, inflammatory and immune status. The Adele significantly higher total serum protein concentrations than the urban individuals, with higher concentrations of a1 globulins (2.35 +/- 0.57 g/L versus 1.94 +/- 0.52 g/L) and g globulins (22.19 +/- 5.67 g/L versus 16.98 +/- 5.23 g/L) and lower concentrations of b globulins (6.83 +/- 1.56 g/L versus 7.34 +/- 1. 52 g/L). The Adele also had lower plasma concentrations of albumin (41.91 +/- 5.74 g/L versus 44.56 +/- 6.32 g/L), tranferrin (2.5 +/- 0.52 g/L versus 3.03 +/- 0.6 g/L), haptoglobin (0.57 +/ 0.59 g/L versus 1.32 +/- 0.89 g/L) and IgA (2.3 +/- 0.89 g/L versus 2.88 +/- 1.12 g/L) and higher plasma concentrations of orosomucoid (0.85 +/- 0.26 g/L versus 0.69 +/- 0.27 g/L); IgG (25.3 +/- 7.11 g/L versus 21. 79 +/- 6.5 g/L) and IgM (4.25 +/- 2.83 g/L versus 2.25 +/- 1.0 g/L). The data obtained for the Adele and urban populations were similar to those obtained for European populations except for IgM (higher in the Adele than in the urban and European populations), IgG and CRP (higher for the Adele and urban populations than for European populations). Nutritional status, as estimated by albumin and transferrin concentrations, was higher in the urban population of Lome than in the Adele population but the Adele population suffered no malnutrition problems. These results are consistent with those of a previous study, using apo A-I concentrations as an index of nutritional status. Apo A-I has also been shown to be a reliable indicator of nutritional status, as prealbumin concentration alone is sufficient for the early diagnosis of protein malnutrition. The very high concentrations of plasma CRP obtained indicate the presence of an inflammatory syndrome in the Adele and urban populations, as this protein is the first acute phase protein to be detected. However, the orosomucoid concentrations obtain-ed provide no evidence of significant inflammation. The high affinity of haptoglobin (Hp) for hemoglobin (Hb) results in the formation of soluble Hp-Hb complexes, reducing the value of Hp as a marker of the acute phase of inflammation. The frequency os sickle cell disease was higher in the Adele population than in the urban population (10-25% versus 2-6%). Hemoglobinopathies are correlated with haptoglobin concentration and thus plasma haptoglobin concentration was lower in the Adele population than in the urban population. The plasma concentrations of a1-antitrypsin in this study were similar to those reported for Europeans. The plasma concentration of protease inhibitors, such as a1-antitrypsin, increased as protease levels increased. These data confirm that the Adele and urban populations suffer no disease due to high levels of protease release into the bloodstream. They also show that a1-antitrypsin is of some value as an acute phase marker protein. The acute nature of the inflammatory syndrome (as assessed by CRP concentration) in the Adele and urban populations was confirmed by the hyperglobulinemia (high levels of production of IgM and IgG antibodies) observed in these populations. The Adele and Lome urban populations live in a tropical environment in which they are continuously in contact with infectious agents. This results in repeated stimulation of the immune system in both these populations. This study of plasma proteins in the Adele provides insight into the physiological conditions of this ethnic group, w PMID- 11111245 TI - [Lagoonal and coastal malaria at Cotonou: entomological findings]. AB - Nowadays, malaria control is planned according to the epidemiological context. Various aspects of malaria have been described in sub-Saharan Africa. We report here entomological data from the coastal area of Benin, West Africa, which has many lakes and lagoons. We carried out a longitudinal study in which we investigated the dynamics of populations of malaria vectors in various zones, the frequency of inoculation in these zones, the infestation rate of the Anopheles gambiae mosquitoes collected, the effect of urbanization on malaria transmission, the effects of inundation and of salinity at mosquito breeding sites. A total of 3, 342 identifications were made on a chromosomal basis. Two species of the Anopheles gambiae complex were detected in the coastal and lagoon areas of Benin: An. melas and An. gambiae ss. The density of the populations of these species was highly dependent on the level of urbanization. In traditional villages on the lagoons (such as Agbalilame, Djegbadji and Ketonou), the density of An. melas (86. 2%) was much higher than that in more urbanized areas (such as Ladji and Abomey-Calavi) (4.9%). We checked for chromosome polymorphism. We detected a 2Rn1 inversion in An. melas, similar to the 2Rn inversion found in mosquitoes in Gambia and Guinea-Bissau. The frequency of the n1 inversion and the density of An. melas populations were correlated and both seemed to depend on a single factor, salinity. The epidemiological situation with respect to malaria was very heterogeneous in the lagoon area of Benin. In the city of Cotonou, transmission was seasonal, sporozoite indices and the frequency of inoculation were high, in contrast to what would normally be expected in an urban area. In communities built on the beach, the level of transmission was markedly lower: about 5 infected bites per person per year versus 29 infected bites per person in the center of the city. In the traditional fishing villages, a paradoxical situation was observed in which the mosquitoes were very aggressive towards humans (4,502 bites per person per year) but the frequency of transmission was low (d = 0. 27%, CS+ = 0.57%). This was largely due to the high density in this area of An. melas, a poor malaria vector. If traditional villages become more urbanized, more freshwater breeding sites are created and the An. gambiae population increases, leading to an increase in malaria transmission. This is the reason for the higher level of malaria transmission at Ladji and Abomey-Calavi (h = 47 infected bites per person per year) than at Agbailame, Djegbadji and Ketonou (h = 12.1 infected bites per person per year) PMID- 11111246 TI - [One-year assessment of the cholera epidemic in Madagascar, from March 1999 to March 2000]. AB - The seventh cholera pandemic reached Madagascar in March 1999, 30 years after its appearance in East Africa. Two waves of infection were observed during the first year. The second wave was the stronger of the two. It occurred in the warm rainy season and spread over six provinces of the country. The incidence of cholera was from 0.1% to 2% and the hospital case fatality rate was between less than 2% and 6%, depending on the geographical area and the period. This outbreak has raised the awareness of communities and their leaders with respect to the environmental, practical and cultural factors that increase the risk of diseases transmitted by feces. PMID- 11111247 TI - [The fight against sexually transmitted diseases in Ivory Coast: what strategies can we use in the face of HIV/AIDS?]. AB - In the AIDS era, sexually transmitted diseases (STDs) have become a major health problem in developing countries, particularly in Africa. Delays in the diagnosis and treatment of such infections may result in complications, many of which primarily affect women. Epidemiological studies in Abidjan have shown that more than 10% of the pregnant women attending antenatal clinics present STDs potentially serious for their own health or that of their infants (gonorrhea, chlamydia infection, genital ulcers or active syphilis). There is evidence that STDs increase the transmission of HIV and that improving the syndromic management of STDs reduces the incidence of HIV infection. This provides a strong argument in favor of controlling STDs in areas of high HIV prevalence. In Ivory Coast, as in other African countries, a STD control program has been integrated into the AIDS control program since 1992, as recommended by the World Health Organization. During the first six years of the STD program, considerable progress was made in some areas, but not without difficulty. Simple syndrome-based decision trees have been adopted for the management of STDs in primary health care. Clinical studies have shown these therapeutic algorithms to be effective. At the same time, effective and affordable drugs for treating STDs were added to the list of essential drugs in Ivory Coast, after an international invitation to tender. The entire staff of the public health sector in Abidjan has been trained in syndromic STD management. Training is now being extended to other parts of Ivory Coast, including the private health sector and, in particular, private nurses. The surveillance of syndromic STDs, mainly genital ulcers in both sexes and urethral discharge in men, facilitates monitoring and evaluation of the STD program, following health care activities and adapting orders for drugs for treating STDs to real needs. In the near future, some parts of the STD program will be strengthened, particularly the management of sexual partners of STD patients and reduction of the cost of STD treatment for pregnant women. PMID- 11111248 TI - Managed care and public health. PMID- 11111249 TI - Not-so-strange bedfellows: public health and managed care. PMID- 11111251 TI - The Ohio Substance Abuse Monitoring Network: constructing and operating a statewide epidemiologic intelligence system. AB - Working with the Ohio Department of Alcohol and Drug Addiction Services (ODADAS) and researchers at the University of Akron, Wright State University's Center for Interventions, Treatment, and Addictions Research developed the Ohio Substance Abuse Monitoring (OSAM) Network to provide a statewide summary of substance abuse trends. Ten key informants across the state collect qualitative and statistical data on substance abuse trends in their regions and prepare biannual reports. The OSAM network has a rapid response capability through which key informants can investigate special issues related to substance abuse identified by ODADAS and provide policymakers with timely, statewide reports. Within 12 months after operations began, the key informants produced reports on drug abuse trends and rapid response issues for the state. These reports prepared policymakers to respond more effectively to prevention and substance abuse treatment needs. PMID- 11111250 TI - Latino child health: need for inclusion in the US national discourse. AB - The "rediscovery" of poverty, as echoed in concepts of social inequality, has contributed to the goal of eliminating racial/ethnic and social class disparities in the United States. This commentary focuses on what we know about the pressing health care needs and issues relevant to Latino children and families and how extant knowledge can be linked to priority policy recommendations to ensure the inclusion of Latino health issues in the national discourse. A systematic review of the literature on Latino children and of expert opinion revealed 4 evidence based themes focused on poverty: economic factors, family and community resources, health system factors, and pitfalls in Latino subgroup data collection. Consensus was found on 4 priority policy recommendations: (1) reduce poverty and increase access to health care coverage, (2) increase funding in targeted primary and preventive health care services, (3) provide funds needed to fully implement relevant health legislation, and (4) improve measurement and quality of data collection. If these recommendations are not instituted, the goals of Healthy People 2010 will not be achieved for the Latino population. PMID- 11111252 TI - After Cairo: women's reproductive and sexual health, rights, and empowerment. PMID- 11111253 TI - The great population debates: how relevant are they for the 21st century? AB - Two great debates--whether population growth is a problem and how to address the problem if one exists--dominated population policy discussions in the 20th century. The debate about whether pitted those who saw rapid population growth as a problem against those who believed the cries of alarm were false. The debate about how was conducted between advocates of the direct delivery of contraceptives through family planning programs and those who counseled a broader, more holistic approach. The debate about how was largely resolved by the 1994 International Conference on Population and Development at Cairo; the debate about whether remains unresolved. Environmentalists, ecologists, and physical scientists generally support the view that rapid population growth is harmful, but economists remain largely unconvinced. Contemporary declines in fertility and the end of the population crisis mentality of the mid- to late 20th century could, ironically, diminish public support for precisely those programs that have been responsible for the rapid fertility decline of the past 3 decades--programs that will be required to complete the "demographic transition" in those parts of the developing world where fertility remains very high. PMID- 11111254 TI - Population and reproductive health: where do we go next? PMID- 11111255 TI - Type of health insurance and the quality of primary care experience. AB - OBJECTIVES: This study examined the association between type of health insurance coverage and quality of primary care as measured by its distinguishing attributes -first contact, longitudinality, comprehensiveness, and coordination. METHODS: The household component of the 1996 Medical Expenditure Panel Survey was used for this study. The analysis primarily focused on subjects aged younger than 65 years who identified a usual source of care. Logistic regressions were used to examine the independent effects of insurance status on primary care attributes while individual sociodemographic characteristics were controlled for. RESULTS: The experience of primary care varies according to insurance status. The insured are able to obtain better primary care than the uninsured, and the privately insured are able to obtain better primary care than the publicly insured. Those insured through fee-for-service coverage experience better longitudinal care and less of a barrier to access than those insured through health maintenance organizations (HMOs). CONCLUSIONS: While expanding insurance coverage is important for establishing access to care, efforts are needed to enhance the quality of primary health care, particularly for the publicly insured. Policymakers should closely monitor the quality of primary care provided by HMOs. PMID- 11111256 TI - Parental employment and health insurance coverage among school-aged children with special health care needs. AB - OBJECTIVES: This study examined parental employment and health insurance coverage among children with and without special health care needs. Special needs were defined as conditions likely to require a high amount of parental care, potentially affecting parental employment. METHODS: Data from the 1994 National Health Interview Survey were analyzed for 21,415 children aged 5 to 17 years, including 1604 children with special needs. Logistic regression was used to estimate the effect of special needs on the odds of full-time parental employment and on the odds of a child's being uninsured, having Medicaid, or having employer sponsored insurance. RESULTS: Parents of children with special needs had less full-time employment. Their children had lower odds of having employer-sponsored insurance (adjusted odds ratio [OR] = 0.7) than other children. Children with special needs had greater odds of Medicaid coverage (adjusted OR = 2.3-5.1, depending on family income). Children with and without special needs were equally likely to be uninsured. CONCLUSIONS: Lower full-time employment among parents of children with special needs contributes to the children's being less likely to have employer-sponsored health insurance. Medicaid covers many children with special needs, but many others remain uninsured. PMID- 11111257 TI - The effect of capitated financing on mental health services for children and youth: the Colorado experience. AB - OBJECTIVES: This study tested 2 propositions concerning the effect of capitated financing on mental health services for Medicaid-eligible children and youth in Colorado. The first is that capitation reduces costs. The second is that shifting providers from fee-for-service to capitated financing will increase their efforts to prevent illness. METHODS: Interrupted time-series designs were applied to a naturally occurring quasi experiment occasioned by the state of Colorado's reorganization of mental health services financing. RESULTS: The cost of services was significantly lower in counties with capitated services compared with counties with fee-for-service financing. Findings also suggested that economic incentives may lead to greater efforts at secondary and tertiary prevention. CONCLUSIONS: Policymakers and the public can expect that capitation will reduce the costs of children's mental health services below those likely with fee-for service financing. Capitation per se, however, may not increase prevention as surely or swiftly as it lowers costs. PMID- 11111259 TI - Serving street-dwelling individuals with psychiatric disabilities: outcomes of a psychiatric rehabilitation clinical trial. AB - OBJECTIVES: This study tested a psychiatric rehabilitation approach for organizing and delivering services to street-dwelling persons with severe mental illness. METHODS: Street-dwelling persons with severe mental illness were randomly assigned to the experimental program (called Choices) or to standard treatment in New York City. We assessed study participants at baseline and at 6 month intervals over 24 months, using measures of service use, quality of life, health, mental health, and social psychological status. The average deviation from baseline summary statistic was employed to assess change. RESULTS: Compared with persons in standard treatment (n = 77), members of the experimental group (n = 91) were more likely to attend a day program (53% vs 27%), had less difficulty in meeting their basic needs, spent less time on the streets (55% vs 28% reduction), and spent more time in community housing (21% vs 9% increase). They showed greater improvement in life satisfaction and experienced a greater reduction in psychiatric symptoms. CONCLUSIONS: With an appropriate service model, it is possible to engage disaffiliated populations, expand their use of human services, and improve their housing conditions, quality of life, and mental health status. PMID- 11111258 TI - An international comparison of cancer survival: metropolitan Toronto, Ontario, and Honolulu, Hawaii. AB - OBJECTIVES: Comparisons of cancer survival in Canadian and US metropolitan areas have shown consistent Canadian advantages. This study tests a health insurance hypothesis by comparing cancer survival in Toronto, Ontario, and Honolulu, Hawaii. METHODS: Ontario and Hawaii registries provided a total of 9190 and 2895 cancer cases (breast and prostate, 1986-1990, followed until 1996). Socioeconomic data for each person's residence at the time of diagnosis were taken from population censuses. RESULTS: Socioeconomic status and cancer survival were directly associated in the US cohort, but not in the Canadian cohort. Compared with similar patients in Honolulu, residents of low-income areas in Toronto experienced 5-year survival advantages for breast and prostate cancer. In support of the health insurance hypothesis, between-country differences were smaller than those observed with other state samples and the Canadian advantage was larger among younger women. CONCLUSIONS: Hawaii seems to provide better cancer care than many other states, but patients in Toronto still enjoy a significant survival advantage. Although Hawaii's employer-mandated health insurance coverage seems an effective step toward providing equitable health care, even better care could be expected with a universally accessible, single-payer system. PMID- 11111260 TI - The effects of race/ethnicity, income, and family structure on adolescent risk behaviors. AB - OBJECTIVES: The study examined the unique and combined contributions of race/ethnicity, income, and family structure to adolescent cigarette smoking, alcohol use, involvement with violence, suicidal thoughts or attempts, and sexual intercourse. METHODS: Analyses were based on the National Longitudinal Study of Adolescent Health. A nationally representative sample of 7th to 12th graders participated in in-home interviews, as did a resident parent for 85.6% of the adolescent subjects. The final sample included 10,803 White, Black, and Hispanic 7th to 12th graders. RESULTS: White adolescents were more likely to smoke cigarettes, drink alcohol, and attempt suicide in the younger years than were Black and Hispanic youths. Black youths were more likely to have had sexual intercourse; both Black and Hispanic youths were more likely than White teens to engage in violence. Controlling for gender, race/ethnicity, income, and family structure together explained no more than 10% of the variance in each of the 5 risk behaviors among younger adolescents and no more than 7% among older youths. CONCLUSIONS: Findings suggest that when taken together, race/ethnicity, income, and family structure provide only limited understanding of adolescent risk behaviors. PMID- 11111261 TI - Suicide acts in 8 states: incidence and case fatality rates by demographics and method. AB - OBJECTIVES: This study examined incidence rates of medically identified suicide acts (self-inflicted injuries, either fatal or nonfatal) and case fatality rates by age, sex, race, and method used. METHODS: The authors analyzed data on 10,892 suicides and 57,439 attempted suicides among hospital-admitted individuals in 8 states, along with 6219 attempted suicides among individuals released from emergency departments in 2 states. RESULTS: The 8 states experienced a mean of 11 suicides and 119 attempted suicides per 100,000 residents each year. Groups with high suicide rates were men, the elderly, and Whites; groups with high attempted suicide rates were teenagers, young adults, women, and Blacks and Whites aged 25 to 44 years. Blacks aged 15 to 44 years evidenced high attempted suicide rates undocumented in previous studies. Poisoning and firearm were the most common methods used among those attempting suicide and those completing suicide acts, respectively. The most lethal method was firearm. CONCLUSIONS: The characteristics of suicides and attempted suicides differ dramatically. Method used is important in the lethality of the act. PMID- 11111262 TI - The effect of income inequality on the health of selected US demographic groups. AB - OBJECTIVES: This study assessed whether documented effects of income inequality on health are consistent across demographic subgroups of the US population. METHODS: Data from the National Health Interview Survey on White and Black non Hispanics were used. Logistic regression models were estimated with SUDAAN software. Perceived health was the outcome variable. RESULTS: The results of the multivariate analysis, in which individual family income and county-level poverty rates were included, were not consistent with existing research. In the presence of covariates, the conditional effects of inequality were restricted to Whites aged 18-44 years in the 2 highest income inequality quartiles and middle-aged Whites in counties with the highest level of income inequality. The health of Blacks of all ages, elderly Whites, and middle-aged Whites outside of the areas of highest inequality was unaffected when controls for individual characteristics and county-level poverty were in place. CONCLUSIONS: For the United States, the independent and direct contribution of income inequality to the determination of self-perceived health net of individual income and county income levels is restricted to certain demographic groups. PMID- 11111263 TI - Excess mortality among urban residents: how much, for whom, and why? AB - OBJECTIVES: The goals of this study were to estimate prospective mortality risks of city residence, specify how these risks vary by population subgroup, and explore possible explanations. METHODS: Data were derived from a probability sample of 3617 adults in the coterminous United States and analyzed via cross tabular and Cox proportional hazards methods. RESULTS: After adjustment for baseline sociodemographic and health variables, city residents had a mortality hazard rate ratio of 1.62 (95% confidence interval [CI] = 1.21, 2.18) relative to rural/small-town residents; suburbanites had an intermediate but not significantly elevated hazard rate ratio. This urban mortality risk was significant among men (hazard rate ratio: 2.25), especially non-Black men, but not among women. Among Black men, and to some degree Black women, suburban residence carried the greatest risk. All risks were most evident for those younger than 65 years. CONCLUSIONS: The mortality risk of city residence, at least among men, rivals that of major psychosocial risk factors such as race, low income, smoking, and social isolation and merits comparable attention in research and policy. PMID- 11111264 TI - Epidemiology of fetal alcohol syndrome in a South African community in the Western Cape Province. AB - OBJECTIVES: This study determined the characteristics of fetal alcohol syndrome in a South African community, and methodology was designed for the multidisciplinary study of fetal alcohol syndrome in developing societies. METHODS: An active case ascertainment, 2-tier methodology was used among 992 first-grade pupils. A case-control design, using measures of growth, development, dysmorphology, and maternal risk, delineated characteristics of children with fetal alcohol syndrome. RESULTS: A high rate of fetal alcohol syndrome was found in the schools--40.5 to 46.4 per 1000 children aged 5 to 9 years--and age specific community rates (ages 6-7) were 39.2 to 42.9. These rates are 18 to 141 times greater than in the United States. Rural residents had significantly more fetal alcohol syndrome. After control for ethnic variation, children with fetal alcohol syndrome had traits similar to those elsewhere: poor growth and development, congruent dysmorphology, and lower intellectual functioning. CONCLUSIONS: This study documented the highest fetal alcohol syndrome rate to date in an overall community population. Fetal alcohol syndrome initiatives that incorporate innovative sampling and active case ascertainment methods can be used to obtain timely and accurate data among developing populations. PMID- 11111265 TI - Working together? Organizational and market determinants of collaboration between public health and medical care providers. AB - OBJECTIVES: This study examines organizational characteristics and market conditions likely to influence collaborative relationships between public health agencies and community medical care providers. METHODS: Public health directors in 60 US counties were surveyed by telephone concerning their relationships with area community hospitals (n = 263) and community health centers (n = 85). Multivariate models were used to estimate the effects of organizational and market characteristics on collaboration. RESULTS: Collaboration was reported among 55% of the hospitals and 64% of the health centers. Certain forms of collaboration were more likely in markets characterized by higher HMO penetration and lower HMO competition. CONCLUSIONS: Targeted efforts to facilitate collaboration may be required in settings where institutional and market incentives are lacking. PMID- 11111266 TI - Attitudes and practices regarding varicella vaccination among physicians in Minnesota: implications for public health and provider education. AB - OBJECTIVES: This study sought to determine physicians' attitudes and practices regarding varicella vaccine. METHODS: A sample of Minnesota family physicians and pediatricians was surveyed in January 1997. RESULTS: Of 255 physicians surveyed, 108 (42%) reported routinely offering varicella vaccine. Physicians who perceived their professional organization's recommendations as "very important" were more likely to routinely offer varicella vaccine. Physicians who preferred natural disease over vaccination and those concerned about waning immunity were less likely to routinely offer vaccine. CONCLUSIONS: Recommendations of professional organizations have encouraged varicella vaccine use and may further enhance future use. Differences in pediatricians' and family physicians' attitudes and practices regarding this vaccine suggest that education of providers by specialty may be needed to increase acceptance of newly licensed vaccines. PMID- 11111267 TI - Length of stay and hospital readmission for persons with disabilities. AB - OBJECTIVES: Length of stay (LOS) and hospital readmission for persons receiving medical rehabilitation were examined. METHODS: A total of 96,473 patient records (1994-1998) were analyzed. Mean age of patients was 68.97 years; 61% were female and 83% were non-Hispanic White. RESULTS: A decrease in LOS of 6.07 days (SD = 3.23) and increase in hospital readmission were found across all impairment groups (P < .001). Readmission increases ranged from 6.7% for amputations to 1.4% for orthopedic conditions. LOS was longer (2.1 days) for readmitted patients (P < .01). Age was not a significant predictor of rehospitalization. CONCLUSIONS: Understanding variables associated with rehospitalization is important as prospective payment systems are introduced for postacute care. PMID- 11111268 TI - The relation of socioeconomic factors and racial/ethnic differences in US asthma mortality. AB - OBJECTIVES: This study described relations between socioeconomic factors and race/ethnicity as risk factors for asthma mortality. METHODS: A cross-sectional study was conducted of US mortality records from 1991 through 1996. RESULTS: Higher standardized mortality ratios were seen for Blacks vs Whites (3.34 vs 0.65), low vs high educational level (1.51 vs 0.69), and low vs high income (1.46 vs 0.71). Excess mortality for Blacks vs Whites was present in the highest and lowest quintiles of median county income and educational level. The disparity in asthma mortality rates according to median county income and education remained after control for race/ethnicity. CONCLUSIONS: Black race/ethnicity appears to be associated, independently from low income and low education, with an elevated risk for asthma mortality. PMID- 11111269 TI - Lessons learned from the tobacco industry's efforts to prevent the passage of a workplace smoking regulation. AB - OBJECTIVES: This study assessed the implementation of tobacco industry strategies to prevent a workplace smoking regulation. METHODS: Tobacco industry internal documents were identified; hearing transcripts for the affiliations, arguments, and positions regarding the regulation of testifiers were coded; and media coverage was analyzed. RESULTS: Tobacco industry strategies sought to increase business participation and economic discussions at public hearings and to promote unfavorable media coverage of the regulation. The percentage of business representatives opposing the regulation grew from 18% (5 to 28) to 57% (13 of 23) between the hearings. Economic arguments opposing the regulation rose from 25% (7 of 28) to 70% (16 of 23). Press coverage was neutral and did not increase during the period of the regulatory hearings. CONCLUSIONS: The tobacco industry was successful in implementing 2 of its 3 strategies but was not able to prevent passage of the comprehensive workplace regulation. PMID- 11111270 TI - Outreach developmental services to children of patients in treatment for substance abuse. AB - OBJECTIVES: This report describes a model for delivering developmental services to children of patients in treatment for substance abuse. METHODS: A multidisciplinary team provides developmental evaluations of children at a substance abuse treatment clinic. RESULTS: In 3 years of operation, 85% of 117 children completed individualized developmental evaluations. Cognitive limitations were diagnosed in 69%, speech and language impairments in 68%, emotional or behavioral problems in 16%, and medical problems in 83%. Follow-up information on children completing evaluation indicated that 72% of eligible children are receiving services as recommended. CONCLUSIONS: This high-risk population of children of substance-abusing parents can be effectively served by providing developmental services at a substance abuse treatment program. PMID- 11111271 TI - Trends in crime and the introduction of a needle exchange program. AB - OBJECTIVES: This study sought to determine whether introduction of a needle exchange program would be associated with increased crime rates. METHODS: Trends in arrests were compared in program and nonprogram areas before and after introduction of a needle exchange program in Baltimore. Trends were modeled and compared via Poisson regression. RESULTS: No significant differences in arrest trends emerged. Over the study period, increases in category-specific arrests in program and nonprogram areas, respectively, were as follows: drug possession, 17.7% and 13.4%; economically motivated offenses, 0.0% and 20.7%; resistance to police authority, 0.0% and 5.3%; and violent offenses, 7.2% and 8.0%. CONCLUSIONS: The lack of association of overall and type-specific arrest data with program implementation argues against the role of needle exchange programs in increasing crime rates. PMID- 11111272 TI - Giving means receiving: the protective effect of social capital on binge drinking on college campuses. AB - OBJECTIVES: We tested whether higher levels of social capital on college campuses protected against individual risks of binge drinking. METHODS: We used a nationally representative survey of 17,592 young people enrolled at 140 4-year colleges. Social capital was operationalized as individuals' average time committed to volunteering in the past month aggregated to the campus level. RESULTS: In multivariate analyses controlling for individual volunteering, sociodemographics, and several college characteristics, individuals from campuses with higher-than-average levels of social capital had a 26% lower individual risk for binge drinking (P < .001) than their peers at other schools. CONCLUSIONS: Social capital may play an important role in preventing binge drinking in the college setting. PMID- 11111273 TI - Self-reported health and prior health behaviors of newly admitted correctional inmates. AB - OBJECTIVES: This study obtained comprehensive health information from newly admitted correctional inmates. METHODS: Interviews were conducted with 1198 inmates on day 3 of their incarceration. RESULTS: Interviewers found a high prevalence of chronic medical and mental health issues, limited access to health care, high rates of infections and sexually transmitted diseases, substantial substance abuse, other unhealthy behaviors and violence, and a strong desire for help with health-related problems. CONCLUSIONS: The data document the need to apply the public health approach to correctional health care, including detection and early treatment of disease, education and prevention to facilitate health and behavior change, and continuity of care into the community. PMID- 11111275 TI - Evaluation of Medicaid managed care for children: access and satisfaction. PMID- 11111274 TI - Factors influencing a communitywide campaign to administer hepatitis A vaccine to men who have sex with men. AB - OBJECTIVES: A hepatitis A outbreak among men who have sex with men (MSM) led to a publicly funded vaccination campaign. We evaluated the MSM community's response. METHODS: A cohort of MSM from 5 community sites was surveyed. RESULTS: Thirty four (19%) of 178 potential vaccine candidates received the vaccine during the campaign. We found a linear relation between the number of exposures to campaign information and the likelihood of vaccination (P < .001). Vaccination was independently associated with awareness of the outbreak and the vaccine, having had sexual relations with men for 12 years or longer, having recently consulted a physician, and routinely reading a local gay newspaper. CONCLUSIONS: The difficult task of vaccinating MSM can be aided by repetitive promotional messages, especially via the gay media. PMID- 11111276 TI - The Social Security Administration "presumed living" search. PMID- 11111277 TI - A survey of men who have sex with men: mainland China. PMID- 11111278 TI - Combating dental disease. PMID- 11111279 TI - Improving dental health. PMID- 11111280 TI - Depression and the association of smoking and suicide. PMID- 11111281 TI - Health insurance problems are not going away. PMID- 11111282 TI - The financial performance of hospitals belonging to health networks and systems. AB - The U.S. health industry is experiencing substantial restructuring through ownership consolidation and development of new forms of interorganizational relationships. Using an established taxonomy of health networks and systems, this paper develops and tests four hypotheses related to hospital financial performance. Consistent with our predictions, we find that hospitals in health systems that had unified ownership generally had better financial performance than hospitals in contractually based health networks. Among health network hospitals, those belonging to highly centralized networks had better financial performance than those belonging to more decentralized networks. However, health system hospitals in moderately centralized systems performed better than those in highly centralized systems. Finally, hospitals in networks or systems with little differentiation or centralization experienced the poorest financial performance. These results are consistent with resource dependence, transaction cost economics, and institutional theories of organizational behavior, and provide a conceptual and empirical baseline for future research. PMID- 11111283 TI - The effect of changing state health policy on hospital uncompensated care. AB - This paper examines the effect of changing state policy, such as Medicaid eligibility, payment generosity, and HMO enrollment on provision of hospital uncompensated care. Using national data from the American Hospital Association for the period 1990 through 1995, we find that not-for-profit and public hospitals' uncompensated care levels respond positively to Medicaid payment generosity, although the magnitude of the effect is small. Not-for-profit hospitals respond negatively to Medicaid HMO penetration. Public and for-profit hospitals respond negatively to increases in Medicaid eligibility. Results suggest that public insurance payment generosity is an effective but inefficient policy instrument for influencing uncompensated care among not-for-profit hospitals. Further, in localities with high HMO penetration or high penetration of for-profit hospitals, it may be necessary to establish explicit payments for care of the uninsured. PMID- 11111284 TI - What determines hospital sponsorship of an HMO? AB - Using a strategic adaptation framework, this study evaluates the underlying institutional, market, organizational, and financial factors leading to hospital sponsorship of a health maintenance organization (HMO) insurance product. Analyzing hospitals in Metropolitan Statistical Areas (MSAs) in 1995 and 1996, the study found that a hospital is more likely to sponsor an HMO in markets that see a combined interaction effect of a large number of competing HMOs and high HMO penetration. HMO sponsorship also is more likely among hospitals with relatively low market share. Only in small MSA markets do hospitals with greater liquidity sponsor an HMO. Finally, hospitals that are affiliated with a multihospital system and under public ownership are more likely to sponsor HMOs. PMID- 11111285 TI - Ownership, competition, and the adoption of new technologies and cost-saving practices in a fixed-price environment. AB - Advances in medical technology have been implicated as the primary cause of rising health care expenditures. It is not yet known whether the increasing prevalence of managed care mechanisms, particularly capitation, will change substantially incentives for acquiring and using cost-increasing innovations. We examined the decisions of dialysis units (a set of providers that has faced capitation and real decreases in payment for several decades) with respect to use of cost-increasing technologies that enhance quality of care, cost-cutting practices that reduce quality of care, and amenities desired by patients that are unrelated to quality of care. We found that the dialysis payment system does not appear to have blocked access to a number of new, quality-enhancing technologies that were developed in the 1980s. However, facilities made adjustments along other valuable margins to facilitate adoption of these technologies; use of new technologies varied with numerous facility, regulatory, and case-mix characteristics including ownership, chain membership, size, market competition, and certificate of need programs. Interestingly, the trade-offs made by for profit and nonprofit facilities when faced with fixed prices appeared quite different. For-profits tended to deliver lower technical quality of care but more amenities, while nonprofits favored technical quality of care over amenities. Our findings may have implications for the response of other types of health care providers to capitation and increasing economic constraints. PMID- 11111286 TI - Explaining the decline in health insurance coverage among young men. AB - This article examines the experience of cohorts of young American men to see how and why their employer-provided health insurance coverage has changed over time. It explores changes in the structure of the labor market, changes in the cost of employer-provided health insurance, and changes in the composition of wages and benefits offered to employees. We find that increases in the cost of health insurance rather than changes in the structure of the labor market are the principal cause of the observed decline in employer-provided health insurance coverage across all cohorts. PMID- 11111287 TI - Does managed care mean more hassle for physicians? AB - Using the results of a 1995 nationally representative survey of physicians, this paper examines the relationship between exposure to managed care and resources expended by physicians on administrative and insurance matters. Our measures of managed care exposure are the degree to which a physician experiences a variety of managed care techniques (i.e., utilization review, capitation payment, restricted panels, gatekeepers, discounted fees, compensation links to utilization measures, profiling, protocols, and salary payment). Physicians report expending, on average, three hours per week on insurance-related matters and 4.8 hours per week on administration. Although managed care techniques affect administrative and insurance-related burdens, the direction of that effect varies according to the form that managed care exposure takes. With the exception of being salaried, none of our variables has an economically significant effect on physicians' administrative/insurance burdens, even at the outer-most edge of the 95% confidence interval. Overall, our findings contradict the widely held notion that managed care dramatically raises the administrative and insurance burden of physicians. PMID- 11111288 TI - Confirming insurance coverage in a telephone survey: evidence from the National Survey of America's families. AB - Until recently, most surveys of insurance coverage have classified people as uninsured if they have not been assigned some coverage in response to one of a series of questions about specific types of insurance. This "residual" approach to measuring uninsurance rates has not required respondents to either verify their insurance status or confirm that they are uninsured. Using the 1997 National Survey of America's Families, this paper examines the impact of a question confirming whether individuals for whom no insurance coverage is reported are, in fact, uninsured. The results of our analysis suggest that a confirmation question as part of a telephone-based survey works to lower estimates of the uninsured. PMID- 11111289 TI - [Consequences of intravascular contrast media on kidney function-- risk and prevention]. AB - The use of iodinated contrast media (CM) continues to be a common cause of hospital-acquired acute renal failure (ARF) and its development increases the in hospital mortality significantly. Alterations in renal hemodynamics and direct tubular toxicity by contrast media are the primary factors believed to be responsible for contrast media-associated nephrotoxicity. We review recent insights into the pathogenesis of this complication and summarize prophylactic strategies focussing on hydration, vasoactive pharmacological agents, alternative contrast media and "prophylactic hemodialysis". PMID- 11111290 TI - MRI evaluation of aseptic osteonecrosis in children over the course of hyperbaric oxygen therapy. AB - PURPOSE: The study aimed at MRI evaluation of aseptic osteonecrosis (AON) in children over the course of hyperbaric oxygen (HBO) therapy. MATERIAL AND METHODS: Retrospective analysis of 72 MRI studies in 20 children presenting with AON during chemotherapy. Two groups were differentiated: Gr. I (n = 8) was treated exclusively with relief of weightbearing structures, Gr. II (n = 12) was additionally treated with HBO therapy. The MRI examinations were evaluated by a point-score system (1-6 points) by two radiologists. RESULTS: Gr. II initially showed more severe findings (average score: 3.4) in comparison to Gr. I (average score: 2.65). During the follow-up time period the average scores rose to 3.2 score-points in Gr. I and 4.1 points in Gr. II. No statistically significant difference was evident between the two groups in the course of AON. CONCLUSION: The majority of chemotherapy associated AON which initially present with advanced findings show a progression in MRI over their further course. HBO therapy in addition to the relief of affected weightbearing structures statistically shows no significant improvement in MRI morphology during the course of the treatment. PMID- 11111291 TI - [Normal myelination in childhood brains using MRI--a meta analysis]. AB - PURPOSE: To establish age limits for the assessment of normal myelination of the brain on T1-weighted (T1w) and T2-weighted (T2w) images. METHOD: Comparison of previous publications (Barkovich et al. 1988, Grodd 1993, Hayakawa et al. 1990, Hittmair et al. 1994, Martin et al. 1988/1990/1991, Nakagawa et al. 1998, Staudt et al. 1993/1994, Stricker et al. 1990). RESULTS: Despite technical and methodological differences, these studies principally agreed on the timing of myelination for most regions of the brain. Thus, a common time-table could be established: At 1 month, myelin is visible on both T1w and T2w in the medulla oblongata, tegmentum pontis, cerebellar peduncles and vermis, quadrigeminal plate, decussation of superior cerebellar peduncles, thalamus, posterior limb of internal capsule, optic radiation, corona radiata. Thereafter, the myelin-typical signal in the different regions of the brain should be present at the following ages (M = months): anterior limb of internal capsule (2 M: T1w; 7 M: T2w), splenium of corpus callosum (4 M: T1w; 6 M: T2w), genu of corpus callosum (6 M: T1w; 8 M: T2w), centrum semiovale (2 M: T1w; 7 M: T2w). Branching of myelin into the gyri of the telencephalon (= arborization) appears at the latest at: occipital lobe (5 M: T1w; 12 M: T2w) and frontal lobe (7 M: T1w; 14 M: T2w). CONCLUSION: These extracted age limits can be used for a more reliable assessment of myelination than the time-tables from a single study. PMID- 11111292 TI - [Is perfusion MRI feasible in lesions with disrupted blood-brain barrier? Pitfalls and possible solutions]. AB - AIM: When using perfusion-weighted magnetic resonance imaging (MRI) in lesions with blood-brain barrier (BBB) disruption two methods are normally applied to eliminate the influence of contrast material uptake on T2* dynamics: injection of contrast agent before starting perfusion imaging (pre-injection) and simultaneous acquisition of T1- and T2*-dynamics using a dual echo (DE)-FLASH sequence. The purpose of this study was to examine both methods with regard to their reliability. MATERIAL AND METHODS: We performed four perfusion measurements in a patient with a primary cerebral lymphoma located in the corpus callosum: two measurements with a DE-FLASH sequence and two measurements with a gradient-echo echo-planar-imaging (GE-EPI) sequence, respectively. In both cases there was one measurement with and one without pre-injection of contrast material. RESULTS: Pre injection of contrast material reduced the influence of T1 and allowed us to monitor the transient signal drop when using the GE-EPI sequence. Analysis of the time-course of DE-FLASH, however, showed that there was still a T1 decrease even after pre-injection. After the first pass the T2*-dynamics showed a distinct long term T2*-decrease. DISCUSSION: Neither pre-injection nor DE-FLASH allow us to eliminate the effects of contrast agent uptake completely. Both methods, however, increase the reliability of perfusion MRI. Their efficacy depends on the extent of the BBB disruption. PMID- 11111293 TI - [The radial scar in contrast media-enhanced MR mammography]. AB - INTRODUCTION: In patients with the mammographic findings of a radial scar, contrast enhanced (CE) MR mammography was evaluated in a retrospective study. MATERIAL AND METHODS: In 24 women with radial opacities and black star configurations, CE MR mammography was performed. Examinations were done on a 1.5 T system using bilateral superficial coil (2D technique, T1-weighted FLASH sequence, TR 336 ms, TE 5 ms, FA 90 degrees). Findings in mammography and MR mammography were compared with the histological results. RESULTS: 15 radial scars (including 4 with additional ADH) and 9 carcinomas (6 in coexistence with a radial scar) presenting with a diameter of 3 mm to 13 mm were evaluated. There was no evidence of malignancy in MRI in 12 of 15 radial scars. In MR mammography 6 of the carcinomas fulfilled the criteria for malignant tumors. There were 3 borderline cases (scored 3 points) corresponding histologically to 1 radial scar, and to 2 carcinomas. Within the results found in MRI there were 2 false positives and 1 false negative. CONCLUSION: CE MR mammography is superior to other imaging modalities in the differentiation between radial scars and carcinomas. However, lesions suggestive of radial scars have to be removed surgically. PMID- 11111294 TI - [Comparison of bolus and infusion of the ultrasound contrast media levovist for color doppler ultrasound of renal arteries]. AB - PURPOSE: To investigate if the duration of Doppler enhancement of the ultrasound contrast agent Levovist can be prolonged by continuous infusion compared to bolus injections in patients. METHODS: 12 patients with suspected renal artery stenosis were included. Each patient received 1 or 2 bolus injections of 4 g each and a "fast" infusion of 3 ml/min (average dose: 7 g) of Levovist. In 8 patients an additional "slow" infusion of 1 ml/min (average dose 4.2 g) was given. The duration of "strong" Doppler signal enhancement and the visualization of vessels were compared. RESULTS: The duration of strong enhancement was substantially prolonged by the slow infusion (bolus 6:21 min, fast infusion: 8:57 min, slow infusion: 15:12 min, p < 0.001). The dose effectiveness (duration of strong enhancement per 1 g Levovist) was markedly improved from 0:57 min (bolus) to 3:36 min (slow infusion). Visualisation of the renal arteries was more complete with the slow infusion. CONCLUSION: Levovist infusions at 1 ml/min are superior to bolus injections in the assessment of renal arteries. They prolong the enhancement, make more efficient use of a given dose of contrast material and thus allow additional clinically relevant information to be obtained at reduced cost. PMID- 11111295 TI - [Vascularization of fibroadenomas: contrast harmonic imaging compared with contrast-enhanced power doppler]. AB - PURPOSE: To compare contrast material-enhanced harmonic power Doppler ultrasonography (CHI) with conventional contrast-enhanced power Doppler ultrasound (CPD) in depicting the vascularity of fibroadenomas. MATERIALS AND METHODS: Forty patients with fibroadenomas (aged 19-61 years) underwent conventional contrast-enhanced and harmonic power Doppler US. According to a standardized examination protocol, serial dynamic scans were obtained before and at 30, 90 and 150 seconds after injection of contrast agent. Video-taped scans were independently reviewed by three blinded readers with regard to parameter artifacts, degree of Doppler signal enhancement, demarcation of vessels and extent of vessel visualization. RESULTS: The number of intratumoral power Doppler signals depicted was similar with both techniques at 30-90 seconds after contrast administration; however, after 90 seconds, CPD depicted significantly more intratumoral signals than did CHI (p < 0.0001). CHI was superior to CPD with regard to "blooming" or motion-related artifacts (p < 0.0001). CONCLUSION: CHI balances the increased susceptibility of CPD to interference from clutter artifacts and thus allows the depiction of vessels that may have been obscured on CPD at similar system settings. PMID- 11111296 TI - [First results after implantation of the new balloon-expanded Bridge-Stent into the iliac artery]. AB - PURPOSE: To determine the success and patency rates of the new, balloon expandable Bridge-stents for treatment of occlusions and stenoses of iliac arteries. MATERIAL AND METHODS: Between April and October 1999, 21 patients suffering from peripheral arterial occlusive disease were treated (22 lesions). In 20 cases high-grade stenoses and in two cases complete occlusions were treated. 23 balloon-expandable Bridge-stents were implanted in the common iliac (n = 16) and the external iliac (n = 6) artery after percutaneous transluminal angioplasty. RESULTS: All interventions were performed with technical success. The mean ankle-brachial index (ABI) increased from 0.7 +/- 0.19 before to 0.86 +/ 0.16 at 24 hours after the procedure (Rutherford +3 [n = 18] and +2 [n = 3]); the ABI measured after 6 months was 0.89 +/- 0.16. The primary cumulative patency rate amounted to 91% (20/22) at 6 months. In two cases restenosis of 75% were observed, requiring repeated angioplasty. CONCLUSION: The new balloon-expandable Bridge-stents achieve very good initial technical success and promising short term patency in the percutaneous treatment of iliac occlusive disease. PMID- 11111297 TI - [Changes in the region and the proximal aneurysm neck after endovascular repair of infrarenal aortic aneurysms]. AB - PURPOSE: To analyze short and mid-term changes of the proximal aneurysm neck and to evaluate renal function after endovascular repair of infrarenal abdominal aortic aneurysms. MATERIALS AND METHODS: 24 of 29 patients, who were treated endoluminally with the bifurcated Vanguard Stent graft between 1997 and 1999 underwent spiral CT follow-up after 1, 3, 6 and every 12 months. Mean follow-up period was 10.5 months (3-32 months). Follow-up included spiral CT scanning. The aortic diameter as well as cross-sectional area were measured. Stent graft position and renal perfusion were checked by spiral CT. Serum creatinine levels were determined preoperatively and during follow-up. RESULTS: Supra- and infrarenal aortic sizes remained stable. No correlation to the distance between the stent-graft and the renal arteries was observed. Caudal migration of the stent-graft with an average of 6.5 (3-15) mm occurred in 13 patients. Cranial migration was observed in 3 patients. Uncovered stent wires partially overlapped at least one renal artery in 18 patients. No renal infarction occurred. No significant increase of the serum creatinine level was found. CONCLUSION: The risk of proximal endoleakage due to post-interventional dilatation of the aorta abdominalis seems to be low. Insufficient stent graft fixation requires a wide distance between the proximal end of the stent graft and the aneurysm. In the mid term uncovered stent wires overlapping the renal arteries had no relevant influence on renal function. PMID- 11111298 TI - [MR-tomographic clarification of aortic isthmus stenosis]. PMID- 11111299 TI - [Symptomatic spinal hemangioma as a cause of spinal root syndrome]. PMID- 11111300 TI - [Diagnosis of tumor-simulating intra-ossicle arachnoid diverticulum using liquid sensitive sequential MRI]. PMID- 11111301 TI - [Talocrural joint imaging: an unremarkable abnormality with significant accompanying symptoms]. PMID- 11111302 TI - [Palliative therapy in an arterial bleeding ureteral tumor-- transfemoral tumor embolization]. PMID- 11111303 TI - [Two unusual cases of arterial and venous collateral circulation the the ligamentum teres hepatis]. PMID- 11111304 TI - [Women's imaging: a challenge for radiology in Austria]. PMID- 11111305 TI - [New drugs for the treatment of influenza]. PMID- 11111306 TI - [The economic and labor impact of no staff replacement in primary care. Why and how we should measure it]. AB - OBJECTIVES: To quantify the effect of not replacing staff in both job (mean overloads in time -MOT- and in users -MOU-) and economic terms. To calculate the effective real time worked (ERTW). DESIGN: Cross-sectional descriptive study. SETTING: Primary care, San Fernando Health Centre, Mostoles, Madrid. PARTICIPANTS: The 41 people in the team. MEASUREMENTS AND MAIN RESULTS: Records of time off in 1999, with economic and labour costs. Comparison of the mean patients per shift -MOU- with the gold standard: Royal Decree 1575/1993. 72.63% (14,227,538 pesetas) of the cost of time off (19,589,226 pesetas) was assumed by the team (negative cost, economic impact). The MOT per health professional was 6.6 weeks per year for doctors, 6.5 for nurses and 4.2 for paediatricians. The ERTW per professional was 44.1 weeks a year for paediatricians, 39.4 for doctors and 39.1 for nurses. The MOU on a professional being absent raises the lists to 2471 (medicine) and 2179 (paediatrics) on the morning shift; and to 2296 (medicine) and 1662 (paediatrics) on the afternoon one. CONCLUSIONS: The team assumes most of the economic burden, with no counter-benefit. The MOT is high and exceeds the number of weeks per year of formally allowed time off. In practice, therefore, each professional covers for their full allowed time off. The ERTW is much below the 52 weeks on which the team was assessed. RD 1575/1993 is frequently broken by current MOU. It is suggested that coordinators use these indicators in negotiating the Contract of Care Management and the Portfolio of Services. PMID- 11111307 TI - [What do family medicine residents expect of their tutors? A qualitative approach]. AB - OBJECTIVE: To find the view of family and community medicine (FCM) residents on the kinds of relationship between tutors and residents and their training. DESIGN: The methodology employed was qualitative; and the technique chosen, that of focus groups. SETTING AND PARTICIPANTS: FCM residents belonging to the teaching units in Murcia and Cartagena. INTERVENTION: Five groups were organised, two second-year resident groups, two third-year ones, and one first-year one. At the start of each group the objectives and norms of the meeting were explained. Residents were then asked four questions about their relationship with their tutor and the training received during their residency. The information arising was classified as a function of the time of debate, the number of residents expressing the same view and the intensity of their emotional involvement on expressing their views. RESULTS: The tutor's confidence in the resident's work was what residents valued most highly, along with the exchange of views as equals. The participants stated that they wanted to take part in planning their training and that they needed to participate in the clinical and organisational decisions taken in the tutor's clinic. CONCLUSIONS: Trust between tutor and resident is essential. Communication between the two must be on an equal basis. Training procedures are valued higher than training content. Residents want to intervene in planning their training. Tutors are models for training. PMID- 11111308 TI - [The quality of the drug treatment in hyperlipemia patients from 4 health areas. The VICAF Group]. AB - OBJECTIVE: To calculate how suitable the lipid-lowering treatment prescribed for pensioners in primary care clinics in four health areas is. DESIGN: Cross sectional descriptive study of quality of pharmacological treatment. SETTING: Four primary care health districts, INSALUD, Madrid. PARTICIPANTS: 1125 patients registered with 49 doctors, chosen at random on the basis of three strata defined by the value of the lipid-lowering drug indicator of prescription. For a year, each doctor filled in a protocol of variables for each pensioner to whom he/she prescribed a lipid-lowering drug. MEASUREMENTS AND MAIN RESULTS: An automated algorithm was designed to evaluate the suitability of the drugs treatment for each patient, according to scientific criteria including: cholesterol levels, LDL, age, and risk factors. Quality of prescription was finally measured for 1009 patients. The indication of the treatment was due to primary prevention in 65% of cases. 32% of patients were correctly treated. If LDL compliance was not demanded, the suitability figure rose to 77%. Drug treatment was more suitable when the doctor him/herself administered it (as against another doctor or a specialist; p = 0.001) or when the patient was on the list of the prescribing doctor (p < 0.0001). Proper indication was lower in patients over 74 (p < 0.0001). CONCLUSIONS: The quality of lipid-lowering drug prescription for pensioners in primary care clinics in four health districts, as a function of the criteria defined above, could be improved. LDL is the factor which most affects the procedure. PMID- 11111309 TI - [The community activity of a hospital geriatrics service: a practical example of coordination between primary and specialized care]. AB - OBJECTIVES: To describe the management of the geriatric hospital home assessment team to support at primary care in the need of health of geriatric patients. DESIGN: Prospective observational study. SETTING: 5-North health district from Madrid. PARTICIPANTS AND METHODS: All patients evaluated at home and the coordination activities between primary care and hospital geriatric service, developed by the geriatric assessment team between january 1997 at december 1999. Inquiry to primary care physicians about the usefulness of geriatric hospital home assessment team. MAIN RESULTS: 524 patients was assisted at home by the geriatric assessment team to request primary care (58.4% at all patients attended), 83.24 +/- 7.21 years old (68.1% females), with pluripathology (4.02 +/ 1.86 diagnostics) and polypharmacy (4.95 +/- 2.8), moderate-severe functional disability (Barthel Index < 60 at 65% and Red Cross functional disability > 2 at 74.4%) and frequently cognitive impairment (41.2% with Pfeiffer > 5). The main reasons of primary care to consult was functional impairment (28.4%), medical process assessment (16.2%), geriatric assessment (13.2%), skin ulcers (13.2%) and behavioral disorders (10.8%). 865 telephone call and 178 meeting in health center of primary care for consultation or medical management or for coordinating medical management was attended. 62% of primary care physician inquired to answer back. The belief of 100% answers was about de usefulness of the geriatric assessment team activity, mainly because their specialization on geriatric care (76.7%) and their responsibility of the hospital resources (65.1%). CONCLUSIONS: The activity of geriatric hospital home assessment team is useful as support of primary care on assessment and management geriatric patients in the community and coordinate hospital and primary care to resolve health problems in this population. PMID- 11111310 TI - [The hospital discharge report in primary care (1). An analysis of its utility]. AB - OBJECTIVES: To calculate the number of hospital discharge reports (HDR) received in primary care for determined kinds of patients suffering chronic pathologies who need on-going care. To evaluate whether the reports' contents are of use to the family doctor's monitoring of the patient. DESIGN: Descriptive study. SETTING: Four reformed health districts that share the same referral hospital in the city of Mataro (Barcelona). PATIENTS AND OTHER PARTICIPANTS: 124 patients were discharged in 1996 for pathologies requiring ongoing care in primary care (diabetes, hypertension, chronic obstructive pulmonary disease, cerebral vascular accident, femur fracture and neoplasms). INTERVENTIONS: Systematic comparison of various sections of the HDR with the information in the primary care clinical notes (PCCN). MEASUREMENTS AND MAIN RESULTS: In 12.9% (95% CI, 8.1-18.8%) of the PCCNs examined, there was no record of the HDR. In 76.6% (69.1-84.1%) of the patients studied, the HDR contributed new information on some of the contents reviewed ("useful" HDR). In the remaining 22.6% (15.2-30.0%) the HDR contributed no new information and left out necessary follow-up and monitoring information ("useless" HDR). Nevertheless, in as many as 89.5% (84.1-94.9%) of the patients, the PCCNs recorded some clinically important piece of information or intervention that did not figure in the HDR. This occurred more often in questions of prevention and quality of life (56.5%), special treatment (42.3% of those who needed it), anamnesis (40.3%) and follow-up (31.5%). CONCLUSIONS: Despite the potential importance of HDRs for on-going care of patients discharged from hospital for chronic complaints, and although a high percentage of HDRs reach the general practitioner, their lack of content negates their usefulness in 22.6% or more of cases. PMID- 11111311 TI - [Sexual dysfunctions induced by serotonin reuptake inhibitors]. AB - OBJECTIVE: To assess the incidence of serotonin reuptake inhibitor (SRI) antidepressant-induced sexual dysfunction (SD) and to compare the sexual side effects of SRI. DESIGN: Naturalistic, prospective, observational study. SETTING: Two urban health centers. PATIENTS: 235 outpatients (164 women, 71 males) who began treatment with some of the following SRI: fluoxetine, sertraline, paroxetine, citalopram and venlafaxine, who had engaged in regular sexual practices with stable partner, who were suffering from different mental disorders who were being treated with SRI. The assignment to each group was according to clinical criteria. INTERVENTIONS: Patients completed questionnaires that allowed reporting of both SD induced by the illness and the treatment, evaluating changes in libido, arousal, and orgasm. The patients were observed over 6 months of treatment. RESULTS: 147 patients (62.6%) reported one or more SD related to SRI treatment. There were differences in the incidence between the different SRI: 39% with fluoxetine, 75.5% with paroxetine, 78.8% with sertraline, 28.9% with citalopram and 80% with venlafaxine. In 78.2% of patients the SD showed no improvement by the end of this period. In a predictive logistical regression model of the presence of SD induced by the SRI, the female category and the presence of previous sexual problems were favourable predictors and the treatment with paroxetine, sertraline or venlafaxine were increased the risk of SD. CONCLUSIONS: SD is one of the most frequent and persistent SRI adverse effect. We recommended to inquiry about SD in patients who were treated with SRI. Significant differences were found in the occurrence of SD between the different SRI. Such data would be particularly valuable to physicians when choosing a specific antidepressant from this therapeutic group. PMID- 11111312 TI - [An alternative method for the diagnosis of an anterosuperior hemiblock]. AB - OBJECTIVE: To develop and validate a rapid method for diagnosing superior anterior hemiblock, comparing it with the system seen as standard. DESIGN: Comparison study of two diagnostic tests. SETTING: Urban health centre. PARTICIPANTS: Adult population with prior diagnosis of superior anterior hemiblock included in a computer file of clinical records. MEASUREMENTS AND RESULTS: The presence or otherwise of superior anterior hemiblock was found by applying the standard criteria for electrocardiograms contained in the clinical records of the participating patients. The proposed test was then applied by finding the precise cardiac axis and the confirmation or rejection of the diagnosis of superior anterior hemiblock. Out of the 121 histories analysed, 40 had superior anterior hemiblock according to the standard criteria, and 81 did not. The system proposed detected 38 of the 40 hemiblock cases, 95% sensitivity, and rejected as normal 76 of the 81 electrocardiograms without superior anterior hemiblock according to the standard pattern, specificity of 93.8%. CONCLUSIONS: Superior anterior hemiblock can be diagnosed by an alternative method which was quicker than the standard pattern, so markedly simplifying the analysis of this feature of the electrocardiogram. PMID- 11111313 TI - [The prevalence of caries and associated factors in children 2-5 years old from the Almanjayar and Cartuja Health Centers of the capital Granada]. AB - OBJECTIVE: To estimate caries prevalence and associated factors in children 2-5 years old in a deprived community. DESIGN: Transversal, cross-sectional study. SETTING: Paediatric services. Health centres of Almanjayar and Cartuja in Granada (Spain). PATIENTS: 173 children attending to a pediatric revision. MEASUREMENTS: Children's odontological examination and revision of clinical records followed by structured interview with the mother or tutor. Target variables were caries, sociodemographic factors, nutritional habits, oral hygiene, medical antecedents, familiar experience of caries and use of dentistry services. RESULTS: Total prevalence of caries was 37%, but 29% among the majority population and 58% in the gipsy group. Statistically significant associated factors with caries were: increasing age (OR = 2.0, 95% CI = 1.2-3.2), father unemployment (OR = 3.1, 95% CI = 1.3-9.9), high consumption of sweets (OR = 3.3, 95% CI = 1.1-8.5), deficient oral hygiene (OR = 9.3, 95% CI = 3.4-24.7), mother's consultation for toothache or tooth extraction (OR = 2.9, 95% CI = 1.1-7.9) and not attendance to dentistry services due to high costs or fear (OR = 4.3, 95% CI = 1.5-12.4). CONCLUSIONS: The prevalence of caries in the gipsy population is very high, and it is probably associated with factors previously reported but not yet controlled. There is a need to initiate therapeutic and preventive measures in this community, and to detect barriers and facilitate the use of public dentistry services. PMID- 11111314 TI - [Care for the menopausal woman: an objective to be developed from primary care]. PMID- 11111315 TI - [Fibromyalgia: a Cinderella diagnosis]. PMID- 11111317 TI - [Local corticoid infiltrations at a health center]. PMID- 11111316 TI - [The prevention of breast cancer in primary care]. PMID- 11111318 TI - [Possible toxicoderma secondary to influenza vaccination]. PMID- 11111319 TI - [The referral of patients from primary care to a rheumatology consultation in a regional hospital]. PMID- 11111320 TI - [Constipation and laxative consumption in the elderly]. PMID- 11111321 TI - [The assignment of population to nursing]. PMID- 11111322 TI - Inhibitory effect of lactoferrin on the adhesion of Prevotella nigrescens ATCC 25261 to hydroxyapatite. AB - Prevotella nigrescens ATCC 25261 (P. nigrescens) cells adhere well to hydroxyapatite treated with citrate (CHA), but the attachment is drastically inhibited by lactoferrin (LF). To determine the nature of the iron-free LF responsible for inhibiting P. nigrescens cell attachment, this study tested the duration and frequency of LF treatment of CHA and the effects of divalent and trivalent ferric ions. The inhibitory effect on the attachment of P. nigrescens was somewhat higher with bovine LF than human LF. Apo LF effectively inhibited P. nigrescens cell attachment to CHA, and almost abolished attachment at a concentration of 0.4 mg/ml. Fe3+ saturated LF was unable to inhibit attachment, whereas Fe2+ showed a slight effect under the same conditions. The LF absorbed rapidly to CHA in less than 10 min. With a lower concentration (0.1 mg/ml) of LF, only three treatments of CHA were required for the maximum inhibition of P. nigrescens cell attachment. The quantity of LF adsorbed to the hydroxyapatite (HA) and to P. nigrescens cells was determined by use of [3H]-LF. Approximately 25 micrograms of LF protein adsorbed to 5 mg of HA at saturation, and approximately 0.25 microgram of LF did so to the 6 x 10(8) cells. PMID- 11111323 TI - A case of localized ridge augmentation from using a titanium membrane: a pilot study. AB - The aim of this investigation was to histologically compare the effects of new titanium membranes with those of proven expanded polytetrafluoroethylene (ePTFE) membranes on alveolar ridge augmentation. The study was carried out using a canine mandible where the right and left premolar teeth were previously extracted. At the second month of healing following extraction, a total of seven similar defects were created on both sides of the mandible. Three defects received the titanium membrane, one of which was in the right side of the mandible. Two defects, one on each side, received the ePTFE membrane. The remaining two defects, both on the right side, served as control. Fixation and stabilization of the membranes were accomplished by using the Frios Augmentation System. After three months of healing, the animal was sacrificed, block sections were taken and processed for histological examination according to the cutting and grinding technique. The results revealed that the ePTFE and titanium-treated defects, and the controls, showed complete bone fill with the exception that there was a more pronounced and thicker connective tissue formation in titanium treated sites as compared to the ePTFE treated and control sites. PMID- 11111324 TI - Topography of periodontally involved human root surfaces after different chemical treatment modalities: an in vitro scanning electron microscopic study. AB - The significance of chemical and conservative treatments of cemental tissue proximal to periodontal pockets has been pointed out in recent years. This in vitro scanning electron microscopy (SEM) study aimed to investigate the surface effects of topical applications of 0.1% cetylpyridinium chloride (CPC) and 2% sodium lauryl sulfate (SLS) and polishing on the periodontally involved root surfaces of human teeth. Ten single-rooted teeth from 8 patients with advanced adult periodontitis were included. Following extraction, any calculus was removed with extreme care to preserve as much cementum as possible. Eighty root specimens were prepared. Fresh solutions of CPC and SLS were applied for 1, 3 and 5 minutes each to 10 segments of root cementum. A total of 20 segments formed the polished (P) and control (C) groups, respectively. The results showed that the surfaces treated with CPC or SLS differed considerably from polished and control specimens. Depending on time, the surface coating was partly or wholly removed, leaving a nodular cementum structure, uncovering a fibrillar collagen substrate and the openings of dentinal tubules. Scarce debris was present on both control and polished surfaces, whereas bacteria were observed only on the control specimens. In view of these results, further definitive in vitro and in vivo research must be done to determine the advantages of chemical treatment and its effect on periodontal regeneration. PMID- 11111325 TI - Persistence of hepatitis B surface antibody levels after vaccination with a recombinant hepatitis B vaccine: a 3-year follow-up study. AB - We conducted a 3-year follow-up study of 2,008 individuals for the persistence of antibody levels after vaccination with a recombinant hepatitis B vaccine. At 1 month after vaccination had been completed 96.3% of subjects had acquired protective HB surface (HBs) antibody titers on passive hemagglutination assay (PHA) of 2, and 3 years after completion of the vaccination 63.1% had acquired similar titers. The titer decreased below this level after 3 years in 34.5% of subjects who had initially acquired protective antibody titers. The mean acquired HBs antibody titer on PHA was 2 at 1 month and 2 at 3 years after completion of the vaccination. The regression line for these changes was expressed as logY = 1.800-0.24X by the least squares method. The antibody level was estimated to decrease to 2 at 37 months and to seronegativity at 75 months. On the basis of the relationship between PHA antibody titers and the period of their persistence, the persistence of antibody levels after vaccination with a recombinant vaccine can be estimated from the acquired antibody titer determined 1 month after completion of the vaccination, as with plasma-derived vaccines. PMID- 11111326 TI - Relationship between gingival health and dental caries in children aged 7-12 years. AB - The purpose of this study was to investigate the relationship between gingival health and dental caries in elementary school children in Japan. The subjects were 474 children aged 7 to 12 years who attended dental check-ups at an elementary school. The Oral Rating Index for Children, which consists of five categories (+2, +1, 0, -1, -2), was used to rate the findings of the gingival health examination. The dental examination was performed using the WHO caries diagnostic criteria for DMFT. Children were divided into three groups: a healthier group (H-group) made up of those scoring +2 (excellent) or +1 (good), an equivocal group (E-group) made up of those scoring 0, and a gingival less healthy group (L-group) made up of those scoring -2 (very poor) or -1 (poor). Overall percentages for the H-group, E-group and L-group were 48.3%, 21.5% and 30.2%, respectively. The number in the L-group increased with increasing age. The mean scores of the DT and DMFT in the H-group were significantly lower than those in the L-group (p < 0.01 and p < 0.05, respectively). The results suggest that oral hygiene instruction should be given to children in order to motivate self care, not only to avoid dental caries but also to prevent gingivitis. PMID- 11111327 TI - Effects of saiko-ka-ryukotsu-borei-to on spontaneous locomotor activity in mice. Oriental Medicine Research Group. AB - The effects of the Japanese Kampo (herbal) medicine, Saiko-ka-ryukotsu-borei-to, on spontaneous locomotor activity were studied in mice. Saiko-ka-ryukotsu-borei to (60 mg, 150 mg and 300 mg/kg/day) was administered for 14 consecutive days in the drinking water and spontaneous locomotor activity was measured for 60 min by a photocell ambulometer. Saiko-ka-ryukotsu-borei-to (60 mg/kg/day) significantly increased the total activity count on the 11th day after the start of administration when compared to vehicle control, whereas failed to significantly affect the activity on the 2nd, 5th, 8th and 14th days. A similar significant increase was also found with a higher dose (150 mg/kg/day) on the 8th day after the start of administration. However, the highest dose (300 mg/kg/day) did not significantly affect locomotor activity throughout the experimental period. We have previously reported that Saiko-ka-ryukotsu-borei-to, at a dose of 60 mg/kg/day, enhances escape attempts assessed by water-wheel rotations in a mouse model of despair, particularly on the 8th, 11th and 14th days after the start of chronic treatment. However, at higher doses (150 and 300 mg/kg/day), Saiko-ka ryukotsu-borei-to decreases the escaped attempts on the 5th and 8th days after the treatment. It is therefore concluded that the previously reported changes in escape attempts of mice are not associated with the changes in their spontaneous locomotor activity. PMID- 11111328 TI - Treatment of ligature-induced peri-implantitis defects by regenerative procedures. Part II: A histometric study in dogs. AB - The purpose of this study was to evaluate, by histometric analysis, re osseointegration following treatment of ligature-induced peri-implantitis in dogs. Five dogs were used in this study. Their mandibular premolars (P2, P3 and P4) were first removed. After 3 additional months of healing, two titanium implants were placed on each side of the mandible. After 3 months, the abutment connection was performed and experimental peri-implantitis was induced by placing cotton ligatures in a submarginal position. Ligatures and abutments were removed after one month and the peri-implant bone defects were randomly assigned to one of the treatments: debridement, debridement plus guided-bone regeneration, debridement plus mineralized-bone graft, and debridement plus guided-bone regeneration associated with mineralized-bone graft. Five months post-treatment, the degree of bone contact with the implant surface and the bone area within the threads were measured in 12 threads, the 6 most coronal at each side of each implant. One-way analysis of variance did not reveal statistically significant differences between the treatment modalities (p > 0.05). Within the limits of the present study, it can be concluded that there is a limited possibility of re osseointegration around implant surfaces previously exposed by ligature-induced peri-implantitis. PMID- 11111329 TI - Observation of the abluminal surface of the atrioventricular endothelium which undergoes transition into cardiac cushion mesenchymal cells in the chick embryo. AB - In the developing chick heart, endothelial cells in the atrioventricular canal (AV) undergo a series of morphological changes and transform into cushion mesenchymal cells. In the present scanning electron microscopic study, we examined the abluminal surface features of the AV endothelium through an artificial window in the myocardial wall. The AV endothelial cell at stages 12 or earlier had a smooth, flattened basal surface with only a few blebs. In the successive stages, the abluminal surface exhibited remarkable changes; 1) the number of blebs increased, 2) elongated microvillous projections emerged, and 3) a thick filopodium, or a migratory appendage developed. It appeared, however, that these changes do not occur synchronously within the entire AV endothelium but were initially observed mostly in the proximity of the endothelial "crease" which was a limited invagination of the endothelial sheet towards the underlying acellular matrix. In addition, even in the proximity of the crease, endothelial cells with flattened basal surfaces were also observed next to endothelial cells that showed apparent morphological indications of transition into mesenchymal cells. These findings suggest that AV endothelial cells are possibly heterogeneous in the competency of transformation into mesenchymal cells and such heterogeneity would be important for maintaining the continuity of the AV endothelium. PMID- 11111330 TI - Cellular hemangioma in an adult. AB - This report describes an adult case of cellular hemangioma arising in the lower lip. A 39-year-old healthy woman presented with a polypoid mass of 4 months duration. The tumor imparted little color to the overlying mucosa and was misdiagnosed as a mucous granuloma preoperatively. The lobular proliferation of plump endothelial cells with inconspicuous vascular spaces was a cardinal morphologic feature of the present tumor. PMID- 11111331 TI - Oxalic acid in PM2.5 particulate matter in California. AB - Ambient air monitoring for organic acids in PM2.5 was conducted at several locations in California. During the study, it was found that oxalic acid (ethanedioc acid) was the most abundant organic acid found in the PM2.5 fraction. Samples from Azuza (in southern California), San Jose (in the San Francisco Bay area), and Fresno (in central California), a PM2.5 Super Site, were collected in 1999 and analyzed. The results for oxalic acid concentrations during this monitoring effort are presented. PMID- 11111332 TI - Nickel speciation of residual oil fly ash and ambient particulate matter using X ray absorption spectroscopy. AB - The chemical speciation of Ni in fly ash produced from approximately 0.85 wt % S residual (no. 6 fuel) oils in laboratory (7 kW)- and utility (400 MW)-scale combustion systems was investigated using X-ray absorption fine structure (XAFS) spectroscopy, X-ray diffraction (XRD), and acetate extraction [1 M NaOAc-0.5 M HOAc (pH 5) at 25 degrees C]-anodic stripping voltammetry (ASV). XAFS was also used to determine the Ni speciation of ambient particulate matter (PM) sampled near the 400-MW system. Based on XAFS analyses of bulk fly ash and their corresponding acetate extraction residue, it is estimated that > 99% of the total Ni (0.38 wt %) in the experimentally produced fly ash occurs as NiSO4.xH2O, whereas > 95% of the total Ni (1.70 and 2.25 wt %) in two fly ash samples from the 400-MW system occurs as NiSO4.xH2O and Ni-bearing spinel, possibly NiFe2O4. Spinel was also detected using XRD. Acetate extracts most of the NiSO4.xH2O and concentrates insoluble NiFe2O4 in extraction residue. Similar to fly ash, ambient PM contains NiSO4.xH2O and NiFe2O4; however, the proportion of NiSO4.xH2O relative to NiFe2O4 is much greater in the PM. Results from this and previous investigations indicate that residual oil ash produced in the 7-kW combustion system lack insoluble Ni (e.g., NiFe2O4) but are enriched in soluble NiSO4.xH2O relative to fly ash from utility-scale systems. This difference in Ni speciation is most likely related to the lack of additive [e.g., Mg(OH)2] injection and residence time in the 7-kW combustion system. PMID- 11111333 TI - The 1999 Fresno particulate matter exposure studies: comparison of community, outdoor, and residential PM mass measurements. AB - Two collaborative studies have been conducted by the U.S. Environmental Protection Agency (EPA) National Exposure Research Laboratory (NERL) and National Health and Environmental Effects Research Laboratory to determine personal exposures and physiological responses to particulate matter (PM) of elderly persons living in a retirement facility in Fresno, CA. Measurements of PM and other criteria air pollutants were made inside selected individual residences within the retirement facility and at a central outdoor site on the premises. In addition, personal PM exposure monitoring was conducted for a subset of the participants, and ambient PM monitoring data were available for comparison from the NERL PM research monitoring platform in central Fresno. Both a winter (February 1-28, 1999) and a spring (April 19-May 16, 1999) study were completed so that seasonal effects could be evaluated. During the spring study, a more robust personal exposure component was added, as well as a more detailed evaluation of physical factors, such as air-exchange rate, that are known to influence the penetration of particles into the indoor environment. In this paper, comparisons are made among measured personal PM exposures and PM mass concentrations measured at the NERL Fresno Platform site, outside on the premises of the retirement facility, and inside selected residential apartments at the facility during the two 28-day study periods. The arithmetic daily mean personal PM2.5 exposure during the winter study period was 13.3 micrograms/m3, compared with 9.7, 20.5, and 21.7 micrograms/m3 for daily mean overall apartment, outdoor, and ambient (i.e., platform) concentrations, respectively. The daily mean personal PM2.5 exposure during the spring study period was 11.1 micrograms/m3, compared with 8.0, 10.1, and 8.6 micrograms/m3 for the daily mean apartment, outdoor, and ambient concentrations, respectively. PMID- 11111334 TI - The formation of reactive oxygen species catalyzed by neutral, aqueous extracts of NIST ambient particulate matter and diesel engine particles. AB - It is important to characterize the chemical properties of particulate matter in order to understand how low doses, inhaled by a susceptible population, might cause human health effects. The formation of reactive oxygen species catalyzed by neutral, aqueous extracts of two ambient particulate samples, National Institute of Standards & Technology (NIST) Standard Reference Materials (SRM) 1648 and 1649, and two diesel particulate samples, NIST SRM 1650 and SRM 2975, were measured. The formation of reactive oxygen species was estimated by measuring the formation of malondialdehyde from 2-deoxyribose in the presence of ascorbic acid; H2O2 was not added to this assay. SRM 1649, ambient particulate matter collected from Washington, DC, generated the most malondialdehyde, while SRM 2975, diesel particulate matter collected from a forklift, yielded the least amount. Desferrioxamine inhibited the formation of malondialdehyde from the particulate samples providing additional data to support the observation that transition metals were involved in the generation of reactive oxygen species. Six transition metal sulfates (iron sulfate, copper sulfate, vanadyl sulfate, cobalt sulfate, nickel sulfate, and zinc sulfate) were assayed for their ability to generate reactive oxygen species under the same conditions used for the particulate samples in order to facilitate comparisons between particles and these transition metals. The concentration of transition metals was measured in aqueous extracts of these particulate samples using ion-coupled plasma mass spectrometry (ICP-MS) analysis. There was qualitative agreement between the concentrations of Fe, Cu, and V and the amount of malondialdehyde produced from extracts of these particulate samples. These data suggest that transition metals can be dissolved from particles in neutral, aqueous solutions and that these metals are capable of catalyzing the formation of reactive oxygen species. PMID- 11111335 TI - An improved inventory of methane emissions from coal mining in the United States. AB - Past efforts to estimate methane emissions from underground mines, surface mines, and other coal mine operations have been hampered, to different degrees, by a lack of direct emissions data. Direct measurements have been completely unavailable for several important coal mining operations. A primary goal of this study was to collect new methane emissions measurements and other data for the most poorly characterized mining operations and use these data to develop an improved methane emission inventory for the U.S. coal mining industry. This required the development and verification of measurement methods for surface mines, coal handling operations, and abandoned underground mines and the use of these methods at about 30 mining sites across the United States. Although the study's focus was on surface mines, abandoned underground mines, and coal handling operations, evaluations were also conducted to improve our understanding of underground mine emission trends and to develop improved national data sets of coal properties. Total U.S. methane emissions are estimated to be 4.669 million tons, and as expected, emissions from underground mine ventilation and methane drainage systems dominate (74% of the total emissions). On the other hand, emissions from coal handling, abandoned underground mines, and surface mines are significant, and collectively they represent approximately 26% of the total emissions. PMID- 11111336 TI - Characterization of products from fluidized-bed combustion of coal. AB - The technology of fluidized-bed combustion (FBC) of coal generates byproducts that have a series of unique characteristics and potential uses in technological practice. In this study, the products of fluidized-bed combustion (FBC-P) of coal derived from Moravian heat stations, a.s. Zlin, Cinergy Global Resources, Czech Republic, were characterized. Particular attention was paid to determining the chemical composition of FBC-P, the content of polycyclic aromatic hydrocarbons (PAHs) and toxic metals in the water leachates of these FBC-P, the content of unburned carbon, the capability of FBC-P to solidify with water and form a solid matrix, and the method for discovering optimum mixing water content for FBC-P solidification. The results suggest that one of the qualitatively more important means of utilizing FBC-P could be their application during solidification/stabilization (S/S) of wastes, particularly wastewater treatment sludges. PMID- 11111337 TI - A fuel-based assessment of off-road diesel engine emissions. AB - The use of diesel engines in off-road applications is a significant source of nitrogen oxides (NOx) and particulate matter (PM10). Such off-road applications include railroad locomotives, marine vessels, and equipment used for agriculture, construction, logging, and mining. Emissions from these sources are only beginning to be controlled. Due to the large number of these engines and their wide range of applications, total activity and emissions from these sources are uncertain. A method for estimating the emissions from off-road diesel engines based on the quantity of diesel fuel consumed is presented. Emission factors are normalized by fuel consumption, and total activity is estimated by the total fuel consumed. Total exhaust emissions from off-road diesel equipment (excluding locomotives and marine vessels) in the United States during 1996 have been estimated to be 1.2 x 10(9) kg NOx and 1.2 x 10(8) kg PM10. Emissions estimates published by the U.S. Environmental Protection Agency are 2.3 times higher for both NOx and exhaust PM10 emissions than estimates based directly on fuel consumption. These emissions estimates disagree mainly due to differences in activity estimates, rather than to differences in the emission factors. All current emission inventories for off-road engines are uncertain because of the limited in-use emissions testing that has been performed on these engines. Regional- and state-level breakdowns in diesel fuel consumption by off-road mobile sources are also presented. Taken together with on-road measurements of diesel engine emissions, results of this study suggest that in 1996, off-road diesel equipment (including agriculture, construction, logging, and mining equipment, but not locomotives or marine vessels) was responsible for 10% of mobile source NOx emissions nationally, whereas on-road diesel vehicles contributed 33%. PMID- 11111338 TI - Production of activated carbons from pyrolysis of waste tires impregnated with potassium hydroxide. AB - Activated carbons were produced from waste tires using a chemical activation method. The carbon production process consisted of potassium hydroxide (KOH) impregnation followed by pyrolysis in N2 at 600-900 degrees C for 0-2 hr. The activation method can produce carbons with a surface area (SA) and total pore volume as high as 470 m2/g and 0.57 cm3/g, respectively. The influence of different parameters during chemical activation, such as pyrolysis temperature, holding time, and KOH/tire ratio, on the carbon yield and the surface characteristics was explored, and the optimum preparation conditions were recommended. The pore volume of the resulting carbons generally increases with the extent of carbon gasified by KOH and its derivatives, whereas the SA increases with degree of gasification to reach a maximum value, and then decreases upon further gasification. PMID- 11111339 TI - Construction and economics of a pilot/full-scale biological trickling filter reactor for the removal of volatile organic compounds from polluted air. AB - The design and the construction of an actual 8.7-m3 pilot/full-scale biotrickling filter for waste air treatment is described and compared with a previous conceptual scale-up of a laboratory reactor. The reactor construction costs are detailed and show that about one-half of the total reactor costs ($97,000 out of $178,000) was for personnel and engineering time, whereas approximately 20% was for monitoring and control equipment. A detailed treatment cost analysis demonstrated that, for an empty bed contact time of 90 sec, the overall treatment costs (including capital charges) were as low as $8.7/1000 m3air in the case where a nonchlorinated volatile organic compound (VOC) was treated, and $14/1000 m3air for chlorinated compounds such as CH2Cl2. Comparison of these costs with conventional air pollution control techniques demonstrates excellent perspectives for more field applications of biotrickling filters. As the specific costs of building and operating biotrickling filter reactors decrease with increasing size of the reactor, the cost benefit of biotrickling filtration is expected to increase for full technical-scale bioreactors. PMID- 11111340 TI - Investigation of the concentration of bacteria and their cell envelope components in indoor air in two elementary schools. AB - Bacterial cell envelope components are widely distributed in airborne dust, where they act as inflammatory agents causing respiratory symptoms. Measurements of these agents and other environmental factors are assessed in two elementary schools in a southeastern city in the United States. Muramic acid (MA) was used as a marker for bacterial peptidoglycan (PG), and 3-hydroxy fatty acids (3-OH FAs) were used as markers for Gram-negative bacterial lipopolysaccharide (LPS). Culturable bacteria were collected using an Andersen sampler with three different culture media. In addition, temperature (T), relative humidity (RH), and CO2 were continuously monitored. Concentrations of airborne MA and 3-OH FAs were correlated with total suspended particulate (TSP) levels. Outdoor MA (mean = 0.78 1.15 ng/m3) and 3-OH FA levels (mean = 2.19-2.18 ng/m3) were similar at the two schools. Indoor concentrations of airborne MA and 3-OH FAs differed significantly between schools (MA: 1.44 vs. 2.84 ng/m3; 3-OH FAs: 2.96 vs. 4.57 ng/m3). Although indoor MA levels were low, they were significantly related to teachers' perception of the severity of indoor air quality (IAQ) problems in their classrooms. Concentrations of CO2 correlated significantly with all bacteria measurements. Because CO2 levels were related to the number of occupants and the ventilation rates, these findings are consistent with the hypothesis that the children and teachers are sources of bacterial contamination. Many culturable bacteria present in indoor air are opportunistic organisms that can be infectious for compromised individuals, while both culturable and nonculturable bacterial remnants act as environmental toxins for both healthy and compromised individuals. Measuring the "total bacteria load" would be most accurate in assessing the biotoxicity of indoor air. Chemical analysis of MA and 3-OH FAs, when coupled with the conventional culture method, provides complementary information for assessing biocontamination of indoor air. PMID- 11111341 TI - Integrating remote sensing and local vegetation information for a high-resolution biogenic emissions inventory--application to an urbanized, semiarid region. AB - This paper presents a methodology for the development of a high-resolution (30 m), standardized biogenic volatile organic compound (BVOC) emissions inventory and a subsequent application of the methodology to Tucson, AZ. The region's heterogeneous vegetation cover cannot be modeled accurately with low-resolution (e.g., 1-km) land cover and vegetation information. Instead, local vegetation data are used in conjunction with multispectral satellite data to generate a detailed vegetation-based land-cover database of the region. A high-resolution emissions inventory is assembled by associating the vegetation data with appropriate emissions factors. The inventory reveals a substantial variation in BVOC emissions across the region, resulting from the region's diversity of both native and exotic vegetation. The importance of BVOC emissions from forest lands, desert lands, and the urban forest changes according to regional, metropolitan, and urban scales. Within the entire Tucson region, the average isoprene, monoterpene, and OVOC fluxes observed were 454, 248, and 91 micrograms/m2/hr, respectively, with forest and desert lands emitting nearly all of the BVOCs. Within the metropolitan area, which does not include the forest lands, the average fluxes were 323, 181, and 70 micrograms/m2/hr, respectively. Within the urban area, the average fluxes were 801, 100, and 100 micrograms/m2/hr, respectively, with exotic trees such as eucalyptus, pine, and palm emitting most of the urban BVOCs. The methods presented in this paper can be modified to create detailed, standardized BVOC emissions inventories for other regions, especially those with spatially complex vegetation patterns. PMID- 11111342 TI - Development of a microscale emission factor model for CO for predicting real-time motor vehicle emissions. AB - The U.S. Environmental Protection Agency's (EPA) National Exposure Research Laboratory has initiated a project to improve the methodology for modeling human exposure to motor vehicle emissions. The overall project goal is to develop improved methods for modeling the source through the air pathway to human exposure in significant microenvironments of exposure. This paper presents the technical description of a newly developed model for CO emissions. The sensitivity analysis and evaluation of this emission model is presented in a companion paper titled "Sensitivity Analysis and Evaluation of MicroFacCO: A Microscale Motor Vehicle Emission Factor Model for CO Emissions." The MOBILE models (used in the United States, except California) and EMFAC models (used in California only) used to estimate emissions are suitable for supporting mostly regional (county)-scale modeling and emission inventory because of their dependence on vehicle-miles-traveled aggregate data. These emission models are not designed to estimate real-time emissions needed for human exposure studies near roadways. A number of independent studies have found that current mobile source emission factor models are not very reliable at estimating microscale emissions and are, therefore, inappropriate for use with microscale modeling necessary to estimate human exposures near roadways. A microscale emission factor model for predicting real-world real-time motor vehicle CO emissions (MicroFacCO) has been developed. It uses available information on the vehicle fleet composition. The algorithm used to calculate emission factors in MicroFacCO is disaggregated based on the on-road vehicle fleet. The emission factors are calculated from a real-time fleet rather than from a fleet-wide average estimated by a vehicle-miles-traveled weighting of the emission factors for different vehicle classes. MicroFacCO uses the same database used to develop the MOBILE6 model. As compared with MOBILE emission models, MicroFacCO requires only a few input variables, which are necessary to characterize the real-time fleet being modeled. The main input variables required are on-road vehicle fleet, time and day of year, ambient temperature, and relative humidity. PMID- 11111343 TI - Idle emissions from heavy-duty diesel and natural gas vehicles at high altitude. AB - Idle emissions of total hydrocarbon (THC), CO, NOx, and particulate matter (PM) were measured from 24 heavy-duty diesel-fueled (12 trucks and 12 buses) and 4 heavy-duty compressed natural gas (CNG)-fueled vehicles. The volatile organic fraction (VOF) of PM and aldehyde emissions were also measured for many of the diesel vehicles. Experiments were conducted at 1609 m above sea level using a full exhaust flow dilution tunnel method identical to that used for heavy-duty engine Federal Test Procedure (FTP) testing. Diesel trucks averaged 0.170 g/min THC, 1.183 g/min CO, 1.416 g/min NOx, and 0.030 g/min PM. Diesel buses averaged 0.137 g/min THC, 1.326 g/min CO, 2.015 g/min NOx, and 0.048 g/min PM. Results are compared to idle emission factors from the MOBILE5 and PART5 inventory models. The models significantly (45-75%) overestimate emissions of THC and CO in comparison with results measured from the fleet of vehicles examined in this study. Measured NOx emissions were significantly higher (30-100%) than model predictions. For the pre-1999 (pre-consent decree) truck engines examined in this study, idle NOx emissions increased with model year with a linear fit (r2 = 0.6). PART5 nationwide fleet average emissions are within 1 order of magnitude of emissions for the group of vehicles tested in this study. Aldehyde emissions for bus idling averaged 6 mg/min. The VOF averaged 19% of total PM for buses and 49% for trucks. CNG vehicle idle emissions averaged 1.435 g/min for THC, 1.119 g/min for CO, 0.267 g/min for NOx, and 0.003 g/min for PM. The g/min PM emissions are only a small fraction of g/min PM emissions during vehicle driving. However, idle emissions of NOx, CO, and THC are significant in comparison with driving emissions. PMID- 11111344 TI - A comparison of nonlinear regression and neural network models for ground-level ozone forecasting. AB - A hybrid nonlinear regression (NLR) model and a neural network (NN) model, each designed to forecast next-day maximum 1-hr average ground-level O3 concentrations in Louisville, KY, were compared for two O3 seasons--1998 and 1999. The model predictions were compared for the forecast mode, using forecasted meteorological data as input, and for the hindcast mode, using observed meteorological data as input. The two models performed nearly the same in the forecast mode. For the two seasons combined, the mean absolute forecast error was 12.5 ppb for the NLR model and 12.3 ppb for the NN model. The detection rate of 120 ppb threshold exceedances was 42% for each model in the forecast mode. In the hindcast mode, the NLR model performed marginally better than the NN model. The mean absolute hindcast error was 11.1 ppb for the NLR model and 12.9 ppb for the NN model. The hindcast detection rate was 92% for the NLR model and 75% for the NN model. PMID- 11111345 TI - Application of the Urban Airshed Model to forecasting next-day peak ozone concentrations in Atlanta, Georgia. AB - Twenty-four to forty-eight-hour ozone air quality forecasts are increasingly being used in metropolitan areas to inform the public about potentially harmful air quality conditions. The forecasts are also behind "ozone action day" programs in which the public and private sectors are encouraged or mandated to alter activities that contribute to the formation of ground-level ozone. Presented here is a low-cost application of the Urban Airshed Model (UAM), an Eulerian 3 dimensional photochemical-transport grid model for generating next-day peak ozone concentration forecasts. During the summer of 1997, next-day peak ozone concentrations in Atlanta, GA, were predicted both by a team of eight forecasters and by the Urban Airshed Model in Forecast Mode (UAM-FM). Results are presented that compare the accuracy of the team and the UAM-FM. The results for the summer of 1997 indicate that the UAM-FM may be a better predictor of peak ozone concentrations when concentrations are high (> 0.095 ppmv), and the team may be a better predictor of ozone concentrations when concentrations are low (< or = 0.095 ppmv). The UAM-FM is also discussed in the context of other forecasting tools, primarily linear regression models and a no-skill, persistence-based technique. PMID- 11111346 TI - DUR: asking and addressing relevant policy questions. PMID- 11111347 TI - Don't label nonadherence a disorder. PMID- 11111348 TI - More on nonadherence. PMID- 11111349 TI - Opportunity for pharmacist involvement in medication error survey. PMID- 11111351 TI - National Legislative Day provides opportunity to showcase pharmacy services. PMID- 11111350 TI - APhA, the Journal of Pharmaceutical Sciences, and the future of the profession. PMID- 11111352 TI - The pharmaceutical sciences in America, 1852-1902. PMID- 11111353 TI - Implementing bone mineral density testing in the community pharmacy. PMID- 11111354 TI - Strategies to improve compensation for pharmaceutical care services. AB - In the past decade, the pharmacy profession has made remarkable strides in implementing a wide range of pharmacy-based patient care services. To foster greater awareness of the value of these services among payers and to ensure the long-term success of pharmaceutical care, pharmacists need to focus more attention on obtaining compensation for these services. In the long run, pharmacists are likely to receive greater net profits from pharmaceutical care than from dispensing. As Norwood et al. noted, pharmacies keep all the revenues they receive from pharmaceutical care as profits and to cover operating expenses, whereas they keep only about 29% of the revenues from the sale of products. By exploring innovative markets for pharmaceutical care services and continuing to improve rates of reimbursement from third party payers, pharmacists can further enhance the revenues they obtain from their growing array of patient care services. PMID- 11111355 TI - How many pharmacists are in our future? The Bureau of Health Professions Projects Supply to 2020. AB - OBJECTIVE: To describe a Bureau of Health Professions model for estimating the numbers and selected demographic characteristics of active pharmacists in the United States and to relate the model's findings. DESIGN: We constructed a model using as base counts data from the Pharmacy Manpower Project census of 1989 to 1991 and advancing the counts annually based on estimates of pharmacists entering and leaving the workforce. The total number of active pharmacists in any year was the sum of the male and female cohorts from age 24 through age 75. The model and its underlying assumptions included consideration of U.S. graduates through 1998, international pharmacy graduates who become licensed in the United States, new schools, type of entry-level degrees, and separation rates. A basic series and high and low alternative series were constructed based on different assumptions. RESULTS: The basic series projected 196,011 active pharmacists in 2000, 224,524 by 2010, and 249,086 by 2020. Estimated pharmacists per-100,000 population were 71.2 in 2000, 74.9 in 2010, and 76.7 in 2020. The workforce was projected to consist increasingly of women: 32% in 1991, 46% in 2000, 50% in 2003, and 64% in 2020. Percentages of graduates receiving the BS degree fell from 94% (1980) to 64.4% (1998) and were projected to decrease to 0% by 2005. Estimated U.S. graduates were 7,945 in 2000, 8,133 in 2010, and 8,452 in 2020. The mean age in 2000 was 38 years for women pharmacists, 46 for men, and 42 overall. Estimates of total pharmacists in 1998 were similar to those from other sources, increasing confidence in the model. CONCLUSION: The Bureau of Health Professions model, which can be readily revised as more and better data become available, provided estimates of active pharmacists by age and sex from 1991 to 2020. The model portrayed an increasingly female pharmacy workforce, with more pharmacists holding the PharmD degree. The model and data are useful for research, analysis, and health care planning. PMID- 11111356 TI - Patient-guided counseling in the community pharmacy setting. AB - OBJECTIVE: To test a new prescription counseling method termed "patient-guided counseling" (PGC) in community pharmacies. DESIGN: Post-test experimental design in which subjects were randomized to three groups. SETTING: Six community pharmacies (three chain and three independent). PATIENTS: Patients presenting new prescriptions. INTERVENTIONS: Patients were randomly assigned to one of three comparison groups. The PGC group was given a written prompt instructing them to write any questions they wished to ask about their prescription or their medical condition. The pharmacist then incorporated these questions into the subsequent verbal counseling. A second group was given a written prompt encouraging them to ask the pharmacist questions. This was followed by customary verbal counseling. A third group served as the control. No prompts were provided, but the pharmacist did provide customary verbal counseling. MAIN OUTCOME MEASURES: Patients' demographics, recall of medication information, and satisfaction with counseling. Patients were contacted by telephone 5 days after the start of drug therapy to measure compliance. Pharmacists rated their satisfaction with the information communicated and with their interactions with patients. RESULTS: Compared with customary verbal counseling, the PGC method was associated with more supplemental questions asked by the patient. Compared with the other two methods, PGC was associated with greater pharmacist satisfaction with the information communicated and slightly longer counseling sessions. No significant differences were found for patients' overall satisfaction with counseling, recall of information, and compliance. CONCLUSION: In the community pharmacy setting, PGC fosters patient participation in medication counseling, a necessary element for the provision of pharmaceutical care. PMID- 11111357 TI - Health promotion beliefs and practices among pharmacists. AB - OBJECTIVE: To examine the health promotion beliefs and practices of pharmacists. DESIGN: Cross-sectional mail survey. SETTING: Indiana. PARTICIPANTS: Staff pharmacists. METHODS: Of the 1,440 registered Indiana pharmacies, one-half were selected using a systematic random process. Questionnaires were mailed to the 720 selected pharmacies asking one staff pharmacist to complete a 73-item questionnaire. RESULTS: A total of 552 pharmacists responded to the questionnaire, providing a 76.7% response rate. The majority of pharmacists believed that 10 of the 20 behaviors encouraged by national health objectives were very important for the average person. There was, however, considerably less agreement among pharmacists about the importance of the other 10 behaviors and practices. Pharmacists' involvement, preparation, and confidence in specific health promotion areas and activities were limited. A number of barriers were cited by respondents that could have hindered pharmacists' involvement in public health education activities. Despite these hindrances, pharmacists appear to be making strides toward providing health promotion education and activities. CONCLUSION: Pharmacists are readily accessible sources of information concerning the importance of needed lifestyle factors on health. They can provide valuable education to patients about improvement of lifestyles as a routine component of pharmaceutical care. PMID- 11111358 TI - Changes in number and distribution of community and ambulatory pharmacies in Virginia from 1994 to 1999. AB - OBJECTIVE: To determine whether market changes have resulted in a decrease in the number or geographic distribution of pharmacies available to ambulatory patients in Virginia. DESIGN: Retrospective review of Virginia Board of Pharmacy records of pharmacy registrations in 1994 and 1999. SETTING: The Commonwealth of Virginia. PARTICIPANTS: All pharmacies classified as outpatient pharmacies (including community and other types of ambulatory pharmacies) and operating in Virginia in 1994 and 1999. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Changes in the total number, geographic distribution, and metropolitan/nonmetropolitan distribution of outpatient pharmacies between 1994 and 1999. RESULTS: The total number of outpatient pharmacies increased from 1,290 to 1,337 between 1994 and 1999. Chain pharmacies, mass merchandiser, and grocery pharmacies increased in number while independent pharmacies declined. There was little change in the geographic or metropolitan/nonmetropolitan distribution of pharmacies. CONCLUSION: Changes in the number and distribution of community and other ambulatory pharmacies in Virginia have not diminished their availability to consumers. PMID- 11111359 TI - Antioxidant nutrients: current dietary recommendations and research update. AB - OBJECTIVE: To review the importance of antioxidant nutrients in the maintenance of health and the prevention and treatment of disease, with a focus on data pertaining to vitamin C, vitamin E, selenium, and carotenoids. A secondary objective was to discuss the new Dietary Reference Intakes released by the Institute of Medicine (IOM) for these nutrients. DATA SOURCES: IOM reports on the use of antioxidant vitamins were reviewed for nutrient recommendations. In addition, a MEDLINE search was performed to identify recent research and review articles on the topic, which were analyzed to identify key research findings in the area. DATA SYNTHESIS: The review discusses the biologic processes of oxidation reactions and antioxidants in biologic systems, provides an overview of information on selected antioxidant nutrients, and explores their role in the prevention and treatment of cancer, cardiovascular disease, ocular disorders, and respiratory disorders. CONCLUSION: There appear to be significant health benefits from dietary antioxidants, as can be found in fruits and vegetables. Some prospective assessment of the effect of supplemental antioxidants also suggests benefit, especially for vitamin E; however, there are conflicting results in this area. Overall, it appears that antioxidant nutrients, especially those from food sources, have important roles in preventing pathogenic processes related to cancer, cardiovascular disease, macular degeneration, cataracts, and asthma, and may enhance immune function. PMID- 11111360 TI - Women's health care during the perimenopause. AB - OBJECTIVE: To review the perimenopause, its associated symptoms, and current management options. DATA SOURCES: Published articles identified through MEDLINE (1966-2000) using the search terms perimenopause and treatment. Additional articles and books were identified from the bibliographies of the retrieved articles. DATA SYNTHESIS: The perimenopause is the transition period from normal ovulatory cycles to menopause. It is associated with erratic fluctuations in reproductive hormone levels, often leading to irregular menstrual cycles, vasomotor symptoms, changes in mood or cognition, and sexual dysfunction. The perimenopause is an ideal time to evaluate a woman's health risks for such common chronic midlife conditions as heart disease, osteoporosis, and some cancers, and to initiate appropriate preventive health measures. Low-dose oral contraceptives and other hormonal therapies are often effective in managing perimenopausal symptoms. CONCLUSION: The transition to menopause is an important time in the female life span that is associated with varied physical and psychological symptoms. Pharmacists should be prepared to provide education about the perimenopause and counsel women on the benefits and risks of various pharmacologic and nonpharmacologic treatments that can ease their passage through this often difficult transition. Pharmacists also are well-positioned to educate and encourage perimenopausal women to initiate lifestyle changes that can enhance their health for the rest of their lives. PMID- 11111361 TI - Reducing the transmission of HIV-1: needle bleaching as a means of disinfection. AB - OBJECTIVE: To review the efficacy, safety, and proper methods for use of bleach (sodium hypochlorite) as a means of needle disinfection. DATA SOURCES: Controlled studies cited in MEDLINE between 1966 and 1999 using indexing terms: needle, bleach, HIV/AIDS, and disinfection. STUDY SELECTION AND DATA EXTRACTION: Studies were categorized based on experimental conditions produced and specific testing procedures used. DATA SYNTHESIS: Used properly, undiluted bleach (sodium hypochlorite 5.25%) appears to be an effective disinfection solution for used needles. Proper needle disinfection with undiluted bleach may reduce the risk of HIV transmission among injection drug users from needle sharing. CONCLUSION: Pharmacists can play a role in reducing HIV transmission among injection drug users by advocating cessation of drug use, drug treatment programs, and avoidance of needle sharing. Pharmacists should be prepared to educate patients who are unwilling to cease illicit drug use or participate in drug treatment programs on the proper methods of bleach disinfection of used needles. PMID- 11111362 TI - Consumers' willingness to pay for pharmacy services that reduce risk of medication-related problems. AB - OBJECTIVE: To measure willingness to pay (WTP) for pharmacists' services directed toward reducing the risk of medication-related problems and to determine factors that have a significant influence on WTP. DESIGN: WTP was measured using a contingent valuation method that involved asking respondents about their maximum WTP for pharmacists' services using a self-administered questionnaire. Respondents' WTP through ex post (out of pocket) and ex ante (insurance premium) methods were measured using three hypothetical scenarios illustrating reductions in the risk of medication-related problems. Logistic regression and semilog regression were performed to evaluate the responses to the survey. SETTING: Outpatient clinic of Robert Wood Johnson University Hospital and two physician offices located in New Jersey. PARTICIPANTS: A convenience sample of 316 patients and/or family members, who visited the study site between March 1 and September 15, 1999, were included. MAIN OUTCOME MEASURES: Amounts patients were willing to pay out of pocket or through insurance premiums for pharmacists' services. RESULTS: The mean WTP out of pocket for pharmacy services ranged from $4.02 to $5.48 per prescription, depending on the level of risk reduction. Mean WTP through an increased insurance premium ranged from $28.79 to $36.29 per year. Overall, the average WTP for a pharmacist's consultation was $5.57, and WTP increased by $0.87 as counseling time increased by 1 minute. WTP was sensitive to changes in the magnitude of risk reduction in both ex post and ex ante scenarios. Income was positively related to WTP, but not to the level of statistical significance. CONCLUSION: Respondents were willing to pay for pharmacists' services that reduce the risk of medication-related problems. Additional investigations to determine the factors that influence WTP for health care professionals' services are warranted. PMID- 11111363 TI - Smoking cessation activities in South Carolina community pharmacies. PMID- 11111364 TI - A student-based introduction to disease state management in pharmacy practice. PMID- 11111365 TI - Improving health literacy: a key to better patient outcomes. PMID- 11111366 TI - Good science is the way to measure vaccine safety. PMID- 11111367 TI - Gene therapy: the first 10 years. PMID- 11111368 TI - Hand-held drug information: applications for hospital pharmacists. PMID- 11111370 TI - A patient's guide to weight loss and weight loss maintenance PMID- 11111369 TI - A pharmacist's guide to weight management. PMID- 11111371 TI - Good report, bad spin. PMID- 11111372 TI - "The medical expert from hell". PMID- 11111373 TI - The medical expert from heaven. PMID- 11111374 TI - Scientific evidence. PMID- 11111375 TI - A global review of HIV infection, its interaction with other infections, and establishment responses. PMID- 11111376 TI - Clinical governance. PMID- 11111377 TI - Structural organization and fine distribution of lymphatic vessels in periodontal tissues of the rat and monkey: a histochemical study. AB - The structural organization and fine distribution of lymphatic vessels in the periodontal tissues (gingiva, periodontium and alveolar process) were examined by light and electron microscopy using an enzyme-histochemical method. Whole mount preparations of periodontal membranes peeled from the teeth and cryostat sections of normal or decalcified tissues treated with EDTA were double-stained using 5' nucleotidase (5'-Nase)-alkaline phosphatase (ALPase) and examined by light microscopy. This staining procedure allowed the lymphatic vessels in the periodontal tissue to be differentiated from blood vessels. Well-developed 5' Nase-positive lymphatics were observed in the gingiva and periodontium. The histochemical aspects of 5'-Nase activity in lymphatic vessels are discussed in detail, with special reference to the supply of Mg++ ions. A network of 5'-Nase positive lymphatics was observed in whole mount preparations of the periodontal membrane for the first time. This network was also observed in the tissue sections. More 5'-Nase-positive lymphatics were seen in the root area of the periodontium than in the cervical area. 5'-Nase-positive lymphatics in residual tissue blocks remaining after cryostat sectioning and in whole mount preparations were highlighted with good contrast and resolution on backscattered electron images produced by scanning electron microscopy. Dense granular precipitations resulting from the 5'-Nase reaction were observed on the luminal surface of the lymphatic endothelial cells as well as on the basal side but were absent in the blood vessels. PMID- 11111378 TI - Immunohistochemical localization of certain inducers related to the proliferation of the pituitary gland and a study on cell contacts between the presumptive neurohypophysis and the adenohypophysis in the foetal mouse. AB - Optic and confocal laser scanning microscopy (CLSM) and the LsAB method were used to immunolocalize TGF alpha, EGF, FGF-2 and their related receptors, which are involved in the regulation of organogenesis in the mouse hypophysis. Internalization of the above receptors and the active proliferation of the presumptive adenohypophysis and neurohypophysis were observed during the mid foetal stage. CLSM images were used to map the distribution of Cx32 in the proliferating hypophysis, particularly between the closely apposed neuro- and adenohypophyses. Using conventional transmission electron microscopy, gap junctions were observed at the boundary of these structures. The results suggest that cell coupling via gap junctions may provide positional information that is then used to control the differentiation of the hypophyseal cells. PMID- 11111379 TI - Sympathetic fibers innervating the extraocular muscles: cells of origin in the cat superior cervical ganglion. AB - Cells of origin of sympathetic fibers innervating the extraocular muscles were demonstrated in the cat superior cervical ganglion (SCG) using the horseradish peroxidase (HRP) technique. Each injection of HRP into the muscles was confirmed by labeling in the oculomotor, trochlear and abducens nuclei. In all cases retrogradely labeled neurons were found to be distributed widely to the ipsilateral SCG. Labeled neurons were round or oval in shape and majority of these were medium in size. There were some large-sized neurons which frequently showed to have long dendrites expanding rostrocaudally. The labeling was characterized by localization in the rostral two-thirds of the SCG after HRP injections into the medial, lateral and inferior and superior oblique muscles. However, the labeling was found in the whole of the SCG after HRP injections into the superior rectus and inferior oblique muscles. Compared with our previous studies demonstrating cells of origin of the sympathetic fibers in the facial and hypoglossal nerves, the present experiments showed quite different pattern of distribution of labeled neurons in the SCG. PMID- 11111380 TI - Immunohistochemical demonstration of inducible nitric oxide and nuclear factor kappa B with reference to age-related changes in the mouse spiral and vestibular ganglion. AB - The morphology and immunohistochemistry of inducible nitric oxide synthase (iNOS) and nuclear factor-kappa B (NF-kappa B) were studied on the spiral and vestibular ganglion of young and old ddy mice. The significant decrease in the number of the spiral ganglion cells and a significant expression of iNOS and NF-kappa B were observed in old mice. In contrast, in the vestibular ganglion of all animals examined, decrease in the number of the ganglion cells or expression of iNOS and NF-kappa B were not observed. Although the relevance of enzymatic systems for the protection of vestibular ganglion cells in old individuals from harmful oxidative stress increased with aging should be further clarified, lack of harmful stress due to nitric oxide (NO) may be one of the plausible reasons for that the vestibular ganglion cells were not decreased in number with aging, since iNOS was not detected in the vestibular ganglion cells in the animals tested in the present study. PMID- 11111381 TI - Two cases of the double inferior venae cavae. AB - Two cases of the double inferior venae cavae (IVC) were found during the student dissection practice in 1997 in Gifu University School of Medicine. On the first case (70-year-old male), the calibers of the right and left IVC were 15 mm and 13 mm, respectively. An anastomosis (4-mm caliber) ran obliquely from the left internal iliac vein to the right IVC. On the second case (86-year-old male), the calibers of the right and left IVC were 15 mm and 10 mm, respectively. We found no anastomosis between the right and left IVC. Each IVC was observed behind the ureter. Both cases belong to Type BC of the classification of McClure and Butler (1925), that is based on the combinations of the right and left IVC, and on their location to the ureters. Both cases also belong to Type II-b-2 of the classification of Takemoto et al. (1978), that is based on the calibers of the right and left IVC and on the running course of the interiliac vein. These are the first and second cases among 808 cadavers in Gifu University School of Medicine and the 93rd and 94th cases in Japan since 1901. PMID- 11111382 TI - [Medical and social impact of mapping the human genome]. PMID- 11111383 TI - [Diagnostic and preventive factors in the background of attempted suicide--review of two-years results of the Pecs Center in the WHO/EURO Multicenter Study of Attempted Suicide]. AB - Since parasuicides are not being systematically recorded in Hungary, a monitoring project within the framework of the WHO/Euro Multicentre Study on Parasuicide was started in Pecs catchment area. The study is aimed both to monitor the medically treated parasuicides and to predicts future acts (repetition prediction study) through a follow-up investigation. Pecs centre has been participated for two years in this study. The recent results of this survey are presented and analysed: 553 monitored suicidal events has been collected; 101 structured interview and 37 follow-up cases has been conducted in this period. The most important epidemiologic sociodemographic and psychosocial factors are discussed. Authors paid special attention to the presuicidal psychological characteristics especially the cry for help of the patients, emphasizing the specific diagnostic and preventive possibilities. PMID- 11111384 TI - [Randomized study of cisplatin-based combination chemotherapy for the treatment of planocellular cancer of the head and neck region]. AB - Between January 1996 and November 1998 38 patients were treated with induction chemotherapy. Patients were distributed in two randomized groups, 19-19 patients in each, receiving (group N) either bleomycin, vincristine and methotrexate (BVM) or the previous medication plus cisplatin (BVCM) chemotherapy (group C). Side effects were low and reversible during the treatments. The clinical regression rate (RR) of the cases was 87% including complete regression 24%. There was no difference between the two groups. There was no difference in the pathological macroscopic regression, however group C was better in microscopic regression. During the 27.5 months of average follow-up time, group N had a significantly better result in the tumor-free survival, with a lower rate of metastatic recidives (N/C = 2/9). There was no significant difference in the overall survival rate, due to the radical neck dissections of recidive metastases. According to our experience we recommend the use of cisplatin in conjunction with neck radiotherapy. PMID- 11111385 TI - [Evaluation of myocardial perfusion using intravenous contrast echocardiography]. AB - The aim of the study was to assess the accuracy of rest myocardial contrast echocardiography (MCE) in detecting perfusion abnormalities after intravenous contrast administration in chronic coronary artery disease. In 21 patients (mean age 49 years) contrast agent was injected intravenously. ATL HDI 5000 ultrasound machine was used. Triggering every fifth cardiac cycle in end-systole apical 2 chamber, 3-chamber and 4-chamber views were used. All patients underwent thallium scintigraphy on the same day and coronary angiography was performed within 24 hours. Second harmonic imaging and power Doppler were used in assessing presence or absence of perfusion, localization and extent of perfusion defects, and their relation with wall motion. In the first group all the 13 patients after myocardial infarction had akinetic segments on echocardiography in accordance with the coronary occlusion detected by coronary angiography. In the second group none of the 8 patients without previous myocardial infarction had wall motion abnormality. Group I: dividing the left ventricle into 16 segments out of 208 segments 44 were akinetic. Perfusion defect was detected by MCE in 29 segments. In 12 segments with wall motion abnormality the normal myocardial perfusion was consistent with viable myocardium, 2 inferior akinetic segments could not be evaluated due to contrast attenuation and in one inferior segment MCE in contrast to the thallium scintigraphy showed no perfusion defect. Group II: good contrast effect was detected in all 128 segments except one inferior segment in which there was a fixed perfusion defect also by thallium scintigraphy and coronary angiography revealed occluded right coronary artery. In conclusion MCE and second harmonic triggered imaging is comparable with thallium scintigraphy in detecting fixed perfusion abnormalities. MCE may contribute to the detection of viable myocardium. PMID- 11111386 TI - [Familial occurrence of Turner syndrome]. AB - The authors report a family in which a woman with the mosaic Turner karyotype 45,X/46,X,r(X) transmitted the ring (X) chromosome to the second daughter and transmitted the X-derived chromosome to the first daughter. The mother had normal fertility with three documented pregnancies and conceptions occurred without any hormonal therapy. Her first pregnancy ended with spontaneous abortion, while the second pregnancy resulted in a 45,X/46,X,+mar female with ovarian failure and the third pregnancy resulted in a 45,X/46,X,r(X) female. The nature of the ring chromosome was confirmed by conventional cytogenetic technics. A small marker chromosome was identified as an X-derived chromosome by fluorescent in situ hibridisation (FISH). PMID- 11111387 TI - Science and technology policy: past and prologue. A companion to Science and Engineering Indicators--2000. PMID- 11111388 TI - [Current significance of surgery in inflammatory bowel disease]. PMID- 11111389 TI - [Current significance of surgery in the treatment of inflammatory digestive tract disease]. PMID- 11111390 TI - [Patient assessment of the long-term benefits of surgery in inflammatory bowel disease]. AB - The objective of surgical treatment in ulcerative colitis (UC) and Crohn's disease (CD) differs. Surgery in UC is more aggressive and potentially curative whereas in CD it is more conservative and palliative. OBJECTIVE: To assess the opinion of patients with inflammatory bowel disease who underwent surgery in the distant past about the results and timing of surgery. MATERIAL AND METHODS: A total of 50 surgical patients (36 with CD and 14 with UC) who had undergone an intestinal surgical procedure at least one year before. The clinical characteristics of patients and details of surgery procedures were recorded. Also, a personal interview was conducted. Patients were asked about their current clinical status, surgical consequences and their opinion about the appropriate timing of surgery. RESULTS: Surgery for UC was total proctocolectomy in 85% of patients and it was on an emergency basis in 43% of them. Surgery for UC was partial intestinal or colonic resection, and it was on an emergency basis in 22% of them. Postsurgical complications were more common in UC than CD patients (50% versus 20%; p < 0.05). In CD surgery, recurrence of disease occurred in 78% of patients within a 2.6 years interval. Among UC and CD patients, 71% and 50%, respectively, reported that their presurgical expectatives had been fulfilled (p = 0.17). CONCLUSIONS: Surgery for UC is associated with an appreciable rate of complications; however, most patients had their expectatives fulfilled with surgery as long-term symptoms were controlled. As for CD, the patient's satisfaction degree was lower than or UC. PMID- 11111391 TI - [Efficacy of four widely used techniques of the diagnosis of Helicobacter pylori infection in gastric ulcer disease]. AB - OBJECTIVE: To study the accuracy of four currently used tests for the diagnostic of Helicobacter pylori infection among gastric ulcer patients with a gold standard as reference which combines several diagnostic methods. MATERIAL AND METHODS: Seventy-three consecutive gastric ulcer patients were prospectively studied. From all patients, three biopsies each were obtained from both antrum and body (two for haematoxylin-eosin staining and one for rapid urease test- Jatrox H.p. Test--. Also, IgG ELISA serology (Helico G) and 13C-urea breath test were performed. According to the gold standard, a patient was considered to be infected with H. pylori when at least two tests were positive; a patient was considered not to be infected with H. pylori when at least three tests were negative. RESULTS: Among gastric ulcer patients, the prevalence of H. pylori infection was 87.6% (95% CI: 78%-93%) with the gold standard as reference. The sensitivity and specificity values were as follows: histology (antrum), 96.8% (89%-99%) and 100% (66%-100%), respectively; histology (body), 98.4% (91%-100%) and 100% (66%-100%); urease test (antrum), 71.8% (60%-81%) and 100% (66%-100%); urease test (body), 96.8% (89%-99%) and 100% (66%-100%); breath test, 100% (94% 100%) and 100% (66%-100%), and serology, 95.3% (87%-98%) and 100% (66%-100%). The sensitivity of the urease test was higher with a body biopsy specimen (McNemar: 15; p < 0.001). CONCLUSIONS: All diagnostic tests (histology, rapid urease test, 13C-urea breath test and serology) are highly accurate for the diagnosis of H. pylori infection among gastric ulcer patients with the exception of the rapid urease test performed with antrum biopsy specimens, where this test displays a lower sensitivity for bacterial detection. PMID- 11111392 TI - [Effect of Helicobacter pylori eradication on histological lesions of gastric mucosa. An 18-month follow-up study]. AB - OBJECTIVE: To evaluate the effect of Helicobacter pylori eradication on pathologic lesions over the gastric mucosa during an 18-month follow-up period. PATIENTS AND METHODS: A total of 122 duodenal ulcer patients infected with H. pylori were prospectively studied. Patients were randomized to receive: ranitidine alone, ranitidine plus antibiotics, or bismuth plus antibiotics. An endoscopy was performed at 3, 6, 12 and 18 months. Haematoxylin-eosin, Giemsa, and Warthin-Starry staining methods were used. Histologic lesions were classified according to the following score: normal (0); superficial chronic gastritis (CG): mild (0.5), moderate (1) and severe (1.5); atrophic CG: mild (2), moderate (3) and severe (4); intestinal metaplasia: absence (0), mild (1), moderate (2) and severe (3). The acute inflammatory activity (active CG) was scored from 0 to 3 (absence, mild, moderate, severe) regarding: inflammatory density in the lamina propria, density of plymorphonuclear leukocytes in the lamina propria, density of intra-epithelial polymorphonuclear leukocytes and superficial erosions. RESULTS: H. pylori eradication was achieved in 31% of patients (0% in the group of ranitidine alone and 48% in patients who received antibiotics). The score corresponding to CG declined progressively after H. pylori eradication, with average values of 2.1 +/- 1.3, 1.98 +/- 1.4, 1.73 +/- 1.6, 1.43 +/- 1.9 and 1.38 +/- 1.9 (p < 0.0001) at 0, 3, 6, 12 and 18 months, respectively. The corresponding score for active CG also improved progressively after eradication: 7.82 +/- 1, 2.51 +/- 0.7, 1.24 +/- 0.6, 0.45 +/- 1.6 and 0.12 +/- 0.5 (p < 0.0001). Nevertheless, no changes were observed regarding atrophia or intestinal metaplasia conditions. CONCLUSIONS: H. pylori eradication is associated with a histologic improvement of gastric mucosa. It begins early and continues for the 18 months after therapy. The improvement in the overall inflammatory component is slow and progressive. In contrast, improvement of the acute component in gastritis is more marked early, and is observed immediately after eradication. Nevertheless, H. pylori eradication is not followed by an improvement in atrophy or intestinal metaplasia. PMID- 11111393 TI - [Retro-auricular inflammation of one month evolution. Acute mastoiditis with subperiosteal abscess]. PMID- 11111394 TI - [Focal seizures in a young man with long term arthromyalgia and recurrent skin lesions. Cardiac myxoma. Neoplastic aneurysm and multiple cerebral infarct due to tumor embolization]. PMID- 11111395 TI - [Persistent neutrophilic meningitis and brain abscesses in a male patient with pulmonary sarcoidosis and corticosteroid therapy. Meningitis and cerebral abscesses due to Nocardia sp. Pulmonary sarcoidosis, steroid treatment]. PMID- 11111396 TI - [Cachectic woman and mediastinal widening. Zenker diverticulitis]. PMID- 11111397 TI - [Cholesterolemia control in Spain, 2000: a tool for cardiovascular disease prevention]. AB - The document "Cholesterolemia control in Spain, 2000: a tool for cardiovascular disease prevention" reviews the current evidence on cardiovascular disease prevention and the therapeutic advances achieved in recent years, in order to aid risk-based clinical decision-making. Cardiovascular diseases rank as the first cause of death in Spain. Their demographic, health and social impact is increasing and it is likely to continue to do so in the next decades. Appropriate treatment for high blood cholesterol and other major risk factors is crucial in cardiovascular disease prevention. Individual risk stratification is essential to determine follow-up periodicity and treatment. Priorities for the control of cholesterolemia and the consequent cardiovascular risk are based on risk stratification. In primary prevention, the therapeutic objective in high risk patients has been established as LDL-cholesterol < 130 mg/dl. In secondary prevention, drug treatment is indicated when LDL-cholesterol > or = 130 mg/dl and the therapeutic objective is LDL-cholesterol < 100 mg/dl. Statins are first line drugs for treatment of high blood cholesterol. In moderate-severe hypertriglyceridemia or low HDL-cholesterol, fibrates are preferred. In acute coronary syndrome, hypolipemiant treatment, should be started as soon as possible, when indicated. Secondary prevention programmes that continually provide good clinical and risk factor control should be provided to coronary heart disease patients. PMID- 11111398 TI - [Diagnostic strategies in inherited hemochromatosis. Value of the genetic test]. PMID- 11111399 TI - [Diagnostic assessment of pleural effusion]. PMID- 11111400 TI - [Diagnostic interpretation of antineutrophil cytoplasm antibodies (ANCA) in vasculitis]. PMID- 11111401 TI - [The use of caloric preparation supplement in chronic renal insufficiency: much data and no proof]. PMID- 11111402 TI - [The use of anticoagulants in patients with cerebral ischemia and atrial fibrillation]. PMID- 11111403 TI - [Follow-up of thyroid function in patient treated with amiodarone]. PMID- 11111404 TI - [Tularemia acquired by tick bites in Castilla-Leon region]. PMID- 11111405 TI - [Polyserositis and arthritis in a patient with meningococcal meningitis]. PMID- 11111406 TI - [What is your diagnosis? Schonlein-Henoch-Glanzmann purpura (also called purpura rheumatica or anaphylactoid purpura) with concomitant glomerulonephritis]. PMID- 11111407 TI - [Percutaneous nephrostomy in treatment of urinary tract obstructions and malformations in the child]. AB - Percutaneous nephrostomy (NP) is a routinely applied procedure in adults, but sparsely reported in the pediatric population. We report the treatment of acute obstructive renal diseases in 15 children aged one day to 6 years. The causes of obstruction included pelvi-ureteral junction stenosis (10 cases) by calculus or complicated by pyonephrosis, vesico-ureteral junction stenosis (2 cases), urethral valves and stenosis (2 cases) and one complex urogenital malformation. Catheter placement was performed under ultrasonographic guidance without immediate complication. NP should be considered as a primary preoperative modality for urinary tract obstruction in the pediatric population. PMID- 11111408 TI - [Development of skin diseases in intravenous drug dependent patients treated with heroin substitution]. AB - Drug addiction is linked with increased prevalence of various illnesses. Of major importance are skin diseases which often have a powerful influence on the health. Analysis of the situation in Switzerland at the outset of the 1990s showed that not all drug addicts could be reached with the existing range of treatments. For this reason, heroin-supported treatment was examined as a new therapy option from 1994 on. The influence on the skin's health are examined in this study. The minimum age of those admitted was set at age 20. Heroin addiction had to date back at least two years, and several treatment efforts had to have failed. Data of 1,035 patients was based on tests at admission and after six, 12, and 18 months of treatment. Some 18% of drug consumers indicated abcesses at admission, and almost 30% showed phlegmones. The prevalence of all skin diseases examined show significant declines over the 18 months of treatment. The relative risk resulting from puncture points fell to 0.35 (SD: 0.26-0.47), from phlegmones to 0.24 (SD: 0.14-0.41), from absesses to 0.31 (SD: 0.15-0.60). This study shows that skin diseases are a frequent and important complication among intravenous drug addicts. Heroin-supported treatment led to favourable progress of the dermatological situation among patients. PMID- 11111409 TI - [Diagnosis and therapy of behavior disorders in dementia]. AB - Alzheimer's disease is one of the most common brain disorder in the elderly. In most patients who develop dementia the core syndrome of cognitive dysfunction is superimposed over the course of the disease by behavioral disorders that manifest at least temporarily to varying degrees. These include depression, anxiety, agitation, restlessness, aggression, disturbances of the sleep-wake cycle, delusions and hallucinations. Classical psychiatric exploration can be complemented by the use of the Neuropsychiatric Inventory (NPI) which allows structured diagnosis and comparative documentation of the clinical course. The majority of psychotropic drugs used in psychogeriatics have not been specifically developed for or tested in elderly, often multimorbid patients. Substances used in psychogeriatrics are subject to special requirements due to the pharmacokinetic and pharmacodynamic changes specific to very old persons. They must show clinical efficacy and a low rate of cardiovascular, peripheral and central anticholinergic effects, a low delirogenic potential and favorable pharmacokinetic and pharmacodynamic properties. Most of the newer atypical neuroleptics (e.g. risperidone) and the newer antidepressants (e.g. from the class of selective serotonin reuptake inhibitors, SSRIs, or selective serotonin and noradrenaline reuptake inhibitors, SNRIs) tend to fulfill these criteria better than the high-potency neuroleptics of the butyrophene type (e.g. haloperidol) or the tricyclic antidepressants. For that reason, these newer products should usually be preferred over the conventional agents. PMID- 11111410 TI - [Chronic abdominal pain in a young diabetic patient]. AB - We report on a 33-year-old patient from Sri Lanka who had been suffering from recurrent episodes of abdominal cramps since he was ten years old. He additionally suffered from postprandial flatulence and an increased frequency of bowel movements. By the age of 24, his condition had worsened with polyuria and polydipsia and he was diagnosed with type II diabetes mellitus. Recently, the patient's compliance deteriorated steadily and his diabetes mellitus was uncontrolled. His flatulence continued and he had six to seven bowel movements daily. He presented to us with renewed bouts of severe stomach cramps, similar to the painful episodes that the patient experienced in his youth. After exclusion of other etiologies and judging by the clinical picture, the patient's origin and the sonographically and radiologically verified pancreatic calcification, we rendered the diagnosis of a tropic calcifying pancreatitis with secondary diabetes mellitus. According to the literature, malignant neoplasia may develop on the basis of this disease. However, we were able to rule out a carcinoma as the cause of the current pain episodes in this patient based on clinical findings and course. We attributed the stomach cramps to compression of the common bile duct by the fibrotic head of pancreas. Pain and cholestasis regressed, thus obviating the need for surgical intervention (pancreaticojejunostomy). On therapy with enzyme substitution and insulin, the patient's exo- and endocrine pancreatic insufficiency was asymptomatic. PMID- 11111411 TI - [Bone manifestations of sarcoidosis Jungling ostitis multiplex cystoides]. PMID- 11111412 TI - [Acute local reaction after intra-articular injection of HYALAN G-F 20 (Synvisc) for treatment of gonarthrosis onset]. PMID- 11111414 TI - Understanding certification PMID- 11111413 TI - International perspectives in occupational and environmental health nursing PMID- 11111415 TI - Health and safety at work in Switzerland: impact of European Union directives. AB - Switzerland, surrounded by European Union (EU) member states, rejected a 1992 referendum to join the European Economic Communities (EEC), which currently includes 15 member states. As a result, the country has had difficulties resolving economic issues with health and safety interests. This study analyzed the consequences of selected EU Directives of Health and Safety at Work in a country that chose not to join the EU. The Directives went into effect throughout the entire EU in 1993. Executive directors and safety advisors from the Swiss company "Migros" participated in a two round Delphi survey focused on timing, feelings, and preference of the legal system in relation to the EU, prioritizing selected EU directives, and implementing health and safety concepts. The results showed the effects of the Directives (although not legally required) demand careful consideration particularly in terms of the timing of the implementation and the priorities of the Swiss health and safety legal system. The two professional groups involved showed congruent opinions on several questions, presenting a solid foundation for planning common action. In conclusion, the growing awareness of occupational health and safety aspects observed during the survey should be pursued among all Migros key staff in decision making positions in occupational safety and health. In this way, Migros could serve as a role model in the occupational health and safety field, much as it has long been recognized as a pioneer in funding social causes throughout Swiss society. PMID- 11111416 TI - The essence of the occupational health nurse's role: a teaching experience in Portugal. AB - Occupational health nurses are in a unique position within the occupational and environmental health and safety team to establish a partnership with the workers to promote their health and safety. The essence of the role of occupational health nurses is the empathetic relationship and approach they are able to develop. Education and training are essential to promote the knowledge, skills, and behaviors of occupational health nurses. PMID- 11111417 TI - Technology enhanced learning for occupational and environmental health nursing: a global imperative. AB - One strategy for decreasing the barriers to higher education and for increasing the competency and performance of the occupational and environmental health nurse in the information age is technology enhanced learning. Technology enhanced learning encompasses a variety of technologies employed in teaching and learning activities of presentation, interaction, and transmission to on campus and distant students. Web based learning is growing faster than any other instructional technology, offering students convenience and a wealth of information. PMID- 11111418 TI - The self employed occupational and environmental health nurse: maximizing business success by managing financial resources. AB - The occupational and environmental health nurse entrepreneur can avoid business failure by engaging in a planning process that maximizes financial resources. Successful financial management involves understanding key financial reports and using those reports as management tools to "keep score" on the business. The prices the occupational and environmental health nurse entrepreneur charges for services will have a direct effect on the success of the business. Payroll, earnings, and expense records are useful management tools to help the occupational and environmental health nurse entrepreneur track the business and meet legal requirements. PMID- 11111419 TI - EEOC's ADA policy guidance: Part II. PMID- 11111420 TI - A potted history of 19th-century remote-area nursing in Australia and, in particular, Queensland. AB - Knowledge of the history of remote-area nursing in Australia is necessary if we are to truly understand the processes that have dictated the boundaries and reality of the scope of practice for today's RANs (remote-area nurses). This paper briefly explores and discusses the social context and history of remote area nursing in Australia and, in particular, Queensland during the 19th century. PMID- 11111421 TI - General practitioners leaving rural practice in Western Victoria. AB - The West Vic Division of General Practice, working with the Department of General Practice, The University of Queensland conducted a qualitative study of GPs who had left western Victoria over the previous 10 years to examine issues relating to the decision to leave rural practice. This study was conducted as part of a project to explore the role of rural Divisions in assisting with general practitioner recruitment and retention. The study supported the conclusions of a similar study in North Queensland and proposed a model that regards rural retention as an interplay of influences both positive and negative with acute trigger factors that can precipitate the decision to leave. Conflict and dissatisfaction with aspects of rural GP hospital work appeared to be a relatively frequent trigger factor that is immediately amenable to intervention from Divisions of general practice and through improvement in negotiation and conflict resolution skills for rural general practitioners. PMID- 11111422 TI - Issues affecting Australia's rural occupational therapy workforce. AB - The unequal distribution of health workers across Australia in favour of urban areas affects the provision of effective health services to rural and remote communities. Additional pressures on the current and future supply of occupational therapists may arise from a restructuring of the health labour force and demographic changes in the age structure of the population. Projections made on the basis of these data indicate that employment growth for occupational therapists will create a demand for 9600 therapists in 2005, or 79.9% more than the number of occupational therapists employed in 1994. Factors such as reductions in the level of immigration and the number of people of working age, and a diminishing population of school leavers to fill student places in universities will make it difficult to meet the projected demand for occupational therapists. Occupational therapy labour force planning suffers from a lack of detailed data on under-serviced areas. Such data are critical for clarifying the magnitude of the projected discrepancy between future demand and supply needs for occupational therapists in rural and urban areas in Australia. PMID- 11111423 TI - Area health services as learning organisations: the rural experience. AB - Staff development units (SDUs) across New South Wales Health are in a state of flux. Traditional models of training may no longer be meeting the continuing professional education needs of staff. This paper outlines how one SDU, the Rural Health Education and Research Centre at Tamworth, with few resources, has successfully negotiated the transformation from delivering ad hoc, face-to-face teaching, to a model encompassing competency-based training, recognition of prior learning, workplace assessment and distance education. PMID- 11111424 TI - The success of men's health nights and health sessions. AB - Men's health nights and men's health sessions have proven to be remarkably successful in rural and some suburban regions of Victoria. In the rural regions, enough interest has been generated to run follow-up health sessions on topics selected at the initial nights. Approximately 2000 men attended these events and 575 filled in questionnaires giving information such as age, occupation, health concerns and perceptions of health professionals. The results indicated that men's health nights appeal to older men who are more likely to be professional or retired. These men saw cardiovascular disease, cancer and stress management as their main health concerns. The majority indicated that they would be interested in attending more men's health sessions. The follow-up sessions provide initial pathways by which men may address the issues of their own health. The data collected contributed to the development of the Men's Awareness Network model for men's health. PMID- 11111425 TI - Inequitable distribution of general practitioners in Australia: analysis by state and territory using census data. AB - The objective of this study was to describe the distribution of general practitioners in each State and Territory, stratified by statistical division and adjusted for estimated community need. The location of general practitioners was obtained from the 1996 Census of Population and Housing. Community need was estimated from crude death rates supplied by the Australian Bureau of Statistics. On average there are 920 people per full-time general practitioner in Australia. Three States are relatively oversupplied by up to 5% (Australian Capital Territory) and the rest are relatively undersupplied by up to 12% (Western Australia). Adjusted for estimated need, the Australian Capital Territory is oversupplied by 71% compared with all of Australia, while Western Australia is undersupplied by 15%. More marked differences occur within the States, with the statistical division containing each capital city in each State that is relatively overserved. The greatest oversupply is in Sydney, where 33% fewer people share a full-time general practitioner than the whole of New South Wales (adjusted for need, oversupply in Sydney is 63%). Relative undersupply is greatest in Queensland, with 133% more people sharing each general practitioner in the north-west statistical division compared with the whole State. The distribution of general practitioners between and within States and Territories is unequal and inequitable. In each State, capital cities tend to be relatively oversupplied compared with more rural areas. While these data do not inform the absolute level of service needed in a community; they do suggest that strategies to redress the inequitable distribution are required. PMID- 11111426 TI - Older patients in the acute care setting: rural and metropolitan nurses' knowledge, attitudes and practices. AB - Many studies reporting nurses' knowledge of and attitudes toward older patients in long-term care settings have used instruments designed for older people. However, nurses' attitudes toward older patients are not as positive as their attitudes toward older people. Few studies investigate acute care nurses' knowledge of and attitudes toward older patients. In order to address these shortcomings, a self-report questionnaire was developed to determine nurses' knowledge of, and attitudes and practices toward, older patients in both rural and metropolitan acute care settings. Rural nurses were more knowledgeable about older patients' activities during hospitalisation, the likelihood of them developing postoperative complications and the improbability of their reporting incontinence. Rural nurses also reported more positive practices regarding pain management and restraint usage. However, metropolitan nurses reported more positive attitudes toward sleeping medications, decision making, discharge planning and the benefits of acute gerontological units, and were more knowledgeable about older patients' bowel changes in the acute care setting. PMID- 11111427 TI - Prevalence and treatment of pressure ulcers in northern New South Wales. AB - This paper reports on a small, cross-sectional study of 18 hospitals in northern New South Wales. The objectives of this study were to collect baseline data on: (i) the prevalence and type of pressure ulcers in a variety of rural hospitals; and (ii) the range of nursing and medical interventions that are used to prevent/treat pressure ulcers. Using a cross-sectional design, the study found that pressure ulcer prevalence, which was 6%, was within the range found by previous reports (4-15%). Pressure ulcer prevention and treatment practices were varied, ranging from turning of the patient and occlusive dressings, to such creative methods as exposure to sunshine and airing the wound. This study demonstrates that despite years of attention to pressure sore prevention and treatment, the prevalence of ulcers is still a significant problem in northern New South, Wales hospitals. PMID- 11111428 TI - Case management: a model for the recruitment of rural general practitioners. AB - Many strategies have been trialed to recruit GPs to rural areas and to retain them once there. The West Vic Division of General Practice, a rural division in central-western Victoria, has developed and piloted a case management model. Case management is the holistic provision of services to meet an agreed outcome. Usually these processes are used to support clients with chronic or complex care needs. In this instance, the Division recognised that the sometimes puzzling and perplexing world of medical and provider registration, rural communities and the politics of bringing a doctor into a community requires specific skills and support. A dedicated and experienced case manager provides a continuous range of services to support the process from identification through to ongoing support within the community. The project has met with considerable success over the past 18 months, with 17 doctors placed in temporary and permanent positions. PMID- 11111429 TI - Bright stars in the sky? AB - Actions to improve the health of people in rural and remote areas are supported by people's ability as individuals and as a community to take part actively in the design, management and evaluation of their own health services. To do this successfully, people need information about their health, resources to support their actions and political support or legitimacy. This article focuses on the third requirement. At the beginning of 2000, there seems to be strong political interest in improving the status of rural communities and rural services overall, including rural health and health services. The constellation of events that contribute to this encouraging state of affairs includes the Regional Australia Summit that was held in October 1999, the existence of a strategic framework for rural health that was agreed on by all health jurisdictions (Healthy Horizons), improved clarity about the relative state of health and health service utilisation in rural and remote areas, and the Prime Minister's recent commitment to improve rural services. The next major national opportunity to convert these positive signs into action comes with the Federal Budget in May 2000, the content of which will be closely watched by people concerned about improving health in country areas. PMID- 11111430 TI - Roles and activities of the Commonwealth Government University Departments of Rural Health. AB - Since 1996, University Departments of Rural Health (UDRH) have been established at Broken Hill, Mount Isa, Shepparton, Launceston, Whyalla, Alice Springs and Geraldton. Each UDRH is underpinned by Commonwealth funding for an initial period of 5 years. The role of the UDRHs is to contribute to an increase in the rural and remote health workforce through education and training programs, as well as a reduction in the health differentials between rural and urban people and between indigenous and non-indigenous peoples. A strong population health focus involving partnerships between existing health providers in a targeted region and the university sector underpins their operation. While UDRHs have been established as a means of addressing a national workforce problem, their organisational arrangements with universities and local service providers vary widely, as does the program mix of activities in education, research service development, facilitation and advocacy. This article outlines some of the activities and progress of the UDRHs to date. PMID- 11111431 TI - An exciting future or has nursing been betrayed? PMID- 11111432 TI - Divided opinions over benefits of NHS Direct. PMID- 11111433 TI - Case 24: nurses with health problems. The UKCC's system for nurses with serious health problems. PMID- 11111434 TI - Applying the principles of infection control to wound care. AB - Human skin in healthy adults is inhospitable organisms. A wound may occur from any accidental or deliberate trauma that breaks the surface of the skin. Once this line of defence is broken there is a risk of infection. All soft tissue injuries, whether chronic, traumatic or surgical, involve the same basic biochemical and cellular processes. This article looks at the risk factors associated with healing of such wounds. The principles of asepsis, wound cleansing agents, choice of dressings and the taking of wound swabs are considered. PMID- 11111435 TI - Developments in wound care for difficult to manage wounds. AB - Research and development in wound healing has ensured that issues relating to chronic wound management remain high in the nursing agenda. Since the advent of modern wound dressings, which retain a moist wound healing environment, work has continued to progress into more advanced, interactive products which aim to alter the wound bed in order to promote a suitable environment for cell migration and growth. Rapid wound healing is advocated and necessary to reduce morbidity and mortality in patients with large chronic wounds and to reduce the financial and manpower implications of long-term wound care in the hospital or community setting. Vacuum-assisted closure, artificial skins, growth factors and larval therapy are discussed in order to give an overview of some of the emerging practices being adopted for difficult to manage wounds. PMID- 11111436 TI - Multiprofessional perceptions of a paediatric nursing development unit. AB - This article describes the final component of the quinquennial evaluation of a King's Fund accredited nursing development unit (NDU) within a paediatric outpatient department of a large district general hospital during the period 1993 1998. The study draws upon descriptive data obtained through a questionnaire distributed to 83 members of staff associated with the outpatient department. Overall, the results demonstrate that NDU status has enabled staff to embrace the change process more readily than might have been possible otherwise. The basic premise that NDUs should promote nursing innovation is explored and critically evaluated. PMID- 11111437 TI - A celebration of 100 years' achievement in child health. AB - This, the second of a two-part article, will reflect upon the gains made in the health and welfare of children within the UK over the last 50 years of the 20th century. It will show how government reforms covering health, welfare, and education have continued to improve all aspects of children's lives. It will also show how nursing, midwifery, and health visiting have tried to continue to meet the changing needs of children and their families. Lastly, it will highlight the increasing rate of change during this time and predict what may await children and young people in the first part of the new millennium. PMID- 11111438 TI - Using biography to enhance the nursing care of older people. AB - This article describes how the use of the biographical approach in nursing practice should enable a fuller understanding of older people as individuals, based on their lived experience. This, in turn, should affect the way in which nurses work with, and care for, older people, encouraging person-centred practice. First, drawing on a life story, the article describes the ways in which people's accounts of their past lives may provide an insight into their present needs, priorities and aspirations; it also helps to reveal and challenge our assumptions about later life. Second, the article discusses why the biographical approach is particularly appropriate for the care of older people in continuing care settings and how this approach could be undertaken. PMID- 11111439 TI - Getting research into practice: developing oral hygiene standards. AB - In 1997, the then Victoria Infirmary NHS Trust established a nursing research and practice development committee (NRPDC) to implement evidence-based practice in nursing care in response to its nursing strategy for 1998-2000. A survey of nursing projects was undertaken in 1996 and repeated in spring of 1998. Initially, 107 projects were identified which included 58 reviews of the literature. In 1998, 95 projects were identified with 42 reviews of literature. The number of research projects being undertaken by nurses in the trust increased from four to 15 and the number of audits increased from nine to 45. The NRPDC established a link nurse system to assist in developing practice at ward level and they have been offering a series of educational seminars. Oral hygiene was the first topic tackled trustwide, with a mouth care standard developed and staff knowledge subsequently surveyed 6 months after it was put into practice. Results demonstrated a good level of knowledge for general oral hygiene among trained and untrained staff. However, specialist oral care and care of stomatitis require some further updating. This survey has identified the increasing sophistication of the projects being undertaken by the nursing staff across the trust and the support they are receiving. This arrangement has provided the opportunity to demonstrate the impact of having a senior researcher available for advice at trust level. The NRPDC can improve the quality of evidence-based care delivered within the trust and it can provide a model for the implementation of evidence based practice. PMID- 11111440 TI - Educational leave for lecturers to regain clinical competence: 1. AB - The clinical credibility and clinical competence of nurse lecturers have presented interesting debate for some years in nursing. While some argue that being clinically competent is an unavoidable necessity for nurse lecturers, others are either ambivalent or believe that they do not have to be clinically competent, being clinically credible should suffice. This article distinguishes between the two terms and presents an analysis of how the latter can be developed through the use of paid educational leave. Such leave, known also as a sabbatical, is one of the possible benefits of being a nurse lecturer within higher education in the UK, and it can be used opportunely as a means of regaining and further developing one's clinical skills and knowledge, if this is perceived as a need. If nurse lecturers value the retention and development of one's own clinical skills and knowledge, then some mechanism should be in place for them to achieve this aim. Johns' (1998) model of structured reflection is used for reflecting on one such experience. The article is in two parts. PMID- 11111441 TI - The use of Clinisan in the skin care of the incontinent patient. AB - The use of skin care regimes for the cleansing of skin when a patient has been incontinent is becoming accepted practice in maintaining healthy skin. Clinisan, produced by Venture Healthcare, is a soap-free, pH balanced foam which has been specifically formulated to cleanse the skin of incontinent patients who are vulnerable to skin breakdown. PMID- 11111442 TI - Nurse prescribing: the sensible way forward? PMID- 11111443 TI - Health and safety ... your problem or mine? AB - This year the topic of Health and Safety has been selected by the Education Committee as the theme for the annual programme. To complement the February Study day held at Birmingham in February, the National Workshop planned for Harrogate in July and the Study Day to be held in Newcastle in November, we are publishing a series of papers on the same theme. Publication can disseminate best practice on a much wider scale and stimulate work based discussion. Over the coming months, different contributors will cover a variety of health and safety topics in the domains of personal care, patient care and environmental considerations, such as latex sensitivity, manual handling, COSHH and risk assessment. In this issue Raymond Rose 'kicks off' the series with a brief outline of social and workplace reforms that have led to key legal statutes and European directives that continue to influence our workbased practices. This is your Journal and we are always keen to respond to your concerns. Please write to either the Editor or myself about topics you feel could be included in this important series. I look forward to hearing from you. PMID- 11111444 TI - How can autologous blood transfusions help our patients? AB - In modern day clinical nursing practice, respecting and encouraging patient autonomy, together with acting as advocate for the patient, is an integral part of holistic patient care. We have a 'duty of care' to our patients. So why, in the early days of the 21st century, are we still transfusing our patients with homologous blood products when there is a safer alternative which has been recognised for almost 200 years? PMID- 11111445 TI - Tension between policy and practice. AB - This is the first in a series of articles in which the political influences on the health service are examined. Niall Dickson, currently Social Affairs Editor with the BBC, gave this paper at Congress 1999--which had the overall theme of Rewind, Pause, Fastforward. Niall Dickson is a former editor of Nursing Times (1983-1988) and he has a continuing interest in matters affecting health and social policy. PMID- 11111446 TI - Adult hypoxaemia in the perioperative period: a review of the literature. AB - As a group of academic skills, learning to search for, obtain, read and critically review literature represents a considerable achievement. For the many perioperative nurses who may never be able to pursue their own research project, reading literature reviews is an excellent alternative which enables individuals to become better and more critically informed. A review of the available literature may provide answers to questions or indicate where a subject is in need of further research. In this article by Denise O'Reilly, perioperative hypoxaemia is defined and a range of literature about the subject is critically reviewed. PMID- 11111447 TI - The burns unit and theatre integration. AB - Disbanding the assigned burns theatre team at St John's Hospital had a major impact on perioperative care. Donna O'Boyle describes how, and what was done to rectify the situation, in her 3M/NATN Award commended work. PMID- 11111448 TI - Professional regulation. Now and in the future. AB - The professional regulation of nursing, midwifery and health visiting is a partnership between the public which expects it, the practitioners who practise it and the UKCC and the four National Boards which co-ordinate and administer the system. PMID- 11111449 TI - Electrosurgery in perioperative practice. AB - No series looking at basics in the perioperative environment would be complete without a good look at electrosurgery. And who better to inform us than Paul Wicker, who gives us all the information and advice we need to maintain patient and staff safety whilst this potentially dangerous but essential apparatus is in use. Paul also answers some of the most common questions about diathermy and its use, and I have certainly been asked all of these myself over the past couple of years. Thanks, Paul. PMID- 11111450 TI - Another kind of battle for the elderly. PMID- 11111451 TI - Mechanism of action of 5-HT3 receptor antagonists: clinical overview and nursing implications. AB - Research has shown that the emetic response is linked to serotonin and the stimulation of the 5-HT3 receptors located in several areas of the body. Consequently, 5-HT3 receptor antagonists have been developed, and many studies have demonstrated their efficacy in controlling chemotherapy-induced nausea and vomiting. Oral formulations of 5-HT3 receptor antagonists are often equally or more effective than i.v. antiemetic regimens and are well tolerated and convenient. Adverse effects associated with the 5-HT3 receptor antagonists are generally mild to moderate in severity and transient in nature. A thorough patient assessment combined with a multidisciplinary approach that involves patients, nurses, physicians, and pharmacists optimizes antiemetic therapy. PMID- 11111452 TI - Therapeutic options for treating advanced colorectal cancer. AB - Despite recent decreases in overall incidence and mortality, colorectal cancer is a major health concern affecting 1 of every 18 Americans. Although potentially curable if detected early, 25% of patients present with metastatic disease, whereas another 10%-60% develop metastases resulting from the spread of microscopic disease not noted at the time of initial surgery. Historical treatment options for advanced colorectal cancer have been unsatisfactory, with survival rates of approximately 9%. This article reviews the newer treatment options that are available to patients while providing nurses with information they need to confidently care for patients receiving these treatments. PMID- 11111453 TI - Metastatic brain tumors: diagnosis, treatment, and nursing interventions. AB - Metastatic tumors are by far the most common cause of brain cancer, with an incidence rate higher than all other types of primary brain tumors combined. Cancer cells can spread to the brain from other sites via the blood supply, lymphatic system, or direct extension. Neurologic symptoms, which are dependent on tumor location, often include headache, weakness, or cognitive deficits. Although the standard of care is evolving, treatment often includes some combination of surgery and radiation therapy. Newer treatments, including radiosurgery and novel chemotherapy approaches, currently are being investigated. These treatment advances have markedly altered the prognosis for patients with brain metastases. Consequently, the status of the systemic components of the malignancy often may be the determinant of outcome. PMID- 11111454 TI - Tyrosine kinase inhibitors: a cure for chronic myeloid leukemia? AB - Chronic myeloid leukemia (CML), a malignant transformation of hematopoietic cells, accounts for one-fifth of all leukemias and will be diagnosed in 4,400 individuals in the United States this year. CML has three phases: chronic, accelerated, and blastic. Interferon, hydroxyurea, busulfan, and bone marrow and stem cell transplantation are used to treat CML, but individuals who are in the accelerated phases or blast crisis usually respond poorly to treatment. A new tyrosine kinase inhibitor, STI 571, is being evaluated in clinical trials. STI 571 is highly bioavailable in oral form and produced minimal toxicities in phase I studies. Further research is needed to determine the rate of response and survival data, but STI 571 holds great promise in the treatment of CML. PMID- 11111455 TI - Chronic lymphocytic leukemia and its impact on the immune system. AB - Myelosuppression and immunosuppression are terms that often are used interchangeably, yet they have very different meanings. Myelosuppression, which is caused by many types of cancer treatments (e.g., chemotherapy, radiation therapy), occurs when the body's population of blood cells is lowered. In contrast, immunosuppression occurs when the body's immune function is compromised. Diseases of either the B or T lymphocytes (e.g., lymphoma, multiple myeloma, CLL) alter the normal functioning of the lymphocytes, rendering them unable to mount an immune response. With CLL, B lymphocytes are unable to mature into immunoglobulin-producing plasma cells (delGiglio et al., 1993). Multiple myeloma occurs when the plasma cells become malignant (Mansen et al., 1997). Knowledge of the basic principles of immunology assists nurses in understanding the complexities of the immune system and the effects of common cancer treatments. Patients with CLL require astute assessment of infectious symptoms, comprehensive nursing care and symptom management, and education about the disease and its effects. Hays and McCartney (1998) also noted that the challenges of caring for patients with CLL include patient management in the outpatient setting, quality-of-life issues, and ongoing support because of the chronicity of the disease. PMID- 11111456 TI - Trastuzumab. PMID- 11111457 TI - Cultural and educational issues in pain management. PMID- 11111458 TI - Catheter malfunction. PMID- 11111459 TI - Pharmacologic and biologic therapies in cancer care. AB - Although Coley first began treating patients more than 100 years ago, he was far ahead of his time in observing the link between boosting the immune system and improving response and survival. The field of immunotherapy has advanced rapidly, and patients' requests to be treated with the newer biologic agents far outweigh requests to be treated with earlier agents such as Coley Toxins (Coley Toxins, 2000). PMID- 11111460 TI - Cancer treatment-related pneumonitis. PMID- 11111461 TI - Women: an endangered species? PMID- 11111462 TI - Social constructions of breast cancer. AB - In contrast to other life-threatening diseases, in which mortality is understood as the fundamental threat, much popular and professional discourse about breast cancer focuses on such issues as the identity, body image, and self-worth of the afflicted woman. Within the Western biomedical tradition, the meaning ascribed to breast cancer has been strongly influenced by competing social interpretations. In this paper, we contend that such social constructions shape the manner in which women experience breast cancer, including their decision making in response to treatment options as well as their strategies for coping with and making sense of breast cancer illness. We argue that an appreciation of the historical and cultural contexts in which breast cancer imagery has been constructed helps to explain the confusing array of ideologies that confront contemporary women diagnosed with breast cancer. PMID- 11111463 TI - The stigma of being named "AIDS carriers" on Haitian-American women. AB - AIDS-related stigma has impacted the lives of many individuals, including Haitians. Little, however, is known about the long-term effects AIDS stigmatization has had on Haitian women living in the United States. A pilot study was conducted to explore Haitian-American women's perceptions of the impact of the AIDS epidemic on their lives. The long-term effects of AIDS-stigmatization fell into five categories: rejection by the dominant society, self-doubt, effect on self-esteem, effect on intimate relationships, and rejection by Haitians within their community. More extensive research is needed to explore the long term effects of the AIDS epidemic on Haitian women. Additionally, to counter the negative effects of the AIDS epidemic, intervention strategies need to be developed and tested. PMID- 11111465 TI - Fatigue in chronic fatigue syndrome: a discourse analysis of women's experiential narratives. AB - Chronic fatigue syndrome (CFS) is a debilitating condition. Approximately 75% of sufferers are women. The etiology of CFS is debated, but remains inconclusive. "Fatigue" is ill defined and conceptually problematic. The international multidisciplinary literature on CFS reveals a paucity of studies on women. Qualitative research to analyze women's discourses on CFS is virtually absent. Eleven New Zealand women of European descent with experience of CFS were interviewed in depth. Within the complex facets of CFS, this article reports specifically on an analysis of discourses on "fatigue." The predominant theme that emerged was that fatigue is articulated as "lack" or absence, which is not representable as an identifiable entity in biomedical terms. Parallels with chronic pain are briefly drawn. We conclude that approaches to CFS must respond to the diverse and complex constructions of the experience of fatigue evident in women's narratives. PMID- 11111464 TI - Feminist perspectives on the social construction of chronic fatigue syndrome. AB - We contrast Western medical views of chronic fatigue syndrome (CFS) etiology, diagnosis, and treatment with views maintained by a predominantly female CFS population. We argue that the failure of Western medicine to demonstrate a viral etiology for CFS led to a paradigmatic shift in research perspectives, which then embraced psychiatric and sociocultural explanations for CFS. As a result, CFS was delegitimized as a biomedical phenomenon within medical, academic, governmental, and public arenas. We compare alternative social constructions of CFS with issues pertaining to multiple sclerosis (MS), an illness that similarly predominates among women. Patient perspectives suggest that the history of medical attitudes toward CFS may eventually parallel the transformations that occurred in relation to MS. In particular, the discovery of biological markers for CFS may lay to rest the categorization of CFS as largely within the psychiatric realm. PMID- 11111466 TI - Exploration of migraine pain, disability, depressive symptomatology, and coping: a pilot study. AB - As many as 29% of women suffer from migraine headache, yet it remains a poorly understood phenomenon. Our purpose in conducting this pilot study was to determine the relationships among migraine pain, disability, depressive symptomatology, and coping in women. A convenience sample of 34 women was recruited from university and workplace populations. Nineteen women met the International Headache Society criteria for migraine, while 15 women served as a nonmigraine comparison group. Participants completed eight instruments measuring migraine pain, disability, depressive symptomatology, and coping. The two groups of women were not significantly different on demographic variables. Migraineurs scored significantly higher for pain characteristics, disability, depressive symptomatology, and total coping scores. PMID- 11111467 TI - Women's perceptions of health care in prison. AB - Fifteen female inmates' perceptions of medical care and the manner in which treatment is provided are explored through individual interviews in a state prison. The women did not hold exclusively negative or positive views about the care and treatment they received; however, the predominant view was negative. Examples of inadequate medical care are described by 14 of the 15 women. Nonempathetic treatment, such as being treated as if undeserving of care, is described by all 15. Examples of adequate medical care and empathetic treatment are offered as well, and the overlap between positive and negative perceptions of care is explored. PMID- 11111469 TI - Measuring outcomes and satisfaction is increasingly important to consumers. PMID- 11111468 TI - Acculturation and attitudes about contraceptive use among Latina women. AB - Previous studies have shown that acculturation among Latinos is associated with increases in health-risk behaviors. This study examined associations between acculturation and contraceptive use among 291 low- to moderately acculturated Latina women. Respondents completed a survey assessing acculturation, contraceptive use, and related attitudes. Moderately acculturated women expressed lower intentions to use contraceptives, were less certain that they would be able to use contraceptives consistently for the next 6 months, and reported lower social support for contraceptive use, than did unacculturated women. Unacculturated women expressed more traditional cultural attitudes favoring large families than did moderately acculturated women. Social norms and low self efficacy may place moderately acculturated Latinas at high risk for unintended pregnancy and STDs. PMID- 11111470 TI - Normative treatment guidelines in home care: building the case. AB - To effectively market programs and maximize resources under a prospective payment system, home healthcare agencies need to look closely at standardizing treatment practices by using normative treatment guidelines as a mechanism to link patient outcomes to the care provided, which can improve quality without increasing costs. This article discusses the issues, methods, and processes used to test treatment guidelines and strategies to improve existing guidelines through evidence-based research. PMID- 11111471 TI - Educating the home health aide about the patient in pain. PMID- 11111472 TI - Parish nursing assessment--what should you know? AB - The prospective payment system will require agencies to become more creative and network with community resources. This article describes a health needs assessment survey that provided the foundation for a parish nurse ministry. The survey revealed that parish nurses could complement home health nurses by filling some of the gaps in healthcare not provided by third-party payers. PMID- 11111473 TI - Insight into accessing the physician. PMID- 11111474 TI - Implementing standard precautions in home care. PMID- 11111475 TI - Using plug-ins and Internet browser extensions. AB - Plug-ins and helper applications are programs that add additional capability to computer Web browsers. These browser extenders allow us to access the most current and most dynamic information on the Web: material that can assist us in fulfilling our professional role now and in the future. We can--and most of us will need to--add browser extensions to our desktop computers both at the time a Web site advises us that such an extension is needed, or we can load the most common browser extensions in advance (see "Essential Plug-Ins"). These steps may seem daunting, but are only inconveniences to our use of the Web as a vital resource and should not be seen as barriers. Happy hunting! PMID- 11111476 TI - Turn the tables on turnover: five keys to maximum retention. AB - Because the current unemployment rate suggests that it's an "employee's marketplace," employers need to do whatever they can to retain their current staff and attract the best possible candidates for future employment. When you transcend typical employer stereotypes and create a work environment that fosters creativity and success, your company will benefit. The end result will be a workforce that stays with you and an agency that thrives. PMID- 11111477 TI - A telehealth primer for managers. AB - Managers must know as much as possible about telehealth and its use in home care. This article gives an overview of the technology and how it is changing home care practice and management. PMID- 11111480 TI - Robin major PMID- 11111479 TI - Nursing image more than what the doctor orders. PMID- 11111481 TI - Exploring multistate licensure. PMID- 11111482 TI - Protecting nursing students from needlestick injury. PMID- 11111483 TI - Accidental exposure to blood and bodily fluids. PMID- 11111484 TI - National ad campaign heightens public awareness of nursing. PMID- 11111485 TI - A new generation of nurses. PMID- 11111486 TI - Nursing in the new millennium: vision, activism, and the role of the media. PMID- 11111488 TI - Clinicals: the start of a new journey PMID- 11111487 TI - Nursing image what meets the eye ... and more. PMID- 11111489 TI - It's all in the mind.... PMID- 11111490 TI - Acute-care nurses' attitudes towards older patients: a literature review. AB - With increases in life expectancy and increasing numbers of older patients utilising the acute setting, attitudes of registered nurses caring for older people may affect the quality of care provided. This paper reviews recent research on positive and negative attitudes of acute-care nurses towards older people. Many negative attitudes reflect ageist streotypes and knowledge deficits that significantly influence registered nurses' practice and older patients' quality of care. In the acute setting, older patients experience reduced independence, limited decision-making opportunities, increased probability of developing complications, little consideration of their ageing-related needs, limited health education and social isolation. Available instruments to measure attitudes towards and knowledge about older people, although reliable and valid, are outdated, country-specific and do not include either a patient-focus or a caring perspective. This paper argues for the development and utilisation of a research instrument that includes both a patient focus and a caring dimension. PMID- 11111491 TI - More than time and place: using historical comparative research as a tool for nursing. AB - Views of history often seem to be inextricably intertwined with assumptions about past and present political systems and the kind of pressure groups they generate. This fact is important when comparing the history of nursing across different time periods. This paper addresses the relevance of the historical comparative method for nurse historians as it applies to remote area nursing in Australia. The method is an under-utilized but legitimate and important methodological approach in health research, particularly when the research involves more than just content analysis of documents from the past. Specifically, when researching the history of nursing in Australia across different time periods, this method has much to contribute to our understanding of health-care practice and policy. Its contribution lies in allowing a student of nursing history to find the lessons from the past, which are many in health care. The historical comparative research method provides the way for comparisons of certain times and events as they relate to the scope of practice of nursing today. PMID- 11111492 TI - Tackling inequalities in health and social wellbeing: evidence of 'good practice' by nurses, midwives and health visitors. AB - This paper details a project which assessed the contribution made by nurses, midwives and health visitors to Targeting Health and Social Need (THSN). Targeting Health and Social Need is an important theme within the Northern Ireland Regional Strategy entitled Health and Wellbeing into the Next Millennium: A Regional Strategy for Health and Social Wellbeing 1997-2002 which is concerned with addressing inequalities in health status and social wellbeing. While there is a considerable body of research and information on variations in health and social wellbeing there is a paucity of corresponding research on those interventions which may improve the health of disadvantaged groups. Health professionals are addressing such inequalities so it is appropriate that their efforts should be recorded and assessed. The project was conducted jointly by the departments of nursing at The Queen's University of Belfast and the University of Ulster, funded by the Department of Health and Social Services and had a timetable that covered three phases. Phase 1 entailed the distribution of a survey questionnaire to all nurses, including those professionals working with community and voluntary groups, to assess their contribution to THSN. The paper describes the results from the 392 questionnaires identified (a response rate of almost 40%) and the work in phase 2 of analysing the responses by selection criteria devised by the Project Advisory Board to allow further investigation. The resulting interventions were reduced to 22 and in an overview of each of these a number of characteristics kept recurring that could serve to define aspects of 'good practice'. These 22 case studies demonstrate the efforts made by nurses and health professionals to change behaviour, practice, the community and the environment. Also highlighted are the nursing competencies gained through working, the lessons learnt and the problems and difficulties encountered. The paper concludes that the use of qualitative research opens the door to measures of social position that reflect the ways in which people define themselves and the relationships that sustain them. Although there is still some way to go in understanding the different approaches to evaluation, an environment that fosters the monitoring and assessment of practice might be created in the future within the nursing community. PMID- 11111493 TI - Chinese husbands' presence during labour: a preliminary study in Hong Kong. AB - This study compares the childbirth outcomes of women whose husbands were present during labour with those whose husbands were absent. A retrospective comparative design was used. Sixty-three Hong Kong Chinese primigravid mothers recruited from childbirth education classes were allocated to one of two groups: those whose husband attended labour (n = 45) and those whose husband was not in attendance (n = 18). Details of demographic characteristics, maternal history and antenatal attendance were obtained along with obstetric measures of maternal anxiety, pain perception, dosage of analgesia used, and length of labour. The results indicated that women whose husbands were present during labour used significantly higher dosage of analgesia than those whose husbands were absent. No significant differences were found between groups in other outcome measures. The researcher concluded that nurse-midwives in Hong Kong need to find ways to help husbands provide the type of support that may help their partners during labour. PMID- 11111494 TI - How first-time mothers cope with child care while still in the maternity ward. AB - The aim of the study was to ascertain those factors that are related to the way first-time mothers assess themselves to be coping with child care when leaving the maternity hospital. The study is part of a wider longitudinal project that follows-up first-time mothers' growth into motherhood for a period of 8 months after the birth. Data collection was done using a questionnaire distributed between January and May 1995. The sample comprised 271 first time mothers. The mothers completed the questionnaire on average on the fifth day after childbirth. A stepwise regression analysis was applied. The mothers who assessed themselves to be coping better with child care were those who went home feeling rested and in a good frame of mind and who had positive experience of child care in the maternity ward. The more functional support the mother perceived herself to receive from members of her social network the better she assessed herself to be coping with child care. Those mothers who had previous experience of taking care of infants also assessed themselves to be coping better with child care. On the basis of the research findings it may be stated that the first-time mother needs support from both nursing professionals and close relatives in order to trust their own abilities to cope better with child care during early motherhood. PMID- 11111495 TI - Hospitals without walls: a journey through the health-care system. AB - The paper describes aspects of a journey through the health-care system following a domestic accident. The journey commenced in the accident and emergency department and, over a 3 month period, traversed the operating theatre, intensive care and an orthopaedic ward before moving into a Hospital in the Home programme and community health and district nursing services. The paper explores the experiences of the accident victim, a 56-year-old man, and his wife who is an experienced nurse and university lecturer. The paper supports the 'seamless delivery' concept of integrated care while, at the same time, sounding notes of caution. The principal messages in this paper are about the relationships that nurses build with patients and their families over long periods of care, the paradoxical sense of 'outsideness' that can occur when the home becomes medicalized and the importance of the home as a healing environment. PMID- 11111496 TI - The effect on staff of essential oil burners in extended care settings. AB - Because hospital wards may be associated with smells, such as chemicals, food, and people and their produce, essential oils in burners were considered a way of improving the ward atmosphere for staff. A study was conducted in three parts. A questionnaire covering a period of 1 month; lavender oil in burners for a 3-month period; and a second questionnaire. A significant number of respondents (88%) indicated in the first questionnaire their belief that the use of essential oil would have a positive effect on the workplace. Eighty-five per cent of respondents to the second questionnaire believed that there had been an improvement in the work environment following the use of the lavender oil burners. PMID- 11111497 TI - Takin' it to the streets. PMID- 11111498 TI - Parenting from prison: helping children and mothers. AB - Incarceration of a mother disrupts the mother-child relationship and the child's emotional development. The researchers evaluated a 15-week parenting program in a women's prison that was designed to enhance mother-child interactions during imprisonment. Pre- and postmeasures for the 104 women were Hudson's (1982) Index of Self-Esteem, Bavolek's (1984) Adult-Adolescent Parenting Inventory, and semistructured questionnaires. Self-esteem and attitudes about expectations of children, corporal punishment, and family roles improved significantly. Empathy and mother-child interactions through visits and letters improved. Participants identified the most helpful components of the program. Those who had been physically, sexually, and emotionally abused and those who had used drugs and alcohol had positive results. Findings support the value of parent education for self-development of incarcerated mothers and for the welfare of their children. PMID- 11111499 TI - Factors predicting postoperative pain in children and adolescents following spine fusion. AB - Despite advances in research and technology, investigators around the world acknowledge the continued undertreatment and unnecessary suffering of children in pain. The dilemma of inadequate relief of children's pain may relate to the complexity of the pain phenomenon. Using the Gate Control Theory as the foundation for this work, the purpose of my study was to determine the extent to which selected variables (severity of operative procedure, postoperative use of pharmacological agents, prior experience with pain, pain tolerance, gender, and age) predicted children's and adolescents' pain following spine fusion. Using a descriptive correlational design, data were collected from 93 children (ages 8 to 21 years) who had undergone spine fusion. For four consecutive postoperative days, children were asked to rate the intensity of their pain using the Adolescent Pediatric Pain Tool and observed pain behaviors were recorded using the Child Pain Scale. Repeated measures MANOVA revealed that children continued to experience moderate to severe pain throughout the four days. Stepwise multiple regression analyses were conducted for each postoperative day on each of the dependent variables. A modest amount of variance in children's pain was explained by the variables studied. R square values suggested that age, pain tolerance, and severity of operative procedure have the greatest potential as predictors of children's postoperative pain and warrant future research. PMID- 11111500 TI - Psychometric testing of the adolescent version of the Cook-Medley hostility scale. AB - Hostility, a phenomenon that has been linked to cardiovascular disease and all cause mortality, is often measured by the Cook-Medley hostility (Ho) scale. Although there is an adolescent version of the Ho scale, it has had little testing with only Anglo-American samples. This pilot study tested the reliability and validity of the adolescent version of the Ho scale with a multiethnic sample of adolescents. Sixty adolescents participated. Reliability was measured using Chronbach's alpha. The Ho scale was correlated with the Speilberger State-Trait Anger Expression Inventory (1988) scales to determine concurrent validity. A factor analysis assessed construct validity; content validity was assessed by analyzing tape-recorded descriptions of a circumstance that provoked anger, as remembered by each adolescent. Internal consistency reliability was .75. The hostility measure was most highly correlated with anger expression (r = .62, p = .000) and trait anger (r = .50, p = .000). The factor analysis generated three factors (suspicious alienation, cynical aggression and justified mistrust), that accounted for 34.5% of the variance. The content analysis resulted in five anger provoking themes: aggression, unfulfilled personal expectations, mistrust/lying, criticism of effort, and rejection. The majority of items in the scale (74%) were related to one of the themes. PMID- 11111501 TI - Maternal sensitivity and responsiveness, limit-setting style, and relationship history in the transition to toddlerhood. AB - During the transition of their children to toddlerhood, mothers must learn to adapt their behaviors in a period of marked developmental change. Maternal sensitivity and responsiveness were examined across interactions with varying levels of control-saliency over children at 12 months. Mothers were significantly less sensitive as a group in more control-salient interactions (teaching task, toy clean-up, and limit-setting), than in less control-salient interactions (play and snack). Mothers' sensitivity and responsiveness, and their perceived relationship history in their own families of origin, also were related to their use of limit-setting strategies. Mothers who were less sensitive and responsive, and who reported more negative relationship histories, displayed power-based limit-setting strategies. In contrast, higher levels of maternal sensitivity and responsiveness and more positive relationship histories were associated with teaching-based limit-setting styles. PMID- 11111502 TI - National grassroots strategic planning for NIMH: you can be a part of it. PMID- 11111503 TI - An organizational framework for conceptualizing resilience in children. AB - TOPIC: An organizational framework for conceptualizing resilience in children. PURPOSE: To propose a framework based on relevant literature that clarifies, differentiates, organizes, and elaborates on pertinent factors associated with resilience in children. SOURCES: Relevant literature from developmental psychology, child, psychiatry, and nursing. CONCLUSIONS: Salient factors affecting resilience in children originate internally or externally to the individual. Internal factors include biological and psychological factors; external factors are reflected in the nature and quality of relationships established within or outside the family group. The influence and importance of each factor, however, may vary in individual situations. The framework can guide research efforts and facilitate interventions for practice. PMID- 11111504 TI - Attention deficit hyperactivity disorder: current concepts and underlying mechanisms. AB - TOPIC: Neuropsychological concentration of attention deficit hyperactivity disorder (ADHD). PURPOSE: Survey of current understanding of underlying neural systems and pathways involved in ADHD and the relationship to specific behavioral patterns. SOURCES: Literature review, author's experience in neuropsychological assessment and clinical treatment. CONCLUSIONS: Clinicians will be able to specify treatment interventions designed to compensate for specific behavioral patterns and functional deficits. PMID- 11111505 TI - Friends and pets as companions: strategies for coping with loneliness among homeless youth. AB - PROBLEMS: Loneliness and negative health outcomes associated with being homeless and living on the streets. METHODS: Qualitative data from 32 homeless youth, ages 16 to 23 years, who participated in focus groups, and a subsample of 10 youth, ages 15 to 23 years, who participated in individual interviews, were analyzed using manifest and content analysis, techniques. FINDINGS: Homeless adolescents who live on the streets or in "squats" described feelings of loneliness that they say "go with the territory." Three themes emerged from the data: how lonely subjects felt, circumstances that provoked feelings of loneliness, and ways of coping with loneliness. Thirteen identified their pets as companions that provided unconditional love, reduced feelings of loneliness, and improved their health status. CONCLUSIONS: Vulnerable adolescents who are homeless often recognize the therapeutic value of pets. Interventions that enhance this coping strategy need to be developed and tested. PMID- 11111506 TI - From the inside looking out: violence in schools. PMID- 11111507 TI - Continuing to gain knowledge. PMID- 11111508 TI - Immunization update: a community-based nursing education program. AB - BACKGROUND: Nurses play key roles as advocates, educators, and researchers to eliminate barriers to access as well as educate the public on the importance of vaccines for children in the United States. METHOD: Fifty-four nurses attending a community-based immunization program completed a pretest questionnaire which included identification of related practice behaviors and general knowledge questions. Twenty-four responded to a similar posttest survey 6 months later. The investigators compared knowledge pretest and posttest scores with a t test. Behavioral changes were compared between the pretest and posttest group by analyzing the percentage of change between the time period. RESULTS: The mean score in the knowledge section increased significantly from a pretest mean of 52% to 75% during the posttest period (p < 005). Behavior change was not statistically significant because of the low number of posttest responses indicating actual immunization practices in the clinical setting. The investigators analyzed trends in practice by comparing percentages of responses. CONCLUSION: These findings suggest that a multifaceted, community-based nursing continuing education program including both local, ethnic community perspectives and information related to the latest standards of practice can impact knowledge levels and some practice behaviors. PMID- 11111509 TI - An outcomes model prototype: integrating continuing education learning into practice. AB - BACKGROUND: A method for measuring the downstream effects of a continuing education conference on nursing practice was developed and tested. METHOD: In collaboration with conference faculty, the authors developed an assessment of knowledge and nursing experience related to the course content. Knowledge about the conference content was measured before the conference and at its conclusion. Experience was measured before the conference and 3 months after. RESULTS: Increases were found in both knowledge and experience, indicating that knowledge gained at the conference was integrated into nursing practice. CONCLUSION: Based on positive findings from this pilot, the method will be tested further. PMID- 11111510 TI - Advanced cardiac life support education: a self-directed, scenario-based approach. AB - BACKGROUND: The American Heart Association adopted new curriculum guidelines for Advanced Cardiac Life Support (ACLS) education. Requirements include the incorporation of case-based learning strategies in the course format. METHOD: A self-directed, scenario-based course format was developed to incorporate principles of adult learning and to meet the newest curriculum guidelines of the American Heart Association. A post-course survey was used to evaluate the participants' perceptions of the course format and its effects on their learning. RESULTS: Twenty-seven participants completed the course. Analysis of the post course survey responses indicated participants favored the self-directed, scenario-based design and felt the format contributed to their learning. Additionally, this format was more cost-effective than the traditional 2-day course design. CONCLUSION: The use of a self-directed, scenario-based format creates an optimal environment that fosters the complex cognitive and psychomotor skills learning required to perform ACLS. This format meets the expectations of the American Heart Association curriculum guidelines and should be considered a viable and valuable option for providing ACLS education. PMID- 11111511 TI - Meeting the challenge of developing courses for distance delivery: two different models for course development. AB - Nursing education is increasingly challenged to convert traditional course offerings to distance delivery modalities to accommodate practicing RNs who wish to pursue continuing education. There is a lack of understanding regarding the experiences of faculty and staff from other departments within the university in the development of distance education courses. The purpose of this descriptive study was to uncover the experiences of nursing faculty and members of a university support unit involved in interdisciplinary development of distance delivery courses. Interviews were conducted with 11 participants. Two organizational models were identified. Other issues that emerged in the development of distance delivery courses were: faculty ownership of distance courses; workload implications for faculty; clinical practice by distance; and faculty expertise in new technologies. Implications for incorporating the challenges facing nursing faculty in the 21st century in the development of distance delivery courses are discussed. PMID- 11111512 TI - Advocating for continuing nursing education in a pediatric hospital: the Prince Scholar and Sabbatical Programs. AB - As nurses gain more experience, they often question the basis of nursing practice and want to find the most current and accepted methods of providing nursing care. Attending seminars, conferences, and continuing education programs is often difficult because of financial and staffing constraints. The authors describe the design and implementation of two funded programs--the Prince Scholars and Sabbatical Programs--that support continuing nursing education in a pediatric tertiary hospital. PMID- 11111513 TI - Advancing the development of human resources in nursing in Jordan. AB - BACKGROUND: This article describes a collaborative international health and development program between a Jordanian and a Canadian university. It presents a human resource development model in Jordan that has been the basis for a variety of developmental activities for practicing nurses and nurse educators in academic and clinical settings. METHOD: Reciprocal visits by leaders of the project, as well as continued collaboration between key members of the two universities were instrumental in ensuring success of this venture. RESULTS: The activities implemented in this project culminated in the development of a pragmatic human resource development model that is sensitive to issues particularly relevant to the Jordanian culture. CONCLUSION: The collaborative venture discussed in this article has enabled nurses and nurse educators to increase their academic and clinical skills and raised the profile of the nursing profession in Jordan. PMID- 11111514 TI - Cardiovascular health for older adults. PMID- 11111515 TI - Age and gender differences in pain management following coronary artery bypass surgery. PMID- 11111516 TI - Factors associated with referral of elderly individuals with cardiac and pulmonary disorders for home care services following hospital discharge. AB - Referrals for home care services initiated prior to hospital discharge may prevent or delay readmission or nursing home placement, especially for elderly individuals with multiple, chronic health problems. While multiple factors could justify the need for home follow-up after hospital discharge, little is known about those patient factors associated with clinicians' decisions to refer older adults with cardiac or pulmonary disorders. Increased understanding of factors that contribute to initiating a home care referral could enhance clinicians' decision-making and thus improve post-discharge outcomes for these patient groups. This study examined patient factors associated with and predictive of the decision to refer for home follow-up, using a sample of older adults hospitalized with chronic obstructive pulmonary disease (COPD) or congestive heart failure (CHF). Study findings suggest a model that includes patients diagnosed with both COPD and CHF, who are not married, need home health aides, and have a longer than average length of hospital stay may be helpful in predicting the need for home care referrals. PMID- 11111517 TI - Dietary problem-solving skills among older adults with coronary heart disease. PMID- 11111518 TI - Cardiovascular risk assessment in elderly individuals. PMID- 11111519 TI - Heart health and older women. PMID- 11111520 TI - What has been your experience with hypodermoclysis? What have been the burdens and benefits to your clients for whom it was used? PMID- 11111521 TI - Regulations governing special care units. PMID- 11111522 TI - Variations on a theme. PMID- 11111523 TI - Fast-tracking the postanesthesia patient: the pros and cons. AB - Fast-tracking patients in the perianesthesia setting is a health care practice initiative that is rapidly increasing in popularity in hospitals and freestanding surgery centers throughout the United States and the world. ASPAN recognizes the efficacy of fast-tracking ambulatory surgery patients through the postanesthesia phase of care and currently recognizes 2 important models of fast-tracking: rapid PACU progression and bypassing Phase I PACU. This article reviews the pros and cons of these fast-tracking initiatives. Finally, ethical issues will be critically evaluated to ensure safe, quality, cost-effective care is maintained when fast-tracking this patient population. PMID- 11111524 TI - Management of the extracorporeal shock wave lithotripsy patient. AB - Over 2 million Americans experience kidney and urinary stone disease each year. Early treatments resulted in high mortality and morbidity rates. With the advent of extracorporeal shock wave lithotripsy less than 20 years ago, treatment for this disease has become far safer with more rapid recovery and fewer complications. The selection of patients eligible for extracorporeal shock wave lithotripsy is dependent on the location and size of the stones and the overall health of the patient. This article discusses the different treatment modalities used for stone disease and the different methods currently available for extracorporeal shock wave lithotripsy. Preprocedure preparation of the patient and postoperative care for this population is reviewed in detail. PMID- 11111525 TI - Limitations to self-care in the ambulatory surgical patient. AB - Myalgia, or muscle pain, is a common postoperative phenomenon. Causes of myalgia include age, gender, medication, positioning, and abdominal insufflation for laparoscopic surgery. For some patients, myalgia may be more severe than surgically induced pain. This study explored postoperative myalgia from the perspective of self-care limitations. Sixty-three individuals were surveyed, and responses were analyzed using unidimensional and multidimensional scaling techniques. Subjects reported back pain and the inability to accomplish high energy activities of daily living to be most incapacitating after surgery. Results of this investigation may be used to structure preoperative teaching and perianesthetic nursing interventions. PMID- 11111526 TI - Building spirited multidisciplinary teams. AB - Managing multidisciplinary teams in complex settings such as the perianesthesia setting is a leadership challenge. With the addition of work redesign and cross training initiatives, the task of helping people form healthy working relationships is substantial. The Team Spirit model (The Expanded Learning Institute, Del Mar, CA) demonstrates how employing the phases of Initiating, Visioning, Claiming, Celebrating, and Letting Go will build team spirit and more effective teams. The model and the phases are described with examples given of how getting to know each other, addressing differences, developing a shared vision, doing the work through effective communication, and celebrating will lead to spirited multidisciplinary teams. PMID- 11111527 TI - Practical points in spinal endoscopy. AB - This article presents an overview of spinal endoscopy, including history, indications, and contraindications, along with the anatomy and physiology of the spine. A description of the procedure is provided, complications are identified, and nursing considerations are highlighted. PMID- 11111529 TI - Recognizing Horner's syndrome. AB - Horner's syndrome is a dramatic finding identified by perianesthesia nurses after regional anesthesia. This article describes the relationship between Horner's syndrome and regional anesthesia while explaining the signs and symptoms as they relate to blockade of the sympathetic nervous system. PMID- 11111528 TI - How will we manage? AB - Management of perianesthesia nursing units is complex. Comfortable practice that upholds the status quo but does not embrace opportunities may stagnate staff and management. The challenge is to focus on what is done well, and what practice should be questioned. Creating a unit atmosphere that supports change and explores growth requires effective nurse managers. Critical qualities set the tone for professionalism and positive growth of the staff in the unit, and the unit itself. PMID- 11111530 TI - Notes from the American Society of Anesthesiologists annual meeting. PMID- 11111531 TI - The state of nursing. PMID- 11111532 TI - Be free, be proud. PMID- 11111534 TI - Seven habits of highly effective school nurses PMID- 11111533 TI - Caught up in the Web ... Part III PMID- 11111535 TI - How a multidisciplinary vascular access care program enables implementation of the DOQI guidelines. Part I. AB - This comprehensive, proactive, multidisciplinary team approach to access management has enabled the achievement of center-specific best-demonstrated clinical practiCes for vascular access care. It has also resulted in significant cost savings to the health care delivery process. It has not been an easy task; if it were, access care outcomes would be better nationally than they are today. The VACP approach to vascular access care improvement employs four key implementation principles that ensure the success of Gambro's program and form the infrastructure supporting any successful team approach to care. These core processes, known as the four "C's, include: 1. Commitment, 2. Continuous Quality Improvement, 3. Core Competency, and 4. Communication. PMID- 11111538 TI - The graft vs. the fistula... AB - As the authors note, much has been written about the problem of vascular access. Yet, it remains a major issue in renal care. What are others saying about determining the best strategy for battling the "Achilles' Heel" of dialysis? Following are two abstracts that support use of the fistula. PMID- 11111537 TI - Using CQI and the DOQI guidelines to improve vascular access outcomes: the Southern California Kaiser Permanente experience. PMID- 11111539 TI - The impact of managed care on providing renal nutrition services. PMID- 11111540 TI - E-commerce gives renal providers a new form of purchasing power. New, old suppliers introduce electronic buying. PMID- 11111541 TI - Using e-mail in renal patient care: panacea for information overload? PMID- 11111542 TI - A survey of job satisfaction and performance factors among technical and patient care practitioners. PMID- 11111543 TI - Important ingredients in dialysis delivery: an international perspective. PMID- 11111544 TI - Advanced practice nurses: roles in the hemodialysis unit. AB - Newly diagnosed cases of end stage renal disease (ESRD) have increased by 9% each year since 1970. It has been estimated that there will need to be a significant increase in the number of nephrologists to care for the ESRD population by the year 2010. Recent reports have advocated the use of advanced practice nurses (APN) to collaborate with nephrologists to meet increasing patient care demands. Clinical evidence has supported the financial and clinical advantages of APN utilization in nephrology. The renal community has stressed an outcome-based practice with a provision of guidelines to improve morbidity and mortality in ESRD. Reimbursement and mortality have been linked to identification of quality care delivery. APNs can be instrumental in assuring that quality patient care is delivered across the ESRD continuum through several different roles: clinician, educator, consultant, researcher, administrator, and case manager. PMID- 11111545 TI - Nephrology APNs: who are we and what do we do? Survey results October 1999. AB - In an attempt to identify characteristics of the nephrology APN, attendees at the January 1999 Advanced Practice/Management Seminar in Savannah, Georgia were surveyed. The results discussed focus on the 46 attendees who identified themselves as APNs. They represented 25 states and were predominately master's prepared nurse practitioners with both CNN and advanced practice certification. Most were over 40 years old (83%) with over 16 years of nursing experience (76%), the majority (78%) were in their current APN role less than 5 years. The modal salary range was $60,000-$65,000 (30%). Further demographics, scope of practice and role satisfaction are discussed. PMID- 11111546 TI - Interviews with advanced practice nurses: a day in the life. Interview by Sherry L. Child. AB - The role of the advanced practice nurse in nephrology is evolving rapidly. The following questions and role descriptions were posted on the ANNAlink Advanced Practice ListServe. The dialogue is just beginning--stay tuned! Stay informed and connect to advance your practice! PMID- 11111547 TI - Lupus nephritis: pathophysiology, diagnosis, and collaborative management. AB - Lupus nephritis (LN) is a complex disease. The pathophysiology involves the glomerulus and mesangium, and its manifestations are exhibited in extensive renal lesions. The World Health Organization (WHO) has developed a classification system to assist clinicians in understanding the severity of renal involvement. The diagnosis of systemic lupus erythematosus (SLE) and LN can be difficult because of their often vague symptoms and the long list of differential diagnoses. However, it is important to identify LN quickly to have an impact on a patient's prognosis. The 11 criteria established by the American Rheumatology Association provide a framework for identifying clinical manifestations of SLE. Management of LN, which may be guided by renal biopsy findings, includes pharmacologic therapy, management of drug toxicities, dialysis, transplantation, controlling symptoms (e.g., hypertension), patient education, and psychosocial support for the patient and family. This article focuses on the pathophysiology, diagnostic approach, and collaborative management of LN. PMID- 11111548 TI - Call week in hemodialysis. PMID- 11111549 TI - Exploring hope in patients with end stage renal disease on chronic hemodialysis. AB - The purpose of this qualitative study was to explore the definitions and sources of hope in patients with end stage renal disease (ESRD) receiving chronic hemodialysis. A convenience sample was recruited from a population of chronic hemodialysis patients from two dialysis centers in a rural area of the Pacific northwest. Study participants consisted of 9 men and 5 women between the ages of 43 and 81 (M = 62.5) who had been on chronic hemodialysis an average of 8.3 years. The data collection process consisted of an audiotaped interview guided by the pre-established research questions. The results of this study provide examples of the experience of hope in patients with ESRD on chronic hemodialysis. Hope is a multifaceted human response. The participants in this study were able to adapt to situational changes by attaching their hopes to reality consideration, therefore developing a cognitive process for maintaining hope. PMID- 11111550 TI - Iron overload in the erythropoietin era. AB - Increasing use of maintenance parenteral iron in the erythropoietin (EPO) era has been accompanied by growing concern about iron overload. This article attempts to put the issue of iron overload in hemodialysis patients into perspective. The condition is less common in all dialysis patients today than it was in the pre EPO era, since fewer patients are being transfused and EPO therapy shifts iron into erythroid cells. Patients with end stage renal disease (ESRD) are less likely than patients with hemochromatosis to develop iron-induced organ dysfunction. Diagnosis of iron overload is best accomplished through liver biopsy. Clinically significant iron overload, which rarely occurs if ESRD patients are properly managed, can be treated in most EPO-treated renal failure patients by simply withholding parenteral iron therapy. PMID- 11111551 TI - Association between hematocrit level and mortality in hemodialysis patients. Case study of the anemic patient. AB - A large, 4-year, retrospective study of the HCFA end-stage renal disease (ESRD) claims database, Parts A and B, was conducted to determine the association between hematocrit (Hct) level and survival in patients on hemodialysis. Patients who survived the last 6 months of each year and had at least 4 Epoetin alfa claims qualified for the study. Cohort entry years were 1990 through 1993, with the relative risk (RR) of mortality evaluated during the following year. Patients were stratified into four groups on the basis of mean Hct levels in the 6-month entry period: < 27%, 27% to < 30%, 30% to < 33%, and 33% to 36%. Using the 30% to < 33% Hct group as reference (relative risk (RR) = 1), patients whose mean 6 month Hct was in the 33% to 36% range were associated with significantly lower RR of mortality, while patients whose mean 6-month Hct was below 30% were associated with significantly higher RR of mortality. This epidemiologic study highlighted an important association between higher hematocrits in hemodialysis patients and lower RR of mortal. PMID- 11111552 TI - Life-threatening illness in a nontransfusable patient: a health care challenge. PMID- 11111553 TI - Heparin-induced thrombocytopenia in hemodialysis patients. PMID- 11111554 TI - Basiliximab (Simulect): simplifying induction therapy. PMID- 11111557 TI - Nurse practitioner versus physician assistant. PMID- 11111558 TI - The impact of prior nursing-like work experience on a student's clinical performance. PMID- 11111559 TI - Computer-based testing system. PMID- 11111560 TI - Authoring software for courses delivered on the Web, part 3: Designer's Edge/net synergy. AB - Designer's Edge is an excellent tool for any course developer who is new to the concepts of instructional design or who wants to formalize the process of designing online courses. Integrated storyboarding and publishing options for the Web make this application even more valuable, if the user is either satisfied with limited control over text formatting or is HTML knowledgeable. The steep price of Designer's Edge makes this tool most useful to larger universities and institutions that want to standardize and guide their online course development. PMID- 11111561 TI - An ongoing initiative to reach rural family nurse practitioner students. AB - The challenges we have encountered in educating rural nurses to become nurse practitioners are not unique to Penn State but rather are similar to those encountered by other programs with similar missions. As emphasized in these reports, providing distance education to distance nurse practitioner students requires patience and flexibility. Also emphasized is that, although the numbers of rural nurses educated to become practitioners may be few, their impact on the rural community is magnified many times over. For our program, the difficulties encountered putting this rural initiative in place have been more than compensated for by the enthusiasm of our students and faculty and the very positive support we have received from practitioners and clients in the rural community. As a result of the plan described, we now have the opportunity to reach into the rural community itself for local, qualified nurses committed to providing primary healthcare to their own neighbors. We encourage other programs to do the same. PMID- 11111562 TI - Developing low-cost clinical teaching tools. AB - Nursing is an art as well as a science. Using artistic resources can help the nurse educator develop creative and low-cost teaching tools that are truly effective and realistic. According to Bowman, these inexpensive teaching tools assist in obtaining and maintaining attention of students, stimulate discussion, encourage questions, and provide a means of skill development. PMID- 11111563 TI - Faculty ways of knowing: the crux of nursing curricula. AB - The authors, using the narrative form, examine three epistemological structures that are found in nursing curricula. Those fundamentally different approaches of knowledge acquisition are portrayed and discussed by the director and three staff members from a fictitious nursing school. Although faculty members naturally tend to adhere to their own embedded view of the way one learns, the authors stress the need for judicious blend of the three epistemic orientations when developing nursing curricula. PMID- 11111564 TI - Camp Can-Do. Outcomes of an experiential learning experience. AB - Because the most effective way to learn is by doing, faculty need to develop experiential learning experiences for students. The authors discuss one such experience, a multiple sclerosis camp where nursing students lived with and cared for their clients for 6 days. The camp experience illustrates how educators can partner with community organizations to the benefit of both. PMID- 11111565 TI - Benefits of cooperative learning groups when preparing for the NCLEX-RN examination. PMID- 11111566 TI - Developing a teaching portfolio in nursing education: a reflection. AB - Presenting teaching accomplishments through a portfolio promotes self-reflection and provides direction for improvement and advancement decisions. The authors examine experiences, insights, and reflections about their participation in a pilot project introducing teaching portfolios. Themes of taking stock, documenting practice, and reflecting emerge. Their experiences of completing teaching portfolios and personal growth as educators and colleagues are described. PMID- 11111567 TI - Stress management: a step-by-step process. PMID- 11111568 TI - Increasing students' cultural sensitivity. A step toward greater diversity in nursing. AB - Increasing minority representation in nursing is essential to assure culturally competent care in the next century. Academic institutions need to recruit, encourage, and retain minority students and faculty. Students need minority mentors in their academic and clinical environments. Euro-American faculties teaching in predominantly white institutions need to explore effective ways to facilitate aggregate understanding and appreciation of cultural diversity within the profession. The author explains a teaching strategy to increase awareness and sensitivity of graduate nursing students to the role of minority nurses within the culture of nursing. Becoming culturally sensitive is a prerequisite to increasing diversity and culturally competent care. PMID- 11111569 TI - Fostering holistic care in oncology nursing. PMID- 11111570 TI - A comparison of teaching strategies for integrating information technology into clinical nursing education. AB - As health care becomes more information-intensive and diverse, there is a need to integrate information technology (IT) into clinical education. Little is known, however, about how to design instructional strategies for integrating information technology into clinical nursing education. This article outlines the instructional strategies used by faculty in five nursing programs who taught students to use a point-of-care information technology system. The article also reports students' computer acceptance and summarizes IT clinical teaching recommendations. PMID- 11111571 TI - Developing an evaluation tool for instructional software programs. AB - The use of instructional software is predominant in many nursing programs. The need for cost-effective, quality programs requires faculty to evaluate instructional software before its purchase and use. The purpose of this work was to develop a tool for evaluating instructional software programs. The tool was based on data from the literature, as well as input and feedback from nurse educators and multimedia specialists. The evaluation tool assists faculty to ensure proper evaluation and selection of instructional software. PMID- 11111573 TI - Nursing across the rural lifecycle. PMID- 11111574 TI - Surfing lessons for nurses. PMID- 11111572 TI - Using standards of practice and key clinical points for teaching psychiatric- mental health nursing. AB - To optimize opportunities for clinical experiences in psychiatric-mental health nursing education, it is essential that the approach be directed to the student patient interaction. Clinical experiences, which rely on the assumption that spending time in a facility prepares students for practice, is unacceptable. One approach to correcting the problem is to develop comprehensive clinical guidelines that direct student experiences and assure mastery of identified outcomes. The authors focus on the use of key clinical points to direct student experiences in psychiatric-mental health nursing. PMID- 11111575 TI - Nurses always there for you. PMID- 11111576 TI - Embracing technology and caring. PMID- 11111577 TI - Beware of the legal implications of the Internet. PMID- 11111578 TI - A refugee camp without boundaries? PMID- 11111579 TI - Developing maternity services in Vietnam. PMID- 11111580 TI - Older people seek services that meet their needs. PMID- 11111581 TI - Providing new forums for experienced delegates. PMID- 11111582 TI - Nightingale's values live on. PMID- 11111583 TI - Internet resources for outcomes management PMID- 11111584 TI - Point-of-care database solutions for critical care: tapping the Internet to communicate critical information. PMID- 11111585 TI - Meta-analytic methods that support outcomes management. PMID- 11111586 TI - Service utilization and outcomes in rural home health agencies. AB - As home health reimbursement moves from fee-for-service to prospective payment, data describing the relationship between service utilization and patient outcomes will be the basis for planning services. The investigators measured the relationship between service utilization and generic patient outcomes for 1,704 home health episodes of care. Few significant relationships were found. The average study patient received 17 visits, well below the average number for the state and nation. Investigators suggested the possibility that visit numbers were too low to stimulate improvement in outcomes and that when services are curtailed, home health staff may do well to focus quality improvement efforts on condition-specific patient outcomes rather than generic outcomes. PMID- 11111587 TI - Survival analysis of three treatment modalities in a residential substance abuse program for women and children. AB - Retaining women in substance abuse treatment is difficult. Increased length of stay (LOS) has been found to be predictive of positive treatment outcomes, including lower drug use, criminal behavior, and unemployment among adults, and improved growth and development for children. This study used survival analysis statistical methods to examine LOS in a residential treatment program for women and children as the program shifted from a traditional therapeutic community, to gender-specific programming, to interdisciplinary, family-focused treatment. Results suggest that with implementation of family-focused treatment, LOS increases more than with gender-specific programming alone. PMID- 11111588 TI - Discussion of generic and disease-specific outcome tools for patients in cardiac rehabilitation. AB - Morbidity and mortality alone are not sufficient outcome measures in evaluating the effectiveness of interventions aimed at controlling or reducing complications of coronary heart disease. Outcomes are considered to be indicators of the quality of care. As a result, the selection of interventions, treatments, and reimbursement for services has become outcome driven. The purpose of this article is to discuss generic and disease-specific health-related quality of life tools and how to select appropriate tools for measuring long-term outcomes of cardiac rehabilitation. Choosing the appropriate tools to evaluate outcomes is critical for evaluation of patient progress and quality of life and for the success of cardiac rehabilitation programs. PMID- 11111589 TI - Relationship of cardiac inpatients' outcomes to mood state. AB - This descriptive study used a computerized charge capture system (CCCS) to explore the differences of cost and length of stay (LOS) between cardiac inpatients with a diagnosis of depression (n = 144) and cardiac inpatients without depression (n = 9,099). Level of severity, gender, and mood state (depression vs. nondepression) were also compared. A matched sample of 352 nondepressed patients was compared with a sample of 94 depressed patients. There were no significant differences between the depressed and nondepressed groups. However, the study did indicate interesting findings regarding mood state, gender, and cardiac outcomes. Depression was significantly overrepresented among females (chi 2 = 24.0, df = 1, P < 0.05). When gender and mood state were considered together, women with cardiac disease who were depressed had significantly longer lengths of stay (LOSs) and increased costs than men with depression (F = 6.6, df = 1, P = 0.01). A major unanticipated finding was the extremely low incidence of depression detected in these patients (1.6%) when compared with patients in other studies. One possible reason for the low incidence of depression was related to the use of a financial, rather than a clinical, data set. PMID- 11111590 TI - Satisfaction and adequacy of prenatal care utilization among rural low-income women. AB - This study was designed to describe adequacy and satisfaction with prenatal care in a group of rural low-income women (n = 60) and to determine whether either was correlated with birth outcomes. Despite less than adequate prenatal care in 50% of the women, they were satisfied with their care, and outcomes for infants were good. When compared with women who received adequate prenatal care, there were no differences between the two groups. Tailoring prenatal care to individual needs, including care provided by certified nurse midwives with fewer prenatal visits, could be cost-effective without sacrificing quality. It is time to reexamine the recommended prenatal visit structure and care delivery in this country. PMID- 11111591 TI - Age, developmental, and job stage influences on nurse outcomes. AB - This descriptive study surveyed 412 nurses in three hospitals and found that older nurses and nurses in more mature developmental stages showed greater job satisfaction, productivity, and organizational commitment. Job stages of entry, mastery, and disengagement were examined, and 24% of nurses reported being disengaged from their jobs, with lower satisfaction and commitment. Implications include the compelling need for nurses and organizations to do career planning together to avoid disengagement of nurses so critical to patient and organizational outcomes. PMID- 11111592 TI - Does your assessment include alternative therapies? PMID- 11111593 TI - Trauma and dissociation: treatment perspectives. AB - TOPIC: How advanced practice nurses can work with trauma survivors to decrease dissociation as a needed coping mechanism. PURPOSE: To review the literature on trauma and dissociation as well as current treatment perspectives. SOURCES: Review of the literature and authors' clinical experience. CONCLUSIONS: Advanced practice nurses can use knowledge of selected psychopharmacological medications and Erikson's stages of psychosocial development to plan treatment for posttrauma clients. PMID- 11111594 TI - Medusa appears: a case study of a narcissistic disturbance. AB - TOPIC: A detailed case study of the brief psychotherapy of a college student with a narcissistic disturbance, discussed through the lens of relational psychoanalytic theory and including a description of important interventions. PURPOSE: To promote greater understanding of psychoanalytic theory and therapy with a particular young adult population, and to underscore the importance of empathy and the therapeutic relationship to successful psychotherapeutic outcome. SOURCE: The author's own clinical practice. CONCLUSIONS: The therapeutic alliance is fundamental to the psychotherapeutic change process. Empathy is not just a means to a better healing relationship; it is in itself healing. Accurate empathy within the therapeutic relationship is essential to developmental unfolding, especially where sustained, age-appropriate emotional responses to developmental needs were lacking and an integrated sense of self failed to develop. PMID- 11111596 TI - Atypical antipsychotic drugs: Part 1. PMID- 11111595 TI - Veterans of abuse and daughters of the dark: the politics of naming and risk of transformation in building partnerships for change. PMID- 11111597 TI - Touch and psychotherapy. PMID- 11111598 TI - Mental health parity. PMID- 11111599 TI - Combination Interferon alfa-2b/ribavirin therapy for the treatment of hepatitis C: nursing implications. AB - An effective new therapeutic option consisting of Intron A (Interferon alfa-2b, recombinant; Schering Corporation, Kenilworth, NJ) Injection and Rebetol (Ribavirin, USP) Capsules is now available for the initial therapy of patients with hepatitis C and for patients who had previously responded to alpha interferon but subsequently relapsed. The combination of recombinant interferon alfa-2b/ribavirin therapy increases hepatitis C viral clearance 10-fold in hepatitis C relapse patients and almost threefold in previously untreated patients compared with alpha interferon monotherapy. There is no synergistic toxicity apparent with the two-drug combination. Ribavirin does not significantly worsen the side effects associated with interferon alfa-2b, which are predictable, manageable, and reversible. The major side effects of combination therapy include flulike symptoms, neutropenia, psychiatric disorders, and anemia; however, these side effects are well known and can be managed with dose modifications and nursing intervention. The assistance of nurses in patient education, in side effect management, in hematologic parameter monitoring, and in medication dosing and administration is crucial to maximizing patient compliance and therapy outcome. PMID- 11111600 TI - The effects of inflammatory bowel disease on adolescents. AB - Adolescence is a time of profound change for individuals. It is a period that witnesses biologic, social, and psychological change in the individual as well as role changes within the family and peer groups. Compounding these difficult transitions with the onset of inflammatory bowel disease leads to additional problems with adolescence adjustment. In the limited studies that have addressed the adolescent and inflammatory bowel disease, several key concepts emerge that point to ways for successfully dealing with these adolescent adjustment problems. Roy's Adaptation Model provides a foundation for identifying and selecting interventions in working with adolescents and chronic illness. A review of the literature finds the need for further study of the difficulties encountered by the changes of adolescence coupled with the onset of inflammatory bowel disease. PMID- 11111601 TI - Hepatitis C, E, F, G, and non-A-G. AB - Inflammation of the liver is known as hepatitis. Six or seven viruses, hepatitis viruses A through G, are responsible for most cases of viral hepatitis. In this second of a series of three articles, current knowledge regarding hepatitis C, E, F, G, and non-A-G is reviewed. Up to 80% of patients infected with hepatitis C progress to chronic liver disease. Hepatitis E is an enteric hepatitis that is endemic in Asia. Hepatitis F may be a mutant hepatitis C. Hepatitis G is a posttransfusion hepatitis. Non-A-G hepatitis consists of all of the hepatitis viruses awaiting identification. Up-to-date information regarding each of these types of hepatitis viruses is presented herein, because every phase of patient care improves when health care professionals are knowledgeable regarding the illnesses of their clients. PMID- 11111602 TI - Self-care interventions for the school-aged child with encopresis. AB - Encopresis, an elimination disorder in children, presents as a challenging problem for gastroenterology nurses working with patients and families confronted with this disorder. This article offers a summary of the literature on encopresis, including pathogenesis, causative factors, early treatment, and clinical interventions focused on self-care. The antecedent factors that facilitate the child's participation in self-care are summarized, along with the intended outcomes of the self-care intervention plan. PMID- 11111603 TI - A description of the gastroenterology nurse endoscopist role in the United States. AB - The use of nurse endoscopists in the specialty of gastroenterology has gained recent support in the United States. While studies using nurse endoscopists have documented positive patient outcomes, including cost effectiveness, public access to cancer screening, and patient satisfaction, research regarding the training and experiences of nurse endoscopists is almost nonexistent. This article presents findings from an exploratory, descriptive study of 17 gastroenterology nurse endoscopists in the United States. Study subjects describe their role as nurse endoscopists, their experiences, and their opinions about basic job and curriculum requirements for further development. These findings support the viability and future expansion of this advanced practice role in gastroenterology nursing. PMID- 11111604 TI - Performance of flexible sigmoidoscopy by registered nurses for the purpose of colorectal cancer screening. SGNA guideline. Society of Gastroenterology Nurses and Associates. PMID- 11111605 TI - Aciphex (rabeprazole). PMID- 11111606 TI - Intervention studies to reduce the prevalence and incidence of pressure sores: a literature review. AB - Much has been written about the prevention of pressure sores. However, electronic and manual searches located only 10 studies within the literature in the UK that described interventions able to reduce either their incidence or prevalence. All the studies located contained serious methodological flaws. Apparent success in reducing the number or severity of pressure sores could have resulted because staff involved in data collection were aware that the study was being undertaken and thus took more interest in pressure area care. From the review findings it is apparent that there is a dearth of research evidence upon which to base practice in the sphere of pressure sore prevention and further research is urgently required. PMID- 11111607 TI - Prevalence of urinary incontinence and its relationship with health status. AB - A postal survey of two random samples of adult populations within two health authorities in the UK was undertaken during 1994. One health authority had an established continence advisory service (HA1) and the other one did not have a continence advisory service (HA2). A total of 12,529 patients (HA1, 6319; HA2, 6210) were mailed a structured questionnaire and 53% (n = 6139) returned completed questionnaires. A point prevalence of current urinary incontinence of 9% (n = 519, 95% CI, or confidence interval, from 7.9% to 9.3%) was found. A large number of people within the populations had experienced urinary incontinence at some time during their adult years (23%, n = 1427, 95% CI from 22.2% to 24.3%). People who were incontinent had a significantly lower health status than people who were continent (mean scores across all eight domains of the Short Form 36, SF36, P < 0.0001), indicative of greater health and social care needs. The prevalence of urinary incontinence in the adult populations of two communities indicates that it is a sizeable public health and primary healthcare issue. PMID- 11111608 TI - Information point: prevalence and incidence. PMID- 11111609 TI - Evaluating support surfaces for patients in transit through the accident and emergency department. AB - Little attention has been given to pressure area care for patients admitted via the accident and emergency department (A & E) in UK hospitals, despite evidence that they may wait for considerable periods on hard surfaces, placing them at risk of tissue damage. The literature was searched, and in the absence of existing guidelines to evaluate the suitability of the standard hospital trolley for use with emergency admission patients, criteria were developed by consensus among stakeholders with relevant expertise who were employed in an acute NHS hospital trust. Audit of the existing patient support surfaces using these criteria revealed deficiencies in all of them. Some of the problems identified were related to deterioration of the equipment; others were related to its design. The criteria were used to inform the purchase of new equipment, and a system of auditing to improve pressure area care for emergency admission patients has been established in the trust. The initiative has also drawn attention to significant omissions in the literature relating to pressure area care, including the need to evaluate patient comfort and inclusion of consumer views. PMID- 11111611 TI - Information point: type I and type II errors. PMID- 11111610 TI - A randomized trial of a Telephone Reassurance Programme for patients recently discharged from an ophthalmic unit. AB - Patients often experience problems after discharge, for instance with housekeeping or a general lack of information. The effect of a nurse-initiated Telephone Reassurance Programme (TRP) on ophthalmic patient outcomes was investigated. Patients in the intervention group were phoned by a nurse 3-6 days after being randomized and discharged home. Patients in both intervention and control groups received a questionnaire 1 week and 1 month after discharge to assess the patient outcomes 'Informational needs', 'Uncertainty', 'Emotional complaints' and 'Functional limitations'. In an attempt to explain the lack of statistically significant results, the limitations related to the participants, intervention and outcomes are discussed. PMID- 11111612 TI - The development of a national guideline on the management of leg ulcers. AB - This paper describes the development of an evidence-linked clinical guideline for the management of uncomplicated venous leg ulcers. Guidelines are developed to provide recommendations for clinical practice which are based on summaries of good quality research evidence. The aim of the guideline discussed in this article is to direct primary health care practitioners to the most effective method of assessment and treatment of venous leg ulcers and to discourage practices that do not have convincing or sufficient evidence of effectiveness. The three most important steps to the development of a valid clinical guideline are: basing recommendations on the best available evidence; explicit linkage between guideline recommendations and quality of evidence; and the involvement of a multidisciplinary group. The steps are discussed in relation to the development of the guideline alongside an introductory presentation on the role guidelines can play in improving practice. Issues arising from guideline development such as valid ways of obtaining patient input and lack of evidence are also discussed. PMID- 11111613 TI - Individualized care in a Finnish healthcare organization. AB - This paper reports the findings of a study exploring the provision of individualized care in a regional hospital in Finland. Individualized care was defined in terms of how patient individuality was taken into account and how patient participation in decision-making was facilitated. The data were collected from hospitalized patients (n = 203) using a questionnaire specially developed for this study. The response rate was 89%. A strong support to facilitate patient participation in decision-making was reported. Most shortcomings concerning the provision of individualized care related to consideration of the patient's cultural background and the involvement of the patient's family in the planning of care. Patients' age and the type of ward were associated with the provision of individualized care: patients aged 65 or over were more satisfied than younger age groups with the way in which their personal life situation had been taken into account. Patients from the gynaecological ward thought, more than patients from the surgical ward, that their situation had been taken into account during hospitalization. PMID- 11111614 TI - Assessment and documentation of bowel care management in palliative care: incorporating patient preferences into the care regimen. AB - Nurses in a palliative care unit (PCU) recognized that there were several inconsistencies relating to assessment and documentation of patient preferences in bowel care management. Although bowel care is recognized as of key importance to the wellbeing of palliative care patients, there is little evidence in current literature about accommodation of patient preferences in bowel care management. A questionnaire was developed to assess whether patient preferences were elicited on admission to the PCU, were documented, and were included in the bowel care regimen. Data were collected from 100 patients in two PCUs in Australia. The findings suggested that little was assessed or documented about bowel care management on admission except functional or pharmacological information. According to patients in the study, their preferences were seldom incorporated into the bowel care regimen. Lack of documentation of bowel care preferences was also found following an audit of patient notes. Techniques for eliciting information, awareness of alternative or complementary methods of bowel care and better documentation procedures are all recommended for inclusion in nursing practice in the palliative care setting. PMID- 11111615 TI - Patients with operable oesophageal cancer: their experience of information-giving in a regional thoracic unit. AB - A qualitative descriptive study was developed to gain an insight into the experiences of patients with operable cancer of the oesophagus and the information they received. Through semistructured interviews, participants related details about different aspects of information-giving. Key sources of information were the consultant surgeon and other patients. Nurses, other medical staff, physiotherapists and dieticians were then mentioned. Family/friends were poor sources. Positive and negative aspects of the verbal and written information given were described, but there was minimal support for audiovisual information. Participants were given information relating to a number of key areas: treatment details, side-effects, extent of their disease, cure and prognosis and return to normality. A number of problem areas were identified. The findings indicate a need for a thoracic nurse specialist, updating of the information booklet and development of a staff education programme. PMID- 11111616 TI - Men living with diabetes: minimizing the intrusiveness of the disease. AB - In this paper we present the findings from the second of four Participatory Action Research (PAR) groups with men and women who have been diagnosed with type two diabetes. The findings of the men's group are reported here. People who have received a diagnosis of diabetes must immediately absorb a great deal of information about how to control their diabetes, care for themselves and make lifestyle changes. In this study, we have asked men about this transition and about what it is like to live with diabetes. We aimed to understand how people with type two diabetes incorporate chronic illness into their lives. Utilizing the processes of PAR, we created a conducive environment for the voices of people with diabetes to be clearly heard in relation to their health. Men who live with type two diabetes met with a researcher and two Clinical Nurse Consultants, for two hours, once a week, for four weeks, during November 1998. The men expressed that diabetes had made a positive impact on their lifestyle; they viewed diabetes as part of life and not as an illness. Men chose foods with confidence; their concern about potential complications meant they chose to take better care of themselves. They were confident in their knowledge of diabetes, and while they took responsibility for themselves, being supported by their partner was helpful in managing their diabetes. They managed their life with diabetes by minimizing the intrusiveness of the disease. PMID- 11111617 TI - How people with stroke and healthy older people experience the eating process. AB - The aim of this study was to describe the process of eating, experiences of eating and oral functions. Participants consisted of 30 people with first stroke and localization of the damage verified by computer topography (CT), and 15 healthy older people. All were observed during test-meals, interviewed about eating, and oral functions were tested. The results demonstrated that most (21) people with stroke had some difficulties in eating and expressed feelings of fear and shame about eating and changed physical and social appearance, mainly related to difficulties in preparing and transporting food to the mouth as well as swallowing deficits. PMID- 11111618 TI - Relationships between partner's support during labour and maternal outcomes. AB - The objective of this study was to measure the relationship between women's ratings of partners' participation during labour and maternal outcomes as measured by anxiety level, pain perception, dosage of pain-relieving drug used and length of labour. A convenience sample of 45 primigravid women was selected from the postpartum unit of a public hospital in Hong Kong. They were all first time Chinese mothers, aged 18 or over, who had attended antenatal classes and had their partners present during labour. The State Scale of the State-Trait Anxiety Inventory was used to measure maternal anxiety during labour. Labour pain was measured by the Visual Analogue Scale. A series of scales were developed to measure partners' participation during labour. Women's ratings of partners' practical support were significantly lower than their ratings of partners' emotional support. There were no significant associations between level of emotional support and maternal outcome measures. However, perceived practical support was positively related to the dosage of pain-relieving drug used and total length of labour. Positive relationships between the duration of partners' presence and women's ratings of perceived support provided by partners during labour were also found. PMID- 11111619 TI - Peer evaluation in nurses' professional development: a pilot study to investigate the issues. AB - Peer evaluation in nursing is a method by which the nurse evaluates the work of a peer, according to set evaluation criteria. The aim of the study was to clarify the potential significance of peer evaluation with regard to nurses' career development and relates to the introduction of a career development programme for nurses in a Finnish University Hospital. The research concepts were created on the basis of literature analysis. The concepts served as a basis for data collection, and five open-ended questions were devised from them. Informants (n = 24) gave free-form essay-type answers to these questions. The material was analysed using qualitative content analysis. The results indicate that self evaluation constitutes the basis for peer evaluation. Peer evaluation allows nurses to give and receive professional and personal support promoting professional development. Professional support offers possibilities for change and alternative action. Personal support requires respect for the peer's equality and individuality. Personal peer support can decrease feelings of uncertainty and insecurity caused by work. The conclusion is drawn that peer evaluation is a means of promoting nurses' professional development to further on-the-job learning in collaboration with peers. PMID- 11111620 TI - The clinical nurse specialist: perceptions of practising CNSs of their role and development needs. AB - This study was carried out to determine the role and development needs of Clinical Nurse Specialists (CNSs) in two acute hospital trusts. One of the aims was to obtain a detailed understanding of the current professional experience of Clinical Nurse Specialists. Data were collected using focus groups and analysed descriptively. Analysis of the content was qualitative, based on a grounded theory approach, and followed the initial steps of open and axial coding. Role transition and development needs are identified on a continuum of novice to expert. PMID- 11111621 TI - Caring in nursing: perceptions of Hong Kong nurses. AB - Ten registered nurses in Hong Kong were interviewed on their perceptions of caring behaviours in their clinical settings, barriers to these behaviours, and possible ways to overcome those barriers. Findings showed that respondents valued the importance of expressive behaviours and interpersonal communication skills in providing holistic patient care. They felt constrained by social, economic, cultural, and personal variables such as staff shortages, the traditional task orientated approach of nursing, the dominance of medicine in the healthcare system, the influence of Chinese culture on work attitudes, and their limited skills and lack of education. The situation could be improved through better staff education, colleague support, effective human resources allocation, and promotion of a democratic working environment. A creative approach is necessary to integrate these strategies into a healthcare system dominated by technology and economic constraints. PMID- 11111622 TI - Stress amongst district nurses: a preliminary investigation. AB - This paper presents the results of a pilot study investigating stress among district nurses in the north-west of England. Nurses completed questionnaires covering job satisfaction, mental health, stress, Type A behaviour, health behavior, coping skills and demographic details. A specific measure of stress was developed following in-depth interviews with primary care professionals, including district nurses. A total of 79 district nurses took part in the study. The major sources of stress isolated by the district nurses related to: time pressure, administrative responsibility, having too much to do, factors not under their control, interruptions, keeping up with National Health Service (NHS) changes, and lack of resources. Factor analysis of stress questionnaire responses identified five major factors: demands of the job and lack of communication, working environment, problems with patients, work/home interface and social life, and career development. The highest levels of satisfaction were reported for the amount of variety in their job and the lowest level of job satisfaction was reported for chance of promotion. The results revealed that the mental wellbeing of the nurses was higher than that of the other population groups. Furthermore, multivariate analysis revealed three major stressors that were predictive of high levels of job dissatisfaction: demands of the job and lack of communication, working environment, and career development. The implications of the findings for further research are considered. PMID- 11111623 TI - Feminist research or humanistic research? Experiences of studying prostatectomy. AB - This paper highlights issues related to men's health research arising from a small-scale study, carried out by a male researcher, to identify the experience of men following transuretheral resection of prostate (TURP) for benign prostatic hypertrophy (BPH). The intention of this paper is to stimulate methodological debate rather than to be a research report. For the study, an informal interview approach was used within a phenomenological framework, and interview experiences raised issues which have been previously discussed under the rubric of feminist research. The conclusion drawn is that a style of research which attempts to gain a holistic view of patients' experiences is better termed 'humanistic research' because the term 'feminist research' clearly cannot be applied to men studying men's health-related experiences. PMID- 11111624 TI - Exercise and older adults. PMID- 11111625 TI - Assessing pain in older adults. PMID- 11111626 TI - Functional performance and exercise of older adults in long-term care settings. AB - Performing functional activities and exercising are important for older adults living in long-term care settings. Participation in these activities not only improves and maintains function in older adults but also can improve physical and emotional health and quality of life. The purpose of this study was to explore the variables that influence functional performance and exercise activity in a group of nursing home residents. Participants included 59 residents in a long term care facility. The mean age of participants was 88 +/- 6.9, and the majority were women (76%), White (97%), and unmarried (76%). Residents participated in a single face-to-face interview. Chart reviews for demographic and health information also were performed. Based on stepwise multiple regression analyses, upper and lower extremity contractures and cognitive status were the only variables that significantly influenced functional performance and accounted for 49% of the variance in function. Self-efficacy and outcome expectations were the only variables to significantly influence exercise behavior and accounted for 57% of the variance in this behavior. These findings can be used to help develop and implement effective restorative nursing care programs in long-term care settings. PMID- 11111627 TI - Exercise and quality of life in elderly individuals. AB - 1. Exercise is important and recommended for older adults. Nurses in community settings are in ideal positions to facilitate older adults' use of exercise programs. 2. Quality of life is complex and multidimensional. Dimensions include well-being, functional status, socioeconomic status, and self-esteem. 3. This article contains a review of empirical evidence that states older adults who exercise have improved quality of life. PMID- 11111628 TI - Changing health behaviors of older adults. AB - 1. Changing lifelong unhealthy habits can have a positive effect on health for older adults. 2. The Transtheoretical Model of behavior change proposes people move through a series of five stages and use a variety of processes as they attempt to change a behavior. 3. Research has shown that tailoring interventions to a individual's stage of change is most effective in promoting behavior change. 4. Specific stage-based strategies are recommended for nurses to use with both individuals and groups of older adults. PMID- 11111629 TI - Understanding what motivates older adults to exercise. AB - The purposes of this study were to explore the factors that influenced adherence to an exercise program for older adults, and compare differences in motivation, efficacy expectations, health status, age, functional performance, and falls between adherers and nonadherers. A combined qualitative and quantitative design was used. Participants included 23 of the 24 members of an existing walking group, with an average age of 81 +/- 7.2 years. Fourteen (60%) participants did not adhere to walking, while 9 (40%) adhered. Those that adhered had fewer functional limitations due to their health, (F = 7.7, p < .05), better functional performance (F = 4.0, p < .05), stronger self-efficacy expectations related to exercise (F = 4.3, p < .05), and fewer falls (F = 4.4, p < .05). Six major themes were identified that impacted adherence: a) beliefs about exercise; b) benefits of exercise; c) past experiences with exercise; d) goals; e) personality; and f) unpleasant sensations associated with exercise. Interventions that focus on teaching older adults about the benefits of exercise, establishing appropriate goals, and decreasing unpleasant and increasing pleasant sensations associated with exercise may be useful to improve adherence to a regular exercise program. PMID- 11111631 TI - A new century: a new vision PMID- 11111630 TI - A fall prevention program for elderly individuals. Exercise in long-term care settings. AB - The purpose of this research was to explore the role of exercise in preventing falls, specifically assessing the effectiveness of an ankle strengthening and walking program to improve balance, ankle strength, walking speed, and falls efficacy and to decrease falls and subjects' fear of falling. Sixteen individuals participated in the study which was conducted at two nursing homes. Subjects were assigned randomly to an intervention or control group. The participants in the intervention group completed a 3-month supervised program of ankle strengthening exercises and walking. Descriptive statistics were used to characterize the sample, and differences in the least square means were used to assess the outcome variables (i.e., balance, ankle strength, walking speed, falls, fear of falling, falls efficacy) before the exercise program, and again at 3 months and 6 months after the program for the intervention and control subjects. Findings for the intervention group from pretest to 3-month posttest were, for the most part, maintained or in the predicted direction, suggesting that regular exercise shows promise for preventing deterioration and improving fall-related outcomes for elderly nursing home residents. PMID- 11111632 TI - An evaluation of knowledge and skill retention following an in-house advanced life support course. AB - The aim of the study was to explore advanced life support (ALS) skills and knowledge retention, in 10 registered nurses, at 6 and 12 weeks following an in house core ALS course. Data was collected by structured interview and structured observation at 6 and 12 weeks following the course. A sharp decline in the number of subjects attaining the 84% pass level at the 6 and 12 week interviews was noted. A modest increase in ALS theoretical knowledge and a decrease in practical skill performance between 6 and 12 weeks was identified. Implications for future practice and research are proposed. PMID- 11111633 TI - Medical consultants' attitudes towards nurse assessment and initiation of thrombolysis prior to medical screening. AB - This study explores medical consultants' attitudes to the concept and introduction of nurse-initiated thrombolysis within a coronary care unit. A self administered questionnaire was used to gauge attitudes of consultants who care for patients admitted to coronary care. Themes that emerged were: conceptual awareness, medico-legal aspects and professional preparedness. Further research is advocated to explore the attitudes and experiences of nurses and physicians involved in the practice of nurse-initiated thrombolysis. PMID- 11111634 TI - Reflection on diagnosing as a nurse in ITU. AB - Brookfield's (1995) four critically reflective lenses are adapted from teaching and applied to nursing within this article. The lenses are used as a framework for exploring the author's feelings associated with diagnosing a life-threatening problem in intensive care in the presence of a disagreeing doctor. The issues surrounding the distinctions between medicine and nursing are explored. Wellington and Austin's (1996) five orientations to reflection provide the framework for the author's 'metacognition'--his thoughts about his thoughts. These are communicated by way of a critical commentary. PMID- 11111635 TI - Beyond diagnosis. PMID- 11111636 TI - Health care support workers in the critical care setting. AB - The 1999/2000 winter demands on the NHS have once again highlighted deficits in UK critical care provision (Daily Telegraph, 2000; London Evening Standard, 2000) Recent years have seen the development of the role of health care support workers in the NHS; some critical care units now employ health care support workers This research examined the views of critical care unit staff on the introduction of health care support workers into the critical care unit It is concluded that the role is viable within the setting of this study A framework is outlined that could form the basis for a critical care health care support worker training programme PMID- 11111637 TI - Liver failure in the critically ill. AB - The pathophysiology and manifestations of liver failure and fibrotic liver disease are discussed The management of patients with liver dysfunction is reviewed Indications for transplant and new modes of liver support are outlined PMID- 11111638 TI - The impact of the promised spending increases for the NHS. PMID- 11111639 TI - The provision for children in the acute independent sector. PMID- 11111640 TI - Managing and leading psychiatric nursing. Part. 2. Basic and continuing education. PMID- 11111641 TI - Culture, religion and patient care. PMID- 11111642 TI - Access to all areas. PMID- 11111643 TI - Managers are made not born. PMID- 11111644 TI - Own goals, how to measure effectiveness. PMID- 11111646 TI - Shaping the future of nursing. PMID- 11111645 TI - All things to all managers. PMID- 11111647 TI - Careering up the ladders. PMID- 11111648 TI - Every nurse is a leader. PMID- 11111649 TI - Documenting a patient's fall risk. PMID- 11111650 TI - Taking a team approach to back pain. PMID- 11111651 TI - Applying a two-piece cervical collar. PMID- 11111652 TI - Action stat. Open fracture. PMID- 11111653 TI - Saying a mouthful about oral diabetes drugs. PMID- 11111654 TI - Lessons learned about insulin therapy. PMID- 11111655 TI - The face in the mirror. PMID- 11111656 TI - Changing an ostomy appliance. PMID- 11111657 TI - Telephone triage: SAVED by the form. PMID- 11111658 TI - Test yourself. The cardiovascular system. PMID- 11111659 TI - Selecting a vascular access device. PMID- 11111660 TI - A healing journey. PMID- 11111661 TI - Going to the dogs ... for help. PMID- 11111662 TI - Understanding hypokalemia. PMID- 11111663 TI - Under the influence? PMID- 11111665 TI - Myths & facts ... about hospice. PMID- 11111666 TI - Managing a diabetic foot ulcer. PMID- 11111664 TI - Handling adverse event reports. PMID- 11111667 TI - Amphotericin B reaction. PMID- 11111668 TI - Looking at lumbar puncture in adults. PMID- 11111669 TI - Little amigo. PMID- 11111670 TI - Varicella-zoster virus. PMID- 11111671 TI - The nurse who gave us candy. PMID- 11111672 TI - [Education by participating in a diabetic food contest]. AB - In order to investigate new methods to use to educate the general public and to foster changes in diet, a cooking course was organized by the "Las Fuentes Norte" Health Center in Zaragoza open to the entire population. 42.1% of the menus selected were prepared by diabetics. In these, their average composition was: 488.3 +/- 114 calories containing 30.7% proteins, 38.8% lipids, and 32.5% carbohydrates. 42.1% of the menus selected were prepared by people in families having no diabetics. In these, their average composition was: 332.8 +/- 114.4 calories containing 25% proteins, 29.9% lipids, and 50.1% carbohydrates. The remaining 15.8% of the menus selected were prepared by relatives of diabetics. In these, their average composition was: 528.1 +/- 237.9 calories containing 40.6% proteins, 45.3% lipids, and 14.1% carbohydrates. There was no finding bearing statistical importance among these groups which prepared these dishes. Analysis of the 1998 contest discovered that calories increased in the menus prepared by diabetics or their relatives; calories increased 101.1 and 330.7 respectively, and this bears statistical importance. In these same groups, lipids increased 11.2 and 25% respectively while carbohydrates dropped 11.2 and 29.6% respectively. In those menus prepared by persons having no diabetics in their families, lipids decreased 4.3% while carbohydrates increased 15.9%. The average number of calories in the 19 menus prepared was 1758.8 +/- 20.7. These menus contained an average of 20.8% proteins, 25.5% lipids, and 53.7% carbohydrates. PMID- 11111673 TI - [Magnetic resonance, an introduction]. AB - What would you explain to a patient if he/she had to undergo a magnetic resonance imagery session? Do you know if a person wearing a pacemaker can undergo an MRI? These and many other questions are answered in the following article since magnetic resonance imagery is a very useful diagnostic medium; however, it is one which not everyone has been able to get to know and use. The authors shed light on this diagnostic technique for nurses starting with its physical foundations; since knowing these aids professionals to correctly plan our treatments and improves the attention provided to patients who undergo this test. The authors also list the specific components in this device, the possible biological effects, the detractions and some basic recommendations. PMID- 11111674 TI - [Popliteal, posterior tibial and peroneal arteries]. PMID- 11111675 TI - [Differences in blood pressure: does the length of time between measurements have an effect?]. AB - Taking blood pressure measurements correctly is essential in order to successfully evaluate a patient's blood pressure from the initial measurement and subsequent measurements over time. With the purpose to determine if the time between blood pressure measurements in an office or clinic bears an influence on any variation in measurement, a study was carried out under which two blood pressure measurements five minutes apart were taken using an automatic instrument in order to evaluate the existence of any differences as well as to identify which arm should be the control arm and to evaluate the absolute measurements between both arms. During this study, the authors observed a decrease in the mean for the repeated blood pressure measurement of 7.7 mmHg (IC 95%: 4.7-10.7 mmHg, p < 0.0001) for the systolic measurement and of 2.9 mmHg (IC 95%: 1.6-4.2 mmHg) for the diastolic measurement. 78% of the patients in this study registered a difference between the two measurements, either systolic or diastolic > or = 5 mmHg. It was not possible to establish a significant relationship between the decrease in these blood pressure measurements and factors such as age, sex, cardiac frequency or hypertension. The control arm was the right arm in 59.3% of the patients. The absolute mean of the differences between both arms was 8.4 mmHg (IC 95%: 6.8-10 mmHg) for the systolic measurements and 5 mmHg (IC 95%: 3.9-6.1 mmHg) for the diastolic measurements. 26 patients, 44.1%, registered a difference in pressure between their arms > or = 10 mmHg. PMID- 11111676 TI - [I was a graduate of the second course of nursing at the University of Alicante]. PMID- 11111677 TI - [Our school is the first to introduce the secondary nursing course with its specific title. Interview by C. Solano and F. Vizcaya]. PMID- 11111678 TI - [Complementary therapies. Phytotherapy]. AB - This article provides insight into the concept of phytotherapy, analyzing the generic term from a pharmaceutical viewpoint and explaining what the "totum" of a plant is. This article also mentions the role of phytotherapy over the course of medical history, what the correct criteria are for preparing a medicinal drink, bearing in mind very practical aspects for all of us, such as: the number of plants to include in a medicinal drink, the manner to prepare one, dosage, toxicity, effectiveness, time period to drink one over, etc. This article is of interest to health professionals and to all those who are drawn to the medicinal, healthy power plants possess. PMID- 11111679 TI - [Antiseptic agents: chlorhexidine]. AB - Chlorhexidine is a broad spectrum antiseptic widely used in clinical practice. This antiseptic works rapidly and its effects last for six hours. Since it is not absorbed through the skin nor through mucus, its systematic toxicity is minimum. It keeps on working in contact with organic matter and, since it is transparent, it does not hide the evolution of injuries. In this article, the authors review the properties and indications for this antiseptic; they also comment on some studies having lesser known indications. PMID- 11111680 TI - [Studies of the personality of nursing students]. AB - This study evaluates diverse psychological variables in 35 third-year nursing students. Personality is studied according to Eysenck's theories, by means of the E.P.Q. questionnaire; assertive behavior is studied by means of the Gambrill and Richey assertiveness questionnaire; and the hostility set is also measured by means of the Buss-Durkee questionnaire. Likewise, due to the results obtained, the authors point out the necessity to work on the theme of assertiveness in nursing students. PMID- 11111682 TI - [Sexual relations after a myocardial infarction]. AB - After an acute miocardiac infarction, sexual disorders may appear in a high percentage of patients. This article analyzes the problems which can lead to a sexual dysfunction, and presents a series of recommendations so that a nurse may explain these to the patient. If the patient possesses sufficient clear information, he/she will be able to overcome the inconveniences which occur with greater ease and thus be able to lead a fuller, happier life. PMID- 11111681 TI - [Systems for the determination of body temperature]. AB - It is essential for a health care professional to know how body temperature is regulated, what factors affect body temperature, and what different measurement systems are available. This file contains descriptions of 1) the thermometers available: electronic, mercury-based, infrared; 2) the places where body temperature can be measured: skin, armpits, rectum, mouth, tympanum, esophagus, or pulmonary artery blood. All content material is justified by a documented bibliography. PMID- 11111683 TI - [A narrative about the world of those who deliver and those who receivecare]. AB - The purpose of the present study is to show the narrative as a methodology that enables the learning about the other's apparent and hidden world. The other may be a client or a member of the nursing team. At first, authors present their general view showing the narrative as a way of reporting the exceptional, the drama experienced by a group, and a form of acting upon a drama situation. Subsequently, authors who use the narrative to represent diseases are presented. Finally, authors show the narrative as a possibility offered to nursing to interpret and act upon its practice. PMID- 11111684 TI - [Iatrogenic nursing care: dealing with danger in everyday nursing]. AB - The study presents the conceptions and experiences of a faculty nurse, built on his quotidian of providing car and, showing, in this context, nursing iatrogenic care. Through the aesthetic language, the author presents reflections and some references of authors who work on the quotidian, characterizing nursing care and, mainly, nursing iatrogenic care within the image of modern and post-modern paradigms. PMID- 11111685 TI - [The pain of burns: terrible for him who feels it, stressful for those who care for him]. AB - The goals of the study were to understand the cultural meaning that the nursing team and burn patients attribute to burn pain. Participant observations were carried out during one year at a Burn Unit. Semi-structured interviews with four nurses, five nursing auxiliaries and 12 patients were recorded. The burn pain is understood by nurses as physical and emotional pain. For professionals and patients, bath and dressing changes are stressful. For patients, burn pain is terrible and they can not explain it. However, they feel they should hold it up. PMID- 11111686 TI - [Development of an alarm system for the prevention of falls in hospitalized patients]. AB - The present study describes the analysis of an online patient assessment system developed to prevent inpatients falls. A chart review was performed in order to identify risk factors present in the Nursing Assessment tool. The identified variables were Functional status including walking, transferring and toilet, sex and patients self-care ability. Authors developed a system using these variables, that is available since February 1997. In a second phase of this study, authors aim to evaluate the effectiveness of this computerized intervention. PMID- 11111687 TI - [Nature and classification of nursing interventions in an adult outpatient clinic for chemotherapy]. AB - The purpose of the present descriptive study was to identify, through a retrospective analysis of records, the nature of nursing interventions as well as to establish the relationship between the nature of these interventions and the problems found. The sample was formed by 184 records of patients in their first nursing consultation at the Adult Chemotherapy Outpatient Unit of Hospital Sao Paulo from January to June, 1997; 37 records were selected at random: 19 male and 18 female patients. Results showed the predominance of interventions centered in patients' psychobiological needs. Educative interventions were more frequently performed when related to patients psychosocial needs. The correspondence between the identified problems and proposed interventions was evidenced when the psychobiological needs were met. PMID- 11111688 TI - [Wearing a pouch shows the difference between "being an "ostomized person" and "being a professional": analysis of a teaching strategy]. AB - This study aimed at analyzing nurses (re)construction process regarding the meanings of ostomy, ostomized patients, nursing care and the professional role after they wore an ostomy appliance. This activity is a teaching strategy applied at the Enterostomal Therapy Nursing Education Program. Two major units were identified after the analysis of students speeches: "being an ostomized person" and "being a professional". When students wore the pouches they felt as ostomized patients experiencing a violation of their identity and quality of life that promotes profound changes in daily life. These symbolic contents cause a crisis regarding the meaning of being a professional as until this experience, nursing care seemed fragmented to them. Recognizing a previous care performed as a technical action mainly directed to the ostomy and the pouch, students were able to visualize a future care, that is holistic and considers the ostomized human being, enabling them to incorporate affective, symbolic and relational dimensions. PMID- 11111689 TI - [Insulin self administration in children with diabetes mellitus, type 1]. AB - The present descriptive study had the goals of characterizing type 1 diabetic children, according to socio-demographic variables and identifying the difficulties related to insulin self-management and home control. 34 type 1 diabetic children were interviewed at a big hospital. Results showed that 82.4% of children were white, 61.8% were female and 54.1% were from nine to eleven years of age, 67.7% were catholic, and 64% had the illness for 3 years. 35.3% of them learned insulin management with their mothers and 32.3% follow a schedule regarding insulin self-administration. Difficulties to perform home control are related to the available resources and lack of information. Results show the need for a planned work integrated by a multiprofessional team and directed to the children whose characteristics meet the mentioned aspects, considering their interdependence and aiming at achieving a successful career. PMID- 11111690 TI - [Evaluation of the rooming-in system at the D. Francisca Cintra Silva Maternity Department, Santa Casa Hospital at the Sao Carlos municipality]. AB - The rooming-in was implemented at a maternity at the municipality of Sao Carlos in 1997. This study analyzes the acceptance of the system by mothers and how nursing participates in this work. Results showed that mothers are satisfied. However, the nursing team does not agree with this evaluation, confirming the need of increasing the number of workers as well as qualifying them. PMID- 11111691 TI - [Neonatal mortality in 1998 in the municipality of Botucatu]. AB - Considering that neonatal mortality is an indicator of the quality of the care provided to pregnant women, at childbirth as well as to the new born, authors developed the present study, whose aim was to analyze the neonatal mortality during the year of 1998 at the municipality Botucatu-SP. The coefficient of neonatal mortality was of 8.3/1000 born alive and the coefficient of precocious neonatal mortality was of 7.3/1000 born alive, confirming the importance of decease in the first week of life. Results showed that approximately 3/4 of the deceases can be reduced through precocious diagnosis and treatment as well as adequate care to birth or partially reduced through appropriate pregnancy control measures, evidencing that in order to decrease the rates of neonatal death, investments must be made to improve the quality of the care to pregnant women, parturients and the new born. PMID- 11111692 TI - [Sexual problems experienced by women in HIV-1 crisis]. AB - This descriptive study was based on the assumptions of qualitative investigation methods and on the Crisis Theory formulated by CAPLAN (1966). The objective of the study was to identify the sexual problems experienced by women undergoing an HIV-1 crisis. Data were collected through a semi-structured interview recorded by the method of FREITAS et al. (1992) and analyzed by the method of MEIHY (1996). Authors concluded that these women maintained themselves unbalanced using negative coping mechanisms. Thus, authors believe that it would be possible to help them using adequate nursing actions such as educational and guidance actions, but mainly by actions directed to the needs of patients undergoing an HIV-1 crisis. PMID- 11111693 TI - [Female sexuality: understanding its significance]. AB - The present qualitative study with a phenomenological focus aimed at understanding the meaning of female sexuality to women who participate in the activities of an orientation group about this subject, organized by a family planning service. Through the analysis of the participants speeches and phenomenological reduction, authors found the central subject: Living sexuality. Results allowed authors to learn a little more on this subject, especially considering that women who took part in this group aimed at solving their problem as they believed they had sexual disorders. However, this fact was demystified, enabling them to understand that they have problems in living their sexuality. PMID- 11111694 TI - [Learning leadership: nurses opinion about their academic education]. AB - The purpose of the present study was to identify nurses opinion about aspects of their academic undergraduate education directed to form leaders of the nursing team. Therefore, authors interviewed 17 nurses from a public federal hospital who experience the condition of being staff leaders. The collected material was analyzed through the technique of content analysis. Results evidenced a dissatisfaction concerning their academic training on leadership. The approached aspects showed the importance of a reflection about the need of teaching-learning strategies that allow students to develop leadership skills. PMID- 11111695 TI - [Group work: reflections on the daily work, report on an experience]. AB - The purpose of the present study was to analyze the meaning of the care delivered by a group of professionals at a basic health unit, in order to propose nursing actions that are not linked to medical actions. As theoretical references, authors used ALMEIDA & ROCHA(1997) and the ideas of CAMPOS (1992) and MERHY(1995). Regarding the group activities, authors used MUNARI & RODRIGUES(1997) and LOPES & MANZOLI(1996) considerations. From August 1995 to December 1997, authors attended 108 clients that spontaneously asked for nursing orientation in order to loose weight. All those showed significant changes in theirs lives and 85% lost weight. Nowadays, the group is recognized by the clientele and by the health team. PMID- 11111696 TI - [Nurses' work in Peru: origin and social conditions]. AB - The study aimed at reviewing the history of the genesis and social formation of nurses work in Peru in order to understand it according to women's social situation in the context of a masculine society organized under capitalism. Thus, nursing formation was influenced by the Nightingale School and the exercise of the Peruvian nurses was characterized by the reproduction of social functions that are historically feminine, with a knowledge and practice subordinated to medical knowledge, that is mainly directed by a masculine ideology. PMID- 11111697 TI - [Measurement of the care directed by nurses: comparison between three psychophysical methods]. AB - The degree of directed care required from nurses who work at the critical care unit environment was assessed and compared through three different psychophysical methods: magnitude estimation, category rating and ordinal scales. The sample was formed by 29 participants whose ages varied from 25 to 44 years. Based on the results, authors concluded that: a) data indicated a high concordance among participants regarding different situations with a Kendall's coefficient of 0.83 (p < 0.0016); b) the results obtained with magnitude estimation scaling proved superiority of this method when compared to others. c) the continuum of directed attention (non metric) has characteristics of a protetic continuum. PMID- 11111698 TI - [Introduction of the nursing process, using nursing diagnoses (taxonomy of the North American Nursing Diagnosis Association) expected results, interventions and collaborative problems]. PMID- 11111699 TI - [Reasons for the registration of occupational accidents of nursing personnel]. PMID- 11111700 TI - [Being a staff nurse in 2000, reflections and theory: two or three items apropos of hospital and management changes]. PMID- 11111701 TI - [Nursing staff yesterday and today]. PMID- 11111702 TI - [From sociological analysis to paramedical research]. PMID- 11111703 TI - [Nursing staff in the year 2000, intercultural teacher]. PMID- 11111704 TI - [Trust in a changing salaried relationship, the case of nursing staff]. PMID- 11111705 TI - [Authority, authorization by the nursing staff]. PMID- 11111706 TI - [Position of the nursing staff]. PMID- 11111708 TI - [Teaching nursing care by the practice of active education]. PMID- 11111707 TI - [From participating management to constructive management]. PMID- 11111709 TI - [Pedagogy. The notebook of nursing students]. PMID- 11111710 TI - [Diagnosis, from its formation to the decision for action]. PMID- 11111711 TI - [Documentation. Administrative documentation]. PMID- 11111712 TI - [Research notes]. PMID- 11111713 TI - [The quality of education in the French university nursing institutes, a tool ... some perspectives]. PMID- 11111714 TI - [The malaise of supervisors. Interview by Paule Bourret]. PMID- 11111715 TI - [Current thinking on health and social issues. Proposing exceptions for euthanasia?]. PMID- 11111716 TI - [Improving the level of qualifications for European nurses concerning public health]. PMID- 11111717 TI - [We want to contribute, in French, to the free circulation of ideas and of the expertise of nurses. Interview by Ghislaine Trabacchi]. PMID- 11111718 TI - [National Qualibio meeting on risk management in health facilities]. PMID- 11111719 TI - [The category of general nurse will celebrate its 50 anniversary in Rheims] [In Process Citation] PMID- 11111720 TI - [Management, what function?]. PMID- 11111721 TI - [Management, science, art, magic?]. PMID- 11111722 TI - [What is the role of the health administrators with regard to the new stakes in health care?]. PMID- 11111723 TI - [Higher hospital management, profession and position]. PMID- 11111724 TI - [Anatomy of a pedagogical sequence]. PMID- 11111725 TI - [The meaning of administration]. PMID- 11111726 TI - [About care, about studies and about nurses]. PMID- 11111727 TI - [Grading, evaluation, what do they mean?]. PMID- 11111728 TI - [How to be a manager in the year 2000]. PMID- 11111730 TI - [Bibliography] [In Process Citation] PMID- 11111729 TI - [Common formation of managers, memory and perspective]. PMID- 11111731 TI - [Management. The general nurse 2000, profile and representations]. PMID- 11111732 TI - [Professional exercise: nursing diagnosis, a catch-all or proof of a disciplinary specialty?]. PMID- 11111734 TI - [Documentation. The medical library of a public health system]. PMID- 11111733 TI - [Relational processes, a pedagogical challenge]. PMID- 11111735 TI - [Speed reading]. PMID- 11111736 TI - [Nursing schools caught between the will and anxiety to change before the start of 2000]. PMID- 11111737 TI - [Psychopathology of the aged living at home]. PMID- 11111738 TI - [Under what circumstances is hospitalization of the aged indicated?]. PMID- 11111739 TI - [Acute hemorrhage of the digestive system in the aged]. PMID- 11111741 TI - [Aging and senility: a psychoanalytic and phenomenologic study]. PMID- 11111740 TI - [Facing an abundance of food, what are the criteria of choice?]. PMID- 11111742 TI - [Physical therapy for decubitus ulcers in the elderly]. PMID- 11111743 TI - [The Alzheimer patient and his relatives. Family care]. PMID- 11111744 TI - [Last will and testament]. PMID- 11111745 TI - [Aged parents and aging handicapped persons, a new situation requiring solution]. PMID- 11111746 TI - [Osteoporosis]. PMID- 11111747 TI - [Mistreatment of the aged]. PMID- 11111748 TI - [Animals in institutions for the aged]. PMID- 11111749 TI - [Human/animal relations, a long, shared history]. PMID- 11111750 TI - [Animals in institutions, the conditions of these locations]. PMID- 11111751 TI - [Dogs in a collective, trained to serve their superiors]. PMID- 11111752 TI - [A pilot study in the Loire-Atlantic region]. PMID- 11111753 TI - [Pruritus]. PMID- 11111754 TI - [Bread and cereals]. PMID- 11111755 TI - [Organizational aspects and the function of linens]. PMID- 11111756 TI - [Equilibrium rehabilitation and prevention of accidental falls in the aged]. PMID- 11111757 TI - [Myosotis, and do not forget the Alzheimer patients]. PMID- 11111758 TI - [Donation between spouses]. PMID- 11111759 TI - [Architecture and quality of life: rehabilitation of the sick elderly patient and treatment planning]. PMID- 11111760 TI - [Horton disease]. PMID- 11111761 TI - [To preserve the relation with the family]. PMID- 11111762 TI - [Therapeutic alliance with the family?]. PMID- 11111763 TI - [Role of the nurse in the relation of the dialogue]. PMID- 11111765 TI - [The family: organizational principles]. PMID- 11111764 TI - [Children's guilt feeling and rivalry in the relation to the nursing staff]. PMID- 11111766 TI - [The symptom of family acceptance]. PMID- 11111767 TI - [Family of dreams, family of paper?]. PMID- 11111768 TI - [Nursing practice: writing a case history]. PMID- 11111769 TI - ["I cannot smell her anymore"]. PMID- 11111770 TI - [American sociology and analysis of a psychiatric hospital]. PMID- 11111771 TI - Asymptomatic common bile duct stones. AB - Patients with asymptomatic bile duct stones exhibit typical signs, such as elevated liver function tests, dilated bile ducts on ultrasound, a history of jaundice, or pancreatitis. The incidence of asymptomatic bile duct stones is about 10%, but up to 2% of patients show no signs of the disease. Bile duct stones can be diagnosed by using clinical judgement, scoring systems, or discriminant function tests. Which diagnostic modality is most reliable, cost effective and safe, varies with different hospitals. Which therapy is most effective, safe and the cheapest also varies with different departments, but in the future an increasing number of departments will use the one-stage laparoscopic approach. PMID- 11111772 TI - Asymptomatic bile duct stones: selection criteria for intravenous cholangiography and/or endoscopic retrograde cholangiography prior to laparoscopic cholecystectomy. AB - OBJECTIVE: Routine use of endoscopic retrograde cholangiography (ERC) and/or intravenous cholangiography (IVC) or magnetic resonance cholangiopancreatography (MRCP) before laparoscopic cholecystectomy (LC) is not cost-effective. The objective of this study was to determine precise and easily applicable criteria to select patients who should undergo IVC, MRCP and/or ERC before LC. DESIGN AND METHODS: Prospectively collected data from 74 consecutive patients who were diagnosed with asymptomatic common bile duct stones (CBDS) before undergoing LC, were compared with data from 74 matched controls without CBDS. Using the chi2 test, those variables were identified which were significantly related to the presence of CBDS. These were inserted into a logistic multiple regression model and, by means of conditional regression analysis, each variable was assigned a score from -2 to +4 proportional to the odds ratio. By adding up the scores obtained, a classification was made as to high, medium and low CBDS risk. RESULTS: As a result, 51 patients were found to be low-risk cases, 53 medium-risk and 44 high-risk. Assuming no further assessment of the bile duct needed to be carried out in low-risk patients, an IVC or MRCP in those at medium risk and an ERC in those at high risk, a calculation was made of the positive predictive value and the sensitivity of the system proposed. The positive predictive value and the sensitivity of the procedure were calculated as being greater than 90%. CONCLUSIONS: This predictive system for the risk of CBDS allows the selective use of ERC, IVC and MRCP to ensure a high yield and improve cost-effectiveness. A controlled prospective study will verify these results. PMID- 11111773 TI - The differential effect of VIP and PACAP on guinea pig gallbladder in vitro. AB - OBJECTIVES: Pituitary adenylate cyclase-activating polypeptide (PACAP) is a recently identified member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family. Like VIP, PACAP is largely inhibitory in the gastrointestinal tract. The aim of our work was to characterize the effects of PACAP on both contraction and relaxation of guinea pig gallbladder (GPGB) muscle. METHODS: Gallbladder muscle strips were obtained from male Dunkin-Hartley guinea pigs (250 350 g). Isometric tension was measured in strips suspended in gassed (95% O2, 5% CO2) Krebs' solution at 37 degrees C and equilibrated for 60 min. Cumulative additions of VIP or PACAP (10(-9)-10(-6) mol/l) were performed with strips at basal tone or with strips pre-contracted with cholecystokinin-octapeptide (CCK 8). RESULTS: VIP had no effect on basal tone, in contrast with PACAP which produced concentration-dependent contraction to a maximum of 57.9 +/- 24.3% of control (CCK 3 x 10(-7) mol/l). The highest concentration (10(-6) mol/l) of VIP produced a 32 +/- 6% relaxation of 3 x 10(-9) mol/l CCK-8-contracted GPGB. With 3 x 10(-8) mol/l CCK-contracted GPGB strips, VIP produced a 26.7 +/- 6.6% relaxation. PACAP produced a further concentration-dependent contraction of 3 x 10(-9) mol/l CCK-contracted strips which reached 17.5 +/- 9.9% at the maximal concentration used (10(-6) mol/l). PACAP produced a concentration-dependent relaxation of 3 x 10(-8) mol/l CCK-contracted strips which reached a maximum relaxation of 19 +/- 5% of the control. CONCLUSIONS: PACAP has a dual effect on guinea pig gallbladder motility in vitro, producing contraction in the basal state, and both contraction and relaxation in the CCK-contracted state. This is in contrast to the effects of VIP, a closely related peptide. PMID- 11111774 TI - Differences in intestinal humoral immunity between healthy volunteers from UK and Bangladesh. AB - BACKGROUND/AIMS: Intestinal morphology has been shown to vary geographically. The impact of this variation on gut mucosal humoral immunity is not well-studied. The technique of peroral whole-gut lavage (WGL) with nonabsorbable cleansing fluid can be utilized for the study of gut immune responses in health and disease. In this study, the WGL technique was employed to compare various gut humoral immune parameters in healthy volunteers from Dhaka in Bangladesh and Edinburgh, UK. METHODS: Eleven healthy individuals (all male, age range 18-32) from Dhaka and 12 healthy individuals (4 male and 8 female, age range 23-48) from Edinburgh underwent WGL with a polyethylene glycol electrolyte-based solution drunk at a rate of 1 l/h. The first clear effluent was collected and processed. An ELISA technique was used to measure total immunoglobulins (A, M and G) and antibodies to bacterial lipopolysaccharide (LPS: endotoxin) ovalbumin and eotaxin. Immunoturbimetry and radioimmunoassy techniques were used to measure protein (albumin and alpha-1 antitrypsin) and eosinophil cationic protein (ECP), respectively, in WGL fluid (WGLF). RESULTS: The total IgA, ECP and eotaxin concentrations in WGLF from the Dhaka group were significantly higher than those of the Edinburgh group (P < 0.03, P < 0.002 and P< 0.005 respectively). The IgA antibody level against the core oligosaccharide of bacterial LPS from several Gram-negative species was significantly higher in the Dhaka group compared to the Edinburgh group (P< 0.0001). Similarly, there was generally higher level of IgA antibody response against the various different LPS core structures of Escherichia coli in the Dhaka group, in particular significantly higher against R1, R3 and R4 LPS cores (P< 0.02, P< 0.03 and P< 0.01 respectively) compared to the Edinburgh group. In contrast to antibacterial antibodies, the IgA and IgM antibodies against ovalbumin were significantly lower in the Dhaka group (P< 0.001 and P< 0.003, respectively) compared to the Edinburgh group. CONCLUSIONS: This study on gut mucosal humoral immunity from two geographically distinct populations suggests that place of residence influences gut mucosal humoral immunity. This difference in stimulation of humoral immunity of the gut might explain different rates of inflammatory bowel diseases in developing and developed countries, and also provides a major challenge for the development of mucosally presented vaccine worldwide. PMID- 11111775 TI - Role of lifestyle factors in the pathogenesis of osteopenia in adult coeliac disease: a multivariate analysis. AB - OBJECTIVES: Coeliac disease is frequently complicated by alterations of bone mass and mineral metabolism. In this condition the degree of malabsorption is a major determinant of bone loss. However, the role of lifestyle factors such as exposure to sunlight, physical activity and cigarette smoking, which have been demonstrated to influence bone mass and mineral metabolism in other conditions, has never been investigated in coeliac disease. DESIGN: We evaluated the impact of potential co-factors on bone homeostasis in coeliac disease by means of a multivariate analysis model. METHODS: Thirty-nine adult patients with untreated coeliac disease (18 symptomatic, 21 subclinical/silent) were studied. Bone mineral density was measured by dual-energy X-ray absorptiometry at lumbar spine and femoral neck levels. Age at diagnosis, gender, duration of symptoms and severity of symptoms were recorded. Nutritional status, cigarette smoking habit, exposure to sunlight, and physical activity were evaluated. The impact of each independent variable on lumbar and femoral bone mineral density was evaluated by means of a multivariate analysis model. RESULTS: The severity of symptoms and nutritional status were significant sources of variability of both lumbar and femoral bone mineral density. Physical activity was a significant source of variability at femoral level, while gender was at lumbar level. Cigarette smoking habit and exposure to sunlight showed no significant effect on bone mineral density. CONCLUSIONS: Gender, malnutrition, global severity of the disease and physical activity are important co-factors in the pathogenesis of bone loss in coeliac disease. PMID- 11111776 TI - 13C-urea breath test for the diagnosis of Helicobacter pylori infection: are basal samples necessary? AB - AIM: The 13C-urea breath test (13C-UBT) is one of the best methods for the diagnosis of Helicobacter pylori infection. Basal breath samples are usually obtained, in addition to those obtained after urea intake, as it has been suggested that basal values may oscillate among a population (e.g. depending on diet). However, the superiority of this strategy has not been sufficiently demonstrated. The elimination of basal samples in the 13C-UBT protocol would have the advantages of higher simplicity and speed. METHODS: The 13C-UBT was performed in 714 consecutive patients. Mean age was 48 +/- 16 years, 49% were males, and in 48% of the patients previous H. pylori eradication therapy had been administered. Basal samples (13C-basal) and at 30 min after taking 100 mg of urea labelled with 13C (13C-post-urea) were obtained, delta over baseline (13C-DOB) being the algebraic difference between the ratio 13C/12C at these two points (which is the parameter usually given in studies, being considered positive when > 5%). A citric acid solution was used prior to urea intake. RESULTS: The prevalence of H. pylori infection was 48%. Mean values of 13C-basal, 13C-post-urea, and 13C-DOB were, respectively, -19 +/- 2, 5.9 +/- 33, and 25 +/- 33. 13C-basal values oscillated between -25 and -14, being between -21 and -16 in 90% of the cases. Linear correlation coefficient for 13C-post-urea and 13C-DOB was 0.999 (determination coefficient, 0.998; P< 0.0001). The area under the receiver operating characteristic (ROC) curve for the diagnosis of infection when only 13C post-urea was used (taking 13C-DOB >5% as the 'gold standard') was 0.997. Best results were obtained with 13C-post-urea cut-off point set at -13, with sensitivity of 97.4% (95% CI, 95-99%), specificity of 99.5% (98-100%), and positive likelihood ratio of 180. The concordance kappa value for both tests (13C post-urea and 13C-DOB) using the aforementioned cut-off point was 0.97. CONCLUSION: When performing the 13C-urea breath test it is sufficient to obtain samples 30 min after urea intake. Basal breath samples would not be necessary, which further simplifies this diagnostic method. PMID- 11111777 TI - Cytotoxin-associated gene A and vacuolating cytotoxin A in human isolates of Helicobacter pylori and their association with the clinical status of ulcer disease. AB - OBJECTIVE: The aim of this study was to evaluate whether different Helicobacter pylori genotypes are associated with different clinical stages of peptic ulcer disease (PUD). DESIGN: We assessed the virulence characteristics of H. pylori isolates from patients with active PUD (presence of an ulcer crater at endoscopy) and from those with PUD in remission (normal endoscopic findings or scar not induced by drugs in PUD patients). METHODS: H. pylori isolates from biopsies of the gastric antrum were examined for cagA and vacA genotypes by PCR amplification and Western blot analysis. Descriptive statistical techniques and multivariate polytomous logistic regression were used to estimate adjusted odds ratio (OR) for cagA and vacA genotypes in patients with active PUD or PUD in remission. Patients with non-ulcer dyspepsia (NUD) were used as negative controls. RESULTS: The cagA genotype and phenotype were found to be differently associated with disease status. In fact, the multivariate regression model showed that gastric colonization by CagA+ H. pylori strains was associated with an increased risk of active PUD (OR 2.58), whereas the OR for patients with PUD in remission was 0.94. CONCLUSIONS: Our data indicate that the active ulcer status is more strongly associated with H. pylori strains carrying the pathogenicity island (PAI) than remission status. These results support the hypothesis that a dynamic equilibrium exists among bacterial populations with or without the PAI, and that the relapse of the peptic ulcer could be consequent to expansion of the H. pylori population carrying the PAI. PMID- 11111778 TI - An epidemiological study of acute upper gastrointestinal bleeding in Crete, Greece. AB - OBJECTIVES: Information about the epidemiology of acute upper gastrointestinal bleeding (UGIB) in southern Europe is very limited and especially in Greece non existent. Our study sought to determine the current epidemiology of acute UGIB (incidence, mortality and case fatality) in the prefecture of Heraklion-Crete. DESIGN/METHODS: From February 1998 to February 1999, we prospectively obtained data on all patients with acute UGIB in the prefecture of Heraklion-Crete. All patients who were permanent residents of the prefecture of Heraklion, aged 16 years and over with acute UGIB were included in the study. RESULTS: During this period, 353 cases of acute UGIB were included in the study. The overall incidence of acute UGIB is 160/100,000 adults per year with a male-to-female ratio of 1.7 and a mean age 66.2 +/- 17.1 years. The incidence rises from 30 in those aged under 30 years to 609 in those aged over 75 years. The overall population mortality was 9/100,000 adults per year. Overall case fatality during hospitalization was 5.6%. All deaths occurred in patients older than 60 years. One or more comorbid illnesses were noted in 61% of cases. Recent intake of non steroidal anti-inflammatory drugs (NSAIDs) was reported in 49% of the cases. The most common recorded diagnoses were erosive disease in 108 (30.5%) patients, duodenal ulcer in 97 (27.4%) and gastric ulcer in 75 (21.2%). Rebleeding occurred in 41 patients (12%). Twelve patients (3.3%) had surgery during hospitalization. CONCLUSIONS: The overall annual incidence of acute UGIB in the prefecture of Heraklion-Crete is one of the highest reported in Europe and increases appreciably with age. Both population mortality and case fatality are slightly lower compared to those reported in most previous studies. PMID- 11111779 TI - Effect of ageing on the upper and lower oesophageal sphincters. AB - OBJECTIVE: To determine the effect of ageing on length and resting pressure of the upper and lower oesophageal sphincters (UOSs, LOSs). BACKGROUND: The effectiveness of upper and lower oesophageal sphincters (UOSs and LOSs, respectively) in the control of retrograde trans-sphincteric flow is influenced by sphincteric pressure and length. METHODS: Nine young and nine elderly healthy volunteers were studied. Resting UOS and LOS pressures were measured by sleeve devices and lengths were measured by the station pull-through technique. RESULTS: The length of the UOS high pressure zone in the elderly (2.1 +/- 0.7 cm posterior; 1.9 +/- 0.1 cm anterior) was significantly shorter than that of the young (2.9 +/- 0.1 cm posterior; 3.1 +/- 0.2 cm anterior) (P< 0.01). Resting UOS pressure in the elderly (42 +/- 5 mmHg) was significantly lower than that of the young (62 +/- 7 mmHg) (P< 0.05). The intersphincteric length of the oesophagus in the elderly (21 +/- 0.2 cm) was similar to that of the young (21 +/- 0.4 cm). Total length of the LOS high pressure zone in the young (4.0 +/- 0.1 cm) was similar to that of the elderly (4.1 +/- 0.1 cm). LOS resting pressure was similar between young and elderly subjects (17 +/- 5 mmHg and 15 +/- 3 mmHg, respectively). CONCLUSIONS: Ageing affects the UOS and LOS differently. With regard to resting pressure and length, ageing weakens the UOS, but has no significant effect on the LOS. PMID- 11111780 TI - 6-mercaptopurine or methotrexate added to prednisone induces and maintains remission in steroid-dependent inflammatory bowel disease. AB - BACKGROUND: As treatment of steroid-dependent patients with inflammatory bowel disease (IBD) is controversial, we analysed the efficacy and tolerance of 6 mercaptopurine (6-MP) and methotrexate (MTX) added to prednisone in increasing and maintaining the disease remission rate. METHODS: Seventy-two steroid dependent IBD patients, 34 with ulcerative colitis (UC) and 38 with Crohn's disease (CD), receiving treatment with prednisone were randomly assigned in a 2:2:1 ratio to additionally receive, orally, over a period of 30 weeks 1.5 mg/kg/day of 6-MP (group A) or 15 mg/week of MTX (group B), or 3 g/day of 5 aminosalicylic acid (5-ASA) (group C). All patients who achieved remission were included in a maintaining remission study for 76 weeks. Remission was defined after stopping prednisone as a CD activity index of <150 and normal serum orosomucoid concentration for CD patients and a Mayo Clinic score <7 for UC patients. RESULTS: With regard to achieved remission, a significantly higher (P< 0.05) rate existed for UC patients in group A (78.6%) than in group C (25%), with no statistical differences in group B (58.3%) versus C. For CD patients, the rates were significantly higher (P< 0.001 and 0.01, respectively) in groups A (93.7%) and B (80%) versus C (14%). With regard to maintaining remission, UC patients in group A (63.6%) presented significantly higher rates (P < 0.0015 and P < 0.001, respectively) versus 14.3% in group B and none in group C. For CD patients, statistical differences (P < 0.001) existed when comparing rates in groups A (53.3%) and B (66.6%) versus none in group C. Noticeable side effects appeared in 13.3% of patients from group A and 11.5% from group B. CONCLUSIONS: These results suggest that 6-MP or MTX added to prednisone could be effective in steroid sparing, as well as in achieving and maintaining remission in steroid dependent IBD patients. MTX was less effective in maintaining remission in UC patients. PMID- 11111781 TI - Digital rectal examination sampling of stool is less predictive of significant colorectal pathology than stool passed spontaneously. AB - OBJECTIVES: To compare the positivity rate and the positive predictive value of an immunochemical faecal occult blood test (FOBT) carried out by using stool samples obtained during a routine screening method and those obtained during digital rectal examination. DESIGN: Screening programme-based, cross-sectional study. METHODS: In a screening programme-based, cross-sectional study, 1,044 subjects who received both an immunochemical FOBT and colonoscopy were divided into two groups according to stool collection techniques--the routine screening method and the digital rectal examination method. The positivity rate of the immunochemical FOBT, as well as the positive predictive value for colorectal cancer and large adenomatous polyp, were determined in the two groups. RESULTS: The positivity rate and positive predictive value were 3.8% and 60.0% (10.0% for cancer and 50.0% for adenomatous polyp) in the routine screening group, and 9.4% and 26.5% (4.1% for cancer and 22.4% for adenomatous polyp) in the digital rectal examination group, respectively, indicating a significant difference in the positivity rate (P < 0.01) as well as the positive predictive value (P< 0.05) between the two groups. CONCLUSIONS: These results show that digital rectal examination sampling of stool is less predictive of significant colorectal pathology than stool passed spontaneously, and therefore the latter is the preferred method for stool sampling. PMID- 11111782 TI - Duodenal and ampullary obstruction by a Peutz-Jeghers polyp. AB - We report a case of Peutz-Jeghers syndrome presenting with obstruction of the second part of the duodenum and the ampulla of Vater by a large intra-luminal polyp leading to duodenal obstruction and obstructive jaundice. CT scan of the abdomen showed a large polypoidal lesion, a caecal polyp and jejuno-jejunal intussusception. At surgery, two intussusceptions were reduced and leading polyps were excised via two enterotomies; the caecal polyp was excised via caecotomy. The duodenal polyp was excised by limited duodenectomy after frozen section has shown no evidence of malignancy. Histopathological study of all the excised polyps including that of the duodenum showed hamartomatous polyps with no malignant changes. Apart from acute bleeding, this case highlights many of the surgical gastrointestinal complications of Peutz-Jeghers syndrome. It also highlights the unusual combined duodenal and common bile duct obstruction by a large Peutz-Jeghers polyp. The controversial association of this syndrome with cancer and management options is also discussed. PMID- 11111783 TI - Cholecystogastric fistula presenting with haematemesis: diagnosed by endoscopic retrograde cholangiography. AB - The case is reported of a 72-year-old woman suffering from morbid obesity, who presented with haematemesis while on anti-coagulant therapy. The source of the bleeding proved to be the gastric exit of a cholecystogastric fistula. Subsequent cholangitis was successfully treated by endoscopic retrograde cholangiography (ERC) and endoscopic sphincterotomy (ES) while simultaneously the extent of the fistula was established. Cholecystectomy and closure of the fistula was contraindicated because of her morbid obesity. She remained well for 6 months but then presented with a gallstone ileus while another stone was found to be escaping from the gastric fistula. Her morbid obesity resulted in surgical procrastination, which eventually proved fatal. This patient experienced both of the most common types of complication in cholecysto-enteral fistulation, cholangitis and gallstone ileus. Although cholecysto-enteral fistulas (CEF) are probably less common than several decades ago, they are now most likely to be diagnosed during ERC. Gastroenterologists therefore need to be aware of their potential to contribute to the diagnosis and treatment of this surgical condition. PMID- 11111784 TI - Hepatic undifferentiated (embryonal) sarcoma in an adult: a case report and review of the literature. AB - Undifferentiated (embryonal) sarcoma of the liver (USL) is a rare malignant tumour with a poor prognosis. The absence of specific symptoms, the rapid tumour growth, the normality of the common tumour markers, and the consequential delay in the diagnosis often result in significant enlargement of the tumour. To our knowledge, there have been only 42 reported cases of USL in adults worldwide during the 40 years since the clinicopathological entity of USL was defined. We report here a 27-year-old male with USL who has been treated successfully with surgical resection and adjuvant chemotherapy using ifosfamide, adriamycin and cisplatin. Although the prognosis of the disease remains generally poor, long term survival has been achieved recently in patients with a combination of surgery and multi-agent chemotherapy. PMID- 11111785 TI - Multiple black hepatocellular adenomas in a male patient. AB - A 65-year-old man presented with multiple liver tumours. Imaging techniques could not differentiate between adenomas and hepatocellular carcinomas. He had no relevant past medical history. Liver function tests were normal except for a 1.5 fold rise in GGT. AFP was normal. Viral markers were negative. During laparoscopy, numerous black tumours of different sizes were seen. These tumours were adenomas without malignant transformation. Tumoral hepatocytes contained a brown pigment in the canalicular area without evidence of cholestasis. This pigment was Fontana positive and looked like Dubin-Johnson pigment by electron microscopy. The expression of the canalicular multispecific organic anion transporter (cMOAT) was decreased in the tumours but normal in the non-tumoral liver ruling out the diagnosis of Dubin-Johnson syndrome. There was mild iron deposition possibly related to an homozygous H63D mutation in the HFE gene. Three years after their discovery, the size of the tumours remained stable. It is concluded that this male patient with multiple adenomas and mild iron overload is at risk of developing an hepatocellular carcinoma and that the black colour of adenomas is probably due to a partial defect in excretion of organic anions. PMID- 11111786 TI - Cardiopulmonary resuscitation-related injuries and homicidal blunt abdominal trauma in children. AB - Defendants accused of inflicting fatal abdominal injuries to children occasionally raise the defense that the injuries were caused by cardiopulmonary resuscitation (CPR). The purpose of this study is to answer the question: Does closed chest CPR result in fatal blunt abdominal injuries that can be mistaken for homicidal assault? To that end, a retrospective study was conducted of all homicidal blunt abdominal injuries in children 10 years and younger from the Dade, Broward, and Palm Beach Medical Examiner's Offices from 1981 through 1997. These were compared to cases of children who died of natural causes during the same time period in Broward County who had CPR (control group 1) and to children who died of nonvehicular accidental blunt abdominal trauma (control group 2). Children with life-threatening head injuries were excluded. Medical examiner records, autopsy reports, documenting photographs, and clinical records were reviewed. The data analyzed included subject demographics, whether CPR was performed and by whom, and autopsy findings. Thirty-three child homicides with fatal abdominal injuries were reviewed. Twenty-four (73%) of the homicides received CPR. There was no difference in the nature and severity of injuries between the 24 children who received CPR and the 9 who did not. Three hundred and twenty-four cases of pediatric natural deaths were reviewed, all of which had CPR. No traumatic abdominal injuries were found in any of the children who died of natural causes. Only four children who died of natural causes had evidence of extraabdominal trauma related to CPR. No cases of nonvehicular accidental blunt abdominal trauma were identified during the 17-year period, although there were nonvehicular accidental fatalities due to extraabdominal injuries. The likelihood of CPR-related primary abdominal trauma in child homicides is very low. PMID- 11111788 TI - Pulmonary macrophage counts in deceased infants: baseline data for further study of infant mortality. AB - Infant lung samples were obtained prospectively at autopsy by medical examiner pathologists in five areas of the United States and regardless of the cause of death. Four sections were examined for each case and were taken from the anterior and posterior aspects of the right and left upper lung lobes. Lung sections were stained with HAM-56 immunostain, which is specific for macrophages. Sixty-one cases were evaluated for the study. Three pathologists independently counted the number of macrophages per 40x field (10x ocular) in 10 contiguous fields near the center of each lung section examined. There was good agreement between pathologists on the average number of macrophages observed in each case. The mean macrophage count for all fields counted was 16.5 per 40x field (range 0-136), and the mean for individual cases was 16 (range 6.6-39.4). There was no observed difference between right, left, anterior, and posterior lung sections. There was a tendency for cases certified as sudden infant death syndrome to show lower macrophage counts than those with other causes of death, but the difference was of only marginal statistical significance. Seven of 10 cases in which infants died after a survival period in the hospital had a mean macrophage count greater than the overall mean of 16 per 40x field. These data suggest that mean pulmonary macrophage counts > 16 per 40x field may be a marker for causes of death other than sudden infant death syndrome or that there was a survival interval. These data may be useful as baseline data for further studies of infant mortality possibly involving pathologic changes in the lungs. PMID- 11111787 TI - Changing trends in the diagnosis of sudden infant death. AB - A study of 114 consecutive cases of unexpected infant death that occurred in South Australia over a 5-year period from January 1994 to December 1998 was undertaken. There were 45 deaths attributed to sudden infant death syndrome (SIDS), 19 to natural causes, 21 to accidents. and 5 to homicides; 24 cases were listed as "undetermined." Although there has been a genuine and continued decline in SIDS numbers in this population, there has also been an increase in the diagnosis of cases of accidental asphyxia due to unsafe sleeping environments and of cases in which the family background and autopsy findings suggested more complex mechanisms. The change in diagnostic profile has followed the introduction of more rigorous clinical history review, death scene examination, and autopsy testing. Thus, although diagnostic outcomes have altered in this population, it is more likely the result of more careful interpretation of the extensive investigations that are now undertaken rather than arbitrary reclassification. PMID- 11111789 TI - Pulmonary hemosiderin in deceased infants: baseline data for further study of infant mortality. AB - Infant lung samples were obtained prospectively at autopsy by medical examiner pathologists in five areas of the United States. Tissues were submitted regardless of the cause of death. Lung sections were stained with Prussian blue to detect deposits of hemosiderin. Fifty-nine cases were evaluated for the study. The four sections examined for each case were taken from the anterior and posterior aspects of the right and left upper lung. Three pathologists independently scanned the lung sections microscopically using a 10x objective lens (with 10x ocular lens) and indicated an "iron score" by indicating for each section if it showed no staining for iron-hemosiderin (Score 0), occasional staining with most fields negative (Score 1), focally abundant staining with most fields having no staining (Score 2), focally abundant staining with most fields showing positive staining (Score 3), or prominent staining throughout the section (Score 4). There was good agreement between pathologists on the iron score for each case. A total iron score was calculated by adding the scores based on each pathologist's observations. The mean total iron score was 6 (range, 1-44), with the range of possible total iron scores being 0 to 48. There was no significant difference between the four lung sections in a given case. Six cases had total iron scores that were at least twice the mean (i.e., total iron score > 12); in five of these cases death was caused by conditions other than sudden infant death syndrome, including one case in which asphyxia was the cause of death. These data are consistent with other reports that pulmonary hemosiderin in deceased infants is suggestive of a cause of death other than sudden infant death syndrome. The data may be useful as baseline data for further studies of infant mortality involving possible pathologic changes in the lungs. PMID- 11111790 TI - Morphine-3-D glucuronide stability in postmortem specimens exposed to bacterial enzymatic hydrolysis. AB - Medical examiners frequently rely on the finding of free morphine present in postmortem specimens to assist in certifying deaths associated with narcotics. In vitro hydrolysis of morphine-3-D glucuronide (M3DG) to free morphine was studied using variable specimen pH, initial degree of specimen putrefaction, storage temperature and time, and the effectiveness of sodium fluoride (NaF) preservation. Reagent M3DG was added to opiate-free fresh blood and urine and to autopsy-derived blood specimens. Reagent bovine glucuronidase was also added to certain specimens. Freshly collected and refrigerated NaF-preserved blood produced minimal free morphine, whereas four of five autopsy blood specimens produced free morphine from M3DG. Increased storage time, temperature, and initial degree of putrefaction resulted in greater free morphine generation despite the absence of viable bacteria. Hydrolysis occurring during specimen storage can generate free morphine from M3DG and may result in erroneous conclusions in certifying narcotic deaths. PMID- 11111791 TI - A review of postmortem alcohol detection as a diagnostic test for substance abuse disorders in suicides. AB - To assess the role of toxicologic detection of alcohol to diagnose substance abuse disorders in suicides, the author reviewed suicide studies with both comprehensive toxicologic and diagnostic data. The sensitivity of alcohol detection to diagnose alcohol and substance abuse disorders in suicides was low in all studies (range, 39%-42%), and the specificity was 80%-95%. A higher cutoff level for alcohol did not increase diagnostic performance. The author concludes that toxicologic detection of alcohol is not a reliable indicator of alcohol and substance abuse disorders in suicides because of the high rate of false negatives. Most of the suicides with positive alcohol detection seem to suffer from chronic substance abuse problems. The role of intoxication is difficult to assess because of methodologic problems. PMID- 11111792 TI - Determination of formic acid tissue and fluid concentrations in three fatalities due to methanol poisoning. AB - Three fatalities caused by methanol ingestion are reported. Admission blood methanol concentrations ranged from 0.28 to 4.6 g/L. Two patients had been admitted after a significant delay (>20 hours), and one patient was observed within 90 minutes following ingestion. Formic acid levels were determined in blood samples at admission and ranged from 302 to 680 mg/L. The patients died 44, 55, and 82 hours after poisoning. Formic acid determinations in postmortem tissues were performed by a gas chromatograph method. The authors found great variability in formic acid distribution among the patients and among organs. PMID- 11111793 TI - The Haddon matrix: applying an epidemiologic research tool as a framework for death investigation. AB - The Haddon matrix is a research tool used by injury epidemiologists. Although this matrix has typically been used only in epidemiologic studies, it may serve as a framework to investigate the circumstances of traumatic deaths. This matrix consists of three rows representing time phases (before the injury incident, during the incident, and after the incident) and four columns representing the energy agent, characteristics of the deceased person, the environment, and the vehicle or vector resulting in the abnormal energy exchange, which are considered in the context of the three time phases. The authors present four cases illustrating how this epidemiologic tool can be useful during death investigations. Although the objectives for epidemiologic studies and medicolegal death investigations differ, this approach can be used to describe the circumstances surrounding an injury-related death. PMID- 11111794 TI - Traumatic rupture of the basilar artery: report of two cases and review of the literature. AB - Two cases of traumatic rupture of the basilar artery are reported. In the first case, severe basal subarachnoid hemorrhage (SAH) due to a complete transverse tear of the basilar artery was observed in a 53-year-old restrained male driver who was involved in a head-on collision while intoxicated and drowsy. He lost consciousness shortly after the accident and was admitted to hospital in cardiopulmonary arrest. Intensive resuscitative therapies produced cardiac response, but he died 50 minutes after the accident. The ethanol concentration in his blood and urine was 0.35 and 0.55 mg/ml, respectively. In the second case, SAH due to a similar tear of the basilar artery was observed in a 47-year-old man who received several fist blows to the face while intoxicated. He suddenly lost consciousness after the final blow and was admitted to hospital in cardiopulmonary arrest. Intensive resuscitative therapies produced cardiac response, but he died 6 hours after the event. In these cases, the mechanism of the traumatic rupture of the basilar artery is thought to be overstretching due to hyperextension of the head, and intoxication, drowsiness, or both may have interfered with the decedents' ability to protect themselves; thus, the hyperextension of the head may have been rather forceful. PMID- 11111795 TI - Coronary arteritis diagnosed at autopsy: three case reports and review of the literature. AB - Coronary arteritis is rare but can be fatal either by itself or in conjunction with other diseases. The authors report cases of three men in whom coronary arteritis was an interesting finding that may have caused or contributed to death. One 45-year-old man collapsed at work, another 56-year-old man was found dead in his parked car, and one 80-year-old man had a recent cerebrovascular accident. All three men had coronary arteritis, arteriosclerotic cardiovascular disease, some form of myocardial disease, and fatty liver change. Two had different lung diseases. The findings suggest that coronary arteritis may be an independent cause of death, part of a systemic disease, or, as these three cases illustrate, part of a constellation of cardiac and cardiovascular pathologies with a possible relation to other medical conditions. Coronary arteritis is an important finding in forensic pathology and merits consideration in a case of unexplained death. PMID- 11111796 TI - Trophoblastic microemboli as a marker for preeclampsia-eclampsia in sudden unexpected maternal death: a case report and review of the literature. AB - The authors report the case of a 25-year-old white woman at 7 months' gestation who died suddenly and unexpectedly at home. Anatomic findings at autopsy included a tongue contusion, glomerulonephritis, changes indicative of systemic hypertension, and trophoblastic microemboli in the lungs. Review of the prenatal care record disclosed 3+ proteinuria 2 days before death. The features of the postmortem examination were consistent with clinically undiagnosed preeclampsia eclampsia and glomerulonephritis. The authors discuss the rarity of fatal preeclampsia-eclampsia, the contribution of concomitant glomerulonephritis, and the significance of trophoblastic microemboli in the lungs. PMID- 11111797 TI - Methadone maintenance programs--a two-edged sword? AB - Retrospective review was undertaken of all autopsies in which methadone was detected at the Forensic Science Centre, South Australia, during a 3-year period from July 1996 to June 1999. Thirty-five cases were found in which methadone had either caused or contributed to death (age range = 14-54 years; average = 31 years; M:F = 3.4:1). Ten victims were participating in a methadone maintenance program, of whom four died within a week of enrollment. Eight victims (23%) not enrolled in a methadone maintenance program were found who had died after the use of "diverted" methadone (i.e., prescribed for someone else) (age range = 14-34 years; average = 25 years; M:F = 6:2). Deaths were directly attributable to methadone toxicity in seven of the eight cases, with additional drugs or alcohol being found in seven cases. Prevention of ongoing deaths caused by methadone diversion could be achieved by allowing only daily releases of methadone, with the addict having to consume the drug under close supervision. PMID- 11111798 TI - Mechanical airway obstruction caused by accidental aspiration of part of a ballpoint pen. AB - The authors present three cases of death in children aged 4, 9, and 10 years, respectively, that were first thought to be caused by herbal or other poisonings but at autopsy were found to be caused by airway obstruction from aspiration of ballpoint pen parts. Aspiration of a foreign body is a leading cause of accidental death in children, but the circumstances in these cases were unique. In the first case, a 4-year-old child died shortly after a visit to a traditional healer. The child's mother blamed him for the death and fatally assaulted him. The second case was a 9-year-old who died at school. Case 3 was a 10-year-old who collapsed while playing with a ballpoint pen in her mouth. In the latter two cases, the relatives alleged poisoning. At autopsy, there was no evidence of trauma, disease, or poisoning in all three cases. Ballpoint pen parts were present in the larynx, carina, and left main bronchus, respectively. Features of "asphyxial" death were present, and included subconjunctival hemorrhages, subendocardial hemorrhages, and congestion of the face and internal organs. These deaths are preventable by education of children, parents, and teachers. Ballpoint pen manufacturers should also modify the design of these pens to improve their safety. PMID- 11111799 TI - Postmortem diagnosis of cerebral malaria. AB - Human cerebral malaria is a frequent encephalopathy that occurs in the endemic tropical-subtropical zones. There are a smaller number of imported cases in continental zones where the diagnosis sometimes remains difficult to establish. Fifteen days after the death of a 36-year-old male French citizen in Africa, an investigation to determine the cause of death was conducted. Histologic examination of the brain permitted the diagnosis of cerebral malaria. Because of the popularity of overseas tourism and because this disorder may appear as "sudden death," these victims may be referred to a forensic pathologist. This case demonstrates the role a forensic pathologist may play in determining the cause of death in cerebral malaria. PMID- 11111800 TI - Lethal combination of tramadol and multiple drugs affecting serotonin. AB - The death of a 36-year-old alcoholic man who died after developing seizure activity while being treated with tramadol, as well as with venlafaxine, trazodone, and quetiapine, all of which interact with the neurotransmitter serotonin, is reported. The decedent, who had a history of chronic back pain, alcoholism, depression, mild hypertensive cardiovascular disease, and gastritis, had just been discharged from the hospital after 4 days of alcohol detoxification treatment. During the admission, no withdrawal seizures were noted. The morning after discharge, a witness observed the decedent exhibiting seizure activity and then collapsing. An autopsy was performed approximately 6 hours after death, and the anatomic findings were consistent with seizure activity and collapse, which included biting injuries of the tongue and soft-tissue injuries of the face. Toxicologic analysis identified tramadol, venlafaxine, promethazine, and acetaminophen in the urine; tramadol (0.70 mg/L) and venlafaxine (0.30 mg/L) in the heart blood, and 0.10 mg of tramadol in 40 ml of submitted stomach contents. No metabolites, such as acetate, acetone, lactate, and pyruvate, were found in the specimens that would be characteristically found in a person with alcohol withdrawal syndrome. The threshold for seizures is lowered by tramadol. In addition, the risk for seizure is enhanced by the concomitant use of tramadol with selective serotonin reuptake inhibitors or neuroleptics, and its use in patients with a recognized risk for seizures, i.e., alcohol withdrawal. The cause of death in this individual was seizure activity complicating therapy for back pain, depression, and alcohol withdrawal syndrome. The data in Adverse Event Reporting System of the Food and Drug Administration from November 1, 1997 to September 8, 1999 was reviewed along with a MEDLINE search from 1966 to the present. This case appears to be the first reported death caused by seizure activity in a patient taking tramadol in combination with drugs that affect serotonin. PMID- 11111801 TI - Influenza A virus infection complicated by fatal myocarditis. AB - Influenza virus typically causes a febrile respiratory illness, but it can present with a variety of other clinical manifestations. We report a fatal case of myocarditis associated with influenza A infection. A previously healthy 11 year-old girl had malaise and fever for approximately 1 week before a sudden, witnessed fatal collapse at home. Autopsy revealed a pericardial effusion, a mixed lymphocytic and neutrophilic myocarditis, a mild lymphocytic interstitial pneumonia, focal bronchial/bronchiolar mucosal necrosis, and histologic changes consistent with asthma. Infection with influenza A (H3N2) was confirmed by virus isolation from a postmortem nasopharyngeal swab. Attempts to isolate virus from heart and lung tissue were unsuccessful. Immunohistochemical tests directed against influenza A antigens and in situ hybridization for influenza A genetic material demonstrated positive staining in bronchial epithelial cells, whereas heart sections were negative. Sudden death is a rare complication of influenza and may be caused by myocarditis. Forensic pathologists should be aware that postmortem nasopharyngeal swabs for viral culture and immunohistochemical or in situ hybridization procedures on lung tissue might be necessary to achieve a diagnosis. Because neither culturable virus nor influenza viral antigen could be identified in heart tissue, the pathogenesis of influenza myocarditis in this case is unlikely to be the result of direct infection of myocardium by the virus. The risk factors for developing myocarditis during an influenza infection are unknown. PMID- 11111802 TI - Sudden death from pelvic hemorrhage after bilateral central fracture dislocations of the hip due to an epileptic seizure. AB - Fracture and dislocation of major joints may be caused by the forceful tonic muscular contractions of seizure activity. A 77-year-old man who was found dead in bed with no sign of external trauma had bilateral central fracture dislocations of the femoral head through the acetabular floor with fatal pelvic hemorrhage and extensive pulmonary fat and bone marrow embolism. He had epilepsy, but the last seizure was 6 years earlier, and he had long discontinued medication. The fractures were attributed to a new unwitnessed seizure. This is the twentieth case of central fracture dislocation of the hip since 1970, when better anesthesia eliminated convulsive therapy-induced fractures. The authors review these 20 cases. Seizures followed inflammation, infarction or neoplasia of the brain, eclampsia, metabolic or iatrogenic causes, or epilepsy (6 cases, 2 of which had no prior seizures for 5 years). There were 11 men (mean age, 64 years) and 9 women (mean age, 47 years). Fractures were unilateral in 13 and bilateral in 7. Additional fractures (in vertebrae, shoulders, or femur) were present in eight. Only eight had prior bone disease. Local symptoms led to diagnosis in most, but two were identified incidentally on imaging. The current patient was the only one to die suddenly, but six other patients presented with shock and three died (one of whom had injuries that led to a suspicion of manslaughter). Central fracture-dislocation of the hip is a rare and little known consequence of seizures, with strong potential for misdiagnosis and lethal complications. PMID- 11111803 TI - Sudden death caused by myocardial tuberculosis: case report and review of the literature. AB - A 25-year-old fit man died suddenly while playing social soccer. Autopsy revealed an infiltrative lesion involving the left ventricle with overlying pericarditis. No other significant pathologic changes were observed. Histologic examination showed necrotizing granulomatous inflammation. No acid-fast bacilli were demonstrated in the pericardial fluid or on histologic examination. The presence of Mycobacterium tuberculosis DNA complex was confirmed by use of the ligase chain reaction technique. The differential diagnosis of myocardial tuberculosis includes sarcoidosis, rheumatic fever, rheumatoid arthritis, giant-cell containing tumors, idiopathic (giant-cell) myocarditis, and bacterial infections such as tularemia and brucellosis. This case illustrates the protean manifestations of tuberculosis and highlights the use of molecular biologic techniques in arriving at a definitive diagnosis in cases of suspected tuberculosis. PMID- 11111804 TI - Unusual infant death: dog attack or postmortem mutilation after child abuse? AB - An unusual form of fatal child abuse is reported in which investigations by the police and the medical examiner were able to distinguish blunt force head trauma followed by postmortem dismemberment from a fatal dog attack. A discussion of the approaches used to ascertain the correct diagnosis is presented, as well as an overview of dog attacks on humans. PMID- 11111805 TI - An analysis of the usefulness of specific stages in the pathologic investigation of sudden infant death. AB - Retrospective analysis of autopsy findings in 60 infants who had been found unexpectedly dead in their cribs or beds in South Australia from 1994 to 1998 was undertaken to determine the diagnostic usefulness of individual stages in the postmortem investigation. Positive findings occurred in 2 of 43 scene examinations (3%), 2 of 60 external examinations (3%), 2 of 11 radiologic examinations (18%), 8 of 60 internal examinations (13%), 7 of 60 histologic examinations (12%), and 3 of 58 microbiologic examinations (5%). No positive findings were detected on toxicologic screening. Not every case underwent each diagnostic step. This gave alternative diagnoses to sudden infant death syndrome (SIDS) in 15 cases (25%). This study demonstrates an increase in the percentage of cases of unexpected infant death due to causes other than SIDS; it also shows the diagnostic yield of individual stages in the postmortem evaluation of such cases. Negative findings were important in giving validity to the diagnosis in the 45 cases that were ultimately designated as SIDS. PMID- 11111806 TI - Plastic bag asphyxia in southeast Scotland. AB - Death resulting from plastic bag asphyxia has been recognized for >40 years, but relatively little is known about either its epidemiology or its pathophysiology. Over 15 years (1984-1998), 30 deaths were attributed to plastic bag asphyxia among the 14,560 autopsies performed in the Forensic Medicine Unit in Edinburgh. These 30 deaths involved 20 male and 10 female subjects, with an age range of 13 to 81 years. Eleven had some alcohol measurable in the blood, with four having levels >80 mg/dl. Only one individual appeared to have ingested a drug overdose, but inhaled substances within the plastic bag may have contributed to death in five cases. The absence of childhood accidental deaths may reflect successful preventive measures. The 3 accidental deaths involved adults (including 2 who died of autoerotic asphyxia), and the remaining deaths were 27 suicides. Of those who committed suicide, most (59%) had chronic psychiatric illness rather than chronic debilitating or terminal physical illness. In contrast with reports from the United States, publicity associated with "self-deliverance" did not result in an increased number of deaths from plastic bag asphyxia (4 deaths in this series). Analysis of the circumstances of all the deaths revealed them to be difficult to predict and hence prevent. PMID- 11111807 TI - Hypotensive hemorrhagic necrosis in basal ganglia and brainstem. AB - Hypotensive hemorrhagic necrosis of the basal ganglia and brainstem has only occasionally been described. Three such cases are reported. Cardiac arrest had occurred in all cases, and it took at least 1 hour to restore adequate circulation. The patients remained comatose for 2 days to 2 weeks until death. Persistent hypotension causing ischemia in the distribution of deep perforating arteries is considered to have been the key underlying mechanism. Hemorrhage is thought to have been caused by extravasation of red blood cells through damaged blood vessels. PMID- 11111808 TI - Effects of individual dental factors on genomic DNA analysis. AB - The use in forensic medicine of methods pertaining to molecular biology has made it possible to identify human remains through the analysis of polymorphic profiles of human DNA. Voluntary, accidental, or natural postmortem degradation, as well as environmental conditions, influences the preservation state of the corpse, making it sometimes difficult to obtain biologic material suitable for genetic analysis (e.g., hair, soft and/or hard tissue). According to their anatomic/morphologic characteristics, dental formations are particularly resistant to external insults and are thus suitable for this kind of research. The purpose of this work, conducted on nonselected dental findings (presenting intrinsic characteristics similar to those usually found in forensic cases) that were homogeneous with regard to environmental factors, was to determine an operative protocol that will enable combination of the maximum availability of genomic DNA with the preservation of the morphologic characteristics of the tooth for classic anthropologic evaluations. PMID- 11111809 TI - Lipids in the proximal tubules of the kidney in diabetic coma. AB - Vacuolization of the renal tubular epithelial cells (the Armanni-Ebstein lesion) associated with diabetic hyperglycemia is usually regarded as an accumulation of glycogen. In a case of death of diabetic coma, the vacuoles were stained strongly for lipids. This observation may have both clinical and therapeutic consequences, and may increase our knowledge of the metabolism in diabetes. PMID- 11111810 TI - Medical examiners and bioterrorism. PMID- 11111811 TI - Restraint asphyxia. PMID- 11111812 TI - Design of the prospective randomized study for the treatment of patients with thrombotic microangiopathy. PRODROMI Study Group. AB - In thrombotic microangiopathies hemolytic uremic syndrome and thrombocytopenic purpura, plasma exchange (PE) therapy using fresh frozen plasma is standard. In almost 20% of the patients, however, this approach is ineffective. This prospective, randomized study for the treatment of patients with thrombotic microangiopathies (PRODROMI) compares PE with fresh frozen plasma (A) and cryosupernatant (B). The participating centers were the University Clinics of Freiburg, Hamburg, Dusseldorf, Essen, Gottingen, Mannheim, Ulm, Jena, Tubingen, Wurzburg, Kreiskrankenhaus Offenburg, Stadt Klinikum Karlsruhe, and Horst-Schmidt Kliniken in Wiesbaden, Germany. Patients (18 to 80 years) were diagnosed by the individual centers based on clinical and laboratory findings (thrombocyte/fragmentocyte count, hemoglobin, serum creatinine, haptoglobin and lactate dehydrogenase levels; negative Coombs-test is obligatory). HIV infection, bone marrow, or solid organ transplantation were exclusion criteria. After written consent, patients were randomized in the A or B group. All patients received 1.5 mg/kg methylprednisolone as a basic therapy. The first PE always was performed with fresh frozen plasma (50 ml/kg). A minimum of 5 and a maximum of 10 PEs were required. Thrombocyte count above 150,000/microl was considered to be a successful therapy. Treatment failure was defined as not responding to 10 PE with a thrombocyte count above 150,000/microl or a fall below this value within 30 days after stopping PE. Patients with clinical and laboratory signs of thrombotic microangiopathy occurring later than 30 days after having stopped PE were considered to have a relapse. Primary endpoints were survival, intensity of required PE sessions (duration, volume, and number), and relapse rate. Follow-up of clinical outcome was 2 years; von Willebrand Factor (vWF), vWF-cleaving protease activity, and Factor H were determined. PMID- 11111813 TI - Comparison of fresh frozen plasma with a standardized serum protein solution following therapeutic plasma exchange in patients with autoimmune disease: a prospective controlled clinical trial. AB - The aim of the study was the comparison of the influence of fresh frozen plasma (FFP) (Freiburg, Germany) and Biseko, Biotest Pharma GmbH (Dreieich, Germany), as a plasma substitute (a standardized, virus inactivated human serum protein solution) on the coagulation factors, inhibitors, proteins, and complement factors in the plasma of autoimmune disease patients following membrane plasma separation. Patients (n = 24) with autoimmune disease were randomized to receive either FFP or Biseko for membrane plasma separation therapy. During each plasma exchange, 100% of the plasma volume was replaced by the respective substitute. Plasma exchange volume was performed once daily for 3 days. Target test parameters of the coagulation system were fibrinogen, fibrinopeptide A, factor VIII (FVIIIC), von Willebrand factor antigen (vWFAg), partial thromboplastin time (PTT), thromboplastin time (Quick value), and antithrombin (AT III). The immunoglobulins were IgG, IgA, and IgM and C-reactive protein (CRP). The thrombocytes were platelet factor 4 (PF4), and complement factors were C3 and C4. Biseko was well tolerated with 1 mild adverse drug reaction (ADR) (n = 1) while FFP gave rise to ADR on 7 occasions (n = 4). Statistically significant differences in the 2 groups were observed for fibrinogen, PTT, Quick value, and AT III. From the clinical point of view, all fluctuations and differences in parameter levels remained clinically silent. The differences had no clinical consequences. Reflecting on a potential decrease in the risk of infections in comparison to FFP therapy and the lower rate of adverse drug reactions, it is possible to postulate an advantage of Biseko for plasma exchange therapy. PMID- 11111814 TI - Therapeutic plasma exchange in treatment of adrenoleukodystrophy. AB - Adrenoleukodystrophy (ALD) is an X-linked disorder of metabolism of very long chain fatty acids (VLCFA) with a frequency of up to 1:20,000 in males. VLCFA C 24:0 and C 26:0 accumulate in the cholesterol ester and ganglioside fraction in plasma and red cells. Symptoms of ALD are ataxia, loss of visual and auditory functions, and cerebral convulsions. Presently, no sure therapeutic approaches are established. Efforts were reported by dietary regimens with VLCFA-restriction and glycerol trioleate and glycerol trierucate intake. In the present trial, we report on a 58-year-old white male suffering from progressive ALD with spastic paraparesis. He has a positive family history back to the 18th century. In this patient, although maximum dietary therapy was applied over a period of 60 months, no normalization of VLCFA C24:0 and C26:0 was reached, and neurological disorders were progressive. As a result, therapeutic plasma exchange (TPE) was applied from 1990 to 1994. Then, for more selective adsorption of VLCFA, dextran-sulfate adsorption (Liposorber, Kaneka, Osaka, Japan) until 1996, and after that, immunoadsorption (Therasorb, Baxter, Munchen, Germany) were used. During these periods (total, 101 months), VLCFA C 24:0 and C 26:0 levels were reduced by 55% and 50% (p < 0.001). The patient experienced a significant improvement in performance and general well-being. No further progression of neuronal disorders was documented. This anecdotal data suggest a very beneficial effect of TPE in treatment of progressive ALD. PMID- 11111815 TI - Surface treated catheters with ion beam based process for blood access. AB - Infection, thrombosis, and stenosis are among the most frequent complications associated with blood contacting catheters. Because these problems are usually related to surface properties of the base catheter material, surface treatment processes such as ion implantation and ion beam assisted deposition (IBAD) (silver based coatings) can be used to mitigate such complications. Because these ion beam based processes affect only the near-surface region (approximately the outer 1 microm), there is little effect on bulk material properties. This study evaluated silver coated large bore catheters used for extracorporeal detoxification. In a 135 patient prospective study, 170 large bore catheters were inserted into the internal jugular or subclavian veins. Seventy-eight surface treated catheters (Spi-Argent, Spire Corporation, Bedford, MA, U.S.A.; n = 32 acute catheters, n = 46 long-term catheters) were inserted in 55 patients. Ninety two untreated catheters placed in 80 patients served as controls (n = 40 acute catheters, n = 52 long-term catheters). After removal, the catheters were cultured for bacterial colonization using standard microbiologic assays. They also were examined using a scanning electron microscope (SEM). Bacterial colonization was observed in 7% of the treated catheters compared with 35.3% of untreated catheters. The SEM investigations showed all treated catheters to possess low thrombogenicity. Results of the study indicate that ion beam based processes can be used to improve thrombus and infection resistance of blood contacting catheters. PMID- 11111816 TI - Rheopheresis: rheologic, functional, and structural aspects. AB - Rheopheresis is a specific application of membrane differential filtration, synonymous with double filtration plasmapheresis for extracorporeal hemorheotherapy, eliminating an exactly defined spectrum of high molecular weight proteins from human plasma (e.g.: fibrinogen, alpha-2-macroglobulin, low-density lipoprotein cholesterol, IgM). This results in the improvement of blood flow and microcirculation initiated by lowering blood and plasma viscosity, and erythrocyte aggregation. In this context, microcirculation stands not only for the patency of small blood vessels, but for the complete interactive network between plasma, blood cells, the vessel wall, and cellular and extracellular compartments of the surrounding tissue. Insufficient tissue oxygenation leads to tissue damage, e.g., a microcirculatory disorder develops, creating acute as well as chronic symptoms. Therefore, impaired microcirculation has a rheologic, functional, and structural dimension with respect to involved organs or tissues. Rheopheresis represents a specific therapeutic approach with an acute rheologic as well as chronic functional and structural effects, which was confirmed in pilot and controlled clinical studies for several organ systems. Data from 2 controlled clinical trials are available for the safe and effective treatment in patients with age-related macular degeneration. PMID- 11111817 TI - Potential of rheopheresis for the treatment of acute ischemic stroke when initiated between 6 and 12 hours. AB - Improvement of hemorheology is one of the most important approaches in the treatment of acute ischemic stroke. We investigated the influence of extracorporal rheopheresis (ER) on cerebral blood flow in patients with acute ischemic stroke and evaluated its therapeutic effect. Thirty-three patients (rheopheresis group, 17; control group, 16; mean age 64 +/- 10 years) with acute ischemic stroke were included in our prospective randomized trial. The first treatment was started within 12 h after onset of symptoms, and treatment was repeated 3 times at an interval of 24 h. Hemorheological parameters were measured before and after each session. The cerebral blood flow was analyzed using 99mTc ECD-SPECT. The functional and neurological outcomes were determined by follow-up investigations after 3 months. The hemorheological parameters were significantly different between the rheopheresis group (18% decrease of plasma viscosity, 55% decrease of red blood cell aggregation) and the control group (no decrease of both parameters). The single photon emission computed tomography (SPECT) analysis showed early reperfusion in 35% of the patients treated with rheopheresis and in 37% of the control group (NS). There were no differences in the neurological outcomes between the 2 groups. Extracorporal rheopheresis is practicable and safe. It rapidly and consistently improved the hemorheological parameters. Although this did not impact on cerebral perfusion or clinical outcome in patients with acute ischemic stroke in this report, we propose that ER deserves to be further evaluated by initiating the first treatment within 6 h post-insult. PMID- 11111818 TI - Immunoadsorption for the treatment of rheumatoid arthritis: final results of a randomized trial. Prosorba Trial Investigators. AB - A double-blind, randomized, placebo controlled study was conducted to determine the efficacy of a promising immunoadsorption treatment device containing staphylococcal protein A (Prosorba Immunoadsorption Column, Cypress Bioscience, Inc., San Diego, CA, U.S.A.) in patients with refractory rheumatoid arthritis (RA). Eligibility criteria required adult RA patients who had failed either methotrexate or 2 other disease modifying antirheumatic drugs (DMARD) and who had predefined active disease. All disease-modifying agents were discontinued at least 30 days prior to entry. Patients received 12 weekly procedures after being randomized to the active treatment arm or to the sham treatment arm (apheresis only). Evaluations were double-blinded and occurred at baseline and periodically for 24 weeks thereafter. Primary efficacy was assessed at 7 and 8 weeks after the completion of 12 treatments (at trial Weeks 19 and 20) using the American College of Rheumatology (ACR) definition of improvement (1,2), and results from the assessments at Weeks 19 and 20 were averaged. Ninety-nine randomized patients had a mean disease duration of 15.4 years and received an average of greater than 5 DMARD regimens prior to entry. Analysis of patients who completed all treatments and follow-up indicated that 15 of 36 (41.7%) column-treated patients responded compared to 5 of 32 (15.6%) sham-treated patients (p < or = 0.003). Intent to treat analysis of all patients who were randomized in the study indicated 15 of 52 (28.9%) column-treated patients responded compared to only 5 of 47 (10.6%) patients who received sham treatments (p = .005). Common adverse events (AEs) included joint pain, fatigue, joint swelling, and hypotension. Central line usage was clearly associated with significant AEs during this trial and is not recommended. Hemoglobin, hematocrit, and mean corpuscular volume values decreased similarly in both treatment arms, attributed to phlebotomy for laboratory and scientific studies and to small, repetitive (normal) apheresis losses. Other AEs such as nausea, rash, pruritus, flushing, and fever occurred in 1 to 6% of treatments in each arm (NS). There was no significant increase in AEs in column treated patients compared to sham-treated patients. Protein A immunoadsorption was proven to be a new therapeutic alternative in patients with severe, refractory disease. PMID- 11111819 TI - Rapid reduction of platelet numbers in thrombocytosis. AB - Thrombocytosis has been reported during chemotherapy with gemcitabine (GEM). A 71 year-old male patient who developed asymptomatic thrombocytosis (1,749 x 10(3)/microl), secondary to GEM chemotherapy for lung carcinoma, was submitted to therapeutic plateletpheresis (TP). We adapted the software of the productive dual needle plateletpheresis of our cell separator in order to reduce the procedure time and the plasma volume harvested. With 2 low-volume TP in 24 h, we achieved a 55% global reduction of the platelet count. While there are no accepted indications for prophylactic plateletpheresis in asymptomatic patients and the measured count should not, by itself, determine whether platelet reduction is indicated, a drastic reduction of an extremely high platelet count, especially if obtained with a relatively short procedure, can be important to prevent acute severe sequelae. PMID- 11111820 TI - Development of a rapid blood transfusion system with the capability of blood purification. AB - Blood is usually irradiated by x-ray to prevent graft-versus-host-disease. However, plasma potassium levels of irradiated blood are rapidly increased during preservation in irradiated blood. The objectives of this study were to develop a rapid blood transfusion system for which irradiated blood can be used and to evaluate the capability of blood purification of the system. Packed red blood cells (RBC) were irradiated (15 Gy x-ray) at 21 days and preserved until 42 days after collection. A blood mixture of RBC and plasma was perfused through a dialyzer at 25, 50, 100, and 200 ml/min. Dialysate was perfused at 100, 100, 500, and 500 ml/min, respectively. Preperfusion levels of sodium, 121; potassium, 35; and chlorine, 76 mEq/L were changed to sodium, 144 to 146; potassium 2.5 to 3.0; and chlorine, 105 to 110 mEq/L, which were comparable with the levels in dialysate after perfusion for 25, 50, and 100 ml/min perfusion groups. For the 200 ml/min perfusion group, potassium was 5.3 mEq/L after perfusion which was slightly higher than other groups, but 84% of the potassium was removed by the system. Citrate levels were significantly decreased to 3.4, 28, 31, and 81 mg/dl for the 25, 50, 100, and 200 ml/min groups, respectively, after perfusions. The rapid transfusion system composed of the dialyzer and the blood pumps was effective in the removal of potassium and in the normalization of electrolytes. Irradiated blood with high levels of potassium can be safely and effectively used for this system in cases requiring massive rapid blood transfusion. PMID- 11111821 TI - Efficacy of different low-density lipoprotein apheresis methods. AB - Low-density lipoprotein (LDL) apheresis is a treatment option in patients with coronary heart disease and drug resistant hypercholesterolemia. Various apheresis systems based on different elimination concepts are currently in use. We compared the efficacy of 4 different apheresis systems concerning the elimination of lipoproteins. The study included 7 patients treated by heparin extracorporeal LDL precipitation (HELP), 10 patients treated by immunoadsorption, 8 patients treated by dextran-sulfate adsorption, and 4 patients treated by cascade filtration. Ten subsequent aphereses were evaluated in patients undergoing regular apheresis for more than 6 months. Total cholesterol decreased by approximately 50% with all 4 systems. LDL cholesterol (LDL-C) (64-67%) and lipoprotein a [Lp(a)] (61-64%) were decreased more effectively by HELP, immunoadsorption, and dextran-sulfate apheresis than by the less specific cascade filtration system [LDL-C reduction 56%, Lp(a) reduction 53%]. Triglyceride concentrations were reduced by 40% (dextran-sulfate) to 49% (cascade filtration) and high-density lipoproteins (HDL) by 9% (dextran-sulfate) to 25% (cascade filtration). On the basis of plasma volume treated, HELP was the most efficient system (LDL-C reduction 25.0%/L plasma), followed by dextran-sulfate (21.0%/L plasma), cascade (19.4%/L plasma), and immunoadsorption (17.0%/L plasma). However, a maximal amount of 3 L plasma can be processed with HELP due to concomitant fibrinogen reduction while there is no such limitation with immunoadsorption. Therefore, the decision of which system should be used in a given patient must be individualized taking the pre-apheresis LDL concentration, concomitant pharmacotherapy, and fibrinogen concentration into account. PMID- 11111822 TI - Outcome of patients on long term low-density lipoprotein apheresis with membrane differential filtration: a case study in three patients 14 years on treatment. AB - Membrane differential filtration (MDF) (1,2) is a variety of cascade filtration. Three patients with primary hyperlipoproteinemia and coronary heart disease (2 patients with foregoing myocardial infarction) were treated with MDF for a period of 14 years. The mean treatment interval was 21 days. The basic level of low density lipoprotein (LDL) cholesterol was about 450 mg/dl, and the level on LDL apheresis with comedication of statins was 180 to 200 mg/dl before treatment. Atherosclerosis progressed slowly during this period, and myocardial infarctions were avoided. However, in all 3 patients angiologic interventions became necessary. MDF is a well tolerated method that can be conducted without allergic hazards. The clinical results compare with those of other apheretic techniques. PMID- 11111823 TI - Removal of anti-A/B antibodies with plasmapheresis in ABO-incompatible kidney transplantation. AB - Because of a shortage of cadaver donors in Japan, ABO-incompatible living kidney transplantation has been carried out. Sixty-seven ABO-incompatible living kidney transplantations (LKT) were performed between January 1989 and December 1995 at our institution. In our previous report on the long-term results of ABO incompatible LKT, graft survival of ABO-incompatible LKT up to 3 years was significantly lower than that of ABO-compatible LKT, but no significant difference was seen from 4 to 8 years. We removed anti-A/B antibodies by immunoadsorption and/or double filtration plasmapheresis before kidney transplantation. There was a significant difference between the anti-A/B antibody titers before and after plasmapheresis. The anti-A/B antibody titers also were well suppressed over the long term after transplantation. PMID- 11111824 TI - Signaling by extracellular nucleotides and nucleosides. PMID- 11111825 TI - Extracellular metabolism of ATP and other nucleotides. PMID- 11111826 TI - Molecular pharmacology of P2Y-receptors. AB - Membrane-bound P2-receptors mediate the actions of extracellular nucleotides in cell-to-cell signalling. P2X-receptors are ligand-gated ion channels, whereas P2Y receptors belong to the superfamily of G-protein-coupled receptors. So far, the P2Y family is composed of eight cloned and functionally defined subtypes. Five of them (P2Y1, P2Y2, P2Y4, P2Y6 and P2Y11) are present in human tissues. The P2Y3-, p2y8- and tp2y-receptors may be species orthologues. The principal physiological agonists of the cloned human P2Y-receptors are ADP (P2Y1), UTP/ATP (P2Y2), UTP (P2Y4), UDP (P2Y6) and ATP (P2Y11). The rat P2Y4-receptor is activated by both UTP and ATP. Specific patterns of polar amino acid residues in the exofacial portions of transmembrane domains (TMs) 6 and 7 of the P2Y-receptors may account for the ligand specificity of the subtypes. Suramin acts as an antagonist at most P2Y-receptors with the exception of P2Y4- and tp2y-receptors. PPADS has been shown to block P2Y1-, the human P2Y4- and P2Y6-receptors. The nucleotide analogue 2'-deoxy-N6-methyladenosine-3',5'-bisphosphate (MRS 2179), in contrast, seems to be a potent and selective antagonist at the P2Y1-receptor. All cloned and functionally expressed P2Y-receptors are able to couple to phospholipase C. The P2Y11-receptor mediates in addition a stimulation of adenylate cyclase and the tp2y-receptor an inhibition of this signal transduction pathway. Other functionally defined subtypes, e.g., the receptor mediating an inhibition of adenylate cyclase in blood platelets, are not yet cloned. The distribution of P2Y1 mRNA is widespread. The receptor plays a crucial role in blood platelet aggregation and mediates the adenine nucleotide-induced release of the endothelium-derived relaxing factor nitric oxide. P2Y1-receptors may also be involved in the modulation of neuro-neural signalling transmission. P2Y2 transcripts are abundantly distributed. One important example for its functional role is the control of chloride ion fluxes in airway epithelia. The P2Y4-receptor is highly expressed in the placenta. The distribution of the P2Y6-receptor is widespread including heart, blood vessels and brain. The P2Y11-receptor may play a role in the differentiation of immunocytes. PMID- 11111827 TI - Neuronal P2X receptors: localisation and functional properties. AB - ATP is a co-transmitter in the central and peripheral nervous system. Extracellular ATP exerts its effects via ionotropic (P2X), as well as metabotropic receptors (P2Y). P2X receptors are involved in fast excitatory synaptic signalling by ATP, whereas the role of P2Y receptors in synaptic transmission is unclear. Seven different mammalian P2X receptor subunits (P2X1-7) have been cloned to date. This article gives an overview about the distribution of these P2X receptor subunits in the nervous system. A comparison is made between the pharmacological properties of recombinant receptors and natively occurring neuronal P2X receptors by means of electrophysiological methods. The subcellular distribution of, developmental influences on, and interspecies differences between P2X receptors are also considered. It is concluded that the properties of native P2X receptors are best explained by a heteromeric assembly of different P2X receptor subunits. PMID- 11111828 TI - Agonists and antagonists acting at P2X receptors: selectivity profiles and functional implications. AB - P2X receptors are nucleotide-gated cation channels composed of homomeric or heteromeric assemblies of three subunits. In the past 7 years, an extended series (P2X1-7) of P2X subunits has been cloned from vertebrate tissues. In this rapidly expanding field, one of the main current challenges is to relate the cloned P2X receptor subtypes to the diverse physiological responses mediated by the native P2X receptors. However, the paucity of useful ligands, especially subtype selective agonists and antagonists as well as radioligands, acts as a considerable impediment to progress. Most of the ligands available are highly limited in terms of their kinetics of action, receptor-affinity, subtype selectivity and P2X receptor-specificity. Their suspected ability to be a substrate for ecto-nucleotidases or to inhibit these enzymes also complicates their use. A number of new antagonists at P2X receptors have recently been described which to some degree are more potent and more selective than earlier antagonists like suramin or pyridoxal-5'-phosphate-6-azophenyl 2',4'-disulfonate (PPADS). This work moves us closer to the ideal goal of classifying the recombinant and native P2X receptor subtypes on the basis of antagonist profiles. This review begins with a brief account of the current status of P2X receptors. It then focuses on the pharmacological properties of a series of key P2 receptor agonists and antagonists and will finish with the discussion of some related therapeutic possibilities. PMID- 11111829 TI - Metabolic flux rates of adenosine in the heart. AB - The quantitatively most important source of adenosine under well-oxygenated conditions is 5'-AMP hydrolyzed by cytosolic 5'-nucleotidase N-I. Hydrolysis of S adenosylhomocysteine and extracellular dephosphorylation of 5'-AMP further contribute to total production. More than 90% of the total production occur intracellularly under well-oxygenated conditions. Besides cardiomyocytes, endothelial cells and smooth muscle contribute significantly to total cardiac adenosine production. Rapid enzymatic conversion of adenosine is provided by adenosine kinase and adenosine deaminase, keeping the cytosolic adenosine concentration in the nanomolar range. Due to the high intracellular rates of adenosine rephosphorylation and deamination the cytosolic is normally below the extracellular adenosine concentration, making the cytosol to a sink rather than a source of adenosine. It is for this reason that blockers of membrane transport enhance the plasma adenosine concentration. With increasing catabolism of adenine nucleotides the rate of intracellular adenosine production exceeds the rate of adenosine deamination and rephosphorylation. Thus, this condition will result in a concentration gradient from intra- to extracellular. Thence, membrane transport blockers would be expected to increase the intracellular adenosine concentration. A considerable insecurity on the importance of experimental data results from species differences of purine metabolism. Cardiac adenosine metabolism has recently been described in quantitative terms using mathematical model analysis. This analysis tool may prove useful in future when (1) clarifying the importance of various regulatory actions described for the different pathways of adenosine metabolism, (2) making quantitative comparisons of different experimental models possible and (3) deepening the insight from experimental data. PMID- 11111830 TI - Structure and function of adenosine receptors and their genes. AB - Four adenosine receptors have been cloned from many mammalian and some non mammalian species. In each case the translated part of the receptor is encoded by two separate exons. Two separate promoters regulate the A1 receptor expression, and a similar situation may pertain also for the other receptors. The receptors are expressed in a cell and tissue specific manner, even though A1 and A2B receptors are found in many different cell types. Emerging data indicate that the receptor protein is targeted to specific parts of the cell. A1 and A3 receptors activate the Gi family of G proteins, whereas A2A and A2B receptors activate the Gs family. However, other G proteins can also be activated even though the physiological significance of this is unknown. Following the activation of G proteins several cellular effector pathways can be affected. Signaling via adenosine receptors is also known to interact in functionally important ways with signaling initiated via other receptors. PMID- 11111831 TI - Functions of neuronal adenosine receptors. AB - Endogenous adenosine in nervous tissue, a central link between energy metabolism and neuronal activity, varies according to behavioral state and (patho)physiological conditions, it may be the major sleep propensity substance. The functional consequences of activation of the four known adenosine receptors, A1, A2A, A2B and A3, are considered here. The mechanisms and electrophysiological actions, mainly those of the A1-receptor, have been extensively studied using in vitro brain-slice preparations. A1-receptor activation inhibits many neurons postsynaptically by inducing or modulating ionic currents and presynaptically by reducing transmitter release. A1-receptors are almost ubiquitous in the brain and affect various K+ (Ileak, IAHP), mixed cationic (Ih), or Ca2+ currents, through activation of Gi/o-proteins (coupled to ion channels, adenylyl cyclase or phospholipases). A2A-receptors are much more localized, their functional role in the striatum is only just emerging. A2B- and A3-receptors may be affected in pathophysiological events, their function is not yet clear. The cAMP-PKA signal cascade plays a central role in the regulation of both neural activity and energy metabolism. Under conditions of increased demand and decreased availability of energy (such as hypoxia, hypoglycemia and/or excessive neuronal activity), adenosine provides a powerful protective feedback mechanism. Interaction with adenosine metabolism is a promising target for therapeutic intervention in neurological and psychiatric diseases such as epilepsy, sleep, movement (parkinsonism or Huntington's disease) or psychiatric disorders (Alzheimer's disease, depression, schizophrenia or addiction). PMID- 11111832 TI - Adenosine receptors and their ligands. AB - The regulatory actions of adenosine are mediated via four subtypes of G protein coupled receptors distinguished as A1, A2A, A2B and A3 receptors. Their presence on basically every cell makes them an interesting target for the pharmacological intervention in many pathophysiological situations. A large number of ligands have been synthesized over the last two decades and provide agonists and antagonists that are more or less selective for the known receptor subtypes. In addition, many radioligands are available in tritiated or radioiodinated form. The comparative pharmacological characterization of all four human adenosine receptor subtypes revealed that some of the compounds thought to be selective from data in other species have unexpected potencies at human receptors. As a result, compounds that exhibit high affinity to only one subtype are an exception. Although the selection of ligands is immense, it is less than satisfying for most subtypes of adenosine receptors. PMID- 11111833 TI - Modified 5-HT3A receptor function by co-expression of alternatively spliced human 5-HT3A receptor isoforms. AB - Serotonin (5-HT) exerts fast excitatory responses by activation of 5-HT3 receptors, irrespective of whether they are homomerically composed of 5-HT3A subunits or heteromerically assembled of 5-HT3A and 5-HT3B subunits. Here we describe a short, truncated (h5-HT3AT) and a long (h5-HT3AL) splice variant of the human 5-HT3A (hS-HT3A) receptor subunit. The deduced protein of the short isoform consists of 238 amino acids (aa) with a single transmembrane domain (M1). Compared to the known 5-HT3A receptor, the long isoform contains 32 additional aa in the extracellular loop between M2 and M3. Both splice variants are co expressed together with the 5-HT3A subunit in the amygdala and hippocampus, whereas in the placenta only the short variant is co-expressed. Both splice variants, when expressed in transfected human embryonic kidney (HEK) 293 cells, are not able to form functional homomeric receptors, but modify 5-HT response at heteromeric h5-HT3A receptors. Co-expression of the short variant considerably decelerates the desensitization of the 5-HT3 receptor; thus, heteromeric assemblies of h5-HT3A and the h5-HT3AT subunit exhibit 5-HT-induced cation fluxes which are much larger than those of homomeric hS-HT3A receptors. In contrast, heteromeric complexes containing the h5-HT3AL subunit display reduced cation fluxes. In conclusion, the splice variants increase the functional diversity of 5 HT3 receptors. PMID- 11111834 TI - Phenotypic expression of the systemic toxicity of cocaine in genetically epilepsy prone rats. AB - The purpose of the present investigation was to determine whether the sensitivity to systemic toxic effects of cocaine is altered in genetically epilepsy-prone rats (GEPRs). Moderate seizure (GEPR-3) and severe seizure (GEPR-9) rats, and the control strain, Sprague-Dawley rats, 10 weeks of age, were lightly anesthetized with halothane and nitrous oxide. Following surgical preparation and stabilization, the animals were given a constant intravenous infusion of cocaine (4 mg/kg per min) until death. Blood pressure, ECG, and EEG were monitored continuously throughout the experiment. Cocaine doses required to produce seizures (i.e., epileptiform activity on the EEG) were not significantly different between GEPRs and control rats (16.8+/-0.6 mg/kg in GEPR-3, 18.7+/-0.7 mg/kg in GEPR-9, and 14.7+/-1.3 mg/kg in Sprague-Dawley). Seizure duration, amplitude and the number of epileptiform bursts were also similar among the three strains. Additionally, there was no significant difference in cocaine doses that produced arrhythmias and cardiac asystole between GEPRs and control. The results indicate that genetically epilepsy-prone rats do not exhibit altered sensitivity to cocaine-induced seizures despite the marked susceptibility to sound-evoked seizures. Local anesthetic-induced seizures and acoustically-evoked seizures apparently have different underlying mechanisms. PMID- 11111835 TI - Interaction of the antidepressant mirtazapine with alpha2-adrenoceptors modulating the release of 5-HT in different rat brain regions in vivo. AB - Mirtazapine (MIR) is a novel antidepressant, reported to raise extracellular noradrenaline (NA) through blockade of alpha2-autoreceptors and serotonin (5-HT) output via (1) indirect activation of facilitatory alpha1-adrenoceptors on the cell bodies of ascending 5-HT neurones and (2) blockade of presynaptic release modulating alpha2-heteroreceptors on 5-HT terminals in the forebrain. To further assess the effect of MIR on NA/5-HT system interplay, including putative regional differences in the effects of the drug on 5-HT release in rat forebrain, we used in vivo microdialysis in anaesthetised rats. Probes were implanted in the dorsal hippocampus (DH) and frontal cortex (FCx), representing median and dorsal raphe 5 HT projection areas, respectively. In the DH, MIR (10 mg/kg s.c.) completely blocked the 5-HT release-suppressing action of the selective alpha2-adrenoceptor agonist clonidine (0.1 mg/kg s.c.), but had no effect per se on the 5-HT output. Neither drug significantly changed the 5-HT levels in the FCx. MIR perfused locally (10 microM via reverse-dialysis) also failed to significantly elevate 5 HT output, and did not affect the clonidine response in either brain area. Thus, the data confirm the basic alpha2-adrenoceptor-blocking properties of MIR, but are only partly concordant with previous studies reporting an increase of 5-HT output after MIR alone. Moreover, we find no elevation in 5-HT by the reference alpha2-adrenoceptor antagonist idazoxan (0.3-1.0 mg/kg s.c.). The discrepancies encountered, and the potential ability of alpha2-adrenoceptor antagonists in general to raise the output of 5-HT, are discussed with particular reference to methodological and other factors that may influence the experimental outcome (e.g., brain regional aspects, different alpha2-adrenoceptor subtypes, potential differences in adrenoceptor tone under varying experimental conditions). PMID- 11111836 TI - Effect of amphetamine-induced dopamine release on radiotracer binding to D1 and D2 receptors in rat brain striatal slices. AB - The in vivo binding of positron emission tomography (PET) and single photon emission computer tomography (SPECT) radiotracers to dopamine D2 receptors in the striatum can be influenced by competition with endogenous dopamine. The present study was undertaken to determine if a similar inhibition of radiotracer binding to dopamine receptors could be observed following pharmacologically-evoked dopamine release in rat brain striatal slices. Striatal slices were incubated in a large volume of oxygenated Krebs saline and exposed to amphetamine or methamphetamine to evoke dopamine release within the slice. Amphetamine and methamphetamine, at concentrations up to 30 microM, reduced [3H]raclopride binding in the slices by 77% and 86%, respectively, with 50% inhibition at 1.6 microM amphetamine or 3.0 microM methamphetamine. Neither drug produced a significant effect on binding of [3H]SCH 23390 in the slices. This suggests that dopamine was able to interfere with radiotracer binding to D2 but not D1 receptors. The dopamine uptake blockers, cocaine and methylphenidate, had relatively little effect by themselves on [3H]raclopride binding but, by inhibiting amphetamine-induced dopamine release, significantly reduced inhibition of [3H]raclopride binding by a low (3 microM) amphetamine concentration. At a higher (30 microM) amphetamine concentration the inhibition of [3H]raclopride binding was not antagonized by uptake blockers and data obtained from homogenate binding experiments indicated a direct displacement of [3H]raclopride binding by amphetamine at this concentration. In conclusion the data obtained in the present study demonstrate that the effects of amphetamine on striatal radiotracer accumulation observed in PET and SPECT can also be observed in brain slices in vitro and, at least at low amphetamine concentrations, are mediated by competition with released dopamine. PMID- 11111837 TI - Characterization of the anticonvulsant and neuroprotectant BIIR 561 CL in vitro: effects on native and recombinant alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) receptors. AB - BIIR 561 CL is a novel blocker of AMPA receptors and voltage-dependent sodium channels. In this study we further describe the effects of BIIR 561 CL on AMPA receptor-mediated membrane currents in rodent neurons, as well as in cells expressing recombinant human GluR1/2 receptors in more detail. BIIR 561 CL suppressed responses to kainate in neuronal cultures from rat cortex with an IC50 of 9.8 microM. Similar effects were observed using acutely dissociated neurons from the CA1 region of rat hippocampus (IC50 = 9.5 microM). Inhibition of kainate responses by BIIR 561 CL was prevented by preapplication of GYKI 53655, suggesting that both non-competitive inhibitors bind to a common site of the receptor. The effect of 10 microM BIIR 561 CL on kainate-induced currents was dependent on extracellular pH, with more pronounced block (84.1%) under acidic conditions (pHextern=6.4), compared to only 30.1% at a pHextern of 8.4. Thus, it can be hypothesized that BIIR 561 CL inhibits AMPA receptors in ischaemic brain regions more effectively than in healthy tissue. BIIR 561 CL inhibited responses to 1 mM glutamate in cells expressing recombinant human GluR1/2 receptors with similar potency, as compared to kainate responses in rat neurons (IC50=17.3 microM). The reference compound NBQX had an IC50 of 25.2 nM. None of the two compounds affected the glutamate-induced receptor desensitization at any tested concentration. The block by BIIR 561 CL was not use-dependent and had fast on- and off-kinetics (tauon=6.8 s; tauoff=1.3 s in hGluR1/2 receptors with 30 microM BIIR 561 CL). Thus, BIIR 561 CL can be anticipated to have a promising profile for the treatment of neurological disorders like brain ischaemia and head trauma. PMID- 11111838 TI - Characterisation of adenosine receptors mediating relaxation in hamster isolated aorta. AB - The aim of this study was to characterise the receptor(s) mediating relaxations to adenosine and its analogues in the hamster isolated aorta. Adenosine relaxed the aorta but there was no significant difference between pIC20 values in the absence and presence of 8-sulphophenyltheophylline (8-SPT, 50 microM), although there was a small right-shift (approximately threefold) of the lower portion of the curve in the presence of 8-SPT. However, in the presence of the adenosine uptake inhibitor nitrobenzylthioinosine (NBTI, 1 microM), curves to adenosine were left-shifted by approximately 100-fold and an apparent pK(B) for 8-SPT of 5.79+/-0.05 was obtained. Likewise, 5'-N-ethylcarboxamidoadenosine (NECA) relaxed the aorta but curves were biphasic. The first phase of the curve was blocked by 8 SPT (10-100 microM, pA2 = 5.75+/-0.14) and the A2A-selective antagonist 4-(2-[7 amino-2-(2-furyl) [1,2,4]-triazolo[2,3-a][1,3,5]triazin-5-ylaminolethyl) phenol (ZM 241385, 3 nM-1 microM, pK(B)=9.17+/-0.10). Similarly, the A2A-selective agonist 2-[p)-(2-carbonylethyl)-phenylethylamino]-5'-N-ethylcarboxam idoadenosine (CGS 21680) relaxed the tissues but curves were biphasic and the first phase was again blocked by ZM 241385 (10 nM, apparent pK(B)=9.06+/-0.34). In contrast, relaxations to N6-R-phenylisopropyladenosine (R-PIA), N6-cyclopentyladenosine (CPA), 2-chloroadenosine (2-CADO) and N6-(3-iodobenzyl)-adenosine-5'-N methyluronamide (IB-MECA) were not blocked by 8-SPT (50 microM). Responses to IB MECA were also not blocked by the A3 receptor antagonist 3-ethyl-5-benzyl-2 methyl-6-phenyl-4-phenylethynyl-1,4-(+/-)-dihyd ropyridine-3,5-dicarboxylate (MRS 1191, 30 microM). The asymptote of the first phase of curves to NECA was markedly reduced (and in some preparations the first phase was completely abolished) both in the presence of N(G)-nitro-L-arginine methyl ester (L-NAME, 0.1 mM), and in the absence of endothelium. Likewise, the first phase of curves to CGS 21680 was abolished both in the presence of L-NAME (0.1 mM) and in the absence of endothelium. In contrast, there were only relatively small shifts to the right of curves to adenosine and the other analogues in the presence of L-NAME or the absence of endothelium (between three- and fivefold). The data suggest the presence of A2A receptors which are located on the endothelium and mediate release of nitric oxide. These receptors are activated by NECA, CGS 21680 and adenosine (in the presence of uptake blockade). The resistance to blockade of relaxations to adenosine (in the absence of uptake inhibitor), CPA, R-PIA, 2 CADO, IB-MECA and high concentrations of NECA and CGS 21680 by 8-SPT or ZM 241385 suggests the presence of an additional mechanism(s). Data obtained with adenosine in the absence and presence of NBTI suggest that the endogenous ligand may cause relaxation via an intracellular mechanism. PMID- 11111839 TI - Quantification of the tissue levels and function of the G-protein regulator phosducin-like protein (PhlP). AB - Phosducin-like protein is a protein with wide-spread expression that has been shown to be capable of inhibiting G-protein function in vitro. However, it is not clear whether it is expressed in sufficient amounts to actually exert such functions in vivo. Here we quantify the expression of the short and the long splice variants of phosducin-like protein, PhlPs and PhlP1. Western blots of various rat tissues showed that PhlP1 was by far the dominant splice variant; its levels were 1.5-2 pmol/mg cytosolic protein in brain, liver and kidney, and about 0.5 pmol/mg cytosolic protein in lung, heart and skeletal muscle. These values correspond to concentrations of 150-200 nM and 50 nM, respectively. The levels of PhlPs were about 20-fold lower. Recombinant phosducin, PhlP1 and PhlPs inhibited the interaction between G-protein alpha- und betagamma-subunits with IC50-values of 6 nM, 6 nM and 90 nM, respectively, as determined by Gbetagamma-dependent ADP ribosylation of Galphai1 by pertussis-toxin. Thus, tissue concentrations of PhlP1 are clearly sufficient to affect G-protein function in vivo, while the expression levels and the Gbetagamma-affinity of PhlPs are most likely too low to have significant inhibitory effects on Gbetagamma (G-protein betagamma-subunits). PMID- 11111840 TI - Acamprosate inhibits Ca2+ influx mediated by NMDA receptors and voltage-sensitive Ca2+ channels in cultured rat mesencephalic neurones. AB - Acamprosate has recently been introduced in relapse prophylaxis in weaned alcoholics. Using fura-2 microfluorimetry, the present study investigates whether acamprosate affects N-methyl-D-aspartate (NMDA) or K+-induced changes in free intracellular Ca2+ concentration ([Ca2+]i) in rat cultured mesencephalic neurones. Both application of NMDA (plus glycine) and elevation of extracellular K+ induced rapid increases in [Ca2+]i which respectively were insensitive and sensitive to omega-conotoxin (omega-CTX) MVIIC, a blocker of voltage-dependent Ca2+ channels (VDCCs). Acamprosate (100 microM and 300 microM) significantly attenuated the response induced by NMDA as well as that induced by K+ in a concentration-dependent manner. Concurrent application of omega-CTX MVIIC and acamprosate impaired the K+-induced increase in [Ca2+]i to the same extent as omega-CTX MVIIC alone. The present data suggest that acamprosate inhibits Ca2+ influx through both NMDA receptors and VDCCs. PMID- 11111841 TI - Comparison of the effects of epibatidine and nicotine on the output of dopamine in the dorsal and ventral striatum of freely-moving rats. AB - The effects of the nicotinic acetylcholine receptor (nAChR) agonist epibatidine on the extracellular concentrations of dopamine (DA) and its metabolites 3,4 dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the dorsal (caudate-putamen) and the ventral striatum (nucleus accumbens) of freely-moving male Wistar rats were studied by in vivo microdialysis. In the dorsal striatum, epibatidine (3.0 microg/kg s.c.) significantly elevated the extracellular concentrations of DA, DOPAC and HVA. In contrast, epibatidine did not alter the extracellular DA concentration in the ventral striatum, but elevated significantly the concentration of DOPAC and also tended to elevate that of HVA. In parallel experiments, nicotine (0.5 mg/kg s.c.) significantly increased DA output in the ventral striatum whereas only a modest and non-significant increase of extracellular DA concentration was found in the dorsal striatum. Earlier studies have shown that the doses of epibatidine and nicotine used in the present study are about equieffective at least with respect to the analgesia-producing or hypothermic effects of the drugs. Comparison of the effects of epibatidine and nicotine suggests that the responses of the mesolimbic and nigrostriatal dopaminergic systems to the two nicotinic receptor agonists differ. Epibatidine, in contrast to nicotine, preferentially stimulates the nigrostriatal vs. the mesolimbic dopaminergic system. Therefore, novel nicotinic AChR ligands structurally related to epibatidine may have low abuse potential. PMID- 11111842 TI - Mechanisms underlying endothelium-dependent, nitric oxide/prostacyclin independent, acetylcholine-induced relaxation in canine corpus cavernosum. AB - The mechanisms underlying endothelium-dependent and nitric oxide (NO)/prostacyclin-independent, acetylcholine-induced relaxation in isolated canine corpus cavernosum were investigated. In isolated canine corpus cavernous strips treated with indomethacin (10(-6) M) and N(G)-nitro-L-arginine (10(-4) M), acetylcholine produced relaxations in a concentration-dependent manner. The relaxations were not affected by treatment with 2-(4-carboxyphenyl)-4,4,5,5 tetramethylimidazole-1-oxyl 3-oxide, sodium salt (carboxy PTIO, 3 x 10(-4) M), glibenclamide (10(-6) M), iberiotoxin (10(-7) M) or charybdotoxin (10(-7) M), but were abolished or reversed to contractions by treatment with apamin (10(-8) M) or scyllatoxin (10(-8) M). Levcromakalim (10(-7)-10(-6) M) induced a concentration dependent relaxation which was abolished by treatment with glibenclamide (10(-6) M), but was not affected by treatment with apamin (10(-8) M) or scyllatoxin (10( 8) M). These findings indicate that endothelial cells of canine corpus cavernosum have an ability to produce a relaxing substance(s) other than NO or prostacyclin in response to acetylcholine. The substance(s) may open solely small conductance Ca2+-dependent K+ channels. PMID- 11111843 TI - The sympathetic skin response in carpal tunnel syndrome. AB - The sympathetic skin response (SSR) is an evoked change in electrical skin potential and is an index of the function of sympathetic pathways. We studied the SSR evoked by electrical stimulation of the median nerve and recording from the contralateral hands in 30 patients with carpal tunnel syndrome (CTS) without clinical autonomic signs and compared the results to the SSR in 30 normal controls. The SSR was absent in the affected hands in seven (23%) patients. In the other carpal tunnel syndrome patients (77%), a significant reduction in the SSR area was seen in the records from the affected hands. Subclinical sympathetic nerve fibre involvement occurs in the affected median nerves in CTS. PMID- 11111844 TI - Autonomic control of heart period in duodenal ulcer patients insights from spectral analysis of heart rate variability. AB - Beyond the fundamental pathogenetic importance of Helicobacter Pylori a possible additional role of vagal innervation in favouring or modulating the clinical history of duodenal ulcer (DU) has been suggested by old studies employing invasive methodologies. Aim of this study was to assess whether vagal prevalence in autonomic modulation was present in healed DU patients (n=20) as compared to controls,(n=50), using a validated non-invasive methodology, based on spectral analysis of cardiovascular variability. This approach provides markers of the sympathetic and vagal modulations of the SA node, respectively by way of the normalized low frequency (LF(RR)) and high frequency (HF(RR)) components of RR interval variability; LF/HF ratio furnishes a marker of sympatho-vagal balance. In addition, sham feeding (SF) provided a means to assess, in DU patients, neurally mediated acid secretion, as the SF acid output (SAO) to basal acid output (BAO) ratio (SAO/BAO). Results showed that LF(RR) was smaller in DU patients than in controls (40.3+/-3.9 vs. 52.3+/-2.3 normalized units, nu; P<0.05). On the contrary, HF(RR) was greater (52.1+/-3.7 vs. 35.7+/-2.3 nu; P<0.05). Conversely the LF component of SAP variability, a marker of sympathetic vasomotor modulations, and the index alpha, a measure of baroreflex control of the SA node, as well as respiratory patterns, were similar in the two groups. SAO/BAO ratio was significantly correlated with markers of autonomic control of the SA node (r = -0.67, P<0.0083 with HF(RR)). In conclusion results suggest an enhanced vagal modulation of heart period in DU patients at rest, that appears linked to indices of neurally mediated gastric acid secretion response. PMID- 11111845 TI - Calbindin-D28k in cerebrovascular extrinsic innervation system of the rat. AB - Calbindin-D28k, one of the calcium-binding proteins, belongs to the EF hand family and is commonly found in neurons. It serves as a representative neuronal marker for neuroanatomical investigations. The authors' knowledge of its precise function, however, is yet very limited. In this study, we examined the existence of nerve fibers with calbindin-D28k immunoreactivity in the cerebral blood vessels and ganglia that innervate the cerebral blood vessels in the rat. Numerous nerve fibers with calbindin-D28k immunoreactivity were observed on the walls of the major extracerebral arteries forming the circle of Willis and its branches. Calbindin-D28k immunoreactivity was seen in many neurons of the trigeminal, dorsal root and jugular ganglia. A small number of neurons showed calbindin-D28k immunoreactivity in the otic and superior cervical ganglia. Calbindin-D28k immunoreactivity was not detected in the sphenopalatine or internal carotid ganglia. Pericellular basket-like formations of nerve terminals with calbindin-D28k immunoreactivity were observed in the sphenopalatine, otic, internal carotid and superior cervical ganglia. The present study demonstrated calbindin-D28k immunoreactivity in the cerebrovascular nerve fibers as well as in their origins--the cranial ganglia. These findings are significant in understanding the calcium-mediated mechanism of the neural control of the cerebral blood vessels. PMID- 11111846 TI - Role of alpha1A-adrenoceptor in the regulation of glucose uptake into white adipocyte of rats in vitro. AB - In an attempt to know the functional role of alpha1A-adrenoceptors in adipose tissue, white adipocytes (WAT) of Wistar rats were used to investigate the change of glucose uptake after pharmacological activation of alpha1-adrenoceptors. Methoxamine enhanced the uptake of radioactive glucose into isolated WAT in a concentration-dependent manner. Translocation of glucose transporter (GLUT4) from cytosol to membrane was also stimulated with methoxamine. Action of methoxamine to raise glucose uptake was abolished in WAT pre-incubated with the antagonists, both tamsulosin and WB 4101, at concentrations sufficient to block alpha1A adrenoceptors. However, chlorethylclonidine (CEC). the antagonist of alpha1B adrenoceptors, showed the inhibition of methoxamine-induced action only at a higher concentration. Even under the treatment with maximal concentration of CEC, methoxamine can produce action about 80% of the vehicle-treated control. The major role of alpha1A-adrenoceptors in the stimulation of glucose uptake by methoxamine can thus be considered. In the presence of specific inhibitor of phospholipase C (PLC), U73312, methoxamine-stimulated glucose uptake into WAT was reduced in a concentration-dependent manner and U73343, the negative control of U73312, did not affect the action of methoxamine. Moreover, chelerythrine and GF 109203X diminished the methoxamine-stimulated glucose uptake at a concentration sufficient to inhibit protein kinase C (PKC). Inhibition of phosphoinositide-3 kinase (PI-3 kinase) by LY294002 also abolished methoxamine-stimulated glucose uptake. Therefore. the obtained data suggest that an activation of alpha1A adrenoceptors, presence in WAT, by agonist and/or neurotransmitter may increase the glucose uptake via PLC-PKC pathway and the activation of PI-3 kinase. PMID- 11111847 TI - P2X2 receptor expression by interstitial cells of Cajal in vas deferens implicated in semen emission. AB - Male reproduction is dependent upon seminal emission mediated by vas deferens contraction. This drives spermatic fluid to the prostatic urethra during ejaculation. We localize interstitial cells of Cajal (ICC), which express P2X2 receptor, subunits of ATP-gated ion channels, to rat, mouse and guinea-pig vas deferens submucosa. Reverse transcription-polymerase chain reaction (RT-PCR) analysis of rat vas deferens resolved two functional splice variant transcripts of the P2X2 receptor subunit. The P2X2 receptor mRNA was localized principally within the lamina propria (submucosal) region of the rat vas deferens using in situ hybridization (ISH) and in situ RT-PCR-ISH. Immunohistochemistry using rat, mouse and guinea-pig vas deferens tissues confirmed expression of P2X2 receptor protein within the lamina propria, particularly within a dense column of small spindle-shaped cells adjacent to the columnar epithelial cells which line the lumen. This immunoreactivity was co-localized with neurone-specific enolase (NSE) and c-Kit protein expression, gene markers for ICC. Mucosal mast cells were distinguished from ICC by toluidine blue staining. Choline acetyltransferase immunoreactivity, a marker for post-ganglionic parasympathetic innervation, occurred on the lateral margin of the lamina propria and extended into the inner longitudinal muscle layer. P2X1 receptor immunolabelling was associated with sympathetic innervation of the smooth muscle in the outer longitudinal and circular muscle layers, but not the inner longitudinal layer. The physiological significance of the vas deferens ICC which express P2X2 receptors remains to be established. Possible roles include regulation of smooth muscle activity or mucosal secretion utilizing local ATP signaling, both of which would affect semen transport. PMID- 11111848 TI - Spontaneous migrating motor complexes occur in both the terminal ileum and colon of the C57BL/6 mouse in vitro. AB - We have studied migrating motor complexes (MMCs) in the isolated terminal ileum or colon (IMMCs and CMMCs respectively) of the C57BL/6 mouse. Periodic contractions occurred spontaneously in both preparations in the absence of intraluminal stimulation. After an initial period, complexes became synchronized between the oral and anal ends of the tissue, and could be observed for in excess of 7 h. The propagation velocity was 3.1+/-1.0 and 3.9+/-0.6 mm s(-1) in the ileum and colon respectively. IMMCs occurred every 6.01+/-0.39 min and had a duration of 86.3+/-10.4 s. The interval between CMMCs was smaller (3.52+/-0.31 min) and contractions were shorter in duration (30.7+/-3.6 s). In both preparations, these motor events were dependent on cholinergic transmission: blocked by hexamethonium (500 microM) and attenuated or blocked by atropine (1 microM). This study is the first demonstration of spontaneous migrating contractions in the isolated ileum or colon of the C57BL/6 mouse, the strain of choice for neurological transgenic and targeted mice. PMID- 11111849 TI - Morphological features of neurons innervating different viscera in the cat stellate ganglion in postnatal ontogenesis. AB - Retrograde axonal transport of horseradish peroxidase (HRP) was used in this study to determine morphological parameters in the stellate ganglion (SG) in newborn, 10-, 20-day- and 1-month-old kittens. Neurons with the largest average size participated in innervation of the heart in newborn kittens and in innervation of the sternocleidomastoid muscle in other animals. The number of neurons sending their axons to target-organs also changed in postnatal ontogenesis. Regardless of the site of HRP injection at animals of all ages labeled neurons in the SG were located in certain zones on a topographical basis. Thus, it is concluded that in postnatal ontogenesis the neuronal organization of the SG changes in parallel to the increase of neuronal sizes and ganglion cross section area and practically finishes at 1 month of age. PMID- 11111850 TI - Axonal degeneration and regeneration in rat uterus during the estrous cycle. AB - Uterine innervation of the adult virgin rat changes throughout the estrous cycle. Nerves immunoreactive for the pan-neuronal marker protein gene product 9.5 and the sympathetic marker dopamine beta-hydroxylase are maximal at diestrus and minimal at estrus, whereas presumptive sensory and parasympathetic axons are unchanged. In the present study, we used quantitative electron microscopy to determine if depletion of immunoreactive nerves from the myometrium is due to loss of structurally intact axons, and whether this occurs through degeneration or retraction. Numbers of intact myometrial axons per unit sectional area were greatest at diestrus and least at estrus, while myometrial area was smallest at diestrus and greatest at estrus. However, depletion of intact axons at estrus was evident even after correcting for changes in uterine size. Varicosities adjacent to smooth muscle cells did not vary significantly with respect to their ultrastructural features or distance to the nearest smooth muscle target cell. Because retracting axons show increases in organelle content and distances to target cells, retraction probably does not play a major role in reducing uterine innervation. In contrast, axons with ultrastructural features consistent with degeneration (organelle and axolemmal disintegration, abnormal electron opacity, dense inclusion bodies) were significantly increased at proestrus and estrus. Growth cones were observed only at metestrus and diestrus. We conclude that cyclical degeneration and regeneration of myometrial innervation is a normal feature of the virgin adult rat. PMID- 11111851 TI - The political economy of nephrology. AB - An issue of Seminars devoted to "The Economics of Nephrology" requires consideration not only of the narrow economic issues affecting the specialty but also the public policies that establish its economic parameters. Some economic issues involve only the balancing of costs and revenues in a dialysis unit. Others turn on the ESRD policies of Medicare. Still others hinge on action by the US Congress and, by definition, are political in character. In nephrology, economics are intertwined with politics, hence the political economy of nephrology. PMID- 11111852 TI - A quarter century of medicare expenditures for ESRD. AB - Medicare's end-stage renal disease (ESRD) program is unique in that it is the only example of an entitlement program based solely on the basis of a clinical condition. Medicare payments on behalf of ESRD beneficiaries is a combination of ESRD-specific payment policies such as those for dialysis, physician oversight, erythropoeitin, and immunosuppression and general Medicare payment policies such as hospital payments, nondialysis physician services, home health, and skilled nursing care. Over the 25-year history of the program, much of the ESRD-related care has been subject to cost controls more stringent than elsewhere in Medicare. Total payments for ESRD beneficiaries continue to consume an ever-increasing percentage of Medicare expenditures, largely because of ever-expanding patient treatment criteria. However, increases in per capita expenditures for ESRD beneficiaries have been far below that of Medicare in general. PMID- 11111853 TI - The cost of clinical dialysis--a historical perspective. PMID- 11111854 TI - Dialyzer reuse: an evolving search for efficiency. AB - The evolution of dialyzer reuse in the United States provides an opportunity to examine the dialysis community's response to changing financial conditions and incentives. As background, we provide a conceptual framework to explain the factors governing the diffusion of dialysis technologies and then describe the clinical context in which reuse programs have developed. Early in its evolution, dialyzer reuse arose principally from the desire to reduce costs under a system of capitated payments. More recently, despite evidence of an adverse health effect, cost-savings from reuse have permitted the adoption of new and expensive technologies. The net effect of the tradeoff's between cost and quality should, ideally, drive the decision to reuse dialyzers. However, even if such tradeoffs can be fully characterized, incentives to implement efficiencies created by capitated systems of payment will continue to influence practice. PMID- 11111855 TI - Prescription drugs in Medicare and the ESRD program. AB - Prescription drugs have improved the length and quality of life for many persons, particularly those with chronic disease. However, their contribution to total health care costs has been increasing, making it difficult for patients to afford necessary medications. Current Medicare policy, derived from statutory enactments by Congress, precludes payment for prescription medications with the exception a few medications for end-stage renal disease (ESRD) patients. A more comprehensive prescription drug benefit for Medicare has been proposed that might benefit ESRD patients. Design and implementation of a medication drug benefit involves several issues including what medications should be covered, under what circumstances, how cost should be shared, the payment structure, level of payment, and management of the benefit. This report describes the experience with Medicare coverage of prescription drugs for ESRD patients and explores issues in offering a general medication insurance benefit under Medicare. PMID- 11111856 TI - The current and future state of the ESRD program: a provider roundtable. AB - The decade of the 1990s have seen substantial consolidation of services in the dialysis industry in the United States. A small number of horizontally and/or vertically integrated companies oversee the care of over two-thirds of dialysis patients. There are many questions regarding this trends as well as the vision of these large organizations regarding the future of the ESRD program. The senior physicians in the four largest such organizations agreed to participate in a provider roundtable to share their thoughts on the following issues: What are the advantages and disadvantages of industry consolidation?; What steps has your organization taken to succeed?; What are the key issues facing this industry in the next decade?; What policy changes by the Federal Government do you anticipate?; What policy changes would you like to see? Although significant differences in specifics are clear in the responses, a recurrent theme relates to how value will be maintained in the program-the balance between high-quality outcomes and the costs of achieving these outcomes. This is clearly the challenge in the years ahead. PMID- 11111857 TI - Economics of an academic renal division: an approach. AB - Nephrology divisions work within the systems and constraints of a department of medicine in a college of medicine, and in an increasing number of cases, also within a larger health care delivery system. Key to the division's stability are a consistently applied practice pan, how the tenure and clinical track systems function, how teaching funds are distributed, mechanisms of incentive for the division, and its faculty and the ability to establish and maintain reasonable financial reserves, especially for investment in research. Dialysis, transplantation, hypertension, research, and the teaching of medical students, residents, and fellows are the key elements the divisional organization must address. Incentives for faculty performance should be included in all of these areas. The division director must recognize his or her control of earning capacity in the distribution of responsibilities; financial returns as compared with effort involved vary considerably. Three main problems confront the future of our divisions: manpower shortages in nephrology; the effects of managed care systems on nephrology practice; and the future existence and success of departments of internal medicine and academic health centers. PMID- 11111858 TI - Challenges and directions for Medicare ESRD payment policy. AB - Since the inception of the ESRD program in 1973, Medicare has been challenged to provide access to high-quality care to beneficiaries with ESRD while trying to contain program payments. Despite implementing policies to control the growth in spending for outpatient dialysis and shifting the risk of certain ESRD beneficiaries to private payers, annual ESRD program payments have grown faster than overall Medicare spending. Some stakeholders contend that these policies have adversely affected beneficiaries' access to high-quality care. Refining the payment systems for caring for beneficiaries with ESRD in traditional Medicare and managed care plans may provide some respite to the growth in ESRD program spending in the short-term. In the long run, ESRD program spending may not be effectively controlled until changes are made in the delivery of health care services to this population. PMID- 11111859 TI - ICSI as an effective therapy for male factor with antisperm antibodies. AB - This study was conducted to evaluate if in vitro fertilization (IVF) with intracytoplasmic sperm injection (ICSI) is an effective treatment for infertility complicated by the presence in the male partner of sperm autoantibodies. Over a 1 year study period comparisons of fertilization, pregnancy, and implantation rates were made in couples where the male partner was negative or weakly positive for sperm autoantibodies (<50%) (gr 1); autoantibodies were strongly positive (>80%) (gr 2); or autoantibodies were moderately positive (50-80%) (gr 3). Only patients having oocytes fertilized by ICSI were included. The fertilization, clinical pregnancy, implantation, and miscarriage rate for group 1 (n = 67) was 56, 43, 21, and 14%. Comparable values for group 2 (n = 20) were 55, 40, 23, and 25%, and for group 3 (n = 6) were 63, 33, 23, and 0%. IVF with ICSI demonstrates comparable fertilization, pregnancy, implantation, and miscarriage rates in female partners of males with and without sperm autoantibodies. PMID- 11111860 TI - Nonsperm cells in human semen and their relationship with semen parameters. AB - The prevalence and clinical significance of leukocytes (WBC) and immature germ cells in semen is currently a matter of controversy. The aim of this work was to assess the prevalence of leukocytospermia in semen samples from Venezuelan men and its possible effects on sperm parameters. The concentration of WBC and round cells (RC) was evaluated in 118 semen samples from 19 fertile subjects (group 1), 62 infertile patients (group II), and 37 men with varicocele (group III). Semen WBC concentration was assessed by peroxidase assay. Twenty-six (22%) of the total samples had more than 10 WBC/mL semen. Twenty of the infertile men had leukocytospermia (32%) compared with 16% in the fertile group and 8% in the varicocele group. Semen RC concentration was lower than 5 x 10(6)/mL in all groups but, in groups II and III was significantly higher compared with group I. Infertile men had the highest WBC concentration. WBC concentration was negatively correlated with progressive motility, percentage of morphologically normal sperm, and hypoosmotic swelling test in infertile men but not in the varicocele group. In this group a negative correlation was obtained between immature germ cells and normal sperm morphology. The data show that leukcytospermia occurs frequently in infertile patients and is associated with poor semen quality parameters. In contrast, in men with varicocele, the increased number of immature germ cells might play a pivotal role in the pathogenesis of abnormal spermatozoa. PMID- 11111861 TI - Effect of neonatal administration of an antidopaminergic drug (metoclopramide) on sexual behavior of male rats. AB - Metoclopramide is an antidopaminergic drug that binds D2 receptor at the central nervous system and peripheral levels. Recent studies have demonstrated the presence of a sexually dimorphic nucleus in the anterior hypothalamus of rats and humans. The sexual differentiation of brain and its expression on sexual behavior is directed by catecholamines and steroids, which in turn will determine the masculinization of the preoptic area of the hypothalamus. Metoclopramide blocking dopaminergic receptor inhibits dopamine action and therefore brain masculinization. To determine the effects of the administration of this antidopaminergic drug in neonatal male rats, the design of the study considered measurements of mounts, penis intromission, and ejaculation in adult male rats treated neonatally with 100 microg metoclopramide. The administration of the drug or the vehicle (control) was by subcutaneous route during the third and fifth days of postnatal life. Six of 10 neonatally treated rats did not express male sexual activity in adulthood, while all control rats showed the characteristic pattern of male sexual behavior. Frequency of ejaculation in the rats treated with metoclopramide was 1.5 +/- 1 as compared to 5.0 +/- 0.9 in control rats (p < .01). There were meaningful differences with respect to the frequency of intromission in experimental rats (16 +/- 8) compared to control rats (115 +/- 18; p < .01). There is strong evidence that metoclopramide neonatally administered inhibits male sexual behavior in adult rats. Dopamine may be implicated in the process of neonatal brain sexual differentiation. PMID- 11111862 TI - Psychosocial response of Chinese infertile husbands and wives. AB - Some 15% or 1 in 6 American couples in the childbearing years have infertility problems. Numerous studies have demonstrated that both infertile men and women have negative emotional responses, such as stress, anxiety and depression. In Europe, Canada, and the United States the response of infertile husbands was different from that of their wives in self-image, marital adjustment, and sexual relations. The differences in psychological distress, marital satisfaction, and sexual satisfaction between Chinese infertile husbands and wives were evaluated. Fifty-nine infertile couples participated in this study. The subjects completed an Infertility Questionnaire, Marital Satisfaction Questionnaire, and Sexual Satisfaction Questionnaire as measures of gender differences in facing infertility problems. Paired t tests revealed that husbands expressed significantly less distress than that of the wives. The husbands' self-esteem was higher than that of the wives. The husbands' marital and sexual satisfaction was also higher than that of the wives. These results propose that although differences exist in cultural, ethnic, and religious norms between Chinese society and Western society, the Chinese couples' response to infertility is similar to that of Western couples. The major difference is that the in-laws play an important role in Chinese society, especially in marital satisfaction. PMID- 11111863 TI - Development of the coping scale for infertile couples. AB - Infertility treatment involves more complicated, uncomfortable, and humbling medical procedures for women, so the coping strategies used by wives may differ from those used by husbands. The purpose of this study was to develop and test a Coping Scale for Infertile Couples that would be sensitive to the differences in gender. What couples actually do to cope with infertility and whether these strategies are different between couples were also addressed. A total of 138 infertile couples participated in this study. The Coping Scale for Infertile Couples was administered with the Infertility Questionnaire, the Perceived Stress Scale, and the Jalowiec Coping Scale as measures of concurrent validity. Factor analysis extracted 4 subscales from the Coping Scale for Infertile Couples. Test retest and internal consistency reliability were high. Paired t tests revealed that wives used the "Increasing space and Sharing the burden" strategies to a greater degree than their husbands. Husbands used the "Being the best" strategy to a greater degree than the wives. A significant correlation with distress, stress, and coping measures provided evidence of concurrent validity. Preliminary results suggest that this measure has good reliability and validity, which can contribute toward the elucidation of coping strategies used by infertile couples and assist in planning effective interventions. PMID- 11111864 TI - Advances in male contraception. AB - The methods devised for male contraception are meager. The authors review the various nonhormonal methods applied for contraception including vas deferens interference and heat. The former comprises the no-scalpel vasectomy, percutaneous vasal injection, the "Shug" method, and the argon laser vasal photocoagulation. Heat methods used wet heat, and artificial cryptorchidism was created by testicular suspension. The testicle was suspended in the superficial inguinal pouch close to the scrotal neck using 2 methods: stitch and ball. Two recently developed methods for male contraception--polyester-induced azoospermia and prolactin injection--are described. The azoospermic effect of the polyester sling seems to be due to (1) creation of an electrostatic field across the intrascrotal structures, and (2) disordered thermoregulation. Prolactin administration as a contraceptive method is efficient and safe and has the potential to be developed as a male contraceptive. The methods, especially testicular suspension and polyester suspensors, are simple and easily applicable and were well accepted by the subjects. PMID- 11111865 TI - Immune responses in rats following oral immunization with attenuated Salmonella typhimurium expressing human sperm antigen. AB - The HSD-I gene codes a human sperm membrane protein (hSMP-1) and has been assigned the accession number U12978. The gene is located on human chromosome 9, region p12-p13. When the 1.7-kb cDNA of HSD-I was digested sequentially with EcoRI, BamHI, and HindIII, a 550-bp cDNA fragment was formed, which codes for the extracellular domain. This fragment was cloned into the asd+ vector pYA3149 to construct pYA3149R. The recombinant plasmid was used to transform an avirulent deltacva, deltacrp, deltaasd vaccine strain of Salmonella typhimurium chi4550. The hSMP-1 component was localized on the surface of the head of mature rat spermatozoa by an immunofluorescence technique using polyclonal anti-hSMP-1 antibodies. Since rat sperm contain hSMP-1, this rodent can be used to assay the immunogenicity of pYA3149R. Female Wistar rats were immunized by oral administration of the recombinant Salmonella. Anti-hSMP-1 antibodies in blood and vaginal washes of immunized animals were determined. Both body fluids contained significant amounts of the antibodies, showing that the recombinant Salmonella is an effective oral immunogen in rats. PMID- 11111866 TI - Pericentric inversion of the Y chromosome of infertile male. AB - The authors report a case with pericentric inversion of the Y chromosome associated with asthenonecrozoospermia. The conventional karyotype was 46, X, inv (Y) (p11q11). Polymerase chain reaction (PCR) analysis revealed the deletion of DYZ3, DYS139, and RBM1. Three-color fluorescent in situ hybridization (FISH) analysis of the sperm chromosomes showed normal ratio between X- and Y-bearing sperm. In this case, the frequencies of aneuploidy of the sperm are not significantly higher compared with those from the normal volunteers. Cytogenetic analysis is recommended when the patients with pericentric inversion of the Y chromosome are attending an infertility clinic. PMID- 11111867 TI - Low plasma testosterone in varicocele patients with impotence and male infertility. AB - To study the affect of bilateral varicocele (grade 3) on impotence and male infertility patients, 29 patients were selected from an outpatient clinic during 15 May 1998 to 15 August 1999 (the mean age was 33.9 +/- 6.3), 15 patients complaining of erectile dysfunction and 14 patients complaining of male infertility. The mean duration of impotence was 3 +/- 2.3 years and for male infertility was 6 +/- 2.5. All organic and psychogenic causes related to impotence and male infertility except bilateral varicocele (grade 3) and low plasma testosterone were excluded by clinical and laboratory investigations. Twenty males with normal erection and fertility were included as controls. Detailed medical history and complete physical examination included measurement of testicular size by orchiometer; semen and hormonal parameters were measured for all patients and control. In impotent patients left and right testicular volume was significantly decreased (p < .05), while in male infertility patients left and right testicular volume was highly significantly and, significantly decreased (p < .005, p < .05) compared to controls. In male infertility patients, left testicular volume was highly significantly decreased compared to impotent patients (p < .005). The sperm count and semen volume in impotent patients was significantly decreased (p < .05, p < .01), but no significant differences were found in sperm motility and abnormal forms, while in male infertility the sperm count was highly significantly decreased (p < .005), the sperm motility was significantly decreased (p < .05), the abnormal form was significantly increased (p) < .05), but in the semen volume there was no significant difference compared to controls. In impotent patients the sperm count was significantly increased and abnormal form was significantly decreased compared to male infertility (p < .05). The mean serum testosterone was significantly decreased in impotent patients (p < .01), and highly significantly decreased in male infertility (p < .005) compared to controls. The mean serum FSH was significantly increased in male infertility (p < .05) and nonsignificant in impotent patients compared to controls. The mean serum LH and prolactin levels were nonsignificant in both impotent and male infertility patients compared to controls, but LH was significantly increased in impotence compared to male infertility patients (p < .025). Therefore, bilateral varicocele (grade 3) is associated with significant reduction in testicular function with significant increase in serum levels of FSH and LH, which may cause erectile dysfunction and male infertility. PMID- 11111868 TI - Histochemical study of human cremaster in varicocele patients. AB - Despite the cremaster's important role in thermoregulation, few morphological and biochemical studies of this muscle in humans have been reported, probably due to limitation of sampling. To gain further insight into the pathology of varicocele, the authors studied the histochemical changes of the cremaster from patients with varicocele. Cremaster was obtained from patients with male infertility and varicocele, grades 1-3. The samples were studied using routine histochemical stains. Fiber size variability and type I predominance were observed in all varicocele cases regardless of the grade, and also in control specimens. Muscle from patients with grades 2 and 3 varicocele showed small group atrophy. It would appear that the hemostasis associated with local tissue edema and hypoxemia may lead to nerve damage and denervation of the cremaster. If denervation of the cremaster persists despite the correction of varicocele, thermoregulation would remain disrupted. PMID- 11111869 TI - Human leukocyte antigen II expression in sperm cells: comparison between fertile and infertile men. AB - Human leukocyte antigens (HLA) class II transcripts in mature spermatozoa of healthy volunteers have recently been demonstrated using reverse transcription polymerase chain reaction (RT-PCR). HLA II expression was investigated on ejaculated sperm cells in fertile and infertile men by RT-PCR and flow cytometry. Among 22 fertile and 20 infertile men, 18 were selected for the study because they showed no contamination with non-sperm cells. HLA II mRNA transcripts were expressed in all but 1 of 8 infertile subjects and in only 2 of 10 fertile ones. The cytofluorometric analysis on three RT-PCR positive samples confirmed the presence of class II antigens on cell surfaces. These data clearly confirm the presence of both HLA II mRNA and surface molecules on human sperm cells. In addition, an interesting nonrandom distribution of positivity among fertile and infertile samples regarding HLA II expression (p < .025) suggests a possible correlation with infertility. PMID- 11111870 TI - Sperm chromatin. AB - Available data on dry and hydrated nuclear volume of mammalian spermatozoa indicate that available volume is clearly insufficient to contain sperm chromatin packed in nucleosome-like structures. Therefore, sperm DNA-protein complexes must be packed differently than somatic DNA-protein complexes. Packing of DNA in fixed, dehydrated mammalian sperm approaches the physical limits of molecular compaction, making mammalian sperm chromatin the most condensed eukaryotic DNA known. The fundamental packaging unit of sperm chromatin is a toroid approximately 900-A outer diameter. 200-A thickness, and 150-A diameter hole. Each toroid contains 60 kilobases of DNA and is linked to other toroids by uncoiled DNA stretches. The factors that contribute to mammalian chromatin structuration are still under study. The role of protamines in sperm chromatin condensation and nuclear shaping has been overstressed to the exclusion of other possible factors. Chromatin organization in sperm nuclei is maintained during sperm condensation by tight interactions with the nuclear matrix at fixed sites, inducing the formation of individual toroid-shaped DNA loop stuctures. Observations that abnormal manchettes affect sperm head shape and chromatin organization inducing sterility speak about manchette importance during chromatin organization. The presence in sperm chromatin of regions packaged in specific ways with several types of protamines or even with histones, indicates that nuclear shaping and chromatin organization must be under DNA control. The structural properties that distinguish sperm DNA from somatic DNA may play the most important role in chromatin organization. PMID- 11111871 TI - Antibodies to human sperm YLP12 peptide that is involved in egg binding inhibit human sperm capacitation/acrosome reaction. AB - Recently, the authors reported a novel dodecamer peptide sequence, designated as YLP12 on human sperm, that is involved in binding to zona pellucida (ZP) of human oocyte [10]. This unique sequence is present on the acrosomal region of the human sperm cell and is expressed only in human testis/ sperm. The aim of the present study was to examine whether YLP12 sequence is involved in capacitation/acrosome reaction. Swim-up sperm were capacitated with anti-YLP12 Fab' antibodies or control Fab's (40 and 85 microg/mL) and then the acrosome reaction was induced with calcium ionophore. An average of 64-73% sperm underwent acrosome reaction when they were capacitated in the presence of 40-85 microg/mL of bovine serum albumin or control Fab's. A significant (p < .01 to < .001) reduction (58-75%) in the percentage of acrosome-reacted sperm was observed when the sperm were capacitated in the presence of YLP12 Fab's. These data indicate that the YLP12 peptide sequence is involved in sperm capacitation / acrosome reaction, and may find clinical applications in the diagnosis and treatment of male infertility and immunocontraception. PMID- 11111872 TI - Effects of cryopreservation on penetration ability of human spermatozoa into bovine cervical mucus. AB - Cryopreserved sperm exhibit lower fertilizing capacity in comparison to fresh sperm, partly due to effects of glycerol as the common cryoprotectant medium. Since standard semen analysis is not a good predictive method to assess sperm fertilizing capacity, functional tests like cervical mucus penetration may provide more useful information. A total of 24 semen samples were examined before and after cryopreservation for sperm parameters as well as number and motility of penetrated sperm into bovine cervical mucus (BCM) as an alternative for human cervical mucus. Freezing and thawing procedures have negative effects on sperm penetration into cervical mucus. No significant relation was noticed between sperm motility percentage or its penetration into BCM before and after cryopreservation, which denotes the variability in resistance of sperm to damaging effects of freezing. PMID- 11111873 TI - Human sperm membrane protein (hSMP-1): a developmental testis-specific component during germ cell differentiation. AB - Serum was obtained from an infertile woman having antibodies with sperm agglutinating activity. The antibodies interacted with a human sperm membrane protein (hSMP-1) with an estimated Mr of 55 kD. The gene (HSD-1) coding hSMP-1 was isolated from a human testis cDNA expression library and assigned the accession number U12978. The cDNA was conjugated to a prokaryotic expression vector to construct the recombinant vector, pRSET-HSD-I, which was expressed in Escherichia coli. The recombinant hSMP-1 was isolated and used to immunize rabbits to raise polyclonal antibodies. Usingan immunocytochemical technique, hSMP-1 protein was immunolocalized in germ cells of human testis at all stages of spermatogenesis. mRNAs were prepared from 16 different human tissues and analyzed by Northern blot using HSD-1 as probe. A positive reaction was elicited only with testis mRNA. The present findings suggest that the expression of hSMP-1 gene is testis-specific and occurs during the early stages of germ cell differentiation. In a comparative study, the location of the hSMP-I protein in sperm and in germ cells of the seminiferous tubules of rats was determined. The target antigen was immunolocated on the head and tail of rat sperm and in late spermatids and spermatozoa of rat testis. These results suggest that, in the rat, the HSD-1 gene is expressed during spermiogenesis. PMID- 11111874 TI - Controversies in the management of abnormal Pap smears. AB - Cytology is not 100% accurate. Overdiagnoses and underdiagnoses occur. Changes in classification systems and new devices have not changed the reliability of the Pap smear. The majority of squamous intraepithelial lesions will remit spontaneously and less than 1% will progress to invasive cancer. HPV-DNA testing has not provided a foolproof way to triage lesions. Therefore, the management of abnormal Pap smears is dependent on the personal experience of the physician, who must consider the following questions. What is the accuracy of your cytology laboratory? Does it overdiagnose or underdiagnose? How often has an ASCUS or LSIL become HSIL or invasive cancer on histologic evaluation? Will your patients come back for follow up? Will their health insurance cover frequent follow-up Pap smears or colposcopies? Do you have the resources to follow every patient who needs it? Do you want to avoid the development of every invasive cancer that may develop during the period of follow up? PMID- 11111875 TI - Adolescent sexuality and pregnancy. AB - The adolescent pregnancy rate has decreased throughout the USA. However, compared with other industrialized countries, US rates remain high. Efforts to decrease the number of teenage pregnancies are centered on sex education, postponing sexual activity, and safe sex practices. PMID- 11111876 TI - Physical and sexual abuse against women and children. AB - There is an increasing awareness of physical and sexual abuse against women and children. However, standardization of research is needed in order to facilitate comparison among studies on abuse. Most of the research has focused on prevalence, incidence and causes of domestic violence, family violence, and physical and sexual abuse. There is also a need to evaluate the effect of different intervention programmes. It is important that the education system, the health sector and the judicial system are motivated to recognize that they have not only the opportunity, but also the responsibility to take action against physical and sexual abuse. A multidisciplinary and multiagency perspective is needed in approaching this issue. All of us, especially educators and clinicians, are in a position to address abuse, and ultimately to reduce violence in our society. Abuse is everyone's business. PMID- 11111877 TI - Management of HIV infection during pregnancy. AB - Optimal management of HIV infection in pregnancy requires maternal use of potent antiretroviral therapy to prevent disease progression in the mother and vertical transmission to the newborn. Combination antiretroviral therapy substantially reduces the risk of perinatal HIV transmission and appears to be more effective than zidovudine monotherapy. The administration of single dose nevirapine to mother intrapartum and infant postpartum effectively reduces vertical HIV transmission and is less costly and cumbersome than zidovudine regimens. Elective cesarean section reduces vertical transmission of HIV but its benefit is less clear when antiretroviral therapy decreases maternal plasma HIV viral load to low levels at delivery. If possible, HIV-infected mothers should avoid breastfeeding. The present review discusses the importance of early identification of maternal HIV infection, strict adherence to combination antiretroviral regimens to prevent drug resistance, developing a better understanding of antiretroviral pharmacokinetics in pregnancy and short/long term safety of anti-HIV drugs. PMID- 11111878 TI - Ectopic pregnancy: an update. AB - This review addresses recent publications that investigate etiology, epidemiology and different modalities in diagnosis and therapy for ectopic pregnancy. A significant proportion of recent work has focused in the development of new diagnostic tools to aid in the early detection of ectopic pregnancy. Diagnostic modalities have included systemic and local markers, vascular endothelial growth factor, vascular cell adhesion molecule-1, urokinase plasminogen activator receptor, cervical fetal fibronectin, and hormonal level determinations. In addition, magnetic resonance imaging, ultrasonography, color flow mapping and endometrial thickness have been evaluated. New studies have investigated controversial issues related to the cost of the medical versus surgical treatment and the use of different medications and techniques for the management of ectopic pregnancy. Most important of all, several lines of investigation have addressed the use of human chorionic gonadotropin, algorithms, and scoring systems as prognostic indicators of successful therapy and to determine the risk of complications. The management of cervical, interstitial and heterotopic pregnancy is evaluated in this review and a summary of recent proposed diagnostic tools and concepts in management is also presented. PMID- 11111879 TI - Endometriosis-associated infertility. AB - This review summarizes the recent literature examining the relationship between endometriosis and infertility. It is clear that the advanced stage of the disease and the mechanical disruption of the pelvic anatomy may cause infertility. The link between early stage endometriosis and infertility remains a source of controversy. Management plans must be individualized contingent upon the stage of disease, the age of the patient and the duration of infertility. The preponderance of data suggests that ablative therapy at the time of laparoscopy is as good as, or superior to expectant or medical therapy. With the exception of IVF/ET, ovarian suppression with GnRH agonists is not warranted in endometriosis associated infertility. Controlled ovarian hyperstimulation with IUI is appropriate therapy in women with minimal-to-mild and surgically corrected endometriosis. PMID- 11111880 TI - Dynamics of voiding in women. PMID- 11111881 TI - Neuronal innervation of urethral and anal sphincters: surgical anatomy and clinical implications. AB - The present review describes the neuronal innervation of the external urethral and anal sphincters. A knowledge of this innervation helps in understanding the clinical symptoms of urinary and anorectal pathology, and in choosing the appropriate technique of nerve localization or block. An ability to locate the pudendal nerve, on the basis of surgically documented anatomy, has important diagnostic and therapeutic advantages. It can be used to study the integrity of pelvic floor muscles, in biofeedback training, nerve blocks, pudendal canal decompression, chronic stimulation trials to treat urinary or faecal incontinence, and in nerve conduction studies or evoked potential recordings. Furthermore, the superficial location of the sphincteric innervation in the perineum and ischiorectal fossa renders the nerve branches susceptible to injury during operative correction of urinary or faecal incontinence. Supported by a knowledge of anatomy, we can make firm recommendations on which to base safe surgical techniques that avoid damage to urethral and anal sphincteric innervation. PMID- 11111882 TI - Recent developments in pelvic organ prolapse. AB - Pelvic organ prolapse is a common worldwide problem. Recent advances in our understanding of its pathophysiology, along with progress made in the evaluation and treatment of pelvic support defects, are discussed. Although the pathophysiology of this condition is still not completely understood, genetic factors and environmental factors are involved. Understanding these factors better will help us to approach treatment of pelvic organ prolapse in a more logical manner. Multiple surgical techniques are available for pelvic relaxation, with a wide range of success rates ranging from 77 to 97% for various procedures. New techniques need to be studied further before being incorporated into routine practice. Better standardization of evaluation methods can help in such clinical studies. PMID- 11111883 TI - Pathophysiology of interstitial cystitis. AB - Despite being described over 80 years ago, interstitial cystitis remains a disease of undetermined aetiology and poor treatment outcomes. Generally agreed diagnostic criteria of this condition, which occurs primarily in females, are frequency, urgency and pain, a low-capacity hypersensitive bladder, and mucosal haemorrhages and tearing on bladder distention. Although current theories of pathophysiology are predominantly conjecture, important elements of the disease process are increased afferent and efferent neuronal activity, an excess of inflammatory mediators, increased epithelial permeability and possibly reduced bladder vascularity. Improved treatment outcome will follow a better understanding of pathophysiology. PMID- 11111884 TI - Biofeedback in urinary incontinence: past, present and future. AB - Recent systematic reviews on the conservative management of urinary incontinence in women have identified a small number of trials that favour conservative management, but the evidence for firm recommendations to include biofeedback in these conservative strategies is lacking and further research is needed to clarify the role of biofeedback. PMID- 11111885 TI - Current status of urethral occlusive devices in management of urinary incontinence. AB - Treatment of outlet incontinence with urethral occlusive devices encompasses simple and complex modalities. The implantable artificial urinary sphincter has been used for 25 years, and a growing fund of clinical information is available to define its indications. Other simple occlusive devices are available that provide an effective, nonsurgical and reversible treatment. The present review summarizes recent clinical investigations into the safety and efficacy of urethral occlusive devices. PMID- 11111887 TI - Bibliography. Current world literature. Adult and pediatric gynecology. PMID- 11111886 TI - Genuine stress incontinence: colpocystourethropexy versus sling procedures. AB - Both colpocystourethropexy (colposuspension) and sling operations have been shown to be effective in treating female stress incontinence. The present review discusses the literature available and compares the results and complications of both procedures. Colposuspension can give excellent results as both primary and secondary surgery. Slings also give excellent results, but are prone to complications relating to the sling material and postoperative voiding difficulties. Slings are arguably best reserved for women in whom vaginal scarring makes colposuspension impossible. Colposuspension remains the gold standard operation against which new techniques should be compared. PMID- 11111888 TI - Bibliography. Current world literature. Urogynecology. PMID- 11111889 TI - The suspensory ligament of the clitoris: connective tissue supports of the erectile tissues of the female urogenital region. AB - We aimed to define the gross anatomy of the supporting structures of the clitoris. We performed a dissection of the perineum of a series of 22 female and four male cadavers. Specific dissection of the clitoral and penile suspensory ligament complex was performed in four female and two male cadavers. Serial written observations and photography were used to document the findings. Our findings were then compared with the anatomical description of these structures in the historical and current anatomical literature. The suspensory ligament of clitoris consistently displayed two components: a superficial fibro-fatty structure extending from a broad base within the mons pubis to converge on the body of the clitoris and extending into the labia majora: in addition there is a deep component with a narrow origin on the symphysis pubis extending to the body and the bulbs of the clitoris. The supporting structures of the clitoris are more substantial and complex than previously described. Their shape, extent, and orientation are different from analogous structures of the penis, the suspensory ligament of which was found as described in the literature. PMID- 11111890 TI - Postprandial decrease in splenic volume demonstrated by magnetic resonance imaging and stereology. AB - The aim of this study was to determine if the volume of the spleen changes after food intake. We applied an unbiased and efficient method for splenic volume estimation using magnetic resonance imaging (MRI) in combination with modern design stereology. MR images of the spleen were obtained for 10 healthy volunteers (five men and five women; mean age 28.9 years [range 23-35 years]) without a history of splenomegaly. The initial scans were performed in the morning after overnight fasting. Each volunteer then consumed a standard balanced meal weighing 500 g [2,460 kJ (627 kcal) energy] with 500 ml of still mineral water. Second identical MR scans were performed approximately 1 hr later. Postprandially, splenic volume decreased by an average of 6.6% (P = 0.005), probably due to increased splanchnic blood flow after food intake. PMID- 11111891 TI - Antireflux mechanisms in veins draining the upper territory of the vertebral column and spinal cord in man. AB - The terminal segments of vertebral, right supreme intercostal and accessory hemiazygos veins from 28 cadavers of various ages were histologically examined to assess the possible presence of intrinsic mechanisms facilitating the delivery of venous blood from the upper vertebral and spinal cord territories. In these veins, blood flow is assisted by gravity (veins of receptive type); however, the tunica adventitia showed a significant amount of longitudinally oriented bundles of smooth muscle fibers together with elastic fibers, i.e., an intrinsic structure similar to that of propulsive type veins. Both the muscular and elastic components increased with age. One or two complete valves were found at the opening of these veins into their major receptors. It can be assumed that the structural changes of the venous wall and the ostial valves play a role in avoiding or limiting undue reversion of the blood flow in many physiologic and pathologic conditions, and in protecting the delicate nervous structures of the vertebral canal from possible damages due to temporary or chronic venous stasis. PMID- 11111892 TI - The minor hepatic veins: anatomy and classification. AB - A detailed description of the distribution and drainage pattern of the minor hepatic veins is presented in this paper. A classification based on the segmentation of the liver divides these veins into four main groups: 1) veins of Segment I which includes the veins of the caudate lobe and the veins of the caudate process; 2) veins of Segment VI; 3) veins of Segment VII; and 4) veins of Segment IX. A knowledge of the anatomy of the minor hepatic veins becomes more clinically valuable as the number of complex dissections of the retrohepatic areas, hepatectomies. and hepatic transplantations grow. PMID- 11111893 TI - Relationship between extracellular matrix both in choroid plexus and the wall of lateral ventricles and intraventricular hemorrhage in preterm neonates. AB - Pathogenesis of intraventricular hemorrhage (IVH) may relate to immature vessels. However extracellular matrix--such as Type IV collagen, laminin, and fibronectin has a direct bearing on the development of the endothelium and the immature microvessel. Twenty specimens taken from lateral ventricular walls and choroid plexus in 10 preterm neonates who died of intraventricular hemorrhage and another 10 preterm neonates who died of non-intraventricular hemorrhage, were marked with antibodies against Type IV collagen, laminin, and fibronectin. Average areas of positive staining around the microvessels in the IVH group showed less staining than that in the non-IVH group (P < 0.05), and the microvessel count of discontinuous positive staining in IVH group was more than that in non-IVH group, P < 0.05. We conclude that lacking Type IV collagen, laminin, and fibronectin may result not only in an incomplete basement membrane in choroid plexus and subependymal immature microvessels but also may be a factor contributing to IVH in preterm neonates. PMID- 11111894 TI - Thickness of the stratum corneum of the volar fingertips. AB - The thickness of the stratum corneum was measured by optical coherence tomography at the center and sides of the tactile elevations of all fingers in 87 healthy volunteers and 18 people with diabetes who performed regular glucose self control. The cornified epidermis was thickest at the thumbs, and thickness decreased toward the little finger. The cornified epidermis was thinner at the sides of the tactile elevations than at the center, and it was thinner in women than in men. In people with diabetes, the cornified epidermis of the fingers most frequently used for capillary blood sampling was not conspicuously thickened. PMID- 11111895 TI - Anatomical study of the accessory head of the flexor pollicis longus and the anterior interosseous nerve in Asians. AB - Anterior interosseous nerve palsy is known to occur uncommonly due to the compression of the nerve by the accessory head of flexor pollicis longus (AHFPL). This study was conducted to investigate the prevalence and origin of the AHFPL and the topographical relationship between the AHFPL and the anterior interosseous nerve in Asians. The AHFPL was present in 48 of 72 arms examined (66.7%), a majority of which originated from the coronoid process. The anterior interosseous nerve was observed to arise from both the medial and posterior aspects of the median nerve. The topographical relationship between the anterior interosseous nerve and the AHFPL was classified into three types depending on if the anterior interosseous nerve crossed the muscular part, or the tendinous part of the AHFPL, or coursed lateral to the AHFPL. The case in which the anterior interosseous nerve crossed the muscular part of the AHFPL occurred most frequently in the current study. The types in which the anterior interosseous nerve may be compressed were also discussed. PMID- 11111896 TI - Approaches to learning spatial relationships in gross anatomy: perspective from wider principles of learning. AB - When students learn spatial relationships in gross anatomy, as in other areas of study, fundamentals should be learned first; otherwise confusion results. The fundamentals in gross anatomy are defined not in conceptual terms but by principles of visual perception. In particular, they derive from Gestalt principles such as collinearity and symmetry, which generally make learning and recognition of visual patterns easier. The collinearity (straight line formations) and symmetry in the body cavities are obvious when one studies the empty cavities, or body cavities with only a few symmetrical structures in place. These principles are, however, totally obscured if one starts one's study, as in traditional dissection, with the body cavities crammed full of a complex mass of interlocking organs. and their ducts, vessels, etc. Therefore, it is recommended that learning gross anatomy (especially of the body cavities) would be an easier exercise if it started with empty body cavities, then building up, with a careful sequence of prosections, to the more complex and realistic anatomy of the full cavities. This system of learning is perceptually preferable to traditional dissection. However, it needs to be enlivened in several ways, e.g., with respect to design principles evident in anatomical structure (especially for the musculoskeletal system), developmental processes, and sometimes by explicit reference to clinical relevance. PMID- 11111897 TI - Learning gross anatomy in a clinical skills course. AB - Recent developments in undergraduate medical education in the United Kingdom have produced changes in the content and delivery of component courses, including human anatomy. Anatomy can retain its place in the medical course in the new world of problem-based learning and clinical skills teaching by gaining recognition as an integral part of the curriculum which underpins much of the practice of clinical medicine. In these new courses, anatomical information is clinically relevant and discussed in the context of medical problems and the acquisition of clinical skills. Students are encouraged to study in a manner in which information is retained (deep learning) and where understanding replaces rote learning of facts. Students take responsibility for their own learning, with appropriate support and resources. In clinical skills courses, anatomy underpins the development and retention of clinical knowledge and skills. PMID- 11111898 TI - The role of three-dimensional information in health care and medical education: the implications for anatomy and dissection. AB - The purposes of medical education can be summarized as learning how to take an effective history, perform a physical examination, and perform diagnostic and therapeutic procedures with minimal risk and maximal benefit to patients. Because patients are three-dimensional (3-D) objects, health care and medical education involve learning and applying 3-D information. The foundation begins in anatomy where students form and confirm or reform their own 3-D ideas and images of the development and structure of the human body at all levels of organization. Students go on to understand the interdependence of structure and function in health and disease. The basic questions for those teaching anatomy are "How do we learn and use 3-D information?" and "How is it taught most effectively?" These are not easy questions for teachers and are rarely asked by those who currently defend or reframe curricula. Unfortunately, there is little information on how we learn 3-D information and no evidence-based literature on the relative long-term vocational effectiveness of methods for teaching it. It is clear that we learn in several distinct modalities and that our students represent a spectrum of learning styles. To support the 3-D learning essential to both medical education and health care, anatomical societies need to provide answers to the following questions: Do the opportunities of dissection (visual, tactile, time, discovery, group process, mentoring) contribute to short- and long-term learning of 3-D information? If so, how? Does dissection offer significant advantages over other methods for learning, confirming, and using 3-D information in anatomy? Answers to these questions will provide a rational basis for decisions about curricular changes in anatomy courses (if, where, and when dissection should occur). This, in turn, will link these changes to society's ultimate purposes for medical education and health care rather than to the fiscal concerns of the businesses of health care and medical education, which is the current practice. PMID- 11111899 TI - Avoiding the postdoctoral glut: an alternative route to a career in academia. AB - Dramatic increases in the number of PhDs granted during the last several decades have not been balanced by comparable increases in the number of academic positions, producing a glut of postdoctoral fellows. One major reason for the burgeoning numbers of postdoctoral fellows is a widespread perception that there is no alternative route to a career in academia other than through gaining expertise in bench research. This paper suggests that considerable numbers of postdoctoral fellows who are interested in securing a tenure-track position at a major medical or research university should consider directing their postgraduate training toward teaching. Teaching at universities is becoming a specialized field partly because researchers are unable to keep up with the explosion of information, new instructional theories, methodologies, and technology. Thus, some individuals who are currently in a postdoctoral holding pattern could use those years to increase their teaching competence. Such an "educational" postdoctoral position may offer a faster route to a career in academia as universities continue to recognize the need for teaching specialists. PMID- 11111900 TI - Bilateral persistent complete sciatic artery. AB - A case with persistent sciatic artery (PSA) was found in a cadaver of a 65-year old female during a medical gross anatomy course. The artery was bilateral and complete and provided the major blood supply to both lower extremities. The vessel arose from the internal iliac artery that was extremely large bilaterally. The sciatic artery passed out of the pelvis through the infrapiriform foramen and descended posterior to the sciatic nerve through the gluteal region. The sciatic nerve was considerably flattened out under the artery. Large articular branches arose from the part of the artery at the buttock just below the piriform muscle. The artery descended along the back of the thigh and was crossed obliquely posteriorly by the long head of the biceps femoris muscle. The sciatic artery continued as the popliteal artery located very superficially in the popliteal fossa. A companion vein, i.e., the sciatic vein, accompanied each artery. The right sciatic vein entered the pelvis posteriorly through the infrapiriform foramen, whereas the left perforated the quadriceps muscle from behind and joined the femoral vein anteriorly. There was a gracile superficial femoral artery bilaterally. The deep femoral arteries of both lower limbs were hypoplastic with slender circumflex branches. There were no macroscopic connections between the sciatic and the deep or superficial femoral arteries on either side. This anomaly should be kept in mind in the evaluation of patients with sciatic or buttock pain or palpable "pulsating" buttock mass. Persistent sciatic artery also could be a potential hazard during orthopedic manipulations, hip joint surgery, and renal transplant surgery. PMID- 11111901 TI - Regulation of toxin and virulence gene transcription in Bacillus thuringiensis. AB - Bacillus thuringiensis is a spore-forming bacterium well known for its insecticidal properties and its ability to produce a crystal inclusion during sporulation. The specific activity of B. thuringiensis against insect larvae is due to the crystal proteins (Cry proteins). Two different transcriptional mechanisms (dependent and independent of sporulation) are responsible for cry gene transcription during the stationary phase. In addition to these specific insecticidal toxins, B. thuringiensis produces potential virulence factors including haemolysins, degradative enzymes and enterotoxins. A pleiotropic regulator (PlcR) that activates the transcription of various genes encoding such extracellular proteins has been identified. Its expression at the onset of the stationary phase is dependent on the growth medium and is controlled by the transition state regulator, SpoOA. PMID- 11111902 TI - Genomics of Bordetella pertussis toxins. AB - Bordetella pertussis, the etiologic agent of whooping cough, produces numerous toxins including pertussis toxin (PTX), adenylate cyclase toxin (AC), dermonecrotic toxin (DNT) and tracheal cytotoxin (TCT). PTX is composed of five different subunits organised in a typical A-B type structure of which the A part possesses an enzymatic ADP-ribosyltransferase activity and the B moiety expresses receptor-binding activity. The secretion of this toxin requires nine other genes (ptl) organised in an operon together with the five structural genes of PTX. To further characterise the genetic locus of this major virulence factor, we analysed the ptx/ptl upstream and downstream sequences. Comparison of these regions between three species of Bordetella (B. pertussis, Bordetella parapertussis and Bordetella bronchiseptica) revealed differences in the upstream region. Analysis of two strains of B. bronchiseptica naturally lacking the ptx genes showed that only the ptx/ptl genes were deleted in these strains, and that the upstream and downstream regions were conserved. Upstream of the PTX structural genes and the promoter, an open reading frame (bugT) was identified, the product of which is homologous with putative proteins from several other Gram negative organisms. Detailed analysis of the genome of B. pertussis which is currently sequenced at the Sanger Centre revealed the presence of 90 genes coding for proteins homologous to BugT, which qualifies the bug gene family as the most populated one of Bordetella. These bug genes are located in various genetic environments, including the proximities of genes coding for other toxins, such as DNT and AC. The Bug proteins are highly conserved in terms of size and periodicity of predicted secondary structure elements, but have also a high variability in their amino acid composition reflected in their wide range of isoelectric points. The function of these genes which is currently unknown is under investigation. To characterise the expression and regulation of these genes, as well as of novel putative B. pertussis virulence factors, we designed a transcriptional fusion vector to be inserted in precise locations of the B. pertussis chromosome by homologous recombination. The reporter gene present in this vector allowed us to show that at least some of the bug genes are expressed. PMID- 11111903 TI - Toxin genes on pathogenicity islands: impact for microbial evolution. AB - Toxin-specific genes are often located on mobile genetic elements such as phages, plasmids and pathogenicity islands (PAIs). The uropathogenic E. coli strain 536 carries two alpha-hemolysin gene clusters, which are part of the pathogenicity islands I536 and II536, respectively. Using different genetic techniques, two additional PAIs were identified in the genome of the E. coli strain 536, and it is likely that further PAIs are located on the genome of this strain. Pathogenicity islands are often associated with tRNA genes. In the case of the E. coli strain 536, the PAI-associated tRNA gene leuX, which encodes a minor leucyl tRNA, affects the expression of various virulence traits including alpha hemolysin production. The exact mode of action of the tRNA5Leu-dependent gene expression has to be identified in the future. PMID- 11111904 TI - Characterization of a plasmid region involved in Bacillus anthracis toxin production and pathogenesis. AB - The germination of spores within the host is the initial step of anthrax infection. We have shown, using immunofluorescence staining, confocal scanning laser microscopy and image cytometry analysis, that the alveolar macrophage is the primary site of B. anthracis germination in a murine inhalation infection model. B. anthracis germinated inside macrophages, in vesicles derived from the phagosomal compartment. We have demonstrated that the toxin genes and their trans activator, AtxA, are expressed within the macrophages after germination. It was also shown that the pXO1 plasmid strongly enhanced capsule formation and that this influence is mediated by AtxA. This indicates the existence of a regulon where AtxA is the regulatory protein acting on genes located on different plasmids. We identified a tricistronic germination operon gerX located between the pag and atxA genes on the 40-kb toxin-encoding fragment of pXO1 . Analysis of a gerX null mutant indicated that gerX-encoded proteins are involved in the virulence of B. anthracis. PMID- 11111905 TI - Structure-function relationships in the Bvg and Evg two-component phosphorelay systems. AB - The unorthodox two-component phosphorelay systems BvgAS and EvgAS of Bordetella pertussis and E. coli, respectively, are suitable model systems to investigate the molecular basis of signalling specificity, because, despite their high relatedness on the sequence level, they do not cross-talk to each other. We could show that the two systems belong to the obligate type of phosphorelay systems and that signalling specificity is mediated by the HPt modules of the histidine kinases and the receiver domains of the effector proteins. To gain more insight into signalling specificity on the molecular level, we started a detailed structural analysis of the respective proteins using a combination of genetic and biochemical methods including limited proteolysis and chemical modification of purified proteins and their mass spectrometrical analysis. PMID- 11111906 TI - Protein secretion mechanisms in Gram-negative bacteria. AB - Gram-negative bacteria have developed a variety of secretion pathways to secrete toxins and enzymes into the extracellular medium. These pathways are very different with respect to their functional mechanism and complexity, and each system has its own advantages and limitations, regarding the number, size, folding state and fate of their substrates. Pseudomonas aeruginosa secretes many different proteins into the extracellular medium, using at least four secretion pathways. Most of the exoproteins are secreted via the type II system, composed of the 12 Xcp proteins. The only outer membrane protein of the system, XcpQ, belongs to a large family of proteins, designated secretins, which participate in a variety of different transport processes. Other Xcp proteins, XcpT-X, show homology to the subunits of the retractile type IV pili. Further analogies between the type II system and the assembly of retractile pili suggest a mechanism for type II secretion, in which a pilus-like structure, composed of XcpT-X, facilitates the transport of exoproteins through the channel formed by the secretin XcpQ. PMID- 11111907 TI - Adenylate cyclase toxin from Bordetella pertussis: current concepts and problems in the study of toxin functions. AB - Adenylate cyclase (AC) toxin produced by Bordella pertussis and other Bordella species is a virulence factor and protective antigen with novel properties and activities, which make it attractive as a prototype toxin for study of membrane insertion and delivery to the target cell interior. It is unique among RTX toxins in that it possesses enzymatic (adenylate cyclase) activity, as well as the capacity to create an ion-permeable pore in target cell membranes and lyse erythrocytes. The current issues in understanding AC toxin, which will be discussed here, include the role of acylation in its various activities and the relationship among those several toxin functions. PMID- 11111908 TI - Colicin import into Escherichia coli cells requires the proximity of the inner and outer membranes and other factors. PMID- 11111909 TI - Characterization of the multimeric Eps complex required for cholera toxin secretion. AB - Vibrio cholerae causes diarrheal disease through colonization of the small intestine. A critical aspect of V. cholerae pathogenesis is its ability to actively secrete cholera toxin to the extracellular environment. This occurs via the type II secretion pathway, where the toxin subunits are first transported to the periplasm through the Sec pathway. Following folding and assembly the toxin is then translocated across the outer membrane by a specialized Extracellular Protein Secretion (Eps) machinery encoded by at least 13 genes. Although the Eps proteins are believed to form a secretion apparatus that spans both membranes, cholera toxin is thought to engage this complex first in the periplasm. In order to determine the organization of the Eps apparatus and to understand the mechanism of secretion, the Eps apparatus has been dissected and three of the components, EpsE, EpsL and EpsM, have been purified and characterized. They were shown to form a stable, multiprotein complex spanning the cytoplasmic membrane. PMID- 11111910 TI - Cholesterol-binding cytolytic protein toxins. AB - Cholesterol-binding cytolysins (CBCs) are a large family of 50- to 60-kDa single chain proteins produced by 23 taxonomically different species of Gram-positive bacteria from the genera Streptococcus, Bacillus, Clostridium, Listeria and Arcanobacterium. Apart pneumolysin, which is an intracytoplasmic toxin, all the other toxins are secreted in the extracellular medium. Among the species producing CBCs, only L. monocytogenes and L. ivanovii are intracellular pathogens which grow and release their toxins in the phagocytic cells of the host. CBCs are lethal to animals and highly lytic toward eukaryotic cells, including erythrocytes. Their lytic and lethal properties are suppressed by sulfhydryl group-blocking agents and reversibly restored by thiols or other reducing agents. These properties are irreversibly abrogated by very low concentrations of cholesterol and other 3beta-hydroxysterols. Membrane cholesterol is thought to be the toxin-binding site at the surface of eukaryotic cells. Toxins molecules bind as monomers to the membrane surface with subsequent oligomerization into arc-and ring-shaped structures surrounding large pores generated by this process. Thirteen structural genes of the toxins (all chromosomal) have been cloned and sequenced to date. The deduced primary structure of the proteins shows obvious sequence homology particularly in the C-terminal part and a characteristic common consensus sequence containing a unique Cys residue (ECTGLAWEWWR) near the C terminus of the molecules (except pyolysin and intermedilysin). However, another Cys residue outside this undecapeptide and closer to the C-terminus occurs in ivanolysin. Genetic replacement of the Cys residue in the consensus undecapeptide by certain amino acids demonstrated that this residue was not essential for toxin function. Other residues in the undecapeptide have been mutagenized, particularly the Trp residues. One of these Trp appeared critical for lytic activity. The recent elucidation of the 3-D structure of perfringolysin O provided interesting information on the structure-activity relationship. The molecule was divided into four domains. Three domains are arranged in a row, giving an elongated shape. Domain 3 is covalently connected to the N-terminal domain 1 and packed laterally against domain 2. Membrane interaction of the monomer appears to be mediated by domain 4, while, oligomerization involves several sites scattered throughout the sequence. The Trp-rich region around the conserved Cys residue within domain 4 is assumed to conformationally adapt to cholesterol, and domain 3 is envisaged to move across the "hinge" by which it is connected to domain 1. PMID- 11111911 TI - Opening of the active site of Clostridium perfringens alpha-toxin may be triggered by membrane binding. AB - On the basis of amino acid sequence homologies with other phospholipases C, the alpha-toxin of Clostridium perfringens was predicted to be a two-domain protein. Using truncated forms of alpha-toxin the phospholipase C active site was shown to be located in the amino-terminal domain. Crystallographic studies have confirmed this organisation and have also revealed that the carboxy-terminal domain is structurally similar to the phospholipid-binding domains in eukaryotic proteins. This information has been used to devise a model predicting how alpha-toxin interacts with membranes via calcium-mediated recognition of phospholipid head groups and the interaction of hydrophobic amino acids with the phospholipid tail group. The binding of alpha-toxin to membranes appears to result in the opening of the active site allowing hydrolysis of membrane phospholipids. PMID- 11111912 TI - Surface dynamics of aerolysin on the plasma membrane of living cells. AB - Aerolysin secreted by the human pathogen Aeromonas hydrophila belongs to a group of bacterial toxins that are hemolytic and form channels in biological membranes. The toxin is secreted as an inactive precursor proaerolysin that must be proteolytically processed at its C-terminus to become active. The toxin then polymerizes into a heptameric ring that is amphipathic and can insert into a lipid bilayer and form a pore. We have examined these various steps at the surface of target cells. The toxin binds to specific receptors. Various receptors have been identified, all of which are anchored to the plasma membrane via a glycosylphosphatidyl inositol (GPI)-anchored moiety. The GPI anchor confers to the protein that is linked to it two usual properties: (i) the protein has a higher lateral mobility in a phospholipid bilayer than its transmembrane counterpart, (ii) the protein has the capacity to transiently associate with cholesterol-glycosphingolipid-rich microdomains. We have shown that both these properties of GPI-anchored proteins are exploited by proaerolysin bound to its receptor. The high lateral mobility within the phosphoglyceride region of the plasma membrane favors the encounter of the protoxin with its converting enzyme furin. The ability to associate with microdomains on the other hand favors the oligomerization process presumably by concentrating the toxin locally. PMID- 11111913 TI - SmcL, a novel membrane-damaging virulence factor in Listeria. AB - We describe here the fourth listerial membrane-damaging virulence factor, a sphingomyelinase C (SMase) that is produced specifically by the ruminant pathogen Listeria ivanovii. Its coding gene, smcL, is a monocistron expressed independently of PrfA. The smcL product, SmcL, is highly similar to the staphylococcal beta-toxin and is responsible for the differential hemolytic properties of L. ivanovii (bizonal hemolysis and CAMP-like reaction with R. equi). The role of SmcL in virulence was assessed by gene disruption and complementation. Our data show that SmcL mediates disruption of the membrane of primary phagosomes, thereby promoting bacterial intracellular proliferation. They also suggest that SmcL may play a role in host tropism. smcL is located in LIPI 2, a novel 18-kb pathogenicity island which also contains a cluster of internalin genes. LIPI-2 is unstable, L. ivanovii-specific and required for full virulence in mice and lambs. PMID- 11111914 TI - A structural overview of the Helicobacter cytotoxin. AB - VacA, the major exotoxin produced by Helicobacter pylori, is composed of identical 87-kDa monomers that assemble into flower-shaped oligomers. The monomers can be proteolytically cleaved into two moieties of 37 and 58 kDa, or P37 and P58. The most studied property of VacA is the alteration of intracellular vesicular trafficking in eukaryotic cells leading to the formation of large vacuoles containing markers of late endosomes and lysosomes. However, VacA also causes a reduction in trans-epithelial electrical resistance in polarized monolayers and forms ion channels in lipid bilayers. The ability to induce vacuoles is localized mostly but not entirely in P37, while P58 is involved in cell targeting. Here, we review the structural aspects of VacA biology. PMID- 11111915 TI - Pseudomonas aeruginosa exoenzyme S, a bifunctional type-III secreted cytotoxin. AB - Our recent studies have shown ExoS to be a bifunctional type-III secreted cytotoxin. Intracellular expression of the amino terminus of ExoS (C234) in eukaryotic cells stimulates actin reorganization without cytotoxicity, which involves small-molecular-weight GTPases of the Rho subfamily. Expression of the carboxyl terminus of ExoS comprises an ADP-ribosyltransferase domain, which is cytotoxic when expressed in cultured cells (Pederson and Barbieri, 1998). Rho and Ras are molecular switches, which control numerous cellular processes. Recent signaling studies suggest that there is crosstalk between Rho and Ras (Keely et al, 1997). Ras and Rho also contribute to wound healing processes and tissue regeneration. Recent studies have shown that microinjection of endothelial cells with activated Ras stimulated their motility, while microinjection of Ras blocking antibodies inhibited cellular motility that is a component of the wound healing process (Fox et al., 1994). In addition, hepatocyte growth factor/scatter factor (HGF/ SF) and epidermal growth factor stimulate cellular motility through the Ras signal transduction pathway (Ridley et al., 1995). Rac and Rho are also involved in motility and tissue regeneration, since dominant negative Rac inhibits the cellular motility stimulated by HGF/SF (Santos et al., 1997) and inhibition of Rho by either C. difficile ToxA and ToxB or the C. botulinum C3 transferase inhibits wound healing (Santos et al., 1997). Inhibition of tissue regeneration and wound healing appear to play a role in the pathogenesis of C. difficile, since treatment of gastrointestinal mucosa with C. difficile ToxA and ToxB alone inhibits regeneration of the gastric mucosa. Thus, ExoS may contribute to the establishment of P. aeruginosa infections by inhibiting wound healing and tissue regeneration by two mechanisms. The amino terminus of ExoS could inhibit Rho function and wound healing in a manner similar to C. difficile. Alternatively, ExoS could inhibit the cellular motility and angiogenesis required for wound healing by ADP-ribosylating Ras. Through the inhibition of tissue regeneration and wound healing, ExoS may play a pivotal role in chronic disease by maintaining sites of colonization. Inhibition of Ras or Rho signaling may also interfere with both innate and acquired immunity. Small-molecular-weight GTP binding proteins of the Ras superfamily are required for cellular processes, such as phagocytosis, as Rho proteins contribute to phagocytosis (Caron and Hall, 1998). Since Ras functions upstream of Rho in cellular signaling processes (Ridley et al., 1995), ADP-ribosylation of Ras by ExoS or the inhibition of Rho function by C234 may inhibit phagocytosis of P. aeruginosa by macrophages. Other studies indicate that Ras plays a role in T cell activation (Cantrell, 1994). Thus, ExoS may inhibit acquired immunity by inhibiting T-cell activation. PMID- 11111916 TI - Structural basis of pore formation by cholesterol-binding toxins. AB - In this paper we describe reconstructions by electron cryo-microscopy of two oligomeric states of the pore-forming toxin pneumolysin. The results are interpreted by the fitting of atomic models of separated domains to the 3 dimensional electron density maps, revealing two steps in the mechanism of pore formation by the family of cholesterol-binding toxins. We briefly describe the observation of the toxin pore in model membranes and contrast the apparent mechanism of pneumolysin with that of other pore-forming toxins. PMID- 11111917 TI - Crystal structure of the F component of the Panton-Valentine leucocidin. AB - Leucocidins and gamma-hemolysins are bi-component staphylococcal toxins that form lytic transmembrane pores. Their cytotoxic activities involve the synergistic association of a class S and a class F component, produced as water-soluble monomers which assemble on the surface of specific cells. The structure of the F protein from Panton-Valentine leucocidin, solved at 2.0 A resolution, and sequence alignment suggest that it represents the fold of any secreted protein in this family of toxins. The comparison of this structure to that of the homoheptameric alpha-hemolysin provides some insights into the molecular events that may occur during pore formation. PMID- 11111918 TI - Floating cholera toxin into epithelial cells: functional association with caveolae-like detergent-insoluble membrane microdomains. AB - In polarized cells, signal transduction by cholera toxin (CT) requires apical endocytosis and retrograde transport into Golgi cisternae and likely endoplasmic reticulum (ER) (Lencer et al., J. Cell Biol. 131, 951-962 (1995)). We have recently found that the toxin's apical membrane receptor ganglioside GM1 acts specifically in this signal transduction pathway, likely by coupling CT with caveolae or caveolae-related membrane domains (lipid rafts) (Wolf et al., J. Cell Biol. 141, 917-927 (1998)). Work in progress shows that 1) cholesterol depletion uncouples the CT-GM1 receptor complex from signal transduction, a characteristic of lipid rafts; 2) the GM1 acyl chains rather than the carbohydrate head groups appear to account for the structural basis of ganglioside specificity in toxin trafficking; and 3) intestinal epithelial cells obtained from normal adult humans exhibit lipid rafts which differentiate between CT-GM1 and LTIIb-GD1a complexes and which contain caveolin 1. PMID- 11111919 TI - Intracellular trafficking and membrane translocation of pertussis toxin into host cells. AB - The translocation of the pertussis toxin (PTX) S1 subunit into the cytoplasm of host cells was analysed in CHO cells producing S1 fused to a signal peptide. This protein channelled into the endoplasmic reticulum (ER) by the signal peptide, was found to ADP-ribosylate its target G proteins, suggesting that membrane translocation can occur from the ER and does not require the B oligomer. Similar results were obtained with a C-terminally truncated S1 subunit, indicating that this hydrophobic tail is not involved in the translocation mechanism. We also analysed the activity of two PTX mutants in which the S3 and S2 subunits were substituted for each other. The mutant protein containing two S3 subunits (PTXAS2) presented a decreased binding to fetuin or haptoglobin but higher in vivo activity than the wild-type PTX, suggesting that replacement of S2 by S3 favours the targeting of PTX to the compartment where translocation occurs and/or the dissociation of S1 from the B oligomer, thereby leading to a better translocation of S1 into the cytoplasm. PMID- 11111920 TI - Ricin transport into cells: studies of endocytosis and intracellular transport. AB - The plant toxin ricin binds to both glycoproteins and glycolipids with terminal galactose, and the toxin will therefore be endocytosed by the different mechanisms operating in a given cell. After endocytosis the toxin is transported to the Golgi apparatus by a process that differs from the Rab9-dependent transport of mannose-6-phosphate receptors. Retrograde toxin transport from the Golgi apparatus to the endoplasmic reticulum (ER) seems to be a requirement for subsequent toxin translocation to the cytosol where the toxin inhibits protein synthesis enzymatically. By using ricin we have characterized different types of endocytosis and the transport steps used by this toxin. PMID- 11111922 TI - The p21 GTP-binding proteins and bacterial toxins. PMID- 11111921 TI - Lethal factor of Bacillus anthracis cleaves the N-terminus of MAPKKs: analysis of the intracellular consequences in macrophages. AB - The lethal toxin of Bacillus anthracis consists of two proteins, PA and LF, which together induce lethal effects in some animal species and cause macrophage lysis. LF is a zinc-binding protein with metalloprotease activity. With a two-hybrid system approach we identified MAP kinase kinases (MAPKKs) Mekl and Mek2 as proteins interacting with LF. LF was shown to cleave Mek1 and Mek2 and an additional MAPKK family member MKK3, within their N-terminal region. We examined macrophage cell lines and primary peritoneal cells with different sensitivities to LF but did not find a direct correlation between MAPKKs cleavage and cell death. On the other hand, sublytic doses of LF cleave MAPKKs and cause a reduction in the LPS/IFNgamma-induced production of proinflammatory mediators. These findings are discussed with respect to the possible role of LF in the initial phase of infection. PMID- 11111923 TI - The diphtheria toxin channel-forming T-domain translocates its own NH2-terminal region and the catalytic domain across planar phospholipid bilayers. AB - The T-domain of diphtheria toxin, which extends from residue 202 to 378, causes the translocation of the catalytic A fragment (residues 1-201) across endosomal membranes and also forms ion-conducting channels in planar phospholipid bilayers. The carboxy-terminal 57-amino acid segment (residues 322-378) in the T-domain is all that is required to form these channels, but its ability to do so is greatly augmented by the portion of the T-domain upstream from this. Here we show that in association with channel formation by the T-domain, its hydrophilic 63-amino acid NH2-terminal region (residues 202-264) as well as the entire catalytic A fragment (residues 1-201) cross the lipid bilayer. The phenomenon that enabled us to demonstrate this was the rapid closure of channels at cis negative voltages when a histidine tag was placed at various positions in the NH2-terminal region of the T-domain or in the A fragment; the inhibition of this effect by trans nickel established that the histidine tag was present on the trans side of the membrane. Thus, all of the machinery necessary to translocate the A fragment across membranes is built into the 114 residues at the carboxy-terminal end of the T domain (residues 265-378), without the requirement of any proteins in the plasma membrane (e.g., toxin receptor) or of any other cellular components. PMID- 11111924 TI - Bordetella pertussis adenylate cyclase toxin as a tool to analyze molecular interactions in a bacterial two-hybrid system. AB - Bordetella pertussis secretes a calmodulin-activated adenylate cyclase toxin (CyaA) that is able to enter into eukaryotic cells. We took advantage of the modular structure of the catalytic domain of CyaA to design a genetic system that can detect protein-protein interactions in Escherichia coli. This bacterial two hybrid system is based on the functional complementation between two complementary fragments, T25 and T18, of the catalytic domain of CyaA, in an E. coli cya strain. This bacterial two-hybrid system could find applications in the studies of structure/function relationships of proteins, in functional analysis of genomes, in high-throughput screening of interacting ligands and in design of new therapeutic agents. PMID- 11111925 TI - Immune modulation by the cholera-like enterotoxin B-subunits: from adjuvant to immunotherapeutic. AB - Cholera toxin (Ctx) and its close relative, Escherichia coli heat-labile enterotoxin (Etx) have long been established as potent mucosal and systemic adjuvants. Problems arising from their inherent toxicity have, however, precluded human use. Here we describe findings which demonstrate that contrary to the established dogma the non-toxic B-subunit of Etx (EtxB) is a highly potent mucosal adjuvant capable of potentiating protective immunity to viral infection. The mechanisms which underlie this activity arise from an ability to trigger specific signaling processes in lymphocyte populations which modulate differentially their activation, differentiation and survival. The elucidation of these properties has led to the further use of EtxB as an agent capable of preventing the establishment of autoimmune diseases. The basis for these activities and their potential applicability to human therapies are discussed. PMID- 11111926 TI - LTK63 and LTR72, two mucosal adjuvants ready for clinical trials. PMID- 11111927 TI - Bacillus thuringiensis and its use in transgenic insect control technologies. AB - The insecticidal activity of Bacillus thuringiensis (Bt) is mainly due to the production of crystals containing insecticidal crystal proteins (ICPs). These proteins are very selectively active against certain insect species, including some agronomically important pest species. Some ICP genes have been used for bioengineered crop protection, resulting in transgenic crop plants with excellent insect protection. PMID- 11111928 TI - Design of toxins that can be activated by cell-specific proteases and their potential use in targeted cell killing. AB - Protein toxins designed to eliminate specific cell types, e.g. disease-associated cells, have mainly made by linking the active domain of the toxin to a protein that only binds to certain cells. A different approach for the construction of toxins capable of killing disease-associated cells is suggested here, based on the knowledge that many of these cells express specific proteases that are not expressed in normal tissue. The construction of toxins that become activated through cleavage by the protease (HIV-1 PR) expressed by the HIV-1 virus is described. These toxins contain a signal for degradation by the N-end rule pathway, which is cleaved off by HIV-1 PR, resulting in increased toxicity. Alternative strategies for the construction of toxins that can be activated by proteases are discussed. PMID- 11111929 TI - The superantigenic toxin of Yersinia pseudotuberculosis: a novel virulence factor? AB - Recently, a superantigenic toxin designated YPM (Yersinia pseudotuberculosis derived mitogen) was characterized in the supernatant of Y. pseudotuberculosis, a Gram-negative bacterium involved in human enteric infection. To assess the role of YPM in pathophysiology of Y. pseudotuberculosis, a superantigen-deficient mutant was constructed and its virulence was tested in a murine model of infection and compared with the virulence of the wild-type strain (wt). Determination of the survival rate after intravenous inoculation of mice clearly demonstrated a higher survival rate when animals were infected with the superantigen-deficient strain. This decreased virulence of the mutant strain could not be explained by a lower bacterial growth rate in spleen, liver or lung of infected animals. Therefore, production of IFNgamma, TNFalpha, IL-2, IL-6 and IL-10 was followed during the course of infection by cytokine assay in the blood and mRNA detection in the spleen. IL-6 and IFNgamma were the two major cytokines detected whereas TNFalpha production was never observed. PMID- 11111930 TI - Murine toxin of Yersinia pestis shows phospholipase D activity but is not required for virulence in mice. AB - Purified murine toxin (Ymt) of Yersinia pestis is highly toxic for mice and rats but less active in other animals such as guinea pigs, rabbits, dogs and monkeys. This suggested that Ymt contributes to the very low infectious dose of Y. pestis in mice. The gene encoding Ymt (ymt) is localised on the 100-kb plasmid pFra, which is unique for Y. pestis. Sequence analysis revealed that Ymt showed homology to proteins of the phospholipase D (PLD) superfamily of proteins. Y. pestis strains expressing Ymt possessed PLD activity whereas strains carrying deletions in the ymt gene showed no detectable PLD activity. Western blot analysis showed that Ymt was associated with bacteria under normal growth conditions, and immunogold EM revealed that Ymt was mainly localised in the bacterial cytoplasm. Ymt was purified to homogeneity, and the purified toxin showed a dose-dependent PLD activity. Substitution of amino acids in the PLD consensus motif of Ymt essentially abolished the enzymatic activity and these variants of the toxin were no longer toxic to mice. Interestingly, an in-frame deletion mutant of ymt in the Y pestis strain KIM was not significantly attenuated for mouse virulence. Together with the observation that expression of Ymt was higher at room temperature compared to 37 degrees C this prompted us to investigate the role of Ymt in the flea vector. Fleas were infected with isogenic ymt+ or ymt- mutant strains of Y. pestis. Preliminary results suggest that Ymt is important for survival of Y. pestis in the flea and thereby also for the flea borne route of infection. PMID- 11111931 TI - The mycoplasma superantigen MAM: role in arthritis and immune-mediated disease. PMID- 11111932 TI - Enterotoxins and the enteric nervous system--a fatal attraction. AB - Although there has been extensive investigation of the biochemical consequences of the interactions between bacterial enterotoxins and intestinal epithelial cells and the mechanisms by which they induce intestinal secretion, relatively little attention has been given to other aspects of the host response to these enterotoxins. There is now compelling evidence that the enteric nervous system has a major role in enhancing the secretory state induced by cholera toxin, the E. coli enterotoxins and possibly C. difficile toxin A. Cholera toxin for example is thought to activate a neural reflex via the release of 5-hydroxytryptamine from enterochromaffin cells. Neurotransmitters involved in the reflex include substance P and vasoactive intestinal polypeptide. Delineation of these neural pathways may offer new possibilities for the pharmacological control of enterotoxin-mediated secretion. PMID- 11111933 TI - The pathogenesis of clostridial myonecrosis. AB - These pieces of evidence can be assimilated into a molecular and cellular model of pathogenesis which is initiated by direct toxin effects upon venous capillary endothelial cell function, leading to expression of pro-inflammatory mediators and adhesion molecules, and initiation of platelet aggregation. Toxin-induced hyperadhesion of leukocytes (see above section) with enhanced respiratory burst activity (due to toxins directly or to toxin-induced IL-8 or PAF synthesis by host cells) and toxin-induced chemotaxis deficits could result in neutrophil mediated vascular injury. Direct toxin-induced cytopathic effects on EC may also contribute to vascular abnormalities associated with gas gangrene. Over prolonged incubation periods, PLC at sublytic concentrations causes EC to undergo profound shape changes similar to those described following prolonged TNF or interferon gamma exposure. In vivo, conversion of EC to this fibroblastoid morphology could contribute to the localized vascular leakage and massive swelling observed clinically with this infection. Similarly, the direct cytotoxicity of PFO could disrupt endothelial integrity and contribute to progressive edema both locally and systemically. Thus, via the mechanisms outlined above, both PLC and PFO may cause local, regional and systemic vascular dysfunction. For instance, local absorption of exotoxins within the capillary beds could affect the physiological function of the endothelium lining the postcapillary venules, resulting in impairment of phagocyte delivery at the site of infection. Toxin-induced endothelial dysfunction and microvascular injury could also cause loss of albumin, electrolytes, and water into the interstitial space resulting in marked localized edema. These events, combined with intravascular platelet aggregation and leukostasis, would increase venous pressures and favor further loss of fluid and protein in the distal capillary bed. Ultimately, a reduced arteriolar flow would impair oxygen delivery thereby attenuating phagocyte oxidative killing and facilitating anaerobic glycolysis of muscle tissue. The resultant drop in tissue pH, together with reduced oxygen tension, might further decrease the redox potential of viable tissues to a point suitable for growth of this anaerobic bacillus. As infection progresses and additional toxin is absorbed, larger venous channels would become affected, causing regional vascular compromise, increased compartment pressures and rapid anoxic necrosis of large muscle groups. When toxins reach arterial circulation, systemic shock and multiorgan failure rapidly ensue, and death is common. PMID- 11111934 TI - Bacterial protein toxins--general conclusions. PMID- 11111935 TI - Experimental intoxication by the mushroom Ramaria flavo-brunnescens in sheep. AB - Ramaria flavo-brunnescens collected in autumn from 1990 to 1994 was orally administered to 11 sheep. These animals were dosed with 100-430 g/kg bw administered over 3-13 d. Six sheep showed clinical signs and 4 of them died. The mininum toxic dose was of 150 g/kg bw. Clinical signs were anorexia, hyperthermia, dyspnea, polyuria, ataxy, muscle tremors and seizures. The eyes had hyperemia of the sclera and, in some cases, hemorrhages of the anterior chamber or corneal opacity. Sheep dosed with higher doses had ulcerations of the tongue and necrotic lesions in the hooves. The main histologic lesions of the feet and tongue were miopachynsis and endotelial degeneration followed by degeneration, necrosis and ulceration of the epithelium. Hemorrhages of the anterior chamber, and severe congestion and hemorrhages of the iris, ciliary body and process were observed in the eyes. Congestion and perivascular hemorrhages occurred in the central nervous system. The similarity of clinical signs and pathologic lesions induced by R flavo-brunnescens and those caused by ergotism in cattle and sheep suggests the presence ofa vasoactive constrictive substance in the mushroom. Fresh R flavo-brunnescens dosed in autumn 1993 was not toxic at doses of 200-400 g/kg demonstrating variations in the toxicity of the mushroom from year to year. PMID- 11111936 TI - Are 2,3-dimercapto-1-propanesulfonic acid or prussian blue beneficial in acute thallotoxicosis in rats? AB - Unithiol (2,3-dimercapto-I-propanesulfonic acid, DMPS) and prussian blue (potassium ferric hexacyanoferrate (II), PB), given alone or in combination, were evaluated as antidotes to treat acute thallotoxicosis in male Sprauge-Dawley rats. Animals were poisoned with 20 mg thallium (TI)/ kg bw PO on day 0 using thallous sulfate. On day 1 (24 h later), treatments began and were continued through day 4 as follows: 50 mg PB/kg bw PO, 2/ d; 5 mg DMPS/kg bw IP, 6/d (day 1), 4/d (day 2), 2/d (days 3-4); or the combination. Animals were sacrificed 24 h after the last treatment (day 5), and TI concentrations in kidney, liver, heart, brain, whole blood and feces determined by electrothermal atomic absorption spectroscopy. The relative accumulation of TI was kidney>>heart>liver approximately equal brain. PB limited incorporation of TI in all tissues. DMPS failed to significantly decrease TI in any organ, but significantly decreased TI in whole blood. PB+DMPS treatment significantly decreased the TI content in all organs, but not to a greater extent than PB alone. PB and PB+DMPS treatments significantly increased TI in feces, whereas DMPS alone produced little effect. This study confirms that PB is beneficial in the treatment of acute thallotoxicosis in rats. The failure of DMPS to significantly affect TI in target organs suggests it is not useful in treating TI poisoning. PMID- 11111937 TI - Acute and repeated vapor exposure toxicology of 3-(methylthio)propionaldehyde. AB - Because of its vapor pressure (0.6 torr at 20 C) there is a potential for vapor exposure to 3-(methylthio)propionaldehyde (3-MTP) vapor. Liquid 3-MTP may contain trace amounts of acrolein (up to 0.1%), and therefore acrolein vapor may also be present. Acute exposure (24 min to 4 h) of rats to substantially saturated atmospheres of 3-MTP generated statically (measured concentrations of 261-951 ppm) resulted in marked ocular and respiratory irritancy followed by death. Deaths occurred either during exposure or a few days postexposure, depending on exposure time. Measured acrolein vapor concentrations in these static studies were 16.7-216 ppm. In contrast, when substantially saturated vapor atmospheres were generated dynamically (277-320 ppm 3-MTP) only minor transient signs of irritancy were present, and only 1/40 exposed animals died. Acrolein vapor concentrations ranged 0-6.8 ppm. These findings indicate that the toxicity associated with acute static exposures to 3-MTP vapor was due to accumulated acrolein vapor, and that 3-MTP per se has a low order of acute vapor inhalation toxicity. In a first 9-d repeated vapor exposure study (6 h/d) rats were exposed to 0, 23.6, 96.8 or 246.2 ppm 3-MTP vapor; the mean acrolein concentration was 1.34 ppm (range 1.08-1.72 ppm). There were no mortalities, but exposure concentration-related indications of toxicity were present. These included reduced body weights, hematology (increased lymphocytes), serum chemistry (reduced total protein and globulin), and respiratory tract histopathology. The latter consisted mainly of squamous metaplasia in the anterior nasal passages at all concentrations, being minimal at 23.6 ppm. At the high concentration there was also olfactory atrophy and squamous metaplasia in the larynx, trachea, and larger bronchi; 23.6 ppm was a threshold effect level. The respiratory tract histopathology was compatible with exposure to acrolein vapor. In a second 9-d study, rats were exposed to 0, 0.47, 4.99 or 50.5 ppm (6 h/d); no acrolein could be detected in the chamber air samples. There were no differences between the controls (air alone) and 3-MTP exposed animals with respect to signs, body weights, food consumption, hematology, serum chemistry, urinalysis, and gross and microscopic pathology. Without detectable acrolein vapor, 50.5 ppm 3-MTP was a no observable effects level. PMID- 11111938 TI - Effects of fescue and clover forage on serum lactate dehydrogenase and glucose 6 phosphate dehydrogenase isoenzymic profiles in steers. AB - We determined the effects of forage type on isoenzymes of lactate dehydrogenase (LDH) and glucose 6-phosphate dehydrogenase (G6PDH). Forty-eight crossbred steers were randomly allotted to replicated pastures consisting of fungus-infected (Neotyphodium coenophialum) fescue or fungus-free fescue each with or without ladino clover overseeding. At the end of the 180-d grazing period, serum was harvested from the steers. Steers were finished in a feedlot and slaughtered after approximately 150 d in the feedlot. Isoenzymes for LDH and G6PDH were separated using PAGE. Five LDH isoenzymes (L1-15) were typically detected. Isoenzyme L1 (most anodic) had the greatest area percent as detected by laser densitometry (72, 12, 10, 5, and 7%, respectively, for L1, L2, L3, L4, and L5). Four proteins had G6PDH activity (G1-G4) with G2 having the greatest area percent (15, 52, 27, and 14, respectively, for G1, G2, G3, and G4). Isoenzymes within a dehydrogenase were correlated (P < .05). In addition, area percentage of L1 was correlated (P < .05; r = .34) with area percentage of G2, and area percentage of L4 was correlated (P < .07; r = .73) with area percentage of G1. Area percentages of L1, L2, and L3 were affected by an interaction (P < .09) of forage types. Body weight gains for steers grazing endophyte-infected fescue were depressed (P < .05); however, steers compensated with increased (P < .05) weight gains during the finishing phase. Fungal toxins produced by Neotyphodium coenophialum may alter an animal's metabolism, growth, and development via shifts in reducing equivalents (NADH). PMID- 11111939 TI - Evaluation of the antibacterial and hemolytic activities of Latvian herbal preparation. AB - Three extracts originating from a combination of various Latvian plant species were tested for their antibacterial activities by evaluating growth delays using a fully automated microturbidimetric method. Ten different human and bovine strains of the genera Staphylococcus and Micrococcus were used as test microorganisms. The inhibitory effect in vitro was defined as the difference between the growth rate without herbs and the growth rate in the presence of an extract. Among the tested strains, Staphylococcus aureus was found sensitive to all 3 extracts. However, extract I was the most effective in slowing the growth of all strains tested. Using appropriate tester strains it should be possible to set up a broad-range microtubidimetry assay for individual herb screening in vitro. The hemolytic effects of the individual extracts on human erythrocytes were also studied at different concentrations. Two of the herbal extracts had minimal lytic effects on eurocaryotic cells. An additional hemolysis test was conducted in the presence of coenzyme Q10 (CoQ10) as a free radical scavenger: CoQ10 had no effect on the hemolytic reaction. PMID- 11111940 TI - Determination of dexamethasone in milk of dairy cows by immuno-enzymatic assay. AB - Eight lactating cows received 3 im injections at 24 h intervals of a commercial formulation containing dexamethasone. Each treatment provided 25 microg/kg bw/d of dexamethasone acetate, equivalent to 22.6 mg of dexamethasone. Milk samples were obtained before treatment (5 d), during the treatment period, and for up to 22 milking after the last injection. The concentrations of dexamethasone in the milk samples were determined by a commercial competitive immunoenzymatic assay for corticosteroids (detection limit 0.15 ng dexamethasone/ml). The conventional therapeutic dose of dexamethasone acetate caused milk drug concentrations exceeding the tolerated maximum residue limit (0.3 mg/kg). A withdrawal time of 3 3.5 d for dexamethasone in milk provided sufficient protection for consumer health. The commercial enzyme immunoassay kit employed in this study was sufficiently sensitive, easy to use, and appropriate to monitor the use of dexamethasone in lactating animals. PMID- 11111941 TI - Intoxication of sheep exposed to ozark milkweed (Asclepias viridis Walter). AB - Some 20 sheep died 1 at a time on a farm in Fleming County, KY, in late July of 1999 after consumption of Asclepias viridis Walter. Major histological lesions were mild multifocal nonsuppurative myocarditis. Gross pathology revealed wet and heavy lungs. Many affected animals had a hunched appearance, and marked posterior paresis was also observed. PMID- 11111942 TI - Diarrhea-associated over-anticoagulation in a patient taking warfarin: therapeutic role of cholestyramine. AB - We present a case of significant over-anticoagulation temporally associated with a bout of protracted diarrhea in a patient on warfarin therapy. Cholestyramine was utilized to interrupt the enterohepatic recycling of warfarin and for its antidiarrheal effects to prevent gastrointestinal vitamin K wasting. Cholestyramine enabled the use of very low doses of sc vitamin K1 (2 mg total) with subsequent attainment of a therapeutic International Normalized Ratio in 39 h. PMID- 11111943 TI - Experimental kerosene poisoning in goats. AB - Eighteen young native male and female goats were divided into 3 equal groups. Kerosene was given to Groups 1, 2 and 3 as single doses of 10, 20 or 40 ml/kg bw respectively. Clinical signs In-Group 1 were mild behavioral changes and in Group 2 were mild to moderate bloat, coughing and behavioral changes. None of the goats of Groups 1 and 2 died. Goats of Group 3 had severe signs of poisoning and died within 4 h to 11 d after dosing with clinical signs of severe bloat, frequent coughing, vomiting, and expelling of kerosene from the mouth and nose. Star gazing, depression, recumbency and dyspnea also occurred. Postmortem changes in Group 3 were gangrenous pneumonia, pleuropneumonia, congestion in brain and kidney, perivascular and perineuronal edema in brain tissue, and renal nephrosis. PMID- 11111944 TI - Rapid method for the detection of cyanide gas release from plant material using CYANTESMO paper. AB - A method for detection of cyanide gas release from solutions and from suspensions of plant material is described which offers advantages in speed, safety and simplicity over the picric acid method. PMID- 11111945 TI - Rapid determination of ethylene glycol and glycolic acid in biological fluids. AB - Ethylene glycol poisoning of companion animals is a common occurrence and is sometimes involved in human intoxication. Ethylene glycol is of limited toxicity, but the metabolites including glycolic acid are responsible for poisoning. Conventional treatment has employed substances to prevent alcohol dehydrogenase from metabolizing the ethylene glycol, but to be effective, therapy must begin within hours of ethylene glycol consumption. We describe a rapid (10 min) analysis of biological fluids for ethylene glycol and glycolic acid using isocratic HPLC, a refractive index detector, and a Waters fast fruit juice analytical column. PMID- 11111946 TI - Biological monitoring of embrio-fetal exposure to methamidophos or chlorothalonil on rat development. AB - Maternal exposure to pesticides during the pre-implantation and very early post implantation periods of pregnancy is correlated with numerous adverse effects on the offspring and in reproductive parameters like an increase in resorption, a decrease in fetal survival and weight, and teratogenic effects. Although the epidemiological evidence is inconclusive as regards the risk of the adverse outcome of pregnancy and developmental toxicity events, the use of biomarkers in exposure assessment may contribute to recognizing a potential health impairment. The present study evaluated the influence of prenatal oral exposure to an insecticide (1.0 mg methamidophos/kg) or a fungicide (200.0 mg chlorothalonil/kg) during gestation days 1 to 6 on maturational and behavioral aspects of offspring development of rats. The pesticides did not affect the body weight gain of dams and offspring, nor did the exposure affect the weight of gravid uterus, fetus, placenta and ovary. There were no observed alterations in the swimming behavior tested at postnatal days 7, 14 and 21, but the pesticides interfered with physical and maturational development landmarks of offspring according to age, showing subtle effects on behavioral and physical development. These findings show the importance of categorizing developmental effects, establishing the relationship between age and important performances, to recognize potential impacts on human populations. PMID- 11111947 TI - Characteristics of adult acute poisoning mortality in a large industrial-agrarian region of Bulgaria during socioeconomic transition and crisis (1990-1998). AB - The severe socioeconomic crisis in Bulgaria, accompanying the transition after the collapse of the totalitarian regime affected unfavorably the health status of the population, increasing the incidence of socially important diseases by making poverty, unemployment, emigration, alcoholism and drug addiction widespread. Systematic studies of acute poisoning suicide and poisoning mortality still lack. To analyze acute poisoning (AP) characteristics during this period in a large industrial-agricultural region of Bulgaria, we retrospectively examined 327 adult AP deaths in Plovdiv region 1990-1998. Males (73.1%) prevail in all age categories. As a caseload men predominate in "working age" categories, women in "pre/retirement age". Mortality rates tended to increase with age, more marked in men. The main cause of death from AP was alcohol intoxication in men (30.1%) and drugs in women (33.7%). Suicides prevailed over accidents (54.1% vs. 45.9%). Accidents were significantly higher in men, suicides in women; 57.5% received no medical aid for more men than women (61.9% vs. 45.5%). Untapped resource for decreasing AP mortality rates are patients who do not receive medical aid and die at home, although that is difficult to correct in a period of crisis. PMID- 11111949 TI - A reminiscence about "lead non-poisoning". PMID- 11111948 TI - Accidental ingestion of acyclovir in dogs: 105 reports. AB - Acyclovir is an antiviral agent that causes termination of viral DNA synthesis by inhibiting viral reverse transcriptase. Acyclovir is used therapeutically to treat herpes simplex, cytomegalovirus, Epstein-Barr, and varicella-Zoster. Although acyclovir is thought to be low in toxicity, it has caused an obstructive nephropathy from accumulation of crystals in renal tissue. A retrospective review (January 1995 through March 2000) was conducted of acyclovir toxicoses in dogs reported to the ASPCA National Animal Poison Control Center. Of 105 ingestions, 10 were considered cases of acyclovir toxicosis. The most common signs seen were vomiting, diarrhea, anorexia, and lethargy. Ingested dosages ranged from 40 to 2195 mg/kg bw. Polyuria and polydipsia were reported in I dog. In 6/10 cases, signs developed within 3 h of ingestion. Treatment included standard decontamination procedures, (ie induction of emesis, administration of activated charcoal), diuresis, and supportive care. PMID- 11111950 TI - The "roots of toxicology" continued... PMID- 11111951 TI - Intracellular mediators of programmed cell death initiated at the cell surface receptor Fas. AB - Apoptosis is a programmed cell death process, which plays a pivotal role in development, in tissue homeostasis and in several human diseases. Fas (CD95/Apo 1) is a member of the "death receptors" family, a group of cell surface proteins that trigger apoptosis upon binding with their natural ligands. In the immune system, intracellular signal transduction triggered from Fas splits into two different pathways. The proteolytic pathway is mediated by a family of cysteine proteases, the caspases, responsible for the morphological changes occurring in the apoptotic process. To complete this death program, another series of events, involving a lipid pathway, is necessary. Upon Fas stimulation, a sequential activation of specific enzymes results in the accumulation of ceramides and GD3 ganglioside. GD3 directly induces mitochondrial damage and triggers the release of apoptogenic factors, allowing efficient execution of Fas-mediated apoptosis. PMID- 11111952 TI - Immunobiology of xenotransplantation. AB - The transplantation of organs between disparate species is hindered by severe immune responses of the recipient against the graft. These immune responses gives rise to hyperacute and acute vascular rejection and to cellular rejection. Research during the past decade has shed light on the elements of the immune system responsible for the rejection of xenografts and has provided novel and incisive therapies which might be applied to these problems. PMID- 11111953 TI - Statistical issues in clinical trials. AB - The care for patients having organ transplants has improved greatly. This improvement is due, in part, to the advances in knowledge gained through clinical trials. These trials are most useful when they address questions which are important (to patients, their families and their clinical care-givers), which are at their most rigorous statistically (by reducing bias and increasing precision), and which relate closely to the real world. Statisticians and clinicians need to work together to achieve these aims. PMID- 11111954 TI - Atherosclerotic complications after renal transplantation. AB - Death with functioning graft, the most frequent cause being cardiac death, continues to be the most frequent cause of long-term graft loss. The risk of cardiovascular death in the transplanted patient is lower than in patients with other modalities of renal replacement therapy, but continues to be substantially higher than in the general population. Amongst the factors predicting patient and graft survival are hypertension, dyslipidemia, smoking and possibly hyperhomocysteinemia. It is concluded that lowering of blood pressure to levels far lower than levels accepted in the past, more widespread administration of statines, cessation of smoking and possibly administration of folate should reduce cardiovascular mortality and possibly also influence chronic allograft vasculopathy. PMID- 11111955 TI - Clinical immunosuppression 2000. PMID- 11111956 TI - Preoperative dobutamine stress echocardiography versus cardiac arteriography for risk assessment prior to renal transplantation. AB - Because coronary artery disease is the leading cause of death in patients with end-stage renal disease, we prospectively studied the prognostic value of dobutamine stress echocardiography (DSE) compared to coronary angiography (CA) as an evaluative tool. Thirty-three patients at high risk for coronary artery disease were selected from a cohort of 133 renal transplant candidates and underwent both DSE and CA. In this study, the value of DSE was found to exist in its strong negative predictive value (92%). A negative DSE coupled with a negative clinical cardiac evaluation was found to practicably exclude the necessity for CA. DSE can thus serve as a non-invasive, low cost screening test. PMID- 11111957 TI - Is hepatitis C virus infection a risk factor for panel-reactive antibody positivity? AB - Patients with high levels of panel-reactive antibody (PRA) represent an increasingly large group in the waiting lists for cadaveric renal transplantation. Hepatitis C virus (HCV) infection has been found to be associated with a high prevalence of positivity of autoimmune serological tests. We planned this study to evaluate the effect of HCV positivity on the PRA levels in our hemodialysis (HD) patients. We included 38 HCV-infected (group I: 20 male, 18 female patients, mean duration of HD 73.6 +/- 50.6 months) and 43 hepatitis marker-negative (group II: 23 male, 20 female patients, mean duration of HD 22.2 +/- 22.4 months) HD patients. The PRA positivity ratio and number of transfusions were not significantly higher in group I than in group II (PRA ABC; 28.9%, 19.4, P > 0.05, PRA DR; 21.8%, 20.9, P > 0.05, respectively, and blood transfusions 7.0 +/- 5.7, 6.6 +/- 5.2, respectively, P = 0.06). HD duration correlated significantly with PRA positivity in our patients (PRA-positive patients: 56.1 +/ 57.9 months, PRA-negative patients: 43.3 +/- 41.9 months, P = 0.021). In conclusion, HD duration was found to be the main factor affecting PRA sensitivity independently of HCV positivity and blood transfusion. PMID- 11111958 TI - Increasing urinary IL-6 levels announce kidney graft rejection. AB - Acute rejection (AR) is the recipient's inflammatory response to the grafted organ. Within the graft-infiltrating cells, a high ratio of IL-6 producing cells can be found, indicating local IL-6 production. Therefore, in cases of kidney transplantation, urinary (u) IL-6 should be detectable. In order to establish the dynamics and diagnostic relevance, uIL-6 levels were determined daily by Quantikine IL-6 immunoassay (R & D Systems, Minneapolis, Minn.) in 101 kidney graft recipients (n = 1915 urine samples) during their post-transplant hospital stay. Immunosuppression consisted of azathioprine, steroids, cyclosporine and an intraoperative high-dose single antithymocyte globulin (ATG)-Fresenius bolus (9 mg/kg). In all the uncomplicated courses (n = 31) mean uIL-6 level was determined, after a post-transplant peak of 174 pg/ml, to be between 4 and 8 pg/ml. In contrast, delayed graft function (n = 16) was always associated with very high uIL-6 levels (> 200 pg/ml), dropping down only with commencement of graft function. Steroid-sensitive AR (n = 14) was consistently associated with significantly increasing uIL-6 levels prior to antirejection therapy (from 23 to 82 pg/ml). In cases of steroid-resistant AR, following antirejection therapy with methylprednisolone (5 days 5 mg/kg), there was no obvious trend towards normalization, indicating the persistence of inflammation (mean uIL-6 peak prior to OKT3 or ATG therapy: 99 pg/ml). In addition, AR-associated uIL-6 levels were found to be of much greater diagnostic relevance than AR-associated serum IL-6 levels. In bacterial urinary tract infections (n = 20), increased uIL-6 levels (peak 53 pg/ml) coincided with the commencement of antibiotic therapy. In mild cytomegalovirus diseases (n = 8), the development of leukocytopenia was associated with a slight increase of uIL-6 (peak 26 pg/ml), showing graft involvement. All increased uIL-6 values returned towards baseline after successful treatment. Thus, uIL-6 provides information about the intragraft inflammatory situation. Its determination is simple, expressive, non-invasive and can be recommended. PMID- 11111959 TI - Does the risk of acute rejection really decrease with increasing recipient age? AB - In corneal transplants the risks of acute rejection and graft failure decrease with increasing recipient age, but kidney graft survival analyses tend to show the opposite effect. Why is this? Cadaveric kidney transplants performed in the UK and Republic of Ireland between 1985 and 1993 (UKTSSA database) were analysed by multifactorial methods to identify major factors affecting graft survival. In a study database that had been censured for technical failure and death with a functioning transplant, it was shown that increasing recipient age was associated with decreasing risk of graft failure at 1 year. This is consistent with the view that kidney transplants, like corneal transplants, are less likely to be acutely rejected as the age of the recipient increases. PMID- 11111960 TI - Careful clinical monitoring in comparison to sequential Doppler sonography for the detection of acute rejection in the early phase after renal transplantation. AB - Acute rejection is the most frequent cause of early graft failure. There is unanimity that Doppler sonography is a helpful method for the detection of complications after kidney transplantation. In the past, the indication for renal biopsy relied mainly on clinical assessment, although this assessment has not been standardised. Therefore, we conducted this prospective study to compare the value of sequential Doppler measurements with a standardised clinical rejection score, based on renal function, weight gain, graft swelling and tenderness. Fifty eight patients (37 males, 21 females, mean age 46 +/- 12 years) after kidney transplantation were consecutively enrolled into the study. Doppler investigations were obtained within the first 24 h after transplantation, followed by an interval of 48-72 h. At the same time, a clinical examination was scored by a transplant physician blinded to the Doppler results. Clinical score and Doppler results, both were referred to the histological results of renal biopsy. In 24 out of 58 patients 25 acute rejections occurred. In seven patients, acute rejection was superimposed on primary graft failure. The cut-off levels for rejection were set at RI > or = 0.80 and PI > or = 1.70 based on receiver operator curves using data from 663 Doppler examinations. Sensitivity and specificity was 72% for RI, and 72% and 74% for PI, respectively. The calculation of the intraindividual increase (deltaRI > or = 3%, deltaPI > or = 10%) did not improve these values. The clinical score revealed a sensitivity and specificity of 82% and 87%, respectively. The combined analysis of Doppler indices and clinical score showed a sensitivity of 96% with a specificity of 66%. Careful clinical monitoring alone using a clinical score is an appropriate procedure with which to decide about renal biopsy. Our data show that Doppler sonography should be performed within the first 24 h after transplantation to evaluate graft perfusion and baseline values. Afterwards, it should be used when clinical signs of rejection occur to underline the decision for renal biopsy even in borderline cases. PMID- 11111961 TI - Sub-clinical acute rejection detected using protocol biopsies in patients with delayed graft function. AB - Acute rejection in renal transplants is difficult to diagnose when patients have delayed graft function (DGF) in the early post-transplant period. In this study protocol, renal transplant biopsies were performed in an attempt to detect sub clinical acute rejection episodes. Eighty-three patients were eligible for the study, of whom 33 had DGE All had protocol renal transplant biopsies performed under ultrasound control at 7 days post-transplant, and those with DGF had further biopsies weekly until the graft functioned. All histologically confirmed acute rejection episodes were treated. Sub-clinical acute rejection was detected in 6/33 (18%) patients with DGF compared to 2/50 (4%) in the other patients (P < 0.05). Borderline rejection was present in 4/33 (12%) and 4/50 (8%) patients, respectively. Because of the high detection rate of sub-clinical acute rejection and the low morbidity of renal transplant biopsies, their use is recommended in patients with DGF. PMID- 11111962 TI - Renal transplantation combined with aortofemoral bypass using a fresh arterial allograft. AB - Aortoiliac atherosclerosis is frequently encountered in renal failure patients waiting for renal transplantation. Staged or simultaneous surgical repair of aortoiliac lesions with renal transplantation is possible at reasonable risk. Arterial reconstruction is most commonly performed using an artificial prosthesis. Another option is the use of a fresh or preserved arterial allograft. In our institute, about 180 cadaveric transplantations are performed each year. Over the past 2 years, three patients with chronic renal failure and obliterative disease of the abdominal aorta and iliac arteries underwent aortofemoral bypass using a fresh arterial allograft combined with kidney transplantation from the same donor. The procedures as well as the postoperative course were uneventful. There was an immediate development of function of the renal transplant. Combined arterial reconstruction and transplantation, managing both conditions at a time, is convenient for the patient mainly because it means undergoing only one general anesthesia during one hospitalization. Moreover, the risk of infection of the vascular prosthesis is somewhat reduced. Disadvantages are that the availability of the arterial allograft is dependent on a suitable donor and the limited body of experience with the behavior of the arterial allograft in patients with chronic immunosuppression. PMID- 11111963 TI - The inferior vena caval conduit--a neglected technique in transplantation of the right cadaveric kidney? AB - A short right renal vein may reduce access or compromise optimal positioning during transplantation of the right cadaveric kidney. This difficulty could be overcome by using the inferior vena cava (IVC) as a venous conduit to lengthen the short right renal vein. This manoeuvre would also facilitate training by ensuring safe tension-free vascular anastomoses since the kidney can be lifted up a comfortable distance, thus improving exposure of the operative field. In a postal survey, only a third of UK renal transplant units utilised the IVC conduit. Despite 81.5% of units claiming that they harvest the IVC during organ retrieval, a 2-year retrospective audit revealed that only 4.3% of imported right kidneys had the IVC. The IVC remains a much under-utilised resource in the UK despite its potential benefit as a venous conduit in transplanting the right cadaveric kidney. We urge all retrieving surgeons to routinely harvest the IVC with right cadaveric kidneys during organ procurement. PMID- 11111964 TI - Combined treatment of hypercholesterolemia of renal transplant allograft recipients with fluvastatin and gemfibrozil. AB - The aim of this study was to investigate the safety and efficacy of combined treatment with fluvastatin (F) and gemfibrozil (G) in hypercholesterolemic renal transplant recipients (RTR). Ten hypercholesterolemic (total cholesterol [TC] > 220 mg/dl) RTR (7 men) with mean age 44 years (range 25-56 years) who remained hypercholesterolemic after 3 months of treatment (period A) with fluvastatin (40 mg/d) continued taking the same dose of F plus G (600 mg/dl) for another 3-month period (B). Serum total cholesterol, high density lipoprotein cholesterol (HDL C), LDL cholesterol (LDL-C), triglyceride, serum creatinine (creatinine phosphokinase (CPK), serum glutamic-oxaloacetic transaminase (SGOT), and serum glutamate pyruvate transaminase (SGPT) were measured before treatment and at the end of periods A and B. Mean TC levels were 360.30 +/- 62.42 mg/dl, 324.10 +/- 100.53 mg/dl, 270.80 +/- 67.77 mg/dl; mean LDL-C levels were 259.33 +/- 71.43 mg/dl, 219.60 +/- 81.31 mg/dl, 189.70 +/- 65.51 mg/dl; mean HDL-C levels were 37.10 +/- 11.68 mg/dl, 39.80 +/- 13.21 mg/dl, 41.00 +/- 12.94 mg/dl; mean triglyceride levels were 354.60 +/- 183.29 mg/dl, 349.30 +/- 242.94 mg/dl, 207.00 +/- 85.35 mg/dl before treatment and at the end of periods A and B, respectively. There was a statistically significant fall of serum TC (P = 0.002), LDL-C (P = 0.016), and triglyceride (P = 0.029) levels at the end of periods A and B. Kidney and liver function did not change. F and G combined treatment is safe and useful in patients who do not respond satisfactorily to monotherapy with F. Gemfibrozil augments the effect of F on TC, LDL-C, and triglyceride levels. PMID- 11111965 TI - Randomised open clinical trial of conversion from mycophenolate mofetil to azathioprine in cadaveric renal transplantation. AB - Mycophenolate mofetil (MMF) is a powerful immunosuppressive drug with established efficacy and safety. The search for a less expensive immunosuppressive protocol has led to an open randomised clinical trial of conversion from MMF to azathioprine (Aza). A total of 28 renal allograft recipients treated with prednisone, cyclosporine, and MMF was randomised into two groups: converted (early conversion) and control (late conversion). Conversion from MMF to Aza was conducted at the end of the 4th post-transplant month in the converted group and after the 12th month in the control. During the 20-month observation period, biopsy-proven acute rejection occurred more frequently in the converted than in the control group, although the difference was not statistically significant. Early conversion from MMF to Aza increased the risk of subsequent rejection in those patients who underwent at least one episode of acute rejection prior to conversion. PMID- 11111967 TI - Risk of early renal allograft failure is increased for patients with antiphospholipid antibodies. AB - Renal allograft thrombosis can cause transplant failure. Because antiphospholipid antibodies (aPA) are associated with thrombosis, we investigated pretransplant sera from patients with early renal allograft failure to determine if aPA were present. Fifty-six final cross-match (FxM) sera from patients whose transplant failed within 16 days were compared to FxM sera from the next sequential transplant patients. The sera were tested for IgG, IgM, and IgA antibodies to cardiolipin, phosphatidylserine, and phosphatidylethanolamine. aPA were identified in 57% of FxM sera from patients with early non-function versus 35% of FxM sera from patients with functioning grafts (P = 0.02). Historical sera from 11 aPA-positive patients contained aPA up to 18 months prior to transplantation. Since aPA were present in historical sera, testing for aPA can identify certain patients at risk for early allograft failure. The involvement of aPA in early allograft loss is supported by studies demonstrating aPA recovery from an explanted failed transplant. PMID- 11111966 TI - Clinicopathological evaluation in non-episode biopsies of renal transplant allograft. AB - Histopathological findings in renal allograft with stable function remain unclear. We therefore performed non-episode biopsy in the long-surviving renal allograft to investigate the histopathological changes. Our data show that, although arteriolopathy is characteristic of drug-induced nephropathy, it is unrelated to dosage and concentration of cyclosporine or tacrolimus in non episode biopsy. We evaluated therefore the clinicopathological findings of arteriolopathy in this study. Non-episode biopsy was defined as follows: as serum creatinine level lower than, 2.0 mg/dl and a urinary protein level lower than 500 mg/day. A total of 65 biopsy specimens were enrolled in this study as non-episode biopsy. Twenty-nine specimens revealed arteriolopathy. There were no statistically significant differences between arteriolopathy and dosage or concentration of cyclosporine or tacrolimus. Arteriolopathy in non-episode biopsy was related to time of biopsy, kidney age, hypertension, and hyperlipidemia, suggesting that it is important for graft survival to strictly control blood pressure and blood lipid level. PMID- 11111968 TI - Withdrawal of cyclosporine in renal transplant recipients with acute tubular necrosis improves renal function. AB - In this study, patients with acute tubular necrosis (ATN) after renal transplantation were prospectively randomized to either conventional immunosuppression or withdrawal of cyclosporine and replacement with anti thymocyte globulin (ATG). The patients treated with cyclosporine withdrawal and ATG had a significantly shorter duration of ATN (8.9 +/- 1.5 vs 10.8 +/- 1.4 days; P < 0.05) and better renal function (mean serum creatinine on day 5 postoperatively: 740 +/- 49 vs 918 +/- 73 micromol/l; P < 0.05). The incidence of acute rejection was lower in the patients with cyclosporine withdrawal and ATG. In conclusion, cyclosporine is toxic to the renal allograft with ATN, and withdrawal of cyclosporine shortens the duration of ATN and improves renal function. PMID- 11111970 TI - Worse outcome in younger adult renal graft recipients with HCV infection. An 8 year prospective study. PMID- 11111969 TI - Effect of increased arterial resistance index on long-term outcome of well functioning kidney grafts. AB - An abnormal vascular status is present in the transplanted kidney. To define whether vascular factors might influence kidney function of the graft, the renal volume, blood flow and vascular resistance of a group of healthy subjects were compared with those of a group of well functioning renal transplants by color Doppler ultrasonography. Sixty healthy subjects and 75 well functioning cadaver renal transplant recipients were compared by color Doppler ultrasonography. Subsequently, 15 couples of donors and recipients of a living related renal graft were compared to observe the differences between the two organs of the same subject in a different environment. The variables studied were: the diameters and the volume of the kidney, renal blood flow and renal resistance index (RI). The group of cadaver renal transplant patients showed higher mean blood pressure (P = 0.009), higher serum creatinine levels (P = 0.0001) and lower endogenous creatinine clearance (P < 0.0001) than healthy controls. The length (P < 0.00001) and volume (P < 0.001) of the kidneys of cadaver transplanted patients were significantly greater than those of healthy subjects, while the length and volume of the living donors kidneys were identical to those of the recipients. RI, measured on renal vessels, showed lower values in healthy subjects and in kidney donors than in transplanted patients (P < 0.00001). Well functioning transplanted kidneys showed increased renal arterial RI. This non-immunologic factor did not appear to be detrimental with renal function in time, at least until 50 months after successful grafting. PMID- 11111971 TI - Does rural follow-up of renal allografts give impaired graft survival? AB - In this study, we compared the patient and graft survival after renal transplantation in patients followed up in rural centers against those in a major transplant center. There was a greater proportion of patients having a living related donor transplant and having prolonged cold ischemic times in the group followed up in a rural centre. The patient and graft survival at 1, 3 and 5 years were similar for local and rural patients. We conclude that a centralized transplant unit with follow-up of patients in rural centers optimizes the use of highly skilled personnel. PMID- 11111972 TI - Transplantation of both kidneys from 408 donors; comparison of results. AB - The outcome of 816 paired kidney transplantations from 408 cadaveric donors was evaluated. The transplantations were divided according to order of transplant surgery into group 1 [mean cold ischemia time (CIT) 22 h] and group 2 (mean CIT 28 h). In group 1 the frequency of delayed onset of graft function (DGF) was 22% versus 35% in group 2 (P < 0.005). The 1-year patient survival and graft survival (GS) in group 1 was 98% and 93% versus 94% (P < 0.005) and 90% in group 2. Hemodialysis patients in group 2 had significantly greater DGF (43%) and poorer GS (88%) than peritoneal dialysis patients and the success of transplantation was particularly poor in recipients over 50 years of age. PMID- 11111973 TI - Application of Prastat ELISA in the determination of anti-HLA specificity for immunized patients awaiting kidney transplant: five years' experience. AB - Three hundred sixty-five patients who underwent cadaver donor kidney transplantation between 1993 and 1998 were divided into four groups: 40 immunized patients with at least one peak panel-reactive antibody (PRA) value more than 50%, 11 hyperimmunized patients with more than three peak PRA values over 50%, 10 retransplanted patients and 304 control patients. Before transplantation, we ascertained the antibody specificities against individual HLA antigens (Prastat Sangstat ELISA method for HLA typing of first donor, husbands of multiparous women and potential donors against whom candidates gave positive cross-matches); thus, patients underwent transplantation excluding the presence of the HLA antigens previously detected and looking for high HLA (class I and II) compatibility. Actuarial graft survival after 12 months was satisfactory in all groups: 87% immunized, 81% hyperimmunized and 80% retransplanted vs 92% controls. Renal function at the end of the first year was similar and the number of rejection episodes in the first 3 months did not significantly differ. PMID- 11111974 TI - Prognostic value of the Banff classification. AB - We evaluated whether classification of renal allograft biopsies according to the Banff schema is a predictive parameter for graft survival. All patients who received renal transplants between 1980 and 1994 at the University of Erlangen Nuremberg (n = 1141) were included. Patients who had undergone a renal biopsy (n = 306) were divided into groups according to the Banff classification. We observed a correlation (P < 0.05) between biopsy findings and the following patient characteristics: donor/recipient age, donor/recipient gender, panel reactive antibodies, maintenance immunosuppression, and primary renal disease. Compared to patients who did not undergo renal biopsy (55.9%), 5-year graft survival was reduced in patients with moderate acute rejection defined by tubulitis (20.6%, P = 0.03) or arteritis (0%; P < 0.0001) and in patients with severe acute rejection (24.4%, P < 0.0001). CONCLUSIONS: (1). The Banff classification is a predictive parameter for renal allograft survival. (2). Certain characteristics predispose patients to certain biopsy findings. PMID- 11111975 TI - An objective method for detecting time-dependent effects in graft survival. AB - The proportional hazards model has become increasingly important in the analysis of censored survival data after transplantation. Neverthless, in clinical transplantation it is still undefined how the influence of covariates changes over time. The additive regression model is an alternative (or extension) to the Cox model. It results in plots (Aalen plots) that may give information on the effect of covariates over time by way of the cumulative regression function plots. A total of 386 primary cadaveric kidney transplants performed between 1984 and 1996 were included in our analysis. The follow-up period ranged from 24 to 156 months. According to Aalen, an additive regression model was used and plots for the detection of time-dependent effects of covariates were determined. Patients dying with functioning grafts were classed as graft failures. Factors potentially affecting graft outcome, such as sex, donor and recipient's age, HLA A-B match, cold ischaemia time (CIT), delayed graft function (DGF), serum creatinine at 1 month (Cr1), rejection episodes within 3 months (R3), and type of brain death (BD), were considered. The slopes of the plots by donor age, DGF, HLA A-B match, R3, Cr1 and BD appear to have a significant influence throughout the observation period, with different time-dependent effects on graft survival. Slopes for DGF, Cr1, and age of donor are positive (increased hazard), while slopes for HLA match and BD are negative (decreased hazard). Estimated regression functions for DGF, donor age and Cr1 show a prompt slope (within 3 months); the covariate R3 has a clear influence for about 5 years, and then seems to disappear; and BD appears to have a consistent effect over the entire period. The additive regression model with Aalen plots represents a useful technique in the analysis of survival data after kidney transplantation. Some covariates, such as R3, may often lose their effects on graft survival, with a relevant clinical impact. Others have a clear and additive influence over the entire period (BD), while the effects of donor age, DGF and CR1 each appear to have a prompt effect in the outcome. PMID- 11111976 TI - The influence of cholesterol on mortality after transplantation is age dependent. AB - There is still no consensus on the treatment of elevated serum cholesterol in patients with a renal transplant. In the general population treatment is age dependent. We studied the influence of serum cholesterol 1 year after transplantation in all 676 recipients of a kidney graft transplanted in Rotterdam that survived and functioned for at least 1 year. The other variables included in this analysis are: donor and recipient age and gender, original disease, race, number of HLA A and B mismatches, number of previous transplantations, postmortal or living related transplantation and transplantation year. At 1 year after transplantation the following variables were included: serum cholesterol, serum creatinine, proteinuria and hypertension. In the Cox proportional hazards analysis, serum cholesterol at 1 year after transplantation turned out to be an important, independent variable influencing patient failure. The influence was linear but there was interaction with recipient age. The negative influence of serum cholesterol on the RR for patient failure decreased with increasing recipient age. For example, the proportional increase in RR of a 20-year-old with a serum cholesterol of 12 mmol/l compared with that of a cholesterol of a patient with serum cholesterol of 6 mmol/l was 6. In a 60-year-old with a cholesterol of 12 mmol/l the proportional increase in RR was only 1.2 compared with a contemporary with a cholesterol of 6 mmol/l. Serum cholesterol levels have an independent influence on patient failure. The RR is influenced by recipient age, so that the negative effect of increasing cholesterol levels in the elderly is overruled by the RR of age and disappears. PMID- 11111977 TI - "Tolerogenic effect" of the liver for a small bowel allograft. AB - A newly developed liver/small bowel transplantation model (LSBTx) was used to investigate the tolerogenic effect of a liver allograft toward a simultaneously transplanted small bowel. Small bowel transplantation (SBTx) under high-dose immunosuppression was compared to LSBTx with a lower FK506 dosage. Syngeneic Lewis [(LEW) to LEW] and two fully allogeneic rat strain combinations (Brown Norway-to-LEW and Dark Agouti-to-LEW) were used. Clinical course and histological findings after SBTx demonstrated a chronic rejection of the small bowel allograft within 100 days. However, after LSBTx long-term acceptance (> 150 days) was achieved after a transient rejection crisis, although initial immunosuppression was significantly lower. Furthermore, indicator heart transplantations demonstrated the induction of donor-specific tolerance in both allogeneic strain combinations. In contrast to other LSBTx rat models, these results reflect observations after human LSBTx, in which the rate of acute and chronic rejection is also significantly lower than after human SBTx. PMID- 11111978 TI - Liver transplantation for alcoholic cirrhosis. AB - Because of the donor shortage, there are concerns for liver transplantation in patients with alcoholic cirrhosis. We therefore analyzed patients transplanted for alcoholic cirrhosis at our center with respect to patient and graft survival, recurrence of disease, and postoperative complications. Out of 1000 liver transplantations performed in 911 patients, 167 patients were transplanted for alcoholic cirrhosis; 91 patients received CsA- and 76 patients FK506-based immunosuppression. Recurrence was diagnosed by patient's or relative's declaration, blood alcohol determination, and delirium. Diagnosis and treatment of acute and chronic rejection was performed as previously described. One- (96.8% versus 91.3%) and 9-year patient survival (83.3% versus 80%) compared well with other indications. Five of 15 patients died due to disease recurrence. Recurrence of disease was significantly related to the duration of alcohol abstinence prior to transplantation. In patients who were abstinent for less than 6 months (17.1%), recurrence rate was 65%, including four of the five patients who died of recurrence. Recurrence rate decreased to 11.8%, when abstinence time was 6-12 months and to 5.5%, when the abstinence times was > 2 years. Next to duration of abstinence, alcohol relapse was significantly related to sex, social environment, and psychological stability. The incidence of acute rejection compared well with other indications (38.1%); CsA: 40.1% versus 33.3% in FK506 patients. In all, 18.2% of CsA patients experienced steroid-resistant rejection compared with 2.6% of FK506 patients. Seven patients (7.6%) in the CsA group and one patient (1.3%) in the FK506 group developed chronic rejection. A total of 57.1% developed infections; 5.7% were life-threatening. CMV infections were observed in 14.3% (versus 25% for other indications). New onset of insulin-dependent diabetes was observed in 8.6% and hypertension in 32.4%. In conclusion, alcoholic cirrhosis is a good indication for liver transplantation with respect to graft and patient survival and development of postoperative complications. FK506 therapy was favourable to CsA treatment. Patient selection is a major issue and established criteria should be strictly adhered to. Patients with alcohol abstinence times shorter than 6 months should be excluded, since recurrence and death due to recurrence was markedly increased in this group of patients. PMID- 11111979 TI - Liberal policy of split liver for pediatric liver transplantation. A single centre experience. AB - We adopted a liberal policy of extensive use of split liver in a pediatric liver transplantation (LT) program. Over a 19-month period, we have performed 64 LT in 54 patients with pediatric indications. One patient received two liver grafts as a part of a liver-small bowel transplantation and was not considered. Of the 60 LT considered, performed in 53 patients, 34 were with split grafts. The 1-year actuarial survival for the patients transplanted with a split graft was 81% and 89% when only elective cases were considered. The median time on the waiting list was 22 days with no mortality. The extensive use of split liver allowed transplantation in a large number of pediatric patients, with good results without the need for living donor liver transplantation. We envisage a trend towards systematic splitting of liver grafts. PMID- 11111980 TI - Evolution of living donor liver transplantation in adults: a single center experience. PMID- 11111981 TI - Factors affecting survival after living-related liver transplantation. AB - The purpose of this study was to determine the perioperative factors that influence patient and graft outcome in living-related liver transplantation (LRLT). Between April 1995 and October 1998, we performed a series of 46 LRLT procedures, including 11 adult cased, at our institute. Mean age and weight of the recipients were 12.0 +/- 2.3 years and 23.7 +/- 2.6 kg, respectively. Seven out of the 46 patients had renal failure and received hemodialysis therapy before and after LRLT or kidney transplantation. The recipients were divided into two groups: those who survived for 7-48 months after LRLT (group 1, n = 36), and those who died within 4 months after surgery (group 2, n = 10). Factors analyzed included recipient age and weight, graft/recipient body weight ratio (G/R ratio), emergent vs elective surgery, United Network for Organ Sharing (UNOS) status, presence of preoperative plasmapheresis (PEX) and renal failure, and so on. Recipients in group 1 compared with group 2 had less advanced liver disease (i. e., a lower rate of emergent surgery, 14% vs 50%, and fewer patients with UNOS status 1, 14% versus 70%; P < 0.05 and P < 0.001, respectively). Group 1 recipients also had a lower percentage of preoperative treatment with plasmapheresis (22% vs 70%, P < 0.01). However, neither the G/R ratio nor the presence of renal failure affected the patient survival rate. In conclusion, factors independently associated with reduced patient survival after LRLT include emergent surgery, Child-Pugh class, UNOS status 1, and preoperative plasmapheresis. PMID- 11111982 TI - Does a visual analogue colour chart carried by the retrieval team help in assessment of the fat content of donor livers? PMID- 11111983 TI - Split liver is an effective tool to transplant paediatric patients. AB - Transplantation activity is dependent upon organ procurement; although great efforts are made to enlarge the cadaver donors' pool, it still remains far too small to meet the recipients' need. Waiting time is a particular problem for paediatric patients, and mortality on the waiting list for liver transplantation is very high. The number of paediatric donors is far too small to satisfy the request. To enlarge the liver pool, the split-liver procedure was introduced in several Transplant Centers. In November 1997, the North Italy Transplant program (NITp) Working Group for Liver Transplantation decided to start an official Split liver Program. A protocol was therefore defined and criteria for donor's and recipient's eligibility were established to minimize the risk. The Working Group also standardized the technical procedure and defined collaboration between centers. Out of 410 cadaver liver donors used in the NITp, from 1 November 1997 until 31 May 1999, 49 patients (37 males and 12 females) were chosen for the split-liver procedure. Mean age was 29.9 +/- 17.5 years. Mean ICU stay of the donors was considerably short (2.5 +/- 2.1 days), and the other conditions foreseen for donor eligibility were met. In all cases (except two) an "in situ" technique was performed. Forty-nine adult recipients and 43 children were transplanted by the split-liver technique in our Transplant Centers. One right lobe and five left liver lobes were sent to Transplant Centers outside the NITp. Adult recipient age ranged from 18 to 60 years (mean 46.4 +/- 11.7 years), and the paediatric one from 2 to 144 months (mean 24.8). Mean patient follow-up was 8.3 +/- 5.5 months. In the paediatric group, the graft was successful in 34 cases (79%), five patients (10.2%) died and four (9.3%) were re-transplanted. In the adult group, graft survival was 67.3%, 11 (22%) patients died and 5 (10%) were re transplanted. On 1 November 1997, 30 paediatric patients were on the liver waiting list. In the preceding 19 months, 52 patients were newly enrolled, and 36 transplants were performed. The mean waiting time of paediatric patients was 259 days (range 1-919 says). From 1 November 1997 to 31 May 1999, 61 paediatric patients were newly enrolled. In this period 70 patients were transplanted. The mean waiting time was 185 days (1-1010 days). At present, the liver waiting list includes eight paediatric patients. Split-liver transplantation is a successful procedure, effective in reducing waiting time for paediatric patients. It should be established if this may be a tool to enlarge the organ pool also for adult liver transplantation. PMID- 11111984 TI - RANTES in the postoperative course after liver transplantation. AB - RANTES (regulated upon activation, normal T-cell expressed and secreted), an inflammatory cytokine, promotes accumulation and activation of leukocytes. In 67 liver transplantations, systemic concentrations of RANTES were correlated to graft survival and incidence of rejection. RANTES levels either increased to highly elevated levels at day 14 (84 +/- 64 ng/ml; group 1; n = 43) or remained within the limit of healthy controls (19 +/- 11 ng/ml at day 14; group 2; n = 24). The 100-day graft function rate was 0.91 in group 1 and 0.63 in group 2 (P = 0.002). The risk ratio for rejection during the first 100 days was increased 2.2 fold in group 2 compared to group 1 (P = 0.02). High postoperative release of RANTES after liver transplantation, a beneficial factor, may reflect a general systemic immunological activation. It can be concluded that high early systemic RANTES levels may play a role in immunological recognition leading to a tolerance of the liver graft. PMID- 11111985 TI - Analysis of risk factors following pediatric liver transplantation. AB - Several recipient, donor and operation factors as well as postoperative complications related to patient survival after liver transplantation (LT) in children were studied by univariate and multivariate analyses. In a 13-year period, 103 patients under 15 years of age underwent 120 LT; the mean age was 63 months and 36% were under 2 years of age. Indications for LT were cholestatic disease in 68 (56%), metabolic diseases in 18 (14%), fulminant hepatic failure in 8 (7.5%), cirrhosis in 7 (5.8%), and retransplants in 17 (14%). Whole liver was transplanted in 79% of cases and partial liver in 21%. Actuarial survival at 1, 5, and 10 years was 70%, 61%, and 57%, respectively. United Network of Organ Sharing (UNOS) I recipients (RR = 2.7), primary non-function (PNF) (RR = 13.9), and hepatic artery thrombosis (HAT) (RR = 3.8) were independent factors for lower patient survival in multivariate analysis. Thus, in our experience, postoperative mortality as a consequence of the patient's condition before transplantation, or complications such as PNF or HAT, are the major causes of decreased survival in pediatric LT. PMID- 11111986 TI - Organ survival after primary dysfunction of liver grafts in clinical orthotopic liver transplantation. AB - In a retrospective analysis of 632 orthototopic liver transplant procedures performed between 1982 and 1997, the incidence of primary dysfunction (PDF) of the liver and its influence on organ survival were studied. Graft function during the first 3 postoperative days was categorized into four groups: (1) good (GOT max < 1000 U/l, spontaneous PT > 50%, bile production > 100 ml/day); (2) fair (GOT 1000-2500 U/l, clotting factor support < 2 days, bile < 100 ml/day); (3) poor (GOT > 2500 U/l, clotting factor support > 2 days, bile < 20 ml/day); (4) primary non-function (PNF; retransplantation required within 7 days). The aim of this study was to evaluate graft survival comparing organs with PDF (poor function) and PNF vs organs with initial good or fair function. After a median follow-up of 45 months, initially good and fair function of liver grafts resulted in a significantly better long-term graft survival compared with grafts with initially poor function or primary non-function (if re-transplanted) (P < 0.01). The Cox model revealed primary function as a highly significant factor in the prediction of long-term graft survival (P < 0.0001). We conclude that these results confirm the hypothesis that primary graft function is of major importance for the long-term survival of liver transplants. Patients with a poor primary function have the worst survival prognosis, which leads to the interpretation that these patients may be candidates for early retransplantation. PMID- 11111987 TI - Perioperative factors influencing patient outcome after liver transplantation. AB - We have previously shown that the development of multiple organ dysfunction syndrome (MODS) after liver transplantation significantly reduced patient survival. Therefore, the question arises of which are the most prominent perioperative donor and recipient factors leading to MODS after transplantation. In total, 634 patients with 700 liver transplants were analyzed. Donor factors included age, increase in transaminases, sex mismatch, requirement for catecholamines, intensive care time, histology, and macroscopic graft appearance. Recipient factors included Child classification, preoperative gastrointestinal (GI) bleeding, mechanical ventilation, hemodialysis, and requirement for catecholamines. MODS was defined by more than two severe organ dysfunctions. The cumulative 2 to 9-year patient survival was 90.9% in patients developing less than 3 severe organ dysfunctions following transplantation. Survival decreased to 60.3% in patients with MODS. Neither any of the donor factors nor the duration of cold ischemia (CIT) was associated with an increase in MODS or decrease in survival. On the other hand, duration of warm ischemia, amount of blood loss, requirement for red packed blood cells, and reoperation had an influence on the development of MODS (40%-56%) and decreased patient survival to 58%-69%. Preoperative therapy with catecholamines, GI bleeding, mechanical ventilation, and hemodialysis were associated with the development of MODS in 54%-88%. Patient survival following MODS decreased to 50%-74%. Initial graft function had a slight influence on the development of MODS, but no influence on the long-term patient survival. In conclusion, patient survival was significantly influenced by the development of postoperative MODS. The most prominent factors in this were recipient and intraoperative ones. No major influence was observed for donor factors, CIT, and initial graft function. Prevention of MODS will further improve the outcome after liver transplantation. PMID- 11111988 TI - Liver transplantation for primary sclerosing cholangitis--a single-center experience. AB - The purpose of this study was to analyze the outcome of liver transplantation for primary sclerosing cholangitis (PSC) at our center. The medical records of 47 consecutive patients transplanted during the period 1985-1996 were reviewed. Actuarial patient survival was determined at 1, 5, and 10 years. Data on reasons for retransplantation, incidental carcinoma, biliary complications that required surgical intervention, and signs of possible disease recurrence were collected. The median follow-up period was 3.6 years. Overall patient survival was 75% at 1 year and 66% at 5 and 10 years. Patients transplanted during the period 1994-1996 (n = 24) had a significantly (P < 0.02) better 1 year (88% vs 61%) and 5-year (83% vs 48%) survival than patients transplanted during the period 1985-1993 (n = 23). Six patients (13%) were retransplanted, and 4 are currently alive. Nine patients (19%) had biliary complications that required surgical intervention. Cholangiocellular carcinoma was found in 4 (9%) explanted livers, and all 4 patients succumbed within 2 years of transplantation. Indications of disease recurrence were seen in 4 patients (9%). In conclusion, the results of liver transplantation for PSC at our center are comparable to those of other benign indications, but a relatively high incidence of biliary complications was noted, and a possible disease recurrence was detected in 9% of patients. PMID- 11111989 TI - Liver transplantation in patients over 60 years of age. AB - Liver transplantation was previously only offered to patients under 60 years of age. We have analyzed the outcome after acceptance on the waiting list and after liver transplantation of patients over 60 years old. A total of 150 patients over 60 years old were listed for a first liver transplantation during 1990-1998. The annual number increased throughout the period. Primary biliary cirrhosis, primary sclerosing cholangitis, and acute hepatic failure were the most frequent diagnoses. A total of 119 patients received a first liver allograft. The patient 1-year survival was 75% and 3-year survival 62%, which was not significantly lower (P = 0.21) than that of the younger patients. When correcting for year of transplantation, the survival was, however, moderately but significantly lower than among the younger patients. Survival among those > 65 years (n = 38) did not differ from that of patients 60-65 years of age (n = 81). We conclude that an increasing number of patients over 60 years old can be listed for liver transplantation and receive a liver allograft with highly satisfying results. PMID- 11111990 TI - Long-term results of liver transplantation in four siblings from the same family with familial amyloidotic polyneuropathy type I TTR Ala-71. AB - Familial amyloidotic polyneuropathy type I (FAP I) is a hereditary systemic amyloidosis usually involving the peripheral nervous system. In this paper we report our experience regarding the survival and the evolution of the sensory motor syndrome of the extremities and autonomic dysfunction in four siblings with the Ala-71 variant who were treated by liver transplantation (LT). The four siblings are alive 2-5 years after LT. After the operation, the seriated determinations of TTR-Ala-71 variant showed a constant decrease in serum levels in all cases. Our results support the proposal that LT should be indicated especially in forms with early clinical onset (3rd and 4th decades) and rapid progress to stop the neurological deterioration of the patients. PMID- 11111991 TI - Long-term medical and psycho-social evaluation of patients undergoing orthotopic liver transplantation for alcoholic liver disease. AB - The major concern in transplanting patients with alcoholic liver disease (ALD) is the high rate of alcohol recidivism. Our aim was to assess the long-term outcome of liver transplantation (OLT) in a group of ALD patients in terms of post-OLT alcohol recidivism and its relationship with pre-OLT psychosocial variables and medical follow up. Fifty-one ALD patients underwent strict medical and psychosocial evaluation before and after OLT. Alcohol abuse was recorded in 60% and alcohol dependence in 40% of patients before OLT. The 5-year survival was similar to the one observed in non-ALD transplanted patients (64 vs 56%). Alcohol recidivism was observed in 33% of transplanted patients, 64% of whom were occasional and 36% were heavy drinkers. The admission of alcoholism by the patient and his/her family prior to OLT significantly predicted abstinence after OLT. A multidisciplinary approach evaluating medical and psycho-social variables before OLT and a close follow up after OLT are mandatory for ALD patients. PMID- 11111992 TI - Small bowel transplantation using grafts from living-related donors. Two case reports. AB - A living-related small bowel transplantation (SBT) was performed in two pediatric patients with short bowel syndrome. In both cases, the donor was the patient's mother. The distal ileum (100 cm, 120 cm) was harvested and the ileocolic vessels, ileocecal valve, and terminal ileum were left intact. The two donors were discharged from the hospital on postoperative days 15 and 6, respectively. Recipient 1 was a 2 year 6 month-old boy with short bowel syndrome who underwent SBT due to loss of venous access. The graft vein was anastomosed to the recipient's infrarenal inferior vena cava. Despite triple immunosuppression (tacrolimus, steroid, and azathioprine), there were four episodes of rejection. The patient had been on total parenteral nutrition for almost his entire posttransplant course. He died from Pneumocystis carinii pneumonia 16 months after the transplantation. Recipient 2 was a 4 year 5 month-old girl with short bowel syndrome who underwent an isolated small bowel transplantation because of recurrent line sepsis. Her pretransplant bilirubin was 8.0 mg/dl and a biopsy showed severe fibrosis. The graft vein was anastomosed to the recipient's inferior mesenteric vein. After transplantation, her bilirubin level became normal within 10 days. Triple immunosuppression (tacrolimus, steroid, and cyclophosphamide) together with a 3-day course of OKT-3 made her post-transplant course feasible. After overcoming a single episode of rejection she left the hospital 4 months after SBT. The patient is currently (10 months after transplantation) hospitalized due to rejection, which is being successfully controlled, and she is off total parenteral nutrition. From our experience, harvesting of the distal ileum for use as a bowel graft can be safely performed. The advantages of living-related grafts, optimal graft length, and choice of vascular reconstruction in SBT are yet to be explored. PMID- 11111993 TI - Vascular complications following bladder drained, simultaneous pancreas-kidney transplantation: the University of Miami experience. AB - Vascular complications remain a significant nonimmunologic source of pancreas allograft loss. From February 1993 through January 1998, we performed 98 simultaneous pancreas-kidney transplantations (SPK) using pancreatic exocrine bladder drainage in patients with type 1 insulin-dependent diabetes mellitus and end-stage renal disease. They originally received quadruple immunosuppression, and since May 1997 triple immunosuppression protocol (tacrolimus, mycophenolate mofetil, and steroids). The patients' mean age was 37 years (range 24-53 years), including 50 women and 48 men with a mean follow-up of 42 months. The overall rate of vascular complications was 6% (5 patients). The vascular complications were as follows: late thrombosis of the Y with persistent pancreas allograft function (n = 1), rupture of a pseudoaneurysm of the superior mesenteric artery (PSMA) with an arteriovenous fistula (AVF) (n = 1), thrombosis of the splenic vein (SV) (n = 3), complete thrombosis of the superior mesenteric vein (SMV) and splenic vein (n = 1). The patient with PSMA underwent surgical correction of the AVF and PSMA with preservation of the allograft pancreas function. The other patient with late thrombosis of the Y-graft required no treatment. All 3 patients with SV thrombosis were systemically heparinized followed by oral anticoagulation. The patient with complete thrombosis required surgical thrombectomy of the SMV and SV followed by heparinization and oral anticoagulation. All 6 patients including the 4 with thrombosis had preservation of the pancreas function. Serial pancreas ultrasound showed resolution and improvement with recanalization of the splenic vein and superior mesenteric vein in those patients with thrombosis. We describe our vascular experience with salvage of the pancreatic allograft function. Surgery seems to be the best treatment option in the case of AVF or complete thrombosis of the allograft. Intravenous heparin followed by oral anticoagultion could be a conservative approach for SV thrombosis. PMID- 11111994 TI - Use of tacrolimus and mycophenolate mofetil as induction and maintenance in simultaneous pancreas-kidney transplantation. AB - Clinical trials using quadruple immunosuppression that include the combination of tacrolimus and mycophenolate mofetil have been shown to reduce the incidence of acute rejection episodes in simultaneous pancreas-kidney (SPK) transplantation. In an attempt to obtain a low rejection rate without antibody induction therapy, we undertook a prospective study of combined tacrolimus and mycophenolate mofetil and steroids as primary immunosuppression for SPK transplantation. In this study, we analyzed 17 patients who received low-dose intravenous tacrolimus as induction therapy. This was combined with oral tacrolimus, mycophenolate mofetil, and steroids as the primary immunosuppression regimen. There was a significant reduction of empirically and biopsy-proven rejection with an incidence of 23% (4 patients). Leukopenia, gastroparesis, and gastrointestinal side-effects were the cause of discontinuation of mycophenolate mofetil, or low tacrolimus trough level in those patients who developed rejection. All rejection episodes were easy to treat, and none of them required antibody therapy. The combination of tacrolimus with mycophenolate mofetil without antibody induction therapy is effective in preventing early acute rejection. This combination is safe and effective as an alternative immunosuppressive regimen after SPK transplantation. PMID- 11111996 TI - Does splitting the lung block into two single lung grafts equate to doubling the societal benefit from bilateral lung donors? Comparisons between two single versus one bilateral lung transplant. UK Cardiothoracic Transplant Audit Steering Group. AB - Multi-organ thoracic transplantation, although beneficial to one recipient, has an opportunity cost of denied transplants to others. This paper compares population based outcomes of splitting lung blocks for two single lung transplants compared to doing one bilateral lung transplant, and suggests that the benefit of splitting lung blocks may not necessarily be double that of using each block for one recipient. PMID- 11111995 TI - Incidence of intraabdominal infection in a consecutive series of 40 enteric drained pancreas transplants with FK506 and MMF immunosuppression. AB - Although the introduction of FK506 and MMF has markedly improved patient and graft outcome after pancreas transplantation, this procedure is still associated with a high surgical complication rate. The aim of the following study was to retrospectively analyze a series of 40 consecutive pancreas transplants with enteric drainage with regard to intraabdominal infection (IAI). Between March 1997 and December 1998 a total of 40 whole pancreas transplants were performed. Prophylactic immunosuppression consisted of an intraoperative single shot ATG (Thymoglobulin), FK506, MMF, and prednisone. The mean observation period was 14.6 (5-26) months. Overall incidence of IAI was 27.5% (n = 11) leading to pancreatectomy in 5 patients (12.5%). In the remaining 6 patients the graft could be rescued by necrosectomy and radical drainage of the abscess (5 patients) or percutaneous drainage (1 patient). Pancreatectomy or local infection did not alter kidney graft function in the 11 patients with simultaneous pancreas kidney transplantation. In 10 patients no evidence for leakage at the site of enteric anastomosis was present, one duodenal leak occurred due to ischemia. IAI in the early postoperative period was the predominat risk factor for graft loss. An early and invasive diagnostic approach is recommended to maximize the chance of graft rescue. PMID- 11111997 TI - Effects of Celsior and University of Wisconsin preservation solutions on hemodynamics and endothelial function after cardiac transplantation in humans: a single-center, prospective, randomized trial. AB - Optimal preservation of the myocardium remains a major concern in clinical and experimental heart transplantation. The present study compared the efficacy of University of Wisconsin (UW) and Celsior preservation solution with respect to myocardial performance, epicardial and microvascular endothelial vasomotor function and myocardial expression of endothelin and nitric oxide synthases in humans. Forty-one cardiac transplant recipients received either UW (n = 20) or Celsior (n = 21) preserved hearts. Catecholamine and vasodilator requirements were assessed within the first 5 postoperative days. Left ventricular performance and endothelial function was assessed 1 month after transplantation. Endothelin and nitric oxide synthase gene expression were detected in myocardial biopsy samples. Celsior preserved hearts required significantly more catecholamines and vasodilators within the first 5 postoperative days. Myocardial performance and endothelial function were comparable 1 month after transplantation. Total ischemic time correlated with impaired endothelial function in the Celsior but not in the UW group. Endothelin and inducible nitric oxide synthase gene expression were significantly higher in the Celsior group. The results of the study show that both solutions provide myocardial protection with regard to left ventricular performance and endothelial function 1 month after cardiac transplantation. The necessity for higher vasodilator and catecholamine therapy in Celsior preserved hearts suggests post-ischemic myocardial stunning within the first 5 postoperative days. The positive correlation between impaired endothelial function and total ischemic time in the Celsior group requires longitudinal investigation in particular with regard to the development of allograft vasculopathy. PMID- 11111998 TI - Quantitative flow cytometry to measure the TNF-alpha and IL-2 system after heart transplantation. AB - After heart transplantation a high incidence of infections and malignancies is found. Not only immunosuppression, but also intrinsic cytokine systems with some unbalance, e. g. TNF-alpha and IL-2, can result in impaired immune competence and may have a role in these complications. The aim of this study was to assess the activity of the TNF-alpha and IL-2 systems after heart transplantation. In peripheral blood we measured expression of activation markers of TNF-alpha (TNF R2) and IL-2 (IL-2R alpha, IL-2R beta-chain) on monocytes and lymphocytes using quantitative flow-cytometric analysis. TNF-R2 expression was significantly enhanced on monocytes and lymphocytes in patients after heart transplantation, while the expression of IL-2R alpha and IL-2R beta was not elevated. Increased TNF-R2 expression in peripheral blood after heart transplantation reflects an activated TNF-alpha system, leading to high levels of active sTNF-R, which impairs TNF-alpha bioavailibility and consequently leads to immune incompetence. PMID- 11111999 TI - High frequency of IL-4 producing helper T lymphocytes associated with a reduced incidence of heart allograft rejection. AB - The reduction in the frequency of rejection episodes several months after heart transplantation (HTX) correlates with the development of donor-specific nonresponsiveness. This is reflected in a reduced frequency of donor-specific cytotoxic T cells (CTL) in the peripheral blood. We investigated whether the reduced CTL frequency and the incidence of rejection episodes coincided with a change in the frequency of either IL-2- or IL-4-producing helper T lymphocytes (HTL). We measured the frequency of HTL before and at several time points after HTX in the blood of ten recipients, using limiting dilution analysis for IL-2 and IL-4. In most patients, HTL frequencies dropped immediately after transplantation, but returned to pre-HTX values later after transplantation. No consistent decrease or increase in frequencies was observed long after HTX. In contrast to IL-2, the HTL frequencies for IL-4 before transplantation were significantly higher in patients without post-HTX rejection episodes requiring treatment than in patients with such episodes. This phenomenon was observed for the in vitro responses towards both donor and third-party cells. In conclusion, relatively high frequencies of IL-4-producing T cells may have a beneficial effect on the outcome of human heart transplantation, because they are associated with a reduced incidence of rejection episodes after transplantation. PMID- 11112000 TI - Non-invasive graft monitoring after heart transplantation: rationale to reduce the number of endomyocardial biopsies. AB - The endomyocardial biopsy is invasive, reduces quality of life and cannot be repeated daily. Initial studies on noninvasive cardiac graft monitoring have been presented recently. During the heart transplant procedure, we implanted wideband telemetric pacemakers and fractally coated, epimyocardial electrodes. On biopsy days and during each follow-up, intramyocardial electrogram sequences were obtained. The maximum T-slew rate from the ventricular evoked response (VER) was automatically calculated and compared to the biopsy results (n = 331, ISHLT grading). The VER T-slew rate was significantly lower during rejection grade 2 or higher. The negative predictive value to exclude rejection was 98%. Using a single threshold diagnosis model, 74% of the biopsies could have been avoided. Noninvasive cardiac graft monitoring can reduce the need for surveillance biopsies and may offer a tool to optimize immunosuppressive therapy after heart transplantation. PMID- 11112002 TI - Elevated plasma homocysteine concentrations after pediatric heart transplantations. AB - Graft coronary artery vasculopathy is the main cause of late morbidity and mortality in cardiac allograft recipients. A high plasma homocysteine (hcy) concentration is now generally accepted as a risk factor for coronary arteriosclerosis, but little information exists for the pediatric age group. We therefore explored the potential role of hcy and antioxidants in 31 pediatric allograft recipients. We found hcy concentrations to be significantly higher in recipients than in control. Hcy continued to rise within the first 2 postoperative years. Vitamin A and E concentrations were significantly lower in transplant patients. Hyperhomocysteinemia was significantly more common in patients with complications than in those without. Our findings suggest that pediatric allograft recipients experience oxidant stress, as highlighted by the high plasma levels of Hcy and the low concentrations of vitamins A and E. Nutritional supplementation may be indicated to lower plasma hcy and to reduce oxidant stress. PMID- 11112001 TI - An association between microvascular endothelial dysfunction, transcardiac nitric oxide production and pro-inflammatory cytokines after heart transplantation in humans. AB - Endothelial dysfunction anticipates the development of transplant coronary artery disease (TxCAD) observed more than 1 year after transplantation (HTx). We investigated whether in patients early after HTx myocardial inducible and constitutive nitric oxide synthases (iNOS; cNOS) are expressed and cardiac nitric oxide production occurs. Moreover, a possible relationship to alterations in endothelium dependent and independent vasomotor function was assessed. Forty-two transplant recipients were studied 37 +/- 5 days after HTx. Microvascular coronary flow velocity reserve (CFVR) was tested endothelium dependent (acetylcholine; 30 microg/min x 5 min/i.c.) and independent (adenosine; 160 microg/min x 5 min/i.c.) by Doppler flow wire. Flow velocity increase by a factor greater than 2 was considered normal. Quantitative coronary angiography was used to assess epicardial vasomotor function in response to the same stimuli. Myocardial iNOS and cNOS gene expression were detected by semiquantitative reversed transcriptase polymerase chain reaction. Plasma nitrite levels (microM) were measured by spectrophotometry. Cytokines (TNF-alpha, IL-6; pg/ml) were measured by enzyme linked immunosorbent assay. In 26.1% of patients (n = 11; group A) an impaired endothelium dependent CFVR (1.65 +/- 0.23 increase) was observed; in 73.9% (n = 31, group B) a normal endothelium dependent CFVR (3.0 +/- 0.7 increase; P = 0.003) was observed. Myocardial iNOS and cNOS gene expression did not differ between the two groups. Transcardiac cytokine production was noted in 58.8% of patients for IL-6 and in 53.3% for TNF-alpha. Coronary sinus (CS) levels of TNF-alpha, IL-6 and nitrite were higher in group A. A significant increase in nitrite production was found only in patients with impaired endothelium dependent CFVR (aorta: 43.9 +/- 3.7 vs CS: 52.8 +/- 5.6, P = 0.05), suggesting transcardiac nitric oxide production. In addition, CS nitrite levels correlated with CS TNF-alpha levels in patients with impaired CFVR (r = 0.44, P = 0.003). Microvascular endothelium dependent CFVR is impaired in 26% of patients early after HTx. Activation of cytokines and the NO pathway seem to be involved in this vasomotor dysfunction The association between cardiac nitric oxide production and TNF-alpha in this group indicates a chronic high immunologic process, which may represent an early and important target for therapy and prevention of TxCAD. PMID- 11112003 TI - Continuous retrograde warm blood reperfusion reduces cardiac troponin I release after heart transplantation: a prospective randomized study. AB - During heart surgery, cardiac troponin I (cTn-I) measurement provides a tool to evaluate different cardioprotective techniques. To investigate myocardial protection during heart transplantation (HTx), cTn-I and creatine kinase (CK)-MB release was measured in 42 patients randomized to receving either continuous retrograde warm blood reperfusion or no reperfusion after cold cardioplegia. A significant linear correlation was found between donor heart ischemic time and peaks and the area under the curve of cTn-I and CK-MB release. In patients with an ischemic time longer than 90 min, cTn-I release was significantly lower in those receiving continuous retrograde warm cardioplegia than in controls. No significant difference was observed for CK-MB, tCK, and myoglobin. Our data suggest that the measurement of postoperative cTn-I release may provide a method to evaluate ischemic cardiac damage after HTx. When the ischemic time is longer than 90 min, warm retrograde blood cardioplegia provides better myocardial protection than no reperfusion. PMID- 11112004 TI - Where are we today with pulmonary transplantation? Current results from a national cohort. UK Cardiothoracic Transplant Audit Steering Group. AB - Lung transplantation is now an accepted therapy for the treatment of end-stage lung disease. This paper presents some current results of lung transplantation as determined from a validated national database. PMID- 11112005 TI - A proposal for scoring marginal liver grafts. AB - The aim of this study is to assess the effect of accumulation of marginal liver graft criteria on the immediate outcome of liver transplantation (LT). The last 325 consecutive LT performed in 293 patients were analyzed retrospectively with respect to donor acceptance criteria. A marginal liver score was elaborated on the basis of the following features: donor > 60 years, ICU stay > 4 days, cold ischemia times > 13 h, hypotensive episodes < 60 mmHg > 1 h, bilirubin > 2.0 mg/dl, ALT > 170 U/l, and AST > 140 U/l were scored with the value 1. The use of dopamine doses > 10 microg/kg per min and peak serum sodium > 155 mEq/l were labeled with value 2. The cut-off point at 6 months after LT revealed 42 deaths (14%), with 65 graft losses (20%) and 32 (9%) retransplants. Recipient survival was not affected by the combined effect of marginal criteria. However, recipients transplanted with marginal livers with score 3 or more showed a decrease in graft survival (log-rank 6.21; P = 0.045) and an increase in delayed non-function rate (10 out of 33 vs 4 out of 156; P = 0.03). The use of marginal liver donors with more than three risk factors must be carefully reviewed or refused because of the cumulative dysfunction of these grafts. PMID- 11112006 TI - Experience with laparoscopic donor nephrectomy at a European transplant centre. AB - Renal grafts from live donors represent an important source for transplantation of end stage renal failure patients. Postoperative short- and long-term comfort is essential. Laparoscopic nephrectomy was performed in 22 cases. The left kidney was preferred for optimal length of the vessels. One procedure was converted to open surgery because of venous bleeding. Warm ischemia time varied between 4 and 7.5 min. Urine production started peroperatively in all cases, and the renal function was excellent. Shoulder pain 1-3 days postoperatively was observed in seven patients; the rest were comfortable on peroral non-opioid analgesia. The patients were discharged at postoperative days 3-9, and returned to work 2-4 weeks later as compared to 4-8 weeks after open nephrectomy at our centre. Laparoscopic donor nephrectomy in the hands of experienced laparoscopic and transplant surgeons is a safe operation with less discomfort to the living kidney donor. PMID- 11112007 TI - Early results of a non-heartbeating donor (NHBD) programme with machine perfusion. AB - Freeman Hospital, Newcastle upon Tyne restarted their non-heartbeating donor (NHBD) programme in September 1998 using machine perfusion, due to early poor results with conventional cold storage (45% graft survival, phase II). Since then, 15 NHBD kidneys have been transplanted. The retrieval protocol consisted of in situ perfusion with a double balloon triple lumen cannula in Maastricht category II male donors age range 13-59 years. Mean primary warm ischaemic time was 24.8 min (range 10-44). All kidneys were machine perfused through a locally developed perfusion system. The viability was assessed by serial measurements of total GST (maximum acceptable limit of 200 units/l) and intrarenal vascular resistance (IRVR) was recorded. Fifteen of the 22 kidneys (68.62%) were transplanted. Delayed graft function (DGF) was seen in ten recipients (66.6%), two kidneys had immediate function (IF), one organ was exported, two recipients died of unrelated causes and a further seven kidneys were discarded (two had high tGST, two were infected and three had poor flow characteristics). In phase III, a success rate of 91.7% was thus achieved, which was better than the phase II period (P = 0.027, Fisher 2-tail test). Machine perfusion has been successfully introduced in phase III to the Newcastle NHBD programme and facilitates viability assessment of NHBD kidneys. PMID- 11112008 TI - An algorithm for cadaver kidney allocation based on a multivariate analysis of factors impacting on cadaver kidney graft survival and function. AB - The large imbalance between cadaver kidney supply and demand makes the implementation of equitable and effective organ allocation systems an urgent need. This has triggered a revision of the criteria used so far for cadaver kidney allocation within the North Italy Transplant program, not least in the light of the many changes that have occurred recently with respect to broader criteria for admission of patients to the waiting list, donor selection, tissue typing methods, organ preservation and immunosuppressive protocols. We based the critical revision of our cadaver kidney allocation algorithm on univariate and multivariate analysis of a number of immunological, clinical, social and administrative factors that impacted on the transplant outcome in 2,917 patients transplanted in the 12 transplant centers operating within our organization from 1 January 1990 to 30 September 1997. This analysis indicated that younger donor age, absence of pretransplant transfusions, patient dialysis center and level of HLA match showed statistically significant positive associations with graft survival. Younger donor age and male donor gender showed a statistically significant association with excellent graft function at 4 years. The results of this analysis were used to develop a new computer-assisted version of our adult kidney allocation algorithm. It works in two steps (local pool first, then the entire waiting list) and four levels (0-1 HLA MM, PRA+; 2 HLA MM, PRA+; 0-1 MM, PRA-; 2-4 HLA MM, PRA-); within each level, selection takes into account waiting time and age difference from donor age. The evaluation of 731 transplants allocated in 19 months with the new algorithm, as against 698 transplants allocated in the preceding 19 months according to the previous algorithm, showed a significantly higher proportion of recipients who had been on the waiting list for more than 3 years (33.2% versus 22.6%). The use of the new algorithm was also associated with a significantly increased number of transplanted alloimmunized patients (18.8% versus 9.2% with the previous algorithm) and recipients with 0-1 HLA mismatches (22% versus 14.3%). Furthermore, the number of kidneys used locally has steadily increased. Differences in 6-month graft survival and percentage of patients with excellent function at 6 months were not statistically significant in recipients transplanted with the new versus the previous algorithm. Survivals were 93.7% versus 91.8%. Percentages of patients with excellent renal function were 69.9% and 71.8%, respectively. These preliminary data suggest that the new algorithm improves HLA match and reduces the number of patients on the waiting list for 3 or more years without determining significant modifications of 6-month graft survival and function. Moreover, it facilitates the achievement of a fair local balance between organs retrieved and transplanted, the compliance of operators with objective allocation rules and the documentation of the whole allocation process. PMID- 11112009 TI - Different diagnostic approaches to adult candidates for cadaveric kidney transplantation in Europe. AB - We investigated which diagnostic procedures are mandatory for all transplant candidates irrespective of their individual situation in European transplant centres, how homogeneously these are applied and what centre characteristics determine differences in the diagnostic approach. A questionnaire was sent to European renal transplant centres asking which of 45 listed diagnostic procedures are mandatory for every transplant candidate. The 154 participating centres require 15.6 +/- 5.6 (4-33) mandatory tests, with significantly less mandatory diagnostics in centres in the UK (8.5 +/- 3.9) and Scandinavia (9.8 +/- 2.3). Geographic location is the single significant factor in multifactorial analysis of possibly related factors. Detailed analysis revealed 16 tests that are required significantly less often in the north of Europe. There are significant differences in the evaluation of renal transplant candidates across Europe. In some parts of Europe transplant candidates are either investigated more discriminately or less comprehensively than in other regions. PMID- 11112010 TI - Efficiency of organ procurement and transplantation programs. AB - The number of donations per million population (pmp) per year and the number of transplants pmp/year enables one to compare donation or transplant programs made in different years in the same area or made the same year in different areas. These pmp indexes may be integrated with an evaluation system by which each organ is evaluated separately in terms of the efficiency of its procurement and transplant systems using the procurement index (percentage in terms of number of organs utilized/number of organs procurable from donors utilized in a single area during 1 year) and the transplant index (percentage in terms of number of transplants performed/number of organs harvested in a single area during 1 year). We have called them Caldes I (procurement) and Caldes II (transplant) indexes. The harvest index evaluates the efficiency of utilization of organs retrieved from suitable donors. It usually ranges between 80-90% for the kidney, 70-95% for the liver, 40-50% for the heart, and 5-15% for the lungs. The transplant index evaluates for each organ the transplant team capacity to use available organs which can be harvested locally or in different areas. It usually ranges between 60-120%. Index determination did not require information different from the standard data available. Both the harvest and transplant indexes could be used to compare the efficiency of donation and transplant programs and policies in the same area during different years or at the same time in different areas. They can be critical in evaluating: (a) marginal donor utilization, (b) marginal organ utilization, and (c) dishomogeneity of organ retrieval and organ transplantation in different regions belonging to the same area. They also enable to evaluate if organs considered not available in a single area are offered to other areas or are not retrieved at all from available donors. PMID- 11112011 TI - Nondestructive and real-time evaluation of liver viability in brain dead donor for liver transplantation using near-infrared spectroscopy. AB - A reliable and less-invasive method is currently desired to assess the hemodynamic and functional alteration associated with brain death in the organs of donor candidates. Near-infrared spectroscopy (NIRs) was applied to rat liver in brain-dead donors for assessing tissue oxygenation and intracellular energy metabolism as a means of monitoring the liver viability in the brain-dead donor. Brain-dead rats were divided into 4 according to doses of epinephrine and vasopressin administered. Arterial ketone bodies ratio (AKBR), hyaluronic acid (HA), and NIRs monitoring of a liver graft were performed in the brain-dead phase before the grafts were transplanted into syngeneic rats. NIRs monitoring of oxygenated hemoglobin (Hb) and cytochrome aa3 oxidase (Cytaa3) redox state reflected changes in the hepatic microcirculation and intracellular oxygenation. The administration of high-dose epinephrine proved to be contraindicated due to catecholamine-induced hypoxic stress, while combined administration of adrenaline and vasopressin at an optimal dose rate was beneficial for preservation of the liver viability. The data obtained by NIRs were significantly correlated with the 7-day survival of recipients after liver transplantation. Thus, we conclude that NIRs is a sensitive and nondestructive method for monitoring alterations in the viability of brain-dead liver and can predict liver graft outcome. PMID- 11112012 TI - Efficacy of HSP72 induction in rat liver by orally administered geranylgeranylacetone. AB - It is well known that heat-shock proteins (HSPs) have a cytoprotective function as "molecular chaperones" when cells are exposed to several stress conditions. Geranylgeranylacetone (GGA) is an antiulcer drug that was developed in Japan and it has recently been reported to induce HSP72 in rat gastric mucosa. In this experiment, we investigated the induction of HSP72 in rat liver in response to oral administration of GGA and assessed its ability to induce tolerance to warm ischemic injury by this approach. We prepared donor rats by orally administering GGA to them and compared HSP72 expression in graft liver, survival rates, and serum TNF-alpha concentrations after liver transplantation with the findings in controls. The survival rates were significantly increased when the livers were obtained from donor rats given GGA. Western blotting revealed expression of HSP72 in graft livers given GGA, and the serum TNF-alpha levels were significantly suppressed in the rats given GGA. Oral administration of GGA induced HSP72 in graft livers, and they were better able to tolerate warm ischemic injury. Oral administration of GGA appears to provide a promising new strategy for preventing ischemia-reperfusion injury. PMID- 11112013 TI - Hormonal changes in brain death and immune activation in the donor. AB - Kidneys obtained from brain dead donors show inferior graft survival compared to living donation. The effects of brain death itself are thought to be partly responsible for these results. We, therefore, examined levels of catecholamines, the vasoconstricting hormones AT II, ET-1 and renin activity, pituitary hormones, and their correlation to pro-inflammatory cytokines and cytokine receptors. In 17 brain dead patients and 19 preoperative neurosurgical patients, these parameters were measured by HPLC, RIA and ELISA. Brain death resulted in massive increases in serum catecholamines, AT II and ET-1, as well as PRA, whereas thyroid and adrenal hormone levels remained unchanged. We found a significant correlation with rises in IL-6 and soluble TNF and IL-2 receptors as markers for the activation of immunological cascades. We concluded that these effects could be directly and indirectly responsible for the impaired organ perfusion and function observed in brain death. PMID- 11112014 TI - Endotoxemia in organ donors: graft function following liver transplantation. AB - Translocation of endotoxin (LPS) to the portal-venous system is produced by multiple factors. In the case of normal liver function, LPS is rapidly cleared from the portal blood by Kupffer cells; in impaired liver function, LPS can reach the systemic circulation. The objective of this study was to investigate whether elevated donor endotoxin levels affect graft function in the recipient. LPS levels in donor plasma were measured in 14 consecutive liver transplantations. Grafts with donor LPS levels < or = 12 pg/ml had a function probability of 100% after 600 days (n = 10). LPS concentrations of > 12 pg/ml in donor plasma led to loss of function in 75% of the liver grafts (n = 4; P = 0.003; Wilcoxon). Elevated LPS values in donor plasma seem to impair the prognosis of the grafts and could predict poor graft function as early as at the time of brain death. PMID- 11112015 TI - The quality of a liver graft. AB - Because transplantation success is influenced by the quality of the graft, the objective of this study was to find parameters to evaluate transplant livers in the recipient centre. In 64 liver grafts, the venous effluates of a portal back table flush were investigated for various parameters. Amongst them, glutathione S transferase (GST), glutamate dehydrogenase (GLDH) and the leucocyte count were found superior in predicting graft survival. Using the combination of these parameters, 100-day graft survival of between 95% (all parameters positive) and 0% (all parameters negative) was predicted. We concluded that good liver grafts are characterized by a low width of injury (cytosolic component: GST), a low depth of injury (mitochondrial component: GLDH), as well as by a potential to induce tolerance (passenger leucocytes). Perfusate analysis seems to be a valuable tool to recognize problematic grafts in advance and to quantify the "graft factor" in considerations concerning quality control. PMID- 11112016 TI - Single-bolus high-dose ATG for prophylaxis of rejection in renal transplantation- a prospective, randomized study. AB - Currently, most centers use antithymocyte globulin (ATG) for induction or treatment of acute rejection. In the literature, postponement of introduction of cyclosporine or delay in acute rejection following ATG induction are well documented [1-4]. In contrast, data are very scant on the reduction of incidence of rejection or improvement of graft survival following ATG prophylaxis [5, 6]. The objective of this study was to compare the efficacy and safety of ATG high dose single-bolus therapy with that of a standard cyclosporine-based protocol in prophylaxis of acute rejection in renal transplantation in an adult population. Rabbit ATG (Fresenius, Oberursel) was administered intraoperatively (before revascularization) to 19 renal transplant recipients as a single intravenous injection in a dose of 9 mg/kg body weight (high dose, single bolus). Treatment results were compared with those of a control group comprising 19 recipients receiving the same cyclosporin-Neoral-based protocol as the study group. In all patients concomitant medication consisted of steroids and azathioprine. The incidence of acute rejection in the high-dose ATG bolus group was 26%, compared with 58% in controls (P < 0.05). In the ATG treated group no grafts were lost to acute rejection in both high- and low-risk recipients, versus compared with a loss of 37% of rejecting grafts in controls. Though the observed difference in 1 year graft survival between study and control groups (84.2% vs 73.6%) did not reach statistical significance, the same trend was also observed in patients (n = 9 and n = 12 respectively) who, at he time of this report, had completed a 2nd post-transplantation year. The bolus and control groups had a similar incidence of complications and comparable renal function. We conclude that a single-bolus high-ATG protocol is efficient and safe in prophylaxis of renal allograft rejection. PMID- 11112017 TI - Active TGF-beta1 expression in kidney transplantation: a comparative study of cyclosporin-A (CyA) and tacrolimus (FK506). AB - Chronic rejection is a major cause of graft dysfunction following kidney transplantation. This fibroproliferative disease may be promoted by overproduction of transforming growth factor beta (TGF-beta). Previous studies have suggested that cyclosporin-A (CyA) might increase production of this growth factor. The current study was designed to measure the expression of TGF-beta in renal transplant biopsies from patients immunosuppressed with either CyA or tacrolimus. Paraffin-embedded renal biopsies were sectioned, dewaxed and incubated with primary antibody against active TGF-beta1 antibody. After washing, the sections were treated with secondary antibody conjugated with fluorescein isothiocyanate (FITC). In each case the sections were assessed by semi quantitative scanning laser confocal microscopy. Biopsies from patients receiving CyA expressed significantly more active TGF-beta1 than biopsies from patients receiving tacrolimus (P < 0.0001, Mann-Whitney test). The increased level of active TGF-beta1 expression in renal biopsies of patients receiving CyA may indicate a mechanism of chronic rejection. PMID- 11112018 TI - Analysis of 100 pregnancy outcomes in women treated systemically with tacrolimus. AB - The aim of this paper is to provide a summary of clinical findings regarding the safety of tacrolimus in pregnancy. From 1992 to 1998 data were collected on 100 pregnancies from 84 mothers who received tacrolimus systemically; 83 cases of solid organ transplantation, and 1 case of Behcet's disease. Maternal mean age at conception was 28 years and pregnancy outcome was live birth in 68%, spontaneous abortion in 12%, induced abortion in 12%, stillbirth/perinatal death in 3%, ongoing pregnancy in 2%, and lost to follow up in 3%. Fifty-nine percent of the neonates were delivered prematurely (< 37 weeks of gestation). Birth weight was appropriate for the gestational age in 90% of the cases. Malformations occurred in 4 neonates: case 1, meningocele and urogenital defects; case 2, alcoholic embryopathy; case 3, ear defect, cleft palate and hypospadia; case 4, multicystic dysplastic kidney. There was no consistent pattern of malformations and 2 mothers subsequently delivered a healthy neonate while on tacrolimus therapy. Nearly 70% of pregnancies following systemic tacrolimus administration resulted in a favourable outcome without any significant effect on intrauterine growth. The incidence of malformations was similar to that reported with other immunosuppressants in transplant recipients. PMID- 11112019 TI - Pharmacokinetics and pharmacodynamics of mycophenolic acid in stable renal transplant recipients treated with low doses of mycophenolate mofetil. AB - Suboptimal doses of mycophenolate mofetil (MMF) are frequently employed in renal transplant (Tx) patients, with drug-related side effects or low weight. The aim of this study was to compare the mycophenolic acid (MPA) pharmacokinetic profile and its pharmacodynamic effect on patients receiving either standard (2 g) or low (1.5 g or 1 g) MMF doses, in order to evaluate the therapeutic efficacy of such low doses in inhibiting IMPDH activity. Twenty-seven stable renal Tx recipients aged 18-65 years, with a post-Tx follow-up of 38.5 +/- 44.8 months (6-166 months), receiving 1 g (n = 10), 0.75 g (n = 7) and 0.5 g (n = 10) MMF twice a day in association with cyclosporine and prednisone, were included. The control group was made up of untreated healthy volunteers (n = 5). Plasma concentrations of MPA were analyzed by reverse-phase HPLC. IMPDH activity was determined in lymphocytes by the measurement of 3H release from [2,8-(3)H] hypoxantine. The mean value of areas under the concentration-time curves (AUC(0-12)) of MPA throughout the 12-h dosing interval in patients treated with 2 g was higher than the corresponding data in patients receiving 1.5 g or 1 g bid, but no statistical differences were observed between the three groups. There was no correlation between MPA-AUC(0-12) values and MMF dose (expressed in g/day or g/kg per day). Predose MPA concentrations correlated only weakly with the respective MPA-AUC(0 12) values (r2 from 0.385 to 0.655), whereas an acceptable correlation was observed between MPA Cmax and MPA-AUC(0-12) (r2 from 0.626 to 0.759) in 2 g, 1.5 g, and 1 g MMF groups. An inverse relationship between MPA concentrations and IMPDH activity was observed. In general, the maximum MPA concentration was achieved from 1 h to 2 h after dosing, and the maximum inhibition of IMPDH was also from 1 h to 2 h after dosing. The evaluation of IMPDH activity demonstrated that there was a significant statistical difference between samples from 0 to 1 h (P = 0.008) and 0 to 2 h (P = 0.04). In conclusion, concentration-time profiles of renal transplant recipients administered 0.75 g and 0.5 g twice a day are slightly lower than those from the 2 g group, but nor significantly. On the other hand, inhibition of IMPDH activity was comparable in the three groups, indicating considerable interindividual pharmacodynamic variability. Pharmacodynamic monitoring of the degree of immunosuppression and its correlation with MPA plasma concentrations will be assessed further in future studies. PMID- 11112020 TI - Rational design of biologically active peptides: inhibition of T cell activation through interference with CD4 function. AB - In our laboratory we generated one synthetic cyclic peptide (Pep4) and tested it in human mitogen stimulation assays (MSA) and mixed lymphocytes reactions (MLR) generating dose-response curves showing a dose-dependent inhibition of MSA up to 80% and MLR up to 98%. MSA and MLR were repeated after pre incubation of the Pep4 with each separate responder cell subset and subsequent reconstitution: these experiments showed inhibition only when the peptide was present in culture. Pep4 showed species specificity since it was ineffective in inhibiting rat MLR. Combination effect analysis with Pep4 and cyclosporine showed a combination index > 1. This rationally designed peptide (Pep4) shows powerful inhibition of human T cell activation and, although the exact mechanism is still undefined, it seems to exert its major action on the T cell surface, interfering with co receptor interaction and disrupting the same activation signal pathway inhibited by cyclosporine A. PMID- 11112021 TI - FTY 720A mediates reduction of lymphocyte counts in human renal allograft recipients by an apoptosis-independent mechanism. AB - The novel immunosuppressive compound FTY 720A posseses a mode of action which is different from all other immunosuppressive drugs. The most prominent feature is a reversible decrease in peripheral lymphocyte counts observed in animal experiments. We investigated in the first human trial (phase 1) whether FTY 720A induces apoptosis of peripheral blood mononuclear cells (PBMC) in stable renal allograft recipients. Monitoring of lymphocyte counts revealed a significant and dose-dependent decrease within 6 h post-FTY 720A dose: placebo 5.1%; 0.25 mg 36.4%; 0.5 mg 40.8%; 0.75 mg 39.4%; 1 mg 45.8%; 2 mg 67.2%; 3.5 mg 64.9%. PBMC apoptosis rates did not change, as determined before intake of FTY 720A and 2 h, 6 h, 24 h and 96 h post-FTY 720A dose. We detected no significant difference in apoptosis rates between patients who received placebo or FTY 720A. However, in vitro experiments showed that high concentrations of FTY 720 A induced apoptosis in human PBMC. PMID- 11112022 TI - Efficacies of sirolimus (rapamycin) and cyclosporine in allograft vascular disease in non-human primates: trough levels of sirolimus correlate with inhibition of progression of arterial intimal thickening. AB - We investigated the efficacies of sirolimus (rapamycin) and cyclosporine for inhibition of graft vascular disease (GVD) in cynomolgus monkey recipients of aortic allografts. Increases in arterial intimal thickening in the midgraft (six consecutive cross-sections) after transplantation were quantified by serial intravascular ultrasound (IVUS) from day 21 to day 105. These data enabled correlations between changes in intimal indexes [II = (intimal area/vessel area) x 100] and trough levels of sirolimus and cyclosporine to be determined. Eighteen recipients received no immunosuppression for 6 weeks to allow alloimmune injury to occur. On day 45, monkeys were treated daily with sirolimus (n = 6) or cyclosporine (n = 6); six monkeys remained untreated. II increased significantly from day 63 to day 105 in untreated monkeys and monkeys treated with cyclosporine, whereas monkeys treated with sirolimus did not have a significant increase in II (P = 0.008, P = 0.006, P = NS; paired t-test). The change in II from days 63 to 105 was significantly greater in untreated monkeys compared to sirolimus-treated monkeys (P = 0.13; one-way ANOVA, P = 0.012 Tukey's post hoc test); other post hoc pairwise comparisons were not significant. Mean sirolimus and cyclosporine levels +/- SEM were 43 +/- 7 ng/ml and 562 +/- 20 ng/ml, respectively. Sirolimus trough levels, but not cyclosporine levels, correlated inversely with changes in II from day 42 to 105 (r2 = 0.73, P = 0.03). This non human primate study shows that inhibition of intimal thickening by sirolimus depends on trough levels and provides the rationale for clinical trials of sirolimus for the control of GVD in organ transplant recipients. PMID- 11112023 TI - Cyclosporine reduces basolateral, but not apical, nitric oxide secretion in medullary thick ascending limb cells. AB - Cyclosporine (CsA) reduces nitric oxide (NO) production in medullary thick ascending limb (mTAL) cells. We postulated that CsA affected NO secretion in a vectorial manner in polarized renal epithelial cells. The experiments were performed in a model of mTAL subcultured cells. The expression of iNOS in mTAL cells was confirmed by RT-PCR. The cells were grown on a non-permeable filter. Nitrite was measured by the modified Griess method. Transepithelial resistance was measured to ensure the integrity of the tight junction. CsA (100 ng/ml) reduced NO production by 22% in mTAL cells. The inhibitory effect was limited to the basolateral side (control: 165 +/- 11; plus CsA: 93 +/- 17 nM/10(6) cells, P < 0.001) without affecting apical NO secretion. The transepithelial resistance through the epithelial monolayer remained unchanged in CsA-treated cells. CsA reduced basolateral NO secretion without affecting apical secretion. The results suggest that CsA might affect intrarenal hemodynamics at the peritubular level. PMID- 11112024 TI - Fas ligand gene transfer combined with low dose cyclosporine A reduces acute lung allograft rejection. AB - The interaction between Fas and its ligand (FasL) induces apoptosis in the Fas expressing cell. We hypothesized that liposome-mediated FasL gene transduction to the lung allograft, in addition to low-dose immunosuppression, might reduce acute rejection. Orthotopic left lung allotransplantation was performed in male rats (Brown Norway to Fischer F344). FasL gene transfer was performed by use of the plasmid pBCMGSNeo carrying the gene coding for murine FasL and the cationic liposome GL#67:DOPE. Six hundred and sixty micrograms of DNA in 250 microl H2O and 0.5 micromol GL#67 in 250 microl H2O were diluted to 5 ml with saline solution. This emulsion (20 degrees C) was instilled retrogradely through the left pulmonary vein after flushing with LPD solution (20 ml, at 4 degrees C). Subsequently, the graft was stored at 10 degrees C for 3 h. A single dose of cyclosporine A (CsA; 2.5 mg/kg i.m.) was given to all groups 48 h after the transplantation. In group 1 (n = 6), FasL/GL#67 was instilled as described. In group 2 (n = 5), GL#67 was given without DNA. Group 3 (n = 5) animals received CsA only. Five days after transplantation, gas exchange was assessed after exclusion of the contralateral native lung (FiO2 = 1.0). Grafts were flushed with saline solution and fixed in formaldehyde for histological evaluation. No statistical difference in gas exchange (PaO2) between the two control groups 2 (6.4 +/- 0.4 kPa) and 3 (7.4 +/- 0.4 kPa) could be detected 5 days postoperatively (P = 0.9). In contrast, grafts transduced with FasL (group 1) had significantly better gas exchange on postoperative day 5 (PaO2: group 1 37.0 +/- 10.6 kPa vs group 2 6.4 +/- 0.41 kPa; P = 0.002). Two animals in group 1 revealed no or only minimal improvement in gas exchange. Histologically, all lung specimen of all groups showed signs of acute rejection (A2). Leukocyte infiltrates, rated by two independent observers, were less severe in all group 1 animals. Liposome mediated FasL gene transfer at the time of harvest in combination with low-dose CsA reduces acute rejection in four out of six animals in this model of rat lung allotransplantation. PMID- 11112025 TI - Adenovirus-mediated gene transfer of CTLA4Ig gene results in prolonged survival of heart allograft. AB - It has been demonstrated that the administration of CTLA4Ig protein can induce the suppression of allograft and xenograft rejection. The purpose of this study is to determine the effect of adenovirus-mediated gene transfer with CTLA4Ig gene on the transgene expression and suppression of alloimmune response in allogeneic cardiac transplantation of rats. Adenoviral vectors with beta galactosidase or human CTLA4Ig cDNA (Adex/LacZ, Adex/hCTLA4Ig) were constructed. These vectors were transduced to the liver of Lewis (LEW) rats by intravenous injection. Then LEW rats received heterotopic cardiac transplantation from Dark Agouti rats. Experiments were performed using both CTLA4Ig gene transduction and/or immunosuppressant FK506. The transgene expression after adenovirus-mediated transfer with CTLA4Ig cDNA lasted for several months, compared with several weeks after that in controls, which was proportional to the blood concentration of CTLA4Ig protein. By the administration of 1 x 10(9) PFU of Adex/hCTLA4Ig, the survival of cardiac grafts was significantly prolonged, compared with controls or the use of 1 x 10(8) PFU of Adex/hCTLA4Ig. In the rats with beating grafts over 100 days, the blood concentrations of CTLA4Ig were undetectable. The combination therapy using a low titer Adex/hCTLA4Ig and low-dose of FK506 was synergistically effective on this cardiac transplantation model. In conclusion, adenovirus mediated gene transfer with CTLA4Ig gene was efficient for the prolongation of both transgene expression and allograft survival. PMID- 11112026 TI - Mycophenolic acid trough levels after kidney transplantation in a cyclosporine free protocol. AB - Twenty-seven stable kidney transplant recipients treated with cyclosporine and prednisone were converted to mycophenolate mofetil (MMF) and prednisone 1 year after transplantation. After conversion the patients were treated with a standard daily dose of 1 g MMF b.i.d. and 10 mg prednisone for 4 months. Thereafter, two MMF dose reductions were performed with a 4-month interval. Mycophenolic acid (MPA) trough levels were measured at regular intervals. A relation was found between MPA trough levels and MMF dose. The median MPA trough level for patients treated with 1 g MMF b.i.d. was 4.3 microg/ml (0.95-15.5) and 3.0 microg/ml (0.73 7.8) for patients treated with 750 mg b.i.d. (P = 0.0002). The MPA trough levels further decreased from 3.0 to 2.3 microg/ml (0.6-6.63) in patients treated with 500 mg MMF b.i.d. (P = 0.01). Dose reduction of MMF from 1 g to 750 mg b.i.d. could be performed without acute rejections. A further dose reduction to 500 mg b.i.d. elicited 3 rejections. Patients experiencing an acute rejection had a median MPA trough level of 2.3 microg/ml (1.26-3.38) compared to 3.8 microg/ml (1.48-6.52) in patients without an acute rejection (P = 0.25). We conclude that there is a significant relation between MPA trough levels and MMF dose. MPA trough levels were not predictive of rejection in the present study. PMID- 11112027 TI - Tacrolimus is highly effective in both dual and triple therapy regimens following renal transplantation. Spanish and Italian Tacrolimus Study Group. AB - This open, multicenter, randomized, parallel-group study evaluated the efficacy and safety of tacrolimus-based dual and triple therapy regimens. For this 3-month study (with 12-month follow up), 491 adult renal transplant patients were randomized and received either dual therapy (tacrolimus/corticosteroids; 246 patients) or triple therapy (tacrolimus/corticosteroids/azathioprine; 245 patients). Patient survival rates at months 3 and 12 were 99.2 (dual) vs 99.6% (triple) and 97.8 vs 98.7%, respectively. Graft survival rates at months 3 and 12 were 94.1 (dual) vs 95.4% (triple) and 92.8 vs 93.3%, respectively. After 3 months, the incidences of treated acute rejection were 28.8 (dual) and 29.7% (triple); and 7.6 (dual) and 5.4% (triple) for corticosteroid-resistant acute rejections. Between months 4 and 12, three new first rejections were reported, (dual: 2, triple: 1). For leukopenia (1.3 vs 11.7%; P < 0.001) and anemia (14.8 vs 23.0%, P = 0.026), significantly higher incidences were reported in the triple therapy group. The incidence of de novo insulin-dependent diabetes was 5.6 (dual) and 4.0% (triple) at month 3. In terms of efficacy, no difference between the treatment groups was observed. PMID- 11112028 TI - Rejection and tacrolimus conversion therapy in paediatric liver transplantation. AB - Rejection and efficacy of rescue therapy with tacrolimus were evaluated in 50 children who underwent primary, ABO-compatible, liver transplantation. Six patients who died within the first week and one child who underwent retransplantation from an ABO-incompatible donor were excluded from the study. No patient or graft were lost due to rejection. We observed 48 episodes of rejection in 33 patients. Fourteen patients required conversion to tacrolimus for steroid resistant rejection with resolution of rejection. One of these children developed PTLD. Other indications for conversion were neurotoxicity and hirsutism. One patient developed blindness of unknown origin after the conversion. Other side effects of tacrolimus were minor and resolved by lowering the dose. Five patients developed rejection after conversion; all achieved resolution with either steroid therapy or increase of tacrolimus dose. In conclusion, our study confirms that tacrolimus is an effective rescue therapy for paediatric liver transplantation. PMID- 11112029 TI - Conversion from cyclosporine A to tacrolimus after kidney transplantation due to hyperlipidemia. AB - As more than 90% of renal grafts retain their function 1 year after renal transplantation, side effects of immunosuppressive therapy gain more and more importance. In a randomised prospective study, we investigated the effects of conversion from cyclosporine A to tacrolimus due to hyperlipidemia in recipients of renal allografts. Fifty-seven patients with stable graft function treated with cyclosporine were randomly assigned to conversion to tacrolimus or continuation of their current therapy and followed for 1 year. Twenty-seven patients were switched and 30 patients remained on cyclosporine A. Cholesterol levels decreased significantly in the tacrolimus group as compared to controls in the intent to treat analysis. When only those patients were evaluated who received cyclosporine or tacrolimus during the whole study, statistical significance was even more pronounced. Triglyceride levels decreased in the tacrolimus group, whereas they increased in controls. Creatinine levels remained stable and no acute rejection was observed. A switch to tacrolimus is a safe alternative in cases of hyperlipidemia after renal transplantation. PMID- 11112030 TI - HHV-6 reactivation is often associated with CMV infection in liver transplant patients. AB - Human herpesvirus 6 (HHV-6) infection has been recently reported in liver transplant patients. HHV-6 is closely related to cytomegalo-virus (CMV), and some interaction between the viruses has been suggested. In this study, the post transplant HHV-6 antigenemia was investigated in relation to symptomatic CMV infections in adult liver transplant patients. CMV infections were diagnosed by the pp65 antigenemia test and by viral cultures. HHV-6 infections were demonstrated by the HHV-6 antigenemia test and by serology. Significant symptomatic CMV infection was diagnosed in 42 of 75 patients during the first 6 months after transplantation. All CMV infections were successfully treated with ganciclovir. Concurrent HHV-6 antigenemia was detected in 21 (50%) of 42 patients with CMV infection. All HHV-6 infections were reactivations. HHV-6 also responded to the antiviral treatment, but with less clear effect. In conclusion, HHV-6 reactivation is often associated with CMV infection in liver transplant patients. The results support the suggestion that CMV and HHV-6 may have interactions. PMID- 11112031 TI - Hyperimmunoglobulin prophylaxis, monitoring and preemptive ganciclovir treatment eliminate the risk of CMV infection to improve patient and renal allograft survival. AB - This study was designed to investigate whether the introduction of ganciclovir to clinical use for anti-CMV treatment changes the risk of CMV infection in renal transplant patients. A total of 1545 cases who had received cadaveric renal transplants were divided into two groups: group 1 (n = 721) was made up of patients who received their transplants within 6 years before the introduction (1991) of ganciclovir and group 2 (n = 824), of individuals transplanted thereafter. Patient and graft survival of CMV D+/R- patients was uni- and multivariately compared with non-CMV D+/R- patients. In CMV D+/R- patients in group 1, survival was significantly lower, and their relative risk for graft loss was 1.32-fold (P = 0.0483) that of non-CMV D+/R- patients. In group 2 patient and graft survival was identical regardless of whether the patients were at risk for CMV infection or not. The risk of CMV infection can be eliminated by hyperimmunoglobulin prophylaxis, CMV monitoring and preemptive ganciclovir treatment in renal transplant patients. PMID- 11112033 TI - Expression of the cytomegalovirus genome in kidney allografts during active and latent infection. AB - Cytomegalovirus (CMV) infection is suggested to be a risk factor for chronic rejection. Here we investigated whether CMV can persist in renal allografts, and in which structures the viral genome is found during an acute infection and a latent period after an active infection. CMV infection was diagnosed in 72/157 patients by CMV antigenemia tests and by viral cultures. CMV antigens were demonstrated in 38 available biopsies by immunohistochemistry, and CMV genome by DNA hybridization in situ. Standard histology was also performed. CMV antigens were detected in 7/15 biopsies obtained during acute infection, in three with acute rejection, and chronic changes in the other biopsies. CMV genome was located in inflammatory cells, in tubuli and in the capillary endothelium. During a latent period without a positive finding in blood or urine, CMV antigens were still found in 6/31 biopsies. CMV DNA was found in inflammatory cells, tubular and glomerular structures and in the endothelium of the arterioles. During the latent period with persistent CMV in the graft, in most cases (10/12) mild to moderate chronic changes were recorded. PMID- 11112032 TI - Antiviral combination therapy for lamivudine-resistant hepatitis B reinfection after liver transplantation. AB - Development of resistance is a major issue in antiviral treatment of hepatitis B reinfection after liver transplantation. Antiviral combination therapy is discussed for therapy or prevention of this breakthrough of viral replication. Eight patients were enrolled into this retrospective analysis after liver transplantation for chronic hepatitis B infection. All had reinfection of the graft and breakthrough of HBV during consecutive famciclovir and lamivudine monotherapy. Subsequently a combination therapy with lamivudine and interferon alpha 2a (group I, n = 4) or lamivudine and famciclovir (group II, n = 4) was initiated. Combination therapy was started 61 months (group I) and 25 months (group II) after liver transplantation. It markedly reduced the viral replication rate in all patients despite lamivudine resistance. In group I three of four patients and in group II two of four patients became HBV-DNA negative. Two long term responders were observed in group I, and none in group II. No patient became HBsAg negative or lost HbeAg. Pretreatment elevated ALT and AST levels were significantly reduced. No severe complications, and especially no rejection episodes, occurred. Lamivudine in combination with other antiviral agents, especially interferon-alpha, might be a therapeutic option for hepatitis B reinfection after liver transplantation. Suppression of virus replication to the point of undetectable values is possible even in patients with lamivudine resistant virus mutations. PMID- 11112034 TI - Diagnosis and monitoring of acute cytomegalovirus infection in peripheral blood of transplant recipients by nested reverse transcriptase polymerase chain reaction (RT-PCR). AB - The aim of this novel diagnostic approach is to monitor cytomegalovirus (CMV) infection in immunocompromised transplant recipients using early, sensitive, and specific predictors before and during antiviral therapy. The peripheral blood cells of 20 patients after transplantation (9 liver, 7 kidney, 4 simultaneous kidney-pancreas) were studied for an early diagnosis of acute infection. The mRNA and DNA of human CMV immediate-early antigen (IEA) were detected by nested polymerase chain reaction (PCR) assay. Results of nested PCR were compared with the immunological detection of antigen pp65 and serological diagnosis of CMV infection. All data were correlated with clinical symptoms like leukopenia, thrombopenia, pneumonia, and allograft-rejection reaction. Of 20 transplant recipients, 12 were infected by CMV, and 9 suffered from a CMV-related disease. CMV mRNA were detected simultaneously with antigen pp65 and CMV DNA in all patients with symptomatic infection. Additionally, CMV mRNA was found over a longer period after ganciclovir treatment of infected recipients. Nested reverse transcriptase (RT)-PCR for CMV-IEA mRNA allows a sensitive and specific diagnosis of an acute CMV infection. CMV mRNA was found to be a good marker of acute viremia and could be a useful tool for CMV monitoring over the whole period of disease management, even during antiviral therapy. PMID- 11112035 TI - The impact of cytomegalovirus serology for 7-year graft survival in cadaveric kidney transplantation--the Newcastle experience. AB - To analyse the contribution of cytomegalovirus (CMV) serology to long-term graft survival in cadaveric kidney transplantation, 404 transplants from a single centre were divided into four subgroups with respect to the combination of donor and recipient CMV antibody status. Graft survival was estimated according to Kaplan-Meier for 1, 3, 5 and 7 years post-transplantation. The single-centre results confirm a negative impact of CMV-positive donor organs for initial graft survival in CMV-negative recipients within the first 3 years after transplantation. However, when 5- and 7-year long-term graft survival was studied, Donor +/Recipient - pairs showed a favourable long-term result, whilst D +/R - pairs had surprisingly a poorer outcome. Therefore, the concept of avoiding transplantation in the D +/R + CMV serology group should be ignored whereas attempts could be made to improve the poor long-term outcome of D +/R + pairs or to reduce its size by organ allocation. PMID- 11112036 TI - Cholestatic syndromes in renal transplant recipients with HCV infection. AB - We present two distinct types of cholestatic syndrome identified in eight renal transplant (RTx) patients with HCV infection. Four patients developed fibrosing cholestatic hepatitis (FCH) and four, vanishing bile duct syndrome (VBDS). All patients with FCH were anti-HCV (-) at the time of Tx and developed a cholestatic profile 1-4 months post-Tx, with high HCV-RNA levels. Immunosuppressive therapy was drastically reduced. Two patients died of sepsis and liver failure 16 and 18 months post-Tx, and the other two showed marked improvement and seroconverted to anti-HCV. Regarding the patients with VBDS, three were anti-HCV (-) and one was anti-HCV (+)/HBsAg (+) at the time of RTx. Two patients became anti-HCV (+) 1 year, and one patient, 3 years post-Tx. Two patients developed progressive VBDS and died of liver failure 2 and 3 years after onset, and two showed marked improvement after withdrawal of immunosuppression. In two of the patients, the progression of the disease coincided with elevation in serum HCV RNA levels. We concluded that a progressive cholestatic syndrome acquiring features of FCH or VBDS may develop in HCV-infected RTx patients. The association with high viral load implicated the virus in the pathogenesis. Drastic reduction of immunosuppression may favourably affect the outcome. PMID- 11112037 TI - Are heart transplant recipients more likely to develop skin cancer than kidney transplant recipients? AB - Non-melanoma skin cancer is frequent in organ transplant recipients. The risk of posttransplant cutaneous squamous cell carcinoma in Norwegian heart transplant recipients (n = 148) and kidney transplant recipients (n = 1020) on triple immunosuppressive therapy with cyclosporine, azathioprine, and prednisolone, transplanted between 1983 and 1992, were studied. After adjustment for age at transplantation in multivariable Cox models, heart transplant recipients had a significantly 2.8-times higher risk of developing squamous cell carcinoma relative to kidney transplant recipients. The risk relative to the general population (standardized incidence ratio) was higher in heart transplant recipients than in kidney transplant recipients. The results indicate that heart transplant recipients are more likely to be diagnosed with skin cancer than kidney transplant recipients, probably due to the higher doses of cyclosporine and azathioprine after heart transplantation used at our center in the study period. PMID- 11112038 TI - Incidence and clinical characteristics of posttransplant lymphoproliferative disorders: report from a single center. AB - In the period 1973-1998, among 2139 allograft recipients treated with standard immunosuppression, posttransplant lymphoproliferative disorders (PTLD) developed in 19 patients (0.9%): one plasmacytic hyperplasia, two polymorphic PTLD, one myeloma, and 15 lymphomas. PTLD developed 1 year after transplantation (tx) in 14 patients. Five patients were diagnosed at autopsy, 2 were lost to follow up, 3 died before therapy could be instituted, and 1 patient has just started chemotherapy. Of the 8 evaluable patients, 2 received acyclovir and are alive in complete remission (CR) and 6 received chemotherapy +/- surgery. Of these 6, 4 died of lymphoma and/or infection, 1 died of unrelated causes in CR, and 1 is alive in CR. PTLD is a severe complication of tx, usually running an aggressive course which may preclude prompt diagnosis and treatment. Nevertheless, therapy is feasible and must be tailored on the histologic subtype. Seventy-four percent of patients were diagnosed with late-onset PTLD stressing the need for long-term follow up. PMID- 11112039 TI - Posttransplant lymphoproliferative disorders in renal allograft recipients: report of 53 cases of a French multicenter study. PTLD French Working Group. AB - New immunosuppressive therapies are currently being developed in renal transplantation and their relative risk in terms of posttransplant lymphoproliferative disorders (PTLD) must be carefully evaluated. For this purpose, a French registry of PTLD occurring after renal transplantation was set up. Among 10,000 patients presently followed up in 30 French renal transplantation centers, we prospectively identified 53 new PTLD (0.5%) since January 1998. Patients (34 male, 19 female) ranged from 3 to 72 years (mean age: 46 years), and the median time between grafting and diagnosis of PTLD was 63 months (2 months to 14 years). Ninety percent of recipients were Epstein-Barr virus (EBV) positive before transplantation. Most patients received a quadruple sequential therapy with polyclonal anti-lymphocyte globulin. Sites involved in PTLD were isolated lymph nodes in 13 cases, stomach or bowel in 10 cases, allograft in 14 cases, central nervous system in 6 cases, oropharynx in 4 cases, and skin or mucosa in 4 cases. Only three PTLD expressed markers of T lineage. Out of 40 studied tumors, 31 (78%) were EBV positive. Tumors were classified as polymorph in 26 cases and monomorph in 23 cases. Genotype studies in 18 PTLD showed a monoclonal pattern in 13 cases. In most patients, treatment consisted of reduction of immune suppression, 21 patients were given additional anti-viral therapy, 13 patients had anti-CD20, 23 patients underwent chemotherapy, and 4 patients were given cerebral radiotherapy. Five patients underwent transplantectomy. Sixteen patients (30%) died within the 1st year and 7 patients returned to dialysis (13%). The outcome of patients with PTLD remains poor, and the optimal approach to therapy is largely unknown. This ongoing registry is not only a national observatory but also a task force designed to improve the treatment strategy of PTLD. PMID- 11112040 TI - Cancer incidence in a kidney-transplanted population. AB - A study on cancer incidence after kidney transplantation was performed using data of national transplant and cancer registries. Since 1964 up to 30 June 1997, 3440 kidney transplantations were performed on 2890 patients. From 1967 to 1997, 230 posttransplantation malignancies were found in 20,817 patient-years of follow up. The standardised incidence ratio (SIR) was 3.33 compared to the general population. The SIR was highest in skin cancer (39.2). The SIRs were high in cancers of the lip (23.0), thyroid (8.08), kidney (8.0), lower urinary tract (3.2), non-Hodgkin lymphoma (4.8), ovary (3.9) and colon (3.9). Skin cancer and lymphomas had much higher SIRs in men than in women whereas lower urinary tract cancer had a higher SIR in women. During the first 10 follow up years, life-table analysis indicates a higher cancer risk in cyclosporine-treated patients, but this may be biased by their shorter follow up as the overall SIR was equal in both groups. This population study shows the increased incidence of cancer in the transplant population and points out the importance of cancer surveillance in the years following kidney transplantation. PMID- 11112041 TI - Circulating Epstein-Barr virus DNA to monitor lymphoproliferative disease following pediatric liver transplantation. AB - Epstein-Barr virus (EBV) infection can induce uncontrolled lymphocyte B proliferation in immunosuppressed transplant patients. Monitoring circulating EBV infected lymphocytes can help in identifying patients at risk of posttransplant lymphoproliferative disease (PTLD). Circulating EBV genome levels were determined in 54 liver transplant pediatric recipients. Ten patients had more than 500 EBV genome/10(5) peripheral blood lymphocytes (PBL) and exhibited clinical manifestations of EBV infection; three developed PTLD. To treat EBV infection, the level of immunosuppression was reduced and acute rejection developed in 4 patients. Three were treated with steroid and one had to be switched from cyclosporine to tacrolimus. Treatment of acute rejection was associated with increases in circulating EBV genome. None of the patients with less than 500 EBV genome/10(5) PBL developed PTLD or EBV infection. Monitoring of EBV DNA is useful in the management of EBV infection and PTLD following pediatric liver transplantation. EBV infection should be treated in ways which do not expose patients to the risk of rejection. PMID- 11112042 TI - Lymphoproliferative disorders in heart transplant recipients: role of hepatitis C virus (HCV) and Epstein-Barr virus (EBV) infection. AB - Post-transplant lymphoproliferative disorders (PTLD) are a well known complication after orthotopic heart transplantation (OHT). Although Epstein-Barr virus (EBV) infection has long been implicated in the pathogenesis of such disorders, other factors may play a part. Because of its lymphotropic properties, hepatitis C virus (HCV) may induce clonal expansion of B-lymphocytes and lead to PTLD. The aim of this study was to evaluate the potential association between HCV and EBV infection and PTLD in OHT patients. The retrospective study considered 404 adult patients screened for HCV. EBV serology, histology, and molecular analysis on tissue biopsies were performed in the PTLD patients (10/404, 2.5%). HCV positivity was found in 36/404 (8.9%) patients. The EBV genome was expressed on all neoplastic tissue samples analyzed. A higher proportion of HCV-positive patients developed PTLD than the HCV-negative cases (8% vs 2%, P = 0.017). EBV has a demonstrated role in the onset of PTLD, but HCV infection probably has to be considered as well. PMID- 11112043 TI - Outcome of liver resection and transplantation for fibrolamellar hepatocellular carcinoma. AB - Fibrolamellar hepatocellular carcinoma (FL HCC) is an uncommon variant of hepatocellular carcinoma occurring usually in non-cirrhotic livers. Hepatic resection or transplantation offers the only chance of cure. We reviewed our experience of surgery for FL HCC from 1985-1998. Twenty patients with FL HCC (13 females and 7 males) median age 27 years (range 12-69) were treated either by hepatic resection [n = 11; extended right hepatectomy (5), extended left hepatectomy (1), right hemihepatectomy (2), left hemihepatectomy (2), left lateral segmentectomy (1)] or, if the disease was non-resectable, by transplantation (n = 9). The median follow up was 25 months (1-63). The prognostic factors analysed included size [less than 5 cm (3 patients), more than 5 cm (17 patients)], number [solitary (16 patients), multiple (4 patients)], capsular invasion (6 patients), vascular invasion (11 patients) and lymph node invasion (6 patients). The overall survival at 1, 3 and 5 years was 89.5, 75 and 50%, respectively. The liver resection survival was better than liver transplantation survival at 3 years 100 vs 76%, respectively (P < 0.025). Although all prognostic factors analysed did not show a significant difference, there is tendency that tumour stage was the most significant for prognosis. Most of the patients in this study are young and presented without specific symptoms, with normal liver function range and had no tumour marker to help in diagnosis. As a result most of our patients were diagnosed late. However the outcome of surgical intervention was favourable. PMID- 11112044 TI - HHV8 in renal transplant recipients. AB - Human herpevirus 8 (HHV8) DNA sequences have been found in lesions from patients with Kaposi's sarcoma (KS) in several forms including immunosuppressed transplant patients. We wanted to study the transmission of HHV8 in kidney transplant recipients and to assess the risk of development of KS related to the viral infection in this group of patients. We tested sera of 120 renal transplant recipients with serological assay for antibodies to HHV8 antigens before transplantation and then we tested sera of 66 patients of the same group after transplantation. Antibodies were detectable in 27.5% of the patients before transplantation. In the seropositive population 15.1% developed KS and in the negative group 1.1%. Analysing 66 posttransplant sera we noticed that 24% of the seronegative patients became positive after transplantation. Our data suggest that being positive for HHV8 before transplantation could be an important risk factor for the development of KS. PMID- 11112045 TI - Cyclosporin-induced endothelial dysfunction and hypertension: are nitric oxide system abnormality and oxidative stress involved? AB - Hypertension is a major side effect of cyclosporin (CsA). While the mechanism(s) responsible are unclear, CsA-induced endothelial dysfunction and CsA-induced hypertension have been attributed to the CsA effect on the endothelial-derived factors controlling vasomotor tone. Endothelial nitric oxide (NO) is crucial in the maintenance of a state of basal vasodilation, and recent studies have suggested an NO-mediated counterregulatory mechanism protective from CsA-induced vasoconstriction. Our study evaluates endothelial nitric oxide synthase (ecNOS) gene status (PCR analysis) and plasma levels of NO metabolites (ELISA) in kidney and heart transplant patients under chronic CsA treatment with CsA-induced hypertension. Since CsA increases superoxide production, which metabolises NO, plasma hydroperoxides from cholesterol esters and from triglycerides and peroxynitrite were also evaluated (HPLC) as an index of the presence of superoxides and of "oxidative stress". Quantification of monocyte ecNOS mRNA and NO metabolites plasma levels from patients and controls (C) demonstrated NO system upregulation in patients notwithstanding the hypertension. The mean ecNOS to beta-actin ratio was 1.80 +/- 0.85 in patients vs 0.40 +/- 0.09 in C (P < 0.04). NO metabolites were 34.03 +/- 14.32 microM in patients vs 11.53 +/- 5.64 microM in C (P < 0.001). Hydroperoxides from cholesterol esters and from triglycerides were also increased in patients, 3.4 +/- 1.4 vs 1.3 +/- 0.6 integrated area units (i. a. u.), P < 0.007 and 10.6 +/- 6.4 vs 1.3 +/- 0.8 i. a. u., P < 0.008, respectively, as well as the peroxynitrite plasma level, 0.32 +/- 0.11 microM/l vs undetectable in C. This study confirms a CsA-induced NO system upregulation in transplanted patients. However, the NO-mediated counterregulatory system to CsA-induced vasoconstriction, present in normals, could be canceled in patients by CsA-induced superoxide (O2-) and free radical production which, by increasing NO metabolism, could contribute to CsA-induced vasoconstriction and hypertension and predispose to atherosclerosis. PMID- 11112046 TI - Risk factors for cardiovascular disease in renal transplant recipients: new insights. AB - Long-term survival of renal transplant recipients appears to be influenced by the occurrence of thromboembolic complications and cardiovascular disease. In order to investigate the prevalence of new hemostasis-related risk factors for venous and arterial thrombosis, we investigated 63 renal transplant recipients and 66 age- and sex-matched control subjects. We assayed antiphospholipid antibodies [lupus anticoagulant (LA) and anticardiolipin antibodies (aCL)], lipoprotein (a) [Lp(a)], plasminogen activator inhibitor-1 (PAI-1), and total homocysteine (tHcy) levels. We found a significantly higher prevalence of positivity for LA (P < 0.001); no difference was detected in the prevalence of aCL between patients and controls. PAI-1 levels were significantly higher in renal transplant recipients than in controls [12.3 IU/ml (2-45.5) vs 7.9 IU/ml (4-18.0); P < 0.0001] with an odd ratio (OR) of 11.8 (4.9-28.5) in univariate analysis and of 5.8 (2.1-15.4) in multivariate analysis. Lp(a) levels were higher in patients then in controls [159 mg/l (1-992) vs 100.5 mg/l (10-412); P < 0.005] with an OR of 5.9 (1.9-18.4) in univariate analysis and of 3.5 (0.9-13.4) in multivariate analysis. Fasting levels of tHcy were significantly higher in renal transplant recipients [7.0 micromol/l (4.0-68) vs 8.1 micromol/l (2.0-24.0); P < 0.00001] with an OR of 40.4 (14.7-111) in univariate analysis and of 33.1 (11.1-115.5) in multivariate analysis. After methionine loading test, we documented levels of tHcy above the 90th percentile of controls in 60/63 patients (95%). Finally, we found a significant correlation between tHcy and PAI-1 plasma levels (r = 0.76; P < 0.000001). Our results show a high prevalence of hemostasis-related risk factors for arterial and venous thrombosis in renal transplant recipients, suggesting the need for the investigation of these patients for the presence of these risk factors in order to improve their long-term survival and to tailor therapy. PMID- 11112047 TI - Efficacy and safety of Palmaz stent insertion in the treatment of renal artery stenosis in kidney transplantation. AB - The aim of this study was to verify the safety and long-term efficacy of Palmaz stent insertion in the treatment of transplant renal artery stenosis (TRAS) in kidney transplantation. Nine of our transplanted patients were submitted to Palmaz stent insertion because of recurrence of renal artery stenosis after previous percutaneous transluminal angioplasty or because of severe ostial stenosis. The post-stenting results were excellent in all patients, with a follow up period ranging from 1 to 3 years. The mean blood pressure (one-third systolic pressure plus two-thirds diastolic pressure) fell from 118.11 +/- 7.44 to 103.21 +/- 9.25 mmHg; P < 0.001. Renal artery peak blood flow velocity as determined by Doppler sonography fell from 352 +/- 73.24 cm/s to 169.8 +/- 23.35 cm/s; P < 0.001. The serum creatinine 1-year after stenting was 1.3 +/- 0.3 mg/dl with a slight reduction with respect to the pre-stenting values (1.5 +/- 0.3 mg/dl; NS). As no complication occurred, we conclude that insertion of the Palmaz stent is a safe and effective way to treat recurrence of artery stenosis or ostial stenosis in renal transplanted patients. PMID- 11112048 TI - Factors influencing vertebral bone density after renal transplantation. AB - To improve our understanding of the mechanisms underlying osteoporosis following renal transplantation, we compared bone mineral density (BMD) in 158 transplant recipients and in 293 patients undergoing maintenance hemodialysis with age- and sex-matched normal controls. Observations in graft recipients were made up to several years following transplantation. Dual-energy X-ray absorptiometry was used to measure BMD. Correlations with clinical variables including serum concentration of parathyroid hormone (PTH) and steroid therapy were evaluated. Lumbar BMD was lower in transplant patients than in dialysis patients at all ages, and continued to decrease with increasing interval posttransplant until the second year after transplantation. Persistent hyperparathyroidism and daily prednisolone dosage were both associated with decreased BMD. Age and creatinine clearance were independent long-term predictors of BMD by multiple regression analysis. Treatment of renal graft recipients with calcium and vitamin D supplements or calcitonin may be indicated in the early months after transplantation. PMID- 11112049 TI - Posttransplant donor-specific antibody characterization and kidney graft survival. AB - This study was designed to investigate the clinical relevance of donor-specific antibodies (DS-Abs) and their influence on graft survival. Among 106 patients who underwent cadaveric kidney donor transplantation and were monitored by flow cytometry crossmatch (FCXM) during the 1st posttransplantation year, 25 (23.6%) resulted positive for DS-Ab production. During a 2-year follow up only 12 of the 81 FCXM-negative patients (14.8%) suffered rejection vs 17 of 25 FCXM-positive patients (68%; P = 0.00001). Correlating graft loss to DS-Ab production, 9 FCXM positive patients lost the graft vs only 1 among the FCXM-negative patients. A worse graft function was evidenced in FCXM-positive subjects who had also suffered rejection episodes than in those which had acute rejection but did not produce DS-Abs. A high incidence of HLA-AB mismatches was found in FCXM-positive subjects which produced anti-class I antibodies. FCXM appears useful in estimating posttransplant alloimmune response. Moreover our findings confirm the harmful effects of anti-class I DS-Abs on long-term graft survival. PMID- 11112050 TI - HLA class I residue mismatch and renal graft outcome. AB - Donor-recipient HLA matching was retrospectively evaluated in 111 cadaveric renal transplants using Takemoto's ten-residue model in which HLA class I antigens are clustered by crossreactive group (CREGs) on the basis of amino acid sequence homology and the sharing of a particular public epitope. The grade and type of HLA residue mismatching were correlated to posttransplant, class I donor-specific antibody production (monitored by flow cytometry crossmatch), rejection occurrence and clinical outcome during the 1st year posttransplant. In 52 patients with 0 mismatchings (MMs) we observed a low incidence of rejection (11.1%) and antibody production (11.1%) for 0 CREG MM grade, while 1 MM was enough to increase immune response against graft (rejection 35%; antibodies 30%). Moreover, a significant correlation was observed between Q144, E163, Q62 and L82/R82 epitopes and the incidence of acute rejection and antibody production ("immunogenic" residues) in patients grouped for a single residue mismatch. PMID- 11112051 TI - Influence of perioperative sHLA I concentrations on the histological development of the liver graft. AB - Soluble HLA I (sHLA I) in human serum are ascribed an immunoregulatory role in the context of organ transplantation. Based on histological findings, the objective of the current study was to evaluate the protective influence of sHLA I in liver transplantation from the time point of reperfusion. The sHLA I concentrations in serum samples derived from the liver vein immediately after reperfusion (flush catheter) of 38 patients with liver transplantations were determined by ELISA. The postoperative histological findings of the transplant biopsies were categorized according to rejection, endothelialitis, cholestasis, and necrosis, as well as fatty degeneration. An evaluation according to Kaplan Meier showed a lower incidence for all of these factors in liver grafts with high sHLA concentrations (P < 0.05). We conclude that low sHLA I concentrations during reperfusion correlate with later complications, thus indicating that sHLA I may have protective potential in liver transplantation. PMID- 11112052 TI - Soluble donor MHC class I gene transfer to thymus promotes allograft survival in a high-responder heart transplant model. AB - Thymic selection of self and non-self-reactive lymphocytes is a process that may be targeted to induce donor-specific immunologic unresponsiveness in organ transplantation. In the present study, gene transfer was used to preexpose the recipient thymus to soluble donor-specific MHC class I molecules prior to heart transplantation in the high-responder ACI (RT1a) to Lewis (RT1l) rat strain combination. Specifically, cultured Lewis hepatocytes were transfected with DNA encoding a secreted form of the donor allo-MHC class I antigen, RT1.Aa. Seven days prior to ACI heart transplantation, genetically altered recipient-strain hepatocytes were injected into the thymus of Lewis recipients which also received a dose of antilymphocyte serum (ALS). Results showed that treatment with both ALS and soluble donor MHC-expressing hepatocytes prolonged transplant survival time by twofold, compared to injection of control hepatocytes and ALS. Therefore, intrathymic gene therapy delivery of soluble donor MHC molecules may be useful for promoting allograft survival in heart transplantation. PMID- 11112053 TI - Modulation of ICAM-1 tissue expression in patients with liver transplantation (LT) and acute rejection (AR) after glucocorticoid treatment. AB - Acute rejection (AR) is a frequent complication following liver transplantation (LT). ICAM-1 may be involved in its pathogenesis. High doses of glucocorticoids are the standard treatment in these patients. The aim of this study was to describe corticoid effects on ICAM-1 tissue expression in liver biopsies of patients with LT and AR. The study included liver biopsies performed before and after treatment in 12 patients with LT and proven AR. In 10 patients AR was reversible and in 2, was steroid resistant. For immunohistochemistry, an indirect immunoperoxidase technique was used. Each histology section was semiquantitatively evaluated as follows: 0: < 10% staining, 1: 10-25%, 2: 25-50%, 3: > 50%. The control group comprised nine patients with LT and normal liver biopsies. In pre-treatment liver biopsy samples, ICAM-1 was markedly expressed on sinusoidal cells (2.41 +/- 0.66), and there was also expression on periportal (0.66 +/- 0.65) and perivenular hepatocytes (0.83 +/- 0.57). By contrast, in the liver tissue from the control group, sinusoidal ICAM-1 reactivity was significantly lower (0.88 +/- 0.33; P < 0.05), and hepatocytes showed no reliable ICAM-1 expression. After steroid treatment the intensity of ICAM-1 decreased significantly in sinusoids (1.5 +/- 0.67; P < 0.05) and in perivenular hepatocytes (0.25 +/- 0.86; P < 0.05). Additionally, we also observed a decreased ICAM-1 reactivity in portal hepatocytes (0.25 +/- 0.62), but these differences did not reach statistical significance. Remarkably, after treatment, hepatocytes did not show ICAM-1 reactivity in resolved AR, but in corticoid-resistant patients AR did not change or increase. In conclusion, in patients with LT and AR, ICAM-1 was expressed in hepatocytes and with more intensity in sinusoid cells. Additionally, a down-regulation of the ICAM-1 tissue expression after corticoid treatment may exist, although in corticoid-resistant AR no modulation on ICAM-1 tissue expression was observed. PMID- 11112054 TI - Donor DNA can be detected in recipient tissues during rejection of allograft. AB - The main source of donor DNA in recipients of allograft are "passenger" cells. It is claimed that they are responsible for the posttransplantation microchimerism and prolongation of allograft survival. We have observed that besides cellular microchimerism, donor DNA can be found in the recipient tissues at the time of rejection of the allograft. In this study, we provide evidence for the presence in the recipient of both DNA in "passenger cells" and free DNA in tissues at the terminal stage of rejection. Male BN (RT1 n) rat heart or skin was transplanted to female LEW (RT1 l) rats followed by a vascularized bone marrow in a hindlimb transplant. In another group, heart and skin were transplanted followed by immediate i.v. infusion of donor-type bone marrow cells. CsA was given in a dose of 17 mg/kg body weight for 30 days, then the rats were followed up until day 100 unless rejection occurred earlier. LEW blood, spleen, mesenteric node and bone marrow cells were stained with moAb OX27 specific for BN but not LEW. Genomic male DNA was isolated and amplified with SRY oligonucleotide. At day 30 and day 100 cellular microchimerism was detected in blood, spleen, nodes and bone marrow cells. Donor DNA was detected in recipient skin, liver and heart extracts, as well as lymphoid organs, at the time of rejection of allograft, but not when the rats were maintained on CsA. Taken together, donor DNAwas detected in recipient tissues at the time of heart or skin rejection. It appeared to be released from cells of rejecting grafts and not from "passenger" cells, representing only a minor cellular mass compared with the graft. PMID- 11112055 TI - Analysis of cellular events in hepatic allografts: donor progenitors induce intragraft chimerism. AB - Long-term graft acceptance and tolerance induction after allogeneic rat liver transplantation are well described. However, the underlying mechanisms remain unclear. In this study we investigated the cellular events within the liver graft during initial immunosuppression and long-term acceptance. Orthotopic liver transplantation was performed in the Dark Agouti (DA)-to-Lewis (LEW) and LEW-to DA rat strain combination. In order to achieve long-term acceptance, LEW recipients of DA livers were treated with two different short-term therapies. Non parenchymal cells (NPC) were isolated from liver allografts on days + 10 and + 100 after transplantation and donor-specific leukocytes were immunophenotyped by flow cytometry. Both the monotherapy and triple therapy prolonged graft survival (> 100 days). Liver allografts from LEW donors into DA recipients were spontaneously accepted across a complete MHC mismatch without immunosuppression. Liver allograft rejection was induced by infiltrating alloreactive immunocompetent cells. But the intensities of cell infiltration in the early and late phases after transplantation did not correlate with eventual outcome. Donor specific NPC decreased to 18-25% on day + 10 in both therapeutic groups, but had rebounded to up to 40% by day + 100. Recurrence of donor-specific cells was caused almost exclusively by rising T cell counts. The persistence of dendritic cells in the late phase after transplantation could be clearly demonstrated. Repopulation by donor-specific T lymphocytes was observed in long-term accepted liver grafts. This recurrence may be based on the differentiation of liver derived progenitor cells. The persistent coexistence of donor and recipient cells within the liver allograft (intrahepatic chimerism) appears to be characteristic and may be important for long-term acceptance. PMID- 11112056 TI - Quantitation of cyclosporine-sensitive and -resistant allospecific cytotoxic cells at birth. AB - In the absence of clinically relevant models of acute rejection we have attempted to develop an assay to measure cyclosporine-resistant allospecific cytotoxic cells in vitro, beginning at birth. The principle of limiting dilution analysis was applied to investigate umbilical cord bloods as responders. Responders were incubated for 1 h in different concentrations of cyclosporine and irradiated HLA mismatched stimulator cells from healthy adults added, followed by recombinant IL 2. After 7 days, responders were tested against three europium-labelled PHA blasts: stimulator, responder and third party. A significant number of cyclosporine-resistant allospecific cytotoxic cell precursors were found in cord blood indicating prior activation. They may have been primed in utero against non inherited maternal HLA antigens. Cyclosporine-resistant allospecific cytotoxic cell precursors were demonstrated in human umbilical cord blood using a quantitative assay. These cells may influence the reaction to subsequent transplants. PMID- 11112057 TI - Allo-tolerance and allorejection responses ex vivo. AB - Spleen cells from fully immune competent mice show different intracellular STAT responses to alloantigen. Cells from mice primed to accept the alloantigen have low STAT 6 and fragmented STAT 4, compared to cells from mice primed to reject the same alloantigen. PMID- 11112058 TI - Mechanisms of tolerance induction in second renal allografts of a chronic rejection model. AB - In a previous experiment we demonstrated the induction of tolerance by the allograft itself. In this model of weak histoincompatibility, second grafts of donor origin replacing chronically rejected first renal allografts were accepted long term. Additionally grafted donor-specific hearts functioned indefinitely while adoptive transfer experiments demonstrated the development of donor specific transferable tolerance. In the current experiment we compared intragraft gene expression of chronically rejected first and tolerant second grafts by RT PCR. Second renal allografts of donor origin (F-344) replaced first grafts 2, 4, 8, 12, and 16 weeks after the initial engraftment. No immunosuppression was used during second engraftment. Grafts were followed by serial proteinuria; morphological and immunohistological studies (APAAP/infiltrating cells, ICAM-1, MHC II expression) and competitive RT-PCR analyses (expressed as arbitary units AU/cDNA) for relevant cells and cytokines (CD-3, IFNgamma, IL-10, and IL-4) were assessed by the end of the observation period (16 weeks). Macrophages/monocytes (ED-1+) and T-cells (CD-5 and CD-4+) infiltrated first allografts in high numbers by 12 weeks associated with strong structural signs of chronic graft rejection (ca. 30% arterio- and glomerulosclerosis, tubular atrophy and interstitial fibrosis). Cellular infiltrates in second grafts were prominent, however significantly reduced, while histological changes were minor. At cDNA levels, CD 3 transcripts were elevated in second renal allografts performed 2, 4, and 8 weeks after the initial engraftment while comparable levels were observed when second engraftment was performed after 12 and 16 weeks. Analyses of relevant cytokines demonstrated a TH1/TH2 shift independent from the time interval between first and second engraftment. These results emphasize the role of alloresponsiveness for the development of chronic graft dysfunction. Mechanisms of tolerance induction in our model are associated with a distinct alloresponsive pattern. A crucial role for regulatory T-cells is suggested. PMID- 11112059 TI - Prolonged survival of baboon renal allografts using idarubicin-conjugated anti CD4 monoclonal antibodies. AB - Tolerance to organ allografts in rodents and pigs can be easily achieved. However, tolerance induction in a large primate model has been more elusive. In this study, we have used an anti-CD4, murine monoclonal antibody as a carrier for the cytotoxic drug idarubicin (IDA) to delete or inactivate alloreactive T-cells responding to a renal allograft in a baboon transplant model. Fourteen Chacma baboons weighing between 15-25 kg received heterotopic renal allografts. Recipient and donor pairs were selected on the basis of ABO compatibility. Seven animals were given no immunosuppression and served as the control group. The remaining 7 animals received anti-CD4 IDA. The first 2 animals in this group received 2 mg IVI intraoperatively and three doses at 48-h intervals thereafter. The last 5 animals received a larger dose of 1 mg/kg, starting 24 h preoperatively and again on postoperative days 2 and 5. The untreated animals promptly rejected their allografts with a mean survival of 10 days. The survival of the 2 animals treated with 2 mg anti-CD4 IDA was 7 days each. However, the animals treated with 1 mg/kg anti-CD4 IDA survived 7, 18, 20, 40 and > 40 days. Peritransplant administration of anti-CD4 IDA prolonged renal allograft survival in a large primate model. This unique immunoconjugate has the potential of tolerance induction. PMID- 11112060 TI - Adenovirus-mediated viral IL-10 gene transfer prolongs survival of xenogeneic spheroidal aggregate-cultured hepatocytes. AB - Xenotransplantation of hepatocytes appears to be a novel promising therapy for some forms of liver disease, and may well overcome the problem of donor shortage. We have previously reported that hepatocytes with a spheroidal shape (spheroids) are ideal for cell transplantation. The application of gene transfer techniques to this hepatocyte transplantation could possibly regulate the xenogeneic rejection reaction and, therefore, result in prolongation of the survival of the transplanted hepatocytes. In this study, we chose the adenovirus as a vector and an immunosuppressive cytokine named viral IL-10 (vIL-10) for transfection. A series of experiments was performed to elucidate the efficacy of transfection to the spheroids with adenovirus vectors and the effect of transfected vIL-10 on the survival of xenogeneic hepatocytes. We examined the cell survival quantitatively by evaluating beta-galactosidase (beta-gal) activity, which was transfected into the hepatocytes in the xenogeneic spleen, and semiquantitatively by the histological findings. The results of in-vitro studies identified an efficient expression of the beta-gal gene within the spheroids infected with Ad-CMVLacZ (LacZ-encoding adenovirus vector with CMV promotor) and the presence of BCRF1 mRNA within the spheroids transfected with AdCMVvIL-10 (vIL-10-expressing adenovirus vector with CMV promotor) under the condition of 1 MOI, for 1 h. Xenogeneic hepatocytes with a spheroidal shape showed comparable survival to syngeneic hepatocytes for up to 4 days after transplantation with co transplantation of the vIL-10-transfected hepatocytes. From this study, we concluded that adenovirus-mediated vIL-10 gene transfer prolongs the survival of xenogeneic hepatocyte spheroids. Furthermore, spheroids possess ideal properties for gene transfection, as well as cell transplantation. PMID- 11112061 TI - Influence of ischemic time on hyperacute xenograft rejection of pig hearts in a working heart perfusion model with human blood. AB - In xenotransplantation long ischemic time of grafts is supposed to have a marked influence on hyperacute rejection (HXR). We investigated the influence of different cold ischemic times on HXR of ex vivo "working pig hearts" perfused with human blood. Xenoreactive natural antibodies (XNAb) as a trigger of HXR were eliminated by Ig-Therasorb immunoadsorption (IA). Explanted Landrace pig hearts of group G1 and group G3 (with additional IA) underwent 4 h of cold ischemia prior to xenoperfusion. Control groups G2 and G4 (with IA) were kept ischemic for only 46.6 +/- 15.8 and 51.2 +/- 4.2 min, respectively. Ischemic time prolonged the perfusion time in our working heart model (G1: 356 +/- 46.1 min; G2: 125 +/- 31 min; P < 0.05). IA had no additional impact on perfusion time but was effective by itself. The heart weight increased fourfold more in G2 as compared to the other groups. IA without ischemia significantly improved cardiac output in G4 (G3: 198.8 +/- 15.4 mL/min; G4: 338.5 +/- 16.0 mL/min). Coronary flow in G2 was significantly lower than in G1 (G1: 157.9 +/- 9.15 mL/min; G2: 59.4 +/- 20.1 mL/min). Histological signs of HXR (light and electron microscopy) could be found in G2 in contrast to the other groups. Parameters of serological damage showed a minimum in G4 and the maximum in G2. In G1 XNAb were nearly equally eliminated immediately after the start of xenoperfusion as in IA groups G4 and G3. Four hours of ischemic time showed beneficial effects in preventing HXR, possibly caused by changes of the endothelial cell surface (for example, glycosylation or loss of alpha1-3Gal epitopes with a hapten effect). PMID- 11112062 TI - Negligible role for NK cells and macrophages in delayed xenograft rejection. AB - Hyperacute rejection (HAR) of a discordant xenograft can be avoided by complement manipulation, but delayed xenograft rejection (DXR) still leads to graft loss. It is generally assumed that macrophages and NK cells play key roles in DXR. In the present study the survival times and cellular infiltrate following guinea pig to rat heart transplantation was analyzed in the course of DXR, following aspecific and specific manipulation of macrophages and NK cells. HAR was overcome by a single injection of cobra venom factor 1 day before heart transplantation. To aspecifically reduce the inflammatory response dominating DXR, dexamethasone (DEXA) was given. Treatment with DEXA markedly reduced infiltration by NK cells, macrophages, and granulocytes. It also led to prolonged graft survival times (median survival of 0.4 days, n = 10, P < 0.05). In the second series of experiments the specific roles of NK cells and macrophages in DXR were further assessed. Monoclonal antibody 3.2.3 was used to selectively deplete NK cells. Liposome-encapsulated dichloromethylene biphosphonate was given to achieve macrophage depletion. Neither of these specific treatments, alone or combined, led to prolonged graft survival. Immunohistology revealed that at day 2 after transplantation no NK cells or macrophages were present in grafts from the combined treatment group. Only a mild infiltration of granulocytes was observed. Collectively, these results strongly suggest that NK cells and macrophages are not likely to be pivotal cell types in DXR. PMID- 11112063 TI - Prevention of hyperacute xenograft rejection in orthotopic xenotransplantation of pig hearts into baboons using immunoadsorption of antibodies and complement factors. AB - To prevent hyperacute xenograft rejection (HXR) caused by preformed natural antibodies (XNAb) after orthotopic heart xenotransplantation (oXHTx) of landrace pig hearts into baboons, we used immunoadsorption of immunoglobulins IgG, IgM and IgA and complement with the reusable Ig-Therasorb column. In addition to functional data, tissue was sampled for histological, immunohistochemical and electron microscopical analysis. We performed three oXHTx of landrace pig hearts to baboons using extracorporeal circulation (ECC) connected to the immunoadsorption unit. Intraoperative treatment consisted of four cycles of immunoabsorption (IA). One oXHTx of a baboon without IA served as a control. A mismatch of donor and recipient heart size was prevented by selecting a 30-40% lower body weight of donor pigs than recipients. Four cycles of IA removed more than 80% of IgG, IgM and IgA, 86% of antipig antibodies and 66% of complement factors C3 and C4 from plasma. The graft of the control animal failed after 29 min. Orthotopic xenotransplantation with IA was selectively terminated after 100 min, 11 h and 21 h, respectively without any histological signs of HXR in light and electron microscopy. After weaning off from ECC these donor xenografts showed sufficient function with normal ECG and excellent cardiac output in echocardiography and invasive measurement (1.93 +/- 0.035 l/min). The myocardium of the control xenograft demonstrated more deposits of Ig and complement components (C3, C4) than in the IA group. Baboons survive HXR after orthotopic pig heart xenotransplantation due to antibody depletion by reusable Ig-Therasorb column treatment. Long-term survival in an orthotopic baboon xenotransplantation model after IA, especially in combination with transgenic pig organs, could be a reliable preclinical trial for future clinical xenotransplantation programs. PMID- 11112064 TI - First human hand transplantation. Case report. PMID- 11112065 TI - Reperfusion injury is dramatically increased by gentle liver manipulation during harvest. AB - Kupffer cell-dependent injury in livers gently manipulated during harvest develops upon reperfusion. The purpose of this study was to characterize this injury and to detect underlying mechanisms. Livers from female Sprague-Dawley rats were harvested for transplantation within 25 min. Minimal dissection was performed during the first 12 min, including freeing the liver from ligaments. After this, for further 13 min, livers were either left alone or manipulated gently. The livers were then cold-stored for 24 h in University of Wisconsin (UW) solution and perfused with oxygen-saturated Krebs-Henseleit buffer at 37 degrees C. Trypan blue in the buffer was used to index microcirculation. Cell damage was assessed with histology. Initial dissection during harvest and cold storage had minimal effects on sinusoidal lining cells; in contrast, the subsequent gentle organ manipulation dramatically increased cell death 6.5-fold, while the time for complete trypan blue distribution increased 2.3-fold (P < 0.05). Manipulation increased proteolysis 2-fold (P < 0.05). At harvest, manipulation increased portal venous pressure significantly by 68%. Treatment of donors with gadolinium chloride, a selective Kupffer cell toxicant, or with dietary glycine, an inhibitor of Kupffer cell activation, prevented effects of organ manipulation on all parameters studied. These findings demonstrate Kupffer cell-dependent reperfusion injury of sinusoidal lining cells caused by manipulation of the liver during its recovery. The mechanisms are those of proteolysis and impaired hepatic microcirculation. PMID- 11112066 TI - Heart valve dysfunction resulting from cellular rejection in a novel heterotopic transplantation rat model. AB - Structural failure of heart valve allografts may be related to technical factors or immunological reactions. To circumvent nonimmunological factors a new rat implantation model was developed to study whether alloreactivity results in histopathological changes and valve dysfunction. Syngeneic (WAG-WAG, DA-DA) and allogeneic (WAG-BN, WAG-DA) transplantation was carried out using this new technique, and the function of explanted valves was assessed 21 days later by retrograde competence testing. Additionally, grafts were examined using standard histological and immunohistochemical techniques. There was no leakage during retrograde injection in nine of tem syngeneic and two of ten allogeneic grafts. Microscopically, syngeneic valves appeared normal without fibrosis or intimal thickening, although CD8+ lymphocytes and macrophages were found in necrotic myocardial rim and adventitia. In contrast, allogeneic valves were deformed and noncellular, with extensive infiltration of CD4+, CD8+ and CD68+ cells in adventitia and media. Absence of fibrosis and intimal thickening in syngeneic transplanted valves indicated circumvention of nonimmunological factors. Allogeneic valve transplantation induces cellular infiltration in the graft with subsequent graft failure. PMID- 11112067 TI - Long-term small bowel allograft function induced by short-term FK 506 application is associated with split tolerance. AB - Functional long-term allograft survival after experimental small bowel transplantation (SBT) is limited by chronic rejection. Initial application of high-dose FK 506 has been shown to induce stable long-term graft function. In order to examine whether this long-term function is associated with donor specific tolerance, we analyzed the functional status of recipient T cells in vivo and in vitro. One-step orthotopic SBT was performed in the allogeneic Brown Norway (BN)-to-Lewis rat strain combination. FK 506 was given daily at a dose of 2 mg/kg from days 0-5 in the rejection model and from days 0-9 in the long-term functional model. Mean survival time in the rejection model was 98 +/- 2.8 days. Histological examination of these small bowel allografts disclosed signs of chronic rejection. In contrast, all animals of the long-term functional model survived long term (> 250 days) without clinical signs of chronic rejection. The latter model, furthermore, produced evidence of donor-specific tolerance. Whereas heterotopic Dark Agouti (DA) hearts were rejected regularly within 7 days, BN hearts survived indefinitely (> 70 days). In vitro, mixed leukocyte reactivity of CD4+ T cells was similarly strong against donor (BN) antigens as against third party (DA) antigens. The split tolerance revealed by our in vivo and in vitro results enabled acceptance of both the small bowel allograft without signs of chronic rejection and of donor-specific heart allografts. PMID- 11112068 TI - Different kinetics of donor cell populations after isolated liver and combined liver/small bowel transplantation. AB - Spontaneous tolerance induction after liver transplantation also supports additional transplants, e.g. a small bowel graft, from the same donor (tolerogenic effect). Chimerism serves as a possible explanation of this phenomenon. Isolated liver (LTx) and combined liver/small bowel transplantation (LSBTx) are compared. LSBTx and LTx were performed in the BN --> LEW rat strain combination without immunosuppression. Parenchymal damage during rejection was monitored by sequential standard histology. Donor/recipient populations were identified and further differentiated for immunohistochemical single and double staining. A small number of donor specific leukocytes can be detected on all days in host organs (microchimerism). A significantly larger donor leukocyte population survives long-term in the sinusoids of liver (graft chimerism). Sinusoidal donor leukocytes survive rejection and recover in number after tolerance induction. Rejection of liver allografts and infiltration by host leukocytes are more pronounced after LSBTx than after LTx. Accordingly, during rejection a steeper decline of sinusoidal donor leukocytes is observed after LSBTx and recovery after tolerance induction is not as marked. Microchimerism apparently plays no significant role in either transplantation model. The number of sinusoidal donor leukocytes, however, mirrors closely host immune responses. PMID- 11112069 TI - Vascularized bone marrow transplanted in orthotopic hind-limb stimulates hematopoietic recovery in total-body-irradiated rats. AB - Hematopoietic recovery after bone marrow transplantation (BMT) is reported to be slow with long-lasting immune deficiency. This may be attributable to lack of a proper microenvironment for hematopoietic cell proliferation and differentiation. We have designed a model in which complete hematopoietic reconstitution of lethally irradiated rats can be achieved by vascularized bone marrow transplantation (VBMT) in an orthotopic hind-limb graft. The aim of the study was to investigate the process of repopulation of bone marrow cavities and peripheral blood of irradiated rats after VBMT and, in particular, to follow the contribution of grafted BM cells and residual recipient BM cells in hematopoietic regeneration. Lewis hind-limbs were transplanted orthotopically to totally irradiated (8 Gy) syngeneic sex-mismatched recipients (VBMT). In the control group 8 x 10(7) BM cells in suspension were injected intravenously (BMCT). After 10 days BM and peripheral blood (PB) cells were collected from the recipient. For cell subset analysis cytomorphological evaluation of BM smears and flow cytometry of PB cells were performed. Additionally, PCR was performed using specific primers for rat Y chromosome (sex-determining region Y-Sry) to detect male (donor or recipient) cells in sex-mismatched BM graft recipients and the products were analyzed by electrophoresis. VBMT brought about much faster replenishment of nucleated cells in BM and PB than did BMCT. Cytometry analysis of PB cells revealed more lymphocytes in VBMT than in BMCT recipients. The amount of donor DNA of bands corresponding to Y-Sry was also higher in PB cells of VBMT than of BMCT recipients. The presence of host DNA was observed in PB cells of VBMT rats but was not detected in PB population of BMCT recipients. VBMT is highly effective in hematopoietic reconstitution of irradiated recipients. The fast cell maturation and repopulation may be due to the presence of stromal cells transplanted in a normal spatial relationship with donor hematopoietic cells in hind-limb graft. Self renewal of radioresistant host cells was seen after VBMT but not after BMCT. PMID- 11112070 TI - In vivo microscopy reveals that complement inhibition by C1-esterase inhibitor reduces ischemia/reperfusion injury in the liver. AB - Complement plays a decisive role in postischemic tissue injury, a process responsible for severe damage after organ ischemia. Several pathophysiologic mechanisms initiated upon reperfusion are mediated by complement inducing microcirculatory disturbances. Here, we demonstrate the effects of complement inhibition using C1-esterase inhibitor (C1-INH) on microcirculation after liver ischemia by in vivo microscopy (IVM). In rats, the left liver lobe was clamped for 70 min. C1-INH was given 1 min prior to reperfusion. Controls received Ringer's solution. IVM was performed 30-100 min after reperfusion. Non-perfused acini decreased and sinusoidal perfusion increased substantially after treatment. Leukocyte adherence to sinusoidal and venular endothelium was markedly reduced by C1-INH. Transaminases were significantly decreased by C1-INH. Our data obtained by IVM suggest that complement activation is an early key event of ischemia/reperfusion injury. These observations demonstrate for the first time that reperfusion related microcirculatory disorders can be minimized by C1-INH. This compound should be evaluated in clinical application. PMID- 11112071 TI - Value of alpha glutathione S-transferase for in vitro evaluation of preservation injury in normal and steatotic livers. AB - Liver steatosis is frequently encountered at organ harvest and, although functionally inapparent in the donor, may seriously affect the functional recovery of the graft after ischemic preservation. The present study was aimed to investigate the diagnostic value of alpha-glutathione S-transferase (GST) in non ischemic and ischemic livers with or without compensated steatosis. A histologically documented mild to moderate steatosis was induced in livers of male Wistar rats by fasting for 2 days and subsequent feeding of a fat-free diet enriched in carbohydrates. Fatty livers (FL) were retrieved and perfused in vitro for 45 min either immediately or after ischemic preservation at 4 degrees C in HTK solution. Effluate was collected during isolated perfusion and later analysed for liver specific enzymes, including GST. Normal livers (NL) were excised from healthy rats and underwent the same protocol. Non-ischemic livers showed similar enzyme release (FL versus NL) for ALT or GLDH but significant differences in GST. After ischemic preservation of NL, enzyme release increased mildly with respect to the non-ischemic reference values for ALT, remained unchanged for GLDH and rose substantially for GST. In FL, there was a more than 10-fold increase in all parameters, being most pronounced for GLDH as a marker of mitochondrial damage. It is concluded that GST may discriminate between healthy and suboptimal steatotic livers prior to ischemia and that the release of GST upon postischemic reperfusion of normal livers proves to be the most sensitive indicator for hepatocellular injury. However, GST turned out to be less useful for the detection of postischemic reperfusion injury in steatotic grafts. PMID- 11112072 TI - Mitochondrial defects by intracellular calcium overload versus endothelial cold ischemia/reperfusion injury. AB - Questions as to the critical stress factor and primary targets of cold ischemia/reperfusion (CIR) injury were addressed by comparing mitochondrial defects caused by (1) CIR injury and (2) intracellular Ca2+ overload. CIR was simulated in transformed human umbilical vein endothelial cell cultures (tEC) by 8 h cold anoxia in University of Wisconsin solution and reoxygenation at 37 degrees C. Intracellular Ca2+ concentrations were changed by permeabilization of suspended cells with digitonin in culture medium (RPMI, 0.4 mM Ca2+). Binding of free Ca2+ by ethylene glycol-bis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid in RPMI or mitochondrial incubation medium served as controls. Extracellular Ca2+ protected the cell membrane against permeabilization. Mitochondrial functions were determined before and after permeabilization of the cell membrane. After CIR, mitochondrial respiratory capacity declined, but oxygen consumption remained coupled to adenosine triphosphate (ATP) production. In contrast, Ca2+ overload caused uncoupling of mitochondrial respiration. High intracellular Ca2+ overload, therefore, does not reproduce cold ischemia/reperfusion injury in endothelial cells. PMID- 11112073 TI - The liver protective effect of ischemic preconditioning may be mediated by adenosine. AB - We investigated the involvement of adenosine in ischemic preconditioning (IPC) by the unspecific antagonist, 8-phenyltheophylline (8-PT). Anesthetized Wistar rats were treated as follows: 1. non-ischemic controls, 2. ischemic controls: 60 min of clamping of the common hepatic artery followed by 60 min reperfusion, 3. IPC: 10 min ischemia followed by 15 min reperfusion, prior to the identical ischemia reperfusion (IR) period as in group 2, 4. 8-PT + IPC: 8-PT 10 mg/kg i.v. was given 10 min prior to the identical procedure as in group 3. The peripheral liver blood flow was monitored by laser-Doppler flowmetry. Blood alanine aminotransferase (ALT) was analyzed once every 60 min. IPC significantly reduced impairment of liver blood flow, as well as ALT increase during reperfusion. This effect was abolished by pretreatment with 8-PT. Adenosine appears to be a crucial effector in IPC. Clinical studies need to be undertaken to explore a possible effect of IPC in liver transplantation. PMID- 11112074 TI - Reduction of hepatic reperfusion injury by antithrombin III and aprotinin. AB - Disturbance in hepatic microcirculation and leucocyte-endothelium interaction after warm ischaemia represents one of the leading mechanisms for postoperative organ dysfunction. Recent studies have shown that pretreatment with antithrombin III (AT III) and aprotinin reduces the leucocyte-endothelium interaction in ischaemic small intestine and during extracorporal circulation in cardiac surgery. Standardized warm hepatic ischaemia and intravital fluorescence videomicroscopy was performed in an experimental study with rats. Animals were pretreated with AT III or aprotinin. Analysis of intravital videomicroscopy showed that the hepatic microcirculation after warm hepatic ischaemia in rats was significantly enhanced by AT III and aprotinin, most likely by reducing the leucocyte-endothelium interaction. We concluded that drug application before the Pringle manoeuvre might reduce the reperfusion damage after liver resection. PMID- 11112075 TI - LF 08-0299 in the prophylaxis and treatment of chronic rejection in a rat aortic allograft model. AB - Chronic rejection is the major cause of late kidney allograft failure. We evaluated the efficacy of LF 08-299 (LF), an analogue of 15-deoxyspergualin, in a rat aortic allograft model of chronic rejection. BN aortic allografts were transplanted to Lew recipients. LF was administered at a dose of 6 mg/kg and 2.5 mg/kg on days 0-20 and 6 mg/kg on days 60-90. CyA was used at a dose of 5 mg/kg on days 0-20. Untreated isografts and allografts were used as controls. Histological changes and immunohistochemistry were monitored sequentially at 8, 12, 16 and 20 weeks. There were no differences in intimal proliferation between LF-treated allografts and untreated or CyA-treated controls. Only a tendency in adventitial infiltration reduction was seen in LF-treated animals. We found a significantly less pronounced reduction in media diameter in LF-treated animals. We concluded that LF 08-0299 is only able to reverse reduction in media thickness in aortic allografts, but not intimal proliferation in this model of chronic rejection. PMID- 11112076 TI - Caspase inhibition protects from liver injury following ischemia and reperfusion in rats. AB - Normothermic ischemia and reperfusion of the liver results in microcirculatory failure followed by necrosis and cell death. Recently, another type of cell death, apoptosis or programmed cell death, was found to be activated during the early phase of reperfusion after liver ischemia. Caspases are cysteine proteinases specifically involved in the initiation and execution phases of apoptosis. The aim of this study was to demonstrate that inhibition of apoptosis by a specific inhibitor of caspases might protect the liver against ischemia/reperfusion injury. Rats were divided into three groups: group 1, control, PBS administration; group 2, Z-Asp-cmk (Z-Asp-2,6-dichlorobenzoyl oxymethylketone) treatment; group 3, sham-operated control animals. Z-Asp-cmk (0.5 mg Z-Asp-cmk dissolved in 300 microl PBS solution containing 1% DMSO) was injected intravenously, 2 min prior to induction of 120 min ischemia. Survival rates were compared and serum activities of aspartate aminotransferases and alanine aminotransferases were assessed in the blood collected from the suprahepatic vena cava. Histology of the liver was assessed 6 h after the end of ischemia. Apoptosis was detected by the terminal deoxynucleotidyl transferase mediated dUTP-FITC nick end-labeling method (TUNEL method) and by electrophoresis for analysis of DNA fragmentation. Caspase activity was determined by measuring hydrolysis of the CPP32-like substrate Ac-DEVD-pNA and absorption of paranitroaniline. Z-Asp-cmk treatment significantly increased 7-day survival (95%) compared with that in nontreated rats (30%, P < 0.001). Serum activities of aminotransferases and the extent of liver congestion and necrosis were significantly (P < 0.001) decreased after treatment with Z-Asp-cmk. TUNEL positive cells were detected 3-6 h after reperfusion in the control group. In Z Asp-cmk pretreated rats, a dramatic decrease in the number of TUNEL-positive cells was observed. Analysis of DNA fragmentation of freshly isolated hepatocytes confirmed these results. Caspase activity was increased 3-6 h after reperfusion in the control group, but significantly (P < 0.001) decreased after treatment with Z-Asp-cmk. These findings demonstrate that liver injury following ischemia and reperfusion can be prevented by inhibition of caspases. Caspase inhibitors may have important implications for therapy in liver disease and after liver transplantation. PMID- 11112077 TI - The effects of the elimination of Kupffer cells in the isolated perfused liver from non-heart-beating rat. AB - We examined the effect of elimination of Kupffer cells on the sinusoidal microcirculation in graft harvested from non-heart-beating donors (NHBD), focusing on the arachidonic acid cascade and cytokines. Cardiac arrest was induced by thoracotomy. Livers were harvested 30 min after thoracotomy and perfused by Krebs-Henseleit bicarbonate buffer for 60 min after 6 h cold preservation. For the elimination of Kupffer cells, rats were pretreated liposome encapsulated dichloromethylene diphosphonate (KE group). Eicosanoids (TXB2, 6 keto-PGF1alpha, LTB4) and cytokines (TNFalpha, IL-1beta) in the perfusate were measured. Histological examination was also carried out. In the KE group, the value of TXB2 was suppressed completely and cytokines were reduced, and sinusoidal structures and hepatocytes were well protected. These results indicated that the elimination of Kupffer cells improved sinusoidal microcirculation in NHBD and liver transplantation using grafts from NHBD could be made to succeed by modulation of Kupffer cells. PMID- 11112078 TI - Mitochondrial calcium overload is restricted to a few mitochondria in endothelial cells after cold ischemia/reperfusion. AB - Changes in cytosolic and mitochondrial calcium content were studied in an endothelial cell model after simulating cold ischemia reperfusion injury. Image analysis demonstrated that only a subpopulation of mitochondria in endothelial cells accumulate calcium. Observations led to the hypothesis that mitochondria which are in close contact with the plasma membrane are mainly affected by the Ca2+ efflux across that membrane, while those in other parts of the cell remain unaffected. PMID- 11112079 TI - Mechanism of primary graft non-function in a rat model for fatty liver transplantation. AB - We established a fatty liver model in rat suitable for the model of human liver with steatosis by cholesterol enriched chow, and investigated the mechanism of primary graft non-function in fatty liver transplantation (LTx) using this model. Grafts with steatosis caused primary graft dysfunction after LTx following even short cold preservation; however, no significant difference was recognized in mitochondrial function of the graft during preservation. Morphological findings were not different at 1 h after reperfusion between non-steatotic and steatotic livers. Focal necrosis of hepatocytes was seen and the sinusoidal endothelial cells were injured 24 h after reperfusion. In addition, the fluidity of the plasma membrane decreased in fatty liver. Our results indicate that deterioration of sinusoidal endothelial cells after reperfusion causes graft dysfunction in LTx of steatotic liver. PMID- 11112080 TI - Gene transfer of endothelial nitric oxide synthase to pulmonary allografts: impact on acute rejection. AB - Experiments were designed to study whether overexpression of nitric oxide (NO) from endothelial nitric oxide synthase (eNOS) affects acute rejection. Allogenic, orthotopic single-lung transplantation was performed after transbronchial adenoviral-mediated gene transfer (3 x 10(8) pfu) of either of eNOS or beta galactosidase to donor lungs of rats (n = 6 each). No immunosuppression was used. After 4 days, transplanted lungs were prepared for enzyme activity, cGMP and histology. Calcium-dependent NOS activity, reflecting eNOS, was greater in eNOS transduced lungs (587 +/- 97 vs 2.1 +/- 1.4 pmol/mg protein per h, P <0.001). In contrast, calcium-independent NOS activity, reflecting iNOS, was comparable. Concentrations of cGMP were higher in eNOS-transduced lungs (13.2 +/- 2.3 vs 4.9 +/- 0.5 pmol/mg protein). Positive immunostaining for eNOS was present in pneumocytes only in eNOS-transduced lungs. No difference in histological grade of rejection was observed. eNOS gene transfer to pulmonary allografts results in a functionally active transgene product and increased NO production. Increasing NO from eNOS does not affect histogically identified acute rejection. PMID- 11112081 TI - Hypoxia-reoxygenation differentially stimulates stress-activated protein kinases in primary-cultured rat hepatocytes. AB - Organ injury after ischemia and reperfusion (I/R) remains one of the most important limiting factors in liver surgery and transplantation. Oxygen-free radical (OFR) generation is considered a major cause of this damage. JNK1/SAPK1, a member of MAPK family, regulates cell adaptation to stressful conditions. The aim of this study was to determine if hypoxia-reoxygenation (H/R) can activate JNK1/SAPK1 and if OFR are involved in this activation. Primary cultured rat hepatocytes isolated from other liver cells and blood flow were submitted to warm and cold H/R phases mimicking surgical and transplant conditions. JNK1/SAPK1 was activated by both warm and cold H/R. Deferoxamine (1 mM), di-phenyleneiodonium (50 microM) and N-acetylcysteine (10 mM) significantly inhibited this kinase activation. PMID- 11112082 TI - Intramucosal pH and liver endotoxin clearance during experimental liver transplantation. AB - The study was designed to assess the gastrointestinal ischaemia and the influence of the specific Kupffer cell toxin gadolinium chloride (GdCl3) on the hepatic and extrahepatic endotoxin [lipopolysaccharide (LPS)] clearance during experimental orthotopic liver transplantation (OLT) in pigs. In eight pig liver transplantations, the donors received 20 mg/kg of GdCl3 24 h before explantation, while controls (n = 8) received normal saline. Gastric and sigmoid intramucosal pH (pHi), LPS and endotoxin-neutralising capacity (ENC) levels were measured in the portal vein and superior vena cava after laparatomy, at the end of the anhepatic phase and 1 h after reperfusion. During the anhepatic phase, the sigmoid pHi decreased significantly from 7.32 +/- 0.02 to 7.29 +/- 0.03 (P < 0.001) and was associated with a substantial increase of portal LPS. Following reperfusion, the systemic LPS concentrations were significantly lower in the pretreated group [39 +/- 23 pg/ml (Control); 14 +/- 7 (GdCl3); P < 0.05] suggesting an improved liver LPS clearance [86% (GdCl3); 58.2% (Control); P < 0.05]. This corresponded to an increased ENC in the pretreated group [118 +/- 52 ENU/ml (GdCl3) vs 81 +/- 45 ENU/ml (Control); P < 0.05]. The anhepatic phase induced splanchnic ischaemia which correlated with portal endotoxaemia. Donor preconditioning with GdCl3 leads to lower systemic LPS concentrations in the recipient and increases ENC values in the early phase after OLT. An improved hepatocellular LPS extraction and/or an activation of the extrahepatic reticulo endothelial system as a result of GdCl3 treatment is discussed. PMID- 11112083 TI - Solidarity model: a way to cope with rationing problems in organ transplantation. AB - This short paper discusses the possibility of implementing a solidarity model as a way of improving organ allocation. PMID- 11112084 TI - Assessment of health-related quality-of-life in patients after heart transplantation under therapy with tacrolimus or cyclosporine. AB - Reduction of allograft rejections remains a primary goal for patients after orthotopic heart transplantation. In an open, multicentre, prospectively randomised, parallel group study, patients with primary orthotopic heart transplantation under oral immunosuppressive treatments with tacrolimus (FK506) or cyclosporine (sandimmun) were compared with respect to medical outcome data. As health-related quality-of-life (HRQOL) is also supposed to be an important outcome parameter, it was assessed as a secondary variable in these two patient groups. Patients' self-rated generic HRQOL was assessed 6 weeks, 3 months, 6 months and 12 months after surgery with the SF-36 questionnaire, a generic HRQOL instrument. For 70 patients (46 under tacrolimus, 24 under cyclosporine), intent to-treat analyses were carried out. The tacrolimus group showed improvements in the different HRQOL subscales of the SF-36 compared to the cyclosporine group. Especially the SF-36 subscales 'vitality' and 'mental health' showed statistically higher scores for the tacrolimus group. Aggregating psychological and cognitive subscales in the 'mental component score', patients treated with tacrolimus showed a statistically significant improvement compared to the cyclosporine group. The assessment of HRQOL variables in the evaluation of treatment effects proved to be an outcome parameter in this study. The results demonstrate the benefit of tacrolimus with respect to the HRQOL of patients, especially in the psychological dimension. PMID- 11112085 TI - Changes in serum levels of quinolones in rats under the influence of experimental trauma. AB - Administration of antibiotics is considered an important factor during, or after, operational procedures in the maxillofacial area, in order to avoid post-surgical complications. In the present study, the levels of quinolones in serum and tissues such as the parotid gland, the tongue and the bone of the jaws were estimated during traumatic injury in the oral cavity. For this purpose, two groups (A and B) of Wistar rats, consisting of 35 animals each were used. Group A (control) and group B (experimental) were divided in five subgroups (A1, A2, A3, A4, A5, and B1, B2, B3, B4, B5). In the experimental group, model traumatic injury was performed through the whole lenght of the cheek. Subjects received orally ciprofloxacin, pefloxacin, norfloxacin, ofloxacin and cinoxacin. The concentration of quinolones in serum and in most of the tissues was significantly higher in the experimental groups than in controls. In addition, the FFA levels and the weight of adrenals (as indicators of stress) were higher in the trauma groups. Stress seemed to affect many pathophysiological mechanisms which are responsible for the alterations observed. PMID- 11112087 TI - A pharmacokinetic study of JOMO-tech in rats. AB - A pharmacokinetic study of 99mTc labelled JOMO-tech in rats (after intravenous administration of a dose of 20 microg/kg body weight) was conducted. JOMO-tech is a heterogeneous extract derived from Nocardia opaca cell walls. An excellent fitting of the three-compartmental disposition model was achieved. The first apparent elimination half-life was very short (t1/2alpha = 0.0572 +/- 0.01383 h) followed by longer second apparent elimination half-life (t1/2beta = 0.817 +/- 0.1922 h), whereas at late post-treatment time the third apparent elimination half-life (t1/2gamma = 21.7 +/- 2.1 h) proved to be long. The peak concentration in the blood extrapolated to t = 0 yielded 32.3 +/- 7.54 ngeq/ml, this being approximately 2-fold the amount of that measured in the 5th post-treatment minute (16.84 +/- 1.447 ngeq/ml). It was determined that the main route of excretion was renal. Up to the 48th post-treatment hour, 30.03 +/- 2.788% of the dose was excreted via the urine, and only 6.71 +/- 0.973% was excreted in the feces by the 7 rats evaluated. The amount of radioactivity detected in selected tissue samples (expressed in ngeq JOMO-tech/g wet tissue) decreased in the sequence liver > kidneys > lungs > blood > plasma. In the time period studied, the highest amount of the dose was found in the liver, whereas up to the 3rd post-treatment day a practically equivalent part of the dose was found in the excreta and in the liver. PMID- 11112086 TI - Microdialysis study of bromocriptine and its metabolites in rat pituitary and striatum. AB - Bromocriptine, a D2 receptor agonist, was administered intravenously (1mg/kg) to anesthetized rats. Microdialysis probes were implanted in the pituitary and the striatum, known sites of D2 agonist action. Bromocriptine and its metabolites were monitored in plasma and tissue dialysates for 4 h. Drug analyses were performed using two different enzyme immunoassays specific for untransformed bromocriptine or a pool of parent drug plus hydroxylated metabolites. The metabolites/parent drug ratio for areas under the curve was 5.5 in plasma and 1 in the pituitary. No metabolites could be detected in the striatum. Bromocriptine penetration was at least 10-fold greater in the pituitary than in the striatum. The kinetics of bromocriptine in the pituitary and striatum did not parallel those in plasma, indicating that the prolonged action of bromocriptine reported by other authors may be due to slow dissociation from receptors. PMID- 11112088 TI - Liquid extraction and HPLC-DAD assay of hydrochlorothiazide from plasma for a bioequivalence study at the lowest therapeutic dose. AB - The main parameters considered in optimizing the liquid extraction and quantitative assay were the yield, precision, limit of quantification, time required for extraction and concentration, and quantity of solvent. The influence on these parameters of the following factors was examined: nature of the extracting solvent, quantity of solvent, co-extraction solvent, and duration of stirring. Instead of equilibrium parameters of the involved thermodynamic system, a kinetic approach was preferred in terms of the effective partition 'constant', which is not really constant but a function of time and extraction conditions. The final selected method, considered to be rapid and simple, was applied to determine the pharmacokinetics of hydrochlorotiazide (HCT) after administration of Capozide (Bristol-Myers Squibb) tablets containing 50 mg Captopril and 25 mg HCT, to 4 healthy volunteers. The results obtained were in accordance with the pharmacokinetic parameters of HCT reported in the literature. PMID- 11112089 TI - Effect of food on the pharmacodynamics and pharmacokinetics of atorvastatin, an inhibitor of HMG-CoA reductase. AB - The pharmacodynamics and pharmacokinetics of atorvastatin, an HMG-CoA reductase inhibitor, were characterized in 16 healthy subjects following administration of 10 mg atorvastatin tablets with, or 3 h after, evening meals for 15 days in an open-label, randomized, 2-way crossover study. Atorvastatin was well tolerated. Atorvastatin administration with evening meals resulted in 25.2% lower mean Cmax and 29.8% longer mean tmax values relative to administration after meals. The mean AUC(0-24) value was 8.6% lower for atorvastatin administration with meals compared to after meals. In contrast to the effect of food on pharmacokinetics, LDL-C reductions were similar after atorvastatin administration with or after evening meals. Average reductions from baseline were 24.4% for total cholesterol, 39.6% for LDL-C and 10% for triglycerides. Therefore, atorvastatin may be administered with or without food. PMID- 11112090 TI - In vivo metabolism of 4-fluorobenzoic acid [(5-nitro-2-furanyl)methylene] hydrazide in rats. AB - It is known that substituted hydrazide hydrazone derivatives have several biological and pharmacological activities; there is limited literature on the metabolism of hydrazide hydrazones in rats. In our previous study, 4 fluorobenzoic acid [(5-nitro-2-furanyl)methylene]hydrazide (S) was found active against Staphylococcus aureus ATCC 29213. Therefore, we planned to study the in vivo metabolism of S in rats. The substrate was administered in doses of 50 mg/kg or 100 mg/kg intraperitoneally. Blood samples were collected at 0, 5, 15, 30, 45 min and 1, 1.5, 2, 4, 8, 12, 24, 48 h after administration. The substrate and its potential metabolites were separated using HPLC on a reverse phase system. 4 Fluorobenzoic acid and one unidentified metabolite were detected together with substrate. PMID- 11112091 TI - Metabolism of clozapine by rat brain: the role of flavin-containing monooxygenase (FMO) and cytochrome P450 enzymes. AB - The atypical antipsychotic clozapine has been reported to be metabolised mainly to its N-oxide and N-demethylated products. Brain, the target organ of clozapine, is known to contain numerous drug-metabolising enzymes which could alter the local concentrations of the drug. The metabolism of clozapine was, therefore, studied in rat brain preparations. Clozapine N-oxide was the major metabolic pathway in rat brain. We characterised the N-oxygenation of clozapine by rat brain preparations. The Km and Vmax values were found to be 319.6 microM and 28.1 pmol/min/mg protein, respectively. The formation of clozapine N-oxide was shown to be inhibited by thiourea (a flavin-containing monooxygenase inhibitor) but not by ketoconazole, quinidine or furafylline. This finding suggests prominent involvement of FMO in the N-oxygenation of clozapine in rat brain. This conclusion was further confirmed by the observation that the formation of clozapine N-oxide is sensitive to heat treatment of the brain preparation and can be partially protected from thermal degeneration by the presence of an NADPH generating system. It was further observed that the rate of clozapine N oxygenation was much higher at pH 8.5 than at pH 7.4. Taken together, the data suggest that N-oxygenation is the major metabolic pathway catalysed by rat brain and this reaction is catalysed mainly by FMO. As significant interindividual differences have been observed in brain FMO activities, these differences may contribute to the interindividual differences in patient response to clozapine. PMID- 11112092 TI - A double-site absorption model fits to pharmacokinetic data of repaglinide in man. AB - The plasma concentrations of repaglinide in 16 male subjects were determined after an oral dose of 4 mg. Two-peak concentrations in plasma were observed. A type of one-compartment model with double sites of drug absorption was developed and successfully used to fit the data. A good agreement between observed and predicted data was found in all subjects with correlation index r2 > 0.97. The corresponding pharmacokinetic parameters were estimated as follows: Tmax1 0.61 +/ 0.14 h, Tmax2 1.45 +/- 0.43 h, Cmax1 40.60 +/- 20.57 ng/ml, Cmax2 42.70 +/- 17.54 ng/ml, T1 0.12 +/- 0.07 h, T2 0.67 +/- 0.30 h and T3 1.03 +/- 0.35 h. PMID- 11112093 TI - Effects of imidazole antimycotics on the liver microsomal cytochrome P450 isoforms in rats: comparison of in vitro and ex vivo studies. AB - We have studied the effects of three imidazole derivatives, clotrimazole (CLO), ketoconazole (KET) and miconazole (MIC) on the liver microsomal diazepam (DZ) metabolism. In in vitro experiments using rats and human liver microsomes, significant inhibition of CYP3A in terms of DZ-3-hydroxylase activity was observed. The inhibition of DZ metabolism was seen 1 h after CLO dosing. On the other hand, the induction of certain cytochrome P450 (CYP) isozymes was observed in in vivo studies 24 h after dosing. That is, CYP1A, CYP2B and CYP3A2, but not CYP2E, were observed 24 h after CLO or KET or MIC treatment. Under these conditions, CLO was the most potent inducer of CYP3A and MIC was a more potent inducer of CYP1A and CYP2B. KET induced CYP1A and CYP2B whereas the inducibility of KET was less than those of CLO and MIC. All of the imidazole derivatives tested here showed significant inhibition of CYP isozymes which overcame the induction of the CYP isozymes by those drugs in the data of Western blot analysis. PMID- 11112094 TI - Effects of subacutely administered saiboku-to, an oriental herbal medicine, on pharmacodynamics and pharmacokinetics of diazepam in rodents. AB - Subacute treatment with saiboku-to (2000 mg/kg, p.o., once a day) for 7 days induced an anxiolytic-like effect in rats. It did not, however, produce any other effects, such as sedative and hypnotic effects, anticonvulsive and muscle relaxant effects except for anxiolytic effect observed in diazepam-injected rats or mice. Diazepam (1.0 mg/kg, s.c.) induced anxiolytic-like effect was enhanced in saiboku-to treated rats as an additional effect of that induced by saiboku-to. To elucidate whether the enhancement of the anxiolytic-like effect following combined administration of diazepam and saiboku-to is due to the inhibition of hepatic drug-metabolizing enzymes, the pharmacokinetics of diazepam were further investigated in saiboku-to treated rats. The pharmacokinetic studies clearly demonstrated that subacute treatment with saiboku-to did not affect plasma concentration and protein binding rate of diazepam, and the activities of hepatic drug-metabolizing enzymes related to diazepam metabolism. These results, taken together, suggest that the enhancement of diazepam-induced anxiolytic-like effect observed in saiboku-to-treated rats is not due to an inhibition of diazepam metabolism. PMID- 11112095 TI - Mechanism of enhancement by fucoidan and CNBr-fibrinogen digest of the activation of glu-plasminogen by tissue plasminogen activator. AB - The interactions of fucoidan with human glutamic type plasminogen (Glu-Plg), porcine pancreatic elastase digested plasminogen fractions and two chain tissue plasminogen activator t-PA) were investigated using fucoidan-Sepharose affinity chromatography. The results showed a high degree of affinity between fucoidan Sepharose and Glu-Plg or PlgK(1-3) but not with PlgK4 or mini-Plg. Fucoidan Sepharose also showed a high affinity for t-PA, which was largely reversed by 0.002 M 6-aminohexanoic acid (6-AH). The addition of fucoidan and CNBr-fibrinogen digest (CNBr-Fbg) gave the highest enhancement of the in vitro activation of Glu Plg by t-PA in the presence of 0.002 M 6-AH. The results of affinity chromatography and enhancement studies suggested a template mechanism, since increasing the concentrations of any one of the two cofactors reversed the enhancement. Enzyme kinetic studies, using double reciprocal plots, showed that the addition of fucoidan-6-AH increased Kcat by 7-fold without affecting Km and addition of CNBr-Fbg lowered Km by 5-fold without significantly affecting Kcat while addition of the two cofactors lowered Km by 16-fold without significantly affecting Kcat. The enhancement by fucoidan-6-AH or by CNBr-Fbg of the in vitro activation of Glu-Plg by t-PA was reversed by plasminogen activator inhibitor 1 (PAI-1). Fucoidan-Sepharose affinity chromatography revealed that the binding of PAI-1 with fucoidan may be responsible for the reversal of the enhancement by fucoidan-6-AH. PMID- 11112096 TI - Studies on the in vitro hepatic microsomal formation of amides during the metabolism of certain secondary and tertiary benzylic amines. AB - Part of our interest during the last few years has been to investigate the possible intermediate(s) and mechanism(s) involved in the formation of amides from N-benzylic amines. A number of benzylic amines with different aryl and alkyl moieties introduced onto the constituent nitrogen were prepared, thus creating a wide variety of secondary, tertiary and heterocyclic benzylic amines with different logP and pKa characteristics (Tables I & II). In some experiments, the possible intermediates of this reaction, i.e. nitrones (Table III), imines (Table IV) and amides themselves (Table V), were used as substrates in our metabolic studies. Their in vitro hepatic microsomal metabolism was studied in order to obtain a structure/metabolic activity relationship for the formation of amides from benzylic amines. This communication reviews these studies and reports our conclusions as to the mechanism of formation of amides from N-benzylic amines. PMID- 11112097 TI - Simultaneous multiorgan presence of human herpesvirus 8 and restricted lymphotropism of Epstein-Barr virus DNA sequences in a human immunodeficiency virus-negative immunodeficient infant. AB - Because a profound dysregulation of the immune system occurs in primary immunodeficiencies, viral infections are not uncommon. Human herpesvirus (HHV)-8 DNA was detected by polymerase chain reaction (PCR) analysis, Southern blotting, and in situ hybridization (ISH) in peripheral blood mononuclear cells and lymphoid organs (bone marrow, spleen, and lymph nodes) and endothelial and epithelial cells and macrophages from several organs (skin, lung, esophagus, intestine, choroid plexus [but not in brain or cerebellum], heart, striated muscle, liver, and kidney) of a human immunodeficiency virus-negative infant with DiGeorge anomaly who died of disseminated infection. Epstein-Barr virus DNA sequences were detected in the spleen and lymph nodes (by PCR and ISH) and in bone marrow (only by ISH) but not in blood or nonlymphoid organs. This report is believed to be the first of multiorgan dissemination of HHV-8 in a primary immunodeficiency. PMID- 11112098 TI - Current status of granulocyte (neutrophil) transfusion therapy for infectious diseases. AB - The transfusion of neutrophils, or granulocyte transfusion therapy, has long been considered as a logical approach to the treatment of severe bacterial and fungal infections in patients with prolonged neutropenia or intrinsic defects in neutrophil function. However, despite numerous clinical trials, the efficacy and safety of granulocyte transfusion therapy remain controversial. Efficacy has been compromised largely by the inability to transfuse sufficient quantities of functionally active neutrophils to patients. The recent use of recombinant granulocyte colony-stimulating factor (G-CSF) to mobilize neutrophils in donors before centrifugation leukapheresis has rekindled interest in the potential clinical applications of granulocyte transfusion therapy. This review focuses on the use of G-CSF for donor stimulation and summarizes the current status of granulocyte transfusion therapy for treatment of infectious diseases. PMID- 11112099 TI - Targeting tuberculosis prevention. PMID- 11112100 TI - Fibrinogen, stroke, and obstructive sleep apnea: an evolving paradigm of cardiovascular risk. PMID- 11112101 TI - Hypertensive pulmonary vascular disease: dawn of the age of prevention? PMID- 11112102 TI - Lung-protective ventilation in acute respiratory distress syndrome: protection by reduced lung stress or by therapeutic hypercapnia? PMID- 11112103 TI - Talc should be used for pleurodesis. PMID- 11112104 TI - Talc should not be used for pleurodesis. PMID- 11112106 TI - Rebuttal from dr. light PMID- 11112105 TI - Rebuttal from dr. sahn PMID- 11112107 TI - A being breathing thoughtful breaths. PMID- 11112108 TI - Legal aspects of withholding and withdrawing life support from critically ill patients in the United States and providing palliative care to them. PMID- 11112110 TI - Fibrinogen levels and obstructive sleep apnea in ischemic stroke. AB - The plasma level of fibrinogen is felt to be an independent risk factor for vascular events. Obstructive sleep apnea (OSA) has a high prevalence in patients with stroke and may also be an independent risk factor. The aim of our study was to determine the association between OSA and plasma levels of fibrinogen in patients with stroke. Polysomnography was performed during neurological rehabilitation in 113 patients (82 men, 31 women, age 58 +/- 11.1 yr, mean +/- SD) with ischemic stroke. OSA was absent (RDI < 5) in 44 patients, 42 had mild OSA (5 < or = RDI < 20), and 27 had moderate to severe OSA (RDI > or = 20). Parameters of OSA (respiratory disturbance index [RDI], oxygen indices) were correlated to plasma levels of fibrinogen, measured in the morning after admission to rehabilitation. Fibrinogen was positively correlated with RDI (r = 0.24, p = 0.007), duration of the longest apnea (r = 0.18, p = 0.049), and negatively correlated with several oxygen indices including average minimal oxygen saturation (r = -0.41, p < 0.001). Correlation coefficients were slightly higher when excluding patients with stroke of presumed cardiac origin. Multiple linear regression identified minimal mean oxygen saturation and sex as independent predictors of fibrinogen level. The correlation between severity of coexisting OSA and fibrinogen level in patients with stroke suggests a possible pathophysiological mechanism for an increased risk of stroke in patients with OSA. PMID- 11112109 TI - Outcomes of contact investigations of infectious tuberculosis patients. AB - The objective of this study was to describe outcomes of tuberculosis (TB) contact investigations, factors correlated with those outcomes, and current successes and ways to improve TB contact investigations. We abstracted clinic records of a representative U.S. urban sample of 1,080 pulmonary, sputum-smear(+) TB patients reported to CDC July 1996 through June 1997 and the cohort of their 6,225 close contacts. We found a median of four close contacts per patient. Fewer contacts were identified for homeless patients. A visit to the patient's residence resulted in two additional (especially child) contacts identified. Eighty-eight percent of eligible contacts received tuberculin skin tests (TSTs). Recording the last exposure date to the infectious patient facilitated follow-up TST provision. Thirty-six percent of contacts were TST(+). Household contacts and contacts to highly smear(+) or cavitary TB patients were most likely to be TST(+). Seventy four percent of TST(+) contacts started treatment for latent TB infection (LTBI), of whom 56% completed. Sites using public health nurses (PHNs) started more high risk TST(-) contacts on presumptive treatment for LTBI. Using directly observed treatment (DOT) increased the likelihood of treatment completion. We documented outcomes of contact investigation efforts by urban TB programs. We identified several successful practices, as well as suggestions for improvements, that will help TB programs target policies and procedures to enhance contact investigation effectiveness. PMID- 11112111 TI - Exhaled nitric oxide in patients with asthma: association with NOS1 genotype. AB - An increased concentration of nitric oxide (NO) in exhaled air (FENO) is now recognized as a critical component of the asthmatic phenotype. When we identified patients with asthma on the basis of a standard case definition alone, we found that they were remarkably heterogeneous with respect to their FENO. However, when we included genotype at a prominent asthma candidate gene (i.e., NOS1) in the case definition, and determined the number of AAT repeats in intron 20, we identified a remarkably homogeneous cohort of patients with respect to FENO. Both mean FENO (p = 0.00008) and variability around the mean (p = 0.000002) were significantly lower in asthmatic individuals with a high number (> or = 12) of AAT repeats at this locus than in those with fewer repeats. These data provide a biologically tenable link between genotype at a candidate gene in a region of linkage, NOS1, and an important component of the asthmatic phenotype, FENO. We show that addition of NOS1 genotype to the case definition of asthma allows the identification of a uniform cohort of patients, with respect to FENO, that would have been indistinguishable by other physiologic criteria. Our isolation of this homogeneous cohort of patients ties together the well-established associations among asthma, increased concentrations of NO in the exhaled air of asthmatic individuals, and variations of trinucleotide repeat sequences as identified in several neurologic conditions. PMID- 11112112 TI - Effects of specific immunotherapy in allergic rhinitic individuals with bronchial hyperresponsiveness. AB - Allergic rhinitis can be associated with bronchial hyperresponsiveness (BHR), and carries an increased risk for the development of asthma. The aim of this study was to evaluate the ability of specific immunotherapy (SIT) to reduce the progression of allergic rhinitis to asthma and prevent the associated increase in BHR. Forty-four subjects monosensitized to Dermatophagoides pteronyssinus, with perennial rhinitis and BHR to methacholine, were randomly assigned to receive SIT or placebo in a double-blind study conducted over a period of 2 yr. After 1 yr of treatment, a 2.88-fold increase in the provocative dose of methacholine producing a 20% decrease in FEV(1) (PD(20)FEV(1)) was recorded in the SIT-treated group (95% confidence interval [CI]: 3.98- to 2.09-fold; p < 0.001), with a further increase to fourfold at the end of Year 2 (95% CI: 2.9- to 5.7-fold; p < 0.001). At the end of the study, the methacholine PD(20)FEV(1) was within the normal range in 50% of treated subjects (p < 0.0001), and was significantly higher in this group than in the group receiving placebo (p < 0.0001). In contrast, no changes in methacholine PD(20)FEV(1) were found in the placebo group throughout the study. Although 9% of subjects given placebo developed asthma, none of those treated with SIT did. This study suggests that SIT, when administered to carefully selected, monosensitized patients with perennial allergic rhinitis, reduces airway responsiveness in subjects with rhinitis, and may be an appropriate prophylactic treatment for rhinitic patients with hyperreactive airways. PMID- 11112113 TI - Therapeutic ratio of inhaled corticosteroids in adult asthma. A dose-range comparison between fluticasone propionate and budesonide, measuring their effect on bronchial hyperresponsiveness and adrenal cortex function. AB - Inhaled corticosteroids have become the mainstay treatment of bronchial asthma. However, simultaneous evaluations of efficacy and side effects are few. This study aimed to compare the relative effect of fluticasone propionate (FP) and budesonide (BUD) on bronchial responsiveness and endogenous cortisol secretion in adults with asthma. The study was double-blind and included 66 adults with asthma, who were randomized to FP (n = 33) or BUD (n = 33). Prestudy, all participants were clinically stable, using inhaled corticosteroids and hyperresponsive to methacholine. Eligible patients were randomized to three consecutive 2-wk periods with either FP 250 microg twice daily, FP 500 microg twice daily, and FP 1,000 microg twice daily, or BUD 400 microg twice daily, BUD 800 microg twice daily, and BUD 1,600 microg twice daily, delivered by Diskhaler and Turbuhaler, respectively. Before randomization and at the end of each treatment, bronchial methacholine PD(20), 24-h urinary cortisol excretion (24-h UC), plasma cortisol, serum osteocalcin, and blood eosinophils were determined. The relative PD(20) potency between FP and BUD was 2.51 (95% CI, 1.05-5.99; p < 0. 05), while the relative 24-h UC potency was 0.60 (95% CI, 0.44-0.83; p < 0.01). The differential therapeutic ratio (FP/BUD) based on PD(20) potency and 24 h UC was 4.18 (95% CI, 1.16-15.03; p < 0.05). The difference in systemic potency was also seen for plasma cortisol, serum osteocalcin, and blood eosinophils. Therapeutic ratio over a wide dose range, determined by impact on bronchial responsiveness and endogenous corticosteroid production, seems to favor FP. PMID- 11112114 TI - Occupational asthma in adults in six Canadian communities. AB - We examined the prevalence, population attributable risk (PAR), and clinical characteristics of occupational asthma (OA) in a randomly selected population in six communities in Canada. Our study followed the European Community Respiratory Health Survey protocol. A randomly selected population of 18,701 (87% response rate) persons from the study communities, ranging in age from 20 to 44 yr, completed an initial questionnaire, of whom 2,974 (39% response rate) attended the laboratory and completed supplementary questionnaires. Of these latter individuals, 383 had asthma. Asthma was defined as physician-diagnosed asthma, and adult-onset asthma was defined as a first attack at age 15 yr or older. We used several methods for estimating OA as follows: (1) reporting of a high-risk job (occupation and industry) for OA at the time of asthma onset (Probable OA); (2) reporting of exposure to a substance that may cause OA (Possible OA) while not in a high-risk job at the time of asthma onset; and (3) combination of the PAR for high-risk jobs and exposures. The prevalence (95% confidence interval [CI]) of Probable OA and Possible OA combined was 36.1% (31.3 to 41.0%) among subjects with adult-onset asthma. The occupations most commonly reported in association with OA were nursing in the Probable OA group and clerical and food preparation in the Possible OA group. The clinical characteristics and exposures reported by both groups were similar. The PAR for adult-onset asthma in high-risk jobs and exposures was 18.2%. The assessment of occupation and industry alone, rather than of exposures, may underestimate the contribution of occupational exposures to asthma prevalence. PMID- 11112115 TI - Community-acquired bacterial pneumonia in human immunodeficiency virus-infected patients: validation of severity criteria. The Grupo Andaluz para el Estudio de las Enfermedades Infecciosas. AB - Severity criteria for community-acquired pneumonia (CAP) have always excluded patients with human immunodeficiency virus (HIV) infection. A 1-yr, multicenter, prospective observational study of HIV-infected patients with bacterial CAP was done to validate the criteria used in the American Thoracic Society (ATS) guidelines for CAP, and to determine the prognosis-associated factors in the HIV infected population with bacterial CAP. Overall, 355 cases were included, with an attributable mortality of 9.3%. Patients who met the ATS criteria had a longer hospital stay (p = 0.01), longer duration of fever (p < 0.001), and higher attributable mortality (13.1% versus 3.5%, p = 0.02) than those who did not. Three factors were independently related to mortality: CD4(+) cell count < 100/microl, radiologic progression of disease, and shock. Pleural effusion, cavities, and/or multilobar infiltrates at admission were independently associated with radiologic progression. A prognostic rule based on the five criteria of shock, CD4(+) cell count < 100/microl, pleural effusion, cavities, and multilobar infiltrates had a high negative predictive value for mortality (97.1%). The attributable mortality for severe pneumonia was 11.3%, as compared with 1.3% for nonsevere disease (p = 0.008). The ATS severity criteria are valid in HIV-infected patients with bacterial CAP. Our study provides the basis for identification of patients who may require hospitalization determined by clinical judgment and the five clinical criteria of shock, a CD4(+) cell count < 100/microl, pleural effusion, cavities, and multilobar involvement. These prognostic factors should be validated in independent cohort studies. PMID- 11112116 TI - Short-term supplementation therapy does not affect elastin degradation in severe alpha(1)-antitrypsin deficiency. The American-Italian AATD Study Group. AB - We evaluated the ability of intravenous supplementation therapy with alpha(1) antitrypsin (AAT) to reduce the rate of urinary excretion of desmosine (DES), a specific marker of elastin degradation, in eight men and four women with emphysema due to severe, congenital deficiency of AAT (range 17-69 mg/dl). Nine were former cigarette smokers, two were current smokers, and one reported never smoking; their mean age was 54 (SD 12) yr and their mean FEV(1) was 41 (18%) of predicted. Urinary DES was measured by isotope dilution and HPLC. Prior to the start of AAT supplementation, mean DES excretion was 13.0 (5.0) microg/g creatinine, 73% higher than in healthy nonsmokers. During 8 wk of supplementation therapy, mean urinary DES excretion was 13.0 (5.9) microg/g creatinine, unchanged from the baseline period (p = 0.85 by repeated measures ANOVA). We conclude that baseline levels of elastin degradation in emphysematous patients with severe AAT deficiency were abnormally high and that 8 wk of AAT supplementation therapy did not appreciably reduce the rate of elastin degradation. These findings raise the possibilities that protective levels of AAT in the lungs are insufficient or that elastin degradation in the lungs of these subjects is not dependent upon neutrophil elastase at this time. PMID- 11112117 TI - Pulmonary blood flow distribution in stage 1 chronic obstructive pulmonary disease. AB - We investigated the hypothesis that lung blood flow distribution is modified in stage 1 chronic obstructive pulmonary disease (COPD). We compared patients with stage 1 COPD (n = 11) with restrictive patients with comparable blood gases (n = 7), to patients with low cardiac index with normal lungs (n = 11) and to control subjects (n = 11). Distribution of transit time (DTT) was computed by deconvolution from first pass radioactivity curves (albumin (99m)Tc) reconstructed from right and left ventricular regions of interest. Distribution descriptors, mean transit time (p < 0.05), standard deviation (p < 0.001), relative dispersion (p < 0.001), and kurtosis (p < 0.001) differed between groups (ANOVA). Cardiac index was the same in COPD and low CI groups but lower compared with normal subjects (p < 0.05). After normalization for cardiac output, the DTT of patients with COPD remained different from low CI and restrictive patients (p < 0.001). Therefore changes in DTT in patients with COPD compared with patients without COPD could not be explained on the basis of difference in cardiac output. Because P(O(2)), PC(O(2)), and pH were similar in COPD and restrictive groups, difference in distribution could not be explained either on the basis of blood gas data. We conclude that changes in DTT occurs in stage 1 COPD and cannot be explained by hypoxemia, hypercapnia, or acidosis alone but must relate to other structural or regulatory responses. PMID- 11112118 TI - Comparison of cost-effectiveness of tuberculosis screening of close contacts and foreign-born populations. AB - Although tuberculosis (TB) screening of immigrants has been conducted for over 50 yr in many industrialized countries, its cost- effectiveness has never been evaluated. We prospectively compared the yield and cost-effectiveness of two immigrant TB screening programs, using close-contact investigation and passive case detection. Study subjects included all immigration applicants undergoing radiographic screening, already arrived immigrants requiring surveillance for inactive TB, and close contacts of active cases resident in Montreal, Quebec, Canada, who were referred from June 1996 to June 1997 to the Montreal Chest Institute (MCI), a referral center specializing in respiratory diseases. For all subjects seen, demographic data, investigations, diagnoses, and therapy were abstracted from administrative data bases and medical charts. Estimated costs of detecting and treating each prevalent active case and preventing future active cases, based on federal and provincial health reimbursement schedules, were compared with the costs for passively diagnosed cases of active TB. Over a period of 1 yr, the three programs detected 27 cases of prevalent active TB and prevented 14 future cases. As compared with passive case detection, close-contact investigation resulted in net savings of $815 for each prevalent active case detected and treated and of $2,186 for each future active case prevented. The incremental cost to treat each case of prevalent active TB was $39,409 for applicant screening and $24,225 for surveillance, and the cost of preventing each case was $33,275 for applicants and $65,126 for surveillance. Close-contact investigation was highly cost effective and resulted in net savings. Immigrant applicant screening and surveillance programs had a significant impact but were much less cost effective, in large part because of substantial operational problems. PMID- 11112119 TI - Influence of ambient fungal spores on emergency visits for asthma to a regional children's hospital. AB - The impact of ambient aeroallergens on morbidity from childhood asthma is largely unknown. To address this issue, we studied the association between daily emergency department visits for asthma to a children's hospital, and daily concentrations of both pollen grains and fungal spores during a 5-yr period between 1993 and 1997. Air pollution and meteorological data accounted for in the analyses included ozone, nitrogen dioxide, sulfur dioxide, sulfates, temperature, barometric pressure, and relative humidity. The daily number of asthma visits ranged from 0 to 36 per day with an average of 7.5. Fungal spores, but not pollen grains, were associated with visits (p < 0.05). The percentage increase associated with each group, independent of the others, was 1.9% (SE 0.9) for deuteromycetes, 4.1% (1.6) for basidiomycetes, 2.8% (1.0) for ascomycetes, and 8.8% for these spores combined. In summary, fungal spores account for a significant proportion of the asthma exacerbations in children that prompt an emergency department visit. PMID- 11112120 TI - Sleep fragmentation, awake blood pressure, and sleep-disordered breathing in a population-based study. AB - Arousal from sleep produces transient increases in systemic blood pressure, leading to the suggestion that repeated arousals are associated with a sustained increase in daytime blood pressure. Using data from the Wisconsin Sleep Cohort Study, a population-based study, we tested the hypothesis that sleep fragmentation is associated with elevated awake blood pressure. Sleep, breathing, and seated blood pressure measurements from 1,021 participants (age 42 +/- 8 yr; 590 males) were analyzed. Sleep fragmentation was defined as the total number of awakenings and shifts to Stage 1 sleep divided by the total sleep time (sleep fragmentation index: SFI). To reduce the confounding influence of sleep disordered breathing, which is related to both increased daytime blood pressure and sleep fragmentation, all participants with an apnea-hypopnea index (AHI) > or = 1 were analyzed separately. Accounting for the influences of sex, age, body mass index, and antihypertensive medication use, the SFI was significantly associated with higher levels of awake systolic blood pressure in people with an AHI < 1; a 2 standard deviation increase in the SFI was associated with a 3.1 mm Hg rise in awake systolic blood pressure. In participants with an AHI > or = 1, there was no independent association between the SFI and awake blood pressure after controlling for the influence of the AHI. PMID- 11112122 TI - Diagnostic value of arterial blood gas measurement in suspected pulmonary embolism. AB - Pulmonary embolism (PE) is a common and lethal yet treatable condition. Several authors have reported on the diagnostic value of combinations of arterial blood gas (ABG) and other clinical data (i. e., prediction rules), and have claimed that these combinations can be safely used to exclude PE. The purpose of this investigation was to evaluate the diagnostic value of ABG measurement and to attempt to validate the ABG prediction rules published by these various authors for the assessment of patients with suspected PE. Two hundred ninety-three consecutive patients referred for imaging to investigate suspected PE were approached to participate in the investigation. ABG and other clinical data were obtained from consenting and eligible patients before an outcome classification (PE versus non-PE) was performed. None of the ABG data or prediction rules had sufficient negative predictive value, specificity, or likelihood ratios to be useful in the management of patients with suspected PE. We conclude that ABG data alone or in combination with other clinical data are not useful in the assessment of suspected PE. PMID- 11112121 TI - Effects of in utero and environmental tobacco smoke exposure on lung function in boys and girls with and without asthma. AB - To investigate whether the effects of in utero exposure to maternal smoking and environmental tobacco smoke (ETS) exposure on lung function vary by sex or asthma status, we examined medical history and tobacco smoke exposure data for 5,263 participants in the Children's Health Study. At study enrollment, parents or guardians of each subject completed a questionnaire, and lung function was measured spirometrically with maximum forced expiratory flow-volume maneuvers. To assess the in utero effects of maternal smoking and ETS exposure on lung function, we used regression splines that accounted for the nonlinear relationship between pulmonary function, height, and age. In utero exposure to maternal smoking was independently associated with deficits in lung function that were larger for children with asthma. Boys and girls with a history of in utero exposure to maternal smoking showed deficits in maximum midexpiratory flow (MMEF) and a decrease in the FEV(1)/FVC ratio. As compared with children without asthma, boys with asthma had significantly larger deficits from in utero exposure in FVC, MMEF, and FEV(1)/FVC, and girls with asthma had larger decreases in FEV(1)/FVC. The effect of ETS exposure varied by children's gender and asthma status. Deficits in flows associated with current ETS exposure were present in children with and without asthma but were significant only among children without asthma. Past ETS exposure was associated with reduced FEV(1), MMEF, and FEV(1)/FVC among boys with asthma. In contrast, past ETS exposure was associated with decreased flow rates in girls without asthma. In summary, both in utero exposure to maternal smoking and ETS exposure were associated with persistent deficits in lung function. The effects of in utero exposure were greatest among children with asthma. PMID- 11112123 TI - Tidal volumes for ventilated infants should be determined with a pneumotachometer placed at the endotracheal tube. AB - Many ventilators measure expired tidal volume (VT) without compensation either for the compliance of the ventilator circuit or for variations in the circuit setup. We hypothesized that the exhaled VT measured with a conventional ventilator at the expiratory valve would differ significantly from the exhaled VT measured with a pneumotachometer placed at the endotracheal tube. To investigate this we studied 98 infants and children requiring conventional ventilation. We used linear regression analysis to compare the VT obtained with the pneumotachometer with the ventilator-measured volume. An additional comparison was made between the pneumotachometer volume and a calculated effective VT. For infant circuits (n = 70), our analysis revealed a poor correlation between the expiratory VT measured with the pneumotachometer and the ventilator-measured volume (r(2) = 0.54). Similarly, the expiratory VT measured with the pneumotachometer did not correlate with the calculated effective volume (r(2) = 0.58). For pediatric circuits (n = 28), there was improved correlation between the expiratory VT measured with the pneumotachometer and both the ventilator measured volume and the calculated effective VT (r(2) = 0.84 and r(2) = 0.85, respectively). The data demonstrate a significant discrepancy between expiratory VT measured at a ventilator and that measured with a pneumotachometer placed at the endotracheal tube in infants. Correcting for the compliance of the ventilator circuit by calculating the effective VT did not alter this discrepancy. In conventionally ventilated infants, exhaled VT should be determined with a pneumotachometer placed at the airway. PMID- 11112124 TI - Delayed distension of contracted airways with lung inflation in vivo. AB - A deep inspiratory sigh is one of the most severe dynamic stresses that lungs normally experience. It typically is a very transient phenomenon, normally lasting only about 2 to 3 s. The airway response to a deep inspiration has been shown to be different in asthmatic and normal individuals. When airway smooth muscle (ASM) is contracted in normal subjects, a deep inspiration results in a subsequent dilation of the airways. However, in asthmatic subjects, a deep inspiration often results in little change in airway function, and sometimes results in an even further contraction of ASM. The mechanism underlying this difference depends on the dynamic behavior of both ASM and the lung parenchyma. If the contracted muscle had slower dynamic responses than the lung parenchyma, the timing of the deep inspiratory maneuver could affect the airway response. In the present study, we designed an experiment to determine how well matched the dynamic response is of airways to that of the lung parenchyma. The results clearly demonstrate that airways contracted with methacholine dilate at about a rate four times slower than that of the lung parenchyma during rapid lung inflation and deflation. This effect may play a role in the unique response of asthmatic subjects to deep inspiration. The mechanism of this dynamic slowness of contracted airways probably involves intrinsic properties of the smooth-muscle contractile processes. PMID- 11112125 TI - Polymorphism of the beta(2)-adrenergic receptor gene and desensitization in human airway smooth muscle. AB - We examined the influence of two common polymorphic forms of the beta(2) adrenergic receptor (beta(2)AR): the Gly16 and Glu27 alleles, on acute and long term beta(2)AR desensitization in human airway smooth muscle (HASM) cells. In cells from 15 individuals, considered without respect to genotype, pretreatment with Isoproterenol (ISO) at 10(-7) M for 1 h or 24 h caused approximately 25% and 64% decreases in the ability of subsequent ISO (10(-6) M) stimulation to reduce HASM cell stiffness as measured by magnetic twisting cytometry. Similar results were obtained with ISO-induced cyclic adenosine monophosphate (cAMP) as the outcome indicator. Data were then stratified post hoc by genotype. Cells containing at least one Glu27 allele (equivalent to presence of the Gly16Glu27 haplotype) showed significantly greater acute desensitization than did cells with no Glu27 allele, whether ISO-induced cell stiffness (34% versus 19%, p < 0.03) or cAMP formation (58% versus 11%, p < 0.02) was measured. Likewise, cells with any Glu27 allele showed greater long-term desensitization of cell stiffness and cAMP formation responses than did cells without the Glu27 allele. The distribution of genotypes limited direct conclusions about the influence of the Gly16 allele. However, presence of the Gly16Gln27 haplotype was associated with less acute and long-term desensitization of ISO-induced cAMP formation than was seen in cells without the Gly16Gln27 haplotype (14% versus 47%, p < 0.09 for short-term desensitization; 32% versus 84%, p < 0.01 for long-term desensitization), suggesting that the influence of Glu27 is not through its association with Gly16. The Glu27 allele was in strong linkage disequilibrium with the Arg19 allele, a polymorphic form of the beta(2)AR upstream peptide of the 5'-leader cistron of the beta(2)AR, and this polymorphism in the beta(2)AR 5'-flanking region may explain the effects of the Glu27 allele. Cells with any Arg19 allele showed significantly greater acute and long-term desensitization of ISO-induced cAMP formation than did cells without the Arg19 allele (54% versus 2%, p < 0.01 for short-term desensitization; 73% versus 35%, p < 0.05 for long-term desensitization). Similar results were obtained for ISO-induced changes in cell stiffness. Thus, the presence of the Glu27 allele is associated with increased acute and long-term desensitization in HASM. PMID- 11112126 TI - Compliance is nonlinear over tidal volume irrespective of positive end-expiratory pressure level in surfactant-depleted piglets. AB - Between the lower and the upper inflection point of a quasistatic pressure-volume (PV) curve, a segment usually appears in which the PV relationship is steep and linear (i.e., compliance is high, with maximal volume change per pressure change, and is constant). Traditionally it is assumed that when positive end-expiratory pressure (PEEP) and tidal volume (V T) are titrated such that the end-inspiratory volume is positioned at this linear segment of the PV curve, compliance is constant over VT during ongoing ventilation. The validity of this assumption was addressed in this study. In 14 surfactant-deficient piglets, PEEP was increased from 3 cm H(2)O to 24 cm H(2)O, and the compliance associated with 10 consecutive volume increments up to full VT was determined with a modified multiple-occlusion method at the different PEEP levels. With PEEP at approximately the lower inflection point, compliance was minimal in most lungs and decreased markedly over VT, indicating overdistension. Compliance both increased and decreased within the same breath at intermediate PEEP levels. It is concluded that a PEEP that results in constant compliance over the full VT range is difficult to find, and cannot be derived from conventional respiratory-mechanical analyses; nor does this PEEP level coincide with maximal gas exchange. PMID- 11112127 TI - Design strategies for longitudinal spirometry studies: study duration and measurement frequency. AB - Measuring the longitudinal change in FEV(1) is useful for assessing the adverse effects of respiratory exposures and pulmonary diseases. Investigators seek to estimate the "true" mean FEV(1) slope (mu(beta)) of an infinite population. The difference between the estimated mean FEV(1) slope (mu(beta)) and the true mean slope, resulting from biological variation and measurement errors, can be minimized by increasing the number of subjects (N), years of follow-up (D), or the frequency of measurements (P). We defined maximum error e(max) such that P[|mu(beta) - mu(beta)| < or = e(max)] = 0.95, and thus e(max) is one-half the width of the 95% confidence interval for mu(beta). We computed the values of e(max) on the basis of actual data obtained from 160 coal miners and working nonminers who had completed 11 spirometry measurements, using recommended equipment and procedures, at 6-mo intervals over 5 yr. Individual 5-yr FEV(1) slopes (Delta FEV(1)) were calculated by linear regression. For a range of values of N, D, and P, tables are provided for e(max), the magnitude of detectable differences in Delta FEV(1) between two groups, and the recommended number of subjects needed in each of two groups to reliably detect the anticipated differences in Delta FEV(1). The tables provide unique guidance for investigators in selecting among various study design options. PMID- 11112128 TI - The effects of inhaled budesonide on circulating eosinophil progenitors and their expression of cytokines after allergen challenge in subjects with atopic asthma. AB - Allergen inhalation by dual responder subjects with atopic asthma is associated with an increase in circulating eosinophil/basophil colony-forming units (Eo/B CFU) and granulocyte-macrophage colony- stimulating factor (GM-CSF) immunolocalization in Eo/B colony cells grown in vitro. The current study examined the effect of the inhaled corticosteroid, budesonide, on the number of allergen- induced circulating eosinophils and Eo/B CFU, and immunolocalization of GM-CSF and interleukin-5 (IL-5) in Eo/B colony cells grown in vitro. Sixteen subjects with mild atopic asthma were treated for either 7 or 8 d with 200 microg inhaled budesonide or placebo twice a day. Peripheral blood was collected before and 24 h after allergen inhalation challenge and nonadherent mononuclear cells (NAMC) were grown in methylcellulose culture. Eo/B CFU were enumerated after 14 d in culture, and prepared on slides for immunocytochemistry. Budesonide attenuated the allergen-induced increase in circulating eosinophils (4.0 +/- 0.4 x 10(5)/ml versus 6.5 +/- 0.7 x 10(5)/ml, p = 0.0001), circulating Eo/B CFU (12.4 +/- 2.3/10(6) NAMC versus 18.8 +/- 4.6/10(6) NAMC, p = 0.05), and immunolocalization of GM-CSF in Eo/B colony cells (11.8 +/- 1.9% positive versus 18.0 +/- 2.2%, p = 0.01) but not immunolocalization of IL-5 (7.9 +/- 1.4% versus 4.5 +/- 0.6%, p > 0.05). Inhaled budesonide attenuated the number of allergen-induced circulating eosinophils and their progenitors grown in the presence of GM-CSF, which may partially be a result of regulating eosinophil progenitor expression of the autocrine growth factor GM-CSF. PMID- 11112129 TI - The production of extracellular matrix proteins by human passively sensitized airway smooth-muscle cells in culture: the effect of beclomethasone. AB - Airway remodeling is a key feature of persistent asthma. Part of the remodeling process involves the laying down of extracellular matrix (ECM) proteins within the airways. In this study we compared the production of ECM proteins by human airway smooth-muscle (ASM) cells in culture after exposure to 10% serum from an asthmatic individual or 10% serum from a nonasthmatic individual with or without beclomethasone (0.01 to 100 nM). Enzyme-linked immunosorbent assays were done with antibodies to human fibronectin; perlecan; elastin; the laminin beta(1), gamma(1), beta(2), alpha(1) chains; thrombospondin; chondroitin sulfate; collagen types I, III, IV, and V; versican; and decorin. Serum from the asthmatic individual, when compared with that from the nonasthmatic individual, caused a significant increase in the production of fibronectin, perlecan, laminin gamma(1), and chondroitin sulfate. Beclomethasone caused a significant reduction in the number of cells exposed to serum from either the asthmatic or nonasthmatic individual, but did not reverse the increase in ECM protein induced by the former. These results suggest an interaction between the ASM and the allergic process that may alter components of the airway wall in asthma, and that corticosteroids may not prevent the fibrosis induced by resident cells within the airways. PMID- 11112130 TI - Gender-related differences in severe, early-onset chronic obstructive pulmonary disease. AB - Men have higher prevalence rates of chronic obstructive pulmonary disease (COPD) than women, which has been attributed to the historically higher rates of cigarette smoking in males. However, the increased rates of cigarette smoking in females within the last several decades have been associated with steadily increasing rates of COPD in women. As part of a study of the genetics of severe, early-onset COPD, we assembled a group of 84 probands with severe, early-onset COPD (without severe alpha(1)-antitrypsin deficiency) and 348 of their first degree relatives. We found a markedly elevated prevalence (71.4%) of females among the early-onset COPD probands. Among the entire group of first-degree relatives of early-onset COPD probands, univariate analysis demonstrated similar spirometric values and bronchodilator responsiveness in males and females; however, among current or ex-smokers, female first-degree relatives had significantly lower FEV(1)/ FVC (81.4 +/- 17.2% in females versus 87.0 +/- 12.9% in males, p = 0.009) and significantly greater bronchodilator responsiveness (expressed as percentage of baseline FEV(1)) (7.7 +/- 9.4% pred in females versus 4.1 +/- 6.4% pred in males, p = 0.002). Female smoking first-degree relatives were significantly more likely to demonstrate profound reductions in FEV(1) (< 40% pred) than male smoking first-degree relatives (p = 0. 03). Multivariate analysis, performed with generalized estimating equations, demonstrated that current or ex-smoking female first-degree relatives had significantly greater risk of FEV(1) < 80% pred (OR 1.91, 95% CI 1.03- 3.54), FEV(1) < 40% pred (OR 3.56, 95% CI 1.08-11.71), and bronchodilator response greater than 10% of baseline FEV(1) (OR 4.74, 95% CI 1.91-11.75). These results suggest that women may be more susceptible to the development of severe COPD. PMID- 11112131 TI - Effect of free radical scavengers on diaphragmatic contractility in septic peritonitis. AB - We investigated the effects of polyethylene glycol-adsorbed superoxide dismutase (PEG-SOD), polyethylene glycol-adsorbed catalase (PEG-CAT), and DMSO on diaphragmatic contractility and malondialdehyde (MDA) concentrations in septic peritonitis in vitro. One hundred eighty-six rats were divided into two groups. One group (CLP group) was treated with cecal ligation and perforation (CLP), and the other (sham group) was treated with laparotomy. PEG-SOD, PEG-CAT, and DMSO were administered intraperitoneally 30 min before and 12 h after CLP. The left hemidiaphragm was removed at 10 h or 16 h after the operation. We assessed the diaphragmatic contractility by twitch characteristics and force-frequency curves in vitro. We measured MDA concentrations, as an index of oxygen-derived free radical-mediated lipid peroxidation, and the activities of two main antioxidant enzymes, superoxide dismutase (SOD) and glutathione peroxidase (GPx), as an index of antioxidant defenses, after CLP. Diaphragmatic force generation capacity was significantly reduced after CLP. Diaphragmatic MDA levels were significantly elevated after CLP. PEG-SOD, PEG-CAT, and DMSO significantly improved diaphragmatic contractility and prevented the elevation in diaphragmatic MDA concentrations after CLP. Diaphragmatic SOD activities were significantly increased after CLP. These results suggest that several types of oxygen-derived free radicals play a role in the reduction in diaphragmatic contractility after CLP. PMID- 11112132 TI - Circulating nitric oxide is suppressed in obstructive sleep apnea and is reversed by nasal continuous positive airway pressure. AB - Epidemiological studies have implicated obstructive sleep apnea (OSA) as an independent comorbid factor in cardiovascular and cerebrovascular diseases. The recurrent episodes of occlusion of upper airways during sleep result in pathophysiological changes that may predispose to vascular diseases, and we postulate that nitric oxide may be one of the mediators involved. This study investigates the levels of circulating nitric oxide (NO), measured as serum nitrites and nitrates, in the early morning in OSA subjects compared with control subjects, and the effect of overnight nasal continuous positive airway pressure (nCPAP) in OSA subjects. Thirty men with moderate to severe OSA (age = 41.9 +/- 9.0; apnea-hypopnea index, AHI = 48.0 +/- 18.1) were compared with 40 healthy men (age = 40.6 +/- 5.4; AHI = 1.4 +/- 1.2). Serum nitrite/nitrate levels were significantly lower in OSA subjects (OSA = 38.9 +/- 22.9 microM, control subjects = 63.1 +/- 47.5 microM, p = 0.015). There was significant negative correlation between serum nitrites/nitrates and the following parameters: AHI (r = -0.389, p = 0.001), oxygen desaturation time (r = -0.346, p = 0.004), and systolic blood pressure (BP) (r = -0.335, p = 0.005). Stepwise multiple linear regression with systolic or diastolic BP as the dependent variable identified serum nitrites/nitrates as the only significant correlate. Twenty-two OSA subjects had measurements of serum NO at baseline and after an overnight application nCPAP. There was significant increase in serum NO after nCPAP (baseline = 30.5 +/- 14.4 microM, after nCPAP = 81.0 +/- 82.1 microM, p = 0.01). This study demonstrates, for the first time, that circulating NO is suppressed in OSA, and this is promptly reversible with the use of nCPAP. The findings offer support for nitric oxide being one of the mediators involved in the acute hemodynamic regulation and long-term vascular remodeling in OSA. PMID- 11112133 TI - Airway nitric oxide levels in cystic fibrosis patients are related to a polymorphism in the neuronal nitric oxide synthase gene. AB - Patients with cystic fibrosis (CF) have decreased concentrations of expired nitric oxide (FENO) as compared with healthy individuals. A number of factors, including viscous mucus as a diffusion barrier for airway NO, consumption of NO by bacterial enzymes, and decreased NO production have been hypothesized to account for these low levels of FENO. We examined the relationship between the size of an AAT repeat polymorphism in intron 20 of the NOS1 gene and FENO in 75 patients with CF. Mean FENO was significantly (p = 0.027) lower in CF patients who harbored two alleles with a high number of repeats (>/= 12) than in those who harbored alleles with fewer repeats at this locus (4.0 +/- 0.8 [mean +/- SEM] ppb versus 6.4 +/- 0.9 ppb). Colonization of the airways with Pseudomonas aeruginosa was significantly (p = 0.0358) more common in CF patients with high numbers of AAT repeats in the NOS1 gene. Significant differences between NOS1 genotypes were also observed among patients homozygous for the cystic fibrosis transmembrane regulator delta F508 mutation for FENO (2.3 +/- 0.4 ppb versus 5.3 +/- 0.7 ppb, p = 0.0006), and this was also true for colonization of the airways with P. aeruginosa (p = 0.0147) and Aspergillus fumigatus (p = 0.0221). These data provide evidence that the NOS1 gene is not only associated with the variability of FENO, but also with P. aeruginosa colonization of airways in CF patients. PMID- 11112134 TI - Association between allergy and asthma from childhood to middle adulthood in an Australian cohort study. AB - A cohort of 378 asthmatic children was studied from 7 to 35 yr of age at 7-yr intervals. On selection for inclusion in the study sample, the children had a wide range of severity of wheezing. At each 7-yr review, asthma severity, the presence of eczema or hay fever, and skin test reactivity to house dust mite or rye grass were recorded by questionnaire or clinical interview. We report on the course of asthma and these atopic conditions over the study period and discuss associations between the two phenomena. The presence of an atopic condition in childhood was found to increase the odds of more severe asthma in later life (odds ratio [OR] = 1.66, 95% confidence interval [CI]: 1.17 to 2.36 in the case of eczema; OR = 1. 39, 95% CI: 1.00 to 1.92 for hay fever; and OR = 2.25, 95% CI: 1.49 to 3.39 for skin test reactivity). Additionally, the odds of eczema and hay fever in later life increased with severity of asthma in childhood. The findings of this study provide substantially new quantitative information on the extent of association between asthma and atopic conditions from childhood into middle adulthood. PMID- 11112135 TI - Post-lung transplant bronchiolitis obliterans syndrome (BOS) is characterized by increased exhaled nitric oxide levels and epithelial inducible nitric oxide synthase. AB - In conditions characterized by airway inflammation, exhaled nitric oxide (eNO) levels are increased. Post-lung transplant bronchiolitis obliterans syndrome (BOS) is characterized by airway inflammation and development of progressive airway narrowing and fibrosis. We have previously shown that in stable lung transplant recipients (LTR), mean eNO levels were not elevated but were still related to the degree of airway neutrophilia within the group. The hypothesis now tested is that in BOS, eNO levels are increased in association with even greater airway neutrophilia and enhanced expression of inducible (iNOS) nitric oxide synthase in the bronchial epithelium. We determined eNO levels in 40 LTR in four groups: well and "stable": LTR (n = 20), BOS (n = 8), bacterial airway infection (BI, n = 6), and acute rejection (AR, n = 6). Following bronchoscopic sampling, we performed a quantitative assessment of iNOS and constitutive nitric oxide synthase (cNOS) expression in endobronchial biopsies by immunohistochemistry. Mean +/- SEM eNO levels in BOS and BI were significantly higher than in stable LTR (20 +/- 1.2 parts per billion [ppb] and 24.7 +/- 1.7 ppb versus 12.5 +/- 0.9 ppb; p < 0.01 for both). In AR, eNO levels (13.4 ppb +/- 0.5) were not different in stable LTR (p = 0.34). When compared with stable LTR, there was increased expression of iNOS in the bronchial epithelium and generally in the lamina propria (LP) in patients with BOS and BI. In AR, iNOS expression was increased but only in the LP in a perivascular distribution. Expression of cNOS was reduced in BOS but not in BI and AR compared with the stable group. Using regression analysis, only iNOS expression in the bronchial epithelium (r(2) = 0.77; p < 0.0001) and %BAL neutrophils (r(2) = 0. 79; p < 0.0001) were positively related to eNO in stable LTR and BOS. We conclude that epithelial iNOS appears to be the major source of eNO. Exhaled NO levels also appear to reflect the degree of airway neutrophilia in both stable LTR and BOS groups. This suggests that serial eNO measurements may be able to predict the early development of BOS. PMID- 11112136 TI - Evidence for genetic associations between asthma, atopy, and bronchial hyperresponsiveness: a study of 8- to 18-yr-old twins. AB - We measured asthma in the last 12 mo, diagnosed by a respiratory physician at interview; atopy, defined by a positive skin prick test to any of eight common allergens; and bronchial hyperresponsiveness (BHR) to hypertonic saline, in 381 twin pairs aged 8 to 18 yr selected from the Australian Twin Registry-183 monozygous (MZ) and 198 dizygous (DZ). The associations between twins, as measured by an odds ratio, were greater in MZ pairs compared with DZ pairs for asthma: 25.6 (95% confidence interval 11.3- 57.8) versus 1.9 (1.0-3. 5); atopy: 14.6 (7.1-30.1) versus 2.5 (1.4- 4.5); and BHR: 14.1 (6. 4-31.0) versus 4.2 (2.1 8.6) (all p < 0.002). The associations between each pair of traits within an individual were slightly greater than the association between one trait in a twin and the other trait in the cotwin (cross-trait cross-pair) in MZ pairs. Further, the associations in MZ pairs were greater than in DZ pairs (p < 0.05). Under the assumptions of the classic twin model, these data suggest that the strong cross sectional associations between these three traits are due to an overlap between the genetic factors involved in each of these three traits. PMID- 11112137 TI - Peripheral and central ventilatory responses in central sleep apnea with and without congestive heart failure. AB - Given that the apnea-ventilation cycle length during central sleep apnea (CSA) with congestive heart failure (CHF) is approximately 70 s, we hypothesized that rapidly responsive peripheral CO(2) ventilatory responses would be raised in CHF CSA and would correlate with the severity of CSA. Sleep studies and single breath and rebreathe hypercapnic ventilatory responses (HCVR) were measured as markers of peripheral and central CO(2) ventilatory responses, respectively, in 51 subjects: 12 CHF with no apnea (CHF-N), 8 CHF with obstructive sleep apnea (CHF OSA), 12 CHF-CSA, 11 CSA without CHF ("idiopathic" CSA; ICSA), and 8 normal subjects. Single breath HCVR was equally elevated in CHF-CSA and ICSA groups compared with CHF-N, CHF-OSA, and normal groups (0.58 +/- 0.09 [mean +/- SE] and 0. 58 +/- 0.07 versus 0.23 +/- 0.06, 0.25 +/- 0.04, and 0.27 +/- 0.02 L/min/PET(CO(2)) mm Hg, respectively, p < 0.001). Similarly, rebreathe HCVR was elevated in both CHF-CSA and ICSA groups compared with CHF-N, CHF-OSA, and normal groups (5.80 +/- 1.12 and 3.53 +/- 0. 29 versus 2.00 +/- 0.25, 1.44 +/- 0.16, and 2.14 +/- 0.22 L/min/PET(CO(2)) mm Hg, respectively, p < 0.001). Furthermore, in the entire CHF group, single breath HCVR correlated with central apnea-hypopnea index (AHI) (r = 0.63, p < 0.001) and percentage central/total apneas (r = 0.52, p = 0.022). Rebreathe HCVR correlated with awake Pa(CO(2)) (r = -0.61, p < 0.001), but not with central AHI or percentage central/total apneas independent of its relationship with single breath HCVR. In conclusion, in subjects with CHF, raised central CO(2) ventilatory response predisposes to CSA promoting background hypocapnia and exposing the apnea threshold to fluctuations in ventilation, whereas raised and faster-acting peripheral CO(2) ventilatory response determines the periodicity and severity of CSA. PMID- 11112138 TI - Noninvasive assessment of inspiratory muscle function during exercise. AB - The use of esophageal and gastric balloons limits measurement of the tension-time index of inspiratory muscles (TTI) during exercise. The aim of this study was to assess whether a noninvasive tension-time index, TT(0.1), given by P(0.1)/PI(max) x TI/Ttot (where P(0.1) is mouth occlusion pressure, PI(max) is maximal inspiratory pressure, and TI/Ttot is duty cycle) could reliably assess TTI during exercise. In seven healthy young men and nine patients with COPD we measured TT(0.1) and TTI (i.e., Pes/Pes(max) x TI/Ttot where Pes is mean esophageal pressure and Pes(max) is maximal static Pes) at rest and during an incremental exercise test. A significant linear correlation (p < 0.02) was found between TT(0.1) and TTI in all normal subjects and patients with COPD. An equation for estimating TTI from TT(0.1) was established for each group. In the normal subjects there was good agreement between estimated and observed data. In five additional normal males studied prospectively, the agreement was also satisfactory and reproducible. In the COPD patients the agreement was poor. In conclusion, in young healthy subjects the changes in TT(0.1) during exercise reflect the changes in TTI, allowing satisfactory estimation of TTI from noninvasive measurements of TT(0.1). PMID- 11112139 TI - Predictive value of pulmonary function parameters for sleep apnea syndrome. AB - Nocturnal polysomnography is the standard diagnostic test for sleep apnea syndrome (SAS) but is both expensive and time-consuming. We developed a predictive index for SAS based on pulmonary function data, including respiratory resistance determined by the forced oscillation technique, from 168 obese snorers with suspected SAS. Our model used logistic regression to obtain case-by-case predictions of the probability of SAS, defined as an apnea-hypopnea index (AHI) > or = 15 during overnight polysomnography. We then tested our model in a prospective group of 101 similar patients. Specific respiratory conductance and daytime oxygen saturation contributed significantly to the model. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the index computed from these parameters were 98%, 86%, 90%, and 97%, respectively. In the prospective group, the model proved repeatable, with 100% sensitivity, 84% specificity, 86% PPV, and 100% NPV. The high NPV may help to identify obese snorers with a SAS risk that is so low as to make polysomnography unnecessary. Based on the 50% prevalence of SAS in our study and on the fact that polysomnography is required in all patients with daytime somnolence, we calculated that using our model would have obviated the need for polysomnography in 38% of our patients. PMID- 11112140 TI - The prognostic significance of the histologic pattern of interstitial pneumonia in patients presenting with the clinical entity of cryptogenic fibrosing alveolitis. AB - Lone cryptogenic fibrosing alveolitis (CFA) is a progressive interstitial lung disease, with a median survival of 3 to 6 yr from the onset of dyspnea. CFA can be subdivided into prognostically significant histopathologic patterns, including nonspecific interstitial pneumonia (NSIP). We reviewed 78 patients with a clinicopathologic diagnosis of CFA, biopsied between 1978 and 1989, to evaluate the prevalence and prognostic significance of these histopathologic patterns, in particular NSIP. Biopsy appearances were reclassified by two pulmonary histopathologists as usual interstitial pneumonia (UIP) (47%), NSIP (36%), or desquamative interstitial pneumonia (DIP)/respiratory bronchiolitis-associated interstitial lung disease (RBILD) (17%). The kappa coefficient of agreement between pathologists was 0.49. In 67 cases, follow-up was complete to death or 10 yr after biopsy, with 50 deaths during a median follow-up of 42 mo (UIP, 89%; NSIP, 61%, DIP/RBILD, 0%). Survival was highest in DIP/RBILD and higher in NSIP than UIP, p < 0.0005. When analysis was confined to patients with UIP or NSIP, the mortality of UIP remained higher, p < 0.01, but the 5-yr survival in patients with fibrotic NSIP was only 45%, indicating that this histologic appearance is often associated with a poor outcome. A response to treatment was more frequent in DIP/RBILD than in NSIP (p < 0.01) or UIP (p < 0.0005). This study confirms the prognostic value of subclassifying patients with CFA according to histopathologic pattern. However, in patients with clinically typical CFA, a histologic diagnosis of fibrotic NSIP needs to be interpreted with caution and does not necessarily denote a good outcome. PMID- 11112141 TI - Reduced nasal nitric oxide in diffuse panbronchiolitis. AB - Diffuse panbronchiolitis (DPB) is a pulmonary disease of unknown origin with inflammation in the respiratory bronchioles, bronchiectasis, and recurrent sinusitis. Patients with DPB suffer from chronic airway infections resulting from mucociliary dysfunction. Whereas a high concentration of nasal nitric oxide (NO) has been documented in healthy subjects, only two diseases are known to reduce nasal NO: primary ciliary dyskinesia syndrome and cystic fibrosis. We hypothesized that patients with DPB have abnormal levels of nasal NO. To test our hypothesis, we measured NO with the chemiluminescence technique. Air was sampled directly from the nose in 15 healthy subjects and eight patients with DPB. Nasal NO was 88% lower in DPB patients than in the age-matched control subjects (69 +/- 70 versus 556 +/- 87 nl/min; p < 0.001). Treatment with erythromycin for 2 wk did not alter the nasal NO in four control subjects. DPB is the third pulmonary disease in which nasal NO is low. The reduced nasal NO may well be involved in the pathogenesis of DPB, and NO measurements may serve as a noninvasive test in the diagnosis of DPB. PMID- 11112142 TI - Eosinophilic granulocytes and interleukin-6 level in bronchoalveolar lavage fluid are associated with the development of obliterative bronchiolitis after lung transplantation. AB - In a prospective cohort study, we assessed whether changes in total cell counts and differentiation and interleukin-6 (IL-6), IL-8, and monocyte chemoattractant protein-1 (MCP-1) concentrations in bronchoalveolar lavage fluid (BALF) are associated with a higher risk to develop obliterative bronchiolitis (OB). We investigated 60 lung transplant patients (follow-up of 2 to 8 yr) with either histologic evidence of OB within 1 yr after lung transplantation (n = 19) or no pathology, good outcome (GO) for at least 24 mo and well-preserved lung function, i.e., FEV > or = 80% of baseline (n = 41). Median time between lung transplantation and the first BAL was 42 d for the GO group and 41 d for the OB group (p > 0.05). In the bronchial fraction, median total cell counts (0.06 x 10(3)/ml versus 0.04 x 10(3)/ml), lymphocyte (9 x 10(3)/ml versus 2 x 10(3)/ml), and eosinophilic granulocyte counts (1 x 10(3)/ml versus 0) were significantly higher in the OB group than in the GO group (p < 0.05). In the alveolar fraction, this was the case for the median value of neutrophilic granulocyte counts (19 x 10(3)/ml versus 4 x 10(3)/ml), respectively. Median values of IL-6 and IL-8 concentrations in both bronchial (IL-6: 23 versus 6 pg/ml, IL-8: 744 versus 102 pg/ml) and alveolar fractions (IL-6: 13 versus 3 pg/ml, IL-8: 110 versus 30 pg/ml) of the BALF were significantly higher in the OB group than in the GO group. By means of logistic regression, we showed that higher total cell, neutrophilic granulocyte, and lymphocyte counts, the presence of eosinophilic granulocytes, and higher concentrations of IL-6 and IL-8 were significantly associated with an increased risk to develop OB. We conclude that monitoring cell counts, neutrophilic and eosinophilic granulocytes, IL-6, and IL-8 in BALF within 2 mo after lung transplantation in addition to the transbronchial lung biopsy (TBB) pathology will contribute to a better identification and management of the group of patients at risk for developing OB within a year. PMID- 11112143 TI - Relationship of upper and lower airway cytokines to outcome of experimental rhinovirus infection. AB - To test the hypothesis that rhinovirus (RV)-induced immune responses influence the outcome of RV infections, we inoculated 22 subjects with allergic rhinitis or asthma with RV16. Nasal secretions and induced sputum were repeatedly sampled over the next 14 d. RV16 infection increased nasal granulocyte colony-stimulating factor (G-CSF) and interleukin (IL)-8, which was accompanied by neutrophilia in blood and nasal secretions. Nasal G-CSF correlated closely with increased blood neutrophils (r(s) = 0.69, p < 0.005), whereas nasal neutrophils correlated with both G-CSF (r(s) = 0.87, p < 0.001) and IL-8 (r(s) = 0.75, p < 0.001). Although similar relationships were present in sputum, changes in sputum neutrophils and G CSF with RV16 infection were relatively modest. In addition, virus-induced changes in the sputum interferon-gamma-to-IL-5 messenger RNA ratio were inversely related to both peak cold symptoms (r(s) = -0.60, p < 0.005) and the time to viral clearance (undetectable picornavirus RNA). These results indicate that airway IL-8 and G-CSF are closely associated with virus-induced neutrophilic inflammation during an experimental RV infection in atopic volunteers. In addition, the balance of airway T-helper cell type 1 (Th1)- and Th2-like cytokines induced by RV infection may help determine the clinical outcome of common cold infections, raising the possibility that the individual subject's immune response, rather than atopic status per se, is important in this regard. PMID- 11112144 TI - Diagnosing acute pulmonary embolism: effect of chronic obstructive pulmonary disease on the performance of D-dimer testing, ventilation/perfusion scintigraphy, spiral computed tomographic angiography, and conventional angiography. ANTELOPE Study Group. Advances in New Technologies Evaluating the Localization of Pulmonary Embolism. AB - In patients with chronic obstructive pulmonary disease (COPD), differentiating a pulmonary embolism (PE) from an exacerbation of COPD can be difficult, since clinical signs and symptoms of the two conditions overlap. Development of reliable noninvasive or minimally invasive techniques for the diagnosis of PE is, especially in these patients, necessary. In this study we assessed the effect of COPD on the accuracy of the clinical probability estimate (CPE), spiral computed tomographic angiography (SCTA), D-dimer analysis, ventilation perfusion (V/Q) scintigraphy, and pulmonary angiography for the diagnosis of PE. From May 1997 through March 1998, 627 consecutive patients with suspected PE were investigated in six teaching hospitals. In these patients, D-dimer testing, CPE, V/Q scintigraphy, and SCTA and/or pulmonary angiography were performed according to a strict diagnostic protocol. The patients were also independently categorized as having COPD or not. A diagnosis of COPD was established in 91 patients (15%). The prevalence of PE was similar in patients with and without COPD (29% and 31%, respectively), notwithstanding the larger proportion of nondiagnostic V/Q scan results in patients with COPD (46% versus 21%, p < 0.001). The distribution of CPEs, diagnostic value of the D-dimer assay and SCTA, and reproducibility of pulmonary angiography were comparable among patients with and without COPD. The presence of COPD does not affect the diagnostic performance of CPE, D-dimer testing, SCTA, or pulmonary angiography. Furthermore, although more nondiagnostic V/Q scan results can be expected in the presence of COPD, V/Q scintigraphy remains a valuable screening test in patients with COPD. PMID- 11112145 TI - Comparison of sputum induction with fiberoptic bronchoscopy in the diagnosis of tuberculosis: experience at an acquired immune deficiency syndrome reference center in Rio de Janeiro, Brazil. AB - Many patients with suspected pulmonary tuberculosis (PTB) do not produce sputum spontaneously or are smear-negative for acid-fast bacilli (AFB). We prospectively compared the yield of sputum induction (SI) and fiberoptic bronchoscopy with bronchoalveolar lavage (BAL) for the diagnosis of PTB in a region with a high prevalence of tuberculosis and human immunodeficiency virus (HIV) infection. Fifty seven percent (143 of 251) of patients had diagnoses of PTB, of whom 17% (25 of 143) were HIV seropositive. There were no significant differences in the yields of AFB smears or cultures whether obtained via SI or BAL. Among 207 HIV seronegative patients, the AFB smear and mycobacterial culture results from specimens obtained by SI and BAL were in agreement in 97% (202 of 207) (kappa test = 0.92) and 90% (186 of 207) (kappa test = 0.78), respectively. Among HIV seropositive patients the agreements between AFB smear and culture results for SI and BAL specimens were 98% (43 of 44) (kappa test = 0.93) and 86% (38 of 44) (kappa test = 0.69), respectively. We conclude that SI is a safe procedure with a high diagnostic yield and high agreement with the results of fiberoptic bronchoscopy for the diagnosis of PTB in both HIV-seronegative and HIV seropositive patients. PMID- 11112146 TI - Multiple combination bactericidal antibiotic testing for patients with cystic fibrosis infected with multiresistant strains of Pseudomonas aeruginosa. AB - We developed a rapid in vitro antibiotic susceptibility test to screen double- and triple-antibiotic combinations for bactericidal activity against 75 multiresistant Pseudomonas aeruginosa isolates referred from 44 cystic fibrosis (CF) patients. When used alone, the most effective intravenous antibiotic, meropenem, was bactericidal against only 44% of the isolates. High-dose tobramycin (200 microg/ml; concentrations achievable by aerosol administration) was bactericidal against 72% of isolates. Adding a second antibiotic significantly improved bactericidal activity. The most effective double antibiotic combinations contained high-dose tobramycin plus meropenem, piperacillin/tazobactam, or ciprofloxacin, and were bactericidal against 88 to 94% of the isolates. Excluding high-dose tobramycin, the most effective intravenous double-antibiotic combinations contained meropenem plus ciprofloxacin, tobramycin (4 microg/ml), or cefipime, and were bactericidal against 85%, 71%, and 70% of isolates, respectively. Adding a third antibiotic did not significantly improve inhibition in vitro. We conclude that double antibiotic combinations containing meropenem or high-dose tobramycin show the best bactericidal activity in vitro against multiresistant strains of P. aeruginosa. Addition of a third antibiotic to these double-antibiotic combinations may be unnecessary. PMID- 11112147 TI - Diagnosis of metabolic acid-base disturbances in critically ill patients. AB - We compare two commonly used diagnostic approaches, one relying on plasma bicarbonate concentration and "anion gap," the other on "base excess," with a third method based on physicochemical principles, for their value in detecting complex metabolic acid-base disturbances. We analyzed arterial blood samples from 152 patients and nine normal subjects for pH, PCO(2), and concentrations of plasma electrolytes and proteins. Ninety-six percent of the patients had serum albumin concentration < or = 3 SD below the mean of the control subjects. In about one-sixth of the patients, base excess and plasma bicarbonate were normal. In a great majority of these apparently normal samples, the third method detected simultaneous presence of acidifying and alkalinizing disturbances, many of them grave. The almost ubiquitous hypoalbuminemia confounded the interpretation of acid-base data when the customary approaches were applied. Base excess missed serious acid-base abnormalities in about one-sixth of the patients; this method fails when the plasma concentrations of the nonbicarbonate buffers (mainly albumin) are abnormal. Anion gap detected a hidden "gap acidosis" in only 31% of those samples with normal plasma bicarbonate in which such acidosis was diagnosed by the third method; when adjusted for hypoalbuminemia, it reliably detected the hidden abnormal anions. The proposed third method identifies and quantifies individual components of complex acid-base abnormalities and provides insights in their pathogenesis. PMID- 11112148 TI - Triptolide attenuates pulmonary arterial hypertension and neointimal formation in rats. AB - This paper reports the effect of triptolide (a diterpenoid triepoxide) on the development of monocrotaline (MCT)-induced pulmonary hypertension in pneumonectomized rats. Male Sprague- Dawley rats were injected with MCT (60 mg/kg) on Day 7 after left pneumonectomy. Rats received therapy from Day 5 to 35 with triptolide (0.25 mg/kg intraperitoneally, every other day, n = 10), or vehicle (0.1 ml of ethanol/cremophor intraperitoneally, every other day, n = 10). By Day 35, triptolide-treated rats demonstrated lower mean pulmonary arterial pressure (mPAP) than vehicle-treated rats (mPAP 21 +/- 3 versus 42 +/- 5 mm Hg, p < 0.001). Triptolide-treated rats also had significantly less right ventricular hypertrophy (RVH) and pulmonary arterial neointimal formation. In a rescue experiment, rats initiated therapy on Day 21. At Day 35, vehicle-treated rats (n = 4) had higher mPAP (40 +/- 9 mm Hg), greater RVH, and more severe pulmonary arterial neointimal formation than rats that received triptolide (0.25 mg/kg every other day, n = 7, mPAP 30 +/- 4 mm Hg) and rats that received triptolide (0.2 mg/kg daily, n = 7, mPAP 25 +/- 5 mm Hg, p < 0.01). In pneumonectomized rats that receive MCT, triptolide attenuates the development of pulmonary hypertension and RVH, and promotes regression of pulmonary arterial neointimal formation. PMID- 11112149 TI - Fibroblast contractility: usual interstitial pneumonia and nonspecific interstitial pneumonia. AB - The aim of this study was to compare the function of lung fibroblasts obtained from surgically biopsied specimens of patients with idiopathic pulmonary fibrosis/usual interstitial pneumonia (UIP; n = 5), nonspecific interstitial pneumonia (NSIP; n = 5), and normal parts of surgically resected lungs (control; n = 5). The results showed that (1) fibroblasts obtained from UIP showed increased contractility compared with those obtained from NSIP or controls (UIP, 72.7 +/- 6.21%; NSIP, 32.8 +/- 5.46; controls, 28.5 +/- 3.51, p < 0.01 in UIP versus NSIP or control); (2) this increase in contractility was consistent with enhanced F-actin content in fibroblasts; (3) conditioned media from UIP fibroblast cultures enhanced control fibroblast contractility, whereas those obtained from NSIP or controls did not; (4) the 180 and 25 kD products representing the contractility in conditioned media were identified as fibronectin (ED-A domain) and TGF-beta1 by immunoblots, respectively; (5) the UIP conditioned media contained higher amounts of fibronectin or TGF-beta 1 (fibronectin: UIP 289 +/- 47.1 ng/ml, NSIP 121 +/- 23.0, control 118 +/- 16.0; TGF-beta1: UIP 798 +/- 119 pg/ml, NSIP 246 +/- 69.1, control 247 +/- 53.6, p < 0.01 in UIP versus NSIP or control); () the contractility positively correlated with the amount of either fibronectin (r = 0.867, p < 0.001, n = 15) or TGF-beta 1 (r = 0.939, p < 0.001, n = 15), respectively. Thus, UIP fibroblasts showed greater contractility than did NSIP fibroblasts and up-regulated control fibroblasts. PMID- 11112150 TI - Optimization of aerosol deposition by pressure support in children with cystic fibrosis: an experimental and clinical study. AB - Nebulized aerosols are commonly used to deliver drugs into the lungs of patients with cystic fibrosis (CF). The aim of this study was to assess the effectiveness of pressure-support (PS) ventilation in increasing aerosol deposition within the lungs of children with CF. An in vitro study demonstrated the feasibility of coupling a breath-actuated nebulizer to a PS device. An in vivo study was done with 18 children (ages 6 to 21 yr) with clinically stable CF, each of whom underwent both a standard and a PS-driven ventilation scan (control session and PS session, respectively). In addition, a perfusion scan was used to determine lung outlines and to construct a geometric model for quantifying aerosol deposition by radioactivity counting in MBq. Homogeneity of nebulization was evaluated from the four first-order moments of aerosol distribution in the peripheral and central lung regions. The time-activity nebulization curve was linear in all patients, with higher slopes during the PS than during the control session (0.43 +/- 0.07 [mean +/- SD] MBq/min and 0.32 +/- 0.23 MBq/min, respectively; p < 0.018). Quantitatively, aerosol deposition was about 30% greater after the PS session (4.4 +/- 2.7 MBq) than after the control session (3.4 +/- 2.1 MBq; p < 0.05). Similarly, deposition efficacy (as a percentage of nebulizer output) was significantly better during the PS session than during the control session (15.3 +/- 8.3% versus 11.5 +/- 5.7%, p < 0.05). No differences in the regional deposition pattern or in homogeneity of uptake were observed. In conclusion, our data show that driving the delivery of a nebulized aerosol by noninvasive PS ventilation enhances total lung aerosol deposition without increasing particle impaction in the proximal airways. PMID- 11112151 TI - Effect of endogenous and exogenous prostaglandin E(2) on interleukin-1 beta induced cyclooxygenase-2 expression in human airway smooth-muscle cells. AB - We studied the effect of endogenous and exogenous prostaglandin E(2) (PGE(2)), a metabolite of arachidonic acid through the cyclooxygenase (COX) pathway, on interleukin (IL)-1 beta-induced COX-2 expression, using primary cultures of human bronchial smooth-muscle cells (HBSMC). Treatment with exogenous PGE(2) resulted in enhanced expression of IL-1 beta-induced COX-2 protein and messenger RNA (mRNA) as compared with the effect of the cytokine per se. Inhibition of PGE(2) production with a nonselective COX inhibitor (flurbiprofen, 10 microM) resulted in a significant reduction in IL-1 beta- induced COX-2 expression, supporting a role of endogenous COX metabolites in the modulation of COX-2 expression. None of the experimental conditions used in the study affected the expression of constitutive cyclooxygenase (COX-1). Treatment with cycloheximide to inhibit translation, and with dexamethasone or actinomycin D to inhibit transcription, linked the effect of PGE(2) to the transcriptional level of COX-2 mRNA rather than to a potential effect on protein and/or mRNA stabilization. PGE(2) increased adenylate cyclase activity in a concentration dependent manner, and forskolin, a direct activator of adenylate cyclase, caused a marked increase in IL-1 beta dependent COX-2, suggesting the existence of a causal relationship between the two events. The same results were observed with salbutamol, a bronchodilator that acts by increasing cyclic adenosine monophosphate. The effect of PGE(2) on COX-2 expression may contribute to the hypothesized antiinflammatory role of PGE(2) in human airways, providing a self-amplifying loop leading to increased biosynthesis of PGE(2) during an inflammatory event. PMID- 11112152 TI - Halothane reduces the early lipopolysaccharide-induced lung inflammation in mechanically ventilated rats. AB - Several studies suggest that anesthetics modulate the immune response. The aim of this study was to investigate the effect of halothane and thiopental on the lung inflammatory response. Rats submitted or not to intratracheal (IT) instillation of lipopolysaccharides (LPS) were anesthetized with either halothane (0. 5, 1, or 1.5%) or thiopental (60 mg. kg(-1)) and mechanically ventilated for 4 h. Control rats were treated or not by LPS without anesthesia. Lung inflammation was assessed by total and differential cell counts in bronchoalveolar lavage fluids (BALF) and by cytokine measurements (tumor necrosis factor-alpha [TNF-alpha], interleukin-6 [IL-6], macrophage inflammatory protein-2 [MIP-2], and monocyte chemoattractant protein-1 [MCP-1]) in BALF and lung homogenates. In the absence of LPS treatment, neither halothane nor thiopental modified the moderate inflammatory response induced by tracheotomy or mechanical ventilation. Cell recruitment and cytokine concentrations were increased in all groups receiving IT LPS. However, in halothane-anesthetized rats (halothane > or = 1%), but not in thiopental-anesthetized rats, the LPS-induced lung inflammation was altered in a dose-dependent manner. Indeed, when using 1% halothane, polymorphonuclear leukocyte (PMN) recruitment was decreased by 55% (p < 0.001) and TNF-alpha, IL-6, and MIP-2 concentrations in BALF and lung homogenates were decreased by more than 60% (p < 0.001) whereas total protein and MCP-1 concentrations remained unchanged. The decrease of MIP-2 (observed at the protein and messenger RNA [mRNA] level) was strongly correlated to the decrease of PMN recruitment (r = 0.73, p < 0.05). This halothane-reduced lung inflammatory response was transient and was reversed 20 h after the end of the anesthesia. Our study shows that halothane > or = 1%, delivered during 4 h by mechanical ventilation, but not mechanical ventilation per se, alters the early LPS-induced lung inflammation in the rat, suggesting a specific effect of halothane on this response. PMID- 11112153 TI - Therapeutic hypercapnia reduces pulmonary and systemic injury following in vivo lung reperfusion. AB - Permissive hypercapnia, involving tolerance to elevated Pa(CO(2)), is associated with reduced acute lung injury (ALI), thought to result from reduced mechanical stretch, and improved outcome in ARDS. However, deliberately elevating inspired CO(2) concentration alone (therapeutic hypercapnia, TH) protects against ALI in ex vivo models. We investigated whether TH would protect against ALI in an in vivo model of lung ischemia-reperfusion (IR). Anesthetized open chest rabbits were ventilated (standard eucapnic settings), and were randomized to TH (FI(CO(2)) 0.12) versus control (FI(CO(2)) 0.00). Pa(CO(2)) and arterial pH values achieved in the TH versus CON groups were 101 +/- 3 versus 44.4 +/- 4 mm Hg and 7.10 +/- 0.03 versus 7.37 +/- 0.03, respectively. Following left lung ischemia and reperfusion, TH versus control was associated with preservation of lung mechanics, attenuation of protein leakage, reduction in pulmonary edema, and improved oxygenation. Indices of systemic protection included improved acid-base and lactate profile, in the absence of systemic hypoxemia. In the TH group, mean BALF TNF-alpha levels were 3.5% of CON levels (p < 0.01), and mean 8-isoprostane levels were 30% of CON levels (p = 0.02). Western blot analysis demonstrated reduced lung tissue nitrotyrosine in TH, indicating attenuation of tissue nitration. Finally, preliminary data suggest that TH may attenuate apoptosis following lung IR. We conclude that in the current model TH is protective versus IR lung injury and mechanisms of protection include preservation of lung mechanics, attenuation of pulmonary inflammation, and reduction of free radical mediated injury. If these findings are confirmed in additional models, TH may become a candidate for clinical testing in critical care. PMID- 11112154 TI - Inflammation and structural changes in the airways of patients with atopic and nonatopic asthma. BHR Group. AB - The aim of the present study was to compare the cellular pattern and structural changes in the airway walls of atopic and nonatopic patients with asthma. Bronchial biopsy specimens were obtained from 13 atopic subjects with asthma, nine nonatopic patients with asthma, and seven healthy control subjects and investigated using immunohistochemical methods. The number of eosinophils increased in both asthma groups, but significantly more in the atopic group. The number of mast cells increased similarly in the two asthma groups, whereas the number of neutrophils increased only in the nonatopic asthma group. The number of T-lymphocytes (CD3-, CD4-, CD8-, CD-25-positive cells) was higher in patients with atopic asthma compared with nonatopic asthma. Interleukin-4 (IL-4) and IL-5 positive cells were more frequently found in the atopic asthma group, whereas cells staining for IL-8 were more frequent in the nonatopic group. The degree of epithelial damage was significantly higher in the atopic asthma group compared with the control subjects and the nonatopic asthmatics. The tenascin and laminin layer was significantly thicker in the atopic group compared with the group of nonatopic asthmatics. In the atopic group, there was a significant negative correlation between epithelial integrity (defined as the relative length of intact epithelium) and the eosinophil count and also between the number of CD25 positive cells and epithelial integrity. The number of mast cells correlated positively with the thickness of tenascin- and laminin-positive layers. In conclusion, we provide evidence of different patterns of involvement of inflammatory cells in atopic and nonatopic patients with asthma. There were also structural differences in the bronchial mucous membrane between atopic asthma and nonatopic asthma. This suggests that there are differences in the extent of the immunopathologic response of these clinically distinct forms of asthma. PMID- 11112155 TI - Hepatocyte growth factor attenuates collagen accumulation in a murine model of pulmonary fibrosis. AB - We investigated the in vivo effects of recombinant human hepatocyte growth factor (HGF) on epithelial cell proliferation in normal mouse lung and on the repair process that follows bleomycin-induced lung injury. Intratracheal administration of 100 micrograms of rhHGF to C57BL/6 mice led to proliferation of bronchial and alveolar epithelial cells as indicated by an increased number of cells staining for proliferating cell nuclear antigen (PCNA). The effect of HGF on the lung repair process was examined by administration of 100 micrograms of rhHGF on Day 3 and Day 6 after intratracheal injection of bleomycin to mice. We found that HGF significantly attenuated collagen accumulation induced by bleomycin as determined by quantitation of hydroxyproline content and by scoring of the extent of fibrosis. To explore the potential mechanisms involved in the beneficial effects of HGF, we performed in vitro studies with A549 pulmonary epithelial cells and found that HGF enhanced cell surface plasmin generation, expression of u-PA activity, and cell migration. In summary, HGF has potent in vivo and in vitro effects on epithelial cells, which suggests it may have a role in the therapy of pulmonary fibrosis. PMID- 11112156 TI - Muscular contractile failure in septic patients: role of the inducible nitric oxide synthase pathway. AB - Skeletal muscle failure is a frequent manifestation of sepsis that affects prognosis and rehabilitation by impairing respiration and ambulation. Animal studies have shown that the inducible NO synthase (NOS2) is expressed in skeletal muscles during sepsis, likely affecting muscular function, by promoting the formation of the strong oxidant peroxynitrite. In contrast, whether human skeletal muscle expresses a functional NOS2 in similar conditions is unknown. We studied NOS2 expression (mRNA and protein) and activity and its role in contractile function in samples from rectus abdominis muscle obtained during surgical procedure in 16 septic patients and in 21 controls. Peroxynitrite formation was detected by immunohistochemical detection of nitrotyrosine residues. The main results of this study are as follows: (1) A significant increase in NOS2 mRNA, protein, and activity was found in muscles from septic patients, the expression of NOS2 protein positively correlating with sepsis severity. (2) Contractile force was significantly lower in septic than in control muscles. This phenomenon was not reverted by muscle incubation ex vivo with the NOS inhibitor L-NMMA, indicating that NO was not involved in force reduction at the time of biopsy. (3) NOS2 expression in skeletal myocytes was strongly co localized with nitrotyrosine, revealing muscular peroxynitrite generation during the septic process, before the muscle was biopsied. Exposure of control muscles to an amount of peroxynitrite similar to that generated in septic muscles during the septic process resulted in a nonreversible reduction in force generation. These results suggest that NOS2 could be involved in the decreased muscular force of septic patients via the local generation of peroxynitrite. PMID- 11112157 TI - Lung inflammation in hyperoxia can be prevented by antichemokine treatment in newborn rats. AB - Hyperoxia may contribute to lung disease in newborns through effects on alveolar neutrophils which predominate in respiratory distress syndrome and other acute lung injuries. Neutrophil chemokines such as interleukin-8 (IL-8) regulate chemoattraction, and are elevated in tracheal aspirates of newborns who develop bronchopulmonary dysplasia (BPD). Blockade of neutrophil chemokines may reduce hyperoxia-induced inflammatory lung injury and BPD. We therefore tested the hypothesis that hyperoxia contributes to elevations of rat neutrophil chemokines, cytokine-induced neutrophil chemoattractant-1 (CINC-1), and macrophage inflammatory protein-2 (MIP-2) in newborn rat lung. Newborn rats were exposed to air or 95% O(2) for 8 d. CINC-1 and MIP-2 were measured in whole lung homogenates by ELISA. Newborn 95% O(2)-exposed animals were given anti-CINC-1 or anti-MIP-2, 1, 5, or 10 microg on Days 3 and 4 of 95% O(2) exposure. Bronchoalveolar lavage (BAL) was performed after perfusion on day 6 to evaluate airway neutrophils, and myeloperoxidase (MPO) was measured in perfused whole lung. Lungs were examined histologically and immunohistochemically for effects of 95% O(2) +/- antichemokine. CINC-1 and MIP-2 increased nearly tenfold by Day 8 95% O(2) treatment versus air control. CINC-1 and MIP-2 immunolabeling was increased in alveolar macrophages and alveolar epithelium in 95% O(2). Anti-CINC-1 and anti MIP-2 treatment at every dose reduced neutrophil number > 90% in BAL. Anti-CINC-1 10 microg reduced tissue MPO by 50%. Antichemokine treatment on days 3 and 4 prevented alveolar septal thickening and reduced chemokine immunolabeling on Day 6. Hyperoxia-induced neutrophil influx is mediated in part by CINC-1 and MIP-2 in newborn rats and can be partially prevented by treatment with anti-CINC-1 and anti-MIP-2. PMID- 11112158 TI - Epithelial desquamation in asthma: artifact or pathology? AB - To determine whether the denudation of the bronchial epithelium observed in endobronchial biopsies from asthmatic subjects is a true pathologic feature or an artifact of tissue sampling, we analyzed epithelial integrity in bronchial biopsies from 14 subjects with mild and moderate asthma and 12 healthy subjects. In each subject, 4 to 8 bronchial biopsies were taken from large airways during bronchoscopy, fixed in 4% paraformaldehyde, embedded in glycomethacrylate, cut into 2-microM sections, and stained with toluidine blue. A x4 image of each biopsy was copied to a computer file using a video camera, and lines were drawn and measured along the basement membrane underlying areas completely denuded of overlying epithelium, areas covered by a single layer of basal cells, and areas of intact epithelium. We found that the percentage of basement membrane that was denuded of epithelium was similar in the healthy and asthmatic subjects (14.8 +/- 11.8 versus 11.4 +/- 9.8% respectively, p = 0.38); the percentage of basement membrane that was covered by a single layer of basal cells was also similar in the two groups (46.4 +/- 11.0 versus 54.5 +/- 9.8%, respectively, p = 0. 11). In the asthmatic subjects, we found no significant correlation between the percentage of basement membrane covered by denuded epithelium or by a single layer of basal cells and the FEV(1) percentage of predicted or the PC(20) methacholine. We conclude that denudation of bronchial epithelium in endobronchial biopsies from asthmatic subjects with stable mild and moderate disease is an artifact of tissue sampling and is not a true pathologic feature of the disease, and that the extent of airway epithelial denudation is not correlated with the severity of airway narrowing or the severity of bronchial hyperresponsiveness. PMID- 11112159 TI - Identification of human lung and skin proteins conjugated with hexamethylene diisocyanate in vitro and in vivo. AB - Diisocyanates are asthma-causing chemicals used in the commercial production of polyurethane. We have previously shown that human lung epithelial cell proteins can become conjugated with hexamethylene diisocyanate (HDI) and may be biologically important in diisocyanate-induced asthma. The objective of this study was to identify specific human lung and skin proteins that become conjugated with diisocyanate after in vitro and in vivo exposure. Following in vitro exposure of human airway epithelial cells (A549), keratin 18, the 78-kD glucose-regulated protein, trans-1, 2-dihyrobenzene-1,2-diol dehydrogenase, and actin were identified as prominent diisocyanate-conjugated proteins through use of a combination of immunocytochemical and mass spectrometric techniques. Following in vivo inhalation of an HDI aerosol, keratin 18 was also identified as the predominant diisocyanate-conjugated protein in human endobronchial biopsy samples, whereas albumin was the predominant diisocyanate-conjugated protein in bronchoalveolar lavage fluid. Keratin was also identified as a predominant diisocyanate-conjugated protein in human skin biopsy samples after epicutaneous exposure to liquid-phase HDI, although the major skin diisocyanate-conjugated protein (56-kD) differed from the predominant lung diisocyanate-conjugated keratin (47-kD). The data from this study identify keratin and other proteins as potential "carriers" for diisocyanates in vivo, and suggest that HDI conjugation of these proteins may play a role in the pathogenesis of diisocyanate-induced asthma. PMID- 11112160 TI - Assessment of neonatal diaphragm function using magnetic stimulation of the phrenic nerves. AB - A nonvolitional test to assess diaphragm strength in neonates has not been previously described. Our aim was to assess the feasibility of cervical (CMS) and anterior (AMS) magnetic stimulation of the phrenic nerves in neonates. Double circular stimulating coils (90-mm) were used. For CMS, one coil was placed over the cervical spine to bilaterally stimulate the phrenic nerve roots, whereas for AMS the coils were placed on the anterolateral aspect of the neck to allow unilateral and bilateral stimulation. Diaphragm contractility was assessed as transdiaphragmatic pressure (Pdi) measured with balloon catheters positioned in the midesophagus and stomach. Stimulus supramaximality was assessed by examining diaphragm twitch Pdi (TwPdi) across a range of stimulator outputs; 85, 90, 95, and 100% of maximum. Pressure signals were measured by differential pressure transducer and displayed in real time on a computer. Patients were studied supine during sleep. CMS was performed on seven neonates (mean gestational age [GA] 38 wk, range 33 to 40 wk) and AMS on 18 neonates (mean GA 37 wk, range 32 to 41 wk). The mean (SD) TwPdi with CMS was 2.5 (0.8) cm H(2)O. CMS was not supramaximal; reducing the stimulator output below 100% caused marked reductions in TwPdi, also the shape of the pressure waveforms suggested that CMS may not have activated the diaphragm alone. Mean (SD) TwPdi with AMS was 4.5 (1.3) cm H(2)O on the left, 4.1 (0.9) cm H(2)O on the right, and 8.7 (3.9) cm H(2)O for bilateral stimulation. The shape of the pressure waveforms suggested that AMS was more specific and a plateau in TwPdi at higher stimulator outputs indicated supramaximality. We conclude that AMS may provide a useful technique to assess diaphragm function in the neonate. PMID- 11112161 TI - Proceedings of the ATS workshop on refractory asthma: current understanding, recommendations, and unanswered questions. American Thoracic Society. PMID- 11112162 TI - Biogeography and community structure of North American seed-harvester ants. AB - Seed-harvester ants are a dominant and conspicuous insect group throughout arid portions of the southwestern United States and northern Mexico, and they include approximately 75 species. Intense study in the late 1970s and early 1980s led to the paradigm that interspecific competition for limited seed resources is the primary factor that structures seed-harvester ant communities. This review attempts to adjust this paradigm, suggesting that interspecific competition for food is probably less important than previously thought, whereas intraspecific competition is common and strong. Abiotic-habitat factors also have important effects on local species composition and attributes. This review also emphasizes several aspects of seed-harvester ants that have been poorly studied, including historical factors, adaptive radiation, and biogeography, to provide a more detailed evolutionary background for understanding existing species assemblages. PMID- 11112163 TI - Mating behavior and chemical communication in the order Hymenoptera. AB - Insects of the order Hymenoptera are biologically and economically important members of natural and agro ecosystems and exhibit diverse biologies, mating systems, and sex pheromones. We review what is known of their sex pheromone chemistry and function, paying particular emphasis to the Hymenoptera Aculeata (primarily ants, bees, and sphecid and vespid wasps), and provide a framework for the functional classification of their sex pheromones. Sex pheromones often comprise multicomponent blends derived from numerous exocrine tissues, including the cuticle. However, very few sex pheromones have been definitively characterized using bioassays, in part because of the behavioral sophistication of many Aculeata. The relative importance of species isolation versus sexual selection in shaping sex pheromone evolution is still unclear. Many species appear to discriminate among mates at the level of individual or kin/colony, and they use antiaphrodisiacs. Some orchids use hymenopteran sex pheromones to dupe males into performing pseudocopulation, with extreme species specificity. PMID- 11112164 TI - Insect biodemography. AB - Biodemography is an emerging subdiscipline of classical demography that brings life table techniques, mortality models, experimental systems, and comparative methods to bear on questions concerned with the fundamental determinants of mortality, longevity, aging, and life span. It is important to entomology because it provides a secure and comprehensive actuarial foundation for life table and mortality analysis, it suggests new possibilities for the use of model insect systems in the study of aging and mortality dynamics, and it integrates an interdisciplinary perspective on demographic concepts and actuarial techniques into the entomological literature. This paper describes the major life table formulae and mortality models used to analyze the actuarial properties of insects; summarizes the literature on adult insect life span, including a discussion of basic concepts; identifies the major correlates of extended longevity; and suggests new ideas for using demographic concepts in both basic and applied entomology. PMID- 11112165 TI - Predicting St. Louis encephalitis virus epidemics: lessons from recent, and not so recent, outbreaks. AB - St. Louis encephalitis virus was first identified as the cause of human disease in North America after a large urban epidemic in St. Louis, Missouri, during the summer of 1933. Since then, numerous outbreaks of St. Louis encephalitis have occurred throughout the continent. In south Florida, a 1990 epidemic lasted from August 1990 through January 1991 and resulted in 226 clinical cases and 11 deaths in 28 counties. This epidemic severely disrupted normal activities throughout the southern half of the state for 5 months and adversely impacted tourism in the affected region. The accurate forecasting of mosquito-borne arboviral epidemics will help minimize their impact on urban and rural population centers. Epidemic predictability would help focus control efforts and public education about epidemic risks, transmission patterns, and elements of personal protection that reduce the probability of arboviral infection. Research associated with arboviral outbreaks has provided an understanding of the strengths and weaknesses associated with epidemic prediction. The purpose of this paper is to review lessons from past arboviral epidemics and determine how these observations might aid our ability to predict and respond to future outbreaks. PMID- 11112166 TI - Evolution of exclusive paternal care in arthopods. AB - Exclusive male care of offspring is the rarest form of postzygotic parental care among animals and has arisen independently in only 13 arthropod taxa. To distinguish the effects of sexual selection from those of natural selection on the evolution of arthropod paternal care, predictions concerning several life history and behavioral traits resulting from both forms of selection are made and tested across all known taxa with exclusive paternal care. Comparisons suggest parallels between prezygotic nuptial gifts and exclusive postzygotic male care and support the hypothesis that, in arthropods, male behaviors that enhance female reproductive success either directly, by releasing females from the fecundity constraints of maternal care (enhanced fecundity hypothesis), or indirectly, by identifying mates with superior genes (handicap principle), are traits on which sexual selection has acted. Under such conditions, males that are willing to guard young become preferred mates for gravid females and enjoy greater promiscuity than males that are unable or unwilling to guard. Females use nest construction or the act of guarding another female's eggs as honest signals of paternal intent and quality. PMID- 11112167 TI - Mating strategies and spermiogenesis in ixodid ticks. AB - Mate-seeking and sperm-transfer in the ixodid hard ticks, which include important vectors of zoonotic pathogens, generally reflect their peculiarly prolonged pattern of feeding. The metastriate ticks, including Dermacentor, Amblyomma, and Rhipicephalus, invariably attain sexual maturity and mate solely on their hosts. The more primitive prostriate Ixodes ticks, however, may copulate both in the absence of hosts and while the female engorges. These expanded opportunities for insemination complicate the mating systems of the Ixodes ricinus complex of species. In these ticks, autogenous spermatogenesis must precede host contact, whereas anautogenous oogenesis requires that the females store sperm. All hard tick males undergo a courting ritual before they can deposit their spermatophores within the female's genital tract. These diverse and prolonged patterns of sexual interaction provide opportunities for interactions between populations and individuals that may be relevant to the role of ticks as vectors of zoonotic pathogens. PMID- 11112168 TI - Genetic and physical mapping in mosquitoes: molecular approaches. AB - The genetic background of individual mosquito species and populations within those species influences the transmission of mosquito-borne pathogens to humans. Technical advances in contemporary genomics are contributing significantly to the detailed genetic analysis of this mosquito-pathogen interaction as well as all other aspects of mosquito biology, ecology, and evolution. A variety of DNA-based marker types are being used to develop genetic maps for a number of mosquito species. Complex phenotypic traits such as vector competence are being dissected into their discrete genetic components, with the intention of eventually using this information to develop new methods to prevent disease transmission. Both genetic- and physical-mapping techniques are being used to define and compare genome architecture among and within mosquito species. The integration of genetic and physical-map information is providing a sound framework for map-based positional cloning of target genes of interest. This review focuses on advances in genome-based analysis and their specific applications to mosquitoes. PMID- 11112169 TI - Insect acid-base physiology. AB - Acid-base status influences many aspects of insect biology, including insect distributions in aquatic systems, insect-plant and insect-pathogen interactions, membrane transport phenomena, and the mode of action of pesticides. Acid-base status in the hemolymph and gut lumen of insects is generally well regulated but varies somewhat within individuals owing to effects of temperature, activity, discontinuous ventilation, and diet. The pH of the midgut lumen varies with the phylogeny and feeding ecology. Insect fluids have buffer values similar to those of vertebrates. The respiratory system participates in acid-base homeostasis primarily by regulating the internal carbon dioxide (partial) pressure via changes in spiracular opening and convective ventilation. The epithelia of the renal system and gut participate in hemolymph acid-base regulation by varying acid-base transport in response to organismal acid-base status. Evidence to date suggests that the dominant mechanisms for control of renal acid-base excretion involve hormonal regulation of H+-V-ATPase activity. PMID- 11112170 TI - Evolution and behavioral ecology of heteronomous aphelinid parasitoids. AB - In almost all species of parasitic wasps in the Coccophaginae, a subfamily of Aphelinidae, males have host relationships different from females. In these "heteronomous" species, females are generally endoparasitoids of sternorrhynchous Hemiptera, such as scale insects, mealybugs, and whiteflies. In contrast, males may be hyperparasitoids, developing in or on conspecific females or other primary parasitoids. In other species, females are endoparasitoids of whiteflies, and males are primary endoparasitoids of eggs of Lepidoptera. Males and females may both be primary parasitoids on the same species of scale insect hosts, but females develop as endoparasitoids, whereas males are ectoparasitoids. Here we review these life histories, focusing on examples of sexually dimorphic host relationships, development, and morphology. Coccophagine species may be sexual or parthenogenetic; we discuss reproductive modes and the interaction of sex ratio distorters with sex-specific host relationships. Sex allocation in the species in which males are hyperparasitoids involves choices of not what sex egg to lay, but whether to accept or reject a host of a given type; study in this area is reviewed as well as research in kin discrimination and ovicide. Last, we present the current understanding of phylogenetic relationships within this lineage and discuss hypotheses for the evolutionary origin of heteronomy in the Aphelinidae. PMID- 11112171 TI - Species traits and environmental constraints: entomological research and the history of ecological theory. AB - The role that entomology has played in the historical (1800s-1970s) development of ecological theories that match species traits with environmental constraints is reviewed along three lineages originating from the ideas of a minister (Malthus TR. 1798. An Essay on the Principle of Population. London: Johnson) and a chemist (Liebig J. 1840. Die Organische Chemie in ihrer Anwendung auf Agricultur und Physiologie. Braunschweig: Vieweg). Major developments in lineage 1 focus on habitat as a filter for species traits, succession, nonequilibrium and equilibrium conditions, and generalizations about the correlation of traits to environmental constraints. In lineage 2, we trace the evolution of the niche concept and focus on ecophysiological traits, biotic interactions, and environmental conditions. Finally, we describe the conceptual route from early demographic studies of human and animal populations to the r-K concept in lineage 3. In the 1970s, the entomologist Southwood merged these three lineages into the "habitat templet concept" (Southwood TRE. 1977. J. Anim. Ecol. 46:337-65), which has stimulated much subsequent research in entomology and general ecology. We conclude that insects have been a far more important resource for the development of ecological theory than previously acknowledged. PMID- 11112172 TI - Genetic transformation systems in insects. AB - The past 5 years have witnessed the emergence of techniques that permit the stable genetic transformation of a number of non-drosophilid insect species. These transposable-element-based strategies, together with virus-based techniques that allow the expression of genes to be quickly examined in insects, provide insect scientists with a first generation of genetic tools that can begin to be harnessed to further increase our understanding of gene function and regulation in insects. We review and compare the characteristics of these gene transfer systems and conclude that, although significant progress has been made, these systems still do not meet the requirements of robust genetic tools. We also review risk assessment issues arising from the generation and probable release of genetically engineered insects. PMID- 11112173 TI - Tests of reproductive-skew models in social insects. AB - Reproductive-skew theory can be broadly divided into transactional models, in which reproduction is shared among group members in return for some fitness benefit, and tug-of-war models, in which reproductive sharing arises solely from an inability of each group member to fully control the others. For small-colony social insects in which complete reproductive control by a single individual is plausible, transactional-concession models account, better than any other existing model, for observed relationships between each of the dependent variables of skew, changes in reproductive partitioning over time, group size, and within-group aggression, and each of the predictor variables of genetic relatedness, ecological constraints on solitary breeding, and benefits of group living. An extension of transactional-concession models via the "workers-as-a collective-dominant" model potentially offers new insights into some of the most striking reproductive patterns in large-colony eusocial Hymenopteran species, from the loss of worker capacity to produce female offspring to patterns of skew and aggression in polygynous societies. PMID- 11112174 TI - Biology and management of grape phylloxera. AB - Grape phylloxera, Daktulosphaira vitifoliae (Homoptera: Phylloxeridae), is a worldwide pest of grapevines. Its life cycle has sexual and asexual portions with forms that feed from leaf and root galls. Not all forms occur throughout the insect's range. Root forms predominate on Vitis vinifera cultivars; leaf forms predominate on other Vitis species characteristic of the American native range. Other conditions influence expression of the life cycle. Hosts and conditions similarly affect life table performance. Damage to grapevines is by secondary soilborne pathogens attacking the feeding site and by physiological interaction of the insect with the grapevine, though the latter has not been well studied. Resistant rootstocks derived from native American Vitis are the primary control tool. The insect varies genetically and relative to performance on hosts. Use of insecticides is limited in effect, and other control methods are not proven. More research on the biology, ecology, and management of phylloxera is needed. PMID- 11112175 TI - Models of division of labor in social insects. AB - Division of labor is one of the most basic and widely studied aspects of colony behavior in social insects. Studies of division of labor are concerned with the integration of individual worker behavior into colony level task organization and with the question of how regulation of division of labor may contribute to colony efficiency. Here we describe and critique the current models concerned with the proximate causes of division of labor in social insects. The models have identified various proximate mechanisms to explain division of labor, based on both internal and external factors. On the basis of these factors, we suggest a classification of the models. We first describe the different types of models and then review the empirical evidence supporting them. The models to date may be considered preliminary and exploratory; they have advanced our understanding by suggesting possible mechanisms for division of labor and by revealing how individual and colony-level behavior may be related. They suggest specific hypotheses that can be tested by experiment and so may lead to the development of more powerful and integrative explanatory models. PMID- 11112176 TI - Population genomics: genome-wide sampling of insect populations. AB - Modern population genetics underwent a major paradigm shift during the last decade of the 20th century with the discovery that thousands of genes of known function and position in a genome can be analyzed simultaneously in a single individual. The impact of this technology on insect population genetics is potentially profound. Sampling distributions of genetic statistics can now be derived from many individual loci or among many segregating sites within a gene. Inferences regarding random mating, gene flow, effective population sizes, disequilibrium, and relatedness among populations can now be based on patterns of variation at many loci. More importantly, genome-wide sampling enables population geneticists to distinguish effects that act on the whole genome from those that act on individual loci or nucleotides. We introduce the term "population genomics" to describe the process of simultaneous sampling of numerous variable loci within a genome and the inference of locus-specific effects from the sample distributions. The four critical assumptions implicit in the population genomics approach are explained in detail. Studies adopting this paradigm are reviewed, and the steps necessary to complete a population genomics study are outlined. PMID- 11112177 TI - The evolution of color vision in insects. AB - We review the physiological, molecular, and neural mechanisms of insect color vision. Phylogenetic and molecular analyses reveal that the basic bauplan, UV blue-green-trichromacy, appears to date back to the Devonian ancestor of all pterygote insects. There are variations on this theme, however. These concern the number of color receptor types, their differential expression across the retina, and their fine tuning along the wavelength scale. In a few cases (but not in many others), these differences can be linked to visual ecology. Other insects have virtually identical sets of color receptors despite strong differences in lifestyle. Instead of the adaptionism that has dominated visual ecology in the past, we propose that chance evolutionary processes, history, and constraints should be considered. In addition to phylogenetic analyses designed to explore these factors, we suggest quantifying variance between individuals and populations and using fitness measurements to test the adaptive value of traits identified in insect color vision systems. PMID- 11112178 TI - Methods for marking insects: current techniques and future prospects. AB - Tracking the movement of insects in their natural habitat is essential for understanding their basic biology, demography, and ethology. A wide variety of markers have been used to assess insect population dynamics, dispersal, territoriality, feeding behavior, trophic-level interactions, and other ecological interactions. The ideal marker should persist without inhibiting the insect's "normal" biology. Furthermore, the marker should be environmentally safe, cost-effective, and easy to use. In this article, we review the current state of knowledge regarding insect marking, document the advantages and limitations of each marking technique, and discuss advances made in marking insects over the past decade. PMID- 11112179 TI - Resistance of Drosophila to toxins. AB - Insects, including Drosophila, readily respond to toxins such as phytotoxins, metal ions, and insecticides in their environment by evolving resistance. Although Drosophila are seldom targets for insecticides, nevertheless populations worldwide have evolved resistance to a variety of insecticides, and these resistance alleles persist in high frequency. In many cases, Drosophila use the same genetic and biochemical mechanisms that underlie resistance in pest insects, including single-site changes in target molecules resulting from point mutations and upregulation of degradative enzymes, particularly cytochrome P450 enzymes and glutathione S-transferases. However, several types of resistance found in pest insects, such as gene amplification and knock-down resistance, have not been reported in Drosophila field populations. Excellent Drosophila-plant models are being studied to understand the adaptation to phytotoxins; P450 enzymes are clearly involved in phytotoxin resistance in one of these models. The genetic advantages of D. melanogaster, including availability of the sequenced genome, should allow further study of these genes and identification of new ones, particularly regulatory genes, responsible for resistance. PMID- 11112180 TI - Chemical ecology and social parasitism in ants. AB - The chemical strategies by which parasites manage to break into the social fortresses of ants offer a fascinating theme in chemical ecology. Semiochemicals used for interindividual nestmate recognition are also involved in the mechanisms of tolerance and association between the species, and social parasites exploit these mechanisms. The obligate parasites are odorless ("chemical insignificance") at the time of usurpation, like all other callow ants, and this "invisibility" enables their entry into the host colony. By chemical mimicry (sensu lato), they later integrate the gestalt odor of this colony ("chemical integration"). We hypothesize that host and parasite are likely to be related chemically, thereby facilitating the necessary mimicry to permit bypassing the colony odor barrier. We also review the plethora of chemical weapons used by social parasites (propaganda, appeasement, and/or repellent substances), particularly during the usurpation period, when the young mated parasite queen synthesizes these chemicals before usurpation and ceases such biosynthesis afterwards. We discuss evolutionary trends that may have led to social parasitism, focusing on the question of whether slave-making ants and their host species are expected to engage in a coevolutionary arms race. PMID- 11112181 TI - Colony dispersal and the evolution of queen morphology in social Hymenoptera. AB - Social Hymenoptera show two contrasting strategies of colony reproduction. A reproductive female can raise the first generation of brood alone (independent foundation), or a colony can divide into autonomous parts in which the reproductive female is helped by sterile relatives (fission, budding, swarming). In independent-founding ants, queens can histolize their flight muscles after dispersal; in many species, large flight muscles and metabolic reserves reduce or eliminate the need for risky foraging trips during the vulnerable solitary stage. Colony division is a derived strategy, and we review the selective pressures leading to its occurrence in the different social taxa. In various ants, fission coexists with independent foundation, and alate queens are retained. However, in ants exhibiting obligate fission (e.g. all army ants and many Ponerinae), queens are permanently wingless (ergatoid), or the queen caste is missing altogether. When reproductive females are flightless, dispersal distances and colonization ability are reduced, and there are extensive modifications in mating behavior and resource allocation. We focus on the characteristics of fission in the phylogenetically primitive ants Ponerinae in which both ergatoid queens and gamergates occur. The ground-living habits of ants have permitted extensive changes in the phenotypes of their reproductive females, unlike in wasps and bees. PMID- 11112182 TI - Joining and avoidance behavior in nonsocial insects. AB - Groups of two or more consexual conspecific adults of many kinds of nonsocial insects have been observed to form at feeding, mating, ovipositional, or sheltering sites. Conversely, adults of these same insects have been observed to avoid joining consexual conspecifics (or their progeny) and to place themselves (or their progeny) at some distance that results in spacing. Examples from various taxa illustrate that mechanisms underlying joining or avoidance behavior differ among species, as do types of benefits and costs to individuals who decide to join or avoid others. Moreover, within a given species, the decision to join or avoid others can be affected markedly by the physiological and informational state of the individual and by contextual response thresholds to resource availability. Decisions that benefit the individual may or may not affect the group in terms of total reproductive output. PMID- 11112183 TI - Biological control of locusts and grasshoppers. AB - Control of grasshoppers and locusts has traditionally relied on synthetic insecticides, and for emergency situations this is unlikely to change. However, a growing awareness of the environmental issues associated with acridid control as well as the high costs of emergency control are expanding the demand for biological control. In particular, preventive, integrated control strategies with early interventions will reduce the financial and environmental costs associated with large-scale plague treatments. The recent development of effective oil formulations of Metarhizium anisopliae spores in Africa, Australia, and Brazil opens new possibilities for environmentally safe control operations. Metarhizium biopesticide kills 70%-90% of treated locusts within 14-20 days, with no measurable impact on nontarget organisms. An integrated pest management strategy, with an emphasis on the use of Metarhizium, that incorporates rational use of chemical pesticides with biological options such as the microsporidian Nosema locustae and the hymenopteran egg parasitoids Scelio spp., has become a realistic option. PMID- 11112184 TI - Neural limitations in phytophagous insects: implications for diet breadth and evolution of host affiliation. AB - This review points out the problem of processing multiple sensory inputs and provides evidence that generalists suffer a disadvantage compared with specialists with respect to efficiency of host plant choice and discrimination. The specialists' mechanisms for improved efficiency are discussed as well as some of the processes that may be selected to increase processing efficiency in generalists. The fitness consequences of differences in efficiency of specialists and generalists are pointed out. One of the major disadvantages for generalists is the increase in vulnerability to ecological risks, especially risks imposed by various natural enemies. Efficiency-related factors are indicated as previously underestimated elements that could influence host affiliations including diet breadth and changes in host plant use. PMID- 11112185 TI - Food webs in phytotelmata: "bottom-up" and "top-down" explanations for community structure. AB - The field study of food webs and the processes maintaining them is hampered by the sheer complexity and unreplicated nature of natural systems. The animal communities in phytotelmata--plant-held waters--are a convenient exception to this generalization. Tree holes, bamboo internodes, pitcher plants, tank bromeliads, and water-retaining plant axils contain a rich fauna, principally of arthropods, which constitute more or less complex, highly discrete food webs. They are widespread and replicated. The explanations for the community structure observed in these systems may call on "bottom-up" mechanisms such as simple environmental limitations, competition, predation, and facilitation, or they may adduce grander "top-down" theories, which explore biogeographic, energetic, dynamic, or biodiversity-related constraints. The existence of the bottom-up mechanisms is well established in experimental systems, and their consequences may be apparent in naturally occurring food webs. Top-down mechanisms demand a more holistic approach and are more difficult to test either by pattern analysis or experimental manipulation. The synoptic explanation of community composition and structure demands a multidimensional approach best expressed as a heuristic "template." Phytotelmata represent nearly ideal natural instruments for further study of food web dynamics, and exciting opportunities exist for the development and testing of community theories through their manipulation. PMID- 11112186 TI - Revision total hip arthroplasty with the use of structural acetabular allograft and reconstruction ring: a case series with a 10-year average follow-up. AB - From 1980 through 1993, 20 consecutive massive structural acetabular allografts and reconstruction rings were performed in 19 patients. In all cases, the magnitude of the acetabular bone deficiency was such that the allograft supported >50% of the cup. The allograft was necessary to restore normal anatomy, bone stock, and leg length. Of the 19 patients who met the inclusion criteria, 7 subjects died of unrelated causes, and 3 subjects failed and underwent resection arthroplasty, 1 (8%) for graft resorption and 2 (15%) for recurrent dislocation. The remaining 9 patients (10 allografts) had a minimum follow-up of 5 years and average follow-up of 10.5 years. The cohort was analyzed using radiographic and outcome data collection questionnaires (AAOS/HKOD, WOMAC, SF-36). The study supports the use of massive structural allografts and reconstruction rings and achieves satisfactory results in 77% (10 of 13) of the patients. We believe these results reveal an impressive outcome for what used to be thought of as a salvage operation. PMID- 11112187 TI - The Burch-Schneider antiprotrusio cage in acetabular revision surgery: a mean follow-up of 12 years. AB - A total of 38 acetabular revisions using a Burch-Schneider antiprotrusio cage in 37 patients (18 women and 19 men), with a mean age at surgery of 75 years (range, 55-88 years), were evaluated retrospectively with a mean follow-up of 12 years (range, 8-21 years). In 2 cases with total hip dislocation and in 1 case with a deep infection, revision of the antiprotrusio cages was required. Defining every revision of the antiprotrusio cage as the endpoint of survivorship, the antiprotrusio cage showed a survival rate of 92% after 21 years. Clinical evaluation of the surviving patients showed a mean Harris hip score of 76 points (range, 20-96). Radiologic evaluation revealed that 1 antiprotrusio cage was loose and that 4 femoral stems were loose. The Burch-Schneider antiprotrusio cage compares favorably with other devices with regard to long-term implant survival rate. PMID- 11112188 TI - The effect of built-in external femoral rotation on patellofemoral tracking in the genesis II total knee arthroplasty. AB - External rotation of the femoral component during total knee arthroplasty (TKA) has been suggested to improve flexion space balancing and patellofemoral tracking. Incorporation of external rotation into the design of the femoral component offers an alternative method to achieve this goal. This study compared 150 TKAs performed with traditional external rotation of the femoral component on the distal femur with a similar group of 150 TKAs performed with an implant that incorporates 3 degrees of external rotation into the femoral component. Statistical improvements were noted in the latter group, with respect to the need for intraoperative lateral retinacular release and in postoperative patellar tracking. Incorporation of 3 degrees of external rotation into the design of the femoral component seems to improve the overall result of TKA, especially with respect to patellofemoral resurfacing. PMID- 11112189 TI - Accuracy of soft tissue balancing in total knee arthroplasty. AB - To the best of our knowledge, this is the first study to assess the accuracy of balancing of the flexion and extension gaps in total knee arthroplasty (TKA). Measurements of the heights of the flexion and extension gaps were obtained during 104 consecutive primary, posterior-stabilized TKAs in osteoarthritic patients. Clinically, all knees appeared to be well balanced intraoperatively. Rectangular flexion and extension gaps almost always were obtained within 1 mm (84%-89%). None of the knees was >3 mm from being perfectly rectangular. Equality of the flexion and extension gaps was more difficult to obtain (47%-57% were within 1 mm). With meticulous attention, perfect soft tissue balance is not always achieved in TKA. PMID- 11112190 TI - Revision total hip arthroplasty using third-generation cementing technique. AB - A total of 83 consecutive first-revision total hip arthroplasties were performed in 83 patients using pressurized vacuum-mixed cement, a third-generation cementing technique. At a median follow-up of 3.6 years (range, 1.5-6.3 years), the overall failure rates (radiographic loosening, re-revision, or both) were 39% for the femoral components and 30% for the acetabular components. With definite radiographic loosening as an endpoint, the cement-bone interface achieved was the only significant factor to predict stem durability, with an adjusted odds ratio of 1.8 (95% confidence interval, 1.1-3.0). On the acetabular side, bone stock was the only significant factor, with an adjusted odds ratio of 5.3 (95% confidence interval, 4.0-27.7). In this retrospective cohort study, femoral cement pressurization per se did not appear to improve the results over those of other studies using second-generation techniques. PMID- 11112191 TI - Revision surgery after failed unicompartmental knee arthroplasty: a study of 35 cases. AB - This retrospective study evaluates the results of 35 revision procedures after failed unicompartmental knee arthroplasty (UKA) in 34 patients, which were done during the period 1986 to 1996. There were 28 women and 6 men with a mean age of 71 years (range, 54-85 years). In all cases, St. Georg and Endo (W. Link, Hamburg, Germany) unicompartmental prostheses were used except 1 PCA unicondylar implant (How medica, Rutherford, NJ) and 1 Bohler unicondylar implant (Allo Pro, Baar, Switzerland). Failures most frequently were due to aseptic loosening followed by polyethylene wear. Two deep infections occurred. Revisions were performed 1 week to 11 years after UKA; 23 were required within the first 5 years. In most cases, revision was to a total knee arthroplasty. Partial component exchange was done in 9 cases. All 34 patients were evaluated clinically after exchange arthroplasty. After a mean follow-up time of 4 years (range, 1 12.2 years), we found 11 excellent, 13 good, 4 fair, and 7 poor results according to the Hospital for Special Surgery score. The fair and poor results were due to aseptic loosening of the knee prosthesis in 6 knees. One of 2 patients with deep infection needed femoral amputation. With correct indication and considerable surgical experience, UKA is still a good alternative, especially in the elderly patient. PMID- 11112192 TI - Patient satisfaction and outcome after septic versus aseptic revision total knee arthroplasty. AB - A consecutive series of revision total knee arthroplasties performed at 3 university-affiliated centers by 3 surgeons was prospectively studied. The same implant was used in all cases. The evaluation included a Knee Society clinical score (KSCS); SF-36; satisfaction survey; and radiographs preoperatively, at 6 and 12 months postoperatively, and annually thereafter. Follow-up averaging 36 months (range, 24-60 months) was obtained in 125 of 138 knees (91%). Twenty-eight knees were infected, and 26 of 28 knees were treated successfully with 2-stage exchange with an interval of 4 to 6 weeks using an antibiotic-impregnated spacer block and intravenous antibiotics. The remaining 99 knees were revised for reasons other than infection, including aseptic component loosening, progressive osteolysis, and component instability. Preoperatively, patients with infection had a significantly decreased arc of motion compared with patients without infection (79 degrees vs 92 degrees; P<.05). There was a strong trend for the infected knees to have a lower preoperative KSCS than the noninfected knees, although this trend did not achieve statistical significance (76 vs. 92; P =.11). Postoperatively, patients with infection continued to have a significantly decreased range of motion (89 degrees vs. 99 degrees; P =.05). The postoperative KSCS was markedly lower in the septic versus aseptic revisions (115 vs. 135; P =.02). Patients with infection had a significantly lower function score (44 vs. 57; P =.03). A significantly higher percentage of patients stated that they were unable to return to normal activities of daily living after septic versus aseptic revision total knee arthroplasty (24% vs. 7%; P<.05). Despite the inferior functional result, patients expressed an equal degree of satisfaction with the results of their treatment in septic versus aseptic revision cases. PMID- 11112193 TI - The effect of epidemiologic and intraoperative factors on survival of the standard Souter-Strathclyde total elbow arthroplasty. AB - Previously published work has revealed an 87% survivorship after 12 years for the standard Souter-Strathclyde total elbow arthroplasty in patients with rheumatoid arthritis. Of the 13% that were revised, 75% were due to loosening of the humeral component. The aim of this research was to identify the specific epidemiologic and intraoperative factors that predisposed to this humeral loosening. Specifically, factors such as age, sex, radiologic staging of the disease, position of the implant in bone, and size of the implant inserted were evaluated. After analysis of 186 cases, we concluded that the position of the humeral component within the humerus is crucial for long-term survivorship. Specifically in the lateral plane, the stem should be aligned in the plane of the humerus and the implant inserted to the correct depth. The articular surface of the implant should lie at the level of the normal trochlea. At the anteroposterior plane, the implant should sit centrally and not be lateralized. We conclude that good surgical technique is crucial to the long-term effectiveness of this implant. PMID- 11112194 TI - Total hip arthroplasty in patients with below-knee amputations. AB - We present a unique small group of 5 patients with below-knee amputation who underwent total hip arthroplasty after a displaced subcapital fracture of the femur. Three patients were operated on after failed fixation of the fracture, and 2 were operated on as a primary procedure. All 5 patients resumed their prefracture level of activity and mobilization with no deterioration during follow-up (average, 69 months [range, 22-98]). These encouraging results call for use of total hip arthroplasty or hemiarthroplasty as the primary treatment modalities of patients with displaced subcapital femoral head fracture in an extremity with below-knee amputation. PMID- 11112195 TI - Total joint arthroplasty in patients surgically treated for morbid obesity. AB - The results of 20 total hip and knee arthroplasties performed in patients with morbid obesity who were treated with bariatric surgery before arthroplasty are reviewed. Bariatric surgery was successful in reducing the Quetelet ratio (weight in kilogram divided by height in square meters) of patients from a mean of 49 kg/m(2) (range, 38-56 kg/m(2)) to a mean of 29 kg/m(2) (range, 25-32 kg/m(2)). The average time from bariatric surgery to arthroplasty was 23 months (range, 7 65 months). The cumulative Knee Society score had improved significantly from a mean of 103.6 (range, 45-165) before arthroplasty to a mean of 148.9 (range, 66 185) at final follow-up in 12 knees undergoing total knee arthroplasty (P<.01). The Harris hip score also had increased significantly from a prearthroplasty mean of 40 (range, 25-55) to 67.5 (range, 50-95) at final follow-up in 8 hips receiving total hip arthroplasties (P<.05). All but 1 patient with continuing patellofemoral pain were satisfied with the result of the arthroplasty at final follow-up. One hip was revised at 5 years for aseptic loosening of the femoral component; no knee revisions were required. All other prostheses were stable with no evidence of radiographic loosening or wear at final surveillance. Morbidly obese individuals, with severe degenerative joint disease, who are considered unsuitable for arthroplasty because of excess weight should be considered for bariatric surgery. Total joint arthroplasty after surgical treatment of obesity has an excellent outcome with an acceptable complication rate. PMID- 11112196 TI - Aprotinin (Trasylol) does not reduce bleeding in primary total hip arthroplasty. AB - This is a randomized, double-blind, controlled study of the effects of aprotinin (Trasylol) during primary total hip arthroplasty. Sixty patients were randomized to receive either 1.5 x 10(6) KIU of aprotinin or a similar volume of normal saline as a bolus preoperatively. Blood loss was measured from the femoral canal at the time of surgery. An estimate of the total blood loss during the operation was made, and the transfusion requirement was recorded. There was no significant difference between the groups in terms of total blood loss, postoperative hemoglobin, or transfusion requirement. In the group that received aprotinin, there was a trend toward reduced blood loss from the femoral canal, but this was not statistically significant. The results of this study do not support the routine use of aprotinin in primary total hip arthroplasty. PMID- 11112197 TI - Slide and flex, tighten, extend (SAFTE): a safe, convenient, effective, and no cost approach to rehabilitation after total knee arthroplasty. AB - Many of the clinical aspects of total knee arthroplasty (TKA) are now standardized; however, treatment protocols for rehabilitation vary according to surgeon and physical therapy departments. The purpose of this article was to determine if the slide and flex, tighten, extend (SAFTE) approach after TKA is a satisfactory method of achieving functional range of motion (full extension and at least 90 degrees of flexion). Of patients in the study group, 70% achieved functional range of motion by the 7-week evaluation period. SAFTE is a safe, effective, and no-cost approach to achieve functional range of motion in TKA using a single-radius, posterior-stabilized knee prosthesis. PMID- 11112198 TI - Two-stage revision with pseudocapsular resection for recurrent dislocation of total hip prostheses. AB - Recurrent dislocation of the prosthesis is a problem with total hip arthroplasty. In the rare cases in which no identifiable cause can be found for the recurrent dislocation, we believe that the answer lies in addressing the large, sterile, fluid-filled cavity bounded by pseudocapsule, which we have noticed to form around the neck of the femoral implant in these cases. We report a 2-stage surgical technique that we have used successfully in 3 cases of recurrent dislocation without an identifiable cause, with an average follow-up of 3 years. PMID- 11112199 TI - The importance of tibial alignment: finite element analysis of tibial malalignment. AB - The influence of the tibial plateau orientation on cancellous bone stress was examined by finite element analysis for a cemented device. The objectives of the study were i) to examine the effect of the plateau-ankle angle on the cancellous bone stress, ii) to analyze the significance of the anteroposterior angles of the tibial component on these stresses, and iii) to compare the finite element predictions with clinical data. In general, positioning the tibial plateau in valgus resulted in lower cancellous bone stresses. These results support previous clinical studies, which suggest that overall alignment in valgus results in lower migration rates and lower incidence of loosening. PMID- 11112200 TI - Fracture and fatigue properties of acrylic bone cement: the effects of mixing method, sterilization treatment, and molecular weight. AB - The purpose of this study was to characterize the relative and combined effects of sterilization, molecular weight, and mixing method on the fracture and fatigue performance of acrylic bone cement. Palacos R brand bone cement powder was sterilized using ethylene oxide gas (EtO) or gamma irradiation. Nonsterile material was used as a control. Molecular weights of the bone-cement powders and cured cements were measured using gel permeation chromatography. Hand and vacuum mixing were employed to mold single edge-notched bend specimens for fracture toughness testing. Molded dog-bone specimens were used for fatigue tests. Electron microscopy was used to study fracture mechanisms. Analysis of variance and Student t-tests were used to compare fracture and fatigue performance between sterilization and mixing groups. Our results indicate that vacuum mixing improved significantly the fracture and fatigue resistance (P<.05, P<.07) over hand mixing in radiation-sterilized and EtO-sterilized groups. In vacuum-mixed cement, the degradation in molecular weight resulting from gamma irradiation decreased fracture resistance significantly when compared with EtO sterilization and control (P<.05). A corresponding decrease in fatigue resistance was observed in the cement that was degraded severely by a radiation dose of 10 MRad (P<.05). In contrast, EtO sterilization did not result in a significantly different fracture resistance when compared with unsterilized controls for vacuum-mixed cement (P>.1). For hand-mixed cement, fracture and fatigue resistance appeared to be independent of sterilization method. This independence is believed to be the result of higher porosity that compromised the mechanical properties and obscures any effect of sterilization. Our results indicate that a combination of nonionizing sterilization and vacuum mixing resulted in the best mechanical performance and is most likely to contribute to enhanced longevity in vivo. PMID- 11112201 TI - Early failure of modern cemented stems. AB - In the late 1970s, improved cement technique was introduced in an attempt to address the problem of early cemented stem loosening. Subsequently, numerous centers reported stem survival rates of >95% beyond 10 years. Long-term cemented stem fixation was believed widely to be consistently obtainable in most patients. Despite the widespread clinical success of these early cemented stems, numerous changes were introduced in stem design and cement technique. In more recent years, a surprising number of series of early failures of cemented stems have been reported. Some designs consistently have had a high early failure rate. Others have failed infrequently, but the failures have occurred early and with extensive osteolysis. Numerous causes have been proposed, including poor cement technique, undersized broaches, increased stem offset, decreased stem length, rough surface finish, and circular stem cross-section. Failures often are multifactorial and defy a simple explanation based on a single parameter. Results of cemented stems are more variable than previously appreciated. There are nuances of cemented stem design, cement technique, and patient selection that can lead to early failure and that are not understood completely at present. Given the availability of many cemented designs with proven records of clinical success, new design features should be introduced prudently with extensive premarket testing, limited clinical release, and careful postmarket surveillance. PMID- 11112202 TI - Allograft sterility as exemplified by human immunodeficiency virus and sterilization by irradiation. PMID- 11112203 TI - Late dislocations after cementless total hip arthroplasty resulting from polyethylene wear. AB - We report 4 cases of polyethylene wear in modular cementless sockets presenting as recurrent dislocation. Before the onset of the dislocations, the patients were functioning without symptoms. This is the first report in the literature of this phenomenon. PMID- 11112204 TI - Complete paraplegia as a result of regional anesthesia. AB - Complications after spinal or epidural anesthesia are rare. We report 2 cases of postoperative, complete paraplegia after regional anesthesia in orthopaedic patients not on anticoagulants. The paralysis was likely the result of spinal cord compression secondary to an epidural hematoma in 1 case and subdural hematoma in 1 case. A review of the literature regarding complications of regional anesthesia is presented. Regional anesthesia should be administered with caution and in selected patients. PMID- 11112205 TI - Two-stage operation for treatment of a large dissecting popliteal cyst after failed total knee arthroplasty. AB - A symptomatic popliteal cyst after total knee arthroplasty (TKA) is rare, occurring most frequently as a result of intra-articular knee pathology. We present a case of a large dissecting popliteal cyst 7 years after TKA with symptoms of severe calf pain and functional disability. The symptomatic cyst was excised completely in a first-stage operation, and the severely worn TKA was corrected by a second-stage surgical procedure. The patient in this report was pain free and had satisfactory range of knee motion 5 years after the index revision TKA, without recurrence of effusion or popliteal cyst formation. PMID- 11112206 TI - Precursor lesions of squamous cell carcinoma of the cervix: are there reliable predictors of biologic behavior? PMID- 11112207 TI - Prognostic significance of presence and reduplication of basal lamina in malignant pleural mesothelioma. AB - The prognosis of patients with malignant pleural mesothelioma (MM) is dependent more on tumor extension and differentiation than on therapeutic effects. Reduplication of the basal lamina (RBL) is an ultrastructural feature of some benign and malignant tumors that has been inversely correlated with aggressiveness and was recently described in MM. To investigate whether RBL is important for predicting the survival of patients with MM, transmission electron microscopy was used to identify the presence of basal lamina or RBL in biopsy specimens obtained by thoracoscopy from 35 patients. Cox's regression analysis was used to study the relation of these ultrastructural features to survival. Better outcomes were found for patients whose tumors expressed either basal lamina (HR 0.48; 95% CI, 0.09-2.47) or RBL (HR 0.38; 95% CI 0.12-1.22) compared with the reference category, where basal lamina or RBL was not found. The expression of basal lamina and RBL is an important novel prognostic factors in MM. HUM PATHOL 31:1341-1345. PMID- 11112208 TI - Patterns of melastatin mRNA expression in melanocytic tumors. AB - The melanocyte-specific gene Melastatin (MLSN1) shows an inverse correlation of mRNA expression with metastatic potential in human and murine cell lines in vitro. Melastatin mRNA expression in primary cutaneous melanoma also has been found to correlate with disease-free survival. The histologic patterns of Melastatin mRNA expression in nevi, primary melanoma, and melanoma metastases have not been described previously. Using in situ hybridization with (35)S labeled probes, we examined Melastatin mRNA expression in 64 cases of normal skin, benign melanocytic nevi, primary cutaneous melanomas, and melanoma metastases. Ubiquitous melanocytic expression of Melastatin mRNA was observed in all benign melanocytic proliferations (14 of 14), although some nevi showed a gradient of reduced Melastatin expression with increased dermal depth (3 of 14). Uniform expression of Melastatin mRNA was observed in 49% of primary cutaneous melanomas (18 of 37 cases, including 1 case of in situ melanoma). Melastatin mRNA loss by a portion of the melanoma was identified in 53% of the invasive melanoma samples (19 of 36) and 100% of the melanoma metastases (11 of 11). Primary melanomas without mRNA loss ranged in thickness from 0.17 to 2.75 mm (median, 0.5 mm; mean, 0.73 mm), whereas tumors that showed Melastatin mRNA down-regulation ranged in thickness from 0.28 to 5.75 mm (median, 1.7 mm; mean, 2.13 mm). A focal aggregate or nodule of melanoma cells without detectable signal was the most commonly observed pattern of Melastatin loss (13 of 19 cases), whereas complete loss of Melastatin mRNA expression by all of the dermal melanoma cells was observed in only 4 of the 19 cases. Two invasive melanomas displayed a scattered, nonfocal pattern of Melastatin mRNA loss. Of the 11 melanoma metastases examined, 64% displayed focal Melastatin mRNA loss, and 36% had complete loss of Melastatin mRNA expression. We observed several patterns of Melastatin mRNA expression in primary melanoma that may be distinguished from expression in benign melanocytic nevi. Melastatin mRNA expression appears to correlate with melanocytic tumor progression, melanoma tumor thickness, and the potential for melanoma metastasis. HUM PATHOL 31:1346:1356. PMID- 11112210 TI - Microscopic nephrocalcinosis and hypercalciuria in nephrotic syndrome. AB - Focal calcification is an occasional tubular abnormality seen in minimal-change nephrotic syndrome. Nephrocalcinosis was also reported in premature infants as a consequence of hypercalciuria resulting from long-term furosemide therapy. We describe 4 nephrotic children (3 minimal change, 1 diffuse proliferative glomerulonephritis) with transient hypercalciuria and intraluminal calcifications in renal histopathological specimens without radiologic evidence of renal calcification. These children were resistant to corticosteroid therapy and were receiving furosemide therapy along with albumin for management of oedema. Two of the children also had urinary infection. We were concerned that children with nephrotic syndrome are at risk for nephrocalcinosis, and urinary calcium and pH should be monitored carefully during prolonged furosemide use, especially in children with nephrotic syndrome with reduced initial responsiveness to corticosteroid therapy. HUM PATHOL 31:1363:1367. PMID- 11112209 TI - Increased carcinoembryonic antigen expression in cervical intraepithelial neoplasia grade 3 and in cervical squamous cell carcinoma. AB - Carcinoembryonic antigen (CEA) is a highly glycosylated cell surface protein that is overexpressed in a variety of human tumors and has been used as a tumor marker for disease progression in colorectal cancer patients. Recently, CEA has been used as a target for vaccine therapy against advanced CEA-expressing colonic adenocarcinomas. Previous reports have found elevated serum CEA levels in patients with cervical cancer, although this did not correlate with disease progression. In this study, cervical biopsy specimens from patients with normal histology, cervical intraepithelial neoplasia (CIN) grades 1 to 3, cervical squamous cell carcinoma (SCC), and adenocarcinoma were evaluated for CEA expression by immunohistochemistry by using the monoclonal antibody COL-1. Staining intensity was graded on a scale of 0 to 3 and was correlated with histologic diagnoses. CEA staining intensity was significantly increased in high grade squamous lesions (CIN III and SCC) compared with normal cervical mucosa and CIN grades I or II (P <.0001). There was a linear correlation between grade of lesion and staining intensity (r = 0.71). CEA expression increased most significantly between CIN grades 2 to 3. Only 1 of 7 primary cervical adenocarcinomas expressed CEA. Only 1 of 10 patients with high CEA expression in their tumors by immunohistochemistry had elevated serum CEA. We thus have shown that lesional CEA expression increases in CIN 3 and SCC without elevations in serum CEA. CEA expression may be a useful diagnostic tool and a useful marker for identifying those at risk for progressive cervical neoplasia. HUM PATHOL 31:1357 1362. PMID- 11112211 TI - Immunohistochemical and statistical studies on the islets of Langerhans pancreas in autopsied patients after gastrectomy. AB - We describe in detail the pathologic features of the islets of Langerhans in specimens of the pancreas from 75 autopsy specimens of patients who underwent gastrectomy and compare them with specimens from 22 patients not having gastrectomy with respect to the duration of the postoperative period and the operative procedure. In comparison with the control group, the islets of Langerhans from the gastrectomy patients showed hyperplasia and increased numbers of endocrine cells within 5 years of gastrectomy but atrophy and decreased numbers of endocrine cells thereafter. These changes are compatible with anti insulin immunoreactivity (B-cells) in the islets of Langerhans, where B-cell counts correlated with the area of the islets. The proliferating cell nuclear antigen (PCNA)-positive cell ratio in the islets was higher during the early phase after gastrectomy, whereas the apoptotic cell ratio was not affected by the time after gastrectomy. Hyalinization of the islets of Langerhans was first recognized after 5 years postoperatively, and the percentage of hyalinization gradually increased. Amylin deposits, detected by immunostaining, increased progressively in the islets in relation to the duration of postoperative period, correlating inversely with B-cell count and PCNA positivity. These histologic changes suggest that B cells proliferated secondary to oxyhyperglycemia in the early phase after gastrectomy followed by the state of waste, finally resulting in atrophy and amylin deposits, which led to hypofunction of the islets of Langerhans with resultant glucose intolerance. HUM PATHOL 31:1368-1376. PMID- 11112212 TI - Clonal analysis of the epithelial component of Warthin's tumor. AB - The proliferation of the epithelial component of Warthin's tumor is generally considered to represent a neoplastic condition. There has been much controversy about the histogenesis of this tumor, and the clonality of the epithelial component has not been clarified. We examined the clonal status of epithelial cells of Warthin's tumor by using a polymerase chain reaction (PCR) method based on trinucleotide repeat polymorphism of the X chromosome-linked human androgen receptor gene (HUMARA) and on random inactivation of the gene by methylation. Total DNA was isolated from formalin-fixed, paraffin-embedded tissue from 16 women with Warthin's tumor. Of the 16 cases analyzed, 7 were heterozygous for the HUMARA polymorphism and informative. The epithelial components of the tumors from the 7 cases were microdissected under the light microscope, and were subjected to extraction of DNA and HUMARA analysis. Using a permanent aqueous mounting medium during microdissection, we succeeded in reducing the rate of contamination by lymphocytes in the samples to less than 10%. All 7 cases showed patterns of polyclonal proliferation in the HUMARA analysis. Our results showed the nonclonal nature of Warthin's tumor, suggesting that Warthin's tumor is a non-neoplastic tumor-like condition. HUM PATHOL 31:1377-1380. PMID- 11112214 TI - Chronic histiocytic intervillositis: a placental lesion associated with recurrent reproductive loss. AB - Chronic (histiocytic) intervillositis (CHIV), defined for the purposes of this study as diffuse histiocytic infiltration of the intervillous space without villitis, is an idiopathic lesion seen in the chorionic sacs of some spontaneous abortion specimens and placentas. In this retrospective study, we evaluated all patients diagnosed with CHIV from 2 hospitals between 1993 and 2000, plus 1 additional patient from 1977. Histopathology, phenotype of the leukocytic infiltrate, perinatal outcome, and other associated clinical features were assessed by review of clinical records and all available pathology specimens plus immunohistochemical staining. CHIV was found in 31 of 45 specimens examined from 21 patients (23 of 31 first trimester, 3 of 5 second trimester, and 5 of 9 third trimester). Recurrence rate was 67% for patients with more than one specimen reviewed. Overall perinatal mortality rate was 77%, and only 18% of pregnancies reached 37 weeks. Eight of 19 patients with 3 or more pregnancies had recurrent spontaneous abortion (RSA); 5 with primary RSA (> or = 3 consecutive spontaneous abortions (SAB) with no living children) and 3 with secondary RSA (> or = 3 consecutive SAB with 1 or more living children). Severe intrauterine growth restriction was seen in 5 of 8 second- and third-trimester placentas with CHIV. Patients were generally not of advanced maternal age (mean, 29.8 +/- 6.2 years), and there was no obvious racial predisposition. Autoimmune or allergic phenomena were identified in 11 patients. Immunohistochemical staining of the intervillous infiltrate showed a near uniform population of monocyte-macrophages at varying stages of maturity and activation: more than 90% CD45Rb and CD68 positive, 30% to 40% MAC387 positive, less than 5% CD3 positive, and CD1a, CD20, CD30, and CD56 negative. We conclude that CHIV is an uncommon but important cause of recurrent spontaneous abortion and, in some cases, loss at later gestational ages. HUM PATHOL 31:1389-1396. PMID- 11112213 TI - Expression of melanoma inhibitory activity in melanoma and nonmelanoma tissue specimens. AB - Melanoma inhibitory activity (MIA) is a small soluble protein secreted by malignant melanoma cells and chondrocytes. Prior studies suggested that MIA expression was relatively tissue-specific, making it a potentially useful marker for melanoma. The current investigations sought to more clearly define the range of tumor/tissue-types where MIA is expressed, compared with expression of 4 other potential melanoma marker genes (tyrosinase melanoma antigen recognized by T cells [MART-1/MelanA], gp100, and melanoma growth-stimulatory activity [MGSA/Gro alpha]). Expression of these genes was assayed by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry in 23 melanoma tumor specimens and in 25 additional nonmelanoma or nonmalignant specimens. MIA, tyrosinase, and MGSA were expressed in most melanoma specimens. Specificity was highest for MART-1, followed by MIA and tyrosinase. Increasing the number of cycles of amplification from 35 to 40 increased sensitivity but decreased specificity of most markers, though MIA was relatively robust. MIA mRNA was also detected in carcinomas of the colon, ovary, kidney, and head/neck, as well as in normal laryngeal epithelium. Although MIA discriminated melanoma from nonmelanoma at least as well as tyrosinase, no single mRNA marker had accuracy greater than 71%, raising potential concern about application of these particular mRNA markers to the minimal disease setting. HUM PATHOL 31:1381-1388. PMID- 11112215 TI - Accumulation of p53 in infectious mononucleosis tissues. AB - Epstein-Barr virus (EBV) infects lymphocytes, where it persists indefinitely for the life of the host; whether the virus interacts with p53 to maintain itself in these cells is unknown. Lymphoid biopsy samples from 10 patients with infectious mononucleosis (IM) were examined for expression of p53 by immunohistochemistry. Accumulation of p53 was detected in all 10 cases, primarily in large lymphocytes of the expanded paracortex. The presence of EBV was confirmed in all 10 cases by EBER1 (EBV-encoded RNA) in situ hybridization, whereas 11 non-IM control samples lacked significant EBER1 and did not express p53 in paracortical lymphocytes. Interestingly, EBV infection alone does not cause accumulation of intracellular p53, because many more cells expressed EBER1 than p53 in the IM tissues. To determine whether p53 was confined to the subset of infected cells in which viral replication was occurring, BZLF1 immunostains were performed. Viral BZLF1 was detected in 8 of 10 IM tissues; however, the paucity and small size of the BZLF1 expressing lymphocytes suggests that they are not the same cells overexpressing p53. To further examine the relationship between p53 and EBV gene expression, the tissues were studied for latent membrane protein 1 (LMP1) expression by immunohistochemistry. Viral LMP1 was observed in the large paracortical lymphocytes of all 10 cases of IM, indicating co-localization of p53 and LMP1 in these cells. Our findings confirm that p53 overexpression is not specific for nodal malignancy and that p53 accumulation is characteristic of IM. Because p53 was not coexpressed in the same cells as BZLF1, it appears that BZLF1 is not directly responsible for p53 accumulation. Nevertheless, co-localization of p53 and LMP1 in activated-appearing lymphocytes suggests that EBV infection is responsible for p53 accumulation. HUM PATHOL 31:1397-1403. PMID- 11112216 TI - Enterocyte apoptosis and proliferation are increased in microvillous inclusion disease (familial microvillous atrophy). AB - Microvillous inclusion disease (MID) is characterized by diffuse villous atrophy without inflammatory changes. While increased apoptosis has been related to mucosal flattening in celiac disease, the role of apoptosis in the pathogenesis of MID is unknown. The aim of this study was to assess the rates of apoptosis and cell proliferation in MID and to compare them with those of normal controls and celiac disease. Small intestinal biopsies from 5 infants with MID, 10 children with normal villous architecture, and 10 children with untreated celiac disease were stained with the terminal uridine deoxynucleotidyl nick end labeling (TUNEL) method to assess apoptotic activity, and with Ki-67 immunohistochemistry to assess cellular proliferation. TUNEL and Ki-67 positive enterocytes were counted in a minimum of 20 well oriented half crypts per section. The percentage of apoptotic cells per crypt (apoptotic index) in normal, MID, and celiac biopsies was 0.03 +/- 0.01%, 0.08 +/- 0.08%, and 0.16 +/- 0.3%, respectively. Significant differences were found between normal and MID, and between normal and celiac cases. The percentage of Ki-67 positive cells per crypt (proliferation index) in normal, MID, and celiac cases was 14 +/- 2.5%, 28 +/- 9.2%, and 56 +/- 14%. Significant differences were found between the 3 groups. In conclusion, (1) enterocyte apoptosis and proliferation are increased in MID; (2) apoptosis appears to be an important factor of cell loss and may be, at least in part, responsible for villous atrophy in MID; and (3) crypts in MID are hyperplastic and not hypoplastic. HUM PATHOL 31:1404-1410. PMID- 11112217 TI - Nasopharyngeal angiofibroma: an APC-gene-associated tumor? AB - Nasopharyngeal angiofibromas are extremely rare, locally invasive tumors of unknown cause exclusively occurring in male adolescents. Recently, 6 cases have been reported in patients with familial adenomatous polyposis coli (Gardners syndrome). Mutations or allelic loss of the adenomatous polyposis coli (APC) gene have therefore been implied in the pathogenesis of nasopharyngeal angiofibroma. The authors analyzed 11 cases of nasopharyngeal angiofibromas from 9 male patients for mutations in the mutation cluster region and allelic loss of the APC gene. Six were primary tumors; 2 first recurrences; and 1, primary tumor with 2 recurrences. Direct sequence analysis was performed on several overlapping polymerase chain reaction (PCR) products. Detection of allelic loss was performed by restriction length polymorphism analysis at a polymorphic locus. No mutation was detected in 11 tumors of 9 different patients. None of the 9 informative (heterozygous) cases carried an allelic loss. We conclude that alterations of the APC gene do not play a major role in the development of nasopharyngeal angiofibroma. The coincidence of nasopharyngeal angiofibromas and adenomatous polyposis coli in some families implies the possibility that another gene in this region might be responsible for the development of nasopharyngeal angiofibromas. HUM PATHOL 31:1411:1413. PMID- 11112218 TI - PCR artifacts in LOH and MSI analysis of microdissected tumor cells. AB - Polymerase chain reaction (PCR) analysis to study loss of heterozygosity (LOH) and microsatellite instability (MSI) in tumors is widely used. Microdissection techniques are applied to obtain tumor-specific tissue cells. By microdissection, however, the amount of template DNA extracted may vary considerably and interfere with optimal PCR amplification. To circumvent LOH and MSI misinterpretations due to low DNA input, we have assessed the critical level of DNA input for reliable PCR analysis. PCR analysis was performed by using 18 polymorphic markers (mono-, di-, tri-, and tetranucleotide) on DNA derived from both paraffin-embedded, formalin-fixed, and fresh frozen tumor specimens at template input levels ranging from 0.05 to 25.0 ng. We show a highly significant relation between DNA input and the occurrence of LOH and MSI artifacts. Furthermore, for DNA extracted from paraffin-embedded material, the percentage of LOH artifacts is significantly higher compared with DNA extracted from frozen tissue. For reliable PCR analyses using a mono-, di-, tri-, or tetranucleotide marker, a minimum of 10.0 ng DNA is required when DNA is isolated from formalin-fixed, paraffin-embedded tissue and 5.0 ng when isolated from fresh frozen tissue. HUM PATHOL 31:1414-1419. PMID- 11112219 TI - Histopathology of familial ovarian tumors in women from families with and without germline BRCA1 mutations. AB - Breast cancers from patients with germline BRCA1 mutations show characteristic histopathologic features. However, similar studies of BRCA1-associated ovarian cancers have reported inconsistent findings. Interobserver differences in histopathologic classification are a significant source of variation, and most studies have obtained histopathologic information from pathology reports rather than from review of histopathology slides. We therefore reviewed the histopathology slides and pathology reports to determine histologic type, grade, and stage for cancers of the ovary or peritoneum in 217 women from 126 families enrolled in the Gilda Radner Familial Ovarian Cancer Registry. Peripheral blood DNA from at least 1 affected member of each family was analyzed for BRCA1 mutations, and tumors from BRCA1 mutation-positive families were compared with those from BRCA1-negative families. Of 66 patients from 36 BRCA1-positive families, 64 had ovarian carcinoma, 1 had an ovarian carcinoma in situ, and 1 had a dysgerminoma. Of 151 patients from 90 BRCA1-negative families, 135 had ovarian carcinoma, 10 had ovarian borderline tumors, 3 had ovarian sex cord/stromal tumors, and 3 had primary peritoneal carcinoma. There were fewer grade 1 (P <.001) and stage I (P =.10) cancers in patients from BRCA1-positive families than in patients from BRCA1-negative families. Neither mucinous nor borderline tumors were found in the BRCA1-positive families. Ovarian cancers arising in women from BRCA1-positive families are more likely to be high grade and nonmucinous than cancers arising in women from BRCA1-negative families. The absence of borderline tumors in patients from BRCA1-positive families adds to accumulating evidence that BRCA1 mutations do not play a role in the development of these tumors. HUM PATHOL 31:1420-1424. PMID- 11112220 TI - Donor origin of neuroendocrine carcinoma in 2 transplant patients determined by molecular cytogenetics. AB - Organ transplant recipients have an increased tumor incidence owing to their immunocompromised state. The origin of such tumors, whether donor or recipient, will have a clinical impact on decision-making concerning immunosuppressive therapy, retransplantation, and for recipients of other organs from the same donors. We report molecular cytogenetic determination of donor origin in 2 cases of small-cell neuroendocrine carcinoma developing in sex-mismatched transplant recipients (kidney and liver). Fluorescence in situ hybridization (FISH) analysis was performed on liver core needle biopsy material from the liver transplant patient and on liver fine needle aspiration cytopreparations from the kidney transplant patient. The results for the liver transplant patient were confirmed with microsatellite allelic analysis and with comparative genomic hybridization. In both cases, FISH showed the presence of only X chromosomes within the tumor cells, indicating the donor origin of the neoplasms. FISH is an excellent method to determine neoplastic origin in sex-mismatched transplant patients. HUM PATHOL 31:1425-1429. PMID- 11112221 TI - Diffuse leiomyomatosis of the uterus: a case report with clonality analysis. AB - Diffuse uterine leiomyomatosis is a rare condition distinguished from the common uterine leiomyomata by involvement of the entire myometrium by innumerable, ill defined, often small and confluent, histologically benign smooth-muscle nodules. Fourteen cases have been previously described in the literature. We report a case of diffuse leiomyomatosis in a 39-year-old woman. Several microscopic foci of the process were microdissected for clonality analysis. All samples showed a non random X-chromosome inactivation pattern, and thus were consistent with a monoclonal neoplastic population of cells. However, in different foci of tumor, different X chromosomes were inactivated, supporting the independent origin of neoplastic clones and rejecting the possibility of a single clonal origin of all tumor cells. The results of the molecular analysis suggest that diffuse uterine leiomyomatosis may be an exuberant example of diffuse and uniform involvement of the entire myometrium by multiple leiomyomata. HUM PATHOL 31:1429-1432. PMID- 11112222 TI - Fatal intrauterine adenoviral endomyocarditis with aortic and pulmonary valve stenosis: diagnosis by polymerase chain reaction. AB - We report a case of fatal hydrops fetalis owing to adenoviral endomyocarditis with aortic and pulmonary valve stenosis. A 1850-g macerated male stillborn delivered 1 week after fetal ultrasonography showed hydrops, cardiomegaly, and possible aortic valve stenosis. Autopsy confirmed hydrops and showed thickened, fibrotic semilunar valves with stenosis. The myocardium was focally fibrotic with areas of calcification. Polymerase chain reaction study of myocardial and aortic valve tissue was positive for adenovirus. Intrauterine viral myocarditis has been reported only rarely, but cases owing to Coxsackie B virus, adenovirus, and parvovirus B19 have appeared in the literature. With the exception of rubella, viral causation of significant valvular lesions in humans has received scanty support in the literature. This report suggests a broader group of causative agents. HUM PATHOL 31:1433-1435. PMID- 11112223 TI - Glioneuronal tumor with neuropil-like islands. AB - Mixed glioneuronal neoplasms are relatively uncommon tumors in the central nervous system. Recently, an unusual glioneuronal tumor arising in adults marked histologically by neuropil-like islands was described. We present a similar case arising in a 23-year-old woman who presented with headaches and seizures and on imaging studies was noted to have a frontal-temporal lobe mass. The patient underwent partial resection of the tumor, which histologically resembled anaplastic astrocytoma, and received a course of radiation therapy and chemotherapy. Increasing seizure frequency and expanding size on neuroimaging prompted a re-excision of the tumor. The second resection was marked by islands of tissue resembling gray matter with slightly atypical neuronal and glial cells situated in the white matter. These islands stained positively with synaptophysin and did not stain with glial fibrillary acid protein. Mild vascular proliferation and moderate nuclear pleomorphism also characterized the tumor. Areas of necrosis were not noted. A MIB-1 labeling index of 18.1% was noted. P53 immunoreactivity was observed in approximately 40% of tumor cell nuclei. This lesion is felt to represent a clinically aggressive glioneuronal neoplasm with an unusual and distinctive histologic phenotype. HUM PATHOL 31:1435-1438. PMID- 11112224 TI - Ovarian serous borderline tumors: the citadel defended. PMID- 11112225 TI - Reply: the citadel defended-The counterattack PMID- 11112226 TI - A bold proposal to achieve near-universal health care coverage in the United States. PMID- 11112227 TI - A 2020 vision for American health care. AB - We enter this century with an unprecedented federal budget surplus-$4.6 trillion over the next 10 years. A substantial portion of the surplus comes from savings in the health care sector. The 1997 Balanced Budget Act cut payments to Medicare providers and raised the premiums for individual beneficiaries, but we overshot the mark. Instead of balancing the budget, we generated a huge surplus. We underestimated the magnitude of Medicare savings. Medicare savings over the period from 1998 to 2007 represent an estimated 15% of the total budget surplus. Fifteen percent of the 10-year budget surplus from 2001 to 2010 comes to $680 billion. We also underestimated the drop-off in Medicaid coverage, as welfare reform took hold. In the year 2000 Medicare and Medicaid outlays were an estimated $104 billion less than projected just 5 years ago-representing an estimated 45% of the budget surplus this year, or about $1 trillion of the 10 year surplus. PMID- 11112228 TI - Hepatitis C in patients with human immunodeficiency virus infection: diagnosis, natural history, meta-analysis of sexual and vertical transmission, and therapeutic issues. AB - Hepatitis C (HCV) infection occurs in as many as 33% of the patients with human immunodeficiency virus (HIV) infection. In view of their improved survival, liver disease will become more clinically significant in patients coinfected with HIV/HCV. Several studies in patients with hemophilia have shown that coinfected patients develop earlier and more severe liver disease, including hepatocellular carcinoma. In nonhemophilic cohorts, lower CD4 counts are associated with an increased prevalence of cirrhosis. However, HCV infection does not seem to alter the natural history of HIV infection in most cases. Human immunodeficiency virus coinfection in pregnant women increases the risk of perinatal HCV transmission 2 fold, with more than 25% of occurrences involving transmission of both viruses: cesarean delivery significantly decreases this risk. The expanded use of highly active antiretroviral therapy may lead to further improvement in morbidity and mortality from HIV infection. Thus, the management of coexistent HCV liver disease will need to be formulated. We suggest that alcohol be disallowed. Interferon and ribavirin in combination are likely to become the therapy of choice, particularly in coinfected patients with higher CD4 counts, lower HCV viremia, and non-1 genotype. During treatment, complete blood cell counts need to be closely monitored. Future controlled trials will determine the efficacy and safety of long-acting interferon preparations. Administration of highly active antiretroviral therapy, with the intent to prevent decreases in CD4 counts, seems crucial in stemming liver disease progression. However, some drugs have clear-cut hepatotoxic potential and patients with known liver disease should be closely monitored. Arch Intern Med. 2000;160:3365-3373. PMID- 11112229 TI - Human tissue research in the genomic era of medicine: balancing individual and societal interests. AB - Advances in DNA sequencing technology and in our understanding of the human genome are ushering in a new era of genomic medicine, one with dramatic potential to not only benefit society through research involving human tissue, but also to cause economic or psychosocial harms to tissue donors and their families. This delicate situation requires that the needs of tissue donors be carefully considered and balanced with those of the medical research community, especially on issues concerning confidentiality, consent, and compensation. We analyzed the tensions between tissue donors and researchers over the research use of human tissue. We also reviewed several approaches, including the establishment of tissue-trustee infrastructures at academic medical centers, aimed at achieving a more equitable balance between individual donor protection and societal benefits derived from tissue-based research. Arch Intern Med. 2000;160:3377-3384. PMID- 11112230 TI - Temporal trends in outcomes of older patients with pneumonia. AB - BACKGROUND: It is unclear how outcomes of care for patients hospitalized for pneumonia have changed as patterns of health care delivery have changed during the 1990s. This study was performed to determine trends in outcomes of care for older patients hospitalized for pneumonia. METHODS: This retrospective analysis was based on Medicare claims and included most patients with pneumonia who were older than 65 years and admitted to acute care hospitals in Connecticut between October 1, 1991, and September 30, 1997 (fiscal years 1992-1997). We assessed the trends in hospital costs, discharge destination, hospital mortality rates, mortality rates within 30 days of discharge, and 30-day readmission rates for pneumonia. Multivariate logistic regression analyses were used to adjust for differences in patient characteristics. RESULTS: The mean (+/- SD) length of stay declined from 11.9 + 11.4 days to 7.7 + 7.2 days between 1992 and 1997. During this period, adjusted in-hospital mortality rates declined (P =.02), while the adjusted risk of discharge to a nursing facility increased (P<.001) and the adjusted risk of hospital readmission for pneumonia within 30 days of discharge increased (P =.05). The adjusted risk of death 30 days after discharge increased, although the difference was not statistically significant (P =.09). CONCLUSIONS: Between 1992 and 1997, the adjusted risks of mortality after discharge, placement in a nursing facility, and hospital readmission for pneumonia increased among older patients hospitalized for pneumonia, in association with a decline in mean hospital length of stay. These findings raise the question of whether the declining hospital length of stay has negatively affected patient outcomes. Arch Intern Med. 2000;160:3385-3391. PMID- 11112231 TI - Coffee consumption and the risk of coronary heart disease and death. AB - OBJECTIVES: To study prospectively the relation of coffee drinking with fatal and nonfatal coronary heart disease (CHD) and all-cause mortality and to perform a cross-sectional analysis at baseline on the association between coffee drinking and CHD risk factors, diagnosed diseases, self-reported symptoms, and use of medicines. METHODS: The study cohort consisted of 20 179 randomly selected eastern Finnish men and women aged 30 to 59 years who participated in a cross sectional risk factor survey in 1972, 1977, or 1982. Habitual coffee drinking, health behavior, major known CHD risk factors, and medical history were assessed at the baseline examination. Each subject was followed up for 10 years after the survey using the national hospital discharge and death registers. Multivariate analyses were performed by using the Cox proportional hazards model. RESULTS: In men, the risk of nonfatal myocardial infarction was not associated with coffee drinking. The age-adjusted association of coffee drinking was J shaped with CHD mortality and U shaped with all-cause mortality. The highest CHD mortality was found among those who did not drink coffee at all (multivariate adjusted). Also, in women, all-cause mortality decreased by increasing coffee drinking. The prevalence of smoking and the mean level of serum cholesterol increased with increasing coffee drinking. Non-coffee drinkers more often reported a history of various diseases and symptoms, and they also more frequently used several drugs compared with coffee drinkers. CONCLUSIONS: Coffee drinking does not increase the risk of CHD or death. In men, slightly increased mortality from CHD and all causes in heavy coffee drinkers is largely explained by the effects of smoking and a high serum cholesterol level. Arch Intern Med. 2000;160:3393-3400. PMID- 11112232 TI - The diffusion of a novel therapy into clinical practice: the case of sildenafil. AB - BACKGROUND: Erectile dysfunction is a common condition, yet in the past most affected men did not seek medical treatment. OBJECTIVE: To examine how sildenafil (Viagra), a new medication for the treatment of erectile dysfunction, has been incorporated into general medical practice. SUBJECTS AND METHODS: The study population consisted of all male members of a group-model Massachusetts health maintenance organization (HMO) whose first prescription for sildenafil was dispensed during the first 24 weeks of its availability through the HMO as a plan benefit (April 24, 1998, through October 8, 1998). Data collected on each member in the study population included age, specialty of the prescribing physician, initial dose, use of prior treatments for erectile dysfunction, receipt of medications known to predispose to impotence, filling of a second prescription for sildenafil, and concomitant medical conditions (including hypertension, ischemic heart disease, hyperlipidemia, diabetes mellitus, and history of radical prostatectomy). Cross tabulations and logistic regression models were constructed to evaluate the potential associations between filling a second prescription for sildenafil and other characteristics of sildenafil users. RESULTS: We identified 899 members who filled a first-time sildenafil prescription in the 24-week period of interest. The majority of sildenafil prescriptions that were filled for the first time (85%) occurred in the first 12 weeks of its availability. Most sildenafil users (84%) were between 45 and 74 years of age (average age, 61 years; age range, 23 to 90 years), and approximately 40% had documentation of prior treatment for erectile dysfunction. Use was highest among those aged 55 to 64 years, with almost 5% of all male HMO members in that age group having received at least 1 sildenafil prescription. Our cohort of sildenafil users was significantly more likely to have hypertension (P<.01), hyperlipidemia (P<.01), and diabetes mellitus (P<.01) than persons who participated in a widely publicized clinical trial of the medication. Prescribing physicians were predominantly primary care physicians (78% were internists, and 11% were family practitioners). More than 60% of sildenafil users filled a second prescription within 3 months of the first prescription; in multivariate analyses, factors associated with filling a second prescription included younger age and prior treatment for erectile dysfunction. CONCLUSIONS: Sildenafil was rapidly adopted into the clinical practice of primary care physicians for the treatment of erectile dysfunction in the managed care setting. The patients for whom the drug was prescribed in the general practice setting differed across many medical characteristics from study subjects who participated in clinical trials of the drug. Arch Intern Med. 2000;160:3401-3405. PMID- 11112234 TI - An epidemiologic study of risk factors for deep vein thrombosis in medical outpatients: the Sirius study. AB - BACKGROUND: Little information is available concerning risk factors for venous thromboembolism (VTE) in nonhospitalized patients. PARTICIPANTS AND METHODS: An epidemiologic case-control study of deep vein thrombosis (DVT) risk factors was conducted in 1272 outpatients by general practitioners. The case population (636 patients presenting with DVT) was paired with the control population (636 patients presenting with influenzal or rhinopharyngeal syndrome) according to sex and age. Deep vein thrombosis was to be documented by at least 1 objective test. Risk factors were classified into "intrinsic" ("permanent") and "triggering" ("transient") factors and were evidenced using univariate analysis. RESULTS: In the medical population, defined as patients who had not undergone surgery or application of a plaster cast to the lower extremities within the 3 weeks preceding inclusion (494 cases and 494 controls), intrinsic factors such as history of VTE, venous insufficiency, chronic heart failure, obesity, immobile standing position, history of more than 3 pregnancies, and triggering factors such as pregnancy, violent effort, or muscular trauma, deterioration of general condition, immobilization, long-distance travel, and infectious disease were significantly more frequent in the case patients than in the controls (odds ratio, >1; P<.05). In the overall population, additional risk factors were cancer, blood group A, plaster cast of the lower extremities, and surgery. In both populations, the number of risk factors per patient was greater in the case patients than in the controls. CONCLUSION: Several risk factors for DVT were identified in medical outpatients presenting with DVT, and their comprehension may improve appropriateness and efficiency of the different methods available for thromboprophylaxis. Arch Intern Med. 2000;160:3415-3420. PMID- 11112233 TI - Potential clinical and economic effects of homocyst(e)ine lowering. AB - BACKGROUND: Elevated total homocyst(e)ine levels (>/=11 micromol/L) have been identified as a potential risk factor for coronary heart disease. However, the benefits expected from lowering homocyst(e)ine levels with folic acid and vitamin B(12) supplementation have yet to be demonstrated in clinical trials. SUBJECTS AND METHODS: We constructed a decision analytic model to estimate the clinical benefits and economic costs of 2 homocyst(e)ine-lowering strategies: (1) "treat all"-no screening, daily supplementation with folic acid (400 microg) and vitamin B(12) (cyanocobalamin; 500 microg) for all; (2) "screen and treat"-screening, followed by daily supplementation with folic acid and vitamin B(12) for individuals with elevated homocyst(e)ine levels. Simulated cohorts of 40-year-old men and 50-year-old women in the general population were evaluated. In the base case analysis, we assumed that lowering elevated levels would reduce excess coronary heart disease risk by 40%; however, this assumption and others were evaluated across a broad range of potential values using sensitivity analysis. Primary outcomes were discounted costs per life-year saved. RESULTS: Although the treat-all strategy was slightly more effective overall, the screen and treat strategy resulted in a much lower cost per life-year saved ($13,600 in men and $27,500 in women) when compared with no intervention. Incremental cost effectiveness ratios for the treat-all strategy compared with the screen and treat strategy were more than $500,000 per life-year saved in both cohorts. Sensitivity analysis showed that cost-effectiveness ratios for the screen and treat strategy remained less than $50,000 per life-year saved under several unfavorable scenarios, such as when effective homocyst(e)ine lowering was assumed to reduce the relative risk of coronary heart disease-related death by only 11% in men or 23% in women. CONCLUSIONS: Homocyst(e)ine lowering with folic acid and vitamin B(12) supplementation could result in substantial clinical benefits at reasonable costs. If homocyst-(e)ine lowering is considered, a screen and treat strategy is likely to be more cost-effective than universal supplementation. Arch Intern Med. 2000;160:3406-3412. PMID- 11112235 TI - A search for sex differences in response to analgesia. AB - BACKGROUND: It is generally accepted that males and females respond differently to painful conditions. With few exceptions, according to the published literature, females demonstrate a lower pain threshold and a lower tolerance of painful stimuli. There is some support in the literature that females experience greater analgesic efficacy than do males after the administration of narcotic analgesics. We compared the analgesic response of females and males to ibuprofen in a post-third-molar extraction dental pain model. METHODS: We performed a meta analysis of 314 subjects included in the ibuprofen treatment arm of 7 double blind, post-third-molar extraction dental pain (moderate to severe) studies, which were submitted to the agency electronically. The inclusion and exclusion criteria were practically identical in all studies. Pain relief and pain intensity measurements used the same metrics in all studies and were recorded just before and at least at 0.5, 1.0, 1.5, 2.0, 3.0, 4.0, 5.0, and 6. 0 hours after drug administration. RESULTS: The study included 195 female subjects and 119 male subjects (mean age, 21 years). Other than requiring dental extractions, the subjects were all healthy. Postoperative baseline pain was greater in females than in males to a statistically significant degree (P =.006). Both pain intensity and pain relief scores demonstrated the well-established analgesic effect of ibuprofen in the pooled data set as well as in all the individual studies. Moreover, the mean pain intensity and pain relief scores over time for the female and male treatment groups were not noticeably different at any time point after drug administration, with no imputation for missing values. Analysis of the data using the "baseline observation carried-forward" technique for remedicated subjects (the technique recommended by the Food and Drug Administration for efficacy analysis of acute analgesic medications) produced the same results, which were confirmed by analysis of variance and t tests at each time point of the study. CONCLUSIONS: Our results demonstrated no sex effect on the analgesic response to ibuprofen. These results were obtained under the post third-molar extraction setting, in which the least possible confounding factors are present. To fully establish the generality of this phenomenon, studies should be carried out in other pain models and using analgesic medications with different mechanisms of action. Arch Intern Med. 2000;160:3424-3428. PMID- 11112236 TI - Clinical risk factors and timing of recurrent venous thromboembolism during the initial 3 months of anticoagulant therapy. AB - BACKGROUND: In patients with venous thromboembolism (VTE), identifying clinical risk factors for recurrence during the initial 3 months of anticoagulant therapy and knowledge of the time course of recurrence may help clinicians decide about the frequency of clinical surveillance and the appropriateness of outpatient treatment. METHODS: Analysis of a randomized controlled trial database involving 1021 patients with VTE (750 with deep vein thrombosis [DVT] and 271 with pulmonary embolism [PE]) who were followed up for 3 months after the start of anticoagulant therapy. All patients received initial treatment with unfractionated heparin or a low-molecular-weight heparin (reviparin) and a coumarin derivative starting the first or second day of treatment, with a target international normalized ratio of 2.0 to 3.0. RESULTS: Four independent clinical risk factors for recurrent VTE were identified: (1) cancer (odds ratio [OR], 2.72; 95% confidence interval [CI], 1. 39-5.32), (2) chronic cardiovascular disease (OR, 2.27; 95% CI, 1. 08-4.97), (3) chronic respiratory disease (OR, 1.91; 95% CI, 0.85-4. 26), and (4) other clinically significant medical disease (OR, 1.79; 95% CI, 1.00-3.21). Older age was associated with a decreased risk for recurrent VTE (OR, 0.76; 95% CI, 0.64-0.92). Previous VTE, sex, and idiopathic VTE were not risk factors for recurrence. In patients with DVT or PE, there was no significant difference in the rates of recurrent nonfatal VTE (4.8% vs 4.1%; P =.62), major bleeding (2.9% vs 2.2%; P =.53), and non-VTE death (6.4% vs 7.8%; P =.45), but recurrent fatal PE was more frequent in patients with PE than DVT (2. 2% vs 0%; P<.01). There was a clustering of recurrent VTE episodes during the initial 2 to 3 weeks after the start of treatment. CONCLUSIONS: During the initial 3 months of anticoagulant therapy, recurrent VTE is more likely to occur in patients with cancer, chronic cardiovascular disease, chronic respiratory disease, or other clinically significant medical disease. Patients with PE are as likely to develop recurrent VTE as those with DVT; however, recurrence is more likely to be fatal in patients who initially present with PE. Arch Intern Med. 2000;160:3431-3436. PMID- 11112237 TI - Out-of-hospital cardiac arrest in octogenarians and nonagenarians. AB - BACKGROUND: Studies of elderly patients who have out-of-hospital cardiac arrest have contradictory results. The studies usually define elderly patients as those older than 70 years, and include relatively few octogenarians and nonagenarians. OBJECTIVES: To compare the survival after out-of-hospital cardiac arrest of octogenarians, nonagenarians, and younger patients and to determine the influence of age on survival after adjusting for factors known to influence out-of-hospital cardiac arrest outcome. METHODS: We conducted a retrospective cohort study in suburban King County, Washington, on 5882 patients who had out-of-hospital cardiac arrest from presumed cardiovascular disease between January 1, 1987, and December 31, 1998, and who received cardiopulmonary resuscitation from bystanders, emergency medical technicians, or both. The main outcome measure was survival to hospital discharge. RESULTS: In patients who had out-of-hospital cardiac arrest due to a cardiac cause, younger patients had higher hospital discharge rates than octogenarians, who in turn had higher hospital discharge rates than nonagenarians (19.4% vs 9.4% vs 4.4%; P<.001). However, survival to hospital discharge improved significantly for younger patients, octogenarians, and nonagenarians who had ventricular fibrillation or pulseless ventricular tachycardia (36% vs 24% vs 17%; P<.001). After multiple logistic regression analysis controlling for other factors, increased age was weakly associated with decreased survival to hospital discharge (odds ratio, 0.92; 95% confidence interval, 0. 85-0.99). CONCLUSIONS: Octogenarians and nonagenarians have lower survival to hospital discharge than younger patients, but age is a much weaker predictor of survival than other factors such as initial cardiac rhythm. Decisions regarding resuscitation should not be based on age alone. Arch Intern Med. 2000;160:3439-3443. PMID- 11112238 TI - Long-term effects of raloxifene on bone mineral density, bone turnover, and serum lipid levels in early postmenopausal women: three-year data from 2 double-blind, randomized, placebo-controlled trials. AB - BACKGROUND: In postmenopausal women, raloxifene hydrochloride has favorable effects on bone and lipid metabolism and does not stimulate reproductive tissues. The studies reported herein evaluated the long-term (3-year) effects of raloxifene treatment on bone mineral density (BMD), serum lipid levels, and drug tolerability in healthy postmenopausal women. METHODS: A total of 1145 healthy European and North American postmenopausal women aged 45 through 60 years were enrolled in 2 parallel, double-blind, randomized, placebo-controlled trials of identical design and randomly assigned to receive raloxifene hydrochloride, 30, 60, or 150 mg, or placebo daily; all groups received 400 to 600 mg of elemental calcium. Assessments included measurements for BMD by dual-energy x-ray absorptiometry, markers of bone turnover, and serum lipid levels. RESULTS: Lumbar spine BMD changed from baseline to 36 months as follows: placebo (mean percentage change + SE), -1. 32% +0.22%; raloxifene, 30 mg, 0.71% +0.23%; raloxifene, 60 mg, 1. 28% +0.23%; and raloxifene, 150 mg, 1.20% +0.24%. Comparable BMD changes were observed in the hip and total body. Biochemical markers of bone turnover were suppressed by raloxifene to normal premenopausal ranges through 3 years. Serum low-density lipoprotein cholesterol was reduced 7% to 12% below baseline through 3 years. Study withdrawals due to any reason (37%) and withdrawals due to adverse events (14%) were not different among groups. The only significant adverse effect of therapy was hot flashes (25% in the 60-mg raloxifene group vs 18% in the placebo group); hot flashes were typically reported as mild and were not associated with study withdrawal (1.7% for 60-mg raloxifene vs 2.4% for placebo). CONCLUSIONS: Raloxifene preserves BMD at important skeletal sites, lowers serum low-density lipoprotein cholesterol levels, and has a tolerability profile comparable to placebo. These results indicate a favorable benefit-risk profile of raloxifene for long-term use in healthy postmenopausal women. Arch Intern Med. 2000;160:3444-3450. PMID- 11112239 TI - The influence of marital adjustment on 3-year left ventricular mass and ambulatory blood pressure in mild hypertension. AB - BACKGROUND: Of psychosocial stressors, job strain has been associated with a sustained increase in blood pressure. The impact of marital factors on blood pressure and target organ has not been explored. OBJECTIVES: To evaluate whether marital adjustment, measured at baseline by self-report (Dyadic Adjustment Scale) influences left ventricular mass index (LVMI) and ambulatory blood pressure measured over 3 years in patients with mild hypertension. METHODS: A prospective cohort study was conducted on 103 cohabiting males or females, including 72 with technically adequate echocardiograms, who at baseline were unmedicated, employed, and living with a significant other, all for a minimum of 6 months and had repeated elevated office diastolic blood pressure. MAIN OUTCOME MEASURES: Left ventricular mass by M-mode echocardiography indexed to body surface area and blood pressure were measured by ambulatory blood pressure every 15 minutes (daytime) and hourly between 11 PM and 7 AM. RESULTS: Marital adjustment, smoking, drinking, and baseline LVMI contributed significantly to the prediction of 3-year LVMI (semipartial correlation, sr(2) = 0.04, 0.07, 0.03, and 0.22; P =.03,.008,.08, and <.001, respectively) together accounting for 36% of the total variability in follow-up LVMI. Three-year ambulatory blood pressure measures were not significantly related to marital adjustment but there were correlations with Dyadic Adjustment Scale subscales. Low or high levels of spousal contact during 3 year ambulatory blood pressure monitoring were associated with an increase or decrease of 3-year, 24-hour diastolic blood pressure, consistent with the quality of marital adjustment (P =.04) or marital satisfaction (Dyadic Adjustment Scale subscale, P =.008). CONCLUSIONS: In a cohort of subjects with mild essential hypertension, marital adjustment had an influence on 3-year LVMI. Depending on the quality of marital adjustment, spousal contact at 3 years was associated with an increase or decrease of 3-year diastolic blood pressure. Confirmation of these results, including objective marital assessment and the participation of normotensive subjects, is required. Arch Intern Med. 2000;160:3453-3458. PMID- 11112240 TI - Results of isoproterenol tilt table testing in monozygotic twins discordant for chronic fatigue syndrome. AB - BACKGROUND: The pathogenesis of chronic fatigue syndrome (CFS) is unknown. Neurally mediated hypotension (NMH) has been suggested as a common comorbid condition or a potential underlying cause. METHODS: We conducted a cotwin control study of 21 monozygotic twins who were discordant for CFS. One twin met the 1994 Centers for Disease Control and Prevention criteria for CFS, and the other twin was healthy and denied chronic fatigue. The twins were selected from a volunteer twin registry in which at least 1 member reported persistent fatigue. As part of a 7-day clinical evaluation, all 21 twin pairs were evaluated with a 3-stage tilt table test with isoproterenol hydrochloride for the assessment of NMH. The presence of NMH was defined as syncope or presyncope associated with a decrease of 25 mm Hg in blood pressure and no associated increase in heart rate. RESULTS: A positive tilt table test result was observed in 4 twins with CFS (19%) and in 4 healthy twins (19%). This difference was not statistically significant (matched pair odds ratio, 1.0; 95% confidence interval, 0.2-5.4; P>.90). Compared with the healthy twins, the twins with CFS reported more severe symptoms of CFS and NMH both in the week before and during the tilt table test. CONCLUSIONS: These results do not support a major role for NMH in CFS. They highlight the importance of selecting well-matched control subjects, as well as the unique value of the monozygotic cotwin control design in the study of this illness. Arch Intern Med. 2000;160:3461-3468. PMID- 11112241 TI - Public health and clinical implications of high hemoglobin A1c levels and weight in younger adult Native American people with diabetes. AB - BACKGROUND: Type 2 diabetes mellitus is a major public health issue for Native American people. Because glycemic levels are predictive of diabetes outcome, understanding determinants of high hemoglobin A(1c) (HbA(1c)) levels may provide targets for prevention efforts. OBJECTIVES: To investigate determinants of high HbA(1c) levels in Native American people. METHODS: We conducted a population based, cross-sectional study of 206 participants with diabetes from 8 Native American communities in New Mexico. We used linear regression to assess the relationship of HbA(1c) level with age, body mass index (BMI), treatment type, duration of diabetes, physical activity, and diet. RESULTS: Age, dietary pattern, and treatment type were determinants of HbA(1c) levels. Participants younger than 55 years had the highest adjusted HbA(1c) levels at 9.5% and those 65 years and older had the lowest levels at 7.8%. According to a participant's dietary intake, HbA(1c) levels were highest for those who consumed the most fat and sugar, and high consumption of fat and sugar affected HbA(1c) levels most among those younger than 55 years. Participants treated with insulin had the highest hemoglobin A(1c) levels. Physical activity was not associated with HbA(1c) level. CONCLUSIONS: We found an increasing severity of diabetes among younger people. To avoid increased morbidity and mortality in the future, young Native American adults with diabetes need vigorous therapy to maintain tight glucose control. Arch Intern Med. 2000;160:3471-3476. PMID- 11112242 TI - Experts practice what they preach: A descriptive study of best and normative practices in end-of-life discussions. AB - BACKGROUND: Advance directives (ADs) are widely regarded as the best available mechanism to ensure that patients' wishes about medical treatment at the end of life are respected. However, observational studies suggest that these discussions often fail to meet their stated goals. OBJECTIVES: To explore best practices by describing what physicians who are considered expert in the area of end of-life bioethics or medical communication do when discussing ADs with their patients and to explore the ways in which best practices of the expert group might differ in content or style from normative practice derived from primary care physicians' discussions of ADs with their patients collected as part of an earlier study. DESIGN: Nonexperimental, descriptive study of audiotaped discussions. SETTING: Outpatient primary care practices in the United States. PARTICIPANTS: Eighteen internists who have published articles in the areas of bioethics or communication and 48 of their patients. Fifty-six academic internists and 56 of their established patients in 5 practice sites in 2 locations-Durham, NC, and Pittsburgh, Pa. Eligible patients were at least 65 years old or suffered from serious medical illness and had not previously discussed ADs with their physician. Expert clinicians had discretion regarding patient selection, while the internists chose patients according to a predetermined protocol. MEASUREMENTS: Coders applied the Roter Interaction Analysis System (RIAS) to audiotapes of the medical visits to describe communication dynamics. In addition, the audiotapes were scored on 21 items reflecting physician performance in specific skills related to AD discussions. RESULTS: Experts spent close to twice as much time (14.7 vs 8.1 minutes, P<.001) and were less verbally dominant (P<.05) than other physicians during AD discussions. When length of visit was controlled statistically, the expert physicians gave less information about treatment procedures and biomedical issues (P<.05) and asked fewer related questions (P<. 05) but tended toward more psychosocial and lifestyle discussion and questions. Experts engaged in more partnership building (P<.05) with their patients. Patients of the expert physicians engaged in more psychosocial and lifestyle discussion (P<.001), and more positive talk (P<.05) than patients of community physicians. Expert physicians scored higher on the 21 items reflecting AD-specific skills (P<.001). CONCLUSIONS: Best practices as reflected in the performance of expert physicians reflect differences in measures of communication style and in specific AD-related proficiencies. Physician training in ADs must be broad enough to include both of these domains. Arch Intern Med. 2000;160:3477 3485. PMID- 11112243 TI - Efficacy and safety of acetaminophen in the treatment of migraine: results of a randomized, double-blind, placebo-controlled, population-based study. AB - BACKGROUND: Although most persons with migraine treat their headaches with over the-counter medication, systematic data on the safety and efficacy of widely used treatment, including acetaminophen, are sparse. METHODS: This is a randomized, double-blind, placebo-controlled study comparing oral acetaminophen, 1000 mg (two 500-mg Extra Strength Tylenol tablets), with identical placebo in the treatment of a single acute migraine attack. Eligible subjects met International Headache Society diagnostic criteria for migraine with or without aura. Patients who usually required bed rest with their headaches or who vomited more than 20% of the time were excluded. MAIN OUTCOME MEASURES: The percentage of subjects who, at 2 hours after dosing, experienced a change in baseline pain intensity from severe or moderate pain to mild or no pain (headache response); and pain intensity difference from baseline at the 2-hour postmedication assessment. RESULTS: The headache response rate 2 hours after dosing was 57.8% in the acetaminophen group and 38.7% in the placebo group (P =.002). Pain-free rates at 2 hours were 22.4% in the acetaminophen group and 11.3% in the placebo group (P =.01). The mean pain intensity difference from baseline 2 hours after dosing was 1.08 in the acetaminophen group and 0.73 in the placebo group (P<.001). At 2 hours, other migraine headache characteristics, such as functional disability (P =.002), photophobia (P =.02), and phonophobia (P =.08), were significantly improved after treatment with acetaminophen vs placebo. CONCLUSIONS: Acetaminophen was highly effective for treating pain, functional disability, photophobia, and phonophobia in a population-based sample of persons with migraine, excluding the most disabled persons with migraine. The drug also had an excellent safety profile and was well tolerated. Arch Intern Med. 2000;160:3486-3492. PMID- 11112244 TI - Some gaps cannot be bridged. PMID- 11112246 TI - Thrombolysis and pulmonary embolism presenting with cardiac arrest. PMID- 11112248 TI - White-coat hypertension or white-coat hypertension syndrome: which is accompanied by target organ damage? PMID- 11112250 TI - Invasive aspergillosis in a patient with ticlopidine-induced agranulocytosis. PMID- 11112251 TI - Reduction of buffalo hump by switching to amprenavir in an HIV-infected patient. PMID- 11112253 TI - Evolution of cooperative problem solving in an artificial economy AB - We address the problem of how to reinforce learning in ultracomplex environments, with huge state-spaces, where one must learn to exploit a compact structure of the problem domain. The approach we propose is to simulate the evolution of an artificial economy of computer programs. The economy is constructed based on two simple principles so as to assign credit to the individual programs for collaborating on problem solutions. We find empirically that starting from programs that are random computer code, we can develop systems that solve hard problems. In particular, our economy learned to solve almost all random Blocks World problems with goal stacks that are 200 blocks high. Competing methods solve such problems only up to goal stacks of at most 8 blocks. Our economy has also learned to unscramble about half a randomly scrambled Rubik's cube and to solve several commercially sold puzzles. PMID- 11112254 TI - Cortical potential distributions and information processing. AB - The use of sets of spatiotemporal cortical potential distributions (CPDs) as the basis for cognitive information processing results in a very large space of cognitive elements with natural metrics. Results obtained from current source density (CSD) analysis suggest that in the CPD picture, action potentials may make only a relatively minor contribution to the brain's code. In order to establish if two CPDs are close, we consider standard metrics in spaces of continuous functions, and these may be employed to ascertain if two stimuli will be identified as the same. The correspondence between the electrical activity within brain regions, including not only action potentials but all postsynaptic potentials (PSPs), and CPDs is considered. We examine the possibility of using the CSD approach to find potential distributions using the descriptive approach in which precise sets of times are ascribed to the occurrence of action potentials and PSPs. Using metrics in the multidimensional space of paths of collections of point processes, we show that closeness of CPDs is implied by closeness of sets of spike times and PSP times if a certain metric is used but not others. We also set forth a dynamical model consisting of a system of reaction-dif fusion equations for ionic concentrations coupled with nerve membrane potential equations and active transport systems. Making the approximation of a descriptive approach, the correspondence between sets of spike times and PSP times and CPDs is obtained as with the CSD method. However, since it is not possible to ascribe precise times to the occurrence of PSPs and action potentials, the descriptive approach cannot be used to describe the configuration of electrical activity in cortical regions accurately. We also discuss how the CPD framework relates to the binding problem and submillisecond timing. PMID- 11112255 TI - Asymptotic bias in information estimates and the exponential (Bell) polynomials. AB - We present a new derivation of the asymptotic correction for bias in the estimate of information from a finite sample. The new derivation reveals a relationship between information estimates and a sequence of polynomials with combinatorial significance, the exponential (Bell) polynomials, and helps to provide an understanding of the form and behavior of the asymptotic correction for bias. PMID- 11112256 TI - Relating macroscopic measures of brain activity to fast, dynamic neuronal interactions. AB - In this article we used biologically plausible simulations of coupled neuronal populations to address the relationship between phasic and fast coherent neuronal interactions and macroscopic measures of activity that are integrated over time, such as the BOLD response in functional magnetic resonance imaging. Event related, dynamic correlations were assessed using joint peristimulus time histograms and, in particular, the mutual information between stimulus-induced transients in two populations. This mutual information can be considered as an index of functional connectivity. Our simulations showed that functional connectivity or dynamic integration between two populations increases with mean background activity and stimulus-related rate modulation. Furthermore, as the background activity increases, the populations become increasingly sensitive to the intensity of the stimulus in terms of a predisposition to transient phase locking. This reflects an interaction between background activity and stimulus intensity in producing dynamic correlations, in that background activity augments stimulus-induced coherence modulation. This is interesting from a computational perspective because background activity establishes a context that may have a profound effect on event-related interactions or functional connectivity between neuronal populations. Finally, total firing rates, which subsume both background activity and stimulus-related rate modulation, were almost linearly related to the expression of dynamic correlations over large ranges of activities. These observations show that under the assumptions implicit in our model, rate-specific metrics based on rate or coherence modulation may be different perspectives on the same underlying dynamics. This suggests that activity (averaged over all peristimulus times), as measured in neuroimaging, may be tightly coupled to the expression of dynamic correlations. PMID- 11112257 TI - Analysis of pointing errors reveals properties of data representations and coordinate transformations within the central nervous system. AB - The execution of a simple pointing task invokes a chain of processing that includes visual acquisition of the target, coordination of multimodal proprioceptive signals, and ultimately the generation of a motor command that will drive the finger to the desired target location. These processes in the sensorimotor chain can be described in terms of internal representations of the target or limb positions and coordinate transformations between different internal reference frames. In this article we first describe how different types of error analysis can be used to identify properties of the internal representations and coordinate transformations within the central nervous system. We then describe a series of experiments in which subjects pointed to remembered 3D visual targets under two lighting conditions (dim light and total darkness) and after two different memory delays (0.5 and 5.0 s) and report results in terms of variable error, constant error, and local distortion. Finally, we present a set of simulations to help explain the patterns of errors produced in this pointing task. These analyses and experiments provide insight into the structure of the underlying sensorimotor processes employed by the central nervous system. PMID- 11112258 TI - Modeling selective attention using a neuromorphic analog VLSI device. AB - Attentional mechanisms are required to overcome the problem of flooding a limited processing capacity system with information. They are present in biological sensory systems and can be a useful engineering tool for artificial visual systems. In this article we present a hardware model of a selective attention mechanism implemented on a very large-scale integration (VLSI) chip, using analog neuromorphic circuits. The chip exploits a spike-based representation to receive, process, and transmit signals. It can be used as a transceiver module for building multichip neuromorphic vision systems. We describe the circuits that carry out the main processing stages of the selective attention mechanism and provide experimental data for each circuit. We demonstrate the expected behavior of the model at the system level by stimulating the chip with both artificially generated control signals and signals obtained from a saliency map, computed from an image containing several salient features. PMID- 11112259 TI - Asymptotic convergence rate of the EM algorithm for gaussian mixtures AB - It is well known that the convergence rate of the expectation-maximization (EM) algorithm can be faster than those of convention first-order iterative algorithms when the overlap in the given mixture is small. But this argument has not been mathematically proved yet. This article studies this problem asymptotically in the setting of gaussian mixtures under the theoretical framework of Xu and Jordan (1996). It has been proved that the asymptotic convergence rate of the EM algorithm for gaussian mixtures locally around the true solution Theta* is o(e(0. 5-epsilon)(Theta*)), where epsilon > 0 is an arbitrarily small number, o(x) means that it is a higher-order infinitesimal as x --> 0, and e(Theta*) is a measure of the average overlap of gaussians in the mixture. In other words, the large sample local convergence rate for the EM algorithm tends to be asymptotically superlinear when e(Theta*) tends to zero. PMID- 11112260 TI - Incremental active learning for optimal generalization. AB - The problem of designing input signals for optimal generalization is called active learning. In this article, we give a two-stage sampling scheme for reducing both the bias and variance, and based on this scheme, we propose two active learning methods. One is the multipoint search method applicable to arbitrary models. The effectiveness of this method is shown through computer simulations. The other is the optimal sampling method in trigonometric polynomial models. This method precisely specifies the optimal sampling locations. PMID- 11112261 TI - A quantitative study of fault tolerance, noise immunity, and generalization ability of MLPs AB - An analysis of the influence of weight and input perturbations in a multilayer perceptron (MLP) is made in this article. Quantitative measurements of fault tolerance, noise immunity, and generalization ability are provided. From the expressions obtained, it is possible to justify some previously reported conjectures and experimentally obtained results (e.g., the influence of weight magnitudes, the relation between training with noise and the generalization ability, the relation between fault tolerance and the generalization ability). The measurements introduced here are explicitly related to the mean squared error degradation in the presence of perturbations, thus constituting a selection criterion between different alternatives of weight configurations. Moreover, they allow us to predict the degradation of the learning performance of an MLP when its weights or inputs are deviated from their nominal values and thus, the behavior of a physical implementation can be evaluated before the weights are mapped on it according to its accuracy. PMID- 11112262 TI - On the computational complexity of binary and analog symmetric hopfield nets AB - We investigate the computational properties of finite binary- and analog-state discrete-time symmetric Hopfield nets. For binary networks, we obtain a simulation of convergent asymmetric networks by symmetric networks with only a linear increase in network size and computation time. Then we analyze the convergence time of Hopfield nets in terms of the length of their bit representations. Here we construct an analog symmetric network whose convergence time exceeds the convergence time of any binary Hopfield net with the same representation length. Further, we prove that the MIN ENERGY problem for analog Hopfield nets is NP-hard and provide a polynomial time approximation algorithm for this problem in the case of binary nets. Finally, we show that symmetric analog nets with an external clock are computationally Turing universal. PMID- 11112263 TI - Macromolecular cross section and cellular localization: determination by radiation target methods. PMID- 11112264 TI - Fluorescent coupled enzyme assays for D-alanine: application to penicillin binding protein and vancomycin activity assays. AB - D-Alanine (D-Ala) is a ubiquitous constituent of bacterial cell walls. Assays for D-Ala can be used to investigate several aspects of cell wall biosynthesis and the effects of antibiotics on this process. High-sensitivity fluorescent assays for D-Ala were developed in a microtiter plate format based on d-aminoacid oxidase/horseradish peroxidase (DAO/HRP)-coupled reactions. For comparative purposes the classic chromogenic (UV-vis) assay using o-phenylenediamine (OPD) was also adapted to microtiter plates. OPD gave a lower limit of sensitivity of 2 nmol and was linear up to 60 nmol. Two commercially available fluorogenic HRP substrates were then tested in this assay. Amplex Red (AR) gave a lower limit of sensitivity of 2 pmol and was linear up to 400 pmol d-Ala. QuantaBlu (QB) based assays exhibited a lag in their response to D-Ala corresponding to 50 pmol D-Ala. This lag complicated calibration, but could be eliminated by addition of 150 pmol D-Ala to all assays. The QB assays were linear up to 3000 pmol D-Ala and gave a lower limit of sensitivity of 10 pmol. These assays are demonstrated for the characterization of the dd-carboxypeptidase activity of a soluble form of Escherichia coli penicillin-binding protein 5 (PBP 5) against the classic PBP substrate diacetyl-L-Lys-D-Ala-D-Ala. AR and QB based assays gave identical v/E(T) profiles, whereas OPD based assays gave slightly (10%) higher activity. This is consistent with the loss of a small amount of E. coli PBP 5 activity during the dilution necessary prior to its use in the highly sensitive fluorescent assays. These assays were then demonstrated for characterization of vancomycin binding to a D-Ala-D-Ala-based substrate. PMID- 11112265 TI - Determination and bioimaging method for nitric oxide in biological specimens by diaminofluorescein fluorometry. AB - A simple and sensitive assay and a cellular bioimaging method for nitric oxide (NO) were developed using a novel diaminofluorescein DAF-FM and its diacetate. DAF-FM is converted via an NO-specific mechanism to an intensely fluorescent triazole derivative. For the measurement of NO, the triazole derivative of DAF-FM was determined by reversed-phase high-performance liquid chromatography with fluorescence detection. In the presence of 1 microM DAF-FM, the concentrations of NOR-1, an NO donor, in the range of 2-200 nM were linearly related to the fluorescence intensity. This sensitive NO assay enabled us to detect the spontaneous and substance P-induced NO release from isolated porcine coronary arteries, both of which were dependent entirely on the NO synthase activity in vascular endothelial cells. We also obtained fluorescence images of cultured smooth muscle cells of the rat urinary bladder after loading with DAF-FM diacetate. In the cells pretreated with cytokines, the fluorescence intensity increased with time after DAF-FM loading. This increase in the fluorescence intensity was blocked by prior treatment of the muscle cells with an NO synthase inhibitor, N(G)-nitro-l-arginine methyl ester. Therefore, the present novel diaminofluorescein fluorometry should be useful not only for sensitive NO assay, but also for NO imaging in a variety of biological specimens. PMID- 11112266 TI - Development of a green fluorescent protein microplate assay for the screening of chemopreventive agents. AB - Here we develop a rapid, cell-based, functional assay to screen and identify naturally occurring or synthetic chemicals with chemopreventive activity. We constructed a reporter gene that consists of the gene-encoding green fluorescent protein (GFP) under the transcriptional control of the thymidine kinase (TK) promoter adjacent to which concatamerized EpRE regulatory elements were inserted. Human hepatoma HepG2 cells were transfected with the EpRE/TK-GFP reporter plasmid, and clones with low GFP background expression and high tBHQ-induced GFP expression were isolated. These GFP reporter cells were seeded into a 96-well microtiter plate, incubated for 24 h, and then treated with test compounds for an additional 24 h. The GFP level and DNA content (as an internal cell survival control) of cells in the 96-well plate were measured subsequently using a fluorescence plate reader. Known inducers of phase II enzymes, such as tert butylhydroquinone, beta-naphthoflavone, and sulforaphane, significantly increased the GFP level in the HepG2 reporter cells. In an initial screening of a chemical library, we identified a synthetic compound whose inducing ability significantly exceeds (1.6-fold) that of the best currently known phase II enzyme inducers. The experimental results indicate that this cell system makes possible a new high throughput screening approach to identify novel chemopreventive molecules. PMID- 11112267 TI - Direct measurement of human lung cancerous and noncancerous tissues by fourier transform infrared microscopy: can an infrared microscope be used as a clinical tool? AB - We have analyzed very small amounts of human lung cancerous tissues directly by a Fourier transform infrared microscopy (FT-IR-MC). The corrected peak heights (H1045 and H1467) obtained from the bands at 1045 and 1467 cm(-1) due to glycogen and cholesterol were chosen for a quantitative evaluation of the malignancy. We found that the H1045/H1467 ratio is an exceptionally useful factor for discrimination of the cancerous tissues from the noncancerous tissues. If the H1045/H1467 ratio from the measured spectrum is larger than 1.4, we can say with confidence that the tissue contains squamous cell carcinoma or adenocarcinoma at least partially. Furthermore, we carried out the microscopic mapping of the tissues containing both cancerous and noncancerous sections, demonstrating that the color map reflects small changes in the spatial distribution of cancer cells in the tissues. The present method may also be applicable to analysis of other cancers, such as colorectal cancerous tissues in which glycogen level has a critical factor for their malignancy. In addition, since FT-IR-MC costs relatively little and does not require a special operator training for collecting and analyzing the spectra, it seems to be perhaps the apparatus best suited to clinical usage, especially in rather small hospitals. PMID- 11112268 TI - A device for the semiautomatic determination of oxygen-radical absorbance capacity. AB - The oxygen-radical absorbance capacity (ORAC) assay has become a standard method to quantify the antioxidative properties of phytonutrients in fruit and vegetable extracts. However, it is usually not possible to determine directly the contribution of specific phytonutrients to the total ORAC value. Separation of the components in the plant extracts by HPLC followed by ORAC analyses of the column fractions might permit the determination of free radical-scavenging profiles. The accurate determination of ORAC values may require 1 to 2 h/sample. Considering the number of samples that would be generated by a single HPLC separation, a device was constructed which permits up to 45 simultaneous ORAC analyses. Varying degrees of automation were included in the design. Furthermore, since the assay has a Q(10) for peroxyl radical-scavenging of about 3, elevation of the assay temperature from the standard 37 to 47 degrees C significantly reduced the assay times. Relatively simple modifications would allow the apparatus to be used in a variety of time-dependent fluorescence and absorbance assays. PMID- 11112269 TI - Continuous flow displacement immunosensors: a computational study. AB - Numerical modeling has been used to investigate the disparity in performance and sensitivity that has been reported for flow displacement immunosensors based on bead-packed columns, membranes, and capillary tubes. The simulations strongly suggest that the high surface areas in the porous media systems may actually be detrimental to sensor performance because of large numbers of free antibody binding sites. Since the free antibody sites are created during the wash step in which the baseline is established, wash protocols are critical in optimizing the sensitivity for a given displacement sensor. PMID- 11112270 TI - Estimation of protein secondary structure from circular dichroism spectra: inclusion of denatured proteins with native proteins in the analysis. AB - We have expanded our reference set of proteins used in the estimation of protein secondary structure by CD spectroscopy from 29 to 37 proteins by including 3 additional globular proteins with known X-ray structure and 5 denatured proteins. We have also modified the self-consistent method for analyzing protein CD spectra, SELCON3, by including a new selection criterion developed by W. C. Johnson, Jr. (Proteins Struct. Funct. Genet. 35, 307-312, 1999). The secondary structure corresponding to the denatured proteins was approximated to be 90% unordered, owing to the spectral similarity of the denatured proteins and unordered structures. We examined the thermal denaturation of ribonuclease T1 by CD using both the original and expanded sets of reference proteins and obtained more consistent results with the expanded set. The expanded set of reference proteins will be helpful for the determination of protein secondary structure from protein CD spectra with higher reliability, especially of proteins with significant unordered structure content and/or in the course of denaturation. PMID- 11112271 TI - Estimation of protein secondary structure from circular dichroism spectra: comparison of CONTIN, SELCON, and CDSSTR methods with an expanded reference set. AB - We have expanded the reference set of proteins used in SELCON3 by including 11 additional proteins (selected from the reference sets of Yang and co-workers and Keiderling and co-workers). Depending on the wavelength range and whether or not denatured proteins are included in the reference set, five reference sets were constructed with the number of reference proteins varying from 29 to 48. The performance of three popular methods for estimating protein secondary structure fractions from CD spectra (implemented in software packages CONTIN, SELCON3, and CDSSTR) and a variant of CONTIN, CONTIN/LL, that incorporates the variable selection method in the locally linearized model in CONTIN, were examined using the five reference sets described here, and a 22-protein reference set. Secondary structure assignments from DSSP were used in the analysis. The performances of all three methods were comparable, in spite of the differences in the algorithms used in the three software packages. While CDSSTR performed the best with a smaller reference set and larger wavelength range, and CONTIN/LL performed the best with a larger reference set and smaller wavelength range, the performances for individual secondary structures were mixed. Analyzing protein CD spectra using all three methods should improve the reliability of predicted secondary structural fractions. The three programs are provided in CDPro software package and have been modified for easier use with the different reference sets described in this paper. CDPro software is available at the website: http://lamar.colostate.edu/ approximately sreeram/CDPro. PMID- 11112272 TI - Implementation of force differentiation in the immunoassay. AB - A technique has been developed to apply force to the antibody-antigen complex in a solid-phase immunoassay. Force was applied to the immunochemical complex by labeling the secondary antibody with a magnetically susceptible, micrometer-size particle and placing the assay chamber in a magnetic field of defined magnitude and orientation. The force was strong enough to displace weakly bound particles but was not strong enough to rupture the immunochemical complex. The number of particles bound to the surface after applying the differentiation force was related to the analyte concentration, thus an optical detection scheme was developed for counting the number of particles on the surface. The sensitivity of the force differentiation assay was demonstrated to be one to two orders of magnitude higher than conventional solid-phase immunoassay techniques for model protein, virus, and bacterial analytes, with 99% specificity. The enhanced sensitivity of this assay appears to result from lowering the assay background through the identification of weakly adhesive, nonspecific interactions. PMID- 11112273 TI - Real-time monitoring of metabolic reactions by microdialysis in combination with tandem mass spectrometry: hydrolysis of CS-866 in vitro in human and rat plasma, livers, and small intestines. AB - Microdialysis combined with tandem mass spectrometry was used to monitor the rapid hydrolysis of CS-866, a new prodrug-type angiotensin II receptor antagonist, in vitro in rat and human plasma as well as in hepatic and intestinal preparations. No chromatographic separation was conducted, and the ion intensity of CS-866 in MS/MS was directly used to perform data acquisition at intervals not longer than several seconds. A methanol-dialyzing medium, flow rate of dialysate, and adoption of sheath liquid were contrived to facilitate this method of measurement. The use of the methanol-dialyzing medium resulted in the effective extraction of the lipophilic analyte through the dialyzing membrane and a substantial reduction of inorganic substances introduced into the ion source. The calibration curve for CS-866 was linear over a concentration range from 0.2 to 20 microM (R(2) = 0.9998), and the intraassay precision was at an acceptable level of not more than 15% in coefficient of variation percentage. CS-866 was hydrolyzed very rapidly to RNH-6270, the pharmacologically active metabolite, in rat and human plasma and rat liver microsomes, and the hydrolysis proceeded extremely rapidly in human plasma with a half-life of less than several seconds. PMID- 11112274 TI - Cloud-point extraction for the determination of the free fraction of antiepileptic drugs in blood plasma and saliva. AB - A method for the determination of the free fraction of antiepileptic drugs in plasma and saliva was developed. The separation of free and protein-bound fractions was obtained by cloud-point extraction under optimum conditions of pH, surfactant type, and concentration. It is shown that the free fraction of carbamazepine and of phenobarbital in plasma coincides with the drug concentration in saliva. The dependence of the free fraction in plasma on the administered dose was studied. The influence of the simultaneous administration of the two drugs on their free concentrations was revealed: In the presence of carbamazepine the free fraction of phenobarbital is decreased markedly whereas phenobarbital does not influence the behavior of carbamazepine. PMID- 11112275 TI - Noninvasive determination of hemoglobin and hematocrit using a temperature controlled localized reflectance tissue photometer. AB - We performed visible/near-infrared optical measurements on the forearm of human subjects. We conducted four studies: one study using a commercial diffuse reflectance spectrometer, and three studies using a breadboard temperature controlled localized reflectance tissue photometer. Calibration relationships were established between skin reflectance signal and either the reference blood hemoglobin (Hb) concentration or the hematocrit values (Hct). Prediction results were expressed as the prediction correlation coefficient (r(p)) and the standard error for cross-validation prediction (CV-SEP). Using diffuse reflectance measurement, r(p) = 0. 8, CV-SEP = 0.9 g/dL for Hb and r(p) = 0.7, CV-SEP = 3.3% for Hct (n = 40). In a localized reflectance study involving calculating the absorption and scattering coefficients and including effect of change of skin temperature in the calibration model, the best prediction results were r(p) = 0.9, CV-SEP = 0.8 g/dL for Hb and r(p) = 0.8, CV-SEP = 3% for Hct (n = 26). In a second localized reflectance study on a population having diverse skin colors at 34 degrees C, the optimal model led to r(p) = 0.8, CV-SEP = 0.9 g/dL for Hb and r(p) = 0.9, CV-SEP = 2.1% for Hct (n = 28) using the localized reflectance values without deducing the absorption and scattering coefficients. Improvement in the correlation was more noticeable for Hct than for the case of Hb. The photometer was used to screen prospective blood donors with low Hb concentration. It was possible to predict anemic subjects in the limited prospective blood donor population. PMID- 11112276 TI - Protein-ribosome-mRNA display: affinity isolation of enzyme-ribosome-mRNA complexes and cDNA cloning in a single-tube reaction. AB - An enzyme-ribosome-mRNA complex was specifically purified by binding to the immobilized enzyme substrate and the cDNA was cloned in a single-tube reaction by one-step reverse transcription-PCR. The ganglioside GM3, used by sialyltransferase II (ST-II) as a substrate, was coated on a 96-well microtiter plate and ST-II was in vitro transcribed and translated from a cDNA library. The isolation of an enzyme-specific protein-ribosome (PRIME) complex was achieved with as little as 0.1 ng ST-II-specific cDNA in 5 microg of a total plasmid preparation or with the cDNA prepared from sublibraries previously inoculated at a density of 2000 clones/culture well. The affinity purification of the PRIME complex was highly specific for GM3 and did not result in cDNA amplification when a different ganglioside (GM1) was used for coating of the microtiter plate. The amplified cDNA was used for cloning or a second round of ribosome display, providing a fast analysis of enzyme affinity to multiple substrates. PRIME display can be used for host-free cDNA cloning from mRNA or cDNA libraries and for binding site mapping of the in vitro translated protein. The use of a single tube reaction in ligand-coated microtiter plates indicates the versatility of PRIME display for cDNA cloning by automated procedures. PMID- 11112277 TI - A luminescence-based test for determining ornithine decarboxylase activity. AB - A sensitive chemiluminescence-based method for the assay of ornithine decarboxylase (ODC) has been developed. This method, which permits the detection of putrescine (the product of ODC) at a picomolar range, can be used to determine ODC activity in cellular extracts. Extracts are incubated with ornithine and spotted onto p81 phosphocellulose paper strips. After drying, the papers are washed with ammonium hydroxide to remove contaminants, which may interfere with the assay. Putrescine is next eluted from the paper by shaking in an elution buffer containing magnesium sulfate. Partially purified hog kidney diamine oxidase is then used to oxidize putrescine in the eluate. The hydrogen peroxide formed during the oxidation is determined by chemiluminescence using luminol and peroxidase. This simple analytical method has the sensitivity of conventional assays based on the use of radioactive ornithine. PMID- 11112278 TI - Preparation of Na,K-ATPase specifically modified on the anti-fluorescein antibody inaccessible site by fluorescein 5'-isothiocyanate. AB - Specific labeling is required for energy transfer measurements and to avoid artifacts in the use of fluorophores as reporter groups. Therefore, a method for specific modification by one of the most popular reagents for P-type ATPases (fluorescein 5'-isothiocyanate) has been developed. Sulfhydryl reagents protected against modification of cysteine residues, and treatment with dithiothreitol eliminated a slow doubling of the fluorescence of conventionally modified Na,K ATPase upon dilution that is attributed to disappearance of self-energy transfer. Removal of nonspecifically bound fluorescein was also confirmed by titration of the modified Na, K-ATPase with anti-fluorescein antibody and by time resolution of the fluorescence change when the modified enzyme was mixed with Na(+) in a stopped-flow instrument. The only fluorescence change when specifically modified Na,K-ATPase was mixed with Na(+) was the signal from fluorescein at the antibody inaccessible, substrate-protectable site that reports the conformational change in unphosphorylated enzyme. The magnitude of the fluorescence change reporting the conformational change increased from between 8 and 12% to between 25 and 30% without affecting the kinetic constants estimated from titrations with Na(+) and K(+). The method should be generally applicable to the preparation of specifically labeled P-type pumps for use in kinetic and equilibrium titrations or energy transfer measurements. PMID- 11112279 TI - Two-wavelength fluorescence assay for DNA repair. AB - A simple and reliable quantitative assay for measuring cellular DNA repair capacity has been developed. It is based on the host cell reactivation of the UV irradiated plasmid pEGFP carrying the marker gene for the enhanced green fluorescent protein (EGFP). As a reference we used the plasmid pEYFP carrying the gene for a red-shifted fluorescent protein (EYFP). Both proteins can be excited by visible light with a maximum at 488 nm, but EGFP emits with a maximum at 509 nm, while EYFP emits with a maximum at 527 nm. This makes it possible to monitor the expression of the two genes simultaneously by measuring the fluorescence at two wavelengths. HEK293 cells were cotransfected with a mixture of UV-irradiated pEGFP and undamaged pEYFP. At different time intervals after transfection the fluorescence of EGFP was determined relative to the fluorescence of EYFP to compensate for any differences in the transfection efficiency or other experimental variables. It was used to calculate the number of UV lesions in DNA and hence the repair capacity of the host cells. It was found that HEK293 cells were able to repair approximately 1.4 UV lesions per 1000 nucleotides DNA for 12 h on the average. PMID- 11112280 TI - Application of a fluorescent histone acetyltransferase assay to probe the substrate specificity of the human p300/CBP-associated factor. AB - Histone N-acetyltransferases (HATs) are a group of enzymes which acetylate specific lysine residues in the N-terminal tails of nucleosomal histones to promote transcriptional activation. Recent structural and enzymatic work on the GCN5/PCAF HAT family has elucidated the structure of their catalytic domain and mechanism of histone acetylation. However, the substrate specificity of these enzymes has not been quantitatively investigated. Utilizing a novel microplate fluorescent HAT assay which detects the enzymatic production of coenzyme A (CoA), we have compared the activities of the HAT domains of human PCAF and its GCN5 homologue from yeast and Tetrahymena and found that they have similar kinetic parameters. PCAF was further assayed with a series of different length histone H3 peptide substrates, which revealed that the determinants for substrate recognition lie within a 19-residue sequence. Finally, we evaluated the acetylation of three putative PCAF substrates, histones H3 and H4 and the transcription factor p53, and have determined that histone H3 is significantly preferred over the histone H4 and p53 substrates. Taken together, the fluorescent acetyltransferase assay presented here should be widely applicable to other HAT enzymes, and the results obtained with PCAF demonstrate a strong substrate preference for the N-terminal residues of histone H3. PMID- 11112281 TI - Use of isosbestic point wavelength shifts to estimate the fraction of a precursor that is converted to a given product. AB - An isosbestic point is observed in overlaid spectra when a chromophoric precursor is converted to a product with a different spectrum, so that it is often assumed that an isosbestic point occurs only when the precursor is quantitatively converted to a single product. We show experimentally and by computer simulations that more complex reactions also exhibit isosbestic points and that the wavelength of the isosbestic point may change. Such wavelength changes will occur if either (i) the molar absorbtivity of the precursor changes or (ii) the fraction of the precursor that is converted to multiple products changes. In the latter case, the isosbestic wavelength and molar absorbtivities of the precursor and product can be used to calculate the fraction of the precursor that is converted to products from the relationship, f = epsilon(Precursor)(M)/epsilon(Product)(M), where f is the fractional conversion, epsilon(Precursor)(M) is the molar absorbtvity of the precursor, and epsilon(Product)(M) is the molar absorbtivity of the product. PMID- 11112282 TI - Microquantity differential display: a strategy for a systematic analysis of differential gene expression with a small quantity of starting RNA. PMID- 11112283 TI - Diphenylamine-aniline-phosphoric acid reagent, a versatile spray reagent for revealing glycoconjugates on thin-layer chromatography plates. PMID- 11112284 TI - Purification of Saccharomcyes cerevisiae mitochondria devoid of microsomal and cytosolic contaminations. PMID- 11112285 TI - Improved method for the production of gold colloid monolayers for use in the phagokinetic track assay for cell motility. PMID- 11112287 TI - Spectrophotometric determination of 3, 4-dihydroxy-2-butanone-4-phosphate synthase activity. PMID- 11112286 TI - Use of sulfobutyl ether beta-cyclodextrin as a vehicle for D-threo-1-phenyl-2 decanoylamino-3-morpholinopropanol-relat ed glucosylceramide synthase inhibitors. PMID- 11112288 TI - One-step directional cloning of blunt-ended polymerase chain reaction products. PMID- 11112289 TI - An ERP investigation of binding and coreference. AB - This study examines the nature of violations in processing one class of binding construction, namely those involving reflexives and their antecedents. When arguments of verbs appear at the point where a syntactic violation is detected, a centroparietal positivity occurs, peaking at 600 ms after the presentation of the stimulus (P600), as is consistent with other types of syntactic anomalies. However, nonarguments in similar sentences fail to elicit the same response. For example, the reflexive in "John's brothers like himself" is in an argument position and elicits the P600 when compared to its grammatical counterpart. The nonargument, participating in the same type of mismatch, "John's brothers like Bill and himself," does not elicit the same positivity. This provides evidence that there are two processes involved in parsing this binding construction, one syntactic and another as yet unidentified, perhaps involving meaning or pragmatics. PMID- 11112290 TI - Language localization in cases of left temporal lobe arachnoid cyst: evidence against interhemispheric reorganization. AB - We investigated whether left-hemisphere arachnoid cysts lead to reorganization of the language function using PET. A group analysis demonstrated that patients showed no more right-hemisphere activation than a matched control group. Several patients had clear language localizations in the left hemisphere during language comprehension; none of the patients showed right-hemisphere activation. We conclude that left-hemisphere tissue must suffer considerable compromise before reorganization of language into the right hemisphere becomes necessary. Language activations within the left hemisphere are clearly displaced. This is consistent with mere physical displacement in some patients rather than reorganization within the left hemisphere; in others intrahemispheric reorganization cannot be excluded. PMID- 11112291 TI - Preserved visual lexicosemantics in global aphasia: a right-hemisphere contribution? AB - Extensive testing of a patient, GP, who encountered large-scale destruction of left-hemisphere (LH) language regions was undertaken in order to address several issues concerning the ability of nonperisylvian areas to extract meaning from printed words. Testing revealed recognition of superordinate boundaries of animals, tools, vegetables, fruit, clothes, and furniture. GP was able to distinguish proper names from other nouns and from nonwords. GP was also able to differentiate words representing living things from those denoting nonliving things. The extent of LH infarct resulting in a global impairment to phonological and syntactic processing suggests LH specificity for these functions but considerable right-hemisphere (RH) participation in visual lexicosemantic processing. The relative preservation of visual lexicosemantic abilities despite severe impairment to all aspects of phonological coding demonstrates the importance of the direct route to the meaning of single printed words. PMID- 11112292 TI - Lexical access of function versus content words. AB - There has been a simmering debate as to whether evidence exists for differential processes of lexical access for function and content words. This has centered around the frequency effect (higher word frequency reducing access times for content words but not function words). Previous work has used the lexical decision paradigm, which has been shown to reflect more than lexical access times. We measured naming times for words in sentences read for meaning. Our findings confirm that lexical access for function words is indeed faster than for content words as predicted by neurolinguistic theory and electrophysiological evidence, but that this difference can be attributed to word predictability (Cloze value) and word familiarity (log frequency). We also show that differences in frequency effect for the two word types holds only for the lower frequency words and not at all for the higher frequency words. We discuss the implications of the results for neurolinguistic theory. PMID- 11112293 TI - An aphasia database on the internet: a model for computer-assisted analysis in aphasiology. AB - A web-based software model was developed as an example for data mining in aphasiology. It is used for educating medical and engineering students. It is based upon a database of 254 aphasic patients which contains the diagnosis of the aphasia type, profiles of an aphasia test battery (Aachen Aphasia Test), and some further clinical information. In addition, the cerebral lesion profiles of 147 of these cases were standardized by transferring the coordinates of the lesions to a 3D reference brain based upon the ACPC coordinate system. Two artificial neural networks were used to perform a classification of the aphasia type. First, a coarse classification was achieved by using an assessment of spontaneous speech of the patient which produced correct results in 87% of the test cases. Data analysis tools were used to select four features of the 30 available test features to yield a more accurate diagnosis. This classifier produced correct results in 92% of the test cases. The neural network approach is similar to grouping performed in group studies, while the nearest-neighbor method shows a design more similar to case studies. It finds the neurolinguistic and the lesion data of patients whose AAT profiles are most similar to the user's input. This way lesion profiles can be compared to each other interindividually. The Aphasia Diagnoser is available on the Web address http://fuzzy.iau.dtu.dk/aphasia.nsf and thus should facilitate a discussion about the reliability and possibilities of data-mining techniques in aphasiology. PMID- 11112294 TI - Comprehension and storage of four serially presented radio news stories by mild aphasic subjects. AB - The present study investigated aphasic subjects' ability to comprehend and store serially presented discourse. Sixteen mild aphasic subjects, eight age-matched normals, and eight younger normals listened to four serially presented radio news stories and a single radio news story. Half of the aphasic subjects performed as well as age-matched normals in a single-news-story comprehension task. However, they demonstrated a drastic deterioration in performance when asked to listen to a series of four news stories. Age-matched normals, and aphasic subjects, to a lesser extent, showed an impairment in the comprehension and storage of the news story heard last in a series of four news stories. These results were discussed in terms of the comprehension and storage resources of working memory. PMID- 11112295 TI - Grammatical encoding in aphasia: evidence from a "processing prosthesis". AB - Agrammatic aphasia is characterized by severely reduced grammatical structure in spoken and written language, often accompanied by apparent insensitivity to grammatical structure in comprehension. Does agrammatism represent loss of linguistic competence or rather performance factors such as memory or resource limitations? A considerable body of evidence supports the latter hypothesis in the domain of comprehension. Here we present the first strong evidence for the performance hypothesis in the domain of production: an augmentative communication system that markedly increases the grammatical structure of agrammatic speech while providing no linguistic information, functioning merely to reduce on-line processing demands. PMID- 11112296 TI - The selective impairment of phonological processing in speech production. AB - We report the naming performance of a patient (DM) with a fluent progressive aphasia who made phonological errors in all language production tasks. The pattern of errors in naming was strikingly clear: DM made very many phonological errors that resulted almost always in nonword responses. The complete absence of semantic errors and the very low ratio of formal errors relative to nonword errors (1.6:30.3) in DM's performance are discussed in the context of recent claims about the nature of naming deficits in fluent aphasics. We argue that DM's performance makes highly improbable the claim that fluent aphasia results from global lesions affecting all levels of the lexical access system equally. PMID- 11112297 TI - Dissociation of semantic and phonological errors in naming. AB - We report the naming performance of a fluent aphasic, DP, who shows a striking dissociation between semantic and phonological (nonword) errors: he produced numerous semantic errors but virtually no phonological errors. DP's pattern of performance is the reverse of that reported for patient DM (Caramazza, Papagno, & Ruml, 2000), who only made phonological errors in a naming task. These patterns of performance are inconsistent with the proposal by Dell, Schwartz, Martin, Saffran, and Gagnon (1997) that the naming deficit in fluent aphasia is the result of global damage to all levels of the lexical access system and support instead the hypothesis that brain damage can selectively disrupt distinct subcomponents of the lexical processing system. PMID- 11112299 TI - Theoretical and experimental neuropsychology (TENNET XII) PMID- 11112298 TI - Kicking over the traces: A note in response to Zurif and Pinango (1999). AB - Zurif and Pinango (1999) claimed that they excluded the four agrammatic patients reported by Druks and Marshall (1991) from their review article because two of the patients were nonnative speakers of Hebrew and because the Hebrew sentences we used in our investigations were ungrammatical. In Druks and Marshall (1991) we have shown that the presence or the absence of a trace in two types of Hebrew passives had no effect on the patients' performance. Two patients, without comprehension deficits, performed equally well on both types of passives and two patients, with comprehension deficits, were equally impaired on both types. We remind Zurif and Pinango of our previous response to the claims of ungrammaticality of our materials (Druks & Marshall, 1992) and argue that there were no justifiable reasons for excluding these cases from the review. We also comment on Zurif and Pinango's (1999) and Grodzinsky's (2000) new proposal that the association of agrammatic comprehension should be with Broca's aphasia and not with agrammatism. PMID- 11112300 TI - Stability in Colloidal Mixtures Containing Particles with a Large Disparity in Size. AB - An experimental approach, based on turbidity measurements, is proposed for studies of the stability in colloidal mixtures containing particles with large disparity in size. The main advantage of this approach is that it permits investigations even under conditions of comparable particle number concentrations of the two colloidal populations. Binary mixtures containing a poly(vinyl acetate) (PVAc) latex and a Ludox AS-40 silica sol were investigated. The silica particles were much smaller than the latex ones. The experimental stability factors were compared with the theoretical values computed on the basis of the Kihira-Ryde-Matijevic model (J. Chem. Soc., Faraday Trans. 88(16), 2379 (1992)) for interaction between spherical particles with unevenly distributed surface charges. All the experimental results support the idea that, even when both sols are negatively charged, the small silica particles are adsorbed onto the latex surface. Under these conditions, the heteroaggregates, which are composed of PVAc cores surrounded with silica particles, can be modeled as PVAc particles having "modified" surface characteristics (i.e., average Stern potential and varying extents of the surface charge segregation). Copyright 2001 Academic Press. PMID- 11112301 TI - Electro-Viscous Effects on Liquid Flow in Microchannels. AB - The presence of the electrical double layer near a solid-liquid interface results in the electro-viscous effect on pressure-driven liquid flow through microchannels. The objective of this paper is to examine the magnitude of the additional flow resistance caused by the electrokinetic effect in microchannels. Deionized ultrafiltered water, 10(-4) and 10(-2) M aqueous KCl solutions, 10(-4) M AlCl(3) solution, and 10(-4) M LiCl solution were used as the testing liquids. Carefully designed flow measurements were conducted in three silicon microchannels with a height of 14.1, 28.2, and 40.5 um, respectively. The measured dP/dx for the pure water, the 10(-4) M KCl solution, and the 10(-4) M LiCl solution was found to be significantly higher than the prediction of the conventional laminar flow theory at the same Reynolds number. Such a high flow resistance and the resulting high apparent viscosity strongly depend on the channel's height, the ionic valence, and the concentration of the liquids. The zeta potentials for the liquid-solid systems were calculated by using the measured streaming potential data. The experimentally determined dP/dx approximately Re relationships were compared with the predictions of a theoretical electro-viscous flow model, and a good agreement was found for pure water, 10(-4) M KCl solution, and 10(-4) MAlCl(3) solution systems. The present electrokinetic flow model cannot interpret the flow characteristics of the LiCl solution. Copyright 2001 Academic Press. PMID- 11112302 TI - Stability Characteristics of Dispersed Oil Droplets Prepared by the Microchannel Emulsification Method. AB - A novel emulsification method, microchannel (MC) emulsification, was developed for making monodispersed regular-sized droplets in our laboratory. An oil-in water dispersed system, in which phosphate buffer was used as the continuous phase, sodium dodecyl sulfate (SDS) as the surfactant, and clove oil as the dispersed phase, was prepared by this technique. The average diameter of oil droplets was about 20 um, with a narrow size distribution. The stability characteristics of the dispersed oil droplets were investigated by an optical microscope and kinetic light scattering method. The stability of the dispersed oil droplets depended on the SDS concentration. When the SDS concentration was above the critical micelle concentration (CMC), the turbidity of the dispersed solution sharply increased at the initial stage. Optical microscopic observation has confirmed that a part of the oil droplets broke up with time, and submicrometer droplets appeared. On the other hand, when the SDS concentration was below the CMC, the turbidity of the dispersed solution had little change in the initial stage, showing that the oil droplets were very stable. The effect of ion concentration was also examined. The results showed that the stability of the oil droplets depended on the balance of the Van der Waals attraction and electrical repulsion between the oil droplets in low ion concentration. Copyright 2001 Academic Press. PMID- 11112303 TI - A Study on the Adsorption Characteristics of Orthophosphates on Rutile-Type Titanium Dioxide in Aqueous Solutions. AB - Rutile-type titanium dioxide is widely used as a pigment for paint, coating ink, paper, plastic products, and cards because of its very whiteness and outstanding hiding property. It has two weak properties to be improved, however, one being its coagulation in compounding and the other its decreasing whiteness owing to poor heat resistance. To solve these problems, a study on the treatment of titanium dioxide surfaces by adsorption technology has been performed. Experiments have been carried out to establish the adsorption isotherms of orthophosphates on titanium dioxide and to investigate the effect of organic solvents on adsorption. It is shown that the adsorption isotherms vary with pH. A Freundlich adsorption isotherm is suitable for the acidic and basic regions, while a Langmuir adsorption isotherm is suitable for the region of pH 5-8 where a maximum adsorption has been achieved. In aqueous solutions containing organic solvents, the adsorption was strongest with aqueous solutions containing 1 wt% toluene and weakest with those containing 1 wt% ethanol. Among the alcohols used, the adsorption was strongest with the aqueous solution containing 1 wt% butanol and weakest with that containing 1 wt% ethanol, thus showing a correlation with the molecular weight of the alcohol. Copyright 2001 Academic Press. PMID- 11112304 TI - Uranium(VI) Sorption Complexes on Montmorillonite as a Function of Solution Chemistry. AB - We have investigated the effect of changes in solution chemistry on the nature of uranyl sorption complexes on montmorillonite (SAz-1) at different surface coverages (1.43-53.6 umol/g). Uranyl uptake onto SAz-1 between pH 3 and 7 was determined in both titration and batch-mode experiments. These pH values result in solutions that contain a range of monomeric and oligomeric aqueous uranyl species. Continuous-wave and time-resolved emission spectroscopies were used to investigate the nature of U(VI) sorbed to SAz-1. A discrete set of uranyl surface complexes has been identified over a wide range of pH values at these low to moderate coverages. For all samples, two surface complexes are detected with spectral characteristics commensurate with an inner-sphere complex and an exchange-site complex; the relative abundance of these two species is similar over these pH values at low coverage (1.43-2.00 umol/g). In addition, surface species having spectra consistent with polymeric hydroxide-like sorption complexes form at the moderate coverages ( approximately 34-54 umol/g), increasing in abundance as the capacity of the amphoteric surface sites is exceeded. Furthermore, a species with spectral characteristics anticipated for an outer-sphere surface complex is observed for wet paste samples at low pH (3.7 4.4) and both low ( approximately 2 umol/g) and moderate ( approximately 40 umol/g) coverage. There are only subtle differences in the nature of sorption complexes formed at different pH values but similar coverages, despite markedly different uranyl speciation in solution. These results indicate that the speciation in the solution has minimal influence on the nature of the sorption complex under these experimental conditions. The primary control on the nature and abundance of the different uranyl sorption complexes appears to be the relative abundance and reactivity of the different sorption sites. Copyright 2001 Academic Press. PMID- 11112305 TI - Noise on Fluorescence Correlation Spectroscopy. AB - The time dependence of the noise and the signal-to-noise (SN) ratio of the fluorescence correlation spectroscopy (FCS) autocorrelation function is obtained from replica measurements of standard dextran solutions. The noise dependence on the delay time is fitted by a hyperbolic function with two fitting parameters. The dependence of these parameters on concentration, fluorescence intensity, and accumulation time is obtained experimentally. The behavior of SN at zero delay time agrees well with the theoretical predictions reported in the literature. The obtained data are useful for the quantitative evaluation of the FCS data fits, as well as for simulation of the FCS autocorrelation functions. Copyright 2001 Academic Press. PMID- 11112306 TI - Electrochemical Investigation of the Commercial Iron Sulfide Stick-Interaction with Complexing Ligands. AB - The present work aims to examine iron sulfide for its interfacial properties in aqueous media containing different ionic/complexing agents. The primary experimental investigation is the potentiometric response of the commercial iron sulfide stick in various aqueous media supplemented by cyclic voltammetry and UV visible and IR spectroscopy. The significance of employing the commercial iron sulfide stick is that it is quite stable under ordinary conditions and possesses appreciable conductivity for employing it as an electrode. Copyright 2001 Academic Press. PMID- 11112307 TI - Effects of Carboxylic Acids on the Crystallization of Calcium Carbonate. AB - The effects of seven carboxylic acids, i.e., acrylic acid, maleic acid, tartaric acid, malic acid, succinic acid, and citric acid, on CaCO(3) crystallization were studied using the unseeded pH-drift method along with a light-scattering technique. Experiments were started by mixing solutions of CaCl(2) and NaHCO(3) in the presence or absence of additives. The crystallization was studied by recording the decrease in pH resulting from the reaction Ca(2+)+HCO(3)(-)- >CaCO(3)+H(+). A given amount of carboxylic acid was added to the solution of CaCl(2) or NaHCO(3) before mixing the reactants. The pH profiles obtained in the case of the CaCl(2) solution containing an additive were similar to those for the NaHCO(3) solution containing one, and when an additive was added after the onset of crystallization, the growth of CaCO(3) immediately stopped. The light scattering observations, in all cases, indicated that CaCO(3) nucleation occurred at 10-20 s after mixing of the reactants. The results indicated that the nucleation of CaCO(3) was not influenced by the presence of carboxylic acids, but CaCO(3) crystal growth was reduced by their adsorption to the surface of the CaCO(3) crystals. These phenomena were explained by assuming a stronger affinity of the carboxylic acids for CaCO(3) particles than for the free Ca(2+) ions in solution. The crystallization of CaCO(3) in the presence of additives was divided into three stages: nucleation, growth incubation, and growth periods. Copyright 2001 Academic Press. PMID- 11112308 TI - Detachment of Particles from Surfaces: An AFM Study. AB - We have measured interactions between hydrophilic and hydrophobic surfaces in an aqueous medium at various pH and ionic strengths as well as in some organic solvents using atomic force microscopy and analyzed them in terms of particle adhesion and detachment from surfaces. In hydrophilic systems the forces observed were found to be well described by DLVO theory at large separation distances. Very long range hydrophobic forces were not observed in hydrophilic-hydrophobic systems. Nevertheless, the jump into contact was found to occur at distances greater that those predicted by just van der Waals attraction. The interaction between two hydrophobic surfaces was dominated by the long-range attraction due to hydrophobic forces. This interaction was found to be sensitive to the type of substrate as well as to the pH and electrolyte concentration. Measured pull-off forces showed poor reproducibility. However, average values showed clear trends and were used to estimate interfacial energies or work of adhesion for all systems studied by means of the Derjaguin approximation. These values were compared to those calculated by the surface tension component theory using the acid-base approach. Good qualitative agreement was obtained, giving support for the usefulness of this approach in estimating interfacial energies between surfaces in liquid media. A comparison of the measured adhesion force with hydrodynamic detachment experiments showed good qualitative agreement. Copyright 2001 Academic Press. PMID- 11112309 TI - Dissolution Kinetics, Selective Leaching, and Interfacial Reactions of a Bioglass Coating Enriched in Alumina. AB - Bioglass coatings are interesting for developing a direct bond between prostheses and bone. But the high solubility of these materials limits their application. The addition of alumina can be used to control their solubility, but may inhibit the bonding mechanisms. In this paper, we study a bioglass in the SiO(2)-Na(2)O CaO-P(2)O(5)-K(2)O-Al(2)O(3)-MgO system. After delays of implantation from 2 to 12 months, the bioglass/bone interface is characterized by energy-dispersive X ray spectroscopy coupled with scanning transmission electron microscopy. Bioglass dissolution can be decomposed into three steps with selective leaching. Results show that, at 2 months after implantation, the bioglass is composed of Al, Si, Ca, and P. Alumina addition increases the coating stability without inhibiting the bonding process. Complex physicochemical reactions take place at the bioglass periphery. The coating bonds to bone through a Ca-P layer on top of a pure Si rich layer. These phenomena are associated with bioactivity properties, which occur for up to 6 months. After 12 months, the bioglass is composed of silicon. Copyright 2001 Academic Press. PMID- 11112310 TI - Effect of gamma Irradiation on Latex Film Dissolution. AB - Latex films have been prepared by annealing pyrene (Py)-labeled poly(methyl methacrylate)-polyisobutylene particles at the glass transition temperature (100 degrees C). These films were then irradiated by gamma rays from (60)Co in a gamma cell at room temperature at various dose rates (rad/h). Before dissolution the films were annealed at 200 degrees C for a 30 min time interval to complete the film formation process. Steady-state fluorescence techniques were used to monitor the dissolution of irradiated latex films. Dissolution of films in a chloroform heptane (80%-20%) mixture was monitored in real time by the Py fluorescence intensity change. Relaxation constants, k(0), and dissolution coefficients, D(d), of polymer chains were measured. Two different regimes of D(d) values were observed during dissolution, which are related to two different molecular weight distributions caused by scission and branching of polymer chains when they were irradiated and annealed. Copyright 2001 Academic Press. PMID- 11112311 TI - Study of Surface-Enhanced Infrared Spectroscopy. AB - The dependence of surface-enhanced infrared absorption (SEIRA) on structure and vibration modes of chemisorbates was studied in a reflection-absorption mode by employing self-assembled monolayers of 4-pyridinethiol and 4-nitrothiophenol formed on Au island films deposited on glass substrates. The present work shows that both the electromagnetic and chemical effects contribute to the total SEIRA enhancement, with the former playing a predominant role. Moreover, the bands due to different vibration modes give rise to distinctly different magnitudes of enhancement, suggesting the presence of a chemical origin in the enhanced IR absorption. Copyright 2001 Academic Press. PMID- 11112312 TI - Study of Surface-Enhanced Infrared Spectroscopy. AB - In this paper we report a versatile and effective strategy to attain strong surface-enhanced infrared absorption by employing a sandwich system consisting of metal island films and self-assembled monolayers (SAMs) of 4-pyridinethiol. The observed larger enhancement factor stems from coupling of the electric fields induced by excitation of the surface plasmon resonance of the overlayer and underlayer Au island films, and from enhanced chemical interactions of the Au island films and the pyridine molecule in the sandwiched structures, compared to the corresponding SAM-Au configuration. Copyright 2001 Academic Press. PMID- 11112313 TI - Uptake of Au(III) Ions by Aluminum Hydroxide and Their Spontaneous Reduction to Elemental Gold (Au(0)). AB - The behavior of AuCl(4)(-) ions during the formation of aluminum hydroxide at pH 6 was examined. With an increase in NaCl concentration, the content of gold taken up by aluminum hydroxide decreased, suggesting that chloro-hydroxy complexes of Au(III) ion were taken up due to the formation of Al-O-Au bonds. It was found unexpectedly that the Au(III) ions taken up were spontaneously reduced to elemental gold without addition of a specific reducing reagent and then colloidal gold particles were formed. The mechanisms for the uptake of Au(III) ions by aluminum hydroxide and for their spontaneous reduction are discussed. Copyright 2001 Academic Press. PMID- 11112314 TI - Asymptotic Particle Size Distributions Attained during Coagulation Processes. AB - The effects of the collision kernel on the self-preserving size distribution and on the gelation phenomenon of aerosol coagulation were investigated. An analytical solution for the asymptotic width of log-normally preserving size distribution during coagulation was obtained as a function of the degree of homogeneity using arbitrary shape of homogeneous collision kernels. From the solution obtained, it was shown that when the degree of homogeneity is larger than 1, self-preserving size distribution does not exist, and gelation occurs. A very accurate numerical coagulation simulation method, the sectional method, was also used for calculating the self-preserving particle size distribution for some specific classes of coagulation kernels and the results were compared with the analytical solution obtained by the log-normal method. Copyright 2001 Academic Press. PMID- 11112315 TI - The Electrostatic Interaction of a Charged Particle with a Surface: The Effect of Surface Charge Rearrangement. AB - In this paper we investigate the electric interaction between a charged particle and a surface in which the charged ions are capable of moving in response to the electric potential disturbance caused by the approach of the charged particle. Such surfaces include ionic surfactants distributed in air-water interface and charged lipids in bilayer membranes. On the basis of the mean field theory, the free energy of the system, which includes the electrostatic internal energy and the entropy of the mobile ions and surface ions, can be written down. The surface charge-potential relation is then derived by the calculus of variation. When the potential disturbance is small enough, a linear charge regulation model is obtained. The interaction energy associated with a long rod parallel to the interface is studied and an analytical expression is obtained. When a rod approaches an oppositely charged surface, the interaction can change from attraction to repulsion, depending on the ratio of the characteristic regulation length to the Debye length. At low surface charge density, the surface behaves as under the condition of constant charge density and acts as that of constant potential for high enough charge density. Copyright 2001 Academic Press. PMID- 11112316 TI - Surface Complexation Modeling of K(+), NO(3)(-), SO(4)(2-), Ca(2+), F(-), Co(2+), and Cr(3+) Ion Adsorption on Silica Gel. AB - Surface complex formation of K(+), NO(3)(-), SO(4)(2-), Ca(2+), F(-), Co(2+), and Cr(3+) ions was determined on the surface of silica gel. Experimental data obtained by acid-base titration of suspensions were interpreted in terms of the triple-layer model. The value of the deprotonation constant of surface OH could be determined precisely but the protonation constant was rather uncertain. The logarithms of ion pair formation constants for K(+), NO(3)(-), Ca(2+), and SO(4)(2-) adsorbed in the beta-plane are log K(ipM,X) approximately 0, therefore these species can be considered inert ions in the investigated pH range. F(-), Co(2+), and Cr(3+) ions were found to be strongly sorbed in the o-plane. In order to provide a good fit and to obtain parameters independent of their initial values, all possible equilibrium must be accounted for in the models. Copyright 2001 Academic Press. PMID- 11112317 TI - Effects of Dissipation on Contact Angle Measurements Using a Dynamic Method. AB - The contribution of viscous friction, slippage, the Marangoni effect, and three phase contact line resistance is considered in the explanation of contact angle measurements using a dynamic method. It is shown that the viscous friction of the liquid and the specific resistance of the three-phase contact line are the basic dissipative forces governing the rheology, while the slip on the solid-liquid interface is not important for this kind of experiment. Since the method is applied to protein solutions, it is demonstrated that the Marangoni effect of the proteins on the liquid-air surface plays an important role in the right juxtaposition of the theoretical and experimental results. Proper application of the theory allows calculation of both receding and equilibrium contact angles from the dropping time measurements as well as their dependence on the concentration of the protein solutions. Copyright 2001 Academic Press. PMID- 11112318 TI - Dynamic Electrophoretic Mobility of a Soft Particle. AB - A theory of the dynamic electrophoretic mobility of a spherical soft particle (that is, a polyelectrolyte-coated spherical particle) in an oscillating electric field is presented. In the absence of the polyelectrolyte layer a spherical soft particle becomes a spherical hard particle, while in the absence of the particle core it tends to a spherical polyelectrolyte. The present theory thus covers two extreme cases, that is, dynamic electrophoresis of hard particles and that of spherical polyelectrolytes. Simple analytic mobility expressions are derived. It is shown how the dynamic electrophoretic mobility of a soft particle depends on the volume charge density distributed in the polyelectrolyte layer, on the frictional coefficient characterizing the frictional forces exerted by the polymer segments on the liquid flow in the polyelectrolyte layer, on the particle size, and on the frequency of the applied oscillating electric field. Copyright 2001 Academic Press. PMID- 11112319 TI - Numerical Study of a Lifshitz-Slyozov-like Metastable Dewetting Model. AB - A Lifshitz-Slyozov-type of model of the coalescence of dry zones in the process of metastable dewetting proposed in V. S. Mitlin (J. Colloid Interface Sci. 227, 371, 2000) is applied to simulating the evolution of the probability distribution of holes by sizes. For the model considered, the average size of a hole increases linearly with time, the surface fraction of holes grows as the time squared, and the maximum value of the probability distribution decreases as the inverse square root of time. Copyright 2001 Academic Press. PMID- 11112320 TI - Obtaining the Osmotic Pressure of Electrostatically Stabilized Colloidal Dispersions from Frontal Ultrafiltration Experiments. AB - A new method for determining the osmotic pressure of electrostatically stabilized colloidal particles from frontal ultrafiltration experiments has been developed. The method is based on a reverse calculation utilizing a previously developed analysis of frontal ultrafiltration (W. R. Bowen and F. Jenner, Chem. Eng. Sci. 50, 1707, 1995; W. R. Bowen and P. M. Williams, Biotechnol. Bioeng. 50, 125, 1996). The method has the following advantages over conventional membrane osmometer measurements: (i) only very simple apparatus is required, (ii) results are obtained quickly, (iii) only small sample quantities are required, and (iv) only dilute initial samples are needed, as the ultrafiltration process creates a concentrated solution at the membrane, making sample preparation a simple task. A comparison of the osmotic pressure determined by this new method with previously measured osmotic pressure data shows excellent agreement. The approach demonstrates how analysis of process data may provide quantitative information on the interactions of concentrated colloidal systems. Copyright 2001 Academic Press. PMID- 11112321 TI - Notch signaling: from the outside in. PMID- 11112322 TI - Protein kinase C-related kinase 2 phosphorylates the protein synthesis initiation factor eIF4E in starfish oocytes. AB - Phosphorylation of eIF4E is required for protein synthesis during starfish oocyte maturation. The activity of protein kinase C-related kinase 2 (PRK2) increases prior to the phosphorylation of eIF4E (G. Stapleton et al., 1998, Dev. Biol. 193, 34-46). We investigate here whether eIF4E is activated by PRK2. A 3.5-kb eIF4E clone isolated from starfish cDNA is 57% identical to human eIF4E and contains the putative phosphorylation site serine-209. The serine-209 environment (SKTGS(209)MAKSRF) is similar to the consensus sequence of the phosphorylation site of protein kinase C and related kinases. A starfish eIF4E fusion protein (GST-4E) was phosphorylated in vitro by PRK2 in the presence of 1,2-diolyl-sn glycerol 3-phosphate. In contrast, replacing the GST-4E serine-209 with an alanine significantly reduced this phosphorylation. Analysis by two-dimensional phosphopeptide mapping reveals a major phosphopeptide in trypsin-digested GST-4E, but not in its serine-209 mutant. Importantly, this major phosphopeptide in GST 4E corresponds to a major phosphopeptide of eIF4E isolated from (32)P-labeled oocytes. Thus, PRK2 may regulate translation initiation during oocyte maturation by phosphorylating the serine-209 residue of eIF4E in starfish. We also demonstrate that high levels of cAMP inhibit the activation of PRK2, eIF4E, and the eIF4E binding protein during starfish oocyte maturation, while PI3 kinase activates these proteins. PMID- 11112323 TI - Multiple roles for four-jointed in planar polarity and limb patterning. AB - Insect cuticles have been a model system for the study of planar polarity for many years and a number of genes required for this process have been identified. These genes organise the polarised arrangement of hairs on the legs, wings, thorax, and abdomen of adult Drosophila. It has previously been shown that four jointed is involved in planar polarity decisions in the eye as well as proximal distal leg and wing development. We now present evidence that four-jointed is expressed in a gradient through the developing wing and show that it is required for planar polarity determination in both the wing and the abdomen. Clones of cells either lacking or ectopically expressing four-jointed cause both autonomous and nonautonomous repolarisation of hairs in these tissues. We propose that the inferred four-jointed expression gradient is important for planar polarity establishment and that local inversions of the gradient by the clones are the probable cause of the observed polarity phenotypes. In addition we observe defects in wing vein development. The subtle phenotypes of mutant flies, and the diverse patterning processes in which it is involved, suggest that four-jointed may act as a modifier of the activity of multiple other signalling factors. PMID- 11112324 TI - Use of large-scale expression cloning screens in the Xenopus laevis tadpole to identify gene function. AB - We have conducted an expression cloning screen of approximately 50, 000 cDNAs from a tadpole stage Xenopus laevis cDNA library to functionally identify genes affecting a wide range of cellular and developmental processes. Fifty-seven cDNAs were isolated for their ability to alter gross tadpole morphology or the expression patterns of tissue-specific markers. Thirty-seven of the cDNAs have not been previously described for Xenopus, and 15 of these show little or no similarity to sequences in the NCBI database. The screen and the identified genes are presented in this paper to demonstrate the power, ease, speed, and flexibility of expression cloning in the X. laevis embryo. Future screens such as this one can be done on a larger scale and will complement the sequence-based screens and genome-sequencing projects which are producing a large body of novel genes without ascribed functions. PMID- 11112325 TI - A gain-of-function mutant of patched dissects different responses to the hedgehog gradient. AB - The Hedgehog (Hh) signal has an inductive role during Drosophila development. Patched is part of the Hedgehog-receptor complex and shows a repressive function on the signaling cascade, which is alleviated in the presence of Hh. Herein, we identify the first dominant gain-of-function allele of patched, Confused (patched(Con)). Analysis of the patched(Con) allele led us to uncover novel features of the reception and function of the Hh signal. At least three different regions of gene expression were identified and a gradient of cell affinities was established in response to Hh. A new state of Cubitus interruptus activity responsible for the activation of araucan and caupolican genes of the iroquois complex, independent of Fused kinase function, was shown. In the disc, patched(Con) behaved like fused mutants and was rescued by Suppressor of fused mutations. However, fused mutants are embryonic lethal while patched(Con) is not, suggesting that Patched could interpret Hedgehog signaling differently in the embryo and in the adult. PMID- 11112327 TI - RNA interference demonstrates a role for nautilus in the myogenic conversion of Schneider cells by daughterless. AB - Schneider SL2 cells activate the myogenic program in response to the ectopic expression of daughterless alone, as indicated by exit from the cell cycle, syncytia formation, and the presence of muscle myosin fibrils. Myogenic conversion can be potentiated by the coexpression of DMEF2 and nautilus with daughterless. In RT-PCR assays Schneider cells express two mesodermal markers, nautilus and DMEF2 mRNAs, as well as very low levels of daughterless mRNA but no twist. Full-length RT-PCR products for nautilus and DMEF2 encode immunoprecipitable proteins. We used RNA-i to demonstrate that both endogenous nautilus expression and DMEF2 expression are required for the myogenic conversion of Schneider cells by daughterless. Coexpression of twist blocks conversion by daughterless but twist dsRNA has no effect. Our results indicate that Schneider cells are of mesodermal origin and that myogenic conversion with ectopic expression of daughterless occurs by raising the levels of daughterless protein sufficiently to allow the formation of nautilus/daughterless heterodimers. The effectiveness of RNA-i is dependent upon protein half-life. Genes encoding proteins with relatively short half-lives (10 h), such as nautilus or HSF, are efficiently silenced, whereas more stable proteins, such as cytoplasmic actin or beta-galactosidase, are less amenable to the application of RNA-i. These results support the conclusion that nautilus is a myogenic factor in Drosophila tissue culture cells with a functional role similar to that of vertebrate MyoD. This is discussed with regard to the in vivo functions of nautilus. PMID- 11112326 TI - DNA replication defects delay cell division and disrupt cell polarity in early Caenorhabditis elegans embryos. AB - In early Caenorhabditis elegans embryos, asymmetric cell divisions produce descendants with asynchronous cell cycle times. To investigate the relationship between cell cycle regulation and pattern formation, we have identified a collection of embryonic-lethal mutants in which cell divisions are delayed and cell fate patterns are abnormal. In div (for division delayed) mutant embryos, embryonic cell divisions are delayed but remain asynchronous. Some div mutants produce well-differentiated cell types, but they frequently lack the endodermal and mesodermal cell fates normally specified by a transcriptional activator called SKN-1. We show that mislocalization of PIE-1, a negative regulator of SKN 1, prevents the specification of endoderm and mesoderm in div-1 mutant embryos. In addition to defects in the normally asymmetric distribution of PIE-1, div mutants also exhibit other losses of asymmetry during early embryonic cleavages. The daughters of normally asymmetric divisions are nearly equal in size, and cytoplasmic P-granules are not properly localized to germline precursors in div mutant embryos. Thus the proper timing of cell division appears to be important for multiple aspects of asymmetric cell division. One div gene, div-1, encodes the B subunit of the DNA polymerase alpha-primase complex. Reducing the function of other DNA replication genes also results in a delayed division phenotype and embryonic lethality. Thus the other div genes we have identified are likely to encode additional components of the DNA replication machinery in C. elegans. PMID- 11112328 TI - A role for Xlim-1 in pronephros development in Xenopus laevis. AB - Xlim-1, a LIM class homeobox gene expressed in Xenopus laevis, is one of the earliest known marker genes of pronephros development and is expressed in pronephros rudiment. In this study, we examined the role of Xlim-1 in pronephros development. Temporal expression of Xlim-1 in explants was analyzed in a series of induction assays using RT-PCR analysis. Xlim-1 was expressed 9 to 15 h after activin/retinoic acid treatment, corresponding to pronephros differentiation in explants. We further examined the role of Xlim-1 using a series of microinjection experiments. Presumptive pronephric anlagen of embryos were injected with various Xlim-1 mutants, and effects of these Xlim-1 mutants on pronephrogenesis in embryos and in explants were analyzed by RT-PCR and immunohistochemistry. Dominant-negative Xlim-1 inhibited differentiation of pronephros in activin/retinoic acid-treated animal caps. In embryos injected with a dominant negative form of Xlim-1, development of pronephric tubules was inhibited at the late tail-bud stage. Our results suggest that Xlim-1 may not initiate differentiation of the pronephros, but that it is necessary for growth and elongation in the development of pronephric tubules. PMID- 11112329 TI - Recovery of developmentally defined gene sets from high-density cDNA macroarrays. AB - New technologies for isolating differentially expressed genes from large arrayed cDNA libraries are reported. These methods can be used to identify genes that lie downstream of developmentally important transcription factors and genes that are expressed in specific tissues, processes, or stages of embryonic development. Though developed for the study of gene expression during the early embryogenesis of the sea urchin Strongylocentrotus purpuratus, these technologies can be applied generally. Hybridization parameters were determined for the reaction of complex cDNA probes to cDNA libraries carried on six nylon filters, each containing duplicate spots from 18,432 bacterial clones (macroarrays). These libraries are of sufficient size to include nearly all genes expressed in the embryo. The screening strategy we have devised is designed to overcome inherent sensitivity limitations of macroarray hybridization and thus to isolate differentially expressed genes that are represented only by low-prevalence mRNAs. To this end, we have developed improved methods for the amplification of cDNA from small amounts of tissue (as little as approximately 300 sea urchin embryos, or 2 x 10(5) cells, or about 10 ng of mRNA) and for the differential enhancement of probe sequence concentration by subtractive hybridization. Quantitative analysis of macroarray hybridization shows that these probes now suffice for detection of differentially expressed mRNAs down to a level below five molecules per average embryo cell. PMID- 11112330 TI - Roles for scalloped and vestigial in regulating cell affinity and interactions between the wing blade and the wing hinge. AB - The scalloped and vestigial genes are both required for the formation of the Drosophila wing, and recent studies have indicated that they can function as a heterodimeric complex to regulate the expression of downstream target genes. We have analyzed the consequences of complete loss of scalloped function, ectopic expression of scalloped, and ectopic expression of vestigial on the development of the Drosophila wing imaginal disc. Clones of cells mutant for a strong allele of scalloped fail to proliferate within the wing pouch, but grow normally in the wing hinge and notum. Cells overexpressing scalloped fail to proliferate in both notal and wing-blade regions of the disc, and this overexpression induces apoptotic cell death. Clones of cells overexpressing vestigial grow smaller or larger than control clones, depending upon their distance from the dorsal-ventral compartment boundary. These studies highlight the importance of correct scalloped and vestigial expression levels to normal wing development. Our studies of vestigial-overexpressing clones also reveal two further aspects of wing development. First, in the hinge region vestigial exerts both a local inhibition and a long-range induction of wingless expression. These and other observations imply that vestigial-expressing cells in the wing blade organize the development of surrounding wing-hinge cells. Second, clones of cells overexpressing vestigial exhibit altered cell affinities. Our analysis of these clones, together with studies of scalloped mutant clones, implies that scalloped- and vestigial dependent cell adhesion contributes to separation of the wing blade from the wing hinge and to a gradient of cell affinities along the dorsal-ventral axis of the wing. PMID- 11112331 TI - FGF signaling restricts the primary blood islands to ventral mesoderm. AB - According to the three-signal model of mesoderm patterning in Xenopus, all mesoderm, with the exception of the Spemann organizer, is originally specified as ventral type, such as lateral plate and primary blood islands. It is proposed that the blood islands become restricted to the ventralmost mesoderm because they are not exposed to the BMP-inhibiting activity of the Spemann organizer. We present evidence here that, contrary to predictions of this model, the blood islands remain ventrally restricted even in the absence of Spemann organizer signaling. We further observed that inhibition of FGF signaling with a dominant negative receptor resulted in the expansion of the blood island-forming territory with a concomitant loss of somite. The requirement for FGF signaling in specifying somite versus blood island territories was observed as early as midgastrulation. The nonoverlapping expression domains of Xnr-2 and Xbra in the gastrula marginal zone appear to mark presumptive blood island and somite, respectively. Inhibition of FGF signaling with dominant negative receptor leads to an expansion of Xnr-2 expression and to a corresponding reduction in Xbra expression. On the other hand, we found no evidence that manipulation of BMP signaling, either positively or negatively, altered the expression domains of Xnr 2 and Xbra. These results suggest that FGF signaling, rather than BMP-inhibiting activity, is essential for restriction of the ventral blood islands to ventral mesoderm. PMID- 11112332 TI - Acidic endomembrane organelles are required for mouse postimplantation development. AB - Vacuolar-type H(+)-ATPase (V-ATPase) plays a major role in endomembrane and plasma membrane proton transport in eukaryotes. We found that the acidic compartments generated by V-ATPase are present from the one-cell stage of mouse preimplantation embryos. Upon differentiation of trophoblasts and the inner cell mass at the blastocyst stage, these compartments exhibited a polarized perinuclear distribution. PL16(-/-) embryos, lacking the V-ATPase 16-kDa proteolipid (c subunit), developed to the blastocyst stage and were implanted in the uterine epithelium, but died shortly thereafter. This mutant showed severe defects in development of the embryonic and extraembryonic tissues at a stage that coincided with rapid cell proliferation. When cultured in vitro, PL16(-/-) blastocysts could hatch and become attached to the surface of a culture dish, but the inner cell mass grew significantly slower and most cells failed to survive for more than 4 days. PL16(-/-) cells showed impaired endocytosis as well as organellar acidification. The Golgi complex became swollen and vacuolated, possibly due to the absence of the luminal acidic pH. These results clearly indicate that acidic compartments are essential for development after implantation. PMID- 11112333 TI - Programmed cell death in the developing somites is promoted by nerve growth factor via its p75(NTR) receptor. AB - Neurotrophins control neuron number during development by promoting the generation and survival of neurons and by regulating programmed neuronal death. In the latter case, the cell death induced by nerve growth factor (NGF) in the developing chick retina is mediated by p75(NTR), the common neurotrophin receptor (J. M. Frade, A. Rodriguez-Tebar, and Y.-A. Barde, 1996, Nature 383, 166-168). Here we show that NGF also induces the programmed death of paraxial mesoderm cells in the developing somites. Both NGF and p75(NTR) are expressed in the somites of chick embryos at the time and the place of programmed cell death. Moreover, neutralizing the activity of endogenous NGF with a specific blocking antibody, or antagonizing NGF binding to p75(NTR) by the application of human NT 4/5, reduces the levels of apoptotic cell death in both the sclerotome and the dermamyotome by about 50 and 70%, respectively. Previous data have shown that Sonic hedgehog is necessary for the survival of differentiated somite cells. Consistent with this, Sonic hedgehog induces a decrease of NGF mRNA in somite explant cultures, thus showing the antagonistic effect of NGF and Sonic hedgehog with respect to somite cell survival. The regulation of programmed cell death by NGF/p75(NTR) in a mesoderm-derived tissue demonstrates the capacity of neurotrophins and their receptors to influence critical developmental processes both within and outside of the nervous system. PMID- 11112334 TI - Epithelial-mesenchymal transformation is the mechanism for fusion of the craniofacial primordia involved in morphogenesis of the chicken lip. AB - We have previously demonstrated that epithelial-mesenchymal transformation (EMT) brings about TGF beta 3-induced confluence of craniofacial primordia that derive from the maxillary processes and give rise to the avian palate. The upper lip of the chick embryo forms by confluence of primordia also derived from the maxillary processes, but in this case, they fuse with the intermaxillary segment of the nasofrontal process. Here, we ask whether the bilateral epithelial seams formed when these primordia contact each other in vivo are removed by apoptosis (as formerly was believed to occur in developing palate) or by EMT. We found that, as is the case in the palate, the periderm of the two-layered embryonic epithelium begins to slough shortly before these primordia fuse, bringing the basal epithelial cells into close contact. We show by TUNEL staining and confirm by TEM that apoptosis occurs only in periderm. TEM reveals that basal epithelial cells contacting each other to form the midline seam produce numerous desmosomes with each other. Then, basement membrane begins to disappear, numerous filopodia extend from the basal surfaces of epithelial cells, the space between them enlarges, and the seam breaks apart, leaving mesenchymal cells in its wake. Transformation of the carboxyfluorescein (CCFSE)-labeled epithelial seam is demonstrated in vivo by detection of CCFSE bodies in mesenchymal cells that replace it. This demonstration of EMT in avian lip development lays important groundwork for understanding the causes of human cleft lip and analyzing the mechanism of action of growth factors, such as SHH and BMPs, that have been shown (J. A. Helms et al., 1997, Dev. Biol. 187, 25-35) to be involved in avian lip confluence. PMID- 11112335 TI - gon-4, a cell lineage regulator required for gonadogenesis in Caenorhabditis elegans. AB - The gon-4 gene is required for gonadogenesis in the nematode Caenorhabditis elegans. Normally, two precursor cells, Z1 and Z4, follow a reproducible pattern of cell divisions to generate the mature somatic gonadal structures (e.g., uterus in hermaphrodites, vas deferens in males). In contrast, in gon-4 mutants, the Z1/Z4 cell lineages are variably aborted in both hermaphrodites and males: Z1 and Z4 divide much later than normal and subsequent divisions are either absent or severely delayed. In gon-4 adults, normal somatic gonadal structures are never observed, and germ-line and vulval tissues, which depend on somatic gonadal cues for their development, are also aberrant. In contrast, nongonadal tissues and the timing of other developmental events (e.g., molts) appear to be normal in gon-4 mutants. The gon-4 alleles are predicted to be strong loss-of-function or null alleles by both genetic and molecular criteria. We have cloned gon-4 in an attempt to learn how it regulates gonadogenesis. The gon-4 gene encodes a novel, acidic protein. A GON-4::GFP fusion protein, which rescues a gon-4 mutant to fertility, is expressed in somatic gonadal cells during early gonadal development. Furthermore, this fusion protein is nuclear. We conclude that gon-4 is a regulator of the early lineage of Z1 and Z4 and suggest that it is a part of a genetic program common to the regulation of both hermaphrodite and male gonadogenesis. PMID- 11112336 TI - The mesoderm specification factor twist in the life cycle of jellyfish. AB - The basic helix-loop-helix (bHLH) transcription factor Twist is highly conserved from Drosophila to vertebrates and plays a major role in mesoderm specification of triploblasts. The presence of a Twist homologue in diploblasts such as the cnidarian Podocoryne carnea raises questions on the evolution of mesoderm, the third cell layer characteristic for triploblasts. Podocoryne Twist is expressed in the early embryo until the myoepithelial cells of the larva differentiate and then again during medusa development. There, the gene is detected first when the myoepithelial cells of the polyp dedifferentiate to form the medusa bud and later Twist is found transiently in the entocodon, a mesoderm-like cell layer which differentiates into the smooth muscle and striated muscle of the bell. On the other hand, in later bud stages and the medusa, expression is seen where non muscle tissues differentiate. Experimental analysis of in vitro transdifferentiation and regeneration demonstrates that Twist activity is not needed when isolated striated muscle regenerate medusa organs. Developmental roles of Twist are discussed with respect to early animal evolution from a common ancestor of cnidarians and bilaterians. PMID- 11112337 TI - Regulation of cyclin-dependent kinase 2 activity by ceramide. AB - Cyclin-dependent kinases have been implicated in the inactivation of retinoblastoma (Rb) protein and cell cycle progression. Recent studies have demonstrated that the lipid molecule ceramide is able to induce Rb hypophosphorylation leading to growth arrest and cellular senescence. In this study, we examined the underlying mechanisms of Rb hypophosphorylation and cell cycle progression utilizing the antiproliferative molecule ceramide. C6-Ceramide induced a G0/G1 arrest of the cell cycle in WI38 human diploid fibroblasts. Employing immunoprecipitation kinase assays, we found that ceramide specifically inhibited cyclin-dependent kinase CDK2, with a mild effect on CDC2 and significantly less effect on CDK4. The effect of ceramide was specific such that C6-dihydroceramide was not effective. Ceramide did not directly inhibit CDK2 in vitro but caused activation of p21, a major class of CDK-inhibitory proteins, and led to a greater association of p21 to CDK2. Using purified protein phosphatases, we showed that ceramide activated both protein phosphatase 1 and protein phosphatase 2A activities specific for CDK2 in vitro. Further, calyculin A and okadaic acid, both potent protein phosphatase inhibitors, together almost completely reversed the effects of ceramide on CDK2 inhibition. Taken together, these results demonstrate a dual mechanism by which ceramide inhibits the cell cycle. Ceramide causes an increase in p21 association with CDK2 and through activation of protein phosphatases selectively regulates CDK2. These events may lead to activation of Rb protein and subsequent cell cycle arrest. PMID- 11112338 TI - The cell-surface-expressed nucleolin is associated with the actin cytoskeleton. AB - Nucleolin is a RNA- and protein-binding multifunctional protein. Mainly characterized as a nucleolar protein, nucleolin is continuously expressed on the surface of different types of cells along with its intracellular pool within the nucleus and cytoplasm. By confocal and electron microscopy using specific antibodies against nucleolin, we show that cytoplasmic nucleolin is found in small vesicles that appear to translocate nucleolin to the cell surface. Translocation of nucleolin is markedly reduced at low temperature or in serum free medium, whereas conventional inhibitors of intracellular glycoprotein transport have no effect. Thus, translocation of nucleolin is the consequence of an active transport by a pathway which is independent of the endoplasmic reticulum-Golgi complex. The cell-surface-expressed nucleolin becomes clustered at the external side of the plasma membrane when cross-linked by the nucleolin specific monoclonal antibody mAb D3. This clustering, occurring at 20 degrees C and in a well-organized pattern, is dependent on the existence of an intact actin cytoskeleton. At 37 degrees C, mAb D3 becomes internalized, thus illustrating that surface nucleolin can mediate intracellular import of specific ligands. Our results point out that nucleolin should also be considered a component of the cell surface where it could be functional as a cell surface receptor for various ligands reported before. PMID- 11112339 TI - Overexpression of Cas-interacting zinc finger protein (CIZ) suppresses proliferation and enhances expression of type I collagen gene in osteoblast-like MC3T3E1 cells. AB - Osteoblasts are the cells which form bone under the regulation not only by hormones and cytokines but also by ECM molecules via their attachment. To obtain insights into the role of intracellular signaling molecules operating to mediate the attachment-related regulation of osteoblastic functions, we investigated in osteoblast-like MC3T3E1 cells the effects of the overexpression of CIZ, a novel signaling protein which interacts with p130Cas. In MC3T3E1 cells, CIZ mRNA is expressed constitutively. Endogenous CIZ was localized in the MC3T3E1 cells with relatively high levels of accumulation at the attachment sites when the cells were cultured on fibronectin, collagen, or BSA. CIZ overexpression increased the number of adhesion plaques and reduced proliferation of the cells compared to that of control cells transfected with an empty vector. Furthermore, CIZ overexpression enhanced type I collagen mRNA expression, the most abundant constituent of bone matrix and a major product of osteoblasts. Analysis of the promoter region of type I collagen gene identified the presence of a consensus CIZ-binding sequence, which indeed conferred responsiveness to CIZ overexpression to a heterologous promoter. These data indicate that CIZ acts as a novel regulatory molecule in controlling osteoblastic function. PMID- 11112340 TI - Role of esterase gp70 and its influence on growth and development of Dictyostelium discoideum. AB - Gp70 is an esterase originally called crystal protein because of its presence in crystalline structures in aggregation-competent Dictyostelium discoideum cells. Although postulated to break down spore coats, the function of gp70 in vivo was incompletely investigated. Our immunolocalization and biochemical studies of vegetative D. discoideum amoebae show that gp70 was recruited to phagosomes and found in lysosomes. Purified gp70 was effective at hydrolyzing naphthyl substrates with acyl chains typical of lipids and lipopolysaccharides, indicating that the gp70 was involved in digesting endocytosed molecules. The activity of purified gp70 was inhibited by reductants that retarded its electrophoretic mobility and verified the presence of intramolecular disulfide bonds predicted by its amino acid sequence. Compared to wild-type cells, cells overexpressing gp70 were more phagocytically active, had shorter generation times, and produced more fruiting bodies per unit area, while cells lacking gp70 were phagocytically less active with longer doubling times, developed more slowly, and had significantly fewer fruiting bodies per unit area. Consistent with the phenotype of a disrupted metabolism, one-third of the gp70-minus cells were large and multinucleated. Together, these results indicated that despite its crystalline appearance, gp70 was an active esterase involved in both the growth and the development of D. discoideum. PMID- 11112341 TI - Type IV collagen induces matrix metalloproteinase 2 activation in HT1080 fibrosarcoma cells. AB - Matrix metalloproteinase 2 (MMP-2) activation has been described as a "master switch" which triggers tumor spread and metastatic progression. We show here that type IV collagen, a major component of basement membranes, promotes MMP-2 activation by HT1080 cells. When plated on plastic, HT1080 cells constitutively processed the 66-kDa pro-MMP-2 into a 62-kDa intermediate activated form, most probably through a membrane type (MT) 1 MMP-dependent mechanism. In the presence of type IV collagen, part of this intermediate form was further processed to fully activated 59-kDa MMP-2. This activation was prevented by tissue inhibitor of MMP (TIMP)-2 and a broad-spectrum hydroxamic acid-based synthetic MMP inhibitor (GI129471). Type IV collagen-mediated pro-MMP-2 activation did not involve either a transcriptional modulation of MMP-2, MT1-MMP, or TIMP-2 expression nor any alteration of MT1-MMP protein synthesis or processing. An inverse relationship between MMP-2 activation and the concentration of secreted TIMP-2 was observed. This is consistent with our previous report that TIMP-2 degradation is probably linked to the MT1-MMP-dependent MMP-2 activation mechanism. Because invasive tumor cells must breach basement membranes at different steps of the metastatic dissemination, the ability of HT1080 cells to activate pro-MMP-2 in the presence of type IV collagen might represent a key regulatory mechanism for the acquisition of an invasive potential. PMID- 11112342 TI - Colon cancer cells adhesion and spreading on autocrine laminin-10 is mediated by multiple integrin receptors and modulated by EGF receptor stimulation. AB - Epidermal growth factor (EGF) receptor ligands such as EGF and transforming growth factor-alpha (TGF-alpha) play an important role in controlling the proliferation, survival, morphology, and motility of colonic epithelial cells. There is also increasing evidence that growth factors and extracellular matrix (ECM) proteins cooperate to regulate these cellular processes. We have reported previously that autocrine TGF-alpha and an unidentified ECM protein in the serum free conditioned medium of the human colon carcinoma cell line LIM1215 synergize to induce spreading of these cells in low-density cultures. We have now purified the ECM protein secreted by LIM1215 cells and show that it synergizes with EGF to induce spreading of LIM1215 cells and other human cell lines from the colon and other tissues. The purified ECM migrated as a single protein band with an apparent molecular mass of approximately 800 kDa on SDS-PAGE under nonreducing conditions and, under reducing conditions, as three protein bands of approximately 360, 210, and 200 kDa. Immunoblotting experiments and mass spectrometry analysis of tryptic digests on the purified protein identified the 360-, 210-, and 200-kDa protein bands as laminin alpha5, beta1, and gamma1 chains, respectively, indicating that LIM1215 cells secrete laminin-10 (alpha5 beta1 gamma1). In serum-free medium, LIM1215 cells adhere to laminin-10 primarily via alpha2 beta1 and alpha3 beta1 integrin receptors. EGF-induced spreading of LIM1215 cells on laminin-10 is partially inhibited by pretreatment of the cells with blocking antibodies directed against integrin alpha3 or beta1 but not alpha2, alpha6, or beta4 subunits. Spreading is almost completely inhibited by blocking alpha3 + alpha2, alpha3 + alpha6, or beta1 + beta4 integrin chains and results in cell death. Increased spreading in the presence of EGF correlates with up-regulation of alpha6 beta4 integrins in these cells after exposure to EGF. These results indicate that colon cancer cells attach and spread on laminin-10 via multiple integrin receptors and suggest a critical role for alpha3 beta1 integrins in the spreading response. Together, our results support the concept that the adhesive properties of colon cancer cells are modulated by autocrine production of TGF-alpha and laminin-10 and autocrine induction of appropriate integrins. PMID- 11112343 TI - A first high-density map of 981 biallelic markers on human chromosome 14. AB - As the largest set of sequence variants, single-nucleotide polymorphisms (SNPs) constitute powerful assets for mapping genes and mutations related to common diseases and for pharmacogenetic studies. A major goal in human genetics is to establish a high-density map of the genome containing several hundred thousand SNPs. Here we assayed 3.7 Mb (154,397 bp in 24 alleles) of chromosome 14 expressed sequence tags (ESTs) and sequence-tagged sites, for sequence variation in DNA samples from 12 African individuals. We identified and mapped 480 biallelic markers (459 SNPs and 21 small insertions and deletions), equally distributed between EST and non-EST classes. Extensive research in public databases also yielded 604 chromosome 14 SNPs (dbSNPs), 520 of which could be mapped and 19 of which are common between CNG (i.e., identified at the Centre National de Genotypage) and dbSNP polymorphisms. We present a dense map of SNP variation of human chromosome 14 based on 981 nonredundant biallelic markers present among 1345 radiation hybrid mapped sequence objects. Next, bioinformatic tools allowed 945 significant sequence alignments to chromosome 14 contigs, giving the precise chromosome sequence position for 70% of the mapped sequences and SNPs. In addition, these tools also permitted the identification and mapping of 273 SNPs in 159 known genes. The availability of this SNP map will permit a wide range of genetic studies on a complete chromosome. The recognition of 45 genes with multiple SNPs, by allowing the construction of haplotypes, should facilitate pharmacogenetic studies in the corresponding regions. PMID- 11112344 TI - The assembly of large BACs by in vivo recombination. AB - We have developed a method for recombining bacterial artificial chromosomes (BACs) and P1 artificial chromosomes (PACs) containing large genomic DNA fragments into a single vector using the Cre-lox recombination system from bacteriophage P1 in vivo. This overcomes the limitations of in vitro methods for generating large constructs based on restriction digestion, ligation, and transformation of DNA into Escherichia coli cells. We used the method to construct a human artificial chromosome vector of 404 kb encompassing long tracts of alpha satellite DNA, telomeric sequences, and the human hypoxanthine phosphoribosyltransferase gene. The specificity of Cre recombinase for loxP sites minimizes the possibility of intramolecular rearrangements, unlike previous techniques using general homologous recombination in E. coli, and makes our method compatible with the presence of large arrays of repeated sequences in cloned DNA. This methodology may also be applied to retrofitting PACs or BACs with markers and functional sequences. PMID- 11112345 TI - A major gene affecting age-related hearing loss is common to at least ten inbred strains of mice. AB - Inbred strains of mice offer promising models for understanding the genetic basis of human presbycusis or age-related hearing loss (AHL). We previously mapped a major gene affecting AHL in C57BL/6J mice. Here, we show that the same Chromosome 10 gene (Ahl) is a major contributor to AHL in nine other inbred mouse strains 129P1/ReJ, A/J, BALB/cByJ, BUB/BnJ, C57BR/cdJ, DBA/2J, NOD/LtJ, SKH2/J, and STOCK760. F1 hybrids between each of these inbred strains and the normal-hearing inbred strain CAST/Ei retain good hearing, indicating that inheritance of AHL is recessive. To follow segregation of hearing loss, F1 hybrids were backcrossed to the parental strains with AHL. Auditory-evoked brain-stem response thresholds were used to assess hearing in more than 1500 N2 mice and analyzed as quantitative traits for linkage associations with Chromosome 10 markers. Highly significant linkage was found in all nine strain backcrosses, with the highest probability (LOD > 70) near the marker D10Mit112. This map position for Ahl is near the waltzer mutation (v) and the modifier of deaf waddler locus (mdfw), suggesting the possibility of allelism. Results from an intercross of C57BL/6J and NOD/LtJ mice indicate that the 6- to 10-month difference in AHL onset between these two strains is not due to allelic heterogeneity of the Ahl gene. PMID- 11112346 TI - Genetic mapping of a mouse modifier gene that can prevent ALS onset. AB - Mutations in the cytoplasmic Cu/Zn superoxide dismutase (SOD1) gene on human chromosome 21q22.1 cause 10-20% of familial amyotrophic lateral sclerosis (ALS) cases. The expression of the ALS phenotype in mice carrying the murine G86R SOD1 mutation is highly dependent upon the mouse genetic background. This is similar to the phenotypic variation observed in ALS patients containing identical SOD1 mutations. In the FVB/N background, mice expressing mG86R SOD1 develop an ALS phenotype at approximately 100 days. However, when these mice were bred into a mixed background of C57Bl6/129Sv, the onset of the ALS phenotype was delayed (143 days to >2 years). Using 129 polymorphic autosomal markers in a whole genome scan, we have identified a major genetic modifier locus with a maximum lod score of 5.07 on mouse chromosome 13 between D13mit36 and D13mit76. This 5- to 8-cM interval contains the spinal muscular atrophy (SMA)-associated gene Smn (survival motor neuron) and seven copies of Naip (neuronal apoptosis inhibitory protein), suggesting a potential link between SMA and ALS. PMID- 11112347 TI - Cloning and characterization of a putative human glycerol 3-phosphate permease gene (SLC37A1 or G3PP) on 21q22.3: mutation analysis in two candidate phenotypes, DFNB10 and a glycerol kinase deficiency. AB - Using multiple exons trapped from human chromosome 21 (HC21)-specific cosmids with homology to a putative Arabidopsis thaliana glycerol 3-phosphate permease, we have determined the full-length cDNA sequence of a novel HC21 gene encoding a putative sugar-phosphate transporter (HGMW-approved symbol SLC37A1, aka G3PP). The predicted protein has 12 putative transmembrane domains and is also highly homologous to bacterial glpT proteins. The transcript was precisely mapped to 21q22.3 between D21S49 and D21S113. Comparison of the SLC37A1 cDNA to genomic sequence revealed that the gene encompasses 82 kb, and it is split into 19 coding exons and 7 untranslated exons, which are alternatively spliced in a complex and tissue-specific manner. Glycerol 3-phosphate (G3P) is produced by glycerol kinase (GK) and is found in several biochemical pathways in different cellular compartments, such as the glycerol phosphate shuttle and glycerophospholipid synthesis. Thus SLC37A1 mutations may cause a phenotype similar to GK deficiency. Mutational analyses of SLC37A1 in seven patients with no mutations in the GK gene and low GK activity revealed only nonpathogenetic sequence variants, excluding SLC37A1 as the gene for the phenotype in these patients. SLC37A1 maps in the refined critical region of the autosomal recessive deafness locus, DFNB10, on 21q22.3. Mutation analyses also excluded SLC37A1 as the gene for DFNB10. PMID- 11112348 TI - The murine Pes1 gene encodes a nuclear protein containing a BRCT domain. AB - Pescadillo was originally identified in the zebrafish Danio rerio as a site of a retrovirus-insertion mutation that caused severe defects during embryogenesis. In particular, growth of the fetal zebrafish liver was significantly affected by loss of pescadillo function. To begin to understand the role of pescadillo during mammalian hepatogenesis we identified the murine homologue of pescadillo and named it Pes1. A single gene localized to chromosome 11 on the mouse genome encodes Pes1. Although Pes1 mRNA was detected in all tissues examined it was present at the highest levels in both adult and fetal liver. Analysis of the predicted amino acid sequence of Pes1 found it to contain a BRCT domain, which has previously been found in several proteins involved in cell-cycle checkpoints and DNA repair. Consistent with a putative role in these processes we found that when recombinant Pes1 protein was expressed in HepG2 cells it localized to the nucleus. PMID- 11112349 TI - Genomic organization of human neuropilin-1 and neuropilin-2 genes: identification and distribution of splice variants and soluble isoforms. AB - Neuropilin-1 (NRP1) and neuropilin-2 (NRP2) are both receptors for semaphorins, which regulate neuronal guidance, and vascular endothelial growth factor (VEGF), an angiogenic factor. The two human NRP1 and NRP2 genes were cloned, and the exon intron boundaries were determined. The NRP1 and NRP2 genes span over 120 and 112 kb, respectively, and are composed of 17 exons. Five of the exons are identical in size in the two genes, suggesting that they arose by gene duplication. Both NRP genes are characterized by multiple alternatively spliced variants. Two NRP2 isoforms, NRP2a and NRP2b, were cloned. A striking feature of these two isoforms is that they have identical extracellular domains but have divergent transmembrane and cytoplasmic domains. In these domains, NRP2a is closer in sequence identity to NRP1 than to NRP2b. As determined by Northern blot analysis, both NRP2a and NRP2b are expressed in a variety of tissues, mostly in a nonoverlapping manner. Within NRP2a and NRP2b, there are several alternatively spliced species: NRP2a(17), NRP2a(22), NRP2b(0), and NRP2b(5). In addition to full-length NRPs, there are truncated NRPs as well, which contain only the extracellular a/CUB and b/coagulation factor domains. These genes encode proteins that are soluble (sNRP) and released by cells. In addition to s12NRP1, which was previously cloned, s11NRP1 and s9NRP2 have now been cloned. These sNRP molecules are characterized by having intron-derived sequences at their C-termini. Altogether, eight NRP isoforms are described in this report. It was concluded that there are multiple NRP1 and NRP2 isoforms including intact and soluble forms. Characterization of these isoforms should help to elucidate the function of NRPs in neuronal guidance and angiogenesis. PMID- 11112350 TI - Human uroporphyrinogen-III synthase: genomic organization, alternative promoters, and erythroid-specific expression. AB - Uroporphyrinogen-III (URO) synthase is the heme biosynthetic enzyme defective in congenital erythropoietic porphyria. The approximately 34-kb human URO-synthase gene (UROS) was isolated, and its organization and tissue-specific expression were determined. The gene had two promoters that generated housekeeping and erythroid-specific transcripts with unique 5'-untranslated sequences (exons 1 and 2A) followed by nine common coding exons (2B to 10). Expression arrays revealed that the housekeeping transcript was present in all tissues, while the erythroid transcript was only in erythropoietic tissues. The housekeeping promoter lacked TATA and SP1 sites, consistent with the observed low level expression in most cells, whereas the erythroid promoter contained GATA1 and NF-E2 sites for erythroid specificity. Luciferase reporter assays demonstrated that the housekeeping promoter was active in both erythroid K562 and HeLa cells, while the erythroid promoter was active only in erythroid cells and its activity was increased during hemin-induced erythroid differentiation. Thus, human URO synthase expression is regulated during erythropoiesis by an erythroid-specific alternative promoter. PMID- 11112351 TI - Characterization of the gene encoding the 100-kDa form of human 2',5' oligoadenylate synthetase. AB - The 2'-5' oligoadenylate synthetases (OAS) represent a family of interferon (IFN) induced proteins implicated in the antiviral action of IFN. When activated by double-stranded (ds) RNA, these proteins polymerize ATP into 2'-5' linked oligomers with the general formula pppA(2'p5'A)n, n greater than or = 1. Three forms of human OAS have been described corresponding to proteins of 40/46, 69/71, and 100 kDa. These isoforms are encoded by three distinct genes clustered on chromosome 12 and exhibit differential constitutive and IFN-inducible expression. Here we describe the structural and functional analysis of the gene encoding the large form of human OAS. This gene has 16 exons with exon/intron boundaries that are conserved among the different isoforms of the human OAS family, reflecting the evolutionary link among them. The promoter region of the p100 gene is composed of multiple features conferring direct inducibility not only by IFNs but also by TNF and all-trans retinoic acid. In contrast, the induction of the p100 promoter by dsRNA is indirect and requires IFN type I production. PMID- 11112352 TI - Cloning and characterization of human NTT5 and v7-3: two orphan transporters of the Na+/Cl- -dependent neurotransmitter transporter gene family. AB - Orphan transporters form a growing subfamily of genes related by sequence similarity to the Na+/Cl- -dependent neurotransmitter superfamily. Using a combination of database similarity searching and cloning methods, we have identified and characterized two novel human orphan transporter genes, v7-3 and NTT5. Similar to other known orphan transporters, v7-3 and NTT5 contain 12 predicted transmembrane domains, intracellular N- and C-terminal domains, and large extracellular loops between transmembrane (TM) domains 3 and 4 and between TM domains 7 and 8. Residues within the extracellular loops are also predicted to contain sites for N-linked glycosylation. Human v7-3, the species orthologue of rat v7-3, contains an open reading frame (ORF) of 730 amino acids. Human NTT5 is a new member of the orphan transporter family and has an ORF of 736 amino acids. The amino acid sequences of human v7-3 and NTT5 are greater than 50% similar to other known orphan neurotransmitter transporters and also show sequence similarity to the human serotonin and dopamine transporters. Radiation hybrid mapping located the human v7-3 and NTT5 genes on chromosomes 12q21.3-q21.4 and 19q13.1-q13.4, respectively. Human mRNA distribution analysis by TaqMan reverse transcription-polymerase chain reaction showed that v7-3 mRNA is predominantly expressed in neuronal tissues, particularly amygdala, putamen, and corpus callosum, with low-level expression in peripheral tissues. In contrast, NTT5 mRNA was highly expressed in peripheral tissues, particularly in testis, pancreas, and prostate. Transient transfection with epitope-tagged transporter constructs demonstrated v7-3 to be expressed at the cell surface, whereas NTT5 was predominantly intracellular, suggestive of a vesicular location. Although the substrates transported by these transporters remain unknown, their specific but widespread distribution suggests that they may mediate distinct and important functions within the brain and the periphery. PMID- 11112353 TI - Cloning and characterization of human VPS35 and mouse Vps35 and mapping of VPS35 to human chromosome 16q13-q21. AB - Maintenance of different organelles in eukaryotic cells depends on sorting proteins, which ensure the proper delivery of organelle-specific proteins. The studies on yeast (Saccharomyces cerevisiae) VPS35, a hydrophilic membrane protein having a direct role in the retrieval of cargo proteins, suggest a mechanism underlying a possible lysosomal protein-sorting pathway in mammalian cells. Here, we report the isolation of human and mouse VPS35 cDNAs, which are 3208 and 3186 bp in length, respectively. The deduced proteins of the two cDNAs, which are both composed of 796 amino acids and share 99% identity, show homology to yeast VPS35 and other VPS35 homologues of various sources ranging from Schizosaccharomyces pombe to Drosophila melanogaster (31-56% identity and 49-71% similarity), especially in their amino- and carboxyl-termini. The conservation of VPS35 suggests that the function of this class of protein is important. The results of Northern hybridization of human VPS35 in 16 tissues showed that one transcript of 3.6 kb was highly expressed in brain, heart, testis, ovary, small intestine, spleen, skeletal muscle, and placenta and expressed at moderate or low levels in other tissues. Another transcript of 3.0 kb was also expressed with proportionally lower levels than the 3.6-kb transcript in all the tissues except that the 3.0-kb transcript was not detected in brain. Mouse Vps35 was widely expressed as a 3.4-kb transcript. In addition, human VPS35 was assigned to human chromosome 16q13-q21 by radiation hybrid mapping. PMID- 11112354 TI - Molecular cloning, genomic organization, and mapping of PRKAG2, a heart abundant gamma2 subunit of 5'-AMP-activated protein kinase, to human chromosome 7q36. AB - 5'-AMP-activated protein kinase (AMPK) acts as a major regulator of cellular ATP levels and protects cells against stresses that cause ATP depletion. AMPK is a protein heterotrimer composed of a catalytic alpha subunit and two regulatory subunits, beta and gamma. In the present study, a homologue of the AMPK gamma1 subunit cDNA with an open reading frame encoding 328 amino acids was identified. The putative protein sequence is about 76% identical to the 331-amino-acid gamma1 subunit and also has four consecutive cystathionine-beta-synthase (CBS) domains, a characteristic structure of AMPK gamma subunits from various species. This cDNA (tentatively termed PRKAG2-b) is identical to a recently reported cDNA (tentatively termed PRKAG2-a) of human AMPK gamma subunits except in their 5'-end regions, suggesting that these two cDNAs are two different transcripts of the same gene. To determine the expression pattern of the gene, two probes, one from the 3'-UTR of PRKAG2-b and the other from the 5'- unique region of PRKAG2-a, were used to hybridize MTN membranes. Three transcripts (3.8, 3.0, and 2.4 kb) were observed when the first probe was used, whereas only 3.8- and 3.0-kb transcripts were seen when the second probe was used. Thus, the PRKAG2-b corresponded to the 2.4-kb transcript, which is ubiquitously expressed except in liver and thymus. The highest level was detected in heart, while abundant expression also existed in placenta and testis. The expression pattern of PRKAG2-b is completely different from those of PRKAG2-a and PRKAG1, whose expression patterns were also determined in the current study. The PRKAG2 gene was located to human chromosome 7q36 between markers D7S2439 and D7S2462 by radiation hybrid mapping. The genomic organization of PRKAG2-b was identified by comparing its cDNA sequence with two genomic sequences AC006358 and AC006966, which showed that PRKAG2-b spanned an approximately 80-kb region and was composed of 12 exons. PMID- 11112355 TI - Gene characterization of sciellin (SCEL) and protein localization in vertebrate epithelia displaying barrier properties. AB - Sciellin is a precursor of the cornified envelope of mammalian stratified epithelia characterized by a central core of nonidentical repeats. We characterized the genomic structure of human sciellin and showed that each homologous repeat was encoded by one exon. We also characterized mouse sciellin and showed that mouse sciellin and human sciellin (HGMW-approved symbol SCEL) share a similar gene organization and protein expression pattern. This one exon/one repeat organization is unique among other cornified envelope precursors characterized by homologous repeats. We identified an alternatively spliced isoform of human sciellin, absent in mouse, characterized by an additional repeat at the beginning of the core domain. During embryonic development, the accumulation of sciellin transcript and the accumulation of sciellin protein in the epidermis correlated with the activation of markers of terminal differentiation in epidermis. Mouse sciellin was also identified in simple epithelia with barrier properties, lending further support to its importance in epithelial function. PMID- 11112356 TI - Conserved worldwide linkage disequilibrium in the human factor XI gene. AB - We have identified, in four diverse human populations, five common single nucleotide polymorphisms (SNPs) in the coding region of the gene for the blood coagulation protease factor XI. Each SNP has an allele frequency >5% in at least one population. Three of the SNPs (C472T, A844G, and T1234C), spread out over approximately 10 kb of genomic DNA, are in marked linkage disequilibrium (LD) with one another (P < 10(-4)). Interestingly, haplotypes associated with the linked SNPs are conserved across all populations studied, despite significantly different allele frequencies between populations. The presence of such common, widely dispersed haplotypes could complicate the interpretation of LD studies and emphasizes the need for a better understanding of general patterns of LD to facilitate identification of genes for common disorders. PMID- 11112357 TI - A disease gene for autosomal hyper-IgM syndrome: more genes associated with more immunodeficiencies. PMID- 11112358 TI - Allergen immunotherapy: novel approaches in the management of allergic diseases and asthma. AB - Currently available pharmacotherapies for allergic diseases and asthma, which are serious public health problems, are aimed primarily at neutralizing effector molecules and inflammatory mediators such as histamine and leukotrienes or at inhibiting the function of inflammatory cells such as eosinophils and Th2 lymphocytes. While this approach is effective in controlling symptoms, these therapies have a limited capacity to alter the natural course of allergic diseases and asthma, and discontinuation of medications results in the redevelopment of symptoms on reexposure to the offending allergens. In contrast, immune-based allergen immunotherapies modify and correct the underlying pathological immune responses in allergy and asthma in an antigen-specific manner. These immunotherapies replicate the regulatory processes that occur in nonallergic individuals and allow patients to tolerate exposure to allergens. Current and future methodologies for immunotherapy involve immunization with allergen, modified allergen, peptides of allergen, cDNA of allergen, with adjuvants, including immunostimulatory DNA sequences, cytokines, and bacterial products such as Listeria monocytogenes. This form of therapy can provide a long lasting cure for allergic diseases without the need for continuous therapeutic intervention and without causing generalized immunosuppression or immune augmentation. PMID- 11112359 TI - Mutations in activation-induced cytidine deaminase in patients with hyper IgM syndrome. AB - Recent studies have shown that mutations in a newly described RNA editing enzyme, activation-induced cytidine deaminase (AID), can cause an autosomal recessive form of hyper IgM syndrome. To determine the relative frequency of mutations in AID, we evaluated a group of 27 patients with hyper IgM syndrome who did not have defects in CD40 ligand and 23 patients with common variable immunodeficiency. Three different mutations in AID were identified in 18 patients with hyper IgM syndrome, including 14 French Canadians, 2 Lumbee Indians, and a brother and sister from Okinawa. No mutations were found in the remaining 32 patients. In the group of patients with hyper IgM syndrome, the patients with mutations in AID were older at the age of diagnosis, were more likely to have positive isohemagglutinins, and were less likely to have anemia, neutropenia, or thrombocytopenia. Lymphoid hyperplasia was seen in patients with hyper IgM syndrome and normal AID as well as the patients with hyper IgM syndrome and defects in AID. PMID- 11112360 TI - In vitro immunomodulatory properties of tucaresol in HIV infection. AB - The immunomodulatory properties of tucaresol (compound 589C80) were tested on in vitro antigen- and mitogen-stimulated proliferation and cytokine production by peripheral blood mononuclear cells (PBMC) of HIV-infected individuals and healthy controls (HC). Results showed that tucaresol: (1) increases influenza A virus-, gp 160 peptide-, and HLA alloantigen-stimulated proliferation as well as interleukin (IL)-2 and interferon gamma (IFN gamma) production by PBMC of HIV infected individuals with higher CD4 counts (>500/microl) but had only a marginal immunomodulatory effect on PBMC of patients with lower CD4 counts (<500/microl); (2) did not modify IL-10 production; (3) augmented CD25 expression on mitogen stimulated T cells of HC but not of HIV-infected individuals; and (4) marginally increased CTL activity. The immunomodulatory properties of tucaresol were confirmed by PCR analyses; additional data showed that tucaresol costimulated CD3 dependent triggering of T cells and that this stimulation was independent of CD28 costimulation. The immunomodulatory effects of tucaresol on T cell functions are characterized by a bell-shaped dose response curve; the action of the compound is optimal in the 100 to 300 microM range. Analyses of mitogen-stimulated apoptosis demonstrated that the lack of effect of tucaresol at higher doses is not the result of increased cell death, suggesting a role of functional impairment. These data confirm that tucaresol can stimulate T helper cell function and enhance the production of type 1 cytokines, thus eliciting cell-mediated immunity, and warrant its potential utility in the therapy of HIV infection. PMID- 11112361 TI - A broad-spectrum caspase inhibitor blocks concanavalin A-induced hepatitis in mice. AB - Fulminant hepatic failure (FHF) is a clinical syndrome resulting from massive death of liver cells or sudden and severe impairment of liver function. The causes of FHF are diverse and the overall mortality is very high. Recently, it became clear that apoptosis of hepatocytes is the critical cause of acute hepatic failure in FHF. It is well known that a family of cysteine proteases called caspase is one of the key mediators of the apoptotic pathway. Thus, caspases are attractive potential targets for the treatment of disorders resulting from excessive apoptosis. In this report, we examined the activity of a new caspase inhibitor, Xyz 033 mp. This nonpeptide inhibitor showed broad-spectrum caspase inhibiting activity and protected primary rat hepatocytes from apoptotic death. In a mouse model of FHF induced by concavalin A (Con A), Xyz 033 mp suppressed elevated AST and ALT and specifically reduced IL-1 beta concentration. Also, Xyz 033 mp rescued mice from lethal experimental hepatitis induced by Con A. In addition, histological examinations indicated that Xyz 033 mp protected hepatocytes from the fatal apoptogenic effect of Con A. These results suggest that Xyz 033 mp may be a candidate therapeutic agent for FHF caused by massive apoptotic death of hepatocytes. PMID- 11112362 TI - Humoral response to recombinant hepatitis B virus vaccine at birth: role of HLA and beyond. AB - From 1991 to 1998 we vaccinated 4835 neonates against hepatitis B virus (HBV) and monitored their humoral response to the recombinant vaccine. In a sample of 184 of these babies we studied the association between HLA class I and II genomic polymorphisms and humoral response to the vaccine and the association between the response and immune-mediated diseases. A subgroup of 96 babies also underwent HLA class III (C4A and C4B) typing. Four levels of humoral response were identified, each with a peculiar MHC restriction. Different HLA products seem to act as agonists (C4AQ0 and HLA-DQB1(*)02) or antagonists (C4AQ0, HLA-DQB1(*)02, and HLA DRB1(*)11, DQB1(*)0301) in lowering humoral response to HBV vaccine. The group of responders was characterized more for lacking "nonresponder" alleles than for having specific "responder" ones. Tolerance to HBV peptides may have clinical implications, possibly being a marker for babies with a genetic risk of immunopathologies. In fact, many of the poor responders carried from two to four HLA-DQ alpha beta heterodimers predisposing to insulin-dependent diabetes mellitus and celiac disease. Two true nonresponders suffered from allergies and two slow responders had transient episodes of hyperglycemia. PMID- 11112363 TI - Preferential S phase entry and apoptosis of CD4(+) T lymphocytes of HIV-1 infected patients after in vitro cultivation. AB - We have studied the relationship between spontaneous apoptosis and cell cycle perturbations in circulating peripheral blood lymphocytes of HIV-1-infected patients and healthy controls. PBMC obtained from HIV-1-infected patients and healthy controls were incubated in culture medium for 48 h. Cells were separated into CD4(+) and CD8(+) populations using immunomagnetic beads. Apoptosis and cell cycle phases were measured by propidium iodide staining and bromodeoxyuridine (BrdU) incorporation followed by flow cytometric analyses. In experiments using cells obtained from HIV-1-infected patients, spontaneous apoptosis was more frequent in CD4(+) T lymphocytes than in CD8(+) T lymphocytes (17.6% vs 9.5%, P < 0.005). Among healthy controls, spontaneous apoptosis in CD4(+) and CD8(+) T lymphocytes was comparable (4.5% vs 5.1%). Lymphocytes obtained from patients were more frequently in S phase than healthy controls' cells (2.2 +/- 0.9% vs 0.5 +/- 0.2%, P < 0.002) and patients' CD4(+) cells tended to enter S phase more frequently than controls' CD4(+) cells (4.2% +/- 3.5% vs 1.8% +/- 0.5% P < 0.04), whereas the frequency of S phase CD8(+) T cells was not different among patients (2.8% +/- 2.9%) and controls (1.8% +/- 0.5%) (P > 0.4). Kinetic analyses using BrdU and PI staining revealed that S phase cells were more likely to become apoptotic than resting (G(0)-G(1)) cells (28.4% +/- 10.3% vs 11.3% +/- 9.9% in patients, P < 0.04, and 15.3% +/- 2.8% vs 1.8% +/- 0.5% in controls, P < 0.003). Lymphocytes obtained from HIV-1-infected persons are activated in vivo to enter S phase and to undergo spontaneous apoptosis after brief in vitro cultivation. The present studies indicate that most apoptotic cells in this system are CD4(+) and kinetic analyses reveal that S phase cells are more likely to undergo spontaneous apoptosis than G(0)-G(1) cells. Accelerated cell death in HIV-1 disease may contribute to the failure of lymphocyte responsiveness to appropriate T cell receptor stimulation. PMID- 11112364 TI - Characterization of the T cell receptor repertoire in patients with common variable immunodeficiency: oligoclonal expansion of CD8(+) T cells. AB - Common variable immunodeficiency is characterized by an impairment in antibody production of at least two immunoglobulin isotypes, and abnormalities of both B and T cells have also been described. In this study, a profound alteration of the T cell receptor repertoire was noted, especially in CD8(+) T cells, in CVID patients when compared to a control group. However, the higher number of oligoclonally expanded populations seen in the patient group was not dependent on age, whereas in the control group an age-related accumulation of these populations was noted. Expansion of CD8(+) CD28(-) T cells in these patients is strongly correlated with T cell receptor repertoire restriction. Furthermore, analysis of cytokine production showed a statistically significant increase in IFN-gamma secretion in the patient group. Lastly, CD94 and perforin were expressed at increased frequencies on CD8(+) T cells in the patient group when compared to the control group. We speculate that these findings support the concept that CD8(+) T cells may play an important regulatory role in CVID. PMID- 11112365 TI - Adjuvant-induced improvement of glucose intolerance in type 2 diabetic KK-Ay mice through interleukin-1 and tumor necrosis factor-alpha. AB - We reported that administration of complete Freund's adjuvant (CFA) improved glucose tolerance test (GTT) results in obese diabetic KK-Ay mice. In this study, we investigated its mechanism. An injection with CFA remarkably improved GTT for more than a week in KK-Ay mice, although insulin response was not changed compared with saline controls. The hypoglycemic effect of insulin was significantly, but partially, potentiated in the CFA-treated mice compared with the controls, suggesting that CFA stimulated insulin-mediated and non-insulin mediated glucose disposal. Improvement in the GTT with CFA was partially transferable to nontreated mice by peritoneal exudative cells, but not spleen or lymph node cells. Pretreatment with anti-interleukin (IL)-1 alpha and -1 beta antibodies or anti-tumor necrosis factor (TNF)-alpha antibody significantly abrogated the improvement in the GTT with CFA. The results indicate that CFA induced improvement in glucose intolerance in KK-Ay mice was mediated at least by IL-1 and TNF-alpha. PMID- 11112366 TI - Interleukin 2 leads to dose-dependent expression of the alpha chain of the IL-2 receptor on CD25-negative T lymphocytes in the absence of exogenous antigenic stimulation. AB - Expression of the alpha chain of the interleukin 2 receptor on T lymphocytes is restricted, increasing in the setting of activation, particularly after antigenic stimulation via the TCR. The effects of IL-2 in vitro on the expression of CD25 and proliferation as well as the cytokine induction in CD25-depleted T cells were studied. CD25-depleted and PBMC of healthy donors were cultured for 7 days with 0, 10, or 100 IU/ml of IL-2. Phenotypic analysis and measurement of cytokines in the culture supernatants were performed. IL-2 led to a dose-dependent induction of the IL-2R alpha chain on both CD4 and CD8 T lymphocytes. In the CD25-depleted cultures, IL-2 treatment (100 IU/ml) increased the percentage of CD4 T cells expressing CD25 by 30.6% (P = 0.05) and of CD8 T cells by 48.2% (P = 0.01) on day 7 compared to no treatment. In the PBMC cultures the increase on day 7 was 36.4% for CD4 (P = 0.01) and 50.8% (P = 0.025) for CD8 T lymphocytes. The patterns of cytokine induction in the CD25-depleted and control cultures were similar with increases of IFN-gamma, GM-CSF, IL-16, TNF alpha, and soluble IL-2 receptor in the IL-2-containing cultures. CFSE experiments demonstrated the proliferative capacity of both CD25-positive and -negative T cells. Interleukin 2 alone can lead to a dose-dependent induction of the alpha chain of its receptor on resting CD4 and CD8 T lymphocytes. IL-2 as a sole stimulant is also associated with generation of a cytokine milieu that includes IFN-gamma, GM-CSF, IL-16, and TNF alpha. PMID- 11112367 TI - Apolipophorin-III and the interactions of lipoteichoic acids with the immediate immune responses of Galleria mellonella. AB - We investigated the effects of lipoteichoic acids, surface components of Gram positive bacteria, on the hemocytes and phenoloxidase activity in last instar Galleria mellonella larvae, as well as the binding of apolipophorin-III, an insect lipid-binding protein, to lipoteichoic acids. Binding of apolipophorin-III to lipoteichoic acid was studied using an assay based on 1,9-dimethylmethylene blue. Apolipophorin-III bound the lipoteichoic acids from Bacillus subtilis, Enterococcus hirae, and Streptococcus pyogenes and to intact cells of E. hirae. E. hirae lipoteichoic acid promoted the binding of apolipophorin-III to the cells of this species. All lipoteichoic acids tested caused a dose- and time-dependent drop in the total counts of hemocytes and, depending on the species of lipoteichoic acid, partial or complete depletion of plasmatocytes. Granulocyte counts were not affected. Apolipophorin-III prevented partially the loss of plasmatocytes due to B. subtilis lipoteichoic acid. All three lipoteichoic acids studied activated phenoloxidase in vitro; injections of B. subtilis lipoteichoic acid into the larvae elevated the phenoloxidase activity, whereas injections of E. hirae or S. pyogenes lipoteichoic acid, or apolipophorin-III alone, suppressed it. Apolipophorin-III decreased the activation of phenoloxidase by B. subtilis lipoteichoic acid. PMID- 11112368 TI - Effects of metal-based environmental pollutants on tunicate hemocytes. AB - Tunicates are filter feeding marine invertebrates that are susceptible to environmental contamination by toxic metals and polyaromatic hydrocarbons. Recently, we have shown that tunicate immune reactions are profoundly affected by exposure to tributyltin (TBT) and copper, both of which are components of marine antifouling paints. This study tests the effects of those pollutants on the hemocytes of tunicates. Immunofluorescence labeling with an anti-hemocyte monoclonal antibody demonstrated that the antigenic structure of the circulating hemocyte population was substantially affected by TBT and copper. Antigen positive hemocytes were also found to accumulate in the pharyngeal papillae of TBT-exposed tunicates. Histological analyses indicated that this cellular accumulation in pharyngeal papillae involved refractile vacuolated hemocytes. Refractile vacuolated cells from TBT-exposed tunicates also occurred at greater frequencies in the circulating hemolymph, and had altered morphologies, compared to cells from nontreated controls. These data confirm that exogenous metals can have profound effects on the hemocytes of tunicates. PMID- 11112369 TI - Light and electron microscopic study of a Rickettsiella species from the cockroach Blatta orientalis. AB - An infection with Rickettsiella sp. was responsible for an illness causing heavy body swelling in the Oriental cockroach Blatta orientalis. Reproduction of the colony stagnated. Vacuoles with parasitic bacteria occurred mainly in the fat body, but also in nearly all other organs, such as gut epithelium, Malpighian tubules, blood cells, and ovarioles. The parasites clearly differed from the symbiotic bacteria of the genus Blattabacterium, which regularly occur in the mycetocytes of B. orientalis. The vacuoles contained four stages of Rickettsiella: (1) infectious, electron-dense, rod-like elementary bodies (mean size 300 x 145 nm); (2) an electron-dense, flat intermedium stage, called flat body (mean size 515 x 255 x 125 nm); (3) an electron-light, spherical intermedium stage, called condensing sphere (mean size 340 nm); portions of cytoplasm condensed crescent-like at the border or in the center of the cell; and (4) large, spherical, electron-light initial bodies that multiplied by binary fission (mean size 600 nm). The initial bodies had a three-layered cell boundary, but all other stages had a five-layered cell boundary. Elementary and flat bodies contained an electron-light, oblique lamella and an oval structure with an array of ribosome-like granules, respectively. In contrast to other species of Rickettsiella, crystal formation or multiple division did not occur. The described species of Rickettsiella is different from "R. blattae," which belongs to the R. popilliae group. Instead, it shares more similarities with the R. chironomi group. To avoid confusion, it was provisionally named "R. crassificans." PMID- 11112370 TI - Immune defense mechanisms of Culex quinquefasciatus (Diptera: Culicidae) against Candida albicans infection. AB - Mosquitoes have an efficient defense system against infection. The cellular immune defense mechanism initiated by the mosquito Culex quinquefasciatus infected with the fungus Candida albicans was investigated in this study. Differences in the hemocyte counts in hemolymph perfused from uninoculated, saline-inoculated, and C. albicans-infected mosquitoes were compared using a light microscope. Phagocytosis was also investigated using electron microscopy. Four types of hemocytes were identified in control mosquitoes: prohemocytes (9.8%), plasmatocytes (38.8%), granular cells (44.2%), and oenocytoids (7.3%). Between 3 and 18 h postinoculation the total hemocyte count was significantly higher in infected, compared to uninfected, mosquitoes. Differential hemocyte counts from infected mosquitoes at 3, 6, and 18 h after inoculation showed that the relative proportion of plasmatocytes (48.6, 50.7, 45%) was higher and, concomitantly, the proportion of granular cells was lower (38, 36.8, 35%, respectively). Yeast cells were phagocytosed and limited growth was observed within the plasmatocytes. Melanized nodules were found attached to different insect tissues at 24 to 72 h following infection. These results suggest that phagocytosis, followed by nodule formation, was capable of clearing the hemolymph of yeast cells. PMID- 11112371 TI - Food utilization values of gypsy moth Lymantria dispar (Lepidoptera: Lymantriidae) larvae infected with the microsporidium Vairimorpha sp. (Microsporidia: Burenellidae). AB - Infection of the gypsy moth, Lymantria dispar, with the microsporidium Vairimorpha sp. strongly influences the development of the host in ways typical of many species of terrestrial entomopathogenic Microsporidia; growth is reduced while development time is extended in infected insects. The appearance of the different stages of the parasite in the host relative to the elapsed time after oral infection, as well as the influence of the parasite proliferation on food utilization of the host, were examined. At 3 days postinfection, midgut muscle cells were infected with primary spores, and the fat body tissues contained meronts, sporonts, and primary spores. Many more fat body cells contained vegetative stages and primary spores at 4 and 5 days postinfection, and diplokaryotic spores and immature octospores were also present. Approximate digestibility of infected larvae increased during this time period, whereas the conversion of ingested and digested food to body substance decreased. The relative growth rate of infected and uninfected groups did not differ significantly between 4 and 5 days postinfection, although the relative consumption rate in infected L. dispar larvae was higher. Between 8 and 10 days postinfection, the relative growth rate of uninfected larvae increased. The infected group did not demonstrate this increase at a time period characterized by maturation of diplokaryotic spores and octospores in larval fat body tissues. Total body weight of uninfected larvae remained higher than that of infected larvae after 8 days postinfection. PMID- 11112372 TI - Toxicity of chitinase-producing Bacillus thuringiensis ssp. kurstaki HD-1 (G) toward Plutella xylostella. AB - One-hundred fifty isolates of Bacillus thuringiensis were tested for their ability to produce chitinase using colloidal chitin agar as the primary plating medium. Of 14 strains that produced chitinase, B. thuringiensis ssp. kurstaki HD 1(G) was identified as the highest chitinase producer and selected for further study. This bacterium produced the highest amount of chitinase (19.3 mU/ml) when it was cultivated in nutrient broth supplemented with 0.3% colloidal chitin on a rotary shaker (200 rpm) at 30 degrees C for 2 days. The toxicities of B. thuringiensis ssp. kurstaki HD-1(G) and B. thuringiensis ssp. kurstaki wa-p-2, a chitinase nonproducer, were assayed toward Plutella xylostella (diamondback moth) larvae, resulting in LC(50)'s of 4.93 x 10(4) and 1.32 x 10(5) spores/ml, respectively. If the culture broth from B. thuringiensis ssp. kurstaki HD-1(G) was used as the suspending liquid instead of phosphate buffer, their LC(50)'s were reduced to 6.23 x 10(3) and 7.60 x 10(4) spores/ml, respectively. The histopathological changes of the midgut epithelial cells of diamondback moth larvae were compared after feeding on B. thuringiensis ssp. kurstaki HD-1(G) with and without the presence of supernatant containing chitinase under light microscopy and transmission electron microscopy. The midgut epithelial cells of larvae fed for 30 min in the presence of chitinase, with or without spores and endotoxin crystals, appeared more elongated and swollen than those of the control larvae. A number of different cellular changes such as extensive cellular disintegration and appearance of numerous vacuoles were observed from the larvae fed on B. thuringiensis ssp. kurstaki HD-1(G) supplemented with supernatant containing chitinase. Thus increased toxicity and changes in epithelial cells were correlated with the presence of chitinase but this was not distinguished from the possible presence of vegetative-stage insecticidal proteins. PMID- 11112373 TI - Responses of giant freshwater prawn (Macrobrachium rosenbergii) to challenge by two strains of Aeromonas spp. AB - The virulence of two Aeromonas strains (A. veronii and A. caviae) isolated from the hepatopancreas of apparently healthy giant freshwater prawns (Macrobrachium rosenbergii) was compared using a challenge by injections. For the A. veronii strain, challenge with 3.7 x 10(5) cells/g of body weight led to 100% mortality; for the A. caviae strain, 3.8 x 10(6) cells/g produced 100% mortality. The 50% lethal doses (LD50) were 2.0 x 10(3) cells/g for A. veronii and 51.2 x 10(3) cells/g for A. caviae. Use of different culture media (trypticase soy broth vs prawn muscle extract) did not significantly affect the virulence of A. veronii. Injection of a sublethal dose (1 x 10(3) cells/g) of A. veronii led to a significant decrease in the total hemocyte count (THC) between 4 and 24 h after injection. Saline injections also caused a similar though less decrease in THC. In the first 24 h after injection of A. veronii (1 x 10(3) cells/g), the change in the percentages of granulocytes (both granular cells and semigranular cells) in the hemolymph was significantly different. After a significant initial increase, the percentage of hyaline cells fell by a factor of 4, from 9 to 2%. Phenoloxidase activity increased fourfold immediately after injection and returned to preinjection levels at 24 h. PMID- 11112375 TI - Development and pathology of two undescribed species of microsporidia infecting the predatory mite, Phytoseiulus persimilis Athias-Henriot. AB - Two undescribed species of microsporidia were found in mass-reared Phytoseiulus persimilis Athias-Henriot from two commercial sources during a routine examination of these predators for pathogens. Both microsporidian species were described from specimens that had been prepared for transmission electron microscopy; live specimens were unavailable for examination. One microsporidium, identified as Species A, was described from two specimens obtained from a commercial insectary in North America. All observed stages of this microsporidium were uninucleate. Rounded-to-ovoid schizonts appeared to develop in direct contact with the cytoplasm of lyrate organ cells (ovarian tissue). Mature spores of Species A were elongate-ovoid and measured 2.88 x 1.21 microm. A polar filament coiled 7 to 10 times in the posterior half of the spore. Sporoblasts and spores were observed in the cytoplasm of cells of numerous tissues and in developing eggs within gravid females. A second species, identified as Species B, was described from five specimens obtained from a commercial source in Israel. All observed stages of this microsporidium were uninucleate. Schizonts of Species B were observed within the cytoplasm of cecal wall cells and within the nuclei of lyrate organ cells. Mature spores were ovoid and measured 2.65 x 1.21 microm. A polar filament coiled 3 to 4 times in the posterior half of the spore. Densely packed ribosomes often concealed the polar filament and other internal spore characteristics. Spores were observed in the cytoplasm of cells of numerous tissues and occasionally within the nuclei of lyrate organ cells. Numerous spores and presporal stages were observed within the ovary and developing eggs. The development and pathology of Species A and B were compared to those of Microsporidium phytoseiuli Bjoornson, Steiner and Keddie, a microsporidium previously described from P. persimilis obtained from a commercial source in Europe. The occurrence of three species of microsporidia within P. persimilis from three sources raises questions regarding the origin of these pathogens. Because microsporidia may have profound impact on the performance of P. persimilis, consideration must be given to the identification and exclusion of microsporidia from field-collected specimens or from predators that may be shared among commercial sources. PMID- 11112374 TI - Interaction of Xenorhabdus nematophilus (Enterobacteriaceae) with the antimicrobial defenses of the house cricket, Acheta domesticus. AB - Fifth instar Acheta domesticus nymphs exhibited a decline in total hemocyte counts during the first hour of exposure to dead Xenorhabdus nematophilus; the bacterial level in the hemolymph also declined during this time. Thereafter bacterial numbers in the hemolymph increased as the level of damaged hemocytes increased. The bacteria lowered phenoloxidase activity in vivo by initially reducing the number of hemocytes containing prophenoloxidase and later by inhibiting enzyme activation. Preincubating X. nematophilus in hemolymph with active phenoloxidase in vitro accelerated the removal of the bacteria from the hemolymph in vivo which may be due to modification of the bacterial surface by serine proteases. Lysozyme activity increased in bacteria-injected insects in parallel with an increase in counts of damaged hemocytes; most of the enzyme was located in hemocytes. Lipopolysaccharides of X. nematophilus caused changes in hemocyte counts and phenoloxidase and lysozyme levels comparable to whole bacteria. Lipopolysaccharides also slowed the removal rate of the bacteria from, and accelerated bacterial emergence into, the hemolymph. PMID- 11112376 TI - Francisella-like endosymbionts of ticks. PMID- 11112377 TI - A new gene-pseudogene fusion allele due to a recombination in intron 2 of the glucocerebrosidase gene causes Gaucher disease. AB - Gaucher disease is the most prevalent sphingolipid storage disorder in humans caused by a recessively inherited deficiency of the enzyme glucocerebrosidase. More than 100 mutations have been described in the glucocerebrosidase gene causing Gaucher disease. Some of them are complex alleles with several mutations due to recombination events between the gene and its highly homologous pseudogene. The generation of these recombinant alleles involves, in most cases, a crossover in the 3' end of the gene, beyond exon 8. However, in a few cases recombination took place in a more upstream location. Here we describe the analysis of a patient with type I Gaucher disease who bears a new complex allele. This allele was originated by a crossover between the gene and the pseudogene at intron 2, the most upstream recombination site described so far, which gave rise to a fusion gene. The patient was first diagnosed as homozygous for the c.1226 A -> G (N370S) mutation but the early onset of the disease prompted us to perform parental DNA analysis which showed that the mother was not a N370S carrier, suggesting deletion of at least part of the gene. Molecular analysis of the complex allele was carried out by Southern blot, PCR, and sequencing. We were able to close down the region of the recombination event to an interval of 18 nucleotides, corresponding to the last 15 nucleotides of intron 2 and the first 3 nucleotides of exon 3 of the gene. These 18 nucleotides are identical between the gene and pseudogene making any further refinement impossible. An exhaustive list of published glucocerebrosidase complex alleles, describing their recombination points, is included for comparison. PMID- 11112379 TI - Phenotype determination of a common Pro-Leu polymorphism in human glutathione peroxidase 1. AB - Oxidative stress has been implicated in human illness such as cardiovascular and neurodegenerative disease. The genetic mechanisms involved are only poorly understood. Here we describe the determination of the allelic frequency and phenotype of a common polymorphism in Se-dependent glutathione peroxidase 1 (GPX1) in Finnish/Swedish populations. A proline/leucine variant occurs at position 197 close to the C-terminus of the protein. The more common allele encoding the Pro variant is present at 59% in a Finnish/Swedish population (n = 66) and at 73% in a Swedish population (n = 315). The genotypes encoding Pro/Pro, Pro/Leu, and Leu/Leu are distributed according to the Hardy-Weinberg relationship. The Swedish population consisted of 101 stroke cases and 214 controls. No significant association between allele frequency and risk to suffer from stroke was evident. Erythrocyte GPX activity was determined in the Finnish/Swedish population and no significant differences were obtained between the genotypes. It can be concluded that the Pro/Leu genetic variation does not appear to compromise the defense against oxidative stress in red blood cells nor to be associated with stroke. PMID- 11112378 TI - Diamond-Blackfan anemia: report of seven further mutations in the RPS19 gene and evidence of mutation heterogeneity in the Italian population. AB - Diamond-Blackfan anemia (DBA) is a congenital disease characterized by defective erythroid progenitor maturation and physical malformations. Most cases are sporadic, but dominant or, more rarely, recessive inheritance is observed in 10% of patients. Mutations in the gene encoding ribosomal protein (RP) S19 have recently been found in 25% of patients with either the dominant or the sporadic form. DBA is the first human disease due to mutations in a ribosomal structural protein. Families unlinked to this locus have also been reported. In an investigation of 23 individuals, we identified eight different mutations in 9 patients. These include five missense, one frameshift, one splice site defect, and one 4-bp insertion in the regulatory sequence. Seven mutations are new; one has so far been found in 8 patients and is a relatively common de novo event. Two mutations are predicted to generate a truncated protein. We also report the prevalence of RPS 19 mutations in the Italian DBA population, as shown by an analysis of 56 patients. No genotype-phenotype correlation was found between patients with the same mutation. The main clinical applications for molecular analysis are clinical diagnosis of patients with an incomplete form of DBA and testing of siblings of a patient with a severe form so as to avoid using those who carry a mutation and a silent phenotype as allogeneic stem cell donors. PMID- 11112380 TI - Effects of lipoxygenase metabolites of arachidonic acid on the growth of human blood CD34(+) progenitors. AB - The influence of lipoxygenase metabolites of arachidonic acid on proliferation and differentiation of CD34(+) cells was studied. Their effects on the CFU-GM and BFU-E progenitors were investigated by culture of CD34(+) cells in liquid or semisolid medium. Only 12-HETE (1 microM) stimulated the [(3)H]thymidine as well as BrdU incorporation and increased the number of cell divisions (PKH2 tracking). Addition of 12-HETE and 15-HETE but not of LXA(4), LXB(4), LTB(4), and LTC(4) to liquid cultures of CD34(+) cells for 3 and 8 days reduced in a time-dependent manner the number of CFU-GM and BFU-E. Both HETEs also increased the percentage of glycophorin A(+) cells while they reduced the percentage of CD34(-)/CD33(+) cells after 3 and 5 days of liquid cultures. These results show that HETE treatment stimulates proliferation and accelerates the differentiation of CD34(+) cells, mostly toward the erythroid lineage. PMID- 11112381 TI - Primary structure of Noetia ponderosa hemoglobins: functional correlates. AB - Homo- and heterodimeric hemoglobins have been isolated from the red cells of the arcid clam Noetia ponderosa (Np). These hemoglobins bind oxygen cooperatively. An extensively studied dimeric hemoglobin from another arcid clam, Scapaharca inaequivalvis, exhibits a molecular mechanism for cooperative ligand binding that is radically different from tetrameric vertebrate hemoglobins. In this study, the two chains found in both Noetia hemoglobins are sequenced and compared to the hemoglobins of the related clam S. inaequivalvis to determine whether Noetia hemoglobins have the structural basis for the same unusual mechanism for cooperative ligand binding and to inquire about the structural basis of absence of tetramers. Although the Noetia sequences are homologous to the Scapharca sequences, critical differences exist. The lack of tetramerization of Np subunits is most likely related to the absence of critical residues in the A and G helices that stabilize the interdimer contact seen in the Scapharca Hb tetramer. The lower affinity of the homodimer (Np-I), but particularly the heterodimer (Np-II) with respect to the homodimer and heterotetramer of Scapharca, can be due to (i) changes in the proximal heme environment and (ii) changes in the dimer interface. Interactions between Asn 100 and the heme of the other subunit are altered in Np II due to the substitution of this residue by methionine, possibly causing the reduced O(2) affinity of the heterodimer of Noetia. (iii) Sequence changes in the E and F helices present in Np-I and Np-II could also contribute to the effect through interfacial changes. In particular, the substitution of Val for Thr in position 72 is expected to have a substantial influence on the interface. We conclude that Np dimers have the structural basis for a direct heme-heme interaction mechanism for cooperativity, as in Scapharca, but there are enough sequence changes to suggest that the pathway of interaction might be somewhat different. PMID- 11112382 TI - Effects of infectious hematopoietic necrosis virus and infectious pancreatic necrosis virus infection on hematopoietic precursors of the zebrafish. AB - The zebrafish Danio rerio is a new model system for studying the genetics of hematopoiesis. To define naturally occurring viruses which could infect and replicate within hematopoietic precursors of the zebrafish, infectious hematopoietic necrosis virus (IHNV) and infectious pancreatic necrosis virus (IPNV) were studied. Infection of whole fish with viral supernatants demonstrated infectious replicants for both viruses, indicating that the virus host range includes the zebrafish. In other species, infection with these viruses leads to prominent hematopoietic necrosis of the head kidney, the major site of adult hematopoiesis. We detected a transient toxicity of the virus to hematopoietic precursors and terminally differentiated red cells after viral infections. The kinetics of hematopoietic defects between IHNV and IPNV infection differed; fish infected with either virus, however, recovered by 6 days postinfection. In contrast to other fish infected with the virus, hematocrit did not change appreciably during this time. These studies are the first to demonstrate IHNV and IPNV infection of the zebrafish and reveal the potential for use of such viruses for gene transfer experiments to infect zebrafish hematopoietic cells. PMID- 11112383 TI - Reduction of the clinical severity of sickle cell/beta-thalassemia with hydroxyurea: the experience of a single center in Greece. AB - The use of hydroxyurea for the prevention of sickle cell crises in patients with homozygous HbS disease is now well established. The beneficial effects of this compound stem from (a) selective enrichment of red cells containing an increased amount of fetal hemoglobin, which inhibits HbS polymerization, and (b) a decrease of leukocytes, platelets, and reticulocytes, which significantly limits their adherence to the vascular wall. We report the results of a clinical trial of hydroxyurea on 55 Greek-origin patients with sickle cell/beta-thalassemia and 14 patients with homozygous HbS disease who have been treated with hydroxyurea for several years. Such patients have a higher probability to benefit from hydroxyurea therapy, since in addition to its antisickling effect, the increase of gamma-chain synthesis is expected to diminish the deleterious effects of the unbound alpha-globin chains. Selection of patients and monitoring throughout the whole trial were done by the same clinicians. Quantitative expression of the clinical condition was done using a system scoring several outcome parameters. For a period of 52 months prior to starting treatment, the total score of severity for 59 evaluable patients was 1182 points (3068 patient-weeks), while for the 12,018 patient-weeks of the trial this parameter fell to only 82 points. Other observations of interest include the significant improvement of a group of patients with hepatic cholestasis, the development of leg ulcers possibly related to the treatment, and the dramatic increase of hemoglobin F, often in association with an increase of the total hemoglobin levels as a result of decreased hemolysis. PMID- 11112384 TI - Erythropoietin-dependent suppression of the expression of the beta subunits of the interleukin-3 receptor during erythroid differentiation. AB - To clarify how erythroid cells lose their response to interleukin-3 (IL-3), we analyzed the expression of the alpha (alpha(IL-3)) and beta (beta(IL 3)/beta(com)) subunits of its receptor in a panel of murine cell lines immortalized at different stages of hemopoietic differentiation. The panel was composed by the mast cell line 32D and by its granulo-monocytic (32D GM), granulocytic (32D G), and erythroid (32D Epo1.1 and Epo) subclones. The 32D Epo cells grow only in erythropoietin (EPO) while the Epo1.1 subclone grows in either EPO or IL-3. The phenotype of these cells is that of early (expression of globins and erythroid-specific carbonic anhydrase II) and late (also expression of the erythroid-specific band 4.1 mRNA) erythroblasts when they grow in IL-3 or EPO, respectively. All the cell lines expressed comparable levels of alpha(IL-3). In contrast, the expression of beta(IL-3)/beta(com) was restricted to cells growing in IL-3 and was barely detectable in 32D Epo and 32D Epo1.1 cells growing in EPO. When switched from EPO to IL-3, 32D Epo1.1 cells expressed 10 times more beta(IL 3)/beta(com) by rapidly activating (within 1 h) their transcription rate. When reexposed to EPO, 32D Epo1.1 cells first expressed (1-6 h) more beta(IL 3)/beta(com) (2 times) but suppressed such an expression at later time points (by 48 h). The beta(IL-3)/beta(com) mRNA half-life was also different when 32D Epo1.1 cells grew in EPO or IL-3 (2-3 h vs >5 h, respectively). These results indicate that EPO specifically induces transcriptional and posttranscriptional downmodulation of beta(IL-3)/beta(com) expression in late erythroid cells. PMID- 11112385 TI - Characterization of zebrafish full-length prothrombin cDNA and linkage group mapping. AB - In this paper, we report the complete cDNA sequence of zebrafish prothrombin. The cDNA sequence predicts that zebrafish prothrombin is synthesized as a pre proprotein consisting of a Gla domain, two kringle domains, and a two-chain protease domain. Zebrafish prothrombin is structurally very similar to human and other vertebrate prothrombins. Zebrafish and human prothrombin share 53% amino acid identity whereas zebrafish and hagfish prothrombin share 51% identity. Amino acid alignments of various prothrombins identified conservation of many of the functional/structural motifs suggesting that the vertebrate prothrombins may have similar functions. The three-dimensional structure of prothrombin predicted by homology modeling also revealed that the prothrombin fragment 1 and the catalytic domain structures are well conserved except for the insertion of an extra 7-amino acid loop in the connecting region (CR) between the Gla and kringle I domain of fragment 1. Linkage analysis revealed that the prothrombin gene locus on linkage group 7 in zebrafish is syntenic to the human chromosome 11-prothrombin region suggesting its preservation through evolution. The availability of this cDNA sequence in zebrafish adds to our knowledge of the zebrafish hemostatic system and provides support for the view that similarities between zebrafish and mammalian coagulation exist, thus underscoring the relevance of the zebrafish model for studying human hemostasis. PMID- 11112386 TI - Telomerase activity and expression and telomere analysis in situ in the course of treatment of childhood leukemias. AB - Samples of blood and marrow from children with leukemia were assayed for telomerase activity and expression on the day of diagnosis and during the course of chemotherapy. A strong correlation between either variables and clinical response was observed in most patients. A unique case was observed in which telomerase activity was only moderately increased on diagnosis; it gradually increased in the course of therapy, and a subsequent decrease occurred only after application of intensified therapy. This patient did not respond to therapy, his disease progressed, and he finally died during intensified therapy. In another patient, analysis of telomere lengths using dideoxy-PRINS revealed a single telomere expansion on a long arm of chromosome 4, suggesting involvement of a telomerase-independent mechanism of telomere elongation. PMID- 11112387 TI - Anticoagulant proteins in childhood venous and arterial thrombosis: a review. AB - Thrombotic disease is less frequent in children than in adults, but may result in severe morbidity and mortality. The coagulation system is balanced to provide rapid activation to stop bleeding and appropriate inhibition to prevent unwanted clot extension. It is regulated by fibrinolysis and by three major anticoagulant pathways: the protein C, antithrombin, and tissue factor pathway inhibitor systems. Acquired or inherited abnormalities of coagulation proteins or hemostatic regulatory mechanisms, particularly when combined with dehydration or the presence of indwelling catheters, may pose a high risk for thrombosis. Thrombosis in a child warrants investigation of potential underlying prothrombotic conditions. These include acquired antiphospholipid antibodies or the lupus anticoagulant as well as abnormalities of the inherited anticoagulant factors including protein C, protein S, antithrombin, and Factor V Leiden. Other abnormalities may result in heightened levels of otherwise normal coagulation proteins such as hyperprothrombinemia due to the prothrombin 20210 mutation. A large survey of children with thrombosis indicated that Factor V Leiden, protein C deficiency, and increased lipoprotein(a) were found most commonly. The most severe predisposition occurs with homozygous protein S or protein C deficiency with resultant purpura fulminans in the newborn. The risk of thrombosis in children with heterozygous deficiencies of anticoagulant proteins is not well defined, although it is clear that combined heterozygotes or a combination of an inherited and an acquired defect heightens the risk for thrombosis. Treatment of thrombosis primarily involves a rapidly acting anticoagulant such as heparin or low-molecular-weight heparin to prevent extension, and long-term anticoagulation with warfarin may be instituted to prevent recurrence. Fibrinolytic therapy is infrequently used because of the risk of serious bleeding complications and is reserved for selected cases of arterial thrombosis to initiate rapid reperfusion of ischemic tissue or used in those patients with a large venous thrombosis and pulmonary emboli causing hemodynamic compromise. PMID- 11112388 TI - Hematologically important mutations: the autosomal recessive forms of chronic granulomatous disease (first update). PMID- 11112389 TI - A new exon 9 glucose-6-phosphate dehydrogenase mutation (G6PD "Rehovot") in a Jewish Ethiopian family with variable phenotypes. AB - Hereditary nonspherocytic hemolytic anemia (HNSHA) is a rare manifestation of glucose-6-phosphate dehydrogenase (G6PD) gene mutations, caused mainly by mutations located in exon 10 of the G6PD gene and less commonly by mutations in other parts of the gene. A new, exon 9, single-base mutation representing a T --> C transition at cDNA nucleotide 964 was found in three brothers and their carrier mother of Jewish Ethiopian descent. Biochemical characterization of the resultant protein was not performed. Though clinical manifestations included HNSHA in all cases, the severity of hemolysis and the transfusion requirement differed markedly. Severe congenital neutropenia (Kostmann's syndrome)--a disorder never reported before in conjunction with G6PD deficiency--was observed in one case. Levels of white blood cell G6PD activity of the three patients were 0-5% of normal controls. Neutrophil oxidative and bactericidal activities were inherently impaired in the patient with Kostmann's syndrome, but were well preserved in his two siblings. PMID- 11112390 TI - Myelodysplasia or myeloneoplasia: thoughts on the nosology of clonal myeloid diseases. PMID- 11112391 TI - Pulsed intravenous high-dose dexamethasone in adults with chronic idiopathic thrombocytopenic purpura. AB - The role of pulsed high-dose dexamethasone (DXM) in the treatment of patients with chronic idiopathic thrombocytopenic purpura (ITP) is still uncertain. Following an early report in which it was described as an effective and well tolerated treatment with a sustained platelet response in 100% of cases, a number of subsequent studies have failed to confirm such favorable results. As all these studies were conducted on small numbers of patients, we investigated further the effectiveness and side effects of this therapeutic modality in a larger cohort. Thirty-two patients with chronic ITP were scheduled to receive six monthly courses of intravenous DXM at the dose of 40 mg/day for 4 consecutive days. All patients had ITP that had been resistant to between two and five different therapeutic regimens, including 9 patients who had already failed splenectomy. All patients had to be seen 2 weeks after each cycle to asses their response as well as secondary effects. Three patients failed to respond and clinically required other therapy. Thirteen patients (41%) had a partial (platelet count between 50 and 100 x 10(9)/liter) or complete (platelet count >100 x 10(9)/liter) response to treatment, responses being mostly transient. Responses were observed early during the course of treatment, usually right after the first cycle of DXM. There were no late responses. Side effects were mild and did not require discontinuation of treatment. No clinical or laboratory parameter was found to predict treatment outcome. We conclude that high-dose DXM has a limited effect in patients with chronic ITP. Novel approaches and controlled multicenter trials may help identify new therapeutic strategies for this disease. PMID- 11112393 TI - The Morel stereotactic atlas of the human thalamus: atlas-to-MR registration of internally consistent canonical model. AB - In 1997, Morel, Magnin, and Jeanmonod presented a microscopic stereotactic atlas of the human thalamus. Parcellations of thalamic nuclei did not only use cyto- and myeloarchitectonic criteria, but were additionally corroborated by staining for calcium-binding proteins, which bears functional significance. The atlas complies with the Anglosaxon nomenclature elaborated by Jones and the data were sampled in three orthogonal planes in the AC-PC reference space. We report on the generation of three-dimensional digital models of the thalamus based on the three sets of sections (sagittal, horizontal, and frontal). Spatial differences between the three anatomical specimens were evaluated using the centers of gravity of 13 selected nuclei as landmarks. Subsequent linear regression analysis yielded equations, which were used to normalize the frontal and horizontal digital models to the sagittal one. The outcome is an internally consistent Canonical Model of Morel's atlas, which minimizes the linear component of the variability between the three sectioned anatomical specimens. In addition, we demonstrate the feasibility of the atlas-to-MRI registration in conjunction with on-line visualization of the trajectory in the digital models. PMID- 11112392 TI - GATA and NF-Y participate in transcriptional regulation of FcgammaRIIA in megakaryocytic cells. AB - Human FcgammaRIIA, expressed on platelets, neutrophils, and macrophages, plays a major role in platelet activation and immune clearance. Clinical observations indicate that regulation of expression of this receptor is an important factor influencing the course of immune thrombocytopenia. We used both transient transfection with FcgammaRIIA promoter constructs and electrophoretic mobility shift assays (EMSA) to study the regulation of FcgammaRIIA transcription. In HEL (erythromegakaryocytic) cells, the 200 bp immediately 5' of the ATG start codon accounted for the majority of the activity of a 3.6-kb promoter fragment. Putative GATA (-161) and NF-Y (-119) sites are present. EMSA analyses demonstrate specific binding of both GATA-1 and GATA-2 to labeled oligonucleotides containing the putative GATA site with HEL but not U937 (myelomonocytic) nuclear extracts. Antibodies to NF-Y supershift the specific -119 NF-Y complex with HEL, U937, Jurkat (T-lymphocytic), and HeLa (nonhematopoietic) nuclear extracts. Comparison of the activity of GATA and NF-Y mutant constructs in HEL and U937 demonstrates that while either GATA or NF-Y mutation results in a large decrease in the promoter activity (2.2- and 2.3-fold, respectively) in HEL cells, neither mutation is effective in reducing activity in U937 cells. This is the first example of a promoter active in the megakaryocyte lineage in which NF-Y cooperates additively with GATA factors to regulate transcription. Identification of other factors that must be operational for FcgammaRIIA transcription in myelomonocytic cells which lack GATA factors will bolster our ongoing efforts to dissect the function of these Fc receptors in megakaryocytic and myelomonocytic cells in vivo. PMID- 11112394 TI - SATSCOM--Selective amobarbital test intraarterial SPECT coregistered to MRI: description of a method assessing selective perfusion. AB - For evaluation of potential functional deficits, an intraarterial amobarbital test is performed prior to neurosurgical or neuroradiological interventions. To visualize individual amobarbital perfusion patterns, simultaneous injection of (99m)Tc-HMPAO was performed previously. The present study describes for the first time a method of coregistration of intraarterial SPECT during selective amobarbital test to MRI. Three patients undergoing selective amobarbital test of the posterior cerebral artery were included. SATSCOM (Selective amobarbital test intraarterial SPECT coregistered to MRI) was performed by skull extraction in SPECT and MRI followed by surface matching. In all three patients, SATSCOM revealed accurate matching results. With this functional-anatomical mapping, suppression of higher cortical functions can be correlated to anatomical regions. Furthermore, a more precise mapping of amobarbital effect improves planning invasive interventions, particularly those close to eloquent areas. PMID- 11112395 TI - Spontaneous low frequency oscillations of cerebral hemodynamics and metabolism in human adults. AB - We investigated slow spontaneous oscillations in cerebral oxygenation in the human adult's visual cortex. The rationale was (1) to demonstrate their detectability by near infrared spectroscopy (NIRS); (2) to analyze the spectral power of as well as the phase relationship between the different NIRS parameters (oxygenated and deoxygenated hemoglobin and cytochrome-oxidase; oxy-Hb/deoxy Hb/Cyt-ox). Also (3) influences of functional stimulation and hypercapnia on power and phase shifts were investigated. The results show that-in line with the literature-low frequency oscillations (LFO) centred around 0.1 s(-1) and even slower oscillations at about 0.04 s(-1) (very low frequency, VLFO) can be distinguished. Their respective power differs between oxy-Hb, deoxy-Hb, and Cyt ox. Either frequency (LFO and VLFO) is altered in magnitude by functional stimulation of the cortical area examined. Also we find a change of the phase shift between the vascular parameters (oxy-Hb, tot-Hb) and the metabolic parameter (Cyt-ox) evoked by the stimulation. It is shown that hypercapnia attenuates the LFO in oxy-Hb and deoxy-Hb. CONCLUSIONS: (1) spontaneous vascular and metabolic LFO and VLFO can be reproducibly detected by NIRS in the human adult. (2) Their spectral characteristics and their response to hypercapnia are in line with those described in exposed cortex (for review see (Hudetz et al., 1998)) and correspond to findings with transcranial doppler sonography (TCD) (Diehl et al., 1995) and fMRI (Biswal et al., 1997). (3) The magnitude of and phase relation between NIRS-parameters at the LFO may allow for a local noninvasive assessment of autoregulatory mechanisms in the adult brain. PMID- 11112396 TI - Validation of partial tissue segmentation of single-channel magnetic resonance images of the brain. AB - We describe and evaluate a practical, automated algorithm based on local statistical mixture modeling for segmenting single-channel, T1-weighted volumetric magnetic resonance images of the brain into gray matter, white matter, and cerebrospinal fluid. We employed a stereological sampling method to assess, prospectively, the performance of the method with respect to human experts on 10 normal T1-weighted brain scans acquired with a three-dimensional gradient echo pulse sequence. The overall kappa statistic for the concordance of the algorithm with the human experts was 0.806, while that among raters, excluding the algorithm, was 0.802. The algorithm had better agreement with the modal expert decision (kappa = 0.878). The algorithm could not be distinguished from the experts by this measure. We also validated the algorithm on a simulated MR scan of a digital brain phantom with known tissue composition. Global gray matter and white matter errors were 1% and <1%, respectively, and correlation coefficients with the underlying tissue model were 0.95 for gray matter, 0.98 for white matter, and 0.95 for cerebrospinal fluid. In both approaches to validation, we evaluated both local and global performance of the algorithm. Human experts generated slightly higher global gray matter proportion estimates on the test brain scans relative to the algorithm (3.7%) and on the simulated MR scan relative to the true tissue model (4.4%). The algorithm underestimated gray in some subcortical nuclei which contain admixed gray and white matter. We demonstrate the reliability of the method on individual 1 NEX data sets of the test subjects, and its insensitivity to the precise values of initial model parameters. The output of this algorithm is suitable for quantifying cerebral cortical tissue, using a commonly performed commercial pulse sequence. PMID- 11112397 TI - Discrimination of speech and of complex nonspeech sounds of different temporal structure in the left and right cerebral hemispheres. AB - The key question in understanding the nature of speech perception is whether the human brain has unique speech-specific mechanisms or treats all sounds equally. We assessed possible differences between the processing of speech and complex nonspeech sounds in the two cerebral hemispheres by measuring the magnetic equivalent of the mismatch negativity, the brain's automatic change-detection response, which was elicited by speech sounds and by similarly complex nonspeech sounds with either fast or slow acoustic transitions. Our results suggest that the right hemisphere is predominant in the perception of slow acoustic transitions, whereas neither hemisphere clearly dominates the discrimination of nonspeech sounds with fast acoustic transitions. In contrast, the perception of speech stimuli with similarly rapid acoustic transitions was dominated by the left hemisphere, which may be explained by the presence of acoustic templates (long-term memory traces) for speech sounds formed in this hemisphere. PMID- 11112398 TI - Spectroscopic analysis of neural activity in brain: increased oxygen consumption following activation of barrel cortex. AB - This research investigates the hemodynamic response to stimulation of the barrel cortex in anaesthetized rats using optical imaging and spectroscopy (Bonhoeffer and Grinvald, 1996; Malonek and Grinvald, 1996; Mayhew et al., 1999). A slit spectrograph was used to collect spectral image data sequences. These were analyzed using an algorithm that corrects for the wavelength dependency in the optical path lengths produced by the light scattering properties of tissue. The analysis produced the changes in the oxy- and deoxygenation of hemoglobin following stimulation. Two methods of stimulation were used. One method mechanically vibrated a single whisker, the other electrically stimulated the whisker pad. The electrical stimulation intensity varied from 0.4 to 1.6 mA. The hemodynamic responses to stimulation increased as a function of intensity. At 0.4 mA they were commensurate with those from the mechanical stimulation; however, the responses at the higher levels were greater by a factor of approximately 10. For both methods of data collection, the results of the spectroscopic analysis showed an early increase in deoxygenated hemoglobin (Hbr) with no evidence for a corresponding decrease in oxygenated hemoglobin (HbO(2)). Evidence for increased oxygen consumption (CMRO(2)) was obtained by converting the fractional changes in blood volume (Hbt) into estimates of changes in blood flow (Grubb et al., 1974) and using the resulting time course to scale the fractional changes in Hbr. The results show an early increase CMRO(2) peaking approximately 2 s after stimulation onset. Using these methods, we find evidence for increased oxygen consumption following increased neural activity even at low levels of stimulation intensity. PMID- 11112399 TI - Bayesian construction of geometrically based cortical thickness metrics. AB - This paper describes the construction of cortical metrics quantifying the probabilistic occurrence of gray matter, white matter, and cerebrospinal fluid compartments in their correlation to the geometry of the neocortex as measured in 0.5-1.0 mm magnetic resonance imagery. These cortical profiles represent the density of the tissue types as a function of distance to the cortical surface. These metrics are consistent when generated across multiple brains indicating a fundamental property of the neocortex. Methods are proposed for incorporating such metrics into automated Bayes segmentation. PMID- 11112400 TI - Specific versus nonspecific brain activity in a parametric N-back task. AB - In this study functional magnetic resonance imaging (fMRI) was used to examine cerebral activity patterns in relation to increasing mental load of a working memory task. Aim of the experiment was to distinguish nonspecific task-related processes from specific workload processes analytically. Twelve healthy volunteers engaged in a spatial n-back task with four levels. FMRI data were acquired with the 3D-PRESTO pulse sequence. Analysis entailed a two-step multiple regression algorithm, which was specifically designed to measure and separate load-sensitive and load-insensitive activity simultaneously, while preserving the original high spatial resolution of the fMRI signal. Load-sensitive and load insensitive activity was found in both dorsolateral-prefrontal and parietal cortex, predominantly bilaterally, and in the anterior cingulate. As expected, the left primary sensorimotor cortex showed predominantly load-insensitive activity. Load-sensitive activity reflects specific working memory functions, such as temporary retention and manipulation of information, while load insensitive activity reflects supportive functions, such as visual orientation, perception, encoding, and response selection and execution. Good performance was correlated with a large area of load-sensitive activity in anterior cingulate, and with a small area of load-insensitive activity in the right parietal cortex. The findings indicate that nonspecific and specific working memory processes colocalize and are represented in multiple frontal and parietal regions. Implication of this analytical strategy for application in research on psychiatric disorders is discussed. PMID- 11112401 TI - The effect of verbal feedback on motor learning--a PET study. Positron emission tomography. AB - The purpose of this study was to investigate brain mechanisms underlying feedback effects on motor learning. We measured human brain activity using positron emission tomography (PET) during length-of-line drawing tasks in the presence or absence of verbal feedback, i.e., information on the precision of motor performance. The average error in responses was significantly lower and the percentage of correct responses was significantly higher in the case of tasks with feedback than those in the absence of feedback. The contralateral sensorimotor, premotor, supplementary motor, the right prefrontal, bilateral parietal and temporal, and anterior cingulate cortices, and the left basal ganglia were activated during all the line-drawing tasks. The right lateral prefrontal and occipital cortices and the left basal ganglia exhibited marked increase in activity after learning. The right inferior parietal and the anterior cingulate cortices were activated in the presence of feedback which provided information on how the subjects should correct their performances. The results indicate that these brain areas may play an important role in representing knowledge of results during motor learning and that appropriate feedback may facilitate motor learning. PMID- 11112402 TI - Abnormal reactivity of the approximately 20-Hz motor cortex rhythm in Unverricht Lundborg type progressive myoclonus epilepsy. AB - The approximately 20-Hz component of the human mu rhythm originates predominantly in the primary motor cortex. We monitored with a whole-scalp neuromagnetometer the reactivity of the approximately 20-Hz rhythm as an index of the functional state of the primary motor cortex in seven patients suffering from Unverricht Lundborg type (ULD) progressive myoclonus epilepsy (PME) and in seven healthy control subjects. In patients, the motor cortex rhythm was on average 5 Hz lower in frequency and its strength was double compared with controls. To study reactivity of the approximately 20-Hz rhythm, left and right median nerves were stimulated alternately at wrists. In controls, these stimuli elicited a small transient decrease, followed by a strong increase ("rebound") of the approximately 20-Hz level. In contrast, the patients showed no significant rebounds of the rhythm. As the approximately 20-Hz rebounds apparently reflect increased cortical inhibition, our results indicate that peripheral stimuli excite motor cortex for prolonged periods in patients with ULD. PMID- 11112403 TI - The role of the dorsolateral prefrontal cortex in random number generation: a study with positron emission tomography. AB - Random number generation (RNG) engages a number of executive processes. We used positron emission tomography (PET) to measure regional cerebral blood flow (rCBF) in six volunteers who performed RNG and a control counting (COUNT) task at six rates paced by a tone. This provided a systematic variation of difficulty of RNG. Relative to COUNT, RNG was associated with significant activation of the left dorsolateral prefrontal cortex (DLPFC), the anterior cingulate, the superior parietal cortex bilaterally, the right inferior frontal cortex, and the left and right cerebellar hemispheres. Faster rates of RNG were associated with a significant decrease in regional cerebral blood flow (rCBF) in the left and right DLPFC and the right superior parietal cortex. rCBF in the left DLPFC was significantly and negatively associated with count score 1, a measure of habitual counting during RNG. These results are discussed in relation to the network modulation model of RNG developed on the basis of our previous studies using transcranial magnetic stimulation and dual task paradigms. This suggests that during RNG, suppression of habitual counting is achieved through the modulatory (inhibitory) influence of the left DLPFC over a number associative network distributed in the superior temporal cortex. At faster rates of RNG the synchronization demands of paced RNG result in the breakdown of this modulatory influence, which is evident from decreased rCBF in the left DLPFC and increased habitual counting at faster rates. PMID- 11112404 TI - Automatic morphology-based brain segmentation (MBRASE) from MRI-T1 data. AB - A method called morphology-based brain segmentation (MBRASE) has been developed for fully automatic segmentation of the brain from T1-weighted MR image data. The starting point is a supervised segmentation technique, which has proven highly effective and accurate for quantitation and visualization purposes. The proposed method automates the required user interaction, i.e., defining a seed point and a threshold range, and is based on the simple operations thresholding, erosion, and geodesic dilation. The thresholds are detected in a region growing process and are defined by connections of the brain to other tissues. The method is first evaluated on three computer simulated datasets by comparing the automated segmentations with the original distributions. The second evaluation is done on a total of 30 patient datasets, by comparing the automated segmentations with supervised segmentations carried out by a neuroanatomy expert. The comparison between two binary segmentations is performed both quantitatively and qualitatively. The automated segmentations are found to be accurate and robust. Consequently, the proposed method can be used as a default segmentation for quantitation and visualization of the human brain from T1-weighted MR images in routine clinical procedures. PMID- 11112405 TI - T2 relaxometry can lateralize mesial temporal lobe epilepsy in patients with normal MRI. AB - In unselected patients with intractable temporal lobe epilepsy (TLE), approximately 15% do not have detectable hippocampal atrophy on MRI. The purpose of this study was to evaluate whether T2 relaxometry can identify hippocampal pathology and lateralize the epileptic focus in patients with intractable TLE, who do not demonstrate hippocampal atrophy on volumetric MRI (MRIV). We selected 14 patients with unilateral TLE who had unilateral atrophy and 11 patients with unilateral TLE who had no evidence of atrophy on MRIV. Images were acquired on a 1.5 T MR scan using a dual echo sequence with 23 contiguous oblique coronal slices in all patients and in 14 healthy subjects. Fitting a single exponential decay equation to the imaging data generated T2 maps. Averages of six slices containing the head, body, and tail of the hippocampus were used to calculate hippocampal T2 relaxation times (HT2). The epileptic focus was defined by history, video-EEG, and surgical response. All TLE patients with hippocampal atrophy and 9/11 (82%) patients with normal MRI had abnormally high HT2 ipsilateral to the epileptic focus. Bilateral abnormal HT2 were found in 6/14 (43%) of patients with unilateral hippocampal atrophy and 2/11 (18%) of patients with normal MRI. However, this increase was always greater ipsilateral to the epileptic focus. Qualitative hippocampal pathology showed gliosis and neuronal loss in 10/14 operated patients with hippocampal atrophy on MRIV and in 5/7 operated patients with normal MRI. In conclusion, hippocampal T2 mapping provides evidence of hippocampal damage in the majority of patients with intractable TLE who have no evidence of atrophy on MRI and can correctly lateralize the epileptic focus in most patients. PMID- 11112406 TI - Dual voxel proton magnetic resonance spectroscopy in the healthy elderly: subcortical-frontal axonal N-acetylaspartate levels are correlated with fluid cognitive abilities independent of structural brain changes. AB - The published literature suggests that degeneration of the subcorticofrontal networks may underlie cognitive ageing, but appropriate methods to examine this in vivo have been lacking. Proton Magnetic Resonance Spectroscopy ((1)H-MRS) has now been used in a number of clinical studies to assess cerebral pathophysicochemistry and recently has been utilized to examine the relationship between neurochemical markers and cognitive functioning in normal individuals. Results have been somewhat conflicting and difficult to interpret. To further clarify the role of the cognitive spectroscopy technique, we measured N acetylaspartate (NAA) levels in the frontal subcortical white matter and the occipitoparietal grey matter and correlated them with performance in different cognitive domains in a group of twenty healthy elderly individuals. Subjects underwent whole brain T(1)- and T(2)-weighted magnetic resonance imaging (MRI), dual voxel short echo-time (1)H-MRS, and a comprehensive neuropsychological assessment. Individual tests of executive and attentional abilities, and a principal components composite score reflecting these skills, but not measures of memory or verbal abilities, were correlated with NAA concentration in the frontal white matter only. These relationships were independent of other neurocognitive predictors of executive impairment such as age, midventricular dilation, frontal white matter disease, and presenescent verbal proficiency. This study suggests the ability of (1)H-MRS to differentiate anatomically distinct neurochemical markers related to specific cognitive abilities. In particular, neurometabolic fitness of the frontal subcortical-cortical axonal fibers may be important in mediating fluid intellectual processing. Longitudinal MRS studies are required to determine if the present results reflect different rates of neurocellular degeneration or preexisting individual differences in neuronal density. PMID- 11112407 TI - Assessing the performance of SPM analyses of spect neuroactivation studies. Statistical Parametric Mapping. AB - Several simulations of SPECT neuroactivation studies have been performed in order to determine the influence of both study size and activation focus characteristics on the detection of brain activation foci following a pixel-based statistical analysis. This was achieved by developing a methodology based on the Hoffman software brain phantom, SPECT acquisition simulation software, standard reconstruction software, and the Statistical Parametric Mapping (SPM96) package. We present results on the minimal activation levels required for focus detection. Furthermore, the improved sensitivity of the analysis resulting from the use of an iterative reconstruction technique (OSEM) with regard to the classical filtered backprojection (FBP) is assessed quantitatively, and the various physical, processing, and physiological parameters that potentially influence the detection of foci are discussed. Finally, the influence is investigated of the height threshold as implemented in SPM96 upon the size of the detected foci. Practical guidelines are proposed with regard to the number of subjects per group for SPECT activation studies following the split-dose design. PMID- 11112408 TI - Synergistic effect of TNF-alpha and IL-4 on the expression of thymus- and activation-regulated chemokine in human corneal fibroblasts. AB - Chemokine production by resident fibroblasts contributes to the recruitment of migratory leukocytes to sites of tissue injury. The effects of cytokines on the expression of thymus- and activation-regulated chemokine (TARC), a potent chemoattractant for Th2 cells, were examined in cultured human corneal fibroblasts. The culture supernatants of cells incubated for 24 h in the absence or presence of either TNF-alpha (10 ng/ml) or IL-4 (10 ng/ml) alone did not contain TARC detectable by ELISA. However, exposure of cells to both cytokines resulted in marked release of TARC into the culture medium in a time- and dose dependent manner. Similarly, quantitative RT-PCR analysis revealed that the two cytokines induced a synergistic increase in the amount of TARC mRNA in the cultured fibroblasts. This synergistic effect of TNF-alpha and IL-4 on TARC production by fibroblasts may contribute to the Th2 cell infiltration and consequent tissue damage associated with allergic inflammation. PMID- 11112409 TI - Interaction of Daxx, a Fas binding protein, with sentrin and Ubc9. AB - Sentrin is a ubiquitin-like protein that can covalently modify cellular proteins, and is a Fas binding protein that protects cells against anti-Fas induced cell death. However, the mechanism by which sentrin exerts its effect upon Fas mediated apoptosis is not well known. Thus, this study examined the interaction of sentrin with Daxx. Sentrin interacted with Daxx but not with FADD when analyzed by yeast two-hybrid assay. In vitro translated Daxx bound to GST-sentrin fusion protein. FLAG-sentrin fusion protein was also coimmunoprecipitated with Daxx in BOSC23 cells. Also, Daxx interacted with Ubc9, an essential protein as a key conjugating enzyme. Amino acids 625-740 of Daxx, known as Fas binding region, was also mapped as sentrin and Ubc9 binding region. Colocalization of Fas, sentrin, and Ubc9 binding regions suggests the importance of that region upon the regulation of Daxx. Our data also demonstrated that sentrin could homooligomerize by protein-protein interaction. PMID- 11112410 TI - Properties and secondary structure analysis of BanI endonuclease: identification of putative active site. AB - Biochemical properties of Type II restriction enzyme BanI were characterized. Kinetic parameters were evaluated and an enhancement of rate was observed when the recognition site was located in a more central position in the substrate, suggesting that BanI locates its recognition site by a sliding mechanism. As BanI has three cysteine residues in its primary sequence, the effect of thiol inhibitors on BanI activity was also studied. Partial inhibition was observed only at a very high concentration of the inhibitor indicating that cysteine residues are not directly involved in catalysis. The gel electrophoretic mobility shift assay demonstrated specific complex formation between BanI and the DNA substrate in the presence of poly dI-dC and Mg(2+). A secondary structure analysis and comparison with EcoRI and BamHI crystal structure revealed a putative active site similar to that seen in BamHI but different in the order in which the catalytic domain (central beta-sheet) and recognition domain (adjacent alpha-helix) were arranged in the protein. PMID- 11112411 TI - delta(6)-desaturase of Mucor rouxii with high similarity to plant delta(6) desaturase and its heterologous expression in Saccharomyces cerevisiae. AB - Gamma-linolenic acid (GLA, gamma-C18:3) is an essential fatty acid that plays a vital role in biological structures and cellular functions. Based on available sequence information and using polymerase chain reaction (PCR) technique, we cloned from the fungus Mucor rouxii the entire coding sequence of a delta(6) desaturase enzyme, which is responsible for the transformation of linoleic acid into GLA. The deduced amino acid sequence of M. rouxii gene showed the highest homology with the plant delta(6)-desaturase. It comprises the characteristics of membrane-bound desaturases, including histidine-rich motifs and hydrophobic regions. A cytochrome b(5)-like domain was observed at the N-terminus. In addition to three conserved histidine-rich motifs, we found an additional histidine-rich motif, HKHHSH, downstream of the cytochrome b(5)-like domain, which is not present in previously cloned delta(6)-desaturase genes. Heterologous expression of the M. rouxii cDNA in Saccharomyces cerevisiae resulted in the synthesis and accumulation of GLA. PMID- 11112412 TI - The effect of ethanol-induced cytochrome p4502E1 on the inhibition of proteasome activity by alcohol. AB - The present investigation was undertaken to determine the effect of CYP2E1 induction by ethanol on the inhibition of proteasomal activity in wild-type and CYP2E1 knockout C57 black mice. The proteasomal chymotrypsin-like activity decreased significantly in ethanol-fed wild-type mice liver, but was not reduced in ethanol-fed knockout mice liver. The 26S proteasomal activity was decreased more by ethanol feeding than was the 20S proteasomal fraction. Individual hepatocytes lost immunostaining of the proteasomes in the centrilobular zone in the livers of ethanol-fed wild-type mice and the knockout mouse liver. There was increased product of protein oxidation in the liver in the wild type but not in the knockout mice given ethanol. Taken together, these results suggest that CYP2E1 induction was responsible for the decrease in proteasome activity seen in the wild-type mice which head to the accumulation of oxidized proteins which were increased as the result of free radicals generated by CYP2E1 metabolism of ethanol. PMID- 11112413 TI - iNOS expression inhibits hypoxia-inducible factor-1 activity. AB - Hypoxia-inducible factor-1 (HIF-1) activates genes important in vascular function such as vascular endothelial growth factor (VEGF), erythropoietin (EPO), and inducible nitric oxide synthase (iNOS). iNOS catalyzes the synthesis of nitric oxide (NO), a free radical gas that mediates a number of cellular processes, including regulation of gene expression, vasodilatation, and neurotransmission. Here we demonstrate that iNOS expression inhibits HIF-1 activity under hypoxia in C6 glioma cells transfected with an iNOS gene and a VEGF promoter-driven luciferase gene. HIF-1 induction of VEGF-luciferase activity in C6 cell is also inhibited by sodium nitroprusside (SNP). Furthermore, pretreatment of C6 cells with N-acetyl-l-cysteine (NAC), an antioxidant, nullified the inhibitory effect of iNOS on HIF-1 binding. These results demonstrate that NO generated by iNOS expression inhibits HIF-1 activity in hypoxic C6 cells and suggest a negative feedback loop in the HIF-1 --> iNOS cascade. PMID- 11112414 TI - Involvement of prolactin-releasing peptide in the preovulatory luteinizing hormone and prolactin surges in the rat. AB - Prolactin (PRL)-releasing peptide (PrRP) is a novel hypothalamic peptide reported as a potent and specific stimulator of PRL secretion. In this study, we examined a possible role of PrRP in the ovarian steroid-induced PRL surge in the rat, simultaneously observing the change in luteinizing hormone (LH) surge. Experiments were performed on both normally-fed and three-day-fasted rats, which were ovariectomized and primed with estradiol and progesterone. From 11:00 to 18:00 h, blood was collected every 30 min to measure LH and PRL. All the following substances were given intracerebroventricularly at 11:00 h. Compared to control serum, anti-rat PrRP31 serum caused a significant reduction of the LH and PRL surges. The antiserum also delayed the onset of PRL surge. Fasted rats were devoid of significant surges of the hormones, while 3.0, but not 0.5 nmol of rat PrRP31 given to these animals produced a significant recovery of PRL surge. Although LH surge was not reinstated, basal LH secretion was transiently stimulated by 3.0 nmol of PrRP31. These results demonstrate for the first time a significant participation of PrRP in the preovulatory LH and PRL surges in the rat. Possible indirect pathways mediating this effect of PrRP were discussed, in view of the unique anatomical distribution of PrRP in the hypothalamus. PMID- 11112415 TI - Differentiation-dependent cytoplasmic distribution and in vivo RNA association of proteins recognized by the 3'-UTR stability element of alpha-globin mRNA in erythroleukemic cells. AB - In this study, we analyzed subcytoplasmic distribution and in vivo RNA association of proteins with specific affinity to cytosine-rich stability determinant sequences of alpha-globin mRNA 3'-UTR in a MEL-707 erythroleukemic model. We took advantage of the possibility that these cells can be reversibly differentiated (as a continuous population, but not at the level of individual cells) which, therefore, allows analysis of various stages of erythroid differentiation within the same cell population. Label transfer experiments revealed four major complexes with molecular mass of 110-, 70-, 55- and 50-kDa in various cytoplasmic fractions. Using the combination of in vitro label transfer and in vivo UV-crosslinking techniques, we also demonstrated that subcytoplasmic distribution as well as in vivo RNA association of various complex-forming proteins is differentiation dependent in MEL-707 cells. These results indicate that changes in the cytoplasmic environment imposed by the differentiating stimulus might direct important biochemical signals as to how the stability determinant 3'UTR elements interact with their binding proteins. These data also suggest that stability complexes are dynamic macromolecular structures with high response capacity to various extra- and intracellular regulatory stimuli. PMID- 11112416 TI - 17 beta-estradiol inhibits tumor necrosis factor-alpha-induced nuclear factor kappa B activation by increasing nuclear factor-kappa B p105 level in MCF-7 breast cancer cells. AB - Tumor necrosis factor-alpha (TNF-alpha) exerts many cytological effects on a wide range of cells. TNF-alpha can activate nuclear factor-kappa B (NF-kappa B). Activation of NF-kappa B by TNF-alpha mediates many functions of TNF-alpha. The NF-kappa B inhibitor, I kappa B alpha, negatively regulates the activity of NF kappa B. In MCF-7 cells (an estrogen and TNF-alpha receptor positive cell line), treatment with 17 beta-estradiol (E(2)) inhibited TNF-alpha-induced NF-kappa B DNA binding activity in the gel retardation assays. But, the level of the I kappa B alpha and the TNF-alpha receptor, TNF-R1, were not obviously affected. The NF kappa B precursor, NF-kappa B p105, has been shown to be associated with NF-kappa B in the cytoplasm and efficiently blocks its nuclear translocation and activation. Treatment of MCF-7 cells with E(2) increased the level of NF-kappa B p105 protein. The anti-estrogen, 4OH-tamoxifen, treatment inhibited E(2)-induced NF-kappa B p105 expression. Our findings indicate that NF-kappa B p105 plays a role in modulating the functions of TNF-alpha in the estrogen receptor positive breast cancer cells. PMID- 11112417 TI - Expression and Up-regulation of alternatively spliced transcripts of melastatin, a melanoma metastasis-related gene, in human melanoma cells. AB - Loss of expression of a novel suppressor of metastasis, melastatin (MLSN1), has recently been reported to correlate with metastatic potential of melanoma cells. Using differential display analysis, we identified MLSN1 among genes overexpressed in pigmented metastatic human melanoma cells treated with the differentiation inducer hexamethylene bisacetamide (HMBA). In this study, we show that multiple short transcripts of MLSN1 are present in melanocytes and pigmented metastatic melanoma cell lines while the full-length 5. 4-kb mRNA is detectable only in melanocytes. Treatment of pigmented melanoma cells with the differentiation-inducing agent, HMBA, results in up-regulation of the 5.4-kb MLSN1 mRNA as well as short RNAs. Analysis of a panel of nonpigmented primary and metastatic melanoma cell lines showed weak expression of a 1.8-kb mRNA in a few melanoma cell lines. Northern blot and RT-PCR analyses with DNA probes and oligonucleotide primers that correspond to distinct regions of full-length MLSN1 mRNA indicated that the short transcripts contained sequences corresponding primarily to either 5'- or 3'-end of the 5.4-kb mRNA. HMBA appears to up-regulate MLSN1 transcripts derived mainly from the 5'-end. Modulators of cAMP and protein kinase C pathways had no significant effect on MLSN1 expression. Our data show that multiple MLSN1 transcripts, both constitutively expressed and inducible, are present in cultured pigmented melanoma cells, and suggest that MLSN1 expression can be regulated at the level of both transcription and mRNA processing. PMID- 11112418 TI - Novel acyl-CoA synthetase in adrenoleukodystrophy target tissues. AB - X-linked adrenoleukodystrophy (X-ALD) is a neurodegenerative disorder characterized by demyelination of white matter. The X-ALD gene product adrenoleukodystrophy protein (ALDP) is expressed broadly among various tissues. However, deficiency of functional ALDP exclusively impairs brain, adrenal gland, and testis. Thus, loss of ALDP function is assumed to involve inactivation of a putative mediating factor that functions in a tissue-specific manner. Here we cloned a mouse cDNA encoding a novel protein, Lipidosin, that possesses long chain acyl-CoA synthetase (LCAS) activity. Lipidosin is expressed exclusively in mouse brain, adrenal gland, and testis, which are affected by X-ALD. LCAS activity of Lipidosin was diminished by mutation of conserved amino acids within the AMP-binding domain. Mutation of the Drosophila homologue of Lipidosin has been reported to cause neuronal degeneration. Thus, Lipidosin may mediate the link between ALDP dysfunction and the impairment of fatty acid metabolism in X ALD. PMID- 11112419 TI - Identification of 6-furfuryladenine (kinetin) in human urine. AB - In contrast to the current view of kinetin (K, N(6)-furfuryladenine) as an unnatural and synthetic cytokinin, recently it has been identified in plant DNA and plant extract. Here we describe identification of K in human urine using chromatography/mass-spectrometry analysis for the first time. The amount of kinetin in urine taken from unhealthy patients lung carcinoma was established to be 0.5 ng in 20 ml and a 100-fold reduced amount in healthy subjects. Since this rare base is a potential source of structural constrains it has to be removed from DNA by enzymatic DNA-repair reactions. It seems that the presence of kinetin in human is linked to oxidative damage processes. PMID- 11112420 TI - A robust cysteine-deficient chitinase-like antifungal protein from inner shoots of the edible chive Allium tuberosum. AB - From the inner shoots of the chive Allium tuberosum, a single-chained protein with a molecular weight of 36 kDa and an N-terminal sequence manifesting resemblance to chitinases but lacking in cysteine residues characteristic of a cysteine-rich domain present in chitinases of other Allium species, was purified. The isolation procedure entailed affinity chromatography on Affi-gel blue gel, ion-exchange chromatography on DEAE-cellulose and Mono S, and gel filtration on Superdex 75. The protein was unadsorbed on DEAE-cellulose and adsorbed on Affi gel blue gel and Mono S. It exhibited antifungal activity against Rhizoctonia solani, Fusarium oxysporum, Coprinus comatus, Mycosphaerella arachidicola, and Botrytis cinerea. The IC(50) for its antifungal effect against Botrytis cinerea was 0.2 microM. The antifungal activity was stable after 1 h at pH 1.6 and 12.3, and up to 60 degrees C for 5 min. Incubation of the protein with trypsin or chymotrypsin at an enzyme:substrate ratio of 1:100 and pH 7.6 up to 150 min did not affect its antifungal activity. The protein did not exhibit antibacterial activity. The protein inhibited cell-free translation in a rabbit reticulocyte system with an IC(50) of 0.8 microM, but did not affect the proliferation of mouse splenocytes. It exerted some cytotoxic effect on breast cancer cells and was inhibitory toward HIV-1 reverse transcriptase. PMID- 11112421 TI - Regulation of the rat liver carnitine palmitoyltransferase I gene transcription by thyroid hormone. AB - L-CPT I isotype is the main locus of control for liver LCFA oxidation. T3 levels have been described as controlling L-CPT I gene expression, and in this paper we demonstrate that rat liver CPT I promoter responds to T3. Using deleted reporter constructs we located the thyroid hormone-responsive element between -2935 and 2918, consisting of a DR4. This response is mediated by the binding of the thyroid to this sequence as a monomer, homodimer, or heterodimer with RXR. PMID- 11112422 TI - Vitamin D receptor expression is linked to cell cycle control in normal human keratinocytes. AB - To improve our understanding of the cutaneous vitamin D system, we studied vitamin D receptor (VDR) gene regulation in cultured human keratinocytes. Because VDR and its ligand 1 alpha,25-dihydroxyvitamin D(3) have been implicated in epidermal growth control, we investigated VDR expression as related to cellular proliferation by using different cell cycle synchronization protocols. Keratinocytes, deprived of growth factors, were forced into quiescence and a concomitant loss of VDR expression was observed. Mitogenic stimulation of these G(0) cells however quickly upregulated VDR levels several hours ahead the G(1)-S transition point. Growth arrest at the G(1)-S border by mimosine treatment or at the metaphase by nocodazole also downregulated VDR levels but a restoration of VDR expression was again quickly achieved after reentering the cell cycle. These findings indicate that VDR expression in keratinocytes is restricted to actively cycling cells, but not limited to one particular phase of the cell cycle. PMID- 11112423 TI - Distinct regions of the chicken p46 polypeptide are required for its in vitro interaction with histones H2B and H4 and histone acetyltransferase-1. AB - We cloned cDNA encoding the chicken p46 polypeptide, chp46, homologous to the p48 subunit of chicken chromatin assembly factor-1, chCAF-1p48. It comprises 424 amino acids including a putative initiation Met, is a member of the WD protein family, with seven WD repeat motifs, and exhibits 90.3% identity to chCAF-1p48 and 94.3% identity to the human and mouse p46 polypeptides (hup46 and mop46). The in vitro immunoprecipitation experiment established that chp46 interacts with histones H2B and H4 and chicken histone acetyltransferase-1, chHAT-1, whereas hup46 interacts with histones H2A and H4 and chHAT-1 and chCAF-1p48 with histone H4 and chHAT-1. The in vitro immunoprecipitation experiment, involving truncated mutants of chp46, revealed not only that two regions comprising amino acids 33 179 and 375-404 are necessary for its binding to H2B, but also that two regions comprising amino acids 1-32 and 405-424 are necessary for its binding to H4. Furthermore, the GST pulldown affinity assay, involving truncated mutants of chp46, revealed that a region comprising amino acids 359-404 (in fact, 375-404) binds to chHAT-1 in vitro. Taken together, these results indicate not only that chp46 should participate differentially in a number of DNA-utilizing processes through interactions of its distinct regions with chHAT-1 and histones H2B and H4, but also that the proper propeller structure of chp46 is not necessary for its interaction with chHAT-1. PMID- 11112424 TI - Overexpression of protein kinase C-eta attenuates caspase activation and tumor necrosis factor-alpha-induced cell death. AB - The protein kinase C (PKC) signal transduction pathway regulates cell death by tumor necrosis factor-alpha (TNF). We previously showed that the induction of novel PKC eta isozyme by PKC activators correlated with their ability to protect MCF-7 breast cancer cells against TNF cytotoxicity. In the present study, we have transfected PKC eta in MCF-7 cells to directly examine its involvement in cell death by TNF. Overexpression of PKC eta delayed TNF-induced cell death in MCF-7 cells. TNF caused a rapid activation of caspase-8 and -7 in cells transfected with a vector. The activation of these caspases was potentiated by the PKC inhibitor bisindolylmaleimide (BIM) which downregulates PKC eta and sensitizes cells to TNF. Overexpression of PKC eta delayed the activation of caspase-8 and 7 by both TNF and the combination of BIM and TNF. These results suggest that PKC eta protects MCF-7 cells against TNF-induced cell death by preventing the activation of caspases. PMID- 11112425 TI - Regulation of HGF/SF gene expression in MRC-5 cells by N-acetylcysteine. AB - The effect of N-acetylcysteine (NAC) on levels of hepatocyte growth factor/scatter factor (HGF/SF) gene transcripts was investigated in the human lung embryonic fibroblast cell line, MRC-5. NAC increased expression of HGF/SF mRNA, in a dose- and time-dependent fashion, by a mechanism independent of glutathione synthesis but sensitive to oxidant stress induced by H(2)O(2). Using actinomycin D to block RNA synthesis, it was observed that NAC had no effect on the stability of the HGF/SF mRNA transcripts. NAC increased HGF/SF promoter activity in cells transiently transfected with chloramphenicol acetyltransferase (CAT) reporter genes driven by HGF/SF gene 5'-flanking sequences. Primer extension analysis demonstrated that NAC enhanced the expression of HGF/SF mRNA transcribed from the main transcription initiation site. Although the 5' flanking region of the HGF/SF gene contains a sequence at -1019 to -1011 with homology to the NF-kappaB response element, electrophoretic mobility shift assay demonstrated that this site did not bind nuclear factors in MRC-5 cells in the presence or absence of NAC. In contrast to the effect on HGF/SF mRNA, NAC did not increase HGF/SF protein production by MRC-5 cells. PMID- 11112426 TI - Myoepithelial-specific CD44 shedding is mediated by a putative chymotrypsin-like sheddase. AB - Our previous studies have demonstrated that myoepithelial cells, which surround incipient carcinomas in situ of the breast and other organs, exert antiinvasive and antiangiogenic effects in vitro through the elaboration of a number of different suppressor molecules among which include the shed membrane CD44. The present study addresses the mechanism of this myoepithelial CD44 shedding. This CD44 shedding is enhanced by PMA pretreatment, is specific for myoepithelial CD44, and inhibited by chymotrypsin-like inhibitors (chymostatin, alpha(1) antichymotrypsin, TPCK, and SCCA-2) but not by trypsin-like inhibitors (TLCK), nor papain-like inhibitors (SCCA-1) nor hydroxamate-based or general metalloproteinase inhibitors (BB2516 (marimastat), 1,10-phenanthroline, and TIMP 1). The effect of PMA can be mimicked by exogenous chymotrypsin but not by other proteases. The CD44 shedding activity cannot be transferred by conditioned media, cell-cell contact, peripheral membrane, or integral membrane fractions. However, cell-free purified integral plasma membrane fractions obtained from myoepithelial cells pretreated with PMA also exhibit CD44 shedding which is inhibited by chymotrypsin-like inhibitors. These findings support the presence and activation of a putative chymotrypsin-like sheddase as the mechanism of CD44 shedding in myoepithelial cells. PMID- 11112427 TI - Carrier-mediated uptake of rhodamine 123: implications on its use for MDR research. AB - We have examined the effects of verapamil and PSC 833 on cellular uptake and release of rhodamine 123 (R123) in two human cancer cell lines. Both verapamil and PSC 833 were able to increase R123 accumulation in the multidrug resistant (MDR) MV522/Q6 and KB-8-5 lines in the release study. However, the effects of these drugs on R123 accumulation during accumulation study were quite different. Incubation with PSC 833 increased R123 accumulation in both MDR lines. By contrast, incubation with verapamil only increased R123 accumulation in the KB-8 5 line. The failure of verapamil to increase R123 accumulation in the MV522/Q6 cells can be attributed to the presence of a carrier system in the parent MV522 cells that recognizes both R123 and verapamil, but not PSC 833, as substrates. These results imply that performing R123 accumulation study without first ascertaining possible role of a carrier system for cellular uptake of R123 and putative P-gp modulators might inadvertently lead one to draw improper conclusions on P-gp activity. PMID- 11112428 TI - Molecular characterization and nuclear localization of rat timeless-like gene product. AB - Among three period genes (per1, per2, per3) in mammals, only per2 gene was shown to be involved in the core clock mechanism. To elucidate the molecular function of rat PERIOD2 (rPER2), we searched for binding proteins to the PAS domain of rPER2. We isolated a binding protein to this domain and identified it as a TIMELESS-like protein (TLP) on the basis of mass analyses. Then, we isolated a rat TLP cDNA from the rat hypothalamus library. RNA blot analysis and in situ hybridization indicates that rTLP mRNA was expressed in all rat tissues from whole brain, the suprachiasmatic nucleus, eye, lung, heart, liver, kidney, placenta, and testis. When rTLP gene product was expressed in COS-1 cells, nuclear localization of rTLP was detected in 99.6% of transfected cells. These results suggest that the interaction of rPER2 with rTLP may influence the regulation of circadian clock components in nucleus after rPER2 is translocated into the nucleus. PMID- 11112429 TI - Characterization of a zebrafish (Danio rerio) sphingosine 1-phosphate receptor expressed in the embryonic brain. AB - Sphingosine 1-phosphate elicits a variety of responses in mammals via at least five G protein-coupled Edg receptors. We cloned zebrafish edg1 and expressed it in Rh7777 cells. In these cultures, S1P inhibited forskolin-driven rises in cAMP and this response was eliminated by pretreatment of the cultures with pertussis toxin. In Rh7777 membranes, S1P stimulated GTPgamma[(35)S] binding 2-3 fold. Zebrafish edg1 is expressed in embryonic brain, particularly ventral diencephalon, optic stalks, and anterior hindbrain. Our findings suggest that nonmammalian vertebrates use S1P to signal during embryogenesis and that the properties of Edg1 receptor have been conserved for 400 million years. PMID- 11112430 TI - Determination of functional regions of p125, a novel mammalian Sec23p-interacting protein. AB - The Sec23p-Sec24p complex is a component of coat protein II-coated vesicles involved in protein export from the endoplasmic reticulum. We previously identified a novel Sec23p-interacting protein, p125, which consists of 1000 amino acids and comprises a proline-rich region and a phospholipase A(1) homology region. p125, when ectopically expressed in cultured cells, localizes to endoplasmic reticulum-Golgi intermediate regions. In the present study we showed that expressed p125 principally colocalizes with p115 and GM130, both of which are involved in vesicle tethering to Golgi membranes. Next, we determined the functional regions of p125 by expressing a p125 series with deletions. The results showed that the proline-rich region (residues 135-259) is responsible for the binding to Sec23p. For the correct localization of p125, a region (residues 135-1000) comprising both the proline-rich and phospholipase A(1) homology regions was required. PMID- 11112431 TI - Regulation of shortening velocity by calponin in intact contracting smooth muscles. AB - To elucidate the function of calponin in intact contracting smooth muscle cells in vivo, we generated mice with a mutated basic calponin (h1) locus (Yoshikawa et al., Genes Cells 3, 685-695, 1998). Crossbridge cycling rates were estimated in aortic smooth muscle by the force redevelopment following an isometric step shortening as a function of time after K(+) depolarization. Evidence is presented that calponin is involved in the inhibition of shortening velocity in the tonic phase of contraction. The phosphorylation levels of myosin regulatory light chain and cytosolic calcium concentrations were not significantly different in paired comparisons between calponin-deficient (-/-) and wild-type (+/+) muscles at any time point after stimulation. The force-velocity relationships in vas deferens smooth muscle showed that the maximum shortening velocity of -/- muscle was significantly faster than that of +/+ muscle. There was no change in the length force relationships in both -/- and +/+ muscles of aorta and vas deferens. The results suggest that calponin plays a role in regulation of the crossbridge cycling and that it may be responsible for reduced shortening velocity during a maintained contraction of mammalian smooth muscle. PMID- 11112432 TI - Inhibitory effects of an antisense oligonucleotide in an experimentally infected mouse model of influenza A virus. AB - The antiviral effects of a 20-mer antisense phosphorothioate oligonucleotide, PB2 as, on influenza A virus infection in mice were examined and compared to those of PB2-as encapsulated with several cationic liposomes. Intravenous injection of PB2 as, as a complex with DMRIE-C, a cationic liposome, was most effective for prolonging the mean survival time in days (MSDs) and increasing the survival rates of mice infected with the influenza A virus. In addition, the liposomal PB2 as significantly inhibited viral growth in lung tissues. These results suggest that PB2-as encapsulated with DMRIE-C may be active against the influenza A virus infection through the inhibition of virus replication in the mouse lung. PMID- 11112433 TI - Down-regulated expression of TWEAK mRNA in acute and chronic inflammatory pathologies. AB - TWEAK is a newly identified member of the Tumor Necrosis Factor (TNF) family of proteins which are involved in many immunoinflammatory mechanisms. The putative role of TWEAK in inflammation was analyzed in mice treated with lipopolysaccharide (LPS), a strong inducer of the immuno-inflammatory responses. TWEAK mRNA rapidly disappeared in all the tissues tested. Analysis of LPS-treated thioglycolate-elicited peritoneal macrophages revealed that the rapid loss of TWEAK mRNA was due to its active destabilization. In chronic pathologies like autoimmune hemolytic anemia in the NZB mouse strain or systemic lupus erythematosus (SLE) in the BXSB mouse strain, TWEAK mRNA was shown to be reduced concomitantly to the development of chronic autoimmune diseases. These results demonstrated that TWEAK mRNA, contrary to TNF mRNA, is stable, ubiquitously distributed in tissues, and is down-regulated after LPS treatment or in chronic inflammation, suggesting that TWEAK could be an important factor, along with TNF, in acute and chronic inflammations. PMID- 11112434 TI - PEA3 and AP-1 are required for constitutive IL-8 gene expression in hepatoma cells. AB - Interleukin-8 (IL-8) mRNA was constitutively expressed in human hepatoma cell line, HepG2 and in human hepatocellular carcinoma (HCC), which often form hypervascular tumors. The sequence 5'-AGGAAG-3' at -137 to -132 bp of IL-8 promoter was shown to be polyomavirus enhancer A binding protein-3 (PEA3) binding site, which can cooperate with activator protein-1 (AP-1). Both PEA3 and AP-1 are essential for constitutive IL-8 expression in HepG2 cells, determined by promoter assays. Moreover, PEA3 and IL-8 proteins coexisted in HCC tissues, but not in uninvolved liver tissues. It is possible PEA3 may have important roles in tumor progression and in angiogenesis in HCC. PMID- 11112435 TI - bFGF induces differentiation and death of olfactory neuroblastoma cells. AB - Olfactory neuroblastoma (ONB) is a highly vascularized and malignant tumor arising in olfactory neuronal precursors from the paranasal sinuses. Previously, we showed that treatment of JFEN cells with transforming growth factor (TGF) alpha caused them to differentiate and respond to chemical odorants, whereas basic fibroblast growth factor (bFGF) treated cells differentiated and died. In the present study we show that established ONB tumors treated with bFGF upregulate the bFGF receptor (FGFR1) prior to differentiation. This cellular differentiation was evidenced by bFGF-induced expression of the human runt homologue AML1 (PEBP2 alpha B, CBFA-2) that is highly expressed in developing olfactory neuroepithelium and TrkA, a preferred nerve growth factor receptor. Since TrkA is expressed in supporting cells, but not in mature olfactory neurons, we hypothesize that the expression of AML1 and TrkA in bFGF-treated JFEN cells induced supporting cell differentiation. Collectively, these results have implications for the treatment of patients afflicted with ONB. PMID- 11112436 TI - Phosphorylation of anchoring protein by calmodulin protein kinase associated to the sarcoplasmic reticulum of rabbit fast-twitch muscle. AB - Regulatory phosphorylation of phospholamban and of SR Ca(2+)-ATPase SERCA2a isoform by endogenous CaM-K II in slow-twitch skeletal and cardiac sarcoplasmic reticulum (SR) is well documented, but much less is known of the exact functional role of CaM K II in fast-twitch muscle SR. Recently, it was shown that RNA splicing of brain-specific alpha CaM K II, gives rise to a truncated protein (alpha KAP), consisting mainly of the association domain, serving to anchor CaM K II to SR membrane in rat skeletal muscle [Bayer, K.-U., et al. (1998) EMBO J. 19, 5598-5605]. In the present study, we searched for the presence of alpha KAP in sucrose-density purified SR membrane fractions from representative fast-twitch and slow-twitch limb muscles, both of the rabbit and the rat, using immunoblot techniques and antibody directed against the association domain of alpha CaM K II. Putative alpha KAP was immunodetected as a 23-kDa electrophoretic component on SDS-PAGE of the isolated SR from fast-twitch but not from slow-twitch muscle, and was further identified as a specific substrate of endogenous CaM K II, in the rabbit. Immunodetected, (32)P-labeled, non-calmodulin binding protein, behaved as a single 23-kDa protein species under several electrophoretic conditions. The 23 kDa protein, with defined properties, was isolated as a complex with 60-kDa delta CaM K II isoform, by sucrose-density sedimentation analysis. Moreover, we show here that putative alphaKAP, in spite of its inability to bind CaM in ligand blot overlay, co-eluted with delta CaM K II from CaM-affinity columns. That raises the question of whether CaM K II-mediated phosphorylation of alpha KAP and triadin together might be involved in a molecular signaling pathway important for SR Ca(2+)-release in fast-twitch muscle SR. PMID- 11112437 TI - Enzymatic formation of unnatural aromatic polyketides by chalcone synthase. AB - Substrate specificity of recombinant chalcone synthase (CHS) from Scutellaria baicalensis (Labiatae) was investigated using chemically synthesized aromatic and aliphatic CoA esters. It was demonstrated for the first time that CHS converted benzoyl-CoA to phlorobenzophenone (2,4,6-trihydroxybenzophenone) along with pyrone by-products. On the other hand, phenylacetyl-CoA was enzymatically converted to an unnatural aromatic polyketide, phlorobenzylketone (2, 4,6 trihydroxyphenylbenzylketone), whose structure was finally confirmed by chemical synthesis. Furthermore, in agreement with earlier reports, S. baicalensis CHS also accepted aliphatic CoA esters, isovaleryl-CoA and isobutyryl-CoA, to produce phloroacylphenones. In contrast, hexanoyl-CoA only afforded pyrone derivatives without formation of a new aromatic ring. It was noteworthy that both aromatic and aliphatic CoA esters were accepted in the active site of the enzyme as a starter substrate for the complex condensation reaction. The low substrate specificity of CHS thus provided further insight into the structure and function of the enzyme. PMID- 11112438 TI - Genomic organization of the human FXYD2 gene encoding the gamma subunit of the Na,K-ATPase. AB - Although the gamma subunit of the Na,K-ATPase has only 66 or 68 amino acids, its human gene (FXYD2) was found to span 9.2 kb and have seven exons, including two alternatively spliced exons encoding different N-termini. Two candidate promoters with consensus sites for transcription factors Sp1, AP-1, and AP-2 are present, consistent with independent transcription of the splice variants. Multiple ESTs support the transcriptional competence of the identified gene elements. In the FXYD2 gene, there are two closely spaced polyadenylation signals, and both are used. A proposed third splice variant encoding a 31-residue N-terminal extension was not found in the gene, nor was the predicted larger protein found in human kidney Na,K-ATPase. Instead, evidence was found for the origin of the larger cDNA clone in homologous recombination with unrelated DNA from chromosome 2. FXYD2 is on chromosome 11q23 close to a site of tumorigenic chromosomal translocations, and has a number of repeat elements. PMID- 11112439 TI - TSH induces insulin receptors that mediate insulin costimulation of growth in normal human thyroid cells. AB - The mitogenic/goitrogenic effects of thyrotropin (TSH) on human thyrocytes in vitro and in vivo depend on permissive comitogenic effects of insulin-like growth factors (IGFs), which are mimicked in vitro by the low-affinity binding of high supraphysiological concentrations of insulin to IGF-I receptors. Contrary to general assumption, we show here that very low concentrations of insulin, acting through insulin receptors but not IGF-I receptors, can also support the stimulation of DNA synthesis by TSH in primary cultures of normal human thyrocytes. Moreover, TSH through cAMP increases the content of insulin receptors demonstrated by Western blotting and the cells' responsiveness to low insulin concentrations. These observations provide the first in vitro evidence in normal human thyroid cells of a functional interaction between TSH and insulin acting through its own receptor. PMID- 11112440 TI - Use of small fluorescent molecules to monitor channel activity. AB - The Mechanosensitive channel of Large conductance (MscL) allows bacteria to rapidly adapt to changing environmental conditions such as osmolarity. The MscL channel opens in response to increases in membrane tension, which allows for the efflux of cytoplasmic constituents. Here we describe the cloning and expression of Salmonella typhimurium MscL (St-MscL). The amino acid sequence encoding for this MscL exhibits a high degree of similarity to Escherichia coli MscL (Eco MscL). Using a fluorescence efflux assay, we demonstrate that efflux through the MscL channel during hypoosmotic shock can be monitored using endogenously produced fluorophores. These fluorophores are synthesized by a cotransformed gene, cobA. In addition, we observe that thermal stimulation, i.e., heat shock, can induce efflux through MscL. PMID- 11112441 TI - Chromatin structure of telomere domain in human sperm. AB - Telomeres in human sperm nucleus are clustered at the nuclear periphery. Chromosomes in the sperm are highly condensed with protamines, however, a small portion of DNA remains associated with histones; the role of the nucleohistone is unknown. To examine structure of the telomeric chromatin, the sperm nuclei were treated with micrococcal nuclease. Chromatin released by the digestion was free from protamines, but contained histones and revealed nucleosomal organization. It was enriched with telomeric DNA organized into closely spaced nucleosomes with a periodicity of 148 +/- bp. Thus, while the most of the sperm genome is packed into extremely dense nucleoprotamine structure, at least a part of the telomeric DNA is arranged into nucleosomes and can be released by the nuclease. We suggest that telomeres might be among the first structures in the sperm nucleus that respond to oocyte signals for male pronucleus development at fertilization. PMID- 11112442 TI - Callipeltin A, a cyclic depsipeptide inhibitor of the cardiac sodium-calcium exchanger and positive inotropic agent. AB - Callipeltin A, a cyclic depsipeptide from the New Caledonian Lithistida sponge Callipelta sp., is a macrocyclic lactone containing four amino acids in the L configuration, Ala, Leu, Thr (2 residues); one (Arg) in the D configuration; two N-methyl amino acids, N-MeAla and N-MeGln; a methoxy tyrosine, a 3, 4-dimethyl-l glutamine; and a 4-amino-7-guanidino-2,3 dihydroxypentanoic acid (AGDHE), formally derived from L-Arg. In cardiac sarcolemmal vesicles Callipeltin A induces a powerful (IC(50) = 0.85 microM) and selective inhibition of the Na(+)/Ca(2+) exchanger. In electrically driven guinea-pig atria, at concentrations ranging between 0.7 and 2.5 microM, Callipeltin A induces a positive inotropic effect, which at the highest concentrations is accompanied by a rise in resting tension. It is suggested that the positive inotropic effect is linked to the inhibition of the Na(+)/Ca(2+) exchanger and that Callipeltin A may be an useful tool to study the role of the cardiac Na(+)/Ca(2+) exchanger in physiological and pathological conditions. PMID- 11112443 TI - Cloning and recombinant expression of human group IIF-secreted phospholipase A(2). AB - Mammalian-secreted phospholipases A(2) (sPLA(2)) form a diverse family of at least nine enzymes that hydrolyze phospholipids to release free fatty acids and lysophospholipids. We report here the cloning and characterization of human group IIF sPLA(2) (hGIIF sPLA(2)). The full-length cDNA codes for a signal peptide of 20 amino acid followed by a mature protein of 148 amino acids containing all of the structural features of catalytically active group II sPLA(2)s. hGIIF sPLA(2) gene is located on chromosome 1 and lies within a sPLA(2) gene cluster of about 300 kbp that also contains the genes for group IIA, IIC, IID, IIE, and V sPLA(2)s. In adult tissues, hGIIF is highly expressed in placenta, testis, thymus, liver, and kidney. Finally, recombinant expression of hGIIF sPLA(2) in Escherichia coli shows that the enzyme is Ca(2+)-dependent, maximally active at pH 7-8, and hydrolyzes phosphatidylglycerol versus phosphatidylcholine with a 15 fold preference. PMID- 11112444 TI - Deficiency of the initiation events of sporulation in Bacillus subtilis clpP mutant can be suppressed by a lack of the Spo0E protein phosphatase. AB - Previous results have shown that the Bacillus subtilis clpP gene is required for developmental processes such as sporulation and competence development. Little is known about its function during the initiation of sporulation. We studied the effect of clpP mutation on the early events of sporulation. The expression of the spo0A and spoIIG genes, whose active transcription requires the phosphorylated Spo0A protein (Spo0A approximately P) as the transcription activator, was significantly decreased in the clpP mutant at the onset of sporulation. The expression of spo0H gene encoding sigma(H) protein was also greatly reduced. As expected from these results, the sigma(H) and Spo0A protein levels in the clpP mutant were also decreased during the initiation of sporulation, indicating that the accumulation of Spo0A approximately P was inhibited in the clpP mutant. We, therefore, introduced the mutation of the spo0E gene, which codes for the Spo0A approximately P-specific phosphatase, into the clpP mutant and found that this double mutant restored the expression of the spo0A as well as spoIIG genes. These results suggest that ClpP had an indirect influence on the intracellular concentration of Spo0A approximately P by regulating the activity of the Spo0E phosphatase during the initiation of sporulation. PMID- 11112445 TI - Glucosamine inhibits inducible nitric oxide synthesis. AB - Glucosamine is widely used in Europe for treatment of arthritis in humans. Based on recent findings that excess production of nitric oxide (NO) by inducible NO synthase (iNOS) mediates the pathogenesis of arthritis, we hypothesized that glucosamine may inhibit NO synthesis. To test this hypothesis, we used an in vivo rat model of lipopolysaccharide (LPS)-induced inflammation. Intravenous administration of d-glucosamine (0.5 mmol/kg) 6 h before, at the time of, and 6 h after intraperitoneal LPS injection (1 mg/kg) decreased urinary excretion of nitrate by 31 and 48%, respectively, at days 1 and 2 post LPS administration. When cultured macrophages were treated with LPS (1 microg/ml) to induce iNOS expression, addition of 0.1, 0.5, 1, and 2 mM d-glucosamine decreased NO production by 18, 38, 60, and 89%, respectively. Glucosamine had no effect on cellular arginine, NADPH or tetrahydrobiopterin concentrations, but dose dependently suppressed iNOS protein expression. Similar decreases in iNOS protein occurred in spleen, lung, and peritoneal macrophages of glucosamine-treated rats. These studies demonstrate that glucosamine is a novel inhibitor of inducible NO synthesis via inhibition of iNOS protein expression, and provide a biochemical basis for the use of glucosamine in treating chronic inflammatory diseases such as arthritis. PMID- 11112446 TI - Mutagenesis studies on the iron binding ligands of clavaminic acid synthase. AB - Mutagenesis studies on conserved histidine residues identified as possible metal binding ligands in clavaminic acid synthase isozyme 2 were consistent with His 145 and His-280 acting as iron ligands, in support of crystallographic and previous mutagenesis studies. Mutagenesis of the four cysteines and a glutamine residue, conserved in both clavaminic acid synthase isozymes 1 and 2, demonstrated that none of these residues is essential for activity. PMID- 11112447 TI - Transcriptional activity of the distal CD40 ligand promoter. AB - CD40 ligand (CD40L, CD154) is a T cell cytokine with highly regulated expression that requires the transcription factor nuclear factor of activated T cells (NF AT) to bind at two sites in the proximal CD40L promoter. We have determined that the distal CD40L promoter (-500 to -1300 bp from start of transcription) conveys superior promoter activity in reporter gene assays. Within the distal promoter, we have identified a third NF-AT binding site, at -761 to -756. Oligonucleotides incorporating each of the three NF-AT sites cross-compete for binding of nuclear extracts from activated T cells and bind NF-ATc2 by antibody supershift. Mutation of the distal NF-AT site reduces activity of the 1300 bp CD40L promoter construct to that of the proximal 500 bp construct, which includes only two NF-AT sites. This suggests that the newly identified NF-AT site is the major mediator of transcriptional activation in the distal CD40L promoter. PMID- 11112448 TI - Characterization of the promoter of the murine mac25 gene. AB - It is important to know the regulation of the expression of the mac25 gene because of its reduced expression in several cancer cells and of its induction by some hormonal factors. We cloned the promoter region of the murine mac25 gene and found five repeats of CCAAT sequences, four Sp1 sites, a TATA-like sequence, and an initiator (INR) sequence. Analysis using luciferase reporter plasmids indicated that CCAAT repeats have a strong enhancer activity and the second to fourth Sp1 sites are essential for basal activity of the expression of the mac25 gene. The 1 kb region that contains the promoter and exon 1 of the mac25 gene was in a typical CpG island. As hypermethylation and reduced expression of the mac25 gene were reported in murine liver tumors, methylation of this CpG island may be directly associated with the expression of the mac25 gene and tumorigenesis. PMID- 11112449 TI - TNF-alpha-dependent activation of NF-kappa B in human osteoblastic HOS-TE85 cells is repressed in vector-averaged gravity using clinostat rotation. AB - Effects of vector-averaged gravity on tumor necrosis factor (TNF)-alpha-dependent activation of nuclear factor kappa B (NF-kappa B) in human osteoblastic HOS-TE85 cells were investigated by culturing the cells using clinostat rotation (clinorotation). Cell cultures were rotated for 72 h at 40 rpm in a clinostat. At the end of clinorotation, the cells were treated with TNF-alpha for 30 min under stationary conditions. Electrophoretic mobility shift assays revealed that TNF alpha-dependent activation of NF-kappa B was markedly reduced in the clinorotated cells when compared with the cells in control stationary cultures or after horizontal rotation (motional controls). The NF-kappa B-dependent transactivation was also impaired in the clinorotated cells, as evidenced by a transient transfection assay with a reporter plasmid containing multimerized NF-kappa B sites. Consistent with these findings, the TNF-alpha-dependent induction of endogenous NF-kappa B-responsive genes p105, I kappa B-alpha, and IL-8, was significantly attenuate in clinorotated cells. These results demonstrate that vector-averaged gravity inhibits the responsiveness of osteoblasts to TNF-alpha by repressing NF-kappa B activation. PMID- 11112450 TI - Oligonucleotide-capped gold nanoparticles for improved atomic force microscopic imaging and enhanced selectivity in polynucleotide detection. AB - A novel assay for selective determination of polynucleotides using atomic force microscopy in conjunction with the formation of the probe/target/DNA-gold nanoparticle sandwich structure at a gold surface is described. A 17-mer probe was attached to the surface for subsequent hybridization with a polynucleotide target. Due to the flat orientation of the probe-target hybrid with respect to the surface and the spatial obstruction of the unhybridized probes near the hybrids, the AFM images are not clear. The hybridization efficiency was estimated to be about 1.1% since certain surface features could not be resolved. The utilization of 30-mer-capped gold nanoparticles not only provides another dimension of selectivity, but also reorients the previously formed probe-target hybrid in such a way that the strands of the target become tethered with respect to the surface. This reorientation improves the resolution in imaging the hybridized target molecules and provides an accurate determination of the hybridization efficiency (16%). PMID- 11112451 TI - Effects of para-nonylphenol on 92 kDa gelatinase secretion by human peripheral lymphocytes and U937 cells in vitro. AB - Much attention has focused on environmental estrogenic chemicals such as para nonylphenol which disrupt various tissues via the estrogen receptor. We studied effects of para-nonylphenol on gelatinase secretion by human lymphocytes in vitro. para-Nonylphenol (0.05-50 microM) dose dependently suppressed 92 kDa gelatinase secretion. The suppressive effect of 25 and 50 microM para-nonylphenol was completely blocked by tamoxifen. We also studied the effects of para nonylphenol (0.05-50 microM) on 92 kDa gelatinase secretion by human leukemia U937 cells. para-Nonylphenol suppressed 92 kDa gelatinase secretion in a dose dependent manner. The suppressive effect of 50 microM para-nonylphenol was completely blocked by tamoxifen. Estradiol did not significantly suppress 92 kDa gelatinase secretion. Our results suggest that para-nonylphenol suppressed 92 kDa gelatinase secretion via the estrogen receptor, however, para-nonylphenol interacts with the estrogen receptor in a manner distinct from estradiol. As this assay system is simple and rapid, it may prove useful to evaluate toxic effects of para-nonylphenol on human blood cells. PMID- 11112452 TI - Intracellular signaling factors--enhanced hepatic nuclear protein binding to TTGGC sequence in the rat regucalcin gene promoter: involvement of protein phosphorylation. AB - The transcriptional mechanism of regucalcin gene expression was determined using gel mobility shift assay with TTGGC oligonucleotide (II-b) which is located between position -523 and -506 in the promoter region, containing a nuclear factor I (NF1) consensus motif TTGGC(N)(6)CC. The mutation analysis in this motif showed that TTGGC sequence was a specific binding region of the nuclear protein in rat liver and the cloned rat hepatoma cells (H4-II-E). When liver nuclei were incubated with ATP (1 mM), the nuclear protein binding to TTGGC sequence was increased. This binding was also increased in the nuclei of H4-II-E cells cultured with 10% FBS. Such an increase was also seen by culture with vanadate (100 microM), a potent inhibitor of protein tyrosine phosphatase. Serum-enhanced nuclear protein binding to TTGGC sequence was decreased in the presence of TFP (10 microM), staurosporine (100 nM), genistein (10 microM), PD98059 (10 microM), or wortmannin (10 nM), which are inhibitors of various protein kinases. Treatment of a monoclonal phosphotyrosine antibody (4G10) caused an alteration in the TTGGC oligonucleotide-nuclear protein complex formation, indicating that tyrosine phosphorylation of nuclear protein is partly involved in the binding to TTGGC sequence. Moreover, when H4-II-E cells were cultured with FBS (10%), Bay K 8644 (5 microM), PMA (1 microM), or insulin (20 nM), the protein binding to TTGGC sequence in the nuclei was increased, while it was reduced in the cytoplasm, indicating a nuclear localization of the TTGGC sequence-binding protein. This study demonstrates that hepatic nuclear protein can specifically bind to the TTGGC sequence in rat regucalcin gene promoter region, and that this binding is enhanced by intracellular signaling factors which are partly mediated through protein phosphorylation. PMID- 11112453 TI - Transcription modulation by a rat nuclear scaffold protein, P130, and a rat highly repetitive DNA component or various types of animal and plant matrix or scaffold attachment regions. AB - The XmnI fragment, a highly repetitive DNA component, and animal and plant matrix or scaffold attachment region (MAR/SAR) were examined for similarity in interaction with nuclear scaffold. As the XmnI fragment bound a 130 kDa scaffold protein (P130) in vitro, various types of MAR/SAR fragments could bind 130 and 123 kDa scaffold proteins. The native XmnI and MAR/SAR fragments clearly augmented SV40 promoter-mediated luciferase gene transcription following transient transfection of recombinant plasmids into various types of recipient cells. In contrast, the XmnI fragment methylated at the cytosine base of the unique HindIII site, and a synthetic variant DNA deficient in base unpairing characteristic of MAR/SAR, could neither bind P130 nor augment this transcription. These two types of genomic regions appeared to have similar properties of interaction with nuclear scaffold, by which the activity of appropriately positioned promoter can be modulated. PMID- 11112454 TI - Discriminating activation of CYP2B9 expression in male C57BL/6 mouse liver by beta-estradiol. AB - The inducible expression of the cytochrome P450 2B subfamily was investigated in male C57BL/6 (B6) and DBA/2 (D2) mice, as well as their hybrids, B6D2F1, at the mRNA level. The expression of hepatic CYP2B mRNAs in B6 was lightly induced by beta-estradiol (ES), while that by phenobarbital (PB) or 1,1,1-trichloro-2, 2 bis(p-chlorophenyl) ethane (DDT) was prominent. Discriminating analysis showed a novelty that ES markedly induced CYP2B9 mRNA expression, whereas PB and DDT increased CYP2B10 more than CYP2B9 expression: albeit both mRNA species responded to all three inducers. Furthermore, the specific induction by ES of CYP2B9 mRNA in B6 male mice, but not D2 male mice, suggests strain dependency in the regulatory pathway of CYP2B9 expression. PMID- 11112455 TI - Erythropoietin inhibits calcium-induced neurotransmitter release from clonal neuronal cells. AB - Erythropoietin (EPO) and EPO receptor (EPO-R) are expressed in the brain but their neuronal functions are not yet clarified. The effects of EPO on neurosecretion were studied using clonal rat pheochromocytoma PC12 cells. EPO suppressed Ca(2+)-induced dopamine release from PC12 cells in a concentration- and time-dependent manner. Inhibition was also produced by an EPO mimetic peptide 1 (EMP1), a small synthetic peptide agonist of EPO-R, but not by its inactive analogue in which Cys residues were substituted with Ser. Inhibition was abolished by genistein but not by genistin. EPO and EMP1 induced autophosphorylation of Janus kinase 2 (JAK 2), a tyrosine kinase that associates with EPO-R, and dephosphorylation of GAP-43 in a tyrosine kinase-dependent fashion. These results suggest that EPO suppresses neurotransmitter release through activation of EPO-R linked to JAK2. PMID- 11112456 TI - The immediate early gene 1 product of human cytomegalovirus is sufficient for up regulation of interleukin-8 gene expression. AB - We previously observed that human cytomegalovirus (CMV) infection induced a massive production of a chemokine with potent neutrophil chemotactic activity, interleukin-8 (IL-8). Hence, we examined the effect of CMV immediate early (IE) gene products on IL-8 production by the human astrocytoma cell line, U373MG. Transient or stable transfection with a CMV IE1 gene expression vector, but not with a IE2 gene expression vector, significantly augmented IL-8 protein secretion and IL-8 mRNA expression. Luciferase activity was enhanced in U373MG cells when the cells were cotransfected with CMV IE1 and chimeric firefly luciferase reporter genes driven by the transcriptional regulatory region of the human IL-8 gene. Moreover, IE1 gene-mediated enhancement of luciferase activity was abolished by the introduction of mutations into the AP-1 or NF-kappa B factor binding elements in the regulatory region of the IL-8 promoter. Furthermore, electrophoretic mobility shift assays demonstrated that CMV IE1 gene products induced the formation of NF-kappa B or AP-1 complexes. Finally, Western blotting analysis demonstrated that the CMV IE1 gene product increased the amount of NF kappa B complexes translocated into the nucleus. Collectively, CMV IE1 gene expression may be sufficient to activate AP-1 and NF-kappa B, resulting in IL-8 gene expression. PMID- 11112457 TI - The current role of rifampicin-impregnated grafts: pragmatism versus science. PMID- 11112458 TI - The prevention and treatment of vascular graft infection with a Triclosan (Irgasan)-bonded Dacron graft: an experimental study in the pig. AB - OBJECTIVES: to evaluate the role of Triclosan (Irgasan(R)) in the prevention of prosthetic graft infection. MATERIAL AND METHODS: fifty-one pigs were assigned randomly to six groups. Group I (graft) and II (graft and Triclosan) were control groups. Groups III (graft) and IV (grafts and Triclosan) were contaminated with 2 x 10(7)CFU/ml S. aureus. Groups V (graft) and VI (graft and Triclosan) were intraoperatively contaminated with 2 x 10(7)CFU/ml S. aureus and reoperated on after 7 days. Remaining animals were sacrificed on day 28. The end point of the investigation was vascular graft infection, defined as the bacteriological and/or histological proof of infection. Results in both control groups no vascular graft infections were detected in Groups I and II. All of the group III animals presented but none of the group IV developed a graft infection (p <0.02). All of the group V animals presented and 10 of 12 animals developed a graft infection. CONCLUSION: in this animal model Triclosan bonding appears effective in preventing prosthetic graft infection. However, the in situ replacement of Triclosan-protected grafts was not successful in the treatment of graft infection. PMID- 11112459 TI - Fucoidan inhibits smooth muscle cell proliferation and reduces mitogen-activated protein kinase activity. AB - OBJECTIVES AND DESIGN: fucoidan has previously been shown to inhibit the proliferation of arterial smooth muscle cells both in animal models and in vitro. However, the mechanisms behind the anti-proliferative effects of this polysulfated polysaccharide are not known in detail. Here, the inhibitory effect of fucoidan on rat aortic smooth muscle cell proliferation was examined and compared with the effects of heparin after stimulation with fetal calf serum, platelet-derived growth factor BB, basic fibroblast growth factor, heparin binding epidermal growth factor, and angiotensin II. MATERIALS AND METHODS: the cultures were analysed with respect to cell proliferation and DNA synthesis by cell counting and measurement of(3)H-thymidine incorporation. Phosphorylation of mitogen-activated protein kinase and nuclear translocation of phosphorylated mitogen-activated protein kinase were studied by immunoblotting and immunocytochemistry. RESULTS: fucoidan was shown to be a more potent inhibitor of smooth muscle cell proliferation than heparin. Fucoidan also reduced growth factor-induced activation of mitogen-activated protein kinase and prevented nuclear translocation of phosphorylated mitogen-activated protein kinase. CONCLUSION: fucoidan is a more potent anti-proliferative polysulphated polysaccharide than heparin and may mediate its effects through inhibition of the mitogen-activated protein kinase pathway in a similar manner as heparin. PMID- 11112460 TI - Determination of wall shear rate in the human carotid artery by magnetic resonance techniques. AB - OBJECTIVES: to measure wall shear rates around the circumference of the human carotid bifurcation throughout the heart cycle. DESIGN: prospective, open study. Materials eight healthy volunteers. METHODS: wall shear rates were determined at the carotid bifurcation using magnetic resonance techniques with high resolution and individually adjusted velocity encoding for imaging and haemodynamic mapping. Wall shear stresses were calculated assuming a constant value of 4 centiPoise. RESULTS: data suitable for postprocessing were obtained in all subjects. The main findings were: unidirectional wall shear rate waveforms and high wall shear rate (775 s(-1)+/-167 s(-1)) at the flow divider; low wall shear rate (60 s(-1+/-40 s( 1)) and a high oscillation index with huge interindividual variation (85+/-65) at the lateral wall. CONCLUSION: these are the first in vivo data describing, in detail, the forces of the blood acting on the wall of the carotid bifurcation. The results do not contradict the hypotheses associating low and oscillating wall shear stress with the development of atherosclerosis.) PMID- 11112461 TI - The significance of the cerebral collateral capacity in patients with carotid atheroma. AB - OBJECTIVES: to identify the echodensity, stenosis of carotid plaques and cerebral collateral capacity that were associated with various ipsilateral presentations (retinal, cerebrovascular, asymptomatic). DESIGN: cross-sectional study. MATERIALS: forty-four patients, with 44 plaques associated with various presentations, were studied. METHODS: the duplex images of the plaques were analysed echomorphologically in a computer by means of Grey Scale Median (GSM) [hypoechoic (low GSM), hyperechoic (high GSM)]. The percentage (%) reduction of the mean velocity in the middle cerebral artery (PRMCA) on transcranial Doppler, during clamping in carotid endarterectomy, was evaluated to distinguish the competent cerebral collateral supply (low PRMCA) from the non-competent one (high PRMCA). RESULTS: the retinal symptoms were associated with plaques of low median GSM (0), severe median stenosis (90%) and low median PRMCA (0.31) as contrasted with the cerebrovascular symptoms (17, 84%, 0.47, respectively) and asymptomatic status (32, 83%, 0.4, respectively) [(p =0.038 (GSM), p =0.67 (stenosis), p=0.15 (PRMCA)]. The retinal and the cerebrovascular symptoms were distinct in terms of PRMCA (p=0.045). CONCLUSIONS: the retinal symptoms were produced by hypoechoic and possibly embologenic plaques, whereas the cerebrovascular ones possibly by the combination of carotid embolism and a non-competent cerebral collateral circulation. Asymptomatic status was associated with the absence of any relevant mechanism. PMID- 11112462 TI - Subintimal angioplasty of tibial vessel occlusions in the treatment of critical limb ischaemia: mid-term results. AB - OBJECTIVES: to evaluate the feasibility and preliminary results at 1 year of subintimal angioplasty of tibial occlusions in critical limb ischaemia (CLI). MATERIAL: from December 1997 to December 1999, we intended to treat 36 patients and 40 limbs by subintimal angioplasty of occlusions of tibial vessels. Thirty one had gangrene or ulceration and nine had rest pain. Twenty-seven occlusions were more than 10 cm, 10 were 5 to 10 cm and three were less than 5 cm in length. Three patients had an occluded previous ipsilateral bypass graft. All patients were followed 3 monthly for a median of 10 months by means of clinical and duplex examination. RESULTS: the technical success rate was 78% (31/40). Nine technical failures were treated by conventional surgery or angioplasty of another diseased tibial vessel. The clinical success rate was 68% (27/40). Four below-the-knee amputations were performed despite a patent recanalisation. Primary and secondary patency rates at 12 months were 56% (72% without technical failures). The 12 month limb salvage rate was 81% and survival rate was 78%. Three of five complications were treated by endovascular procedures. The length of occlusion (>10 cm) but not the location of distal re-entry, the type of vessel re-entry and the presence of diabetes are predictors of technical success and patency. CONCLUSIONS: subintimal angioplasty can be used to treat tibial occlusions in patients with CLI. Technical failure does not preclude conventional surgery and complications may often be treated by endovascular procedures. However, the durability of angioplasty is as yet uncertain. PMID- 11112463 TI - Preoperative angiographic score and intraoperative flow as predictors of the mid term patency of infrapopliteal bypass grafts. AB - OBJECTIVES: preoperative angiographic characteristics of the outflow tract have emerged as a predictive factor for the outcome of infrapopliteal reconstructions. Direct flow measurement can be routinely performed intraoperatively, but little is known regarding its impact on graft outcome. The present study was undertaken to compare the value of these parameters in predicting the mid-term patency of infrapopliteal bypass grafts. DESIGN: retrospective clinical study. PATIENTS: 172 infrapopliteal reconstructions using autogenous vein were performed, of which 92 had a crural and 80 a pedal recipient artery. METHODS: the preoperative angiogram was scored according to the SVS/ISCVS Ad Hoc Committee. At the end of the operation flow was measured with a transit-time flowmeter. Follow-up consisted of pressure measurements and duplex scanning. RESULTS: the runoff score had no impact on femorocrural graft patency. For pedal grafts there was a tendency for inferior outcome with high runoff score, as the 1-year assisted primary patency for grafts with a completely occluded pedal arch was 11% compared with 52% for grafts with lower scores (p=0.056). Both intraoperative volume graft flow and maximum flow capacity had a highly significant influence on the outcome on crural reconstructions on univariate analysis. For pedal reconstructions only a a severely reduced maximum flow capacity after injection of papaverin was associated with an adverse outcome. Multivariate analysis revealed that maximum flow capacity was an independent significant factor affecting patency of femoroinfrapopliteal grafts (relative risk=0.53 per 30 ml/min increase, p<0.001). The runoff score was also a weak independent predictor of 1-year assisted primary patency in these grafts (relative risk=1.9 for a score >4 in crural and a score >5.5 in pedal grafts, p=0.036). CONCLUSIONS: a completely occluded pedal arch in preoperative angiography was associated with poor infrapopliteal bypass outcome. Graft flow and maximal flow capacity are good predictors of the 1-year graft patency of femorocrural bypasses. PMID- 11112464 TI - Reconstruction for renal artery aneurysm and its effect on hypertension. AB - OBJECTIVES: many renal artery aneurysms (RAA) are diagnosed incidentally in the course of investigations for hypertension and their management is controversial. AIM: to review the results of renal artery reconstruction for RAA. METHODS: between January 1978 and December 1998 111 RAR were performed in 81 kidneys in 71 patients. RESULTS: fifty-nine patients were hypertensive, three had a creatinine >2.0 mg/dl and one was on dialysis. The principal underlying pathology was fibromuscular dysplasia (39) and atherosclerosis (17). The mean RAA diameter was 2.2 (range 1-15) cm overall and 3.5 (range 2-10) cm in four patients who presented with rupture. Fifty-one patients had renal artery stenosis. Autogenous material was used in 105 RAR. There was no 30-day mortality and the morbidity rate was 16%. The 5-year cumulative patency rate was 69%. Hypertension was cured in 25% and improved in 39%. CONCLUSIONS: RAR tested for RAA treats hypertension and reduces the risk of rupture and distal embolisation. PMID- 11112465 TI - Activity of matrix metalloproteinase-2 and -9 in abdominal aortic aneurysms. Relation to size and rupture. AB - OBJECTIVES: to investigate the activity of matrix metalloproteinase (MMP)-2 and 9 in asymptomatic abdominal aortic aneurysms (aAAAs) and ruptured abdominal aortic aneurysms (rAAAs). DESIGN: cross-sectional study. MATERIALS AND METHODS: MMP-2 and MMP-9 activity was estimated in biopsies from the anterior wall of 60 AAAs using gelatin zymography. There were 20 medium-sized (diameter 5<7 cm) aAAAs, 20 large (>57 cm) aAAAs and 20 rAAAs. MMP activity was quantified using a laser densitometer and expressed as arbitrary units (au). RESULTS: mean (SEM) MMP 9 activity was significantly lower in large aAAAs (1190 au +/-247) than in rAAAs (2647 au +/-498, p<0.05). There was no difference in MMP-2 activity. CONCLUSION: High MMP-9 activity in the AAA wall is associated with rupture. PMID- 11112466 TI - Abdominal aortic aneurysm and aortic occlusive disease: a comparison of risk factors and inflammatory response. AB - OBJECTIVE: to compare patients with abdominal aortic aneurysm (AAA) and aortic occlusive disease (AOD) with regard to risk factors for atherosclerosis, co morbid conditions and inflammatory activity. PATIENTS AND METHODS: a total of 155 patients undergoing abdominal aortic surgery between January 1993 and October 1997: 82 (53%) had aneurysmal disease and 73 (47%) had occlusive disease. Principal risk factors were compared: age; gender; smoking; hypertension; hyperlipidaemia; diabetes mellitus; severe peripheral vascular disease (PVD) and ischaemic heart disease. Aortic wall tissue samples were obtained during surgery. A prospective blind analysis was performed for the presence of inflammatory cytokines TNF-alpha, IL-1 beta, IL-6 and TGF-beta. RESULTS: the average age of AAA patients was 74 years (50-88), while that of AOD patients was 61 years (43 82) (p<0.0001). Diabetes mellitus was found to be much more prevalent in the AOD group (p<0.001), while hypertension and severe PVD were more prevalent in the AAA group (p<0.001). No differences were found concerning any of the risk factors. Inflammatory cytokine activity: AAA tissue samples contained significantly higher mean TNF-alpha and IL-6 levels compared to the AOD samples (5.6+/-2.7 x 10 E-4 vs. 4.4+/-2.7 x 10 E-5 atmoles/microl (p=0. 01), and 0.6+/-0.4 vs. 0.01+/-0.006 atmoles/microl (p=0.02) respectively). No differences were found related to IL-1 beta and TGF-beta. CONCLUSIONS: (1) Patients with AAA have fewer atherosclerotic risk factors than do patients with AOD. (2) Patients with AAA and AOD have significantly different inflammatory activity. (3) The data supports the hypothesis that AAA and AOD are probably two different pathological entities. PMID- 11112467 TI - Growth of thrombus may be a better predictor of rupture than diameter in patients with abdominal aortic aneurysms. AB - OBJECTIVES: to investigate the relationships between diameter, surface and thrombus area in abdominal aortic aneurysms (AAAs) <5 cm. METHODS AND MATERIAL: sixty-seven patients with AAA underwent at least 2 CT examinations. At the point of maximal diameter, surface area and thrombus area were calculated and related to rupture, or impending rupture, during follow-up. RESULTS: the mean increase in measured diameter, surface area and thrombus area were 3.4 mm, 1.9 cm(2)and 1.7 cm(2)per year respectively. Patients with AAA >4 cm and whose thrombus area increased >1.5 cm(2)/year were more likely to rupture (6/24 vs 1/23). CONCLUSIONS: a rapid increase of thrombus area may be a better predictor of AAA rupture than increase in maximal diameter. PMID- 11112468 TI - Patterns of reflux and severity of varicose veins in the general population- Edinburgh Vein Study. AB - OBJECTIVES: several studies have used duplex ultrasound to assess valvular incompetence in symptomatic patients. This cross-sectional survey was performed to determine, for the first time in a general population sample, the relationship between trunk varices and the presence of reflux in lower limb venous segments. MATERIALS AND METHODS: 1238 subjects (554 male, 684 female), aged 18-64 years, were randomly selected. The presence and severity of any trunk varices was assessed and classified into Grades 1-3; hyphenweb and reticular varices were noted. The presence or absence of reflux > or = 0.5 s in five deep and three superficial venous segments in each leg was assessed using duplex ultrasound. RESULTS: the prevalence of reflux increased in both superficial and deep segments with more severe grades of trunk varices (p< or = 0.01), except the upper SFV in women. Fifteen per cent of females showed reflux confined to the superficial system, compared with 9% of males (p< or = 0.001); 22% of males showed deep reflux, compared with 11% of females (p < or = 0.001). 71% of men and 48% of women with Grade 2-3 trunk varices had mixed deep and superficial reflux. Above knee popliteal segments had a higher prevalence of reflux than lower SFV segments in all groups. There was no consistent relationship between the presence of hyphenweb or reticular varices and deep or superficial reflux. CONCLUSION: segmental venous reflux can be detected in the deep and superficial leg veins of subjects without trunk varices, but superficial and mixed reflux are increasingly found in subjects with more severe varices. PMID- 11112469 TI - Misdiagnosed post-traumatic occlusion of the internal carotid artery in a young woman. PMID- 11112470 TI - Aneurysm sac enlargement after conventional inflammatory aneurysm repair with a polytetrafluoroethylene aortobiiliac graft. PMID- 11112471 TI - Duodenal obstruction following aortic aneurysm repair caused by an aneurysm sac seroma. PMID- 11112472 TI - Arterial haemorrhage from a chronic venous ulcer--pseudoaneurysm formation of the posterior tibial artery. PMID- 11112473 TI - Axilla false aneurysm following late anastomotic disruption of an old axillofemoral bypass graft. PMID- 11112474 TI - Patterns of genome organization and content in lepidopteran baculoviruses. PMID- 11112475 TI - Predominance of canine parvovirus (CPV) in unvaccinated cat populations and emergence of new antigenic types of CPVs in cats. AB - Serological, sequence, and in vitro host range analyses of feline parvovirus (FPV) isolates in Vietnam and Taiwan revealed that more than 80% of the isolates were of the canine parvovirus (CPV) type, rather than feline panleukopenia virus (FPLV). Although parvovirus isolates from three Vietnamese leopard cats were genetically related to CPV type 2a or 2b, they had a natural mutation of VP2 residue 300 Gly to an Asp, resulting in remarkable changes in their antigenic properties. These results indicated the possibility that CPV-2a/2b-type viruses can spread in cats more efficiently than conventional FPLV under natural conditions and that CPV-2a/2b viruses are further evolving in cats. PMID- 11112476 TI - Downregulation of beta1 integrins by Ebola virus glycoprotein: implication for virus entry. AB - Filoviruses, including Ebola virus, are cytotoxic. To investigate the role of the Ebola virus glycoprotein (GP) in this cytopathic effect, we transiently expressed the GP in human kidney 293T cells. Expression of wild-type GP, but not the secretory form of the molecule lacking a membrane anchor, induced rounding and detachment of the cells, as did a chimeric GP containing its ectodomain and influenza virus hemagglutinin transmembrane-cytoplasmic domain. These results indicate that the GP ectodomain and its anchorage to the membrane are required for GP-induced morphologic changes in host cells. Since cell rounding and detachment could be associated with reduced levels of cell adhesion molecules, we also studied the expression of integrins, which are major molecules for adhesion to extracellular matrices, and found that the beta1 integrin group is downregulated by the GP. This result was further extended by experiments in which anti-beta1 monoclonal antibodies or purified integrins inhibited the infectivity of vesicular stomatitis virus pseudotyped with the GP. We suggest that integrins, especially the beta1 group, might interact with the GP and perhaps be involved in Ebola virus entry into cells. PMID- 11112477 TI - Plasmids encoding foot-and-mouth disease virus VP1 epitopes elicited immune responses in mice and swine and protected swine against viral infection. AB - VP1 is a capsid protein of foot-and-mouth disease virus (FMDV) and contains epitopes of the virus. Plasmids encoding two VP1 epitopes (amino acid residues 141-160 and 200-213) and a host-self immunoglobulin molecule were constructed to produce a new type of FMD DNA vaccine. Two plasmids, namely, pCEIM and pCEIS, containing mouse immunoglobulin (IgG) or swine IgG were subjected to immunogenicity testing in mice and swine, respectively. In mice administrated pCEIM in the abdomen using a genegun, both FMDV-specific T-cell proliferation and neutralizing antibodies were detected. In swine immunized with pCEIS at the back of the ear, immune responses were achieved after the second administration. Swine showed a T-cell proliferative response with a stimulation index (SI) of up to 8.1 and a neutralizing antibody response that was able to protect suckling mice from 10(2) LD(50) (lethal dose 50) FMDV challenge. To compare the immunogenicity of the DNA-based vaccine candidate, versus the protein-based vaccine candidates, a second group of swine was immunized with the protein F1-scIgG, which was encoded by the plasmid pCEIS. Injection with F1-scIgG elicited a T-cell proliferative response of SI < 1.7 and a neutralizing antibody response that protected suckling mice from up to 10(5) LD(50) FMDV challenge. In the challenge test, three of three swine immunized with pCEIS were fully protected from FMDV challenge. PMID- 11112478 TI - H9N2 subtype influenza A viruses in poultry in pakistan are closely related to the H9N2 viruses responsible for human infection in Hong Kong. AB - Following the outbreak of H5N1 "bird flu" in Hong Kong in 1997, the isolation of H9N2 subtype viruses from patients in southern China and Hong Kong SAR once again raised the spectre of a possible influenza pandemic. H9N2 viruses have recently been responsible for disease in poultry in various parts of the world and preliminary studies of the H9 haemagglutinin (HA) genes of viruses isolated during 1998 and 1999 in Germany, Iran, Pakistan, and Saudi Arabia showed a close relationship to the HA genes of the viruses that infected two children in Hong Kong SAR. Analysis of the complete genome of a Pakistan isolate, A/chicken/Pakistan/2/99, showed that it is closely related in all eight genes (97 99% homology) to the human H9N2 isolates and furthermore that the six genes encoding internal components of the virus are similar to the corresponding genes of the H5N1 viruses that caused 6 (out of 18) fatal cases of human infection. Thus H9N2 viruses similar to those that caused human infections in Hong Kong are circulating more widely in other parts of the world. Whether or not these H9N2 viruses also have features that facilitate avian-to-human transmission is not known. Since avian H9N2 viruses are currently perceived to represent a significant threat to human health it is important to determine whether or not viruses of this subtype circulating in poultry in various parts of the world have the potential to infect people. PMID- 11112479 TI - The vaccine origin of the 1968 epidemic of type 3 poliomyelitis in Poland. AB - A clear association was demonstrated between the use of USOL-D-bac type 3 poliovirus live-attenuated vaccine and the 1968 poliomyelitis epidemic in Poland. The epidemic followed small-scale trials with Sabin and USOL-D-bac type 3 vaccine strains carried out in seven countries including Poland. Factors that might have contributed to the genesis and development of the epidemic were the pattern of virus excretion from vaccinees, mutations found in viruses from the epidemic, and the particular vaccination policies in Poland during the previous years. These findings may provide essential insights into the strategies for stopping polio immunisation once wild poliovirus has been eradicated. PMID- 11112480 TI - Integrin alpha2beta1 mediates the cell attachment of the rotavirus neuraminidase resistant variant nar3. AB - It was previously reported that integrins alpha2beta1, alpha4beta1, and alphaXbeta2 are involved in rotavirus cell infection. In this work we studied the role of integrin subunits alpha2, alpha4, and beta2 on the attachment of rotaviruses RRV and nar3 to MA104 cells. Integrin alpha2beta1 was found to serve as the binding receptor for the neuraminidase-resistant virus nar3, whereas the neuraminidase-sensitive strain RRV interacted with this integrin at a postattachment step. It was shown that nar3 binds alpha2beta1 through the DGE integrin-recognition motif located in the virus surface protein VP5. Integrin subunits alpha4 and beta2 do not seem to be involved in the initial cell binding of either virus. PMID- 11112481 TI - Influenza virus (A/HK/156/97) hemagglutinin expressed by an alphavirus replicon system protects chickens against lethal infection with Hong Kong-origin H5N1 viruses. AB - Venezuelan equine encephalitis virus replicon particles (VRP) containing the gene expressing hemagglutinin (HA) from the human Hong Kong Influenza A isolate (A/HK/156/97) were evaluated as vaccines in chicken embryos and young chicks. Expressed HA was readily detected in bird-tissue staining with anti-H5 HA antibody and in chicken cells infected with the replicon preparations following immunoprecipitation with monoclonal antibody. Birds challenged with a dose of the lethal parent virus were protected to different extents depending on the age of the bird. In ovo and 1-day-old inoculated animals that received no boost with the VRP were partially protected; birds 2 weeks of age were completely protected with a single dose of VRP. PMID- 11112482 TI - Molecular epidemiology of human herpesvirus 8 in africa: both B and A5 K1 genotypes, as well as the M and P genotypes of K14.1/K15 loci, are frequent and widespread. AB - We have studied 52 new HHV8 strains by sequencing the complete hypervariable K1 gene and genotyping the K14.1/K15 loci located at both sides, respectively, of the viral genome. The samples originated from 49 patients with Kaposi's sarcoma (KS; 32 patients), multicentric Castleman's disease (MCD; 12 patients), or primary effusion lymphoma (PEL; 5 patients). Among these patients, 32 were of African origin (West and Central African countries and Creoles from French Guiana) and the 17 others were mostly French homosexuals. Comprehensive phylogenetic studies allowed the identification of distinct groups within the three already known main subtypes. Interestingly, two new sequences that did not cluster within a known subtype or group could be considered as prototypes of early/ancient variants of the C subtype and A/C set, respectively. Among the 32 African strains, the majority were either of the B subtype (13 cases) or of the A5 group (11 cases), indicating that this latter genotype is frequent and widespread in Africa. In contrast, a subtype C strain infected most of the 17 other patients. PCR-based genotyping of the K14.1/K15 loci revealed an overall predominance of P subtype, except in the A5 and B K1 groups, in which the P and M alleles were equally represented. The implications of these data on the evolution and spread of HHV8 among human African populations are discussed. PMID- 11112483 TI - Identification of a noncanonical signal for transcription of a novel subgenomic mRNA of mouse hepatitis virus: implication for the mechanism of coronavirus RNA transcription. AB - Subgenomic RNA transcription of coronaviruses involves the interaction between the leader (or antileader) and the intergenic (IG) sequences. However, it is not clear how these two sequences interact with each other. In this report, a previously unrecognized minor species of subgenomic mRNA, termed mRNA5-1, was identified in cells infected with mouse hepatitis virus (MHV) strains JHM2c, JHM(2), JHM(3), A59, and MHV-1. Sequence analysis revealed that the leader-body fusion site of the mRNA is located at approximately 150 nucleotides (nt) downstream of the consensus IG sequence for mRNA 5 and did not have sequence homology with any known IG consensus sequences. To determine whether this sequence functions independently as a promoter, we cloned a 140-nt sequence (from approximately 70 nt upstream to approximately 70 nt downstream of the fusion site) from viral genomic RNA and placed it in front of a reporter gene in the defective-interfering (DI) RNA-chloramphenicol acetyltransferase (CAT) reporter vector. Transfection of the reporter RNA into MHV-infected cells resulted in synthesis of a CAT-specific subgenomic mRNA detected by reverse transcription polymerase chain reaction (RT-PCR). The strength of this promoter was similar to that of the IG7 (for mRNA 7) as measured by the CAT activity. Deletion analysis showed that the sequence as few as 13 nt was sufficient to initiate mRNA transcription, while mutations within the 13-nt abolished mRNA transcription. In vitro translation study confirmed that the envelope (E) protein was translated from mRNA5-1, which encodes the open reading frame (ORF) 5b at its 5'-end, indicating that mRNA5-1 is a functional message. Furthermore, when the ORF5b was replaced with the CAT gene and placed in the DI in the context of viral mini genome, CAT was expressed not only from the first ORF of mRNA5-1 but also from the second and third ORF of mRNA5 and genomic DI RNA, respectively, suggesting that more than one mechanism is involved in regulation of ORF5b expression. Our findings thus support the notion that base-pairing between the leader (or antileader) and the IG is not the sole mechanism in subgenomic RNA transcription. PMID- 11112484 TI - Antibody-dependent induction of type I interferons by poliovirus in human mononuclear blood cells requires the type II fcgamma receptor (CD32). AB - The induction of type I interferons (IFNs) in peripheral blood mononuclear cells (PBMCs) can be triggered by viral infection or exposure to viral glycoproteins. Here we show that the IFN-alpha-inducing capacity of attenuated poliovirus vaccine strains is dramatically enhanced in the presence of human polyvalent immunoglobulin G (IgG). The transcription of both IFN-alpha and IFN-beta genes was detected by RT-PCR in stimulated cells. This antibody-dependent activation of type I IFNs genes was also observed with Formalin-inactivated or UV-inactivated poliovirus, but not with empty poliovirus capsids. The ability of poliovirus antibody complexes to induce IFN-alpha was specifically inhibited when PBMCs were preincubated with an excess of the Fc fragment of IgG. Monoclonal antibodies directed to FcgammaRII (CD32) were also inhibitory, whereas antibodies to the two other classes of Fcgamma receptors, CD16 and CD64, were not. Also, aggregation of FcgammaRII by anti-CD32 antibodies alone failed to induce IFN-alpha production. Our results suggest that induction of type I interferons by poliovirus-antibody complexes depends on CD32-mediated phagocytosis of RNA-containing viral particles. As suggested by the results of an ELISPOT analysis, only a fraction of the IFN-alpha-producing cells are able to synthesize IFN-alpha in response to poliovirus-IgG complexes. PMID- 11112485 TI - Inhibition of murine AIDS (MAIDS) development in C57BL/6J mice by tyrphostin AG 1387. AB - We previously showed that certain tyrphostin derivatives, known as protein tyrosine kinase inhibitors, also act as topoisomerase I-specific antagonists and inhibit Moloney murine leukemia virus replication in vitro in acutely and chronically infected cells. However, an accurate portrayal of retroviral-induced disease cannot rely exclusively on extrapolations from in vitro data. Therefore, experiments with animal models are essential for evaluating the efficacy of a specific drug in vivo. In this study, we examined the effect of tyrphostin AG 1387 on murine AIDS (MAIDS) development in C57BL/6J mice injected with the LP-BM5 virus mixture. A single dose of tyrphostin, administered together with or 24 h post virus inoculation, decreased the development of MAIDS symptoms as measured by spleen and lymph node weight, the T-cell response to concanavalin A (con A), and spleen architecture. Furthermore, weekly treatment with tyrphostins totally abolished MAIDS symptoms and prevented the viral infection of the spleen cells as measured by the absence of viral RNA and the restoration of T-cell function in these spleens. These results implicate that prolonged treatment with tyrphostins is needed for the prevention of MAIDS development in infected mice and suggest that it may be applied as a legitimate remedy for the treatment of retroviral induced diseases. PMID- 11112486 TI - Construction and biological characterization of infectious molecular clones of HIV-1 subtypes B and E (CRF01_AE) generated by the polymerase chain reaction. AB - We previously described the use of extended polymerase chain reaction (PCR) to amplify contiguous 9.2-kilobase (kb) single-long terminal repeat (LTR) proviral sequences from HIV-1 genetic subtypes A through G. We now extend these findings by describing a novel vector system to recover infectious molecular clones from long PCR amplicons. Directional ligation of 9.2-kb proviral amplicons into a recovery vector reconstitutes missing LTR sequences, providing candidate molecular clones for infectivity screening. We show that a previously characterized infectious molecular clone of HIV-1 retains its biological properties upon recovery with this strategy. Three additional infectious molecular clones generated, from primary isolates of subtype B (HIV-1(WR27)) and circulating recombinant form 01_AE (subtype E) (HIV-1(CM235)) by subtype-specific LTR reconstitution, displayed biological properties reflecting their cognate parental isolates. This represents the first report of infectious molecular clones from circulating recombinant form 01_AE (subtype E). PMID- 11112487 TI - Ubiquitination of HIV-1 and MuLV Gag. AB - Our previous biochemical studies of HIV-1 and MuLV virions isolated and identified mature Gag products, HIV-1 p6(Gag) and MuLV p12(Gag), that were conjugated to a single ubiquitin. To study the importance of the monoubiquitination of Gag, a series of lysine to arginine mutants were constructed that eliminated ubiquitination at one or both of the lysines in HIV 1(NL4-3) p6(Gag) and both lysines in Moloney MuLV p12(Gag). HPLC and immunoblot analysis of the HIV-1 mutants demonstrated that either of the lysines in p6(Gag), K27 or K33, could be monoubiquitinated. However, infectivity assays showed that monoubiquitination of HIV-1 p6(Gag) or MuLV p12(Gag) is not required for viral replication in vitro. Pulse-chase radiolabeling of HIV-1-producing cells revealed that monoubiquitination of p6(Gag) does not affect the short-term release of virus from the cell, the maturation of Pr55(Gag), or the sensitivity of these processes to proteasome inhibitors. Experiments with protease-deficient HIV-1 showed that Pr55(Gag) can be monoubiquitinated, suggesting that p6(Gag) is first modified as a domain within Gag. Examination of the proteins inside an HIV-1 mutant found that free ubiquitin was incorporated into the virions in the absence of the lysines in p6(Gag), showing that the ubiquitin inside the virus is not initially brought in as a p6(Gag) conjugate. Although our results establish that monoubiquitination of p6(Gag) and p12(Gag) is not required for viral replication in vitro, this modification may be a by-product of interactions between Gag and cellular proteins during assembly and budding. PMID- 11112488 TI - Involvement of minor structural proteins in recombination of polyoma virus DNA. AB - We have previously observed that a polyoma-mouse chimeric DNA molecule (RmI) in which the murine DNA insert is flanked by directly repeated viral sequences is effectively converted into unit-length polyoma DNA upon transfection of permissive mouse cells. This intramolecular recombination event appears to be dependent on VmP1, a protein encoded by RmI which includes the 328 N-terminal amino acids of polyoma VP1, and nine amino acids of murine origin carrying the C terminus of the protein. We report here that introducing mutations into the VP2/VP3 coding sequence reduces the ability of RmI to generate polyoma DNA, even though the same mutations seem to exert little or no effect on the ability of polyoma DNA to either replicate or accumulate inside transfected cells. A mutation affecting VP2 alone being as effective as one that affects both VP2 and VP3, VP2 appears to be playing a critical role in recombination. PMID- 11112489 TI - In vivo retroviral integration: fidelity to size of the host DNA duplication might Be reduced when integration occurs near sequences homologous to LTR ends. AB - Integrated retroviral DNAs are flanked by a short duplication of target DNA whose size is virus specific and invariable. We have sequenced the junctions between an ALSV (Avian Leukemia and Sarcoma Viruses)-based vector and quail DNA from five individual proviruses. Three proviruses were flanked by the expected 6-bp duplication of host DNA, whereas the two others were flanked by a 5-bp duplication. Nucleotide sequencing of the native integration sites of these two proviruses showed that these integrations had occurred at the immediate vicinity of either a CA or a TG dinucleotide, revealing striking microhomologies between the integration sites and viral LTR ends. These results suggest that size duplication of the target DNA might be influenced by nucleotidic sequence at the site of integration. PMID- 11112490 TI - Biological properties of herpes simplex virus 2 replication-defective mutant strains in a murine nasal infection model. AB - We used a mouse nasal model of herpes simplex virus 2 (HSV-2) infection to examine the biological properties of HSV-2 wild-type (wt), TK-negative, and replication-defective strains in vivo. Nasal septa tissue is the major site of wt viral replication post intranasal (i.n.) inoculation. The HSV-2 strain 186 syn(+) 1 wt virus caused lethal encephalitis at doses of 10(4) PFU and above per nostril, and at lower doses no neurons in the trigeminal ganglia were positive for the latency-associated transcript, indicating a lack of latent infection. The 186DeltaKpn TK-negative mutant virus replicated in nasal septa tissue but showed low-level replication in trigeminal ganglia at only one timepoint. In situ hybridization of trigeminal ganglia showed that the number of LAT-positive neurons was proportional to the inoculum dose from 10(3) to 10(6) PFU per nare. The replication-defective mutant virus 5BlacZ showed no replication in nasal septa tissue and no persistence of viral DNA at the inoculation site or the trigeminal ganglia. Nevertheless, inoculation of 5BlacZ or the double-mutant dl5 29 at distal sites reduced acute replication and latent infection of 186DeltaKpn following intranasal challenge. This infection model provides a biological system to test the properties of HSV-2 strains and shows that replication-defective mutant strains do not persist at sites of inoculation or in sensory ganglia but can induce immune protection that reduces the latent viral load of a challenge virus. PMID- 11112491 TI - Regulation of MVM NS1 by protein kinase C: impact of mutagenesis at consensus phosphorylation sites on replicative functions and cytopathic effects. AB - Minute virus of mice NS1, an 83-kDa mainly nuclear phosphoprotein, is the only viral nonstructural protein required in all cell types and it is involved in multiple processes necessary for virus propagation. The diversity of functions assigned to NS1, together with the variation of its complex phosphorylation pattern during infection, suggested that the various activities of NS1 could be regulated by distinct phosphorylation events. So far, it has been demonstrated that NS1 replicative functions, in particular, DNA-unwinding activities, are regulated by protein kinase C (PKC), as exemplified by the modulation of NS1 helicase activity by PKClambda phosphorylation. In order to determine further impact of phosphorylation on NS1 functions, including the induction of cytopathic effects, a mutational approach was pursued in order to produce NS1 variants harboring amino acid substitutions at candidate PKC target residues. Besides the determination of two additional in vivo phosphorylation sites in NS1, this mutagenesis allowed the segregation of distinct NS1 functions from one another, generating NS1 variants with a distinct activity profile. Thus, we obtained NS1 mutants that were fully proficient for trans activation of the viral P38 promoter, while being impaired in their replicative functions. Moreover, the alterations of specific PKC phosphorylation sites gave rise to NS1 polypeptides that exerted reduced cytotoxicity, leading to sustained gene expression, while keeping functions necessary for progeny virus production, i.e., viral DNA replication and activation of the capsid gene promoter. These data suggested that in the course of a viral infection, NS1 may undergo a shift from productive to cytotoxic functions as a result of a phosphorylation-dependent regulation. PMID- 11112492 TI - Recovery of a virulent strain of newcastle disease virus from cloned cDNA: expression of a foreign gene results in growth retardation and attenuation. AB - A recombinant mesogenic NDV strain, Beaudette C, and an engineered recombinant NDV expressing an additional gene were generated entirely from cloned cDNAs. For this purpose, a full-length cDNA clone of the virus genome, represented in eight different subgenomic fragments, was assembled in a transcription plasmid between a T7 RNA polymerase promoter and a hepatitis delta virus ribozyme sequence. Infectious NDV could be generated in the cells infected with recombinant vaccinia virus, which expressed T7 RNA polymerase, by simultaneous expression of antigenome-sense NDV RNA from the full-length plasmid and NDV NP, P, and L proteins from cotransfected plasmids. Recombinant virus was then amplified and recovered, either after inoculation of transfection supernatant into the allantoic cavity of embryonated specific-pathogen-free eggs or after further passage in cell culture. Characterization of the recombinant NDV showed similarities in growth and pathogenicity to that of the parental wild-type virus. By using this system, a recombinant NDV containing a foreign gene encoding chloramphenicol acetyltransferase (CAT) was generated. To do this, the CAT transcription cassette containing the CAT open reading frame, flanked by NDV gene start and gene end sequence motifs, was inserted into the region between the HN and L genes of the full-length cDNA. This construct was then used in the generation of a recombinant NDV expressing CAT protein. The CAT gene was maintained stably for at least eight passages without any detectable loss of the gene from the recombinant. Generation of the recombinant virus, however, was associated with reduced plaque size, slower replication kinetics, and more than 100-fold decrease in yield. In addition, the virus showed an increase in mean death time for eggs and a lower intracerebral pathogenicity index in day-old chicks, implicating attenuation of the recombinant virus. Thus, introduction of an additional gene into the NDV genome represents a method to achieve growth retardation and attenuation. These results also indicate that NDV can be engineered to express foreign protein stably and can be manipulated in the future for use as a vaccine vector. PMID- 11112493 TI - Simian foamy virus infections in a baboon breeding colony. AB - The prevalence, transmission, and variation of simian foamy viruses (SFVs) in baboons was investigated. Over 95% of adult baboons in the breeding colony as well as recently imported adult animals had high titers of anti-SFV serum IgG. Maternal antibody was detectable in infants' serum up to 6 months of age. Approximately 30% of infants in breeding harems experienced SFV infections by 1 year of age. Shedding of SFV in oral secretions was common, with 13% of samples from normal adult animals and 35% from immunosuppressed animals containing infectious SFV. SFV was isolated from three baboon subspecies (olive, yellow, and chacma baboons) and sequences from both the pol and the LTR regions of the provirus were amplified by PCR and sequenced. Phylogenetic analysis indicated that all baboon isolates formed a single lineage distinct from SFVs of other African monkey species. Within the baboon SFV lineage, two distinct clades were apparent, which consisted of isolates from yellow and olive baboons and isolates from chacma baboons. Competition ELISAs indicated that, while SFV isolates of these two groups were very closely related, antigenic differences do exist between them. SFV isolates from a drill and a mandrill were distinct from baboon SFV isolates, both genetically and antigenically. PMID- 11112494 TI - Induction of long-term protective effects against heterologous challenge in SIVhu infected macaques. AB - A group of three rhesus macaques were inoculated with SIV isolated from a human (SIVhu) accidentally exposed and infected with SIVsm. Extensive sequence analyses of SIVhu obtained from the human and macaques following infection indicated the presence of truncated nef. Not only did nef fail to repair itself in vivo postinfection (p.i.), but instead, further mutations added additional stop codons with increasing time p.i. Infection of these animals was associated with minimal acute viral replication, followed by undetectable plasma viral loads and only intermittent PCR detection up to 5 years p.i. The three SIVhu infected and three control monkeys were then challenged with the heterologous highly pathogenic SHIV89.6p. All three controls became infected and showed rapid declines in peripheral CD4(+) lymphocytes, disease, and death at 10 and 32 weeks p.i., respectively. In contrast, all three animals previously infected with SIVhu are healthy and exhibit stable CD4(+) lymphocyte levels and undetectable plasma viral loads at >20 months post-SHIV89. 6p challenge. Only transient, low levels of SHIV replication were noted in these animals. Whereas responses to SIVgag/pol were noted, no evidence for SIV/SHIV envelope cross-reactivity was detected by antibody or CTL analyses, suggesting that the protective immune mechanisms to the heterologous challenge isolate were most likely not directed to envelope but rather to other viral determinants. PMID- 11112495 TI - Persistent elevated expression of cytokine transcripts in ganglia latently infected with herpes simplex virus in the absence of ganglionic replication or reactivation. AB - Infection of mouse trigeminal ganglia by herpes simplex virus induces cytokine expression that persists long after infectious virus or viral antigens become undetectable. To examine mechanisms underlying this phenomenon, we used a thymidine kinase mutant, dlsptk, which fails to replicate in ganglia and does not reactivate upon ganglionic explant. Using quantitative reverse transcriptase polymerase chain reaction assays, we found that levels of interferon-gamma and tumor necrosis factor-alpha transcripts in dlsptk-infected ganglia were lower than those in wild type-infected ganglia, but were significantly (eight- to 10 fold) higher than those in mock-infected ganglia from Day 3 to Day 100 postinfection. We also studied latency-associated transcript (LAT) negative mutants that exhibit increased expression of productive cycle transcripts in ganglia. Ganglia infected with these mutants contained levels of cytokine transcripts similar to those in wild type-infected ganglia; any increases in viral antigen expression mediated by the LAT deletion were not accompanied by increased cytokine expression. Thus, neither viral replication, the ability to reactivate, nor LAT expression in ganglia is required for persistent elevated cytokine expression. The results provide indirect evidence that low-level expression of viral productive cycle genes in neurons can provide signals that elicit cytokine expression. PMID- 11112496 TI - Second-site mutations encoding residues 34 and 78 of the major capsid protein (VP5) of herpes simplex virus type 1 are important for overcoming a blocked maturation cleavage site of the capsid scaffold proteins. AB - During assembly of the herpes simplex type 1 capsid, the major capsid protein VP5 interacts with the C-terminal residues of the scaffold proteins encoded by UL26 and UL26.5. Subsequent to capsid assembly the scaffold proteins are cleaved at the maturation site by a serine protease also encoded by UL26, thereby enabling the bulk of the scaffold proteins to be released from the capsid. Previously, a mutant virus (KUL26-610/611) was isolated in which this maturation cleavage site was blocked by replacing the Ala/Ser at the 610/611 cleavage site by Glu/Phe. This mutation was lethal and required a transformed cell line expressing wild type UL26 gene products for growth. Although the mutation was lethal, spontaneous reversions occurred at a high frequency. Previously, a small number of revertants were isolated and all were found to have second-site mutations in VP5. The purpose of the present study was to do a comprehensive determination of the sites altered in VP5 by the second-site mutations. To do this, an additional 25 independent spontaneous revertants were characterized. Seven of the 25 arose by GC --> GT changes in codon 78, giving rise to an alanine to valine substitution. Four were the result of base changes at codon 34 but two different amino acids were produced as the changes were at different positions in the codon. Two mutations were detected at position 41 and mutations that occurred once were found at codons 69 and 80. Thus, 15 of the 25 second-site mutants were localized to codons 34 to 80 of VP5, which contains 1374 amino acids. The remaining 10 revertants had codon changes at nine different sites, of which the most N terminal was altered at codon 187 and the most C-terminal at codon 1317. As noted in the much smaller study a preponderance of the second-site mutants in VP5 were altered in codons at the extreme N-terminus of VP5. It is especially noteworthy that 11 out of 25 of the mutations occurred at codons 34 and 78. As expected, all of the revertants isolated were shown to retain the original KUL26-610/611 mutation, and the scaffold proteins remain uncleaved. All showed decreased retention of VP24 in the B capsids compared to the wild-type KOS, but more than the KUL26-610/611 parental virus. The revertants all had decreased growth rates of 2 to 18% compared to that of KOS and showed varying degrees of sensitivity when grown at 39.5 degrees C. The mutations in VP5 of three of the previously isolated viruses (PR5, PR6, and PR7) were transferred into a wild-type background, i.e., a virus encoding wild-type UL26 and UL26.5 gene products. All replicated in nonpermissive (Vero) cells and cleaved scaffold proteins. PR5 and PR6 in the wild-type background gave wild-type burst sizes and gave C-capsids that retained VP24 at approximately wild-type levels. The third revertant, PR7, in the wild-type background showed only a twofold increase of burst size (to 20% of wild-type) and the capsids showed little or no increase of VP24 retention. Therefore, the second-site mutations of PR7 (R69C) by itself had a negative effect on virus replication. By contrast the temperature sensitivity of PR6 and PR7 remained unchanged in the wild-type background. Thus the temperature sensitivity of PR6 and PR7 resides in VP5 independently of the mutation in the UL26 cleavage site. PMID- 11112497 TI - Establishment of latent HIV-1 infection of resting CD4(+) T lymphocytes does not require inactivation of Vpr. AB - The introduction of highly active antiretroviral therapy (HAART) for the treatment of HIV-1 infection has allowed dramatic reductions in plasma virus levels to below the limit of detection in many patients. However, latently infected CD4(+) memory T lymphocytes persist as an important reservoir for the virus in the presence of this aggressive therapy and represent a major barrier to HIV-1 eradication with HAART. The mechanism through which the latent compartment is formed has not yet been established. It may involve actively proliferating CD4(+) T-cell intermediates that are infected with HIV-1 and revert back to a resting state, carrying integrated provirus at some low frequency. The HIV-1 accessory protein Vpr, which mediates G(2) cell cycle arrest in host cells, may interfere with the formation of the latently infected T cells by preventing them from exiting the cell cycle to return to a resting state. To investigate the role of the Vpr in the formation of latently infected memory T cells, we cloned and characterized vpr genes from viruses in the latent reservoir. Both sequence analysis and functional assays demonstrated that the vpr gene products of the viruses isolated from the latent pool did not differ significantly from those of a functional Vpr (NL4-3). These results indicate that the generation of resting G(0) memory T lymphocytes that carry latent HIV-1 provirus occurs despite the G(2) arrest function of the vpr gene product. PMID- 11112498 TI - Recombinant bovine adenovirus type 3 expressing bovine viral diarrhea virus glycoprotein E2 induces an immune response in cotton rats. AB - Recombinant bovine adenovirus is being developed as a live vector for animal vaccination and for human gene therapy. In this study, two replication-competent bovine adenovirus 3 (BAV-3) recombinants (BAV331 and BAV338) expressing bovine viral diarrhea virus (BVDV) glycoprotein E2 in the early region 3 (E3) of BAV-3 were constructed. Recombinant BAV331 contains chemically synthesized E2 gene (nucleotides modified to remove internal cryptic splice sites) under the control of BAV-3 E3/major late promoter (MLP), while recombinant BAV338 contains original E2 gene under the control of human cytomegalovirus immediate early promoter. Since E2, a class I membrane glycoprotein, does not contain its own signal peptide sequence at the 5' end, the bovine herpesvirus 1 (BHV-1) glycoprotein D signal sequence was fused in frame to the E2 open reading frame (ORF) for proper processing of the E2 glycoprotein in both the recombinant viruses. Recombinant E2 protein expressed by BAV331 and BAV338 recombinant viruses was recognized by E2 specific monoclonal antibodies as a 53-kDa protein, which also formed dimer with an apparent molecular weight of 94 kDa. Insertion of an E2-expression cassette in the E3 region did not effect the replication of recombinant BAV-3s. Intranasal immunization of cotton rats with these recombinant viruses generated E2-specific IgA and IgG responses at the mucosal surfaces and in the serum. In summary, these results show that the pestivirus glycoprotein can be expressed efficiently by BAV 3. In addition, mucosal immunization with replication-competent recombinant bovine adenovirus 3 can induce a specific immune response against the expressed antigen. PMID- 11112499 TI - Vaccinia virus E10R protein is associated with the membranes of intracellular mature virions and has a role in morphogenesis. AB - This study provides the initial biochemical, microscopic, and genetic characterization of the product of the vaccinia virus E10R gene, which belongs to the ERV1/ALR family of eukaryotic proteins, is conserved in all poxviruses and has homologs in other cytoplasmic DNA viruses. DNA encoding a short epitope tag was appended to the C-terminus of the 95-amino-acid open-reading frame without affecting virus reproduction. The E10R protein was synthesized after DNA replication and was associated with purified intracellular mature virions (IMV), from which it could be extracted with a nonionic detergent. Antibody to the tag decorated the surface of IMV, consistent with the anchorage of the E10R protein to the membrane via its hydrophobic N-terminus. Immunoelectron microscopy revealed that the E10R protein was associated with crescent membranes, immature virions, IMV, and extracellular particles. To investigate the role of E10R in the virus life cycle, we constructed an inducer-dependent null mutant. In the absence of inducer, the formation of infectious virus was severely inhibited and electron microscopy revealed an assembly block with accumulation of crescent membranes and immature virions. Cysteines 43 and 46, comprising a putative redox motif common to all poxvirus E10R homologs, were essential for complementation of the mutant virus by transfected E10R DNA. PMID- 11112500 TI - Closterovirus encoded HSP70 homolog and p61 in addition to both coat proteins function in efficient virion assembly. AB - Assembly of the viral genome into virions is a critical process of the virus life cycle often defining the ability of the virus to move within the plant and to be transmitted horizontally to other plants. Closteroviridae virions are polar helical rods assembled primarily by a major coat protein, but with a related minor coat protein at one end. The Closteroviridae is the only virus family that encodes a protein with similarity to cellular chaperones, a 70-kDa heat-shock protein homolog (HSP70h). We examined the involvement of gene products of Citrus tristeza virus (CTV) in virion formation and found that the chaperone-like protein plus the p61 and both coat proteins were required for efficient virion assembly. Competency of virion assembly of different CTV mutants was assayed by their ability to be serially passaged in Nicotiana benthamiana protoplasts using crude sap as inoculum, and complete and partial virus particles were analyzed by serologically specific electron microscopy. Deletion mutagenesis revealed that p33, p6, p18, p13, p20, and p23 genes were not needed for virion formation. However, deletion of either minor- or major-coat protein resulted in formation of short particles which failed to be serially transferred in protoplasts, suggesting that both coat proteins are required for efficient virion assembly. Deletion or mutation of HSP70h and/or p61 dramatically reduced passage and formation of full-length virions. Frameshift mutations suggested that the HSP70h and p61 proteins, not the RNA sequences, were needed for virion assembly. Substitution of the key amino acid residues in the ATPase domain of HSP70h, Asp(7) to Lys or Glu(180) to Arg, reduced assembly, suggesting that the chaperone like ATPase activity is involved in assembly. Both HSP70h and p61 proteins appeared to contribute equally to assembly, consistent with coordinate functions of these proteins in closterovirus virion formation. The requirement of two accessory proteins in addition to both coat proteins for efficient assembly is uniquely complex for helical virions. PMID- 11112501 TI - Co-replication of a reovirus and a polydnavirus in the ichneumonid parasitoid Hyposoter exiguae. AB - A recently established colony of the ichneumonid parasitoid, Hyposoter exiguae, was found to carry both a reovirus (HeRV) and a polydnavirus (HePDV). Morphogenesis of these viruses was observed in all cells comprising the ovarian calyx epithelium, apparently without detrimental effect to the parasitoid. While polydnavirus replication in H. exiguae was restricted to the calyx region, HeRV was detected in ovarioles, oviducts, midguts, malpighian tubules, and accessory glands associated with the male reproductive system. In addition, HeRV was able to infect the fat body of parasitized host larvae and to establish a persistent infection in vitro. Electron microscopy revealed that both viruses were released into the calyx fluid compartment exclusively by budding, a phenomenon rarely observed among the Reoviridae; HeRV envelopes thus obtained, however, appeared to be subsequently shed within the oviducts. HeRV particles were concentrated to near homogeneity by differential centrifugation; mature virions consisted of seven to eight structural polypeptides and 10 dsRNA genome segments. Prominent spikes were observed at the vertices of icosahedral core particles. Most, but not all, individuals comprising the H. exiguae colony appeared to be infected with HeRV, suggesting a commensal relationship between wasp and virus; however, while this association is of obvious benefit to the virus, it seems unlikely that any advantage accrues to the parasitoid which carries it. PMID- 11112502 TI - CD4-Dependent and CD4-independent utilization of coreceptors by human immunodeficiency viruses type 2 and simian immunodeficiency viruses. AB - More than 10 G protein-coupled receptors (GPCRs) have been reported to act as coreceptors for entry of human and simian immunodeficiency viruses (HIV and SIV). We investigated the utilization of six GPCRs as coreceptors by T-cell-line adapted HIV-2 strains (CBL-20, CBL-21, CBL-23, GH-1, ROD, and SBL6669) and SIV strains (SIVagmTYO-1, SIVmac251, and SIVmndGB-1). NP-2/CD4 cells were transduced with CCR3, CCR5, CCR8, CXCR4, GPR1, or APJ, and examined for susceptibilities to cell-free HIV/SIV. HIV-2 strains were grouped into two types by their coreceptor usage. The first group, CBL-20 and CBL-21, used CXCR4 exclusively; the other four strains used a few or all of the six coreceptors. These strains could further infect CD4-negative NP-2/CXCR4 or NP-2/CCR5 cells in the presence (all strains) or absence (SBL6669 and ROD strains) of soluble CD4. SIVagm and SIVmnd infected NP-2/CD4/GPR1 cells. The coreceptors CCR3, CCR8, GPR1, and APJ did not mediate the CD4-independent infection. Although HIV-2ROD and SIVmnd infected both NP 2/CD4/CXCR4 and NP-2/CD4/CCR5 cells, only CXCR4 and CCR5, respectively, were used in CD4-independent infection. Binding of virions to CD4-negative cells occurred at 4 degrees C. These findings suggest that there may be a correlation between the promiscuous use of coreceptors by HIV-2/SIV strains and their ability to infect CD4-negative cells. PMID- 11112503 TI - Assembly and release of human immunodeficiency virus type 1 Gag proteins containing tandem repeats of the matrix protein coding sequences in the matrix domain. AB - We have constructed human immunodeficiency virus (HIV) gag mutants by increasing the matrix protein (MA) sequences via tandemly repeated duplication of the central 107-MA codons. Instead of a total of 132 amino acid residues for the wild type MA, the resultant mutants designated as MA2, MA3, and MA4 contained a total of 242, 352, and 462 codons in the MA domains, respectively. Analysis indicated that the addition of 110 or 220 amino acid residues to the MA did not significantly affect the assembly, release, and processing of particles; however, particle production was markedly reduced when another copy of 110 residues was added to the MA. Subcellular fractionation analysis suggested that the MA tandem repeat mutations enhanced the Gag membrane affinity, in a manner which correlated with the copy number of MA sequences. The effects of enhanced membrane affinity were substantially reduced when sequences downstream of the capsid (CA) domain were deleted. Sucrose density gradient fractionation analysis showed that particles produced by the large insertion mutants possessed wild-type (wt) HIV particle density. Truncation of sequences downstream of the nucleocapsid (NC) domains of the mutants did not influence the budding of particles. In contrast, particle budding was severely impaired when sequences downstream of the CA domain were truncated. Particle densities for the large Gag proteins, which were truncated at the C-terminus of CA, were about 1.12-1.14 g/ml lower than that for wt. Our results suggest that the HIV MA domain could adopt insertions of large protein sequences, and strongly support the proposal that the NC and p2 domains play a crucial role in the process of correct Gag protein packing. PMID- 11112504 TI - The 2000 Olympic Games of protein structure prediction; fully automated programs are being evaluated vis-a-vis human teams in the protein structure prediction experiment CAFASP2. AB - In this commentary, we describe two new protein structure prediction experiments being run in parallel with the CASP experiment, which together may be regarded as the 2000 Olympic Games of structure prediction. The first new experiment is CAFASP, the Critical Assessment of Fully Automated Structure Prediction. In CAFASP, the participants are fully automated programs or Internet servers, and here the automated results of the programs are evaluated, without any human intervention. The second new experiment, named LiveBench, follows the CAFASP ideology in that it is aimed towards the evaluation of automatic servers only, while it runs on a large set of prediction targets and in a continuous fashion. Researchers will be watching the 2000 protein structure prediction Olympic Games, to be held in December, in order to learn about the advances in the classical 'human-plus-machine' CASP category, the fully automated CAFASP category, and the comparison between the two. PMID- 11112505 TI - Mapping protein sequence spaces by recurrence quantification analysis: a case study on chimeric structures. AB - Recurrence quantification analysis (RQA) was used to characterize the folding properties of 22 chimeric sequences derived from two parent proteins of similar length but different three-dimensional arrangement. A non-linear relation between sequence data and their RQA representation was revealed, which points to new information carried by this method as compared with classical best-alignment methods. This new information is significantly correlated with the folding properties of the hybrid polypeptide chains, as substantiated by careful statistical analysis of the recurrence plots' numerical descriptors, thus encouraging their systematic use to complement sequence data in both proteomics and protein engineering tasks. Even the direct visual screening of the qualitative graphical features of recurrence plots is shown to provide useful hints to discriminate between different recurrence structures of protein sequences. PMID- 11112506 TI - Disulfide recognition in an optimized threading potential. AB - An energy potential is constructed and trained to succeed in fold recognition for the general population of proteins as well as an important class which has previously been problematic: small, disulfide-bearing proteins. The potential is modeled on solvation, with the energy a function of side chain burial and the number of disulfide bonds. An accurate disulfide recognition algorithm identifies cysteine pairs which have the appropriate orientation to form a disulfide bridge. The potential has 22 energy parameters which are optimized so the Protein Data Bank (PDB) structure for each sequence in a training set is the lowest in energy out of thousands of alternative structures. One parameter per amino acid type reflects burial preference and a single parameter is used in an overpacking term. Additionally, one optimized parameter provides a favorable contribution for each disulfide identified in a given protein structure. With little training, the potential is >80% accurate in ungapped threading tests using a variety of proteins. The same level of accuracy is observed in a threading test of small proteins which have disulfide bonds. Importantly, the energy potential is also successful with proteins having uncrosslinked cysteines. PMID- 11112507 TI - Relationship between local structure and stability in hen egg white lysozyme mutant with alanine substituted for glycine. AB - We prepared five mutant lysozymes in which glycines whose dihedral angles are located in the region of the left-handed helix, Gly49, Gly67, Gly71, Gly102 and Gly117, were mutated to an alanine residue. From analyses of their thermal stabilities using differential scanning calorimetry, most of them were more destabilized than the native lysozyme, except for the G102A mutant, which has a stability similar to that of the native lysozyme at pH 2.7. As for the destabilized mutant lysozymes, their X-ray crystallographic analyses showed that their global structures did not change but that the local structures changed slightly. By examining the dihedral angles at the mutation sites based on X-ray crystallographic results, it was found that the dihedral angles at these mutation sites tended to adopt favorable values in a Ramachandran plot and that the extent and direction of their shifts from the original value had similar tendencies. Therefore, the change in dihedral angles may be the cause of the slight local structural changes around the mutation site. On the other hand, regarding the mutation of G102A, the global structure was almost identical with that of the native structure but the local structure was drastically changed. Therefore, it was suggested that the drastic local conformational change might be effective in releasing the unfavorable interaction of the native state at the mutation site. PMID- 11112508 TI - Structural and thermodynamic consequences of introducing alpha-aminoisobutyric acid in the S peptide of ribonuclease S. AB - The S protein-S peptide interaction is a model system to study binding thermodynamics in proteins. We substituted alanine at position 4 in S peptide by alpha-aminoisobutyric acid (Aib) to investigate the effect of this substitution on the conformation of free S peptide and on its binding to S protein. The thermodynamic consequences of this replacement were studied using isothermal titration calorimetry. The structures of the free and complexed peptides were studied using circular dichroic spectroscopy and X-ray crystallography, respectively. The alanine4Aib replacement stabilizes the free S peptide helix and does not perturb the tertiary structure of RNase S. Surprisingly, and in contrast to the wild-type S peptide, the DeltaG degrees of binding of peptide to S pro, over the temperature range 5-30 degrees C, is virtually independent of temperature. At 25 degrees C, the DeltaDeltaG degrees, DeltaDeltaH degrees, DeltaDeltaS and DeltaDeltaCp of binding are 0.7 kcal/mol, 2.8 kcal/mol, 6 kcal/mol x K and -60 kcal/mol x K, respectively. The positive value of DeltaDeltaS is probably due to a decrease in the entropy of uncomplexed alanine4Aib relative to the wild-type peptide. The positive value of DeltaDeltaH: degrees is unexpected and is probably due to favorable interactions formed in uncomplexed alanine4Aib. This study addresses the thermodynamic and structural consequences of a replacement of alanine by Aib both in the unfolded and complexed states in proteins. PMID- 11112509 TI - Charge engineering of a protein domain to allow efficient ion-exchange recovery. AB - We have created protein domains with extreme surface charge. These mutated domains allow for ion-exchange chromatography under conditions favourable for selective and efficient capture, using Escherichia coli as a host organism. The staphylococcal protein A-derived domain Z (Zwt) was used as a scaffold when constructing two mutants, Zbasic1 and Zbasic2, with high positive surface charge. Far-ultraviolet circular dichroism measurements showed that they have a secondary structure content comparable to the parental molecule Zwt. Although melting temperatures (Tm) of the engineered domains were lower than that of the wild-type Z domain, both mutants could be produced successfully as intracellular full length products in E. coli and purified to homogeneity by ion-exchange chromatography. Further studies performed on Zbasic1 and Zbasic2 showed that they were able to bind to a cation exchanger even at pH values in the 9 to 11 range. A gene fusion between Zbasic2 and the acidic human serum albumin binding domain (ABD), derived from streptococcal protein G, was also constructed. The gene product Zbasic2-ABD could be purified using cation-exchange chromatography from a whole cell lysate to more than 90% purity. PMID- 11112510 TI - Analysis of the psychrotolerant property of hormone-sensitive lipase through site directed mutagenesis. AB - Mammalian hormone-sensitive lipase (HSL) has given its name to a family of primarily prokaryotic proteins which are structurally related to type B carboxylesterases. In many of these alpha/beta hydrolases, a conserved HG dipeptide flanks the catalytic pocket. In HSL this dipeptide is followed by two additional glycine residues. Through site-directed mutagenesis, we have investigated the importance of this motif for enzyme activity. Since the presence of multiple glycine residues in a critical region could contribute to cold adaptation by providing local flexibility, we studied the effect of mutating these residues on the psychrotolerant property of HSL. Any double mutation rendered the enzyme completely inactive, without any major effect on the enzyme stability. The partially active single mutants retained the same proportion of activity at reduced temperatures as the wild-type enzyme. These results do not support a role for the HGGG motif in catalysis at low temperatures, but provide further validation of the current three-dimensional model of HSL. Rat HSL was found to be relatively more active than human HSL at low temperatures. This difference was, however, not due to the 12 amino acids which are present in the regulatory module of the rat enzyme but absent in human HSL. PMID- 11112511 TI - Control of antibody-antigen interaction using anion-induced conformational change in antigen peptide. AB - The binding of a monoclonal antibody to an epitope peptide was controlled by the conformational change of the epitope peptide induced by anions. We synthesized peptides in which the epitope sequence DTYRYI for the monoclonal antibody AU1 is located between amphiphilic peptides (KKLL)n and (LLKK)n. In the absence of an appropriate anion, the peptide was in a random coil state and the epitope was linear. In contrast, in the presence of an appropriate anion, the peptide exhibited an anti-parallel alpha-helical structure and the epitope was subsequently 'bent'. In the presence of 41 microM triphosphate, the association constant between the antibody and the peptide was decreased by one order of magnitude in the case of n = 3 and at least three orders of magnitude in the case of n = 4 or 5. Oligo-DNAs, as anions, dissociated the antibody-peptide complex, whereas triphosphate did not. The DNA concentrations required for 50% dissociation of the antibody-peptide complex at pH 7.5 were 4x10(-8), 1x10(-7) and 6x10(-6) M for decamer, octamer and hexamer DNA, respectively. PMID- 11112512 TI - Expression of a bispecific dsFv-dsFv' antibody fragment in Escherichia coli. AB - A bispecific disulfide-stabilized Fv antibody fragment (dsFv-dsFv') consisting of two different disulfide-stabilized Fv antibody fragments connected by flexible linker peptides was produced by secretion of three polypeptide chains into the periplasm of Escherichia coli. The dsFv-dsFv' molecules were enriched by immobilized metal affinity chromatography and further purified by anion-exchange chromatography. The recombinant antibody constructs retained the two parental antigen binding specificities and were able to cross-link the two different antigens. The described dsFv-dsFv' design might be of particular value for therapeutic in vivo applications since improved stability is expected to be combined with minimal immunogenicity. PMID- 11112513 TI - Minor structural consequences of alternative CUG codon usage (Ser for Leu) in Candida albicans exoglucanase. AB - In some species of Candida the CUG codon is encoded as serine and not leucine. In the case of the exo-beta-1,3-glucanase from the pathogenic fungus C. albicans there are two such translational events, one in the prepro-leader sequence and the other at residue 64. Overexpression of active mature enzyme in a yeast host indicated that these two positions are tolerant to substitution. By comparing the crystal structure of the recombinant protein with that of the native (presented here), it is seen how either serine or leucine can be accommodated at position 64. Examination of the relatively few solved protein structures from C. albicans indicates that other CUG encoded serines are also found at non-essential surface sites. However such codon usage is rare in C. albicans, in contrast to C. rugosa, with direct implications for respective recombinant protein production. PMID- 11112514 TI - Protein conformation and disease: pathological consequences of analogous mutations in homologous proteins. AB - The antibody light chain variable domain (V(L))(1) and myelin protein zero (MPZ) are representatives of the functionally diverse immunoglobulin superfamily. The V(L) is a subunit of the antigen-binding component of antibodies, while MPZ is the major membrane-linked constituent of the myelin sheaths that coat peripheral nerves. Despite limited amino acid sequence homology, the conformations of the core structures of the two proteins are largely superimposable. Amino acid variations in V(L) account for various conformational disease outcomes, including amyloidosis. However, the specific amino acid changes in V(L) that are responsible for disease have been obscured by multiple concurrent primary structure alterations. Recently, certain demyelination disorders have been linked to point mutations and single amino acid polymorphisms in MPZ. We demonstrate here that some pathogenic variations in MPZ correspond to changes suspected of determining amyloidosis in V(L). This unanticipated observation suggests that studies of the biophysical origin of conformational disease in one member of a superfamily of homologous proteins may have implications throughout the superfamily. In some cases, findings may account for overt disease; in other cases, due to the natural repertoire of inherited polymorphisms, variations in a representative protein may predict subclinical impairment of homologous proteins. PMID- 11112515 TI - Interaction of a mitochondrial presequence with lipid membranes: role of helix formation for membrane binding and perturbation. AB - The binding of a peptide to a biological membrane is often accompanied by a transition from a random coil structure to an amphipathic alpha-helix. Recently, we have presented a new approach which allows the determination of the thermodynamic parameters of membrane-induced helix formation [Wieprecht et al. (1999) J. Mol Biol. 294, 785]. It involves a systematic variation of the helix content of a given peptide by double D-substitution and a correlation of the binding parameters with the helicity. Here we have used this method to study membrane-induced helix formation for the presequence of rat mitochondrial rhodanese (RHD). The thermodynamic parameters of binding of the peptide RHD and of four of its double D-isomers were determined for 30 nm (SUVs) and 100 nm (LUVs) unilamellar vesicles composed of phosphatidylcholine/phosphatidylglycerol (3:1) using circular dichroism spectroscopy, fluorescence spectroscopy, and isothermal titration calorimetry. The incremental changes of the thermodynamic parameters of helix formation were found to be very similar for SUVs and LUVs. Membrane-induced helix formation of RHD entailed a negative enthalpy of Delta H(helix) = -0.5 to -0.6 kcal/mol/residue and was opposed by an entropy of about Delta S(helix) = -1 to -1.4 cal/mol K/residue. The free energy of helix formation, Delta G(helix), was about -0.2 kcal/mol, and helix formation accounted for 50-60% of the total free energy of membrane binding. Dye-release experiments were used to assess the role of helix formation for the membrane perturbation potential of the peptides. While helix formation plays a major role for membrane binding, it appears to have little importance for inducing membrane leakiness. PMID- 11112516 TI - Proximity relationships between residue 6 of troponin I and residues in troponin C: further evidence for extended conformation of troponin C in the troponin complex. AB - Skeletal muscle troponin C (TnC) adopts an extended conformation when crystallized alone and a compact one when crystallized with an N-terminal troponin I (TnI) peptide, TnI(1-47) [Vassylyev et al. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 4847-4852]. The N-terminal region of TnI (residues 1-40) was suggested to play a functional role of facilitating the movement of TnI's inhibitory region between TnC and actin [Tripet et al. (1997) J. Mol. Biol. 271, 728-750]. To test this hypothesis and to investigate the conformation of TnC in the intact troponin complex and in solution, we attached fluorescence and photo cross-linking probes to a mutant TnI with a single cysteine at residue 6. Distances from this residue to residues of TnC were measured by the fluorescence resonance energy transfer technique, and the sites of photo-cross-linking in TnC were determined by microsequencing and mass spectrometry following enzymatic digestions. Our results show that in the troponin complex neither the distance between TnI residue 6 and TnC residue 89 nor the photo-cross-linking site in TnC, Ser133, changes with Ca(2+), in support of the notion that this region plays mainly a structural rather than a regulatory role. The distances to residues 12 and 41 in TnC's N-domain are both considerably longer than those predicted by the crystal structure of TnC.TnI(1-47), supporting an extended rather than a compact conformation of TnC. In the binary TnC.TnI complex and the presence of Ca(2+), Met43 in TnC's N-domain was identified as the photo-cross-linking site, and multiple distances between TnI residue 6 and TnC residue 41 were detected. This was taken to indicate increased flexibility in TnC's central helix and that TnC dynamically changes between a compact and an extended conformation when troponin T (TnT) is absent. Our results further emphasize the difference between the binary TnC.TnI and the ternary troponin complexes and the importance of using intact proteins in the study of structure-function relationships of troponin. PMID- 11112517 TI - Farnesyl diphosphate synthase. Altering the catalytic site to select for geranyl diphosphate activity. AB - Farnesyl diphosphate synthase (FPPase) catalyzes chain elongation of the C(5) substrate dimethylallyl diphosphate (DMAPP) to the C(15) product farnesyl diphosphate (FPP) by addition of two molecules of isopentenyl diphosphate (IPP). The synthesis of FPP proceeds in two steps, where the C(10) product of the first addition, geranyl diphosphate (GPP), is the substrate for the second addition. The product selectivity of avian FPPase was altered to favor synthesis of GPP by site-directed mutagenesis of residues that form the binding pocket for the hydrocarbon residue of the allylic substrate. Amino acid substitutions that reduced the size of the binding pocket were identified by molecular modeling. FPPase mutants containing seven promising modifications were constructed. Initial screens using DMAPP and GPP as substrates indicated that two of the substitutions, A116W and N144'W, strongly discriminated against binding of GPP to the allylic site. These observations were confirmed by an analysis of the products from reactions with DMAPP in the presence of excess IPP and by comparing the steady-state kinetic constants for the wild-type enzyme and the A116W and N114W mutants. PMID- 11112518 TI - Conformational energetics of a reverse turn in the Clostridium beijerinckii flavodoxin is directly coupled to the modulation of its oxidation-reduction potentials. AB - A surface loop in the flavodoxin from Clostridium beijerinckii comprised of residues -Met(56)-Gly-Asp-Glu(59)- forms a four-residue reverse turn which undergoes a conversion from a mix of cis/trans peptide configurations that approximate a type II configuration in the oxidized state to a type II' turn upon reduction of the bound flavin mononucleotide (FMN) cofactor. This change results in the formation of a new hydrogen bond between the N(5)H of the reduced cofactor and the carbonyl group of Gly57 of the central peptide bond of the turn, an interaction that is thought to contribute to the modulation of the oxidation reduction potentials of the cofactor [Ludwig, M. L., Pattridge, K. A., Metzger, A. L., Dixon, M. M., Eren, M., Feng, Y., and Swenson, R. P. (1997) Biochemistry 36, 1259-1280]. In this study, the direct linkage of the conformational energetics of this turn to the stabilization of the FMN semiquinone was established by systematically replacing the second and third residues of the turn (Gly57 and Asp58) with the -Gly-Gly-, -Gly-Ala-, -Ala-Gly-, and -Ala-Ala- dipeptidyl sequences. On the basis of published position specific preferences for residues with side chains (mimicked by Ala) and glycine, a strong correlation was observed between E(ox/sq) and the calculated free-energy differences between the type II and type II' conformations of each of these sequence combinations. The Ala-Gly- sequence, which favors the type II turn configuration primarily adopted in the oxidized state, displays a E(ox/sq) value that is about 150 mV more negative than that for the wild-type-like -Gly-Ala- sequence, which prefers the type II' conformation observed in the reduced states. The -Gly-Gly- and -Ala-Ala- mutants exhibit intermediate E(ox/sq) values consistent with their ambivalent turn preferences. The potential changes are primarily the result of alterations in the stability of the semiquinone state. These results provide more conclusive evidence for the crucial role of this conformational change in the modulation of the redox potentials of this flavodoxin. Furthermore, this study establishes a direct association between the conformational energetics of the protein, induced in this case by the sequence specificity of a beta-turn, and the differential thermodynamic stabilization of specific redox states of the cofactor, demonstrating another means by which flavoproteins can modulate the redox potentials of the bound cofactor. PMID- 11112519 TI - Determinants of allosteric activation of yeast pyruvate kinase and identification of novel effectors using computational screening. AB - We have analyzed the structural determinants of the allosteric activation of yeast pyruvate kinase (YPK) by mutational and kinetic analysis and initiated a structure-based design project to identify novel effectors that modulate its allosteric response by binding to the allosteric site for fructose-1,6 bisphosphate (FBP). The wild-type enzyme is strongly activated by fructose-1,6 bisphosphate and weakly activated by both fructose-1-phosphate and fructose-6 phosphate; the strength of the activation response is proportional to the affinity of the allosteric effector. A point mutation within the 6'-phosphate binding loop of the allosteric site (T403E) abolishes activation of the enzyme by fructose-1, 6-bisphosphate. The mutant enzyme is also not activated by F1P or F6P. The mutation alone (which incorporates a glutamic acid that is strictly conserved in mammalian M1 isozymes) slightly reduces cooperativity of substrate binding. Three novel compounds were identified that effect the allosteric regulation of YPK by FBP and/or act as novel allosteric activators of the enzyme. One is a physiologically important diphospho sugar, while the other two are hydrophobic compounds that are dissimilar to the natural effector. These results demonstrate that novel allosteric effectors may be identified using structure based screening and are indicative of the potential of this strategy for drug discovery. Regulatory sites are generally more divergent than catalytic sites and therefore offer excellent opportunities for discrimination and specificity between different organisms or between different tissue types. PMID- 11112520 TI - Rhodamine 123 binds to multiple sites in the multidrug resistance protein (MRP1). AB - The mechanisms of MRP1-drug binding and transport are not clear. In this study, we have characterized the interaction between MRP1 and rhodamine 123 (Rh123) using the photoreactive-iodinated analogue, [(125)I]iodoaryl azido-rhodamine 123 (or IAARh123). Photoaffinity labeling of plasma membranes from HeLa cells transfected with MRP1 cDNA (HeLa-MRP1) with IAARh123 shows the photolabeling of a 190 kDa polypeptide not labeled in HeLa cells transfected with the vector alone. Immunoprecipitation of a 190 kDa photolabeled protein with MRP1-sepcific monoclonal antibodies (QCRL-1, MRPr1, and MRPm6) confirmed the identity of this protein as MRP1. Analysis of MRP1-IAARh123 interactions showed that photolabeling of membranes from HeLa-MRP1 with increasing concentrations of IAARh123 was saturable, and was inhibited with excess of IAARh123. Furthermore, the photoaffinity labeling of MRP1 with IAARh123 was greatly reduced in the presence of excess Leukotreine C(4) or MK571, but to a lesser extent with excess doxorubicin, colchicine or chloroquine. Cell growth assays showed 5-fold and 14 fold increase in the IC(50) of HeLa-MRP1 to Rh123 and the Etoposide VP16 relative to HeLa cells, respectively. Analysis of Rh123 fluorescence in HeLa and HeLa-MRP1 cells with or without ATP suggests that cross-resistance to Rh123 is in part due to reduced drug accumulation in the cytosol of HeLa-MRP1 cells. Mild digestion of purified IAARh123-photolabeled MRP1 with trypsin showed two large polypeptides (approximately 111 and approximately 85 kDa) resulting from cleavage in the linker domain (L1) connecting the multiple-spanning domains MSD0 and MSD1 to MSD2. Exhaustive proteolysis of purified IAARh123-labeled 85 and 111 kDa polypeptides revealed one (6 kDa) and two (approximately 6 plus 4 kDa) photolabeled peptides, respectively. Resolution of total tryptic digest of IAARh123-labeled MRP1 by HPLC showed three radiolabeled peaks consistent with the three Staphylococcus aureus V8 cleaved peptides from the Cleveland maps. Together, the results of this study show direct binding of IAARh123 to three sites that localize to the N- and C-domains of MRP1. Moreover, IAARh123 provides a sensitive and specific probe to study MRP1-drug interactions. PMID- 11112521 TI - Interface sliding as illustrated by the multiple quaternary structures of liganded hemoglobin. AB - Initial crystallographic studies suggested that fully liganded mammalian hemoglobin can adopt only a single quaternary structure, the quaternary R structure. However, more recent crystallographic studies revealed the existence of a second quaternary structure for liganded hemoglobin, the quaternary R2 structure. Since these quaternary structures can be crystallized, both must be energetically accessible structures that coexist in solution. Unanswered questions include (i) the relative abundance of the R and R2 structures under various solution conditions and (ii) whether other quaternary structures are energetically accessible for the liganded alpha(2)beta(2) hemoglobin tetramer. Although crystallographic methods cannot directly answer the first question, they represent the most direct and most accurate approach to answering the second question. We now have determined and refined three different crystal structures of bovine carbonmonoxyhemoglobin. These structures provide clear evidence that the dimer-dimer interface of liganded hemoglobin has a wide range of energetically accessible structures that are related to each other by a simple sliding motion. The dimer-dimer interface acts as a "molecular slide bearing" that allows the two alpha beta dimers to slide back and forth without greatly altering the number or the nature of the intersubunit contacts. Since the general stereochemical features of this interface are not unusual, it is likely that interface sliding of the kind displayed by fully liganded hemoglobin plays important structural and functional roles in many other protein assemblies. PMID- 11112522 TI - Structure of the bacteriophage lambda Ser/Thr protein phosphatase with sulfate ion bound in two coordination modes. AB - The protein phosphatase encoded by bacteriophage lambda (lambda PP) belongs to a family of Ser/Thr phosphatases (Ser/Thr PPases) that includes the eukaryotic protein phosphatases 1 (PP1), 2A (PP2A), and 2B (calcineurin). These Ser/Thr PPases and the related purple acid phosphatases (PAPs) contain a conserved phosphoesterase sequence motif that binds a dinuclear metal center. The mechanisms of phosphoester hydrolysis by these enzymes are beginning to be unraveled. To utilize lambda PP more effectively as a model for probing the catalytic mechanism of the Ser/Thr PPases, we have determined its crystal structure to 2.15 A resolution. The overall fold resembles that of PP1 and calcineurin, including a conserved beta alpha beta alpha beta structure that comprises the phosphoesterase motif. Substrates and inhibitors probably bind in a narrow surface groove that houses the active site dinuclear Mn(II) center. The arrangement of metal ligands is similar to that in PP1, calcineurin, and PAP, and a bound sulfate ion is present in two novel coordination modes. In two of the three molecules in the crystallographic asymmetric unit, sulfate is coordinated to Mn2 in a monodentate, terminal fashion, and the two Mn(II) ions are bridged by a solvent molecule. Two additional solvent molecules are coordinated to Mn1. In the third molecule, the sulfate ion is triply coordinated to the metal center with one oxygen coordinated to both Mn(II) ions, one oxygen coordinated to Mn1, and one oxygen coordinated to Mn2. The sulfate in this coordination mode displaces the bridging ligand and one of the terminal solvent ligands. In both sulfate coordination modes, the sulfate ion is stabilized by hydrogen bonding interactions with conserved arginine residues, Arg 53 and Arg 162. The two different active site structures provide models for intermediates in phosphoester hydrolysis and suggest specific mechanistic roles for conserved residues. PMID- 11112523 TI - Estimation of the hydrophobic effect in an antigen-antibody protein-protein interface. AB - Antigen-antibody complexes provide useful models for analyzing the thermodynamics of protein-protein association reactions. We have employed site-directed mutagenesis, X-ray crystallography, and isothermal titration calorimetry to investigate the role of hydrophobic interactions in stabilizing the complex between the Fv fragment of the anti-hen egg white lysozyme (HEL) antibody D1.3 and HEL. Crystal structures of six FvD1.3-HEL mutant complexes in which an interface tryptophan residue (V(L)W92) has been replaced by residues with smaller side chains (alanine, serine, valine, aspartate, histidine, and phenylalanine) were determined to resolutions between 1.75 and 2.00 A. In the wild-type complex, V(L)W92 occupies a large hydrophobic pocket on the surface of HEL and constitutes an energetic "hot spot" for antigen binding. The losses in apolar buried surface area in the mutant complexes, relative to wild-type, range from 25 (V(L)F92) to 115 A(2) (V(L)A92), with no significant shifts in the positions of protein atoms at the mutation site for any of the complexes except V(L)A92, where there is a peptide flip. The affinities of the mutant Fv fragments for HEL are 10-100-fold lower than that of the original antibody. Formation of all six mutant complexes is marked by a decrease in binding enthalpy that exceeds the decrease in binding free energy, such that the loss in enthalpy is partly offset by a compensating gain in entropy. No correlation was observed between decreases in apolar, polar, or aggregate (sum of the apolar and polar) buried surface area in the V(L)92 mutant series and changes in the enthalpy of formation. Conversely, there exist linear correlations between losses of apolar buried surface and decreases in binding free energy (R(2) = 0.937) as well as increases in the solvent portion of the entropy of binding (R(2) = 0.909). The correlation between binding free energy and apolar buried surface area corresponds to 21 cal mol(-1) A(-2) (1 cal = 4.185 J) for the effective hydrophobicity at the V(L)92 mutation site. Furthermore, the slope of the line defined by the correlation between changes in binding free energy and solvent entropy approaches unity, demonstrating that the exclusion of solvent from the binding interface is the predominant energetic factor in the formation of this protein complex. Our estimate of the hydrophobic contribution to binding at site V(L)92 in the D1.3-HEL interface is consistent with values for the hydrophobic effect derived from classical hydrocarbon solubility models. We also show how residue V(L)W92 can contribute significantly less to stabilization when buried in a more polar pocket, illustrating the dependence of the hydrophobic effect on local environment at different sites in a protein-protein interface. PMID- 11112524 TI - Orientation of alpha-neurotoxin at the subunit interfaces of the nicotinic acetylcholine receptor. AB - The alpha-neurotoxins are three-fingered peptide toxins that bind selectively at interfaces formed by the alpha subunit and its associating subunit partner, gamma, delta, or epsilon of the nicotinic acetylcholine receptor. Because the alpha-neurotoxin from Naja mossambica mossambica I shows an unusual selectivity for the alpha gamma and alpha delta over the alpha epsilon subunit interface, residue replacement and mutant cycle analysis of paired residues enabled us to identify the determinants in the gamma and delta sequences governing alpha-toxin recognition. To complement this approach, we have similarly analyzed residues on the alpha subunit face of the binding site dictating specificity for alpha-toxin. Analysis of the alpha gamma interface shows unique pairwise interactions between the charged residues on the alpha-toxin and three regions on the alpha subunit located around residue Asp(99), between residues Trp(149) and Val(153), and between residues Trp(187) and Asp(200). Substitutions of cationic residues at positions between Trp(149) and Val(153) markedly reduce the rate of alpha-toxin binding, and these cationic residues appear to be determinants in preventing alpha-toxin binding to alpha 2, alpha 3, and alpha 4 subunit containing receptors. Replacement of selected residues in the alpha-toxin shows that Ser(8) on loop I and Arg(33) and Arg(36) on the face of loop II, in apposition to loop I, are critical to the alpha-toxin for association with the alpha subunit. Pairwise mutant cycle analysis has enabled us to position residues on the concave face of the three alpha-toxin loops with respect to alpha and gamma subunit residues in the alpha-toxin binding site. Binding of NmmI alpha-toxin to the alpha gamma interface appears to have dominant electrostatic interactions not seen at the alpha delta interface. PMID- 11112525 TI - High-level expression and mutagenesis of recombinant human phosphatidylcholine transfer protein using a synthetic gene: evidence for a C-terminal membrane binding domain. AB - Phosphatidylcholine transfer protein (PC-TP) is a 214-amino acid cytosolic protein that promotes intermembrane transfer of phosphatidylcholines, but no other phospholipid class. To probe mechanisms for membrane interactions and phosphatidylcholine binding, we expressed recombinant human PC-TP in Escherichia coli using a synthetic gene. Optimization of codon usage for bacterial protein translation increased expression of PC-TP from trace levels to >10% of the E. coli cytosolic protein mass. On the basis of secondary structure predictions of an amphipathic alpha-helix (residues 198-212) in proximity to a hydrophobic alpha helix (residues 184-193), we explored whether the C-terminus might interact with membranes and promote binding of phosphatidylcholines. Consistent with this possibility, truncation of five residues from the C-terminus shortened the predicted amphipathic alpha-helix and decreased PC-TP activity by 50%, whereas removal of 10 residues eliminated the alpha-helix, abolished activity, and markedly decreased the level of membrane binding. Circular dichroic spectra of synthetic peptides containing one ((196-214)PC-TP) or both ((183-214)PC-TP) predicted C-terminal alpha-helices in aqueous buffer were most consistent with random coil structures. However, both peptides adopted alpha-helical configurations in the presence of trifluoroethanol or phosphatidylcholine/phosphatidylserine small unilamellar vesicles. The helical content of (196-214)PC-TP increased in proportion to vesicle phosphatidylserine content, consistent with stabilization of the alpha-helix at the membrane surface. In contrast, the helical content of (183-214)PC-TP was not influenced by vesicle composition, implying that the more hydrophobic of the alpha-helices penetrated into the membrane bilayer. These studies suggest that tandem alpha helices located near the C-terminus of PC-TP facilitate membrane binding and extraction of phosphatidylcholines. PMID- 11112526 TI - Identification of tyrosine 204 as the photo-cross-linking site in the DNA-EcoRI DNA methyltransferase complex by electrospray ionization mass spectrometry. AB - We describe a highly sensitive strategy combining laser-induced photo-cross linking and HPLC-based electrospray ionization mass spectrometry to identify amino acid residues involved in protein-DNA recognition. The photoactivatible cross-linking thymine isostere, 5-iodoracil, was incorporated at a single site within the sequence recognized by EcoRI DNA methyltransferase (GAATTC). UV irradiation of the DNA-protein complex at 313 nm results in a >60% cross-linking yield. SDS-polyacrylamide gel electrophoresis and mass spectrometry were used to analyze the covalent cross-linked complex. The total mass is consistent with covalent bond formation between one strand of DNA and the protein with 1:1 stoichiometry. Protease digestion of the cross-linked complex yields several peptide-DNA adducts that were purified by anion-exchange column chromatography. A combination of mass spectrometric analysis and amino acid sequencing revealed that tyrosine 204 was cross-linked to the DNA. Electrospray mass spectrometric analysis of the peptide-nucleoside adduct confirmed this assignment. Tyrosine 204 resides in a peptide motif previously thought to be involved in AdoMet binding and methyl transfer. Thus, amino acids within loop segments but outside of "DNA binding" motifs can be critical to DNA recognition. Our method provides an accurate characterization of picomole quantities of DNA-protein complexes. PMID- 11112527 TI - The substrate activation process in the catalytic reaction of Escherichia coli aromatic amino acid aminotransferase. AB - Aromatic amino acid aminotransferase is active toward both aromatic and dicarboxylic amino acids, and the mechanism for this dual substrate recognition has been an issue in the enzymology of this enzyme. Here we show that, in the reactions with aromatic and dicarboxylic ligands, the pK(a) of the Schiff base formed between the coenzyme pyridoxal 5'-phosphate and Lys258 or the substrate increases successively from 6.6 in the unliganded enzyme to approximately 8.8 in the Michaelis complex and to >10.5 in the external Schiff base complex. Mutations of Arg292 and Arg386 to Leu, which mimic neutralization of the positive charges of the two arginine residues by the ligand carboxylate groups, increased the Schiff base pK(a) by 0.1 and 0.7 unit, respectively. In contrast to these moderate effects of the Arg mutations, the cleavage of the Lys258 side chain of the Schiff base, which was brought about by preparing a mutant enzyme in which Lys258 was changed to Ala and the Schiff base was reconstituted with methylamine, produced the Schiff base pK(a) value of 10.2, that being 3.6 units higher than that of the wild-type enzyme. The observation indicates that the Schiff base pK(a) in the enzyme is lowered by the torsion around the C4-C4' axis of the Schiff base and suggests that the pK(a) is mainly controlled by changing the torsion angle during the course of catalysis. This mechanism, first observed for the reaction of aspartate aminotransferase with aspartate [Hayashi, H., Mizuguchi, H., and Kagamiyama, H. (1998) Biochemistry 37, 15076-15085], does not require the electrostatic contribution from the omega-carboxylate group of the substrate, and can explain why in aromatic amino acid aminotransferase the aromatic substrates can increase the Schiff base pK(a) during catalysis to the same extent as the dicarboxylic substrates. This is the first example in which the torsion pK(a) coupling of the pyridoxal 5'-phosphate Schiff base has been demonstrated in pyridoxal enzymes other than aspartate aminotransferase, and suggests the generality of the mechanism in the catalysis of aminotransferases related to aspartate aminotransferase. PMID- 11112528 TI - Hierarchical protein "un-design": insulin's intrachain disulfide bridge tethers a recognition alpha-helix. AB - A hierarchical pathway of protein folding can enable segmental unfolding by design. A monomeric insulin analogue containing pairwise substitution of internal A6-A11 cystine with serine [[Ser(A6),Ser(A11),Asp(B10),Lys(B28),Pro(B29)]insulin (DKP[A6-A11](Ser))] was previously investigated as a model of an oxidative protein-folding intermediate [Hua, Q. X., et al. (1996) J. Mol. Biol. 264, 390 403]. Its structure exhibits local unfolding of an adjoining amphipathic alpha helix (residues A1-A8), leading to a 2000-fold reduction in activity. Such severe loss of function, unusual among mutant insulins, is proposed to reflect the cost of induced fit: receptor-directed restoration of the alpha-helix and its engagement in the hormone's hydrophobic core. To test this hypothesis, we have synthesized and characterized the corresponding alanine analogue [[Ala(A6),Ala(A11),Asp(B10),Lys(B28), Pro(B29)]insulin (DKP[A6-A11](Ala))]. Untethering the A6-A11 disulfide bridge by either amino acid causes similar perturbations in structure and dynamics as probed by circular dichroism and (1)H NMR spectroscopy. The analogues also exhibit similar decrements in thermodynamic stability relative to that of the parent monomer as probed by equilibrium denaturation studies (Delta Delta G(u) = 3.0 +/- 0.5 kcal/mol). Despite such similarities, the alanine analogue is 50 times more active than the serine analogue. Enhanced receptor binding (Delta Delta G = 2.2 kcal/mol) is in accord with alanine's greater helical propensity and more favorable hydrophobic-transfer free energy. The success of an induced-fit model highlights the applicability of general folding principles to a complex binding process. Comparison of DKP[A6 A11](Ser) and DKP[A6-A11](Ala) supports the hypothesis that the native A1-A8 alpha-helix functions as a preformed recognition element tethered by insulin's intrachain disulfide bridge. Segmental unfolding by design provides a novel approach to dissecting structure-activity relationships. PMID- 11112529 TI - Orientation of the Mn(II)-Mn(III) dimer which results from the reduction of the oxygen-evolving complex of photosystem II by NO: an electron paramagnetic resonance study. AB - The central part of the oxygen-evolving complex of photosystem II is a cluster of four manganese atoms. The known EPR spectra in the various oxidation states of the cluster are complicated by the magnetic interactions of the four Mn ions and accordingly are difficult to analyze. It has been shown recently that NO at -30 degrees C slowly reduces the cluster to a Mn(II)-Mn(III) state [Sarrou, J., Ioannidis, N., Deligiannakis, Y., and Petrouleas, V. (1998) Biochemistry 37, 3581 3587). We study herein the orientation dependence of the Mn(II)-Mn(III) EPR spectrum with respect to the thylakoid membrane plane. Both the powder and the oriented spectra are satisfactorily simulated with the same set of fine and hyperfine parameters assuming axial symmetry and collinear g and A tensors. The axial component of the tensors is found to be oriented at an angle of 20 degrees +/- 10 degrees to the membrane plane normal (mosaic spread Omega = 40 degrees ). We make the reasonable assumption that the Mn(II)-Mn(III) dimer is one of the di mu-oxo units that has been suggested to comprise the Mn tetramer. On the basis of the sign of the hyperfine tensor anisotropy, the axial direction is assigned to the d(z(2)) orbital of Mn(III), which by comparison with synthetic model complexes is assumed to be oriented perpendicular to the Mn-(mu-oxo)-Mn plane. The present results complement earlier orientation studies by EXAFS and suggest that the Mn-(mu-oxo)-Mn plane makes a small angle (approximately 20 degrees) with the membrane plane and the axis connecting the bridging oxygens is approximately parallel to the plane. PMID- 11112530 TI - Reaction of nitric oxide with the turnover intermediates of cytochrome c oxidase: reaction pathway and functional effects. AB - The reactions of nitric oxide (NO) with the turnover intermediates of cytochrome c oxidase were investigated by combining amperometric and spectroscopic techniques. We show that the complex of nitrite with the oxidized enzyme (O) is obtained by reaction of both the "peroxy" (P) and "ferryl" (F) intermediates with stoichiometric NO, following a common reaction pathway consistent with P being an oxo-ferryl adduct. Similarly to chloride-free O, NO reacted with P and F more slowly [k approximately (2-8) x 10(4) M(-1) s(-1)] than with the reduced enzyme (k approximately 1 x 10(8) M(-1) s(-1)). Recovery of activity of the nitrite inhibited oxidase, either during turnover or after a reduction-oxygenation cycle, was much more rapid than nitrite dissociation from the fully oxidized enzyme (t(1/2) approximately 80 min). The anaerobic reduction of nitrite-inhibited oxidase produced the fully reduced but uncomplexed enzyme, suggesting that reversal of inhibition occurs in turnover via nitrite dissociation from the cytochrome a(3)-Cu(B) site: this finding supports the hypothesis that oxidase may have a physiological role in the degradation of NO into nitrite. Kinetic simulations suggest that the probability for NO to be transformed into nitrite is greater at low electron flux through oxidase, while at high flux the fully reduced (photosensitive) NO-bound oxidase is formed; this is fully consistent with our recent finding that light releases the inhibition of oxidase by NO only at higher reductant pressure [Sarti, P., et al. (2000) Biochem. Biophys. Res. Commun. 274, 183]. PMID- 11112531 TI - The proton/electron coupling ratio at heme a and Cu(A) in bovine heart cytochrome c oxidase. AB - Measurements of the H(+)/heme a, Cu(A) ratios for proton-electron coupling at these centers (redox Bohr effect) in CO-inhibited cytochrome c oxidase purified from bovine heart mitochondria, both in the soluble state and reconstituted in liposomes, are presented. In the soluble oxidase, the H(+)/heme a, Cu(A) ratios were experimentally determined upon oxidation by ferricyanide of these centers as well as upon their reduction by hexammineruthenium(II). These measurements showed that in order to obtain H(+)/heme a, Cu(A) ratios approaching 1, one-step full oxidation of both metal centers by ferricyanide had to be induced by a stoicheiometric amount of the oxidant. Partial stepwise oxidation or reduction of heme a and Cu(A) did produce H(+)/heme a, Cu(A) ratios significantly lower or higher than 1, respectively. The experimental H(+)/heme a, Cu(A) ratios measured upon stepwise reduction/oxidation of the metals were reproduced by mathematical simulation based on the coupling of oxido-reduction of both heme a and Cu(A) to pK shifts of common acid-base groups. The vectorial nature of the proton-electron coupling at heme a/Cu(A) was analyzed by measuring pH changes in the external bulk phase associated with oxido-reduction of these redox centers in the CO inhibited oxidase reconstituted in liposomes. The results show that the proton release associated with the oxidation of heme a and Cu(A) takes place in the external aqueous phase. Protons taken up by the oxidase upon rereduction of the centers derive, on the other hand, from the inner space. These results provide evidence supporting the view that cooperative proton-electron coupling at heme a/Cu(A) is involved in the proton pump of the oxidase. PMID- 11112532 TI - Synthesis and biophysical analysis of transmembrane domains of a Saccharomyces cerevisiae G protein-coupled receptor. AB - The Ste2p receptor for alpha-factor, a tridecapeptide mating pheromone of the yeast Saccharomyces cerevisiae, belongs to the G protein-coupled family of receptors. In this paper we report on the synthesis of peptides corresponding to five of the seven transmembrane domains (M1-M5) and two homologues of the sixth transmembrane domain corresponding to the wild-type sequence and a mutant sequence found in a constitutively active receptor. The secondary structures of all new transmembrane peptides and previously synthesized peptides corresponding to domains 6 and 7 were assessed using a detailed CD analysis in trifluoroethanol, trifluoroethanol-water mixtures, sodium dodecyl sulfate micelles, and dimyristoyl phosphatidyl choline bilayers. Tryptophan fluorescence quenching experiments were used to assess the penetration of the membrane peptides into lipid bilayers. All peptides were predominantly (40-80%) helical in trifluoroethanol and most trifluoroethanol-water mixtures. In contrast, two of the peptides M3-35 (KKKNIIQVLLVASIETSLVFQIKVIFTGDNFKKKG) and M6-31 (KQFDSFHILLINleSAQSLLVPSIIFILAYSLK) formed stable beta-sheet structures in both sodium dodecyl sulfate micelles and DMPC bilayers. Polyacrylamide gel electrophoresis showed that these two peptides formed high molecular aggregates in the presence of SDS whereas all other peptides moved as monomeric species. The peptide (KKKFDSFHILLIMSAQSLLVLSIIFILAYSLKKKS) corresponding to the sequence in the constitutive mutant was predominantly helical under a variety of conditions, whereas the homologous wild-type sequence (KKKFDSFHILLIMSAQSLLVPSIIFILAYSLKKKS) retained a tendency to form beta-structures. These results demonstrate a connection between a conformational shift in secondary structure, as detected by biophysical techniques, and receptor function. The aggregation of particular transmembrane domains may also reflect a tendency for intermolecular interactions that occur in the membrane environment facilitating formation of receptor dimers or multimers. PMID- 11112533 TI - Probing the role of the Fe-S subunit hinge region during Q(o) site catalysis in Rhodobacter capsulatus bc(1) complex. AB - The ubihydroquinone:cytochrome c oxidoreductase, or bc(1) complex, functions according to a mechanism known as the modified Q cycle. Recent crystallographic data have revealed that the extrinsic domain containing the [2Fe2S] cluster of the Fe-S subunit of this enzyme occupies different positions in various crystal forms, suggesting that this subunit may move during ubihydroquinone oxidation. As in these structures the hydrophobic membrane anchor of the Fe-S subunit remains at the same position, the movement of the [2Fe2S] cluster domain would require conformational changes of the hinge region linking its membrane anchor to its extrinsic domain. To probe the role of the hinge region, Rhodobacter capsulatus bc(1) complex was used as a model, and various mutations altering the hinge region amino acid sequence, length, and flexibility were obtained. The effects of these modifications on the bc(1) complex function and assembly were investigated in detail. These studies demonstrated that the nature of the amino acid residues located in the hinge region (positions 43-49) of R. capsulatus Fe-S subunit was not essential per se for the function of the bc(1) complex. Mutants with a shorter hinge (up to five amino acid residues deletion) yielded functional bc(1) complexes, but contained substoichiometric amounts of the Fe-S subunit. Moreover, mutants with increased rigidity or flexibility of the hinge region altered both the function and the assembly or the steady-state stability of the bc(1) complex. In particular, the extrinsic domain of the Fe-S subunit of a mutant containing six proline residues in the hinge region was shown to be locked in a position similar to that seen in the presence of stigmatellin. Interestingly, the latter mutant readily overcomes this functional defect by accumulating an additional mutation which shortens the length of the hinge. These findings indicate that the hinge region of the Fe-S subunit of bacterial bc(1) complexes has a remarkable structural plasticity. PMID- 11112534 TI - Proteolytic cleavage of the Fe-S subunit hinge region of Rhodobacter capsulatus bc(1) complex: effects of inhibitors and mutations. AB - The three-dimensional structure of the mitochondrial bc(1) complex reveals that the extrinsic domain of the Fe-S subunit, which carries the redox-active [2Fe2S] cluster, is attached to its transmembrane anchor domain by a short flexible hinge sequence (amino acids D43 to S49 in Rhodobacter capsulatus). In various structures, this extrinsic domain is located in different positions, and the conformation of the hinge region is different. In addition, proteolysis of this region has been observed previously in a bc(1) complex mutant of R. capsulatus [Saribas, A. S., Valkova-Valchanova, M. B., Tokito, M., Zhang, Z., Berry E. A., and Daldal, F. (1998) Biochemistry 37, 8105-8114]. Thus, possible correlations between proteolysis, conformation of the hinge region, and position of the extrinsic domain of the Fe-S subunit within the bc(1) complex were sought. In this work, we show that thermolysin, or an endogenous activity present in R. capsulatus, cleaves the hinge region of the Fe-S subunit between its amino acid residues A46-M47 or D43-V44, respectively, to yield a protease resistant fragment with a M(r) of approximately 18 kDa. The cleavage was affected significantly by ubihydroquinone oxidation (Q(o)) and ubiquinone reduction (Q(i)) site inhibitors and by specific mutations located in the bc(1) complex. In particular, using either purified or detergent dispersed chromatophore-embedded R. capsulatus bc(1) complex, we demonstrated that while stigmatellin blocked the cleavage, myxothiazol hardly affected it, and antimycin A greatly enhanced it. Moreover, mutations in various regions of the Fe-S subunit and cyt b subunit changed drastically proteolysis patterns, indicating that the structure of the hinge region of the Fe-S subunit was modified in these mutants. The overall findings establish that protease accessibility of the Fe-S subunit of the bc(1) complex is a useful biochemical assay for probing the conformation of its hinge region and for monitoring indirectly the position of its extrinsic [2Fe2S] cluster domain within the Q(o) pocket. PMID- 11112535 TI - The C-terminal domain of nucleolin accelerates nucleic acid annealing. AB - We report that the abundant nucleolar protein nucleolin accelerates nucleic acid annealing. Nucleolin accelerates annealing of complementary oligonucleotides and of oligonucleotides that contain a limited number of mismatches. The annealing activity of nucleolin can be localized to a C-terminal region consisting of two RNA binding domains (RBD3 and RBD4) and the RGG(9) domain (RBD3-RBD4-RGG(9)). This same region mediates self-association of nucleolin. The RGG(9) domain of nucleolin, believed to mediate interactions between nucleolin and several ribosomal proteins, is neither sufficient for self-association, as determined by small-angle X-ray scattering, nor can it independently accelerate annealing. Acceleration of nucleic acid annealing by nucleolin is likely to depend on self association of nucleolin molecules bound to nucleic acid. PMID- 11112536 TI - Evidence for changes in the nucleotide conformation in the active site of H(+) ATPase as determined by pulsed EPR spectroscopy. AB - The conformation of di- and triphosphate nucleosides in the active site of ATPsynthase (H(+)-ATPase) from thermophilic Bacillus PS3 (TF1) and their interaction with Mg(2+)/Mn(2+) cations have been investigated using EPR, ESEEM, and HYSCORE spectroscopies. For a ternary complex formed by a stoichiometric mixture of TF1, Mn(2+), and ADP, the ESEEM and HYSCORE data reveal a (31)P hyperfine interaction with Mn(2+) (|A((31)P)| approximately 5.20 MHz), significantly larger than that measured for the complex formed by Mn(2+) and ADP in solution (|A((31)P)| approximately 4.50 MHz). The Q-band EPR spectrum of the Mn.TF1.ADP complex indicates that the Mn(2+) binds in a slightly distorted environment with |D| approximately 180 x 10(-4) cm(-1) and |E| approximately 50 x 10(-4) cm(-1). The increased hyperfine coupling with (31)P in the presence of TF1 reflects the specific interaction between the central Mn(2+) and the ADP beta phosphate, illustrating the role of the enzyme active site in positioning the phosphate chain of the substrate for efficient catalysis. Results with the ternary Mn.TF1.ATP and Mn.TF1.AMP-PNP complexes are interpreted in a similar way with two hyperfine couplings being resolved for each complex (|A((31)P(beta))| approximately 4.60 MHz and |A((31)P(gamma))| approximately 5.90 MHz with ATP, and |A((31)P(beta))| approximately 4.20 MHz and |A((31)P(gamma))| approximately 5.40 MHz with AMP-PNP). In these complexes, the increased hyperfine coupling with (31)P(gamma) compared with (31)P(beta) reflects the smaller Mn.P distance with the gamma-phosphate compared with the beta-phosphate as found in the crystal structure of the analogous enzyme from mitochondria [3.53 vs 3.70 A (Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621-628)] and the different binding modes of the two phosphate groups. The ESEEM and HYSCORE data of a complex formed with Mn(2+), ATP, and the isolated beta subunit show that the (31)P hyperfine coupling is close to that measured in the absence of the protein, indicating a poorly structured nucleotide site in the isolated beta subunit in the presence of ATP. The inhibition data obtained for TF1 incubated in the presence of Mg(2+), ADP, Al(NO(3))(3), and NaF indicate the formation of the inhibited complex with the transition state analogue namely Mg.TF1.ADP.AlF(x) with the equilibrium dissociation constant K(D) = 350 microM and rate constant k = 0.02 min(-1). The ESEEM and HYSCORE data obtained for an inhibited TF1 sample, Mn.TF1.ADP.AlF(x), confirm the formation of the transition state analogue with distinct spectroscopic footprints that can be assigned to Mn.(19)F and Mn.(27)Al hyperfine interactions. The (31)P(beta) hyperfine coupling that is measured in the inhibited complex with the transition state analogue (|A((31)P(beta))| approximately 5.10 MHz) is intermediate between those measured in the presence of ADP and ATP and suggests an increase in the bond between Mn and the P(beta) from ADP upon formation of the transition state. PMID- 11112538 TI - Reconstitution and characterization of the Vibrio cholerae vibriobactin synthetase from VibB, VibE, VibF, and VibH. AB - Vibriobactin [N(1)-(2,3-dihydroxybenzoyl)-N(5),N(9)-bis[2-(2, 3-dihydroxyphenyl) 5-methyloxazolinyl-4-carboxamido]norspermidine] , is an iron chelator from the cholera-causing bacterium Vibrio cholerae. The six-domain, 270 kDa nonribosomal peptide synthetase (NRPS) VibF, a component of vibriobactin synthetase, has been heterologously expressed in Escherichia coli and purified. VibF has an unusual NRPS domain organization: cyclization-cyclization-adenylation-condensation peptidyl carrier protein-condensation (Cy(1)-Cy(2)-A-C(1)-PCP-C(2)). VibF activates and covalently loads its PCP with L-threonine, and together with vibriobactin synthetase proteins VibE (adenylation) and VibB (aryl carrier protein) condenses and heterocyclizes 2, 3-dihydroxybenzoyl-VibB with L-Thr to 2 dihydroxyphenyl-5-methyloxazolinyl-4-carboxy-VibF in the first demonstration of oxazoline formation by an NRPS cyclization domain. This enzyme-bound aryl oxazoline can be transferred by VibF to various amine acceptors but most efficiently to N(1)-(2, 3-dihydroxybenzoyl)norspermidine (k(cat) = 122 min(-1), K(m) = 1.7 microM), the product of 2,3-dihydroxybenzoyl-VibB, norspermidine, and VibH. This diacylated product undergoes a second aryl oxazoline acylation on its remaining secondary amine, also catalyzed by VibF, to yield vibriobactin. Vibriobactin biosynthesis in vitro has thus been accomplished from four proteins, VibE, VibB, VibF, and VibH, with the substrates 2,3-dihydroxybenzoic acid, L-Thr, norspermidine, and ATP. Vibriobactin synthetase is an unusual NRPS in that all intermediates are not covalently tethered as PCP thioesters and in that it represents an NRPS pathway with two branch points. PMID- 11112537 TI - Vibriobactin biosynthesis in Vibrio cholerae: VibH is an amide synthase homologous to nonribosomal peptide synthetase condensation domains. AB - The Vibrio cholerae siderophore vibriobactin is biosynthesized from three molecules of 2,3-dihydroxybenzoate (DHB), two molecules of L-threonine, and one of norspermidine. Of the four genes positively implicated in vibriobactin biosynthesis, we have here expressed, purified, and assayed the products of three: vibE, vibB, and vibH. All three are homologous to nonribosomal peptide synthetase (NRPS) domains: VibE is a 2,3-dihydroxybenzoate-adenosyl monophosphate ligase, VibB is a bifunctional isochorismate lyase-aryl carrier protein (ArCP), and VibH is a novel amide synthase that represents a free-standing condensation (C) domain. VibE and VibB are homologous to EntE and EntB from Escherichia coli enterobactin synthetase; VibE activates DHB as the acyl adenylate and then transfers it to the free thiol of the phosphopantetheine arm of VibB's ArCP domain. VibH then condenses this DHB thioester (the donor) with the small molecule norspermidine (the acceptor), forming N(1)-(2, 3 dihydroxybenzoyl)norspermidine (DHB-NSPD) with a k(cat) of 600 min(-1) and a K(m) for acyl-VibB of 0.88 microM and for norspermidine of 1.5 mM. Exclusive monoacylation of a primary amine of norspermidine was observed. VibH also tolerates DHB-acylated EntB and 1,7-diaminoheptane, octylamine, and hexylamine as substrates, albeit at lowered catalytic efficiencies. DHB-NSPD possesses one of three acylations required for mature vibriobactin, and its formation confirms VibH's role in vibriobactin biosynthesis. VibH is a unique NRPS condensation domain that acts upon an upstream carrier-protein-bound donor and a downstream amine, turning over a soluble amide product, in contrast to an archetypal NRPS embedded C domain that condenses two carrier protein thioesters. PMID- 11112539 TI - Psychrophilic elongation factor Tu from the antarctic Moraxella sp. Tac II 25: biochemical characterization and cloning of the encoding gene. AB - The elongation factor Tu was isolated from a psychrophilic eubacterial Antarctic Moraxella strain (MoEF-Tu) and its molecular and functional properties were determined. It catalyzed the synthesis of poly(Phe) and bound specifically guanine nucleotides with an affinity for GDP about 12-fold higher than that for GTP. The affinity toward guanine nucleotides was lower than that of other eubacterial EF-Tu. The intrinsic GTPase activity of MoEF-Tu was hardly detectable but was accelerated by 2 orders of magnitude in the presence of the antibiotic kirromycin (GTPase(k)). Such a property resembled Escherichia coli EF-Tu (EcEF Tu) even though the affinity of MoEF-Tu for the antibiotic was lower. MoEF-Tu showed a thermophilicity higher than that of EcEF-Tu; its temperature for half denaturation was 44 degrees C. The MoEF-Tu encoding gene corresponding to E. coli tufA was cloned and sequenced. The translated protein had a calculated molecular weight of 43 288 and contained the GTP-binding sequence motifs. Concerning its primary structure, MoEF-Tu showed sequence identity with E. coli and Thermus thermophilus EF-Tu equal to 84% and 74%, respectively, while the identity with EF 1 alpha from the archaeon Sulfolobus solfataricus was equal to 32%. PMID- 11112540 TI - Evolutionary coadaptation of the motif 2--acceptor stem interaction in the class II prolyl-tRNA synthetase system. AB - Known crystal structures of class II aminoacyl-tRNA synthetases complexed to their cognate tRNAs reveal that critical acceptor stem contacts are made by the variable loop connecting the beta-strands of motif 2 located within the catalytic core of class II synthetases. To identify potential acceptor stem contacts made by Escherichia coli prolyl-tRNA synthetase (ProRS), an enzyme of unknown structure, we performed cysteine-scanning mutagenesis in the motif 2 loop. We identified an arginine residue (R144) that was essential for tRNA aminoacylation but played no role in amino acid activation. Cross-linking experiments confirmed that the end of the tRNA(Pro) acceptor stem is proximal to this motif 2 loop residue. Previous work had shown that the tRNA(Pro) acceptor stem elements A73 and G72 (both strictly conserved among bacteria) are important recognition elements for E. coli ProRS. We carried out atomic group "mutagenesis" studies at these two positions of E. coli tRNA(Pro) and determined that major groove functional groups at A73 and G72 are critical for recognition by ProRS. Human tRNA(Pro), which lacks these elements, is not aminoacylated by the bacterial enzyme. An analysis of chimeric tRNA(Pro) constructs showed that, in addition to A73 and G72, transplantation of the E. coli tRNA(Pro) D-domain was necessary and sufficient to convert the human tRNA into a substrate for the bacterial synthetase. In contrast to the bacterial system, base-specific acceptor stem recognition does not appear to be used by human ProRS. Alanine-scanning mutagenesis revealed that motif 2 loop residues are not critical for tRNA aminoacylation activity of the human enzyme. Taken together, our results illustrate how synthetases and tRNAs have coadapted to changes in protein acceptor stem recognition through evolution. PMID- 11112541 TI - RNA-Catalyzed CoA, NAD, and FAD synthesis from phosphopantetheine, NMN, and FMN. AB - A novel in vitro selection method was developed to isolate RNA sequences with coenzyme-synthesizing activities. We used size-heterogeneous libraries containing randomized ribonucleotide sequences of four different lengths (30N, 60N, 100N, and 140N), all with 5'-ATP initiation. Two RNAs, CoES7 (30N) and CoES21 (60N), are able to catalyze the synthesis of three common coenzymes, CoA, NAD, and FAD, from their precursors, 4'-phosphopantetheine, NMN, and FMN, respectively. Both ribozymes require divalent manganese for activities. The results support the availability of these coenzymes in an RNA world, and point to a chemical explanation for the complex bipartite structures of many coenzymes. PMID- 11112542 TI - Recognition of nucleoside triphosphates during RNA-catalyzed primer extension. AB - In support of the idea that certain RNA molecules might be able to catalyze RNA replication, a ribozyme was previously generated that synthesizes short segments of RNA in a reaction modeled after that of proteinaceous RNA polymerases. Here, we describe substrate recognition by this polymerase ribozyme. Altering base or sugar moieties of the nucleoside triphosphate only moderately affects its utilization, provided that the alterations do not disrupt Watson-Crick pairing to the template. Correctly paired nucleotides have both a lower K(m) and a higher k(cat), suggesting that differential binding and orientation each play roles in discriminating matched from mismatched nucleotides. Binding of the pyrophosphate leaving group appears weak, as evidenced by a very inefficient pyrophosphate exchange reaction, the reverse of the primer-extension reaction. Indeed, substitutions at the gamma-phosphate can be tolerated, although poorly. Thio substitutions of oxygen atoms at the reactive phosphate exert effects similar to those seen with cellular polymerases, leaving open the possibility of an active site analogous to those of protein enzymes. The polymerase ribozyme, derived from an efficient RNA ligase ribozyme, can achieve the very fast k(cat) of the parent ribozyme when the substrate of the polymerase (GTP) is replaced by an extended substrate (pppGGA), in which the GA dinucleotide extension corresponds to the second and third nucleotides of the ligase. This suggests that the GA dinucleotide, which had been deleted when converting the ligase into a polymerase, plays an important role in orienting the 5'-terminal nucleoside. Polymerase constructs that restore this missing orientation function should achieve much more efficient and perhaps more accurate RNA polymerization. PMID- 11112543 TI - Characterization of Aspergillus niger pectate lyase A. AB - The Aspergillus niger plyA gene encoding pectate lyase A (EC 4.2.99. 3) was cloned from a chromosomal lambda(EMBL4) library using the Aspergillus nidulans pectate lyase encoding gene [Dean, R. A., and Timberlake, W. E. (1989) Plant Cell 1, 275-284] as a probe. The plyA gene was overexpressed using a promoter fusion with the A. niger pyruvate kinase promoter. Purification of the recombinant pectate lyase A resulted in the identification of two enzyme forms of which one appeared to be N-glycosylated and the other appeared to be free of N glycosylation. The two enzyme forms showed identical specific activities. The N glycosylation free pectate lyase A was further characterized with respect to product formation on polygalacturonic acid (alpha-1,4 linked D-galacturonic acid) and mode of action on oligogalacturonides of degree of polymerization 2-8. The bond cleavage frequencies for tetra-, penta-, and hexagalacturonides were studied as a function of [CaCl(2)]. The bond cleavage frequencies changed in a [CaCl(2)] dependent way for penta- and hexagalacturonide. Kinetic studies using tetra- and hexagalacturonide revealed a strong sigmoidal [CaCl(2)]-dependent relation. The role of Ca(2+) ions in substrate binding is discussed. PMID- 11112544 TI - Differential effects on N(2) binding and reduction, HD formation, and azide reduction with alpha-195(His)- and alpha-191(Gln)-substituted MoFe proteins of Azotobacter vinelandii nitrogenase. AB - In contrast to the wild-type MoFe protein, neither the alpha-195(Asn) nor the alpha-191(Lys) MoFe protein catalyzed N(2) reduction to NH(3), when complemented with wild-type Fe protein. However, N(2) was bound by the alpha-195(Asn) MoFe protein and inhibited the reduction of both protons and C(2)H(2). The alpha 191(Lys) MoFe protein did not interact with N(2). With the alpha-195(Asn) MoFe protein, the N(2)-induced inhibition of substrate reduction was reversed by removing the N(2). Surprisingly, even though added H(2) also relieved N(2) inhibition of substrate reduction, the alpha-195(Asn) MoFe protein did not catalyze HD formation under a N(2)/D(2) atmosphere. This observation is the first indication that these two reactions have different chemical origins, prompting a revision of the current hypothesis that these two reactions are consequences of the same nitrogenase chemistry. A rationale that accounts for the dichotomy of the two reactions is presented. The two altered MoFe proteins also responded quite differently to azide. It was a poor substrate for both but, in addition, azide was an electron-flux inhibitor with the 195(Asn) MoFe protein. The observed reactivity changes are correlated with likely structural changes caused by the amino acid substitutions and provide important details about the interaction(s) of N(2,) H(2), D(2), and azide with Mo-nitrogenase. PMID- 11112545 TI - Spectral and kinetic studies on the formation of eosinophil peroxidase compound I and its reaction with halides and thiocyanate. AB - Compound I of peroxidases takes part in both the peroxidation and the halogenation reaction. This study for the first time presents transient kinetic measurements of the formation of compound I of human eosinophil peroxidase (EPO) and its reaction with halides and thiocyanate, using the sequential-mixing stopped-flow technique. Addition of 1 equiv of hydrogen peroxide to native EPO leads to complete formation of compound I. At pH 7 and 15 degrees C, the apparent second-order rate constant is (4.3 +/- 0.4) x 10(7) M(-1) s(-1). The rate for compound I formation by hypochlorous acid is (5.6 +/- 0.7) x 10(7) M(-1) s(-1). EPO compound I is unstable and decays to a stable intermediate with a compound II like spectrum. At pH 7, the two-electron reduction of compound I to the native enzyme by thiocyanate has a second-order rate constant of (1.0 +/- 0. 5) x 10(8) M(-1) s(-1). Iodide [(9.3 +/- 0.7) x 10(7) M(-1) s(-1)] is shown to be a better electron donor than bromide [(1.9 +/- 0.1) x 10(7) M(-1) s(-1)], whereas chloride oxidation by EPO compound I is extremely slow [(3.1 +/- 0.3) x 10(3) M(-1) s( 1)]. The pH dependence studies suggest that a protonated form of compound I is more competent in oxidizing the anions. The results are discussed in comparison with those of the homologous peroxidases myeloperoxidase and lactoperoxidase and with respect to the role of EPO in host defense and tissue injury. PMID- 11112546 TI - Ring opening of benzo[a]pyrene in the germ-free rat is a novel pathway for formation of potentially genotoxic metabolites. AB - The metabolism of benzo[a]pyrene (BP) is known to lead to a large number of oxygenated compounds, some of which can bind covalently to DNA. We have studied the integrated metabolism of BP in vivo in germ-free rats given (14)C-labeled BP. Urinary metabolites were separated into groups according to acidity using lipophilic ion exchangers. The groups were analyzed by mass spectrometry and were further fractionated by high-performance liquid chromatography. The fraction of urinary metabolites previously shown to contain N-acetylcysteine and glucuronic acid conjugates was found to contain derivatives of 7-oxo-benz[d]anthracene-3,4 dicarboxylic acid as major components. These compounds, which were identified by mass spectrometry and NMR, accounted for about 30% of the total metabolites in urine, demonstrating that, surprisingly, ring opening is a major pathway for metabolism of BP in the germ-free rat. The dicarboxylic acid may be excreted in urine as an ester glucuronide. By using the single cell gel electrophoresis or COMET assay, we were able to demonstrate that the anhydride of 7-oxo benz[d]anthracene-3, 4-dicarboxylic acid was an efficient inducer of DNA damage. Taken together, these results indicate that the novel ring opening metabolic pathway may provide alternative mechanisms for the toxicity of BP. PMID- 11112547 TI - Abietadiene synthase from grand fir (Abies grandis): characterization and mechanism of action of the "pseudomature" recombinant enzyme. AB - The oleoresin secreted by grand fir (Abies grandis) is composed of resin acids derived largely from the abietane family of diterpene olefins as precursors which undergo subsequent oxidation of the C18-methyl group to a carboxyl function, for example, in the conversion of abieta-7,13-diene to abietic acid. A cDNA encoding abietadiene synthase has been isolated from grand fir and the heterologously expressed bifunctional enzyme shown to catalyze both the protonation-initiated cyclization of geranylgeranyl diphosphate to the intermediate (+)-copalyl diphosphate and the ionization-dependent cyclization of (+)-copalyl diphosphate, via a pimarenyl intermediate, to the olefin end products. Abietadiene synthase is translated as a preprotein bearing an N-terminal plastidial targeting sequence, and this form of the recombinant protein expressed in Escherichia coli proved to be unsuitable for detailed structure-function studies. Since the transit peptide mature protein cleavage site could not be determined directly, a truncation series was constructed to delete the targeting sequence and prepare a "pseudomature" form of the enzyme that resembled the native abietadiene synthase in kinetic properties. Both the native synthase and the pseudomature synthase having 84 residues deleted from the preprotein converted geranylgeranyl diphosphate and the intermediate (+)-copalyl diphosphate to a nearly equal mixture of abietadiene, levopimaradiene, and neoabietadiene, as well as to three minor products, indicating that this single enzyme accounts for production of all of the resin acid precursors of grand fir. Kinetic evaluation of abietadiene synthase with geranylgeranyl diphosphate and (+)-copalyl diphosphate provided evidence for two functionally distinct active sites, the first for the cyclization of geranylgeranyl diphosphate to (+)-copalyl diphosphate and the second for the cyclization of (+)-copalyl diphosphate to diterpene end products, and demonstrated that the rate-limiting step of the coupled reaction sequence resides in the second cyclization process. The structural implications of these findings are discussed in the context of primary sequence elements considered to be responsible for binding the substrate and intermediate and for initiating the respective cyclization steps. PMID- 11112548 TI - c-Raf-1 RBD associates with a subset of active v-H-Ras. AB - Mutational analysis of the cRaf-1 Ras binding domain (RBD) identified several point mutants with elevated Ras binding. Detailed examination of the binding kinetics of one mutant (A85K) suggests that it associates with a greater range of isomeric conformers of v-H-Ras than wt-RBD. At limiting v-H-Ras concentrations, saturation binding to A85K-RBD is higher than to wt-RBD. Notably, in assay systems where the RBD concentration is limiting, no difference exists between wt RBD and A85K-RBD saturation levels in the presence of a sufficiently large molar excess of Ras. The inability of wt-RBD to saturate all bindable Ras/GTP (defined by its binding to A85K-RBD) suggests that Ras/GTP exists as several isoforms and that only a minority of these isoforms are capable of associating with wt-RBD. These findings provide the first experimental evidence in support of functionally distinct Ras/GTP isoforms. We also describe a novel analysis of such isoforms. PMID- 11112549 TI - Allosteric linkage between voltage and Ca(2+)-dependent activation of BK-type mslo1 K(+) channels. AB - The activation of BK type Ca(2+)-activated K(+) channels depends on both voltage and Ca(2+). We studied three point mutations in the putative voltage sensor S4 or S4-S5 linker regions in the mslo1 BK channels to explore the relationship between voltage and Ca(2+) in activating the channel. These mutations reduced the steepness of the open probability - voltage (P(o) - V) relation and increased the shift of the P(o) - V relations on the voltage axis in response to increases in the calcium concentration. It is striking that these two effects were reciprocally related for all three mutations, despite different effects of the mutations on other aspects of the voltage dependence of channel gating. This reciprocal relationship suggests strongly that the free energy contributions to channel activation provided by voltage and by calcium binding are simply additive. We conclude that the Ca(2+) binding sites and the voltage sensors do not directly interact. Rather they both affect the mslo1 channel opening through an allosteric mechanism, by influencing the conformational change between the closed and open conformations. The mutations changed the channel's voltage dependence with little effect on its Ca(2+) affinitiy. PMID- 11112550 TI - Transient formation of ubisemiquinone radical and subsequent electron transfer process in the Escherichia coli cytochrome bo. AB - To elucidate a unique mechanism for the quinol oxidation in the Escherichia coli cytochrome bo, we applied pulse radiolysis technique to the wild-type enzyme with or without a single bound ubiquinone-8 at the high-affinity quinone binding site (Q(H)), using N-methylnicotinamide (NMA) as an electron mediator. With the ubiquinone bound enzyme, the reduction of the oxidase occurred in two phases as judged from kinetic difference spectra. In the faster phase, the transient species with an absorption maximum at 440 nm, a characteristic of the formation of ubisemiquinone anion radical, appeared within 10 micros after pulse radiolysis. In the slower phase, a decrease of absorption at 440 nm was accompanied by an increase of absorption at 428 and 561 nm, characteristic of the reduced form. In contrast, with the bound ubiquinone-8-free wild-type enzyme, NMA radicals directly reduced hemes b and o, though the reduction yield was low. These results indicate that a pathway for an intramolecular electron transfer from ubisemiquinone anion radical at the Q(H) site to heme b exists in cytochrome bo. The first-order rate constant of this process was calculated to be 1.5 x 10(3) s(-1) and is comparable to a turnover rate for ubiquinol-1. The rate constant for the intramolecular electron transfer decreased considerably with increasing pH, though the yields of the formation of ubisemiquinone anion radical and the subsequent reduction of the hemes were not affected. The pH profile was tightly linked to the stability of the bound ubisemiquinone in cytochrome bo [Ingledew, W. J., Ohnishi, T., and Salerno, J. C. (1995) Eur. J. Biochem. 227, 903-908], indicating that electron transfer from the bound ubisemiquinone at the Q(H) site to the hemes slows down at the alkaline pH where the bound ubisemiquinone can be stabilized. These findings are consistent with our previous proposal that the bound ubiquinone at the Q(H) site mediates electron transfer from the low-affinity quinol oxidation site in subunit II to low-spin heme b in subunit I. PMID- 11112551 TI - Consequences of cAMP and catalytic-subunit binding on the flexibility of the A kinase regulatory subunit. AB - A combination of site-directed labeling and time-resolved fluorescence anisotropy was used to further elucidate the structure and underlying dynamic features of the type I regulatory (R(I)(alpha)) subunit of the cAMP-dependent protein kinase. Specifically, the consequences of cAMP and the catalytic (C)-subunit binding on the backbone flexibility around seven sites of cysteine substitution and fluorescein maleimide labeling (Thr(6)Cys, Leu(66)Cys, Ser(75)Cys, Ser(81)Cys, Ser(99)Cys, Ser(145)Cys, and Ser(373)Cys) in the R(I)(alpha) subunit were assessed. Focusing on the fast rotational correlation time, the results indicate that most of the interdomain segment connecting the dimerization/docking (D/D) and tandem cAMP-binding domains is probably weakly associated with the latter domain. Also, this segment becomes more tightly bound to the C subunit upon holoenzyme formation. The results also suggest that there is a short 'hinge' segment (around Leu(66)Cys) that could allow the structured interdomain/cAMP binding and D/D domains to pivot about each other. Finally, cAMP binding dramatically reduces the backbone flexibility around only the two sites of cysteine substitution in the cAMP-binding domains, suggesting a selective structural stabilization caused by cAMP and a "tight" coupling of low-nanosecond fluctuations selectively within the tandem cAMP-binding domains. PMID- 11112552 TI - Commitment to folded and aggregated states occurs late in interleukin-1 beta folding. AB - A point mutation, lysine 97 to isoleucine, in the all-beta cytokine interleukin-1 beta (IL-1 beta) exhibits an increased propensity to form inclusion bodies in vivo and aggregates in vitro. In an effort to better understand the aggregation reaction and determine when intervention may allow rescue of protein from aggregation during renaturation, we developed a novel application of mass spectrometry using isotopic labeling to determine the step(s) at which K97I commits to either the native or aggregated state. Interestingly, despite the early formation of a folding intermediate ensemble at an observed rate lambda(2) of 4.0 s(-1), K97I commits to folding at a significantly slower rate lambda(CF) of 0.021 s(-1). This rate of commitment to folding is in excellent agreement with the observed rate of K97I native state formation (lambda(1) = 0.018 s(-1)). K97I also commits slowly to aggregation at an observed rate lambda(CA) of 0.023 s(-1). Earlier folding species and aggregates present prior to these commitment steps are likely to be in a reversible equilibrium between monomeric folding intermediates and higher-order oligomers. Kinetic and equilibrium experimental measurements of folding and aggregation processes are consistent with a nucleation-dependent model of aggregation. PMID- 11112554 TI - Hypermodified nucleosides in the anticodon of tRNA(Lys) stabilize a canonical U turn structure PMID- 11112553 TI - Equilibrium and kinetic studies on folding of the authentic and recombinant forms of human alpha-lactalbumin by circular dichroism spectroscopy. AB - The equilibrium and kinetics of the unfolding and refolding of authentic and recombinant human alpha-lactalbumin, the latter of which had an extra methionine residue at the N-terminus, were studied by circular dichroism spectroscopy, and the results were compared with the results for bovine and goat alpha-lactalbumins obtained in our previous studies. As observed in the bovine and goat proteins, the presence of the extra methionine residue in the recombinant protein remarkably destabilized the native state, and the destabilization was entirely ascribed to an increase in the rate of unfolding. The thermodynamic stability of the native state against the unfolded state was lower, and the thermodynamic stability of the molten globule state against the unfolded state was higher for the human protein than for the other alpha-lactalbumins previously studied. Thus, the population of the molten globule intermediate was higher during the equilibrium unfolding of human alpha-lactalbumin by guanidine hydrochloride. Unlike the molten globule states of the bovine and goat proteins, the human alpha lactalbumin molten globule showed remarkably more intense circular dichroism ellipticity than the native state in the far-ultraviolet region below 225 nm. During refolding from the unfolded state, human alpha-lactalbumin thus exhibited overshoot kinetics, in which the alpha-helical peptide ellipticity exceeded the native value when the molten globule folding intermediate was formed in the burst phase. The subsequent folding involved reorganization of nonnative secondary structures. It should be noted that the rate constant of the major refolding phase was approximately the same among the three types of alpha-lactalbumin and that the rate constant of unfolding was accelerated 18-600 times in the human protein, and these results interpreted the lower thermodynamic stability of this protein. PMID- 11112555 TI - Kinetic mechanism of human histone acetyltransferase P/CAF PMID- 11112556 TI - Peroxycarbenium-mediated C-C bond formation: applications to the synthesis of hydroperoxides and peroxides. AB - The Lewis acid-mediated reaction of alkene nucleophiles with peroxyacetals provides an effective route for the synthesis of homologated peroxides and hydroperoxides. In the presence of Lewis acids such as TiCl(4), SnCl(4), and trimethylsilyl triflate, peroxyacetals and peroxyketals undergo reaction with allyltrimethylsilane, silyl enol ethers, and silyl ketene acetals to afford homoallyl peroxides, 3-peroxyketones, and 3-peroxyalkanoates, respectively. Reactions of peroxyacetals are Lewis acid dependent; TiCl(4) promotes formation of ethers while SnCl(4) and trimethylsilyl triflate promote formation of peroxides. Lewis acid-promoted reactions of silylated hydroperoxyacetals furnish silylated hydroperoxides, which can be deprotected to homologated hydroperoxides. Hydroperoxyketals undergo Lewis acid-mediated allylation to furnish 1,2 dioxolanes via attack of hydroperoxide on the intermediate carbocation. Lewis acid-mediated cyclization of unsaturated peroxyacetals furnishes 1,2-dioxanes, 1,2-dioxepanes, and 1,2-dioxacanes through 6-endo/exo, 7-endo/endo, and 8 endo/endo pathways. The corresponding reactions involving 6-endo/endo and 5 endo/exo pathways were unsuccessful. PMID- 11112557 TI - A practical synthesis of a COX-2-specific inhibitor. AB - A number of synthetic strategies to the Cox-2 specific inhibitor 1 have been described. These studies have led to the identification of a novel pyridine construction using annulation of ketone 2 using a vinamidinium species 29 and ammonia in 97% assay yield. Three approaches to the synthesis of ketone 2 are described that allow for its preparation in large quantities in >65% overall yield from methyl 6-methylnicotinate. PMID- 11112558 TI - Density functional theory study of the mechanism of the BF(3)-catalyzed rearrangement of 2,3,3-trimethyl-1,2-epoxybutane to 2,3,3-trimethylbutanal. AB - The potential energy surface for the rearrangement of BF(3)-coordinated 2,3,3 trimethyl-1,2-epoxybutane to 2,3, 3-trimethylbutanal has been investigated at the B3LYP/6-31G level of theory. SCRF(SCI-PCM) solvent calculations and theoretical primary and secondary kinetic isotope effects at the same level of theory provide support for a two-step process with ring opening of the BF(3)-coordinated epoxide to a tertiary carbocation intermediate followed by hydride/deuteride migration to give aldehyde. The experimentally measured primary isotope effect (k(H)(D)/k(D)(H)) requires a correction for an appropriate secondary isotope effect to give a true isotope effect k(H)(H)/k(D)(H). For the lowest energy pathway for hydride migration, the calculated secondary kinetic isotope effect is 0.92, which when applied to the experimentally measured isotope effect of k(H)(D)/k(D)(H) = 1.73 gives a revised "true" primary kinetic isotope effect of k(H)(H)/k(D)(H) = 1.59. This compares with a calculated value of 2.01. From intermediate 15, migration of the C1-H(a) proton via 19 is energetically favored over C1-H(b) migration via 18 and this result is consistent with the experimental results in which hydride migration of the proton cis to the methyl is favored. PMID- 11112559 TI - Synthetic utilization of polynitroaromatic compounds. 1. S-derivatization of 1 substituted 2,4,6-trinitrobenzenes with thiols. AB - Reactions of 1-R-2,4,6-trinitrobenenes (R = alkyl, protected aldehyde, aminocarbonyl, cyano groups, or isoxazole ring) with thiol salts were investigated. In most cases, these reactions gave a mixture of minor para and major ortho substitution products. Reactions of N,N-disubstituted 2,4,6 trinitrobenzamides with S-,O-, and N-nucleophiles afforded products of substitution of the p-nitro group exclusively. 1-Cyano-2,4,6-trinitrobenzene was found to be the most reactive and the least selective: all three nitro groups can be substituted using an excess of thiol salts. 2-R-4, 6-dinitrobenzamides showed no regioselectivity under similar conditions to yield 1:1 mixtures of para and ortho isomers. PMID- 11112560 TI - Synthetic utilization of polynitroaromatic compounds. 2. Synthesis Of 4,6-dinitro 1,2-benzisothiazol-3-ones and 4,6-dinitro-1, 2-benzisothiazoles from 2-benzylthio 4,6-dinitrobenzamides. AB - Cyclization of 2-benzylthio-4,6-dinitrobenzamides to 4,6-dinitro-1, 2 benzisothiazol-3-ones was achieved using SO(2)Cl(2) in CH(2)Cl(2) at room temperature. Alkylation of these heterocycles proceeded in a nonregioselective manner to yield a mixture of corresponding O- and N-alkylated products. Oxidation of 4,6-dinitro-1,2-benzisothiazoles (50% H(2)O(2) in AcOH) afforded the corresponding S-oxides and S, S-dioxides, depending on oxidation conditions. Unexpectedly, oxidation of a 3-methoxy derivative resulted in ring opening with the formation of the corresponding sulfamide. Chlorination of these 4,6-dinitro 1,2-benzisothiazol-3-ones (PCl(5)-POCl(3)) gave the expected 3 chloroisothiazoles. PMID- 11112561 TI - Ultrasound effects on the photopinacolization of benzophenone. AB - Ultrasonic irradiation is able to modify the course of several photochemical reactions, especially bimolecular, proceeding via triplet states. These effects were illustrated in the study of benzophenone photopinacolization in ethanol. The rates and yields increase when sonication is applied simultaneously to UV irradiation. An explanation is based on a 2-fold effect: (i) light-absorbing transient species undergo sonolytic decomposition, making the photoconversion more efficient, and (ii) sonication induces the triplet state quenching, as shown by Stern-Volmer plots from experiments run in the presence of naphthalene, probably due to the easier collisional deactivation processes favored by the homogeneous distribution of the activated species. PMID- 11112563 TI - BF(3)-Induced rearrangement of aziridino cyclopropanes derived from 2 phenylsulfonyl 1,3-dienes. Application to the total synthesis of (+/-) ferruginine. AB - Total synthesis of the alkaloid (+/-)-ferruginine (1) has been developed via the 2-phenylsulfonyl 1,3-diene approach. BF(3)-induced rearrangement of the N protected cyclohexane aziridino cyclopropane 8, derived from its corresponding epoxy cyclopropane, afforded the desired tropane alkaloid skeleton 9 in good yield. Michael addition of nitroethane (as an acyl anion equivalent) and transformation of the nitro group of the adduct 10 to a keto function gave 11. Elimination of benzenesulfinic acid and subsequent replacement of the tosyl group by a methyl group afforded the title compound 1. PMID- 11112562 TI - Thiophosphorylation of histidine. AB - The preparation of a novel phosphorus species, thiophosphoramidate, has enabled the specific thiophosphorylation of histidine at its 3-position. The rates of phosphorylation and thiophosphorylation of histidine are reported, as well as the spectroscopic properties of both thiophosphoramidate and 3-thiophosphohistidine. Structural assignment of the latter was made by analogy to the NMR properties of the known 3-phosphohistidine. The alkylation of 3-thiophosphohistidine by phenacyl bromide serves as a model for the introduction of labeling or probe reagents into histidine phosphorothioate-containing proteins. PMID- 11112564 TI - New insights on the origins of the stereocontrol of the staudinger reaction: [2 + 2] cycloaddition between ketenes and N-silylimines. AB - Density-functional theory studies on the [2 + 2] reaction between N-silylimines and ketenes to form the corresponding 2-azetidinones (beta-lactams) help to clarify several aspects on the mechanism of the Staudinger reaction. This reaction has been studied experimentally by Panunzio et al. It is shown that the formation of the 2-azetidinone ring takes place via two consecutive reactions. The first reaction consists of the nucleophilic addition of the iminic nitrogen to the sp-hybridized carbon atom of the ketene, with simultaneous migration of the silyl group from the imine to the oxygen atom of the ketene. This leads to silyl enol intermediates, in good agreement with the experimental results. Formation of the N-silylated beta-lactam takes place via a domino reaction consisting of a conrotatory thermal electrocyclization followed by a new silatropic rearrangement. It is also found that isomerization of the starting N silylimine has a lower activation barrier than that associated with the formation of the C-N bond, which explains the stereochemical outcome experimentally observed. Further considerations on the asymmetric torquoelectronic effects involved in this reaction are also reported. PMID- 11112565 TI - 'Upenamide: An unprecedented macrocyclic alkaloid from the Indonesian sponge Echinochalina sp. AB - 'Upenamide (1) represents a new class of macrocyclic marine alkaloid possessing both spirooxaquinolizidinone and hemiaminal ring systems. It was isolated from the Indonesian sponge Echinochalina sp. The gross structure of 1 was elucidated by spectroscopic methods and accurate mass measurements. A suggestion is made as to its biogenetic origin. PMID- 11112566 TI - Substituent dependence of the diastereofacial selectivity in iodination and bromination of glycals and related cyclic enol ethers. AB - The stereochemical course of the electrophilic iodination and bromination of tri O-benzyl-D-glucal under various conditions has been compared to that of substituted dihydropyrans 2-5. IN(3) addition in acetonitrile affords trans-alpha iodoazides (80-87%), besides small amounts of trans-beta-adducts, in the presence or the absence of benzyloxy substituents at C-3 or C-4, and in agreement with bridged iodonium ion intermediates. In contrast, the diastereofacial selectivity of bromine addition in dichloroethane going through open bromo oxocarbenium ions depends strongly on the substituents. Whereas the trans-alpha-dibromides are the main (85-95%) adducts in the absence of C-4 and C-5 substituents, in their presence a moderate to exclusive selectivity for cis-alpha-addition (60-99%) is observed. The predominance of trans-alpha-addition is again observed whatever the substituents when the bromination is carried out in the same solvent but with a tribromide ion salt, supporting a concerted addition of the two bromine atoms under these conditions. Finally, bromine addition in methanol exhibits a completely different behavior with the nonselective formation of trans-alpha- and trans-beta-methoxybromides and a small dependence on the substituents. In agreement with the absence of azide trapping of any cationic intermediate, it is concluded that these brominations which do not go through an ionic intermediate are concerted additions of bromine and methanol with very loose rate- and product determining transition states. Finally, the substituent conformation at C-4 influences drastically the stereoselectivity in all these brominations. Evidence for alpha-anomeric control of the nucleophile approach at C-1 is given by the highly predominant formation of alpha-adducts, except in the methanolic bromination. The factors determining the versatile selectivity of the electrophile approach are discussed in terms of PPFMO theory and of the special mechanisms of glycal reactions. PMID- 11112567 TI - New syntheses of the C,D-ring pyrromethenones of phytochrome and phycocyanin. AB - Pyrromethenone 7, the C,D-ring segment of phytochrome (Pr, 4), has been prepared in an efficient fashion employing three new strategies. Each of these has potential advantages for the synthesis of labeled material. Our first approach is related to the Gossauer synthesis, with the difference that strong alkali is avoided in the condensation of the C- and D-ring components 8 and 17. The key silyloxypyrrole 17 was readily prepared on multigram scales beginning with inexpensive butyrolactone (10). A second synthesis began with 2 acetylbutyrolactone (41). The key steps involved conversion of 41 to the Z enoltriflate 42, followed by Pd(0)-catalyzed coupling with trimethylsilylacetylene, p-chlorophenylselenide ring opening, and finally, amidation to afford the ring-D synthon 45 having the proper geometry and oxidation state for conversion to 7. Sonogashira coupling of 45 with the iodopyrrole 22, followed by oxidative elimination, and F(-)-induced 5-exo-dig cyclization of the resultant pyrroloalkyne 47, then completed the synthesis. In similar fashion, we have also prepared pyrromethenone 6, the C,D-ring segment of phycocyanin (2). PMID- 11112568 TI - Total synthesis of (+/-)-deoxypenostatin A. Approaches to the syntheses of penostatins A and B. AB - A short synthesis of (+/-)-deoxypenostatin A (28) has been carried out using the convergent coupling of dienal 11, epoxide 13, and methylenetriphenylphosphorane (17) to prepare trienol 19 in only two steps. The key step is the Yb(OTf)(3) catalyzed intramolecular Diels-Alder reaction of hydrated trienyl glyoxylate 23, which gives lactone 24 stereoselectively. Elaboration of lactone 24 to enone 27 by an intramolecular Horner-Emmons Wittig reaction and epimerization completes the synthesis of 28. Modest yields of Diels-Alder adducts 45a and 46a could be prepared analogously from MEM ether 44c, but the sensitivity of several of the intermediates precluded the elaboration of 45a to penostatin A (1). PMID- 11112569 TI - An efficient, general asymmetric synthesis of carbocyclic nucleosides: application of an asymmetric aldol/ring-closing metathesis strategy. AB - A general and efficient synthesis of carbocyclic and hexenopyranosyl nucleosides has been developed. The strategy combines three key transformations: an asymmetric aldol addition to establish the relative and absolute configuration of the pseudosugar, a ring-closing metathesis to construct the pseudosugar ring, and a Trost-type palladium(0)-mediated substitution to assemble the pseudosugar and the aromatic base. Carbovir, abacavir, and their 2'-methyl derivatives as well as hexenopyranosyl nucleoside analogues have been prepared by this sequence. PMID- 11112570 TI - Molecular modeling (MM2 and PM3) and experimental (NMR and thermal analysis) studies on the inclusion complex of salbutamol and beta-cyclodextrin. AB - The inclusion complex of salbutamol and beta-cyclodextrin (beta-CD) is studied by computational (MM2 and PM3) and experimental techniques. Molecular modeling calculations predict two different orientations of salbutamol in the beta-CD cavity in vacuo and in aqueous solution. In vacuo calculations show that the introduction of the aromatic ring of salbutamol is preferred to the introduction of the tert-butyl group into the beta-CD cavity. However, in aqueous solution both computational methods predict the introduction of the alkyl chain instead of the aromatic ring in the beta-CD cavity contrary to experimental results published previously. These quantitative predictions were experimentally confirmed here by studying the inclusion complex in solution by NMR. A 1:1 stoichiometry was found by (1)H NMR studies for this complex. A 2D ROESY (rotating-frame Overhauser enhancement spectroscopy) experiment shows that there are no cross-peaks between the aromatic protons of salbutamol and any of the protons of beta-CD. Cross-peaks for the protons of the tert-butyl group and protons inside the cavity of beta-CD demonstrate the full involvement of this group in the complexation process and confirm the orientation of the complex predicted by molecular modeling. The solid-state complex was prepared and its stoichiometry (beta-CD.C(13)H(21)NO(3).8H(2)O) and dissociation process studied by thermogravimetric analysis. PMID- 11112571 TI - Chemoenzymatic synthesis of sialyl oligosaccharides with sialidases employing transglycosylation methodology. AB - A series of sialyloligosaccharides was synthesized using the transglycolytic activity of the sialidases from Vibrio cholerae, Clostridium perfringens, Salmonella typhimurium, and Newcastle disease virus. According to their hydrolytic activities the sialidases from V. cholerae and C. perfringens catalyze preferentially the formation of sialyl alpha(2-6)-linkages whereas the sialidases from S.typhimurium and Newcastle disease virus show a distinct preference for alpha(2-3) directed sialylations. Using combined chemical and enzymatic methodologies structures such as T-(Thomsen-Friedenreich) antigen [beta-D-Gal-(1 3)-alpha-D-GalNAc-OThr], Tn-(Thomsen nouveau) antigen (alpha-D-GalNAc-OThr) and beta-D-Gal-(1-4)-alpha-D-2-deoxy-Gal-OMe were sialylated in alpha(2-3)- and alpha(2-6)-positions regioselectively or in high regioisomeric excess and purified by simple isolation procedures. Depending on the enzyme source and acceptor structure yields for transsialylation varied between 10 and 30%. PMID- 11112572 TI - Soluble polymer-supported chemoenzymatic synthesis of the C(21)-C(27) fragment of the bryostatins. AB - A chemoenzymatic synthesis of the C(21)-C(27) fragment of the marine macrolide family of bryostatin antibiotics is presented. The approach commences from achiral starting materials and has as its crucial step the enzymatic resolution of a racemic mixture of soluble polymer-supported alcohols (syn-10 and syn-11). The immobilized lipase from Candida antarctica (Novozym 435) catalyzes the enantioselective acetylation of syn-10 (in 40% conversion and >99% ee), allowing isolation of the key intermediate (R)-14 in enantiomerically pure form following its cleavage from the poly(ethylene) glycol (PEG) scaffold. The PEG matrix is both compatible with the multipolymer enzymatic transformation and allows for rapid purification and facile NMR characterization of all intermediates throughout the synthesis. PMID- 11112573 TI - Studies on the oxidative addition reaction of 1,1-dibromo-1-alkenes, alpha dehalopalladation, and the intramolecular bis(carbopalladation) reaction of alkenes. An efficient entry to fused bicycles. AB - Twenty-three examples of 1,1-dihalo-1-alkenes were synthesized by the conventional alkylation methods. The oxidative addition reactions of 1,1-dibromo 2,2-diphenylethene or 1, 1-dibromo-2-phenylpropene with a stoichiometric amount of Pd(PPh(3))(4) afforded 1,2-diphenylacetylene and 1-phenylpropyne, respectively, indicating that alpha-dehalopalladation reaction occurred to afford vinylic carbene intermediates. However, alpha-dehalopalladation reaction was not observed in all 1, 1-dihalo-1-alkenes containing an extra C=C bond suitable for cyclic carbopalladation under the current reaction conditions probably due to the fast cyclic carbopalladation reaction of 40A-type of palladium intermediates; A series of bicycles, i.e., fused 5,6-, 6, 6-, 6,7-, and 7,7-bicyclic compounds, were prepared efficiently via this bicyclic carbopalladation protocol. Under condition A, within 0. 5 h, 10 afforded the monocyclic product 37 in 79%. With prolonged reaction time, 37 was converted to bicycle 36. Even with isolated 37, the corresponding reaction under condition A afforded 36 in 92% NMR yield, indicating a stepwise oxidative addition-cyclic carbopalladation-beta-elimination mechanism for this bicyclization reaction. PMID- 11112574 TI - Design, synthesis, and 1,3-dipolar cycloaddition of (5R)- [and (5S)]-5,6-dihydro 5-phenyl-2H-1,4-oxazin-2-one N-oxides as chiral (E)-geometry-fixed alpha alkoxycarbonylnitrones. AB - Optically pure (5R)- [and (5S)]-5,6-dihydro-5-phenyl-2H-1, 4-oxazin-2-one N oxides [(5R)- and (5S)-2] were designed as chiral (E)-geometry-fixed alpha alkoxycarbonylnitrones 1. The nitrones (5R)- and (5S)-2 were synthesized by three step oxidation of (R)- and (S)-phenylglycinols [(R)- and (S)-3], condensation of the resulting (R)- and (S)-2-hydroxylamino-2-phenylethanols [(R)- and (S)-5] with glyoxylic acid, and cyclization of the intermediary nitrones (R)- and (S)-6b. The nitrone (5R)-2reacted with olefins 7-14 under mild conditions to afford the corresponding cycloadducts 15-22 as the main products via the least sterically demanding exo modes. Cycloadduct 30 obtained from (5S)-2 and cyclopentadiene was effectively elaborated to (1S,4S, 5R)-4-benzyloxycarbonylamino-2 oxabicyclo[3.3.0]oct-7-en-3-one (28), the key synthetic intermediate of carbocyclic polyoxin C. PMID- 11112575 TI - Synthesis and biological evaluation of the geometric farnesylated analogues of the a-factor mating peptide of Saccharomyces cerevisiae. AB - The a-factor of Saccharomyces cerevisiae is a dodecapeptide pheromone (YIIKGVFWDPAC(Farnesyl)-OCH(3), 1), in which post-translational modification with a farnesyl isoprenoid and carboxymethyl group is required for full biological activity. This peptide has been used as a model system to explore the biological function of the farnesylcysteine moiety, which is found on and required for the biological activity of many key mammalian proteins. The objective of this particular study was the determination of the biological effect of double bond isomerization of the natural E, E-farnesyl moiety on the biological activity of the a-factor. A unified, stereoselective synthetic route to the three geometric isomers of E,E-farnesol (12, 13, and 14) has been developed. The key feature of this synthesis is the ability to control the stereochemistry of triflation of the beta-ketoester 22 to give either 23 or 25. The three farnesol isomers were converted to the corresponding isomeric a-factors (9, 10 and 11) via a modified version of a previously utilized synthetic route. Biological evaluation of these peptides indicates that, surprisingly, all three possess nearly equivalent activity to the natural a-factor bearing the E,E-farnesyl moiety. PMID- 11112576 TI - Synthesis of angular triquinanes from 1-Alkynylbicyclo[3.2. 0]hept-2-en-7-ones. A tandem alkoxy-cope ring Expansion/Transannular ring closure reaction. AB - Addition of ethenyllithium reagents to the carbonyl group of dialkyl squarate derived 1-alkynylbicyclo[3.2.0]hept-2-ene-7-ones (15), followed by a TBAF workup, results in a low-temperature anion-accelerated alkoxy-Cope rearrangement which proceeds by way of a strained cyclic allene intermediates (e.g.,17). This leads to the formation of angularly fused triquinanes (e.g., 20) in which each of the rings is functionally differentiated. Bicyclo[6.3. 0]undecadienones (e.g., 36) are the major products when the reactions are quenched with aqueous bicarbonate rather than TBAF. Under analogous conditions 2-alkylidene-1-alkynylbicyclo[3.2. 0]heptan-7-ones also give bicyclo[6.3.0]undecadienones by a mechanism that was established to involve a 1,5-hydrgen shift in a strained allene intermediate. The synthetic scope and mechanism of these and related transformations are discussed. PMID- 11112577 TI - On the aromaticity and Meisenheimer rearrangement of strained heterocyclic amine, phosphine, and arsine oxides. AB - A theoretical investigation (AIM and ELF analyses together with NMR chemical shifts) has been conducted for three-membered heterocycle (N, P, and As) oxides. An aromatic stabilization was found for the P and As rings. However, the N derivatives displayed a net negative hyperconjugation in the N-O bond formation, without ring aromaticity observed for their electronic properties. The calculated delta(C) and delta(H) shifts also supported the ring delocalization for the P and As unsaturated heterocycle oxides (delta(C) approximately 165 and delta(H) approximately 9 ppm). In addition, these values for 1H-azirine oxide resembled standard C=C double bond values (delta(C) approximately 130 and delta(H) approximately 7 ppm). The different behavior for the N oxides was also observed in their Meisenheimer rearrangement (MR). All the reaction paths, yielding the corresponding hydroxyl structures, were exothermic (G2 method). However, the N derivatives showed the lowest values for activation enthalpy, DeltaH(). The C=C bond influence in the MR was slight, with the same DeltaH values for the saturated and unsaturated paths. This rearrangement for the P and As oxides yielded TSs closer to the reactives; however, the corresponding TSs resembled the products for the N-derivatives. The different reaction paths have been examined by their corresponding AIM and ELF analyses at the B3LYP/6-311G level. PMID- 11112578 TI - Synthesis of dideuterated and enantiomers of monodeuterated tridecanoic acids at C-9 and C-10 positions. AB - We report a route for the preparation of mono and dideuterated tridecanoic acids: (R)-[9-(2)H(1)]-, (S)-[9-(2)H(1)]-, (R)-[10-(2)H(1)]-, (S)-[10-(2)H(1)]-, [9,9 (2)H(2)]-, and [10, 10-(2)H(2)]-tridecanoic acids required as probes for biochemical studies on desaturases. The key intermediates in the synthesis of all these probes are ketones 9, which give rise to the corresponding alcohols 10 and 13 by reduction with LiAlD(4) and LiAlH(4), respectively. Derivatization of nondeuterated racemic alcohols 13 with (S)-(+)-9-anthranylmethoxyacetic acid ((S) (+)-9-AMA) and chromatographic resolution of both diastereoisomers allowed us to determine the absolute configuration of the stereogenic centers by (1)H NMR using an adaptation of the model proposed by Riguera and co-workers which was validated with alcohols of known absolute configuration. Both enantiomeric alcohols (R)- and (S)-13 were recovered by reduction of each diastereomeric ester with LiAlH(4). Mesylation of alcohols 10 and 13 followed by nucleophilic substitution by LiAlD(4) generated the saturated methoxymethyl derivatives 12 and 16, respectively. Final deprotection and Jones oxidation of the resulting alcohols afforded the above deuterated tridecanoic acids. PMID- 11112579 TI - Organometallic reactions in aqueous media. Indium- and zinc-mediated allylation of sulfonimines. AB - Sulfonimines derived from aryl and nonenolizable aliphatic aldehydes can be effectively allylated to the corresponding homoallylic sulfonamides with allylic bromides promoted by indium or zinc. The solvent used can be water, THF, or a mixed aqueous THF solvent. The regioselectivity and stereoselectivity of the reaction were studied. PMID- 11112580 TI - Total synthesis of (-)-mniopetal E, a novel biologically intriguing drimane sesquiterpenoid. AB - We have achieved the total synthesis of (-)-mniopetal E, a drimane sesquiterpenoid which inhibits the reverse transcriptase of human immunodeficiency virus (HIV)-1. Our enantiospecific total synthesis of this target molecule in naturally occurring form commenced with a known 2,3-anhydro-D arabinitol derivative, which was prepared using the Sharpless asymmetric epoxidation strategy. The key steps of our total synthesis were as follows: (1) a combination of highly stereocontrolled inter- and intramolecular Horner-Emmons carbon elongations for construction of a butenolide tethering a 1,2,4, 9 functionalized nona-5,7-diene moiety at the beta-carbon, (2) stereoselective thermal intramolecular Diels-Alder reaction of the thus-formed trienic compound, providing preferentially an endo-cycloadduct with the desired pi-facial selection, and (3) efficient transformation of the gamma-lactone moiety in the major cycloadduct to the gamma-hydroxy-gamma-lactone part in mniopetal E. Our total synthesis of (-)-mniopetal E established the unsettled absolute stereochemistry of the antibiotic. PMID- 11112581 TI - Stereoselective formation of Bis(alpha-hydroxy ketones) via enzymatic carboligation. AB - The enzymatic approach to a novel class of chiral bis(alpha-hydroxy ketones) of type 5 and 8, which enable the synthesis of new multidentate ligands for asymmetric transition metal catalysis, is described. The key step is the second benzoylformate decarboxylase catalyzed C-C-bond formation between an aromatic dialdehyde and acetaldehyde, which proceeds with complete stereocontrol. Transformation of enantiomerically enriched monoadduct (S)-4 (ee 88%) and (S)-7 (ee 79%) resulted in optical pure (S,S)-5 and (S,S)-8 besides minor amounts of the corresponding diastereomeric meso-forms. PMID- 11112582 TI - Resolution and rotational barriers of quinolinone and acridone sulfenamide derivatives: demonstration of the S-N chiral axis. AB - Achiral (8a) and chiral (8b) N-(2,4-dinitrobenzenesulfenyl)acridone derivatives were synthesized. Addition of the chiral solvating agent (S)- 2,2,2-trifluro-1 (anthryl)ethanol to 8a rendered the enantiotopic groups on the acridone ring diastereotopic and anisochronous, thus allowing the estimation of a lower limit for the rotational barrier about the S-N bond (18.7 kcal mol(-)(1)) by NMR spectroscopy. 8b and the previously reported chiral sulfenamide 5 (Raban, M.; Martin, V. A.; Craine, L. J. Org. Chem. 1990, 55, 4311) were resolved on a Chiracel OD HPLC column. This constitutes the first resolution of stereostable enantiomers of a compound in which the chirality is due only to the presence of the S-N chiral axis. The rotational barriers of both compounds are nearly equal (22.7-22. 8 kcal mol(-)(1) at 303.7 K) and are the largest determined to date for the rotation about the S-N bond in sulfenamides. The relatively high enantiomerization barrier for 8b is remarkable since the peri positions are unsubstituted. PMID- 11112583 TI - Preparation and conformation of octaethylbiphenylene. AB - Dimerization of tetraethylbenzyne (generated by reaction of 1, 2-dibromo-3,4,5,6 tetraethylbenzene (8) with 1 equiv of BuLi) afforded in low yield octaethylbiphenylene (3), together with a major product which was characterized as 2,3,4,5,3',4', 5'-heptaethyl-2'-vinylbiphenyl (9). X-ray diffraction indicates that biphenylene 3 adopts in the crystal a conformation of approximate C(2)(h )()symmetry with the ethyl groups within each phenylene ring arranged in an alternated up-down fashion. Notably, pairs of vicinal ethyl groups located at peri positions are oriented in a syn arrangement in the crystal. Low temperature NMR spectroscopy is consistent with the presence in solution of either the crystal conformation or a fully alternated conformation lacking any syn interaction. Molecular mechanics (MM3), semiempirical (AM1, PM3), and ab initio calculations indicate that the crystal conformation is a high energy form, and that the lowest energy conformation is the fully alternated form. The topomerization barrier of the methylene protons of the ethyl groups of 3 is 9.4 +/- 0.1 kcal mol(-)(1), which is between the rotational barriers of 8 and 1,2,3, 4-tetraethylbenzene 7 (9.9 +/- 0.1 and 8.2 +/- 0.1 kcal mol(-)(1), respectively). The similarity in rotational barriers suggests that a given tetraethylphenylene subunit does not markedly affect the rotational barrier of the ethyl groups of the other subunit. PMID- 11112584 TI - Reactivity of alkyl versus silyl peroxides. The consequences of 1, 2-silicon bridging on the epoxidation of alkenes with silyl hydroperoxides and bis(trialkylsilyl)peroxides. AB - The bond dissociation energies for a series of silyl peroxides have been calculated at the G2 and CBS-Q levels of theory. A comparison is made with the O O BDE of the corresponding dialkyl peroxides, and the effect of the O-O bond strength on the activation barrier for oxygen atom transfer is discussed. The O-O bond dissociation enthalpies (DeltaH(298)) for bis (trimethylsilyl) peroxide (1) and trimethylsilyl hydroperoxide (2) are 54.8 and 53.1 kcal/mol, respectively at the G2 (MP2) and CBS-Q levels of theory. The O-O bond dissociation energies computed at G2 and G2(MP2) levels for bis(tert-butyl) peroxide and tert-butyl hydroperoxide are 45.2 and 48.3 kcal/mol, respectively. The barrier height for 1,2-methyl migration from silicon to oxygen in trimethylsilyl hydroperoxide is 47.9 kcal/mol (MP4//MP2/6-31G). The activation energy for the oxidation of trimethylamine to its N-oxide by bis(trimethylsilyl) peroxide is 28.2 kcal/mol (B3LYP/6-311+G(3df,2p)// B3LYP/6-31G(d)). 1,2-Silicon bridging in the transition state for oxygen atom transfer to a nucleophilic amine results in a significant reduction in the barrier height. The barrier for the epoxidation of E-2-butene with bis(dimethyl(trifluoromethyl))silyl peroxide is 25.8 kcal/mol; a reduction of 7.5 kcal/mol relative to epoxidation with 1. The activation energy calculated for the epoxidation of E-2-butene with F(3)SiOOSiF(3) is reduced to only 2.2 kcal/mol reflecting the inductive effect of the electronegative fluorine atoms. PMID- 11112585 TI - New asymmetric reactions of 2-formyl- and 2-acyl-1-[(2,4, 6 triisopropylphenyl)sulfinyl]naphthalenes via diastereomeric rotamers. AB - Nucleophilic reactions with Grignard reagents and the Mukaiyama aldol reactions of the naphthaldehydes having the (2,4, 6-triisopropylphenyl)sulfinyl group produced products with high stereoselectivity. In these reactions, the stereochemistry of the major products changes depending on the Lewis acids used. Reduction of the 2-acyl-1-[(2,4,6-triisopropylphenyl)sulfinyl]naphthalenes also proceeds with high stereoselectivity but with a different stereochemistry depending on the reducing agents. We have demonstrated, by the mechanistic consideration based on the X-ray crystal structures as well as the (1)H and (13)C NMR spectral data, that the extremely high and specific stereoselectivities of these reactions are due to the predominant rotamer around the C(naph)-S axis. Synthesis of enantiomerically pure 2-naphthylmethanol is provided as an example. PMID- 11112586 TI - Efficient epoxidation of alkenes with aqueous hydrogen peroxide catalyzed by methyltrioxorhenium and 3-cyanopyridine. AB - The epoxidation of alkenes with 30% aqueous hydrogen peroxide is catalyzed efficiently by methyltrioxorhenium (MTO) in the presence of pyridine additives. The addition of 1-10 mol % of 3-cyanopyridine increases the system's efficiency for terminal and trans-disubstituted alkenes resulting in high isolated yields of the corresponding epoxides. The system allows for epoxidation of alkenes with various functional groups. Alkenes leading to acid-sensitive products are efficiently epoxidized using a mixture of pyridine and 3-cyanopyridine as additives. This method is operationally very simple and uses an environmentally benign oxidant. The effects of different pyridine additives on the alkene conversion and the catalyst lifetime are discussed. PMID- 11112587 TI - Alkynyliodonium salts in organic synthesis. Dihydrofuran formation via a formal stevens shift of a carbon substituent within a disubstituted-carbon oxonium ylide. AB - The addition of p-toluenesulfinate to the silyl, 1-furanyl, and 1-pyranyl ethers of 1-hydroxybut-3-ynyl(phenyl)iodonium triflate triggers a sequence of reactions that ultimately delivers 2-substituted 3-p-toluenesulfonyldihydrofuran products in variable yields. A putative 1,2-group shift within an unsaturated oxonium ylide (Stevens rearrangement) accounts for the oxygen-to-carbon transfer of the ether substituent. Deuterium labeling studies clarify the mechanistic course of this shift by providing evidence consistent with intramolecular substituent transfer and by identifying the primary source of the proton that intercepts the ylide in the major yield-limiting process. PMID- 11112588 TI - Cascade radical reaction of 2-alkynyl-substituted aryl radicals with aryl isothiocyanates: a novel entry to benzothieno[2,3-b]quinolines through alpha (Arylsulfanyl)imidoyl radicals. AB - The novel cascade radical reaction of 2-(phenylalkynyl)aryl radicals with 4-Y phenyl isothiocyanates (Y = H, OMe, Me, Cl, CN) provides a useful one-pot protocol for the production of 8-Y-substituted (12) and/or 9-Y-substituted benzothieno[2,3-b]quinolines (11). The whole process entails primary formation of an alpha-(2-alkynylarylsulfanyl)imidoyl radical that undergoes smooth 5-exo-dig cyclization onto the alkynyl triple bond. The derived 1-phenylvinyl radical then exhibits six-membered cyclization onto the isothiocyanate ring, to give 11, and/or five-membered ipso-cyclization to an azaspiro intermediate, whose eventual rearrangement affords 12. The overall findings clearly showed that the relative proportion of the outcoming isomeric benzothienoquinolines 11 and 12 can be markedly affected by the nature of the original isothiocyanate substituent. Moreover, the findings also furnished the first chemical evidence that enhancing the electrophilic power of the employed radical can properly enhance the reactivity of aryl radicals toward isothiocyanates. PMID- 11112589 TI - Synthesis and properties of isoxazolo[60]fullerene-donor dyads. AB - A series of isoxazolo[60]fullerenes has been prepared in one pot from aldoximes under microwave irradiation. Several donors and acceptors were used as substituents. The absorption and emission spectra of these compounds in polar solvents suggest a weak charge-transfer interaction between the oxygen atom of the isoxazoline moiety and the C(60) cage, as well as a stronger interaction between the donor and the fullerene cage when the attached groups are p-N,N dimethylaniline or ferrocene. The electrochemical properties of the compounds were investigated and they show the same or better acceptor character than C(60) in all cases. Theoretical calculations support the results obtained. Solvent effects in the (1)H NMR spectra have been determined and provide useful information concerning the polarization of dyads. PMID- 11112590 TI - Electrolytic partial fluorination of organic compounds. 42.(1) marked solvent effects on regioselective anodic monofluorination of 4-oxo-2-pyrimidyl sulfides. AB - Regioselective anodic monofluorination of 4-oxo-2-pyrimidyl sulfides was investigated under various electrolytic conditions. Anodic fluorination was successfully carried out using Et(4)NF.4HF in dimethoxyethane (DME) to provide the corresponding alpha-fluorinated products in good yields. In contrast, acetonitrile (MeCN) was not suitable for the anodic fluorination due to the severe anode passivation during the electrolysis. A mixed solvent of DME and MeCN was found to be also effective for the fluorination, and the product yield increased with an increase of the ratio of DME to MeCN. The superiority of DME can be explained mainly in terms of the suppression of the anode passivation and enhancement of the nucleophilicity of the fluoride ions. Such marked solvent effects on the anodic fluorination were discussed in detail. PMID- 11112591 TI - Diastereoselective tandem addition-cyclization reactions of unsaturated tertiary amines initiated by photochemical electron transfer (PET). AB - Polycyclic molecules and tetrahydroquinoleines were obtained in a tandem reaction involving the diastereoselective addition of alpha-aminoalkyl radicals to (5R)-5 menthyloxy-2[5H]-furanone 1. The facial diastereoselectivity on 1 is >/=90%. The alpha-aminoalkyl radicals were produced from tertiary amines by photochemical induced electron transfer. When N,N-dialkylanilines 19 were used as starting tertiary amines, a rearomatization step was involved and important side reactions of 1 were observed. A mechanistic study involving isotopic labeling of the starting amine indicated that the byproducts resulted from reduction of 1 during the rearomatization process. An efficient optimization of the reaction was obtained by simply adding acetone or cyclopentanone as mild oxidants to the reaction mixture. The side products resulting from reduction of the furanone 1 were completely suppressed under these conditions, and the yields of the tetrahydroquinolines 21a-i, 22a-f, and 26g-i were doubled. PMID- 11112592 TI - Applications of the sulfinimine-mediated asymmetric strecker synthesis to the synthesis of alpha-alkyl alpha-amino acids. AB - Addition of Et(2)AlCN and i-PrOH to ketosulfinimines (N-sulfinyl imines) affords corresponding alpha-alkyl alpha-amino nitriles in moderate to good yields. The diastereoselectivity is largely dependent on the E/Z isomer ratio of the ketosulfinimine. Hydrolysis of the diastereomerically pure amino nitriles affords enantiopure alpha-alkyl alpha-amino acids in moderate to good yields. PMID- 11112593 TI - Competitive formation of helical cycloocta- and cyclododecapyrroles. AB - In connection with a study aimed at the evaluation of electronic effects in spiro dicorrole (1a) and its binuclear Ni(II) complex (1b) we became interested in gem dimethyl-substituted cyclotetrapyrrole (2a) and the corresponding Ni(II) complex (2b). Attempts to prepare 2a as the 12,13,16,17-tetraethyl-2,3,7, 8-tetramethyl derivative (5) by an acid-catalyzed (1 + 1) condensation of dimethyldipyrrylmethane 3 and diformylbipyrrole 4 resulted in the formation of the (2 + 2) and (3 + 3) condensation products, i.e., the cyclooctapyrrole 6 and the cyclododecapyrrole 7, respectively, rather than in that of the desired gem dimethyl cyclotetrapyrrole. The cyclododecapyrrole 7, isolated as the major product, is among the largest cyclopolypyrroles known to date. These two new macrocycles have been structurally characterized by variable temperature 1D and 2D NMR experiments, as well as by single-crystal X-ray diffraction analysis. In solution both the cyclooctapyrrole 6 and cyclododecapyrrole 7 exhibit dynamic behavior. At 337 K 6 adopts a D(2)-symmetric conformation, whereas at 196 K two equivalent C(2) conformers that interconvert through the D(2)-symmetric intermediate are observed. The energy barrier for the interconversion process between these two degenerate conformers is found to be 10.6 kcal mol(-)(1). The solution dynamics of 7 could be described in an analogous manner, with the time averaged conformation at 378 K displaying D(3)(h)() symmetry. X-ray analyses showed that for both macrocycles, 6 and 7, the solid state structures were nearly identical to the low-temperature solution conformers. PMID- 11112594 TI - Conjugate addition reactions of alpha-aminoalkylcuprates with alpha, beta-alkenyl , alpha,beta-alkynyl-, alpha,beta-beta,gamma-allenyl-, and alpha,beta-gamma,delta dienyl carboxylic acid derivatives, nitriles, and sulfoxides. AB - alpha-Aminoalkylcuprates prepared from alpha-lithio carbamates and CuCN.2LiCl participate in 1,4-addition reactions with alpha, beta-unsaturated esters, thiol esters, imides, and nitriles in poor to excellent yields depending upon the electron-withdrawing substituent and the substitution pattern of the unsaturated substrate. These reagents also undergo conjugate addition reactions with alpha,beta-alkynyl esters, sulfoxides, and nitriles and with alpha,beta beta,gamma-unsaturated allenyl esters. Excellent stereocontrol is achieved in the conjugate additions of alpha-aminoalkylcuprates to the allenyl esters, while poor stereoselectivity results in the conjugate additions to the alkynyl derivatives. Deprotection and cyclization of the alkynyl adducts affords pyrrolin-2-ones, while similar treatment of the allenyl adducts affords 4-alkylidine- pyrrolidin-2 ones and pyrrolizidinones. PMID- 11112595 TI - P-Nitrosophosphate compounds: new N-O heterodienophiles and nitroxyl delivery agents. AB - P-Nitrosophosphates, such as 9, react as N-O heterodienophiles with 1,3-dienes to form highly functionalized cycloadducts that can be directly transformed into allylic phosphoramidates. The in situ periodate oxidation of the unstable N hydroxyphosphoramidate precursors provides an efficient preparation of these new reactive intermediates. P-Nitrosophosphate (9) regioselectively reacts with 1 methoxy-1,3-butadiene to provide cycloadduct 16. P-Nitrosophosphate (9) also reacts with 9,10-dimethylanthracene to give cycloadduct 17, which undergoes retro Diels-Alder dissociation to re-form 9. In the absence of a 1,3-diene, the decomposition of 17 produces nitrous oxide, evidence for nitroxyl, the one electron-reduced form of nitric oxide. An asymmetric P-nitrosophosphate reacted with 1,3-cyclohexadiene to form a mixture of diastereomeric cycloadducts (19 and 20) in a 1.6:1 ratio. These results identify P-nitrosophosphates as new species that react similarly to acyl nitroso compounds, making them useful synthetic intermediates and potential nitroxyl delivery agents. PMID- 11112596 TI - Synthesis of the C(29)-C(45) bis-pyran subunit (E-F) of spongistatin 1 (Altohyrtin A). AB - A synthesis of the C(29)-C(45) bis-pyran subunit 2 of spongistatin 1 (1a) is described. The synthesis proceeds in 19 steps from the chiral aldehyde ent-7, and features highly diastereoselective alpha-alkoxyallylation reactions using the gamma-alkoxy substituted allylstannanes 17 and 19, as well as a thermodynamically controlled intramolecular Michael addition to close the F-ring pyran. The E ring was assembled via the Mukaiyama aldol reaction of F-ring methyl ketone 3 and the 2,3-syn aldehyde 4. PMID- 11112597 TI - Enantioselective synthesis of a fluorinated analogue of the orsellinic acid-type twelve-membered lactone lasiodiplodin. AB - The chemoenzymatic synthesis of the racemate and the one enantiomer of the fluorinated analogue 8 of the natural cyclooxygenase inhibitor lasiodiplodin is decribed. A lipase-mediated deracemization of the fluorohydrin 18 provided the chiral, nonracemic building block for the enantioselective synthesis of the title compound. The key step was the formation of the 12-membered lactone by a ring closing metathesis. PMID- 11112598 TI - Synthesis of persialylated beta-cyclodextrins. AB - The synthesis of homogeneous hepta-antennated C-6 branched sialosyl cyclomaltoheptose derivatives (persialylated beta-cyclodextrins) has been performed in good to excellent yields, and the compounds have been fully characterized. The thioacetate N-acetylneuraminic acid derivative 6 was selectively de-S-acetylated and coupled by nucleophilic displacement in a one-pot reaction to the heptakis(chloroacetamido) beta-CDs 2 and 5, yielding multivalent sialosides 8 and 9, respectively. The thiourea-linked sialyl-CD 10 was obtained by reaction of the 4-isothiocyanatophenyl N-acetylneuraminic acid derivative 7 with the per-tert-butoxycarbonylamino beta-CD derivative 2 after suitable deprotection of the amino function. PMID- 11112599 TI - Coordination control of intramolecular electron transfer in boronate-bridged zinc porphyrin-diimide molecules. AB - Three sets of dyads, in which a zinc-porphyrin (ZP) electron donor is connected to an aromatic diimide electron acceptor,either pyromellitimide (PI) or naphthalene-1,8:4,5-tetracarboxylic acid diimide (NI), via a boronate-ester bridge, a piperidine bridge, and a 1,3-dioxolane bridge, respectively, were prepared for the purpose of control of intramolecular electron transfer (ET) by acid-base reactions at the connecting bridge. Boronate-ester bridge is a Lewis acidic site and confers a chance to regulate intramolecular ET reaction upon base coordination. This has been demonstrated by suppression of photoinduced ET from ZP to PI or NI in highly electron-pair donating solvents or upon addition of a fluoride anion. To extend this strategy to control of ET-path selectivity, we prepared triad 18, which consists of a ZP donor bearing NI and PI acceptors at similar distances through a boronate-ester bridge and an acetal bridge, respectively. Photoexcitation of 18 in a free form led to intramolecular ET from (1)()ZP preferentially to NI, but the ET path was completely switched toward PI in F(-)-coordinated form without a serious drop in the rate, constituting a novel ET-switching molecular system. PMID- 11112600 TI - Ring-closing alkyne metathesis. Application to the total synthesis of sophorolipid lactone. AB - The first total synthesis of a major component of the microbial biosurfactant sophorolipid has been achieved. This approach to the 26-membered macrolide 1 containing a Z-configured alkene group in its lipidic tether spanning the sophorose backbone is based on a ring-closing metathesis reaction of diyne 21 catalyzed by Mo[N(t-Bu)(Ar)](3) (5; Ar = 3,5-dimethylphenyl) activated in situ by CH(2)Cl(2), followed by Lindlar reduction of the resulting cycloalkyne 22. The two beta-glycosidic linkages of compound 21 were installed by means of the glucal epoxide method and a modified Koenigs-Knorr reaction promoted by AgOTf/lutidine, respectively. PMID- 11112601 TI - A facile synthesis of perfluoroalkyl vinyl iodides and their palladium-mediated cross-coupling reactions. AB - Catalytic amounts of zinc, as low as 10 mol %, in the presence of trifluoroacetic acid (TFA) initiate the radical addition of perfluoroalkyl iodides to terminal alkynes with high regio- and stereoselectivities. Palladium-mediated cross coupling of these (E)-perfluoroalkyl vinyl iodides allows for a facile synthesis of potentially useful fluoroorganic intermediates. PMID- 11112602 TI - The hydroboration of propargyl bromide. Simple one-Pot three-component routes to (Z)-1-bromoalk-1-en-4-ols and to anti-homoallylic alcohols. AB - The hydroboration of propargyl bromide with dialkylboranes takes place regioselectively to give 3-bromoprop-1-en-1-yl dialkylboranes 13 which, upon quaternization with bromide ion, undergo a series of transformations into a number of allylic boron species. By a suitable choice of the experimental conditions it is possible to trap the reaction intermediates with aldehydes and to steer the process toward either the synthesis of (Z)-1-bromoalk-1-en-4-ols 6 or anti-homoallylic alcohols 8. Two one-pot three-component processes were developed based on a sequence of four reactions; preparation of dialkylborane and hydroboration of propargyl bromide are the first steps. Then, quaternization with TEBABr may be carried out either in the presence of the aldehyde when (Z)-1 bromoalk-1-en-4-ols 6 are requested, or in the absence of the aldehyde in order to allow the formation of gamma-substituted allyl borane 18 which, successively, adds to the aldehyde affording anti-homoallylic alcohols 8. PMID- 11112603 TI - Expeditious procedure to synthesize ethers and esters of tri- and Tetrahydroxy[6]helicenebisquinones from the dye-intermediates disodium 4-hydroxy- and 4,5-dihydroxynaphthalene-2,7-disulfonates. AB - A procedure is described for synthesizing appreciable quantities of both the tetradodecyloxy[6]helicenebisquinone 1 (R = dodecyl), which exhibits unique optical properties but previously was difficult to prepare, and a variety of analogues. The synthesis starts from disodium 4,5-dihydroxynaphthalene-2,7 disulfonate, the commercially available dye-intermediate known as chromotropic acid. It gives enantiopure 1, with R = (i-Pr)(3)Si, whose silyl groups can be replaced by dodecyl and hexanoyl groups. The same procedure applied to disodium 4 hydroxynaphthalene-2,7-disulfonate, also an inexpensive, commercially available chemical, works equally well to produce the corresponding molecules that have one fewer side chain. Key steps are the use of tosyl groups to protect phenols and of a method described seven years ago by Satoh, Itoh, Miura, and Nomura to transform the sulfonic acid functions to iodides. The structure of tetra-(1S)-camphanate 20, the ester of the reduction product of (-)-1 [R = (i-Pr)(3)Si], was analyzed by X-ray diffraction. It shows the absolute configurations and supports the presumed basis for the rule that the (1S)-camphanates of (P)-helicen-1-ols are more polar than their (M)-diastereomers. PMID- 11112604 TI - Easy access to 3,8-Diaryldifurano[2,3-a:2',3'-f]naphthalenes. A new extended aromatic system. AB - 3,8-Diaryldifurano[2,3-a:2',3'-f]naphthalenes were prepared in two simple steps. First, 1,5-dihydroxynaphthalene and 1-aryl-2-bromodecan-1-ones were condensed to the corresponding naphthalene 1,5-diethers. Second, these intermediates were cyclized using methanesulfonic acid in methylene chloride. Seven examples are given, three of which are doubly substituted with octyl chains to enhance the solubility. This increased solubility allowed further modification of the 3,8 aryl groups to attach electron-withdrawing groups (formyl, nitrile, dicyanovinyl, and benzoyl). PMID- 11112605 TI - Protonated and methylated dimethyl sulfoxide cations and dications. DFT/GIAO-MP2 NMR studies and comparison with experimental data. AB - Energies, electronic structures, and thermodynamics of protonated and methylated dimethyl sulfoxide (DMSO) cations and dications were calculated using the density functional theory (DFT) method. The O-protonated structure 2 was found to be 37.0 kcal/mol more stable than the S-protonated 3. For diprotonated DMSO dication, the O, O-diprotonated form 6 was found to be the global minimum, more stable by 20.8 kcal/mol than O,S-diprotonated 7. Interestingly, for dimethylated DMSO dication, O,O-dimethylated 11 and O,S-dimethylated 12 are isoenergetic. (13)C, (17)O, and (33)S NMR chemical shifts of the cations and dications were calculated using the GIAO-MP2 method and compared with the available experimental data. PMID- 11112606 TI - Wavelength-selective photodenitrogenation of azoalkanes to high-spin polyradicals with cyclopentane-1,3-diyl spin-carrying units and their photobleaching: EPR/UV spectroscopy and product studies of the matrix-isolated species. AB - The photolysis of the mono-, bis-, and trisazoalkanes 1, 2, and 3 in a toluene matrix at 77 K has been studied by EPR and UV spectroscopy. The purpose was to find the optimal conditions for the generation of the corresponding organic high spin polyradicals (the triplet diradicals D-1, D-2, and D-3, the tetraradicals T 2 and T-3, and the hexaradical H-3) all with localized cyclopentane-1,3-diyl spin carrying units, connected by m-phenylene (except D-1) as ferromagnetic coupler. Irradiation of these azoalkanes at 333, 351, or 364 nm gave different polyradical compositions. This observed wavelength dependence is due to the secondary photoreaction (photobleaching) of the polyradical intermediate. The photobleaching process has been examined in detail for the triplet diradical D-1, for which pi,pi excitation affords the cyclopentenes 5 instead of the housane 4 (the usual product of the diradical D-1 on warm-up of the matrix). The pi,pi excited diradical D-1 fragments into a pair of allyl and methyl radicals (the latter was observed by EPR spectroscopy of a photobleached sample), and recombination affords the cyclopentene. Similar photochemical events are proposed for the photobleaching of the tetraradical T-2 and hexaradical H-3, derived from the respective azoalkanes 2 and 3. Thus, photobleaching of the polyradicals competes effectively with their photogeneration from the azoalkane. This unavoidable event is the consequence of spectral overlap between the cumyl radical pi,pi chromophore of the polyradical and the n,pi chromophore of the azoalkane at the wavelength (364 nm), at which the latter is photoactive for the required extrusion of molecular nitrogen. PMID- 11112607 TI - Generation, characterization, and kinetics of triplet Di[1,2,3,4,5,6, 7,8 octahydro-1,4:5,8-di(ethano)anthryl]carbene. AB - The title carbene has been generated by photolysis of the corresponding diazo precursors and studied by spectroscopic means, i. e., electron paramagnetic resonance (EPR) and UV/vis spectroscopy in matrixes at low temperature and laser flash photolysis in solution at room temperature, with the product analysis. The results are compared with triplet di(2,3,5,6-tetramethylphenyl)carbene, an open chain counterpart, which revealed that bicycloalkyl groups are acting as a fairly good kinetic protector for the triplet carbene as opposed to the open-chain counterpart. The formation of all-hydrocarbon triplet carbenes having a half-life over a second under normal conditions was realized for the first time. Effects of para-substituents on the structure and reactivities of the carbene are also investigated and discussed in terms of polar and spin electronic effects. PMID- 11112608 TI - An efficient ketone-catalyzed epoxidation using hydrogen peroxide as oxidant. PMID- 11112609 TI - Preparation of new anti-tubulin ligands through a dual-mode, addition-elimination reaction to a bromo-substituted alpha, beta-unsaturated sulfoxide. PMID- 11112610 TI - Oxidation of alkynes by the HOF.CH(3)CN complex. PMID- 11112611 TI - Efficient one-Pot synthesis of polysubstituted pyrroles. PMID- 11112612 TI - Rearrangement of spiroacetals of the 1,6-Dioxaspiro[4.5]decan-10-yl methanesulfonate type. Synthesis of cis-fused 1,6-dioxadecalins. PMID- 11112613 TI - Chemistry of N,N-bis(silyloxy)enamines. 3. N,N-bis(silyloxy)enamines as beta-C nucleophiles in reaction with acetals mediated by trimethylsilyl trifluoromethanesulfonate. PMID- 11112614 TI - General base and general acid catalyzed intramolecular aminolysis of esters. Cyclization Of esters of 2-aminomethylbenzoic acid to phthalimidine PMID- 11112615 TI - 2000 division of organic chemistry fellowship awards PMID- 11112616 TI - Design and synthesis of novel [60]fullerene derivatives as potential HIV aspartic protease inhibitors. AB - [structure] Two water-soluble fullerene derivatives have been computer-designed and synthesized. They may exhibit interesting anti-HIV activity owing to the presence of two ammonium groups strategically located on the spheroid surface. PMID- 11112617 TI - A highly regioselective sonogashira coupling as a key step in the preparation of the first phenanthroline with two diverse reactive groups in 3,8-positions AB - The preparation of 3,8-unsymmetric phenanthrolines is described. Desymmetrization of 3,8-dibromophenanthroline was achieved after monoarylation followed by regioselective Pd-catalyzed monoalkynylation that was controlled by the methoxy group of the dimethoxyphenyl substituent. PMID- 11112618 TI - Column asymmetric catalysis for beta-lactam synthesis. AB - [structure] A catalytic asymmetric reaction process was designed involving the use of solid-phase reagents and catalysts that constitute the packing of a series of "reaction columns". This process was applied to the catalytic asymmetric synthesis of beta-lactams, yielding pure product after crystallization with exceptional enantio- and diastereoselectivity. PMID- 11112619 TI - A formal total synthesis of dysidiolide. AB - [structure] A formal total synthesis of the natural product dysidiolide is described. Starting from a Diels-Alder reaction between an enoate and a Rawal diene, the cyclohexenone 4 was synthesized. A subsequent stereospecific methyl cuprate addition established the desired trans configuration in the cyclohexane 3. Wacker oxidation of the pentenyl side chain to the diketone 17 followed by an intramolecular aldol condensation led to the bicyclic enone 2, a key intermediate in a recently reported synthesis of dysidiolide. PMID- 11112620 TI - Variable and stereoselective synthesis of azasugar analogues by a ruthenium catalyzed ring rearrangement AB - A novel ruthenium-catalyzed ring opening/ring closing tandem metathesis reaction with a catalytic transfer of stereocenters from a ring to an olefinic chain is described. This ring rearrangement serves as the key step in the stereoselective synthesis of the new azasugar analogues 1 and 2. PMID- 11112621 TI - Enantioselective allyltitanation. Efficient synthesis of the C1-C14 polyol subunit of amphotericin B. AB - [structure] An efficient synthesis of the C(1)-C(14) fragment of amphotericin B is described. This synthesis is based on the formation of syn-1,3-diols from enantioselective allyltitanation of unprotected beta-hydroxyaldehydes. PMID- 11112623 TI - pi-stacking interactions in cis-Bisfullerene AB - Experimental, molecular modeling, and model compound studies suggest that favorable fullerene[60] pi-stacking interactions in the ground state and in syn transition states account for the high cis stereoselectivities observed in the reactions between C(60) and 6, 13-disubstituted pentacenes. PMID- 11112622 TI - Completely regioselective, highly stereoselective syntheses of cis-Bisfullerene AB - cis-Bisfullerene[60] adducts of 6,13-disubstituted pentacenes (R = Ph, 4' hydroxymethylphenyl) are synthesized in 75% to 85% isolated yields under kinetically controlled Diels-Alder conditions. The cycloadditions are completely regioselective and highly stereoselective, with only traces of the diastereomeric trans-bisfullerene[60] adducts forming. PMID- 11112624 TI - Oligomers of enantiopure bicyclic gamma/delta-amino acids (BTAa). 1. Synthesis and conformational analysis of 3-aza-6,8-dioxabicyclo AB - A series of dimeric through pentameric oligomers of a bicyclic gamma/delta-amino acid (BTG) were synthesized using peptide coupling methods in solution with PyBroP or HATU. The analysis of (1)H NMR and CD spectra suggests that these oligomers could have a partially ordered structure in alcohol solutions. PMID- 11112625 TI - An EPR investigation of persistent radicals from the photolysis of p, p'-dialkyl substituted phenyl benzyl ketones adsorbed on MFI zeolites AB - The photolysis of isomeric pairs of p,p'-dialkyl-substituted phenyl benzyl ketones adsorbed on MFI zeolites has been investigated by EPR spectroscopy. Photolysis produces persistent "benzoyl type" and "benzyl type" radicals. The dominant persistent radical produced by photolysis of any particular isomeric pair depends on the length and position of the p-alkyl chain. The results are attributed to supramolecular stereoisomers resulting from preferential adsorption of the longer alkyl chain into the pores of the zeolite. PMID- 11112626 TI - 2'-O-[2-[N,N-(dialkyl)aminooxy]ethyl]-modified antisense oligonucleotides. AB - [structure] Oligonucleotides with two novel modifications, 2'-O-?2-[N, N (dimethyl)aminooxy]ethyl? (2'-O-DMAOE) and 2'-O-?2-[N, N-(diethyl)aminooxy]ethyl? (2'-O-DEAOE), have been synthesized. These modifications exhibit high binding affinity to target RNA (and not to DNA) and enhance the nuclease stability of oligonucleotides considerably with t(1/2) > 24 h as a phosphodiester. PMID- 11112627 TI - A modular synthetic approach toward exhaustively stereodiversified ligand libraries. AB - [structure] This report describes a modular approach to the synthesis of stereodiversified natural product-like libraries. Monomers 2 and 3 were coupled in parallel by silyl-tethered olefin metathesis to generate all 16 stereoisomers of cis-enediols 1. All 16 stereoisomers were incorporated into chimerae having flanking peptidic segments. These chimerae exhibited a broad range of hydrophobicities, raising the possibility that stereochemical variation might be used to tune the pharmacologic properties of small molecules. PMID- 11112628 TI - Photochemical cycloaddition reagents for rigidly attaching the 1, 4 dimethoxynaphthalene chromophore to scaffold alkenes AB - The norbornanecyclobutene epoxides 1a-1c containing a fused 1, 4 dimethoxynaphthalene chromophore have been reacted with cyclobutenes, cyclohexenes, norbornenes, 7-isopropylidenenorbornenes, 7-azanorbornenes, and other cyclic or electron-deficient alkenes at room temperature to form 1:1 adducts in stereoselective 1,3-dipolar cycloaddition reactions; alkynes can also participate in this reaction. The ability to form 2:1 adducts has also been demonstrated, thereby opening up opportunities for preparing functionalized products with large chromophore separations. PMID- 11112629 TI - Methodology for regioselective synthesis of substituted pyridines via intramolecular oximino malonate hetero Diels-Alder reactions. AB - [structure] Various substituted pyridines can be prepared regioselectively by a sequence involving an intramolecular thermal or high-pressure Diels-Alder cycloaddition of an oximino malonate dienophile tethered to a dienic carboxylic acid, followed by mild aromatization of the resulting cycloadduct with cesium carbonate in DMF at room temperature. PMID- 11112630 TI - Titanium(IV) alkoxide ligand exchange with alpha-hydroxy acids: the enantioselective aldol addition AB - Ligand exchange of titanium(IV) alkoxides with alpha-hydroxy acids presents an unexpected and novel approach to enantioselective aldol additions of aldehydes and ketones. The aldol products were isolated in a high degree of syn diastereoselectivity. High enantioselectivities were observed by using simple optically pure alpha-hydroxy acids in this novel aldol addition. PMID- 11112631 TI - Asymmetric synthesis of quaternary centers. Total synthesis of (-)-malyngolide. AB - [structure] The deracemization of 3-nonyl-3,4-epoxybut-1-ene with Pd(0) in the presence of chiral ligands using p-methoxybenzyl alcohol as a nucleophile proceeds regio- and enantioselectively to form the monoprotected vinylglycidol in 99% ee. This chiral building block was converted in seven steps to (-) malyngolide, an antibiotic showing significant activity against Mycobacterium smegmatis and Streptococcus pyogenes. An interesting aspect involves controlling the diastereoselectivity of protonation of an enolate via a distal hydroxyl group. PMID- 11112632 TI - all-Z-Tetrabenzo AB - Crystal structures of the silver complexes derived from tetrabenzo[16]annulene with AgOTf and AgClO(4) are different, although these two complexes show similar (1)H NMR spectra reflecting a similar clathrate structure in solution. The silver complex of pentabenzo[20]annulene with AgClO(4) adopts a clathrate structure both in the solid state and in solution. PMID- 11112633 TI - A new photoactive and highly soluble C(60)-TTF-C(60) dimer: charge separation and recombination AB - The covalent linkage of two [60]fullerene cores to a tetrathiafulvalene (TTF) donor affords a soluble and photoactive C(60)-TTF-C(60) triad. Spectroscopic and photophysical characterization of the C(60)-TTF-C(60) triad are given. Although the cyclic voltammetry measurements reveal no notable interaction between the chromophores in the ground state, photophysical data show that in the excited state an intramolecular electron transfer, evolving from the TTF donor to the singlet state of C(60), prevails, yielding a long-lived charge separated radical pair. PMID- 11112634 TI - Enzyme-catalyzed asymmetric deacylation for the preparation of lasofoxifene (CP 336156), a selective estrogen receptor modulator. AB - [structure] selective estrogen receptor modulator (SERM), Lasofoxifene (CP 336156), was prepared by an enzyme-catalyzed asymmetric deacylation with high optical purity and excellent yield even though the hydrolytic site is remote from the chiral centers. PMID- 11112635 TI - Efficient conversion of vicinal diols to alkenes by treatment of the corresponding dimesylates with a catalytic, minimally fluorous, recoverable diaryl diselenide and sodium borohydride AB - In conjunction with sodium borohydride as stoichiometric reagent a catalytic quantity of bis(4-perfluorohexylphenyl) diselenide converts vicinal dimesylates to the corresponding alkenes in good yield on warming in ethanol. The diselenide is recovered in high yield by continuous fluorous extraction. PMID- 11112636 TI - Enantioselective synthesis of the papulacandin ring system: conversion of the mannose diastereoisomer into a glucose stereoisomer. AB - [structure] An enantioselective synthesis of three diastereoisomers of the C arylglycoside tricyclic spiroketal nucleus of the papulacandins has been achieved, in which the initial asymmetry was introduced via a Sharpless dihydroxylation of substituted 5-aryl-2-vinylfurans. A selective oxidation reduction sequence converted the mannose isomer into the glucose isomer. This sequence can conveniently produce both the papulacandin ring system along with its enantiomer and diastereomers in only 10-14 steps from 3,5-dimethoxybenzyl alcohol in 5-8% overall yield. PMID- 11112637 TI - Synthesis of the sialidase inhibitor siastatin B. AB - [structure] The resolved piperidinecarboxylate (R)-7 was converted to siastatin B (1) by an efficient and stereoselective sequence that includes a bromo-beta lactonization and an N-acyliminium azidation. Two analogues (3 and 4) of siastatin were also prepared. PMID- 11112638 TI - Facile and efficient total synthesis of (+)-preussin. AB - [structure] The enantioselective total synthesis of (+)-preussin, a potent antifungal agent, has been achieved. The key steps are a Pd(0)-catalyzed oxazoline-forming reaction from L-phenylalanine, hydrogenolysis, and subsequent diastereoselective reductive cyclization of the intermediate aminoketone to pyrrolidine using Pearlman's catalyst. PMID- 11112639 TI - The cytosporones, new octaketide antibiotics isolated from an endophytic fungus. AB - [structure] Organic extracts from cultures of endophytic fungi collected in the Guanacaste Conservation Area of Costa Rica were screened for antibiotic activity. Two endophytes CR200 (Cytospora sp.) and CR146 (Diaporthe sp.) were found to have potent antibiotic activity. Bioassay-guided fractionation of the extracts from these fungi led to the identification of cytosporones D and E, antibacterial active trihydroxybenzene lactones, and three related but inactive metabolites. The five new octaketides were characterized using X-ray crystallography and NMR. PMID- 11112640 TI - Cytoskyrins A and B, new BIA active bisanthraquinones isolated from an endophytic fungus. AB - [structure] The biochemical induction assay (BIA) is a rapid (colorimetric) bacterial assay used to identify compounds that damage DNA or inhibit DNA synthesis and thereby identify potential natural product anticancer agents. Bisanthraquinones based on a 1,3,6, 8-tetrahydroxyanthraquinone-type carbon skeleton were isolated from an endophytic fungus and characterized by NMR and X ray crystallography. Cytoskyrin A (1) is highly active in the biochemical induction assay, while the closely related cytoskyrin B (2) has no detectable activity in this assay. PMID- 11112641 TI - Radical-mediated synthesis of alpha-C-glycosides based on N-acyl galactosamine. AB - [structure] C-Glycosides of N-acyl 2-amino-2-deoxygalactose (acyl = MeCO, CF(3)CO, t-BuOCO) are available in a stereoselective manner by trapping of an anomeric radical with an activated alkene. Using anomeric selenides, radical generation and trapping is carried out under conditions that avoid competitive reduction, and this chemistry has been applied to the synthesis of the novel C glycoside analogue of O-benzyl alpha-D-GalNAc. PMID- 11112642 TI - Synthetic studies on the trans-chlorocyclopropane dienyne side chain of callipeltoside A. AB - [structure] Enantiomerically enriched trans-chlorocyclopropanemethanol was obtained by lipase kinetic resolution of dichlorocyclopropanemethanol 3, followed by reduction. The sp-sp(2) bond of the trans-chlorocyclopropane dienyne side chain of callipeltoside A was constructed via a Stille coupling reaction of 1, 1 dibromo-1-alkene 7 and a vinylstannane in a highly dipolar solvent capable of promoting HBr elimination to give internal alkynes. PMID- 11112643 TI - Chiral Palladium(II)-catalyzed asymmetric glyoxylate-Ene reaction: alternative approach to the enantioselective synthesis of alpha-hydroxy esters AB - An effective chiral palladium catalyst [Pd(CH(3)CN)(2)(S)-Tol-BINAP](SbF(6))(2) (2b) is developed for asymmetric glyoxylate-ene reactions. This palladium dicationic catalyst provides a simple but efficient approach to the asymmetric synthesis of alpha-hydroxy esters in excellent yields with high enantioselectivities at relatively higher reaction temperature (60 degrees C). PMID- 11112644 TI - 2H-Chromenes from salicylaldehydes by a catalytic petasis reaction AB - The Petasis condensation of vinylic or aromatic boronic acids, aromatic aldehydes, and amines is assisted by a hydroxy group adjacent to the aldehyde moiety. The products derived from salicylaldehydes and vinylboronic acids undergo cyclization to 2H-chromene compounds with ejection of amine upon heating. A catalytic preparation of 2H-chromenes using resin-bound amine is reported, allowing the convenient incorporation of a variety of components. PMID- 11112645 TI - S-(4-methoxyphenyl) benzenethiosulfinate (MPBT)/trifluoromethanesulfonic anhydride: a convenient system for the generation of glycosyl triflates from thioglycosides. AB - [structure] The combination of S-(4-methoxyphenyl) benzenethiosulfinate (MPBT, 1) and trifluoromethanesulfonic anhydride forms a powerful, metal-free, thiophile which readily activates thioglycosides, via glycosyl triflates, at -60 degrees C in dichloromethane, in the presence of 2,6-di-tert-butyl-4-methylpyridine. The glycosyl triflates are rapidly and cleanly converted to glycosides, upon treatment with alcohols, in good yield and selectivity. PMID- 11112646 TI - A new alternative to the mannich reaction: tandem radical addition-cyclization reaction for asymmetric synthesis of gamma-butyrolactones and beta-amino acids AB - The radical addition-cyclization reaction of substrates having two different radical acceptors such as acrylate and aldoxime ether moieties was studied. This new free radical-mediated Mannich-type reaction proceeded smoothly via a tandem C C bond-forming process. Furthermore, the diastereoselective tandem reaction provides the novel method for asymmetric synthesis of gamma-butyrolactones and beta-amino acid derivatives. PMID- 11112647 TI - Simply assembled and recyclable polymer-supported olefin metathesis catalysts AB - Polymer-supported ruthenium catalysts (PCy(3))(2)Ru(=C(H)Ph)Cl(2), (PCy(3))Ru(IMes)(=C(H)Ph)Cl(2), and (PCy(3))Ru(SIMes)(=C(H)Ph)Cl(2), where IMes = 1,3-bis(2,4,6-trimethylphenyl)imidazol-2-ylidene and SIMes = 1,3-bis(2,4,6 trimethylphenyl)-4,5-dihydroimidazol-2-ylidene, have been prepared and found to be effective "boomerang" catalysts for ring-closing metathesis. They are recyclable, show comparable or better reactivity than their homogeneous counterparts, tolerate functional groups, and perform very well with dienes and moderately well with highly hindered substrates. PMID- 11112648 TI - Novel carbon-carbon bond formation reaction of methoxyallene oxide promoted by TiI(4) AB - Methoxyallene oxide was readily prepared in situ by epoxidation of methoxyallene with 3-chloroperbenzoic acid, and the subsequent reaction with aldehydes or acetals was promoted by titanium tetraiodide and additives to give 2,3-dialkoxy- or 3-hydroxy-2-methoxy ketones in good yields. PMID- 11112650 TI - A regio- and stereodivergent synthesis of vic-amino alcohols AB - A regio- and stereodivergent synthesis of vic-amino alcohols starting from vinylepoxides is described. The developed strategy focuses on the propensity of vinylepoxides and vinylaziridines to be ring-opened at the allylic position by suitable nucleophiles and makes use of reactions that perform such tasks selectively with either retention or inversion of configuration. PMID- 11112649 TI - Enantiocontrolled synthesis of spirooxindoles based on the [5 + 2] cycloaddition of a Tp(CO)(2)Mo(pyridinyl) scaffold (Tp = hydridotrispyrazolylborate). AB - [structure] A [5 + 2] cycloaddition of the pyridinyl pi-complex (-)-1 (98% ee) to methyleneoxindole 2 afforded the spirooxindole complex (-)-3 in high enantiomeric purity. Complex (-)-3 was converted to pyrrolidine (-)-8 (97% ee), which is related to potent cytotoxic analogues of the spirotryprostatins alkaloids. PMID- 11112651 TI - A one-flask synthesis of Weinreb amides from chiral and achiral carboxylic acids using the deoxo-fluor fluorinating reagent. AB - [structure] The reagent [bis(2-methoxyethyl)amino]sulfur trifluoride (Deoxo-Fluor reagent) converts carboxylic acids to the corresponding acid fluorides, which then react with N,N-dimethylhydroxylamine to give the corresponding Weinreb amides in high yields. The reaction proceeds without racemization when optically active acids are used as the starting material. This method is operationally simple and provides the products in high purity. PMID- 11112652 TI - Convergent regiodirected assembly of 2,3-disubstituted furans AB - The conjugate addition of organocopper reagents to alpha, beta-unsaturated enones results in the regiospecific generation of enolate anions, which can be made to undergo the aldol reaction with (tetrahydropyranyloxy)acetaldehyde under zinc chloride catalysis. Treatment of the resulting product with p-toluenesulfonic acid in THF affords the targeted 2,3-disubstituted furan. PMID- 11112653 TI - Two general routes to 1,4-disubstituted-2,3,4,5-tetrahydro- 1H-3-benzazepines. AB - [structure] Two general routes to 1,4-disubstituted-2,3,4, 5-tetrahydro-1H-3 benzazepines are described. Both routes utilize an appropriately functionalized phenethylamino alcohol as the penultimate intermediate: the first route makes use of the reductive amination of a benzyl alkyl ketone with alpha (aminomethyl)benzyl alcohol, while the second route utilizes the addition of a Grignard reagent to the oxazolidine derived from a substitued phenylacetaldehyde and alpha-(methylaminomethyl)benzyl alcohol. In all cases studied, the cis-1,4 disubstituted-2,3,4, 5-tetrahydro-1H-3-benzazepine was obtained as the major product. PMID- 11112654 TI - Total syntheses of (-)-fumiquinazolines A, B, and I. AB - [structure] The first total syntheses of (-)-fumiquinazolines A, B, and I have been accomplished efficiently using the Pd-catalyzed cyclization of an iodoindole carbamate to construct the imidazoindolone moiety and the dehydrative cyclization of a diamide followed by rearrangement through an amidine to construct the quinazolone moiety. PMID- 11112655 TI - Highly selective aziridination of imines using trimethylsilyldiazomethane and applications of C-silylaziridines in synthesis AB - Trimethylsilyldiazomethane has been found to add directly to N-sulfonyl (Ts and SES) imines to afford aziridines in good yields and high cis stereoselectivities. The silyl group can be substituted by treatment with a fluoride source and electrophiles again with high selectivity. Complete regioselectivity is observed in ring opening of these aziridines with nucleophiles. PMID- 11112657 TI - Palladium-mediated formation of bowl-shaped molecules: synthesis of as-Indaceno PMID- 11112656 TI - Deuterium isotope effects on (13)C NMR chemical shifts reflect the smaller steric size of CD(3) compared to CH(3) groups AB - A CD(3) group close in space to (but many bonds distant from) a carbon atom A causes a substituent effect on the chemical shift of C(A) that is algebraically smaller than the effect of a CH(3) group, in agreement with the notion of shorter C-D relative to C-H bonds. Hence, the deuterium isotope effect of CD(3) upon delta(C(A)) is shielding when the substituent effect is deshielding, and vice versa. PMID- 11112658 TI - Genotypic and phenotypic spectrum in tricho-rhino-phalangeal syndrome types I and III. AB - Tricho-rhino-phalangeal syndrome (TRPS) is characterized by craniofacial and skeletal abnormalities. Three subtypes have been described: TRPS I, caused by mutations in the TRPS1 gene on chromosome 8; TRPS II, a microdeletion syndrome affecting the TRPS1 and EXT1 genes; and TRPS III, a form with severe brachydactyly, due to short metacarpals, and severe short stature, but without exostoses. To investigate whether TRPS III is caused by TRPS1 mutations and to establish a genotype-phenotype correlation in TRPS, we performed extensive mutation analysis and evaluated the height and degree of brachydactyly in patients with TRPS I or TRPS III. We found 35 different mutations in 44 of 51 unrelated patients. The detection rate (86%) indicates that TRPS1 is the major locus for TRPS I and TRPS III. We did not find any mutation in the parents of sporadic patients or in apparently healthy relatives of familial patients, indicating complete penetrance of TRPS1 mutations. Evaluation of skeletal abnormalities of patients with TRPS1 mutations revealed a wide clinical spectrum. The phenotype was variable in unrelated, age- and sex-matched patients with identical mutations, as well as in families. Four of the five missense mutations alter the GATA DNA-binding zinc finger, and six of the seven unrelated patients with these mutations may be classified as having TRPS III. Our data indicate that TRPS III is at the severe end of the TRPS spectrum and that it is most often caused by a specific class of mutations in the TRPS1 gene. PMID- 11112659 TI - Primate DAX1, SRY, and SOX9: evolutionary stratification of sex-determination pathway. AB - The molecular evolution of DAX1, SRY, and SOX9, genes involved in mammalian sex determination, was examined in six primate species. DAX1 and SRY have been added to the X and Y chromosomes, respectively, during mammalian evolution, whereas SOX9 remains autosomal. We determined the genomic sequences of DAX1, SRY, and SOX9 in all six species, and calculated K(a), the number of nonsynonymous substitutions per nonsynonymous site, and compared this with the K(s), the number of synonymous substitutions per synonymous site. Phylogenetic trees were constructed by means of the DAX1, SRY, and SOX9 coding sequences, and phylogenetic analysis was performed using maximum likelihood. Overall measures of gene and protein similarity were closer for DAX1 and SOX9, but DAX1 exhibited nonsynonymous amino acid substitutions at an accelerated frequency relative to synonymous changes, similar to SRY and significantly higher than SOX9. We conclude that, at the protein level, DAX1 and SRY are under less selective pressure to remain conserved than SOX9, and, therefore, diverge more across species than does SOX9. These results are consistent with evolutionary stratification of the mammalian sex determination pathway, analogous to that for sex chromosomes. PMID- 11112660 TI - A second locus for an axonal form of autosomal recessive Charcot-Marie-Tooth disease maps to chromosome 19q13.3. AB - Autosomal recessive Charcot-Marie-Tooth disease (CMT) represents a heterogeneous group of disorders affecting the peripheral nervous system. The axonal form of the disease is designated as "CMT type 2" (CMT2), and one locus (1q21.2-q21.3) has been reported for the autosomal recessive form. Here we report the results of a genomewide search in an inbred Costa Rican family (CR-1) affected with autosomal recessive CMT2. By analyzing three branches of the family we detected linkage to the 19q13.3 region, and subsequent homozygosity mapping defined shared haplotypes between markers D19S902 and D19S907 in a 5.5-cM range. A maximum two point LOD score of 9.08 was obtained for marker D19S867, at a recombination fraction of.00, which strongly supports linkage to this locus. The epithelial membrane protein 3 gene, encoding a PMP22 homologous protein and located on 19q13.3, was ruled out as being responsible for this form of CMT. The age at onset of chronic symmetric sensory-motor polyneuropathy was 28-42 years (mean 33.8 years); the electrophysiological data clearly reflect an axonal degenerative process. The phenotype and locus are different from those of demyelinating CMT4F, recently mapped to 19q13.1-13.3; hence, the disease affecting the Costa Rican family constitutes an axonal, autosomal recessive CMT subtype (ARCMT2B). PMID- 11112661 TI - Population structure in admixed populations: effect of admixture dynamics on the pattern of linkage disequilibrium. AB - Gene flow between genetically distinct populations creates linkage disequilibrium (admixture linkage disequilibrium [ALD]) among all loci (linked and unlinked) that have different allele frequencies in the founding populations. We have explored the distribution of ALD by using computer simulation of two extreme models of admixture: the hybrid-isolation (HI) model, in which admixture occurs in a single generation, and the continuous-gene-flow (CGF) model, in which admixture occurs at a steady rate in every generation. Linkage disequilibrium patterns in African American population samples from Jackson, MS, and from coastal South Carolina resemble patterns observed in the simulated CGF populations, in two respects. First, significant association between two loci (FY and AT3) separated by 22 cM was detected in both samples. The retention of ALD over relatively large (>10 cM) chromosomal segments is characteristic of a CGF pattern of admixture but not of an HI pattern. Second, significant associations were also detected between many pairs of unlinked loci, as observed in the CGF simulation results but not in the simulated HI populations. Such a high rate of association between unlinked markers in these populations could result in false positive linkage signals in an admixture-mapping study. However, we demonstrate that by conditioning on parental admixture, we can distinguish between true linkage and association resulting from shared ancestry. Therefore, populations with a CGF history of admixture not only are appropriate for admixture mapping but also have greater power for detection of linkage disequilibrium over large chromosomal regions than do populations that have experienced a pattern of admixture more similar to the HI model, if methods are employed that detect and adjust for disequilibrium caused by continuous admixture. PMID- 11112662 TI - A narrow segment of maternal uniparental disomy of chromosome 7q31-qter in Silver Russell syndrome delimits a candidate gene region. AB - Maternal uniparental disomy of chromosome 7 (matUPD7), the inheritance of both chromosomes from only the mother, is observed in approximately 10% of patients with Silver-Russell syndrome (SRS). It has been suggested that at least one imprinted gene that regulates growth and development resides on human chromosome 7. To date, three imprinted genes-PEG1/MEST, gamma2-COP, and GRB10-have been identified on chromosome 7, but their role in the etiology of SRS remains uncertain. In a systematic screening with microsatellite markers, for matUPD7 cases among patients with SRS, we identified a patient who had a small segment of matUPD7 and biparental inheritance of the remainder of chromosome 7. Such a pattern may be explained by somatic recombination in the zygote. The matUPD7 segment at 7q31-qter extends for 35 Mb and includes the imprinted gene cluster of PEG1/MEST and gamma2-COP at 7q32. GRB10 at 7p11.2-p12 is located within a region of biparental inheritance. Although partial UPD has previously been reported for chromosomes 6, 11, 14, and 15, this is the first report of a patient with SRS who has segmental matUPD7. Our findings delimit a candidate imprinted region sufficient to cause SRS. PMID- 11112663 TI - Hereditary nonpolyposis colorectal cancer in 95 families: differences and similarities between mutation-positive and mutation-negative kindreds. AB - Hereditary nonpolyposis colorectal cancer (HNPCC) describes the condition of a disparate group of families that have in common a predisposition to colorectal cancer in the absence of a premalignant phenotype. The genetic basis of this disease has been linked to mutations in genes associated with DNA mismatch repair. A large proportion of families harbor changes in one of two genes, hMSH2 and hMLH1. Approximately 35% of families in which the diagnosis is based on the Amsterdam criteria do not appear to harbor mutations in DNA-mismatch-repair genes. In this report we present data from a large series of families with HNPCC and indicate that there are subtle differences between families that harbor germline changes in hMSH2 and families that harbor hMLH1 mutations. Furthermore, there are differences between the mutation-positive group (hMSH2 and hMLH1 combined) of families and the mutation-negative group of families. The major findings identified in this study focus primarily on the extracolonic disease profile observed between the mutation-positive families and the mutation-negative families. Breast cancer was not significantly overrepresented in the hMSH2 mutation-positive group but was overrepresented in the hMLH1 mutation-positive group and in the mutation-negative group. Prostate cancer was not overrepresented in the mutation-positive groups but was overrepresented in the mutation-negative group. In age at diagnosis of colorectal cancer, there was no difference between the hMSH2 mutation-positive group and the hMLH1 mutation-positive group, but there was a significant difference between these two groups and the mutation negative group. PMID- 11112664 TI - Gene preference in maple syrup urine disease. AB - Untreated maple syrup urine disease (MSUD) results in mental and physical disabilities and often leads to neonatal death. Newborn-screening programs, coupled with the use of protein-modified diets, have minimized the severity of this phenotype and allowed affected individuals to develop into productive adults. Although inheritance of MSUD adheres to rules for single-gene traits, mutations in the genes for E1alpha, E1beta, or E2 of the mitochondrial branched chain alpha-ketoacid dehydrogenase complex can cause the disease. Randomly selected cell lines from 63 individuals with clinically diagnosed MSUD were tested by retroviral complementation of branched-chain alpha-ketoacid dehydrogenase activity to identify the gene locus for mutant alleles. The frequencies of the mutations were 33% for the E1alpha gene, 38% for the E1beta gene, and 19% for the E2 gene. Ten percent of the tested cell lines gave ambiguous results by showing no complementation or restoration of activity with two gene products. These results provide a means to establish a genotype/phenotype relationship in MSUD, with the ultimate goal of unraveling the complexity of this single-gene trait. This represents the largest study to date providing information on the genotype for MSUD. PMID- 11112666 TI - Successful treatment of enterovirus infection with the use of pleconaril in 2 infants with severe combined immunodeficiency. AB - Two patients with severe combined immunodeficiency and enterovirus infections were successfully treated with pleconaril. There were no adverse affects. PMID- 11112665 TI - Mutational analysis in a cohort of 224 tuberous sclerosis patients indicates increased severity of TSC2, compared with TSC1, disease in multiple organs. AB - Tuberous sclerosis (TSC) is a relatively common hamartoma syndrome caused by mutations in either of two genes, TSC1 and TSC2. Here we report comprehensive mutation analysis in 224 index patients with TSC and correlate mutation findings with clinical features. Denaturing high-performance liquid chromatography, long range polymerase chain reaction (PCR), and quantitative PCR were used for mutation detection. Mutations were identified in 186 (83%) of 224 of cases, comprising 138 small TSC2 mutations, 20 large TSC2 mutations, and 28 small TSC1 mutations. A standardized clinical assessment instrument covering 16 TSC manifestations was used. Sporadic patients with TSC1 mutations had, on average, milder disease in comparison with patients with TSC2 mutations, despite being of similar age. They had a lower frequency of seizures and moderate-to-severe mental retardation, fewer subependymal nodules and cortical tubers, less-severe kidney involvement, no retinal hamartomas, and less-severe facial angiofibroma. Patients in whom no mutation was found also had disease that was milder, on average, than that in patients with TSC2 mutations and was somewhat distinct from patients with TSC1 mutations. Although there was overlap in the spectrum of many clinical features of patients with TSC1 versus TSC2 mutations, some features (grade 2-4 kidney cysts or angiomyolipomas, forehead plaques, retinal hamartomas, and liver angiomyolipomas) were very rare or not seen at all in TSC1 patients. Thus both germline and somatic mutations appear to be less common in TSC1 than in TSC2. The reduced severity of disease in patients without defined mutations suggests that many of these patients are mosaic for a TSC2 mutation and/or have TSC because of mutations in an as-yet-unidentified locus with a relatively mild clinical phenotype. PMID- 11112667 TI - Comparison of amplicor, in-house polymerase chain reaction, and conventional culture for the diagnosis of tuberculosis in children. AB - A total of 251 clinical specimens (235 gastric aspirates and 16 bronchoalveolar lavages) from 88 children were prospectively tested in a blinded manner for the presence of Mycobacterium tuberculosis complex, by use of the Amplicor M. tuberculosis test and by means of in-house polymerase chain reaction (PCR). The results were compared with those obtained by conventional culture and by direct microscopy. All of the children underwent extended follow-up to verify or exclude the clinical diagnosis of tuberculosis. The results of the different tests, when compared to the final clinical diagnosis, were a sensitivity of 60% and a specificity of 96.8% for in-house PCR, 44% and 93.7% respectively for the Amplicor test, 44% and 100% for mycobacterial culture and 12% and 100% for microscopy. Amplicor tests presented false-positive findings in children without tuberculous infection. We conclude that both in-house PCR and the Amplicor test are rapid methods that can be helpful for difficult or urgent diagnosis of tuberculosis in children. However, efforts should be aimed toward improvement of the sensitivity and specificity of an easy-to-use PCR kit. PMID- 11112668 TI - Spinal epidural abscesses in children: a 15-year experience and review of the literature. AB - We reviewed medical records and laboratory and diagnostic evaluations for 8 pediatric patients with spinal epidural abscesses who were treated during the last 15 years at our institution. Staphylococcus aureus was isolated from 5 of 8 epidural abscesses, including 2 abscesses with methicillin-resistant S. aureus. Unusual isolates were group B Streptococcus in a patient with chronic vesicouretral reflux associated with the posterior urethral valves and Aspergillus flavus in a patient with acute myelogenous leukemia. An analysis incorporating our results and a review of the English-language literature about abscesses in children and adults revealed differences related to age. Abscesses in children were more posterior in epidural location, had greater spinal column extension, and were associated with more favorable clinical outcomes than were abscesses in adults. Magnetic resonance imaging is the diagnostic procedure of choice; however, radionuclide bone scans should be considered for associated distant osteomyelitis in children. Prompt diagnosis and combined medical and surgical treatment remain the cornerstones for the prevention of adverse outcomes. PMID- 11112669 TI - Travel-associated Burkholderia pseudomallei infection (Melioidosis) in a patient with cystic fibrosis: a case report. AB - In September 1997, a 25-year-old Italian woman with cystic fibrosis (CF) spent 3 weeks in Thailand. In August 1998, her pulmonary function rapidly declined, with productive cough and intermittent fever. Chest x-ray films revealed diffuse, small, patchy opacities in the upper lobes. Burkholderia pseudomallei (BP) was isolated from specimens of the patient's sputum and was identified by means of 16S rDNA sequencing. The diagnosis of melioidosis was serologically confirmed. Continuous therapy with ceftazidime and co-trimoxazole and maintenance with co trimoxazole, doxycycline, and chloramphenicol resulted in eradication of BP. We present the issue of whether patients with CF represent a population particularly at risk for melioidosis. PMID- 11112670 TI - Subacute sclerosing panencephalitis in an American-born adult. AB - We describe a case of an adult born in the United States who had subacute sclerosing panencephalitis (SSPE). We discuss the possibility that the patient contracted subclinical measles during the 1989-1991 measles epidemic in the United States. PMID- 11112671 TI - Bartonella infection associated with systemic juvenile rheumatoid arthritis. AB - A 4-year-old girl with systemic juvenile rheumatoid arthritis had Bartonella infection diagnosed serologically. This case suggested that Bartonella (most probably Bartonella henselae) infection may in part be responsible for the development of systemic juvenile rheumatoid arthritis. PMID- 11112672 TI - High frequency of serious infections in patients with panhypopituitarism: a case control study. AB - We reviewed the records of 65 patients with panhypopituitarism (PHP) for the frequency and types of infections requiring hospitalization, and documented serious infections in 13 of 65 patients with PHP. The increased frequency of serious infectious diseases in patients with PHP is likely to contribute to increased age-specific mortality. PMID- 11112673 TI - Burden of meningitis and other severe bacterial infections of children in africa: implications for prevention. AB - Apart from meningococcal disease in the sub-Saharan meningitis belt, the incidence and impact of life-threatening bacterial diseases in children across Africa have not been quantified. The clinical and epidemiological data on pneumococcal, Haemophilus influenzae type b (Hib), and other forms of bacterial meningitis, as well as data on other severe bacterial infections throughout the continent were scrutinized. Pneumococci were the leading causative agents of nonepidemic meningitis and other bacteremic diseases, followed by Hib. Meningococcal diseases were less common. Mortality rates associated with pneumococcal, Hib, and meningococcal meningitis were 549 (45%) of 1211 patients, 389 (29%) of 1352 patients, and 104 (8%) of 1236 patients, respectively; sequelae occurred in 50%, 40%, and 10% of cases. At 0-4 years of age, the estimated incidences of Hib meningitis and all classic Hib diseases were 70 and 100 cases per 100,000 population per year, accounting for approximately 90,000 and 120,000 cases per year, respectively. Including older age groups and, especially, nonbacteremic Hib pneumonia in the estimates of Hib disease in Africa increased the overall numbers manifold; the numbers of pneumococcal infections were even greater. The only realistic way to combat these severe infections efficaciously would be through widespread vaccination, starting with Hib conjugates. PMID- 11112674 TI - Failure of cidofovir therapy in progressive multifocal leukoencephalopathy unrelated to human immunodeficiency virus. AB - We describe the first reported human immunodeficiency virus (HIV)-seronegative patient treated with cidofovir for progressive multifocal leukoencephalopathy (PML). Marked clinical and radiological progression of PML occurred during cidofovir therapy. The improvement observed during cidofovir therapy of HIV infected patients may be due to the effect of concomitant antiretroviral therapy rather than cidofovir. PMID- 11112675 TI - High rate of tuberculosis reinfection during a nosocomial outbreak of multidrug resistant tuberculosis caused by Mycobacterium bovis strain B. AB - We present a study of a nosocomial outbreak of multidrug-resistant tuberculosis caused by Mycobacterium bovis in 31 patients, 30 of whom were infected with human immunodeficiency virus; all 31 died of progressive tuberculosis. All M. bovis strains had identical spoligotyping patterns and showed resistance to 12 antituberculosis drugs. Reinfection was suggested in 11 cases and confirmed in 4 by molecular typing methods. The causative strain was named "B strain." PMID- 11112676 TI - Vancomycin-resistant enterococci among chronic hemodialysis patients: a prospective study of acquisition. AB - To determine the prevalence and rate of acquisition of vancomycin-resistant enterococci (VRE) among patients undergoing chronic (i.e., long-term) hemodialysis who were admitted to a tertiary care center, serial rectal cultures for VRE were performed at hospital admission and every 5 days until hospital discharge. A total of 7 (6%) of the 119 patients were colonized with VRE at admission. Six (19%) of the 32 patients who remained in the hospital > or =4 days acquired VRE. A nonambulatory status was significantly associated with colonization at admission (OR, 9.7; 95% CI, 1.8-53; P=.01), and vancomycin exposure was significantly associated with VRE acquisition (relative risk, 1.8; 95% CI, 1.1-2.9; P=.02). All patients acquired VRE from epidemiologically linked dialysis patients colonized with similar VRE genotypes. Hospital acquisition of VRE contributes substantially to the increasing prevalence of VRE in the chronic hemodialysis patient population. PMID- 11112677 TI - Impact of fluoroquinolone administration on the emergence of fluoroquinolone resistant gram-negative bacilli from gastrointestinal flora. AB - We assessed the risk factors for acquisition of fluoroquinolone-resistant, gram negative organisms in the gastrointestinal tract of hospitalized patients. We analyzed stool samples from 204 patients and recovered fluoroquinolone-resistant, gram-negative organisms from 63. Receipt of fluoroquinolone during the month preceding admission was the only risk factor identified, whereas female sex, duration of hospitalization, exposure to indwelling devices, admission from another hospital, and history of infection were risk factors for fecal colonization after day 4. PMID- 11112678 TI - Oral cefixime is similar to continued intravenous antibiotics in the empirical treatment of febrile neutropenic children with cancer. AB - Empiric oral antibiotic therapy for febrile neutropenic cancer patients has been suggested as a means to decrease hospitalization, but the safety of this approach has not been adequately studied in children. We compared continued iv antibiotic therapy with switching treatment to orally administered cefixime in a group of selected febrile neutropenic children for whom blood cultures were sterile after 48 h of incubation. Two hundred episodes of febrile neutropenia were studied (156 patients), and 100 episodes were randomized to receive each treatment. Failure to respond to therapy was defined by documented or suspected bacterial infection, recurrent fever, or discontinuation of assigned therapy for any reason before neutropenia resolved. Rates of treatment failure were similar in the oral cefixime group (28%) and in the iv antibiotic group (27%; P=1.0). Results support the safety of oral cefixime therapy for low-risk febrile neutropenic children, a therapeutic approach that would facilitate earlier outpatient management and decrease the costs of treatment. PMID- 11112679 TI - Dancing pointers, preachers, sweating slides and other distractions during talks. "By Caveman". PMID- 11112680 TI - Actin dynamics. PMID- 11112685 TI - Lamins in disease: why do ubiquitously expressed nuclear envelope proteins give rise to tissue-specific disease phenotypes? AB - The nuclear lamina is a filamentous structure composed of lamins that supports the inner nuclear membrane. Several integral membrane proteins including emerin, LBR, LAP1 and LAP2 bind to nuclear lamins in vitro and can influence lamin function and dynamics in vivo. Results from various studies suggest that lamins function in DNA replication and nuclear envelope assembly and determine the size and shape of the nuclear envelope. In addition, lamins also bind chromatin and certain DNA sequences, and might influence chromosome position. Recent evidence has revealed that mutations in A-type lamins give rise to a range of rare, but dominant, genetic disorders, including Emery-Dreifuss muscular dystrophy, dilated cardiomyopathy with conduction-system disease and Dunnigan-type familial partial lipodystrophy. An examination of how lamins A/C, emerin and other integral membrane proteins interact at the INM provides the basis for a novel model for how mutations that promote disease phenotypes are likely to influence these interactions and therefore cause cellular pathology through a combination of weakness of the lamina or altered gene expression. PMID- 11112686 TI - Subversion of integrins by enteropathogenic Yersinia. AB - Enteropathogenic Yersinia are gram-negative bacterial species that translocate from the lumen of the intestine and are able to grow within deep tissue sites. During the earliest stages of disease, the organism is able to bind integrin receptors that are presented on the apical surface of M cells in the intestine, which allows its internalization and subsequent translocation into regional lymph nodes. The primary integrin substrate is the outer-membrane protein invasin, which binds with extraordinarily high affinity to at least five different integrins that have the (beta)(1) chain. Bacterial uptake into host cells is modulated by the affinity of receptor-substrate interaction, receptor concentration and the ability of the substrate to aggregate target receptors. PMID- 11112687 TI - Interferon gamma regulates accumulation of the proteasome activator PA28 and immunoproteasomes at nuclear PML bodies. AB - PA28 is an interferon (gamma) (IFN(gamma)) inducible proteasome activator required for presentation of certain major histocompatibility (MHC) class I antigens. Under basal conditions in HeLa and Hep2 cells, a portion of nuclear PA28 is concentrated at promyelocytic leukemia oncoprotein (PML)-containing bodies also commonly known as PODs or ND10. IFN(gamma) treatment greatly increased the number and size of the PA28- and PML-containing bodies, and the effect was further enhanced in serum-deprived cells. PML bodies are disrupted in response to certain viral infections and in diseases such as acute promyelocytic leukemia (APL). Like PML, PA28 was delocalized from PML bodies by expression of the cytomegalovirus protein, IE1, and in NB4 cells, an APL model line. Moreover, retinoic acid treatment, which causes remission of APL in patients and reformation of PML-containing bodies in NB4 cells, relocalized PA28 to this site. In contrast, the proteasome, the functional target of PA28, was not detected at PML bodies under basal conditions in HeLa and Hep2 cells, but IFN(gamma) promoted accumulation of 'immunoproteasomes' at this site. These results establish PA28 as a novel component of nuclear PML bodies, and suggest that PA28 may assemble or activate immunoproteasomes at this site as part of its role in proteasome dependent MHC class I antigen presentation. PMID- 11112688 TI - Df31 is a novel nuclear protein involved in chromatin structure in Drosophila melanogaster. AB - We originally isolated the Df31 protein from Drosophila embryo extracts as a factor which could decondense Xenopus sperm, by removing the sperm specific proteins and interacting with histones to facilitate their loading onto DNA. We now believe that this protein has a more general function in cellular DNA metabolism. The Df31 gene encodes a very hydrophilic protein with a predicted molecular mass of 18.5 kDa. Immunostaining showed that Df31 was present in a wide range of cell types throughout differentiation and in both dividing and non dividing cells. In all cases the protein is present in large amounts, comparable with the level of nucleosomes. Injection of antisense oligonucleotides to lower the level of Df31 in embryos caused severe disruption of the nuclear structure. Large irregular clumps of DNA were formed, and in most cases the amount of DNA associated with each clump was more than that found in a normal nucleus. Immunofluorescence, cell fractionation, and formaldehyde cross-linking show that Df31 is associated with chromatin and that a significant fraction of it binds very tightly. It also shows the same binding characteristics when loaded onto chromatin in vitro. Chromatin fractionation shows that Df31 is tightly associated with nucleosomes, preferentially with oligonucleosomes. Despite this no differences were observed in the properties of nucleosomes loaded in the in vitro system in the presence and absence of Df31. These results suggest that Df31 has a role in chromosomal structure, most likely acting as a structural protein at levels of folding higher than that of nucleosomes. PMID- 11112689 TI - Mitotic segregation of viral and cellular acentric extrachromosomal molecules by chromosome tethering. AB - Mitotic chromosome segregation is mediated by spindle microtubules attached to centromeres. Recent studies, however, revealed that acentric DNA molecules, such as viral replicons and double minute chromosomes, can efficiently segregate into daughter cells by associating with mitotic chromosomes. Based on this similarity between viral and cellular acentric molecules, we introduced Epstein-Barr virus vectors into cells harboring double minute chromosomes and compared their mitotic behaviors. We added lac operator repeats to an Epstein-Barr virus vector, which enabled us to readily identify the transgene in cells expressing a fusion protein between the lac repressor and green fluorescent protein. Unexpectedly, we found that Epstein-Barr virus vectors integrated into the acentric double minute chromosomes, but not into normal chromosomes, in all of the six stably transfected clones examined. While transiently transfected Epstein-Barr virus vectors randomly associated with wheel-shaped prometaphase chromosome rosettes, the chimeras of double minute chromosomes and Epstein-Barr virus vectors in stably transfected clones always attached to the periphery of chromosome rosettes. These chimeric acentric molecules faithfully represented the behavior of native double minute chromosomes, providing a tool for analyzing their behavior in living cells throughout the cell cycle. Further detailed analyses, including real-time observations, revealed that double minute chromosomes appeared to be repelled from the spindle poles at the same time that they attached to the chromosome periphery, while centromeric regions were pulled poleward by the attached microtubules. Disrupting microtubule organization eliminated such peripheral localization of double minute chromosomes, but it did not affect their association with chromosomes. The results suggest a model in which double minute chromosomes, but not Epstein-Barr virus vectors, are subject to the microtubule-mediated antipolar force, while they both employ chromosome tethering strategies to increase their segregation to daughter cells. PMID- 11112690 TI - Evidence for separate ND10-binding and homo-oligomerization domains of Sp100. AB - Nuclear domains called ND10 or PML nuclear bodies consist of an aggregation of several proteins, most notably PML and Sp100. PML is essential in the nucleation and formation of ND10 as well as in the recruitment of other ND10-associated proteins such as Daxx, pRb, BLM and Sp100. In cells induced to overexpress Sp100, ND10 binding of Sp100 was saturable and excess Sp100 formed new aggregation sites devoid of other ND10-associated proteins, suggesting that homo-oligomerization is the basis for aggregation. To determine whether Sp100 binds to ND10 through hetero- or oligomerization, Sp100 deletion variants fused with GFP were transfected into cells with and without endogenous Sp100, and the localization of the GFP-labeled fragments was determined relative to ND10. Amino acids 29-152 were sufficient for deposition of the GFP-labeled fragments at ND10 in the absence of endogenous Sp100 (heterologous binding) and for self-aggregation (formation of new Sp100 deposits). None of the shorter fragments was deposited at ND10 or self-aggregated. The 29-152 amino acid fragment and some larger fragments, but not the full-size Sp100, induced elongation of ND10, which at their ends contain only Sp100, probably due to self-aggregation. By fusing a peptide consisting of the p53-binding domain from hMDM2 to the Sp100(29-152) fragment, this self-aggregation could be blocked while retaining the limited ND10 binding capacity, indicating that the Sp100 self-aggregation domain and the ND10 binding domain are separate entities. This fusion peptide was used to demonstrate the potential of ND10 to recruit p53 as a protein not usually present at this site. Such deposited p53 was protected from turnover. The capacity of ND10 to recruit Sp100 may serve primarily to reduce its availability. PMID- 11112691 TI - Two type V myosins with non-overlapping functions in the fission yeast Schizosaccharomyces pombe: Myo52 is concerned with growth polarity and cytokinesis, Myo51 is a component of the cytokinetic actin ring. AB - The fission yeast genome project has identified five myosin genes: one type I myosin, myo1(+), two type II myosins, myo2(+) and myp2(+), and two type V myosins, myo51(+) and myo52(+). Cells deleted for myo51(+) show normal morphology and growth rates whereas deletion of myo52(+) results in a partial loss of cell polarity, slow growth and cytokinetic defects. Combining both deletions in a single strain is phenotypically non-additive, myo52(delta) being epistatic to myo51(delta). Overproduction of Myo51 gives rise to elongated cells which fail to form functional septa whereas overproduction of Myo52 results in branched cells with aberrant septa that fail to cleave. Myo52 localises to the poles of growing cells but during cell division it relocalises to the cell equator as a bar that is bisected by the cytokinetic septum. Myo51 shows no obvious localisation during interphase but at cytokinesis it is associated with the contractile cytokinetic actin ring (CAR). Both myosins are dependent upon an intact actin cytoskeleton for localisation. Myo52 partially colocalises with the (alpha)-glucan synthase Mok1 at the cell tips and to a lesser extent at the septum. Mok1 is delocalised and upregulated in myo52(delta) and myo52(delta) cell walls are resistant to digestion by the cell wall degrading enzyme zymolyase. Thus myo52(+) appears to be involved in the local delivery or positioning of vesicles containing cell wall precursors at the cell tips and has a role in the maturation or cleavage of the septum. Myo51 has a non-essential role in cytokinesis as a component of the cytokinetic actin ring. PMID- 11112692 TI - Overexpression of Akt inhibits NGF-induced growth arrest and neuronal differentiation of PC12 cells. AB - To investigate the role of Akt in nerve growth factor (NGF)-induced neuronal differentiation, PC12 cells ectopically expressing wild-type or dominant inhibitory forms of Akt were analyzed. NGF-induced neurite outgrowth was greatly accelerated in cells expressing dominant-inhibitory Akt, compared to parental PC12 cells, but was almost completely blocked in cells expressing wild-type Akt. Since neuronal differentiation requires an arrest of cell growth, several aspects of cell growth of the different cell lines were compared. Cells expressing wild type Akt were not susceptible to the growth-arresting effect of NGF, whereas parental PC12 cells and notably cells expressing mutant Akt were so affected. Accompanying this, the expressions of CDKs and p21(WAF1) were down- and up regulated, respectively, in both parental PC12 cells and cells expressing mutant Akt. When treated with some growth arrest-inducing agents such as sodium nitroprusside, forskolin and butyrolactone I, cells expressing wild-type Akt regained their responsiveness to the effects of NGF on differentiation. In summary, our results indicate that Akt overrides the growth-arresting effect of NGF and thereby, negatively regulates neuronal differentiation. PMID- 11112693 TI - Nuclear localization of Ku antigen is promoted independently by basic motifs in the Ku70 and Ku80 subunits. AB - The Ku antigen is a heteromeric (Ku70/Ku80), mostly nuclear protein. Ku participates in multiple nuclear processes from DNA repair to V(D)J recombination to telomere maintenance to transcriptional regulation and serves as a DNA binding subunit and allosteric regulator of DNA-dependent protein kinase. While some evidence suggests that subcellular localization of Ku may be subject to regulation, how Ku gains access to the nucleus is poorly understood. In this work, using a combination of indirect immunofluorescence and direct fluorescence, we have demonstrated that transfer of the Ku heterodimer to the nucleus is determined by basic nuclear localization signals in each of the Ku subunits that function independently. A bipartite basic nuclear localization signal between amino acids 539-556 of Ku70 was observed to be required for nuclear import of full-length Ku70 monomer, while a short Ku80 motif of four amino acids from 565 568 containing three lysines was required for the nuclear import of full-length Ku80. Ku heterodimers containing only one nuclear localization signal accumulated in the nucleus as efficiently as wild-type Ku, while site directed mutagenesis inactivating the basic motifs in each subunit, resulted in a Ku heterodimer that was completely localized to the cytoplasm. Lastly, our results indicate that mutations in Ku previously proposed to abrogate Ku70/Ku80 heterodimerization, markedly reduced the accumulation of Ku70 without affecting heterodimer formation in mammalian cells. PMID- 11112694 TI - Molecular characterization of the major membrane skeletal protein in the ciliate Tetrahymena pyriformis suggests n-plication of an early evolutionary intermediate filament protein subdomain. AB - Epiplasmin C is the major protein component of the membrane skeleton in the ciliate Tetrahymena pyriformis. Cloning and analysis of the gene encoding epiplasmin C showed this protein to be a previously unrecognized protein. In particular, epiplasmin C was shown to lack the canonical features of already known epiplasmic proteins in ciliates and flagellates. By means of hydrophobic cluster analysis (HCA), it has been shown that epiplasmin C is constituted of a repeat of 25 domains of 40 residues each. These domains are related and can be grouped in two families called types I and types II. Connections between types I and types II present rules that can be evidenced in the sequence itself, thus enforcing the validity of the splitting of the domains. Using these repeated domains as queries, significant structural similarities were demonstrated with an extra six heptads shared by nuclear lamins and invertebrate cytoplasmic intermediate filament proteins and deleted in the cytoplasmic intermediate filament protein lineage at the protostome-deuterostome branching in the eukaryotic phylogenetic tree. PMID- 11112695 TI - Release of an invasion promoter E-cadherin fragment by matrilysin and stromelysin 1. AB - The function of many transmembrane molecules can be altered by cleavage and subsequent release of their ectodomains. We have investigated ectodomain cleavage of the cell-cell adhesion and signal-transducing molecule E-cadherin. The E cadherin ectodomain is constitutively shed from the surface of MCF-7 and MDCKts.srcC12 cells in culture. Release of the 80 kDa soluble E-cadherin fragment is stimulated by phorbol-12-myristate-13-acetate and is inhibited by overexpression of the tissue inhibitor of metalloproteinases-2. The metalloproteinases matrilysin and stromelysin-1 both cleave E-cadherin at the cell surface and release sE-CAD into the medium. The soluble E-cadherin fragment thus released inhibits E-cadherin functions in a paracrine way, as indicated by induction of invasion into collagen type I and inhibition of E-cadherin-dependent cell aggregation. Our results, therefore, suggest a novel mechanism by which metalloproteinases can influence invasion. PMID- 11112696 TI - Involvement of actin filaments and integrins in the binding step in collagen phagocytosis by human fibroblasts. AB - In physiological conditions, collagen degradation by fibroblasts occurs primarily via phagocytosis, an intracellular pathway that is thought to require collagen receptors and actin assembly for fibril internalization and degradation. Currently it is unclear which specific steps of collagen phagocytosis in fibroblasts involve actin filament assembly. As studies of phagocytosis in fibroblasts are complicated by the relatively slow rate of particle internalization compared to professional phagocytes, we have examined the role of collagen receptors and actin only in the initial collagen binding step. Prior to the binding of collagen-coated fluorescent beads by human gingival fibroblasts, a cell type that is avidly phagocytic in vitro, cells were treated with cytochalasin D (actin filament barbed-end capping) or swinholide A (actin dimer sequestering and severing) or latrunculin B (actin monomer sequestering). Bead binding and immunostaining of (alpha)(2)(beta)(1) and (alpha)(3)(beta)(1) integrin collagen receptors were measured by flow cytometry. After 1-3 hours of coincubation with beads, cytochalasin D or swinholide A eliminated actin filaments stained by rhodamine-phalloidin and inhibited collagen bead binding (reductions of 25% and 50%, respectively), possibly because of cell rounding and restricted interactions with beads. In contrast, latrunculin enhanced binding dose-dependently over controls (twofold at 1 microM) and induced the formation of brightly staining aggregates of actin and the retention of long cytoplasmic extensions. Latrunculin also reduced surface (beta)(1), (alpha)(2) and (alpha)(3) integrin staining up to 40% in bead-free and bead-loaded cells, indicating that latrunculin enhanced collagen receptor internalization. As determined by fluorescence recovery after photobleaching, latrunculin increased the mobility of surface-bound (beta)(1) integrin. The stimulatory effect of latrunculin on collagen bead binding was reduced to control levels by treatment with a (beta)(1) integrin inactivating antibody while a (beta)(1) integrin blocking antibody abrogated both bead binding and the latrunculin-induced stimulation. Immunoblotting of bead-associated proteins showed that latrunculin completely eliminated binding of (beta)-actin to collagen beads but did not affect (beta)(1) integrin binding. These data indicate that latrunculin-induced sequestration of actin monomers facilitates the disengagement of actin from (beta)(1) integrin receptors, increases collagen bead binding and enhances collagen receptor mobility. We suggest that these alterations increase the probability of adhesive bead-to-cell interactions. PMID- 11112697 TI - TNF-alpha stimulates activation of pro-MMP2 in human skin through NF-(kappa)B mediated induction of MT1-MMP. AB - Tumor necrosis factor-alpha (TNF-(alpha)) is an important mediator during the inflammatory phase of wound healing. Excessive amounts of pro-inflammatory cytokines such as TNF-(alpha) are associated with inflammatory diseases including chronic wounds. Matrix metalloproteinases (MMPs) are involved in matrix re modeling during wound healing, angiogenesis and tumor metastasis. As with pro inflammatory cytokines, high levels of MMPs have been found in inflammatory states such as chronic wounds. In this report we relate these two phenomena. TNF (alpha) stimulates secretion of active MMP-2, a type IV collagenase, in organ cultured full-thickness human skin. This suggests a mechanism whereby excess inflammation affects normal wound healing. To investigate this observation at the cellular and molecular levels, we examined TNF-(alpha) mediated activation of pro MMP-2, induction of MT1-MMP, and the intracellular signaling pathways that regulate the proteinase in isolated human dermal fibroblasts. We found that TNF (alpha) substantially promoted activation of pro-MMP-2 in dermal fibroblasts embedded in type-I collagen. In marked contrast, collagen or TNF-(alpha) individually had little influence on the fibroblast-mediated pro-MMP-2 activation. One well-characterized mechanism for pro-MMP-2 activation is through a membrane type matrix metalloproteinase, such as MT1-MMP. We report that TNF (alpha) significantly induced MT1-MMP at the mRNA and protein levels when the dermal fibroblasts were grown in collagen. Although the intracellular signaling pathway regulating mt1-mmp gene expression is still obscure, both TNF-(alpha) and collagen activate the NF-(kappa)B pathway. In this report we provide three sets of evidence to support a hypothesis that activation of NF-(kappa)B is essential to induce MT1-MMP expression in fibroblasts after TNF-(alpha) exposure. First, SN50, a peptide inhibitor for NF-(kappa)B nuclear translocation, simultaneously blocked the TNF-(alpha) and collagen mediated MT1-MMP induction and pro-MMP-2 activation. Secondly, TNF-(alpha) induced I(kappa)B to breakdown in fibroblasts within the collagen lattice, a critical step leading to NF-(kappa)B activation. Lastly, a consensus binding site for p65 NF-(kappa)B (TGGAGCTTCC) was found in the 5'-flanking region of human mt1-mmp gene. Based on these results and previous reports, we propose a model to explain TNF-(alpha) activation of MMP-2 in human skin. Activation of NF(kappa)B signaling in fibroblasts embedded in collagen induces mt1-mmp gene expression, which subsequently activates the pro-MMP-2. The findings provide a specific mechanism whereby TNF-(alpha) may affect matrix remodeling during wound healing and other physiological and pathological processes. PMID- 11112698 TI - Targeting of cytoskeletal proteins to the flagellum of Trypanosoma brucei. AB - The eukaryotic flagellum represents one of the most complex macromolecular structures found in any organism and contains more than 250 proteins. Due to the relative ease of genetic manipulation the flagellum of Trypanosoma brucei has emerged as an accessible model system to study the morphogenesis and dynamics of this organelle. We have recently started to characterise the mechanisms by which components of the cytoskeletal fraction of the flagellum, such as the axoneme, the paraflagellar rod and the flagellar attachment zone, are targeted by proteins synthesised in the cytoplasm and assembled. Here, we present the identification of a novel actin-related protein as a component of the axoneme. We show that this protein shares the tripeptid motif histidine-leucine-alanine (HLA) with one of the major proteins of the paraflagellar rod, PFRA. Building on previous work from this lab which showed that a deletion comprising this motif abolished targeting of PFRA to the flagellum we demonstrate in this study that the deletion of the tripeptid motif is sufficient to achieve mistargeting both of the PFRA and the actin-related protein. We propose that this motif represents an essential part of a flagellar targeting machinery in trypanosomes and possibly in other flagellated organisms. PMID- 11112699 TI - Abnormal localisation and hyperclustering of (alpha)(V)(beta)(3) integrins and associated proteins in Src-deficient or tyrphostin A9-treated osteoclasts. AB - The non-receptor tyrosine kinase Src was shown to be essential for osteoclast function in vivo. We have previously reported that engagement of (alpha)(v)(beta)(3) integrin in osteoclasts induces tyrosine phosphorylation and activation of the adhesion kinase PYK2 and the adaptor protein p130(Cas) in a Src dependent manner. The objective of this study was to analyse the role of c-Src in the (alpha)(v)(beta)(3) integrin-dependent recruitment of signalling and cytoskeletal molecules in osteoclasts during bone resorption. Using prefusion osteoclasts (pOCs) obtained from cocultures of osteoblasts and spleen cells isolated from Src(-/-) mice or their normal littermates, we found: (1) similar expression levels and ligand binding affinities of (alpha)(v)(beta)(3) integrins in Src(-/-) and Src(+/?) pOCs, (2) reduced adhesion and spreading of Src(-/-) pOCs, (3) defective organisation of the microfilament proteins, F-actin, vinculin and paxillin, and of PYK2 and p130(Cas) in the sealing zone of Src(-/-)OCLs, and (4) hyperclustering of (alpha)(v)(beta)(3) integrins together with microfilament and signalling proteins in the basal membrane of Src-deficient OCLs. In normal OCLs, the tyrosine kinase inhibitor tyrphostin A9 inhibits actin ring formation, bone resorption and tyrosine phosphorylation of several proteins, including c Src. Furthermore, tyrphostin A9 induced similar hyperclustering of (alpha)(v)(beta)(3) integrins in osteoclasts as observed in Src(-/-) OCLs. Taken together, these findings suggest that normal localisation of (alpha)(v)(beta)(3) and recruitment of its downstream effectors to the appropriate compartments of the osteoclast during resorption depend on Src kinase activity. PMID- 11112700 TI - Characterization of the microtubule binding domain of microtubule actin crosslinking factor (MACF): identification of a novel group of microtubule associated proteins. AB - MACF (microtubule actin cross-linking factor) is a large, 608-kDa protein that can associate with both actin microfilaments and microtubules (MTs). Structurally, MACF can be divided into 3 domains: an N-terminal domain that contains both a calponin type actin-binding domain and a plakin domain; a rod domain that is composed of 23 dystrophin-like spectrin repeats; and a C-terminal domain that includes two EF-hand calcium-binding motifs, as well as a region that is homologous to two related proteins, GAR22 and Gas2. We have previously demonstrated that the C-terminal domain of MACF binds to MTs, although no homology was observed between this domain and other known microtubule-binding proteins. In this report, we describe the characterization of this microtubule binding domain of MACF by transient transfection studies and in vitro binding assays. We found that the C-terminus of MACF contains at least two microtubule binding regions, a GAR domain and a domain containing glycine-serine-arginine (GSR) repeats. In transfected cells, the GAR domain bound to and partially stabilized MTs to depolymerization by nocodazole. The GSR-containing domain caused MTs to form bundles that are still sensitive to nocodazole-induced depolymerization. When present together, these two domains acted in concert to bundle MTs and render them stable to nocodazole treatment. Recently, a study has shown that the N-terminal half of the plakin domain (called the M1 domain) of MACF also binds MTs. We therefore examined the microtubule binding ability of the M1 domain in the context of the entire plakin domain with and without the remaining N-terminal regions of two different MACF isoforms. Interestingly, in the presence of the surrounding sequences, the M1 domain did not bind MTs. In addition to MACF, cDNA sequences encoding the GAR and GSR-containing domains are also found in the partial human EST clone KIAA0728, which has high sequence homology to the 3' end of the MACF cDNA; hence, we refer to it as MACF2. The C terminal domain of mouse MACF2 was cloned and characterized. The microtubule binding properties of MACF2 C-terminal domain are similar to that of MACF. The GAR domain was originally found in Gas 2 protein and here we show that it can associate with MTs in transfected cells. Plectin and desmoplakin have GSR containing domains at their C-termini and we further demonstrate that the GSR containing domain of plectin, but not desmoplakin, can bind to MTs in vivo. PMID- 11112701 TI - A novel nucleolar G-protein conserved in eukaryotes. AB - We describe here a novel, evolutionarily conserved set of predicted G-proteins. The founding member of this family, TbNOG1, was identified in a two-hybrid screen as a protein that interacts with NOPP44/46, a nucleolar phosphoprotein of Trypanosoma brucei. The biological relevance of the interaction was verified by co-localization and co-immunoprecipitation. TbNOG1 localized to the trypanosome nucleolus and interacted with domains of NOPP44/46 that are found in several other nucleolar proteins. Genes encoding proteins highly related to TbNOG1 are present in yeast and metazoa, and related G domains are found in bacteria. We show that NOG1 proteins in humans and Saccharomyces cerevisae are also nucleolar. The S. cerevisae NOG1 gene is essential for cell viability, and mutations in the predicted G motifs abrogate function. Together these data suggest that NOG1 may play an important role in nucleolar functions. The GTP-binding region of TbNOG1 is similar to those of Obg and DRG proteins, which, together with NOG, form a newly recognized family of G-proteins, herein named ODN. The ODN family differs significantly from other G-protein families, and shows several diagnostic sequence characteristics. All organisms appear to possess an ODN gene, pointing to the biological significance of this family of G-proteins. PMID- 11112702 TI - Latent TGF-beta binding protein LTBP-1 contains three potential extracellular matrix interacting domains. AB - Latent TGF-beta binding proteins (LTBPs) are components of the extracellular matrix (ECM). They belong to the fibrillin/LTBP-superfamily, and are high molecular weight glycoproteins characterized by EGF-like repeats and 8-Cys repeats. Most LTBPs associate with the small latent forms of TGF-beta. Their roles include to facilitate the secretion of latent TGF-beta and to target it to the ECM. In order to identify new matrix-binding domains of LTBP-1 and to characterize their association with the extracellular matrix, we have produced (in a mammalian expression system) partly overlapping recombinant fragments of its shorter form, LTBP-1S, and analyzed the binding of the purified fusion proteins to extracellular matrices of cultured human dermal and lung fibroblasts. Recombinant fragments from three different regions of the N- and C-termini showed affinity to the matrix. These interacting regions contain either the first (hybrid), second or fourth 8-Cys domains of the LTBP-1S molecule. They bound independently to the matrix. Each of them had an ability to inhibit the association of native exogenous LTBP-1 with fibroblast extracellular matrix. The interactions of the LTBP-1 fragments with the extracellular matrix resisted treatment with sodium deoxycholate, suggesting strong, possibly covalent binding. The binding occurred in a time- and dose-dependent fashion. The N-terminal fragments bound more readily to the matrices. With all fragments the binding took place both with intact fibroblast matrices and with matrices isolated by sodium deoxycholate. When using CHO cell layers, which form sparse matrices, only the N terminal fragment of LTBP-1 was efficiently incorporated. The association of the binding fragments with isolated matrices was enhanced by soluble, cell-derived factors. The current data suggest that LTBP-1 contains three different domains with an ability to associate with the extracellular matrix. PMID- 11112703 TI - Glycosaminoglycan synthesis and secretion by the retinal pigment epithelium: polarized delivery of hyaluronan from the apical surface. AB - Hyaluronan and chondroitin sulfate glycosaminoglycan secretion from retinal pigment epithelial cells was established in confluent cultures with high transepithelial resistance. Cell cultures were maintained on Millicell-PCF culture plates, which allow separation of culture medium exposed to apical and basal epithelial surfaces. Following various times in culture, apical and basal culture media were sampled at three day intervals and the glycosaminoglycan content was quantified. Samples were digested with proteinase K to free the glycosaminoglycans from their core proteins, the glycosaminoglycans were ethanol precipitated, and subjected to hyaluronidase SD and chondroitinase ABC digestion to release hyaluronan and chondroitin sulfate disaccharides. Disaccharides were fluorotagged with 2-aminoacridone, separated on polyacrylamide gels and the molar fluorescence in each disaccharide band quantitated. Hyaluronan in the apical medium was significantly higher than in the basal medium (5-12 times) at all recovery intervals (P<0.0001). In contrast, the distribution of unsulfated chondroitin, 4-sulfated chondroitin and 6-sulfated chondroitin disaccharides in apical and basal media was non-polar. Confocal microscopy of cultures probed with a hyaluronan-specific fluorotag established that the HA evident in these cultures is restricted to the apical border of the RPE cultures. Collectively, these data indicate that hyaluronan synthesized by the retinal pigment epithelium is secreted preferentially from the apical surface, suggesting that this tissue is an important source of hyaluronan present in the interphotoreceptor matrix. PMID- 11112704 TI - Cdc28-Clb mitotic kinase negatively regulates bud site assembly in the budding yeast. AB - In the budding yeast Saccharomyces cerevisiae, a prospective mother normally commences the formation of a daughter (the bud) only in the G(1) phase of the cell division cycle. This suggests a strict temporal regulation of the processes that initiate the formation of a new bud. Using cortical localization of bud site components Spa2 and Bni1 as an indicator of bud site assembly, we show that cells assemble a bud site following inactivation of the Cdc28-Clb mitotic kinase but prior to START. Interestingly, an untimely inactivation of the mitotic kinase is sufficient to drive cells to assemble a new bud site inappropriately in G(2) or M phases. The induction of Cdc28/Clb kinase activity in G(1), on the other hand, dramatically reduces a cell's ability to construct an incipient bud site. Our findings strongly suggest that the Cdc28-Clb kinase plays a critical role in the mechanism that restricts the timing of bud formation to the G(1) phase of the cell cycle. PMID- 11112705 TI - Localization of cellubrevin-related peptide, endobrevin, in the early endosome in pancreatic beta cells and its physiological function in exo-endocytosis of secretory granules. AB - Cellubrevins are integral membrane proteins expressed in a wide variety of tissues and usually localized in recycling vesicles. Here, we investigated the cellular localization of a cellubrevin-related peptide, endobrevin, in pancreatic (beta) cells and its implication in the exo-endocytosis of insulin and (gamma) amino butyric acid (GABA). Immunocytochemistry showed that endobrevin is associated with tubulo-vesicular structures, which are colocalized with early endosomes labeled by early endosome antigen (EEA)-1 in insulinoma MIN6 cells. To determine the cellular localization of endobrevin, we appended the green fluorescent protein (GFP) to endobrevin and the fusion protein was introduced into MIN6 cells. The subcellular localization of GFP-endobrevin was visualized by confocal laser microscopy. Colocalization study based on the expressed GFP endobrevin and endocytosed Texas-Red(Tx-R) labeled transferrin receptor and immunocytochemistry with anti-EEA1 antibody revealed that endobrevin was preferentially localized in the early endosome. Then, we examined the functional role of endobrevin in the exocytosis of insulin and GABA from pancreatic (beta) cells. Endobrevin overexpression increased the amount of GABA released from MIN6 cells; in contrast, it decreased the glucose-stimulated insulin release from rat islets, MIN6 and INS1-D cells to approximately 50% of the control levels. Both in vitro and in vivo binding studies showed that endobrevin binds to syntaxin 1. Finally, using the fluorescent probe FM4-64, it was revealed that endobrevin overexpression accelerates vesicle recycling. We conclude that (1) endobrevin is localized in the early endosome in pancreatic (beta) cells and (2) endobrevin plays a physiological role in the exo-endocytosis of insulin and GABA from pancreatic (beta) cells, probably via an interaction between endocytic vesicles and the endosome. PMID- 11112706 TI - Immunolocalization of cytoplasmic dynein and dynactin subunits in cultured macrophages: enrichment on early endocytic organelles. AB - Cytoplasmic dyneins and their cofactor, dynactin, work together to mediate the movement of numerous cargo organelles toward the minus-ends of microtubules. In many cases, there is compelling evidence that dynactin functions in part to attach dyneins to cargo organelles, but this may not always be the case. We have localized three dynactin subunits (Arp1, p62 and p150(Glued)) and two subunits of conventional cytoplasmic dynein (dynein intermediate chain and dynein heavy chain 1) in murine macrophages using immunogold labeling of thawed cryosections. Using stereological techniques, we have quantified the relative distributions of each of these subunits on specific membrane organelles to generate a comprehensive analysis of the distribution of these proteins in a single cell type. Our results show that each of the subunits tested exhibits the same distribution with respect to different membrane organelles, with highest levels present on early endosomes, and lower levels present on later endocytic organelles, the mitochondrial outer membrane, the plasma membrane and vesicles in the Golgi region. An additional pool of punctate dynactin labeling was detected in the cell periphery, in the absence of dynein labeling. Even when examined closely, membrane organelles could not be detected in association with these dynactin-positive sites; however, double labeling with anti-tubulin antibody revealed that at least some of these sites represent the ends of microtubules. The similarities among the labeling profiles with respect to membrane organelles suggest that dynein and dynactin bind to membrane organelles as an obligate unit. In contrast, our results show that dynactin can associate with microtubule ends in the absence of dynein, perhaps providing sites for subsequent organelle and dynein association to form a functional motility complex. PMID- 11112707 TI - The two fundamental duties of the physician? PMID- 11112709 TI - Radiation treatment PMID- 11112708 TI - The two fundamental duties of the physician? PMID- 11112710 TI - Residents' work hours: the Achilles heel of the profession? PMID- 11112711 TI - Implementing a comprehensive relative-value-based incentive plan in an academic family medicine department. AB - The authors describe the implementation and first three years (1997-1999) of a department-wide incentive plan of the Department of Family Medicine at the State University of New York at Buffalo School of Medicine and Biomedical Sciences. By using a consensus approach, a representative elected committee designed a clinical relative value unit (explained in detail) that could be translated to equally value and reward faculty efforts in patient care, education, and research and which allowed the department to avoid the imposition of a model that could have undervalued scholarship and teaching. By 1999, the plan's goal of eight patient-care-equivalent points per four-hour session had been exceeded for pure clinical care. Clearly, only a small financial incentive was necessary (in 1999, an incentive pool of 4% of providers' gross salary) to motivate the faculty to be more productive and to self-report their efforts. Long-term productivity for pure clinical care rose from 9.8 points per session in 1997 to 10.4 in 1999. Of the mean total of 3,980 points for the year 1999, the contribution from teaching was 1,146, or 29%, compared with 25% in 1997. For scholarship, the number of points was 775, or 20%, in 1999, compared with 11% in 1997. The authors describe modifications to the original plan (e.g., integration of quality measures) that the department's experience has fostered. Problems encountered included the lack of accurate and timely billing information from the associated teaching hospitals, the inherent problems of self-reported information, difficulties of gaining buy-in from the faculty, and inherent risks of a pay-for-performance approach. But the authors conclude that the plan is fulfilling its goal of effectively and fairly quantifying all areas of faculty effort, and is also helping the department to more effectively demonstrate clinical productivity in negotiations with teaching hospitals. PMID- 11112712 TI - Continuing medical education: a new vision of the professional development of physicians. AB - The authors describe their vision of what continuing medical education (CME) should become in the changing health care environment. They first discuss six types of literature (e.g., concerning learning and adult development principles, problem-based/practice-based learning, and other topics) that contribute to ways of thinking about and understanding CME. They then state their view that the Association of American Medical Colleges (AAMC) has made a commitment to helping CME be more effective in the professional development of physicians. In presenting their new vision of CME, the authors describe their interpretation of the nature and values of CME (e.g., optimal CME is highly self-directed; the selection and design of the most relevant CME is based on data from each physician's responsibilities and performance; etc.). They then present seven action steps, suggestions to begin them, and the institutions and organizations they believe should carry them out, and recommend that the AAMC play a major role in supporting activities to carry out these steps. (For example, one action step is the generation and application of new knowledge about how and why physicians learn, select best practices, and change their behaviors). Six core competencies for CME educators are defined. The authors conclude by stating that collaboration among the appropriate academic groups, professional associations, and health care institutions, with leadership from the AAMC, is essential to create the best learning systems for the professional development of physicians. PMID- 11112713 TI - If students are not customers, what are they? AB - The answers to questions about the relationship between faculty and students including medical students-depend on an understanding of the nature of teaching and the underlying ethical principles of our society. The authors maintain that teaching is purposive, rational, communal, and moral. They assert that Western society is based on the values of liberal democracy and that the key ethical principles for the professions that are derived from those values are autonomy, standard of care, and respect for democratic institutions. There are three candidates for ethical models on which to base the relationship between students and faculty. Two of them (clientism and paternalism) the authors reject. The one that they favor (the fiduciary model) is based on mutual trust and respect, which both students and faculty have responsibilities to maintain. Using that model, the authors conclude that students are, in some aspects, customers of faculty. This student-centered approach is balanced by treating society and other faculty as customers as well. Pathologies in medical education attributed to clientism (such as an obsession with marks and overemphasis on memorization) existed well before medical students were purportedly being treated as customers; perhaps it is not the student who is "broken" but the system in which the student is made to function. Whether students are called customers, clients, knowledge workers, or simply students, faculty must involve them more in shaping their education and in dealing with enduring problems that profoundly affect their learning. PMID- 11112714 TI - Competency-based residency training: the next advance in graduate medical education. AB - The goal of all graduate medical education is to ensure that the graduating physician is competent to practice in his or her chosen field of medicine. The evaluation of a resident's competency to practice, however, has never been clearly defined, nor has the fixed period of time given for residency training in each specialty been shown to be the right amount of time for each individual resident to achieve competency. To better ensure that new physicians have the competencies they need, the author proposes the replacement of the current approach to residents' education, which specifies a fixed number of years in training, with competency-based training, in which each resident remains in training until he or she has been shown to have the required knowledge and skills and can apply them independently. Such programs, in addition to tailoring the training time to each individual, would make it possible to devise and test schemes to evaluate competency more surely than is now possible. The author reviews the basis of traditional residency training and the problems with the current training approach, both its fixed amount of time for training and the uncertainty of the methods of evaluation used. He then explains competency-based residency education, notes that it is possible, indeed probable, that some trainees will become competent considerably sooner than they would in the current required years of training, quotes a study in which this was the case, and explains the implications. He describes the encouraging experience of his neurosurgery department, which has used competency-based training for its residents since 1994. He then discusses issues of demonstrating competency in procedural and nonprocedural fields, as well as the evaluation of competency in traditional and competency-based training, emphasizing that the latter approach offers hope for better ways of assessing competency. PMID- 11112715 TI - What evidence supports teaching evidence-based medicine? PMID- 11112716 TI - Strategic planning and the balanced scorecard for faculty practice plans. PMID- 11112717 TI - Premedical education. 1926. PMID- 11112718 TI - Medicine and the arts. Patchwork Sky and A Close Shave. PMID- 11112719 TI - A close shave PMID- 11112720 TI - Patchwork Sky PMID- 11112721 TI - Certification and specialization: do they matter in the outcome of acute myocardial infarction? AB - PURPOSE: To learn whether there are differences among certified and self designated cardiologists, internists, and family practitioners in terms of the mortality of their patients with acute myocardial infarction (AMI). METHOD: Data on all patients admitted with AMI were collected for calendar year 1993 by the Pennsylvania Health Care Cost Containment Council and analyzed. Certified and self-designated family practitioners, internists, and cardiologists (n = 4,546) were compared with respect to the characteristics of their patients' illnesses. In addition, a regression model was fitted in which mortality was the dependent measure and the independent variables were the probability of death, hospital characteristics (location and the availability of advanced cardiac care), and physician characteristics (patient volume, years since graduation from medical school, specialty, and certification status). RESULTS: On average, cardiologists treated more patients than did generalists, and their patients were less severely ill. In the regression analysis, all variables were statistically significant except the availability of advanced cardiac care. Holding all other variables constant, treatment by a certified physician was associated with a 15% reduction in mortality among patients with AMI. CONCLUSIONS: Less patient mortality was associated with treatment by physicians who were cardiologists, cared for larger numbers of AMI patients, were closer to their graduation from medical school, and were certified. PMID- 11112722 TI - Does a physician's ability to accurately assess the likelihood of pulmonary embolism increase with training? AB - PURPOSE: Pulmonary embolism (PE), an elusive diagnosis, is detected by a diagnostic work-up that is often guided by the physician's level of clinical suspicion. The ability to accurately assess PE risk on solely clinical grounds may increase with the physician's level of training. This study documented the ability of house staff practicing in an academic teaching hospital to accurately assess the clinical likelihood of PE in patients. METHOD: During a seven-month period, all 245 patients with suspected acute PE who had had lung scans ordered via a computerized order-entry system were enrolled in the study. When ordering the lung scans, all physicians (interns, residents, and attending physicians) were required to also enter their levels of clinical suspicion on a scale of 0 to 100. The physicians' levels of clinical suspicion were correlated with the final determinations of PE, and receiver operating characteristic (ROC) curves were calculated for patients' and physicians' subgroups. RESULTS: Attending physicians were most able to diagnose PE; residents were moderately able to make the diagnosis, and interns were least able to diagnose PE. The area under the ROC curve for a correct identification of patients with PE was greatest for attending physicians (0.839), intermediate for residents (0.601), and least for interns (0.594). CONCLUSION: The ability to correctly assess a patient's likelihood of PE increases with a physician's level of training, suggesting that more senior physicians should be involved in the diagnostic work-up of patients with suspected acute PE. More instruction may help medical students, interns, and residents navigate clinical scenarios in which the diagnosis is uncertain or in which sequential tests must be performed to reach the correct diagnosis. PMID- 11112723 TI - A comparison of physician examiners', standardized patients', and communication experts' ratings of international medical graduates' English proficiency. AB - PURPOSE: To assess the quality of ratings of interviewing skills and oral English proficiency provided on a clinical skills OSCE by physician examiners, standardized patients (SPs), and communication skills experts. METHOD: In 1998, 73 candidates to the Ontario International Medical Graduate (OIMG) Program completed a 29-station OSCE-type clinical skills selection examination. Physician examiners, SPs, and communication skills experts assessed components of oral English proficiency and interview performance. Based on these results, the frequency and generalizability of English-language flags, physician examiners' indications that spoken English skills were bad enough to significantly impede communication with patients; the reliability of the OIMG's Interview and Oral Performance Scales and generalizability of overall interview and oral performance ratings; and comparisons of repeated assessments by experts were calculated. Principal-components analysis was applied to the panels' ratings to determine a more economical expression of the language proficiency and interview communication skills results. RESULTS: The mean number of English-language flags per candidate was 2.1, the median was 1.0, and Cronbach's alpha of the ratings was 0.63. Means, SDs, and alphas of the physician examiners' and SPs' ratings of the interview performance scale were 9.15/10, 0.43, 0.36, and 9.30/10, 0. 56, 0.50, respectively. Corresponding values for overall interview performance ratings were 3.08/4, 0.30, 0.33, and 3.34/4, 0.32, 0.47. Means, SDs, and alphas of the physician examiners' and SPs' ratings of the oral performance scale were 8.54/10, 0.74, 0.78, and 8.74/10, 1.00, 0.76. Corresponding values for overall ratings of oral performance were 3.85/5, 0.51, 0.68, and 4.08/5, 0.60, 0.68. For the two experts' ratings of two contiguous five-minute interview stations, internal consistencies were 0.88 and 0.78. For the two experts' ratings of standardized ten-minute interviews, internal consistencies were 0.81 and 0.92. Correlations between the mean values of the experts' ratings of the ten- and five minute stations were 0.45 and 0.51. Three factors emerged from the PCA, language proficiency, physician examiners' ratings of interview proficiency, and SPs' ratings of interview proficiency. CONCLUSIONS: Consistency between the physician examiners' and SPs' ratings of English proficiency was observed; less agreement was observed in their ratings of interviewing skills, and little agreement was observed between the experts' ratings. Communication skills results may be validly expressed by three measures: one overall global rating of language proficiency provided by physician examiners or SPs, and overall global ratings of interview proficiency provided separately by physician examiners and SPs. PMID- 11112724 TI - Effect of an evidence-based medicine seminar on participants' interpretations of clinical trials: a pilot study. AB - PURPOSE: To evaluate the effect of evidence-based medicine (EBM) education on physicians' short-term and long-term understanding of research methods and statistics. METHOD: Twenty-four gastroenterology (GI) fellows attended a three day seminar about evidence-based medicine and the critical appraisal of medical literature. Attendees completed the same 14-item test on this material at the start of the seminar, at the conclusion of the seminar, and six months after the seminar. A student's t-test and chi-square analysis were performed to determine the differences between test scores by testing date and performance on test items. RESULTS: Seminar attendees improved their test scores between pre-seminar and post-seminar tests (mean test score: 57% +/- 16% versus 82 +/- 14%, respectively; p <.001) and between pre-seminar and six-month post-seminar tests (mean test score: 57% +/- 16% versus 78% +/- 13%, respectively; p <.001). Seminar attendees showed significant improvement in frequency of correct answers with individual questions on concealment of allocation, relative risk reduction, and meta-analysis trial methods. CONCLUSIONS: In this pilot study, the critical appraisal skills necessary to practice EBM were taught to GI fellows in a seminar format that led to significant improvement in their understanding of research methods and statistics. Data from this pilot study justify a definitive trial examining the educational value of EBM seminars for physicians. PMID- 11112725 TI - Differences in medical students' empathy. AB - Medical students' Balanced Emotional Empathy Scale scores were compared by year, sex, and expressed specialty choice. Scores were lower for students choosing non core specialties, and for M4 men students compared with M3 men students. PMID- 11112726 TI - Using formal evaluation sessions for case-based faculty development during clinical clerkships. AB - Developing housestaff and faculty in their roles as medical educators is a dynamic process. The rigorous clinical evaluation method used during the third year internal medicine clerkship at the Uniformed Services University uniquely incorporates faculty development into the process of evaluation and generating feedback for students. Formal evaluation sessions are held monthly at all clerkship sites throughout the 12-week clerkship and are moderated by either the internal medicine clerkship director or the on-site clerkship directors. Although designed to provide an opportunity for faculty to evaluate student performance and prepare formative feedback, the sessions also function as formal, planned, and longitudinal forums of "real-time," "case-based" faculty development that address professional, instructional, and leadership development. The evaluation sessions are used as a means to model and teach the key concepts of the Stanford Faculty Development Program. Providing a unifying form of evaluation across multiple teaching sites and settings makes formal evaluation sessions a powerful, state-of-the-art tool for faculty development. PMID- 11112727 TI - Faculty development in communication skills instruction: insights from a longitudinal program with "real-time feedback". AB - Responsibility for teaching communication skills often falls to a multidisciplinary group of faculty who lack both a common model for teaching and prior experience teaching communication in small groups. This article describes East Tennessee State University's multifaceted faculty development program in teaching communication skills. The program was developed and implemented in three phases. First, a two-step Delphi approach helped identify core communication skills. Phase two gave faculty the opportunity to practice identifying communication teaching issues and effective strategies for working with small groups. The third phase involved the videotaping of faculty teaching small groups of students. These tapes were reviewed both individually and in faculty groups. The tapes were also reviewed by students, who provided realtime, moment-to-moment feedback to the faculty. Implementation and review of the program has helped to identify new strategies for effectively facilitating small-group teaching of communication skills. PMID- 11112728 TI - Evidence-based morning report for inpatient pediatrics rotations. AB - The authors describe a patient-centered method for teaching evidence-based medicine that is part of the inpatient morning report for pediatrics residents at the University of Illinois at Chicago. With library support, residents search for evidence to answer their own questions about patients, and present it at morning report PMID- 11112729 TI - Medical school problems. 1926. PMID- 11112731 TI - Editor's pick PMID- 11112730 TI - Measuring contributions to the clinical mission of medical schools and teaching hospitals. AB - This is the final report of a panel convened as part of the Association of American Medical College's (AAMC's) Mission-based Management Program to examine the use of metrics (i.e., measures) in assessing faculty and departmental contributions to the clinical mission. The authors begin by focusing on methods employed to estimate clinical effort and calculate a "clinical full-time equivalent," a prerequisite to comparing productivity among faculty members and departments. They then identify commonly used metrics, including relative-value units, total patient-care gross charges, total net patient fee-for-service revenue, total volume per CPT (current procedural terminologies) code by service category and number of patients per physician, discussing their advantages and disadvantages. These measures reflect the "twin pillars" of measurement criteria, those based on financial or revenue information, and those based on measured activity. In addition, the authors urge that the assessment of quality of care become more highly developed and integrated into an institution's measurement criteria. The authors acknowledge the various ways users of clinical metrics can develop standards against which to benchmark performance. They identify organizations that are sources of information about external national standards, acknowledge various factors that confound the interpretation of productivity data, and urge schools to identify and measure secondary service indicators to assist with interpretation and provide a fuller picture of performance. Finally, they discuss other, non-patient-care, activities that contribute to the clinical mission, information about which should be incorporated into the overall assessment. In summary, the authors encourage the use of clinical productivity metrics as an integral part of a comprehensive evaluation process based upon clearly articulated and agreed-upon goals and objectives. When carefully designed, these measurement systems can provide critical information that will enable institutional leaders to recognize and reward faculty and departmental performance in fulfillment of the clinical mission. PMID- 11112732 TI - This month in wjm PMID- 11112733 TI - Your spouse/partner gets a skin infection and needs antibiotics: is it ethical for you to prescribe for them? Yes: it is ethical to treat short-term, minor problems. PMID- 11112734 TI - Your spouse/partner gets a skin infection and needs antibiotics: is it ethical for you to prescribe for them? No: such treatment rarely leads to comprehensive care. PMID- 11112735 TI - The ritalin wars continue. PMID- 11112736 TI - Living wills can help doctors and patients talk about dying PMID- 11112737 TI - Rocket science. wjm launches a fast-track system for publishing important papers. PMID- 11112739 TI - How to submit your medical images for publication in Med.Pix PMID- 11112738 TI - A netlike rash. PMID- 11112740 TI - Personalized medicine comes a step closer for asthma. PMID- 11112741 TI - Laparoscopic surgery: two thirds of injuries initially missed. PMID- 11112742 TI - Online pharmacies: industry growing, but safety questions remain. PMID- 11112743 TI - Everything you ever wanted to know about your anatomy. PMID- 11112744 TI - A book to make you think PMID- 11112745 TI - Severe varicella in an immunocompromised adult presenting with abdominal pain. PMID- 11112747 TI - Any Questions? any answers? PMID- 11112746 TI - Inequality and political power. PMID- 11112748 TI - US adolescent food intake trends from 1965 to 1996. AB - OBJECTIVE: To examine adolescent food consumption trends in the United States with important chronic disease implications. METHODS: Analysis of dietary intake data from 4 nationally representative US Department of Agriculture surveys of persons aged 11 to 18 years (n = 12,498). RESULTS: From 1965 to 1996, a considerable shift occurred in the adolescent diet. Total energy intake decreased, as did the proportion of energy from total fat (39%-32%) and saturated fat (15%-12%). Concurrent increases occurred in the consumption of higher-fat potatoes and mixed dishes (pizza and macaroni and cheese). Lower-fat milks replaced higher-fat milks, but total milk consumption decreased by 36%. This decrease was accompanied by an increase in the consumption of soft drinks and noncitrus juices. An increase in high-fat potato consumption led to an increase in vegetable intake, but the number of servings for fruits and vegetables is still lower than the recommended 5 per day. Iron, folic acid, and calcium intakes continue to be below those recommended for girls. CONCLUSIONS: These trends, far greater than for US adults, may compromise the health of the future US population. PMID- 11112749 TI - Clinical implications of trends in food intake among US adolescents. PMID- 11112750 TI - Qualitative interview study of communication between parents and children about maternal breast cancer. AB - OBJECTIVE: To examine parents' communication with their children about the diagnosis and initial treatment of breast cancer in the mother. DESIGN: Qualitative interview study within a cross-sectional cohort. SETTING: Two breast cancer treatment centers. PARTICIPANTS: 32 women with stage I or stage II breast cancer with 56 school-aged children. Main outcome measures Semistructured interview regarding timing and extent of communication with children about the diagnosis and initial treatment of the mother's illness, reasons for talking to children or withholding information, and help available and requested from health professionals. RESULTS: Women were most likely to begin talking to their children after their diagnosis had been confirmed by biopsy, but a few waited until after surgery or said nothing at all. Family discussion did not necessarily include mention of cancer. There was considerable consistency in the reasons given for either discussing or not discussing the diagnosis. The most common reason for not communicating was to avoid children's questions, particularly those about death. Although most women had helpful discussion with a physician concerning their illness, few were offered help with talking to their children; many would have liked help, particularly the opportunity for both parents to talk to a health professional with experience in understanding and talking to children. CONCLUSION: Parents diagnosed with cancer or other serious illnesses should be offered help to think about whether, what, and how to tell their children and about what children can understand, especially as they may well be struggling themselves to come to terms with their illness. PMID- 11112751 TI - Treating cancer as a family disease. PMID- 11112752 TI - A survey of physician attitudes and practices concerning cost-effectiveness in patient care. AB - OBJECTIVE: To identify physicians' views regarding cost-containment and cost effectiveness and their attitudes and experience using cost-effectiveness in clinical decision making. DESIGN: A close-ended 30-item written survey. SUBJECTS: 1,000 randomly selected physicians whose practices currently encompass direct patient care and who work in the California counties of Sacramento, Yolo, Placer, Nevada, and El Dorado. OUTCOME MEASURES: Physician attitudes about the role of cost and cost-effectiveness in treatment decisions, perceived barriers to cost effective medical practice, and response of physicians and patients if there are conflicts about treatment that physicians consider either not indicated or not cost-effective. RESULTS: Most physicians regard cost-effectiveness as an appropriate component of clinical decisions and think that only the treating physician and patient should decide what is cost-worthy. However, physicians are divided on whether they have a duty to offer medical interventions with remote chances of benefit regardless of cost, and they vary considerably in their interactions with patients when cost-effectiveness is an issue. CONCLUSION: Although physicians in the Sacramento region accept cost-effectiveness as important and appropriate in clinical practice, there is little uniformity in how cost-effectiveness decisions are implemented. PMID- 11112753 TI - Cost-effectiveness: no easy choices or answers. PMID- 11112754 TI - Are naturalistic weight-reducing efforts associated with weight gain and onset of obesity in adolescent girls? PMID- 11112755 TI - Are naturalistic weight-reducing efforts associated with weight gain and onset of obesity in adolescent girls? PMID- 11112756 TI - In children receiving antibiotics, does coadministration of Lactobacillus GG reduce the incidence of diarrhea? PMID- 11112757 TI - In children receiving antibiotics, does coadministration of lactobacillus GG reduce the incidence of diarrhea? PMID- 11112758 TI - Drug treatment of multiple sclerosis PMID- 11112759 TI - Chest pain evaluation units PMID- 11112760 TI - Withdrawal of medical treatment in children. PMID- 11112761 TI - Withdrawing and withholding life-sustaining treatment in children. PMID- 11112764 TI - Students may delay putting on condoms PMID- 11112762 TI - The cardiac complications of recreational drug use. PMID- 11112765 TI - Chronic fatigue syndrome: is it physical? PMID- 11112766 TI - Hospitals in rural America. PMID- 11112767 TI - Myth: identifying classic coronary risk factors helps to predict the likelihood of acute ischemia. PMID- 11112768 TI - Civilization and the colon. Constipation as "the disease of diseases". PMID- 11112770 TI - Meeting needs with available resources PMID- 11112769 TI - The power of song. PMID- 11112771 TI - Mitochondrial control of iron homeostasis. A genome wide analysis of gene expression in a yeast frataxin-deficient strain. AB - Deletion of YFH1, the yeast frataxin homologue gene, elicits mitochondrial iron accumulation and alters cellular iron homeostasis. Here, we report a genome wide analysis of gene expression in a yfh1(DeltaYFH1) deleted strain. Frataxin deficiency results in enhanced expression of some 70 genes including a set of genes, called the iron regulon, that are under the control of the iron-sensing transcription factor AFT1. Five new AFT1-dependent genes, YOR382w, YOR383c, YDR534c, YLR136c, and YLR205c were found. The first three genes presumably encode cell-wall glycosylphosphatidylinositol anchor proteins and exhibit a 30-100-fold increased expression. The triple deletion of these genes decreases efficiency in utilization of the iron of ferrioxamine B by the yeast cell. YLR136c bears homology to tristetraproline proteins, which are post-transcriptional regulators in mammalian cells. Deletion of YLR136c increases the mRNA levels of iron regulon members. YLR205c bears homology to heme oxygenases. Our data show that frataxin deficiency elicits iron mobilization from all iron sources in an AFT1-dependent manner. Wild-type and DeltaYFH1 glycerol-grown cells exhibit similar high respiration rates, no mitochondrial iron accumulation, and high expression of the iron regulon, suggesting that under these conditions little iron is extruded from mitochondria. These data suggest that the activity of Yfh1p is not essential in cells grown on glycerol. This study has also revealed unexpected links between mitochondria and remote metabolic pathways since frataxin deficiency also enhances the expression of genes such as HSP30, that escape to AFT1 control. Finally, no oxidative stress gene is induced. PMID- 11112772 TI - DSIF and NELF interact with RNA polymerase II elongation complex and HIV-1 Tat stimulates P-TEFb-mediated phosphorylation of RNA polymerase II and DSIF during transcription elongation. AB - Control of transcription elongation requires a complex interplay between the recently discovered positive transcription elongation factor b (P-TEFb) and negative transcription elongation factors, 5,6-dichloro-1-beta-d ribofuranosylbenzimidazole (DRB) sensitivity inducing factors (DSIF) and the negative elongation factor (NELF). Activation of HIV-1 gene expression is regulated by a nascent RNA structure, termed TAR RNA, in concert with HIV-1 Tat protein and these positive and negative elongation factors. We have used a stepwise RNA pol II walking approach and Western blotting to determine the dynamics of interactions between HIV-1 Tat, DSIF/NELF, and the transcription complexes actively engaged in elongation. In addition, we developed an in vitro kinase assay to determine the phosphorylation status of proteins during elongation stages. Our results demonstrate that DSIF/NELF associates with RNA pol II complexes during early transcription elongation and travels with elongation complexes as the nascent RNA is synthesized. Our results also show that HIV-1 Tat protein stimulated DSIF and RNA pol II phosphorylation by P-TEFb during elongation. These findings reveal a molecular mechanism for the negative and positive regulation of transcriptional elongation at the HIV-1 promoter. PMID- 11112773 TI - Caveolin-1 associates with TRAF2 to form a complex that is recruited to tumor necrosis factor receptors. AB - Tumor necrosis factor (TNF) receptor-associated factor (TRAF) 2 is an intracellular adapter protein, which, upon TNF stimulation, is directly recruited to the intracellular region of TNF receptor 2 (TNFR2) or indirectly, via TRADD, to the intracellular region of TNF receptor 1 (TNFR1). In cultured human umbilical vein endothelial cells, endogenous TRAF2 colocalizes with the membrane organizing protein caveolin-1 at regions of enrichment subjacent to the plasma membrane as detected by confocal fluorescence microscopy. Both endogenous and transfected TRAF2 protein coimmunoprecipitate with caveolin-1 in the absence of ligand. Upon TNF treatment, the TRAF2-caveolin-1 complex transiently associates with TRADD, and upon overexpression of TNFR2, the TRAF2-caveolin-1 complex stably associates with and causes redistribution of this receptor as detected by confocal fluorescence microscopy. In human embryonic kidney 293 cells, which have minimal endogenous expression of caveolin-1, cotransfection of TRAF2 and caveolin 1 results in spontaneous association of these proteins which can further associate with and redistribute transfected TNFR2 molecules. The association of caveolin-1 with TNFR2 depends upon TRAF2. Cotransfection of caveolin-1 protein increases TRAF2 protein expression levels in HEK 293 cells, which correlates with enhancement of TNF and TRAF2 signaling, measured as transcription of a NF-kappaB promoter-reporter gene, although the caveolin-enhanced response to TNF is attenuated at higher caveolin levels. These findings suggest that intracellular distribution of activated TNF receptors may be regulated by caveolin-1 via its interaction with TRAF2. PMID- 11112774 TI - Modulation of the herpes simplex virus type-1 UL9 DNA helicase by its cognate single-strand DNA-binding protein, ICP8. AB - The mechanism of stimulation of a DNA helicase by its cognate single-strand DNA binding protein was examined using herpes simplex virus type-1 UL9 DNA helicase and ICP8. UL9 and ICP8 are two essential components of the viral replisome that associate into a complex to unwind the origins of replication. The helicase and DNA-stimulated ATPase activities of UL9 are greatly elevated as a consequence of this association. Given that ICP8 acts as a single-strand DNA-binding protein, the simplest model that can account for its stimulatory effect predicts that it tethers UL9 to the DNA template, thereby increasing its processivity. In contrast to the prediction, data presented here show that the stimulatory activity of ICP8 does not depend on its single-strand DNA binding activity. Our data support an alternative hypothesis in which ICP8 modulates the activity of UL9. Accordingly, the data show that the ICP8-binding site of UL9 constitutes an inhibitory region that maintains the helicase in an inefficient ground state. ICP8 acts as a positive regulator by neutralizing this region. ICP8 does not affect substrate binding, ATP hydrolysis, or the efficiency of translocation/DNA unwinding. Rather, we propose that ICP8 increases the efficiency with which substrate binding and ATP hydrolysis are coupled to translocation/DNA unwinding. PMID- 11112775 TI - Identification of a novel cis element required for cell density-dependent down regulation of insulin-like growth factor-2 P3 promoter activity in Caco2 cells. AB - The activity of the exogenous, full-length insulin-like growth factor-2 (IGF-2) P3 promoter is significantly up-regulated during the logarithmic growth phase but rapidly declines in confluent CaCo2 cells undergoing differentiation. Nuclear run on assays confirmed cell density-dependent regulation of endogenous P3 promoter. To identify regulatory elements in the P3 promoter that may be required for regulating cell density-dependent transcriptional activity, we used the methods of promoter truncation, electrophoretic mobility shift assay, DNase footprinting, and mutation analysis. The relative activity of the full-length (-1229/+140) and truncated (-1090/+140) promoter was identical, being approximately 19, 27, 7, and 3% of pSV-luc activity on days 3, 5, 7, and 9 of cell culture, respectively. However, truncation to -1048 resulted in complete loss of cell density-dependent down-regulation of P3 promoter activity on days 7 and 9, suggesting the presence of regulatory elements between -1091 and -1048 sequence. Further stepwise truncation to -515 did not change promoter activity. Truncation to -138/+140 resulted in complete loss of promoter activity, suggesting that the core promoter was within the -515/-138 segment. A 14-base pair footprint (-1084/-1070) was identified by DNase footprinting within the distal -1091/-1048 segment. Electrophoretic mobility shift assay with wild type and mutant probes confirmed the presence of a novel 7-base pair (CGAGGGC) (-1084/-1078) cis element (P3-D); its mutation abolished binding. Functionality of P3-D cis element was confirmed by measuring the activity of core P3 promoter ligated to distal P3 segment containing either the mutant or wild type P3-D element. We have, therefore, identified a novel cis element, P3-D, that appears to play a critical role in regulating IGF-2 P3 promoter activity in a cell density/differentiation-dependent manner. PMID- 11112776 TI - Localization and insulin-regulated relocation of phosphoinositide 5-kinase PIKfyve in 3T3-L1 adipocytes. AB - The mammalian phosphoinositide kinase PIKfyve catalyzes the synthesis of phosphatidylinositol 5-P and phosphatidylinositol 3,5-P(2), thought essential in cellular functions, including membrane trafficking. To discern the intracellular loci of PIKfyve products' formation, we have examined the localization of PIKfyve protein versus enzymatic activity and a possible acutely regulated redistribution in 3T3-L1 adipocytes. Subcellular fractions of resting cells that were positive for immunoreactive PIKfyve, such as cytosol ( approximately 76%), internal structures (low density microsomal fraction (LDM), composed of recycling endosomes, GLUT4 storage compartment, Golgi, and cytoskeletal elements) ( approximately 20%), and plasma membrane (( approximately )4%), expressed enzymatically active PIKfyve. While the presence of a FYVE finger in PIKfyve predicts early endosome targeting, density gradient sedimentation, immunoadsorption, and fluorescence microscopy analyses segregated the LDM associated PIKfyve from the membranes of the recycling endosomes and GLUT4. PIKfyve fluorescence staining largely coincided with trans-Golgi network/multivesicular body markers, indicating PIKfyve's role in the late endocytic/biosynthetic pathways. A subfraction of particulate PIKfyve resisted nonionic detergent treatment, implying association with cytoskeletal structures, previously found positive for key members of the insulin signaling cascade. Upon acute stimulation of 3T3-L1 adipocytes with insulin or pervanadate, a portion of the cytosolic PIKfyve was recruited onto LDM, which was coupled with a commensurate increase of PIKfyve lipid kinase activity and an electrophoretic mobility shift. We suggest the recruited PIKfyve specifies the site and timing of phosphoinositide signals that are relevant to the acute insulin action. PMID- 11112777 TI - Rational design of a peptide agonist that interacts selectively with the orphan receptor, bombesin receptor subtype 3. AB - The orphan receptor, bombesin (Bn) receptor subtype 3 (BRS-3), shares high homology with bombesin receptors (neuromedin B receptor (NMB-R) and gastrin releasing peptide receptor (GRP-R)). This receptor is widely distributed in the central nervous system and gastrointestinal tract; target disruption leads to obesity, diabetes, and hypertension, however, its role in physiological and pathological processes remain unknown due to lack of selective ligands or identification of its natural ligand. We have recently discovered (Mantey, S. A., Weber, H. C., Sainz, E., Akeson, M., Ryan, R. R. Pradhan, T. K., Searles, R. P., Spindel, E. R., Battey, J. F., Coy, D. H., and Jensen, R. T. (1997) J. Biol. Chem. 272, 26062-26071) that [d-Tyr(6),beta-Ala(11),Phe(13),Nle(14)]Bn-(6-14) has high affinity for BRS-3 and using this ligand showed BRS-3 has a unique pharmacology with high affinity for no known natural Bn peptides. However, use of this ligand is limited because it has high affinity for all known Bn receptors. In the present study we have attempted to identify BRS-3 selective ligands using a strategy of rational peptide design with the substitution of conformationally restricted amino acids into the prototype ligand [d-Tyr(6),beta Ala(11),Phe(13),Nle(14)]Bn-(6-14) or its d-Phe(6) analogue. Each of the 22 peptides synthesized had binding affinities determined for hBRS-3, hGRPR, and hNMBR, and hBRS-3 selective ligands were tested for their ability to activate phospholipase C and increase inositol phosphates ([(3)H]inositol phosphate). Using this approach we have identified a number of BRS-3 selective ligands. These ligands functioned as receptor agonists and their binding affinities were reflected in their potencies for altering [(3)H]inositol phosphate. Two peptides with an (R)- or (S)-amino-3-phenylpropionic acid substitution for beta-Ala(11) in the prototype ligand had the highest selectivity for the hBRS-3 over the mammalian Bn receptors and did not interact with receptors for other gastrointestinal hormones/neurotransmitters. Molecular modeling demonstrated these two selective BRS-3 ligands had a unique conformation of the position 11 beta-amino acid. This selectivity was of sufficient magnitude that these should be useful in explaining the role of hBRS-3 activation in obesity, glucose homeostasis, hypertension, and other physiological or pathological processes. PMID- 11112778 TI - Human Ku70 interacts with heterochromatin protein 1alpha. AB - Ku is involved in the metabolism of DNA ends, DNA repair, and the maintenance of telomeres. It consists of a heterodimer of 70- and 80-kDa subunits. Recently we have demonstrated that Ku70 interacted with TRF2, a mammalian telomere-binding protein. Using the same yeast two-hybrid screening system, we now show that Ku70 also interacts with heterochromatin protein 1alpha (HP1alpha), a protein known to be associated with telomeres as well as heterochromatin. HP1 is a suppressor of the position effect variegation in Drosophila and acts as a transcriptional suppressor in mammalian cells. The interaction with Ku70 in the two-hybrid system was confirmed by a glutathione S-transferase pull-down study using bacterial recombinant proteins in vitro. The interaction was also reproduced in vivo in HeLa cells, where endogenous Ku70 coimmunoprecipitated with HP1alpha. This interaction was more effective in acidic pH and weakened considerably as the pH of the reaction buffer was elevated up to 7.5. Ku80 did not interact with HP1alpha directly. The interaction domains of Ku70 and HP1alpha included the Leu Ser repeat (amino acids 200-385) and the chromo shadow domain, respectively. Ku70 was largely colocalized with transfected HP1alpha but not with a C-terminal deletion mutant, HP1alpha(Delta)C. In contrast to HP1alpha, Ku70 did not repress transcriptional activity of the reporter gene when tethered to DNA after transfection to mammalian cells. The implication of this interaction is discussed. PMID- 11112779 TI - The rat liver Na(+)/bile acid cotransporter. Importance of the cytoplasmic tail to function and plasma membrane targeting. AB - To understand the potential functions of the cytoplasmic tail of Na(+)/taurocholate cotransporter (Ntcp) and to determine the basolateral sorting mechanisms for this transporter, green fluorescent protein-fused wild type and mutant rat Ntcps were constructed and the transport properties and cellular localization were assessed in transfected COS 7 and Madin-Darby canine kidney (MDCK) cells. Truncation of the 56-amino acid cytoplasmic tail demonstrates that the cytoplasmic tail of rat Ntcp is involved membrane delivery of this protein in nonpolarized and polarized cells and removal of the tail does not affect the bile acid transport function of Ntcp. Using site-directed mutagenesis, two tyrosine residues, Tyr-321 and Tyr-307, in the cytoplasmic tail of Ntcp have been identified as important for the basolateral sorting of rat Ntcp in transfected MDCK cells. Tyr-321 appears to be the major basolateral-sorting determinant, and Tyr-307 acts as a supporting determinant to ensure delivery of the transporter to the basolateral surface, especially at high levels of protein expression. When the two Tyr-based basolateral sorting motifs have been removed, the N-linked carbohydrate groups direct the tyrosine to alanine mutants to the apical surface of transfected MDCK cells. The major basolateral sorting determinant Tyr-321 is within a novel beta-turn unfavorable tetrapeptide Y(321)KAA, which has not been found in any naturally occurring basolateral sorting motifs. Two-dimensional NMR spectroscopy of a 24-mer peptide corresponding to the sequence from Tyr-307 to Thr-330 on the cytoplasmic tail of Ntcp confirms that both the Tyr-321 and Tyr 307 regions do not adopt any turn structure. Since the major motif YKAA contains a beta-turn unfavorable structure, the Ntcp basolateral sorting may not be related to the clathrin-adaptor complex pathway, as is the case for many basolateral proteins. PMID- 11112780 TI - A chicken gonadotropin-releasing hormone receptor that confers agonist activity to mammalian antagonists. Identification of D-Lys(6) in the ligand and extracellular loop two of the receptor as determinants. AB - Mammalian receptors for gonadotropin-releasing hormone (GnRH) have over 85% sequence homology and similar ligand selectivity. Biological studies indicated that the chicken GnRH receptor has a distinct pharmacology, and certain antagonists of mammalian GnRH receptors function as agonists. To explore the structural determinants of this, we have cloned a chicken pituitary GnRH receptor and demonstrated that it has marked differences in primary amino acid sequence (59% homology) and in its interactions with GnRH analogs. The chicken GnRH receptor had high affinity for mammalian GnRH (K(i) 4.1 +/- 1.2 nM), similar to the human receptor (K(i) 4.8 +/- 1.2 nM). But, in contrast to the human receptor, it also had high affinity for chicken GnRH ([Gln(8)]GnRH) and GnRH II ([His(5),Trp(7),Tyr(8)]GnRH) (K(i) 5.3 +/- 0.5 and 0.6 +/- 0.01 nM). Three mammalian receptor antagonists were also pure antagonists in the chicken GnRH receptor. Another three, characterized by D-Lys(6) or D-isopropyl-Lys(6) moieties, functioned as pure antagonists in the human receptor but were full or partial agonists in the chicken receptor. This suggests that the Lys side chain interacts with functional groups of the chicken GnRH receptor to stabilize it in the active conformation and that these groups are not available in the activated human GnRH receptor. Substitution of the human receptor extracellular loop two with the chicken extracellular loop two identified this domain as capable of conferring agonist activity to mammalian antagonists. Although functioning of antagonists as agonists has been shown to be species-dependent for several GPCRs, the dependence of this on an extracellular domain has not been described. PMID- 11112781 TI - The dhb operon of Bacillus subtilis encodes the biosynthetic template for the catecholic siderophore 2,3-dihydroxybenzoate-glycine-threonine trimeric ester bacillibactin. AB - Bacillus subtilis was reported to produce the catecholic siderophore itoic acid (2,3-dihydroxybenzoate (DHB)-glycine) in response to iron deprivation. However, by inspecting the DNA sequences of the genes dhbE, dhbB, and dhbF as annotated by the B. subtilis genome project to encode the synthetase complex for the siderophore assembly, various sequence errors within the dhbF gene were predicted and confirmed by re-sequencing. According to the corrected sequence, dhbF encodes a dimodular instead of a monomodular nonribosomal peptide synthetase. We have heterologously expressed, purified, and assayed the substrate selectivity of the recombinant proteins DhbB, DhbE, and DhbF. DhbE, a stand-alone adenylation domain of 59.9 kDa, activates, in an ATP-dependent reaction, DHB, which is subsequently transferred to the free thiol group of the cofactor phosphopantetheine of the bifunctional isochorismate lyase/aryl carrier protein DhbB. The third synthetase, DhbF, is a dimodular nonribosomal peptide synthetase of 264 kDa that specifically adenylates threonine and, to a lesser extent, glycine and that covalently loads both amino acids onto their corresponding peptidyl carrier domains. To functionally link the dhb gene cluster to siderophore synthesis, we have disrupted the dhbF gene. Comparative mass spectrometric analysis of culture extracts from both the wild type and the dhbF mutant led to the identification of a mass peak at m/z 881 ([M-H](1-)) that corresponds to a cyclic trimeric ester of DHB-glycine-threonine. PMID- 11112782 TI - Cystic fibrosis transmembrane conductance regulator gating requires cytosolic electrolytes. AB - Cystic fibrosis transmembrane conductance regulator (CFTR), which causes cystic fibrosis when nonfunctional, is an anion channel and a member of the ATP binding cassette superfamily. After phosphorylation, CFTR gates by binding and hydrolyzing ATP. We show that CFTR open probability (P(o)) also depends on the electrolyte concentration of the cytosol. Inside-out patches from Calu-3 cells were transiently exposed to solutions of 160 mm salt or solutions in which up to 90% of the salt was replaced by nonionic osmolytes such as sucrose. In lowered salt solutions, CFTR P(o) declined within 1 s to a stable lower value that depended on the electrolyte concentration, (K(1/2) approximately 80 mm NaCl). P(o) was rapidly restored in normal salt concentrations without regard to the electrolyte species. Reducing external electrolytes did not affect CFTR P(o). The same results were obtained when CFTR was stably phosphorylated with adenosine 5' O-(thiotriphosphate). The decrease in P(o) resulted entirely from an increase in mean closed time. Increasing ATP levels up to 20-fold did not counteract the effect of low electrolytes. The same effect was observed for CFTR expressed in C127 cells but not for a different species of anion channel. Cytosolic electrolytes are an unsuspected, essential cofactor for CFTR gating. PMID- 11112783 TI - Role of the differentially spliced carboxyl terminus in thromboxane A2 receptor trafficking: identification of a distinct motif for tonic internalization. AB - The thromboxane A(2) receptor (TP) is a G protein-coupled receptor that is expressed as two alternatively spliced isoforms, alpha (343 residues) and beta (407 residues) that share the first 328 residues. We have previously shown that TPbeta, but not TPalpha, undergoes agonist-induced internalization in a dynamin-, GRK-, and arrestin-dependent manner. In the present report, we demonstrate that TPbeta, but not TPalpha, also undergoes tonic internalization. Tonic internalization of TPbeta was temperature- and dynamin-dependent and was inhibited by sucrose and NH(4)Cl treatment but unaffected by wild-type or dominant-negative GRKs or arrestins. Truncation and site-directed mutagenesis revealed that a YX(3)phi motif (where X is any residue and phi is a bulky hydrophobic residue) found in the proximal portion of the carboxyl-terminal tail of TPbeta was critical for tonic internalization but had no role in agonist induced internalization. Interestingly, introduction of either a YX(2)phi or YX(3)phi motif in the carboxyl-terminal tail of TPalpha induced tonic internalization of this receptor. Additional analysis revealed that tonically internalized TPbeta undergoes recycling back to the cell surface suggesting that tonic internalization may play a role in maintaining an intracellular pool of TPbeta. Our data demonstrate the presence of distinct signals for tonic and agonist-induced internalization of TPbeta and represent the first report of a YX(3)phi motif involved in tonic internalization of a cell surface receptor. PMID- 11112784 TI - Mitogen-activated protein kinases mediate activator protein-1-dependent human inducible nitric-oxide synthase promoter activation. AB - Inducible nitric-oxide synthase (iNOS) is an important signaling protein involved in the regulation of biological processes (e.g. vasodilation, inflammation) and is subject to transcriptional regulation by cytokines and lipopolysaccharide (LPS). Full activation of the human iNOS (hiNOS) promoter by cytokines (i.e., tumor necrosis factor-alpha, interleukin-1beta, interferon-gamma (IFN-gamma)) required downstream and upstream nuclear factor-kappaB (-115, -8283) and activator protein-1 (AP-1) (-5115, -5301) transcription factor binding sites. Human lung epithelial (A549) cells were transiently transfected with luciferase reporter plasmids containing an 8.3-kilobase human iNOS promoter to examine the molecular signaling events necessary for hiNOS transcriptional activation. The combination of LPS and IFN-gamma, but neither alone, increased hiNOS promoter activity 28-fold, in a reaction requiring two critical AP-1 (JunD-Fra-2) promoter binding sites. Mitogen-activated protein kinases (MAPKs) were assessed as potential activators of AP-1 and the hiNOS promoter. Both pharmacological and molecular inhibitors of the extracellular signal-related kinase (ERK) and p38 pathways reduced cytokine mixture (CM)- and LPS/IFN-gamma-induced promoter activation. By gel retardation analysis, the addition of MAP/ERK kinase-1 and p38 inhibitors significantly diminished AP-1 binding in both CM- and LPS/IFN-gamma stimulated cells. Thus, p38- and ERK-dependent pathways, through effects on the AP-1 complex, activate the hiNOS promoter in cells stimulated with CM or LPS/IFN gamma. PMID- 11112785 TI - A mutational epitope for cytochrome C binding to the apoptosis protease activation factor-1. AB - Cytochrome c (Cc) binding to apoptosis protease activation factor-1 (Apaf-1) is a critical activation step in the execution phase of apoptosis. Here we report studies that help define the Cc:Apaf-1 binding surface. It is shown that a large number of Cc residues, including residues 7, 25, 39, 62-65, and 72, are involved in the Cc:Apaf-1 interaction. Mutation of residue 72 eliminated Cc activity whereas mutations of residues 7, 25, 39, and 62-65 showed reduced activity in an additive fashion. The implications of this binding model for both recognition and modulation of protein-protein interactions are briefly discussed. PMID- 11112786 TI - A role for poly(ADP-ribose) polymerase in the transcriptional regulation of the melanoma growth stimulatory activity (CXCL1) gene expression. AB - The melanoma growth stimulatory activity/growth-regulated protein, CXCL1, is constitutively expressed at high levels during inflammation and progression of melanocytes into malignant melanoma. It has been shown previously that CXCL1 overexpression in melanoma cells is due to increased transcription as well as stability of the CXCL1 message. The transcription of CXCL1 is regulated through several cis-acting elements including Sp1, NF-kappaB, HMGI(Y), and the immediate upstream region (IUR) element (nucleotides -94 to -78), which lies immediately upstream to the nuclear factor kappaB (NF-kappaB) element. Previously, it has been shown that the IUR is necessary for basal and cytokine-induced transcription of the CXCL1 gene. UV cross-linking and Southwestern blot analyses indicate that the IUR oligonucleotide probe selectively binds a 115-kDa protein. In this study, the IUR element has been further characterized. We show here that proximity of the IUR element to the adjacent NF-kappaB element is critical to its function as a positive regulatory element. Using binding site oligonucleotide affinity chromatography, we have selectively purified the 115-kDa IUR-F. Mass spectrometry/mass spectrometry/matrix-assisted laser desorption ionization/time of flight spectroscopy and amino acid analysis as well as microcapillary reverse phase chromatography electrospray ionization tandem mass spectrometry identified this protein as the 114-kDa poly(ADP-ribose) polymerase (PARP1). Furthermore, 3 aminobenzamide, an inhibitor of PARP-specific ADP-ribosylation, inhibits CXCL1 promoter activity and reduces levels of CXCL1 mRNA. The data point to the possibility that PARP may be a coactivator of CXCL1 transcription. PMID- 11112787 TI - Human complex I defects can be resolved by monoclonal antibody analysis into distinct subunit assembly patterns. AB - Complex I defects are one of the most frequent causes of mitochondrial respiratory chain disorders. Therefore, it is important to find new approaches for detecting and characterizing Complex I deficiencies. In this paper, we introduce a new set of monoclonal antibodies that react with 39-, 30-, 20-, 18-, 15-, and 8-kDa subunits of Complex I. These antibodies are shown to aid in diagnosis of Complex I deficiencies and add understanding to the genotype phenotype relationships of different mutations. A total of 11 different patients were examined. Four patients had undefined Complex I defects, whereas the other patients had defects in NDUFV1, NDUFS2 (two patients), NDUFS4 (two patients), NDUFS7, and NDUFS8. We show here that Western blotting with these antibodies, particularly when used in conjunction with sucrose gradient studies and enzymatic activity measurements, helps distinguish catalytic versus assembly defects and further distinguishes between mutations in different subunits. Furthermore, different mutations in the same gene are shown to give very similar subunit profiles, and we show that one of the patients is a good candidate for having a defect in a Complex I assembly factor. PMID- 11112788 TI - Glucose and insulin function through two distinct transcription factors to stimulate expression of lipogenic enzyme genes in liver. AB - Transcription of a number of genes involved in lipogenesis is stimulated by dietary carbohydrate in the mammalian liver. Both insulin and increased glucose metabolism have been proposed to be initiating signals for this process, but the pathways by which these effectors act to alter transcription have not been resolved. We have previously defined by electrophoretic mobility shift assay a factor in nuclear extracts from rat liver, designated the carbohydrate-responsive factor (Cho- RF), that binds to liver-type pyruvate kinase and S(14) promoters at sites critical for regulation by carbohydrate. The sterol regulatory element binding protein-1c (SREBP-1c) has also emerged as a major transcription factor involved in this nutritional response. In this study, we examined the relationship between SREBP-1c and ChoRF in lipogenic gene induction. The two factors were found to possess distinct DNA binding specificities both in vitro and in hepatocytes. Reporter constructs containing binding sites for ChoRF were responsive to glucose but not directly to insulin. On the other hand, reporter constructs with an SREBP-1c site responded directly to insulin. The S(14) gene possesses binding sites for both ChoRF and SREBP, and both sites were found to be functionally important for the response of this promoter to glucose and insulin in hepatocytes. Consequently, we propose that SREBP-1c and ChoRF are independent transcription factors that mediate signals generated by insulin and glucose, respectively. For many lipogenic enzyme genes, these two factors may provide an integrated signaling system to support the overall nutritional response to dietary carbohydrate. PMID- 11112789 TI - Hypoxia inhibits G1/S transition through regulation of p27 expression. AB - Mammalian cellular responses to hypoxia include adaptive metabolic changes and a G1 cell cycle arrest. Although transcriptional regulation of metabolic genes by the hypoxia-induced transcription factor (HIF-1) has been established, the mechanism for the hypoxia-induced G1 arrest is not known. By using genetically defined primary wild-type murine embryo fibroblasts and those nullizygous for regulators of the G1/S checkpoint, we observed that the retinoblastoma protein is essential for the G1/S hypoxia-induced checkpoint, whereas p53 and p21 are not required. In addition, we found that the cyclin-dependent kinase inhibitor p27 is induced by hypoxia, thereby inhibiting CDK2 activity and forestalling S phase entry through retinoblastoma protein hypophosphorylation. Reduction or absence of p27 abrogated the hypoxia-induced G1 checkpoint, suggesting that it is a key regulator of G1/S transition in hypoxic cells. Intriguingly, hypoxic induction of p27 appears to be transcriptional and through an HIF-1-independent region of its proximal promoter. This demonstration of the molecular mechanism of hypoxia induced G1/S regulation provides insight into a fundamental response of mammalian cells to low oxygen tension. PMID- 11112790 TI - Identification of ERSE-II, a new cis-acting element responsible for the ATF6 dependent mammalian unfolded protein response. AB - Herp is a 54-kDa membrane protein in the endoplasmic reticulum (ER). The mRNA expression level of Herp is increased by the accumulation of unfolded proteins in the ER. Transcriptional changes designed to deal with this type of ER stress is called the unfolded protein response (UPR). Most mammalian UPR-target genes encode ER-resident molecular chaperones: GRP78, GRP94, and calreticulin. The promoter regions of these genes contain a cis-acting ER stress response element, ERSE, with the consensus sequence of CCAAT-N(9)-CCACG. Under conditions of ER stress, p50ATF6 (the active form of the transcription factor, ATF6) binds to CCACG when CCAAT is bound by the general transcription factor, NF-Y/CBF. Here, we report the genomic structure of human Herp and the presence of a new ER stress response element, ERSE-II, in its promoter region. The gene for Herp consists of eight exons, localized to chromosome 16q12.2-13. The promoter region contains a single ERSE-like sequence. In reporter gene assays, disruption of this cis element resulted in a partial reduction of the transcriptional response to ER stress, suggesting that the element is functional for the UPR. These results also suggest the involvement of additional elements in the UPR. Further analysis, using an optimized plasmid containing an mRNA-destabilizing sequence, revealed ERSE-II (ATTGG-N-CCACG) as the second ER stress response element. Interestingly, ERSE-II was also dependent on p50ATF6, in a manner similar to that of ERSE, despite the disparate structure. The strong induction of Herp mRNA by ER stress would be achieved by the cooperation of ERSE and ERSE-II. PMID- 11112795 TI - Maze navigation by honeybees: learning path regularity. AB - We investigated the ability of honeybees to learn mazes of four types: constant turn mazes, in which the appropriate turn is always in the same direction in each decision chamber; zig-zag mazes, in which the appropriate turn is alternately left and right in successive decision chambers; irregular mazes, in which there is no readily apparent pattern to the turns; and variable irregular mazes, in which the bees were trained to learn several irregular mazes simultaneously. The bees were able to learn to navigate all four types of maze. Performance was best in the constant-turn mazes, somewhat poorer in the zig-zag mazes, poorer still in the irregular mazes, and poorest in the variable irregular mazes. These results demonstrate that bees do not navigate such mazes simply by memorizing the entire sequence of appropriate turns. Rather, performance in the various configurations depends on the existence of regularity in the structure of the maze and on the ease with which this regularity is recognized and learned. PMID- 11112796 TI - Perception and recognition memory in monkeys following lesions of area TE and perirhinal cortex. AB - Monkeys with lesions of perirhinal cortex (PR group) and monkeys with lesions of inferotemporal cortical area TE (TE group) were tested on a modified version of the delayed nonmatching to sample (DNMS) task that included very short delay intervals (0.5 sec) as well as longer delay intervals (1 min and 10 min). Lesions of the perirhinal cortex and lesions of area TE produced different patterns of impairment. The PR group learned the DNMS task as quickly as normal monkeys (N) when the delay between sample and choice was very short (0.5 sec). However, performance of the PR group, unlike that of the N group, fell to chance levels when the delay between sample and choice was lengthened to 10 min. In contrast to the PR group, the TE group was markedly impaired on the DNMS task even at the 0.5 sec delay, and three of four monkeys with TE lesions failed to acquire the task. The results provide support for the idea that perirhinal cortex is important not for perceptual processing, but for the formation and maintenance of long-term memory. Area TE is important for the perceptual processing of visual stimuli. PMID- 11112797 TI - Impairment of spatial learning and hippocampal synaptic potentiation in c-kit mutant rats. AB - The c-kit receptor tyrosine kinase encoded by the white-spotting (W) gene is highly expressed in rat hippocampal CA1-CA4 regions. We found an impaired spatial learning and memory in homozygous c-kit (Ws/Ws) mutant rats that have a 12-base deletion in the tyrosine kinase domain of the c-kit gene and a very low kinase activity. Electrophysiological studies in hippocampal slices revealed that the long-term potentiation (LTP) induced by the tetanic stimulation (100 Hz, 1 sec) in the mossy fiber (MF)-CA3 pathway, but not in the Schaffer collaterals/commissural-CA1 pathway, was significantly reduced in c-kit mutants compared with wild-type (+/+) rats. The paired-pulse facilitation (PPF) was measured before the tetanus and after the establishment of the LTP in each slice. The initial PPF in the MF-CA3 pathway positively correlated with the amplitude of the LTP in the wild-type rats but not in the c-kit mutant rats. Furthermore, they failed to show the normal characteristics observed in the MF-CA3 pathway of +/+ rats; that is, the negative correlation between the initial PPF and the changes in PPF measured after the LTP. These findings suggest an involvement of SCF/c-kit signaling in hippocampal synaptic potentiation and spatial learning and memory. PMID- 11112798 TI - Blockade of NMDA receptors in the amygdala prevents latent inhibition of fear conditioning. AB - The association between a conditioned stimulus (CS) and an unconditioned stimulus (US) in fear-conditioning depends on N-methyl-D-aspartate (NMDA) receptors in the basolateral amygdala complex (BLA). Latent inhibition (LI) is the retardation in learning due to nonreinforced presentation of the prospective CS before conditioning. Disruption of LI in rats is an animal model of schizophrenia, reflecting the deficits of schizophrenic patients in neglecting irrelevant information. We investigated whether the BLA is involved in LI of fear potentiated startle. Infusions of the NMDA receptor antagonist D,L-2-amino-5 phosphonopentanoic acid (AP-5; 12.5 nmoles) into the BLA before preexposure of rats to the neutral stimulus prevent LI of fear-conditioning. We also demonstrated by the same method that a complex of thalamic nuclei, comprising the medial part of the medial geniculate nucleus, the posterior intralaminar nucleus, and the suprageniculate nucleus, is involved in fear-conditioning, but not in LI. This suggests that the presentation of an innocuous stimulus during preexposure leads to an NMDA receptor-dependent change of neurotransmission in the BLA, but not in the thalamus. Our data show that the BLA but not the thalamus regulates in LI of fear-potentiated startle. Furthermore, it supports the hypothesis that the inability of schizophrenic patients to ignore irrelevant stimuli may be caused by hypofunction of the glutamatergic transmission in the brain and suggests an involvement of the amygdala in the neuropathology of schizophrenia. PMID- 11112799 TI - Cytokine responses to LTP induction in the rat hippocampus: a comparison of in vitro and in vivo techniques. AB - Because exogenous application of a number of cytokines and growth factors can alter synaptic properties, we sought to determine if endogenous cytokine expression is affected by neuronal activity. In addition, we examined whether cytokine expression is altered by the techniques used to stimulate and record from hippocampal neurons. Using semi-quantitative RNase protection and RT-PCR assays, we studied the expression of 18 cytokine, growth factor, and receptor genes in the hippocampus following the induction of Schaffer collateral-CA1 long term potentiation (LTP). We found that various cytokines are dramatically induced following preparation of slices for in vitro recording and as a result of injury following acute electrode placement in vivo. These increases can be overcome in vivo, however, using permanent electrodes implanted three weeks prior to testing. Using this chronic preparation, we found that interleukin-6 (IL-6) mRNA was upregulated nearly 20-fold by LTP induction in vivo, marking the first demonstration of endogenous regulation of this cytokine in response to LTP. In situ hybridization for IL-6 revealed that upregulation is tightly localized near the site of stimulation and is detected only in non-neuronal cells, identified as GFAP+ astrocytes and GFAP- cells within proximal blood vessels. Coupled with previous results showing that exogenously applied IL-6 can prevent the induction of LTP, this finding suggests a mechanism by which the local release of a cytokine could regulate LTP at nearby sites. PMID- 11112800 TI - Isoproterenol increases CREB phosphorylation and olfactory nerve-evoked potentials in normal and 5-HT-depleted olfactory bulbs in rat pups only at doses that produce odor preference learning. AB - Norepinephrine (NE) and serotonin (5-HT) are important modulators of early odor preference learning. NE can act as an unconditioned stimulus (UCS), whereas 5-HT facilitates noradrenergic actions. In this study, we examined the phosphorylation of an important transcription factor, cAMP response element binding protein (CREB), which has been implicated in long-term-memory formation (McLean et al. 1999) during NE-induced odor preference learning in normal and olfactory bulb 5 HT-depleted rat pups. We also examined NE modulation of olfactory nerve-evoked field potentials (ON-EFPs) in anesthetized normal and bulbar 5-HT depleted pups. Systemic injection of 2 mg/kg isoproterenol (beta-adrenoceptor agonist) induced odor preference learning, enhanced pCREB expression in the olfactory bulbs at 10 min after odor pairing, and increased ON-EFPs in normal rat pups but not in bulbar 5-HT-depleted rat pups. A dose of 6 mg/kg isoproterenol, which was ineffective in modulating these measures in normal rat pups, induced odor preference learning, enhanced phosphorylated CREB (pCREB) expression, and increased ON-EFPs in bulbar 5-HT-depleted pups. These outcomes suggest that NE and 5-HT promote specific biochemical and electrophysiological changes that may critically underlie odor preference learning. PMID- 11112801 TI - Optical recording of odor-evoked responses in the olfactory brain of the naive and aversively trained terrestrial snails. AB - Regular spontaneous oscillations were recorded both electro- and optophysiologically using a voltage-sensitive absorption dye in the olfactory part of the brain (procerebral lobe of the cerebral ganglia) of the gastropod mollusk Helix lucorum. Odor application caused transient changes in procerebral oscillations, and an odor-evoked potential was recorded in the procerebrum (PC). The wave of evoked potential originated near the place of olfactory nerve entrance into the PC and propagated via the procerebral neuropile toward the cell body layer. The spread of the odor-evoked potential corresponded roughly to the neuropile area, whereas the spontaneous oscillations were recorded in the cell body layer of the PC and were not observed in the neuropile. Evoked potential did not produce additional events intercalated into the ongoing spontaneous oscillations. Changes in parameters of spontaneous oscillations to the repeated presentations of the same odor were variable. To estimate the role of spontaneous oscillations in odor encoding, we trained the snail to avoid cineole, using paired presentations of cineole and electric shock. Elaboration of conditioned aversion to cineole applications resulted in distinct pairing-specific changes in behavior of the snails and procerebral activity. Responses to odor (cineole) applications were not different in amplitude or frequency of spontaneous oscillations in control and trained snails, whereas ratio of amplitudes of the same oscillation wave in proximal and distal regions of the procerebrum was significantly different in control and aversively trained snails, reflecting changes in neural firing in certain areas of the olfactory lobe. PMID- 11112802 TI - Imaging high-resolution structure of GFP-expressing neurons in neocortex in vivo. AB - To detect subtle changes in neuronal morphology in response to changes in experience, one must image neurons at high resolution in vivo over time scales of minutes to days. We accomplished this by infecting postmitotic neurons in rat and mouse barrel cortex with a Sindbis virus carrying the gene for enhanced green fluorescent protein. Visualized with 2-photon excitation laser scanning microscopy, infected neurons showed bright fluorescence that was distributed homogeneously throughout the cell, including axonal and dendritic arbors. Single dendritic spines could routinely be resolved and their morphological dynamics visualized. Viral infection and imaging were achieved throughout postnatal development up to early adulthood (P 8-30), although the viral efficiency of infection decreased with age. This relatively noninvasive method for fluorescent labeling and imaging of neurons allows the study of morphological dynamics of neocortical neurons and their circuits in vivo. PMID- 11112805 TI - Congratulations to the 2000 editorial fellows PMID- 11112803 TI - Fornix and hippocampal atrophy in traumatic brain injury. AB - This study compared a fornix cross-sectional-area measurement and hippocampal volume in 86 traumatic brain injury (TBI) subjects with 46 normal controls. The TBI group showed a significant reduction in fornix area and hippocampal volume. It was also shown that initial injury severity was related to the degree of atrophy in both structures. Although fornix size and hippocampal volume correlated, such a modest correlation between these two structures suggests differential and potentially independent mechanisms of injury. The General Memory Index score from the Wechsler Memory Scale-Revised was shown to be significantly correlated with hippocampal volume following TBI. PMID- 11112806 TI - CT of blunt trauma bowel and mesenteric injury: typical findings and pitfalls in diagnosis. AB - Detection of bowel and mesenteric injury can be challenging in patients after blunt abdominal trauma. Early diagnosis and treatment are critical to decrease patient morbidity and mortality. Computed tomography (CT) has become the primary modality for the imaging of these patients. Signs of bowel perforation such as free air and contrast material are virtually pathognomonic. Bowel-wall thickening, free fluid, and mesenteric infiltration may be seen with this type of injury and partial thickness injuries. The authors present and discuss the range of CT findings seen with bowel and mesenteric injuries. Examples of observation and interpretation errors are also provided to highlight pitfalls encountered in the evaluation of abdominopelvic CT scans in patients after blunt trauma. PMID- 11112807 TI - Invited commentary PMID- 11112808 TI - Practice corner: advice from the expert. PMID- 11112809 TI - MR imaging of cervical carcinoma: a practical staging approach. AB - Cervical carcinoma is the third most common gynecologic malignancy and is typically seen in younger women, often with serious consequences. The International Federation of Gynecology and Obstetrics (FIGO) staging system provides worldwide epidemiologic and treatment response statistics. However, there are significant inaccuracies in the FIGO staging system, and magnetic resonance (MR) imaging, although not included in that system, is now widely accepted as optimal for evaluation of important prognostic factors such as lesion volume and metastatic lymph node involvement that will help determine the treatment strategy. MR imaging examination obviates the use of invasive procedures such as cystoscopy and proctoscopy, especially when there is no evidence of local extension. Brachytherapy and external beam therapy are optimized with MR imaging evaluation of the shape and direction of lesion growth. In general, T2-weighted MR imaging more clearly delineates cervical carcinoma and is preferred for evaluation of the lymph nodes. Dynamic gadolinium-enhanced T1 weighted imaging may help identify smaller tumors, detect or confirm invasion of adjacent organs, and identify fistulous tracts. MR imaging staging, when available, is invaluable for identifying important prognostic factors and optimizing treatment strategies. PMID- 11112810 TI - Comprehensive MR imaging of acute gynecologic diseases. AB - Rapid advances in techniques of magnetic resonance (MR) imaging have enabled diagnosis of acute gynecologic conditions, which are characterized by sudden onset of lower abdominal pain, fever, genital bleeding, intraperitoneal bleeding, or symptoms of shock. The chemical-selective fat-suppression technique not only helps establish the characteristics of lesions that contain fat components but also increases the conspicuity of inflammatory lesions. When a T2-weighted image is obtained with a very long effective echo time (>250 msec), even a small amount of ascites can be easily identified and the contrast between urine and complex fluid becomes more conspicuous. T2*-weighted images are useful for identification of hemorrhagic lesions by demonstrating deoxyhemoglobin and hemosiderin. Contrast material-enhanced dynamic subtraction MR imaging performed with a three dimensional fast field-echo sequence and a rapid bolus injection of gadopentetate dimeglumine allows evaluation of lesion vascularity and the anatomic relationship between pelvic vessels and a lesion and allows identification of the bleeding point by demonstrating extravasation of contrast material. To optimize the MR imaging examination, attention should be given to the parameters of each pulse sequence and proper combination of the sequences. PMID- 11112811 TI - Dynamic MR imaging of pelvic organ prolapse: spectrum of abnormalities. AB - Pelvic organ prolapse is a relatively common condition in women that can have a significant impact on quality of life. Pelvic organ prolapse typically demonstrates multiple abnormalities and may involve the urethra, bladder, vaginal vault, rectum, and small bowel. Patients may present with pain, pressure, urinary and fecal incontinence, constipation, urinary retention, and defecatory dysfunction. Diagnosis is made primarily on the basis of findings at physical pelvic examination. Imaging is useful in patients in whom findings at physical examination are equivocal. Fluoroscopy, ultrasonography, and magnetic resonance (MR) imaging can be useful in evaluating pelvic organ prolapse. Advantages of MR imaging include lack of ionizing radiation, depiction of the soft tissues of the pelvic floor, and multiplanar imaging capability. Dynamic imaging is usually necessary to demonstrate pelvic organ prolapse, which may be obvious only when abdominal pressure is increased. Treatment is more likely to be successful if a survey of the entire pelvis is performed prior to therapy. Therapy is usually undertaken only in symptomatic patients. In all patients, imaging findings must be interpreted in conjunction with physical examination findings and the patient's symptoms. PMID- 11112812 TI - Why IHE? PMID- 11112814 TI - Breast US in children and adolescents. AB - Ultrasonography (US) is of value in the evaluation and characterization of breast masses in children. Most masses represent either normal breast tissue, cysts, or fibroadenomas. Premature thelarche may be unilateral, and normal breast tissue is found at US. Cysts are commonly retroareolar; when they become infected, they appear sonographically as a complex mass. Fibroadenoma is the most frequent breast tumor in adolescent girls, and it is usually solitary, homogeneous, and hypoechoic. Malignant breast lesions are very rare in children; most are due to metastatic disease secondary to rhabdomyosarcoma, leukemia, lymphoma, and neuroblastoma, and their US appearance is nonspecific. Gynecomastia in boys can be mimicked by general obesity and pectoral hypertrophy; US is helpful in the diagnosis, especially when gynecomastia is asymmetric. Most breast lesions in children and adolescents are benign, and surgery should be avoided to prevent later deformity. US is the ideal imaging modality to evaluate breast lesions and may be used to guide a fine-needle aspiration biopsy. Color Doppler US evaluation is helpful; cysts are avascular, fibroadenomas may be avascular or hypovascular, and abscesses show peripheral increased flow. Bloody nipple discharge is more common in prepubertal patients, may occur in infants, and may be secondary to mammary ductal ectasia. Discharge commonly resolves spontaneously, and findings at US are frequently normal. PMID- 11112813 TI - Pediatric renal masses: Wilms tumor and beyond. AB - A variety of pediatric renal masses may be differentiated from Wilms tumor on the basis of their clinical and imaging features. Wilms tumor is distinguished by vascular invasion and displacement of structures and is bilateral in approximately 10% of cases. Nephroblastomatosis occurs most often in neonates and is characterized by multiple bilateral subcapsular masses, often associated with Wilms tumors. Renal cell carcinoma is unusual in children except in association with von Hippel-Lindau syndrome and typically occurs in the 2nd decade. Mesoblastic nephroma is the primary consideration in a neonate with a solid renal mass. Multilocular cystic renal tumor is suggested by a large mass with multiple cysts and little solid tissue. Clear cell sarcoma is distinguished by frequent skeletal metastases, and rhabdoid tumor is distinguished by its association with brain neoplasms. Angiomyolipoma frequently contains fat and is associated with tuberous sclerosis. Renal medullary carcinoma occurs in patients with sickle cell trait or hemoglobin SC disease and manifests as an infiltrative mass with metastases. Ossifying renal tumor of infancy is differentiated from mesoblastic nephroma by the presence of ossified elements. Metanephric adenoma lacks specific features but is always well defined. Renal lymphoma is characterized by multiple homogeneous masses, often with associated adenopathy. PMID- 11112816 TI - MRI videotape series 8850 AB - Minnetonka, Minn: EduMed, 1999. Videotape, $95.00 for each tape. PMID- 11112815 TI - Spectrum of US findings in pediatric and adolescent patients with palpable breast masses. AB - Palpable breast masses arising in pediatric and adolescent patients are uncommon. A careful physical examination should be performed first, followed by an ultrasonographic evaluation when a suspect mass is present. In this study population, palpable findings were all due to benign causes, which is concordant with the literature. Benign causes included gynecomastia, cyst, fibroadenoma, lymph node, galactocele, duct ectasia, and infection. Though extremely rare, breast malignancies do occur in the pediatric and adolescent population. PMID- 11112817 TI - Thoracic manifestations of systemic autoimmune diseases: radiographic and high resolution CT findings. AB - The systemic autoimmune diseases include collagen vascular diseases, the systemic vasculitides, Wegener granulomatosis, and Churg-Strauss syndrome. They can cause a variety of thoracic abnormalities that are influenced by the pathophysiologic characteristics of the underlying disease process. Although many of the abnormalities can be detected at chest radiography, high-resolution computed tomography (CT) has been shown to be superior in depicting parenchymal, airway, and pleural abnormalities. Thoracic manifestations of collagen vascular diseases include pleural disease, pulmonary fibrosis, diaphragm weakness, aspiration pneumonia, bronchiolitis obliterans organizing pneumonia, bronchiolitis obliterans, and bronchiectasis. Wegener granulomatosis may be associated with multiple nodules or masses with irregular margins that are frequently cavitated. Patients with Churg-Strauss syndrome often have consolidation or ground-glass attenuation at chest radiography and CT. Goodpasture syndrome is associated with extensive bilateral air-space consolidation. PMID- 11112818 TI - CT of heart transplant recipients: spectrum of disease. AB - The emergence of heart transplantation as the ultimate treatment for end-stage heart failure has been accompanied by new diagnostic challenges. Computed tomography (CT) has emerged as an important diagnostic tool in the evaluation of heart transplant recipients because many infectious, ischemic-hemorrhagic, and neoplastic complications are amenable to early detection with this modality. In the early postoperative period, CT is mostly indicated in the evaluation of infectious complications or cerebral symptoms. Later, CT is mostly performed for staging of infectious or neoplastic disease. Infectious complications include mediastinitis, soft-tissue inflammation, abscess formation, cerebral infarction, and aspergillosis. Complications related to ischemia or hemorrhage include allograft rejection and coronary allograft vasculopathy, the latter being the leading long-term cause of death in heart transplant recipients. CT is also indicated in malignant disease (eg, lymphoma, visceral carcinoma, skin tumors), which is the second most important long-term cause of death. Moreover, CT is helpful in identifying disease caused by immunosuppressive therapy (eg, leukoencephalopathy, osteoporosis, thoracic lipomatosis). CT has proved superior to both ultrasound and magnetic resonance imaging in the evaluation of heart transplant recipients. It has become the diagnostic modality of choice for many transplant-related complications and may help improve postoperative treatment of affected patients. PMID- 11112819 TI - Role of radionuclide imaging in the diagnosis of postoperative infection. AB - Postoperative infections are a serious cause of morbidity and mortality and are difficult to diagnose. Signs and symptoms that are generally associated with infection may be masked by, or mistaken for, normal postoperative changes. Anatomic imaging modalities provide high-quality anatomic detail and are the procedures of choice in affected patients because of their availability, ease of performance, accuracy, and value in the selection of treatment options. However, radionuclide studies demonstrate physiologic processes, which often precede anatomic changes, and can help distinguish normal postoperative inflammation from infection. Radionuclide studies are also useful in identifying complicated orthopedic infections, in which the often extensive distortions produced by metallic hardware can confound the interpretation of anatomic images. Of the three agents (gallium-67 citrate, indium-111-labeled leukocytes, technetium-99m labeled leukocytes) that are currently approved in the United States for imaging of infection, In-111-labeled leukocyte imaging is the procedure of choice for diagnosing postoperative infection. Gallium scintigraphy is best reserved for those situations in which leukocyte imaging is not available or there is concern that the suspected infection may not incite a neutrophil response. In general, the value of radionuclide imaging is maximized when used only in those patients for whom the results of anatomic imaging are negative, nondiagnostic, or at odds with the clinical impression. PMID- 11112820 TI - Invited commentary PMID- 11112821 TI - Imaging of surgical paraphernalia: what belongs in the patient and what does not. AB - Many radiologists are not familiar with the names of various instruments, surgical sponges, and needles that may be seen on intraoperative and postoperative radiographs. These devices may be intentionally placed for localization or therapeutic intervention, discovered on radiographs obtained to evaluate incorrect sponge or needle counts, or incidentally encountered on postoperative radiographs. These paraphernalia are usually described in vague nonspecific terms in radiology reports. In this article, photographs and radiographs of several instruments commonly used for intraoperative localization or therapy are presented, as well as examples of sponges, needles, and other devices that should not be found on postoperative radiographs. Familiarity with their appearances will allow a more precise and knowledgeable description in radiology reports. PMID- 11112822 TI - Motion artifacts in subsecond conventional CT and electron-beam CT: pictorial demonstration of temporal resolution. AB - To visually demonstrate the effective temporal resolution of subsecond conventional (slip-ring) and electron-beam computed tomographic (CT) systems, two phantoms containing high-contrast test objects were scanned with a slip-ring CT system (effective exposure time, 0.5 second) and an electron-beam CT system (exposure time, 0.1 second). Images were acquired of each phantom at rest, during translation along the x axis at speeds of 10-100 mm/sec, and during rotation about isocenter at speeds of 0.1 and 0.5 revolution per second. Motion artifacts and loss of spatial resolution were judged to be absent, noticeable, or severe. For 0.5-second conventional CT images, motion artifacts and loss of spatial resolution were noticeable at 10 mm/sec and 0.1 revolution per second and were severe at speeds greater than or equal to 20 mm/sec and at 0.5 revolution per second. For 0.1-second electron-beam CT scans, noticeable, but not severe, motion artifacts and loss of spatial resolution occurred at speeds between 40 and 100 mm/sec and at 0.5 revolution per second. Over the range of physiologic speeds examined, the images provide visually compelling evidence of the effect of improving temporal resolution in CT. PMID- 11112823 TI - A note of thanks PMID- 11112824 TI - Vertebral artery dissection: spectrum of imaging findings with emphasis on angiography and correlation with clinical presentation. AB - A study was performed to evaluate the relationship between the imaging features and clinical presentation of vertebral artery (VA) dissection. Twenty-two patients with 24 VA dissections at angiography and clinical evaluation also underwent computed tomography and magnetic resonance imaging. The angiographic patterns of VA dissection were categorized as aneurysmal (n = 10) or steno occlusive (n = 14). All 10 patients (10 lesions) with the aneurysmal pattern had dissection in the V4 (intradural) segment and presented with headache (n = 5), neurologic deficit (n = 2), dizziness (n = 2), or altered mentality (n = 1). However, the 12 patients (14 lesions) with the steno-occlusive pattern had dissection from the V1 segment to the V4 segment and presented with neurologic deficits caused by infarction of an embolic nature. Overall, the most frequent VA dissection site was the V4 segment. The distribution of the dissection sites and the clinical presentation tended to differ according to the angiographic patterns of aneurysm or stenosis-occlusion. PMID- 11112825 TI - Advances in imaging of lymph flow disorders. AB - Conventional oil-contrast lymphography has long been the mainstay for lymphatic imaging. However, the emergence of computed tomography (CT) and magnetic resonance (MR) imaging has severely curtailed its use. Because of recent improvements and refinements, lymphangioscintigraphy now permits high-resolution imaging of peripheral lymphatic vessels and provides insight into lymph flow dynamics. It is indispensable for patients with known or suspected lymphatic circulatory disorders in confirming the diagnosis and delineating the pathogenesis and evolution of lymphedema. In addition, lymphangioscintigraphy helps evaluate lymphatic truncal anatomy and radiotracer transport. It can also be used to evaluate the efficacy of various treatment options designed to facilitate lymph flow or reduce lymph formation. The procedure is essentially noninvasive, can easily be repeated, and does not adversely affect the lymphatic vascular endothelium. MR imaging complements lymphangioscintigraphy in the monitoring and treatment of more complex lymphatic circulatory disorders, whereas CT facilitates catheter-guided percutaneous sclerosis or obliteration of specific lymphangiectasia or lymphangioma syndromes. Ultrasonography has proved useful in the setting of filariasis. Patients with a provisional diagnosis of peripheral lymphatic dysfunction or idiopathic edema should undergo diagnostic lymphangioscintigraphy and, in some cases, MR imaging to verify diagnostic accuracy, pinpoint the specific abnormality, and help guide subsequent therapy. PMID- 11112826 TI - Neoplasms of the spinal cord and filum terminale: radiologic-pathologic correlation. AB - Intramedullary spinal cord neoplasms are rare, accounting for about 4%10% of all central nervous system tumors. Despite their rarity, these lesions are important to the radiologist because magnetic resonance (MR) imaging is the preoperative study of choice to narrow the differential diagnosis and guide surgical resection. On contrast materialenhanced MR images, intramedullary spinal tumors almost always manifest as expansion of the spinal cord and show enhancement. Syringohydromyelia and cystic lesions are frequently associated with intramedullary tumors. Nontumoral cysts tend to be located at the poles of the tumors and do not enhance on contrast-enhanced MR images, whereas cysts within the substance of the tumor are considered tumoral cysts and typically demonstrate peripheral enhancement. Spinal cord ependymomas are the most common type in adults, and cord astrocytomas are most common in children. Both entities constitute up to 70% of all intramedullary neoplasms. A central location within the spinal cord, presence of a cleavage plane, and intense homogeneous enhancement are imaging features that favor an ependymoma. Intramedullary astrocytomas are usually eccentrically located within the cord, are ill defined, and have patchy enhancement after intravenous contrast material administration. Even with these characteristics, it may not be possible to differentiate these two entities on the basis of imaging features alone. Cord hemangioblastomas are the third most common type of intramedullary spinal tumor. Gangliogliomas commonly extend over more than eight vertebral segments. Paragangliomas and primitive neuroectodermal tumors have an affinity for the filum terminale and cauda equina. Other spinal cord tumors include metastatic disease, which is characterized by prominent cord edema for the size of the enhancing portion, and primary lymphoma. PMID- 11112827 TI - A review of selective salpingography and fallopian tube catheterization. AB - Use of selective salpingography and fallopian tube recanalization has revolutionized the diagnosis and treatment of infertility. Selective salpingography, a diagnostic procedure in which the fallopian tube is directly opacified through a catheter placed in the tubal ostium, has been used since the late 1980s to differentiate spasm from true obstruction and to clarify discrepant findings from other tests. In fallopian tube recanalization, a catheter and guide wire system is used to clear proximal tubal obstructions. The recanalization procedure is simple for interventional radiologists to perform and is successfully completed in most patients (71%-92%). Pregnancy rates after the procedure have been variable, with an average rate of 30%. The combination of selective salpingography with fallopian tube recanalization has improved the overall management of infertility caused by tubal obstruction. The same catheterization technique used in fallopian tube recanalization is currently being explored for use in tubal sterilization. PMID- 11112828 TI - The AAPM/RSNA physics tutorial for residents: fluoroscopy: optical coupling and the video system. AB - In fluoroscopic/fluorographic systems, an image intensifier is optically coupled to recording cameras. The optical distributor is responsible for transmitting a focused image from the output phosphor of the image intensifier to the focal planes of the cameras. Each camera has an aperture, which is used to control the level of light reaching its focal plane. The aperture setting determines the patient x-ray exposure level and the image noise level. Increasing the x-ray exposure reduces image noise; reducing the x-ray exposure increases image noise. Fluoroscopic/fluorographic systems always include a video camera. The functions of the video system are to provide for multiple observers and to facilitate image recording. The camera head contains an image sensor, which converts the light image from the image intensifier into a voltage signal. The device used to generate the video signal is a pickup tube or a charge-coupled device sensor. The method used is raster scanning, of which there are two types: progressive and interlaced. The vertical resolution of the system is primarily determined by the number of scan lines; the horizontal resolution is primarily determined by the bandwidth. Frame rate reduction can be a powerful tool for exposure reduction. PMID- 11112829 TI - Multisection CT: scanning techniques and clinical applications. AB - Multisection computed tomography (CT) was introduced in 1992 with the advent of dual-section-capable scanners and was improved in 1998 following the development of quad-section technology. With a recent increase in gantry speed from one to two revolutions per second, multisection CT scanners are now up to eight times faster than conventional single-section helical CT scanners. The benefits of quad section CT relative to single-section helical CT are considerable. They include improved temporal resolution, improved spatial resolution in the z axis, increased concentration of intravascular contrast material, decreased image noise, efficient x-ray tube use, and longer anatomic coverage. These factors substantially increase the diagnostic accuracy of the examination. The multisection CT technique has enabled faster and superior evaluation of patients across a wide spectrum of clinical indications. These include isotropic viewing, musculoskeletal applications, use of multiplanar reformation in special situations, CT myelography, long coverage and multiphase studies, CT angiography, cardiac scoring, evaluation of brain perfusion, imaging of large patients, evaluation of acute chest pain or dyspnea, virtual endoscopy, and thin-section scanning with retrospective image fusing. Multisection CT is superior to single section helical CT for nearly all clinical applications. PMID- 11112830 TI - Computer-aided design and modeling of workstations and radiology reading rooms for the new millennium. AB - Three-dimensional (3D) computer modeling, simulation, and rendering techniques were used to redesign the diagnostic workstations and radiology reading rooms for a proposed hospital with particular attention given to lighting conditions, noise reduction, and optimal use of limited workspace. The results were presented to a panel of multidisciplinary experts and iteratively improved and redesigned with the development or addition of new design criteria or requirements. These 3D techniques allowed faster, more efficient design and presentation of multiple options than is possible with traditional two-dimensional drawings, thereby expediting decision making and resulting in significant savings. The current workstation designs can easily be developed and implemented with available technology at a reasonable cost. They can also accommodate anticipated advances in computer and display technology as well as new imaging paradigms (eg, changes in keyboard and control ergonomics such as adjustable virtual keys on touch sensitive screens, digital drawing tablets for annotations and controls, direct film digitizing, personal identification devices, offline media readers such as compact disks and digital videodisks, and speech recognition and voice activation). Use of 3D techniques in designing other parts of the radiology department (eg, examination rooms, technologists' areas, physicians' offices) could greatly improve and facilitate the design and implementation of complex settings in these work areas. PMID- 11112832 TI - Errata PMID- 11112831 TI - Application of a "Jini-enabled" patient data storage system. PMID- 11112833 TI - Neonatal cholestasis. AB - In simplest terms, cholestasis is defined as a decrease in bile flow. The clinical manifestations of cholestasis occur because of accumulation of substances normally excreted in the bile; namely bilirubin, bile acids, and cholesterol. Accumulation of bilirubin leads to jaundice and dark urine. Accumulation of bile acids is associated with pruritus, and accumulation of cholesterol causes hypercholesterolemia and xanthomas. There are many causes of cholestasis in early infancy ranging from normal physiologic jaundice to complete biliary obstruction associated with biliary atresia. PMID- 11112834 TI - Cholelithiasis and cholecystitis in children. AB - With advances in medical technology, including intensive care, new medications, alterations in the composition of parenteral nutrition, and the institution of minimally invasive surgery, our understanding of the spectrum of diseases of the gallbladder resulting in stone formation or inflammation, and the management of these disorders has changed over the past few decades. The discussion herein focuses on our thinking about the current diagnosis and treatment for these disorders. PMID- 11112835 TI - Biliary atresia. AB - Although the prognosis of biliary atresia has been dramatically improved in the era of liver transplantation, the Kasai operation is still the first line of surgical treatment. Successful hepatic portoenterostomy depends on early diagnosis and surgery, adequate surgical technique, prevention of cholangitis, and precise postoperative management. PMID- 11112836 TI - Congenital biliary dilatation. AB - Congenital biliary dilatation is commonly associated with pancreaticobiliary malunion. Its etiology remains unknown. With the advent of accurate cholangiography, combined abnormalities of the intrahepatic duct, common channel, and pancreatic duct are being identified more frequently in patients with congenital biliary dilatation. Early diagnosis followed by cyst excision is the treatment of choice, even in asymptomatic cases. The treatment of intrahepatic and intrapancreatic ductal diseases such as intrahepatic duct dilatation and stone debris in the intrahepatic duct and common channel are also discussed. The value of intraoperative endoscopy as an adjunct to cyst excision for the prevention of postoperative complications is explained. PMID- 11112837 TI - Liver tumors. AB - Liver tumors in children are rare, potentially complex, and encompass a broad spectrum of disease processes. Any age group may be affected, including the fetus. Most present with abdominal distension and/or a mass. Accurate preoperative diagnosis is usually possible using a combination of ultrasound scanning and cross-sectional imaging techniques (CT and/or MR), supplemented by liver biopsy and measurement of tumor markers. The most common benign tumors are hemangiomas, but mesenchymal hamartoma, focal nodular hyperplasia, and adenoma also are found. In Western countries, hepatoblastoma is the most common primary malignant liver tumor; disease-free survival is now possible in more than 80% of affected patients because of advances in combination chemotherapy, improved techniques of surgical resection, and the selective use of liver transplantation. In contrast, there has been less progress in the management of hepatocellular cancer, which still poses many therapeutic challenges. PMID- 11112838 TI - Cysts and tumors of the pancreas. AB - Both pancreatic cysts and pancreatic tumors rarely occur in childhood. However, when encountered they can present a diagnostic and therapeutic challenge to the pediatric surgeon. The aim of this review is to discuss the different types of pancreatic cysts and tumours that may be encountered in the pediatric population, to note the modes available for their diagnosis, and to outline treatment options. PMID- 11112839 TI - Catalytic antibodies (abzymes) induced by stable transition-state analogs. AB - This review deals with recent advances in the generation of catalytic antibodies by the immunization of animals with stable transition-state analogs. Characteristic features in the functioning of such abzymes are considered in comparison with traditional enzymes. PMID- 11112840 TI - Natural catalytic antibodies (abzymes) in normalcy and pathology. AB - This review summarizes literature data on natural abzymes. Peculiar features of their functioning and substrate specificity are considered in comparison with traditional enzymes. Working hypotheses on the possible biological roles of natural abzymes in autoimmune processes and diseases accompanied by disorders of immune status are analyzed. PMID- 11112841 TI - Antigen-binding activity of monoclonal antibodies after incubation with organic solvents. AB - Effects of four organic solvents--methanol, trifluoroethanol, dimethylsulfoxide, and dimethylformamide (DMF)--on the ferritin-binding activity of three monoclonal mouse antibodies of IgG2a and IgG1 subclasses were studied. The ferritin-binding constants of monoclonal antibodies G10 and F11 (the IgG2a subclass) were increased 2-6-fold after incubation with DMF and removal of the organic solvent by gel filtration. The maximum effect on the F11 antibodies was found in the presence of 5-13% DMF and on the G10 antibodies at 11-40% DMF. The effect remained after the removal of DMF from the incubation medium, and this suggests that the incubation with DMF resulted in irreversible conformational changes of the antibodies and in production of active conformers of the G10 and F11 antibodies. These conformations occurred within 15-60 min. The long-term stability and the fluorescence of the antibodies exposed to DMF suggest that the conformational changes were not global, but involved small and relatively independent structural elements of the antibodies, either of hypervariable CDR loops in variable domains or of the hinge region of the antibodies. The affinity of the C5 antibodies of the mouse IgG1 subclass was decreased after incubation with DMF. The activation was a solvent-specific effect because incubation of the G10 antibodies with methanol and dimethylsulfoxide decreased the affinity for the antigen, and incubation with trifluoroethanol virtually did not affect it. Relatively small changes in the antigen-binding activity of the antibodies were found even after the incubation with 5% organic solvent. PMID- 11112842 TI - The widely accepted model of primary photosynthetic processes in purple bacteria must be revised. AB - Using computer simulation, it was found that, at least in case of purple bacteria, the widely accepted model for the primary photosynthetic processes (energy migration to reaction centers and its further trapping) is unable to provide a fully consistent explanation for the available experimental data. Two physical mechanisms suggested in this work are thought to be able to resolve or at least decrease the severity of this problem. PMID- 11112843 TI - Synthesis of lipopolysaccharides in the bacterium Yersinia pseudotuberculosis: effect of the pVM82 plasmid and growth temperature. AB - The composition and structure of lipopolysaccharides (LPS) of three isogenic strains of Yersinia pseudotuberculosis serovar O:1b (without plasmids (82-) and with plasmids pVM82 (82+) or p57 (57+)) grown at 8 or 37 degrees C were studied by chemical and immunochemical methods, SDS-polyacrylamide gel electrophoresis, and 13C-NMR spectroscopy. At the lower temperature, the (82-) and (82+) strains synthesized S-form of LPS with similar structure characterized by high acylation and immunochemical activity. On the other hand, LPS of the (82+) strain had shorter carbohydrate chains than LPS of the (82-) strain. The contents of LPS were decreased in cells of the plasmid-free strain grown at the higher temperature. LPS isolated from these cells were of the R-form and had low acylation and immunochemical activity. Total LPS content in cells of the (82+) strain did not significantly depend on the growth temperature. LPS of the warm variant of these bacteria contained a polysaccharide fragment and had moderate immunochemical activity. The cells of the (57+) strain at both growth temperatures had low LPS contents and produced LPS of low acylation without O specific chains (cold variant) or containing O-polysaccharide with low polymerization degree (bacteria grown at 37 degrees C). The data indicate that in the absence of the plasmids, LPS synthesis is encoded by the chromosomal genes in pseudotuberculosis bacteria. Expression of the genes involved in LPS synthesis is regulated by the temperature of bacterial growth. Genes responsible for temperature-dependent regulation of LPS biosynthesis are located on chromosomal DNA. The pVM82 plasmid includes two gene groups; one group is localized in a 57 mD fragment of DNA and inhibits LPS synthesis, suppressing temperature-dependent regulation of the synthesis. The genes located in a 25-mD fragment of the pVM82 plasmid are de-repressors of the 57-mD fragment, and they restore the ability of pseudotuberculosis bacteria to synthesize relatively long LPS at both growth temperatures. PMID- 11112844 TI - Hydrogen peroxide-induced DNA repair and death of resting human blood lymphocytes. AB - DNA single-strand breaks induced by cell treatment with hydrogen peroxide are repaired and simultaneously trigger programmed cell death in resting human blood lymphocytes. Apoptosis is accompanied by special morphological changes in lymphocytes (15% of total cell number), internucleosomal DNA degradation, and p53 level elevation. According to morphological criteria, a major part (up to 40% of total cell number) displayed necrotic death features. Nicotinamide inhibited repair in cells with 2.5-fold elevation of the apoptotic cell proportion, whereas the fraction of cells with necrotic nuclear morphology decreased 4.5-fold. Both the inhibition of repair and the protective effect of nicotinamide against necrotic death indicate that the repair process and related poly(ADP ribose)polymerase (PARP) activation induce a decrease in intracellular NAD+ and ATP contents below the threshold at which necrosis becomes the preferential mechanism of cell death. The mixed pattern of cell death induced by hydrogen peroxide observed in resting lymphocytes can be explained in the context of a concept of cell de-energization as a consequence of effective single-stand break repair during the first hours after removing the genotoxic agent. PMID- 11112845 TI - Influence of metal ions on hydrogenase from the purple sulfur bacterium Thiocapsa roseopersicina. AB - The effects of some metal ions on the activity and activation of Thiocapsa roseopersicina hydrogenase have been studied. Inhibitory effects of Ni2+ and Cd2+ on the catalytic activity of the enzyme were reversible and competitive with respect to methyl viologen (MV) in the reaction of hydrogen oxidation. The affinity of these metal ions to the enzyme increased significantly with increasing pH, suggesting that their interactions are determined by electrostatic forces. Cu2+ and Hg2+ irreversibly inhibited the hydrogenase activity. A decrease in absorption of hydrogenase at 400 nm in the presence of these metal ions is indicative of the destruction of the FeS cluster in the enzyme. PMID- 11112846 TI - Active oxygen-associated control of rice blast disease by riboflavin and roseoflavin. AB - Exogenous riboflavin and its dimethylated amino(nor)-derivative roseoflavin were studied in their ability to protect susceptible rice plants from blast disease and to induce fungitoxicity mediated by active oxygen. Both compounds, either added to the inoculum (10 microg/ml) or to soil (40 mg/kg, two days prior to inoculation), induced disease resistance, i.e., diminished the frequency of compatible-type lesions on infected leaves, mainly at the expense of the appearance of hypersensitive spots. Leaf diffusates of untreated plants possessed a weak fungitoxicity that increased slightly after leaf infection or illumination of diffusate. The flavins added to inoculum, to soil, or to a collected diffusate augmented significantly the light-activated part of the diffusate toxicity. In some instances, the light-independent part was stimulated as well. The effect was not due to direct fungitoxicity of flavins as they alone did not interfere with spores regardless of illumination. Antioxidant reagents (superoxide dismutase, catalase, scavengers of hydroxyl radical, and the iron ion chelator desferrioxamine) protected spores from intoxication in almost all cases. This implies the involvement of active oxygen in the toxic and, probably, disease controlling effects of the flavins. Roseoflavin was a better inducer of disease resistance than riboflavin but was similar in stimulation of diffusate toxicity. However, roseoflavin did not produce superoxide and exhibited only weak fungitoxicity if substituted for riboflavin in the well-known O2--generating model photosystem containing methionine. Therefore, the superoxide generation due to photo-oxidation of methionine or similar substrates is not the cause of the increase of leaf diffusate fungitoxicity and of disease resistance of plants supplied with roseoflavin. It is suggested that the rise in active oxygen production favors a forthcoming hypersensitive reaction, and both phenomena contribute to resistance induced by flavo-compounds. The light-driven activation of oxygen may be of interest as a mode of action of novel fungicides. PMID- 11112847 TI - Cytotoxic activity, accumulation, and intracellular distribution of anthracycline antibiotics and their conjugates with the epidermal growth factor in sensitive and resistant MCF-7 cells. AB - Cytotoxic activities, accumulation levels and dynamics, and intracellular distribution of the anthracycline antibiotics doxorubicin (DR) and carminomycin (CM) in the free forms or within conjugates with the epidermal growth factor (EGF) were for the first time compared in human breast carcinoma cell lines MCF 7Wt and MCF-7AdrR. The cytotoxic activities of DR and CM conjugates with EGF were higher than the cytotoxic activities of the free antibiotics in both cell lines. The accumulation levels of the free anthracyclines in both cell lines were lower than those of the conjugates and significantly depended on the cell sensitivities to the antibiotics. On receptor-mediated endocytosis of the anthracycline-EGF conjugates, the accumulation levels did not significantly depend on the cell sensitivities to the antibiotics. Both DR and CM, either free or conjugated with EGF, were mainly accumulated in nuclei. The free drugs were accumulated more rapidly, and the accumulation rates of both free and EGF-conjugated CM were higher than those of DR preparations. The intracellular distribution of the free antibiotics significantly depended on the cell sensitivities to the anthracyclines, whereas the cell sensitivities had no effect on the distribution of the conjugates between the nucleus and cytoplasm. The rate of intracellular degradation of DR and CM delivered to target cells within conjugates with EGF was twice lower than that of the free antibiotics. The difference in the accumulation levels and dynamics and in the intracellular distribution of the free and conjugated DR and CM is likely to underlie the higher cytotoxic activities of the anthracycline conjugates with EGF compared to the free drugs. PMID- 11112848 TI - Isolation and properties of peroxidase produced by the fungus Panus tigrinus. AB - Synthesis of peroxidase and laccase by the fungus Panus tigrinus was significantly stimulated by addition of the lignocellulose substrate to the culture media. Peroxidase was isolated from the culture liquid and some properties of the enzyme were investigated. P. tigrinus peroxidase belongs to a group of extracellular peroxidases similar to the plant type peroxidases. PMID- 11112849 TI - Effects of alcohol on the major steps of reverse cholesterol transport. AB - The effects of moderate alcohol consumption on the capacity of blood sera to promote acceptance of cholesterol (C) from Fu5AH hepatoma cells, esterification of delivered free C, and transfer of produced cholesteryl esters to apolipoprotein (apo) B-containing lipoproteins have been studied. Twenty male subjects with relatively high (>50 mg/dl, n = 10) and low (<50 mg/dl, n = 10) high density lipoprotein (HDL) C levels consumed for eight weeks red grape wine (0.3 g ethanol/kg body mass per day). Alcohol consumption reduced total C and low density lipoprotein C levels in both groups of subjects. Low HDL C subjects showed an increase in HDL C, apo AI, apo AII, and lipoprotein (Lp) AI particle levels after alcohol consumption. Alcohol did not affect free C efflux from the cells. However, after the following period of substitution of alcohol with an isocaloric amount of red grape juice, cellular C efflux markedly reduced. While lecithin:cholesterol acyltransferase (LCAT) activity increased during alcohol consumption only in subjects with low HDL C, high HDL C subjects showed a significant decrease in cholesteryl ester transfer protein (CETP) activity. At the same time, alcohol consumption reduced the endogenous C esterification rate and increased the transfer of endogenous cholesteryl esters to apo B-containing lipoproteins in both groups. Thus, alcohol consumption in moderate doses enhanced the anti-atherogenicity of the serum lipoprotein spectrum, supporting more effective C efflux from peripheral cells and transport of accepted C to apo B containing lipoproteins. The effects of alcohol on the reverse cholesterol transport depend on the initial HDL C level. PMID- 11112850 TI - Cellular localization of the galactose-binding lectin from human serum. AB - An SL2 lectin was isolated from human serum and characterized previously; cellular localization of the lectin was studied using polyclonal rabbit antibodies. According to cytofluorimetry, anti-SL2 antibodies bound only to lymphocytes and monocytes but not to other blood cells. Antibodies bound to Jurkat T cell lymphoma but did not interact with IM-9 cells of B cell origin. Moreover, the Jurkat cells bound oligosaccharides having the highest affinity to SL2 (GalNAcalpha_and Fucalpha1-2Gal), and this interaction was inhibited by anti SL2 antibodies. Lysis of the Jurkat cells with subsequent electrophoresis and Western blotting indicates that anti-SL2 antibodies recognized a 14-kD protein. PMID- 11112851 TI - Analysis of HI0220 protein from Haemophilus influenzae, a novel structural and functional analog of ArcB protein from Escherichia coli. AB - A Haemophilus influenzae gene encoding a protein with high homology to ArcB receptor protein from Escherichia coli has been cloned. An error in the previously reported sequence of this gene has been found, thus increasing its open reading frame. The cloned gene comprising the entire open reading frame restores oxygen-dependent regulation of succinate dehydrogenase in an ArcB deficient E. coli strain. Thus, this gene is a functional analog of ArcB from E. coli. By screening partially sequenced bacterial genomes using the BLAST program, proteins with high homology to ArcB protein from E. coli were found in Salmonella typhi, Yersinia pestis, Vibrio cholerae, and Pasteurella multocida. Comparison of these proteins with ArcB protein from E. coli and H. influenzae revealed conserved amino acid regions. Transmembrane helix II was shown to be highly homologous in all the ArcB-type proteins. The involvement of this region in ArcB mediated oxygen-dependent regulation is suggested. PMID- 11112852 TI - Molecular cloning and characterization of the promoter region of the inducible nitric oxide synthase gene of the rat. AB - To investigate the mechanism by which inducible nitric oxide synthase (iNOS) within vascular smooth muscle cells (VSMC) is expressed following exposure to proinflammatory cytokines, we cloned the promoter region of the rat iNOS gene by a single specific primer PCR cloning strategy. The 5;-flanking region of the rat iNOS gene contains interferon-gamma (IFN-gamma)- and tumor necrosis factor-alpha (TNF-alpha)-responsive elements and an NF-kappaB-binding consensus sequences. The position and alignment of these consensus sequences are distinct from those in the mouse iNOS and human iNOS genes. It was shown that some nuclear factor(s) which bound specifically to the fragment of the rat iNOS gene containing several consensus sequences were produced after VSMC were treated with interleukin 1 (IL 1) and IFN-gamma. PMID- 11112853 TI - A rapid effect of thyroxine on accumulation of diacylglycerols and on activation of protein kinase C in liver cells. AB - L-Thyroxine rapidly stimulated the accumulation of diacylglycerols in isolated hepatocytes and in liver when lipids were prelabeled with [14C]oleic acid or with [14C]CH3COONa. Perfusion of the liver of hypothyroid animals with L-thyroxine containing solution or incubation of liver fragments with the hormone increased the content of diacylglycerols in the liver cells. The increase in [14C]diacylglycerol level in the liver cells was accompanied by a decrease in the level of [14C]phosphatidylcholine, whereas contents of other 14C-labeled phospholipids, such as phosphatidylethanolamine, sphingomyelin, lysophosphatidylcholine, phosphatidylinositol (PtdIns), phosphatidylinositol-4 phosphate (PtdIns4P), and phosphatidylinositol-4,5-bis-phosphate (PtdIns(4,5)P2), and of 14C-labeled fatty acids were the same as in the control. The L-thyroxine induced accumulation of diacylglycerols in hepatocytes was not affected by neomycin but was inhibited by propranolol. Incubation of hepatocytes prelabeled with [14C]oleic acid with L-thyroxine and ethanol (300 mM) was accompanied by generation and accumulation of [14C]phosphatidylethanol that was partially suppressed by 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7). L-Thyroxine was responsible for the translocation of protein kinase C from the cytosol into the membrane fraction and for a many-fold activation of the membrane-bound enzyme. D Thyroxine failed to affect the generation of diacylglycerols in hepatocytes and the activity of protein kinase C. PMID- 11112854 TI - Evolution in the understanding of cross-reactivities of respiratory allergens: the role of recombinant allergens. AB - The aim of this review is to show the impact of the use of purified and recombinant allergens to discriminate between co- and cross-sensitization to respiratory allergens. The author describes the evolution of diagnostic tests over the last decades; the tests initially allowed the detection of simultaneously positive cutaneous tests and/or simultaneous positivity of specific IgE to different allergen extracts, but they did not differentiate cross sensitization from co-sensitization. RAST inhibition studies with crude extracts then established cross-reactivity, but did not identify the cross-reactive allergens involved. Later, immunoblot and CRIE inhibition were able to detect multiple cross-reactive allergens and to assess their physicochemical properties. But it is only since purified and recombinant allergens have been used in the different investigations that identification of cross-reactive allergens has been made possible at a molecular level. This historical approach is illustrated by examples selected from some of the main respiratory allergen sources: tree pollen, grass pollen, weed pollen, acarids, cockroaches and mammalians. For each of these allergen sources, the author gives an updated presentation of major and minor cross-reactive allergen molecules and refers to the last decade's major publications concerning immunochemical investigations carried out in the field of cross-reactive respiratory allergens. Emphasis is placed on the clinical applications for allergic patients: improvement in the accuracy of the diagnosis of sensitization, new concepts of immunotherapy based on genetically engineered hypoallergenic variants of cross-reactive allergens used alone or in combination, evaluation of allergen load with environmental tests using monoclonal antibodies against cross-reactive allergens. PMID- 11112855 TI - Anti-IgE therapy in asthma: rationale and therapeutic potential. AB - Airway inflammation is found in virtually all individuals with asthma symptoms. The factors contributing to asthma-related airway inflammation are multiple and involve a number of different inflammatory cells and mediators. Allergic responses at the level of the respiratory system are mostly mediated by IgE dependent mechanisms. After sensitization of a susceptible individual and the synthesis and binding of allergen-specific IgE to target cells, atopic individuals respond immunologically to common, naturally occurring allergens by releasing mast cell-derived mediators. Subsequent allergen exposure in susceptible individuals produces a characteristic cascade of events orchestrated by immune effector cells, most prominently, mast cells, T lymphocytes and eosinophils. A new strategy, neutralizing IgE antibodies, inhibits expression of allergic symptoms by preventing the initial trigger of the allergic reaction, IgE binding to IgE-receptor bearing cells. rhuMAb-E25 is a recombinant humanized monoclonal anti-IgE antibody currently under investigation that has been shown to reduce allergic responses in atopic individuals and to improve symptoms and reduce rescue medication and corticosteroid use in patients with allergic asthma. Thus, the clinical effectiveness of rhuMAb-E25 supports the central role of IgE in allergic reactions and the viability of anti-IgE therapy as a potentially effective treatment option for asthma. PMID- 11112856 TI - A soybean G2 glycinin allergen. 1. Identification and characterization. AB - BACKGROUND: Multiple allergens have been documented in soybean extracts. IgE from individuals allergic to soybeans, but not to peanut, was shown by immunoblot analysis to bind to proteins with a molecular weight of approximately 21 kD. These findings suggested that unique proteins in soybeans might be responsible for soybean allergic reactivity. The objective of the present study was to identify unique proteins in soybean extracts that bind to specific IgE from soybean-sensitive individuals, and to characterize the allergen using physicochemical methods and IgE binding. METHODS: Two-dimensional and preparative SDS-PAGE/IgE immunoblot analysis was used to identify a 22-kD soybean-specific allergen from crude soybean extracts. N-terminal sequence analysis was used to determine the identification of the protein binding IgE from soybean-sensitive individuals. RESULTS: IgE immunoblot and amino acid sequence analysis identified the 22-kD protein as a member of the G2 glycinin soybean protein family. Further investigation revealed that the IgEs reacted with basic chains from each member of the glycinin family of soybean storage proteins. CONCLUSIONS: Each of the subunits from glycinin, the storage protein that is the most prevalent component of soybean, are major allergens. PMID- 11112858 TI - Characterisation of horse dander allergen glycoproteins using amino acid and glycan structure analyses. a mass spectrometric method for glycan chain analysis of glycoproteins separated by two-dimensional electrophoresis. AB - Separation of horse dander allergens using two-dimensional PAGE resulted in the identification of 16 proteins that react with allergic patient sera. A sensitive method has been developed for analysing the structures of the glycan chains of individual glycoprotein allergens transferred to blots following two-dimensional PAGE, and has allowed the structural identification of the glycan chains of the most abundant isoforms of Equ c 1, a glycosylated horse dander major allergen. The method involves separation of the allergens by two-dimensional PAGE, transfer to polyvinylidene difluoride membranes, release of the glycan chains using peptide N-glycosidase F, permethylation and mass spectrometric analysis of the derivatised glycans. The amino acid compositions of the 16 horse dander allergens separated by two-dimensional PAGE have been determined, allowing the identification of the various isoforms of Equ c 1. These results also confirmed that the two non-glycosylated major allergens, Equ c 2.0101 and Equ c 2.0102, belong to the lipocalin family, and support the idea that these two allergens are most probably isoforms of the same protein. The glycan structures identified using the mass spectrometric method are common biantennary and triantennary glycan chains. These carbohydrate moieties may have a role in the binding of IgE; however, it is more likely that the overall glycoprotein structure involving both the glycan and protein moieties, rather than the structure of the glycan chains alone, is responsible for eliciting allergic responses. PMID- 11112857 TI - A soybean G2 glycinin allergen. 2. Epitope mapping and three-dimensional modeling. AB - BACKGROUND: Multiple allergens have been documented in soybean extracts. IgE from individuals allergic to soybeans, but not to peanut, has been shown by immunoblot analysis to bind to proteins with a molecular weight of approximately 22 kD. These findings suggested that this unique protein fraction from soybean might be responsible, in part, for soybean allergic reactivity. The objective of the present study was to characterize specific B cell epitopes, to determine if any amino acid was critical to IgE binding and to model the 22-kD G2 soybean allergen to the three-dimensional (3-D) phaseolin molecule. METHODS: B cell epitopes were identified using SPOTs peptide analysis. Structural orientation of the IgE binding regions was mapped to the 3-D phaseolin molecule using molecular modeling of the protein tertiary structure. RESULTS: Eleven linear epitopes, representing 15 amino acid peptide sequences, bound to IgE in the glycinin molecule. These epitopes were predicted to be distributed asymmetrically on the surface of G2 trimers. CONCLUSIONS: Only 1 epitope could be rendered non-IgE binding by alanine substitutions in the peptide. The nonrandom distribution of the IgE binding sites provides new insight into their organization in trimers in 11S complexes of the G2 glycinin allergen. PMID- 11112860 TI - Development of a monoclonal antibody-based sandwich ELISA for detection of the latex allergen Hev b 1. AB - BACKGROUND: Natural rubber latex (NRL) products are complex mixtures consisting of different allergenic components. Among them, Hev b 1 belongs to the important and well-characterized ones. To quantify the relevant allergen Hev b 1 in NRL products, a two-site monoclonal antibody (mAb)-based assay was developed. METHODS: Two Hev b 1-specific mAbs with different epitope recognition and ability to bind simultaneously to an Hev b 1 molecule were used in the study. Both mAbs (II4F9 and II4G9) were enriched by in vitro production in a modular minifermenter and affinity purified. Wells of micro-ELISA plates coated with captured mAb II4G9 were incubated with samples containing Hev b 1. Bound Hev b 1 was detected by a combination of biotinylated mAb II4F9 as detection antibody and peroxidase labeled avidin. RESULTS: The optimized sandwich ELISA was highly reproducible in the linear range of the standard curve and Hev b 1 concentrations ranging from 12.5 to 400 ng/100 microl could be detected. The assay was suitable for the detection of Hev b 1 concentrations in latex sap and latex products, e.g. gloves, with a detection limit of 1.25 microg of Hev b 1/g of rubber. In a preliminary study with five different brands of latex gloves, Hev b 1 concentrations were found to be in the range of 18-40 microg per gram of rubber material, corresponding to 2-4% of the total extractable protein content in latex glove extracts. CONCLUSIONS: A sensitive sandwich assay was developed to quantify the latex allergen Hev b 1. This assay can be used to standardize latex extracts with regard to the content of the major allergen Hev b 1. PMID- 11112859 TI - Identification of a sequential B-cell epitope on major allergen (Cry j 1) of Japanese cedar (Cryptomeria japonica) pollen in mice. AB - BACKGROUND: Japanese cedar (Cryptomeria japonica; CJ) pollinosis is one of the most common allergic diseases in Japan. B cell epitopes on Cry j 1, a major allergen of CJ pollen, have been analyzed by the specific monoclonal antibodies to Cry j 1, and most of these epitopes may be conformational, but no previous report has addressed the analysis of sequential epitope mapping with synthetic peptides. The main purpose of the present study is to identify IgE and IgG B cell epitopes on Cry j 1 by using a synthetic peptide approach in mice. METHODS: We synthesized 35 overlapping peptides that cover the entire length of Cry j 1 and examined whether mouse IgE and IgG antibodies produced by immunization with Cry j 1 reacted to the Cry j 1 peptides. RESULTS AND CONCLUSION: We found that mouse IgE and IgG antibodies reacted strongly to Cry j 1 peptide No. 15 ((141)GVEPVHPQDGDALTLRTATN(160)), though those antibodies did not react with other peptides. IgE and IgG antibody binding to peptide No. 15 was completely inhibited by Cry j 1 and the peptide. To determine the minimum epitope in peptide No. 15, we conducted an ELISA inhibition test. IgE and IgG antibody binding to peptide No. 15 was inhibited by smaller peptides of this peptide. We found the core of the epitope to be (145)VHPQDGDA(152). PMID- 11112861 TI - Monoclonal antibodies against Der s 1, a major allergen of Dermatophagoides siboney. AB - Six stable clones secreting murine monoclonal antibodies (Mabs) against Der s 1 were obtained. The binding of Mabs showed cross-reactivity with Dermatophagoides farinae, as determined by enzyme-linked immunosorbent assay (ELISA). In a Western blot assay, antibodies reacted with a 24-kD protein considered to represent the major allergen Der s 1. The repertoire of antigenic sites on Der s 1 was studied using a panel of Mabs. Epitope specificity of the Mabs was determined by both competitive inhibition and sandwich ELISA assays. The results defined six different, non-overlapping and non-repeated antigenic sites on the allergen molecule. Der s 1 allergen from Dermatophagoides siboney extracts was purified by Mab affinity chromatography, this procedure gave 43% recovery of >90% pure allergen. The purified allergen had capacity to bind specific human IgE and demonstrated an allergenic activity of up to 77% of total D. siboney extract. An Mab-ELISA was developed using Mabs directed against different epitopes on Der s 1. This assay could detect up to 1 ng/ml of Der s 1 and Der f 1 in allergen preparations. PMID- 11112862 TI - Immunologic stimulation of mast cells leads to the reversible exposure of phosphatidylserine in the absence of apoptosis. AB - BACKGROUND: Loss of phospholipid asymmetry represents one of the hallmarks of apoptosis and results in the surface exposure of phosphatidylserine (PS) which can be indirectly monitored by the calcium-dependent binding of annexin V. METHODS AND RESULTS: Here, we provide evidence that the IgE-dependent stimulation of a rat mast cell line, as well as murine and human nontransformed mast cells, leads to the exposure of PS at the plasma membrane. The appearance of PS was quantitatively related to allergic mediator release. Pharmacological agents that prevent stimulus-secretion coupling blocked PS cell surface exposure and calcium ionophore-induced PS appearance, suggesting that it is a direct consequence of exocytosis rather than early signaling events initiated by the aggregation of the high-affinity IgE receptor (FcepsilonRI). The surface exposure of PS in mast cells was reversible even in the continuous presence of stimulus and was not associated with the appearance of apoptotic nuclei, demonstrating that it was independent of physiological cell death. CONCLUSIONS: In addition to providing a means of monitoring exocytosis at the single cell level, our results indicate that PS externalization in mast cells is not necessarily related to apoptosis but could be an important feature of the degranulation process. PMID- 11112863 TI - MMP-2- and MMP-9-linked gelatinolytic activity in the sputum from patients with asthma and chronic obstructive pulmonary disease. AB - BACKGROUND: The course of asthma and chronic obstructive pulmonary disease (COPD) is associated with bronchial morphological changes. Metalloproteinases are thought to play a role in these structural changes. METHODS: We studied the gelatinolytic activity present in the induced sputum from 20 patients with asthma, 20 with COPD and 19 healthy controls. The assessment of gelatinolytic activity was performed by quantitative zymography, and gelatinolytic species were identified by Western blot analysis. Tissue inhibitor of metalloproteinase-1 (TIMP-1) was detected by reverse zymography and ELISA. RESULTS: From zymography, we found significantly higher gelatinolytic activity linked to pro-matrix metalloproteinase-9 (pro-MMP-9) in the sputum from asthmatics (p < 0.0001) and COPD patients (p < 0.0001) compared to the control group. Furthermore, the activated form of MMP-9 (85 kD) was found in the sputum from 60% of asthmatics and 85% of COPD patients, but was absent in that of control subjects (p < 0.0001). Importantly, although less frequently detectable than pro-MMP-9, pro- MMP-2 (72 kD) was found more frequently in asthmatics (50%) than in control subjects (5%) (p < 0. 005). We also described two unusual gelatinolytic species of 45 and 120 kD and showed that they derived from MMP-9 according to their ability to bind gelatin and anti-MMP-9 antibody. Levels of TIMP-1 were higher in asthmatics (p < 0.05) and COPD patients (p < 0.05) than in controls. CONCLUSION: Asthmatics and COPD patients display an increased gelatinolytic activity linked to MMP-2 and MMP-9 and higher levels of TIMP-1 in their sputum. PMID- 11112864 TI - Double sensitization to honeybee and wasp venom: immunotherapy with one or with both venoms? Value of FEIA inhibition for the identification of the cross reacting ige antibodies in double-sensitized patients to honeybee and wasp venom. AB - BACKGROUND: Double sensitization to honeybee (Apis mellifera) and wasp venom (Vespula spp.) as determined by skin test and measurement of specific IgE is common in hymenoptera sting allergy. Double-sensitized patients have either distinct antibodies for each venom or cross-reacting antibodies that recognize similar or identical epitopes in both venoms. Unfortunately, patients often fail to identify the stinging insect which makes it difficult to distinguish cross reactors from non cross-reactors. However, for economic reasons as well as for the benefit of the patients, it would be useful to identify complete cross reactors. METHODS: In this study we investigated 24 double-sensitized patients who were candidates for venom immunotherapy. Homologous and heterologous FEIA inhibition was carried out with honeybee (Apis mellifera) and wasp venom (Vespula spp.) preparations from two different providers. The inhibitor concentrations were ranging from 0 to 100 microg protein/ml. RESULTS: Sera of 4 patients were completely cross-reacting for one venom (3 honeybee, 1 wasp), 8 patients were partially cross-reacting and 10 patients were not cross-reacting. Two patients were excluded from the study due to insufficient homologous inhibition. Data from specific IgE measurements, skin test, and clinical history were not useful for the identification of cross-reacting patients. CONCLUSION: FEIA inhibition is easy to perform and useful for the identification of patients with complete cross reactivity. In these patients immunotherapy might be restricted to one venom which is beneficial for the patient and cost-effective. PMID- 11112865 TI - Qualitative determination of urinary human complement factor H-related protein (hcfHrp) in patients with bladder cancer, healthy controls, and patients with benign urologic disease. AB - OBJECTIVES: To assess the diagnostic quality of human complement factor H-related protein (hcfHrp, BTA STAT(TM)) as a qualitative urinary marker for bladder cancer. METHODS: Urine samples of 354 individuals (76 healthy volunteers, 111 patients with benign urologic disorders, 167 patients with histologically proven bladder cancer) were examined prior to therapy for the presence of hcfHrp. RESULTS: Overall test sensitivity was 62.9%. Sensitivity of low-grade/low-stage tumors was <50% and was thus comparable to published sensitivities of urinary cytology. In high-stage, high-grade tumors sensitivity was 100 and 77.3%, respectively. Superficial tumors with high risk of progression (pT1G3) were detected significantly better (88.9% sensitivity) than low-risk superficial tumors (pTa G1-3/pT1G1-2; 48.2%; p < 0.001). The overall specificity (healthy individuals and patients with benign urologic disease) was 93.0%. CONCLUSION: Since test sensitivity is comparable to urinary cytology and specificity is excellent, hcfHrp should be further evaluated in prospective studies focussing on the follow-up of patients with bladder cancer. PMID- 11112866 TI - Long-term analysis of suprapubic cystostomy drainage in patients with neurogenic bladder. AB - OBJECTIVE: We assessed the roles of suprapubic cystostomy in patients with neurogenic bladder and analyzed the complications and their courses. PATIENTS AND METHODS: We reviewed 118 patients with neurogenic bladder managed with suprapubic cystostomy. The original diseases were spinal cord injury in 90, degenerative disease of the central nervous system in 15, spina bifida in 6, cerebral palsy in 3, pontine bleeding in 1, Parkinson's disease in 1, brain tumor in 1, and dysgenesis of the external sphincter in 1. Fifty-six (62.2%) of spinal cord injured patients demonstrated cervical damage. Renal function, urinary pH and white blood cell values were measured and evaluated after insertion. The stone free rate after insertion was estimated by the Kaplan-Meier method. RESULTS: Indications for cystostomy were failure of clean intermittent catheterization in 62 (52.5%) and Crede's maneuver in 2, severe urethral damage in 30 (25.4%), replacement of urethral catheter in 3, worsening of the original disease in 15 (12.7%), deterioration of the general condition in 2, mental retardation in 2, and traumatic vesical rupture in 1. Frequent complications were formation of the bladder calculi in 30 (25%) and urinary leakage through the urethra in 11 (10%). No fatal complications occurred. The stone-free rates 5 and 10 years after insertion were 77 and 64%, respectively. The urinary pH of the stone-forming group was significantly higher than that of the stone-free group. The high urinary pH group (>7.24) had a higher risk of stone formation. CONCLUSIONS: Although continuous cystostomy drainage is not considered to be ideal management for bladder emptying, some patients with neurogenic bladder may benefit from this procedure. PMID- 11112867 TI - Acupuncture on clinical symptoms and urodynamic measurements in spinal-cord injured patients with detrusor hyperreflexia. AB - OBJECTIVES: We investigated the possible use of acupuncture for the treatment of urinary incontinence caused by detrusor hyperreflexia in patients with chronic spinal cord injuries. METHODS: A total of 13 patients (11 males, 2 females) suffering from urinary incontinence due to spinal cord injuries were treated by acupuncture, which was carried out with disposable stainless steel needles inserted into the bilateral BL-33 (Zhongliao) points on the skin of the third posterior sacral foramina. Urodynamic studies were also performed before acupuncture, immediately after the 1st acupuncture and 1 week after the 4th acupuncture. In 6 patients, these urodynamic studies were performed again 1 month after the 4th acupuncture. RESULTS: No side effects were recognized throughout the treatment period. Of the 13 patients, incontinence disappeared in 2 (15%) and decreased to 50% or less compared to baseline in a further 6 (46%). Maximum cystometric bladder capacity increased significantly from 76.2 +/- 62.3 to 148.1 +/- 81.5 ml 1 week after the 4th acupuncture (p < 0.01). In the 6 patients in whom cystometry was repeated 1 month after the 4th acupuncture, bladder capacity decreased from 187.5 +/- 90.4 ml 1 week after the 4th acupuncture to 128.3 +/- 93.4 ml. CONCLUSION: In spinal cord injury patients acupuncture could represent another valuable therapeutic alternative to the treatment of urinary incontinence caused by detrusor hyperreflexia. PMID- 11112868 TI - Evaluation of a fibrin sealant free of bovine-derived components in an experimental vas anastomosis study. AB - OBJECTIVES: The risk of transmission of bovine spongiform encephalopathy cannot be excluded from the use of bovine-derived products. The present study was undertaken to evaluate the performance of a new fibrin glue free of bovine derived components in vas anastomosis and to compare this product to conventional vas anastomosis with fibrin glue. METHODS: Bilateral delayed vas anastomosis was performed in 40 Sprague-Dawley rats. All animals underwent a fibrin glue-assisted vas anastomosis with three transmural sutures tied prior to fibrin glue application. The composition and preparation of fibrin glue was similar for all vas anastomoses except the fibrinolysis inhibitor component which was aprotinin (3,000 KUI/ml) in group 1 and tranexamic acid (10 mg/ml) in group 2. The animals (20 rats in both groups) were sacrificed 7 weeks postoperatively and evaluated for gross patency, presence of sperm granuloma and tensile strength measurements at the anastomosis site. RESULTS: No difference was found between the 2 groups for all parameters evaluated whether a bovine-derived or a synthetic fibrinolysis inhibitor component was used. CONCLUSION: This study showed that tranexamic acid, a fibrinolysis inhibitor, can be substituted for conventional fibrin glue thereby avoiding the risks of bovine products. PMID- 11112869 TI - Relationship between antisperm antibodies, sperm movement, and semen quality. AB - The presence of antisperm antibodies might impair fertilization by affecting sperm function in different ways. The aim of this study was to analyze the incidence of antisperm antibodies and evaluate its impact on sperm movement, in normozoospermic and abnormal semen samples. Samples from 144 patients were classified as normozoospermic or abnormal, according to WHO guidelines and Kruger's strict criteria. Motilility was assessed by a computer-assisted system by placing a drop of semen in a Makler chamber. Antisperm antibodies were detected using the Mar Test and Immunobead tests to identify immunoglobulin location and isotype. Results showed no association between the presence of antisperm antibodies and semen quality. Antibody incidence was 15 and 9.5% for normozoospermic and abnormal semen samples, respectively. Motion analysis showed that, although sperm linearity was significantly reduced in abnormal patients with antibodies, the existence of an immune response did not affect the sperm pathway in most cases. This study suggests that sperm motion analysis would not be of great assistance in the screening of an immunological male fertility factor. PMID- 11112870 TI - Risk factors for an early increase in dose of vasoactive agents for intracavernous pharmacotherapy. AB - AIMS: This study was to identify factors that influence the early increase in the dose of intracavernous vasoactive agents to maintain erection for satisfactory intercourse and to elucidate when the dose increase would begin. METHODS: Seventy nine patients using intracavernous pharmacotherapy with tri-mix (PGE(1), papaverine, and phentolamine) over a 3- to 4-year period were enrolled. At 3 month intervals, patients were questioned about changes in dose to maintain erection for satisfactory intercourse and frequency of injection, and underwent examination of the penis. The patients were divided into 2 groups: group A, dose increase of > or = 20% after initiating home therapy, and group B, no change or an increase in dose < 20%. RESULTS: A significant increase (p < 0.01) in dose was started 2-3 years after initiating pharmacotherapy. The initial doses of group A at 2-3 and 3-4 years were significantly higher than those of group B (p < 0.01). There were no significant differences in the age of the patients, duration of erectile dysfunction, incidence of accompanying vascular risk factors, frequency of injection, or incidence of fibrous plaques between group A and group B, both at 2-3 and 3-4 years. CONCLUSION: The initial dose of intracavernous vasoactive agents (tri-mix) may be a unique risk factor for the early increase in dose to maintain erection for satisfactory intercourse. PMID- 11112871 TI - The shrinking seminoma--fact or fiction? AB - We report 2 cases of testicular seminoma with unusual but dissimilar clinical presentations, which may represent different stages of spontaneous regression of a primary testicular seminoma, the so-called 'shrinking seminoma'. The aetiology of this phenomenon is discussed. These cases also illustrate that, like enlargement, reduction in the size of a testicle warrants investigation for malignancy. PMID- 11112872 TI - Sertoli cell tumor in a prepubertal boy mimicking testicular torsion. AB - A 9-year-old boy presented with left, intermittent testicular pain that was present for 3 days. On physical examination, left testis was grossly enlarged and firm but mildly tender. Serum alpha-fetoprotein and beta-human chorionic gonadotropin levels were within normal range. Color doppler ultrasonography which was performed to rule out testicular torsion revealed an intratesticular mass located at the upper pole of left testis and left radical orchiectomy was performed. The histopathological diagnosis was Sertoli cell tumor. PMID- 11112873 TI - Sacral hydatid cysts: an uncommon cause of neurogenic bladder. AB - Hydatid cysts of the sacrum are rare entities, characterized by chronicity without any clinical manifestation and are usually misdiagnosed in the early stage resulting in significant loss of bone and destruction of surrounding tissue. One should keep this possibility in mind in cases of early sphincteric involvement with minimal sensorimotor deficit in the lower limbs and bone destruction on radiography. PMID- 11112874 TI - Infected hydrocele in a neonate. AB - A case of infected hydrocele in a neonate is presented. We describe this unusual condition, and discuss the diagnosis, pathophysiology and treatment. PMID- 11112875 TI - Idiopathic congenital dysmorphic megascrotum. AB - An enlarged scrotum in the pediatric population constitutes a relatively frequent physical finding requiring evaluation. Most cases of scrotomegaly have a clearly identifiable etiology. We present a patient with an idiopathic congenital dysmorphic megascrotum. PMID- 11112876 TI - Telomerase activity in giant condyloma acuminatum. AB - A 46-year-old male came to our hospital 1 month after noticing a 2-cm penile tumor. Since malignant findings such as atypical cells and mitosis were not observed in the frozen sections obtained at operation, the pathological diagnosis of this tumor was giant condyloma acuminatum. This tumor was analyzed by a telomeric repeat amplification protocol method, and telomerase activity was revealed. For comparison, a case of squamous cell carcinoma and a case of condyloma acuminatum were examined. Telomerase activity was observed in our case and in the case of squamous cell carcinoma. To our knowledge, this is the first case of telomerase activity in giant condyloma acuminatum ever reported. In addition to the histological examination, measurement of telomerase activity may provide valuable objective diagnostic information on evaluating the degree of malignancy of giant condyloma acuminatum and in obtaining a differential diagnosis between the benign and malignant. PMID- 11112877 TI - Extravesical foreign body presenting as a bladder tumor. AB - Although foreign bodies left in the abdominal cavity may remain asymptomatic for long periods, they may also cause serious complications. We present a case of gauze forgotten in the lower abdomen which remained asymptomatic for almost 1 year. When a granuloma had formed, it infiltrated the bladder wall giving the clinical and imaging appearance of an invasive bladder tumor. PMID- 11112878 TI - An accessory spleen mimicking a nonfunctional adrenal tumor: a potential pitfall in the diagnosis of a left adrenal tumor. AB - We describe a case of accessory spleen mimicking a left adrenal tumor. A 66-year old woman was referred to our hospital because of a suspected left adrenal mass detected by US. A laparoscopic adrenalectomy was performed, and examination of the surgical specimen revealed that the resected adrenal gland contained no tumorous lesion. A further investigation of the intraperitoneal space revealed an accessory spleen. This indicates that urologists should be aware of the possible existence of accessory spleens when left adrenal tumors are suspected on CT and MRI. PMID- 11112879 TI - Cystic leiomyosarcoma of the kidney: an unusual clinical presentation. AB - Primary sarcomas of the kidney are rare, accounting for 1-3% of all renal malignancies. We describe an unusual case of renal leiomyosarcoma in a 41-year old white woman who presented with a large smooth mass, which was mobile to the overlying structures and which occupied the right hypochondria and flank. Radical nephrectomy was carried out and the patient is well, without symptoms of relapse, 1 year after surgery. Leiomyosarcomas of the kidney have an aggressive and rapidly progressive natural history, with 5-year survival rates of 29-36%. Size <5 cm, low histological grade, absence of lymph node metastases and radical surgical treatment are all associated with a better prognosis. Irradiation and chemotherapy do not appear to alter the clinical course. PMID- 11112881 TI - Effect of allergist intervention on patient-centered and societal outcomes: allergists as leaders, innovators, and educators. AB - Atopic disorders, which afflict millions of Americans and hundreds of millions worldwide, are at epidemic levels with concomitant increases in morbidity and mortality. Environmental and lifestyle changes over the past three to five decades are proposed causes for this pandemic and as such present major burdens to reverse. The scope of allergy practice bridges directly on this challenge. Allergy as a specialty is a major leader in developing effective strategies to confront this epidemic. Allergists have made major contributions to the understanding of the risk factors, immunology, pathophysiology, immunomodulation, and prevention of atopic and immunologic disorders. Allergist epidemiologists and clinicians have helped develop and implement national and international guidelines in the recognition, management, and prevention of asthma and rhinitis. Allergist clinical researchers are active in (1) outcomes research that demonstrates convincingly the value of allergy as a specialty in asthma, allergic rhinitis, anaphylaxis, drug and food allergy, and other atopic disorders, (2) National Institutes of Health clinical trials that will form the basis for the future treatment of asthma and allergic disease, and (3) pharmaceutical trials that evaluate new, effective, and safe medication to treat atopic disease. Allergist educators, comprising academic and practicing allergists, supported by allied health professionals, national associations, and affiliated lay organizations, provide comprehensive education to fellows, residents, colleague physicians, media, the public, and patients. Documentation of the value of allergists in improving patient-centered and societal outcomes in their core domain, allergy, is the appropriate final topic contribution in the important series "New millennium: The conquest of allergy." PMID- 11112882 TI - Allergic and immunologic disorders of the eye. Part II: ocular allergy. AB - Allergy affects more than 15% of the world population, and some studies have shown that up 30% of the US population has some form of allergy. Most of these patients have various target organs for their allergies, and most have ocular involvement. The ocular component may be the most prominent and sometimes disabling feature of their allergy. Some are affected for only a few weeks to months, whereas others have symptoms that last throughout the year. The seasonal forms may present to clinical allergists, whereas the more chronic forms may present to ophthalmologists. Thus, in the second of this 2-part review series (Part I: Ocular Immunology appeared in the November issue of the Journal), an overview is provided of the spectrum of ocular allergy that ranges from acute seasonal allergic conjunctivitis to chronic variants of atopic keratoconjunctivitis. With a better understanding of the immunologic mechanisms, we now can develop better treatment approaches and design further research in intervention of allergic eye diseases. PMID- 11112883 TI - Pathophysiology of severe asthma. AB - Although asthma affects nearly 8% of the adult population, most of these patients have mild-to-moderate disease that can be controlled with appropriate treatment. It is estimated, however, that 5% to 10% of patients with asthma have severe disease that is unresponsive to typical therapeutics, including corticosteroids. Because patients with severe asthma are disproportionately affected by their disease, in terms of both impaired lifestyle and health care costs, the National Heart, Lung, and Blood Institute sponsored a workshop on the pathogenesis of severe asthma. The goals of this workshop were to begin to define the characteristics of severe asthma. In these discussions, it was clear that many characteristics need to be considered in defining this phenotype of asthma, including symptoms, intensity of therapy (including administration of systemic corticosteroids), and impairment of lung function. Also discussed were potential mechanisms of severe asthma including the role of allergic diseases, which may play less of a role in severe asthma than in mild-to-moderate disease, and infections. A major limitation to control of severe asthma is the recalcitrant response of these patients to usual therapy including systemic corticosteroids; the potential of other therapies was reviewed. From these discussions, recommendations were made for future research needs to gain insights into a difficult therapeutic and possibly novel mechanistic area of asthma. PMID- 11112884 TI - New opportunities, new products. PMID- 11112885 TI - Natural and induced allergic responses increase the ability of the nose to warm and humidify air. AB - BACKGROUND: We have previously shown that subjects with seasonal allergic rhinitis out of season had a reduced ability to warm and humidify air compared with normal subjects. OBJECTIVE: We sought to investigate whether allergic reactions induced by either seasonal exposure or nasal challenge with antigen would decrease the capacity of the nose to condition cold, dry air. METHODS: We performed two prospective studies comparing the effects of allergic inflammation, induced by either seasonal exposure or nasal challenge with antigen, on nasal conditioning capacity (NCC). The total water gradient (WG) across the nose was used to represent the NCC. In the first study, the NCC was measured and compared before and during the allergy season in 10 subjects with seasonal allergic rhinitis. In the second study, 20 subjects with seasonal allergic rhinitis were recruited outside of the allergy season. NCC was measured and compared before and 24 hours after challenge with antigen. RESULTS: In the first study, seasonal allergic subjects in season showed a significant increase in NCC when compared with their preseason baseline (total WG in season: 2050 +/- 138 mg vs total WG preseason: 1524 +/- 100 mg; P <.01). In the second study, antigen challenge led to early-phase and late-phase responses. There was a statistically significant increase in NCC 24 hours after antigen challenge compared with that before antigen challenge (total WG after antigen challenge: 1938 +/- 101 mg vs total WG before antigen challenge: 1648 +/- 84 mg; P =.01). CONCLUSION: Allergic reactions induced by either seasonal exposure or antigen challenge increase the ability of the nose to condition inspired air. We speculate that allergic inflammation increases this ability by changing the perimeter of the nasal cavity. PMID- 11112886 TI - Effects of topical glucocorticoids on in vitro lactoferrin glandular secretion: comparison between human upper and lower airways. AB - BACKGROUND: Mucus hypersecretion is a hallmark of upper and lower airway diseases, such as rhinitis, asthma, and chronic obstructive pulmonary disease. Although topical glucocorticoids are widely used to treat mucosal inflammation, their effect on mucus hypersecretion remains uncertain. OBJECTIVE: The aim of this study was to investigate the effect of budesonide and beclomethasone dipropionate on in vitro lactoferrin glandular secretion from both human nasal and bronchial mucosa and the potential mediating role of lipocortin 1. METHODS: Nasal and bronchial explants obtained from patients undergoing surgery were cultured in a controlled atmosphere. Lactoferrin (ELISA) was measured in culture supernatants, and lipocortin 1 (Western blot) was analyzed in explant tissues. RESULTS: Both budesonide and beclomethasone dipropionate (10(-6) mol/L) decreased spontaneous lactoferrin secretion in nasal and bronchial mucosa. The maximum effect of cortico-steroids (10(-6) mol/L) was obtained at day 3 in bronchial mucosa (budesonide: -56% +/- 9%, P <.05; beclomethasone dipropionate: -32% +/- 6%, P <.05) and at day 5 in nasal mucosa (budesonide: -34% +/- 10%, P <.05; beclomethasone dipropionate: -37% +/- 10%, P <.05). Methacholine (10(-4) mol/L) increased lactoferrin secretion in both bronchial (248% +/- 72%, P <.05) and nasal (107% +/- 28%, P <.05) explants, with this effect being completely abrogated by atropine. Budesonide caused a dose-related inhibitory effect on methacholine-induced lactoferrin secretion that was similar in both bronchial (down to -86% at 10(-6) mol/L) and nasal (down to -73% at 10(-6) mol/L) mucosa. Budesonide (10(-6) mol/L) did not show any effect on lipocortin 1 expression. CONCLUSIONS: These results suggest that glucocorticoid effects on airway inflammation may include a reduction of mucus hypersecretion in both nasal and bronchial mucosa. PMID- 11112887 TI - The relationship of sputum eosinophilia and sputum cell generation of IL-5. AB - BACKGROUND: Eosinophil recruitment to the airway after antigen challenge is regulated by many factors, including airway cell generation of cytokines. OBJECTIVES: The purpose of this study was to determine the relationship between sputum cell generation of IL-5 and the appearance of eosinophils in the sputum after antigen challenge. METHODS: Sputum samples from 11 allergic subjects were collected before and again 4 and 24 hours after antigen challenge. In 6 of these subjects, induced sputum samples were also obtained 48 hours and 7 days after challenge. Sputum leukocyte differential and cell counts and eosinophil-derived neurotoxin levels were determined. Sputum cells were then cultured with PHA (10 microg/mL) to stimulate IL-5 and IFN-gamma, which were measured in culture supernatants. RESULTS: An increase in sputum eosinophils and eosinophil-derived neurotoxin levels was detected at 4 hours after antigen challenge, with peak values at 24 hours. In contrast, significant increases in ex vivo generation of IL-5 by sputum cells was not seen until 24 hours after challenge. At 24 hours, PHA-induced IL-5 correlated with airspace eosinophil values (r (s) = 0.78, P <.01). In addition, the ratio of IFN-gamma/IL-5 decreased at 24 hours (P <.05) and had an inverse correlation with sputum eosinophils (r (s)= -0.68, P <.05). CONCLUSION: Although eosinophils are increased in the airway lumen as early as 4 hours, the ex vivo generation of IL-5 by sputum cells is first noted in samples obtained 24 hours after antigen challenge. This suggests that the early (4 hours) recruitment of eosinophils to the airway lumen may be regulated by factors other than IL-5 or that mucosal cells (rather than airspace cells) contribute to the IL 5 generation at this time point. Furthermore, IL-5 generation by airspace cells may be more responsible for either eosinophil recruitment or retention at later time points. PMID- 11112888 TI - Mouse allergen. I. The prevalence of mouse allergen in inner-city homes. The National Cooperative Inner-City Asthma Study. AB - BACKGROUND: Although mouse allergen is a well-defined cause of IgE-mediated hypersensitivity in occupational settings, it has not been well studied in the general population. OBJECTIVE: We sought to determine the prevalence of mouse allergen in inner-city homes. METHODS: A subset of 608 homes from the National Cooperative Inner-City Asthma Study population had dust samples adequate for analysis of mouse allergen. In addition, data regarding the demographics and housing of the subjects were related to the mouse allergen levels. RESULTS: Ninety-five percent of all homes had detectable mouse allergen (Mus m 1) in at least one room, with the highest levels found in kitchens (kitchen: range, 0-618 microg/g; median, 1.60 microg/g; bedroom: range, 0-294 microg/g; median, 0.52 microg/g; television-living room: range, 0-203 microg/g; median, 0. 57 microg/g). By city, 100% of the kitchens in Baltimore had detectable mouse allergen, with the lowest percentage (74%) in Cleveland. Mouse allergen levels correlated among rooms (R = 0.65-0. 75). Forty-nine percent of the homes had reported problems with mice within the last year, and 29% of the homes had evidence of mice in one or more rooms on home inspection and had higher levels of mouse allergen (P =.0001). Higher allergen levels were also associated with evidence of cockroach infestation in any room (P =.006). None of the other subject or housing demographics evaluated were associated with a higher prevalence or level of mouse allergen. CONCLUSIONS: We conclude that mouse allergen is widely distributed in inner-city homes and that cockroach infestation is associated with high mouse allergen levels. PMID- 11112889 TI - Mouse allergen. II. The relationship of mouse allergen exposure to mouse sensitization and asthma morbidity in inner-city children with asthma. AB - BACKGROUND: Although mouse allergen is known to cause occupational asthma in laboratory workers, its potential significance in home environments has never been studied. OBJECTIVE: This study was designed to define the prevalence of mouse sensitivity and its relationship to mouse allergen exposure and disease activity in inner-city children with asthma. METHODS: A subset of 499 subjects from the National Cooperative Inner-City Asthma Study had dust samples adequate for mouse allergen analysis, as well as valid puncture skin test (PST) results. Data were analyzed to relate mouse allergen exposure and other risk factors to mouse sensitization and asthma morbidity. RESULTS: Eighty-nine (18%) of the 499 children had a positive mouse skin test response. Children whose homes had mouse allergen levels above the median (1.60 microg/g) in the kitchen had a significantly higher rate of mouse sensitization (23% vs 11%, P =. 007). Atopy was also significantly related to mouse sensitization, with 40% of those with more than 4 positive PST responses having mouse sensitivity compared with 4% of those with no other positive PST responses (P <.0001). When atopy and exposure were considered together, 53% of those with more than 4 positive PST responses and allergen levels above the median had a positive PST response to mouse allergen compared with 22% of those with more than 4 positive PST responses and allergen levels below the median (P <.0001). The relationship among mouse allergen exposure, sensitization, and any measures of asthma morbidity was not statistically significant. CONCLUSIONS: Mouse allergen may be an important indoor allergen in inner-city children with asthma, with exposure and atopy contributing to mouse sensitization. PMID- 11112890 TI - Role of endogenous nitric oxide in exercise-induced airway narrowing in patients with bronchial asthma. AB - BACKGROUND: Increased amounts of nitric oxide (NO) in expired air and induced sputum have been found in asthmatic patients, and the role of excessively produced NO in the pathogenesis of bronchial asthma is under active investigation. OBJECTIVE: This study was designed to investigate the involvement of endogenous NO in exercise-induced bronchoconstriction (EIB) in asthmatic patients by using the sputum induction method. METHODS: The concentration of NO derivatives and inflammatory indices in induced sputum were examined in 18 asthmatic subjects and 10 normal control subjects. All asthmatic subjects performed an exercise test for 6 minutes. For 8 weeks after the first exercise testing, 400 microg of beclomethasone dipropionate twice daily was administered for asthmatic subjects with EIB, and the exercise testing and sputum induction were repeated in these patients. RESULTS: The concentration of NO derivatives in induced sputum was significantly higher in 9 asthmatic subjects with EIB (1580 +/ 280 micromol/L) than in 9 asthmatic subjects without EIB (1130 +/- 210 micromol/L) and normal control subjects (510 +/- 150 micromol/L). Moreover, there was a significant correlation between the concentration of NO derivatives and the percentage of maximal fall in FEV(1) (r = 0.569, P =.019). The concentration of NO derivatives was also more closely correlated with the area under the curve of the percentage fall in FEV(1) plotted against time for 30 minutes (AUC(0-30); r = 0.812, P <.001). After treatment with inhaled beclomethasone dipropionate in asthmatic subjects with EIB, there was a significant decrease in the concentration of NO derivatives in induced sputum. The change in the concentration of NO derivatives was significantly correlated with the change in the AUC(0-30) (r = 0.896, P =.0114) but not with the change in the percentage of maximal fall in FEV(1). CONCLUSION: These findings suggest that excessive production of NO is associated with EIB in patients with asthma and contributes to the prolonged airway narrowing phase rather than to the maximal airway narrowing evoked by exercise. PMID- 11112891 TI - Fluticasone propionate/salmeterol combination provides more effective asthma control than low-dose inhaled corticosteroid plus montelukast. AB - BACKGROUND: Asthma is a disease of chronic inflammation and bronchoconstriction. Inhaled corticosteroids (ICSs) provide important anti-inflammatory treatment but may not provide optimal control of asthma when taken alone. Two therapeutic alternatives for enhanced asthma control are to substitute the combination of fluticasone propionate (FP) and salmeterol (FP/Salm Combo) through the Diskus inhaler or to add montelukast to existing ICS therapy. OBJECTIVE: We compared the efficacy and safety of FP/Salm Combo through the Diskus inhaler versus montelukast added to FP (FP + montelukast) in patients whose symptoms were suboptimally controlled with ICS therapy. METHODS: We performed a multicenter, double-blind, double-dummy, parallel-group, 12-week study in 447 patients with asthma who were symptomatic at baseline while receiving low-dose FP. Patients were treated for 12 weeks with one of the following: (1) combination of FP 100 microg plus salmeterol 50 microg twice daily through the Diskus inhaler, or (2) FP 100 microg twice daily through the Diskus inhaler plus oral montelukast 10 mg once daily. RESULTS: FP/Salm Combo treatment provided better overall asthma control than FP + montelukast with significantly greater improvements in morning peak expiratory flow (+24.9 L/min vs +13.0 L/min, P <.001), evening peak expiratory flow (+18.9 L/min vs +9.6 L/min, P <.001), and forced expiratory volume in 1 second (+0.34 L vs +0.20 L, P <.001), as well as a change in the percentage of days with no albuterol use (+26.3% vs +19.1%, P =.032) and the shortness of breath symptom score (-0.56 vs -0.40, P =.017). The groups had comparable improvements in chest tightness, wheeze, and overall symptom scores. Asthma exacerbation rates were significantly lower (P =.031) in the FP/Salm Combo group (4 patients, 2%) than in the FP + montelukast group (13 patients, 6%). Adverse event profiles were comparable. CONCLUSION: Symptomatic patients on low dose ICS therapy had significantly greater improvement in asthma control when switched to the FP/Salm Combo than when montelukast was added to ICS therapy. PMID- 11112892 TI - Allergic fungal sinusitis: an immunohistologic analysis. AB - BACKGROUND: Allergic fungal sinusitis is a noninvasive form of fungal sinusitis that has recently been delineated as a distinct clinicopathologic entity. It is increasingly recognized as a cause of chronic sinusitis, with the primary causative agents being members of the Dematiaceae fungus family. Although its immunopathogenesis has not been elucidated, the eosinophil is a prominent inflammatory cell on histologic examination. OBJECTIVE: We sought to characterize the involvement of eosinophils in sinus tissue and accompanying mucin from patients with allergic fungal sinusitis. As a comparison, neutrophil and mast cell involvement was also evaluated in the same group of patients. METHODS: Tissue specimens from 8 patients with allergic fungal sinusitis, along with 8 nasal polyp specimens from patients without allergic fungal sinusitis, were stained by using indirect immunofluorescence for eosinophil granule major basic protein (MBP). Neutrophil elastase and mast cell tryptase staining was also performed on the same allergic fungal sinusitis and nasal polyp tissues. RESULTS: MBP was diffusely localized within the mucin, showing intense staining at the periphery and variable staining of degenerated cell clusters throughout. Extracellular MBP in the mucin was strikingly greater than intact eosinophil staining. Diffuse extracellular neutrophil elastase was also present in the mucin. Mucinous areas showed no tryptase localization. Adjacent nonmucinous areas of respiratory mucosa showed predominantly cellular staining with eosinophil MBP, neutrophil elastase, and mast cell tryptase. MBP staining of nasal polyps showed a predominantly cellular pattern with focal areas of extracellular deposition. CONCLUSIONS: Given the known toxicities of eosinophil granule MBP and neutrophil elastase, their extracellular presence supports the contribution of these proteins in the pathogenesis of allergic fungal sinusitis and further indicates that eosinophil and neutrophil activation occurs in the disease. PMID- 11112893 TI - Perception of induced bronchoconstriction in a community sample of adolescents. AB - BACKGROUND: Poor perception of asthma symptoms has been cited as a risk factor for asthma death, yet there is no consensus as to the best way to characterize perception, and little is known about perception in normative samples. Hypoperceivers are of clinical interest because of risks of undertreatment; hyperperceivers are at risk for adverse iatrogenic effects caused by overtreatment. OBJECTIVE: This study investigates perception of methacholine induced bronchoconstriction in 175 adolescents. METHODS: Breathlessness was rated after each inhalation by using the Borg scale. Perception groups were calculated on the basis of change from placebo Borg to high Borg scores (perception score at the highest methacholine dose). Subjects were called hypoperceivers if their Borg change score was greater than 1 SD below the mean for their FEV(1) group, hyperperceivers if their Borg change score was greater than 1 SD above the mean for their FEV(1) group, and accurate perceivers otherwise. RESULTS: For subjects with an FEV(1) drop of less than 10%, accurate perceivers had a change in Borg score of 1.4 or less, and hyperperceivers had a change of greater than 1.4. For a drop in FEV(1) between 10% and 19%, hypoperceivers had a change in Borg score of less than 0.2, accurate perceivers had a change between 0.2 and 2.1, and hyperperceivers had a change of greater than 2.1. For those with an FEV(1) drop of 20% or greater, hypoperceivers had a Borg change of less than 0.2, accurate perceivers had a change between 0.2 and 2.6, and hyperperceivers had a change of greater than 2.6. No differences in age, sex, placebo Borg ratings, baseline pulmonary functions, PC(20) values, or psychologic variables were found among perception groups. CONCLUSION: This study provides reference Borg values during methacholine challenge for 175 community adolescents. PMID- 11112894 TI - Asthma pharmacotherapy and utilization by children in 3 managed care organizations. The Pediatric Asthma Care Patient Outcomes Research Team. AB - BACKGROUND: Asthma is the most common chronic disease among children and the most frequent cause of hospitalization. Appropriate pharmacotherapy is a cornerstone of published national guidelines for the care of children with asthma. OBJECTIVE: The goal was to compare the baseline pharmacotherapy and health care utilization from 1996 to 1997 in children with asthma at managed care organizations (MCOs). METHODS: A common protocol was used to extract the study sample from 3 MCOs with automated claims and pharmacy databases. Children were selected if they were 3 to 15 years old as of June 1997 with 1 or more encounters (outpatient, emergency department visit, hospitalization) with an asthma diagnosis in the previous year. RESULTS: Of the 13,352 children studied, less than 40% were given controllers during the 12-month interval, with ranges of 15% to 77% by level of bronchodilator use, 31% to 44% by age, and 38% to 42% by MCO. Among children given 6 or more bronchodilators, controller dispensing ranged from 73% to 89% among the 3 MCOs. Variability was most evident for inhaled corticosteroids, for which dispensing ranged from 51% to 70%. Rates of asthma hospitalization and emergency department visits also differed among the MCOs, ranging from 21 to 37 per 1000 person-years and 37 to 142 per 1000 person-years, respectively. CONCLUSION: Five years after dissemination of national guidelines for care, the pattern of asthma therapy does not reflect guideline recommendations. Variation among health care organizations with respect to asthma therapy and utilization of health services exists. In addition, controller medications may not be used by all children who could benefit from them. PMID- 11112895 TI - Peripheral blood and airway tissue expression of transforming growth factor beta by neutrophils in asthmatic subjects and normal control subjects. AB - BACKGROUND: Airway remodeling may play an important role in asthma pathophysiology. Transforming growth factor beta (TGF-beta) has a critical role in the remodeling process. Although cellular sources for TGF-beta have been previously investigated in asthma airways, the expression, release, or both of TGF-beta from asthmatic airways and blood neutrophils has not been reported. OBJECTIVE: The current study evaluated the TGF-beta protein and messenger (m)RNA expression by airway and peripheral blood neutrophils in asthmatic and normal subjects. METHODS: TGF-beta protein expression by airway and peripheral blood neutrophils was detected by using immunocytochemistry. TGF-beta protein levels in blood neutrophil supernatant were measured by using an enzyme immunoassay. TGF beta mRNA expression was evaluated by using reverse transcription-PCR. RESULTS: Higher numbers of TGF-beta(+) cells and neutrophils were found in airway tissue of asthmatic (n = 15) compared with normal subjects (n = 10). Although neutrophils in both asthmatic and normal airway tissue expressed TGF-beta protein and the percentage of neutrophils expressing TGF-beta was similar between the two groups, the total number of TGF-beta(+) neutrophils was higher in the asthmatic subjects (P =.01). Peripheral blood neutrophils from asthmatic (n = 5) and normal subjects (n = 7) also expressed TGF-beta protein and mRNA. Blood neutrophils from asthmatic subjects spontaneously released significantly higher levels of TGF-beta than those from normal subjects (P =.007). CONCLUSION: These data suggest that airway and blood neutrophils from both asthmatic and normal subjects can express and release TGF-beta. Higher levels of TGF-beta expression-release from asthmatic neutrophils indicate that neutrophils may be involved in the airway remodeling process of asthmatic subjects. PMID- 11112896 TI - Expression of c-erbB receptors and ligands in human nasal epithelium. AB - BACKGROUND: The epidermal growth factor (EGF) family of growth factors plays an important role in maintenance and repair in a variety of epithelial tissues. However, very little is known about coexpression of these factors and their receptors, the c-erbB family of receptor tyrosine kinases, in human nasal epithelium. OBJECTIVE: We sought to investigate the expression of these molecules in cultured nasal epithelial cells and nasal mucosa from healthy individuals. METHODS: Identification of c-erbB receptors and their ligands was sought by using reverse transcription PCR, Western blotting, and immunohistochemistry. RESULTS: Messenger RNA encoding the EGF receptors (EGFR) c-erbB2 and c-erbB3, but not c erbB4, was detected in primary cultures of human nasal epithelial cells. Transcripts encoding EGF, heparin-binding EGF, transforming growth factor (TGF) alpha, and amphiregulin were also detected. Receptor and ligand expression was confirmed by using immunocytochemical staining of the cells and Western blotting of the cell lysates. Immunohistochemical analysis of tissue sections obtained from biopsy specimens of nasal mucosa revealed intense membrane staining for the EGFR within the respiratory nasal epithelium, which was predominantly localized at the level of the columnar epithelial layers. Similar staining patterns were observed for c-erbB2 and c-erbB3 in the respiratory nasal epithelium. Evidence for EGF, transforming growth factor alpha, heparin-binding EGF, amphiregulin, and betacellulin immunostaining in the nasal epithelium was also obtained; their staining patterns paralleled that of EGFR immunostaining. CONCLUSION: Colocalization of c-erbB receptors and ligands establishes a basis on which to investigate c-erbB receptor- mediated effects in human nasal epithelium. PMID- 11112897 TI - Glucocorticoids preferentially upregulate functional CXCR4 expression in eosinophils. AB - BACKGROUND: Chemokines play an important role in accumulation of eosinophils at allergic inflammatory sites. Systemic administration of glucocorticoids (GCCs) attenuates tissue eosinophilia. In vivo chemokine actions are regulated at levels of both ligand production and receptor expression. The inhibitory effects of GCCs on the production of eosinophil-active chemokines, such as eotaxin, have been well established. However, no data exist regarding the effects of GCCs on expression of chemokine receptors in eosinophils per se. OBJECTIVE: The objective of this study was to investigate the regulation of chemokine receptor expression in eosinophils by GCCs. METHODS: Chemokine receptor expression was analyzed by using flow cytometry and reverse transcriptase PCR. Intracellular Ca(2+) influx and chemotaxis were also analyzed. RESULTS: Eosinophil CCR3 expression was slightly downregulated by 24-hour treatment with dexamethasone (DEX). On the other hand, DEX-treated eosinophils showed markedly increased CXCR4 expression ( approximately 6 fold) in a time- and dose-dependent fashion. In contrast to eosinophils, CXCR4 expression in neutrophils was only marginally affected by DEX. In DEX-treated eosinophils, stromal cell-derived factor 1alpha, a natural ligand for CXCR4, induced a higher level of Ca(2+) influx and chemotaxis compared with untreated cells. CONCLUSION: GCCs upregulate the expression of CXCR4 in eosinophils but not in neutrophils. Because stromal cell-derived factor 1alpha may play a role in baseline trafficking of eosinophils into extravascular tissues rather than recruiting them directly to inflammatory sites, upregulation of CXCR4 by GCCs may mediate the antiallergic property of these drugs by sequestering eosinophils from the circulation to extravascular tissues. PMID- 11112898 TI - Diesel exhaust particles directly induce activated mast cells to degranulate and increase histamine levels and symptom severity. AB - BACKGROUND: The ability of combustion products, such as diesel exhaust particles (DEPs), to modulate the immune system has now been firmly established. DEPs can synergize with allergen at the human upper respiratory mucosa to enhance allergen specific IgE production, initiate a T(H)2 cytokine environment, and even promote primary allergic sensitization. Experiments suggest that these effects result from the initial activation of mast cells to produce IL-4. OBJECTIVE: We sought to demonstrate that in vivo mast cell activation by DEPs plus allergen will also affect the release of classic mast cell mediators and consequently enhance the immediate-phase response. METHODS: Dust mite-sensitive subjects were challenged intranasally with allergen, and symptom scores and histamine levels in nasal wash samples were compared after prechallenge with 0.3 mg of DEPs. RESULTS: If the subjects were first sprayed with DEPs, mean symptom scores rose from 3.7 to 9.9; additionally, only one fifth of the amount of intranasal dust mite allergen was required to induce clinical symptoms. DEPs alone had no effect. The changes in symptoms correlated with histamine levels measured in nasal lavage specimens from these subjects. Although challenge with DEPs alone did not induce histamine release, challenge with both DEPs and allergen resulted in 3-fold higher histamine concentrations than those seen with allergen alone. In contrast, carbon black particles (elemental carbon devoid of chemicals) had no effect. The role of chemicals was confirmed because degranulation of a murine mast cell line by FcepsilonRI cross-linking was increased significantly (by 72%) by the soluble organic chemicals extracted from DEPs. CONCLUSIONS: Overall, these results suggest that exposure to DEPs can enhance the severity of clinical symptoms to allergen by enhancing mast cell degranulation. PMID- 11112899 TI - Frequent de novo mutations and exon deletions in the C1inhibitor gene of patients with angioedema. AB - BACKGROUND: Cases of angioedema with no family history but with functionally low levels of C1 inhibitor and recurrent attacks are often observed. Clinical and biochemical data do not distinguish these cases from proven inherited forms of hereditary angioedema. OBJECTIVE: We sought to test the hypothesis of de novo mutations in patients affected by angioedema without a family history of the disease. METHODS: Among 137 independent kindreds followed for hereditary angioedema, 45 (32.8%) patients with early onset of the disease were registered as sporadic cases. Nineteen patients with unaffected parents were screened for point mutations and microdeletions-insertions by using fluorescence-assisted mismatch analysis. The biologic paternity of these patients was verified by determining their alleles at 4 microsatellite loci. Gross deletions were detected with Southern blot analysis. RESULTS: C1 inhibitor plasma levels measured in both parents of 24 sporadic patients were normal in all but 3 patients. Among the 19 patients studied at the DNA level, 9 de novo single nucleotide substitutions and 6 de novo microdeletions were found. De novo exon deletions were detected in 3 additional patients with Southern blot analysis. CONCLUSIONS: De novo C1inhibitor mutations and exon deletions account for at least 25% of all unrelated cases of angioedema. This finding has implications relevant to the genetic epidemiology and genetic counseling of this disease. The observation that 5 of the 9 de novo point mutations reproduce previously reported changes underlines the presence of multiple hot spots, two of which contain a CpG dinucleotide. PMID- 11112900 TI - Differences in antigen-specific T-cell responses between infants with atopic dermatitis with and without cow's milk allergy: relevance of TH2 cytokines. AB - BACKGROUND: Cow's milk is the most important food antigen in infancy and may lead to acute cutaneous symptoms and atopic dermatitis (AD). The role of circulating allergen-specific T cells in the pathogenesis of food-allergic skin symptoms is still under investigation. OBJECTIVE: This study was designed to analyze the cow's milk protein (CMP)-specific T-cell response at the clonal level in infants with AD and cow's milk allergy (CMA) in comparison with infants with AD without CMA. METHODS: We used an antigen-specific culturing system with autologous B cells as antigen-presenting cells to establish CMP-specific T-cell clones derived from PBMCs in infants with AD. T-cell reactivity, measured by using a lymphocyte stimulation test, and cytokine production, measured by using ELISA, was compared between infants with AD with and without CMA. RESULTS: Both infants with and without allergy to cow's milk had a CMP-specific T helper cell response directed against the major proteins in milk. Analysis of antigen-specific cytokine production showed that this response was T(H)2 skewed in infants with CMA, with production of high levels of IL-4, IL-5, and IL-13. In contrast, infants without CMA had a T(H)1-skewed response, with high levels of IFN-gamma and low levels of IL-4, IL-5, and IL-13. CONCLUSION: These data confirm for the first time at the clonal level that food allergy in infants with AD is associated with production of T(H)2 cytokines by circulating antigen-specific CD4(+) T cells, whereas tolerance to food antigens is associated with low levels of these cytokines. This suggests a key role for the T helper cell-derived T(H)2 cytokines in food allergy related skin symptoms. PMID- 11112901 TI - Characterization of airway inflammation after repeated exposures to occupational agents. AB - BACKGROUND: Little is known about the comparative kinetics of eosinophil recruitment after exposure to low- and high-molecular-weight sensitizers in subjects with occupational asthma (OA). OBJECTIVES: The aims of the study were to investigate the kinetics of changes in inflammatory mediators associated with eosinophil infiltration (IL-5 and eotaxin) and to examine the nature of the airway inflammation induced in response to different types of occupational agents. METHODS: We investigated 15 subjects with OA caused by high- and low molecular-weight agents. The subjects were exposed to increasing doses of the relevant occupational agent over 3 to 4 days until a 20% fall in FEV(1) occurred. Methacholine challenge and sputum induction were performed at the end of each day of exposure. Sputum samples were assessed for differential cell counts, including eosinophils, IL-5, and eotaxin messenger RNA. RESULTS: There was an increase in sputum eosinophils, eotaxin, and IL-5 on the day preceding the occurrence of asthmatic reaction, although there was no change in functional parameters (FEV(1) and PC(20)). Increase in sputum eosinophils was more prominent in subjects exposed to low-molecular-weight agents than to high-molecular-weight agents. CONCLUSION: Changes in eosinophils, IL-5, and eotaxin precede functional changes after exposure to occupational agents in subjects with OA. Eosinophil inflammation is a feature of exposure to both high- and low-molecular-weight agents. Induced sputum may be a useful tool in the early diagnosis of OA. PMID- 11112902 TI - Evidence for a role for IL-5 and eotaxin in activating and recruiting eosinophils in drug-induced cutaneous eruptions. AB - BACKGROUND: Cutaneous drug reactions may be associated with increased numbers of eosinophils in the blood and tissue. However, the factors leading to the generation of eosinophilia have not been fully delineated. OBJECTIVE: The aim of this study was to investigate the in situ expression of IL-5, eotaxin, RANTES, monocyte chemoattractant protein 3, and IL-8 together with the appearance of eosinophils in acute cutaneous drug reactions. METHODS: Skin biopsy specimens were obtained from drug-induced maculopapular exanthems (n = 9), from normal skin of control subjects (n = 9), and from the skin of patients with psoriasis (n = 8). The in situ expression of IL-5, eotaxin, RANTES, monocyte chemoattractant protein 3, and IL-8 was analyzed by using immunohistochemistry. Furthermore, the corresponding numbers of eosinophils were determined in the blood and skin sections. RESULTS: Compared with normal skin and psoriatic skin, a significantly higher number of eosinophils was found both in the blood and tissue of patients with a drug-induced exanthem. In comparison with normal skin, immunoreactivity for IL-5 and all the chemokines was also significantly enhanced in drug-induced exanthem, whereas significant differences in psoriatic were only observed for IL 5 and eotaxin. CONCLUSION: Our data indicate that IL-5 and eotaxin may particularly contribute to the activation and recruitment of eosinophils and thereby play an important pathogenic part in the development of skin inflammation in drug-induced maculopapular exanthems. PMID- 11112903 TI - Immediate allergic reactions to cephalosporins: cross-reactivity and selective responses. AB - BACKGROUND: After penicillins, cephalosporins are the most important beta-lactams inducing IgE-mediated reactions. Responses may be selective or cross-reactive with common beta-lactam determinants. Unlike determinants derived from benzylpenicillin, cephalosporin allergenic determinants have not been properly identified, even though a wide variety of these beta-lactams is currently used. OBJECTIVE: We sought to evaluate the IgE response in subjects with immediate allergic reactions to injectable cephalosporins and to assess their reactivity to different penicillins and cephalosporins. METHODS: We studied 30 subjects with immediate reactions to one or more of the following cephalosporins: ceftriaxone, cefotaxime, ceftazidime, and cefuroxime. Skin tests and in vitro-specific IgE antibody assays were performed for major and minor determinants of penicillin G, amoxicillin, and ampicillin, as well as for the culprit cephalosporins. Responses to cephalosporins other than the culprit ones were also studied by using skin testing. RESULTS: Twenty-six patients (group A, 86.7%) displayed skin test and RAST negativity to penicillin determinants and skin test positivity to cephalosporins, with RAST confirmation in 9 patients. Four subjects (group B, 13.3%) had a positive response to penicillin determinants. In group A two patterns of reactivity were observed: one characterized by a response only to the culprit cephalosporin (n = 15, 57.7%) and the other by positive responses to different cephalosporins, including the responsible cephalosporins (n = 11, 42. 3%). CONCLUSION: Most patients with a history of immediate reactions to cephalosporins are sensitized to determinants generated only by cephalosporins (group A), although a small percentage react to penicillin determinants (group B). Some patients from group A responded only to the culprit cephalosporin, but others reacted to different cephalosporins. These findings can be explained in terms of either selective response to unique determinants or cross-reactivity. PMID- 11112904 TI - Insect sting-inflicted systemic reactions: attitudes of patients with insect venom allergy regarding after-sting behavior and proper administration of epinephrine. AB - BACKGROUND: Patients with insect venom allergy are at higher risk for development of a recurrent systemic reaction after re-sting. This risk significantly decreases with venom immunotherapy. Patients with insect venom allergy should be able to distinguish a life-threatening systemic reaction from all other various reactions after an insect sting. Accidental epinephrine injection by EpiPen has been reported in the past. Therefore patients with venom allergy should also be well trained in self-administration of their epinephrine when needed. OBJECTIVE: Our objective was to assess patients' attitudes regarding after-sting behavior and their capability to correctly self-administer the epinephrine autoinjector. METHODS: All patients with venom allergy attending our allergy unit either before commencement of or during venom immunotherapy answered a questionnaire addressing various aspects of their intended after-sting behavior. Using an EpiPen trainer device, patients' performance of EpiPen self-administration was evaluated. RESULTS: Ninety-six patients participated in the study. Seventy-six of them were equipped with an EpiPen device. Less than 30% of these patients carried it at all times. After re-sting, 50 (54%) patients planned to wait for the development of other symptoms before taking any further action. Twenty-two percent of the patients said that after re-sting they would immediately administer their EpiPen. Proper EpiPen administration technique was demonstrated by 44% of the patients. Having not reached the maintenance dose correlated with a better compliance with carrying of the EpiPen. EpiPen instruction provided by an allergist correlated with a better EpiPen administration technique by the patients. CONCLUSION: Many patients with venom allergy hold wrong ideas about after-sting behavior. Compliance with carrying EpiPen at all times and the ability to correctly administer it are both poor in most patients. Thorough and probably repeated instruction, both written and oral, provided by knowledgeable physicians is mandatory. PMID- 11112905 TI - Surface membrane antigen alteration on blood basophils in patients with Hymenoptera venom allergy under immunotherapy. AB - BACKGROUND: Venom immunotherapy (VIT) provides widespread protection against systemic anaphylactic reactions after a sting of the respective insect. This effect is attributed to a shift from T(H)2 to T(H)1. However, because basophils also produce and release cytokines, such as IL-4 and IL-13, they may be part of the cytokine network. The cytokines may regulate basophilic granulocytes, as suggested by the presence of cytokine receptors IL-2Ralpha, GM-CSFRalpha, IL 1RII, IL-3R, IL-4R, IL-5R, and IL-6R on basophils from nonallergic donors. OBJECTIVE: The purpose of this study was to demonstrate that human basophils from subjects allergic to wasp venom undergoing VIT are regulated by cytokines, as shown by the alteration of the expression of cytokine receptors (and other markers). METHODS: The expression of the surface interleukin receptors and activation antigens on basophils from 19 nonallergic subjects and 48 patients with wasp venom allergy was investigated before, immediately after, and 1 week after VIT (20 patients only). RESULTS: Basophilic granulocytes in allergic subjects, compared with those in healthy persons, showed elevated expression of CD32 (FcgammaRII), CD122 (IL-2Rbeta), CD124 (IL-4Ralpha), CD130 (IL-6 and 11Rbeta), CD154 (CD40L), and HLA-DR. Activation of basophils clearly increased during VIT indicated by increased expression of CD32, CD33, CD35 (CR1), CD63, CD116 (GM-CSFRalpha), CD122, CD124, CD130, and CD154. HLA-DR expression also tended to increase. The expression of IL-5R (CD125) decreased. A significant decrease of the basophilic surface antigens CD11c, CD32, CD35, CD63, CD116, CD122, CD124, CD130, and CD132 (interleukin receptor gamma) was detected 1 week after the end of rush VIT. CONCLUSION: The rise in CD63 during VIT indicates a partial basophil degranulation with release of stored protein mediators, including IL-4. IL-4 may cause a transient upregulation of different surface antigens in an autocrine manner. Thereafter, cytokines released by T cells, which as a result of VIT have changed from a T(H)2 type to a more T(H)1 type, downregulate the activation of the basophilic granulocytes. PMID- 11112906 TI - Atypical fatal hypocomplementemic urticarial vasculitis with involvement of native and homograft aortic valves in an African American man. PMID- 11112907 TI - BCG scar diameter and asthma: a case-control study. PMID- 11112908 TI - Latex allergy in spina bifida: at the turning point? PMID- 11112909 TI - Selective cyclo-oxygenase 2 inhibitor in patients with aspirin-induced asthma. PMID- 11112910 TI - IgE-mediated reaction to a banana-flavored drug additive. PMID- 11112911 TI - Contact allergy to polidocanol, 1992 to 1999. PMID- 11112912 TI - Monitoring peak flow using control charts: comments from experience. PMID- 11112914 TI - Efficacy and safety overview of a new inhaled corticosteroid, QVAR (hydrofluoroalkane-beclomethasone extrafine inhalation aerosol), in asthma. AB - Chlorofluorocarbon (CFC)-containing inhalers are gradually being phased out and replaced with hydrofluoroalkane (HFA)-based alternatives. The reformulation provided the opportunity to improve the inhalation technology and physical characteristics of corticosteroid formulations. QVAR contains HFA-beclomethasone dipropionate (HFA-BDP) with the steroid in solution rather than suspension, which, in combination with improved inhaler technology, produces an extrafine aerosol with a mass median aerodynamic diameter of 1.1 microm (smaller than the 3.5-4.0 microm found with CFC-BDP). It was predicted and demonstrated that the smaller particle size of QVAR would be deposited in the lung to a greater extent than that found with CFC-BDP, particularly in the small airway, a major site of inflammation. Increased lung deposition of QVAR permits a reduction in dosage relative to CFC-BDP. Clinical evidence confirms that adult and elderly patients required approximately half the dose of QVAR to achieve the same degree of asthma control as with CFC-BDP. In long-term assessments, patients taking CFC-BDP could be switched to QVAR at half the daily dose without exacerbation of their asthma symptoms. QVAR was associated with a low overall incidence of side effects and, at the maximum recommended dose of 640 microg/d, caused no more adrenal suppression than 672 microg/d CFC-BDP. PMID- 11112917 TI - The gifts we give and receive. PMID- 11112916 TI - Primary care and primary health care: defining what we do. PMID- 11112918 TI - Early-onset neonatal group B streptococcal infection: implications for practice. AB - Group B streptococcus (GBS) is the leading bacterial infection associated with morbidity and mortality of newborns in the United States. Most neonatal infections can be prevented through the use of intrapartum antimicrobial prophylaxis in women who are at increased risk for transmitting infection to their newborns. However, prevention strategies have not been implemented widely or consistently, and the incidence of neonatal GBS disease has not declined. An understanding of GBS epidemiology, clinical presentation, and prevention strategies enhances the PNP's decision-making skills in the nursery and strengthens the PNP's ability to evaluate and compare new approaches to GBS prevention. PMID- 11112919 TI - Comprehensive primary care follow-up for premature infants. AB - Advances in perinatal and neonatal care have led to an increased incidence of survival of premature infants. Although most premature infants have normal outcomes, they are at increased risk for morbidity and mortality and require comprehensive primary care follow-up after they are discharged from the hospital. This article will review guidelines for general follow-up of premature infants and the associated problems related to prematurity. General follow-up is performed by the pediatric nurse practitioner, with subspecialty consultant referrals as needed. Knowledge of the problems of prematurity and treatment regimes will assist the pediatric nurse practitioner in providing high-quality care to these high-risk infants. PMID- 11112920 TI - Making the transition: pediatric lung transplantation. AB - INTRODUCTION: Little information exists regarding how parents whose children have undergone lung transplantation perceive the meaning of their return home to face the reality of the posttransplant experience. METHOD: A naturalistic approach was selected to investigate the meaning of the experience from the perspective of the parents. Fifteen parents of 12 children were interviewed. Colaizzi's method of phenomenologic analysis guided the analysis of the interview data. RESULTS: A theme cluster, "making the transition," emerged from the data. The themes that reflected parents' perceptions of the issues they faced in the posttransplant period included reuniting the family, assuming a new role, returning to school, facing the risk of infection, facing the threat of rejection, and striking a balance. DISCUSSION: Parents' perceptions of the situation reflected the shifting impact of uncertainty on their daily lives and coping strategies. The need exists to recognize the parents' unique and changing needs and to implement individualized nursing care to meet their needs. PMID- 11112921 TI - Perceived stress, social support, and affectionate behaviors of adolescent mothers with infants in neonatal intensive care. AB - INTRODUCTION: This article focuses on affectionate behaviors of adolescent mothers with their infants in a neonatal intensive care unit (NICU). Hypotheses derived from behavioral science theory posited the direct influence of social support and perceived stress on affectionate behaviors, the statistical interaction of social support and perceived stress on affectionate behaviors, and perceived stress as a mediator of the relationship between social support and affectionate behaviors. METHOD: Subjects were enrolled from July 1993 through September 1994. Information about perceived stress and social support was obtained twice by means of an interview. Affectionate behaviors were measured by NICU nurse observations. Analyses were conducted on subsamples ranging from 57 to 107 subjects. RESULTS: All hypotheses were rejected. Neither social supports nor perceived stress were related to affectionate behaviors, and no statistical interactions among the 3 variables were identified. DISCUSSION: The findings are considered in the context of the methodology used, stress and social support theory, and implications for practice and future research. PMID- 11112922 TI - Measuring imaging ability in children. AB - INTRODUCTION: Guided imagery has been suggested as an intervention to help children cope with noxious symptoms associated with medical care. A measure of imaging ability, that is, the ability to generate vivid mental images and to experience those images as if they were real, could be helpful in identifying children most likely to succeed in relieving symptoms with guided imagery. The purpose of this study was to test psychometric properties of a new instrument, the Kids Imaging Ability Questionnaire (KIAQ). METHOD: Three expert clinicians and researchers were asked to review the KIAQ to assess content validity. A convenience sample of 58 children were invited to complete the questionnaire twice to obtain data for tests of reliability and criterion-related validity. RESULTS: Content validity, internal consistency (alpha =.75-.76), and test-retest reliability (r =.73) were acceptable. Criterion-related validity using the Singer Fantasy Proneness Interview as a standard was poor (rho =.31-.46). DISCUSSION: Some psychometric properties were acceptable; however, continued research will be necessary to test validity of the questionnaire and demonstrate a relationship between KIAQ score and success with imagery. With continued research, pediatric nurses may use the KIAQ in practice to identify children most likely to benefit from guided imagery. PMID- 11112923 TI - Pediatric end-stage renal disease: incidence, management, and prevention. AB - Although pediatric end-stage renal disease affects a small number of children, it is a serious and growing health problem in the United States. In the past decade, the incidence of the disease has increased steadily in all racial groups. However, poor and minority children are disproportionately affected. Recent research results make it clear that appropriate prenatal and pediatric care can reduce the incidence of this complex and expensive-to-treat condition. PMID- 11112924 TI - Dietary supplements for children. PMID- 11112925 TI - Hypertensive emergencies in children. PMID- 11112926 TI - Adolescent with upper extremity numbness after exercise. PMID- 11112927 TI - Marketing nurse practitioners is the key to lobbying. PMID- 11112931 TI - Personality traits and smoking in patients with obsessive-compulsive disorder. AB - As opposed to other psychiatric populations, subjects with obsessive-compulsive disorder (OCD) smoke less than the general population. The present study aims at further investigating the relationship between smoking in OCD subjects and personality traits. Sixty-four subjects with OCD were interviewed concerning their smoking habits. Personality traits were evaluated using the Karolinska Scales of Personality, and specific obsessive-compulsive personality traits were elicited through self-report questionnaires. Non-smokers were more easily fatigued, more inclined to worry, more remorseful, less self-confident, less impulsive and became uneasy more frequently when urged to speed up, than smokers with OCD. Additionally, non-smokers fulfilled significantly more obsessive compulsive personality disorder criteria as compared to the smokers (P < 0.001). We propose a clinical subtype of OCD related to non-smoking, psychasthenia, anxiety, and pronounced obsessive-compulsive personality disorder traits. PMID- 11112928 TI - The garden of good and evil: pharmaceutical companies and the perspective practices of PNPs. PMID- 11112932 TI - Hemispheric function in disorganized type schizophrenia: performance on the quality extinction test. AB - We assessed hemisphere function in right-handed male chronic, disorganized type schizophrenic patients (N = 60, age range 18-45 years) using the Quality Extinction Test (QET), in comparison to 20 right-handed male healthy controls in the same age range. The QET analysis discriminated between the disorganized schizophrenic patients and the controls. QET results indicated that chronic schizophrenic patients were less sensitive to tactile stimuli in both hands as compared to controls. Furthermore, the sensitivity to tactile stimuli of the left hand was less than that of the right hand in the schizophrenic patients. In contrast, in the normal controls the sensitivity was similar in both hands. These results indicate possible right hemisphere dysfunction together with disturbance in interhemispheric transmission through the corpus callosum in chronic, disorganized type schizophrenic patients. PMID- 11112933 TI - Evaluation of the supervisory system in elderly subjects with and without disinhibition. AB - Disinhibition and irritability, defined as loss of behavioral and emotional control, are frequent in the elderly. The working hypothesis for this study was that these disorders are associated with a cognitive alteration of control processes that manifests as non-routine behavior because of the dysfunction of a general executive component known as the supervisory attentional system (SAS). METHODS: A total of 28 elderly subjects with mild cognitive impairment were recruited and divided into two groups using the Neuropsychiatric Inventory. Fourteen subjects were allocated to the disinhibited group and 14 subjects matched for age, sex and educational level formed a disinhibition-free control group. The neuropsychological battery included the following tests: Mini Mental Score Evaluation, Boston Naming test, Token test, Trail Making and Verbal Fluency. Two tasks were specifically designed to stress the SAS: 1) A specific verbal sentence arrangement task in which subjects had to use sequential reasoning with verbal material. Each test sequence consisted of a series of words shown in jumbled order. The construction of some sequences had to be done by using familiar routine associations (valid conditions). In contrast, other sequences required the overriding selection of familiar routine associations, which were inappropriate within the general context of the task (invalid conditions). 2) Using the Continuous Performance Test, four aspects were evaluated: sustained, selective, preparation and suppressive attention. RESULTS: The only group differences in neuropsychological test results were the following: 1) the sentence arrangement task. In comparison with the control group, the disinhibited group was impaired in invalid conditions and the calculated difference between the number of correct responses in invalid conditions minus that in valid conditions was significantly higher; and 2) the CPT. Disinhibited subjects had a significantly lower number of hits, exclusively in the 'suppressive attention' paradigm. These results suggest that subjects with disinhibition have impaired supervisory system function. PMID- 11112934 TI - The suicide assessment scale: an instrument assessing suicide risk of suicide attempters. AB - The Suicide Assessment Scale (SUAS), a scale constructed to measure suicidality over time, was administered to 191 suicide attempters. Its predictive validity was tested. SUAS ratings were compared to ratings from other scales, and related to age and psychiatric diagnoses including co-morbidity. Eight patients committed suicide within 12 months after the SUAS assessment. Apart from advanced age, high scores in the SUAS were significant predictors of suicide. From a receiver operating characteristic (ROC) analysis, we identified cutoff SUAS scores which alone and in combination with certain diagnostic and demographic factors are of apparent value in the clinical evaluation of suicide risk after a suicide attempt. PMID- 11112935 TI - Valpromide increases amplitude of heart rate circadian rhythm in remitted bipolar and unipolar disorders. A placebo-controlled study. AB - The aim of this study was to investigate for the effects of valpromide on heart rate circadian rhythm in remitted recurrent unipolar and bipolar disorders (DSM III-R). It consisted of a comparative, randomized, double-blind, repeated cross over study of valpromide versus placebo over four four-week periods. The primary evaluation criteria was heart rate (HR). Secondary criteria comprised motor activity (MA) and the Bech and Rafaelsen mania assessment scale, Horne and Ostberg questionnaire, Montgomery and Asberg depression rating scale, Spiegel questionnaire, a sleep diary, and Clinical Global Impression. Fifteen patients were included, giving 60 one-month periods (30 valpromide periods and 30 placebo periods). Cosinor analysis of HR and MA data revealed a difference in amplitude (P = 0.037, analysis of variance, one-tailed test). The clinical sleep study shows that the duration of sleep was greater with valpromide than with placebo (P = 0.007, one-tailed test). Similarly, evaluation of the quality of sleep by patients themselves showed valpromide to be superior to placebo (P = 0.045, one tailed test). The results of analysis of the Spiegel questionnaire also confirm the superiority of valpromide over placebo. Safety and compliance were comparable for the active drug and the placebo. In conclusion, the relatively small sample size requires cautious interpretation of this study. Nevertheless, these initial results show a definite effect of valpromide on a biological rhythm that leads one to suppose that it may be effective through a 'synchronizing' effect. PMID- 11112936 TI - Adjunctive gabapentin treatment of bipolar disorder. AB - INTRODUCTION: The aim of this study was to analyze the effectiveness of gabapentin administration to bipolar patients who had an incomplete response to other mood stabilizers. SUBJECTS AND METHODS: Twenty-two RDC bipolar 1 and II patients were assessed by means of the SADS and entered if they gave their consent to participate. All them had suffered from frequent relapses, subsyndromal features (mostly depressive) and incomplete response to other drugs. They all received open-label increasing doses of gabapentin until clinical response. The patients were assessed through the CGI-BP and a specific questionnaire at baseline and at 12 weeks of follow-up. RESULTS: Six out of the 22 patients dropped out for various reasons (four because of relapse, one because of side effects and one more because of poor compliance). Eight of the 16 patients that completed the 12-week follow-up showed at least two stages of improvement in the CGI. Using the last observation-carried forward analysis, the improvement was statistically significant for the depression subscale, and apparently related to social functioning, irritability and anxiety. Only one patient dropped out because of intolerance (mild rash). The mean dose of gabapentin was 1,310 mg/day. CONCLUSION: Gabapentin may be a useful drug for the add-on treatment of bipolar patients with poor response to other mood stabilizers. Gabapentin may improve depressive residual symptoms such as irritability, social withdrawal or anxiety. These results should be confirmed in randomized clinical trials. PMID- 11112937 TI - Improvement of tardive dyskinesia in a bipolar patient with olanzapine. PMID- 11112939 TI - Ensuring shelter in Eastern Europe. PMID- 11112940 TI - A vision for public health development. PMID- 11112941 TI - The social patterning of teenage pregnancy. PMID- 11112942 TI - The life course, childhood housing conditions and adult health. PMID- 11112943 TI - The completeness of AIDS surveillance in Europe: difficulties and new challenges. PMID- 11112944 TI - Population change and mortality in men and women. PMID- 11112945 TI - Childhood housing conditions and later mortality in the Boyd Orr cohort. AB - STUDY OBJECTIVES: To examine associations between five measures of housing conditions during childhood and subsequent mortality from all causes, coronary heart disease, stroke, and cancer. DESIGN: Historical cohort study. SETTING: Data on housing conditions were collected from survey centres in 16 areas of England and Scotland. PARTICIPANTS: Children of families participating in the Carnegie Survey of Family Diet and Health in pre-war Britain (1937-1939). Analyses are based on a subset of 4168 people who were traced and alive on 1 January 1948. MAIN RESULTS: Poorer housing conditions were generally associated with increased adult mortality. After adjustment for childhood and adult socioeconomic factors, statistically significant associations were only found between lack of private indoor tapped water supply and increased mortality from coronary heart disease (hazard ratio 1.73, (95% CI 1.13, 2.64); and between poor ventilation and overall mortality (hazard ratio for people from households with poorest ventilation relative to best ventilation 1.30, 95% CI 0.97, 1.74). CONCLUSIONS: This study provides evidence that associations between housing conditions in childhood and mortality from common diseases in adulthood are not strong, but are in some respects distinguishable from those of social deprivation. PMID- 11112946 TI - Teenage childbearing in Great Britain and the spatial concentration of poverty households. AB - STUDY OBJECTIVE: To investigate the association between the spatial concentration of deprived households and teenage non-marital childbearing. Associations with area deprivation are tested before and after allowing for levels of personal deprivation. DESIGN AND SETTING: The individual data are derived from the 2% sample of anonymised records (SAR) from the census of 1991 in Great Britain, and are combined with area data from the 278 districts of residence identifiable in the SAR. PARTICIPANTS: Sample is restricted to unmarried women living at home (with at least one parent) and aged 16 to 19. MAIN RESULTS: The results suggest generally higher risk of teenage childbearing for women who are economically inactive, women from households with no access to a car or households resident in local authority accommodation. Without adjusting for personal circumstances, the risk of teenage pregnancy shows a clear, significant and approximately linear association with social deprivation of area of residence in 1991. Residual analysis shows that many urban areas have much higher levels of teenage childbearing than expected. When adjustment is made for personal disadvantage the simple association with local area deprivation is attenuated. A higher risk of teenage childbearing is still seen in urban areas while the areas having the highest negative differentials are heterogeneous. CONCLUSIONS: Both individual and spatial characteristics are important in influencing levels of teenage childbearing. Teenage childbearing shows an association with residence in more deprived areas. The association seems to be largely because residence in more deprived areas is associated with personal disadvantage, which increases the risk of teenage childbearing. Area characteristics are of lesser significance in determining teenage non-marital childbearing than individual and household characteristics. PMID- 11112947 TI - Decline of the relative risk of death associated with low employment grade at older age: the impact of age related differences in smoking, blood pressure and plasma cholesterol. AB - STUDY OBJECTIVE: To explore whether the observed age related decline in the relative risk of death associated with low employment grade can be explained by the profiles of smoking, blood pressure and plasma cholesterol changing differently with age between the employment grades. DESIGN: Prospective cohort study with 25 years of mortality follow up. SETTING: Whitehall study. PARTICIPANTS: There were 16,984 men aged 40 to 69 years at baseline with complete information on smoking, blood pressure and plasma cholesterol. MAIN RESULTS: The relative risk of death associated with low employment grade decreased from 2.1 at 55-59 years of age to 1.3 at 85-89 years of age. Adjustment for smoking status and blood pressure, attenuated the age related decline of the relative risk by 18% and 3% respectively; adjustment for plasma cholesterol increased the decline by 3%. Taken together, these risk factors explain 20% of the observed age related decline. CONCLUSIONS: A small part of the observed age related decline in the relative risk of death associated with low employment grade can be explained by differential changes in the profiles of smoking, blood pressure and plasma cholesterol with age between the employment grades. PMID- 11112948 TI - Dietary patterns among a national random sample of British adults. AB - STUDY OBJECTIVES: To identify groups within the UK male and female population who report similar patterns of diet. DESIGN: National representative dietary survey, using seven day weighed dietary records, of men and women aged 16-64 years living in private households in Great Britain in 1986-7. Cluster analysis was used to aggregate participants into diet groups. SETTING: Great Britain. PARTICIPANTS: 1087 men and 1110 women. RESULTS: 93% of men and 86% of women fell into one of four distinct diet groups. Among men the most prevalent diet group was "beer and convenience food" (34% of the male population); second was "traditional British diet" (18%); third was "healthier but sweet diet" (17.5%) and fourth was "healthier diet " (17%). Among women, the most prevalent diet group was " traditional British diet" (32%); second, was "healthy cosmopolitan diet" (25%); third was a "convenience food diet" (21%); and fourth was "healthier but sweet diet" (15%). There were important differences in nutrient profile, sociodemographic and behavioural characteristics between diet groups. CONCLUSIONS: Cluster analysis identified four diet groups among men and four among women, which differed not only in terms of reported dietary intakes, but also with respect to nutrient, social and behavioural profiles. The groups identified could provide a useful basis for development, monitoring and targeting of public health nutrition policy in the UK. PMID- 11112949 TI - Life expectancy as a summary of mortality in a population: statistical considerations and suitability for use by health authorities. AB - OBJECTIVE: To investigate the sampling distribution and usefulness of expectation of life in comparisons of mortality at health district level or below. DESIGN: Derivation of a formula for the variance of the expectation of life, confirmation of the result and generation of the sampling distribution by Monte Carlo simulation; comparison of expectation of life with standardised mortality ratio (SMR) and other summary indices of mortality. SETTING: A health district in Trent Region, England. SUBJECTS: Routinely available mortality statistics at electoral ward level and above. MAIN RESULTS: Given reasonable and simple assumptions the sampling distribution of the expectation of life is approximately normal. Expectation of life shows a high negative correlation with SMR even if the oldest age band for the SMR is open ended. CONCLUSIONS: Where sampling error is an issue, inference concerning differences in mortality rates between populations can be based on expectation of life, which is better for illustrative purposes than SMR. The formula for the variance of the expectation of life is more complex however. If the final age band is open ended, its lower bound should be as high as possible to avoid misleading results caused by hidden differences in age structure. PMID- 11112950 TI - A multilevel analysis of the effects of rurality and social deprivation on premature limiting long term illness. AB - STUDY OBJECTIVE: To examine the geographical variation in self perceived morbidity in the south west of England, and assess the associations with rurality and social deprivation. DESIGN: A geographically based cross sectional study using 1991 census data on premature Limiting Long Term Illness (LLTI). The urban rural and intra-rural variation in standardised premature LLTI ratios is described, and correlation and regression analyses explore how well this is explained by generic deprivation indices. Multilevel Poisson modelling investigates whether Customized Deprivation Profiles (CDPs) and area characteristics improve upon the generic indices. SETTING: Nine counties in the south west of England PARTICIPANTS: The population of the south west enumerated in the 1991 census. MAIN RESULTS: Intra-rural variation is apparent, with higher rates of premature LLTI in remoter areas. Together with high rates in urban areas and lower rates in the semi-rural areas this indicates the existence of a U shaped relation with rurality. The generic deprivation indices have strong positive relations with premature LLTI in urban areas, but these are a lot weaker in semi-rural and rural locations. CDPs improve upon the generic indices, especially in the rural settings. A substantial reduction in unexplained variation in rural areas is seen after controlling for the level of local isolation, with higher isolation, at the wider geographical scale, being related to higher levels of LLTI. CONCLUSIONS: This study highlights the need to treat rural areas as heterogeneous, although this has not been the tendency in health research. Generic deprivation indices are unlikely to be a true reflection of levels of deprivation in rural environments. The importance of CDPs that are specific to the area type and health outcome is emphasised. The significance of physical isolation suggests that accessibility to public and health services may be an important issue, and requires further research. PMID- 11112951 TI - How complete is AIDS surveillance in Europe? An eagle eye comparison with mortality data. AB - OBJECTIVES: Preliminary assessment of the adequacy of AIDS surveillance efforts in Europe by comparing data from two official sources-AIDS surveillance and mortality statistics. METHODS: The study used ENAADS (European Non-Aggregate AIDS Data Set) data compiled by the European Centre for the Epidemiological Monitoring of AIDS in St Maurice, France, and mortality statistics from WHO. As ENAADS provides information about AIDS incidence as well as AIDS mortality, both series were compared with WHO mortality data. Western European countries with more than 1000 adult AIDS cases as of July 1997 were included in the cross country comparative analyses. RESULTS: AIDS surveillance and mortality statistics in Europe depict four different patterns: (1) high overall concordance (Austria, Italy, Switzerland); (2) concordance between incidence by ENAADS and mortality by WHO, but a delay in mortality reporting in ENAADS (France, Spain); (3) more cases in WHO mortality data than in ENAADS data (Germany, Portugal); (4) more cases in ENAADS data than in WHO mortality data (Sweden, United Kingdom, Greece, Belgium). CONCLUSIONS: National AIDS surveillance systems in Europe exhibit important differences in terms of completeness and functionality. New challenges such as the introduction of effective but expensive and complex treatments will exert demands on surveillance efforts. Countries with discrepant AIDS and mortality data should try to improve and update their surveillance systems. PMID- 11112952 TI - Air pollution and emergency hospital admissions for cardiovascular diseases in Valencia, Spain. AB - STUDY OBJECTIVE: To estimate the short-term association between air pollution levels and emergency hospital admissions for cardiovascular diseases in Valencia, within 1994-1996 period. DESIGN: Daily levels of air pollution and emergency admissions for cardiovascular diseases were related to using an ecological time series design. The number of admissions was obtained from the hospital records database. Selected groups of causes were all cardiovascular diseases, heart admissions, and admissions for cerebrovascular diseases. The number of admissions for digestive diseases was used as control. Pollutants were black smoke, sulphur dioxide (SO(2)), nitrogen dioxide (NO(2)), carbon monoxide (CO) and ozone (O(3)). Magnitude of association was estimated by Poisson autoregressive regression. Estimations were calculated according the hottest (May to October) and the coldest (November to April) periods. SETTING: City of Valencia, Spain, about 750,000 inhabitants. PARTICIPANTS: People being admitted to the two major hospitals in the city, with a catchment area of nearly 400,000 inhabitants. MAIN RESULTS: For the whole period, a significant association for SO(2)-24 h was found so a rise in its levels of 10 microg/m(3) was associated with an increment of 3% (95%CI 0.4 to 5.7%) in the expected number of cardiovascular admissions. A significant association for black smoke, SO(2)-24 h, SO(2)-1 h, and CO-1 h was found in the hottest semester. All these associations were verified with a lag of two days. The estimates of the associations for particles, SO(2), and CO were affected by the inclusion of the other pollutants in their models. NO(2) was independently associated with cerebrovascular admissions. There were no significant associations between air pollution and admissions for digestive diseases. CONCLUSIONS: Current levels of air pollution and emergency cardiovascular admissions are significantly related in Valencia. PMID- 11112953 TI - A characterisation of patient drop outs in a cohort of HIV positive homosexual/bisexual men and intravenous drug users. PMID- 11112954 TI - Tobacco consumption and bladder cancer in non-coffee drinkers. PMID- 11112955 TI - Botulinum toxin type A (BOTOX) for treatment of migraine headaches: an open-label study. AB - OBJECTIVE: The object of this clinical experience was to evaluate the correlation between pericranial botulinum toxin type A (BOTOX, Allergan Corp, Irvine, CA) administration and alleviation of migraine headache symptoms. STUDY DESIGN AND SETTING: A nonrandomized, open-label study was performed at 4 different test sites. The subjects consisted of 106 patients, predominantly female, who either (1) initially sought BOTOX treatment for hyperfunctional facial lines or other dystonias with concomitant headache disorders, or (2) were candidates for BOTOX treatment specifically for headaches. Headaches were classified as true migraine, possible migraine, or nonmigraine, based on baseline headache characteristics and International Headache Society criteria. BOTOX was injected into the glabellar, temporal, frontal, and/or suboccipital regions of the head and neck. Main outcome measures were determined by severity and duration of response. The degrees of response were classified as: (1) complete (symptom elimination), (2) partial > or =50% reduction in headache frequency or severity), and (3) no response [neither (1) nor (2)]. Duration of response was measured in months for the prophylactic group. RESULTS: Among 77 true migraine subjects treated prophylactically, 51% (95% confidence interval, 39% to 62%) reported complete response with a mean (SD) response duration of 4.1 (2.6) months; 38% reported partial response with a mean (SD) response duration of 2.7 (1.2) months. Overall improvement was independent of baseline headache characteristics. Seventy percent (95% confidence interval, 35% to 93%) of 10 true migraine patients treated acutely reported complete response with improvement 1 to 2 hours after treatment. No adverse effects were reported. CONCLUSIONS: BOTOX was found to be a safe and effective therapy for both acute and prophylactic treatment of migraine headaches. Further research is needed to explore and develop the complete potential for the neuroinhibitory effects of botulinum toxin. PMID- 11112956 TI - Apoptosis in the developing human cricoid cartilage: a pilot study. AB - Apoptosis is widely recognized as a major phenomenon in normal development. Deficiencies in this process may lead to developmental abnormalities such as congenital subglottic stenosis. We studied apoptosis using in situ end labeling of the 3'-OH ends of fragmented DNA in 5 progressively older, normal, human cricoid cartilage specimens. Results show that apoptosis is a very active process in fetal and neonatal tissue. The process gradually slows with advancing age. In the 4- and 13-year-old specimens, minimal to no apoptosis was seen. We conclude that apoptosis plays a critical role in the intraluminal and extraluminal expansion of the cricoid cartilage. PMID- 11112957 TI - A meta-analysis of dexamethasone use with tonsillectomy. AB - OBJECTIVE: To determine the quantitative impact of intravenous dexamethasone on recovery after tonsillectomy using established principles for meta-analysis. STUDY DESIGN/SETTING: Double-blind randomized-control trials in which subjects were treated identically except for the presence or absence of perioperative intravenous dexamethasone. Six articles met inclusion criteria. Two investigators extracted data regarding postoperative emesis and return to a soft/regular diet. RESULTS: Pooled analysis using a random effects model revealed a 27% decrease (P<0.00001) in postoperative emesis attributable to dexamethasone (95% CI, 12% to 42%). Dexamethasone increased the tolerance of a soft/regular diet at 24 hours by 22% (P< 0.001), but studies were heterogenous with low precision (95% CI, 1% to 44%). CONCLUSION: To prevent emesis in 1 child after tonsillectomy, approximately 4 children must receive perioperative dexamethasone. An additional benefit is earlier tolerance of a soft/regular diet, but low precision and heterogeneity among studies preclude definitive conclusions. SIGNIFICANCE: Perioperative dexamethasone administration had a positive impact on recovery from tonsillectomy. PMID- 11112958 TI - Chronic rhinosinusitis: allergy and sinus computed tomography relationships. AB - The management of chronic or recurrent rhinosinusitis problems is multifaceted and should include consideration of contributory and potentially correctable medical and anatomic factors. To date, the relationship between allergy and rhinosinusitis has not been clearly defined. The purpose of this study is to improve understanding of the relative roles of perennial and seasonal allergens in the cause of chronic rhinosinusitis. A retrospective review of 200 consecutive patients was carried out on patients who had chronic rhinosinusitis refractory to medical therapy and who subsequently underwent functional endoscopic sinus surgery. All of these patients had allergy testing for common perennial and seasonal inhalant allergens before surgery. Each patient had sinus CT imaging before undergoing the surgery. The CT scans of each patient were staged according to a validated, standardized grading system by investigators blinded to allergic profile. Allergy testing indicated that 84% of all patients tested positive for allergies. Moreover, 60% of all patients had significant allergic sensitivity; 52% of all patients had multiple allergen sensitivities. Furthermore, there was a predominance of perennial allergens, especially house dust mite over seasonal allergens. The vast majority of our patients undergoing functional endoscopic sinus surgery had concomitant allergy. This study highlights the potential contribution of perennial allergies to the development of rhinosinusitis. Given this direction, future studies may reveal that in the care of patients with perennial allergic rhinitis, early intervention with identification of the offending allergen(s), and subsequent treatment through avoidance, pharmacotherapy, and/or immunotherapy may help in the prevention of recurrent and chronic rhinosinusitis. PMID- 11112959 TI - Management of nasopharyngeal stenosis after uvulopalatoplasty. AB - OBJECTIVE: The objective of this study is to evaluate the management of nasopharyngeal stenosis (NPS) with the CO(2) laser and a customized nasopharyngeal obturator. STUDY DESIGN: An 8-year retrospective study based at a tertiary care teaching hospital consisting of 18 patients with NPS after uvulopalatoplasty treated over an 8-year period with the CO(2) laser and a nasopharyngeal obturator. Patients with grade I stenosis were treated in the office and did not require a nasopharyngeal obturator. More severe cases (grades II and III) were treated in the operating room and required a nasopharyngeal obturator. RESULTS: Eighteen patients with NPS, stages I to III, were treated with a CO(2) laser with or without a nasopharyngeal obturator with good results. CONCLUSION: The repair of NPS with a CO(2) laser and a nasopharyngeal obturator in severe cases helps in restoring nasopharyngeal patency. SIGNIFICANCE: This technique provided a reliable method of correcting postuvulopalatoplasty NPS. PMID- 11112960 TI - Value of flow cytometry with fine needle aspiration biopsy in patients with head and neck lymphoma. AB - Lymphomas frequently present in the head and neck clinically as painless adenopathy or as a swelling of an extranodal site, such as salivary gland or Waldeyer's ring. Most non-Hodgkin's lymphomas are B-cell neoplasms with distinctive clonal proteins that can be readily detected using flow cytometric analysis. In our experience, flow cytometric analysis has been a useful, convenient adjunct in the diagnosis of head and neck non-Hodgkin's lymphomas by fine needle aspiration biopsy. PMID- 11112961 TI - Ultrasound-guided fine-needle aspiration and thyroid disease. AB - BACKGROUND: Fine-needle aspiration represents a critical diagnostic test in determining proper management of thyroid disease and the use of ultrasound-guided fine-needle aspiration (USGFNA) has increased over the years. METHODS: A retrospective chart review of patients undergoing USGFNA. Two hundred fifteen patients underwent 234 procedures with 362 nodules aspirated within a 2 (1/2) year period. RESULTS: The mean ages of women and men were 51.9 and 57.8, respectively. The average size of nodules was 2.1 cm. A difficult to assess gland or nodule was the most common indication for USGFNA (33%). The sensitivity was 88.2%, specificity was 80.0%, the PPV was 65.2%, the negative predictive value was 94.1%, and the accuracy was 82.5%. The cancer yield, inadequacy, and complication rates were 44%, 10.5%, and 8.5%, respectively. CONCLUSIONS: USGFNA aspiration is a safe and effective diagnostic modality in the management of thyroid disease, especially for nodules that are difficult to palpate. PMID- 11112962 TI - Lymphatic malformation: predictive factors for recurrence. AB - OBJECTIVE: Although lymphatic malformations are often found to be well circumscribed when surgery is undertaken in early childhood, complete surgical excision can be difficult when the lesion is infiltrative. This study retrospectively evaluates these patients in an attempt to identify prognostic factors that may predict recurrence. STUDY DESIGN AND SETTING: A retrospective chart review was conducted covering the years 1991 to 1998. Seventeen patients were identified having undergone 32 surgical resections of tumors described as lymphatic malformations. Data abstracted from the charts included the site of the lesion, surgical and histologic assessment of encapsulation, and status at follow up examination. RESULTS: Six of 17 patients developed a recurrence after surgery. Correlation between recurrence and histologic or operative impressions of encapsulation was significant by chi(2) analysis (P<0.01). CONCLUSION: On the basis of the findings of this case series, lymphatic malformations that are found to be nonencapsulated and infiltrative by intraoperative or histologic assessment are more likely to recur. PMID- 11112963 TI - Osteocutaneous radial forearm free flap: its use without significant donor site morbidity. AB - While the fasciocutaneous radial forearm free flap has gained increasing popularity, the osteocutaneous radial forearm free flap has been condemned because of a high rate of pathologic donor radius fracture. On the basis of studies that demonstrated increased strength in ostectomized radii after dynamic compression plating, we believed that internal fixation at the time of graft harvest would significantly reduce the incidence of donor radius fracture. This is a retrospective review of the first 54 patients undergoing osteocutaneous radial forearm free flap reconstruction of the head and neck at our institution; 52 underwent prophylactic plating of their donor radii. No clinically significant donor radius fractures have occurred in plated patients. Five asymptomatic fractures were discovered on routine radiographs and required no treatment. Objective evaluation of forearm range of motion and strength after graft harvest demonstrated excellent function compared with unoperated arms. Serial radiographs have shown remodeling and reconstitution of donor radii without localized osteopenia. PMID- 11112964 TI - Bone-screw mandible fixation: an intraoperative alternative to arch bars. AB - OBJECTIVES: Bone-screw mandible fixation (BSMF) is evaluated as an alternative to intraoperative arch-bar maxillomandibular fixation before plating of mandibular fractures. BSMF is achieved by wire ligation of opposing bone-screws placed in the maxilla and mandible. METHODS: A retrospective evaluation of 23 patients with 40 mandibular fractures who underwent mandibular fracture repairs. BSMF was used instead of arch bars to ensure proper dental occlusion. All fractures were then plated, after which BSMF was removed before termination of anesthesia. RESULTS: Normal occlusion was observed in 21 patients (91.3%), Class II malocclusion was noted in 1 patient (4. 3%), and 1 patient was edentulous. No complications related to the use of BSMF were observed. CONCLUSION: BSMF can serve as a viable alternative to arch-bar maxillomandibular fixation for obtaining temporary intraoperative occlusion. BSMF produces acceptable malocclusion rates and offers the advantages of decreased intraoperative time, lower risk for percutaneous and mucosal wire punctures, and ease of use. PMID- 11112965 TI - Results of a humanitarian otologic and audiologic project performed outside of the United States: lessons learned from the "Oye, Amigos!" project. AB - From 1989 to 1993, "Oye, Amigos!" a combined group of hearing health and other medical professionals performed 18 humanitarian medical and audiologic trips to Tepic, Nayarit, Mexico. The group saw 1500 patients, issued over 800 hearing aids, and performed 150 surgeries on 123 patients. Our tympanoplasty success rate, defined as an intact tympanic membrane, was 41% during the first 2 years of the project but increased to 74% during the last 3 years. Two hundred eighteen patients who were candidates for surgery did not return for care. We present the lessons learned from the surgical care and overall management of this project, and present suggestions to improve future projects. PMID- 11112966 TI - Revision stapedectomy. AB - OBJECTIVE: To evaluate results of revision stapedectomy with and without use of the laser and determine factors predictive of hearing outcome. STUDY DESIGN AND SETTING: Retrospective review of 356 revision stapedectomy operations performed at the House Ear Clinic, a tertiary neurotologic private practice, between 1983 and 1995. RESULTS: A postoperative gap of < or =10 dB was obtained in 60% of cases. Results were similar with and without the use of a laser. Sensorineural hearing loss of >10 dB occurred in 7.7%, with 3 (1.4%) ears with profound hearing loss. A poorer outcome was related to incus necrosis, multiple revisions, and indications for surgery other than conductive hearing loss. CONCLUSION: Revision stapedectomy can provide good gap closure in 60% of cases, with small risk of sensorineural hearing loss. SIGNIFICANCE: Although not as satisfactory as primary stapedectomy, revision stapedectomy can be offered to patients with reasonable expectations for good gap closure. PMID- 11112967 TI - Complications associated with labyrinthine fistula in surgery for chronic otitis media. AB - OBJECTIVE: Labyrinthine fistula (LF) is encountered during surgery for chronic otitis media (COM) with an average frequency of 7.5%. This study was undertaken to investigate the prevalence of coexisting complications with LF in surgery for COM. STUDY DESIGN AND SETTING: A retrospective review of 111 consecutive cases of COM surgery performed by the same surgeon in a large metropolitan teaching public hospital in the United States serving the inner city population. RESULTS: There were 23 (21%) LFs in 111 patients. The majority of cases were revision surgery with an average of 2.6 operations per patient. Other complications were encountered in 18 (78%) of the 23 patients with an LF: 5 (22%) encephaloceles, 9 (39%) dehiscences of the tegmen, 12 (52%) patients with facial nerve involvement by cholesteatoma, and 2 (8.7%) patients with facial nerve paralysis. These patients required extensive surgery for correction of their concurrent complications. CONCLUSION: Compared with those cases in which a fistula was not found, the ratio of the above complications were as follows: encephalocele, 1:9; facial nerve involvement by cholesteatoma, 1:11.3; facial nerve paralysis, 1:3.8. SIGNIFICANCE: Patients with neglected COM have an incidence of an LF twice that reported in the literature. The prevalence of associated complications with LF requiring extensive surgery is significantly higher than that observed in noncomplicated COM cases. PMID- 11112968 TI - Cochlear implantation in patients with compromised healing. AB - BACKGROUND: The indications for cochlear implantation (CI) are continually evolving and, as experience accumulates, the relative contraindications for CI continue to decrease. However, there is little information regarding CI in patients who may be considered to be at risk for poor wound healing due to immunosuppression or intercurrent disease. OBJECTIVE: To assess and report the complication rates, postoperative course, postimplant rehabilitation, and long term performance of patients considered at risk due to presumably impaired healing capability. We hypothesized that these patients had outcomes similar to other implanted patients. METHODS: This is a retrospective chart review of 277 patients who have received CI at the University of Miami Ear Institute between 1990 and 1999. The clinical courses of 6 patients on immunosuppressive medications and 7 patients with diseases believed to be associated with poor healing are reported. RESULTS: Long-term follow-up (mean, 33 months) showed postoperative complication rates, performance, and rehabilitation compliance that were similar to published reports of noncompromised patients. CONCLUSION: CI of selected patients with potentially reduced healing capabilities is safe and effective. PMID- 11112969 TI - Endobronchial placement of a safety pin during intubation. PMID- 11112970 TI - Cavernous sinus thrombosis complicating odontogenic parapharyngeal space neck abscess: a case report and discussion. PMID- 11112971 TI - Unusual presentation of a rare nasal tumor. PMID- 11112972 TI - Primary melanoma of the sphenoid sinus. PMID- 11112973 TI - Nasal septal clamp. PMID- 11112974 TI - Otogenic brain abscess: review of 41 cases. AB - Forty-one patients in whom otogenic brain abscess was diagnosed and has been treated since 1968 are presented. Sixty-five percent of the patients were between 5 and 15 years of age. All patients had chronic otitis media, and 95% had cholesteatoma. Abscess was located in the temporal lobe in 54%, in the cerebellum in 44%, and in both locations in 2% of the cases. Most patients had radical mastoidectomy and evacuation of the abscess through the mastoidectomy (61%). In addition to mastoidectomy, burr hole drainage was used in 20% and craniotomy in 15%. The most common micro-organism involved was Proteus. Overall mortality in this series is 29%, but after 1976, when CT became available for the diagnosis and follow-up, the mortality rate was reduced to 10%. PMID- 11112975 TI - Development and application of a health-related quality-of-life instrument for adults with cochlear implants: the Nijmegen cochlear implant questionnaire. AB - OBJECTIVE: The goal was to develop a quantifiable, self-assessment health-related quality of life (QoL) instrument for use in cochlear implant (CI) users. DESIGN: Three principal domains were distinguished: physical, psychological, and social. Forty-five postlingually deaf adult multichannel CI users and 46 deaf candidates on the waiting list for CIs (control group) participated in the study. RESULTS: Retrospective scores for the CI group corresponded very well with the scores for the control group. Current QoL scores were substantially higher for all 6 subdomains. Internal consistency and testretest reliability coefficients proved to be satisfactory, and the ability to detect clinical changes with the Nijmegen Cochlear Implantation Questionnaire (NCIQ) proved to be good. CONCLUSIONS: The psychometric characteristics of the NCIQ proved to be reliable and probably valid and sensitive to clinical changes. The data obtained with the NCIQ reflected that the instrument was able to detect that a CI had significant effects on several health-related QoL aspects, including the social and psychological domains. PMID- 11112976 TI - Increased retinal image velocity after vestibular lesion. AB - The vestibulo-ocular reflex stabilizes gaze during head movements by producing compensatory eye movements. Retinal image velocity (RIV) is defined as the difference between the eye and head velocities. The RIV of 20 vestibular schwannoma (VS) patients and 17 healthy controls was measured with a head autorotation test. The head autorotation test had a sensitivity of 80% and a specificity of 88%. The mean RIV (degree/second) +/- 95% confidence intervals of the VS patients in the 5 frequency bands of 1 to 5 Hz was respectively 4.8 (4.2 to 5.5), 11.5 (8.6 to 14.4), 21.7 (15.5 to 27.9), 25.2 (17.1 to 33.4), and 26.1 (13.1 to 39.1). The RIV of the VS patients was asymmetrically larger on the operated side (P<0.05) in the frequency band of 1 Hz. The mean RIV was significantly (P<0.05) larger in the VS patients than in the controls in the frequency bands of 1 to 4 Hz. The vestibulo-ocular reflex is inaccurate after VS surgery; but the inaccuracy may not lead to the occurrence of any symptoms. PMID- 11112977 TI - Nuclear morphometry for the prediction of regional lymph nodes metastases in patients with cancer of the larynx. AB - The changes in cell nuclei reflect their activity. Quantitative morphometric analyses of tumor nuclei could be instrumental in providing prognostic information. We studied whether, and if so, which specific nuclear parameters and histoclinical factors in patients with cancer of the larynx could be related to the lymph node metastases. Specimens were taken from 61 patients surgically treated in the Department of Otolaryngology, Jagiellonian University, Cracow, Poland, between 1987 and 1988. The period between the onset of the first symptoms and the actual commencement of the treatment spanned no more than 9 months. The follow-up period was no shorter than 5 years. Histologically confirmed metastases in the regional lymph nodes were found in 16 patients. The histologic grading and tumor front grading was pursued in all cases. Fourteen parameters of the nuclei were studied with the aid of a computer-assisted system of image analysis. The morphometric parameters and the histoclinical factors were analyzed by the chi(2) test and the stepwise logistic regression. It was established that nuclear area > or =66 microm (P = 0.042), perimeter > or =32 microm (P = 0.087), optical density > or =22,500 (P = 0.027), long axis > or =10.15 microm (P = 0.025), short axis > or =7.3 microm (P = 0.003), TFG assessed (> or =15 points) and tumor advancement (T3, T4) were related to more frequent metastases to the lymph nodes. The morphometric parameters of the greatest significance were short axis and optical density. The quantitative morphometric analysis could prove a useful tool in predicting metastases to the lymph nodes. PMID- 11112978 TI - Improvement in bone conduction threshold after tympanoplasty. AB - OBJECTIVE: To investigate the causes of bone conduction threshold impairment associated with middle ear pathoses and the factors influencing improvement in bone conduction threshold after tympanoplasty. STUDY DESIGN AND SETTING: The records of 98 consecutive patients with unilateral chronic otitis media who underwent tympanoplasty were reviewed. Pre-operatively, 15 dB or more depression of bone conduction threshold at least in 2 frequencies between 500 and 6000 Hz was considered to be significant. Similarly in the postoperative period, 15 dB or more improvement of bone conduction threshold at least in 2 frequencies between 500 and 6000 Hz was regarded as significant. RESULTS: Twelve (12.5%) of 98 cases were found to have depressed bone conduction threshold; 6 of 12 cases had improved bone conduction threshold after tympanoplasty. CONCLUSION: In cases with cholesteatoma and extensive middle ear disease, successful results could be achieved after tympanoplasty disregarding the air-bone gap and deteriorated bone conduction threshold. SIGNIFICANCE: Bone conduction threshold may improve after tympanoplasty. PMID- 11112979 TI - Correlation between Ki-67 index and some clinical aspects of acoustic neuromas (vestibular schwannomas). AB - Evaluation of the proliferation activity of neuromas has a practical meaning when there are doubts about the complete resection of the tumor. Evaluation of the clinical aspects connected with increased proliferation activity may have a much broader application. The aim of this study was to correlate selected clinical and radiologic aspects of vestibular schwannomas with the results of the Ki-67 index. The studied group included 23 males and 20 females. Unilateral neuromas were stated in 38 cases (mean age, 52.2 years) and bilateral tumors in 5 cases (mean age, 44.2 years). The immunohistochemical tests (Ki-67) were performed on the specimens preserved in formalin and stored in paraffin. The Ki-67 index was estimated in a semiquantitative study. The mean value of Ki-67 index was 1.86%. In case of unilateral neuromas (n = 38), the average Ki-67 index was 1.74%. In 5 cases of bilateral tumors, the index amounted to 2.79% (P = 0.278). No significant correlation was found by comparing the value of the Ki-67 index with the age of patients (P = 0.410: r = 0.128). Significant differences in the value of the Ki-67 index were noted in the sub-groups of tumors that were evaluated radiologically as growing and stable. The mean value of Ki-67 index was 3.17% in the first subgroup; in stable neuromas, it was significantly lower, amounting to 1.11% (P = 0.020). Such results may confirm that the growth rate of vestibular schwannomas varies and may explain the difficulties in estimating the growth of neuromas on the basis of clinical aspects only. PMID- 11112980 TI - Head and neck manifestations of B-cell chronic lymphocytic leukemia. PMID- 11112982 TI - Reply PMID- 11112981 TI - Malignant neoplasms of the nasal cavity and paranasal sinuses. PMID- 11112983 TI - Imaging of dynamic changes of the actin cytoskeleton in microextensions of live NIH3T3 cells with a GFP fusion of the F-actin binding domain of moesin. AB - BACKGROUND: The cell surface undergoes continuous change during cell movement. This is characterized by transient protrusion and partial or complete retraction of microspikes, filopodia, and lamellipodia. This requires a dynamic actin cytoskeleton, moesin, components of Rho-mediated signal pathways, rearrangement of membrane constituents and the formation of focal adhesion sites. While the immunofluorescence distribution of endogenous moesin is that of a membrane-bound molecule with marked enhancement in some but not all microextensions, the C terminal fragment of moesin co-distributes with filamentous actin consistent with its actin-binding activity. By taking advantage of this property we studied the spontaneous protrusive activity of live NIH3T3 cells, expressing a fusion of GFP and the C-terminal domain of moesin. RESULTS: C-moesin-GFP localized to stress fibers and was enriched in actively protruding cellular regions such as filopodia or lamellipodia. This localization was reversibly affected by cytochalasin D. Multiple types of cytoskeletal rearrangements were observed that occurred independent of each other in adjacent regions of the cell surface. Assembly and disassembly of actin filaments occurred repeatedly within the same space and was correlated with either membrane protrusion and retraction, or no change in shape when microextensions were adherent. CONCLUSIONS: Shape alone provided an inadequate criterion for distinguishing between retraction fibers and advancing, retracting or stable filopodia. Fluorescence imaging of C-moesin-GFP, however, paralleled the rapid and dynamic changes of the actin cytoskeleton in microextensions. Regional regulatory control is implicated because opposite changes occurred in close proximity and presumably independent of each other. This new and sensitive tool should be useful for investigating mechanisms of localized actin dynamics in the cell cortex. PMID- 11112984 TI - Hepatic glucokinase promoter polymorphism is associated with hepatic insulin resistance in Asian Indians. AB - BACKGROUND: The role of glucokinase (GCK) in the pathogenesis of maturity-onset diabetes of the young is well established. However, its role in the common form of type 2 diabetes is far from convincing. We investigated the role of the G-to-A polymorphism in the hepatic GCK promoter on insulin sensitivity and beta cell function in 63 normotensive Asian Indians with normal glucose tolerance. As proposed by Matsuda and DeFronzo, hepatic insulin sensitivity (ISIH) and total body insulin sensitivity (ISIM) were estimated from the oral glucose tolerance test. Beta cell function was estimated using %B from the Homeostasis Model Assessment and insulingenic index (dI/dG). RESULT: We identified 38 GG, 24 GA, and one AA subjects. The AA subject was pooled with the GA subjects during the analysis. No difference was noted in the demographic features between the two genotypic groups (GG vs. GA/AA). Compared to the GG group, the GA/AA group had a lower ISIH (p=0.002), a lower ISIM (p=0.009), a higher %B (p=0.014), and a higher dI/dG (p=0.030). Multivariate analysis revealed that this polymorphism is an independent determinant for ISIH (p=0.019) and along with age, waist-hip ratio, gender, and diastolic blood pressure accounted for 51.5% of the variation of ISIH. However, this polymorphism was a weak, but independent determinant for ISIM (p=0.089) and %B (p=0.083). Furthermore, it had no independent effect on dI/dG (p=0.135). CONCLUSIONS: These data suggest that the G-to-A polymorphism in the hepatic GCK promoter is associated with hepatic insulin resistance in Asian Indians. PMID- 11112985 TI - In vivo labelling of functional ribosomes reveals spatial regulation during starvation in Podospora anserina. AB - BACKGROUND: To date, in eukaryotes, ribosomal protein expression is known to be regulated at the transcriptional and/or translational levels. But other forms of regulation may be possible. RESULTS: Here, we report the successful tagging of functional ribosomal particles with a S7-GFP chimaeric protein, making it possible to observe in vivo ribosome dynamics in the filamentous fungus Podospora anserina. Microscopic observations revealed a novel kind of ribosomal protein regulation during the passage between cell growth and stationary phases, with a transient accumulation of ribosomal proteins and/or ribosome subunits in the nucleus, possibly the nucleolus, being observed at the beginning of stationary phase. CONCLUSION: Nuclear sequestration can be another level of ribosomal protein regulation in eukaryotic cells. This may contribute to the regulation of cell growth and division. PMID- 11112986 TI - The immune response following myocardial infarction: a role for T-cell-mediated myocyte damage. PMID- 11112987 TI - Titin: the "missing link" of diastole. PMID- 11112988 TI - Atherosclerosis in AIDS: potential pathogenetic roles of antiretroviral therapy and HIV. PMID- 11112989 TI - Phospholamban and cardiac contractile function. PMID- 11112990 TI - Cytotoxic T lymphocytes are activated following myocardial infarction and can recognize and kill healthy myocytes in vitro. AB - The damage of myocardial infarction (MI) is often progressive. A possible mechanism for subsequent myocardial damage and heart failure after MI is immune response against cardiac self-antigens. The purpose of our study was to test the hypothesis that cytotoxic T lymphocytes are activated following acute MI and may have a role in producing further myocardial damage. Rats were allocated into three experimental groups: acute MI, Sham MI and non-operated control. One, two and three weeks after surgery, lymphocytes were obtained from rat spleens and incubated with neonatal cardiac myocytes. Lymphocyte proliferation was assessed by a thymidine incorporation assay and calculated as proliferation index (PI). Myocyte destruction was measured by a crystal-violet staining assay and expressed as percentage of cell destruction. Proliferation index was significantly higher among lymphocytes obtained from MI animals (44. 3+/-5.8 and 44.9+/-5.1, at 2 and 3 weeks after MI, respectively) than sham MI (29.3+/-5.3, 27.1+/-4.7) (P<0.05) or control animals (17.1+/-2.5, 16.2+/-2.8) (P=0.03). Cytotoxic activity of the MI lymphocytes against the cultured cardiomyocytes was significantly higher 2 and 3 weeks after MI, (36.4+/-7.3%, 69.3+/-4.9%) compared to sham MI (17.9+/-3.14%, 36.6+/-5.3%) (P<0.001) and control animals respectively (13.3+/-5.4%, 17.4+/ 6.1%) (P<0.001). The cytotoxic activity against healthy cardiomyocytes was myocyte-specific, induced by CD8 lymphocytes and major-histocompatibility complex (MHC) restricted. Cytotoxic T lymphocytes (CD8) are activated following MI and can recognize and kill normal cardiomyocytes in vitro. The newly described pathophysiological insights may provide novel oportunities to prevent death of non-ischemic cardiomyocytes and heart failure following myocardial infarction. PMID- 11112992 TI - Effect of interleukin-18 on viral myocarditis: enhancement of interferon- gamma and natural killer cell activity. AB - Encephalomyocarditis virus causes viral myocarditis with myocyte necrosis and inflammatory cell infiltration in mice. Because previous studies have shown that some cytokines prevent the sequelae of myocarditis, we assessed the effect of a newly identified cytokine, interleukin-18 (IL-18), in preventing the sequelae of myocarditis. Murine IL-18 (10 microg/day/mouse) was given peritoneally for 10 days in C3H mice infected with EMC virus. Mice were divided into group IL-18 (infected-treated), saline group (infected-untreated), group IL-18-2 (treatment started on day 2), group IL-18-5 (treatment started on day 5). Although the 14 day survival rate in saline group was 20%, that in the group IL-18 increased to 80% (P<0.05). Either mice in group IL-18-2 or in group IL-18-5 did not survive longer than saline group. The viral titer of the heart on day 3 was lower in group IL-18 compared to the saline group (1.00+/-0.20 log(10)tissue culture infected dose (TCID)(50)/mg wet weight v 1.42+/-0.12 log(10)TCID(50)/mg, n=3 of each). Mice in group IL-18 had less myocardial necrosis and cellular infiltration than the saline group. The myocardial expression of interferon- gamma (IFN- gamma) mRNA in group IL-18 was significantly (P<0.05) greater than the saline group on days 1 and 3 after viral inoculation. Circulating IFN- gamma was significantly elevated on days 1, 5, and 7, but significantly reduced on day 3. The natural killer cell activities in the spleen on days 1, 3, and 5 were significantly (P<0.05) greater in group IL-18 than in the saline group (41+/-9%v 10+/-6% on day 3, 4 of each). We conclude that IL-18 reduces the severity of EMC viral myocarditis by inducing cardiac expression of IFN- gamma mRNA and increasing splenic natural killer cell activity. PMID- 11112991 TI - Changes in titin and collagen underlie diastolic stiffness diversity of cardiac muscle. AB - Small (N2B) and large (N2BA) cardiac titin isoforms are differentially expressed in a species-specific and heart location-specific manner. To understand how differential expression of titin isoforms may influence passive stiffness of cardiac muscle we investigated the mechanical properties of mouse left ventricular (MLV) wall muscle (expressing predominantly the small titin isoform), bovine left atrial (BLA) wall muscle (predominantly the large isoform), and bovine left ventricular (BLV) wall muscle (expressing small and large isoforms at similar levels). Results indicate that the overall passive muscle stiffness of the muscle types varies nearly ten-fold, with stiffness increasing in the following order: BLA, BLV and MLV. To investigate the basis of the variation in the overall muscle stiffness, the contributions of titin and collagen to muscle stiffness were determined. Results showed that increased muscle stiffness results from increases in both titin- and collagen-based passive stiffness, indicating that titin and collagen change in a co-ordinated fashion. The expression level of the small titin isoform correlates with titin's contribution to overall muscle stiffness, suggesting that differential expression of titin isoforms is an effective means to modulate the filling behavior of the heart. PMID- 11112993 TI - Phosphorylation of phospholamban at threonine-17 in the absence and presence of beta-adrenergic stimulation in neonatal rat cardiomyocytes. AB - The site-specific phospholamban phosphorylation was studied with respect to the interplay of cAMP- and Ca(2+)signaling in neonatal rat cardiomyocytes. To elucidate the signal pathway(s) for the activation of Ca(2+)/calmodulin-dependent protein kinase (CaMKII) we studied Thr17 phosphorylation of phospholamban in dependence of Ca(2+)channel activation by S(-)-Bay K8644 and in dependence of the depletion of the sarcoplasmic reticulum Ca(2+)stores by ryanodine or thapsigargin in the absence or presence of beta -adrenergic stimulation. The isoproterenol (0.1 microM)-induced Thr17 phosphorylation was potentiated 2.5-fold in presence of 1 microM S(-)-Bay K8644. Interestingly, S(-)-Bay K8644 alone was also able to induce Thr17 phosphorylation in a dose- and time-dependent fashion. Ryanodine (1.0 microM) reduced both the isoproterenol (0.1 microM) and S(-)-Bay K8644-(1 microM) mediated Thr17 phosphorylation by about 90%. Thapsigargin (1 microM) diminished the S(-)-Bay K8644 and isoproterenol-associated Thr17 phosphorylation by 53.5+/-6.3% and 92. 5+/-11.1%, respectively. Ser16 phosphorylation was not affected under these conditions. KN-93 reduced the Thr17 phosphorylation by S(-) Bay K8644 and isoproterenol to levels of 1.1+/-0.3% and 8.6+/-2. 1%, respectively. However, the effect of KN-93 was attenuated (47. 8+/-3.6%) in isoproterenol prestimulated cells. Protein phosphatase inhibition by okadaic acid increased exclusively the Ser16 phosphorylation. In summary, our results reflect a cross-talk between beta -adrenoceptor stimulation and intracellular Ca(2+)at the level of CaMKII-mediated phospholamban phosphorylation in neonatal rat cardiomyocytes. We report conditions which exclusively produce Thr17 or Ser16 phosphorylation. We postulate that Ca(2+)transport systems of the sarcoplasmic reticulum are critical determinants for the activation of CaMKII that catalyzes phosphorylation of phospholamban. PMID- 11112994 TI - An immortalized myocyte cell line, HL-1, expresses a functional delta -opioid receptor. AB - The present study characterizes opioid receptors in an immortalized myocyte cell line, HL-1. Displacement of [(3)H]bremazocine by selective ligands for the mu (mu), delta (delta), and kappa (kappa) receptors revealed that only the delta selective ligands could fully displace specific [(3)H]bremazocine binding, indicating the presence of only the delta -receptor in these cells. Saturation binding studies with the delta -antagonist naltrindole afforded a B(max)of 32 fmols/mg protein and a K(D)value for [(3)H]naltrindole of 0.46 n M. The binding affinities of various delta ligands for the receptor in HL-1 cell membranes obtained from competition binding assays were similar to those obtained using membranes from a neuroblastomaxglioma cell line, NG108-15. Finally, various delta -agonists were found to stimulate the binding of [(35)S]GTP gamma S, confirming coupling of the cardiac delta -receptor to G-protein. DADLE (D-Ala-D-Leu enkephalin) was found to be the most efficacious in this assay, stimulating the binding of [(35)S]GTP gamma S to 27% above basal level. The above results indicate that the HL-1 cell line contains a functionally coupled delta -opioid receptor and therefore provides an in vitro model by which to study the direct effects of opioids on cardiac opioid receptors. PMID- 11112995 TI - Abciximab inhibits the migration and invasion potential of human coronary artery smooth muscle cells. AB - In the EPIC trial, high-risk patients received the integrin receptor antagonist abciximab v placebo during and for 12 h following percutaneous coronary intervention with a significant 23% decrease of repeat revascularisation at 6 months. However, EPILOG and CAPTURE trials could not confirm these promising long term results. Recently presented data from the EPISTENT trial suggested a beneficial effect of abciximab on restenosis in patients with diabetes. Based on these divergent results the aim of this study was to test whether alpha v beta 3 receptor blockade by abciximab could cause inhibition of human coronary smooth muscle cell (hcSMC) proliferation, migration, and invasion which represent crucial steps during restenosis development. In contrast to quiescent hcSMCs, proliferating cells were capable to migrate towards chemoattractive stimuli and even capable to invade through a basement membrane equivalent. Abciximab and LM609, an alpha v beta 3 specific inhibiting antibody, caused only a modest dose dependent inhibition of hcSMC proliferation. On the contrary, the chemotactic and invasive potential of hcSMCs was significantly inhibited by abciximab administration 24 h prior to and during migration. (IC(50)=33.0 microg/ml for chemotaxis and IC(50)=0.5 microg/ml for invasion). For LM609 similar results were obtained. Administration of the drugs just during migration without pretreatment inhibited migration equally but invasion to a lower extent (abciximab: IC(50)=32.6 microg/ml for chemotaxis and IC(50)=44.9 microg/ml for invasion; LM609 IC(50)=3.1 microg/ml for chemotaxis and IC(50)=2.0 microg/ml for invasion). The attachment to the extracellular matrix proteins collagen I, collagen IV, laminin and vitronectin was not influenced. Pretreatment for 24 h with abciximab or LM609 did not cause a downregulation of the alpha v beta 3-integrin receptor. The results of this study indicate that the alpha v beta 3 antagonist abciximab is a potent inhibitor of hcSMC migration and invasion which could explain the observed lower reintervention rate after PTCA and stent implantation. PMID- 11112997 TI - Thyroid hormone regulates slow skeletal troponin I gene inactivation in cardiac troponin I null mouse hearts. AB - Two main troponin I genes, cardiac (cTnI) and slow skeletal (ssTnI), are expressed in the mammalian heart under the control of a developmentally regulated program. ssTnI is expressed first in embryonic and fetal heart, and is then downregulated by an unknown mechanism after birth. Unlike other contractile protein genes, ssTnI is not re-expressed during hypertrophy or end-stage heart failure in rats and humans. In the present study, we also show that ssTnI re expression does not occur in hypertrophic mouse heart. To investigate ssTnI downregulation further, cTnI knockout mice were used to examine a possible role for thyroid hormone. Northern blot analysis of euthyroid animals showed a time dependent loss of ssTnI mRNA that was similar for wild-type, heterozygous and homozygous cTnI mutant mice. In cTnI null mice made hyperthyroid by l -thyroxine, the duration of ssTnI expression assessed by both mRNA and protein content was abbreviated compared with the euthyroid group. Hyperthyroid cTnI null mice also died significantly earlier than euthyroids (postnatal day 14 v day 18). In cTnI null mice made hypothyroid by addition of phenylthiouracil to the drinking water, ssTnI expression was prolonged and mice survived until day 20 or 21. Overall, the results indicate that inactivation of the ssTnI gene occurs even in the absence of cTnI mRNA and protein indicating that these are not critical signals for ssTnI down regulation in the heart. In contrast, thyroid hormone influences the time course of ssTnI expression and the life span of cTnI null mice probably through a transcriptional regulation of ssTnI in the heart. PMID- 11112996 TI - SSeCKS gene expression in vascular smooth muscle cells: regulation by angiotensin II and a potential role in the regulation of PAI-1 gene expression. AB - Rat aortic smooth muscle cells (RASM) express the src suppressed C-kinase substrate (SSeCKS), which is thought to be an integral regulatory component of cytoskeletal dynamics and G-protein coupled-receptor signaling modules. The specific sub-classes of growth factor receptors that regulate the genomic changes in SSeCKS expression in smooth muscle cells have not been characterized. In this study we identify SSeCKS as an angiotensin type 1 (AT(1)) receptor-dependent target gene in RASM cells treated with angiotensin II (Ang II). SSeCKS mRNA levels increase up to three-fold relative to the control within 3.5 h of Ang II treatment and are followed by a slight decrease of mRNA relative to the control levels after 24 h of stimulation. SSeCKS gene expression and plasminogen activator inhibitor-1 (PAI-1) gene expression correlate in RASM cells treated with Ang II. By co-transfecting plasmids bearing recombinant-SSeCKS and a PAI-1 promoter/luciferase reporter into Cos-1 cells, we show that alternative forms of recombinant-SSeCKS protein differentially influence PAI-1 promoter activity. These data indicate a biochemical linkage between SSeCKS activity and one or more of the cytoplasmic signaling pathways that are involved in the control of PAI-1 promoter activity. Finally, we show that the alternative forms of recombinant SSeCKS protein differentially influence cell-spreading when ectopically expressed in ras -transformed rat kidney (KNRK) fibroblasts. Taken together, our data suggest that SSeCKS interacts with intracellular signaling pathways that control cytoskeletal remodeling and extracellular matrix remodeling following Ang II stimulation of the RASM cell. PMID- 11112998 TI - Cytoprotective mechanism of heat shock protein 70 against hypoxia/reoxygenation injury. AB - The role of heat shock protein 70 (HSP70) in the cytoprotection against hypoxia/reoxygenation injury was examined. Adult rat cardiomyocytes were isolated, subjected to hyperthermia at 42 degrees C for 15 min (heat shock treatment), and then incubated at 37 degrees C for 3 to 24 h (HSP production process). Heat shock treatment increased HSP70 production (80-260% increase); the peak increase was seen after 9 h of HSP production process. Thereafter, the cells were subjected to 120-min hypoxia and 15-min reoxygenation. Heat shock treatment increased the survival of the cells subjected to hypoxia/reoxygenation (1.5-2.5 fold); the maximal cytoprotection was observed after 12 h of HSP production process. Heat shock treatment increased HSP70 content in the nucleus when cells were subjected to 12 h of HSP production process. To examine the role of HSP70 accumulation in the nuclear fraction, the activity of poly(ADP-ribose) synthetase (PARS), which functions in the nucleus and consumes high-energy phosphates excessively in the reoxygenated state, were measured in the cells with heat shock and 12 h of HSP production process. Heat shock treatment attenuated the hypoxia/reoxygenation-induced increase in the PARS activity (50% decrease). Treatment of the cells with 3-aminobenzamide, an inhibitor of PARS, exerted the effects similar to those of heat shock treatment. These results suggest that attenuation of the PARS activity in the nucleus may play an important role in the cytoprotective effect of HSP70 on hypoxia/reoxygenation injury. PMID- 11112999 TI - Angiotensin II type 1 receptor blockade abolishes specific K(ATP)channel gene expression in rats with myocardial ischemia. AB - The cardiac ATP-sensitive potassium (K(ATP)) channel is potentially composed of an inward rectifier potassium channel (Kir6.1 and/or Kir6.2) subunit and the cardiac type of sulfonylurea receptor (SUR2A). We reported that cardiac Kir6.1 mRNA and protein are specifically upregulated in the non-ischemic as well as the ischemic regions in rats with myocardial ischemia, suggesting that humoral and/or hemodynamic factors are responsible for this regulation. In the present study, pretreatment with TCV-116, an angiotensin (Ang) II type 1 receptor antagonist, completely inhibited the upregulation of Kir6.1 mRNA and protein expression in both regions of rat hearts subjected to 60 min of coronary artery occlusion followed by 24 h of reperfusion; whereas pretreatment with lisinopril, an Ang converting enzyme (ACE) inhibitor, partly inhibited this upregulation. Except for rats pretreated with TCV-116, Kir6.1 mRNA levels were positively correlated with those for brain natriuretic peptide (BNP), a molecular indicator of regional wall stress, in both the non-ischemic and the ischemic regions. Plasma Ang II levels were not elevated in rats with control myocardial ischemia compared with sham rats. Thus, the stress-related induction of cardiac Kir6.1 mRNA and protein expression under myocardial ischemia is inhibited by pretreatment with an AT1 antagonist, but also in part by an ACE inhibitor, suggesting that activation of local renin-angiotensin system may play a role. PMID- 11113000 TI - Differences in Ca(2+)-handling and sarcoplasmic reticulum Ca(2+)-content in isolated rat and rabbit myocardium. AB - We made novel measurements of the influence of rest intervals and stimulation frequency on twitch contractions and on sarcoplasmic reticulum (SR) Ca(2+) content (using rapid cooling contractures, RCCs) in isolated ventricular muscle strips from rat and rabbit hearts at a physiological temperature of 37 degrees C. In addition, the frequency-dependent relative contribution of SR Ca(2+)-uptake and Na(+)/Ca(2+)-exchange for cytosolic Ca(2+)-removal was assessed by paired RCCs. With increasing rest intervals (1-240 s) post-rest twitch force and RCC amplitude decreased monotonically in rabbit myocardium (after 240 s by 45+/-10% and 61+/-11%, respectively P<0. 05, n=14). In contrast, rat myocardium (n=11) exhibited a parallel increase in post-rest twitch force (by 67+/-16% at 240 s P<0.05) and RCC amplitude (by 20+/-14%P<0.05). In rabbit myocardium (n=11), increasing stimulation frequency from 0.25 to 3 Hz increased twitch force by 295+/-50% (P<0.05) and RCC amplitude by 305+/-80% (P<0.05). In contrast, in rat myocardium (n=6), twitch force declined by 43+/-7% (P<0.05), while RCC amplitude decreased only insignificantly (by 16+/-7%). The SR Ca(2+)-uptake relative to Na(+)/Ca(2+)-exchange (based on paired RCCs) increased progressively with frequency in rabbit, but not in rat myocardium (;66+/-2% at all frequencies). We conclude that increased SR Ca(2+)-load contributes to the positive force frequency relationship in rabbits and post-rest potentiation of twitch force in rats. Decreased SR Ca(2+)-load contributes to post-rest decay of twitch force in rabbits, but may play only a minor role in the negative force-frequency relationship in rats. SR Ca(2+)-release channel refractoriness may contribute importantly to the negative force-frequency relationship in rat and recovery from refractoriness may contribute to post-rest potentiation. PMID- 11113001 TI - Modulation of myocardial SOD and iNOS during ischemia-reperfusion by antisense directed at ACE mRNA. AB - Renin-angiotensin system (RAS) is involved in the regulation of superoxide dismutase (SOD) and nitric oxide (NO) equilibrium, and its modulation protects hearts from ischemic dysfunction. We examined the effect of a new antisense oligodeoxynucleotides (AS-ODNs) directed at ACE mRNA on SOD and iNOS expression during myocardial ischemia. Sprague-Dawley rats were treated with saline, AS ODNs, or inverted-ODNs (IN-ODNs), given with liposome DOTAP/DOPE. Hearts were excised and subjected to 25 min of ischemia followed by 30 min of reperfusion. Ischemia-reperfusion in saline-treated hearts resulted in a decrease in the expression of SOD and an increase in the expression of inducible NOS (iNOS) genes concurrently with myocardial dysfunction. AS-ODNs, but not IN-ODNs, protected hearts against functional deterioration, and upregulated SOD expression and inhibited the expression of iNOS. ACE protein expression was decreased in the rat hearts of the AS-ODNs-treated group, but not in the IN-ODNs group. Thus manipulation of RAS with AS-ODNs directed at ACE mRNA can ameliorate cardiac dysfunction and modulate expression of SOD and iNOS at genomic level. PMID- 11113002 TI - Protection against murine coxsackievirus B3 myocarditis by T cell vaccination. AB - T cell vaccination regulates autoimmunity by the modification of helper and suppressor T cells. The present study was performed to examine whether T cell vaccination can prevent viral myocarditis in vivo. We used coxsackievirus B3 myocarditis in mice as an animal model with the analysis of lymphokine-activated killer cell activity. Vaccination of the mice with T lymphocytes significantly prolonged survival and improved cardiac histology of murine myocarditis. The effects of T cell vaccination were most evident when T cells sensitized with the same virus were used. Vaccination of the mice with T cells from other strains of mice showed lesser protective effects. Clearance of myocardial virus was not affected by this treatment. The efficacy of T cell vaccination was confirmed in vitro by the decrease of the lymphokine-activated killer cell activity against EL 4 tumor cells and cultured myocytes. T cell vaccination of mice prolonged survival and improved myocardial lesions of animals inoculated with coxsackievirus B3. PMID- 11113003 TI - No confirmation for a causal role of volume-regulated chloride channels in ischemic preconditioning in rabbits. AB - Volume-regulated chloride channels have recently been proposed to be end effectors in ischemic preconditioning. The present study attempted to confirm this hypothesis by looking both at cardioprotection and channel activity. In isolated rabbit cardiomyocytes, hypo-osmotic stress (167 mosm/l) induced a current with a magnitude of 2-5 pA/pF at 60 mV. That current could be blocked by the selective chloride channel blockers 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) or indanyloxyacetic acid 94 (IAA-94), but only at 100 microM and 1 m M respectively. Lower concentrations were not effective. Because the channel blocking concentrations were toxic in isolated perfused rabbit hearts, as evidenced by cessation of cardiac contraction and massive infarction, neither agent could be tested against preconditioning's anti-infarct effect. NPPB and IAA 94 at 1 microM and 10 microM, respectively (the doses used in a previous report), did not affect coronary flow, heart rate and developed pressure, and also did not prevent the infarct size reduction of ischemic preconditioning with 5 min global ischemia/10 min reperfusion preceding 30 min of regional ischemia and 120 min of reperfusion [11. 4(+/-3.6) and (11.1(+/-3.7)% infarction of risk area, respectively]. The volume-regulated chloride and organic osmolyte channel blocker 4, 4;-diisothiocyanostilbene-2,2;-disulfonic acid (DIDS) at 100 microM blocked the hypo-osmotically induced current in myocytes, but again could not be used, since it induced total cessation of cardiac contraction and reduced infarct size in non-preconditioned hearts. Our data do not confirm a prior study on a causal role for volume-regulated chloride channels in the protection of ischemic preconditioning. This hypothesis remains to be adequately tested. PMID- 11113004 TI - Metabolic adaptation of the hypertrophied heart: role of the malate/aspartate and alpha-glycerophosphate shuttles. AB - Activation of the malate/aspartate and alpha -glycerophosphate shuttles (the NADH shuttles) has been identified in glycolytically active newborn myocardium. The goal of this study was to determine if the NADH shuttles and their regulatory genes are activated in hypertrophied myocardium as substrate utilization shifts away from fatty acids and toward glucose and lactate. Capacity of the shuttles was determined in cardiac mitochondria isolated one week, one month, and three months following aortic banding or sham operation. Myocardial steady-state mRNA and protein levels of regulatory enzymes were also measured. Despite a significant increase in left ventricular mass and activation of the atrial natriuretic peptide gene, no change in malate/aspartate nor alpha glycerophosphate shuttle capacity was found at any of the three time points studied. Reactivation of the genes encoding the regulatory inner mitochondrial membrane proteins was not found in the hypertrophied myocardium, though these genes were down regulated one week following aortic-banding. These results suggest that sufficient malate/aspartate and alpha -glycerophosphate shuttle capacity exists in cardiac mitochondria to accommodate increased shuttle flux as hypertrophied myocardium becomes more glycolytically active. PMID- 11113005 TI - Impaired nitric oxide modulation of myocardial oxygen consumption in genetically cardiomyopathic hamsters. AB - We investigated the role of kinin and nitric oxide (NO) in the modulation of cardiac O(2)consumption in Syrian hamsters with overt heart failure (HF) and age matched normal hamsters. Using echocardiography, the hamsters with heart failure had reduced ejection fraction [31(+/-8) v 76(+/-5)%] and LV dilation [4.9(+/-0. 2) v 5.7(+/-0.3) mm, both P<0.05 from normal]. O(2)consumption in the left ventricular free wall was measured using a Clark-type O(2)electrode in an air tight chamber, containing Krebs solution buffered with Hepes (37 degrees C, pH 7.4). Concentration response curves to bradykinin (BK), ramiprilat (RAM), amlodipine (AMLO) and the NO donor, S -nitroso- N -acetyl-penicillamine (SNAP) were performed. Basal myocardial O(2)consumption was lower in the HF group compared to normal [316(+/-21) v 404(+/-36) nmol O(2)/min/g, respectively, P<0.05]. In the hearts from normal hamsters BK (10(-4)mol/l), RAM (10(-4)mol/l), and AMLO (10(-5)mol/l) all significantly reduced myocardial O(2)consumption by 42(+/-6)%, 29(+/-7)% and 27(+/-5)% respectively. This reduction was attenuated in the presence of N -nitro- l -arginine methyl ester (l -NAME) [BK: 3.3(+/-1.5)%, RAM: 3.3(+/-1.2)%, AMLO: 2.3(+/-1.2)%, P<0.05]. Interestingly in the hearts from HF group, BK, RAM and AMLO caused a significantly smaller reduction in myocardial O(2)consumption [10(+/-2)%, 2.5(+/-1.3)%, 6.3(+/-2.3)%, P<0.05]. In contrast, the NO donor SNAP reduced myocardial O(2)consumption in both groups and all those responses were not affected by l -NAME. These data indicate that endogenous NO production through the kinin-dependent mechanism is impaired at end-stage heart failure. The loss of kinin and NO control of mitochondrial respiration may contribute to the pathogenesis of heart failure. PMID- 11113006 TI - Malignant hypertrophic cardiomyopathy caused by the Arg723Gly mutation in beta myosin heavy chain gene. AB - Mutations causing hypertrophic cardiomyopathy have been described in nine genes encoding sarcomeric proteins. We report a new mutation in three families, with a C-->G transversion in nucleotide 12 307 of the beta-myosin heavy chain gene, located at the essential light chain interacting region, resulting in the replacement of arginine by glycine at amino acid residue 723. PCR amplification of the selected regions followed by single strand conformation polymorphism analysis, DNA sequencing of the polymorphic patterns and restriction analysis were used to detect the mutation. A total of 23 individuals were diagnosed as carriers, and seven were obligate carriers or had been clinically diagnosed. The Arg723Gly mutation was associated with a malignant phenotype. Ten out of 30 affected members died suddenly or needed an implantable cardioverter defibrillator at a mean age of 42, and seven members developed progressive heart failure, leading to death or heart transplant in five, at a mean age of 50 years. Echocardiography showed non-obstructive left ventricular hypertrophy in affected members older than 20 (sensitivity 68%). Mean survival of affected members was 51 years. In conclusion, a new mutation Arg723Gly in beta-myosin heavy chain gene is reported which shortens life expectancy because of sudden death and end-stage heart failure. PMID- 11113007 TI - The cardiac beta-adrenoceptor-G-protein(s)-adenylyl cyclase system in monocrotaline-treated rats. AB - In rats, injection of the alkaloid monocrotaline (MCT) causes right ventricular hypertrophy and cardiac failure. In order to study whether, in MCT-treated rats, changes in the cardiac beta -adrenoceptor-G-protein(s)-adenylyl cyclase system might be comparable to those found in human primary pulmonary hypertension, we assessed in right and left ventricles from MCT-treated rats the components of the beta -adrenoceptor system: the receptor number and subtype distribution (by (-) [(125)I]iodocyanopindolol binding), the G-proteins (by quantitative Western blotting), and the activity of adenylyl cyclase. A single injection of 60 mg/kg i.p. MCT caused in rats right ventricular hypertrophy (RVH); part of the rats developed cardiac failure (RVF). In these rats the cardiac beta -adrenoceptor-G protein(s)-adenylyl cyclase system was markedly changed beta -adrenoceptors were desensitized due to a decrease in receptor number, an uncoupling of the receptor from the G(s)-adenylyl cyclase system, a decrease in G(s)and a decrease in the activity of the catalytic unit of adenylyl cyclase. In general, these changes were more pronounced in right ventricles v left ventricles, and in rats with RVF v rats with RVH. On the other hand, cardiac muscarinic receptors and G(i)appeared not to be altered. We conclude that in MCT-treated rats changes in the cardiac beta -adrenoceptor-G-protein(s)-adenylyl cyclase system occur that resemble those observed in human primary pulmonary hypertension. Thus, MCT-treated rat appears to be a suitable animal model to study in more detail the pathophysiology of the development of right heart failure, and to identify new therapeutic possibilities. PMID- 11113009 TI - Limitation of infarct size in rabbit hearts by the novel adenosine receptor agonist AMP 579 administered at reperfusion. AB - The novel A(1)/A(2)adenosine receptor agonist AMP 579 has been reported to reduce myocardial infarct size in pig and dog. The present study tested the effect of AMP 579 in two rabbit models. In open-chest rabbits undergoing 30 min of regional ischemia and 3 h of reperfusion AMP 579 (3 microg/min/kg) reduced infarct size when treatment was started either 10 min before ischemia or 10 min prior to reperfusion from 36.4+/-3.1% of the risk zone in untreated hearts to 11.8+/-4.4 and 12.3+/-1.0%, respectively. To determine whether protection observed when the drug was administered shortly before reperfusion represented a long-lasting effect rather than merely a transient delay of necrosis, the chest wound was closed in layers and the rabbits permitted to recover. After 3 days the hearts were removed to evaluate infarct size. Continued limitation of infarct size after 3 days of reperfusion (8.2+/-2.8% of the risk zone) confirmed that sustained tissue salvage had been conferred by the drug. In isolated, buffer-perfused rabbit hearts undergoing 30 min of regional ischemia and 2 h of reperfusion, AMP 579 again limited infarct size (8.6+/-2.9% of the risk zone) when treatment started 10 min prior to reperfusion, arguing against an anti-leukocyte mechanism of protection. AMP 579's protective effect in this in vitro model was abrogated by 8-(p-sulfophenyl)theophylline, indicating that it was mediated through adenosine receptors. We conclude that AMP 579 given just prior to reperfusion may be an effective anti-infarct intervention. PMID- 11113008 TI - Retention in the endoplasmic reticulum as a mechanism of dominant-negative current suppression in human long QT syndrome. AB - Mutations in the cardiac potassium channel HERG (KCNH2) cause chromosome 7-linked long QT syndrome (LQT2) characterized by a prolonged QT interval, recurrent syncope and sudden cardiac death. Most mutations in HERG exhibit "loss of function" phenotypes with defective channels either inserted into the plasma membrane or retained in the endoplasmic reticulum. "Loss of function" mutations reduce I(Kr), the cardiac delayed rectifier current encoded by HERG, due to haploinsufficiency or suppression of wild-type function by a dominant-negative mechanism. One explanation for dominant-negative current suppression is that mutant subunits render tetrameric channel complexes non-conducting on co assembly. In the present paper we describe an alternative mechanism for this phenomenon. We show (1) that the dominant-negative HERG mutation A561V is retained in the endoplasmic reticulum and (2) that wild-type channels are tagged for retention in the ER by co-assembly with trafficking deficient A561V subunits. Thus, in HERG A561V dominant-negative suppression of wild-type function is the result of an acquired trafficking defect. PMID- 11113010 TI - Regional expression of protein phosphatase type 1 and 2A catalytic subunit isoforms in the human heart. AB - In mammalian species, including man, the duration of myocardial contraction is shorter in atria than ventricles. Total contraction time depends at least in part on phosphorylation and dephosphorylation of cardiac regulatory proteins. Dephosphorylation reactions are mediated by protein phosphatases. In the mammalian heart more than 90% of the protein phosphatase (PP) activity consists of PP1 and PP2A. Therefore, the aim of this study was to investigate which isoforms of PP1 and PP2A are present in human myocardium and whether their expression is regionally different. RT-PCR and Northern blotting revealed that all isoforms of PP1 and PP2A presently known are expressed in the human heart. Expression levels of PP1 alpha, delta, and gamma as well as 2A alpha were higher in right ventricles than in right atria. However, there was no such difference for PP2A beta. At the protein level PP1 alpha was unchanged, whereas PP2A was by 56% higher in right ventricles compared to atria. The phosphorylation state of TnI was lower in right ventricle than in right atrium. Thus, lower protein expression of PP2A in atrium could contribute to the faster relaxation by increasing the phosphorylation state of TnI. We conclude that expression of PP1 and PP2A isoforms is regionally regulated in the human heart. PMID- 11113011 TI - Abnormal mitochondrial respiration in failed human myocardium. AB - Chronic heart failure (HF) is associated with morphologic abnormalities of cardiac mitochondria including hyperplasia, reduced organelle size and compromised structural integrity. In this study, we examined whether functional abnormalities of mitochondrial respiration are also present in myocardium of patients with advanced HF. Mitochondrial respiration was examined using a Clark electrode in an oxygraph cell containing saponin-skinned muscle bundles obtained from myocardium of failed explanted human hearts due to ischemic (ICM, n=9) or idiopathic dilated (IDC, n=9) cardiomyopathy. Myocardial specimens from five normal donor hearts served as controls (CON). Basal respiratory rate, respiratory rate after addition of the substrates glutamate and malate (V(SUB)), state 3 respiration (after addition of ADP, V(ADP)) and respiration after the addition of atractyloside (V(AT)) were measured in scar-free muscle bundles obtained from the subendocardial (ENDO) and subepicardial (EPI) thirds of the left ventricular (LV) free wall, interventricular septum and right ventricular (RV) free wall. There were no differences in basal and substrate-supported respiration between CON and HF regardless of etiology. V(ADP)was significantly depressed both in ICM and IDC compared to CON in all the regions studied. The respiratory control ratio, V(ADP)/V(AT), was also significantly decreased in HF compared to CON. In both ICM and IDC, V(ADP)was significantly lower in ENDO compared to EPI. The results indicate that mitochondrial respiration is abnormal in the failing human heart. The findings support the concept of low myocardial energy production in HF via oxidative phosphorylation, an abnormality with a potentially impact on global cardiac performance. PMID- 11113012 TI - Variants of trophic factors and expression of cardiac hypertrophy in patients with hypertrophic cardiomyopathy. AB - Patients with hypertrophic cardiomyopathy (HCM) exhibit variable expression of left ventricular hypertrophy (LVH), a major determinant of mortality and morbidity, which is partly due to the diversity of causal mutations, genetic background (modifier genes), and probably environmental factors. We determined association of functional variants of tumor necrosis factor (TNF)- alpha, interleukin-6 (IL6), insulin-like growth factor-2 (IGF2), transforming growth factor- beta 1 (TGFB1), and aldosterone synthase (CYP11B2) genes, all previously implicated in cardiac hypertrophy, with the severity of LVH in patients with HCM. Two-dimensional echocardiography was performed and demographic variables were recorded in 142 genetically independent patients. Indices of LVH including interventricular septal thickness (IVST), left ventricular mass index (LVMI), and LVH score were measured/calculated. TNF-alpha-308G/A, IL6-174G/C, IGF2 820G/A, TGFB1-509C/T, and CYP11B2-344T/C genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Genotypes were identified by the presence of specific electrophoretic patterns and their distributions were according to the Hardy-Weinberg equilibrium. Demographic variables were not significantly different among the genotypes. Subjects with the AA genotype of TNF-alpha (n=8) were approximately 13 years younger at the time of clinical diagnosis. Despite a younger age, they had a greater mean LVMI than those with the GG (n=94) or GA (n=33) genotypes (191.8+/-59.5 v 139.1+/-47.3 v 132.1+/-34.3, respectively, P=0.004). TNF-alpha-308G/A genotypes accounted for 6.0% of variability of LVMI (P=0.002). Mean IVST, LVEDD, and LVH score were not significantly different. Variants of IL6, IGF2, TGFB1, and CYP11B2 were not associated with indices of LVH. The uncommon allele of TNF-alpha-308G/A polymorphism, known to produce more TNF- alpha, was associated with greater LVMI and clinical diagnosis at a younger age in patients with HCM. Functional variants of other trophic factors, previously implicated in cardiac hypertrophy, were not associated with the indices of LVH. These results suggest that TNF-alpha is a modifier gene for HCM. PMID- 11113013 TI - Calreticulin and calsequestrin are differentially distributed in canine heart. AB - Cardiac sarcoplasmic reticulum (SR) consists of three continuous yet distinct regions: longitudinal, corbular, and junctional. Ca(2+)uptake, catalyzed by the Ca(2+)-dependent ATPase, is thought to occur throughout the SR whereas Ca(2+)release occurs in the region of the junctional SR. In the SR, Ca(2+)is stored in a complex with Ca(2+)-binding proteins such as calsequestrin. In the present study, the distribution of the high-affinity calcium-binding protein, calreticulin, in canine cardiac SR was determined and compared with the distribution of other SR proteins. Three types of distribution were observed. Many proteins, including the Ca(2+)-ATPase and two mannose-containing glycoproteins (52 and 131 kDa), were present in all subfractions. These proteins were absent or depleted from the sarcolemma-enriched fraction. The ryanodine receptor/Ca(2+)release channel and calsequestrin were localized to the junctional SR. Calreticulin immunoreactivity was detected in fractions containing longitudinal SR but not in fractions enriched in sarcolemma or junctional SR. Hence calreticulin is present in a unique vesicular fraction and the Ca(2+) binding proteins calreticulin and calsequestrin display different patterns of distribution in canine heart. PMID- 11113014 TI - Downregulation and nuclear relocation of MLP during the progression of right ventricular hypertrophy induced by chronic pressure overload. AB - The cardiac LIM domain protein MLP plays a crucial role in the architecture and mechanical function of cardiac myocytes. Mice lacking the MLP gene develop cardiac hypertrophy, dilated cardiopathy and heart failure. We investigated whether downregulation of MLP is induced by pressure overload and contributes to the physiopathology of cardiac hypertrophy and failure. We studied this mechanism in rat right ventricles submitted to pulmonary arterial hypertension, because it is known that this ventricle is very vulnerable to the deleterious effects of pressure overload. During the progression of cardiac hypertrophy to failure over a 31 days period there was a dramatic decrease by 50% of the MLP transcripts level. Consistently, immunohistochemistry detected very weak protein signals in the cytoplasms of cardiomyocytes at the failing stage, but myocytes nuclei were heavily labeled. The nuclear relocation was confirmed by the immunodetection of MLP on the nuclear and cytosolic fractions. This nuclear localization is the hallmark of a retro-differentiated phenotype, since it has been observed only in differentiating myoblasts. These changes were associated with ultrastructural disorganization of the myofibrils similar to that observed in MLP -/- mice. Therefore, MLP dowregulation occurring during gene reprogramming may critically contribute to mechanical failure of the myocardium. PMID- 11113016 TI - Introduction: antipyretic therapy PMID- 11113015 TI - Intracoronary, adenovirus-mediated Akt gene transfer in heart limits infarct size following ischemia-reperfusion injury in vivo. AB - BACKGROUND: Previous data have shown that enhanced Akt signaling inhibits cardiac myocyte apoptosis in vitro and in vivo. To elucidate the contribution of apoptosis to the pathogenesis of the infarct, we investigated whether intra coronary Akt gene delivery could reduce gross infarct size following ischemia/reperfusion injury. METHODS AND RESULTS: Replication-defective adenoviral constructs encoding a myristoylated, constitutively-active form of Akt (myrAkt) or beta -galactosidase were delivered to rat hearts by intracoronary perfusion. Twenty-four h after gene transduction, hearts in both groups underwent 45 min of ischemia followed by 4 h of reperfusion. A third group of animals also underwent ischemia-reperfusion injury but were not transduced with an adenoviral vector. The proportion of the left ventricle at risk was not different among the experimental groups. However, infarct size as a proportion of the area at risk was significantly lower in myrAkt-treated group than in the beta -galactosidase treated group or in the control group that was not subject to intracoronary perfusion (myrAkt=20.9+/-2.7%v beta -galactosidase=56.1+/-3.9% and control=46.2+/ 4.6%, P<0.05), as was infarct size as a proportion of the total left ventricle (myrAkt=11.4+/-3.2 v beta -galactosidase=32. 9+/-3.3 and control=23.5+/-3.0, P<0.05). CONCLUSIONS: These data demonstrate that Akt signaling limits infarct size following ischemia/reperfusion injury and they indicate that the activation of this pathway may be useful in protecting against myocardial loss in the diseased heart. PMID- 11113017 TI - Brief history of antipyretic therapy. AB - Both external cooling and pharmacotherapy have been used to treat fever since time immemorial. In the past century such treatments have proliferated at an astonishing rate. The COX-2 inhibitors are the most recent additions to the antipyretic pharmacopoeia. Additional research is needed to determine whether they represent an important new chapter in antipyretic therapy's long history or, for that matter, if the benefits of any currently available treatment for fever outweigh its cost. PMID- 11113018 TI - Role of the preoptic-anterior hypothalamus in thermoregulation and fever. AB - Lesion and thermal stimulation studies suggest that temperature regulation is controlled by a hierarchy of neural structures. Effector areas for specific thermoregulatory responses are located throughout the brain stem and spinal cord. The preoptic region, in and near the rostral hypothalamus, acts as a coordinating center and strongly influences each of the lower effector areas. The preoptic area contains neurons that are sensitive to subtle changes in hypothalamic or core temperature. Preoptic thermosensitive neurons also receive a wealth of somatosensory input from skin and spinal thermoreceptors. In this way, preoptic neurons compare and integrate central and peripheral thermal information. As a result of this sensory integration and its control over lower effector areas, the preoptic region elicits the thermoregulatory responses that are the most appropriate for both internal and environmental thermal conditions. Thermosensitive preoptic neurons are also affected by endogenous substances, such as pyrogens. By reducing the activity of warm-sensitive neurons and increasing the activity of cold-sensitive neurons, pyrogens cause fever, a state in which all thermoregulatory responses have elevated set-point temperatures. PMID- 11113019 TI - Neural route of pyrogen signaling to the brain. AB - In the pathogenesis of systemic inflammation and fever, peripheral inflammatory and pyrogenic signals gain access to the brain via humoral and neural routes. One of the neural routes is represented by chemosensitive afferent fibers of the abdominal vagus. We summarize our recent studies of the role of the abdominal vagus in fever. We conclude that capsaicin-sensitive fibers traveling within the hepatic vagal branch constitute a necessary component of the afferent mechanism of the febrile response to low, but not high, doses of circulating pyrogens. We speculate that this mechanism is triggered by blood-borne prostaglandins of the E series. PMID- 11113020 TI - Pyrogen sensing and signaling: old views and new concepts. AB - Fever is thought to be caused by endogenous pyrogenic cytokines, which are elaborated and released into the circulation by systemic mononuclear phagocytes that are activated by exogenous inflammatory agents and transported to the preoptic-anterior hypothalamic area (POA) of the brain, where they act. Prostaglandin (PG) E2 is thought to be an essential, proximal mediator in the POA, and induced by these cytokines. It seems unlikely, however, that these factors could directly account for early production of PGE2 following the intravenous administration of bacterial endotoxic lipopolysaccharides (LPS), because PGE2 is generated before the cytokines that induce it are detectable in the blood and the before cyclooxygenase-2, the synthase that they stimulate, is expressed. Hence other, more quickly evoked mediators are presumed to be involved in initiating the febrile response; moreover, their message may be conveyed to the brain by a neural rather than a humoral pathway. This article reviews current conceptions of pyrogen signalling from the periphery to the brain and presents new, developing hypotheses about the mechanism by which LPS initiates fever. PMID- 11113021 TI - Circulating cytokines as mediators of fever. AB - The febrile response is thought to be mediated by endogenous mediators, generically called "endogenous pyrogens." In the classical model of pathogenesis, induction of fever is mediated by the release of pyrogenic cytokines such as tumor necrosis factor (TNF), interleukin (IL)-1, IL-6, and interferons into the bloodstream in response to exogenous pyrogens. These mediators act at the level of the organum vasculosum of the lamina terminalis in the central nervous system (CNS), inducing synthesis of prostaglandins, which are the central mediators of the coordinated responses leading to fever. However, analysis of recent data suggests that multiple pathways may be involved in the induction of fever by cytokines, such as local cytokine production leading to signaling through vagal fibers, release of cytokine-induced circulating mediators at the tissue level, the use of membrane-bound cytokines as mediators, or the local release of cytokines in the hypothalamus by circulating activated monocytes. In addition, certain bacterial products can stimulate cytokine production directly at the level of hypothalamus, probably by activation of Toll-like receptors. A multipathway mechanism for the induction of fever is therefore suggested. PMID- 11113022 TI - Physiological rationale for suppression of fever. AB - Two critical assumptions are made when prescribing antipyretic therapy. One is that fever is, at least in part, noxious, and the other is that suppression of fever will reduce, if not eliminate, the noxious effects of fever. At present, neither assumption has been validated experimentally. PMID- 11113023 TI - Endogenous antipyretics. AB - Fever is the hallmark of the stereotyped host response to microbial infection, although it is just one of a number of high-risk strategies employed by the infected host to clear itself of invading pathogens. The febrile response is accompanied by activation of multiple endogenous antipyretic systems that serve to suppress its magnitude or duration. These include neuroactive substances of neural and humoral origin, some of which (e.g., glucocorticoids, melanocortins, and IL-10) have broad-ranging anti-inflammatory actions. Glucocorticoids, vasopressin, and melanocortins appear to exert their antipyretic effects by acting on receptors within the brain, but beyond this the mechanisms involved are unknown. It is hypothesized, but not proven, that endogenous antipyretic systems protect the host against the destructive consequences of unchecked fever. Importantly, pharmacological blockade of the actions of endogenous antipyretic systems increases fevers of even low to moderate intensity. Therefore, in addition to protecting against catastrophic consequences of high fever, endogenous antipyretic systems seem to play a fundamental physiological role in determining the normal course of fever. Elucidating the neural and biochemical mechanisms involved in suppression of fever by physiological antipyretic systems will yield a rich benefit, both by advancing the basic understanding of host defense strategies, and by permitting the design of novel antipyretic and anti inflammatory strategies for therapeutic intervention in human disease. PMID- 11113024 TI - Mechanism of action of acetaminophen: is there a cyclooxygenase 3? AB - Acetaminophen, also known as paracetamol, is a nonsteroidal anti-inflammatory drug with potent antipyretic and analgesic actions but with very weak anti inflammatory activity. When administered to humans, it reduces levels of prostaglandin metabolites in urine but does not reduce synthesis of prostaglandins by blood platelets or by the stomach mucosa. Because acetaminophen is a weak inhibitor in vitro of both cyclooxygenase (COX)-1 and COX-2, the possibility exists that it inhibits a so far unidentified form of COX, perhaps COX-3. In animal studies, COX enzymes in homogenates of different tissues vary in sensitivity to the inhibitory action of acetaminophen. This may be evidence that there are >2 isoforms of the enzyme. Recently, a variant of COX-2 induced with high concentrations of nonsteroidal anti-inflammatory drugs was shown to be highly sensitive to inhibition by acetaminophen. Therefore COX-3 may be a product of the same gene that encodes COX-2, but have different molecular characteristics. PMID- 11113025 TI - Nonsteroidal anti-inflammatory drugs, acetaminophen, cyclooxygenase 2, and fever. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently used antipyretic agents that most probably exert their antifever effect by inhibiting cyclooxygenase (COX)-2. Thus, COX-2-selective drugs or null mutation of the COX-2 gene reduce or prevent fever. Acetaminophen is antipyretic and analgesic, as are NSAIDs, but it lacks the anti-inflammatory and anticoagulatory properties of these drugs. This has led to the speculation that a COX variant exists that is inhibitable by acetaminophen. An acetaminophen-inhibitable enzyme is inducible in the mouse J774.2 monocyte cell line. Induction of acetaminophen-inhibitable prostaglandin E(2) synthesis parallels induction of COX-2. Thus, inhibition of pharmacologically distinct COX-2 enzyme activity by acetaminophen may be the mechanism of action of this important antipyretic drug. PMID- 11113026 TI - Toxicities of drugs used in the management of fever. AB - Fever is frequently managed outside the purview of medical professionals, and antipyretic therapy, on the whole, is generally considered safe. However, each of the drugs used in the management of fever has significant toxicities. The purpose of this review is to examine the relative safety of such agents with a focus on the nonsteroidal anti-inflammatory drugs and acetaminophen. Toxicity to the gastrointestinal, renal, and hepatic systems are considered; the comparative safety profile of acetaminophen and ibuprofen as antipyretics are highlighted; and specific recommendations to improve the safe use of these therapies are advanced. PMID- 11113027 TI - External cooling in the management of fever. AB - Although physical methods of cooling are the treatment of choice for hyperthermia, their value in the treatment of fever remains uncertain. Methods involving convection and evaporation are more effective than those involving conduction for the treatment of hyperthermia. These same methods, combined with antipyretic medication, are preferable to immersion as treatment for fever in young children but are generally not practical in adults. Febrile children treated with tepid-water sponging plus antipyretic drugs are more uncomfortable that those treated with antipyretic drugs alone, although they exhibit slightly more rapid reductions in temperature. When febrile, seriously ill patients are externally cooled and are sedated or paralyzed with drugs that suppress shivering, they may have a more rapid reduction of fever and reduced energy expenditure than if treated with antipyretic drugs alone. A risk/benefit assessment of the consequences of such treatment is not yet possible. PMID- 11113028 TI - Diagnostic implications and clinical consequences of antipyretic therapy. AB - It has been suggested that the response to antipyretic therapy might differentiate between fevers due to serious illness and fevers caused by less severe disorders; that neoplastic fevers are more responsive to nonsteroidal anti inflammatory drugs than are infectious fevers; that the metabolic costs of fever can exceeds its clinical benefits; that antipyretic therapy can prevent or reverse febrile seizures in children and fever-associated mental dysfunction in frail elderly patients. This article examines the data on which these assertions are based. PMID- 11113029 TI - Antipyretic therapy in patients with sepsis. AB - Sepsis is a clinical syndrome characterized by a systemic inflammatory response to infection. Mortality rates in sepsis have remained high, despite recent advances in our understanding of the immunological mechanisms that cause sepsis. Fever, a nonspecific acute-phase response, has been associated with improved survival and shortened disease duration in some infections. This article reviews the biological effects of fever and the influence of antipyretic therapy on the outcome in sepsis in experimental models and in humans and offers clinical recommendations for antipyretic therapy in early and late stages of the disorder. PMID- 11113030 TI - Antipyretic Therapy's future. AB - There is little doubt that clinicians will continue to seek new and, one hopes, more intelligent ways to suppress fever. In the process, new agents will be developed, new uses will be identified for existing antipyretic agents, new measures will be designed to maximize the benefits of antipyretic therapy while minimizing its adverse effects, and a concerted effort will be made to define more clearly and to promote appropriate indications for such therapy. PMID- 11113031 TI - Cortical change in Alzheimer's disease detected with a disease-specific population-based brain atlas. AB - We report the first detailed population-based maps of cortical gray matter loss in Alzheimer's disease (AD), revealing prominent features of early structural change. New computational approaches were used to: (i) distinguish variations in gray matter distribution from variations in gyral patterns; (ii) encode these variations in a brain atlas (n = 46); (iii) create detailed maps localizing gray matter differences across groups. High resolution 3D magnetic resonance imaging (MRI) volumes were acquired from 26 subjects with mild to moderate AD (age 75.8+/ 1.7 years, MMSE score 20.0+/-0.9) and 20 normal elderly controls (72.4+/-1.3 years) matched for age, sex, handedness and educational level. Image data were aligned into a standardized coordinate space specifically developed for an elderly population. Eighty-four anatomical models per brain, based on parametric surface meshes, were created for all 46 subjects. Structures modeled included: cortical surfaces, all major superficial and deep cortical sulci, callosal and hippocampal surfaces, 14 ventricular regions and 36 gyral boundaries. An elastic warping approach, driven by anatomical features, was then used to measure gyral pattern variations. Measures of gray matter distribution were made in corresponding regions of cortex across all 46 subjects. Statistical variations in cortical patterning, asymmetry, gray matter distribution and average gray matter loss were then encoded locally across the cortex. Maps of group differences were generated. Average maps revealed complex profiles of gray matter loss in disease. Greatest deficits (20-30% loss, P<0.001-0.0001) were mapped in the temporo parietal cortices. The sensorimotor and occipital cortices were comparatively spared (0-5% loss, P>0.05). Gray matter loss was greater in the left hemisphere, with different patterns in the heteromodal and idiotypic cortex. Gyral pattern variability also differed in cortical regions appearing at different embryonic phases. 3D mapping revealed profiles of structural deficits consistent with the cognitive, metabolic and histological changes in early AD. These deficits can therefore be (i) charted in a living population and (ii) compared across individuals and groups, facilitating longitudinal, genetic and interventional studies of dementia. PMID- 11113032 TI - Hemispheric and gender-related differences in the gross morphology of the anterior cingulate/paracingulate cortex in normal volunteers: an MRI morphometric study. AB - The sulci and gyri found within the anterior cingulate (AC), and across the cerebrum generally, have been found to vary in location and complexity from one individual to the next, making it difficult to analyze imaging data accurately and systematically. In this study, we examined the nature of morphometric variance in the AC of the left and right cerebral hemispheres using high resolution structural magnetic resonance imaging (MRI) acquired from 176 healthy volunteers. Depending on the presence of a paracingulate sulcus (PCS) and its antero-posterior extent, three types of AC patterns were identified: 'prominent', 'present' and 'absent'. Hemispheric comparisons across the whole sample showed the PCS to be more commonly 'prominent' in the left hemisphere and more commonly 'absent' in the right hemisphere. There was a significant gender difference, such that males showed an asymmetric pattern characterized by increased fissurization of the left AC, while females showed greater symmetry, with less fissurization of the left AC. Overall cerebral morphology, namely hemispheric volume and hemispheric fissurization, were also measured and used as independent variables as well as covariates in the analyses in order to ascertain the specificity of the results regarding AC morphology. Results showed that cerebral volume for males was larger on the right than on the left while fissurization showed the reverse asymmetry of greater leftward fissurization. In contrast, females were symmetric in both respects. The findings regarding AC morphology could not be explained by differences in these overall cerebral measures or by differences in age and handedness within the population. The results suggest that in the normal male brain, there exist morphological asymmetries at both the global and local levels that are less apparent in the female brain. The findings have implications for future studies examining the organization, development and functional anatomy of the AC. PMID- 11113033 TI - Differences in somatosensory processing in S1 barrel cortex between normal and monoamine oxidase A knockout (Tg8) adult mice. AB - Spatio-temporal processing of whisker information was analysed in vivo for single neurons in D2 barrel columns of S1 cortex in Tg8 mutant mice, which lack barrels. Findings were compared with normal C3H mice of the same genetic background. The topographical organization of functional columns was similar in Tg8 and normal mice. Response magnitudes (RMs) to D2 principal whisker deflections in D2 columns for Tg8 were similar to normals for layers I-III and layer IV cells but short latency responses (>10 ms post-stimulus) were twice the magnitude of normal mice. The surrounding whiskers D1 and D3 yielded smaller RMs in layer IV of mutants than normal mice whereas RMs in layers I-III were equipotent (P>0.5). Modal latencies were shorter in Tg8 mice in all layers. Latency distributions for whisker D2 responses in both laminae were bimodal in normal mice, peaking at 6-8 and 12 ms post-stimulus, but unimodal in Tg8 mice in both laminae, peaking at 6-8 ms. Hence, despite an absence of barrels, segregation of columns is enhanced in layer IV and sensory processing is faster in layers I-IV compared with normal mice. This contrasts with adenylyl cyclase knockout mice where both an absence of barrels and enhanced surrounding whisker responses have been observed. These findings suggest that factors other than barrels and clustering of thalamo cortical terminals define receptive field geometry. PMID- 11113034 TI - A neural model of how horizontal and interlaminar connections of visual cortex develop into adult circuits that carry out perceptual grouping and learning. AB - A neural model suggests how horizontal and interlaminar connections in visual cortical areas V1 and V2 develop within a laminar cortical architecture and give rise to adult visual percepts. The model suggests how mechanisms that control cortical development in the infant lead to properties of adult cortical anatomy, neurophysiology and visual perception. The model clarifies how excitatory and inhibitory connections can develop stably by maintaining a balance between excitation and inhibition. The growth of long-range excitatory horizontal connections between layer 2/3 pyramidal cells is balanced against that of short range disynaptic interneuronal connections. The growth of excitatory on-center connections from layer 6-to-4 is balanced against that of inhibitory interneuronal off-surround connections. These balanced connections interact via intracortical and intercortical feedback to realize properties of perceptual grouping, attention and perceptual learning in the adult, and help to explain the observed variability in the number and temporal distribution of spikes emitted by cortical neurons. The model replicates cortical point spread functions and psychophysical data on the strength of real and illusory contours. The on-center, off-surround layer 6-to-4 circuit enables top-down attentional signals from area V2 to modulate, or attentionally prime, layer 4 cells in area V1 without fully activating them. This modulatory circuit also enables adult perceptual learning within cortical area V1 and V2 to proceed in a stable way. PMID- 11113035 TI - Separate neural correlates for the mnemonic components of successive discrimination and working memory tasks. AB - We have used positron emission tomography to map the mnemonic components of two tasks at the extremes of the visual short-term/ working memory spectrum. The successive discrimination task requires only storage of a single item for very short time (ultra-short- term memory), while the 2back task requires both maintenance (i.e. storage and rehearsal) and manipulation of several items (working memory). We tested whether or not the storage component, common to the two tasks, engaged the same cerebral regions. To remove unnecessary confounds, we reduced the cues available to the subjects to a single elementary attribute, the orientation of a grating presented in central vision. This prevented subjects from using verbal strategies or vestibular cues and allowed equating of difficulty among tasks. Ultra-short-term memory for orientation engaged a large expanse of occipito-temporal cortex with a rate-dependent antero-posterior gradient: a fast trial rate engaged posterior regions, a slow trial rate anterior regions. On the other hand, working memory for orientation involved the left inferior parietal cortex, left dorsolateral prefrontal cortex and a left superior frontal sulcus region, and to a lesser degree the symmetrical right superior frontal region and a left superior parietal region. Direct comparison of the two orientation memory networks confirmed their functional segregation. We conclude that at least the storage of orientation information engages distinct regions depending on whether or not short-term memory/working memory involves rehearsal and/or manipulative processes. PMID- 11113036 TI - The noradrenergic alpha2 agonist clonidine modulates behavioural and neuroanatomical correlates of human attentional orienting and alerting. AB - We examined whether the known noradrenergic attenuation of the alerting effect (the beneficial effect of a warning cue) results from an underlying effect of noradrenaline on temporal orienting (orienting toward a particular moment in time). Following a within-subjects, counterbalanced design, 10 healthy human volunteers received placebo, 200 microg clonidine or 1 mg guanfacine (alpha2 agonists) in three separate testing sessions. Subjects were scanned by fMRI while performing attentional orienting tasks containing spatially informative, temporally informative, non-informative or no cues. The alerting effect primarily activated left-lateralized prefrontal, premotor and parietal regions. Clonidine, but not guanfacine, impaired behavioural measures of the alerting effect while attenuating activity in the left temporo-parietal junction. Replicating previous results, the temporal orienting task activated left parietal and frontal cortex, while parietal cortex was activated bilaterally during spatial orienting. Of these networks, clonidine, but not guanfacine, attenuated left prefrontal cortex and insula activity during temporal orienting and attenuated right superior parietal cortex activity during spatial orienting,. To complement these neuroanatomical changes, clonidine produced selective behavioural effects on both temporal and spatial orienting. The anatomical dissociation between the effects of clonidine during temporal orienting versus alerting suggests that noradrenergic modulation of the alerting effect does not result only from an underlying effect on temporal orienting. Furthermore, we have demonstrated lateralized neuroanatomical substrates for the noradrenergic modulation of human attentional orienting in the spatial and temporal domains. PMID- 11113037 TI - Dissociable mechanisms of attentional control within the human prefrontal cortex. AB - Neuropsychological tests that require shifting an attentional set, such as the Wisconsin Card Sorting Test, are sensitive to frontal lobe damage. Although little information is available for humans, an animal experiment suggested that different regions of the prefrontal cortex may contribute to set shifting behavior at different levels of processing. Behavioral studies also suggest that set shifting trials are more time consuming than non-set shifting trials (i.e. switch cost) and that this may be underpinned by differences at the neural level. We determined whether there were differential neural responses associated with two different levels of shifting behavior, that of reversal of stimulus-response associations within a perceptual dimension or that of shifting an attentional set between different perceptual dimensions. Neural activity in the antero-dorsal prefrontal cortex increased only in attentional set shifting, in which switch costs were significant. Activity in the postero-ventral prefrontal cortex increased not only in set shifting but also in reversing stimulus-response associations, in which switch costs were absent. We conclude that these distinct regions in the human prefrontal cortex provide different levels of attention control in response selection. Thus, the antero-dorsal prefrontal cortex may be critical for higher order control of attention, i.e. attentional set shifting, whereas the postero-ventral area may be related to a lower level of shift, i.e. reorganizing stimulus-response associations. PMID- 11113038 TI - Provisional versus routine stenting: routine stenting is here to stay. PMID- 11113039 TI - Effect of intracoronary gamma-radiation therapy on in-stent restenosis: An intravascular ultrasound analysis from the gamma-1 study. AB - BACKGROUND: The aim of this study was to use serial volumetric intravascular ultrasound to evaluate the effect of gamma-radiation on recurrent in-stent restenosis. METHODS AND RESULTS: After successful reintervention, patients were randomized to receive either (192)Ir or placebo. Intravascular ultrasound studies with motorized pullback (0.5 mm/s) were performed immediately after irradiation and at 8-month follow-up in 70 patients. Paired volumetric analysis of the stented segment and of 5-mm proximal and distal reference segments was performed; this included measurements of the external elastic membrane, lumen, plaque and media (external elastic membrane minus lumen), stent, and intimal hyperplasia (stent minus lumen). Baseline proximal reference, stent, and distal reference measurements were similar in both groups. The changes in proximal and distal reference measurements of the external elastic membrane, plaque and media, and lumen areas were similar in both groups. However, the decrease in stented segment lumen volume was less in the (192)Ir patients than the placebo patients (-25+/-34 mm(3) versus -48+/-42 mm(3); P:=0.0225), and the increase in the volume of intimal hyperplasia in the stented segment was less in the (192)Ir patients than in the placebo patients (28+/-37 mm(3) versus 50+/-40 mm(3); P:=0.0352). When averaged over the length of the stented segment (32+/-13 mm versus 33+/-14 mm; P:=0.9), the increase in mean area of intimal hyperplasia was 0.8+/-1.0 mm(2) in the (192)Ir group and 1.6+/-1.2 mm(2) in the control group (P:=0.0065). Late stent-vessel wall malapposition was noted in one placebo patient and no (192)Ir patients. CONCLUSIONS: gamma-Radiation therapy can effectively prevent recurrent in-stent restenosis by inhibiting neointimal formation within the stent. At the stent edge, there were no significant differences between (192)Ir and placebo patients. PMID- 11113040 TI - Transfer of CD4(+) T cells aggravates atherosclerosis in immunodeficient apolipoprotein E knockout mice. AB - BACKGROUND: Atherosclerosis is associated with immune responses to oxidized lipoproteins and certain microorganisms, but the role of specific immunity has remained unclear. METHODS AND RESULTS: To study the role of immunity in atherosclerosis, we crossed atherosclerosis-prone apoE(-/-) mice with immunodeficient scid/scid mice. The offspring showed a 73% reduction in aortic fatty streak lesions when compared with immunocompetent apoE(-/-) mice. Transfer of CD4(+) T cells from apoE(-/-) to immunodeficient apoE(-/-)/scid/scid mice increased lesions by 164%. This was associated with the infiltration of transferred T cells into lesions, increased circulating interferon-gamma levels, and increased I-A expression in lesions. CONCLUSIONS: CD4(+) T cells carry disease-promoting immunity in atherosclerosis. PMID- 11113041 TI - Economic assessment of platelet glycoprotein IIb/IIIa receptor blockade with abciximab and low-dose heparin during percutaneous coronary revascularization: results from the EPILOG randomized trial. Evaluation in PTCA to Improve Long-term Outcome with abciximab GP IIb/IIIa blockade. AB - BACKGROUND: In the EPILOG trial (Evaluation in PTCA to Improve Long-term Outcome with abciximab GP IIb/IIIa blockade), abciximab administered with weight-adjusted heparin diminished the risk of ischemic complications within 30 days by 56% among patients undergoing percutaneous coronary revascularization, without increased bleeding complications. METHODS AND RESULTS: A prospective economic assessment was performed in the 2792 patients enrolled in EPILOG. Patients were randomized to receive placebo with standard-dose weight-adjusted heparin, abciximab with low dose weight-adjusted heparin, or abciximab with standard-dose weight-adjusted heparin during percutaneous coronary intervention. Hospital billing data for the baseline hospitalization were collected for 2581 patients (92.4% of total) and imputed for the remainder, with physician fees estimated from the Medicare Fee Schedule. For the baseline hospitalization, medical costs (hospitalization and physician fees) averaged $9632 for the placebo arm compared with $8758 (P:=0.005) and $9092 (P:=0.176) for the abciximab with low-dose and standard-dose heparin arms, respectively. Inclusive of average drug cost ($1454 to $1457), the net incremental baseline cost of these 2 abciximab strategies was $583 with low-dose weight-adjusted heparin and $914 with standard-dose weight-adjusted heparin. During 6-month follow-up, average hospital costs were not significantly different in the 3 treatment groups; cumulative net incremental costs were $1236 and $1268 in the abciximab with low-dose and standard-dose heparin groups, respectively. CONCLUSIONS: Treatment with abciximab and low-dose, weight-adjusted heparin during percutaneous coronary revascularization reduces ischemic events and associated costs, thereby offsetting some of the cost of the drug. The suppression of bleeding complications associated with this agent by heparin dose reduction optimizes the economic attractiveness of this treatment strategy. PMID- 11113042 TI - Randomized comparison of primary stenting and provisional balloon angioplasty guided by flow velocity measurement. Doppler Endpoints Balloon Angioplasty Trial Europe (DEBATE) II Study Group. AB - BACKGROUND: Coronary stenting improves outcomes compared with balloon angioplasty, but it is costly and may have other disadvantages. Limiting stent use to patients with a suboptimal result after angioplasty (provisional angioplasty) may be as effective and less expensive. METHODS AND RESULTS: To analyze the cost-effectiveness of provisional angioplasty, patients scheduled for single-vessel angioplasty were first randomized to receive primary stenting (97 patients) or balloon angioplasty guided by Doppler flow velocity and angiography (523 patients). Patients in the latter group were further randomized after optimization to either additional stenting or termination of the procedure to further investigate what is "optimal." An optimal result was defined as a flow reserve >2.5 and a diameter stenosis <36%. Bailout stenting was needed in 129 patients (25%) who were randomized to balloon angioplasty, and an optimal result was obtained in 184 of the 523 patients (35%). There was no significant difference in event-free survival at 1 year between primary stenting (86.6%) and provisional angioplasty (85.6%). Costs after 1 year were significantly higher for provisional angioplasty (EUR 6573 versus EUR 5885; P:=0.014). Results after the second randomization showed that stenting was also more effective after optimal balloon angioplasty (1-year event free survival, 93.5% versus 84.1%; P:=0. 066). CONCLUSIONS: After 1 year of follow-up, provisional angioplasty was more expensive and without clinical benefit. The beneficial value of stenting is not limited to patients with a suboptimal result after balloon angioplasty. PMID- 11113043 TI - Randomized comparison of elective stent implantation and coronary balloon angioplasty guided by online quantitative angiography and intracoronary Doppler. DESTINI Study Group (Doppler Endpoint STenting INternational Investigation). AB - BACKGROUND: The purpose of this study was to compare long-term outcomes of coronary stenting in all lesions (elective stenting) or only in lesions with inadequate morphological and functional results after balloon angioplasty (guided PTCA). METHODS AND RESULTS: Treatment of multivessel disease, with any lesion length and vessel size, was allowed provided that all lesions were suitable for stent implantation. Patients were randomized to elective stent implantation (n=370) or guided PTCA (n=365). An optimal PTCA result (residual diameter stenosis 2.0, absence of threatening dissections) was achieved in 166 lesions (43%). The remaining 218 lesions underwent stent implantation (provisional stenting). Final residual diameter stenosis was lower in the elective and provisional stent groups (9.3% and 10.2%) than in the optimal PTCA group (24.8%, P:<0. 00001). On an intention-to-treat analysis, the probability of >/=1 major adverse cardiac event at 12 months was 17.8% in the elective stenting group and 18.9% in the guided PTCA group (20.1% for optimal PTCA and 18.0% for the provisional stenting subgroup, P:=NS). The incidence of repeat target lesion revascularization at 1 year was 14. 9% in the elective stent group and 15.6% in the guided PTCA group (17.6% for optimal PTCA and 14.1% for the provisional stenting subgroup, P:=NS). CONCLUSIONS: When balloon angioplasty is guided by online quantitative angiography and Doppler-derived coronary flow reserve, with provisional stenting reserved for suboptimal results, early and late clinical outcomes are comparable to those achieved by elective stenting of all patients. PMID- 11113044 TI - Percutaneous coronary intervention in the current era compared with 1985-1986: the National Heart, Lung, and Blood Institute Registries. AB - BACKGROUND: Although refinements have occurred in coronary angioplasty over the past decade, little is known about whether these changes have affected outcomes. METHODS AND RESULTS: Baseline features and in-hospital and 1-year outcomes of 1559 consecutive patients in the 1997-1998 Dynamic Registry who were having first coronary intervention were compared with 2431 patients in the 1985-1986 National Heart, Lung, and Blood Institute Registry. Compared with patients in the 1985 1986 Registry, Dynamic Registry patients were older (mean age, 62 versus 58 years; P:<0.001) and more often female (32.1% versus 25.5%; P:<0.001). In the Dynamic Registry, procedures were more often performed for acute myocardial infarction (22.9% versus 9.9%; P:<0.001) and treated lesions were more severe (84.5% versus 82.5% diameter reduction; P:<0.001), thrombotic (22.1% versus 11.3%; P:<0.001) or calcified (29.5% versus 10.8%; P:<0.001). Stents were used in 70.5% of Dynamic Registry patients, whereas 1985-1986 patients received balloon angioplasty alone. Procedural success was higher in the Dynamic Registry (92.0% versus 81.8%; P:<0.001) and the rate of in-hospital death, myocardial infarction, and emergency coronary bypass surgery combined was lower (4.9% versus 7.9%; P:=0.001) than in the 1985-1986 Registry. The 1-year rate for CABG was lower in the Dynamic Registry (6.9% versus 12.6%; P:<0.001). CONCLUSIONS: Although Dynamic Registry patients had more unstable and complex coronary disease than those in the 1985-1986 Registry, their rate of procedural success was higher whereas rates of complications and subsequent CABG were lower. Results of percutaneous coronary intervention have improved substantially over the past decade. PMID- 11113045 TI - Enhanced efficacy of eptifibatide administration in patients with acute coronary syndrome requiring in-hospital coronary artery bypass grafting. PURSUIT Investigators. AB - BACKGROUND: Patients with a recent episode of non-ST-segment elevation acute coronary syndrome before CABG have higher rates of operative morbidity and mortality than patients with stable coronary syndromes. The efficacy of administering eptifibatide to these patients undergoing in-hospital CABG is unknown. METHODS AND RESULTS: The Platelet Glycoprotein IIb-IIIa in Unstable Angina: Receptor Suppression Using Integrilin Therapy (PURSUIT) trial randomized 10 948 patients to receive either eptifibatide or placebo. There were 1558 study participants who underwent in-hospital CABG: 692 received placebo, and 866 received eptifibatide. The main substudy analysis end point was death or myocardial infarction (MI) rates at the 6-month follow-up. The 30-day death or MI rates were 30. 8% and 26.1% for the placebo and eptifibatide groups, respectively (P:=0.041). The benefit of eptifibatide administration persisted through 6-months of follow-up (32.7% versus 27.6% for placebo versus eptifibatide, respectively; P:=0.029). There was a greater reduction in the 6-month death or MI rate for patients who received eptifibatide within 72 hours of CABG (33.6% versus 23.8%; P:=0.002) compared with the >72-hour group (31.6% versus 32%; P:=1.0). The incidence of major bleeding was 56.6% for placebo-treated patients versus 58.2% for eptifibatide-treated patients (P:=0.7). CONCLUSIONS: Eptifibatide administration in patients undergoing in-hospital CABG with a recent episode of a non-ST-segment elevation acute coronary syndrome results in a significant reduction in death or MI that is evident at 7 days and persists through the 6 month follow-up without a significant increase in perioperative bleeding rates. PMID- 11113046 TI - Immediate changes of collateral function after successful recanalization of chronic total coronary occlusions. AB - BACKGROUND: Coronary collaterals are essential to maintain myocardial function in chronic total coronary occlusions (TCOs). The aim of the present study was to assess the collateral circulation in TCOs before coronary angioplasty and to determine the recruitable collateral perfusion after recanalization by use of intracoronary Doppler flow velocimetry. METHODS AND RESULTS: In 21 patients with TCOs (duration >4 weeks), Doppler recordings of basal collateral flow were obtained before the first balloon inflation. Angioplasty was performed with stent implantation in all lesions. At the end of the procedure, recruitable collateral flow was measured during a repeat balloon inflation. The collateral flow index (CFI) was calculated from the velocity integral during the occlusion/velocity integral of antegrade flow. In 17 of 21 patients, angiography was repeated after 24 hours, and CFI was reassessed. Average peak velocity of collateral flow was 10.9+/-5.6 cm/s with a predominantly systolic flow (diastolic/systolic velocity ratio <0.5) compared with antegrade flow (diastolic/systolic velocity ratio >1.5). After recanalization, the average peak velocity of recruitable collateral flow dropped by >50% to 4.7+/-2.5 cm/s. CFI fell from 0.48+/-0.25 to 0.21+/-0.16 (P:<0.001). There was no further change of CFI during the following 24 hours. CFI was higher in patients with preserved regional ventricular function than in those with akinetic myocardium (0.57+/-0.23 versus 0.38+/-0.12, P:<0.05). CONCLUSIONS: Collateral circulation in TCO provided 50% of antegrade coronary flow. A considerable fraction of collateral flow was immediately lost after recanalization, indicating that TCO may not remain protected from future ischemic events by a well-developed collateral function. PMID- 11113047 TI - Are patients with renal failure good candidates for percutaneous coronary revascularization in the new device era? AB - BACKGROUND: Patients with end-stage renal disease undergoing conventional balloon angioplasty have reduced procedural success and increased complication rates. This study was designed to determine the immediate and long-term outcomes of patients with varying degrees of renal failure undergoing percutaneous coronary intervention in the current device era. METHODS AND RESULTS: We compared the immediate and long-term outcomes of 362 renal failure patients (creatinine >1.5 mg/dL) with those of 2972 patients with normal renal function who underwent percutaneous coronary intervention between 1994 and 1997. Patients with renal failure were older and had more associated comorbidities. They had reduced procedural success (89.5% versus 92.9%, P:=0.007) and greater in-hospital combined major event (death, Q-wave myocardial infarction, emergent CABG; 10.8% versus 1.8%; P:<0.0001) rates. Renal failure was an independent predictor of major adverse cardiac events (MACEs) (OR, 3.41; 95% CI, 1.84 to 6.22; P:<0.00001). Logistic regression analysis identified shock, peripheral vascular disease, balloon angioplasty strategy, and unstable angina as independent predictors of in-hospital MACEs in the renal group. Compared with 362 age- and sex-matched patients selected from the control group, patients with renal failure had a lower survival rate (27.7% versus 6.1%, P:<0.0001) and a greater MACE rate (51% versus 33%, P:<0.001) at long-term follow-up. Cox regression analysis identified age and PTCA strategy as independent predictors of long-term MACEs in the renal group. Finally, within the renal failure population, the dialysis and nondialysis patients experienced remarkably similar immediate and long-term outcomes. CONCLUSIONS: Although patients with renal failure can be treated with a high procedural success rate in the new device era, they have an increased rate of major events both in hospital and at long-term follow-up. Nevertheless, utilization of stenting and debulking techniques improves immediate and long-term outcomes. PMID- 11113048 TI - Risks of morbidity and mortality in dialysis patients undergoing coronary artery bypass surgery. Northern New England Cardiovascular Disease Study Group. AB - BACKGROUND: Although dialysis patients are undergoing CABG with increasing frequency, large studies specifically comparing patient characteristics and procedure-related risks in this population have not been performed. METHODS AND RESULTS: We conducted a regional prospective cohort study of 15,500 consecutive patients undergoing CABG in northern New England from 1992 to 1997. We used multiple logistic regression analysis to examine associations between preoperative dialysis-dependent renal failure and postoperative events and to adjust for potentially confounding variables. The 279 dialysis-dependent renal failure patients (1.8%) were 4.4 times more likely to experience in-hospital mortality than were other CABG patients (12.2% versus 3.0%, respectively; P:<0.001). Dialysis-dependent renal failure patients were older and had more comorbidities and more severe cardiac disease than did other CABG patients. After adjusting for these factors in multivariate analysis, however, dialysis-dependent renal failure patients remained 3.1 times more likely to die after CABG (adjusted odds ratio [OR] 3.1, 95% CI 2.1 to 4.7; P:<0.001). Dialysis-dependent renal failure patients compared with other CABG patients also had a substantially increased risk of postoperative mediastinitis (3.6% versus 1.2%, respectively; adjusted OR 2.4, 95% CI 1.2 to 4.7; P:=0.011) and postoperative stroke (4.3% versus 1.7%, respectively; adjusted OR 2. 1, 95% CI 1.1 to 3.9; P:=0.016), even after controlling for potentially confounding variables. Risks of reexploration for bleeding were similar for patients with and without dialysis-dependent renal failure. CONCLUSIONS: Preoperative dialysis-dependent renal failure is a strong independent risk factor for in-hospital mortality and mediastinitis after CABG. PMID- 11113049 TI - Evidence of Trypanosoma cruzi infection (Chagas' disease) among patients undergoing cardiac surgery. AB - BACKGROUND: Trypanosoma cruzi, the agent of Chagas' heart disease, is transmitted by triatomine insects and by blood transfusion. The emigration of several million people from T cruzi-endemic countries to the United States has raised concerns regarding a possible increase in cases of Chagas' heart disease here, as well as an increased risk of transfusion-transmitted T cruzi. To investigate these 2 possible outcomes, we tested a repository of blood specimens from multiply transfused cardiac surgery patients for antibodies to T cruzi. METHODS AND RESULTS: Postoperative blood specimens from 11 430 cardiac surgery patients were tested by enzyme immunoassay, and if repeat-reactive, were confirmed by radioimmunoprecipitation. Six postoperative specimens (0.05%) were confirmed positive. Corresponding preoperative specimens, available for 4 of these patients, were also positive. The other 2 patients had undergone heart transplantations. Tissue samples from their excised hearts were tested for T cruzi by polymerase chain reaction and were positive. Despite the fact that several of these 6 patients had histories and clinical findings suggestive of Chagas' disease, none of them were diagnosed with or tested for it. Patient demographics showed that 5 of 6 positive patients were Hispanic, and overall, 2. 7% of Hispanic patients in the repository were positive. CONCLUSIONS: No evidence for transfusion-transmitted T cruzi was found. All 6 seropositive patients apparently were infected with T cruzi before surgery; however, a diagnosis of Chagas' disease was not known or even considered in any of these patients. Indeed, Chagas' disease may be an underdiagnosed cause of cardiac disease in the United States, particularly among patients born in countries in which T cruzi is endemic. PMID- 11113050 TI - Effects of estrogen replacement on infarct size, cardiac remodeling, and the endothelin system after myocardial infarction in ovariectomized rats. AB - BACKGROUND: Estrogen may increase the long-term survival of women who have suffered from a myocardial infarction (MI). We examined the acute and chronic influence of estrogen on MI in the rat left coronary artery ligation model. METHODS AND RESULTS: Female Sprague-Dawley rats (10 to 12 weeks, n=93), divided into 3 groups (rats with intact ovaries, ovariectomized rats administered 17beta estradiol [17beta-E(2)] replacement, and ovariectomized rats administered placebo 2 weeks before MI), were randomized to left coronary artery ligation (n=66) or sham-operated (n=27) groups. Ten to 11 weeks after MI, rats were randomly assigned to either (1) assessment of left ventricular (LV) function and morphometric analysis or (2) measurement of cardiopulmonary mRNA expression of preproendothelin-1 and endothelin A and B receptors. Acutely, estrogen was associated with a trend toward increased mortality. Infarct size was increased in the 17beta-E(2) group compared with the placebo group (42+/-2% versus 26+/-3%, respectively; P:=0.01). Chronically, wall tension was normalized through a reduction in LV cavity size with estrogen treatment (419+/-41 mm Hg/mm for 17beta E(2) versus 946+/-300 mm Hg/mm for placebo, P:=0.039). In the LV, there was a 2.5 fold increase in endothelin B mRNA expression after MI in placebo-treated rats (P:=0.004 versus sham-operated rats) that was prevented in the 17beta-E(2) group (P:=NS versus sham-operated rats). CONCLUSIONS: These results suggest that estrogen is detrimental at the time of MI or early post-MI period, resulting in an increased size of infarct or infarct expansion, but chronically, it can normalize wall tension and inhibit LV dilatation, which may in turn lead to increased long-term survival. Regulation of the endothelin system, particularly the expression of the endothelin B receptor, may contribute to these estrogenic effects. PMID- 11113051 TI - Antioxidant effect of estrogen on cytomegalovirus-induced gene expression in coronary artery smooth muscle cells. AB - BACKGROUND: Pathogens infecting the arterial wall with resultant inflammation may contribute to atherogenesis. Human coronary artery smooth muscle cells (SMCs) infected with human cytomegalovirus (CMV) demonstrate a rapid increase in reactive oxygen species (ROSs), with activation of genes involved in viral replication and inflammation. Because estrogen appears to have antioxidant properties, we wished to determine whether this hormone attenuates SMC responses to CMV infection. METHODS AND RESULTS: Using confocal microscopy and an intracellular fluorescent dye activated by ROSs, we found that 17beta-estradiol (0.1 to 10 nmol/L) and its stereoisomer 17alpha-estradiol (which has low affinity for the estrogen receptor) dose-dependently inhibited ROS generation in CMV infected SMCs. These effects were not blocked by the estrogen receptor inhibitor ICI 182,780. 3-Methoxyestrone, which lacks the phenolic hydroxyl group, did not interfere with ROS generation. We found that 17beta-estradiol and 17alpha estradiol, but not 3-methoxyestrone, prevented binding of nuclear factor (NF) kappaB to DNA. Furthermore, in SMCs transfected with the reporter constructs 3XkappaB-CAT, MIEP-CAT, or ICAM-CAT, cotransfection with a CMV-IE72 expression plasmid caused promoter and CAT activation. Treatment with 17beta-estradiol and 17alpha-estradiol, but not 3-methoxyestrone, inhibited CAT activity and, in CMV infected SMCs, prevented IE72 and ICAM-1 protein expression and cytopathic effects. CONCLUSIONS: These findings indicate that estrogen molecules with an A ring hydroxyl group have estrogen receptor-independent anti-CMV effects at physiological concentrations by inhibiting ROS generation, NF-kappaB activation, NF-kappaB-dependent transcription, and viral replication. To the extent that chronic infection of the vascular wall with CMV contributes to atherogenesis, these antioxidant actions of estrogen may be of therapeutic importance. PMID- 11113052 TI - Cyclic nucleotide phosphodiesterase type 5 activity limits blood flow to hypoperfused myocardium during exercise. AB - BACKGROUND: Nitric oxide (NO) causes vasodilation by stimulation of guanylate cyclase in vascular smooth muscle to produce cGMP. The resultant vasodilator effect is regulated by a family of cGMP phosphodiesterases (PDEs). Sildenafil, a selective inhibitor of PDE5 used for treatment of erectile dysfunction, has been found to cause relaxation of isolated epicardial coronary artery segments. The present study examined the effects of sildenafil on coronary blood flow and hemodynamics during exercise in normal and ischemic heart. METHODS AND RESULTS: In chronically instrumented normal dogs, sildenafil 2 mg/kg PO caused a slight but significant increase in left anterior descending (LAD) coronary blood flow during resting conditions, with a nonsignificant trend toward increased coronary flow during treadmill exercise. Exercise in the presence of LAD stenosis that decreased distal coronary pressure to 57+/-2 mm Hg reduced LAD flow during exercise from 69+/-8 to 41+/-7 mL/min (P:<0. 05), with hypoperfusion most severe in the subendocardium. At the same distal coronary pressure, sildenafil increased LAD flow in the ischemic region to 50+/-11 mL/min (P:<0.05). Increase in ischemic region blood flow produced by sildenafil was uniform across the LV wall, given that no change occurred in the transmural distribution of perfusion. CONCLUSIONS: Inhibition of PDE5 with sildenafil caused vasodilation of coronary resistance vessels with an increase of blood flow into an ischemic myocardial region during exercise in the presence of coronary artery stenosis. PMID- 11113053 TI - Trypanosoma cruzi-infected cardiomyocytes produce chemokines and cytokines that trigger potent nitric oxide-dependent trypanocidal activity. AB - BACKGROUND: The pathogenesis of myocarditis that occurs in Trypanosoma cruzi infected mice is still poorly understood. Therefore, it is important to know the mediators that trigger leukocyte migration to the heart as well as the cellular source of these possible mediators. In this study, we investigated (1) NO synthase (NOS) induction, (2) NO synthesis, (3) trypanocidal activity, and (4) chemokine and cytokine mRNA expression by isolated cardiomyocytes infected with T cruzi. METHODS AND RESULTS: Mouse cardiomyocytes were isolated, infected with T cruzi, and evaluated for induction of inducible NOS (iNOS), nitrite production, trypanocidal activity, and cytokine and chemokine mRNA expression. We found that T cruzi-infected murine embryonic cardiomyocytes produced nitrite and expressed mRNAs for the chemokines chemokine growth-related oncogene, monokine induced by interferon-gamma, macrophage inflammatory protein-2, interferon-gamma-inducible protein, RANTES, and monocyte chemotactic protein, for iNOS, and for the cytokines tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta. Separate addition of IL-1beta, interferon-gamma, TNF-alpha or monocyte chemotactic protein, macrophage inflammatory protein-2, and interferon-gamma-inducible protein, to cultured cardiomyocytes resulted in NO production but low trypanocidal activity. However, simultaneous addition of IL-1beta, interferon gamma, and TNF-alpha or the chemokines to cultures resulted in the induction of iNOS, high levels of nitrite, and a marked trypanocidal activity. The iNOS/L arginine pathway mediated the latter activity, inasmuch as it was inhibited by treatment with N:(G)-monomethyl-L-arginine. CONCLUSIONS: These results indicate that iNOS activation and the proinflammatory cytokines and chemokines produced by cardiomyocytes are likely to control parasite growth and cell influx, thus contributing to the pathogenesis of chagasic cardiomyopathy seen in T cruzi infected mice. PMID- 11113054 TI - Endogenous Na,K pump ligands are differentially regulated during acute NaCl loading of Dahl rats. AB - BACKGROUND: Two mammalian digitalis-like factors, an ouabain-like compound (OLC) and marinobufagenin (MBG), exhibit specificity to alpha-3 and alpha-1 Na(+),K(+) ATPase isoforms, respectively. We compared regulation of MBG and OLC by acute NaCl loading in Dahl salt-sensitive (DS) and salt-resistant (DR) rats. METHODS AND RESULTS: An intraperitoneal NaCl load (0.8 g/kg) was given to adult male rats (24 DS and 24 DR). Diuresis, natriuresis, renal excretion, and tissue levels of MBG and OLC were measured. Inhibition of renal Na(+),K(+)-ATPase by MBG and ouabain was compared in DS, DR, and Wistar rats. DS (versus DR) exhibited a smaller peak (2 hours) natriuretic response (1.34+/-0.10 versus 2.08+/-0.14 mmol. kg(-)(1). h(-)(1); P:<0.01), despite a greater plasma Na(+) (153+/-2 versus 145+/ 1 mmol/L; P:<0.01). In DS and DR, pituitary, adrenal, and plasma OLC exhibited transient 2-fold to 3-fold increases, followed by a decrease to baseline levels. Plasma and adrenal MBG doubled in both strains within 1 hour of NaCl loading and remained elevated. Eight-hour MBG excretion in DS was 4-fold greater than in DR (15. 8+/-0.8 versus 3.6+/-0.4 pmol; P:<0.01), whereas OLC excretion in DS was only 30% greater than in DR (16.1+/-1.1 and 11.9+/-0.8 pmol; P:<0.05). Kidney Na(+),K(+)-ATPase (alpha-1 isoform) from Wistar rats and DS exhibited greater sensitivity to MBG than to ouabain. CONCLUSIONS: NaCl loading of DS causes transient increase in OLC but sustained increases in MBG tissue levels and excretion. We hypothesize that increased MBG production occurs in an attempt to compensate for genetically impaired pressure-natriuresis mechanisms. PMID- 11113055 TI - Endotoxin-induced mortality is related to increased oxidative stress and end organ dysfunction, not refractory hypotension, in heme oxygenase-1-deficient mice. AB - BACKGROUND: Heme oxygenase (HO)-1 is an enzyme that degrades heme to generate CO (a vasodilatory gas), iron, and the potent antioxidant bilirubin. A disease process characterized by decreases in vascular tone and increases in oxidative stress is endotoxic shock. Moreover, HO-1 is markedly induced in multiple organs after the administration of endotoxin (lipopolysaccharide [LPS]) to mice. METHODS AND RESULTS: To determine the role of HO-1 in endotoxemia, we administered LPS to mice that were wild-type (+/+), heterozygous (+/-), or homozygous null (-/-) for targeted disruption of HO-1. LPS produced a similar induction of HO-1 mRNA and protein in HO-1(+/+) and HO-1(+/-) mice, whereas HO-1(-/-) mice showed no HO-1 expression. Four hours after LPS, systolic blood pressure (SBP) decreased in all the groups. However, SBP was significantly higher in HO-1(-/-) mice (121+/-5 mm Hg) after 24 hours, compared with HO-1(+/+) (96+/-7 mm Hg) and HO-1(+/-) (89+/-13 mm Hg) mice. A sustained increase in endothelin-1 contributed to this SBP response. Even though SBP was higher, mortality was increased in HO-1(-/-) mice, and they exhibited hepatic and renal dysfunction that was not present in HO 1(+/+) and HO-1(+/-) mice. The end-organ damage and death in HO-1(-/-) mice was related to increased oxidative stress. CONCLUSIONS: These data suggest that the increased mortality during endotoxemia in HO-1(-/-) mice is related to increased oxidative stress and end-organ (renal and hepatic) damage, not to refractory hypotension. PMID- 11113056 TI - Granulomatous aortitis presenting as an acute myocardial infarction in Crohn's disease. PMID- 11113058 TI - The naming of jugular venous valves. PMID- 11113057 TI - Fen/Phen and valvular heart disease: the final link has now been established. PMID- 11113059 TI - Ventilatory and heart rate responses to exercise: better predictors of heart failure mortality than peak exercise oxygen consumption. PMID- 11113060 TI - Smoking and aldosterone synthase polymorphism. PMID- 11113061 TI - Late-breaking trials at American Heart Association's Scientific Sessions 2000. PMID- 11113062 TI - Odysseus's lament: death of mentor. PMID- 11113063 TI - A randomized trial comparing wireless capsule endoscopy with push enteroscopy for the detection of small-bowel lesions. AB - BACKGROUND & AIMS: Wireless capsule endoscopy is a new, painless method of imaging the entire small bowel. It has not been compared with push enteroscopy. We compared the sensitivity, specificity, and safety of capsule and push enteroscopy in detecting small-bowel lesions. METHODS: Nine to 13 radiopaque, colored beads (3-6 mm diameter) were sewn in random order inside 9 canine small bowels, half within the first meter, and confirmed on x-ray. After recovery, the number, order, and color of beads were assessed in 23 capsule enteroscopies and 9 push enteroscopies in a random order. The surgeons, push enteroscopists, capsule video interpreters, and pathologist were blinded to the others' findings. RESULTS: The capsules identified more beads than push enteroscopy (median, 6 [range, 2-9] vs. 3 [range, 2-6 beads]; P < 0.001). The sensitivity of the capsule was 64% compared with 37% for push enteroscopy. The specificity was 92% for capsule enteroscopy and 97% for push enteroscopy. The capsules identified significantly more beads beyond the reach of the push enteroscope (median, 4 [range, 2-7] vs. 0; P < 0.0001). Hair, ingested plastic, ulceration, submucosal swelling, and worms were clearly identified by the capsule. The capsules passed safely through the animals with no significant histologic findings. CONCLUSIONS: Wireless capsule endoscopy detected more abnormalities in the small bowel than push enteroscopy. PMID- 11113064 TI - Excess gastroesophageal reflux in patients with hiatus hernia is caused by mechanisms other than transient LES relaxations. AB - BACKGROUND & AIMS: Esophageal acid exposure is higher in gastroesophageal reflux disease (GERD) patients with hiatus hernia than in those without. We investigated the effect of a sliding hiatus hernia on the mechanisms underlying spontaneous gastroesophageal reflux over 24 hours. METHODS: Twelve GERD patients with and 10 GERD patients without hiatus hernia were studied for 24 hours. Combined esophageal pH and manometric recordings of the pharynx, lower esophageal sphincter (LES), and stomach were performed using a multiple-lumen assembly incorporating a Dent sleeve connected to a portable water-perfused manometric system and a pH glass electrode. RESULTS: Patients with hiatus hernia had greater esophageal acid exposure (7.6% vs. 3.3%; P < 0.01) and more reflux episodes (3.1 vs. 1.8/h; P < 0.001) than those without. LES pressure, the incidence of transient LES relaxations (TLESRs), and the proportion of TLESRs associated with acid reflux were comparable in both groups. Both groups had equal numbers of reflux episodes associated with TLESRs and swallow-associated prolonged LES relaxations. Patients with hiatus hernia had more reflux associated with low LES pressure, swallow-associated normal LES relaxations, and straining during periods with low LES pressure. CONCLUSIONS: The excess reflux in GERD patients with hiatus hernia compared with those without is caused by malfunction of the gastroesophageal barrier during low LES pressure, swallow-associated normal LES relaxations, deep inspiration, and straining. PMID- 11113065 TI - Very high risk of cancer in familial Peutz-Jeghers syndrome. AB - BACKGROUND & AIMS: The Peutz-Jeghers syndrome (PJS) is an autosomal dominant polyposis disorder with increased risk of multiple cancers, but literature estimates of risk vary. METHODS: We performed an individual patient meta-analysis to determine the relative risk (RR) of cancer in patients with PJS compared with the general population based on 210 individuals described in 6 publications. RESULTS: For patients with PJS, the RR for all cancers was 15.2 (95% confidence limits [CL], 2, 19). A statistically significant increase of RR was noted for esophagus (57; CL, 2.5, 557), stomach (213; CL, 96, 368), small intestine (520; CL, 220, 1306), colon (84; CL, 47, 137), pancreas (132; CL, 44, 261), lung (17.0; CL, 5.4, 39), breast (15.2; CL, 7.6, 27), uterus (16.0; CL, 1.9, 56), ovary (27; CL, 7.3, 68), but not testicular or cervical malignancies. Cumulative risk for all cancer was 93% from age 15 to 64 years old. CONCLUSIONS: Patients with PJS are at very high relative and absolute risk for gastrointestinal and nongastrointestinal cancers. PMID- 11113066 TI - Ileorectal anastomosis is appropriate for a subset of patients with familial adenomatous polyposis. AB - BACKGROUND & AIMS: This study reevaluates the risk of rectal cancer and the frequency of subsequent proctectomy for nonmalignant causes in patients with familial adenomatous polyposis (FAP) who have undergone colectomy with ileorectal anastomosis (IRA). Potential risk factors for rectal cancer in this setting are also examined, and recommendations for the choice of surgical procedure are made. METHODS: The national polyposis registries in Denmark, Finland, The Netherlands, and Sweden included 659 patients undergoing surgery with IRA in 1940-1997. Kaplan Meier analysis and Cox regression analysis were performed to evaluate cumulative risk, survival, and predictive risk factors. RESULTS: Rectal carcinoma was diagnosed in 47 patients, with a cumulative 40-year risk of 0.32. The cumulative risk according to chronologic age was 0.30 at age 60, and higher in patients undergoing surgery above age 25 (P = 0.0016). Chronologic age was the only independent risk factor (P = 0.0016). The cumulative 5-year survival rate after rectal carcinoma was 0.60. The apc mutation was known in 167 patients, of whom 7 had rectal cancer. The cumulative 40-year risk of secondary proctectomy was 0.70, and higher in patients with a mutation in codon 1250-1500 than outside this region (P = 0.005). However, all 7 rectal cancers were found in the latter group. None of the 18 patients with attenuated FAP (mutation in codon 0-200 or >1500) had a secondary proctectomy. CONCLUSIONS: IRA is recommended in (1) young patients with few rectal adenomas and a family history of a mild phenotype and (2) patients with attenuated FAP (a mutation in codon 0-200 or >1500), provided there is acceptance of life-long rectal surveillance. Patients with many rectal polyps and/or a family history of severe polyposis should be offered a restorative proctocolectomy with an ileal pouch-anal anastomosis. PMID- 11113067 TI - Safety and efficacy of recombinant human interleukin 10 in chronic active Crohn's disease. Crohn's Disease IL-10 Cooperative Study Group. AB - BACKGROUND & AIMS: Interleukin (IL)-10 is a cytokine with potent anti inflammatory properties. We investigated the safety and efficacy of different doses of human recombinant (rhu)IL-10 in patients with Crohn's disease (CD). METHODS: A prospective, multicenter, double-blind, placebo-controlled study was conducted in 329 therapy-refractory patients with CD. Clinical improvement was defined by a reduction of the Crohn's Disease Activity Index (CDAI) by 100 points or more and clinical remission by a decrease of the CDAI to <150 points. At selected centers, patients underwent ileocolonoscopies and activation of the nuclear factor-kappa B (NF-kappa B) system was assessed in biopsy specimens. RESULTS: Subcutaneous treatment with rhuIL-10 over 28 days induced a fully reversible, dose-dependent decrease in hemoglobin and thrombocyte counts but no clinically significant side effects. No differences in the induction of remission were observed between rhuIL-10 groups (1 microg, 18% [9.6-29.2]; 4 microg, 20% [11.3-32.2]; 8 microg, 20% [11.1-31.8]; 20 microg, 28% [18-40.7]; and placebo, 18% [9.6-29.6]). Clinical improvement was observed in 46% (33.7-59) in the 8 microg/kg rhuIL-10 group in comparison with 27% (17-39.6) in patients taking placebo. Responders to rhuIL-10 showed inhibition of NF-kappaB p65 activation in contrast to nonresponders. CONCLUSIONS: Up to 8 microg/kg of rhuIL-10 was well tolerated. A tendency toward clinical improvement but not remission was observed in the 8-microg/kg dose group. Further studies should delineate which subgroups of patients with CD benefit from rhuIL-10 therapy. PMID- 11113068 TI - Recombinant human interleukin 10 in the treatment of patients with mild to moderately active Crohn's disease. The Interleukin 10 Inflammatory Bowel Disease Cooperative Study Group. AB - BACKGROUND & AIMS: Interleukin 10 (IL-10) is an anti-inflammatory, immunomodulatory cytokine that regulates mucosal inflammation. This study evaluated the safety, tolerance, and efficacy of recombinant human IL-10 (rhuIL 10) for mild to moderately active Crohn's disease. METHODS: We conducted a 24 week multicenter, prospective, randomized, double-blind, placebo-controlled, and sequential-escalating-dose study. Ninety-five patients with Crohn's Disease Activity Index of 200-350, not presently undergoing corticosteroid, mesalamine, or immunosuppressive therapy, were treated with subcutaneous rhuIL-10 (1, 5, 10, or 20 microg/kg) or placebo once daily for 28 consecutive days. Patients were followed up for 20 weeks after treatment. Evaluation of safety and tolerance was the first objective, and efficacy was the second objective. RESULTS: Adverse effects were dose-related, mild-to-moderate in severity, and reversible. Asymptomatic and reversible anemia and thrombocytopenia were observed at higher doses. No withdrawal or delayed adverse effects were evident during 20 weeks of follow-up. At the end of treatment (day 29), intent-to-treat analysis showed that 23.5% (confidence interval [CI], 6.8%-49.9%) of patients receiving 5 micro/kg rhuIL-10 experienced clinical remission and endoscopic improvement; 0% (CI, 0% 14.8%) of patients in the placebo group did. Higher doses of recombinant human IL 10 were less effective than 5 microg/kg. No rhuIL-10 serum accumulation and no antibody against IL-10 were detected after 4 weeks. CONCLUSIONS: Subcutaneous rhuIL-10 administered daily for 28 days to patients with mild to moderately active Crohn's disease is safe, well-tolerated, and shows clinical and endoscopic improvement. PMID- 11113069 TI - Linkage heterogeneity for the IBD1 locus in Crohn's disease pedigrees by disease onset and severity. AB - BACKGROUND & AIMS: There is evidence for the IBD1 Crohn's disease (CD) susceptibility locus on chromosome 16 in several but not all populations studied. Genetic and phenotypic heterogeneity may underlie ability to replicate IBD1. We determined if age and severity stratification could identify a clinical subgroup at risk for IBD1. METHODS: Linkage analysis at microsatellites spanning chromosome 16 was performed in 2 groups of CD pedigrees: group 1, 57 pedigrees with at least one affected relative classified as having "severe" disease, by history of surgical resection or immunomodulator therapy, and with disease diagnosed before age 22; and group 2, 33 pedigrees with no history of early onset, severe CD. RESULTS: Group 1 pedigrees demonstrated genomewide significant linkage evidence for the IBD1 locus (nonparametric multipoint logarithm of the odds [Mlod], 3.84; P = 1.3 x 10(-5)) with linkage evidence greater than all 90 pedigrees (Mlod, 2.12; P = 9.0 x 10(-4)). Group 2 pedigrees had near zero nonparametric 2-point and Mlod scores for the IBD1 region. Heterogeneity between groups 1 and 2 was significant (P = 0.002). CONCLUSIONS: Presence of early-onset, more severe CD identifies pedigrees at high risk for IBD1. These pedigrees will have more power to refine the IBD1 locus and identify the causative gene. PMID- 11113070 TI - Susceptibility to severe ulcerative colitis is associated with polymorphism in the central MHC gene IKBL. AB - BACKGROUND & AIMS: IKBL gene lies telomeric of the tumor necrosis factor cluster in the central major histocompatibility complex and carries a structural polymorphism at position +738. In the Spanish white population, we found the IKBL+738(C) allele in haplotypes carrying either HLA-DRB1(*)1501 or -DRB1(*)0103. Because these HLA class II alleles may confer susceptibility to ulcerative colitis, we investigated an association between IKBL+738(C) and this disease. METHODS: DNA-based techniques were used to type individual alleles of HLA-DRB1 and IKBL+738. The frequencies of these alleles were compared among ethnically matched populations comprising 155 patients and 298 controls. RESULTS: IKBL+738(C) allele was exclusively increased in patients with extensive and/or intractable disease. HLA-DRB1(*)0103 was the only HLA-DRB1 allele to be significantly increased in frequency in patients with UC compared with controls. It was found in patients with extensive and distal disease. In the HLA DRB1(*)0103-negative population, patients with extensive disease still had a significant association with IKBL+738(C). The difference between the 2 groups of patients was statistically significant (13.7% vs. 1.7% in patients with distal disease; odds ratio, 9.25; P = 0.01). CONCLUSIONS: HLA-DRB1(*)0103 is associated with susceptibility to ulcerative colitis, and IKBL+738(C) marks a propensity to extensive and more severe disease. PMID- 11113071 TI - Postabsorptive plasma citrulline concentration is a marker of absorptive enterocyte mass and intestinal failure in humans. AB - BACKGROUND & AIMS: No blood marker assessing the functional absorptive bowel length has been identified. Plasma citrulline, a nonprotein amino acid produced by intestinal mucosa, is one candidate. We tested this hypothesis in adult patients with the short-bowel syndrome, whose condition can lead to intestinal failure. METHODS: In 57 patients, after a minimal follow-up of 2 years subsequent to final digestive circuit modification, postabsorptive citrulline concentration was measured and parenteral nutrition dependence was used to define permanent (n = 37) and transient (n = 20) intestinal failure. Absorptive function, studied over a 3-day period, was evaluated by net digestive absorption for protein and fat (n = 51). Relations between quantitative values were assessed by linear regression analysis and cutoff citrulline threshold, for a diagnosis of intestinal failure by linear discriminant analysis. Cox model was used to compare citrulline threshold and anatomic variables of the short bowel as indicators of transient as opposed to permanent intestinal failure. RESULTS: In patients with short-bowel syndrome, citrulline levels were lower than in controls (n = 51): 20 +/- 13 vs. 40 +/- 10 micromol/L (mean +/- SD), respectively (P < 0.001). After multivariate analysis, citrullinemia was correlated to small bowel length (P < 0.0001, r = 0.86) and to net digestive absorption of fat, but to neither body mass index nor creatinine clearance. A 20-micromol/L threshold citrullinemia, (1) classified short bowel patients with permanent intestinal failure with high sensitivity (92%), specificity (90%), positive predictive value (95%), and negative value (86%); and (2) was a more reliable indicator (odds ratio, 20.0; 95% confidence interval, 1.9-206.1) than anatomic variables (odds ratio, 2.9; 95% confidence interval, 0. 5-15.8) to separate transient as opposed to permanent intestinal failure. CONCLUSIONS: In patients with short-bowel syndrome, postabsorptive plasma citrulline concentration is a marker of functional absorptive bowel length and, past the 2-year adaptive period, a powerful independent indicator allowing distinction of transient from permanent intestinal failure. PMID- 11113072 TI - Congenital sodium diarrhea is an autosomal recessive disorder of sodium/proton exchange but unrelated to known candidate genes. AB - BACKGROUND & AIMS: Congenital sodium diarrhea (CSD) is caused by defective sodium/proton exchange with only 6 sporadic cases reported. The genetics of the disease have not been established. We studied 5 infants with secretory diarrhea, identified in a circumscribed rural area in Austria, to define the mode of transmission and the involvement of candidate genes known to encode for sodium/proton exchangers (NHEs). METHODS: We collected clinical and laboratory data from 5 affected patients, analyzed the pedigrees of their families, and performed homozygosity mapping and multipoint linkage analysis studies in 4 candidate regions known to contain NHE genes. RESULTS: The diagnosis of CSD in 4 of 5 patients was based on daily fecal sodium excretion between 98 and 190 mmol/L, hyponatremia, metabolic acidosis, and low-to-normal urinary sodium concentrations. Pedigree analysis of the affected 2 CSD families revealed parental consanguinity and a common single ancestor 5 generations ago. Homozygosity mapping and/or multipoint linkage analysis excluded the NHE1 locus on chromosome 1, NHE2 locus on chromosome 2, NHE3 locus on chromosome 5, and NHE5 locus on chromosome 16 as potential candidate genes for CSD in this pedigree. Results on NHE4 were inconclusive because the precise chromosomal location of this NHE gene in humans is currently unknown. CONCLUSIONS: Our data indicate that CSD is an autosomal recessive disorder but is not related to mutations in the NHE1, NHE2, NHE3, and NHE5 genes encoding for currently known sodium/proton exchangers. PMID- 11113073 TI - Interleukin 18 is a potent proliferative factor for intestinal mucosal lymphocytes in Crohn's disease. AB - BACKGROUND & AIMS: Crohn's disease (CD) is characterized by a marked accumulation of activated Th1 type CD4(+) T cells and macrophages in inflamed intestinal mucosa. Interleukin (IL)-18 is a recently described cytokine that mainly exists in activated macrophages and shares biological activities with IL-12 in driving the development of Th1 type CD4(+) T cells by inducing interferon gamma. To clarify the role of IL-18 in intestinal inflammation in CD, we assessed the functional role of IL-18 in regulating intestinal mucosal lymphocytes. METHODS: Serum IL-18 concentration was measured by enzyme-linked immunosorbent assay. Expression of IL-18 and IL-18 receptor in human intestinal mucosa was determined using immunohistochemistry and flow cytometry. The functional activity of IL-18 was assessed by the use of recombinant IL-18 to stimulate both the growth of intestinal mucosal lymphocytes and IL-2 receptor induction activity. RESULTS: The serum IL-18 concentration was significantly higher in patients with CD than normal controls. In the inflamed colonic mucosa of CD, many IL-18(+)CD68(+) macrophages had infiltrated the lamina propria. Intestinal mucosal lymphocytes from CD expressed functional IL-18 receptors. Recombinant IL-18 induced significant proliferative responses in freshly isolated mucosal lymphocytes from CD patients, but not from normal controls. IL-18 up-regulated IL-2 receptor expression in mucosal lymphocytes from patients with CD, but not from normal controls. CONCLUSIONS: These findings suggest that infiltrated macrophages in the inflamed intestinal mucosa in CD produce IL-18, and that macrophage-derived IL-18 may serve as a potent regulatory factor for intestinal mucosal lymphocytes, thereby contributing to chronic intestinal inflammation in CD. PMID- 11113074 TI - Modulation of barrier function during Fas-mediated apoptosis in human intestinal epithelial cells. AB - BACKGROUND & AIMS: Intestinal epithelial cell apoptosis occurs continually without apparent permeability defects and is increased in response to intestinal inflammation. We hypothesized that increased, immune-mediated apoptosis during inflammation might result in barrier dysfunction of the epithelium. METHODS: T84 cells were cultured as a polarized monolayer and exposed to agonist antibody to Fas. Barrier function was assessed by transepithelial resistance and permeability measurements. Immunofluorescent staining was used to examine junctional protein expression. RESULTS: Fas expression is predominantly basolateral in polarized T84 monolayers. Basolateral cross-linking of the Fas receptor resulted in T84 cell apoptosis and a loss of 50% of the cells within 24 hours. Apoptosis was coincident with a decrease in transepithelial electrical resistance and increased flux of small but not large molecules. Preservation of barrier function was associated with dramatic rearrangement of tight junctions and desmosomal junctions in apoptotic monolayers. E-cadherin-mediated cell contact was maintained between intact cells in the monolayer, thus sealing gaps created by apoptotic cells. Apoptosis and barrier dysfunction could be prevented by caspase inhibition. CONCLUSIONS: Immune-mediated apoptosis of intestinal epithelial cells may contribute to the permeability defects associated with inflammatory conditions of the bowel, but the intestinal epithelium is remarkably resilient in the face of apoptosis. PMID- 11113075 TI - Drug enterocyte adducts: possible causal factor for diclofenac enteropathy in rats. AB - BACKGROUND & AIMS: Enteropathy is a frequent complication of diclofenac and other nonsteroidal anti-inflammatory drugs, yet little is known about the underlying mechanism. One possibility is that reactive metabolites of diclofenac form adducts with enterocyte macromolecules, as previously shown for liver. We addressed this possibility by using immunohistochemistry to detect diclofenac adducts. METHODS: Rats were treated orally with diclofenac (10-100 mg/kg) and killed after 1-24 hours, and their gastrointestinal (GI) tracts were evaluated for ulcer number and area. Adduct distribution and intensity were assessed by immunohistochemistry by using a technique to simultaneously process and stain multiple intestinal rings. RESULTS: Drug treatment led to dose-dependent formation of both adducts and ulcers only in small intestine and only in animals with intact enterohepatic circulation. Adducts formed within enterocytes by 1 hour, translocated to the brush border, preceded ulceration and vascular protein leakage, and were intense at sites of ulceration. Adducts and ulcers exhibited a parallel distribution within intestinal quintiles: 3rd > 5th >> 1st. CONCLUSIONS: Diclofenac treatment resulted in the formation of drug adducts in enterocytes. Because this molecular change occurred before ulceration, was dose dependent, and exhibited concordant distribution with extent of ulceration, the results suggest a causal role for drug adduct formation in diclofenac enteropathy. PMID- 11113076 TI - Uptake and presentation of antigen to T cells by primary colonic epithelial cells in normal and diseased states. AB - BACKGROUND & AIMS: The immunoregulatory properties of primary colonic epithelial cells (CECs) have not been defined. The ability of CECs from wild-type and interleukin 2-deficient (IL-2(-/-)) mice to take up a complex protein antigen and present peptides via MHC molecules to T cells was assessed and contrasted with that of primary small intestinal epithelial cells (SIECs). METHODS: Uptake of fluorescein isothiocyanate (FITC)-labeled ovalbumin (FITC-OVA) by CECs and SIECs from wild-type and IL-2(-/-) mice was measured by flow cytometry. The effect of disrupting cytoskeleton organization and metabolic activity of CEC on antigen uptake was assessed. An OVA/I-A(b)-specific CD4(+) T-cell line transfected with an NFAT-lacZ reporter gene construct was used to evaluate the ability of CECs and SIECs as well as CECs from healthy and colitic IL-2(-/-) mice to present antigen to T cells. RESULTS: Uptake of FITC-OVA by CECs is concentration dependent, is not saturated at physiologic concentrations, and requires metabolically active cells. CECs from IL-2(-/-) mice take up significantly more antigen than those from wild-type mice. CECs are more efficient APCs than SIECs, and antigen-pulsed CECs from IL-2(-/-) mice induce the highest levels of T-cell activation. CONCLUSIONS: Primary CECs are efficient APCs for CD4 MHC class II-restricted T cells. Antigen uptake and presentation is up-regulated in animals prone to develop intestinal inflammation. PMID- 11113077 TI - Adrenal cortical activation in murine colitis. AB - BACKGROUND & AIMS: Proper adrenal glucocorticoid secretion is crucial in the course of inflammatory diseases. However, the function and structure of the adrenal glands have not been examined in inflammatory bowel diseases. METHODS: After induction of trinitrobenzene sulfonic acid (TNBS) colitis in SJL/J mice, plasma hormone and cytokine levels were measured, adrenal structure was analyzed by immunohistochemistry and electron microscopy, and adrenal cytokine/cytokine receptor expression were studied by RNase protection. RESULTS: Adrenals of colitic animals were enlarged and hypervascularized. These animals had a marked increase in plasma corticosterone levels during the course of colitis (270 +/- 34 vs. 16 +/- 11 ng/mL; P < 0.0001) but only a modest elevation of their concurrent adrenocorticotropin levels (57 +/- 13 vs. 29 +/- 9 pmol/L; NS). On electron microscopy, adrenocortical cells showed ultrastructural signs of marked stimulation, and intra-adrenal lymphocytes were frequently found in direct contact with these cells. Concurrent plasma levels of interleukin (IL)-6, the major cytokine activating the hypothalamic-pituitary-adrenal axis, were markedly increased (495 +/- 131 vs. 20 +/- 1.5 pg/mL; P < 0.0001), and this cytokine directly stimulated corticosterone secretion by adrenocortical cells in vitro. Intra-adrenal expression of IL-6 in animals with colitis was increased 80-fold, and the IL-6 receptor subunits IL-6R alpha and gp130 were present in the adrenal cells. Treatment of animals with neutralizing anti-IL-6 antibody reduced the TNBS induced growth and activation of the adrenal cortices. CONCLUSIONS: Colitis is associated with a profound stimulation of adrenocortical cell function and glucocorticoid release. Direct immune-adrenal interactions seem to contribute to this activation of the adrenal glands during colitis. PMID- 11113078 TI - Peripheral corticotropin-releasing factor and stress-stimulated colonic motor activity involve type 1 receptor in rats. AB - BACKGROUND & AIMS: Corticotropin-releasing factor (CRF) exerts its action through CRF receptors 1 and 2 (CRF-R1 and CRF-R2). CRF has preferential affinity for CRF R1, whereas urocortin displays high affinity for both. We investigated changes in colonic motor function after intraperitoneal (IP) injection of CRF-related peptides. METHODS: Colonic motility was recorded in vivo in conscious rats equipped with electrodes chronically implanted in the cecum and proximal colon or in vitro in distal colon; fecal output was monitored in naive rats. RESULTS: Rat CRF, rat urocortin, and amphibian sauvagine (10 microg/kg, IP) induced a new pattern of cecocolonic myoelectric activity characterized by clustered spike bursts of long duration; the percentage of occurrence was highest after CRF. The rank order of potency to increase fecal pellet output after IP peptide injection (0.3-10 microg/kg, IP) was CRF > urocortin = sauvagine. The CRF-R1/R2 antagonist astressin (33 microg/kg, IP) and the CRF-R1 antagonist CP-154,526 (20 mg/kg, subcutaneously) inhibited IP CRF-induced changes in cecocolonic myoelectric activity and IP CRF- and water avoidance stress-induced fecal output. In vitro, CRF injected into the inferior mesenteric artery increased distal colonic myoelectric activity compared with saline injection. CONCLUSIONS: These results demonstrate that CRF acts peripherally to stimulate colonic motility and that CRF R1 is primarily involved in mediating IP CRF/urocortin- and water avoidance stress-induced colonic motor response. PMID- 11113079 TI - Prostaglandin EP receptor subtypes have distinctive effects on jejunal afferent sensitivity in the rat. AB - BACKGROUND & AIMS: Tissue levels of prostaglandin (PG) E(2) are increased in inflammatory bowel disease. The aim of this study was to characterize the potential for PGE(2) to modulate the sensitivity of intestinal afferents. METHODS: Electrophysiologic recordings were obtained from mesenteric afferent supplying the proximal jejunum of anesthetized rats. RESULTS: PGE(2) evoked a dose-dependent increase in afferent nerve discharge that was biphasic at higher doses. An early response phase, peak discharge frequency of 165.4 +/- 14.3 imp. s(-1), and duration of 20.2 +/- 1.2 seconds were followed by a plateau of elevated afferent nerve discharge lasting several minutes. The increase in afferent nerve discharge was accompanied by an increase in intestinal pressure of 4.4 +/- 0.5 cm H(2)O. Nifedipine (1 mg. kg(-1)) attenuated the pressure response and the plateau phase of afferent discharge, whereas the early component remained unchanged. In contrast, the early phase, but not the plateau phase, was reduced by luminal anesthetic. Experiments with EP receptor-selective agonists and the EP(1)-receptor antagonist AH-6809 (500 microg. kg(-1)) implicate EP1 receptors in the early response, and EP(2) receptors appeared to play a major role in the plateau phase. CONCLUSIONS: PGE(2) has complex actions on intestinal afferent discharge acting by direct and indirect mechanisms and mediated by different receptor subtypes. PMID- 11113080 TI - Interstitial cells of cajal and inflammation-induced motor dysfunction in the mouse small intestine. AB - BACKGROUND & AIMS: Interstitial cells of Cajal (ICC) play an important role in the control of gastrointestinal motility. We aimed to determine a potential role for ICC in the pathophysiology of inflammation-induced motor disorders. METHODS: Effects of Trichinella spiralis infection on electrical pacemaker activity, the structure of ICC associated with Auerbach's plexus, and in vivo motor patterns were studied in the mouse small intestine. RESULTS: Between days 1 and 15 after infection, structural damage occurred in the network of ICC, particularly in the processes connecting ICC to each other and to smooth muscle cells. This was associated with desynchronization of electrical pacemaker activity. Abnormal slow wave activity occurred, including doubling of frequency and electrical quiescence, leading to the development of ectopic pacemaker activity in vivo. In vivo motor patterns in the small intestine changed from consistent peristaltic contractile activity in control animals to periods of quiescence alternating with both orally and aborally propagating contractile activity in the presence of inflammation. Sixty days after infection, all parameters studied had returned to normal values. CONCLUSIONS: Inflammation-induced alterations in the network of ICC of the small intestine associated with Auerbach's plexus lead to disorganization of motor patterns. Because of the strong temporal correlation between damage to the ICC network, electrical uncoupling, the appearance of ectopic pacemaker activity, and the occurrence of retrograde peristalsis, it is concluded that ICC can play a major role in inflammation-induced motor disturbances. PMID- 11113081 TI - Lower esophageal sphincter relaxation and activation of medullary neurons by subdiaphragmatic vagal stimulation in the mouse. AB - BACKGROUND & AIMS: Isolated lower esophageal sphincter (LES) relaxation associated with belching and vomiting and the transient LES relaxation associated with gastroesophageal reflux are gastric afferent-mediated vagovagal reflexes. We aimed to identify the brain stem vagal subnuclei involved in these reflexes. METHODS: In anesthetized mice, LES pressures were recorded using a manometric technique and response to electrical stimulation of the ventral trunk of subdiaphragmatic vagus was investigated. Anatomy of the vagal subnuclei was defined, and activated subnuclei with ventral subdiaphragmatic vagus stimulation were detected by c-fos immunohistochemical staining. RESULTS: Ventral subdiaphragmatic vagal stimulation elicited frequency-dependent LES relaxation without evoking esophageal contractions and induced c-fos expression in interneurons in medial, dorsomedial, and commissural subnuclei along with outer shell of area postrema and motoneurons in the caudal dorsal motor nucleus of vagus. Brain stem subnuclei including interstitial, intermediate, and central subnuclei, and nucleus ambiguous, which have been reported to be involved in the response to swallowing, were not activated. CONCLUSIONS: Stimulation of the ventral subdiaphragmatic vagus causes isolated LES relaxation and activates neurons in select vagal subnuclei that may represent the brain stem circuit involved in the abdominal vagal-afferent-evoked isolated LES relaxation. These observations suggest that different brain stem circuits are involved in swallow induced and gastric afferent-mediated isolated LES relaxations. PMID- 11113082 TI - Neutrophil chemoattractant 2 beta regulates expression of the Reg gene in injured gastric mucosa in rats. AB - BACKGROUND & AIMS: Regenerating (Reg) protein has a trophic effect on gastric mucosal cells. We have shown that Reg gene expression is increased in enterochromaffin-like (ECL) cells during the healing of damaged gastric mucosa around mucosal erosion. This study was designed to explore the stimulants of Reg expression during the healing of gastric mucosal damage. METHODS: Time course changes of the expression of genes for various proinflammatory cytokines and Reg were investigated after induction of gastric mucosal lesions in rats. The direct effect of proinflammatory cytokines on Reg gene expression and Reg protein production were investigated in vitro using counterflow elutriation-enriched rat ECL cells. CXC receptor 2 (CXCR-2) expression was investigated in ECL cells by reverse-transcription polymerase chain reaction. Reg gene expression was also investigated in rats treated by the neutralizing antibody of cytokine-induced neutrophil chemoattractant (CINC-2 beta). RESULTS: During healing, the gene expression of several proinflammatory cytokines and Reg was markedly augmented. Among the proinflammatory cytokines, CINC-2 beta is the only cytokine in which augmented expression preceded the increase of Reg gene expression. In rats treated with CINC-2 beta neutralizing antibody, the augmentation of Reg gene expression was significantly inhibited. When ECL cells were incubated with these proinflammatory cytokines, CINC-2 beta dose-dependently increased Reg messenger RNA and Reg protein in ECL cells. CXCR-2 was identified in isolated ECL cells. CONCLUSIONS: CINC-2 beta, expressed in damaged gastric mucosa, stimulates the production of Reg protein in ECL cells via CXCR-2 and may be involved in the accelerated healing of injured gastric mucosa. PMID- 11113083 TI - Nutritional regulation of nucleoside transporter expression in rat small intestine. AB - BACKGROUND & AIMS: Concentrative nucleoside transporters CNT1 (pyrimidine preferring) and CNT2 (purine preferring) may be involved in the uptake of nucleoside-derived drugs used in antiviral and chemical therapies. The possibility that nucleoside carrier isoform expression is modulated by nutrient availability has been studied. METHODS: CNT1 and CNT2 tissue distribution was determined by Western blot analysis. The effect of 48-hour starvation on CNT expression was then studied. Nucleoside transporter expression and uptake activity were measured in jejunal brush border plasma membrane vesicles from fed and starved rats. The expression of nucleoside transporters was later determined in a second model of nutrient deficiency: rats fed a purified diet with or without nucleotides for 10 days. RESULTS: CNT1 and CNT2 nucleoside transporters were expressed in a wider variety of tissues than expected from messenger RNA distribution analysis. CNT1 was sensitive to nutrient availability in small intestine and, accordingly, jejunal brush border membrane vesicles from 48-hour fasted rats showed increased expression of CNT1 and enhanced Na(+)-dependent thymidine and gemcitabine uptake. This effect was mimicked by feeding semipurified diets lacking nucleotides. CONCLUSIONS: Substrate availability modulates nucleoside transporter expression (CNT1) in rat jejunum in vivo. PMID- 11113084 TI - Epidemiology and natural history of primary biliary cirrhosis in a US community. AB - BACKGROUND & AIMS: The epidemiology of primary biliary cirrhosis (PBC) has not been studied systematically in the United States. We report the incidence and prevalence of this condition in the general population. We also examined the validity of the Mayo natural history model for PBC among these unselected patients from the community. METHODS: The Rochester Epidemiology Project entails a computerized index of diagnoses from the health care encounters of residents of Olmsted County, Minnesota. For potential cases identified using this database, the complete (inpatient and outpatient) medical records were reviewed to verify the diagnosis and extract information necessary for the application of the Mayo model. We estimated the incidence and prevalence of PBC in this population and compared the actual survival of patients with PBC in the community with the survival predicted for PBC patients by the Mayo natural history model. RESULTS: The age-adjusted (to 1990 U.S. whites) incidence of PBC per 100,000 person-years for years 1975-1995 was 4.5 (95% confidence interval [CI], 3.1-5.9) for women, 0.7 (95% CI, 0.1-1.3) for men, and 2.7 (95% CI, 1.9-3.5) overall. The age- and sex-adjusted prevalence per 100,000 persons as of 1995 was 65.4 (95% CI, 43.0 87.9) for women, 12.1 (95% CI, 1.1-23.1) for men, and 40.2 (95% CI, 27.2-53.1) overall. The Mayo natural history model accurately predicted the actual survival of these patients. CONCLUSIONS: This first description of the epidemiology of PBC in the United States indicates that its incidence and prevalence in this country are among the highest reported. Outcomes among these unselected patients from a community population further validated the Mayo natural history model of PBC. PMID- 11113085 TI - Pentoxifylline improves short-term survival in severe acute alcoholic hepatitis: a double-blind, placebo-controlled trial. AB - BACKGROUND & AIMS: An earlier pilot study from our liver unit suggested benefit from treatment with pentoxifylline (PTX), an inhibitor of tumor necrosis factor (TNF), in severe acute alcoholic hepatitis. The aim of the present study was to evaluate this treatment in a larger cohort of patients. METHODS: One hundred one patients with severe alcoholic hepatitis (Maddrey discriminant factor > or = 32) entered a 4-week double-blind randomized trial of PTX (400 mg orally 3 times daily) vs. placebo. Primary endpoints of the study were the effect of PTX on (1) short-term survival and (2) progression to hepatorenal syndrome. On randomization, there were no differences in demographic and clinical characteristics or laboratory values (including TNF) between the 2 groups. RESULTS: Twelve (24.5%) of the 49 patients who received PTX and 24 (46.1%) of the 52 patients who received placebo died during the index hospitalization (P = 0.037; relative risk, 0.59; 95% confidence interval, 0.35-0.97). Hepatorenal syndrome was the cause of death in 6 (50%) and 22 (91.7%) patients (P = 0.009; relative risk, 0.29; 95% confidence interval, 0.13-0.65). Three variables (age, creatinine level on randomization, and treatment with PTX) were independently associated with survival. TNF values on randomization were not predictive of survival; however, during the study period they increased markedly in nonsurvivors compared with survivors in both groups. CONCLUSIONS: Treatment with PTX improves short-term survival in patients with severe alcoholic hepatitis. The benefit appears to be related to a significant decrease in the risk of developing hepatorenal syndrome. Increasing TNF levels during the hospital course are associated with an increase in mortality rate. PMID- 11113086 TI - Two PKR inhibitor HCV proteins correlate with early but not sustained response to interferon. AB - BACKGROUND & AIMS: The NS5A and the E2 proteins of hepatitis C virus (HCV)-1b can bind and inhibit in vitro the interferon (IFN)-induced cellular kinase PKR. The role of such interaction in modulating the antiviral effect of IFN is still controversial. We have analyzed the E2 and the NS5A sequences in HCV-1b-infected patients treated with IFN to assess whether and how different combinations of wild-type and mutant proteins correlated with early and long-term virological response. METHODS: In 30 patients, sequences of pretreatment and on-treatment E2 PePHD and NS5A-PKR binding domain (including the putative ISDR) were analyzed in parallel by sequencing cDNA-polymerase chain reaction products and up to 25 independent clones. RESULTS: The E2-PePHD sequence was highly conserved with a homogeneous quasispecies and was identical in 29 of 30 cases with no association with the pattern of response and no evidence of evolution during therapy. Patients with a mutated NS5A-ISDR had a higher rate of early virological response (67%) than cases with wild-type ISDR (17%). This association was lost in long term responders (33% vs. 17%). CONCLUSIONS: Although the highly conserved E2 PePHD motif might contribute to reduce IFN responsiveness, variations within this region do not seem to play a role in modulating IFN sensitivity. The NS5A-ISDR sequence influenced the early, but not the sustained response, to IFN, suggesting that other factors may be more important for the long-term outcome of therapy. PMID- 11113087 TI - Elevated bound leptin correlates with energy expenditure in cirrhotics. AB - BACKGROUND & AIMS: Leptin, found to be elevated in patients with liver cirrhosis, may contribute to the inadequate energy expenditure and malnutrition associated with a negative prognosis for these patients. Our aim was to characterize leptin components and their relationships to body composition, resting energy expenditure (REE), and substrate use in patients with posthepatic liver cirrhosis. METHODS: Using specific radioimmunoassays, we measured free leptin and bound leptin in 27 cirrhotics and 27 matched control subjects. In the cirrhotic group, body composition and REE were determined. RESULTS: Free leptin was not different in cirrhotics and control subjects and was related to body mass index (controls: r = 0.34, P < 0.05; cirrhotics: r = 0.55, P < 0.005) and to fat mass (cirrhotics: r = 0.76, P < 0.0001). Bound leptin was significantly higher in cirrhotic subjects than in controls (P < 0.001) and was related to REE x fat-free mass(-1) (r = 0.57, P < 0.005) or to the difference between measured and estimated REE (r = 0.55, P < 0.005). CONCLUSIONS: Free leptin reflects fat mass in controls and cirrhotics. Increased serum leptin in cirrhotics is a result of increased bound leptin serum concentrations, which are positively related to energy expenditure. Moreover, bound leptin may be a useful marker for inadequate energy expenditure in patients with liver cirrhosis. PMID- 11113088 TI - Combined interleukin 6 and soluble interleukin 6 receptor accelerates murine liver regeneration. AB - BACKGROUND & AIMS: Liver regeneration after loss of hepatic tissue leads to hepatocyte and nonparenchymal cell proliferation and rapid restoration of liver parenchyma. Interleukin (IL)-6 is a key inducer of transcription factors involved in liver regeneration. Whenever IL-6 activates target cells, it binds to a specific IL-6 receptor (IL-6R). The IL-6/IL-6R complex then associates with the signal transducer gp130, leading to activation of intracellular signaling. METHODS: We have recently constructed the designer cytokine Hyper-IL-6 consisting of soluble IL-6R covalently linked to IL-6, which directly stimulates gp130 even in the absence of membrane-bound IL-6R. We compared the influence of IL-6 and Hyper-IL-6 on liver regeneration after partial hepatectomy in mice. RESULTS: The IL-6/soluble IL-6 fusion protein Hyper-IL-6, but not IL-6 alone, led to an earlier onset of hepatocellular proliferation resulting in an acceleration of liver weight restoration. Also, during liver regeneration, soluble IL-6R levels were increased. CONCLUSIONS: These results emphasize a central role for IL-6 and soluble IL-6R in liver regeneration and indicate a possible therapeutic potential for the designer cytokine Hyper-IL-6 in clinical situations associated with liver regeneration such as acute hepatic failure or resection of chronically damaged liver tissue. PMID- 11113089 TI - Perfused rat intrahepatic bile ducts secrete and absorb water, solute, and ions. AB - BACKGROUND & AIMS: We report a novel approach to study biliary water, bile acid, and HCO(3)(-) transport: the microperfusion of intrahepatic bile duct units (IBDUs) isolated from normal rat liver. METHODS: To study water transport, IBDUs were perfused in vitro with a membrane-impermeant fluorescent volume marker, fluorescein sulfonate; net water movement (J(v)) and osmotic water permeability (P(f)) were then calculated. To study solute transport, IBDUs were perfused with taurocholic acid (TCA) and bile acid uptake was calculated from the concentrations of TCA in the perfused and collected solutions. To study ion transport, IBDUs were perfused with the cell-impermeant pH-sensitive dye BCECF dextran; luminal pH was determined from fluorescence excitation ratios. RESULTS: When inward (secretory) or outward (absorptive) osmotic gradients were established across IBDUs, water movement was observed from bath to lumen (i.e., secretion) and from lumen to bath (i.e., absorption). The perfused IBDUs absorbed TCA in a saturable, sodium-dependent manner; in addition, TCA absorption was blocked in a dose-dependent fashion by S0960, a specific inhibitor of the Na(+)/bile acid cotransporter. Addition of forskolin to HCO(3)(-)-containing (but not HCO(3)(-)-free) bath buffer resulted in lumen alkalinization reflecting HCO(3)(-) transport into the lumen of perfused IBDUs. CONCLUSIONS: The results provide direct functional evidence that intrahepatic bile ducts both secrete and absorb water in response to osmotic gradients, actively absorb bile acid, and transport HCO(3)(-). PMID- 11113090 TI - Estrogens stimulate proliferation of intrahepatic biliary epithelium in rats. AB - BACKGROUND & AIMS: We investigated the expression of estrogen receptor (ER) alpha and beta subtypes in cholangiocytes of normal and bile duct-ligated (BDL) rats and evaluated the role and mechanisms of estrogens in the modulation of cholangiocyte proliferation. METHODS: ER-alpha and ER-beta were analyzed by immunohistochemistry, reverse-transcription polymerase chain reaction, and Western blotting in normal and BDL rats. The effects of the ER antagonists tamoxifen and ICI 182,780 on cholangiocyte proliferation were evaluated. RESULTS: Cholangiocytes expressed both ER-alpha and ER-beta subtypes, whereas hepatocytes expressed only ER-alpha. In association with a marked cholangiocyte proliferation and with enhanced estradiol serum levels, the immunoreactivity for ER-alpha involved a 3-fold higher percentage of cholangiocytes in 3-week BDL than in normal rats; immunoreactivity for ER-beta showed a 30-fold increase. Western blot analysis showed that during BDL, the total amount of ER-beta in cholangiocytes was markedly increased (5-fold), whereas that of ER-alpha decreased slightly ( 25%). Treatment with tamoxifen or ICI 182,780 of 3-week BDL rats inhibited cholangiocyte proliferation and induced overexpression of Fas antigen and apoptosis in cholangiocytes. In vitro, 17 beta estradiol stimulated proliferation of cholangiocyte, an effect blocked to the same extent by tamoxifen or ICI 182,780. CONCLUSIONS: This study suggests that estrogens and their receptors play a role in the modulation of cholangiocyte proliferation. PMID- 11113091 TI - Apical endocytosis in rat hepatocytes In situ involves clathrin, traverses a subapical compartment, and leads to lysosomes. AB - BACKGROUND & AIMS: This study demonstrates and characterizes apical (canalicular) endocytic pathways in hepatocytes in situ. METHODS: Endocytic markers were administered by retrograde infusion through the common bile duct. Colocalization with proteins that are specific for various endocytic compartments was performed on stacks of deconvoluted confocal immunofluorescence images. The subcellular distribution of marker proteins was assessed by electron microscopy (EM). RESULTS: Bulk-phase, as well as membrane-associated markers, were internalized readily at the apical cell pole. At the EM level, marker was found initially in 60-100-nm tubulovesicular structures and 150-200-nm cup-shaped vesicles, whereas multivesicular bodies and lysosomes became labeled after longer time intervals. Apical endocytosis involved clathrin and delivered marker to late endosomes (rab7(+), cathepsin D(+)), as well as lysosomes (rab7(-), cathepsin D(+)). Simultaneous labeling of the basolateral endocytic route resulted in overlap of both pathways in the late endosomal and lysosomal compartments. In addition, apical endocytosis involved a subapical compartment (endolyn-78(+), rab11(+), polymeric IgA receptor [pIgA-R(+)]) that is passed by the transcytotic route, thus constituting a crossroads. pIgA-R immunoreactivity, probably reflecting the cleaved receptor fragment, was associated with apical endocytic marker and colocalized with clathrin and later with cathepsin D. CONCLUSIONS: Apical endocytosis involves coated pits/vesicles, leads to a subapical compartment, and plays a role in the retrieval of canalicular plasma membrane components for lysosomal degradation. PMID- 11113092 TI - Sterol carrier protein 2 gene transfer changes lipid metabolism and enterohepatic sterol circulation in mice. AB - BACKGROUND & AIMS: Sterol carrier protein 2 (SCP-2) enhances sterol cycling and facilitates cholesterol translocation between intracellular organelles and plasma membrane in cultured cells, including hepatocytes. We examined the role of SCP-2 in hepatic cholesterol and lipid trafficking through the sinusoidal and canalicular secretory pathways of the liver in vivo. METHODS: Recombinant adenovirus-mediated SCP-2 gene transfer was used to obtain hepatic overexpression of SCP-2 in C57BL/6 mice. RESULTS: SCP-2 overexpression in the mouse liver resulted in an 8-fold increase of SCP-2 protein levels and determined various effects on lipid metabolism. It decreased high-density lipoprotein cholesterol and increased low-density lipoprotein (LDL) cholesterol concentrations. The expressions of hepatic LDL receptor, apolipoprotein (apo) A-I, apoB, and apoE were decreased. SCP-2 overexpression also increased hepatic cholesterol concentration, associated with decreased cholesterol neosynthesis. Increased biliary cholesterol and bile acid secretion, bile acid pool size, and intestinal cholesterol absorption were also observed. CONCLUSIONS: These results indicate that modulation of SCP-2 expression in the liver determines important modifications on lipoprotein metabolism, hepatic cholesterol synthesis and storage, biliary lipid secretion, bile acid metabolism, and intestinal cholesterol absorption. PMID- 11113093 TI - Correction of liver disease by hepatocyte transplantation in a mouse model of progressive familial intrahepatic cholestasis. AB - BACKGROUND & AIMS: Patients with progressive familial intrahepatic cholestasis (PFIC) type 3 have a mutation in the MDR3 gene, encoding the hepatocanalicular phospholipid translocator. In general, liver failure develops within the first decade of life in these patients. Previous studies have shown that in the mdr2 knockout mouse, the animal model for this disease, the absence of phospholipids in bile causes chronic bile salt-induced damage to hepatocytes. We aimed to test the efficacy of hepatocyte transplantation and liver repopulation in this disease model. METHODS: Transgenic MDR3-expressing hepatocytes as well as normal mdr2(+/+) hepatocytes were transplanted in mdr2(-/-) mice, and liver repopulation was assessed by immunohistochemistry and measurement of biliary lipid secretion. RESULTS: Transplanted hepatocytes partially repopulated the liver, restored phospholipid secretion, and diminished liver pathology. Repopulation was stronger when hepatocellular damage was enhanced by a bile salt-supplemented diet. After 1 year, however, these animals developed multiple hepatic tumors, and biliary phospholipid secretion decreased. In transplanted animals receiving a control diet, repopulation was slower but eventually remained stable at 21%, while liver pathology was completely abrogated and tumor formation was prevented. CONCLUSIONS: These results suggest that moderate liver pathology is a safe condition for the induction of effective hepatocyte repopulation and that this therapy is potentially applicable to patients with PFIC type 3. PMID- 11113094 TI - Regulation of the initiation of pancreatic digestive enzyme protein synthesis by cholecystokinin in rat pancreas in vivo. AB - BACKGROUND & AIMS: Cholecystokinin (CCK) is known to stimulate the synthesis of digestive enzymes in the pancreas at the translational level. We investigated in vivo the biochemical regulation of initiation factors important for the stimulation of translation of digestive enzyme protein in rat pancreas by CCK. METHODS: Intraperitoneal injection of CCK or intragastric administration of a trypsin inhibitor to elicit endogenous CCK release was followed by removal and preparation of pancreas for protein evaluation. Isoelectric focusing was used to evaluate the phosphorylation of the initiation factor eIF4E, and Western blotting and immunoprecipitation followed by Western blotting were used to study the phosphorylation state and amount of other interacting factors. RESULTS: CCK treatment induced a time- and dose-dependent phosphorylation of pancreatic eIF4E and its binding protein (PHAS-I). Because the release of eIF4E from its binding protein as a result of phosphorylation is followed by formation of a messenger RNA cap-binding complex that includes the initiation factor eIF4G, we evaluated the association of eIF4G with released eIF4E and showed that it was increased by CCK. These events occurred over a range of CCK doses from 0.2 to 5 microg/kg. We also evaluated the effect of endogenous CCK by administering a synthetic trypsin inhibitor, camostat (100 mg/kg). Camostat treatment markedly increased the phosphorylation of both PHAS-I and eIF4E and the formation of eIF4E-eIF4G complex. Thus, both exogenous and endogenous CCK activate translational initiation factors in vivo. CONCLUSIONS: Activation of translational machinery necessary for initiation of protein synthesis likely contributes to the normal postprandial synthesis of pancreatic digestive enzymes. PMID- 11113095 TI - Increased intestinal permeability precedes the onset of Crohn's disease in a subject with familial risk. AB - Increased intestinal permeability to several specific molecular probes has been observed in patients with Crohn's disease and their first-degree relatives. A positive family history is also a potent risk factor for inflammatory bowel disease. Although it has been argued that increased permeability in relatives may confer an increased future risk of developing Crohn's disease, long-term follow up of such family members has been lacking. We describe a 24-year-old woman with a positive family history of Crohn's disease who had an elevated gut permeability to (51)Cr-EDTA at age 13, as part of a cross-sectional cohort study in patients and their first-degree relatives. She was asymptomatic at the time, and extensive investigation found no evidence of microscopic or macroscopic Crohn's disease. Repeat investigation because of symptom onset at age 21 revealed ileocolonic Crohn's disease, which required treatment with systemic corticosteroids to induce a clinical remission. In this case, a permeability defect was clearly identified to precede the onset of Crohn's disease in a subject at increased risk. This observation provides support for the hypothesis that increased gut permeability to macromolecules is an early step in the pathogenesis of this disorder. PMID- 11113096 TI - Identical T-cell expansions in the colon mucosa and the synovium of a patient with enterogenic spondyloarthropathy. AB - Intestinal T lymphocytes activated by antigen are suspected to play a key role in enterogenic spondyloarthropathies (SpA). Therefore, we aimed to identify and functionally characterize T-cell clones that are coexpanded in the intestinal mucosa and the synovium. Colon, peripheral blood, and synovium of a patient with enterogenic SpA were screened for clonal T-cell expansions by TCRB-CDR3 length analysis and sequencing. T-cell clones expanded in vivo were isolated from archived synovial cells by targeted T-cell cloning and characterized for phenotype, cytokine production, and antigen specificity. The synovial TCRBV18(+) T-cell repertoire of the patient was dominated by 2 CD8(+) T-cell clones using related CDR3. Both clones were expanded throughout the colon and were present in the peripheral blood. Upon in vitro stimulation with PDB/ionomycin, they showed predominantly interferon gamma and interleukin (IL)-4 but also tumor necrosis factor alpha and IL-10 production and did not specifically lyse autologous T-cell blasts, B-cell lines, or other autologous or allogeneic target or CD1d transfected cells. These findings strongly suggest that T lymphocytes activated by antigen in the intestinal mucosa contribute to joint inflammation in enterogenic SpA by recognition of antigens specific for the inflamed synovium. PMID- 11113097 TI - Genetic counseling and testing for germline p16 mutations in two pancreatic cancer-prone families. AB - The mortality from pancreatic cancer coincides closely with its incidence, indicating a dismal outlook. Hereditary factors probably account for approximately 5%-10% of the pancreatic cancer burden. The molecular genetic etiology of pancreatic cancer is only beginning to be identified. We describe our genetic counseling experience with 2 large families prone to pancreatic cancer malignant melanoma in which p16 (CDKN2) germline mutations had been identified. Members of each family underwent intensive counseling before and at the time of disclosure of p16 germline mutation findings. Two non-cancer-affected siblings from each of the 2 families had p16 mutations identified in DNA from their peripheral blood lymphocytes. In each case, a parent affected with pancreatic cancer also harbored the p16 mutation identified in DNA from their respective tumor blocks. The sibling pairs stated that they would seriously consider prophylactic pancreatectomy if biomarkers or imaging findings suggested a precancerous state. Our experience highlights limited options for managing these families and emphasizes the need for better tools to diagnose pancreatic cancer at a curable stage. PMID- 11113098 TI - American Gastroenterological Association Medical Position Statement: guidelines on constipation. AB - This document presents the official recommendations of the American Gastroenterological Association (AGA) on constipation. It was approved by the Clinical Practice and Practice Economics Committee on March 4, 2000, and by the AGA Governing Board on May 21, 2000. PMID- 11113099 TI - AGA technical review on constipation. American Gastroenterological Association. AB - This literature review and the recommendations therein were prepared for the American Gastroenterological Association Clinical Practice and Practice Economics Committee. The paper was approved by the Committee on March 4, 2000, and by the AGA Governing Board on May 21, 2000. PMID- 11113100 TI - The fall and rise of the hiatal hernia. PMID- 11113101 TI - Recombinant interleukin 10 for the treatment of active Crohn's disease: lessons in biologic therapy. PMID- 11113102 TI - Maintaining a defense as the injured leave the field: apoptosis and barrier function in the intestine. PMID- 11113103 TI - Pentoxifylline and alcoholic hepatitis. PMID- 11113104 TI - Connecting apical endocytosis to the intracellular traffic infrastructure in polarized hepatocytes. PMID- 11113105 TI - Population screening and treatment of Helicobacter pylori. PMID- 11113106 TI - Chemoprevention comes to clinical practice: COX-2 inhibition in familial adenomatous polyposis. PMID- 11113107 TI - MARS: a new treatment for hepatorenal failure. Molecular adsorbent and recirculating system. PMID- 11113109 TI - Gastrointestinal disease in primary care PMID- 11113108 TI - Hepatogenous diabetes: reduced insulin sensitivity and increased awareness. PMID- 11113110 TI - An acknowledgment PMID- 11113111 TI - Proofreading of DNA polymerase eta-dependent replication errors. AB - Human DNA polymerase eta, the product of the skin cancer susceptibility gene XPV, bypasses UV photoproducts in template DNA that block synthesis by other DNA polymerases. Pol eta lacks an intrinsic proofreading exonuclease and copies DNA with low fidelity, such that pol eta errors could contribute to mutagenesis unless they are corrected. Here we provide evidence that pol eta can compete with other human polymerases during replication of duplex DNA, and in so doing it lowers replication fidelity. However, we show that pol eta has low processivity and extends mismatched primer termini less efficiently than matched termini. These properties could provide an opportunity for extrinsic exonuclease(s) to proofread pol eta-induced replication errors. When we tested this hypothesis during replication in human cell extracts, pol eta-induced replication infidelity was found to be modulated by changing the dNTP concentration and to be enhanced by adding dGMP to a replication reaction. Both effects are classical hallmarks of exonucleolytic proofreading. Thus, pol eta is ideally suited for its role in reducing UV-induced mutagenesis and skin cancer risk, in that its relaxed base selectivity may facilitate efficient bypass of UV photoproducts, while subsequent proofreading by extrinsic exonuclease(s) may reduce its mutagenic potential. PMID- 11113112 TI - vCLAP, a caspase-recruitment domain-containing protein of equine Herpesvirus-2, persistently activates the Ikappa B kinases through oligomerization of IKKgamma. AB - vCLAP, the E10 gene product of equine herpesvirus-2, is a caspase-recruitment domain (CARD)-containing protein that has been shown to induce both apoptosis and NF-kappaB activation in mammalian cells. vCLAP has a cellular counterpart, Bcl10/cCLAP, which is also an activator of apoptosis and NF-kappaB. Recent studies demonstrated that vCLAP activates NF-kappaB through an IkappaB kinase (IKK)-dependent pathway, but the underlying mechanism remains unknown. In this report, we demonstrate that vCLAP associates stably with the IKK complex through direct binding to the C-terminal region of IKKgamma. Consistent with this finding, IKKgamma was found to be essential for vCLAP-induced NF-kappaB activation, and the association between vCLAP and the IKK complex induced persistent activation of the IKKs. Moreover, enforced oligomerization of the isolated C-terminal region of vCLAP, which interacts with IKKgamma, can trigger NF-kappaB activation. Finally, substitution of the C-terminal region of IKKgamma, which interacts with vCLAP, with the CARD of vCLAP or Bcl10 produced a molecule that was able to activate NF-kappaB when ectopically expressed in IKKgamma deficient cells. These data suggest that vCLAP-induced oligomerization of IKKgamma, which is mediated by the CARD of vCLAP, could be the mechanism by which vCLAP induces activation of NF-kappaB. PMID- 11113113 TI - Novel role of phosphatidylinositol 3-kinase in CD28-mediated costimulation. AB - Ligation of the CD28 surface receptor provides a major costimulatory signal for full scale T cell activation. Despite extensive studies, the intracellular signaling pathways delivered by CD28 ligation are not fully understood. A particularly controversial matter is the role of phosphatidylinositol 3-kinase (PI3K) in CD28-mediated costimulation. It is known that the binding site for PI3K and Grb-2 lies nested within the YMNM motif of the CD28 cytoplasmic domain. To elucidate the role of PI3K during CD28-mediated interleukin-2 (IL-2) production, CD28 YMNM point and deletion mutants were expressed in Jurkat cells. We then measured IL-2 promoter activation after CD28 ligation. Our results showed that the Y189F mutant, which disrupts binding by PI3K, and the YMNM deletion mutant both demonstrated reduced but significant activity for IL-2 promoter activation. In contrast, the N191A mutant, which retains PI3K binding ability, resulted in a complete abrogation of activity, suggesting that PI3K mediates a negative effect upon transcriptional activation of the IL-2 gene. Consistent with this idea, we found that the addition of a PI3K pharmacological inhibitor augmented IL-2 promoter activity, whereas coexpression of a constitutively active form of PI3K reduced this activity. Taken together, these data indicate that PI3K, when associated with the YMNM motif, may act as a negative mediator in CD28-mediated IL-2 gene transcription. PMID- 11113114 TI - Differential regulation of alternatively spliced endothelial cell myosin light chain kinase isoforms by p60(Src). AB - The Ca(2+)/calmodulin-dependent endothelial cell myosin light chain kinase (MLCK) triggers actomyosin contraction essential for vascular barrier regulation and leukocyte diapedesis. Two high molecular weight MLCK splice variants, EC MLCK-1 and EC MLCK-2 (210-214 kDa), in human endothelium are identical except for a deleted single exon in MLCK-2 encoding a 69-amino acid stretch (amino acids 436 505) that contains potentially important consensus sites for phosphorylation by p60(Src) kinase (Lazar, V., and Garcia, J. G. (1999) Genomics 57, 256-267). We have now found that both recombinant EC MLCK splice variants exhibit comparable enzymatic activities but a 2-fold reduction of V(max), and a 2-fold increase in K(0.5 CaM) when compared with the SM MLCK isoform, whereas K(m) was similar in the three isoforms. However, only EC MLCK-1 is readily phosphorylated by purified p60(Src) in vitro, resulting in a 2- to 3-fold increase in EC MLCK-1 enzymatic activity (compared with EC MLCK-2 and SM MLCK). This increased activity of phospho-MLCK-1 was observed over a broad range of submaximal [Ca(2+)] levels with comparable EC(50) [Ca(2+)] for both phosphorylated and unphosphorylated EC MLCK 1. The sites of tyrosine phosphorylation catalyzed by p60(Src) are Tyr(464) and Tyr(471) within the 69-residue stretch deleted in the MLCK-2 splice variant. These results demonstrate for the first time that p60(Src)-mediated tyrosine phosphorylation represents an important mechanism for splice variant-specific regulation of nonmuscle MLCK and vascular cell function. PMID- 11113115 TI - A novel enhancer of the Apaf1 apoptosome involved in cytochrome c-dependent caspase activation and apoptosis. AB - Apaf1/CED4 family members play central roles in apoptosis regulation as activators of caspase family cell death proteases. These proteins contain a nucleotide-binding (NB) self-oligomerization domain and a caspase recruitment domain (CARD). A novel human protein was identified, NAC, that contains an NB domain and CARD. The CARD of NAC interacts selectively with the CARD domain of Apaf1, a caspase-activating protein that couples mitochondria-released cytochrome c (cyt-c) to activation of cytosolic caspases. Cyt-c-mediated activation of caspases in cytosolic extracts and in cells is enhanced by overexpressing NAC and inhibited by reducing NAC using antisense/DNAzymes. Furthermore, association of NAC with Apaf1 is cyt c-inducible, resulting in a mega-complex (>1 MDa) containing both NAC and Apaf1 and correlating with enhanced recruitment and proteolytic processing of pro-caspase-9. NAC also collaborates with Apaf1 in inducing caspase activation and apoptosis in intact cells, whereas fragments of NAC representing only the CARD or NB domain suppress Apaf1-dependent apoptosis induction. NAC expression in vivo is associated with terminal differentiation of short lived cells in epithelia and some other tissues. The ability of NAC to enhance Apaf1-apoptosome function reveals a novel paradigm for apoptosis regulation. PMID- 11113116 TI - A truncated plasminogen activator inhibitor-1 protein induces and inhibits angiostatin (kringles 1-3), a plasminogen cleavage product. AB - Plasminogen activator inhibitor-1 (PAI-1) is a serpin protease inhibitor that binds plasminogen activators (uPA and tPA) at a reactive center loop located at the carboxyl-terminal amino acid residues 320-351. The loop is stretched across the top of the active PAI-1 protein maintaining the molecule in a rigid conformation. In the latent PAI-1 conformation, the reactive center loop is inserted into one of the beta sheets, thus making the reactive center loop unavailable for interaction with the plasminogen activators. We truncated porcine PAI-1 at the amino and carboxyl termini to eliminate the reactive center loop, part of a heparin binding site, and a vitronectin binding site. The region we maintained corresponds to amino acids 80-265 of mature human PAI-1 containing binding sites for vitronectin, heparin (partial), uPA, tPA, fibrin, thrombin, and the helix F region. The interaction of "inactive" PAI-1, rPAI-1(23), with plasminogen and uPA induces the formation of a proteolytic protein with angiostatin properties. Increasing amounts of rPAI-1(23) inhibit the proteolytic angiostatin fragment. Endothelial cells exposed to exogenous rPAI-1(23) exhibit reduced proliferation, reduced tube formation, and 47% apoptotic cells within 48 h. Transfected endothelial cells secreting rPAI-1(23) have a 30% reduction in proliferation, vastly reduced tube formation, and a 50% reduction in cell migration in the presence of VEGF. These two studies show that rPAI-1(23) interactions with uPA and plasminogen can inhibit plasmin by two mechanisms. In one mechanism, rPAI-1(23) cleaves plasmin to form a proteolytic angiostatin-like protein. In a second mechanism, rPAI-1(23) can bind uPA and/or plasminogen to reduce the number of uPA and plasminogen interactions, hence reducing the amount of plasmin that is produced. PMID- 11113117 TI - Cloning and expression of a pig liver taurochenodeoxycholic acid 6alpha hydroxylase (CYP4A21): a novel member of the CYP4A subfamily. AB - A cytochrome P450 expressed in pig liver was cloned by polymerase chain reaction using oligonucleotide primers based on amino acid sequences of the purified taurochenodeoxycholic acid 6alpha-hydroxylase. This enzyme catalyzes a 6alpha hydroxylation of chenodeoxycholic acid, and the product hyocholic acid is considered to be a primary bile acid specific for the pig. The cDNA encodes a protein of 504 amino acids. The primary structure of the porcine taurochenodeoxycholic acid 6alpha-hydroxylase, designated CYP4A21, shows about 75% identity with known members of the CYP4A subfamily in rabbit and man. Transfection of the cDNA for CYP4A21 into COS cells resulted in the synthesis of an enzyme that was recognized by antibodies raised against the purified pig liver enzyme and catalyzed 6alpha-hydroxylation of taurochenodeoxycholic acid. The hitherto known CYP4A enzymes catalyze hydroxylation of fatty acids and prostaglandins and have frequently been referred to as fatty acid hydroxylases. A change in substrate specificity from fatty acids or prostaglandins to a steroid nucleus among CYP4A enzymes is notable. The results of mutagenesis experiments indicate that three amino acid substitutions in a region around position 315 which is highly conserved in all previously known CYP4A and CYP4B enzymes could be involved in the altered catalytic activity of CYP4A21. PMID- 11113118 TI - Transcriptional activation of the rat vesicular monoamine transporter 2 promoter in gastric epithelial cells: regulation by gastrin. AB - Vesicular monoamine transporter 2 is important for the accumulation of monoamine neurotransmitters into synaptic vesicles and histamine transport into secretory vesicles of the enterochromaffin-like cell of the gastric corpus. In this study we have investigated the mechanisms regulating the transcriptional activation of the rat vesicular monoamine transporter 2 (VMAT2) promoter in gastric epithelial cells. Maintenance of basal levels of transcription was dependent on the presence of SP1, cAMP-response element (CRE), and overlapping AP2/SP1 consensus sequences within the region of promoter from -86 to +1 base pairs (bp). Gastrin stimulation increased transcriptional activity, and responsiveness was shown to be dependent on the CRE (-33 to -26 bp) and AP2/SP1 (-61 to -48 bp) consensus sites but independent of the SP1 site at -86 to -81 bp. Gastrin-induced transcription was dependent on the cooperative interaction of an uncharacterized nuclear factor of approximately 23.3 kDa that bound to the putative AP2/SP1 site, CRE-binding protein (CREB), and CREB-binding protein/p300. Gastrin stimulation resulted in the increased binding of phosphorylated CREB to the promoter, but it did not result in the increased binding of the AP2/SP1-binding protein. The gastrin responsiveness of the promoter was shown to be dependent on both the protein kinase C and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase-signaling pathways, which may converge on the AP2/SP1-binding protein. PMID- 11113119 TI - Inotropic stimulation induces cardiac dysfunction in transgenic mice expressing a troponin T (I79N) mutation linked to familial hypertrophic cardiomyopathy. AB - The cardiac troponin T (TnT) I79N mutation has been linked to familial hypertrophic cardiomyopathy and a high incidence of sudden death, despite causing little or no cardiac hypertrophy. In skinned fibers, I79N increased myofilamental calcium sensitivity (Miller, T., Szczesna, D., Housmans, P. R., Zhao, J., deFreitas, F., Gomes, A. V., Culbreath, L., McCue, J., Wang, Y., Xu, Y., Kerrick, W. G., and Potter, J. D. (2001) J. Biol. Chem. 276, 3743-3755). To further study the functional consequences of this mutation, we compared the cardiac performance of transgenic mice expressing either human TnT-I79N or human wild-type TnT. In isolated hearts, cardiac function was different depending on the Ca(2+) concentration of the perfusate; systolic function was significantly increased in Tg-I79N hearts at 0.5 and 1 mmol/liter. At higher Ca(2+) concentrations, systolic function was not different, but diastolic dysfunction became manifest as increased end-diastolic pressure and time to 90% relaxation. In vivo measurements by echocardiography and Doppler confirmed that base-line systolic function was significantly higher in Tg-I79N mice without evidence for diastolic dysfunction. Inotropic stimulation with isoproterenol resulted only in a modest contractile response but caused significant mortality in Tg-I79N mice. Doppler studies ruled out aortic outflow obstruction and were consistent with increased chamber stiffness. We conclude that in vivo, the increased myofilament Ca(2+) sensitivity due to the I79N mutation enhances base-line contractility but leads to cardiac dysfunction during inotropic stimulation. PMID- 11113120 TI - Structure of a sialic acid-activating synthetase, CMP-acylneuraminate synthetase in the presence and absence of CDP. AB - The x-ray crystallographic structure of selenomethionyl cytosine-5'-monophosphate acylneuraminate synthetase (CMP-NeuAc synthetase) from Neisseria meningitidis has been determined at 2.0-A resolution using multiple-wavelength anomalous dispersion phasing, and a second structure, in the presence of the substrate analogue CDP, has been determined at 2.2-A resolution by molecular replacement. This work identifies the active site residues for this class of enzyme for the first time. The detailed interactions between the enzyme and CDP within the mononucleotide-binding pocket are directly observed, and the acylneuraminate binding pocket has also been identified. A model of acylneuraminate bound to CMP NeuAc synthetase has been constructed and provides a structural basis for understanding the mechanism of production of "activated" sialic acids. Sialic acids are key saccharide components on the surface of mammalian cells and can be virulence factors in a variety of bacterial species (e.g. Neisseria, Haemophilus, group B streptococci, etc.). As such, the identification of the bacterial CMP NeuAc synthetase active site can serve as a starting point for rational drug design strategies. PMID- 11113122 TI - Binding of levosimendan, a calcium sensitizer, to cardiac troponin C. AB - Levosimendan is an inodilatory drug that mediates its cardiac effect by the calcium sensitization of contractile proteins. The target protein of levosimendan is cardiac troponin C (cTnC). In the current work, we have studied the interaction of levosimendan with Ca(2+)-saturated cTnC by heteronuclear NMR and small angle x-ray scattering. A specific interaction between levosimendan and the Ca(2+)-loaded regulatory domain of recombinant cTnC(C35S) was observed. The changes in the NMR spectra of the N-domain of full-length cTnC(C35S), due to the binding of levosimendan to the primary site, were indicative of a slow conformational exchange. In contrast, no binding of levosimendan to the regulatory domain of cTnC(A-Cys), where all the cysteine residues are mutated to serine, was detected. Moreover, it was shown that levosimendan was in fast exchange on the NMR time scale with a secondary binding site in the C-domain of both cTnC(C35S) and cTnC(A-Cys). The small angle x-ray scattering experiments confirm the binding of levosimendan to Ca(2+)-saturated cTnC but show no domain domain closure. The experiments were run in the absence of the reducing agent dithiothreitol and the preservative sodium azide (NaN(3)), since we found that levosimendan reacts with these chemicals, commonly used for preparation of NMR protein samples. PMID- 11113121 TI - A role for the homeobox protein Distal-less 3 in the activation of the glycoprotein hormone alpha subunit gene in choriocarcinoma cells. AB - Synthesis and secretion of chorionic gonadotropin in trophoblast cells of the placenta is required for establishment of early pregnancy in primates. Chorionic gonadotropin is a heterodimeric glycoprotein hormone consisting of alpha and beta subunits. Regulation of the alpha subunit gene within the placenta requires an array of cis elements within the 5'-flanking region of the promoter. Within this array of elements, the junctional regulatory element (JRE) putatively binds a placental-specific transcription factor. The aim of our studies was to determine the identity and role of the transcriptional regulator that binds to the JRE in choriocarcinoma cells (JEG3 cells). Mutations within the JRE resulted in reduction in basal expression of an alpha subunit reporter gene, suggesting that the JRE binding factor was necessary for full basal activity. Using electrophoretic mobility shift assays, we determined that the JRE was capable of serving as a homeobox factor-binding site. The homeobox factor, Distal-less 3 (Dlx 3) was found to be expressed in JEG3 cells and in the trophoblast layer of human chorionic villus but not in a gonadotrope cell line that also expresses the alpha subunit gene. Electrophoretic mobility shift assays revealed that recombinant Dlx 3 could bind specifically to the JRE and endogenous Dlx 3 was present in JRE/JEG3 nuclear protein complexes. Overexpression of Dlx 3 resulted in activation of an alpha subunit reporter gene. A JRE mutation resulted in attenuated activation of the alpha subunit reporter via an adjacent cis element, suggesting that JRE/Dlx 3 interactions may facilitate regulation of the alpha subunit gene at sites immediately upstream of the JRE. Our studies support the conclusion that Dlx 3 is a placental-specific transcriptional regulator that binds to the JRE and contributes to expression of the alpha subunit gene in cells of trophoblast origin. PMID- 11113123 TI - Transporters on demand: intrahepatic pools of canalicular ATP binding cassette transporters in rat liver. AB - ABC transporter trafficking in rat liver induced by cAMP or taurocholate and [(35)S]methionine metabolic labeling followed by subcellular fractionation were used to identify and characterize intrahepatic pools of ABC transporters. ABC transporter trafficking induced by cAMP or taurocholate is a physiologic response to a temporal demand for increased bile secretion. Administration of cAMP or taurocholate to rats increased amounts of SPGP, MDR1, and MDR2 in the bile canalicular membrane by 3-fold; these effects abated after 6 h and were insensitive to prior treatment of rats with cycloheximide. Half-lives of ABC transporters were 5 days, which suggests cycling of ABC transporters between canalicular membrane and intrahepatic sites before degradation. In vivo [(35)S]methionine labeling of rats followed by immunoprecipitation of (sister of P-glycoprotein) (SPGP) from subcellular liver fractions revealed a steady state distribution after 20 h of SPGP between canalicular membrane and a combined endosomal fraction. After mobilization of transporters from intrahepatic sites with cAMP or taurocholate, a significant increase in the amount of ABC transporters in canalicular membrane vesicles was observed, whereas the decrease in the combined endosomal fraction remained below detection limits in Western blots. This observation is in accordance with relatively large intracellular ABC transporter pools compared with the amount present in the bile canalicular membrane. Furthermore, trafficking of newly synthesized SPGP through intrahepatic sites was accelerated by additional administration of cAMP but not by taurocholate, indicating two distinct intrahepatic pools. Our data indicate that ABC transporters cycle between the bile canaliculus and at least two large intrahepatic ABC transporter pools, one of which is mobilized to the canalicular membrane by cAMP and the other, by taurocholate. In parallel to regulation of other membrane transporters, we propose that the "cAMP-pool" in hepatocytes corresponds to a recycling endosome, whereas recruitment from the "taurocholate pool" involves a hepatocyte-specific mechanism. PMID- 11113124 TI - The complement regulator factor H binds to the surface protein OspE of Borrelia burgdorferi. AB - Spirochete bacteria of the Borrelia burgdorferi sensu lato complex cause Lyme borreliosis. The three pathogenic subspecies Borrelia garinii, Borrelia afzelii, and Borrelia burgdorferi sensu stricto differ in their disease profiles and susceptibility to complement lysis. We investigated whether complement resistance of Borreliae could be due to acquisition of the main soluble inhibitors of the alternative complement pathway, factor H and the factor H-like protein 1. When exposed to nonimmune EDTA-plasma, the serum-resistant B. afzelii and B. burgdorferi sensu stricto strains bound factor H/factor H-like protein 1 to their surfaces. Assays with radiolabeled proteins showed that factor H bound strongly to the B. burgdorferi sensu stricto strain. To identify factor H ligands on the borrelial surface, we analyzed a panel of outer surface proteins of B. burgdorferi sensu stricto with the surface plasmon resonance technique. The outer surface lipoprotein OspE was identified as a specific ligand for factor H. Using recombinant constructs of factor H, the binding site for OspE was localized to the C-terminal short consensus repeat domains 15-20. Specific binding of factor H to B. burgdorferi sensu stricto OspE may help the pathogen to evade complement attack and phagocytosis. PMID- 11113125 TI - Chromatin assembly enhances binding to the CYP2B1 phenobarbital-responsive unit (PBRU) of nuclear factor-1, which binds simultaneously with constitutive androstane receptor (CAR)/retinoid X receptor (RXR) and enhances CAR/RXR-mediated activation of the PBRU. AB - Phenobarbital induction of CYP2B genes is mediated by a complex phenobarbital responsive enhancer (PBRU), which contains a binding site for nuclear factor-1 (NF-1) flanked by two DR-4 nuclear receptor (NR) binding sites for a heterodimer of constitutive androstane receptor (CAR) and retinoid X receptor (RXR). To examine potential interactions between NF-1 and CAR/RXR, binding of purified recombinant proteins to DNA, or to chromatin assembled using Drosophila embryo extract, was examined. NF-1 and CAR/RXR bound simultaneously and independently to the overlapping NF-1 and NR-1 sites; binding of CAR/RXR to the NR-2 site was modestly increased by NF-1 binding; and CAR/RXR bound to a new site in the PBRU region, designated NR-3. Assembly of plasmid DNA into chromatin using Drosophila extract resulted in linearly phased nucleosomes in the PBRU region. The apparent binding affinity of NF-1 was increased by about 10-fold in assembled chromatin compared with DNA, whereas CAR/RXR binding was decreased. As observed for DNA, however, simultaneous, largely independent, binding to the NF-1 and NR sites was observed. CAR-mediated transactivation of the PBRU in cultured cells of hepatic origin was inhibited by mutations in the NF-1 site, and overexpression of NF-1 increased CAR transactivation in HepG2 cells. These studies demonstrate that NF-1 and CAR/RXR can both bind to the PBRU at the same time and that chromatin assembly increases NF-1 binding, which is consistent with previous in vivo footprinting studies in which the NF-1 site was occupied in untreated animals and the NF-1 and flanking NR sites were occupied after phenobarbital treatment. CAR mediated trans-activation of the PBRU was increased by NF-1, analogous to NF-1 effects on phenobarbital induction in previous transient transfection studies and consistent with mediation of phenobarbital induction by CAR. PMID- 11113126 TI - Point mutations in anthrax protective antigen that block translocation. AB - The protective antigen (PA) moiety of anthrax toxin delivers the toxin's enzymatic moieties to the cytosol of mammalian cells by a mechanism associated with its ability to heptamerize and form a transmembrane pore. Here we report that mutations in Lys-397, Asp-425, or Phe-427 ablate killing of CHO-K1 cells by a cytotoxic PA ligand. These mutations blocked PA's ability to mediate pore formation and translocation in cells but had no effect on its receptor binding, proteolytic activation, or ability to oligomerize and bind the toxin's enzymatic moieties. The mutation-sensitive residues lie in the 2beta(7)-2beta(8) and 2beta(10)-2beta(11) loops of domain 2 and are distant both in primary structure and topography from the 2beta(2)-2beta(3) loop, which is believed to participate in formation of a transmembrane beta-barrel. These results suggest that Lys-397, Asp-425, and Phe-427 participate in conformational rearrangements of a heptameric pore precursor that are necessary for pore formation and translocation. Identification of these residues will aid in elucidating the mechanism of translocation and may be useful in developing therapeutic and prophylactic agents against anthrax. PMID- 11113127 TI - Structure-function analysis of TFII-I. Roles of the N-terminal end, basic region, and I-repeats. AB - The transcription factor TFII-I can bind specifically to several DNA sequence elements and is implicated in both basal and activated transcription. There are four alternatively spliced isoforms of TFII-I, all characterized by the presence of six I-repeats, R1-R6, each containing a potential helix-loop-helix motif implicated in protein-protein interactions. These isoforms exhibit both homomeric and heteromeric interactions that lead to nuclear localization. In this study we mapped two distinct regions in TFII-I that affect its DNA binding. Deletion of either of these regions led to abrogation of DNA binding and transcriptional activation from both the Vbeta and c-fos promoters. The I-repeats, as expected, were capable of mediating homomeric interactions either individually or in combination. Unexpectedly, an additional homomeric interaction domain was found within the N-terminal end of TFII-I that includes a putative leucine zipper motif. These data suggest a model in which TFII-I undergoes regulated homomeric interaction mediated by both the N-terminal end and the I-repeats. PMID- 11113128 TI - Structure/function relationships in OxlT, the oxalate-formate transporter of oxalobacter formigenes. Assignment of transmembrane helix 11 to the translocation pathway. AB - OxlT, the oxalate:formate antiporter of Oxalobacter formigenes, has a lone charged residue, lysine 355 (Lys-355), at the center of transmembrane helix 11 (TM11). Because Lys-355 is the only charged residue in the hydrophobic sector, we tested the hypothesis that lysine 355 contributes to the binding site for the anionic substrate, oxalate. This idea was supported by mutational analysis, which showed that of five variants studied (Lys-355 --> Cys, Gly, Gln, Arg, or Thr), residual function was found for only the K355R derivative, in which catalytic efficiency had fallen 2,600-fold. Further insight came from a study of TM11 single-cysteine mutants, using the impermeant, thiol-specific reagents, carboxyethyl methanethiosulfonate and ethyltrimethylammonium methanethiosulfonate. Of the five reactive positions identified in TM11, four were at the cytoplasmic or periplasmic ends of TM11 (S344C and A345C, and G366C and A370C, respectively), whereas the fifth was at the center of the helix (S359C). Added study with carboxyethyl methanethiosulfonate and ethylsulfonate methylthiosulfonate showed that the attack on S359C could be blocked by the presence of the substrate, oxalate, and that protection could be predicted quantitatively by a kinetic model in which S359C is accessible only in the unliganded form of OxlT. Parallel study showed that the proteoliposomes used in such work contained OxlT of right side-out and inside-out orientations in about equal amounts. Accordingly, full inhibition of S359C by the impermeable methanethiosulfonate-linked probes must reflect an approach from both the cytosolic and periplasmic surfaces of the protein. This, coupled with the finding of substrate protection, leads us to conclude that S359C lies on the translocation pathway through OxlT. Since position 359 and 355 lie on the same helical face, we suggest that Lys-355 also lies on the translocation pathway, consistent with the idea that the essential nature of Lys-355 reflects its role in binding the anionic substrate, oxalate. PMID- 11113129 TI - Evaluation of prototype transmembrane 4 superfamily protein complexes and their relation to lipid rafts. AB - Recent literature suggests that tetraspanin proteins (transmembrane 4 superfamily; TM4SF proteins) may associate with each other and with many other transmembrane proteins to form large complexes that sometimes may be found in lipid rafts. Here we show that prototype complexes of CD9 or CD81 (TM4SF proteins) with alpha(3)beta(1) (an integrin) and complexes of CD63 (a TM4SF protein) with phosphatidylinositol 4-kinase (PtdIns 4-K) may indeed localize within lipid raft-like microdomains, as seen by three different criteria. First, these complexes localize to low density light membrane fractions in sucrose gradients. Second, CD9 and alpha(3) integrin colocalized with ganglioside GM1 as seen by double staining of fixed cells. Third, CD9-alpha3beta1 and CD81 alpha3beta1 complexes were shifted to a higher density upon cholesterol depletion from intact cells or cell lysate. However, CD9-alpha3beta1, CD81-alpha3beta1, and CD63-PtdIns 4-K complex formation itself was not dependent on localization into raftlike lipid microdomains. These complexes did not require cholesterol for stabilization, were maintained within well solubilized dense fractions from sucrose gradients, were stable at 37 degrees C, and were small enough to be included within CL6B gel filtration columns. In summary, prototype TM4SF protein complexes (CD9-alpha3beta1, CD81-alpha3beta1, and CD63-PtdIns 4-K) can be solubilized as discrete units, independent of lipid microdomains, although they do associate with microdomains resembling lipid rafts. PMID- 11113130 TI - Expression and regulation of normal and polymorphic epithelial sodium channel by human lymphocytes. AB - Gene expression, protein expression, and function of amiloride-sensitive sodium channels were examined in human lymphocytes from normal individuals and individuals with Liddle's disease. Using reverse transcriptase polymerase chain reactions, expression of all three cloned epithelial sodium channel (ENaC) subunits was detected in lymphocytes. Polyclonal antibodies to bovine alpha-ENaC bound to the plasma membrane of normal and Liddle's lymphocytes. A quantitative analysis of fluorescence-tagged ENaC antibodies indicated a 2.5-fold greater surface binding of the antibodies to Liddle's lymphocytes compared with normal lymphocytes. The relative binding intensity increased significantly (25%; p < 0.001) for both normal and Liddle's cells after treatment with 40 microM 8-CPT cAMP. Amiloride-sensitive whole cell currents were recorded under basal and cAMP treated conditions for both cell types. Liddle's cells had a 4.5-fold larger inward sodium conductance compared with normal cells. A specific 25% increase in the inward sodium current was observed in normal cells in response to cAMP treatment. Outside-out patches from both cell types under both treatment conditions revealed no obvious differences in the single channel conductance. The P(open) was 4.2 +/- 3.9% for patches from non-Liddle's cells, and 27.7 +/- 5.4% in patches from Liddle's lymphocytes. Biochemical purification of a protein complex, using the same antibodies used for the immunohistochemistry, yielded a functional sodium channel complex that was inhibited by amiloride when reconstituted into lipid vesicles and incorporated into planar lipid bilayers. These four independent methodologies yielded findings consistent with the hypotheses that human lymphocytes express functional, regulatable ENaC and that the mutation responsible for Liddle's disease induces excessive channel expression. PMID- 11113131 TI - Hepcidin, a urinary antimicrobial peptide synthesized in the liver. AB - Cysteine-rich antimicrobial peptides are abundant in animal and plant tissues involved in host defense. In insects, most are synthesized in the fat body, an organ analogous to the liver of vertebrates. From human urine, we characterized a cysteine-rich peptide with three forms differing by amino-terminal truncation, and we named it hepcidin (Hepc) because of its origin in the liver and its antimicrobial properties. Two predominant forms, Hepc20 and Hepc25, contained 20 and 25 amino acid residues with all 8 cysteines connected by intramolecular disulfide bonds. Reverse translation and search of the data bases found homologous liver cDNAs in species from fish to human and a corresponding human genomic sequence on human chromosome 19. The full cDNA by 5' rapid amplification of cDNA ends was 0.4 kilobase pair, in agreement with hepcidin mRNA size on Northern blots. The liver was the predominant site of mRNA expression. The encoded prepropeptide contains 84 amino acids, but only the 20-25-amino acid processed forms were found in urine. Hepcidins exhibited antifungal activity against Candida albicans, Aspergillus fumigatus, and Aspergillus niger and antibacterial activity against Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, and group B Streptococcus. Hepcidin may be a vertebrate counterpart of cysteine-rich antimicrobial peptides produced in the fat body of insects. PMID- 11113132 TI - A new mouse liver-specific gene, encoding a protein homologous to human antimicrobial peptide hepcidin, is overexpressed during iron overload. AB - Considering that the development of hepatic lesions related to iron overload diseases might be a result of abnormally expressed hepatic genes, we searched for new genes up-regulated under the condition of iron excess. By suppressive subtractive hybridization performed between livers from carbonyl iron-overloaded and control mice, we isolated a 225-base pair cDNA. By Northern blot analysis, the corresponding mRNA was confirmed to be overexpressed in livers of experimentally (carbonyl iron and iron-dextran-treated mice) and spontaneously (beta(2)-microglobulin knockout mice) iron-overloaded mice. In addition, beta(2) microglobulin knockout mice fed with a low iron content diet exhibited a decrease of hepatic mRNA expression. The murine full-length cDNA was isolated and was found to encode an 83-amino acid protein presenting a strong homology in its C terminal region to the human antimicrobial peptide hepcidin. In addition, we cloned the corresponding rat and human orthologue cDNAs. Both mouse and human genes named HEPC are constituted of 3 exons and 2 introns and are located on chromosome 7 and 19, respectively, in close proximity to USF2 gene. In mouse and human, HEPC mRNA was predominantly expressed in the liver. During both in vivo and in vitro studies, HEPC mRNA expression was enhanced in mouse hepatocytes under the effect of lipopolysaccharide. Finally, to analyze the intracellular localization of the predicted protein, we used the green fluorescent protein chimera expression vectors. The murine green fluorescent protein-prohepcidin protein was exclusively localized in the nucleus. When the putative nuclear localization signal was deleted, the resulting protein was addressed to the cytoplasm. Taken together, our data strongly suggest that the product of the new liver-specific gene HEPC might play a specific role during iron overload and exhibit additional functions distinct from its antimicrobial activity. PMID- 11113133 TI - Extracellular matrix metalloproteinase 2 levels are regulated by the low density lipoprotein-related scavenger receptor and thrombospondin 2. AB - We have recently shown that the adhesive defect observed in dermal fibroblasts derived from thrombospondin 2 (TSP2)-null mice results from an increase in matrix metalloproteinase 2 (MMP2) levels (Yang, Z., Kyriakides, T. R., and Bornstein, P. (2000) Mol. Biol. Cell 11, 3353-3364). Adhesion was restored by replacement of TSP2 and by inhibitors of MMP2 activity. In pursuing the observation that TSP2 and MMP2 interact, we now demonstrate that this interaction is required for optimal clearance of extracellular MMP2 by fibroblasts. Since TSP2 is known to be endocytosed by the scavenger receptor, low density lipoprotein receptor-related protein (LRP), we determined whether interference with LRP function affected fibroblast adhesion and/or extracellular MMP2 levels. Addition of heparin, which competes for the binding of TSP2 to LRP coreceptor proteoglycans, inhibited adhesion of control but not TSP2-null cells, and a blocking antibody to LRP as well as the LRP inhibitor, receptor-associated protein, also inhibited adhesion and increased MMP2 levels only in control fibroblasts. TSP2 did not inhibit active MMP2 directly and did not inhibit the activation of pro-MMP2. Finally, the internalization of 125I-MMP2 was reduced in TSP2-null compared with control fibroblasts. We propose that clearance of MMP2-TSP2 complexes by LRP is an important mechanism for the regulation of extracellular MMP2 levels in fibroblasts, and perhaps in other cells. Thus, some features of the phenotype of TSP2-null mice, such as abnormal collagen fibrillogenesis, accelerated wound healing, and increased angiogenesis, could result in part from increased MMP2 activity. PMID- 11113134 TI - Amphiphysin 1 binds the cyclin-dependent kinase (cdk) 5 regulatory subunit p35 and is phosphorylated by cdk5 and cdc2. AB - Amphiphysin 1 is a phosphoprotein expressed at high levels in neurons, where it participates in synaptic vesicle endocytosis and neurite outgrowth. It is a substrate for cyclin-dependent kinase (cdk) 5, a member of the cyclin-dependent protein kinase family, which has been functionally linked to neuronal migration and neurite outgrowth via its action on the actin cytoskeleton. The yeast homologue of amphiphysin, Rvs167, functions in endocytosis and actin dynamics, is phosphorylated by the cdk5 homologue Pho85, and binds the Pho85 regulatory subunit Pcl2. We show here that amphiphysin 1 interacts with the cdk5-activating subunit p35 and that this interaction is mediated by the conserved NH2-terminal region of amphiphysin. Amphiphysin 1 colocalizes with p35 in the growth cones of neurons and at actin-rich peripheral lamellipodia in transfected fibroblasts. Amphiphysin is phosphorylated by cdk5 in a region including serines 272, 276, and 285. Amphiphysin 1 is also phosphorylated by the cdc2/cyclin B kinase complex in the same region and undergoes mitotic phosphorylation in dividing cells. These data indicate that phosphorylation by members of the cyclin-dependent kinase family is a conserved property of amphiphysin and suggest that this phosphorylation may play an important physiological role both in mitosis and in differentiated cells. PMID- 11113135 TI - The gene for a variant form of the polyadenylation protein CstF-64 is on chromosome 19 and is expressed in pachytene spermatocytes in mice. AB - Many mRNAs in male germ cells lack the canonical AAUAAA but are normally polyadenylated (Wallace, A. M., Dass, B., Ravnik, S. E., Tonk, V., Jenkins, N. A., Gilbert, D. J., Copeland, N. G., and MacDonald, C. C. (1999) Proc. Natl. Acad Sci. U. S. A. 96, 6763-6768). Previously, we demonstrated the presence of two distinct forms of the M(r) 64,000 protein of the cleavage stimulation factor (CstF-64) in mouse male germ cells and in brain, a somatic M(r) 64,000 form and a variant M(r) 70,000 form. The variant form was specific to meiotic and postmeiotic germ cells. We localized the gene for the somatic CstF-64 to the X chromosome, which would be inactivated during male meiosis. This suggested that the variant CstF-64 was an autosomal homolog activated during that time. We have named the variant form "tau CstF-64," and we describe here the cloning and characterization of the mouse tauCstF-64 cDNA, which maps to chromosome 19. The mouse tauCstF-64 protein fits the criteria of the variant CstF-64, including antibody reactivity, size, germ cell expression, and a common proteolytic digest pattern with tauCstF-64 from testis. Features of mtauCstF-64 that might allow it to promote the germ cell pattern of polyadenylation include a Pro --> Ser substitution in the RNA-binding domain and significant changes in the region that interacts with CstF-77. PMID- 11113136 TI - Characterization of CAF4 and CAF16 reveals a functional connection between the CCR4-NOT complex and a subset of SRB proteins of the RNA polymerase II holoenzyme. AB - The CCR4-NOT transcriptional regulatory complex affects transcription both positively and negatively and consists of the following two complexes: a core 1 x 10(6) dalton (1 MDa) complex consisting of CCR4, CAF1, and the five NOT proteins and a larger, less defined 1.9-MDa complex. We report here the identification of two new factors that associate with the CCR4-NOT proteins as follows: CAF4, a WD40-containing protein, and CAF16, a putative ABC ATPase. Whereas neither CAF4 nor CAF16 was part of the core CCR4-NOT complex, both CAF16 and CAF4 appeared to be present in the 1.9-MDa complex. CAF4 also displayed physical interactions with multiple CCR4-NOT components and with DBF2, a likely component of the 1.9-MDa complex. In addition, both CAF4 and CAF16 were found to interact in a CCR4 dependent manner with SRB9, a component of the SRB complex that is part of the yeast RNA polymerase II holoenzyme. The three related SRB proteins, SRB9, SRB10, and SRB11, were found to interact with and to coimmunoprecipitate DBF2, CAF4, CCR4, NOT2, and NOT1. Defects in SRB9 and SRB10 also affected processes at the ADH2 locus known to be controlled by components of the CCR4-NOT complex; an srb9 mutation was shown to reduce ADH2 derepression and either an srb9 or srb10 allele suppressed spt10-enhanced expression of ADH2. In addition, srb9 and srb10 alleles increased ADR1(c)-dependent ADH2 expression; not4 and not5 deletions are the only other known defects that elicit this phenotype. These results suggest a close physical and functional association between components of the CCR4-NOT complexes and the SRB9, -10, and -11 components of the holoenzyme. PMID- 11113137 TI - Regulation of platelet factor Va-dependent thrombin generation by activated protein C at the surface of collagen-adherent platelets. AB - Recent studies have indicated that factor Va bound to activated platelets is partially protected from inactivation by activated protein C (APC). To explore whether this sustained factor Va activity could maintain ongoing thrombin generation, the kinetics of platelet factor Va-dependent prothrombinase activity and its inhibition by APC were studied. In an attempt to mimic physiologically relevant conditions, platelets were adhered to collagen type I-coated discs. These discs were then spun in solutions containing prothrombin and factor Xa either in the absence or presence of APC. The experiments were performed in the absence of platelet-derived microparticles, with thrombin generation and inhibition confined to the surface of the adherent platelets. APC completely inactivated platelet-associated prothrombinase activity with an overall second order rate constant of 3.3 x 10(6) m(-)1 s(-)1, which was independent of the prothrombin concentration over a wide range around the apparent K(m) for prothrombin. Kinetic studies on prothrombinase assembled at a planar phospholipid membrane composed of 25 mol % phosphatidylserine and 75 mol % phosphatidylcholine revealed a similar second order rate constant of inhibition (2.5 x 10(6) m(-1) s( 1)). Collectively, these data demonstrate that ongoing platelet factor Va dependent thrombin generation at the surface of collagen-adherent platelets is effectively inhibited by APC. No differences were observed between the kinetics of APC inactivation of plasma-derived factor Va or platelet factor Va as part of the prothrombinase associated with, respectively, a planar membrane of synthetic phospholipids or collagen-adherent platelets. PMID- 11113138 TI - A novel chitin-binding protein from the vestimentiferan Riftia pachyptila interacts specifically with beta-chitin. Cloning, expression, and characterization. AB - A cDNA from Riftia pachyptila was cloned. It encodes a novel 21.3-kDa protein from the worm protective tube, named RCBP (for Riftia chitin-binding protein). On the basis of partial tube-peptide sequences previously obtained, experiments using reverse transcriptase-mediated polymerase chain reaction and rapid amplification of cDNA ends led to the complete cDNA sequence. Analysis of its deduced amino acid sequence shows the presence of two chitin-binding domains. These domains are closely related to type 2 chitin-binding domains that are restricted to the animal kingdom. We showed by affinity assay and immunogold labeling that RCBP is the first protein so far known that binds specifically beta chitin and that is unable to bind the most common alpha-form found in chitin secreting animals. The RCBP mRNA was found to be present in specific epidermal cells from the worm body wall, but never in the chitin-secreting gland cells. This unexpected result clearly indicates that this tube protein is synthesized in specialized areas of the outer epithelium and that at least two different tissues are involved in this exoskeleton synthesis. PMID- 11113139 TI - Cloning of ACP33 as a novel intracellular ligand of CD4. AB - CD4 recruitment to T cell receptor (TCR)-peptide-major histocompatibility class II complexes is required for stabilization of low affinity antigen recognition by T lymphocytes. The cytoplasmic portion of CD4 is thought to amplify TCR-initiated signal transduction via its association with the protein tyrosine kinase p56(lck). Here we describe a novel functional determinant in the cytosolic tail of CD4 that inhibits TCR-induced T cell activation. Deletion of two conserved hydrophobic amino acids from the CD4 carboxyl terminus resulted in a pronounced enhancement of CD4-mediated T cell costimulation. This effect was observed in the presence or absence of p56(lck), implying involvement of alternative cytosolic ligands of CD4. A two-hybrid screen with the intracellular portion of CD4 identified a previously unknown 33-kDa protein, ACP33 (acidic cluster protein 33), as a novel intracellular binding partner of CD4. Since interaction with ACP33 is abolished by deletion of the hydrophobic CD4 C-terminal amino acids mediating repression of T cell activation, we propose that ACP33 modulates the stimulatory activity of CD4. Furthermore, we demonstrate that interaction with CD4 is mediated by the noncatalytic alpha/beta hydrolase fold domain of ACP33. This suggests a previously unrecognized function for alpha/beta hydrolase fold domains as a peptide binding module mediating protein-protein interactions. PMID- 11113140 TI - Suppression of apoptosis induced by growth factor withdrawal by an oncogenic form of c-Cbl. AB - The v-Cbl oncogene induces myeloid and B-cell leukemia; however, the mechanism by which transformation occurs is not understood. An oncogenic form of c-Cbl (Cbl DeltaY371) was expressed in the interleukin-3 (IL-3)-dependent cell line 32Dcl3 to determine whether it was able to induce growth factor-independent proliferation. We were unable to isolate clones of transfected 32Dcl3 cells expressing Cbl-DeltaY371 that proliferated in the absence of IL-3. In contrast, 32Dcl3/Cbl-DeltaY371 cells did not undergo apoptosis like parental 32Dcl3 cells when cultured in the absence of IL-3. Both 32Dcl3 and 32D/CblDeltaY371 cells arrested in G(1) when cultured in the absence of IL-3. Approximately 18% of the 32Dcl3 cells cultured in the absence of IL-3 for 24 h were present in a sub-G(1) fraction, while only 4% of the 32D/Cbl-DeltaY371 and 2% of the 32D/Bcl-2 cells were found in a sub-G(1) fraction. There was no difference in the pattern of tyrosine-phosphorylated proteins observed following stimulation of either cell type with IL-3. The phosphorylation of JAK2, STAT5, and endogenous c-Cbl was identical in both cell types. No differences were detected in the activation of Akt, ERK1, or ERK2 in unstimulated or IL-3-stimulated 32D/Cbl-DeltaY371 cells compared with parental 32Dcl3 cells. Likewise, there was no difference in the pattern of phosphorylation of JAK2, STAT5, ERK1, ERK2, or Akt when 32Dcl3 and 32D/CblDY371 cells were withdrawn from medium containing IL-3. The protein levels of various Bcl-2 family members were examined in cells grown in the absence or presence of IL-3. We observed a consistent increased amount of Bcl-2 protein in five different clones of 32D/Cbl-DeltaY317 cells. These data suggest that the Cbl DeltaY371 mutant may suppress apoptosis by a mechanism that involves the overexpression of Bcl-2. Consistent with this result, activation of caspase-3 was suppressed in 32D/Cbl-DeltaY371 cells cultured in the absence of IL-3 compared with 32Dcl3 cells cultured under the same conditions. PMID- 11113141 TI - Mass spectrometric resolution of reversible protein phosphorylation in photosynthetic membranes of Arabidopsis thaliana. AB - The use of mass spectrometry to characterize the phosphorylome, i.e. the constituents of the proteome that become phosphorylated, was demonstrated using the reversible phosphorylation of chloroplast thylakoid proteins as an example. From the analysis of tryptic peptides released from the surface of Arabidopsis thylakoids, the principal phosphoproteins were identified by matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry. These studies revealed that the D1, D2, and CP43 proteins of the photosystem II core are phosphorylated at their N-terminal threonines (Thr), the peripheral PsbH protein is phosphorylated at Thr-2, and the mature light-harvesting polypeptides LCHII are phosphorylated at Thr-3. In addition, a doubly phosphorylated form of PsbH modified at both Thr-2 and Thr-4 was detected. By comparing the levels of phospho- and nonphosphopeptides, the in vivo phosphorylation states of these proteins were analyzed under different physiological conditions. None of these thylakoid proteins were completely phosphorylated in the steady state conditions of continuous light or completely dephosphorylated after a long dark adaptation. However, rapid reversible hyperphosphorylation of PsbH at Thr-4 in response to growth in light/dark transitions and a pronounced specific dephosphorylation of the D1, D2, and CP43 proteins during heat shock was detected. Collectively, our data indicate that changes in the phosphorylation of photosynthetic proteins are more rapid during heat stress than during normal light/dark transitions. These mass spectrometry methods offer a new approach to assess the stoichiometry of in vivo protein phosphorylation in complex samples. PMID- 11113142 TI - Cholera toxin is found in detergent-insoluble rafts/domains at the cell surface of hippocampal neurons but is internalized via a raft-independent mechanism. AB - A number of studies have demonstrated that cholera toxin (CT) is found in detergent-insoluble, cholesterol-enriched domains (rafts) in various cells, including neurons. We now demonstrate that even though CT is associated with these domains at the cell surface of cultured hippocampal neurons, it is internalized via a raft-independent mechanism, at both early and late stages of neuronal development. CT transport to the Golgi apparatus, and its subsequent degradation, is inhibited by hypertonic medium (sucrose), and by chlorpromazine; the former blocks clathrin recruitment, and the latter causes aberrant endosomal accumulation of clathrin. Moreover, both internalization of the transferrin receptor (Tf-R), which occurs via a clathrin-dependent mechanism, and CT internalization, are inhibited to a similar extent by sucrose. In contrast, the cholesterol-binding agents filipin and methyl-beta-cyclodextrin have no effect on the rate of CT or Tf-R internalization. Finally, once internalized, CT becomes more detergent-soluble, and chlorpromazine treatment renders internalized CT completely detergent-soluble. We propose two models to explain how, despite being detergent-insoluble at the cell surface, CT is nevertheless internalized via a raft-independent mechanism in hippocampal neurons. PMID- 11113143 TI - Interleukin-5-mediated allergic airway inflammation inhibits the human surfactant protein C promoter in transgenic mice. AB - Allergen challenge in the lung of humans and animals is associated with surfactant dysfunction, but the mechanism of this effect has not been established. By using a murine model of asthma we now report the effect of allergen-induced airway inflammation on the expression of transgenes regulated by the human surfactant protein (hSP)-C promoter. The hSP-C 3.7-kilobase pair promoter was used to direct the expression of eotaxin, an eosinophil-selective chemokine, into the lungs of several transgenic lines. As expected, the transgenic mice expressed increased amounts of eotaxin mRNA and protein compared with wild-type mice. Surprisingly, following allergen challenge, there was a marked down-regulation of transgene mRNA in three independent transgenic lines. The down-regulation was in contrast to other related proteins such as endogenous eotaxin and surfactant protein D levels, which were both increased following allergen challenge. Consistent with specific down-regulation of the eotaxin transgene, there was no increase in pulmonary eosinophil levels in the transgenic mice above that found in wild-type mice. Analysis of hSP-C transgenic mice with distinct reporter genes and 3'-untranslated regions revealed that allergen challenge was directly affecting the hSP-C promoter. We hypothesized that allergen-induced down-regulation of the hSP-C promoter was related to the eosinophilic inflammation. To test this, we blocked eosinophilic inflammation in the lungs by treating mice with neutralizing antiserum against interleukin-5. Interestingly, this treatment also blocked allergen-induced inhibition of the hSP C promoter. These results establish that allergic airway inflammation is associated with up-regulation of the surfactant proteins primarily involved in immunity, whereas down-regulation of the surfactant protein primarily involved in maintaining airway patency. Furthermore, the marked down-regulation of the hSP-C promoter is interleukin-5-dependent, implying a critical role for eosinophilic inflammation. These results suggest that alterations in surfactant protein levels may contribute to immune and airway dysfunction in asthma. PMID- 11113144 TI - An interaction between ricin and calreticulin that may have implications for toxin trafficking. AB - Here we demonstrate that ricin is able to interact with the molecular chaperone calreticulin both in vitro and in vivo. The interaction occurred with ricin holotoxin, but not with free ricin A chain; and it was prevented in the presence of lactose, suggesting that it was mediated by the lectin activity of the ricin B chain. This lectin is galactose-specific, and metabolic labeling with [(3)H]galactose or treating galactose oxidase-modified calreticulin with sodium [(3)H]borohydride indicated that Vero cell calreticulin possesses a terminally galactosylated oligosaccharide. Brefeldin A treatment indicated that the intracellular interaction occurred initially in a post-Golgi stack compartment, possibly the trans-Golgi network, whereas the reductive separation of ricin subunits occurred in an earlier part of the secretory pathway, most probably the endoplasmic reticulum (ER). Intoxicating Vero cells with ricin whose A chain had been modified to include either a tyrosine sulfation site or the sulfation site plus available N-glycosylation sites, in the presence of Na(2)35SO(4), confirmed that calreticulin interacted with endocytosed ricin that had already undergone retrograde transport to both the Golgi and the ER. Although we cannot exclude the possibility that the interaction between ricin and calreticulin is an indirect one, the data presented are consistent with the idea that calreticulin may function as a recycling carrier for retrograde transport of ricin from the Golgi to the ER. PMID- 11113145 TI - Cytosolic phospholipase A2 is required for optimal ATP activation of BK channels in GH(3) cells. AB - To test the hypothesis that ATP activation of BK channels in GH(3) cells involves cytosolic phospholipase A(2) (cPLA(2)) as a potential protein target for phosphorylation, we first inhibited the activity of cPLA(2) by both pharmacologic and molecular biologic approaches. Both approaches resulted in a decrease rather than an increase in BK channel activity by ATP, suggesting that in the absence of cPLA(2), phosphorylation of other regulatory elements, possibly the BK channel protein itself, results in inactivation rather than activation of the channel. The absence of changes in activity in the presence of the non-substrate ATP analog 5'-adenylyl-beta,gamma-imidodiphosphate verified that ATP hydrolysis was required for channel activation by ATP. Experiments with an activator and inhibitor of protein kinase C (PKC) support the hypothesis that PKC can be involved in the activation of BK channels by ATP; and in the absence of PKC, other kinases appear to phosphorylate additional elements in the regulatory pathway that reduce channel activity. Our data point to cPLA(2)-alpha (but not cPLA(2)-gamma) as one target protein for phosphorylation that is intimately associated with the BK channel protein. PMID- 11113146 TI - Identification and enzymatic characterization of two diverging murine counterparts of human interstitial collagenase (MMP-1) expressed at sites of embryo implantation. AB - Remodeling of fibrillar collagen in mouse tissues has been widely attributed to the activity of collagenase-3 (matrix metalloproteinase-13 (MMP-13)), the main collagenase identified in this species. This proposal has been largely based on the repeatedly unproductive attempts to detect the presence in murine tissues of interstitial collagenase (MMP-1), a major collagenase in many species, including humans. In this work, we have performed an extensive screening of murine genomic and cDNA libraries using as probe the full-length cDNA for human MMP-1. We report the identification of two novel members of the MMP gene family which are contained within the cluster of MMP genes located at murine chromosome 9. The isolated cDNAs contain open reading frames of 464 and 463 amino acids and are 82% identical, displaying all structural features characteristic of archetypal MMPs. Comparison for sequence similarities revealed that the highest percentage of identities was found with human interstitial collagenase (MMP-1). The new proteins were tentatively called Mcol-A and Mcol-B (Murine collagenase-like A and B). Analysis of the enzymatic activity of the recombinant proteins revealed that both are catalytically autoactivable but only Mcol-A is able to degrade synthetic peptides and type I and II fibrillar collagen. Both Mcol-A and Mcol-B genes are located in the A1-A2 region of mouse chromosome 9, Mcol-A occupying a position syntenic to the human MMP-1 locus at 11q22. Analysis of the expression of these novel MMPs in murine tissues revealed their predominant presence during mouse embryogenesis, particularly in mouse trophoblast giant cells. According to their structural and functional characteristics, we propose that at least one of these novel members of the MMP family, Mcol-A, may play roles as interstitial collagenase in murine tissues and could represent a true orthologue of human MMP 1. PMID- 11113147 TI - Mutagenic analysis of functional residues in putative substrate-binding site and acidic domains of vacuolar H+-pyrophosphatase. AB - Vacuolar H(+)-translocating inorganic pyrophosphatase (V-PPase) uses PP(i) as an energy donor and requires free Mg(2+) for enzyme activity and stability. To determine the catalytic domain, we analyzed charged residues (Asp(253), Lys(261), Glu(263), Asp(279), Asp(283), Asp(287), Asp(723), Asp(727), and Asp(731)) in the putative PP(i)-binding site and two conserved acidic regions of mung bean V-PPase by site-directed mutagenesis and heterologous expression in yeast. Amino acid substitution of the residues with alanine and conservative residues resulted in a marked decrease in PP(i) hydrolysis activity and a complete loss of H(+) transport activity. The conformational change of V-PPase induced by the binding of the substrate was reflected in the susceptibility to trypsin. Wild-type V PPase was completely digested by trypsin but not in the presence of Mg-PP(i), while two V-PPase mutants, K261A and E263A, became sensitive to trypsin even in the presence of the substrate. These results suggest that the second acidic region is also implicated in the substrate hydrolysis and that at least two residues, Lys(261) and Glu(263), are essential for the substrate-binding function. From the observation that the conservative mutants K261R and E263D showed partial activity of PP(i) hydrolysis but no proton pump activity, we estimated that two residues, Lys(261) and Glu(263), might be related to the energy conversion from PP(i) hydrolysis to H(+) transport. The importance of two residues, Asp(253) and Glu(263), in the Mg(2+)-binding function was also suggested from the trypsin susceptibility in the presence of Mg(2+). Furthermore, it was found that the two acidic regions include essential common motifs shared among the P-type ATPases. PMID- 11113148 TI - Biologically active recombinant human progastrin(6-80) contains a tightly bound calcium ion. AB - Evidence is accumulating that gastrin precursors may act as growth factors for the colonic mucosa in vivo. The aims of this study were to prepare recombinant human progastrin(6-80) and to investigate its structure and biological activities in vitro. Human progastrin(6-80) was expressed in Escherichia coli as a glutathione S-transferase fusion protein. After thrombin cleavage progastrin(6 80) was purified by reverse phase high pressure liquid chromatography and characterized by radioimmunoassay, amino acid sequencing, and mass spectrometry. Assays for metal ions by atomic emission spectroscopy revealed the presence of a single tightly bound calcium ion. Progastrin(6-80) at concentrations in the pm to nm range stimulated proliferation of the conditionally transformed mouse colon cell line YAMC. The observations that progastrin(6-80) did not bind to either the cholecystokinin (CCK)-A or the gastrin/CCK-B receptor expressed in COS cells and that antagonists selective for either receptor did not reverse the proliferative effects of progastrin(6-80) suggested that progastrin(6-80) stimulated proliferation independently of either the CCK-A or the gastrin/CCK-B receptor. We conclude that recombinant human progastrin(6-80) is biologically active and contains a single calcium ion. With the exception of the well known zinc dependent polymerization of insulin and proinsulin, this is the first report of selective, high affinity binding of metal ions to a prohormone. PMID- 11113149 TI - Properties of a native cation channel activated by Ca2+ store depletion in vascular smooth muscle cells. AB - Depletion of intracellular Ca(2+) stores activates capacitative Ca(2+) influx in smooth muscle cells, but the native store-operated channels that mediate such influx remain unidentified. Recently we demonstrated that calcium influx factor produced by yeast and human platelets with depleted Ca(2+) stores activates small conductance cation channels in excised membrane patches from vascular smooth muscle cells (SMC). Here we characterize these channels in intact cells and present evidence that they belong to the class of store-operated channels, which are activated upon passive depletion of Ca(2+) stores. Application of thapsigargin (TG), an inhibitor of sarco-endoplasmic reticulum Ca(2+) ATPase, to individual SMC activated single 3-pS cation channels in cell-attached membrane patches. Channels remained active when inside-out membrane patches were excised from the cells. Excision of membrane patches from resting SMC did not by itself activate the channels. Loading SMC with BAPTA (1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid), which slowly depletes Ca(2+) stores without a rise in intracellular Ca(2+), activated the same 3-pS channels in cell-attached membrane patches as well as whole cell nonselective cation currents in SMC. TG- and BAPTA-activated 3-pS channels were cation-selective but poorly discriminated among Ca(2+), Sr(2+), Ba(2+), Na(+), K(+), and Cs(+). Open channel probability did not change at negative membrane potentials but increased significantly at high positive potentials. Activation of 3-pS channels did not depend on intracellular Ca(2+) concentration. Neither TG nor a variety of second messengers (including Ca(2+), InsP3, InsP4, GTPgammaS, cyclic AMP, cyclic GMP, ATP, and ADP) activated 3-pS channels in inside-out membrane patches. Thus, 3-pS nonselective cation channels are present and activated by TG or BAPTA-induced depletion of intracellular Ca(2+) stores in intact SMC. These native store-operated cation channels can account for capacitative Ca(2+) influx in SMC and can play an important role in regulation of vascular tone. PMID- 11113150 TI - An essential cytoplasmic domain for the Golgi localization of coiled-coil proteins with a COOH-terminal membrane anchor. AB - Giantin is a resident Golgi protein that has an extremely long cytoplasmic domain (about 370 kDa) and is anchored to the Golgi membrane by the COOH-terminal membrane-anchoring domain (CMD) with no luminal extension. We examined the essential domain of giantin required for Golgi localization by mutational analysis. The Golgi localization of giantin was not affected by the deletion of its CMD or by substitution with the CMD of syntaxin-2, a plasma membrane protein. The giantin CMD fused to the cytoplasmic domain of syntaxin-2 could not retain the chimera in the Golgi apparatus. Sequential deletion analysis showed that the COOH-terminal sequence (positions 3059--3161) adjacent to the CMD was the essential domain required for the Golgi localization of giantin. We also examined two other Golgi-resident proteins, golgin-84 and syntaxin-5, with a similar membrane topology as giantin. It was confirmed that the cytoplasmic domain of about 100 residues adjacent to the CMD was required for their Golgi localization. Taken together, these results suggest that the COOH-terminally anchored Golgi proteins with long cytoplasmic extensions have the Golgi localization signal(s) in the cytoplasmic sequence adjacent to the CMD. This is in contrast to previous observations that a transmembrane domain is required for Golgi localization by other Golgi proteins transported from the endoplasmic reticulum. PMID- 11113151 TI - Mutational definition of RNA-binding and protein-protein interaction domains of heterogeneous nuclear RNP C1. AB - The heterogeneous nuclear ribonucleoprotein (hn- RNP) C proteins, among the most abundant pre-mRNA-binding proteins in the eukaryotic nucleus, have a single RNP motif RNA-binding domain. The RNA-binding domain (RBD) is comprised of approximately 80-100 amino acids, and its structure has been determined. However, relatively little is known about the role of specific amino acids of the RBD in the binding to RNA. We have devised a phage display-based screening method for the rapid identification of amino acids in hnRNP C1 that are essential for its binding to RNA. The identified mutants were further tested for binding to poly(U) Sepharose, a substrate to which wild type hnRNP C1 binds with high affinity. We found both previously predicted, highly conserved residues as well as additional residues in the RBD to be essential for C1 RNA binding. We also identified three mutations in the leucine-rich C1-C1 interaction domain near the carboxyl terminus of the protein that both abolished C1 oligomerization and reduced RNA binding. These results demonstrate that although the RBD is the primary determinant of C1 RNA binding, residues in the C1-C1 interaction domain also influence the RNA binding activity of the protein. The experimental approach we described should be generally applicable for the screening and identification of amino acids that play a role in the binding of proteins to nucleic acid substrates. PMID- 11113152 TI - The regulation of type 7 adenylyl cyclase by its C1b region and Escherichia coli peptidylprolyl isomerase, SlyD. AB - Mammalian membrane-bound adenylyl cyclase consists of two highly conserved cytoplasmic domains (C1a and C2a) separated by a less conserved connecting region, C1b, and one of two transmembrane domains, M2. The C1a and C2a domains form a catalytic core that can be stimulated by forskolin and the stimulatory G protein subunit alpha (Galpha(s)). In this study, we analyzed the regulation of type 7 adenylyl cyclase (AC7) by C1b. The C1a, C1b, and C2a domains of AC7 were purified separately. Escherichia coli SlyD protein, a cis-trans peptidylprolyl isomerase (PPIase), copurifies with AC7 C1b (7C1b). SlyD protein can inhibit the Galpha(s)- and/or forskolin-activated activity of both soluble and membrane-bound AC7. Mutant forms of SlyD with reduced PPIase activity are less potent in the inhibition of AC7 activity. Interestingly, different isoforms of mammalian membrane-bound adenylyl cyclase can be either inhibited or stimulated by SlyD protein, raising the possibility that mammalian PPIase may regulate enzymatic activity of mammalian adenylyl cyclase. Purified 7C1b-SlyD complex has a greater inhibitory effect on AC7 activity than SlyD alone. This inhibition by 7C1b is abolished in a 7C1b mutant in which a conserved glutamic acid (amino acid residue 582) is changed to alanine. Inhibition of adenylyl cyclase activity by 7C1b is further confirmed by using 7C1b purified from an E. coli slyD-deficient strain. This inhibitory activity of AC7 is also observed with the 28-mer peptides derived from a region of C1b conserved in AC7 and AC2 but is not observed with a peptide derived from the corresponding region of AC6. This inhibitory activity exhibited by the C1b domain may result from the interaction of 7C1b with 7C1a and 7C2a and may serve to hold AC7 in the basal nonstimulated state. PMID- 11113153 TI - Overexpression of a modified human malonyl-CoA decarboxylase blocks the glucose induced increase in malonyl-CoA level but has no impact on insulin secretion in INS-1-derived (832/13) beta-cells. AB - The long-chain acyl-CoA (LC-CoA) model of glucose-stimulated insulin secretion (GSIS) holds that secretion is linked to a glucose-induced increase in malonyl CoA level and accumulation of LC-CoA in the cytosol. We have previously tested the validity of this proposal by overexpressing goose malonyl-CoA decarboxylase (MCD) in INS-1 cells, but these studies have been criticized due to: 1) the small insulin secretion response (2-4-fold) of the INS-1 cells used; 2) unknown contribution of the ATP-sensitive K(+) (K(ATP)) channel-independent pathway of GSIS in INS-1 cells, which has been implicated as the site at which lipids regulate insulin granule exocytosis; and 3) deletion of the N-terminal mitochondrial targeting sequence, but not the C-terminal peroxisomal targeting sequence in the goose MCD construct, raising the possibility that a significant fraction of the overexpressed enzyme was localized to peroxisomes. To address these outstanding concerns, INS-1-derived 832/13 cells, which exhibit robust K(ATP) channel-dependent and -independent pathways of GSIS, were treated with a new adenovirus encoding human MCD lacking both its mitochondrial and peroxisomal targeting sequences (AdCMV-MCD Delta 5), resulting in large increases in cytosolic MCD activity. Treatment of 832/13 cells with AdCMV-MCD Delta 5 completely blocked the glucose-induced rise in malonyl-CoA and attenuated the inhibitory effect of glucose on fatty acid oxidation. However, MCD overexpression had no effect on K(ATP) channel-dependent or -independent GSIS in 832/13 cells. Furthermore, combined treatment of 832/13 cells with AdCMV-MCD Delta 5 and triacsin C, an inhibitor of long chain acyl-CoA synthetase that reduces LC-CoA levels, did not impair GSIS. These findings extend our previous observations and are not consistent with the LC-CoA hypothesis as originally set forth. PMID- 11113154 TI - Assembly of scaffold-mediated complexes containing Cdc42p, the exchange factor Cdc24p, and the effector Cla4p required for cell cycle-regulated phosphorylation of Cdc24p. AB - In budding yeast cells, the cytoskeletal polarization and depolarization events that shape the bud are triggered at specific times during the cell cycle by the cyclin-dependent kinase Cdc28p. Polarity establishment also requires the small GTPase Cdc42p and its exchange factor, Cdc24p, but the mechanism whereby Cdc28p induces Cdc42p-dependent polarization is unknown. Here we show that Cdc24p becomes phosphorylated in a cell cycle-dependent manner, triggered by Cdc28p. However, the role of Cdc28p is indirect, and the phosphorylation appears to be catalyzed by the p21-activated kinase family member Cla4p and also depends on Cdc42p and the scaffold protein Bem1p. Expression of GTP-Cdc42p, the product of Cdc24p-mediated GDP/GTP exchange, stimulated Cdc24p phosphorylation independent of cell cycle cues, raising the possibility that the phosphorylation is part of a feedback regulatory pathway. Bem1p binds directly to Cdc24p, to Cla4p, and to GTP bound Cdc42p and can mediate complex formation between these proteins in vitro. We suggest that Bem1p acts to concentrate polarity establishment proteins at a discrete site, facilitating polarization and promoting Cdc24p phosphorylation at specific times during the cell cycle. PMID- 11113155 TI - The molecular adapter SLP-76 relays signals from platelet integrin alphaIIbbeta3 to the actin cytoskeleton. AB - Platelet adhesion to fibrinogen through integrin alpha(IIb)beta(3) triggers actin rearrangements and cell spreading. Mice deficient in the SLP-76 adapter molecule bleed excessively, and their platelets spread poorly on fibrinogen. Here we used human platelets and a Chinese hamster ovary (CHO) cell expression system to better define the role of SLP-76 in alpha(IIb)beta(3) signaling. CHO cell adhesion to fibrinogen required alpha(IIb)beta(3) and stimulated tyrosine phosphorylation of SLP-76. SLP-76 phosphorylation required coexpression of Syk tyrosine kinase and stimulated association of SLP-76 with the adapter, Nck, and with the Rac exchange factor, Vav1. SLP-76 expression increased lamellipodia formation induced by Syk and Vav1 in adherent CHO cells (p < 0.001). Although lamellipodia formation requires Rac, SLP-76 functioned downstream of Rac by potentiating adhesion-dependent activation of PAK kinase (p < 0.001), a Rac effector that associates with Nck. In platelets, adhesion to fibrinogen stimulated the association of SLP-76 with the SLAP-130 adapter and with VASP, a SLAP-130 binding partner implicated in actin reorganization. Furthermore, SLAP 130 colocalized with VASP at the periphery of spread platelets. Thus, SLP-76 functions to relay signals from alpha(IIb)beta(3) to effectors of cytoskeletal reorganization. Therefore, deficient recruitment of specific adapters and effectors to sites of adhesion may explain the integrin phenotype of SLP-76(-/-) platelets. PMID- 11113156 TI - Aspects of seasonality. AB - The seasons are astronomical, astrological, meteorological, biological, and agricultural. From a perspective outside the biological sciences, the questions of interest about plant seasonality are linked to this wider context. In this review I try to see flowering time, as one important aspect of seasonality, from an outsider's point of view, and describe what is known about it in different types of plants. What is known about it is conditioned by what particular scientists have asked about it, so the variety of approaches to seasonality is another point of emphasis. Detailed consideration is given to flowering seasonality in perennials compared with annuals, and both molecular and whole plant perspectives are presented. PMID- 11113157 TI - Annuality, perenniality and cell death. AB - This essay considers annuality and perenniality as quantitative traits and discusses the application of established and new genetic tools to the analysis of plant life histories. Annual/perennial status is a function of meristem determinacy in combination with the processes of cell death and disposal employed by plants to generate well-adapted anatomies and morphologies. Creeping perennials, like clover or bracken, seem to move around in the environment. They do this by extending into unoccupied space while the oldest tissues behind the growing and mature regions senesce, die and decompose. Trees do essentially the same thing, except that they develop vertically and the old dead tissue does not disappear but instead persists as wood. A root system is a kind of upended vertical perennial. The balance between exploratory growth and the wave of tissue death that succeeds it is a major determinant of perenniality. So although perenniality and annuality may appear to be dramatically different traits, extremes of behaviour can arise by a relatively minor change in the relationship between growth and death. This conclusion is supported by evidence from genome dosage studies, from the practical experiences of breeding perennial-type traits into annual backgrounds and from molecular cladistics. Applications of methods for the genetic analysis of quantitative characters are described, including the exploitation of introgression mapping in Lolium-Festuca and quantitative trait locus mapping in cereals and other species. PMID- 11113158 TI - Isolation of three distinct CycD3 genes expressed during fruit development in tomato. AB - Tomato (Lycopersicon esculentum Mill.) is an important fruit crop world-wide and a model for studying fruit development. As determined using flow cytometry, fruit growth was characterized by high cell division activity in tomato during the first week after anthesis and followed by endoreduplications (DNA replication without cell divisions). D-type cyclins are considered to be important parts of the signal transduction for stimulation of DNA replication and cell division. To study the function of D cyclins in fruit development, full-length cDNA clones for three D cyclin genes were isolated from young tomato fruit. They were classified as D3 cyclins by sequence similarities and a phylogenetic analysis and named as LeCycD3;1, LeCycD3;2 and LeCycD3;3. The deduced amino acid sequences for LeCycD3;1-3 contained a retinoblastoma-binding motif and a PEST-destruction motif. Pollination and fertilization were followed by a high increase in the transcript levels of LeCycD3;1-3 in young fruit. Using in situ hybridization, high expression of LeCycD3;3 was detected in the vascular tissue of young fruit suggesting a role in vascular development. The D3 cyclins are probably involved in transducing the signals leading to fruit growth by cell divisions. Distinct differences were detected in their temporal and spatial expression patterns suggesting that they play different roles in fruit development as well as in the development of other plant organs. PMID- 11113159 TI - The fungal elicitor cryptogein induces cell wall modifications on tobacco cell suspension. AB - Upon addition of the fungal elicitor cryptogein, suspension cells of tobacco (Nicotiana tabacum cv. Xanthi) aggregated in clusters. Cytochemical experiments indicated that elicited cells displayed fibrillar expansions of pectin along the primary cell wall. Immunocytochemical detection of pectin epitopes indicated that the fibrillar material surrounding the treated cells was mostly composed of low methylated galacturonan sequences, but the use of the cationic probe did not reveal the presence of negatively charged carboxyl groups: the presence of important amounts of calcium ions in these pectic fibrillar expansions accounts for these observations. These data indicate that tobacco cells treated with cryptogein show a cell wall altered by the presence of a calcium pectate gel, resulting from the reorganization of pectin in the middle lamellae. These results are consistent with a drastic reduction in wall digestibility, partially reversed by increasing the pectolyase concentration in the hydrolytic solution. Diphenylene iodonium, an inhibitor of the oxidative burst triggered by cryptogein on tobacco cells, partially prevents elicited cell walls from this loss of digestibility, suggesting a possible role of active oxygen species in the cell wall strengthening. This work represents a new element of the signal transduction cascade triggered on tobacco cells by cryptogein. PMID- 11113160 TI - Biomechanical study of the effect of a controlled bending on tomato stem elongation: global mechanical analysis. AB - An experiment was designed to apply a controlled bending to a tomato stem and simultaneously to measure its effect on stem elongation. Stem elongation was measured over 2 d until steady and equal rates were obtained for the control and the treated plants. Thereafter, the basal part of the stem was submitted to a transient controlled bending at constant displacement rate using a motorized dynamometer. After load removal, stem elongation was again measured for 2 d. The tested plants were mature (height visible internodes) and only the basal part of the stem, which had already finished elongation, was loaded (hypocotyl and the first three internodes). A few minutes after the application of bending, elongation stopped completely for 60 min. Thereafter it took 120-1000 min to recover a rate of elongation similar to the control. The growth response was exclusively due to the bending of the basal part of the stem. It was shown that the side mechanical perturbations on the roots and on the stem tissues interacting directly with the clamp were not significantly involved on the elongation response. These results give evidence for mechanical perception and plant signalling from the basal stem to the upper elongating zone. However, none of the variables characterizing the global mechanical state of the bent part of the stem (i.e. the maximal force, bending moment, inclination, mean curvature of the stem, stored mechanical energy) could quantitatively explain the variability of the growth response. A more local mechanical analysis is therefore needed to elucidate how the mechanical stimulus is perceived. PMID- 11113161 TI - Biomechanical study of the effect of a controlled bending on tomato stem elongation: local strain sensing and spatial integration of the signal. AB - In a previous paper it has been demonstrated that tomato stems, submitted to a controlled basal bending, had a reduced terminal primary elongation, indicating mechanosensing and intra plant signalling. The 'intensity' of the growth response, as measured by the time to recover an elongation rate similar to the control, varied hugely between plants. However, no relation was found between the intensity of this response and the mechanical variables characterizing the global mechanical state of the stem. In this paper, a local analysis of mechanical state of each bent stem is performed in the context of beam theory. The spatial distributions of local variables all along the stem (curvature, bending moment, strains and stresses) are established. The validity of hypotheses underlying the mechanical analysis is demonstrated. To investigate the relationships between the mechanical stimulus and the growth response, a novel biomechanical analysis based on spatial integration of the mechanical stimulus is presented. It revealed that the mechanosensing is local and scattered through the stem and that the variability of the growth response is only explained by the integrals of the longitudinal strain field. PMID- 11113162 TI - Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.). AB - The effects of the plant growth regulator ethylene, and of ethylene inhibitors, on barley (Hordeum vulgare L.) germination and seedling growth were investigated. Exogenous 1-aminocyclopropane-1-carboxylic acid (ACC) at 100 microM enhanced ethylene production by barley seedlings and stimulated shoot growth, whereas both germination and seedling growth were inhibited by antagonists of ethylene perception (75 microM silver ions, 100 microM 2,5-norbornadiene (NBD)). In contrast, germination was unaffected by, and root and shoot growth of seedlings was strongly stimulated by inhibitors of ethylene biosynthesis (10 microM cobalt chloride, 10 microM aminoethoxyvinylglycine (AVG)). Since the ethylene and polyamine biosynthetic pathways are linked through S:-adenosylmethionine, this prompted further explorations into the role of polyamines in germination and seedling growth. Exogenous polyamines (putrescine, spermidine and spermine) at 1 microM concentration stimulated barley seedling growth in a similar fashion to the ethylene biosynthetic inhibitors. Both polyamines and ethylene biosynthetic inhibitors reversed the inhibitory effects of ethylene perception inhibitors on germination and seedling growth. Blocking endogenous ethylene production with aminoethoxyvinylglycine enhanced the free putrescine and spermidine content of germinating barley grains. Thus endogenous polyamines may play a complementary, growth-promotive, role to ethylene in the normal course of barley germination. Further, experiments that have been carried out using inhibitors of ethylene biosynthesis may have to be re-evaluated to take the possible effect of polyamines into account. PMID- 11113163 TI - Role of ethylene in cotyledon development of microspore-derived embryos of Brassica napus. AB - Ethylene production during seed development in Brassica napus occurs first at 20 d after pollination (DAP), while a second greater peak occurs at 35 DAP. Because of the inaccessible location of the embryo within the maternal tissue, microspore derived embryos (MDEs) of B. napus were used as a model for studying the role of ethylene during embryo development. The MDEs also produced a peak in ethylene evolution at 20 DAC (i.e. the early cotyledonary stage), dropping to minimal levels by 25-30 DAC. At 20 DAC the excised cotyledon evolved 85% of the ethylene found in the whole MDE. To determine the role of ethylene, MDEs were treated with aminoethoxyvinylglycine (AVG, an inhibitor of ethylene biosynthesis), CoCl(2) (an inhibitor of 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase), and silver thiosulphate (STS, an inhibitor of ethylene action). An inhibition in ethylene production or action at 20 DAC resulted in diminished lateral cotyledon expansion, due to a reduction in the lateral expansion of cells within the cotyledon. Recovery to 'control-type' levels of cotyledon cell expansion was achieved by application of ACC (the metabolic precursor of ethylene) to AVG treated MDEs. Thus, ethylene production at 20 DAP likely controls cotyledon expansion during embryo development. PMID- 11113164 TI - The role of abscisic acid in controlling leaf water loss, survival and growth of micropropagated Tagetes erecta plants when transferred directly to the field. AB - Plants of Tagetes erecta L. (marigold) cultivated in vitro in ventilated containers exhibited greater control of leaf water loss and increased survival in the field than plants cultivated in sealed containers. Increased field survival of plants cultivated in ventilated containers was attributed to higher levels of endogenous abscisic acid (ABA). Therefore, ABA was supplied exogenously to plants in sealed or ventilated containers by adding ABA (10(-6), 10(-5), 10(-4) M) to the in vitro culture media in order to evaluate control of leaf water loss, growth and field survival. The addition of 10(-4) M ABA to the culture media in sealed containers produced plants that had similar control of leaf water loss and were morphologically similar to plants cultivated in ventilated containers without the addition of ABA. Field survival of 10(-4) M ABA plants (75%) was increased compared to plants cultivated in sealed containers without ABA (31%), with survival being closer to that of plants cultivated in ventilated containers (90-100%). Plants cultivated with 10(-4) M ABA (sealed and ventilated) also exhibited increased plant vigour and leaf area in the field compared to plants cultivated without ABA. The results suggest that the limited field survival and growth of plants cultured in vitro are related to the limited ABA concentrations they accumulate while in vitro. Consequently, conditions that increase the endogenous ABA concentrations of in vitro plants (like ventilation or ABA addition to the medium) would improve the control of leaf water loss, field survival and plant vigour. PMID- 11113165 TI - Overexpression of Mn-superoxide dismutase in maize leaves leads to increased monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase activities. AB - The effect of increased Mn-superoxide dismutase (SOD) on antioxidant enzymes and metabolites was studied using transformed maize, TG1+ and TG2+. The progeny of the backcross of each of the primary transformants with the parental line generated two populations denoted M6884 and M6885. These were grown at optimal (25 degrees C) and sub-optimal (18, 14 and 10 degrees C) temperatures to assess the impact of elevated SOD activity on cold tolerance and the antioxidant defences in maize. The plants of the M6885 population had similar foliar SOD activities to the untransformed maize plants. Within the segregating M6884 population 50% of the plants had elevated SOD activity (up to four times the activity of the untransformed controls) and 50% of the plants contained the product of the transgene. In untransformed plants grown at 25 degrees C and 18 degrees C, SOD activity was not detectable in mesophyll extracts. Similarly, increased foliar SOD activity in the M6884 transformed maize did not lead to detectable mesophyll SOD activity. Increased foliar KCN-insensitive SOD activities were accompanied by enhancement of monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase activities; enzymes which are localized exclusively in the leaf mesophyll tissues. Increased foliar SOD activity had no effect on the hydrogen peroxide, glutathione or ascorbate contents of the leaves. This suggests that increased recycling of reduced ascorbate was required to compensate for enhanced hydrogen peroxide production in transformed plants. PMID- 11113166 TI - The nitrogen handling characteristics of white clover (Trifolium repens L.) cultivars and a perennial ryegrass (Lolium perenne L.) cultivar. AB - Ryegrass (Lolium perenne L.) and white clover (Trifolium repens L.) have contrasting responses to soil mineral N availability and clover has the ability to fix atmospheric N(2) symbiotically. It has been hypothesized that these differences are the key to understanding grass-clover coexistence and vegetative dynamics in pastures. However, the whole plant response of clover and ryegrass to mineral N availability has not been fully characterized and inter-cultivar variability in the N-handling dynamics of clover has not been assessed. A detailed experimental study to address these issues was undertaken. For all clover cultivars and ryegrass, mass specific mineral N uptake rates (of whole plants) were similar saturating functions of mineral N availability. For all clover cultivars total N assimilation rates, whole plant C : N ratios and root : shoot ratios were independent of mineral N availability. Clover growth rates were also independent of mineral N availability except for a slight (<10%) reduction at very low N availability levels. Specific N(2) fixation rate (whole plant) was precisely controlled to ensure fixation balanced the deficit between mineral N uptake and the total N assimilation required to maintain constant whole plant C : N ratio. There was always a deficit between N uptake and the total N assimilation required to maintain C : N ratio. Consequently, some N(2) fixation remained engaged even at high mineral N availability levels. All inter-cultivar variation in N(2) fixation dynamics could be attributed to variations in growth rate. Clover mass specific growth rate declined as plant size increased. Ryegrass specific growth rate, whole plant C : N ratio and root : shoot ratio were dependent on N availability. Increased N availability led to increased growth rate and decreased C : N and root : shoot ratios. Specific growth rate was also dependent on plant size, growth rate declining as plant size increased. It is concluded that clover inter-cultivar variation in field performance is unlikely to be a consequence of variation in N-handling characteristics. Inter-cultivar differences in growth rate are likely to be a much more important source of variation. PMID- 11113167 TI - An overnight chill induces a delayed inhibition of photosynthesis at midday in mango (Mangifera indica L.). AB - The effect of a cold night on photosynthesis in herbaceous chilling-sensitive crops, like tomato, has been extensively studied and is well characterized. This investigation examined the behaviour of the sub-tropical fruit tree, mango, to enable comparison with these well-studied systems. Unlike tomato, chilling between 5 degrees C and 7 degrees C overnight produced no significant inhibition of light-saturated CO(2) assimilation (A:) during the first hours following rewarming, measured either under controlled environment conditions or in the field. By midday, however, there was a substantial decline in A:, which could not be attributed to photoinhibition of PSII, but rather was associated with an increase in stomatal limitation of A: and lower Rubisco activity. Overnight chilling of tomato can cause severe disruption in the circadian regulation of key photosynthetic enzymes and is considered to be a major factor underlying the dysfunction of photosynthesis in chilling-sensitive herbaceous plants. Examination of the gas exchange of mango leaves maintained under constant conditions for 2 d, demonstrated that large depressions in A: during the subjective night were primarily the result of stomatal closure. Chilling did not disrupt the ability of mango leaves to produce a circadian rhythm in stomatal conductance. Rather, the midday increase in stomatal limitation of A: appeared to be the result of altered guard cell sensitivity to CO(2) following the dark chill. PMID- 11113168 TI - Phosphoenolpyruvate carboxylase in cucumber (Cucumis sativus L.) roots under iron deficiency: activity and kinetic characterization. AB - Phosphoenolpyruvate carboxylase (PEPCase) activity was investigated in cucumber roots grown under iron starvation. The enzyme extracted from plants grown in the presence and in the absence of Fe was characterized both kinetically and biochemically. Extractable PEPCase activity was increased by 4-fold in the absence of Fe. This increase began about 5 d after Fe starvation. Western blot analysis revealed the presence of two polypeptides with apparent molecular masses of 103 and 108 kDa. At the beginning both the polypeptides were equally present in the control and in the Fe-deficient roots. After 10 d of Fe starvation the increase was clearly evident and concerned, in particular, the polypeptide of 103 kDa whose enhancement was around 3-fold with respect to the control. Re-supply of iron to Fe-starved roots decreased both the activity and the concentration of the enzyme to the control values. Determination of kinetic parameters revealed that the K:(m) values for the substrates were the same, while the V:(max) was increased by four times for the enzyme extracted from Fe-deficient roots. Also the responses to pH changes and to the allosteric modulators malate and glucose-6 phosphate were different. The kinetic data, the increase in PEPCase specific activity and in the PEPCase polypeptides concentration seem to indicate that under Fe deficiency the enzyme regulation might be, in part, exerted at the transcriptional level. PMID- 11113169 TI - Desiccation tolerance of recalcitrant Theobroma cacao embryonic axes: the optimal drying rate and its physiological basis. AB - Recalcitrant seed axes were reported to survive to lower water contents under fast-drying conditions. The present study was to examine the hypothesis that drying rate and dehydration duration could interact to determine desiccation tolerance through different physico-chemical mechanisms. The effect of drying rate on desiccation tolerance of Theobroma cacao seed axes at 16 degrees C was examined. Rapid-drying at low relative humidity (RH) and slow-drying at high RH were more harmful to cocoa axes, because electrolyte leakage began to increase and axis viability began to decrease at high water contents. Maximum desiccation tolerance was observed with intermediate drying rates at RH between 88% and 91%, indicating the existence of an optimal drying rate or optimal desiccation duration. This maximum level of desiccation tolerance for cocoa axes (corresponding to a critical water potential of -9 MPa) was also detected using the equilibration method, in which axes were dehydrated over a series of salt solutions or glycerol solutions until the equilibrium. These data confirmed that the physiological basis of the optimal drying rate is related to both mechanical stress during desiccation and the length of desiccation duration during which deleterious reactions may occur. The optimal drying rate represents a situation where combined damages from mechanical and metabolic stresses become minimal. PMID- 11113170 TI - The strategy of the wheat plant in acclimating growth and grain production to nitrogen availability. AB - Two cultivars of spring wheat (Triticum aestivum L.) were grown to maturity in hydroponic cultures. Nitrogen accumulation was controlled by daily growth limiting additions of nitrate together with all other nutrients in excess. Six different curves of N accumulation were used, with the same relative changes from day to day, but with different amplitudes. These curves were obtained by using the same mathematic formula of the N accumulation curves but varying the value of initial N content. The total amount of nitrogen added varied from 20 mg plant(-1) to 65 mg plant(-1). Plant bioproductivity showed a linear response to accumulated N. The number of grains per plant increased linearly with increased N availability whereas grain weights were essentially unaffected. Grain N concentrations and N content varied slightly, with highest values generally at the lower N availability levels. The quantitatively most important response to increased N availability was an increased number of earbearing tillers per plant. This varied from 0.1 tiller plant(-1) at maturity when given 20 mg N plant(-1), up to about 2 tillers plant(-1) when given 65 mg N plant(-1). Not all tillers that were initiated developed ears. The reduction of tillers seems to be one important mechanism in adapting plant productivity to N availability. Other individual characters influenced by N availability were straw height and the number of spikelets per spike. The two cultivars behaved in a qualitatively similar manner over the range of N availability even though they quantitatively differed in grain size, N concentrations and yield. PMID- 11113171 TI - Atmospheric CO(2) and mycorrhiza effects on biomass allocation and nutrient uptake of nodulated pea (Pisum sativum L.) plants. AB - The effect of ambient and elevated atmospheric CO(2) on biomass partitioning and nutrient uptake of mycorrhizal and non-mycorrhizal pea plants grown in pots in a controlled environment was studied. The hypothesis tested was that mycorrhizae would increase C assimilation by increasing photosynthetic rates and reduce below ground biomass allocation by improving nutrient uptake. This effect was expected to be more pronounced at elevated CO(2) where plant C supply and nutrient demand would be increased. The results showed that mycorrhizae did not interact with atmospheric CO(2) concentration in the variables measured. Mycorrhizae did not affect photosynthetic rates, had no effect on root weight or root length density and almost no effect on nutrient uptake, but still significantly increased shoot weight and reduced root/shoot ratio at harvest. Elevated CO(2) increased photosynthetic rates with no evidence for down-regulation, increased shoot weight and nutrient uptake, had no effect on root weight, and actually reduced root/shoot ratio at harvest. Non-mycorrhizal plants growing at both CO(2) concentrations had lower shoot weight than mycorrhizal plants with similar nutritional status and photosynthetic rates. It is suggested that the positive effect of mycorrhizal inoculation was caused by an enhanced C supply and C use in mycorrhizal plants than in non-mycorrhizal plants. The results indicate that plant growth was not limited by mineral nutrients, but partially source and sink limited for carbon. Mycorrhizal inoculation and elevated CO(2) might have removed such limitations and their effects on above-ground biomass were independent, positive and additive. PMID- 11113172 TI - Abscisic acid and hypoxic induction of anoxia tolerance in roots of lettuce seedlings. AB - Lettuce (Lactuca sativa L.) seedlings were subjected to anoxic stress after ABA pretreatment (ABA-PT) or hypoxic-pretreatment (H-PT). The H-PT increased the survivability of the anoxia in roots of the seedlings by 5.2-fold compared to that of non-pretreated (N-PT) seedlings. ABA-PT also increased the survivability at concentrations greater than 1 microM, and the survivability increased with increasing ABA doses. At 100 microM ABA, the survivability was 4.5-fold greater than that of N-PT seedlings. During pretreatment periods, alcohol dehydrogenase (ADH, EC 1.1.1.1) activity in the roots became 3.1- and 3.4-fold greater than that of N-PT seedlings following 100 microM ABA-PT and H-PT seedlings, respectively. After the onset of anoxic stress, ADH activities in all roots increased, but the activities in H-PT and ABA-PT roots remained much greater than that in N-PT roots, and the average ethanol production rate for the initial 6 h was 5.3, 4.0 and 1.4 micromol g(-1) FW h(-1) for H-PT, ABA-PT and N-PT roots, respectively. Roots of the seedlings lost ATP rapidly under anoxic stress; however, the decrease in ATP was much slower in the ABA-PT and H-PT seedlings than in the N-PT seedlings. These results suggest that the ABA-PT and H-PT may maintain ATP levels due to activation of ethanolic fermentation, which may be one of the causes of the increasing anoxia tolerance in the seedling roots. Measurement of endogenous ABA levels, however, showed that ABA levels did not increase during the H-PT, suggesting that the H-PT does not increase tolerance through an increase in ABA levels. PMID- 11113174 TI - Rubisco activation state decreases with increasing nitrogen content in apple leaves PMID- 11113173 TI - A novel DNA-binding protein associated with DNA polymerase-alpha in pea stimulates polymerase activity on infrequently primed templates. AB - A 42 kDa DNA-binding protein is associated with DNA polymerase-alpha-primase in pea (Pisum sativum). In a previous publication it was shown that the protein has strong preference for ds-ss junctions in DNA, including the cohesive termini generated by restriction endonucleases. In this paper it is shown that when the DNA-binding protein is added back to polymerase-primase, the protein stimulates the activity of the polymerase. The stimulation is particularly marked when M13 DNA, primed with a single sequencing primer or primed with oligoribonucleotides by the polymerase's associated primase activity, is used as a template. The stimulation of polymerase activity is not caused by an increase in processivity. These data lead to the suggestion that the 42 kDa DNA-binding protein is a primer recognition protein. PMID- 11113175 TI - Regulation of leaf and fruit growth in plants growing in drying soil: exploitation of the plants' chemical signalling system and hydraulic architecture to increase the efficiency of water use in agriculture PMID- 11113177 TI - The Sko1p repressor and Gcn4p activator antagonistically modulate stress regulated transcription in Saccharomyces cerevisiae. AB - In the transcriptional response of Saccharomyces cerevisiae to stress, both activators and repressors are implicated. Here we demonstrate that the ion homeostasis determinant, HAL1, is regulated by two antagonistically operating bZIP transcription factors, the Sko1p repressor and the Gcn4p activator. A single CRE-like sequence (CRE(HAL1)) at position -222 to -215 with the palindromic core sequence TTACGTAA is essential for stress-induced expression of HAL1. Down regulation of HAL1 under normal growth conditions requires specific binding of Sko1p to CRE(HAL1) and the corepressor gene SSN6. Release from this repression depends on the function of the high-osmolarity glycerol pathway. The Gcn4p transcriptional activator binds in vitro to the same CRE(HAL1) and is necessary for up-regulated HAL1 expression in vivo, indicating a dual control mechanism by a repressor-activator pair occupying the same promoter target sequence. gcn4 mutants display a strong sensitivity to elevated K(+) or Na(+) concentrations in the growth medium. In addition to reduced HAL1 expression, this sensitivity is explained by the fact that amino acid uptake is drastically impaired by high Na(+) and K(+) concentrations in wild-type yeast cells. The reduced amino acid biosynthesis of gcn4 mutants would result in amino acid deprivation. Together with the induction of HAL1 by amino acid starvation, these results suggest that salt stress and amino acid availability are physiologically interconnected. PMID- 11113176 TI - Studies of nematode TFIIE function reveal a link between Ser-5 phosphorylation of RNA polymerase II and the transition from transcription initiation to elongation. AB - The general transcription factor TFIIE plays important roles in transcription initiation and in the transition to elongation. However, little is known about its function during these steps. Here we demonstrate for the first time that TFIIH-mediated phosphorylation of RNA polymerase II (Pol II) is essential for the transition to elongation. This phosphorylation occurs at serine position 5 (Ser 5) of the carboxy-terminal domain (CTD) heptapeptide sequence of the largest subunit of Pol II. In a human in vitro transcription system with a supercoiled template, this process was studied using a human TFIIE (hTFIIE) homolog from Caenorhabditis elegans (ceTFIIEalpha and ceTFIIEbeta). ceTFIIEbeta could partially replace hTFIIEbeta, whereas ceTFIIEalpha could not replace hTFIIEalpha. We present the studies of TFIIE binding to general transcription factors and the effects of subunit substitution on CTD phosphorylation. As a result, ceTFIIEalpha did not bind tightly to hTFIIEbeta, and ceTFIIEbeta showed a similar profile for binding to its human counterpart and supported an intermediate level of CTD phosphorylation. Using antibodies against phosphorylated serine at either Ser-2 or Ser-5 of the CTD, we found that ceTFIIEbeta induced Ser-5 phosphorylation very little but induced Ser-2 phosphorylation normally, in contrast to wild-type hTFIIE, which induced phosphorylation at both Ser-2 and Ser-5. In transcription transition assays using a linear template, ceTFIIEbeta was markedly defective in its ability to support the transition to elongation. These observations provide evidence of TFIIE involvement in the transition and suggest that Ser-5 phosphorylation is essential for Pol II to be in the processive elongation form. PMID- 11113178 TI - Insulin receptor substrate 3 (IRS-3) and IRS-4 impair IRS-1- and IRS-2-mediated signaling. AB - To investigate the roles of insulin receptor substrate 3 (IRS-3) and IRS-4 in the insulin-like growth factor 1 (IGF-1) signaling cascade, we introduced these proteins into 3T3 embryonic fibroblast cell lines prepared from wild-type (WT) and IRS-1 knockout (KO) mice by using a retroviral system. Following transduction of IRS-3 or IRS-4, the cells showed a significant decrease in IRS-2 mRNA and protein levels without any change in the IRS-1 protein level. In these cell lines, IGF-1 caused the rapid tyrosine phosphorylation of all four IRS proteins. However, IRS-3- or IRS-4-expressing cells also showed a marked decrease in IRS-1 and IRS-2 phosphorylation compared to the host cells. This decrease was accounted for in part by a decrease in the level of IRS-2 protein but occurred with no significant change in the IRS-1 protein level. IRS-3- or IRS-4-overexpressing cells showed an increase in basal phosphatidylinositol 3-kinase activity and basal Akt phosphorylation, while the IGF-1-stimulated levels correlated well with total tyrosine phosphorylation level of all IRS proteins in each cell line. IRS-3 expression in WT cells also caused an increase in IGF-1-induced mitogen-activated protein kinase phosphorylation and egr-1 expression ( approximately 1.8- and approximately 2.4-fold with respect to WT). In the IRS-1 KO cells, the impaired mitogenic response to IGF-1 was reconstituted with IRS-1 to supranormal levels and was returned to almost normal by IRS-2 or IRS-3 but was not improved by overexpression of IRS-4. These data suggest that IRS-3 and IRS-4 may act as negative regulators of the IGF-1 signaling pathway by suppressing the function of other IRS proteins at several steps. PMID- 11113179 TI - Analysis of the steroid receptor coactivator 1 (SRC1)-CREB binding protein interaction interface and its importance for the function of SRC1. AB - The transcriptional activity of nuclear receptors is mediated by coactivator proteins, including steroid receptor coactivator 1 (SRC1) and its homologues and the general coactivators CREB binding protein (CBP) and p300. SRC1 contains an activation domain (AD1) which functions via recruitment of CBP and and p300. In this study, we have used yeast two-hybrid and in vitro interaction-peptide inhibition experiments to map the AD1 domain of SRC1 to a 35-residue sequence potentially containing two alpha-helices. We also define a 72-amino-acid sequence in CBP necessary for SRC1 binding, designated the SRC1 interaction domain (SID). We show that in contrast to SRC1, direct binding of CBP to the estrogen receptor is weak, suggesting that SRC1 functions primarily as an adaptor to recruit CBP and p300. In support of this, we show that the ability of SRC1 to enhance ligand dependent nuclear receptor activity in transiently transfected cells is dependent upon the integrity of the AD1 region. In contrast, the putative histone acetyltransferase domain, the Per-Arnt-Sim basic helix-loop-helix domain, the glutamine-rich domain, and AD2 can each be removed without loss of ligand-induced activity. Remarkably, a construct corresponding to residues 631 to 970, which contains only the LXXLL motifs and the AD1 region of SRC1, retained strong coactivator activity in our assays. PMID- 11113180 TI - Ptc1, a type 2C Ser/Thr phosphatase, inactivates the HOG pathway by dephosphorylating the mitogen-activated protein kinase Hog1. AB - The HOG (high-osmolarity glycerol) mitogen-activated protein kinase (MAPK) pathway regulates the osmotic stress response in the yeast Saccharomyces cerevisiae. Three type 2C Ser/Thr phosphatases (PTCs), Ptc1, Ptc2, and Ptc3, have been isolated as negative regulators of this pathway. Previously, multicopy expression of PTC1 and PTC3 was shown to suppress lethality of the sln1Delta strain due to hyperactivation of the HOG pathway. In this work, we show that PTC2 also suppresses sln1Delta lethality. Furthermore, the phosphatase activity of these PTCs was needed for suppression, as mutation of a conserved Asp residue, likely to coordinate a metal ion, inactivated PTCs. Further analysis of Ptc1 function in vivo showed that it inactivates the MAPK, Hog1, but not the MEK, Pbs2. In the wild type, Hog1 kinase activity increased transiently, approximately 12-fold in response to osmotic stress, while overexpression of PTC1 limited activation to approximately 3-fold. In contrast, overexpression of PTC1 did not inhibit phosphorylation of Hog1 Tyr in the phosphorylation lip, suggesting that Ptc1 does not act on Pbs2. Deletion of PTC1 also strongly affected Hog1, leading to high basal Hog1 activity and sustained Hog1 activity in response to osmotic stress, the latter being consistent with a role for Ptc1 in adaptation. In vitro, Ptc1 but not the metal binding site mutant, Ptc1D58N, inactivated Hog1 by dephosphorylating the phosphothreonine but not the phosphotyrosine residue in the phosphorylation lip. Consistent with its role as a negative regulator of Hog1, which accumulates in the nucleus upon activation, Ptc1 was found in both the nucleus and the cytoplasm. Thus, one function of Ptc1 is to inactivate Hog1. PMID- 11113181 TI - Activation of NF-kappaB by double-stranded RNA (dsRNA) in the absence of protein kinase R and RNase L demonstrates the existence of two separate dsRNA-triggered antiviral programs. AB - Double-stranded RNA (dsRNA) of viral origin triggers two programs of the innate immunity in virus-infected cells. One is intended to decrease the rate of host cell protein synthesis and thus to prevent viral replication. This program is mediated by protein kinase R (PKR) and by RNase L and contributes, eventually, to the self-elimination of the infected cell via apoptosis. The second program is responsible for the production of antiviral (type I) interferons and other alarmone cytokines and serves the purpose of preparing naive cells for the viral invasion. This second program requires the survival of the infected cell and depends on the expression of antiapoptotic genes through the activation of the NF kappaB transcription factor. The second program therefore relies on ongoing transcription and translation. It has been proposed that PKR plays an essential role in the activation of NF-kappaB by dsRNA. Here we present evidence that the dsRNA-induced NF-kappaB activity and the expression of beta interferon and inflammatory cytokines do not require either PKR or RNase L. Our results indicate, therefore, that the two dsRNA-activated programs are separate and can function independently of each other. PMID- 11113182 TI - p45(NFE2) is a negative regulator of erythroid proliferation which contributes to the progression of Friend virus-induced erythroleukemias. AB - In previous studies, we identified a common site of retroviral integration designated Fli-2 in Friend murine leukemia virus (F-MuLV)-induced erythroleukemia cell lines. Insertion of F-MuLV at the Fli-2 locus, which was associated with the loss of the second allele, resulted in the inactivation of the erythroid cell- and megakaryocyte-specific gene p45(NFE2). Frequent disruption of p45(NFE2) due to proviral insertion suggests a role for this transcription factor in the progression of Friend virus-induced erythroleukemias. To assess this possibility, erythroleukemia was induced by F-MuLV in p45(NFE2) mutant mice. Since p45(NFE2) homozygous mice mostly die at birth, erythroleukemia was induced in +/- and +/+ mice. We demonstrate that +/- mice succumb to the disease moderately but significantly faster than +/+ mice. In addition, the spleens of +/- mice were significantly larger than those of +/+ mice. Of the 37 tumors generated from the +/- and +/+ mice, 10 gave rise to cell lines, all of which were derived from +/- mice. Establishment in culture was associated with the loss of the remaining wild type p45(NFE2) allele in 9 of 10 of these cell lines. The loss of a functional p45(NFE2) in these cell lines was associated with a marked reduction in globin gene expression. Expression of wild-type p45(NFE2) in the nonproducer erythroleukemic cells resulted in reduced cell growth and restored the expression of globin genes. Similarly, the expression of p45(NFE2) in these cells also slows tumor growth in vivo. These results indicate that p45(NFE2) functions as an inhibitor of erythroid cell growth and that perturbation of its expression contributes to the progression of Friend erythroleukemia. PMID- 11113183 TI - Altered extracellular signal-regulated kinase signaling and glycogen metabolism in skeletal muscle from p90 ribosomal S6 kinase 2 knockout mice. AB - The p90 ribosomal S6 kinase (RSK), a cytosolic substrate for the extracellular signal-regulated kinase (ERK), is involved in transcriptional regulation, and one isoform (RSK2) has been implicated in the activation of glycogen synthase by insulin. To determine RSK2 function in vivo, mice lacking a functional rsk2 gene were generated and studied in response to insulin and exercise, two potent stimulators of the ERK cascade in skeletal muscle. RSK2 knockout (KO) mice weigh 10% less and are 14% shorter than wild-type (WT) mice. They also have impaired learning and coordination. Hindlimb skeletal muscles were obtained from mice 10, 15, or 30 min after insulin injection or immediately after strenuous treadmill exercise for 60 min. While insulin and exercise significantly increased ERK phosphorylation in skeletal muscle from both WT and KO mice, the increases were twofold greater in the KO animals. This occurred despite 27% lower ERK2 protein expression in skeletal muscle of KO mice. KO mice had 18% less muscle glycogen in the fasted basal state, and insulin increased glycogen synthase activity more in KO than WT mice. The enhanced insulin-stimulated increases in ERK and glycogen synthase activities in KO mice were not associated with higher insulin receptor or with IRS1 tyrosine phosphorylation or with IRS1 binding to phosphatidylinositol 3-kinase. However, insulin-stimulated serine phosphorylation of Akt was significantly higher in the KO animals. c-fos mRNA was increased similarly in muscle from WT and KO mice in response to insulin (2. 5-fold) and exercise (15-fold). In conclusion, RSK2 likely plays a major role in feedback inhibition of the ERK pathway in skeletal muscle. Furthermore, RSK2 is not required for activation of muscle glycogen synthase by insulin but may indirectly modulate muscle glycogen synthase activity and/or glycogen content by other mechanisms, possibly through regulation of Akt. RSK2 knockout mice may be a good animal model for the study of Coffin-Lowry syndrome. PMID- 11113184 TI - Reciprocal activation by cyclin-dependent kinases 2 and 7 is directed by substrate specificity determinants outside the T loop. AB - Cyclin-dependent kinase 7 (CDK7) is the catalytic subunit of the metazoan CDK activating kinase (CAK), which activates CDKs, such as CDC2 and CDK2, through phosphorylation of a conserved threonine residue in the T loop. Full activation of CDK7 requires association with a positive regulatory subunit, cyclin H, and phosphorylation of a conserved threonine residue at position 170 in its own T loop. We show that threonine-170 of CDK7 is phosphorylated in vitro by its targets, CDC2 and CDK2, which also phosphorylate serine-164 in the CDK7 T loop, a site that perfectly matches their consensus phosphorylation site. In contrast, neither CDK4 nor CDK7 itself can phosphorylate the CDK7 T loop in vitro. The ability of CDC2 or CDK2 and CDK7 to phosphorylate each other but not themselves implies that each kinase can discriminate among closely related sequences and can recognize a substrate site that diverges from its usual preferred site. To understand the basis for this paradoxical substrate specificity, we constructed a chimeric CDK with the T loop of CDK7 grafted onto the body of CDK2. Surprisingly, the hybrid enzyme, CDK2-7, was efficiently activated in cyclin A-dependent fashion by CDK7 but not at all by CDK2. CDK2-7, moreover, phosphorylated wild type CDK7 but not CDK2. Our results suggest that the primary amino acid sequence of the T loop plays only a minor role, if any, in determining the specificity of cyclin-dependent CAKs for their CDK substrates and that protein-protein interactions involving sequences outside the T loop can influence substrate specificity both positively and negatively. PMID- 11113185 TI - Sequence-specific recognition and cleavage of telomeric repeat (TTAGG)(n) by endonuclease of non-long terminal repeat retrotransposon TRAS1. AB - The telomere of the silkworm Bombyx mori consists of (TTAGG/CCTAA)(n) repeats and harbors a large number of telomeric repeat-specific non-long terminal repeat retrotransposons, such as TRAS1 and SART1. To understand how these retrotransposons recognize and integrate into the telomeric repeat in a sequence specific manner, we expressed the apurinic-apryrimidinic endonuclease-like endonuclease domain of TRAS1 (TRAS1 EN), which is supposed to digest the target DNA, and characterized its enzymatic properties. Purified TRAS1 EN could generate specific nicks on both strands of the telomeric repeat sequence between T and A of the (TTAGG)(n) strand (bottom strand) and between C and T of the (CCTAA)(n) strand (top strand). These sites are consistent with insertion sites expected from the genomic structure of boundary regions of TRAS1. Time course studies of nicking activities on both strands revealed that the cleavages on the bottom strand preceded those on the top strand, supporting the target-primed reverse transcription model. TRAS1 EN could cleave the telomeric repeats specifically even if it was flanked by longer tracts of nontelomeric sequence, indicating that the target site specificity of the TRAS1 element was mainly determined by its EN domain. Based on mutation analyses, TRAS1 EN recognizes less than 10 bp around the initial cleavage site (upstream 7 bp and downstream 3 bp), and the GTTAG sequence especially is essential for the cleavage reaction on the bottom strand (5'. TTAGGTT downward arrow AGG. 3'). TRAS1 EN, the first identified endonuclease digesting telomeric repeats, may be used as a genetic tool to shorten the telomere in insects and some other organisms. PMID- 11113186 TI - Tsc13p is required for fatty acid elongation and localizes to a novel structure at the nuclear-vacuolar interface in Saccharomyces cerevisiae. AB - The TSC13/YDL015c gene was identified in a screen for suppressors of the calcium sensitivity of csg2Delta mutants that are defective in sphingolipid synthesis. The fatty acid moiety of sphingolipids in Saccharomyces cerevisiae is a very long chain fatty acid (VLCFA) that is synthesized by a microsomal enzyme system that lengthens the palmitate produced by cytosolic fatty acid synthase by two carbon units in each cycle of elongation. The TSC13 gene encodes a protein required for elongation, possibly the enoyl reductase that catalyzes the last step in each cycle of elongation. The tsc13 mutant accumulates high levels of long-chain bases as well as ceramides that harbor fatty acids with chain lengths shorter than 26 carbons. These phenotypes are exacerbated by the deletion of either the ELO2 or ELO3 gene, both of which have previously been shown to be required for VLCFA synthesis. Compromising the synthesis of malonyl coenzyme A (malonyl-CoA) by inactivating acetyl-CoA carboxylase in a tsc13 mutant is lethal, further supporting a role of Tsc13p in VLCFA synthesis. Tsc13p coimmunoprecipitates with Elo2p and Elo3p, suggesting that the elongating proteins are organized in a complex. Tsc13p localizes to the endoplasmic reticulum and is highly enriched in a novel structure marking nuclear-vacuolar junctions. PMID- 11113187 TI - Insertion of a telomere repeat sequence into a mammalian gene causes chromosome instability. AB - Telomere repeat sequences cap the ends of eucaryotic chromosomes and help stabilize them. At interstitial sites, however, they may destabilize chromosomes, as suggested by cytogenetic studies in mammalian cells that correlate interstitial telomere sequence with sites of spontaneous and radiation-induced chromosome rearrangements. In no instance is the length, purity, or orientation of the telomere repeats at these potentially destabilizing interstitial sites known. To determine the effects of a defined interstitial telomere sequence on chromosome instability, as well as other aspects of DNA metabolism, we deposited 800 bp of the functional vertebrate telomere repeat, TTAGGG, in two orientations in the second intron of the adenosine phosphoribosyltransferase (APRT) gene in Chinese hamster ovary cells. In one orientation, the deposited telomere sequence did not interfere with expression of the APRT gene, whereas in the other it reduced mRNA levels slightly. The telomere sequence did not induce chromosome truncation and the seeding of a new telomere at a frequency above the limits of detection. Similarly, the telomere sequence did not alter the rate or distribution of homologous recombination events. The interstitial telomere repeat sequence in both orientations, however, dramatically increased gene rearrangements some 30-fold. Analysis of individual rearrangements confirmed the involvement of the telomere sequence. These studies define the telomere repeat sequence as a destabilizing element in the interior of chromosomes in mammalian cells. PMID- 11113188 TI - Identification of DNA cis elements essential for expansion of ribosomal DNA repeats in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae carries approximately 150 ribosomal DNA (rDNA) copies in tandem repeats. Each repeat consists of the 35S rRNA gene, the NTS1 spacer, the 5S rRNA gene, and the NTS2 spacer. The FOB1 gene was previously shown to be required for replication fork block (RFB) activity at the RFB site in NTS1, for recombination hot spot (HOT1) activity, and for rDNA repeat expansion and contraction. We have constructed a strain in which the majority of rDNA repeats are deleted, leaving two copies of rDNA covering the 5S-NTS2-35S region and a single intact NTS1, and whose growth is supported by a helper plasmid carrying, in addition to the 5S rRNA gene, the 35S rRNA coding region fused to the GAL7 promoter. This strain carries a fob1 mutation, and an extensive expansion of chromosomal rDNA repeats was demonstrated by introducing the missing FOB1 gene by transformation. Mutational analysis using this system showed that not only the RFB site but also the adjacent approximately 400-bp region in NTS1 (together called the EXP region) are required for the FOB1-dependent repeat expansion. This approximately 400-bp DNA element is not required for the RFB activity or the HOT1 activity and therefore defines a function unique to rDNA repeat expansion (and presumably contraction) separate from HOT1 and RFB activities. PMID- 11113189 TI - Gene disruption of tissue transglutaminase. AB - Transglutaminase 2 (TGase 2), or tissue transglutaminase, catalyzes either epsilon-(gamma-glutamyl)lysine or N(1), N(8)-(gamma-glutamyl)spermidine isopeptide bonds. TGase 2 expression has been associated with apoptosis, and it has been proposed that its activation should lead to the irreversible assembly of a cross-linked protein scaffold in dead cells. Thus, TGase 2-catalyzed protein polymerization contributes to the ultrastructural changes typical of dying apoptotic cells; it stabilizes the integrity of the apoptotic cells, preventing the release of harmful intracellular components into the extracellular space and, consequently, inflammation and scar formation. In order to perform a targeted disruption of the enzyme, we prepared a construct deleting part of exons 5 and 6, containing the active site, and intron 5. Complete absence of TGase 2 was demonstrated by reverse transcription-PCR and Western blot analysis. TGase activity measured on liver and thymus extracts showed, however, a minimal residual activity in TGase 2(-/-) mice. PCR analysis of mRNA extracted from the same tissues demonstrated that at least TGase 1 (normally present in the skin) is also expressed in these tissues and contributes to this residual activity. TGase 2(-/-) mice showed no major developmental abnormalities, and histological examination of the major organs appeared normal. Induction of apoptosis ex vivo in TGase 2(-/-) thymocytes (by CD95, dexamethasone, etoposide, and H(2)O(2)) and in vitro on TGase 2(-/-) mouse embryonal fibroblasts (by retinoids, UV, and H(2)O(2)) showed no significant differences. A reduction in cross-linked apoptotic bodies with a modestly increased release of lactate dehydrogenase has been detected in some cases. Together our results show that TGase 2 is not a crucial component of the main pathway of the apoptotic program. It is possible that the residual enzymatic activity, due to TGase 1 or redundancy of other still unidentified TGases, can compensate for the lack of TGase 2. PMID- 11113190 TI - Oligomerization of ETO is obligatory for corepressor interaction. AB - Nearly 40% of cases of acute myelogenous leukemia (AML) of the M2 subtype are due to a chromosomal translocation that combines a sequence-specific DNA binding protein, AML1, with a potent transcriptional repressor, ETO. ETO interacts with nuclear receptor corepressors SMRT and N-CoR, which recruit histone deacetylase to the AML1-ETO oncoprotein. SMRT-N-CoR interaction requires each of two zinc fingers contained in C-terminal Nervy homology region 4 (NHR4) of ETO. However, here we show that polypeptides containing NHR4 are insufficient for interaction with SMRT. NHR2 is also required for SMRT interaction and repression by ETO, as well as for inhibition of hematopoietic differentiation by AML1-ETO. NHR2 mediates oligomerization of ETO as well as AML1-ETO. Fusion of NHR4 polypeptide to a heterologous dimerization domain allows strong interaction with SMRT in vitro. These data support a model in which NHR2 and NHR4 have complementary functions in repression by ETO. NHR2 functions as an oligomerization domain bringing together NHR4 polypeptides that together form the surface required for high-affinity interaction with corepressors. As nuclear receptors also interact with corepressors as dimers, oligomerization may be a common mechanism regulating corepressor interactions. PMID- 11113191 TI - A novel GATA factor transcriptionally represses yolk protein precursor genes in the mosquito Aedes aegypti via interaction with the CtBP corepressor. AB - In anautogenous mosquitoes, vitellogenesis, the key event in egg maturation, requires a blood meal. Consequently, mosquitoes are vectors of many devastating human diseases. An important adaptation for anautogenicity is the previtellogenic arrest (the state of arrest) preventing the activation of the yolk protein precursor (YPP) genes Vg and VCP prior to blood feeding. A novel GATA factor (AaGATAr) that recognizes GATA binding motifs (WGATAR) in the upstream region of the YPP genes serves as a transcriptional repressor at the state of arrest. Importantly, AaGATAr can override the 20-hydroxyecdysone transactivation of YPP genes, and its transcriptional repression involves the recruitment of CtBP, one of the universal corepressors. AaGATAr transcript is present only in the adult female fat body. Furthermore, in nuclear extracts of previtellogenic fat bodies with transcriptionally repressed YPP genes, there is a GATA binding protein forming a band with mobility similar to that of AaGATAr. The specific repression of YPP genes by AaGATAr in the fat body of the female mosquito during the state of arrest represents an important molecular adaptation for anautogenicity. PMID- 11113192 TI - Screening for modulators of spermine tolerance identifies Sky1, the SR protein kinase of Saccharomyces cerevisiae, as a regulator of polyamine transport and ion homeostasis. AB - Although most cells are capable of transporting polyamines, the mechanism that regulates polyamine transport in eukaryotes is still largely unknown. Using a genetic screen for clones capable of restoring spermine sensitivity to spermine tolerant mutants of Saccharomyces cerevisiae, we have demonstrated that Sky1p, a recently identified SR protein kinase, is a key regulator of polyamine transport. Yeast cells deleted for SKY1 developed tolerance to toxic levels of spermine, while overexpression of Sky1p in wild-type cells increased their sensitivity to spermine. Expression of the wild-type Sky1p but not of a catalytically inactive mutant restored sensitivity to spermine. SKY1 disruption results in dramatically reduced uptake of spermine, spermidine, and putrescine. In addition to spermine tolerance, sky1Delta cells exhibit increased tolerance to lithium and sodium ions but somewhat increased sensitivity to osmotic shock. The observed halotolerance suggests potential regulatory interaction between the transport of polyamines and inorganic ions, as suggested in the case of the Ptk2p, a recently described regulator of polyamine transport. We demonstrate that these two kinases act in two different signaling pathways. While deletion or overexpression of SKY1 did not significantly affect Pma1p activity, the ability of overexpressed Sky1p, Ptk1p, and Ptk2p to increase sensitivity to LiCl depends on the integrity of PPZ1 but not of ENA1. PMID- 11113193 TI - Requirement of DNA polymerase eta for error-free bypass of UV-induced CC and TC photoproducts. AB - The yeast RAD30-encoded DNA polymerase eta (Poleta) bypasses a cis-syn thymine thymine dimer efficiently and accurately. Human DNA polymerase eta functions similarly in the bypass of this lesion, and mutations in human Poleta result in the cancer prone syndrome, the variant form of xeroderma pigmentosum. UV light, however, also elicits the formation of cis-syn cyclobutane dimers and (6-4) photoproducts at 5'-CC-3' and 5'-TC-3' sites, and in both yeast and human DNA, UV induced mutations occur primarily by 3' C to T transitions. Genetic studies presented here reveal a role for yeast Poleta in the error-free bypass of cyclobutane dimers and (6-4) photoproducts formed at CC and TC sites. Thus, by preventing UV mutagenesis at a wide spectrum of dipyrimidine sites, Poleta plays a pivotal role in minimizing the incidence of sunlight-induced skin cancers in humans. PMID- 11113194 TI - Cross talk between tRNA and rRNA synthesis in Saccharomyces cerevisiae. AB - Temperature-sensitive RNA polymerase III (rpc160-112 and rpc160-270) mutants were analyzed for the synthesis of tRNAs and rRNAs in vivo, using a double-isotopic labeling technique in which cells are pulse-labeled with [(33)P]orthophosphate and coextracted with [(3)H]uracil-labeled wild-type cells. Individual RNA species were monitored by Northern blot hybridization or amplified by reverse transcription. These mutants impaired the synthesis of RNA polymerase III transcripts with little or no influence on mRNA synthesis but also largely turned off the formation of the 25S, 18S, and 5.8S mature rRNA species derived from the common 35S transcript produced by RNA polymerase I. In the rpc160-270 mutant, this parallel inhibition of tRNA and rRNA synthesis also occurred at the permissive temperature (25 degrees C) and correlated with an accumulation of 20S pre-rRNA. In the rpc160-112 mutant, inhibition of rRNA synthesis and the accumulation of 20S pre-rRNA were found only at 37 degrees C. The steady-state rRNA/tRNA ratio of these mutants reflected their tRNA and rRNA synthesis pattern: the rpc160-112 mutant had the threefold shortage in tRNA expected from its preferential defect in tRNA synthesis at 25 degrees C, whereas rpc160-270 cells completely adjusted their rRNA/tRNA ratio down to a wild-type level, consistent with the tight coupling of tRNA and rRNA synthesis in vivo. Finally, an RNA polymerase I (rpa190-2) mutant grown at the permissive temperature had an enhanced level of pre-tRNA, suggesting the existence of a physiological coupling between rRNA synthesis and pre-tRNA processing. PMID- 11113195 TI - Matrix attachment region-dependent function of the immunoglobulin mu enhancer involves histone acetylation at a distance without changes in enhancer occupancy. AB - Nuclear matrix attachment regions (MARs), which flank the immunoglobulin mu heavy chain enhancer on either side, are required for the activation of the distal variable-region (V(H)) promoter in transgenic mice. Previously, we have shown that the MARs extend a local domain of chromatin accessibility at the mu enhancer to more distal sites. In this report, we examine the influence of MARs on the formation of a nucleoprotein complex at the enhancer and on the acetylation of histones, which have both been implicated in contributing to chromatin accessibility. By in vivo footprint analysis of transgenic mu gene constructs, we show that the occupancy of factor-binding sites at the mu enhancer is similar in transcriptionally active wild-type and transcriptionally inactive DeltaMAR genes. Chromatin immunoprecipitation experiments indicate, however, that the acetylation of histones at enhancer-distal nucleosomes is enhanced 10-fold in the presence of MARs, whereas the levels of histone acetylation at enhancer-proximal nucleosomes are similar for wild-type and DeltaMAR genes. Taken together, these data indicate that the function of MARs in mediating long-range chromatin accessibility and transcriptional activation of the V(H) promoter involves the generation of an extended domain of histone acetylation, independent of changes in the occupancy of the mu enhancer. PMID- 11113196 TI - Identification and characterization of human orthologues to Saccharomyces cerevisiae Upf2 protein and Upf3 protein (Caenorhabditis elegans SMG-4). AB - Nonsense-mediated mRNA decay (NMD), also called mRNA surveillance, is an important pathway used by all organisms that have been tested to degrade mRNAs that prematurely terminate translation and, as a consequence, eliminate the production of aberrant proteins that could be potentially harmful. In mammalian cells, NMD appears to involve splicing-dependent alterations to mRNA as well as ribosome-associated components of the translational apparatus. To date, human (h) Upf1 protein (p) (hUpf1p), a group 1 RNA helicase named after its Saccharomyces cerevisiae orthologue that functions in both translation termination and NMD, has been the only factor shown to be required for NMD in mammalian cells. Here, we describe human orthologues to S. cerevisiae Upf2p and S. cerevisiae Upf3p (Caenorhabditis elegans SMG-4) based on limited amino acid similarities. The existence of these orthologues provides evidence for a higher degree of evolutionary conservation of NMD than previously appreciated. Interestingly, human orthologues to S. cerevisiae Upf3p (C. elegans SMG-4) derive from two genes, one of which is X-linked and both of which generate multiple isoforms due to alternative pre-mRNA splicing. We demonstrate using immunoprecipitations of epitope-tagged proteins transiently produced in HeLa cells that hUpf2p interacts with hUpf1p, hUpf3p-X, and hUpf3p, and we define the domains required for the interactions. Furthermore, we find by using indirect immunofluorescence that hUpf1p is detected only in the cytoplasm, hUpf2p is detected primarily in the cytoplasm, and hUpf3p-X localizes primarily to nuclei. The finding that hUpf3p-X is a shuttling protein provides additional indication that NMD has both nuclear and cytoplasmic components. PMID- 11113197 TI - Defining roles for HOX and MEIS1 genes in induction of acute myeloid leukemia. AB - Complex genetic and biochemical interactions between HOX proteins and members of the TALE (i.e., PBX and MEIS) family have been identified in embryonic development, and some of these interactions also appear to be important for leukemic transformation. We have previously shown that HOXA9 collaborates with MEIS1 in the induction of acute myeloid leukemia (AML). In this report, we demonstrate that HOXB3, which is highly divergent from HOXA9, also genetically interacts with MEIS1, but not with PBX1, in generating AML. In addition, we show that the HOXA9 and HOXB3 genes play key roles in establishing all the main characteristics of the leukemias, while MEIS1 functions only to accelerate the onset of the leukemic transformation. Contrasting the reported functional similarities between PREP1 and MEIS1, such as PBX nuclear retention, we also show that PREP1 overexpression is incapable of accelerating the HOXA9-induced AML, suggesting that MEIS1 function in transformation must entail more than PBX nuclear localization. Collectively, these data demonstrate that MEIS1 is a common leukemic collaborator with two structurally and functionally divergent HOX genes and that, in this collaboration, the HOX gene defines the identity of the leukemia. PMID- 11113199 TI - Biochemical and biological functions of the N-terminal, noncatalytic domain of extracellular signal-regulated kinase 2. AB - Extracellular signal-regulated kinase 1 (ERK1) and ERK2 are important components in signal transduction pathways involved in many cellular processes, including cell differentiation and proliferation. These proteins consist of a central kinase domain flanked by short N- and C-terminal noncatalytic domains. While the regulation of ERK2 by sequences within the kinase domain has been extensively studied, little is known about the small regions outside of the kinase domain. We performed mutational analysis on the N-terminal, noncatalytic domain of ERK2 in an attempt to determine its role in ERK2 function and regulation. Deleting or mutating amino acids 19 to 25 (ERK2-Delta19-25) created an ERK2 molecule that could be phosphorylated in response to growth factor and serum stimulation in a MEK (mitogen-activated protein kinase kinase or ERK kinase)-dependent manner but had little kinase activity and was unable to bind to MEK in vivo. Since MEK acts as a cytoplasmic anchor for the ERKs, the lack of a MEK interaction resulted in the aberrant nuclear localization of ERK2-Delta19-25 mutants in serum-starved cells. Assaying these mutants for their ability to affect ERK signaling revealed that ERK2-Delta19-25 mutants acted in a dominant-negative manner to inhibit transcriptional signaling through endogenous ERKs to an Elk1-responsive promoter in transfected COS-1 cells. However, ERK2-Delta19-25 had no effect on the phosphorylation of RSK2, an ERK2 cytoplasmic substrate, whereas a nonactivatable ERK (T183A) that retained these sequences could inhibit RSK2 phosphorylation. These results suggest that the N-terminal domain of ERK2 profoundly affects ERK2 localization, MEK binding, kinase activity, and signaling and identify a novel dominant-negative mutant of ERK2 that can dissociate at least some transcriptional responses from cytoplasmic responses. PMID- 11113198 TI - Different domains of the essential GTPase Cdc42p required for growth and development of Saccharomyces cerevisiae. AB - In budding yeast, the Rho-type GTPase Cdc42p is essential for cell division and regulates pseudohyphal development and invasive growth. Here, we isolated novel Cdc42p mutant proteins with single-amino-acid substitutions that are sufficient to uncouple functions of Cdc42p essential for cell division from regulatory functions required for pseudohyphal development and invasive growth. In haploid cells, Cdc42p is able to regulate invasive growth dependent on and independent of FLO11 gene expression. In diploid cells, Cdc42p regulates pseudohyphal development by controlling pseudohyphal cell (PH cell) morphogenesis and invasive growth. Several of the Cdc42p mutants isolated here block PH cell morphogenesis in response to nitrogen starvation without affecting morphology or polarity of yeast form cells in nutrient-rich conditions, indicating that these proteins are impaired for certain signaling functions. Interaction studies between development specific Cdc42p mutants and known effector proteins indicate that in addition to the p21-activated (PAK)-like protein kinase Ste20p, the Cdc42p/Rac-interactive binding domain containing Gic1p and Gic2p proteins and the PAK-like protein kinase Skm1p might be further effectors of Cdc42p that regulate pseudohyphal and invasive growth. PMID- 11113200 TI - MAT1-modulated CAK activity regulates cell cycle G(1) exit. AB - The cyclin-dependent kinase (CDK)-activating kinase (CAK) is involved in cell cycle control, transcription, and DNA repair (E. A. Nigg, Curr. Opin. Cell. Biol. 8:312-317, 1996). However, the mechanisms of how CAK is integrated into these signaling pathways remain unknown. We previously demonstrated that abrogation of MAT1 (menage a trois 1), an assembly factor and targeting subunit of CAK, induces G(1) arrest (L. Wu, P. Chen, J. J. Hwang, L. W. Barsky, K. I. Weinberg, A. Jong, and V. A. Starnes, J. Biol. Chem. 274:5564-5572, 1999). This result led us to investigate how deregulation of CAK by MAT1 abrogation affects the cell cycle G(1) exit, a process that is regulated most closely by phosphorylation of retinoblastoma tumor suppressor protein (pRb). Using mammalian cellular models that undergo G(1) arrest evoked by antisense MAT1 abrogation, we found that deregulation of CAK inhibits pRb phosphorylation and cyclin E expression, CAK phosphorylation of pRb is MAT1 dose dependent but cyclin D1/CDK4 independent, and MAT1 interacts with pRb. These results suggest that CAK is involved in the regulation of cell cycle G(1) exit while MAT1-modulated CAK formation and CAK phosphorylation of pRb may determine the cell cycle specificity of CAK in G(1) progression. PMID- 11113201 TI - Wsc1 and Mid2 are cell surface sensors for cell wall integrity signaling that act through Rom2, a guanine nucleotide exchange factor for Rho1. AB - Wsc1 and Mid2 are highly O-glycosylated cell surface proteins that reside in the plasma membrane of Saccharomyces cerevisiae. They have been proposed to function as mechanosensors of cell wall stress induced by wall remodeling during vegetative growth and pheromone-induced morphogenesis. These proteins are required for activation of the cell wall integrity signaling pathway that consists of the small G-protein Rho1, protein kinase C (Pkc1), and a mitogen activated protein kinase cascade. We show here by two-hybrid experiments that the C-terminal cytoplasmic domains of Wsc1 and Mid2 interact with Rom2, a guanine nucleotide exchange factor (GEF) for Rho1. At least with regard to Wsc1, this interaction is mediated by the Rom2 N-terminal domain. This domain is distinct from the Rho1-interacting domain, suggesting that the GEF can interact simultaneously with a sensor and with Rho1. We also demonstrate that extracts from wsc1 and mid2 mutants are deficient in the ability to catalyze GTP loading of Rho1 in vitro, providing evidence that the function of the sensor-Rom2 interaction is to stimulate nucleotide exchange toward this G-protein. In a related line of investigation, we identified the PMT2 gene in a genetic screen for mutations that confer an additive cell lysis defect with a wsc1 null allele. Pmt2 is a member of a six-protein family in yeast that catalyzes the first step in O mannosylation of target proteins. We demonstrate that Mid2 is not mannosylated in a pmt2 mutant and that this modification is important for signaling by Mid2. PMID- 11113202 TI - DNA damage-dependent nuclear dynamics of the Mre11 complex. AB - The Mre11 complex has been implicated in diverse aspects of the cellular response to DNA damage. We used in situ fractionation of human fibroblasts to carry out cytologic analysis of Mre11 complex proteins in the double-strand break (DSB) response. In situ fractionation removes most nucleoplasmic protein, permitting immunofluorescent localization of proteins that become more avidly bound to nuclear structures after induction of DNA damage. We found that a fraction of the Mre11 complex was bound to promyelocyte leukemia protein bodies in undamaged cells. Within 10 min after gamma irradiation, nuclear retention of the Mre11 complex in small granular foci was observed and persisted until 2 h postirradiation. In light of the previous demonstration that the Mre11 complex associated with ionizing radiation (IR)-induced DSBs, we infer that the protein retained under these conditions was associated with DNA damage. We also observed increased retention of Rad51 following IR treatment, although IR induced Rad51 foci were distinct from Mre11 foci. The ATM kinase, which phosphorylates Nbs1 during activation of the S-phase checkpoint, was not required for the Mre11 complex to associate with DNA damage. These data suggest that the functions of the Mre11 complex in the DSB response are implicitly dependent upon its ability to detect DNA damage. PMID- 11113203 TI - Stimulation of homologous recombination through targeted cleavage by chimeric nucleases. AB - Chimeric nucleases that are hybrids between a nonspecific DNA cleavage domain and a zinc finger DNA recognition domain were tested for their ability to find and cleave their target sites in living cells. Both engineered DNA substrates and the nucleases were injected into Xenopus laevis oocyte nuclei, in which DNA cleavage and subsequent homologous recombination were observed. Specific cleavage required two inverted copies of the zinc finger recognition site in close proximity, reflecting the need for dimerization of the cleavage domain. Cleaved DNA molecules were activated for homologous recombination; in optimum conditions, essentially 100% of the substrate recombined, even though the DNA was assembled into chromatin. The original nuclease has an 18-amino-acid linker between the zinc finger and cleavage domains, and this enzyme cleaved in oocytes at paired sites separated by spacers in the range of 6 to 18 bp, with a rather sharp optimum at 8 bp. By shortening the linker, we found that the range of effective site separations could be narrowed significantly. With no intentional linker between the binding and cleavage domains, only binding sites exactly 6 bp apart supported efficient cleavage in oocytes. We also showed that two chimeric enzymes with different binding specificities could collaborate to stimulate recombination when their individual sites were appropriately placed. Because the recognition specificity of zinc fingers can be altered experimentally, this approach holds great promise for inducing targeted recombination in a variety of organisms. PMID- 11113204 TI - Position effects are influenced by the orientation of a transgene with respect to flanking chromatin. AB - We have inserted two expression cassettes at tagged reference chromosomal sites by using recombinase-mediated cassette exchange in mammalian cells. The three sites of integration displayed either stable or silencing position effects that were dominant over the different enhancers present in the cassettes. These position effects were strongly dependent on the orientation of the construct within the locus, with one orientation being permissive for expression and the other being nonpermissive. Orientation-specific silencing, which was observed at two of the three site tested, was associated with hypermethylation but not with changes in chromatin structure, as judged by DNase I hypersensitivity assays. Using CRE recombinase, we were able to switch in vivo the orientation of the transgenes from the permissive to the nonpermissive orientation and vice versa. Switching from the permissive to the nonpermissive orientation led to silencing, but switching from the nonpermissive to the permissive orientation did not lead to reactivation of the transgene. Instead, transgene expression occurred dynamically by transcriptional oscillations, with 10 to 20% of the cells expressing at any given time. This result suggested that the cassette had been imprinted (epigenetically tagged) while it was in the nonpermissive orientation. Methylation analysis revealed that the methylation state of the inverted cassettes resembled that of silenced cassettes except that the enhancer had selectively lost some of its methylation. Sorting of the expressing and nonexpressing cell populations provided evidence that the transcriptional oscillations of the epigenetically tagged cassette are associated with changes in the methylation status of regulatory elements in the transgene. This suggests that transgene methylation is more dynamic than was previously assumed. PMID- 11113205 TI - Efficiency alleles of the Pctr1 modifier locus for plasmacytoma susceptibility. AB - The susceptibility of BALB/c mice to pristane-induced plasmacytomas is a complex genetic trait involving multiple loci, while DBA/2 and C57BL/6 strains are genetically resistant to the plasmacytomagenic effects of pristane. In this model system for human B-cell neoplasia, one of the BALB/c susceptibility and modifier loci, Pctr1, was mapped to a 5.7-centimorgan (cM) chromosomal region that included Cdkn2a, which encodes p16(INK4a) and p19(ARF), and the coding sequences for the BALB/c p16(INK4a) and p19(ARF) alleles were found to be polymorphic with respect to their resistant Pctr1 counterparts in DBA/2 and C57BL/6 mice (45). In the present study, alleles of Pctr1, Cdkn2a, and D4Mit15 from a resistant strain (BALB/cDAG) carrying DBA/2 chromatin were introgressively backcrossed to the susceptible BALB/c strain. The resultant C.DAG-Pctr1 Cdkn2a D4Mit15 congenic was more resistant to plasmacytomagenesis than BALB/c, thus narrowing Pctr1 to a 1.5 cM interval. Concomitantly, resistant C57BL/6 mice, from which both gene products of the Cdkn2a gene have been eliminated, developed pristane-induced plasma cell tumors over a shorter latency period than the traditionally susceptible BALB/cAn strain. Biological assays of the p16(INK4a) and p19(ARF) alleles from BALB/c and DBA/2 indicated that the BALB/c p16(INK4a) allele was less active than its DBA/2 counterpart in inducing growth arrest of mouse plasmacytoma cell lines and preventing ras-induced transformation of NIH 3T3 cells, while the two p19(ARF) alleles displayed similar potencies in both assays. We propose that the BALB/c susceptibility/modifier locus, Pctr1, is an "efficiency" allele of the p16(INK4a) gene. PMID- 11113206 TI - Essential role of insulin receptor substrate 1 in differentiation of brown adipocytes. AB - The most widely distributed members of the family of insulin receptor substrate (IRS) proteins are IRS-1 and IRS-2. These proteins participate in insulin and insulin-like growth factor 1 signaling, as well as the actions of some cytokines, growth hormone, and prolactin. To more precisely define the specific role of IRS 1 in adipocyte biology, we established brown adipocyte cell lines from wild-type and IRS-1 knockout (KO) animals. Using differentiation protocols, both with and without insulin, preadipocyte cell lines derived from IRS-1 KO mice exhibited a marked decrease in differentiation and lipid accumulation (10 to 40%) compared to wild-type cells (90 to 100%). Furthermore, IRS-1 KO cells showed decreased expression of adipogenic marker proteins, such as peroxisome proliferator activated receptor gamma (PPARgamma), CCAAT/enhancer-binding protein alpha (C/EBPalpha), fatty acid synthase, uncoupling protein-1, and glucose transporter 4. The differentiation deficit in the KO cells could be reversed almost completely by retrovirus-mediated reexpression of IRS-1, PPARgamma, or C/EBPalpha but not the thiazolidinedione troglitazone. Phosphatidylinositol 3-kinase (PI 3 kinase) assays performed at various stages of the differentiation process revealed a strong and transient activation in IRS-1, IRS-2, and phosphotyrosine associated PI 3-kinase in the wild-type cells, whereas the IRS-1 KO cells showed impaired phosphotyrosine-associated PI 3-kinase activation, all of which was associated with IRS-2. Akt phosphorylation was reduced in parallel with the total PI 3-kinase activity. Inhibition of PI 3-kinase with LY294002 blocked differentiation of wild-type cells. Thus, IRS-1 appears to be an important mediator of brown adipocyte maturation. Furthermore, this signaling molecule appears to exert its unique role in the differentiation process via activation of PI 3-kinase and its downstream target, Akt, and is upstream of the effects of PPARgamma and C/EBPalpha. PMID- 11113207 TI - Inhibition of the Wnt signaling pathway by Idax, a novel Dvl-binding protein. AB - In attempting to clarify the roles of Dvl in the Wnt signaling pathway, we identified a novel protein which binds to the PDZ domain of Dvl and named it Idax (for inhibition of the Dvl and Axin complex). Idax and Axin competed with each other for the binding to Dvl. Immunocytochemical analyses showed that Idax was localized to the same place as Dvl in cells and that expression of Axin inhibited the colocalization of Dvl and Idax. Further, Wnt-induced accumulation of beta catenin and activation of T-cell factor in mammalian cells were suppressed by expression of Idax. Expression of Idax in Xenopus embryos induced ventralization with a reduction in the expression of siamois, a Wnt-inducible gene. Idax inhibited Wnt- and Dvl- but not beta-catenin-induced axis duplication. It is known that Dvl is a positive regulator in the Wnt signaling pathway and that the PDZ domain is important for this activity. Therefore, these results suggest that Idax functions as a negative regulator of the Wnt signaling pathway by directly binding to the PDZ domain of Dvl. PMID- 11113208 TI - CIA, a novel estrogen receptor coactivator with a bifunctional nuclear receptor interacting determinant. AB - Coregulators for nuclear receptors (NR) are factors that either enhance or repress their transcriptional activity. Both coactivators and corepressors have been shown to use similar but functionally distinct NR interacting determinants containing the core motifs LxxLL and PhixxPhiPhi, respectively. These interactions occur through a hydrophobic cleft located on the surface of the ligand-binding domain (LBD) of the NR and are regulated by ligand-dependent activation function 2 (AF-2). In an effort to identify novel coregulators that function independently of AF-2, we used the LBD of the orphan receptor RVR (which lacks AF-2) as bait in a yeast two-hybrid screen. This strategy led to the cloning of a nuclear protein referred to as CIA (coactivator independent of AF-2 function) that possesses both repressor and activator functions. Strikingly, we observed that CIA not only interacts with RVR and Rev-ErbAalpha in a ligand independent manner but can also form complexes with estrogen receptor alpha (ERalpha) and ERbeta in vitro and enhances ERalpha transcriptional activity in the presence of estradiol (E(2)). CIA-ERalpha interactions were found to be independent of AF-2 and enhanced by the antiestrogens EM-652 and ICI 182,780 but not by 4-hydroxytamoxifen and raloxifene. We further demonstrate that CIA-ERalpha interactions require the presence within CIA of a novel bifunctional NR recognition determinant containing overlapping LxxLL and PhixxPhiPhi motifs. The identification and functional characterization of CIA suggest that hormone binding can create a functional coactivator interaction interface in the absence of AF-2. PMID- 11113209 TI - Translational and structural requirements of the early nodulin gene enod40, a short-open reading frame-containing RNA, for elicitation of a cell-specific growth response in the alfalfa root cortex. AB - A diversity of mRNAs containing only short open reading frames (sORF-RNAs; encoding less than 30 amino acids) have been shown to be induced in growth and differentiation processes. The early nodulin gene enod40, coding for a 0.7-kb sORF-RNA, is expressed in the nodule primordium developing in the root cortex of leguminous plants after infection by symbiotic bacteria. Ballistic microtargeting of this gene into Medicago roots induced division of cortical cells. Translation of two sORFs (I and II, 13 and 27 amino acids, respectively) present in the conserved 5' and 3' regions of enod40 was required for this biological activity. These sORFs may be translated in roots via a reinitiation mechanism. In vitro translation products starting from the ATG of sORF I were detectable by mutating enod40 to yield peptides larger than 38 amino acids. Deletion of a Medicago truncatula enod40 region between the sORFs, spanning a predicted RNA structure, did not affect their translation but resulted in significantly decreased biological activity. Our data reveal a complex regulation of enod40 action, pointing to a role of sORF-encoded peptides and structured RNA signals in developmental processes involving sORF-RNAs. PMID- 11113210 TI - A therapeutic target and biomarker in Friedreich's ataxia. PMID- 11113211 TI - New tests for dementia. PMID- 11113212 TI - Population norms for the MMSE in the very old: estimates based on longitudinal data. Mini-Mental State Examination. AB - OBJECTIVE: To report the percentile distribution of Mini-Mental State Examination (MMSE) scores in older people by age, sex, and education level, estimated from longitudinal data, after correcting for loss due to dropout. METHODS: The Cambridge City over 75 Cohort is a population-based study of a cohort of 2106 subjects age 75 years and older at study entry followed up over 9 years. At each of the four waves, cognitive function was assessed using MMSE. Based on these data, the relationship between age and MMSE score was modeled. Percentile distributions by age, sex, and education level were provided using inverse probability weighting to correct for dropouts. RESULTS: Performance on MMSE was related to age in men and women. In women, at age 75, MMSE score ranged from 21 (10th percentile) to 29 (90th percentile). At age 95, the range was 10 (10th percentile) to 27 (90th percentile). The upper end of MMSE distribution was slightly modified with age, whereas the lower end of the distribution was very sensitive to age effect. A similar pattern was observed in both sexes. CONCLUSION: These findings provide norms for MMSE scores in subjects age 75 years and older from longitudinal population-based data. Such norms can be used as reference values to determine where an individual's score lies in relation to his or her age, sex, and education level. PMID- 11113213 TI - A brief cognitive test battery to differentiate Alzheimer's disease and frontotemporal dementia. AB - OBJECTIVES: To validate a simple bedside test battery designed to detect mild dementia and differentiate AD from frontotemporal dementia (FTD). METHODS: Addenbrooke's Cognitive Examination (ACE) is a 100-point test battery that assesses six cognitive domains. Of 210 new patients attending a memory clinic, 139 fulfilled inclusion criteria and comprised dementia (n = 115) and nondementia (n = 24) groups. The composite and the component scores on the ACE for the two groups were compared with those of 127 age- and education-matched controls. Norms and the probability of diagnosing dementia at different prevalence rates were calculated. To evaluate the ACE's ability to differentiate early AD from FTD, scores of the cases diagnosed with dementia with a Clinical Dementia Rating < or = 1 (AD = 56, FTD = 24, others = 20) were compared. RESULTS: Two cut-off values for the ACE composite score (88 and 83) were of optimal utility depending on the target population. The ACE had high reliability, construct validity, and sensitivity (93%, using 88 as cut-off). Using the lower cut-off of 83, the ACE had a higher sensitivity (82%) and predictive value than the Mini-Mental State Examination for a wide range of dementia prevalence. The ACE differentiated AD from FTD, and the VLOM ratio (derived using component scores: [verbal fluency + language]/[orientation + memory]) of <2.2 for FTD and >3.2 for AD was highly discriminating. CONCLUSION: The ACE is a brief and reliable bedside instrument for early detection of dementia, and offers a simple objective index to differentiate AD and FTD in mildly demented patients. PMID- 11113214 TI - The FAB: a Frontal Assessment Battery at bedside. AB - OBJECTIVE: To devise a short bedside cognitive and behavioral battery to assess frontal lobe functions. METHODS: The designed battery consists of six subtests exploring the following: conceptualization, mental flexibility, motor programming, sensitivity to interference, inhibitory control, and environmental autonomy. It takes approximately 10 minutes to administer. The authors studied 42 normal subjects and 121 patients with various degrees of frontal lobe dysfunction (PD, n = 24; multiple system atrophy, n = 6; corticobasal degeneration, n = 21; progressive supranuclear palsy, n = 47; frontotemporal dementia, n = 23). RESULTS: The Frontal Assessment Battery scores correlated with the Mattis Dementia Rating Scale scores (rho = 0.82, p < 0.01) and with the number of criteria (rho = 0.77, p < 0.01) and perseverative errors (rho = 0.68, p < 0.01) of the Wisconsin Card Sorting Test. These variables accounted for 79% of the variance in a stepwise multiple regression, whereas age or Mini-Mental State Examination scores had no significant influence. There was good interrater reliability (kappa = 0.87, p < 0.001), internal consistency (Cronbach's coefficient alpha = 0.78), and discriminant validity (89.1% of cases correctly identified in a discriminant analysis of patients and controls). CONCLUSION: The Frontal Assessment Battery is easy to administer at bedside and is sensitive to frontal lobe dysfunction. PMID- 11113216 TI - Early DAT is distinguished from aging by high-dimensional mapping of the hippocampus. Dementia of the Alzheimer type. AB - OBJECTIVE: To determine the feasibility of using high-dimensional brain mapping (HDBM) to assess the structure of the hippocampus in older human subjects, and to compare measurements of hippocampal volume and shape in subjects with early dementia of the Alzheimer type (DAT) and in healthy elderly and younger control subjects. BACKGROUND: HDBM represents the typical structures of the brain via the construction of templates and addresses their variability by probabilistic transformations applied to the templates. Local application of the transformations throughout the brain (i.e., high dimensionality) makes HDBM especially valuable for defining subtle deformities in brain structures such as the hippocampus. METHODS: MR scans were obtained in 18 subjects with very mild DAT, 18 healthy elderly subjects, and 15 healthy younger subjects. HDBM was used to obtain estimates of left and right hippocampal volume and eigenvectors that represented the principal dimensions of hippocampal shape differences among the subject groups. RESULTS: Hippocampal volume loss and shape deformities observed in subjects with DAT distinguished them from both elderly and younger control subjects. The pattern of hippocampal deformities in subjects with DAT was largely symmetric and suggested damage to the CA1 hippocampal subfield. Hippocampal shape changes were also observed in healthy elderly subjects, which distinguished them from healthy younger subjects. These shape changes occurred in a pattern distinct from the pattern seen in DAT and were not associated with substantial volume loss. CONCLUSIONS: Assessments of hippocampal volume and shape derived from HDBM may be useful in distinguishing early DAT from healthy aging. PMID- 11113215 TI - Hippocampal and cortical atrophy predict dementia in subcortical ischemic vascular disease. AB - BACKGROUND: The cause of dementia in subcortical ischemic vascular disease (SIVD) is controversial. OBJECTIVES: To determine whether cognitive impairment in SIVD 1) correlates with measures of ischemic brain injury or brain atrophy, and/or 2) is due to concomitant AD. METHODS: Volumetric MRI of the brain was performed in 1) elderly subjects with lacunes (L) and a spectrum of cognitive impairment normal cognition (NC+L, n = 32), mild cognitive impairment (CI+L, n = 26), and dementia (D+L, n = 29); 2) a comparison group with probable AD (n = 28); and 3) a control group with normal cognition and no lacunes (NC). The authors examined the relationship between the severity of cognitive impairment and 1) volume, number, and location of lacunes; 2) volume of white matter signal hyperintensities (WMSH); and 3) measures of brain atrophy (i. e., hippocampal, cortical gray matter, and CSF volumes). RESULTS: Among the three lacune groups, severity of cognitive impairment correlated with atrophy of the hippocampus and cortical gray matter, but not with any lacune measure. Although hippocampal atrophy was the best predictor of severity of cognitive impairment, there was evidence for a second, partially independent, atrophic process associated with ventricular dilation, cortical gray matter atrophy, and increase in WMSH. Eight autopsied SIVD cases showed variable severity of ischemic and neurofibrillary degeneration in the hippocampus, but no significant AD pathology in neocortex. The probable AD group gave evidence of only one atrophic process, reflected in the severity of hippocampal atrophy. Comparison of regional neocortical gray matter volumes showed sparing of the primary motor and visual cortices in the probable AD group, but relatively uniform atrophy in the D+L group. CONCLUSIONS: Dementia in SIVD, as in AD, correlates best with hippocampal and cortical atrophy, rather than any measure of lacunes. In SIVD, unlike AD, there is evidence for partial independence between these two atrophic processes. Hippocampal atrophy may result from a mixture of ischemic and degenerative pathologies. The cause of diffuse cortical atrophy is not known, but may be partially indexed by the severity of WMSH. PMID- 11113217 TI - Effect of APOE genotype and promoter polymorphism on risk of Alzheimer's disease. AB - OBJECTIVE: To verify the association between APOE gene promoter polymorphisms and the development of AD and to determine whether the effect of promoter polymorphisms on AD is independent of the APOE epsilon4 allele. BACKGROUND: Three polymorphisms in the APOE promoter have been shown to modify APOE expression in vitro. Several studies have suggested that these polymorphisms may also modulate risk for AD, either independently or by modifying the effect of the APOE coding polymorphism. METHODS: The authors analyzed allele and genotype distributions for APOE and all three known APOE promoter polymorphisms (-491 A/T, -427 T/C, and 219 G/T) in a study group consisting of 237 subjects with AD and 274 age-matched controls. They then used log-linear and logistic regression analyses to test for possible interactions between APOE genotype and the promoter polymorphisms on risk of AD. CONCLUSION: A strong association between the APOE epsilon 4 allele and AD was detected regardless of promoter polymorphism status. In addition, the 491 AA genotype appears to be an independent genetic risk factor for AD. The -427 T/C polymorphism and the -219 T/G polymorphism were not directly associated with AD. PMID- 11113218 TI - Early stroke treatment associated with better outcome: the NINDS rt-PA stroke study. AB - BACKGROUND: The National Institute of Neurological Disorders and Stroke (NINDS) rt-PA Stroke Study showed a similar percentage of intracranial hemorrhage and good outcome in patients 3 months after stroke treatment given 0 to 90 minutes and 91 to 180 minutes after stroke onset. At 24 hours after stroke onset more patients treated 0 to 90 compared to 91 to 180 minutes after stroke onset had improved by four or more points on the NIH Stroke Scale (NIHSS). The authors performed further analyses to characterize the relationship of onset-to-treatment time (OTT) to outcome at 3 months, early improvement at 24 hours, and intracranial hemorrhage within 36 hours. METHODS: Univariate analyses identified potentially confounding variables associated with OTT that could mask an OTT treatment interaction. Tests for OTT-treatment interactions adjusting for potential masking confounders were performed. An OTT-treatment interaction was considered significant if p < or = 0.10, implying that treatment effectiveness was related to OTT. RESULTS: For 24-hour improvement, there were no masking confounders identified and there was an OTT-treatment interaction (p = 0.08). For 3-month favorable outcome, the NIHSS met criteria for a masking confounder. After adjusting for NIHSS as a covariate, an OTT-treatment interaction was detected (p = 0.09): the adjusted OR (95% CI) for a favorable 3-month outcome associated with recombinant tissue-type plasminogen activator (rt-PA) was 2.11 (1.33 to 3.35) in the 0 to 90 minute stratum and 1.69 (1.09 to 2.62) in the 91 to 180 minute stratum. In the group treated with rt-PA, after adjusting for baseline NIHSS, an effect of OTT on the occurrence of intracranial hemorrhage was not detected. CONCLUSIONS: If the NINDS rt-PA Stroke Trial treatment protocol is followed, this analysis suggests that patients treated 0 to 90 minutes from stroke onset with rt PA have an increased odds of improvement at 24 hours and favorable 3-month outcome compared to patients treated later than 90 minutes. No effect of OTT on intracranial hemorrhage was detected within the group treated with rt-PA, possibly due to low power. PMID- 11113219 TI - Rebleeding, secondary ischemia, and timing of operation in patients with subarachnoid hemorrhage. AB - OBJECTIVE: To assess the time course of secondary ischemia and first rebleeding and the relation between the timing of operation and the time course of secondary ischemia in a consecutive series of patients with aneurysmal subarachnoid hemorrhage (SAH). METHODS: Life table methods were used to assess the daily rates of ischemia and of rebleeding on day 0, day 1 to 3, day 4 to 10, day 11 to 14, and day 15 to 21. The authors compared the time course of secondary ischemia between patients operated within 4 days of SAH and those operated after 10 days. RESULTS: Of 346 patients included, 220 were operated, 131 within 4 days and 74 after 10 days. The rebleed rate was highest on the day of the initial hemorrhage, then diminished, and increased slightly again during the second week. The rate of secondary ischemia was highest on day 4, diminished after day 10, but peaked again from day 14 to 18 for patients who were operated later than 10 days after aneurysmal rupture. The peak rate of ischemia was much higher after early than after late operation. Although patients with early operation were in a better clinical condition on admission, with a relatively low risk of secondary ischemia, the overall rate of secondary ischemia was as high as in patients with delayed operation. From day 11 to 21 the rebleed rate was higher than the rate of secondary ischemia. CONCLUSIONS: These results indicate that operation is a risk factor for ischemia, especially when performed early. If operation is postponed, it should be planned soon after day 10, because of the relatively high rebleed rate from day 11 to 21. PMID- 11113220 TI - Familial occipital calcifications, hemorrhagic strokes, leukoencephalopathy, dementia, and external carotid dysplasia. AB - OBJECTIVE: To describe a new familial association of late-onset dementia, patchy leukoencephalopathy, intracerebral hemorrhages, bilateral occipital calcifications (BOC), and external carotid artery dysplasia (ECAD). METHODS: At age 62, the proband, who was of Spanish descent, had left temporal hemorrhage in a background of progressive mental deterioration. Neuroimaging revealed fine tram line BOC, extensive leukoencephalopathy, and bilateral ECAD. Biologic screening for celiac disease was negative. Skin biopsy with ultrastructural study revealed heretofore unreported changes in the basal lamina of capillaries, with multilayered appearance and round-shaped microcalcifications. Of 19 next-of-kin who survived beyond 60 years of age, six had brain disorders; four of the six presented at least three components of the syndrome. The proband's mother had died at age 83 with profound dementia; one sister, who was diagnosed with dementia with BOC and leukoencephalopathy at age 67, died 2 years later from intracerebral hemorrhage; a brother had an occipital hemorrhage at age 58, at which time BOC and leukoencephalopathy were discovered; and another brother died after a minor stroke at age 70 with dementia, leukoaraiosis, BOC, and ECAD. A proband's cousin also had an unexplained ischemic stroke at age 55, but without other features of the entity. In no subject was there evidence of seizures, facial angioma, or intracranial vascular malformation, and arterial hypertension was neither constant nor severe. CONCLUSION: These clinical, neuroradiologic, and histologic features suggest a new familial cerebrovascular entity with widespread microvascular calcifications and autosomal (presumably dominant) inheritance. We suggest the acronym FOCHS-LADD, for familial occipital calcifications, hemorrhagic strokes, leukoencephalopathy, arterial dysplasia, and dementia. PMID- 11113221 TI - Subtraction peri-ictal SPECT is predictive of extratemporal epilepsy surgery outcome. AB - OBJECTIVES: To determine whether localization of extratemporal epilepsy with subtraction ictal SPECT coregistered with MRI (SISCOM) is predictive of outcome after resective epilepsy surgery, whether SISCOM images provide prognostically important information compared with standard tests, and whether blood flow change on SISCOM images is useful in determining site and extent of excision required. BACKGROUND: The value of SISCOM in predicting surgical outcome for extratemporal epilepsy is unknown, especially if MRI findings are nonlocalizing. METHODS: SISCOM images in 36 consecutive patients were classified by blinded reviewers as "localizing and concordant with site of surgery," "localizing but nonconcordant with site of surgery," or "nonlocalizing." SISCOM images were coregistered with postoperative MRI, and reviewers visually determined whether cerebral cortex underlying the SISCOM focus had been completely resected, partially resected, or not resected. RESULTS: Twenty-four patients (66.7%) had localizing SISCOM, including 13 (76.5%) of those without a focal MRI lesion. Eleven of 19 patients (57.9%) with localizing SISCOM concordant with the surgical site, compared with 3 of 17 (17.6%) with nonlocalizing or nonconcordant SISCOM, had an excellent outcome (p < 0.05). With logistic regression analysis, SISCOM findings were predictive of postsurgical outcome, independently of MRI or scalp ictal EEG findings (p < 0.05). The extent of resection of the cortical region of the SISCOM focus was significantly associated with the rate of excellent outcome (100% with complete resection, 60% with partial resection, and 20% with nonresection, p < 0.05). CONCLUSION: SISCOM images may be useful in guiding the location and extent of resection in extratemporal epilepsy surgery. PMID- 11113222 TI - MRI, (1)H-MRS, and functional MRI during and after prolonged nonconvulsive seizure activity. AB - BACKGROUND: Various structural and functional changes, such as focal edema, blood flow, and metabolism, occur in the cerebral cortex after focal status epilepticus. These changes can be assessed noninvasively by means of MRI techniques, such as fluid-attenuated inversion recovery (FLAIR), EEG-triggered functional MRI (EEG-fMRI), and proton MR spectroscopy (MRS). METHODS: The authors report on a 40-year-old patient with nonlesional partial epilepsy in the left posterior quadrant in whom these MRI techniques were applied in an active seizure focus and repeated during a follow-up of 1 year. RESULTS: FLAIR imaging taken at the time of status epilepticus showed a signal hyperintensity in the occipital region. (1)H-MRS of this cortical region showed elevated lactate, decreased N: acetylaspartate (NAA), and elevated choline (Cho). In the same region, EEG-fMRI revealed an area of signal enhancement. After seizure control, recovery of lactate and Cho was observed, whereas the NAA level remained reduced. The structural abnormality demonstrated on FLAIR disappeared within 3 months. CONCLUSIONS: Repetitive MRI with sensitive sequences during clinically critical periods may disclose the structural correlate in a previously nonlesional epilepsy case. Corresponding to the clinical evolution, reversible and irreversible focally abnormal metabolism can be determined with (1)H-MRS, reflecting both increased neuronal activity and neuronal damage. PMID- 11113223 TI - Lissencephaly: fetal pattern of glucose metabolism on positron emission tomography? AB - BACKGROUND: In classical lissencephaly, the cerebral cortex is four-layered, containing neurons that have failed to complete their migration between 12 and 16 weeks of gestation. METHODS: The authors studied the functional activity of lissencephalic cortex using 2-deoxy-2[(18)F]fluoro-D-glucose PET (FDG PET) in eight patients (six girls and two boys, mean age 7.5 years) with isolated lissencephaly sequence. RESULTS: The PET scans revealed a remarkably similar and bilaterally symmetric pattern of glucose metabolism in all eight patients. The cerebral cortex of lissencephaly showed two layers that could be differentiated based on metabolic activity. The inner layer, which probably corresponds to the inner cellular layer of lissencephalic cortex, showed 8 to 63% higher glucose utilization rate than the outer layer, which probably represents a composite of the molecular, outer cellular, and cell-sparse layers. Patients with a higher metabolic ratio between the cortical layers (inner/outer) showed greater delay in communication (p = 0.007) and socialization (p = 0.03). CONCLUSIONS: These findings are consistent with [(14)C]-2-deoxyglucose autoradiography studies in fetal sheep that have shown that before the development of significant numbers of axons, dendrites, and synapses, glucose metabolism appears to be highest in regions with the highest density of cell bodies, compared to the more mature state when glucose metabolism is highest in areas of greatest dendritic arborization. FDG PET studies of classical lissencephaly provide a different perspective in the analysis of brain gyral anomalies than those with traditional neuroanatomic imaging techniques. PMID- 11113224 TI - A novel ryanodine receptor gene mutation causing both cores and rods in congenital myopathy. AB - BACKGROUND: Central core disease (CCD) and nemaline rod myopathy are generally considered two genetically and histologically distinct disorders. CCD is defined by the presence of well-demarcated round cores within most myofibers. Nemaline rod myopathy is distinguished by the presence of characteristic nemaline bodies within myofibers. The simultaneous occurrence of both cores and rods in the same muscle biopsy has been described, but no gene mutations have been reported yet for this condition. OBJECTIVE: To describe a family containing 16 affected individuals in six generations with an autosomal dominant congenital myopathy that shows clinical and histologic features of both CCD and nemaline myopathy, and to determine the genetic etiology and protein composition of the cores/rods in this family. METHODS AND RESULTS: The results of linkage analyses excluded involvement of the two autosomal dominant nemaline myopathy loci on chromosome 1, but were consistent with a localization of the disease gene at the CCD locus on chromosome 19q13.1 (ryanodine receptor). SSCP analysis and DNA sequencing identified a novel Thr4637Ala mutation in the transmembrane region of the ryanodine receptor protein. Immunofluorescence studies of patient muscle biopsies showed the central cores to stain for ryanodine receptor. CONCLUSIONS: These data suggest that the occurrence of nemaline bodies can be a secondary feature of CCD, and that genetic studies on previously reported core/rod families should be targeted to the ryanodine receptor locus. The results of the immunofluorescence studies suggest that the cores contain excess abnormal ryanodine receptor protein. PMID- 11113225 TI - A "dystrophic" variant of autosomal recessive myotonia congenita caused by novel mutations in the CLCN1 gene. AB - OBJECTIVES: To identify the disease-causing mutation and its molecular consequence for a clinically distinct type of myotonic myopathy. BACKGROUND: The authors encountered a unique myotonic disorder of early onset in a 37-year-old man and his 47-year-old sister. METHODS: After examining known loci of inherited myotonic disorders, the authors looked for mutations within the CLCN1 gene using single strand conformation polymorphism and direct sequencing. To investigate the disease mechanism, reverse transcriptase PCR analyses of total RNA were performed. RESULTS: In the proband and his affected sister, two novel mutations comprising a compound heterozygous state in the CLCN1 gene were identified: 1) a base (G) insertion in exon 7 generating a premature termination codon (fs289X) in the D5 domain, and 2) a C-to-T substitution in exon 23 resulting in a missense mutation (P932L). These mutations accompanied a clinical phenotype that is distinguishable from recessive myotonia congenita by progressive generalized muscle weakness, severe distal muscle atrophy, joint contractures, high serum creatine kinase levels, and conspicuous myopathic changes on muscle histopathology. Reverse transcriptase PCR analyses detected only the P932L mutant mRNA in skeletal muscle, suggesting that the fs289X mRNA is degraded rapidly. CONCLUSIONS: These data suggest that fs289X is a null mutation, rendering the patients with the compound heterozygous genotype of fs289X/P932L to exclusively express P932L homomeric channels that may have caused the "dystrophic" phenotype. PMID- 11113226 TI - Mechanisms of immunomodulation by glatiramer acetate. AB - OBJECTIVE: To define the mechanism of action of glatiramer acetate (GA; formerly known as copolymer-1) as an immunomodulatory treatment for MS. BACKGROUND: The proposed mechanisms of action of GA include 1) functional inhibition of myelin reactive T cells by human leukocyte antigen (HLA) blocking, 2) T-cell receptor (TCR) antagonism, and 3) induction of T helper 2 (Th2) immunomodulatory cells. In this report, the authors examined the effects of GA on the functional activation of human T-cell clones (TCC) specific for myelin basic protein (MBP) and for foreign antigens. Several questions were addressed: Is the inhibitory effect of GA specific for autoantigens? Is it mediated by blocking the interaction between peptide and HLA molecule? Is GA a partial agonist or TCR antagonist, or does it induce anergy? Does it induce Th2 modulatory T cells? METHODS: The effects of GA on antigen-induced activation of human TCC specific for MBP, influenza virus hemagglutinin, and Borrelia burgdorferi were studied by proliferation and cytokine measurements, TCR downmodulation, and anergy assays. GA-specific TCC were generated in vitro from the peripheral blood of patients and healthy controls by limiting dilution. RESULTS: GA more strongly inhibited the proliferation of MBP, as compared with foreign antigen-specific TCC; in some MBP specific TCC, the production of Th1-type cytokines was preferentially inhibited. In addition to HLA competition, the induction of anergy, but not direct TCR antagonism, was observed. Numerous GA-specific TCC were generated from the peripheral blood of both MS patients and normal controls, and a fraction of these showed a Th2 phenotype. CONCLUSIONS: This study confirms a preferential inhibitory effect of GA on autoreactive TCC. With respect to cellular mechanisms, although HLA competition appears to play the most important role in functional inhibition in vitro, a direct effect on the TCR may be involved at least in some autoreactive T cells as shown by anergy induction. Although not confirmed at the clonal level, it is demonstrated further that GA induces T cells that crossreact with myelin proteins. GA-specific, Th2-modulatory cells may play an important role in mediating the effect of the drug in vivo. PMID- 11113227 TI - Whole brain volume changes in patients with progressive MS treated with cladribine. AB - OBJECTIVE: To compare changes in whole brain volume measured using MRI scans in patients with progressive MS enrolled in a double-blind, placebo-controlled trial assessing the efficacy of two doses of cladribine (0.7 and 2.1 mg/kg) and to assess the correlations between change in whole brain volume and change in other conventional MRI measures. BACKGROUND: Measuring brain parenchymal volumes is an objective and reliable surrogate for the destructive pathologic process in MS. The dynamics and the mechanisms of tissue loss in progressive MS are unclear. METHODS: Whole brain volumes were measured using postcontrast T1-weighted scans with 3 mm slice thickness from 159 patients with progressive MS (70% secondary progressive and 30% primary progressive) enrolled in a double-blind, placebo controlled trial of 12-month duration. RESULTS: Whole brain volumes were similar in the placebo and cladribine-treated patients on the baseline scans. A significant decrease of brain volume over time was observed both in the entire population of patients (p = 0.001) and in the placebo patients in isolation (p = 0.04). No significant treatment effect of either dose of cladribine on brain volume changes over time was found. In the 54 patients who received placebo, the change in brain volume was not significantly correlated with other MRI measures at baseline (enhancing lesion number and volume and T2-hyperintense and T1 hypointense lesion volumes) or at follow-up (cumulative number of enhancing lesions and absolute and percentage changes of enhancing T2- and T1-hypointense lesion volumes). CONCLUSIONS: This study shows in a large cohort of patients that brain parenchymal loss occurs, even over a short period of time, in progressive MS and that cladribine is not able to alter this process significantly. It also suggests that MRI-visible inflammation and new lesion formation has a marginal role in the development of brain atrophy in patients with progressive MS. PMID- 11113228 TI - Oxidative stress in patients with Friedreich ataxia. AB - Increased generation of reactive oxygen species may underlie the pathophysiology of Friedreich ataxia (FRDA). The authors measured concentrations of 8-hydroxy-2' deoxyguanosine (8OH2'dG), a marker of oxidative DNA damage, in urine and of dihydroxybenzoic acid (DHBA), a marker of hydroxyl radical attack, in plasma of 33 patients with FRDA. They found a 2.6-fold increase in normalized urinary 8OH2'dG but no change in plasma DHBA as compared with controls. Oral treatment with 5 mg/kg/day of the antioxidant idebenone for 8 weeks significantly decreased urinary 8OH2'dG concentrations, indicating that 8OH2'dG may be useful in monitoring therapeutic interventions in patients with FRDA.-1721 PMID- 11113229 TI - Absent headache despite CSF volume depletion (intracranial hypotension). AB - CSF volume depletions, whether from leak or shunt overdrainage, typically cause low CSF opening pressures, orthostatic headaches, and diffuse pachymeningeal gadolinium enhancement on MRI. The authors report three patients-two with overdraining CSF shunts and one with proven CSF leak-with the typical pachymeningeal enhancement but without headaches. In CSF leaks and CSF shunt overdrainage, like MRI abnormalities and CSF alterations, the clinical features also show considerable variability. The independent variable remains the CSF volume depletion. PMID- 11113230 TI - The value of informant versus individual's complaints of memory impairment in early dementia. AB - Self-reported versus informant-reported memory problems in nondemented elderly adults and in individuals with very mild and mild dementia of the Alzheimer type (DAT) were correlated with cognitive outcomes. No significant correlations were found between self-reported memory complaints and cognitive performance or (in controls) later development of dementia. In contrast, informant-reported memory loss distinguished nondemented from demented individuals and predicted future diagnosis of DAT. PMID- 11113231 TI - Expression of tau immunoreactivity in the spinal motor neurons of Alzheimer's disease. AB - Tau-related pathology was investigated in the spinal cord of 11 patients with AD. Ten ALS and 10 age-matched neurologically intact patients served as controls. Tau immunoreactivity was detected in neurons of the anterior horn in all AD cases and to a lesser extent in the intermediate zone and in the posterior horn. Tau immunoreactivity was rare in controls. Neurofibrillary tangles were identified in seven AD cases, but none was observed in the controls. PMID- 11113232 TI - The pathology of REM sleep behavior disorder with comorbid Lewy body dementia. AB - A patient with REM sleep behavior disorder who subsequently developed probable Lewy body dementia is now reported to have a definite pathologic diagnosis of Lewy body dementia. Examination of brain revealed Lewy bodies as well as marked neuronal loss in brainstem monoaminergic nuclei-particularly locus coeruleus and substantia nigra-that inhibit cholinergic neurons in the pedunculopontine nucleus mediating atonia during REM sleep. PMID- 11113233 TI - Improvement of sleep architecture in PD with subthalamic nucleus stimulation. AB - High-frequency stimulation of the subthalamic nucleus (STN) was used to investigate the relationship of sleep disorders with motor handicap in PD. In 10 insomniac patients with PD, stimulation reduced nighttime akinesia by 60% and completely suppressed axial and early morning dystonia, but did not alleviate periodic leg movements (n = 3) or REM sleep behavior disorders (n = 5). Total sleep time increased by 47%; wakefulness after sleep onset decreased by 51 minutes. Insomnia in patients with PD may predominantly result from nighttime motor disability. PMID- 11113234 TI - Dopa-responsive dystonia: mutation analysis of GCH1 and analysis of therapeutic doses of L-dopa. German Dystonia Study Group. AB - Analysis of the gene GCH1 in 58 patients with dystonia and a positive response to L-dopa revealed mutations in 30 individuals from 22 families. Thirteen of the mutations observed were familial, three occurred de novo, and inheritance could not be determined in six cases. There was no mutation in the promoter region of GCH1 in any patient. The doses of L-dopa given to members of the two groups were not significantly different. PMID- 11113235 TI - Cerebral microembolism in acute spontaneous internal carotid artery dissection. AB - The rate and risk factors for early ischemic recurrence in patients with internal carotid artery dissection (ICAD) are largely unknown. Serial transcranial Doppler (TCD) monitoring of microembolic signals (MES) was performed in 28 consecutive patients with acute ICAD. MES were identified in 13 patients, and early ischemic recurrence occurred in 7. Six of 13 patients with MES and 1 of 15 without MES experienced early ischemic recurrence (p = 0.029). MES detection on serial TCD monitoring may be associated with an increased risk of early ischemic recurrence in patients with acute ICAD. PMID- 11113236 TI - Arteriovenous bubbles following cold water sport dives: relation to right-to-left shunting. AB - Neurologic injury subsequent to decompression from diving may be due to paradoxical arterialization of venous gas emboli. Of 40 divers who performed 53 open water dives after being tested for a patent foramen ovale (PFO), arterial gas emboli were detected in 7 of 13 dives, which resulted in venous bubbles. In five of these seven dives, there was evidence of a PFO by contrast transcranial Doppler sonography, indicating an increased risk of arterializing venous bubbles in divers with a PFO. PMID- 11113237 TI - Neurologic outcome of controlled compressed-air diving. AB - The authors compared the neurologic, neuropsychological, and neuroradiologic status of military compressed-air divers without a history of neurologic decompression illness and controls. No gross differences in the neuropsychometric test results or abnormal neurologic findings were found. There was no correlation between test results, diving experience, and number and size of cerebral MRI lesions. Prevalence of cerebral lesions was not increased in divers. These results suggest that there are no long-term CNS sequelae in military divers if diving is performed under controlled conditions. PMID- 11113238 TI - Patients with intractable epilepsy have low melatonin, which increases following seizures. AB - Melatonin, which is used to treat sleep disorders, has anticonvulsant properties. The authors measured salivary melatonin and cortisol, at baseline and following seizures, in patients with intractable temporal lobe epilepsy and controls. Melatonin was reduced in patients with epilepsy at baseline compared with controls, and increased threefold following seizures. Cortisol also increased following seizures. Patients with intractable epilepsy have low baseline melatonin levels that increase dramatically following seizures. PMID- 11113239 TI - A placebo-controlled crossover trial of creatine in mitochondrial diseases. AB - To test the efficacy and safety of creatine (Cr) monohydrate in mitochondrial diseases, 16 patients with chronic progressive external ophthalmoplegia or mitochondrial myopathy were randomized in a crossover design to receive double blind placebo or 20 g Cr/day for 4 weeks. Cr was well tolerated, but there were no significant effects with regard to exercise performance, eye movements, or activities of daily life. The power of this pilot study was limited and future multicenter trials are needed. PMID- 11113240 TI - MRI correlate of Kernohan's notch. PMID- 11113241 TI - Increased levels of plasma malondialdehyde in Friedreich ataxia. PMID- 11113242 TI - Treatment of drug-induced psychosis with quetiapine and clozapine in Parkinson's disease. PMID- 11113243 TI - Bilateral pallidal stimulation for cervical dystonia: dissociated pain and motor improvement. PMID- 11113244 TI - Amyloidosis presenting with intractable epistaxis and multiple cranial neuropathies. PMID- 11113245 TI - Cerebral manifestations of cefepime toxicity in a dialysis patient. PMID- 11113246 TI - Exacerbation of juvenile myoclonic epilepsy with lamotrigine. PMID- 11113247 TI - Patients' versus caregivers' report of poor memory in relation to dementia and tested abilities. PMID- 11113248 TI - Poststroke depression and emotional incontinence. PMID- 11113249 TI - Comparison of diffusion-weighted MRI and CT in acute stroke. PMID- 11113250 TI - Developmental apraxia arising from neonatal brachial plexus palsy. PMID- 11113251 TI - Physiologic-pathologic correlation in Guillain-Barre syndrome in children. PMID- 11113253 TI - Giardiasis as a re-emerging infectious disease and its zoonotic potential. AB - The reasons for considering giardiasis as a re-emerging infectious disease are presented, with emphasis on Giardia infections in child care centres, livestock and pets, and the role of zoonotic transmission. However, the aetiology and control of giardiasis is complicated by the genetic and phenotypic variability of Giardia species infective to mammals. Of particular significance has been the uncertainty about host specificity and the question of zoonotic transmission. The recent application of molecular characterisation procedures based on PCR has made an enormous contribution to an understanding of the genetic structure of Giardia populations, and this is reviewed in the context of the zoonotic transmission and molecular epidemiology of Giardia infections. PMID- 11113254 TI - The leishmaniases as emerging and reemerging zoonoses. AB - The 20 or so species of Leishmania which have been recorded as human infections are all either zoonotic, or have recent zoonotic origins. Their distribution is determined by that of their vector, their reservoir host, or both, so is dependent on precise environmental features. This concatenation of limiting factors leads to specific environmental requirements and focal distribution of zoonotic or anthroponotic sources. Human infection is dependent on the ecological relationship between human activity and reservoir systems. Examples are available of the emergence of leishmaniasis from the distant past to the present, and can be postulated for the future. These emergences have been provoked by the adoption of new, secondary reservoir hosts, the adoption of new vector species, transport of infection in humans or domestic animals, invasion by humans of zoonotic foci, and irruption of reservoir hosts beyond their normal range. The leishmaniases therefore present an excellent model for emerging disease in general, and for the generation of the principles governing emergence. The model is, however, limited by gaps in our knowledge, usually quantitative, sometimes qualitative, of the structure of reservoir systems. PMID- 11113255 TI - Echinococcosis: an emerging or re-emerging zoonosis? AB - The aim of this review is a critical discussion of factors actually or potentially contributing to persistence or emergence of echinococcosis in humans. Alveolar echinococcosis (AE), a life-threatening infection of humans, is caused by a larval stage of Echinococcus multilocularis. The adult parasite inhabits the intestine of foxes and other carnivores and has a wide distribution in the northern hemisphere (North America and northern and central Eurasia). Recent surveys in central Europe have extended the known geographical occurrence of E. multilocularis in foxes from four countries at the end of the 1980s to at least 11 countries in 1999. Cases of human AE previously regularly reported from only four countries are now recorded from seven countries, but the annual incidences are low. Since adequate information from earlier surveys is not available, it is not possible to conclude if the new findings reflect a recent extension of the parasite's range or just the first identification of hitherto unnoticed endemic areas. Evidence of parasite spreading has been reported from North America and Japan. Factors with the potential of enhancing the infection risk for humans in the future include increasing fox populations and parasite prevalences, progressing invasion of cities by foxes, the establishment of urban cycles of the parasite, and the spill-over of the E. multilocularis infection from wild carnivores to domestic dogs and cats. In view of the potential severity and fatality of AE in humans health authorities should initiate internationally coordinated countermeasures. Although control programmes against human cystic echinococcosis (CE), caused by E. granulosus, have been established in some countries and effective control strategies are available, the parasite has still a wide geographical distribution affecting many countries of all continents. Thus, human CE is persisting in many parts of the world with high incidences, and in some areas it is a re-emerging problem. For example, alarming increases of the number of human cases have been reported from Bulgaria and Kazakhstan, and the People's Republic of China. Progress in control can only be expected if health authorities attribute a higher priority to this disease and if all modern diagnostic and control options (for example vaccination of intermediate host animals) can be used. PMID- 11113256 TI - Neuro-angiostrongyliasis: unresolved issues. AB - Angiostrongylus cantonensis, the rat lungworm, probably evolved with its hosts, members of the genus Rattus and closely related species, in south-east Asia. Since its first discovery in rats in China and in a case of human infection in Taiwan, the parasite has been found to infect humans and other mammals across a wide and ever-increasing territory, which now encompasses much of south-east Asia, Melanesia, Polynesia and eastern Australia. It has also established a foothold in Africa, India, the Caribbean and south-eastern USA. This dispersal has been a direct result of human activity, and in some cases has been linked with the spread of the African giant land snail, Achatina fulica. However, this snail is not critical to the extension of the parasite's range, as numerous other indigenous molluscan species serve as adequate intermediate hosts; the importance of Achatina to the life cycle may have been over-emphasized. In Australia, the parasite is established along parts of the east coast, and the presence of an indigenous close relative, Angiostrongylus mackerrasae, suggests a long association of the parasite with its local rat hosts, a situation analogous to that of Angiostrongylus malaysiensis in south-east Asia. These three Angiostrongylus species share virtually the same life cycle, but only A. cantonensis has been confirmed to be a human pathogen. PMID- 11113257 TI - Epidemiology of Cryptosporidium: transmission, detection and identification. AB - There are 10 valid species of Cryptosporidium and perhaps other cryptic species hidden under the umbrella of Cryptosporidium parvum. The oocyst stage is of primary importance for the dispersal, survival, and infectivity of the parasite and is of major importance for detection and identification. Because most oocysts measure 4-6 microm, appear nearly spherical, and have obscure internal structures, there are few or no morphometric features to differentiate species and in vitro cultivation does not provide differential data as for bacteria. Consequently, we rely on a combination of data from three tools: morphometrics, molecular techniques, and host specificity. Of 152 species of mammals reported to be infected with C. parvum or an indistinguishable organism, very few oocysts have ever been examined using more than one of these tools. This paper reviews the valid species of Cryptosporidium, their hosts and morphometrics; the reported hosts for the human pathogen, C. parvum; the mechanisms of transmission; the drinking water, recreational water, and food-borne outbreaks resulting from infection with C. parvum; and the microscopic, immunological, and molecular methods used to detect and identify species and genotypes. PMID- 11113258 TI - Human babesiosis: an emerging tick-borne disease. AB - Human babesiosis is an important emerging tick-borne disease. Babesia divergens, a parasite of cattle, has been implicated as the most common agent of human babesiosis in Europe, causing severe disease in splenectomized individuals. In the US, Babesia microti, a babesial parasite of small mammals, has been the cause of over 300 cases of human babesiosis since 1969, resulting in mild to severe disease, even in non-splenectomised patients. Changing ecology has contributed greatly to the increase and expansion of human babesiosis in the US. A relatively recently described babesial parasite, the WA1-type, has been shown to be the causative agent in seven human cases in the western US. This parasite is closely related to babesial parasites isolated from large wild ungulates in California. Like B. microti, WA1-type parasites cause mild to severe disease and the immunopathogenesis of these parasites is distinctly different from each other in experimental infections of hamsters and mice. A B. divergens-like parasite was also identified as the cause of a fatal human babesiosis case in Missouri. Isolated cases of human babesisosis have been described in Africa and Mexico, but the causative parasites were not well characterized. Standard diagnostic techniques for human infection, such as examination of Giemsa-stained thin blood smears and serology, have been complemented with molecular techniques, such as PCR. Current treatment for babesiosis is focused on a regimen of clindamycin and quinine, although new drugs have shown promise. Prevention of infection relies on self-monitoring for the presence of ticks and, in some locations, targeted application of pesticides to decrease tick abundance. Identification of human infection with Babesia spp. will probably increase as physicians and the public become more aware of the disease, as people live and recreate in rural tick infested areas, and as the numbers of immunocompromised individuals increase. PMID- 11113252 TI - Toxoplasma gondii: from animals to humans. AB - Toxoplasmosis is one of the more common parasitic zoonoses world-wide. Its causative agent, Toxoplasma gondii, is a facultatively heteroxenous, polyxenous protozoon that has developed several potential routes of transmission within and between different host species. If first contracted during pregnancy, T. gondii may be transmitted vertically by tachyzoites that are passed to the foetus via the placenta. Horizontal transmission of T. gondii may involve three life-cycle stages, i.e. ingesting infectious oocysts from the environment or ingesting tissue cysts or tachyzoites which are contained in meat or primary offal (viscera) of many different animals. Transmission may also occur via tachyzoites contained in blood products, tissue transplants, or unpasteurised milk. However, it is not known which of these routes is more important epidemiologically. In the past, the consumption of raw or undercooked meat, in particular of pigs and sheep, has been regarded as a major route of transmission to humans. However, recent studies showed that the prevalence of T. gondii in meat-producing animals decreased considerably over the past 20 years in areas with intensive farm management. For example, in several countries of the European Union prevalences of T. gondii in fattening pigs are now <1%. Considering these data it is unlikely that pork is still a major source of infection for humans in these countries. However, it is likely that the major routes of transmission are different in human populations with differences in culture and eating habits. In the Americas, recent outbreaks of acute toxoplasmosis in humans have been associated with oocyst contamination of the environment. Therefore, future epidemiological studies on T. gondii infections should consider the role of oocysts as potential sources of infection for humans, and methods to monitor these are currently being developed. This review presents recent epidemiological data on T. gondii, hypotheses on the major routes of transmission to humans in different populations, and preventive measures that may reduce the risk of contracting a primary infection during pregnancy. PMID- 11113259 TI - Trichinellosis: the zoonosis that won't go quietly. AB - Trichinellosis, is normally not included among those regarded as emerging zoonoses because it has been a public health threat for more than 150 years. However, its dramatic re-emergence in many areas around the world over the past 10-20 years, inspite of a century of veterinary public health efforts to control and eradicate it, justifies it being included in this group. The reasons for this re-emergence are diverse, and include human pertubation and manipulation of ecosystems, war and political turmoil, rapidly changing food distribution and marketing systems, and even, surprisingly, rising affluence in developing countries. These influences, and their impact on the epidemiology of both domestic and sylvatic trichinellosis, are discussed, along with recommendations for confronting this altered status as a public health threat. PMID- 11113260 TI - Emerging helminth zoonoses. AB - As our ability to recognise and diagnose human disease caused by helminth parasites has improved, so our understanding of the epidemiology and clinical manifestations of these diseases has improved. Humans can develop patent infection with a wide range of helminth parasites, whose natural host is another vertebrate. Rather than focusing on a comprehensive review of zoonotic helminth infections, this review describes in detail examples of zoonotic helminth infections that have newly appeared in human populations, or have existed but are increasing in incidence or geographic range. Examples include intestinal capillariasis, anisakidosis, eosinophilic enteritis, oesophagostomiasis and gnathostomiasis. Potential reasons for the emergence of these infections, including changes in social, dietary or cultural mores, environmental changes, and the improved recognition of heretofore neglected infections often coupled with an improved ability to diagnose infection are discussed. PMID- 11113261 TI - Emerging parasite zoonoses: the role of host-parasite relationship. AB - Human-parasite relationships have played an essential role in the emergence or re emergence of some parasitic diseases. These interactions are due to numerous causes. Some are linked to humans (immunodeficiencies due to AIDS among other causes, treatments, nosocomial contaminations, genetic predisposition), others concern the parasite (particular genotypes having modified their parasitic specificity). Several of these causes were predominant in the emergence of parasitoses such as cryptosporidiasis, microsporidioses or, to a certain point, pneumocystosis, the transmission of which has become zoonotic or even anthroponotic, inter-human. Re-emergent diseases (toxoplasmosis, leishmaniasis, giardiasis, strongyloidiasis, scabies) had already been described in human pathology, but their frequency or symptomatology have been drastically modified. In this case also, the unbalanced host-parasite relationship is largely responsible but it can not be dissociated from other causes, especially environmental and nutritional. PMID- 11113262 TI - The role of companion animals in the emergence of parasitic zoonoses. AB - Pets offer individuals and the community significant benefits, however cognisance must be taken of the potential for transmission of infectious agents from these animals to humans. The prevalence of many parasites, such as Giardia and Cryptosporidium, has increased over the past few decades while others, such as Toxocara and Ancylostoma, have decreased. These changes could be real, associated with the ready availability of efficacious anthelmintic products or could be artificial due to the type of surveys conducted, the animals surveyed and the diagnostic tests used. Immunocompromised people, in particular, must be aware of the potential risk of acquiring parasitic infections from their pets. However, with the adoption of good hygiene and a thorough knowledge of the transmission of these parasites, immunocompromised people should be able to continue to enjoy the significant benefits of pet ownership. As many owners are not aware of the zoonotic parasites that could be carried by their pets or their mode of transmission, it is concluded that veterinarians need to play a greater role in the education of their clients. PMID- 11113263 TI - Emerging parasite zoonoses associated with water and food. AB - The environmental route of transmission is important for many protozoan and helminth parasites, with water, soil and food being particularly significant. Both the potential for producing large numbers of transmissive stages and their environmental robustness, being able to survive in moist microclimates for prolonged periods of time, pose a persistent threat to public and veterinary health. The increased demands on natural resources increase the likelihood of encountering environments and produce contaminated with parasites. For waterborne diseases, the protozoa, Cryptosporidium, Giardia and Toxoplasma, are the most significant causes, yet, with the exception of Toxoplasma, the contribution of zoonotic transmission remains unclear due to the absence of 'standardised' methods. The microsporidia have been documented in one waterborne outbreak, but the role of animals as the cause of contamination was not elucidated. In foods, surface contamination is associated with the faecal-oral pathogens, and some data are available to indicate that animal wastes remain an important source of contamination (e.g. cattle faeces and apple cider outbreaks), however, further work should focus on examining the source of contamination on fruit and vegetables. Increasing recognition of the burden of human fascioliasis has occurred; it is now recognised as an emerging zoonosis by the WHO. Toxoplasma, Trichinella and Taenia spp. remain important meatborne parasites, however, others, including Pleistophora-like microsporidians may be acquired from raw or lightly cooked fish or crustaceans. With increased international travel, the public health importance of the foodborne trematodiases must also be realised. Global sourcing of food, coupled with changing consumer vogues, including the consumption of raw vegetables and undercooking to retain the natural taste and preserve heat-labile nutrients, can increase the risk of foodborne transmission. A greater awareness of parasite contamination of our environment and its impact on health has precipitated the development of better detection methods. Robust, efficient detection, viability and typing methods are required to assess risks and to further epidemiological understanding. PMID- 11113264 TI - Effects of environmental change on emerging parasitic diseases. AB - Ecological disturbances exert an influence on the emergence and proliferation of malaria and zoonotic parasitic diseases, including, Leishmaniasis, cryptosporidiosis, giardiasis, trypanosomiasis, schistosomiasis, filariasis, onchocerciasis, and loiasis. Each environmental change, whether occurring as a natural phenomenon or through human intervention, changes the ecological balance and context within which disease hosts or vectors and parasites breed, develop, and transmit disease. Each species occupies a particular ecological niche and vector species sub-populations are distinct behaviourally and genetically as they adapt to man-made environments. Most zoonotic parasites display three distinct life cycles: sylvatic, zoonotic, and anthroponotic. In adapting to changed environmental conditions, including reduced non-human population and increased human population, some vectors display conversion from a primarily zoophyllic to primarily anthrophyllic orientation. Deforestation and ensuing changes in landuse, human settlement, commercial development, road construction, water control systems (dams, canals, irrigation systems, reservoirs), and climate, singly, and in combination have been accompanied by global increases in morbidity and mortality from emergent parasitic disease. The replacement of forests with crop farming, ranching, and raising small animals can create supportive habitats for parasites and their host vectors. When the land use of deforested areas changes, the pattern of human settlement is altered and habitat fragmentation may provide opportunities for exchange and transmission of parasites to the heretofore uninfected humans. Construction of water control projects can lead to shifts in such vector populations as snails and mosquitoes and their parasites. Construction of roads in previously inaccessible forested areas can lead to erosion, and stagnant ponds by blocking the flow of streams when the water rises during the rainy season. The combined effects of environmentally detrimental changes in local land use and alterations in global climate disrupt the natural ecosystem and can increase the risk of transmission of parasitic diseases to the human population. PMID- 11113265 TI - Detection and characterisation of parasites causing emerging zoonoses. AB - Understanding the epidemiology of zoonotic parasitic infections is dependent upon the availability of accurate and sensitive diagnostic techniques. The development of molecular diagnostic methods, particularly those utilising PCR for the detection of zoonoses will contribute greatly to the identification and control of these pathogens, by increasing the speed of diagnosis, specificity and sensitivity, reproducibility and ease of interpretation. Molecular characterisation studies allow us to distinguish between closely related infectious agents and to document the patterns of transmission of 'strains' and species within populations. This will allow precise determinations to be made about the aetiological agent, its characteristics and the source of infection. This review focuses on recent detection and characterisation techniques for both emerging and re-emerging parasite zoonoses. PMID- 11113266 TI - Micro-evolution and emergence of pathogens. AB - Changes in the epidemiology of infectious diseases are the direct result of ecological and evolutionary changes in hosts and parasites. Precisely what the causal processes are is rarely known in any particular case, and this hinders the design of appropriate control strategies. This is particularly so for emerging infections, as opportunity is rapidly lost to study the ecological parameters which might have affected initial emergence. However, molecular evolutionary studies of the pathogens can yield data which discriminate between possible causes. The current distribution of DNA sequence variation is important information which may reveal past and current changes in pathogen population structures, and can also identify adaptive changes in pathogen genes which have affected their evolution. Such studies have been quite intensively performed on particular viral and bacterial pathogens, and some of the successes of these are noted here. Approaches to understanding the recent evolution of eukaryotic pathogens are outlined, with particular reference to current problems of emerging zoonoses, and changes in virulence and drug resistance. PMID- 11113267 TI - Emerging parasite zoonoses. PMID- 11113268 TI - Enhanced shear-induced von Willebrand factor binding to platelets in acute myocardial infarction. AB - Recent investigations have suggested that von Willebrand factor (vWF) plays a crucial role in platelet thrombosis under flow conditions. The effects of plasma obtained from 15 patients with acute myocardial infarction and 10 patients with the chest pain syndrome as controls, on shear-induced vWF binding to platelets and subsequent platelet activation, as evidenced by microparticle release, were investigated by quantitative flow cytometry. Platelet-rich plasma was obtained from a 38-year-old healthy male volunteer with blood type O. Stored plasma from either the acute myocardial infarction or control patients was then added to the freshly prepared platelet-rich plasma in equal volumes. The mixtures were then exposed to specific shear rates in an optically modified cone-plate viscometer. The number of vWF molecules bound to the platelet surface and the number of microparticles released from the platelets were then measured by quantitative flow cytometry using an FITC-conjugated anti-vWF monoclonal antibody. The shear induced increase in vWF binding to the platelet surface was enhanced in the presence of plasma from patients with acute myocardial infarction (acute myocardial infarction plasma). The shear-induced release of microparticles from platelets was enhanced from 889+/-134 in the presence of control plasma to 1045+/ 222 in the presence of the acute myocardial infarction plasma (p<0.05). Acute myocardial infarction plasma also reduced the shear rate threshold required to induce measurable shear-induced vWF binding from 10800 s(-1) to 9000 s(-1). We conclude that the plasma of acute myocardial infarction patients contains factors that enhance shear-induced vWF binding and vWF-mediated platelet activation, which may contribute to thrombotic re-occlusions of the coronary arteries in patients who have received reperfusion treatments. PMID- 11113269 TI - Gene polymorphisms in the TNF locus and the risk of myocardial infarction. AB - We investigated two genetic polymorphisms in the tumor necrosis factor locus (TNF alpha -308 G-->A and LT-alpha +252 A-->G) as risk factors for coronary atherothrombotic disease (CAD) by determining its prevalence in 148 survivors of myocardial infarction (MI) with angiographically-proven severe CAD, and in 148 age-, gender- and race-matched controls. The odds ratio (OR) for MI related to the mutant TNF-alpha and LT-alpha alleles was 0.8 (CI95: 0.4-1.3) and 1. 3 (CI95: 0.8-2.0), respectively. We also sought interaction of smoking and metabolic risk factors for MI with each mutant genotype. Smokers not carrying the LT-alpha +252 A-->G mutation had a risk of MI of 2.7 (CI95: 1.4-5.4) whereas in smoking carriers the risk was 6. 9 (CI95: 3.4-14.1). An interactive effect of the LT alpha mutation may also exist with dyslipidemia (OR for MI in non-carriers was 12 [CI95: 3.2-41.3] and in carriers the OR was 39, [CI95: 5.1-301] and with obesity (OR for MI was 2.7, [CI95: 1-7.2] in non-carriers and in carriers the OR was 6 [CI95: 2.1-16.8]). Lastly, the OR for MI in obese non-carriers of TNF-alpha -308 G-->A was 2.8 (CI95: 1.3-6) and in obese carriers the OR was 14.5 (CI95: 1.8 113). Although significant interactive effects could not be detected, the findings suggest that interaction of polymorphisms in the TNF locus with major risk factors for CAD may exist, and should be explored in larger studies. PMID- 11113270 TI - Mild hyperhomocysteinemia and fibrinolytic factors in patients with history of venous thromboembolism. AB - Mild hyperhomocysteinemia is recognized as a risk factor for venous thromboembolism (VTE), though its role in the thrombogenic processes is not understood. Its possible association with impaired fibrinolysis was investigated in 157 patients (61 women, 96 men) below the age of 60 years (43+/-11, mean+/-SD) with a history of objectively confirmed VTE. Patients had significantly higher fasting total plasma homocysteine (tHcy) levels than 138 apparently healthy subjects (8.0, 6.6-9.9 micromol/L vs. 7.2, 5.9-8.6 micromol/L, P=0. 001; median, range between first and third quartile). In 17 of 157 patients (12%) hyperhomocysteinemia (tHcy>11.4 micromol/L for women and tHcy>12.6 micromol/L for men) was established. The adjusted odds ratio as an estimate of relative risk for VTE was 2.3 (0.8-7.0; 95% confidence interval). When patients with hyperhomocysteinemia were compared to patients without hyperhomocysteinemia, no significant differences in t-PA (antigen 9.2+/-5.5 microg/L and 9.7+/-4.7 microg/L, respectively; activity 1.3+/-0.5 IU/mL and 1.3+/-0.7 IU/mL, respectively) and PAI-1 (antigen 19.3+/-17.5 microg/L and 22.6+/-20. 4 microg/L, respectively; activity 15.0+/-12.6 and 15.8+/-13.3 IU/mL, respectively) were observed. In conclusion, this study showed an association between mild hyperhomocysteinemia and VTE, but provided no evidence for an independent association between hyperhomocysteinemia and alterations in fibrinolytic proteins. PMID- 11113271 TI - Accuracy of a portable prothrombin time monitor (Coagucheck) in patients on chronic oral anticoagulant therapy: a prospective multicenter study. AB - A portable prothrombin time (PT) monitor allows patients on oral anticoagulant therapy (OAT) to measure their PT at home. The purpose of the study was to evaluate the accuracy and precision of a portable PT monitor (Coagucheck, Roche Diagnostics, Mannheim, Germany) as compared with laboratory methods. The prospective study was conducted in four centers of the Italian Federation of Anticoagulation Clinics. A one-month instruction phase was followed by a six month surveillance phase. Seventy-eight subjects on stable OAT (48 men, 30 women, age range: 18-75) were selected on a volunteer basis. Dual measurements of INR values were performed in each subject both from finger capillary blood by the monitor and from venous blood by the Anticoagulation Clinic laboratory in three instruction sessions. The mean difference (bias) of the monitor INR results when compared with the average of laboratory INR and monitor INR results was -0.025 (limits of agreement-LA: -0.84/+0.81 INR units). The mean bias was -0.0675 (LA: 0.37/+0.23), +0.018 (LA: -0.39/+0.35), and +0.039 (LA: -0.49/+0.55), respectively, for INR values lower than 2.0, between 2.0 and 3.0, and greater than 3.0. The overall precision coefficient of monitor INR was 0.370, while it was 0.23, 0.46, 0.29, and 0.21, respectively, in Centers 1, 2, 3, and 4. The overall variation coefficient was 6.5% while it was 3.7%, 8.5%, 4.7%, and 4.9%, respectively, in Centers 1, 2, 3, and 4. Coagucheck has an acceptable level of accuracy for INR values in the range between 2.0 and 3.0. A wide variation in monitor performance was found among centers. PMID- 11113272 TI - Diagnostic value of the Nycocard, Nycomed D-dimer assay for the diagnosis of deep venous thrombosis and pulmonary embolism: a retrospective study. AB - The relatively new D-dimer assay from Nycomed Pharma (Nycocard), which has shown both high sensitivity and specificity in diagnosing deep venous thrombosis (DVT) and pulmonary embolism (PE) in earlier studies, was re-evaluated retrospectively. The diagnostic value of the D-dimer assay for DVT was evaluated with contrast venography as reference. The diagnostic value of the D-dimer assay for PE was evaluated with pulmonary scintigraphy as reference. D-dimer tests were examined from 54 consecutive patients. The D-dimer assay was found to have a sensitivity and specificity of 50% and 58%, respectively, for the diagnosis of DVT, with a positive predictive value (PPV) and negative predictive value (NPV) of 55% and 54%, respectively. Using reference diagnostic results of high probability and low probability for the diagnosis of PE, the D-dimer test sensitivity and specificity was found to be 40% and 94%, respectively (PPV: 86%, NPV: 64%). The diagnostic value of the Nycocard(R) assay appears to be very limited for the diagnosis of DVT and PE. This retrospective study suggests that it is unsuitable as a screening method. Further re-evaluation of D-dimer assays is recommended prior to routine clinical use. PMID- 11113273 TI - Adrenaline potentiates type 2B von Willebrand factor-induced activation of human platelets by enhancing both the formation and action of thromboxanes. AB - Von Willebrand factor (vWF) is a large plasma glycoprotein that mediates platelet adhesion at sites of vascular injury. We have previously reported that the pathological type 2B (formerly named type IIB) variant of vWF promotes platelet activation through phospholipase A(2)-mediated release of arachidonic acid. The present report shows that adrenaline (1 microM) potentiates type 2B vWF-induced platelet aggregation, serotonin secretion, rise in cytosolic Ca(2+) concentration, and pleckstrin phosphorylation, as well as thromboxane B(2) production. The hormone also increases the partially inhibited release of serotonin observed in platelets pretreated with the anti-GPIIb-IIIa antibody LJCP8 but does remove the total inhibition on the secretion caused by the anti GPIb antibody LJIB1. Adrenaline also increases type 2B vWF-elicited tyrosine phosphorylation of proteins with apparent molecular masses of 60 and 80 kDa. Furthermore, adrenaline potentiates the rise in cytosolic Ca(2+) and the release of thromboxane B(2) in platelets stimulated with arachidonic acid (2 microM) as well as the increase in Ca(2+) induced by the thromboxane mimetic U46619 (0.3 microM). Platelet pretreatment with yohimbine or 13-azaprostanoic acid, which are antagonists of the alpha(2)-adrenergic and thromboxane receptors, respectively, or with acetylsalicylate and indomethacin, both of which act as inhibitors of thromboxane formation, abolishes the potentiating effect of adrenaline. These observations lead to the conclusion that the potentiating action of adrenaline on type 2B vWF-promoted platelet responses is due to an increase in both the formation and activating action of thromboxanes. PMID- 11113275 TI - Time related neutralization of two doses acetyl salicylic acid. AB - Aspirin has a well established role in the prevention of arterial thrombosis. Discussion on the efficacy and safety of aspirin in the treatment and prophylaxis of thrombosis has become an important issue. In fact, hemorrhage complications are often associated with its use. On the other hand, previous studies showed unexpected thrombotic potencies associated with the presence of this drug at ultra low doses (ULD) in the circulation. In this study, we aimed to evaluate the effect of aspirin at ULD, injected 1, 2, or 3 hours after the administration of aspirin at 100 mg/kg, on hemostasis and bleeding in rats. We used an experimental model of thrombosis induced by laser beams to evaluate these effects. Platelet aggregation was determined by Cardinal and Flower method. Results from this investigation demonstrate that the neutralizing effect of aspirin at ULD did not operate significantly 1 hour after the injection of aspirin at 100 mg/kg. This effect was observed 2 and 3 hours after. The use of aspirin at ULD to neutralize the side effects of aspirin at high doses will reduce the hemorrhagic risk during extra corporeal circulation. The therapeutic benefit and safety of aspirin therapy in the treatment of cardiovascular diseases can be obtained. PMID- 11113274 TI - Antithrombotic potential of olive oil administration in rabbits with elevated cholesterol. AB - Olive oil is the main source of dietary fatty acids in the Mediterranean region. The objective of this study was to evaluate the effect of dietary supplementation with virgin olive oil in an experimental model with rabbits fed an atherogenic diet (saturated fat 48% of total fat). Four different groups of 10 animals each were studied: (1) normolipemic diet (NLD), (2) atherogenic diet or saturated fatty acid-enriched diet (SFAED), (3) NLD with 15% olive oil (NLD+OLIV), and (4) SFAED with 15% virgin olive oil (SFAED+OLIV). The animals were fed the experimental diets for 6 weeks, after which we determined serum lipid profile (total cholesterol, HDL-cholesterol, and triglycerides), platelet aggregation, platelet thromboxane B(2), aortic prostacyclin, and platelet and vascular lipid peroxidation. Scanning electron microscopic images of the vascular endothelium were studied, as were morphometric parameters in the arterial wall and thrombogenicity of the subendothelium (annular perfusion chamber). Animals fed the SFAED showed platelet hyperactivity and increased subendothelial thrombogenicity. Animals fed the SFAED+OLIV showed, compared with the SFAED group, an improved lipid profile with decreased platelet hyperactivity and subendothelial thrombogenicity and less severe morphological lesions of the endothelium and vascular wall. We conclude that supplementation of the SFAED with 15% olive oil reduced vascular thrombogenicity and platelet activation in rabbits. Although the percentage of olive oil in the diet was higher than the amount in the human diet, these results may be helpful in determining the effect of olive oil in the human thrombogenic system. PMID- 11113276 TI - Synergistic effect of endothelin-1 and serotonin in rabbit platelets: effect on tyrosine phosphorylation. PMID- 11113277 TI - Acceleration of fibrinolysis by high-frequency ultrasound: the contribution of acoustic streaming and temperature rise. AB - High-frequency ultrasound has been shown to accelerate enzymatic fibrinolysis. One of the supposed mechanisms of this effect is the enhancement of mass transport by acoustic streaming, i.e., ultrasound-induced macroscopic flow around the clot. In this study, which is aimed at further elucidating the mechanisms of the acceleration of fibrinolysis by ultrasound, we investigated whether ultrasound would accelerate fibrinolysis if the flow around the thrombus is already present, as may occur in vivo. The effect of the ultrasound-induced temperature rise was also studied. In a model of a plasma clot submerged in plasma, containing tissue-type plasminogen activator, mild stirring of the outer plasma producing a shear rate of 40 seconds(-1) at the surface of the clot resulted in a two-fold acceleration of lysis. A similar effect was obtained with ultrasound (1 MHz, 2 W/cm(2)). Furthermore, if ultrasound was applied together with stirring, only 30% acceleration by ultrasound was documented, fully attributable to the concomitant temperature rise. In a model with tissue-type plasminogen activator incorporated throughout a plasma clot, the effect of ultrasound (two-fold shortening of lysis time) was fully attributable to the concomitant temperature rise of a few degrees. We concluded that the acceleration of enzymatic plasma clot lysis by high-frequency ultrasound in the models we used can be largely explained by a combination of the effects of heating and acoustic streaming, equivalent to mild stirring. The thermal effects can hardly be utilized in vivo due to the danger of tissue overheat. The therapeutic advantage of transcutaneous high-frequency ultrasound as an adjunct to thrombolytic therapy may appear limited to the situations where there is no flow in the direct environment of the thrombus. PMID- 11113278 TI - Differences between neonates and adults in the urokinase-plasminogen activator (u PA) pathway of the fibrinolytic system. AB - This study deals with plasminogen activation kinetics of fetal and adult Glu plasminogen types 1 and 2 as well as fetal and adult Lys-plasminogen by urokinase in the presence and absence of the lysine analogues epsilon-amino-n-caproic acid (EACA) and tranexamic acid. In addition, activation kinetics of single-chain urokinase-plasminogen activator (scu-PA) by adult and fetal plasmin types were investigated in the absence and presence of soluble fibrin. All Lys-plasminogen isoforms were more readily activated by urokinase than their corresponding Glu plasminogen types. No substantial differences of the catalytic constants of urokinase-catalyzed plasminogen activation could be obtained when all fetal plasminogen types were compared to the respective adult types. In the case of all Glu-plasminogen isoforms, EACA as well as tranexamic acid first stimulated the activation process and, at higher concentrations, showed inhibitory properties. Again, the relative ability of all fetal plasminogen types to interact with lysine analogues revealed no differences compared to the respective adult glycoforms. In the absence of soluble fibrin, the catalytic efficiency of scu-PA activation by plasmin was significantly lower for both fetal plasmin isoforms. However, there were no differences in catalytic efficiency between fetal and adult plasmin types in the presence of 4 microM soluble fibrin. In conclusion, no substantial differences exist in urokinase-catalyzed plasminogen activation between neonates and adults, which is in contrast to reported data on plasminogen activation by tissue-type plasminogen activator. In the absence of soluble fibrin, scu-PA activation by fetal plasmin is markedly slower than by adult plasmin. However, this is compensated when fibrin is added at a concentration that is close to the physiological fibrinogen concentration in plasma. It can be summarized that the differences in carbohydrate structures of fetal and adult plasminogen are not associated with major differences in the global function of this part of fibrinolysis, despite functional alterations of scu-PA activation. PMID- 11113279 TI - Inhibition of plasmin activity by sulfated polyvinylalcohol-acrylate copolymers. AB - The effect of four sulfated polyvinylalcohol-acrylate copolymers and heparin on plasminogen activation and on plasmin activity is studied. The molecules differing in charge (proportion of negatively charged units 40.5%-73.5% of the total) and in size (5600 Da-8800 Da) accelerate plasminogen activation by 2- up to 4-fold at a 7-fold molar excess of the polyvinylacrylates over plasminogen. They, however, exert a concentration and charge-dependent effect on plasmin: both the amidolytic (half-maximal effect at a 1.33-3.66 molar excess of the polyvinylacrylates) and fibrinolytic (half-maximal effect at 1.23-1.72 molar excess of the polyvinylacrylates) activities of plasmin are inhibited. In contrast, heparin (a similarly carboxylated and sulfated polymer) and polyvinylacrylates with a low number of sulfate groups (30% sulfated monomers) at concentrations up to 2.2 microM do not affect plasminogen activation and plasmin activity in a milieu of physiological ionic strength. Experiments with plasmin derivatives lacking N-terminal peptides of different length (des-kringle(1-4) and des-kringle(1-5) plasmin) show identical changes in the protease activities, precluding involvement of the kringle-domain in the interaction with the polyvinylacrylates. Fluorescence studies evidence the charge-dependent binding of the polyvinylacrylates to plasmin, but not to plasminogen. Thus, through non covalent interaction with the protease-domain of plasmin the polyvinylacrylates inhibit fibrinolysis. Since these sulfated copolymers inhibit both thrombin [4] and plasmin activity, they may be a useful therapeutic tool in situations when both the blood coagulation and the fibrinolytic system are activated (such as intravascular coagulation and fibrinolysis, ICF). PMID- 11113280 TI - Isolation and characterization of a human alternative complement pathway inhibiting protein from larval hemolymph of the silkworm, Bombyx mori. AB - An alternative complement pathway-inhibiting protein (ACPIP), which inhibits the activation of the alternative complement pathway (ACP) of the human serum, was isolated from larval hemolymph of the silkworm, Bombyx mori, by using ammonium sulfate fractionation and column chromatographies to homogeneity. About 400microg of ACPIP was routinely obtained from 20ml hemolymph. The purified ACPIP preparation consisted of two distinct polypeptides (34 and 32kDa) on SDS-PAGE. The amino acid compositions of the two polypeptides were nearly identical; 21% of the amino acid residues were acidic. The amino terminal amino acid sequences up to 20 residues in these two polypeptides were also identical. Addition of the ACPIP to human serum resulted in a dose-dependent inhibition of the hemolysis of intact rabbit erythrocytes via the ACP, whereas in no inhibition of hemolysis of sensitized-sheep erythrocytes (EA) via the classical pathway. PMID- 11113281 TI - Products of proteolytic cleavage of transferrin induce nitric oxide response of goldfish macrophages. AB - Enzymatic cleavage product of transferrin induced the production of nitric oxide (NO) by LPS-stimulated goldfish macrophages. A NO-inducing factor was purified from the supernatants of mitogen-stimulated goldfish kidney leukocytes using fast performance liquid chromatography (FPLC) and the purified proteins analyzed by microcapillary reverse-phase HPLC nano-electrospray tandem mass spectrometry. The proteins were identified as truncated forms of transferrin, having approximate molecular weights (MW) of 33, 35, and 37kDa (kilodaltons). The precursor form (i.e. full-length) of transferrin did not enhance NO production by LPS-stimulated goldfish macrophages, but enzymatic cleavage of this precursor form correlated with enhanced production of NO by goldfish macrophages. Enzymatic cleavage of transferrin was dependent on the presence of stimulated kidney leukocytes and was shown to occur in response to both mixed lymphocyte reactions (MLR) and the mitogenic stimulation of goldfish kidney leukocytes. Time course analysis revealed that 24h after kidney leukocyte MLR or mitogen stimulation, cleaved transferrin products appeared in the supernatants of cultured cells, which was related to the on-set of NO-inducing activity of these preparations. To confirm these findings, bovine transferrin was digested in vitro using protease XXVII. The resulting cleavage products had approximate MW of 33, 35, and 37kDa. When these peptides were subjected to the purification protocols used to purify a NO inducing factor from goldfish leukocyte supernatants, they were shown to elute to identical fractions. To examine the potential role of fish transferrin in mediating goldfish NO production, carp transferrin was purified from serum and following protease-digestion and purification by FPLC, the truncated proteins were found to elute to similar fractions as bovine transferrin. Furthermore, mitogen-stimulated leukocyte supernatants prepared in the absence of bovine serum (carp serum only) retained NO-inducing activity, indicating that this response was not an artifact of bovine serum components (i.e. bovine transferrin). Anti bovine and anti-carp transferrin polyclonal antibodies identified the presence of truncated forms of transferrin in the active fractions of FPLC-separated mitogen stimulated leukocyte supernatants prepared in the presence of bovine or carp serum, respectively. Thus, our results suggest a novel role for fish transferrin as one of the factors that mediates teleost macrophage antimicrobial functions. PMID- 11113282 TI - T-cell antigen receptors in Atlantic cod (Gadus morhua l.): structure, organisation and expression of TCR alpha and beta genes. AB - By using short degenerate primers complementing conserved T-cell antigen receptor (TCR) variable and constant region segments for PCR, we were able to isolate putative TCRalpha and beta chain full length cDNAs in Atlantic cod. The Valpha and Vbeta domains have the canonical features of known teleost and mammalian TCR V domains, including conserved residues in the beginning of FR2 and at the end of FR3. The Jalpha and Jbeta region possess the conserved Phe-Gly-X-Gly motif found in nearly all TCR and immunoglobulin light chain J regions. Similar to other vertebrates, the Atlantic cod Calpha and Cbeta sequences exhibit distinct immunoglobulin, connecting peptide, transmembrane and cytoplasmic regions. The Atlantic cod Cbeta sequence lacks a cysteine in its connecting peptide region, but other motifs proposed to be important for dimerisation and cell surface expression are observed. Four different cod Cbeta sequences were identified, two of which share 3' untranslated regions different from one of the other two sequences, suggesting the existence of isotypic gene variants of Cbeta. Based on Southern blot analyses, the TCRalpha and beta gene loci appear to be arranged in translocon organisation (as opposed to multicluster) with multiple V gene segments, some (D) and J gene segments and a single or few C gene segments. Northern blot analyses show expression of the TCRalpha and beta chains in thymus, spleen and head kidney, expression of the TCRbeta chain was also detected in the ovary. Interestingly, no expression was detected in intestine even though the existence of T-cells in intestine has been proposed in other teleost species. PMID- 11113283 TI - Cloning and developmental expression of a family of pleurocidin-like antimicrobial peptides from winter flounder, Pleuronectes americanus (Walbaum). AB - Low molecular weight antimicrobial peptides are an important component of the innate immune system in animals, yet they have not been examined widely in fish. Of particular interest is their expression during development and in response to environmental conditions and disease. Here, we report the isolation of four genomic sequences encoding putative antimicrobial peptides from the winter flounder, Pleuronectes americanus (Walbaum), as well as reverse transcription-PCR products from two tissues that form the first defensive barrier to microbes - skin and intestine. Alignment of the predicted polypeptide sequences shows a conserved hydrophobic signal peptide of 22 amino acids followed by 25 amino acids that are identical (WF2) or homologous to the amino acid sequence of pleurocidin, followed by a conserved acidic portion. Southern hybridisation analysis indicates that related peptides are encoded in the genomes of other flatfish species. Northern and RT-PCR analyses of RNA from multiple tissues show that two of the pleurocidin genes are expressed predominantly in the skin whereas two other genes are expressed mainly in the intestine. RT-PCR assays of total RNA from larvae of different ages provide the first evidence of developmental expression of antimicrobial peptides in fish and indicate that the pleurocidin gene is first expressed at 13 days post-hatch in winter flounder. PMID- 11113284 TI - Major histocompatibility complex and immunoglobulin loci visualized by in situ hybridization on Xenopus chromosomes. AB - A technique for fluorescent in situ hybridization (FISH) on chromosomes of the amphibian Xenopus laevis is described. Positive results were obtained with cDNA probes of about 1kb when at least three adjacent copies of the gene are present. The immunoglobulin heavy chain locus is in the centre of the long arm of chromosome 1. Previously, family studies showed that bona fide MHC class Ib genes segregated independently. Now we show that MHC class II alpha and beta genes and class Ib genes are on the same acrocentric chromosome, with MHC in the middle of the long arm, the class Ib complex (XNC) at the tip or the same arm. Each locus or complex is found on only one pair of chromosomes confirming the diploidization of these genes in the pseudotetraploid X. laevis. PMID- 11113285 TI - Induction of the respiratory burst in turtle peritoneal macrophages by Salmonella muenchen. AB - Peritoneal macrophages were collected from juvenile turtles 72h after intraperitoneal inoculation with a 3% Sephadex suspension. The macrophages were assayed for their chemiluminescent (CL) properties, reflecting their respiratory burst activity, after stimulation with Zymosan A, phorbol 12-myristate 13-acetate (PMA), N-formyl-methionyl-leucyl-phenylalanine (fMLP), and calcium ionophore A23187. Except for fMLP, all triggering agents induced a marked CL response. Luminol was used as the chemiluminescent probe. When comparing CL responses in temperatures ranging from 15 to 35 degrees C, lower assay temperatures induced lower and slower CL responses. Stimulation with viable Salmonella muenchen resulted in a distinct response. Bacteria, inactivated by means of heat or acetone, induced a faster and stronger oxidative burst. Opsonization of either viable or heat-inactivated S. muenchen with non-inactivated anti-S. muenchen serum, prepared in turtles, induced faster and higher CL responses. On the other hand, opsonization of acetone-inactivated S. muenchen caused CL responses to be slower and weaker. S. muenchen, opsonized with heat-inactivated turtle anti S. muenchen serum, induced higher responses than non-opsonized bacteria, but slower and weaker responses than bacteria opsonized with native turtle antiserum. No response was recorded after stimulation with LPS and the supernatant of heat inactivated bacteria. PMID- 11113286 TI - Intestinal anaphylaxis in chickens: epithelial ion secretion as a determinant and potential component of functional immunity. AB - Immunity to secondary protozoan infections in chickens is accompanied by rapid onset of intestinal permeability to serum proteins, an event in mammals associated with local anaphylaxis. The permeability changes in the chicken intestine are hypothesized to be mediated by mast cell-derived paracrine factors. In a test of this hypothesis, we demonstrated, using an electrophysiological correlate of intestinal anaphylaxis (antigen-induced Cl(-) secretion), that the response of the chicken intestine to antigenic stimulation is consistent with type I hypersensitivity reactions. Day-old, single-comb-white-Leghorn chickens were sensitized to bovine serum albumin (BSA). At 3 weeks of age ileal segments were mounted in Ussing-type chambers. Serosal challenge with BSA elevated the transmural short circuit current (DeltaIsc) within 1min and was maximally expressed (DeltaIsc=50-60microA/cm(2)) within 2-3 min. The magnitude of the DeltaIsc was directly related to the concentration of antigen (10-200 microg antigen/ml), was only expressed in immunized chickens, and was blocked by the mucosal application of a Cl(-) channel blocker. Data obtained in the present investigation identify epithelial ion secretion as a potential mechanism of functional immunity in the mucosal immune system of the chicken small intestine. PMID- 11113287 TI - Pain following spinal cord injury: animal models and mechanistic studies. PMID- 11113288 TI - Abnormal brain chemistry in chronic back pain: an in vivo proton magnetic resonance spectroscopy study. AB - The neurobiology of chronic pain, including chronic back pain, is unknown. Structural imaging studies of the spine cannot explain all cases of chronic back pain. Functional brain imaging studies indicate that the brain activation patterns are different between chronic pain patients and normal subjects, and the thalamus, and prefrontal and cingulate cortices are involved in some types of chronic pain. Animal models of chronic pain suggest abnormal spinal cord chemistry. Does chronic pain cause brain chemistry changes? We examined brain chemistry changes in patients with chronic back pain using in vivo single- voxel proton magnetic resonance spectroscopy ((1)H-MRS). In vivo (1)H-MRS was used to measure relative concentrations of N-acetyl aspartate, creatine, choline, glutamate, glutamine, gamma-aminobutyric acid, inositol, glucose and lactate in relation to the concentration of creatine. These measurements were performed in six brain regions of nine chronic low back pain patients and 11 normal volunteers. All chronic back pain subjects underwent clinical evaluation and perceptual measures of pain and anxiety. We show that chronic back pain alters the human brain chemistry. Reductions of N-acetyl aspartate and glucose were demonstrated in the dorsolateral prefrontal cortex. Cingulate, sensorimotor, and other brain regions showed no chemical concentration differences. In chronic back pain, the interrelationship between chemicals within and across brain regions was abnormal, and there was a specific relationship between regional chemicals and perceptual measures of pain and anxiety. These findings provide direct evidence of abnormal brain chemistry in chronic back pain, which may be useful in diagnosis and future development of more effective pharmacological treatments. PMID- 11113289 TI - Comparison of autotomy behavior induced in rats by various clinically-used neurectomy methods. AB - When a peripheral nerve is cut, a neuroma develops at its proximal end. Nerve-end neuromas are known to be a source of ectopic sensory input. In some humans this input may cause spontaneous and evoked neuropathic pain. There is currently no available animal model for developing better methods of cutting nerves that produce less painful neuromas than those currently in clinical use. Transection of the sciatic and saphenous nerves in rats also produces nerve-end neuromas. Afferent fibers in such neuromas spontaneously emit ectopic input that coincides with the outbreak of licking, scratching and self-mutilation of the denervated limb ('autotomy'). This behavior is considered to be the expression of spontaneous disagreeable sensations such as paresthesias, dysesthesias or neuropathic pain. We propose here that the autotomy model can be used as the first step for development of better neurectomy methods. As a demonstration, in this report we compared the course of autotomy expressed by rats following several methods of cutting peripheral nerves that are currently in clinical use. We found that the lowest extent of autotomy was caused by sciatic and saphenous neurectomy with a CO(2) laser. Tight ligation of the nerve, or a simple cut with scissors, also yielded significantly lower autotomy scores compared to cryoneurolysis and electrocut. The differing scores of autotomy caused by these neurectomy methods may derive from different properties of the injury discharge produced by these methods at the time of nerve cut. Our results raise the possibility that a higher incidence of neuropathic pain or related sensory disorders in humans may be expected following cryosurgical and electrocut neurectomies. If validated by further studies, neurectomy methods eliciting lower incidence of autotomy, and sensory disorders in models not based on autotomy may produce lower levels of neuropathic pain in humans. PMID- 11113290 TI - A Pain Monitoring Program for nurses: effect on the administration of analgesics. AB - Both physicians and nurses are responsible for adequate pain management. The aim of this study was to assess pain management behavior of physicians and nurses, and to evaluate the effects of a Pain Monitoring Program for nurses on the extent to which nurses administer analgesics. The Pain Monitoring Program consisted of two components: educating nurses about pain, pain assessment and pain management; and implementing daily pain assessment by means of a numeric rating scale. Several outcomes were distinguished to evaluate the administration of analgesics by nurses: the prescribed analgesics by physicians, the administered analgesics by nurses, and the discrepancy between the ordered and the administered analgesics. The effects of the Pain Monitoring Program on these outcomes were measured in a quasi-experimental design with a non-equivalent control group. In total, 703 patients participated: 358 patients in the control group and 345 in the intervention group. Patients were interviewed twice, i.e. at the beginning and at the end of hospitalization. Results of the control group showed that at the first interview 70% of the patients were prescribed analgesics by physicians and only 74% of those patients were actually administered analgesics by nurses. Consequently, 50% of the patients in pain received analgesics. The administered analgesics was in absolute agreement with the prescribed analgesics in 60% of the patients with routine analgesics and in 85% of the patients with PRN analgesics. The relative difference between ordered and administered routine analgesics was small, namely 15% for opioids and 20% for non-opioids. Similar results of the control group were found for the second interview. In addition, the results showed that the Pain Monitoring Program was effective in improving nurses' administration of analgesics. At the first interview more patients received analgesics that were prescribed on a PRN basis and the doses of administered routine non-opioids including PRN increased. At the time of the second interview, more patients received weak opioids. The Pain Monitoring Program was especially effective in patients with moderate to severe pain. However, the discrepancy between the analgesics ordered by physicians and actually administered by nurses did not change as a result of the Pain Monitoring Program. Based on this study it can be concluded that the use of a simple method such as a numeric rating scale together with pain education for nurses is effective in improving the administration of analgesics by nurses. These are important results because nurses play an essential role in helping patients to cope with their pain. Because the Pain Monitoring Program (PMP) was effective in a heterogeneous population in multiple care settings, the possibility of implementing the PMP in routine nursing practice should be considered. PMID- 11113291 TI - The interaction between IL-1beta and morphine: possible mechanism of the deficiency of morphine-induced analgesia in diabetic mice. AB - It is known that diabetic mice are less sensitive to the analgesic effect of morphine. Some factor(s) derived from mononuclear cells, e.g. interleukin-1beta (IL-1beta), may be responsible for the diminished analgesic effect of morphine in diabetic mice. Therefore, we examined direct effects of IL-1beta, intracerebroventricularly (i.c.v.), on morphine-induced analgesia, subcutaneously (s.c.), in diabetic and control mice by using the tail-flick test. Morphine at doses of 1, 2 and 5 mg/kg (s.c.) produced dose-dependent analgesia in diabetic and control mice but diabetic mice were less sensitive to the analgesic effect of morphine when compared to the controls. IL-1beta at a dose of 0.1 ng/mouse produced analgesia in control mice but not in diabetics, whereas IL-1beta at a dose of 10 ng/mouse produced a hyperalgesic effect both in diabetic and control mice. IL-1beta at a dose of 1 ng/mouse has neither an analgesic nor a hyperalgesic effect in control and diabetic mice. Administration of a neutral (neither analgesic nor hyperalgesic) dose of IL-1beta, 1 ng/mouse (i.c.v.), just prior to administration of morphine (s.c.) abolished the analgesic effect of morphine at doses of 1, 2 and 5 mg/kg in control mice and the analgesic effect of morphine became similar to that in diabetics. The diminished analgesic effect of morphine in diabetes was attenuated further with IL-1beta at a dose of 1 ng/mouse (i.c.v.). These results suggest that the decreased analgesic effect of morphine in diabetes may be related to IL-1beta. PMID- 11113292 TI - Tetrodotoxin-resistant action potentials in dorsal root ganglion neurons are blocked by local anesthetics. AB - Evidence from animal models and studies of human sensory nerves demonstrate that tetrodotoxin (TTX)-resistant Na(+) channels are present in sensory neurons and might play an important role in pain conduction and chronic pain. Recent investigations suggest that TTX-resistant Na(+) channels in the peripheral nervous system are less sensitive to local anesthetics than TTX-sensitive Na(+) channels. To test the effects of the clinically used local anesthetics lidocaine and bupivacaine on TTX-resistant action potentials (APs) in sensory neurons, we performed electrophysiological experiments on small dorsal root ganglion (DRG) neurons from young rats. Amplitudes, time to peak and duration of TTX-resistant APs were measured in Adelta- and C-type neurons using the patch-clamp technique in a thin slice preparation (150 microm), thus avoiding enzymatic treatment. With increasing concentrations of the local anesthetics, the AP amplitude was gradually reduced but the AP did not disappear abruptly. The concentrations needed to reduce the amplitudes of TTX-resistant APs by half were 760 microM for lidocaine and 110 microM for bupivacaine. Time to peak and duration of TTX resistant APs were prolonged by local anesthetics. Trains of APs could be elicited in some neurons by long-lasting current injections, and the half-maximal concentrations needed to suppress these trains were 30 microM lidocaine or 10 microM bupivacaine. We suggest that the reduction in firing frequency at low concentrations of local anesthetic may explain the phenomenon of paresthesia when sensory information is gradually suppressed during spinal anesthesia. PMID- 11113293 TI - The formalin test in the mouse: a parametric analysis of scoring properties. AB - We investigated the scoring properties of the mouse formalin test using the time sampling method recently developed for infant and adult rats. Formalin was injected under the plantar surface of one rear paw (10 microl, 1-8%), and pain behaviours (paw favouring, lifting and licking) and behavioural state were recorded. Correlational and regression analyses indicated that scores composed of combinations of all three pain behaviours, either summed or weighted, provided less variable indices of pain than licking alone. The maximum percent effect (MPE(50); i.e. pain behaviour 50% of the time) for the log formalin concentration effect curves was 3-4% in both phases. Habituation to the test environment prior to testing did not alter the MPE(50)s, but slopes were lower in unhabituated mice, dramatically increasing the size of the confidence interval. Formalin dose dependently reduced locomotion, rearing and sniffing in both the first phase and the early part of the second phase. The combination measures were sensitive to morphine (2-8 mg/kg), amphetamine (1-4 mg/kg), dipyrone (50-200 mg/kg), xylazine (0.25-1 mg/kg), and acepromazine (0.25-1 mg/kg), and resistant to diazepam (0.5-2 mg/kg), pimozide (0.05-0.25 mg/kg), pentobarbital (10 and 15 mg/kg) and indomethacin (2-8 mg/kg). Decreased pain was correlated with increased motor activity for morphine and amphetamine, and with decreased activity for xylazine and acepromazine; dipyrone and indomethacin did not alter activity levels. PMID- 11113294 TI - In vivo pharmacology of SDZ 249-665, a novel, non-pungent capsaicin analogue. AB - Capsaicin and analogues are valuable analgesic agents when administered to mammals, including humans. However, their pungency and the effects on the cardiovascular and respiratory systems through their general activation of small calibre (nociceptive) primary afferents severely limit their use. Recently, structure activity analysis revealed that the initial pungent and general excitatory effects can be prevented by structural modifications in such a way that the analgesic activity is retained. In this paper we present SDZ 249-665, a capsaicin analogue which produced analgesia in the mouse and anti-hyperalgesic effects in the rat and guinea pig. SDZ 249-665 was administered p.o., s.c. and i.v. in models of nociceptive pain, such as tail flick latency in response to a noxious thermal stimulus and acetic acid-induced writhing in mice, and in models of inflammatory mechanical hyperalgesia induced by turpentine or carrageenan in the rat and guinea pig, respectively. SDZ 249-665 was effective in the tail flick and the writhing assays and produced significant anti-hyperalgesic effects in the inflammatory models. The efficacy of SDZ 245-665 was similar to that of capsaicin, however, it was significantly more potent. SDZ 249-665 did not produce any irritancy in a nose wipe assay in guinea pigs or an eye irritancy assay in rats, while capsaicin was clearly irritant in both cases. Furthermore, unlike capsaicin, SDZ 249-665 did not produce unwanted side effects such as bronchoconstriction and blood pressure changes in the analgesic/anti-hyperalgesic dose range. Thus, a clear analgesic therapeutic window exists for SDZ 249-665. In summary, SDZ 249-665 is a potent orally active, analgesic/anti-hyperalgesic agent in mouse, rat and guinea pig. It lacks the excitatory effects associated with capsaicin and other close analogues, and therefore provides a clear therapeutic window for use in painful conditions. In addition to this favourable profile, no sign of tolerance was detected after a 5 day repeated dose treatment. PMID- 11113295 TI - Expectations of analgesia do not affect spinal nociceptive R-III reflex activity: an experimental study into the mechanism of placebo-induced analgesia. AB - The purpose of this study was to investigate whether placebo analgesia is mediated by the release of beta-endorphin. In addition to subjective pain reports, we included an objective physiological parameter of nociception reflected by the opioid sensitive nociceptive R-III reflex. Placebo consisted of strong suggestions of pain relief and an intravenous injection of saline. Forty minutes after placebo, either the opioid antagonist naloxone or saline was administered intravenously without subjects noticing (hidden). Sixty healthy males, aged 18-30 years, voluntarily participated in this study. Subjects were randomized into one of four groups: group 1 received placebo and hidden naloxone, group 2 received hidden naloxone only, group 3 received placebo and hidden saline and group 4 received hidden saline only. Pain was induced by electrical stimulation of the sural nerve and evaluated with a visual analogue scale (VAS). In addition, changes in the magnitude of the nociceptive R-III reflex activity were assessed. We determined to what extent R-III reflex activity and subjective pain reports were decreased by placebo and we investigated whether these placebo induced changes in reflex activity and subjective pain reports were naloxone reversible. Furthermore, we measured the degree of association between pain relief as measured on VAS and changes in R-III reflex activity. Finally, the role of beta-endorphin was assessed by measuring plasma endorphin levels before and after the administration of placebo. This study could not demonstrate a placebo effect as measured on VAS and R-III responses. The administration of placebo did not appear to have an effect on the release of beta-endorphins. Consistently, the antagonizing effects of naloxone were negligible. A subgroup analysis of those who did show a placebo response as indicated on the VAS did not support the supposition that beta-endorphin is released due to placebo suggestion. It is suggested that intensified stimuli and a more effective procedure to induce placebo analgesia (e.g. conditioning) may produce a proper placebo effect. PMID- 11113296 TI - An excitatory role for 5-HT in spinal inflammatory nociceptive transmission; state-dependent actions via dorsal horn 5-HT(3) receptors in the anaesthetized rat. AB - The involvement of 5-HT(3) receptor mediated modulation of formalin and carrageenan induced inflammatory transmission was investigated. The effects of the selective 5-HT(3) receptor antagonist ondansetron on the electrically evoked responses of dorsal horn neurones in normal animals were compared to those following carrageenan. The effect of pre-treatment on the formalin response was also studied. Ondansetron had no significant effect on the electrically evoked responses of dorsal horn neurones in normal animals or following carrageenan induced inflammation, but significantly inhibited both phases of the formalin response. Our results suggest that 5-HT(3) receptors in the spinal cord have no significant role under normal conditions. However, during formalin (but not carrageenan) induced inflammation this system is activated, maintaining the response of nociceptive spinal neurones to peripheral formalin. PMID- 11113297 TI - Age-related differences in the endogenous analgesic response to repeated cold water immersion in human volunteers. AB - Recent animal studies using stress-induced analgesia have suggested a general age related decline in endogenous pain inhibitory systems. The aim of the current study was to examine age-related differences in the magnitude of endogenous analgesia in human volunteers, using psychophysical measures of neuroselective electrical, and thermal CO(2) laser induced pain thresholds, before, immediately after and 1 h after repeated cold water immersion of the hand. Sensory detection thresholds did not differ between age groups indicating that the functional integrity of primary afferent sensory fibres appears to be intact in older people. Consistent with many previous studies, older adults required a higher intensity of noxious stimulation in order to first report the presence of pain. The cold water immersion task was effective in eliciting a powerful analgesic response, regardless of age; pain thresholds were shown to increase by up to 100% immediately after the cold pressor test. This effect was relatively transient with thresholds returning to baseline within 1 h. The magnitude of analgesic response, however, was found to be significantly less in older people. Age differences in the efficacy of endogenous analgesic systems may be expected to reduce the ability of older adults to cope with severe persistent pain states and may help explain some of the variation in the literature on pain report. PMID- 11113298 TI - Partial sciatic nerve transection as a model of neuropathic pain: a qualitative and quantitative neuropathological study. AB - One of the most commonly used experimental animal models for neuropathic pain is the chronic constriction injury (CCI) where four loose ligatures are tied around the sciatic nerve. One disadvantage of this model is the introduction of foreign material into the wound, which causes a local inflammatory reaction. Thus the distinction between the neuropathic and the inflammatory component of pain is difficult in this model. In order to produce a pure nerve lesion, we performed a partial sciatic nerve transection (PST; a modification of the Seltzer model) in female Sprague-Dawley rats and compared behavior and nerve pathology. These rats developed thermal hyperalgesia and mechanical allodynia comparable to the CCI model. Recovery of these symptoms was found between days 40 and 60 after the nerve lesion. Some animals still showed symptoms on day 101, which was associated with a neuroma formation. The main pathological findings in the endoneurium in nerve segments distal to the lesion were edema, loss of myelinated fibers and increase in endoneurial cells, especially macrophages. In the epineurium the number of macrophages was strikingly increased after CCI compared with PST, indicating that the response of the immune system is different in a structural lesion with and without foreign material. In conclusion, PST is a pure nerve injury model without an epineurial inflammatory component due to foreign material and is therefore well suited for studying the role of local endoneurial processes in the development and maintenance of neuropathic pain. Also, the importance of regeneration in the termination of hyperalgesia can convincingly be shown in this model. PMID- 11113299 TI - The nature of the effect of female gonadal hormone replacement therapy on cognitive function in post-menopausal women: a meta-analysis. AB - We reviewed epidemiological and experimental studies of female gonadal hormone replacement therapy (HRT) on cognitive function in post-menopausal women and carried out meta-analyses. In healthy ageing women, HRT has small and inconsistent effects that include enhancement of verbal memory, abstract reasoning and information processing. Epidemiological studies show larger effects than experimental studies, which is not related to sample size. Important confounds may be that women who start using HRT are healthier than women who do not. Also, controlling for socio-economic status diminishes the effect of HRT. The effects of HRT may depend on the age and type of menopause and the therapeutic intervention used, with the most widely used drug, Premarin, having least effect. However, the effects are independent of mood and climacteric symptom alleviation. There is a paucity of experimental studies that include healthy elderly women. The evidence for an estrogen deficiency in women with dementia and cognitive dysfunction is inconsistent. Nevertheless, epidemiological studies suggest that HRT protects against the development of clinically diagnosed Alzheimer's disease. However, poor recall of HRT use by patients and altered physician behaviour may have confounded the effects. Surprisingly, both healthy and demented women with low education seem to benefit most from HRT. Three recent controlled experimental studies using Premarin showed no effects of HRT in preventing further cognitive decline in women who already have Alzheimer's disease. Duration of treatment seems to play an important role, with beneficial effects declining-and even reversing-with longer treatment in women with Alzheimer's disease.Future research should further investigate the cognitive effect of different HRT preparations, serum estrogen levels, and the interactions of HRT with age, menopausal status and existing protective (e.g. education) and risk factors (e.g. smoking and apolipoprotein E genotype) for cognitive decline and Alzheimer's disease. PMID- 11113300 TI - Bilateral lesions of the central but not anterior or posterior parts of the piriform cortex retard amygdala kindling in rats. AB - The piriform cortex is thought to be involved in temporal lobe seizure propagation, such as that occurring during kindling of the amygdala or hippocampus. A number of observations suggested that the circuits of the piriform cortex might act as a critical pathway for limbic seizure discharges to assess motor systems, but direct evidence for this suggestion is scarce. Furthermore, the piriform cortex is not a homogeneous structure, which complicates studies on its role in limbic epileptogenesis. We have previously reported data indicating that the central part of the piriform cortex might be particularly involved during amygdala kindling. In order to further evaluate the role of different parts of the piriform cortex during kindling development, we bilaterally destroyed either the central, anterior or posterior piriform cortex by microinjections of ibotenate two weeks before onset of amygdala kindling. Lesions of the anterior piriform cortex hardly affected kindling acquisition, except that fewer animals exhibited stage 3 (unilateral forelimb) seizures compared to sham controls. Lesions of the central piriform cortex significantly retarded kindling, which was due to a decreased progression from stage 3 to stage 4/5 seizures, i.e. the lesioned rats needed significantly longer for the acquisition of generalized clonic seizures in the late stages of kindling development. Lesions of the posterior piriform cortex did not significantly affect kindling development. The data demonstrate that different parts of the piriform cortex mediate qualitatively different effects on amygdala kindling. The central piriform cortex seems to be a neural substrate involved in the continuous development of kindling from stage 3 to stages 4/5, indicating that this part of the piriform cortex may have preferred access, either directly or indirectly, to structures capable of supporting generalized kindled seizure expression. PMID- 11113301 TI - Dual electroencephalogram markers of human sleep homeostasis: correlation between theta activity in waking and slow-wave activity in sleep. AB - To investigate the relationship between markers of sleep homeostasis during waking and sleep, the electroencephalogram of eight young males was recorded intermittently during a 40-h waking episode, as well as during baseline and recovery sleep. In the course of extended waking, spectral power of the electroencephalogram in the 5-8Hz band (theta activity) increased. In non-rapid eye movement sleep, power in the 0.75-4.5Hz band (slow-wave activity) was enhanced in the recovery night relative to baseline. Comparison of individual records revealed a positive correlation between the rise rate of theta activity during waking and the increase in slow-wave activity in the first non-rapid eye movement sleep episode. A topographic analysis based on 27 derivations showed that both effects were largest in frontal areas. From these results, we suggest that theta activity in waking and slow-wave activity in sleep are markers of a common homeostatic sleep process. PMID- 11113302 TI - Increased synaptic inhibition in dentate gyrus of mice with reduced levels of endogenous brain-derived neurotrophic factor. AB - The aim of this study was to explore the role of endogenous neurotrophins for inhibitory synaptic transmission in the dentate gyrus of adult mice. Heterozygous knockout (+/-) mice or neurotrophin scavenging proteins were used to reduce the levels of endogenous brain-derived neurotrophic factor and neurotrophin-3. Patch clamp recordings from dentate granule cells in brain slices showed that the frequency, but not the kinetics or amplitude, of miniature inhibitory postsynaptic currents was modulated in brain-derived neurotrophic factor +/- compared to wild-type (+/+) mice. Furthermore, paired-pulse depression of evoked inhibitory synaptic responses was increased in brain-derived neurotrophic factor +/- mice. Similar results were obtained in brain slices from brain-derived neurotrophic factor +/+ mice incubated with tyrosine receptor kinase B immunoglobulin G, which scavenges endogenous brain-derived neurotrophic factor. The increased inhibitory synaptic activity in brain-derived neurotrophic factor +/- mice was accompanied by decreased excitability of the granule cells. No differences in the frequency, amplitude or kinetics of miniature inhibitory postsynaptic currents were seen between neurotrophin-3 +/- and +/+ mice. From these results we suggest that endogenous brain-derived neurotrophic factor, but not neurotrophin-3, has acute modulatory effects on synaptic inhibition onto dentate granule cells. The site of action seems to be located presynaptically, i.e. brain-derived neurotrophic factor regulates the properties of inhibitory interneurons, leading to increased excitability of dentate granule cells. We propose that through this mechanism, brain-derived neurotrophic factor can change the gating/filtering properties of the dentate gyrus for incoming information from the entorhinal cortex to hippocampus. This will have consequences for the recruitment of hippocampal neural circuitries both under physiological and pathological conditions, such as epileptogenesis. PMID- 11113303 TI - Apoptotic protein expression and activation of caspases is changed following cholinergic denervation and hippocampal sympathetic ingrowth in rat hippocampus. AB - Following cholinergic denervation of the hippocampus by medial septal lesions, an unusual neuronal reorganization occurs in which peripheral adrenergic fibers arising from superior cervical ganglia grow into the hippocampus (hippocampal sympathetic ingrowth). Recent studies suggest that a similar process, in which sympathetic noradrenergic axons invade the hippocampus, can occur in Alzheimer's disease patients. In the last few years, the occurrence of apoptotic cell death has been studied in Alzheimer's disease patients and in animal models of this disorder. Several studies suggest that the hippocampus is an important area to be considered for apoptotic cell death. In our studies in the rat hippocampus, we have measured the expression of inducers and blockers of apoptosis in membrane, cytosolic and mitochondrial fractions, and the activity of caspases. The level of cytosolic Fas was increased in cholinergic denervation compared to control and hippocampal sympathetic ingrowth groups. The membrane Fas ligand expression was significantly increased in hippocampal sympathetic ingrowth and in cholinergic denervation compared to the control group. The level of caspase-3 (CPP32) was increased in the cholinergic denervation group compared to control and hippocampal sympathetic ingrowth groups. The cytosolic expression of bcl-x was increased in hippocampal sympathetic ingrowth compared to control and cholinergic denervation. The cytosolic activity of caspase-3 appeared to be significantly decreased in hippocampal sympathetic ingrowth and increased in cholinergic denervation groups compared to control and cholinergic denervation/hippocampal sympathetic ingrowth, respectively. From the present results, we suggest that cholinergic denervation may be responsible for pro-apoptotic responses, while hippocampal sympathetic ingrowth may protect neurons from apoptosis in rat dorsal hippocampus. PMID- 11113304 TI - Seizures and neurodegeneration induced by 4-aminopyridine in rat hippocampus in vivo: role of glutamate- and GABA-mediated neurotransmission and of ion channels. AB - Infusion of the K(+) channel blocker 4-aminopyridine in the hippocampus induces the release of glutamate, as well as seizures and neurodegeneration. Since an imbalance between excitation and inhibition, as well as alterations of ion channels, may be involved in these effects of 4-aminopyridine, we have studied whether they are modified by drugs that block glutamatergic transmission or ion channels, or drugs that potentiate GABA-mediated transmission. The drugs were administered to anesthetized rats subjected to intrahippocampal infusion of 4 aminopyridine through microdialysis probes, with simultaneous collection of dialysis perfusates and recording of the electroencephalogram, and subsequent histological analysis. Ionotropic glutamate receptor antagonists clearly diminished the intensity of seizures and prevented the neuronal damage, but did not alter substantially the enhancement of extracellular glutamate induced by 4 aminopyridine. None of the drugs facilitating GABA-mediated transmission, including uptake blockers, GABA-transaminase inhibitors and agonists of the A type receptor, was able to reduce the glutamate release, seizures or neuronal damage produced by 4-aminopyridine. In contrast, nipecotate, which notably increased extracellular levels of the amino acid, potentiated the intensity of seizures and the neurodegeneration. GABA(A) receptor antagonists partially reduced the extracellular accumulation of glutamate induced by 4-aminopyridine, but did not exert any protective action. Tetrodotoxin largely prevented the increase of extracellular glutamate, the electroencephalographic epileptic discharges and the neuronal death in the CA1 and CA3 hippocampal regions. Valproate and carbamazepine, also Na(+) channel blockers that possess general anticonvulsant action, failed to modify the three effects of 4-aminopyridine studied. The N-type Ca(2+) channel blocker omega-conotoxin, the K(+) channel opener diazoxide, and the non-specific ion channel blocker riluzole diminished the enhancement of extracellular glutamate and slightly protected against the neurodegeneration. However, the two former compounds did not antagonize the 4 aminopyridine-induced epileptiform discharges, and riluzole instead markedly increased the intensity and duration of the disharges. Moreover, at the highest dose tested (8mg/kg, i.p.), riluzole caused a 75% mortality of the rats. We conclude that 4-aminopyridine stimulates the release of glutamate from nerve endings and that the resultant augmented extracellular glutamate is directly related to the neurodegeneration and is involved in the generation of epileptiform discharges through the concomitant overactivation of glutamate receptors. Under these conditions, a facilitated GABA-mediated transmission may paradoxically boost neuronal hyperexcitation. Riluzole, a drug used to treat amyotrophic lateral sclerosis, seems to be toxic when combined with neuronal hyperexcitation. PMID- 11113305 TI - Mitochondrial superoxide production in kainate-induced hippocampal damage. AB - The objective of this study was to determine the role of mitochondrial superoxide radical-mediated oxidative damage in seizure-induced neuronal death. Using aconitase inactivation as an index of superoxide production, we found that systemic administration of kainate in rats increased mitochondrial superoxide production in the hippocampus at times preceding neuronal death. 8-Hydroxy-2 deoxyguanosine, an oxidative lesion of DNA, was also increased in the rat hippocampus following kainate administration. Manganese(III) tetrakis(4-benzoic acid)porphyrin, a catalytic antioxidant, inhibited kainate-induced mitochondrial superoxide production, 8-hydroxy-2-deoxyguanosine formation and neuronal loss in the rat hippocampus. Kainate-induced increases of mitochondrial superoxide production and hippocampal neuronal loss were attenuated in transgenic mice overexpressing mitochondrial superoxide dismutase-2. We propose that these results demonstrate a role for mitochondrial superoxide production in hippocampal pathology produced by kainate seizures. PMID- 11113307 TI - Developmental increase in asynchronous GABA release in cultured hippocampal neurons. AB - Developmental changes in GABAergic synaptic transmission were examined in cultured hippocampal neurons using patch-clamp recordings and Ca(2+) imaging. In paired recordings, tetanization of the presynaptic GABAergic neuron with 80 pulses at either 40 or 80Hz was accompanied by tetanic depression of inhibitory postsynaptic responses. In neurons that had been cultured for more than two weeks, asynchronous inhibitory postsynaptic currents often appeared during the tetanus and continued for several seconds following stimulation. There was little asynchronous activity in neurons that had been cultured for shorter times. However, no age-related changes were observed in the amplitude of single synchronous inhibitory postsynaptic currents, paired-pulse depression or post tetanic potentiation of inhibitory postsynaptic currents. Following equimolar replacement of extracellular Ca(2+) with strontium ions (Sr(2+)), single autaptic inhibitory postsynaptic currents were depressed in amplitude and asynchronous inhibitory postsynaptic currents were present on the decaying phase. Sr(2+) induced asynchronous inhibitory postsynaptic currents showed no dependence on age in culture. Imaging of Ca(2+) in single GABAergic boutons was performed by including Fluo-3 in the patch pipette. During action potential firing induced by stimulating at 80Hz for 1s, intracellular calcium [Ca(2+)](i) increased rapidly in individual boutons. Following the stimulus, [Ca(2+)](i) decayed back to baseline within 10-15s. The half-time of decay increased from 1. 7+/-0.2s at 15days in vitro to 4.0+/-0.2s at 30days in vitro (P<0. 05), with a developmental profile that closely matched the increase in asynchronous inhibitory postsynaptic currents. We propose that the increase in tetanus-induced asynchronous GABA release during the first month of synapse maturation in vitro is caused by a slowing of the Ca(2+)-clearing mechanisms in the GABAergic boutons. This results in larger and more prolonged elevations of [Ca(2+)](i) during tetanic stimulation, which leads to enhanced asynchronous transmitter release. We propose that the results of this study demonstrate a potentially important aspect of synapse maturation during development, and also imply that GABA release is up regulated in conditions of decreased Ca(2+) buffering and clearing. PMID- 11113306 TI - Neuronal activity and stress differentially regulate hippocampal and hypothalamic corticotropin-releasing hormone expression in the immature rat. AB - Corticotropin-releasing hormone, a major neuromodulator of the neuroendocrine stress response, is expressed in the immature hippocampus, where it enhances glutamate receptor-mediated excitation of principal cells. Since the peptide influences hippocampal synaptic efficacy, its secretion from peptidergic interneuronal terminals may augment hippocampal-mediated functions such as learning and memory. However, whereas information regarding the regulation of corticotropin-releasing hormone's abundance in CNS regions involved with the neuroendocrine responses to stress has been forthcoming, the mechanisms regulating the peptide's levels in the hippocampus have not yet been determined. Here we tested the hypothesis that, in the immature rat hippocampus, neuronal stimulation, rather than neuroendocrine challenge, influences the peptide's expression. Messenger RNA levels of corticotropin-releasing hormone in hippocampal CA1, CA3 and the dentate gyrus, as well as in the hypothalamic paraventricular nucleus, were determined after cold, a physiological challenge that activates the hypothalamic pituitary adrenal system in immature rats, and after activation of hippocampal neurons by hyperthermia. These studies demonstrated that, while cold challenge enhanced corticotropin-releasing hormone messenger RNA levels in the hypothalamus, hippocampal expression of this neuropeptide was unchanged. Secondly, hyperthermia stimulated expression of hippocampal immediate-early genes, as well as of corticotropin-releasing hormone. Finally, the mechanism of hippocampal corticotropin-releasing hormone induction required neuronal stimulation and was abolished by barbiturate administration. Taken together, these results indicate that neuronal stimulation may regulate hippocampal corticotropin-releasing hormone expression in the immature rat, whereas the peptide's expression in the hypothalamus is influenced by neuroendocrine challenges. PMID- 11113308 TI - Microinfusion of the non-competitive N-methyl-D-aspartate receptor antagonist MK 801 (dizocilpine) into the dorsal hippocampus of wistar rats does not affect latent inhibition and prepulse inhibition, but increases startle reaction and locomotor activity. AB - Latent inhibition (the retarded conditioning to a stimulus following its repeated non-reinforced pre-exposure) and prepulse inhibition (the reduction in the startle response to an intense acoustic stimulus when this stimulus is immediately preceded by a prepulse) reflect cognitive and sensorimotor gating processes, respectively, and are deficient in schizophrenic patients. The disruption of latent inhibition and prepulse inhibition in the rat is used as an animal model for the attentional deficits associated with schizophrenia. The present study tested the extent to which latent inhibition and prepulse inhibition, startle reaction and locomotor activity in the open field were affected by infusing the non-competitive N-methyl-D-aspartate receptor antagonist MK-801 (dizocilpine) into the dorsal hippocampus of Wistar rats. We used the same dose of MK-801 (6.25microg/0.5microl per side) previously found to be effective in the disruption of prepulse inhibition when infused into the dorsal hippocampus of Sprague-Dawley rats [Bakshi V. P. and Geyer M. A. (1998) J. Neurosci. 18, 8394 8401; Bakshi V. P. and Geyer M. A. (1999) Neuroscience 92, 113-121]. Bilateral infusion of MK-801 into the dorsal hippocampus did not disrupt latent inhibition. Furthermore, in contrast to previous studies, we failed to find a significant disruption of prepulse inhibition after MK-801 infusion into the dorsal hippocampus, although MK-801 infusion was effective in increasing the startle amplitude as well as locomotor activity in an open field. From our results, we suggest that N-methyl-D-aspartate receptor-mediated processes within the dorsal hippocampus are not necessary for the normal maintenance of the attentional processes reflected by latent inhibition and prepulse inhibition. PMID- 11113309 TI - Molecular evidence for the involvement of NR2B subunit containing N-methyl-D aspartate receptors in the development of morphine-induced place preference. AB - The present study was designed to clarify the role of the NR1, NR2A and NR2B subunits of N-methyl-D-aspartate receptors in the development of morphine-induced place preference using specific antibodies to N-methyl-D-aspartate receptor subunits in the mouse. An i.c.v. injection with specific antibodies against the carboxyl-terminal region of either NR1, NR2A or NR2B subunits caused the attenuation of seizures induced by i.v. infusion of N-methyl-D-aspartate in the mouse. Using western blotting, NR1, NR2A and NR2B subunits were found to be highly expressed in the cerebral cortex and hippocampus of the mouse brain, which are key areas in producing seizures regulated by N-methyl-D-aspartate receptors. These findings indicate that all three subunits of the N-methyl-D-aspartate receptor in these areas are likely to be critical for the seizure induced by i.v. infusion of N-methyl-D-aspartate. Furthermore, present data provide evidence that these antibodies when given into the brain specifically act on the target sites, resulting in the blockade of the respective N-methyl-D-aspartate receptor subunit. Under these conditions, i.c.v. treatment with the antibody against NR2B subunits abolished the morphine-induced place preference, whereas antibodies against NR1 and NR2A subunits did not affect the rewarding effect of morphine. Furthermore, the NR2B subunit protein was specifically up-regulated in the limbic forebrain of morphine-conditioned mice, whereas the protein levels of either NR1 or NR2A subunits were not altered.From these results, we suggest that the stimulation of the carboxyl-terminal region of NR2B subunits in the limbic forebrain may contribute to the development of morphine-induced rewarding effect. PMID- 11113311 TI - Kappa-opioid receptor activation prevents alterations in mesocortical dopamine neurotransmission that occur during abstinence from cocaine. AB - In vivo microdialysis was used to characterize basal dopamine dynamics and cocaine-evoked dopamine levels in the medial prefrontal cortex of male Sprague Dawley rats that had previously received once daily injections of cocaine (days 1 5; 20mg/kg, i.p.) in combination with the selective kappa-opioid receptor agonist U-69593 (days 3-5; 0.32mg/kg, s.c.) or its vehicle. The influence of these treatments on [3H]dopamine uptake in medial prefrontal cortex synaptosomes was also determined. Three days following the cessation of drug treatment, animals with prior history of cocaine administration exhibited enhanced psychomotor stimulation in response to a subsequent cocaine challenge. This effect was not apparent in animals that had previously received the cocaine treatment regimen in combination with the kappa-opioid receptor agonist U-69593. Cocaine challenge increased prefrontal dopamine levels in all pretreatment groups, but cocaine-pre exposed animals had lower cocaine-evoked dopamine levels and higher basal in vivo extraction fraction, indicative of an increase in basal dopamine uptake relative to controls. Pretreatment with U-69593 prevented these effects of cocaine. Measurement of [3H]dopamine uptake in synaptosomes revealed a significant increase in uptake three days after the cessation of cocaine treatment. No increase in uptake was observed in animals that had received the cocaine treatment regimen in combination with U-69593. These results demonstrate that the early phase of abstinence from cocaine is associated with marked alterations in medial prefrontal cortex dopamine neurotransmission and that these neuroadaptations are prevented by the activation of kappa-opioid receptors. Furthermore, they raise the possibility that mesocortical dopamine neurons may be an important neural substrate upon which kappa-opioid agonists act to prevent the development of cocaine-induced behavioral sensitization. PMID- 11113310 TI - N-Methyl-D-aspartate receptors and p38 mitogen-activated protein kinase are required for cAMP-dependent cyclase response element binding protein and Elk-1 phosphorylation in the striatum. AB - In vivo cyclic adenosine monophosphate (cAMP)-induced N-methyl-D-aspartate receptor and mitogen-activated protein kinase activation was investigated in the dorsal striatum by semiquantitative immunocytochemistry. Intracerebroventricular infusion of 8-bromo-adenosine 3',5'-cyclic monophosphorothioate, Sp isomer (Sp-8 Br-cAMPS), increased phosphorylated cAMP-responsive element binding protein, phosphorylated Elk-1 and Fos immunoreactivity in a dose-dependent manner. Intracerebroventricular infusion of the N-methyl-D-aspartate antagonist, MK801, decreased, but tetrodotoxin or the mitogen-activated extracellular-regulated kinase inhibitor, PD98059, did not affect Sp-8-Br-cAMPS-induced phosphorylated c AMP-responsive element binding protein, phosphorylated Elk-1, phosphorylated extracellular-signal-regulated kinase and Fos immunoreactivity. The p38 mitogen activated protein kinase inhibitor, SB203580, decreased the Sp-8-Br-cAMPS-induced increase in all markers, except phosphorylated extracellular-signal-regulated kinase, in a dose-dependent manner. We suggest that N-methyl-D-aspartate receptors couple c-AMP to phosphorylation events and immediate early gene induction in the nucleus of striatal medium spiny neurons. These events are mediated by crosstalk between protein kinase A and mitogen-activated protein kinase cascades in vivo. PMID- 11113312 TI - Dopamine D(1A) receptor function in a rodent model of tardive dyskinesia. AB - Tardive dyskinesia develops as a common complication of long-term neuroleptic use. The emergence of such dyskinesias may reflect a shift in the balance of dopamine D(1) and D(2) receptor-mediated activity, with a relative increase in activity in the D(1) receptor-regulated direct striatonigral pathway. In rats, chronic treatment with the antipsychotic fluphenazine triggers a syndrome of vacuous chewing movements, which are attenuated by dopamine D(1) receptor antagonists. A similar syndrome can be seen in drug-naive animals following acute administration of selective dopamine D(1) receptor agonists. However, not all dopamine D(1) receptor agonists elicit these mouth movements. Thus, some investigators have suggested the existence of novel subtypes of the dopamine D(1) receptor. In these studies, we sought to clarify the role of the dopamine D(1A) receptor in vacuous chewing movements induced both by the selective dopamine D(1) receptor agonist SKF 38393, as well as by chronic neuroleptic administration, using in vivo oligonucleotide antisense to dopamine D(1A) receptor messenger RNA. Intrastriatal antisense treatment significantly and selectively attenuated striatal dopamine D(1) receptor binding, accompanied by reductions in SKF 38393- and chronic fluphenazine-induced vacuous chewing movements. These findings suggest that the dopamine D(1A) receptor plays an important role in the expression of vacuous chewing movements in a rodent model of tardive dyskinesia and may contribute to the pathogenesis of the human disorder. This may have important implications for the treatment of tardive dyskinesia in humans. PMID- 11113313 TI - Localization of glutamatergic/aspartatergic neurons projecting to the hypothalamic paraventricular nucleus studied by retrograde transport of [3H]D aspartate autoradiography. AB - Morphological and functional data indicate that glutamatergic innervation of the hypothalamic paraventricular nucleus plays an important role in the control of this prominent cell group. Sources of this neural input are unknown. The present investigations were aimed at studying this question. The retrograde tracer [3H]D aspartate, which is selectively taken up by the terminals of neurons that use glutamate or aspartate as a neurotransmitter, and is retrogradely transported to their perikarya, was injected into the paraventricular nucleus. The brain was examined for labelled neurons visualized by autoradiography. Labelled neurons were detected in the paraventricular nucleus itself, in several hypothalamic areas including medial and lateral preoptic area, suprachiasmatic nucleus, anterior hypothalamic area, ventromedial nucleus, dorsomedial nucleus, lateral hypothalamic area, posterior part of arcuate nucleus, ventral premammillary nucleus and supramammillary nucleus. Outside the hypothalamus labelled neurons were found in the thalamic paraventricular nucleus and in certain telencephalic regions including lateral septum, bed nucleus of the stria terminalis and amygdala. All of them are known to project to the hypothalamic paraventricular nucleus. We failed to detect labelled neurons in the lower brainstem. From these findings we conclude that firstly, there are glutamatergic/aspartatergic interneurons in the paraventricular nucleus; secondly, all intrahypothalamic and telencephalic, but not lower brainstem afferents to this nucleus contain glutamatergic/aspartatergic fibres; and thirdly, the glutamatergic/aspartatergic innervation of this heterogeneous cell group is extremely complex. PMID- 11113314 TI - Synchronized release of dopamine and serotonin in the medial and lateral hypothalamus of rats. AB - A positive linear correlation between dopamine and serotonin release was found in the ventromedial hypothalamus and in the lateral hypothalamic area in fasting rats and in fed rats during intermeal intervals. Dopamine release in the ventromedial hypothalamus positively correlated with dopamine and serotonin release in the lateral hypothalamic area, which occurred only during intermeal intervals and was non-significant during the meal consumption periods or during fasting. Meal size correlated significantly only with a decrease in serotonin release in the lateral hypothalamic area. The study was designed to evaluate the relationship between dopamine and serotonin release in these hypothalamic areas and their dependence on feeding status. Microdialysis was performed simultaneously via two probes, one in the ventromedial hypothalamus and the other in the contralateral lateral hypothalamic area, of freely moving male lean Zucker rats over 24h with preserved light and dark phase, either with ad libitum access to food and water, or when no food was available. Dopamine and serotonin concentrations were measured by high-performance liquid chromatography with electrochemical detection in 20-min dialysis samples. Time-series analysis was applied to determine linear correlations between monoamines and in relation to food intake. Data showed that release of dopamine and serotonin is synchronized within the ventromedial hypothalamus and lateral hypothalamic area, particularly in the dark phase and when no food was ingested. However, synchronized release of monoamines between these nuclei occurred only during intermeal intervals: the periods of satiety. These findings suggest a tight relationship between dopaminergic and serotonergic systems of the lateral hypothalamic area and ventromedial hypothalamus, which is influenced by the feeding state and which may be involved in maintaining the balance within and between the centers of the parasympathetic and sympathetic nervous systems. The data also illustate that food intake is coupled unequivocally to the release of dopamine and serotonin in the hypothalamus, suggesting it as a mechanism of activation of postsynaptic neurons associated with new metabolic status. PMID- 11113315 TI - Developmental regulation of tryptophan hydroxylase messenger RNA expression and enzyme activity in the raphe and its target fields. AB - Tryptophan hydroxylase is the rate-limiting enzyme in the synthesis of serotonin and during development, brain serotonin levels and tryptophan hydroxylase activities increase. Increased tryptophan hydroxylase activity could result from alterations in tryptophan hydroxylase messenger RNA levels, translation, and/or post-translational regulation. Tryptophan hydroxylase messenger RNA levels in the dorsal raphe nucleus increased 35-fold between embryonic day 18 and postnatal day 22, measured by quantitative in situ hybridization, then decreased by 40% between postnatal days 22 and 61. These changes correlated with tryptophan hydroxylase enzyme activities in the raphe nuclei as expected, but not in cortical or hippocampal targets. Tryptophan hydroxylase messenger RNA expression in the nucleus raphe obscuris increased 2.5-fold between postnatal days 8 and 22 but did not correlate with enzyme activity in the spinal cord. Using an in vitro model of serotonergic raphe neuron differentiation, serotonergic differentiation was associated with an increase in both tryptophan hydroxylase promoter activity and protein expression. In vivo, tryptophan hydroxylase messenger RNA levels per single cell and per brain section were correlated during development up to postnatal day 22, but not beyond for both the dorsal raphe nucleus and nucleus raphe obscuris. Between postnatal days 22 and 61 single cell levels of tryptophan hydroxylase messenger RNA in the dorsal raphe nucleus did not change yet the levels per brain section significantly decreased by 40%. During the same period in the nucleus raphe obscuris, tryptophan hydroxylase messenger RNA levels per single cell signifcantly increased by 30% yet levels per brain section did not change. Comparison of tryptophan hydroxylase messenger RNA levels per cell and per brain section indicated a serotonergic loss between postnatal days 22 and 61 in both the dorsal raphe nucleus and nucleus raphe obscuris and may reflect either a loss of neurotransmitter phenotype or cell death. This study is the first to characterize the expression of brain tryptophan hydroxylase messenger RNA during rat development. In addition, this study is the first to report the activity of tryptophan hydroxylase in the spinal cord and hippocampus in the embryonic and neonatal rat. Together, the data provide a better understanding of the intricate relationship between patterns of tryptophan hydroxylase messenger RNA expression and enzyme activity. PMID- 11113316 TI - In vivo expression of the intermediate filament peripherin in rat motoneurons: modulation by inhibitory and stimulatory signals. AB - Peripherin is a type III intermediate filament which, in contrast to the neurofilaments, is strongly up-regulated after nerve injury. Although peripherin expression is stimulated in vitro by neurotrophins and cytokines, little is known about its in vivo regulation. In this report, we show that the in vivo down regulation of peripherin expression to normal levels during regeneration closely correlates with target reconnection in rat facial motoneurons. Prevention of reconnection, by transection and suture, results in the persistence of strong peripherin expression for prolonged periods of up to 11months. This contrasts with the modulation of the p75 low-affinity neurotrophin receptor, whose expression returns to normal even in the absence of reconnection. We further demonstrate that blockade of the axonal transport in non-injured motoneurons increases the expression of peripherin. Blockade of the axonal transport simultaneously to, or after injury of, facial motoneurons does not abolish the axotomy-induced peripherin up-regulation. These data demonstrate that the in vivo expression of peripherin is normally restrained by a distal retrogradely transported inhibitory signal. Thus, peripherin up-regulation results primarily from a lack of supply in this factor. Our results show that stimulatory factors released at the injury site are not required for the initial up-regulation and maintenance of high peripherin expression. However, they appear to enhance this increase during the acute post-lesion phase. Peripherin expression is thus finely tuned by both glial cell-derived stimulatory and distal inhibitory signals that reflect neuron-target interactions. PMID- 11113317 TI - Adrenergic sensitivity of neurons with non-periodic firing activity in rat injured dorsal root ganglion. AB - In this study, we compared the sensitivity of non-periodically and periodically active neurons in chronically compressed dorsal root ganglion in rats to norepinephrine and sympathetic stimulation. Forty-nine of 58 (84.5%) neurons with non-periodic activity showed responses to norepinephrine, whereas only five of 48 (10.4%) neurons with periodic activity displayed any response. The dose-response relationship of norepinephrine to the irregular burst pattern neurons shifted towards the left significantly compared to that of the periodic activity neurons. Responses to norepinephrine became apparent in eight neurons after their periodic firing activity was transformed into the non-periodic firing activity through the increase in Ca(2+). Changes in the time-response curves indicate a higher sensitivity of irregular burst pattern neurons to sympathetic stimulation than the periodic activity neurons. Finally, deterministic dynamics contained within the interburst interval series for non-periodic activity were identified. From these results, we suggest that the non-periodic activity neurons have a higher adrenergic sensitivity than those displaying periodic activity, and that this sensitivity may depend on the deterministic chaos within its firing dynamic system. PMID- 11113318 TI - Localization of N-methyl-D-aspartate NR2B subunits on primary sensory neurons that give rise to small-caliber sciatic nerve fibers in rats. AB - In the present study we have used immunohistochemical staining and retrograde tracing techniques to investigate the relationship between the N-methyl-D aspartate receptor NR2B subunits and small-diameter primary afferent dorsal root ganglion neurons that give rise to the sciatic nerve fibers. Three days after an intra-sciatic nerve injection of tetramethyl rhodamine isothiocyanate-conjugated wheat germ agglutinin which labels small-diameter primary afferents, many NR2B and wheat germ agglutinin-double-labeled cells ( approximately 70% of wheat germ agglutinin-labeled neurons) were observed in the L5 dorsal root ganglia. Three days after an intra-sciatic nerve injection of fluorescein isothiocyanate conjugated Bandeiraea simplicifolia agglutinin isolectin B4 which labels predominantly non-peptidergic C-fiber primary afferents, NR2B and Bandeiraea simplicifolia agglutinin isolectin B4 double-labeled neurons ( approximately 90% of Bandeiraea simplicifolia agglutinin isolectin B4-labeled neurons) were also observed in the L5 dorsal root ganglion. Three days after an intra-sciatic nerve injection of fluorescein isothiocyanate-conjugated cholera toxin B subunit, only approximately 40% of cholera toxin B subunit-labeled neurons were NR2B positive and those labeled neurons tended to be small-sized. When calcitonin gene-related peptide and NR2B were labeled by a double immunofluorescent staining technique, we found that the majority of calcitonin gene-related peptide-positive neurons was NR2B immunoreactive (>90% of calcitonin gene-related peptide-positive neurons, and approximately 60% of NR2B-positive neurons) as well. Size frequency analysis also demonstrated that NR2B subunits were predominantly localized on the small and medium-sized neurons. These results suggest that NR2B subunits are predominantly expressed on small diameter primary afferents, and these NR2B containing N-methyl-D-aspartate receptors may play a role in the modulation of neurotransmitter release from primary afferent terminals. PMID- 11113319 TI - Disparate spinal and supraspinal opioid antinociceptive responses in beta endorphin-deficient mutant mice. AB - The role of endogenous opioid systems in the analgesic response to exogenous opiates remains controversial. We previously reported that mice lacking the peptide neurotransmitter beta-endorphin, although unable to produce opioid mediated stress-induced antinociception, nevertheless displayed intact antinociception after systemic administration of the exogenous opiate morphine. Morphine administered by a peripheral route can activate opioid receptors in both the spinal cord and brain. However, beta-endorphin neuronal projections are confined predominantly to supraspinal nociceptive nuclei. Therefore, we questioned whether the absence of beta-endorphin would differentially affect antinociceptive responses depending on the route of opiate administration. Time- and dose-response curves were obtained in beta-endorphin-deficient and matched wild-type C57BL/6 congenic control mice using the tail-immersion/withdrawal assay. Null mutant mice were found to be more sensitive to supraspinal (i.c.v.) injection of the micro-opioid receptor-selective agonists, morphine and D-Ala(2) MePhe(4)-Gly-ol(5) enkephalin. In contrast, the mutant mice were less sensitive to spinal (i.t.) injection of these same drugs. Quantitative receptor autoradiography revealed no differences between genotypes in the density of mu, delta, or kappa opioid receptor binding sites in either the spinal cord or pain relevant supraspinal areas. Thus we report that the absence of a putative endogenous ligand for the mu-opioid receptor results in opposite changes in morphine sensitivity between discrete areas of the nervous system, which are not simply caused by changes in opioid receptor expression. PMID- 11113320 TI - Vanilloid receptor 1-like receptor-immunoreactive primary sensory neurons in the rat trigeminal nervous system. AB - Immunohistochemistry for vanilloid receptor 1-like receptor (VRL-1), a candidate transducer for high-threshold noxious heat, was performed on rat trigeminal primary sensory neurons. The immunoreactivity was detected in 14% of the trigeminal ganglion cell bodies, while the neurons in the mesencephalic trigeminal tract nucleus were almost devoid of it (0.5%). The immunoreactive neurons in the trigeminal ganglion were mostly of medium to large size (mean+/ S.D. of 956+/-376microm(2)). Nerve bundles in the tooth pulp, periodontal ligament, facial skin and oral mucosa contained VRL-1-positive smooth nerve fibers. The immunoreactivity could not be traced to the isolated nerve fibers, except in the tooth pulp. In the brainstem trigeminal nuclear complex, a notable concentration of the immunoreactivity was seen in laminae I and II of the medullary dorsal horn. Thirty-seven per cent of the trigeminal ganglion neurons retrogradely labeled from the tooth pulp exhibited VRL-1 immunoreactivity, while the immunoreactivity was detected in only 9% of those labeled from the skin. Co expression of calcitonin gene-related peptide was common among the VRL-1 immunoreactive tooth pulp neurons (45%) and cutaneous neurons (25%). Moreover, as many as 41% of the VRL-1-immunoreactive tooth pulp neurons co-expressed parvalbumin immunoreactivity. Parvalbumin immunoreactivity was never detected in the VRL-1-immunoreactive cutaneous neurons. From the findings of the present study, we propose that large primary neurons responding to high-threshold noxious heat are abundant in the tooth pulp, but not in the facial skin. PMID- 11113321 TI - Differential effects of capsaicin on rat visceral sensory neurons. AB - Nodose neurons play an important role in the regulation of visceral function. Recent studies demonstrated that about 80% of these neurons contain messenger RNA for the capsaicin receptor, a heat-sensitive ion channel. Nodose neurons express voltage-sensitive sodium currents that can be differentiated based on their sensitivity to tetrodotoxin. Considering the potential role of tetrodotoxin resistant sodium currents in somatosensory neurons, sodium channel expression and sodium currents were studied in nodose neurons. The results were correlated with the response to capsaicin. Nodose neurons contain messenger RNA for the tetrodotoxin-resistant sodium channel PN3. Consistent with these findings, about half of the neurons predominantly expressed tetrodotoxin-resistant sodium currents. In 54% (47/87) of the cells, capsaicin triggered an increase in intracellular calcium. Similarly, in 42% (18/43) of the cells, capsaicin elicited an inward current. There was no relationship between cell size (r=0.07) or sodium current properties (r=0.14) and the response to capsaicin. Micromolar concentrations of capsaicin inhibited voltage-dependent sodium, calcium and potassium currents. This effect was use dependent and did not involve the capsaicin receptor. In conclusion, capsaicin changed the excitability of visceral sensory neurons by blocking voltage-dependent ion channels, an effect that may contribute to the analgesic properties of capsaicin. PMID- 11113322 TI - Monocyte chemoattractant protein-1 is a mediator of acute excitotoxic injury in neonatal rat brain. AB - Monocyte chemoattractant protein-1 is a chemokine with potent monocyte activating and chemotactic effects. Monocyte chemoattractant protein-1 gene and protein expression is rapidly up-regulated in response to a variety of acute and chronic central nervous system disorders. The activation and recruitment of microglia and monocytes into areas of inflammation may play a critical role in the pathogenesis of acute brain injury. Monocyte chemoattractant protein-1 could be a pathophysiologically important mediator of the microglial and monocyte responses in the brain. Using a well-characterized model of acute excitotoxic brain injury in neonatal rats, experiments were designed to evaluate whether monocyte chemoattractant protein-1 plays a role in the progression of tissue damage. Direct co-administration of recombinant monocyte chemoattractant protein-1 with the excitotoxin N-methyl-D-aspartate exacerbated injury, both in the striatum and in the hippocampus, by 55% and 167%, respectively. Complementary experiments to determine the effect of functional inhibition of monocyte chemoattractant protein 1, using an anti-monocyte chemoattractant protein-1-neutralizing antibody, revealed that co-administration of the antibody with N-methyl-D-aspartate attenuated tissue injury in the striatum and hippocampus by 57% and 39%, respectively.Together, these data suggest that monocyte chemoattractant protein-1 is a mediator of acute excitotoxic brain injury in neonatal rats and that inflammatory mechanisms contribute significantly to the pathogenesis of acute neonatal brain injury. Whether chemokines are pathophysiologically relevant mediators of neuronal injury in human neonates remains to be determined. PMID- 11113323 TI - Hyperalgesia due to nerve injury: role of neutrophils. AB - The hypothesis that the early inflammatory cell, the neutrophil, contributes to the hyperalgesia resulting from peripheral nerve injury was tested in rats in which the sciatic nerve was partially transected on one side. The extent and time course of neutrophilic infiltration of the sciatic nerve and innervated paw skin after partial nerve damage was characterized using immunocytochemistry. The number of endoneurial neutrophils was significantly elevated in sections of operated nerve compared to sections of sham-operated nerve for the entire period studied, i.e. up to seven days post-surgery. This considerable elevation in endoneurial neutrophil numbers was only observed at the site of nerve injury. Depletion of circulating neutrophils at the time of nerve injury significantly attenuated the induction of hyperalgesia. However, depletion of circulating neutrophils at day 8 post-injury did not alleviate hyperalgesia after its normal induction. It is concluded that endoneurial accumulation of neutrophils at the site of peripheral nerve injury is important in the early genesis of the resultant hyperalgesia. The findings support the notion that a neuroimmune interaction occurs as a result of peripheral nerve injury and is important in the subsequent development of neuropathic pain. PMID- 11113324 TI - Intrathecal S-nitroso-N-acetylpenicillamine and L-cysteine attenuate nerve injury induced allodynia through noradrenergic activation in rats. AB - Spinal norepinephrine release and activation of spinal alpha(2)-adrenergic receptors represent important components of descending control of nociception. Recent studies have shown that nitric oxide is capable of stimulating neuronal norepinephrine release in the presence of thiol-containing compounds such as L cysteine. In the present study, we tested a hypothesis in a rodent model of neuropathic pain that intrathecal injection of the nitric oxide donor S-nitroso-N acetylpenicillamine and L-cysteine produces an antiallodynic action mediated by the spinal alpha(2)-adrenergic receptors. Allodynia was induced in rats by ligation of the left lumbar L5/L6 spinal nerves. Mechanical allodynia was quantified by application of von Frey filaments to the left hindpaw. Intrathecal injection of 20-100microg of S-nitroso-N-acetylpenicillamine in the presence of 200microg of L-cysteine, but not D-cysteine, dose-dependently attenuated the allodynia. Intrathecal injection of a combination of 100microg of S-nitroso-N acetylpenicillamine and 50-200microg of L-cysteine also inhibited the allodynia in a dose-dependent manner. Pretreatment with a nitric oxide scavenger, carboxy PTIO, or depletion of norepinephrine with a specific neurotoxin, N-(2 chloroethyl)-N-ethyl-2-bromobenzylamine, prevented the antiallodynic action of intrathecal S-nitroso-N-acetylpenicillamine and L-cysteine. Furthermore, the antiallodynic effect produced by intrathecal injection of a combination of S nitroso-N-acetylpenicillamine and L-cysteine was abolished by pretreatment with intrathecal injection of a non-specific alpha-adrenergic receptor antagonist, phentolamine, or an alpha(2) receptor antagonist, idazoxan. This study provides the first functional evidence that spinal nitric oxide interacts with the thiol containing compounds to produce an antiallodynic effect in neuropathic pain. We propose that such an action is mediated by endogenous norepinephrine and spinal alpha(2)-adrenergic receptors. PMID- 11113325 TI - Nerve injury-associated kinase: a sterile 20-like protein kinase up-regulated in dorsal root ganglia in a rat model of neuropathic pain. AB - Partial injury of the rat sciatic nerve elicits a variety of characteristic chemical, electrophysical and anatomical changes in primary sensory neurons and constitutes a physiologically relevant model of neuropathic pain. To elucidate molecular mechanisms that underlie the physiology of neuropathic pain, we have used messenger RNA differential display to identify genes that exhibit increased ipsilateral expression in L4/5 dorsal root ganglia, following unilateral partial ligation of the rat sciatic nerve. One set of partial complementary DNA clones identified in this screen was found to encode a protein kinase, nerve injury associated kinase. Cloning of the full-length human nerve injury-associated kinase complementary DNA, together with recombinant expression analysis, reveal nerve injury-associated kinase to be a functional member of a subgroup of sterile 20-like protein kinases characterised by the presence of a putative carboxy terminal autoregulatory domain. Induction of nerve injury-associated kinase expression in dorsal root ganglia in the rat neuropathic pain model was confirmed by quantitative reverse transcription-polymerase chain reaction, and RNA in situ hybridization analysis revealed enhanced levels of nerve injury-associated kinase within neurons.Together, our data implicate nerve injury-associated kinase as a novel upstream component of an intracellular signalling cascade that is up regulated in dorsal root ganglia neurons in response to sciatic nerve injury. PMID- 11113326 TI - Prenatal exposure to diethylstilbestrol decreases the number of estrogen receptor alpha-containing neurons innervating the ovary in rat celiac ganglion. AB - Recently, we reported that, in rats, transplacental exposure to diethylstilbestrol, a potent synthetic estrogen, decreases the density of the ovarian sympathetic nerve network compared to that in control rats not exposed to diethylstilbestrol. To clarify the mechanism of the decrease in the density, we performed a series of experiments using rats prenatally exposed to diethylstilbestrol and unexposed control rats. First, a retrograde tract tracer, Fast Blue, was microinjected into the ovaries of both groups of rats at four months of age, and the number of Fast Blue-positive neurons in the celiac ganglion was counted. The number of neurons in the ganglion was smaller in diethylstilbestrol-exposed rats than in the controls. Second, double labeling of the neurons with antibody against estrogen receptor alpha and Fast Blue in the celiac ganglion was carried out in both groups of rats. The results showed that estrogen receptor alpha-containing neurons in the ganglion innervated the ovary, and their number was decreased selectively by prenatal diethylstilbestrol exposure. Finally, a decrease in the celiac ganglion volume in rats prenatally exposed to diethylstilbestrol was also detected on day 23 of gestation, as compared to the volume in the control rats. From these observations, we propose that prenatal exposure to diethylstilbestrol can induce a loss of estrogen receptor alpha-containing neurons innervating the ovary during development, resulting in paucity of the neural network in the ovary. PMID- 11113327 TI - Anthelmintic actions on homomer-forming nicotinic acetylcholine receptor subunits: chicken alpha7 and ACR-16 from the nematode Caenorhabditis elegans. AB - Two homomer-forming nicotinic acetylcholine receptor subunits with 47% identity in their amino acid sequences were employed to compare the actions of cholinergic anthelmintics and ivermectin on expressed vertebrate and nematode nicotinic receptors of known molecular composition. Voltage-clamp electrophysiology was used to study recombinant nicotinic receptors expressed in Xenopus laevis oocytes following nuclear injection of cDNA encoding either chicken alpha7 or Caenorhabditis elegans ACR-16 (Ce21) subunits. Butamisole, morantel and metyridine were without agonist actions on either alpha7 or ACR-16 nicotinic receptors in the range 10nM-1mM. However, butamisole (pIC(50)=4.9 for both alpha7 and ACR-16) and morantel (pIC(50)=5.6 for alpha7 and 5.7 for ACR-16) antagonized responses of both alpha7 and ACR-16 receptors to acetylcholine. Metyridine (1mM) did not affect responses to acetylcholine of either receptor. Oxantel was without agonist actions on ACR-16, but was an acetylcholine antagonist (pIC(50)=5.4). In contrast, it was found to have low efficacy agonist action (pEC(50)=4.4) on alpha7 at concentrations in the range 10-300microM. In agreement with a previous study, ivermectin (30microM), an agonist of L-glutamate-gated chloride channels, enhanced the amplitude of responses to acetylcholine of alpha7 nicotinic receptors. However, this same concentration of ivermectin (30microM) did not potentiate the acetylcholine-induced responses of ACR-16, but rather resulted in a slight attenuation. We conclude that oxantel and ivermectin have identified new pharmacological differences between the chicken alpha7 nicotinic receptor and its C. elegans homologue ACR-16. PMID- 11113328 TI - Role of calcium in neurotransmitter release evoked by alpha-latrotoxin or hypertonic sucrose. AB - At the synapse, neurotransmitter release is triggered physiologically by Ca(2+) influx through voltage-gated Ca(2+) channels. Non-physiologically, release can be evoked by a potent neurotoxin, alpha-latrotoxin, and by hypertonic sucrose. Controversy has arisen on whether release evoked by alpha-latrotoxin and hypertonic sucrose requires extracellular Ca(2+) or Ca(2+) from intracellular stores. Using synaptosomes, we have studied the Ca(2+) dependence of alpha latrotoxin and sucrose action in different neurotransmitter systems. In agreement with previous data, no requirement for extracellular Ca(2+) in sucrose-induced secretion of norepinephrine, dopamine, glutamate or GABA was detected. Unexpectedly, we observed large differences between these neurotransmitters in the Ca(2+) dependence of alpha-latrotoxin-stimulated release: norepinephrine release required Ca(2+), dopamine release was only partially Ca(2+) dependent, and glutamate and GABA release did not require Ca(2+). To test if Ca(2+) derived from intracellular Ca(2+) stores participates in neurotransmitter release triggered by alpha-latrotoxin or hypertonic sucrose, we employed thapsigargin, a Ca(2+)-ATPase inhibitor that empties Ca(2+) stores. Thapsigargin did not induce neurotransmitter release, nor did it inhibit subsequent release stimulated by KCl depolarization, hypertonic sucrose or alpha-latrotoxin. However, intracellular Ca(2+) performs an important regulatory function, since thapsigargin increased the size of the readily releasable pool as measured by stimulation with hypertonic sucrose. This effect required extracellular Ca(2+) and protein kinase C, suggesting that depletion of internal Ca(2+) stores leads to store-operated Ca(2+) entry. The resulting Ca(2+) influx does not trigger release by itself, but activates protein kinase C that increases the readily releasable pool of neurotransmitters. Our data show that internal and external Ca(2+) is not acutely involved in hypertonic sucrose-evoked neurotransmitter release, while alpha latrotoxin-triggered release requires external Ca(2+) for a subset of neurotransmitters. Although internal Ca(2+) is not essential for release, it modulates its extent, implying that the emptying of intracellular stores by activation of presynaptic receptors plays an important regulatory role in neurotransmitter release. PMID- 11113329 TI - Linear coupling between functional magnetic resonance imaging and evoked potential amplitude in human somatosensory cortex. AB - The interpretation of task-induced functional imaging of the brain is critically dependent on understanding the relationship between observed blood flow responses and the underlying neuronal changes. However, the exact nature of this neurovascular coupling relationship remains unknown. In particular, it is unclear whether blood oxygen level-dependent functional magnetic resonance imaging (BOLD fMRI) responses principally reflect neuronal synaptic activity. In order to address this issue directly in humans, we measured the increase in somatosensory evoked potential amplitude and fMRI BOLD changes to increases in intensity of median nerve electrical stimulation in five healthy non-anaesthetized subjects. We found that mean N20-P22 amplitudes increased significantly with stimulus intensity in all subjects, as did fMRI BOLD percentage signal intensity change. Moreover, the intensity of the BOLD signal was found to correlate linearly with evoked potential amplitude in four of the five subjects studied. This suggests that the BOLD response correlates with synchronized synaptic activity, which is the major energy consuming process of the cortex. PMID- 11113330 TI - Increase in AMPA receptors in aged memory-impaired rats is not associated with increase in monoamine oxidase B levels. AB - Aged rats may be behaviorally classified as either cognitively impaired or unimpaired based upon their performance in the Morris water maze task. In aged Long-Evans rats, emergence of functional deficits has been related to the increase in the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor subtype in most hippocampal subfields, not observed in other brain structures. As AMPA receptors expressed in astrocytes may participate in the delayed and long-term glial response to injury, we investigated whether astrocytes participate in the increase of AMPA receptor observed in these aged rats. To this end, distribution of monoamine oxidase B, used as an astroglial marker, was characterized by quantitative autoradiography in the hippocampus and septum of young adults (six months) and aged (24-25 months) rats using [3H]lazabemide. Specific binding to brain sections of young, aged unimpaired, and aged impaired animals were calculated densitometrically. Compared to young animals, all hippocampal subfields in the aged unimpaired group showed a significant age-related increased labeling, which was not present in the aged impaired group. This contrasts with the increased glial transcription described in this last group. We propose that increase in AMPA receptors in the aged memory impaired animals may be related to an atypic astrocytic reactivity. PMID- 11113331 TI - Prosaptide exacerbates ischemia-induced behavioral deficits in vivo; an effect that does not involve mitogen-activated protein kinase activation. AB - Prosaposin is a 517 amino acid membrane component and secreted protein(5,7,9) that is proteolytically cleaved to generate the four small glycoproteins; saposins A, B, C and D.(9,13,19) Prosaposin's ability to promote neurite outgrowth(31) and to protect neurons from programmed cell death(28) in vitro, as well as to rescue neurons from ischemia and other damage in vivo(11,12,15,25) implied that prosaposin was neurotrophic/neuroprotectant.(1,7,24,31) The neurotrophic sequence of prosaposin was isolated to smaller peptide fragments termed prosaptides(15,31) within the amino terminal portion of saposin C.(1,6,8,10,17,20,21,28) The proposed use of synthetic prosaptides as peripherally administered neuroprotective and/or neurotrophic therapeutic agents has stemmed from their ability to cross the blood-brain barrier,(27) as well as their reported neurotrophic activity in vitro.(15,23,31) Few studies, however, have attempted to characterize these peptides, presumably due to their reported instability following peripheral administration.(27) With the recent design of a stable 11-mer retro-inverso prosaptide,(15,31) it has become feasible to investigate the pharmacological effects of a stable version of these peptides in the validated rabbit spinal cord ischemia model that has been used extensively in the development of therapeutics to treat ischemic stroke.(4,14,16,18) Our results show not only that prosaptide was not neurotrophic/neuroprotectant in vivo, but rather it worsened ischemia-induced behavioral deficits. PMID- 11113332 TI - GABA(A) receptors: immunocytochemical distribution of 13 subunits in the adult rat brain. AB - GABA(A) receptors are ligand-operated chloride channels assembled from five subunits in a heteropentameric manner. Using immunocytochemistry, we investigated the distribution of GABA(A) receptor subunits deriving from 13 different genes (alpha1-alpha6, beta1-beta3, gamma1-gamma3 and delta) in the adult rat brain. Subunit alpha1-, beta1-, beta2-, beta3- and gamma2-immunoreactivities were found throughout the brain, although differences in their distribution were observed. Subunit alpha2-, alpha3-, alpha4-, alpha5-, alpha6-, gamma1- and delta immunoreactivities were more confined to certain brain areas. Thus, alpha2 subunit-immunoreactivity was preferentially located in forebrain areas and the cerebellum. Subunit alpha6-immunoreactivity was only present in granule cells of the cerebellum and the cochlear nucleus, and subunit gamma1-immunoreactivity was preferentially located in the central and medial amygdaloid nuclei, in pallidal areas, the substantia nigra pars reticulata and the inferior olive. The alpha5 subunit-immunoreactivity was strongest in Ammon's horn, the olfactory bulb and hypothalamus. In contrast, alpha4-subunit-immunoreactivity was detected in the thalamus, dentate gyrus, olfactory tubercle and basal ganglia. Subunit alpha3 immunoreactivity was observed in the glomerular and external plexiform layers of the olfactory bulb, in the inner layers of the cerebral cortex, the reticular thalamic nucleus, the zonal and superficial layers of the superior colliculus, the amygdala and cranial nerve nuclei. Only faint subunit gamma3-immunoreactivity was detected in most areas; it was darkest in midbrain and pontine nuclei. Subunit delta-immunoreactivity was frequently co-distributed with alpha4 subunit immunoreactivity, e.g. in the thalamus, striatum, outer layers of the cortex and dentate molecular layer. Striking examples of complementary distribution of certain subunit-immunoreactivities were observed. Thus, subunit alpha2-, alpha4-, beta1-, beta3- and delta-immunoreactivities were considerably more concentrated in the neostriatum than in the pallidum and entopeduncular nucleus. In contrast, labeling for the alpha1-, beta2-, gamma1- and gamma2-subunits prevailed in the pallidum compared to the striatum. With the exception of the reticular thalamic nucleus, which was prominently stained for subunits alpha3, beta1, beta3 and gamma2, most thalamic nuclei were rich in alpha1-, alpha4-, beta2- and delta immunoreactivities. Whereas the dorsal lateral geniculate nucleus was strongly immunoreactive for subunits alpha4, beta2 and delta, the ventral lateral geniculate nucleus was predominantly labeled for subunits alpha2, alpha3, beta1, beta3 and gamma2; subunit alpha1- and alpha5-immunoreactivities were about equally distributed in both areas. In most hypothalamic areas, immunoreactivities for subunits alpha1, alpha2, beta1, beta2 and beta3 were observed. In the supraoptic nucleus, staining of conspicuous dendritic networks with subunit alpha1, alpha2, beta2, and gamma2 antibodies was contrasted by perykarya labeled for alpha5-, beta1- and delta-immunoreactivities. Among all brain regions, the median emminence was most heavily labeled for subunit beta2-immunoreactivity. In most pontine and cranial nerve nuclei and in the medulla, only subunit alpha1-, beta2- and gamma2-immunoreactivities were strong, whereas the inferior olive was significantly labeled only for subunits beta1, gamma1 and gamma2. In this study, a highly heterogeneous distribution of 13 different GABA(A) receptor subunit immunoreactivities was observed. This distribution and the apparently typical patterns of co-distribution of these GABA(A) receptor subunits support the assumption of multiple, differently assembled GABA(A) receptor subtypes and their heterogeneous distribution within the adult rat brain. PMID- 11113333 TI - Morphological and electrophysiological properties of atypically oriented layer 2 pyramidal cells of the juvenile rat neocortex. AB - We used whole-cell patch clamp recordings combined with intracellular dye-filling to examine the morphological and electrophysiological properties of atypically oriented pyramidal cells located at the layer 1/2 border of the juvenile rat neocortex. Orientation of the apical dendrite varied from oblique (>20 degrees from vertical) to truly horizontal (90 degrees from vertical). The length of the apical dendrite ranged from 150 to 400 microm. The total horizontal domain of the dendritic tree (including basal dendrites) of the longest horizontal pyramids exceeded 500 microm, but we also found short horizontal cells with horizontal dendritic domains of less than 300 microm. In addition, atypically oriented pyramids had long horizontal axon collaterals in layer 1/2. Electrophysiologically, atypically oriented pyramidal cells had intrinsic membrane properties similar to regularly oriented pyramids that have been described in the superficial layers at this age in the rat. Cells that fired repetitively were all regular spiking. In addition, we identified a subgroup of neurons (20%) in this sample, which were unable to fire more than a few spikes at the beginning of the current pulse. We suggest that the unique orientation and size of their dendritic trees and the length and arrangement of their local axons collaterals make atypically oriented pyramids in layer 2 ideally suited to perform horizontal integration of synaptic inputs in the neocortex. PMID- 11113334 TI - Effect of partial sensory deprivation on monoaminergic neuromodulators in striate cortex of adult cat. AB - The role of monoaminergic neuromodulators in the reorganization of cortical topography following limited sensory deprivation in the adult cat was investigated. The total concentrations of dopamine, noradrenaline, serotonin and their major metabolites were measured in the visual cortex of both normal control and experimental animals using microbore high-performance liquid chromatography coupled with electrochemical detection. The experimental animals were subjected to a binocular retinal lesion corresponding to the central 10 degrees of vision and killed two weeks post-lesion. The sensory deprivation was confirmed in area 17 by measuring immediate-early gene zif-268 messenger RNA expression. Following the retinal lesion, the total concentrations of noradrenaline and dopamine were significantly higher in the non-deprived cortex of retinal lesion cats than in the deprived cortex of retinal lesion cats and the cortex of normal animals. This pattern follows the release of the excitatory neurotransmitter glutamate under the same conditions. Serotonin levels were significantly lower in the deprived cortex, and its metabolite 5-hydroxyindole-3-acetic acid was significantly higher in the non-deprived cortex than in deprived cortex and normal cortex. From these results, we suggest that the modulation of noradrenaline, dopamine and serotonin is regulated by visual afferent activity. PMID- 11113336 TI - Susceptibility of transgenic mice expressing human apolipoprotein E to closed head injury: the allele E3 is neuroprotective whereas E4 increases fatalities. AB - Apolipoprotein E, the major brain lipid-binding protein, is expressed in humans as three common isoforms (E2, E3 and E4). Previous studies revealed that the allele apolipoprotein E4 is a major genetic risk factor of Alzheimer's disease and that traumatic brain injury is associated with increased risk for developing this disease. Furthermore, it has been suggested that the effects of traumatic head injury and apolipoprotein E4 in Alzheimer's disease are synergistic. To test the hypothesis that the apolipoprotein E genotype affects susceptibility to brain injury, we subjected transgenic mice, expressing either human apolipoprotein E3 or human apolipoprotein E4 on a null mouse apolipoprotein E background and apolipoprotein E-deficient knockouts, to closed head injury and compared mortality, neurological recovery and the extent of brain damage of the survivors. More than 50% of the transgenic mice expressing human apolipoprotein E4 died following closed head injury, whereas only half as many of the transgenic mice expressing human apolipoprotein E3, and of the control and apolipoprotein E deficient mice died during this period (P<0.02). A neurological severity score used for clinical assessment of the surviving mice up to 11 days after closed head injury revealed that the four mouse groups displayed similar severity of damage at 1h following injury. At three and 11 days post-injury, however, the neurological severity scores of the transgenic mice expressing human apolipoprotein E3 were significantly lower than those of the other three groups whose scores were similar, indicating better recovery of the transgenic mice expressing human apolipoprotein E3. Histopathological examination of the mice performed 11 days post-injury revealed, consistent with the above neurological results, that the size of the damaged brain area of the transgenic mice expressing human apolipoprotein E3 was smaller than that of the other head injured groups. These findings show that transgenic mice expressing human apolipoprotein E4 are more susceptible than those expressing apolipoprotein E3 to closed head injury. We suggest that this effect is due to both a protective effect of apolipoprotein E3 and an apolipoprotein E4-related pathological function. PMID- 11113335 TI - Regional cerebral cortical activation in monoamine oxidase A-deficient mice: differential effects of chronic versus acute elevations in serotonin and norepinephrine. AB - Mice deficient in monoamine oxidase A have previously been shown to demonstrate a chronic elevation of serotonin and norepinephrine in the brain. Using the autoradiographic [14C]iodo-antipyrine method, we examined cerebral cortical blood flow in conscious, restrained four- to five-month-old knock-out and wild-type animals following the intraperitoneal administration of either saline or D fenfluramine. Knock-out animals administered saline, compared to their wild-type counterparts, demonstrated a significantly higher regional cortical blood flow in somatosensory and barrel field neocortex, an area which previous histological studies have shown to be characterized by abnormal serotonergic projection fibers and absent barrel formation. Regional cortical blood flow was significantly lower in knock-out than in wild-type mice in the entorhinal and midline motor cortex, with non-significant decreases noted in the olfactory, piriform and retrosplenial cortices and the amygdala. We compared the above findings to those obtained in response to D-fenfluramine which, in conjunction with its metabolite D norfenfluramine, results in acute elevations of brain levels of serotonin and norepinephrine. Administration of D-fenfluramine (21. 2mg/kg) resulted in changes in regional cortical perfusion in most brain regions of both knock-out and wild type mice that were opposite to the genotypic differences seen in perfusion in response to saline. Fenfluramine significantly increased regional cortical blood flow in the allocortex (olfactory, piriform, entorhinal) and the amygdala, and significantly decreased regional cortical blood flow in the somatosensory, barrel field, midline motor and retrosplenial cortices. Changes in regional perfusion in response to fenfluramine were topographically equivalent in knock-out and wild type mice, although in knock-out mice such changes were of greater magnitude. Our study suggests that the effects on regional cortical blood flow of a lifelong absence of monoamine oxidase A, and the consequent chronic increase in serotonin and norepinephrine, differ from those attributable to acute increases in these neurotransmitters following fenfluramine administration. Such a differential response may reflect neurodevelopmental abnormalities and/or effects of a chronic physiological adaptation on the regulation of cortical activation. PMID- 11113337 TI - Microtubule-associated protein-2 located in growth regions of rat hippocampal neurons is highly phosphorylated at its proline-rich region. AB - Neuronal morphogenesis is regulated, among other factors, by microtubule associated proteins (MAPs). A family of these proteins, MAP2, which is very abundant in the mammalian nervous system, has been associated with the formation of neurites at early developmental stages and with the dendritic scaffold upon maturation. The function of MAP2 is regulated by its phosphorylation state. One of the phosphorylation sites that has been described is located in the proline rich region of the protein. It comprises of the residues 1616-1626 and is specifically recognized by the antibody 305. However, little is known about the functional consequences of its modification in vivo. To gain insight into this, we have analysed the expression levels and intracellular distribution of MAP2 phosphorylated at this site (MAP2-P), in primary cultures of rat hippocampal neurons at different developmental stages. Western blot analysis of hippocampal neuron protein extracts revealed that the ratio of MAP2-P:MAP2 was 4:1 at early developmental stages and became 1:4 at later developmental stages, suggesting a role of such phosphorylated forms of the protein in neuritogenesis. Consistent with this view, immunofluorescence microscopy analysis showed that the ratio MAP2 P:MAP2 was 2 in the neurite growth cones, sites where net elongation takes place. A higher presence of phosphorylated MAP2 was observed in growth regions with higher levels of microfilaments, which may be related with the growth region stability. Indeed, when growth-cone collapse was induced in hippocampal neurons after cytochalasin D treatment, which depolymerizes microfilaments, the ratio MAP2-P:MAP2 in these growing regions decreased down to 1. Finally, acceleration of neuronal maturation induced by the activation of glutamate-receptors triggered a dramatic decrease in the phosphorylation of MAP2 at the site recognized by antibody 305. From these results we suggest that the phosphorylation of MAP2 at its proline-rich region is an important event during neuritogenesis. PMID- 11113338 TI - Progestin receptors mediate progesterone suppression of epileptiform activity in tetanized hippocampal slices in vitro. AB - Clinical and laboratory studies suggest that progesterone reduces epileptic seizure activity. The mechanisms underlying this effect are not known. The present study determined the effects of progesterone on extracellular evoked responses recorded in the CA1 field of hippocampal slices, as well as epileptiform responses recorded from tetanized slices. Slices were prepared from ovariectomized rats, with or without estrogen replacement. Hippocampal slices were superfused in vitro with one of the following treatments: progesterone with or without RU486 (a progesterone receptor antagonist); allopregnanolone (a progesterone metabolite that potentiates GABA action at GABA(A) receptors); RU5020 (a high-affinity progesterone receptor agonist); or cholesterol (control). In non-tetanized slices, a twofold increase in the excitatory postsynaptic field potential and population spike amplitude occurred during both cholesterol and progesterone superfusion. In contrast, under the same conditions, exposure to allopreganolone caused a 25% reduction in both field potential and population spike amplitude of evoked responses within 30min of treatment. In tetanized slices, progesterone and RU5020, but not allopregnanolone or cholesterol, caused significant reductions in the field potential and population spike amplitude of evoked responses. Progesterone and RU5020 also significantly reduced the duration of tetanic stimulus-induced afterdischarges and the frequency of spontaneous interictal discharges. The effects of allopregnanolone were restricted to a reduction in the primary afterdischarge duration. Estrogen replacement slightly attenuated progesterone's suppression of spontaneous discharges and depression of evoked responses. All responses to progesterone were blocked by prior or concurrent exposure to RU486. These data indicate that allopregnanolone suppresses evoked potentials in non-tetanized hippocampal slices, consistent with previous reports that this neurosteroid has marked anxiolytic and anticonvulsant effects. After tetanization, however, progesterone receptor-mediated responses become quantitatively more important as a mechanism for suppressing hippocampal electrical activity. PMID- 11113339 TI - A transient increase in temperature induces persistent potentiation of synaptic transmission in rat hippocampal slices. AB - Previous studies have shown that increasing the temperature of rat hippocampal brain slices from 32.5 to 38.5 degrees C initiates a profound, adenosine-mediated decrease in excitatory synaptic transmission in the CA1 region. Here we found that upon lowering the temperature back to 32.5 degrees C, the amplitude of the field excitatory postsynaptic potential often recovers to a level that is significantly potentiated with respect to the initial baseline. This potentiation is rapid in onset (< 5min following return to 32.5 degrees C) and long lasting (>60min following the termination of the increase in temperature). Similar effects could not be induced by superfusion with adenosine alone, and adenosine receptor antagonists did not block the potentiation. Therefore, although an adenosine-mediated decrease in excitatory synaptic transmission occurs during the temperature increase, it is unrelated to the potentiation. Likewise, N-methyl-D aspartate receptor activation is not required, as N-methyl-D-aspartate receptor antagonists do not influence this form of potentiation. In summary, we propose that transiently increasing brain slice temperature represents a novel way to induce synaptic plasticity in the hippocampus, and may provide a paradigm to elucidate additional cellular mechanisms involved in functional plasticity. PMID- 11113340 TI - Long-term induction of Fos-related antigen-2 after methamphetamine-, methylenedioxymethamphetamine-, 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine- and trimethyltin-induced brain injury. AB - A long-term induction of Fos-related antigens has been shown in neurons after brain injury, suggesting that Fos-related antigens are involved in enhancing the transcription of genes related to the process of regeneration and repair. In the present study, we report that levels of Fos-related antigen-2 are elevated in several models of chemically induced brain injury. Trimethyltin, which causes degeneration of neurons primarily in the hippocampus and other limbic regions, results in a five-fold induction of Fos-related antigen-2 immunoreactivity in neurons in the pyramidal and dentate layers of the hippocampus starting at seven days post-treatment and persisting for 60days. Methamphetamine and methylenedioxymethamphetamine, agents which cause degeneration of dopaminergic nerve terminals in the striatum of the mouse, cause an increase in Fos-related antigen-2 immunoreactivity which begins at three days post-treatment and returns to basal levels by days 5 and 15, respectively. Treatment with 1-methyl-4-phenyl 1,2,3, 6-tetrahydropyridine elevated levels of Fos-related antigen-2 in the mouse striatum at three days post-treatment. This abbreviated time-course of Fos related antigen-2 induction is consistent with less severe insult (terminal damage) relative to trimethyltin (cell death), but induction occurs during the period of regeneration and repair in both models. Dexfenfluramine, a non neurotoxic amphetamine, does not induce Fos-related antigen-2 expression. Decreasing core temperature of the mouse, which blocks amphetamine-induced neurotoxicity, also blocks Fos-related antigen-2 induction. In summary, Fos related antigen-2 is induced in models of both cell death and terminal degeneration, suggesting that this transcription factor may serve as a universal signal transduction molecule involved in the regulation of genes related to regeneration and repair in the CNS. PMID- 11113341 TI - Modulation of latent inhibition in the rat by altered dopamine transmission in the nucleus accumbens at the time of conditioning. AB - Latent inhibition describes a process by which pre-exposure of a stimulus without consequence retards the learning of subsequent conditioned associations with that stimulus. It is well established that latent inhibition in rats is impaired by increased dopamine function and potentiated by reduced dopamine function. Previous evidence has suggested that these effects are modulated via the meso accumbens dopamine projections. We have now undertaken three experiments to examine this issue directly, especially in the light of one study in which latent inhibition was reported to be unaffected by direct injection of amphetamine into the accumbens. Latent inhibition was studied using the effect of pre-exposure of a tone stimulus on the subsequent formation of a conditioned emotional response to the tone. 6-Hydroxydopamine-induced lesions of dopamine terminals in the nucleus accumbens resulted in potentiation of latent inhibition. Bilateral local injections of the dopamine antagonist haloperidol into the nucleus accumbens (0.5 microg/side) before conditioning also potentiated latent inhibition. Moreover, such injections were able to reverse the disruptive effect of systemic amphetamine (1mg/kg, i.p.) on latent inhibition. Bilateral local injection of amphetamine (5 microg/side) into the nucleus accumbens before conditioning was able to disrupt latent inhibition, provided that it was preceded by a systemic injection of amphetamine (1mg/kg) 24h earlier.We conclude that the attenuation of latent inhibition by increased dopamine function in the nucleus accumbens is brought about by impulse-dependent release of the neurotransmitter occurring at the time of conditioning. The previously reported failure to disrupt latent inhibition with intra-accumbens amphetamine is probably due to impulse independent release of dopamine. The implications of these conclusions for theories linking disrupted latent inhibition to the attentional deficits in schizophrenia, and to the dopamine theory of this disorder, are discussed. PMID- 11113342 TI - Effects of gonadal hormone replacement on measures of basal forebrain cholinergic function. AB - The effects of different hormone replacement regimens on basal forebrain cholinergic function were examined by measuring changes in choline acetyltransferase activity and high affinity choline uptake in adult, ovariectomized, rats. Increases in choline acetyltransferase activity were detected in the frontal cortex (20. 1%) and olfactory bulbs (30.4%) following two weeks, but not four weeks, of repeated treatment with estrogen plus progesterone. Increases in high affinity choline uptake were detected in the frontal cortex (39.5-55.1%), hippocampus (34.9-48.9%), and olfactory bulbs (29.9%) after two weeks, but not four weeks, of either continuous estrogen administration, repeated progesterone administration, or repeated treatment with estrogen plus progesterone. Repeated administration of estradiol (2-25 microg/250 g body weight) for two or four weeks, and continuous estrogen administration for four weeks and six months, produced no significant changes in choline acetyltransferase activity or high affinity choline uptake in the hippocampus, frontal cortex or olfactory bulbs. Continuous estrogen administration for 13 months produced a significant decrease in high affinity choline uptake across all regions with the largest effect (-28.1%) detected in the hippocampus. The findings demonstrate that short-term treatment with estrogen and/or progesterone can significantly enhance cholinergic function within specific targets of the basal forebrain cholinergic projections. Most important is the fact that the effects varied considerably according to the manner and regimen of hormone replacement and did not persist with prolonged treatment. These findings could have important implications for the effective use of hormone replacement strategies in the prevention and treatment of Alzheimer's disease and age-related cognitive decline in women. PMID- 11113343 TI - Quantitative histological analysis of amyloid deposition in Alzheimer's double transgenic mouse brain. AB - The development of transgenic mice has created new opportunities for the generation of animal models of human neurodegenerative diseases where previously there was no animal counterpart. The first successful transgenic mouse model of Alzheimer's disease expressed increased levels of mutant human amyloid precursor protein, exhibiting neuritic-type amyloid deposits and behavioral deficits at six to nine months of age. More recently, it was shown that transgenic mice expressing both mutant human amyloid precursor protein and presenilin 1 exhibit neuritic-type amyloid deposits and behavioral deficits in as little as 12 weeks. This accelerated Alzheimer phenotype greatly reduces the time necessary to conduct preclinical drug trials, as well as animal housing costs. The purpose of this study was to quantify the deposition of amyloid in five regions of the cortex and two regions of the hippocampus of transgenic mice expressing amyloid precursor protein (K670N, M671L) and presenilin 1 (M146L) mutations at various ages, using quantitative methods of confocal laser scanning microscopy and image analysis. Amyloid burden, expressed as the percentage area occupied by thioflavin S-positive amyloid deposits, increased an average of 179-fold from 12 to 54 weeks of age (0.02+/-0.01% to 3.57+/-0.29%, mean+/-S.E.M., respectively) in five regions of the cortex and two of the hippocampus. This was a function of increases in both deposit number and size. This transgenic mouse provides an ideal animal model for evaluating the efficacy of potential therapeutic agents aimed at reducing amyloid deposition, such as inhibitors of amyloid fibril formation or secretase inhibitors. PMID- 11113344 TI - Conditionally immortal neuroepithelial stem cell grafts reverse age-associated memory impairments in rats. AB - In order to investigate the effects of stem cell grafts on water maze deficits in aged (22-month-old) rats, three groups of aged rats, assigned by pre-training latency scores to unimpaired, impaired control and impaired grafted groups, were compared with young (five-month-old) controls, six to eight weeks after implantation of cells from the conditionally immortal Maudsley hippocampal stem cell line, clone 36 (MHP36 stem cell line), in the cortex, striatum and hippocampus. Grafted rats were substantially superior to their matched impaired aged controls, and learned to find the platform as rapidly as unimpaired aged rats, although young controls were more efficient than all aged groups in several measures of spatial search during training. On the probe trial, however, aged rats with grafts showed significantly better recall of the precise position of the platform than any other group, including young controls, possibly indicating some perseveration. A further comparison found that groups of unimpaired and moderately impaired aged rats showed far less improvement from water maze pre training to acquisition phases than young controls, indicative of progressive deficits over time. Histological investigation showed that beta-galactosidase positive MHP36 cells migrated widely from the implantation sites to infiltrate the striatal matrix, all hippocampal fields and areas of the cortex. Grafted cells showed both astrocytic and neuronal morphologies, with cells of pyramidal and granular appearance in appropriate hippocampal strata.Taken together, these results indicate that neuroepithelial stem cell grafts extensively colonize the aged rat brain and substantially reverse progressive cognitive decline associated with ageing. PMID- 11113345 TI - Diffusion and action of intracerebroventricularly injected interleukin-1 in the CNS. AB - Interleukin-1beta acts on the CNS to induce fever, neuroendocrine activation and behavioural depression. We have previously demonstrated that interleukin-1beta is synthesized in glial cells and macrophages of circumventricular organs and choroid plexus after intraperitoneal administration of bacterial lipopolysaccharide. Whether, and how, interleukin-1beta produced in glial cells affects neuronal functioning is unknown. Diffusion throughout the extracellular space is an important pathway by which factors produced by glial cells act on distant cells, a phenomenon coined "volume transmission". The present study assessed diffusion of recombinant rat interleukin-1beta, recombinant human interleukin-1 receptor antagonist and 10mol. wt dexran in the rat CNS after intracerebroventricular administration to model interleukin-1beta release from choroid plexus. Immunocytochemistry with specific antibodies directed against interleukin-1beta and interleukin-1 receptor antagonist revealed that these molecules rapidly penetrated into periventricular tissue and spread along white matter fibre bundles and blood vessels in the caudoputamen, hypothalamus and amygdala. The transcription factor nuclear factor kappa B and the immediate-early gene product Fos were detected immunocytochemically to reveal interleukin-1beta action. Intracerebroventricular infusion of interleukin-1beta induced nuclear factor kappa B translocation in choroid plexus, ependymal cells, basolateral amygdala, cerebral vasculature and meninges. Fos immunoreactivity was found in the supraoptic and paraventricular hypothalamus and central amygdala. We propose that intracerebroventricular injected interleukin-1beta can enter the brain parenchyma and act as a "volume transmission" signal in, for example, the basolateral amygdala where it might activate a neuronal projection to the central amygdala. PMID- 11113346 TI - Differential synaptic distribution of AMPA receptor subunits in the ventral posterior and reticular thalamic nuclei of the rat. AB - Although the mechanisms by which the cerebral cortex controls its ascending input are still poorly understood, it is known that cortical control at the thalamic level is via direct glutamatergic projections to relay nuclei and to the reticular nucleus. Here we confirm previous light microscopic reports of a high expression of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit, GluR4, in reticular and ventral posterior thalamic nuclei of the rat, and moderate staining using an antibody recognizing both GluR2 and GluR3. In contrast only low levels of staining for GluR2, and barely detectable levels of GluR1 immunoreactivity were observed. After injections of biotinylated dextran, electron microscopy revealed that anterogradely-labeled cortical synapses in both thalamic nuclei were small with fewer mitochondria and more densely-packed vesicles than terminals likely to arise from intrinsic and ascending pathways. We performed post-embedding immunogold to provide quantitative data on the density of AMPA receptor subunits at morphologically defined groups of synapses. We found that corticothalamic synapses in the reticular thalamic nucleus contain twice as much GluR2/3, and at least three times more GluR4 protein than do intrathalamic synapses. In the ventral posterior nucleus, corticothalamic synapses contain similar amounts of GluR2/3, but four times more GluR4 than do those from ascending afferents. Corticothalamic synapses in reticular nucleus contain slightly more GluR2/3, and three times more GluR4, than those in ventral posterior nucleus. We conclude that enrichment of GluR4 at morphologically-defined cortical synapses is a feature common to both thalamic nuclei, and those in the reticular nucleus express higher levels of AMPA receptors. The rapid kinetics of GluR4-rich AMPA receptors we suggest indicate that cortical descending control may be more temporally precise than previously recognized. PMID- 11113347 TI - Fos expression in the rostral thalamic nuclei and associated cortical regions in response to different spatial memory tests. AB - Using the quantification of the Fos protein as an indicator of neuronal activation, we studied the involvement of the rostral thalamic nuclei and associated structures in different spatial memory tasks in two experiments. In both experiments, tasks were matched for sensorimotor factors but differed in their spatial and mnemonic demands. In Experiment 1, matched groups of rats either ran in a standard eight-arm radial maze or ran up and down just one arm of the maze while the number of runs and rewards were matched across pairs of rats. In Experiment 2, both groups were trained on the eight-arm radial maze but in different rooms. On the test day, one group was moved so that both groups now performed the same radial-maze task in the same room but for one group the extramaze cues were novel. There were significant increases in Fos in all three of the anterior thalamic nuclei (anterodorsal, anteroventral and anteromedial) as well as the adjacent nucleus reuniens and rostral reticular thalamic nucleus, in both the eight-arm versus one-arm condition (Experiment 1) and the novel room versus familiar room condition (Experiment 2). There were no significant differences in the mediodorsal thalamic nucleus in either experiment. The more spatially demanding task in each experiment also resulted in increased Fos expression in the subicular complex (postsubiculum, presubiculum and parasubiculum), as well as in the prelimbic cortex. Performing the standard radial-arm maze task also produced significant Fos increases in both rostral and caudal levels of the retrosplenial cortex when compared to rats running up and down a single arm in the same maze (Experiment 1); performing the task in a novel room did not, however, result in any further Fos increases in this region (Experiment 2). The specificity of the changes in levels of Fos was shown by a lack of any consistent difference in levels in six control sites.The present results reveal a group of anatomically related structures that work together in the intact rat brain during tasks that tax allocentric spatial working memory. PMID- 11113348 TI - Unilateral lesion of the nigrostriatal pathway induces a transient decrease of firing rate with no change in the firing pattern of neurons of the parafascicular nucleus in the rat. AB - Electrophysiological recordings of thalamic parafascicular nucleus neurons were done in normal rats and in three groups of rats at different time intervals after injection of 6-hydroxydopamine into the pars compacta of substantia nigra. In normal rats, parafascicular neurons exhibited low firing rates (3.88+/-0.80 spikes/s). Concerning the pattern, 59% of the units discharged irregularly and 41% exhibited bursty pattern. In rats with 6-hydroxydopamine lesions, the firing rate decreased significantly during the first week post-lesion (1.15+/-0.36 spikes/s, P<0.01). During the second week, the firing rate was slightly, but not significantly, lower (2.59+/-0.41 spikes/s, P>0.05) than that of normal rats to return to the basal level three weeks post-lesion (3. 66+/-0.41 spikes/s, P>0.05). In these three groups of 6-hydroxydopamine-lesioned rats, the firing pattern showed no change when compared to control animals. These results show that the lesion of nigral dopaminergic neurons induced a transient decrease of the firing rate of parafascicular neurons with no change in the firing pattern demonstrating the absence of a stable influence of the dopaminergic system on the spontaneous activity of parafascicular neurons. PMID- 11113349 TI - Expression of peptides and other neurochemical markers in hypothalamus and olfactory bulb of mice devoid of all known thyroid hormone receptors. AB - We have investigated with histochemical techniques the expression of peptides and other neurochemical markers in the hypothalamus and olfactory bulb of male mice, in which the genes encoding the alpha and beta thyroid hormone receptors (TRalpha1, TRbeta1 and TRbeta2) have been deleted. Thyrotropin-releasing hormone messenger RNA levels were increased in the hypothalamic paraventricular nucleus and in the medullary raphe nuclei of mutant mice lacking the thyroid hormone receptors alpha1 and beta (alpha1(-/-)beta(-/-)), as compared to wild-type mice. In contrast, galanin messenger RNA levels were lower in the hypothalamic paraventricular nucleus of mutant animals, as was galanin-like immunoreactivity in the internal layer of the median eminence. Substance P messenger RNA levels were unchanged in the medullary raphe nuclei. Thyrotropin-releasing hormone receptor messenger RNA levels were increased in motoneurons, unchanged in the subiculum, and lower in the amygdala of mutant animals. Galanin messenger RNA levels were unchanged in the hypothalamic dorsomedial and arcuate nuclei of the thyroid hormone receptor alpha1(-/-)beta(-/-) mice, as was the immunocytochemistry for oxytocin and for vasopressin in the hypothalamic paraventricular nucleus. A reduction in tyrosine hydroxylase messenger RNA levels was found in the arcuate nucleus of mutant mice. In the olfactory bulb, immunohistochemistry for calbindin and for tyrosine hydroxylase revealed a reduction in the intensity of labeling of nerve processes in the glomerular layer of thyroid hormone receptor alpha1(-/-)beta(-/-) mice. The tyrosine hydroxylase messenger RNA levels were also slightly reduced. In contrast, the levels of galanin and neuropeptide Y messenger RNA in this region were unchanged in thyroid hormone receptor alpha1(-/-)beta(-/-) mice as compared to wild-type mice. Together these studies reveal many regional and neurochemically selective alterations in neuronal phenotype of mice devoid of all known thyroid hormone receptors. PMID- 11113350 TI - Direct pathways to the supraoptic nucleus from the brainstem and the main olfactory bulb are activated at parturition in the rat. AB - Sensory input from female reproductive structures is paramount for the co ordination of neuroendocrine changes at parturition. Using a retrograde tracer (fluorescent latex microspheres) in combination with Fos (as an indicator of neuronal activation) and tyrosine hydroxylase (to identify catecholaminergic neurons) immunocytochemistry we identified cells within the brainstem and main olfactory bulb that project to the supraoptic nucleus, and which become significantly activated at parturition (compared to virgin rats and rats on the day of expected parturition). Within the A2/C2 region in the nucleus tractus solitarii, 60% of the projecting activated cells were catecholaminergic, as were 59% of such cells in the A1/C1 region of the ventrolateral medulla. This suggests that oxytocin and vasopressin neurons within the supraoptic nucleus are stimulated at parturition via afferent inputs from the brainstem, but the input is not exclusively noradrenergic. Within the mitral layer of the main olfactory bulb, cells that projected to the supraoptic nucleus were significantly activated, suggesting that the olfactory system may regulate supraoptic nucleus cell firing at parturition. The preoptic area, organum vasculosum of the lamina terminalis and medial amygdala contained cells that projected to the supraoptic nucleus but these projections were not significantly activated at parturition, although non-projecting cells in these regions were. On the expected day of parturition, but before birth, projections from the organum vasculosum of the lamina terminalis to the supraoptic nucleus became significantly activated. These findings provide evidence of direct afferent pathways to the supraoptic nucleus from the brain stem and olfactory bulbs that are activated at parturition. PMID- 11113351 TI - Differential responses of phosphorylated mitogen-activated protein kinase and phosphorylated cyclic-AMP response element-binding protein immunoreactivity in the rat brain to sub-convulsive pentylenetetrazol. AB - The possible advantage of using multiple phospho-specific antibodies to study changes in brain activity was assessed. For this purpose, rats were injected intraperitoneally with either a control treatment or 15 mg/kg pentylenetetrazol. The sub-convulsive dose of pentylenetetrazol did not induce marked behavioural effects. Ten minutes after treatment, the rats were perfused and the brains were dissected. Adjacent brain sections were immunohistochemically stained for phosphorylated cyclic-AMP response element-binding protein and phosphorylated mitogen-activated protein kinase. Opposite effects of pentylenetetrazol treatment were observed on the immunoreactivity of these two antibodies within the hypothalamic paraventricular nucleus, the supraoptic nucleus and the arcuate nucleus. In these regions, pentylenetetrazol treatment increased phosphorylated mitogen-activated protein kinase immunoreactivity, but decreased phosphorylated cyclic-AMP response element-binding protein immunoreactivity. These findings show that changes in the activity of a brain nucleus can be accompanied by differential changes in the activity of two signal transduction pathways, which can be detected immunohistochemically. Therefore, the use of multiple phospho specific antibodies may enhance our potential to monitor changes in brain activity. PMID- 11113352 TI - Decreased messenger RNA expression of key markers of the nigrostriatal dopaminergic system following vitamin E deficiency in the rat. AB - We have evaluated the effect of a vitamin E-deficient diet on the rat nigrostriatal dopaminergic system. After 15 days of deficient diet, the amount and activity of striatal and nigral tyrosine hydroxylase increased, which contrasted with a decreased messenger RNA expression for tyrosine hydroxylase and the dopamine transporter in the ventral mesencephalon. When we prolonged the deficiency of vitamin E for 30 days, dopamine levels did not differ in both areas. In contrast, messenger RNA levels for tyrosine hydroxylase and the dopamine transporter were markedly reduced in 30-day deficient rats. In addition, the number of oxidatively modified proteins significantly increased in the striatal and nigral areas studied. Overall, we propose that these changes suggest an important role of vitamin E in maintaining the normal equilibrium of the dopaminergic nigrostriatal system. PMID- 11113353 TI - Synaptic localization of ionotropic glutamate receptors in the rat substantia nigra. AB - Glutamatergic neurotransmission in the substantia nigra pars compacta and pars reticulata is mediated through N-methyl-D-aspartate and alpha-amino-3-hydroxy-5 methyl-4-isoxaline propionic acid/kainate (AMPA) type receptors as well as other glutamate receptors and is critical for basal ganglia functioning. A major glutamatergic input to the substantia nigra originates in the subthalamic nucleus, and the long-lasting stimulation of the dopaminergic cells of the substantia nigra pars compacta by the subthalamic neurons has been implicated in the pathophysiology of Parkinson's disease. The objectives of the present study were to determine the subcellular and subsynaptic localization of subunits of the N-methyl-D-aspartate and AMPA receptors in the substantia nigra, and also to determine whether co-localization of N-methyl-D-aspartate and AMPA receptor subunits occur at individual synapses. To achieve this, pre-embedding and post embedding immunocytochemistry was applied to sections of substantia nigra using antibodies that recognize the NR1 and NR2A/B subunits of the N-methyl-D-aspartate receptor, and GluR2/3 subunits of the AMPA receptor. In both regions of the substantia nigra, immunolabelling for each of the subunits was observed in numerous perikarya and proximal dendrites. At the subcellular level, silver intensified immunogold particles localizing N-methyl-D-aspartate and AMPA receptor subunits were most commonly present within dendrites where they were associated with a variety of intracellular organelles and with the internal surface of the plasma membrane. Post-embedding immunogold labelling revealed immunoparticles labelling for NR1, NR2A/B and GluR2/3 to be enriched at asymmetric synaptic specializations, although a large proportion of asymmetric synapses were immunonegative. Double immunolabelling revealed, in addition to single-labelled synapses, the co-localization of subunits of the N-methyl-D aspartate receptor and subunits of the AMPA receptor at individual asymmetric synapses. Similarly, double immunolabelling also revealed the co-localization of the NRl and NR2A/B subunits of the N-methyl-D-aspartate receptor at individual asymmetric synapses. Labelling for NR1 and GluR2/3 was, on average, relatively evenly distributed across the width of the synapse with a gradual reduction towards the periphery when analysed in single sections. In summary, the present results demonstrate that AMPA and N-methyl-D-aspartate receptors are selectively localized at a subpopulation of asymmetric synapses in the substantia nigra pars compacta and reticulata and that the two receptor types, at least partially co localize at individual synapses. It is concluded that glutamatergic transmission in the substantia nigra pars compacta and pars reticulata occurs primarily at asymmetric synapses and, at least in part, is mediated by both N-methyl-D aspartate and AMPA receptors. PMID- 11113354 TI - Micturition evoked by glutamate microinjection in the ventrolateral periaqueductal gray is mediated through Barrington's nucleus in the rat. AB - Neural tracing experiments have demonstrated a direct spinal projection to Barrington's nucleus and a possible indirect pathway to Barrington's nucleus via the periaqueductal gray. We sought to identify the role of the periaqueductal gray matter in micturition in urethane-anesthetized rats. Blockade of micturition by focal injection of cobalt chloride was used to identify sites critical to micturition. These sites were located near the ventral margin of the caudal ventrolateral periaqueductal gray and in Barrington's nucleus. L-Glutamate injections into caudal regions of the periaqueductal gray evoked bladder contraction with coordinated sphincter activation. Additional L-glutamate sites with a similar pattern of response and sites where sphincter activation was produced without bladder contraction were found more rostrally and dorsally in the periaqueductal gray. Activation of bladder contractions by L-glutamate injection in the ventrolateral periaqueductal gray was blocked by prior injection of cobalt chloride into Barrington's nucleus. From these data we propose that ventrolateral periaqueductal gray is functionally important to micturition in the urethane-anesthetized rat. Further, we have shown that a periaqueductal gray to Barrington's nucleus pathway is functionally relevant to central mediation of bladder contraction. PMID- 11113355 TI - Pharmacological inactivation of the vesicular monoamine transporter can enhance 1 methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced neurodegeneration of midbrain dopaminergic neurons, but not locus coeruleus noradrenergic neurons. AB - The vesicular monoamine transporter in the brain can sequester the neurotoxin 1 methyl-4-phenylpyridinium into synaptic vesicles and protect catecholamine containing neurons from degeneration. Mouse nigrostriatal dopaminergic neurons, and to a lesser extent locus coeruleus noradrenergic neurons, are vulnerable to toxicity produced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. The present study sought to determine whether pharmacological inactivation of the vesicular monoamine transporter in the brain would enhance the degeneration of substantia nigra dopaminergic neurons and locus coeruleus noradrenergic neurons in 1-methyl 4-phenyl-1,2,3, 6-tetrahydropyridine-treated animals. Mice were treated subacutely with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine alone, or in combination with vesicular monoamine transporter inhibitors (tetrabenazine or Ro4 1284), and 10-24 days later striatal dopamine and cortical norepinephrine levels were measured using chromatographic methods. In the same animals, substantia nigra and locus coeruleus catecholaminergic neurons were counted using tyrosine hydroxylase immunohistochemical staining with computer imaging techniques. Mice in which pharmacological blockage of the vesicular monoamine transporter enhanced the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity in the depletion of striatal dopamine concentrations also exhibited enhanced degeneration of substantia nigra dopaminergic neurons. In the same animals, however, vesicular monoamine transporter blockade did not enhance the effects of 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine in the locus coeruleus noradrenergic system. These data are consistent with the hypothesis that the vesicular monoamine transporter can protect catecholamine-containing neurons from 1-methyl-4-phenylpyridinium-induced degeneration by sequestration of the toxin within brain vesicular monoamine transporter-containing synaptic vesicles. Since the amount of vesicular monoamine transporter in locus coeruleus neurons is more than in substantia nigra neurons, and because 1-methyl-4-phenylpyridinium is sequestered within locus coeruleus neurons to a far greater extent than within substantia nigra neurons, it may be that a greater amount of vesicular monoamine transporter inhibition is required for 1-methyl-4-phenylpyridinium to be toxic to locus coeruleus neurons than to substantia nigra dopaminergic neurons. PMID- 11113356 TI - Role of locus coeruleus in foot shock-evoked Fos expression in rat brain. AB - The robust activation of locus coeruleus neurons in response to a variety of stressors, in conjunction with the widespread outputs of the locus coeruleus, suggest that the locus coeruleus may be important in mediating responses to stress. Previous studies in rats have demonstrated that exposure to foot shock elicits Fos expression, a marker of neuronal activation, in the locus coeruleus and other brain sites. In order to evaluate the involvement of the locus coeruleus in foot shock-induced activation of other brain sites, shock-induced Fos expression was examined in the locus coeruleus and other brain areas known to be activated by foot shock, following direct inhibition of the locus coeruleus by local infusion of muscimol, a GABA agonist, prior to foot shock. Control rats received infusions of artificial cerebrospinal fluid into the locus coeruleus or muscimol into areas outside of locus coeruleus. Rats infused with artificial cerebrospinal fluid and then exposed to foot shock had significant increases in Fos expression in several brain areas, including locus coeruleus, nucleus O, several subdivisions of the hypothalamus, subnuclei of amygdala, bed nucleus of the stria terminalis and cingulate cortex. Inhibition of the locus coeruleus prior to foot shock significantly inhibited Fos expression in the locus coeruleus, nucleus O, some subdivisions of the hypothalamus including the magnocellular and medial parvicellular paraventricular hypothalamic nucleus, subnuclei of amygdala, and cingulate cortex. In contrast, inhibition of the locus coeruleus did not affect shock-induced Fos expression in other areas, including certain subdivisions of the hypothalamus and bed nucleus of the stria terminalis. We suggest that foot shock may activate multiple pathways, with activation of certain discrete nuclei requiring input from the locus coeruleus and activation of others occurring independently of locus coeruleus input. PMID- 11113357 TI - Cellular localization of corticotropin releasing factor receptors in the adult mouse cerebellum. AB - Corticotropin releasing factor is a 41 amino acid peptide that is present in afferent systems that project to the cerebellum. In the adult, this peptide modulates the activity of Purkinje cells by enhancing their responsiveness to excitatory amino acids. Two different types of corticotropin releasing factor receptors, designated type 1 and type 2, have been identified. The purpose of this study is to use immunohistochemistry to identify which corticotropin releasing factor receptors are present in the cerebellum of the adult mouse and to determine their cellular distribution. Receptor type 1 immunostaining is present throughout all lobules of the cerebellar cortex. Distinct labeling is present over the somas of most, if not all, Purkinje cells as well as the primary dendrites of Purkinje cells located at the base of vermal folia. In vermal lobules V, VI, VIII and IX numerous glial fibrillary acidic protein immunoreactive processes, oriented radially in the molecular layer, also are immunoreactive for receptor type 1. In the granule cell layer, scattered type 1 immunoreactive puncta are present throughout most cerebellar lobules. Receptor type 2 immunoreactive puncta are present throughout the molecular layer in all lobules. In addition, scattered basket and/or stellate cells, identified with a GABA antibody, are immunopositive for the type 2 receptor. In the Purkinje cell layer, the type 2 receptor immunolabeling is confined to the basal pole of the Purkinje cell including the initial axonal segment. In the granule cell layer, labeling is present over large cell bodies, and their initial axonal segments. These are likely to be Golgi cells, based on their co-staining with GABA. Finally, numerous elongated processes within the white matter, which are likely to be axons, also are type 2 immunoreactive. These data indicate that both types of corticotropin releasing factor receptor are present in the mouse cerebellum. However, the unique distribution of the two types of receptor strongly suggests a differential role for corticotropin releasing factor in modulating the activity of neurons, axons and glial cells via cell-specific ligand-receptor interactions. PMID- 11113359 TI - Electrophysiological properties of identified trigeminal ganglion neurons innervating the cornea of the mouse. AB - The cornea is innervated by three functional types of neurons: mechanosensory, polymodal and cold-sensitive neurons, all of which are presumed to be nociceptive. To explore if corneal neurons constitute a heterogeneous population according to their electrophysiological properties, intracellular recordings were made in vitro from trigeminal ganglion neurons innervating the cornea of the mouse. Corneal neurons were labelled with FluoroGold applied after a corneal epithelial wound. Five days later, the trigeminal ganglion attached to the eye by its nerves was removed and placed in a superfusion chamber. FluoroGold-positive cells that also responded to electrical stimulation of the cornea were considered corneal neurons. Non-corneal neurons were also studied. Based on their conduction velocity at room temperature, corneal neurons were classified as myelinated A (>1.5m/s) or non-myelinated C (< or =1.5m/s) neurons. A and C neurons differed significantly in their passive and active electrical properties. Virtually all corneal C neurons and about two-thirds of A neurons exhibited a hump in the falling phase of the action potential (S neurons), while the remaining A neurons (F neurons) showed faster and narrower action potentials without a hump. Among non-corneal neurons, A neurons of the F type were found in a proportion of about 50%. Based on their ability to produce somatic action potentials in tetrodotoxin (0.1 microM), non-corneal neurons were classified as fully or partially tetrodotoxin sensitive, which were mainly of the Adelta type, and tetrodotoxin resistant, which were C neurons. Among the corneal neurons, those with a faster action potential, possibly associated to the expression of tetrodotoxin-sensitive Na(+) channels, may be pure corneal mechanosensory neurons, all of which are known to belong to the Adelta type. Neurons with a slower action potential showing a hump in the repolarization phase are both corneal Adelta and C polymodal nociceptive neurons, a type of cell in which tetrodotoxin-resistant Na(+) channels have been identified. The possibility is raised that the small population of neurons with a very high input resistance are cold-sensitive neurons. From the present results, we suggest that the electrophysiological properties of primary sensory neurons innervating the cornea are attributable not only to their conduction velocities, but also to the functional characteristics of their peripheral nerve terminals. PMID- 11113358 TI - Localization and regulation of cyclo-oxygenase-1 and -2 and neuronal nitric oxide synthase in mouse spinal cord. AB - Prostaglandins are important mediators in spinal nociceptive processing. They are produced by cyclo-oxygenase isoforms, cyclo-oxygenase-1 and -2, which are both constitutively expressed in the central nervous system. The present immunohistochemical study details localization and regulation of cyclo-oxygenase 1 and -2 and neuronal nitric oxide synthase in lumbar spinal cord before and after induction of a painful paw inflammation in mice. Cyclo-oxygenase-1 immunoreactivity was found in glial cells of the dorsal and ventral horns, but not in neurons. In unstimulated mice, cyclo-oxygenase-2 immunoreactivity was found in motoneurons of the ventral horns and in lamina X, but not in dorsal horn neurons. After induction of a paw inflammation with zymosan, cyclo-oxygenase-2 immunoreactivity increased dramatically in dorsal horn neurons of laminae I-VI and X, paralleled by a significant increase in prostaglandin E(2) release from lumbar spinal cord. Cyclo-oxygenase-2 was co-localized with neuronal nitric oxide synthase immunoreactivity in several neurons in superficial laminae of the dorsal horns and in the area surrounding the central canal. Nitric oxide synthase was distributed in the cytoplasm and extended to processes of some neurons. In contrast, electron microscopy revealed that cyclo-oxygenase-2 immunoreactivity was restricted to the nuclear membrane and rough endoplasmic reticulum. It is shown in the present study that both cyclo-oxygenase isoforms are constitutively expressed in the spinal cord, cyclo-oxygenase-1 in glial cells of the dorsal and ventral horns and cyclo-oxygenase-2 in motoneurons. After induction of a hindpaw inflammation, several dorsal horn neurons express cyclo-oxygenase-2. Some of them are also positive for neuronal nitric oxide synthase, which is also induced following peripheral inflammation. Intracellularly, cyclo-oxygenase-2 is bound to the membranes of the nucleus and endoplasmic reticulum, whereas neuronal nitric oxide synthase is found in the cytoplasm. PMID- 11113360 TI - Adenosine triphosphate-evoked currents in cultured dorsal root ganglion neurons obtained from rat embryos: desensitization kinetics and modulation of glutamate release. AB - Sensory neurons express purinergic P2X receptors on their central and peripheral terminals as well as their cell bodies. ATP activation of these receptors drives action potential firing and glutamate release with potentially important consequences for sensory function. Here we show ATP-gated currents activated in cultured embryonic dorsal root ganglion neurons have heterogeneity of time courses comparable to those observed in different subpopulations of acutely dissociated adult dorsal root ganglion neurons. The distribution of time-courses across the population of cultured neurons is strongly influenced by culture conditions. Heterogeneity in ATP current kinetics occurs even though immunocytochemical staining reveals a relatively homogeneous and widespread expression of the P2X2 and P2X3 subunits. We show that the time-courses of ATP gated currents recorded at the cell bodies are mirrored by the time-courses of transmitter release from the dorsal root ganglion nerve terminals, indicating similar P2X receptor properties on the soma and their associated terminals. Our results illustrate a functional heterogeneity of P2X receptor-mediated currents that is strongly influenced by external factors. This heterogeneity in current kinetics may have implications for neuronal function as it constrains the time course of ATP-mediated modulation of neurotransmitter release at sensory nerve terminals. PMID- 11113361 TI - Relationship between nociceptor activity, peripheral edema, spinal microglial activation and long-term hyperalgesia induced by formalin. AB - To determine whether initial nociceptive inputs caused by subcutaneous injection of formalin into the hindpaw are necessary and/or sufficient for allodynic behavior and microglial activation observed at one week following behavior, we examined Sprague-Dawley rats under five test conditions. Test condition 1. Formalin alone group (six rats), 5% formalin was injected subcutaneously into the dorsal side of the right hind paw. Test condition 2. Bupivacaine/Formalin group (six rats), bupivacaine was injected into the ankle area and into the site of formalin injection 10 min before formalin injection. Test condition 3. Saline/Formalin group (six rats), saline was injected 10min before formalin in the same manner as bupivacaine. Test condition 4. Formalin/Bupivacaine group 1 (six rats), bupivacaine was injected 10 min after formalin. Test condition 5. Formalin/Bupivacaine group 2 (six rats), bupivacaine was injected similarly 1h after formalin. The magnitude of paw edema and paw withdrawal thresholds to mechanical stimuli applied to the plantar surface of the injected paw and on the dorsal surface of the contralateral side were evaluated prior to and one week after formalin injection. The lumbar spinal cord was immunohistochemically processed at one week to assess the expression of a marker for activated microglia. The results showed: (i) pre-treatment with bupivacaine blocked both phases of formalin-evoked pain behaviors and the mechanical allodynia that developed one week post-formalin injection, but did not block microglial activation; (ii) treatment with bupivacaine 1h after formalin injection reduced paw edema and prevented skin ulceration, but one week allodynia and microglial activation were still present; and (iii) prolonged spinal microglial activation was not dependent on acute formalin-induced nociceptor activity, but was strongly associated with the amount of tissue destruction. Our studies suggest that: (i) the central sensitization associated with the phase II of formalin-evoked behaviors and spinal microglial activation are both necessary to permit the development of the long-term hyperalgesia produced by the subcutaneous administration of formalin into the rat's hindpaw; and (ii) acute nociceptive inputs following formalin injection are not necessary for central microglial activation that may be triggered by nerve damage or prolonged signals from peripherally inflamed tissue PMID- 11113362 TI - Detection of substance P and its receptor in human fetal microglia. AB - Substance P, the most abundant neurokinin in the CNS, is a major modulator of the immune system. We have examined the gene expression of substance P and its receptor in human fetal brain microglia. Using reverse transcription-polymerase chain reaction and Southern blotting assay, the four isoforms of preprotachykinin A gene transcripts (alpha, beta, gamma and delta) were detected in the microglia. The human fetal microglia produced significantly higher levels of endogenous substance P protein (640-850 pg/10(6) cells) than did human peripheral blood monocyte-derived macrophages (25-50 pg/10(6) cells), as determined by an enzyme immunoassay. Using immunohistochemical staining with an anti-substance P antibody, cell membrane substance P immunoreactivity was observed. In addition, we identified the presence of messenger RNA for neurokinin-1 receptor, a primary receptor for substance P in human fetal microglia.From these data, we propose that substance P and its receptor are biologically involved in regulating the functions of microglia, and potentially play an important role in host defense of the central nervous system. PMID- 11113363 TI - beta1 integrin antagonism on adherent, differentiated human neuroblastoma cells triggers an apoptotic signaling pathway. AB - Integrin receptors mediate several functions including prevention of matrix detachment-induced apoptosis (anoikis) of several adherent cell types. We report here that antagonists of beta1 integrins trigger an apoptotic signaling pathway in adherent differentiated LAN-5 human neuroblastoma cells, a cell line which represents a model system for the study of human neurons. The pathway is characterized by cytochrome c release into the cytoplasm, and activation of caspase-9 and caspase-3, 4-6h after treatment; cleavage products of caspase-8 and caspase-2 were not detectable in the cells. Coordinate inactivation of cell survival pathways, including cleavage of focal adhesion kinase, decreased expression of protein kinase B, and reduced phosphorylation of the pro-apoptotic protein, Bad, also characterized the signaling pathway. These events occurred in adherent cells; DNA fragmentation and detachment followed as late events 18-24h after addition of beta1 integrin antagonists. zDEVD-fmk, an irreversible inhibitor of caspase-3-like enzymes, and cytochalasin D, an actin depolymerizing agent, blocked caspase-3 cleavage and delayed cell death. In contrast to these results, undifferentiated, adherent and dividing LAN-5 cells did not die in response to beta1 integrin antagonists. These studies identify a distinct apoptotic pathway which is triggered by antagonists of beta1 integrins on differentiated adherent neuronal cells. PMID- 11113365 TI - A new day. PMID- 11113364 TI - Regulation of N-methyl-D-aspartate receptor expression and N-methyl-D-aspartate induced cellular response during chronic hypoxia in differentiated rat PC12 cells. AB - The purpose of the present study was to examine the effect of chronic hypoxia on N-methyl-D-aspartate-mediated cellular responses in differentiated PC12 cells. PC12 cells were differentiated by treatment with nerve growth factor. Patch-clamp analysis in differentiated PC12 cells showed that extracellularly applied N methyl-D-aspartate induced an inward current that was abolished by the presence of the N-methyl-D-aspartate receptor antagonist MK-801. Results from Ca(2+) imaging experiments showed that N-methyl-D-aspartate induced an elevation in intracellular free Ca(2+) which was also abolished by MK-801. We also examined the effect of hypoxia on the N-methyl-D-aspartate-induced current in nerve growth factor-treated cells. We found that the N-methyl-D-aspartate-induced inward current and the N-methyl-D-aspartate-induced elevation in intracellular free Ca(2+) were markedly attenuated by chronic hypoxia. We next examined the possibility that the reduced N-methyl-D-aspartate responsiveness was due to down regulation of N-methyl-D-aspartate receptor levels. Northern blot and immunoblot analyses showed that both messenger RNA and protein levels for N-methyl-D aspartate receptor subunit 1 were markedly decreased during hypoxia. However, the messenger RNA for N-methyl-D-aspartate receptor subunit 2C was increased, whereas the protein level for subunit 2C did not change. Our results indicate that differentiated PC12 cells express functional N-methyl-D-aspartate receptors and that chronic exposure to hypoxia attenuates the N-methyl-D-aspartate-induced Ca(2+) accumulation in these cells via down-regulation of N-methyl-D-aspartate receptor subunit 1. This mechanism may play an important role in protecting PC12 cells against hypoxic stress. PMID- 11113366 TI - Detection of telomerase activity in prostatic fluid specimens. AB - We report here the extended use of the telomeric repeat amplification protocol (TRAP) assay for the detection of telomerase activity in fresh prostatic fluid obtained from anesthetized patients. Telomerase activity was detected in pellet extract and/or supernatant fluid of specimens obtained from 25 of 30 prostate cancer (PCa) patients (83%), whereas no activity was similarly detectable in specimens taken from 8 of 9 patients (89%) without clinical evidence of PCa. The positive predictive value (PPV) of the TRAP assay for PCa in this pilot study was 96%. We found a strong correlation between telomerase activity in prostatic fluid specimens and serum prostate specific antigen (PSA) values. Telomerase activity was found in 84% of specimens from patients with PSA values >4 ng/ml, whereas in specimens from patients with PSA values 4. In prostatic fluid from PCa patients with Gleason scores of /=pT2 tumors tended to be higher than those in 0.05). Lumbar spine BMD increased 3% (p < 0.001) and distal tibia BMD increased 5% (p < 0.001). TBMC showed a 7.5% increase over the study period. In this group of young men, gain in BMD and TBMC is greatest to age 21 years, with minimal further increase after age 21. The significance of this rise in bone mass during early adulthood on risk for osteoporotic fractures in later life or its impact on exercise-related bone injuries is unknown and warrants further examination. PMID- 11113396 TI - Comparison of morphometric assessment of prevalent vertebral deformities in women using different reference data. AB - The semiquantitative assessment of vertebral deformities is based on visual evaluation. The quantitative approach is based on different morphometric criteria. This study is aimed at comparing the impact of different reference groups to define normal vertebral shape on the diagnosis of verterbral deformities. Reference normal values were obtained in three groups of women: French, mixed European, and Argentinian. All these women had normal lumbar spine bone mineral density and no vertebral deformities according to the semiquantitative assessment. In a group of 135 women having vertebral deformities according to Genant's semiquantitative assessment, three different morphometric criteria were applied. Morphometric diagnosis disclosed a good agreement with semiquantitative assessment. Agreement of diagnosis was higher for a given cutoff using thresholds obtained in different reference groups (kappa = 0.84-0.96) and lower when different criteria were compared using thresholds obtained in the same reference group (kappa = 0. 75-0.85). When fracture thresholds obtained in three different cohorts were compared separately for the three morphometric criteria, agreement was the highest when the cutoff was based only on the arithmetical mean of vertebral heights and was independent of its standard deviation (SD). Average vertebral height ratios did not differ between the three reference cohorts, whereas SDs of vertebral height ratios were the highest in the mixed European cohort and the lowest in the French cohort (F = 7.41, p < 0.001). In the three groups of women of different nationality, SDs of vertebral height ratios, but not the arithmetical means, were significantly higher in the radiographs of poor quality compared with those of good quality. Thus, the main source of difference of diagnosis was related to different SDs whereas average height ratios were not different. Differences in SDs between the three groups were found to be related, at least partly, to poor quality of radiographs. The impact of the differences between populations seems less important, however, only three countries were compared. These findings suggest that those techniques that take into account the SD of vertebral height ratios will provide different reference values for vertebral morphometry. Because differences in SDs depend mainly on the quality of radiographs, they can be reduced by improving the X-ray technique and by the use of standardized protocols. This variability will result in the identification of a variable number of vertebral deformities in osteoporotic women. These results may be of importance especially for multicentric studies. PMID- 11113397 TI - A fifteen-year longitudinal study in young adults on the relation of physical activity and fitness with the development of the bone mass: The Amsterdam Growth And Health Longitudinal Study. AB - Although positive effects of physical activity are often reported, there are still uncertainties about the type, intensity, duration, and frequency of these activities that are most effective for (re)modeling bone mass during youth. In the Amsterdam Growth and Health Longitudinal Study, daily physical activity and fitness were monitored from age 13 to 29 years in a group of 182 males and females. At a mean age of 28 years, bone mineral density (BMD) was measured at three sites with dual X-ray absorptiometry (DXA): in the lumbar region (lumbar BMD), the femoral neck (hip BMD), and the distal radius (wrist BMD). Physical activity (PA) was estimated from a cross-check activity interview taking in consideration all daily physical activities during the last 3 months; PA was scored in two different ways: (1) metabolic physical activity score (METPA) by weighting the intensity (multiples of basic metabolic rate [METs]) and duration (minutes per week); and (2) mechanic physical activity score (MECHPA) by weighting the peak strain (ground reaction forces as multiples of body mass) irrespective of frequency and duration of the physical activities. Physical fitness was measured with a neuromotor fitness test (composite of six strength, flexibility, and speed tests) and as cardiopulmonary fitness (maximal oxygen uptake). The physical activity and fitness scores were calculated over two age periods: during adolescence (13-16 years) and during adulthood (21-27 years). The standardized regression coefficients (corrected for gender, biological age, body composition, and calcium intake) show that weight, physical activity (both METPA and MECHPA), and neuromotor fitness during adolescence and in young adulthood are significantly and positively related with the lumbar BMD (beta = 0. 11-0.40) and hip BMD (beta = 0.18-0.26), measured at the mean age of 28 years. This was not the case for cardiorespiratory fitness. No significant correlations at all are found with wrist BMD, a bone site that is less involved in physical activity and fitness. It can be concluded that daily physical activity during adolescence and in the young adult period is significantly related to the BMD at the lumbar spine and femoral neck at age 28 of males and females. Only neuromotor fitness and not cardiopulmonary fitness during adolescence and young adulthood is related to the BMD of males and females at age 28 years. PMID- 11113399 TI - Fibrogenesis imperfecta ossium (Baker's disease): a case studied at autopsy. AB - A man aged 40 years showed radiographic changes in the form of generalized increased bone density and patchy rarefaction. Urinary calcium was increased and serum alkaline phosphatase was elevated; serum calcium and phosphate levels were normal. Multiple fractures developed. At autopsy, all parts of the skeleton showed partial replacement of bone and bone marrow by a tissue deficient in collagen fibers. Much of this tissue was unmineralized, but lesions in cortical bone showed hypermineralization on microradiographic examination. Electron microscopy showed replacement of collagen fibers by amorphous material in the affected areas. Electron probe analysis showed a normal Ca:P ratio for bone mineral in the hypermineralized areas. PMID- 11113398 TI - Exercise and oral contraceptive use suppress the normal age-related increase in bone mass and strength of the femoral neck in women 18-31 years of age. AB - Women who exercise during their second and third decades may increase their peak bone mass and lower their eventual risk for postmenopausal fracture. However, the effects of exercise in younger women can be modulated by the use of oral contraceptives, which may prevent the normal accretion of bone mass that would otherwise occur. We hypothesized that exercise intervention in young adult women would significantly increase both bone mass and the bending rigidity of the femoral neck. We further hypothesized that exercise intervention in the presence of oral contraceptive use would have a negative effect on bone mass and bending rigidity. Women 18-31 years of age (n = 123) were classified by oral contraceptive use (OC, NOC) and age (18-23, 24-31 years), and then randomized into exercise or nonexercise groups. The exercise protocol consisted of three sessions/week of aerobic and nonaerobic exercises, and continued for 2 years. Each 6 months, the femoral neck of each subject was scanned using a Lunar dual energy X-ray absorptiometry (DEXA) scanner, and bone mineral content, density and geometric information were used to calculate estimated stresses and bending rigidity at the hip. Percent changes from baseline were analyzed using two-way analysis of variance (ANOVA) at 6, 12, 18, and 24 months. Women who neither exercised nor took oral contraceptives (NE/NOC) had the greatest percentage increases in cross-sectional area (4.98 +/- 2.29%), cross-sectional moment of inertia (9.45 +/- 2.37%), total bone mineral density (2.07 +/- 2.09%), fracture index (8.03 +/- 2.03%), and safety factor (20.03 +/- 5.79%) over the 24 month exercise program. Women who exercised and did not take oral contraceptives (E/NOC) declined on most variables related to femoral strength and bone mass, whereas those women who took oral contraceptives were usually intermediate between NE/NOC and E/NOC, whether they exercised or not. These data show that either exercise or OC use is associated with a suppression of the normal increase in bone mass and mechanical strength in the femoral neck in women 18-31 years old, but the combination of exercise and OC use appears to have a less suppressive effect. PMID- 11113400 TI - Bone fractures. PMID- 11113401 TI - Beneficial effects of pravastatin (+/-colestyramine/niacin) initiated immediately after a coronary event (the randomized Lipid-Coronary Artery Disease [L-CAD] Study). AB - Secondary prevention of coronary heart disease by antilipidemic therapy beginning at > or =3 months after an acute coronary syndrome is well documented. The impact, however, of immediate initiation of antilipidemic therapy on coronary stenoses and clinical outcome in patients with acute coronary syndrome is unknown. In our study, patients were randomized, on average, 6 days after an acute myocardial infarction and/or percutaneous transluminal coronary angioplasty secondary to unstable angina, to pravastatin (combined, when necessary, with cholestyramine and/or nicotinic acid) to achieve low-density lipoprotein cholesterol levels of < or =130 mg/dl (group A, n = 70). In controls (group B, n = 56), antilipidemic therapy was determined by family physicians. Quantitative coronary angiography was performed at inclusion, and at 6- and 24-month follow up. The combined clinical end points were total mortality, cardiovascular death, nonfatal myocardial infarction, need for coronary intervention, stroke, and new onset of peripheral vascular disease. Minimal lumen diameter in group A increased by 0.05 +/- 0.20 mm after 6 months and 0.13 +/- 0.29 mm after 24 months, whereas it decreased by 0.08 +/- 0.20 mm and 0.18 +/- 0.27 mm, respectively, in group B (p = 0.004 at 6 months and p <0.001 at 24 months). After 2 years, 29 patients of 56 patients in group B, but only 16 of 70 patients in group A, experienced a clinical end point (p = 0.005; odds ratio 0.28, confidence intervals 0.13 to 0.6). We conclude that pravastatin-based therapy initiated immediately after an acute coronary syndrome is well tolerated and safe, lessens coronary atherosclerosis, and has a pronounced clinical benefit. PMID- 11113402 TI - Comparison of gated single-photon emission computed tomography with magnetic resonance imaging for evaluation of left ventricular function in ischemic cardiomyopathy. AB - To perform a head-to-head comparison between magnetic resonance imaging (MRI) and gated single-photon emission computed tomography (SPECT) for the evaluation of left ventricular (LV) function (LV ejection fraction [LVEF], LV volumes, and regional wall motion) in patients with ischemic cardiomyopathy, we studied 22 patients with chronic coronary artery disease and LV dysfunction. Multislice, multiphase echoplanar MRI was performed with Philips Gyroscan ACS-NT15. Image analysis was performed using the MASS software package to determine LV end systolic volume, LV end-diastolic volume, and LVEF. The same parameters were calculated using quantitative gated SPECT software (QGS, Cedars-Sinai Medical Center). The different parameters were compared using linear regression, and correlation coefficients were calculated. Regional wall motion was also determined from both techniques, according to a 13-segment model and a 3-point scoring system (from 1 = normokinesia to 3 = akinesia or dyskinesia). A summed wall motion score was also calculated for MRI and gated SPECT. Good correlations were found between MRI and gated SPECT for all parameters: (1) summed wall motion scoreMRI versus summed wall motion scoreSPECT: y = 0.74x + 8.0, r = 0.88, p <0.01; (2) LV end-systolic volumeMRI versus LV end-systolic volumeSPECT: y = 0.94x - 12.3, r = 0.87, p <0.01; (3) LV end-diastolic volumeMRI versus LV end diastolic volumeSPECT: y = 0.93x - 18.4, r = 0.84, p <0.01; and (4) LVEFMRI versus LVEFSPECT: y = 0.97x + 0.68, r = 0.90, p <0.01. For regional wall motion, an exact agreement of 83% was found, with a kappa statistic of 0.77 (95% confidence intervals 0.71 to 0.83, SE 0.03), indicating essentially excellent agreement. Thus, close and significant correlations were observed for assessment of LVEF, LV volumes, and regional wall motion by MRI and gated SPECT in patients with ischemic cardiomyopathy. PMID- 11113403 TI - Coronary calcium and anti-cardiolipin antibody are elevated in patients with typical chest pain. AB - The aim of this study was to examine whether detection of coronary calcium and the autoimmune response associated with atherosclerosis, either solely or in combination, are different in patients with typical and atypical chest pain. Coronary calcium as detected by spiral computerized tomography and levels of antibodies against cardiolipin (CL), oxidized low-density lipoprotein (ox-LDL), and beta2-glycoprotein-I (beta2-GPI) were studied in patients with typical chest pain (n = 52), atypical chest pain (n = 19), or without chest pain (n = 21). Patients with typical chest pain had higher mean levels of coronary calcium (expressed as natural transformation of total coronary calcium score) compared with patients with atypical chest pain and controls (5.04 vs 3.21 and 2.75, respectively; p < 0.001). The levels of anti-CL were (mean +/- SD of optical density multiplied by 1,000): 262 +/- 140, 170 +/- 82, and 230 +/- 115 for patients with typical chest pain, atypical chest pain, and controls, respectively (p = 0.016). No significant difference was found between groups regarding anti-ox LDL and anti-beta2-GPI autoantibody levels. In the typical chest pain group, there was a higher prevalence of high total coronary calcium scores (p = 0.03) and high anti-CL levels (p = 0.01) than in the atypical chest pain group. Eighteen of 52 patients with typical chest pain (35%) had both high calcium scores and high antibody levels, whereas none of the 19 patients (0%) who had atypical chest pain had high levels of both (p = 0.003). A combination of both coronary calcium and anti-CL was associated with higher area under the receiver operator characteristic curves than for each separately. High coronary calcium scores or high anti-CL levels are found more often in typical than in atypical chest pain patients, but a combination of high levels of both can better differentiate typical from atypical chest pain patients. PMID- 11113404 TI - Usefulness of QT dispersion immediately after exercise as an indicator of coronary stenosis independent of gender or exercise-induced ST-segment depression. AB - Several recent studies suggest that QT dispersion on a standard 12-lead electrocardiogram is a clinically useful indicator of significant coronary stenosis. In this study, we compared the diagnostic accuracy of QT dispersion immediately after exercise as an indicator of coronary stenosis in men and women, and in the presence or absence of exercise-induced significant ST-segment depression. The subjects were 273 consecutive patients (mean age 56 +/- 9 years; 190 men and 83 women) without a history of myocardial infarction who underwent treadmill exercise electrocardiography and coronary angiography for evaluation of angina. Of these, 146 patients had no significant coronary stenosis, 61 had single-vessel disease, 56 had multivessel disease, and 10 had left main coronary artery disease. QT dispersion immediately after exercise was significantly greater in patients with significant coronary stenosis than in those without (64 +/- 14 vs 39 +/- 14 ms, p <0.01). QT dispersion immediately after exercise was significantly more sensitive in men (sensitivity 75%; specificity 85%) and significantly more specific in women (sensitivity 77%, specificity 88%) than exercise-induced significant ST-segment depression (men: sensitivity 62%, specificity 74%; women: sensitivity 81%, specificity 68%) as an indicator of significant coronary stenosis. The addition of factors such as gender and the presence or absence of exercise-induced significant ST-segment depression did not significantly alter the sensitivity and specificity of QT dispersion immediately after exercise for detecting significant coronary stenosis (patients with significant ST-segment depression: sensitivity 77%, specificity 88%; patients without significant ST-segment depression: sensitivity 72%, specificity 86%). In conclusion, QT dispersion immediately after exercise is a clinically useful indicator of significant coronary stenosis independent of gender or the presence or absence of exercise-induced significant ST-segment depression. PMID- 11113405 TI - Effect of preintervention plaque burden on subsequent intimal hyperplasia in stented coronary artery lesions. AB - We sought to determine if axial and circumferential distribution of plaque before stenting determines the axial and circumferential distribution of subsequent intimal hyperplasia (IH). We studied 22 patients with a single Palmaz-Schatz stent implanted in a native coronary artery, who underwent intravascular ultrasound (IVUS) imaging before intervention, after stenting, and at 6-month follow-up. For each lesion, 7 locations were analyzed: proximal and distal reference, proximal and distal edge of the stent, proximal and distal location within the body of the stent, and the articulation. Pre- and postintervention and follow-up image slices were precisely aligned and analyzed for pre- and postintervention plaque area and follow-up IH area and thickness. The location of maximal IH area was at or adjacent to the location of maximal preintervention plaque in 17 of 22 of the patients (77%). Similiarly, the circumferential distribution of IH at follow-up paralleled the eccentricity pattern of the native plaque burden in 69% (24 of 35 slices). Using multivariant analysis, the strongest predictor of IH was preintervention plaque area (p = 0.001). IH accumulates axially and circumferentially preferentially at the site of maximal preintervention plaque. PMID- 11113406 TI - Comparison of medicine alone, coronary angioplasty, and left internal mammary artery-coronary artery bypass for one-vessel proximal left anterior descending coronary artery disease. AB - Despite the deleterious and sometimes catastrophic consequences of proximal left anterior descending (LAD) artery occlusion, there is a paucity of data to guide the treatment of patients with such disease. Our aim was to describe outcomes with medical therapy, angioplasty, or left internal mammary artery (LIMA) bypass grafting in patients with 1-vessel, proximal LAD disease. We retrospectively analyzed prospectively collected data from 1,188 patients first presenting only with proximal LAD disease at 1 center over 9 years. We assessed the rates of death, acute myocardial infarction, and repeat intervention by initial treatment over a median 5.7 years of follow-up. Patients undergoing angioplasty or LIMA bypass were more often men and had progressive or unstable angina; those receiving medical therapy had a lower median ejection fraction. Both revascularization procedures offered slightly better adjusted survival versus medicine (hazard ratio for angioplasty, 0.82; 95% confidence interval, 0.60 to 1.11; hazard ratio for bypass, 0.74; 95% confidence interval, 0.44 to 1.23). Bypass, but not angioplasty, was associated with significantly fewer composite end point events (death, infarction, or reintervention, p <0.0001), and angioplasty was associated with a higher composite event rate than bypass or medical therapy (p <0.0001 and p = 0.0003, respectively). The initial advantages of bypass and medicine over angioplasty diminished over time; angioplasty became more advantageous than medicine after 1 year (p = 0.05) and not significantly different from bypass. Treatment of 1-vessel, proximal LAD disease with medicine, angioplasty, or UMA bypass resulted in comparable adjusted survival. However, LIMA bypass alone reduced the long-term incidence of infarctions and repeat procedures. PMID- 11113407 TI - Control of paroxysmal atrial fibrillation recurrence using combined administration of propafenone and quinidine. AB - The frequent recurrence of paroxysmal atrial fibrillation (PAF) despite the use of standard antiarrhythmic agents prompted the use of new therapeutic approaches. There are few data on systematic assessment of PAF control with stepwise dose escalation and the use of a drug combination. Low-dose quinidine may promote the efficacy of propafenone by inhibiting its degradation through the cytochrome P450 pathway (CYP2D6). We prescribed propafenone 300 to 450 mg/day to 60 patients with PAF for 8 weeks, and 62% were symptomatically controlled. The 19 refractory patients were randomized in a double-blinded fashion to receive either a higher dose of propafenone (450 to 675 mg/day) or the standard dose of propafenone with low-dose quinidine 150 mg/day, each for an 8-week study period, and subsequently crossed over to the alternative treatment. The resulting serum propafenone concentrations were 259 +/- 208 and 336 +/- 237 mg/day (p >0.5), respectively. Both treatment arms prolonged the time to the first symptomatic atrial fibrillation (AF) recurrence and the interval between attacks, and AF was controlled in 37% of patients. However, the higher dose of propafenone was associated with gastrointestinal side effects not present with the low-dose quinidine combination. Of the 10 refractory patients, 7 were further controlled with a standard dose of propafenone plus quinidine (600 mg/day). Overall, control of PAF was achieved in 85% of patients at the end of 8 months; adverse effects necessitating withdrawal were observed in 6%, and uncontrolled AF in 5% of patients. There was no difference in the mean AF rate during recurrences in all phases, and ventricular proarrhythmia was not seen. This study documents the role of stepwise antiarrhythmic treatment of PAF. The use of a standard dose of propafenone, followed by low-dose quinidine combination to reduce propafenone degradation, and the combined standard dose of propafenone and quinidine may be used to maximize efficacy and tolerability. PMID- 11113408 TI - Electrophysiologic characteristics of diverse accessory pathway locations of antidromic reciprocating tachycardia. AB - This study assessed antidromic reciprocating tachycardia (ART) in patients with paraseptal accessory pathways (APs). Previous clinical experience suggests that paraseptal APs are unable to serve as the anterograde limb during ART. Based on the reentry wavelength concept, we hypothesized that anatomic location of a paraseptal AP may not preclude occurrence of ART. If wavelength criteria were met due to prolonged conduction time retrogradely in the atrioventricular node or anterogradely in the AP, ART may be sustained. All patients who had ART in the electrophysiologic laboratory at our institution (1991 to 1998) were studied. Based on fluoroscopically guided electrophysiologic mapping and radiofrequency ablation, AP location was classified as paraseptal, posterior, or lateral. Conduction time and refractoriness measurements were made for all components of the ART circuit. Of 24 patients with ART, 5 (21%) had ART utilizing a paraseptal AP. Anterograde conduction time through the AP and retrograde atrioventricular nodal conduction time were significantly longer in patients with paraseptal versus lateral pathways. Isoproterenol was required for ART induction in 38% of patients with a posterior AP, 36% with lateral AP location, but not in patients with a paraseptal AP. There were no significant differences in tachycardia cycle length or refractoriness of anterograde and/or retrograde components of the macroreentry circuit between the 3 pathway locations. Thus, ART can occur in patients with a paraseptal AP. Slower anterograde pathway conduction, or retrograde atrioventricular nodal conduction renders the wavelength critical for completion of the antidromic re-entrant circuit. PMID- 11113409 TI - Usefulness of a history of tobacco and alcohol use in predicting multiple heart failure readmissions among veterans. AB - Multiple hospital readmissions for heart failure (HF) are progressively increasing and may be related to continued tobacco and alcohol use. To study this relation, we conducted a retrospective chart audit of all veterans discharged with HF at a large Veterans Administration (VA) facility from 1997 to 1998. Using a multivariate logistic regression model, the smoking and alcohol use of patients who required > 1 HF admission within 1 year were compared with those who did not. Demographic, clinical, and psychosocial variables were also included in the model. Of 753 patients admitted with HF during the review period (mean age 69.1 years, 99% men), 220 patients (29.2%) were readmitted to the hospital at least once (range 1 to 8 readmissions, mean 1.79 +/- 0.27) after the index admission. In a multivariate analysis, current smoking (odds ratio [OR] 1.82; confidence interval [CI] 1.17 to 2.82) and current alcohol use (OR 5.92; CI 3.83 to 9.13) were independent predictors of readmissions. Other predictors included living alone (OR 2.09; CI 1.42 to 3.09), HF associated with ischemic etiology (OR 3.99; CI 2.58 to 6.18), higher New York Heart Association class (OR 2.57; CI 1.86 to 3.55), and care provided by a primary care physician compared with a cardiologist (OR 2.41; CI 1.57 to 3.67). This study confirms that noncompliance to smoking and alcohol restrictions, which are amenable to change, dramatically increases the risk for multiple hospital readmissions among patients with HF. Consequently, evaluation of noncompliance to smoking and alcohol consumption with targeted interventions in this population may be a key component for the reduction of multiple hospital readmissions. PMID- 11113410 TI - Three-dimensional color Doppler reconstruction of intracardiac blood flow in patients with different heart valve diseases. AB - An improved perception of the magnitude and dynamics of intracardiac flow disturbances has been made possible by the advent of 3-dimensional (3-D) color Doppler, a new diagnostic procedure developed at our institution. This study describes the new insights derived from 3-D reconstruction of color Doppler flow patterns in patients with different heart valve diseases. The color Doppler flow data from 153 multiplanar transesophageal or transthoracic echocardiographic examinations has been obtained from 133 patients with heart valve disease; 73 patients had mitral regurgitation, 15 had mitral stenosis, 18 had aortic regurgitation, 26 had aortic stenosis, and 21 patients had tricuspid regurgitation. Four patients had pulmonary regurgitation associated with mitral valve disease. The 3-D reconstructions of color Doppler flow signals were accomplished by means of the "Heidelberg Raytracing model," developed at our institution. The 3-D color Doppler reconstructions were obtained in all patients. The 3-D images revealed for the first time the complex spatial distribution of the blood flow abnormalities in the heart chambers caused by different heart valve diseases. New patterns of intracardiac blood flow disturbances were observed and classified. Three-dimensional color Doppler provides a unique noninvasive method that can be easily applied for studying intracardiac blood flow disturbances in clinical practice. PMID- 11113411 TI - Echocardiographic assessment of the mechanisms of correction of bileaflet prolapse causing mitral regurgitation with only posterior leaflet repair surgery. AB - Recent data suggest that posterior leaflet repair alone corrects mitral regurgitation in patients with bileaflet prolapse and normal anterior chordae. The purpose of this study was to use echocardiography to define the anatomic differences between posterior and bileaflet prolapse and to determine if posterior leaflet repair alone leads to correction of bileaflet prolapse. We studied patients who underwent quadrangular resection of the posterior mitral valve leaflet to treat bileaflet prolapse (group I, n = 20) or isolated posterior leaflet prolapse (group II, n = 20). Echocardiographic characteristics were compared before and after the procedure. There were no differences in the left ventricular end-diastolic or end-systolic dimensions or function between the 2 groups. However, anterior leaflet length was greater in patients with bileaflet prolapse (3.3 +/- 0.6 cm vs 2.6 +/- 0.4 cm, p = 0.003). In group I, posterior leaflet repair changed anterior leaflet displacement from -0.8 +/- 0.2 to 0.5 +/- 0.4 cm (p <0.001) and posterior leaflet displacement from -0.8 +/- 0.3 cm below to 0.5 +/- 0.4 cm (p <0.001) in front of the mitral annular plane. In group II, anterior leaflet displacement was unchanged from 0.2 +/- 0.1 to 0.3 +/- 0.2 cm (p = 0.22), whereas posterior leaflet displacement changed from -0.7 +/- 0.2 to 0.4 +/- 0.2 cm (p <0.001). Thus, patients with bileaflet prolapse and no ruptured chords have excessive anterior leaflet length. In such patients, posterior leaflet repair alone corrects anterior and posterior leaflet prolapse. PMID- 11113412 TI - Right and left ventricular systolic function late after repair of tetralogy of Fallot. AB - Right ventricular (RV) dysfunction has adverse effects on long-term outcome in patients with repaired tetralogy of Fallot (TOF). We employed serial radionuclide angiography (RNA) to examine RV and left ventricular (LV) systolic function in adults late after TOF repair and its relation to clinical outcome. We reviewed 10 year records of 95 patients (53 men) with TOF followed in our clinic (mean age at repair 12.6 +/- 10.5 years, mean age at last follow-up 37.7 +/- 9.8 years) who underwent at least 2 RNAs between 1987 and 1997. Most patients were well by the end of the study (80% were New York Heart Association class I, 17% were class II, and 3% were in class III). Sixteen patients experienced sustained tachyarrhythmias (8 had atrial; 8 patients had ventricular). One patient died suddenly. Fifteen patients underwent RV outflow reoperations (15 underwent pulmonary valve replacement; 7 had relief of RV outflow obstruction); RV systolic function during exercise in these 15 patients was significantly impaired before and returned to similar levels after surgery, compared with the rest of the patients. Overall, RV and LV function remained stable in the whole group at a mean interval of 5.7 +/- 2.2 years between first and last RNA. This group of closely followed adults with TOF remained well over 10 years with a low incidence of sudden death and stable RV and LV systolic function, despite a relatively large number of RV outflow reoperations. Aggressive intervention for right-sided hemodynamic abnormalities may have contributed to this outcome. Preserved ventricular function may herald a favorable long-term outlook in this group. PMID- 11113413 TI - Comparison of four echocardiographic techniques for measuring left ventricular ejection fraction. AB - Accurate quantitative measurement of left ventricular (LV) ejection fraction (EF) by 2-dimensional echocardiography is limited by subjective visual endocardial border detection. Both harmonic and precision contrast microbubbles provide better delineation of endocardial borders than fundamental imaging. The aim of this study was to correlate 2-dimensional echocardiographic quantification of LVEF measured by 4 currently available techniques with radionuclide angiography. A total of 50 patients who underwent radionuclide (EF) measurement (47 of 50 had technically difficult echocardiograms by fundamental imaging) underwent echocardiography by 4 methods: fundamental alone, fundamental with contrast, harmonic alone, and harmonic with contrast. Three echocardiologists measured the biplane 2-dimensional echocardiographic LVEF independently and were blinded to radionuclide angiography. The correlation of echocardiographic EF with radionuclide EF improved incrementally with each method. However, contrast with harmonic imaging provided the closest correlation (r = 0.95, 0.96, and 0.95 as assessed by the 3 independent analysts. PMID- 11113414 TI - Clinical impact of borderline and undiagnosed diabetes mellitus in patients with coronary artery disease. AB - The present study was aimed to evaluate the prevalence and prognostic significance of unrecognized and newly defined borderline diabetes (with fasting blood sugar 126 to 139 mg/dl) by the American Diabetes Association criteria in coronary patients over a 7.7-year follow-up. Both undiagnosed and newly diagnosed borderline diabetes were associated with an unfavorable metabolic profile. The mortality of the borderline diabetics tended to be higher than in their nondiabetic counterparts. but this tendency did not reach statistical significance. A significant excess in long-term mortality was observed among the undiagnosed diabetes group. PMID- 11113415 TI - Comparison between strategies using creatine kinase-MB(mass), myoglobin, and troponin T in the early detection or exclusion of acute myocardial infarction in patients with chest pain and a nondiagnostic electrocardiogram. AB - Different strategies using creatine kinase-MB(mass), myoglobin, and troponin T were compared in 738 patients admitted because of chest pain and an electrocardiogram not diagnostic of acute myocardial infarction. We conclude that a combination of creatine kinase-MB and troponin T during the first 6 hours enables early detection or exclusion of acute myocardial infarction in this population. PMID- 11113416 TI - Diagnostic and prognostic role of myoglobin in patients with suspected acute coronary syndrome. North-Wurttemberg Infarction Study (NOWIS) Group. AB - In patients with suspected acute coronary syndrome, myoglobin is, according to IFCC and NACB guidelines, the marker of choice for early determination of acute infarction, in particular in combination with creatine kinase-MB, 4 hours after admission with a sensitivity of 96%, and correctly excludes Q-wave infarctions. In patients without acute myocardial infarction, a positive troponin T (relative risk 31.5%), but not an elevated myoglobin (relative risk 4.5%), is highly predictive for adverse in-hospital outcome. PMID- 11113417 TI - Global impairment of coronary blood flow in the setting of acute coronary syndromes (a RESTORE substudy). Randomized Efficacy Study of Tirofiban for Outcomes and Restenosis. AB - Acute coronary syndromes result in a global impairment of coronary blood flow with nonculprit artery blood flow being associated with culprit artery flow and vice versa. Improvements in nonculprit artery flow are related to improvements in culprit artery flow after percutaneous intervention; nonculprit arteries with abnormal flow sustain greater improvements in their flow after culprit artery intervention. PMID- 11113418 TI - Impact of injections during diagnostic coronary arteriography on coronary patency in the setting of acute myocardial infarction from the TIMI trials. Thrombolysis In Myocardial Infarction. AB - The mechanical force of injection at 90 minutes opens 13.4% of occluded arteries, but overall, only 2.4% of all culprit arteries (already open and occluded combined) are opened. Thus, although some arteries are opened by the force of hand injection, the frequency of mechanical opening among all arteries is low, and hand injections appear to alter current 80% patency rates by approximately 2.5%. PMID- 11113419 TI - Initial results and long-term clinical and angiographic outcome of coronary stenting in women. AB - To assess whether gender influences the results of coronary stenting, 158 consecutive women undergoing coronary stenting were compared with 823 consecutive men. Women had more adverse baseline characteristics, a higher hospital mortality, and were independently associated with procedural failure/complications (relative risk 2.4, 95% confidence interval 1.2 to 4.8); however, the long-term event-free survival and the restenosis rate were not influenced by gender. PMID- 11113420 TI - Immediate reinitiation of atrial fibrillation after electrical cardioversion predicts subsequent pharmacologic and electrical conversion to sinus rhythm and amiodarone. AB - Ninety-one percent of patients with atrial fibrillation (AF) resisting electrical cardioversion because of immediate reinitiation of AF after the shock maintain long-term sinus rhythm with amiodarone and repeat cardioversion. Thus, immediate reinitiation of AF is an important sign that should guide further treatment in patients with electrical cardioversion-resistant AF. PMID- 11113421 TI - Acute electrophysiologic effect of estradiol 17beta in menopausal women. AB - Sixteen postmenopausal women underwent electrophysiologic study before and 20 minutes after the administration of sublingual estradiol 17beta or placebo. Estradiol 17beta significantly affected electrophysiologic parameters, thereby suggesting its role in the development of palpitations in women. PMID- 11113422 TI - Internal transcardiac pericardiocentesis for acute tamponade. AB - If the catheter is still in the pericardium when tamponade is recognized during catheterization or electrophysiologic procedures, it can be used for definitive aspiration and relief of tamponade. This is physiologically beneficial to the patient, and psychologically beneficial to both patient and medical staff. PMID- 11113423 TI - Dynamics of the uncorrected QT interval during vagal-induced lengthening of RR intervals. AB - Analysis of 21 episodes of vagal-induced atrioventricular block showed that the uncorrected QT intervals at the end of the corresponding RR pauses were not prolonged, in reference to the pre-block QT intervals, with pauses shorter than 1,280 ms. Subsequently, they gradually lengthened as the RR pauses progressively increased to 13,710 ms. This dynamic behavior of the QT interval in subjects without structural heart disease could have resulted from a complex interaction between the cumulative effects of previous cycle lengths (memory effect?) and the autonomic nervous system. PMID- 11113424 TI - Effects of carvedilol on heart rate and blood pressure variability in subjects with chronic heart failure. AB - In this study we observed that carvedilol administration to patients with heart failure improves hemodynamic function, baroreflex sensitivity, and heart rate variability. These findings contribute to improvement in survival in such patients. PMID- 11113425 TI - Effect of surgical repair of secundum-type atrial septal defect on right atrial, right ventricular, and left ventricular volumes in adults. AB - Surgical repair of atrial septal defect in adults reduces right ventricular and right atrial diameters and volumes, and improves left ventricular filling. PMID- 11113427 TI - The surgeon and breast ultrasonography procedures. PMID- 11113426 TI - Both atenolol and propranolol blunt the fibrinolytic response to exercise but not resting fibrinolytic potential. AB - This randomized, double-blind trial found that tissue plasminogen activator activity increased and plasminogen activator inhibitor-1 activity decreased significantly more with exercise during placebo treatment than during treatment with beta blockade. These results suggest that beta blockade blunts the fibrinolytic response to maximal exercise. PMID- 11113428 TI - Breast surgery: past, present, and future. PMID- 11113429 TI - Breast cancer chemoprevention. AB - Chemoprevention of breast cancer is a rapidly growing field. Chemoprevention was initiated with the development of the antiestrogen tamoxifen. A major clinical trial in the United States found that tamoxifen reduced the incidence of breast cancer by almost 50% in women at an increased risk for the disease. Although two European trials did not confirm these findings, the Food and Drug Administration found the American studies significant enough to approve tamoxifen for the delaying of breast cancer in women at high risk for the disease. However, adverse effects associated with tamoxifen include a minimally increased rate of endometrial cancer, cataracts, and strokes. Newer classes of antiestrogens, called selective estrogen receptor modulators (SERMs), are being investigated as potential chemopreventive agents. These SERMS, such as raloxifene, will hopefully provide some of the benefits of estrogen without its inherent risks. In addition, naturally occurring compounds and their analogues are also under investigation. PMID- 11113430 TI - Presentation, management and outcome of axillary recurrence from breast cancer. AB - BACKGROUND: The impact of axillary node dissection on breast cancer survival is unclear. Limited axillary surgery has been proposed but may increase regional recurrence rates. Optimal management for axillary recurrence is poorly understood. METHODS: Axillary recurrences were initial treatment failure sites in 44 of 4,255 breast cancer patients (1%) seen at M.D. Anderson Cancer Center, 1982 to 1992. RESULTS: Twenty-one patients (48%) had early stage disease (0, I, II) at diagnosis. With 70.8 months median follow-up, complete control of axillary recurrence was achieved in 31 patients (71%). Distant metastases developed in 50% and were more likely with uncontrolled axillary recurrences. Failure to receive multimodality therapy and failure to undergo surgery for the recurrence correlated with resistant axillary disease. CONCLUSIONS: Axillary recurrence from breast cancer is uncommon but may follow any stage of disease. One half of affected patients develop distant metastases. Durable disease control is best achieved with multimodality therapy including a surgery component. PMID- 11113431 TI - The details of successful sentinel lymph node staging for breast cancer. AB - Sentinel lymphadenectomy is an effective and accurate tool for staging breast cancer. In recent years the details of a successful program have become better defined. The authors outline practical considerations for the performance of successful sentinel lymph node staging from a multidisciplinary perspective. PMID- 11113432 TI - State-of-the-art lymph node staging for breast cancer in the year 2000. AB - Axillary staging for breast cancer is vitally important for determining appropriate adjuvant hormone and chemotherapy. In the absence of distant metastases, axillary lymph node status remains the most accurate predictor of clinical outcome. Sentinel lymph node biopsy is a minimally invasive approach with enhanced accuracy and less morbidity than conventional axillary dissection. The stage is now set for the sentinel lymphadenectomy staging to move from state of-the-art care to the standard care in coming years. PMID- 11113433 TI - Credentialing issues with sentinel lymph node staging for breast cancer. AB - Sentinel lymphadenectomy (SL) is a minimally invasive approach for staging patients with breast cancer. SL, when performed in lieu of axillary dissection, is associated with less morbidity and is potentially more cost effective and more accurate than the historical axillary dissection in the detection of regional nodal metastases. The credentialing and privileging of SL, as with any surgical procedure, is by the policies of the local hospital or institution. The suggested credentialing criteria for local hospitals has been an area of controversy. Herein the authors outline the credentialing controversy and suggest criteria for the implementation of sentinel lymph node staging for breast cancer. PMID- 11113434 TI - The role of sentinel lymph node biopsy in women undergoing prophylactic mastectomy. AB - BACKGROUND: Indications for prophylactic mastectomy (PM) range from LCIS to BRCA 1-2 positive, cosmesis, and cancer phobia. Occult cancers have been found in up to 5% of PM cases. Consequently, consideration must be given to the role of sentinel lymph node (SLN) biopsy as a diagnostic procedure in these patients as PM excludes the subsequent option of SLN biopsy. METHODS: From April 1994 to November 1999, all patients undergoing PM had SLN biopsy after four quadrant periareolar injections of radiocolloid (450 mci) and blue dye (5 cc). All patients were prospectively accrued to the computerized database of breast patients. The SLN were all evaluated with hematoxylin and eosin (H&E) as well as CAM5.2 cytokeratin immunohistochemical (CK-IHC) stains. RESULTS: Over a 67-month period, 1,356 patients were mapped; 57 patients underwent PM in which 148 nodes (2.6 nodes per patient) were evaluated. Nodes were examined by routine H&E and CK IHC staining. Two patients, neither of whom was found to have a cancer in the prophylactic mastectomy breast, were found to have a positive SLN by CK-IHC staining. Infiltrating carcinoma was discovered within the PM breasts of 2 additional patients. Sentinel lymph node biopsy was negative for malignancy by H&E as well as CK-IHC stains. No lymphedema has been detected in PM patients. CONCLUSIONS: Sentinel node biopsy has been shown to be an accurate and minimally invasive method of evaluating the lymphatic basin. This study shows that the absence of known disease within the breast does not preclude the presence of occult cancer or metastatic nodal disease. Four patients (7%) had a significant change in their surgical management as a direct result of sentinel lymph node biopsy. Two patients were spared the complications of a complete axillary node dissection. This minimally invasive procedure accurately evaluated the known disease status and provided new diagnostic information. Most important, once a mastectomy is performed, the opportunity for SLN biopsy is lost should a cancer be found within the breast specimen. PMID- 11113435 TI - Axillary dissection in the context of the biology of lymph node metastases. AB - BACKGROUND: Modern breast surgery, as the primary treatment of invasive breast carcinoma, has been evolving over the last century. Aggressive radical surgery, which included chest wall resection, complete axillary clearance and internal mammary node dissection, has slowly changed to a less aggressive approach. This has been based on an improved understanding of the biology of the disease. Over the years, randomized prospective trials, performed at centers all over the world, have demonstrated that axillary dissection does not impact on the overall survival while it helps with loco-regional control of breast cancer. Its major role, at the present time, is limited to staging and prognostication; functions that are equally well served by the limited approach of a sentinel node biopsy. SOURCES: This review is based on the available medical literature involving the biology and organ specificity of the metastatic process, not only in breast cancer but also in other malignancies. In addition, studies pertaining to clinical breast cancer, and the role of surgery in its treatment, were reviewed. The ongoing trials on the role of sentinel node biopsy in the management of the clinically node negative patients are discussed. CONCLUSIONS: This review covers the history, pathophysiology, and clinical basis of the current role of axillary dissection for invasive breast cancer. From the data presented we hope that the medical community will agree that there is no therapeutic role for extended axillary dissection at the current time. PMID- 11113436 TI - Diagnostic and interventional ultrasound for breast disease. AB - The availability of reliable, portable computer-enhanced ultrasonography with high-frequency transducers has improved breast ultrasonography such that its role has increased dramatically. Diagnostic characteristics of breast lesions may be used to categorize these lesions according to their relative risk for malignancy. Furthermore, breast ultrasonography may be used to guide needle aspiration and biopsy of lesions so indicated by diagnostic evaluation. Results of ultrasound guided aspiration and core biopsy accurately diagnose specific histopathology thereby avoiding unnecessary open biopsy for benign lesions and facilitating therapeutic planning for malignant lesions. PMID- 11113437 TI - Mammographic screening of the high-risk woman. AB - Annual screening mammography beginning at age 40 is recommended for the general population. For some women at high risk for developing breast cancer at a younger age, annual screening may be appropriate starting at an earlier age. These women include those with a personal history of breast cancer, nontherapeutic radiation to the breasts especially for Hodgkin's disease, BRCA positive women, women with a family history of a first-degree relative with breast cancer at a young age, and women with a biopsy diagnosis of lobular carcinoma in situ or atypical ductal hyperplasia. Women with a biopsy diagnosis of atypical lobular hyperplasia develop breast cancer after age 40 and do not need earlier screening, unless they have a family history of breast cancer. Although increasing a woman's risk for breast cancer, radial scar does not increase risk for women younger than 40 years old and therefore does not require screening at a young age. PMID- 11113438 TI - Skin-sparing mastectomies. PMID- 11113439 TI - The breast surgeon's role in BRCA1 and BRCA2 testing. AB - Five percent to 10% of all women who develop breast cancer carry a hereditary mutation in the genes BRCA1 or BRCA2. Genetic testing is now clinically available, and the results of such testing can dramatically alter a patient's risks for an ipsilateral or contralateral primary breast cancer and ovarian cancer. Therefore, genetic testing will become integral in tailoring surveillance, chemoprevention, and surgical management plans for patients at risk for hereditary cancer syndromes. Such results will also impact the cancer risks for the patient's nuclear and extended family members. Surgeons will play a pivotal role in eliciting personal and family histories from patients, determining which of those histories is suggestive of a germline mutation, facilitating referrals for genetic counseling and testing, and incorporating the results of genetic testing into the patient's short- and long-term management plans. PMID- 11113440 TI - Long-term results of wide-field brachytherapy as the sole method of radiation therapy after segmental mastectomy for T(is,1,2) breast cancer. AB - BACKGROUND: We hypothesized that wide-field brachytherapy (BRT) after margin negative excision would result in complication rates, local recurrence rates, and cosmesis scores equivalent to external beam radiotherapy (ERT). METHODS: Patients with T(is,1,2) tumors less than or equal to 4 cm, 0 to 3 positive axillary nodes, and negative inked surgical margins were entered prospectively into BRT phase I/II trial. Patients who met the eligibility criteria for BRT but were treated with ERT during the same time period were retrospectively identified as controls. A blinded panel of healthcare professionals graded cosmetic outcome. RESULTS: Fifty patients with 51 breast cancers received BRT from January 1992 to October 1993. We identified 94 patients eligible for BRT but concurrently treated with ERT. At a median follow-up of 75 months, the two groups were similar for grade III treatment toxicities, local/regional recurrence rates, and cosmesis scores. CONCLUSIONS: For selected breast cancer patients undergoing breast-conserving therapy, BRT is an attractive alternative to ERT. PMID- 11113441 TI - Are microinvasion and micrometastasis in breast cancer mountains or molehills? AB - BACKGROUND: The increased rate of early detection of breast cancer due to widespread mammographic screening has led to an increased incidence of in situ as well as microinvasive carcinoma. The enhanced pathological examination to which sentinel lymph nodes are subjected has led to an increased rate of detection of micrometastatic carcinoma. Despite the augmented rate of diagnoses of both diseases, the pathological diagnoses as well as clinical management of these entities continue to be controversial. DATA SOURCES: A computerized literature search was performed on the Medline and PubMed database from 1990 to date. Relevant earlier publications were also perused. The database of the Department of Pathology at New York Presbyterian Hospital-Well Medical College of Cornell University were also accessed. CONCLUSIONS: Based on cumulative data, patients diagnosed with either microinvasive or micrometastatic carcinoma of breast have a relatively favorable, albeit guarded, prognosis. Treatment recommendations for both of these disease entities continue to be controversial, and may remain so until additional refined clinicopathological data becomes available. PMID- 11113442 TI - Bone marrow micrometastases in breast cancer patients. AB - BACKGROUND: The status of the axillary nodes has historically been the most important indicator of prognosis in breast cancer patients. However, approximately one third of node-negative patients recur with systemic disease. The detection of bone marrow micrometastases (BMM) may represent additional information in predicting distant recurrence and survival. METHODS: Bone marrow aspiration is obtained from the patient's anterior iliac crest at the time of breast cancer surgery. Cytospins are prepared from this aspirate and stained for polymerase chain reaction (PCR) analysis. RESULTS: Multiple studies have evaluated the clinical implications of BMM. The majority of studies have found a significant correlation between the presence of BMM and decreased survival. The information of bone marrow status may serve as a complement to axillary lymph node status in assessing the prognosis of breast cancer patients. CONCLUSIONS: There is a strong correlation between the presence of bone marrow micrometastases and poorer survival. These results may have an impact upon therapeutic recommendations in breast cancer patients. PMID- 11113443 TI - Is surgical excision necessary for atypical ductal hyperplasia of the breast diagnosed by Mammotome? AB - BACKGROUND: Core biopsy findings of atypical ductal hyperplasia (ADH) underestimates the diagnosis of malignancy by 18% to 88%. Using the Mammotome biopsy technique, more accurate assessment of the lesion is possible, making selective excision of these lesions a consideration. METHODS: The records of 62 patients who were found to have ADH at Mammotome biopsy and subsequently underwent excision of the lesion were reviewed. Patient data were statistically analyzed for predictors of malignancy at the time of surgical excision. RESULTS: Of the 62 patients, 9 (15%) had malignancy at excision. Variables predicting for malignancy included markedly atypical hyperplasia and incomplete removal of calcifications at Mammotome biopsy, a previous contralateral breast cancer, and a family history of breast cancer, with a combined sensitivity of 100% and specificity of 80%. CONCLUSIONS: Mild ADH found on Mammotome, not associated with a personal or family history of breast cancer, may not need excision if all calcifications have been removed. PMID- 11113444 TI - The effects of cysteine to alanine mutations of CD18 on the expression and adhesion of the CD11/CD18 integrins. AB - Of the 56 cysteines in the extracellular domain of the CD18 antigen (beta2 integrin subunit), corresponding ones are not found in 12 positions in the beta4, beta7, or beta8 integrin subunits. These 12 cysteines were mutated to alanines, either singly or in pairs, in CD18. All these mutants can support the expression of all three CD11/CD18 integrins. Transfectants expressing these variant integrins are generally more adhesive than the wild-type, suggesting that the cysteine residues, perhaps by engaging in disulphide bonds, may contribute to the maintenance of the CD11/CD18 integrins in a resting state. PMID- 11113445 TI - GORK, a delayed outward rectifier expressed in guard cells of Arabidopsis thaliana, is a K(+)-selective, K(+)-sensing ion channel. AB - Here we report on the molecular identification, guard cell expression and functional characterization of AtGORK, an Arabidopsis thaliana guard cell outward rectifying K(+) channel. GORK represents a new member of the plant Shaker K(+) channel superfamily. When heterologously expressed in Xenopus oocytes the gene product of GORK mediated depolarization-activated K(+) currents. In agreement with the delayed outward rectifier in intact guard cells and protoplasts thereof, GORK is activated in a voltage- and potassium-dependent manner. Furthermore, the single channel conductance and regulation of GORK in response to pH changes resembles the biophysical properties of the guard cell delayed outward rectifier. Thus GORK very likely represents the molecular entity for depolarization-induced potassium release from guard cells. PMID- 11113446 TI - Disruption of SMN function by ectopic expression of the human SMN gene in Drosophila. AB - Spinal muscular atrophy is a neurodegenerative disorder caused by mutations or deletions in the survival motor neuron (SMN) gene. We have cloned the Drosophila ortholog of SMN (DmSMN) and disrupted its function by ectopically expressing human SMN. This leads to pupal lethality caused by a dominant-negative effect, whereby human SMN may bind endogenous DmSMN resulting in non-functional DmSMN/human SMN hetero-complexes. Ectopic expression of truncated versions of DmSMN and yeast two-hybrid analysis show that the C-terminus of SMN is necessary and sufficient to replicate this effect. We have therefore generated a system which can be utilized to carry out suppressor and high-throughput screens, and provided in vivo evidence for the importance of SMN oligomerization for SMN function at the level of an organism as a whole. PMID- 11113447 TI - Rice 1Cys-peroxiredoxin over-expressed in transgenic tobacco does not maintain dormancy but enhances antioxidant activity. AB - Possible functions that have been proposed for the plant 1Cys-peroxiredoxin, include activity as a dormancy regulator and as an antioxidant. The transcript level of rice 1Cys-peroxiredoxin (R1C-Prx) rapidly decreased after imbibition of rice seeds, but the protein was detected for 15 days after imbibition. To investigate the function of this protein, we generated transgenic tobacco plants constitutively expressing the R1C-Prx gene. The transgenic R1C-Prx plants showed a germination frequency similar to control plants. However, the transgenic lines exhibited higher resistance against oxidative stress, suggesting that antioxidant activity may be its primary function. PMID- 11113448 TI - Domain architecture of a Caenorhabditis elegans AKAP suggests a novel AKAP function. AB - A-kinase anchoring proteins (AKAPs) are adapter proteins that are involved in directing cAMP-dependent protein kinase and some other signaling enzymes to certain intracellular locations. In this study, we investigate the domain architecture of an AKAP from Caenorhabditis elegans (AKAP(CE)). We show that AKAP(CE) shares two domains with the Smad anchor for receptor activation, a FYVE finger and a transforming growth factor beta (TGFbeta) receptor binding domain, suggesting that AKAP(CE) may interact with a receptor belonging to the TGFbeta receptor family. This predicted novel AKAP function supports the recent view of AKAPs as adapter proteins that can be involved in various signaling pathways. PMID- 11113449 TI - Two novel calcium-binding proteins from cytoplasmic granules of the protozoan parasite Entamoeba histolytica. AB - We report on the molecular characterisation of two novel granule proteins of the protozoon and human pathogen Entamoeba histolytica. The proteins, which were named grainin 1 and 2, show a considerable structural similarity to calcium binding proteins, particularly within EF-hand motifs. Each grainin possesses three of these putative calcium-binding sites. Based on careful inspection of known structures of protein families containing EF-hands, a domain of grainin 1 covering two EF-hand motifs was modeled by homology. Calcium-binding activity of grainins was demonstrated by two independent methods. These granule proteins may be implicated in functions vital for the primitive phagocyte and destructive parasite such as control of endocytotic pathways and granule discharge. PMID- 11113450 TI - Tc1, from Tityus cambridgei, is the first member of a new subfamily of scorpion toxin that blocks K(+)-channels. AB - A new peptide, Tc1, containing only 23 amino acids closely packed by three disulfide bridges was isolated from the Amazonian scorpion Tityus cambridgei. It blocks reversibly the Shaker B K(+)-channels with a K(d) of 65 nM and displaces binding of noxiustoxin to mouse brain synaptosome membranes. It is the shortest known peptide from scorpion venom that recognizes K(+)-channels and constitutes a new structural subfamily of toxin, classified as alphaKTx 13.1. PMID- 11113451 TI - Immature human NT2 cells grafted into mouse brain differentiate into neuronal and glial cell types. AB - NT2 cells are a transfectable human embryonal carcinoma cell line, that can be differentiated into postmitotic neuron-like cells (NT2N cells), and transplanted into rodent brains. Differentiation requires a 5-week-long treatment with retinoic acid prior to transplantation. Here, we show that this step can be omitted, and that undifferentiated NT2 cells migrate over long distances and differentiate into both neuron- and oligodendrocyte-like cell types upon grafting into brains of immunocompetent newborn mice. Grafted cells can be traced by fluorogold, with no evidence for tumor formation. Our approach provides an experimental model system which allows the immunohistological and biochemical study of neuronal and glial differentiation of human cells in vivo, and which may be suitable as an in vivo model for pharmacological studies. PMID- 11113452 TI - Inhibition of Mycobacterium smegmatis topoisomerase I by specific oligonucleotides. AB - DNA topoisomerase I from Mycobacterium smegmatis unlike many other type I topoisomerases is a site specific DNA binding protein. We have investigated the sequence specific DNA binding characteristics of the enzyme using specific oligonucleotides of varied length. DNA binding, oligonucleotide competition and covalent complex assays show that the substrate length requirement for interaction is much longer ( approximately 20 nucleotides) in contrast to short length substrates (eight nucleotides) reported for Escherichia coli topoisomerase I and III. P1 nuclease and KMnO(4) footprinting experiments indicate a large protected region spanning about 20 nucleotides upstream and 2-3 nucleotides downstream of the cleavage site. Binding characteristics indicate that the enzyme interacts efficiently with both single-stranded and double-stranded substrates containing strong topoisomerase I sites (STS), a unique property not shared by any other type I topoisomerase. The oligonucleotides containing STS effectively inhibit the M. smegmatis topoisomerase I DNA relaxation activity. PMID- 11113453 TI - Cycloamylose as an efficient artificial chaperone for protein refolding. AB - High molecular weight cyclic alpha-1,4-glucan (referred to as cycloamylose) exhibited an artificial chaperone property toward three enzymes in different categories. The inclusion properties of cycloamylose effectively accommodated detergents, which keep the chemically denatured enzymes from aggregation, and promoted proper protein folding. Chemically denatured citrate synthase was refolded and completely recovered it's enzymatic activity after dilution with polyoxyethylenesorbitan buffer followed by cycloamylose treatment. The refolding was completed within 2 h, and the activity of the refolded citrate synthase was quite stable. Cycloamylose also promoted the refolding of denatured carbonic anhydrase B and denatured lysozyme of a reduced form. PMID- 11113454 TI - Serotonin transporter phosphorylation modulated by tetanus toxin. AB - Tetanus toxin (TeTx) modifies Na(+)-dependent, high-affinity 5-hydroxytryptamine (5-HT, serotonin) uptake in a synaptosomal-enriched P(2) fraction from rat brain. The effect corresponds to a rapid and non-competitive uptake inhibition, and it is preceded by induction of phospholipase C (PLC) activity and translocation and down-regulation of the classical protein kinase C (PKC-alpha, -beta and -gamma) isoforms. The effects on serotonin transport and on cPKC activation were similar to the effects exhibited by phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA). Moreover, after treatment with TeTx, an increase in Ser- and Tyr-specific phosphorylation was found. Activation of PKC by both TeTx and TPA results in a loss of transport capacity and serotonin transporter (SERT) phosphorylation, which are abolished by coapplication of the specific PKC inhibitor bisindolylmaleimide-1. Since a specific PLCgamma1 phosphorylation prior to TeTx's inducing SERT phosphorylation was found, the studies suggest that part of the action of TeTx consists of modifying the signal cascade initiated in tyrosine kinase receptors on nerve tissue previous to its cellular internalization, resulting in transporter phosphorylation. PMID- 11113455 TI - Cross-talk between signal transducer and activator of transcription 3 and estrogen receptor signaling. AB - Interleukin-6 (IL-6) is a multifunctional cytokine that plays important roles in the immune system, hematopoiesis, and acute phase reactions. Estrogens have significant roles in a variety of biological events, such as the development and maintenance of female reproductive organs, and bone and lipid metabolism. Previous studies demonstrated that estrogens suppress IL-6-induced osteoporosis and the growth of multiple myeloma cells by repressing IL-6 and IL-6 receptor gene expression. Here we present a novel mechanism for the inhibitory effect of estrogens on IL-6 function. IL-6-induced activation of signal transducer and activator of transcription 3 (STAT3) activity and STAT3-mediated gene expression were suppressed by 17beta-estradiol (E2) in breast cancer cells. E2-mediated inhibition of STAT3 activation was reversed by tamoxifen, an estrogen receptor (ER) antagonist. We provide evidence that the inhibitory action of ER on STAT3 activity was due to direct physical interactions between STAT3 and ER which represents a novel form of cross-talk between STAT3 and ER signaling pathways. PMID- 11113456 TI - PA-FABP, a novel marker of human epidermal transit amplifying cells revealed by 2D protein gel electrophoresis and cDNA array hybridisation. AB - Human epidermal stem cells express higher levels of beta1 integrins than their more differentiated daughters, transit amplifying cells. In a search for additional stem and transit cell markers we used proteomics and differential cDNA hybridisation to compare keratinocytes fractionated on the basis of beta1 integrin expression. There were remarkably few differences between the two populations and none of the RNAs differed in abundance by more than 2-fold. Nevertheless, proteomics revealed upregulated expression of epidermal fatty acid binding protein (PA-FABP, also known as E-FABP), Annexin II and two keratin related proteins in the transit population. An unknown high molecular mass protein was upregulated in the stem cell population. The upregulation of PA-FABP was confirmed by Northern blotting and conventional and whole mount labelling of human epidermis. We conclude that PA-FABP is a novel marker of epidermal transit amplifying cells. PMID- 11113457 TI - Upregulation of vacuolar H(+)-translocating pyrophosphatase by phosphate starvation of Brassica napus (rapeseed) suspension cell cultures. AB - The influence of phosphate (Pi) deprivation on the vacuolar H(+)-translocating pyrophosphatase (PPiase) and ATPase in tonoplast vesicles from Brassica napus suspension cells was assessed. Pi starvation significantly elevated the ratios of PPi-:ATP-dependent H(+) translocation rate and H(+)-PPiase:H(+)-ATPase hydrolytic activities. These increases were reversed 36 h following resupply of 2.5 mM Pi to the Pi-starved cells. Immunoblotting indicated that Pi starvation also induced a two-fold increase in the amount of H(+)-PPiase protein, whereas the amount of H(+)-ATPase remained unchanged. It is proposed that H(+)-PPiase facilitates the conservation of limited ATP pools, and Pi recycling during Pi stress. PMID- 11113458 TI - Effects of homocysteine on the binding of extracellular-superoxide dismutase to the endothelial cell surface. AB - Homocysteine is known to be a risk factor for several vascular diseases. Previously, we found a significant association between plasma homocysteine and plasma extracellular-superoxide dismutase (EC-SOD) levels. The binding of EC-SOD to human and bovine aortic endothelial cell cultures showed significant decreases after incubation with 10 microM homocysteine, whereas the expression of EC-SOD in fibroblast cell cultures was inhibited with a high concentration (1 mM) of homocysteine. Furthermore, binding of EC-SOD to heparin immobilized on plates was decreased with homocysteine. These observations suggested that homocysteine decreases the binding of EC-SOD to vascular endothelial cell surfaces by degradation of endothelial heparan sulfate proteoglycan, which results in a loss of the ability to protect endothelial cell surfaces from oxidative stress. PMID- 11113460 TI - Genetics and evolution of ultraviolet vision in vertebrates. AB - Various vertebrates use ultraviolet (UV) vision for such basic behaviors as mating, foraging, and predation. We have successfully interchanged the color sensitivities of the mouse UV pigment and the human blue pigment by introducing forward and reverse mutations at five sites. This unveils for the first time the general mechanism of UV vision. Most contemporary UV pigments in vertebrates have maintained their ancestral functions by accumulating no more than one of the five specific amino acid changes. The avian lineage is an exception, where the ancestral pigment lost UV-sensitivity but some descendants regained it by one amino acid replacement at an entirely different site. PMID- 11113459 TI - Location of the epoxide function determines specificity of the allelic variants of human glutathione transferase Pi toward benzo[c]chrysene diol epoxide isomers. AB - Carcinogenic activity of many polycyclic aromatic hydrocarbons (PAHs) is mainly attributed to their respective diol epoxides, which can be classified as either bay or fjord region depending upon the location of the epoxide function. The Pi class human glutathione (GSH) transferase (hGSTP1-1), which is polymorphic in humans with respect to amino acid residues in positions 104 (isoleucine or valine) and/or 113 (alanine or valine), plays an important role in the detoxification of PAH-diol epoxides. Here, we report that the location of the epoxide function determines specificity of allelic variants of hGSTP1-1 toward racemic anti-diol epoxide isomers of benzo[c]chrysene (B[c]C). The catalytic efficiency (k(cat)/K(m)) of V104,A113 (VA) and V104,V113 (VV) variants of hGSTP1 1 was approximately 2.3- and 1.7-fold higher, respectively, than that of the I104,A113 (IA) isoform toward bay region isomer (+/-)-anti-B[c]C-1,2-diol-3,4 epoxide. On the other hand, the IA variant was approximately 1.6- and 3.5-fold more efficient than VA and VV isoforms, respectively, in catalyzing the GSH conjugation of fjord region isomer (+/-)-anti-B[c]C-9,10-diol-11,12-epoxide. The results of the present study clearly indicate that the location of the epoxide function determines specificity of the allelic variants of hGSTP1-1 in the GSH conjugation of activated diol epoxide isomers of B[c]C. PMID- 11113461 TI - Rational re-design of the substrate binding site of flavocytochrome P450 BM3. AB - Bacillus megaterium P450 BM3 is a fatty acid hydroxylase with selectivity for long chain substrates (C(12)-C(20)). Binding or activity with substrates of chain length 13-fold with butyrate, while the L75T/L181K double mutant has k(cat)/K(M) increased >15-fold with hexanoate and binding (K(d)) improved >28 fold for butyrate. Removing the arginine 47/lysine 51 carboxylate binding motif at the mouth of the active site disfavours binding of all fatty acids, indicating its importance in the initial recognition of substrates. PMID- 11113462 TI - The asp-rich region at the carboxyl-terminus of calsequestrin binds to Ca(2+) and interacts with triadin. AB - Calsequestrin (CSQ) is a high capacity Ca(2+) binding protein in the junctional sarcoplasmic reticulum of striated muscles, and has been shown to regulate the ryanodine receptor (RyR) through triadin and junctin. In order to identify the functional roles of specific regions on CSQ, several CSQ deletion mutants were prepared by molecular cloning and Escherichia coli expression. 45Ca(2+) overlay assay using a native gel system revealed that the major Ca(2+) binding motif of CSQ resides in the asp-rich region (amino acids 354-367). In an in vitro binding assay using a glutathione-S-transferase affinity column, the interaction between CSQ and triadin was found to be Ca(2+)-dependent, and the site of interaction was confined to the asp-rich region of CSQ. Our results suggest that the asp-rich region of CSQ could participate in the RyR-mediated Ca(2+) release process by offering a direct binding site to luminal Ca(2+) as well as triadin. PMID- 11113463 TI - Editorial PMID- 11113464 TI - Strategies and testing methods for identifying mutagenic risks. AB - The evolution of testing strategies and methods for identification of mutagenic agents is discussed, beginning with the concern over potential health and population effects of chemical mutagens in the late 1940s that led to the development of regulatory guidelines for mutagenicity testing in the 1970s and 1980s. Efforts to achieve international harmonization of mutagenicity testing guidelines are summarized, and current issues and needs in the field are discussed, including the need for quantitative methods of mutagenic risk assessment, dose-response thresholds, indirect mechanisms of mutagenicity, and the predictivity of mutagenicity assays for carcinogenicity in vivo. Speculation is offered about the future of mutagenicity testing, including possible near-term changes in standard test batteries and the longer-term roles of expression profiling of damage-response genes, in vivo mutagenicity testing methods, and models that better account for differences in metabolism between humans and laboratory model systems. PMID- 11113465 TI - Practical aspects of mutagenicity testing strategy: an industrial perspective. AB - Genetic toxicology studies play a central role in the development and marketing of new chemicals for pharmaceutical, agricultural, industrial, and consumer use. During the discovery phase of product development, rapid screening tests that require minimal amounts of test materials are used to assist in the design and prioritization of new molecules. At this stage, a modified Salmonella reverse mutation assay and an in vitro micronucleus test with mammalian cell culture are frequently used for screening. Regulatory genetic toxicology studies are conducted with a short list of compounds using protocols that conform to various international guidelines. A set of four assays usually constitutes the minimum test battery that satisfies global requirements. This set includes a bacterial reverse mutation assay, an in vitro cytogenetic test with mammalian cell culture, an in vitro gene mutation assay in mammalian cell cultures, and an in vivo rodent bone marrow micronucleus test. Supplementary studies are conducted in certain instances either as a follow-up to the findings from this initial testing battery and/or to satisfy a regulatory requirement. Currently available genetic toxicology assays have helped the scientific and industrial community over the past several decades in evaluating the mutagenic potential of chemical agents. The emerging field of toxicogenomics has the potential to redefine our ability to study the response of cells to genetic damage and hence our ability to study threshold phenomenon. PMID- 11113466 TI - The Ames Salmonella/microsome mutagenicity assay. AB - The Ames Salmonella/microsome mutagenicity assay (Salmonella test; Ames test) is a short-term bacterial reverse mutation assay specifically designed to detect a wide range of chemical substances that can produce genetic damage that leads to gene mutations. The test employs several histidine dependent Salmonella strains each carrying different mutations in various genes in the histidine operon. These mutations act as hot spots for mutagens that cause DNA damage via different mechanisms. When the Salmonella tester strains are grown on a minimal media agar plate containing a trace of histidine, only those bacteria that revert to histidine independence (his(+)) are able to form colonies. The number of spontaneously induced revertant colonies per plate is relatively constant. However, when a mutagen is added to the plate, the number of revertant colonies per plate is increased, usually in a dose-related manner. The Ames test is used world-wide as an initial screen to determine the mutagenic potential of new chemicals and drugs. The test is also used for submission of data to regulatory agencies for registration or acceptance of many chemicals, including drugs and biocides. International guidelines have been developed for use by corporations and testing laboratories to ensure uniformity of testing procedures. This review provides historical aspects of how the Ames was developed and detailed procedures for performing the test, including the design and interpretation of results. PMID- 11113467 TI - The bacterial tryptophan reverse mutation assay with Escherichia coli WP2. AB - The Escherichia coli WP2 tryptophan reverse mutation assay detects trp(-) to trp(+) reversion at a site blocking a step in the biosynthesis of tryptophan prior to the formation of anthranilic acid. The different WP2 strains all carry the same AT base pair at the critical mutation site within the trpE gene. The assay is currently used by many laboratories in conjunction with the Ames Salmonella assay for screening chemicals for mutagenic activity. In general the WP2 strains are used as a substitute for, or as an addition to Salmonella strain TA102 which also carries an AT base pair at the mutation site. The assay is also recommended together with the Ames assay for data submission to regulatory agencies. National and international guidelines have been established for performing these mutagenicity assays. The E. coli WP2 assay procedures are the same as those described elsewhere in this volume for the Ames Salmonella assay (Mortelmans and Zeiger, 2000) with the exception that limited tryptophan instead of limited histidine is used. This chapter is an addendum to the previous chapter and the reader should refer to the previous chapter for details regarding experimental procedures and assay design. PMID- 11113468 TI - The Escherichia coli lacZ reversion mutagenicity assay. AB - The Escherichia coli lacZ reversion assay, based on the set of episomal lacZ alleles engineered by Miller et al., provides an attractive system for studies of mutagenesis and mutational specificity. Each strain in the lacZ set reverts by a specific base substitution or frameshift event. Revertants are selected by growth on lactose minimal medium. In this review, I describe the development of the assay and its subsequent modifications and improvements. Examples of its application are presented and detailed protocols for the implementation of the assay are given. PMID- 11113469 TI - The in vitro micronucleus technique. AB - The study of DNA damage at the chromosome level is an essential part of genetic toxicology because chromosomal mutation is an important event in carcinogenesis. The micronucleus assays have emerged as one of the preferred methods for assessing chromosome damage because they enable both chromosome loss and chromosome breakage to be measured reliably. Because micronuclei can only be expressed in cells that complete nuclear division a special method was developed that identifies such cells by their binucleate appearance when blocked from performing cytokinesis by cytochalasin-B (Cyt-B), a microfilament-assembly inhibitor. The cytokinesis-block micronucleus (CBMN) assay allows better precision because the data obtained are not confounded by altered cell division kinetics caused by cytotoxicity of agents tested or sub-optimal cell culture conditions. The method is now applied to various cell types for population monitoring of genetic damage, screening of chemicals for genotoxic potential and for specific purposes such as the prediction of the radiosensitivity of tumours and the inter-individual variation in radiosensitivity. In its current basic form the CBMN assay can provide, using simple morphological criteria, the following measures of genotoxicity and cytotoxicity: chromosome breakage, chromosome loss, chromosome rearrangement (nucleoplasmic bridges), cell division inhibition, necrosis and apoptosis. The cytosine-arabinoside modification of the CBMN assay allows for measurement of excision repairable lesions. The use of molecular probes enables chromosome loss to be distinguished from chromosome breakage and importantly non-disjunction in non-micronucleated binucleated cells can be efficiently measured. The in vitro CBMN technique, therefore, provides multiple and complementary measures of genotoxicity and cytotoxicity which can be achieved with relative ease within one system. The basic principles and methods (including detailed scoring criteria for all the genotoxicity and cytotoxicity end-points) of the CBMN assay are described and areas for future development identified. PMID- 11113470 TI - The mouse lymphoma assay. AB - In this paper, the current status of the protocol for the Mouse Lymphoma Assay is discussed. A brief history describes the events leading to current protocol recommendations. Areas for further development such as cytotoxicity, 24-h treatments, acceptability criteria and statistical analysis are also considered. Recent guidelines are reviewed, and consensus issues from the Mouse Lymphoma workgroup assembled as part of the International Workshop on Genotoxicity Test Procedures (IWGTP) are included. There are two versions of the assay - soft agar and microwell - and both will be discussed. For assay procedures, the emphasis will be on a typical microwell protocol but an attempt will be made to highlight protocol variations between laboratories and between the microwell and agar versions of the assay. PMID- 11113471 TI - Microgels for estimation of DNA strand breaks, DNA protein crosslinks and apoptosis. AB - This report describes a part of the evolution of microgel electrophoresis in the author's laboratory for the last 15 years. It also describes the importance of estimation of DNA single and double strand breaks, DNA crosslinks and apoptosis. Some experiments based on each methodology are included here. A new protocol for rapid and efficient precipitation of DNA in microgel is included. A step by step description of laboratory protocol is also included. PMID- 11113472 TI - A refined protocol for conducting the low pH 6.7 Syrian hamster embryo (SHE) cell transformation assay. AB - The Syrian hamster embryo (SHE) cell transformation assay evaluates the potential of chemicals to induce morphological transformation in karyotypically normal primary cells. Induction of transformation has been shown to correlate well with the carcinogenicity of many compounds in the rodent bioassay. Historically the assay has not received wide-spread use due to technical difficulty. An improved protocol for a low pH 6.7 assay was developed by LeBoeuf et al. [R.A. LeBoeuf, G.A. Kerckaert, M.J. Aardema, D.P. Gibson, R. Brauninger, R.J. Isfort, Mutat. Res., 356 (1996) 85-127], that greatly reduced many of the technical difficulties associated with the SHE assay. The purpose of this paper is to describe the most current execution of the pH 6.70 protocol including protocol refinements made since the publication of a comprehensive protocol for this assay in Kerckaert et al. [G.A. Kerckaert, R.J. Isfort, G.J. Carr, M.J. Aardema, Mutat. Res., 356 (1996) 65-84]. PMID- 11113473 TI - Parallel monitoring of mitotic recombination, clastogenicity and teratogenic effects in eye tissue of Drosophila. AB - Loss of heterozygosity (LOH) of the wild-type allele by structural chromosome aberrations (SCAs), homologous mitotic recombination (HMR) or intra-chromosomal (deletion/amplification) recombination (ICR) plays a crucial role in multistage carcinogenesis. We describe here an in vivo system, enabling the detection of all three chromosome breakage-related events in the same genetic experiment, with eye tissue of Drosophila as targets. This modification of the white/white(+) system enables to measure, simultaneously, HMR and ICR on the X-chromosome, and loss of a ring-shaped X-chromosome, utilizing the eye color gene white. Optimal conditions for the detection and quantification of SCAs (ring-X loss) compared to HMR are discussed in detail. Emerging new techniques comprise the parallel detection of HMR on chromosomes X and 3, using the tumor suppressor gene warts in addition to the X-linked marker white. Another modification of the white/white(+) system measures, again in parallel, HMR and chromosome duplication (non disjunction). PMID- 11113474 TI - In vivo rodent micronucleus assay: protocol, conduct and data interpretation. AB - In vivo rodent micronucleus assay has been widely used to detect genotoxicity. Evaluation of micronucleus induction is the primary in vivo test in a battery of genotoxicity tests and is recommended by the regulatory agencies around the globe to be conducted as part of product safety assessment. The assay, when performed appropriately, detects both clastogenicity and aneugenicity. Methods for performing micronucleus evaluation have evolved since its initial description in the 1970s. In recent years, the focus has been directed toward improving micronucleus detection with high efficiency by proposing data-based recommendations to the standard initial protocol design. Such improvements include, e.g., the use of appropriate harvest time(s), inclusion of one or both sexes, number of doses tested, limit dose, integrating micronucleus assessment into the routine toxicology studies, use of fluorescent staining, automation of micronucleus detection and assessment of micronuclei in multiple tissues. This protocol paper describes: the mechanism of micronucleus formation, a generalized protocol for manual detection, enumeration of micronuclei, and data interpretation in light of published information thus far, on the regulatory aspects of this assay. Certain recent protocol issues that are practical in nature are equally valid in relation to standard manual method and provide robust database, which are also included for consideration. It is expected that such improvements of the protocol will continue to drive the utility of this assay in the product safety assessment. PMID- 11113475 TI - In vivo cytogenetics: mammalian germ cells. AB - This chapter summarizes the most relevant methodologies available for evaluation of cytogenetic damage induced in vivo in mammalian germ cells. Protocols are provided for the following endpoints: numerical and structural chromosome aberrations in secondary oocytes or first-cleavage zygotes, reciprocal translocations in primary spermatocytes, chromosome counting in secondary spermatocytes, numerical and structural chromosome aberrations, and sister chromatid exchanges (SCE) in spermatogonia, micronuclei in early spermatids, aneuploidy in mature sperm. The significance of each methodology is discussed. The contribution of novel molecular cytogenetic approaches to the detection of chromosome damage in rodent germ cells is also considered. PMID- 11113476 TI - Recent advances in the protocols of transgenic mouse mutation assays. AB - Transgenic mutation assays were developed to detect gene mutations in multiple organs of mice or rats. The assays permit (1) quantitative measurements of mutation frequencies in all tissues/organs including germ cells and (2) molecular analysis of induced and spontaneous mutations by DNA sequencing analysis. The protocols of recently developed selections in the lambda phage-based transgenic mutation assays, i.e. cII, Spi(-) and 6-thioguanine selections, are described, and a data set of transgenic mutation assays, including those using Big Blue and Muta Mouse, is presented. PMID- 11113477 TI - The effects of neonatal ethanol and/or cocaine exposure on isolation-induced ultrasonic vocalizations. AB - Isolation-induced ultrasonic vocalizations (USVs) are emitted by young rat pups when isolated from their dam and conspecifics. These USVs play an important role in maternal/offspring interactions, and have been used as an indicator of response to stress and isolation. This study examined the effects of neonatal ethanol and/or cocaine exposure on USVs in neonatal rats. The neonatal exposure paradigm serves as a model for the "human third trimester of pregnancy" in terms of CNS development. There were five treatment groups including an artificially reared (AR) ethanol-exposed group (6 g/kg/day), an AR cocaine-exposed group (60 mg/kg/day), an AR ethanol- and cocaine-exposed group (6 g/kg/day+60 mg/kg/day), an AR isocaloric control, and a normally reared control. Both groups that received ethanol took longer to vocalize, and displayed fewer vocalizations than non-ethanol-exposed pups when tested on clean bedding (Experiment 1) or on chips from the nest of a lactating dam (Experiment 2). These results suggest that neonatal ethanol exposure alters the pup's immediate response to isolation. This could have direct effects on maternal/infant interactions, and might help explain some of the long-term effects of ethanol exposure on social behaviors. PMID- 11113478 TI - 7-OH-DPAT, unlike quinpirole, does not prime a yawning response in rats. AB - Repeated treatment in ontogeny with the dopamine (DA) D(2)/D(3) receptor agonist quinpirole is associated with enhanced quinpirole-induced yawning and other behaviors such as vacuous chewing, vertical jumping, and antinociception. To determine if the reputedly DA D(3) agonist (+/-)-2-(dipropylamino)-7-hydroxy 1,2,3, 4-tetrahydronaphthalene (7-OH-DPAT) would prime for yawning in a manner analogous to that for quinpirole, rats were treated for the first 11 days after birth with an equimolar dose of either quinpirole or 7-OH-DPAT (195.4 nmol/kg/day) and tested for agonist-induced yawning in adulthood. While enhanced quinpirole-induced and 7-OH-DPAT-induced yawning was observed in quinpirole primed rats, acute treatments with quinpirole and 7-OH-DPAT did not produce an enhanced yawing response in 7-OH-DPAT-"primed" rats. Our findings indicate that 7 OH-DPAT, unlike quinpirole, does not prime for quinpirole- or 7-OH-DPAT-induced yawning in rats. PMID- 11113479 TI - DBA/2J mice develop stronger lithium chloride-induced conditioned taste and place aversions than C57BL/6J mice. AB - Genetic differences in lithium-induced conditioned aversion were examined using both place- and taste-conditioning procedures. In the place-conditioning procedure, adult male C57BL/6J (B6) and DBA/2J (D2) mice were exposed to a differential conditioning procedure in which each mouse received four 30-min pairings of a distinctive floor cue immediately after IP injections of either 0.75, 1.5, or 3. 0 mEq/kg LiCl. A different floor cue was paired with saline injections. A separate group of control mice received saline injections paired with both floor types. Subsequent floor preference testing revealed greater conditioned aversion in D2 mice compared to B6 mice in groups receiving 3.0 mEq/kg LiCl. Lower LiCl doses did not produce conditioning in either strain. In a conditioned taste-aversion procedure, fluid-restricted mice received four trials in which access to 0.2 M NaCl solution was followed by IP injection of either 0.75, 1.5, 3.0, or 6.0 mEq/kg LiCl. D2 mice showed stronger conditioned taste aversion than B6 mice at all doses, suggesting that taste conditioning may be a more sensitive index of aversive drug sensitivity than place conditioning. These findings are not well explained by strain differences in general learning ability or by strain differences in stimulus salience or innate preference. Rather, these data appear more consistent with previous studies showing strain differences in lithium pharmacokinetics and in general sensitivity to aversive events. PMID- 11113480 TI - Plasma concentrations of beta-endorphin in smokers who consume different numbers of cigarettes per day. AB - The harmful effects of smoking on health have been widely documented, although it is as yet unclear whether tobacco dependence is only psychological in nature, or both psychological and physical. We studied plasma concentrations of beta endorphin, cortisol, and adrenocorticotropic hormone (ACTH) in healthy persons who consumed different numbers of cigarettes per day, and compared the findings with those in a control group of nonsmokers. Beta-endorphin levels were significantly higher than in controls only in persons who smoked fewer than 10 cigarettes per day. Cortisol levels were significantly higher in smokers who consumed more than 20 cigarettes per day. There were no significant differences between any of the groups in plasma ACTH concentrations. PMID- 11113481 TI - Neurosteroids and reward: allopregnanolone produces a conditioned place aversion in rats. AB - The neurosteroid 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone) has been reported to have rewarding properties in mice tested for place conditioning. Another study found that allopregnanolone reduced dopamine (DA) output in the nucleus accumbens (NAc) of rats. As many rewarding stimuli increase accumbens DA, these results may appear contradictory. Thus, the present study examined the rewarding properties of allopregnanolone in rats tested for place conditioning using an unbiased conditioning procedure. In control studies, a place preference was observed following conditioning with intraperitoneal (2.0 mg/kg) or intracerebroventricular (i.c.v.) (100 microg/0.5 microl) amphetamine. Conditioning with i.c.v. allopregnanolone produced a significant aversion at a dose of 5.0 microg (in 5.0 microl) and a near aversion at 25.0 microg (in 8.3 microl); doses of 0 microg (i.e., vehicle alone, in 10 microl) or 30.0 microg (in 10 microl) produced little effect on place preference. During conditioning, locomotor activity was stimulated by amphetamine using either route of administration, but allopregnanolone had no significant main effect on locomotor activity. Thus, there was a dissociation between the effects of drugs on locomotor activity vs. place conditioning. Results show that i.c.v. amphetamine produces a place preference, whereas allopregnanolone produces either no effect or an aversion, depending on the dose. PMID- 11113482 TI - Behavioral and neural toxicity of the artemisinin antimalarial, arteether, but not artesunate and artelinate, in rats. AB - Three artemisinin antimalarials, arteether (AE), artesunate (AS), and artelinate (AL) were evaluated in rats using an auditory discrimination task (ADT) and neurohistology. After rats were trained on the ADT, equimolar doses of AE (25 mg/kg, in sesame oil, n=6), AS (31 mg/kg, in sodium carbonate, n=6), and AL (36 mg/kg, in saline, n=6), or vehicle (sodium carbonate, n=6) were administered (IM) for 7 consecutive days. Behavioral performance was evaluated, during daily sessions, before, during, and after administration. Histological evaluation of the brains was performed using thionine staining, and damaged cells were counted in specific brainstem nuclei of all rats. Behavioral performance was not significantly affected in any rats treated with AS, AL, or vehicle. Furthermore, histological examination of the brains of rats treated with AS, AL, and vehicle did not show damage. In stark contrast, all rats treated with AE showed a progressive and severe decline in performance on the ADT. The deficit was characterized by decreases in accuracy, increases in response time and, eventually, response suppression. When performance on the ADT was suppressed, rats also showed gross behavioral signs of toxicity that included tremor, gait disturbances, and lethargy. Subsequent histological assessment of AE-treated rats revealed marked damage in the brainstem nuclei, ruber, superior olive, trapezoideus, and inferior vestibular. The damage included chromatolysis, necrosis, and gliosis. These results demonstrate distinct differences in the ability of artemisinins to produce neurotoxicity. Further research is needed to uncover pharmacokinetic and metabolic differences in artemisinins that may predict neurotoxic potential. PMID- 11113483 TI - The influence of buspirone, and its metabolite 1-PP, on the activity of paroxetine in the mouse light/dark paradigm and four plates test. AB - Although numerous animal procedures have been employed in the study of antidepressants (ADs) in anxiety, the results following acute administration remain highly variable. The present study investigated the effect of the SSRI paroxetine (4, 8, and 16 mg/kg, IP) in two tests of anxiety in mice: the light/dark test paradigm, and the four plates test (FPT). In both tests, it was found that paroxetine resulted in an anxiolytic-like effect at doses that did not modify motor performance (at the doses of 4 and 8 mg/kg in the light/dark test and at the doses of 4, 8, and 16 mg/kg in the four plates test). In the light/dark paradigm, both doses of buspirone significantly potentiated paroxetine, while in the four plates only one dose of buspirone (a 5HT(1A) partial agonist) (0.06 mg/kg) increased the anxiolytic-like effect of paroxetine. Prior administration of 1-PP was without effect in the light/dark paradigm but antagonized the effect of paroxetine (at the dose of 0.06 and 0. 5 mg/kg) in the FPT. The results suggested that a balance between pre- and postsynaptic 5-HT(1A) receptor was implicated in the anxiolytic-like effect of paroxetine. Buspirone seemed to emphasize the role of paroxetine in 5-HT(1A) receptor modulation and exerted a biphasic influence in the two tests. PMID- 11113484 TI - A9 and A10 dopamine nuclei as a site of action for effects of 8-OH-DPAT on locomotion in the rat. AB - The 5-hydroxytryptamine (5-HT) 5-HT(1A) receptor agonist 8-hydroxy-2-(di-n propylamino)tetralin (8-OH-DPAT) was applied locally (0-5 microg bilaterally) into either the substantia nigra (A9) or the ventral tegmental area (A10) of adult male Wistar rats, and 10 min later spontaneous motor activity was observed in an open field ( approximately 0.5 m(2)) for 30 min. The rate of dopamine synthesis was estimated in neostriatal areas, the amygdala, and the prefrontal cortex, by measuring the accumulation of DOPA, following inhibition of cerebral decarboxylase by means of 3-hydroxybenzylhydrazine (NSD-1015). The A10 application of 8-OH-DPAT resulted in an increase in all aspects of spontaneous motor activity in the open field. A9 application of 8-OH-DPAT produced a stereotyped forward locomotion, characterized by a modest decrease in total horizontal activity, almost complete inhibition of rearing activity and an increase in proportion forward locomotion along the perimeter of the open-field arena. The injection of 8-OH-DPAT into the A9 was accompanied by an increased neostriatal DA rate of synthesis, whereas the A10 injection was followed by a decreased DA rate of synthesis in the amygdala and in the prefrontal cortex. It is concluded that mesencephalic dopaminergic mechanisms are involved in the stereotyped forward locomotion characteristically seen after systemic administration of the 5-HT(1A) receptor agonist 8-OH-DPAT. PMID- 11113485 TI - The role of opioid receptors in hypoxic preconditioning against seizures in brain. AB - Preconditioning is defined as an adaptive mechanism produced by short periods of hypoxia/ischemia, resulting in protection against subsequent ischemic insult, and development of seizures. Results of the present study demonstrate that an episode of normobar hypoxia reduces the susceptibility to convulsions induced by pentylenetetrazol (PTZ) 30 min, 24 h, as well as 4 and 7 days later. Administration of morphine showed similar effects after 24 h. Naloxone, given before ischemic preconditioning, as well as morphine, blocked the development of the protection. Administration of D-Ala-Met-enkephalin-Gly-ol (DAMGO - a selective mu-opioid receptor agonist), as well as trans-3, 4-dichloro-N-methyl-N [7-(1-pyrrolidinyl) cycloexilbenzeneacetamide ethane sulfonate] (U-69,593 - a selective kappa-opioid receptor agonist), mimicked the effects of hypoxic preconditioning (HPC). (-)-N-(Cyclopropylmethyl)-4,14-dimethoxymorphinan-6-one (cyprodime - a selective mu-opioid receptor antagonist, as well as nor binaltorphimine dihydrochloride (nor-BNI - selective kappa-opioid receptors antagonist), given before HPC as well as before respective opioid receptor agonists, blocked the development of the protection. This study provides evidence that mu- and kappa-opioid receptors are involved in HPC against seizures in the brain. PMID- 11113486 TI - Dissociating nicotine and nonnicotine components of cigarette smoking. AB - To dissociate the sensorimotor aspects of cigarette smoking from the pharmacologic effects of nicotine, smokers rated the subjective effects of nicotine-containing or denicotinized cigarettes, and intravenous (IV) nicotine or saline infusions. Three groups of participants (n=20 per group) received either: (1) continuous nicotine, (2) pulsed nicotine, or (3) saline. Each group was exposed to an IV condition once while smoking a denicotinized cigarette and once while not smoking, in a 3x2 mixed design. A fourth group (n=20) received saline while smoking their usual brand of cigarette. The dose and rate of nicotine administration were individualized based on previous measures of ad lib smoke intake. Denicotinized cigarette smoke significantly reduced craving and was rated significantly more satisfying and rewarding than the no-smoking conditions. IV nicotine reduced craving for cigarettes, and increased ratings of lightheadedness and dizziness. However, no significant satisfaction or reward was reported after IV nicotine. The combination of IV nicotine and denicotinized cigarette smoke produced effects similar to those of smoking the usual brand of cigarette. The results suggest that sensorimotor factors are critical in mediating the immediate subjective response to smoking, and that the immediate subjective effects of nicotine administered in doses obtained from cigarette smoking are subtle. Thus, addressing smokers' needs for both for the sensorimotor aspects of smoking as well as for the direct CNS effects of nicotine may be critical in enhancing smoking cessation treatment outcome. PMID- 11113487 TI - Non-opioid antinociceptive effects of supraspinal histogranin and related peptides: possible involvement of central dopamine D(2) receptor. AB - The antinociceptive effects of intracerebroventricular (ICV) administration of histogranin (HN) and related peptides were assessed in the mouse writhing and tail-flick assays. In the writhing test, the peptides displayed dose-dependent analgesic effects with an AD(50) of 23.9 nmol/mouse for HN and the following order for other peptides: HN-(7-15)/=AMPA threshold. Type 1 cells were least mature, and Type 4 cells most mature as assessed by cell properties, dendritic arborization, and penetration of dendrites into the molecular layer. Thus NMDA-mediated currents predominate early in GC development as is consistent with their role in processes that determine dentate architecture - neuronal migration, dendritic outgrowth and regression, and synapse stabilization. By analogy with 'silent synapses' (i.e. synapses that contain only NMDA receptors), Type 2 cells are candidate 'silent cells' that may undergo activity-dependent acquisition of functional fast-conducting AMPA receptors with maturation. PMID- 11113510 TI - Sexual dimorphism in number and proportion of neurons in the human median raphe nucleus. AB - The number and proportion of neurons in the median raphe nuclei stained by the Golgy-Cox and Nissl methods was compared in males and females infants. When subjects are matched by age and cause of death the number and proportion of fusiform, ovoid and multipolar cells differs significantly between sexes at different ages. PMID- 11113511 TI - Perinatal exposure to environmental tobacco smoke alters cell signaling in a primate model: autonomic receptors and the control of adenylyl cyclase activity in heart and lung. AB - Perinatal exposure to environmental tobacco smoke (ETS) is known to have adverse effects on respiratory function in conjunction with changes in autonomic responses. In the current study, Rhesus monkeys were exposed to ETS during late gestation and in the early neonatal period. Hearts and lungs were examined for changes in beta-adrenergic and m2-muscarinic cholinergic receptors, and for alterations in adenylyl cyclase activity. Whereas there were no changes in the heart, there was robust induction of adenylyl cyclase in the lung; previous work with prenatal nicotine exposure in rodent models has shown that adenylyl cyclase induction is associated with a shift towards predominance of cholinergic over adrenergic responses. These data indicate that perinatal ETS exposure evokes changes in cells signaling that they are selective for the lung and that may ultimately reflect adverse effects at the level of physiological function. PMID- 11113512 TI - Neuropeptide Y- and somatostatin-immunoreactive axons in the corpus callosum during postnatal development of the rat. AB - Corpus callosum (CC) projections in adult mammals were generally thought to be excitatory and to use excitatory amino acids as their transmitters. Little information has been available about the electrical properties and neurochemical status of developing CC connections. The present study investigated the chemical status of rat CC axons during postnatal development by using antibodies to neuropeptide Y (NPY) and to somatostatin (SOM). Both NPY-immunoreactive (ir) and SOM-ir axons were found in the CC of the rat from newborn through adult; however, the number of SOM-ir CC axons is less than that of NPY-ir CC axons at corresponding ages. The density of both NPY-ir and SOM-ir CC axons initially increased, then peaked, and finally decreased to the mature level. In the adult, only a few NPY-ir and SOM-ir CC axons were found in the CC. These results indicate that many NPY-ir and SOM-ir CC axons are transitory during early postnatal development. The results also suggest that the functions of CC connections in adult mammals may be different from that of developing ones. The present results as well as the previous results demonstrate that both developing and mature CC connections are chemically heterogeneous. PMID- 11113513 TI - Acute hypoxic hypoxia transiently reduces GABA(A) binding site number in developing chick optic lobe. AB - The Central Nervous System is known to be critically affected in the prenatal perinatal period by hypoxic-ischemic insults, which produce several disorders such as loss of neural projections, increased susceptibility to seizures, apoptosis and an imbalance in normal activity of glutamatergic and GABAergic neurones, resulting in acute cell excitotoxicity. The aim of the present work was to establish a chick embryo model of normobaric acute hypoxic hypoxia as well as to evaluate modifications in GABA(A) receptor complex from chick optic lobe, that may result from this injury. Fertile chicken (Gallus gallus domesticus) eggs from White Leghorn were incubated and at embryonic days (ED) 12 to 18, subjected to a stream of 8%O(2)/92%N(2) during1 h, and then were either returned to their shelves in the incubator for recovery, or immediately processed for biochemical studies. Hypoxic treatment produced a significant age dependent reduction in GABA binding sites showing the greatest decrease at the earliest stages studied (ED12 ED16). Saturation curves of GABA binding performed at ED12 showed a decrease in B(max), (control, 5.48+/-0.20, hypoxic, 3.90+/-0.39 pmol/mg protein), but no significant change in K(d). Following 48 h in normoxic atmosphere post-hypoxia reduction in [3H]GABA binding was reversed. Pharmacological properties of GABA(A) receptor at ED12 showed that positive allosteric modulation effects of the steroid 3alpha-hydroxy-5alpha-pregnan-20-one and the barbiturate pentobarbital sodium were enhanced by the treatment. This model of acute prenatal hypoxic hypoxia produced marked alterations in inhibitory CNS neurotransmission that proved reversible and age dependent. PMID- 11113515 TI - Expression of cholinergic system molecules during development of the chick nervous system. AB - There are suggestions of the participation of nicotinic acetylcholine receptors (nAChRs), the acetylcholine degradation enzyme, acetylcholinesterase (AChE), and the acetylcholine synthesizing enzyme, choline acetyltransferase (ChAT), in the development of the nervous system. In this study, we aimed at comparing the development of some subunits of the nAChRs, AChE, and ChAT in the chick nervous system by standard immunohistochemical methods. The expression of all molecules investigated here appeared very early in ganglia (embryonic day 3.5-4), persisting into posthatching, except for ChAT, which is not detected after hatching in ganglia. A differential development was observed for nAChR subunits, with these receptors appearing around embryonic day 6 in some sites. The time course of development of different nAChR subunits revealed several instances of transient expression (such as in the cerebellum), increasing expression (such as in the nucleus spiriformis lateralis), and diminishing expression into posthatching stages (such as in the oculomotor and throclear nuclei). Expression of AChE and ChAT also starts around embryonic day 6 in some structures and follows mainly increasing time-courses in the chick brain. The results of this study reveal a developmentally regulated expression of cholinergic system-related molecules in the chick nervous system and characterize differential time-courses of expression for nAChR subunits, AChE, and ChAT during development. PMID- 11113514 TI - The role of opioid receptors in morphine withdrawal in the infant rat. AB - Exposure to opiates such as morphine can lead to psychological and physical dependence in both adult and infant humans. Infant rats experience opiate withdrawal behaviors that are qualitatively different from the withdrawal behaviors displayed by adult rats. In the adult, withdrawal is largely mediated by the mu-opioid receptor. We sought to understand more about what role each opioid receptor (mu, kappa, and delta) plays in the display of the physical withdrawal in the infant rat. Beginning on postnatal day 1, infant rats were injected with morphine sulfate twice a day for 6.5 days. On the afternoon of the seventh day the infant rats were given an i.c. injection of a vehicle, the mu opioid receptor antagonist CTOP, the kappa-opioid receptor antagonist nor-BNI, or the delta-opioid receptor antagonist naltrindole. CTOP precipitated withdrawal behaviors in the 7-day-old rat in a dose-dependent manner. Neither nor-BNI nor naltrindole induced any significant changes in the frequency of the withdrawal behaviors. These data suggest that in the infant rat control of certain behavioral withdrawal signs is modulated primarily by the mu-opioid receptor, as is the case in the adult rat. PMID- 11113516 TI - Glial growth factor-2 promotes the survival, migration and bromodeoxyuridine incorporation of mammalian neural crest cells in caudal neural tube explant cultures. AB - Using an in vitro assay system, we found that GGF-2 increases the number of nascent trunk neural crest cells (NCC) present in the dorsal outgrowth derived from E12 caudal neural tube explants. Data is presented which suggests that this increased outgrowth was due to a combination of GGF-2 mediated effects, including its ability to promote (A) NCC survival by decreasing the percentage of NCC that undergo cell death via a mechanism involving DNA fragmentation, (B) the initial phases of NCC migration, (C) mitosis of peripherally migrating NCC. We also show that GGF-2 can promote the long-term survival of NCC in the absence of the neural tube. An immunohistochemical analysis indicates that NCC express erbB-2 and erbB 4 neuregulin receptors. PMID- 11113517 TI - Maturation of vulnerability to excitotoxicity: intracellular mechanisms in cultured postnatal hippocampal neurons. AB - Neuronal vulnerability to excitotoxicity changes dramatically during postnatal maturation. To study the intracellular mechanisms by which maturation alters vulnerability in single neurons, we developed techniques to maintain hippocampal neurons from postnatal rats in vitro. After establishing their neuronal phenotype with immunohistochemistry and electrophysiology, we determined that these neurons exhibit developmentally regulated vulnerability to excitotoxicity. At 5 days in vitro, NMDA-induced cell death at 24 h increased from 3.6% in 3-day-old rats to >90% in rats older than 21 days. Time-lapse imaging of neuronal morphology following NMDA demonstrated increasingly prevalent and severe injury as a function of postnatal age. Neither high- nor low-affinity calcium dyes demonstrated differences in peak NMDA-induced [Ca(2+)](i) increases between neurons from younger and older animals. However, neurons from older animals were uniformly distinguished from those from younger animals by their subsequent loss of [Ca(2+)](i) homeostasis. Because of the role of mitochondrial Ca(2+) buffering in [Ca(2+)](i) homeostasis, we measured NMDA-induced changes in mitochondrial membrane potential (DeltaPsi) as a function of postnatal age. NMDA markedly dissipated DeltaPsi in neurons from mature rats, but minimally in those from younger rats. These data demonstrate that, in cultures of postnatal hippocampal neurons, (a) vulnerability to excitotoxicity increases as a function of the postnatal age of the animal from which they were harvested, and (b) developmental regulation of vulnerability to NMDA occurs at the level of the mitochondrion. PMID- 11113518 TI - Cytochrome oxidase activity in rat retinal ganglion cells during postnatal development. AB - In this study, the metabolic activity of rat retinal ganglion cells during postnatal development has been examined in vivo using cytochrome oxidase histochemistry. The intensity of staining was measured by optical densitometry. The activity of cytochrome oxidase in retinal ganglion cells progressively increased from postnatal day 0 (P0) and reached a peak during the second week of postnatal development (P10-P14) and declined thereafter. Our data show that the increased levels of cytochrome oxidase seen in developing retinal ganglion cells occur at the same time, when neuronal maturity and synaptogenesis reach their peaks. PMID- 11113519 TI - Neonatal halothane anesthesia affects cortical morphology. AB - Neonatal cryoanesthesia has recently been documented to affect morphology and behavior after a single exposure [Dev. Brain Res. 111 (1998) 89; Horm. Behav. 37 (2000) 169]. In the current experiment, we investigated the effect of one-time exposure to halothane inhalant anesthesia on neonatal rats of both sexes. Fifteen minutes of exposure on postnatal day one resulted in detectable changes in the volume of the visual cortex at 3 months. Thus, neonatal halothane alters neural development and its effects are observable in the adult rat. PMID- 11113520 TI - Increased DOI-induced head shakings in adult rats neonatally treated with MK-801. AB - We examined the effects of neonatal treatment with MK-801 on 1-(2, 5-dimethoxy-4 iodophenyl)-2-aminopropane (DOI)-induced head shaking as well as [(3)H]ketanserin binding in adult rats. Neonatal rats were injected with MK-801 (0.25 mg/kg, s.c., twice daily) or with saline from postnatal days (PND) 7-18. At PND 60, a statistically significant increase in the frequency of head shaking induced by DOI (1.0 mg/kg, s.c.) was observed in the rats neonatally treated with MK-801, compared to saline-treated rats, without any change in the specific [(3)H]ketanserin binding in the frontal cortex. These results suggest that repeated NMDA receptor blockades during the critical period of brain development produce a long lasting hyper-responsiveness in the 5-HT(2A) receptor-mediated behavior, interfering with the development of neural circuits related to the behavior. PMID- 11113521 TI - Retinoic acid enhances the rate of olfactory recovery after olfactory nerve transection. AB - In the olfactory system, retinoic acid (RA) plays an important role in development and may affect growth in the adult animal. To explore the potential effects of RA on recovery after injuries, adult mice were trained in a buried food paradigm and were given a single oral supplement of RA after olfactory nerve transection. Results demonstrate that RA accelerates the recovery of olfactory functions after injury. PMID- 11113522 TI - Central allopregnanolone is increased in rat pups in response to repeated, short episodes of neonatal isolation. AB - This experiment investigated whether neonatal isolation stress alters central concentrations of progestins. Whole brain progesterone (P), dihydroprogesterone (DHP), and allopregnanolone (3alpha, 5alpha-THP) were measured in pups that were isolated from the nest, dam, and siblings for 1 h on postnatal days (PND) 2-9 and were compared to control litters of pups that were not isolated. Isolated 2-day old pups had significantly lower central P and higher P to DHP and 3alpha, 5alpha THP metabolism ratios. On PND 9, pups that had been repeatedly isolated (PND 2 8), had significantly lower whole brain DHP and significantly greater whole brain 3alpha, 5alpha-THP compared to controls. Thus, the biosocial stress of isolation in neonatal rats alters central pregnane steroids. PMID- 11113523 TI - Pentobarbital-activated Cl(-) channels in cultured adult and embryonic human DRG neurons. AB - Developmental differences in pentobarbital-activated Cl(-) currents were studied in adult and embryonic human dorsal root ganglia (DRG) neurons using whole-cell patch-clamp recordings. Pentobarbital-induced Cl(-) conductance was significantly greater in adult DRGs (28.4 pS) than in embryonic cells (19.1 pS). Fluctuation analysis of the spectral density plots of Cl(-) channel activation by pentobarbital showed age differences in the length and number of open time constants (adult cells, tau(1), tau(2), tau(3) were 224, 8. 4, 1.5 ms, respectively; embryonic cells tau(1) and tau (2) were 165 and 26.3 ms, respectively). The different kinetic properties of human adult and embryonic DRG Cl(-) channels opened by pentobarbital may reflect the presence of different subunits in the two populations of neurons. PMID- 11113524 TI - Developmental and regional expression patterns of Type I Nitric Oxide Synthase mRNA and protein in fetal sheep brain during the last third of gestation. AB - Type I NOS (nNOS) catalytic activity represents the activity of full-size protein and truncated protein variants originated from many different spliced mRNA variants. Splice mRNA variants are thought to be important in determining the differential organ and subcellular expression of Type I NOS. The present study was directed to increase our understanding of the developmental regulation of Type I NOS in fetal brain. In four discrete areas of the fetal brain, we measured steady-state mRNA levels and catalytic activity and protein mass in the soluble and particulate fractions. Under general anesthesia, we collected sensory-motor cortex, striatum, hippocampus and cerebellum from sheep fetuses at 105, 115, 125 and 135 days gestation (32 fetuses). NOS protein in the soluble and particulate fractions was characterized using Western blot (molecular weight) and arginine to citrulline conversion (enzymatic activity). At the mRNA level, steady state levels were determined using probes directed against exon 2 and exon 21/22 by ribonuclease protection assay (RPA). Our data show that NOS catalytic activity is regulated in a region, subcellular and temporal manner. NOS activity was higher in the soluble fraction in all brain regions and significantly higher levels were found in the soluble fraction of striatum and particulate fraction of hippocampus (P<0.05 by ANOVA). Western blot analysis revealed three distinct molecular weight bands for Type I NOS (155, 144 and 136 kDa). The bands were present in all brain regions and in both cellular compartments with the 155 kDa band being the most abundant molecular form. Truncated protein variants accounted for 25% and 15% of total Type I NOS protein in the soluble fraction and particulate fraction respectively. RPA analysis showed a differential regulation of mRNA variants with exon 2 frame deletions in striatum and hippocampus. A coordinated increase with advancing gestational age of catalytic activity, the full-length protein, the protein variants and steady state mRNA levels was observed in cortex and striatum as demonstrated by higher levels at 125 and 135 days gestation (P<0.05 by ANOVA). NOS enzymatic activity was Ca(2+) and calmodulin dependent. However, in the particulate fraction 20% of the NOS activity was resistant to calmodulin inhibition. In summary, fetal brain Type I NOS mRNA variants are differentially regulated according to brain regions. Our data suggests that exon 2 deleted mRNA variants have low translation efficiency as indicated by the lack of parallel expression of truncated Type I NOS protein variants. PMID- 11113525 TI - Erratum to: differential expression of S100B(1) and S100A6(1) in the human fetal and aged cerebral cortex. PMID- 11113526 TI - ZFOR2, a new opioid receptor-like gene from the teleost zebrafish (Danio rerio). AB - A new opioid receptor-like (ZFOR2) has been cloned and characterized in an anamniote vertebrate, the teleost zebrafish (Danio rerio). ZFOR2 encodes a 384 amino-acid protein with seven potential transmembrane domains, and its predicted amino acid sequence presents an overall 74% degree of identity to mammalian mu opioid receptors. Its inclusion in a dendrogram generated from the alignment of the opioid receptor's protein sequences, confirms its classification as a mu opioid receptor. Divergences in sequence are greater in the regions corresponding to extracellular loops, suggesting possible differences in ligand selectivity with respect to the classical mu opioid receptors. The genomic structure of ZFOR2 is also highly conserved throughout the phylogenetic scale, supporting the origin of opioid receptors early in evolution. Nevertheless, ZFOR2 lacks the fourth exon found in human and rodent mu opioid receptors, that is known to be involved in desensibilization and internalization processes. PMID- 11113527 TI - Low selenium diet induces tyrosine hydroxylase enzyme in nigrostriatal system of the rat. AB - We have studied the effect of a selenium-deficient diet on the nigrostriatal dopaminergic system for 15 and 30 days. The neurochemical analysis demonstrated significant elevations in nigral DA levels after 15 and 30 days of selenium deficiency. The most significant change in striatum was an elevation in dopamine (DA) in 30-day-deficient animals. As a further step, we measured the levels of activity and mRNA expression of tyrosine hydroxylase (TH) and dopamine transporter (DAT). There were significant elevations in all of these parameters in the nigrostriatal system following selenium deficiency at the two time-points studied except for the striatal DA uptake at day 30, which dropped to control levels. Altogether, our results suggest that the decrease in antioxidant capacity due to a selenium deficiency promotes an increase in DA synthesis and turnover, which are clearly associated to the induction of TH. This effect may trigger a positive feed-forward mechanism that could increase the oxidative stress considering the capacity of DA to generate free radicals. PMID- 11113528 TI - A tyrosine hydroxylase-neurofilament chimeric promoter enhances long-term expression in rat forebrain neurons from helper virus-free HSV-1 vectors. AB - Helper virus-free herpes simplex virus (HSV-1) plasmid vectors are attractive for neural gene transfer, but a promoter that supports neuronal-specific, long-term expression is required. Although expression from many promoters is unstable, a 6.8-kb, but not a 766-bp, fragment of the tyrosine hydroxylase (TH) promoter supports long-term expression. Thus, 5' upstream sequences in this promoter may enhance expression. In this study, we evaluated expression from vectors that contain 5' upstream sequences from this promoter (-0.5 to -6.8 kb) inserted at the 5' end of either a neurofilament heavy subunit (NF-H) promoter or the cytomegalovirus (CMV) immediate early promoter. The TH-NFH promoter supported expression for 6 months in the striatum, 2 months in the hippocampus, and for 1 month in both perirhinal and postrhinal cortex (the longest time points examined). Expression was targeted to neurons. The enhanced expression may require specific sequences in the TH promoter fragment because replacing this fragment with a similar sized fragment of bacteriophage lambda DNA did not enhance expression. The reverse orientation of the TH promoter fragment also enhanced expression. Insertion of insulators from the chicken beta-globin locus between the TH-NFHlac transcription unit and the vector backbone may support a modest additional enhancement in expression. Other eucaryotic sequences may also enhance expression; a S. cerevisiae (40-kb fragment)-NFH promoter enhanced expression. In contrast, the TH-CMV promoter did not enhance expression. Thus, the TH-NFH promoter may support some physiological studies that require long-term expression in forebrain neurons. PMID- 11113529 TI - Homocysteine-induced changes in mRNA levels of genes coding for cytoplasmic- and endoplasmic reticulum-resident stress proteins in neuronal cell cultures. AB - Elevated homocysteine levels have been suggested to contribute to various pathological states of the brain. However, the basic mechanisms underlying homocysteine-induced neurotoxicity have not yet been fully elucidated. In the present series of experiments, we investigated the effect of homocysteine on mRNA levels of genes coding for cytoplasmic- or endoplasmic reticulum-resident stress proteins. Primary neuronal cell cultures were exposed to different homocysteine levels for 1-24 h. Cell injury was evaluated using the MTT assay, protein synthesis was studied by measuring the incorporation of L-[4,5-3H]leucine into proteins, mRNA levels of hsp70, gadd153, grp78, and grp94 were evaluated by quantitative PCR, and changes in protein levels of hsp70, grp78 and grp94 were analyzed by immunoblotting. Exposure of cells to 5 or 10 mM homocysteine for 24 h induced marked cell injury (decrease of viability to 58 or 45% of control respectively). After 6 h treatment, gadd153, grp78 and grp94 mRNA levels increased markedly, but only when cells were exposed to levels of homocysteine high enough to induce cell injury. In addition, hsp70 mRNA levels and protein synthesis were significantly reduced. At earlier (1 or 3 h) or later (12 or 24 h) time intervals, homocysteine exposure induced a marked increase in mRNA levels of all genes studied. GRP78 and GRP94 protein levels were increased in cells exposed to 5 mM homocysteine for 24 h but not in cells exposed to 10 mM homocysteine. HSP70 protein levels, in contrast, were decreased in cells exposed to homocysteine for different periods. The expression of genes coding for ER resident stress proteins is specifically activated under conditions of ER stress. The close relationship between the extent of cell injury and increase in grp78 mRNA levels suggests that ER dysfunction may contribute to the pathological process. The results imply that the ER is an intracellular target of homocysteine toxicity. PMID- 11113530 TI - The differential molecular mechanisms underlying proenkephalin mRNA expression induced by forskolin and phorbol-12-myristic-13-acetate in primary cultured astrocytes. AB - In rat astrocytes, forskolin (FSK; 5 microM) and phorbol-12-myristic-13-acetate (PMA; 2.5 microM) increase the proenkephalin (proENK) mRNA level via different pathways. FSK-induced proENK mRNA expression is independent of protein de novo synthesis, and well correlated with CREB phosphorylation. This is in contrast to PMA-induced proENK mRNA expression that is dependent on protein de novo synthesis and is well correlated with the increase of AP-1 DNA binding activity rather than CREB phosphorylation. Differential regulation of AP-1 proteins by PMA and FSK was also observed. While c-Fos, Fra-2 and JunB were increased in response to either stimuli, only Fra-1, c-Jun and JunD were increased by PMA. The combined treatment with FSK and PMA additively increased the proENK mRNA level, which was correlated with AP-1 or ENKCRE-2 DNA binding activity, and CREB phosphorylation. Dexamethasone (DEX; 1 microM) further enhanced FSK- or PMA-induced proENK mRNA expression, which was not correlated with the activation of AP-1 expression and CREB phosphorylation, suggesting that synergistic interaction of glucocorticoid with PKA or PKC pathway for the regulation of proENK mRNA expression appears to be mediated by other pathways rather than CREB and AP-1 families. PMID- 11113531 TI - Pivagabine-induced increases in the abundance of CRF mRNA in the cerebral cortex and hypothalamus of rats. AB - The effect of treatment of rats with pivagabine (4-[(2,2-dimethyl-1-oxopropyl) amino] butanoic acid) for 4 days on the abundance of corticotropin-releasing factor (CRF) mRNA in the brain was investigated. Such treatment resulted in dose dependent (100-300 mg/kg, i.p.) increases in the amount of CRF mRNA in both the hypothalamus and cerebral cortex. The maximal increases were thus apparent with the dose of 300 mg/kg in the hypothalamus (+108%) and cerebral cortex (+49%) 30 or 60 min, respectively, after the last drug injection. Foot-shock stress administered 30 min after the final drug injection had no effect on the pivagabine-induced increases in the abundance of CRF mRNA in the hypothalamus or cerebral cortex. Such stress also had no effect on the amounts of CRF mRNA in these brain regions of vehicle-treated rats. These results demonstrate that pivagabine increases the amount of CRF mRNA in both the hypothalamus and cerebral cortex of rats, effects that might be relevant to the action of this drug in preventing the stress-induced changes in CRF hypothalamic concentration. PMID- 11113532 TI - Postischemic changes in the immunophilin FKBP12 in the rat brain. AB - An immunosuppressant tacrolimus (FK506) protects against neuronal damage following cerebral ischemia. On the other hand, the major physiological role of the immunophilin FK506-binding protein-12 (FKBP12) is a modulation of intracellular calcium flux. Since an increase in intracellular calcium concentration is a major mediator of ischemic neuronal death, we investigated the changes in FKBP12 following cerebral ischemia in the rat. We induced focal cerebral ischemia by intraluminal occlusion of the middle cerebral artery for 1 h, and global cerebral ischemia for 10 min by bilateral carotid artery occlusion combined with hypotension. The animals were killed at 4 h to 7 days after reperfusion. Immunohistochemistry was performed on paraffin sections using a monoclonal antibody raised against recombinant FKBP12. Immunoreactivity to FKBP12 in control brains was most pronounced in the CA1 subfield of the hippocampus and the striatum, the localization being primarily neuronal. Following focal ischemia, FKBP12 immunoreactivity decreased rapidly in the ischemic core by 4 h, but increased in surviving neurons in penumbra areas (4 h-7 days). Within an area of infarction, invading leukocytes and macrophages exhibited immunoreactivity to FKBP12 (3-7 days). Following global ischemia, FKBP12 immunoreactivity in CA1 neurons decreased after 1 day, and then it was lost between 2 and 7 days, although many CA1 neurons showed a transient increase in FKBP12 at 2 days. No FKBP12 immunoreactivity was observed in reactive glial cells. Thus, FKBP12 declined in dying neurons, whereas FKBP12 was upregulated in less severely injured neurons. The findings suggest that (1) FKBP12 plays an important role in the process of neuronal survival and death following cerebral ischemia, and (2) FKBP12 is involved in inflammatory reactions that occur within an area of infarction. PMID- 11113534 TI - Mild deficits in mice lacking pituitary adenylate cyclase-activating polypeptide receptor type 1 (PAC1) performing on memory tasks. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptor subtype 1 (PAC1) have been suggested to play a role in the modulation of learning and memory. However, behavioral evidence for altered mnemonic function due to altered PAC1 activity is missing. Therefore, the role of PAC1 in learning and memory was studied in mouse mutants lacking this receptor (PAC1 knock-out mice), tested in water maze two-choice spatial discrimination, one-trial contextual and cued fear conditioning, and multiple-session contextual discrimination. Water maze spatial discrimination was unaffected in PAC1 mutants, while a mild deficit was observed in multiple session contextual discrimination in PAC1 knock-out mice. Furthermore, PAC1 knock-out mice were able to learn the association between context and shock in one-trial contextual conditioning, but showed faster return to baseline than wild-type mice. Thus, the effects of PAC1 knock-out on modulating performance in these tasks were subtle and suggest that PAC1 only plays a limited role in learning and memory. PMID- 11113533 TI - Nigrostriatal innervation is preserved in Nurr1-null mice, although dopaminergic neuron precursors are arrested from terminal differentiation. AB - Various factors, including the orphan nuclear receptor Nurr1, have been implicated in dopamine biosynthesis, but many of the specific events involved in this process have to be determined. Using genetic manipulations in mice, the obligatory role for Nurr1 in dopamine (DA) biosynthesis has been documented; however, the mechanism remains unclear. DA biosynthetic enzymes, transporters and receptors are absent in the substantia nigra (SN) and the ventral tegmental area (VTA) of Nurr1-null neonates. The current study establishes that the loss of Nurr1 function does not affect the normal ventralization of neuroepithelial cells to the ventral midbrain, their differentiation into neurons, and their topographical pattern in the SN and VTA. Futhermore, the absence of Nurr1 does not affect the survival of these DA precursor cells in the ventral midbrain, as determined by quantitative analysis of cells, expressing the general neuronal nuclear marker (NeuN) and the TUNEL assay for apoptosis. These neurons express cholecystokinin (CCK), a co-transmitter of dopaminergic neurons in this area. The untranslated exon 1-2 of the Nurr1 gene, which remains intact after homologous recombination, revealed the presence of dopaminergic precursors in the ventral midbrain of the Nurr1-null mice. In addition, these neurons establish their nigrostriatal projections, as shown by axonal transport of a fluorescent tracer, DiI. These results provide evidence that Nurr1 is essential for terminal differentiation of the dopaminergic neurons in the ventral midbrain but does not affect the early steps of their neurogenesis, migration, survival and striatal projections. Our findings suggest that activation of Nurr1 might be therapeutically useful in Parkinson's disease. PMID- 11113535 TI - Conserved helix 7 tyrosine functions as an activation relay in the serotonin 5HT(2C) receptor. AB - The function of the helix VII Tyr in the conserved Asn-Pro-X-X-Tyr segment of rhodopsin-like G protein coupled receptors has been investigated in many receptors. Various effects of site-directed mutation of this locus have been found, including altered coupling, sequestration and agonist affinity. We report the first constitutively active mutations of this Tyr. In the serotonin 5HT(2C) receptor, substituting Ala or Cys for Tyr resulted in a marked increase in the basal level of inositol phosphate accumulation in transfected COS-1 cells. This constitutive signaling was abolished by the inverse agonist SB206553. Introducing Phe at this locus eliminated both basal and agonist-stimulated signaling. All three mutant receptors showed an increase in binding affinity for the structurally dissimilar agonists 5-hydroxytryptamine (5HT), (+/-)-1-(2,5 dimethoxy-4-iodophenyl)-2-aminopropane (DOI), and quipazine, suggesting that both the activating and inactivating mutations stabilize a high affinity state. These results implicate the conserved Tyr in the conformational rearrangements that occur during agonist complexing and receptor activation. PMID- 11113536 TI - Hypothalamic CART and serum leptin levels are reduced in the anorectic (anx/anx) mouse. AB - Cocaine- and amphetamine-regulated transcript (CART) is expressed in the hypothalamus, and putative peptides encoded by CART potently inhibit feeding when administered centrally. CART is strongly down-regulated in the lateral hypothalamic area and the arcuate nucleus in animal models of obesity with disrupted leptin signaling. Here we have used in situ hybridization and immunohistochemistry to study CART expression in mice homozygous for the anorexia (anx) mutation which are characterized by a much reduced food intake and premature death. anx/anx mice had significantly decreased levels of CART mRNA label and peptide-immunoreactive cell bodies and fibers in the arcuate nucleus and a lower number of detectable CART-expressing cells in the dorsomedial hypothalamic nucleus/lateral hypothalamic area. Moreover, serum leptin levels were significantly lower in anx/anx mice compared to normal littermates, most likely due to the prominent depletion of body fat in these animals. The decrease in the anorexigenic agents leptin and CART, may reflect a compensatory down regulation in response to the energy-deprived state of anx/anx mice. Alternatively, the reduced arcuate CART expression may be a consequence of a molecular defect in the arcuate nucleus of these animals. PMID- 11113537 TI - Molecular characterization and distribution of the opioid growth factor receptor (OGFr) in mouse. AB - The native opioid growth factor (OGF), [Met(5)]-enkephalin, is a tonic inhibitory peptide that modulates cell proliferation and tissue organization during development, cancer, cellular renewal, wound healing, and angiogenesis. OGF action is mediated by a receptor mechanism. The receptor for OGF, OGFr, has been cloned and sequenced in humans and rats. Using primers based on the rat OGFr cDNA, and a mouse embryo expressed sequence tag, the full-length 2.1 kb mouse OGFr cDNA was sequenced. The open reading frame was found to encode a protein of 634 amino acids, and 14 imperfect repeats of 9 amino acids each were a prominent feature. The molecular weight of OGFr was calculated as 70679, and the isoelectric point was 4.5. Northern blot analysis revealed a 2.1 kb OGFr mRNA transcript in adult mouse brain, heart, lung, liver, kidney, and triceps surae muscle. The amino acids for mouse and rat OGFr were 93% similar and 91% identical, but the mouse and human shared only a 70% similarity and a 58% identity. These results emphasize the molecular validity of OGFr, and explain the interaction of OGF with respect to normal and abnormal growth in mouse cells and tissues. PMID- 11113538 TI - Positive-negative epitope-tagging of beta amyloid precursor protein to identify inhibitors of A beta processing. AB - In this report, a novel positive-negative epitope tagging approach was developed to study the cellular processing of beta amyloid precursor protein (beta APP). Amino acids centered around the alpha-secretase cleavage site within the A beta sequence were replaced with residues comprising an epitope for which high affinity monoclonal antibodies are commercially available. The resulting mutant beta APP cDNAs were expressed in human embryonic kidney cells (HEK 293). Cleavage of labeled beta APP by beta- and gamma-secretase(s) results in the release of an epitope-tagged A beta peptide, whereas cleavage by alpha-secretase results in destruction of the epitope. Highly sensitive and specific immunoassays were developed to study processing of this labeled beta APP via the amyloidogenic pathway. Secretion of epitope-tagged A beta was prevented by MDL 28170, a previously described gamma-secretase inhibitor. Confocal microscopic studies revealed that processing and cellular trafficking of epitope-tagged beta APP was not different from wild-type beta APP. These results suggest that positive negative epitope-tagged beta APP is normally processed within the cell and may be used to identify secretase inhibitors as therapeutics for Alzheimer's disease. PMID- 11113539 TI - Simultaneous induction of mitochondrial heat shock protein mRNAs in rat forebrain ischemia. AB - Several investigations have postulated evidence of the involvement of apoptosis in delayed neuronal death following brief periods of global cerebral ischemia. Apoptosis may be closely linked to mitochondrial dysfunction. Heat shock protein (HSP) 60 and HSP10 are mitochondrial matrix proteins induced by stress and form the chaperonin complex that is implicated in protein folding and assembly within the mitochondria. This study investigated the induction of these mitochondrial stress protein genes in the hippocampal CA1 region and less vulnerable regions following transient forebrain ischemia. In situ hybridization analysis revealed that the induction pattern of HSP60 mRNA was identical to that of HSP10 mRNA throughout the entire ischemic course. No changes occurred in the expression of both mRNAs after 2 min ischemia. Strong induction of both mRNAs occurred in the CA1 region after 10 min ischemia and persisted until 1 d after reperfusion. In contrast, induction of both mRNAs in the less vulnerable regions was terminated by 1 d after reperfusion. These results demonstrate that mitochondrial stress conditions persist concomitantly with cytosolic stress conditions in regions vulnerable to transient forebrain ischemia. PMID- 11113540 TI - Tyrosine hydroxylase and norepinephrine transporter mRNA expression in the locus coeruleus in Alzheimer's disease. AB - Despite the loss of locus coeruleus (LC) noradrenergic neurons in Alzheimer's disease (AD), cerebrospinal fluid norepinephrine (NE) levels are normal or increased in AD. This paradox suggests compensatory upregulation of NE synthetic capacity or downregulation of the NE transporter (NET) in the remaining LC neurons. LC tyrosine hydroxylase (TH) mRNA expression in the LC was measured in AD subjects (n=5) and in age and gender comparable non-demented subjects (n=6). When AD subjects were divided into those still ambulatory prior to death (CDR 3/4) and those in a prolonged 'vegetative' state prior to death (CDR 5), differences among groups became apparent at specific levels of the LC. In CDR 3/4 AD subjects there was increased TH mRNA expression per neuron compared to non demented subjects in the caudal half of the LC. However, expression of NET mRNA in the same subjects was not significantly different at any level of the LC. These preliminary results suggest an upregulation of NE biosynthetic capacity in at least some LC neurons in AD prior to the very late stage of the disease. PMID- 11113541 TI - Expression of the Sox10 gene during mouse inner ear development. AB - Mutations in the SOX10 gene, encoding a cell-lineage specific transcription factor, are associated with congenital deafness. We analyzed the expression of Sox10 mRNA in developing mouse inner ear by in situ hybridization. Sox10 mRNA is expressed in the entire epithelia of the otic vesicle at embryonic day 11.5 (E11.5) and in the developing cochlea and vestibule at E13.5. In postnatal day 8 and adult cochleas, Sox10 expression is restricted to the supporting cells of the organ of Corti. These expression profiles suggest that Sox10 is important for development of the cochlea. PMID- 11113542 TI - Differential expression of neurexin mRNA in CA1 and CA3 hippocampal neurons in response to ischemic insult. AB - A short period of cerebral ischemia will trigger a cascade of events leading to neuronal death. In an effort to elucidate molecular mechanisms underlying differential vulnerability of CA1 and CA3 hippocampal neurons to neurodegeneration, we performed a transcriptional analysis of rat hippocampal neurons following transient global ischemia. In response to 15-min ischemia, the mRNA level of neurexins II alpha and III alpha was elevated in CA1 neurons and CA3 neurons, respectively. Interestingly, the up-regulated neurexin III alpha mRNA in postischemic CA3 consisted of the insert corresponding to the fourth splicing site, while the transcripts in postischemic CA1 neurons and control CA3 neurons lacked the insert. The observed tissue specific expression and the splicing pattern suggest functional importance of neurexins in postischemic degeneration of hippocampal neurons. PMID- 11113544 TI - Determination of D1 and D2 dopamine receptor expression by Ntera-2 cells. AB - There is evidence that D1 and D2 dopamine receptors are co-expressed on some neurons. As a potential model of co-expression we examined Ntera-2 cells using RT PCR, and showed that they express D2 but not D1 receptors. D2 dopamine receptor expression was confirmed by quinpirole inhibition of forskolin-stimulated cAMP formation. Absence of D1 dopamine receptors was confirmed by the inability of dopamine or SKF 81297 to increase cAMP. PMID- 11113543 TI - Molecular cloning and expression analysis of human glycogen synthase kinase-3 alpha promoter. AB - Human glycogen synthase kinase-3 alpha (GSK-3 alpha) is a serine/threonine kinase that phosphorylates a variety of cytoplasmic and nuclear proteins. It also phosphorylates components of the neuronal cytoskeleton including tau and neurofilament heavy chain. Hyperphosphorylated tau is found in neurofibrillary tangles, a hallmark of Alzheimer's disease and aberrant phosphorylation of neurofilament heavy chain is observed in motor neuron disease. Alterations in GSK 3 alpha activity may therefore contribute to the disease process in these disorders. As a first step to understand the transcriptional regulation of GSK-3 alpha, a 2-kb (p-1751/+243) DNA fragment upstream of the GSK-3 alpha initiation codon was obtained from a YAC clone and characterised. Using primer extension assays, a putative transcriptional start site was located to a G nucleotide 244 bp upstream of the ATG codon. Several transcription factor-binding sites were identified on the promoter region, but no TATA-like element was located close to the start site. Deletion mutants of the 2-kb DNA fragment were generated and fused to a promoterless chloramphenicol acetyltransferase (CAT) gene. Transfection study in a neuroblastoma cell line revealed the 1-kb (p-719/+243) fragment carried strong promoter activity, while the 2-kb construct that contains an Alu-like sequence was only 50% active. PMID- 11113545 TI - Hepatozoon canis infection associated with dog ticks of rural areas of Rio de Janeiro State, Brazil. AB - Hepatozoon canis is a tick-borne protozoan that infects dogs and has been reported throughout the world. Manifestation of H. canis infection varies from being sub-clinical in apparently healthy dogs to severe illness. The main vector of the infection is the dog tick, Rhipicephalus sanguineus although other species may also transmit this agent. H. canis has been reported previously in Brazil, but mostly as an occasional finding during laboratory exams and always associated with other diseases. The prevalence of H. canis in dogs of rural areas of Brazil has been little studied. For this study, 250 dogs from seven counties of Rio de Janeiro state were examined. All the dogs were from rural areas, near forest. Of the dogs examined, 26 dogs were from Seropedica, 82 from Itaguai, 41 from Paracambi, 26 from Mangaratiba, 32 from Barra do Pirai, 32 from Pirai and 11 from Miguel Pereira. Blood smears from the peripheral blood of the ear were taken and ticks found on the dogs were collected for identification in the laboratory. Using blood smear evaluation, H. canis was identified in 39.2% of the animals examined. Other hemoparasites identified were Babesia canis (5.2%) and Ehrlichia canis (4.8%). Four tick species were found parasitizing the dogs: Amblyomma cajennense (23.6%), R. sanguineus (12.4%), Amblyomma aureolatum (2.8%) and Amblyomma ovale (2.0%). There was a positive correlation between the presence of A. cajennense and H. canis infection. PMID- 11113547 TI - Detection of Cryptosporidium oocysts in soil samples by enzyme-linked immunoassay. AB - An ELISA protocol was adapted for detection of Cryptosporidium parvum oocysts in soil samples and the limit of detection of the test was determined. A modified indirect antigen capture ELISA protocol was developed using monoclonal antibodies against the oocyst outer wall. The accuracy of the ELISA was compared to spiked soil samples and measured in terms of sensitivity and specificity of the test. The performance of the ELISA was evaluated in field soil samples and measured using the kappa-statistics. Similarly, the performance of the ELISA was compared to the concentration flotation method, to a modified concentration flotation method and to a commercial ELISA (ProSpecT) in field fecal and soil samples. The limit of detection of the test was selected to be 10,000 oocysts/g. At this limit of detection, the ELISA had a sensitivity of 95% and specificity of 100%. The agreement between the ELISA and the modified flotation-concentration method in detecting Cryptosporidium oocysts in soil samples was 32% (kappa=0.32). The ELISA had the same relative sensitivity (82%) in comparison to both the flotation and ProSpecT in determining Cryptosporidium-infection status of an animal. The kappa statistics was 0.26 for both tests. The developed ELISA proved to be a valuable diagnostic test for detecting oocysts in soil samples and has a potential application in determining the infection status of animals. PMID- 11113546 TI - Individual intake and antiparasitic efficacy of free choice mineral and fenbendazole in range calves. AB - The current study was conducted to assess the feasibility of fenbendazole (FB) administration to steers in a free choice mineral supplement. Provision of free choice FB reduces the need for handling of animals as well as decreases the level of animal parasitism. Two separate trials were conducted using 400 +/- 19 kg Holstein steers (n=14 and 17) during the months of July and August. Each steer was tagged with a unique electronic identification (EID) ear tag and randomly allocated into one of two groups. The tags worked in conjunction with a mineral feeder equipped with a load cell by registering the steer's EID number every time the animal entered the electromagnetic field. Individual daily mineral intake and feeding times were determined over two 8-day periods of non-medicated mineral (no FB), separated by a 14-day period of medicated mineral (0.55% FB). Fecal samples were collected at the beginning and end of each trial period and were analyzed for gastrointestinal nematode eggs and Giardia cyst. There was a consistently high level of attendance for the entire experimental period, with the exception of the first six days of the adaptation period. There were three preferential times for visiting the mineral feeder, approximately 07:00, 12:00 and 18:00 h. Individual daily mineral and FB intake was 229 +/- 27.21 g/day and 2 +/- 0.14 mg/kg BW/day, respectively, for the 14-day drug delivery period. The levels of fecal nematode eggs and Giardia cysts decreased significantly (<0.01) between pre and post-sampling, with reductions of 92% for nematode eggs and 85% for Giardia cysts. Free choice medication for the control of gastrointestinal parasites is potentially effective, provided that the appropriate drug concentration, adaptation period, intake level and duration of treatment is utilized. PMID- 11113548 TI - Application of PCR and DNA probes in the characterisation of trypanosomes in the blood of cattle in farms in Morogoro, Tanzania. AB - Polymerase chain reaction (PCR) and deoxyribonucleic acid (DNA) probes were used to characterise trypanosomes from cattle in Morogoro region of Tanzania. Blood samples collected from 390 beef and dairy cattle in selected farms in Morogoro region were examined for presence of trypanosomes using the buffy coat technique (BCT) and blood smears (BSs). Fifty-two animals were found infected: 40 with Trypanosoma congolense, 10 with T. vivax and two with both T. congolense and T. vivax. DNA extracted from all the parasitologically positive and 62 randomly selected parasitologically negative samples were subjected to PCR amplification using primers specific for different trypanosome species. Using a set of seven specific-pairs of primers on the parasitologically positive samples, we detected only T. congolense, either the Savannah- or the Kilifi-type, as single or mixed infections. With the PCR, trypanosome DNA could be detected in 27 (43%) out of 62 samples that were parasitologically negative. DNA hybridisation using probes specific for Savannah- or Kilifi-types T. congolense, or T. vivax, confirmed the presence of these parasites in cattle kept on some farms in Morogoro region of Tanzania. From these studies, it is clear that there is a need to undertake molecular epidemiological studies to determine the distribution of trypanosome species and subspecies, and to assess the economic impact of these parasites in the productivity of livestock in Tanzania. In particular, it would be desirable to verify the assumed association between the different presentations of trypanosomosis on one hand and genotypes of T. congolense on the other. PMID- 11113549 TI - Experimental cross-infections of Haemonchus placei (Place, 1893) in sheep and cattle. AB - To examine effects on biology and morphology, Haemonchus placei infections were propagated in cattle or sheep and infective larvae were introduced into the same or opposite host. Ovine source larvae had a geometric mean (GM) prepatent period of 26.5 days regardless of host species, compared to 30.6 days for bovine source larvae regardless of host species. Similarly, ovine source H. placei had a higher GM percentage establishment versus bovine source larvae (9.6% versus 3.4%) regardless of host species. The patent daily egg count for ovine source versus bovine source H. placei was 109.1 versus 50.0 FEC regardless of host species. Mean spicule length for ovine source H. placei was 492.5 microm while bovine source measured 496.5 microm. Mean female tail length for ovine source H. placei was 586.5 and 589.5 microm for bovine source H. placei. Using synlophe morphology, all worms that were measured were identified as H. placei. Sixty three percent of females examined for vulvar flap morphology had knob-like vulvar flaps while the remaining 37% had linguiform vulvar flaps. PMID- 11113550 TI - Relationship between microfilaria count and sensitivity of the direct smear for diagnosis of canine dirofilariosis. AB - Direct blood smear examination (using 0.05 ml of whole blood) detected 168 (80.9%) of 204 microfilaremic canine blood samples as determined by the modified Knott test for microfilariae (mff) of Dirofilaria immitis (using 1 ml of whole blood). Direct smear examination detected all of 134 microfilaremias greater than 50 mff ml(-1), but only 31 of 70 (44.3%) microfilaremias having less than 50 mff ml(-1). In a separate retrospective query of a database of 963 dogs with necropsy confirmed heartworm infections, 834 (86.6%) were positive by the DiroCHEK heartworm antigen test, and 504 (52.3%) were microfilaremic by the modified Knott test. Only 2 (0.4%) of the microfilaremic dogs were DiroCHEK negative and another 18 (3.6%) were very weak positives. Although these microfilaremic dogs were not tested by direct smear, only one of the two DiroCHEK-negative and six of 18 weakly DiroCHEK-positive dogs had microfilaremias so low that a direct smear may have given a false negative result. Significant adverse reactions to either diethylcarbamazine or the macrolide endectocides have not been reported for microfilaremias less than 500 mff ml(-1), thus substitution of the direct smear for a concentration test for mff, such as the modified Knott test or membrane filtration, does not appear to increase the risk of an unexpected adverse reaction to heartworm prophylactic drugs. Such a substitution results in only a very slight decrease (on the order of 0.1%) in the overall sensitivity of heartworm screening, provided a test for mff is run concurrently with an antigen test. If a test for mff is the only screening test used, then substitution of a direct smear for a concentration test may decrease the sensitivity of heartworm screening by nearly 20%, depending on the prevalence of low level microfilaremias in the population of dogs tested. PMID- 11113551 TI - Survival away from sheep and alternative methods of transmission of sheep lice (Bovicola ovis). AB - Transmission of sheep lice is thought to occur mainly by sheep to sheep contact although the possibility of other sources of infestation is often suggested. This study investigated the period of survival of Bovicola ovis after removal from sheep under varying conditions and assessed the likelihood of new infestations arising from contaminated facilities, wool caught on fences and shearers' footwear. In laboratory studies with lice held away from sheep at 4, 20, 25 and 36.5 degrees C, adults and nymphs survived longest at 25 degrees C (LT90 of 11.7 and 24.1 days for adults and large nymphs, respectively). Nymphs survived longer than adults and lice provided with raw wool survived longer than lice provided with wool that had been degreased. Nymphal lice survived for up to 29 days on unscoured wool at 36.5 degrees C, but the LT50 was less than 9 days in most experiments. In shearing sheds in winter and early spring lice survived for up to 14 and 16 days, respectively. These periods of survival are considerably longer than previously indicated for B. ovis. Most lice dropped out of wool staples attached to a fence within 1 h and only two of a total of 225 lice were still present after 24 h, suggesting that sheep are unlikely to become infested from wool caught on fences. Adult and nymphal lice readily transferred to shearers' moccasins and survived there for up to 10 days, indicating that transmission of lice on the footwear of shearers or other sheep handlers may be a cause of new infestations. Microwaving each moccasin for 5 min killed all lice and may provide a simple method of reducing the likelihood of transmission of B. ovis between properties. PMID- 11113552 TI - Postnatal neosporosis in dairy cattle in northeast Thailand. AB - A total of 83 dairy cows in Loei Province (Muang) and Nong Bua Lamphu (NBL) Province, northeast Thailand were sampled three times within 6 months in 1998 and their sera were examined for antibodies to Neospora caninum at a dilution of 1:100 in the indirect fluorescent antibody test (IFAT). In Muang the seroprevalence of N. caninum was 37.5% (June), 60% (August), and 62.5% (November). In NBL the prevalence was 50% (August) and 70% (November). In both areas abortions were observed between 1 and 3 months after the introduction of these cattle from another area. Nine of 14 and seven of 17 calves were descendants of seropositive dams, of which only two calves from Muang and two calves from NBL were positive for N. caninum antibodies. These findings suggest postnatal N. caninum transmission. PMID- 11113553 TI - A method for the sequential study of eimerian chromosomes by light and electron microscopy. AB - In the present study, the authors describe a simple method to isolate chromosomes from eimerian oocysts and to submit them to sequential study by light and electron microscopy. This method includes a reliable and reproducible technique for transferring eimerian chromosomes from slides to grid that fulfills the essential requirements for generalized use in cytogenetics. In addition, this method overcomes the difficulty of the resistance of protozoan oocysts to disruption and permits the release of intact meiotic chromosomes. The observation by the authors of synaptonemal complexes in meiotic chromosomes of different Eimeria species by applying the above-mentioned method to oocysts revealed its importance to future applications. PMID- 11113554 TI - Expression, purification and characterization of beta domain and beta domain dimer of metallothionein. AB - In order to examine the independent self-assembly of the beta fragment of metallothionein and the interaction between two domains with the linker sequence, Lys-Lys-Ser, the chemically synthesized genes of the beta domain and its dimer (beta-KKS-beta) were cloned into vector pGEX-4T-1 and expressed as carboxyl terminal extension of glutathione-S-transferase (GST). After the GST fusion proteins had been digested with thrombin on a glutathione-Sepharose 4B affinity chromatography column, the beta domain and its dimer were purified with gel filtration and analyzed for their biochemical and spectroscopic properties. Amino acid composition and molecular mass are determined to be consistent with the expected value. The analysis of metal content shows that the beta domain and its dimer can bind with about 3eq and 6eq divalent metals, respectively. The characteristic peak presented around 254 nm in the UV and CD spectrum indicated that both the beta domain and its dimer are able to form the cadmium-thiolate clusters without the aid of the alpha domain. Furthermore, the absorption peak of the beta domain dimer is much higher than that of the beta domain, which suggested that there is an interaction between two beta domains. Finally, the metal-binding ability was determined by DTNB competitive reaction and the value of half dissociation pH, the results reveal that the beta domain dimer has stronger metal-binding ability than the single beta domain, which provides further evidence of the interaction between the two domains. PMID- 11113555 TI - Characterization and immunohistochemical localization of nucleoside triphosphate diphosphohydrolase (NTPDase) in pig adrenal glands (presence of a non sedimentable isoform). AB - Considering that adrenal glands possess a variety of purinoceptors associated with various cell types and that some of these cells (chromaffin cells) secrete large amounts of adenine nucleotides, it was of interest to localize nucleoside triphosphate diphosphohydrolase (NTPDase) in these glands and to define the biochemical characteristics of this ectonucleotidase. Immunolocalization produced a moderate reaction in capsula and medulla, with no signal in zona glomerulosa and zona reticularis. In contrast, a very strong reaction was found in zona fasciculata. Biochemical analysis of particulate fractions isolated from whole glands revealed high levels of ATPase and ADPase activities. This appeared to be attributable to the NTPDase since the level of ADPase was as high as ATPase. Both ATPase and ADPase activities were similarly inhibited by sodium azide. Additionally electrophoretograms with these two substrates showed comparable patterns. Western blots with 'Ringo', an antibody that recognizes the different isoforms of mammalian NTPDases, showed the presence of isoforms of NTPDases at 54 and 78 kDa, respectively. Interestingly, the 54 kDa isoform remains in the supernatant of a chromaffin granule lysate after ultracentrifugation. Up until now little interest has been given to the relationship between adrenal medulla and cortex. Presence of purinoceptors and ectonucleotidases in both these regions together with the effects of ATP in vivo and in vitro in different species indicate that purines play a significant role in adrenal glands. PMID- 11113556 TI - Differential protein profile in the ear-punched tissue of regeneration and non regeneration strains of mice: a novel approach to explore the candidate genes for soft-tissue regeneration. AB - Wound repair/regeneration is a genetically controlled, complex process. In order to identify candidate genes regulating fast wound repair/regeneration in soft tissue, the temporal protein profile of the soft-tissue healing process was analyzed in the ear-punched tissue of regeneration strain MRL/MpJ-Fas(lpr) (MRL) mice and non-regeneration strain C57BL/6J(B6) mice using surface-enhanced laser desorption and ionization (SELDI) ProteinChip technology. Five candidate proteins were identified in which responses of MRL to the ear punch were 2-4-fold different compared to that of B6. Their corresponding genes were predicted using an antigen-antibody assay validated mass-based approach. Most of the predicted genes are known to play a role or are likely to play a role in the wound repair/regeneration. Of the five candidate proteins, the amount of the 23560 Da protein in the ear-punched tissue was significantly correlated with the rate of ear healing in six representative strains of mice, making it a good candidate for fast wound repair/regeneration. We speculate that the increased concentration of the 23560 Da protein in the wound tissue could stimulate the expression of various growth-promoting proteins and consequently speed up the wound repair/regeneration processes. Here, we have shown that examination of protein expression profile using SELDI technology, coupled with database search, is an alternative approach to search for candidate genes for wound repair/regeneration. This novel approach can be implemented in a variety of biological applications. PMID- 11113557 TI - Gel-sol transition can describe the proteolysis of extracellular matrix gels. AB - We monitored the cell-free solubilization of extracellular matrix and fibronectin gels, catalyzed by exogenous proteinases. The corresponding measurements could not be interpreted according to usual proteinase kinetics. The observation that this experimental system did not consist in surface but in bulk degradation and appeared specific to gel substrates, incited us to use gelation-related approaches to describe these kinetics. We show that the gel-sol transition theory adequately describes the enzyme reactions. This allowed formulation and experimental confirmation of a power law relating macroscopic changes of the gel to enzyme kinetics. This approach could also be used for other power laws predicted by the gel-sol transition theory, allowing a better understanding of macroscopic modification of the extracellular matrix during proteolysis, which is implied in many biological situations, especially tumor dissemination. PMID- 11113558 TI - In vivo cell electrofusion. AB - In vitro electrofusion of cells brought into contact and exposed to electric pulses is an established procedure. Here we report for the first time the occurrence of fusion of cells within a tissue exposed in vivo to permeabilizing electric pulses. The dependence of electrofusion on the ratio of applied voltage to distance between the electrodes, and thus on the achievement of in vivo cell electropermeabilization (electroporation) is demonstrated in the metastasizing B16 melanoma tumor model. The kinetics of the morphological changes induced by cell electrofusion (appearance of syncytial areas or formation of giant cells) are also described, as well as the kinetics of mitosis and cell death occurrence. Finally, tissue dependence of in vivo cell electrofusion is reported and discussed, since electrofusion has been observed neither in liver nor in another tumor type. Particular microenvironmental conditions, such as the existence of reduced extracellular matrices, could be necessary for electrofusion achievement. Since biomedical applications of in vivo cell electropermeabilization are rapidly developing, we also discuss the influence of cell electrofusion on the efficacy of DNA electrotransfer for gene therapy and of antitumor electrochemotherapy, in which electrofusion could be an interesting advantage to treat metastasizing tumors. PMID- 11113559 TI - Identification of putrescine-responsive mRNAs in Chinese hamster ovary cells using representational difference analysis. PMID- 11113560 TI - Placental arylamine N-acetyltransferase type 1: potential contributory source of urinary folate catabolite p-acetamidobenzoylglutamate during pregnancy. AB - Human arylamine N-acetyltransferase type 1 (NAT1), better known as a drug metabolising enzyme, has been proposed to acetylate the folate catabolite p aminobenzoylglutamate (p-abaglu) to N-acetamidobenzoylglutamate (ap-abaglu) which is a major urinary folate catabolite. Using mass spectroscopic analysis, we demonstrate the formation of ap-abaglu by recombinant human NAT1 and human placental homogenates. Using density gradient centrifugation the placental enzymic activity which acetylates p-aba and the placental enzymic activity acetylating p-abaglu both have an S(20,w) value of 3.25 S. This is the expected value for a monomer of human NAT1 (33 kDa). The specific NAT1 inhibitor 5 iodosalicylate inhibits acetylation of both p-aba and p-abaglu catalysed by either recombinant human NAT1 or placental samples as the source of enzyme. These data demonstrate that NAT1 is the major placental enzyme involved in acetylating p-abaglu. PMID- 11113561 TI - Expression and fast-flow purification of a polyhistidine-tagged myoglobin-like aerotaxis transducer. AB - A Co(2+)-affinity, fast-flow perfusion chromatography method to purify a polyhistidine-tagged myoglobin-like aerotaxis transducer HemAT-Hs has been developed. The method relies upon a six-histidine affinity tag fused to the C terminus and N-terminus of HemAT-Hs for expression in the native host, an extremely halophilic Archaeon Halobacterium salinarum, and in the heterologous host Escherichia coli, respectively. The His-tagged HemAT-Hs can be purified rapidly using either low or high ionic strength buffers. Purified His-tagged HemAT-Hs in high or low salt buffers demonstrated no difference in spectral characteristics and retained reversible oxygen binding capacity. This fast-flow Co(2+)-affinity perfusion chromatography provides a simple method for preparation of halophilic heme containing soluble proteins for biophysical and structural studies. PMID- 11113562 TI - Stage-associated expression of ceramide structures in glycosphingolipids from the human trematode parasite Schistosoma mansoni. AB - Glycosphingolipids of Schistosoma mansoni adults, cercariae and eggs comprise ceramide monohexosides (CMH) with glucose or galactose and ceramide dihexosides (CDH) with the schistosome-specific structure GalNAc(beta1-4)Glc(1-1)ceramide. Ceramide analysis revealed C18- and C20-phytosphingosines in egg CMH, C18 sphinganine as well as C18-, C19- and C20-phytosphingosines in cercarial CMH, and C18- and C20-phytosphingosines as well as C18-sphingosine and C18-sphinganine in adult CMH. For all three life cycle stages, the predominant fatty acid was C16h:0. As a characteristic feature, a range of saturated, unsaturated and hydroxylated long-chain fatty acids with 24-28 carbon atoms were additionally found in minor cercarial CMH species. The corresponding ceramides represented major constituents in cercarial CDH, while adult and egg CDH were dominated by ceramides with short fatty acid chains. The resultant ceramide patterns could be correlated with the differential expression of carbohydrate antigens on schistosomal glycolipids at various stages. A possible impact of ceramide structure on the biosynthesis of the carbohydrate moieties is discussed. PMID- 11113563 TI - Enhanced oxidative damage by the familial amyotrophic lateral sclerosis associated Cu,Zn-superoxide dismutase mutants. AB - Some cases of familial amyotrophic lateral sclerosis (FALS), a degenerative disorder of motor neurons, is associated with mutation in the Cu,Zn-superoxide dismutase (SOD) gene SOD1. The purified FALS mutant and wild-type Cu,Zn-SODs expressed in Escherichia coli cells have identical dismutation activity whereas the hydroxyl radical formation of FALS mutants was enhanced relative to that of the wild-type enzyme. These higher free radical-generating activities of mutants facilitated the release of copper ions from their own molecules. The reaction of the mutants with hydrogen peroxide enhanced DNA strand breaks and lipid peroxidation. The results suggested that the enhanced oxidative damage of macromolecules is mediated in the Cu,Zn-SOD mutants and hydrogen peroxide system via the generation of hydroxyl radicals by a combination of the higher free radical-generating activities of mutants and a Fenton-like reaction of copper ions released from oxidatively damaged Cu,Zn-SODs. Carnosine has been proposed to act as antioxidant in vivo. We investigated whether carnosine could protect the oxidative damage induced by FALS mutants. Carnosine effectively inhibited the DNA cleavage and lipid peroxidation. These results suggest that the higher free radical-generating function of FALS mutants can lead to increased oxidative damage of macromolecules which further implicates free radical-mediated motor neuronal injury in the pathogenesis of FALS and carnosine may be explored as potential therapeutic agents for FALS patients. PMID- 11113564 TI - In vitro evaluation of antioxidant activity by electrophoresis and high performance liquid chromatography. AB - Two methods for the analysis of antioxidants, based on polyacrylamide gel electrophoresis (PAGE) and gel permeation high performance liquid chromatography (HPLC) were developed. Both of them exploit the variations of the signal (band or peak) given by human serum albumin (0.2% w/v in 100 mM sodium phosphate pH 7) upon oxidation with hypochlorite (1% of a solution containing 4% active Cl), quantitatively determined by densitometric analysis or peak integration. Based on such changes, two formulas were defined which allowed the determination of the antioxidant activity of ascorbic acid (EC(50,PAGE)=4.8x10(-4) M, EC(50,HPLC)=3.6x10(-4) M), glutathione (EC(50,PAGE)=1.5x10(-4) M, EC(50,HPLC)=2.0x10(-4) M) and melatonin (EC(50,PAGE)=5.2x10(-4) M, EC(50,HPLC)=3.2x10(-4) M), chosen as reference compounds. A good correlation was found between the activities of these substances in the two assays, which are also in good agreement with literature data, indicating that the two methods are essentially equivalent. These assays could be useful for the screening of new antioxidant drugs for pathological conditions such as cataract, rheumatic diseases, atherosclerosis and Alzheimer's disease. PMID- 11113565 TI - Lipopolysaccharide induces the expression of cellular inhibitor of apoptosis protein-2 in human macrophages. AB - Apoptosis is an important process in normal animal development as well as in diseases, and inhibitor of apoptosis protein (IAP) is one of the important factors that regulate apoptotic cell death. We found that lipopolysaccharide (LPS) enhances the expression of mRNA and protein of cellular IAP-2 (cIAP2) in human monoblastic U937 cells differentiated by phorbol ester pretreatment. cIAP2 mRNA was not detected in undifferentiated U937 cells. mRNAs of cIAP1 and X chromosome-linked IAP (XIAP) were expressed constitutively and not affected by LPS in both undifferentiated and differentiated cells. LPS stimulated the expression of cIAP2 mRNA and protein in time- and concentration-dependent manners. LPS enhanced the expression of cIAP2 mRNA and protein in human monocyte derived macrophages, which was associated with the inhibition of the caspase-3 activation, i.e., decrease in active p17 fragment of caspase-3 with simultaneous accumulation of precursor p20 fragment. We conclude that LPS may inhibit apoptosis of macrophages, at least in part, through the induction of cIAP2. PMID- 11113566 TI - Purification and characterization of UDP-glucose:tetrahydrobiopterin glucosyltransferase from Synechococcus sp. PCC 7942. AB - Tetrahydrobiopterin (BH4)-glucoside was identified from Synechococcus sp. PCC 7942 by HPLC analysis and the enzymatic activity of a glycosyltransferase producing the compound from UDP-glucose and BH4. The novel enzyme, named UDP glucose:BH4 glucosyltransferase, has been purified 846-fold from the cytosolic fraction of Synechococcus sp. PCC 7942 to apparent homogeneity on SDS-PAGE. The native enzyme exists as a monomer having a molecular mass of 39.2 kDa on SDS PAGE. The enzyme was active over a broad range of pH from 6.5 to 10.5 but most active at pH 10.0. The enzyme required Mn(2+) for maximal activity. Optimum temperature was 42 degrees C. Apparent K(m) values for BH4 and UDP-glucose were determined as 4.3 microM and 188 microM, respectively, and V(max) values were 16.1 and 15.1 pmol min(-1) mg(-1), respectively. The N-terminal amino acid sequence was Thr-Ala-His-Arg-Phe-Lys-Phe-Val-Ser-Thr-Pro-Val-Gly-, sharing high homology with the predicted N-terminal sequence of an unidentified open reading frame slr1166 determined in the genome of Synechocystis sp. PCC 6803, which is known to produce a pteridine glycoside cyanopterin. PMID- 11113567 TI - Demonstration of the central dark line in crystals of dental calculus. AB - Using an electron microscope and Fourier transform infrared (FTIR) microspectroscopy, we studied the lattice images of crystallites of dental calculus to demonstrate the presence of the central dark line (CDL) in its crystallite and to compare this CDL with that of bone and synthetic hydroxyapatite crystals. Ultrastructural observations revealed clearly a number of crystallites, which displayed a proper lattice image and CDL similar to that of bone, in the dental calculus. FTIR microspectroscopy revealed that the dental calculus displayed a set of major spectra analogous to that of bone. These results suggest that the formation process of hydroxyapatite crystals with CDL in dental calculus, which is considered to be an unusual type of calcified structure in association with microorganisms, is basically similar to that of the ordinary calcifying hard tissues (bone, enamel, etc.). PMID- 11113568 TI - Opposite regulation of tenascin-C and tenascin-X in MeLiM swine heritable cutaneous malignant melanoma. AB - Interactions between tumour cells and surrounding extracellular matrix (ECM) influence the growth of tumour cells and their ability to metastasise. It is thus interesting to compare ECM composition in tumours and healthy tissues. Using the recently described MeLiM miniature pig model of heritable cutaneous malignant melanoma, we studied the expression of two ECM glycoproteins, the tenascin-C (TN C) and tenascin-X (TN-X), in normal skin and melanoma. Using semiquantitative RT PCR, we observed a 3.6-fold mean increase of TN-C RNAs in melanoma compared to normal skin. Both stromal and tumour cells synthesise TN-C. On the contrary, TN-X RNAs decreased 30-fold on average in melanoma. This opposite regulation of TN-C and TN-X RNAs was confirmed at the protein level by indirect immunofluorescence. Whereas pig normal skin displayed a discrete TN-C signal at the dermo-epidermal junction, around blood vessels and hair bulbs, the swine tumour showed enhanced expression of TN-C in these areas and around stromal and tumour cells. In contrast, normal skin showed a strong TN-X staining at the dermo-epidermal junction and in the dermis, whereas this signal almost completely disappeared in the tumour. The results presented here describe a dramatic alteration of the ECM composition in swine malignant melanoma which might have a large influence on tumourigenesis or invasion and metastasis of melanoma cells. PMID- 11113569 TI - Endothelial NOS expression and ischemia-reperfusion in isolated working rat heart from hypoxic and hyperoxic conditions. AB - Induction of endothelial nitric oxide synthase (eNOS) contributes to the mechanism of heart protection against ischemia-reperfusion damage. We analyzed the effects of hypoxia and hyperoxia on eNOS expression in isolated working rat hearts after ischemia-reperfusion damage. Adult male Wistar rats were submitted to chronic hypoxia (2 weeks) and hyperoxia (72 h). The hearts were submitted to 15 min of ischemia and reperfused for 60 min, then we evaluated hemodynamic parameters and creatine phosphokinase (CPK) release. eNOS expression was estimated by RT-PCR; enzyme localization was evaluated by immunohistochemistry and the eNOS protein levels were detected by Western blot. All hemodynamic parameters in hypoxic conditions were better with respect to other groups. The CPK release was lower in hypoxic (P<0.01) than in normoxic and hyperoxic conditions. The eNOS deposition was significantly higher in the hypoxic group versus the normoxic or hyperoxic groups. The eNOS protein and mRNA levels were increased by hypoxia versus both other groups. Chronic hypoxic exposure may decrease injury and increase eNOS protein and mRNA levels in heart subjected to ischemia-reperfusion. PMID- 11113570 TI - Interactions of racemic mefloquine and its enantiomers with P-glycoprotein in an immortalised rat brain capillary endothelial cell line, GPNT. AB - The brain distribution of the enantiomers of the antimalarial drug mefloquine is stereoselective according to the species. This stereoselectivity may be related to species-specific differences in the properties of some membrane-bound transport proteins, such as P-glycoprotein (P-gp). The interactions of racemic mefloquine and its individual enantiomers with the P-glycoprotein efflux transport system have been analysed in immortalised rat brain capillary endothelial GPNT cells. Parallel studies were carried out for comparison in human colon carcinoma Caco-2 cells. The cellular accumulation of the P-glycoprotein substrate, [(3)H]vinblastine, was significantly increased both in GPNT cells and in Caco-2 cells when treated with racemic mefloquine and the individual enantiomers. In GPNT cells, the (+)-stereoisomer of mefloquine was up to 8-fold more effective than its antipode in increasing cellular accumulation of [(3)H]vinblastine, while in Caco-2 cells, both enantiomers were equally effective. These results suggest that racemic mefloquine and its enantiomers are effective inhibitors of P-gp. Furthermore, a stereoselective P-glycoprotein inhibition is observed in rat cells but not in human cells. The efflux of [(14)C]mefloquine from GPNT cells was decreased when the cells were incubated with the P-gp modulators, verapamil, cyclosporin A or chlorpromazine, suggesting that MQ could be a P-gp substrate. PMID- 11113571 TI - Mechanism of superoxide anion generation in the toxic red tide phytoplankton Chattonella marina: possible involvement of NAD(P)H oxidase. AB - Red tide phytoplankton Chattonella marina is known to produce reactive oxygen species (ROS), such as superoxide anion (O(2)(-)), hydrogen peroxide (H(2)O(2)) and hydroxyl radical (&z.rad;OH), under normal physiological conditions. Although several lines of evidence suggest that ROS are involved in the mortality of fish exposed to C. marina, the mechanism of ROS generation in C. marina remains to be clarified. In this study, we found that the cell-free supernatant prepared from C. marina cells showed NAD(P)H-dependent O(2)(-) generation, and this response was inhibited by diphenyleneiodonium, an inhibitor of mammalian NADPH oxidase. When the cell-free supernatant of C. marina was analyzed by immunoblotting using antibody raised against the human neutrophil cytochrome b558 large subunit (gp91phox), a main band of approximately 110 kDa was detected. The cell surface localization of the epitope recognized with this antibody was also demonstrated in C. marina by indirect immunofluorescence. Furthermore, Southern blot analysis performed on genomic DNA of C. marina with a probe covering the C-terminal region of gp91phox suggested the presence of a single-copy gene coding for gp91phox homologous protein in C. marina. These results provide evidence for the involvement of an enzymatic system analogous to the neutrophil NADPH oxidase as a source of O(2)(-) production in C. marina. PMID- 11113572 TI - Inhibition of collagenase by Cr(III): its relevance to stabilization of collagen. AB - Bacterial collagenase has now been reacted with a select series of Cr(III) complexes and modifications in the activity of chromium-modified collagenase has been deduced from the extent of hydrolysis of (2-furanacryloyl-L-leucyl-glycyl-L prolyl-L-alanine), FALGPA. A homologous series of Cr(III) complexes with dimeric, trimeric and tetrameric structures as in 1, 2 and 3 respectively has been investigated for their ability to inhibit the action of collagenase against FALGPA. Whereas competitive and non-competitive modes of inhibition of collagenase are expressed by 1, (dimer) and 2, (trimer) respectively, the tetramer, 3, exhibits poor affinity to collagenase and the inhibition of the enzyme activity is uncompetitive. Evidence for different modes of inhibition of collagenase depending on the nature of Cr(III) species has been presented in this work. Circular dichroism and gel electrophoresis data on Cr(III) modified collagenase corroborate the hypothesis that the inhibition of collagenase by the heavy metal ion arises from secondary and quaternary structural changes in the enzyme. The implications of the observed Cr(III) species specific inhibition of collagenase in gaining new insight into the mechanism of stabilization of collagen by Cr(III) are discussed. PMID- 11113573 TI - Isolation of a high affinity scFv from a monoclonal antibody recognising the oncofoetal antigen 5T4. AB - The oncofoetal antigen 5T4 is a 72 kDa glycoprotein expressed at the cell surface. It is defined by a monoclonal antibody, mAb5T4, that recognises a conformational extracellular epitope in the molecule. Overexpression of 5T4 antigen by tumours of several types has been linked with disease progression and poor clinical outcome. Its restricted expression in non-malignant tissue makes 5T4 antigen a suitable target for the development of antibody directed therapies. The use of murine monoclonal antibodies for targeted therapy allows the tumour specific delivery of therapeutic agents. However, their use has several drawbacks, including a strong human anti-mouse immune (HAMA) response and limited tumour penetration due to the size of the molecules. The use of antibody fragments leads to improved targeting, pharmacokinetics and a reduced HAMA. A single chain antibody (scFv) comprising the variable regions of the mAb5T4 heavy and light chains has been expressed in Escherichia coli. The addition of a eukaryotic leader sequence allowed production in mammalian cells. The two 5T4 single chain antibodies, scFv5T4WT19 and LscFv5T4, described the same pattern of 5T4 antigen expression as mAb5T4 in normal human placenta and by FACS. Construction of a 5T4 extracellular domain-IgGFc fusion protein and its expression in COS-7 cells allowed the relative affinities of the antibodies to be compared by ELISA and measured in real time using a biosensor based assay. MAb5T4 has a high affinity, K(D)=1.8x10(-11) M, as did both single chain antibodies, scFv5T4WT19 K(D)=2.3x10(-9) M and LscFv5T4 K(D)=7.9x10(-10) M. The small size of this 5T4 specific scFv should allow construction of fusion proteins with a range of biological response modifiers to be prepared whilst retaining the improved pharmacokinetic properties of scFvs. PMID- 11113574 TI - Synthesis of [(14)C]pyrroloquinoline quinone (PQQ) in E. coli using genes for PQQ synthesis from K. pneumoniae. AB - Radiochemical forms of pyrroloquinoline quinone (PQQ) are of utility in studies to determine the metabolic role and fate of PQQ in biological systems. Accordingly, we have synthesized [(14)C]PQQ using a tyrosine auxotrophic strain of Escherichia coli (AT2471). A construct containing the six genes required for PQQ synthesis from Klebsiella pneumoniae was used to transform the auxotrophic strain of E. coli. E. coli were then grown in minimal M9 medium containing 3.7x10(9) Bq/mmol [(14)C]tyrosine. At confluence, the medium was collected and applied to a DEAE A-25 anionic exchange column; [(14)C]PQQ was eluted using a KCl gradient (0-2 M in 0.1 M potassium phosphate buffer, pH 7.0). Radioactivity co eluting as PQQ was next pooled, acidified and passed through a C-18 column; [(14)C]PQQ was eluted with a phosphate buffer (0.1 M, pH 7.0). Reverse phase HPLC (C-18) using either the ion-pairing agent trifluoroacetic acid (0. 1%) and an acetonitrile gradient or phosphoric acid and a methanol gradient were used to isolate [(14)C]PQQ. Fractions were collected and analyzed by liquid scintillation counting. (14)C-labelled compounds isolated from the medium eluted corresponding to the elution of various tyrosine-derived products or PQQ. The radioactive compound corresponding to PQQ was also reacted with acetone to form 5-acetonyl PQQ, which co-eluted with a 5-acetonyl-PQQ standard, as a validation of [(14)C]PQQ synthesis. The specific activity of synthesized [(14)C]PQQ was 3.7x10(9) Bq/mmol [(14)C]PQQ, equal to that of [U-(14)C]tyrosine initially added to the medium. PMID- 11113575 TI - Role of paraquat in the uncoupling of nitric oxide synthase. AB - Nitric oxide synthase (NOS) oxidizes L-arginine to NO(&z.ccirf;) and L citrulline. Recent studies have shown that this enzyme can also generate O(2)(&z.ccirf;-) during its enzymatic cycling. Herein, we used spin trapping and electron paramagnetic resonance (EPR) spectroscopy to investigate the impact paraquat has on the transport of electrons through purified neuronal NOS (NOS I). In a concentration-dependent manner, ranging from 10-100 microM of paraquat, paraquat free radical was observed under anaerobic conditions. This demonstrates that NOS shunts electrons to paraquat, thereby uncoupling this enzyme. This resulted in enhanced production of O(2)(&z.ccirf;-) at the expense of NO(&z.ccirf;). Experiments demonstrated that the reductase domain is the site of paraquat-mediated uncoupling of NOS. PMID- 11113576 TI - Biodistribution characteristics of mannosylated, fucosylated, and galactosylated liposomes in mice. AB - The in vivo disposition behavior and pharmacokinetic characteristics of galactosylated (Gal), mannosylated (Man) and fucosylated (Fuc) liposomes were compared in this study. For the preparation of the glycosylated liposomes, cholesten-5-yloxy-N-(4-((1-imino-2-beta-D-thiogalactosyle thyl)amino)a lkyl)formamide (Gal-C4-Chol) (Kawakami et al., Biochem. Biophys. Res. Commun. 252 (1998) 78-83) and its mannosylated and fucosylated derivatives (Man-C4-Chol and Fuc-C4-Chol, respectively) were synthesized. The glycosylated liposomes are composed of distearoylphosphatidylcholine (DSPC), cholesterol (Chol), and Gal-C4 Chol (or Man-C4-Chol or Fuc-C4-Chol) with the molar ratio of 60:35:5. After intravenous injection in mice, these three types of [(3)H]cholesteryl hexadecyl ether-labeled glycosylated liposomes were rapidly eliminated from the circulating blood and preferentially recovered in the liver. In contrast, DSPC/Chol (60:40) liposomes without glycosylation were retained for a long time in the circulating blood. The uptake ratios by parenchymal cells (PC) and nonparenchymal cells (NPC) (PC/NPC ratios) for 0.5% Gal, Man and Fuc liposomes were found to be 15.1, 0.6 and 0.2, respectively. The effect of predosing glycosylated proteins and liposomes on the hepatic uptake of 0.5% (3)H-labeled Gal, Man, and Fuc liposomes was investigated and the results support the conclusion that Gal, Man, and Fuc liposomes are taken up by the liver via asialoglycoprotein receptors in PC, mannose receptors in NPC, and fucose receptors in NPC, respectively. Interestingly, Gal liposomes were taken up by NPC rather than by PC at a high dose (5%). Together with the finding that 5% Gal liposomes inhibit the hepatic uptake of (3)H-labeled Fuc liposomes, this suggests that Gal-liposomes administered at a high dose will also be taken up by fucose receptors in NPC, that are considered to act as galactose particle receptors. PMID- 11113577 TI - Role of sodium channel inhibition in neuroprotection: effect of vinpocetine. AB - Vinpocetine (ethyl apovincaminate) discovered during the late 1960s has successfully been used in the treatment of central nervous system disorders of cerebrovascular origin for decades. The increase in the regional cerebral blood flow in response to vinpocetine administration is well established and strengthened by new diagnostical techniques (transcranial Doppler, near infrared spectroscopy, positron emission tomography). The latest in vitro studies have revealed the effect of the compound on Ca(2+)/calmodulin dependent cyclic guanosine monophosphate-phosphodiesterase 1, voltage-operated Ca(2+) channels, glutamate receptors and voltage dependent Na(+)-channels; the latest being especially relevant to the neuroprotective action of vinpocetine. The good brain penetration profile and heterogenous brain distribution pattern (mainly in the thalamus, basal ganglia and visual cortex) of labelled vinpocetin were demonstrated by positron emission tomography in primates and man. Multicentric, randomized, placebo-controlled clinical studies proved the efficacy of orally administered vinpocetin in patients with organic psychosyndrome. Recently positron emission tomography studies have proved that vinpocetine is able to redistribute regional cerebral blood flow and enhance glucose supply of brain tissue in ischemic post-stroke patients. PMID- 11113578 TI - Action of 4-aminopyridine on extracellular amino acids in hippocampus and entorhinal cortex: a dual microdialysis and electroencehalographic study in awake rats. AB - In order to study the role of amino acids in the hippocampus and the entorhinal cortex during the convulsive process induced by 4-aminopyridine (4-AP), we have used a device allowing the simultaneous microdialysis and the recording of their electrical activity of both regions in freely moving rats. We found that infusion of 4-AP into the entorhinal cortex resulted in a large increase in extracellular glutamate and glutamine and small increases in glycine and taurine levels. Likewise, infusion of 4-AP into the hippocampus resulted in a major increase in glutamate, as well as slight increases in taurine and glycine. In both infused regions the peak concentration of extracellular glutamate was observed 15 min after 4-AP administration. No significant changes were found in the non-infused hippocampus or entorhinal cortex of the same rats. Simultaneous electroencephalographic recordings showed intense epileptiform activity starting during 4-AP infusion and lasting for the rest of the experiment (1 h) in both the entorhinal cortex and the hippocampus. The discharges were characterized by poly spikes and spike-wave complexes that propagated almost immediately to the other region studied. These findings suggest that increased glutamatergic synaptic function in the circuit that connects both regions is involved in the epileptic seizures induced by 4-AP. PMID- 11113579 TI - In vitro and in vivo characterization of hNT neuron neurotransmitter phenotypes. AB - The hNT neuron exhibits many characteristics of neuroepithelial precursor cells, making them an excellent model to study neuronal plasticity in vitro and in vivo. These cells express a number of neurotransmitters in vitro, including dopamine, gamma-aminobutyric acid and acetylcholine. However, there have been few reports of the neurotransmitters that hNT neurons express in vivo. The present study examined whether hNT neurons express the same neurotransmitters in vivo as they do in vitro. First, the expression of tyrosine hydroxylase (TH), glutamic acid decarboxylase (GAD), choline acetyltransferase (ChAT) and the human specific nuclear marker NuMA by hNT neurons was confirmed. Nineteen normal animals were then transplanted with 80,000 hNT neurons aimed at the striatum, hippocampus or cerebral cortex. Five additional animals received injections of medium. All animals received daily intraperitoneal injections of cyclosporine (10 mg/kg) and survived 30 days. Sections through the transplants were examined for NuMA positive hNT neurons, and for the presence of the three neurotransmitter markers: TH, GAD and ChAT. The hNT neurons were found in the striatum and cortex. Of the hNT neurons found within the rat striatum, 33% were ChAT-positive. In the cortex, only 4% of the neurons expressed ChAT. No GAD-positive hNT neurons were detected at either site. No NuMA-positive neurons were found in the hippocampus. The implanted hNT neurons did not induce activation of astrocytes as determined by immunocytochemistry for glial fibrillary acidic protein (GFAP). Moreover, no hNT neuron was found to express GFAP in vivo. Together, these data suggest that the hNT neurons engraft in the new host tissue, maintain their neuronal identity and may be guided in differentiation according to local environmental cues. PMID- 11113580 TI - Estradiol masculinizes the posteromedial cortical nucleus of the amygdala in the rat. AB - It has been demonstrated that the posteromedial cortical amygdaloid nucleus (PMCo), is sexually dimorphic. It is shown (Experiment 1) that male orchidectomy on the day of birth (D1) decreases the volume and number of neurons of the PMCo, while a single injection of propionate testosterone to the female on D1 masculinizes the PMCo in this gender. Since male gonadectomy on D1 (Experiment 2) is counteracted by a single injection of estradiol benzoate in males it has been suggested that the masculinization of the PMCo is due to the aromatization of testosterone to estradiol in this structure. These findings support the hypothesis that the development of sex differences in structures that belong to the vomeronasal system are due to the aromatization of testosterone to estradiol shortly after birth. PMID- 11113581 TI - Cholinergic inputs to rostral ventrolateral medulla pressor neurons from hypothalamus. AB - The rostral ventrolateral medulla (RVLM) has cholinergic mechanisms responsible for pressor responses. Stimulation of the hypothalamic paraventricular nucleus (PVN) causes an increase of arterial pressure via activation of neurons in the RVLM. In this study, we examined whether PVN stimulation causes a pressor response via activation of cholinergic mechanisms in the RVLM. Male Wistar rats were used and they were anesthetized, paralyzed and artificially ventilated. Electrical stimulation of the PVN produced a pressor response. Microinjection of the muscarinic receptor antagonist scopolamine and the cholinesterase inhibitor physostigmine into the RVLM inhibited and potentiated, respectively, the pressor response induced by PVN stimulation. PVN stimulation also increased the firing rate of RVLM barosensitive neurons and the increase in the firing rate was inhibited and potentiated by scopolamine and physostigmine, respectively, iontophoretically applied on neurons. Microinjection of L-glutamate into the PVN produced a release of ACh in the RVLM. The inhibitory amino acid gamma aminobutyric acid injected into the lateral parabrachial nucleus (LPBN) inhibited the pressor response induced by PVN stimulation. These results suggest that PVN stimulation causes an increase in arterial pressure via activation of cholinergic inputs in the RVLM. It appears that the pressor response is mediated, at least in part, via cholinergic inputs from the LPBN. PMID- 11113582 TI - Diazepam induces FGF-2 mRNA in the hippocampus and striatum. AB - starting by 6 h following diazepam injection and returning to approximately control values by 24 h. In situ hybridization showed elevated FGF-2 mRNA labeling in the hippocampal formation, mostly in the pyramidal layer of the CA1 and CA2 subfields and in the dentate gyrus hilar region. These results indicate that diazepam treatment up-regulates FGF-2 expression in select regions of the brain and suggest that GABA may promote neuroplasticity in concert with FGF-2. PMID- 11113583 TI - Prefrontocortical serotonin depletion results in plastic changes of prefrontocortical pyramidal neurons, underlying a greater efficiency of short term memory. AB - The prefrontal cortex activity is involved in organizing the short-term memory. Although the involvement of serotonin for an appropriate performance in learning and memory tests is well known, its role is still unclear; as is the cellular basis of short-term memory behavioral performance. Sprague-Dawley rats were stereotactically injected with 1 microg/microl of 5, 7-dihydroxitryptamine to cause a lesion to the dorsal raphe nucleus. Sham-operated or intact rats were also studied as control groups. Before surgery and 20 days post-operatively, each animal was placed in the Biel maze for five consecutive trials. In the pre treatment test, all three groups decreased significantly the number of errors beginning with the fourth trial. The same occurred in the post-treatment test, except for the experimental group, whose animals committed less errors beginning with the second trial. After behavioral testing, the dorsomedial prefrontal cerebral cortex was dissected out, and the Golgi study of the third-layer pyramidal neurons revealed that the length of both the apical and the basilar dendrites was smaller than that of controls, and that the apical and oblique dendrites had a greater spine density. A major proportion of thin spines was also seen on the basilar and oblique dendrites, and more stubby spines were seen on the apical dendrite. Serotonin depletion in the prefrontal cerebral cortex resulted in cytoarchitectural alterations of the prefrontocortical pyramidal neurons, which may be underlying partially the greater efficiency observed in the short-term memory behavioral performance. PMID- 11113584 TI - Age-related alterations of nitric oxide production in the brains of seizure susceptible EL mice. AB - We evaluated age-related changes in nitric oxide (NO) production in the brains of EL mice, a strain highly susceptible to seizures. A group of EL(s) mice were tossed up weekly to induce convulsive seizures, while in a nonstimulated EL(ns) group induction of convulsive seizures was avoided. Brain levels of nitrite plus nitrate (NOx) in EL(ns) mice were significantly higher than in nonstimulated mice at 10 days, and also higher than levels at 15 and 50 weeks in either EL(s) or EL(ns) mice. A significantly higher number of NO-producing cells were demonstrated in the hippocampus and parietal cortex by staining for nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase in EL(s) mice at the ages of 15 and 50 weeks than in EL(ns) mice at the age of 6 weeks. In EL(ns) mice, significantly fewer neurons showed NADPH-diaphorase staining in the hippocampus, striatum and parietal cortex at the age of 50 weeks than at 6 weeks. The present results suggest that whole-brain NOx levels in EL(ns) and EL(s) mice and numbers of NADPH-diaphorase-positive neurons in EL(ns) mice decreased with aging, while increasing of numbers of such neurons in EL(s) mice were assumed to develop in compensation for reduction in whole-brain NOx levels. PMID- 11113585 TI - Expression of fos in the hypothalamus of rats exposed to warm and cold temperatures. AB - Fos-like immunoreactivity was investigated in hypothalamic areas involved in central thermoregulatory processes. Different groups of urethane anaesthetized rats (n = 36) were exposed to: (1) 23.5 degrees C for 1 h (control); (2) 5 degrees C for 20 min (short cold); (3) 5 degrees C for 1 h (long cold); (4) 47 degrees C for 10 min (short warm) and (5) 47 degrees C for 1 h (long warm). Fos was present in the supramammillary nucleus, supraoptic nucleus and paraventricular hypothalamus of all (control and long and short, warm- and cold exposed) rats. Fos was seen in the dorsomedial, medial and ventromedial hypothalamus of rats with long or short exposure to both warm and cold temperatures, and in the medial preoptic area and lateral anterior hypothalamus of long and short warm-exposed, and long cold-exposed, rats. Fos was present in the hypothalamus of long and short cold-exposed animals only in the posterior hypothalamus, and in the anterior hypothalamus (central and anterior divisions), suprachiasmatic nucleus and ventrolateral preoptic area of short and long warm exposed rats. These results provide information on the location of neurons in rat hypothalamus activated by exposure to warm and cold temperatures and may aid in the functional identification of central thermoregulatory pathways. PMID- 11113586 TI - Identification of urocortin mRNA antisense transcripts in rat tissue. AB - Urocortin (UCN) has 45% sequence homology with corticotropin releasing factor (CRF) and binds to CRF receptors. We used reverse-transcriptase-polymerase chain reaction to demonstrate the presence of UCN RNA in various brain regions and in peripheral tissues. Ribonuclease protection assay (RPA) using sense and antisense riboprobes demonstrated the presence of a naturally occurring antisense UCN RNA transcript in a number of tissues. Northern blot indicated that the antisense transcript was the same size as sense UCN. RPA, using probes that covered bases 1 to 560 of 579 bp sequence of rat UCN, indicated that the antisense sequence was complementary to sense UCN but did not contain an open reading frame. Sense and antisense UCN RNA were co-expressed in all tissues that contained levels of either transcript detectable by RPA. Sense RNA expression was greater than antisense in the midbrain, the two transcripts were expressed equally in the hypothalamus and antisense was expressed at higher levels than sense in the liver, heart, and skeletal muscle. Antisense RNA expression was stress responsive, suggesting that it may play a role in regulating transcription or translation of UCN mRNA. PMID- 11113587 TI - Neurokinin A inhibits oxytocin and GABA release from the posterior pituitary by stimulating nitric oxide synthase. AB - Neurokinin A (NKA) is a tachykinin that participates in the control of neuroendocrine functions. The posterior pituitary lobe (PP) contains abundant nitric oxide synthase (NOS), suggesting that nitric oxide (NO) may play a role in controlling the release of neuropeptides and neurotransmitters. In the present project, we investigated the in vitro effect of NKA on oxytocin release from hypothalamic explants and PP of male rats and the possible involvement of NO in the action of NKA. Since NKA inhibits gamma-aminobutyric acid (GABA) release from PP, we also examined the role of NO in the effect of NKA on basal and K(+)-evoked GABA release. NKA (10(-7)-10(-5) M) significantly decreased oxytocin release from PP, whereas it did not affect its release from hypothalamic explants. The inhibitory effect of NKA on oxytocin release from PP was completely blocked by the NOS inhibitors N(G)-monomethyl-L-arginine (L-NMMA, 0.5 mM) or N(G)-nitro-L arginine-methyl-ester (L-NAME, 1 mM). Sodium nitroprusside (0.5 mM), an NO releaser, had no effect on basal GABA release but significantly decreased K(+) evoked GABA release. L-NMMA (0.3 mM) and L-NAME (0.5 mM) increased K(+)-evoked GABA release, indicating that NO plays an inhibitory role in GABA release from PP. The inhibition in both basal and K(+)-evoked GABA release induced by NKA (10( 7) M) was reduced by L-NAME (1 mM). Also, NKA (10(-7) M) increased NO synthesis as measured by [(14)C] citrulline production. Considered all together, our data indicate that NO may mediate the inhibitory effect of NKA on the release of both oxytocin and GABA from PP. PMID- 11113588 TI - Lordosis-inhibiting effect of progesterone in male and female rats with septal lesions. AB - The inhibitory role of progesterone (P) in regulating lordosis was investigated in male and female rats with septal lesions (SL). Male rats with SL showed lordosis quotients (LQ) as high as female rats with SL and female control rats without brain surgery after injection of 50 microg/kg estradiol benzoate (EB) followed by 0.5 mg P 44 h later. Even when primed with 5 mg P 1 h prior to the 50 microg EB-injection, the mean LQs were still high in all groups. When the dose of EB was decreased to 5 microg/kg, all rats showed high-score LQs. In contrast, all animals in both male and female in which 5 mg P was injected 1 h before 5 microg EB, showed low LQs. These results suggest that P is effective in suppressing lordosis enhanced by estrogen in either male rats or females. Furthermore, the high dose of estrogen overcomes the inhibitory action of P on lordosis in both sexes. PMID- 11113589 TI - Antihyperalgesia effect of BmK IT2, a depressant insect-selective scorpion toxin in rat by peripheral administration. AB - The study was undertaken to assess the antihyperalgesia effect of BmK IT2, a sodium channel-specific ligand purified from the venom of Chinese scorpion Buthus martensi Karsch in rat by peripheral injection. The peripheral inflammation of rat was induced by carrageenan resulted in hyperalgesia to heat stimulus. The heat hyperalgesia was measured by paw withdrawal latency (PWL). PWL was increased to 272 +/- 18%, 217 +/- 19% and 186 +/- 16% of the control by application of 10 microl BmK IT2 at the concentration of 0.1, 0. 01 and 0.001 mg/ml in inflammatory rats, respectively. In contrast, PWL was enhanced to about 177 +/- 16%, 141 +/- 15% and 133 +/- 15% of the control at the same applied concentrations of BmK IT2 in normal rats. The results thus suggest that BmK IT2 can produce peripheral antihyperalgesia and antinociception, which might be attributed a pathway of modulating the sodium channels on nociceptor. PMID- 11113590 TI - Endogenous dopamine potentiates the effects of glutamate on extracellular GABA in the prefrontal cortex of the freely moving rat. AB - Using microdialysis, the effects of endogenous dopamine on basal extracellular concentrations of gamma-aminobutyric acid (GABA) and on the increases of GABA produced by glutamate were investigated in the medial prefrontal cortex of the awake rat. The dopamine uptake inhibitor nomifensine (1, 100 and 1000 microM), used to increase extracellular dopamine, produced a dose-related increase of dialysate dopamine (0.1-1 nM) but did not change dialysate concentrations of GABA or glutamate at any dose used. The glutamate uptake inhibitor L-trans-pyrrolidine 2,4-dicarboxilic acid (PDC; 0.5 and 2 mM), used to increase extracellular glutamate, produced a dose-related increase of dialysate glutamate (1.5-5.5 microM) and increased dialysate GABA by 125%. When a simultaneous increase of endogenous dopamine and glutamate was produced, the increases of dialysate GABA were significantly higher (185% of baseline) than those produced by glutamate alone. These effects on dialysate GABA were attenuated by the D2 receptor antagonist (-) sulpiride, but not by the D1 receptor antagonist SCH-23390, all of which suggests that extracellular dopamine plays an important role in modulating endogenous glutamate-GABA interactions in the prefrontal cortex of the rat. PMID- 11113591 TI - Effects of N-methyl-D-aspartate, kainate or veratridine on extracellular concentrations of free D-serine and L-glutamate in rat striatum: an in vivo microdialysis study. AB - Using an in vivo microdialysis technique, we have investigated the effect of N methyl-D-aspartate (NMDA) or kainate on the extracellular concentrations of free D-serine and L-glutamate in the striatum. A intrastriatal perfusion of NMDA or kainate caused a significant increase in the extracellular release of L glutamate, but a significant decrease in that of D-serine. Co-perfusion of an NMDA receptor antagonist, MK-801, or an alpha-amino-3-hydroxy-5-methyl-isoxazole 4-propionic acid/kainate receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3 dione (CNQX), with NMDA or kainate significantly reversed the NMDA- or kainate induced decrease in the extracellular level of D-serine, respectively. The NMDA- or kainate-evoked increase in the extracellular L-glutamate level was also reversed by co-perfusion of MK-801 or CNQX, respectively. Because D-serine acts as a potent and selective agonist for the glycine site of the NMDA receptor and because intracerebroventricularly injected D-serine is accumulated in the astrocytes, D-serine could be taken up by the astrocytes following synaptic activation. Furthermore, because cortical ablation to remove corticostriatal glutamatergic inputs attenuates the excitotoxic effects of kainate in the striatum, L-glutamate may enhance its own release through a presynaptic NMDA and/or non-NMDA receptor-mediated mechanism. PMID- 11113592 TI - Serotonin-mediated striatal dopamine release involves the dopamine uptake site and the serotonin receptor. AB - Modulation of striatal dopamine (DA) release by serotonin (5HT) and its antagonists was studied utilizing in vitro perfusion techniques. In isolated striatal tissue, 5HT (10 microM) increased the fractional basal release of labeled DA. The 5HT(2/1c) antagonist ketanserin (5 microM) also stimulated the basal release. These two effects were mediated by different mechanisms as cocaine (10 microM) greatly inhibited the 5HT-mediated response, but slightly increased the ketanserin-mediated response. 6-Nitroquipazine maleate (10 microM, 5HT uptake inhibitor) partially inhibited both responses. Inhibition by GBR 12909 (DA uptake inhibitor) at 1 microM of the 5HT-mediated DA release was similar to that of cocaine, but at 10 microM it increased release before addition of 5HT, and maintained elevated DA release while present in the incubation medium. At 1 microM GBR 12909, ketanserin-mediated DA release was stimulated and a much greater release was seen at 10 microM, but the prolonged release was not observed as after 5HT-mediated release. Among other antagonists methiothepin (5HT(1,2,6) antagonist) also enhanced DA release, whereas oxymetazoline (5HT(1A,1B,1D) agonist) had no effect. RS2359-190 (5HT(4) antagonist) had a small effect (slight stimulation) on 5HT-mediated DA release, and no effect on ketanserin-mediated DA release. CGS 12066A (5HT(1B) agonist) inhibited 5HT-mediated DA release. The glutamate antagonist MK-801 and the GABA(A) antagonist bicuculline had no affect on either response. These results indicate that 5HT-mediated DA release occurs via reversal of the DA transporter and that inhibitory presynaptic 5HT heteroreceptors and both inhibitory and stimulatory somato-dendritic 5HT receptors regulate release. In addition to the reversal of the transporter, an inhibitory 5HT(2) component was identified. PMID- 11113593 TI - The polymodal sensory cortex is crucial for controlling lateral postural stability: evidence from stroke patients. AB - In modern literature, internal models are considered as a general neural process for resolving sensory ambiguities, synthesising information from disparate sensory modalities, and combining efferent and afferent information. The polymodal sensory cortex, especially the temporoparietal junction (TPJ), is thought to be a nodal point of the network underlying these properties. According to this view, a pronounced disruption of the TPJ functioning should dramatically impair body balance. Surprisingly, little attention has been paid to this possible relationship, which was the subject of investigation in this study. Twenty-two brain-damaged patients and 14 healthy subjects were subjected to a self-regulated lateral balance task, performed while sitting for 8 s on a rocking platform. Their lateral body balance was analysed both with and without vision (darkness). Support displacements in the frontal plane were recorded by means of an accelerometer. Two criteria were taken into account to evaluate body stability in each trial: the number of aborted trials due to balance loss and the angular dispersion of the supporting surface. Lesions involving the temporoparietal junction were found to markedly increase body instability, both with and without vision. Therefore, the temporoparietal junction plays a pivotal role in lateral body stabilisation, irrespective of the sensory condition in which the task is performed. This suggests that body stability is controlled throughout internal model(s). PMID- 11113595 TI - Proceedings of the Fifth International Symposium on the Neurobiology and Neuroendocrinology of Aging, Bregenz, Austria, July 23-28, 2000. PMID- 11113594 TI - The action of isatin (2,3-dioxoindole) an endogenous indole on brain natriuretic and C-type natriuretic peptide-induced facilitation of memory consolidation in passive-avoidance learning in rats. AB - Isatin is an endogenous indole which has been shown to counteract some of the effects of atrial natriuretic peptide (ANP) both in vitro and in vivo. The present study was designed to determine whether it could antagonise in vivo effects of the related peptides brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP). The model used was consolidation of memory in a one trial step-through passive-avoidance paradigm in the rat. Previous studies have shown that all three peptides (1 microg intracerebroventricular) can consolidate such learning and increase latency to entry a dark box. Isatin was given intraperitoneally at doses of 5, 10 and 50 mg/kg before the peptide, or a saline control. Both BNP and CNP significantly increased the latency of entry. Isatin alone had no effect. Isatin reduced the effect of both BNP and CNP; this was significant for its effect on BNP at 50 mg/kg and on CNP at both 10 and 50 mg/kg. These results show that isatin can inhibit behavioural effects of BNP and CNP as well as ANP. PMID- 11113596 TI - Demography of longevity: past, present, and future trends. AB - Life expectancy at birth has roughly tripled over the course of human history. Early gains were due to a general improvement in living standards and organized efforts to control the spread of infectious disease. Reductions in infant and child mortality in the late 19th and early 20th century led to a rapid increase in life expectancy at birth. Since 1970, the main factor driving continued gains in life expectancy in industrialized countries is a reduction in death rates among the elderly. In particular, death rates due to cardiovascular disease and cancer have declined in recent decades thanks to a variety of factors, including successful medical intervention. Based on available demographic evidence, the human life span shows no sign of approaching a fixed limit imposed by biology or other factors. Rather, both the average and the maximum human life span have increased steadily over time for more than a century. The complexity and historical stability of these changes suggest that the most reliable method of predicting the future is merely to extrapolate past trends. Such methods suggest that life expectancy at birth in industrialized countries will be about 85 87years at the middle of the 21st century. PMID- 11113597 TI - Dietary restriction and aging in rhesus monkeys: the University of Wisconsin study. AB - Dietary restriction (DR) retards aging and extends the maximum lifespan of laboratory mice and rats. To determine whether DR has similar actions in a primate species, we initiated a study in 1989 to investigate the effects of a 30% DR in 30 adult male rhesus monkeys. In 1994, an additional 30 females and 16 males were added to the study. Although the animals are still middle-aged, a few differences have developed between the control and DR animals suggesting that DR may induce physiologic changes in the rhesus monkey similar to those observed in rodents. Fasting basal insulin and glucose concentrations are lower in DR compared to control animals while insulin sensitivity is higher in the restricted animals. DR has also altered circulating LDL in a manner that may inhibit atherogenesis. These results suggest that DR may be slowing some age-related physiologic changes. In addition to measures of glucose and lipid metabolism, the animals are evaluated annually for body composition, energy expenditure, physical activity, hematologic indices, and blood or urinary hormone concentrations. In the next few years, the first animals will reach the average lifespan ( approximately 26 years) of captive rhesus monkeys and it will become possible to determine if DR retards the aging process and extends the lifespan in a primate species. PMID- 11113598 TI - Viral oncogenes as tools to study replicative senescence of human cells. AB - Human aging is correlated with reduced proliferation of various cell types, a phenomenon that can be reproduced in in vitro models of replicative senescence. We study senescence of several human primary cell types by analysis of age related changes in gene expression and gene function. In a second approach, my group uses immortalizing oncogenes derived from DNA tumor viruses as genetic tools to study genetic and biochemical mechanisms underlying the progression of cells into senescence. Specifically, our work is guided by the hypothesis that cellular proteins binding to the E7 gene product of human papillomavirus are good candidates for senescence-inducing cellular factors. For several of these cellular factors, e.g. the inhibitor of cyclin-dependent kinases p21(WAF-1), a functional role in senescence has already been demonstrated. PMID- 11113599 TI - Targeted neuronal gene expression and longevity in Drosophila. AB - Earlier studies from this laboratory have shown that in the insect, Drosophila melanogaster, the motorneuron is an important cellular nexus between the metabolism of reactive oxygen species (ROS) and adult lifespan. This was demonstrated by experiments in which expression of CuZn SOD (SOD1) specifically in motorneurons was shown to extend the mean and maximum adult lifespans to 140% of normal, and to rescue the majority of deliterious phenotypes displayed by SOD1 null mutants. We have interpreted these results to mean either that the lifespan of the organism is normally limited by the functional lifespan of this post mitotic cell type, or that ROS metabolism in motorneurons affects organismic lifespan via a systemic, perhaps neuroendocrine, signaling mechanism. We have now extended these studies to ask: (i) whether expression of catalase (CAT) or of the mitochondrially-localized Mn SOD (SOD2) in motorneurons, either singly or in combination with SOD1, have similar effects on lifespan; (ii) if expression of SOD2 can rescue SOD1-null mutant phenotypes; and (iii) if ROS metabolism in cell types other than motorneurons has significant impact on aging and lifespan determination. PMID- 11113600 TI - Neuronal plasticity and stressor toxicity during aging. AB - Brain aging, Alzheimer disease and stroke share common elements of deficits in calcium regulation, declines in mitochondrial function, increases in generation of reactive oxygen species (ROS), accumulated damage from ROS and immune system dysfunction. The problem is to distinguish less significant side reactions, such as gray hair, from aspects of aging that contribute to disease. Toward establishing cause and effect relationships, a neuron cell culture system is described that allows comparisons with age under uniform environmental conditions. This neuron culture model indicates that susceptibility to death by apoptosis and consequences of the inflammatory response from beta-amyloid are age related and an inherent characteristic of the neurons. Further mechanistic investigations are possible. New therapeutic approaches are suggested that combine inhibition of calcium overloads (calcium channel blockers), reduced ROS damage (melatonin, N-acetyl-cysteine), and bolstered mitochondrial function and energy generation (creatine). Together with newly demonstrated capabilities for adult and aged neuron regeneration and multiplication, i.e. plasticity, these approaches offer new hope toward reversing age-related decrements and damage from neurodegenerative disease. PMID- 11113601 TI - NMDA receptor activation in the aged rat hippocampus. AB - Age-related alterations of N-methyl-D-aspartate receptor (NMDAr) activation were investigated in the CA1 field of hippocampal slices from young (3-6 months old) and aged (25-33 months old) Sprague-Dawley rats by using ex vivo extracellular electrophysiological recording techniques. NMDAr-mediated field excitatory postsynaptic potentials (fEPSPs) were induced by electrical stimulation of glutamatergic fibers in a magnesium (Mg(2+))-free medium supplemented with the non-NMDAr antagonist CNQX. The fEPSPs were significantly smaller in aged rats, whereas the response of presynaptic afferent fibers remained unaffected. No significant age-related differences were found in the ability of Mg(2+) to depress the magnitude of NMDAr-mediated fEPSPs. The responsiveness of postsynaptic NMDAr to the agonist was assessed in both groups of animals. No age related differences were recorded either in the depolarizing effect of bath applied NMDA or in the magnitude of the depolarization after altering extracellular Mg(2+) concentration. Finally, short-term potentiation (STP) of excitatory transmission was studied in young and aged rats considering the pivotal role of NMDAr in synaptic plasticity. No age-related alterations of the magnitude and the time course of STP in response to 10 or 30Hz conditioning stimulation were found. Because of the decrease in the magnitude of NMDAr mediated synaptic transmission in aged animals, the absence of obvious modifications of synaptic plasticity suggests the occurrence of compensatory mechanisms that are discussed. PMID- 11113602 TI - 5-Lipoxygenase and cyclooxygenase mRNA expression in rat hippocampus:early response to glutamate receptor activation by kainate. AB - Recent research has identified in central nervous system neurons the expression of two enzymes from the inflammatory pathway of the metabolism of arachidonic acid, the 5-lipoxygenase (5LOX) and the cyclooxygenase-2 (COX2). Expression of both enzymes appears to be upregulated during aging; upregulated 5LOX/COX2 expression in neurons may be responsible for the increased neuronal vulnerability to degeneration. Involvement of the excitatory neurotransmitter glutamate in aging-associated neurodegeneration has also been suggested. Stimulation of glutamate receptors by kainic acid (kainate) has been shown independently to affect the brain expression of 5LOX or COX2. Using a quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay to measure the contents of mRNAs we found 3h after kainate injection (intraperitoneally; 10 mg/kg) increased mRNA levels of 5LOX and COX2, but not that of COX1 in the hippocampus of rats. Pretreatment with the COX2 inhibitor NS-398 (9 mg/kg, 1h prior to kainate) inhibited the kainate-stimulated increase of 5LOX and COX2 mRNA levels. Our results indicate that hippocampal expression of both 5LOX and COX2 increases rather promptly when glutamate receptors are stimulated by kainate. The mechanism of how NS-398 inhibits this action of kainate should be further investigated. PMID- 11113603 TI - Brain plasticity: to what extent do aged animals retain the capacity to coordinate gene activity in response to acute challenges. AB - The ability of the rodent brain to support plasticity-related phenomena declines with increasing age. A decreased coordination of genes implicated in brain plasticity may be one factor contributing to this decline. Synaptic rearrangement that occurs after seizure activity is regarded as a model of brain plasticity. In a rat model of seizure-related brain plasticity, we found that the induction of immediate-early genes, as exemplified by c-fos and tissue plasminogen activator (TPA) is not impaired in the aged rat brain. However, the aged rat brain responded more slowly to chemically induced seizure and the levels of c-fos and TPA mRNAs induction are decreased in the cortex and in the hippocampus of 30 month-old rats, as compared to the levels expressed by 3-month-old rats. In addition, at the peak induction the TPA transcripts were restricted to certain cortical layers of the older rats. Surprisingly, in applying the same experimental paradigm to late genes we found that there was a shift toward earlier times in the maximum expression of growth-related molecule, the microtubule-associated protein 1B (MAP1B) mRNA, which was very evident in 18 month-old rats. Aberrant immunolabeling of MAP1B occurred in cortical layer VI of the aged rats where, unlike in young rats, there was heavy staining of neuronal somata. These results suggest that (i) one consequence of aging, besides decreases in the levels of mRNA, is a progressive loss of coordination in gene activity following the administration of a stimulus; (ii) since c-fos, TPA and MAP1B have been implicated in neuronal plasticity, these findings could explain, in part, the limited plasticity of the aging brain. PMID- 11113604 TI - Circadian and sleep disturbances in the elderly. AB - The incidence of disturbed sleep is strongly increased in healthy and demented elderly. Age-related alterations in the circadian timing system appear to contribute strongly to these problems. With increasing age, a lack of input to the suprachiasmatic nucleus (SCN), the biological clock of the brain, may accelerate de-activation of neurons involved in the generation of 24-h rhythm or output of this rhythm. This process appears to be reversible, since supplementation of stimuli that impinge on the SCN can re-activate these neurons and ameliorate disturbances in the sleep-wake rhythm. PMID- 11113605 TI - Pineal and pituitary-adrenocortical function in physiological aging and in senile dementia. AB - The simultaneous evaluation of the circadian rhythm of plasma melatonin and ACTH and of serum cortisol and DHEAS represents a clinically reliable tool to appreciate the neuroendocrine changes occurring in physiological and pathological brain aging.A selective impairment of the nocturnal melatonin secretion has been observed in elderly subjects, being significantly related either to the age or to the severity of dementia. A significant increase of serum cortisol levels during evening- and night-times was found in elderly subjects, particularly if demented, when compared to young controls. Besides, both the circadian amplitude of cortisol rhythm and the nocturnal cortisol increase were significantly reduced in relation either to age or to cognitive impairment. By comparison to vascular dementia, patients with Alzheimer's disease exhibited the highest cortisol concentrations throughout the 24h. The sensitivity of the hypothalamic-pituitary adrenal axis to the steroid feedback was significantly impaired in old subjects and particularly in the demented ones. The serum DHEAS levels were significantly lower in elderly subjects and even more in demented patients than in young controls. Consequently, a significant increase of the cortisol/DHEAS molar ratio was evident when going from young controls to healthy elderly subjects and to demented patients. In conclusion, the aging process affects many neuroendocrine functions resulting in subtle but clinically relevant consequences; the occurrence of senile dementia seems to play an additive role. PMID- 11113606 TI - Aging and the neuroendocrine regulation of reproduction and body weight. AB - Aging in men is associated with a decline in trophic factors such as testosterone (T), alterations in body composition and impaired energy and body weight regulation. We performed studies to investigate the mechanisms underlying age related changes in the neuroendocrine control of testis function, body composition, food intake and body weight in the Brown Norway (BN) rat. We found that similar to aging men, male BN rats demonstrate both primary and secondary testicular failure with aging without confounding age-related tumors, hormonal changes and systemic illnesses. With aging, these animals have blunted circadian variations in luteinizing hormone (LH) and T, and decreased hypothalamic gonadotropin-releasing hormone (GnRH) synthetic capacity with preserved pituitary gonadotropin responses to GnRH. We found that aging male BN rats have increased peripheral and visceral adiposity associated with increased insulin and leptin levels, and decreased relative lean body mass and muscle mass. We found that these rats exhibit reduced food intake and body weight gain associated with decreased hypothalamic neuropeptide Y (NPY) gene expression in the arcuate nucleus (ARC), both during ad-libitum feeding and after a 72-h fast. Recently, we found that old male BN rats treated chronically with troglitazone, an insulin sensitizer, lowered high insulin and leptin levels, decreased body fat, and corrected the blunted food intake and body weight gain response to fasting without affecting basal ARC NPY gene expression. These findings suggested that hyperinsulinemia and/or hyperleptinemia associated with aging may contribute to the age-related impairment in energy and weight regulation. Our studies suggest that the aging male BN rat is an excellent model to investigate the mechanisms underlying the age-associated changes in the neuroendocrine control of body composition, energy intake and body weight. PMID- 11113607 TI - Aging of the male reproductive system. AB - Reproductive and sexual physiology, changes in body composition and mental performance in the aging male cannot simply be reduced to presumptive hypogonadism defined by low androgen serum levels or by decreasing levels of growth hormone (GH) and melatonin. Morphological changes in organs at different regulatory levels of hormonal networks governing, for example reproduction, such as diminished hypothalamic pulse generator mass, focal degeneration and loss of Leydig cells in testicular tissue, lead to diminished reserve capacities in production and to loss of coordinated pulsatile release of hypothalamic neuropeptides (e.g. gonadotropin releasing hormone, GnRH) and consequently diminished release of pituitary protein and glycoprotein hormones and testicular steroid hormones. Owing to presumptive alterations in feedback sensitivity, decreased testosterone levels do not necessarily upregulate pituitary LH secretion. Alternatively, increased serum levels of LH and FSH can be observed in old men either because of primary hypogonadism or to decreased hypothalamic opioid tone. In general, endocrine functions are sufficient to maintain fertility in elderly men because, except for sperm motility, quantitative and qualitative functional semen parameters are apparently not affected by age. Nevertheless, reduced endocrine and organic functions might become critical at different levels, with high inter-individual variability, of the hypothalamo/pituitary/gonadal-axis. One of the most intriguing organic manifestations of male aging is benign prostatic hyperplasia (BPH), the pathologic prevalence of which closely matches age. Age-associated changes in the endocrine system and in local networks of epithelial, stromal and luminal factors may play important roles in BPH development. PMID- 11113608 TI - Recent neuroendocrine facets of male reproductive aging. AB - We recently identified consistent attenuation of LH and testosterone secretory pulse amplitude and associated disruption of their orderly patterns of release in healthy older men. These dynamic changes emerge despite young-adult concentrations of LH and total testosterone. Moreover, we could document disruption of synchrony between LH secretion and oscillations in FSH, prolactin, sleep-stage and NPT (nocturnal penile tumescence), thus pointing to loss of coordinate neurohormone outflow. Such data suggest that CNS-hypothalamically based regulatory defects may be important in aging, as inferred indirectly in the old male rat and mouse more than 15 years ago. How such alterations are related to specific hypothalamic neurotransmitter changes in aging will be critical to unravel. PMID- 11113609 TI - Estrogen blocks inducible nitric oxide synthase accumulation in LPS-activated microglia cells. AB - Estrogens are thought to play a protective role against neurodegeneration through a variety of mechanisms including the activation of growth factors and neurotransmitter synthesis, the control of synaptic plasticity and functions, and the blockade of oxidative reactions. We here propose a novel mechanism to explain the neuroprotective effects of estradiol by showing that estrogens may antagonize nitric oxide synthase activity and reduce the accumulation of nitrites and nitrates consequent to various inflammatory stimuli. The potential anti inflammatory activity of estradiol is analyzed in vitro in cells in culture including primary cultures of microglia and in vivo in a well-known model of inflammation. PMID- 11113610 TI - Ovariectomy and 17beta-estradiol modulate the levels of Alzheimer's amyloid beta peptides in brain. AB - Alzheimer's disease (AD) is a neurodegenerative disorder characterized by accumulation of aggregated forms of the 40- and 42-amino acid Abeta peptides (Abeta40 and Abeta42). Estrogen replacement therapy (ERT) in postmenopausal women is associated with decreased risk for AD and/or delay in disease onset. The mechanism by which estrogen exerts this neuroprotective effect is elusive. 17beta estradiol (E2) was shown to reduce the release of Abeta peptides by primary neuronal cultures of murine and human origin. To test whether estrogen can modulate the metabolism of Abeta peptides in vivo, four experimental sets of guinea pigs were used: intact animals, ovariectomized animals, and ovariectomized animals that received E2 at two different doses. Ovariectomy was associated with a 1.5-fold average increase in total brain Abeta levels as compared to intact controls. E2 treatment significantly reversed the ovariectomy-induced increase in brain Abeta levels. The high-dose E2 treatment did not lead to further decrease in brain Abeta beyond the one observed with the low-dose E2 treatment. Our results infer that cessation of ovarian estrogen production in postmenopausal women might facilitate Abeta deposition by increasing the local concentrations of Abeta40 and Abeta42 peptides in brain and suggest that modulation of Abeta metabolism may be one of the ways by which ERT prevents and/or delays the onset of AD in postmenopausal women. PMID- 11113611 TI - A retinyl palmitate model of the phenomenon of the intrinsic fluorescence increase in ceroid-lipofuscin cytosomes. AB - We report here on a retinoid model of the phenomenon discovered in 1980, in which ceroid-lipofuscin cytosomes (CLC) increased their intrinsic fluorescence when exposed to fluorescence-exciting (lambda(ex)=365nm) ultraviolet (UV) light. We modeled this effect in vitro, irradiating a methanolic solution of retinyl palmitate (RP) either synthetic or extracted from the aged rat liver retinoid. Following our model, the mechanism of this phenomenon can be explained by the photodecomposition of RP during fluorescence. Palmitic acid (PA) that quenched fluorescence was separated by photohydrolysis and some products of RP photodegradation shifted absorption spectra towards the fluorescence-exciting UV band compared with the absorption spectrum of the initial RP. The total intensity of intrinsic fluorescence at 490nm of RP photodegradation products increased several times, because relative absorption at 365 nm wavelength was higher. We report here also the established chemical formulae of two retinoids, derivatives of RP photohydrolysis tentatively called R(368) and R(346). R(368) was determined as anhydroretinol, a naturally occurring intracellular messenger in the transduction pathway of retinoids regulating growth inhibition in lymphoid line cells. Retinoid R(346) was determined to be a methanolic intermediate of RP photodegradation (4,5-dihydro-5-methoxy-Anhydroretinol). PMID- 11113612 TI - Identification of novel genes in late-onset Alzheimer's disease. AB - Four genes affecting Alzheimer's Disease (AD)(AP, PS1, PS2, and APOE) have been identified and a fifth potential gene localized to chromosome 12. Collectively, these genes explain at most half of the genetic effect in AD. Understanding the genetics of AD is critical to developing new treatments. The quest to find the remaining AD genes led us to undertake a large genomic screen using over 466 families (730 affected sibpairs) in late-onset AD. In conjunction with this increase in power, we initiated several novel approaches to identify potential AD related genes. This included stratification of the data into an autopsy-confirmed subset of 199 AD families. Each of these targeted analyses resulted in the identification of novel regions containing potential AD genetic risk factors. Our most significant finding was on chromosome 9 in the autopsy-confirmed subset where we obtained an MLS of 4.31. These approaches, together with new methodologies such as conditional linkage analysis, generalized family-based association tests (PDT), and a new generation of genetic markers (SNPs), opens the door for additional AD gene discovery. Such strategies are necessary if we are to understand the subtle and complex threads that, woven together, create the intricate tapestry of AD. PMID- 11113614 TI - Brain glucose and energy metabolism abnormalities in sporadic Alzheimer disease. Causes and consequences: an update. AB - It is discussed that Alzheimer disease does not form a nosologic entity. 5 to 10% of all Alzheimer cases are due to inherited abnormalities on chromosomes 1, or 14, or 21, whereas the majority of 90-95% is sporadic in origin. Age-related changes in the composition of membranes and in glucose/energy metabolism along with a sympathetic tone in the brain are assumed to be cellular/molecular risk factors for this disease. In its pathogenesis, the desensitization of the neuronal insulin receptor similar to non-insulin dependent diabetes mellitus may be of pivotal significance. This abnormality along with a reduction in insulin concentration is assumed to induce a cascade-like process of disturbances including decreases in cellular glucose, acetylcholine, cholesterol, and ATP, associated with changes in the metabolism of amino acids and fatty acids. There is evidence that the reductions in the availability of both glucose/energy and insulin contribute to the formation of amyloidogenic derivatives and hyperphosphorylated tau protein. This may indicate that the amyloid cascade hypothesis in not valid for sporadic Alzheimer disease but that the formation of both, amyloidogenic derivatives and hyperphosphorylated tau protein is downstream the origin of this neurodegenerative disease. PMID- 11113613 TI - Candidate genes showing no evidence for association or linkage with Alzheimer's disease using family-based methodologies. AB - Alzheimer's disease (AD) is a genetically complex and heterogeneous disorder. To date, a large number of candidate genes have been associated with the disease, however none of these findings has been consistently replicated in independent datasets. In this study we report the results of family-based analyses for polymorphisms of five such candidates on chromosomes 2 (interleukin-1beta, IL 1B), 3 (butyrylcholinesterase, BCHE), 11 (cathepsin D, CTSD; Fe65, APBB1) and 12 (lipoprotein receptor-related protein-1, LRP1) that were all suggested to be associated with AD in recent case-control studies. To minimize the possibility of spurious findings due to population admixture, we used a family-based design applying the sibship disequilibrium test (SDT) as well as two-point parametric linkage analyses on families from the National Institute of Mental Health (NIMH) Genetics Initiative. Contrary to the initial reports, none of the polymorphisms that were analyzed showed evidence for association or linkage with AD in our families. Our results suggest that the previously reported associations from case control studies are either (a) false positive results, e.g. due to type I error or population admixture, (b) smaller than initially proposed, or (c) due to linkage disequilibrium with an as yet unidentified polymorphism nearby. PMID- 11113615 TI - The metabolism of plasma glucose and catecholamines in Alzheimer's disease. AB - Several lines of evidence suggest that the cholinergic system in the hippocampus plays a pivotal roll in regulating the peripheral metabolism of glucose and catecholamines. The injection of cholinergic stimulators including neostigmine, the acetylcholine esterase inhibitor, into the third ventricle or the hippocampus induces the elevation of glucose or catecholamines in plasma in rats. Under stress conditions, release of acetylcholine in the hippocampus increases, which coincides with the elevation of plasma glucose and catecholamines. Age-related reduction in responsivity of the cholinergic system in the hippocampus has been well documented. The intrahippocampal neostigmine injection induces significantly attenuated responses in plasma glucose and catecholamines in rats, the finding suggested that changes in cholinergic system activity in the hippocampus could result in alteration of the peripheral metabolism of glucose and catecholamines. In Alzheimer's disease (AD), the most common type of dementia, degeneration of the hippocampal cholinergic system is one of the most robust pathological features. Measurement of plasma catecholamines during a fasting state in the groups of AD subjects, vascular dementia subjects, and non-demented control subjects showed significantly lower plasma epinephrine levels in the AD subjects. PMID- 11113616 TI - Structural brain aging in inbred mice: potential for genetic linkage. AB - To identify genetic factors involved in brain aging, we have initiated studies assessing behavioral and structural changes with aging among inbred mouse strains. Cognitive performance of C57BL/6J mice is largely maintained with aging, and stereological analysis revealed no significant age-related change in neuron number, synaptic bouton number or glial number in the hippocampus. Moreover, no change in cortical neuron number and cholinergic basal forebrain neuron number has been found in this strain. 129Sv/J mice have more pronounced age-related cognitive deficits, although hippocampal and basal cholinergic forebrain neuron number also appear unchanged with aging. Differences in neurogenesis and neuron vulnerability in the adult CNS of C57BL/6, 129/Sv and other inbred strains have been reported, which in turn may have important consequences for brain aging. Age related lesions, such as thalamic eosinophilic inclusions and hippocampal clusters of polyglucosan bodies also vary greatly among inbred strains although the functional significance of these lesions is not clear. The continued assessment of such age-related structural and behavioral changes among inbred mouse strains offers the potential to identify genes that control age-related changes in brain structure and function. PMID- 11113617 TI - Mouse models of alpha-synucleinopathy and Lewy pathology. AB - The discovery of two missense mutations (A53T and A30P) in the gene encoding the presynaptic protein alpha-synuclein (alphaSN) that are genetically linked to rare familial forms of Parkinson's disease and its accumulation in Lewy bodies and Lewy neurites has triggered several attempts to generate transgenic mice overexpressing human alphaSN. Analogous to a successful strategy for the production of transgenic animal models for Alzheimer's disease we generated mice expressing wildtype and the A53T mutant of human alphaSN in the nervous system under control of mouse Thy1 regulatory sequences. These animals develop neuronal alpha-synucleinopathy, striking features of Lewy pathology, neuronal degeneration and motor defects. Neurons in brainstem and motor neurons appeared particularly vulnerable. Motor neuron pathology included axonal damage and denervation of neuromuscular junctions, suggesting that alphaSN may interfere with a universal mechanism of synapse maintenance. Thy1-transgene expression of wildtype human alphaSN resulted in comparable pathological changes thus supporting a central role for mutant and wildtype alphaSN in familial and idiopathic forms of diseases with neuronal alpha-synucleinopathy and Lewy pathology. The mouse models provide means to address fundamental aspects of alpha-synucleinopathy and to test therapeutic strategies. PMID- 11113618 TI - Nutraceutical interventions may delay aging and the age-related diseases. AB - Largely due to better control of infectious diseases and to year-round access to a more nutritious diet, life expectancy in developed countries has increased dramatically in the twentieth century. However, as the average age of the population has risen, the incidence of chronic age-related diseases such as Alzheimer's, Parkinson's, cardiovascular disease, cancer, arthritis, osteoporosis, benign prostatic hyperplasia, late-onset diabetes, and macular degeneration have increased. To obtain further significant improvements in both lifespan and the quality of life in this century, treatments and nutritional changes that address the age-related diseases and the aging process itself need to be examined and validated. There are many reports suggesting that oxidative stress and certain nutritional deficiencies may contribute to the aging process and to many age-related diseases. In this article, we report on two human clinical trials using novel antioxidant supplements in which urinary oxidative stress is significantly reduced. We also discuss the conceptual basis and existing literature for several nutritional supplements that may have beneficial effects on aging and age-related diseases. Based on the available data, we suggest that human life expectancy can be significantly increased in the twenty first century by optimizing diet and using nutritional supplements. PMID- 11113619 TI - Serotonin transporter gene polymorphisms in alcohol dependence. AB - The serotonin transporter (5-HTT) gene is a candidate gene in alcohol dependence because serotonin reuptake inhibitors (SRIs) can alleviate alcohol withdrawal. Studies of the 5-HTT gene in alcohol dependence have not resulted in a consensus. Recent studies have examined the transcriptionally active promoter polymorphism, a 44-bp deletion resulting in short (S) or long (L) alleles. In this study, 131 alcohol-dependent patients of Northern and Western European descent were genotyped. Seventy of these patients were diagnosed with alcohol dependence without comorbid disorders. Sixty-one patients were diagnosed with alcohol dependence comorbid with Tourette syndrome (alcoholic-TS). We found an excess of the S allele in alcohol-dependent patients (47%) compared with 125 ethnically matched controls (39%). A similar trend was found in 150 ethnically matched TS patients without alcohol dependence comorbidity (51%). However, the statistical significance of this trend in the data was not present after Bonferroni correction. The data presented suggests a trend toward increased frequency of the S promoter allele in alcohol-dependent, alcoholic-TS and TS patients. PMID- 11113620 TI - Acute tolerance during intravenous infusion of alcohol: comparison of performance during ascending and steady state concentrations--a pilot study. AB - Although acute tolerance (AT) to alcohol has been demonstrated in many single dose studies, the existence of AT at steady state concentrations of alcohol has been questioned. In the present study, six subjects were examined as (1) 7.5% alcohol or (2) placebo was administered intravenously (IV). The order of the infusions was randomized. The alcohol infusions were designed to result in similar blood alcohol concentrations at 20, 60, and 140 min (approximately 0. 7 per thousand). At 20 min, the concentrations were rising; the steady state (+/ 0.10 per thousand) was reached after 60 min and continued until 140 min. Three reaction time (RT) tests from the automated psychological test system were used (simple RT, two-choice RT, and two-choice RT with auditory inhibition). When the performance of the subjects was compared at rising and steady-state concentrations of alcohol, AT was shown for the most complex task requiring parallel processing, i.e., RT with failed inhibition, test. However, at steady state (i.e., 60 vs. 140 min), AT was not found for any of the tests. Further, the analysis showed that the test results of different individuals were related to their estimated normal alcohol consumption and that these differences presumably influenced the test results in accordance with our earlier findings. PMID- 11113621 TI - Prenatal alcohol exposure decreases the number of nitric oxide synthase positive neurons in rat superior colliculus and periaqueductal gray. AB - Because nitric oxide (NO) is involved in the development and refinement of axonal projections and synapses, it is of interest to know if developmental alcohol exposures affect NO producing neurons. Pregnant rats were fed artificial liquid diet throughout gestation as the only fluid or caloric source. The diet for experimental dams contained ethanol (6.7% v/v) while the pair-fed diet for control dams contained isocaloric maltose-dextrin instead of ethanol. This ethanol diet regime is known to produce peak blood alcohol concentrations of approximately 140 mg%. Cells stained histochemically for nitric oxide synthase (NOS) were counted at postnatal day 15 (P15) and 35 (P35) in cross-sections of the stratum griseum superficiale (SGS) of the superior colliculus (SC) and in the dorsolateral column of the periaqueductal gray (dlPAG). Compared to control tissues, alcohol caused the following effects: In the SC, the areal density of NOS+ neurons was decreased 24% at P15 but a similar decrease in means at P35 was not statistically significant (P=0.10); soma size was unaffected at either P15 or P35. In the dlPAG, both the areal density and the total number of NOS+ neurons per section were unaffected at P15 but were decreased at P35 (33% and 37% decreases); soma size was unaffected at either P15 or P35. The decrease in NOS+ neurons in the SC at P15 could be expected to have a negative impact on the refinement of neuronal connections while the decreases in NOS+ neurons in the dlPAG at P35 likely represent more permanent effects that could alter the function of that nucleus. PMID- 11113622 TI - The effect of antagonists selective for mu- and delta-opioid receptor subtypes on alcohol consumption in C57BL/6 mice. AB - Several studies have demonstrated that non-selective opioid receptor antagonists effectively reduce alcohol consumption in both animal models and at the clinical level. However, research examining the contribution of specific opioid receptor subtypes to this effect has yielded conflicting results. Some of these studies have shown that the effect is contingent upon the action of mu receptors while others have suggested that delta receptors are primarily responsible. The data reported here re-examine this question using the alcohol-preferring C57BL/6 mice. The results of this experiment demonstrate that D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen Thr-NH(2) (CTOP), a mu-selective antagonist, and naltrindole, a delta-selective antagonist, are equally effective at reducing alcohol consumption in a limited access model compared to a saline control group. While there was no specific comparison of the effects of these drugs on alternative appetitive behavior, neither of these drugs had effects on measured off-session food or water consumption. The results of this experiment suggest that alcohol consumption is mediated by both mu- and delta-opioid receptor subtypes. PMID- 11113623 TI - Alcohol differentially affects noradrenergic and purinergic responses in the bisected rat vas deferens. AB - This study investigates the effect of acute in vivo or in vitro ethanol administration on the adrenergic and purinergic responses in the epididymal and prostatic portion of rat vas deferens. Acute in vivo ethanol treatment (3.0 g/kg, i.p.) selectively impaired the response to noradrenaline in the prostatic portion of rat vas deferens, leaving unaffected the epididymal portion. In addition, the response evoked by the maximal concentration of alpha, beta-methylene-ATP was significantly depressed by acute in vivo ethanol treatment in both the epididymal and prostatic segments. Ethanol 50 mM in vitro was devoid of any action on the response to exogenous noradrenaline in both tissues. Ethanol in vitro left unaltered the response to alpha,beta-methylene-ATP in the epididymal portion, while potentiated the contractile response in the prostatic one. These results provide, for the first time, evidence that ethanol in vitro and in vivo differentially affects the noradrenergic and purinergic responses in the bisected vas deferens, suggesting that this substance may alter the male reproductive tract function. PMID- 11113624 TI - Neuronal subserving of behavior before and after chronic ethanol treatment. AB - We have previously shown that an acute ethanol dose (1 g/kg), sufficient to impair the performance of a healthy rabbit, also reversibly depresses the activity of those limbic-cortex neurons that are specifically activated during recently learned behavioral acts. Our new morphological and neurophysiological data suggest a death of such neurons after 9-month chronic ethanol treatment. The effect of acute ethanol administration on neurons and performance speed in alcoholic rabbits was opposite to that found in healthy animals. Our results help to understand why neurocognition of alcoholics changes and why acute low-level alcohol ingestion influences them differently than healthy individuals. PMID- 11113625 TI - Ethanol-induced increases in dopamine extracellular concentration in rat nucleus accumbens are accounted for by increased release and not uptake inhibition. AB - We performed a quantitative microdialysis study to determine whether the increase in dialysate dopamine from the nucleus accumbens caused by intraperitoneal administration of ethanol (1 g/kg) was due to enhanced dopamine release or inhibition of dopamine uptake. The Lonnroth method (no net flux), adapted for transient conditions, was used to follow the time course of true extracellular dopamine concentrations in the nucleus accumbens simultaneously with the in vivo recovery of dopamine across the microdialysis probe. Separate groups of rats were perfused with artificial cerebral spinal fluid containing 0, 4, 8, or 12 nM dopamine for the entire experiment. Samples were taken every 10 min. Each rat received a saline or an ethanol injection. The concentration of dopamine gained by or lost from the probe was plotted as a function of the concentration of dopamine perfused into the probe for each time point. Linear regression was used to determine the slope of the line (in vivo recovery) and the x-intercept (point of no net flux) for each plot. The in vivo recovery did not significantly change over time for the saline- or the ethanol-injected rats. However, the point of no net flux (true extracellular concentration of dopamine) significantly increased after the ethanol injection from 9.4+/-0.4 nM (mean of six basal samples) to 13.2+/-1.8 nM, at the maximum, but did not change after the saline injection. On the basis of these results, it is suggested that the primary mechanism by which ethanol increases dialysate dopamine levels in the nucleus accumbens after intraperitoneal administration is by increasing dopamine release from the terminals, rather than by inhibiting the dopamine transporter. PMID- 11113626 TI - Microsomal fatty aldehyde dehydrogenase catalyzes the oxidation of aliphatic aldehyde derived from ether glycerolipid catabolism: implications for Sjogren Larsson syndrome. AB - The enzyme that catalyzes the oxidation of fatty aldehyde derived from ether glycerolipid catabolism has not been identified. To determine whether microsomal fatty aldehyde dehydrogenase (FALDH) is responsible, we investigated the metabolism of 1-O-[9, 10-(3)H-octadecyl]-glycerol ([(3)H]OG) in FALDH-deficient cultured cells from patients with Sjogren-Larsson syndrome (SLS) and in mutant Chinese hamster ovary (CHO) cells. Intact fibroblasts from SLS patients incubated with [(3)H]OG showed a selective deficiency (38+/-7% of normal) in the incorporation of radioactivity into fatty acid, but no decrease in incorporation of radioactivity into fatty alcohol, total lipids and phosphatidylethanolamine (PE). Consistent with fatty aldehyde accumulation, incorporation of radioactivity into N-alkyl-phosphatidylethanolamine, which is derived from Schiff base formation of free aldehyde with PE, was 4-fold higher in SLS fibroblasts compared to normal controls. Similar results were seen with SLS keratinocytes, whereas FALDH-deficient CHO cells showed a more profound reduction in radioactive fatty acid to 12+/-2% of normal. These results implicate FALDH in the oxidation of ether-derived fatty aldehyde in human and rodent cells. Metabolism of ether glycerolipids is a previously unrecognized source of fatty aldehyde that may contribute to the pathogenesis of SLS. PMID- 11113627 TI - Influence of diabetes on cardiac nitric oxide synthase expression and activity. AB - There is some evidence that the endothelium dependent vasodilatation of coronary arteries is impaired in both types of diabetes. The underlying mechanisms are not yet clear, in particular whether this defect is caused by a direct effect of diabetes on the activity and the expression of nitric oxide synthases (NOS) or indirectly by an enhanced inactivation of nitric oxide. METHODS: To study this question we determined the activity (conversion of L-arginine to citrulline) and the mRNAs encoding the isoforms of NOS (using polymerase chain reaction after reverse transcription of the mRNAs into cDNAs by reverse transcriptase) in hearts of streptozotocin diabetic rats and in rat heart endothelial cells (RHEC). The formation of reactive oxygen intermediates (ROI) was measured by the dichloro dihydro-fluorescein method. RESULTS: The activity of total NOS and the amounts of mRNAs encoding ecNOS and iNOS were dependent on the duration of diabetes. After a diabetes duration of 4 to 6 weeks both the total activity as well as the mRNAs encoding ecNOS and iNOS were elevated. A reduction of NOS activity and the amounts of mRNAs of ecNOS and iNOS was only seen after a diabetes duration longer than 20 weeks, a time at which a loss of endothelium has been described. In RHEC, high glucose (22 mM) and H(2)O(2) (100 microM) were able to increase the mRNA encoding ecNOS, but not iNOS. This increase in ecNOS mRNA was inhibited by lipoic acid (1 microM). In addition, high glucose (22 and 30 mM) led to an enhanced formation of ROI and to activation of the transcription NF kappa B. CONCLUSION: These observations suggest that diabetes causes a temporary increase in NOS activity and ecNOS mRNA in the rat heart which is presumably the consequence of an enhanced oxidative stress exerted by hyperglycaemia. Together with previously published observations, our data suggest that the impairment of endothelium dependent vasodilatation in rat heart is not the consequence of a reduced activity and expression of NOS, but is caused by an enhanced inactivation of nitric oxide by ROI. PMID- 11113628 TI - Modification of the composition of polycystin-1 multiprotein complexes by calcium and tyrosine phosphorylation. AB - Mutations in the PKD1 gene are responsible for >85% of autosomal dominant polycystic kidney disease (ADPKD). The protein product of PKD1, polycystin-1, is a large, modular membrane protein, with putative ligand-binding motifs in the extracelluar N-terminal portion, 9-11 transmembrane domains and an intracellular C-terminal portion with phosphorylation sites. A role for polycystin-1 as a cell surface receptor involved in cell-matrix and cell-cell interactions has been proposed. In this study, we have analyzed polycystin-1 and associated protein distribution in normal human epithelial cells and examined the role of cell matrix versus cell-cell interactions in regulation of the assembly of polycystin 1 multiprotein complexes. Immunocytochemistry, sucrose density gradient sedimentation, co-immunoprecipitation analyses and in vitro binding assays have shown that polycystin-1 associates with the focal adhesion proteins talin, vinculin, p130Cas, FAK, alpha-actinin, paxillin and pp60c-src in subconfluent normal human fetal collecting tubule (HFCT) epithelia when cell-matrix interactions predominate. Polycystin-1 also forms higher S value complexes with the cell-cell adherens junction proteins E-cadherin, beta- and gamma-catenins in confluent cultures when cell-cell interactions are predominant. Polycystin-1 multiprotein complexes can be disrupted by cytochalasin D but not by colchicine, suggesting involvement of the actin cytoskeleton. Although inhibition of tyrosine phosphorylation by tyrphostin inhibits polycystin-1-FAK interactions, E-cadherin interactions are enhanced. High calcium treatment also increases polycystin-1-E cadherin interactions. PMID- 11113629 TI - Sustained elevation of norepinephrine depresses hepatocellular function. AB - The sympathetic-adrenal system is rapidly activated in response to sepsis or hemorrhagic shock, resulting in an increase in circulating levels of catecholamines. Although it has been shown that the occurrence of hepatocellular dysfunction under such conditions is associated with an early and sustained increase in plasma norepinephrine (NE), it remains unknown whether the increased NE per se plays any role in producing hepatocellular dysfunction. To study this, exogenous NE was administered, by implantation of a peritoneal mini-osmotic pump (consistently releasing NE), to produce a plasma level of NE similar to that observed after sepsis or hemorrhage. At 24 h after the pump implantation, cardiac output was determined by dye dilution technique and hepatocellular function [i.e., the maximal velocity (V(max)) and the efficiency of the transport (K(m)) of in vivo indocyanine green clearance) was assessed without blood sampling. In addition, tissue perfusion in various organs was determined using laser Doppler flowmetry. Plasma levels of interleukin-6 were measured by bioassay and liver enzymes were assayed enzymatically. The results indicate that sustained (24 h) elevation of plasma levels of NE caused a decrease in cardiac output and microvascular blood flow in the liver, spleen, and small intestine. In addition, the increase in plasma NE produced significant depression in hepatocellular function as evidenced by reduced V(max) and K(m). This was associated with elevated plasma levels of liver enzymes, indicating hepatocyte injury. In addition, plasma levels of interleukin-6 increased significantly. These findings suggest that sustained elevated levels of NE, observed after sepsis or hemorrhagic shock, may play an important role in producing hepatocellular dysfunction and altered hepatocyte integrity. PMID- 11113630 TI - A role for linoleic acid in erythrocytes infected with Plasmodium berghei. AB - Unesterified fatty acids were measured in mouse erythrocytes infected either with chloroquine-susceptible (CS) or with chloroquine-resistant (CR) lines of Plasmodium berghei. This work was undertaken to identify candidates for the lipid involved in ferriprotoporphyrin IX (FP) polymerization. Linoleic, oleic, palmitic, and stearic acids were quantified by gas chromatography/mass spectrometry. In total, they increased 4-fold with CS infections and 6-fold with CR infections. Treating infected mice with chloroquine did not affect the amounts of unesterified fatty acids in erythrocytes. Of the four fatty acids, only linoleic acid increased disproportionately to the total. It increased 16-fold for the CS line and 35-fold for the CR line. The method could detect monoglycerides but they were below the limit of detection. It could not detect diglycerides, triglycerides or phospholipids. Triglycerides and phospholipids have been tested previously, however, and found to be ineffective at promoting FP polymerization. Therefore, other than linoleic acid, the lipids most likely to be involved in FP polymerization are diglycerides. We tested dilinoleolyglycerol in the present work and found it to be an effective promoter of FP polymerization. These results suggest that linoleic acid or a diglyceride containing it has the critical role of promoting FP polymerization in malaria parasites. PMID- 11113631 TI - CINC blockade prevents neutrophil Ca(2+) signaling upregulation and gut bacterial translocation in thermal injury. AB - In this study, we have evaluated the role of cytokine-induced neutrophil chemoattractant (CINC), in the upregulation of neutrophil Ca(2+) signaling in neutrophils from thermally injured rats treated with anti-CINC antibody. Additionally, we have determined the effect of the treatment with CINC antibody on the accumulation of activated neutrophils in the intestinal wall, and the effect of such accumulation on gut bacterial translocation. Measurements of myeloperoxidase (MPO) activity and immunohistochemical localization of neutrophils determined neutrophil sequestration in the rat intestine. Agar culture analyses and a specific Escherichia coli beta-galactosidase gene polymerase chain reaction was carried out to detect gut indigenous bacterial invasion into intestinal wall and extraintestinal mesenteric lymph nodes (MLN). The results showed that pretreatment of rats with anti-CINC antibody attenuated the thermal injury-induced enhancement in [Ca(2+)](i) responses in neutrophils both in the basal and Formyl-Met-Leu-Phe stimulated conditions. Moreover, treatment with the CINC antibody decreased neutrophil infiltration into the gut and attenuated thermal injury-caused translocation of bacteria into the MLN. PMID- 11113632 TI - Hippocampal level of neural specific adenylyl cyclase type I is decreased in Alzheimer's disease. AB - Previous studies reported disruption of adenylyl cyclase (AC)-cyclic AMP (cAMP) signal transduction in brain of Alzheimer's disease (AD). We also demonstrated that basal and stimulated AC activities in the presence of calcium and calmodulin (Ca(2+)/CaM) were significantly decreased in AD parietal cortex. In the present study, we examined the amounts of Ca(2+)/CaM-sensitive types I and VIII AC, and Ca(2+)/CaM-insensitive type VII AC in the postmortem hippocampi from AD patients and age-matched controls using immunoblotting. The specificities of the anti-type VII and VIII AC antibodies were confirmed by preabsorption with their specific blocking peptides. We observed a significant decrease in the level of type I AC and a tendency to decrease in the level of type VIII AC in AD hippocampus. On the other hand, the level of type VII AC showed no alteration between AD and controls. A body of evidence from the studies with invertebrates and vertebrates suggests that types I and VIII AC may play an essential role in learning and memory. Our finding thus firstly demonstrated that a specific disruption of the Ca(2+)/CaM-sensitive AC isoforms is likely involved in the pathophysiology in AD hippocampus. PMID- 11113633 TI - Does ethanol metabolism affect erythrocyte hemolysis? AB - The effects of ethanol and acetaldehyde on the hemolytic stability of rabbit erythrocytes have been compared. Incubation of normal erythrocytes with ethanol facilitated both acidic and oxidative hemolysis and increased the percentages of cells that were hemolyzed at maximal rate. Acetaldehyde exerted a similar destabilizing effect on erythrocytes only in the case of oxidative hemolysis. The destabilizing effect of ethanol was observed in catalase-inactivated erythrocytes under acidic, but not oxidative, hemolysis conditions. It is concluded that the destabilizing effect of unmetabolized ethanol occurs under conditions of acidic hemolysis, whereas the destabilizing effect of the oxidation of ethanol to acetaldehyde takes place only under the conditions of oxidative hemolysis. PMID- 11113635 TI - Microdialysis for the evaluation of penetration through the human skin barrier - a promising tool for future research? AB - The direct measurement of local drug concentration levels at discreet skin locations with minor trauma has recently become possible with the introduction of cutaneous microdialysis. Cutaneous microdialysis is an in vivo sampling technique for measuring solutes in the extracellular fluid of the dermis. When used in combination with other experimental approaches, for example with a variety of non invasive techniques to describe the functional status of the skin (bioengineering methods), it may help investigators to gain new insights into the fields of skin diseases, metabolism and drug absorption/penetration. An important parameter to describe the efficacy of microdialysis is the relative recovery. This is the ratio between the concentration of a substance in the dialysate and the true extracellular concentration. Several methods are in common use to describe the relative recovery (no-net-flux method or retrodialysis). Parameters such as probe design, depth of the probe in the dermis, physico-chemical properties of the compound of interest, and analytical aspects are important factors influencing microdialysis. Microdialysis has been used to investigate the influence of penetration enhancers, vehicles or iontophoresis on percutaneous absorption, performed by in vivo studies in rats. In human volunteers, most of the experiments have been performed to study the kinetics of fast penetrating substances, e.g. nicotine, non-steroidal antiinflammatory drugs, local anaesthetics, or solvents. Problems have been encountered in the detection of lipophilic and highly protein-bound substances. Further, dermal metabolism and the influence of barrier perturbation on percutaneous absorption have been analyzed. Investigations suggest that microdialysis, in combination with traditional techniques, might give valuable information regarding the assessment of the penetration of drugs and other exogenous agents through the skin. In spite of the clearly defined and accepted advantages of microdialysis technology for studies of transdermal drug delivery, to date no standardized test procedure exists nor has the reproducibility of the results been evaluated. In the future, these problems have to be solved to enable this method to find its place in standard research. PMID- 11113634 TI - Stimulation of cannabinoid receptor agonist 2-arachidonylglycerol by chronic ethanol and its modulation by specific neuromodulators in cerebellar granule neurons. AB - In an earlier study, we reported that chronic ethanol (EtOH) stimulates the formation of anandamide in human SK-N-SH cells. In the present study, we investigated the effect of chronic EtOH on the formation of yet another cannabinoid receptor (CB1) agonist, 2-arachidonylglycerol (2-AG), in cerebellar granule neurons (CGNs). The formation of 2-[(3)H]AG without any stimulation was more pronounced in the older cultures than in younger cultures. Exposure of CGNs to EtOH led to a significant increase in the level of 2-[(3)H]AG (P<0.05). Incubation with the anandamidehydrolase inhibitor phenylmethylsulfonyl fluoride and EtOH did result in an additive increase in 2-[(3)H]AG, but did not with E-6 (bromomethylene)tetrahydro-3-(1-naphthelenyl)-2H-pyran-2-one. The formation of 2 [(3)H]AG was enhanced by ionomycin in both the control and EtOH-exposed CGNs, and the ionomycin-stimulated 2-[(3)H]AG synthesis was inhibited by the intracellular chelating agent 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Further, glutamate increased the formation of 2-[(3)H]AG only in control CGNs. MK 801 inhibited the EtOH-induced 2-[(3)H]AG synthesis, suggesting the participation of intracellular Ca(2+) in EtOH-induced 2-[(3)H]AG synthesis. The dopamine receptor (D2) agonist did not modify the 2-AG synthesis in either the control or EtOH-exposed CGNs. However, the D2 receptor antagonist inhibited the EtOH-induced formation of 2-[(3)H]AG. The EtOH-induced 2-[(3)H]AG formation was inhibited by SR141716A and pertussis toxin, suggesting the CB1 receptor- and Gi/o-protein mediated regulation of 2-AG. The observed increase in 2-AG level in CGNs is possibly a mechanism for neuronal adaptation to the continuous presence of EtOH. These findings indicate that some of the pharmacological actions of EtOH may involve alterations in the endocannabinoid signaling system. PMID- 11113636 TI - Submicron emulsions as drug carriers. Studies on destabilization potential of various drugs. AB - Few drugs have been successfully formulated as submicron emulsions. The physicochemical stability of submicron emulsions on admixture of drugs is the main factor limiting a wider use of this type of vehicle for parenteral or other routes of administration. In a short-term stability studies we have investigated 35 drugs with regard to their compatibility with 20 or 5% parenteral fat emulsions. The study has shown that interaction of drugs with submicron emulsions is complex in nature. Taking into consideration the physicochemical properties of a drug e.g. lipophilicity or ionization, it is difficult to predict changes in the physical stability of the system. If destabilization occurs it is maximal at saturated concentrations and the presence of even significant amounts of the solid phase formed by undissolved drug does not influence the short-term stability of the system. Due to the increased ratio of emulgator to oil, emulsions containing 5% of oily phase are less susceptible to destabilization than 20% emulsions. PMID- 11113637 TI - A study of the binding requirements of calyculin A and dephosphonocalyculin A with PP1, development of a molecular recognition model for the binding interactions of the okadaic acid class of compounds with PP1. AB - The interactions of the okadaic acid class of compounds, with special emphasis on the solution structures of calyculin A and dephosphonocalyculin A with PP1 are reported. After examination of the interactions of all docked structures, a receptor based pharmacophore model for the interactions of the protein phosphatase inhibitors has been developed. Calyculin A or dephosphonocalyculin A can interact with the enzyme in either a manner similar to the reported crystal structure, or in an extended form. The inhibitors require two essential regions interacting with the hydrophobic region and the central metal binding regions of the enzyme. This simplified model is consistent with previously published models of the okadaic acid class of compounds with PP1. PMID- 11113638 TI - Capsaicin and nonivamide as novel skin permeation enhancers for indomethacin. AB - The study was conducted in vitro to investigate the changes of indomethacin transdermal permeation pretreated by capsaicin and nonivamide, two compounds chemically similar to Azone. The combined effect of low frequency ultrasound (20 kHz) and enhancers on the indomethacin permeation was also evaluated. The experimental data demonstrated that capsaicin and nonivamide significantly enhanced the flux of indomethacin across nude mouse skin. Enhancement effects of both analogues were very similar and depended predominantly on the concentration tested. Histological examination coupled with visual scores indicated the safety of capsaicin and nonivamide on skin structure. Simultaneous application of ultrasound and enhancers significantly increased skin permeation of indomethacin compared with either ultrasound or enhancers alone. Better effect was obtained by the combination with capsaicin than nonivamide. PMID- 11113639 TI - Evaluation of a vincristine resistant Caco-2 cell line for use in a calcein AM extrusion screening assay for P-glycoprotein interaction. AB - AIM: To develop a fast fluorometric screening assay based on vincristine resistant Caco-2 cells (Caco-2VCR) in order to elucidate potential P-glycoprotein (Pgp) interactions of compounds, and to characterise Caco-2VCR cells with regard to their expression of the efflux transporters Pgp, MRP1 and MRP2. METHODS: We applied the Caco-2VCR cells to a 96-well plate-based calcein AM extrusion assay. The Caco-2VCR cells were cultured as monolayers and incubated with calcein AM with/without addition of Pgp modulators. Fourteen known Pgp modulators were tested in the assay (chloropromazine, cyclosporin A, domperidone, digoxin, ivermectin, ketoconazole, loperamide, metoprolol, propranolol, progesterone, quinidine, quinine, verapamil and vincristine). For each compound an EC50 value was calculated. Protein and mRNA levels of the efflux transporters were analysed by Western blot and polymerase chain reaction techniques. RESULTS: All compounds with the exception of digoxin displayed increased calcein levels. Protein and mRNA analysis showed increased levels of Pgp after vincristine exposure, while expression of the efflux transporters MRP1 and MRP2 remained unchanged. CONCLUSIONS: The calcein AM extrusion assay applied to Caco-2VCR cells can be a valuable tool as a screening assay for new compounds and their potential interaction with P-glycoprotein. PMID- 11113640 TI - Establishment and functional characterization of an in vitro model of the blood brain barrier, comprising a co-culture of brain capillary endothelial cells and astrocytes. AB - OBJECTIVE: The aim was to establish a flexible, abundantly available, reproducible and functionally characterized in vitro model of the blood-brain barrier (BBB). METHODS: In a first step, bovine brain capillaries and newborn rat astrocytes were isolated. Subsequently, a co-culture of primary brain capillary endothelial cells (BCEC) on semi-permeable filter inserts, with astrocytes on the bottom of the filter was established. The cell material was characterized on the basis of specific cell-type properties and (functional expression of) specific BBB properties. RESULTS: BCEC displayed: (1) characteristic endothelial cell morphology; (2) expression of endothelial cell markers (i.e., CD51, CD62P, CD71 and cadherin 5); (3) marginal F-actin localization; (4) tight junction formation between the cells; (5) expression of gamma-glutamyl-transpeptidase (gamma-GTP); (6) expression of P-glycoprotein (Pgp); (7) functional transendothelial transferrin transport and uptake; (8) restriction of paracellular transport; and (9) high transendothelial electrical resistance (TEER). Astrocytes displayed characteristic astrocyte morphology and expressed glial fibrillary acidic protein (GFAP). Co-culture with astrocytes increased TEER and decreased paracellular transport. In addition, expression of the glucocorticoid receptor (GR) was demonstrated in the endothelial cells of the BBB, while no expression of the mineralocorticoid receptor (MR) was found. CONCLUSIONS: A high quality and mass production in vitro BBB model was established in which experiments with physiological (e.g., regulation of BBB permeability), pharmacological (e.g., pharmacokinetics and pharmacodynamics) and pathophysiological (e.g., disease influence on BBB permeability) objectives can be reproducibly performed. PMID- 11113641 TI - Determination of the passive absorption through the rat intestine using chromatographic indices and molar volume. AB - Immobilized artificial membrane (IAM) chromatography coupled to physicochemical descriptors was evaluated to model the passive intestinal absorption of drugs through rat gut sacs. The chromatographic capacity factors (logk'(IAM)) of 12 structurally diverse compounds were determined on a IAM PC DD2 column. The passive permeabilities (P(a)) of the drugs were determined through rat everted gut sacs or through non-everted sacs for actively transported molecules. Correlation studies between logk'(IAM), physicochemical descriptors and P(a) were conducted by stepwise multiple linear regression (MLR) and back-propagation neural network (BPNN). MLR and BPNN showed that logk'(IAM) was the descriptor which correlated best with P(a). Considering the molar volume as an additional descriptor, the correlation was improved. Retention indices on IAM and the molar volume can be used concurrently to predict passive drug absorption. PMID- 11113642 TI - Biochemical and immunohistological changes in the brain of 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine (MPTP)-treated mouse. AB - We investigated neurochemically and neuropathologically the utility of 1-methyl-4 phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice as a model of Parkinson's disease. The changes in dopamine D1 and D2 receptors and dopamine uptake sites were determined by quantitative autoradiography using [3H]SCH23390, [3H]raclopride and [3H]mazindol, respectively. Dopamine and 3,4-dihydroxyphenyl acetic acid (DOPAC) contents in the striatum were measured by high-performance liquid chromatography. The distribution of nigral neurons and reactive astrocytes was determined by immunohistochemical staining with antibody against tyrosine hydroxylase (TH) and glial fibrillary acidic protein (GFAP). The mice received four intraperitoneal injections of MPTP (10 mg/kg) at 1-h intervals and then the brains were analyzed at 3 and 7 days after the treatments. No significant change in dopamine D1 receptors was observed in the striatum and substantia nigra after acute treatment with MPTP. Dopamine D2 receptors were reduced significantly in the substantia nigra only 7 days after the MPTP treatment, whereas striatum showed no significant change in the binding throughout the experiments. In contrast, dopamine uptake sites were reduced markedly in the striatum and substantia nigra 3 and 7 days after the MPTP treatment. Dopamine and DOPAC content were also reduced in the striatum 3 and 7 days after the MPTP treatment. An immunohistochemical study indicated a loss of the number of TH-positive neurons in the substantia nigra 7 days after the MPTP treatment. In contrast, numerous GFAP-positive astrocytes were evident in the striatum 7 days after the MPTP treatment. These results provide valuable information for the pathogenesis of acute stage of Parkinson's disease. PMID- 11113643 TI - Direct estimation of the in vivo dissolution of spironolactone, in two particle size ranges, using the single-pass perfusion technique (Loc-I-Gut) in humans. AB - AIM: The objective of this in vivo dissolution study was to investigate the usefulness of the Loc-I-Gut technique for differentiating between the in vivo dissolution rate of two particle sizes of spironolactone, and to compare these in vivo results with corresponding in vitro data. METHODS: The study included six volunteers, and consisted of three sequential parts (I, II, III). In parts I and III the in vivo dissolution was measured directly by perfusing a semi-open segment in the proximal jejunum. In part II, a solution of spironolactone was administered orally, and the plasma concentration time profile was followed for 48 h. The in vitro dissolution was measured using flow-through cells and different dissolution media simulating human gastrointestinal fluids. RESULTS: A difference in in vivo dissolution rate of the two different particle sizes was observed, based on perfusion data. This difference was not pronounced in the relative bioavailability of spironolactone administered in two different particle sizes. The relative bioavailability was dependent on the bile acid concentration in vivo. In vitro, dissolution rate of the smaller particles was improved at fasted state bile acid concentrations, while the larger particles were only significantly affected at fed state bile acid concentrations. CONCLUSION: In vivo dissolution studies discriminated between the dissolution rate of the two different particle sizes of spironolactone, based on the perfusate samples. The lack of difference in relative bioavailability, might be explained by the insufficient wash-out of particles after ending the perfusion, reabsorption of surface active ingredients along the GI tract, relatively small difference in particle size and the large inter- and intra-individual differences in pharmacokinetic variables. PMID- 11113644 TI - Ganciclovir-loaded albumin nanoparticles: characterization and in vitro release properties. AB - Ganciclovir is one of the most widely used antiviral drug for the treatment of cytomegalovirus retinitis. Due to its short half-life in the vitreous, frequent administrations are necessary to maintain the therapeutic levels. In this context, the aim of this study was to characterise and in vitro evaluate the drug release properties of three different formulations of ganciclovir-loaded albumin nanoparticles. These carriers were prepared by a coacervation method and chemical cross-linking with glutaraldehyde. Depending on the step where the drug and/or cross-linking agent were added three different formulations were obtained, named models A, B and C. For model A nanoparticles, ganciclovir was incubated with the just-formed albumin nanoparticles. For the other two types of nanoparticulate formulations, the drug was added to a solution of albumin (model B) and glutaraldehyde (model C) prior to the formation of the carriers by coacervation. In all cases, the size of the different nanoparticulate formulations was comprised between 200 and 400 nm and the yield ranged from 50%, in model A, to 65% in model B. Concerning the ganciclovir loading, model B nanoparticles offered the higher capacity to carry this antiviral drug (around 30 microg ganciclovir/mg nanoparticle). On the contrary, the drug loading calculated for model A nanoparticles was only 14.6 microg/mg. The in vitro release profiles of the nanoparticles showed a biphasic pattern, with an initial and rapid release, followed by a slower step for up 5 days. This burst effect was especially relevant in model A (around 60% in 1 h), followed by model B (40%) and less important in model C (20%). The addition of trypsin to the release medium did not have a significant influence on the release characteristics. However, the release of the drug was increased in acidic or basic mediums, due to the disruption of the covalent binding between ganciclovir and the protein matrix via glutaraldehyde. This strong linkage was also confirmed by TLC experiences. In summary, a first step of incubation between the drug and the protein, prior to the preparation of nanoparticles, enabled us to obtain albumin carriers able to release ganciclovir in a sustained way. PMID- 11113645 TI - Physical stabilisation of amorphous ketoconazole in solid dispersions with polyvinylpyrrolidone K25. AB - The glass forming properties of ketoconazole were investigated using differential scanning calorimetry (DSC), by quench cooling liquid ketoconazole from T(m)+10 to 273.1 K, followed by subsequent heating at 5 K/min to T(m)+10 K. It was shown that liquid ketoconazole forms a glass which did not recrystallise following reheating, indicating its stability; T(g) was found to be 317.5+/-0.3 K. However, the presence of a small amount of crystalline ketoconazole was able to convert the amorphous drug back to the crystalline state: the addition of only 4.1% (w/w) of crystalline material converted 77.1% of the glass back to the crystalline state, and this value increased as the amount of added crystals increased. PVP K25 was found to be highly effective in the prevention of such recrystallisation, but only if the amorphous drug was formulated in a solid dispersion, since physical mixing of amorphous ketoconazole with the polymer resulted in recrystallisation of the former compound. Storage of the solid dispersions for 30 days at 298.1 K (both 0 and 52% RH) in the presence or absence of crystals did not result in recrystallisation of the amorphous drug. Solid dispersions formed compatible blends as one single T(g) was observed, which gradually increased with increasing amounts of PVP K25, indicating the anti-plasticising property of the polymer. The values of T(g) followed the Gordon-Taylor equation, indicating no significant deviation from ideality and suggesting the absence of strong and specific drug-polymer interactions, which was further confirmed with 13C NMR and FT-IR. It can be concluded therefore that the physical mechanism of the protective effect is not caused by drug-polymer interactions but due to the polymer anti-plasticising effect, thereby increasing the viscosity of the binary system and decreasing the diffusion of drug molecules necessary to form a lattice. PMID- 11113646 TI - Cause of high variability in drug dissolution testing and its impact on setting tolerances. AB - Considering a variable mixing/stirring and flow pattern in a drug dissolution vessel as a likely source of high variability in results, experiments were conducted using USP paddle apparatus by placing (aligned to the walls) a metal strip (1.7 mm thickx6.4 mm wide) in a dissolution vessel. The metal strip forces the undisintegrated tablet to settle about 3 mm away from the centre, facilitates spread of disintegrated material and diminishes the cone formation at the bottom of the vessel. To assess the impact of this altered environment in the vessel, but still maintaining the vessel dimensions within required specifications, drug release characteristics were evaluated for products having different formulation/manufacturing attributes. Tests were conducted with calibrator tablets (USP prednisone and salicylic acid tablets and FDA proposed NCDA No. 2 prednisone tablets) and two commercially available products (250 mg amoxicillin capsules and 5 mg glibenclamide tablets). Except for the glibenclamide tablet product, all products gave significantly (P<0.01) higher dissolution results with vessels containing metal strip than without. The extent of increased dissolution with the metal strip varied from product to product i.e. USP prednisone tablet was the smallest (14.4%) and NCDA No. 2 was the largest (88.4%). Based on the results obtained from this study, it is concluded that employing the current apparatuses, in many cases products will provide lower than anticipated results which may not be reflective of the product drug release characteristics. Test-to test variability, within or between laboratories, can also be very high depending on the settling position of the product once dropped in the vessel and/or due to slight aberration in the walls of the vessel by altering the extent of spread of disintegrated material at the bottom of the vessel. Thus, dissolution testing will require wider tolerances to be useful for comparison of batch-to-batch or interlaboratory results. PMID- 11113647 TI - Synthesis and binding affinity of cis-(-)- and cis-(+)-N-ethyleneamino-N nordeoxymetazocine and cis-(-)-N-normetazocine analogues at sigma1, sigma2 and kappa opioid receptors. AB - The synthesis of cis-(+)- and cis-(-)-N-ethyleneamino-N-nordeoxymetazocine and cis-(-)-N-normetazocine analogues is described and their affinities to sigma1, sigma2 and kappa opioid receptors are evaluated. The cis-(+)-deoxy compounds displayed high sigma/kappa selectivity with nanomolar K(i) values for sigma1 receptors, whereas in the cis-(-)-N-normetazocine series the compound (-)-7b was found to bind with nanomolar affinity to the kappa opioid receptor (K(i)=21.5 nM). Compound (-)-7b showed good selectivity for the kappa opioid receptor in comparison to the sigma1 and sigma2 sites and to the mu and delta opioid receptors. A correlation of the binding affinities between cis-(-)- and cis-(+)-N deoxynormetazocine derivatives show that both isomers of the deoxy analogs have similar sigma1 and sigma2 binding profiles as the cis-(+)-N-normetazocine derivatives. PMID- 11113648 TI - The possibility of achieving an interactive mixture with high dose homogeneity containing an extremely low proportion of a micronised drug. AB - The homogeneity of binary mixtures containing coarse particulate mannitol and three different size fractions of sodium salicylate particles was investigated. The proportion of sodium salicylate (0.15% w/w and 0.015% w/w) and the sample size of the mixture were also varied. The homogeneity of these experimental mixtures was compared with the theoretical values that would be expected from ordered, adhesive interactive (pseudo-random) and random model mixtures. The results indicated that adhesive interactive forces were created between the drug and the carrier particles in most of the experimental mixtures, since their homogeneity tended to be higher than that of a theoretical randomised mixture. In fact, there was some resemblance to an ideal ordered state in mixtures containing the highest drug proportion (0.15%) and the smallest particles. However, the effect of sample size was not negligible and it was concluded that the number of drug particles interacting with individual carrier particles was not constant but was in fact related to a process of randomisation. The homogeneity of the mixture was predicted using two approaches: the ratio of the number of drug to carrier particles in the mixture and the absolute number of drug particles, estimated by weight, in the sample. It is suggested that calculating the relative numbers of particles by weight in the mixture could aid in predicting the feasibility of achieving an interactive powder mixture with a high dose-homogeneity. PMID- 11113649 TI - Effects of interactions between powder particle size and binder viscosity on agglomerate growth mechanisms in a high shear mixer. AB - A study was performed in order to elucidate the effects of the interactions between powder particle size and binder viscosity on the mechanisms involved in agglomerate formation and growth. Calcium carbonates having mean particle sizes in the range of 5-214 microm and polyethylene glycols having viscosities in the range of approximately 50-100000 mPas were melt agglomerated in a high shear mixer. Agglomerate growth by nucleation and coalescence was found to dominate when agglomerating small powder particles and binders with a low viscosity. Increasing the binder viscosity increased the formation of agglomerates by immersion of powder particles in the surface of the binder droplets. With a larger powder particle size, an increasing binder viscosity was necessary in order to obtain an agglomerate strength being sufficient to avoid breakage. Due to a low agglomerate strength, a satisfying agglomeration of very large particles (214 microm) could not be obtained, even with very viscous binders. The study demonstrated that the optimum agglomerate growth occurred when the agglomerates were of an intermediate strength causing an intermediate deformability of the agglomerates. In order to produce spherical agglomerates (pellets), a low viscosity binder has to be chosen when agglomerating a powder with a small particle size, and a high viscosity binder must be applied in agglomeration of powders with large particles. PMID- 11113650 TI - In vitro study on fluoxetine adsorption onto charcoal using potentiometry. AB - This in vitro investigation was performed to study the adsorption rate constant as well as the adsorption characteristics of fluoxetine (F) to activated charcoal and its commercial formulation Carbomix powder in simulated gastric (pH 1.2) fluid environment. Ion-selective electrode (ISE) potentiometry, based on the selective, direct and continuous monitoring of F with an F-ISE constructed in our laboratory was used. The method used in the kinetic experiments consists of the rapid addition of a slurry containing the charcoal into the drug solution under stirring and continuous recording of the F-ISE potential until the establishment of equilibrium. The free ionized drug concentration at appropriate time intervals was calculated from the recorded adsorption curve and the apparent adsorption rate constant was estimated assuming pseudo first order kinetics. Within run R.S.D. of the estimates ranged from 0.24 to 11.5%, while between run R.S.D. (n=3 4) ranged from 0.90 to 13.8%. A linear relationship was found between the apparent adsorption rate constants and the amount of charcoal used with slopes (+/-S.D.) for activated charcoal and Carbomix equal to 1.14(+/-0.21) and 0.146(+/ 0.009) s(-1)g(-1), respectively. Successive additions of microvolumes of F solution were made into a charcoal slurry with measurement of the F-ISE potential at equilibrium. The maximum adsorption capacity values (+/-S.D.) of activated charcoal and Carbomix were 254.8+/-1.8 and 405+/-41 mg/g, respectively while the affinity constant values (+/-S.D.) were 45.6+/-2.2 and 55.5+/-2.9 l/g, respectively. The adsorption of F to charcoals was rapid and for amounts of charcoal 10 times greater than the amount of the drug, 95% of F was adsorbed within the first 5 min. Relative to the toxic and lethal doses in cases of F intoxications, both types of charcoals tested adsorbed effectively F at gastric pH. Carbomix can be considered as appropriate charcoal formulation for medical treatment in cases of F poisoning. PMID- 11113651 TI - Measuring the water retention capacities (MRC) of different microcrystalline cellulose grades. AB - The ability of various types of microcrystalline cellulose (MCC) to hold water when subjected to an applied force has been assessed by a centrifuge technique. By considering the final percent of water retained after a standard centrifugation, as a function of the initial water content, a moisture retention capacity (MRC) can be determined. Statistical analysis of the results identified that it was possible to divide the nine samples of MCC into six sub-sets in terms of final moisture content retained. Those types which contained added polymer had by far the highest water level remaining. In terms of the MRC value, statistical analysis again sub-divided the celluloses into six sub-sets, although different from those for the final water content. Again those types containing added polymer gave much higher MRC values. As experiments were carried out with initial water/MCC ratios of different levels, statistical analysis of the influence of initial water content on the MRC values was undertaken with each type of MCC. The results showed a varying dependence on initial water level with the different types of cellulose. To provide a value of MRC which characterised the cellulose, the maximum value of MRC at the lowest initial water level was identified. Samples of MCC with low values of MRC have been shown previously to require less water for processing by extrusion/spheronization, while celluloses with a high MRC value appear better for the limitation of water movement during the process of extrusion/spheronization. The water retention must, however, also be considered in association with the rheological properties of the wet powder mass. Thus, while the Avicel RC591 had the highest MRC value, its rheological properties are so that the production of pellets with such a type can be less effective than with other types of MCC. PMID- 11113652 TI - Prediction of drug transport processes using simple parameters and PLS statistics. The use of ACD/logP and ACD/ChemSketch descriptors. AB - A method of modelling and predicting biopharmaceutical properties using simple theoretically computed molecular descriptors and multivariate statistics has been investigated for several data sets related to solubility, IAM chromatography, permeability across Caco-2 cell monolayers, human intestinal perfusion, brain blood partitioning, and P-glycoprotein ATPase activity. The molecular descriptors (e.g. molar refractivity, molar volume, index of refraction, surface tension and density) and logP were computed with ACD/ChemSketch and ACD/logP, respectively. Good statistical models were derived that permit simple computational prediction of biopharmaceutical properties. All final models derived had R(2) values ranging from 0.73 to 0.95 and Q(2) values ranging from 0.69 to 0.86. The RMSEP values for the external test sets ranged from 0.24 to 0.85 (log scale). PMID- 11113653 TI - New trends in antiretroviral therapy for HIV infection. AB - The treatment of HIV infection is a dynamic topic. More than 15 compounds, derived from three classes of antiretroviral drugs, are now available. A large number of clinical trials have determined the combinations that are optimal to use as first-line therapy; further investigations are needed to establish the value of simplified regimens (reduced number of doses and/or pills per day) in any attempt to increase the adherence of patients to therapy. In patients experiencing virological failure, assessment of adherence to treatment is helpful to determine the mechanisms of failure and to choose an alternative therapeutic option. The value of in vitro viral resistance evaluation by genotypic and/or phenotypic methods is currently being investigated. The value of therapeutic regimens available at the present time to treat HIV infection is hampered by their side effects. Metabolic toxicity has recently been identified as a major point of concern, and a better understanding of its mechanisms is urgently needed. Finally, in patients with long-term response to treatment, the value of passive immunotherapy to reduce the size of the viral reservoir and the effect of transient treatment interruption on specific immunological response have recently been investigated, but the first results are disappointing. PMID- 11113654 TI - Systematic, immediate in-hospital initiation of lipid-lowering drugs during acute coronary events improves lipid control. AB - Background: Patients who have had a coronary heart attack often go completely untreated for hypercholesterolemia. We investigated whether immediate initiation of lipid-lowering drugs during hospitalization for acute coronary events increases the proportion of correctly treated patients compared to referred treatment as recommended by current guidelines. Methods: This prospective, multicenter study randomized 57 hypercholesterolemic patients hospitalized for acute coronary events to immediate in-hospital initiation or to referred initiation of lipid-lowering drugs by primary care physicians 3 months after unsuccessful nutritional intervention. Results: After 6 months, 53 patients were available for follow-up. More patients in the immediate initiation group (26/30 patients, 87%) were treated with lipid-lowering drugs than in the referred initiation-group (13/23 patients, 57%, P=0.03). Twenty-seven patients (87%) in the immediate initiation group versus 17 patients (65%) in the referred initiation group had a 10% or greater decrease in total cholesterol or a 15% or greater decrease in LDL-cholesterol (P=0.18). Although statistically not significant, there was a trend to improved lipid values in the immediate initiation group compared to the referred initiation group (TC, -21.1 vs. -13.8% (P=0.08); LDL-C, -28.2 vs. -18.9% (P=0.13); HDL-C, +10.8 vs. +5% (P=0.44); TC/HDL C ratio, -24.7 vs. -15.1% (P=0.22)), and the LDL-C/HDL-C ratio was -34.1 vs. 19.1% (P=0.04, P=NS after Bonferroni correction). Conclusion: The immediate initiation of lipid-lowering drugs in hypercholesterolemic patients hospitalized for acute coronary events increases the rate of correctly treated patients and has the potential to improve lipid control. PMID- 11113655 TI - Biochemical expression of heterozygous hereditary hemochromatosis. AB - Background: Hereditary hemochromatosis (HH) is a common autosomal recessive disease caused by an iron overload. Two mutations (C282Y and H63D) on the responsible HFE gene have been described. HH heterozygotes may have a slight iron overload that does not cause clinical disease. Compound heterozygosity may be associated with higher iron stores than C282Y heterozygosity. We studied biochemical iron parameters in HH C282Y and compound heterozygotes without a clinically significant iron overload. Methods: Data on hemoglobin, hematocrit, mean corpuscular volume, serum ferritin, serum iron, transferrin, and transferrin saturation were obtained from 40 C282 wild type controls (irrespective of H63D genotype), 61 C282Y heterozygotes, and 18 compound (C282Y/H63D) heterozygotes without clinical iron overload disease. Results: Serum ferritin levels were significantly higher in female HH heterozygotes, particularly in compound heterozygotes, than in normal women. In male heterozygotes, no difference in serum ferritin was found. We found higher mean serum iron and transferrin saturation levels in male and female HH heterozygotes than in normal controls, the highest in the group of compound heterozygotes. Conclusions: Mean serum ferritin (only in women), serum iron, and transferrin saturation are highest in compound heterozygotes and lowest in controls. C282Y heterozygotes seem to be an intermediate group between compound heterozygotes and the normal population. PMID- 11113656 TI - Association of increased C-peptide serum levels and testosterone in type 2 diabetes. AB - Background: The purpose of our study was to investigate the association of C peptide and steroid hormones in males and females with type 2 diabetes compared to controls. Methods: In 562 subjects, matched for age and body mass index (BMI), 126 female type 2 diabetic patients (known diabetes duration: 7.8+/-0.8 years, HbA(1c): 7.6+/-0.14%), 126 healthy female subjects, 155 male type 2 diabetic patients (known diabetes duration: 6.4+/-0.6 years, HbA(1c): 7.7+/-0.11%), and 155 healthy male controls, C-peptide levels and serum levels of steroid hormones (progesterone, cortisol, DHEAS, estradiol, and testosterone) were measured by immunometric assays. Ratios of steroid hormones were calculated to investigate shifts in steroidogenesis. Results: In female patients, testosterone was significantly higher than in controls (1.7+/-0.1 vs. 1.4+/-0.2 pmol/l; P<0.05), something that was also demonstrated for the ratio of testosterone/estradiol (P<0.05). In male patients, lower levels of testosterone (11.8+/-0.5 vs. 14.3+/ 0.5 pmol/l; P<0.05) and higher cortisol levels (257.5+/-9.9 vs. 228.2+/-7.9 umol/l, P<0.01) were found than in controls. The progesterone/DHEAS ratio (P<0.05) and the progesterone/testosterone ratio (P<0.001) were significantly higher and DHEAS/cortisol significantly lower in type 2 diabetic males than in controls. In a multiple linear regression analysis that controlled for age, BMI, C-peptide, HDL-cholesterol, and HbA(1c), testosterone was significantly and positively correlated with C-peptide levels in female (P<0.05) but not in male type 2 diabetic patients. Conclusions: Testosterone levels were higher in female patients than in controls and correlated with C-peptide levels while testosterone levels were lower in male patients than in controls and showed no correlation with C-peptide. A higher ratio of testosterone/estradiol in type 2 diabetic females and of progesterone/DHEAS and progesterone/testosterone in type 2 diabetic males than in controls may indicate gender-dependent shifts in steroidogenesis. Whether the shifts in steroidogenesis contribute to insulin resistance in diabetic patients should be the subject of further studies. PMID- 11113657 TI - Palliative hormone therapy, low-dose chemotherapy, and bisphosphonate in breast cancer patients with bone marrow involvement and pancytopenia: report of a pilot experience. AB - Background: The choice between supportive care or specific anticancer treatment for poor performance status (PS) breast cancer patients with multimetastatic disease and pancytopenia due to bone marrow involvement (BMI) often remains a clinical dilemma. Patients and methods: Five consecutive patients with poor PS (WHO 2 or 3) and severe pancytopenia due to BMI received concomitant hormone therapy, weekly low-dose chemotherapy (anthracycline or anthracenedione), and either oral clodronate or intravenous pamidronate. Hormone therapy was adjusted to the patients' menopausal status and/or previous (adjuvant) therapy and consisted of either tamoxifen+/-LH-RH agonist or an aromatase inhibitor. Results: In the five treated patients, one treatment-unrelated death was observed in the early phase. Significant PS improvement, pain relief, and rapid normalization of the blood counts were observed in the remaining cases. No major toxic phenomenon was observed. No severe infection occurred, and red cell or platelet transfusions were not required after 1-4 months of treatment. Three patients showed objective tumor response. Overall survival ranged from 12 to 38 months. Conclusion: Due to its low aggressiveness and potentially high activity, this combined treatment should be preferred to supportive care alone. A significant survival gain can be obtained in patients who respond, even with initially poor PS. PMID- 11113658 TI - Differential diagnosis between community-acquired pneumonia and non-pneumonia diseases of the chest in the emergency ward. AB - Background: The differential diagnosis of community-acquired pneumonia and some non-pneumonia diseases involving the chest may sometimes be cumbersome. Adding some objective variables to the diagnostic strategy may be helpful.We evaluated the main objective variables that are usually available in the emergency ward and that may be valuable in this differential diagnosis. Methods: We recorded epidemiological, clinical, and analytical data, as well as that obtained from physical examination, from 284 consecutive patients diagnosed in the emergency ward as having community-acquired pneumonia. The diagnosis was reviewed by the investigators applying pre-set diagnostic criteria. Statistical analysis was then performed comparing data from patients with a definitive diagnosis of community acquired pneumonia with those with a final diagnosis of non-pneumonia disease excluding acute exacerbations of chronic bronchitis. Results: In the univariate analysis, C-reactive protein (difference of means 93 mg/l; 95% C.I. 47, 140), erythrocyte sedimentation rate (d.m. 19 mm/h; 95% C.I. 3, 35), leukocyte count (d.m. 3.5x10(9)/l; 95% C.I. 0.5, 6.4), and temperature (d.m. 0.5 degrees C; 95% C.I. 0.1, 0.9) discriminated between community-acquired pneumonia and non pneumonia diseases. In the multivariate analysis, only C-reactive protein remained in the equation. Conclusions: C-reactive protein, erythrocyte sedimentation rate, leukocyte count, and temperature were measurable variables that proved to be useful in the differential diagnosis between community-acquired pneumonia and non-pneumonia diseases. C-reactive protein appears to be the most suitable for this purpose. PMID- 11113659 TI - Tuberculous myocarditis presenting as long QT syndrome. AB - An increasing QT interval can precipitate life-threatening tachyarrhythmias such as ventricular fibrillation. Tuberculous myocarditis is a very unusual diagnosis commonly made at autopsy. Mycobacterium tuberculosis can invade the cardiac conduction system and produce potentially dangerous arrhythmias. This case presents an HIV-infected man with tuberculous infection and long QT syndrome. We comment on the pathology, clinical features and outcome of this rare form of tuberculous infection. PMID- 11113660 TI - A rare manifestation of Kingella kingae infection. AB - Kingella kingae is an aerobic gram-negative coccobacillus that has been associated predominantly with bone and joint infection but also with septicemia and endocarditis. Until now, only four cases of proven K. kingae meningitis have been reported. We describe a case of a K. kingae meningitis in a male adolescent who presented with a history of fever of unclear origin. PMID- 11113661 TI - A patient with essential thrombocytosis and multiple sclerosis. AB - We report on a patient with a 5-year history of essential thrombocytosis (ET) who developed multiple sclerosis (MS) during the last 5 months. The patient was treated for MS with interferon-beta (IFN-beta), which also had a beneficial effect on the ET. We describe the patient's history and the beneficial effect of IFN-beta administration in reducing the number of platelets. We also discuss the possible link between the pathogenesis of ET and MS. PMID- 11113662 TI - Hypercalcaemia of hypoadrenal crisis mistaken for hypercalcaemia of malignancy in a patient with known bone metastases: a case report. AB - Hypercalcaemia is a well-recognised feature of hypoadrenalism. The adrenal glands are often involved with metastatic disease subclinically although full hypoadrenal crisis is not uncommon. Ill patients with known malignancy should generate a high degree of clinical suspicion for the possibility of adrenal involvement. This case highlights the need to look beyond confirmed bone metastases as the cause of hypercalcaemia in patients with widespread carcinomatosis. PMID- 11113663 TI - Internal medicine in Poland. PMID- 11113664 TI - Cancer cachexia. AB - Cachexia is a common cause of morbidity and mortality in patients with advanced cancer. It is characterised by numerous metabolic abnormalities including inefficient substrate utilisation, alterations in the balance of energy intake and expenditure and the acute-phase protein response. These changes seem to be driven by pro-inflammatory cytokines, alterations of the neuro-endocrine axis and tumour-derived catabolic factors. This results in the loss of both fat and lean tissue. Trials of conventional nutritional supplements in patients with cancer cachexia have failed to show any benefit in terms of weight gain or quality of life and this may be because the ongoing metabolic abnormalities prevent the efficient use of additional calories supplied. A variety of pharmacological agents have been studied in an attempt to normalise these metabolic changes with only limited success. However, it is possible that the combination of an agent to normalise the metabolic milieu along with the provision of additional nutritional support may have the potential to reverse cachexia in advanced cancer. PMID- 11113665 TI - The role of NF-kappaB/IkappaB proteins in cancer: implications for novel treatment strategies. AB - The nuclear factor-kappaB (NF-kappaB) family of transcription factors are involved in multiple cellular processes, including cytokine gene expression, cellular adhesion, cell cycle activation, apoptosis and oncogenesis. Constitutive activation of NF-kappaB has been described in a number of solid tumors and this activation appears to affect cancer cell survival. Inhibition of NF-kappaB has been shown to enhance the sensitivity of some cancer cell lines to antineoplastic or radiation-induced apoptosis. Furthermore, suppression of NF-kappaB results in attenuation of cancer cachexia in a mouse tumor model. Studies are underway to further delineate the role of NF-kappaB in cancer cell survival, growth and resistance to standard chemotherapy and radiation regimens. Moreover, the effects of novel therapeutic agents which specifically target NF-kappaB proteins are currently being assessed in experimental models of cancer cell growth both in vitro and in vivo. In this review, we discuss the possible involvement of NF kappaB in the growth of various solid tumors and potential future treatment strategies based on NF-kappaB inhibition. PMID- 11113667 TI - Concluding remarks by the retiring Editor. PMID- 11113666 TI - Parathyroid cancer: biology and management. AB - A review of all reports in the literature of parathyroid carcinoma (PTC) was undertaken to define an optimal management strategy for this rare condition. PTC is uncommon and its etiology of PTC is largely unknown although patients with familial hyperparathyroidism, multiple endocrine neoplasia type 1 and irradiation to the head and neck are at increased risk for developing the disease. PTC occurs with equal frequency in both sexes and is usually diagnosed in the fifth decade. En bloc resection of the carcinoma and the adjacent structures in the neck is the surgical treatment and is associated with an 8% local recurrence rate and a long term overall survival rate of 89% (mean follow up 69 months). In contrast simple parathyroidectomy results in a 51% local recurrence rate and 53% long-term survival rate (mean follow up 62 months). Adverse prognostic factors for survival were initial management with simple parathyroidectomy alone, the presence of nodal or distant metastatic disease at presentation and non-functioning PTC. PMID- 11113668 TI - Perfection and compassion--essentials in cardio-thoracic surgery. PMID- 11113669 TI - The language of science. PMID- 11113670 TI - Combined photodynamic therapy and hyperbaric oxygenation in carcinoma of the esophagus and the esophago-gastric junction. AB - OBJECTIVES: The photochemical reaction of photodynamic therapy (PDT) depends on the presence of molecular oxygen. Due to anoxic regions in tumor tissue and vascular shutdown during PDT the efficiency is limited. Therefore, the use of hyperbaric oxygen which increases the oxygen in tumor tissue, as well as the amount of singlet oxygen, may enhance the efficiency of PDT. PATIENTS AND METHODS: After diagnostic work-up, photosensitization was carried out with a hematoporphyrin-derivate 2 mg/kg BW 48 h prior to PDT. The light dose was calculated as 300 J/cm fiber tip. Thirty-one patients were treated by PDT alone and 44 patients received PDT under hyperbaric oxygen at a level of two absolute atmospheric pressure. RESULTS: Improvement regarding stenosis-diameter could be obtained in both treatment arms with no significant difference (P=0.82). The dysphagia-score and tumor-length also decreased in both groups and showed a significant difference in favour of the PDT/HBO-group (P=0. 0064 and P=0.0002, respectively). The median overall survival for the PDT-group was 7 months and for the PDT/HBO-group 12 months (P=0. 0098). CONCLUSION: According to this prospective non-randomized study, combined PDT/HBO represents a new approach in the treatment of esophageal and cardia cancer which appears to have enhanced the efficiency of PDT. PMID- 11113671 TI - Muscle sparing thoracotomy: a biomechanical analysis confirms preservation of muscle strength but no improvement in wound discomfort. AB - OBJECTIVES: This study compares the posterior auscultatory triangle thoracotomy incision (muscle sparing) with full posterolateral thoracotomy (where latissimus dorsi muscle is always cut across its full width), with particular attention to the difference between latissimus dorsi muscle strength, post operative pain and chronic wound related symptoms. METHODS: Ten patients who had undergone auscultatory triangle thoracotomy (ATT) at least 1 year previously were matched with ten patients who had undergone posterolateral thoracotomy (PLT). Each pair was matched for age, sex, dominant hand, side of the operation, time since operation and presence or absence of history of previous muscle training. Latissimus dorsi muscle strength was assessed by testing the shoulder adduction strength through an arc of 90-0 degrees using isokinetic technique. Early post operative pain was assessed indirectly by calculating the analgesic requirement in the first 5 post-operative days. A subjective assessment of chronic post thoracotomy pain was made using a questionnaire presented to the patients at the time of muscle testing. Variability of the torque curves, recorded as coefficient of variance at the time of muscle strength testing, provided objective measurements of chronic pain. Data were analysed using two sample t-tests. RESULTS: All patients reported at least one chronic post-thoracotomy symptom. There was no significant difference between the two groups in terms of acute or chronic wound pain and other long term wound related symptoms. Shoulder adduction strength was 24% greater in ATT than PLT (95% confidence limits=1-43%, P=0.04). CONCLUSIONS: All thoracotomy patients have long term wound related symptoms. This situation is not improved by performing a muscle sparing incision. However thoracotomy through the triangle of auscultation can preserve latissimus dorsi strength which is compromised in a posterolateral thoracotomy incision. We therefore recommend that a muscle sparing thoracotomy be considered for patients where preservation of muscle strength is deemed important, providing the operation is not compromised due to inadequate access. PMID- 11113672 TI - Pulmonary complications after surgical treatment of lung cancer in octogenarians. AB - OBJECTIVE: The purpose of this study was to analyze the risks associated with pulmonary resection for primary non-small cell lung cancer in octogenarians to help better management in these patients. METHODS: We reviewed the outcome in our 35 patients aged 80 years and older who underwent pulmonary resection between 1981 and 1998. RESULTS: The 5-year survival rate was 39.8%. The operative mortality rate was 0% and the morbidity 60%. There were ten major pulmonary complications, including respiratory insufficiency following bacterial pneumonia and sputum retention. Preoperative arterial pO(2) was significantly lower, A aDO(2) was significantly higher, and operation time were significantly longer in patients with pulmonary complications after surgical treatment than in patients without complications (P<0.05). CONCLUSIONS: Surgical treatment was not contraindicated for octogenarians with lung cancer. However, a relatively preoperative low arterial pO(2), high A-aDO(2), and long operation time may be risk factors for postoperative pulmonary complications in such patients. Surgeons must assess the preoperative data prudently to determine appropriate surgical strategy. PMID- 11113673 TI - Intrathoracic muscle flap transposition in the treatment of fibrocavernous tuberculosis. AB - BACKGROUND AND OBJECTIVE: Conventionally, pulmonary resection with thoracoplasty is used to treat fibrocavernous complication of pulmonary tuberculosis. This operation is usually bloody, time-consuming with complicated postoperative course. To prevent massive blood loss and preserved pulmonary function, a more simplified operative procedure, cavernostomy combined intrathoracic muscle flap transposition was used and the outcome was evaluated in this study. DESIGN: Retrospective review. METHODOLOGY: Between December 1989 and June 1996, a total of ten patients with fibrocavernous pulmonary tuberculosis were managed using cavernostomy combined with intrathoracic muscle flap transposition. Five of them had concomitant aspergilloma within the cavity while three had multiple drug resistant pulmonary tuberculosis. The muscle flap was used to plombage the cavity and reinforce the closure of bronchopleural fistula after cavernostomy. RESULTS: Six postoperative complications occurred in five patients, including reformation of cavity (2), bronchopleurocutaneous fistulae (3), and postoperative bleeding (1). The success or failure of intrathoracic muscle flap transposition on patients with fibrocavernous tuberculosis was significantly correlated with the size of the cavity (194.0+/-11.2 vs. 283.0+/-44.6 cm(3), P=0.016) and the number of bronchopleural fistulae (1.6+/-0.4 vs. 4.0+/-0.4, P=0.008). There was no operative death and in long term follow-up, there was no recurrence of hemoptysis or deterioration of pulmonary function in the successful group of patients. CONCLUSIONS: Cavernostomy combined with intrathoracic muscle flap transposition can be used to treat well-selected fibrocavernous pulmonary tuberculosis patients, except on patients with large size cavity, multiple bronchopleural fistulae or multiple drug resistance tuberculosis. PMID- 11113674 TI - Therapeutic video-assisted thoracoscopic surgical resection of colorectal pulmonary metastases. AB - OBJECTIVE: Careful patient selection is vital when video-assisted thoracoscopic surgical (VATS) therapeutic pulmonary metastasectomy of colorectal carcinoma is considered. Complete resection of all metastatic disease remains a vital concept. We reviewed our VATS experience for therapeutic metastasectomy of peripheral colorectal pulmonary metastases. METHODS: Over 90 months, therapeutic VATS metastasectomy was accomplished upon 80 patients with colorectal metastases. Thin cut computed tomography (CT) was central in identifying lesions. The mean interval from primary carcinoma to VATS resection was 41 months (1-156 months; median, 33). A solitary lesion was resected in 60 patients and multiple (2-7) lesions resected in 20 patients. Statistics were obtained using the Student's t test. RESULTS: No operative mortality or major postoperative complications occurred. The hospital stay was 4.5+/-2. 2 days (range, 1-13). All lesions were resected by VATS, with four conversions to thoracotomy to improve the margins. The mean survival of patients with one lesion was 34.8 months compared with 26.5 months for patients with multiple lesions (P=0.37). The mean survival was 20.5 months when metastases occurred <3 years vs. 28.1 months for >3 years from primary carcinoma resection (P=0.20). Twenty-five (31%) patients are disease free; with a mean interval of 38.7 (3-84; median, 35) months. Sixty-nine percent (55/80) of patients developed a recurrence: 6/80 (8%) local; 19/80 (24%) regional (same hemithorax away from resection); and 30/80 (38%) distant. The overall survival at 1 year was 81.2%, 48.4% at 3 years and 30.8% at 5 years. CONCLUSIONS: Therapeutic VATS resection of colorectal metastases appears efficacious. Preoperative CT can identify peripheral colorectal metastases amenable to VATS. Conversion to thoracotomy is indicated when none of the lesions identified by CT are found or when clear surgical margins are jeopardized. PMID- 11113675 TI - Right submammary minithoracotomy for repair of congenital heart defects. AB - OBJECTIVE: The initial experience with the right submammary minithoracotomy incision for correction of intracardiac congenital defects is reported. METHODS: Between March 1997 and March 1999, 100 children underwent repair of congenital heart disease through this approach. Their mean age and weight were 4.6 years and 20 kg, respectively. Diagnosis included: atrial septal defect (78), ventricular septal defect (7), tetralogy of Fallot (6), partial atrioventricular canal (5), double-chambered right ventricle (3) and single ventricle with dextrocardia (1). The standard technique entailed a 5 to 6 cm right submammary incision, entering the chest through the third or fourth intercostal space (depending on the body weight), direct aortic and bicaval cannulation and aortic cross-clamping with cardioplegic protection. RESULTS: There were no hospital deaths. Postoperative morbidity included bleeding in two cases, recurrent atrial septal defect in one, spleen injury in one. The average hospital stay was 3.5 days. All patient are currently free of symptoms and medications. CONCLUSIONS: (1) This approach for repair of selected congenital cardiac malformations is technically feasible, safe and effective; (2) younger age is a facilitating factor; (3) hospital stays are effectively reduced. PMID- 11113676 TI - The short- and mid-term results of bidirectional cavopulmonary shunt with additional source of pulmonary blood flow as definitive palliation for the functional single ventricular heart. AB - OBJECTIVE: The purpose of this study was to demonstrate the early and late outcomes of bidirectional cavopulmonary shunt (BCPS) as a definitive procedure for the functional single ventricular heart. METHOD: From September 1991 to December 1997, 34 patients underwent a BCPS procedure without a routine conversion to Fontan circulation. The additional source of pulmonary blood flow was left in all patients. Conversion was performed only when it was required for excessive cyanosis. RESULTS: The hospital mortality rate was 8.8% (3/34, 95% confidence limit; 1.9-23%) and the 5-year survival rate was 75% for a mean follow up period of 33+/-22 months. Seven patients underwent a conversion procedure for remnant or recurrent cyanosis and deterioration of exercise tolerance. Four of these patients died after conversion to Fontan circulation. Twenty-five long-term survivors with BCPSs maintained an arterial oxygen saturation of 84+/-6.1%, and 52% of them had a normal exercise tolerance or mild limitation. No patients developed severe late complications other than recurrent cyanosis. CONCLUSION: Due to the high mortality after conversion to Fontan circulation in patients whose conditions had deteriorated, we could not demonstrate the clear superiority of long-term BCPS over the construction of Fontan circulation for management of the functional single ventricular heart. If deteriorated conditions were successfully managed in the late period, the outcome of long-term BCPS would have been better. PMID- 11113677 TI - Optimal conduit size for extracardiac Fontan operation. AB - BACKGROUND: Lack of conduit growth potential and thrombogenicity are the main drawbacks of the extracardiac Fontan operation (ECFO). Optimal size of the conduit according to the patients age and inferior vena cava diameter has not been established. OBJECTIVES: We set out to ascertain whether the optimal dimensions of the conduit could be determined before an ECFO. METHODS: Actual and expected age-related inferior vena cava diameters were compared with the extracardiac conduit diameter in 20 patients after ECFO. In 50 other pediatric and adult patients, the distance between intrapericardial part of the inferior vena cava and the undersurface of the right pulmonary artery (IVC-RPA) was measured. Cases of conduit thrombosis were analyzed. RESULTS: The actual diameter of the inferior vena cava was variable and has a weak correlation with anthropometric data and expected diameter (R=0.07-0.23, P=0.32-0.76). The IVC-RPA distance correlated with height (R=0.87, P=0.0001), but was also variable. At the age of 2-4 years and body weight 12-15 kg IVC diameter and IVC-RPA distance are equal to 60-80% of adult values. Conduit thrombosis developed in two patients with unfavorable Fontan hemodynamics and oversized conduits. CONCLUSIONS: Considering the inferior vena cava size, ECFO may be performed at the age of 2-3 years and at a body weight 12-15 kg, when a hemodynamically optimal almost adult sized conduit can be implanted. Optimization of the conduit is necessary on the basis of the actual inferior vena cava diameter and IVC-RPA distance. Anticoagulation postoperatively should be considered to prevent conduit thrombosis in patients with suboptimal Fontan circulation PMID- 11113678 TI - Pulmonary thromboendarterectomy in patients with chronic thromboembolic pulmonary hypertension: hemodynamic characteristics and changes. AB - OBJECTIVE: To see whether degree of pulmonary hypertension or severity of cardiac failure affect the success of pulmonary thromboendarterectomy (PTE) in chronic thromboembolic pulmonary hypertension. METHODS: From May 1996 to June 1999, 33 patients, all in New York Heart Association (NYHA) class 3 or 4 were treated with PTE. Preoperative hemodynamic values were: central venous pressure (CVP) 8+/-6 (1 23), mean pulmonary artery pressure (mPAP) 50+/-10 (30-69), cardiac output (CO) 3.3+/-0.9 (1.8-5.2), pulmonary vascular resistance (PVR) 1056+/-344 (523-1659), and right ventricle ejection fraction (RVEF) 12+/-5 (5-21). To establish whether some hemodynamic or cardiac variables correlate with surgical failure (early death or functional non-success), these patients were divided into a low risk or a high risk group for each variable: CVP (<9 or > or =9), mPAP (<50 or > or =50), CO (> or =3.5 or <3.5), PVR (> or =1100 or <1100), and RVEF (> or = 10 or <10). The duration of 3-4 NYHA class period (<24 or > or = 24 months) was also included in the study. RESULTS: Three patients (9. 1%) died in hospital, one (3.0%) underwent lung transplant shortly after PTE, and in five cases (15.2%) mPAP and PVR at the 3-month follow-up examination corresponded with our definition of functional nonsuccess (mPAP and PVR decreased by less than 40% of preoperative values). One of the five functional nonsuccess patients underwent lung transplant 3 months after the operation and another died 17 months after the operation from a non-related cause. Thus PTE was successful in 24 patients and unsuccessful in nine. None of the hemodynamic variables considered was found to be associated with the disparate outcomes. At the 3-month examination, all surviving patients were in NYHA class 1 or 2 except for three in NYHA class 3. At 2 years, hemodynamic values were: CVP 2+/-2 (0-4), mPAP 16+/-3 (12-21), CO 5.0+/-1.0 (3.4 6.5), PVR 182+/-51 (112-282), and RVEF 35+/-5 (26-40). All differences were significant with respect to baseline values (P<0.001). Preoperative mPAP and RVEF values had a strict linear correlation (R=0.45; P=0.014). CONCLUSIONS: None of the variables considered was correlated with early death or functional nonsuccess. Neither preoperative severity of pulmonary hypertension nor degree of cardiac failure influenced the outcome of the operation. PTE leads to hemodynamic recovery even in very compromised patients. PMID- 11113680 TI - Surgical aspects of chronic post-thoracotomy pain. AB - Chronic post-thoracotomy pain is a continuous dysaesthetic burning and aching in the general area of the incision that persists at least 2 months after thoracotomy. It occurs in approximately 50% of patients after thoracotomy and is usually mild or moderate. However, in 5% the pain is severe and disabling. No one technique of thoracotomy has been shown to reduce the incidence of chronic postthoracotomy pain. The most likely cause is intercostal nerve damage, although the precise mechanism for this is not known. Future work needs to examine surgical technique in detail. Until then, patients need to be adequately warned of this sequela of thoracotomy. PMID- 11113679 TI - Intra-institutional prediction of outcome after cardiac surgery: comparison between a locally derived model and the EuroSCORE. AB - OBJECTIVE: To construct models for predicting mortality, morbidity and length of intensive care unit (ICU) stay after cardiac surgery and to compare the performance of these models with that of the EuroSCORE in two independent validation databases. METHODS: Clinical data on 4592 cardiac surgery patients operated between 1992 and 1996 were retrospectively collected. In order to derive predictive models and to validate them, the patient population was randomly divided into a derivation database (n=3061) and a validation database (n=1531). Variables that were significant in univariate analyses were entered into a backward stepwise logistic regression model. The outcome was defined as mortality within 30 days after surgery, predefined morbidity, and the length of ICU stay lasting >2 days. In addition to the retrospective database, the models were validated also in a prospectively collected database of cardiac surgical patients operated in 1998-1999 (n=821). The EuroSCORE was tested in two validation databases, i.e. the retrospective and prospective one. Hosmer-Lemeshow goodness of-fit was used to study the calibration of the predictive models. Area under the receiver operating characteristic (ROC) curve was used to study the discrimination ability of the models. RESULTS: The overall mortality in the retrospective and the prospective data was 2 and 1%, and morbidity 22 and 18%, respectively. The created predictive models fitted well in the validation databases. Our models and the EuroSCORE were equally good in discriminating patients. Thus, in the prospective validation database, the mean areas under the ROC curve for our models and for the EuroSCORE were similar, i.e. 0.84 and 0.77 for mortality, 0.74 and 0.74 for morbidity, and 0.81 and 0.79 for the length of intensive care unit stay lasting for 2 days or more, respectively. CONCLUSIONS: Our models and the EuroSCORE were equally good in discriminating the patients in respect to outcome. However, our model provided also well calibrated estimation of the probability of prolonged ICU stay for each patient. As was originally suggested, the EuroSCORE may be an appropriate tool in categorizing cardiac surgical patients into various subgroups in interinstitutional comparisons. Our models may have additive value especially in resource allocation and quality assurance purposes for local use. PMID- 11113681 TI - Radial artery graft inflow from the undetached, unharvested RIMA: a method to avoid proximal anastomosis to the aorta in CABG surgery. AB - Performing the proximal anastomosis of a free arterial graft to the ascending aorta is problematic, especially if the wall of the aorta is calcified or thickened. We describe a method, which makes it possible to avoid this procedure. PMID- 11113682 TI - Congenital cystic adenomatoid malformation in an adult presenting as lung abscess. AB - The case of a 21-year-old male with congenital cystic adenomatoid malformation is presented. His medical history started after his birth with recurrent pulmonary infections during his infancy. Lung abscess of the right lower lobe was suspected and right lower lobectomy was performed to remove a sizeable mass infiltrating the largest part of the lobe. The clinical features, diagnostic procedures, differential diagnosis, pathologic characteristics, therapeutic assessment, etiopathology and prognosis of the tumor are discussed. PMID- 11113683 TI - Correction of cor triatriatum sinistrum in a Jehovah's Witness infant. AB - A cor triatriatum sinistrum was successfully treated by operation in a 14-week old infant of a Jehovah's Witness family. The child was pretreated with erythropoietin until a hemoglobin level of 14 g/dl was obtained. There was no cardiac catheterization before the operation. The operation was performed with cardiopulmonary bypass. No blood products were transfused and the hemoglobin level after performing modified ultrafiltration was 11.5 g/dl. The infant was extubated on the same day and discharged from our institution on the eighth day after surgery. Two years after surgery the child is in sinus rhythm and is developing well. PMID- 11113684 TI - Health-related quality of life in patients with diabetes mellitus and foot ulcers. AB - To investigate health-related quality of life (HRQL) in diabetes patients separately for those with current foot ulcers, those with primary healed ulcers, and those who have undergone minor or major amputations. The EuroQol quality of life (QoL) questionnaire including a visual analogue scale (VAS) was sent to 457 patients treated for foot ulcers by a multidisciplinary team between 1995 and 1998. Patients who never had undergone any lower extremity amputation (LEA) were classified according to whether an ulcer was present at time of the survey or if they had healed primarily. Patients who had undergone any amputation were classified into a minor or a major amputation group according to their maximal amputation status. Patient characteristics and ulcer status at time of the survey were collected using patient records and pre-set forms used to follow-up of foot ulcer patients specifically. A response rate of 70% was obtained. Completion rates on single items were high. There were no differences in patient characteristics between respondents and non-respondents. Patients with current foot ulcers rated their HRQL significantly lower than patients who had healed primarily without amputation. Major amputation reduced the EuroQol index value, while the VAS value was reduced by other diabetic complications and increased by living with a healthy partner. Both values were reduced by a current foot ulcer. EuroQol can be used to investigate HRQL in diabetic patients with foot complications. Patients with current foot ulcers value their QoL lower than primary healed patients. QoL is reduced after major amputations. PMID- 11113685 TI - Epidemiological analysis of patients with Type 2 diabetes in France. AB - This paper presents the baseline epidemiological data from 5548 patients with type 2 diabetes enrolled in a French observational study that aims to examine the safety, tolerability and use of acarbose as prescribed by general practitioners (GPs). Patients were recruited and monitored by a representative sample of GPs. Recruitment did not depend on a patient's suitability for acarbose treatment. The data revealed that the mean age of the patient population was 63 years, and that more than 50% of patients were over 65 years old. The population was markedly overweight [mean body mass index(BMI): males, 28.4 kg/m(2); females, 29.1 kg/m(2)] and the mean duration of diabetes was 10 (+/-7.3) years. Over 37% of patients had at least one diabetic complication, and the frequency of complications increased with both age and the duration of diabetes. The most frequently reported complications were cardiac (17.8%), vascular (14.5%) and ocular (12%). At recruitment, almost 90% of patients were being treated with oral antidiabetic agents (OADs). Sulphonylureas (74%) and biguanides (50%) were the most commonly prescribed agents. Acarbose was used to treat 17% of patients and 1% were receiving insulin. GPs set glycaemic treatment goals for 44% of patients in the study. Fasting glycaemia was the primary goal for 37% of the total study population, and HbA(1c) levels for 21% of patients. Postprandial glycaemia was generally given as a secondary or tertiary goal. In conclusion, this study provides the most up-to-date epidemiological data for patients with type 2 diabetes in France. PMID- 11113686 TI - Comparative effects of pioglitazone, glibenclamide, and voglibose on urinary endothelin-1 and albumin excretion in diabetes patients. AB - Urinary endothelin (ET)-1 excretion is present in non-insulin dependent diabetes (NIDDM) patients with microalbuminuria, and an increase in circulating ET-1 precedes the microalbuminuric phase of renal injury related to diabetes. The aim of the present study was to determine whether various drugs alter urinary ET-1 levels and urinary albumin excretion (UAE) in NIDDM patients with microalbuminuria. Forty-five NIDDM patients with microalbuminuria were randomly assigned to three groups: those treated with pioglitazone at 30 mg/day (n=15), those treated with glibenclamide at 5 mg/day (n=15), and those treated with voglibose at 0.6 mg/day (n=15). Patients received these drugs for 3 months. UAE, urinary ET-1, and plasma ET-1 levels were measured in these patients before and after treatment. Before treatment, UAE, urinary ET-1, and plasma ET-1 levels differed little among the three groups. UAE in the 45 NIDDM patients (156.2+/ 42.8 microg/min) was greater than that in 30 healthy controls (8.2+/-2.6 microg/min) (P<.001). Urinary ET-1 levels in the NIDDM patients (8.7+/-1.3 ng/g urinary creatinine (UC)) were significantly higher than that in the controls (2.4+/-0.2 ng/g UC) (P<.01). Plasma ET-1 levels, however, in the NIDDM patients (1.3+/-0.4 pg/ml) did not differ significantly from the levels in healthy controls (1.0+/-0.6 pg/ml). Pioglitazone but no glibenclamide or voglibose reduced UAE from 142.8+/-42.2 to 48. 4+/-18.2 microg/min (P<.01) and urinary ET-1 levels from 8.6+/-1.3 to 3.4+/-0.5 ng/g UC (P<.01). These data suggest pioglitazone to be effective in reducing UAE and urinary ET-1 concentrations in NIDDM patients with microalbuminuria. PMID- 11113687 TI - Efficacy of infrainguinal bypass for limb salvage in young diabetic patients. AB - The efficacy of infrainguinal bypass for limb salvage in young diabetic patients has not been well established. The purpose of this study is to determine the intermediate-term results (patency and limb salvage) of infrainguinal revascularization carried out for limb salvage (rest pain or ulceration) in young (<50 years old) diabetic atherosclerotic patients. Thirty-nine bypasses in 31 patients with a mean age of 44 years were retrospectively reviewed. There were no perioperative deaths. Minor or major complications occurred in 23% of cases. By life table analysis, the 18-month primary patency rate was 60+/-11%, assisted primary patency rate was 78+/-9%, and limb salvage rate was 71+/-9%. Most major amputations (five of nine) were required in patients with functional bypasses, either because of persistent infection or failure of wound healing. The presence of severe stenoses (>70%) in all three major named foot vessels (dorsalis pedis, medial and lateral plantar arteries) was associated with a high likelihood of limb loss despite a patent bypass (p<0.05). We could not identify any other factors statistically predictive of thrombosis, amputation, or the need for graft revision. Infrainguinal revascularization in this patient population can be carried out with acceptable limb salvage rates. However, patients should be made aware of the high incidence of amputation regardless of the success of the revascularization procedure, particularly in the presence of severe occlusive disease within the foot. PMID- 11113688 TI - Urinary protein excretion in Type 2 diabetes with complications. AB - This study examined the association between urinary markers of early diabetic nephropathy and non-renal diabetic complications in 946 patients with type 2 diabetes mellitus. The association with hypertension was also studied. Data on macrovascular complications (ischaemic heart disease, stroke, peripheral vascular disease) and microvascular complications (retinopathy, peripheral neuropathy) were obtained from case records and clinical examination. Urine samples collected were analysed for albumin, beta(2)-microglobulin, retinol-binding protein (RBP), and N-acetyl-beta-D-glucosaminidase (NAG). Results showed that urinary albumin, RBP and beta(2)-microglobulin levels were higher in patients with macro- and/or microvascular complications, compared to those without. NAG levels were higher only in patients with both types of complications. A higher proportion of patients with complications had abnormally raised urinary protein and enzyme levels, compared to those without. Patients with associated hypertension had higher urinary levels of albumin and beta(2)-microglobulin, regardless of whether complications were present or not. RBP excretion was, however, markedly higher only in patients with microvascular complications, whereas hypertension did not influence NAG excretion. Urine albumin and RBP excretion were predictive of microvascular, as well as both macrovascular and microvascular complications, whereas NAG excretion was predictive of macro- and microvascular complications. These findings could mean that increased urinary protein and enzyme excretion were associated with more severe disease in these patients. PMID- 11113689 TI - Prognostic factors in Brazilian diabetic patients starting dialysis: a 3.6-year follow-up study. AB - The objective of this study was to analyze the prognostic factors of a cohort of diabetic patients starting dialysis. This prospective, 3. 6-year population-based cohort study included 111 diabetic patients starting dialysis in all 18 dialysis centers of the metropolitan area of Porto Alegre, Brazil, between July 1995 and October 1996. The survival rate was analyzed by Kaplan-Meier curves and prognostic factors for death by Cox's proportional-hazards model. During the study period, 685 patients started dialysis; 182 (26.5%) had diabetes and 111 patients were included. Eighty-four percent of the 111 patients were classified as type 2 diabetes (random C-peptide>0. 6 ng/ml), and these patients presented more coronary artery disease (60% vs. 29%; P<0.02) than type 1 patients. In type 2 patients, later diagnosis of diabetes was associated with a shorter interval until beginning of dialysis (r=0.67; P=0.001). Diabetic nephropathy was the primary renal disease in 61% of all patients. Overall median survival (26 months) was similar for types 1 and 2 diabetic patients. Survival in the first, second, and third year was 69%, 51%, and 28%, respectively. Cardiovascular disease was the most common cause (63%) of death. According to Cox's proportional-hazards model, history of stroke (HR: 4.53, CI: 2.09-9.86, P<0.0001), amputations (HR: 3.2, CI: 1.61-6.35, P<0.0009), and coronary artery disease (HR: 1.67, CI: 0.95 2.96, P<0.076) at baseline were significantly associated with mortality. In conclusion, macrovascular complications were the main predictors of mortality in this cohort of diabetic patients starting dialysis. Intensive treatment of cardiovascular risk factors during dialysis might reduce the mortality rate of diabetic patients. PMID- 11113690 TI - Hypomagnesemia is linked to low serum HDL-cholesterol irrespective of serum glucose values. AB - Hypomagnesemia is common in diabetic subjects, and is especially common in poorly controlled diabetes, suggesting that diabetes low serum magnesium status is osmotic diuresis-dependent. To assess the relationship between serum magnesium and HDL-cholesterol concentration adjusted by serum glucose values. We assessed the serum magnesium levels of 50 controlled (HbA(1c)7.5% and FPG>/=126 mg/dl) type II diabetic patients, 40 subjects with impaired fasting glucose (IFG) (FPG>/=110 mg/dl and <126 mg/dl) and 190 healthy volunteers (FPG<110 mg/dl). Healthy volunteers were required to have normal blood pressure and normal laboratory tests. Subjects in the groups included were matched by age and body mass index (BMI). The average of diabetes duration was of 11.4+/-6.6, and 10.9+/-6.2 years, P=NS, for the controlled and non-controlled diabetic patients, respectively. Thirty (60.0%) controlled diabetic subjects, 58 (52. 7%) non-controlled diabetic patients, 21 (52.5%) subjects with IFG, and 39 (20.5%) healthy volunteers had serum magnesium levels 90% of dibenz[a,h]anthracene (4.18mg/kg soil), benzo[a]pyrene (1.96mg), benzofluoranthenes (0.14mg), and benz[a]anthracene (0. 18mg) were present in the acetone subfraction of cyclohexane solubles. Concentrations and mutagenic activities, however, did not correlate. Additional preparative and analytical HPLC of the solvent fractions of polar neutrals and polar aromatics, resulted in the tentative identification of 2 nitrofluorene. Analysis of the vertical profile of soil revealed an increase of mutagenicity per gram from the surface to a maximum at 5-15cm depth and a subsequent decrease with very little activity remaining deeper than 35cm. In human lymphocyte cultures, the fraction of polar aromatics, 0.01-0. 3microg/ml, induced 11.27+/-4.76-20.70+/-6.19 sister-chromatid exchanges (SCE) per cell in the absence of S9 (solvent control: 10. 16+/-4.83 SCE per cell) and 12.77+/-6.53 17.87+/-4.93 SCE per cell in the presence of S9 (solvent control: 8.37+/-3.92 SCE per cell). However, no activities could be detected in the fractions of polar neutrals and non-polar neutrals. Again, negative results were obtained in the in vivo mouse bone marrow micronucleus assay at 2000mg/kg p.o. with all fractions. PMID- 11113693 TI - Using semi-automated oscillometric blood pressure measurement in diabetic patients and their offspring. AB - To determine whether a semi-automatic oscillometric blood pressure (BP) monitor Dinamap 1846SX (DIN) can replace the standard mercury sphygmomanometer (SMS) for BP measurements in diabetic patients and their offspring, we compared SMS with DIN in 105 diabetic patients and their families. Their mean age was 50.6 (range 24-86) years, of whom 41 had diabetes mellitus (DM), 32 impaired glucose tolerance and 32 non-DM. After resting quietly for 10 min, their right arm BP were measured twice with each device at random and with 1-min intervals between each measurement. Agreement between measurements was tested by plotting the differences between the methods against means and by intraclass correlation coefficient (r(I)). The DIN was also evaluated by the criteria of American Association for the Advancement of Medical Instrumentation (AAMI), the British Hypertension Society (BHS) criteria and clinical criteria of O'Brien. All measurements by DIN [first readings or averaged readings of duplicate measurements of systolic BP (SBP) or diastolic BP (DBP)] satisfied the AAMI criteria and had good agreement with SMS (r(I)=. 951 for SBP and r(I)=.905 for DBP). The first readings of systolic BP measured by DIN vs. SMS failed to satisfy the criteria by O'Brien and reached BHS grade C level. Other measurements passed the limits of O'Brien and reached BHS grade A or B. In conclusion, averaged readings of duplicate BP measurements by DIN are interchangeable with that by SMS in Chinese diabetic patients and their offspring. Only one single DIN measurement is not acceptable for clinical application. PMID- 11113695 TI - Soil mutagens are airborne mutagens: variation of mutagenic activities induced in Salmonella typhimurium TA 98 and TA 100 by organic extracts of agricultural and forest soils in dependence on location and season. AB - As our hypothesis was that soil mutagens are airborne mutagens, possibly modified by soil microorganisms, we checked solvent extracts from agricultural and forest soils collected during late summer in the environment of Mainz, a region highly charged by anthropogenic air pollution, or near Bayreuth, a rural low charged region of Germany, or in a remote region of western Corsica without anthropogenic air pollution for the presence of mutagenicity in Salmonella typhimurium. Levels of mutagenic activities were quantified by calculation of revertants/g from the initial slope of dose-response curves applying tester strains S. typhimurium TA 98 and TA 100 in the absence and presence of an activation system from rat liver (S9). Three soils from Corsica did not induce mutagenicity under any test condition. However, most soils from Germany exhibited mutagenic activities, though preferentially in strain TA 98, but no statistically significant differences could be detected between 27 soils from the Mainz and nine soils from the Bayreuth regions. On the other hand, no correlation could be detected between the levels of mutagenic activities at any test condition and agricultural practice - rye growing, viniculture, fruit growing, meadow, and fallow - texture of soils - % composition of clay, slit, and sand - or the contents of organic matter. The only significant difference of mutagenicity was, however, found with S. typhimurium TA 98-S9 between forest soils of pH approximately 4.0 as compared with agricultural soils of pH approximately 7.0. The presence of antimutagens in soil as demonstrated by the course of dose-response curves of the three soils from Corsica may be another possible confounder. Calculation of mean values of mutagenic activities for all soils from Germany gave the following results: S. typhimurium TA 98: 69.7+/-153.2 (-S9); 63.0+/-176.3 (+S9); S. typhimurium TA 100: 144.7+/-399.4 (-S9); 43.3+/-172.0 (+S9) revertants/g of dry soil. In another series of experiments, soil mutagenicity in 10 rye fields near Mainz was monitored for 1 year. It became evident that low levels of mutagenic activities in late summer increased during autumn, reached a peak in late winter, and subsequently, decreased during spring and summer. These results agree with the hypothesis of an airborne origin of soil mutagens, deposition, and an adjacent transformation to non-mutagenic compounds by soil microorganisms. PMID- 11113696 TI - Analyses of spontaneous pink mutant events in the stamen hairs of Tradescantia clone BNL 4430 cultivated in a nutrient solution circulating growth chamber. AB - In order to obtain more fundamental data on Tradescantia clone BNL 4430, one of the most suitable testers for environmental mutagens, the occurrences of spontaneous somatic pink mutations in the stamen hairs were scored for 52 weeks from 12 December 1998 to 10 December 1999, cultivating the young inflorescence bearing shoots with roots in a nutrient solution circulating (NSC) growth chamber. The environmental conditions in the chamber were 22.0+/-0.5 degrees C during the 16h day with the light intensity of 7.5klx from white fluorescent tubes, and 20.0+/-0.5 degrees C at night. During the scoring period, 697,443 stamen hairs with an average cell number of 25.36 were observed and 2642 pink mutant events (PMEs) were detected. The overall spontaneous mutation frequency was 1.56+/-0.03 PMEs per 10(4) hair-cell divisions, and the frequency was significantly lower in May, July and August and significantly higher in November and December. By analyzing the sectoring patterns of 1856 PMEs (70.25% of PMEs detected), the most of 172 cases of multiple (two to five) pink sectors observed in the same hairs (scored as 232 PMEs for calculating mutation frequency) were found to be the results of events involving somatic recombinations occurred in single cells or cell lineages, rather than those of two or more independent somatic mutations occurred in different cells. This finding clearly shows the significance of somatic recombinations in producing such multiple sectors (382 sectors in total) which occupied 19.0% of the 2006 pink sectors in total analyzed. Somatic recombinations were considered to be playing a significant role also in producing single PMEs in the stamen hairs. PMID- 11113697 TI - Induction of micronuclei in human lymphocytes exposed in vitro to microwave radiation. AB - Increasing applications of electromagnetic fields are of great concern with regard to public health. Several in vitro studies have been conducted to detect effects of microwave exposure on the genetic material leading to negative or questionable results. The micronucleus (MN) assay which is proved to be a useful tool for the detection of radiation exposure-induced cytogenetic damage was used in the present study to investigate the genotoxic effect of microwaves in human peripheral blood lymphocytes in vitro exposed in G(0) to electromagnetic fields with different frequencies (2.45 and 7.7GHz) and power density (10, 20 and 30mW/cm(2)) for three times (15, 30 and 60min). The results showed for both radiation frequencies an induction of micronuclei as compared to the control cultures at a power density of 30mW/cm(2) and after an exposure of 30 and 60min. Our study would indicate that microwaves are able to cause cytogenetic damage in human lymphocytes mainly for both high power density and long exposure time. PMID- 11113698 TI - Inter- and intra-individual variation of faecal water - genotoxicity in human colon cells. AB - Exogenous nutritional factors modulate the faecal contents leading to an enhanced or reduced burden with toxic and cancerogenic factors. These factors are thought to contribute to colon cancer by inducing mutations or enhancing proliferation in colon cells. Faecal water more or less causes these effects in model systems and thus could be the basis for valuable biomarker approaches. Our investigations are aimed at determining geno- and cytotoxicity of faecal water in human colon cell lines in vitro. We are developing techniques for their applicability as biomarker tests during dietary intervention studies. Faecal water is isolated by centrifugation of the faeces at 25000xg and added to cultured human colon cells (HT29). Membrane damage as assessed by trypan blue exclusion is determined as a measure for cytotoxicity. Semiquantitative analysis of inducible DNA damage (breaks and alkali labile sites) are analysed with the single cell microgelelectrophoresis assay (comet-assay) and oxidised DNA bases by the additional use of repair specific enzymes. We have now determined baseline toxic activities and calculated inter- and intra-individual and -experimental coefficients of variation for faecal water from different subjects consuming similar or different diets. Most faecal water induced DNA damage and oxidised DNA bases in HT29 clone 19a cells (0.9-9.14 fold and 1.7-4.9 fold, respectively in comparison to the NaCl controls). Intra- and inter-experimental coefficients (CV) of variation, were in a similar order of magnitude and ranged from 6.9 to 31.4. In contrast both intra- and inter-individual variability were considerably higher (CV-ranges of 29.7-76.6 and 21.3-64.0, respectively). Interestingly, these inter individual values were not lowered when subjects consumed identical diets (CV ranges of 28.4-126.0). However, following intervention with certain protective dietary regimens (e.g. lignan containing bread) significant reductions of faecal water-induced genotoxicity can be observed. Therefore, in spite of the expected and observed degrees of variation in this methodology, effective experimental protocols may still lead to detectable modulations of the level of toxic and genotoxic effects. PMID- 11113699 TI - Evidence that chlorophyllin (CHLN) may behave as an inhibitor or a promoter of radiation-induced genetic damage in somatic cells of drosophila. AB - Irradiation of 96h old Drosophila following a 24h pretreatment with 5% chlorophyllin (CHLN) was delayed 0-4 days. The antimutagenic effect of CHLN in somatic cells monitored by the wing spot test persisted for 3 days after completion of the pretreatment and appeared to terminate at a time corresponding to the cessation of mitotic divisions of wing anlagen cells. Within the same population of cells, CHLN demonstrated both an inhibitory effect as measured in mwh single spot classes, and contrarily, a promoting effect in the class of mwh/flr twin spots and to an extent in the class of large flr spots. The reason for the contrasting effects of CHLN remains to be determined. PMID- 11113700 TI - Evaluation of the in vitro genetic toxicity of 4-(2, 4-dichlorophenoxy)butyric acid. AB - The herbicide 4-(2,4-dichlorophenoxy)butyric acid (2,4-DB) is principally used in the USA on peanuts, soybeans and alfalfa. In Europe, it is used on undersown spring barley and grassland (with clover). The genetic toxicity in vitro of the dimethylamine salt of 2,4-DB was examined by employing a range of end points including gene mutation in bacteria (Ames test) and mammalian cell cultures (CHO/HGPRT assay), cytogenetic abnormalities in mammalian cells (CHO/chromosomal aberration assay), and induction of DNA damage and repair in rat hepatocytes. There were no indications of genotoxic potential for 2,4-DB in the first three of these assays. One of the two criteria for a positive response in the UDS assay was exceeded but the increases did not exceed the second criteria for a positive response. The test material was therefore evaluated as weakly active in this assay. The weight of the evidence clearly indicates that 2, 4-DB is not genotoxic to mammals and are consistent with the reported lack of carcinogenic potential for 2,4-DB in both mice and rats. PMID- 11113701 TI - Cytogenetic response without changes in peripheral cholinesterase enzymes following exposure to a sheep dip containing diazinon in vivo and in vitro. AB - Occupational exposure to organophosphorus insecticides (OPs), such as diazinon, may be monitored by the measurement of the activity of peripheral cholinesterase enzymes, including erythrocyte acetylcholinesterase (EAChE) and plasma or serum cholinesterase (plasma or serum ChE). Exposures have also been measured by the analysis of dialkyl phosphate metabolites of OPs in urine. The potential health risks associated with exposure, especially those of a neurological nature, may then be estimated, and appropriate measures to reduce or eliminate exposures can be implemented. There is evidence that some OP pesticides may have in vivo genotoxic effects, suggesting a possible link with cancer with long term or repeated heavy exposures. This paper describes work performed in 17 subjects with a single or two exposures to a sheep dip containing diazinon. Urine samples revealed OP metabolites dimethylphosphate (DMP), dimethylthiophosphate (DMTP), diethylphosphate (DEP) and diethylthiophosphate (DETP) in 37% of subjects at low levels which were not elevated after exposure. EAChE and plasma ChE were also unchanged before and after exposure, and were similar to those measured in unexposed control groups. Sister chromatid exchanges (SCE), a marker of chromosome damage, was significantly elevated in peripheral blood lymphocytes after exposure compared with before. SCE were unchanged in a group of non occupationally exposed workers. In vitro studies with both authentic diazinon (98%) and diazinon in a sheep dip formulation (45%) showed increased SCE and decreased replicative indices, suggesting toxic and genotoxic effects of diazinon. PMID- 11113702 TI - The genotoxicity of 3-nitrobenzanthrone and the nitropyrene lactones in human lymphoblasts. AB - Polycyclic aromatic hydrocarbons (PAH) and nitrated polycyclic aromatic compounds (nitro-PAC) have been found to be mutagenic in bacterial and human cells as well as carcinogenic in rodents. In this investigation, the genotoxic effects of 3 nitrobenzanthrone (3NB) and a mixture of nitropyrene lactones (NPLs) were determined using forward mutation assays performed in two human B-lymphoblastoid cell lines, MCL-5 and h1A1v2, which are responsive to the nitro-PAC class of compounds. Mutagenicity of the compounds was determined at the heterozygous tk locus and the hemizygous hprt locus, thus, identifying both large-scale loss of heterozygosity (LOH) events as well as intragenic mutagenic events. Genotoxicity was also determined using the CREST modified micronucleus assay, which detects chromosomal loss and breakage events. Results indicate 3NB is an effective human cell mutagen, significantly inducing mutations at the tk and hprt loci in both cell lines, and inducing micronuclei in the h1A1v2 cell line. The NPL isomers are also mutagenic, inducing mutations at the two loci as well as micronuclei in both cell lines. Because of their mutagenic potencies and their presence in ambient air, further assessments should be made of human exposures to these nitro-PAC and the potential health risks involved. PMID- 11113703 TI - Micronuclei and gene mutations in transgenic big Blue((R)) mouse and rat fibroblasts after exposure to the epoxide metabolites of 1, 3-butadiene. AB - 1,3-Butadiene (BD) is a commodity compound and by-product in the manufacture of synthetic rubber that elicits a differential carcinogenic response in rodents after chronic exposure. Mice are up to approximately 1000-fold more sensitive to the tumorigenicity of inhaled BD than rats, thereby confounding human risk assessment analyses. Rodent transgenic in vivo and in vitro models have been recently utilized for generating genetic toxicology data in support of risk assessment studies. However, studies have not been extended to investigate multiple endpoints of genetic damage using in vitro transgenic models. The goal of this study was to evaluate possible differences in the production of genetic damage in transgenic Big Blue((R)) mouse (BBM1) and rat (BBR1) fibroblasts exposed to three predominant epoxide metabolites of BD. Analyses of cytotoxicity, micronucleus (MN) formation, cII mutant frequency (MF) and apoptosis were assessed after in vitro exposure of BBM1 and BBR1 cells exposed to various concentrations of butadiene monoepoxide (BMO), diepoxybutane (DEB) and butadiene diolepoxide (BDE). Both BMO and DEB reduced cell survival in BBM1 and BBR1 cells. However, BDE decreased cell survival only in BBM1 cells at the concentrations evaluated. Concentration-dependent increases in the formation of MN was observed in both BBM1 and BBR1 cells, with DEB being the most potent followed by BDE and then BMO. The dose-response for mutations induced at the cII locus was essentially equal after DEB exposure of BBM1 and BBR1 fibroblasts. In contrast, the cII MF was significantly increased only in BBM1 cells after exposure to either BMO or BDE. These data demonstrate a differential genetic response for gene mutations but not for MN formation in transgenic BBM1 and BBR1 fibroblasts and suggest a rodent species-specific difference in the persistence of DNA damage that results in gene mutations. In addition, apoptosis was observed in BBR1 cells but not in BBM1 cells when treated with any of the three BD epoxide metabolites. This response may partially explain the differential response to mutations induced by BMO and BDE. These data offer insight into specific differences in mouse and rat cells with respect to their response to BD epoxide metabolites. Such data may help to explain the different tumorigenicity results observed in rodent BD carcinogenicity studies. PMID- 11113704 TI - Analysis of DNA adducts by accelerator mass spectrometry in human breast tissue after administration of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and benzo[a]pyrene. AB - Epidemiological evidence has suggested an association between meat consumption and the risk of breast cancer. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a heterocyclic amine found in cooked meat, has been implicated in the aetiology of breast cancer and has been shown to induce tumour formation in rodent mammary glands. In addition, polycyclic aromatic hydrocarbons, such as benzo[a]pyrene (B[a]P) which has also been shown to induce tumour formation at a number of sites in rodents including the breast, are produced during the cooking of meat through the pyrolysis of fats. The aim of this study was to examine the bioavailability of these compounds to human breast tissue and their ability to bind to DNA to form DNA adducts. Patients undergoing breast surgery at York District Hospital were orally administered prior to surgery a capsule containing 20microg of 14C PhIP (182kBq, specific activity 2.05GBq/mmol) or 5microg of 14C B[a]P (36kBq, specific activity 1.81GBq/mmol). At surgery, normal and tumour breast tissue was resected and tissue concentrations of carcinogen measured by liquid scintillation counting and DNA adduct levels by accelerator mass spectrometry (AMS) were subsequently determined. It was found that both 14C PhIP and 14C B[a]P were able to reach the target organ where they had the ability to form DNA adducts. The level of adducts ranged from 26.22-477.35 and 6.61-208. 38 adducts/10(12) nucleotides following administration of 14C PhIP and 14C B[a]P, respectively, with no significant difference observed between levels in normal or tumour tissue. In addition, the data obtained in this study were comparable to adduct levels previously found in colon samples following administration of the same compounds to individuals undergoing colorectal surgery. This is the first report that these two carcinogens bind to human breast DNA after administration of a defined low dose. PMID- 11113705 TI - Bioactivation of diesel exhaust particle extracts and their major nitrated polycyclic aromatic hydrocarbon components, 1-nitropyrene and dinitropyrenes, by human cytochromes P450 1A1, 1A2, and 1B1. AB - The genotoxicities of four samples of diesel exhaust particle (DEP) extracts (DEPE) and nine nitroarenes found in DEPE were investigated after activation catalyzed by human cytochrome P450 (P450) family 1 enzymes co-expressed with NADPH-cytochrome P450 reductase (NPR) in Escherichia coli membranes. The DEPE samples induced umu gene expression in Salmonella typhimurium TA1535/pSK1002 without any P450 system and were further activated by human P450 1B1/NPR membranes. Moderate activation of the DEPE sample by P450 1A2/NPR membranes was also observed, but not by either P450 1A1/NPR or NPR membranes. 1-Nitropyrene (1 NP) was strongly activated by human P450 1B1/NPR membranes. 1,8-Dinitropyrene (1,8-DNP) was most highly activated by P450 1A1 and 1B1 systems for the three DNPs tested. In contrast, 1, 3-DNP was inactivated by P450 1A1/NPR, 1A2/NPR, and 1B1/NPR systems and slightly activated by NPR membranes. 2-Nitrofluoranthene (2 NF) and 3-nitrofluoranthene (3-NF) showed activities similar to 1-NP after bioactivation by P450 1B1/NPR membranes. However, the genotoxicities of 6 nitrochrysene, 7-nitrobenz[a]anthracene, and 6-nitrobenzo[a]pyrene were all weak in the present assay system. Apparent genotoxic activities of DEPE were very low compared with standard nitroarenes in the presence of P450s, possibly because unknown component(s) of DEPE had inhibitory effects on the bioactivation of 1-NP and 1,8-DNP catalyzed by human P450 1B1. These results suggest that environmental chemicals existing in airborne DEP, in addition to 1-NP, 1,6-DNP, 1,8-DNP, 2-NF, and 3-NF, can be activated by human P450 1B1. Biological actions of air pollutants such as nitroarenes to human extrahepatic tissues may be of concern in tissues in which P450 1B1 is expressed. PMID- 11113706 TI - Protective effects of hemin and tetrakis(4-benzoic acid)porphyrin on bacterial mutagenesis and mouse skin carcinogenesis induced by 7, 12 dimethylbenz[a]anthracene. AB - Porphyrins which are widespread in nature can interfere with the actions of certain carcinogens and mutagens, and have also been used clinically in photodynamic therapy (PDT) of tumors. Porphyrins such as chlorophyll, chlorophyllin (CHL) and hemin are known to inactivate various mutagens by forming complexes with them. Tetrakis(4-benzoic acid)porphyrin (TBAP) has been developed as a photosensitizer for PDT and its metal complex, MnTBAP has been shown to be efficacious in a variety of in vitro and in vivo oxidative stress models of human diseases. In the present study, we have found that TBAP and hemin exert concentration-related inhibition of his(+) reversion in Salmonella typhimurium TA100 induced by 7, 12-dimethylbenz[a]anthracene (DMBA), and significantly reduced both incidence and multiplicity of skin tumors when topically applied prior to treatment of 12-O-tetradecanoylphorbol-13-acetate in female ICR mice. Covalent DNA binding of DMBA in mouse skin was also significantly inhibited by topical application of TBAP or hemin as well as CHL. These results suggest the chemopreventive potential of compounds containing a porphyrin nucleus. PMID- 11113707 TI - The genetic toxicity of 3,3',4,4'-tetrachloroazobenzene and 3,3',4, 4' tetrachloroazoxybenzene: discordance between acute mouse bone marrow and subchronic mouse peripheral blood micronucleus test results. AB - 3,3',4,4'-Tetrachloroazobenzene (TCAB) and 3,3',4, 4'-tetrachloroazoxybenzene (TCAOB) are dioxin-like chemicals that were investigated for toxicity in 13-week gavage studies in male and female B6C3F(1) mice and F344N rats by the National Toxicology Program. As part of the comprehensive toxicological investigation of these chemicals, peripheral blood smears from mice treated 5 days per week for 13 weeks with 0.1-30mg/kg/day TCAB or TCAOB were analyzed for the frequency of micronucleated (MN) normochromatic erythrocytes (NCE). Both chemicals produced significant increases in MN-NCE in male and female mice. In contrast to these positive results in subchronic exposure studies, no significant increases were seen in acute bone marrow MN tests in male mice administered three daily injections of 50-200mg/kg/day TCAB and TCAOB. The results with TCAB and TCAOB suggest that the routine integration of MN tests with subchronic toxicity studies may allow detection of mutagenic activity for some chemicals that fail to elicit responses in short-term, high dose tests. In addition, the integration of mutagenicity tests into general toxicity tests reduces the use of laboratory animals and the cost of the testing. PMID- 11113708 TI - Micronucleus induction and chromosome loss in transformed human white cells indicate clastogenic and aneugenic action of the cyanobacterial toxin, cylindrospermopsin. AB - Cylindrospermopsin (CYN) is a potent inhibitor of protein synthesis produced by a number of cyanobacterial species, the most common being Cylindrospermopsis raciborskii. CYN contains a uracil moiety attached to a sulphated guanidino moiety, suggesting that it may have carcinogenic activity. This report describes the use of the WIL2-NS lymphoblastoid cell-line in the well-validated cytokinesis block micronucleus (CBMN) assay to test this hypothesis. Centromeres (CENs) were identified in micronuclei (MNi) of binucleated cells (BNCs) by fluorescent in situ hybridisation of alpha centromeric DNA sequence repeats. The results indicate that CYN induced a significant increase in the frequency of MNi in BNCs exposed to 6 and 10microg/ml, and a significant increase in CEN-positive MNi at all concentrations of CYN tested (1, 3, 6, and 10microg/ml). However, despite this apparently greater sensitivity of WIL2-NS cells to induction of CEN-positive MNi at low CYN concentrations, at the higher concentrations the magnitude of the increase in CEN-positive MNi did not account for the greater increase in MNi in BNCs, indicating that both CEN-positive and CEN-negative MNi were induced. This suggests that CYN acts to induce cytogenetic damage via two mechanisms, one at the level of the DNA to induce strand breaks, the other at the level of kinetochore/spindle function to induce loss of whole chromosomes (aneuploidy). C. raciborskii occurs in a number of human drinking water sources worldwide and so these findings may have important public health implications. PMID- 11113709 TI - Assessment of potential mutagenic activities of a novel benzothiazole MAO-A inhibitor E2011 using Salmonella typhimurium YG1029. AB - The potential initiation activities of a novel monoamine oxidase type-A (MAO-A) inhibitor E2011, which induced preneoplastic foci in the rat liver, were investigated by comparing the mutagenic activity of E2011, 6-aminobenzothiazole (6-ABT, a structural scaffold of E2011) and its derivatives, which are suggested primary reactive metabolites for E2011-induced hepatotoxicity in the rats in vivo, in the Ames assay system employing two Salmonella tester strains, TA100 and YG1029, a bacterial O-acetyltransferase-overproducing strain of TA100. E2011, a tertiary amine, showed no mutagenic activity both in the Salmonella typhimurium TA100 and YG1029 with and without S9 mix. On the other hand, a secondary aromatic amine ER-174238-00, a typical decarbonated metabolite of E2011, showed weak but significant mutagenicity in YG1029 in the presence of S9 mix, and a primary aromatic amine ER-174237-00, an N-dealkylated derivative of ER-174238-00, exhibited S9-dependent potent mutagenicity in YG1029. Thus, it appears that primary and secondary amino moieties of benzothiazole derivatives at C(6) position are the specific structures contributing to their mutagenic activity. In addition, the alkyl group at C(2)-position of E2011, ER-174237-00 and ER-174238 00 is suggested to intensify the mutagenic activity, since the mutagenicity of ER 174237-00 is approximately two-fold higher than that of 6-ABT, which has hydrogen at C(2)-position in the place of the alkyl group. These results strongly suggest that E2011 has potential initiation activities in the rat liver in vivo after undergoing decarbonation, one of the metabolic pathways, at the carbonyl moiety of oxazolidinone ring to form mutagenic amine(s). PMID- 11113710 TI - Annual scientific and financial report Working Group on Heart Failure European Society of Cardiology: September 1999-August 2000. PMID- 11113711 TI - Thromboembolism in heart failure: who should be treated? AB - The risk of thromboembolic complications in patients with heart failure and/or chronic left-ventricular systolic dysfunction is increased. Nevertheless, anticoagulant therapy in these patients is still a subject of debate. Atrial fibrillation is the only prospectively evaluated, proven thromboembolic risk factor and patients with atrial fibrillation benefit from long term anticoagulant therapy. The significance of other proposed thromboembolic risk factors in heart failure and/or chronic left-ventricular dysfunction such as gender, cause of myocardial disease, severity of heart failure, left-ventricular ejection fraction, left-ventricular thrombus, left ventricular aneurysm and history of previous thromboembolic event is less clear. This article summarizes key studies, assesses the incidence of thromboembolism, evaluates risk factors and proposes guidelines for anticoagulation of patients with heart failure and/or left ventricular systolic dysfunction. PMID- 11113712 TI - Mitral valve repair in heart failure. AB - Mitral regurgitation (MR) is a frequent complication of end-stage heart failure. Historically, these patients were either managed medically or with mitral valve replacement, both associated with poor outcomes. Mitral valve repair via an 'undersized' annuloplasty repair is safe and effectively corrects MR in heart failure patients. All of the observed changes contribute to reverse remodeling and restoration of the normal left-ventricular geometric relationship. Mitral valve repair offers a new strategy for patients with MR and end-stage heart failure. PMID- 11113713 TI - Progressive cardiac dysfunction in adriamycin-induced cardiomyopathy rats. AB - Cardiotoxicity is a limiting factor in the treatment of cancer with adriamycin. We administered adriamycin by a method which minimizes the risk of peritonitis in an adriamycin-induced cardiomyopathy rat model. Sixty male Wistar rats were given 1 mg/kg of adriamycin intraperitoneally 15 times over a 3-week period (total dose, 15 mg/kg) to induce the cardiomyopathy model. Fifteen control rats received 10 ml/kg body wt. saline 15 times over 3 weeks. The animals were observed for 12 weeks and assessed for mortality, and cardiac volume and function was analyzed by echocardiography at 4, 8, and 12 weeks. In rats treated with adriamycin, the cumulative mortality was 35.8% while in the controls, none of the rats died. Left ventricular diameter of the systole (LVDs) was significantly increased at 4 weeks (4.5 vs. 3.3 mm; P<0.001). Left ventricular diameter of the diastole (LVDd) was significantly increased at 12 weeks (7.9 vs. 7.0 mm; P<0.01) and the % fractional shortening (FS) was significantly decreased at 8 weeks (33.4% vs. 50.0%; P<0.01) in the adriamycin-treated rats. This administration method appears to be useful for investigating the cardiac effect of adriamycin while avoiding the influence of peritonitis typically caused by an intraperitoneal injection of higher single doses of adriamycin. PMID- 11113714 TI - Relationship between left ventricular wall stress and ANP gene expression during the evolution of rapid ventricular pacing-induced heart failure in the dog. AB - We have recently described a modified model of progressive rapid ventricular pacing-induced heart failure which evolves over a period of 38 days. To further characterize left ventricular remodeling during the progression of heart failure, we assessed left ventricular geometry, wall stress, and atrial natriuretic peptide (ANP) gene expression and protein content during control conditions, asymptomatic left ventricular dysfunction, and overt congestive heart failure (CHF). Although asymptomatic left ventricular dysfunction was characterized by a significant increase in systolic and diastolic left ventricular dimension (+30% and +6%, respectively, P<0.05 each) and a marked increase in left ventricular systolic wall stress (+68%, P<0.01), left ventricular ANP gene expression was unchanged as compared to control. In contrast, strong left ventricular ANP gene expression (+449%, P<0.05) was observed during overt CHF in the absence of further significant increases in left ventricular systolic wall stress. The onset of strong left ventricular ANP gene expression was associated with increased ANP content (+88%, P<0.05) and left ventricular mass index (+13%, P<0.05). In contrast, left atrial ANP gene expression and left ventricular diastolic wall stress increased progressively during asymptomatic left ventricular dysfunction (+39%, P=n.s. and +131%, P<0.01) and overt CHF (+76% and +336% vs. control, P<0.01 each). Progressive rapid ventricular pacing is associated with the induction of left ventricular ANP gene expression and protein synthesis exclusively during overt CHF. The current studies provide new insight into the temporal pattern of ANP-activation and the disparity between left ventricular systolic wall stress and ANP-activation in a large animal model of progressive CHF. PMID- 11113715 TI - The effect of valvular regurgitation on plasma Cardiotrophin-1 in patients with normal left ventricular systolic function. AB - BACKGROUND: Cardiotrophin-1 (CT-1), a member of the interleukin-6 related cytokine family that act via the gp130 signalling pathway, has been shown to stimulate the assembly of sarcomeric units in series in cardiomyocytes resulting in eccentric hypertrophy, ventricular dilatation and finally loss of function. In situations of volume overload a similar form of eccentric hypertrophy occurs with time. AIMS: We hypothesised that plasma CT-1 would be raised in patients with significant mitral, tricuspid and/or aortic regurgitation (MR/TR or AR, respectively) when compared to those with no (or mild) valvular regurgitant lesion. METHODS: A novel competitive immunoluminometric assay using an in-house polyclonal antibody to amino acids 105-120 of the CT-1 sequence was developed. Seventy-eight patients (31 male, mean+/-S.D. age 63.5+/-17.9 years), all with normal left ventricular systolic function were studied. Results are expressed as mean+/-S.D. fmol/ml. RESULTS: Sixty-three subjects had no significant valvular lesion, seven had moderate/severe MR, nine had moderate/severe TR and four had moderate/severe AR. These subjects had CT-1 concentrations of 53. 3+/-23.2, 90.5+/-44.4, 72.6+/-43.8 and 48.4+/-24.4, respectively (P=0.02, ANOVA). Mean log CT-1 was higher in those with moderate/severe MR when compared to those without a significant regurgitant valvular lesion (P<0.03). The only predictor of moderate/severe MR was log CT-1 (P=0.004). CONCLUSION: These results suggest that plasma CT-1 is raised in those patients with moderate/severe MR in the presence of normal left ventricular systolic function. This secretion of CT-1 could potentially be the cause of ventricular dilatation and subsequent loss of contractile function in these patients. It also offers the intriguing possibility that plasma CT-1 could be used to monitor progression of mitral regurgitation biochemically. PMID- 11113716 TI - Serum erythropoietin in heart failure patients treated with ACE-inhibitors or AT(1) antagonists. AB - BACKGROUND: Erythropoietin (Epo), a growth factor produced by the kidney, is important in heart failure patients to promote oxygen delivery to tissues. Seventy-two chronic heart failure (CHF) patients at our outpatient clinic were subjected to morning serum Epo-level measurements and classified according to NYHA criteria. RESULTS: Forty-eight patients of classes III and IV had a significantly elevated serum Epo-level of 42.9+/-40.3 mIU/ml (mean+/-1 S.D.) when compared to the mean level of 24 patients of classes I and II who had a normal range mean value of 13.4+/-6.2 mIU/ml (P<0.05). Patients on angiotensin converting enzyme (ACE) inhibitors showed a trend towards lower serum Epo-levels compared to patients treated with angiotensin-II type-1 receptor antagonists (AT(1) antagonists) (levels: 33.3+/-35.6 mIU/ml and 43.6+/-38.1 mIU/ml). This trend did not, however, reach statistical significance (P=0.36). CONCLUSION: We suggest that a desirable Epo increase in class III and IV CHF patients could be achieved by either recombinant human Epo administration or, possibly, by appropriate selection of the concomitant medical therapy. A large prospective study shall investigate the possible advantage of AT(1) antagonists over ACE inhibitors with regard to Epo effect. PMID- 11113717 TI - Reduction of hospital days by biventricular pacing. AB - The health care costs for heart failure are substantial. Studies indicate that hospital treatment constitutes 65-75% of these. The aim of this study was to assess total and heart failure related hospital days as well as safety and efficacy of biventricular pacing in 16 patients with severe heart failure and delayed intraventricular conduction (QRS duration >150 ms). They were implanted with a biventricular pacemaker and followed by NYHA class, 6-min walk test and quality of life for a mean of 291+/-76 days. Total number of hospital days and the need for hospitalisations were monitored. Thirteen responders improved by at least one functional class. After 6 months of pacing the 6-min walk test improved from 375+/-83 m to 437+/-73 m (P<0.001) and Minnesota Living with Heart Failure quality of life score from 41+/-19 to 24+/-17 (P<0.001) compared to baseline. The need for hospital care decreased significantly after biventricular pacing. The total number of hospital days in all patients was 253 the year before compared to 45 the year after biventricular pacing (P<0.01). For heart failure related hospital days the corresponding figures were 183 and 39 days, respectively (P<0.01). Biventricular pacing improved 13/16 patients with severe heart failure and wide QRS complexes in this open study. The improvement resulted in a reduced need for hospital care. PMID- 11113718 TI - Effect of beta 1 blockade with atenolol on progression of heart failure in patients pretreated with high-dose enalapril. AB - BACKGROUND: The survival benefit of beta-blocker treatment in patients with heart failure has been established in recent trials. Yet, the impact of beta-blockers added on high dose angiotensin converting enzyme inhibitors has not been reported. AIMS: To investigate the effect of atenolol, a hydrophilic, selective beta1-adrenergic antagonist, added on enalapril 40 mg/day in patients with advanced left ventricular dysfunction in a double-blind placebo-controlled trial. METHODS: One hundred and nineteen patients with class II or III heart failure, left ventricular ejection fraction < or = 25% and treatment with 40 mg enalapril daily were given an initial challenge dose of atenolol 12. 5 mg. One hundred patients (54 with idiopathic, 28 with ischemic, 18 with other dilated cardiomyopathy) tolerated challenge and were randomized to atenolol (maintenance dose 89+/-11 mg/day, range 50-100 mg/day) or placebo. The primary endpoint was combined worsening heart failure or death within 2 years, the secondary endpoint was hospitalization for cardiac events. RESULTS: After 395+/-266 days interim analysis revealed a significant difference between the atenolol and placebo group (log rank P<0.01) and the trial was concluded. Twenty-seven patients had developed worsening heart failure (8 in the atenolol group vs. 19 in the placebo group) and 13 patients had died (5 in the atenolol vs. 8 in the placebo group). Overall there were 23 hospitalizations for cardiac events (6 in the atenolol group vs. 21 in the placebo group, P=0.07); 17 hospitalizations were due to worsening heart failure (5 in the atenolol group, 12 in the placebo-group, P=0.05) and 10 due to arrhythmias (1 in the atenolol group vs. 9 in the placebo group, P<0.01) CONCLUSIONS: The data suggest that in patients with advanced left ventricular dysfunction, beta-blockers can provide substantial benefits supplementary to that already achieved with high dose enalapril treatment. PMID- 11113719 TI - Heart failure in patients seeking medical help at outpatients clinics. Part I. General characteristics. AB - BACKGROUND: During the last decade, the beneficial changes in lifestyle and in medical care increased average life expectancy, particularly in patients with chronic diseases such as hypertension and coronary heart disease. Unfortunately this also increased the number of patients, particularly among the elderly, who are susceptible to complications of these conditions such as heart failure. Uncontrolled hypertension is known to be a primary cause of heart failure and is also known to be very prevalent and frequently uncontrolled in the Polish population. AIM: To estimate the prevalence and characteristics of heart failure among patients of 65 years and older seeking medical care in outpatient clinics in Poland. METHODS: The study is a cross-country epidemiological project in which 417 physicians from outpatient clinics were asked to register 50 consecutive patients aged 65 years and above seeking medical care for any cause. Information on case history, physical examination (diagnosis of heart failure, NYHA class, heart failure symptoms), laboratory tests (resting ECG, chest X-ray, echocardiogram) and data concerning pharmacology management during the 2 weeks prior to the index visit was obtained. RESULTS: Over 5 months, 19877 eligible patients (7324 men and 12553 women) presented to the 417 participating physicians (90% physicians registered 46-50 patients). Among the patients, 53% were diagnosed with heart failure (3901 men and 6678 women), prevalence did not differ by gender. Among patients with heart failure there were 38% of men in NYHA class III or IV and 34% of women. Coronary heart disease was a predominant cause of heart failure in 87% of men (26% of cases with isolated coronary heart disease, 53% with concomitant hypertension and 8% with other diseases), while percentages for women were 80% (15%, 61% and 4%, respectively). Isolated hypertension was a further cause of heart failure in 8% of men and 13% of women. Cardiac arrhythmia was found in approximately 20% of patients, enlargement of heart size in 32% of patients and peripheral leg edema in 54% of men and 64% of women. These symptoms increased with age. Chest X-ray revealed cardiomegaly in 68% of men and women and increased cardiothoracic ratio (>50%) in approximately 40% of patients. From resting ECGs, cardiac arrhythmia was recorded in 21% of patients with heart failure, with atrial fibrillation as a predominant disorder (19%). Left ventricular hypertrophy on resting ECG was noted in 42% of men and women and old myocardial infarction or cardiac ischemia was diagnosed in 71% of men and 66% of women. CONCLUSIONS: (1) Heart failure was diagnosed in over half of outpatients aged 65 and older; in more than a third of these it was NYHA class III and IV. (2) Outpatients with heart failure had a high frequency of co-existing diseases such as arrhythmia, coronary heart disease and hypertension. PMID- 11113720 TI - A Rapid Access Heart Failure Clinic provides a prompt diagnosis and appropriate management of new heart failure presenting in the community. AB - BACKGROUND AND AIMS: The diagnosis of heart failure is an important clinical problem and yet reported diagnostic accuracy in primary care is less than 50%. We established a Rapid Access Heart Failure Clinic (RAHFC) in a district general hospital serving a population of 292,000 in SE London, UK, to diagnose and manage new cases of heart failure presenting for the first time in the community. METHODS: Patients with suspected new onset heart failure were referred by their Primary Care Physician without appointment for clinical assessment on the same or next working day. Assessment by a specialist registrar in cardiology included history, examination, chest X-ray, electrocardiogram (ECG) and echocardiogram. When a diagnosis of heart failure was made appropriate treatment, including angiotensin converting enzyme inhibitors (ACEI), was started. RESULTS: Over 15 months 383 patients were seen (0.4 cases/100,000 population/weekday) 178/383 (46%) were considered to have definite or possible heart failure at the initial assessment in the RAHFC. A normal ECG (negative predictive value 94%) and chest X ray virtually excluded the diagnosis of heart failure. After subsequent specialist investigations and follow-up, including a trial of therapy where appropriate, 101/383 (26%) were finally diagnosed as clinical heart failure. ACEI therapy was commenced in 56/57 (98%) of patients in whom it was considered appropriate. CONCLUSION: The RAHFC provided rapid assessment, prompt diagnosis and early introduction of life prolonging therapy for patients presenting with suspected heart failure in the community. PMID- 11113721 TI - Survival and prognosis: investigation of Crataegus extract WS 1442 in congestive heart failure (SPICE)--rationale, study design and study protocol. AB - SPICE is the first, international, randomized, placebo-controlled, double-blind study to investigate the influence of the herbal drug Crataegus Special Extract WS 1442 (hawthorn leaves with flowers) on mortality of patients suffering from congestive heart failure. BACKGROUND: In vitro and experimental animal studies have suggested the following pharmacological modes of action of standardized Crataegus extracts: (1) cAMP-independent positive inotropy; (2) peripheral and coronary vasodilation; (3) protection against ischemia-induced ventricular arrhythmias; (4) antioxidative properties; and (5) anti-inflammatory effects. STUDY DESIGN: In this randomized, placebo-controlled, double-blind, international trial (approximately 120 investigational centers in seven European countries), up to 2300 patients with congestive heart failure, New York Heart Association class II and III and markedly impaired left ventricular function, will be enrolled and treated over a period of 24 months. During this time patients receive either two film-coated tablets of 450 mg of the Special Extract WS 1442 standardized to 84.3 mg of oligomeric procyanidines or matched placebo per day in addition to standard therapy for congestive heart failure, such as diuretics, digoxin or digitoxin, beta-adrenoceptor blockers and angiotensin-converting-enzyme inhibitors. The primary outcome variable is the combined endpoint of cardiac death, non-lethal myocardial infarction, and hospitalization due to progression of heart failure. Secondary outcome variables are total mortality, exercise duration, echocardiographic parameters, quality of life as well as pharmacoeconomic parameters. The first patient was included in October 1998. The trial is expected to be completed at the end of 2002. PMID- 11113722 TI - Baseline demographics of the Valsartan Heart Failure Trial. Val-HeFT Investigators. AB - BACKGROUND: The Valsartan Heart Failure Trial (Val-HeFT) is the first large-scale randomized, multinational clinical study to assess the efficacy and safety of valsartan, an angiotensin II receptor blocker, added to conventional therapy, including angiotensin-converting enzyme inhibitors, in heart failure patients. A total of 5010 patients with an ejection fraction <40% have been randomized to either valsartan titrated to 160 mg b.i.d. or to placebo. AIMS: Baseline characteristics of patients in Val-HeFT are presented and compared with other major clinical trials in heart failure. METHODS: Baseline data were collected and summary statistics calculated. RESULTS: The study population has a mean age of 62.7 years and is 80% male, 90.3% white, 6.9% black, and 2.8% Asian. Antecedent coronary heart disease is reported in 57.2% of patients. Angiotensin-converting enzyme inhibitors are prescribed for 92.7% of patients, diuretics for 85.8%, digoxin for 67.3%, and beta-blockers for 35.6%. Subgroup comparisons by age, sex, race and ejection fraction quartile show small differences in baseline characteristics. CONCLUSION: Overall the Val-HeFT population is generally representative of the population of patients with mild to moderate heart failure in industrialized countries. PMID- 11113724 TI - Clinical trials update: IMPROVEMENT-HF, COPERNICUS, MUSTIC, ASPECT-II, APRICOT and HEART. AB - Important new studies relevant to the field of heart failure reported at the annual congress of the European Society of Cardiology (ESC), held in Amsterdam in August 2000, are reviewed. The IMPROVEMENT of Heart Failure survey investigated the knowledge and perceptions of over 1300 primary care physicians from 14 ESC member nations and the actual practice in over 11000 of their patients. Guidelines and clinical practice were compared. The survey suggested, in this large sample, that the quality of care was higher than previous smaller surveys have suggested but have also identified important deficiencies in knowledge and management that should be rectified. The COPERNICUS study demonstrated that carvedilol was remarkably well tolerated even in patients with very severe heart failure and that treatment was associated with a substantial reduction in mortality even among patients that would conventionally not be considered, by many, for beta-blocker therapy. The MUSTIC trial suggested that cardiac resynchronisation using biventricular pacing improved patients symptomatically whether or not the patient was in atrial fibrillation. Morbidity and mortality studies of cardiac resynchronisation are now underway. The ASPECT-II and APRICOT II studies investigated the role of warfarin, aspirin and their combination for the long-term management of myocardial infarction. One interpretation of the data from these studies is that the combination of aspirin and warfarin is about as effective as warfarin alone but with a much higher incidence of side effects. Warfarin alone appeared superior to aspirin alone. In summary, the annual congress of the ESC provided important new information for clinical practice and, to date, was, by far, the most important cardiology congress in the world this year. PMID- 11113723 TI - Low doses vs. high doses of the angiotensin converting-enzyme inhibitor lisinopril in chronic heart failure: a cost-effectiveness analysis based on the Assessment of Treatment with Lisinopril and Survival (ATLAS) study. The ATLAS Study Group. AB - OBJECTIVE: A cost-effectiveness analysis of high and low doses of the angiotensin converting enzyme (ACE) inhibitor lisinopril in the treatment of chronic heart failure. METHODS: A cost-effectiveness analysis using data from a randomized controlled trial, ATLAS, where 3164 patients with chronic heart failure were allocated to a high-dose (daily target dose 32.5-35 mg) or low-dose strategy (daily target dose 2.5-5.0 mg) of lisinopril. Differential costs were based on resource use data collected in the trial costed using UK unit costs. Cost effectiveness analysis related differential costs to differential life-years during a 4-year trial follow-up. RESULTS: The mean total number of hospital in patient days per patient was 18. 5 in the high dose group and 22.5 in the low dose group. Over the whole duration of the trial, the mean (S.D.) daily dose of lisinopril in the high-dose group was 22.5 mg (15.7 mg) compared to 3.2 mg (2.5 mg) in the low-dose group. The mean difference in cost per patient was pound sterling 397 lower in the high-dose group [95% CI (high-dose-low-dose) - pound sterling 1263 to pound sterling 436]. Mean life-years per patient were 0.085 years higher in the high-dose group [95% CI (high-dose-low-dose) -0.0074 to 0.1706). Based on mean costs and life-years, high-dose therapy dominates low-dose (less costly and more effective). Allowing for uncertainty in mean costs and life years, the probability of high-dose therapy being less costly than low dose was 82%. If a decision maker is willing to pay at least pound sterling 3600 per life year gained, the probability of high-dose being more cost-effective was 92%. CONCLUSIONS: The ATLAS Study showed that the treatment of heart failure with high doses of lisinopril has a high probability of being more cost-effective than low dose therapy. PMID- 11113726 TI - Prostate-specific antigen and other prostate cancer markers. PMID- 11113727 TI - Development and validation of the expanded prostate cancer index composite (EPIC) for comprehensive assessment of health-related quality of life in men with prostate cancer. AB - OBJECTIVES: Health-related quality of life (HRQOL) is an increasingly important endpoint in prostate cancer care. However, pivotal issues that are not fully assessed in existing HRQOL instruments include irritative urinary symptoms, hormonal symptoms, and multi-item scores quantifying bother between urinary, sexual, bowel, and hormonal domains. We sought to develop a novel instrument to facilitate more comprehensive assessment of prostate cancer-related HRQOL. METHODS: Instrument development was based on advice from an expert panel and prostate cancer patients, which led to expanding the 20-item University of California-Los Angeles Prostate Cancer Index (UCLA-PCI) to the 50-item Expanded Prostate Index Composite (EPIC). Summary and subscale scores were derived by content and factor analyses. Reliability and validity were assessed by test retest correlation, Cronbach's alpha coefficient, interscale correlation, and EPIC correlation with other validated instruments. RESULTS: Test-retest reliability and internal consistency were high for EPIC urinary, bowel, sexual, and hormonal domain summary scores (each r >/=0.80 and Cronbach's alpha >/=0.82) and for most domain-specific subscales. Correlations between function and bother subscales within domains were high (r >0.60). Correlations between different primary domains were consistently lower, indicating that these domains assess distinct HRQOL components. EPIC domains had weak to modest correlations with the Medical Outcomes Study 12-item Short-Form Health Survey (SF-12), indicating rationale for their concurrent use. Moderate agreement was observed between EPIC domains relevant to the Functional Assessment of Cancer Therapy Prostate module (FACT-P) and the American Urological Association Symptom Index (AUA-SI), providing criterion validity without excessive overlap. CONCLUSIONS: EPIC is a robust prostate cancer HRQOL instrument that complements prior instruments by measuring a broad spectrum of urinary, bowel, sexual, and hormonal symptoms, thereby providing a unique tool for comprehensive assessment of HRQOL issues important in contemporary prostate cancer management. PMID- 11113728 TI - Sildenafil taken at bedtime significantly increases nocturnal erections: results of a placebo-controlled study. AB - OBJECTIVES: Nighttime erections occur at all ages and contribute to the maintenance of the morphodynamic integrity of smooth muscle cells within the corpora cavernosa. This study was aimed at evaluating the effect on nocturnal erections of sildenafil versus a placebo taken at bedtime. METHODS: A double blind, crossover, placebo-controlled study design was used to examine the effects of sildenafil and placebo on sleep-related erectile activity. Thirty selected patients with erectile dysfunction (vasculogenic etiology, 22 patients [73%]; psychogenic etiology, 8 patients [27%]) were submitted to a polysomnographic recording of nocturnal erections, using a RigiScan device during 3 consecutive nights. After a first night of adaptation, the 2 following nights were used to study patients after the administration of sildenafil (100 mg) or a placebo taken at bedtime. RESULTS: Twenty-three patients (77%) showed a significantly improved nocturnal erectile activity (according to the calculation of rigidity and tumescence activity units) after the administration of sildenafil (P <0.01), 5 patients (17%) showed comparable nocturnal erections with sildenafil and placebo, and 2 patients (6%) showed a significantly improved nocturnal erectile activity after taking the placebo (P <0.05). Overall, mean rigidity and tumescence activity values at the tip and base of the penis were significantly improved after sildenafil rather than placebo administration (P <0.001). The duration of tip rigidity greater than 60% was significantly longer during the night with sildenafil (P <0. 001). Although the number of erectile episodes was greater during the sildenafil night, this did not reach statistical significance. CONCLUSIONS: In most patients with good sleep efficiency and who have erectile dysfunction, sildenafil, rather than a placebo, taken at bedtime produces a significantly improved nocturnal erectile activity. Further studies are needed to verify whether this preliminary finding may constitute the basis for the use of sildenafil as a tool for preventing erectile dysfunction. PMID- 11113729 TI - Gahat: a Napalese cure for urolithiasis? AB - OBJECTIVES: Gahat (Vigna unguiculata) is a legume used for centuries in Nepal and Pakistan to treat the symptoms associated with urinary calculi. We prospectively evaluated the effect of Gahat consumption on 24-hour urine parameters in an attempt to assess its in vivo effect in normal volunteers. METHODS: Eight non stone-forming volunteers collected 24-hour urine specimens while on their routine diets for baseline data. Urine was analyzed for pH, volume, calcium, citrate, phosphate, sodium, magnesium, uric acid, and oxalate. The Gahat was prepared according to local custom. No additives were used to enhance flavor. The pureed mixture (8 ounces) was ingested three times daily for 2 days. Subjects were instructed to maintain their normal diet, including fluid intake and activity during the study period. Twenty-four hours after the start of Gahat intake, a second 24-hour urine collection was initiated while volunteers continued the Gahat. Results of the urine samples before and after Gahat intake were analyzed, using the paired Student t test. RESULTS: There were no significant differences in urinary electrolytes between the urine samples before and after Gahat intake. Magnesium, urine volume, and uric acid differences approached clinical significance. CONCLUSIONS: Gahat increased urinary magnesium through an unknown mechanism and had no effect on other routine 24-hour urine electrolytes. The increase in urinary volume is attributed to the increase in fluid consumption by the subjects. If this legume is effective in preventing or dissolving urinary calculi, it may act through mechanisms not identified in 24-hour urine electrolytes. PMID- 11113730 TI - Metabolic investigation of recurrent nephrolithiasis: compliance with recommendations. AB - OBJECTIVES: Nephrolithiasis is a recurrent condition with significant associated morbidity and economic impact. Although urologic intervention addresses symptomatic stone episodes, prevention of recurrences with proven medical therapy is indicated. METHODS: This retrospective study examined 97 patients who presented in 1997 and 1998 with recurrent nephrolithiasis in a large tertiary care center for the presence of an appropriate metabolic investigation as recommended by the National Institutes of Health Consensus Conference. Complete data were abstracted from the hospital and private clinic charts. RESULTS: The average patient age was 50.5 years; 61.9% of patients were men. The mean number of stones per patient was 5.6 (range 2 to 62), with stone analysis performed for 78 patients. Fifty-eight stones (74.4%) were calcium oxalate and/or phosphate, 14 (17.9%) urate, 8 (10.3%) struvite, and 3 (3.8%) cystine. Five patients had two stone types on different occasions. Either lithotripsy or a urologic procedure was required for at least one stone presentation in 89 patients (91.8%). An investigation for stone disease was pending in 54 patients (55.7%). A complete evaluation, satisfying the preset criteria, was performed in 34 patients (35.1%). Six patients who did not undergo evaluation were lost to follow-up. Univariate analysis revealed that referral to a nephrologist (P = 0.001), treatment with medications used for stone disease (P = 0.008), and urate stones (P = 0.005) were associated with a complete investigation. Similarly, these were independently associated with a complete evaluation in regression analysis of 77 complete data sets, with odds ratios of 24.4 (nephrology referral), 4.9 (medication use), and 5.6 (urate stones). CONCLUSIONS: The results of this study demonstrate that a significant proportion of patients with recurrent nephrolithiasis do not undergo appropriate metabolic investigations. Efforts should be made to improve the evaluation of these patients. PMID- 11113732 TI - Editorial comment PMID- 11113731 TI - Retroperitoneal laparoscopic adrenalectomy: clinical experience in 52 procedures. AB - OBJECTIVES: Laparoscopic adrenalectomy has become an effective option for removal of small adrenal tumors. The aim of this prospective study was to evaluate the retroperitoneal approach with regard to intraoperative complications, morbidity, and length of hospital stay. METHODS: Between September 1996 and October 1999, we performed 52 laparoscopic adrenalectomies (31 left, 21 right) for benign lesions by a retroperitoneal approach in 44 patients (27 women, 17 men) with a mean age of 46.9 years (range 17 to 74). The average adrenal tumor size was 32 mm (range 10 to 63). All procedures required four trocars and a mean operative time of 135 minutes (range 75 to 240). RESULTS: There was no mortality, conversion rate to open surgery was 1.9%, and estimated blood loss was 80 mL (range 30 to 200). With a mean follow-up of 16 months, morbidity was 17.2%, which included intraoperative complications (5. 7%) with two vascular injuries, and postoperative complications (11. 5%) with wound infections, deep hematoma, and parietal dehiscence. Average length of hospital stay was 5 days with a mean analgesic consumption of 2 days (range 1 to 5). CONCLUSIONS: The retroperitoneal approach in laparoscopic adrenalectomy appears to be a minimally invasive and safe therapeutic option that may become the standard for unilateral or bilateral adrenal tumors not larger than 7 cm. However, a learning curve in laparoscopy is indispensable before starting this type of procedure. PMID- 11113734 TI - Editorial comment PMID- 11113733 TI - Laparoscopic nephrectomy in the markedly obese living renal donor. AB - OBJECTIVES: To determine whether laparoscopic living donor nephrectomy is safe and efficacious in markedly obese renal donors. METHODS: From 1996 to 1999, 431 laparoscopic living donor nephrectomies were performed. The markedly obese group consisted of 41 patients with a body mass index (BMI) greater than 35. Forty-one controls with a BMI less than 30 were matched to the obese donors by sex, age, race, and date of surgery. RESULTS: The markedly obese and control groups were closely matched in sex, race, age, serum creatinine level, creatinine clearance, HLA match to recipient, side of donated kidney, and experience level of the surgeons. The obese patients had a BMI range of 35.2 to 53.9 (mean 39.3), and the control patients had a BMI range of 18.4 to 29.0 (mean 24.3). Donor operations in the markedly obese were significantly longer by an average of 40 minutes. The greater intraoperative blood loss and longer extraction incision length seen in the markedly obese did not reach statistical significance. More and larger laparoscopic ports were used in the markedly obese. Obese donors were more likely to require conversion from laparoscopic nephrectomy to open nephrectomy than ideal-sized donors. The postoperative recovery of the gastrointestinal tract, hospitalization time, analgesic requirements, and total complications were equal in the two groups, although the obese donors' complications tended to be cardiopulmonary problems. The recipient graft function was equivalent between the two groups. CONCLUSIONS: Laparoscopic living donor nephrectomy is more difficult to perform in the markedly obese but is associated with an equivalent donor morbidity and recipient renal outcome. PMID- 11113735 TI - Limitations of computed tomography in the preoperative staging of upper tract urothelial carcinoma. AB - OBJECTIVES: Computed tomography (CT) of the abdomen and pelvis has been used for staging of upper tract urothelial carcinoma. This study was initiated to evaluate the utility of this modality in guiding the management of patients with upper tract urothelial malignancies. METHODS: We performed a retrospective chart review of 37 consecutive patients with urothelial carcinoma of the upper urinary tract (21 renal pelvic, 16 ureteral) who underwent preoperative CT staging of the abdomen and pelvis before open surgical management. RESULTS: CT was not required to establish the diagnosis in any of the patients, but in 16.2% helped to confirm the diagnosis when the disease was suspected based on other studies. CT accurately provided evidence of metastatic disease in only 3% of the patients. CT was accurate in predicting pathologic TNM stage in 59. 5% of patients. The study understaged or did not detect in 16.2% and 24.3%, respectively. Most importantly, CT did not alter the management of any patient. CONCLUSIONS: CT was rarely helpful in establishing the diagnosis of the upper tract urothelial carcinoma and did not influence the management of any patient. We conclude that preoperative CT scan in those patients who are to undergo open surgical management of confirmed urothelial malignancies of the upper urinary tract without suspicion of advanced disease will rarely influence the management of the disease and its use should be selective and not routine. PMID- 11113736 TI - Nitinol stone retrieval-assisted ureteroscopic management of lower pole renal calculi. AB - OBJECTIVES: Current ureteroscopic intracorporeal lithotripsy devices and stone retrieval technology allow for the treatment of calculi located throughout the intrarenal collecting system. Difficulty accessing lower pole calculi, especially when the holmium laser fiber is used, is often encountered. We retrospectively reviewed our experience with cases in which lower pole renal calculi were ureteroscopically managed by holmium laser fragmentation, either in situ or by first displacing the stone into a less dependent position with the aid of a nitinol stone retrieval device. METHODS: Thirty-four patients (36 renal units) underwent ureteroscopic treatment of lower pole renal calculi between April 1998 and November 1999. Lower pole stones less than 20 mm were primarily treated by ureteroscopic means in patients who were obese, in patients who had a bleeding diathesis, in patients with stones resistant to shock wave lithotripsy, and in patients with complicated intrarenal anatomy, or as a salvage procedure after failed shock wave lithotripsy. Lower pole calculi were fragmented with a 200 micrometer holmium laser fiber by way of a 7.5F flexible ureteroscope. For those patients in whom the laser fiber reduced the ureteroscopic deflection, precluding re-entry into the lower pole calix, a 3.2F nitinol basket or a 2.6F nitinol grasper was used to displace the lower pole calculus into a more favorable position, allowing easier fragmentation. RESULTS: In 26 renal units, routine in situ holmium laser fragmentation was successfully performed. In the remaining 10 renal units, a nitinol device was passed into the lower pole, through the ureteroscope, for stone displacement. Only a minimal loss of deflection was seen. Irrigation was significantly reduced by the 3.2F nitinol basket, but improved with the use of the 2.6F nitinol grasper. This factor did not impede stone retrieval in any of the patients. At 3 months, 85% of patients were stone free by intravenous urography or computed tomography. CONCLUSIONS: Ureteroscopic management of lower pole calculi is a reasonable alternative to shock wave lithotripsy or percutaneous nephrolithotomy in patients with low-volume stone disease. If the stone cannot be fragmented in situ, nitinol basket or grasper retrieval, through a fully deflected ureteroscope, allows one to reposition the stone into a less dependent position, thus facilitating stone fragmentation. PMID- 11113737 TI - Treatments used in women with interstitial cystitis: the interstitial cystitis data base (ICDB) study experience. The Interstitial Cystitis Data Base Study Group. AB - OBJECTIVES: To evaluate the frequency and types of treatments reported at baseline in women who entered the Interstitial Cystitis Data Base (ICDB) cohort study. METHODS: From 1993 to 1997, 581 women were enrolled and followed in the ICDB. All treatments reported at study entry, including those prescribed for interstitial cystitis (IC) and concomitant medications, were reviewed. The number and types of treatments were evaluated with respect to baseline factors such as prior diagnosis of IC and symptom severity. RESULTS: One hundred five (18%) women were receiving no therapy at baseline. Single-mode therapy was reported by 195 (34%) women, and a combination of two treatments was reported by 119 (21%) women. Three or more treatments were reported in 162 (28%) women. A total of 183 different types of therapies were recorded. The five most commonly used therapies for IC symptoms were cystoscopy and hydrodistention, amitriptyline, phenazopyridine, special diet, and intravesical heparin. Because most patients entered the ICDB before the approval of oral pentosan polysulfate sodium (PPS), only 6% of women reported oral PPS use at baseline. There were statistically significant associations between the number and types of treatments and clinical center, a prior diagnosis of IC, and symptom severity. CONCLUSIONS: The diversity of IC therapies underscores the lack of understanding about the treatment of this syndrome. Further research in IC is essential to develop and to evaluate rational therapies and treatment algorithms. These algorithms should be "evidence based" and should be revised as the underlying etiology and pathophysiology of IC is delineated. PMID- 11113738 TI - Electromyographic study of the striated urethral sphincter in type 3 stress incontinence: evidence of myogenic-dominant damages. AB - OBJECTIVES: To determine the electromyographic features of the striated urethral sphincter in patients with type 3 stress incontinence (SI) due to intrinsic sphincteric deficiency (ISD). METHODS: We performed electromyography (EMG) of the striated urethral sphincter muscle and urodynamic studies in a total of 51 women, 41 female patients with type 3 SI and 10 women with normal urinary control (NUC). The results were analyzed in both groups, and the correlation of EMG findings with the patient characteristics and urodynamic parameters was evaluated. RESULTS: Motor unit potentials (MUP) of SI patients showed significantly shorter duration (P = 0.0014), lower amplitude (P = 0.0008), and larger number of phases (P = 0.0022) compared with those in the NUC group. Thirty (73%) of the SI patients showed an obvious low amplitude (less than 350 microV)/short duration (less than 4.5 milliseconds)/polyphasic pattern and early recruitment of interference activity with low amplitude at voluntary contraction of the striated sphincter, both indicating existence of myogenic damages. These patients showing myogenic damages had significantly lower Valsalva leak point pressure (P = 0.002) and more leakage in the pad-weigh test (P = 0.010) compared with the SI patients without myogenic damage findings. CONCLUSIONS: These results suggested that myogenic-dominant damages of the striated urethral sphincter may contribute to the etiology of ISD in most patients with type 3 SI. PMID- 11113739 TI - Biofeedback, pelvic floor re-education, and bladder training for male chronic pelvic pain syndrome. AB - OBJECTIVES: Pelvic floor tension myalgia may contribute to the symptoms of male patients with chronic pelvic pain syndrome (CPPS). Therefore, measures that diminish pelvic floor muscle spasm may improve these symptoms. Based on this hypothesis, we enrolled 19 patients with CPPS in a 12-week program of biofeedback directed pelvic floor re-education and bladder training. METHODS: Pre-treatment and post-treatment symptom assessments included daily voiding logs, American Urological Association (AUA) symptom score, and 10-point visual analog pain and urgency scores. Pressure-flow studies were obtained before treatment in most patients. Instruction in pelvic floor muscle contraction and relaxation was achieved using a noninvasive form of biofeedback at biweekly sessions. Home exercises were combined with a progressive increase in timed-voiding intervals. RESULTS: Mean age of the 19 patients was 36 years (range 18 to 67). Four patients completed less than three treatment sessions, 5 patients completed three to five sessions, and 10 attended all six sessions. Mean follow-up was 5.8 months. Median AUA symptom scores improved from 15.0 to 7.5 (P = 0.001), and median bother scores decreased from 5.0 to 2.0 (P = 0.001). Median pain scores decreased from 5.0 to 1.0 (P = 0.001), and median urgency scores decreased from 5.0 to 2.0 (P = 0.002). Median voiding interval increased from 0.88 hours to 3.0 hours (P = 0.003). Presence of detrusor instability, hypersensitivity to filling, or bladder sphincter pseudodyssynergia on pretreatment urodynamic studies was not predictive of treatment results. CONCLUSIONS: This preliminary study confirms that a formalized program of neuromuscular re-education of the pelvic floor muscles together with interval bladder training can provide significant and durable improvement in objective measures of pain, urgency, and frequency in patients with CPPS. PMID- 11113740 TI - Transvaginal bone-anchored synthetic sling for the treatment of stress urinary incontinence: an outcomes analysis. AB - OBJECTIVES: To evaluate the results and complications of a new transvaginal minimally invasive procedure for the treatment of stress urinary incontinence. METHODS: Sixty-seven women aged 37 to 77 years underwent a pervaginal bone anchoring synthetic sling procedure between April 1997 and February 1999. Sixty three patients had a defect of the anatomic support, and 4 had iatrogenic intrinsic sphincteric deficiency (ISD). Patients were assessed at least 1 year postoperatively, underwent physical examination, and filled in the self assessment questionnaire with the help of a nonpartisan health care provider. The questionnaire inquired about urine leakage, obstructive and irritative symptoms, quality of life, and satisfaction with the treatment received. The physician and questioner had no knowledge of each other. RESULTS: Mean follow-up was 17 months. Perfect dryness was seen in 82% of patients with improvement in 9% and failure in 9%. Patients reporting a failed outcome were significantly older than those reporting improvement or cure (P = 0.01). All patients with ISD failed. Moderate obstructive symptom scores have been noticed in 31% of patients. Irritative voiding symptoms have been recorded in 22% of cured patients and 83% and 80% of improved and failed patients, respectively (P <0.001). Three percent of patients experienced pain during intercourse. Mild pelvic pain was found in 8% of patients. The only main complication of the operation was the vaginal erosion and sling removal in 16% of patients. Seventy-two percent of patients were completely satisfied with the treatment received. CONCLUSIONS: This procedure allows a high cure rate in patients with urinary incontinence due to a defect of anatomic support, and it is unsuitable when incontinence is due to ISD. The only main complication came from the use of the gelatin-coated Dacron sling that resulted in vaginal erosion, often necessitating the sling removal. The use of different synthetic or nonsynthetic materials may be advisable. PMID- 11113741 TI - Reconstructive surgery in voiding dysfunction: experience with 69 patients. AB - OBJECTIVES: To present our reconstructive surgery experience with voiding dysfunction due to both neurologic and non-neurologic etiology. METHODS: From March 1993 to January 2000, 69 patients (43 men and 26 women) with voiding dysfunction underwent lower urinary tract reconstruction. Mean patient age at the time of surgery was 34. 5 years (range 9 to 75). Voiding dysfunction had a neurologic etiology in 65.2% of the patients and a non-neurologic etiology in 34.8%. Urodynamic investigation revealed poor bladder compliance in 52%, detrusor hyperreflexia in 19%, and a combination of the two in 29% of the patients. Thirteen patients (19%) had coexistent intrinsic sphincteric deficiency. A total of 56.5% of the patients had upper urinary tract deterioration. Most patients (78%) had severe urinary incontinence. Augmentation cystoplasty was performed in 60 patients. Nine patients had augmentation cystoplasty with a continent stoma. Concomitant procedures were performed in 11 patients. RESULTS: Mean follow-up was 36.6 months (range 8 to 108). Marked improvement of the upper tracts was documented in 79% of the patients in the neuropathic and 73% in the non neuropathic group. High continence rates were achieved in both groups (82% and 94%, respectively). Intermittent catheterization rate was 88.6% in the neuropathic and 44% in the non-neuropathic groups and patient satisfaction rate was 84% and 94%, respectively. Three major complications in 2 patients required surgery. CONCLUSIONS: Surgical reconstruction to treat urinary incontinence and upper urinary tract deterioration gives satisfactory results in voiding dysfunction in the case of medical treatment failure. PMID- 11113742 TI - Reliability of color Doppler ultrasound urodynamics in the evaluation of bladder outlet obstruction. AB - OBJECTIVES: To analyze the intrarater and interrater reliability of a newly developed noninvasive urodynamic technique based on color Doppler ultrasound for the evaluation of bladder outlet obstruction. We previously demonstrated the feasibility of this new technique. METHODS: Color Doppler ultrasound urodynamic evaluations were performed on 31 men with and without bladder outlet obstruction. An ultrasound image-directed Doppler system with a 3.75-MHz probe operated by a remote control robotic manipulator was used to obtain color scale data using the transperineal approach in men during voiding. We measured the flow velocities in the distal prostatic (V1) and membranous urethra (V2) and used them to obtain the velocity ratio (VR = V1/V2). Combining this information with simultaneous uroflowmetry, the functional cross-sectional area of the distal prostatic urethra (A1) was calculated. These parameters were independently reinterpreted by an inexperienced investigator using the color image data stored in a personal computer. The intrarater and interrater reliability for VR were assessed using a classification based on cutoff values previously shown to indicate the presence or absence of obstruction. RESULTS: The retest correlation using Spearman's rho for VR in terms of intrarater and interrater reliability was 0.95 and 0.57, respectively; that for A1 was 0.97 and 0.64, respectively. Using a VR of less than 1.1 (to indicate the absence of obstruction) and a VR greater than 1.6 (to indicate the presence of obstruction) for classification, intrarater and interrater agreement occurred in 93.6% to 96.8% and 77.4% to 83. 9% of cases, respectively. CONCLUSIONS: The color Doppler ultrasound urodynamic technique for noninvasive evaluation of bladder outlet obstruction can be performed with reasonable reliability. PMID- 11113744 TI - Cost effectiveness of microwave thermotherapy in patients with benign prostatic hyperplasia: part II--results. AB - OBJECTIVES: To evaluate the cost effectiveness of transurethral microwave thermotherapy relative to medical therapy (alpha-blocking agents) and transurethral resection of the prostate (TURP) for patients with moderate-to severe benign prostatic hyperplasia (BPH) symptoms. METHODS: A cost-effectiveness analysis was performed from the societal perspective for a hypothetical cohort of 65-year-old men with moderate-to-severe BPH symptoms. We calculated the incremental cost effectiveness of thermotherapy relative to medical therapy and TURP during 5 years after treatment initiation. Event probabilities were obtained from published reports, a consensus panel, and the Targis System (Urologix) randomized clinical trial. Costs were estimated using the national Medicare reimbursement schedules. Costs are reported in 1999 U.S. dollars. Total thermotherapy procedure costs were estimated at $2629. Quality-of-life and utility estimates were obtained by interviewing 13 patients with moderate-to severe BPH symptoms. On the basis of their risk attitudes, patients were classified into risk-averse or non-risk-averse groups. The costs and health effects were discounted at 3% annually. RESULTS: In a hypothetical cohort of 10,000 non-risk-averse patients who were candidates for all three modalities, the 5-year costs were highest for patients undergoing TURP and lowest for those receiving medical therapy ($7334 and $6294, respectively). The thermotherapy group exhibited the highest 5-year utility value (53.52 quality-adjusted life months). Compared with medical therapy, thermotherapy resulted in an additional 0.23 quality-adjusted life-months, with an incremental cost of $741. This yielded an incremental cost per quality-adjusted life-year gained of $38,664 for thermotherapy compared with medical therapy. Thermotherapy had a higher utility (difference of 1.71 quality-adjusted life-months) and lower cost (difference of $299) compared with TURP and thus was dominant over TURP. The results were similar for a hypothetical cohort of 10,000 risk-averse patients. CONCLUSIONS: From a societal perspective, thermotherapy appears to be a reasonable and cost effective alternative to both medical and surgical treatment. However, the actual treatment decision should be based on multiple factors, only one of which is cost effectiveness. PMID- 11113743 TI - Cost effectiveness of microwave thermotherapy in patients with benign prostatic hyperplasia: part I-methods. AB - OBJECTIVES: To present the method used to evaluate the cost effectiveness, from the societal perspective, of transurethral microwave thermotherapy relative to medical therapy (alpha-blocking agents) and transurethral resection of the prostate (TURP) for a hypothetical cohort of 65-year-old men with moderate-to severe benign prostatic hyperplasia (BPH) symptoms. METHODS: We constructed a decision-analytic Markov model with 25 health states describing the 3 treatments, 5 short-term clinical events, and 17 possible long-term outcomes. Each health state had an associated cost and utility. Utility weights, reflecting an individual's preference for a specific health outcome, range from 0, indicating death, to 100, indicating perfect health. Utility estimates were obtained by interviewing 13 men with moderate-to-severe BPH symptoms using the standard gamble preference measurement technique. On the basis of their risk attitudes, the patients were classified as risk averse or non-risk averse. The rates of remission, temporary and permanent adverse events, retreatment, and mortality were obtained from the Targis System (Urologix) randomized clinical trial, published reports, and a consensus panel. The costs during the 5 years after treatment initiation were estimated using national Medicare reimbursement schedules. The costs are reported in 1999 U.S. dollars. RESULTS: Eliciting utility values from patients with BPH was feasible and generated internally consistent and externally valid measures. In the non-risk-averse group, the utility value for significant remission, moderate remission, no remission, and worsening BPH symptoms without an adverse event was 99.1, 97.1, 94.4, and 87.3, respectively. As expected, the risk-averse individuals (n = 6) exhibited higher utility values than those in the non-risk-averse group (n = 7). In the non-risk averse group, thermotherapy was the preferred treatment, and in the risk-averse group, medical therapy was preferred. In both groups, TURP was the least preferred therapy. The initial thermotherapy procedure costs without complications were estimated at $2629, and the initial TURP procedure costs without complications were estimated at $4597. Time-dependent probabilities were developed to reflect treatment durability. CONCLUSIONS: The resulting model parameters appear to be suitable for evaluating the cost effectiveness of thermotherapy relative to medical therapy and TURP in 65-year-old men with moderate-to-severe BPH symptoms. PMID- 11113745 TI - Multicenter ProstaScint imaging findings in 2154 patients with prostate cancer. The ProstaScint Imaging Centers. AB - OBJECTIVES: To report the results of a retrospective study of 2290 ProstaScint scans of 2154 patients with prostate carcinoma done at 15 institutions. METHODS: The results were analyzed by logistic regression after stratification of the patients into four groups: group 1, newly diagnosed; group 2, after radical prostatectomy with a rising prostate-specific antigen (PSA) level; group 3, after radiation therapy with a rising PSA level; and group 4, after hormonal therapy. RESULTS: The PSA level and ProstaScint scans positive in the prostate bed (P <0.001) and for pelvic metastases (P <0.001), but not for extrapelvic metastases, correlated significantly in group 1 patients. In group 2, the association for detecting fossa recurrence was weaker (P = 0.033) and was insignificant for pelvic and extrapelvic metastases. Patients in group 3 also exhibited a weak PSA ProstaScint association for detecting fossa recurrence (P = 0.038), and was insignificant for pelvic and extrapelvic metastases. No significant PSA ProstaScint correlation was found in patients in group 4 for fossa recurrence, pelvic or extrapelvic metastases. The distribution of positive ProstaScint results among the prostate/prostate bed, pelvic nodes, and extrapelvic nodes was nearly equal for all groups, except that a significantly greater percentage of extrapelvic metastases was found in the hormonal group (group 4). The ProstaScint results were independent of the Gleason score for 260 patients before and 285 patients after therapy. CONCLUSIONS: The results of this study underscore the complementary diagnostic value of ProstaScint to PSA level and Gleason score as an independent indicator of prostate cancer recurrence and metastases and in identifying extrapelvic metastases in both newly diagnosed and recurrent prostate cancer. PMID- 11113746 TI - A neural network predicts progression for men with gleason score 3+4 versus 4+3 tumors after radical prostatectomy. AB - OBJECTIVES: To determine the significance of Gleason scores 3+4 (GS3+4) versus 4+3 (GS4+3) with respect to biochemical recurrence in a retrospective review of a series of men with clinically localized prostate cancer who underwent radical retropubic prostatectomy (RRP) and to develop and test an artificial neural network (ANN) to predict the biochemical recurrence after surgery for this group of men using the pathologic and clinical data. METHODS: From 1982 to 1998, 600 men had pathologic Gleason score 7 disease without lymph node or seminal vesicle involvement. We analyzed the freedom from biochemical (prostate-specific antigen) progression after RRP on 564 of these men on the basis of their GS3+4 versus GS4+3 (Gleason 7) status. The Cox proportional hazards model was used to determine the importance of Gleason 7 status as an independent predictor of progression. In addition, an ANN was developed using randomly selected training and validation sets for predicting biochemical recurrence at 3 or 5 years. Different input variable subsets, with or without Gleason 7 status, were compared for the ability of the ANN to maximize the prediction of progression. Standard logistic regression was used concurrently on the same random patient population sets to calculate progression risk. RESULTS: A significant recurrence-free survival advantage was found in men who underwent RRP for GS3+4 compared with those with GS4+3 disease (P <0.0001). The ANN, logistic regression, and proportion hazard models demonstrated the importance of Gleason 7 status in predicting patient outcome. The ANN was better than logistic regression in predicting patient outcome, in terms of prostate-specific antigen progression, at 3 and 5 years. CONCLUSIONS: A simple modification of the Gleason scoring system for men with Gleason 7 disease revealed a difference in the patient outcome after RRP. ANN models can be developed and used to better predict patient outcome when pathologic and clinical features are known. PMID- 11113747 TI - Performance of a neural network in detecting prostate cancer in the prostate specific antigen reflex range of 2.5 to 4.0 ng/mL. AB - OBJECTIVES: To explore the potential role of a neural network-derived algorithm in enhancing the specificity of prostate cancer detection compared with the determination of prostate-specific antigen (PSA) and free PSA (fPSA) while maintaining a 90% detection rate. Recent information suggests that the incidence of detectable prostate cancer is similar in men whose PSA values range from 2.5 to 4.0 ng/mL and from 4.0 to 10.0 ng/mL. If the PSA threshold triggering a prostate biopsy is lowered to 2.5 ng/mL, approximately 13% of men older than 50 would be added to the patient biopsy pool. METHODS: One hundred fifty-one men were enrolled in a prospective, Institutional Review Board-approved protocol to evaluate the incidence of cancer in a population of men who participated in an early-detection program and whose PSA level was between 2.5 and 4.0 ng/mL. All the men underwent biopsy using an 11-core multisite-directed biopsy scheme, and all biopsy specimens were examined by one pathologist. All men had a second blood specimen drawn before the biopsy for a determination of serum PSA, creatinine kinase, prostatic acid phosphatase, and fPSA. A new neural network algorithm was developed with PSA, creatinine kinase, prostatic acid phosphatase, fPSA, and age as input variables to produce a single-valued prostate cancer detection index (PCD-I). This new algorithm was then prospectively tested in the 151 men. Performance parameters (including sensitivity, specificity, positive and negative predictive values, and biopsies saved) were calculated, and a comparative analysis was performed to evaluate the differences among the new algorithm, percent fPSA, PSA density, and PSA density-transition zone. RESULTS: Cancer was histologically confirmed in 24.5% (37 of 151) of the men. The median age of the men was 62 years (range 43 to 74). At a sensitivity of 92%, the specificity for percent fPSA was 11%. The new algorithm (PCD-I) demonstrated an additional enhancement of specificity to 62% at 92% sensitivity. Clinically, the PCD-I would result in a savings of 49% (74 of 151) of all biopsies or 63.6% (71 of 114) of all unnecessary biopsies. CONCLUSIONS: A new generation algorithm, derived from a neural network (PCD-I) incorporating the parameters of age, creatinine kinase, PSA, prostatic acid phosphatase, and fPSA can significantly enhance the specificity and reduce the number of biopsies while maintaining a 92% sensitivity rate. PMID- 11113748 TI - Ethnic differences in the age-related distribution of serum prostate-specific antigen values: a study in a healthy Korean male population. AB - OBJECTIVES: To further improve the use of prostate-specific antigen (PSA) as a screening test for prostate cancer in Asian countries, we sought to establish the normal distribution of serum PSA values in Korean men, because, until recently, studies conducted to establish normal serum PSA values have involved few Asian populations. METHODS: Between May 1995 and June 1997, 5805 healthy Korean men 30 to 79 years old who visited our hospital for a routine health checkup were entered into a prospective study of early screening for prostate cancer. All men underwent detailed clinical examinations, including a digital rectal examination and serum PSA determination. All men who were more than 50 years old with abnormal digital rectal examination findings and/or an elevated serum PSA level (greater than 4.0 ng/mL) also underwent transrectal ultrasound-guided sextant biopsy. Four were found to have cancer and were excluded from the analysis. RESULTS: The median serum PSA concentration (5th to 95th percentile range) was 0.8 ng/mL (0.2 to 1.8) for patients 30 to 39 years old (n = 1382); 0.8 ng/mL (0.2 to 2.0) for patients 40 to 49 years old (n = 1776); 0.9 ng/mL (0.2 to 2.4) for those 50 to 59 years old (n = 1775); 1.0 ng/mL (0.2 to 3.9) for men 60 to 69 years old (n = 746); and 1.3 ng/mL (0.5 to 6.3) for patients 70 to 79 years old (n = 122). The serum PSA concentration correlated with age (P <0.001), with an increase by approximately 1.2% annually, although the statistical correlation was weak (r = 0.16). Almost no change occurred in the median serum PSA value in patients 50 years old or younger; a gradual increase was observed in patients older than 50. In those 50 years old or older, the median and 95th percentile serum PSA values for Korean men were lower than those for white men. CONCLUSIONS: Contrary to earlier observations that the serum PSA level strongly correlates with age, the influence of age on serum PSA was found to be weaker in this study. Moreover, the results also demonstrated that the distribution of the serum PSA level differs along ethnic lines. The cutoff value for serum PSA in mass screening for prostate cancer should be adjusted in nonwhite races. PMID- 11113749 TI - Neuroendocrine differentiation is not prognostic of failure after radical prostatectomy but correlates with tumor volume. AB - OBJECTIVES: To study neuroendocrine (NE) tumor cell differentiation in prostate cancer in relation to failure after radical prostatectomy. METHODS: Radical prostatectomy specimens from 103 of 111 patients randomized to 3-month neoadjuvant luteinizing hormone-releasing hormone-analogue treatment (neoadjuvant group) or to surgery alone (control group) were available for analysis. Immunohistochemistry using antibodies to chromogranin A (CGA) enabled detection of tumor cells with NE differentiation. NE differentiation was scored as NE negative (0 to 1+) or NE-positive (2 to 3+). The number of CGA-positive cells/cm(2) tumor area on the slides was assessed in a separate analysis. The patients were followed up for 39 months after surgery, and a prostate-specific antigen value of 0.5 ng/mL or greater in two consecutive blood samples was considered biochemical failure. RESULTS: Kaplan-Meier analysis stratified for neoadjuvant hormonal treatment showed the failure rate to be significantly greater among those with NE-positive tumors than among those with NE-negative tumors. However, the number of CGA-positive cells/cm(2) was not a variable of prognostic significance. Instead, both NE differentiation and the CGA-positive cell count correlated with the tumor area on the slides (P = 0.0001). Multivariate analysis revealed the tumor area on the slide (P <0.0001) and positive surgical margins (P = 0.03) to be the only significant predictors of biochemical failure. CONCLUSIONS: The extension of NE differentiation in prostate cancer correlates with tumor volume and is not an independent prognostic factor of failure after radical prostatectomy. PMID- 11113750 TI - Impact of socioeconomic status and race on clinical parameters of patients undergoing radical prostatectomy in an equal access health care system. AB - OBJECTIVES: To analyze the relationships among socioeconomic status (SES), race, and the clinical parameters of patients undergoing radical prostatectomy (RP) in an equal access health care system. METHODS: The Department of Defense Center for Prostate Disease Research longitudinal prostate cancer database from multiple military institutions was used to analyze the clinical, pathologic, and outcome data of 1058 patients with localized (Stage T2c or lower) prostate cancer and a preoperative prostate-specific antigen (PSA) level of 20 ng/mL or less who underwent RP between January 1987 and December 1997. Military rank (officer versus enlisted) was used as a surrogate measure of SES. RESULTS: The percentage of patients with pathologic Gleason grade 7 or greater prostate cancer was higher in enlisted (45%) than in officer (37%) patients (P = 0. 021). However, no difference was found between these groups with respect to pathologic stage or biochemical recurrence rates. African Americans presented at a younger age (P = 0.003), with a higher pretreatment PSA level (P = 0.001), and demonstrated higher biochemical recurrence rates than other ethnic groups (P = 0.037). The Cox proportional hazards analysis showed that a lower SES (P = 0.010) but not African American race (P = 0.696) was an independent predictor of a higher grade (Gleason grade 7 or higher) cancer. However, biochemical progression was more common in African American men (P = 0.035) and was not related to SES (P = 0.883). CONCLUSIONS: In an equal access health care system, patients of lower SES presented with higher grade prostate cancer at the time of RP. However, only African American race predicted biochemical progression after RP. PMID- 11113751 TI - Reassessment of the definition of castrate levels of testosterone: implications for clinical decision making. AB - OBJECTIVES: Based on methods introduced in the late 1960s and no longer used, serum testosterone level in men after surgical castration was reported to be 50 ng/dL or less. Radioimmunoassay and, subsequently, chemiluminescent methods have supplanted the early analytic methods because of their improved accuracy and ease of testing. The purpose of this study was to define the castrate testosterone level in the era of chemiluminescent testing. METHODS: After bilateral orchiectomy, serum testosterone (total) levels were measured prospectively in 35 prostate cancer patients. RESULTS: The median testosterone value in this patient cohort was 15 ng/dL (0.5 nmol/L; 95% confidence interval 12 to 17 ng/dL). CONCLUSIONS: In a contemporary series, castrate testosterone should be defined as less than 20 ng/dL (0.7 nmol/L). The important biologic and economic implications are discussed. PMID- 11113752 TI - Evaluation of the cytokines interleukin 8 and epithelial neutrophil activating peptide 78 as indicators of inflammation in prostatic secretions. AB - OBJECTIVES: Chronic prostatitis/chronic pelvic pain syndrome (CPPS) is a disorder characterized by pelvic pain and varying degrees of inflammation exhibited in expressed prostatic secretions (EPS). To provide objective parameters of inflammation, we measured the cytokines interleukin 8 (IL-8) and epithelial neutrophil activating peptide 78 (ENA-78) in EPS of healthy men, men with benign prostatic hyperplasia (BPH), men with bacterial prostatitis (BP), and men with chronic prostatitis/CPPS. METHODS: Enzyme-linked immunosorbent assays of the EPS for IL-8 and ENA-78 were done in 63 men: control (n = 9), BPH (n = 6), BP (n = 3), inflammatory CPPS (National Institutes of Health [NIH] category IIIa) (n = 17), noninflammatory CPPS (NIH category IIIb) (n = 17), and asymptomatic inflammatory prostatitis (NIH category IV) (n = 11). RESULTS: IL-8 was detectable in all patients, and ENA-78 was detectable in all except 2 patients (threshold of detection 10 pg/mL for IL-8, 15 pg/mL for ENA-78). Mean levels of IL-8 [ENA-78] were similar in control (3010 pg/mL [423 pg/mL]), BPH (3341 pg/mL [98 pg/mL]), and IIIb (2751 pg/mL [335 pg/mL]) groups. Both cytokine levels were higher in BP (11,175 pg/mL [13,761 pg/mL]), IIIa (10,418 pg/mL [2240 pg/mL]), and IV (8571 pg/mL [1865 pg/mL]) groups. A statistically significant difference between the control group versus BP, IIIa, and IV (P <0.05) groups was found for IL-8 but not for ENA-78. CONCLUSIONS: IL-8 and ENA-78 are frequently elevated in the EPS of men with BP, CPPS IIIa, and asymptomatic inflammatory prostatitis category IV. These cytokines are direct mediators of leukocyte accumulation and activation at inflammatory sites and may be responsible, in part, for the presence of inflammatory reaction in the prostate. PMID- 11113754 TI - Editorial comment PMID- 11113753 TI - Current sexual functioning in 106 patients with Peyronie's disease treated with radiotherapy 9 years earlier. AB - OBJECTIVES: To analyze retrospectively the sexual functioning and treatment outcome in patients after radiotherapy (RT) for Peyronie's disease. METHODS: During 1982 to 1997, 179 patients with Peyronie's disease were treated at our radiation oncology department. One hundred thirty patients whose address could be traced were sent a questionnaire about their symptoms before RT, treatment outcome, and current sexual functioning (the past 4 weeks). One hundred six patients, mean age 59 years, responded. RESULTS: Before RT, 44% reported painful erections, 97% penile curvature, and 18% erectile dysfunction. Sixty-nine percent reported that after RT, penile pain was diminished and 29% that penile curvature was decreased; 13% reported improved erections. With respect to their current sex life, 51% reported sometimes or always having problems getting an erection and 61% in maintaining an erection; 36% never experienced spontaneous erections. Fifty-four percent reported not having erections rigid enough for sexual activity. Since RT (mean 9 years), there had been a moderate or severe decrease in sexual interest (17%), sexual activity (41%), and sexual pleasure (32%). Overall, 49% of patients were dissatisfied with their current sexual functioning. CONCLUSIONS: Low-dose external RT resulted in relief of pain in two thirds of patients with Peyronie's disease. An improvement in penile curvature was reported in one third of patients. One half of men reported problems in getting an erection. The rigidity of erections was satisfactory in only 54%. There was a moderate to severe decrease in sexual interest, activity, and pleasure after RT; however, this was associated with age, although comorbidity or medications cannot be excluded. PMID- 11113755 TI - Reply by the authors PMID- 11113756 TI - Normative values for female genital sensation. AB - OBJECTIVES: Neurologic disorders might be responsible for many cases of female sexual dysfunction. Yet, they are currently undiagnosed because of the lack of measurement tools to assess genital neural function. Therefore, our objective is to provide norms for sensory thresholds in the vagina and clitoris, for a wide range of patient ages. METHODS: Vaginal and clitoral warm, cold, and vibratory sensory thresholds were measured in 89 healthy paid volunteers by the method of limits. Normograms were derived from this group of healthy volunteers. An additional 61 patients were also tested, for a total of 150 individuals. Sixty two individuals (42 healthy volunteers and 20 patients) from the total group were tested twice to provide test-to-test repeatability data across the range of clinical (normal and abnormal) responses. RESULTS: Normograms are presented, providing age-corrected upper and lower normal values, expressed as 95% confidence limits for warm, cold, and vibratory thresholds. Intertest repeatability is also reported. CONCLUSIONS: Thermal and vibratory thresholds of both the vaginal and clitoral region are clinically feasible, valid, and repeatable. These can be applied as a valuable diagnostic tool to assess neural dysfunction through sensory assessment of the female genitalia. PMID- 11113758 TI - Abdominoscrotal hydrocele in childhood: is it really a rare entity? AB - OBJECTIVES: Abdominoscrotal hydrocele (ASH) is an apparently highly uncommon clinical entity especially in childhood, with only about 80 pediatric cases reported in the modern literature. METHODS: The incidence, diagnosis, and treatment of ASH are discussed with reference to 9 cases observed at our institution and to cases in the literature. RESULTS: Surgical correction was successful in all our cases and no hydrocele or hernia recurrence was registered. CONCLUSIONS: ASH incidence appears to be higher than reported; undescended testis is a frequent association and surgical correction through an inguinal approach is always possible and curative. Ligation of the processus vaginalis may prevent recurrence. PMID- 11113757 TI - Y-chromosome microdeletion and its effect on reproductive decisions in taiwanese patients presenting with nonobstructive azoospermia. AB - OBJECTIVES: To investigate the position, extent, and frequency of Y chromosome microdeletions in Taiwanese patients presenting with nonobstructive azoospermia, and to investigate the effect of microdeletions on reproductive decisions. METHODS: We studied 176 consecutive men with azoospermia in our urology clinic. Polymerase chain reaction tests were performed in 94 patients with nonobstructive azoospermia, and a series of 27 sequence-tagged sites (STSs) mapped within intervals 5 and 6 of Yq11 was selected for analysis. Clinical genetics counseling was provided to couples with microdeletions, and these couples made their own choices about further treatment modalities. RESULTS: Among 94 patients screened for microdeletion, 11 (11.7%) showed microdeletions of one or more STSs. One had a deletion confined to the azoospermia factor b (AZFb) region (encompassing the RBM gene). Two were found to have deletions of both the AZFb and AZFc regions. Eight patients had deletions in the AZFc region (encompassing the DAZ gene). Five had deletions distal to the DAZ gene family. One had multiple, noncontiguous deletions. In 8 patients with testicular histology available, a lack of genotype/phenotype correlation was noted. Of the 11 couples with deletions, 3 thought microdeletion was a serious defect and opted for an artificial insemination of donor or adoption, 5 chose intracytoplasmic sperm injection, and the other 3 decided to undergo treatment with Chinese medicinal herbs. CONCLUSIONS: The most commonly deleted region in the Taiwanese population is AZFc. The genes implicated in Taiwanese spermatogenesis defects are the DAZ and RBM gene families. Twenty-seven percent of couples with microdeletions deferred assisted reproductive technologies because of concern about their underlying genetic defects. PMID- 11113759 TI - The thimble: a useful adjunct to needle suspension procedures for female stress incontinence. AB - All needle suspension procedures carry the risk of inadvertent puncture of the surgeon's finger. The thimble provides simple, inexpensive protection for the surgeon while maintaining tactile feedback during surgery. PMID- 11113760 TI - Prospective multicenter study of transperineal prostatic block for transurethral needle ablation of the prostate. AB - The choice of anesthesia during thermal therapy of the prostate plays a significant role in the morbidity profile, patient convenience, and cost. We report 39 men with symptomatic benign prostatic hyperplasia who underwent transurethral needle ablation of the prostate under transperineal prostatic block. This method of local anesthesia proved safe, convenient, and satisfactory during the procedure. PMID- 11113761 TI - Percutaneous management of stones in a patient with sacral agenesis. AB - We report a case of a patient with sacral agenesis and nephrolithiasis in whom percutaneous nephrostolithotomy was used to treat the stone disease. Sacral agenesis is an uncommon congenital anomaly involving the lower vertebral bodies and is associated with urinary tract dysfunction. Nephrolithiasis in a patient with sacral agenesis poses a problem in access for percutaneous nephrostolithotomy because of the associated presence of renal ectopia. We describe our technique and the special considerations necessary for successful access. PMID- 11113762 TI - Lesch-Nyhan syndrome presenting as acute renal failure secondary to obstructive uropathy. AB - Lesch-Nyhan syndrome is a rare genetic disorder characterized by mental retardation, self-mutilation, choreoathetosis, and hyperuricemia. The disease is caused by a mutation in the hypoxanthine-guanine phosphoribosyltransferase gene and is transmitted as a sex-linked recessive disorder. Since hyperuricemia is the primary metabolic problem caused by a hypoxanthine-guanine phosphoribosyltransferase mutation, urologic evaluation and treatment is often necessary for children with this disease. We report a 3-year-old boy who presented with anuric renal failure secondary to bilateral obstructing uric acid calculi. The evaluation of T lymphocytes revealed a hypoxanthine-guanine phosphoribosyltransferase mutation consistent with Lesch-Nyhan syndrome. The diagnosis and urologic management of this disorder is discussed. PMID- 11113763 TI - Concomitant laparoscopic hand-assisted radical nephrectomy and open radical prostatectomy using a single lower midline incision. AB - The hand-assist technique offers the urologic surgeon several advantages. The technique provides the novice laparoscopist a logical segue into minimally invasive surgery by literally allowing one hand to remain in the realm of open surgery. Hand-assist access affords the laparoscopist the use of tactile sensation and blunt manual dissection and retraction. We describe an additional benefit of the hand-assist technique. In clinical situations in which more than one procedure is required, a properly positioned hand-assist device will avoid the need for two large incisions. We present simultaneous hand-assisted laparoscopic radical nephrectomy and open radical prostatectomy performed through a single midline incision. PMID- 11113764 TI - Paclitaxel-induced stomal neuropathy: a unique cause of pain in a patient with ileal conduit. AB - We present a case of unusual chemotherapy-induced neurotoxicity in a patient who had undergone radical cystoprostatectomy and ileal conduit diversion for invasive bladder cancer. On routine computed tomography scan several years later, he was diagnosed with metastatic transitional cell carcinoma involving the retroperitoneal lymph nodes. The patient received systemic chemotherapy, including a combination of paclitaxel (Taxol) and gemcitabine (Gemzar). During this treatment, the patient developed spasmodic pain and dysesthesia in the stoma area, with no apparent skin irritation or any other local finding. These symptoms resolved about 3 months after completion of the therapy. PMID- 11113765 TI - Diagnosis of prostate adenocarcinoma using transurethral resection of the prostate after multiple negative transrectal biopsies and persistently elevated prostate-specific antigen level. AB - We report a case of prostate cancer diagnosis by transurethral resection of the prostate in a man who underwent more than 50 needle biopsies by three different physicians for an increasing prostate-specific antigen level. Radical prostatectomy resulted in removal of an organ-confined tumor (T2aN0M0), and a short follow-up revealed an undetectable prostate-specific antigen level. PMID- 11113766 TI - Simultaneous bilateral tubeless percutaneous nephrolithotomy. AB - We present what is to our knowledge the first reported case of simultaneous bilateral tubeless (no nephrostomy tube) percutaneous nephrolithotomy. The 64 year-old man was rendered stone free with a single general anesthetic and discharged within 24 hours. The role, indications, and potential benefits of this novel technique are discussed. PMID- 11113767 TI - Priapism: ecstasy related? AB - Priapism, an uncommon urological emergency, is commonly drug-induced. We present a previously unreported case of a young man with priapism probably related to Ecstasy. PMID- 11113768 TI - Retroperitoneal minilaparoscopic nephrectomy in the rat model. AB - OBJECTIVES: Development of small animal models for laparoscopic surgery is important for basic pathophysiologic and oncologic studies, instrument development, and surgical training. Although transperitoneal laparoscopy has been described in the rat, the technical feasibility of the retroperitoneoscopic approach for major renal surgery has not been reported previously. Herein, we describe the development of a rat model for retroperitoneal minilaparoscopic nephrectomy. METHODS: Sixteen male Sprague-Dawley rats underwent a three-port bilateral retroperitoneoscopic nephrectomy using 2 and 3-mm instruments and optics exclusively. After developing the technique in 10 animals, the study was conducted in 6 animals. Following retroperitoneal balloon dilation and CO(2) pneumoretroperitoneum (mean 4.5 mm Hg), nephrectomy was accomplished by intracorporeal en bloc ligation of the renal pedicle. To prevent peritoneal entry, the anterior surface of the kidney was mobilized subcapsularly. Volume of the created retroperitoneal space and peritoneal integrity were confirmed by a contrast x-ray study. Intraperitoneal pressure was monitored constantly during the procedure. RESULTS: Mean surgical time was 74.5 minutes (range 60 to 95) and estimated blood loss was less than 1 mL. Mean volume of the retroperitoneal space was 8.4 mL after initial balloon dilation, and 11.5 mL after nephrectomy. Mean weight of the excised kidneys was 1. 4 g. Inadvertent peritoneotomy occurred during 3 of 12 study nephrectomies. Complications included renal artery hemorrhage leading to death in 1 animal and renal vein injury in 1 animal. CONCLUSIONS: Laparoscopic retroperitoneal nephrectomy in the rat model is technically feasible. This novel small animal model can be used for further studies of the retroperitoneal laparoscopic approach. PMID- 11113769 TI - Absorption of oxybutynin from vaginal inserts: drug blood levels and the response of the rabbit bladder. AB - OBJECTIVES: Oxybutynin has been used for treatment of urge urinary incontinence for more than 20 years. However, one of the major problems with its use is uncomfortable anticholinergic side effects that can lead to discontinuation of treatment. Alternative forms of drug administration may reduce side effects and thus improve patient compliance. METHODS: A cylinder-shaped, curved silicone elastomer insert containing oxybutynin was anchored in the vagina of female rabbits. The inserts were designed to release oxybutynin at rates of 0.5, 1.0, and 5.0 mg/day, respectively. Blood drug and metabolite levels were monitored for 1 to 7 days and cystometry was carried out after 7 days of treatment. RESULTS: There was a consistent dose-dependent absorption of the oxybutynin resulting in stable plasma concentrations by 3 days. Levels of N-desethyloxybutynin, the active metabolite that is thought to be responsible for side effects, were less than 1.0 ng/mL in all groups. The cystometrograms showed a decrease in the detrusor pressures for the higher oxybutynin groups and a dose-dependent decrease in micturition pressure. The vaginal wall in contact with the insert showed no irritation. CONCLUSIONS: The inserts produced stable blood levels and released sufficient amounts of oxybutynin to have measurable effects on the bladder. There was no irritating effect of the insert on the vaginal wall after a 1-week treatment. Vaginal inserts containing oxybutynin may be an interesting alternative method for the chronic delivery of oxybutynin. PMID- 11113770 TI - DNA organization in patients with a history of cryptorchidism. AB - OBJECTIVES: Cryptorchidism is associated with infertility, even in those patients with unilateral undescended testes. The mechanism for this infertility is not understood. We demonstrated recently that in mice, a stable nuclear matrix, a structural component of the nucleus that organizes DNA, is necessary for proper embryogenesis. We tested the hypothesis that spermatozoa from cryptorchid patients had unstable nuclear matrices. METHODS: Semen samples from 7 patients with a history of undescended testes and decreased fertility were tested for sperm nuclear matrix stability using our halo assay. RESULTS: All 7 patients were found to have unstable nuclear matrices, as compared with controls. CONCLUSIONS: This preliminary study suggests that one factor in the decreased fertility of cryptorchid patients may be unstable sperm nuclear matrices. PMID- 11113771 TI - Overexpression and regulation of expression of scatter factor/hepatocyte growth factor in prostatic carcinoma. AB - OBJECTIVES: Scatter factor (hepatocyte growth factor) (SF/HGF) is a multifunctional polypeptide growth factor that has been implicated in tumor proliferation, angiogenesis, invasiveness, and metastasis. Little is known of the expression of SF/HGF in human prostatic carcinoma. The aims of this investigation were to quantitate the level of SF/HGF expression in benign versus malignant human prostatic tissues and to assess regulation of SF/HGF expression by human prostatic stromal myofibroblasts. METHODS: We determined the level of SF/HGF expression in 10 human prostatic tissue samples (5 benign, 5 carcinoma) by Western blot analysis. Five purified growth factors-basic fibroblast growth factor (bFGF), interleukin-1beta (IL-1beta), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), and endothelial growth factor (EGF)-were tested for their capacity to induce SF/HGF expression by a human prostatic stromal myofibroblastic cell line, as assessed by enzyme-linked immunosorbent assay. Supernatant from the normal PrEC prostatic epithelial cell line and the DU 145 carcinoma cell line were assayed for SF/HGF-inducing activity. RESULTS: SF/HGF exhibited a mean fourfold overexpression in carcinoma tissues compared with benign prostatic tissue. Significant stimulation of SF/HGF expression by prostatic stromal myofibroblasts was detected for IL-1beta (8.1 fold), PDGF (6.2-fold), bFGF (4.0-fold), VEGF (3. 7-fold), and EGF (2.9-fold). DU 145-conditioned media, but not the PrEC-conditioned media, contained SF/HGF inducing activity, which was determined to include IL-1beta, bFGF, and PDGF by antibody-blocking experiments. CONCLUSIONS: SF/HGF is overexpressed in human prostatic carcinoma tissues. Prostatic carcinoma cell stimulation of SF/HGF expression by adjacent benign myofibroblastic cells as a type of epithelial stromal paracrine interaction could potentially influence prostatic carcinoma cell behaviors. PMID- 11113772 TI - Evaluation of cadaveric pericardium in the rat for the surgical treatment of Peyronie's disease. AB - OBJECTIVES: To evaluate the intermediate-term efficacy of cadaveric pericardium (Tutoplast) as a grafting material in the surgical correction of Peyronie's disease using a rat model. Peyronie's disease is a connective tissue disorder of the tunica albuginea. When less invasive modalities fail to correct the penile deformity, surgical excision of the plaque and coverage with various grafting materials has been advocated. MFETHODS:Twenty male Sprague-Dawley rats (300 to 325 g) constituted the study population. The animals were divided into two groups: group 1, control rats (n = 10) and group 2, rats that underwent wedge excision of the tunica albuginea and replacement with cadaveric pericardial grafts (n = 10). All rats underwent electrical stimulation of the cavernosal nerve to assess erectile function after 4 months. Tissues obtained after death were stained with trichrome and Verhoff's van Giesen for collagen and elastic fibers. RESULTS: Erectile function as studied by cavernosal nerve stimulation was not significantly different in either group (P >0.05), and histologic studies of penile cross sections of the pericardial graft group revealed a mild to moderate degree of fibrosis surrounding the patch at 4 months. CONCLUSIONS: We found that pericardial cadaveric grafts in a rat model are a suitable tunica albuginea substitute. They allow for penile expansion after cavernosal nerve stimulation and are strong enough to withstand normal intracorporeal pressures. Our early experimental data in the rat support the use of pericardial cadaveric material for coverage of excised Peyronie's plaques. However, long-term follow-up in humans is mandatory. PMID- 11113773 TI - Influence of semen processing technique on human sperm DNA integrity. AB - OBJECTIVES: To compare the effects of density-gradient centrifugation and swim-up technique on sperm DNA integrity. METHODS: Semen samples (n = 22) were obtained from consecutive nonazoospermic men presenting for infertility evaluation. Individual samples were divided into three aliquots (whole semen, density gradient centrifugation, and swim-up) for subsequent analysis of sperm motility and DNA integrity. Sperm DNA integrity was evaluated by flow cytometry analysis of acridine orange-treated spermatozoa and expressed as the percentage of spermatozoa demonstrating denatured DNA. RESULTS: Mean sperm motility (+/-SEM) improved significantly after processing with two-layer density-gradient and swim up compared with whole semen (65.6% +/- 4.0% and 73.0% +/- 3.0% versus 52.0% +/- 3.6%, respectively, P <0.005), with no significant difference in motility between Percoll-treated and swim-up-treated spermatozoa. In contrast, the percentage of spermatozoa with denatured DNA was reduced significantly in swim-up-treated but not in Percoll-treated spermatozoa compared with whole semen (4.8% +/- 1. 2% and 13.6% +/- 3.6% versus 10.1% +/- 2.3%, respectively, P <0. 0001). CONCLUSIONS: Although density-gradient centrifugation is comparable to swim-up technique in recovering spermatozoa with enhanced motility, spermatozoa recovered after swim up possess higher DNA integrity. These data urge us to reexamine our current sperm processing techniques in order to minimize sperm DNA damage. PMID- 11113774 TI - Physical activity and the prevention of breast cancer. AB - Epidemiologists have identified several risk factors for breast cancer, yet clinical advice to women to change these risk factors has been uncommon. Physical activity promises to be one modifiable risk factor through which women can reduce their risk for breast cancer. Clinicians can now advise women that reducing risk for breast cancer may be one additional reason to adopt an active lifestyle. There are still questions about the type and amount of exercise needed, the ages at which exercise should be done, and the interactions with other risk factors such as reproductive and menstrual history, diet, body mass, alcohol intake, genetics, and hormone therapy. Finding answers to these questions will require a research agenda focused on the biology of exercise and breast cancer. PMID- 11113775 TI - Risk factors for osteoporosis: prevalence, change, and association with bone density. AB - OBJECTIVES: To describe the prevalence of risk factors for osteoporosis in a population-based cohort of Australian-born midlife women; determine the effect of these risk factors on premenopausal and early perimenopausal bone mineral density (BMD); and describe changes in risk factors and any effect of these on bone loss. DESIGN: 4-year longitudinal community-based study. BMD of the lumbar spine (LS) and femoral neck (FN) was measured using dual x-ray absorptiometry (DXA). SETTING: Melbourne, Australia PARTICIPANTS: 224 Australian-born women aged 46-56 years MAIN OUTCOME MEASURES: Risk factors for osteoporosis, LS-BMD, FN-BMD, and change in risk factors and BMD. RESULTS: At baseline, 52% reported a calcium intake of less than 800 mg/day and 46% reported a caffeine intake of more than 360 mg/day; 29% exercised less than 1.5 hours/week; 5% had a body mass index (BMI) of less than 20; 14% were current smokers; 23% were past smokers; 10% reported abnormal menstrual histories; and 25% reported a family history of osteoporosis. BMD was positively associated with weight; BMI; and waist, hip, and trunk skin-fold measure (P less than .0005). At 4-year follow-up, there were increases in weight (P less than .0005), waist/hip ratio (P less than. 05), trunk skin-fold measurements (P less than.005), and calcium intake (P less than.05). In women who became late perimenopausal or postmenopausal, bone loss was associated with time in relation to the final menstrual period but not with other variables. CONCLUSIONS: There are multiple risk factors for osteoporosis in this Australian born population of midlife women, but only anthropometric variables were associated with BMD at baseline. Significant changes during the menopausal transition in anthropometric variables and calcium intake were in the direction that could decrease the risk of osteoporosis but were not found to affect menopausal bone loss. PMID- 11113777 TI - Breast cancer risk reduction: what do we know and where should we go? AB - Clinicians should be aware of the advances in breast cancer risk assessment and risk-reduction therapy. The modified Gail model is appropriate for predicting the risk of developing breast cancer within the next 5 years for most women between ages 35 and 75. Tamoxifen has been approved by the U.S. Food and Drug Administration (FDA) for reduction of breast cancer risk in women aged 35 and older who meet the threshold risk for breast cancer. Raloxifene is being compared with tamoxifen in the clinical trial, STAR (a Study of Tamoxifen and Raloxifene), which is now enrolling postmenopausal women aged 35 or older. The risks and benefits of therapy to reduce breast cancer risk are reviewed here. Processes for comparison of risks and benefits and for shared decision making are outlined. PMID- 11113779 TI - Do we need to lighten up? PMID- 11113778 TI - The parathyroid hormones: bone-forming agents for treatment of osteoporosis. AB - This is a 3-part article. Part 1 is an overview of bone formation and resorption and the consequences of estrogen loss on bone. Part 2 comprehensively reviews the most current data on the ability of a family of potent osteoblast-stimulating bone-builders, the native 84-amino-acid parathyroid hormone (PTH), and certain of its 31- to 38- amino-acid fragments to stimulate the growth of animal and human bones. These PTHs are currently in, or about to enter, clinical trial as treatment for postmenopausal osteoporosis. Part 3 provides a detailed consideration of how these PTHs might stimulate bone growth via PTH receptor signaling. PMID- 11113780 TI - Helping terminally ill clients experience a "good death". PMID- 11113781 TI - Creative food tips and dysphagia. PMID- 11113782 TI - Managed care: a maze of new models and terminology. PMID- 11113783 TI - Intensity of care by discipline for selected home health diagnoses. PMID- 11113784 TI - Culturally competent health care in the home. PMID- 11113785 TI - Client expectations and satisfaction of quality in home care services. A consumer perspective. AB - This study examines clients' expectations of quality in home care services and their perceived satisfaction with services among a random sample of 76 home care recipients in Vancouver, Canada. The researchers conducted face-to-face interviews that applied Multiattribute Utility Technology, a procedure that organizes several quality attributes of "ideal" home care into a tree structure to compare their relative importance and ranking from the clients' perspective. Participants also were asked to state their satisfaction or dissatisfaction with the services received in these domains. Among the five main quality attributes identified, the subjects ranked suitability of the home helper and its subset, personal competence, as the most important indicators of quality, followed by continuity in service. In addition, clients tended to have a high level of satisfaction with regard to the attributes of overall home care services. The highest level of satisfaction was reported for elements of personal dispositions of home care staff. The lowest level of satisfaction involved the time/availability components of the service. Finally, comparisons between client expectations and satisfaction of received home care services showed the highest discrepancy for the attributes of influence and time/availability and the greatest congruence for personal attributes of the staff. The results are discussed in terms of their implications for the delivery of home care services. PMID- 11113786 TI - 2001 standards for home health and personal care organizations. PMID- 11113788 TI - Early treatment. PMID- 11113789 TI - Early treatment. PMID- 11113790 TI - Evidence-based orthodontics. PMID- 11113791 TI - Autotransplantation of premolars to replace maxillary incisors: a comparison with natural incisors. AB - The published literature contains no comprehensive studies that compare the outcome of premolar autotransplantation to the maxillary anterior region with natural incisors in the same patients. This article describes the gingival and periodontal conditions around premolars transplanted to the maxillary incisor region, subsequent to restoration. Forty-five premolars autotransplanted to the maxillary incisor region in 40 adolescent patients were evaluated after a mean observation period of 4.0 years. Mean age at surgery was 11.0 years. Established clinical criteria were used to assess tooth mobility, plaque and gingival indexes, probing pocket depth, and percussion. Recession and hyperplasia of interproximal gingival papillae were assessed according to a recently proposed index. Standardized radiography was used to evaluate presence of pathosis, pulp obliteration, root length, and crown-root ratios. Clinical variables for transplants did not differ from those of the natural incisors, except for increased mobility and more plaque in a few transplanted premolars. The interproximal gingival papillae adjacent to all transplanted teeth were normal or slightly hyperplastic. Radiographically, all transplants showed varying degrees of pulp obliteration, but no signs of pathosis. Crown-root ratios were similar for natural and transplanted teeth as were distances from cementoenamel junction to marginal bone. The overall status of the transplanted premolars and surrounding tissues indicated that this treatment modality may be recommended when maxillary incisors are missing in adolescents. In addition, tooth transplantation represents an inherent potential for bone induction and reestablishment of a normal alveolar process. PMID- 11113792 TI - Endosseous titanium implants as anchors for mesiodistal tooth movement in the beagle dog. AB - The purpose of this study was to determine the anchorage potential of titanium implants (Branemark; 3.75 x 7 mm) with the use of a sectional arch wire technique for orthodontic mesiodistal tooth movement, as assessed by the osseointegration of implants and tooth movement. Two implants were surgically placed in healed mandibular extraction sites of the second and third premolars on each side in 4 adult male beagle dogs. The implants were surgically uncovered 18 weeks later, and second-stage abutments with soldered edgewise tubes were attached. Segmented edgewise rectangular archwires (0.017 x 0. 025 inch) with a T-loop or an L-loop were placed between the implants and the fourth premolars on both sides as the anchorage unit. One segment in each dog served as a loaded side, and the archwire was calibrated to produce 200 g of lateral force on the fourth premolar. The contralateral segment served as an unloaded side and was not subjected to orthodontic force. Sectional wires were activated biweekly 24, 28, 28, and 32 weeks, respectively, depending on the magnitude and the appearance of mesial tipping movement of the fourth premolar. After mandibular impressions were taken to measure the distance between the first molar and the fourth premolar, the animals were euthanized and dissected mandibles were prepared. The specimens were then embedded in polyester resin and cut to take backscattered electron images. On the basis of these images, the percentage of peri-implant bone volume was calculated and defined as an index of osseointegration. The differences between the initial and final fourth premolar to first molar distances varied (7.40, 8.85, 10.50, and 3.30 mm) on the loaded side, whereas the unloaded side showed no movement. Not only was there no statistical difference in the percent of peri implant bone volume between the loaded and unloaded sides, but there was also no statistical difference between the compression and tension sides in both loaded and unloaded implants, which suggests that the implants maintained rigid osseointegration. In conclusion, the present study demonstrated that endosseous titanium implants can function as anchors for long-term orthodontic mesiodistal movement. PMID- 11113793 TI - Tooth anomalies associated with failure of eruption of first and second permanent molars. AB - The occurrence of tooth anomalies in association with failure of the first and second molars to erupt was assessed in a sample of 1520 nonsyndromic subjects with uncrowded dental arches (mean age, 14 years 4 months) and compared with the prevalence rate calculated in a matched control group of 1000 subjects. The tooth anomalies examined included infraocclusion of deciduous molars, palatal displacement of maxillary canines, rotation of maxillary lateral incisors, aplasia of second premolars, and small size of maxillary lateral incisors. Associations among arrested eruption of first and second permanent molars and anomalies in tooth eruption and position (infraoccluded deciduous molars, palatally displaced canines, rotated maxillary lateral incisors) were highly significant (P <. 001). No significant association was found among the occurrence of molar eruption disturbances, aplasia of premolars, and small-sized laterals. These findings point to a common biologic cause for the appearance of failure of eruption of molar teeth and other disturbances in tooth eruption and position, most likely under genetic influence. PMID- 11113794 TI - Effect of serial extraction alone on crowding: spontaneous changes in dentition after serial extraction. AB - We examined changes in dentition after serial extraction in subjects who wore no appliances to determine the relationships between changes in dentition and improvement in dental crowding. Mandibular dental casts and lateral cephalograms from 31 subjects who had undergone serial extraction without orthodontic treatment were analyzed at 3 stages: before extraction of the deciduous canines (T1), after extraction of first premolars (T2), and at the end of the observation period (T3). Although movements of the first molar cusp and apex from T2 to T3 were significantly greater than from T1 to T2, movements of the incisor cusp and apex from T1 to T2 were significantly greater than from T2 to T3. The first molar tipped mesially from T1 to T2 but tipped distally from T2 to T3. The distal tipping of the incisor from T1 to T2 was significantly greater than from T2 to T3. These results suggest that the main changes in dentition from T1 to T2 are different from those from T2 to T3. The correlations between the annual change in the canine movement or tipping from T2 to T3 and the annual change in the irregularity index at the same time were significant. These results suggest that the canine movement or tipping contributed to the correction of the anterior crowding from T2 to T3. PMID- 11113795 TI - Short-term effects of fiberotomy on relapse of anterior crowding. AB - The effects of fiberotomy were evaluated in alleviating dental relapse of incisors after orthodontic treatment. The study sample consisted of 23 patients with crowded maxillary and mandibular incisors before orthodontic treatment. The amount of initial crowding was determined according to Little's irregularity index. Fiberotomy procedures were performed on 11 of the patients 1 week before debonding. The other 12 subjects served as the control group. All patients wore Hawley retainers. Lateral cephalometric headfilms and dental casts of the patients were taken at the beginning (T1) and at the end (T2) of treatment, 6 months into the retention phase (T3), and 1 year after orthodontic treatment (T4). Significant increase of irregularity index was noted in the control group at T3 and T4 for both maxillary and mandibular anterior segments (P <.05, P <.01). Meanwhile, in the group where circumferential supracrestal fiberotomy was performed, no significant increase of the irregularity index was noted. PMID- 11113796 TI - Triangular-shaped incisor crowns and crowding. AB - The purpose of this study was to establish if there is a correlation between the shape of the labial crowns of the incisors and crowding. Plaster cast models of 69 untreated individuals (30 males and 39 females) were evaluated. The casts were selected randomly from the collection at Seoul National University and Ajou University. With the use of Little's irregularity index, the sample was divided into 2 groups, a crowded group and a normal group. Repeated measurements of the maximum mesiodistal width of the incisal and cervical areas of the incisors were taken by means of a digital vernier caliper and a ratio of these measurements was calculated. The mean value for the crowded group was significantly larger in the incisal area (P <.01) and smaller in the cervical areas (P <.01) than corresponding values in the normal group. These ratios were correlated with the irregularity index (Pearson r(2) from 55% to 65%). The value obtained from the incisor width ratio in the normal group can be useful for the diagnosis and treatment of crowded malocclusion. PMID- 11113797 TI - An evaluation of preoperative ibuprofen for treatment of pain associated with orthodontic separator placement. AB - Patients undergoing orthodontic treatment can experience significant levels of pain. This study assessed the effectiveness of preoperative ibuprofen in reducing the incidence and the severity of pain after orthodontic separator placement. Sixty-three adolescent patients (mean age, 13 years) were included in this randomized, double-blind, placebo-controlled, prospective study. Patients were randomly assigned to 1 of 3 experimental conditions: (1) 400 mg of ibuprofen taken orally 1 hour before separator placement and a lactose placebo taken orally immediately after the appointment, (2) a lactose placebo taken orally 1 hour before separator placement and 400 mg of ibuprofen taken orally immediately after the appointment, or (3) a lactose placebo taken orally 1 hour before separator placement and again immediately after the appointment. The patient's level of discomfort was assessed with a visual analog scale at 2, 6, and 24 hours, as well as at 2, 3, and 7 days after placement of the orthodontic separators. An analysis of variance and Duncan's multiple range test revealed that 2 hours after their orthodontic appointment the patients who had taken ibuprofen 1 hour before separator placement had significantly less pain with chewing than did the patients who received either ibuprofen postoperatively or a placebo. Additional measures suggest a trend for less pain for this group of patients. These results support the use of pretreatment ibuprofen for patients requiring analgesics for orthodontic discomfort. Future study of the use of preemptive analgesics in orthodontics is warranted. PMID- 11113798 TI - Cephalometric variables as predictors of Class II treatment outcome. AB - Cephalometric analysis of skeletodental features is accepted as an integral part of orthodontic diagnosis and treatment planning. This assumes that diagnostic cephalometric variables affect prognosis and thus help reduce malocclusion severity, which is the aim of orthodontic treatment. The aim of this study was to assess the predictive value of 41 commonly used cephalometric parameters with regard to pretreatment severity and treatment outcomes. Pretreatment severity was assessed by using the Peer Assessment Rating (PAR) occlusal index, an instrument that has been shown to be valid and reliable. Treatment outcomes consisted of (1) posttreatment malocclusion severity (post-PAR), (2) relative improvement (percent PAR reduction), and (3) treatment duration. Complete records, including cephalograms, of 223 treated Class II cases were analyzed by means of separate multiple linear regression models. Each of the outcome variables and the pretreatment severity served as the respective dependent variables, and the cephalometric parameters served as the independent or predictor variables. The cephalometric parameters explained 39.2% of the pretreatment severity variance, 17. 9% of posttreatment severity variance, 15.7% of relative treatment improvement variance, and 20.0% of treatment duration variance. PMID- 11113799 TI - Skeletal and dental changes with nonextraction Begg mechanotherapy in patients with Class II Division 1 malocclusion. AB - This prospective cephalometric study was undertaken to assess the mode and magnitude of Class II correction with nonextraction Begg mechanotherapy in growing children. The sample comprised subjects with similar malocclusion and age range (9-12 years) who were specifically selected for nonextraction Begg mechanotherapy. Cephalograms were analyzed to assess the skeletal, dental, and soft tissue changes that occurred after correction of the molar relationship, the overjet, and the overbite during the 9-month treatment period. The results revealed a significant improvement in the anteroposterior jaw relationship, suggested by the significant reduction in the ANB angle (1.62 degrees ) and in Wits AO-BO (1.42 mm). The mandibular length increase of 0.56 mm suggests that the Class II elastics used in nonextraction Begg mechanotherapy had a minimal stimulatory effect on mandibular growth. There was a significant increase in the anterior and posterior facial heights and the ramal height. Almost all of the dental changes were significant. The most striking feature were a significant retraction and extrusion of the maxillary incisors and proclination and intrusion of the lower incisors accompanied by extrusion of the mandibular molars. The maxillary incisors extruded by 1.64 mm under the influence of the undesirable downward component of the Class II elastic forces. The major contribution to overjet and molar correction was predominantly dentoalveolar. PMID- 11113800 TI - Growth of the anterior dental arch in black American children: a longitudinal study from 3 to 18 years of age. AB - Dental arch size and form change systematically because of tooth emergence and because teeth migrate into shorter and broader arch forms in the deciduous dentition and again in the permanent dentition. The present longitudinal analysis describes changes in arch form in a cohort of 52 black American children (Nashville, Tenn) between the ages of 3 and 18 years. The anterior (incisor canine) arch dimensions were analyzed. Incisor-to-canine depth remained static in both arches between 3 and 5 years but shortened significantly between 12 and 18 years. Intercanine width broadened significantly in both arches, first during the deciduous dentition, then again as the primary teeth were supplanted by the permanent incisors and canines. But there was no change in intercanine width once the permanent canines were in functional occlusion (approximately 11-18 years). These changes alter anterior arch form (the ratio depth/width). This index decreased significantly during the deciduous phase as the arches broadened; the index increased from 6 to 10 years as teeth were replaced, then again decreased during the duration of the study (approximately 10-18 years). Dimensions of these black American children all exceed comparable values for white American children, although the anterior shapes are indistinguishable. The present data focus attention on the dynamics of arch form in which considerable, protracted change occurs by physiologic migration, not just during the short phase of tooth replacement. PMID- 11113801 TI - The relationship of the glenoid fossa to the functional occlusal plane. AB - The geometric relationship of the functional occlusal plane to the center of the glenoid fossa, as seen in the sagittal view, is described for males and females from age 7 through 25 years. This relationship is fully described by the distance from the geometric center of the glenoid fossa perpendicular to the functional occlusal plane (L, in millimeters) and its angulation (theta, in degrees) relative to a constructed Frankfort horizontal (SN-7 degrees ). Regression formulas with 95% confidence levels are described for L and theta for both genders combined. The differences between genders were found to be statistically insignificant. No correlation was found between these dimensions relative to the presence of temporomandibular joint dysfunction symptoms. A statistically significant difference in both L and theta was found in males who exhibited a Class III relationship compared with males exhibiting a Class II relationship. PMID- 11113802 TI - The friction and wear patterns of orthodontic brackets and archwires in the dry state. AB - Frictional resistance at the bracket-archwire interface has been demonstrated to impede tooth movement when sliding mechanics are used. Thus, the coefficients of friction of titanium and stainless steel brackets used in conjunction with stainless and ion-implanted beta-titanium archwires were investigated using a single contact interface between the brackets and archwires. The wear patterns between the brackets and the.016- in flat archwire surfaces were also examined using scanning electron microscopy and energy dispersive x-ray analysis. Stainless steel brackets tested with. 016-in flat stainless steel wire surfaces recorded the lowest coefficient of static friction mean (0.289), whereas titanium brackets paired with.016-in flat ion-implanted beta-titanium wire surfaces produced the highest mean (0.767). Stainless steel brackets had significantly (P <.05) lower coefficients of friction than titanium brackets for all wires except.020-in round stainless steel wires. Ion-implanted beta-titanium wires generally had significantly larger coefficients of friction than stainless steel wires. The increased friction of the titanium and ion-implanted beta-titanium alloys is also reflected in the severity of their wear patterns. An inverse relationship between friction and archwire surface dimension was generally found for ion-implanted beta-titanium wires. Round stainless steel wires demonstrated lower coefficients of kinetic friction than the flat stainless steel wire surfaces. PMID- 11113803 TI - Evaluation of titanium brackets for orthodontic treatment: Part II--The active configuration. AB - After each archwire was ligated into a bracket with a 0.010-in stainless steel wire, both stainless steel and beta-titanium archwires (0.017- x 0.025-in) were slid through commercially pure titanium brackets (0.018-in slot size) at 34 degrees C in both the dry and wet conditions. As controls, stainless steel archwire versus stainless steel bracket couples were used with comparable dimensions. The drawing forces were measured at 5 angulations (0 degrees, 3 degrees, 7 degrees, 9 degrees, and 11 degrees ) for 5 normal forces (nominally 0.2, 0.4, 0.6, 0.8, and 1.0 kg). Regression lines were determined for each frictional couple (P <.05). In the passive configuration, the kinetic frictional coefficients of control and test couples in the dry condition were comparable to previously reported values at 0.11 +/- 0.01 for stainless steel versus stainless steel, 0.12 +/- 0.00 for stainless steel versus titanium, and 0.26 +/- 0.02 for beta-titanium versus titanium. As the angulation was increased from 0 degrees to 11 degrees and the normal force was maintained at 0.2 kg, the resistance to sliding values increased by 208 g for stainless steel versus stainless steel, by 222 g for stainless steel versus titanium, and by 185 g for beta-titanium versus titanium. When the normal force was increased to 1.0 kg, the resistance to sliding values increased to 277 g, 246 g, and 245 g, respectively. Although resistance to sliding increased with angulation and normal force, the passive layer did not breakdown. Titanium brackets remained comparable to stainless steel brackets in the active configuration. PMID- 11113804 TI - Nickel-titanium alloys: stress-related temperature transitional range. AB - The inducement of mechanical stress within nickel-titanium wires can influence the transitional temperature range of the alloy and therefore the expression of the superelastic properties. An analogous variation of the transitional temperature range may be expected during orthodontic therapy, when the archwires are engaged into the brackets. To investigate this possibility, samples of currently used orthodontic nickel-titanium wires (Sentalloy, GAC; Copper Ni-Ti superelastic at 27 degrees C, 35 degrees C, 40 degrees C, Ormco; Nitinol Heat Activated, 3M-Unitek) were subjected to temperature cycles ranging between 4 degrees C and 60 degrees C. The wires were mounted in a plexiglass loading device designed to simulate clinical situations of minimum and severe dental crowding. Electrical resistivity was used to monitor the phase transformations. The data were analyzed with paired t tests. The results confirmed the presence of displacements of the transitional temperature ranges toward higher temperatures when stress was induced. Because nickel-titanium wires are most commonly used during the aligning stage in cases of severe dental crowding, particular attention was given to the performance of the orthodontic wires under maximum loading. An alloy with a stress-related transitional temperature range corresponding to the fluctuations of the oral temperature should express superelastic properties more consistently than others. According to our results, Copper Ni-Ti 27 degrees C and Nitinol Heat-Activated wires may be considered suitable alloys for the alignment stage. PMID- 11113805 TI - Treatment of a patient with a Class II malocclusion, impacted canine, and severe malalignment. AB - A case report of the orthodontic treatment of a male adolescent with a unilateral dental Class II malocclusion, an impacted canine, severe maxillary malalignment, and a canted maxillary anterior occlusal plane. Treatment consisted of full fixed appliances, extraction of the maxillary right first premolar, and surgical exposure of the impacted canine. Treatment vastly improved the patient's facial and dental esthetics. A Class I skeletal and dental relationship was established, along with a functional anterior guidance. The dental arches were coordinated and the dental midlines coincident with the midsagittal plane. This case report was presented to the American Board of Orthodontics in partial fulfillment of the requirements for the certification process conducted by the Board. PMID- 11113806 TI - Orthodontic tooth movement after extraction of previously autotransplanted maxillary canines and ridge augmentation. AB - A case report is detailed in which autotransplanted maxillary canines were removed and the spaces closed. Substantial surrounding bone loss was associated with the upper right canine, and a bone graft was needed to reestablish normal dentoalveolar ridge morphology. Bone was taken from the maxillary tuberosity and placed in the canine extraction site, fixed with a bone screw, and covered with GoreTex. Seven months after placement of the bone graft, the GoreTex and stabilizing screw were removed to allow for consolidation of the bone. The upper left canine and lower second premolars were extracted, and fixed appliances were placed in both arches to align the teeth and close the spaces. Protraction of the upper right first premolar and retraction of the lateral incisor into the graft site were kept slow and constant with continued periodontal assessment. During the space closure, there was some concern that the bone in the graft site might resorb, leaving the teeth with compromised periodontal support. However, no significant periodontal attachment loss occurred despite ongoing concern about the amount of keratinized tissue. Perhaps the relatively slow rate of tooth movement provided for bone to be maintained and recreated ahead of the tooth. Almost complete closure of the upper canine extraction spaces was achieved. The upper premolars were substituted for the maxillary canines, and unfavorable prosthetic options were thus avoided. The lower arch was aligned, and the extraction spaces completely closed. PMID- 11113808 TI - Parameters for digital imaging: Part 2. PMID- 11113809 TI - Litigation, Legislation, and Ethics. Show me the money: A brief history of the expert witness for hire. PMID- 11113810 TI - Evaluation of microvascular bone graft reconstruction of the head and neck with 3 D 99mTc-DPD SPECT scans. AB - OBJECTIVE: We conducted a prospective investigation to evaluate the diagnostic accuracy of computer-aided 3-dimensional (3-D) technetium 99m dicarboxypropane methylene diphosphonate ((99m)Tc-DPD) single photon emission computed tomography (SPECT) reconstruction in the evaluation of microvascular bone flaps used for maxillofacial reconstruction. STUDY DESIGN: Twenty patients who received 20 autogenous microvascular bone flaps for reconstruction of the mandible and maxilla were evaluated. Forty bone scans with subsequent computer-aided reconstruction were performed. Each graft could be assessed within 48 to 72 hours after surgery. The second bone scan was performed between 12 and 14 days after surgery. RESULTS: Complications were observed in 5 grafts. SPECT investigation performed at the 2 time points after reconstruction showed a significantly higher tracer uptake in grafts with an uncomplicated further course than in those that developed complications. CONCLUSIONS: Computer-aided 3D (99m)Tc-DPD SPECT reconstruction serves as a useful prognostic tool and helps in the very early recognition of complications. This technique adds significantly to the value of planar bone scintigraphy and conventional SPECT images. PMID- 11113811 TI - Genioglossus muscle attachments: an anatomic analysis and the implications for genioglossus advancement. AB - PURPOSE: To determine the exact dimensions of the attachments of musculature to the genial tubercles and to describe their relationships to the anterior mandible. MATERIALS AND METHODS: Ten randomly selected adult cadavers with intact mandibular dentition and without periodontal disease underwent midline sectioning and precise measurements with a caliper to evaluate the dimensions of the attachments of associated musculature. RESULTS: The average thickness of the mandible at the genial tubercle was 12.6 mm. The average thickness of the mandible at the inferior border at pogonion was 14.5 mm. The average distance from the genial tubercle to the inferior border was 14.2 mm. The average distance from the apex of the incisors to the genial tubercle was 11.8 mm. The width of the genioglossus muscle itself was 13.8 mm. CONCLUSIONS: This study precisely evaluates the attachment area of the genioglossus muscle and its relationship to the anterior mandible. These findings help estimate the dimensions of osteotomies made in the anterior mandible for the treatment of obstructive sleep apnea to offer the greatest amount of muscular advancement. PMID- 11113812 TI - Management of tongue cancer in the patient who is systemically immunosuppressed: a preliminary report. AB - OBJECTIVE: Renal transplantation is being improved, and life expectancy for patients with transplanted kidneys is being prolonged, but the incidence of malignant tumors in other body organs or tissues is increasing progressively. We investigated effective clinical plans for treating lingual cancer in persons who are long-term renal transplant recipients. STUDY DESIGN: The group included 3 cases of lingual cancer after renal transplantation. All were men. The mean age was 47.5 years (range, 40 to 55). The lingual cancer occurred a mean of 9 years after renal transplantation. The lesions of two patients were in the left lingual margins; the other patient had multiple bilateral lesions. All diagnoses were confirmed microscopically to be squamous cell carcinomas stage I to II. RESULTS: The doses of immunosuppressive agents were decreased, and two patients accepted surgery, chemotherapy, and radiotherapy. The third patient died 1 year after the cancer was diagnosed because of metastases. The other 2 patients had regular follow-up for 2 to 3 years and did not have either local recurrence or metastasis. CONCLUSION: Patients who have lingual cancer develop after renal transplantation have received long-term immunosuppressive therapy. We suggest the following clinical protocol: (1) Besides regular evaluation after renal transplantation, physical examination and biopsy of suspicious oral lesions are necessary because of the possibility of postrenal malignancy. (2) Surgery, along with chemotherapy and radiotherapy, is the main treatment of lingual cancer after renal transplantation. (3) The lingual tumor should be comprehensively treated despite the poor immune state of the body, or tumor recurrence can accelerate and metastases can occur. (4) Treating cancer thoroughly and maintaining the function of the transplanted kidney can decrease the patient's immune status. We should continue to observe and to report the patients' protocol, even if the patients' clinical follow-up periods are limited. PMID- 11113813 TI - Human herpesvirus 8 and oral health care: an update. AB - OBJECTIVE: The purpose of this report was to review the current literature on human herpesvirus 8 (HHV-8) with particular attention to the aspects of interest for dental health care workers. MATERIAL AND METHODS: The authors searched original research and review articles on specific aspects of HHV-8 infection, including virology, epidemiology, transmission, diagnosis, natural history, therapy, and oral aspects. The relevant material was evaluated and reviewed. RESULTS: HHV-8 is a recently discovered DNA virus that is present throughout the world but with major geographic variation. In the Western world, the virus, transmitted mainly by means of sexual contact, is strongly associated with Kaposi's sarcoma and body cavity-based lymphoma and more controversially with multiple myeloma and non-neoplastic disorders. There is no specific effective treatment, but human immunodeficiency virus protease inhibitors may play an indirect role in the clearance of HHV-8 DNA from peripheral blood mononuclear cells of patients infected with human immunodeficiency virus. HHV-8 DNA is present in saliva, but as yet, there are no documented instances of nosocomial transmission to health care workers. The prevalence of HHV-8 among dental health care workers is probably similar to that in the general population. CONCLUSION: HHV-8 does not appear to be ubiquitous in most populations, particularly in western Europe and the United States, where it may be restricted to a population at risk of having Kaposi's sarcoma develop (men infected with human immunodeficiency virus and patients who are iatrogenically immunosuppressed). Most serologic studies suggest a global HHV-8 seroprevalence of 2% to 10% and show that the virus may be under immunologic control in people who are healthy but infected with HHV-8. Also, HHV-8 certainly has the means to overcome cellular control and immune responses and thus predispose to malignancy. To date, there are no data to suggest that health care staff members are at particular risk of HHV-8 acquisition through occupational routes. PMID- 11113814 TI - Porphyria cutanea tarda affecting lower lip. AB - In porphyria cutanea tarda, the most common type of porphyria, blisters of the skin occur because of high sensitivity to sunlight. This case report describes porphyria cutanea tarda of the lower lip mimicking an actinic cheilitis in a 62 year-old man with a history of Hodgkin's lymphoma. PMID- 11113815 TI - Pediatric sialolithiasis. AB - Pediatric sialolithiasis is a rare condition. This article characterizes 15 cases in children between 5 and 14 years of age. The diagnoses of this condition were made with routine radiograph and ultrasound, as well as with sialography whenever possible. Sialoendoscopy was performed as a diagnostic and treatment modality. Thirteen of the 15 affected children were boys, and 12 of 15 cases occurred in the submandibular gland. We were able to diagnose 67% by our imaging methods; the remainder were diagnosed by clinical examination. PMID- 11113816 TI - Resolution of recalcitrant human papillomavirus gingival infection with topical cidofovir. AB - Cidofovir, a purine nucleotide analogue of cytosine, is a promising new drug that acts against a wide number of DNA viruses. In 1997, the Food and Drug Administration approved intravenous cidofovir for the treatment of cytomegalovirus retinitis in patients with acquired immunodeficiency syndrome. Recent studies have shown cidofovir (1% gel or cream) to be effective for the treatment of recalcitrant and unmanageable viral cutaneous lesions induced by herpes, pox, and papillomavirus families. We report the case of a 45-year-old man who had been diagnosed as having acquired immunodeficiency syndrome in 1995. Recalcitrant to conventional therapies, the human papillomavirus lesions on his gingival mucosa were successfully treated with cidofovir 1% cream. To the best of our knowledge, this is the first case in which topical cidofovir has been used for the treatment of a human papillomavirus infection of the oral mucosa. PMID- 11113817 TI - Ameloblastic carcinoma ex ameloblastoma of the mandible with malignancy associated hypercalcemia. AB - Ameloblastoma is a rare, locally destructive, benign neoplasm of the jawbones, which arises from epithelium derived from the epithelial components of the developing tooth. Ameloblastic carcinoma is the term used to designate any ameloblastoma in which there is histologic evidence of malignancy in the primary tumor, regardless of whether it has metastasized. Most ameloblastic carcinomas are presumed to have arisen de novo, with few cases of malignant transformation of ameloblastoma being apparent. Hypercalcemia is the most common metabolic complication of malignancy. Although malignancy-associated hypercalcemia is often reported in association with other malignancies, it is exceedingly unusual in association with ameloblastoma, malignant ameloblastoma, or ameloblastic carcinoma. We describe a patient with multiple recurrences of ameloblastoma, with subsequent malignant transformation presenting with malignancy-associated hypercalcemia. PMID- 11113818 TI - Proliferative verrucous leukoplakia of the gingiva. AB - OBJECTIVE: The purpose of this study was to describe the clinical-pathologic features of what appears to be a gingival form of proliferative verrucous leukoplakia. STUDY DESIGN: Ten adult patients with recurrent and histologically progressive gingival leukoplakias who were diagnosed and treated at the University of California, San Francisco between 1994 and 1999, comprised the subject group for this investigation. Clinical and microscopic features were reviewed. Proliferation indices and p53 expression were evaluated immunohistochemically, and the presence of human papillomavirus (HPV) DNA was determined by using polymerase chain reaction (PCR) amplification. RESULTS: Lesions presented as solitary or regional flat/papillary/verrucal leukoplakias of the free and attached gingiva (tooth-bearing areas only). With time, flat lesions developed a papillary or verruciform profile. Although lesions were recurrent, they were confined to the gingiva, and multiple lesions did not develop. Half the patients used tobacco, and HPV could not be detected by using PCR. Microscopically, 6 cases began as hyperkeratotic lesions, and 4 initially exhibited a psoriasiform pattern with a marked inflammatory component. With recurrences, the lesions became progressively atypical histologically. The proliferation indices for these lesions showed modest increases over normal epithelium, and positive p53 staining was evident in 4 of 10 cases, indicating a disruption of the keratinocyte cell cycle in these lesions. The mechanism associated with the positive p53 staining (protein binding to wild type p53 versus mutation of the p53 gene) was not determined. Lesions recurred after conservative scalpel or laser excision, and many developed into verrucous or squamous cell carcinoma. CONCLUSIONS: Proliferative verrucous leukoplakia of the gingiva (PVLG) appears to be a subset of oral proliferative verrucous leukoplakia. It can be characterized as a solitary, recurring, progressive white patch that develops a verruciform architecture and may not be associated with HPV. PVLG has an unpredictable course and is at risk for development into verrucous or squamous cell carcinoma. Currently, there is no way to determine or predict which gingival white lesions will follow the clinical course described for this group of patients with PVLG. PMID- 11113819 TI - Superficial arteriovenous hemangioma of the oral cavity. AB - OBJECTIVE: Superficial arteriovenous hemangioma (AH) is a benign vascular lesion that often affects the head and neck, but only 3 histologically proven intraoral lesions have been previously reported. The aim of this study was to analyze the clinical and histologic features of 36 oral AHs from 35 patients. STUDY DESIGN: All vascular lesions, other than pyogenic granulomas, accessioned between 1952 and 2000 were retrieved, and clinical details were gleaned from the request forms or, when available, from the case notes. Histologic sections stained with hematoxylin-eosin were reviewed, and all selected cases were stained for smooth muscle actin and elastin. RESULTS: The age range was 12 to 90 (mean, 53 years; median, 59). Of all patients, 54% were female. All the lesions were solitary. The labial mucosa or vermilion was affected in 17 (49%) patients, the tongue in 5, the hard palate and cheek mucosa in 4 each, the gingiva or alveolar mucosa in 3, and the floor of the mouth in 1. Clinical presentation was most often a raised lesion smaller than 20 mm. Duration ranged from months to many years. Four lesions recurred. The consistent histopathologic feature was an unencapsulated mass of blood vessels located in the lamina propria, the submucosa, or both, but 3 patterns could be discerned. The most common (in 44% of cases) comprised a diffuse mixture of thick-walled and thin-walled vessels in variable proportions. The second pattern (42%) showed a lobular arrangement of smaller vessels of uniform wall thickness, sometimes associated with an arteriole. The remaining 14% showed a tortuous pattern of angular, branching, thick-walled vessels lacking inner elastic laminae; one lesion showed both this pattern and the first pattern, one showed both the lobular morphology and the third pattern. Of the patients with the lobular arrangement, 64% were female. Smooth muscle actin was a major component of the lesional vessel walls regardless of thickness, but although most contained some fibrillary elastin, none had inner elastic laminae as prominent as those seen in adjacent true arteries. All AHs contained plump endothelial cells, and mast cell numbers were increased. CONCLUSIONS: The etiology of AH is uncertain; endocrine and inflammatory stimuli may activate an underlying vascular malformation. Some lesions, especially those in younger patients, may be true hamartomas. PMID- 11113820 TI - Periapical central giant cell granuloma: a potential endodontic misdiagnosis. AB - This retrospective study ascertained the incidence and clinicopathologic features of central giant cell granulomas (CGCGs) associated with teeth with necrotic pulps or teeth that had received previous endodontic treatment and determined whether periapical CGCGs can result in endodontic misdiagnosis. Clinical and histopathologic data of biopsy specimens diagnosed as CGCG were collected from the archives of the Oral Pathology Laboratory, Temple University, and were reviewed. Over the 9-year period, 16 of 79 cases (20%) of CGCG were associated with a tooth that had a history of pulp necrosis. Of those, 14 (88%) were associated with previous root canal treatment. The data from this series of 79 cases of CGCG also showed that CGCGs were less common in women, less common before age 30, and did not cross the midline of the jaw as often as previously reported. Clinical and histopathologic data were compared from (1) CGCGs associated with teeth with vital pulps or that occurred in edentulous areas; (2) CGCGs associated with teeth with necrotic pulps; and (3) 194 cases of periapical granulomas and radicular cysts. These data strongly suggest that CGCGs associated with teeth with necrotic pulps are not directly related to periapical inflammation and may be misdiagnosed as endodontic lesions. Posttreatment follow up and routine submission of periapical surgical specimens are emphasized. PMID- 11113821 TI - Paresthesia of the mental nerve induced by periapical infection: a case report. AB - Paresthesia can be a rare complication of infections of dental origin. This article presents a case of anesthesia/paresthesia caused by a periapical infection of the right mandibular second premolar. The sensory disturbance disappeared 2 weeks after conventional endodontic treatment associated with antibiotic therapy. Twelve months later, the tooth was still asymptomatic. The possible mechanisms responsible for paresthesia associated with periapical infection are discussed. PMID- 11113822 TI - The use of SPECT bone scans to evaluate patients with idiopathic jaw pain. AB - OBJECTIVE: The purpose of this study was to investigate the potential usefulness of single photon emission computed tomography (SPECT) bone scanning with technetium-99m methylene diphosphonate (Tc-99m MDP) in the diagnosis of idiopathic jaw pain. Unlike planar bone scanning, SPECT uses tomographic technology to provide 3-dimensional images, which are more useful in localizing small lesions. STUDY DESIGN: Twenty patients, each with a diagnosis of chronic idiopathic jaw pain, were compared after SPECT bone scanning with 20 age-matched and gender-matched normal controls. Uptake was identified and compared in sites with previously detected jaw pathoses and jaw pain. RESULTS: Nineteen of 20 patients with jaw pain evaluated with SPECT had positive scans, in contrast with 12 of 20 control subjects (P <.04). Positive scans were correlated with painful sites in 15 of 20 patients, with the remaining 5 patients demonstrating no uptake in painful locations. Patients with jaw pain demonstrated 37 of 80 mouth quadrants with positive scans, in contrast with 21 of 80 mouth quadrants in the controls (P <.01). Nineteen of 24 painful mouth quadrants had uptake in the pain group. Of the 21 quadrants positive in the controls, 17 were correlated with previously detected jaw pathoses. The sensitivity and specificity for detecting painful sites were 0.79 and 0.68, respectively. The sensitivity and specificity for detecting previously identified pathoses in the jaws of normal controls were 0. 80 and 0.93, respectively. CONCLUSION: Patients with idiopathic jaw pain had a significantly greater frequency of positive SPECT bone scans when compared with normal controls. However, the sensitivity and specificity of SPECT bone scans in detecting painful sites were low. These findings suggest that SPECT bone scanning with Tc-99m MDP is not indicated as a routine imaging procedure for the detection of jaw pathoses, but may be considered as a potential research tool in the future study of chronic idiopathic jaw pain. PMID- 11113823 TI - Synovial chondromatosis of the temporomandibular joint: varying presentation in 4 cases. AB - Synovial chondromatosis is a rare condition in which cartilage is formed in the synovial membrane of a joint. The manifestations of this benign neoplastic process can mimic many common temporomandibular joint and parotid diseases. Four cases of synovial chondromatosis are presented. In each case, atypical presentation, coexisting joint disease, or both caused diagnostic confusion. The histories and physical examinations were initially consistent with more common joint diseases in each case. Imaging provided some insight into diagnosis and was a definitive indication for surgical treatment. Treatment by subtotal synovectomy and by removal of chondromatous nodules were undertaken in each case. No patient in our series has had recurrence of disease or symptoms after surgical treatment. PMID- 11113824 TI - Cherubism: a 36-year long-term follow-up of 2 generations in different families and review of the literature. AB - OBJECTIVE: To clarify the relationships between the varying clinical or radiographic features of cherubism. STUDY DESIGN: Nonparametric statistics were used in a long-term follow-up of 18 patients through 2 generations from 6 Danish families. RESULTS: The radiographic grade of cherubism was significantly related to sex, maximal buccal bone expansion, course of cherubism, and number of aplasia or ectopic impacted teeth, but it was not related to families. Normal dentition in nonaffected regions was present or was obtained in 14 of 14 patients (age, >14 years). Surgical treatment did not provoke growth of lesional tissue in 22 of 22 cases. Radiographically, the bone structure in the lesional areas was related to age in all grades of cherubism: new bone formation in radiolucent areas (age, >20 years), normal bone structure with multilocular sketches (age, 32 to 39 years), and completely normal bone structure (age, >41 years), also found in 7 of 7 carriers of cherubism (age, >32 years). CONCLUSIONS: This group analysis verifies the knowledge of cherubism previously based on cumulative reviews of findings in single-family and case reports. PMID- 11113825 TI - What should we be doing for children with peanut allergy? PMID- 11113826 TI - Celiac disease and Down syndrome. PMID- 11113827 TI - Promises and pitfalls in the evaluation of pediatric asthma scores. PMID- 11113828 TI - Confusion over consent. PMID- 11113829 TI - The natural history of peanut allergy in young children and its association with serum peanut-specific IgE. AB - OBJECTIVES: To observe the nature and frequency of adverse reactions caused by accidental peanut exposure in young children with clinical peanut hypersensitivity and to determine the value of serum peanut-specific IgE levels during follow-up. STUDY DESIGN: Eighty-three children with clinical peanut hypersensitivity diagnosed before their fourth birthdays were contacted yearly to track adverse peanut reactions. Serum peanut-specific IgE levels were determined in 51 of 83 subjects. RESULTS: Fifty-eight percent (31/53) of subjects followed up for 5 years experienced adverse reactions from accidental peanut exposure. Regardless of the nature of their initial reaction, the majority with subsequent reactions (52%, 31/60) experienced potentially life-threatening symptoms. The group with isolated skin symptoms (11/51, 22%) had lower serum peanut-specific IgE levels than the group with respiratory and/or gastrointestinal symptoms (40/51, 78%) (median: 1.25 kU(A)/L vs 11. 65 kU(A)/L, P =.004, Wilcoxon rank sums test). Despite this, there was no threshold level below which only skin symptoms appeared to occur. Four selected subjects had negative double-blind placebo controlled food challenge responses to peanuts during follow-up. CONCLUSIONS: The majority of children with clinical peanut hypersensitivity followed up for 5 years will have adverse reactions from accidental peanut exposure. Symptoms experienced during subsequent adverse peanut reactions may not be consistent with symptoms reported during initial reactions. Therefore proper education regarding peanut avoidance and treatment of adverse reactions is necessary in all cases of clinical peanut hypersensitivity. Young children who are allergic to peanuts can lose clinical hypersensitivity. PMID- 11113830 TI - Accuracy and cost-effectiveness of a new strategy to screen for celiac disease in children with Down syndrome. AB - OBJECTIVES: To investigate the best approach to screen for celiac disease (CD) in patients with Down syndrome (DS). STUDY DESIGN: One hundred thirty-seven children with DS were followed up longitudinally. CD screening was offered in 1994, 1996, and 1999 by determination of serum immunoglobulin A-anti-endomysium antibodies (AEA). The HLA-DQA1*0501/DQB1*02 allelic combination known to be strongly positively associated with CD was typed. All IgA-AEA-positive children were given the opportunity to undergo a small bowel biopsy: if villous atrophy was found, the diagnosis of CD was established. RESULTS: CD was diagnosed in 11 (8%) children: 8 in 1994 and 3 in 1996. All of them carried the HLA-DQ alleles associated with CD. The presence of symptoms was not useful in discriminating which children could have CD. CONCLUSIONS: Screening once in a lifetime is not enough to detect CD in patients with DS. We propose a new, accurate, and cost sparing 2-step strategy for screening, based on selection of the individuals with potential CD by HLA-DQ typing and on longitudinal serologic CD screening in this selected group. PMID- 11113831 TI - The Preschool Respiratory Assessment Measure (PRAM): a responsive index of acute asthma severity. AB - OBJECTIVE: To elaborate and validate a Preschool Respiratory Assessment Measure (PRAM) that would accurately reflect the severity of airway obstruction and the response to treatment in young patients with asthma. STUDY DESIGN: A prospective cohort study was performed in 217 children aged 3 to 6 years who presented to a pediatric emergency department with acute asthma. Respiratory resistance measured by forced oscillation served as a gold standard. Children were randomized to either the test group, in which multivariate analyses were performed to elaborate the PRAM, or the validation group, in which the characteristics of the PRAM were tested. RESULTS: For the test group (N = 145), the best multivariate model contained 5 variables: wheezing, air entry, contraction of scalenes, suprasternal retraction, and oxygen saturation. In the validation group (N = 72), the PRAM correlated substantially with the change in resistance (r = 0.58) but modestly with the % predicted resistance measured before (r = 0.22) and after bronchodilation (r = 0.36). A change of 3 (95% CI: 2.2, 3.0) indicated a clinically important change. CONCLUSIONS: PRAM appears to be a responsive but moderately discriminative tool for assessing acute asthma severity. This measure, designed for preschool-aged children, has been validated against a concurrent measure of lung function. PMID- 11113832 TI - A randomized controlled trial of alternative modes of service provision to young children with cerebral palsy in Bangladesh. AB - OBJECTIVE: To compare the efficacy of an outreach program for young children with cerebral palsy with center-based and "minimal intervention" control groups. DESIGN: Randomized controlled trial conducted in a group of 85 children between the ages of 1.5 and 5 years. Urban children were allocated to a daily center based mother-child group or to monthly training of their parents along with a pictorial guidance manual. Rural children were allocated either to parent training or health advice. Outcome measures were changes in children's adaptive skills, maternal stress and adaptation to the child, satisfaction with social support, and knowledge of handling a physically disabled child. RESULTS: Fifty eight children were successfully followed up. The pattern of change in children's adaptive skills was as predicted (ie, least progress in the health advice group). Positive effects of intervention also included increased maternal knowledge and perceived helpfulness of support from formal sources. However, maternal adaptation increased most in the health advice group with minimal intervention. When children had attended a program at least 4 times, their skills improved, and mothers' adaptation did increase. CONCLUSIONS: Outreach training for mothers in Bangladesh can help them to improve the skills of their young children with cerebral palsy and is perceived as helpful. PMID- 11113833 TI - Neurophysiologic evaluation of auditory recognition memory in healthy newborn infants and infants of diabetic mothers. AB - OBJECTIVES: We evaluated the integrity of neural pathways for auditory recognition memory in normal newborn infants (n = 32) and infants of diabetic mothers (IDMs, n = 25). IDMs are at risk for fetal metabolic abnormalities that potentially damage recognition memory pathways. We hypothesized that newborn IDMs would have recognition memory deficits that would be correlated with later cognitive development. STUDY DESIGN: Recognition memory was assessed with event related potentials (ERPs). Neonatal ERPs elicited by the maternal voice were compared with those elicited by a stranger's voice. The Bayley Scales of Infant Development were administered at 1 year of age. RESULTS: Infants in both the control and IDM groups demonstrated recognition of the maternal voice, but their ERP patterns differed. Both groups demonstrated increased amplitude and latency for the "P2" peak elicited by the maternal voice compared with the stranger's voice. In the control group the stranger's voice also elicited a negative slow wave, which was attenuated in the IDMs. The negative slow wave correlated significantly with the 1-year Mental Developmental Index. CONCLUSIONS: The presence of a specific neonatal ERP pattern indicated better 1-year cognitive development in infants in the control and IDM groups. ERPs from IDMs demonstrated subtle evidence of recognition memory impairments. PMID- 11113834 TI - Reduced serum amino acid concentrations in infants with necrotizing enterocolitis. AB - OBJECTIVE: To determine whether premature infants who have necrotizing enterocolitis (NEC) have deficiencies in glutamine (GLN) and arginine (ARG), which are essential to intestinal integrity. STUDY DESIGN: A 4-month prospective cohort study of serum amino acid and urea levels in premature infants was done. Serum amino acid and urea levels were measured by high-pressure liquid chromatography and enzymatic methods, respectively, on samples obtained on days of life 3, 7, 14, and 21. RESULTS: Infants in the control (n = 32) and NEC groups (n = 13) were comparable for birth weight, gestational age, and Apgar scores. NEC began on mean day of life 14.5 (95% CI, day of life 11 to 18). Median values of GLN were 37% to 57% lower in the NEC group on days 7, 14, and 21 compared with those in the control group (P <.05). On days 7 and 14, median values of ARG, GLN, alanine, lysine, ornithine, and threonine were decreased 36% to 67% (P <.05) in the NEC group. Total nonessential amino and total essential amino acids were 35% to 50% lower in the NEC group on days 7 and 14 (P <.05). Infants in the NEC group had significant reductions in GLN and ARG 7 days before the onset of NEC. CONCLUSIONS: Infants who have NEC have selective amino acid deficiencies including reduced levels of GLN and ARG that may predispose to the illness. PMID- 11113835 TI - Oral tacrolimus treatment of severe colitis in children. AB - OBJECTIVE: To evaluate the efficacy of oral tacrolimus as an induction agent in steroid-refractory severe colitis. STUDY DESIGN: Open-label, multicenter trial of oral tacrolimus in patients with severe colitis. Patients not responding to conventional therapy received tacrolimus, 0.1 mg/kg/dose given twice a day, and the dosage was adjusted to achieve blood levels between 10 and 15 ng/mL. Response was defined as improvement in a number of clinical parameters (including abdominal pain, diarrhea, rectal bleeding, and cessation of transfusions). Patients who responded by 14 days continued to receive tacrolimus, and 6 mercaptopurine or azathioprine was added as a steroid-sparing agent 4 to 6 weeks after the tacrolimus was instituted. RESULTS: Fourteen patients were enrolled in the study. One patient elected to withdraw after 48 hours. Of the 13 remaining, 9 (69%) responded and were discharged. Tacrolimus was continued for 2 to 3 months in the responders, except for 1 patient who was given tacrolimus for 11 months. After 1 year of follow-up, only 5 (38%) patients were receiving maintenance therapy; the other 4 responders had undergone colectomy. CONCLUSION: Although tacrolimus is effective induction therapy for severe ulcerative or Crohn's colitis, fewer than 50% of patients treated will successfully achieve a long-term remission. PMID- 11113836 TI - Omeprazole for treatment of chronic erosive esophagitis in children: a multicenter study of efficacy, safety, tolerability and dose requirements. International Pediatric Omeprazole Study Group. AB - OBJECTIVES: To determine the efficacy, safety, and tolerability of omeprazole in children and to determine the doses required to heal chronic, severe esophagitis. STUDY DESIGN: Open multicenter study in children aged 1 to 16 years with erosive reflux esophagitis. The healing dose of omeprazole used was that with which the duration of acid reflux was <6% of a 24-hour intraesophageal pH study. Follow-up endoscopy was performed after 3 months of treatment with the healing dose. RESULTS: At entry, two thirds of 57 patients who completed the study had esophagitis grade 3 or 4 (scale 0-4); some 50% had neurologic impairment or repaired esophageal atresia. Of the 57 patients, 54 healed; 3 did not heal and left the study, and 3 healed with a second course. Doses required for healing were 0.7 to 3.5 mg/kg/d: 0.7 mg/kg/d in 44% of patients and 1.4 mg/kg/d in another 28%. Healing dose correlated with grade of esophagitis but not with age or underlying disease. Reflux symptoms improved dramatically in almost all of the 57 patients, including the unhealed patients. CONCLUSIONS: Omeprazole is well tolerated, highly effective, and safe for treatment of erosive esophagitis and symptoms of gastroesophageal reflux in children, including children in whom antireflux surgery or other medical therapy has failed. On a per-kilogram basis, the doses of omeprazole required to heal erosive esophagitis are much greater than those required for adults. PMID- 11113837 TI - Lack of benefit of laxatives as adjunctive therapy for functional nonretentive fecal soiling in children. AB - OBJECTIVES: To determine whether the combination of laxative treatment and biofeedback therapy (BF) is more effective for management of functional nonretentive fecal soiling than biofeedback therapy alone. STUDY DESIGN: In a prospective nonblinded study, 48 children were randomized in 2 groups: treatment with oral laxatives (LAX) and 5 sessions of BF (BF + LAX) or 5 sessions of BF alone (BF) during a treatment intervention period of 7 weeks. Biofeedback was performed with perfused manometry catheters and rectal balloon distension. Training focused on awareness of balloon distension and instruction in correct defecation dynamics. Successful treatment was defined as <1 encopresis episode per 2 weeks. RESULTS: At the end of the intervention period, the number of encopresis episodes was significantly decreased in both groups: from 7 (2 to 24) to 2 (0 to 17) in the BF group and from 7 (3 to 25) to 2 (0 to 14) in the BF + LAX group. However, children given BF alone had significantly higher success rates than children treated with BF and additional oral laxatives (44% to 11%). CONCLUSIONS: There is no additional effect of laxative treatment in functional nonretentive fecal soiling. Children treated with BF in combination with laxatives showed a significantly lower success percentage compared with those treated with BF alone. These results suggest that children with functional nonretentive fecal soiling should be treated differently from children with constipation and encopresis. PMID- 11113838 TI - Daytime urinary incontinence in primary school children: a population-based survey. AB - OBJECTIVES: To determine the prevalence and severity of, and risk factors for, daytime urinary incontinence in children starting primary school. DESIGN AND SETTING: Population-based cross-sectional survey of new entrant primary school children in Sydney, Australia. METHODS: A random cluster sample of 2020 primary school children was surveyed by using a daytime incontinence questionnaire with known substantial repeatability (mean kappa = 0.70). RESULTS: The questionnaire was returned for 1419 (70%) children with a mean age of 5.9 years; 16.5% of children had experienced one or more episodes of wetting in the last 6 months (mild), 2.0% had wet twice or more per week (moderate), and 0.7% were wet every day (severe) (overall prevalence of 19.2%). On multivariate analysis, recent emotional stress (odds ratio 5.7), a history of daytime wetting along the paternal line (odds ratio 9.3), and a history of wetting among male siblings (odds ratio 5.3) were independent risk factors for moderate to severe daytime wetting. Expressed as population attributable risk, 59% and 28% of moderate severe and mild daytime wetting, respectively, can be attributed to these 3 factors. Only 16% of families with affected children had sought medical help. CONCLUSIONS: Daytime urinary incontinence in the first year of primary school is more common than previously reported, and only a small proportion of affected children seek medical help. Emotional stress and family history are likely to be major causal factors. PMID- 11113839 TI - Do all girls with apparent idiopathic precocious puberty require gonadotropin releasing hormone agonist treatment? AB - OBJECTIVE: To evaluate prospectively pubertal and predicted adult height progression until final height (FH) or near FH in girls with apparent idiopathic precocious puberty who were not treated. STUDY DESIGN: The decision not to treat at the time of initial evaluation was based on evidence of slowly progressive puberty as shown by bone age (BA) advancement <2 years above the chronologic age, whatever the hypothalamic pituitary ovarian axis activation, or no evidence of hypothalamic pituitary ovarian axis activation, whatever the BA advancement. During follow-up, patients who showed a significant decrease in predicted FH were treated with gonadotropin-releasing hormone agonist. RESULTS: Twenty-six girls with idiopathic precocious puberty were studied at a mean chronologic age of 7.4 +/- 0.9 years during a follow-up period of 6.6 +/- 2.2 years until FH or near FH. During the first 2 years of follow-up, most of the patients (group 1, n = 17; 65% of the cases) showed no substantial changes in predicted FH. They never required treatment, and menarche occurred at a mean chronologic age of 11.9 +/- 0.6 years. Their mean FH (or near FH) at 160.7 +/- 5.7 cm was close to their target height (161.3 +/- 4.7 cm). On the other hand, after a mean follow-up period of 1.4 +/- 0.8 years, 9 patients (group 2) had acceleration of bone maturation and deterioration of their predicted FH (from 162.1 +/- 6. 2 cm to 155.3 +/- 5.6 cm; P <.01), which was at that time significantly lower than their target height (P <.05) (mean target height = 159.8 +/- 4.6 cm). They received a gonadotropin releasing hormone agonist for 2.1 +/- 0.7 years, resulting in a restoration of growth prognosis (mean FH or near FH = 160.2 +/- 6.7 cm). CONCLUSIONS: This study demonstrates that not all patients with apparent idiopathic precocious puberty require medical treatment, notably when there is no evidence of hypothalamo pituitary ovarian activation or no significantly advanced BA to impair height potential. Most show a slowly progressing puberty. However, careful follow-up of these patients is necessary up to at least 9 years of age, because until then height prediction may deteriorate, necessitating gonadotropin-releasing hormone agonist treatment in one third of the cases. PMID- 11113840 TI - Insulin-like growth factors and bone mineral density in African American and White girls. AB - OBJECTIVES: African American children have greater bone mineral density (BMD) and bone mineral content (BMC) than white children. We examined the hypothesis that differences in insulin-like growth factors (IGFs) are important determinants of BMD during childhood. METHODS: We measured IGFs and IGF binding proteins in 59 African American and 59 white girls matched for age, body mass index, socioeconomic status, and pubertal stage. BMD and BMC were determined by dual emission x-ray absorptiometry. RESULTS: African American girls had greater total BMD (P <.001), BMC (P <.01), total IGF-1 (P <.001), and free IGF-1 (P <.01) than white girls. IGFBP-1, IGFBP-2, and IGFBP-3 were similar in both groups or lower in African Americans. IGF-1 was positively correlated with IGF-2 in white girls (P =.012) but was negatively correlated with IGF-2 in African Americans (P =.015). IGF-1 and free IGF-1 were positively correlated with BMD/BMC. Multiple regression analyses showed 80% of the variance in BMC could be accounted for by the use of body weight, height, and IGF-1 in the model. When IGF-1 was included as a factor, race did not add to the model's predictive power. CONCLUSION: IGF-1 and free IGF-1 are greater in African American than in white girls and may contribute to the greater BMD of African Americans. PMID- 11113841 TI - Risk factors for premature ovarian failure in females with galactosemia. AB - The risk for premature ovarian failure (POF) in females with galactosemia can be predicted by analyzing 3 areas of risk pathology: the patient's molecular genotype for galactose-1-phosphate uridyltransferase (GALT), alternate pathways for galactose metabolism, and the patient's environment at diagnosis and during treatment. STUDY DESIGN: Retrospective cross-sectional information was collected on 53 females with classic galactosemia, and their ovarian function was analyzed by determination of serum follicle-stimulating hormone and luteinizing hormone levels and by clinical observation. The associations were analyzed between POF and the mutations in GALT, the highest erythrocyte galactose-1-phosphate (Gal-1 P) level at diagnosis, the age at which dietary treatment was initiated, mean erythrocyte Gal-1-P level during treatment, and whole-body carbon 13-labeled galactose oxidation to (13)CO(2). RESULTS: The most prevalent mutation, Q188R, had a significant effect of genotype category (Q188R/Q188R, Q188R/Other, Other/Other) on POF (P =.04, Fisher exact test and an odds ratio of 8.3). Mean erythrocyte Gal-1-P level during treatment was a significant risk factor for POF (P =.04). Also, all patients studied with less than 5% total body oxidation of galactose to (13)CO(2) had POF, whereas those with more than 5% did not have POF (P =.008, Fisher exact test). CONCLUSION: The development of POF in females with galactosemia is more likely if the patient's genotype is Q188R/Q188R, if the mean erythrocyte Gal-1-P is >3.5 mg/dL during therapy, and if the recovery of (13)CO(2) from whole-body (13)C-galactose oxidation is reduced below 5% of administered (13)C-galactose. PMID- 11113842 TI - The importance of language and culture in pediatric care: case studies from the Latino community. AB - BACKGROUND: Few studies have examined culture's effect on pediatric care. OBJECTIVES: To analyze 3 cases illustrating the importance of culture in pediatrics. METHODS: Case analysis with a cultural competency model. RESULTS: No interpreter was available for the parents of a 3-year-old brought to the emergency department because of abdominal pain; she was discharged twice, returned with an acute abdomen, and was hospitalized for treatment of appendiceal rupture and peritonitis. A 2-year-old fractured her clavicle in a fall but was placed in social services' custody because of a pediatrician's misinterpretation. Parents of a ventilation-dependent 2-week-old with encephalopathy, seizures, and renal failure were unaware of the infant's poor prognosis, despite use of an interpreter. CONCLUSIONS: Culture can have a significant impact on pediatric care; use of a simple model can ensure that pediatricians provide culturally competent care. PMID- 11113843 TI - A simplified cyclic adenosine monophosphate-mediated sweat rate test for quantitative measure of cystic fibrosis transmembrane regulator (CFTR) function. AB - OBJECTIVE: Sweat production is stimulated by both cholinergic and beta-adrenergic pathways in the sweat gland secretory coil. beta-Adrenergic pathway-mediated sweating is absent in cystic fibrosis (CF) because cyclic adenosine monophosphate (cAMP)-mediated chloride transport through the cystic fibrosis transmembrane regulator (CFTR) is disrupted. We report the development of a rapid, reproducible, macroscopic, and quantitative methodology to test the hypothesis that beta-adrenergic sweat rate discriminates among 3 different CFTR phenotypes CF, heterozygote CF carriers, and non-CF. STUDY DESIGN: Intradermal injection of a mixture of 50 micromol/L isoproterenol, 5 mmol/L aminophylline (to potentiate the beta-adrenergic stimulation), and 140 micromol/L atropine (to block potential cholinergic stimulation) in lactated Ringer's solution was performed in duplicate on one forearm. A single injection of 0.5 mmol/L methacholine to stimulate sweat production by the cholinergic pathway was performed on the other forearm. Sweat rate was determined as the amount of sweat collected on filter paper over 20 minutes. RESULTS AND CONCLUSIONS: Median cAMP-mediated sweat rates were 1.45 mg/20 min (CF, n = 29), 2.55 mg/20 min (CF heterozygote carriers, n = 30), and 3.65 mg/20 min (non-CF, n = 30) and were significantly different in all 3 groups (P =.0001, Kruskal-Wallis test). Methacholine-stimulated sweat rates were similar for all 3 groups. The cAMP-mediated sweat rate test may be a useful endpoint for studies of new agents to increase the function of CFTR. PMID- 11113844 TI - The burden of influenza illness in children with asthma and other chronic medical conditions. AB - OBJECTIVE: Although influenza immunization is recommended for children with high risk medical conditions, the majority of such children do not receive influenza vaccine. This study was designed to measure the burden of influenza among children with asthma and other chronic medical conditions. STUDY DESIGN: We performed a retrospective cohort study of children younger than 15 years with medically treated asthma or other chronic medical conditions enrolled in the Tennessee Medicaid program from 1973 to 1993. We determined rates of hospitalization for acute cardiopulmonary disease, outpatient visits, and antibiotic courses throughout the year. Annual differences between event rates when influenza virus was circulating and event rates during winter months when there was no influenza in the community were used to calculate influenza attributable morbidity. RESULTS: Influenza accounted for an average of 19, 8, and 2 excess hospitalizations for cardiopulmonary disease yearly per 1000 high-risk children aged <1 year, 1 to <3 years, and 3 to <15 years, respectively. For every 1000 children, an estimated 120 to 200 outpatient visits and 65 to 140 antibiotic courses were attributable to influenza annually. CONCLUSIONS: Children younger than 15 years with asthma and other chronic medical conditions experience substantial morbidity requiring inpatient and outpatient care during influenza season. More effective targeting of this population for annual influenza immunization is warranted. PMID- 11113845 TI - Rates of hospitalization for respiratory syncytial virus infection among children in medicaid. AB - OBJECTIVE: To determine rates of hospitalization associated with respiratory syncytial virus (RSV) infection among children with and without specific medical conditions. STUDY DESIGN: Retrospective cohort study of all children <3 years old enrolled in the Tennessee Medicaid program from July 1989 through June 1993 (248,652 child-years). RESULTS: During the first year of life, the estimated number of RSV hospitalizations per 1000 children was 388 for those with bronchopulmonary dysplasia, 92 for those with congenital heart disease, 70 for children born at < or = 28 weeks' gestation, 66 for those born at 29 to <33 weeks, 57 for those born at 33 to <36 weeks, and 30 for children born at term with no underlying medical condition. In the second year of life, children with bronchopulmonary dysplasia had an estimated 73 RSV hospitalizations per 1000 children, whereas those with congenital heart disease had 18 and those with prematurity 16 per 1000. Overall, 53% of RSV hospitalizations occurred in healthy children born at term. CONCLUSIONS: Children with bronchopulmonary dysplasia have high rates of RSV hospitalization until 24 months of age. In contrast, after the first year of life, children with congenital heart disease or prematurity have rates no higher than that of children at low risk who are <12 months old. PMID- 11113846 TI - The natural history of varicella embryopathy: a 25-year follow-up. AB - A patient with clinically and immunologically proven varicella embryopathy achieved substantial recovery after initial severe developmental delay and manages well with her residual physical disabilities in adulthood. PMID- 11113847 TI - Coronary flow abnormalities in neonates with aortic stenosis. AB - Left coronary artery flow velocities in neonates were determined noninvasively with transthoracic pulsed wave Doppler ultrasonography. In normal subjects (n = 30) there was diastolic flow predominance with median (range) peak flow velocity in diastole, 23.8 cm/s (12.7 to 51.3 cm/s), and median peak flow velocity in systole, 12.7 cm/s (7.8 to 35.0 cm/s). In 3 neonates with severe aortic stenosis, retrograde left coronary flow throughout systole was observed before surgery. In these patients there was forward systolic flow 4 to 8 days after successful surgical valvulotomy was performed. PMID- 11113848 TI - Presymptomatic diagnosis of X-linked adrenal hypoplasia congenita by analysis of DAX1. AB - A novel DAX1 mutation (L381H) was discovered in the asymptomatic 8-month-old brother of a boy with primary adrenal failure. The infant had impaired adrenal reserve despite normal basal adrenal steroid concentrations. This case highlights the value of genetic testing in children at risk of the development of X-linked adrenal hypoplasia congenita before the onset of a potentially life-threatening adrenal crisis. PMID- 11113849 TI - Successful bone marrow transplantation in a patient with Schimke immuno-osseous dysplasia. AB - Early death in Schimke immuno-osseous dysplasia often results from renal failure and/or cell-mediated immunodeficiency. Kidney transplants have improved renal function, but effective therapy for the immunodeficiency has not yet been reported. We describe markedly improved marrow function 2 years after bone marrow transplantation in a boy with Schimke immunoosseous dysplasia. PMID- 11113850 TI - Peripheral harlequin-like thermal imbalance after Wilms' tumor. PMID- 11113851 TI - Poor imaging technique produces poor results: again! PMID- 11113852 TI - Reply PMID- 11113854 TI - Reply PMID- 11113853 TI - Exhaled carbon monoxide in asthma. PMID- 11113855 TI - Why young children are resistant to acetaminophen poisoning. PMID- 11113856 TI - Reply PMID- 11113857 TI - The role of cytochrome P450 in tumour development and progression and its potential in therapy. AB - The cytochromes P450 (P450) are a large group of constitutive and inducible haem containing enzymes, which have a central role in the oxidative metabolism of a diverse range of xenobiotics. Many P450 substrates are carcinogenic, while other substrates are anti-cancer drugs; the P450s thus have various potentially important roles in tumour biology. Several P450 genes are polymorphic and are associated with the increased risk of cancer development in specific tissues. Individual P450s, especially CYP1B1, are overexpressed in different types of tumours. The increased expression of P450s in tumours is highly significant and is important for understanding tumour development and progression. The tumour specific expression of P450 provides the basis for the development of novel diagnostic and therapeutic strategies. PMID- 11113858 TI - Expression of CD44v6 but not E-cadherin or beta-catenin influences prognosis in primary pulmonary adenocarcinoma. AB - Primary pulmonary adenocarcinoma was studied, looking for relationships between the expression of cell adhesion molecules (CAMs) E-cadherin, beta-catenin and CD44v6, and clinicopathological tumour parameters and patient post-operative survival. Formalin-fixed, paraffin-embedded tissue from 120 primary lung adenocarcinomas, including 23 poorly differentiated tumours, 17 of probable bronchial origin, and 29 with a prominent bronchioloalveolar pattern, together with nodal metastatic tumour from 34 of these patients was stained using monoclonal antibodies and immunohistochemistry. Sections were scored either high level (>10% cells positive) or low level (<10% positive). High level expression of CD44v6 was retained in 28.4% (34/120) of tumours, while high levels of E cadherin (57.5%, 69/120) and beta-catenin (80. 8%, 97/120) were more frequent. For all CAMs, staining levels did not correlate with nodal status, stage or tumour type. The apical or basal staining seen in normal bronchial and alveolar epithelium was often seen in papillary, glandular, and bronchioloalveolar areas of tumour, while solid invasive tumour more often showed pericellular staining. When the staining for each CAM in 34 nodal metastases was compared with that in the corresponding primary tumour, a high degree of concordance was found, with no tendency for metastases to show less staining than the primary tumour. Expression of E-cadherin and beta-catenin in the primary tumour had no influence on post operative survival, but patients whose tumours had low level CD44v6 expression had a poorer post-operative survival than those with high levels of CD44v6 (p=0.0014 for all patients, p=0.0012 for stage I patients only). In primary pulmonary adenocarcinoma, the levels of expression of E-cadherin, beta-catenin, and CD44v6 are not associated with lymph node metastases or tumour stage but the staining pattern is associated with tumour morphology. Low levels of CD44v6 expression predict a poor post-operative survival, independently of stage, while there is no such relationship with the expression of E-cadherin or beta-catenin. PMID- 11113859 TI - Expression of the E-cadherin/catenin (alpha-, beta-, and gamma-) complex correlates with the macroscopic appearance of early gastric cancer. AB - E-cadherin and its associated cytoplasmic proteins, alpha-, beta-, and gamma catenins, play an essential role in the control of epithelial differentiation. We have previously shown that loss or down-regulation of E-cadherin/catenin correlates with poor survival in advanced gastric adenocarcinoma. The aim of this study was to assess the expression of E-cadherin and catenins in early gastric cancers (EGCs). Immunohistochemical staining for E-cadherin and alpha-, beta-, and gamma-catenins was performed on 41 paraffin-embedded gastrectomy specimens of EGC using an indirect immunoperoxidase technique. The pattern of expression and cellular localization of the E-cadherin/catenin complex in tumour cells were correlated with the macroscopic appearance of the tumour according to the Japanese Endoscopic Society classification. The tumours were classified as follows: three type I (protruding) and 38 type II (superficial), of which ten were type IIa (elevated), one was type IIb (flat), and 27 were type IIc (depressed). E-cadherin and alpha-, beta-, and gamma-catenins were expressed at the cell-cell junctions in normal mucosa. Forty out of 41 tumours showed abnormal expression (loss of membranous immunoreactivity and/or nuclear staining) of at least one component of the E-cadherin catenin complex. Loss of E-cadherin immunoreactivity was more frequently seen in type IIb (1/1, 100%) and type IIc (27/27, 100%) than in type I (1/3, 33%) and type IIa (1/10, 10%) (p<0.01). Abnormal expression of E-cadherin and alpha-catenin was more frequently seen in diffuse-type than in intestinal type tumours (p<0.05). Abnormal immunoreactivity of beta- and gamma-catenin, including nuclear localization, was observed in 34% and 7.3% of tumours, respectively, but there was no significant correlation with tumour type or endoscopic appearance. In conclusion, abnormal expression of the E cadherin/catenin complex occurs in EGC and seems to correlate with macroscopic appearances. PMID- 11113860 TI - Specific patterns of chromosomal abnormalities are associated with RER status in sporadic colorectal cancer. AB - Current opinion of the genetic events driving colorectal tumourigenesis focuses on genomic instability. At least two apparently independent mechanisms are recognized, microsatellite instability and chromosomal instability. The genetic defects underlying each type of instability are only partially understood and controversy remains as to the role of p53 in the generation of chromosomal defects in colorectal cancer. This study sought to clarify the relationships between chromosomal abnormalities and defects of both p53 and mismatch repair. Extensive chromosomal analysis was undertaken, using flow cytometry and comparative genomic hybridization, of a series of sporadic colorectal cancers which had been grown to early passage as subcutaneous xenografts in SCID mice. Overall levels of chromosomal defects were observed to be low in RER+ cancers compared with RER- and distinctive patterns of chromosomal anomalies were found to be associated with both the RER+ and RER- phenotype. No particular level or pattern of chromosomal anomalies appeared to be associated with p53 status, supporting recent observations that abnormal p53 function is not sufficient to cause chromosomal anomalies in colorectal tumours. PMID- 11113861 TI - Validation of a model of colon cancer progression. AB - A unique feature of SW480 and SW620 colon carcinoma cell lines is that they are derived from primary and secondary tumours resected from a single patient. As such, they may represent a valuable resource for examining genetic changes late in colon cancer progression. In order to verify this, both cell lines have been characterized to determine whether phenotypic differences have been retained despite long-term cell culture in vitro. The primary tumour-derived SW480 cells have an epithelioid morphology in vitro, while metastasis-derived SW620 cells have a fibroblast-like appearance. Xenografts of SW480 cells form gland-like structures in vivo, while SW620 xenografts form solid sheets of tumour cells. SW620 cells have a higher BrdU labelling index than SW480 cells, and are more highly tumourigenic and metastatic. Furthermore, SW620 cells show less susceptibility to apoptosis induction by TNFalpha and anti-Fas monoclonal. Findings from these investigations therefore indicate that SW480 and SW620 cell lines do show appropriate phenotypic differences and represent an interesting model for studying the genetic events in the late stages of colon cancer progression. PMID- 11113862 TI - Increased expression of tissue inhibitor of metalloproteinases type 1 (TIMP-1) in a more tumourigenic colon cancer cell line. AB - Genetic changes occurring in the late stages of colonic tumour progression have received much less attention than those occurring in the early stages. As described in the accompanying paper, SW480 and SW620 cell lines provide a useful model for studying the advanced stages of progression for colon cancer. Comparison of the two cell lines by differential display reveals that SW620 cells express lower levels of the CC3 tumour suppressor gene and also lower levels of the tissue inhibitor of metalloproteinases-3 (TIMP-3) gene. Northern blot analysis for TIMP-3 confirms this finding and shows a similar difference in the expression of TIMP-2, which seems logical since TIMPs inhibit enzymes that play a role in tumour invasion. For this reason, it was surprising to find that TIMP-1 messenger RNA expression is markedly increased in SW620 cells. Consistent with this finding, western blot analysis shows a ten-fold increase in TIMP-1 protein secretion by SW620 cells. It is noteworthy that high TIMP-1 expression is associated with poor prognosis in colorectal cancer. This association between TIMP-1 expression and tumour progression may be related to additional growth factor-like effects described for TIMP-1 in some systems. PMID- 11113864 TI - Significantly different bcl-2 expression profiles in gastric and non-gastric primary extranodal high-grade B-cell lymphomas. AB - Fifty-five cases of primary extranodal high-grade B-cell non-Hodgkin's lymphoma were investigated for bcl-2 and p53 protein expression as well as for t(14;18) translocations and p53 mutations. Phenotypic and genotypic profiles were compared between tumours of gastric (27 cases) and non-gastric (28 cases) origin. bcl-2 protein expression was significantly lower in gastric (11/27) than in non-gastric (28/28) lymphomas (p<0.0001), while nuclear p53 protein expression did not differ significantly between these two groups. In the stomach, there were no significant differences in either bcl-2 or p53 expression profiles between high-grade lymphomas with (n=14) and without (n=13) evidence of a low-grade component of MALT type. However, secondary high-grade lymphomas showed a significant down regulation of bcl-2 protein (p<0.0001) and, conversely, an up-regulation of p53 protein (p<0.0001) as compared with their low-grade tumour components. In extranodal high-grade B-cell lymphomas, bcl-2 protein expression was not associated with t(14;18) translocation. Only one gastric lymphoma had a p53 point mutation with potential alteration of the amino acid sequence. These findings indicate that primary gastric high-grade B-cell lymphomas are immunohistologically distinct from primary extranodal high-grade B-cell lymphomas of an origin other than in the stomach. PMID- 11113863 TI - E-cadherin and alpha-, beta-, and gamma-catenin protein expression is up regulated in ovarian carcinoma cells in serous effusions. AB - The aims of this study were firstly, to investigate the expression of E-cadherin complex proteins in ovarian carcinoma cells in serous effusions and in primary and metastatic lesions; and secondly to study the value of these four proteins and calretinin, a mesothelial marker, in the differential diagnosis of ovarian carcinoma cells from reactive mesothelial cells in effusions. Sixty-seven malignant effusions and 97 corresponding primary (n=36) and metastatic (n=61) lesions were immunohistochemically stained for E-cadherin and alpha-, beta-, and gamma-catenin. Staining extent and intensity were scored. Effusion specimens were additionally analysed for calretinin immunoreactivity. Membrane immunoreactivity for E-cadherin and alpha-, beta-, and gamma-catenin was detected on carcinoma cells in the majority of the effusions, but rarely on reactive mesothelial cells (p<0.001 for all markers). Calretinin immunoreactivity was confined to mesothelial cells (p<0.001). An association was seen between E-cadherin and alpha catenin expression, in both effusions and solid tumours, and for beta-catenin in solid tumours (range p<0. 001 to p=0.014). Up-regulation of all four cadherin complex proteins was seen in carcinoma cells in effusions, when compared with corresponding primary tumours (range p<0.001 to p=0.028). As with effusions, metastatic lesions showed up-regulation of alpha-, beta-, and gamma-catenin when compared with primary carcinomas (p=0.002-0. 015). Carcinoma cells in effusions showed in addition elevated levels of E-cadherin when compared with metastatic lesions (p<0.001). Staining results in effusions showed no association with effusion site, tumour type or histological grade. Immunoblotting on 29 malignant effusions confirmed the presence of all four proteins in the majority of samples and co-precipitation of E-cadherin and beta-catenin was seen in ten specimens examined. E-cadherin complex proteins are widely expressed in ovarian carcinoma cells. Together with calretinin, they form a powerful battery of markers for the cytological diagnosis of carcinoma cells in effusions. The up-regulation of E cadherin complex proteins in serous effusions and metastatic lesions may mark an early metastatic phenotype and possibly mediates survival of tumour cells at these sites through the inhibition of apoptosis. PMID- 11113865 TI - Molecular analysis reveals somatically mutated and unmutated clonal and oligoclonal B cells in T-cell-rich B-cell lymphoma. AB - This paper describes the immunohistology and molecular genetics of 18 cases of T cell-rich B-cell lymphoma (TCRBL). In all cases, the large B cells stained strongly for CD20, with more variable expression of CD79a, and were negative for CD30 and CD15. The majority of T cells were predominantly positive for TIA-1 and negative for CD57; a large population of histiocytes was present in all cases. Epstein-Barr virus (EBV)-coded RNA (EBER) was found in B blasts from four cases and in one case was present among the background lymphoid cells. IgH PCR products were generated in 16/18 cases and revealed clonal, oligoclonal and polyclonal PCR products in 12, two and two cases, respectively. In addition, TCRG clonal gene rearrangements were identified in two cases. TCRB gene rearrangements were polyclonal. Sequence analysis of seven cases with clonal/oligoclonal IgH gene rearrangements revealed functional sequences with predominant V(H)3 gene usage associated with various D genes and J(H)4 or J(H)6 gene segments. Four cases displayed varying degrees of replacement and silent mutations (1.8-21%), with one case exhibiting intraclonal heterogeneity; the distribution of mutations was indicative of antigen selection in three cases. The remaining three cases, including two cases with functional oligoclonal IgH rearrangements, harboured unmutated V region genes. The EBV-positive cases were associated with clonal, oligoclonal and polyclonal PCR products and with mutated and germline clonal sequences. These data indicate that TCRBL may be a heterogeneous entity associated with clonal and oligoclonal B cells derived from both germinal centre and naive B cells. PMID- 11113866 TI - Clone-specific PCR reveals wide dissemination of gastric MALT lymphoma to the gastric mucosa. AB - The development of low-grade gastric mucosa-associated lymphoid tissue (MALT) lymphoma is closely associated with Helicobacter pylori infection. Despite its indolent clinical course and prolonged localization to the site of origin, the lymphoma frequently presents with multifocal lesions. However, the true extent of tumour involvement in the gastric mucosa is unclear, since reactive appearing lymphocytic infiltrates are always present and could contain tumour cells that are not readily identifiable on cytological grounds. Gastrectomy specimens of four MALT lymphoma cases were studied by microdissection and clone-specific polymerase chain reaction (CS-PCR) and of a further case with t(1;14)(p22;q32) by immunohistochemistry for BCL10 protein, which acted as a tumour marker for tumour cells carrying the translocation. CS-PCR revealed that tumour cells were commonly present in histologically non-lymphomatous lymphocytic infiltrates microdissected from areas well separated from tumour lesions. Tumour cells were also frequently found in infiltrates microdissected from the resection margins. These findings were reinforced by direct identification of tumour cells, as recognized by strong BCL10 nuclear staining, in non-lymphomatous lymphocytic infiltrates in the case with t(1;14)(p22;q32). The results show that gastric MALT lymphoma disseminates widely within the gastric mucosa without necessarily forming diagnostic lesions. PMID- 11113867 TI - Decreased synthesis and expression of TGF-beta1, beta2, and beta3 in epithelium of HPV 16-positive cervical precancer: a study by microdissection, quantitative RT-PCR, and immunocytochemistry. AB - Cervical carcinogenesis is a multistep process initiated by 'high-risk' human papillomaviruses (HR-HPVs), most commonly HPV 16. Transforming growth factor-beta (TGF-beta) inhibits epithelial proliferation and down-regulates transcription of E6/E7 genes of HPV. Altered TGF-beta expression may be important in carcinogenesis. Quantitative RT-PCR was used to investigate TGF-beta1, beta2, and beta3 mRNA levels in nine specimens of normal cervix and 15 cervical precancers (eight HPV-positive, including five HPV 16-positive). Immunocytochemical expression of TGF-beta1, beta2, and beta3 was examined in cervical intraepithelial neoplasia (CIN) positive for HPV 16 (26), and in HPV-negative, normal ectocervical epithelium (9); reserve cell hyperplasia (12); and immature (7) and mature (15) squamous metaplasia. The intensity of staining for TGF-beta1 was measured using grey-scale image analysis. Microdissection was used to investigate epithelial and stromal (excluding crypts) levels of TGF-beta1 mRNA in HPV 16-positive cervical precancer. Normal cervix, including reserve cells and immature and mature metaplasia, showed strong immunocytochemical expression of all TGF-beta isoforms. Expression was decreased in the basal third of the epithelium in CIN 1, in the basal and middle thirds in CIN 2, and in all layers in CIN 3. Quantitative analysis of TGF-beta1 expression showed that the changes in CIN compared with normal ectocervix and mature metaplasia were statistically highly significant (p<0.001, ANOVA). TGF-beta1, beta2, and beta3 mRNA levels showed a significant decrease only in the five HPV 16-positive CIN samples when compared with normal (p=0. 0034, 0.0033, and 0.029, respectively). TGF-beta mRNA levels in HPV 16-positive epithelium also decreased from normal through low-grade to high-grade precancer. Stromal TGF-beta1 was absent or very low compared with epithelial production and was not altered in HPV 16 precancer. Progressive loss of epithelial TGF-beta expression and synthesis may be important in HPV 16 associated human cervical carcinogenesis. PMID- 11113868 TI - Deletion of the FHIT gene in neoplastic and invasive cervical lesions is related to high-risk HPV infection but is independent of histopathological features. AB - The fragile histidine triad (FHIT) gene encompasses the common chromosomal fragile site FRA3B. Human papilloma virus (HPV), which is the main aetiological agent in cervical cancers, has been found to be able to integrate its genes into the chromosome 3 fragile site of cultured cells, deleting a piece of DNA which includes the FHIT gene. Eighty-six microdissected archival cervical LLETZ biopsies comprising cases of cervical intraepithelial neoplasia (CIN) 1 (n=27), CIN3 (n=30) and microinvasive carcinoma (n=29) were evaluated for HPV infection and FHIT gene loss of heterozygosity (LOH). FHIT gene LOH was detected by polymerase chain reaction (PCR) using fluorescently labelled intragenic microsatellite markers D3S1300 and D3S4103. PCR products were analysed on a semi automated DNA sequencer using Fragment Manager(trade mark) software to determine allele loss. The HPV status of the lesions was determined by PCR using generic and type-specific primers in conjunction with restriction endonuclease digestion. The results were analysed using Epi-Info and SPSS-PC statistical analysis software. Haematoxylin and eosin-stained sections from the 86 cases were profiled for six histopathological features, some of which have been previously shown to be associated with microinvasive cancer. FHIT gene LOH was found in 36% of CIN1 cases, 52% of CIN3 cases and 73% of microinvasive cases (p=0.029). HPV 16 DNA was found in 68% of CIN3 cases and 93% of microinvasive cases (p<0.001). The second most prevalent HPV type found was HPV 31, which was present in only four lesions, three of which had FHIT gene LOH. When FHIT gene LOH was evaluated versus HPV 16 and 31 infection using the chi-square test, a statistically significant relationship was found (p=0.014). FHIT gene LOH was found to be independent of the histopathological features evaluated. The finding of a statistically significant relationship between FHIT gene LOH and oncogenic HPV infection suggests a link between the integration of viral DNA and subsequent gene deletion in the progression of cervical cancer. FHIT gene anomalies may prove to be excellent markers of progression in early uterine cervical cancers. PMID- 11113869 TI - Abnormalities of the transforming growth factor-beta pathway in ocular melanoma. AB - The majority of ocular melanomas occur in the uveal tract. Chemotherapy is generally ineffective and large tumours requiring enucleation have a greater than 50% mortality at 5 years. Monosomy for chromosome 3 is common in uveal melanoma and it is known that there is loss of responsiveness to transforming growth factor beta (TGFbeta) in melanoma cell lines. Since the gene for TGFbeta receptor II (TGFbetaR2) is located on chromosome 3p22, this study investigates the possibility that the TGFbeta pathway, and TGFbetaR2 in particular, might be involved in the pathogenesis of this rare eye tumour. To this end, the expression of molecules in the pathway has been examined by immunocytochemistry (TGFbeta, TGFbetaR2, SMAD2, SMAD3, SMAD4, and p27), backed up by a cell culture assay of TGFbeta-mediated growth suppression, RT-PCR for SMAD4, and loss of heterozygosity (LOH) on 3p22. There was LOH at 3p22 in 6/19 tumours and loss of TGFbetaR2 expression in 10/27 tumours. Immunohistochemistry for SMADs 2, 3, and 4 showed potential loss of signal transduction in 14/27 tumours. The results indicate abnormality of the TGFbeta pathway in 61% of tumours for which unequivocal results were obtained and suggest that abrogation of control of melanocyte growth by the TGFbeta pathway may be important in the formation of uveal melanoma. PMID- 11113870 TI - Epidermal growth factor up-regulates CD44-dependent astrocytoma invasion in vitro. AB - CD44/hyaluronan interactions and epidermal growth factor (EGF) stimulation are both known to enhance tumour invasion in vitro. The frequent amplification of the EGF receptor (EGFR) in high-grade astrocytomas led to the examination of the hypothesis that CD44-dependent astrocytoma invasion is regulated by EGF. It has been shown that human astrocytoma cells express only the standard (haemopoietic) form of CD44 (CD44s) and that EGF up-regulates CD44 mRNA and protein in a time- and dose-dependent (10-100 ng/ml) manner. EGF stimulation did not result in induction of additional splice variants. No EGF-induced increase in CD44s was observed after treatment of cells with the wild-type EGFR tyrosine kinase inhibitor Tyrphostin AG1478 (30 nM). Up-regulation of CD44 by EGF is also prevented by the transcriptional inhibitor actinomycin D (5 microg/ml) and by blocking the MAP kinase (MAPK) pathway using the MEK inibitor U0126 (100 microM). CD44 up-regulation was associated with a 50% increase in invasion through hyaluronan-supplemented Matrigel(trade mark), which was abrogated by ligating CD44 with the specific antibody KM201. These results suggest that increased CD44 expression in response to EGF stimulation plays a significant role in astrocytoma invasion. PMID- 11113871 TI - Changes in gastrointestinal morphology associated with obstructive jaundice. AB - Bacterial translocation has been consistently demonstrated in experimental models of obstructive jaundice. An important factor which promotes this phenomenon is physical injury of the intestinal mucosa. Some previous studies have presented suggestive evidence of this, following bile duct ligation. The aims of this study were to analyse objectively intestinal mucosal morphometric characteristics, to examine for evidence of bacterial translocation, and to assess enterocytes for ultrastructural abnormalities. Adult female Wistar rats were assigned to one of three groups: control (n=8), bile duct ligation (BDL; n=11), or sham operation (n=10). One week later, portal blood, mesenteric lymph nodes, liver, and spleen were harvested and cultured aerobically and anaerobically for evidence of bacterial translocation. Segments of jejunum, ileum, caecum, and large bowel were examined histologically, using light microscopy and morphometrically, using an image analysis system. Electron microscopy was performed on regions of the gastrointestinal tract where significant morphometric alterations had been identified. Significant bacterial translocation was identified following BDL (63. 6% BDL vs. 0% sham vs. 0% control, p<0.01, Fisher's exact test). There was a significant reduction in total mucosal thickness (standard error) [650 microm (23) BDL vs. 731 microm (27) sham vs. 744 microm (95) control] and villous height [451 microm (20) BDL vs. 515 microm (18) sham vs. 559 microm (79) control] in jaundiced animals, compared with sham-operated and control animals (p<0.02, Mann Whitney U-test). Electron microscopy revealed oedematous change associated with mild inflammation, disruption of desmosomes, and the formation of lateral spaces between enterocytes. In addition, enterocytes showed vacuolation of their cytoplasm and mitochondrial swelling. Increased numbers of bacteria appeared to be attached to the mucosa. These data provide evidence of physical disruption of intestinal mucosa in jaundiced animals, most marked in the distal ileum. Significant bacterial translocation occurs following bile duct ligation and this supports the hypothesis of gut barrier dysfunction with obstructive jaundice. PMID- 11113872 TI - Distribution of the interleukin-8 receptors, CXCR1 and CXCR2, in inflamed gut tissue. AB - There is increasing evidence to suggest that the potent neutrophil chemoattractant interleukin-8 (IL-8) has an important role in the pathogenesis of inflammatory bowel disease. IL-8 mediates its actions via two cell surface receptors, CXCR1 and CXCR2. This paper describes the distribution of these IL-8 receptors in the normal gastrointestinal tract and how this is modified in ulcerative colitis (UC). Paraffin-embedded colonic resection specimens were stained with monoclonal antibodies directed against CXCR1 and CXCR2 in ten cases of total UC, 16 cases of appendicitis, and 11 histologically normal sections. A semiquantitative scale of 0-4 was used to assess the proportion and intensity of positively stained cells within certain defined areas of tissue. A comparative assessment was made of the distribution of various cell populations. Dual immunostaining was used to confirm the phenotype of positively staining cells. In the histologically normal colon, the antibody against CXCR1 stained a subpopulation of macrophages deep to the epithelium and germinal centre lymphocytes. A similar pattern of staining was seen in acute appendicitis, with in addition some positively stained neutrophil polymorphs. In UC, there was up regulation of CXCR1, with a striking increase in positively stained macrophages throughout the mucosa and of B and T lymphocytes outside the germinal centre areas. There was also intense up-regulation of CXCR1 expression by the luminal epithelium, reflected in the epithelial staining score (mean+/-SE=1.8+/-0.44 for UC cases, vs. 0.23+/-0.16 for controls and 0.25+/-0.14 for acute appendicitis). CXCR2 was only expressed on a small population of lamina propria mononuclear cells and crypt epithelial cells, with no significant differences observed between the groups. These results suggest that IL-8 may, through CXCR1, have a role beyond neutrophil recruitment in mediating the immune response in UC and that this is not merely a consequence of the acute inflammation seen in UC. PMID- 11113873 TI - Identification of MRP-8 (calgranulin A) as a major responsive protein in chronic periodontitis. AB - The purpose of the study was to analyse how the protein composition of the inflammatory exudate associated with chronic periodontitis differed from the exudate in periodontal health. Gingival crevicular fluid (GCF) was collected from sites with chronic periodontal inflammation and from non-diseased sites in healthy control subjects. Microbore HPLC analysis revealed one major difference in GCF protein profiles between healthy controls and periodontitis patients. The protein enhanced in periodontitis patients was identified as migration inhibitory factor-related protein-8 (MRP-8) by a combination of N-terminal amino acid sequencing, mass spectrometry, and SDS-PAGE. Together, these data demonstrate, for the first time, the presence of monomeric MRP-8 in an inflammatory exudate. Whether monomeric MRP-8 is a unique feature of chronic periodontal inflammation is not yet clear, but the chemotactic properties of this peptide support a functional role for MRP-8 in periodontal inflammation. PMID- 11113874 TI - Validation of endobronchial biopsy specimens for nerve quantitation by computer assisted image analysis. AB - The aim of this study was to assess the validity of endoscopic bronchial biopsy specimens for the quantitation of nerves. To this end, endobronchial biopsy was simulated ex vivo on surgically resected lung specimens and nerve densities were compared in airway smooth muscle of biopsy and surrounding tissue. Specimens were stained immunohistochemically for the general neural marker protein gene product 9.5 (PGP 9.5) and for vasoactive intestinal peptide (VIP), and nerve densities were quantitated using computer-assisted image analysis. Nerve density for total (PGP 9.5-immunoreactive) nerves was slightly higher in biopsies than in corresponding lung tissue, but this difference did not reach statistical significance (p=0.08). There was also no significant difference in the density of VIP-immunoreactive nerves (p=0.60). These findings support the use of endobronchial biopsy specimens to quantitate nerves in asthma and other airway diseases. PMID- 11113875 TI - Multiple tissue core arrays in histopathology research: a validation study. AB - The use of multiple tissue arrays allows the examination of large cohorts of tumour tissue with economies of material and technical resources. It also permits the direct comparison of tissues on the same slide. In the present study, a series of 157 breast cancers was labelled with antibodies which recognize oestrogen (ER) and progesterone (PR) receptors and the staining obtained on whole tissue sections was compared with that from a series of multicore arrays. A highly significant association was found between the staining scores (0-7) obtained from the individual tissue sections and from the multicore arrays, although there was some discordance between the receptor status (positive/negative) of the whole section and the tissue core in 5% of cases for ER and in 6.5% of cases for PR. Multiple tissue cores represent an attractive way of dealing with large cohorts of tumours for research studies, because of the significant reduction in reagents and technical time required and the overall speed with which a study can be completed. A proportion of individual tissue cores were not representative of the diagnostic section, which limits the value of multicore arrays as a tool for patient management. However, the technique provides an efficient way of assessing the potential predictive value of novel proteins in different tumour types and in large cohorts. PMID- 11113876 TI - An assessment of the long-term preservation of the DNA of a bacterial pathogen in ethanol-preserved archival material. AB - To examine the potential for DNA recovery from spirit-preserved medical material, a set of specimens from the Hunterian Collection of the Royal College of Surgeons was investigated. Using a range of DNA extraction techniques and the PCR, no replicable positive amplifications were made from this material of either human or Helicobacter DNA. Experiments with modern stomach biopsies of H. pylori positive patients suggest that the bacterial DNA is typically present in a much lower concentration (10(3)-fold) than that of the host. The potential for recovery of this organism from spirit specimens is therefore low. The absence of DNA in this material is probably due to several factors, chiefly the incomplete fixation of the specimen by the ethanol storage fluid. Studies such as this demonstrate the need for a good understanding of specimen history when working with archival material. PMID- 11113877 TI - Re. article entitled 'Keratin 20 is a specific marker of submicroscopic lymph node metastases in colorectal cancer: validation by K-RAS mutations'. PMID- 11113878 TI - Authors' reply PMID- 11113879 TI - Mitochondrial DNA 'common' deletion in Hurthle cell lesions of the thyroid. PMID- 11113880 TI - Authors' reply. Mitochondrial DNA damage and oncocytic neoplasia PMID- 11113882 TI - Characteristics of intestinal absorption and disposition of glycyrrhizin in mice. AB - As basic studies to apply an intestinal pressure-controlled colon delivery capsule (PCDC) for glycyrrhizin (GZ), the characteristics of intestinal absorption and disposition of GZ were investigated in mice. In the in vivo study, after intravenous (iv) administration of GZ, 10 mg/kg dose, plasma GZ disappeared from the systemic circulation with t(1/2(alpha)) of 0.0063 h, thereafter, it slowly declined with t(1/2(beta)) of 15.23 h. The area under the plasma drug concentration versus time curve (AUC) values of iv (10 mg/kg), intracolonic (50 mg/kg) and intraduodenum (50 mg/kg) administrations were 115.1, 16.7 and 2.7 microgh/mL, respectively. The AUC values of plasma glycyrrhetic acid (GA), a degradation product after intracolonic and intraduodenum administrations were 2.8 and 8.4 microgh/mL, respectively. In the in situ closed loop study, the concentrations of GZ in plasma and liver after intracolonic administration were significantly increased (p<0.05) in comparison with those after intrajejunum or intraileum administration, while the concentration of GA in plasma and liver after intracolonic administration had trends to increase. These observations clearly suggest that the intracolonic administration is a useful way to improve the oral bioavailability of GZ and to enhance its pharmacological efficacy. These pharmacokinetic results of GZ suggest that GZ is a subject drug to be applied for the PCDC system we previously developed. The PCDCs formulation of GZ will enable us to carry GZ to the colon and enhance the oral bioavailability of GZ. PMID- 11113881 TI - Active efflux kinetics of etoposide from rabbit small intestine and colon. AB - The aim of the present study was to investigate the directional transport kinetics of etoposide in rabbit intestinal tissues using side-by-side diffusion chambers. Etoposide is a routinely used mixed-mechanism 'efflux' inhibitor; however, its absorptive and secretory transport kinetics in rabbit intestinal tissues, a commonly used animal model, have not yet been reported. Kinetic studies revealed that the apical (AP) to basolateral (BL) (i.e. absorptive) transport of etoposide was not apparently mediated by specialized transporters, whereas secretion (i.e. BL to AP transport) by intestinal tissues was concentration dependent and saturable. Half-saturation constant values (K(m), mean+/-standard deviation (S.D.)) ranged from 53.6+/-35.8 microM to 168.7+/-127.3 microM, consistent with previous results from our group in intestinal tissues from other species and Caco-2 cell monolayers. Secretory permeability was greatest in the ileum, whereas values in the upper small intestine and colon were approximately equal, and represented only 50% of the value in the ileum. The ileal secretory transport of etoposide was temperature dependent, with the activation energy (E(a)) >4 kCal/mole at 5 microM, suggesting the involvement of the active, energy dependent mechanism. Etoposide inhibition by verapamil and saquinavir, known inhibitors of intestinal secretion, was characterized as competitive with K(i)'s equal to 193.0+/-164.4 microM and 72.6+/-53.5 microM, respectively. The current results demonstrate that the absorptive transport of etoposide in rabbit tissue was not mediated by specialized carriers, and that secretory transport was regionally dependent, mediated by a transporter or transporters, the K(m)'s were in the micromolar range, and involved the energy dependent mechanism(s). The relatively low k(m) of etoposide compared with its aqueous solubility (0.25-0.34 mM, pH 5-6.5, 25 degrees C) makes it the excellent mixed-mechanism competitive inhibitor for determining the secretory transport properties of putative drug substrates. Understanding the in vitro secretory transport kinetics of etoposide provides a mechanistic basis for ongoing studies exploring the functional role of 'efflux' in vivo. PMID- 11113883 TI - The effect of body size on the metabolic clearance of carbamazepine. AB - OBJECTIVES: To examine the profile of the known pathways of carbamazepine (CBZ) metabolism in a group of children and adolescents, and to test for associations with physical measurements, age and plasma hormonal levels. STUDY DESIGN: Cross sectional study of children and adolescents attending a neurological outpatients department who were medicated with CBZ. Partial clearances of CBZ to CBZ-epoxide (CBZ-ep), CBZ-10,11-trans-diol (CBZ-diol), 2-hydroxy-CBZ (CBZ-2-OH), 3-hydroxy CBZ (CBZ-3-OH), CBZ-acridan (CBZ-acr) and their respective glucuronides were calculated by relating 24-h recovery of these metabolites from urine to trough steady-state serum CBZ levels. CBZ and its metabolites were measured by a gradient high performance liquid chromatography (HPLC) method. Serum CBZ-ep, LH, FSH, prolactin, IGF-I, and testosterone or oestradiol and progesterone were also measured. Surface area (SA) and liver volume (LV) were calculated from height and weight. RESULTS: Twelve males and nine females with an age range of 6-17 years participated in the study. Partial clearance to each of the metabolites was most strongly correlated with the calculated size of the liver relative to body weight. These associations persisted when corrected for potential confounders using multiple regression analysis. CONCLUSION: In the age group studied, urinary clearance of CBZ to its known metabolites is proportional to the size of the liver relative to body weight. PMID- 11113884 TI - Pharmacokinetics, skin absorption, stability, blood partition, and protein binding of AS 2-006A, a new wound healing agent. AB - After intravenous administration of AS 2-006A, 20, 50, and 90 mg/kg, to rats, the pharmacokinetic parameters, terminal half-life (69.8-86. 6 min), mean residence time (56.2-75.2 min), apparent volume of distribution at steady state (809-1040 mL/kg), and total body clearance (11.4-11.9 mL/min/kg), were dose-independent. After topical application of 0.5 or 1% AS 2-006A ointment, 300 mg, to abraded rat skin, the absorbed amounts were dose (0.5 and 1%) and time (30, 60, 120, 240, 360 and 480 min)-independent; the value was approximately 20%. The tissue-to-plasma ratios of AS 2-006A were greater than unity in all rat tissues studied, except in the muscle and large intestine. AS 2-006A was stable for up to 24 h incubation in rat plasma, and human plasma and urine; however, it seemed not to be stable in rat urine; the disappearance rate constant was 0.0218/h. AS 2-006A reached equilibrium fast between plasma and blood cells, and the equilibrium plasma/blood cells partition ratios were independent of the initial rabbit blood concentrations of AS 2-006A, 10, 20, and 50 microg/mL; the mean values were in the range of 2.38-2.75 for three rabbit blood. The protein binding of AS 2-006A to rat plasma was high, as the drug was under detection limit in the filtrate at the plasma concentrations of the drug, ranging from 7.21 to 228 microg/mL. PMID- 11113885 TI - The practical importance of pedigree analysis in women considering invasive prenatal diagnosis for advanced maternal age or abnormal serum screening tests. AB - Genetic counselling prior to prenatal diagnosis subserves several functions, one of them to put the planned prenatal test into the wider context of the personal and familial medical history. Even though considered a pivotal part of counselling, little is known about the informational yield and practical relevance of a comprehensive pre-test pedigree analysis. This is particularly true for patients who do not consider prenatal diagnosis for a specific heritable disorder with a high recurrence risk in the ongoing pregnancy, but for a moderate risk for conditions such as Down syndrome that mostly arise de novo. We analysed the informational yield of pedigree analysis for such patients through a retrospective analysis of 1356 consecutive genetic counselling sessions. All cases were referred for advanced maternal age or an abnormal result upon triple serum marker screening. 148 cases (10.9%) were classified as having a significant and previously unknown genetic or teratologic risk factor for the fetus that was uncovered through pedigree analysis. Of these cases, 55% could be recommended a specific prenatal test covering the previously unknown genetic risk factor. PMID- 11113886 TI - Evaluation of prenatal diagnosis of cleft lip with or without cleft palate and cleft palate by ultrasound: experience from 20 European registries. EUROSCAN study group. AB - Ultrasound scans in the mid-trimester of pregnancy are now a routine part of antenatal care in most European countries. Using data from registries of congenital anomalies a study was undertaken in Europe. The objective of the study was to evaluate prenatal detection of cleft lip with or without cleft palate (CL(P)) and cleft palate (CP). All CL(P) and CPs suspected prenatally and identified at birth in the period 1996-98 were registered from 20 Congenital Malformation Registers from the following European countries: Austria, Croatia, Denmark, France, Germany, Italy, Lithuania, Spain, Switzerland, The Netherlands, UK, Ukraine. These registries followed the same methodology. A total of 709,027 births were covered; 7758 cases with congenital malformations were registered. Included in the study were 751 cases reported with facial clefts: 553 CL(P) and 198 CP. The prenatal diagnosis by transabdominal ultrasound of CL(P) was made in 65/366 cases with an isolated malformation, in 32/62 cases with chromosomal anomaly, in 30/89 cases with multiple malformations and in 21/36 syndromic cases. The prenatal diagnosis of CP was made in 13/198 cases. One hundred pregnancies were terminated (13%); in 97 of these the cleft was associated with other malformations. PMID- 11113887 TI - Prenatal diagnosis of a novel COL1A1 mutation in osteogenesis imperfecta type I carried through full term pregnancy. AB - Prenatal diagnosis was performed in a family where the father has osteogenesis imperfecta (OI) type I, with a novel mutation in the COL1A1 gene: a C to T change at position c3076 (c.3076C-->T) leading to a change of arginine at codon 848 to a stop codon (R848X). Prenatal diagnosis by chorionic villous sampling (CVS) was performed during the fourth pregnancy, and revealed that the fetus is a carrier of the same COL1A1 mutation. The possibility of phenotypic variability was discussed with the parents. They elected to carry the pregnancy to term, and a male child with mild OI was born. This is the first reported case where OI was diagnosed prenatally, and the parents opted to carry the pregnancy to term. It illustrates the potential use of DNA-based analysis for early prenatal diagnosis of OI, and the complexities of genetic counselling. PMID- 11113888 TI - Survival analysis of transfused fetuses affected by Rh-alloimmunization. AB - The aim of the present study was to evaluate the survival rate of a group of 86 fetuses affected by Rh-alloimmunization submitted to intrauterine red blood cell transfusion. All the women had antibody titres> or = 1:32 at the time of their enrollment in the study. Crude fetal survival rate was 89.5% (77/86 cases). Data were stratified according to specific cut-off points of (1) pre-transfusion fetal haemoglobin expressed as the rate between the observed and the estimated value for each gestational age at the time of the first transfusion; (2) the difference between the haemoglobin at the beginning of the second-transfusion less that at the end of the first transfusion (delta haemoglobin); and (3) presence of ultrasound detected anomalies. Statistically significant stratification of the survival rate was observed for the level of pre-transfusion fetal haemoglobin (95% and 76.9%, respectively, p= 0.009) using a cut-off value of < 70% and > or = 70% of the expected value. Again, delta haemoglobin showed a different survival rate when a cut-off value of 6 g/dl was used to generate subgroups of fetuses: 94.6% and 80%, respectively (p= 0.0145). Among the ultrasound anomalies, the presence of hydrops showed a correlation with the survival rates. The quoted values were 97.83% (absence) and 80.0% (presence) respectively (p= 0.0058). Cox regression showed a significant association of the studied variables with the outcome (survival). The presence of hydrops was the best predictor (Odds ratio= 8.7, p= 0.0073) followed by Delta haemoglobin (Odds ratio= 2.0, p= 0.0422). The rate of pre-treatment fetal haemoglobin < 70% of the expected value did not add any significant valu and was thus removed from the final model. Weight at delivery expressed in grams showed a direct correlation with the survival rate (Odds ratio= 0.9, p= 0.1529) and was added into the model as an adjustment quantitative variable. PMID- 11113889 TI - Apoptosis in fetal nucleated erythrocytes circulating in maternal blood. AB - The purpose of this study was to determine if apoptosis occurs in fetal cells that have crossed into the maternal circulation, which would potentially explain the difference between the number of intact fetal cells and the amount of fetal DNA detectable in maternal plasma. We flow-sorted fetal nucleated erythrocytes (FNRBCs) using antibody to the gamma chain of fetal haemoglobin and confirmed them to be fetal in origin by FISH analysis using chromosome-specific probes. Fetal cells were then analysed microscopically for the presence of terminal UdTP nuclear end labelling (TUNEL) staining. Apoptotic change was observed in 42.7% of fetal NRBCs (106/246) and 3.5% of maternal cells (29/818). Results of this study indicate that a significant number of fetal cells in maternal blood are undergoing apoptosis at the time of sampling. Apoptosis may be one mechanism by which fetal cells are cleared by the maternal circulation. PMID- 11113890 TI - The subarachnoid space: normal fetal development as demonstrated by transvaginal ultrasound. AB - Enlargement of the subarachnoid spaces can be seen in the following conditions: communicating hydrocephalus, brain atrophy and benign enlargement of the subarachnoid spaces. These disorders may begin in utero. There are no established normograms for the fetal subarachnoid spaces. This study was conducted in order to determine its normal development. Transvaginal sonography was used to examine the subarachnoid space in 80 fetuses between 16 and 40 weeks' gestation. The sinocortical width (SCW) and craniocortical width (CCW) were measured in a coronal plane at the level of the foramen of Monro. The SCW remained relatively constant during the gestational period. The CCW increased in size from the 20th to the 28th week of pregnancy, with a subsequent gradual decrease until term. Determination of fetal subarachnoid space normograms may potentially help in the diagnosis of pathological conditions affecting this space and allow prenatal counselling. PMID- 11113891 TI - Axial growth of the fetal eye and evaluation of the hyaloid artery: in utero ultrasonographic study. AB - The aims of this prospective, cross-sectional study were to report axial ocular growth during human gestation, to determine the presence of the hyaloid artery (HA) and its blood flow, and to provide a timetable for HA regression. The study group comprised 231 low-risk singleton pregnancies between 14 and 38 weeks' gestation. Ocular axial length (OAL), anterior chamber depth (ACD) and posterior chamber depth (PCD) were measured using high-resolution ultrasound. The growth of these eye segments in correlation with gestational age (GA) was established. The presence of the HA and its regression were determined. By using power Doppler, ultrasound blood flow within the HA was estimated. HA regression is a gradual process that is not evident before 18 weeks' gestation. In all fetuses beyond 29 weeks' gestation, no HA could be detected (P<0.001). Blood flow within the HA was documented only until the 16th week of gestation. The correlation coefficients, r=0.924, 0.784 and 0.929, for OAL, ACD and PCD, respectively, were found to be highly statistically significant (P<0.0001). The present data offer normative measurements of the fetal axial eye lengths, timetable for HA regression and flow cessation. PMID- 11113893 TI - Isolated polydactyly: prenatal diagnosis and perinatal outcome. AB - Our objective was to determine the clinical significance of isolated polydactyly identified on prenatal sonogram. All patients with sonographically detected isolated polydactyly scanned over an 11-year period were identified from our database. All patients underwent detailed surveys, and follow-up was obtained by review of the medical records and telephone conversations with parents and referring physicians. Thirteen patients with isolated polydactyly were identified. Follow-up was available in 12 patients. Indications for referral included advanced maternal age (2), second-opinion polydactyly (4), family history of polydactyly (1), uncertain dates (5), and growth (1). The gestational ages at the times of sonographic diagnosis ranged from 17.5 to 34 weeks with all but one case being identified before 23 weeks. Prenatal identification included polydactyly of the upper limb (8), lower limb (4), and both upper and lower limbs (1). Postaxial polydactyly was seen in 12 patients and preaxial in one. Polydactyly was confirmed in all 12 cases in which follow-up was available. Karyotypes were normal in all five fetuses in which amniocentesis was performed. Ten of 12 fetuses were born alive, one died in utero at 34 weeks as a complication of severe pre-eclampsia and one died at term as a result of a cord accident. No surviving neonate had any other identifiable malformation or suspected karyotypic abnormality. In conclusion isolated polydactyly identified by prenatal sonography is associated with good perinatal outcome. PMID- 11113892 TI - Prenatal diagnosis of haemoglobin Bart's disease by cordocentesis at 12-14 weeks- experience with the first 59 cases. AB - We have shown that fetuses affected by haemoglobin (Hb) Bart's disease can be reliably identified by their sonographic manifestation of cardiac enlargement at 12-14 weeks. Between 1995 and 1999, 282 couples were seen before 15 weeks. They were offered the options of chorionic villus sampling, or amniocentesis and DNA study, or ultrasound examination at 12-14 weeks, followed by cordocentesis and Hb study only when the ultrasound findings were abnormal. Two hundred and thirty four at-risk pregnancies had ultrasound assessment at 12-14 weeks, 62 fetuses showed enlarged cardiothoracic ratio [mean (SD) 0.54 (0.02)] and four of them also had hydropic changes. Fifty-nine women agreed to undergo cordocentesis at 12 14 weeks and the procedure was successful in 57 cases (97%). Cordocentesis were performed by a freehand technique using a 26- or 24-gauge spinal needle with a 20 gauge introducer. Fifteen fetuses (25%) had bleeding from the cord and 12 fetuses (20%) had bradycardia following cordocentesis. The fetal loss rate was 8% (5/59). Hb Bart's disease was confirmed in all the 62 fetuses with cardiac enlargement. Their Hb concentration ranged between 3.1 to 8.4 g/dl. One hundred and seventy two fetuses had normal ultrasound assessment and 148 of them were confirmed to be unaffected by Hb Bart's disease. Twenty-three pregnancies were ongoing and one miscarried at 15 weeks. We believe that sonographic assessment followed by selective cordocentesis at 12-14 weeks is a feasible prenatal diagnostic option for Hb Bart's disease. PMID- 11113894 TI - Hyperechogenic bowel in the second trimester fetus: a review. PMID- 11113895 TI - Retrospective diagnosis of trisomy 15 in formalin-fixed, paraffin-embedded placental tissue in a newborn girl with Prader-Willi syndrome. AB - Paternal deletion of 15q11-q13 and maternal uniparental disomy (UPD) of chromosome 15 are the main causes of Prader-Willi syndrome (PWS). The finding of an UPD(15) is associated with increased maternal age. We present a retrospective diagnosis of a trisomy 15 mosaicism confined to the placenta (CPM) after birth of a girl with clinical features of PWS born to a 43-year-old mother. Chromosome analysis after amniocentesis, performed because of advanced maternal age, had shown a normal female karyotype. In peripheral blood cells molecular studies showed the absence of the paternal allele at the SNRPN locus and fluorescence in situ hybridization (FISH) analysis excluded a deletion of the SNRPN locus on both chromosomes 15. Trisomic cells were detected by FISH on nuclei isolated from formalin-fixed, paraffin-embedded placental tissue using a DNA-probe specific for the centromeric region of chromosome 15. PMID- 11113896 TI - An unusual case of trisomy and triploidy in a chorion villus biopsy. AB - A case is reported of a 35-year-old woman who underwent a chorion villus biopsy (CVB) at 17 weeks' gestation after intrauterine growth retardation and oligohydramnios were diagnosed by ultrasound scan. Chromosome analysis of the CVB direct preparations showed a 47,XX,+6 karyotype in all cells. The pregnancy was terminated and subsequent analysis of cultured cells from both the CVB and the post-mortem placenta showed three cell lines: 46,XX, 47,XX,+6 and 69,XXX, while fetal skin and muscle were entirely 69,XXX. An explanation is proposed for the origin and distribution of the three cell lines. PMID- 11113897 TI - 3-M syndrome: a prenatal ultrasonographic diagnosis. AB - The ultrasonographic imaging of a fetus affected by 3-M syndrome is described. This is a primordial dwarfism with low birthweight, short stature, facial dysmorphism and normal mental development. The biparietal diameter and head circumference were in accordance with the gestational age at 18 weeks. The femur and tibia lengths were on the fifth centile and the radius, ulna and humerus lengths were below the fifth centile. A second scan at 22 weeks showed slowing of growth of all long bones, with the femur, tibia, fibula, humerus, radius and ulna lengths further below the fifth centile. The pregnancy was terminated and postmortem examination confirmed the prenatal diagnosis. The differential diagnosis of skeletal dysplasias characterized by a slow growth of long bones is discussed and the conclusion reached that the detection of shortened long bones (below the fifth centile) is the only ultrasonographic finding of 3-M syndrome. PMID- 11113898 TI - Transposition of the great arteries, ventricular septal defect and diaphragmatic hernia in a fetus: the role of prenatal diagnosis in helping to predict postnatal survival. AB - Congenital heart disease and congenital diaphragmatic hernia are frequently associated. The combination of these lesions is predictive of poor postnatal survival. The identification of both lesions during prenatal life may facilitate improved survival in carefully selected cases. We present a case of left-sided diaphragmatic hernia with transposition of the great arteries and a ventricular septal defect (VSD) that survived following repair of both defects within the first six weeks of life. PMID- 11113899 TI - Prolidase deficiency among an Israeli population: prenatal diagnosis in a genetic disorder with uncertain prognosis. AB - Prolidase deficiency is an autosomal recessive disorder that is characterized by considerable inter- and intrafamilial variability in its clinical presentation, ranging from asymptomatic to severe and fatal illness. We report here, for the first time, prenatal diagnosis of prolidase deficiency in a family whose first child was severely affected since birth and died at an early age. However, unexpectedly, the parents decided to continue the second pregnancy, which produced a full-term, healthy-appearing baby. The diagnosis of severe prolidase deficiency was confirmed in the baby's leukocytes. At age 4 months the baby is asymptomatic. Since the clinical severity of the disorder cannot be predicted, genetic counselling remains problematic despite the feasibility of prenatal diagnosis. PMID- 11113900 TI - A case of maternal uniparental disomy of chromosome 9 diagnosed prenatally and the related problem of residual trisomy. AB - Non-mosaic trisomy 9 was found in a chorionic villus (CV) sample taken from a 43 year-old woman referred for prenatal chromosome analysis due to advanced maternal age. Follow-up amniocentesis revealed level 2 mosaicism for trisomy 9. Trisomy 9 was not detected at fetal blood sampling. Molecular analysis of fetal (amniocyte) DNA showed maternal uniparental heterodisomy (UPD) for chromosome 9. Two crossovers resulted in a region of isodisomy in the distal long arm. Trisomy rescue of a meiosis 1 segregation error seems to have been responsible for the uniparental disomy of chromosome 9. The pregnancy continued and neonatal blood testing showed a mosaic trisomy 9 karyotype, i.e. 4/50 cells analysed. Clinical postnatal follow-up for a period of 1 year has documented only minor facial dysmorphism and skeletal abnormalities. Development appears unremarkable. This case is the second report of maternal uniparental disomy for chromosome 9 detected prenatally and is the first case followed up post-term. This report highlights the difficulty of making informed prognostic assessments in such cases despite extensive laboratory investigation. PMID- 11113901 TI - Cytogenetic discrepancy between fetal tissue and body fluid in a fetus with cystic hygroma colli. PMID- 11113902 TI - Prenatally detected true double trisomy mosaic 12 and 13 not confirmed in fetal and placental tissues by conventional cytogenetic methods, but suggested by FISH analysis. PMID- 11113903 TI - Prenatal diagnosis of a partial trisomy 7q in two fetuses with bilateral ventriculomegaly. PMID- 11113904 TI - Current awareness in prenatal diagnosis. PMID- 11113905 TI - Prenatal diagnosis of holoprosencephaly (HPE) in a fetus with a recombinant (18)dup(18q)inv(18)(p11.31q11.2)mat. AB - Alobar holoprosencephaly (HPE) was identified by ultrasonography at 18 weeks' gestation in a fetus of a 29-year-old G2P0A1 woman. HPE has been described in association with various chromosomal anomalies. Amniocentesis was performed and a rearrangement of chromosome 18 resembling an isochromosome for the long arm of chromosome 18 was found. Subsequently, the mother was found to have a pericentric inversion of chromosome 18 with breakpoints at p11.31 and q11.2. The karyotype of the fetus was re-interpreted as 46,XX, rec(18)dup(18q)inv(18)(p11.31q11.2)mat. This is the first case of a parental inversion leading to a deficiency of 18p11.31 to 18pter associated with HPE. PMID- 11113906 TI - Four years' cytogenetic experience with the culture of chorionic villi. AB - In 1958 chorionic villus samples, investigated by culture method, we found 137 (7%) abnormalities. The abnormal results were classified in certain abnormal (generalised abnormal at high probability) and uncertain abnormal (potentially confined to the placenta) results. Certain abnormal were 73 cases (3.7%). Uncertain abnormal were 64 cases (3.3%), in which confirmation studies were done in 47 cases. In 12 cases of these 47, the abnormality was confirmed and in 35 cases (1.8%) the abnormality was confined to the placenta. Among the latter cases, poor pregnancy outcome [16% intrauterine death (IUD), 6% intrauterine growth retardation (IUGR)] was increased. Total maternal cell contamination was not seen. The positive predictive value of all confirmed abnormal cases was 66%. The positive predictive value was 100% for indications 'ultrasound abnormalities' and 'carrier' and between 50 and 60% for all other indications. Predictive value among uncertain abnormal cases was low (26%). However, the positive predictive value depends of the type of abnormality. Therefore we conclude that the culture method for chorionic villi is a good test for indications 'ultrasound abnormalities' and 'carrier' and reliable for all other indications. Whether or not follow-up investigations should be offered to the parents depends of the type of abnormality. We conclude that the culture method is reliable for prenatal diagnosis and can be used as the sole investigative method. PMID- 11113907 TI - Accuracy of abnormal karyotypes after the analysis of both short- and long-term culture of chorionic villi. AB - We report in detail the cytogenetic results of 1838 consecutive chorionic villus samples with the availability of both short-term culture (STC-villi) and long term culture (LTC-villi) preparations in 1561 cases (84.9%). A high degree of laboratory success (99.5%) and diagnostic accuracy (99.8%) was observed; in four cases of low mosaicism, all four associated with the final birth of a normal child, a small risk of uncertainty was accepted. The combined analysis of STC- and LTC-villi reduced follow-up amniocenteses by one-third in comparison with the analysis of STC-villi alone. We believe that the desired level of quality and accuracy of prenatal cytogenetics in chorionic villi can only be achieved when both STC- and LTC-villi are available. We conclude that CVS might then be the mode of prenatal diagnosis of first choice in pregnancies with a high (cytogenetic) risk. PMID- 11113908 TI - Correlation of prenatal clinical findings with those observed in fetal autopsies: pathological approach. AB - Our objective was to present a comprehensive description of the clinicopathological findings of 173 abortions, including 121 therapeutic and 52 spontaneous ones in the period between 1992 and 1998. In all of these fetuses pathological examination was carried out. It was complemented when indicated by immunohistochemistry, in situ hybridization, flow cytometry, and X-ray examination. In the 121 therapeutic abortions the distribution of malformations was: 45 central nervous system anomalies (37%), 12 genitourinary anomalies (10%), 25 gastrointestinal anomalies (21%), two respiratory system anomalies (1.65%), eight cardiac anomalies (6.6%) and 28 other anomalies (17.2%) as revealed by autopsy. From the clinically selected 52 spontaneous abortions, major malformations were seen in 15/52 cases. With the comparison of the pathological and clinical findings in 121 therapeutic abortions, the percentage of cases with correct clinical designation and no missed anomalies amounted for 49%. However in 51% additional or different lethal, severe, or major malformations were revealed or excluded by fetopathological examinations. In 4% the clinical observation and diagnosis were modified, but without implications for the therapeutic termination of pregnancy. The clinical indication could not be supported in another 3% of the cases. PMID- 11113909 TI - Urinary hyperglycosylated hCG in first trimester screening for chromosomal abnormalities. AB - Hyperglycosylated human chorionic gonadotrophin (H-hCG), also known as Invasive Trophoblast Antigen or ITA, is a unique metabolic variant of hCG with more complex oligosaccharide side chains. Concentrations are independent of regular hCG. Urine H-hCG has recently proved to be a highly sensitive marker for Down syndrome screening in the second trimester of pregnancy. We evaluated H-hCG as a potential marker in the first trimester of pregnancy. Maternal urine samples were collected from 10(+0) to 11(+6) weeks of gestation prior to genetic analysis and stored in frozen form. Samples from eight cases of Down syndrome, two cases of trisomy 13, one case of trisomy 18, and 55 control pregnancies were hand-carried frozen to the USA and tested blindly. Samples were tested in a specific H-hCG immunoassay and values were normalized to creatinine concentration. Values were plotted against gestational age, and multiples of control pregnancy median (MoM) calculated. The median level of the MoMs of the eight Down syndrome cases was 3.6 MoM. Five of the eight Down syndrome cases exceeded the 90th centile of the 55 unaffected cases. The MoMs of the trisomy 13 and 18 pregnancies were 0.2, 0.2 and 0.3. All three cases were under the 10th centile of unaffected pregnancies. The results of this study indicate that H-hCG testing may be useful in screening for Down syndrome in the first trimester of pregnancy. Further studies are needed to assess the potential screening values of urine H-hCG and the combination of this test with free beta-subunit, PAPP-A and other markers for Down syndrome in the first trimester of pregnancy. PMID- 11113910 TI - Cost-effective screening methods for various single gene defects in single cells using high magnesium and total ionic strength and restriction enzymes. AB - A reliable cost-effective protocol for the diagnosis of various defective genes in single blastomeres from preimplantation embryos has been established. Single cells were lysed in alkali buffer followed by neutralization and addition of a solution containing a high concentration of sulfhydryl reducing agents and MgCl(2) in relatively high ionic strength (0.45) (solution M) with or without restriction enzyme(s). The reaction mixture was incubated at 37 degrees C for 15 min followed by heat denaturation at 95 degrees C for 10 min. Respective polymerase chain reaction (PCR) mixture was then added to amplify each designated DNA region. The treatment of neutralized single cell lysate with adequate restriction enzyme(s) which do not cleave the target DNA sequences but shortens the genomic template DNA strands. This may facilitate primer-template annealing. The subsequent heat denaturation of the cell lysate in solution M indeed gave better signals of amplified DNA fragments on polyacrylamide gels. Defects in Tay Sachs exons 11 and 12, CF-DeltaF508 and CF-N1303K, and genomic sequences of ZFX/ZFY were successfully detected on gels after one-step PCR amplification, especially those cell lysates treated with restriction enzymes. In conclusion, a cost-effective one-step PCR method for amplifying various specific genomic regions containing a single gene defect in single cells has been established. This protocol may be applied to genetic screening for many single defective genes of biopsied single blastomeres from preimplantation in vitro fertilization (IVF) embryos. PMID- 11113911 TI - Accuracy of prenatal diagnosis for haemoglobin disorders in the UK: 25 years' experience. AB - We have reviewed the accuracy of prenatal diagnosis for the thalassaemias and sickle cell disorders performed for UK residents since the service began in 1974. Prenatal diagnosis has been performed in 3254 pregnancies: 517 by fetal blood analysis, 681 by Southern blotting and 2056 by polymerase chain reaction (PCR) methods, the majority using the amplification refractory mutation system (ARMS). The number of homozygotes diagnosed was 808 (24.8%). Twenty-five diagnostic errors have been recorded, ten arising from non-laboratory errors (0.31%) and 15 due to technical problems associated with the diagnostic techniques. The latter group consisted of eight misdiagnoses by globin chain synthesis (1.55%), five by Southern blot analysis (0.73%) and two by PCR methods (0. 10%). The data show that the accuracy of prenatal diagnosis has improved with each development of diagnostic technique, and confirms that prenatal diagnosis of beta-thalassaemia and sickle cell disorders by ARMS-PCR is very accurate and reliable. The overall error rate for prenatal diagnosis by PCR methods in the UK is now 0. 41%. PMID- 11113912 TI - Prenatal diagnosis of del(15)(q26.1) and del(18)(q21.3) due to an unbalanced de novo translocation: ultrasound, molecular cytogenetic and autopsy findings. AB - A case of partial deletion of the distal parts of chromosomes 15 and 18 [(15)(q26.1)(18)(q21.3)] due to a de novo translocation is reported. Cordocentesis and fetal karyotyping was done because of severe oligohydramnios and bilateral absence of kidneys. Renal defects are a frequent finding in fetuses with different chromosomal anomalies; this particular chromosomal rearrangement however has not been reported yet in a fetus with bilateral renal agenesis. FISH was performed for detailed clarification of the chromosomal anomaly. Prenatal karyotyping appears to be important in fetuses with renal agenesis. PMID- 11113913 TI - Prospective ultrasound diagnosis of Pallister-Killian syndrome in the second trimester of pregnancy: the importance of the fetal facial profile. AB - The Pallister-Killian syndrome (PKS) represents a rare polymalformative complex characterized by a tissue-specific mosaic distribution of an additional isochromosome 12p and characterized by diaphragmatic hernia, rhizomelic limb shortening, facial anomalies and, rarely, acral hypoplasia. Since diaphragmatic hernia and acral hypoplasia can be also found in Fryns syndrome, the differential diagnosis between the two conditions depends on the demonstration of the 12p isochromosome by FISH. Prenatal diagnosis of PKS has been reported in cases submitted to karyotyping due to advanced maternal age or congenital anomalies detected on second trimester ultrasound. Among the ultrasound-detected malformations, little attention has been paid to facial anomalies. We describe a case in which PKS was prospectively suspected on the basis of the various anomalies detected at ultrasound, namely diaphragmatic hernia, rhizomelic limb shortening, and abnormal facial profile. The diagnosis was then confirmed by FISH on amniocytes and peripheral lymphocytes. In the present case, the disclosure of typical facial abnormalities significantly contributed to the differentiation between PKS and Fryns syndrome. PMID- 11113914 TI - Prenatal diagnosis of heterokaryotypic mosaic twins discordant for fetal sex. AB - The presence of a monozygotic twin gestation with discordant sex of the twins is a very rare constellation, which is referred to as heterokaryotypic monozygotic pregnancy. This constellation can develop either due to a chromosomal aberration after twinning or is - as in the following case - due to a mitotic error before twinning and an unequal distribution of mosaicism in both embryos. So far the diagnosis of heterokaryotypic monozygotic pregnancy has always been made postnatally, with only one exception (Gonsoulin et al., 1990). In this case we suspected the presence of monozygotic twins ultrasonically because of the chorionic and amniotic membrane characteristics. Surprisingly the sex of the fetuses was discrepant. As one of them had hydrops and a structural heart defect, we carried out an amniocentesis, which revealed mosaicism [45,X/46,X,i(Y)(p10)] of both fetuses. The female fetus with a predominant 45,X set of chromosomes and the typical intrauterine signs of the Ullrich-Turner syndrome (massive hygroma colli, hydrops fetalis and multiple cardiac defects) died during the 25th week of gestation due to cardiac decompensation. The other fetus appeared to be male with a predominance of a 46,X,i(Y)(p10) set of chromosomes and was born a few days after the intrauterine death of the hydropic fetus. In conclusion, our observation shows that ultrasonic evidence of discordant fetal sex in twins does not necessarily exclude monozygosity. PMID- 11113915 TI - Prenatal MRI in a fetus with a giant neck hemangioma: a case report. AB - We report a fetus with a giant neck hemangioma which was examined by MRI in utero. The initial diagnosis was made by ultrasonography. The sonolucent aspect of the mass, together with the presence of pulsating Doppler flow signals, was highly suggestive of a fetal hemangioma. In late pregnancy, fetal MRI revealed the location, size and characteristics of the neck tumor. Following prenatal corticosteroid treatment and premature delivery of the pregnancy due to fetal cardiac failure, the newborn received angiography and coil embolization of the tumor vessels. Despite vigorous treatments, the newborn died 12 h after birth. Evaluation of a fetal neck hemangioma by MRI is recommended late in pregnancy for precise information on the tumor and adjacent organs since the image is valuable for planning optimal perinatal treatment. PMID- 11113916 TI - Greenberg dysplasia: first reported case with additional non-skeletal malformations and without consanguinity. AB - In 1988 Greenberg et al. reported an association of fetal hydrops with irregular calcification and moth-eaten skeletal dysplasia. Here, we report on the first case of this disorder accompanied by additional malformations (omphalocele, intestinal malrotation, disturbed fingernails and toes, hypolobated lungs) in a German couple without consanguinity (karyotype 46,XY). Sonograpically, the fetus was characterised by tetraphokomelia, severe generalised hydrops, pulmonal hypoplasia and hepato-splenomegaly. Greenberg dysplasia should be considered in differential diagnosis in cases with severe fetal hydrops and phokomelia on antenatal sonography. PMID- 11113917 TI - MASA syndrome: ultrasonographic evidence in a male fetus. AB - The recent identification of a common etiology among MASA syndrome (McKusick 303300), X-linked hydrocephalus (HSAS) (McKusick 307000) and other related neurological disorders, which had previously been considered distinct nosological entities, allowed us to diagnose MASA syndrome in a male fetus in a primigravida at the 29th week of gestation by sonographic signs of the MASA spectrum such as hydrocephalus and hypoplasia of corpus callosum. Indeed, the evidence of an X linked neurological disease in the brother and the maternal uncle of the pregnant women enabled us to estimate a 25% risk of a male fetus being an affected hemizygote. The way in which a prenatal diagnosis, based on instrumental procedures, was reached is described since the authors were unable to perform, at the time of the observation, a molecular confirmation which was carried out only after birth. PMID- 11113918 TI - High maternal serum inhibin A levels following the loss of one fetus in a twin pregnancy. AB - Inhibin A levels are elevated in the second trimester of pregnancies affected with fetal Down syndrome, on average, two times the level in unaffected pregnancies. Inhibin A levels are also two times higher in twin than in singleton pregnancies. Prenatal serum screening using inhibin A levels as a second trimester marker began at the Women and Infants Hospital in March 1998. We describe a case of a 17-year-old woman thought to have had a complete spontaneous abortion of a twin pregnancy but later found to be continuing the pregnancy with a single fetus. Routine second trimester prenatal serum screening revealed an extremely elevated inhibin A level of 39 MoM (multiples of the median). The patient delivered an apparently healthy female infant at 41 weeks of gestation. Therefore, inhibin A may be extremely elevated in the second trimester of a twin pregnancy after the loss of one fetus and this increased inhibin A level does not have any obvious adverse maternal or fetal effects. PMID- 11113919 TI - Risks of genetic amniocentesis in women less than 35 years old. PMID- 11113920 TI - Quality control in components of comparative genomic hybridization technique used for diagnostic cytogenetic analysis. PMID- 11113921 TI - Prenatal diagnosis of limb-body wall complex using two- and three-dimensional ultrasound. PMID- 11113922 TI - Current awareness in prenatal diagnosis. AB - In order to keep subscribers up-to-date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of prenatal diagnosis. Each bibliography is divided into 17 sections: 1 Books, Reviews & Symposia; 2 General Interest; 3 Normal Fetal Development; 4 Gametogenesis and Pre-implantation Diagnosis; 5 First Trimester Diagnosis; 6 Second Trimester Diagnosis; 7 Fetal Diagnosis by Ultrasound and Other Imaging; 8 Maternal Screening; 9 Screening for Carriers of Genetic Abnormality; 10 Technological Developments; 11 Confined Placental Mosaicism and Uniparental Disomy; 12 Molecular Cytogenetics; 13 Fetal Cells in Maternal Circulation; 14 Fetal Therapy; 15 Psychosocial Aspects; 16 Epidemiology and Environmental Factors; 17 Developmental Pathology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. PMID- 11113923 TI - Redox properties of isradipine and its electrochemical detection in the HPLC determination of the compound in human serum. AB - The electrochemical behaviour of isradipine in a mixed solution of Britton Robinson buffer (pH 11.8):acetonitrile:methanol (6:3:1, v/v) was studied by cyclic voltammetry and spectroelectrochemistry using an optically transparent thin layer electrode of carbon cloth. The cyclic voltammogram showed several peaks whose shape and potentials depended on the pH. The peak at 330 nm, corresponding to the absorbance of the dihydropyridine ring, disappeared after electrolysis at a potential that was more positive than the oxidation peak. The oxidation peak corresponds to the oxidation of the dihydropyridine ring. Peak height at pH 11.8 was proportional to isradipine concentration. On the basis of the redox properties of isradipine, HPLC was conducted applying electrochemical detection on a polybutadiene coated alumina column using an alkaline mobile phase. The method was applied for the determination of isradipine content in human serum. A good linear relationship between isradipine concentration and peak height was found in the concentration range of 2-200 ng/mL with a correlation coefficient of 0.9924. The detection limit was 0.5 ng/mL. The within-day and day to-day variation were examined for control human serum and percentage relative standard deviation ranged from 0.5 to 6.7. Interference from many other coadministered drugs was studied in the specified experimental conditions. Photo and heat stabilities of the compound were also studied. PMID- 11113924 TI - Excretion pattern investigation of urinary normal and modified nucleosides of breast cancer patients by RP-HPLC and factor analysis method. AB - Modified nucleosides, formed post-transcriptionally in RNA by a number of modification enzymes, are excreted in abnormal levels in the urine of patients with malignant tumors. To test their usefulness as tumor markers, and to compare them with the conventional tumor markers, a reversed-phase high-performance liquid chromatographic (RP-HPLC) method and a factor analysis method have been used to study the excretion pattern of nucleosides of breast cancer patients. A clear cut differentiation of the breast cancer group and the healthy individuals in two clusters without overlapping was obtained. PMID- 11113925 TI - Isolation of 5 alpha- and 5 beta-dihydrorubrosterone from Silene otites L. (Wib). AB - 5 alpha-Dihydrorubrosterone (2 beta, 3 beta, 14 alpha, 17 beta-tetrahydroxy-5 alpha-androst-7-ene-6-one), a new 19-carbon 5 alpha-ecdysteroid, was isolated together with its 5 beta counterpart from the aerial parts of Silene otites L. (Wib.) (Caryophyllaceae) by a combination of solvent partition, low-pressure column chromatography, thin-layer chromatography (normal-phase and reversed phase) and finally HPLC. Mass spectrometry and nuclear magnetic resonance spectroscopic procedures were used for compound characterization. PMID- 11113927 TI - Enantiomeric determination of L- and D-lactic acid in human cerebrospinal fluid by chiral ligand exchange high-performance liquid chromatography. AB - Enantiomeric determination of L- and D-lactate in human cerebrospinal fluid (CSF) was achieved by HPLC on a chiral stationary phase with UV detection. Samples were submitted to a solid-phase extraction procedure using Oasis HLB Plus Extraction Cartridge and L- and D-lactate in the extract were separated by Shodex ORpac CRX 453 B column, a ligand exchange column for chiral separation, using a mobile phase containing copper (II) ion. L- and D-lactate were determined in 25 min. Intra-assay precision in CSF was 4.98% (mean 1.85 mmol/L) for L-lactate and 10.1% (mean 4.96 micromol/L) for D-lactate (n = 5). Detection limits were between 1.0 (L-lactate) and 1.5 (D-lactate) pmol. The mean values (n = 3) of analytical recovery for L- and D-lactate were 95% and 107%, respectively. The mean +/- SD of concentrations of L- and D-lactate in CSF (n = 20) were 1.52 +/- 0.54 mmol/L and 10.98 +/- 5.15 micromol/L, respectively. PMID- 11113926 TI - Simultaneous determination of reduced and oxidized glutathione in freshly isolated rat hepatocytes and cardiomyocytes by HPLC with electrochemical detection. AB - Glutathione and glutathione disulphide constitute an essential thiol redox system present in the cell. The balance in favour of the latter is an indication of oxidative stress. Glutathione and glutathione disulphide quantification in isolated cells may therefore be essential for the evaluation of mechanistic and comparative studies of toxic xenobiotics. In this study, a rapid and sensitive isocratic reverse-phase high-performance liquid chromatographic method using coulometric detection was implemented for the simultaneous detection of glutathione and glutathione disulphide, in freshly isolated hepatocytes and cardiomyocytes of the rat. The method implemented proved to be effective for the measurement of glutathione and glutathione disulphide in control conditions and for the detection of variations in this redox system, induced by tert butylhydroperoxide. tert-Butylhydroperoxide is an organic peroxide, which has been used as a model molecule for inducing oxidative stress in isolated cells. A comparative study with a previously published HPLC-electrochemical detection method was performed. PMID- 11113928 TI - Chromatography of human prothrombin from Nitschmann fraction III on DEAE Sepharose Fast Flow using axial and radial flow column. AB - An axial column (3 x 2.6 cm) and a radial flow column (3.5 x 5 cm) packed with DEAE Sepharose Fast Flow media was evaluated for the separation of human prothrombin from Nitschmann fraction III. Under radial flow conditions, a sample flow rate up to 14 mL/min (approximately 18 bed vols/h) was achieved. Breakthrough capacity was determined and both columns had almost the same breakthrough capacity per mL media, indicating that the sample loading was independent of radial column geometry. PMID- 11113929 TI - The combined use of high performance liquid chromatography and immuno-biochemical techniques for protein isolation: a new approach for identification of an individual protein from a pool of proteins. AB - HPLC was used in combination with immuno-bead separation technique for identification of an individual protein from a pool of proteins. This was carried out using an in-house monoclonal antibody (ATC2) specific for placental alkaline phosphatase (PLAP) as a primary antibody for conjugation to CNBr beads. The phosphatase activity (ALP) of PLAP was measured by colorimetric assay (MEDC). The data from this study has so far indicated that: 1. HPLC analysis of molecules following isolation with ATC2-conjugated beads showed high degree of purity. This could be achieved using protein mixtures prepared from lysates of tumour cell lines or tumour fragments. 2. HPLC-isolated PLAP maintained phosphatase activity. 3. Out of the four dissociation reagents used, diethyl amine (DEA) was found to be the best reagent for dissociation of antigen, ie PLAP, but not mAb from CNBr beads. 4. The profile of ALP activity was different for samples prepared from testis and kidney fragments, both in terms of the HPLC peak profile as well as the sensitivity. These data confirmed that the immuno-bead separation technique in conjunction with HPLC were powerful tools for identifying an individual protein from a pool of proteins. These approaches are being used for the identification of PLAP molecules, as a tumour marker in patients suspected of testicular malignancies with equivocal ultrasound. PMID- 11113930 TI - Application of capillary electrophoresis to the analysis of soluble chromatin. AB - Capillary electrophoresis (CE) has been applied to study DNA-protein complexes using as the test system soluble chromatin from chicken erythrocytes and rapidly proliferated cultured Chinese hamster fibroblast-like cells B11-dii-FAF-28. Separation was performed with home-made CE apparatus, using a regulated high voltage power supply, UV-detector and fused silica capillaries with inner diameter 75 microm. The heterogeneity of nucleosomal particles with different DNA lengths after micrococcal nuclease digestion was detected. PMID- 11113931 TI - The advantages of cell lysis before blood sample preparation by extraction for HPLC propofol analysis. AB - Propofol (2,6-diisopropylphenol) is a short-acting drug with a large volume of distribution and high body clearance. It is suitable both for the induction of anaesthesia by bolus injection and the maintenance of anaesthesia by repeated injections or a continuous infusion. Examining the drug concentration its analysis in whole blood is recommended. This results from the fact that propofol molecules strongly bind with plasma proteins and cellular blood constituents and blood composition variations are observed between individuals or in different disease states or resulting from transfusion etc. In most cases the HPLC analysis follows the extraction of samples. The degree of propofol binding with blood cells can be different, depending on the blood type, and it can change in time, which may affect the results of the analysis. The paper discusses and shows the necessity of blood cell lysis before the extraction procedure. The cell lysis makes possible to determine the total amount of propofol in blood independently of the degree of propofol binding with cellular blood constituents and its changes. PMID- 11113932 TI - Stereoselective determination of propafenone enantiomers in transgenic Chinese hamster CHL cells expressing human cytochrome P450. AB - An enantioselective assay for S(+)- and R(-)-propafenone in transgenic Chinese hamster CHL cells expressing human cytochrome P450 was developed. The method involved extraction of propafenone from the S9s incubates, and formation of propafenone diastereomeric derivatives with the chiral reagent 2,3,4, 6-tetra-O beta-D-glucopranosyl isothiocyanate. Separation and quantitation of diastereomeric propafenone derivatives were carried out in a reverse-phase-HPLC system with UV detection. The assay was linear from 2 to 200 microg/mL for each enantiomer. The analytical method gave average recoveries of 97.5% and 97.0% for S(+)- and R(-)-propafenone, respectively. The limits of detection and quantitation for the method are 0.1 and 2.0 microg/mL for both S(+)- and R(-) propafenone, respectively. The reproducibility of the assay was good (RSD <10%). The method allowed study of the depletion of S(+)- and R(-)-propafenone in transgenic Chinese hamster CHL cells expressing human cytochrome P450. The stereoselectivity of propafenone phase I metabolism via cDNA-expressed CYP3A4 was observed. PMID- 11113933 TI - Proposal of sampling process for collecting human sweat and determination of caffeine concentration in it by using GC/MS. AB - Caffeine concentration in human sweat was estimated by measuring separately the amounts of water and caffeine. After washing a finger with tap water for 15 s and waiting 2 min for drying, 70 microL aqueous ethanol solution in a small vial (0.6 mL) was used to sample for several minutes. Then 3 microL of the aliquot was used for GC/MS analysis of caffeine. As a first-order relationship between the sweat amount secreted on the left and right hands was obtained (correlation factor 0.848), the amount of sweat secretion during sampling on one hand was estimated by the value obtained on the other hand. This new indirect evaluation was used for the estimation of the amount of sweat secreted during sampling. Typical variations of caffeine concentration in sweat were demonstrated. Thirty minutes after the intake of caffeine, it was secreted in sweat, and the secretion had continued for more than 4 h. PMID- 11113934 TI - Alkaline treatment of the cellulose fiber affecting membrane column behaviour for high-performance immunoaffinity chromatography. AB - The original cellulose fibers and those treated by alkaline solution were both used to prepare the acrylic membranes. The two kinds of membranes were packed into the columns for high-performance immunoaffinity chromatography by the immobilization of protein A on them. It was observed that the alkaline treatment of the cellulose fiber decreased the pressure resistance of the membrane to the mobile phases and greatly increased the accessible volume to the proteins, but affected the adsorption capacity of human IgG on the protein A membrane columns less. There is little difference between those two kinds of membranes on the adsorption capacities of HIgG, which means that the alkaline treatment of the cellulose fiber only significantly changes the void volume inter-membrane, and the porosity and surface area of membrane less. Alkaline treatment of the cellulose fiber reduced the membrane-column efficiency significantly. Some typical examples for the immunoaffinity analysis of IgG from human and dog plasma on the protein A membrane columns are illustrated. PMID- 11113935 TI - Screening for pharmaceutically important taxoids in Taxus baccata var. Aurea corr. with CC/SPE/HPLC-PDA procedure. AB - Needles of 'the golden yew' Taxus baccata var. Aurea Corr. were extracted with methanol followed by pre-purification of the crude extract and column chromatographic (CC) separation on florisil in gradient mode (an increasing concentration of acetone in dichloromethane). The obtained fractions were concentrated and purified on silanized silica gel SPE cartridges and taxoids eluted with 75% methanol were analysed by HPLC-PDA procedure using Waters Symmetry C(18) column with gradient elution. The applied method enabled not only determination of four taxoids commonly occurring in yew extracts (10 deacetylbaccatin III, baccatin III, paclitaxel and cephalomannine), but also, on the basis of chromatographic behaviour and UV spectrum, 10-deacetylated taxoids (10-deacetylpaclitaxel, 10-deacetylcephalomannine, 7-xyloside-10 deacetylpaclitaxel and 10-deacetyltaxol C) could be detected together with 7-epi 10-deacetylpaclitaxel. From the needles of Taxus baccata var. Aurea Corr. the largest amounts isolated were of 10-deacetylbaccatin III, then 10 deacetylpaclitaxel and 7-xyloside-10-deacetylpaclitaxel, all compounds considered to be paclitaxel precursors in semisynthesis. The efficient mechanism of the separation of 10-acetylated taxoids from their 10-deacetylated derivatives on florisil on the basis of electron acceptor-electron donor interactions is discussed. PMID- 11113936 TI - A cellular automata model of chromatography. AB - Dynamic models of the behavior of solvent and solute molecules can be made using cellular automata. A chromatographic column was represented by use of a cellular automata grid of 43 x 200 spaces. Solvent (mobile phase), solute and stationary phase cells were designated to simulate the chromatographic situation. The movements of solute and solvent cells down the grid were monitored for different numbers of iterations, different flow rates and different affinities of the solutes for the stationary phase and the solvent for itself. The cellular automata dynamics were successfully able to model expected chromatographic behavior except in a few cases where the number of cells was not large enough to provide an average value reflective of the molecular situation. PMID- 11113937 TI - High performance liquid chromatography analysis of D-penicillamine by derivatization with N-(1-pyrenyl)maleimide (NPM). AB - D-Penicillamine (2-amino-3-mercapto-3-methylbutanoic acid), a well-known heavy metal chelator, is the drug of choice in the treatment of Wilson's disease and is also effective for the treatment of several disorders including rheumatoid arthritis, primary biliary cirrhosis, scleroderma, fibrotic lung diseases and progressive systemic sclerosis. The method proposed incorporates a technique, previously developed in our laboratory, that utilizes the derivatizing agent N-(1 pyrenyl)maleimide (NPM) and reversed-phase high-performance liquid chromatography (HPLC). The coefficients of variation for within-run precision and between-run precision for 500 nM standard D-penicillamine (D-pen) were 2.27% and 2.23%, respectively. Female Sprague-Dawley rats were given 1 g/kg D-pen i.p. and the amounts of D-pen in liver, kidney, brain and plasma were subsequently analyzed. This assay is rapid, sensitive and reproducible for determining D-pen in biological samples. PMID- 11113938 TI - Separation and determination of strychnine and brucine in Strychnos nux-vomica L. and its preparation by capillary zone electrophoresis. AB - A capillary zone electrophoresis method was developed for the separation and determination of strychnine and brucine in Strychnos nux-vomica L. and its preparation. The factors that could affect the separation were studied, such as the types and concentrations of electrolytes, pH, ionic strength and organic modifier. The optimum running buffer was 20 mmol/L of ammonium acetate containing 0.2 mol/L of glacial acetic acid (pH 3.64). The applied voltage was 25 kV and the wavelength of the UV detector was set at 214 nm. The established method with dopamine hydrochloride as internal standard was linear in the range of 5-100 microg/mL for both strychnine and brucine. The recovery was 102.96% for strychnine and 98.56% for brucine. The extracts of Strychnos nux-vomica and its preparation could be directly injected for analysis. PMID- 11113939 TI - Simultaneous HPLC of twelve monoamines and metabolites shows neuroblastoma cell line releases HVA and HIAA. AB - Neuroblastoma is a solid tumor occurring usually in children less than 5 years old. It has been difficult to distinguish neuroblastoma from other childhood tumors through morphological diagnosis. Urine homovanillic acid (HVA), which is a metabolite of dopamine, has been proposed as a diagnostic index. Although increased levels of a serotonin metabolite, 5-hydroxyindole-3-acetic acid (HIAA), have also been observed in urine samples of the patients, they were largely attributed to dietary amines. By using an HPLC system with electrochemical detection, which can simultaneously assay 12 monoamines and metabolites, we showed that HVA and HIAA are two of the most prominent monoamine metabolites in the medium after a neuroblastoma cell line (IMR-32) was cultured for 3 days. Moreover, we found that the levels of HVA and HIAA in the media are proportional to the cell densities. These results suggest that the levels of HVA and HIAA in tissue culture media, or in urine from patients whose dietary amines are well controlled, may provide a valuable diagnostic index for neuroblastoma. PMID- 11113940 TI - Group sequential designs for cure rate models with early stopping in favour of the null hypothesis. AB - Ewell and Ibrahim derived the large sample distribution of the logrank statistic under general local alternatives. Their asymptotic results enable us to extend several group sequential designs which allow for early stopping in favour of the null hypothesis to the setting in which the cure rate model is appropriate. In particular, we derive stopping rules for the cure rate model using conditional power, predictive power and repeated confidence intervals methods. We illustrate the methods proposed using a hypothetical phase III clinical trial which is typical for melanoma studies. PMID- 11113941 TI - Power comparisons for tests of trend in dose-response studies. AB - The Cochran-Armitage test for trend is a popular statistical procedure for detecting increasing or decreasing probabilities of response when a categorical exposure is ordered. Such associations may arise in a variety of biomedical research settings, particularly in dose-response designs such as carcinogenicity experiments. Previously, computing limitations mandated the use of the asymptotic trend test, but with the availability of new algorithms, increased computing power, and appropriate software the exact trend test is now a practical option. Nevertheless, the exact test is sometimes criticized on the grounds that it is conservative. In this paper we investigate the implications of this conservatism by comparing the true type I error and power of three alternative tests of trend the asymptotic test, the exact test and an admissible exact test proposed by Cohen and Sackrowitz. The computations are performed by an extension to the network algorithm of Mehta et al. This allows us to make precise power comparisons between the tests under any given design without resorting to simulation. We show how this tool can guide investigators in choosing the most appropriate test by considering the design of two-year carcinogenicity studies carried out by the National Toxicology Program. We additionally compare the tests for various other combinations of sample sizes and number of groups or levels of exposure. We conclude that the asymptotic test, while more powerful where it is valid, generally does not preserve the type I error. This violation of the a priori testing level can be greatly affected by imbalance in the data or unequal spacing of dose levels. PMID- 11113942 TI - Exact sequential tests for single samples of discrete responses using spending functions. AB - Sequential tests are increasingly used to reduce the expected sample size of trials in medical research. The majority of such methods are based on the assumption of normality for test statistics. In clinical trials yielding a single sample of discrete data, that assumption is often poorly satisfied. In this paper we show how a novel application of the spending function approach of Lan and DeMets can be used together with exact calculation methods to design sequential procedures for a single sample of discrete random variables without the assumption of normality. A special case is that of binomial data and the paper is illustrated by the design of a cytogenetic study which motivated this work. PMID- 11113944 TI - Reproducibility of the six-minute walking test in chronic heart failure patients. AB - The six-minute walking test (WT) is used in trials and clinical practice as an easy tool to evaluate the functional capacity of chronic heart failure (CHF) patients. As WT measurements are highly variable both between and within individuals, this study aims at assessing the contribution of the different sources of variation and estimating the reproducibility of the test. A statistical model describing WT measurements as a function of fixed and random effects is proposed and its parameters estimated. We considered 202 stable CHF patients who performed two baseline WTs separated by a 30 minute rest; 49 of them repeated the two tests 3 months later (follow-up control). They had no changes in therapy or major clinical events. Another 31 subjects performed two baseline tests separated by 24 hours. Collected data were analysed using a mixed model methodology. There was no significant difference between measurements taken 30 minutes and 24 hours apart (p = 0.99). A trend effect of 17 (1.4) m (mean (SE)) was consistently found between duplicate tests (p < 0.001). REML estimates of variance components were: 5189 (674) for subject differences in the error-free value; 1280 (304) for subject differences in spontaneous clinical evolution between baseline and follow-up control, and 266 (23) for the within-subject error. Hence, the standard error of measurement was 16.3 m, namely 4 per cent of the average WT performance (403 m) in this sample. The intraclass correlation coefficient was 0.96. We conclude that WT measurements are characterized by good intrasubject reproducibility and excellent reliability. When follow-up studies > or = 3 months are performed, unpredictable changes in individual walking performance due to spontaneous clinical evolution are to be expected. Their clinical significance, however, is not known. PMID- 11113943 TI - Comparison of competing risks failure time methods and time-independent methods for assessing strain variations in vaccine protection. AB - In a preventive vaccine efficacy trial of a vaccine for a genotypically and phenotypically diverse pathogen, it is important to assess if and how vaccine protection against infection or disease varies with characteristics of the exposing pathogen. Gilbert, Self and Ashby developed statistical methods for this problem when the outcome data are counts of the number of vaccinated and unvaccinated trial participants infected by each pathogen strain. However, in many vaccine trials time-to-case information is available, and the extent to which this information improves investigation of differential vaccine protection is unclear. We describe how cause-specific proportional hazards models and other popular competing risks failure time techniques can be applied to this problem. This includes new results on the assumptions required for these methods to give valid inferences about strain-specific vaccine efficacy, and a comparison of theoretical and finite-sample properties between these methods and the time independent methods. Theoretical considerations, a cholera vaccine trial example, and an extensive simulation study of a human immunodeficiency virus type 1 (HIV 1) vaccine trial show that information about failure times does not appreciably improve estimation or testing unless the pathogen has a high attack rate and the relative prevalence of pathogen strains shifts substantially during the trial follow-up period. An important implication is that practically optimal evaluation of strain-specific vaccine efficacy in HIV-1 vaccine trials will not require knowledge of infection times. PMID- 11113945 TI - Empirical Bayes approach to estimating the number of HIV-infected individuals in hidden and elusive populations. AB - In this paper we estimate the numbers of intravenous drug users (IVDUs) and commercial sex workers (CSWs) in Thailand infected with human immunodeficiency virus (HIV) who have not developed acquired immunodeficiency syndrome (AIDS) directly from the semi-annual HIV serosurveillance data of Thailand from June 1993 to June 1995. We propose a 'generalized removal model for open populations' for estimating HIV-infected population size within a hidden, elusive, and perhaps high-risk population group, for all sampling time when capture probabilities vary with time. We apply empirical Bayes methodology to the generalized removal model for open populations by using the Gibbs sampler, a Markov chain Monte Carlo method. No assumption on the size of the hidden population in question is needed to implement this procedure. The statistical method proposed here requires very little computing and only a minimum of two sets of serosurvey data to obtain an estimate, thereby providing a simple and viable option in epidemiological studies when either powerful computing facilities or abundant sampling data are lacking. PMID- 11113946 TI - Sympercents: symmetric percentage differences on the 100 log(e) scale simplify the presentation of log transformed data. AB - The results of analyses on log transformed data are usually back-transformed and interpreted on the original scale. Yet if natural logs are used this is not necessary--the log scale can be interpreted as it stands. A difference of natural logs corresponds to a fractional difference on the original scale. The agreement is exact if the fractional difference is based on the logarithmic mean. The transform y = 100 log(e)x leads to differences, standard deviations and regression coefficients of y that are equivalent to symmetric percentage differences, standard deviations and regression coefficients of x. Several simple clinical examples show that the 100 log(e) scale is the natural scale on which to express percentage differences. The term sympercent or s% is proposed for them. Sympercents should improve the presentation of log transformed data and lead to a wider understanding of the natural log transformation. PMID- 11113947 TI - A simple method for converting an odds ratio to effect size for use in meta analysis. AB - A systematic review may encompass both odds ratios and mean differences in continuous outcomes. A separate meta-analysis of each type of outcome results in loss of information and may be misleading. It is shown that a ln(odds ratio) can be converted to effect size by dividing by 1.81. The validity of effect size, the estimate of interest divided by the residual standard deviation, depends on comparable variation across studies. If researchers routinely report residual standard deviation, any subsequent review can combine both odds ratios and effect sizes in a single meta-analysis when this is justified. PMID- 11113948 TI - Repeated measures in clinical trials: analysis using mean summary statistics and its implications for design by L. Frison and S.J. Pocock, Statistics in Medicine 1992; 12: 1685-1704. PMID- 11113950 TI - Common sense and figures: the rhetoric of validity in medicine (Bradford Hill Memorial Lecture 1999). AB - Austin Bradford Hill was once a friend to The Lancet, but, as occasionally happens, friends fall out. The great legacy of his association with the journal, however, was Principles of Medical Statistics. As each edition was succeeded by another--the first in 1937, the last in 1991--he seemed to shift his view about the influence of statistical method on clinical practice from one of assured certainty to one of modest advantage. That change paralleled a move away from an emphasis on the importance of internal validity in the randomized trial to one of understanding the inescapably practical significance of generalizability. Writers on medical research have explored notions of external validity in various ways. One view, for example, is to seek a close correlation between the participants in a clinical trial and patients seen in practice. The argument goes that such a correspondence has to be made before any decision can be taken about whether to apply the result of that trial to the clinical setting. Another view, first worked out by the American logician Charles Sanders Peirce, is that one must simply rely on the informed guess, based on a reasonable estimate of the limits of extrapolation. The tensions between and implications of these two different approaches are worked through using the example of coronary stents. A solution is, perhaps, to write explicit rules of interpretation that provide a framework for judging the strength of a claim to applicability. Five questions are posed, which try to lay a foundation for such a framework. PMID- 11113951 TI - Reflections on statistics at the London School of Hygiene and Tropical Medicine 30 years ago. AB - A course leading to the Master of Science (MSc) degree in Medical Statistics was started at the London School of Hygiene and Tropical Medicine in 1968. The events leading up to this initiative are outlined in the context of earlier developments in statistics at the School and the general growth of opportunities in statistical education. PMID- 11113952 TI - Analysis and design issues for studies using censored biomarker measurements with an example of viral load measurements in HIV clinical trials. AB - For many biomarkers, the range (L,R) over which they can be quantified is restricted by technical limitations, leading to some measurements that are left or right censored. However, despite the widespread availability of statistical methods for the analysis of censored data, many studies use an imputed value for censored measurements (for example, replacing a value 23. Memory complaints in highly educated elderly subjects may be predictive of dementia even when there is no indication of cognitive impairment on short cognitive screen tests. The shift in methodology which is noticeable in the recently published major studies is discussed as a possible explanation for the established association between memory complaints and decline in memory (or dementia) in elderly subjects. Three methodological factors, in particular, are responsible for the results: community-based sampling, longitudinal design and the treatment of variables such as depression, cognitive impairment and level of education. CONCLUSION: Memory complaints in elderly people should no longer be considered merely as an innocent age-related phenomenon or a symptom of depression. Instead, these complaints deserve to be taken seriously, at least as a possible early sign of dementia. PMID- 11113978 TI - Dementia: the cost of care for behaviourally disturbed patients living in the community. AB - OBJECTIVE: To determine the cost of care of people with dementia and behavioural disturbances living in the community, which is currently unknown. DESIGN, SETTING & PATIENTS: This is a study of 12 randomly selected subjects attending a continuing care dementia day facility for people with behavioural disturbances. MEASURES: Hours of care received from different sources were recorded and costed. RESULTS: The total mean direct costs to the public services was pound400 per person per week. Caregiver's time costed at professional rates were pound1208 per person per week. CONCLUSIONS: Unpaid caregivers supported by publicly funded agencies provide a less expensive service compared to specialised institutional care. PMID- 11113977 TI - Symptoms of striatofrontal dysfunction contribute to disability in geriatric depression. AB - OBJECTIVE: To examine whether symptoms of striatofrontal dysfunction contribute to disability in geriatric depression. DESIGN: Cross-sectional evaluation of the relationship of specific cognitive impairments, psychomotor retardation, severity of depression, and medical burden to impairment of instrumental activities of daily living. SETTING: Inpatient and outpatient services of a psychiatric university hospital located in a suburban metropolitan area.Patients. One hundred and fifty elderly psychiatric inpatients and outpatients with major depression and cognitive function ranging from normal to moderate dementia. MEASURES: Psychomotor retardation was evaluated with the Hamilton retardation item and executive dysfunction was assessed with the initiation/perseveration (IP) domain of the Dementia Rating Scale. Disability, severity of depression and medical burden were assessed with the Instrumental Activities of Daily Living Index of the Multilevel Assessment Instrument, the Hamilton Depression Rating Scale and the Cumulative Illness Rating Scale-Geriatric, respectively. RESULTS: In the entire sample (N = 150) and in the non-demented subjects (N = 101), stepwise regression analyses revealed that IP and psychomotor retardation were associated with IADL impairment. Additionally, a 'striatofrontal component', which consisted of IP and psychomotor retardation was also significantly associated with IADL impairment in the whole sample, as well as in the non-demented patients. CONCLUSION: Clinical symptoms and neuropsychological findings associated with striatofrontal dysfunction contribute to disability in depressed elderly patients. PMID- 11113979 TI - End of life treatment decisions in people with dementia: carers' views and the factors which influence them. AB - OBJECTIVE: Treatment decisions in life threatening situations (TD) are poorly studied in people with dementia. METHOD: The carers of people with dementia were asked four TD questions, pertaining to cardiac resuscitation, intravenous fluids, oral antibiotics and intravenous antibiotics. The impact of key variables (age, dementia severity, psychiatric co-morbidity, physical illness, family relationship of carer) on TD were evaluated. RESULTS: Fifty carers participated, 46% wanted cardiac resuscitation, 60% wanted treatment with intravenous fluids, 52% wanted treatment with intravenous antibiotics and 60% wanted treatment with oral antibiotics. Agreement between questions was high (76 - 89%), suggesting that relatives were either for or against intervention. There was an association between more severe dementia and a reduced wish for intravenous antibiotics. None of the variables significantly influenced other TD. CONCLUSION: The 'global' view of carers, was not influenced greatly by key disease variables. There are potential implications for the way in which carers are used as proxy decision makers. PMID- 11113980 TI - Age and nortriptyline concentrations in plasma ultrafiltrate. AB - OBJECTIVES: Since plasma protein binding of tricyclic antidepressants may be relevant to treatment effects and can be influenced by age-associated factors, we examined both plasma ultrafiltrate and total concentrations of nortriptyline (NT) in patients and compared these to age. We hypothesized negative associations with age of both ultrafiltrate NT and the ratio of ultrafiltrate NT to total plasma NT. METHODS: Patients with major depression at a psychiatric service treated with a stable dose of NT were studied. Trough plasma ultrafiltrate NT concentrations and total plasma NT concentrations were measured by high performance liquid chromatography. Concentrations were corrected for dose. RESULTS: Eighty-seven patients aged 26 - 88 years were studied. Ultrafiltrate NT concentrations and the ratio of ultrafiltrate NT to total NT concentrations were both significantly negatively associated with age. Total NT concentrations were not significantly associated with age. CONCLUSIONS: Relatively low ultrafiltrate NT concentrations in older patients may reflect lower tissue exposure at a given total plasma NT concentration. This could be relevant to toxic and therapeutic effects. Studies of relationships between non-bound drug concentrations and NT treatment outcomes across the age span are needed. PMID- 11113981 TI - Resident characteristics associated with wandering in nursing homes. AB - OBJECTIVES: This retrospective cohort study examined the association between resident characteristics and the development of wandering behavior. METHODS: Subjects included a total of 8982 residents from the states of Mississippi, Texas, and Vermont who had baseline and 3-month follow-up Minimum Data Set assessments between 1 January 1996 and 31 December 1997. RESULTS: Residents who had a short-term memory problem (Odds Ratio (OR) = 3.05), had pneumonia (OR = 3.15), asked repetitive questions (OR = 2.19), had a long-term memory problem (OR = 2.06), exhibited dementia (OR = 19.4), constipation (OR = 1.82), expressed sadness or pain (OR = 1.65), and used antipsychotic medication (OR = 1.70), were at an increased risk for developing wandering behavior compared to residents without these characteristics. Residents with functional impairment (OR = 0.28) and women (OR = 0.61) were less likely to develop wandering behavior. CONCLUSIONS: Results of this study may be useful in constructing causal theories for the development of wandering behavior. PMID- 11113982 TI - The mini-cog: a cognitive 'vital signs' measure for dementia screening in multi lingual elderly. AB - OBJECTIVES: The Mini-Cog, a composite of three-item recall and clock drawing, was developed as a brief test for discriminating demented from non-demented persons in a community sample of culturally, linguistically, and educationally heterogeneous older adults. SUBJECTS: All 129 who met criteria for probable dementia based on informant interviews and 120 with no history of cognitive decline were included; 124 were non-English speakers. METHODS: Sensitivity, specificity, and diagnostic value of the Mini-Cog were compared with those of the Mini-Mental State Exam (MMSE) and Cognitive Abilities Screening Instrument (CASI). RESULTS: The Mini-Cog had the highest sensitivity (99%) and correctly classified the greatest percentage (96%) of subjects. Moreover, its diagnostic value was not influenced by education or language, while that of the CASI was adversely influenced by low education, and both education and language compromised the diagnostic value of the MMSE. Administration time for the Mini Cog was 3 minutes vs 7 minutes for the MMSE. CONCLUSIONS: The Mini-Cog required minimal language interpretation and training to administer, and no test forms of scoring modifications were needed to compensate for the extensive linguistic and educational heterogeneity of the sample. Validation in clinical and population based samples is warranted, as its brevity and ease of administration suggest that the Mini-Cog might be readily incorporated into general practice and senior care settings as a routine 'cognitive vital signs' measure. PMID- 11113983 TI - A comparison of sleep profiles in patients with dementia with lewy bodies and Alzheimer's disease. AB - INTRODUCTION: Sleep disturbances are common in healthy old age and in dementia syndromes. Polysomnography has demonstrated typical changes in both Alzheimer's disease (AD) and dementia with Lewy bodies (DLB) with AD being characterised by sundowning and sleep apnoea and DLB patients showing more disturbances of movement control during sleep. The technical difficulties associated with EEG sleep recordings mean that polysomnography is not possible out of specialist centres. OBJECTIVES: To use questionnaires to assess the frequency of sleep disturbances in patients with Alzheimer's disease and dementia with Lewy bodies. METHOD: The sleep profiles of twenty patients with AD and 17 with DLB were assessed using three questionnaires, one designed to assess night time sleep disturbance, one day time sleepiness and the last carer burden. The sleep questionnaires were repeated in a subgroup after treatment with a cholinesterase inhibitor (rivastigmine). RESULTS: Level of sleep disturbance in both groups was high. DLB patients had more overall sleep disturbance, more movement disorders whilst asleep and more abnormal day time sleepiness. Treatment with rivastigmine produced a trend towards normalisation of sleep profile in a small number of subjects. CONCLUSIONS: Both groups have extensive sleep problems. The DLB and AD groups have different sleep profiles that are of diagnostic importance and may suggest different treatment strategies. The results are consistent with those found from polysomnographic assessment and suggest that the questionnaires used are sensitive to detect differences previously documented with polysomnography. PMID- 11113984 TI - Dementia with lewy bodies: findings from an international multicentre study. AB - OBJECTIVES: To describe the baseline demographic, neuropsychiatric and neurological data of a large selected clinical sample of patients with dementia with Lewy Bodies (DLB) from an international multicentre trial with rivastigmine. To examine the usefulness of the Consensus Criteria for the diagnosis of DLB in different countries. METHODS: Seventeen centres from Spain, the UK and Italy recruited patients diagnosed clinically as probable DLB according to recent Consensus Criteria (McKeith et al., 1996). A standard clinical protocol including inclusion/exclusion criteria, collection of demographic and medical data, cognitive (Mini Mental State Examination: MMSE), motor (Unified Parkinson's Disease Rating Scale: UPDRS) and neuropsychiatric (Neuropsychiatric Inventory: NPI) examinations, was applied after obtaining informed consent. Data were summarised and compared across countries with uni- and multivariate analyses. RESULTS: One hundred and twenty patients were recruited: 56.7% males, mean (SD) age 73.9 (6.4) years, range 57 - 87 years. Sixty percent fulfilled all three core diagnostic features of DLB, and 40% only two ('parkinsonism' 92.4%, 'cognitive fluctuations' 89.1%, 'visual hallucinations' 77.3%). 'Systematised delusions' (46%) and 'repeated falls' (42%) were the most frequent supportive diagnostic features. There were no differences across countries in demographic, diagnostic or clinical features. Patients showed a wide range of psychopathology which was weakly correlated with cognitive impairment. Some mild extrapyramidal signs (EPS) were observed in most patients. CONCLUSIONS: The Consensus Criteria for DLB can be consistently applied across many different sites for multicentre studies. 'Parkinsonism' and 'cognitive fluctuations' as core features and 'systematised delusions' and 'repeated falls' as supportive features are the most frequent diagnostic clues. Neuropsychiatric disturbances, in particular apathy, delusions, hallucinations and anxiety, and mild symmetric EPS are frequent in DLB and are only related weakly to cognitive impairment. PMID- 11113985 TI - Mental health of elderly Asians in Britain: a comparison of Hindus from nuclear and extended families of differing cultural identities. AB - OBJECTIVE: To compare the psychological adjustment of grandmothers from nuclear and extended families within British Hindu communities, and to investigate the influence of cultural identity. DESIGN: Interviews were carried out with Hindu grandmothers, mothers and granddaughters living in both nuclear and extended families. SETTING: The sample was drawn from the total population of Asian Hindu girls aged between 13 and 17 years at four comprehensive schools in the London Borough of Redbridge. PARTICIPANTS++: The final sample consisted of 36 and 34 sets of individuals in extended and nuclear families, respectively. MEASURES: Questionnaire measures were obtained of the following variables: cultural integrity, traditionalism, religious participation, ethnic identity, anxiety, depression, self-esteem. RESULTS: Grandmothers were better adjusted in extended families than in nuclear families. This adjustment was in part mediated by the level of traditional belief within the family. Elders whose granddaughters had an exclusively 'Indian' or 'Hindu' ethnic identity were better adjusted than those whose granddaughters included a 'British' ethnic identity. CONCLUSIONS: This study confirmed findings from earlier studies that grandmothers in extended families were significantly better adjusted in comparison to those from nuclear families. Ethnic identity of the adolescent, independent of its salience and commitment to the adolescent, had a significant relationship to the grandmother's mental health. PMID- 11113986 TI - A retrospective study of neuroradiological abnormalities detected on structural magnetic resonance imaging of the brain in elderly patients with cognitive impairment. AB - BACKGROUND: The aim of the study was to investigate the relationship between neuroradiological and clinical diagnosis in patients presenting with cognitive impairment, and also the relationship between the neuroradiological abnormalities and cognitive function as assessed by the Mini-Mental State Examination (MMSE) score. METHODS: One hundred and four elderly subjects (65 years and over) with cognitive impairment, referred to secondary hospital services and who had brain magnetic resonance imaging (MRI) scans as part of routine clinical investigations, were studied by review of their MRI scans using a standardized procedure and by examination of the case notes. RESULTS: In patients with a clinical diagnosis of senile dementia Alzheimer-type (SDAT), the diagnosis was reviewed in 11.1%. In patients with vascular dementia, the diagnosis was reviewed in 62. 5%. In patients without a firm clinical diagnosis, radiological features compatible with SDAT were seen in 44.4% and with vascular dementia in 27.0%. Only 2/104 patients showed a significant focal lesion on MRI. Of the variables studied (age, sex, degree of hippocampal atrophy, extent of T2 hyper-intensities, and enlargement of the sulcal and ventricular cerebrospinal fluid (CSF) spaces) only hippocampal atrophy predicted the MMSE score ( p < 0.002). CONCLUSION: MRI brain scanning has an important role in aiding and refining the clinical diagnosis of cognitive impairment/dementia in the elderly. PMID- 11113987 TI - Gender differences in survival of 234 patients referred to a psychogeriatric service. AB - OBJECTIVE: To explore the survival characteristics of psychogeriatric patients. Participants and settingAn historical cohort of 234 patients consecutively referred to a specialized psychogeriatric service proximal to a general health service in Perth, Western Australia. METHODS: Linked health service data were analysed using relative survival analysis and actuarial methods. RESULTS: Relative survival of the cohort after 40 months was significantly lower than the general population of the same age, sex and calendar period (0.78; 95% CI=0.70 0.86). Male patients experienced twice the mortality rate of female patients after adjustment for age (MRR=2.10; 95% CI=1.37-3.20). Age, dementia, mood disorder and ethnicity had no independent effects on mortality in male patients. Female patients with a diagnosis of dementia experienced twice the mortality of female patients without this diagnosis. The distribution of major underlying causes of death was similar in males and females. CONCLUSIONS: Gender-specific factors appear to affect survival in psychogeriatric patients. Male patients are younger on average, but experience higher mortality than female patients. Ethnic background does not influence mortality in either male or female patients. PMID- 11113988 TI - Travel-induced psychosis in the elderly. AB - We present three cases of travel-induced psychosis in an elderly population. The clinical presentations are described with a review of the literature. PMID- 11113989 TI - Do elderly victims of fatal self-harm with history of deliberate self-harm use the same methods in their final act as they did in previous attempts? PMID- 11113991 TI - Current awareness in geriatric psychiatry. PMID- 11113990 TI - Evaluation of the Chinese version of the Psychogeriatric Assessment Scales (PAS) PMID- 11113992 TI - The plant kingdom as a source of anti-ulcer remedies. AB - Phytogenic agents have traditionally been used by herbalists and indigenous healers for the prevention and treatment of peptic ulcer. This article reviews the anti-acid/anti-peptic, gastro-protective and/or anti-ulcer properties of the most commonly employed herbal medicines and their identified active constituents. Botanical compounds with anti-ulcer activity include flavonoids (i.e. quercetin, naringin, silymarin, anthocyanosides, sophoradin derivatives) saponins (i.e. from Panax japonicus and Kochia scoparia), tannins (i.e. from Linderae umbellatae), gums and mucilages (i.e. gum guar and myrrh). Among herbal drugs, liquorice, aloe gel and capsicum (chilli) have been used extensively and their clinical efficacy documented. Also, ethnomedical systems employ several plant extracts for the treatment of peptic ulcer. Despite progress in conventional chemistry and pharmacology in producing effective drugs, the plant kingdom might provide a useful source of new anti-ulcer compounds for development as pharmaceutical entities or, alternatively, as simple dietary adjuncts to existing therapies. PMID- 11113993 TI - Beneficial effects of flavonoids from Sesamum indicum, Emblica officinalis and Momordica charantia. AB - Flavonoids from Sesamum indicum (gingili), Emblica officinalis (gooseberry) and Momordica charantia (bittergourd) were analysed for their biological activities. Of the three sources, flavonoids isolated from Emblica officinalis exerted the maximum beneficial action by eliciting highly potent hypolipidaemic and hypoglycaemic activities. Moreover these flavonoids were effective in raising the haemoglobin levels in rats. PMID- 11113994 TI - Inhibitory effects of cimicifugae rhizoma extracts on histamine, bradykinin and COX-2 mediated inflammatory actions. AB - Rhizoma Cimicifugae (RC) has been used traditionally to treat pain and inflammation in Korea. The present study was conducted to gain insights into the mechanism of action regarding analgesic and antiinflammatory activities of RC extracts. RC was first extracted with methanol. The methanol extract (A) was fractionated to an ether-soluble fraction (B) and a water-soluble fraction (C). Fraction C was fractionated to a butanol-soluble fraction (D) and a water-soluble fraction (E). Each fraction (100 mg/kg, i.p.) was tested for analgesic and antiinflammatory activities. Administration of fractions A and D caused dramatic analgesic effects based on acetic acid writhing and tail-flick assays. However, fraction E had an analgesic effect only based on the acetic acid writhing assay. Fractions A, D and E exerted antiinflammatory effects on the rat paw oedema assay. The fractions A, D, E had an inhibitory action on the bradykinin/histamine mediated contractions of guinea-pig ileum. In addition, fractions A, D and E had the ability to inhibit the production of LPS-induced 6-keto-PGF1alpha production in macrophage cultures. Taken together, these results provide scientific evidence that RC extracts exert analgesic and antiinflammatory effects by inhibiting bradykinin/histamine mediated actions and inhibiting 6-keto-PGF1alpha induction. PMID- 11113995 TI - Antibacterial activity of plant extracts from the families Fabaceae, Oleaceae, Philadelphaceae, Rosaceae and Staphyleaceae. AB - The selected plant extracts exhibited antibacterial activity. The strongest effect was manifested by extracts prepared from Gymnocladus dioicus, Amelanchier ovalis, Exochorda racemosa, Holodiscus discolor, Philadelphus microphyllus, Philadelphus coronarius and Pelargonium tabulare. The percentage inhibition of bacterial growth was 0-41.8%. In addition it was found that extracts isolated from Amelanchier ovalis, Exochorda racemosa and Pelargonium tabulare were specifically effective only against the bacterial strains tested. PMID- 11113996 TI - Search for antiviral activity in higher plant extracts. AB - In the course of our search for plant natural products as antiviral agents, extracts of ten plants from the Iberian Peninsula were tested for antiviral activity against herpes simplex type I (HSV-1), vesicular stomatitis virus (VSV) and poliovirus type 1. Aqueous extracts of five of these medicinal plants, namely Nepeta nepetella (150-500 microg/mL), Nepeta coerulea (150-500 microg/mL), Nepeta tuberosa (150-500 microg/mL), Dittrichia viscosa (50-125 microg/mL) and Sanguisorba minor magnolii (50-125 microg/mL), showed a clear antiviral activity against two different DNA and RNA viruses, i.e. HSV-1 and VSV. Only the medicinal plant Dittrichia viscosa was active against an additional virus, poliovirus type 1. PMID- 11113997 TI - Toxicity and genotoxicity of antimalarial alkaloid rich extracts derived from Mitragyna inermis O. Kuntze and Nauclea latifolia. AB - The toxicity and the genotoxicity of antimalarial alkaloid rich extracts derived from two plants used in traditional medicine in Mali (Mitragyna inermis (Willd.) O. Kuntze Rubiaceae and Nauclea latifolia (Sm.) Rubiaceae) were evaluated on in vitro and in vivo systems. The results demonstrated that an alkaloid rich extract derived from M. inermis induced a strong inhibition of protein synthesis in mammalian cells but did not exhibit mutagenic or genotoxic activity. An alkaloid rich extract derived from N. latifolia could interact in vitro with DNA of bacteria and mammalian cells, leading to G2-M cell cycle arrest and heritable DNA damage, as well as inducing in vivo single-strand breaks in liver, kidney and blood cells. PMID- 11113998 TI - Antioxidants in medicinal plant extracts. A research study of the antioxidant capacity of Crataegus, Hamamelis and Hydrastis. AB - The antioxidant capacity of extracts of Crataegus oxyacantha, Hamamelis virginiana, Hydrastis canadensis, plants native to Europe and North America which have long been used in herbal medicine for the treatment of cardiac and circulatory functions, has been investigated. The total antioxidant potential conferred by all hydrogen donating antioxidants present in these extracts has been assessed by the ABTS assay and the relative order of antioxidant potential has been established. Gas chromatography coupled to mass spectrometry (GC-MS) has been used for the chemical identification of the antioxidant volatile compounds present in the extracts. The GC-MS data were related to the results obtained using the ABTS assay. PMID- 11114000 TI - Pharmacological and antimicrobial studies on different tea-tree oils (Melaleuca alternifolia, Leptospermum scoparium or Manuka and Kunzea ericoides or Kanuka), originating in Australia and New Zealand. AB - Three different species of Myrtaceae growing in Australia and New Zealand are known as 'Tea-tree': the Australian Tea tree (Melaleuca alternifolia), the New Zealand Manuka (Leptospermum scoparium) and Kanuka (Kunzea ericoides). All three essential oils are used by aromatherapists, although only Melaleuca has been tested for toxicity, and its antimicrobial effects studied. The pharmacology and antimicrobial activity of the three 'tea-tree' oils was determined using guinea pig ileum, skeletal muscle (chick biventer muscle and the rat phrenic nerve diaphragm) and also rat uterus in vitro. Differences were shown between the three essential oils in their action on smooth muscle: Manuka had a spasmolytic action, while Kanuka and Melaleuca had an initial spasmogenic action. Using the diaphragm, Manuka and Melaleuca decreased the tension and caused a delayed contracture; Kanuka had no activity at the same concentration. The action on chick biventer muscle was, however, similar for all three oils, as was the action on the uterus, where they caused a decrease in the force of the spontaneous contractions. The latter action suggests caution in the use of these essential oils during childbirth, as cessation of contractions could put the baby, and mother, at risk. The comparative antimicrobial activity showed greater differences between different samples of Manuka and Kanuka than Melaleuca samples. The antifungal activity of Kanuka was inversely proportional to its strong antibacterial activity, whilst Manuka displayed a stronger antifungal effect, though not as potent as Melaleuca. The antioxidant activity of Manuka samples was more consistent than that of Kanuka, while Melaleuca showed no activity. The variability in the Manuka and Kanuka essential oils suggests caution in their usage, as does the fact that the oils have not been tested for toxicity. PMID- 11113999 TI - Immunoenhancing properties of Plantago major leaf extract. AB - Plantago major (PM), also known as plantain, is a weed found in temperate zones worldwide. PM leaves have been associated with various biological properties ranging from antiinflammatory, antimicrobial and antitumour to wound healing. However, its mechanism of action associated with boosting of the immune function remains to be elucidated. We found that endotoxin-free methanol extracts from PM leaves, at doses of 50, 100, 250, and 500 microg/mL, were associated with 4.4 +/- 1, 6 +/- 1, 12 +/- 0.4, and 18 +/- 0.4-fold increases of nitric oxide (NO) production, and increased TNF-alpha production (621 +/- 31, 721 +/- 36, 727 +/- 36, and 1056 +/- 52 U/mL, respectively) by rat peritoneal macrophages, in the absence of IFN-gamma or LPS. NO and TNF-alpha production by untreated macrophages was negligible. In addition, PM extracts potentiated Con A-induced lymphoproliferation (3- to 12-fold increases) in a dose-dependent fashion, compared with the effect of Con A alone. The regulation of immune parameters induced by plant extracts may be clinically relevant in numerous diseases including chronic viral infections, tuberculosis, AIDS and cancer. PMID- 11114001 TI - Inhibition of glutathione S-transferases (GSTs) from parasitic nematodes by extracts from traditional Nigerian medicinal plants. AB - Piliostigma thonningii, Ocimum gratissimum, Nauclea latifolia and Alstonia boonei are used in Nigerian traditional medicines against gastrointestinal helminths of animals and man. Proanthocyanidins were detected in Piliostigma and Nauclea, but not Alstonia or Ocimum. Extracts of these plants killed 50% of brine shrimp nauplii at <10 ppm (Nauclea), 100 ppm (Piliostigma) and <1000 ppm (Ocimum and Alstonia), the Nauclea LD50 being similar to the anthelmintic drug piperazine. Extracts were also toxic to the parasitic nematode Haemonchus infective L3 stage. Nematode glutathione-S-transferases (GSTs) are potential drug targets. Apart from Alstonia all the medicinal plants contained heat-stable inhibitory activities against recombinant Ascaris and Onchocerca GSTs in vitro. Piliostigma, Ocimum and Nauclea had IC50s of 2, 10 and 15 microg/mL respectively for Ascaris GST and 4, 8, 28 microg/mL respectively for Onchocerca GST. We suggest that the inhibitory properties of some of these Nigerian plant extracts against GST may contribute to the pharmacological basis of their efficacy against helminths in traditional herbal use. PMID- 11114002 TI - Analgesic and antiinflammatory activities of an extract from Parkia biglobosa used in traditional medicine in the Ivory Coast. AB - In the Ivory coast, Parkia biglobosa (Mimosaceae) is used in traditional medicine as an analgesic drug, especially against dental pain. Of the three extracts obtained from the plant bark, the hexane fraction was studied to determine its analgesic and/or antiinflammatory activities. The results show that this extract possesses a marked analgesic activity when evaluated with the abdominal writhing test in mice, but, like paracetamol, was ineffective with the hot-plate method, a feature suggesting a peripheral mechanism of action. This activity was accompanied by an antiinflammatory effect, somewhat weaker than the analgesic one. PMID- 11114004 TI - Biologically active steroid from the green alga Ulva lactuca. AB - The marine green alga, Ulva lactuca, was shown to contain 3-O-beta-D glucopyranosyl-stigmasta-5,25-dien. The structure of the compound was established on the basis of its spectroscopic data and it was extracted for the first time from this alga. The topical antiinflammatory activity of this compound was examined using the mouse ear oedema assay as an experimental model of topical inflammation. Also, the antimicrobial activity of the isolated compound was tested against 10 various microorganisms (G+, G-, fungi and yeast strains). PMID- 11114003 TI - Antiinflammatory activity of extracts and fractions from the leaves of Gochnatia polymorpha. AB - The aqueous and ethanol extracts from the leaves of Gochnatia polymorpha and further fractions obtained from the latter extract using solvents with increasing polarity, including its aqueous residue and the amino acid, 4-hydroxy-N-methyl proline were investigated by carrageenin-induced pedal oedema formation. It was shown that the aqueous and ethanol extracts and the ethyl acetate fraction demonstrated significant antiinflammatory activity. The chemical investigation of the latter fraction revealed the presence of caffeic acid, chlorogenic acid, 3-0 methylquercetin, hyperosid and rutin. The amino acid 4-hydroxy-N-methyl-proline, a nonprotein amino acid that has not been reported before in the Asteraceae was isolated as a major compound and identified by spectroscopic methods. PMID- 11114005 TI - A simple microbiological assay for the stereospecific differentiation of alpha and beta isomers of arteether. AB - A new rapid bioassay has been developed which can precisely differentiate between stereospecific alpha and beta isomers of the antimalarial drug arteether. This method was developed through the disc diffusion bioactivity tests wherein semisynthetically produced alpha arteether was able to inhibit the growth of E. coli strains which are defective in DNA gyrase enzyme. The wild type E. coli with intact DNA gyrase did not show this sensitivity to alpha arteether. The beta isomer of arteether was, however, ineffective against both the mutant and wild type strains. Direct experimental proof of gyrase involvement was obtained through mobilization of gyr genes by transformation of E. coli gyr- mutant strains with wild type gyrA clone pMK90 (carried on the thermo-inducible lambda Col E1 vector). This resulted in alpha arteether resistant and nalidixic acid sensitive phenotype clearly demonstrating the use of gyrA mutant strains in differentiating alpha and beta isomers of arteether by this simple bioassay. PMID- 11114006 TI - Antioxidant activity of the extracts from fruiting bodies of cultured Cordyceps sinensis. AB - Cordyceps sinensis is one of the most valued herbs in traditional Chinese medicine. We investigated the antioxidant activities of the cultured fruiting bodies of Cordyceps sinesis. The water and ethanol extracts of Cordyceps sinensis were found to possess a potent antioxidant activity. The scavenging effects of the extracts on superoxide were very weak, but the extracts moderately inhibited malondialdehyde formation via hydroxyl radical induced by SIN-1, a peroxynitrite generator. Of the extracts examined, the hot water extract (70 degrees C for 5 min) showed the greatest oxygen free radical scavenging activity. Also, when low density lipoprotein (LDL) was incubated with macrophages in the presence of CuCl2 (1 microM), the hot water extract showed a strong inhibitory effect against lipid peroxidation in the medium and consequent accumulation of cholesteryl ester in macrophages. Their activities were comparable to that of authentic Cu/Zn SOD. These results suggest that the extracts of cultured Cordyceps sinensis possess potent antioxidant and anti-lipid peroxidation activities and inhibit accumulation of cholesteryl ester in macrophages via suppression of LDL oxidation. PMID- 11114007 TI - Inhibitory effects of water extracts from fruiting bodies of cultured Cordyceps sinensis on raised serum lipid peroxide levels and aortic cholesterol deposition in atherosclerotic mice. AB - We investigated the effects of the water extracts of the fruiting bodies of cultured Cordyceps sinensis (WECS) on lipid metabolism in mice fed an atherogenic diet. WECS was orally administered at doses of 50, 100 and 200 mg/kg/day for 12 weeks. WECS showed no toxic effects on the growth rate, liver or kidney weights of the mice. Mice fed the atherogenic diet showed marked increases in serum lipid and lipid peroxide levels and also aortic cholesterol levels, particularly cholesteryl ester level, a major lipid constituent in atherosclerotic lesions. WECS significantly suppressed the increased serum lipid peroxide level but not other lipid levels in a dose-dependent manner. WECS also suppressed the increased aortic cholesteryl ester level in a dose-dependent manner. These results suggest that WECS prevents cholesterol deposition in the aorta by inhibition of LDL oxidation mediated by free radicals rather than by reduction in serum lipid level. WECS may exert beneficial effects on the formation of the atherosclerotic lesion induced by oxidative stress with few side effects. PMID- 11114008 TI - New protopine alkaloids from Aristolochia constricta reduce morphine withdrawal in vitro. AB - The present study examines the effect of four new protopine alkaloids (1-4) isolated and purified from the aerial parts of Aristolochia constricta (Aristolichiaceae) on morphine withdrawal in vitro. The results of our experiments indicate that the pure compounds (1-4) significantly and in a concentration-dependent manner reduced the morphine withdrawal. The results of the present study suggest that these new protopine alkaloids may be potential anti-addictive agents. PMID- 11114010 TI - Patents alert PMID- 11114011 TI - Selected bibliography PMID- 11114012 TI - R. Daudt, G. L. Von poser, G. Neves and S. M. K. Rates, 'Screening for the antidepressant activity of some species of hypericum from south Brazil'. Phytotherapy research14(5) 2000, 344-346 AB - The original article to which this Erratum refers was published in Phytotherapy Research 14(5) 2000, 344-346. Following the publication of this paper in the August 2000 issue of Phytotherapy Research (14(5):344-346), it has come to our attention that there is a misleading statement regarding conclusions cited from the work of Butterweck et al. 1998. The discussion in the recent PTR paper states that those authors are 'in favour of the hypothesis that the antidepressant activity is due to the hypericin only'. We wish to make it clear that this is not the case, and the Butterweck paper actually concludes that 'both naphthodianthrones must be considered as active constituents of the crude extract of H. perforatum. However, previous studies indicate that the other consitutuents of the crude drug also have activity'. The authors apologize for this error and are happy to correct it. PMID- 11114013 TI - S. K. Adesina, O. Idowu, A. O. Ogundaini, H. Oladimeji, T. A. Olugbade, G. O. Onawunmi and M. Pais, 'Antimicrobial constituents of the leaves of EAcalypha wilkesiana and acalypha hispida'. Phytotherapy research14(5) 2000, 371-374 AB - The original article to which this Erratum refers was published in Phytotherapy Research 14(5) 2000, 371-374. Following publication of this paper in the August 2000 issue of Phytotherapy Research (14(5):371-374), it has come to our attention that the dates of receipt and acceptance were printed incorrectly. The corrected dates appear below. The publishers would like to apologise for any confusion caused. Received 28 August 1996 Accepted 15 June 1997 PMID- 11114014 TI - Analysis of neutral isomeric low molecular weight carbohydrates using ferrocenyl boronate derivatization and tandem electrospray mass spectrometry. AB - A new analytical technique for small carbohydrates utilizing the cyclic ferrocenyl boronic esters (FcBors) of several neutral mono- and disaccharides is demonstrated. Distinction between the diastereomers of mono- and disaccharides is obtained. Analysis is by tandem electrospray-mass spectrometry (ES-MS) using a modified ion-source that promotes the preformation of ions. Selection of the molecular ion produced during single-electron oxidation of the ferrocene moiety of a specific population of saccharide isomers permits a variety of collisionally induced dissociation (CID) experiments. The resultant MS2/MS3 spectra reflect the ensemble of possible cyclic esters in equilibrium. An array of stable cross pyranose ring fragment ions representing sequential carbon loss as 30 u is observed. Consequently, the system provides an information-rich set of MSn spectra containing large amounts of structural information. Identification of D glucose (D-Glc), its two commonly found epimers (D-mannose and D-galactose), and the two major L-diastereomers of 6-dideoxymonosaccharides, L-fucose (L-Fuc) and L rhamnose (L-Rhm), are demonstrated. Selected pairs of disaccharides can be distinguished in terms of their anomeric linkage by reference to their MS3 spectra. PMID- 11114015 TI - Differentiating alpha- and beta-aspartic acids by electrospray ionization and low energy tandem mass spectrometry. AB - Spectra obtained by low-energy electrospray ionization tandem mass spectrometry (ESI-MS/MS) of 34 peptides containing aspartic acids at position n were studied and unambiguously differentiated. beta-Aspartic acid yields an internal rearrangement similar to that of the C-terminal rearrangements of protonated and cationized peptides. As a result of this rearrangement, two different ions containing the N- and the C-terminal ends of the original peptide are formed, namely, the bn-1 + H2O and y"l - n + 1 - 46 ions, respectively, where e is the number of amino acid residues in the peptide. The structure suggested for the y"l - n + 1 - 46 ion is identical to that proposed for the vn ions observed upon high energy collision-induced dissociation (CID) experiments. The intensity of these ions in the low-energy MS/MS spectra is greatly influenced by the presence and position of basic amino acids within the sequences. Peptides with a basic amino acid residue at position n - 1 with respect to the beta-aspartic acid yield very intense bn-1 + H2O ions, while the y"l - n + 1 - 46 ion was observed mostly in tryptic peptides. Comparison between the high- and low-energy MS/MS spectra of several isopeptides suggests that a metastable fragmentation process is the main contributor to this rearrangement, whereas for long peptides (40 AA) CID plays a more important role. We also found that alpha-aspartic acid containing peptides yield the normal immonium ion at 88 Da, while peptides containing beta-aspartic acid yield an ion at m/z 70, and a mechanism to explain this phenomenon is proposed. Derivatizing isopeptides to form quaternary amines, and performing MS/MS on the sodium adducts of isopeptides, both improve the relative intensity of the bn + 1 + H2O ions. Based on the above findings, it was possible to determine the isomerization sites of two aged recombinant growth proteins. PMID- 11114016 TI - On-line coupling of solid-phase extraction with mass spectrometry for the analysis of biological samples. II. Determination of clenbuterol in urine using multiple-stage mass spectrometry in an ion-trap mass spectrometer. AB - Solid-phase extraction (SPE) was coupled to ion-trap mass spectrometry to determine clenbuterol in urine. For SPE a cartridge exchanger was used and, after extraction, the eluate was directly introduced into the mass spectrometer. For two types of cartridges, i.e. C18 and polydivinylbenzene (PDVB), the total SPE procedure (including injection of 1 mL urine, washing, and desorption) has been optimised. The total analysis, including SPE, elution, and detection, took 8.5 min with PDVB cartridges, while an analysis time of 11.5 min was obtained with C18 cartridges. A considerable amount of matrix was present after extraction of urine over C18 cartridges, resulting in significant ion suppression. With PDVB cartridges, the matrix was less prominent, and less ion suppression was observed. For single MS, a detection limit (LOD) of about 25 ng/mL was found with PDVB cartridges. With C18 cartridges an LOD of only about 50 ng/mL could be obtained. Applying tandem mass spectrometry (MS/MS) did not lead to an improved LOD due to an interfering compound. However, a considerable improvement in the LOD was obtained with MS3. The selectivity and sensitivity were increased by the combination of efficient fragmentation of clenbuterol and reduction of the noise. Detection limits of 2 and 0.5 ng/mL were obtained with C18 and PDVB cartridges, respectively. The ion suppression was 4 to 45% (concentration range: 250 to 1.0 ng/mL) after extraction of urine using PDVB cartridges, and up to 70% ion suppression was observed using C18 cartridges. With MS4, no further improvement in selectivity and sensitivity was achieved, due to inefficient fragmentation of clenbuterol and no further reduction of noise. PMID- 11114017 TI - Evidence of nucleophilic addition to chemiluminescent N-sulfonylacridinium-9 carboxamides from electrospray ionization mass spectrometry. AB - Addition of nucleophiles to the chemiluminescent acridinium-9-carboxamide 1 resulted in the formation of the acridan adduct 7. The relative ratio of 1/7 present in solution could be determined by electrospray ionization mass spectrometry (ESI-MS) for a number of nucleophiles that are commonly employed during the use of 1 as a label in medical diagnostic assays. PMID- 11114018 TI - Analysis of some commercial polysorbate formulations using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - Analyses of polysorbate formulations (Tween 20, Tween 40, Tween 60, and Tween 80) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) reveal a complex mixture of oligomers that include polyethylene glycols, polyethylene glycol esters, isosorbide polyethoxylates, sorbitan polyethoxylates, polysorbate monoesters, polysorbate diesters, and sorbitol polyethoxylate esters. The MALDI-TOF mass spectra for these formulations show the presence of sodiated molecules in which the major signals are attributed to the presence of polyethylene glycols, isosorbide polyethoxylates, and sorbitan polyethoxylates. Additionally, the complexity of the spectra was correlated to the constituent fatty acid moieties in the polysorbate formulations. Thus Tween 20 showed the presence of polysorbate monolaurates, polysorbate monomyristates, and polysorbate monopalmitates. Tween 40 contained polysorbate mono- and dipalmitates. Tween 60 contained polysorbate monopalmitates and polysorbate monostearates. For the Tween 80, mass assignment for polysorbate monooleates and polysorbate dioleates was equivocal, because both of these oligomeric series have the same molecular weight as the sorbitan polyethoxylates, and thus the Tween 80 MALDI-TOF spectrum appeared to be the least complicated of the four commercial polysorbate formulations. PMID- 11114019 TI - Reaction monitoring of succinylation of collagen with matrix-assisted laser desorption/ionization mass spectrometry. AB - Succinylated collagen was synthesized by the reaction of collagen with succinic anhydride under basic conditions for one hour. Using the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) technique, the reaction products were directly identified without multi-step separation processes. MALDI-MS monitored the reaction more accurately than the conventional method of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). From the change in molecular mass during the reaction, it is observed that about 28 succinyl groups are attached to the collagen strand. PMID- 11114020 TI - Characterizing the transmission properties of an ion funnel. AB - The characteristic features of ion transmission properties in a radio-frequency (RF) electric-field driven ion funnel using the SIMION ion trajectory simulation package is presented. A user program applying the Douglas ion-neutral collisional drag coefficient model is incorporated to properly account for the ion focusing and transport effect of the background gas under the effect of the driving RF and a superimposed DC field. The simulated m/z transmission window compares favorably with the experimental results reported by Smith et al. RF amplitude and pressure dependence of experimentally observed m/z transmission windows are also examined, and an approximated effective potential model based on Gerlich's equation is proposed to interpret the low-m/z cutoff behavior. A modified ion funnel configuration is described. PMID- 11114021 TI - Ionisation and fragmentation of complex glycans with a quadrupole time-of-flight mass spectrometer fitted with a matrix-assisted laser desorption/ionisation ion source. AB - This paper reports the use of an experimental matrix-assisted laser desorption/ionisation (MALDI) ion source fitted to a quadrupole time-of-flight (Q Tof) mass spectrometer for the analysis of carbohydrates, particularly the N linked glycans from glycoproteins. Earlier work on the Q-Tof instrument, using electrospray ionisation, gave excellent MS/MS spectra, particularly from the [M + Na]+ ions, but suffered from the major disadvantages that the signal was often split between singly and multiply charged ions and that sensitivity fell dramatically as the molecular weight of the carbohydrate rose. The MALDI ion source did not suffer from these problems and the instrument produced excellent MS and MS/MS spectra from small amounts of complex, underivatised glycans as well as those derivatised at the reducing terminus. Positive ion MS spectra of sialylated glycans recorded on the new instrument were much less complex than those recorded with a conventional MALDI-TOF instrument because of the absence of ions resulting from metastable (post-source decay, (PSD)) fragmentations occurring in the flight tube. However, considerable fragmentation by loss of sialic acid still occurred. MS/MS spectra of the [M + Na]+ ions from all compounds were almost identical to those recorded earlier with the electrospray-Q Tof combination and far superior to MALDI-PSD spectra recorded with reflectron TOF instruments. Spectra are shown for neutral and sialylated N-linked glycans from chicken ovalbumin, riboflavin binding protein, alpha1-acid glycoprotein, bovine fetuin and ribonuclease B, both as free glycans and as those derivatised at their reducing termini. The technique was applied to the structural determination of N-linked glycans from human secretory IgA and Apo-B 100 from human low-density lipoprotein. PMID- 11114022 TI - Linked multiple-stage mass spectrometry experiments in an ion-trap using filtered noise fields techniques. PMID- 11114023 TI - Enhancing the intensities of lysine-terminated tryptic peptide ions in matrix assisted laser desorption/ionization mass spectrometry. AB - Tryptic digests of three proteins are reacted with O-methylisourea in order to convert lysine residues to homoarginines. The resulting homoarginine-terminated peptides exhibit more intense MALDI mass spectral peaks than their lysine terminated predecessors. This simple chemical reaction should therefore facilitate protein sequencing and mass mapping. PMID- 11114024 TI - Observation of Na(+)-bound oligomers of quercetin in the gas phase. AB - While developing a liquid chromatography/tandem mass spectrometry method for the analysis of the flavonoid quercitin, it was observed that quercetin (3,3',4',5,7 pentahydroxyflavone) exhibited clustering in both the positive and negative ion mode. Two series of positive ion clusters were observed; the first series corresponds to singly charged [2M + Na](+) at m/z 627.2 to [13M + Na](+) at m/z 3947.5, while the second series corresponds to doubly charged [7M + 2Na](2+) at m/z 1080.4 to [25M + 2Na](2+) at m/z 3798.5. In the negative ion mode, the behavior of quercetin parallels that of apigenin (4',5,7-trihydroxyflavone) in that [M + NO(3)](-), [2M + NO(3)](-), and [3M + NO(3)](-) were observed at m/z 364.1, 666.0, and 968.9, respectively; in addition, quercitin clusters with chloride ions ([2M + Cl](-) at m/z 638.9 and [3M + Cl](-) at m/z 940. 9) were observed. The results of tandem mass spectrometric examination of several cluster ions are reported. PMID- 11114025 TI - Identification of by-products from an orthogonal peptide ligation by oxime bonds using mass spectrometry and tandem mass spectrometry. AB - Synthetic proteins with unusual architecture are obtained through chemoselective ligation, a method based on the condensation of unprotected peptides under mild aqueous conditions. The last step of a new procedure leading to a tri-branched conjugate consists of the chemoselective ligation reaction between an (aminooxy)acetyl peptide and a peptide aldehyde resulting from a first ligation via an oxime bond. In order to optimize the reaction conditions, electrospray ionization mass spectrometry combined with liquid chromatography and tandem mass spectrometry has been used. In addition to the target tri-branched conjugate, two other conjugates were characterized allowing documentation of transoximation reactions in peptide chemistry. A fourth conjugate was identified as a side product appearing after the first ligation. Data obtained by low-energy collision induced dissociation led to a rapid and reliable identification of impurities observed in the (aminooxy)acetyl peptide despite a previous high performance liquid chromatography (HPLC) purification. This highlights the great reactivity of the aminooxy group towards carbonyl-containing compounds. PMID- 11114026 TI - Surface chemistry of RuO(2)/IrO(2)/TiO(2) mixed-oxide electrodes: secondary ion mass spectrometric study of the changes induced by electrochemical treatment. AB - The IrO(2)/RuO(2)/TiO(2) ternary system is well known for its electrocatalytic activity towards oxygen- and chlorine-evolution reactions. Electrochemical processing induces noticeable chemical and morphological modifications on these electrodes, depending on the noble metal oxide content. In this work, cathodic/anodic polarization and the oxygen-evolution reaction were studied in order to evaluate the electrocatalytic activity at various noble metal oxide percentages. The best performing electrode (30 mol% noble metal oxides) was analyzed before and after electrochemical tests by means of secondary ion mass spectrometry (SIMS) in order to determine the chemical composition modification which occurred on the surface and in deeper regions of the mixed-oxide film. PMID- 11114027 TI - Study of the structure of poly(methyl methacrylate) obtained in the presence of potassium hydride. AB - Electrospray ionization mass spectrometry (ESI-MS) in positive- and negative-ion mode and multi-step tandem mass spectrometry (ESI-MS ( n) ) were selected for the determination of the structure of poly(methyl methacrylate) obtained using potassium hydride as initiator. Macromolecules with methoxy and methyl starting groups, i. e. those situated at the beginning of the polymer backbone, were observed in this case. No other differences in the chain structure were found in the polymer. PMID- 11114028 TI - Effects of matrix-assisted laser desorption/ionization experimental conditions on quantitative compositional analysis of ethylene oxide/propylene oxide copolymers. AB - Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry has the potential to become a valuable tool for the compositional analysis of copolymers. For a copolymer composed of structurally very similar building blocks with minor chain length changes, one would expect the relative peak intensities observed in the MALDI mass spectra to reflect its composition, at least within a narrow mass range. However, we show that variations in experimental conditions in MALDI can have a significant effect on the mass spectral appearance of a copolymer. The effects of concentration, laser power, type of matrices and solvents on mass spectra of an ethylene oxide/propylene oxide copolymer are illustrated. These somewhat surprising results show that great care needs to be exercised when interpreting copolymer spectra for compositional analysis, even for copolymers with structurally similar monomers. This work also points out that further studies are needed to better understand and optimize spectral acquisition conditions for reliable copolymer compositional analysis by MALDI. PMID- 11114029 TI - Characterisation of variant forms of prophenin: mechanistic aspects of the fragmentation of proline-rich peptides. AB - Prophenin 1 (PF-1) is a 79-residue polypeptide originally isolated from porcine leukocytes. Its amino acid sequence has been determined by a combination of mass spectrometry and Edman degradation (Harwig SSL. et al. FEBS Lett. 1995; 362: 65). Prophenin (PF) and variants thereof are also found in organic extracts of porcine pulmonary tissue (Wang Y. et al. FEBS Lett. 1999; 460: 257). In the present study we have characterised the variant forms of PF found in these extracts using nano electrospray (nano-ES) high resolution and tandem mass spectrometry. The major forms of PF found in these extracts by nano-ES mass spectrometry are the 80 residue polypeptides prophenin-2-Pyr (PF-2-Pyr) and prophenin-2-Gln (PF-2-Gln). Prophenin-2-Pyr is refractory to Edman degradation due to the presence of an N terminal pyroglutamic residue. In PF-2-Gln the N-terminal residue is glutamine and the C-terminus is amidated. In porcine pulmonary extracts PF-1 is present to only a minor extent. Other shorter polypeptides are also found in these extracts including 18- and 17-residue C-terminal fragments of PF. The primary structure of PF is highly unusual in that it shows four almost perfect decamer repeats of FPPPN(V/F)PGPR and, out of the 79/80 residues, 42 are proline and 14 are phenylalanine. Tryptic digestion of PF gives peptides containing the decamer repeat and collision-induced dissociation of these peptides provides an insight into the fragmentation mechanisms of proline-rich peptides. Facile cleavage within the Pro-Pro-Pro sequence of these peptides suggests the involvement of a cyclic peptide in the fragmentation mechanism. Fragmentation mechanisms that account for the formation of fragment ions at other cleavage sites are also discussed. PMID- 11114030 TI - Applications of mass spectrometry in cultural heritage: identification of ligands employed for mordant gilding. AB - Three different samples of gilding from wall paintings, from the Palazzo della Ragione in Padua, have been investigated by GC/MS analysis of their basic hydrolysis products obtained using trimethyl(alpha,alpha,alpha-trifluoro-m tolyl)ammonium hydroxide (TMTFTH). The method employed allowed the identification of the mordant used for the paints, and consequently to distinguish Menabuoi's original painting from the others produced by extensive restorations. This was achieved by characterizing, among the hydrolysis products, molecular species characteristic of natural resins, siccative oils and waxes. PMID- 11114031 TI - Measurement of oilfield chemical residues in produced water discharges and marine sediments. AB - During oil production, significant quantities of water are produced with the crude oil which, following treatment on the platform, are discharged to the marine environment. This produced water contains residues of oilfield chemicals added by the platform operators to the topside processing equipment to aid oil water separation and mitigate operational problems. The levels of oilfield chemicals entering the marine environment via this route were investigated using electrospray ionisation tandem mass spectrometry (ESI-MS/MS) and wet chemical analysis techniques. The generic nature of different chemical types was shown by ESI-MS/MS. Studies of the partitioning behaviour of corrosion inhibitors and demulsifiers between the oil and water phases of the produced fluids suggested corrosion inhibitors partitioned primarily into the aqueous phase and demulsifiers into the oil phase. This was reflected in levels observed in produced water although, in the case of a corrosion inhibitor, lower than expected concentrations were measured. Scale inhibitors were discharged with the produced water at their dosing concentrations. Marine sediments in the proximity of two North Sea oil platforms contained low levels of benzalkonium quaternary ammonium salts (0.74-10.84 ng/g), typical corrosion inhibitor chemicals. PMID- 11114032 TI - On-probe sample pretreatment for the detection of proteins above 15 KDa from whole cell bacteria by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - A rapid methodology is described for the enhancement of the signal-to-base-line (S/B) ratio of high molecular weight protein signals from whole cell bacteria analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). The procedure involves depositing growing bacteria colonies from culture dishes directly onto the MALDI probe followed by treatment of the sample spot with a 2 microL aliquot of 40% ethanol prior to the addition of a ferulic acid matrix solution (12.5 mg dissolved in 17% formic acid/33% acetonitrile/50% H(2)O). Protein signals of more than 20 kDa were routinely produced from both Gram positive and Gram negative bacteria prepared in this manner. Moreover, a substantial number of intense protein signals were also produced in the more 'conventional' fingerprint region extending from 4 to 20 kDa. This approach is rapid, easy to implement into existing methodologies, and does not require any special hardware. PMID- 11114033 TI - Identification of triacylglycerols containing two short-chain fatty acids at sn-2 and sn-3 positions from bovine udder by fast atom bombardment tandem mass spectrometry. AB - Several triacylglycerol (TAG) molecular species, that contain two short-chain fatty acids (C4 to C8) at the sn-2 and sn-3 positions of the glycerol backbone, were isolated from bovine udder by using solvent extraction and silica gel column chromatography. Their structures were identified by fast atom bombardment (FAB) tandem mass spectrometry (MS/MS), based on the information obtained from collision-induced dissociation (CID) spectra of sodium-adducted molecules ([M + Na](+)) of model TAG compounds which had been synthesized from glycerol and appropriate fatty acids. For each species, the relative positions of the three fatty acids on the glycerol backbone, as well as fatty acid composition and double-bond position in the fatty acyl group, were determined. A majority of sodium-adducted molecules observed in the FAB mass spectrum were mixtures of at least two components that have different fatty acid composition but the same molecular mass. In addition, all the components present in mixtures of all the species contain a long-chain fatty acid (C12 to C18) at the sn-1 position, a short-chain fatty acid (C4 to C8) at the sn-2 position, and a butyric acid uniquely at the sn-3 position. PMID- 11114035 TI - Electron capture dissociation of b (2+) peptide fragments reveals the presence of the acylium ion structure. AB - Electron capture dissociation (ECD) of peptides and their fragments has now been extended to b ( n) ( 2+) ions, where it also produced far more structural information than collisional activation. Interestingly, b ( n) ( 2+) ions revealed abundant loss of CO from radical monocations and the presence of c ((n - 1)) ( +.) fragments. The CO loss from peptide radical cations is unusual and was attributed to neutralization of the -C identical with O(+) group in the acylium ion structure, supported by the observation of c ( (n - 1)) ( +.) ions that can only be formed from an open-chain ion. These characteristic features were most prominent for b ( 12)( 2+) ions of renin substrate and least prominent for b ( n) ( 2+) ions of substance P (n = 9,10). Totally, out of seven b ( n) ( 2+) ions studied, CO loss above 3% level was present in all spectra as well as c ( (n - 1))( +.) fragments of three species, suggesting that the acylium ion structure plays a significant role for at least some b ( 2+) ions. This is an unexpected result in view of the literature data for small, singly charged b ions, for which the protonated oxazolone structure is favoured in ab initio calculations. Apparently, more studies are required before extrapolating the small molecule results to large species. The CO loss in ECD can be used for distinguishing between b and y ions in the MS/MS spectrum of larger molecules. PMID- 11114034 TI - A rapid and simple method for quantitation of urinary hydroxylysyl glycosides, indicators of collagen turnover, using liquid chromatography/tandem mass spectrometry. AB - Some glycosides of hydroxylysine, viz., alpha-1, 2-glucosylgalactosyl-O hydroxylysine and beta-1-galactosyl-O-hydroxylysine, appear to be good indicators of collagen turnover. A simple liquid chromatography/tandem mass spectrometry (LC/MS/MS) method is proposed for measuring these analytes in urine, with no sample preparation except for a dilution step. Quantitation is performed using external calibration with no internal standard. A preliminary survey indicates good intra- and inter-day reproducibility (better than 5 and 8%, respectively). With the present method, the estimated limits of detection (S/N > 3) in urine are 0.8 and 0.5 microM/L for beta-1-galactosyl-O-hydroxylysine and alpha-1,2 glucosylgalactosyl-O-hydroxylysine, respectively. The method is proposed as a robust tool for a large-scale research investigation on collagen turnover. PMID- 11114037 TI - A study of the electrospray ionisation and ion trap fragmentation of hemiterpenoid and dimeric coumarin derivatives. AB - The electrospray ionisation (ESI) of selected hemiterpenoid and dimeric coumarin derivatives and their subsequent fragmentation using an ion trap mass spectrometer are reported and discussed. Sequential product ion fragmentation experiments (MS(n)) were performed in order to elucidate the degradation pathways for these compounds. The results illustrate that the observed characteristic fragmentation patterns are of considerable utility in the application of ESI mass spectrometry to the characterisation of this class of compounds. PMID- 11114036 TI - Molecular characterization of a highly heterogeneous mixture of glucosylceramides from a deep-water Mediterranean scleractinian coral Dendrophyllia cornigera. AB - We report here on the structural characterization of a highly heterogeneous mixture of glucosylceramides (GlcCers) isolated from a deep-water Mediterranian dendrophylliid coral, Dendrophyllia cornigera. The neutral glycosphingolipid (GSL) components of the coral were separated into three HPLC fractions which were structurally characterized by nuclear magnetic resonance (NMR) and mass spectrometry (MS). NMR analysis revealed a beta-glucosylpyranose, a methyl branched conjugated sphingadienine and alpha-hydroxy fatty acid moieties characteristic for the species. Molecular mass distributions of the HPLC fractions were monitored using single-stage MS. At least 17 different GlcCer constituents with variable long-chain base and fatty acid residues were observed based on the molecular ion peaks in the liquid secondary ion (LSI) survey spectra. Structures of the individual components were revealed by product ion spectra of the alkali-cationized molecules ([M + Cat](+)), which resulted in two characteristic fragment ions, F(F) and F(S). Tandem MS of the same fragment ions formed in the ion source showed that F(F) carries the hydoxy fatty acid, while F(S) carries the long-chain sphingoid base, thus providing complementary structural information for the characterization of ceramide composition. Based on the tandem mass spectra of the molecular ions [M + Na](+), 26 different GlcCers of the coral were identified. The ceramide moiety showed heterogeneity in both the sphingoid portion (d18:2, d19:2, d20:2 and d20:3) and the alpha-hydroxy fatty acid chain (h19-h24, either saturated or unsaturated), forming an extremely heterogeneous mixture. The method is generally applicable to the characterization of structurally heterogeneous GlcCer mixtures. PMID- 11114038 TI - Gas chromatography/ion trap tandem mass spectrometry for the analysis of halobenzenes in soils by solid-phase microextraction. AB - Headspace solid-phase microextraction combined with gas chromatography/ion trap tandem mass spectrometry (HS-SPME/GC/ITMS/MS) was used for the analysis of 12 halobenzenes from soil samples. For MS/MS optimisation, the experiments were performed by precursor ion selection and software controlled operations. Collision-induced dissociation (CID) can be achieved by two different approaches, resonant and non-resonant excitation modes. Different results were obtained using the two approaches, and the resonant excitation mode was chosen as the best for all halobenzenes. Parameters such as the CID excitation amplitude, excitation RF storage level and CID bandwidth frequency were optimised to maximise the formation of halobenzene product ions. A 100-microm polydimethylsiloxane fibre was used for the isolation and preconcentration of the analytes. The HS SPME/GC/ITMS/MS method was applied to the analysis of halobenzenes in an agricultural soil sample. The halobenzenes were quantified by standard addition, which led to good reproducibility (RSD between 4.7 and 9.2%) and detection limits in the low pg/g range. The method was validated by comparing the results with those obtained in a European inter-laboratory exercise. PMID- 11114039 TI - High-throughput approaches to the quantitative analysis of ketoconazole, a potent inhibitor of cytochrome P450 3A4, in human plasma. AB - Ketoconazole, an imidazole-piperazine compound, is an orally active antimycotic agent. In addition, ketoconazole is a specific inhibitor of cytochrome P450 3A4. As about 60% of oxidized drugs are biotransformed by this isoform, the potential effect of a concomitant administration of ketoconazole on drug disposition may be of interest during drug development. The present paper describes three different approaches (methods A, B, and C) to attain high-throughput sample preparation and analysis in the quantification of ketoconazole in human plasma. Method A consisted of acetonitrile precipitation in a 96-well plate, transfer of the supernatant via a Tomtec Quadra 96 Model 320, and subsequent injection onto a 50 x 4.6 mm (i.d.) Develosil Combi-RP-5 column (packed with C30 bonded silica particles). Method B consisted of an identical sample preparation to method A with the exception that a Michrom Magic Bullet(trade mark) column, 2.0 --> 0.50 mm (i.d., tapered bore) x25 mm length, was used. Lastly, in method C, a turbulent flow chromatography (TurboFlow LC/APCI-MS/MS) module was used for the direct analysis of ketoconazole in human plasma. A Sciex API 3000 was used in methods A and B, while a Micromass Quattro LC was employed in method C. Based on the values obtained for the calibrator (standard) and quality control samples, all three protocols yielded satisfactory accuracy, precision, and reduced manual sample preparation time. PMID- 11114040 TI - A rapid screening assay for measuring urinary androsterone and etiocholanolone delta(13)C ( per thousand) values by gas chromatography/combustion/isotope ratio mass spectrometry. AB - A gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) method is described and validated for measurement of delta(13)C values of the acetate derivatives of urinary etiocholanolone and androsterone. The analysis was performed with only 2 mL of urine. The sample preparation consisted of deconjugation with beta-glucuronidase, solid phase extraction, and derivatization with acetic anhydride and pyridine. The within-assay precision of two quality control (QC) urine samples ranged from 0.5 to 2.1 CV%. The between-assay precision in the same QC urines ranged from 1.7 to 3.4 CV%. Administration of testosterone enanthate to a subject resulted in a 6 per thousand decrease in delta(13)C values from -25 per thousand (baseline) to -31 per thousand. Two weeks after testosterone administration was discontinued, the delta(13)C values remained abnormally low while the urine testosterone/epitestosterone (T/E) ratio returned to less than 6. This relatively simple method is useful for rapidly screening a large number of urine samples, including those with T/E <6. PMID- 11114041 TI - Sensitivity enhancement for nitrophenols using cationic surfactant-modified activated carbon for solid-phase extraction surface-assisted laser desorption/ionization mass spectrometry. AB - Previous work has demonstrated that a combination of solid-phase extraction with surface-assisted laser desorption/ionization (SPE-SALDI) mass spectrometry can be applied to the determination of trace nitrophenols in water. An improved method to lower the detection limit of this hyphenated technique is described in this present study. Activated carbon powder is used as both the SPE adsorbent and the SALDI solid in the analysis by SPE-SALDI. The surface of the activated carbon is modified by passing an aqueous solution of a cationic surfactant through the SPE cartridge. The results demonstrate that the sensitivity for nitrophenols in the analysis by SPE-SALDI can be improved by using cationic surfactants to modify the surface of the activated carbon. The detection limit for nitrophenols is about 25 ppt based on a signal-to-noise ratio of 3 by sampling from 100 mL of solution. PMID- 11114042 TI - New results on photoion pair formation from application of the velocity imaging photoionisation coincidence technique. AB - The velocity imaging photoionisation coincidence (VIPCO) technique is shown to be a powerful tool for studies of ion pair formation. Sequential mechanisms are demonstrated for some three-body ion pair formation reactions. Observation of a new type of reaction producing one negatively and two positively charged ions plus an electron is reported. PMID- 11114043 TI - Characterization of methylation site of monomethylflavan-3-ols by liquid chromatography/electrospray ionization tandem mass spectrometry. AB - Liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI MS/MS) was used for the structural characterization and differentiation of four isomeric O-monomethylated catechins (on phenolic positions) by the analysis of the fragmentation behaviour of catechin. The catechin fragmentation routes were rationalized and it is shown that several diagnostic ions such as (1,3)A(+), (1,2)B(+), and (1,4)B(+) allow the unambiguous identification of the methylated ring. The precise position of the methyl group on each ring is determined by the difference in the relative intensities of the diagnostic ions. Isomeric O methylepicatechins were also differentiated using this methodology. PMID- 11114044 TI - Analysis of process impurities in the basic drug SB-253149 using capillary electrophoresis and on-line mass spectrometric detection. AB - Capillary electrophoresis with on-line electrospray ionisation mass spectrometry (CE/ESI-MS) has been used to identify process impurities in a batch of the anti atherosclerotic drug, SB-253149. The impurities were separated from the main drug compound by capillary electrophoresis (CE) using an ammonium formate buffer at low pH in an untreated fused silica capillary. The CE method was initially developed using UV as the detection mode and then later structural elucidation work was achieved using an ion trap mass spectrometer. To maintain peak resolution and peak shape when the CE system was coupled to the mass spectrometer, a modified capillary cassette linked to a coaxial sheath flow electrospray ionisation (ESI) interface was used. By performing MS/MS experiments in conjunction with chemical knowledge of the reactivities of SB-253149, it was possible to propose molecular structures for impurities detected in the batch of SB-253149. The results from this study revealed that most of the process impurities in SB-253149 were dimeric derivatives of the parent molecule as well as trace levels of the starting material. This type of information was vital in process control and optimisation for the synthetic route for this drug. PMID- 11114045 TI - Direct peptide mapping of real samples containing sickle-cell and fetal hemoglobin by electrospray mass spectrometry. PMID- 11114046 TI - Determination of eight sulfonamides in bovine kidney by liquid chromatography/tandem mass spectrometry with on-line extraction and sample clean up. AB - A sensitive, high performance liquid chromatography/tandem mass spectrometric (i.e. mass spectrometry/mass spectrometry; LC/MS/MS) method with on-line extraction and sample clean-up for the screening and confirmation of residues of sulfonamides in kidney is described. The sulfonamides are extracted from homogenized kidney with methanol. After centrifugation of the extract, an aliquot of the extract is directly injected on the LC/MS/MS system with further extraction and clean-up of the sample on-line. Detection of the analytes was achieved by positive electrospray ionization (ESI) followed by multiple reaction monitoring. For each sulfonamide the collisional decomposition of the protonated molecule to a common, abundant fragment ion was monitored. The method has been validated for sulfadimethoxine, sulfaquinoxaline, sulfamethazine, sulfamerazine, sulfathiazole, sulfamethoxazole, sulfadiazine and sulfapyridine. Calibration curves resulting from spiked blank kidney samples at the 10-200 microg/kg level showed good linear correlation. At the level of 50, 100 and 200 microg/kg both within- and between-day precision, as measured by relative standard deviation (RSD), were less than 16%. The limits of detection (LODs) ranged from 5 to 13.5 microg/kg. The recoveries ranged from 78 to 82%. The procedure provides a rapid, reliable and sensitive method for the determination of residues of sulfonamides in bovine kidney. The advantage of this method over existing methods is its decreased sample preparation and analysis time, which makes the method more suitable for routine analysis. PMID- 11114047 TI - Electron impact mass spectral fragmentation of 3a,5-disubstituted 1, 3-diphenyl 3a,4,5,6-tetra-hydro-3H-1,2,4-triazolo[4,3-a][1, 5]benzo-diazepines. AB - The mass spectrometric behaviour of six 3a,5-disubstituted 1, 3-diphenyl-3a,4,5,6 tetrahydro-3H-1,2,4-triazolo[4,3-a][1, 5]benzodiazepines has been studied with the aid of mass-analyzed ion kinetic energy spectrometry and accurate mass measurements under electron impact ionization. All compounds show a tendency to eliminate (substituted) styrene molecules, aryl radicals, arylmethyl radicals or phenylnitrene (PhN:). All of the resulting fragment ions, except [M - PhN:](+.), could further undergo a reverse [2 + 3] cycloaddition. The [M - PhN:](+.) ions could further lose styrene derivatives and undergo a ring enlargement rearrangement. The molecular ions also show a tendency to eliminate a phenyl radical, and the [M - Ph](+) ions could eliminate styrene derivatives. The [M - R(1)CH = CH(2)](+.) ions could further lose NH(2) to yield stable tetracyclic 1,3 diphenyl-1,2,4-triazolo[4,3-d]phenanthridine ions, which could further lose benzonitrile, or undergo a reverse [2 + 3] cycloaddition. The molecular ions could also undergo a reverse [2 + 3] cycloaddition to produce N phenylbenzonitrile imine ions and 2, 4-disubstituted 2,3-dihydro-1H-1,5 benzodiazepine ions, whose further fragmentations were also investigated. PMID- 11114048 TI - Detection of exogenous intake of natural corticosteroids by gas chromatography/combustion/isotope ratio mass spectrometry: application to misuse in sport. AB - A detailed procedure for the analysis of exogenous hydrocortisone and cortisone in urine by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) is proposed. As urinary levels of hydrocortisone are rather low for GC/C/IRMS analysis, the focus is on the main corticosteroid metabolites, tetrahydrocortisone (THE) and tetrahydrocortisol (THF). Following different solid phase extraction purifications, THE and THF are oxidized to 5beta androstanetrione before analysis by GC/C/IRMS. Significant differences in delta(13)C per thousand values of synthetic natural corticosteroids and endogenous human corticosteroids have been observed. Therefore, a positive criterion, to detect exogenous administration of synthetic corticosteroids in anti-doping control, is proposed. PMID- 11114049 TI - Derivatization procedures to facilitate de novo sequencing of lysine-terminated tryptic peptides using postsource decay matrix-assisted laser desorption/ionization mass spectrometry. AB - Guanidination of the epsilon-amino group of lysine-terminated tryptic peptides can be accomplished selectively in one step with O-methylisourea hydrogen sulfate. This reaction converts lysine residues into more basic homoarginine residues. It also protects the epsilon-amino groups against unwanted reaction with sulfonation reagents, which can then be used to selectively modify the N termini of tryptic peptides. The combined reactions convert lysine-terminated tryptic peptides into modified peptides that are suitable for de novo sequencing by postsource decay matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The guanidination reaction is very pH dependent. Product yields and reaction kinetics were studied in aqueous solution using either NaOH or diisopropylethylamine as the base. Methods are reported for derivatizing and sequencing lysine-terminated tryptic peptides at low pmole levels. The postsource decay (PSD) MALDI tandem mass spectra of a model peptide (VGGYGYGAK), the homoarginine analog and the sulfonated homoarginine analog are compared. These spectra show the influence that each chemical modification has on the peptide fragmentation pattern. Finally, we demonstrate that definitive protein identifications can be achieved by PSD MALDI sequencing of derivatized peptides obtained from solution digests of model proteins and from in-gel digests of 2D gel separated proteins. PMID- 11114050 TI - Electron impact fragmentation mechanisms of some cyclic esters with helical structures AB - The electron impact mass spectra of several cyclic esters with helical structures have been studied. Their fragmentation pathways were proposed and confirmed by mass-analyzed ion kinetic energy (MIKE) and high-resolution data. In general, the dominant fragmentation pathways in the spectra of these compounds originate from a alpha-cleavage with loss of a hydrogen or methyl group. The difference between hydrogen and methyl group loss greatly affects the subsequent fragmentations. Although, due to their helicity, these cyclic esters are optically active no stereo-related fragmentation pathway was observed. Copyright 2000 John Wiley & Sons, Ltd. PMID- 11114051 TI - Identification of an N-(hydroxysulfonyl)oxy metabolite using in vitro microorganism screening, high-resolution and tandem electrospray ionization mass spectrometry. AB - Preliminary metabolic profiling of a drug under pre-clinical development revealed the presence of a minor unknown metabolite with a positive ion electrospray ionization (ESI) mass spectrum identical to that of the unchanged compound. Since the low concentration of the compound did not allow any additional experiments, preparative bioconversion using fungi was used to obtain a substantial amount of the molecule. Negative ion ESI-MS and tandem mass spectrometry (MS/MS) in combination with accurate mass measurements obtained on a quadrupole/time-of flight instrument (Q-TOF) led to the positive identification of a hydroxylamide sulfoconjugated metabolite. PMID- 11114052 TI - Mass spectrometric studies of the pterocarpan skeleton. AB - The mass spectrometric characteristics of the pterocarpan skeleton have been studied with the aid of metastable decomposition (CAD-MIKES), high resolution mass measurements and specific deuterium labeling. New structural rotations have been postulated for the [M - H](+) ions which can explain the fragmentation routes for the pterocarpan skeleton. PMID- 11114053 TI - Electron impact mass spectral studies of 2a,4-disubstituted 2-phthalimido 2,2a,3,4-tetrahydro-1H-azeto[2,1-d][1, 5]benzothiazepin-1-ones. AB - The mass spectrometric behaviour of seven 2a,4-disubstituted 2-phthalimido 2,2a,3,4-tetrahydro-1H-azeto[2,1-d][1, 5]benzothiazepin-1-ones has been studied with the aid of mass-analyzed ion kinetic energy spectrometry and accurate mass measurements under electron impact ionization. All compounds show a tendency to eliminate a CO molecule, a phthalimido (PhthN) radical, or a phthalimide (PhthNH) molecule. All of the resulting fragment ions could further lose a propene or (substituted) styrene molecule. The molecular ions could also undergo a reverse [2 + 2] cycloaddition reaction. PMID- 11114054 TI - Directly coupled liquid chromatography with inductively coupled plasma mass spectrometry and orthogonal acceleration time-of-flight mass spectrometry for the identification of drug metabolites in urine: application to diclofenac using chlorine and sulfur detection. AB - We report the application of high-performance liquid chromatography (HPLC) linked to inductively coupled plasma mass spectrometry (ICPMS) and orthogonal acceleration time-of-flight mass spectrometry (oa-TOFMS) for the identification of phase I and II urinary metabolites of diclofenac. The metabolites were separated by reversed-phase HPLC monitored with a UV diode array detector (UV DAD) after which 90% of the eluent was directed to an ICPMS source, with the remainder going to an oa-TOF mass spectrometer. Compounds containing (35)Cl, (37)Cl and (32)S were detected specifically using ICPMS and identified by oa TOFMS. The metabolites detected and identified in this way included glucuronic acid and sulfate conjugates, mono- and dihydroxylated and free diclofenac. In addition a previously unreported in vivo metabolite, an N-acetylcysteinyl conjugate of diclofenac, was also characterised. This is the first application of the combination of HPLC/UV-DAD/ICPMS/oa-TOFMS for the investigation of the metabolic fate of chlorinated xenobiotics by direct biofluid analysis. PMID- 11114055 TI - Electrospray ionization tandem mass spectrometric studies of competitive pyridine loss from platinum(II) ethylenediamine complexes by the kinetic method. AB - The competition between pyridine ligand loss in square planar Pt(II) complexes has been examined using the doubly and singly charged ions of complexes consisting of platinum(ethylenediamine) coordinated to two different substituted pyridines. Collision induced dissociation (CID) of [Pt(en)Py(1)Py(2)](2+) (where Py(1) = one of ten different substituted pyridines and Py(2) = pyridine) results in loss of the protonated pyridines to yield the singly charged platinum ions [Pt(en)Py(1)-H](+) and [Pt(en)Py(2)-H](+). In contrast, fragmentation of [Pt(en)Py(1)Py(2)-H](+) results in neutral pyridine loss to yield the ions [Pt(en)Py(1)-H](+) and [Pt(en)Py(2)-H](+). In the latter case, the correlation between relative losses of each pyridine compared to their gas-phase proton affinities is poor. A novel chloride ion abstraction reaction occurs for the fragmentation of [Pt(en)Py(1)Py(2)](2+) when Py(1) = o-C(5)H(4)CIN and Py(2) = C(5)H(5)N, to yield the [Pt(en)(Cl)Py(2)](+) and [o-C(5)H(4)N](+) pair of ions. In order to model this process the competition between nitrogen and chlorine binding in [Pt(NH(3))(3)(o-NC(5)H(4)Cl)](2+) has been examined using density functional theory (DFT) calculations at the B3LYP/LANL2DZ level of theory. Both adducts are minima with the N adduct being more stable than the Cl adduct by 22.7 kcal mol(-1). Furthermore, the Cl adduct exhibits a significant stretching of the C-Cl bond (to 1.935 A), consistent with the observed chloride ion abstraction reaction, which is endothermic by 9.0 kcal mol(-1) (relative to the N adduct). PMID- 11114056 TI - Characterization of Aspergillus spores by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - The intact fungal spores of several strains of four Aspergillus species, Aspergillus flavus, A. oryzae, A. parasiticus, and A. sojae, were directly analyzed by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry. Very simple MALDI mass spectra are obtained by directly mixing spores with a matrix such as alpha-cyano-4-hydroxycinnamic acid or sinapinic acid. The mass spectra are obtained from the ablation of cell walls of spores owing to the acidity of the matrix solution. The MALDI results show that aflatoxigenic strains and non-aflatoxigenic strains have different mass peak profiles. Furthermore, the MALDI results of non-aflatoxigenic A. flavus and A. parasiticus spores resemble those of the closely related A. oryzae and A. sojae spores, respectively. PMID- 11114057 TI - Determination of gamma-hydroxybutyric acid (GHB) in plasma and urine by headspace solid-phase microextraction and gas chromatography/positive ion chemical ionization mass spectrometry. AB - A new method for the qualitative and quantitative analysis of gamma hydroxybutyric acid (GHB) in plasma and urine samples is described. It involves the conversion of GHB to gamma-butyrolactone (GBL), its subsequent headspace solid-phase microextraction (SPME), and detection by gas chromatography/positive ion chemical ionization mass spectrometry (GC/PICI-MS), using D(6)-GBL as internal standard. The assay is linear over a plasma GHB range of 1-100 microg/mL (n = 5, r = 0.999) and a urine GHB range of 5-150 microg/mL (n = 5, r = 0. 998). Relative intra- and inter-assay standard deviations, determined for plasma and urine samples at 5 and 50 microg/mL, are all below 5%. The method is simple, specific and reasonably fast. It may be applied for clinical and forensic toxicology as well as for purposes of therapeutic drug monitoring. PMID- 11114058 TI - Miniature time-of-flight mass spectrometer using a flexible circuitboard reflector AB - An innovative design for a miniature time-of-flight mass spectrometer has been developed employing several newly designed components. These include: (1) a gridless, focusing ion source allowing for the use of very high extraction energies in a maintenance-free design, (2) a new method of construction for an ion reflector using rolled flexible circuitboard material, and (3) an improved center-hole microchannel plate detector assembly that significantly reduces the noise (or 'ringing') inherent in the coaxial design. A prototype time-of-flight instrument was constructed and used to evaluate the performance of these components. Compared to previous designs, results indicate that background noise for data acquired on this instrument is substantially reduced, resolution is improved, and the potential for mass producing this instrument in an inexpensive and rugged package for field-portable and remote installations is significantly enhanced. Copyright 2000 John Wiley & Sons, Ltd. PMID- 11114059 TI - Exploratory pharmacokinetics and brain distribution study of a neuropeptide FF antagonist by liquid chromatography/atmospheric pressure ionization tandem mass spectrometry. AB - Dansyl-Pro-Gln-Arg-NH(2), an N-terminally modified tripeptide amide and a putative neuropeptide FF antagonist, was amenable to both positive-ion ESI and APCI. The protonated molecule yielded several fragment ions upon collision induced dissociation in a quadrupole ion trap instrument for the development of LC/MS/MS assay methods. ESI clearly outperformed APCI in limits of detection, and was the method of choice for coupling with narrow-bore reversed-phase liquid chromatography to assess the pharmacokinetic profile and brain concentration of the neuropeptide FF antagonist in experimental animals. While plasma could be analyzed after rapid sample preparation, brain tissue required cleanup (solid phase extraction) and preconcentration before injection, and the assay was prone to matrix interference. This study indicated a rapid disappearance of dansyl-Pro Gln-Arg-NH(2) from the plasma and the brain, and modest CNS bioavailability after intravenous administration to rats. PMID- 11114060 TI - Quantitation by (1)H-NMR of dolichol, cholesterol and choline-containing lipids in extracts of normal and phathological thyroid tissue. AB - Proton magnetic resonance spectroscopy at 1.9 T was used to quantify dolichols, cholesterols, choline-containing phospholipids and double bonds in unsaturated acyl chains in lipid extracts of four types of thyroid tissue [normal (n = 27), papillary cancer (n = 15), adenoma (n = 13) and Basedow disease (n = 6)]. In normal thyroid the mean concentrations of dolichol, cholesterol and phospholipids were 1.2, 3.6 and 2.1 micromol/g wet weight, respectively. The concentrations of these lipids exhibited positive mutual correlations and positive correlations with patient age. The increase in dolichol in elderly human thyroid may be due to the accumulation of lysosomes and may help to compensate for the decrease in the activity of lysosomal enzymes and in thyroid hormone production and release. Dolichol concentrations were significantly lower in papillary cancer (0.4 micromol/g) and Basedow disease (0.3 micromol/g) compared to normal thyroid (p < 0.01 and p < 0.05, respectively), while cholesterol was enhanced only in cancer tissue (10.7 micromol/g). Benign adenoma exhibited normal levels of both dolichol and cholesterol. These results suggest that the synthesis and accumulation of isoprenoids are normal in adenoma but not in cancer. PMID- 11114061 TI - Regional cerebral blood volume response to hypocapnia using susceptibility contrast MRI. AB - We used steady-state susceptibility contrast MRI to evaluate the regional cerebral blood volume (rCBV) response to hypocapnia in anesthetised rats. The rCBV was determined in the dorsoparietal neocortex, the corpus striatum, the cerebellum, as well as blood volume in extracerebral tissue (group 1). In addition, we used laser-Doppler flow (LDF) measurements in the left dorsoparietal neocortex (group 2), to correlate changes in CBV and in cerebral blood flow. Baseline values, expressed as a percentage of blood volume in each voxel, were higher in the brain regions than in extracerebral tissue. Hypocapnia (P(a)CO(2) approximately 25 mmHg) resulted in a significant decrease in CBV in the cerebellum (-17 +/- 9%), in the corpus striatum (-15 +/- 6%) and in the neocortex (-12 +/- 7%), compared to the normocapnic CBV values (group 1). These changes were in good agreement with the values obtained using alternative techniques. No significant changes in blood volume were found in extracerebral tissue. The CBV changes were reversed during the recovery period. In the left dorsoparietal neocortex, the reduction in LDF (group 2) induced by hypocapnia (-21 +/- 8%) was in accordance with the values predicted by the Poiseuille's law. We conclude that rCBV changes during CO(2) manipulation can be accurately measured by susceptibility contrast MRI. Abbreviations used: ANOVA analysis of variance CBF cerebral blood flow CBV cerebral blood volume CPMG Carr-Purcell-Meiboom-Gill FiO(2) fractional inspired oxygen ICP intracranial pressure LDF laser-Doppler flow MABP mean arterial blood pressure MRI magnetic resonance imaging MTT mean transit time PaCO(2) arterial partial pressure of carbon dioxide PaO(2) arterial partial pressure of oxygen PET positron emission tomography rCBV regional cerebral blood volume SPECT single-photon emission computed tomography PMID- 11114062 TI - Magnetic susceptibility shift selected imaging (MESSI) and localized (1)H(2)O spectroscopy in living plant tissues. AB - Maize root segments permeated with aqueous solutions of the paramagnetic agents GdDTPA(2-) or DyDTPA-BMA display two well-resolved NMR peaks corresponding to the signals from intracellular and extracellular (1)H(2)O, which arise from well understood bulk magnetic susceptibility effects. This allows each component to be studied separately. Images obtained at each frequency with MESSI editing, and single- and multiple-voxel ('spectroscopic imaging') localized spectra, clearly indicate that the agents permeate into the interstitial spaces, and into the longitudinal (xylem/phloem) channels in the stele (core) of the root, confirming earlier assessments. We believe these are the first images of a multicellular tissue acquired in vivo exclusively from the intracellular water proton resonance. This method can be further exploited to study water transport in similar systems. PMID- 11114063 TI - A precise and user-independent quantification technique for regional comparison of single volume proton MR spectroscopy of the human brain. AB - The aim of this work was to study and correct the influence of varying coil load and local B(1) field in single volume MR spectroscopy. A simple, precise, and user-independent way to adjust the transmitter gain has been developed and validated. It is based on a fit of the localized signal to flip angle variation around 90 degrees. This method proved to be robust against B(1) gradients and suitable for in vivo applications. Local B(1) correction was combined with an external reference and decomposition of the volume into CSF and tissue to obtain a comprehensive absolute quantification of tissue water content and metabolite concentrations in human brain. STEAM localized spectra of parietal and insular gray matter and subparietal white matter (n = 11, TE = 30 ms) were analyzed using a linear combination of model spectra (LCModel). Coefficients of variation (CV) between 1.5% and 4% were obtained for the tissue water content (1-2% in a single subject). The CVs of major metabolite concentrations (4-21%) were dominated by the errors of the spectral analysis. The largest B(1) variation in the in vivo experiments (range 30%) was due to changes in coil load. Differences in regional sensitivity due to B(1) inhomogeneity (parietal: 8% and 9%; insular: 16%) were found to be the second largest source of variation. Correction for local B(1) improved standard deviations and intra-subject reproducibility. On average, sensitivity was 9% less in insular than in parietal gray matter. If ignored, significant differences were introduced for water and N-acetyl-aspartate or were obscured for creatine and cholines. Hence, local sensitivity correction proved to be necessary for regional comparison of absolute metabolite concentrations. PMID- 11114064 TI - Hyperpolarized (129)Xe T (1) in oxygenated and deoxygenated blood. AB - The viability of the new technique of hyperpolarized (129)Xe MRI (HypX-MRI) for imaging organs other than the lungs depends on whether the spin-lattice relaxation time, T(1), of (129)Xe is sufficiently long in the blood. In previous experiments by the authors, the T(1) was found to be strongly dependent upon the oxygenation of the blood, with T(1) increasing from about 3 s in deoxygenated samples to about 10 s in oxygenated samples. Contrarily, Tseng et al. (J. Magn. Reson. 1997; 126: 79-86) reported extremely long T(1) values deduced from an indirect experiment in which hyperpolarized (129)Xe was used to create a 'blood foam'. They found that oxygenation decreased T(1). Pivotal to their experiment is the continual and rapid exchange of hyperpolarized (129)Xe between the gas phase (within blood-foam bubbles) and the dissolved phase (in the skin of the bubbles); this necessitated a complicated analysis to extract the T(1) of (129)Xe in blood. In the present study, the experimental design minimizes gas exchange after the initial bolus of hyperpolarized (129)Xe has been bubbled through the sample. This study confirms that oxygenation increases the T(1) of (129)Xe in blood, from about 4 s in freshly drawn venous blood, to about 13 s in blood oxygenated to arterial levels, and also shifts the red blood cell resonance to higher frequency. PMID- 11114065 TI - The effect of transient hypercapnia on task-related changes in cerebral blood flow and blood oxygenation in awake normal humans: a functional magnetic resonance imaging study. AB - It has recently been reported in alpha-chloralose anesthetized rats that the hemodynamic response to somatosensory stimulation almost doubled following transient hypercapnia (THC). In principle, this effect could be employed to enhance the sensitivity of perfusion-based fMRI experiments. To investigate whether a comparable effect was detectable in awake normal humans, changes in cerebral blood flow (DeltaCBF) and the effective transverse relaxation time (DeltaT(2)*) induced by a visual search task were measured in 10 healthy volunteers before and after THC. Concerning DeltaT(2)* no significant differences were found, whereas in four subjects DeltaCBF was significantly decreased (p < 0.01) following THC. These results demonstrate no increase in the CBF response following THC for awake humans. We conclude that the most likely explanation for this discrepancy with the earlier results obtained with animals is an as yet unknown mechanism of modulation of the cholinergic system by the anesthesia. PMID- 11114067 TI - Insight into protein structure and protein-ligand recognition by Fourier transform infrared spectroscopy. AB - An overview of the application of Fourier transform infrared spectroscopy for the analysis of the structure of proteins and protein-ligand recognition is given. The principle of the technique and of the spectra analysis is demonstrated. Spectral signal assignments to vibrational modes of the peptide chromophore, amino acid side chains, cofactors and metal ligands are summarized. Several examples for protein-ligand recognition are discussed. A particular focus is heme proteins and, as an example, studies of cytochrome P450 are reviewed. Fourier transform infrared spectroscopy in combination with the various techniques such as time-resolved and low-temperature methods, site-directed mutagenesis and isotope labeling is a helpful approach to studying protein-ligand recognition. PMID- 11114068 TI - Mimotopes: realization of an unlikely concept. AB - Theoretically it seems highly unlikely that relatively small peptides could mimic functionally discontinuous epitopes of antigens. Nevertheless various recent reports show this to be the case. Peptide mimics of protein-, polysaccharide- and DNA-epitopes have been shown to be able to replace the native epitope. Moreover, some of them are able to induce, when used in a vaccine, antibodies with the same activity as that of the antibody used as a template. These mimics, called mimotopes, can be used in vaccines and diagnostics and can be developed more or less systematically using solely antibodies and random, semi-random and dedicated peptide arrays or libraries. Furthermore, the mimotope concept which seems to have proven itself for antibody and antigen interaction can be applied equally well to many receptor ligand interactions and thus may form a new generic approach to the development of drugs. Ltd. PMID- 11114069 TI - New feature of angiotensin-converting enzyme: carbohydrate-recognizing domain. AB - Self carbohydrate-mediated dimerization of glycoprotein angiotensin-converting enzyme (ACE) was demonstrated. The dimerization was studied in the reverse micelle experimental system as a model of biomembrane situation. Asialo-ACE or agalacto-ACE was able to form a dimer, whereas deglycosylated ACE and sequentially desialylated and degalactosylated ACE failed to dimerize. ACE-ACE interaction was competitively inhibited by Neu5Ac- or Gal-terminated saccharides. The results have allowed us to propose the existence of carbohydrate-recognizing domain (CRD) on ACE molecule. The structural requirements of this CRD were estimated based on the ability of saccharides to inhibit ACE dimerization. The most effective monosaccharides with equal inhibition potencies were shown to be galactose (as GalbetaOMe) and N-acetylneuraminic acid (as Neu5AcalphaOMe). Among oligosaccharides, the most effective ones were found to be 3'SiaLac and, especially, the whole pool of ACE oligosaccharide chains and biantennae complex oligosaccharide chains of other glycoproteins. Bovine and human ACEs were shown to be similar in terms of recognition of carbohydrates. PMID- 11114070 TI - Design, synthesis and evaluation of biomimetic affinity ligands for elastases. AB - A low-molecular-weight biomimetic affinity ligand selective for binding elastase has been designed and synthesized. The ligand was based on mimicking part of the interaction between a natural inhibitor, turkey ovomucoid inhibitor and elastase, and modelled from the X-ray crystallographic structure of the enzyme-inhibitor complex. Limited solid-phase combinatorial chemistry was used to synthesize 12 variants of the lead ligand using the triazine moiety as the scaffold for assembly. The ligand library was screened for its ability to bind elastase and trypsin, and two ligands were studied further. Ligand C4/6 [2-alanyl-alanyl-4 tryptamino-6-(alpha-lysyl)-s-triazine] was found to bind porcine pancreatic elastase, but not trypsin, with a dissociation constant of 6 x 10(-5) M and a binding capacity of 21 mg elastase per ml gel. The adsorbent was used to purify elastase from a crude extract of porcine pancreas. Immobilized ligand C4/5 6 [2 alanyl-alanyl-4-tyramino-6-(alpha-lysyl)-s-triazine] was similarly chosen for optimal binding of elastase from cod and used to purify the enzyme from a crude extract of cod pyloric caeca. Ligand C4/6 was subsequently synthesized in solution and its structure verified by 1H-NMR. PMID- 11114072 TI - Survey of the 1999 surface plasmon resonance biosensor literature. AB - The application of surface plasmon resonance biosensors in life sciences and pharmaceutical research continues to increase. This review provides a comprehensive list of the commercial 1999 SPR biosensor literature and highlights emerging applications that are of general interest to users of the technology. Given the variability in the quality of published biosensor data, we present some general guidelines to help increase confidence in the results reported from biosensor analyses. PMID- 11114071 TI - Phage-displayed peptides as biosensor reagents. AB - This study investigated the potential to utilize phage-displayed peptides as reagents in sensor applications. A library of random 12-mers displayed on phage was panned against staphylococcal enterotoxin B (SEB), a causative agent of food poisoning. Nine SEB binding phage clones were isolated, all of which share the consensus sequence Trp His Lys at their amino terminus. Binding of several of these phage was shown to be inhibited when they were assayed in a competitive enzyme-linked immunosorbent assay (ELISA) format with synthesized peptide corresponding to the peptide-encoding region of one of the clones. Whole phage were labeled with the dye Cy5, and incorporated into fluoroimmunoassays. Labeled phage were able to detect SEB down to a concentration of 1.4 ng/well in a fluorescence-based immunoassay. When incorporated into an automated fluorescence based sensing assay, Cy5-labeled phage bound to probes coated with SEB generated a robust signal of about 10,000 pA, vs a signal of 1,000 pA using a control fiber coated with streptavidin. These results demonstrate the potential for development of phage-based sensor reagents. PMID- 11114073 TI - Skerra A, 2000. Engineered scaffolds for molecular recognition. Journal of Molecular Recognition13:167-187. PMID- 11114074 TI - Biological synergism between infectious agents. PMID- 11114075 TI - Spliced segments at the 5' terminus of adenovirus 2 late mRNA. 1977. PMID- 11114076 TI - Herpes simplex virus: discovering the link between heparan sulphate and hereditary bone tumours. AB - To gain entry into the host, viruses use host cell surface molecules that normally serve as receptors for other ligands. Herpes simplex virus type 1 (HSV 1) uses heparan sulphate (HS) glycosaminoglycans (GAGs) as receptors for initial attachment to the host cell surface. HS GAGs are both ubiquitous and structurally diverse, and normally serve as critical mediators of interactions between the cell and the extracellular environment. We have used the HS binding ability of HSV-1 to identify the function of a cellular gene, EXT1, which is involved in HS polymerisation. Cellular factors that affect virus growth and replication are often key regulators of the cell cycle and EXT1 is no different-humans with inherited mutations in EXT1 have developmental defects that lead to bone tumours (hereditary multiple exostoses, HME) and sometimes chondrosarcomas. Thus, as a result of using HSV-1 as a molecular probe, a functionally orphaned disease gene now has a defined function. These findings highlight the utility of viruses for investigating important cellular processes. PMID- 11114077 TI - Limits to potent antiretroviral therapy. AB - The availability of potent combination antiretroviral therapy (ART), also known as highly active antiretroviral therapy or HAART has changed the prognosis of HIV infection. However, the benefits have to be seen in the context of deficiencies of current therapy: failure to eradicate the virus, the slow and potentially incomplete recovery of the immune system, the high prevalence of resistance, and the potential for long-term toxicity. Treatment strategies need to take into account these limits to better target those HIV-infected patients who could benefit the most from antiretroviral therapy. PMID- 11114078 TI - Innate defences against viraemia. AB - Human blood plasma has been reported to possess nonspecific antiviral activity. This activity is due to several preexisting naturally occurring molecules that are either active against individual members or a family of viruses. These molecules, however, have not been adequately studied to reveal their molecular structures and mechanisms of action presumably because of their low and nonspecific antiviral action. Therefore, their possible role against viraemia remains unknown. Recently, two naturally occurring nonspecific broad-spectrum antiviral agents, University of Texas Inhibitor beta (UTIbeta) glycoprotein and high density lipoprotein, have been described in human serum. They are active against DNA and RNA viruses and one of them, UTIbeta, possesses significant antiviral activity of 40 units/mL. Since preexisting antiviral molecules in serum appear to be the only defence mechanisms available at the onset of viral infection they may have protective significance against viraemia. In view of this potential, we have undertaken to review the properties of these innate viral inhibitory molecules. PMID- 11114080 TI - Secondary ion-molecule reactions in matrix-assisted laser desorption/ionization AB - Ion-molecule charge- and proton-transfer reactions in the desorption plume are considered for the case of matrix-assisted laser desorption/ionization (MALDI) with ultraviolet laser excitation, and it is proposed that they are major determinants of the observed mass spectrum. Specific MALDI phenomena which are discussed include the dominance of singly charged ions and analyte-matrix or analyte-analyte signal suppression. Should any be formed, highly charged products can be reduced by reaction with neutral matrix, yet singly charged ions cannot generally be neutralized in the same manner. Ion suppression effects can also be explained by similar reactions, which in some cases involve interconversion of dissimilar ion types. The plume is proposed often to be more under thermodynamic rather than kinetic control owing to these secondary reactions. UV/MALDI mass spectra should therefore be largely predictable, given sufficient thermodynamic information, and appropriate experimental conditions of sufficient analyte and plume density. Copyright 2000 John Wiley & Sons, Ltd. PMID- 11114081 TI - Identification and characterization of Podophyllum emodi by API-LC/MS/MS. AB - An API-LC/MS/MS method was developed for the identification of the medicinal herb Podophyllum emodi based on the profile of its aryltetrahydronapthalene and related lignan marker compounds. This was done by matching the structural information from the tandem mass spectrometric data with those lignan marker compounds already reported for the herb. The method could be employed in the absence of reference standards for the markers and was particularly useful in view of the scarcity of supply of these chemical standards. It has been used successfully to differentiate Podophyllum emodi from two commonly used medicinal herbs of a different genus but having similar appearance, Radix clematidis and Radix gentiana, as well as a closely related herb, Podophyllum peltatum. PMID- 11114082 TI - High-resolution LC/MS for analysis of minor components in complex mixtures: negative ion ESI for identification of impurities and degradation products of a novel oligosaccharide antibiotic. AB - High-resolution mass spectrometry has been routinely used for structural confirmation and identification; however, it has mostly been applied to relatively pure samples. Exact mass measurement of minor components such as impurities, degradation products or metabolites in complex mixtures has been difficult without prior separation and isolation. Here we report the utilization of on-line liquid chromatography in combination with high-resolution mass spectrometry for the identification of impurities and base degradation products of Sch 27899, a member of the everninomicin class of antibiotics. Nine Sch 27899 related impurities and degradation products were detected by negative ion electrospray ionization using a magnetic sector mass spectrometer. Exact mass measurements were obtained at a resolution of 5000 using polyethylene glycol (PEG) sulfates as internal standards. Corresponding elemental compositions were determined within a 2 ppm error tolerance and structures were proposed for all components. PMID- 11114083 TI - Characterization of metabolites of Celecoxib in rabbits by liquid chromatography/tandem mass spectrometry. AB - The metabolism of the anti-inflammatory drug Celecoxib in rabbits was characterized using liquid chromatography (LC)/tandem mass spectrometry (MS/MS) with precursor ion and constant neutral loss scans followed by product ion scans. After separation by on-line liquid chromatography, the crude urine samples and plasma and fecal extracts were analyzed with turbo-ionspray ionization in negative ion mode using a precursor ion scan of m/z 69 (CF(3)) and a neutral loss scan of 176 (dehydroglucuronic acid). The subsequent product ion scans of the [M H] ions of these metabolites yielded the identification of three phase I and four phase II metabolites. The phase I metabolites had hydroxylations at the methyl group or on the phenyl ring of Celecoxib, and the subsequent oxidation product of the hydroxymethyl metabolite formed the carboxylic acid metabolite. The phase II metabolites included four positional isomers of acyl glucuronide conjugates of the carboxylic acid metabolite. These positional isomers were caused by the alkaline pH of the rabbit urine and were not found in rabbit plasma. The chemical structures of the metabolites were characterized by interpretation of their product ion spectra and comparison of their LC retention times and the product ion spectra with those of the authentic synthesized standards. PMID- 11114084 TI - Mass spectrometry of bis-quinolizidine alkaloids: 2- and 17-alkyl-substituted derivatives of sparteine and lupanine. AB - The mass spectral fragmentations of 2-methylsparteine (1), 2, 17 dimethylsparteine (2), 2-methyl-17-isopropylsparteine (3), 2-methyl-17 oxosparteine (4), 2-oxo-17-methylsparteine (17-methyllupanine) (5) and 2-oxo-17 isopropylsparteine (17-isopropyllupanine) (6) were investigated. Fragmentation pathways, whose identification was assisted by accurate mass measurements and a correlation between the abundances of the M(+.) and selected fragment ions of the investigated compounds, are discussed. The data obtained create the basis for distinguishing the structural isomers and metamers. PMID- 11114085 TI - Using surfactants to enhance the analyte signals in activated carbon, surface assisted laser desorption/ionization (SALDI) mass spectrometry. AB - The effect of surface activity in surface-assisted laser desorption/ionization (SALDI) mass spectrometry was examined. Several surfactants, including p tolunensulfonic acid (PTSA), sodium dodecyl sulfate and alkyltrimethylammonium bromide, were used as analytes or additives in the SALDI matrix to demonstrate the surface activity effect. The experimental results demonstrate that analytes that have good surface activity have good sensitivity. Adding suitable amounts of surfactants to the SALDI matrix can dramatically enhance the sensitivity of analytes lacking surface activity. We propose that the enhancement of analyte signals is due to the ionic interaction between ionic surfactants and analytes because non-ionic surfactant additives in the SALDI matrix do not affect the analyte signals. The detection limit of methylephedrine can be as low as 100 pg in the SALDI analysis of 0.5 M PTSA additive in the SALDI matrix. Although other surfactants can also be used as matrix additives to enhance the analyte signal, they do not improve the ion abundance as much as PTSA does. PMID- 11114086 TI - Derivatization procedures for the detection of beta(2)-agonists by gas chromatographic/mass spectrometric analysis. AB - An evaluation of derivatization procedures for the detection of beta(2)-agonists is presented. The study was performed with the beta(2)-agonists bambuterol, clenbuterol, fenoterol, formoterol, salbutamol, salmeterol and terbutaline. Different derivatizating agents were employed, aiming to obtain derivatives with high selectivity to be used in the gas chromatographic/mass spectrometric analysis of beta(2)-agonists in biological samples. Trimethylsilylation was compared with different agents and the role of some catalysts was evaluated. Acylation, combined trimethylsilylation and acylation, and the formation of cyclic methylboronates were also studied. Sterical hindrance caused by different substituents at the nitrogen atom of the beta-ethanolamine lateral chain of beta(2)-agonist molecules is mainly responsible for differences in the abundances of the derivatives obtained. The use of catalysts produces an increase in the derivatization yield, especially for compounds with low steric hindrance (substituents with primary and secondary carbon atoms). The formation of trimethylsilyl (TMS) ethers is not influenced by structural molecular differences when only hydroxy groups are involved in derivatization. Combined trimethylsilylation and acylation showed that compounds with a secondary carbon atom linked to the nitrogen atom form mainly N-TFA-O-TMS derivatives, with a small amount of N-TMS-O-TMS derivatives. Compounds with tert-butyl substituents at the amino group (bambuterol, salbutamol and terbutaline) formed O-TMS derivatives as the main products, although a limited amount of trifluoroacylation at the nitrogen atom also occurred. Cyclic methylboronates were formed with bambuterol, clenbuterol, formoterol, salbutamol and salmeterol. Owing to hydroxy substituents in unsuitable positions for ring formation, this procedure was not effective for fenoterol and terbutaline. Mass spectra of different derivatives and tentative fragmentation profiles are also shown. For screening purpose (e.g. sports drug testing), derivatization with MSTFA or BSTFA alone is recommended as a comprehensive derivatization technique for beta(2)-agonists owing to minimal by product formation; formation of cyclic methylboronates can be useful for confirmation purposes. Detection limits were obtained for the TMS and cyclic methylboronate derivatives using the derivatizing reagents MSTFA and trimethylboroxine, respectively. For most of the compounds, lower detection limits were found for the TMS derivatives. PMID- 11114087 TI - Electrospray ionization mass spectrometric study of Cu(I) and Cu(II) bipyridine complexes employed in atom transfer radical polymerization AB - We report an electrospray ionization mass spectrometric study of Cu(I) and Cu(II) bipyridine complexes employed in atom transfer radical polymerization. Mass spectra of Cu(I)Br complexed with 2 equiv. of 4,4'-di(5-nonyl)-2,2'-bipyridine (dNbpy) in toluene, methyl acrylate or styrene showed the presence of [Cu(I)(dNbpy)(2)](+) cation and [Cu(I)Br(2)](-) anion. For the Cu(II)Br(2)/2dNbpy system, [Cu(II)(dNbpy)(2)Br](+), [Cu(II)(dNbpy)Br](+), [Cu(I)Br(2)](-), [Cu(II)Br(3)](-) and [Cu(II)(dNbpy)Br(3)](-) species were observed. In addition, for mixed Cu(I)Br/2dNbpy and Cu(II)Br(2)/2dNbpy systems, the negative ion mode showed only the presence of [Cu(I)Br(2)](-) anions, which are potentially formed through halogen exchange between [Cu(II)Br(3)](-) and [Cu(I)(dNbpy)(2)](+). Copyright 2000 John Wiley & Sons, Ltd. PMID- 11114088 TI - Post-source decay fragmentation analyses of linkage isomers of Lewis-type oligosaccharides in curved-field reflectron matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: combined in-source decay/post-source decay experiments and relative ion abundance analysis. AB - Linkage isomers of Lewis(X) trisaccharide (Le(X)) and Lewis(a) trisaccharide (Le(a)) were distinguished by the post-source decay (PSD) fragment spectra obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) without permethylation. Both Y- and Z-type fragmentations were observed at the C-3 position of N-acetylhexosamine. beta Elimination at C-3 of the reducing-end N-acetylglucosamine in Le(X) formed a double bond, which conjugated to an N-acetyl group, making the chemical species stable. In contrast, the double bond formed in the reducing end glucose of 3 fucosyllactose was unstable owing to the lack of a conjugated system. Therefore, beta-elimination of N-acetylglucosamine occurred predominantly rather than that of hexose in MALDI-PSD fragmentation. The measurements of the PSD fragment mass spectra using pseudo precursor ions originating from in-source decay were useful for the analyses of the fragmentation mechanisms and for the assignments of the chemical species of the fragment ions. The combined in-source decay/post-source decay experiments revealed the formation of a double bond between C-2 and C-3 in N-acetylglucosamine of Le(X). Abundance analysis of the PSD ions indicated that the 1-3 glycosyl linkage cleaves more easily than does the 1-4 linkage in MALDI PSD fragmentation. Ion abundance analyses were useful in estimating the degree of Y- and Z-type fragmentation at the C-3 position of hexose and N-acetylhexosamine. The analysis of the relative ion abundances was a powerful tool for the assignments of the chemical species of the PSD ions. PMID- 11114089 TI - Direct sample fraction deposition using electrospray in narrow-bore size exclusion chromatography/matrix-assisted laser desorption/ionization time-of flight mass spectrometry for polymer characterization AB - A direct sample fraction deposition method was developed for off-line size exclusion chromatography (SEC)/matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry. By using electrospray, the SEC eluent, together with a suitable matrix solution added coaxially, was directly deposited on the MALDI plate. Owing to the formation of very small droplets in electrospray, solvent evaporation is much faster. The fractionation volume in narrow-bore SEC, which can directly be collected in one MALDI spot, can easily be optimized in the range of a few microlitres. In addition, fairly homogeneous sample spots were obtained. The possible influence of composition variation of the SEC effluent on the analytical results using direct fraction deposition was investigated; no substantial effects were observed. The applicability of the method was demonstrated by characterizing a broad poly(methyl methacrylate) sample. Copyright 2000 John Wiley & Sons, Ltd. PMID- 11114090 TI - Determination of lodenosine and its major metabolite in human plasma by liquid chromatography/electrospray ionization tandem mass spectrometry. AB - A sensitive and selective method for the determination of 2'-beta-fluoro-2',3' dideoxyadenosine (lodenosine, F-ddA), an experimental anti-AIDS drug, and its major metabolite, 2'-beta-fluoro-2',3'-dideoxyinosine (F-ddI), in human plasma was developed and validated. The procedure employs two internal standards and a simple ultrafiltration step followed by chromatography on a Betasil C(18) minibore column. An in-line valve is used to remove salts before reaching the ion source. Detection is by electrospray ionization tandem mass spectrometry with selected reaction monitoring. The method has a limit of quantitation of 4 ng ml( 1) (16 nM) for F-ddA and 8 ng ml(-1) (32 nM) for F-ddI with a linear range up to 2000 ng ml(-1) (7.9 microM) for each. Predicted concentrations from a three-day validation study were within 5% of the nominal values for F-ddA and 16% for F ddI. Intra- and inter-assay precision, as measured by relative standard deviation, was 13% or better for both compounds. To achieve good reproducibility, many variables related to the electrospray ionization were optimized for both precision and sensitivity. The method was successfully employed to analyze samples and evaluate plasma pharmacokinetics from a Phase I clinical trial. PMID- 11114091 TI - Influence of the laser fluence in infrared matrix-assisted laser desorption/ionization with a 2.94 microm Er : YAG laser and a flat-top beam profile. AB - The dependence of the signal intensity of analyte and matrix ions on laser fluence was investigated for infrared matrix-assisted laser desorption/ionization (IR-MALDI) mass spectrometry using a flat-top laser beam profile. The beam of an Er : YAG laser (wavelength, 2.94 microm; pulse width, 90 ns) was coupled into a sapphire fiber and the homogeneously illuminated end surface of the fiber imaged on to the sample by a telescope. Three different laser spot sizes of 175, 350 and 700 microm diameter were realized. Threshold fluences of common IR matrices were determined to range from about 1000 to a few thousand J m(-2), depending on the matrix and the size of the irradiated area. In the MALDI-typical fluence range, above the detection threshold ion signals increase strongly with fluence for all matrices, with a dependence similar to that for UV-MALDI. Despite the strongly different absorption coefficients of the tested matrices, varying by more than an order of magnitude at the excitation laser wavelength, threshold fluences for equal spot sizes were found to be comparable within a factor of two. With the additional dependence of fluence on spot size, the deposited energy per volume of matrix at threshold fluence ranged from about 1 kJ mol(-1) for succinic acid to about 100 kJ mol(-1) for glycerol. PMID- 11114092 TI - Determination of ondansetron and its hydroxy metabolites in human serum using solid-phase extraction and liquid chromatography/positive ion electrospray tandem mass spectrometry. AB - Ondansetron and its hydroxylated metabolites were determined in human serum using solid-phase extraction (SPE) and liquid chromatography/positive ion electrospray tandem mass spectrometry. Pyrimethamine was used as the internal standard. The analytes were eluted from the SPE cartridge using 2 x 1 ml of methanol containing 0.5% triethylamine, evaporated under vacuum and the residue was reconstituted in the mobile phase. The liquid chromatographic separation was achieved on a silica column using a mobile phase of aqueous 20 mM ammonium acetate (pH 4.7) acetonitrile (85 : 15, v/v) at a flow-rate of 0.4 ml min(-1). The method was linear over the range 1-500 ng ml(-1) for ondansetron and each of the metabolites in human serum. The intra-day accuracy was better than 9.1% and the precision was <10.3%; the inter-day accuracy was better than 9.5% and the precision was <12.6%. The limit of detection was 250 pg ml(-1) based on a signal-to-noise ratio of 3. The absolute recovery from serum for all analytes was >90%. PMID- 11114093 TI - The information encrypted in accurate peptide masses-improved protein identification and assistance in glycopeptide identification and characterization. AB - Analytically useful information from accurate mass data for peptides with an error of 0.99 and goodness of fit <10%). Limits of quantification of 25.0 ng g(-1) were obtained for the determination of gentamicin in muscle, fat, liver and kidney tissues of swine and calf, which correspond in all cases to at least half the MRLs. Limits of detection ranged between 0.5 and 2.5 ng g(-1) for the tissues. The within-day and between-day precisions (RSD) and the results for accuracy fell within the ranges specified. The method was successfully used for the determination of gentamicin in tissue samples of swines and calves medicated with gentamicin by intramuscular injection. PMID- 11114095 TI - Proton affinity of peroxyacetyl nitrate. A computational study of topical proton affinities. AB - The structure and energetics of the peroxyacetyl nitrate conformers syn- and anti PAN and several cations formed by PAN protonation were investigated by a combination of density functional theory and ab initio calculations. syn-PAN is the more stable conformer that is predicted to predominate in gas-phase equilibria. The acetyl carbonyl oxygen was found to be the most basic site in PAN, the oxygen atoms of the peroxide and NO(2) groups being less basic. The 298 K proton affinity of syn-PAN was calculated as 759-763 kJ mol(-1) by effective QCISD(T)/6-311 + G(3df,2p) and 771-773 kJ mol(-1) by B3-MP2/6-311 + G(3df,2p). The calculated values are 25-39 kJ mol(-1) lower than the previous estimate by Srinivasan et al. (Rapid Commun. Mass Spectrom. 1998; 12: 328) that was based on competitive dissociations of proton-bound dimers (the kinetic method). The calculated threshold dissociation energies predicted the formation of CH(3)CO(+) + syn - HOONO(2) and CH(3)COOOH + NO(2)(+) to be the most favorable fragmentations of protonated PAN that required 83 and 89 kJ mol(-1) at the respective thermochemical thresholds at 298 K. The previously observed dissociation to CH(3)COOH + NO(3)(+) was calculated by effective QCISD(T)/6-311 + G(3df,2p) to require 320 kJ mol(-1). The disagreement between the experimental data and calculated energetics is discussed. PMID- 11114096 TI - High-throughput screening of tocopherols in natural extracts. PMID- 11114097 TI - Current awareness. PMID- 11114099 TI - Insulin receptors and insulin actions in the nervous system. AB - Insulin receptors are widely distributed in the brain. They are also present in peripheral nerve. Insulin signaling through its receptors in the brain is responsible for the hormone's effects on the regulation of food intake, body weight, and reproduction. Signaling through the insulin receptor also appears to influence higher cognitive functions. In peripheral nerve, insulin signaling may play a role in the maintenance and repair of myelinated fibers. Future studies should determine the extent to which a defective insulin signal may be linked to the pathogenesis of diabetic neuropathies and neurodegenerative disorders such as Alzheimer's disease. PMID- 11114100 TI - Role of vasoactive factors in the pathogenesis of early changes in diabetic retinopathy. AB - Several interactive and mutually perpetuating abnormal biochemical pathways, such as protein kinase C (PKC) activation, augmented polyol pathway, and non-enzymatic glycation, may be activated as a result of sustained hyperglycemia in diabetes. These abnormal pathways may in turn influence several vasoactive factors, which are probably instrumental in the production of functional and morphological changes in the retina in diabetes. The vasoactive factors such as endothelins, nitric oxide, vascular endothelial growth factors, etc., are of importance in mediating functional and structural alterations in early diabetic retinopathy. Intricate and interactive regulatory mechanism(s) among these factors may control ultimate availability of these molecules to produce biologically significant effects. A better understanding of these factors and their interactions would aid the development of adjuvant therapies for the treatment of diabetic retinopathy. PMID- 11114101 TI - Diabetic neuropathy--a continuing enigma. AB - In this article we will review the clinical signs and symptoms of diabetic somatic polyneuropathy (DPN), its prevalence and clinical management. Staging and classification of DPN will be exemplified by various staging paradigms of varied sophistication. The results of therapeutic clinical trials will be summarized. The pathogenesis of diabetic neuropathy reviews an extremely complex issue that is still not fully understood. Various recent advances in the understanding of the disease will be discussed, particularly with respect to the differences between neuropathy in the two major types of diabetes. The neuropathology and natural history of diabetic neuropathy will be discussed pointing out the heterogeneities of the disease. Finally, the various prospective therapeutic avenues will be dealt with and discussed. PMID- 11114102 TI - Insulin receptor substrate (IRS) transduction system: distinct and overlapping signaling potential. AB - Insulin receptor substrate (IRS) proteins play a central role in maintaining basic cellular functions such as growth and metabolism. They act as an interface between multiple growth factor receptors possessing tyrosine kinase activity, such as the insulin receptor, and a complex network of intracellular signalling molecules containing Src homology 2 (SH2) domains. Four members (IRS-1, IRS-2, IRS-3, IRS-4) of this family have been identified which differ in their subcellular distribution and interaction with SH2 domain proteins. In addition, differential IRS tissue- and developmental-specific expression patterns may contribute to specificity in their signaling potential. PMID- 11114103 TI - Glutamic acid decarboxylase antibodies (GADA) is the most important factor for prediction of insulin therapy within 3 years in young adult diabetic patients not classified as Type 1 diabetes on clinical grounds. AB - BACKGROUND: Differentiation between Type 1 and Type 2 diabetes in adults is difficult at diagnosis. In this study we tested the hypothesis that autoantibodies at diagnosis are predictive for insulin treatment within 3 years in patients initially not classified as Type 1 diabetes. METHODS: In a nationwide population-based study, blood samples were obtained from 764 patients, all diagnosed with diabetes during a 2-year period. At diagnosis, 583 (76%) were classified as Type 1, 110 (14%) as Type 2 and 71 (9.3%) could not be classified. RESULTS: Among patients not classified as Type 1 diabetes, 52 (47%) of Type 2 and 42 (59%) of unclassified patients were positive for islet cell antibodies (ICA), glutamic acid decarboxylase antibodies (GADA) or tyrosine phosphatase antibodies (IA-2A). These patients (n=94) had lower body mass index (BMI) (p<0.001) and lower C-peptide (p<0.001) compared to the autoantibody negative patients (n=87). Compared to clinically classified Type 1 diabetes patients positive for autoantibodies (n=477), they have higher BMI (p<0.001), higher C-peptide (p<0.001) and the same levels of ICA, GADA and IA-2A. After 3 years, 93% of autoantibody positive patients initially not classified as Type 1 were on insulin. When ICA, GADA, IA-2A, BMI and C-peptide were tested in a multiple logistic regression, only GADA was significant for insulin treatment within 3 years (OR=18.8; 95% CI 1.8-191) in patients treated with diet or oral drugs at diagnosis. CONCLUSIONS: A correct classification is difficult in adult diabetic patients. The presence of pancreatic autoantibodies, especially GADA, at diagnosis of diabetes are highly predictive for insulin therapy within 3 years from diagnosis. PMID- 11114105 TI - Pancreatitis in fibrocalculous pancreatic diabetes mellitus is not associated with common mutations in the trypsinogen gene. AB - BACKGROUND: A distinct type of pancreatitis associated with diabetes, termed fibrocalculous pancreatic diabetes (FCPD), has been reported in tropical developing countries including Bangladesh. The molecular basis for autosomal dominant hereditary pancreatitis (HP) has recently been attributed to mutations in exons 2 and 3 of the trypsinogen gene. We have investigated the hypothesis that mutations in the aforementioned exons of this gene might also predispose to FCPD. METHODS: Seventy Bangladeshi and 50 South Indian unrelated FCPD patients and seven South Indian families with FCPD probands were studied. Pancreatic calcification was confirmed by abdominal X-ray, ultrasound and/or ERCP. Established mutations of exons 2 and 3 of the trypsinogen gene were studied in these subjects by PCR-RFLP analysis and DNA sequencing. RESULTS: The mutations found in hereditary pancreatitis were not observed in this collection of FCPD subjects, and complete DNA sequencing of exons 2 and 3 of the fourth cationic trypsinogen gene also excluded any new mutations. CONCLUSIONS: These results indicate that chronic pancreatitis of FCPD is unlikely to be caused by common mutations in the trypsinogen gene. PMID- 11114104 TI - Insulin resistance, cardiovascular risk factors and ultrasonically measured early arterial disease in normotensive Type 2 diabetic subjects. AB - BACKGROUND: The objective of the study was to examine the relationship between serum fasting insulin, insulin sensitivity S(i), cardiovascular risk factors, and asymptomatic early atherosclerosis in normotensive Type 2 diabetic subjects. METHODS: Specific insulin was measured using an enzyme-linked immunosorbent assay (ELISA) and insulin sensitivity was assessed with an insulin-modified frequently sampled intravenous glucose tolerance test (FSIVGTT). Early atherosclerotic change was assessed using carotid intima media thickness (IMT) and an arterial ultrasound score (AUS) measured by high-resolution B-mode ultrasound. RESULTS: On bivariate analysis, there was a positive correlation between S(i) and high density lipoprotein (HDL) cholesterol (r(s)=0.27, p<0.05), and a negative correlation between S(i) and body mass index (BMI) (r(s)=-0.42, p<0.001), HbA(1c) (r(s)=-0.29, p<0.05) and serum triglyceride (r(s)=-0.30, p<0.05). There was a positive correlation between carotid IMT and age (r(s)=0.41, p<0.0005), and a positive association with male sex (p<0.0001) as well as with smoking (p<0.0001). However, we found no correlation between carotid IMT and fasting specific insulin (r(s)=-0.04) or S(i) (r(s)=-0.08). On multiple regression analyses, only age and serum triglycerides appeared to be significant independent variables with respect to carotid IMT whereas age, male sex and smoking emerged jointly significant with respect to AUS. There were no independent associations between carotid IMT or AUS with other variables including using either fasting specific insulin or S(i) as markers on insulin resistance separately. CONCLUSION: Carotid IMT and AUS in Type 2 diabetes are closely associated with age, male sex and smoking. The relationships between serum insulin and insulin resistance with ultrasonically measured early arterial disease in Type 2 diabetes remain unclear. PMID- 11114106 TI - Current literature in diabetes. PMID- 11114107 TI - Editorial. PMID- 11114108 TI - Luminescent cryptands. 3-aroylcoumarin macrobicyclic complexes of europium(III) and terbium(III): the effect of coumarin substitution. AB - Substituted 3-aroylcoumarins incorporated in a polyethylenoxy cryptand (2.2.2) by the 3-aroyl group were synthesized with the purpose of developing new markers to be used in time-resolved fluorimetric bioaffinity assays based on the unique luminescence properties of Eu(III) and Tb(III) ions. Some spectral properties and luminescence intensities of the complexes were measured in acetonitrile and MeOH. The mechanism of metal sensitization depended on lanthanide and will be discussed in detail. PMID- 11114109 TI - A time-resolved study of the mechanism of the energy transfer from a ligand to the lanthanide(III) ion in solutions and solid films. AB - The photochemical properties of some lanthanide chelates developed for immunohistochemistry have been studied in water and in solid Langmuir-Blodgett films. The fluorescence and triplet-state lifetimes of 4-(phenylethynyl)pyridine (PET), di[(phenylethynyl)pyridine] (D-PET), phenylterpyridine (PTP) and their tetra- or penta-acid derivatives (-TA or -PA) were measured in the presence and absence of Gd(III)-, Tb(III)- and Eu(III)-ions. The mechanism for the total process and the rate constants and quantum yields for the individual reaction steps and for the total process were determined in water solution. Time-resolved absorption and luminescence methods were also used to study the energy transfer between an amphiphilic 4-[4-[(C(10)H(12))(2)NCO]phenylethynyl]-pyridine tetra acid (A-PET-TA) and the Tb(III)- and Eu(III)-ions in solid Langmuir-Blodgett films. Luminescence and transient absorption rate constants were determined. PMID- 11114110 TI - Zeptomole detection sensitivity of prostate-specific antigen in a rapid microtitre plate assay using time-resolved fluorescence. AB - Prostate-specific antigen (PSA) was detected in microtitre wells coated with a PSA-specific antibody using biotinylated antibody and streptavidin-coated, highly fluorescent 107 nm nanoparticles, which contained more than 30000 europium ions entrapped by beta-diketones. PSA was monitored directly on the surface of a well without any additional enhancement step. The sensitivity of the assay was 1.6 ng/L, corresponding to 50 fmol/L or 250 zeptomoles (250 x 10(-21) mol/L) of PSA. The high specific activity and low non-specific binding of the streptavidin coated nanoparticles improved the sensitivity of the PSA assay 100-fold compared to the conventional europium-labelled streptavidin tracer in the same assay format. Additionally, the streptavidin-coated nanoparticle label made very rapid assays possible, due to the high affinity of the streptavidin-biotin complex and a high number of binding sites available for tracing the biotinylated antibody on the surface. Due to the inherent problems of tracing analyte with a complex of biotinylated antibody and streptavidin-coated nanoparticles, the streptavidin coated nanoparticles reacting with the surface-captured analyte and biotinylated antibody was favoured and factors influencing this are discussed. This universal labelling technology can be applied to detect any biotinylated molecule, either in solution or on a solid phase, in order to improve detection sensitivities in many areas of biochemical analysis, such as cyto- and histochemistry, multianalyte DNA-chip assays and single-particle assays. PMID- 11114111 TI - The measurement of prion protein in bovine brain tissue using differential extraction and DELFIA as a diagnostic test for BSE. AB - A simple diagnostic test for the detection of bovine spongiform encephalopathy (BSE), based on a commercially available time-resolved fluorescence immunoassay (DELFIA) for the measurement of the normal and disease-associated isoforms of prion protein (PrP), is described. The isoforms are sequentially extracted from homogenized bovine brain tissue using two concentrations of guanidine hydrochloride. This procedure initially extracts a soluble isoform and subsequently a less soluble disease-associated aggregated isoform. Following quantification of the two fractions, the percentage of the insoluble prion becomes a measurable parameter, independent of protein concentration, clearly identifying normal from infected animals displaying clinical signs of BSE. The mean percentages of insoluble PrP in brain tissue from 60 BSE-confirmed-positive cattle and 100 cattle that had never been exposed to the disease were 52.6% (SD = 22.8) and 3.9% (SD = 1.5), respectively. The assay is sensitive, with a detection limit of less than 50 pg PrP, and is robust and precise (CVs < 10%) over the appropriate working range. PMID- 11114112 TI - Determination of methyl anthranilate in food samples by coupling stopped-flow mixing technique and time-resolved luminescence detection. AB - A sensitive and fast approach for the determination of methyl anthranilate in grape must and honey samples, using time-resolved luminescence measurements, has been reported for the first time. The method involves the alkaline hydrolysis of the ester to anthranilic acid and the formation of a chelate with terbium(III) and tri-n-octylphosphine oxide in presence of Triton X-100. Kinetic and equilibrium measurements were obtained in 0.1 and 15 s, respectively, by using a stopped-flow mixing technique. The dynamic ranges of the calibration graphs of the kinetic and equilibrium methods were 21.9 nmol/L-29.2 micromol/L and 19.7 nmol/L-21.9 micromol/L, respectively, and the detection limits were 7.3 and 6.6 nmol/L, respectively. The precision, expressed as relative standard deviation, was less than 3%. Although both-kinetic and equilibrium methods exhibited very similar analytical features, only the better selectivity of the former allowed the content of methyl anthranilate to be determined in the samples, as the initial rate measurements avoided the negative effect that the sample matrix caused in the equilibrium measurements. The analytical recoveries obtained by applying the kinetic method to the analysis of grape must and flower honey samples were in the range 92.5-105.0%. PMID- 11114113 TI - Detection of unwanted residues of ivermectin in bovine milk by dissociation enhanced lanthanide fluoroimmunoassay. AB - Avermectins are frequently used to control parasitic infestations in many animal species. Previous studies have shown the long-term persistence of unwanted residues of these drugs in animal tissues and fluids. An immunoassay screening test for the detection and quantification of ivermectin residues in bovine milk has been developed. After an extensive extraction procedure, milk samples were applied to a competitive dissociation-enhanced lanthanide fluoroimmunoassay using a monoclonal antibody against an ivermectin-transferrin conjugate. The monoclonal antibody, raised in Balb C mice, showed cross-reactivity with eprinomectin (92%), abamectin (82%) and doramectin (16%). The limit of detection of the assay (mean + 3 SD), calculated from the analysis of 17 known negative samples, was calculated as 4.6 ng/mL. Intra- and inter-assay RSDs were determined as 11.6% and 15.8%, respectively, using a negative bovine milk sample fortified with 25 ng/mL ivermectin. Six Friesian milking cows were treated with ivermectin, three with a pour-on formulation of the drug and three with an injectable solution at the manufacturer's recommended dose rate. An initial mean peak in ivermectin residue concentration was detected at day 4 (mean level = 47.5 ng/mL) and day 5 post treatment (mean level = 26.4 ng/mL) with the injectable form and pour-on treatment, respectively. A second peak in residue concentration was observed using the DELFIA procedure 28 days post-treatment in both treatment groups (23.1 ng/mL injectable and 51.9 ng/mL pour-on). These second peaks were not confirmed by HPLC and must at this time be considered to be false-positive results. By day 35 after treatment the mean ivermectin residue concentration of both groups fell below the limit of detection of the assay. PMID- 11114115 TI - Time-resolved fluorometry in end-point and real-time PCR quantification of nucleic acids. AB - Two time-resolved fluorescence-based methods for nucleic acid quantification are described and their results are compared. Both methods use an exogenous internal standard to eliminate errors arising from different steps of the assay. The first method is a competitive end-point assay, where the standard competes for the same primers with the actual target sequence, prostate-specific antigen (PSA) cDNA. The standard and target are quantified in a dual-label plate hybridization with lanthanide-labelled probes after a fixed number of PCR cycles. The second method is based on real-time monitoring of PCR and on the use of a novel homogeneous signal generation principle that relies on the use of a 5'-->3' exonucleolytic DNA polymerase and a probe labelled with an environment sensitive, stable and fluorescent lanthanide chelate. In this assay, a non-competitive, exogenous internal standard is used. Both assays have a wide linear range (50-5 x 10(6) and 10-5 x 10(7) input PSA cDNA molecules for the end-point and real-time assays, respectively) and there is a strong correlation between the results obtained with the two assays (r = 1.0). Being somewhat faster to perform, the real-time format is better suited for assays that require high throughput. PMID- 11114114 TI - Inhibitors of protein tyrosine phosphorylation: preliminary assessment of activity by time-resolved fluorescence. AB - Epidermal growth factor (EGF) receptor (ErbB1)-associated tyrosine kinase inhibitors may act as potential chemotherapeutic agents. In order to assess the inhibitory activity of these compounds, we developed a simple and sensitive assay based on time-resolved fluorescence. In this technique, crude cell lysates bearing the ErbB1 receptor were captured in microtitre plates immobilized with monoclonal anti-ErbB1 antibody SG 565. Subsequently, the phosphotyrosine content of the cell lysates was quantified by a europium-labelled anti-phosphotyrosine antibody. Thus, genistein, a tyrosine kinase inhibitor, was capable of reducing by half the tyrosine phosphorylation caused by the binding of EGF to A431 cells, whereas 6-carboxymethyl genistein did not inhibit protein tyrosine phosphorylation. This assay is simple to perform, does not use radioactive substrates, and can be useful for screening EGF receptor tyrosine kinase inhibitors from natural products or synthetic compounds. Moreover, the assay has a high signal:noise ratio and is suitable for large-scale screening. PMID- 11114116 TI - Time-resolved fluorescence imaging for quantitative histochemistry using lanthanide chelates in nanoparticles and conjugated to monoclonal antibodies. AB - Tissue and cell examinations have a potential to produce extremely valuable information about antigen quantities in samples. Using currently available methods, a truly quantitative analysis is nearly impossible. We have previously shown that immunohistochemical (IHC) detection of prostate-specific antigen and human glandular kallikrein from prostatic tissue, together with time-resolved fluorescence imaging (TRFI), is a suitable method for obtaining quantitative data from biological samples and that the signal response is linear. In this paper we show that Eu-chelate containing particles in the nanometer range are suitable labels for quantitative IHC. Even single nanoparticle molecules can be detected by TRFI and the signals measured can be readily quantitated. The signal intensity correlates very well with the amount of bound label, and the use of nanoparticles could markedly improve the sensitivity of quantitative IHC methods. TRFI provides a powerful tool for providing quantitative data about antigens or transcripts in tissue sections or cultured cells. It is also of major importance in standardization and optimization of protocols for fixation and tissue preparation, including antigen retrieval methods. PMID- 11114117 TI - Time-resolved fluorometry (TRF)-based immunoassay concept for rapid and quantitative determination of biochemical myocardial infarction markers from whole blood, serum and plasma. AB - We report the development of a time-resolved fluorometry-based immunoassay concept for the rapid measurement of three cardiac markers from whole blood, serum or plasma. Using a universal all-in-one (AIO) dry reagent concept, all the analyte specific reagents are built into a single microtire well, to which an identical assay protocol is applied. Addition of 5-20 microL sample (whole blood, serum or plasma) together with a universal buffer initiates the reaction, which is brought close to equilibrium in 15 min. After the wash step the Eu chelate derived signal is measured directly from the dried surface. Application of this concept to the three cardiac markers illustrates its ability to provide rapid, highly sensitive and fully quantitative results over a large dynamic range with good reproducibility. Such a performance, especially when using whole blood specimens, is largely a consequence of the inherently fluorescent and stable Eu chelate employed in the system. Correlation to commercial assays was excellent for all three analytes, as was between-sample matrix correlation using the AIO assays. The presented assay concept enabling a simple automation is particularly suited for point-of-care applications, where the performance characteristics are fully comparable to state-of-the-art central laboratory immunoassays. PMID- 11114118 TI - Preliminary examination of time-resolved fluorometry for protein array applications. AB - The advantages of time-resolved fluorometry over conventional fluorometric analysis are well known. However, time-resolved fluorescence has not as yet found wide applications in protein microarray or other multiparametric methods of analysis. Here we describe a general method which is suitable for multiparametric and microarray analysis, based on time-resolved fluorometry. A polystyrene surface is coated with different monoclonal antibodies, specific for certain analytes. The analyte mixtures are then universally biotinylated, using an active biotin ester. After removing excess biotin, the biotinylated samples are applied on the polystyrene surface, incubated and the excess is washed away. The bound moieties are then quantified by adding a universal detection reagent containing streptavidin, labelled with a fluorescent europium chelate. After washing and drying of the solid surface, the immobilized moieties are detected by using solid phase, laser-excited time-resolved fluorometric analysis. In a preliminary examination of this principle, we have demonstrated that we can correctly identify upregulation of three secreted proteins, following stimulation of a breast carcinoma cell line with various steroids. Our method should be suitable for high-density microarray analysis of proteins, captured by specific monoclonal antibodies or other binding reagents. PMID- 11114119 TI - Precaution: belief, regulatory system, and overarching principle. AB - The precautionary principle has been incorporated as a belief statement in international agreements for more than a decade. The 1998 Wingspread definition of the principle was the first to bring together four components that, in the past two years, have formed the elements of a broader, overarching approach to precaution that is a robust basis for its specific implementation: prompt action even in the face of scientific uncertainty, burden of proof and persuasion on proponents of potentially hazardous technologies, assessment of alternatives, and transparency. This broad approach to precaution is in direct conflict with the simplistic, easily manipulated principles and methods of risk-assessment-based risk management being exported by U.S. officialdom. In contrast to risk assessment, precaution, broadly defined, incorporates the full range of human intelligence in the task of protecting human health and the environment: flexibility, foresight, fairness, thoughtful consideration, and honesty. PMID- 11114120 TI - Precautionary principle in international law. AB - The deregulatory nature of trade rules frequently brings them into conflict with the precautionary principle. These rules dominate debate over the content and legal status of the precautionary principle at the international level. The World Trade Organization (WTO), because of its power in settling disputes, is a key player. Many States are concerned to define the precautionary principle consistent with WTO rules, which generally means defining it as simply a component of risk analysis. At the same time, many States, especially environmental and public health policymakers, see the principle as the legal basis for preserving domestic and public health measures in the face of deregulatory pressures from the WTO. The precautionary principle has begun to acquire greater content and to move into the operative articles of legally binding international agreements. It is important to continue this trend. PMID- 11114121 TI - Children's environmental health: a case study in implementing the precautionary principle. AB - The plausible threat to children from environmental exposures and uncertainty as to the magnitude and nature of potentially harmful effects provide a rationale for taking precautionary measures to prevent such exposures. The authors present principles for applying precaution to children's environmental health, and policy tools for implementing them. A stronger focus on primary prevention and a better understanding of the risks are needed. PMID- 11114122 TI - An environmentalist's vision of operationalizing the precautionary principle in the management of chemicals. AB - In Europe and elsewhere, there is compelling evidence that humans, wildlife, and the environment are being damaged by man-made chemicals. Recognizing that regulatory initiatives are needed to try to prevent harm before it occurs in the future, the precautionary principle has become a focus of attention on both sides of the Atlantic. The authors suggest a way to implement the precautionary principle in the management of chemicals, outlining how this guiding principle can be given a practical relevance within regulatory initiatives to reduce the risks posed by chemicals currently traded in the European Union and elsewhere. PMID- 11114123 TI - Precautionary approaches to the appraisal of risk: a case study of a genetically modified crop. AB - There are strong scientific reasons for holding the broader scope of precautionary approaches to be more consistent with the scientific foundations of rational choice and probability theory than are conventional narrow risk assessment techniques. The imperatives both of science and precaution can be seen to pull in the same direction. The regulatory appraisal of risk should become more systematic and broader in scope. In particular, a set of criteria can be developed concerning the need for greater humility, completeness, transparency, and participation in regulatory appraisal, with specific attention to the comparison of different options (including mixtures of options), the consideration of benefits and justifications, and the systematic "mapping" of the ways in which different framing assumptions lead to different pictures of performance. A case study of a pilot exercise applying a multi-criteria mapping method to the regulatory appraisal of a genetically modified crop is reported. The results are more complete than orthodox risk assessment, in that they embody consideration of an unlimited array of issues and include consideration of a wide range of different strategic alternatives to the use of GM technologies. It is concluded that conventional regulatory appraisal might be adapted to better address the imperatives of both science and precaution. PMID- 11114124 TI - Risk assessment in a third-world reality: an endosulfan case history. AB - The author describes the chain of frustrated attempts to regulate the use of the toxic pesticide endosulfan in the Philippines in the face of opposition from its internationally powerful manufacturer. Risk assessment, although purportedly a science-based system to protect public health and the environment, has failed to do so, particularly in vulnerable third-world countries. The precautionary principle, based on preventing risk rather than assessing established risk, holds hope for resolution of the problem. PMID- 11114125 TI - Beyond risk: an ecological paradigm to prevent global chemical pollution. AB - Since World War II, synthetic chemical pollutants have accumulated in the environment and food webs on a global basis, have damaged wildlife populations, and may pose large-scale hazards to human health. Despite the global nature of this problem, the vast majority of environmental regulations focus on preventing local risks using risk assessment of individual compounds, discharge permits, and control and disposal technologies. The current approach has failed to prevent global contamination and environmental damage because it underestimates the scale, complexity, and diversity of the hazards of chemical pollution. Fundamental shifts in the mode of chemical assessment and policy are required; a new framework should focus on chemical classes rather than individual substances, convert industrial processes to prevent the production and use of persistent and/or bioaccumulative substances, and shift the default state of pollution policy in the face of uncertainty from permission to restriction. PMID- 11114126 TI - The international trade in toxic waste: the case of Sihanoukville, Cambodia. AB - In December 1998, 2,700 metric tons of industrial waste containing high levels of mercury and other metals and possibly other toxic compounds were shipped illegally from Taiwan to Sihanoukville, Cambodia. There the waste was unloaded and transferred to a nearby inland dumpsite. An estimated 2,000 Sihanoukville residents were exposed to the waste occupationally or environmentally, and at least six deaths and hundreds of injuries have been associated with the incident. The authors describe the human exposures and associated morbidity and mortality, recount the medical and public health response, and consider the issues complicating epidemiologic analysis of the incident. They also consider the major issues highlighted by the incident, including the trade in toxic waste between developed and less developed countries, the factors that shape emergency public health responses in resource-poor environments, and the importance of prevention and preparedness in containing emergencies in developing countries. PMID- 11114127 TI - Oupatient programs of myeloablative chemotherapy, autologous and allogeneic bone marrow transplantation. PMID- 11114128 TI - Educational Book for the Fourth Congress of the European Haematology Association, held in Barcelona, Spain, on June 9-12, 1999. PMID- 11114129 TI - Cytokine gene expression and T-cell proliferative responses in lymph node mononuclear cells from children with early stage human immunodeficiency virus infection. AB - BACKGROUND AND OBJECTIVES: The immunologic events taking place in secondary lymphoid tissue from children with early stage human immunodeficiency virus (HIV) infection are poorly understood. The aim of this study was to investigate cytokine gene expression and proliferative responses in lymph node (LN) biopsies from five children with early stage HIV infection, in the context of LN morphology and viral load. DESIGN AND METHODS: The design of the study was approved by the local Ethical Committee. Cytokine gene expression was studied in LN biopsies and in paired peripheral blood (PB) samples from HIV-infected children by reverse transcriptase-polymerase chain reaction. T-cell proliferation was assessed by 3H-thymidine incorporation. Viral burden in germinal centers was assessed by video densitometric analysis following immunohistochemical staining for HIV p24. RESULTS: Interleukin (IL)-2, IL-4 and IL-5 mRNA were not detected in any LN or PB sample from HIV-infected children. Interferon (IFN)-gamma mRNA was found only in CD8+ cells. IL-12 p35, IL-10, transforming growth factor-(TGF) beta1, regulated on activation normal T-cell expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, MIP-1beta and IL-16 transcripts were detected in all samples. Proliferation of LN and PB mononuclear cells to polyclonal mitogens and soluble (recall and HIV-related) antigens was impaired as compared with the responses in a group of age-matched healthy controls. INTERPRETATION AND CONCLUSIONS: Changes in cytokine gene expression and T-cell proliferative responses are already detectable in lymph nodes from HIV-infected children at an early stage of disease. PMID- 11114130 TI - Rapid quantitative detection of BCR-ABL transcripts in chronic myeloid leukemia patients by real-time reverse transcriptase polymerase-chain reaction using fluorescently labeled probes. AB - BACKGROUND AND OBJECTIVES: The limited value of qualitative reverse transcription polymerase chain-reaction (RT-PCR) for monitoring chronic myeloid leukemia (CML) patients has prompted the development of quantitative assays. We have developed a quantitative real-time PCR (QC-PCR) method in the LightCycler, based on the use of fluorescently labeled probes (HybProbes), to estimate BCR-ABL fusion gene transcripts in samples from CML patients. DESIGN AND METHODS: Fifty-two samples (45 peripheral blood, five bone marrow, and two apheresis product samples) from nine patients with CML were analyzed. Seven patients were studied at diagnosis and during follow-up after hematopoietic stem cell transplantation (HSCT), whereas two were evaluated only after HSCT. The PCR reaction was carried out in capillary tubes in a final volume of 10 microL, using 2 microL cDNA, the Mensik et al. primers, and two HybProbes. The results for BCR-ABL were normalized with reference to ABL. The PCR program is completed in only 45 min. RESULTS: The sensitivity attained allowed the detection of rearrangements at dilutions of between 5-10(-4) and 10(-5) K562 cDNA. The within-assay coefficient of variation was 11% for BCR-ABL, and 9% for ABL. A greater than 2 log reduction in the BCR ABL/ABL ratio was evident shortly after transplantation in all allografted patients. INTERPRETATION AND CONCLUSIONS: We may conclude that the TaqMan probe technology can be easily adapted to HybProbes with equivalent results. Besides, the results of BCR-ABL quantification in the follow-up of patients clearly confirm that real-time PCR with HybProbes is a reliable and sensitive method for monitoring minimal residual leukemia after HSCT in CML patients. PMID- 11114131 TI - Incidence and outcome of pneumonia in patients with acute leukemia receiving first induction therapy with anthracycline-containing regimens. AB - BACKGROUND AND OBJECTIVES: Even though the risk of pneumonia is higher in patients with advanced disease, the potential risk of death is particularly relevant during induction therapy, when patients can be potentially cured of their hematologic disease: our study was aimed at evaluating the risk and outcome of pneumonia in these patients. DESIGN AND METHODS: We retrospectively studied all 458 patients affected by acute leukemia receiving an anthracycline-containing induction regimen in the years 1984-1989. RESULTS: Of the 458 patients, 109 (23.8%) developed pneumonia: 91 had acute myelogenous leukemia (AML) and 18 had acute lymphoblastic leukemia (ALL). At univariate analysis, advanced age, AML and total blast count significantly correlated with the risk of pneumonia. At multivariate analysis, only age (p< 0.0001) and total blast count (p=0.002) retained their prognostic significance. Pneumonia responded to treatment in 67 (61.5%) patients, while 42 (38.5%) patients died. Among patients with pneumonia, 51 (46.8%) patients achieved a complete remission: 9/18 ALL and 42/91 AML. At univariate analysis, the most significant determinant of a positive outcome was the achievement of complete remission; a higher absolute neutrophil count at the onset of pneumonia, the absence of rales, a single infiltrate and the absence of microbiological demonstration of infection were also related to a positive outcome. At multivariate analysis, the achievement of complete remission and, with borderline significance, a single infiltrate maintained their prognostic value. INTERPRETATION AND CONCLUSIONS: Pneumonia remains one of the most relevant risks of morbidity and mortality during induction therapy for acute leukemia. A fatal outcome is associated, in most cases, with a failure to achieve remission of leukemia. PMID- 11114132 TI - The relevance of multidrug resistance-associated P-glycoprotein expression in the treatment response of B-cell chronic lymphocytic leukemia. AB - BACKGROUND AND OBJECTIVES: The aim of this study was to determine whether expression of P-glycoprotein (Pgp) is an intrinsic feature of B-lymphocytes in B cell chronic lymphocytic leukemia (B-CLL) and how it correlates with hematologic indices and tumor load in the disease. Furthermore, the change of Pgp expression under cytotoxic treatment and its correlation to treatment outcome were studied. DESIGN AND METHODS: In 42 B-CLL patients, of whom 13 were sequentially monitored, expression of extracellular (MRK-16) and intracellular (C-219) Pgp epitopes on peripheral blood lymphocytes was determined by flow cytometry analysis and quantified by ratio of the mean fluorescence (RMF) in flow cytometry analysis. RESULTS: Median RMF values in B-CLL patients were higher than in age-matched controls. Pgp expression did not correlate with any of the hematologic features or clinical stage of the disease. Patients who received some type of cytoreductive treatment prior to the study had lower Pgp values for both measured epitopes (median RMF for C-219 and MRK-16 of 1.99 and 2.03 in comparison to those of non-treated patients: 3.11 and 2.88, respectively). In 13 patients monitored during treatment the decrease in RMF was noted after treatment with chlorambucil, with RMF values for both Pgp epitopes decreasing in responders. This was in contrast to unchanged or even increased RMF values in those patients who did not respond to therapy. INTERPRETATION AND CONCLUSIONS: Our study confirms the importance of quantitative evaluation of Pgp expression by flow cytometry. At the clinical level, cross-sectional, single test evaluation of Pgp is of limited value whereas sequential follow-up values correlate with treatment response. PMID- 11114133 TI - Low-dose fludarabine and cyclophosphamide in elderly patients with B-cell chronic lymphocytic leukemia refractory to conventional therapy. AB - BACKGROUND AND OBJECTIVES: In recent years fludarabine alone or in combination with other drugs has been reported to be effective in the treatment of B-cell chronic lymphocytic leukemia (B-CLL), both as first line and salvage therapy. Among the different combination regimens, the association of fludarabine and cyclophosphamide has shown a considerable therapeutic efficacy, although a relevant number of infectious complications have been described, particularly in elderly patients. The aim of this work was to evaluate the efficacy, the toxicity, and the incidence of infectious episodes of a regimen combining lower doses of fludarabine and cyclophosphamide in elderly patients with B-CLL refractory to conventional therapy. DESIGN AND METHODS: Twenty patients with progressive B-CLL with a median age of 75 years (4 in stage B and 16 in stage C) and refractory to conventional therapy were enrolled in this study. The combination regimen was as follows: fludarabine 15 mg/m2/day i.v. [max 25 mg] and cyclophosphamide 200 mg/m2/day i.v. for four days. RESULTS: All patients enrolled were evaluable for response. Three out of 20 (15%) patients achieved a complete remission (CR), 14/20 (70%) a partial response (PR) with an overall response rate (CR+PR) of 85%, according to National Cancer Institute-Working Group response criteria. Three patients were considered resistant. In four out of 20 patients (20%), a severe neutropenia (neutrophils < 0.5x10(9)/L) occurred and one of them developed an infectious complication which required treatment with systemic antibiotics and granulocyte colony- stimulating factor (G-CSF). Non-hematologic toxicity was negligible in all patients but one, who despite a adequate therapy with allopurinol and hydration, experienced a tumor lysis syndrome with transient but severe renal impairment. INTERPRETATION AND CONCLUSIONS: The association of low-dose fludarabine and cyclophosphamide appeared to be effective in this subset of B-CLL patients, reproducing a similar overall response rate obtained with other fludarabine-based combination therapies. In addition, in this group of elderly patients, toxic side effects were negligible and infectious complications remarkably low. PMID- 11114134 TI - Risk of venous thrombosis in carriers of the prothrombin G20210A variant and factor V Leiden and their interaction with oral contraceptives. AB - BACKGROUND AND OBJECTIVES: The prothrombin G20210A mutation and factor V Leiden have been found to be associated with an increased risk of venous thrombosis, but the reported prevalences of the prothrombin gene variant both in the normal population and in patients with deep venous thrombosis (DVT) vary greatly in the literature. Moreover, the influence of oral contraceptives (OC) on thrombotic events in patients with the prothrombin G20210A variant has not been well established. In this study we evaluate both circumstances. DESIGN AND METHODS: A case-control study was run on 229 patients with DVT and 246 healthy controls. The patients' history of thrombosis and acquired thrombotic risk factors, especially OC, were recorded. Prothrombin G20210A mutation, factor V Leiden, antithrombin, heparin II cofactor, plasminogen and proteins C and S were evaluated. RESULTS: Seven and a half percent of the patients and 2.9% of the controls were carriers of the prothrombin mutation, while 12.2% of the patients and 1.6% of the controls had factor V Leiden. Among the 229 DVT patients there were 130 patients with clinically suspected thrombophilia (first thrombotic event occurring before the age of 45 years or positive family history of thrombosis or recurrent venous thrombosis). Ten percent of these 130 patients were carriers of the prothrombin G20210A mutation and 18.5% had the factor V Leiden mutation. The odds ratios (OR) for DVT risk were: 2.4 (95% CI, 1.0-6.3) for the total DVT patients and 5.2 (95% CI, 1.4-19.5) for the patients with clinically suspected thrombophilia with the prothrombin mutation. The risk of thrombosis was 6.9 (95% CI, 2.3-20.6) for the DVT patients and 14.3 (95% CI, 3.3-64.6) for the patients with clinically suspected thrombophilia with factor V Leiden. Fifty-five percent of the patients with combined congenital defects (prothrombin mutation G20210A plus another congenital defect) had recurrent thrombosis. In women receiving OC the risk of DVT was 3.5 (95% CI, 1.5-8.2) that of the patients not receiving OC. When women with combined defects were also taking OC, the risk of thrombosis increased significantly. INTERPRETATION AND CONCLUSIONS: The prevalence of the prothrombin G20210A mutation in the healthy population in our study is similar to that observed in other southern European countries. The prothrombin G20210A mutation does not by itself seem to be a high thrombotic risk factor. However, when it is present together with other thrombotic risk factors, the predicted risk of thrombotic events increases. The use of OC by women with the prothrombin G20210A variant or FV Leiden, either alone or combined with other thrombotic risk factors, was associated with a significant increase in the risk of venous thrombosis. PMID- 11114135 TI - The effect of two different doses of aprotinin on hemostasis in cardiopulmonary bypass surgery: similar transfusion requirements and blood loss. AB - BACKGROUND AND OBJECTIVES: Various dosages of aprotinin have proven to be effective in reducing blood loss and allogeneic transfusion requirements in cardiopulmonary bypass surgery, despite the controversy surrounding the precise hemostatic mechanisms of this drug. The aim of our prospective, randomized, double-blind study was to assess differences in blood loss and transfusion requirements and the effect of two dosages of aprotinin on hemostatic activation. DESIGN AND METHODS: Patients undergoing coronary artery bypass grafting received high-dose aprotinin (n=28), pump-prime-only (PPO) aprotinin (n=28), or placebo (n=28). RESULTS: The high-dose and the PPO groups had a significantly lower blood loss (985 mL [95%CI 845-1,102] and 1,255 mL, [95% CI 1,025-1,406], respectively) than the placebo group (1,416 mL, 95%CI 1,248-1,612]. Transfusion requirements were lower in the aprotinin-treated groups than in the patients receiving placebo (21 units and 15 units in the high and low-dose groups vs 59 units in placebo group). As far as concerns parameters of thrombin generation, the aprotinin groups showed a significant reduction of F1+2 prothrombin fragment but not of thrombin-antithrombin complexes. There were higher levels of natural anticoagulants, i.e. antithrombin, protein C and protein S, in the high-dose aprotinin group. As regards fibrinolysis parameters, D-dimer was lower in the aprotinin groups, and the levels of alpha2-antiplasmin and plasmin-alpha2 antiplasmin complexes were raised. In summary, both dosages of aprotinin were equally effective in reducing blood loss and transfusion requirements. There was a lower activation of coagulation and fibrinolysis in cardiopulmonary bypass patients treated with aprotinin: the levels of natural anticoagulants were less decreased in the high-dose group. No differences in thrombotic complications were observed between aprotinin groups. INTERPRETATION AND CONCLUSIONS: Our study shows that both dosages of aprotinin are safe and effective in reducing transfusion requirements. Considering the difference in cost of using a low-dose or high-dose schedule, the former should be recommended for patients undergoing cardiopulmonary bypass surgery. PMID- 11114136 TI - Bone marrow aspirate on the 14th day of induction treatment as a prognostic tool in de novo adult acute myeloid leukemia. AB - BACKGROUND AND OBJECTIVES: In adult acute myeloid leukemia (AML), a variety of clinical and biological parameters have been examined for their potential value in predicting treatment response. Early response to induction therapy could be an important prognostic factor in this disease. DESIGN AND METHODS: We studied the relationship between reduced blasts in bone marrow aspirate on the 14th day (BMA14th) of induction chemotherapy and treatment outcome in 198 adult AML patients of whom 124 were < 60 years old (group A) and 74 > or = 60 years old (group B). Receiver operating characteristic curve analysis was used to assess the prognostic performance of BMA14th. Using the percentages of blasts of < or = 22% and < or = 15% as criteria for predicting treatment outcome gave the highest accuracy in terms of sensitivity and specificity in groups A and B, respectively. RESULTS: In group A, of 97 patients with a BMA14th < or = 22%, 77 (79%) achieved complete remission (CR), whereas of 27 patients with a BMA14th > 22%, 22 (81%) were non-responders (NR) (p < 0.0001). The test sensitivity and specificity were 93.9% and 71.4%, respectively. In group B, of 27 patients with a BMA14th < or = 15%, 18 (67%) achieved CR, whereas of 47 patients with a BMA14th >15%, 38 (81%) were NR (p = 0.0001). The test sensitivity and specificity were 66.7% and 80.9%, respectively. INTERPRETATION AND CONCLUSIONS: Our data suggest that BMA14th may be a predictive test for CR, helping to identify NR patients early in their disease. Further studies are needed to establish the practical implications of the results of our study. PMID- 11114137 TI - The pathologist's view point. Part I--indolent lymphomas. AB - BACKGROUND AND OBJECTIVES: The REAL/WHO classification constitutes a new tool for the better understanding and treatment of malignant lymphomas. The authors focus on the key features of B-cell lymphomas with an indolent behavior, aiming to contribute to the cross-talk between pathologists and clinicians. DATA SOURCES AND METHODS: Each lymphoma entity is analyzed on the basis of the most representative contributions in the literature and the authors' experience gained in studying more than 20,000 lymphoid tumors over a 20-year period. RESULTS: Guidelines for diagnosis and areas of interest for future clinico-pathologic studies are identified and discussed. Within this context, selected data obtained by the application of novel markers are presented. INTERPRETATION AND CONCLUSIONS: The present know- ledge and organization of malignant lymphomas now make the development of tailored therapies a feasible goal. PMID- 11114138 TI - The pathologist's view point. Part II --aggressive lymphomas. AB - BACKGROUND AND OBJECTIVES: The REAL/WHO classification constitutes a new tool for the better understanding and treatment of malignant lymphomas. The authors focus on the key features of aggressive B- and T-cell lymphomas, aiming to contribute to the cross-talk between pathologists and clinicians. DATA SOURCES AND METHODS: Each lymphoma entity is analyzed on the basis of the most representative contributions in the literature and the authors' experience gained in studying more than 20,000 lymphoid tumors over a 20-year period. RESULTS: Guidelines for diagnosis and areas of interest for future clinico-pathologic studies are identified and discussed. Within this context, selected data obtained by the application of novel markers are presented. INTERPRETATION AND CONCLUSIONS: The present know- ledge and organization of malignant lymphomas now make the development of tailored therapies a feasible goal. PMID- 11114139 TI - The irreplaceable image: Platelet satellitism to granulated lymphocytes. PMID- 11114140 TI - The irreplaceable image: Unique intranuclear inclusion in an IgG kappa multiple myeloma. PMID- 11114141 TI - The S65C mutation in Spain. Implications for iron overload screening. AB - Hereditary hemochromatosis is related to mutations of the HFE gene. The role of the S65C mutation of this gene was evaluated in a Spanish population, consisting of 100 controls and 41 patients who had resulted positive to screening for iron overload. Only one patient was heterozygous for the S65C mutation, so the S65C mutation is infrequent in our area. Nevertheless, it is advisable to search for this mutation in cases with iron overload and heterozygosity for the C282Y or H63D mutations of the HFE gene. PMID- 11114142 TI - Differences in phenotype, growth factor requirements, pattern of expression of adhesion molecules and rate of apoptosis displayed by three new myeloid sister leukemic cell lines. AB - We established three new human myeloid cell lines from one patient, in the presence of granulocyte-macrophage colony-stimulating factor (UPM1-GM), interleukin-3 (UMP1-IL-3) or without exogenous growth factors (UPM1). The 3 lines were characterized by phenotypic, genotypic and functional studies. These cell lines may provide useful tools to study different aspects of leukemic cell biology PMID- 11114143 TI - Acute lymphoblastic leukemia in the elderly. A twelve-year retrospective, single center study. AB - Acute myeloid leukemia in elderly patients is a well-studied disease, while only a few studies on acute lymphoid leukemia (ALL) in elderly patients have been reported and their results are not encouraging. The aims of the present study were to review the characteristics of acute lymphoblastic leukemia developing in patients aged over 65 years old during a twelve-year period at our Institution and to analyze the clinical and laboratory characteristics. PMID- 11114144 TI - Unsuccessful treatment of resistant thrombotic thrombocytopenic purpura with prostacyclin. AB - Prostacyclin has been suggested as a useful agent for patients with thromobotic thrombocytopenic pupura (TTP) refractory to plasma-exchange. We report our unsuccessful experience with iloprost in a patient with TTP resistant to plasma exghange, vincristine and high dose immunoglobulins. PMID- 11114146 TI - Odor and odorant: a terminological clarification. PMID- 11114145 TI - Percutaneous removal of a knotted pulmonary artery catheter using a tracheostomy dilator. AB - STATEMENT OF FINDINGS: This case report describes removal of a knotted, subclavian, pulmonary artery catheter using a tracheostomy dilator. With this simple method an invasive procedure might be averted. PMID- 11114147 TI - EOG responses in anesthetized freely breathing rats. AB - In mammals, access of odor molecules to the olfactory receptor neurons is controlled by respiratory activity. Thus, anesthetized, freely breathing rats were used to record from the olfactory mucosa in the intact nasal cavity (electroolfactogram or EOG) so as to study global response characteristics to odor stimuli. During alternation of the inspiratory phases of odor sampling and expiratory phases, the response was a succession of individual EOG events synchronized with respiration. These were characterized by a steep decrease that started approximately 100-150 ms after the beginning of inhalation, reached its maximum at the transition between inspiration and expiration and was followed by a slower rise until the next inhalation. They were greater during the first respiratory cycles following odor stimulation onset. Thereafter their amplitudes decreased throughout odor delivery, but a significant EOG signal was still present at the end of short (10 s) and long (60 s) odor presentations. Amplitude increased with odor concentration, but much less than expected from concentration changes. Lastly, for some odors EOG responses persisted well beyond the end of stimulation. These results are in agreement with the respiratory synchronization of mitral cell activities observed during short odor presentations and long duration odor exposures. They underline again the importance of taking into account the respiratory activity in studies on the functioning of the olfactory system. PMID- 11114148 TI - Evidence for separate perceptive and semantic memories for odours: a priming experiment. AB - Sixty-four subjects participated in an olfactory priming experiment comprising separate study and test phases. Priming was measured within the olfactory modality (intramodal condition) and from the visual modality to the olfactory modality (intermodal condition). In the study phase of the intramodal condition, subjects were exposed twice to a series of odours: once performing a semantic orientation task (deciding which of seven categories odour stimuli belonged to) and once performing a perceptual orientation task (judging the intensity, the hedonicity and the familiarity of odour stimuli). Half of the odour stimuli corresponded to edible products, the other half did not. The study phase of the intermodal condition was similar, with the exception that the names of the odours (instead of the odours themselves) were presented. In the test phase, subjects were presented with primed and non-primed odour targets and had to decide as fast as possible whether the target corresponded to an edible product or not. Response times and types were recorded by a computer. The analysis of response times revealed a priming effect in the intramodal condition only. Results are discussed in terms of separate perceptual and semantic subsystems that store odour representations. PMID- 11114149 TI - Autonomic nervous system responses associated with primary tastes. AB - The hedonic dimension of the taste sensation plays a crucial role in the control of many taste-mediated responses related to food ingestion or rejection. The purpose of this study was to evaluate the emotional reactivity associated with each primary taste (sweet, salty, sour and bitter) through analysis of the variations of autonomic nervous system (ANS) parameters. Thirty-four healthy non smoker volunteer subjects (17 males and 17 females, mean age = 28 years) participated in the experiment. Taste stimuli were solutions of 0.3 M sucrose (sweet), 0.15 M NaCl (salty), 0.02 M citric acid (sour) and 0.00015 M quinine sulfate (bitter). Evian mineral water was used as the diluent and control (neutral taste). Throughout the test, five ANS parameters (skin potential and skin resistance, skin blood flow and skin temperature, and instantaneous heart rate) were simultaneously and continuously recorded. Results of the ANOVA evidenced a significant effect of primary taste on skin resistance amplitude (P: < 0.001) and duration (P: < 0.0001), skin temperature amplitude (P: < 0.001), skin blood flow amplitude (vasoconstriction) (P: < 0.0001) and instantaneous heart rate increase (P: < 0.0001). Skin resistance and cardiac responses were the most relevant ANS parameters to distinguish among the taste solutions. The four primary tastes could be associated with significantly different ANS responses in relation to their hedonic valence: the pleasantly connoted and innate-accepted sweet taste induced the weakest ANS responses whereas the unpleasant connoted tastes (salty, sour and bitter) induced stronger ANS responses, the innate rejected bitter taste inducing the strongest ones. Such a neurovegetative characterization of each primary taste could provide references for the hedonic analysis of the more complex gustative sensation attached to foods. PMID- 11114150 TI - Lack of functional and morphological susceptibility of the greater superficial petrosal nerve to developmental dietary sodium restriction. AB - Restriction of dietary sodium during gestation has major effects on taste function and anatomy in the offspring. The chorda tympani nerve of offspring that are maintained on sodium-reduced chow throughout life (NaDep) has reduced neurophysiological responses to sodium and altered morphology of its terminal field in the nucleus of the solitary tract. There are many anatomical and physiological similarities between the chorda tympani nerve that innervates taste buds on the anterior tongue and the greater superficial petrosal nerve (GSP) that innervates taste buds on the palate. To determine if the GSP is similarly susceptible to the effects of dietary sodium restriction, the present study examined neurophysiological responses and the terminal field of the GSP in NaDep and control rats. Neurophysiological responses of the GSP to a variety of sodium and non-sodium stimuli did not differ between NaDep and control rats. Furthermore, the volume and shape of the GSP terminal field in the nucleus of the solitary tract did not differ between the groups. Therefore, despite the high degree of functional and anatomical correspondence between the chorda tympani nerve and the GSP, the GSP does not appear to be susceptible to the effects of lifelong dietary sodium restriction. PMID- 11114151 TI - Human foetuses learn odours from their pregnant mother's diet. AB - Olfactory responsiveness was assessed in 24 neonates born to mothers who had or had not consumed anise flavour during pregnancy. Both groups of infants were followed-up for behavioural markers of attraction and aversion when exposed to anise odour and a control odour immediately after birth and on day 4. Infants born to anise-consuming mothers evinced a stable preference for anise odour over this period, whereas those born to anise non-consuming mothers displayed aversion or neutral responses. This study provides the first clear evidence that through their diet human mothers influence the hedonic polarity of their neonates' initial olfactory responses. The findings have potential implications for the early mother-to-infant transmission of chemosensory information relative to food and addictive products. PMID- 11114152 TI - Diazepam-binding inhibitor-like activity in rat cerebrospinal fluid after stimulation by an aversive quinine taste. AB - Cerebrospinal fluid (CSF) taken from rats after stimulation by an aversive quinine taste (hereafter called quinine CSF) administered into the fourth ventricle of mice suppressed their intake of 5% sucrose solution. We examined the effects of CSF on glutathione-induced tentacle ball formation (TBF) of hydra to determine the change in CSF components associated with aversive taste stimuli. The suppressive activity of quinine CSF on TBF in the presence of 3 microM S: methyl-glutathione (GSM) was markedly lower than that of CSF obtained from control rats (control CSF). Pronase-treated quinine CSF had suppressive activity similar to that of control CSF. The active principle passed through an ultrafiltration membrane, with a molecular weight cut-off of 30 kDa, but not through one with a cut-off of 3 kDa. A peptide fragment of diazepam-binding inhibitor (DBI) nullified the suppression of TBF at 3 microM GSM by control CSF. The nullifying activity of quinine CSF was not observed after treatment with a benzodiazepine receptor preparation that was able to bind DBI. When flumazenil, a benzodiazepine receptor antagonist, was given to mice, the suppression of the intake of 5% sucrose solution by quinine CSF was partially reversed. It is suggested that quinine CSF contains a DBI-like substance. PMID- 11114153 TI - Chemosensory context effects: role of perceived similarity and neural commonality. AB - Seven experiments investigated how stimulus context affects judgements of the magnitude of chemosensory stimuli. In each experiment, subjects gave magnitude estimates of the intensity of several concentrations of two substances, with the contextual set of concentrations varying across experimental conditions. Different experiments used different pairs of substances, which could be tastants (sucrose or NaCl) that were sipped, odorants (orange or vanillin) that were sipped (i.e. presented retronasally) or the same odorants sniffed (i.e. presented orthonasally). Varying the stimulus context affected the judgements differentially when the two substances were compositionally different (sucrose and NaCl; sucrose and orange; sucrose and vanillin) but not when they were the same (vanillin or orange presented orally and nasally). Judgements of qualitative similarity of the same pairs of substances, obtained in a separate experiment, failed to predict accurately the pattern of differential context effects. Taken together, the results suggest that differential effects of context relate only indirectly to perceptual dissimilarity per se but may primarily reflect the result of stimulus-specific adaptation-like processes. PMID- 11114154 TI - Effects of air flow on rat electroolfactogram. AB - The electroolfactogram (EOG) previously has been used to demonstrate the regional distribution of rat olfactory epithelial odorant responses. Here, we evaluated the effects of airflow parameters on EOGs in two preparations: one where odorants were directly applied to the epithelium (opened preparation) and one where odorants were drawn through the nasal passages by an artificial sniff (closed preparation). EOG rise times served as one measure of odorant access. For isoamyl acetate (but not for limonene), rise times were slower in the lateral recesses of the closed (but not the opened) preparation. Polar odorants (amyl acetate, carvone and benzaldehyde) evoked smaller responses in the closed preparation than in the opened preparation, and these responses were particularly depressed in the lateral regions of the closed preparation. Responses to nonpolar hydrocarbon odorants (limonene and benzene) were equal in the lateral regions of both preparations, but were somewhat depressed in the medial region of the closed preparation. The responses to some polar odorants in the closed preparation were sensitive to changes in airflow parameters. These data suggest that the sorptive properties of the nose contribute substantially to determining the response of the epithelium and act to increase differences produced by inherent receptor mechanisms. PMID- 11114156 TI - Chemosensory bioresponses in man II PMID- 11114155 TI - Sensory properties of citric acid: psychophysical evidence for sensitization, self-desensitization, cross-desensitization and cross-stimulus-induced recovery following capsaicin. AB - In a first experiment, human subjects used a bipolar scale to rate the irritant sensation elicited by 10 sequentially repeated applications of either 3 ppm capsaicin or 250 mM citric acid on one side of the dorsal surface of the tongue, at 1 min intervals (30 s inter-stimulus interval). Citric acid-evoked irritation significantly increased across trials, consistent with sensitization. With capsaicin there was a large degree of inter- and intra-individual variation in successive ratings with no overall sensitization. Following the sequential stimulation series and a 10 min rest period, self- and cross-desensitization effects were tested in a two-alternative forced choice (2-AFC) paradigm by placing either citric acid or capsaicin on both sides of the tongue and asking subjects to indicate which side of the tongue yielded a stronger irritant sensation. Subjects also gave separate intensity ratings for irritation on each side of the tongue. Capsaicin self-desensitization was confirmed, while cross desensitization to citric acid was not observed. In addition, citric acid self desensitization and cross-desensitization to capsaicin were observed. In a second experiment a stronger capsaicin solution (33 ppm) was applied to one side of the tongue using cotton swabs. After the burning sensation elicited by capsaicin had disappeared, citric acid was applied bilaterally and cross-desensitization was observed using the same 2-AFC and rating procedures. This was followed by repeated re-application of citric acid at 1 min intervals to the capsaicin treated side. The irritant sensation elicited by citric acid increased significantly, indicating a 'cross-stimulus-induced recovery' from capsaicin desensitization. In a final experiment we investigated the effect of the sodium channel blocker amiloride on the perceived irritation elicited by citric acid or capsaicin. Following application of amiloride to one side of the tongue with cotton swabs, either citric acid or capsaicin was applied bilaterally and subjects asked to perform a 2-AFC and intensity ratings. Amiloride significantly, albeit weakly, reduced the irritation elicited by citric acid while it weakly but significantly enhanced capsaicin-evoked irritation. These findings are discussed in terms of involvement of vanilloid and acid-sensitive ion channels in acid evoked irritation and pain. PMID- 11114157 TI - Direct gating by retinoic acid of retinal electrical synapses. AB - Retinoic acid (RA), a signaling molecule derived from vitamin A, controls growth and differentiation of a variety of cell types through regulation of gene transcription. In the vertebrate retina, RA also regulates gap junction-mediated physiological coupling of retinal neurons through a nontranscriptional mechanism. Here we report that RA rapidly and specifically modulates synaptic transmission at electrical synapses of cultured retinal horizontal cells through an external RAR(beta)(/gamma)-like binding site, the action of which is independent of second messenger cascades. External application of all-trans retinoic acid (at-RA) reversibly reduced the amplitude of gap junctional conductance in a dose dependent manner, but failed to affect non-gap-junctional channels, including glutamate receptors. In contrast, internal dialysis with at-RA was ineffective, indicating an external site of action. Selective RAR(beta)(/gamma) ligands, but not an RAR(alpha)-selective agonist, mimicked the action of at-RA, suggesting that gating of gap junctional channels is mediated through an RAR(beta)(/gamma) like binding site. At-RA did not act on gap junctional conductance by lowering [pH](i) or by increasing [Ca(2+)](i). A G protein inhibitor and protein kinase inhibitors did not block at-RA uncoupling effects indicating no second messenger systems were involved. Direct action of at-RA on gap junction channels was further supported by its equivalent action on whole-cell hemi-gap-junctional currents and on cell-free excised patch hemichannel currents. At-RA significantly reduced single-channel open probability but did not change unitary conductance. Overall, the results indicate that RA modulates horizontal cell electrical synapses by activation of novel nonnuclear RAR(beta)(/gamma)-like sites either directly on, or intimately associated with, gap junction channels. PMID- 11114158 TI - Acetylation of adenovirus E1A regulates binding of the transcriptional corepressor CtBP. AB - Adenovirus E1A mediates its effects on cellular transformation and transcription by interacting with critical cellular proteins involved in cell growth and differentiation. The amino terminus of E1A binds to CBP/p300 and associated histone acetyltransferases such as P/CAF. The carboxyl terminus binds to the carboxyl-terminal binding protein (CtBP), which associates with histone deacetylases. We show that 12S E1A can be acetylated by p300 and P/CAF and map one of the acetylation sites to Lys-239. This Lys residue is adjacent to the consensus CtBP binding motif, PXDLS. Mutation of Lys-239 to Gln or Ala blocks CtBP binding in vitro and disrupts the E1A-CtBP interaction in vivo. Peptide competition assays demonstrated that the interaction of E1A with CtBP is also blocked by Lys-239 acetylation. Supporting a functional role for Lys-239 in CtBP binding, mutation of this residue to Ala decreases the ability of E1A to block cAMP-regulated enhancer (CRE)-binding protein (CREB)-stimulated gene expression. Finally, we demonstrate that Lys-239 is acetylated in cells by using an antibody directed against an acetyl-Lys-239 E1A peptide. CtBP interacts with a wide variety of other transcriptional repressors through the PXDLS motif, and, in many instances, this motif is followed by a Lys residue. We suggest that acetylation of this residue by histone acetyltransferases, and the consequent disruption of repressor complexes, might be a general mechanism for gene activation. PMID- 11114159 TI - Straightening of bulged RNA by the double-stranded RNA-binding domain from the protein kinase PKR. AB - The human interferon-induced protein kinase, PKR, is an antiviral agent that is activated by long stretches of double-stranded (ds)RNA. PKR has an N-terminal dsRNA-binding domain that contains two tandem copies of the dsRNA-binding motif and interacts with dsRNA in a nonsequence-specific fashion. Surprisingly, PKR can be regulated by certain viral and cellular RNAs containing non-Watson-Crick features. We found that RNAs containing bulges in the middle of a helix can bind to p20, a C-terminal truncated PKR containing the dsRNA-binding domain. Bulges are known to change the global geometry of RNA by bending the helical axis; therefore, we investigated the conformational changes of bulged RNA caused by PKR binding. A 66-mer DNA-RNA(+/- A(3) bulge)-DNA chimera was constructed and annealed to a complementary RNA strand. This duplex forces the protein to bind in the middle. A 66-mer duplex with a top strand composed of DNA-DNA(+/-A(3) bulge) RNA was used as a control. Gel mobility-shift changes among the RNA-protein complexes are consistent with straightening of bulged RNA on protein binding. In addition, a van't Hoff analysis of p20 binding to bulged RNA reveals a favorable DeltaDeltaH degrees and an unfavorable DeltaDeltaS degrees relative to binding to straight dsRNA. These thermodynamic parameters are in good agreement with predictions from a nearest-neighbor analysis for RNA straightening and support a model in which the helical junction flanking the bulge stacks on protein binding. The ability of dsRNA-binding motif proteins to recognize and straighten bent RNA has implications for modulating the topology of RNAs in vivo. PMID- 11114160 TI - Negative regulation of defense responses in plants by a conserved MAPKK kinase. AB - The enhanced disease resistance 1 (edr1) mutation of Arabidopsis confers resistance to powdery mildew disease caused by the fungus Erysiphe cichoracearum. Resistance mediated by the edr1 mutation is correlated with induction of several defense responses, including host cell death. Double mutant analysis revealed that all edr1-associated phenotypes are suppressed by mutations that block salicylic acid (SA) perception (nim1) or reduce SA production (pad4 and eds1). The NahG transgene, which lowers endogenous SA levels, also suppressed edr1. In contrast, the ein2 mutation did not suppress edr1-mediated resistance and associated phenotypes, indicating that ethylene and jasmonic acid-induced responses are not required for edr1 resistance. The EDR1 gene was isolated by positional cloning and was found to encode a putative MAP kinase kinase kinase similar to CTR1, a negative regulator of ethylene responses in Arabidopsis. Taken together, these data suggest that EDR1 functions at the top of a MAP kinase cascade that negatively regulates SA-inducible defense responses. Putative orthologs of EDR1 are present in monocots such as rice and barley, indicating that EDR1 may regulate defense responses in a wide range of crop species. PMID- 11114161 TI - Malaria parasite exit from the host erythrocyte: a two-step process requiring extraerythrocytic proteolysis. AB - Intraerythrocytic malaria parasites replicate by the process of schizogeny, during which time they copy their genetic material and package it into infective merozoites. These merozoites must then exit the host cell to invade new erythrocytes. To better characterize the events of merozoite escape, erythrocytes containing Plasmodium falciparum schizonts were cultured in the presence of the cysteine protease inhibitor, l-transepoxy-succinyl-leucylamido-(4 guanidino)butane (E64). This treatment resulted in the accumulation of extraerythrocytic merozoites locked within a thin, transparent membrane. Immunomicroscopy demonstrated that the single membrane surrounding the merozoites is not erythrocytic but rather is derived from the parasitophorous vacuolar membrane (PVM). Importantly, structures identical in appearance can be detected in untreated cultures at low frequency. Further studies revealed that (i) merozoites from the PVM-enclosed merozoite structures (PEMS) are invasive, viable, and capable of normal development; (ii) PEMS can be purified easily and efficiently; and (iii) when PEMS are added to uninfected red blood cells, released merozoites can establish a synchronous wave of infection. These observations suggest that l-transepoxy-succinyl-leucylamido-(4-guanidino)butane (E64) causes an accumulation of an intermediate normally present during the process of rupture. We propose a model for the process of rupture: merozoites enclosed within the PVM first exit from the host erythrocyte and then rapidly escape from the PVM by a proteolysis-dependent mechanism. PMID- 11114162 TI - spr-2, a suppressor of the egg-laying defect caused by loss of sel-12 presenilin in Caenorhabditis elegans, is a member of the SET protein subfamily. AB - Presenilin plays critical roles in the genesis of Alzheimer's disease and in LIN 12/Notch signaling during development. Here, we describe a screen for genes that influence presenilin level or activity in Caenorhabditis elegans. We identified four spr (suppressor of presenilin) genes by reverting the egg-laying defective phenotype caused by a null allele of the sel-12 presenilin gene. We analyzed the spr-2 gene in some detail. We show that loss of spr-2 activity suppresses the egg laying defective phenotype of different sel-12 alleles and requires activity of the hop-1 presenilin gene, suggesting that suppression is accomplished by elevating presenilin activity rather than by bypassing the need for presenilin activity. We also show that SPR-2 is a nuclear protein and is a member of a protein subfamily that includes human SET, which has been identified in numerous different biochemical assays and at translocation breakpoints associated with a subtype of acute myeloid leukemia. PMID- 11114163 TI - A secondary drug resistance mutation of TEM-1 beta-lactamase that suppresses misfolding and aggregation. AB - In Gram-negative bacteria, TEM-1 beta-lactamase provides the major mechanism of plasmid-mediated beta-lactam resistance. Natural variants of TEM-1 with increased antibiotic resistance have appeared in response to the use of extended-spectrum beta-lactam antibiotics (e.g., ceftazidime) and beta-lactamase inhibitors (e.g., clavulanic acid). Some of the variant enzymes are more efficient at catalyzing beta-lactam hydrolysis, whereas others are more resistant to inhibitors. M182T is a substitution observed in both types of variant TEM-1 beta-lactamases. This mutation is found only in combination with other amino acid substitutions, suggesting that it may correct defects introduced by other mutations that alter the specificity. An engineered core mutation, L76N, which diminishes the periplasmic beta-lactamase activity by 100-fold, was used as a model to understand the mechanism of suppression of the M182T mutation. Biochemical studies of the L76N enzyme alone and in combination with the M182T mutation indicate that the M182T substitution acts at the level of folding but does not affect the thermodynamic stability of TEM-1 beta-lactamase. Thus, the M182T substitution is an example of a naturally occurring mutation that has evolved to alter the folding pathway of a protein and confer a selective advantage during the evolution of drug resistance. PMID- 11114164 TI - The Sir2 protein family: A novel deacetylase for gene silencing and more. PMID- 11114165 TI - A recombinant rabies virus expressing vesicular stomatitis virus glycoprotein fails to protect against rabies virus infection. AB - To investigate the importance of the rabies virus (RV) glycoprotein (G) in protection against rabies, we constructed a recombinant RV (rRV) in which the RV G ecto- and transmembrane domains were replaced with the corresponding regions of vesicular stomatitis virus (VSV) glycoprotein (rRV-VSV-G). We were able to recover rRV-VSV-G and found that particle production was equal to rRV. However, the budding of the chimeric virus was delayed and infectious titers were reduced 10-fold compared with the parental rRV strain containing RV G. Biochemical analysis showed equal replication rates of both viruses, and similar amounts of wild-type and chimeric G were present in the respective viral particles. Additional studies were performed to determine whether the immune response against rRV-VSV-G was sufficient to protect against rabies. Mice were primed with rRV or rRV-VSV-G and challenged with a pathogenic strain of RV 12 days later. Similar immune responses against the internal viral proteins of both viruses indicated successful infection. All mice receiving the rRV vaccine survived the challenge, whereas immunization with rRV-VSV-G did not induce protection. The results confirm the crucial role of RV G in an RV vaccine. PMID- 11114167 TI - Pivotal role of cyclic nucleoside phosphodiesterase 4 in Tat-mediated CD4+ T cell hyperactivation and HIV type 1 replication. AB - We show here that HIV type 1 (HIV-1) Tat protein, in combination with anti CD3/CD28 mAbs, promotes IL-2 production and proliferation of primary CD4(+) T lymphocytes, obtained from HIV-1-seronegative donors. This effect was observed when Tat was immobilized on a solid support, but it was not observed with soluble Tat. Such hyperactivation was accomplished by recruiting the rolipram-sensitive cyclic nucleoside phosphodiesterase 4 and resulted in increased susceptibility to HIV-1 infection. Accordingly, rolipram potently inhibited HIV-1 replication in cultures stimulated by anti-CD3/CD28 +/- Tat. These results add to the concept that decreasing Tat activity is an important addition to anti-HIV-1 therapy, and they suggest a target for anti-HIV-1 chemotherapy, phosphodiesterase 4. PMID- 11114166 TI - Cytoplasmic-nuclear shuttling of FKBP12-rapamycin-associated protein is involved in rapamycin-sensitive signaling and translation initiation. AB - Translation initiation is one of the key events regulated in response to mitogenic stimulation and nutrient availability, tightly coupled to mammalian cell cycle progression and growth. FKBP12-rapamycin-associated protein (FRAP; also named mTOR or RAFT1), a member of the ataxia telangiectasia mutated (ATM) related kinase family, governs a rapamycin-sensitive membrane-to-cytoplasm signaling cascade that modulates translation initiation via p70 S6 kinase (p70(s6k)) and eIF-4E binding protein 1 (4E-BP1). Our studies reported here reveal a surprising regulatory mechanism of this signaling, which involves cytoplasmic-nuclear shuttling of FRAP. By using leptomycin B (LMB), a specific inhibitor of nuclear export receptor Crm1, we show that FRAP is a cytoplasmic nuclear shuttling protein. Inhibition of FRAP nuclear export by LMB coincides with diminished p70(s6k) activation and 4E-BP1 phosphorylation. Further investigation by altering FRAP's nuclear shuttling activity with exogenous nuclear import and export signals has yielded results that are consistent with a direct link between nuclear shuttling of FRAP and mitogenic stimulation of p70(s6k) activation and 4E-BP1 phosphorylation. Furthermore, by using a reporter system, we provide evidence suggesting that nuclear shuttling of FRAP regulates mitogen-stimulated rapamycin-sensitive translation initiation. These findings uncover a function for the nucleus in the direct regulation of the protein synthesis machinery via extracellular signals. PMID- 11114168 TI - Gene expression profiles in normal and Otx2-/- early gastrulating mouse embryos. AB - The mouse Otx2 gene is a homeobox transcription factor required as early as gastrulation for the proper development of the head. We compared gene expression profiles in wild-type and Otx2(-/-) 6.5 days postcoitum embryos by using a serial analysis of gene expression assay adapted to microdissected structures. Among a broader list, the study of six genes found to be differentially expressed allows defining a role for Otx2 in the orchestration of cell movements leading to the adequate organization of the embryo before gastrulation. PMID- 11114169 TI - Another role highlighted for estrogens in the male: sexual behavior. PMID- 11114170 TI - Mass spectrometry of proteins of known mass. PMID- 11114171 TI - Polarity of transcriptional enhancement revealed by an insulator element. AB - Transcriptional enhancers for genes transcribed by RNA polymerase II may be localized upstream or downstream of the stimulated promoter in their normal chromosomal context. They stimulate transcription in an orientation-independent manner when assayed on circular plasmids. We describe a transient transformation system to evaluate the orientation preference of transcriptional enhancers in Drosophila. To accomplish this, the gypsy insulator element was used to block bidirectional action of an enhancer on circular plasmids. In this system, as in the chromosome, blocking of enhancer activity requires wild-type levels of the su(Hw) protein. We evaluated the orientation preference for the relatively large (4.4 kb) Adh larval enhancer from Drosophila melanogaster, used in conjunction with a luciferase reporter gene under the control of a minimal Adh promoter. An orientation preference was revealed by insertion of a single copy of the insulator between the enhancer and the promoter. This orientation effect was greatly amplified when the promoter was weakened by removing binding sites for critical transcription factors, consistent with a mechanism of insulator action in which the insulator intercepts signals from the enhancer by competing with the promoter. The orientation preference, as much as 100-fold, is a property of the enhancer itself because it is displayed by gene constructions introduced into the chromosome regardless of the presence of the insulator in a distal location. These findings are most easily reconciled with a facilitated tracking mechanism for enhancer function in a native chromosomal environment. PMID- 11114172 TI - Associative memory hamiltonians for structure prediction without homology: alpha helical proteins. AB - Energy landscape theory is used to obtain optimized energy functions for predicting protein structure, without using homology information. At short sequence separation the energy functions are associative memory Hamiltonians constructed from a database of folding patterns in nonhomologous proteins and at large separations they have the form of simple pair potentials. The lowest energy minima provide reasonably accurate tertiary structures even though no homologous proteins are included in the construction of the Hamiltonian. We also quantify the funnel-like nature of these energy functions by using free energy profiles obtained by the multiple histogram method. PMID- 11114173 TI - Protein sequence signatures support the African clade of mammals. AB - DNA sequence evidence supports a superordinal clade of mammals that comprises elephants, sea cows, hyraxes, aardvarks, elephant shrews, golden moles, and tenrecs, which all have their origins in Africa, and therefore are dubbed Afrotheria. Morphologically, this appears an unlikely assemblage, which challenges-by including golden moles and tenrecs-the monophyly of the order Lipotyphla (Insectivora). We here identify in three proteins unique combinations of apomorphous amino acid replacements that support this clade. The statistical support for such "sequence signatures" as unambiguous synapomorphic evidence for the naturalness of the Afrotherian clade is reported. Using likelihood, combinatorial, and Bayesian methods we show that the posterior probability of the mammalian tree containing the Afrotherian clade is effectively 1.0, based on conservative assumptions. Presenting sequence data for another African insectivore, the otter shrew Micropotamogale lamottei, we demonstrate that such signatures are diagnostic for including newly investigated species in the Afrotheria. Sequence signatures provide "protein-morphological" synapomorphies that may aid in visualizing monophyletic groupings. PMID- 11114174 TI - Mixed model analysis of quantitative trait loci. AB - We develop a mixed model approach of quantitative trait locus (QTL) mapping for a hybrid population derived from the crosses of two or more distinguished outbred populations. Under the mixed model, we treat the mean allelic value of each source population as the fixed effect and the allelic deviations from the mean as random effects so that we can partition the total genetic variance into between- and within-population variances. Statistical inference of the QTL parameters is obtained by using the Bayesian method implemented by Markov chain Monte Carlo (MCMC). This unified QTL mapping algorithm treats the fixed and random model approaches as special cases of the general mixed model methodology. Utility and flexibility of the method are demonstrated by using a set of simulated data. PMID- 11114175 TI - Autosomal dominant myopathy: missense mutation (Glu-706 --> Lys) in the myosin heavy chain IIa gene. AB - We here report on a human myopathy associated with a mutation in a fast myosin heavy chain (MyHC) gene, and also the genetic defect in a hereditary inclusion body myopathy. The disorder has previously been described in a family with an "autosomal dominant myopathy, with joint contractures, ophthalmoplegia, and rimmed vacuoles." Linkage analysis and radiation hybrid mapping showed that the gene locus (Human Genome Map locus name: IBM3) is situated in a 2-Mb region of chromosome 17p13, where also a cluster of MyHC genes is located. These include the genes encoding embryonic, IIa, IIx/d, IIb, perinatal, and extraocular MyHCs. Morphological analysis of muscle biopsies from patients from the family indicated to us that the type 2A fibers frequently were abnormal, whereas other fiber types appeared normal. This observation prompted us to investigate the MyHC-IIa gene, since MyHC-IIa is the major isoform in type 2A fibers. The complete genomic sequence for this gene was deduced by using an "in silico" strategy. The gene, found to consist of 38 exons, was subjected to a complete mutation scan in patients and controls. We identified a missense mutation, Glu-706 --> Lys, which is located in a highly conserved region of the motor domain, the so-called SH1 helix region. By conformational changes this region communicates activity at the nucleotide-binding site to the neck region, resulting in the lever arm swing. The mutation in this region is likely to result in a dysfunctional myosin, compatible with the disorder in the family. PMID- 11114176 TI - Integrin alpha(v)beta(3) mediates rotavirus cell entry. AB - Rotavirus strains differ in their need for sialic acid (SA) for initial binding to the cell surface; however, the existence of a postattachment cell receptor, common to most, if not all, rotavirus strains, has been proposed. In the present study, antibodies to the alpha(v) and beta(3) integrin subunits, and the alpha(v)beta(3) ligand, vitronectin, efficiently blocked the infectivity of the SA-dependent rhesus rotavirus RRV, its SA-independent variant nar3, and the neuraminidase-resistant human rotavirus strain Wa. Vitronectin and anti-beta(3) antibodies, however, did not block the binding of virus to cells, indicating that rotaviruses interact with alpha(v)beta(3) at a postbinding step, probably penetration. This interaction was shown to be independent of the tripeptide motif arginine-glycine-aspartic acid present in the natural ligands of this integrin. Transfection of CHO cells with alpha(v)beta(3) genes significantly increased their permissiveness to all three rotavirus strains, and the increment of virus infectivity was reverted by incubation of these cells either with antibodies to beta(3) or with vitronectin. These findings implicate alpha(v)beta(3) integrin as a cellular receptor common to neuraminidase-sensitive and neuraminidase-resistant rotaviruses, and support the hypothesis that this integrin could determine, at least in part, the cellular susceptibility to rotaviruses. PMID- 11114177 TI - Referred phantom sensations and cortical reorganization after spinal cord injury in humans. AB - To test the hypothesis that cortical remapping supports phantom sensations, we examined referred phantom sensations and cortical activation in humans after spinal-cord injury (SCI) at the thoracic level (T3-T12). Of 12 SCI subjects, 9 reported phantom sensations, and 2 reported referred phantom sensations. In both of these subjects, referred phantom sensations were evoked by contact in reference zones (RZ) that were not adjacent in the periphery and were not predicted to be adjacent in the postcentral gyrus (PoCG), suggesting that representations separated by centimeters of cortical space were simultaneously engaged. This finding was supported by functional MRI (fMRI). In a subject with a T6-level complete SCI, contact in RZ on the left or right forearm projected referred phantom sensations to the ipsilateral chest. During fMRI, contact in either forearm RZ evoked activity in the central PoCG (the position of the forearm representation) and the medial PoCG (the position of the chest representation) with >/=1.6 cm of nonresponsive cortex intervening. In contrast, stimulation in non-RZ forearm and palm regions in this subject and in lesion matched SCI subjects evoked central but not medial PoCG activation. Our findings support a relation between PoCG activation and the percept of referred phantom sensations. These results, however, present an alternative to somatotopic cortical reorganization, namely, cortical plasticity expressed in coactivation of nonadjacent representations. The observed pattern suggests that somatotopic subcortical remapping, projected to the cortex, can support perceptual and cortical reorganization after deafferentation in humans. PMID- 11114178 TI - Ama1p is a meiosis-specific regulator of the anaphase promoting complex/cyclosome in yeast. AB - Meiosis is the developmental program by which diploid organisms produce haploid gametes capable of sexual reproduction. Here we describe the yeast gene AMA1, a new member of the Cdc20 protein family that regulates the multisubunit ubiquitin ligase termed the anaphase promoting complex/cyclosome (APC/C). AMA1 is developmentally regulated in that its transcription and splicing occur only in meiotic cells. The meiosis-specific processing of AMA1 mRNA depends on the previously described MER1 splicing factor. Several results indicate that Ama1p is required for APC/C function during meiosis. First, coimmunoprecipitation assays indicate that Ama1p associates with the APC/C in vivo. Second, Ama1p is required for the degradation of the B-type cyclin Clb1p, an APC/C substrate in both meiotic and mitotic cells. Third, ectopic overexpression of AMA1 is able to stimulate ubiquitination of Clb1p in vitro and degradation of Clb1p in vivo. Mutants lacking AMA1 revealed that it is required for the first meiotic division but not the mitotic-like meiosis II. In addition, ama1 mutants are defective for both spore wall assembly and the expression of late meiotic genes. In conclusion, this study indicates that Ama1p directs a meiotic APC/C that functions solely outside mitotic cell division. The requirement of Ama1p only for meiosis I and spore morphogenesis suggests a function for APC/C(Ama1) specifically adapted to germ cell development. PMID- 11114179 TI - DNA tightens the dimeric DNA-binding domain of human papillomavirus E2 protein without changes in volume. AB - The recognition of palindromic specific DNA sequences by the human papillomavirus (HPV) E2 proteins is responsible for regulation of virus transcription. The dimeric E2 DNA-binding domain of HPV-16 (E2c) dissociates into a partially folded state under high hydrostatic pressure. We show here that pressure-induced monomers of E2c are highly structured, as evidenced by NMR hydrogen-deuterium exchange measurements. On binding to both specific and nonspecific DNA, E2c becomes stable against pressure. Competitive binding studies using fluorescence polarization of fluorescein-labeled DNA demonstrate the reversibility of the specific binding. To assess the thermodynamic parameters for the linkage between protein dissociation and DNA binding, urea denaturation curves were obtained at different pressures in the presence of specific and nonspecific DNA sequences. The change in free energy on denaturation fell linearly with increase in pressure for both protein-DNA complexes, and the measured volume change was similar to that obtained for E2c alone. The data show that the free energy of dissociation increases when E2c binds to a nonspecific DNA sequence but increases even more when the protein binds to the specific DNA sequence. Thus, specific complexes are tighter but do not entail variation in the volume change. The thermodynamic data indicate that DNA-bound E2c dissociates into monomers bound to DNA. The existence of monomeric units of E2c bound to DNA may have implications for the formation of DNA loops, as an additional target for viral and host factors binding to the loosely associated dimer of the N-terminal module of the E2 protein. PMID- 11114180 TI - Caveolin-1 down-regulates inducible nitric oxide synthase via the proteasome pathway in human colon carcinoma cells. AB - To investigate whether caveolin-1 (cav-1) may modulate inducible nitric oxide synthase (iNOS) function in intact cells, the human intestinal carcinoma cell lines HT29 and DLD1 that have low endogenous cav-1 levels were transfected with cav-1 cDNA. In nontransfected cells, iNOS mRNA and protein levels were increased by the addition of a mix of cytokines. Ectopic expression of cav-1 in both cell lines correlated with significantly decreased iNOS activity and protein levels. This effect was linked to a posttranscriptional mechanism involving enhanced iNOS protein degradation by the proteasome pathway, because (i) induction of iNOS mRNA by cytokines was not affected and (ii) iNOS protein levels increased in the presence of the proteasome inhibitors N-acetyl-Leu-Leu-Norleucinal and lactacystin. In addition, a small amount of iNOS was found to cofractionate with cav-1 in Triton X-100-insoluble membrane fractions where also iNOS degradation was apparent. As has been described for endothelial and neuronal NOS isoenzymes, direct binding between cav-1 and human iNOS was detected in vitro. Taken together, these results suggest that cav-1 promotes iNOS presence in detergent insoluble membrane fractions and degradation there via the proteasome pathway. PMID- 11114181 TI - The inducible N-acetylglucosamine catabolic pathway gene cluster in Candida albicans: discrete N-acetylglucosamine-inducible factors interact at the promoter of NAG1. AB - The catabolic pathway of N-acetylglucosamine (GlcNAc) in Candida albicans is an important facet of its pathogenicity. One of the pathway genes, encoding glucosamine-6-phosphate deaminase (NAG1) is transcriptionally regulated by GlcNAc. Sequence analysis of a 4-kb genomic clone containing NAG1 indicates that this gene is part of a cluster containing two other genes of the GlcNAc catabolic pathway, i.e., DAC1, GlcNAc-6-phosphate deacetylase, and HXK1, hexokinase. All three genes are temporally and coordinately induced by GlcNAc suggesting a common regulatory mechanism for these genes. The NAG1 promoter is up-regulated when induced by GlcNAc in C. albicans but not in Saccharomyces cerevisiae. In vivo analysis of the deletion constructs delineated the minimal promoter to -130 bp and mapped two regions at -200 and -400 bp upstream of +1 (ATG) responsible for GlcNAc induction. Gel mobility-shift assays and "footprinting" (DNase protection method) analyses revealed two regions, 5'-GGAGCAAAAAAATGT 3' (-164 to -150, box A) and 5'-ACGGTGAGTTG 3' (-291 to -281, box B), that are recognized and bound by at least two inducible activator proteins directing the regulation of gene expression. PMID- 11114182 TI - Fast kinetics of chromatin assembly revealed by single-molecule videomicroscopy and scanning force microscopy. AB - Fluorescence videomicroscopy and scanning force microscopy were used to follow, in real time, chromatin assembly on individual DNA molecules immersed in cell free systems competent for physiological chromatin assembly. Within a few seconds, molecules are already compacted into a form exhibiting strong similarities to native chromatin fibers. In these extracts, the compaction rate is more than 100 times faster than expected from standard biochemical assays. Our data provide definite information on the forces involved (a few piconewtons) and on the reaction path. DNA compaction as a function of time revealed unique features of the assembly reaction in these extracts. They imply a sequential process with at least three steps, involving DNA wrapping as the final event. An absolute and quantitative measure of the kinetic parameters of the early steps in chromatin assembly under physiological conditions could thus be obtained. PMID- 11114183 TI - Abolition of male sexual behaviors in mice lacking estrogen receptors alpha and beta (alpha beta ERKO). AB - Male mice with a knockout of the estrogen receptor (ER)-alpha gene, a ligand activated transcription factor, showed reduced levels of intromissions and no ejaculations whereas simple mounting behavior was not affected. In contrast, all components of sexual behaviors were intact in male mice lacking the novel ER-beta gene. Here we measure the extent of phenotype in mice that lack both ER-alpha and ER-beta genes (alphabetaERKO). alphabetaERKO male mice did not show any components of sexual behaviors, including simple mounting behavior. Nor did they show ultrasonic vocalizations during behavioral tests with receptive female mice. On the other hand, reduced aggressive behaviors of alphabetaERKO mice mimicked those of single knockout mice of ER-alpha gene (alphaERKO). They showed reduced levels of lunge and bite aggression, but rarely showed offensive attacks. Thus, either one of the ERs is sufficient for the expression of simple mounting in male mice, indicating a redundancy in function. Offensive attacks, on the other hand, depend specifically on the ER-alpha gene. Different patterns of natural behaviors require different patterns of functions by ER genes. PMID- 11114184 TI - Effect of Bradyrhizobium photosynthesis on stem nodulation of Aeschynomene sensitiva. AB - Some leguminous species of the genus Aeschynomene are specifically stem-nodulated by photosynthetic bradyrhizobia. To study the effect of bacterial photosynthesis during symbiosis, we generated a photosynthesis-negative mutant of the Bradyrhizobium sp. strain ORS278 symbiont of Aeschynomene sensitiva. The presence of a functional photosynthetic unit in bacteroids and the high expression of the photosynthetic genes observed in stem nodules demonstrate that the bacteria are photosynthetically active during stem symbiosis. Stem inoculation by the photosynthetic mutant gave a 50% decrease in stem-nodule number, which reduced nitrogen fixation activity and plant growth in the same proportion. These results indicate an important role of bacterial photosynthesis in the efficiency of stem nodulation. PMID- 11114185 TI - Reprogramming leukemic cells to terminal differentiation by inhibiting specific cyclin-dependent kinases in G1. AB - Some tumor cells can be stimulated to differentiate and undergo terminal cell division and loss of tumorigenicity. The in vitro differentiation of murine erythroleukemia (MEL) cells is a dramatic example of tumor-cell reprogramming. We found that reentry of MEL cells into terminal differentiation is accompanied by an early transient decline in the activity of cyclin-dependant kinase (CDK) 2, followed by a decline of CDK6. Later, as cells undergo terminal arrest, CDK2 and CDK4 activities decline. By analyzing stable MEL-cell transfectants containing vectors directing inducible expression of specific CDK inhibitors, we show that only inhibitors that block the combination of CDK2 and CDK6 trigger differentiation. Inhibiting CDK2 and CDK4 does not cause differentiation. Importantly, we also show that reprogramming through inhibition of CDKs is restricted to G(1) phase of the cell cycle. The results imply that abrogation of normal cell-cycle controls in tumor cells contributes to their inability to differentiate fully and that restoration of such controls in G(1) can lead to resumption of differentiation and terminal cell division. The results also indicate that CDK4 and CDK6 are functionally distinct and support our hypothesis that the two CDKs regulate cell division at different stages of erythroid maturation. PMID- 11114186 TI - Mutational studies on HslU and its docking mode with HslV. AB - HslVU is an ATP-dependent prokaryotic protease complex. Despite detailed crystal and molecular structure determinations of free HslV and HslU, the mechanism of ATP-dependent peptide and protein hydrolysis remained unclear, mainly because the productive complex of HslV and HslU could not be unambiguously identified from the crystal data. In the crystalline complex, the I domains of HslU interact with HslV. Observations based on electron microscopy data were interpreted in the light of the crystal structure to indicate an alternative mode of association with the intermediate domains away from HslV. By generation and analysis of two dozen HslU mutants, we find that the amidolytic and caseinolytic activities of HslVU are quite robust to mutations on both alternative docking surfaces on HslU. In contrast, HslVU activity against the maltose-binding protein-SulA fusion protein depends on the presence of the I domain and is also sensitive to mutations in the N-terminal and C-terminal domains of HslU. Mutational studies around the hexameric pore of HslU seem to show that it is involved in the recognition/translocation of maltose-binding protein-SulA but not of chromogenic small substrates and casein. ATP-binding site mutations, among other things, confirm the essential role of the "sensor arginine" (R393) and the "arginine finger" (R325) in the ATPase action of HslU and demonstrate an important role for E321. Additionally, we report a better refined structure of the HslVU complex crystallized along with resorufin-labeled casein. PMID- 11114187 TI - Identification of XPR-1, a progesterone receptor required for Xenopus oocyte activation. AB - Quiescent full-grown Xenopus oocytes remain arrested at the G(2)/M border of meiosis I until exposed to progesterone, their natural mitogen. Progesterone triggers rapid, nontranscriptional responses that lead to the translational activation of stored mRNAs, resumption of the meiotic cell cycles, and maturation of the oocyte into a fertilizable egg. It has long been presumed that progesterone activates the oocyte through a novel nontranscriptional signaling receptor. Here, we provide evidence that a conventional transcriptional progesterone receptor cloned from Xenopus oocytes, XPR-1, is required for oocyte activation. Overexpression of XPR-1 through mRNA injection increases sensitivity to progesterone and accelerates progesterone-activated cell cycle reentry. Injection of XPR-1 antisense oligonucleotides blocks the ability of oocytes to respond to progesterone; these oocytes are rescued by subsequent injection of XPR 1 or the human progesterone receptor PR-B. Antisense-treated oocytes can be activated in response to inhibition of protein kinase A, one of the earliest known changes occurring downstream of progesterone stimulation. These results argue that the conventional progesterone receptor also functions as the signaling receptor that is responsible for the rapid nontranscriptional activation of frog oocytes. PMID- 11114188 TI - Histone deacetylase 4 associates with extracellular signal-regulated kinases 1 and 2, and its cellular localization is regulated by oncogenic Ras. AB - Histone deacetylase 4 (HDAC4) is a member of a family of enzymes that catalyze the removal of acetyl groups from core histones, resulting in a compact chromatin structure that is generally associated with repressed gene transcription. Protein phosphorylation has been implicated in the regulation of the corepressor activity of the deacetylase. Here we report that serine/threonine kinases are found in association with HDAC4 and phosphorylate HDAC4 in vitro, and HDAC4 is phosphorylated in cells. The extracellular signal-regulated kinases 1 and 2 (ERK1/2), also known as p44(MAPK) and p42(MAPK), respectively, are two of the kinases associated with HDAC4. ERK1/2 are components of the Ras-mitogen-activated protein kinase (MAPK) signal transduction pathway. Activation of the Ras-MAPK pathway by expression of oncogenic Ras or constitutively active MAPK/ERK kinase 1 results in an increased percentage of cells (from approximately 10% to approximately 70%) that express HDAC4 in the nucleus in C2C12 myoblast cells. In cells transfected with oncogenic Ras, nuclear HDAC4 is associated with kinase activity. Our results provide evidence that protein kinase activity is present in a protein complex with HDAC4 and directly links the Ras-MAPK signal transduction pathway to a mechanism for chromatin remodeling (i.e., histone deacetylation). PMID- 11114189 TI - Topology to geometry in protein folding: beta-lactoglobulin. AB - Evolution of protein structure from random coil to native is first represented topologically by its time-dependent sequences of discretized Ramachandran basins occupied by successive backbone residues. Introducing energetic and entropic criteria at each instant of observation transforms the description from a structurally ambiguous topological representation to an unambiguous geometric picture of the folding process. The method is applied with success to folding of beta-lactoglobulin, traditionally perplexing because of its reputed nonhierarchical folding pattern. This molecule passes through a stage, ca. 0.1 microseconds duration, of transient, "flickering" alpha-helical structure, until a bit of tertiary structure forms that stabilizes the system long enough to allow it to pass to its native beta-sheet. PMID- 11114190 TI - Efficient transduction of neural cells in vitro and in vivo by a baculovirus derived vector. AB - Gene delivery to the central nervous system is central to the development of gene therapy for neurological diseases. We developed a baculovirus-derived vector, the Bac-CMV-GFP vector, containing a reporter gene encoding for the green fluorescent protein (GFP) under the control of the cytomegalovirus (CMV) promoter. Two neuroblastomal cell lines and three human primary neural cultures could be efficiently transduced. In all cases, addition of butyrate, an inhibitor of histone deacetylase, increased the level of expression in terms of the number of GFP-expressing cells and the intensity of fluorescence. The level of expression in a human telencephalic culture was over 50% of transduced cells with a multiplicity of infection of 25. GFP expression was demonstrated to be genuine expression and not pseudotransduction of the reporter protein. Most interestingly, Bac-CMV-GFP could transduce neural cells in vivo when directly injected into the brain of rodents and was not inactivated by the complement system. Thus, baculovirus is a promising tool for gene transfer into the central nervous system both for studies of the function of foreign genes and the development of gene therapy strategies. PMID- 11114191 TI - Structural organization of yeast and mammalian mediator complexes. AB - Structures of yeast Mediator complex, of a related complex from mouse cells and of thyroid hormone receptor-associated protein complex from human cells have been determined by three-dimensional reconstruction from electron micrographs of single particles. All three complexes show a division in two parts, a "head" domain and a combined "middle-tail" domain. The head domains of the three complexes appear most similar and interact most closely with RNA polymerase II. The middle-tail domains show the greatest structural divergence and, in the case of the tail domain, may not interact with polymerase at all. Consistent with this structural divergence, analysis of a yeast Mediator mutant localizes subunits that are not conserved between yeast and mammalian cells to the tail domain. Biochemically defined Rgr1 and Srb4 modules of yeast Mediator are then assigned to the middle and head domains. PMID- 11114192 TI - Role of the conserved WHXL motif in the C terminus of synaptotagmin in synaptic vesicle docking. AB - Synaptotagmin (Syt) I, an abundant synaptic vesicle protein, consists of one transmembrane region, two C2 domains, and a short C terminus. This protein is essential for both synaptic vesicle exocytosis and endocytosis via its C2 domains. Although the short C terminus is highly conserved among the Syt family and across species, little is known about the exact role of the conserved C terminus of Syt I. In this paper, we report a function of the Syt I C terminus in synaptic vesicle docking at the active zones. Presynaptic injection of a peptide corresponding to the C-terminal 21 amino acids of Syt I (named Syt-C) into the squid giant synapse blocked synaptic transmission without affecting the presynaptic action potential or the presynaptic Ca(2+) currents. The same procedure repeated with a mutant C-terminal peptide (Syt-CM) had no effect on synaptic transmission. Repetitive presynaptic stimulation with Syt-C produced a rapid decrease in the amplitude of the postsynaptic potentials as the synaptic block progressed, indicating that the peptide interferes with the docking step rather than the fusion step of synaptic vesicles. Electron microscopy of the synapses injected with the Syt-C peptide showed a marked decrease in the number of docked synaptic vesicles at the active zones, as compared with controls. These results indicate that Syt I is a multifunctional protein that is involved in at least three steps of synaptic vesicle cycle: docking, fusion, and reuptake of synaptic vesicles. PMID- 11114193 TI - Functional genomics reveals a family of eukaryotic oxidation protection genes. AB - Reactive oxygen species (ROS) are toxic compounds produced by normal metabolic processes. Their reactivity with cellular components is a major stress for aerobic cells that results in lipid, protein, and DNA damage. ROS-mediated DNA damage contributes to spontaneous mutagenesis, and cells deficient in repair and protective mechanisms have elevated levels of spontaneous mutations. In Escherichia coli a large number of genes are involved in the repair of oxidative DNA damage and its prevention by detoxification of ROS. In humans, the genes required for these processes are not well defined. In this report we describe the human OXR1 (oxidation resistance) gene discovered in a search for human genes that function in protection against oxidative damage. OXR1 is a member of a conserved family of genes found in eukaryotes but not in prokaryotes. We also outline the procedures developed to identify human genes involved in the prevention and repair of oxidative damage that were used to identify the human OXR1 gene. This procedure makes use of the spontaneous mutator phenotype of E. coli oxidative repair-deficient mutants and identifies genes of interest by screening for antimutator activity resulting from cDNA expression. PMID- 11114194 TI - Functional redundancy of cryptochromes and classical photoreceptors for nonvisual ocular photoreception in mice. AB - The daily light-dark (LD) cycle exerts a powerful influence on the temporal organization of behavior and physiology. Much of this influence is preserved in behaviorally blind retinally degenerate mice; the photoreceptors underlying this nonvisual phototransduction are unknown. The mammalian eye contains at least two classes of photoactive pigments, the vitamin A-based opsins and the vitamin B(2) based cryptochromes. To genetically define the roles of these pigments in light modulation of behavior, we generated rd/rd;mCry1(-)/mCry1(-);mCry2(-)/mCry2(-) mutant mice lacking rods and most cones as well as both cryptochrome proteins. The response of the mutant mouse to photic input was analyzed at both behavioral and molecular levels. Behaviorally, mice lacking either classical photoreceptors or cryptochromes exhibited strongly rhythmic locomotor responses to 10 and 100 lux daily LD 12 h/12-h cycles; however, triple mutant mice carrying both cryptochrome and retinal degenerate mutations were nearly arrhythmic under both LD cycles and in constant darkness. At the molecular level, the light induction of c-fos transcription in the suprachiasmatic nucleus was markedly reduced in the triple mutant mouse compared with either rd/rd or cryptochrome mutant mice. These data indicate that classical opsins and cryptochromes serve functionally redundant roles in the transduction of light information to behavioral modulation and suggest a pleomorphic role for cryptochromes in both photoreception and central clock mechanism. PMID- 11114195 TI - Identification of ligands and coligands for the ecdysone-regulated gene switch. AB - The ecdysone-inducible gene switch is a useful tool for modulating gene expression in mammalian cells and transgenic animals. We have identified inducers derived from plants as well as certain classes of insecticides that increase the versatility of this gene regulation system. Phytoecdysteroids share the favorable kinetics of steroids, but are inert in mammals. The gene regulation properties of one of these ecdysteroids have been examined in cell culture and in newly developed strains of ecdysone-system transgenic mice. Ponasterone A is a potent regulator of gene expression in cells and transgenic animals, enabling reporter genes to be turned on and off rapidly. A number of nonsteroidal insecticides have been identified that also activate the ecdysone system. Because the gene controlling properties of the ecdysone switch are based on a heterodimer composed of a modified ecdysone receptor (VgEcR) and the retinoid X receptor (RXR), we have tested the effect of RXR ligands on the VgEcR/RXR complex. Used alone, RXR ligands display no activity on the ecdysone switch. However, when used in combination with a VgEcR ligand, RXR ligands dramatically enhance the absolute levels of induction. This property of the heterodimer has allowed the development of superinducer combinations that increase the dynamic range of the system. PMID- 11114196 TI - Calsarcins, a novel family of sarcomeric calcineurin-binding proteins. AB - The calcium- and calmodulin-dependent protein phosphatase calcineurin has been implicated in the transduction of signals that control the hypertrophy of cardiac muscle and slow fiber gene expression in skeletal muscle. To identify proteins that mediate the effects of calcineurin on striated muscles, we used the calcineurin catalytic subunit in a two-hybrid screen for cardiac calcineurin interacting proteins. From this screen, we discovered a member of a novel family of calcineurin-interacting proteins, termed calsarcins, which tether calcineurin to alpha-actinin at the z-line of the sarcomere of cardiac and skeletal muscle cells. Calsarcin-1 and calsarcin-2 are expressed in developing cardiac and skeletal muscle during embryogenesis, but calsarcin-1 is expressed specifically in adult cardiac and slow-twitch skeletal muscle, whereas calsarcin-2 is restricted to fast skeletal muscle. Calsarcins represent a novel family of sarcomeric proteins that link calcineurin with the contractile apparatus, thereby potentially coupling muscle activity to calcineurin activation. PMID- 11114197 TI - Activation of the myocyte enhancer factor-2 transcription factor by calcium/calmodulin-dependent protein kinase-stimulated binding of 14-3-3 to histone deacetylase 5. AB - Skeletal muscle differentiation is controlled by interactions between myocyte enhancer factor-2 (MEF2) and myogenic basic helix-loop-helix transcription factors. Association of MEF2 with histone deacetylases (HDAC) -4 and -5 results in repression of MEF2 target genes and inhibition of myogenesis. Calcium/calmodulin-dependent protein kinase (CaMK) signaling promotes myogenesis by disrupting MEF2-HDAC complexes and stimulating HDAC nuclear export. To further define the mechanisms that confer CaMK responsiveness to HDAC4 and -5, we performed yeast two-hybrid screens to identify HDAC-interacting factors. These screens revealed interactions between HDAC4 and members of the 14-3-3 family of proteins, which function as signal-dependent intracellular chaperones. HDAC4 binds constitutively to 14-3-3 in yeast and mammalian cells, whereas HDAC5 binding to 14-3-3 is largely dependent on CaMK signaling. CaMK phosphorylates serines -259 and -498 in HDAC5, which subsequently serve as docking sites for 14 3-3. Our studies suggest that 14-3-3 binding to HDAC5 is required for CaMK dependent disruption of MEF2-HDAC complexes and nuclear export of HDAC5, and implicate 14-3-3 as a signal-dependent regulator of muscle cell differentiation. PMID- 11114198 TI - Prenatal exposure to high levels of glucocorticoids increases the susceptibility of cerebellar granule cells to oxidative stress-induced cell death. AB - There is growing concern that prenatal exposure to excessive glucocorticoids may have deleterious effects on the development of various organs, including the nervous system. This study aimed at evaluating whether prenatal exposure to high levels of glucocorticoids might produce long-term effects on neuronal cell survival. Pregnant rats were injected i.p. with 0.1 mg/kg dexamethasone (DEX) from day 14 postconception, and cerebellar granule cells (CGC) were prepared from 1-week-old rats from DEX-treated and control dams. After 7 days in culture, cells were exposed to H(2)O(2), methylmercury, or colchicine at concentrations known to induce apoptotic cell death. After exposure to H(2)O(2) or methylmercury, both inducing oxidative stress, the number of apoptotic cells was significantly higher in DEX- than in control-CGC. Because mitochondria play a key role in apoptosis, mitochondrial function was investigated, and a decrease in the threshold level of Ca(2+) necessary for induction of mitochondrial permeability transition, in Ca(2+) accumulation rate, and in oxygen consumption was detected in DEX-CGC. Moreover, the activity of the antioxidant enzyme catalase was significantly decreased in DEX-CGC. A similar decrease in catalase activity was observed in cerebellar homogenate from newborn and 40-day-old DEX-rats. In conclusion, these results indicate that prenatal exposure to high levels of glucocorticoids induces long-lasting changes in CGC rendering them more sensitive to oxidative stress. With the increasing use of multiple doses of glucocorticoids in preterm infants, the possibility that prenatal exposure to excess glucocorticoids may lead to long term neurological consequences becomes a relevant issue. PMID- 11114199 TI - A kinetically significant intermediate in the folding of barnase. AB - A series of studies on the small protein barnase in the 1990s established it as a paradigm for protein folding in which there is a kinetically important intermediate. But, a recent study in PNAS claims that there are no stable intermediates on the folding pathway. I summarize the evidence that proves that the folding kinetics of barnase is inconsistent with the absence of a folding intermediate. I reinterpret the major evidence presented against the intermediate (an inflection in the unfolding limb of a chevron plot) and show that the inflection is precisely what is predicted from the energy diagram for a three state reaction with a kinetically significant on-pathway intermediate. The inflection is indicative of a change of rate determining step from the formation to breakdown of an intermediate on unfolding. Other evidence presented against the intermediate is, in fact, consistent with a kinetically important intermediate. I show how the complexities in the kinetics provide a means for measuring otherwise unobtainable rate constants and provide a strategy for mapping the structure of the early transition state in folding. Rather than refute multistate kinetics, the presence of the inflection in the unfolding plot constitutes a novel type of evidence for on-pathway folding intermediates. PMID- 11114200 TI - The SUR2 gene of Arabidopsis thaliana encodes the cytochrome P450 CYP83B1, a modulator of auxin homeostasis. AB - Genetic screens have been performed to identify mutants with altered auxin homeostasis in Arabidopsis. A tagged allele of the auxin-overproducing mutant sur2 was identified within a transposon mutagenized population. The SUR2 gene was cloned and shown to encode the CYP83B1 protein, which belongs to the large family of the P450-dependent monooxygenases. SUR2 expression is up-regulated in sur1 mutants and induced by exogenous auxin in the wild type. Analysis of indole-3 acetic acid (IAA) synthesis and metabolism in sur2 plants indicates that the mutation causes a conditional increase in the pool size of IAA through up regulation of IAA synthesis. PMID- 11114201 TI - Crystal structure of an intracellular protease from Pyrococcus horikoshii at 2-A resolution. AB - The intracellular protease from Pyrococcus horikoshii (PH1704) and PfpI from Pyrococcus furiosus are members of a class of intracellular proteases that have no sequence homology to any other known protease family. We report the crystal structure of PH1704 at 2.0-A resolution. The protease is tentatively identified as a cysteine protease based on the presence of cysteine (residue 100) in a nucleophile elbow motif. In the crystal, PH1704 forms a hexameric ring structure, and the active sites are formed at the interfaces between three pairs of monomers. PMID- 11114202 TI - Chemical defense: bestowal of a nuptial alkaloidal garment by a male moth on its mate. AB - Males of the moth Cosmosoma myrodora (Arctiidae) acquire pyrrolizidine alkaloid by feeding on the excrescent fluids of certain plants (for instance, Eupatorium capillifolium). They incorporate the alkaloid systemically and as a result are protected against spiders. The males have a pair of abdominal pouches, densely packed with fine cuticular filaments, which in alkaloid-fed males are alkaloid laden. The males discharge the filaments on the female in bursts during courtship, embellishing her with alkaloid as a result. The topical investiture protects the female against spiders. Alkaloid-free filaments, from alkaloid deprived males, convey no such protection. The males also transmit alkaloid to the female by seminal infusion. The systemic alkaloid thus received, which itself may contribute to the female's defense against spiders, is bestowed in part by the female on the eggs. Although paternal contribution to egg defense had previously been demonstrated for several arctiid moths, protective nuptial festooning of a female by its mate, such as is practiced by C. myrodora, appears to be without parallel among insects. PMID- 11114203 TI - RbcS suppressor mutations improve the thermal stability and CO2/O2 specificity of rbcL- mutant ribulose-1,5-bisphosphate carboxylase/oxygenase. AB - In the green alga Chlamydomonas reinhardtii, a Leu(290)-to-Phe (L290F) substitution in the large subunit of ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco), which is coded by the chloroplast rbcL gene, was previously found to be suppressed by second-site Ala(222)-to-Thr and Val(262)-to Leu substitutions. These substitutions complement the photosynthesis deficiency of the L290F mutant by restoring the decreased thermal stability, catalytic efficiency, and CO(2)/O(2) specificity of the mutant enzyme back to wild-type values. Because residues 222, 262, and 290 interact with the loop between beta strands A and B of the Rubisco small subunit, which is coded by RbcS1 and RbcS2 nuclear genes, it seemed possible that substitutions in this loop might also suppress L290F. A mutation in a nuclear gene, Rbc-1, was previously found to suppress the biochemical defects of the L290F enzyme at a posttranslational step, but the nature of this gene and its product remains unknown. In the present study, three nuclear-gene suppressors were found to be linked to each other but not to the Rbc-1 locus. DNA sequencing revealed that the RbcS2 genes of these suppressor strains have mutations that cause either Asn(54)-to-Ser or Ala(57)-to Val substitutions in the small-subunit betaA/betaB loop. When present in otherwise wild-type cells, with or without the resident RbcS1 gene, the mutant small subunits improve the thermal stability of wild-type Rubisco. These results indicate that the betaA/betaB loop, which is unique to eukaryotic Rubisco, contributes to holoenzyme thermal stability, catalytic efficiency, and CO(2)/O(2) specificity. The small subunit may be a fruitful target for engineering improved Rubisco. PMID- 11114204 TI - An alternative cytokinin biosynthesis pathway. AB - Studies of de novo cytokinin biosynthesis in isopentenyltransferase (ipt) transformed Arabidopsis thaliana, involving in vivo deuterium labeling and mass spectrometry, showed that the biosynthetic rate of zeatinriboside-5' monophosphate was around 66-fold higher than that of isopentenyladenosine-5' monophosphate (iPMP), the proposed primary product of the Agrobacterium ipt. Double tracer analysis, using [(2)H(6)] isopentenyladenosine and deuterium oxide, provided evidence for an alternative, iPMP-independent, biosynthetic pathway for zeatin-type cytokinins, present in both ipt-expressing and wild-type Arabidopsis thaliana. Reduction of the biosynthetic flux in the alternative pathway by use of mevastatin, an inhibitor for 3-hydroxy-3-methylglutaryl CoA reductase, indicated a terpenoid origin for the side-chain precursor of the iPMP independent pathway. PMID- 11114205 TI - Evolutionary biogeography on Mexico's Baja California peninsula: A synthesis of molecules and historical geology. PMID- 11114206 TI - Development of tolerance during chronic treatment of kindled rats with the novel antiepileptic drug levetiracetam. AB - PURPOSE: Levetiracetam (LEV) is an interesting novel antiepileptic drug with proven efficacy in both animal models and patients with partial epilepsy. To study whether the efficacy of the drug changes during chronic treatment, we evaluated the anticonvulsant activity of LEV during prolonged daily administration in amygdala-kindled rats. METHODS: On the basis of the anticonvulsant potency and duration of action after acute doses, LEV was administered in fully kindled rats three times daily at 108 mg/kg i.p. for 3 weeks. For the study of anticonvulsant efficacy, the afterdischarge threshold was determined before, during, and after the drug treatment period. To determine whether the pharmacokinetics of the drug change during prolonged treatment, LEV levels were repeatedly determined in plasma during the treatment period. RESULTS: LEV markedly increased the afterdischarge threshold and decreased the seizure severity and duration after initial dosing, but a marked loss of anticonvulsant efficacy developed during chronic treatment. This loss of efficacy was not due to alterations in drug elimination, indicating the development of functional tolerance to LEV during repeated administration. After the termination of treatment, no withdrawal signs (e.g., changes in behavior, body temperature, body weight, or seizure threshold) were observed. CONCLUSIONS: The data demonstrate that chronic administration of LEV leads to a significant reduction in anticonvulsant efficacy in the kindling model of temporal lobe epilepsy. Whether this experimental observation has clinical relevance must await monotherapy trials with long-term follow-up of patients who initially responded to LEV. PMID- 11114207 TI - Valproate prevents epileptiform activity after trauma in an in vitro model in neocortical slices. AB - PURPOSE: Epileptogenesis is a hallmark of severe cortical trauma, with up to approximately 80% of patients experiencing seizures in the first 24 hours after penetrating head injury. An in vitro model of traumatic brain injury was developed to investigate hyperexcitability and epileptogenesis and their prevention. We determined whether sodium valproate would prevent epileptiform activity in this experimental model. METHODS: Rat cortical slices were prepared and maintained in vitro using standard methods. Trauma was effected by removing the superficial 450-500 microm of slices. Traumatized slices were exposed to valproate at various time points. Intracellular and extracellular recordings were used to assess evoked activities. RESULTS: In untreated traumatized deep segments, hyperexcitability was manifested by depressed inhibition and often (54%) by epileptiform activity. Preparations exposed to valproate at 30 minutes or later after trauma showed abnormal activity similar to control traumatized slices. Epileptogenicity in deep segments was significantly reduced when slices were exposed to valproate (a) continuously immediately after trauma, (b) after a 20-minute delay, or (c) immediately after trauma for 1 hour and then returned to physiological solution. Finally, slices that were exposed to valproate or pentobarbital beginning 20 minutes after trauma for only 1 hour and then returned to physiological medium showed a significant reduction in abnormal activity. Valproate was found to enhance fast gamma-aminobutyric acid(A)-ergic inhibitory strength. CONCLUSIONS: Valproate significantly reduces epileptiform activity after trauma to the neocortex, likely by restoring the excitation-inhibition balance, perhaps through augmentation of gamma-aminobutyric acid transmission. The timing of this action may have implications for mechanisms of seizure genesis and may suggest a role for rapid treatment. PMID- 11114208 TI - Lamotrigine treatment during amygdala-kindled seizure development fails to inhibit seizures and diminishes subsequent anticonvulsant efficacy. AB - PURPOSE: Lamotrigine (LTG) is an anticonvulsant that is currently in use for the treatment of various seizure disorders and that shows promise in the treatment of affective illness. LTG is also effective in the suppression of amygdala-kindled seizures. Because many drugs show a differential efficacy profile as a function of the phase of kindling evolution, we evaluated LTG for its potential antiepileptogenic effects on the development of amygdala-kindled seizures. METHODS: In two separate studies, LTG (5 or 15 mg/kg versus vehicle) was administered before each daily amygdala stimulation (biphasic square wave pulses, 100 pulse pairs per second for a total of 0.5 second, 1-millisecond pulse width) at an intensity of 50 microA over the AD threshold. Seizure development was assessed, as well as the effect of this pretreatment on subsequent efficacy of LTG on completed kindled seizures. RESULTS: LTG at 5 mg/kg failed to block seizure development. At 15 mg/kg, LTG paradoxically enhanced seizure development and produced running fits in four of the nine animals tested. Animals previously treated with either dose of LTG during kindling development showed a diminished response to the anticonvulsant effects of LTG on fully kindled seizures compared with the vehicle-treated controls. CONCLUSIONS: Although LTG possesses potent anticonvulsant effects on completed amygdala-kindled seizures, it is either without effect (5 mg/kg) or facilitates (15 mg/kg) the initial phase of kindling development. In addition, exposure to LTG during kindled seizure development leads to a reduced subsequent response to the drug in fully kindled animals. These observations parallel those with carbamazepine and suggest that different stages of kindling (epileptogenesis versus fully manifest seizures) may have different underlying neural mechanisms that require distinct pharmacotherapies. PMID- 11114209 TI - Childhood epilepsy with occipital paroxysms: clinical variants in 134 patients. AB - PURPOSE: In its recent proposal, the Commission on Classification and Terminology of the International League Against Epilepsy classified childhood epilepsy with occipital paroxysms (CEOP) into two syndromes with different predominant seizure types: early onset (Panayiotopoulos type) with eye deviation and ictal vomiting and late onset (Gastaut type) with initial ictal visual symptoms. We documented the clinical features of a large group of patients with CEOP to confirm whether the classification is justified. METHODS: A file review of all patients with partial-onset seizure and interictal occipital spikes referred to our pediatric seizure unit between January 1975 and May 1997 yielded 134 who met the criteria for CEOP. Data were collected with a specially developed protocol and classified according to the two International League Against Epilepsy systems: (a) seizure classification, to test age-specific differences associated with the predominant seizure type, and (b) syndrome classification, to determine whether the clusters of signs and symptoms are sufficiently delineated. RESULTS: Three groups were defined according to the predominant ictal manifestations. Group 1 (visual) consisted of 24 patients (17.9%) with ictal visual symptoms; 19 (14%) of these patients also had overlapping adversive manifestations, either as a separate seizure or as part of the same event (median age at first and last seizure, 7 years 11 months and 10 years). Group 2 (adversive) consisted of 72 patients (53.7%) with tonic eye deviation (median age at first and last seizure, 5 years 2 months and 7 years 2 months). Group 3 (nonvisual, nonadversive) consisted of 38 patients (28.4%) with various seizure spread patterns (median age at first and last seizure, 6 and 7 years 2 months). Two syndromes were identified. The Gastaut type included all 24 patients in the visual group (group 1); seizures were brief and frequent and were diurnal in 83%. The Panayiotopoulos type included all 72 patients in group 2; ictal eye deviation occurred in 100% of the patients and ictal vomiting in 44%; prolonged seizures were observed in 35% and were more frequent in patients who had ictal vomiting than in those who did not (46.8% versus 25%, respectively; p < 0.027). Seizures were infrequent; 24% of patients had a single seizure and 58% had nocturnal seizures. Onset was earlier than for the Gastaut type (p < 0.002). The 38 patients with nonoccipital manifestations did not satisfy the criteria for the complete form of either syndrome. CONCLUSIONS: The most common type of CEOP, the Panayiotopoulos type, is characterized by a cluster of signs and symptoms sufficiently delineated to justify their separate classification from Gastaut-type CEOP, despite the absence of ictal vomiting in more than 50% of the patients. PMID- 11114210 TI - Distribution of seizure precipitants among epilepsy syndromes. AB - PURPOSE: Previous studies of patient-reported seizure precipitants have not evaluated whether different epilepsy syndromes are differentially affected. METHODS: Patients of a tertiary-care epilepsy center were consecutively surveyed with the use of a standardized questionnaire that lists precipitants that might trigger or exacerbate seizures (alcohol, caffeine, fasting, fatigue, fever or illness, flashing lights, heat or humidity, menstrual cycle, sleep, sleep deprivation, emotional stress, unknown, or other). Patients were classified into epilepsy syndromes according to International League Against Epilepsy criteria. Age and gender within groups defined by major precipitants were compared. Pearson's correlation was performed to evaluate common patterns of precipitants. RESULTS: Of 400 patients, 62% cited at least one precipitant. In order of frequency, stress (30%), sleep deprivation (18%), sleep (14%), fever or illness (14%), and fatigue (13%) were noted by at least 10% of patients. Stress, fatigue, and sleep deprivation positively correlated, but sleep tended to negatively correlate with other major precipitants. Rankings of precipitants varied within epilepsy syndromes, with patients with temporal lobe epilepsy citing sleep infrequently compared with patients with other epilepsy syndromes. Menstrual effects were ranked highly within major precipitants among women over age 12 and were especially noted by women with temporal lobe epilepsy (28%). CONCLUSIONS: Most patients with epilepsy identify a precipitant that triggers or exacerbates seizures. The high correlation of stress, sleep deprivation, and fatigue suggests that they act through common mechanisms to worsen seizure control. Through identification of the effect of both endogenous and exogenous precipitants among syndromes, more research and counseling can be directed to specific precipitants. PMID- 11114211 TI - Predictors of hippocampal, cerebral, and cerebellar volume reduction in childhood epilepsy. AB - PURPOSE: We examined the factors related to brain volume reduction in a pediatric sample of patients that included those with nonintractable epilepsy. METHODS: Entry criteria were children less than 18 years old with epilepsy referred for MRI, including a whole brain volumetric sequence. The sample size was 231. Risk factors were ascertained from interviews and reviews of medical records. Factors included age of onset, seizure years, family history, status epilepticus, intellectual disability, and febrile convulsions. MRI data were obtained for 44 normal childhood control subjects. RESULTS: Cerebral and cerebellar volumes were significantly associated with age, gender, moderate-to-severe intellectual disability (p < 0.001), seizure years, and status epilepticus (p < 0.03). Compared with controls, the brain volume of all patients was reduced by 10% (p < 0.001). Hippocampal volume was significantly associated with total brain volume, age (p < 0.001), focal cerebral ischemic injury, and complex febrile convulsions (p < 0.05). CONCLUSIONS: Significant brain volume reduction is present in children with epilepsy. A component of this reduction is due to acquired insults. The reduction is seen even in children with infrequent seizures over a brief time, suggesting an innate structural abnormality. When evaluating possible etiologic factors in the development of hippocampal volume reduction, one must control for total brain volume. We have confirmed the association of complex febrile convulsions with unilateral hippocampal volume reduction. PMID- 11114212 TI - Occipitoparietal epilepsy, hippocampal atrophy, and congenital developmental abnormalities. AB - PURPOSE: Diagnostic uncertainty may arise in patients with occipitoparietal epilepsy when there is neuroimaging evidence of a posterior quadrant lesion and coexistent hippocampal abnormalities ("dual pathology"). It is not known whether hippocampal atrophy (HA) in these patients results from seizure propagation to temporolimbic structures or whether it is part of the pathological process underlying the occipitoparietal epilepsy. Clarification of this issue may have a significant bearing on the management of these patients. METHODS: We studied 20 patients with occipitoparietal epilepsy and neuroimaging or pathologic evidence of a congenital developmental abnormality. Normalized hippocampal volumes were obtained in all patients. The medical records and video-EEG recordings were analyzed to correlate the MRI findings with clinical data, seizure semiology, and EEG findings. RESULTS: HA was found in seven patients (35%). Neuroimaging abnormalities concordant with the side of HA were seen in all cases. There was clinical or EEG evidence of temporal spread in 12 patients. There was no correlation between the presence of HA and temporal lobe spread. The only clinical factor associated with HA in this series was a younger age of seizure onset. CONCLUSIONS: HA in patients with occipitoparietal epilepsy due to congenital developmental abnormalities is most likely to be a marker of a more widespread process related to a common pathogenesis during prenatal or perinatal development. HA in these patients is unlikely to be the result of secondary spread from an extrahippocampal focus. Surgical treatment should be tailored toward the primary epileptogenic zone rather the site of seizure spread. PMID- 11114213 TI - Correlation of temporal lobe glucose metabolism with the Wada memory test. AB - PURPOSE: The goal of the present study was to examine the relationship of 18F fluorodeoxyglucose positron emission tomography (FDG-PET) and the Wada memory test in lateralizing memory dominance and epileptic focus. METHODS: FDG-PET and the Wada test were performed in 18 patients with temporal lobe epilepsy (TLE). The asymmetry indices of FDG-PET (PET-AI) were calculated in mesial, polar, anterolateral, midlateral, and posterolateral regions of the temporal lobe, and those of Wada memory test (Wada-AI) were obtained as well. RESULTS: The Wada-AI was significantly correlated with PET-AI in mesial (r = 0.67, p = 0.003), polar (r = 0.55, p = 0.019), anterolateral (r = 0.55, p = 0.019), and midlateral (r = 0.51, p = 0.031) regions of the temporal lobe. However, after a linear regression analysis, PET-AI of only the mesial temporal region was significantly correlated with Wada-AI (p = 0.008). Wada-AI could correctly lateralize the seizure focus in 90% of the left TLE and 75% of the right TLE patients. The PET-AI of the mesial temporal region showed the highest sensitivity of seizure lateralization (80% of left TLE and 87.5% of right TLE). PET-AI of other temporal regions had lower sensitivities (50-80% of left TLE, 20-75% of right TLE). One or two patients showed false seizure lateralization by PET-AI on each temporal region. CONCLUSIONS: Although FDG-PET hypometabolism is observed at both mesial and lateral regions of the temporal lobe in mesial TLE, mesial temporal region appeared to be a dominant and leading area for lateralizing Wada memory dominance and epileptic focus. PMID- 11114214 TI - Ratio-images calculated from interictal positron emission tomography and single photon emission computed tomography for quantification of the uncoupling of brain metabolism and perfusion in epilepsy. AB - PURPOSE: Image processing techniques were applied to interictal positron emission tomography (PET) and single-photon emission computed tomography (SPECT) brain images to aid in the localization of epileptogenic foci by calculating a functional image that represents the degree of coupling between perfusion and metabolism. Uncoupling of these two functions has been demonstrated to be a characteristic of epileptogenic tissue in temporal lobe epilepsy and has the potential to serve as a diagnostic measure for localization in other areas as well. METHODS: Interictal PET ((18)F-FDG) and interictal SPECT ((99m)Tc-HMPAO) scans were acquired from 11 epilepsy patients. The metabolism and perfusion images were three-dimensionally spatially registered, and a functional ratio image was computed. These functional maps are overlaid onto a three-dimensional rendering of the same patient's magnetic resonance imaging anatomy. RESULTS: In all patients, an average uniform perfusion-to-metabolism ratio showed approximately constant values throughout most of the whole brain. However, the epileptogenic area (confirmed on surgery) demonstrated an area of elevated perfusion/metabolism in the grey matter. CONCLUSIONS: Although hypometabolism in the PET image was observed in most of these patients, the calculation of a functional ratio-image demonstrated localized foci that in some cases could not be observed on the PET image alone. The ratio-image also yields a quantitative measure of the uncoupling phenomenon. PMID- 11114215 TI - Combining ictal surface-electroencephalography and seizure semiology improves patient lateralization in temporal lobe epilepsy. AB - PURPOSE: The study goal was to assess the concordance of ictal surface-EEG and seizure semiology data in lateralizing intractable temporal lobe epilepsy (TLE) and to examine the benefits of the combined use of these two methods. METHODS: We independently analyzed the ictal recordings and clinical symptoms associated with 262 seizures recorded in 59 TLE patients. Each seizure was lateralized on the basis of (i) its associated ictal surface-EEG pattern according to a predefined lateralization protocol and (ii) its associated ictal and postictal seizure semiology according to strictly defined clinical criteria. Individual patients were also lateralized based on these data. RESULTS: Ictal surface-EEG findings lateralized 62.6% of seizures and 64.4% of patients. Seizure semiology findings lateralized 46.2% of seizures and 78.0% of patients. There was a high degree of concordance between lateralizations based on these two methods, for both individual seizures and individual patients. Combination of the information from the two methods allowed for lateralization in a greater proportion of both seizures (79.8%) and patients (94.9%). Combined EEG-seizure lateralization was concordant with the side of operation in 33 of 34 patients who underwent successful surgery (Engel's surgical outcome class I/II). CONCLUSIONS: In TLE, there is a high agreement between the lateralization of individual seizures and patients, which is based on ictal surface-EEG findings and seizure semiology. Furthermore, combination of these two methods improves the lateralization of individual seizures and patients. Thus, standardized combined EEG-seizure analysis is a valuable noninvasive tool in the presurgical evaluation of TLE. PMID- 11114216 TI - Source reconstruction of mesial-temporal epileptiform activity: comparison of inverse techniques. AB - PURPOSE: To evaluate whether advanced source reconstruction such as current density reconstruction (CDR) provides additional hints for clinical presurgical evaluation, different source reconstruction techniques with idealized spherical as well as realistically shaped head models (boundary element method, BEM) were applied on interictal and ictal epileptiform activity in presurgical evaluated patients with temporal lobe epilepsy. It is discussed whether CDR and BEM give additional information for presurgical evaluation compared to "conventional" strategies, such as single moving, and spatio-temporal dipole modeling with spherical head models. METHODS: A variety of source reconstruction procedures were applied to the data of five patients with pharmacoresistent temporal lobe epilepsy with probable mesial origin: (1) single-moving dipole in a spherical head model and (2) in BEM, (3) spatio-temporal dipole modeling in a spherical head model and (4) in BEM; and (5) deconvolution with fixed locations and orientations and (6) with cortically constrained L1-norm CDR in BEM. In addition, simulated sources of temporal lobe origin were calculated in each subject with CDR to prove the basic feasibility of this technique in the particular application. RESULTS: Source activity was correctly localized within the affected temporal lobe by all source reconstruction techniques used. Neither single moving dipole, spatio-temporal modeling, nor CDR was able to localize sources at a sublobar level. In the case of two sources, single moving dipole solutions showed changes in dipole orientation in time and spatio-temporal modeling separated two sources, whereas CDR at the peak latency failed to distinguish among different origins. BEM enhanced localization accuracy. CONCLUSION: There was no advantage of using CDR. Single moving dipole as well as spatio-temporal dipole modeling in BEM leads to more precise localization within the individual anatomy and provides a simple algorithm, which is capable of indicating both the time course and the number of sources. PMID- 11114217 TI - Electroclinical and magnetoencephalographic analysis of epilepsy in patients with congenital bilateral perisylvian syndrome. AB - PURPOSE: Epilepsies in four patients with congenital bilateral perisylvian syndrome (CBPS) were studied electroclinically and by magnetoencephalography (MEG) to determine whether cortical dysplasia associated with this syndrome is epileptogenic. METHODS: We studied three women and one man (mean +/- SD age 30.8 +/- 3.4 years) with a diagnosis of CBPS based on magnetic resonance imaging (MRI). All had drug-resistant epilepsy. In addition to electroclinical analysis, the distribution of equivalent-current dipoles (ECDs) analyzed from MEG signals was examined. RESULTS: Patient 1 had left-sided parietal lobe epilepsy and ECDs clustered in the left temporoparietal lobe. Patient 2 had bilateral, independent temporoparietal lobe epilepsy, and ECDs were found in both hemispheres, with clustering in the right temporoparietal lobe but not in the left hemisphere. Patient 3 presumably had parietal lobe epilepsy with unidentified focus side and ECDs clustered in the left parietal and right temporal lobes. Patient 4 had symptomatic generalized epilepsy, and ECDs were observed in both hemispheres without clustering. In the three patients who had partial epilepsy (patients 1 3), clusters of ECDs overlaid the perisylvian dysplasia and were not found elsewhere. CONCLUSIONS: Patients with CBPS may have either symptomatic partial epilepsy or symptomatic generalized epilepsy, and those with partial epilepsy are likely to have temporoparietal foci. In addition, cortical dysplasia is closely related to the generation of partial seizures and may be epileptogenic per se. However, this study did not elucidate the relationship between cortical dysplasia and CBPS-generated generalized epilepsy. PMID- 11114218 TI - Long-term retention rates of lamotrigine, gabapentin, and topiramate in chronic epilepsy. AB - PURPOSE: We sought to determine the long-term retention rates of lamotrigine (LTG), gabapentin (GBP), and topiramate (TPM) therapy for patients at a tertiary referral clinic for chronic, refractory epilepsy. METHODS: We analyzed 424 consecutive patients with chronic, refractory partial and/or generalized epilepsy who were started on LTG, 158 patients who were started on GBP, and 393 patients who were started on TPM. The percentages of patients who continued therapy with LTG, GBP, and TPM were estimated with the use of Kaplan-Meier survival analysis. Factors that influence retention were analyzed with the use of Cox regression analysis. RESULTS: Kaplan-Meier survival analysis showed that at 3 years, 30% continued therapy on TPM compared with 29% on LTG and fewer than 10% on GBP. Adverse events resulted in therapy withdrawal in 40% of patients on TPM compared with GBP (37%) and LTG (22%). Perceived lack of efficacy led to treatment withdrawal in 39% of patients on GBP compared with 34% on LTG and 19% on TPM. Cox regression estimated that a fourth or fewer of patients with chronic partial epilepsy are likely to continue therapy with a new antiepileptic drug beyond 5 years. CONCLUSIONS: The impact of these new antiepileptic drugs on the long-term course of chronic partial epilepsy is likely to be small, as approximately three of four patients will discontinue therapy. More patients appear to continue on TPM compared with LTG or GBP, with a possible reason being better perceived efficacy of TPM, despite having the highest incidence of adverse events. PMID- 11114219 TI - Oxcarbazepine placebo-controlled, dose-ranging trial in refractory partial epilepsy. AB - PURPOSE: The goal of the study was to evaluate the safety and efficacy of a broad oxcarbazepine (OXC) dosage range (600, 1200, and 2400 mg/d) as adjunctive therapy for uncontrolled partial seizures and to determine the relationship between trough plasma 10-monohydroxy derivative concentrations and OXC safety and efficacy. METHODS: This multinational, multicenter, randomized, 28-week, double blind, placebo-controlled, four-arm, parallel-group trial enrolled 694 patients aged 15-65 years with uncontrolled partial seizures with or without secondarily generalized seizures. The primary efficacy variable was percentage change in seizure frequency per 28 days relative to baseline. RESULTS: The median reduction in seizure frequency was 26%, 40%, 50%, or 8% for patients receiving 600, 1200, or 2400 mg/d OXC or placebo, respectively (all p < or = 0.0001). Of patients in the 600, 1200, or 2400 mg/d OXC groups, 27%, 42%, and 50% respectively, had more than 50% reduction in seizure frequency compared with 13% for placebo (all p < 0.001). Higher plasma 10-monohydroxy derivative concentrations were associated with larger decreases in seizure frequency (p = 0.0001). During the double-blind treatment phase, 84%, 90%, 98%, and 76% of patients receiving 600, 1200, or 2400 mg/d OXC or placebo, respectively, reported one or more adverse events. The most common adverse events were related to the nervous and digestive systems. CONCLUSIONS: OXC is safe and effective as adjunctive therapy in patients with uncontrolled partial seizures. OXC 600 mg/d was the minimum effective dosage; effectiveness of OXC increased with dose. The rapid and fixed titration to high doses was associated with an increased risk of adverse events, which could potentially be reduced by adjusting concomitant antiepileptic medication and by using a slower, flexible OXC titration schedule. PMID- 11114220 TI - Psychiatric outcome after temporal lobectomy: a predictive study. AB - PURPOSE: The occurrence of psychiatric symptoms after temporal lobectomy is well documented. The aim of the present study was to identify preoperative factors that predict postoperative psychiatric outcome. METHODS: We studied the case notes of 121 patients (from an initial sample of 167) who underwent temporal lobectomy at the National Hospital of Neurology and Neurosurgery, Queen Square, London, between 1988 and 1997. Data concerning gender, laterality of lesion, pathology, seizure outcome, psychiatric history, psychiatric outcome, resection volume, telemetry, and MRI scans were systematically collected. Factors that predict the occurrence of postoperative psychiatric symptomatology were investigated using correlational, chi(2), and logistic regression techniques. RESULTS: Poor postoperative psychiatric outcome in general was positively associated with preoperative bilateral independent spike discharges at telemetry. The size of surgical resection was positively correlated with the occurrence of postoperative emotional lability. The laterality of the epileptogenic lesion was not associated with a poor psychiatric outcome. Developmental lesions were associated with a good psychiatric outcome at a marginally significant level. Patients with a preoperative psychiatric history and de novo psychiatric symptomatology had a poorer surgical outcome in terms of seizure frequency, also at a marginally significant level. A significant correlation was found between a past psychiatric history and seizure outcome. CONCLUSIONS: We identified a high frequency of psychiatric symptoms both before and after temporal lobectomy, demonstrating that it is not a benign procedure from the point of view of psychopathology. Our results show that there are certain predictive factors that may help identify patients most at risk for postoperative psychiatric disorders. PMID- 11114221 TI - Adherence to treatment in children with epilepsy: who follows "doctor's orders"? AB - PURPOSE: The goal of the present study was to examine sociocultural, medical, family environment, and individual cognitive factors that predict adherence to treatment in children with epilepsy. METHODS: The study subjects (4-13 years old) were enrolled in a longitudinal seizure study at the first visit to the seizure clinic, attended at least 6 months, and had at least two appointments. Baseline predictors, which were obtained by interview, chart review, and psychometric testing, included sociocultural and family environment, seizure and previous treatment history, child behavior, cognitive functioning (IQ), and family stress. Four latent factors tapping these indicators of risk (acculturative risk, seizure severity, behavior problems, family environment) and two measured variables (IQ and life events) were hypothesized. Outcomes were visit adherence (proportion of scheduled appointments kept, plus proportion without unscheduled contacts), medication report (proportion of visits at which parent report of medication agreed with records), and medication levels (proportion of serum anticonvulsant levels within expected range for dosage). Two-step analytic procedure included confirmatory factor analysis to validate the hypothetical structure of the baseline risk indicators, followed by structural equation modeling to examine longitudinal relations between baseline risk and subsequent adherence outcomes. RESULTS: Significant prospective relationships included acculturative risk associated positively with visit adherence and medication levels, behavior problems associated negatively with visit adherence and medication levels, family environment associated negatively with medication report, life events associated positively with medication levels and visit adherence, and cognitive functioning (IQ) associated positively with medication levels. Seizure severity was not associated significantly with any adherence outcome. There also were no significant within-time associations between adherence outcomes. CONCLUSIONS: Contrary to clinical expectations, families at higher acculturative risk and with higher life events reported greater adherence. Seizure severity did not influence adherence. The three adherence measures were statistically independent of each other. PMID- 11114222 TI - Wisconsin Card Sorting performance in patients with temporal lobe epilepsy: clinical and neuroanatomical correlates. AB - PURPOSE: A sizable proportion of patients with temporal lobe epilepsy (TLE) display impairments on tests of executive function. Previous studies have suggested several factors that may explain such performance, including the presence of hippocampal sclerosis, electrophysiological disruption to extratemporal regions, and early age of seizure onset. However, no clear determinants have been found that consistently explain such executive dysfunction. The present study investigated the contribution of several clinical variables and temporal lobe neuroanatomic features to performance on the Wisconsin Card Sorting Test (WCST) in a series of patients with TLE. METHODS: Eighty-nine patients with lateralized TLE (47 left, 42 right) were examined. Seventy-two patients from this series underwent anterior temporal lobectomy (ATL). Regression analysis was used to examine the effects of age, education, age at seizure onset, seizure duration, seizure laterality, history of secondary generalized seizures, and MRI-based volumes of the right and left hippocampi on preoperative WCST performance (number of categories completed, perseverative errors). Further univariate analyses examined whether the presence of bilateral hippocampal sclerosis, mesial temporal lobe abnormalities beyond the hippocampus, or temporal neocortical abnormalities affected preoperative WCST performance. In addition, we examined whether becoming seizure free after ATL affected change in WCST performance. RESULTS: Overall regression analysis was not significant. However, an examination of individual partial correlations revealed that patients with a history of secondary generalized seizures performed more poorly on the preoperative WCST than did patients without such history. In addition, patients who were seizure free after ATL did not exhibit better WCST outcome than patients who did not become seizure free. The presence of bilateral hippocampal sclerosis, extrahippocampal mesial temporal atrophy, or temporal neocortical lesions did not affect WCST performance. CONCLUSIONS: These results indicate that the presence of temporal lobe structural abnormalities do not significantly affect executive function as measured by the WCST. The present study does suggests that the critical determinants of WCST performance in patients with TLE lie outside the temporal lobe and likely relate to metabolic disruption to frontostriatal neural network systems. PMID- 11114223 TI - Idiopathic generalized epilepsy presenting with hemiconvulsive seizures. AB - PURPOSE: Unilateral seizures, or hemiconvulsive attacks, are motor seizures with tonic and/or clonic phenomena that involve only one side of the body. METHODS: We describe three adolescents who presented with hemiconvulsive seizures and were found to have 3-cps generalized spike-and-wave discharges on ictal and/or interictal EEG. All had normal neuroimaging studies. Two patients had been previously treated with carbamazepine, which led to a partial response in one patient. RESULTS: All three patients, however, are now seizure free on either sodium valproate or a combination of sodium valproate and lamotrigine. We believe the electroclinical diagnosis is that of idiopathic generalized epilepsy. CONCLUSIONS: Idiopathic generalized epilepsy presenting with hemiconvulsive seizures has not, to our knowledge, been previously described. However, the correct diagnosis of an idiopathic generalized seizure disorder, as opposed to a partial seizure disorder, has important treatment implications. The possible mechanism of hemiconvulsive seizures in idiopathic generalized epilepsy is discussed. PMID- 11114224 TI - Central nystagmus induced by deep-brain stimulation for epilepsy. AB - PURPOSE: The goal of the present study was to describe the localization of central nystagmus induced as a side effect of electrical deep-brain stimulation for epilepsy. METHODS: Bilateral deep-brain stimulating electrodes were inserted in the centromedian nucleus of the thalamus to control seizures in a patient with intractable epilepsy. RESULTS: Cathodal high-frequency stimulation through the deepest contact of each electrode elicited cycles of slow ipsiversive conjugate eye deviations, each followed by rapid contralateral jerks. The involved electrode contacts were situated at the mesodiencephalic junction just inferior to the centromedian nucleus of the thalamus and rostral to the superior colliculus. Right-sided stimulation evoked left beating nystagmus and left-sided stimulation evoked right beating nystagmus. Stimulation through other electrode contacts did not induce nystagmus. Electronystagmography showed the nystagmus to have constant velocity slow phases. CONCLUSIONS: A central nystagmogenic area exists in humans that appears to be homologous to the nucleus of the optic tract, a region described in nonhuman primates to play a role in the generation of optokinetic nystagmus. PMID- 11114225 TI - Renal tubular acidosis associated with zonisamide therapy. AB - PURPOSE: We sought to report a previously undescribed adverse effect, renal tubular acidosis associated with zonisamide (ZNS) therapy. METHODS: Ammonium chloride, bicarbonate, and furosemide loading tests were performed in an epileptic patient with metabolic acidosis and episodic hypokalemia who was treated with ZNS. RESULTS: Distal renal tubular acidosis was diagnosed. On reexamination 7 weeks after ZNS had been replaced with phenytoin, the renal tubular acidosis disappeared. CONCLUSIONS: This case indicates, for the first time, that ZNS might be a potential cause of renal tubular acidosis. Blood gases and serum electrolytes should be measured in patients undergoing ZNS therapy. PMID- 11114226 TI - Dynamic, stochastic, and topological aspects of polyrhythmic performance. AB - Previous research on polyrhythmic performance can be broadly summarized in terms of 2 classes of models: timekeeper models and nonlinear dynamical models. In the former approach, research has been focused on patterns of covariance among time intervals, and in the latter approach, the concentration has been on pattern (in)stability and the spatiotemporal properties of oscillating limbs. It is suggested that one can achieve a more comprehensive theory that incorporates the strengths of each of these approaches by endowing timekeeper models with nonlinear dynamics or by endowing nonlinear oscillator models with stochastic variability. Additionally, those models are complemented by a topological description of performance based on knot theory. Knot theory provides a new index of difficulty for polyrhythmic tapping, a spatial interpretation of transitions between different stable rhythms, and a possible instantiation of N. A. Bernstein's (1967a) notion of a topological motor program. PMID- 11114227 TI - Compensatory coordination of release parameters in a throwing task. AB - The consistency and coordination of release parameters in ball-throwing movements were investigated. The authors used a newly developed index of coordination for release parameters (ICRP) that quantifies the degree of improvement of performance consistency caused by compensatory relationships among parameters (i.e., not caused by consistency of parameters). Eight participants practiced for 150 trials, with the nondominant hand, a ball-throwing task aimed at a stationary target. The magnitude of the ball-release velocity vector, among release parameters, as well as the performance was found to become consistent with practice. The ICRP score suggested that the release parameters were complementarily coordinated with one another, and that the coordination improved with practice. Those results indicate that compensatory relationships among varying release parameters contribute to reducing the variability of performance in a ball-throwing task whose goal is accuracy. PMID- 11114228 TI - Monocular and binocular vision in the control of goal-directed movement. AB - In the present research the authors examined the time course of binocular integration in goal-directed aiming and grasping. With liquid-crystal goggles, the authors manipulated vision independently to the right and left eyes of 10 students during movement preparation and movement execution. Contrary to earlier findings reported in catching experiments (I. Olivier, D. J. Weeks, K. L. Ricker, J. Lyons, & D. Elliott, 1998), neither a temporal nor a spatial binocular advantage was obtained in 1 grasping and 2 aiming studies. That result suggests that, at least in some circumstances, monocular vision is sufficient for the precise control of limb movements. In a final aiming experiment involving 3 dimensional spatial variability and no trial-to-trial visual feedback about performance, binocular vision was associated with greater spatial accuracy. Binocular superiority appeared to be most pronounced when participants were unable to adjust their limb control strategy or procedure on the basis of terminal feedback about performance. PMID- 11114229 TI - A dynamical systems approach to manual tracking performance. AB - Manual tracking performance was investigated from the perspective of dynamical systems theory. The authors manipulated the type of visual display, the control system dynamics, and the frequency of the sinusoidal input signal to examine couplings with various phases between the visual signal and control movements. Analyses of the system output amplitude ratio and relative phase showed that participants (N = 24) performed poorly with 90 degrees relative phase coupling. All the couplings became less stable as the movement frequency increased. The authors developed an adaptive oscillator model with linear damping to describe the coupled system consisting of the human performer, the visual display, and the control system dynamics. A geometric account of the stability of performance at different relative phases is also presented. PMID- 11114230 TI - Trunk-assisted prehension: specification of body segments with imposed temporal constraints. AB - The authors used a trunk-assisted prehension task to examine intersegment coordination. Participants (N = 7) reached to grasp an object placed beyond full arm extension, thus requiring trunk flexion to achieve the target object, under 4 varying temporal constraints. Kinematic analyses were performed in which the motions of the arm, the trunk, and the endpoint were characterized. The spatial trajectories and the segments' peak velocity data revealed that under high temporal constraints the arm was more responsible for endpoint motion than the trunk, whereas in the unconstrained condition the trunk was more involved. In addition, the arm exhibited a decline in spatial variability toward the end of the movement in all conditions, whereas the trunk did not. The present study is the first to show that when temporal demand is increased for a trunk-assisted prehensile task, the arm plays a larger role than the trunk in the transport of the hand to the object. The data also suggest that the arm participates in the fine accuracy control of the reach, whereas the trunk does not. PMID- 11114231 TI - Effect of age and gender in the control of elbow flexion movements. AB - In previous studies of rapid elbow movements in young healthy men, characteristic task-dependent changes in the patterns of muscle activation when movement speed or distance was varied have been reported. In the present study, the authors investigated whether age or gender is associated with changes in the patterns of muscle activity previously reported in young men. Arm movements of 10 healthy older and 10 healthy younger participants (5 men and 5 women in each group) were studied. Surface electromyograms (EMGs) from agonist (biceps) and antagonist (triceps) muscles, kinematic and kinetic parameters, as well as anthropometric and strength measures were recorded. All 4 groups of participants showed similar task- (distance or speed) dependent changes in biphasic EMG activity. Similar modulation of the initial rate of rise of the EMG, integrated agonist and antagonist EMG activity, as well as their relative timing were observed in all 4 groups. Those results suggest that older individuals of both genders retain the control strategies for elbow movements used by young individuals. Despite the qualitative similarities in the patterns of muscle activation, the men moved more quickly than the women, and younger participants moved more quickly than older participants. Those performance differences could not be explained in terms of differences in body size and strength alone. PMID- 11114232 TI - The visuomotor system resists the horizontal-vertical illusion. AB - The effect of the horizontal-vertical illusion on the visual and visuomotor systems was investigated. Participants (N = 8) viewed horizontal and vertical lines in an inverted-T stimulus and judged whether the two line segments were the same or different lengths. Participants also reached out and grasped either the vertical or the horizontal line segment of the stimulus. Perceptually, participants succumbed to the illusion; that is, they judged Ts of equal horizontal and vertical line lengths to be different and Ts of unequal line lengths to be the same. When reaching toward the same stimuli, however, the size of their grip aperture was scaled appropriately for the various line lengths. Thus, whereas the perceptual system succumbed to the illusion, the visuomotor system did not. Those results support a model proposed by M. A. Goodale and A. D. Milner (1992), who posited separate cortical pathways for visual perception and visually guided action. PMID- 11114233 TI - Center of gravity motions and ankle joint stiffness control in upright undisturbed stance modeled through a fractional Brownian motion framework. AB - The authors modeled the center of gravity vertical projection (CG(v)) and the difference, CP - CG(v), which, combined, constitute the center of pressure (CP) trajectory, as fractional Brownian motion in order to investigate their relative contributions and their spatiotemporal articulation. The results demonstrated that CG(v) and CP - CG(v) motions are both endowed in complementary fashion with strong stochastic and part-deterministic behaviors. In addition, if the temporal coordinates remain similar for all 3 trajectories by definition, the switch between the successive control mechanisms appears for shorter displacements for CP - CG(v) and CG(v) than for CP trajectories. Results deduced from both input (CG(v)) and muscular stiffness (CP - CG(v)) thus provide insight into the way the central nervous system regulates stance control and in particular how CG and CP - CG are controlled. PMID- 11114234 TI - Protein targeting to flagella of trypanosomatid protozoa. PMID- 11114235 TI - Effects of the myosin inhibitor 2,3-butanedione monoxime (BDM) on cell shape, locomotion and fluid pinocytosis in human polymorphonuclear leucocytes. AB - We investigated the role of myosin in polymorphonuclear leucocyte (PMN) shape changes, locomotion, and fluid pinocytosis using the myosin inhibitor 2,3 butanedione monoxime (BDM). Treatment of resting spherical PMNs with BDM produced spheroid cells showing small continuous shape changes (IC(50)=15.5 m m BDM) and occasionally small blebs. Cell polarity, as induced by the chemotactic peptide fNLPNTL or by colchicine, and locomotion were completely suppressed (IC(50)=8.4 to 10 m m). Suppression of fNLPNTL- or colchicine-induced cell polarity produced spheroid cells, suppression of PMA-induced shape changes and fluid pinocytosis produced non-motile spherical cells (IC(50)=25 to 30 m m BDM). BDM suppressed formation of lamellipodia but not formation of blebs. Suppression of microvilli by BDM as observed in resting spherical cells was partially antagonized by PMA. The results suggest that myosin is involved in stabilizing the shape of resting spherical cells, including microvilli, and that myosin is required for cell polarity, locomotion, fluid pinocytosis and for formation of lamellipodia, but not for formation of blebs. PMID- 11114236 TI - Changes in polyamine metabolism during glucocorticoid-induced programmed cell death in mouse thymus. AB - When mice are injected with dexamethasone, cortical thymocytes are deleted through programmed cell death (PCD). We have used this in vivo model system to investigate the kinetics of PCD and cell proliferation in relation to polyamine metabolism for 16 h after injection of dexamethasone. As a marker for PCD, we used the appearance of a sub-G(1)peak in the DNA histogram. When a sub-G(1)peak appeared at 4 h after dexamethasone treatment, the activity of the polyamine catabolic enzyme spermidine/spermine N(1)-acetyltransferase (SSAT) was significantly increased and the activity of the polyamine biosynthetic enzyme S adenosylmethionine decarboxylase (AdoMetDC) was significantly decreased compared to the activities found in the thymi of control mice. Despite the significant changes in the activities of SSAT and AdoMetDC, the only change in the polyamine pool during the experimental period was that of putrescine. Presumably the complexity of this in vivo system masks changes in the spermidine and spermine pools that were expected in relation to the increased SSAT activity and decreased AdoMetDC activity. PMID- 11114237 TI - Changes in the expression of small intestine extracellular matrix proteins in streptozotocin-induced diabetic rats. AB - Diabetes mellitus is characterized by anatomical and functional alterations of the intestinal tract. However, the aetiology of these disturbances remains unclear. The aim of the present work was to investigate the effects of diabetes on the expression of laminin-1 and fibronectin in the small intestine of Streptozotocin (STZ)-induced diabetic rats. The Western immunoblotting of the extracts from the small intestine revealed that experimental diabetes resulted in a marked increase in the intensity of the bands corresponding to laminin-1 and fibronectin. Immunohistochemical studies demonstrated a strong labelling to these two extracellular matrix (ECM) proteins in the small intestine of diabetic rats, mainly localized in the smooth muscle layer. These results occur together with a thickening of the basement membrane (BM) of the smooth muscle cells, demonstrated by transmission electron microscopy (TEM). We propose that the accumulation of ECM proteins in the smooth muscle layer may be an effect mediated by hyperglycaemia, since insulin treatment of diabetic rats reversed this accumulation. These results could provide information on the potential role of the ECM in the intestine, an organ which is known to exhibit important alterations in diabetes. PMID- 11114238 TI - Recombinant human interleukin-8, but not human interleukin-1beta, induces bovine neutrophil migration in an in vitro co-culture system. AB - A co-culture system was established by culturing a bovine mammary epithelial cell line (MAC-T) and a bovine aortic endothelial cell line on calf tail collagen pre coated inserts. This system allowed us to study bovine neutrophil migration across endothelium, extracellular matrix (ECM), and epithelium in the correct sequence and direction in vitro. The effect of recombinant interleukin-1beta (rHIL-1beta) and interleukin-8 (rHIL-8) on bovine neutrophil migration was investigated using this system. rHIL-8 stimulated bovine neutrophil migration in a dose-dependent fashion. The level of migrating bovine neutrophils increased up to approximately 25% when 100 ng/ml of rHIL-8 was used. On the other hand, rHIL 1beta at concentrations up to 100 ng/ml did not directly induce bovine neutrophil migration. Furthermore, pre-incubation with 5 ng/ml of rHIL-1beta in the co culture system for 4 or 24 h failed to have any effect. These results suggest that IL-8 plays an important role in neutrophil migration into bovine mammary glands during mastitis. PMID- 11114239 TI - Changes in liver gangliosides in streptozotocin-induced diabetic rats. AB - Diabetes mellitus is associated with various structural and functional liver abnormalities that affect the glycogen and lipid metabolisms. The effects of streptozotocin-induced diabetes and of insulin supplementation to Sprague-Dawley diabetic rats on ganglioside patterns in liver were determined. Diabetic livers showed a tendency to hepatomegaly 3 weeks after STZ-induction of diabetes. The concentration of total gangliosides in diabetic and non-diabetic livers was similar, but the concentration of total gangliosides in the liver of insulin stabilized rats was slightly increased. Bidimensional TLC chromatographic analysis of gangliosides isolated from normal diabetic and insulin-stabilized diabetic livers showed quantitative and qualitative changes. In comparison with normal controls, the densitometric analyses of diabetic liver ganglioside patterns had increased amounts of GM3, GM1, GD1b, and GT1b gangliosides, while GM2 could not be detected. The hepatic ganglioside pattern of insulin-stabilized diabetic rats was partially restored, resembling the profile of normal rats. The activity of GalNAcT, GalT-2 and SialT-4 transferases was measured in liver microsomal fractions of the different groups of animals. Diabetic rats showed an increased activity of GalNAcT and a decrease in the activity of GalT-2 and SialT 4 compared with the controls. The enzymatic activities found in insulin-treated rats showed a tendency to return to the values observed in normal control animals. The results evidenced that streptozotocin-induced diabetes affects the liver ganglioside pattern and the ganglioside synthesis enzyme activity. The alterations found in ganglioside metabolism could represent one of the earliest changes associated with the diabetic pathology. PMID- 11114241 TI - INDEX AND VOLUME CONTENTS: VOLUME 24, NOS. 1-12 2000. PMID- 11114240 TI - Automated detection of morphological alterations during apoptosis. AB - A convenient means of measuring apoptosis is described using an automated analyzer (MF-1; Sysmex), which normally provides rapid measurement of malarial parasites. By means of this MF-1 procedure, apoptotic cells exhibited characteristic changes of light scatter (size) and fluorescence (DNA content) relating to cell shrinkage and nuclear fragmentation of apoptosis. PMID- 11114242 TI - The subunit 1 of the COP9 signalosome suppresses gene expression through its N terminal domain and incorporates into the complex through the PCI domain. AB - The COP9 signalosome is a conserved protein complex composed of eight subunits. Individual subunits of the complex have been linked to various signal transduction pathways leading to gene expression and cell cycle control. However, it is not understood how each subunit executes these activities as part of a large protein complex. In this study, we dissected structure and function of the subunit 1 (CSN1 or GPS1) of the COP9 signalosome relative to the complex. We demonstrated that the C-terminal half of CSN1 encompassing the PCI domain is responsible for interaction with CSN2, CSN3, and CSN4 subunits and is required for incorporation of the subunit into the complex. The N-terminal fragment of CSN1 cannot stably associate with the complex but can translocate to the nucleus on its own. We further show that CSN1 or the N-terminal fragment of CSN1 (CSN1-N) can inhibit c-fos expression from either a transfected template or a chromosomal transgene ( fos-lacZ). Moreover, CSN1 as well as CSN1-N can potently suppress signal activation of a AP-1 promoter and moderately suppress serum activation of a SRE promoter, but is unable to inhibit PKA-induced CRE promoter activity. We conclude that the N-terminal half of CSN1 harbors the activity domain that confers most of the repression functions of CSN1 while the C-terminal half allows integration of the protein into the COP9 signalosome. PMID- 11114243 TI - Concerted kinetic folding of a multidomain ribozyme with a disrupted loop receptor interaction. AB - The free energy landscape for the folding of large, multidomain RNAs is rugged, and kinetically trapped, misfolded intermediates are a hallmark of RNA folding reactions. Here, we examine the role of a native loop-receptor interaction in determining the ruggedness of the energy landscape for folding of the Tetrahymena ribozyme. We demonstrate a progressive smoothing of the energy landscape for ribozyme folding as the strength of the loop-receptor interaction is reduced. Remarkably, with the most severe mutation, global folding is more rapid than for the wild-type ribozyme and proceeds in a concerted fashion without the accumulation of long-lived kinetic intermediates. The results demonstrate that a complex interplay between native tertiary interactions, divalent ion concentration, and non-native secondary structure determines the ruggedness of the energy landscape. Furthermore, the results suggest that kinetic folding transitions involving large regions of highly structured RNAs can proceed in a concerted fashion, in the absence of significant stable, preorganized tertiary structure. PMID- 11114244 TI - The efficiency of strand invasion by Escherichia coli RecA is dependent upon the length and polarity of ssDNA tails. AB - RecA protein is essential for homologous recombination and the repair of DNA double-strand breaks in Escherichia coli. The protein binds DNA to form nucleoprotein filaments that promote joint molecule formation and strand exchange in vitro. RecA polymerises on ssDNA in the 5'-3' direction and catalyses strand exchange and branch migration with a 5'-3' polarity. It has been reported previously, using D-loop assays, in which ssDNA (containing a heterologous block at one end) invades supercoiled duplex DNA that 3'-homologous ends are reactive, whereas 5'-ends are inactive. This polarity bias was thought to be due to the polarity of RecA filament formation, which results in the 3'-ends being coated in RecA, whereas 5'-ends remain naked. Using a range of duplex substrates containing ssDNA tails of various lengths and polarities, we now demonstrate that when no heterologous block is imposed, 5'-ends are just as reactive as 3'-ends. Moreover, using short-tailed substrates, we find that 5'-ends form more stable D-loops than 3'-ends. This bias may be a consequence of the instability of short 3'-joints. With more physiological substrates containing long ssDNA tails, we find that RecA shows no intrinsic preference for 5' or 3'-ends and that both form D-loop complexes with high efficiency. PMID- 11114245 TI - Structural rearrangements and insertions of dispersed elements in pericentromeric alpha satellites occur preferably at kinkable DNA sites. AB - Centromeric region of human chromosome 21 comprises two long alphoid DNA arrays: the well homogenized and CENP-B box-rich alpha21-I and the alpha21-II, containing a set of less homogenized and CENP-B box-poor subfamilies located closer to the short arm of the chromosome. Continuous alphoid fragment of 100 monomers bordering the non-satellite sequences in human chromosome 21 was mapped to the pericentromeric short arm region by fluorescence in situ hybridization (alpha21 II locus). The alphoid sequence contained several rearrangements including five large deletions within monomers and insertions of three truncated L1 elements. No binding sites for centromeric protein CENP-B were found. We analyzed sequences with alphoid/non-alphoid junctions selectively screened from current databases and revealed various rearrangements disrupting the regular tandem alphoid structure, namely, deletions, duplications, inversions, expansions of short oligonucleotide motifs and insertions of different dispersed elements. The detailed analysis of more than 1100 alphoid monomers from junction regions showed that the vast majority of structural alterations and joinings with non-alphoid DNAs occur in alpha satellite families lacking CENP-B boxes. Most analyzed events were found in sequences located toward the edges of the centromeric alphoid arrays. Different dispersed elements were inserted into alphoid DNA at kinkable dinucleotides (TG, CA or TA) situated between pyrimidine/purine tracks. DNA rearrangements resulting from different processes such as recombination and replication occur at kinkable DNA sites alike insertions but irrespectively of the occurrence of pyrimidine/purine tracks. It seems that kinkable dinucleotides TG, CA and TA are part of recognition signals for many proteins involved in recombination, replication, and insertional events. Alphoid DNA is a good model for studying these processes. PMID- 11114246 TI - Recognition of the lipoyl domain is the ultimate determinant of substrate channelling in the pyruvate dehydrogenase multienzyme complex. AB - Reductive acetylation of the lipoyl domain (E2plip) of the dihydrolipoyl acetyltransferase component of the pyruvate dehydrogenase multienzyme complex of Escherichia coli is catalysed specifically by its partner pyruvate decarboxylase (E1p), and no productive interaction occurs with the analogous 2-oxoglutarate decarboxylase (E1o) of the 2-oxoglutarate dehydrogenase complex. Residues in the lipoyl-lysine beta-turn region of the unlipoylated E2plip domain (E2plip(apo)) undergo significant changes in both chemical shift and transverse relaxation time (T(2)) in the presence of E1p but not E1o. Residue Gly11, in a prominent surface loop between beta-strands 1 and 2 in the E2plip domain, was also observed to undergo a significant change in chemical shift. Addition of pyruvate to the mixture of E2plip(apo) and E1p caused larger changes in chemical shift and the appearance of multiple cross-peaks for certain residues, suggesting that the domain was experiencing more than one type of interaction. Residues in both beta strands 4 and 5, together with those in the prominent surface loop and the following beta-strand 2, appeared to be interacting with E1p, as did a small patch of residues centred around Glu31. The values of T(2) across the polypeptide chain backbone were also lower than in the presence of E1p alone, suggesting that E2plip(apo) binds more tightly after the addition of pyruvate. The lipoylated domain (E2plip(holo)) also exhibited significant changes in chemical shift and decreases in the overall T(2) relaxation times in the presence of E1p, the residues principally affected being restricted to the half of the domain that contains the lipoyl-lysine (Lys41) residue. In addition, small chemical shift changes and a general drop in T(2) times in the presence of E1o were observed, indicating that E2plip(holo) can interact, weakly but non-productively, with E1o. It is evident that recognition of the protein domain is the ultimate determinant of whether reductive acetylation of the lipoyl group occurs, and that this is ensured by a mosaic of interactions with the Elp. PMID- 11114247 TI - Electron microscopic analysis supports a dual role for the mitochondrial telomere binding protein of Candida parapsilosis. AB - Linear mitochondrial genomes exist in several yeast species which are closely related to yeast that harbor circular mitochondrial genomes. Several lines of evidence suggest that the conversion from one form to another occurred accidentally through a relatively simple mechanism. Previously, we (L.T. & J.N.) reported the identification of the first mitochondrial telomere-binding protein (mtTBP) that specifically binds a sequence derived from the extreme end of Candida parapsilosis linear mtDNA, and sequence analysis of the corresponding nuclear gene MTP1 revealed that mtTBP shares homology with several bacterial and mitochondrial single-stranded (ss) DNA-binding (SSB) proteins. In this study, the DNA-binding properties of mtTBP in vitro and in vivo were analyzed by electron microscopy (EM). When M13 ssDNA was used as a substrate, mtTBP exhibited similar DNA binding characteristics as human mitochondrial SSB: mtTBP formed protein globules along the DNA substrate, and the bound proteins were randomly distributed, indicating that the binding of mtTBP to M13 ssDNA is not highly cooperative. EM analysis demonstrated that mtTBP is able to recognize the 5' single-stranded telomeric overhangs in their natural context. Using isopycnic centrifugation of mitochondrial lysates of C. papsilosis we show that mtTBP is a structural part of mitochondrial nucleoids of C. parapsilosis and is predominantly bound to the mitochondrial telomeres. These data support a dual role of mtTBP in mitochondria of C. parapsilosis, serving both as a typical mitochondrial SSB and as a specific component of the mitochondrial telomeric chromatin. PMID- 11114248 TI - Projection structure of the monomeric porin OmpG at 6 A resolution. AB - The Escherichia coli porin OmpG, which acts as an efficient unspecific channel for mono-, di- and trisaccharides, has been purified and crystallized in two dimensions. Projection maps of two different crystal forms of OmpG at 6 A resolution show that the protein has a beta-barrel structure characteristic for outer membrane proteins, and that it does not form trimers, unlike most other porins such as OmpF and OmpC, but appears in monomeric form. The size of the barrel is approximately 2.5 nm, indicating that OmpG may consist of 14 beta strands. The projection map suggests that the channel is restricted by internal loops. PMID- 11114249 TI - Structural basis for enantiomer binding and separation of a common beta-blocker: crystal structure of cellobiohydrolase Cel7A with bound (S)-propranolol at 1.9 A resolution. AB - Cellobiohydrolase Cel7A (previously called CBH 1), the major cellulase produced by the mould fungus Trichoderma reesei, has been successfully exploited as a chiral selector for separation of stereo-isomers of some important pharmaceutical compounds, e.g. adrenergic beta-blockers. Previous investigations, including experiments with catalytically deficient mutants of Cel7A, point unanimously to the active site as being responsible for discrimination of enantiomers. In this work the structural basis for enantioselectivity of basic drugs by Cel7A has been studied by X-ray crystallography. The catalytic domain of Cel7A was co crystallised with the (S)-enantiomer of a common beta-blocker, propranolol, at pH 7, and the structure of the complex was determined and refined at 1. 9 A resolution. Indeed, (S)-propranolol binds at the active site, in glucosyl-binding subsites -1/+1. The catalytic residues Glu212 and Glu217 make tight salt links with the secondary amino group of (S)-propranolol. The oxygen atom attached to the chiral centre of (S)-propranolol forms hydrogen bonds to the nucleophile Glu212 O(epsilon1) and to Gln175 N(epsilon2), whereas the aromatic naphthyl moiety stacks with the indole ring of Trp376 in site +1. The bidentate charge interaction with the catalytic glutamate residues is apparently crucial, since no enantioselectivity has been obtained with the catalytically deficient mutants E212Q and E217Q. Activity inhibition experiments with wild-type Cel7A were performed in conditions close to those used for crystallisation. Competitive inhibition constants for (R)- and (S)-propranolol were determined at 220 microM and 44 microM, respectively, corresponding to binding free energies of 20 kJ/mol and 24 kJ/mol, respectively. The K(i) value for (R)-propranolol was 57-fold lower than the highest concentration, 12.5 mM, used in co-crystallisation experiments. Still several attempts to obtain a complex with the (R)-enantiomer have failed. By using cellobiose as a selective competing ligand, the retention of the enantiomers of propranolol on the chiral stationary phase (CSP) based on Cel7A mutant D214N were resolved into enantioselective and non- selective binding. The enantioselective binding was weaker for both enantiomers on D214N-CSP than on wild-type-CSP. PMID- 11114250 TI - Crystal structure and novel recognition motif of rho ADP-ribosylating C3 exoenzyme from Clostridium botulinum: structural insights for recognition specificity and catalysis. AB - Clostridium botulinum C3 exoenzyme inactivates the small GTP-binding protein family Rho by ADP-ribosylating asparagine 41, which depolymerizes the actin cytoskeleton. C3 thus represents a major family of the bacterial toxins that transfer the ADP-ribose moiety of NAD to specific amino acids in acceptor proteins to modify key biological activities in eukaryotic cells, including protein synthesis, differentiation, transformation, and intracellular signaling. The 1.7 A resolution C3 exoenzyme structure establishes the conserved features of the core NAD-binding beta-sandwich fold with other ADP-ribosylating toxins despite little sequence conservation. Importantly, the central core of the C3 exoenzyme structure is distinguished by the absence of an active site loop observed in many other ADP-ribosylating toxins. Unlike the ADP-ribosylating toxins that possess the active site loop near the central core, the C3 exoenzyme replaces the active site loop with an alpha-helix, alpha3. Moreover, structural and sequence similarities with the catalytic domain of vegetative insecticidal protein 2 (VIP2), an actin ADP-ribosyltransferase, unexpectedly implicates two adjacent, protruding turns, which join beta5 and beta6 of the toxin core fold, as a novel recognition specificity motif for this newly defined toxin family. Turn 1 evidently positions the solvent-exposed, aromatic side-chain of Phe209 to interact with the hydrophobic region of Rho adjacent to its GTP-binding site. Turn 2 evidently both places the Gln212 side-chain for hydrogen bonding to recognize Rho Asn41 for nucleophilic attack on the anomeric carbon of NAD ribose and holds the key Glu214 catalytic side-chain in the adjacent catalytic pocket. This proposed bipartite ADP-ribosylating toxin turn-turn (ARTT) motif places the VIP2 and C3 toxin classes into a single ARTT family characterized by analogous target protein recognition via turn 1 aromatic and turn 2 hydrogen-bonding side chain moieties. Turn 2 centrally anchors the catalytic Glu214 within the ARTT motif, and furthermore distinguishes the C3 toxin class by a conserved turn 2 Gln and the VIP2 binary toxin class by a conserved turn 2 Glu for appropriate target side-chain hydrogen-bonding recognition. Taken together, these structural results provide a molecular basis for understanding the coupled activity and recognition specificity for C3 and for the newly defined ARTT toxin family, which acts in the depolymerization of the actin cytoskeleton. This beta5 to beta6 region of the toxin fold represents an experimentally testable and potentially general recognition motif region for other ADP-ribosylating toxins that have a similar beta-structure framework. PMID- 11114251 TI - NMR structure of Streptomyces killer toxin-like protein, SKLP: further evidence for the wide distribution of single-domain betagamma-crystallin superfamily proteins. AB - A protein isolated from the culture supernatant of the soil bacterium, Streptomyces sp. F-287, exhibits cytocidal effects for both budding and fission yeasts, and causes morphological changes of yeasts and filamentous fungi. This protein, which was the first killer toxin-like protein for yeasts identified in the Streptomyces microorganism, was named SKLP (Streptomyces killer toxin-like protein). Since the amino acid sequence of the protein, as determined by sequential Edman degradations, seemed to be unique, we determined the structure by NMR spectroscopy. Although the actual target of SKLP in yeasts has not been determined yet, the structure might give us a clue to characterize the targets. The solution structure of SKLP determined by NMR, however, turned out to be a single-domain crystallin-like protein, with two Greek key motifs and a short extra beta-strand at the N terminus. The final ensemble of 20 NMR structures overlaid onto their mean coordinate with rmsd values of 0.32(+/-0.06) A for the backbone atoms involved in the secondary structure elements. As a yeast killer toxin, WmKT, isolated from the yeast strain Williopsis mrakii also has a Greek key beta-barrel fold, we have made a detailed comparison of the structural features of SKLP with the other crystallin superfamily proteins. It is very interesting that SKLP has a unique electrostatic potential distribution on the molecular surface. Namely, one surface of the beta-barrel fold in SKLP has a large negatively charged region, with an isolated positive charge of the Arg62 side-chain at the center. The edge of this surface is surrounded by positively charged residues, including Arg31, Arg65 and Arg74. The salient features of the charge distribution on this surface and the cluster of Arg residues might be related to the target binding of SKLP. PMID- 11114252 TI - Structure-activity relationships in flexible protein domains: regulation of rho GTPases by RhoGDI and D4 GDI. AB - The guanine dissociation inhibitors RhoGDI and D4GDI inhibit guanosine 5' diphosphate dissociation from Rho GTPases, keeping these small GTPases in an inactive state. The GDIs are made up of two domains: a flexible N-terminal domain of about 70 amino acid residues and a folded 134-residue C-terminal domain. Here, we characterize the conformation of the N-terminal regions of both RhoGDI and D4GDI using a series of NMR experiments which include (15)N relaxation and amide solvent accessibility measurements. In each protein, two regions with tendencies to form helices are identified: residues 36 to 58 and 9 to 20 in RhoGDI, and residues 36 to 57 and 20 to 25 in D4GDI. To examine the functional roles of the N terminal domain of RhoGDI, in vitro and in vivo functional assays have been carried out with N-terminally truncated proteins. These studies show that the first 30 amino acid residues are not required for inhibition of GDP dissociation but appear to be important for GTP hydrolysis, whilst removal of the first 41 residues completely abolish the ability of RhoGDI to inhibit GDP dissociation. The combination of structural and functional studies allows us to explain why RhoGDI and D4GDI are able to interact in similar ways with the guanosine 5' diphosphate-bound GTPase, but differ in their ability to regulate GTP-bound forms; these functional differences are attributed to the conformational differences of the N-terminal domains of the guanosine 5'-diphosphate dissociation inhibitors. Therefore, the two transient helices, appear to be associated with different biological effects of RhoGDI, providing a clear example of structure-activity relationships in a flexible protein domain. PMID- 11114253 TI - Membrane binding motif of the P-type cardiotoxin. AB - Carditoxins (CTXs) from cobra snake venoms, the basic 60-62 residue all-beta sheet polypeptides, are known to bind to and impair the function of cell membranes. To assess the membrane induced conformation and orientation of CTXs, the interaction of the P-type cardiotoxin II from Naja oxiana snake venom (CTII) with perdeuterated dodecylphosphocholine (DPC) was studied using ( 1 )H-NMR spectroscopy and diffusion measurements. Under conditions where the toxin formed a well-defined complex with DPC, the spatial structure of CTII with respect to the presence of tightly bound water molecules in loop II, was calculated using the torsion angle dynamics program DYANA. The structure was found to be similar, except for subtle changes in the tips of all three loops, to the previously described "major" form of CTII in aqueous solution illustrated by the "trans" configuration of the Val7-Pro8 peptide bond. No "minor" form with the "cis" configuration of the above bond was found in the micelle-bound state. The broadening of the CTII backbone proton signals by 5, 16-doxylstearate relaxation probes, together with modeling based on the spatial structure of CTII, indicated a periphery mode of binding of the toxin molecule to the micelle and revealed its micelle interacting domain. The latter includes a hydrophobic region of CTII within the extremities of loops I and III (residues 5-11, 46-50), the basement of loop II (residues 24-29,31-37) and the belt of polar residues encircling these loops (lysines 4,5,12,23,50, serines 11,46, histidine 31, arginine 36). It is suggested that this structural motif and the mode of binding can be realized during interaction of CTXs with lipid and biological membranes. PMID- 11114255 TI - Localization of the protein L2 in the 50 S subunit and the 70 S E. coli ribosome. AB - The protein L2 is found in all ribosomes and is one of the best conserved proteins of this mega-dalton complex. The protein was localized within both the isolated 50 S subunit and the 70 S ribosome of the Escherichia coli bacteria with the neutron-scattering technique of spin-contrast variation. L2 is elongated, exposing one end of the protein to the surface of the intersubunit interface of the 50 S subunit. The protein changes its conformation slightly when the 50 S subunit reassociates with the 30 S subunit to form a 70 S ribosome, becoming more elongated and moving approximately 30 A into the 50 S matrix. The results support a recent observation that L2 is essential for the association of the ribosomal subunits and might participate in the binding and translocation of the tRNAs. PMID- 11114254 TI - Folding pathway mediated by an intramolecular chaperone: the structural and functional characterization of the aqualysin I propeptide. AB - Aqualysin I, a thermostable homologue of subtilisin, requires its propeptide (ProA) to function as an intramolecular chaperone (IMC). To decipher the mechanisms through which propeptides can initiate protein folding, we characterized ProA in terms of its sequence, structure and function. Our results show that, in contrast to ProS (propeptide of subtilisin), ProA can fold spontaneously, reversibly and cooperatively into a stable monomeric alpha-beta conformation, even when isolated from its cognate protease-domain. ProA displays an indiscernible amount of tertiary structure with a considerable solvent accessible hydrophobic surface, but is not a classical molten-globule folding intermediate. Moreover, despite showing only 21 % sequence identity with ProS, ProA can not only inhibit enzymatic activity with a magnitude tenfold greater than ProS, but can also chaperone subtilisin folding, albeit with a lower efficiency. The structure of ProA complexed with subtilisin is different from that of isolated ProA. Hence, additional interactions seem necessary to induce ProA into a compact structure. Our results also suggest that: (a) propeptides that are potent inhibitors are not necessarily better IMCs; (b) propeptides within the subtilase family appear polymorphic and; (c) the intrinsic instability within propeptides may be necessary for rapid activation of the cognate protein. PMID- 11114256 TI - PMP22 carrying the trembler or trembler-J mutation is intracellularly retained in myelinating Schwann cells. AB - Missense mutations in the murine peripheral myelin protein-22 gene (Pmp22) underly the neuropathies in the trembler (Tr) and trembler-J (Tr-J) mice and in some humans with Charcot-Marie-Tooth disease. We have generated replication defective adenoviruses containing epitope-tagged, wild-type-, Tr-, or Tr-J-PMP22 bicistronic with the Lac-Z reporter gene. These viruses were microinjected into the sciatic nerves of 10-day-old Sprague-Dawley rats and, later, analyzed by immunohistochemistry to determine the distribution of mutant protein in infected myelinating Schwann cells. We found that epitope-tagged, wild-type PMP22 is successfully transported to compact myelin, whereas the Tr and the Tr-J mutant proteins are retained in cytoplasmic compartment, colocalizing with the endoplasmic reticulum. These results provide in vivo evidence that the pathogenesis of the Tr and Tr-J mutations are most likely a function of abnormal retention within the endoplasmic reticulum of myelinating Schwann cells. PMID- 11114257 TI - Brain-derived neurotrophic factor in astrocytes, oligodendrocytes, and microglia/macrophages after spinal cord injury. AB - Recent studies suggest that the injured adult spinal cord responds to brain derived neurotrophic factor (BDNF) and neurotrophin-3 (NT3) with enhanced neuron survival and axon regeneration. Potential neurotrophin sources and cellular localization in spinal cord are largely undefined. We examined glial BDNF localization in normal cord and its temporospatial distribution after injury in vivo. We used dual immunolabeling for BDNF and glial fibrillary acidic protein (GFAP) in astrocytes, adenomatous polyposis coli tumor suppressor protein (APC) for oligodendrocytes or type III CDH receptor (OX42) for microglia/macrophages. In normal cord, small subsets of astrocytes and microglia/macrophages and most oligodendrocytes exhibited BDNF-immunoreactivity. Following injury, the number of BDNF-immunopositive astrocytes and microglia/macrophages increased dramatically at the injury site over time. Most oligodendrocytes contained BDNF 1 day and 1 week following injury, but APC-positive cells were largely absent at the injury site 6 weeks postinjury. Glial BDNF-immunolabeling was also examined 10 and 20 mm from the wound. Ten millimeters from the lesion, astrocyte and microglia/macrophage BDNF-immunolabeling resembled that at the injury at all times examined. Twenty millimeters from injury, BDNF localization in all three glial subtypes resembled controls, regardless of time postlesion. Our findings suggest that in normal adult cord, astrocytes, oligodendrocytes, and microglia/macrophages play roles in local trophin availability and in trophin mediated injury and healing responses directly within and surrounding the wound site. PMID- 11114258 TI - Homozygous deletion in the coding sequence of the c-mer gene in RCS rats unravels general mechanisms of physiological cell adhesion and apoptosis. AB - The RCS rat presents an autosomal recessive retinal pigment epithelium dystrophy characterized by the outer segments of photoreceptors being phagocytosis deficient. A systematic genetic study allowed us to restrict the interval containing the rdy locus to that between the markers D3Mit13 and D3Rat256. We report the chromosomal localization of the rat c-mer gene in the cytogenetic bands 3q35-36, based on genetic analysis and radiation hybrid mapping. Using a systematic biocomputing analysis, we identified two strong related candidate genes encoding protein tyrosine kinase receptors of the AXL subfamily. The comparison of their expression patterns in human and mice tissues suggested that the c-mer gene was the best gene to screen for mutations. RCS rdy- and RCS rdy+ cDNAs were sequenced. The RCS rdy- cDNAs carried a significant deletion in the 5' part of the coding sequence of the c-mer gene resulting in a shortened aberrant transcript encoding a 20 amino acid peptide. The c-mer gene contains characteristic motifs of neural cell adhesion. A ligand of the c-mer receptor, Gas6, exhibits antiapoptotic properties. PMID- 11114259 TI - Apoptosis in the central nervous system of cerebral adrenoleukodystrophy patients. AB - The childhood cerebral form of adrenoleukodystrophy (ALD) is a fatal demyelinating disease, yet mice deficient in the ALD gene do not show such clinicopathological phenotype. We have therefore investigated in human autopsy tissues whether the ALD gene mutation results in apoptosis of CNS cells. Specimens from telencephalic and brainstem regions of four patients, and three controls were examined for internucleosomal DNA fragmentation, in situ detection of DNA breaks by the TUNEL method, and caspase-3 immunostaining. None of the controls showed significant apoptosis in white matter, while apoptotic nuclei with chromatin alterations were detected in areas of active demyelination in three ALD patients. A large proportion of apoptotic cells were oligodendrocytes and some express activated caspase-3. TUNEL-positive nuclei and/or caspase-3 staining were also detected in perivascular infiltrates and, occasionally, in neurons. We conclude that apoptosis of oligodendrocytes may account, at least in part, for the demyelinating process in the ALD brain. PMID- 11114260 TI - p53 is abnormally elevated and active in the CNS of patients with amyotrophic lateral sclerosis. AB - Motor neuron degeneration in amyotrophic lateral sclerosis (ALS) is a form of apoptosis, but the mechanisms for this neuronal cell death are not known. We evaluated whether motor neuron degeneration in ALS is associated with changes in the levels and function of the apoptosis regulating protein p53. The protein levels and localizations of p53 are abnormal in ALS. By immunoblotting, p53 is elevated in the nuclear compartment of selectively vulnerable CNS regions in individuals with ALS compared to age-matched controls. The levels of a carboxyl terminal degradation fragment of p53 were decreased in cases of ALS. DNA binding assay demonstrated that the increased p53 in individuals with ALS had competent DNA binding activity. Immunocytochemistry revealed that in normal human CNS p53 is expressed in subsets of nonneuronal cells, but it is found only rarely in neurons; in contrast, in individuals with ALS, p53 is frequently found in motor neurons of spinal cord and motor cortex and is upregulated in astroglia. It is concluded that p53 may participate in the mechanisms for motor neuron apoptosis in ALS. PMID- 11114261 TI - Human Cu/Zn superoxide dismutase (SOD1) overexpression in mice causes mitochondrial vacuolization, axonal degeneration, and premature motoneuron death and accelerates motoneuron disease in mice expressing a familial amyotrophic lateral sclerosis mutant SOD1. AB - Cytosolic Cu/Zn superoxide dismutase (SOD1) is a ubiquitous small cytosolic metalloenzyme that catalyzes the conversion of superoxide anion to hydrogen peroxide (H(2)O(2)). Mutations in the SOD1 gene cause a familial form of amyotrophic lateral sclerosis (fALS). The mechanism by which mutant SOD1s causes ALS is not understood. Transgenic mice expressing multiple copies of fALS-mutant SOD1s develop an ALS-like motoneuron disease resembling ALS. Here we report that transgenic mice expressing a high concentration of wild-type human SOD1 (hSOD1(WT)) develop an array of neurodegenerative changes consisting of (1) swelling and vacuolization of mitochondria, predominantly in axons in the spinal cord, brain stem, and subiculum; (2) axonal degeneration in a number of long fiber tracts, predominantly the spinocerebellar tracts; and (3) at 2 years of age, a moderate loss of spinal motoneurons. Parallel to the development of neurodegenerative changes, hSOD1(WT) mice also develop mild motor abnormalities. Interestingly, mitochondrial vacuolization was associated with accumulation of hSOD1 immunoreactivity, suggesting that the development of mitochondrial pathology is associated with disturbed SOD1 turnover. In this study we also crossed hSOD1(WT) mice with a line of fALS-mutant SOD1 mice (hSOD1(G93A)) to generate "double" transgenic mice that express high levels of both wild-type and G93A mutant hSOD1. The "double" transgenic mice show accelerated motoneuron death, earlier onset of paresis, and earlier death as compared with hSOD1(G93A) littermates. Thus in vivo expression of high levels of wild-type hSOD1 is not only harmful to neurons in itself, but also increases or facilitates the deleterious action of a fALS-mutant SOD1. Our data indicate that it is important for motoneurons to control the SOD1 concentration throughout their processes, and that events that lead to improper synthesis, transport, or breakdown of SOD1 causing its accumulation are potentially dangerous. PMID- 11114262 TI - Neurotoxicity of the putative transmembrane domain of the prion protein. AB - It has been shown recently that the generation of an abnormal transmembrane form of the prion protein ((Ctm)PrP) is involved in the neurodegeneration process during inherited and infectious prion diseases but a causative relationship has never been established. We wanted to know if and how the proposed transmembrane domain of PrP could induce neuronal dysfunction. Thus, we investigated the neurotoxic properties of two peptides whose sequences are encompassed within this domain. We show that PrP peptides 118-135 and 105-132 as well as an amidated more soluble peptide 105-132 induce the death of pure cortical neurons originating from normal and PrP knockout mice. This can be correlated with the high propensity of these peptides to insert stably into and to destabilize cell membranes. Through this study, we have identified a novel mechanism of neurotoxicity for PrP, which directly involves membrane perturbation; this mechanism is independent of fibril formation and probably corresponds to the effect of the transmembrane insertion of (Ctm)PrP. PMID- 11114263 TI - Neurodegeneration is associated to changes in serum insulin-like growth factors. AB - Serum levels of insulin and insulin-like growth factors and their binding proteins (IGFs and IGFBPs, respectively) are changed in human neurodegenerative diseases of very different etiology, such as Alzheimer's disease, amyotrophic lateral sclerosis, or cerebellar ataxia. However, the significance of these endocrine disturbances is not clear. We now report that in two very different inherited neurodegenerative conditions, ataxia-telangiectasia (AT) and Charcot Marie-Tooth 1A (CMT-1A) disease, serum levels of IGFs are also altered. Both types of patients have increased serum IGF-I and IGFBP-2 levels, and decreased serum IGFBP-1 levels, while only AT patients have high serum insulin levels. Furthermore, serum IGFs are also changed in three different animal models of neurodegeneration: neurotoxin-induced motor discoordination, diabetic neuropathy, and hereditary cerebellar ataxia. In these three models, serum insulin levels are significantly decreased, serum IGF-I and IGFBP-1, -2, and -3 are decreased in diabetic and neurotoxin-injected rats, while serum IGFBP-1 is increased in hereditary ataxic rats. Altogether, these observations indicate that a great variety of neurodegenerative diseases show endocrine perturbations, resulting in changes in serum IGFs levels. These perturbations are disease-specific and are probably due to metabolic and endocrine derangements, nerve cell death, and sickness-related disturbances associated to the neurodegenerative process. Our observations strongly support the need to evaluate serum IGFs in other neurodegenerative conditions. PMID- 11114264 TI - Cellular expression of alpha7 nicotinic acetylcholine receptor protein in the temporal cortex in Alzheimer's and Parkinson's disease--a stereological approach. AB - Cognitive deficits in Alzheimer's and Parkinson's disease are closely related to disturbed cholinergic transmission. The decrease of nicotinic acetylcholine receptor protein has been assessed by Western blotting and immunohistochemistry. Stereology, however, has not been used to assess numbers of receptor-expressing human cerebrocortical neurons. Our approach applies a combination of alpha7 subunit-immunohistochemistry with a stereological technique using defined stretches of pial surface as reference standard. The number of alpha7 subunit protein-expressing neurons in the Alzheimer temporal cortices amounted to approximately half of that of controls while numbers in Parkinson patients lay in between. No differences in the total number of neurons were seen. These results corroborate nonstereological studies on Alzheimer cortices and for the first time show a similar decrease in receptor expression in Parkinson's disease. They provide evidence that not only Alzheimer dementia but also cognitive deficits in Parkinson's disease may be related to decreased nicotinic receptor expression. PMID- 11114265 TI - FAD mutations in presenilin-1 or amyloid precursor protein decrease the efficacy of a gamma-secretase inhibitor: evidence for direct involvement of PS1 in the gamma-secretase cleavage complex. AB - To investigate the mechanism of regulation of Ass production by familial Alzheimer's disease (FAD)-linked presenilin 1 (PS1), we used a cell-free system that allows de novo Ass generation to examine whether PS1 participates directly in the gamma-secretase reaction. Optimal Ass generation in vitro was achieved at mildly acidic pH and could be inhibited by the aspartyl protease inhibitor pepstatin A, consistent with the suggestion that gamma-secretase is an aspartyl protease. Dominant negative mutations of the critical transmembrane aspartates in PS1 or full deletion of PS1 did not alter the maturation of APP in the secretory pathway. Instead, PS1 had a direct effect on the inhibition of Ass production by a designed peptidomimetic inhibitor: the inhibition was significantly less effective in cells expressing FAD-causing mutations in either APP or PS1 than in cells expressing the wild-type proteins. Taken together, these findings suggest that PS1 participates physically in a complex with APP during the gamma-secretase cleavage event. PMID- 11114266 TI - Costimulatory effects of interferon-gamma and interleukin-1beta or tumor necrosis factor alpha on the synthesis of Abeta1-40 and Abeta1-42 by human astrocytes. AB - Chronic inflammation and astrocytosis are characteristic histopathological features of Alzheimer's Disease (AD). Astrocytes are one of the predominant cell types in the brain. In AD they are activated and produce inflammatory components such as complement components, acute phase proteins, and cytokines. In this study we analyzed the effect of cytokines on the production of amyloid beta (Abeta) in the astrocytoma cell line U373 and in primary human astrocytes isolated postmortem from healthy aged persons as well as from patients with AD. Astrocytes did not produce Abeta in the absence of stimuli or following stimulation with IL 1beta, TNFalpha, IL-6, and TGF-beta1. Neither did combinations of TNFalpha and IL 1beta, IL-6 or TGF-beta1, or the coadministration of IFNgamma and IL-6 or TGF beta1 induce Abeta production. In contrast, pronounced production of Abeta1-40 and Abeta1-42 was observed when primary astrocytes or astrocytoma cells were stimulated with combinations of IFNgamma and TNFalpha or IFNgamma and IL-1beta. Induction of Abeta production was accompanied by decreased glycosylation of APP as well as by increased secretion of APPsbeta. Our results suggest that astrocytes may be an important source of Abeta in the presence of certain combinations of inflammatory cytokines. IFNgamma in combination with TNFalpha or IL-1beta seems to trigger Abeta production by supporting beta-secretase cleavage of the immature APP molecule. PMID- 11114267 TI - The origins and consequences of public trust in government: a time series analysis. AB - The study of citizens' trust in the national government has been primarily individual-level, cross-sectional analysis. In the current research, we develop a quarterly time series measure of trust in the U.S. national government from 1980 to 1997 and conduct the first multivariate time series examination of public trust in government. We find that negative perceptions of the economy, scandals associated with Congress, and increasing public concern about crime each lead to declining public trust in government. Declining trust in government in turn leads to less positive evaluations of Congress and reduced support for government action to address a range of domestic policy concerns. These results provide new evidence of the influence of public concern about crime and the centrality of Congress in understanding public evaluations of the national government and new evidence of how declining levels of trust in government may influence elections and domestic policy making. PMID- 11114268 TI - Question wording and the house vote choice : some experimental evidence. AB - Since 1978, the vote reported for House incumbents in the American National Election Studies (NES) has been significantly higher than the actual incumbents' vote in the districts surveyed; in NES surveys before 1978, the reported vote was much closer to the actual vote. The prime suspect for the source of this bias is the new question format introduced in 1978 and used in all subsequent studies. We document the problem and review the results of several question-wording experiments that confirm the superior accuracy of a format that does not mention the candidates' names over the ballot format currently in use. We also find evidence that a modified version of the ballot format may reduce the pro incumbent bias, so that improvement may be possible without a major interruption of the post-1978 NES times series. PMID- 11114269 TI - The measurement of personal values in survey research: a test of alternative rating procedures. AB - When survey researchers are interested in measuring the personal values of respondents, they often use a rating rather than a ranking method because it is easier and faster to administer and yields data that are amenable to parametric statistical analyses. However, because personal values are inherently positive constructs, respondents often exhibit little differentiation among the values and end-pile their ratings toward the positive end of the scale. Such lack of differentiation may potentially affect the statistical properties of the values and the ability to detect relationships with other variables. Two experiments were conducted via mail surveys to general population samples to test alternative rating methods designed to increase differentiation and reduce end-piling in the rating of personal values. The results suggest that a procedure in which respondents first pick their most and least important values, then rate them (most-least), provides more differentiation and less end-piling than a simple rating procedure (rate-only). Increased differentiation for the most-least method influenced the fit of latent structure and resulted in more robust relations between the values ratings and other criterion variables. These results generalized across type of values scale, number of values rated, and number of rating points. PMID- 11114270 TI - Leverage-saliency theory of survey participation: description and an illustration. PMID- 11114271 TI - The polls--trends : support for the women's movement. PMID- 11114272 TI - How to treat women with antiphospholipid antibodies in pregnancy? PMID- 11114273 TI - Gold treatment, nephrotic syndrome, and multi-organ failure in a patient with adult onset Still's disease. PMID- 11114274 TI - Diagnosis and nonsurgical management of osteoarthritis PMID- 11114275 TI - Validated measurement of periarticular bone mineral density at the knee joint by dual energy x ray absorptiometry. AB - OBJECTIVE: The association of inflammatory arthritis with loss of periarticular bone mineral density (BMD) has been well established. However, changes in bone density cannot be quantified by conventional radiography. This study aimed at developing a new technique for measurement of periarticular bone density at the knee joint by dual energy x ray absorptiometry (DXA) and assessing the precision of this technique for selected areas around the knee. METHODS: To validate this technique for bone density assessment in both patient and control subjects, knee joints from healthy subjects and patients with inflammatory arthritis were selected for study. Posteroanterior (PA) and lateral scans of both knees were acquired with the Hologic 4500 elite bone densitometer. Each scan was repeated three times, with repositioning between scans. Knee scans were obtained with the forearm software and evaluated by subregion analysis. Seven femoral and seven tibial subregions of interest (ROIs) were selected on PA scans. Six ROIs were selected on lateral scans. Precision was determined for each ROI selected. RESULTS: 14 knee joints were studied in each group. Precision, expressed as percentage coefficient of variation (CV%), varied widely between subregions. PA scans were most appropriate for measurement of femoral bone density (CV% = 1.89 2.64%), whereas the best value obtained for ROIs within the tibia was on the lateral scan, where CV% for measurement of the proximal 5 mm was 2.67% in the patient group. CV% for BMD of the patella was excellent at 0.84% in the patient group. CONCLUSION: This new application of DXA can be used to measure periarticular bone density at the knee joint. Regions within the distal femur and patella have been identified as the optimal areas to study PMID- 11114276 TI - Bone scintigraphy for osteonecrosis of the knee in patients with non-traumatic osteonecrosis of the femoral head: comparison with magnetic resonance imaging. AB - OBJECTIVE: To determine whether technetium bone scintigraphy (BS) is useful for screening of non-traumatic osteonecrosis of the knee (ONK), which was a major affected site, secondary to the femoral head, among multiple osteonecrosis, in patients with non-traumatic osteonecrosis of the femoral head (ONFH). METHODS: A total of 214 knee joints in 107 patients with ONFH were evaluated by BS and a comparison made with magnetic resonance imaging (MRI). ONK was classified into five sites, including the femoral condyles (ONFC), distal femoral metaphysis (ONFM), tibial plateau (ONTP), proximal tibial metaphysis (ONTM), and patella (ONP). RESULTS: Based on the diagnosis by MRI, ONK was detected in 103 knees of 62 patients (48%). ONFC was most common (86 knees, 40%), ONFM (15%), followed by ONTM (10%), ONP (3%), and ONTP (0.9%). Sensitivity, specificity, and accuracy of BS for ONFC detection were 63%, 71%, and 68%, respectively. When the ONFC lesions on the coronal views of MRI were large or medium sized and occupied two thirds, or the entire anteroposterior joint surface on the sagittal views, the sensitivity of BS for ONFC detection increased to 89% (34/38 knees). The sensitivity of BS for ONFM, ONTM, and ONP detection was 3%, 0%, and 0%, respectively, but these lesions showed a low likelihood of collapse. CONCLUSION: BS is useful for screening large ONK in patients with ONFH given that 89% of patients with ONFC who had a high risk of collapse of the knee were identified. PMID- 11114277 TI - Strength training induced adaptations in neuromuscular function of premenopausal women with fibromyalgia: comparison with healthy women. AB - OBJECTIVE: To investigate the effects of 21 weeks' progressive strength training on neuromuscular function and subjectively perceived symptoms in premenopausal women with fibromyalgia (FM). METHODS: Twenty one women with FM were randomly assigned to experimental (FM(T)) or control (FM(C)) groups. Twelve healthy women served as training controls (H(T)). The FM(T) and H(T) groups carried out progressive strength training twice a week for 21 weeks. The major outcome measures were muscle strength and electromyographic (EMG) recordings. Secondary outcome measures were pain, sleep, fatigue, physical function capacity (Stanford Health Assessment Questionnaire), and mood (short version of Beck's depression index). RESULTS: Female FM(T) subjects increased their maximal and explosive strength and EMG activity to the same extent as the H(T) group. Moreover, the progressive strength training showed immediate benefits on subjectively perceived fatigue, depression, and neck pain of training patients with FM. CONCLUSIONS: The strength training data indicate comparable trainability of the neuromuscular system of women with FM and healthy women. Progressive strength training can safely be used in the treatment of FM to decrease the impact of the syndrome on the neuromuscular system, perceived symptoms, and functional capacity. These results confirm the opinion that FM syndrome has a central rather than a peripheral or muscular basis. PMID- 11114278 TI - Release of cartilage and bone macromolecules into synovial fluid: differences between psoriatic arthritis and rheumatoid arthritis. AB - OBJECTIVE: To elucidate whether differences in the destructive tissue process in cartilage and bone in psoriatic arthritis (PsA) and rheumatoid arthritis (RA) can be recognised by different release patterns of molecular fragments derived from joint tissue. METHODS: Aggrecan, cartilage oligomeric matrix protein (COMP), and bone sialoprotein (BSP) were quantified by immunoassays in knee joint synovial fluid samples. These were obtained early in the disease course of patients with PsA and RA. At the time of arthrocentesis radiographs of their knee and hip joints were normal. RESULTS: At follow up no destruction had developed in the knees and hips of most patients with PsA (n=18), whereas the patients with RA could be separated into one "destructive" group (n=18) and one "non-destructive" group (n=25). Patients with PsA had low synovial fluid aggrecan concentrations (p<0.001 v the RA destructive group) but high COMP concentrations (p<0.01 and p<0.05 v destructive and non-destructive RA groups, respectively). Consequently, the aggrecan/COMP ratio was lowest in the PsA group (p<0.001 and p<0.01 v the destructive and non-destructive RA group, respectively). The synovial fluid concentrations of BSP did not differ between the three patient groups. CONCLUSIONS: The release pattern of aggrecan and COMP, reflecting cartilage turnover, differed between the PsA group and, particularly, the destructive RA group. This suggests that different pathophysiological mechanisms for cartilage involvement operate in these conditions, with different destructive potential. The BSP concentrations did not differ between the patients groups, which indicates similar levels of bone involvement. PMID- 11114279 TI - Association of autoantibodies to filaggrin with an active disease in early rheumatoid arthritis. AB - OBJECTIVE: To evaluate the clinical significance of antifilaggrin antibodies (AFA) measured by an enzyme linked immunosorbent assay (ELISA) in serial specimens from patients with recent onset rheumatoid arthritis (RA). METHODS: Filaggrin was purified from human skin and used as an antigen in ELISA. The AFA test was applied to five serial specimens from 78 patients with recent onset RA followed up for three years. Rheumatoid factor (RF) had been measured earlier from the same samples by quantitative immunoturbidimetry. RESULTS: The mean AFA level was highest at entry (54% positive), followed by a statistically significant decline at six months and a slight increase at three years. AFA were persistently positive in 23 patients and persistently negative in 28 patients. Eleven of the latter patients were persistently negative for RF. At study entry AFA levels correlated to some degree with RF levels. In general, raised AFA levels at entry were associated with an active and treatment resistant disease, but they did not predict radiological progression. CONCLUSIONS: The test for AFA has potential for an additional immunological test for RA. PMID- 11114280 TI - Low T cell production of TNFalpha and IFNgamma in ankylosing spondylitis: its relation to HLA-B27 and influence of the TNF-308 gene polymorphism. AB - OBJECTIVE: To test the hypothesis that ankylosing spondylitis (AS) is a T helper cell type 2 polarised disease by quantifying the T cell cytokines interferon gamma (IFNgamma), interleukin 4 (IL4), tumour necrosis factor alpha (TNFalpha), and IL10 at the single cell level in patients with AS in comparison with healthy HLA-B27 negative and HLA-B27 positive controls. METHODS: Peripheral blood mononuclear cells from 65 subjects (25 HLA-B27 positive patients with active AS, 18 healthy HLA-B27 positive controls, and 22 healthy HLA-B27 negative controls) were stimulated with phorbol myristate acetate/ionomycin for six hours, surface stained for CD3 and CD8, intracellularly stained for the cytokines IFNgamma, TNFalpha, IL4, and IL10, and analysed by flow cytometry. TNFalpha production was related to the genotype of the TNFalpha promoter at the -308 and -238 polymorphisms. RESULTS: In peripheral blood the percentage of TNFalpha+ T cells was significantly lower in HLA-B27 positive patients with AS (median 5.1% for CD4+ T cells) than in healthy HLA-B27 negative controls (median 9.5%; p=0.008). Surprisingly, the percentage of TNFalpha+ T cells was also significantly lower in healthy HLA-B27 positive controls (median 7.48%) than in healthy HLA-B27 negative controls (p=0.034). Furthermore, the percentage of IFNgamma+ T cells was lower in patients with AS and in healthy HLA-B27 positive controls than in healthy HLA-B27 negative controls (p=0.005 and p=0.003, respectively). The percentage of IL10+/CD8+ T cells was higher in patients with AS than in both control groups. In HLA-B27 positive subjects, TNF1/2 heterozygosity at -308 (n=6) was associated with a higher percentage of TNFalpha+ T cells than TNF1/1 homozygosity (n=25; median 9.97% v 5.11% for CD4+ T cells; p=0.017). In contrast, in HLA-B27 negative controls (n=18) there was no such genotype/phenotype correlation (median 9.4% v 10.6%). CONCLUSIONS: The lower T cell production of TNFalpha and IFNgamma shown at the single cell level in HLA-B27 positive patients with AS and healthy HLA-B27 positive controls may contribute to the increased susceptibility of HLA-B27 positive subjects to develop AS. Preliminary genotype-phenotype correlations suggest that in HLA-B27 positive subjects TNF2 at -308 or a linked gene results in higher TNFalpha production and, therefore, might be a marker for a protective haplotype. PMID- 11114281 TI - Right ventricular diastolic dysfunction in patients with anticardiolipin antibodies and antiphospholipid syndrome. AB - OBJECTIVE: To evaluate the prevalence of diastolic dysfunction in patients with anticardiolipin antibodies (aCL) and to examine whether the antiphospholipid syndrome (APS) is associated with diastolic dysfunction independently of valvular abnormalities and systolic dysfunction. METHODS: Pulsed, continuous, colour Doppler echocardiography was performed in 179 subjects, of whom 15 were excluded from the analysis because of systolic dysfunction or severe valvular disease. The remaining 164 subjects included 29 patients with primary APS, 26 patients with secondary APS (APS in the presence of systemic lupus erythematosus (SLE)), and 30 patients with SLE and aCL but without APS; 43 patients with SLE without aCL and 36 normal volunteers served as control groups. RESULTS: The groups compared differed significantly in all measures of right ventricular function. There was a gradation of increasing diastolic function impairment as manifested by prolonged deceleration time (DT) and isovolumic relaxation time (IVRT) across the groups of patients with SLE without aCL, SLE with aCL, secondary APS, and primary APS. Differences in left ventricular diastolic function measures were less prominent. In regression analysis, DT increased by 19.6 ms (p=0.002) in the presence of primary APS and by 20.1 ms (p=0.038) in the presence of pulmonary hypertension. The titre of IgG aCL was the strongest predictor of a prolonged IVRT. CONCLUSION: Diastolic dysfunction, in particular of the right ventricle-that is, independent of valvular disease and systolic dysfunction, is a prominent feature of APS and may be related to the pathogenesis of the syndrome. PMID- 11114282 TI - Recognition of YKL-39, a human cartilage related protein, as a target antigen in patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate whether autoimmunity to YKL-39, a recently cloned cartilage protein, occurs in patients with rheumatoid arthritis (RA). METHODS: Autoantibody to YKL-39 was assayed by enzyme linked immunosorbent assay (ELISA) and western blotting in serum samples from patients with RA, systemic lupus erythematosus (SLE), and healthy donors, using recombinant YKL-39 protein. This reactivity was compared with that against a YKL-39 homologue, YKL-40 (human cartilage gp-39/chondrex), which has been reported to be an autoantigen in RA. RESULTS: Autoantibody to YKL-39 was detected in seven of 87 patients with RA (8%), but not in serum samples from patients with SLE or healthy donors. YKL-40 reactivity was found in only one of 87 RA serum samples (1%), with no cross reactivity to YKL-39. CONCLUSION: The existence of anti-YKL-39 antibody in a subset of patients with RA is reported here for the first time. Further, it was shown that the immune response to YKL-39 was independent of that to YKL-40. Clarification of the antibody and T cell responses to autoantigens derived from chondrocyte, cartilage, or other joint components may lead to a better understanding of the pathophysiology of joint destruction in patients with RA. PMID- 11114283 TI - Decline after immobilisation and recovery after remobilisation of synovial fluid IL1, TIMP, and chondroitin sulphate levels in young beagle dogs. AB - OBJECTIVE: To monitor the concentration of markers of cartilage and synovium metabolism in the knee (stifle) joint synovial fluid of young beagles subjected to immobilisation and subsequent remobilisation. METHODS: The right hind limb of 17 dogs was immobilised in flexion for 11 weeks. Simultaneously, the contralateral left knee was exposed to increased weight bearing. The remobilisation period lasted 50 weeks. Litter mates served as controls. The concentration in joint lavage fluid of interleukin 1alpha (IL1alpha) was measured by immunoassay, the activity of phospholipase A(2) (PLA(2)) was determined by an extraction method, chondroitin sulphate (CS) concentration by precipitation with Alcian blue, hyaluronan (HA) by an ELISA-like assay using biotinylated HA-binding complexes, matrix metalloproteinase 3 (MMP-3), and tissue inhibitor of metalloproteinases 1 (TIMP-1) by sandwich ELISA, and synovitis was scored by light microscopy. RESULTS: Synovitis or effusion was absent in all experimental and control groups. Immobilisation decreased the joint lavage fluid levels of IL1alpha (p<0.05), TIMP (p< 0.05), and the concentration of CS down to 38% (p<0.05) in comparison with untreated litter mates with normal weight bearing. Immobilisation did not affect the activity of PLA(2), or the concentration of MMP 3 or HA in synovial fluid. Joint remobilisation restored the decreased concentrations of markers to control levels. Increased weight bearing did not change the concentrations of markers in comparison with the control joints with normal weight bearing. CONCLUSIONS: 11 weeks' joint immobilisation decreased the concentration of markers of cartilage and synovium metabolism in the synovial fluid, and remobilisation restored the concentrations to control levels. The changes in joint metabolism induced by immobilisation, as reflected by the markers, are thus different from those found in osteoarthritis, where increased levels of these markers are associated with enhanced degradation and synthesis. These findings suggest that the change induced in joint metabolism by immobilisation is reversible in its early stages. PMID- 11114285 TI - Intestinal permeability in Behcet's syndrome. AB - OBJECTIVE: To measure the intestinal permeability in patients with Behcet's syndrome (BS) and to compare the results with those obtained from healthy and diseased controls. METHOD: The study group comprised 34 patients with BS without known gastrointestinal disease. Ten patients with ankylosing spondylitis (AS), 6 with inflammatory bowel diseases (IBD), 17 with systemic lupus erythematosus (SLE), and 15 healthy subjects (HC) constituted the controls. All patients received 100 microCi (3.7 MBq) of chromium-51 EDTA ((51)Cr-EDTA) as a radioactive tracer after a 72 hour abstinence from all drugs. The percentage of the isotope excreted in a 24 hour urinary specimen was the measure of permeability. RESULTS: The percentage (SD) rate of excretion of (51)Cr-EDTA was 4.6 (2.6) in BS, 6 (2.4) in AS, 5.2 (1. 9) in IBD, 5.56 (1.78) in SLE, and 2.3 (1) in healthy controls. (Analysis of variance: f=6.4, p=0.0002. BS v HC, AS v HC, SLE v HC significant.) CONCLUSION: The intestinal permeability in BS was significantly more than that seen among the healthy controls. Similar results in all the diseased controls cast doubt on its specificity. PMID- 11114284 TI - Short term effects of corticosteroid pulse treatment on disease activity and the wellbeing of patients with active rheumatoid arthritis. AB - OBJECTIVE: To investigate the short term effects of corticosteroid pulse treatment (CPT) on disease activity, functional ability, and psychological wellbeing of patients with active rheumatoid arthritis (RA). METHODS: Of 66 consecutive patients with active RA admitted for CPT, erythrocyte sedimentation rate, C reactive protein level, haemoglobin concentration, platelet count, duration of early morning stiffness, a joint score, and grip strength were assessed before and after CPT. Additionally, a health status questionnaire was administered. Effects of CPT were expressed as before to after intervention effect sizes and, to place them in perspective, compared with the (long term) effect sizes of disease modifying antirheumatic drug (DMARD) treatment in a historical contrast group of patients with early RA. RESULTS: Statistically significant improvement from baseline in disease activity, physical functioning, and psychological wellbeing after CPT was seen, with moderate to large effect sizes, resembling the effects seen after DMARD treatment. Neither depression nor psychosis occurred during and after CPT. CONCLUSION: Qualitatively and quantitatively the short term effects of CPT in patients with active established RA on various dimensions of health status resemble the long term effects of conventional DMARD treatment in patients with early RA. Psychological disorders do not seem to be common short term side effects of CPT in patients with active RA. PMID- 11114286 TI - Use of anti-citrullinated peptide and anti-RA33 antibodies in distinguishing erosive arthritis in patients with systemic lupus erythematosus and rheumatoid arthritis. AB - OBJECTIVES: Systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) can both present with an erosive arthritis with the small joints of the hands affected. Therefore a serological marker would be useful to distinguish between these two diseases at onset. In this study anti-RA33 antibodies, which are found in patients with SLE and RA, and anti-citrullinated peptide antibodies (anti CCP), which have recently been described as highly specific for RA, were assessed. METHODS: Two hundred and thirty one patients receiving long term follow up for SLE were evaluated for arthritis and classified as erosive and non-erosive disease. Sixty six patients were tested for anti-RA33 and anti-CCP antibodies. All the patients were tested for rheumatoid factor (RF) and HLA-DR4 status. RESULTS: Ten patients had erosive disease, six of whom were RF positive (60%), and six anti-RA33 positive (60%), whereas only two were anti-CCP positive (20%). Two hundred and twenty one patients had non-erosive disease, 40 of whom were RF positive (18%), 14 were anti-RA33 positive (6%), whereas only one patient was found to be anti-CCP positive (0.5%). CONCLUSION: The presence of anti-CCP antibodies may be useful in distinguishing RA from erosive SLE. Anti-RA33 antibodies and RF are unhelpful. PMID- 11114287 TI - Prolonged prodrome, systemic vasculitis, and deafness in Cogan's syndrome. AB - Cogan's syndrome is a rare, multisystem disease which occurs predominantly in children and young adults. It was originally described as the combination of interstitial keratitis and audiovestibular disturbance, but other forms of ocular disease, as well as systemic vasculitis, have since been recognised as part of the syndrome. Diagnosis can be difficult if the various manifestations occur separately, but early recognition is important because prompt treatment may prevent deafness. Two cases are presented here illustrating the features of this disease, and providing histological evidence of systemic vasculitis in both. PMID- 11114288 TI - Dephosphorylation of autoantigenic ribosomal P proteins during Fas-L induced apoptosis: a possible trigger for the development of the autoimmune response in patients with systemic lupus erythematosus. AB - OBJECTIVES: Autoimmune diseases are characterised by the production of autoantibodies against various autoantigens. In the past few years data have been published on a possible role of apoptosis in the development of autoimmunity. These include the finding that several autoantigens become modified (for example, by cleavage) during apoptosis, and the observation that these modified antigens are translocated to the cell surface. When the normal clearance of apoptotic cells somehow is disturbed, such modified antigens might become exposed to the immune system. Because acidic ribosomal P (phospho-) proteins targeted by autoantibodies in systemic lupus erythematosus (SLE) are also concentrated at the surface of apoptotic cells, this study aimed at investigating what modifications occur on these antigens during apoptosis. METHODS: Apoptosis in Jurkat cells was induced by Fas ligand (Fas-L), and the fate of autoantigenic P proteins was analysed in both normal and apoptotic total cell extracts. RESULTS: The autoantigenic P proteins were not cleaved but dephosphorylated during Fas-L induced apoptosis. This dephosphorylation was prevented when caspase activity was inhibited. CONCLUSIONS: As has been shown for other autoantigens targeted by autoantibodies in SLE, P proteins also are modified during apoptosis. P1 and P2 are completely dephosphorylated while P0 is partly dephosphorylated. Because the epitope targeted by autoantibodies normally is phosphorylated, it is possible that the apoptotic dephosphorylation of the antigen might be the trigger for the development of the autoimmune response against P proteins. PMID- 11114289 TI - Acceptance of the different denominations for reflex sympathetic dystrophy. AB - OBJECTIVE: To elucidate the real impact in the medical literature of the different denominations for reflex sympathetic dystrophy (RSD). METHODS: A search was performed through the Medline database (WinSPIRS, SilverPlatter International, NS), from 1995 to 1999, including the following descriptors: RSD, complex regional pain syndrome (CRPS), CRPS type I, algodystrophy, Sudeck, shoulder-hand syndrome, transient osteoporosis, causalgia, and CRPS type II. RESULTS: The descriptor RSD was detected in 576 references, algodystrophy in 54, transient osteoporosis in 42, CRPS type I in 24, Sudeck in 16, and shoulder-hand syndrome in 11. One hundred records were obtained for the descriptor causalgia and five for CRPS type II. The descriptor RSD was detected in the title of 262 references, algodystrophy in 29, transient osteoporosis in 29, CRPS type I in 15, Sudeck in 3, shoulder-hand syndrome in 5, causalgia in 17, and CRPS type II in 3 references. CONCLUSIONS: The new CRPS terminology has not effectively replaced the old one. RSD and causalgia are the most used denominations. PMID- 11114290 TI - Preservation of calcium pyrophosphate dihydrate crystals: effect of Mayer's haematoxylin staining period. AB - OBJECTIVE: To clarify the deleterious effects of Mayer's haematoxylin staining procedure which result in a decrease in, or complete loss of, the number of calcium pyrophosphate dihydrate (CPPD) crystals, and to determine the proper staining period for preserving the crystals in a histological paraffin section of articular tissues. METHODS: Paraffin sections of CPPD crystal-bearing articular tissues of six patients were stained with Mayer's haematoxylin for 3, 8, or 15 minutes, and subsequently with eosin for one minute. The specimens were examined with an Olympus BHS polarised light microscope. The pH of Mayer's haematoxylin solution was measured with a TOA pH meter. RESULTS: Positive birefringent CPPD crystals were seen clearly in all specimens stained with Mayer's haematoxylin for three minutes. The specimens stained for eight minutes showed a reduced number of crystals. No crystals were seen in the specimens stained for 15 minutes. Ordinary light microscopy showed no notable differences in the stainability of nucleus, cell membrane, and their surrounding tissues among specimens when stained with Mayer's haematoxylin for either 3, 8, or 15 minutes. The pH of Mayer's haematoxylin solution was 2.31. CONCLUSIONS: To find CPPD crystals in the paraffin sections of articular tissues, the staining period with Mayer's haematoxylin should be limited to three minutes. The longer the staining period, the greater the reduction in the number of crystals owing to the strong acidity of the haematoxylin solution. A staining period of 15 minutes causes a complete loss of CPPD crystals. PMID- 11114291 TI - HHV8 associated Kaposi's sarcoma during triple immunosuppressive treatment with cyclosporin A, azathioprine, and prednisolone for ocular Behcet's disease and complete remission of both disorders with interferon alpha. PMID- 11114292 TI - Coffee or decaff? PMID- 11114293 TI - Sp1 and Smad proteins cooperate to mediate transforming growth factor-beta 1 induced alpha 2(I) collagen expression in human glomerular mesangial cells. AB - The mechanism(s) by which Smads mediate and modulate the transforming growth factor (TGF)-beta signal transduction pathway in fibrogenesis are not well characterized. We previously showed that Smad3 promotes alpha2(I) collagen gene (COL1A2) activation in human glomerular mesangial cells, potentially contributing to glomerulosclerosis. Here, we report that Sp1 binding is necessary for TGF beta1-induced type I collagen mRNA expression. Deletion of three Sp1 sites (GC box) between -376 and -268 or mutation of a CAGA box at -268/-260 inhibited TGF beta1-induced alpha2(I) collagen promoter activity. TGF-beta1 inducibility was also blocked by a Smad3 dominant negative mutant. Chemical inhibition of Sp1 binding with mithramycin A, or deletion of the GC boxes, inhibited COL1A2 activation by Smad3, suggesting cooperation between Smad3 and Sp1 in the TGF beta1 response. Electrophoretic mobility shift assay showed that Sp1 and Smads form complexes with -283/-250 promoter sequences. Coimmunoprecipitation experiments demonstrate that endogenous Sp1, Smad3, and Smad4 form complexes in mesangial cells. In a Gal4-LUC reporter assay system, Sp1 stimulated the TGF beta1-induced transcriptional activity of Gal4-Smad3, Gal4-Smad4 (266), or both. Using the transactivation domain B of Sp1 fused to the Gal4 DNA binding domain, we show that, in our system, the transcriptional activity of this Sp1 domain is not regulated by TGF-beta1, but it becomes responsive to this factor when Smad3 is coexpressed. Finally, combined Sp1 and Smad3 overexpression induces marked ligand-independent and ligand-dependent promoter activity of COL1A2. Thus, Sp1 and Smad proteins form complexes and their synergy plays an important role in mediating TGF-beta1-induced alpha2(I) collagen expression in human mesangial cells. PMID- 11114294 TI - NF-kappa B mediates up-regulation of CFTR gene expression in Calu-3 cells by interleukin-1beta. AB - Inflammation of the airways is a major feature of the inherited disease cystic fibrosis. Previous studies have shown that the pro-inflammatory cytokines tumor necrosis factor alpha and interferon gamma reduce the expression of the cystic fibrosis transmembrane conductance regulator (CFTR) gene (CFTR) in HT-29 and T84 cells by acting post-transcriptionally. We have investigated the effect of the pro-inflammatory peptide interleukin 1beta (IL-1beta) on the expression of the CFTR in Calu-3 cells. IL-1beta increased the production of CFTR mRNA in a dose- and time-dependent manner. Its action was inhibited by inhibitors of the NF kappaB pathway, including N-acetyl-l-cysteine, pyrrolidine dithiocarbamate, and a synthetic cell-permeable peptide containing the NF-kappaB nuclear localization signal sequence. Gel shift analysis showed that IL-1beta activated NF-kappaB in Calu-3 cells, and transfection experiments using p50 and RelA expressing vectors showed that exogenous transfected NF-kappaB subunits increased the concentration of CFTR mRNA. Gel shift analysis with antibody supershifting also showed that IL 1beta caused the binding of NF-kappaB to a kappaB-like response element at position -1103 to -1093 in the CFTR 5'-flanking region. Transfection experiments using -2150 to +52 CFTR reporter gene constructs showed that the activity of the CFTR promoter is enhanced by exogenous transfected NF-kappaB and IL-1beta and that this enhancement is due, at least in part, to the -1103 to -1093 kappaB site. We conclude that the intracellular signaling that leads to increased CFTR mRNA in response to IL-1beta in Calu-3 cells includes the binding of NF-kappaB to the -1103 kappaB element and a subsequent increase in CFTR promoter activity. PMID- 11114295 TI - MRG1 expression in fibroblasts is regulated by Sp1/Sp3 and an Ets transcription factor. AB - MRG1 (melanocyte-specific gene 1 (MSG1)-related gene), a ubiquitously expressed transcription factor that interacts with p300/CBP, TATA-binding protein and Lhx2, is the founding member of a new family of transcription factors. Initial characterization of this newly discovered transcription factor has underscored its potential involvement in many important cellular processes through transcriptional modulation. We previously demonstrated that MRG1 can be induced by various biological stimuli (Sun, H. B., Zhu, Y. X., Yin, T., Sledge, G., and Yang, Y. C. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 13555-13560). As a first step in understanding its role in different biological processes, we investigated mechanisms that regulate transcription of the mouse MRG1 gene in fibroblasts. Transient transfection of Rat1 fibroblast cells with sequential 5'-deletions of mouse MRG1 promoter-luciferase fusion constructs indicated that the -104 to +121 region contains the full promoter activity. Deletion and site-directed mutations within this region revealed that the Ets-1 site at -97 to -94 and the Sp1 site at -51 to -46 are critical for MRG1 expression in fibroblasts. Gel mobility shift and supershift assays performed with Rat1 nuclear extracts identified nucleoprotein complexes binding to the Ets-1 site and the Sp1 site. In Drosophila SL2 cells, which lack the Sp and Ets family of transcription factors, expression of Sp1, Sp3, and Ets-1 or Elf-1 functionally stimulated MRG1 promoter activity in a synergistic manner. These results suggest that multiple transcription factors acting in synergy are responsible for MRG1 expression and the responsiveness of cells to different biological stimuli. PMID- 11114297 TI - Identification of the E2A gene products as regulatory targets of the G1 cyclin dependent kinases. AB - The E2A gene products, E12 and E47, are multifunctional transcription factors that as homodimers regulate B cell development, growth, and survival. In this report, the E2A gene products are shown to be targets for regulation by the G1 cyclin-dependent kinases. Two novel G1 cyclin-dependent kinase sites are identified on the N-terminal domain of E12/E47. One site displays homology to a preferential D-type cyclin-dependent kinase site (serine 780) on the retinoblastoma susceptibility gene product (pRB) and, consistent with this homology, is more efficiently phosphorylated by cyclin D1-CDK4 than by the other cyclin-dependent kinases (CDK) that were tested. The second kinase site is phosphorylated by both cyclin D1-CDK4 and cyclin A/E-CDK2 complexes. Mutation studies indicated that phosphorylation of the cyclin D1-CDK4 site, or more potently, of both the cyclin D1-CDK4 and cyclin A/E-CDK2 sites, negatively regulates the growth suppressor function associated with the N-terminal domain of E12/E47. Transient expression studies showed that ectopic expression of cyclin D1 or E negatively regulates sequence-specific activation of gene transcription by E12/E47. Analysis of site mutants, however, indicated that inhibition of E12/E47 transcriptional activity did not require the N-terminal G1 cyclin-dependent kinase sites. Together, the results suggest that the growth suppressor and transcriptional activator functions of E12/E47 are targets for regulation by G1 cyclin-dependent kinases but that the mechanisms of regulation for each function are distinct. PMID- 11114298 TI - Granzyme B induces BID-mediated cytochrome c release and mitochondrial permeability transition. AB - Many cell death pathways converge at the mitochondria to induce release of apoptogenic proteins and permeability transition, resulting in the activation of effector caspases responsible for the biochemical and morphological alterations of apoptosis. The death receptor pathway has been described as a triphasic process initiated by the activation of apical caspases, a mitochondrial phase, and then the final phase of effector caspase activation. Granzyme B (GrB) activates apical and effector caspases as well as promotes cytochrome c (cyt c) release and loss of mitochondrial membrane potential. We investigated how GrB affects mitochondria utilizing an in vitro cell-free system and determined that cyt c release and permeability transition are initiated by distinct mechanisms. The cleavage of cytosolic BID by GrB results in truncated BID, initiating mitochondrial cyt c release. BID is the sole cytosolic protein responsible for this phenomenon in vitro, yet caspases were found to participate in cyt c release in some cells. On the other hand, GrB acts directly on mitochondria in the absence of cytosolic S100 proteins to open the permeability transition pore and to disrupt the proton electrochemical gradient. We suggest that GrB acts by two distinct mechanisms on mitochondria that ultimately lead to mitochondrial dysfunction and cellular demise. PMID- 11114299 TI - Molecular cloning of a dendritic cell-associated transmembrane protein, DC-HIL, that promotes RGD-dependent adhesion of endothelial cells through recognition of heparan sulfate proteoglycans. AB - We isolated a novel molecule (DC-HIL) expressed abundantly by the XS52 dendritic cell (DC) line and epidermal Langerhans cells, but minimally by other cell lines. DC-HIL is a type I transmembrane protein that contains a heparin-binding motif and an integrin-recognition motif, RGD, in its extracellular domain (ECD). A soluble fusion protein (DC-HIL-Fc) of the ECD and an immunoglobulin Fc bound to the surface of an endothelial cell line (SVEC). This binding induced adhesion of SVEC to its immobilized form. Sulfated polysaccharides (e.g. heparin and fucoidan) inhibited binding of soluble DC-HIL-Fc and adhesion of SVEC. By contrast, an integrin inhibitor (RGDS tetramer) had no effect on binding to SVEC, but prevented adhesion of SVEC. This differential RGD requirement was confirmed by the finding that DC-HIL-Fc mutant lacking the RGD motif can bind to SVEC but is unable to induce adhesion of SVEC. Furthermore, DC-HIL appears to recognize directly these sulfated polysaccharides. These results suggest that DC-HIL binds to SVEC by recognizing heparan sulfate proteoglycans on endothelial cells, thereby inducing adhesion of SVEC in an RGD-dependent manner. We propose that DC HIL serves as a DC-associated, heparan sulfate proteoglycan-dependent integrin ligand, which may be involved in transendothelial migration of DC. PMID- 11114301 TI - Membrane raft association of CD47 is necessary for actin polymerization and protein kinase C theta translocation in its synergistic activation of T cells. AB - CD47 is a ubiquitously expressed membrane protein with an extracellular Ig domain and a multiple membrane-spanning domain that can synergize with antigen to induce interleukin (IL)-2 secretion by T lymphocytes. Ligation of CD47 induced actin polymerization and increased protein kinase Ctheta (PKCtheta) association with the cytoskeleton independent of antigen receptor ligation, but ligation of mutant forms of the molecule missing either the Ig domain or the multiple membrane spanning domain did not. Simultaneous ligation of CD47 and CD3 led to additive effects on F-actin and synergistic effects on PKCtheta cytoskeletal association. Disruption of membrane rafts by removal of cholesterol with cyclodextrin blocked CD47-induced actin polymerization, and mutant forms of CD47 that localized poorly to rafts failed to effect cytoskeletal rearrangement. However, raft association alone was not sufficient, because a raft-localized CD47 Ig domain bound to the membrane by a glycan phosphoinositol anchor was unable to induce actin polymerization. A mutant form of CD47 without its Ig domain that did not induce actin polymerization or localize to rafts still enhanced T cell receptor (TCR) dependent tyrosine phosphorylation of PLCgamma and associated Ca(2+) signaling but did not augment IL-2 secretion. Thus, CD47 synergy with TCR to increase [Ca(2+)](i) is independent of actin and rafts but is insufficient to explain CD47 cooperation with TCR in IL-2 synthesis. Full synergy with TCR requires CD47 localization to membrane rafts where ligation leads to TCR-independent signals causing actin polymerization and PKCtheta translocation. PMID- 11114302 TI - Two novel residues in M2 of the gamma-aminobutyric acid type A receptor affecting gating by GABA and picrotoxin affinity. AB - An amino acid residue was found in M2 of gamma-aminobutyric acid (GABA) type A receptors that has profound effects on the binding of picrotoxin to the receptor and therefore may form part of its binding pocket. In addition, it strongly affects channel gating. The residue is located N-terminally to residues suggested so far to be important for channel gating. Point mutated alpha1beta(3) receptors were expressed in Xenopus oocytes and analyzed using the electrophysiological techniques. Coexpression of the alpha(1) subunit with the mutated beta(3) subunit beta(3)L253F led to spontaneous picrotoxin-sensitive currents in the absence of GABA. Nanomolar concentrations of GABA further promoted channel opening. Upon washout of picrotoxin, a huge transient inward current was observed. The reversal potential of the inward current was indicative of a chloride ion selectivity. The amplitude of the inward current was strongly dependent on the picrotoxin concentration and on the duration of its application. There was more than a 100 fold decrease in picrotoxin affinity. A kinetic model is presented that mimics the gating behavior of the mutant receptor. The point mutation in the neighboring residue beta(3)A252V resulted in receptors that displayed an about 6-fold increased apparent affinity to GABA and an about 10-fold reduced sensitivity to picrotoxin. PMID- 11114300 TI - Regulation of ROMK1 channels by protein-tyrosine kinase and -tyrosine phosphatase. AB - We have used the two-electrode voltage clamp technique and the patch clamp technique to investigate the regulation of ROMK1 channels by protein-tyrosine phosphatase (PTP) and protein-tyrosine kinase (PTK) in oocytes coexpressing ROMK1 and cSrc. Western blot analysis detected the presence of the endogenous PTP-1D isoform in the oocytes. Addition of phenylarsine oxide (PAO), an inhibitor of PTP, reversibly reduced K(+) current by 55% in oocytes coinjected with ROMK1 and cSrc. In contrast, PAO had no significant effect on K(+) current in oocytes injected with ROMK1 alone. Moreover, application of herbimycin A, an inhibitor of PTK, increased K(+) current by 120% and completely abolished the effect of PAO in oocytes coexpressing ROMK1 and cSrc. The effects of herbimycin A and PAO were absent in oocytes expressing the ROMK1 mutant R1Y337A in which the tyrosine residue at position 337 was mutated to alanine. However, addition of exogenous cSrc had no significant effect on the activity of ROMK1 channels in inside-out patches. Moreover, the effect of PAO was completely abolished by treatment of oocytes with 20% sucrose and 250 microg/ml concanavalin A, agents that inhibit the endocytosis of ROMK1 channels. Furthermore, the effect of herbimycin A is absent in the oocytes pretreated with either colchicine, an inhibitor of microtubules, or taxol, an agent that freezes microtubules. We conclude that PTP and PTK play an important role in regulating ROMK1 channels. Inhibiting PTP increases the internalization of ROMK1 channels, whereas blocking PTK stimulates the insertion of ROMK1 channels. PMID- 11114303 TI - Purification and characterization of A61. An angiostatin-like plasminogen fragment produced by plasmin autodigestion in the absence of sulfhydryl donors. AB - Plasmin, a broad spectrum proteinase, is inactivated by an autoproteolytic reaction that results in the destruction of the heavy and light chains of the protein. Recently we demonstrated that a 61-kDa plasmin fragment was one of the major products of this autoproteolytic reaction (Fitzpatrick, S. L., Kassam, G., Choi, K. S., Kang, H. M., Fogg, D. K., and Waisman, D. M. (2000)Biochemistry 39, 1021-1028). In the present communication we have identified the 61-kDa plasmin fragment as a novel four kringle-containing protein consisting of the amino acid sequence Lys(78)-Lys(468). To avoid confusion with the plasmin(ogen) fragment, angiostatin(R) (Lys(78)-Ala(440)), we have named this protein A(61). Unlike angiostatin, A(61) was produced in vitro from plasmin autodigestion in the absence of sulfhydryl donors. A(61) bound to lysine-Sepharose and also underwent a large increase in fluorescence yield upon binding of the lysine analogue, trans 4-aminomethylcyclohexanecarboxylic acid. Circular dichroism suggested that A(61) was composed of 21% beta-strand, 14% beta-turn, 18% 3(1)-helix and 8% 3(10) helix. A(61) was an anti-angiogenic protein as indicated by the inhibition of bovine capillary endothelial cell proliferation. Plasminogen was converted to A(61) by HT1080 cells and bovine capillary endothelial cells. Furthermore, a plasminogen fragment similar to A(61) was present in the serum of humans as well as normal and tumor-bearing mice. These results establish that plasmin turnover can generate anti-angiogenic plasmin fragments in a nonpathological setting. PMID- 11114304 TI - PU.1 is a lineage-specific regulator of tyrosine phosphatase CD45. AB - The hematopoietic cell-specific ets family transcription factor PU.1 regulates many lymphoid and myeloid genes. We have determined that PU.1 is critical for lineage-specific expression of the tyrosine phosphatase CD45. CD45 is expressed exclusively in hematopoietic cells at all stages of development, except for mature red cells and platelets. Although CD45 is normally expressed in all leukocyte lineages, it is critically regulated by PU.1 only in myeloid cells. Whereas myeloid cells from PU.1 null mice failed to express CD45, lymphoid cells were CD45(+) by flow cytometry. Additionally, mRNA for CD45 was absent from PU.1 deficient myeloid cells. To understand the molecular basis for these observations, we characterized a transcriptional regulatory region of the murine CD45 gene containing exons 1a, 1b, and 2. Distinct transcriptional initiation sites for CD45 were demonstrated in T and B cells versus myeloid cells. A transcriptional initiation site in exon 1b (P1b) was principally utilized by myeloid cells. A PU.1 binding site was identified upstream of exon 1b by sequence analysis and DNA binding assays. Using this region of the CD45 locus we demonstrated that PU.1 directly transactivated reporter gene expression. Finally, retrovirus-mediated restoration of PU.1 expression to PU.1-deficient myeloid cells resulted in expression of cell surface CD45 and restored phosphatase activity, confirming the role of PU.1 in the positive regulation of this well known signaling molecule. We conclude that CD45 is regulated differentially in myeloid and lymphoid cells and that sequences critical to direct myeloid expression include a PU.1 binding site upstream of the P1b transcriptional initiation site. PMID- 11114305 TI - Interleukin-6-induced tethering of STAT3 to the LAP/C/EBPbeta promoter suggests a new mechanism of transcriptional regulation by STAT3. AB - LAP/C/EBPbeta is a member of the C/EBP family of transcription factors and contributes to the regulation of the acute phase response in hepatocytes. Here we show that IL-6 controls LAP/C/EBPbeta gene transcription and identify an IL-6 responsive element in the LAP/C/EBPbeta promoter, which contains no STAT3 DNA binding motif. However, luciferase reporter gene assays showed that STAT3 activation through the gp130 signal transducer molecule is involved in mediating IL-6-dependent LAP/C/EBPbeta transcription. Southwestern analysis indicated that IL-6 induces binding of a 68-kDa protein to the recently characterized CRE-like elements in the LAP/C/EBPbeta promoter. Transfection experiments using promoter constructs with mutated CRE-like elements revealed that these sites confer IL-6 responsiveness. Further analysis using STAT1/STAT3 chimeras identified specific domains of the protein that are required for the IL-6-dependent increase in LAP/C/EBPbeta gene transcription. Overexpression of the amino-terminal domain of STAT3 blocked the IL-6-mediated response, suggesting that the STAT3 amino terminus has an important function in IL-6-mediated transcription of the LAP/C/EBPbeta gene. These data lead to a model of how tethering STAT3 to a DNA bound complex contributes to IL-6-dependent LAP/C/EBPbeta gene transcription. Our analysis describes a new mechanism by which STAT3 controls gene transcription and which has direct implication for the acute phase response in liver cells. PMID- 11114306 TI - Internal ribosome entry site-mediated translation of a mammalian mRNA is regulated by amino acid availability. AB - The cationic amino acid transporter, Cat-1, facilitates the uptake of the essential amino acids arginine and lysine. Amino acid starvation causes accumulation and increased translation of cat-1 mRNA, resulting in a 58-fold increase in protein levels and increased arginine uptake. A bicistronic mRNA expression system was used to demonstrate the presence of an internal ribosomal entry sequence (IRES) within the 5'-untranslated region of the cat-1 mRNA. This study shows that IRES-mediated translation of the cat-1 mRNA is regulated by amino acid availability. This IRES causes an increase in translation under conditions of amino acid starvation. In contrast, cap-dependent protein synthesis is inhibited during amino acid starvation, which is well correlated with decreased phosphorylation of the cap-binding protein, eIF4E. These findings reveal a new aspect of mammalian gene expression and regulation that provides a cellular stress response; when the nutrient supply is limited, the activation of IRES-mediated translation of mammalian mRNAs results in the synthesis of proteins essential for cell survival. PMID- 11114307 TI - ATM is required for IkappaB kinase (IKKk) activation in response to DNA double strand breaks. AB - Following challenge with proinflammatory stimuli or generation of DNA double strand breaks (DSBs), transcription factor NF-kappaB translocates from the cytoplasm to the nucleus to activate expression of target genes. In addition, NF kappaB plays a key role in protecting cells from proapoptotic stimuli, including DSBs. Patients suffering from the genetic disorder ataxia-telangiectasia, caused by mutations in the ATM gene, are highly sensitive to inducers of DSBs, such as ionizing radiation. Similar hypersensitivity is displayed by cell lines derived from ataxia-telangiectasia patients or Atm knockout mice. The ATM protein, a member of the phosphatidylinositol 3-kinase (PI3K)-like family, is a multifunctional protein kinase whose activity is stimulated by DSBs. As both ATM and NF-kappaB deficiencies result in increased sensitivity to DSBs, we examined the role of ATM in NF-kappaB activation. We report that ATM is essential for NF kappaB activation in response to DSBs but not proinflammatory stimuli, and this activity is mediated via the IkappaB kinase complex. DNA-dependent protein kinase, another member of the PI3K-like family, PI3K itself, and c-Abl, a nuclear tyrosine kinase, are not required for this response. PMID- 11114308 TI - Differential regulation of the uridine nucleotide-activated P2Y4 and P2Y6 receptors. SER-333 and SER-334 in the carboxyl terminus are involved in agonist dependent phosphorylation desensitization and internalization of the P2Y4 receptor. AB - Agonist-promoted regulation of the uridine nucleotide-activated human P2Y4 receptor (P2Y4-R) and P2Y6 receptor (P2Y6-R) was studied. Incubation of P2Y4-R expressing 1321N1 human astrocytoma cells with the cognate agonist UTP resulted in rapid desensitization of the inositol phosphate response and a 50% loss of cell surface receptors. In contrast, incubation of P2Y6-R-expressing cells with the cognate agonist UDP caused neither rapid desensitization nor rapid loss of cell surface receptors. Removal of UTP from the medium of UTP-pretreated cells resulted in rapid and complete recovery of surface P2Y4-R even after 12 h of agonist treatment. Although extended incubation with UDP also caused a loss of surface P2Y6-R, rapid recovery of surface P2Y6-R did not occur following removal of agonist. Pharmacological studies indicated that neither protein kinase C nor other Ca(2+)-activated kinases were involved in agonist-promoted desensitization or loss of surface P2Y4-R or P2Y6-R. Mutational analyses were carried out to identify domains involved in agonist-dependent regulation of P2Y4-R. Sequential truncation of the carboxyl-terminal domain revealed that sequence between amino acids 332 and 343 was necessary for UTP-promoted desensitization and internalization. Further mutational analyses of the three serines in this domain confirmed that Ser-333 and Ser-334 play a major role in these agonist-promoted changes in P2Y4-R. Experiments were carried out with [(32)P]P(i)-labeled cells to ascertain the role of phosphorylation in regulation of P2Y4-R. Incubation with UTP for 2 min caused a marked increase in phosphorylation of both the wild-type P2Y4-R and the P2Y4-343 truncation mutant. In contrast, no UTP-promoted phosphorylation of the P2Y4-332 truncation mutant was observed. Taken together, these results demonstrate differential regulation of uridine nucleotide-activated P2Y4-R and P2Y6-R and indicate that Ser-333 and Ser-334 in the carboxyl terminus of P2Y4-R are important for UTP-dependent phosphorylation, desensitization, and loss of surface receptors. PMID- 11114309 TI - Identification of novel, functional genetic variants in the human matrix metalloproteinase-2 gene: role of Sp1 in allele-specific transcriptional regulation. AB - Matrix metalloproteinase-2 (MMP-2) is an enzyme with proteolytic activity against matrix and nonmatrix proteins, particularly basement membrane constituents. Thus, any naturally occurring genetic variants that directly affect gene expression and/or protein function would be expected to impact on progression of pathological processes involving tissue remodeling. We scanned a 2-kilobase pair promoter region and all 13 exons of the human MMP-2 gene, from a panel of 32 individuals, and we identified the position, nature, and relative allele frequencies of 15 variant loci as follows: 6 in the promoter, 1 in the 5' untranslated region, 6 in the coding region, 1 in intronic sequence, and 1 in the 3'-untranslated region. The majority of coding region polymorphisms resulted in synonymous substitutions, whereas three promoter variants (at -1306, -790, and +220) mapped onto cis-acting elements. We functionally characterized all promoter variants by transient transfection experiments with 293, RAW264.7, and A10 cells. The common C --> T transition at -1306 (allele frequency 0.26), which disrupts an Sp1-type promoter site (CCACC box), displayed a strikingly lower promoter activity with the T allele. Electrophoretic mobility shift assays confirmed that these differences in allelic expression were attributable to abolition of Sp1 binding. These data suggest that this common functional genetic variant influences MMP-2 gene transcription in an allele-specific manner and is therefore an important candidate to test for association in a wide spectrum of pathologies for which a role for MMP-2 is implicated, including atherogenesis and tumor invasion and metastasis. PMID- 11114310 TI - The insect attractant 1-octen-3-ol is the natural ligand of bovine odorant binding protein. AB - Bovine odorant-binding protein (bOBP) is a dimeric lipocalin present in large amounts in the respiratory and olfactory nasal mucosa. The structure of bOBP refined at 2.0-A resolution revealed an elongated volume of electron density inside each buried cavity, indicating the presence of one (or several) naturally occurring copurified ligand(s) (Tegoni et al. (1996) Nat. Struct. Biol. 3, 863 867; Bianchet et al. (1996) Nat. Struct. Biol. 3, 934-939). In the present work, by combining mass spectrometry, x-ray crystallography (1.8-A resolution), and fluorescence, it has been unambiguously established that natural bOBP contains the racemic form of 1-octen-3-ol. This volatile substance is a typical component of bovine breath and in general of odorous body emanations of humans and animals. The compound 1-octen-3-ol is also an extremely potent olfactory attractant for many insect species, including some parasite vectors like Anopheles (Plasmodium) or Glossina (Trypanosoma). For the first time, a function can be assigned to an OBP, with a possible role of bOBP in the ecological relationships between bovine and insect species. PMID- 11114311 TI - Stable insertion of the early light-induced proteins into etioplast membranes requires chlorophyll a. AB - Etiolated plant seedlings exposed to light respond by transient accumulation of the nucleus-encoded, plastid-located early light-inducible proteins (Elips). These proteins are distant relatives of the light-harvesting chlorophyll a/b binding gene family and bind pigments with unusual characteristics. To investigate whether accumulation of Elips in plastid membranes is post translationally regulated by pigments, reconstitution studies were performed, where in vitro transcribed and translated low molecular mass Elip precursors of barley were combined with lysed barley etioplasts complemented with various compositions of isolated pigments. We showed that the membrane insertion of Elips, as proven by protease protection assays and washes with a chaotropic salt or alkali, depended strictly on chlorophyll a but not on chlorophyll b or xanthophyll zeaxanthin. The amount of inserted Elips increased almost linearly with the chlorophyll a concentration, and the insertion efficiency was not significantly influenced by a light intensity between 1 and 1,000 micromol x m( 2) x s(-1). In contrast, in vitro import of Elip precursors into greening plastids was enhanced by high intensity light. Thus, we conclude that although chlorophylls bound to Elips seem to not be involved in light harvesting, they are crucial for a stable insertion of these proteins into the plastid membrane. PMID- 11114312 TI - Fine particulate air pollution and mortality in 20 U.S. cities, 1987-1994. AB - BACKGROUND: Air pollution in cities has been linked to increased rates of mortality and morbidity in developed and developing countries. Although these findings have helped lead to a tightening of air-quality standards, their validity with respect to public health has been questioned. METHODS: We assessed the effects of five major outdoor-air pollutants on daily mortality rates in 20 of the largest cities and metropolitan areas in the United States from 1987 to 1994. The pollutants were particulate matter that is less than 10 microm in aerodynamic diameter (PM10), ozone, carbon monoxide, sulfur dioxide, and nitrogen dioxide. We used a two-stage analytic approach that pooled data from multiple locations. RESULTS: After taking into account potential confounding by other pollutants, we found consistent evidence that the level of PM10 is associated with the rate of death from all causes and from cardiovascular and respiratory illnesses. The estimated increase in the relative rate of death from all causes was 0.51 percent (95 percent posterior interval, 0.07 to 0.93 percent) for each increase in the PM10 level of 10 microg per cubic meter. The estimated increase in the relative rate of death from cardiovascular and respiratory causes was 0.68 percent (95 percent posterior interval, 0.20 to 1.16 percent) for each increase in the PM10 level of 10 microg per cubic meter. There was weaker evidence that increases in ozone levels increased the relative rates of death during the summer, when ozone levels are highest, but not during the winter. Levels of the other pollutants were not significantly related to the mortality rate. CONCLUSIONS: There is consistent evidence that the levels of fine particulate matter in the air are associated with the risk of death from all causes and from cardiovascular and respiratory illnesses. These findings strengthen the rationale for controlling the levels of respirable particles in outdoor air. PMID- 11114313 TI - Fludarabine compared with chlorambucil as primary therapy for chronic lymphocytic leukemia. AB - BACKGROUND: Fludarabine is an effective treatment for chronic lymphocytic leukemia that does not respond to initial treatment with chlorambucil. We compared the efficacy of fludarabine with that of chlorambucil in the primary treatment of chronic lymphocytic leukemia. METHODS: Between 1990 and 1994, we randomly assigned 509 previously untreated patients with chronic lymphocytic leukemia to one of the following treatments: fludarabine (25 mg per square meter of body-surface area, administered intravenously daily for 5 days every 28 days), chlorambucil (40 mg per square meter, given orally every 28 days), or fludarabine (20 mg per square meter per day for 5 days every 28 days) plus chlorambucil (20 mg per square meter every 28 days). Patients with an additional response at each monthly evaluation continued to receive the assigned treatment for a maximum of 12 cycles. RESULTS: Assignment of patients to the fludarabine-plus-chlorambucil group was stopped when a planned interim analysis revealed excessive toxicity and a response rate that was not better than the rate with fludarabine alone. Among the other two groups, the response rate was significantly higher for fludarabine alone than for chlorambucil alone. Among 170 patients treated with fludarabine, 20 percent had a complete remission, and 43 percent had a partial remission. The corresponding values for 181 patients treated with chlorambucil were 4 percent and 33 percent (P< 0.001 for both comparisons). The median duration of remission and the median progression-free survival in the fludarabine group were 25 months and 20 months, respectively, whereas both values were 14 months in the chlorambucil group (P<0.001 for both comparisons). The median overall survival among patients treated with fludarabine was 66 months, which was not significantly different from the overall survival in the other two groups (56 months with chlorambucil and 55 months with combined treatment). Severe infections and neutropenia were more frequent with fludarabine than with chlorambucil (P=0.08), although, overall, toxic effects were tolerable with the two single-drug regimens. CONCLUSIONS: When used as the initial treatment for chronic lymphocytic leukemia, fludarabine yields higher response rates and a longer duration of remission and progression-free survival than chlorambucil. PMID- 11114314 TI - Predictors of rehospitalization for symptomatic venous thromboembolism after total hip arthroplasty. AB - BACKGROUND: Recent studies have shown that symptomatic venous thromboembolism after total hip arthroplasty most commonly develops after the patient is discharged from the hospital. Risk factors associated with these symptomatic thromboembolic events are not well defined. METHODS: Using administrative data from the California Medicare records for 1993 through 1996, we identified 297 patients 65 years of age or older who were rehospitalized for thromboembolism within three months after total hip arthroplasty. We compared demographic, surgical, and medical variables potentially associated with the development of thromboembolism in these patients and 592 unmatched controls. RESULTS: A total of 89.6 percent of patients with thromboembolism and 93.8 percent of control patients were treated with pneumatic compression, warfarin, enoxaparin, or unfractionated heparin, alone or in combination. In addition, 22.2 percent and 29.7 percent, respectively, received warfarin after discharge. A body-mass index (the weight in kilograms divided by the square of the height in meters) of 25 or greater was associated with rehospitalization for thromboembolism, with an odds ratio of 2.5 (95 percent confidence interval, 1.8 to 3.4). In a multivariate model, the only prophylactic regimens associated with a reduced risk of thromboembolism were pneumatic compression in patients with body-mass indexes of less than 25 (odds ratio, 0.3; 95 percent confidence interval, 0.2 to 0.6) and warfarin treatment after discharge (odds ratio, 0.6; 95 percent confidence interval, 0.4 to 1.0). CONCLUSIONS: In patients who underwent total hip arthroplasty, a body-mass index of 25 or greater was associated with subsequent hospitalization for thromboembolism. Pneumatic compression in patients with a body-mass index of less than 25 and prophylaxis with warfarin after discharge were independently protective against thromboembolism. PMID- 11114315 TI - Familial aggregation of Parkinson's disease in Iceland. AB - BACKGROUND: The role of genetics in early-onset Parkinson's disease has been established, but whether there is a genetic contribution to the more common, late onset form remains uncertain. METHODS: We reviewed the medical records and confirmed the diagnosis of Parkinson's disease in 772 living and deceased patients in whom the disease had been diagnosed during the previous 50 years in Iceland. With the use of an extensive computerized data base containing genealogic information on 610,920 people in Iceland during the past 11 centuries, several analyses were conducted to determine whether the patients were more related to each other than random members of the population (control subjects). RESULTS: Patients with Parkinson's disease, including a subgroup of 560 patients with late-onset disease (onset at >50 years of age), were significantly more related to each other than were subjects in matched groups of controls, and this relatedness extended beyond the nuclear family. The risk ratio for Parkinson's disease was 6.7 (95 percent confidence interval, 4.3 to 9.6) for siblings, 3.2 (95 percent confidence interval, 1.2 to 7.8) for offspring, and 2.7 (95 percent confidence interval, 1.6 to 3.9) for nephews and nieces of patients with late onset Parkinson's disease. CONCLUSIONS: Late-onset Parkinson's disease has a genetic component as well as an environmental component. PMID- 11114316 TI - Images in clinical medicine. Mediastinal abnormalities in systemic sclerosis. PMID- 11114317 TI - Association of the California Tobacco Control Program with declines in cigarette consumption and mortality from heart disease. AB - BACKGROUND: The California Tobacco Control Program, a large, aggressive antitobacco program implemented in 1989 and funded by a voter-enacted cigarette surtax, accelerated the decline in cigarette consumption and in the prevalence of smoking in California. Since the excess risk of heart disease falls rapidly after the cessation of smoking, we tested the hypothesis that this program was associated with lower rates of death from heart disease. METHODS: Data on per capita cigarette consumption and age-adjusted rates of death from heart disease in California and the United States from 1980 to 1997 were fitted in multiple regression analyses. The regression analyses included the rates in the rest of the United States and variables that allowed for changes in the rates after 1988, when the tobacco-control program was approved, and after 1992, when the program was cut back. RESULTS: Between 1989 and 1992, the rates of decline in per capita cigarette consumption and mortality from heart disease in California, relative to the rest of the United States, were significantly greater than the pre-1989 rates, by 2.72 packs per year per year (P = 0.001) and by 2.93 deaths per year per 100,000 population per year (P<0.001). These rates of decline were reduced (by 2.05 packs per year per year, [P=0.04], and by 1.71 deaths per year per 100,000 population per year, [P=0.031) when the program was cut back, beginning in 1992. Despite these problems, the program was associated with 33,300 fewer deaths from heart disease between 1989 and 1997 than the number that would have been expected if the earlier trend in mortality from heart disease in California relative to the rest of the United States had continued. The diminished effectiveness of the program after 1992 was associated with 8300 more deaths than would have been expected had its initial effectiveness been maintained. CONCLUSIONS: A large and aggressive tobacco-control program is associated with a reduction in deaths from heart disease in the short run. PMID- 11114318 TI - Prevention and treatment of influenza. PMID- 11114319 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 38-2000. A 45-year-old woman with exertional dyspnea, hemoptysis, and pulmonary nodules. PMID- 11114320 TI - Particulate air pollution and mortality--clearing the air. PMID- 11114321 TI - When and how to treat chronic lymphocytic leukemia. PMID- 11114322 TI - Tobacco, the Food and Drug Administration, and Congress. PMID- 11114323 TI - Protecting the public health by strengthening the Food and Drug Administration's authority over tobacco products. PMID- 11114324 TI - p53: death star. PMID- 11114325 TI - Bugs on drugs go GAGAA. PMID- 11114326 TI - Unequal access: regulating V(D)J recombination through chromatin remodeling. PMID- 11114327 TI - RNA processing marches on. PMID- 11114328 TI - Evolutionary implications of the frequent horizontal transfer of mismatch repair genes. AB - Mutation and subsequent recombination events create genetic diversity, which is subjected to natural selection. Bacterial mismatch repair (MMR) deficient mutants, exhibiting high mutation and homologous recombination rates, are frequently found in natural populations. Therefore, we have explored the possibility that MMR deficiency emerging in nature has left some "imprint" in the sequence of bacterial genomes. Comparative molecular phylogeny of MMR genes from natural Escherichia coli isolates shows that, compared to housekeeping genes, individual functional MMR genes exhibit high sequence mosaicism derived from diverse phylogenetic lineages. This apparent horizontal gene transfer correlates with hyperrecombination phenotype of MMR-deficient mutators. The sequence mosaicism of MMR genes may be a hallmark of a mechanism of adaptive evolution that involves modulation of mutation and recombination rates by recurrent losses and reacquisitions of MMR gene functions. PMID- 11114329 TI - Adaptive amplification: an inducible chromosomal instability mechanism. AB - Adaptive mutation is an induced response to environmental stress in which mutation rates rise, producing permanent genetic changes that can adapt cells to stress. This contrasts with neo-Darwinian views of genetic change rates blind to environmental conditions. DNA amplification is a flexible, reversible genomic change that has long been postulated to be adaptive. We report the discovery of adaptive amplification at the lac operon in Escherichia coli. Additionally, we find that adaptive amplification is separate from, and does not lead to, adaptive point mutation. This contradicts a prevailing alternative hypothesis whereby adaptive mutation is normal mutability in amplified DNA. Instead, adaptive mutation and amplification are parallel routes of inducible genetic instability allowing rapid evolution under stress, and escape from growth inhibition. PMID- 11114330 TI - Transcription factor dosage affects changes in higher order chromatin structure associated with activation of a heterochromatic gene. AB - The mechanisms of transcriptional activation in heterochromatin were investigated by using FISH to directly visualize changes in chromatin organization during activation of a heterochromatic lambda5 transgene. A DNase I hypersensitive site was shown to relocate the transgene to the outside of the pericentromeric heterochromatin complex in the absence of transcription. Activation of transcription, which is dependent on the transcription factor EBF, occurs in a stochastic manner that resembles telomeric silencing in yeast, with the transcribed gene remaining closely associated with the heterochromatin complex. Reducing the dosage of EBF results in a reduced frequency of localization of the transgene to the outside of the heterochromatin complex and lower levels of transcription. These data provide evidence that transcription factors can initiate changes in higher order chromatin structure during the earliest stages of gene activation. PMID- 11114331 TI - c-Jun and JunB antagonistically control cytokine-regulated mesenchymal-epidermal interaction in skin. AB - Interactions between mesenchymal and epithelial cells are responsible for organogenesis and tissue homeostasis. This mutual cross-talk involves cell surface proteins and soluble factors, which are mostly the result of regulated transcription. To elucidate dimer-specific functions of the AP-1 family of transcription factors, we reconstituted skin by combining primary human keratinocytes and mouse wild-type, c-jun(-/-), and junB(-/-) fibroblasts. We have discovered an antagonistic function of these AP-1 subunits in the fibroblast mediated paracrine control of keratinocyte proliferation and differentiation, and traced this effect to the IL-1-dependent regulation of KGF and GM-CSF. These data suggest that the relative activation state of these AP-1 subunits in a non-cell autonomous, transregulatory fashion directs regeneration of the epidermis and maintenance of tissue homeostasis in skin. PMID- 11114332 TI - The leukotriene C(4) transporter MRP1 regulates CCL19 (MIP-3beta, ELC)-dependent mobilization of dendritic cells to lymph nodes. AB - Adaptive immune responses begin after antigen-bearing dendritic cells (DCs) traffic from peripheral tissues to lymph nodes. Here, we show that DC migration from skin to lymph nodes utilizes the leukotriene C(4) (LTC(4)) transporter multidrug resistance-associated protein 1 (MRP1). DC mobilization from the epidermis and trafficking into lymphatic vessels was greatly reduced in MRP1(-/-) mice, but migration was restored by exogenous cysteinyl leukotrienes LTC(4) or LTD(4). In vitro, these cysteinyl leukotrienes promoted optimal chemotaxis to the chemokine CCL19, but not to other related chemokines. Antagonism of CCL19 in vivo prevented DC migration out of the epidermis. Thus, MRP-1 regulates DC migration to lymph nodes, apparently by transporting LTC(4), which in turn promotes chemotaxis to CCL19 and mobilization of DCs from the epidermis. PMID- 11114333 TI - Transmembrane electron transfer by the membrane protein DsbD occurs via a disulfide bond cascade. AB - The cytoplasmic membrane protein DsbD transfers electrons from the cytoplasm to the periplasm of E. coli, where its reducing power is used to maintain cysteines in certain proteins in the reduced state. We split DsbD into three structural domains, each containing two essential cysteines. Remarkably, when coexpressed, these truncated proteins restore DsbD function. Utilizing this three piece system, we were able to determine a pathway of the electrons through DsbD. Our findings strongly suggest that the pathway is based on a series of multistep redox reactions that include direct interactions between thioredoxin and DsbD, and between DsbD and its periplasmic substrates. A thioredoxin-fold domain in DsbD appears to have the novel role of intramolecular electron shuttle. PMID- 11114334 TI - X-Ray structures of the universal translation initiation factor IF2/eIF5B: conformational changes on GDP and GTP binding. AB - X-ray structures of the universal translation initiation factor IF2/eIF5B have been determined in three states: free enzyme, inactive IF2/eIF5B.GDP, and active IF2/eIF5B.GTP. The "chalice-shaped" enzyme is a GTPase that facilitates ribosomal subunit joining and Met-tRNA(i) binding to ribosomes in all three kingdoms of life. The conserved core of IF2/eIF5B consists of an N-terminal G domain (I) plus an EF-Tu-type beta barrel (II), followed by a novel alpha/beta/alpha-sandwich (III) connected via an alpha helix to a second EF-Tu-type beta barrel (IV). Structural comparisons reveal a molecular lever, which amplifies a modest conformational change in the Switch 2 region of the G domain induced by Mg(2+)/GTP binding over a distance of 90 A from the G domain active center to domain IV. Mechanisms of GTPase function and ribosome binding are discussed. PMID- 11114335 TI - Structural basis for double-sieve discrimination of L-valine from L-isoleucine and L-threonine by the complex of tRNA(Val) and valyl-tRNA synthetase. AB - Valyl-tRNA synthetase (ValRS) strictly discriminates the cognate L-valine from the larger L-isoleucine and the isosteric L-threonine by the tRNA-dependent "double sieve" mechanism. In this study, we determined the 2.9 A crystal structure of a complex of Thermus thermophilus ValRS, tRNA(Val), and an analog of the Val-adenylate intermediate. The analog is bound in a pocket, where Pro(41) allows accommodation of the Val and Thr moieties but precludes the Ile moiety (the first sieve), on the aminoacylation domain. The editing domain, which hydrolyzes incorrectly synthesized Thr-tRNA(Val), is bound to the 3' adenosine of tRNA(Val). A contiguous pocket was found to accommodate the Thr moiety, but not the Val moiety (the second sieve). Furthermore, another Thr binding pocket for Thr-adenylate hydrolysis was suggested on the editing domain. PMID- 11114336 TI - The amnesiac gene product is expressed in two neurons in the Drosophila brain that are critical for memory. AB - Mutations in the amnesiac gene in Drosophila affect both memory retention and ethanol sensitivity. The predicted amnesiac gene product, AMN, is an apparent preproneuropeptide, and previous studies suggest that it stimulates cAMP synthesis. Here we show that, unlike other learning-related Drosophila proteins, AMN is not preferentially expressed in mushroom bodies. Instead, it is strongly expressed in two large neurons that project over all the lobes of the mushroom bodies, a finding that suggests a modulatory role for AMN in memory formation. Genetically engineered blockade of vesicle recycling in these cells abbreviates memory as in the amnesiac mutant. Moreover, restoration of amn gene expression to these cells reestablishes normal olfactory memory in an amn deletion background. These results indicate that AMN neuropeptide release onto the mushroom bodies is critical for normal olfactory memory. PMID- 11114337 TI - The C termini of Arabidopsis cryptochromes mediate a constitutive light response. AB - Cryptochrome blue light photoreceptors share sequence similarity to photolyases, flavoproteins that mediate light-dependent DNA repair. However, cryptochromes lack photolyase activity and are characterized by distinguishing C-terminal domains. Here we show that the signaling mechanism of Arabidopsis cryptochrome is mediated through the C terminus. On fusion with beta-glucuronidase (GUS), both the Arabidopsis CRY1 C-terminal domain (CCT1) and the CRY2 C-terminal domain (CCT2) mediate a constitutive light response. This constitutive photomorphogenic (COP) phenotype was not observed for mutants of cct1 corresponding to previously described cry1 alleles. We propose that the C-terminal domain of Arabidopsis cryptochrome is maintained in an inactive state in the dark. Irradiation with blue light relieves this repression, presumably through an intra- or intermolecular redox reaction mediated through the flavin bound to the N-terminal photolyase-like domain. PMID- 11114371 TI - HFE--a novel nonclassical class I molecule that is involved in iron metabolism. PMID- 11114372 TI - DNA double-strand breaks in immunoglobulin genes undergoing somatic hypermutation. AB - How rearranged immunoglobulin (Ig) genes are further diversified by somatic hypermutation is unknown. Using VDJ passenger Ig heavy chain (IgH) knockin mouse strains, we now demonstrate a high frequency of DNA double-strand breaks (DSBs) in the targeted VDJ passenger gene of germinal center (GC) B cells. These DSBs parallel the distribution of mutations in the targeted hypermutation domain and are found preferentially at RGYW motifs, the intrinsic hot spots of somatic hypermutation. The introduction of DSBs appears to depend on transcriptional activity. Thus, secondary diversification of rearranged V gene segments relates to an error-prone nonhomologous DSB repair system acting in B cells of the GC. PMID- 11114373 TI - Control of B cell production by the adaptor protein lnk. Definition Of a conserved family of signal-modulating proteins. AB - Lnk is an SH2 domain-containing adaptor protein expressed preferentially in lymphocytes. To illuminate the importance of Lnk, we generated lnk(-/-) mice. Whereas T cell development was unaffected, pre-B and immature B cells accumulated in the spleens. In the bone marrow, B-lineage cells were proportionately increased, reflecting enhanced production of pro-B cells that resulted in part from hypersensitivity of precursors to SCF, the ligand for c-kit. Hence, Lnk ordinarily acts to regulate B cell production. Further characterization of lnk(-/ ) mice also revealed that full-length Lnk is a 68 kDa protein containing a conserved proline-rich region and a PH domain. Lnk is a representative of a multigene adaptor protein family whose members act, by analogy with Lnk, to modulate intracellular signaling. PMID- 11114374 TI - Maturation of an antibody response is governed by modulations in flexibility of the antigen-combining site. AB - Although affinity maturation constitutes an integral part of T-dependent humoral responses, its structural basis is less well understood. We compared the physicochemical properties of antigen binding of several independent antibody panels derived from both germline and secondary responses. We found that antibody maturation essentially reflects modulations in entropy-control of the association, but not dissociation, step of the binding. This influence stems from variations in conformational heterogeneity of the antigen-combining site, which in turn regulates both the affinity and specificity for antigen. Thus, the simple device of manipulating conformational flexibility of paratope provides a mechanism wherein the transition from a degenerate recognition capability to a high-fidelity effector response is readily achieved, with the minimum of somatic mutations. PMID- 11114375 TI - Identification of CD72 as a lymphocyte receptor for the class IV semaphorin CD100: a novel mechanism for regulating B cell signaling. AB - We have identified the lymphocyte semaphorin CD100/Sema4D as a CD40-inducible molecule by subtractive cDNA cloning. CD100 stimulation significantly enhanced the effects of CD40 on B cell responses. Administration of soluble CD100 markedly accelerated in vivo antigen-specific antibody responses. CD100 receptors with different binding affinities were detected on renal tubular cells (K(d) = approximately 1 x 10(-9)M) and lymphocytes (K(d) = approximately 3 x 10(-7)M). Expression cloning revealed that the CD100 receptor on lymphocytes is CD72, a negative regulator of B cell responsiveness. CD72 thus represents a novel class of semaphorin receptors. CD100 stimulation induced tyrosine dephosphorylation of CD72 and dissociation of SHP-1 from CD72. Our findings indicate that CD100 plays a critical role in immune responses by the novel mechanism of turning off negative signaling by CD72. PMID- 11114376 TI - The class IV semaphorin CD100 plays nonredundant roles in the immune system: defective B and T cell activation in CD100-deficient mice. AB - The class IV semaphorin CD100/Sema4D differentially utilizes two distinct receptors: plexin-B1 in nonlymphoid tissues, such as brain and kidney, and CD72 in lymphoid tissues. We have generated CD100-deficient mice and demonstrated that they have functional defects in their immune system, without apparent abnormalities in other tissues. The number of CD5(+) B-1 cells was considerably decreased in the mutant mice, whereas conventional B cells and T cells appeared to develop normally. In vitro proliferative responses and immunoglobulin production were reduced in CD100-deficient B cells. The humoral immune response against a T cell-dependent antigen and in vivo priming of T cells were also defective in the mutant mice. These results demonstrate nonredundant and essential roles of CD100-CD72 interactions in the immune system. PMID- 11114377 TI - CD8(+) T cell-mediated skin disease in mice lacking IRF-2, the transcriptional attenuator of interferon-alpha/beta signaling. AB - The balanced action of cytokines is known to be critical for the maintenance of homeostatic immune responses. Here, we report the development of an inflammatory skin disease involving CD8(+) T cells, in mice lacking the transcription factor, interferon regulatory factor-2 (IRF-2). CD8(+) T cells exhibit in vitro hyper responsiveness to antigen stimulation, accompanied with a notable upregulation of the expression of genes induced by interferon-alpha/beta (IFN-alpha/beta). Furthermore, both disease development and CD8(+) T cell abnormality are suppressed by the introduction of nullizygosity to the genes that positively regulate the IFN-alpha/beta signaling pathway. IRF-2 may represent a unique negative regulator, attenuating IFN-alpha/beta-induced gene transcription, which is necessary for balancing the beneficial and harmful effects of IFN-alpha/beta signaling in the immune system. PMID- 11114378 TI - Fratricide among CD8(+) T lymphocytes naturally infected with human T cell lymphotropic virus type I. AB - Infection and gene expression by the human T lymphotropic virus type I (HTLV-I) in vivo have been thought to be confined to CD4(+) T lymphocytes. We show here that, in natural HTLV-I infection, a significant proportion of CD8(+) T lymphocytes are infected by HTLV-I. Interestingly, HTLV-I-specific but not Epstein-Barr virus-specific CD8(+) T lymphocytes were shown to be infected. Furthermore, HTLV-I protein expression in naturally infected CD8(+) T lymphocytes renders them susceptible to fratricide mediated by autologous HTLV-I-specific CD8(+) T lymphocytes. Fratricide among virus-specific CTLs could impair the immune control of HTLV-I and possibly other lymphotropic viruses. PMID- 11114379 TI - On the dynamics of TCR:CD3 complex cell surface expression and downmodulation. AB - TCR downmodulation following ligation by MHC:peptide complexes is considered to be a pivotal event in T cell activation. Here, we analyzed the dynamics of TCR:CD3 cell surface expression on resting and antigen-activated T cells. We show that the TCR:CD3 complex is very stable and is rapidly internalized and recycled in resting T cells. Surprisingly, the internalization rate is not increased following TCR ligation by MHC:peptide complexes, despite significant TCR downmodulation, suggesting that constitutive internalization rather than ligation induced downmodulation serves as the force that drives serial ligation. Furthermore, TCR downmodulation is mediated by the intracellular retention of ligated complexes and degradation by lysosomes and proteasomes. Thus, our data demonstrate that ligation induces TCR downmodulation by preventing recycling rather than inducing internalization. PMID- 11114380 TI - NF-kappa B activation by the pre-T cell receptor serves as a selective survival signal in T lymphocyte development. AB - Activation of the transcription factor NF-kappa B and pre-T cell receptor (pre TCR) expression is tightly correlated during thymocyte development. Inhibition of NF-kappa B in isolated thymocytes in vitro results in spontaneous apoptosis of cells expressing the pre-TCR, whereas inhibition of NF-kappa B in transgenic mice through expression of a mutated, superrepressor form of I kappa B alpha leads to a loss of beta-selected thymocytes. In contrast, the forced activation of NF kappa B through expression of a dominant-active I kappa B kinase allows differentiation to proceed to the CD4(+)CD8(+) stage in a Rag1(-/-) mouse that cannot assemble the pre-TCR. Therefore, signals emanating from the pre-TCR are mediated at least in part by NF-kappa B, which provides a selective survival signal for developing thymocytes with productive beta chain rearrangements. PMID- 11114381 TI - Gut cryptopatches: direct evidence of extrathymic anatomical sites for intestinal T lymphopoiesis. AB - Athymic cytokine receptor gamma chain mutant mice that lack the thymus, Peyer's patches, cryptopatches (CP), and intestinal T cells were reconstituted with wild type bone marrow cells. Bone marrow-derived TCR(-) intraepithelial lymphocytes (IEL) first appeared within villous epithelia of small intestine overlying the regenerated CP, and these TCR(-) IEL subsequently emerged throughout the epithelia. Thereafter, TCR(+) IEL increased to a comparable number to that in athymic mice and consisted of TCRgammadelta and TCRalphabeta IEL. In gut associated lymphoid tissues of wild-type mice, only CP harbored a large population of c-kit(high)IL-7R(+)CD44(+)Thy-1(+/-)CD4(+/-)CD25(low/-)alpha(E) beta(7)(-)Lin(-) (Lin, lineage markers) lymphocytes that included cells expressing germline but not rearranged TCRgamma and TCRbeta gene transcripts. These findings provide direct evidence that gut CP develop progenitor T cells for extrathymic IEL descendants. PMID- 11114382 TI - Deletion of germline promoter PD beta 1 from the TCR beta locus causes hypermethylation that impairs D beta 1 recombination by multiple mechanisms. AB - The role of the germline transcriptional promoter, PD beta 1, in V(D)J recombination at the T cell receptor beta locus was investigated. Deletion of PD beta 1 caused reduced germline transcription and DNA hypermethylation in the Dbeta1-J beta 1 region and decreased D beta 1 rearrangement. Analyses of methylation levels surrounding recombination signal sequences (RSS) before, during, and after recombination revealed that under physiological conditions cleavage of hypomethylated alleles was preferred over hypermethylated alleles. Methylation of a specific CpG site within the heptamer of the 3' D beta 1 RSS was incompatible with cleavage by the V(D)J recombinase. These findings suggest that methylation can regulate V(D)J recombination both at a general level by influencing regional chromatin accessibility and specifically by blocking RSS recognition or cleavage by the V(D)J recombinase. PMID- 11114383 TI - Novel p19 protein engages IL-12p40 to form a cytokine, IL-23, with biological activities similar as well as distinct from IL-12. AB - A novel sequence discovered in a computational screen appears distantly related to the p35 subunit of IL-12. This factor, which we term p19, shows no biological activity by itself; instead, it combines with the p40 subunit of IL-12 to form a novel, biologically active, composite cytokine, which we term IL-23. Activated dendritic cells secrete detectable levels of this complex. IL-23 binds to IL-12R beta 1 but fails to engage IL-12R beta 2; nonetheless, IL-23 activates Stat4 in PHA blast T cells. IL-23 induces strong proliferation of mouse memory (CD4(+)CD45Rb(low)) T cells, a unique activity of IL-23 as IL-12 has no effect on this cell population. Similar to IL-12, human IL-23 stimulates IFN-gamma production and proliferation in PHA blast T cells, as well as in CD45RO (memory) T cells. PMID- 11114384 TI - Crystal structure and ligand binding properties of the D1D2 region of the inhibitory receptor LIR-1 (ILT2). AB - LIR-1 is an inhibitory receptor that recognizes class I MHC molecules and the human cytomegalovirus class I homolog UL18. Here, we report the 2.1 A resolution crystal structure of the ligand binding portion of LIR-1 (domains 1 and 2 [D1D2]) and localize the binding region for UL18. LIR-1 D1D2 is composed of two immunoglobulin-like domains arranged at an acute angle to form a bent structure resembling the structures of natural killer inhibitory receptors (KIRs). The LIR 1 binding site comprises a portion of D1 distant from the interdomain hinge region that constitutes the KIR binding site, consistent with differences in LIR 1 and KIR recognition properties and functions. PMID- 11114385 TI - Tissue-specific inducible expression of antimicrobial peptide genes in Drosophila surface epithelia. AB - The production of antimicrobial peptides is an important aspect of host defense in multicellular organisms. In Drosophila, seven antimicrobial peptides with different spectra of activities are synthesized by the fat body during the immune response and secreted into the hemolymph. Using GFP reporter transgenes, we show here that all seven Drosophila antimicrobial peptides can be induced in surface epithelia in a tissue-specific manner. The imd gene plays a critical role in the activation of this local response to infection. In particular, drosomycin expression, which is regulated by the Toll pathway during the systemic response, is regulated by imd in the respiratory tract, thus demonstrating the existence of distinct regulatory mechanisms for local and systemic induction of antimicrobial peptide genes in Drosophila. PMID- 11114386 TI - Mitochondrial genomes of human helminths and their use as markers in population genetics and phylogeny. AB - To date, over 100 complete metazoan mitochondrial (mt) genomes of different phyla have been reported. Here, we briefly summarise mt gene organisation in the Metazoa and review what is known of the mt genomes of nematodes and flatworms parasitic in humans. The availability of complete or almost complete mtDNA sequences for several parasitic helminths provides a rich source of genetic markers for phylogenetic analysis and study of genetic variability in helminth groups. Examples of the application of mtDNA in studies on Ascaris, Onchocerca, Schistosoma, Fasciola, Paragonimus, Echinostoma, Echinococcus and Taenia are described. PMID- 11114387 TI - Spur cell anaemia and acute haemolysis in patients with hyperreactive malarious splenomegaly. Experience in an isolated Yanomamo population of Venezuela. AB - A prospective study, aimed to investigate the aetiology of an unusual clustering of cases of severe acute haemolytic anaemia affecting a high percentage of the adult population, was carried out in two isolated Yanomamo communities of the Upper Orinoco basin in Venezuela. Twenty-six patients with active or recent episodes of severe haemolysis were evaluated. All of them exhibited massive liver and spleen enlargement and fulfilled the diagnostic criteria of the hyperreactive malarious splenomegaly (HMS) syndrome. In four cases with advanced non-alcohol related chronic liver disease, hypersplenism, severe haemolytic anaemia and acanthocytosis, the characteristic clinical and laboratory findings of spur cell anaemia were documented. Chronic infection by the HBV and HCV was present in three of them. However, in most of the 22 additional HMS cases, the acute haemolytic condition appeared associated with the occurrence of a cold agglutinin mediated autoimmune response. The clustering of a significant number of cases of severe acute haemolysis in HMS patients from this small isolated aboriginal community is most unusual, and represents a serious complicating factor for a population already beleaguered by a high prevalence of malaria due to multiresistant strains of Plasmodium falciparum. Moreover, the coexistence of HMS and severe chronic HBV or HCV infection may further aggravate the course of the haemolytic disorder, because of the occurrence of spur cell anaemia. PMID- 11114388 TI - The decline of anti-trypanosomal antibody levels in cattle after treatment with trypanocidal drugs and in the absence of tsetse challenge. AB - The decline of anti-trypanosomal antibody levels in cattle after treatment with trypanocidal drugs was investigated using an anti-trypanosomal antibody-detection enzyme-linked immunosorbent assay (ELISA). The decline of antibody levels differed between experimental animals but went through two phases. During the first 5 months after trypanocidal drug treatment, the decline is rapid with a monthly average decline of 10% of the average percentage positivity during the treatment. Between months 6 and 13 after treatment, the monthly average decline was only 3.6% of the average percentage positivity at the moment of treatment. It took a total of 13 months for all the experimental animals to become seronegative. The usefulness of the anti-trypanosomal antibody-detection enzyme linked immunosorbent assay (ELISA) in the monitoring of tsetse control operations is discussed. PMID- 11114389 TI - Two different vibratory signals in Rhodnius prolixus (Hemiptera: reduviidae). AB - In this study the substrate-borne stridulatory vibrations produced by Rhodnius prolixus females were recorded and analysed in two different behavioural contexts. In the context of sexual communication females spontaneously stridulated to reject copulatory attempts performed by males. These male deterring stridulations were fully effective: out of 61 attempts, no copulation occurred. These stridulations consisted of short series of repetitive syllables, each one composed by a single chirp. In the context of defensive behaviour, bugs stridulated if they were clasped or restrained. These disturbance stridulations consisted of long series of repetitive syllables, each one composed by a series of short chirps and a long one. Male-deterring and disturbance stridulations differed in their temporal pattern and frequency spectra, having a main carrier frequency of about 1500 and 2200 Hz, respectively. As no differences in the inter ridge distances along the whole stridulatory organ were found, the differences in the frequency between both signals could be explained on the basis of a different velocity of rubbing of the proboscis against the prosternal stridulatory organ. It was found that R. prolixus and the related species Triatoma infestans rubbed only the central region of the stridulatory groove (around 1/3 of the total length) to produce disturbance stridulations. The results are discussed in relation to previous work on vibrational sensitivity in R. prolixus and are also compared with results reported for T. infestans. PMID- 11114390 TI - Development and persistence of antibodies to Orientia tsutsugamushi in the roof rat, Rattus rattus and laboratory mice following attachment of naturally infected Leptotrombidium deliense. AB - The development and persistence of antibodies to Orientia tsutsugamushi in Rattus rattus and laboratory mice following infection from the bite of naturally infected Leptotrombidium deliense is reported. Antibodies in R. rattus were first detected using an indirect immunoperoxidase method 2 weeks after attachment of an infected mite. The IgM antibody response was mild, and was detected 2, 4 and 6 weeks after infection, while the IgG response was stronger and was detected from 2 to 19 weeks after infection, when the tests ceased. Active rickettsiae were isolated from R. rattus 1-8 weeks after attachment of infected L. deliense, but rats were not adversely affected by infection and appeared to behave in the same way as uninfected rats. In contrast to rats, laboratory mice were killed by O. tsutsugamushi following infection by attachment of infected L. deliense. The development of antibodies in mice was not detected before gross symptoms of infection were observed. Antibodies were first detected 14 days after attachment of infected mites, when mice were close to death. PMID- 11114391 TI - The burden of hydrocele on men in Northern Ghana. AB - The social and economic impact of lymphatic filariasis was studied in Northern Ghana. Qualitative methods of gathering information revealed that even though the disease was a problem to both men and women, men with hydrocele suffered a greater psychosocial burden. Particular attention was paid to them, distinguishing men with small hydroceles and men with large ones. Out of frustration men with small hydroceles sought health care from a wider range of places than men with larger ones. The pain associated with adenolymphangitis (ADL) renders them inactive for up to 5 days. Complications of lymph scrotum and ridicule from community members were a problem. Unmarried men in particular found it difficult to find a spouse with their condition, and various degrees of sexual dysfunction were reported amongst married men. The clinical significance and the value of time and attention for counseling to mitigate the effects of the disease on damaged male identity and the need for gender studies to address male issues and the need for including psychosocial issues in the calculating of disability adjusted life years (DALY's) is also discussed. PMID- 11114392 TI - Behaviour of Glossina morsitans morsitans Westwood (Diptera: Glossinidae) on waterbuck Kobus defassa Ruppel and feeding membranes smeared with waterbuck sebum indicates the presence of allomones. AB - The behavioural responses of caged individual teneral Glossina morsitans morsitans on waterbuck and ox and on feeding membranes with and without smears of different doses of waterbuck sebum were compared. No significant difference was found in the initial landing behaviour on the two animals, nor on treated and control parts of the membrane. However, the subsequent behaviours of the flies were significantly different. Whereas none of the flies that landed on the ox showed any escape behaviour, more than a third of those that initially landed on waterbuck departed before probing. Similar results were obtained on feeding membranes treated in part with 1.0 or 1.4 mg cm(-2) of waterbuck sebum. Moreover, flies that landed on waterbuck or its sebum changed probing sites more often and probed significantly longer. The proportions that initiated feeding during the 10 min observation period were also significantly less. Our results suggest the presence of both volatile and non-volatile allomones on waterbuck which would account for low numbers of flies found attracted to and feeding on waterbuck in the wild. PMID- 11114393 TI - In vitro leishmanicidal activity of monomeric and dimeric naphthoquinones. AB - A series of monomeric and dimeric naphthoquinones with potential for treatment of Leishmania infections was identified in vitro using both a direct cytotoxicity assay against extracellular promastigotes of Leishmania donovani, L. infantum, L. enriettii, and L. major and a test against intracellular amastigote L. donovani residing within murine macrophages. Several naphthoquinones proved to be active at concentrations in the microgram range (EC(50) 0.9-17.0 microg/ml). When tested against a panel of human cancer cell lines (KB, SKMel, A549, MDA) and murine bone marrow culture-derived macrophages (BMMPhi) as mammalian host cell controls, compounds with anti-Leishmania-activity showed moderate (EC(50)>25 microg/ml) to pronounced (EC(50)<10 microg/ml) toxic effects. PMID- 11114394 TI - Pharmacological properties of native metabotropic glutamate receptors in freshly dissociated Golgi cells of the rat cerebellum. AB - We have examined the pharmacological properties of native metabotropic glutamate (mGlu) receptors in freshly isolated rat cerebellar Golgi cells using the whole cell configuration of the patch-clamp technique. Group II mGlu receptor agonists inhibited voltage-gated Ca(2+) channels (VGCC) currents in a reversible and concentration-dependent manner with a rank order of potency being LY354740> DCG IV > L-CCG-I > glutamate >>1S,3R-ACPD > NAAG. The maximum degree of inhibition obtained was similar for all drugs tested, saturating at about 33-41%, except for NAAG that had a non saturating effect of 50% at 1mM. Two novel group II mGlu receptor antagonists, LY341495 and Ro 65-3479, reversed VGCC current inhibition by LY354740 with pK(B) values of 7.0 and 6.3, respectively. In a subpopulation of Golgi cells, the antagonistic effect of LY341495 was only partial, suggesting a remaining effect of group I mGlu receptors. This was confirmed by experiments with S-DHPG, a selective group I mGlu receptor agonist. These experiments suggest that Golgi cells of the cerebellum express group II mGlu receptors that couple to the inhibition of VGCCs. Therefore, inhibition of VGCCs in cerebellar Golgi cells is a useful model system to evaluate novel group II mGlu receptor ligands. PMID- 11114395 TI - Pharmacological profiles of the metabotropic glutamate receptor ligands. AB - Metabotropic glutamate receptors (mGluRs) are a family of G-protein coupled receptors that are expressed in the central and peripheral nervous systems. The purpose of this study was to compare the ligand binding selectivity profiles of the mGluR agonist [(3)H]L-AP4 and the novel radiolabeled phenylglycine antagonist [(3)H]CPPG at all eight rat mGluR subtypes expressed in transfected human embryonic kidney cells. At a concentration of 30 nM [(3)H]L-AP4, no specific binding was detected in membranes expressing the group I receptors mGluR1a or mGluR5a, or in membranes expressing the group II mGluRs, mGluR2 and mGluR3. Among the group III mGluRs, specific [(3)H]L-AP4 binding was detected in cells expressing mGluR4a and mGluR8a but not in cells expressing mGluR6 or mGluR7a. The binding of [(3)H]CPPG showed an exceptional pattern of selectivity amongst the mGluR subtypes; at a concentration of 20 nM [(3)H]CPPG, a high level of specific binding was seen in membranes containing mGluR8a but not in any of the other mGluR subtypes. The affinity constant (K(D)) calculated for [(3)H]CPPG binding to mGluR8a was 183 nM. In competition experiments, the phosphono-substituted phenylglycine congeners including MPPG, (RS)-PPG, and unlabeled CPPG were the most potent inhibitors of [(3)H]CPPG binding while non-phosphonated compounds such as L-glutamate and MCPG were substantially less potent. These results demonstrate that [(3)H]L-AP4 and [(3)H]CPPG can be used as probes to selectively label group III mGluRs and that CPPG and related phenylglycine derivatives are useful for studying differences in the ligand recognition sites of highly homologous mGluRs. PMID- 11114396 TI - Effects of membrane cholesterol on the sensitivity of the GABA(A) receptor to GABA in acutely dissociated rat hippocampal neurones. AB - The effects of membrane cholesterol on the GABA(A) receptor were investigated in acutely dissociated rat hippocampal neurones, using the whole-cell patch clamp technique. Neuronal cholesterol was manipulated within the range 56-250% control by incubation with methyl-beta-cyclodextrin for depletion and a complex of cholesterol and methyl-beta-cyclodextrin for enrichment. Manipulation over a narrower range was achieved with cholesterol + phosphatidylcholine liposomes. A complex of epicholesterol and methyl-beta-cyclodextrin was used to insert epicholesterol. Cholesterol enrichment reduced the potency of GABA, as did cholesterol depletion, with increases in EC(50) of up to 4-fold. Cholesterol enrichment reduced the potency of the competitive antagonist bicuculline but did not affect that of the non-competitive antagonist picrotoxinin. Cholesterol depletion did not affect the potencies of either antagonist. Epicholesterol substituted functionally for cholesterol with respect to the effects of enrichment. In cholesterol-depleted neurones, however, only incubation with cholesterol was able to restore GABA potency to normal. These results suggest a specific requirement for cholesterol at control levels to maintain optimal GABA potency, which may involve specific binding of cholesterol to the GABA(A) receptor. The reduction in GABA potency by enrichment with cholesterol or epicholesterol is more likely to be due to reduced plasma membrane fluidity. PMID- 11114397 TI - Ontogenic expression of anterior pituitary GABA(B) receptor subunits. AB - gamma-Aminobutyric acid (GABA) is involved in the neuroendocrine control of hypophyseal secretion, acting both in the central nervous system and directly at the pituitary. We have characterized the properties of anterior pituitary GABA(B) receptors. In this work the ontogeny of rat anterior pituitary GABA(B) receptors and the pattern of subunit expression in rats of both sexes were determined. Western blot analysis showed a temporal decrease in GABA(B) subunits GABA(B(1a)) and GABA(B(1b)) expression in female anterior pituitary membranes from day 4 to adulthood, with GABA(B(1a)) being significantly more abundant than GABA(B(1b)) at early stages of development; the GABA(B(2)) subunit was barely detectable. In the male, GABA(B(1a)) followed a similar pattern and appeared to be significantly less abundant than in 4- and 12-day-old females; GABA(B(1b)) and GABA(B(2)) expression in the male was barely detectable. Scatchard plot analysis showed a temporal decrease in binding sites in female anterior pituitary membranes, in agreement with the western blot results. The number of binding sites was significantly higher in female than in male 4-day-old membranes. Dissociation constant values were similar for both sexes at all ages studied. This study reports for the first time the ontogeny of anterior pituitary GABA(B) receptors, showing a particular developmental pattern of subunit expression and a clear sexual dimorphism. PMID- 11114398 TI - The effects of GABA(B) receptor activation on spontaneous and evoked activity in the dentate gyrus of kainic acid-treated rats. AB - Enhancement of GABAergic inhibition is central to the treatment of epilepsy. The role of the GABA(B) receptor, however, is poorly understood. The current study investigates the effects of r-baclofen (a GABA(B) receptor agonist) on spontaneous and evoked electrophysiological activity in the dentate gyrus of normal and epileptic rats in vivo. Administration of kainic acid (KA), which causes similar pathology to that seen in human temporal lobe epilepsy, was used to prepare chronically epileptic rats. Bursts of spontaneous high-amplitude field potentials (spiking) were observed in isoflurane-anaesthetised control and KA treated rats in vivo; however, this activity was significantly more frequent in KA-treated rats (223+/-26.1 spikes min(-1)) than in control rats (124+/-17.4 spikes min(-1)). Feedback inhibition, measured using paired-pulsed stimulation, was also greater in KA-treated rats; 50% inhibition of the second response was observed at 43.05+/-4.46 ms in KA-treated animals, as opposed to 26.27+/-2.39 ms in control animals. r-Baclofen (10 mg kg(-1) i.v.) abolished spontaneous spiking and also reduced feedback inhibition in both control and KA-treated rats. These effects of r-baclofen may be due to inhibition of GABA release, through activation of pre-synaptic GABA(B) receptors on terminals of interneurones in the inhibitory feedback pathway. These results suggest a link between feedback inhibition and spontaneous spiking, and provide support for the hypothesis that mechanisms of synchronisation may give rise to seizure activity in human temporal lobe epilepsy. PMID- 11114399 TI - Epileptiform activity and EPSP-spike potentiation induced in rat hippocampal CA1 slices by repeated high-K(+): involvement of ionotropic glutamate receptors and Ca(2+)/calmodulin-dependent protein kinase II. AB - We have previously demonstrated that repeated brief increases in extracellular K(+) (K(+)(o)) induce a hyperexcitability in CA1 pyramidal cells that persists for a long time after the final application of K(+) [Neurosci. Lett. 223 (1997) 177; Epilepsy Research (2000) 75]. This epileptiform activity, which was associated with a lasting excitatory postsynaptic potential (EPSP)-spike potentiation, presented some of the characteristic features of traditional in vivo kindling. We have also found that Ca(2+) influx through L-type voltage sensitive Ca(2+) channels is essential for the development of both in vitro kindling and EPSP-spike potentiation. The aims of this study were to investigate the involvement of ionotropic glutamate receptors, especially those of the NMDA subtype, and the requirement for Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) in these phenomena. Field EPSPs with presynaptic fibre volleys from the stratum radiatum, and population spikes from the stratum pyramidale, were recorded in the CA1 area of rat hippocampal slices in response to electrical stimulation of the Schaffer collateral/commissural fibres. Repeated (three episodes) brief (30 s) increases in extracellular K(+) induced a sustained decrease in the threshold for development of evoked epileptiform discharges (i.e. an in vitro kindling-like state) and a lasting potentiation of the EPSP-spike transfer in CA1 pyramidal neurons (EPSP-spike potentiation). The selective antagonist of NMDA receptors, APV (50 microM), blocked the EPSP-spike potentiation, depressed the induction phase of the in vitro kindling-like state, and blocked the maintenance phase of this state. In contrast to APV, the blockade of AMPA/kainate receptors by CNQX (10 microM) had no effect. Like APV, KN62 (3 microM), a selective membrane permeable inhibitor of CaMKII, blocked the EPSP spike potentiation and the maintenance phase of the in vitro kindling-like state. Our previous and present results therefore demonstrate that Ca(2+) influxes through L-type voltage-dependent-and NMDA receptor-dependent-Ca(2+) channels contribute differentially to the development of an in vitro kindling-like state, and both induce EPSP-spike potentiation in CA1 hippocampal pyramidal cells in response to repeated brief increases in K(+)(o). It is suggested that these effects of intracellular Ca(2+) on the maintenance phase of the in vitro kindling like state and EPSP-spike potentiation are mediated by CaMKII-dependent mechanisms. PMID- 11114400 TI - Supraspinal vs spinal sites of the antinociceptive action of the subtype selective NMDA antagonist ifenprodil. AB - The N-methyl-D-aspartate (NMDA) antagonist ifenprodil and several structurally related compounds are highly selective for the NR2B-containing receptor subtype. This selectivity could provide an explanation for the reported difference of the analgesic and side-effect profile of ifenprodil-like compounds from other NMDA antagonists. In this work, we have queried if the ifenprodil-induced antinociception can be attributed to the block of NMDA receptors in the spinal cord. Ifenprodil and some other NMDA antagonists (MK-801, memantine) were tested in a model of inflammatory pain (Randall-Selitto) in rats. The in vivo NMDA antagonism was assessed in anaesthetised rats on responses of spinal dorsal horn (DH) neurones to iontophoretic NMDA and in the model of single motor unit (SMU) wind-up. Ifenprodil, MK-801 and memantine dose-dependently increased nociceptive thresholds in the Randall-Selitto model. Antinociceptive doses of the channel blockers selectively antagonised NMDA responses of DH neurones and inhibited wind up. In contrast, antinociceptive doses of ifenprodil did not show any NMDA antagonism in electrophysiological tests. Although ifenprodil did not inhibit the SMU responses to noxious stimuli in spinalised rats, it markedly and dose dependently inhibited nociceptive SMU responses in sham-spinalised rats. These results argue against the spinal cord being the principal site of antinociceptive action of ifenprodil; supraspinal structures seem to be involved in this effect. PMID- 11114401 TI - Actions of cannabinoid receptor ligands on rat cultured sensory neurones: implications for antinociception. AB - Cannabinoids modulate nociceptive processing in models of acute, inflammatory and neuropathic pain. We have investigated the location and function of cannabinoid receptors on cultured neonatal dorsal root ganglion (DRG) neurones and F-11 cells, a dorsal root ganglionxneuroblastoma hybridoma which displays several of the features of authentic DRG neurones. CB(1) receptor immunolabelling was observed on the cell bodies and as fine puncta on processes of both cultured DRG neurones and F-11 cells. Additionally, fluorescence-activated cell sorting (FACS) analysis provided evidence that both CB(1) and CB(2) receptors are expressed on populations of cells within the cultured DRG and F-11 cells. The cannabinoid receptor agonist (+)-WIN55212 (10 and 100 nM) inhibited the mean voltage activated Ca(2+) current in DRG neurones by 21% and 30%, respectively. The isomer, (-)-WIN55212 (10 and 100 nM) produced significantly less inhibition of 6% and 10% respectively. The CB(1) selective receptor antagonist SR141716A (100 nM) enhanced the peak high voltage-activated Ca(2+) current by 24% and simultaneous application of SR141716A (100 nM) and (+)-WIN55212 (100 nM) resulted in a significant attenuation of the inhibition obtained with (+)-WIN55212 alone. These data give functional evidence for the hypothesis that the analgesic actions of cannabinoids may be mediated by presynaptic inhibition of transmitter release in sensory neurones. PMID- 11114402 TI - New channel blocker BIIA388CL blocks delayed rectifier, but not A-type potassium current in central neurons. AB - A new substance (R,S)-(3,4-dihydro-6,7-dimethoxyisoquinoline-1-yl)-2-cyclohexyl-N (3,3-diphenylpropyl)-acetamide hydrochloride (BIIA388Cl), which demonstrates neuroprotective properties in animal models, was examined for its action on K(+) currents in acutely isolated rat hippocampal neurons using the patch clamp/concentration clamp techniques in the whole-cell configuration. The delayed rectifier K(+)-current (I(DR)) was strongly inhibited by externally applied BIIA388Cl, while the transient A-current (I(A)) remained virtually unaffected. Block of I(DR) by the pre-applied BIIA388Cl was revealed as a rapid decay of the current indicating direct interaction of the drug with the open state of the channel. The removal of the block upon repolarization was also rapid (tau=22 ms). The dose-response relationship for the blocking action of BIIA388Cl revealed an IC(50) value of 300 nM for the peak I(DR), whereas the IC(50) value for I(DR) measured 300 ms after the onset of depolarization was 120 nM. The blocking action of BIIA388Cl on I(A) was at least 200 times less potent. These data allow us to conclude that BIIA388Cl is an effective and selective blocker of I(DR). This current is the main pathway for the loss of intracellular potassium by depolarized neurons. Selective obstruction of this pathway could be useful for neuroprotection. PMID- 11114403 TI - Detailed distribution of Neurokinin 3 receptors in the rat, guinea pig and gerbil brain: a comparative autoradiographic study. AB - The neurokinin 3 (NK3) receptor is predominantly expressed in the central nervous system (CNS). Species differences in neurokinin 3 (NK3) receptor pharmacology have led to the preferential use of guinea pigs and gerbils in the characterization of non-peptide NK3 antagonists. Little is known about the central localization of NK3 receptors in the CNS of these species. To study this, [(3)H]senktide and [(3)H]SR 142801 were used in autoradiography experiments to visualize the NK3 receptors in the guinea pig and gerbil brain and compared to with the distribution of [(3)H]senktide binding sites in the rat brain. In the three species, the NK3 receptor was similarly distributed within the cerebral cortex, the zona incerta, the medial habenula, the amygdaloid complex, the superior colliculus and the interpeduncular nucleus. Outside of these structures, our study has revealed that each species displayed a specific distribution pattern of central NK3 receptors. The rat was the only species where NK3 receptors could be visualized in the striatum, the supraoptic nucleus and the paraventricular nucleus of the hypothalamus. The guinea pig differed mainly from the two other species by the absence of detectable binding sites in the substantia nigra pars compacta and the ventral tegmental area. A specific localization of NK3 receptors in the anterodorsal and anteroventral thalamic nuclei characterized the gerbil. This last species is also unique by in the higher level of NK3 receptors in the dorsal and median raphe nuclei. All these differences suggest that the NK3 receptor mediates different functions in different species. PMID- 11114404 TI - Pharmacological evidence for the role of central alpha 1B-adrenoceptors in the motor activity and spontaneous movement of mice. AB - Central alpha 1-noradrenergic neurotransmission has been shown to be an important complement of dopaminergic transmission in the control of motor activity but the identity of the responsible alpha 1 receptor subtype has not yet been identified. This was investigated in the present experiment by measuring the effects of intraventricular administration of a series of alpha 1 antagonists varying in affinities for the three known receptor subtypes--1a, 1b and 1d--on active behavior in mice in response to a cage change. It was found that the potency of the drugs to block both gross and small movements correlated highly with published affinities for the cloned 1b receptor but not for those of either the cloned 1a or 1d receptors. It is concluded that central alpha 1B receptors are critically involved in the mediation of the (nor)adrenergic influence on active behavior, a finding which has implications for basic and clinical research in both movement and mood disorders. PMID- 11114405 TI - PDE4 inhibitors induce emesis in ferrets via a noradrenergic pathway. AB - The objective of this work was to assess the role of alpha(2)-adrenoceptors in emesis induced by inhibitors of type 4 phosphodiesterase (PDE4) in ferrets. Pre treatment with yohimbine, MK-912 or MK-467 (alpha(2)-adrenoceptor antagonists) caused sudden and unexpected vomiting. In contrast, clonidine (alpha(2) adrenoceptor agonist) did not induce emesis at doses ranging from 62.5-250 microg/kg s.c. At the dose of 250 microg/kg, clonidine also provided protection against emesis induced by the PDE4 inhibitors, PMNPQ (i.e. 6-(4-pyridylmethyl)-8 (3-nitrophenyl)quinoline, CT-2450 and R-rolipram. It was postulated that PDE4 inhibitors trigger emesis by mimicking the pharmacological actions of alpha(2) adrenoceptor antagonists. This hypothesis was strengthened by the demonstration that PDE4 inhibitors can reverse the hypnotic effect of an alpha(2)-adrenoceptor mediated anaesthetic regimen in rats and ferrets. Similar to alpha(2) adrenoceptor antagonists, PMNPQ, R-rolipram and S-rolipram dose-dependently decreased the duration of anaesthesia in rats injected with the combination xylazine/ketamine. While subcutaneous injections of CT-2450 (3-30 mg/kg) were without effect, a central infusion (6 microg i.c.v.) decreased the duration of anaesthesia. These studies suggest that the ferret is an appropriate model to study emesis induced by PDE4 inhibitors and that these compounds trigger the emetic reflex via a noradrenergic pathway, mimicking the pharmacological actions of a pre-synaptic alpha(2)-adrenoceptor inhibition. PMID- 11114406 TI - Influence of nitric oxide donors and of the alpha(2)-agonist UK-14,304 on acetylcholine release in the pig gastric fundus. AB - This study in circular muscle strips of the pig gastric fundus aimed to measure the release of acetylcholine directly and to investigate whether NO and alpha(2) adrenoceptor agonists can modulate acetylcholine release from cholinergic neurones. After incubation of the tissues with [(3)H]-choline, basal and electrically induced release of tritium and [(3)H]-acetylcholine were analyzed in a medium containing physostigmine (10(-5) M) as well as atropine (10(-6) M). The NO synthase inhibitor L-N(G)-nitroarginine methyl ester (3x10(-4) M), and the NO donors sodium nitroprusside (10(-5) M) and 3-morpholino-sydnonimine (10(-5) M) did not influence the basal release nor the electrically evoked release, indicating that NO does not modify [(3)H]-acetylcholine release. The alpha(2) adrenoceptor agonist UK-14,304 (10(-5) M) significantly inhibited the electrically evoked release of [(3)H]-acetylcholine, and this effect was prevented by the alpha(2)-adrenoceptor antagonist rauwolscine (2x10(-6) M), suggesting that presynaptic alpha(2)-adrenoceptors are present on cholinergic neurones of the pig gastric fundus. PMID- 11114407 TI - SCH 23390 affords protection against soman-evoked seizures in the freely moving guinea-pig: a concomitant neurochemical, electrophysiological and behavioural study. AB - We studied the role of striatal dopamine (DA) release in seizure activity evoked by the subcutaneous administration of the cholinesterase inhibitor pinacolyl methylphosphonofluoridate (soman), in the guinea-pig. The involvement of the dopamine receptor subtypes was studied by systemic administration of the D(1) like receptor antagonist SCH 23390 (0.5 mg kg(-1)) or the D(2)-like receptor antagonist sulpiride (30 mg kg(-1)). Microdialysis and HPLC with electrochemical detection were used to monitor changes in extracellular levels of striatal DA and its metabolites, acetylcholine and choline. These data were correlated with changes in the striatal and cortical electroencephalogram and observation of predefined clinical signs. We found that the blockade of the D(1) receptor with SCH 23390 can inhibit seizure activity, while blockade of the D(2) receptor with sulpiride can augment the evoked seizure activity. These results clarify the involvement of the dopaminergic system in soman-evoked seizures. PMID- 11114408 TI - Combinations of clozapine and phencyclidine: effects on drug discrimination and behavioral inhibition in rats. AB - Phencyclidine (PCP) produces psychotomimetic effects in humans that resemble schizophrenia symptoms. In an effort to screen compounds for antipsychotic activity, preclinical researchers have investigated whether these compounds block PCP-induced behaviors in animals. In the present study, the atypical antipsychotic clozapine was tested in combination with an active dose of PCP in two-lever drug discrimination and mixed signalled-unsignalled differential reinforcement-of-low-rates (DRL) procedures. PCP produced distinctive effects in each task: it substituted for the training dose in PCP discrimination and it increased the number of responses with short (<3 s) interresponse times as well as increasing overall response rates in the DRL schedule. Acute dosing with clozapine failed to alter the behavioral effects of PCP in either procedure even when tested up to doses that produced pharmacological effects alone. These results suggest that acute dosing with clozapine would not affect behaviors most closely associated with PCP intoxication. Further, they bring into question the utility of using PCP combination procedures in animals to screen for antipsychotic potential. Since chronic dosing is required for therapeutic efficacy of antipsychotics, future studies should focus on investigation of chronic dosing effects of these drugs in combination with PCP. PMID- 11114409 TI - Blockade of sensitization to methylphenidate by MK-801: partial dissociation from motor effects. AB - The role of MK-801's locomotor effect in blocking the development of sensitization to methylphenidate was investigated utilizing a computerized locomotor activity monitoring system. After 7 days of acclimation to a 12:12 light-dark cycle (lights on at 07:00), male Sprague-Dawley rats (n=62) were housed in test cages and motor activity was recorded continuously for 16 days. The first 2 days of recording served as a baseline for each rat, and on day 3 each rat received a saline injection. On days 4 to 9 rats were randomly divided into seven groups: Rats received either six daily s.c. injections of methylphenidate (2.5 mg/kg; Group 1), or six daily i.p. injections of 0.30 mg/kg, 0.05 mg/kg MK-801 (Groups 2 and 3, respectively); two MK-801 pre-treatment groups received a single i.p. injection of 0.05, or 0.30 mg/kg MK-801 one hour prior to 2.5 mg/kg methylphenidate (n=8 each) on day 4 followed by five daily injections of 2.5 mg/kg methylphenidate; and finally, two cotreatment groups received a challenge dose of 2.5 mg/kg methylphenidate on day 4 followed by either 0.05 or 0.30 mg/kg MK-801 i.p. one hour prior to 2.5 mg/kg methylphenidate from days 5 to 9. All groups were allowed five days of no treatment before being re-challenged on day 15 with the same treatment they received on day 4. Methylphenidate and 0.30 mg/kg MK-801 sensitized to their own locomotor effects, but 0.05 mg/kg MK 801, which had no acute motor effects, did not. The administration of MK-801 (0.30 mg/kg) prior to methylphenidate either singly on day 4, or coadministered throughout the repeated methylphenidate treatment phase, blocked the development of sensitization to methylphenidate. However, MK-801 at 0.05 mg/kg delayed the development of sensitization when co-administered on days 5 to 9, but a single injection 1 h prior to methylphenidate on day 4 did not prevent sensitization to subsequent methylphenidate administration. In conclusion, MK-801 prevents sensitization to methylphenidate; motor stimulation by MK-801 is not necessary for short-term prevention or delay of sensitization to methylphenidate but may be necessary for a persistent blockade of sensitization. PMID- 11114410 TI - Innate antiviral defenses in body fluids and tissues. AB - Innate, non-specific, resistance mechanisms are important barriers to pathogens, particularly delaying virus multiplication at the onset of infections. These innate defense mechanisms include a series of mechanical barriers, pre-existing inhibitory molecules, and cellular responses with antimicrobial activity. The antiviral activities of these innate inhibitors reside in a variety of partly characterized substances. This review presents the innate antiviral inhibitors in cell cultures, urine, serum, the gastrointestinal tract, the nervous system, tissues of crustaceans, and saliva. Medical adaptation of the innate antiviral defense mechanisms may be useful for prevention and treatment of viral infections. PMID- 11114411 TI - Alpha interferon combined with ribavirin potentiates proliferative suppression but not cytokine production in mitogenically stimulated human lymphocytes. AB - The improved clinical outcome observed among patients with hepatitis C treated with the combination of alpha interferon (IFN) and ribavirin (RBV) is presumed to result from immunomodulation and viral inhibition. However, the impact of the drug combination upon lymphocyte activity is unknown. The present study evaluated the effects of IFN and RBV, singly and in combination, upon proliferation, cell cycle sensitivity and cytokine elaboration following PHA stimulation of lymphocytes. Two formulations of IFN, interferon-a-2b (IFN-2b) and interferon-a con-1 (CIFN), were included. Titration of each drug over a wide range of concentrations showed dose dependent proliferative suppression without cytotoxicity. Proliferation was suppressed 57-99% (P<0.001) by IFN-2b (10(5) 10(7) IU/ml), 41-74% (P<0.001) by CIFN (1.5-150 ng/ml), and 10-94% (P<0.001) by RBV (0.5-50 microg/ml). Isobologram analysis showed that the interaction between IFN-2b and RBV on proliferative suppression was additive. In contrast, the interaction between CIFN and RBV was weakly antagonistic. Proliferative suppression by both the IFNs was cell cycle restricted. IFN-2b and CIFN added at the onset of PHA stimulation (G0/G1) versus 24 h later (S phase) inhibited proliferation by 50 versus 5%, respectively (P<0.05). The onset of IFN resistance correlated with a 50% reduction (P<0.05) in IFN receptors on the cell surface. In contrast, RBV caused equivalent proliferative suppression (P=NS) when added at any time during PHA activation. Cytokine secretion after 24 h of PHA stimulation showed that IFN-2b versus CIFN increased the secretion of IL2, TNF and gamma IFN by 4.5-, 4.1- and 8.3-fold (P<0.005) versus 1-, 1.9- and 1.9-fold (P<0.05), respectively, above control levels. Neither IFN affected IL10 secretion. RBV, singly and in combination with IFN, had no impact on cytokine expression (P=NS). This study identifies several potential mechanisms by which the combination of IFN and RBV may exert a more potent effect upon cellular immunity than either agent alone and shows that different formulations of IFN may have non-identical effects upon lymphocyte responses. PMID- 11114413 TI - Ribavirin and mycophenolic acid potentiate the activity of guanine- and diaminopurine-based nucleoside analogues against hepatitis B virus. AB - Mycophenolic acid [the active metabolite of the immunosuppressive agent mycophenolate mofetil (MMF)] and ribavirin were found to potentiate the anti-HBV activity of the guanine-based nucleoside analogues penciclovir (PCV), lobucavir (LBV) and 3'-fluorodideoxyguanosine (FLG) and diaminopurine dioxolane (DAPD). Ribavirin and mycophenolic acid are both inhibitors of inosine 5'-monophosphate dehydrogenase and cause a depletion of intracellular dGTP levels. It may be assumed that the 5'-triphosphorylated derivatives of the guanine-based nucleoside analogues, in the presence of reduced levels of dGTP, inhibit more efficiently the priming reaction as well as the reverse transcription and DNA-dependent DNA polymerase activity of the HBV polymerase. This assumption is corroborated by the observation that exogenously added guanosine reversed the potentiating effect of ribavirin and mycophenolic acid on the anti-HBV activity of the guanosine analogues. Our observations may have implications for those (liver) transplant recipients that receive MMF as (part of their) immunosuppressive regimen and that, because of de novo or persistent infection with HBV, need specific anti-HBV therapy. PMID- 11114412 TI - The neuraminidase inhibitor GS4104 (oseltamivir phosphate) is efficacious against A/Hong Kong/156/97 (H5N1) and A/Hong Kong/1074/99 (H9N2) influenza viruses. AB - In 1997, an H5N1 avian influenza A/Hong Kong/156/97 virus transmitted directly to humans and killed six of the 18 people infected. In 1999, another avian A/Hong/1074/99 (H9N2) virus caused influenza in two children. In such cases in which vaccines are unavailable, antiviral drugs are crucial for prophylaxis and therapy. Here we demonstrate the efficacy of the neuraminidase inhibitor GS4104 (oseltamivir phosphate) against these H5N1 and H9N2 viruses. GS4071 (the active metabolite of oseltamivir) inhibited viral replication in MDCK cells (EC(50) values, 7.5-12 microM) and neuraminidase activity (IC(50) values, 7.0-15 nM). When orally administered at doses of 1 and 10 mg/kg per day, GS4104 prevented death of mice infected with A/Hong Kong/156/97 (H5N1), mouse-adapted A/Quail/Hong Kong/G1/97 (H9N2), or human A/Hong Kong/1074/99 (H9N2) viruses and reduced virus titers in the lungs and prevented the spread of virus to the brain of mice infected with A/Hong Kong/156/97 (H5N1) and mouse-adapted A/Quail/Hong Kong/G1/97 (H9N2) viruses. When therapy was delayed until 36 h after exposure to the H5N1 virus, GS4104 was still effective and significantly increased the number of survivors as compared with control. Oral administration of GS4104 (0.1 mg/kg per day) in combination with rimantadine (1 mg/kg per day) reduced the number of deaths of mice infected with 100 MLD(50) of H9N2 virus and prevented the deaths of mice infected with 5 MLD(50) of virus. Thus, GS4104 is efficacious in treating infections caused by H5N1 and H9N2 influenza viruses in mice. PMID- 11114414 TI - In vivo effect of EICAR (5-ethynyl-1-beta-D-ribofuranosylimidazole-carboxamide) on experimental infected rainbow trout (Oncorhynchus mykiss) and coho salmon (Oncorhynchus kisutch) fry with infectious pancreatic necrosis virus. AB - The in vivo antiviral effect of 5-ethynyl-1-beta-D-ribofuranosylimidazole carboxamide (EICAR) was evaluated in coho salmon and rainbow trout fry, experimentally infected with infectious pancreatic necrosis virus (IPNV). Treatment consisted of a daily bath of 2 h in 0.4 microg ml(-1) or 0.8 microg ml( 1) of EICAR, for approximately 20 days. The behavior of the fish was studied for 45 days post-infection. The survival of the infected treated groups was compared with the survival of non-infected and infected untreated control groups. The results showed that the survival of coho salmon and rainbow trout fry in the infected group treated with both doses of EICAR was similar to the survival observed in the healthy control group (approximately 94%). While, the survival of the infected and untreated control fish was 56% for salmon and 28% for trout, there were no significant difference in the weight of coho salmon fry between those treated with EICAR and non-infected and infected untreated control groups. However, in rainbow trout there was a statistically significant weight decrease in infected untreated group. Finally, the analysis of tissue samples of the fish by reverse transcription associated with the polymerase chain reaction (RT-PCR) suggest that EICAR have decreased the viral load in infected treated fry. Consequently, the results indicate that EICAR is an effective inhibitor of IPNV replication in vivo and could be a promissory antiviral compound for the treatment of IPNV disease. PMID- 11114416 TI - Putting the rap on integrin activation. PMID- 11114415 TI - In vivo anti-papillomavirus activity of nucleoside analogues including cidofovir on CRPV-induced rabbit papillomas. AB - A series of nucleoside analogues were tested for in vivo anti-papillomavirus activity using the cottontail rabbit papillomavirus (CRPV) domestic rabbit model. Compounds were delivered either topically, injected into growing papillomas, or delivered subcutaneously at a site remote from the papillomas. Compounds tested included cidofovir [(S)-1-(3-hydroxy-2-phosphonylmethoxypropyl)cytosine] (HPMPC); cyclic HPMPC (cHPMPC); cyclopentenylcytosine (CPE-C); lobucavir [1R(1alpha,2beta,3alpha)]-9-[2, 3-bis(hydroxymethyl)cyclobutyl]guanine; 9-((2 phosphonylmethoxy)propyl)adenine (PMPA); adefovir 9-((2 phosphonylmethoxy)ethyl)adenine(PMEA) and cyclopropyl 9-(2 phosphonylmethoxyethyl)-2,6-diaminopurine (cyclopropylPMEDAP). Dose response curves and time-course treatments were included for most compounds tested. Strong anti-viral activity was detected using cidofovir and cHPMPC when delivered either topically or by the intralesional route. Complete cures were obtained using 1% (w/v) topical cidofovir at dosing schedules of twice daily for 8 weeks beginning at 4 weeks after CRPV infection, which represents a time when papillomas were clearly visible. Complete cures of large established papillomas were obtained by intralesional injection of 1% cidofovir three times per week for 8 weeks. Topical treatments with adefovir had strong anti-viral activity, cyclopropyl PMEDAP had moderate anti-viral activity, and CPE-C, PMPA and lobucavir showed no effects. These data indicate that certain nucleoside analogues have strong in vivo anti papillomavirus activity and that the CRPV/rabbit model is a good model for assessing clinical responses of anti-viral treatments for patients with HPV disease. PMID- 11114418 TI - How does TAP pump peptides? insights from DNA repair and traffic ATPases. PMID- 11114419 TI - Stress-free T-cell development: glucocorticoids are not obligatory. AB - A role for glucocorticoids in thymopoiesis has been suggested by studies using glucocorticoid receptor (GR) anti-sense transgenic mice, glucocorticoid synthesis inhibitors and GR antagonists. Unfortunately, no consensus has been reached on exactly how glucocorticoids influence T-cell development. The most recent approach, using GR knockout (GR(-/-)) mice, indicates that GR signaling is, in fact, dispensable in this entire process. PMID- 11114420 TI - The ITAM-bearing transmembrane adaptor DAP12 in lymphoid and myeloid cell function. AB - DAP12, an ITAM-bearing transmembrane adaptor protein, associates non-covalently with receptors in natural killer (NK) and myeloid cells, and provides signaling function via the Syk and ZAP-70 tyrosine kinase activation pathways. Humans and mice lacking DAP12 (DAP12(-/-)) show normal development of hematopoietic cells. However, DAP12(-/-) humans develop presenile dementia and bone cysts, and DAP12( /-) mice show impaired immune responses. PMID- 11114421 TI - Helicobacter pylori vaccine strategies--triggering a gut reaction. AB - Helicobacter pylori causes peptic ulcer disease and some forms of gastric cancer; it is one of the most common chronic bacterial infections of humans. Although several prototype protein-based vaccines have shown promising results, they have not cleared infection and/or prevented reinfection. Nucleic acid vaccination offers a useful alternative to protein immunization, especially now that two complete H. pylori genome sequences are available, and facilitates the selection of antigenic targets. PMID- 11114422 TI - A natural history of melanoma: serial gene expression analysis. AB - Ena Wang and Francesco Marincola propose a novel strategy whereby tumor-host interactions are studied within the melanoma microenvironment by serial gene expression analysis. Methodological constraints and ways to circumvent them are discussed. This approach might improve our understanding of the molecular basis of tumor regression in response to immune manipulation. PMID- 11114423 TI - Natural antibodies and complement link innate and acquired immunity. AB - Natural or spontaneous antibodies are an essential part of the first line of defense against hematogenically spreading infections, including viruses. These antibodies target virus-antibody complexes and complement to the spleen. This prevents infections from reaching vital organs and enhances neutralizing antibody responses, particularly when the antibody is bound to a highly repetitive antigen. PMID- 11114424 TI - Molecular interactions of coreceptor CD8 and MHC class I: the molecular basis for functional coordination with the T-cell receptor. AB - In recent years, substantial progress has been made towards understanding the molecular basis for CD8 binding to class I MHC and the coreceptor's role in cytotoxic T-cell activation. Here, we review the structural, mechanistic and functional studies that point to a model of coordination of T-cell receptor and CD8 signaling that might provide the key to cytotoxic T-cell activation. PMID- 11114426 TI - Erratum to 'How to develop globular proteins into adhesives'. PMID- 11114427 TI - Gene network dynamics and the evolution of development. PMID- 11114425 TI - Branching out to gain control: how the pre-TCR is linked to multiple functions. AB - How is signaling specificity achieved by the pre-TCR during selection of T-cell fate? Like the TCR, this receptor controls many functions, and recent studies define which pathways couple the pre-TCR to the molecular events controlling survival, proliferation, allelic exclusion at the TCRbeta locus, and further differentiation. PMID- 11114428 TI - Speciation: Dobzhansky-Muller incompatibilities, dominance and gene interactions. PMID- 11114429 TI - The evolutionary stability of mixed strategies. PMID- 11114430 TI - Global warming and the spread of disease: the debate heats up. PMID- 11114431 TI - Conflict begets diversity? PMID- 11114432 TI - How does recombination affect phylogeny estimation? PMID- 11114433 TI - Finding Mr Right: good genes and multiple mating by females. PMID- 11114434 TI - Evidence that specialists are special. PMID- 11114435 TI - Paleoecology and coalescence: phylogeographic analysis of hypotheses from the fossil record. AB - The application of principles from coalescence theory to genealogical relationships within species can provide insights into the process of diversification and the influence of biogeography on distributional patterns. There are several features that make some organisms more suitable for detailed studies of historical processes; in particular, limited dispersal, which serves to conserve the patterns of genetic variation that developed during colonization. We describe the potential benefits of studies that integrate analyses of genetic variation with information from the fossil pollen record and present recent examples of the application of quantitative methods of phylogeographic analysis. PMID- 11114436 TI - Statistical methods for detecting molecular adaptation. AB - The past few years have seen the development of powerful statistical methods for detecting adaptive molecular evolution. These methods compare synonymous and nonsynonymous substitution rates in protein-coding genes, and regard a nonsynonymous rate elevated above the synonymous rate as evidence for darwinian selection. Numerous cases of molecular adaptation are being identified in various systems from viruses to humans. Although previous analyses averaging rates over sites and time have little power, recent methods designed to detect positive selection at individual sites and lineages have been successful. Here, we summarize recent statistical methods for detecting molecular adaptation, and discuss their limitations and possible improvements. PMID- 11114437 TI - Nice snake, shame about the legs. AB - Snakes are one of the most extraordinary groups of terrestrial vertebrates, with numerous specializations distinguishing them from other squamates (lizards and their allies). Their musculoskeletal system allows creeping, burrowing, swimming and even gliding, and their predatory habits are aided by chemo- and thermoreceptors, an extraordinary degree of cranial kinesis and, sometimes, powerful venoms. Recent discoveries of indisputable early fossil snakes with posterior legs are generating intense debate about the evolutionary origin of these reptiles. New cladistic analyses dispute the precise significance and phylogenetic placement of these fossils. These conflicting hypotheses imply radically different scenarios of snake origins and relationships with wide biological implications. PMID- 11114438 TI - Alternative reproductive strategies: a queen perspective in ants. AB - Ant colonies are commonly thought to have a stable and simple family structure, with one or a few egg-laying queens and their worker daughters. However, recent genetic studies reveal that the identity of breeding queens can vary over time within colonies. In several species, some queens are apparently specialized to enter established colonies instead of initiating a new colony on their own. The previously overlooked occurrence of queen turnover within colonies has important consequences not only on the genetic structure and nature of kin conflict within colonies, but also on the evolution of social parasitism. PMID- 11114439 TI - Predicting extinction risks for plants: environmental stochasticity can save declining populations. AB - An emerging generalization from theoretical and empirical studies on conservation biology is that high levels of environmental stochasticity increase the likelihood of population extinction. However, coexistence theory has illustrated that there are circumstances under which environmental stochasticity can increase the chance of population persistence. These theoretical studies have shown that the sign of the effect of environmental stochasticity on population persistence is determined by interactions between life history and environmental stochasticity. These interactions mean that the stochastic and deterministic rates of population growth might differ fundamentally. Although difficult to demonstrate in real systems, observed life histories and variance in the vital rates of populations suggest that this phenomenon is likely to be common, and is therefore of much relevance to conservation biologists. PMID- 11114440 TI - Biomechanics of the cervical spine Part 2. Cervical spine soft tissue responses and biomechanical modeling. AB - OBJECTIVE: The responses and contributions of the soft tissue structures of the human neck are described with a focus on mathematical modeling. Spinal ligaments, intervertebral discs, zygapophysial joints, and uncovertebral joints of the cervical spine are included. Finite element modeling approaches have been emphasized. Representative data relevant to the development and execution of the model are discussed. A brief description is given on the functional mechanical role of the soft tissue components. Geometrical characteristics such as length and cross-sectional areas, and material properties such as force-displacement and stress-strain responses, are described for all components. Modeling approaches are discussed for each soft tissue structure. The final discussion emphasizes the normal and abnormal (e.g., degenerative joint disease, iatrogenic alteration, trauma) behaviors of the cervical spine with a focus on all these soft tissue responses. A brief description is provided on the modeling of the developmental biomechanics of the pediatric spine with a focus on soft tissues. Relevance. Experimentally validated models based on accurate geometry, material property, boundary, and loading conditions are useful to delineate the clinical biomechanics of the spine. Both external and internal responses of the various spinal components, a data set not obtainable directly from experiments, can be determined using computational models. Since soft tissues control the complex structural response, an accurate simulation of their anatomic, functional, and biomechanical characteristics is necessary to understand the behavior of the cervical spine under normal and abnormal conditions such as facetectomy, discectomy, laminectomy, and fusion. PMID- 11114441 TI - Intervertebral disc herniation: studies on a porcine model exposed to highly repetitive flexion/extension motion with compressive force. AB - OBJECTIVE: To determine whether repeated motion with low magnitude joint forces, and flexion/extension moments consistently produce herniation in a non degenerated, controlled porcine spine motion segment. DESIGN: Combined loading (flexion/extension motions and compressive forces) was applied to in vitro porcine functional spinal units. Biomechanical and radiographic characteristics were documented. BACKGROUND: While most studies performed in vitro have examined uniaxial or fixed position loading to older specimens, there have been few studies that have examined whether 'healthy' intervertebral discs can be injured by low magnitude repeated combined loading. METHODS: Porcine cervical spine motion segments (C3-C4) were mounted in a custom jig which applied axial compressive loads with pure flexion/extension moments. Dynamic testing was conducted to a maximum of 86400 bending cycles at a rate of 1 Hz with simultaneous torques, angular rotations, axial deformations recorded for the duration of the test. RESULTS: Herniation (posterior and posterior-lateral regions of the annulus) occurred with relatively modest joint compression but with highly repetitive flexion/extension moments. Increased magnitudes of axial compressive force resulted in more frequent and more severe disc injuries. CONCLUSIONS: The results support the notion that intervertebral disc herniation may be more linked to repeated flexion extension motions than applied joint compression, at least with younger, non-degenerated specimens. Relevance. While intervertebral disc herniations are observed clinically, consistent reproduction of this injury in the laboratory has been elusive. This study was designed to examine the biomechanical response and failure mechanics of spine motion segments to highly repetitive low magnitude complex loading. PMID- 11114442 TI - Female and male trunk geometry: size and prediction of the spine loading trunk muscles derived from MRI. AB - OBJECTIVE: Develop a gender specific database of trunk muscle cross-sectional areas across multiple levels of the thoracic and lumbar spine and develop prediction equations for the physiological cross-sectional area as a function of gender and anthropometry. DESIGN: This study quantified trunk muscle cross sectional areas of male and female spine loading muscles. BACKGROUND: There is a lack of comprehensive data regarding the female spine loading muscle size. Although biomechanical models often assume females are the same as males, little is known regarding gender differences in terms of trunk muscle areas and no data exist regarding the prediction of trunk muscle physiological cross-sectional areas from commonly used external anthropometric measures. METHODS: Magnetic resonance imaging scans through the vertebral bodies from T(8) through S(1) were performed on 20 females and 10 males. Muscle fiber angle corrected cross sectional areas were recorded at each vertebral level. Linear regression techniques taking into account anthropometric measures were utilized to develop prediction equations for the physiological cross-sectional area for each muscle of interest, as well as tests for differences in cross-sectional areas due to gender and side of the body. RESULTS: Significant gender differences were observed for the prediction of the erector spinae, internal and external obliques, psoas major and quadratus lumborum physiological cross-sectional areas. Anthropometric measures about the xyphoid process and combinations of height and weight resulted in better predictions of cross-sectional areas than when using traditional anthropometry. CONCLUSIONS: This study demonstrates that the trunk muscle geometry of females and males are different, and that these differences should be considered in the development of biomechanical models of the torso. Relevance. The prediction of physiological cross-sectional areas from external anthropometric measures provide gender specific equations to assist in estimation of forces of muscles which load the spine for biomechanical purposes. PMID- 11114443 TI - Injury initiation and progression in the anterior cruciate ligament. AB - OBJECTIVE: To develop a theoretical model to identify mechanisms by which total and partial tears of the anterior cruciate ligament could occur. DESIGN: A sagittal-plane knee model was used to investigate anterior cruciate ligament injury due to excessive anterior tibial translation. The ligament was modelled as an ordered array of fibres linking femur and tibia. BACKGROUND: Despite years of research, the detailed biomechanics of anterior cruciate ligament injury is not well understood. METHODS: A "critical strain criterion" was used to identify the onset and progression of model ligament fibre disruption. The associated forces were also calculated. RESULTS: At low flexion angles (<20 degrees ), the posterior fibre of the model ligament failed first, and the tear progressed anteriorly through the ligament. At higher flexion angles, the anterior fibre failed first, and the tear progressed posteriorly. Near the flexion angle at which the progression of injury changed direction, all fibres failed at approximately the same anterior tibial translation. At all but very high flexion angles, the force supported by the injured ligament was maximum when initial fibre failure occurred; the force then decreased with increasing anterior tibial translation. CONCLUSIONS: Near (20 degrees ) flexion, all model anterior cruciate ligament fibres fail at approximately the same anterior tibial translation, implying that a partial ligament tear may be impossible in this flexion region. Relevance. This study provides insight into possible mechanisms of initiation and progression of anterior cruciate ligament injury. It suggests that a partial tear of the posterior half of the ligament may be difficult to detect clinically. PMID- 11114444 TI - Quantified kinematics of the injury to the posterior cruciate ligament: a computer-aided design simulation study. AB - OBJECTIVE: To quantify the kinematics of the injury to the posterior cruciate and the other major knee ligaments as a function of the knee flexion angle at the moment of impact. DESIGN: Computer-aided design modelling was used to investigate the strain response of all major knee ligaments during antero-posterior abnormal tibio-femoral translation at 0-90 degrees knee flexion. BACKGROUND: It is generally believed that the likelihood of injury to the posterior cruciate ligament following anterior impact is higher in the flexed knee. However, there are no kinematical studies to quantify this clinical observation or investigate the role of the other knee ligaments in the above situation. METHODS: Computer calculations of the individual ligament strain were plotted against the magnitude of posterior tibial translation. Additionally, the strain rate for each ligament (defined as the ligament strain produced per mm of posterior tibial linear translation) was calculated as the slope of the strain-displacement curve for all tested degrees of knee flexion. RESULTS: The posterior cruciate ligament has been shown to be the primary restraint to posterior tibial translation in all degrees of knee flexion. However, at 90 degrees of knee flexion the strain rate of the posterior cruciate ligament is approximately half that in the fully extended knee and the posterior cruciate ligament is the only ligament to resist posterior tibial translation. CONCLUSIONS: The strain behaviour of the posterior cruciate ligament during injury is highly dependent on the knee flexion during the moment of impact. Forced posterior tibial translation in the 90 degrees flexed knee may result in isolated posterior cruciate ligament deficit rather than a complex ligament disruption. The strain rate of a ligament as introduced in the present study is a quantified parameter related to the resistance that the ligament imposes to an abnormal joint movement. Relevance. This study provides insight into the differential strain of the knee ligaments during impacts that result in posterior cruciate ligament injury. Studies that quantify the strain behaviour of individual knee ligaments are important to the understanding, diagnosis and prevention of injuries sustained during contact sports and high-energy road traffic accidents. PMID- 11114445 TI - Assessment of functional knee bracing: an in vivo three-dimensional kinematic analysis of the anterior cruciate deficient knee. AB - OBJECTIVE: To describe three-dimensional tibial and femoral movements in vivo and examine the effect of a brace on knee kinematics during moderate to intense activity. DESIGN: Skeletal kinematics of anterior cruciate ligament deficient knees was measured with and without braces during moderate to intense activity. BACKGROUND: Invasive markers implanted into the tibia and femur are the most accurate means to directly measure skeletal motion and may provide a more sensitive measure of the differences between brace conditions. METHODS: Steinmann traction pins were implanted into the femur and tibia of four subjects having a partial or complete anterior cruciate ligament rupture. Non-braced and braced conditions were randomly assigned and subjects jumped for maximal horizontal distance to sufficiently stress the anterior cruciate ligament. RESULTS: Intra subject peak vertical force and posterior shear force were generally consistent between conditions. Intra-subject kinematics was repeatable but linear displacements between brace conditions were small. Differences in angular and linear skeletal motion were observed across subjects. Bracing the anterior cruciate ligament deficient knee resulted in only minor kinematic changes in tibiofemoral joint motion. CONCLUSION: In this study, no consistent reductions in anterior tibial translations were observed as a function of the knee brace tested. Relevance. Investigations have reported that knee braces fail when high loads are encountered or when load is applied in an unpredictable manner. Questions remain regarding tibiofemoral joint motion, in particular linear displacements. The pin technique is a means for direct skeletal measurement and may provide a more sensitive measure of the differences between brace conditions. PMID- 11114446 TI - A spring network model for the analysis of load transfer and tissue reactions in intra-medullary fixation. AB - OBJECTIVE: A spring network can be used to represent the load transfer from a prosthetic stem into its surrounding bone. The study seeks to test the hypothesis that clinical patterns of bone remodelling can be simulated using a feedback that modifies the properties of the network depending on the load transfer. DESIGN: A mathematical model is used to simulate the initial properties of the linear system and its subsequent remodelling behaviour. BACKGROUND: A stable and pain free transfer of physiological forces is essential for a clinically successful arthroplasty. Following surgery, bone remodelling and osteolysis can modify this load transfer. METHODS: The combined effect of all factors that influence prosthesis-bone load transfer are summarised in the properties of 'inter-link' springs that connect springs representing the prosthesis and bone in the linear network. It is on these inter-links that a remodelling feedback operates, and their properties can be varied with time in response to deformation or force values. RESULTS: Reducing inter-link stiffness leads to a broad distribution of load transfer, whilst an iso-elastic stem concentrates this transfer through the proximal and distal portions of a prosthesis. Physiological patterns of bone resorption and osteolysis become apparent in a time-series analysis of the feedback in the linear system. Specifically, osseo-integration requires a fixation of sufficient stiffness otherwise loosening will occur. Simulated osteolysis following osseo-integration loosens the implant from a distal to a proximal direction. CONCLUSIONS: Complex physiological bone remodelling patterns can emerge from a simple feedback within a linear system. Relevance. Implant loosening is presented here as an adverse response of a stable dynamic system caused by mechanical or biological stimuli. PMID- 11114447 TI - A triaxial dynamometer to monitor lateral bending and axial rotation moments during static trunk extension efforts. AB - OBJECTIVE: The purpose of the study was to describe a new static triaxial dynamometer designed to monitor the lateral bending and axial rotation moments during trunk extension efforts. BACKGROUND: Most studies on back muscle function using electromyographic spectral analysis have not controlled moments produced about the three orthopaedic axes during trunk extension efforts. Criteria to control lateral bending and axial rotation moments during extension efforts have not been proposed in the literature. METHODS: Fourteen healthy subjects performed three trunk extension ramp contractions (0-100% of the maximal voluntary contraction). Triaxial L5/S1 moments at 20%, 40%, 60% and 80% of the maximal voluntary contraction in extension were extracted. RESULTS: During the extension efforts, the lateral bending and axial rotation moments at L5/S1 increased significantly across the force levels and reached 6.2 Nm (SD: 6.6) and 6.1 Nm (SD: 4.5), respectively, at 80% of the maximal voluntary contraction. Tolerance limits were proposed to control these associated efforts in the context of the electromyographic analysis of back muscles. Relevance. Simultaneous measurement of lateral bending and axial rotation moments at L5/S1 during extension efforts might help to explain and control load sharing between back muscles during extension efforts. PMID- 11114449 TI - In this issue PMID- 11114448 TI - Graphical presentation of the range of hip and knee rotations for clinical evaluation of gait. AB - OBJECTIVE: This paper reports a method of using a multi-axis graph to represent the range of rotations at the hip and knee joints during gait for clinical evaluation of a patient's performance. DESIGN AND METHODS: The multi-axis graph uses 12 symmetrically arranged axes to represent each component of range of rotations at the hip and knee joints of both legs. The range of joint rotations of thirteen normal subjects and two patients were measured using an electromagnetic motion tracking system with four foot switches. RESULTS: The range of joint rotations of normal subjects shows a symmetrical star in the multi axis graph. Abnormal function of a patient shows an asymmetrical star. CONCLUSIONS: This representation provides a simple way to examine a patient's performance in a time effective manner. Relevance. In clinical practice, the multi-axis representation of joint rotations can be used as an initial evaluation of a patient's performance to identify problems for further investigation. PMID- 11114450 TI - Virginia Apgar and the newborn Apgar score. PMID- 11114451 TI - Resuscitation in the hospital: relationship of year and rhythm to outcome. AB - OBJECTIVE: determine the frequency of initial rhythms in in-hospital resuscitation and examine its relationship to survival. Assess changes in outcome over time. METHODS: retrospective cohort (registry) including all admissions to the Medical Center of Central Georgia in which a resuscitation was attempted between 1 January, 1987 and 31 December, 1996. RESULTS: the registry includes 3327 admissions in which 3926 resuscitations were attempted. Only the first event is reported. There were 961 hospital survivors. Survival increased from 24.2% in 1987 to 33.4% in 1996 (chi(2)=39.0, df=1, P<0.0001). Survival was affected strongly by initial rhythm (chi(2)=420.0, df=1, P<0.0001) and decreased from 63.2% for supraventricular tachycardia (SVT) to 55.3% for ventricular tachycardia (VT), 51.0% for perfusing rhythms (PER), 34.8% for ventricular fibrillation (VF), 14.3% for pulseless electrical activity (PEA) and 10.0% for asystole (ASYS). PEA was the most frequent rhythm (1180 cases) followed by perfusing (963), asystole (580), VF (459), VT (94) and SVT (38). DISCUSSION: the powerful effect of initial rhythm on survival has been reported in pre-hospital and in-hospital resuscitation. VF is considered the dominant rhythm and generally accounts for the most survivors. We report good outcome for each; however, VF represents only 13.8% of events and 16.7% of survivors. PEA accounts for more survivors (169) than does VF (160). Our improved outcome is partially explained by changes in rhythms, but other institutional variables need to be identified to fully explain the results. Further studies are needed to see if our findings can be sustained or replicated. PMID- 11114452 TI - Neurological rehabilitation of severely disabled cardiac arrest survivors. Part I. Course of post-acute inpatient treatment. AB - BACKGROUND: Some survivors of out-of-hospital cardiac arrest (CA) sustain anoxic brain injury. The aim of this study was to offer these patients a new treatment approach, to describe the course and outcome of rehabilitation, and to judge whether rehabilitation provided benefit. METHODS: Twenty severely disabled patients (mean age 47.6 years, 17 M:3 F) were admitted for inpatient rehabilitation after sustaining anoxic brain damage secondary to CA. The multidisciplinary treatment approach aimed at orientation, communication, mobility, and self care. Function was assessed using Barthel index (BI) score. On discharge, placement and global outcome was noted. Medical charts of consecutive patients were reviewed retrospectively. RESULTS: Inpatient rehabilitation lasted on for average 12 weeks. Improvement in function was slow with a median increase of 1.88 BI score per week. Patients achieved clinically significant functional improvement as measured by pre-post comparison of BI (P<0.001). On discharge, overall disability was mild in 2 (10%), moderate in 7 (35%), and severe in 11 (55%) patients. CONCLUSION: Rehabilitation of selected CA survivors is appropriate, reducing the subsequent burden of care. Although in 55%, only minor dependence on care persisted, on a group level, the potential for rehabilitation was modest, and recovery curve was flat. Before admission, families should be given realistic information about the possible outcome, because independence was rarely achieved. PMID- 11114453 TI - Neurological rehabilitation of severely disabled cardiac arrest survivors. Part II. Life situation of patients and families after treatment. AB - BACKGROUND: About half of out-of-hospital cardiac arrest survivors experience secondary anoxic brain damage. Neurological outcome can be influenced by rehabilitative treatment approaches, but the nature and severity of persistent disabilities remain unclear. The aim of the study was to explore persistent neuropsychiatric symptoms, global function and life situation of these patients, and to evaluate quality of life in families. METHODS: 25 months after inpatient rehabilitation, 12 individuals (mean age=51 years; ten M: two F) attended a cross sectional interdisciplinary follow-up assessment with their carers. Function was investigated by clinical rating scales, neuropsychological standard tests, and clinical psychological inventories. Family members were asked about quality of life and satisfaction with social support. RESULTS: All patients had deficits in at least one or more cognitive areas such as orientation, memory, alertness, and awareness. Three different clinical syndromes were observed: very severe intellectual and physical impairment, (two), mild to moderate dementia, (five), and amnesic syndrome, (five). Prevalence of multiple disabilities, was high. A striking discrepancy was found between self and proxy rating of disabilities (P<0.01). Family members faced dramatically altered life situations after CA; 60% of spouses suffered from psychosomatic problems, 50% complained of lack of social support. CONCLUSION: Despite optimal in-hospital treatment, severe anoxic brain damage resulted in permanent cognitive decline, impaired awareness and self care ability. Families felt isolated, and more than half need more support to prevent burn out. PMID- 11114454 TI - Teaching cardiopulmonary resuscitation to CEGEP students in Quebec--a pilot project. AB - In order to increase CPR training in Quebec, we designed a pilot study to test out the efficacy of training CEGEP (junior college) students in CPR. We tried out four different methods of teaching CPR on students (Group A 'control', 4 h course, manikin to student ratio 1:4; Group B, 4 h course, manikin to student ratio 1:1; Group C, 2 h course, manikin to student ratio 1:1; Group D, video assisted CPR instruction, manikin to student ratio 1:1). CPR skills were tested on a computerized manikin at the end of the initial course and again at the end of the semester in order to evaluate short and long-term retention of skills. There were no significant differences between the test groups and the control group in terms of compressions or ventilations at the beginning and end of the semester, however groups C and D performed significantly better primary surveys (Airway, Breathing, Circulation - ABC sequences) during the initial testing. The most common reasons reported by students for not taking CPR courses were the cost of courses (49.2%) and the inconvenience of courses (26.2%), similarly the two most common incentives which could get students to take CPR courses were; free courses (65.6%) and greater accessibility of courses (54.1%). Video-assisted CPR training appears to be feasible, enjoyable and as, if not more effective than traditional CPR courses. Instituting a mandatory video-assisted CPR program in the CEGEP system in Quebec and in high schools and colleges throughout the world, would be a cost-effective way to train massive amounts of young people in CPR. PMID- 11114455 TI - Life supporting first aid (LSFA) teaching to Brazilians by television spots. AB - Accidents in developing countries are frequent and have high mortality and morbidity rates. In Brazil, in 1995-1996, the year of this study, life supporting first aid (LSFA), which includes cardiopulmonary resuscitation (CPR) basic life support (BLS) was not taught in schools. With the population of 165 million, the only way to teach the adult population on a large scale would be by television (TV), that is widely viewed. This study compares two groups of factory employees 86 controls without TV exposure to LSFA and 116 exposed to brief LSFA skill demonstrations on TV. Their ability to acquire eight LSFA skills was evaluated: external hemorrhage control; immobilization of a suspected forearm fracture; treatment of a skin burn by cold flush; body alignment after a fall; positioning for shock and coma; airway control by backward tilt of the head; and CPR (steps A B-C). Simulated skill performance on the evaluating nurse or manikin was tested at 1 week, 1 month, and 13 months. In the control group, 1-31% performed individual skills correctly; as compared to 9-96% of the television group (P<0.001). There was excellent retention over 13 months. Over 50% of the television group performed correctly five of the eight skills, including positioning and hemorrhage control. Television viewing increased correct airway control performance from 5 to 25% of trainees, while it remained at 3% in the control group. CPR-ABC performance, however, was very poor in both groups. We conclude that a significant proportion of factory workers can acquire simple LSFA skills through television viewing alone, except for the skill acquisition of CPR steps B (mouth-to-mouth ventilation) and C (external chest compressions) which need coached manikin practice. PMID- 11114456 TI - Does concurrent or previous illness accurately predict cardiac arrest survival? PMID- 11114457 TI - Attitudes towards CPR training and performance in family members of patients with heart disease. AB - Considering that heart patients may be at higher risk for cardiac arrest, this study was conducted to evaluate the preparedness and willingness of cardiac patient family members to perform cardiopulmonary resuscitation (CPR). A cross sectional survey of 100 family members of cardiac patients was conducted at a tertiary care emergency department over a 1.5-month period. Response rate was 95%. While 49% reported prior CPR training, only 7% trained within the past year. The majority received training (59%) because of a school or job requirement with only 8% trained because of 'concern for a family member.' The most frequent reasons for not being trained were 'never thought about it' or 'not interested' (57%). However, 49% of the untrained group did report an interest in future training. While 2% of respondents recalled a healthcare professional suggesting such training, 58% stated they would be influenced positively by such a recommendation. The most frequently reported barriers to performing CPR included fear of harming the patient or a lack of knowledge and skill to help. Despite a presumed higher risk for sudden cardiac death, most family members of cardiac patients do not maintain skills in basic CPR. Healthcare professionals may have the ability to significantly alter this concerning statistic through education and routine recommendations to patients' families. PMID- 11114458 TI - Is emergency open chest cardiopulmonary resuscitation accepted by patients' families? AB - Emergency open chest cardiopulmonary resuscitation (OCCPR) is sometimes performed on patients with cardiopulmonary arrest (CPA), especially those resulting from trauma. Since OCCPR is frequently carried out without the permission of patients' families, we surveyed the opinions of the families. A total of 1058 CPA patients were transferred to our department during the last 15 years. We sent questionnaires individually to the families of these patients to ask their opinions about OCCPR. The questionnaire provided the six questions allowing multiple answers; (1) Do you unconditionally agree with OCCPR? (2) Do you agree with OCCPR in children? (3) Do you agree with OCCPR in elderly patients? (4) Do you agree with OCCPR without permission from patient's families? (5) Do you entrust OCCPR to the doctors in charge? and (6) others. The questionnaire reached 846 families, of which 277 (32.7%) responded. The percentage response to each question was (1) 70.2, (2) 5.8, (3) 21.8, (4) 7.1, (5) 4.2 and (6) 5.0%. The younger the age of the responders the more they agreed with OCCPR. All the responders less than 30 years old agreed with the procedure. Of the 277 families, 95 had CPA patients treated with OCCPR. This group of families responded to six questions at the following rates: (1) 79.5, (2) 6.0, (3) 13.3, (4) 2. 4, (5) 4.8 and (6) 4.8%, suggesting that families with OCCPR patients are more cooperative to our treatment than those with non-OCCPR patients. The results of this study suggest that OCCPR in CPA patients is generally accepted by the patients' families, especially by young generations, although post-OCCPR careful explanation to patients' families is still indispensable. PMID- 11114459 TI - Minimally invasive direct cardiac massage versus closed-chest cardiopulmonary resuscitation in a porcine model of prolonged ventricular fibrillation cardiac arrest. AB - Open chest cardiac massage has been shown to be superior to closed-chest cardiopulmonary resuscitation for both hemodynamics produced during resuscitation and ultimate resuscitation success. The inexperience of many rescuers with emergency thoracotomy, along with the associated morbidity contributes to the continued reluctance in the use of invasive cardiopulmonary resuscitation techniques. A device has been developed for performing 'minimally invasive' direct cardiac massage. This technique was compared to standard closed-chest CPR for resuscitation results in 20 swine during prolonged ventricular fibrillation cardiac arrest. Minimally invasive direct cardiac massage was superior to closed chest CPR for return of spontaneous circulation (7/10 vs. 2/10; P<0.02) and coronary perfusion pressure at 30 min of CPR (17+/-9 vs. 6+/-6 mmHg; P<0.05). No significant injuries altering outcome were found with the invasive device. Throughout most of the time course of the study no significant differences in end tidal expired carbon dioxide levels were noted. Nor were there any differences in 24-h survival. Improvements in assuring proper placement of the device on the epicardium should make this technique a potent advanced cardiac life support adjunct. PMID- 11114460 TI - Interactions between CPR and defibrillation waveforms: effect on resumption of a perfusing rhythm after defibrillation. AB - BACKGROUND: Cardiopulmonary resuscitation (CPR) improves survival from cardiac arrest. The interactions between CPR and the new biphasic (BiP) defibrillation waveforms have not been defined. Our purpose was to compare the effect of CPR versus no CPR during BiP and damped sinusoidal (DS) shocks on the termination of ventricular fibrillation (VF) and the resumption of a perfusing rhythm. METHODS: We studied 20 pigs; VF was induced electrically and allowed to persist for 6 min. During VF episodes each pig received (in random order): (a) 6 min of full CPR (continuous ventilation and closed chest mechanical compression (Thumper, Michigan Instruments)) followed by DS defibrillation at 100 J; (b) no CPR, DS defibrillation; (c) 6 min of full CPR and BiP defibrillation at 100 J; and (d) no CPR, BiP defibrillation. RESULTS: BiP shocks with CPR terminated VF in 83% of attempts versus 45% without CPR (15/18 and 5/11 respectively, P<0.05). DS shocks with CPR were successful in terminating VF in 53% of attempts; DS shocks without CPR were successful in 44% (8/15 and 7/16, respectively, P=NS). No animal achieved a perfusing rhythm after shocks of either waveform if CPR did not precede the shocks during the 6-min VF period, whereas if CPR was administered during VF 46% (11/24) of the combined BiP/DS shocks restored a perfusing rhythm (P<0.01). CONCLUSION: In this experimental long duration VF model, CPR was essential for a perfusing rhythm after termination of VF by shocks with either waveform. CPR facilitated the termination of VF and resumption of a perfusing rhythm after biphasic waveform defibrillation but not after damped sinusoidal waveform defibrillation. PMID- 11114461 TI - Resuscitation research and emergency waiver of informed consent. PMID- 11114462 TI - Successful cardiopulmonary resuscitation outcome reviews. AB - An implicit question in every pre-hospital cardiopulmonary resuscitation (CPR) scenario is 'what will be the quality of life if a save is achieved?' This issue has implications for doctrine, policy, training and post-CPR counselling of both resuscitator and victim. Post-salvage neurological syndromes in surviving victims include amnesia, personality change, cognitive loss, depression, Parkinsonian syndromes, decorticate and decerebrate states and permanent brain damage with vegetative existence. Children who are salvaged by CPR rarely have pre-existing co-morbidities; but 75% of adults have pre-existing cardiac disease, cancer or diabetes. Such, of course, continue after a successful resuscitation. In the case of children who are resuscitated from acute hypoxic insults, the quality of life is generally good and, in the specific instance of survivors from near-drowning, some 95% will lead lives relatively unmodified. Although successful CPR resuscitation rates remain low in adults, the quality of life of those who leave hospital remains generally high. CPR involves two feature subjects, the resuscitator and the victim. Just as for the victim, so too the resuscitator's life is modified by CPR and its aftermath, whether immediate salvage has been achieved or not. This review addresses these issues, as a successful CPR (dramatic as it is) is not a conclusion but the beginning of a new phase of life for both resuscitator and victim. PMID- 11114463 TI - Successful defibrillation in profound hypothermia (core body temperature 25.6 degrees C). AB - We report a case of successful defibrillation in a severely hypothermic patient with a core body temperature of 25.6 degrees C as measured oesophageally. Ventricular fibrillation is a recognised life threatening arrhythmia in severely hypothermic patients. The traditional wisdom is that this arrhythmia is refractory to defibrillation at temperatures below 28 degrees C. PMID- 11114464 TI - A survey of basic life support training in various undergraduate health care professions. AB - Basic life support (BLS) is a core skill in which all healthcare professionals should be proficient. It is logical to provide BLS training during undergraduate years ensuring basic competence in all graduating healthcare students. Previous surveys of medical and dental schools have highlighted deficiencies in BLS training. This survey sought to assess the level of BLS training provided for students across a broad range of disciplines in the North West region of the UK. This included courses leading to an entry qualification into medicine, dentistry, nursing, midwifery or a profession allied to medicine (PAM). Information was collected by self-administered postal questionnaire with a response rate of 87%. The survey highlighted major variations in BLS training provided at undergraduate level across disciplines. PMID- 11114465 TI - Minimally invasive direct cardiac massage and defibrillation. AB - Direct massage of the heart may be performed using a specifically designed device inserted via a limited thoracic incision. The technique is simple to perform and has been shown in experimental studies to be physiologically similar to formal open-chest cardiac massage. Preliminary human studies have been encouraging and pre-hospital and in-hospital studies are in progress. Modification of the device allows combined epicardial-transthoracic defibrillation. Energy requirements for successful defibrillation using this method are considerably lower than standard external defibrillation. PMID- 11114466 TI - The European Resuscitation Council's paediatric life support course 'Advanced Paediatric Life Support'. AB - The poor outcome for resuscitation from cardiopulmonary arrest in childhood is widely recognised. The European Resuscitation Council has adopted the Advanced Paediatric Life Support course (originating in the UK and now available in a number of countries) as its course for providers caring for children. This paper outlines the course content and explains its remit, which is to reduce avoidable deaths in childhood by not only resuscitation from cardiac arrest but, more effectively, by recognising and treating in a timely and effective fashion life threatening illness and injury in infants and children. Two related courses Paediatric Life Support, a less intense course for less advanced providers, and Pre-Hospital Paediatric Life Support for immediate care providers are also described. PMID- 11114467 TI - Management of acute severe asthma. PMID- 11114468 TI - Rational use of vasoactive drugs after cardiac resuscitation: focus on inotropic agents. PMID- 11114469 TI - Associated techniques for the tracheal intubation. PMID- 11114470 TI - Human event-related potentials and distraction during selective listening. AB - Event-related brain potentials were recorded from healthy human subjects while they attended to one of two auditory stimulus channels (defined by location, left and right of a fixation point, and pitch) in order to detect rare target events. The distracting properties of periodic noise (vs. continuous noise, experiment 1) and backward speech (vs. forward speech, experiment 2) presented from a third speaker located behind the subjects were investigated. A typical attention effect with a larger negativity for attended tones was observed in both experiments. Backward speech led to a significantly reduced target detection rate for the first four stimuli after onset of the distractor accompanied by a reduced event related brain potential (ERP)-attention effect and a reduced fronto-central N2b component for the target stimuli. This indicates that irrelevant information leads to an attention decrement of about 1 s duration. PMID- 11114471 TI - Inhibitory effect of nicergoline on superoxide generation by activated rat microglias measured using a simple chemiluminescence method. AB - We evaluated the effect of nicergoline on superoxide production by rat microglias using a 2-methyl-6-(p-methoxyphenyl)-3, 7-dihydroimidazo[1,2-a]pyrazin-3-one dependent chemiluminescence assay. Nicergoline dose-dependently inhibited superoxide production by microglias stimulated with phorbol myristate acetate or opsonized zymosan, while it had no effect on superoxide production by a hypoxanthine-xanthine oxidase system. These results indicate that nicergoline does not have a scavenging effect, but has an inhibitory effect on superoxide generation by microglias. Although this drug is commonly used for treating chronic cerebral infarction, it may also have a protective effect on progression of Parkinson's disease or Alzheimer's disease. PMID- 11114472 TI - Nitric oxide synthase isoenzymes during in vitro development of rat neuronal and human non-neuronal cells. AB - We studied the expression of neuronal (n), endothelial (e) and inducible (i) nitric oxide synthase (NOS) in cell cultures of rat mesencephalic neurons (embryonic day 14), human keratinocytes from juvenile epidermis, human endothelial cells from juvenile coronary arteries, and human osteoblasts. All cell types were cultured for 5, 10 or 15 days. During proliferation (round cells without processes), the intracellular distribution and the intracellular amount of the calcium-dependent NOS isoforms (n- and e-NOS) did not change whereas the calcium-independent i-NOS changed from a cytosolic distribution pattern to compartmentalized distribution. A striking decrease of i-NOS immunoreactivity was measured by means of image analysis. Our results support the opinion that i-NOS acts as a switch between proliferation and differentiation of cells. PMID- 11114474 TI - Intracortical inhibitory circuits and sensory input: a study with transcranial magnetic stimulation in humans. AB - We compared intracortical inhibition (ICI) following paired transcranial magnetic stimulation (TMS) (interstimulus interval, ISI: 3 ms) and the inhibition of motor evoked potentials (MEPs) to TMS induced by stimulation of the median nerve (ISI: 200 ms) in six normal subjects. MEPs evoked by focal TMS were recorded in the relaxed opponens pollicis muscle and the size of the conditioned responses was expressed as a percentage of the size of the non-conditioned responses. Maximal ICI, ranging from 4 to 40%, and inhibition after median nerve stimulation, ranging from 11 to 68%, were significantly correlated. The results suggest that both phenomena are possibly mediated by the same gamma aminobutyric acid-ergic (GABAergic) inhibitory circuits and that afferent inputs to the cortex may contribute to their physiological activation. PMID- 11114473 TI - Regulation of rat hippocampal neural cadherin in the kainic acid induced seizures. AB - Regulation of neural (N-) cadherin expression in the hippocampus was examined by in situ hybridization and immunohistochemistry methods in the rat model of kainic acid (KA) induced seizures. After 12 and 24 h of KA administration, mRNA expression level of N-cadherin decreased in the hippocampal CA1 and CA3 area in parallel with decrease of the number of neural cells. In contrast, after 48 h and 7 days, mRNA expression level recovered partially, although the number of neural cells remained small. In addition, immunohistochemical staining indicated that N cadherin protein expression of survived neurons increased significantly after 48 h of KA administration. These results indicated that N-cadherin might be involved in neuronal reconstruction at the hippocampus. PMID- 11114475 TI - Acupuncture increases cell proliferation in dentate gyrus after transient global ischemia in gerbils. AB - The effects of acupuncture on cell proliferation in the dentate gyrus of gerbils after transient global ischemia were investigated in this study. Acupuncture was performed on Zusanli (ST36), which is a well known acupoint in animals and humans. In Oriental medicine, Zusanli has been commonly used for the enhancement of functional recovery in stroke patients. Through 5-bromo-2'-deoxyuridine (BrdU) immunohistochemistry, an increase in cell birth in the dentate gyrus of gerbils after ischemic injury was detected. Interestingly, acupunctural treatment in ischemic gerbils resulted in a significant increase in the number of BrdU positive cells in the dentate gyrus. The present findings indicate that acupuncture may affect cell proliferation in the dentate gyrus of gerbils after ischemic injury. PMID- 11114476 TI - Exaggerated responses to chronic nociceptive stimuli and enhancement of N-methyl D-aspartate receptor-mediated synaptic transmission in mutant mice lacking D amino-acid oxidase. AB - Formalin-induced nociceptive behaviors and N-methyl-D-aspartate (NMDA) subtype glutamate receptor-mediated excitatory synaptic transmission were analyzed in mutant mice lacking D-amino-acid oxidase, which catalyzes the oxidative deamination of D-amino acids. The second phase of the formalin-induced licking response, a part of which is known to be mediated by NMDA receptors in the spinal cord, was significantly augmented in mutant mice. NMDA receptor-mediated excitatory postsynaptic currents recorded from spinal cord dorsal horn neurons by tight-seal whole-cell methods were significantly potentiated in mutant mice. The present observations provide another line of evidence that D-serine functions as an endogenous coagonist at the glycine site of NMDA receptors, and raise the possibility that D-amino-acid oxidase exerts a neuromodulatory function by controlling the concentration of D-serine in the central nervous system. PMID- 11114477 TI - Domain-specific distribution of working memory processes along human prefrontal and parietal cortices: a functional magnetic resonance imaging study. AB - This study reinvestigated the functional neuroanatomy of phonological and visual working memory in humans. Articulatory suppression was used to deprive the human subjects of species-specific verbal strategies in order to make the functional magnetic resonance imaging results more comparable to findings in non-human primates. Both phonological and visual working memory processes activated similar prefronto-parietal networks but were found to be differentially distributed along several cortical structures, in particular along the anterior and posterior parts of the intermediate frontal sulcus. These results suggest that a domain-specific topographical organization of neural working memory mechanisms in the primate brain is conserved in evolution. However, the findings also underline the critical dynamic influence that the additional availability of language may have on working memory processes and their functional implementation in the human brain. PMID- 11114478 TI - Okadaic acid-induced upregulation of nitrotyrosine and heme oxygenase-1 in rat cortical neuron cultures. AB - Hyperphosphorylation of tau is a characteristic feature of the neurodegenerative pathology in Alzheimer's disease (AD). Okadaic acid (OA) is currently used in models of AD research to increase the phosphorylation of tau. Using immunocytochemistry and fluorescent study, we found that markers of oxidative activity such as nitrotyrosine, c-jun, 2',7'-dichlorofluorescein diacetate (DCF), and heme oxygenase-1 (HO-1) were altered in OA-treated culture. Immunoreactivity of nitrotyrosine and c-jun, and DCF-oxidation were increased in degenerating neurons, while HO-1 expression was increased in astrocyte in response to OA. The data suggest that tau phosphorylation and oxidative damage be implicated in OA induced neurodegeneration. PMID- 11114479 TI - Helium-oxygen pressure induces striatal glutamate increase: a microdialysis study in freely-moving rats. AB - In rat, helium pressures induce locomotor and motor activity which requires dopaminergic and N-methyl-D-aspartate (NMDA) receptor activities at striatal level. However, biochemical studies have suggested that pressure exposure may increase striatal glutamate level. We used microdialysis technique to study the effects of pressure on glutamate level in the striatum and the effects of local administration of D1 (SCH23390) or D2 (sulpiride) on these changes. Pressures increase both glutamate and glutamine levels in striatal microdialysates. Administration of sulpiride (1 microM) or SCH23390 (1 microM) by reverse microdialysis did not affect significantly pressure induced glutamate increase. So, protective effects of D1 and D2 antagonists against locomotor and motor hyperactivity (LMA) are probably independent of the processes involved in the striatal glutamate increase evoked by pressure. PMID- 11114480 TI - A visuomotor reaction time task increases the irregularity and complexity of inspiratory airflow pattern in man. AB - Previous studies have suggested that motor cortex may be involved in complex control of respiration in man. In this study, we estimated the influence of a simple visuomotor reaction time task (VMT) on the inspiratory airflow curve (IFC) by comparing measurements of standard deviation (SD), C(0) complexity and approximate entropy (ApEn) of the IFC at rest and during VMT conditions. It was found that both C(0) complexity and ApEn, but not SD of the IFC were significantly increased during the VMT challenge. The results suggest that visuomotor pathways may modulate respiratory motor pathways, because an increase in ApEn is suggestive of increased coupling or communication between (motor) systems. PMID- 11114481 TI - Reduction of gamma-aminobutyric acid-ergic neurotransmission as a putative mechanism of radiation induced activation of the gonadotropin releasing-hormone pulse generator leading to precocious puberty in female rats. AB - Brain irradiation in prepubertal children with malignomas can cause precocious puberty. A selective cranial cobalt (Co(60))-irradiation technique has been developed in rats. In two experiments early juvenile (13-15 days old) female rats received a single dose of 5 Gy or sham irradiation. At pubertal age (post-natal days 33-34) irradiated rats had higher serum estradiol and luteinizing hormone levels. In experiment 1 irradiated rats had higher gonadotropin releasing-hormone (GnRH) mRNA levels in the preoptic area compared to controls (P<0.05). In experiment 2 the release rates of gamma-aminobutyric acid (GABA) in vitro from preoptic mediobasal hypothalamic areas of irradiated rats were significantly reduced after stimulation with the GABA(A) receptor agonist muscimol (maximum values 4607+/-804 vs. 7399+/-1048 pM in controls, mean+/-SEM, P<0.05). Radiation induced central precocious puberty might be caused by damage to inhibitory GABAergic neurons leading to premature activation of the GnRH-pulse generator. PMID- 11114482 TI - Not growth associated protein GAP-43 (B-50), but its fragment GAP-43-3 (B-60) predominates in rat brain during development. AB - The participation of the nerve termini growth associated protein GAP-43 in neurite outgrowth and targeting is well documented. Commonly, besides GAP-43 itself, two big fragments devoid of four (GAP-43-2, IB-50) and of about 40 (GAP 43-3, B-60) N-terminal residues were co-isolated from brain. In adult brain, GAP 43 significantly prevails over the fragments. To find their relative amounts during development, rat brain proteins were isolated on different stages of embryonal and post-natal development and subjected to gel electrophoresis in 0.9 M acetic acid-2.5 M urea system. The bands of GAP-43 protein family were detected on Western blots. We show that in developing brain (until 5th post-natal day), a proteolysis of GAP-43 near Ser(41) that results in GAP-43-3 accumulation is activated. We hypothesize that just the functions that can be performed by the GAP-43 fragments are of importance for developing brain. PMID- 11114484 TI - The organization of rapid eye movement activity during rapid eye movement sleep is further impaired in very old human subjects. AB - Rapid eye movement activity (REMA) during rapid eye movement (REM) sleep was studied in seven very old ('old-old') subjects. Beyond global quantitative features (REMA density), we evaluated the organizational aspects of REMA, that is its occurrence in burst mode, which were compared to a group of younger elderly subjects ('young-old'). REMA density in 'old-old' subjects is not significantly different from that of 'young-old' subjects. The same lack of difference in the two groups is found for the number of REMA bursts. By contrast, the duration of REMA bursts is reduced in the 'old-old' subjects, as well as the 'burst state-to burst-state' probability, i.e. the probability for successive inter-REMA time intervals to be part of the same REMA burst. Our results clearly show that the trend towards an impairment of REMA organizational aspects continues with aging. This is consistent with the hypothesis that sleep disorganization keeps worsening with age. However, it is of interest to observe that the capability of producing REMA bursts is preserved despite aging. PMID- 11114483 TI - Differential sensitivities to the lethal, but not the neurotoxic, effects of p chloroamphetamine in inbred rat strains. AB - The Lewis, Fischer 344, Brown Norway, Spontaneously Hypertensive, and Wistar Kyoto inbred rat strains were, respectively, compared for the lethal and neurotoxic effects of acute p-chloroamphetamine (PCA, 2.5-10 mg/kg i.p.). The lethal properties of the amphetamine were recorded within 24 h after its administration whereas neurotoxicity (as assessed by frontocortical 5 hydroxytryptamine (5-HT) reuptake and 5-HT transporter binding assays) was analyzed 1 week after PCA administration. Preliminary experiments indicated that neither the rapid hyperlocomotor and/or the hypoexploratory effect of PCA nor the in vitro potency of PCA to inhibit frontocortical [(3)H]5-HT reuptake varied between strains. On the other hand, strain differences were observed with respect to the rapid fatal effects of the 5 and 10 mg/kg doses of PCA administration. Lastly, frontocortical [(3)H]5-HT reuptake and [(3)H]citalopram binding at 5-HT transporters diminished in a dose-dependent, but strain-independent, manner 1 week after the acute injection of PCA. This study reveals an independency between the mechanisms underlying the fatal effects of PCA on the one hand, and the long term damaging effects of PCA on serotonergic neurons on the other hand. PMID- 11114485 TI - Effects of pleasant and unpleasant ambient odors on human voice pitch. AB - The perception of odors is well identified as having strong emotional correlates. It is also well known that the acoustic characteristics of the voice differ according to the emotional state. This study compared some acoustic features of the voice of 18 subjects reading the same text in pleasant (lavender) and unpleasant (pyridine) ambient odor conditions. The results revealed that the pitch of the voice was higher in the pleasant than in the unpleasant condition. These findings are consistent with the hypothesis of local and functional convergences of encoding vocal emotion and hedonic perception of odors. PMID- 11114486 TI - Foreword PMID- 11114487 TI - Managing depression and anxiety in the elderly patient. AB - Depression in the elderly is under-recognised and under-treated, even more than in younger patients. Late-life depression is a chronic and disabling illness, but there is a common misconception that it is a normal feature of ageing. It can also be difficult to differentiate depression from coexisting anxiety and cognitive disorders. Antidepressants are as effective in acute treatment in older, as in younger, depressed adults. Maintenance antidepressant therapy also appears as effective in older patients. It may, however, be difficult to attain therapeutic levels of tricylic antidepressants (TCAs) in the elderly because of side effects. The selective serotonin re-uptake inhibitors (SSRIs) have more favourable safety and tolerability profiles, making them a suitable treatment option for older patients, who are more vulnerable to adverse effects. In selecting an SSRI for treating late-life depression, its pharmacokinetic profile and effects on coexisting illnesses that are prevalent in the elderly, such as anxiety, dementia and cardiovascular disease, also need to be considered. PMID- 11114488 TI - Treatment of depression and concomitant anxiety. AB - The prevalence of depression and concomitant anxiety is high in the community. Patients with depression and concomitant anxiety experience increased functional disability, increased disruption to social, work and family life, and frequently report more symptoms. Treatment needs to be primarily efficacious for depression and secondly for anxiety; both sets of symptoms require prompt and effective treatment. Although benzodiazepines are still prescribed for depression and associated anxiety, they are essentially ineffective in treating depression and therefore are inappropriate. The anxiolytic effect of tricyclic antidepressants (TCAs) takes longer to develop than their antidepressant effect and their adverse tolerability profile can hinder treatment compliance. Selective serotonin re uptake inhibitors (SSRIs) are a newer class of antidepressants that are as effective as TCAs in treating depression and are well tolerated in the long term and have demonstrated efficacy in the treatment of anxiety. This paper will provide evidence to demonstrate the short- and long-term efficacy of SSRIs in the treatment of depression and comorbid anxiety, with most clinical evidence supporting the anxiolytic profile of the SSRI paroxetine. PMID- 11114489 TI - Depressive illness and disability. AB - Depression is a prevalent disorder that imposes a severe burden of functional disability. With the exception of advanced coronary artery disease, depression is more disabling than the many other chronic medical illnesses often encountered in primary care. Depressed patients also suffer reduced workplace productivity, take more days off work due to illness and are very high users of healthcare resources. Where depression is comorbid with anxiety, the burden of disability and cost increases considerably. Treatment of depressed patients results in a lower level of disability and increased workplace productivity. Moreover, reduced usage of healthcare resources can offset the costs of the treatment. Depression is frequently under-recognized or misdiagnosed in the primary care setting, and many patients do not receive appropriate treatment that could help relieve their disability. Improving the recognition and treatment of depression is essential to reduce the burden of disability, on depressed patients and on society as a whole. PMID- 11114490 TI - Depression in Europe: experience from the DEPRES II survey. Depression Research in European Society. AB - Despite being one of the most prevalent psychiatric conditions in the community, depression is commonly unrecognised in clinical practice. The Depression Research in European Society (DEPRES) survey examined depression in a pan-European population (N=78463) and identified a 6-month prevalence of depression of 17%. In DEPRES II (N=1884), more than 50% of the individuals were categorised as being currently depressed, with one-third receiving antidepressant treatment. Cluster analysis grouped patients within six clearly differentiated types. Individuals with patient type 'severe depression and anxiety' (Group III), had the greatest number of symptoms and were more likely to be taking antidepressants than the other patient groups. In Group III, depression prevented individuals from undertaking normal activities for 6 weeks and prevented them from working for 1 month. The patient type making the most demands on healthcare resources are those with depression and anxiety. Prompt and effective treatment would be of benefit to all patient types, and the use of a selective serotonin reuptake inhibitor (SSRI) with activity against anxiety symptoms is an appropriate management strategy. PMID- 11114491 TI - Structural issues and policy in the management of depressive illness. Overview and expert panel commentary. AB - Opening with an overview of the current state of knowledge regarding management of depression and anxiety spectrum disorders, this paper then reports commentary from the expert panel on the question: When there are effective treatments, why are optimal outcomes not achieved in clinical practice? PMID- 11114492 TI - Hippocampal perfusion in mild Alzheimer's disease. AB - Perfusion and metabolic studies in patients with Alzheimer's disease (AD) have so far yielded conflicting results on the functional status of the hippocampal region, whose deep location in the brain makes it critical to optimize the image reconstruction technique employed in emission tomography. We used a brain dedicated device (CERASPECT) to perform single photon emission computed tomography (SPECT) studies with 99mTc-hexamethylpropylene-amine-oxime in 22 consecutive patients (mean age: 74+/-6.5 years) with mild [mini-mental status examination (MMSE) score > or =15, mean 20.8+/-3.2], probable AD. The control subjects were 11 healthy elderly people (mean age: 70.5+/-6.5 years). In patients, the total score on the selective reminding test (SRT) was used as an index of memory function. Counts from a hippocampal and a temporoparietal region of interest in each hemisphere were referred to the average thalamic counts. To optimize SPECT images, we used conventional filtered back-projection (FBP) reconstruction and a new iterative method of conjugate gradients (CG), which takes into account the geometrical and physical characteristics of the gamma camera. Hippocampal perfusion in the two hemispheres was significantly lower in patients than in control subjects, regardless of which reconstruction method was used, and correlated with the MMSE score. The correlation between hippocampal perfusion and the SRT score was significantly (bootstrap procedure) higher with the CG method than with the FBP method (CG: r=0.52 and 0.54; FBP: r=0.39 and 0.47, for the right and left hemisphere, respectively). These results show hippocampal hypoperfusion in patients with mild AD, a correlation between hippocampal perfusion and the severity of cognitive impairment, and enhanced identification of these subtle perfusional changes with the use of an alternative image-reconstruction method that improves the spatial resolution of SPECT images. PMID- 11114493 TI - Regional cerebral blood flow changes in drug-resistant depressed patients following treatment with transcranial magnetic stimulation: a statistical parametric mapping analysis. AB - Changes of regional cerebral blood flow (rCBF) in five drug-resistant depressed patients were examined by single photon emission computed tomography (SPECT) with 99mTc-hexamethylpropyleneamine oxime (99mTc-HMPAO) before and after treatment with transcranial magnetic stimulation (TMS). The SPECT images were analysed with the Statistical Parametric Mapping (SPM) package. TMS administered in the region of the left dorsolateral prefrontal cortex (DLPFC) of the depressed patients was associated with an increase of rCBF at a focal region some distance from the stimulation site. No change was observed at any other remote region. PMID- 11114494 TI - Effect of the 5-HT(1A) partial agonist buspirone on regional brain electrical activity in man: a functional neuroimaging study using low-resolution electromagnetic tomography (LORETA). AB - In a double-blind, placebo-controlled study, the effects of 20 mg buspirone - a 5 HT(1A) partial agonist - on regional electrical generators within the human brain were investigated utilizing three-dimensional EEG tomography. Nineteen-channel vigilance-controlled EEG recordings were carried out in 20 healthy subjects before and 1, 2, 4, 6 and 8 h after drug intake. Low-resolution electromagnetic tomography (LORETA; Key Institute for Brain-Mind Research, software: http://www.keyinst.unizh.ch) was computed from spectrally analyzed EEG data, and differences between drug- and placebo-induced changes were displayed as statistical parametric maps. Data were registered to the Talairach-Tournoux human brain atlas available as a digitized MRI (McConnell Brain Imaging Centre: http://www.bic.mni.mcgill.ca). At the pharmacodynamic peak (1st hour), buspirone increased theta and decreased fast alpha and beta sources. Areas of theta increase were mainly the left temporo-occipito-parietal and left prefrontal cortices, which is consistent with PET studies on buspirone-induced decreases in regional cerebral blood flow and fenfluramine-induced serotonin activation demonstrated by changes in regional cerebral glucose metabolism. In later hours (8th hour) with lower buspirone plasma levels, delta, theta, slow alpha and fast beta decreased, predominantly in the prefrontal and anterior limbic lobe. Whereas the results of the 1st hour speak for a slight CNS sedation (more in the sense of relaxation), those obtained in the 8th hour indicate activation. Thus, LORETA may provide useful and direct information on drug-induced changes in central nervous system function in man. PMID- 11114496 TI - Web alert. Proteins catalysis and regulation. AB - A selection of World Wide Web sites relevant to reviews published in this issue of Current Opinion in Structural Biology. PMID- 11114495 TI - Cerebral cortex: a topographic segmentation method using magnetic resonance imaging. AB - Remarkable developments in magnetic resonance imaging (MRI) technology provide a broad range of potential applications to explore in vivo morphological characteristics of the human cerebral cortex. MR-based parcellation methods of the cerebral cortex may clarify the structural anomalies in specific brain subregions that reflect underlying neuropathological processes in brain illnesses. The present study describes detailed guidelines for the parcellation of the cerebral cortex into 41 subregions. Our method conserves the topographic uniqueness of individual brains and is based on our ability to visualize the three orthogonal planes, the triangulated gray matter isosurface and the three dimensional (3D) rendered brain simultaneously. Based upon topographic landmarks of individual sulci, every subregion was manually segmented on a set of serial coronal or transaxial slices consecutively. The reliability study indicated that the cerebral cortex could be parcelled reliably; intraclass correlation coefficients for each subregion ranged from 0.60 to 0.99. The validity of the method is supported by the fact that gyral subdivisions are similar to regions delineated in functional imaging studies conducted in our center. Ultimately, this method will permit us to detect subtle morphometric impairments or to find abnormal patterns of functional activation in circumscribed cortical subregions. The description of a thorough map of regional structural and functional cortical abnormalities will provide further insight into the role that different subregions play in the pathophysiology of brain illnesses. PMID- 11114497 TI - Where is the information in structural space?. Editorial overview. PMID- 11114498 TI - The end of the beginning: structural studies of ribosomal proteins. AB - Work on the structural biology of ribosomes has progressed rapidly over the past few years. It has come to a stage at which the structures of the individual components are no longer of interest, except for those that still present ambiguous information about their structure because of conformational dynamics, as well as for those that show very little homology with their counterparts from other species or other kingdoms. The recently solved structure of protein L7/L12 and its proposed modes of dimerization have helped to understand the structural flexibility of this protein, which occurs as two dimers in the ribosome. The structure provides a missing link for many previous biochemical and functional studies. PMID- 11114499 TI - EF-hand calcium-binding proteins. AB - The EF-hand motif is the most common calcium-binding motif found in proteins. Several high-resolution structures containing different metal ions bound to EF hand sites have given new insight into the modulation of their binding affinities. Recently determined structures of members of several newly identified protein families that contain the EF-hand motif in some of their domains, as well as of their complexes with target molecules, are throwing light on the surprising variety of functions that can be served by this simple and ingenious structural motif. PMID- 11114500 TI - The tumour necrosis factor receptor family: life or death choices. AB - The past twelve months have seen a renewal of interest in the therapeutic potential of members of the tumour necrosis factor receptor family and their cytokine ligands. This biomedical interest has spawned a number of structural studies, which have significantly deepened our understanding of the molecular basis for the function of these cell-surface signalling systems. The fresh data have revealed unexpected mechanisms conferring ligand-receptor specificity and have highlighted the structural requirements for the initiation of intracellular signalling. PMID- 11114501 TI - Caspases: key players in programmed cell death. AB - Research in apoptosis has established a central role for caspases. The recent determination of structures of caspase-1, caspase-3 and caspase-8, together with biochemical studies, has greatly enhanced our understanding of the structure, function and specificity of these enzymes. This provides a basis for the further elucidation of the biological role of caspases and a guide to the design of selective inhibitors to treat caspase-mediated diseases. PMID- 11114502 TI - Structural basis of cell-cell interactions in the immune system. AB - During the past year, advances in our understanding of receptor-ligand interactions between opposing cell surfaces have occurred at a structural level. These include adhesion involving CD2-CD58, antigen-specific T-cell receptor interactions with peptides bound to major histocompatibility complex molecules (both pMHCI and pMHCII), the CD8alphaalpha co-receptor-pMHCI interaction and the binding of two distinct classes of natural killer receptors to self-MHC ligands. PMID- 11114503 TI - Protein-protein interactions in intracellular membrane fusion. AB - The fusion of intracellular vesicles with their target membranes is an essential feature of the compartmental structure of eukaryotic cells. This process requires proteins that dictate the targeting of a vesicle to the correct cellular location, mediate bilayer fusion and, in some systems, regulate the precise time at which fusion occurs. Recent biophysical and structural studies of these proteins have begun to provide a foundation for understanding their functions at a molecular level. PMID- 11114504 TI - The rotary mechanism of ATP synthase. AB - Since the chemiosmotic theory was proposed by Peter Mitchell in the 1960s, a major objective has been to elucidate the mechanism of coupling of the transmembrane proton motive force, created by respiration or photosynthesis, to the synthesis of ATP from ADP and inorganic phosphate. Recently, significant progress has been made towards establishing the complete structure of ATP synthase and revealing its mechanism. The X-ray structure of the F(1) catalytic domain has been completed and an electron density map of the F(1)-c(10) subcomplex has provided a glimpse of the motor in the membrane domain. Direct microscopic observation of rotation has been extended to F(1)-ATPase and F(1)F(o) ATPase complexes. PMID- 11114505 TI - AB(5) toxins: structures and inhibitor design. AB - High-resolution crystal structures of AB(5) toxins in their native form or in complex with a variety of ligands have led to the structure-based design and discovery of inhibitors targeting different areas of the toxins. The most significant progress is the development of highly potent multivalent ligands that block binding of the toxins to their receptors. PMID- 11114506 TI - The rhamnose pathway. AB - L-Rhamnose is a deoxy sugar found widely in bacteria and plants. Evidence continues to emerge about its essential role in many pathogenic bacteria. The crystal structures of two of the four enzymes involved in its biosynthetic pathway have been reported and the other two have been submitted for publication. This pathway does not exist in humans, making enzymes of this pathway very attractive targets for therapeutic intervention. PMID- 11114507 TI - Novel reactions catalysed by antibodies. AB - New structural data on nonhydrolytic antibody catalysts gained over the past two years confirm that antibodies elicited against transition-state analogues function by differential stabilisation of the transition-state over the ground state through electrostatic, van der Waals, cation-pi and hydrogen-bonding interactions. The lack of chemical catalysis correlates with the low catalytic efficiency. Novel strategies that precisely position a key functional residue in the antibody catalyst combining site have therefore emerged, as demonstrated by crystallographic studies. Whereas antibodies with a bulky residue at position H100c of hypervariable loop H3 adopt different cavity shapes, other antibodies share a common deep combining site. This structural restriction might reflect the use of similar hydrophobic haptens to generate the antibody; novel hapten design or new immunisation strategies may, in the future, lead to more structurally diversified active sites. PMID- 11114508 TI - Clarity through structures. Editorial overview. PMID- 11114509 TI - The structural basis for the remarkable catalytic proficiency of orotidine 5' monophosphate decarboxylase. AB - The three-dimensional structures of orotidine 5'-monophosphate decarboxylases from four different organisms have been determined by X-ray crystallography. The structures reveal an active site in which the pyrimidine base and phosphate groups are rigidly held in place. Surprisingly, both pyrimidine carbonyl groups are hydrogen bonded to amide groups, rather than to strong active site acids, as was previously predicted. The positioning of a conserved aspartate sidechain close to the substrate carboxylate and a conserved lysine ammonium group close to the C6 of the pyrimidine suggests a novel mechanism to explain the extreme catalytic proficiency of this enzyme. PMID- 11114510 TI - Combining structural genomics and enzymology: completing the picture in metabolic pathways and enzyme active sites. AB - An important goal of structural genomics is to complete the structural analysis of all the enzymes in metabolic pathways and to understand the structural similarities and differences. A preliminary glimpse of this type of analysis was achieved before structural genomics efforts with the glycolytic pathway and efforts are underway for many other pathways, including that of catecholamine metabolism. Structural enzymology necessitates a complete structural characterization, even for highly homologous proteins (greater than 80% sequence homology), as every active site has distinct structural features and it is these active site differences that distinguish one enzyme from another. Short cuts with homology modeling cannot be taken with our current knowledge base. Each enzyme structure in a pathway needs to be determined, including structures containing bound substrates, cofactors, products and transition state analogs, in order to obtain a complete structural and functional understanding of pathway-related enzymes. PMID- 11114511 TI - Ribonucleotide reductases: the link between an RNA and a DNA world? AB - The structures of a class III ribonucleotide reductase (RNR) and pyruvate formate lyase exhibit striking homology within their active site domains with respect to each other and to the previously published structure of a class I RNR. The common structures and the common complex-radical-based chemistry of these systems, as well as of the class II RNRs, suggest that RNRs evolved by divergent evolution and provide an essential link between the RNA and DNA world. PMID- 11114512 TI - Floundering about at cell membranes: a structural view of phospholipid signaling. AB - Structures are now available for the majority of the enzyme families involved in the phosphorylation, dephosphorylation and hydrolysis of signaling phospholipids. Lipid kinase and phosphatase structures recapitulate catalytic motifs involved in protein phosphorylation and dephosphorylation, whereas cytosolic phospholipase A(2) manifests novel catalytic geometry. Structures have been determined for most known intracellular phospholipid 'receptor' domains, both those that bind membrane-embedded phospholipids and those that bind lipid monomers. PMID- 11114513 TI - Trapping intermediates in the crystal: ligand binding to myoglobin. AB - Crystal structures of the reactive short-lived species that occur in chemical or binding reactions can be determined using X-ray crystallography via time-resolved or kinetic trapping approaches. Recently, various kinetic trapping methods have been used to determine the structure of intermediates in ligand binding to myoglobin. PMID- 11114514 TI - The Caenorhabditis elegans heterochronic gene lin-29 coordinates the vulval uterine-epidermal connections. AB - BACKGROUND: The development of a connection between the uterus and the vulva in the nematode Caenorhabditis elegans requires specification of a uterine cell called the utse, and its attachment to the vulva and the epidermal seam cells. The uterine pi cells generate the utse and uv1 cells, which also connect the uterus to the vulva. The uterine anchor cell (AC) induces the vulva through LIN 3/epidermal growth factor (EGF) signaling, and the pi cells through LIN-12/Notch signaling. Here, we report that a gene required for seam cell maturation is also required for specification of the utse and for vulval differentiation, and thus helps to coordinate development of the vulval-uterine-seam cell connection. RESULTS: We cloned the egl-29 gene, which is necessary for induction of uterine pi cells, and found it to be allelic to lin-29, which encodes a zinc finger transcription factor that is necessary for the terminal differentiation of epidermal seam cells. In the uterus, lin-29 functioned upstream of lin-12 in the induction of pi cells and was necessary to maintain expression in the AC of lag 2, which encodes a ligand for LIN-12. CONCLUSIONS: The lin-29 gene controls gene expression in the epidermal seam cells, uterus and vulva, and may help to coordinate the terminal development of these three tissues by regulating the timing of late gene expression during organogenesis. PMID- 11114515 TI - Myosin VI is required for asymmetric segregation of cellular components during C. elegans spermatogenesis. AB - BACKGROUND: The asymmetric division of cells and unequal allocation of cell contents is essential for correct development. This process of active segregation is poorly understood but in many instances has been shown to depend on the cytoskeleton. Motor proteins moving along actin filaments and microtubules are logical candidates to provide the motive force for asymmetric sorting of cell contents. The role of myosins in such processes has been suggested, but few examples of their involvement are known. RESULTS: Analysis of a Caenorhabditis elegans class VI myosin deletion mutant reveals a role for this motor protein in the segregation of cell components during spermatogenesis. Mutant spermatocytes cannot efficiently deliver mitochondria and endoplasmic reticulum/Golgi-derived fibrous-body membranous organelle complexes to budding spermatids, and fail to remove actin filaments and microtubules from the spermatids. The segregation defects are not due to a global sorting failure as nuclear inheritance is unaffected. CONCLUSIONS: C. elegans myosin VI has an important role in the unequal partitioning of both organelles and cytoskeletal components, a novel role for this class of motor protein. PMID- 11114516 TI - The role of the proteins Kar9 and Myo2 in orienting the mitotic spindle of budding yeast. AB - BACKGROUND: Two genetic 'pathways' contribute to the fidelity of nuclear segregation during the process of budding in the yeast Saccharomyces cerevisiae. An early pathway, involving Kar9p and other proteins, orients the mitotic spindle along the mother-bud axis. Upon the onset of anaphase, cytoplasmic dynein provides the motive force for nuclear movement into the bud. Loss of either pathway results in nuclear-migration defects; loss of both is lethal. Here, to visualize the functional steps leading to correct spindle orientation along the mother-bud axis, we imaged live yeast cells expressing Kar9p and dynein as green fluorescent protein fusions. RESULTS: Transport of Kar9p into the bud was found to require the myosin Myo2p. Kar9p interacted with microtubules through the microtubule-binding protein Bim1p and facilitated microtubule penetration into the bud. Once microtubules entered the bud, Kar9p provided a platform for microtubule capture at the bud cortex. Kar9p was also observed at sites of microtubule shortening in the bud, suggesting that Kar9p couples microtubule shortening to nuclear migration. CONCLUSIONS: Thus, Kar9p provides a key link between the actin cytoskeleton and microtubules early in the cell cycle. A cooperative mechanism between Kar9p and Myo2p facilitates the pre-anaphase orientation of the spindle. Later, Kar9p couples microtubule disassembly with nuclear migration. PMID- 11114517 TI - R7 photoreceptor specification requires Notch activity. AB - The eight photoreceptors in each ommatidium of the Drosophila eye are assembled by a process of recruitment [1,2]. First, the R8 cell is singled out, and then subsequent photoreceptors are added in pairs (R2 and R5, R3 and R4, R1 and R6) until the final R7 cell acquires a neuronal fate. R7 development requires the Sevenless receptor tyrosine kinase which is activated by a ligand from R8 [3]. Here, we report that the specification of R7 requires a second signal that activates Notch. We found that a Notch target gene is expressed in R7 shortly after recruitment. When Notch activity was reduced, the cell was misrouted to an R1/R6 fate. Conversely, when activated Notch was present in the R1/R6 cells, it caused them to adopt R7 fates or, occasionally, cone cell fates. In this context, Notch activity appears to act co-operatively, rather than antagonistically, with the receptor tyrosine kinase/Ras pathway in R7 photoreceptor specification. We propose two models: a ratchet model in which Notch would allow cells to remain competent to respond to sequential rounds of Ras signalling, and a combinatorial model in which Notch and Ras signalling would act together to regulate genes that determine cell fate. PMID- 11114518 TI - FGF-8 stimulates neuronal differentiation through FGFR-4a and interferes with mesoderm induction in Xenopus embryos. AB - The role of fibroblast growth factors (FGFs) in neural induction is controversial [1,2]. Although FGF signalling has been implicated in early neural induction [3 5], a late role for FGFs in neural development is not well established. Indeed, it is thought that FGFs induce a precursor cell fate but are not able to induce neuronal differentiation or late neural markers [6-8]. It is also not known whether the same or distinct FGFs and FGF receptors (FGFRs) mediate the effects on mesoderm and neural development. We report that Xenopus embryos expressing ectopic FGF-8 develop an abundance of ectopic neurons that extend to the ventral, non-neural, ectoderm, but show no ectopic or enhanced notochord or somitic markers. FGF-8 inhibited the expression of an early mesoderm marker, Xbra, in contrast to eFGF, which induced ectopic Xbra robustly and neuronal differentiation weakly. The effect of FGF-8 on neurogenesis was blocked by dominant-negative FGFR-4a (DeltaXFGFR-4a). Endogenous neurogenesis was also blocked by DeltaXFGFR-4a and less efficiently by dominant-negative FGFR-1 (XFD), suggesting that it depends preferentially on signalling through FGFR-4a. The results suggest that FGF-8 and FGFR-4a signalling promotes neurogenesis and, unlike other FGFs, FGF-8 interferes with mesoderm induction. Thus, different FGFs show specificity for mesoderm induction versus neurogenesis and this may be mediated, at least in part, by the use of distinct receptors. PMID- 11114519 TI - Directed differentiation of dendritic cells from mouse embryonic stem cells. AB - Dendritic cells (DCs) are uniquely capable of presenting antigen to naive T cells, either eliciting immunity [1] or ensuring self-tolerance [2]. This property identifies DCs as potential candidates for enhancing responses to foreign [3] and tumour antigens [4], and as targets for immune intervention in the treatment of autoimmunity and allograft rejection [1]. Realisation of their therapeutic potential would be greatly facilitated by a fuller understanding of the function of DC-specific genes, a goal that has frequently proven elusive because of the paucity of stable lines of DCs that retain their unique properties, and the inherent resistance of primary DCs to genetic modification. Protocols for the genetic manipulation of embryonic stem (ES) cells are, by contrast, well established [5], as is their capacity to differentiate into a wide variety of cell types in vitro, including many of hematopoietic origin [6]. Here, we report the establishment, from mouse ES cells, of long-term cultures of immature DCs that share many characteristics with macrophages, but acquire, upon maturation, the allostimulatory capacity and surface phenotype of classical DCs, including expression of CD11c, major histocompatibility complex (MHC) class II and co-stimulatory molecules. This novel source should prove valuable for the generation of primary, untransformed DCs in which candidate genes have been overexpressed or functionally ablated, while providing insights into the earliest stages of DC ontogeny. PMID- 11114520 TI - Yeast Eap1p, an eIF4E-associated protein, has a separate function involving genetic stability. AB - A rate-limiting step during translation initiation in eukaryotic cells involves binding of the initiation factor eIF4E to the 7-methylguanosine-containing cap of mRNAs. Overexpression of eIF4E leads to malignant transformation [1-3], and eIF4E is elevated in many human cancers [4-7]. In mammalian cells, three eIF4E-binding proteins each interact with eIF4E and inhibit its function [8-10]. In yeast, EAP1 encodes a protein that binds eIF4E and inhibits cap-dependent translation in vitro [11]. A point mutation in the canonical eIF4E-binding motif of Eap1p blocks its interaction with eIF4E [11]. Here, we characterized the genetic interactions between EAP1 and NDC1, a gene whose function is required for duplication of the spindle pole body (SPB) [12], the centrosome-equivalent organelle in yeast that functions as the centrosome. We found that the deletion of EAP1 is lethal when combined with the ndc1-1 mutation. Mutations in NDC1 or altered NDC1 gene dosage lead to genetic instability [13,14]. Yeast strains lacking EAP1 also exhibit genetic instability. We tested whether these phenotypes are due to loss of EAP1 function in regulating translation. We found that both the synthetic lethal phenotype and the genetic instability phenotypes are rescued by a mutant allele of EAP1 that is unable to bind eIF4E. Our findings suggest that Eap1p carries out an eIF4E-independent function to maintain genetic stability, most likely involving SPBs. PMID- 11114521 TI - The protein tyrosine phosphatase Shp-2 regulates RhoA activity. AB - Remodeling of filamentous actin into distinct arrangements is precisely controlled by members of the Rho family of small GTPases [1]. A well characterized member of this family is RhoA, whose activation results in reorganization of the cytoskeleton into thick actin stress fibers terminating in integrin-rich focal adhesions [2]. Regulation of RhoA is required to maintain adhesion in stationary cells, but is also critical for cell spreading and migration [3]. Despite its biological importance, the signaling events leading to RhoA activation are not fully understood. Several independent studies have implicated tyrosine phosphorylation as a critical event upstream of RhoA [4]. Consistent with this, our recent studies have demonstrated the existence of a protein tyrosine phosphatase (PTPase), sensitive to the dipeptide aldehyde calpeptin, acting upstream of RhoA [5]. Here we identify the SH2 (Src homology region 2)-containing PTPase Shp-2 as a calpeptin-sensitive PTPase, and show that calpeptin interferes with the catalytic activity of Shp-2 in vitro and with Shp-2 signaling in vivo. Finally, we show that perturbation of Shp-2 activity by a variety of genetic manipulations results in raised levels of active RhoA. Together, these studies identify Shp-2 as a PTPase acting upstream of RhoA to regulate its activity and contribute to the coordinated control of cell movement. PMID- 11114522 TI - An oncogenic role of sphingosine kinase. AB - Sphingosine kinase (SphK) is a highly conserved lipid kinase that phosphorylates sphingosine to form sphingosine-1-phosphate (S1P). S1P/SphK has been implicated as a signalling pathway to regulate diverse cellular functions [1-3], including cell growth, proliferation and survival [4-8]. We report that cells overexpressing SphK have increased enzymatic activity and acquire the transformed phenotype, as determined by focus formation, colony growth in soft agar and the ability to form tumours in NOD/SCID mice. This is the first demonstration that a wild-type lipid kinase gene acts as an oncogene. Using a chemical inhibitor of SphK, or an SphK mutant that inhibits enzyme activation, we found that SphK activity is involved in oncogenic H-Ras-mediated transformation, suggesting a novel signalling pathway for Ras activation. The findings not only point to a new signalling pathway in transformation but also to the potential of SphK inhibitors in cancer therapy. PMID- 11114523 TI - Higher concentrations of histone macroH2A in the Barr body are correlated with higher nucleosome density. AB - Histone macroH2A, which is a subtype of histone H2A, possesses a histone H2A-like portion fused to a relatively long non-histone portion. MacroH2A has been shown to associate preferentially with the inactive X chromosome [1]. To investigate the specificity of this association, the nuclear distribution of macroH2A was compared with that of regular core histones. In normal human female fibroblasts, all anti-histone antibodies that were tested (including anti-macroH2A antibody) preferentially labeled the inactive X chromosome. Moreover, when expressed as green fluorescent protein (GFP) fusions, both histone H2A and macroH2A were concentrated in the Barr body. These data clearly show the presence of a higher density of nucleosomes in the inactive X chromosome. Accordingly, the specificity of the macroH2A association with the inactive X chromosome should be reconsidered. While investigating the role of macroH2A, we found that the proximity of the non-histone region of macroH2A to a promoter could lead to a specific repression of transcription, suggesting that the incorporation of macroH2A into chromatin might help to establish the stable pattern of gene expression in differentiated cells. PMID- 11114524 TI - Interaction of the Arabidopsis polycomb group proteins FIE and MEA mediates their common phenotypes. AB - Genes of the FERTILISATION INDEPENDENT SEED (FIS) class regulate cell proliferation during reproductive development in Arabidopsis [1-5]. The FIS genes FERTILISATION INDEPENDENT ENDOSPERM (FIE) and MEDEA (MEA) encode homologs of animal Polycomb group (Pc-G) proteins, transcriptional regulators that modify chromatin structure and are thought to form multimeric complexes [3-11]. To test whether similarities in fis mutant phenotypes reflect interactions between their protein products, we characterised FIE RNA and protein localisation in vivo, and FIE protein interactions in yeast and in vitro. Expression of FIE mRNA overlaps with that of MEA during embryo sac and seed development and is unaffected in mea mutants. Results from the yeast two-hybrid system and an in vitro pull-down assay indicate that MEA and FIE proteins interact. The relevance of this interaction in vivo is supported by the finding that FIE and MEA co-localise in the nucleus in transfected plant cells. Interaction of MEA and FIE is mediated by the amino terminal region of MEA. Despite sequence divergence in this domain, MEA can interact with its corresponding animal partner Extrasexcombs (ESC) in the yeast two-hybrid system. We conclude that FIE and MEA act together as part of a multimeric complex and that this accounts for the similarities in mutant phenotypes. We propose that an ancient mechanism for chromatin modification has been independently recruited to different developmental processes in the two kingdoms. PMID- 11114525 TI - Salmonella typhimurium proliferates and establishes a persistent infection in the intestine of Caenorhabditis elegans. AB - Genetic analysis of host-pathogen interactions has been hampered by the lack of genetically tractable models of such interactions. We showed previously that the human opportunistic pathogen Pseudomonas aeruginosa kills Caenorhabditis elegans, that P. aeruginosa and C. elegans genes can be identified that affect this killing, and that most of these P. aeruginosa genes are also important for mammalian pathogenesis. Here, we show that Salmonella typhimurium as well as other Salmonella enterica serovars including S. enteritidis and S. dublin can also kill C. elegans. When C. elegans is placed on a lawn of S. typhimurium, the bacteria accumulate in the lumen of the worm intestine and the nematodes die over the course of several days. This killing requires contact with live bacterial cells. The worms die with similar kinetics when placed on a lawn of S. typhimurium for a relatively short time (3-5 hours) before transfer to a lawn of E. coli. After the transfer to E. coli, a high titer of S. typhimurium persists in the C. elegans intestinal lumen for the rest of the worms' life. Furthermore, feeding for 5 hours on a 1:1000 mixture of S. typhimurium and E. coli followed by transfer to 100% E. coli, also led to death after several days. This killing correlated with an increase in the titer of S. typhimurium in the C. elegans lumen, which reached 10,000 bacteria per worm. These data indicate that, in contrast to P. aeruginosa, a small inoculum of S. typhimurium can proliferate in the C. elegans intestine and establish a persistent infection. S. typhimurium mutated in the PhoP/PhoQ signal transduction system caused significantly less killing of C. elegans. PMID- 11114526 TI - Caenorhabditis elegans is a model host for Salmonella typhimurium. AB - The idea of using simple, genetically tractable host organisms to study the virulence mechanisms of pathogens dates back at least to the work of Darmon and Depraitere [1]. They proposed using the predatory amoeba Dictyostelium discoideum as a model host, an approach that has proved to be valid in the case of the intracellular pathogen Legionella pneumophila [2]. Research from the Ausubel laboratory has clearly established the nematode Caenorhabditis elegans as an attractive model host for the study of Pseudomonas aeruginosa pathogenesis [3]. P. aeruginosa is a bacterium that is capable of infecting plants, insects and mammals. Other pathogens with a similarly broad host range have also been shown to infect C. elegans [3,4]. Nevertheless, the need to determine the universality of C. elegans as a model host, especially with regards pathogens that have a naturally restricted host specificity, has rightly been expressed [5]. We report here that the enterobacterium Salmonella typhimurium, generally considered to be a highly adapted pathogen with a narrow range of target hosts [6], is capable of infecting and killing C. elegans. Furthermore, mutant strains that exhibit a reduced virulence in mammals were also attenuated for their virulence in C. elegans, showing that the nematode may constitute a useful model system for the study of this important human pathogen. PMID- 11114527 TI - Hindsight blinkers Britain's mad cow disease response. PMID- 11114528 TI - Peering into review. PMID- 11114529 TI - The 3F3/2 anti-phosphoepitope antibody binds the mitotically phosphorylated anaphase-promoting complex/cyclosome. PMID- 11114530 TI - G-protein signaling: satisfying the basic necessities of life. AB - Recent studies have shed light on the role of G-protein signaling, and in particular the regulatory RGS proteins, in behavioral adaptations of the round worm Caenorhabditis elegans; similar signaling pathways underlie analogous physiology and behaviors in mammals. PMID- 11114531 TI - Developmental genetics: vertebrates and insects see eye to eye. AB - The results of a number of recent studies indicate that eye development in insects and vertebrates may have more features in common than hitherto suspected. The results support the possibility that insect and vertebrate eyes evolved from a complex ancestral organ. PMID- 11114532 TI - Microtubule dynamics: the view from the tip. AB - Recent studies have suggested that proteins found at the tips of microtubules in vertebrate cells may play an important role in intracellular membrane transport processes. Evidence from fission yeast indicates that such proteins can also regulate microtubule dynamics. PMID- 11114533 TI - Multisensory integration: attending to seen and felt hands. AB - The neglect of one side of space exhibited by some brain-damaged patients can be ameliorated by cueing the patient to the neglected side of space. A related effect has been found to depend on the hand being seen and felt at the same time. The results add to a growing literature on somatosensory-visual interactions. PMID- 11114534 TI - Evolutionary genomics: is Buchnera a bacterium or an organelle? AB - The first genome sequence of an intracellular bacterial symbiont of a eukaryotic cell has been determined. The Buchnera genome shares features with the genomes of both intracellular pathogenic bacteria and eukaryotic organelles, and it may represent an intermediate between the two. PMID- 11114535 TI - Membrane biology: membrane-regulated transcription. AB - The activation of membrane-bound transcription factors involves release from the membrane by proteolysis. Recent studies show that, for some proteins, cleavage is performed by the proteasome, whereas others require specific proteases. PMID- 11114536 TI - Genetic imprinting: silencing elements have their say. AB - Tissue-specific silencing elements have been identified that are required for imprinting of the individual genes in the Igf2-H19 domain of the mouse genome. These elements further elaborate the differences between the two parental chromosomes, and add a new feature to parent-of-origin-specific gene regulatory complexes. PMID- 11114537 TI - Membrane targeting: what a difference a G makes. AB - Pleckstrin homology domains are modular domains that direct membrane targeting of their host proteins by binding to polyphosphoinositides; recent results have increased our appreciation of how some of these domains actually bind 3 phosphoinositides, and along the way thrown up some unexpected observations. PMID- 11114539 TI - Nuclear export of yeast signal recognition particle lacking srp54p by the Xpo1p/Crm1p NES-dependent pathway PMID- 11114538 TI - Comparative genetics: a third model nematode species. AB - Recent studies have introduced Oscheius sp. CEW1 as a third nematode species accessible to genetic analysis, joining the better known Caenorhabditis elegans and Pristionchus pacificus. A group of vulva-defective mutants in Oscheius has been identified, with defects not seen in C. elegans. PMID- 11114540 TI - The dynamics of velocity adaptation in human vision PMID- 11114541 TI - A permissive function of phosphoinositide 3-kinase in ras activation mediated by inhibition of GTPase-activating proteins PMID- 11114542 TI - Phosphorylation of the pro-apoptotic protein BAD on serine 155, a novel site, contributes to cell survival PMID- 11114543 TI - Ventricular dysfunction clinical research in infants, children and adolescents. AB - These two issues of Progress in Pediatric Cardiology comprehensively illustrate the wealth of currently available information on the pathophysiology of heart failure, age-related myocardial responsiveness, energy metabolism, cardiopulmonary interactions, the pressure-volume relationship, the systemic inflammatory response, the management of heart failure, pediatric pharmacology, the use of heart failure therapies including digoxin, ACE inhibitors, beta adrenergic blockers, inotropic agents, diuretics, vasodilators, calcium sensitizers, angiotensin and aldosterone receptor blockers, growth hormone, and future gene therapy. The etiology and course of ventricular dysfunction in children is poorly characterized. Furthermore, many changing developmental properties of the pediatric myocardium and differences in the etiologies of ventricular dysfunction in children compared with adults are illustrated in these articles, invalidating the concept that children can safely be considered small adults for the purpose of understanding heart failure pathophysiology and treatment. However, these articles reveal that strikingly little research in children with ventricular dysfunction exists in terms of well-designed large scale studies of the epidemiology or multicenter controlled clinical therapeutic trials. A future research agenda is proposed to improve understanding etiologies, course and treatment of ventricular dysfunction in children that is based on organized and funded cooperative groups since no one pediatric cardiac center treats enough children with a particular etiology of ventricular dysfunction. In conclusion, significant understanding of basic mechanisms of pediatric ventricular dysfunction and effective therapies for adults with ventricular dysfunction exist. A multicenter pediatric cardiac ventricular dysfunction network would allow improved understanding of diseases and treatments, and result in evidence-based medicine for pediatric patients with ventricular dysfunction. PMID- 11114544 TI - Clinical pharmacology research in the pediatric patient: the challenge continues. AB - For most of the 20th century, most drugs labeled by the United States Food and Drug Administration (USFDA) have not been adequately studied in the pediatric population. This lack of data has necessitated the continued dependence of practitioners on sub-optimal prescribing data placing pediatric patients at great risk of serious therapeutic misadventures. Recently, the USFDA has enacted and begun to enforce the Final Rule of 1997 which became effective on 1 April 1999. This rule is the culmination of the persistent efforts of numerous professional organizations, clinicians, academicians, the USFDA and others, to ensure the ready availability of appropriate data for medications intended for or that will be used in children. Unlike the 1994 Rule which voluntarily required pharmaceutical manufacturers to submit pediatric data, the Final Rule mandates submission of such data and, most importantly, empowers the USFDA to afford incentives and penalties for non-compliance including possible removal of already marketed products. This overview addresses many of the important components which must be included in the performance of a comprehensive clinical pharmacologic evaluation serving as the foundation for optimal dosing across the broad age range encompassing pediatric practice. Furthermore, the possible risk and/or benefits of the study must be reasonably defined prior to undertaking the study and clearly shared with the patient's caregivers. Consent should always be obtained from the caregiver and, when appropriate, assent obtained from the underage child. To facilitate such clinical investigations and to foster collaborative efforts with innovators and clinical research programs, the National Institutes of Health through the National Institute of Child Health and Human Development of the NIH established a network of Pediatric Pharmacology Research Units. These units have worked closely together and with other pediatric research centers to facilitate USFDA labeling of a number of commonly used medications. All of these very positive efforts highlight the many challenges that remain for the pediatric investigator and practitioner while underscoring the very positive environment in support of these efforts. PMID- 11114545 TI - Digitalis use in children: an uncertain future. AB - The therapeutic indications for digoxin in the treatment of children with large left-to-right shunts continue to be reassessed. New insights into the alterations in cardiac function imposed by this hemodynamic burden have shown preserved systolic performance. Pharmacological interventions that improve cardiac output by afterload reduction or other modalities have proven useful and potentially have low risk for serious toxicity. Early and successful surgical treatment of most conditions causing pulmonary overcirculation has shortened the duration of medical management and prevented many of the untoward complications of this pathology. As new agents are forthcoming to treat various cardiac conditions, use by-tradition of cardiac glycosides has appropriately diminished. While the understanding of complicated molecular aspects of cation transport have been enhanced by the unique actions of cardiac glycosides, their clinical utility has decreased. This report summarizes studies on the use of cardiac glycosides in the treatment of large left-to-right shunts in children. PMID- 11114546 TI - Diuretic therapy of heart failure in infants and children. AB - Diuretics are the mainstay of traditional therapy for congestive heart failure. The syndrome of heart failure is now understood to involve complex interactions of neurohumoral substances released in response to poor cardiac function. Developmental changes during infancy and childhood will affect both the activation of systemic neurohumoral responses and the pharmacokinetic and pharmacodynamic actions of diuretics. Few human studies directly evaluate the efficacy of diuretic therapy in heart failure in adults. The pharmacodevelopmental aspects of diuretic therapy in infants and children are also incompletely studied. This review will describe the kidney's role in the pathogenesis of sodium and water retention in heart failure and the developmental changes in the kidney related to fluid retention. Known principles of diuretic therapy in congestive heart failure will be described. All these factors can then be used by the reader to evaluate the role of diuretic therapy in the complex syndrome of heart failure in infants and children. PMID- 11114547 TI - Pharmacology of inotropic agents in infants and children. AB - Inotropic agents are drugs which increase the stroke work of the heart at a given pre-load and after-load. All of these agents work through a final common pathway involving the modulation of calcium interactions with various myocardial contractile proteins. The agents employed with pediatric patients include the cardial glycosides, catecholamine beta-agonists and the selective phosphodiesterase III inhibitors. Digoxin is the prototypic cardiac glycoside which has a long history of safe and effective use in infants and children. Its utility in improving right ventricular dysfunction in patients with cor pulmonale leading to biventricular dysfunction makes it ideally suited to the pediatric population. Monitoring digoxin pharmacokinetics in infants is confounded by the presence of an endogenous digoxin-like substance. Nevertheless, the drug is well suited for subacute and chronic myocardial support. In contrast, the catecholamines are the drugs of choice for acute intervention. Their pharmacokinetics permit rapid dosing titration. In infants and children the greatest experience has been accrued with dopamine, a mixed alpha- and beta agonist but both epinephreine and norepinephrine are being used with increasing frequency as the need for drugs with increased potency and pressor activity becomes more common. The phosphodiesterase inhibitors amrinone and milrinone are the newest additions to our therapeutic armamentarium. In addition to their modest inotropic effects, amrinone and to a greater extent, milrinone offer significant pulmonary vasodilatation as part of their therapeutic package. These effects occur with little or any impact on myocardial oxygen consumpton while their lusitropic effects enhance relaxation in hypertrophied ventricular muscle. Of the two agents milrinone is probably preferred due to its greater therapeutic index and shorter elimination half-life. All of these agents remain important tools in the care of critically ill infants and children. The rational use of these drugs based upon their pharmacokinetic and pharmacodynamic properties is essential to achieve their optimal effects. PMID- 11114548 TI - Vasodilators in the treatment of pediatric heart failure. AB - The goals of heart failure therapy have shifted from purely hemodynamic manipulation to a combination of hemodynamic and neurohumoral modulation. Vasodilators with neurohumoral modulatory properties [such as ACE inhibitors (ACEi) and third generation beta-blockers] have become the cornerstone of chronic heart failure therapy. These newer agents have proven to improve morbidity and mortality in adults with chronic heart failure. Pure vasodilators still have a place in the treatment of acute decompensated heart failure and in patients who are intolerant to ACEi or beta-blocker therapy. In decompensated heart failure management, improvement of cardiac output is of paramount importance and restoration of normal hemodynamics takes priority over modulation of cardiac maladaptation. Under these circumstances agents that improve contractility and modify cardiac preload and afterload are used. In the intensive care unit setting inodilators offer the advantage of an added positive inotropic effect. NO donors play an important role when close titration of blood pressure is also needed. It is the purpose of this manuscript to address principles and current practice regarding the use of vasodilators in pediatric heart failure. ACE inhibitors and third generation beta-blockers due to their importance in today's therapeutic approach to heart failure are the focus of more detailed articles in this issue of Progress in Pediatric Cardiology. PMID- 11114549 TI - Angiotensin-converting enzyme inhibitor therapy for ventricular dysfunction in infants, children and adolescents: a review. AB - Angiotensin-converting enzyme (ACE) inhibitors have become an important part of the pharmacologic armamentarium in the battle against treatment of ventricular dysfunction. There have been a number of large controlled, randomized trials in adults with both asymptomatic and symptomatic ventricular dysfunction, which confirm the safety and efficacy of this category of drugs for the treatment of this potentially lethal condition. ACE inhibitors may be used to treat infants, children and adolescents with asymptomatic and symptomatic ventricular dysfunction as well. The data supporting their use in children is less complete than that concerning the treatment of adults. We review here the various causes of ventricular dysfunction and congestive heart failure (CHF) in infants, children, and adolescents; the data available regarding treatment of these conditions with ACE inhibitors, and the safety and efficacy of these drugs for the various conditions. The pharmacokinetics and proposed mechanisms of action of ACE inhibitors in children are reviewed, as are speculated long-term results of ACE inhibitor use in cohorts of growing children. Recommendations are made for future studies. PMID- 11114550 TI - Beta-adrenergic blockers in the treatment of pediatric heart failure. AB - Beta-adrenergic blocking agents for the treatment of chronic congestive heart failure in adults have gained wide acceptance. Although the exact mechanism of their effect is still not entirely clear, the preponderance of data support the hypothesis that the primary mechanism of action of beta-blocking agents in chronic heart failure is to prevent and reverse adrenergically-mediated intrinsic myocardial dysfunction and remodeling. Large, multicenter trials in adults with chronic congestive heart failure have definitively shown that beta-blockers improve left ventricular ejection fraction, symptoms, and survival when compared to placebo. Although experience with beta-blockers in children is limited, anecdotal evidence suggests that children with chronic congestive heart failure may also benefit from this therapy. Large trials in children are currently in the planning stages in order to attempt to determine the indications, dosages, and optimal use of beta-blockers in children with chronic congestive heart failure. PMID- 11114551 TI - New drugs in the treatment of heart failure. AB - New therapeutic strategies as well as the development of drugs with more specific targets have been fueled by disappointments in the treatment of adult heart failure. Calcium sensitizers, vesnarinone and angiotensin channel blockers will be addressed in this manuscript. The physiologic and pharmacologic principles that justify their use in the management of heart failure are reviewed. Calcium sensitizers increase myocardial contractility and in part they bypass the adenylyl cyclase cascade, which gives them a more favorable energy profile. Vesnarinone is a quinolinone derivative with ion channel modulation properties, which result in a positive inotropic effect and prolongation of the action potential. In addition vesnarinone has immunomodulatory properties. Angiotensin converting enzyme inhibitors are the cornerstones for the treatment of heart failure. The discovery of some putative drawbacks to ACE inhibition has challenged this supremacy. Angiotensin receptor blockers have been developed hoping to overcome these deficiencies. Myocardial developmental differences highlight the shortcomings of attempting to extrapolate data on drugs and cellular physiology in adults to children. Studies are needed addressing standards of care, quality of life, morbidity and mortality, neurohumoral activation, its modulation and the consequences of these therapies in pediatric heart failure. PMID- 11114552 TI - Exogenous growth hormone: a new therapy for dilated cardiomyopathy. AB - Heart failure is an epidemic within the United States and, despite current medical therapy, carries a high mortality rate. Growth hormone and insulin-like growth factor-1 have known direct effects on the cardiovascular system. Improvement in contractility, reduction in wall stress, and increase in cardiac performance have been noted in animal experiments. Furthermore, preliminary data from human trials are encouraging. This report outlines the biology of growth hormone, the experimental and human data to support clinical trials of growth hormone treatment, and the outcome of trials reported to date. PMID- 11114553 TI - Prospects for gene therapy for inherited cardiomyopathies. AB - Over the last few years the genes responsible for a number of genetic diseases of the cardiovascular system have been identified. These have included X-linked and autosomal dominant dilated cardiomyopathy, and hypertrophic cardiomyopathy. Genetic heterogeneity has been described in both of these diseases but a commonality of function has been apparent: defects in cytoskeletal proteins cause dilated cardiomyopathy and mutations in sarcomeric proteins cause hypertrophic cardiomyopathy. This led us to develop a 'final common pathway' hypothesis as a framework for selecting candidate genes for mutation screening in families with these diseases. The characterization of gene mutations has led to the development of therapies specifically targeting the defective protein or the pathway in which it is involved. These have included the use of pharmaceutical agents to replace or to antagonize the mutated protein, and replacement of the defective gene with a functional one (gene therapy). While early studies using gene therapy vectors were promising, translating studies in animals to viable therapeutic options for humans has remained problematic. There have been many publications describing the use of vectors to transduce target cells for the correction of gene defects, including recombinant retroviruses, adenoviruses, and adeno-associated viruses, as well as non-viral vectors. In this review we will discuss the identification of gene defects associated with cardiomyopathies, and the potential of gene therapy for the treatment of these diseases, as well as addressing some concerns related to the use of adenovirus-based vectors, a virus known to be an etiologic agent of acquired dilated cardiomyopathy. PMID- 11114554 TI - Next generation therapeutics. Drug discovery in the 21st Century Editorial overview. PMID- 11114555 TI - Introduction PMID- 11114556 TI - Outcome in 104 pubovaginal slings using freeze-dried allograft fascia lata from a single tissue bank. AB - We describe our experience with the use of allograft fascia lata for the treatment of stress urinary incontinence. One hundred and four patients underwent allograft fascia lata pubovaginal slings. Preoperatively, all were evaluated by a detailed urogynecologic evaluation, voiding diary, and pelvic examination. The pubovaginal sling was performed using a 2x15-cm freeze-dried nonirradiated cadaveric fascia lata specimen. Outcome measures were assessed by a urogynecologic questionnaire, pad usage, and disease-specific quality-of-life questionnaires. Eighty-eight percent (91 of 104) responded to a mailed urogynecology and disease-specific quality-of-life questionnaire with an average follow-up period of 19. 4 +/- 10.3 months. The mean preoperative daily pad usage was 4.6 +/- 3.0, postoperatively pad usage was 1.1 +/- 1.4 (P < 0.0001). Urge incontinence resolved in 41% (n = 24) of the 59 patients who complained of this preoperatively. Eighty-seven percent of the responders indicated that urinary incontinence was not substantially affecting their daily life. As in our preliminary report, the use of freeze-dried allograft pubovaginal sling continues to provide good results without adverse outcomes. A prospective, randomized comparison of autologous versus allograft slings and a review of preparation techniques used by tissue banks are needed. PMID- 11114557 TI - A new technique for cystocele repair and transvaginal sling: the cadaveric prolapse repair and sling (CAPS). AB - A new technique using cadaveric fascia lata for the simultaneous repair of a cystocele and placement of a pubovaginal sling by means of a transvaginal approach is described, and our early results are reported. We refer to this as the cadaveric prolapse repair with sling (CaPS). Fifty patients, ages 37 to 90 years, underwent a new technique for simultaneous cystocele repair and transvaginal pubovaginal sling using a single piece of cadaveric fascia. Maximum follow-up was 6 months (range 1 to 6). A 6 x 8 cm segment of cadaveric fascia lata is placed transvaginally to repair the defect through which the bladder herniates into the vagina and to provide sling support at the bladder neck/proximal urethra. The sling is anchored to the pubic bone with transvaginal bone anchors. The remainder of the fascia is then secured to the medial edge of the levator muscles/pubocervical fascia bilaterally and at the vaginal cuff or cervix with absorbable sutures to reduce the cystocele. Patients are being evaluated with preoperative and postoperative stress, emptying, anatomy, protection, instability (SEAPI) scores as well as with grading of the prolapse based on a 3-grade anatomic classification system. Presenting symptoms have included stress urinary incontinence (SUI) in 13 (26%), urge incontinence in 4 (8%), mixed incontinence in 6 (12%), and pelvic prolapse in 20 (40%). These symptoms are not mutually exclusive; some patients presented with a combination of symptoms. The mean SEAPI scores were 5.51 preoperatively and 0.63 postoperatively, representing a significant improvement (P <0.001). Of the 40 patients whose prolapse was quantified, 1 patient (2.5%) had a minimal cystocele, 16 (40.0%) had moderate cystoceles, and 23 (57.5%) had large cystoceles. After the CaPS, 36 (72%) were completely dry, 3 (6%) had persistent SUI, 1 (2%) had de novo urinary incontinence (UI), 3 (6%) had persistent UI, and 1 (2%) had mixed incontinence. No patient had permanent urinary retention. Transvaginal placement of cadaveric fascia for concomitant sling and cystocele repair provides material of excellent strength for the repair without relying on the inherently weak tissues in the patient with pelvic prolapse. Thus far, the early results with CaPS are extremely encouraging. Long-term follow-up is underway to evaluate the efficacy of this procedure. PMID- 11114558 TI - Use of bone anchors in female urology. AB - Stress urinary incontinence remains one of the most prevalent conditions encountered by urologists. In many cases, surgical correction of this condition is carried out using a pubovaginal sling procedure. Bone anchors were initially used in transvaginal needle suspension procedures to improve stabilization of the bladder neck. This technology has been extended to sling procedures, allowing completion of these procedures by an entirely transvaginal approach. Early results of these procedures are encouraging, and overall morbidity appears much less when compared with conventional pubovaginal sling procedures. In this article, the application of bone anchors in female urology is reviewed. Techniques of pubovaginal sling and abdominal sacrocolpopexy using bone anchors and potential complications of bone anchor implantation are discussed. Surgeons performing procedures for the treatment of stress incontinence should be aware of the benefits and potential risks of bone anchor implantation. PMID- 11114559 TI - Transvaginal therapy of genuine stress incontinence. AB - Two minimally invasive techniques for treatment of genuine stress incontinence, a transvaginal retropubic urethropexy and a transvaginal sling, using Cooper's ligament as the anchoring structure, are reported along with the early results. These surgeries can be done easily in conjunction with vaginal reconstructive procedures. Twenty-seven women were operated on between October 1998 and September 1999. Seventeen women underwent the transvaginal retropubic urethropexy for genuine stress incontinence, whereas 10 women underwent the transvaginal sling for genuine stress incontinence with maximal urethral closure pressures less than 20 cm H(2)O. Postoperative urodynamics were done routinely at 4 months. Subjective follow-up was by routine postoperative visits or telephone survey. The mean follow-up of the retropubic urethropexy group was 6.5 months (range 1 to 12). Of these 17 women, 16 (94%) had no stress incontinence. Seven of 14 patients (50%) resolved their urge incontinence also. The mean follow-up of the transvaginal sling group was 2.8 months (range 1 to 4.5). Of the 10 patients in this group, 7 (70%) were cured of their stress incontinence. One patient in each group underwent only the anti-incontinence procedure without concomitant vaginal reconstructive procedures; both of these women were dry. Additional follow-up is required to see if these minimally invasive techniques produce successful results in the long term. PMID- 11114560 TI - The tension-free vaginal tape procedure for the treatment of stress incontinence in the female patient. AB - The newest development in the treatment of female stress incontinence is the tension-free vaginal tape (TVT) procedure. This procedure was first described by Ulmsten et al. in 1996 and involves recreating suburethral support with a polypropylene mesh, without repositioning the bladder or urethra. In their initial study, Ulmsten et al. reported an 84% cure rate at 2-year follow-up. The purpose of this report is to discuss the evolution and technical aspects of the TVT procedure and to outline the pre- and postoperative care recommended. A brief summary of our initial experience in our first 100 patients is included. PMID- 11114561 TI - Injectable agents in the treatment of stress urinary incontinence in women: where are we now? AB - Periurethral bulking agents have been used for decades. The only currently available agents (in the United States) include glutaraldehyde cross-linked collagen, autologous fat, and carbon bead technology. Initial subjective cure rates with collagen are acceptable, but with the majority of women requiring reinjection. The risk of allergic phenomena complicates collagen use. Autologous fat injection is initially effective in >50% of women, but resorption and fibrous replacement hamper the stability of the transplanted graft. Polytetrafluoroethylene and silicone are not currently approved by the US Food and Drug Administration because of particle migration. Materials in development include biologic agents such as allogeneic human collagen and autologous cartilage. Developmental synthetic agents include microballoon technology, hyaluronic acid with or without microsphere technology, hydroxylapatite, and a variety of polymeric technologies. Patient selection and material characteristics influence the optimal choice for injectable agent. PMID- 11114563 TI - The levator myorraphy repair for vaginal vault prolapse. AB - Various techniques have been described for the repair of vaginal vault prolapse after hysterectomy. Because of inherent difficulties associated with the sacrospinous fixation, a new repair, the levator myorraphy, was devised to simplify reconstruction of the pelvic floor 10 years ago. This repair results in a restored vaginal axis and prevention of posterior enterocele recurrence by recreating the levator shelf high within the peritoneal cavity and fixing the vault in that position. Because this procedure can be performed from a vaginal approach, morbidity is minimized. Over this 10-year time period, we have performed the levator myorraphy in over 120 women. We believe this approach can be easily taught, is highly effective, and results in a repair that most closely restores the normal anatomic position. PMID- 11114562 TI - Advancements in pharmacologic management of the overactive bladder. AB - Continued developments in the understanding of lower urinary tract function have led to improvements in the pharmacologic manipulation of bladder dysfunction. Drug delivery changes have produced drugs that provide better efficacy and tolerability, thus improving patient compliance. Improvements in drug delivery systems have altered drug bioavailability and pharmacokinetics. Active current investigation in new agents and delivery systems for intravesical delivery has yielded intriguing early results that may substantially add to the armamentarium for the management of the overactive bladder (urgency, frequency, urge incontinence). New developments in the understanding of the neuropharmacology of the bladder, peripheral pelvic nerves, and sacral cord may provide agents with entirely new drug effects, either as primary agents or agents to be used in combination with currently available drugs. We herein review newer agents and drug delivery systems. PMID- 11114564 TI - Abdominal sacral colpopexy and abdominal enterocele repair in the management of vaginal vault prolapse. AB - Vaginal vault prolapse and enterocele represent challenging forms of female pelvic organ relaxation. These conditions are most commonly associated with other pelvic organ defects. Proper diagnosis and management is essential to achieve long-term successful outcomes. Physical examination should be carried out in the lithotomy and standing positions (if necessary) in order to detect a loss of vaginal vault support. With proper identification of the vaginal cuff, one should assess the degree of mobility of the vaginal cuff with a Valsalva maneuver. If there is significant descent of the vaginal cuff, vaginal vault prolapse is present, and correction should be considered. The abdominal sacral colpopexy is an excellent means to provide vaginal vault suspension. This procedure entails suspension of the vaginal cuff to the sacrum with fascia or synthetic mesh. This procedure should always be accompanied by an abdominal enterocele repair and cul de-sac obliteration. In addition, many patients require surgical procedures to correct stress urinary incontinence, which is either symptomatic or latent (occurs postoperatively after prolapse correction). Complications include: mesh infection, mesh erosion, bowel obstruction, ileus, and bleeding from the presacral venous complex. If the procedure is carried out using meticulous technique, few complications occur and excellent long-term reduction of vaginal vault prolapse and enterocele are achieved. The purpose of this article is to review the preoperative evaluation of women with pelvic organ prolapse, and provide a detailed description of the surgical technique of an abdominal sacral colpopexy. PMID- 11114565 TI - Laparoscopic paravaginal repair plus burch colposuspension: review and descriptive technique. AB - The objective of this article was to review the available literature on laparoscopic Burch urethropexy cure rates and describe the authors' laparoscopic technique and experience with Burch urethropexy and paravaginal repair. A MEDLINE search (1991 to 1999) was performed for articles describing the laparoscopic Burch urethropexy using suture to elevate and stabilize the paraurethral tissue. Also a retrospective chart review of the authors' 171 consecutive patients between January 1997 and December 1999 was done. The laparoscopic Burch urethropexy and paravaginal repair is described using an open laparoscopic technique with 3 accessory ports for access. A transperitoneal approach is taken to gain access to the space of Retzius. The anterior vaginal wall and its paravaginal defects, if present, are identified. Nonabsorbable sutures are placed in a conventional fashion. The paravaginal repair is used for support of the anterior vaginal wall proximal to the urethral vesical junction and the Burch urethropexy distal to the vesical neck. An average of 6 sutures are used for the paravaginal repair and 4 sutures for the Burch urethropexy. Cystoscopy is performed to ensure no breech of lower urinary tract integrity. In all, 20 articles describing a laparoscopic Burch urethropexy and postoperative cure rate were identified. Cure rates ranged from 69% to 100%. A review of our experience revealed 130 of 171 patients had a Burch urethropexy and paravaginal repair, 23 of 171 patients a Burch urethropexy alone, and 18 of 171 patients a paravaginal repair alone. Of the authors' 171 patients, 4 (2.3%) had injury to the lower urinary tract during laparoscopic Burch urethropexy or paravaginal repair. All 4 injuries were cystotomies, 2 in patients with previous open retropubic urethropexies. No ureteral ligations or intravesical placement of suture was diagnosed. Other surgical parameters for the laparoscopic Burch uethropexy and paravaginal repair include an estimated blood loss of 50 mL, average hospital stay of less than 23 hours, and an average operative time of 70 minutes. All patients had their surgery completed via laparoscopy. The literature review and our personal experience suggests that the laparoscopic Burch urethropexy and paravaginal repair are safe and effective alternatives to traditional laparotomy for the treatment of genuine anatomic stress urine incontinence and cystourethrocele resulting from lateral vaginal wall defects. PMID- 11114566 TI - Anterior approach bilateral sacrospinous ligament fixation for vaginal vault prolapse. AB - The sacrospinous ligament fixation (SSLF) was first described as a unilateral fixation; however, bilateral fixation, when possible, allows a symmetrical vaginal reconstruction and provides additional vaginal vault support. We evaluated the outcome of treating total vault prolapse using a bilateral SSLF through an anterior vaginal approach. From July 1996 to July 1999, 28 patients (mean age 67) underwent bilateral SSLF procedures through an anterior vaginal approach. All patients had either grade 3 or 4 vault prolapse, and all patients had associated enteroceles, cystoceles, and rectoceles. All patients underwent fluorourodynamic evaluation including an abdominal leak point pressure (ALPP) with reduction of the vaginal prolapse. A pubovaginal sling was performed in 25 patients and all 28 patients underwent an anterior colporrhaphy, rectocele, and enterocele repair. A vaginal paravaginal repair was performed in 22 cases. At a mean follow-up of 17 months (range 5 to 35), 27 of 28 patients were cured, 1 patient had an asymptomatic unilateral grade 1 vault prolapse, 2 patients had developed small asymptomatic cystoceles and there had been no recurrence of rectoceles or enteroceles. Stress incontinence had been cured in all patients; however, 2 patients continued to have mild urge incontinence requiring <1 pad per day. Two patients complained of transient gluteal pain. We believe the anterior approach bilateral SSLF is a safe procedure with excellent medium term results in women with grade 3 to 4 vaginal prolapse. PMID- 11114567 TI - Recent advances in the management of the neurogenic bladder. AB - Proper evaluation of the neurogenic bladder remains the cornerstone for accurate management of the neurologically impaired patient. Due to the inherent progressive nature of many neurologic disorders causing bladder dysfunction and lack of targeted medical therapy, much work has been done and needs to be done to advance the management of this often-difficult patient population. This article reviews the latest advances in managing the neurogenic bladder. For ease of review, the neurogenic bladder can be divided into 2 basic categories: first, bladders that fail to empty successfully and, second, those that fail to store urine adequately. Therapy should be based on these categories because poor therapeutic results are seen when a standard treatment is prescribed for the wrong bladder condition. Given the success of other specialties (physical medicine and rehabilitation, orthopedics, and neurology) at improving and prolonging the lives of the neurologically impaired patient, the urologist has an increasing responsibility to evaluate and treat the neurogenic bladder effectively over a life span that is approaching that of the normal population. PMID- 11114568 TI - Update on extracorporeal magnetic innervation (EXMI) therapy for stress urinary incontinence. AB - Pulsed magnetic technology has been developed for pelvic floor muscle strengthening for the treatment of urinary incontinence. This report includes an update of the prospective multicenter study of extracorporeal magnetic innervation (ExMI) therapy for stress incontinence and a discussion of the possible mechanisms of action. Issues of patient selection for ExMI therapy will also be discussed. One hundred and eleven women with demonstrable stress urinary incontinence were studied. The mean age was 55 +/- 13 years, and the mean duration of symptoms was 11 years. Ninety-seven completed ExMI therapy and analysis. Evaluation before treatment included bladder diaries, dynamic pad weight test, urodynamics, and a quality-of-life survey. For treatment the patients were seated fully clothed in a Neocontrol chair with a magnetic field therapy head in the seat. Treatment sessions were for 20 minutes, twice a week, for 6 weeks. After ExMI therapy, all of the measures were repeated at 8 weeks, including the dynamic pad weight testing and quality-of-life survey. At 6 months, further data were added, including repeat bladder diary, pad use, and quality-of life survey. Forty-seven women completed 6 months of follow-up; of the 47, 13 patients were completely dry (28%) and 25 used no pad or less than 1 pad per day (53%). Pad use was reduced in 33 patients (70%). The median number of pads was reduced from 2.16 to 1 per day (Wilcoxon signed rank test, P <0.005). The frequency of leak episodes was reduced from 3.0 to 1.7 at 6 months (Wilcoxon signed rank test, P = 0.004). Detrusor instability was demonstrated in 10 before and 6 after ExMI (P <0.05). ExMI offers an alternative approach for the treatment of urinary incontinence. ExMI therapy is effective for both stress and urge incontinence. The best results are achieved in those patients who use no more than 3 pads a day and have had no prior continence surgery. PMID- 11114569 TI - Long-term results of a multicenter study on sacral nerve stimulation for treatment of urinary urge incontinence, urgency-frequency, and retention. AB - Many patients have chronic, debilitating symptoms of voiding dysfunction that are refractory to conventional medical or surgical therapies. This multicenter, prospective study evaluated the long-term effectiveness of sacral nerve stimulation using the implantable Medtronic InterStim therapy for urinary control in patients with otherwise intractable complaints of urinary urge incontinence, urgency-frequency, or retention. Each patient first underwent temporary, percutaneous sacral nerve test stimulation. If at least a 50% reduction in target symptoms was documented for at least 3 days, patients received a permanent Medtronic InterStim sacral nerve stimulation system that includes a surgically implanted lead and neurostimulator. Regular follow-up was conducted with outcome data. We report here on patients who have been observed from 1.5 to 3 years postimplantation. The results demonstrate that after 3 years, 59% of 41 urinary urge incontinent patients showed greater than 50% reduction in leaking episodes per day with 46% of patients being completely dry. After 2 years, 56% of the urgency-frequency patients showed greater than 50% reduction in voids per day. After 1. 5 years, 70% of 42 retention patients showed greater than 50% reduction in catheter volume per catheterization. We conclude that the Medtronic InterStim therapy for urinary control system is an effective therapy with sustained clinical benefit for patients with intractable symptoms of urinary urge incontinence, urgency-frequency, or retention. PMID- 11114570 TI - Diffusion kinetics in blood during haemodialysis and in vivo clearance of inorganic phosphate. AB - Contradictory data are reported in the literature concerning the diffusion kinetics of inorganic phosphates (iPh) between red blood cells and plasma during haemodialysis. Accordingly, we performed mass balance and equilibration studies to analyze the diffusion kinetics of iPh in vivo and in vitro. Mass balance analysis shows that iPh is only cleared from the plasma volume and thus that it practically does not diffuse from red blood cells to plasma during the short time lapse of blood transit through the haemodialyzer. In vitro equilibration studies of blood drawn at the filter outlet show that at room temperature there is a slow, limited, and almost linear net efflux of iPh during the 4 h that follow blood drawing. Our results point out: (1) that the in vivo clearance of iPh should be exclusively determined as plasma clearance, and (2) that for accurate clearance determinations the iPh concentrations should be measured in blood samples centrifuged within at most 1 h after blood drawing. Whole-blood clearance determinations--as well as the in vitro dialyzer data--largely overestimate (>30%) the real in vivo dialyzer performance. PMID- 11114571 TI - Fitness of biocompatible high-flux hemodiafiltration for dialysis-related amyloidosis. AB - AIMS: In order to assess the long-term influence of high-flux hemodiafiltration (HDF) on hemodynamic stability in patients with dialysis-related amyloidosis. METHODS: HDF with high-flux dialyzers was performed for a year in 11 patients who had undergone the surgery for carpal tunnel syndrome. Patients were divided into two groups, and for each group synthetic or cellulose membrane dialyzers were applied. RESULTS: A year after the exchange from standard hemodialysis to HDF with cellulose high-flux dialyzers, beta(2)-microglobulin was decreased (45+/-3 to 28+/-2 mg/l, n = 5, p< 0.05), but plasma non-refilling ratio was not altered. However, the continuance of synthetic high-flux HDF for a year decreased plasma non-refilling ratio (17+/-5 to 7+/-4%, p<0.05), hypotensive episodes (4.3+/-0.6 to 2.5+/-0.4 sessions/month, p<0.05) and muscle cramps (3.8+/-0.5 to 1.8+/-0.5 sessions/month, p<0.01) in addition to the decrements in serum beta(2) microglobulin (45+/-5 to 33+/-2 mg/l, n = 6, p<0.01). CONCLUSIONS: Although the study on a large number of patients may be required to draw a final conclusion, our present data suggest that biocompatible high-flux HDF is uniquely suited for dialysis-related amyloidosis. PMID- 11114572 TI - Validation of a simple method for assessing sodium intake in dialysis patients. AB - It has been reported that sodium intake can be estimated in dialysis patients by the increment in the body sodium pool from the end of a dialysis session to the beginning of the following one. To verify the reliability of this method we compared the sodium intake, estimated by the interdialytic changes in plasma sodium concentration (C) and body water volume (V), to sodium removal during three consecutive sessions. For this purpose we investigated 9 nondiabetic patients, 5 females and 4 males, under chronic hemofiltration treatment. Sodium intake was estimated by the formula (C(pre) V(pre)) - (C(post) V(post)) using a flame photometer and electrical bioimpedance to determine the plasma sodium concentration and total body water, respectively. Sodium removal was calculated by the difference between sodium loss into the ultrafiltrate and sodium gain with the reinfusion fluid. The mean values of sodium intake calculated during the three intervals corresponded with the sodium losses measured during the three hemofiltration sessions in each patient (338+/-55 vs. 329+/-67 mEq; r = 0.92, p<0.0001). A direct relationship was also observed between sodium intake and both interdialytic body weight increase (r = 0.89, p< 0.0001) and fluid loss during the sessions (r = 0.88, p<0.0001). This data demonstrates that sodium intake can be properly estimated by the interdialytic changes in body water and plasma sodium concentrations. They also suggest that fluid intake may be influenced by sodium consumption and that sodium intake monitoring could be useful for the control of excessive interdialytic fluid gain. PMID- 11114573 TI - Endocrine-disrupting chemicals in CAPD dialysate and effluent. AB - Environmental contamination by endocrine-disrupting chemicals (EDC) has been a major focus of recent research and policy discussions. EDC-suspected man-made chemicals used as raw materials or plasticizers have been shown to elute from plastic products. To examine whether the dialysate for continuous ambulatory peritoneal dialysis (CAPD) is contaminated with EDC, we determined bisphenol A (BPA), nonylphenol (NP), di-(2-ethylhexyl)phthalate (DEHP) and di-n-butyl phthalate (DBP) in the pre-used dialysate and in the peritoneal effluent from renal failure patients by gas chromatography/mass spectrometry. Concentrations of BPA, NP, DEHP and DBP were 0.02-0.23 ppb (microg/l), 0.09-0.22, 1.1-3.7, and <0.1 2.1 ppb, respectively, in the pre-used dialysate, and <0.01-0.07, <0.1-0.45, 0.35 1.23, and 0.42-1.76 ppb, respectively, in the effluent, from which the maximal daily contamination of BPA and NP by CAPD was estimated at the microgram level and that of phthalate esters at the 10-microg level. These concentrations are far less than the toxic dosage reported so far, so that CAPD is unlikely to contaminate patients seriously. PMID- 11114574 TI - Effects of L-carnitine supplementation on renal anemia in poor responders to erythropoietin. AB - While renal anemia can be successfully treated by use of erythropoietin (EPO) in most hemodialysis (HD) patients, some patients have anemia that is refractory to treatment with a high dose of EPO. We examined whether L-carnitine treatment could raise hematocrit (Hct) levels in such patients. Fourteen HD patients who showed a poor response to EPO and no evident factors which inhibit a response to EPO were selected to receive oral L-carnitine (500 mg/day) in a 3-month trial. During the study, 36% of the patients showed Hct increases of more than 2%. Statistical analysis revealed significant increases of Hct (p = 0.003) and total iron-binding capacity (TIBC) (p = 0.050) and a significant decrease of ferritin (p = 0.005). In addition, we found that red blood cells (RBCs) in HD patients contained a comparable level of carnitine to normal controls, despite the presence of serum carnitine deficiency, and that RBC carnitine was not removed through HD, in contrast to serum carnitine. These results suggest that RBC carnitine may be essential for RBCs to perform their metabolic function in renal anemia and that oral L-carnitine treatment could improve anemia in poor responders to EPO. PMID- 11114575 TI - Increase in blood volume during dialysis without ultrafiltration. AB - Combined dialysis and ultrafiltration leads to more frequent episodes of hypotension than isolated ultrafiltration. It has been suggested that decreased plasma volume preservation could be responsible for this phenomenon. The present study evaluates the effects of diffusive dialysis on the changes in relative blood volume (RBV). Six stable hemodialysis patients, without the need of ultrafiltration, were studied during 10 sessions of diffusive dialysis (bicarbonate) lasting 4 h. RBV was monitored continuously by measurement of hematocrit. During the 1st and 2nd h RBV increased by 2.4+/-1.4 and 2.5+/-0.8% respectively, returning to baseline levels at the end of dialysis. No changes in blood pressure or heart rate were noted. We conclude that during diffusive dialysis without ultrafiltration RBV is increased. A decrease in vascular resistance, or changes in regional blood distribution could explain these findings. PMID- 11114576 TI - Successful use of central venous catheter as permanent hemodialysis access: 84 month follow-Up in lucania. AB - Cuffed tunneled venous access catheters are commonly used for temporary and permanent access in hemodialysis (HD) patients. These catheters serve an essential role in providing permanent access in subjects in whom all other access options have been exhausted. The predominant complications are catheter thrombosis, catheter fibrin sheating and infection. The aim of this study was to evaluate long-term survival and complications of permanent venous catheters (PVC) placed for the purpose of HD during the period from January 1992 to December 1998, at the Dialysis Units of Lucania (a southern Italian region). A total of 98 PVC were placed in 88 patients during this period. The catheters used were of three types: (a) 72 VasCath Soft Cell catheters (Bard Instrument Company, Toronto, Ont., Canada); (b) 22 PermCath catheters (Quinton Instrument Company, Seattle, Wash., USA), and (c) 4 Tesio catheters (Bellco SpA, Mirandola, Italy). Survival curves of catheters were calculated using the Kaplan-Meier product-limit estimator. The patient survival was 60% at the 78th month. Actually, 52 patients (27 males, 25 females) are still alive: 15 (26.9%) of these patients have diabetes mellitus and 1 has been transplanted. The actuarial survival rate of PVC was 89% in the whole population studied and 82% in subjects alive after 84 months. Twenty-five patients (28.4%) had PVC as the first reliable vascular access. Long-term complications occurred 27 times (1 episode every 44.81 month/patient) as: breakage (3.1%); thrombosis (10.2%); displacement (2.0%); subcutaneous tunnel bleeding (3.1%); inadequate blood flow (7.1%), and infection (10.2%). In conclusion, our data confirm that PVC might represent an effective long-term blood access route for HD. Again, PVC are getting the access of choice for selected patients (i.e., older subjects with cardiovascular diseases and cancer patients) and are enjoying a dramatic increase in use for subjects who are terrified of repetitive venopuncture. PMID- 11114577 TI - Resource limitations and strategies for the treatment of uremia. A dialysis unit in the Himalayan foothills. AB - India has made great strides in health care since gaining independence in 1947. Much still needs to be done. Scarce health care resources are directed at priorities that include infant and maternal mortality, immunizations, malnutrition, communicable disease prevention, and access to protected water and sanitation. There is, therefore, no government reimbursement for treatment of patients with ESRD (estimated incidence of 100 per million population). Transplantation, the modality of choice in India, benefits only 2-3% of ESRD patients. Hemodialysis is primarily used short-term for pretransplant stabilization. A very small minority of patients is placed on maintenance hemodialysis or CAPD. The annual cost of renal replacement therapies is more than 10 times the per capita gross national product. Financial constraints pose ethical problems for the nephrologist related to adequate prescriptions and compliance. Preventing the progression of kidney disease and reducing the cost of disposables through indigenous manufacture are initiatives that need to be explored. PMID- 11114578 TI - Inflammatory and atherosclerotic interactions in the depleted uremic patient. AB - Despite the improvements in dialysis technology, the cardiovascular mortality rate is still unacceptably high among dialysis patients. It is obvious that traditional risk factors, such as hypertension, chronic heart failure (CHF), dyslipidemia and diabetes mellitus, may account for a large part of the increased cardiovascular mortality rate in these patients. However, based on recent research it could be speculated that other, non-traditional risk factors might also contribute to the high cardiovascular mortality rate in dialysis patients. Chronic inflammation, as evidenced by increased levels of pro-inflammatory cytokines and C-reactive protein (CRP), is a common feature in dialysis patients and is associated with an increased cardiovascular morbidity and mortality. Indeed, elevated levels of pro-inflammatory cytokines (such as TNF-alpha, IL-1 and IL-6) may cause malnutrition and progressive atherosclerotic cardiovascular disease by several pathogenetic mechanisms, which will be discussed in this review. Based on the strong associations observed between malnutrition, inflammation and atherosclerosis in patients with chronic renal failure (CRF) we have proposed that these features constitute a specific syndrome (MIA), which carries a high mortality rate. As elevated levels of pro-inflammatory cytokines may play a central part in the vicious circle of malnutrition, inflammation and atherosclerosis, further research is needed to investigate whether or not different anti-cytokine treatment strategies may improve survival in dialysis patients. PMID- 11114579 TI - An in vitro bacterial touch contamination risk assessment of two CAPD twinbag systems. AB - The objective of this study was to compare, using in vitro quantitative microbiology, the ability of two commercially available peritoneal dialysis solution delivery systems to prevent and remove, via convective fluid flow, intralumenal fluid path bacterial contamination. The two systems (A and B) differed in both the configuration of their flow control, or Y-junction and the method of fluid flow control and also in the design of their Luer tubing connectors. System A had a tubing type Y-junction that requires clamps to control fluid flow and uses a connector with a male Luer that is deeply recessed within a shroud. System B has a dial-type rigid Y-junction with in-line flow control and a connector with a male Luer that is shrouded but not recessed. System A connectors allowed significantly (p<0.0001) fewer bacteria to be transferred into the fluid path than System B after simulated touch contamination. Also, when an equivalent number of bacteria were deliberately placed into the fluid paths of both systems, System A was more effective in removal of the bacterial contamination by convective fluid flow than System B (p<0.0001), resulting in fewer organisms infused into the simulated peritoneum. Specific design features of System A, such as a recessed male Luer, and a Y-junction fluid flow path with low turbulence were likely explanations for its superior results. This study emphasizes the importance of connector and fluid path flow design in the aseptic performance of peritoneal dialysis delivery systems. PMID- 11114580 TI - Variations in hematocrit induced by hemodialysis. PMID- 11114581 TI - The role of dialyzer biocompatibility in acute renal failure. PMID- 11114582 TI - Reply PMID- 11114583 TI - Towards defining the role of glycans as hardware in information storage and transfer: basic principles, experimental approaches and recent progress. AB - The term 'code' in biological information transfer appears to be tightly and hitherto exclusively connected with the genetic code based on nucleotides and translated into functional activities via proteins. However, the recent appreciation of the enormous coding capacity of oligosaccharide chains of natural glycoconjugates has spurred to give heed to a new concept: versatile glycan assembly by the genetically encoded glycosyltransferases endows cells with a probably not yet fully catalogued array of meaningful messages. Enciphered by sugar receptors such as endogenous lectins the information of code words established by a series of covalently linked monosaccharides as letters for example guides correct intra- and intercellular routing of glycoproteins, modulates cell proliferation or migration and mediates cell adhesion. Evidently, the elucidation of the structural frameworks and the recognition strategies within the operation of the sugar code poses a fascinating conundrum. The far reaching impact of this recognition mode on the level of cells, tissues and organs has fueled vigorous investigations to probe the subtleties of protein carbohydrate interactions. This review presents information on the necessarily concerted approach using X-ray crystallography, molecular modeling, nuclear magnetic resonance spectroscopy, thermodynamic analysis and engineered ligands and receptors. This part of the treatise is flanked by exemplarily chosen insights made possible by these techniques. PMID- 11114584 TI - Biosynthesis and expression of zona pellucida glycoproteins in mammals. AB - The zona pellucida (ZP) is an extracellular matrix surrounding the oocyte and the early embryo that exerts several important functions during fertilization and early embryonic development. The ZP of most mammalian species is composed of three glycoproteins (ZPA, ZPB, ZPC), products of the gene families ZPA, ZPB and ZPC that have been found to be highly homologous within mammalian species. Most data on the structure and function of the ZP are obtained from studies in mouse. New data from pig and other domestic animals, however, indicate that the mouse model does not hold for all other species. Whereas in the mouse ZPB is the primary sperm receptor, in the pig ZPA has been shown to possess receptor activity. Contrary to the mouse, where the growing oocyte is the only source of zona glycoproteins, in domestic animals these proteins are expressed in both the oocyte and granulosa cells in a stage-specific pattern and may play also a role in granulosa cell differentiation. In several mammalian species, the epithelial secretory cells of the oviduct synthesize and secrete specific glycoproteins (oviductins) that become closely associated with the ZP of the ovulated oocyte. Once bound to the ZP, oviductin molecules could act as a protective layer around the oocyte and early embryo by virtue of their densely glycosylated mucin-type domains. PMID- 11114585 TI - Towards the molecular basis of sperm and egg interaction during mammalian fertilization. AB - During the past 2 decades, a number of genes have been cloned from mammals which encode polypeptides that participate in the process of fertilization. Among these are glycoproteins ZP1-3 that constitute the zona pellucida of eggs from mice to human beings. In mice, one of these glycoproteins, mZP3, acts as a primary sperm receptor and acrosome reaction-inducer. The evidence suggests that acrosome intact sperm recognize and bind to a specific class of mZP3 oligosaccharides present on two serine residues (O-linked) located near the carboxy-terminus of the polypeptide. Mutagenesis of either of these residues results in the synthesis of an inactive form of the receptor. Therefore, mammalian fertilization is a carbohydrate-mediated event. It is possible that changes in the structure of these oligosaccharides (e.g., composition, sequence, linkages, modifications, etc.) could account for species-specific binding of sperm to eggs. Stably transfected somatic cells, null mutant animals, and DNA constructs are now available to test this possibility both in vivo and in vitro. PMID- 11114586 TI - Galactosyltransferase function during mammalian fertilization. AB - Gamete recognition has been studied extensively in the mouse. In this system, it is generally believed that sperm bind to a class of O-linked oligosaccharides on the zona pellucida glycoprotein, ZP3. The best characterized sperm receptor for ZP3 is beta1, 4-galactosyltransferase (GalT), which functions in a lectin-like capacity by binding to N-terminal N-acetylglucosamine residues on ZP3 oligosaccharides. Multivalent oligosaccharides on ZP3, as well as synthetic polymers terminating in N-acetylglucosamine aggregate GalT, leading to activation of a heterotrimeric G protein cascade and culminating in the acrosome reaction. Following fertilization, cortical granules release N-acetylglucosaminidase, which removes the binding site for sperm GalT and facilitates the zona block to polyspermic binding. Genetic manipulation of GalT expression has confirmed its function as a ZP3 receptor. Overexpressing GalT on sperm leads to increased binding of ZP3, increased G protein activation, and precocious acrosome reactions. In contrast, sperm from mice made null for GalT by homologous recombination are refractory to ZP3, in that they are unable to bind soluble ZP3 and fail to undergo the acrosome reaction in response to zona glycoproteins. Surprisingly, GalT null sperm still bind to the zona and achieve low rates of fertilization in vitro. This then suggests that sperm-egg binding involves receptor-ligand interactions independent of GalT and ZP3. The current model suggests that GalT functions as the ZP3 receptor that is responsible for inducing the acrosome reaction, whereas initial sperm-zona binding is dictated by other sperm surface receptors. Consistent with this, at least three other zona pellucida monosaccharides have been implicated in sperm binding, and novel sperm surface glycoproteins have been suggested to function in gamete binding. A large scaffolding protein has been identified that associates with the GalT cytoplasmic domain and may be responsible for orchestrating its signal transduction capacities that lead to the acrosome reaction. PMID- 11114587 TI - Molecular basis of human sperm-zona pellucida interaction. AB - The recognition of carbohydrate sequences by complimentary receptors has been shown to be a critical factor in gamete interaction in many different animal species. We have proposed the hypothesis that, in the human, sperm binding to the zona pellucida requires a 'selectin-like' interaction. We have used the hemizona assay (a unique internally controlled bioassay that evaluates tight binding of human sperm to the homologous zona pellucida) and advanced methods of carbohydrate analysis to test this hypothesis. We provide compelling evidence that demonstrates that oligosaccharide recognition is also required for specific, tight human gamete binding. Hapten inhibition tests, zona pellucida lectin binding studies and removal/modification of functional carbohydrates by chemical and enzymatic methods provide evidence for the involvement of defined carbohydrate moieties in initial binding. Our studies suggest the existence of distinct zona-binding proteins on human sperm that can bind to selectin ligands. Additionally, results suggest a possible convergence in the types of carbohydrate sequences recognized during initial human gamete binding and immune/inflammatory cell interactions. Full characterization of the glycoconjugates that manifest selectin-ligand activity on the human zona pellucida will allow for a better understanding of human gamete interaction in physiological and pathological situations. PMID- 11114588 TI - Localization of sperm ligand carbohydrate chains in pig zona pellucida glycoproteins. AB - Neutral N-linked carbohydrate chains from pig ZPB/ZPC mixture are shown to possess sperm ligand activity. Of these complex-type chains, triantennary/tetraantennary chains exhibit the activity stronger than that of diantennary chains. Intact ZPB and ZPC cannot be separated from each other unless acidic N-acetyllactosamine regions of their carbohydrate chains are removed by endo-beta-galactosidase digestion. The endo-beta-galactosidase-digested ZPB and its N-terminal fragment of 111 residues retain the sperm ligand activity. Three glycopeptides, having one Asn residue to which the carbohydrate chain is linked, are obtained by lysylendopeptidase digestion of the heat-solubilized zonae containing intact ZPB, and lysylendopeptidase and chymotryptic digestions of endo beta-galactosidase-digested ZPB. On the basis of sugar mapping analysis of the N linked chains from these glycopeptides and comparison with the carbohydrate structures of the main intact neutral N-linked chains of ZPB/ZPC, the triantennary and tetraantennary chains are shown to be localized mainly at Asn220 of ZPB, whereas diantennary chains are present on all the three N-glycosylation sites (Asn203, Asn220 and Asn333). These results suggest that the carbohydrate chains linked to Asn220 of ZPB participate predominantly in sperm-egg binding. ZPC has been shown to support the expression of sperm ligand activity of ZPB. Three glycopeptides, each having one of the N-glycosylation sites, are obtained by tryptic digestion of endo-beta-galactosidase-digested ZPC. Triantennary and tetraantennary chains are found mainly at Asn271 of ZPC, whereas diantennary chains are present at all three N-glycosylation sites (Asn124, Asn146 and Asn271). Thus, the localization of triantennary and tetraantennary chains in ZPC is different from that in ZPB. PMID- 11114589 TI - Carbohydrate-mediated sperm-egg interaction and species specificity: a clue from the Unio elongatulus model. AB - As a first step fertilization requires that sperm bind to the extracellular coat of the egg. It is generally accepted that binding is mediated by complex carbohydrates on the egg coat, recognized by carbohydrate-binding proteins on the sperm surface. In the mollusc bivalve Unio elongatulus, the main functional epitope of the carbohydrate ligand has been determined, and it shares several characteristics with other sugar structures involved in the fertilization process in different animal models. A polyclonal antibody against the Unio epitope reacts with the human ZP3 glycoprotein and the Unio ligand binds human spermatozoa in an in vitro assay. These findings are discussed in the light of the species specificity of sperm-egg binding at fertilization. PMID- 11114590 TI - Sperm adhesion molecules: structure and function. AB - Fertilisation is a unique event in which the morphologically disparate gametes recognise, bind and fuse with each other. This event follows a highly regulated schedule of biochemical interactions, in which molecules are involved that mediate cell adhesion, signal transduction and the initiation of metabolic pathways. A plethora of molecules has been found on the male gamete and with regard to the different protein structures it is almost impossible to overlook the structures involved. Even more, carbohydrate structures cause an additional diversity with regard to the generation of surface structures. In this communication we try to elucidate the structures of proteins that have been known so far. We have focussed on spermadhesins, the zonadhesin, proacrosin and the PH 20 antigen. The variety of structures and also the common features among them as well as the presence of redundant systems are attributable to the evolutionary force of intraspecific sperm competition. This evolutionary force is assumed to be also responsible for the species selectivity observed in these adhesion molecules, which explains the preferential binding of gametes in a homologous system. PMID- 11114592 TI - Carbohydrate-mediated formation of the oviductal sperm reservoir in mammals. AB - Mammalian sperm are trapped in a reservoir in the oviduct until ovulation is imminent. Then, they are gradually released, such that a few meet the oocytes as they enter the ampulla of the oviduct. In the three eutherian species studied to date, sperm are trapped in the reservoir by carbohydrate-mediated binding to the oviductal mucosa. Evidence indicates that a molecule on the surface of the plasma membrane overlying the acrosome binds to a carbohydrate moiety on the surface of the oviduct. While sperm remain bound, they appear to be protected from degradation. When sperm become capacitated, they lose binding affinity for the oviductal mucosa. The mechanism initiating capacitation in the reservoir is unknown; however, it must be tied to the hormonal signalling of ovulation. Hyperactivated motility may assist sperm in pulling off from the mucosal surface as binding affinity declines. The function of the reservoir appears to be to prevent polyspermy and ensure fertilization by providing a small number of sperm in the proper physiological condition for fertilization at the time the oocytes enter the oviduct. PMID- 11114591 TI - New insights into the origin, structure and role of CD52: a major component of the mammalian sperm glycocalyx. AB - The sperm glycocalyx represents the primary interface between the male gamete and its environment, and gamete interaction inevitably involves interaction with this structure. Thus, it has potential significance as a target for antibodies that inhibit sperm function. Still, little is known about the components and biological role of the sperm glycocalyx. Despite the apparent complexity of the sperm membrane, surface carbohydrate labelling experiments show a high selectivity suggesting that carbohydrate side chains of CD52, an unusually short, bipolar glycopeptide of epididymal origin, form major components of the sperm glycocalyx in all mammalian species investigated. Acquisition of the highly sialylated, lipid-anchored CD52 antigen is one of the few well-defined modifications that occur to the sperm membrane during epididymal passage. It would explain changes in lectin-binding patterns and also the remarkable surface charge differences occurring during epididymal transit, most probably attributable to its terminal sialic acid residues. CD52 seems to be immunodominant on human spermatozoa, and antibodies directed against it can agglutinate and completely immobilize human sperm in the presence of complement. Expression of the same peptide backbone in lymphocytes had largely discounted its consideration as a candidate for contraceptive development. However, the recent proof of male-specific modifications indicates the feasibility of this approach. PMID- 11114593 TI - The species recognition system: a new corollary for the human fetoembryonic defense system hypothesis. AB - We have previously suggested that the human fetus is protected during human development by a system of both soluble and cell surface associated glycoconjugates that utilize their carbohydrate sequences as functional groups to enable them to evoke tolerance. The proposed model has been referred to as the human fetoembryonic defense system hypothesis (hu-FEDS). In this paradigm, it has previously been proposed that similar oligosaccharides are used to mediate crucial recognition events required during both human sperm-egg binding and immune-inflammatory cell interactions. This vertical integration suggested to us that the sperm-egg binding itself is related to universal recognition events that occur between immune and inflammatory cells, except that in this case recognition of 'species' rather than recognition of 'self' is being manifested. In this paper, we have designated this component of hu-FEDS as the species recognition system (SRS). We propose that the SRS is an integral component of the hu-FEDS used to enable sperm-egg recognition and protection of the gametes from potential immune responses. Recent structural data indicates that the glycan sequences implicated in mediating murine gamete recognition are also expressed on CD45 in activated murine T lymphocytes and cytotoxic T lymphocytes. This overlap supports our contention that there is an overlap between the immune and gamete recognition systems. Therefore the hu-FEDS paradigm may be a subset of a larger model that also applies to other placental mammals. We therefore propose that the hu-FEDS model for protection should in the future be referred to as the eutherian fetoembryonic defense system hypothesis (eu-FEDS) to account for this extension. The possibility exists that the SRS component of eu-FEDS could predate eutherians and extend to all sexually reproducing organisms. Future investigation of the interactions between the immune and gamete recognition system will be required to determine the degree of overlap. PMID- 11114594 TI - Radical prostatectomy: options and issues. AB - Radical prostatectomy is an effective treatment for patients with clinically localized prostate cancer and is associated with a very low level of mortality. However, many men with untreated clinically localized prostate cancer do not die from the disease and, following radical prostatectomy, some patients will suffer from a loss of potency and/or incontinence. A major challenge faced by the clinician is to identify the individual patient who will benefit from radical prostatectomy. In this review, we discuss the natural history of clinically localized prostate cancer and the factors likely to affect the treatment decision for an individual patient. Recent studies by other investigators and ourselves have revealed that the T1/T2 tumour is heterogeneous with respect to pathological stage and outcome, and that the quantity of Gleason grade 4/5 tumour is a significant prognostic factor predicting lymph node progression and capsular penetration. Classification and Regression Trees (CART) analysis including such preoperative parameters can be used to predict the probability of an individual patient having a pT2 tumour and, therefore, whether he could have a nerve-sparing radical prostatectomy - a procedure which offers better outcomes in terms of potency and continence. PMID- 11114595 TI - Neoadjuvant hormone therapy and radical prostatectomy: the jury is still out. AB - Radical prostatectomy may lead to cure as long as the cancer is confined to the prostate and all malignant cells are removed. However, clinical staging is inaccurate and a significant proportion of cT1-T2 patients have positive margins which increases the likelihood of disease progression within 5 years of surgery. Neoadjuvant hormone therapy is one option being used to increase the likelihood of prostate cancer cure after radical prostatectomy. Randomized clinical trials using neoadjuvant hormone therapy and radical prostatectomy have been conducted mainly in patients with cT1 and cT2 prostate cancer. A decrease in the number of positive surgical margins was found in cT1 and cT2 prostate cancer patients receiving neoadjuvant hormone therapy, with a further decrease in those receiving treatment over longer periods. In cT3 prostate cancer patients equivocal results have been obtained and further research is needed. None of the studies reported so far were able to define the impact of neoadjuvant treatment on the surgical management of locally advanced prostate cancer. Additional studies are required to determine the optimal type and duration of hormone treatment. Furthermore, long-term follow-up is needed to evaluate whether neoadjuvant therapy will improve overall survival. In the meantime, patients must be informed of the advantages and disadvantages of treatment to allow them to make informed treatment decisions. PMID- 11114596 TI - Advanced prostate cancer: immediate or deferred hormone therapy? AB - The timing of hormone therapy in patients with advanced prostate cancer remains controversial. Although evidence from the Veterans Administration Cooperative Urological Group (VACURG) data indicated the potential benefit of immediate hormone treatment in terms of time to progression and disease-specific survival, it also supported the possibility of deferred treatment. This review looks at the results of subsequent studies of immediate versus deferred hormone therapy including the Medical Research Council (MRC) study, the European Organisation for Research and Treatment of Cancer (EORTC) studies, the Radiation Therapy Oncology Group (RTOG) study and the Eastern Cooperative Oncology Group (ECOG) study. Data from these studies and the availability of newer, better tolerated hormone therapies may well mean that the survival and welfare of patients with advanced prostate cancer could be considerably better with immediate hormone therapy rather than deferred treatment. PMID- 11114597 TI - Treatment of locally advanced prostate cancer--a new role for antiandrogen monotherapy? AB - Men with locally advanced prostate cancer face a high risk of disease progression and cancer-related death. The traditional active treatment options for locally advanced disease, either following failure of treatment of primary curative intent or newly diagnosed, are radiotherapy and castration. Radiotherapy alone has a high failure rate, although outcome can be improved by adjuvant hormonal therapy. Castration is associated with loss of libido, sexual dysfunction, osteoporosis and hot flushes, which are significant drawbacks when patients may receive treatment for several years. Monotherapy with a non-steroidal antiandrogen offers potential benefits with respect to quality of life. Studies in the adjuvant setting are in progress. In the setting of previously untreated locally advanced disease, pooled mature data (56% deaths) from two major studies indicate no significant difference in survival outcome between bicalutamide ('Casodex') 150 mg and castration. Bicalutamide 150 mg offers quality of life benefits with respect to sexual interest and physical capacity. Preliminary data suggest that bicalutamide maintains bone mineral density. Bicalutamide 150 mg is well tolerated; gynaecomastia and breast pain, common side effects of antiandrogen monotherapy, may be managed by prophylactic irradiation or surgery. Bicalutamide 150 mg monotherapy is an alternative to castration for locally advanced prostate cancer. PMID- 11114598 TI - Matrix metalloproteinase 2 activation in cultured corneas. AB - PURPOSE: Corneas that are maintained in tissue culture medium shed their epithelial cells and repopulation following graft surgery is an essential facet of the healing process. Failure to do so may be a result of structural damage to the epithelial basement membrane of a donor cornea. The purpose of the present investigation was to ascertain whether MMP-2, the matrix metalloproteinase produced by corneal keratocytes, may be activated during storage and hence cleave the type IV collagen component of the epithelial cell basement membrane. METHODS: Fresh and transplant rejected corneas that had been stored in culture medium for varying time periods and of known donor age were collected. The soluble protein fractions of these corneas were obtained. Their MMP-2 proteins were visualised by zymography on SDS gelatin polyacrylamide gels and assayed for activity against nitrophenyl acetate and denatured [(3)H]type I collagen. RESULTS: The stromal tissue of fresh, normal corneas produced inactive MMP-2 of M(r) 66,000. Although the cultured corneas did not up-regulate MMP-2 production, they contained additional MMP-2 activities of M(r) 62,000 and M(r) 43,000. The appearance of these additional MMP-2 activities correlated with corneal culture time but not donor age. The ability to cleave denatured [(3)H]type I collagen correlated with the appearance of the M(r) 43,000 activity but not the M(r) 62,000 activity. CONCLUSION: Activated MMP-2 is produced in cultured corneas. For this reason the corneas donated for all graft procedures should not be held in culture medium for periods exceeding 4 weeks. PMID- 11114599 TI - Mineralocorticoid hormone signaling regulates the 'epithelial sodium channel' in fibroblasts from human cornea. AB - We investigated the regulation of sodium absorption by steroid hormones in embryologically diverse cells from the human eye. A cell extract from human corneal fibroblasts was positive for both the epithelial sodium channel (ENaC) and the mineralocorticoid receptor (MCR) as 82- to 85-kD and 102-kD bands, respectively, by the Western blot technique. In fluorescent, confocal and electron microscopy, the MCR was revealed as a nucleocytoplasmic protein, whereas the ENaC was almost exclusively membrane bound; both appeared aligned along actin filaments of corneal keratocytes, and both were widely colocalized in various cell types of human cornea in situ. Following reverse transcription and amplification of total RNA isolated from corneal fibroblasts, the ENaC and MCR genes in the PCR product were evident as predicted bands of 520 and 843 bp, respectively, whose sequence exhibited 100% identity with those from known human sources. The multiplication of corneal fibroblasts was influenced by both the MCR specific antagonist RU 26752 and the natural hormone aldosterone, and these steroids also stimulated protein phosphorylation. In quantitative PCR, both the basal and aldosterone-induced levels of ENaC were diminished by the MCR-specific antagonist ZK 91587. Consequently, the ocular sodium channel appears to be regulated by steroid signalling in cells of diverse embryological origins, contrary to the existing notions where (a) this process would be limited exclusively to the epithelial cells and (b) ocular sodium transport would be regulated via the Na(+)-K(+)-ATPase in the basolateral membrane. PMID- 11114601 TI - Antisense basic fibroblast growth factor oligonucleotide reduced adhesion of retinal pigment epithelial cells to extracellular matrix molecules and their proliferation. AB - We investigated the effect of extracellular matrix molecules on the adhesion of retinal pigment epithelial cells and their subsequent proliferation. Fibronectin, collagen type I and vitronectin enhanced their adhesion and proliferation. In addition, the effect of basic fibroblast growth factor (bFGF) on their adhesion and proliferation was studied. bFGF enhanced their adhesion and proliferation, whereas antisense bFGF reduced their adhesion to and their proliferation on extracellular matrix molecules. PMID- 11114600 TI - The effect of prinomastat (AG3340), a synthetic inhibitor of matrix metalloproteinases, on posttraumatic proliferative vitreoretinopathy. AB - In a search for a pharmacologic adjuvant in the management of posttraumatic proliferative vitreoretinopathy (PVR), we investigated the effect of intravitreal injection of prinomastat (AG3340) on an experimental model. Posterior penetrating eye trauma was created in one eye each of 24 New Zealand white rabbits. One week after the surgery, all rabbits were randomized (1:1) to receive 0.5 mg prinomastat or the vehicle of the drug intravitreally every week for 6 weeks. The degree of PVR for each hemiretina was scored, and the two scores were summed to obtain a total eye score. The mean total eye score was 3.58 in the treatment group and 5.75 in the control group (p = 0.0307). The numbers of eyes with tractional retinal detachment in the prinomastat-treated (n = 12) and control (n = 12) groups were 3 and 9, respectively (p = 0.0391). These results suggest that intravitreally administered prinomastat has an inhibitory effect on posttraumatic PVR. PMID- 11114602 TI - Effect of particle size of polymeric nanospheres on intravitreal kinetics. AB - In this study, we injected nanospheres containing a fluorescein derivative into the vitreous cavity of pigmented rabbit eyes and evaluated their intraocular kinetics as drug carriers in vivo. Polystyrene nanospheres (2 microm, 200 nm and 50 nm in diameter) containing a fluorescein derivative were used in this study. A suspension of each particle was prepared by diluting with distilled water at a concentration of 10 microg/ml equivalent to sodium fluorescein. The suspension of nanospheres was injected once into the vitreous cavity of unilateral eyes of pigmented rabbits. A sodium fluorescein solution of the same concentration was injected once into the vitreous cavity of the other eye as the control. The intraocular kinetics of nanospheres was evaluated by measuring vitreous fluorescence using a scanning fluorophotometer. To investigate elimination pathways of nanospheres in detail, serial cross-sections of the eyes were examined with a fluorescence microscope. The fluorescence derived from nanospheres was observed in the vitreous cavity for over 1 month (2 microm: t(1/2) = 5.4 +/- 0.8 days, 200 nm: t(1/2) = 8.6 +/- 0.7 days, 50 nm: t(1/2) = 10.1 +/- 1.8 days), whereas that in the control eyes completely disappeared within 3 days (t(1/2) = 7.8 +/- 0.7 h). The elimination half-life from the vitreous cavity correlated well with the particle diameter (r = -0.997, p = 0.007). Histological studies using a fluorescence microscope revealed that nanospheres with a diameter of 2 microm were seen in the vitreous cavity and trabecular meshwork, while nanospheres with a diameter of smaller than 200 nm were also observed in the retina as well as these tissues. Our findings indicated that nanospheres may be beneficial as a drug carrier to the retina, vitreous and trabecular meshwork. PMID- 11114603 TI - Disruption of the blood-aqueous barrier following retinal laser photocoagulation and cryopexy in pigmented rabbits. AB - Disruption of the blood-aqueous barrier (BAB) induced by retinal photocoagulation and cryopexy in pigmented rabbits was evaluated by laser flare photometry. A significant increase in flare values after retinal photocoagulation was measured from the 1st postoperative day, with values returning to baseline levels by day 7. Cryopexy induced consistently high flare values for 14 days. Intravitreal injection of interleukin (IL) 1, IL-6 and prostaglandin (PG) E(2) induced a significant increase in flare values. Following these treatments, introduction of a PG synthetase inhibitor can partially ameliorate BAB disruption. IL-1, IL-6 and PGE(2) may be involved in BAB disruption following retinal photocoagulation and cryopexy. PMID- 11114604 TI - Influence of dehydroepiandrosterone on rabbit intraocular pressure. AB - PURPOSE: The systemic concentration of dehydroepiandrosterone decreases with age in primates while in humans intraocular pressure (IOP) increases with aging. This study was designed to investigate if a relationship existed between dehydroepiandrosterone and IOP in pigmented rabbits. METHODS: Animals were treated unilaterally for 6 weeks with topical 3% dehydroepiandrosterone in 30% 2 hydroxypropyl-beta-cyclodextrin; the contralateral eye received vehicle alone. Drops were applied, and IOP measured, twice daily. RESULTS: Small, but statistically significant, drug-related effects were found. IOP was consistently higher in the afternoon; the afternoon minus morning difference in IOP, however, decreased with time. Topical, radioactive drug application indicated very low level penetration into aqueous humor, iris, corneal epithelium, the rest of the cornea, or bulbar conjunctiva. CONCLUSION: The small drug-related effects may be due, in large part, to poor intraocular drug penetration. The circadian rhythm of IOP appears to be time-dependent in chronic studies with a gradual loss of IOP difference between a.m. and p.m. readings. PMID- 11114605 TI - Tear nitrite and nitrate levels as nitric oxide end products in patients with Behcet's disease and non-Behcet's uveitis. AB - To evaluate the role of nitric oxide (NO) in ocular inflammation, tear nitrite and nitrates (NN) as NO end products were determined in 11 patients with Behcet's disease (BD) and in 11 with non-Behcet's uveitis (NBU) during the active and remission stages and in 12 healthy controls. Median (with the range) NN levels were 82.29 (59. 60-98.25) micromol/l in the active and 98.25 (52.88-246.92) micromol/l in the remission stage of BD; 88.17 (25.99-116.73) micromol/l in the active and 83.00 (31.04-250.28) micromol/l in the remission stage of NBU and 109.17 (88.17-158.74) micromol/l in the controls. The NN levels in the active stage of BD and NBU were significantly decreased when compared to the controls (p < 0.05; Kruskal-Wallis test). Decreased NN levels at the activation stage may be caused by the rapid transformation of the NO to peroxynitrites, which are highly oxidizing and cytotoxic substances. PMID- 11114606 TI - Preventive effect of diethyldithiocarbamate on selenite-induced opacity in cultured rat lenses. AB - Increasing evidence suggests the involvement of reactive oxygen species in the development of cataracts. In this study, we investigated the preventive effect of diethyldithiocarbamate (DDC) on the selenite-induced opacification of cultured rat lenses. Lens opacity was produced by incubation with 0.2 mM selenite for 24 h, which resulted in an increase in selenium content in the cultured lenses. The increase in selenium content and the onset of opacification and lens membrane damage were inhibited by preincubation with DDC. It is reasonable to assume that DDC contributed to anticataract ability. In addition, selenite resulted in a significant decrease in glutathione and protein thiol content and an increase in lipid peroxidation levels in the lenses. These alterations were also depressed by DDC, suggesting a contribution of an antioxidative effect by DDC in the inhibition of lens opacification. At the same lens selenium content, DDC treatment inhibited opacification and lipid peroxide. In conclusion, we propose that the antioxidative properties of DDC play a major role in its contribution to the anticataract effect. PMID- 11114607 TI - Turning information into knowledge to prevent health care-associated infections and other adverse events: the electronic ICP as an agent of change. PMID- 11114608 TI - Status of infection surveillance and control programs in the United States, 1992 1996. Association for Professionals in Infection Control and Epidemiology, Inc. AB - BACKGROUND: Nosocomial infections have been recognized as a source of morbidity and mortality throughout the world for several decades. In the United States, an estimated 2.1 million nosocomial infections occur annually in acute care hospitals alone. Infection surveillance and control programs (ISCPs) play a vital role in addressing this problem, but no national studies have described the status and composition of these programs since the 1970s. METHODS: In January 1997, a voluntary survey was sent by mail to members of the Association for Professionals in Infection Control and Epidemiology, Inc. Only one response per facility was requested. The survey asked for information for the years 1992 to 1996 (study period), and questions pertained to characteristics of the health care facility in which the respondent worked, characteristics of the ISCP and its personnel, and the overall level of administration support for infection control activities. RESULTS: Completed questionnaires were received from personnel at 187 health care facilities located in 40 states and the District of Columbia. The majority (76.5%) of responding facilities were nongovernment owned, and 57.2% were classified as general acute care facilities. The number of licensed beds at these facilities remained stable throughout the study period, but all other measures of facility size and activity (eg, number of patient days and number of nurses) decreased by as much as 28.9%. In 1992, ISCPs were most likely to be organizationally located in the Nursing Department, but by 1996, many had been transferred to departments of Medical Records, Quality Assurance, or Risk Management. Throughout the course of the study period, the number of facilities performing surveillance for health care-associated infections in outpatient settings increased by 44.0%, from 100 to 144. In 1996, only 47.6% of facilities had a hospital epidemiologist (HE), and HEs devoted a median of 15% or less of their time to infection control activities. For the most part, HEs were trained in infectious diseases, and few had certification in infection control. Infection control professionals (ICPs) were much more common than were HEs (ICPs were reported at 97.9% of respondents' facilities in 1996), and they spent the majority (80% in 1996) of their time on infection control activities. During the course of the study period, increasing numbers of facilities had ICPs who had certification in infection control. Furthermore, most respondents did not report a change over time in the level of administration support for infection control activities. CONCLUSIONS: Health care delivery has changed dramatically during the past 20 years. This study presents an updated description of ISCPs in the United States. Our results illustrate several changing parameters, such as departmental shifts and increased outpatient surveillance, that reflect adjustments in health care priorities during the study period. As the transformation of the health care system continues, continued evaluation of the status of ISCPs on a national level will be necessary. Diligent monitoring, proactive measures, and collaboration between infection control organizations and government agencies will be vital for the prevention and control of health care-associated infections in the future. PMID- 11114609 TI - Pilot testing standardized surveillance: Hospital Infection Standardised Surveillance (HISS). On behalf of the HISS Reference Group. AB - In Australia the time-consuming nature of double handling of surveillance data has meant that surveillance methodology rarely included prospective monitoring of patients at risk for the acquisition of a nosocomial infection. To streamline surveillance activities, infection control professionals favored the collection of case data either from the ward or pathology laboratories. By default, this method introduced a variety of definitions resulting in inconsistencies across health care facilities and artificial fluctuations in the magnitude of infection. In June 1998, the New South Wales Health Department funded its first attempt to develop and implement a standardized approach to collection of nosocomial infection data-Hospital Infection Standardized Surveillance (HISS). Six months later, in December 1998, 10 public acute care hospitals pilot tested the content and methodology of HISS. HISS members tested the application of the National Nosocomial Infection Surveillance system definitions for infection, active and passive surveillance methodology, the handheld computer for data collection, and the Electronic Infection Control Automated Technology (eICAT) version for HISS software and analysis. HISS member hospitals selected from several sentinel monitoring programs such as intravascular device-related bacteremia and nonintravascular device-related bacteremia infections, surgical site infections, respiratory syncytial virus infections, and rotavirus infections. Hospitals continued to perform active surveillance in the first 12 months, collecting demographic variables, risk factors, and outcomes. The completeness of the data sets for the two most frequently monitored programs, surgical site infections and intravascular device-related bacteremia, was high, with 99.6% of the required 36, 372 surgical site infection data fields and 99.4% of the 572,717 intravascular device-related bacteremia data fields completed. PMID- 11114611 TI - Antibiotic use and cost indicators at a rural hospital: a pilot project. AB - BACKGROUND: Recently, simple antibiotic use and cost indicators were developed for use in long-term care facilities. It was hypothesized that these indicators also may be applicable to the acute hospital setting. METHODS: For a 24-month period, data were collected quarterly on antibiotic use and cost indicators for 11 primary care physicians in a 40-bed rural hospital. Indicators included antimicrobial use ratio (AUR, ratio of the number of antibiotic days to the number of patient care days), cost per antibiotic day, and cost of antibiotics per patient care day. One-way analysis of variance and simple linear regression were used to analyze data. RESULTS: Quinolones (oral plus parenteral) accounted for 26% of the total antibiotic days (N = 6020) followed by ceftriaxone (19%) and cefuroxime (11.8%; oral plus parenteral). Overall trends in antibiotic use and cost included a significant increase in quarterly AUR (R(2) = 0.78, P =.004) and cost per patient care day (R(2) = 0. 82, P =.002) but no significant change in quarterly total antibiotic costs or cost per antibiotic day. Among physicians there was a significant difference in mean quarterly AUR (P <.001) and mean quarterly cost per patient care day (P <.001) but no significant difference in mean quarterly cost per antibiotic day. Variation in physician-specific cost per patient care day was best explained by variation in AUR (R(2) = 0.75, P <.001). CONCLUSIONS: Significant variation in simple antibiotic use and cost indicators was identified at a rural hospital from both the facility and physician perspective. Standardized methods for antibiotic use and cost monitoring, like the one described in this article, are required before the relationship between antibiotic use and resistance can be fully understood. PMID- 11114610 TI - Practical risk-adjusted quality control charts for infection control. AB - BACKGROUND: Control chart methodology has been widely touted for monitoring and improving quality in the health care setting. P charts and U charts are frequently recommended for rate and ratio statistics, but their practical value in infection control may be limited because they (1) are not risk-adjusted, and (2) perform poorly with small denominators. The Standardized Infection Ratio is a statistic that overcomes both these obstacles. It is risk-adjusted, and it effectively increases denominators by combining data from multiple risk strata into a single value. SETTING: The AICE National Database Initiative is a voluntary consortium of US hospitals ranging in size from 50 to 900 beds. The infection control professional submits monthly risk-stratified data for surgical site infections, ventilator-associated pneumonia, and central line-associated bacteremia. METHODS: Run charts were constructed for 51 hospitals submitting data between 1996 and 1998. Traditional hypothesis tests (P values <.05) flagged 128 suspicious points, and participating infection control professionals investigated and categorized each flag as a "real problem" or "background variation." This gold standard was used to compare the performance of 5 unadjusted and 11 risk adjusted control charts. RESULTS: Unadjusted control charts (C, P, and U charts) performed poorly. Flags based on traditional 3-sigma limits suffered from sensitivity <50%, whereas 2-sigma limits suffered from specificity <50%. Risk adjusted charts based on the Standardized Infection Ratio performed much better. The most consistent and useful control chart was the mXmR chart. Under optimal conditions, this chart achieved a sensitivity and specificity >80%, and a receiver operating characteristic area of 0. 84 (P <.00001). CONCLUSIONS: These findings suggest a specific statistic (the Standardized Infection Ratio) and specific techniques that could make control charts valuable and practical tools for infection control. PMID- 11114612 TI - Health care workers' experience with postexposure management of bloodborne pathogen exposures: a pilot study. AB - PURPOSE: This descriptive study of health care workers enrolled in a postexposure bloodborne pathogen management program had 3 goals: (1) to characterize their exposure incidents, (2) to assess health care workers' experience with the program, and (3) to identify strategies to improve the management of exposure incidents. METHODS: A confidential, self-administered, 5-page survey was mailed to 150 hospital employees who were recently evaluated in the employee health clinic for a blood/body fluid exposure. RESULTS: Sixty-five usable surveys were returned to the study office, representing a 43% response rate. Although the majority of the employees enrolled in the postexposure management program were generally satisfied with the overall quality of care they received, many respondents perceived a lack of social support during the lengthy follow-up period. Long-term distress related to the exposure was not uncommon. The respondents' suggestions for improvement focused on the need for department managers to become more personally involved when their staff members have an exposure incident. CONCLUSION: These qualitative data suggest that additional studies are needed to assess both the short-term and long-term impact of exposure incidents on the health and well being of affected health care workers. In addition, because of a paucity of information in this area, studies are needed to assess both the effectiveness of the United States Public Health Service recommendations for postexposure management and the degree to which they have been implemented by health care facilities. PMID- 11114613 TI - National Nosocomial Infections Surveillance (NNIS) system report, data summary from January 1992-April 2000, issued June 2000. PMID- 11114614 TI - Draft definitions for surveillance of infections in home health care. PMID- 11114615 TI - Multicenter surveillance study for nosocomial infections in major hospitals in Korea. Nosocomial Infection Surveillance Committee of the Korean Society for Nosocomial Infection Control. AB - BACKGROUND: The goals of a surveillance for nosocomial infections (NIs) are to observe the magnitude and characteristics of NIs and to plan and evaluate policies and guidelines of infection control. This study was designed to determine the rate and distribution of NIs and their causative pathogens in Korean hospitals. METHODS: Prospective surveillance was performed at 15 acute care hospitals with more than 500 beds during a 3-month period from June to August 1996. The case-finding methods were laboratory-based surveillance for patients in the general wards and a direct review of medical charts done regularly for all the patients in the intensive care units. RESULTS: A total of 3162 NIs were found among 85,547 discharged patients, with an overall nosocomial infection rate of 3.70 per 100 patients discharged. Urinary tract infections constituted 30.3% of all NIs. Other infections were pneumonias, 17.2%, surgical site infections, 15.5%, and primary bloodstream infections, 14.5%. The infection rate was the highest in neurosurgery (14.21), followed by neurology (8. 62) and ontology services (6.70). The infection rate in intensive care units was higher than it was in the general wards (10.74 vs 2. 57, P =.001). The commonly isolated organisms were Staphylococcus aureus (17.2%), Pseudomonas aeruginosa (13.8%), and Escherichia coli (12.3%). CONCLUSIONS: This first multicenter surveillance study provided extensive information on the current status and trends of NIs in major hospitals in Korea. The results may contribute to the evaluation of infection control programs and the development of effective strategies in these hospitals. PMID- 11114616 TI - The Internet: a practical example of the use of new technology in the assessment of vancomycin use in pediatrics. The Pediatric Prevention Network. AB - BACKGROUND: The rapid emergence of both new infections and new technologies has revolutionized health care during the past 50 years. Increased use of the Internet has enabled health care professionals to educate, interact, and collaborate throughout the world in ways never before possible. Increased use of vancomycin has been associated with the emergence of organisms with decreased susceptibility to vancomycin, such as Enterococcus and staphylococcal species. The purpose of this article is to describe our experience using Internet technology to assess vancomycin use at children's hospitals in the United States. METHODS: A Web-based evaluation was developed and distributed on the Internet to 57 Pediatric Prevention Network hospitals. The evaluation was structured to collect summary statistics on vancomycin use and admissions data by service for 1997 and 1998. RESULTS: Twenty-four hospitals were able to provide archived vancomycin use and patient admissions data; completed evaluations were returned from 15 hospitals (62.5% response rate). Personnel at 6 (40%) hospitals completed the evaluation directly on the Internet. CONCLUSIONS: In our study, Internet technology facilitated a more efficient evaluation of vancomycin use, but fewer than half of the personnel at Pediatric Prevention Network hospitals completed the evaluation directly on the Internet. It is unclear whether personnel at these hospitals were limited in Internet access, support, or understanding. Efforts should be directed to educate health care personnel on the advantages of the Internet. Furthermore, many of the pharmacy databases used in our assessment were not standardized across hospitals nor systematically validated. Understanding that limitations still remain-within the source of the data studied, the health care system sampled, and the Internet tools available-is essential because the Internet offers health care professionals today a tool both to protect patients and to improve quality throughout the world. PMID- 11114617 TI - Computer keyboards and faucet handles as reservoirs of nosocomial pathogens in the intensive care unit. AB - PURPOSE: We postulate that computer keyboards and faucet handles are significant reservoirs of nosocomial pathogens in the intensive care unit (ICU) setting. METHODS: Sterile swab samples were obtained from 10 keyboards and 8 pairs of faucet handles in the medical ICU at Tripler Army Medical Center during a period of 2 months. Methicillin-resistant Staphylococcus aureus (MRSA) obtained from the environmental and patient specimens were sent for DNA identification by using pulsed-field gel electrophoresis. RESULTS: A total of 144 samples were obtained (80 keyboards and 64 faucet handles), yielding 33 isolates. The colonization rate for keyboards was 24% for all rooms and 26% in occupied rooms. Rates for faucet handles in all rooms and occupied rooms were 11% and 15%, respectively. The environmental isolates annd their prevalence were: MRS, 49%; Enterococcus, 18%; Enterobacter, 12%; and all other gram-negative rods, 21%. Fourteen individual patient isolates were recorded: MRSA, 43%; Enterobacter, 21%; other gram-negative rods, 36%; and Enterococcus, 0%. By using pulsed-field gel electrophoresis, an indistinguishable strain of MRSA was identified in two patients, the keyboards and faucet handles in their respective rooms, and on other keyboards throughout the ICU, including the doctors' station. CONCLUSIONS: The colonization rate for keyboards and faucet handles, novel and unrecognized fomites, is greater than that of other well-studied ICU surfaces in rooms with patients positive for MRSA. Our findings suggest an associated pattern of environmental contamination and patient infection, not limited to the patient's room. Pulsed-field gel electrophoresis results have documented an indistinguishable strain of MRSA present as an environmental contaminant on these two fomites and in two patients with clinical infections patients during the same period. We believe these findings add evidence to support the hypothesis that these particular surfaces may serve as reservoirs of nosocomial pathogens and vectors for cross transmission in the ICU setting. New infection control policies and engineering plans were initiated on the basis of our results. PMID- 11114618 TI - Interdisciplinary care--the future of endocrine surgery. PMID- 11114619 TI - Papillary thyroid cancer with pulmonary metastases in children: long-term prognosis. AB - BACKGROUND: Papillary thyroid cancer (PTC) in young patients may rarely be encountered with pulmonary metastases. Previous studies have suggested that, in the pediatric population, this may not portend a lethal outcome. Our present study, children with pulmonary metastases, was designed to clarify this issue. METHODS: Fourteen children and young adolescents (mean age, 13.5 years; range, 9.8-17 years) with PTC and pulmonary metastases were treated at our institution between 1937 and 1998. Surgical treatment consisted of total thyroidectomy (n = 10 patients), subtotal thyroidectomy (n = 3 patients), and a biopsy only procedure (n = 1 patient). All patients who underwent thyroidectomy also underwent a variety of cervical lymph node dissections, and all patients proved to have regional nodal disease. After the operation, 12 patients were treated with ablative doses of (131)I, 1 patient was treated with external beam irradiation, and all patients were placed on suppressive thyroid hormone therapy. The mean length of follow-up was 19.3 years (range, 1-45 years). RESULTS: Regional recurrent disease developed in 2 patients (15%). No patient experienced the development of worsening pulmonary disease or extra-pulmonary metastases. All patients with recurrent disease underwent selective nodal resections. No patient died of metastatic PTC. Seven patients (50%) remain completely free of disease and are probably cured; 7 patients (50%) are asymptomatic with residual pulmonary disease. CONCLUSIONS: A stepwise treatment approach allows long-term survival and frequent cure for young patients with PTC and concomitant pulmonary metastases. PMID- 11114620 TI - Screening for genetic aberrations in papillary thyroid cancer by using comparative genomic hybridization. AB - BACKGROUND: Determination of the genetic composition of papillary thyroid cancers may help explain differences in observed clinical behavior. Comparative genomic hybridization (CGH) is a novel molecular cytogenetic assay that allows simultaneous detection of gains, losses, and amplification of genetic information, making it an ideal screening tool. The aim of this study was to identify genetic aberrations occurring in papillary thyroid cancers by using CGH analysis. METHODS: CGH analysis was performed on 21 individual cases of papillary thyroid cancers. Nonparametric statistical comparisons were performed with the Fisher exact test. RESULTS: Genetic abnormalities were identified by CGH in 10 of 21 cases (48%). A recurrent pattern of aberrations was seen in cases where genetic changes were detected, involving losses at chromosome arms 1p and 9q and chromosomes 17, 19, and 22, and gains at chromosome 4 and chromosome arms 5q, 6q, 9q, and 13q. The loss of chromosome 22 was unique to younger patients (P =.05) and was associated with a higher rate of regional lymphatic metastasis (19% vs 80%, P =.02). CONCLUSIONS: Two genetically unique groups of patients were identified by using CGH analysis. One group had no detectable aberrations; the other had a recurrent pattern of aberrations, localizing to the identical chromosomal loci. This pattern of aberrations suggests that the involved loci may contain genes important in thyroid carcinogenesis. The clinical significance of the presence of copy number changes detected by CGH needs to be determined. In addition, molecular cloning of involved genes in each of the aberrations is warranted. PMID- 11114621 TI - Left ventricular systolic and diastolic function and exercise testing in primary hyperparathyroidism-effects of parathyroidectomy. AB - BACKGROUND: Nontraditional manifestations of primary hyperparathyroidism (HPT) are controversial and may include morbidity, mortality, and risk factors for cardiovascular diseases. This study evaluates cardiovascular functions at rest and during exercise in HPT. METHOD: Thirty patients with HPT (mean serum calcium, 2.97 +/- 0.24 mmol/L) and 30 control people with normocalcemia, matched for age and sex, underwent symptom-limited exercise testing and echocardiography before and 13 months (mean) after having a parathyroidectomy. RESULTS: Despite similar maximal workload and blood pressures at rest in patients and healthy controls, HPT associated with higher systolic blood pressure during exercise (P =.03) and increased number of ventricular extrasystolic beats (P =.04). There was also an operatively reversible increase in ST-segment depression during exercise. Echocardiography showed an increased left ventricular (LV) isovolemic relaxation time (P =.02) and mitral deceleration time (P =.08), which indicate an LV diastolic dysfunction that could be partially reversed by operation. LV systolic function (ejection fraction and shortening fraction) tended to be elevated in HPT (P =.07 and.06, respectively) and diminished after parathyroidectomy. There was a trend toward higher LV mass, especially among the men with HPT (P =.06), which was unchanged postoperatively. CONCLUSIONS: HPT couples to reversible signs of myocardial ischemia and LV dysfunctions with a possible increased risk of life threatening arrhythmia. PMID- 11114622 TI - Value of intra-arterial calcium stimulated venous sampling for regionalization of pancreatic insulinomas. AB - BACKGROUND: Intra-arterial calcium stimulation with hepatic venous sampling (ASVS) for insulin gradients has been reported to be the most sensitive preoperative localizing technique for insulinomas. We reviewed our experience with ASVS to localize and guide the treatment of insulinomas over the past decade. METHODS: Eighteen patients who underwent ASVS before surgical exploration for insulinoma were studied. The accuracy of ASVS was compared with intraoperative findings and other localizing studies. RESULTS: There were no complications arising from the procedures. A more than 2-fold step-up in insulin level 30 to 60 seconds after injection to at least 1 feeding artery was observed in 16 patients. Fourteen of the 16 solitary tumors (87.5%) were correctly located; 100% (6/6 tumors) at the head and 80% (8/10 tumors) at the body/tail. The overall accuracy of this test was 89%, compared with 11%, 33%, 38%, and 63% of ultrasonography, computed tomography, magnetic resonance imaging, and endoscopic ultrasonography, respectively. Six enucleations and 10 distal resections were performed, which included 2 laparoscopic procedures. The combination of intraoperative ultrasonography with preoperative ASVS identified all tumors. CONCLUSIONS: ASVS is the most accurate preoperative localization tool for the localization of insulinomas and, in combination with intraoperative ultrasonography, can enhance surgical success. PMID- 11114623 TI - Overexpression of heme oxygenase-1 protects allogeneic thyroid grafts from rejection in naive mice. AB - BACKGROUND: Endocrine allografts are an option for the treatment of endocrine failure. METHODS: One lobe of the thyroid was transplanted under the kidney capsule. RESULTS: C57BL/10 (H2(b)) thyroids were rejected in naive CBA (H2(k)) mice within 14 days after transplantation. When mice were treated with anti-CD4 monoclonal antibodies (mAb), all grafts survived for more than 60 days. The first grafts still survived after second C57BL/10 or Balb/c (H2(d)) thyroid grafts that were transplanted into the same recipients were rejected acutely, which suggests that the primary grafts were modified under anti-CD4 mAb treatment. To confirm this hypothesis, C57BL/10 thyroid grafts from anti-CD4 mAb-treated mice were retransplanted. All grafts survived in naive mice; this correlated with the overexpression of heme oxygenase-1 (HO-1) in the grafts. Next, an inhibitor of HO 1 (zinc protoporphyrin) or control compound (copper protoporphyrin) was injected intraperitoneally after transplantation of C57BL/10 thyroid grafts into the primary CBA recipients that had been treated with anti-CD4 mAb. The grafts in mice that had been treated with zinc protoporphyrin, but not copper protoporphyrin, were rejected when retransplanted to naive recipients. CONCLUSIONS: Overexpression of HO-1 correlated with the protection of fully allogeneic thyroid grafts from rejection when retransplanted into naive recipients. PMID- 11114624 TI - Management of incidentally discovered adrenal masses and risk of malignancy. AB - BACKGROUND: Incidentally discovered adrenal masses of more than 1 cm in size are relatively frequent, but the correct management of such lesions is not well established. METHODS: Between 1980 and 1999, 158 patients (73 men [46.2%] and 85 women [53.8%]; median age, 58 years) with adrenal incidentalomas of more than 2 cm in size were observed. Sixty-eight patients (43.0%) underwent adrenalectomy. The main reasons for surgery were (1) suspicious computed tomography (CT) scan or magnetic resonance imaging (MRI) appearance or no uptake at the 75-Se norcholesterol scintigraphy (NCS; n = 22 patients), (2) an increase in the size of the mass of more than l cm (n = 15 patients), (3) subclinical endocrine hyperfunction (n = 14 patients), and (4) mass size of more than 5 cm (n = 22 patients), with imaging-guided fine-needle aspiration biopsy with spinal-type narrow-gauge needle (FNAB) that suggested malignancy (n = 5 patients). RESULTS: Pathologic examination showed 39 adrenocortical adenomas (57.4%), 20 adrenal malignancies (29.4%; carcinomas, 15; unsuspected metastases, 3; nonfunctioning malignant pheochromocytomas, 2), and 9 various benign lesions (13.2%). All masses that increased in size were benign. Seven malignant tumors (46.7%) were 3 to 4 cm in size, and 14 benign lesions (29.1%) were 5 to 6 cm in size. Sensitivity and specificity in the detection of malignancy were 100% and 100% for NCS (n = 34 patients) and FNAB (n = 19 patients), 75.0% and 93.7% for CT scan (n = 68 patients), and 87.5% and 100% for MRI (n = 26 patients), respectively. CONCLUSIONS: To differentiate benign and malignant incidentalomas, NCS and FNAB are more sensitive than CT scan and MRI; size criteria are of little value. PMID- 11114625 TI - Late parathyroid function after successful parathyroidectomy guided by intraoperative hormone assay (QPTH) compared with the standard bilateral neck exploration. AB - BACKGROUND: Controversy continues between bilateral neck exploration and limited parathyroidectomy. One approach depends on gland size and histopathologic factors; the other approach limits excision to only hypersecreting glands. Both have excellent early operative success, but late recurrence rates with limited exploration are unknown. METHODS: Three hundred twenty consecutive patients with primary hyperparathyroidism were followed 6 to 313 months after successful parathyroidectomy. One hundred seventy-six patients had bilateral neck exploration with excision of enlarged glands (group I); 144 patients had glands excised based on hyper-secretion (group II). Calcium and intact parathyroid hormone (iPTH) levels were measured yearly. Parathyroid gland hypersecretion was determined by elevated iPTH levels. RESULTS: In group I, 1 gland was excised in 160 patients (91%); 19 of 176 patients (11%) had elevated iPTH levels. In group II, 139 patients (97%) had 1 gland excised; 19 of 144 patients (13%) had high iPTH levels. The number of patients with more than 1 gland excised in group I (9%) is 3 times higher than in group II (3%) (P <.05). There was no significant difference in the incidence of recurrent hyperfunctioning glands between the 2 different operative approaches (chi-squared test). CONCLUSIONS: Late parathyroid gland function was comparable with both approaches. Multiple gland excision based on size alone may lead to excision of normal functioning glands. PMID- 11114626 TI - Normalization of intraoperative parathyroid hormone does not predict normal postoperative parathyroid hormone levels. AB - BACKGROUND: Intraoperative intact parathyroid hormone (iPTH) is being used to confirm complete excision of hyperfunctioning parathyroid tissue. It is uncertain whether normalization of intraoperative iPTH levels accurately predicts long-term postoperative iPTH values. METHODS: Fifty-two consecutive patients with primary or secondary hyperparathyroidism underwent parathyroidectomy with measurement of intraoperative iPTH. Ten patients were excluded due to incomplete laboratory follow-up. Follow-up serum calcium and iPTH levels were measured at 1- and 3 month intervals. RESULTS: Before operation, the mean serum iPTH level was 249 pg/mL (SD=208) and mean serum calcium level was 11.4 +/- 0.9 mg/dL (+/- SD). In all but 4 patients, final intraoperative iPTH levels normalized to less than 67 +/- 41 pg/mL (mean, 35 pg/mL). One week after operation, serum calcium levels had returned to normal (mean, 9.4 +/- 1.1 pg/mL), which directly correlated with the final intraoperative serum iPTH values (Pearson correlation, r = -.434; P <.01). By 1 month, all but 2 patients were normocalcemic (mean, 9.4 +/- 0.9 pg/mL) with a mean iPTH level of 74.8 +/- 82 pg/mL. There was no correlation between final intraoperative and postoperative serum iPTH values (r =.099; P <.533). Both patients with persistent hypercalcemia at 1 month had appropriate intraoperative decreases in iPTH values. CONCLUSIONS: Intraoperative serum iPTH levels significantly correlate with postoperative serum calcium levels but not with postoperative serum iPTH levels. There was a 4.8% failure rate in the correction of postoperative serum calcium levels and a 29% failure rate in the normalization of postoperative serum iPTH levels. PMID- 11114627 TI - Noninsulinoma pancreatogenous hypoglycemia syndrome: an update in 10 surgically treated patients. AB - BACKGROUND: Neuroglycopenia from endogenous hyperinsulinism usually is caused by insulinomas in adults. We recently reported a novel hypoglycemic disorder in 5 patients (patients 1 to 5) with postprandial neuroglycopenia, negative 72-hour fasts, negative perioperative imaging studies, but positive calcium stimulation tests and islet hypertrophy and nesidioblastosis in the gradient-guided resected pancreata. METHODS: In this report we compare our experience with 5 additional patients (patients 6 to 10) with this syndrome to that in the original report. RESULTS: The clinical features of patients 6 to 10 were similar to those of patients 1 to 5. Each had positive calcium stimulation testing that guided the extent of the distal pancreatectomy and histologic evidence of islet cell hypertrophy or nesidioblastosis. All 10 patients are alive from 9 to 50 months after operation, 1 of whom had no amelioration of neuroglycopenia. Minor perioperative complications occurred in 3 patients. One patient has experienced repeated bouts of acute pancreatitis, pseudocyst formation, and exocrine insufficiency. CONCLUSIONS: We have identified adult patients with severe, postprandial hyperinsulinemic hypoglycemia from diffuse islet cell disease, 80% of whom have been well palliated with surgery. The results in 7 men have been better than those in the 3 women for reasons that are not obvious. PMID- 11114628 TI - Strategy for identification of novel glucose transporter family members by using internet-based genomic databases. AB - BACKGROUND: We previously reported that medullary thyroid carcinomas and pheochromocytomas avidly take up the glucose analog fluoro-deoxyglucose on positron emission tomography but do not express any of the known human facilitative glucose transporters. We therefore hypothesized that a novel glucose transporter is responsible for glucose uptake in these tumors. METHODS: Internet based Expressed Sequence Tags and high throughput genome sequence databases were screened for novel sequences homologous to the known glucose transporters. Derived clones were used to screen cDNA libraries. Sequence comparison and hydropathic analysis of the putative proteins were performed. RESULTS: We identified 2 novel genes (GLUT8 and GLUT9) that are members of the facilitative glucose transporter family. The putative GLUT8 and GLUT9 proteins have 44% and 31% sequence identity to GLUT5 and GLUT3, respectively. Hydropathic analysis showed both have exofacial and transmembrane domains consistent with a hexose transporter. CONCLUSIONS: By using the Expressed Sequence Tags database, we identified novel members of the glucose transporter family. Further work will establish function and expression patterns in medullary thyroid carcinomas and pheochromocytomas. Internet-based genomic databases allow rapid screening and identification of candidate sequences of novel members of human gene families. PMID- 11114629 TI - The helix-loop-helix transcription factor, Id-1, is overexpressed in medullary thyroid cancer. AB - BACKGROUND: The Id-1 helix-loop-helix protein inhibits differentiation and enhances cell proliferation. It is required for cell cycle progression. The Id-1 gene is highly expressed in a variety of tumor-derived cell lines. It increases after mitogen stimulation and is overexpressed in some human neoplasms. Therefore, we hypothesized that the Id-1 gene may play a role in medullary thyroid carcinogenesis. METHODS: The expression of the Id-1 protein in human medullary thyroid cancer (MTC) and the corresponding normal thyroid tissue was determined by Id-1 immunohistochemistry. In a human MTC cell line (TT), the effects of growth stimulation and redifferentiation on Id-1 expression were determined by Northern blot analysis. RESULTS: Id-1 immunostaining intensity in 9 MTC samples (6 sporadic, 2 familial, and 1 MEN 2A) was moderate to strong. However, it was absent or faint in the corresponding normal thyroid tissue. The Id-1 protein was significantly overexpressed in MTC compared with corresponding normal thyroid tissue on the basis of the percentage of positive cells and immunostaining intensity (P =.002). In the TT cell line, Id-1 messenger RNA (mRNA) expression was increased 4-fold after growth stimulation with serum. Phorbol ester (which induces redifferentiation in the TT cell line) downregulated Id-1 mRNA expression. CONCLUSIONS: Id-1 is overexpressed in MTC. The Id-1 gene may play a role in the regulation of MTC differentiation and proliferation. PMID- 11114630 TI - Pancreaticoduodenal endocrine tumors in multiple endocrine neoplasia type 1: surgery or surveillance? AB - BACKGROUND: The management of pancreaticoduodenal endocrine tumors (PETs) remains controversial in multiple endocrine neoplasia type 1 (MEN 1). METHODS: Twenty-one patients with MEN 1 and PETs were analyzed for outcome of surgery and surveillance with special regard to the genotype based on MEN1 gene mutation analysis. RESULTS: Nine patients had gastrinomas, 5 had nonfunctioning tumors, 4 had insulinomas, 2 had insulinomas and gastrinomas, and 1 had a VIPoma. Seven patients (33%) had malignant tumors. Sixteen patients (76%) were initially treated by pancreatic resections or tumor enucleations or both. Six patients underwent reoperations for recurrences or lymph node metastases or both. Fifteen of the 16 operated patients are alive, and 12 have no evidence of disease after a median follow-up of 78 months (range, 1-198 months). Five patients with gastrinomas or nonfunctioning tumors, but no symptoms, underwent surveillance; 1 of them developed lymph node metastases. Patients with truncating mutations in the N- or C-terminal region (exons 2, 9, or 10) of the MEN1 gene had a significantly higher rate of malignant tumors (55% vs 10%; P <.05) than patients with other mutations. CONCLUSIONS: An aggressive surgical approach is justified for PETs in patients with MEN 1. However, MEN1 gene mutations in exons 3 to 8 seem to be associated with mild behavior of PETs, possibly allowing surveillance in asymptomatic patients. PMID- 11114631 TI - Is calciphylaxis best treated surgically or medically? AB - BACKGROUND: Calciphylaxis is a rare, painful, life-threatening problem of cutaneous necrosis and refractory healing in patients with uremia and secondary hyperparathyroidism. The pathogenesis involves abnormalities in calcium and phosphorus metabolism and acute deposition of calcium in tissues. METHOD: The clinical course of 16 patients who were diagnosed with calciphylaxis at our institution from 1994 through 1998 was reviewed. RESULTS: Fourteen female patients and 2 male patients had chronic renal disease, secondary hyperparathyroidism, and characteristic skin necrosis (mean age, 56 years; range, 39-70 years). All patients underwent intensive medical therapy, including ongoing hemodialysis (n = 16 patients), parathyroidectomy (n = 7 patients), and debridement of cutaneous lesions (n = 8 patients). Mean serum values in surgical and nonsurgical patients were significantly different for phosphorus, calcium phosphorus product, and parathormone levels. Median survival was 9.4 months; 15 patients (93%) have died. The median survival time for parathyroidectomy versus nonparathyroidectomy was 14.8 and 6.3 months (P =.22), for skin debridement versus nondebridement was 14.1 and 6.1 months (P =.08), and for diabetic versus nondiabetic patients was 6.5 and 13.9 months (P =.11). CONCLUSIONS: Calciphylaxis has a female preponderance, with a dismal prognosis. A multidisciplinary approach that uses frequent hemodialysis to normalize calcium and phosphorus levels and local debridement of skin lesions seems prudent. Parathyroidectomy cannot be recommended routinely in all patients, unless severe hyperparathyroidism mandates intervention. PMID- 11114632 TI - Limitations of size as a criterion in the evaluation of adrenal tumors. AB - BACKGROUND: Size has been considered to be the single best predictor of malignancy in adrenal neoplasms that have been identified incidentally. However, small adrenal cortical cancers have been reported from multiple centers. METHODS: We retrospectively evaluated the value of tumor size and other clinical parameters in the prediction of the presence of adrenal malignancy. RESULTS: The records of 117 patients who underwent evaluation for tumors of the adrenal gland were reviewed. The median tumor size of the adrenal cortical carcinomas (n = 38 carcinomas) was 9.2 cm (range, 1.7-30 cm); 5 cancers (13.5%) were smaller than 5.0 cm. The median overall size of the benign tumors, excluding pheochromocytomas, was 4.0 cm (n = 38 carcinomas); 10 benign tumors (26%) were larger than 5.0 cm. The imaging features of 4 of 5 small adrenal cancers predicted malignancy; the remaining patients had hormonally functioning tumors. The imaging features of 7 of 10 large benign adrenal tumors predicted benign histologic features, including 5 of 5 myelolipomas. CONCLUSIONS: Although size remains a good predictor of the histologic features and clinical behavior of adrenal neoplasms, both small adrenal cortical cancers and large benign tumors occur with measurable frequency. High-quality imaging studies may be helpful in the identification of relatively small adrenal cancers and of characteristic benign lesions that may be selectively followed. PMID- 11114633 TI - Prognostic significance of RET and NTRK1 rearrangements in sporadic papillary thyroid carcinoma. AB - BACKGROUND: The expression of RET/PTC chimeras was demonstrated in 10% to 20% of sporadic papillary thyroid carcinomas (PTCs), whereas rearrangements of NTRK1 were detected less frequently. Some investigators have hypothesized that RET/PTC activation is preferentially associated with slow-growing tumors of low malignancy in elderly patients; other studies support the contrary. METHODS: Expression analysis of RET and NTRK1 was performed by duplex reverse transcription-polymerase chain reaction in tumor tissues from 119 patients with PTC. Samples with suspected rearrangements were further analyzed for the expression of the hybrid messenger RNAs RET/PTC 1 to RET/PTC 7 and for known NTRK1 chimeras, respectively. RESULTS: Seventeen of 119 tumors (14.3%) revealed somatic rearrangements of RET; NTRK1-derived hybrids were demonstrated in 15 cases (12.6%). In patients with RET/PTC chimeras, a statistically not significant tendency towards younger age, lower recurrence rate, and improved survival was observed, despite increased incidence of lymph node metastasis. Cumulative survival analysis of NTRK1 rearrangement-positive individuals demonstrated a worse outcome when compared with patients with expression of RET hybrids (P =.055). CONCLUSIONS: The high incidence of yet uncharacterized NTRK1 hybrid mRNAs in our patient cohort leads to the speculation that activating chromosomal rearrangements of several tyrosine kinase receptors may be a common feature of PTCs and that the expression of distinct chimeras may potentially be of prognostic significance. PMID- 11114634 TI - Hypothyroidisim following hemithyroidectomy: incidence, risk factors, and management. AB - BACKGROUND: The purpose of this study was to characterize the hypothyroidism that occurs following hemithyroidectomy. METHODS: The records of all euthyroid patients who underwent hemithyroidectomy from 1992 to 2000 were reviewed to determine the frequency of postsurgical hypothyroidism and the predisposing factors. All patients were evaluated for age, gender, serum thyrotropin (TSH) levels, weight of resected thyroid tissue, and associated thyroiditis. Hypothyroid patients were evaluated for symptoms, timing of diagnosis, and treatment doses of levothyroxine (L-T(4)). RESULTS: Hypothyroidism was diagnosed in 25 (35%) of 71 patients, subclinical in 16 and overt in 9 with a mean postoperative TSH level of 8.51 +/- 6.53 microIU/L. The mean preoperative TSH level was 1.94 +/- 1.00 microIU/L in hypothyroid compared with 1.10 +/- 0.74 microIU/L in euthyroid patients (P <.05). Lymphocytic thyroiditis was present in 10 (40%) of 25 hypothyroid compared with 10 (22%) of 46 euthyroid patients (P = not significant). There were no significant differences in age, gender, or weight of resected thyroid tissue. The average therapeutic dose of L-T(4) was 1.3 microg/kg (range, 0.5 to 1.9 microg/kg). All but 2 hypothyroid patients were diagnosed within 2 months of operation. CONCLUSIONS: Hypothyroidism following hemithyroidectomy occurs in patients with higher preoperative TSH levels, is usually mild and asymptomatic, and can be treated with reduced doses of L-T(4). PMID- 11114635 TI - A murine model of allogeneic adrenocortical cell transplantation: perspectives for the treatment of Addison's disease in humans. AB - BACKGROUND: Hormone substitution for the treatment of adrenocortical insufficiency (Addison's disease) does not adequately substitute the hormone peaks required in stress situations. Therefore, allogeneic transplantation of adrenal cortex could offer an intriguing alternative. METHODS: Major histocompatibility complex (MHC) class I transgenic mice were used for the implementation of an animal model of adrenocortical cell transplantation in adrenalectomized mice. K(b)-transgenic cells and allogeneic adrenocortical cells were cocultured in mixed lymphocyte cultures to examine the alloimmune response. Lymphocytes from T-cell receptor transgenic mice and normal allogeneic mice served as responder cells. The effect of corticosteroids secreted by adrenocortical cells was antagonized by the steroid receptor antagonist mifepristone (RU486). RESULTS: In vitro coculture experiments showed that MHC class I disparate adrenocortical cells failed to activate B10.BR and T-cell receptor transgenic lymph node cells. In the presence of mifepristone this inhibitory effect was antagonized, resulting in strong lymphocyte proliferation. Activation of B10.BR lymphocytes by K(b)-disparate spleen cells was also abolished in the presence of adrenocortical cells. This effect, however, could not be reversed by mifepristone. CONCLUSIONS: In vitro, the presence of adrenocortical cells potently suppressed allogeneic immune responses. This effect was only in part due to the secretion of corticosteroids, pointing to an additional immunomodulatory property of adrenocortical cells. PMID- 11114636 TI - Pheochromocytoma: inherited associations, bilaterality, and cortex preservation. AB - BACKGROUND: Hereditary pheochromocytoma (HP) is characterized by early onset, bilateral adrenal involvement, low malignancy rate, and genetic linkage with certain familial syndromes. This retrospective review is intended to show the high yield of surveillance, predictable bilaterality, and the challenge of cortex sparing adrenalectomy. METHODS: From 1964 to 1999, 32 patients with HP were treated at a single institution and followed for a mean of 7 years. There were 15 cases of multiple endocrine neoplasia type 2A (MEN 2A), 12 cases of von Hippel Lindau (VHL) disease, 3 cases of von Recklinghausen's disease (VRD), and 2 cases of familial pheochromocytoma. Twenty-four of 32 patients underwent bilateral adrenalectomy (9 metachronous). Subtotal resection with orthotopic cortex preservation was performed in 5 patients, and heterotopic autografting was performed in 14 patients. RESULTS: Pheochromocytoma was the first manifestation in 50% of patients with VHL disease and in 27% of patients with MEN 2A. Surveillance uncovered medullary thyroid cancer in 5 of 15 patients with MEN 2A and hemangioblastomas, renal cell carcinoma, and islet cell tumors in 7 of 15 patients with VHL disease and VRD. HP was bilateral in 24 of 32 patients (14/15 in patients with MEN 2A, 7/12 in patients with VHL disease, 2/3 in patients with VRD, and 1/2 in patients with familial pheochromocytoma). In 9 cases of metachronous adrenalectomy, the mean interval was 67 months (range, 9-156 months). Three of 5 patients who underwent orthotopic preservation of the adrenal cortex experienced recurrence compared with 0 of 14 patients with heterotopic autotransplantation of cortical tissue. CONCLUSIONS: Pheochromocytoma frequently heralds coexisting silent VHL disease or MEN-2, mandating surveillance for inherited associations. The long interval of metachronous pheochromocytoma argues against prophylactic removal of the contralateral "normal" adrenal gland. Total adrenalectomy and heterotopic autotransplantation of medulla-free cortex may diminish the need for lifelong steroid substitution and eliminates recurrence. PMID- 11114637 TI - Randomized trial of parathyroidectomy in mild asymptomatic primary hyperparathyroidism: patient description and effects on the SF-36 health survey. AB - BACKGROUND: The treatment of patients with asymptomatic primary hyperparathyroidism remains controversial despite a National Institutes of Health consensus statement. This statement also recommended a randomized clinical trial because none exists to address this issue. METHODS: Informed consent was obtained from 53 asymptomatic patients with confirmed asymptomatic primary hyperparathyroidism who participated in this randomized trial of parathyroidectomy versus observation. Patients completed the SF-36 Health Survey, an instrument that measures wellness, every 6 months for 2 years. Average annual changes were compared. RESULTS: Fifty-three patients (42 female, 11 male) with asymptomatic, mild (serum calcium level, 10.1-11.5 mg/dL) asymptomatic primary hyperparathyroidism who agreed to participate were randomized into either a surgical group or an observation group. The mean calcium level was 10.31 mg/dL. The only demographic difference between groups was age, with the operative group being older (66.7 vs 62.6 years; P <.03). The scores on 2 of the 9 domains of the SF-36 were significantly different (P <.007 and <.012, respectively); both favored the operative group. CONCLUSIONS: Improved function is seen after parathyroidectomy when compared with patients who did not undergo operation. This study supports surgical management of mild primary hyperparathyroidism at the time of diagnosis because many patients have reversible nonclassic symptoms of the disease. PMID- 11114638 TI - Clinical and genetic analysis of patients with pancreatic neuroendocrine tumors associated with von Hippel-Lindau disease. AB - BACKGROUND: Patients with von Hippel-Lindau disease (VHL) may develop pancreatic neuroendocrine tumors (PNETs), which can behave in a malignant fashion. We prospectively evaluated size criteria for resection of lesions and the role of genotype/phenotype analysis of germline VHL mutations in predicting clinical course. METHODS: From December 1988 through December 1999 we screened 389 patients with VHL. The diagnosis of PNET was made by pathologic analysis of tissues or by radiographic appearance. Germline mutations were determined by quantitative Southern blotting, fluorescence in situ hybridization and complete gene sequencing. RESULTS: Forty-four patients with PNETs have been identified; 25 have undergone surgical resection, 5 had metastatic disease, and 14 are being monitored. No patient who has undergone resection based on tumor size criteria has developed metastases. Patients with PNETs were more likely to have missense mutations (58%), and 4 of 5 patients (80%) with metastatic disease had mutations in exon 3 compared with 18 of 39 (46%) patients without metastatic disease. CONCLUSIONS: Imaging for detection and surgical resection based on size criteria have resulted in the successful management of VHL patients with PNETs. Analysis of germline mutations may help identify patients at risk for PNET and which patients may benefit from surgical intervention. PMID- 11114639 TI - Intraoperative decay profile of intact (1-84) parathyroid hormone in surgery for renal hyperparathyroidism--a consecutive series of 80 patients. AB - BACKGROUND: The utility of intraoperative parathyroid hormone (PTH) monitoring is unclear in the surgical management of renal hyperparathyroidism. Our goal was to define the normal pattern of decay during operation for renal hyperparathyroidism by using the rapid intact (1-84) parathyroid hormone (PTH) assay. METHODS: Eighty consecutive patients underwent neck exploration for renal hyperparathyroidism. Intact PTH levels were monitored with a rapid immunochemiluminometric assay. Samples were assayed at the induction of anesthesia, after dissection before resection, and 20 and 40 minutes after resection. Follow-up ranged from 3 to 24 months. RESULTS: Twenty minutes after resection, PTH levels remained many-fold supranormal. Seventy-seven patients (96%) were cured. Of these, 75 patients (94%) had PTH decay of more than 50% from the preoperative level; 74 (99%) were cured. Only 1 of 3 patients (33%) in whom the PTH level decreased less than 40% from the preoperative level was cured. Two patients had intermediate values and both were cured. CONCLUSIONS: The intraoperative decay of PTH during operation for renal hyperparathyroidism is slower than for patients with normal renal function. However, 20 minutes after resection, a decline to less than 50% of the preoperative level predicts cure, while a level greater than 60% predicts failure. PMID- 11114640 TI - Endoscopic endocrine neck surgery with carbon dioxide insufflation: the effect on intracranial pressure in a large animal model. AB - BACKGROUND: Endoscopic endocrine neck surgery requires insufflation with carbon dioxide (CO(2)) at 10 to 15 mm Hg, which may decrease the cerebral venous return and increase intracranial pressure. This study evaluated the effect of CO(2) neck insufflation on intracranial pressure (ICP) and hemodynamic parameters. METHODS: Fifteen pigs underwent endoscopic thyroid dissection. Insufflation was performed with CO(2) at 0 (sham), 10, 15, and 20 mm Hg and with helium at 20 mm Hg with 3 pigs in each group. ICP, mean arterial pressure, central venous pressure (CVP), cardiac output, and blood gas were measured at baseline, 30, 60, and 120 minutes. RESULTS: There were no differences in mean ICP between the sham group and CO(2) insufflation at 10 mm Hg. Mean ICP increased significantly with CO(2) at 15 and 20 mm Hg and with helium at 20 mm Hg. A significant increase in CVP occurred in pigs operated with CO(2) at 20 mm Hg. We observed jugular vein collapse under all insufflation pressures; however, pigs operated at 10 mm Hg were able to maintain an intermittent blood flow. CONCLUSIONS: A severe increase in ICP occurs with insufflation pressures higher than 15 mm Hg, possibly as a result of decreased cervical venous blood flow. Carbon dioxide insufflation up to 10 mm Hg does not alter ICP and is recommended for clinical application in endoscopic neck surgery. PMID- 11114641 TI - Is familial non-medullary thyroid carcinoma more aggressive than sporadic thyroid cancer? A multicenter series. AB - BACKGROUND: The aggressiveness of familial non-medullary thyroid cancer (FNMTC) has been a subject of debate. The purpose of the study was to determine whether FNMTC is more aggressive than sporadic thyroid cancer. METHODS: A multicenter retrospective matched-case control study of FNMTC versus sporadic non-medullary thyroid cancer was conducted. Disease-free survival (time to recurrence) for both groups was compared. RESULTS: Forty-eight familial cases were compared with 144 age-, gender-, and stage-matched controls. Patients with FNMTC had a significantly shorter disease-free survival compared with sporadic non medullary thyroid cancer. Patients with FNMTC who presented with evidence of distant metastasis, or who were from families with more than 2 thyroid cancer-affected members, had the worst prognosis. The available staging systems were less likely to predict the outcome in patients with FNMTC than in patients with sporadic non medullary thyroid cancer unless one accounted for the strength of family history in the staging system. CONCLUSIONS: FNMTC is more aggressive than sporadic non medullary thyroid cancer. The best predictors of a poor outcome in patients with FNMTC are the number of family members affected by thyroid cancer and evidence of distant metastasis. PMID- 11114642 TI - Update on the MEN 2A c804 RET mutation: is prophylactic thyroidectomy indicated? AB - BACKGROUND: Mutations of the RET proto-oncogene co-segregate with multiple endocrine neoplasia type 2A. A rare sequence abnormality at codon 804 (c804) has been reported in 6 kindreds and linked to mild C-cell disease, which raises the question of the appropriateness of thyroidectomy in childhood. The purpose of this study was to (1) report the clinical correlates of 5 additional c804 kindreds, and (2) clarify therapeutic options in children. METHODS: Thirty-eight members from five c804 kindreds underwent genetic analysis. Biochemical, operative, and pathology reports were reviewed. RESULTS: Twenty-three gene carriers were identified, of whom 14 had thyroidectomy. Medullary thyroid carcinoma was found in 7 patients (aged 5-56 years), C-cell hyperplasia in 6 patients (aged 13-40 years), and normal histology in a single patient (aged 27 years). One patient with medullary thyroid carcinoma died of metastases (aged 12 years). Nine of the 23 gene carriers delayed operation, 4 of whom had calcitonin testing. Three of the 4 patients had abnormal calcitonin levels and a single patient was negative (aged 40 years). Of the remaining 9 patients, 2 await thyroidectomy, and 3 have refused evaluation. CONCLUSIONS: Penetrance of the c804 mutation is highly variable. Medullary thyroid carcinoma associated with this genotype has aggressive potential. Prophylactic thyroidectomy in childhood is a viable approach. PMID- 11114643 TI - Neutralizing vascular endothelial growth factor activity inhibits thyroid cancer growth in vivo. AB - BACKGROUND: Without angiogenesis, tumor growth is limited to a few millimeters, the limit of diffusion. Vascular endothelial growth factor (VEGF) is an endothelial-specific mitogen and a major regulator of angiogenesis. METHODS: To investigate the relationship between VEGF and thyroid tumor angiogenesis, we xenografted human dermal matrix inoculated with FTC-133 cells into nude mice or directly injected FTC-133 cells subcutaneously. To block the function of VEGF, the neutralizing anti-VEGF monoclonal antibody A.4.6.1 (mAb A.4.6.1) was injected intraperitoneally twice weekly. As control, an antibody of the same isotype (Ab 5B6) or phosphate buffer saline solution (PBS) was used. To evaluate the dermal matrix as a model for angiogenesis studies, recombinant human VEGF was inoculated into the dermal matrix pocket and xenografted into mice. RESULTS: In the dermal matrix angiogenesis model, the number of blood vessels paralleled the concentration of recombinant human VEGF and was highest at 100 ng/mL. Mice that were treated with the mAb A4.6.1 developed fewer blood vessels (mean, 6.6 per HPF) than control mice (18 per HPF in Ab 5B6 and 22 per HPF in PBS; P <.01). Tumors from mice that were treated with mAb A.4.6.1 were much smaller (mean +/- SD, 0.09 +/- 0.02 gm) at 5 weeks, compared with the tumors treated with Ab 5B6 (5.38 +/- 1.15 gm) or PBS (4.0 +/- 0.72 gm; P <.001). CONCLUSIONS: VEGF is produced by the follicular thyroid cancer cell line and stimulates angiogenesis and growth of thyroid cancer. This stimulation can be blocked by mAb A.4.6.1. PMID- 11114644 TI - Value of preoperative diagnostic modalities in patients with recurrent thyroid carcinoma. AB - BACKGROUND: Patients with well-differentiated thyroid cancer (WDTC) regularly have an excellent prognosis. However, tumor recurrence either involving the thyroid bed or the regional lymph nodes, or both, can be associated with significant morbidity and even mortality. The aim of the follow-up after primary surgery is to detect recurrent disease at its earliest stage. We assessed the value of different diagnostic methods in detecting locoregional recurrence in patients with WDTC. METHODS: We prospectively identified 150 patients with WDTC. Of those, 43 (28.7%) presented with recurrent disease. Ultrasonography-guided fine needle biopsy (US-FNB), iodine 131 ((131)I) wholebody scintigraphy, thyroglobulin (Tg) measurement, and fluorine-18 fluorodeoxyglucose positron emission tomography (FDG-PET) were carried out. RESULTS: Ultrasonography detected malignant lesions in 95.3% of the patients. The true positive rate of US-FNB was 95.3%. (131)I scanning had true positive, false negative, and false positive results in 54.2%, 40.0%, and 5.7% of the cases, respectively. In 85.7% of the patients, Tg levels were within pathologic range. Among the 13 patients who underwent FDG-PET, 84.6% showed pathologic uptake indicating malignancy. US and US-FNB provided the highest specificity for detecting recurrence (P <.001). CONCLUSIONS: In patients with WDTC and locoregional recurrence, US and US-FNB are the most sensitive methods in detecting local recurrence or regional lymph node metastases. FDG-PET is valuable in case of negative (131)I scanning results and elevated serum Tg levels. The method has limitations in finding minimal disease. PMID- 11114645 TI - The role of frozen section, gender, age, and tumor size in the differentiation of follicular adenoma from carcinoma: a meta-analysis. AB - BACKGROUND: The purpose of this study was to reanalyze modern trials and use meta analysis to determine how well frozen section gender, age, and tumor size could differentiate follicular adenoma from follicular carcinoma. METHOD: Inclusion criteria were studies where patients had a permanent pathologic diagnosis of follicular adenoma or follicular carcinoma and underwent frozen section or had clinical features recorded. Data were pooled, and the random effects model of meta-analysis was used. A probability value of less than.05 was considered significant. RESULTS: Nineteen studies were included (n = 3486 patients). Frozen section was evaluated in 11 studies (n = 2204 patients). Frozen section had an 87% sensitivity, a 48% specificity, a 92% and 35% positive and negative predictive value, respectively, an 82% accuracy, an odds ratio of 0.181, a 95% confidence interval (CI) of 0.07 to 0.49, and a probability value of.001. Clinical features were evaluated in 10 studies (n = 1954 patients). Of the patients with follicular carcinoma, 27.5% were male compared with patients with follicular adenoma, of whom 17.7% were male (P <.01; odds ratio, 2.17; CI 1.3 3.6; P =.003). Of the patients with follicular carcinoma, 52.2% were older than 50 years (52.2%) compared with patients with follicular adenoma, of whom 28.5% were older than 50 years (P <.001). Of patients with follicular carcinoma, 36.8% had tumors larger than 3 to 5 cm compared with patients with follicular adenoma, of whom 14.7% had tumors larger than 3 to 5 cm (P <.001; odds ratio, 3.99; CI 1.5 10.8; P =.006). CONCLUSIONS: Meta-analysis suggests that frozen section is not a specific test and cannot be used to confidently rule out follicular carcinoma. Male gender and large tumor size are significantly associated with carcinoma. PMID- 11114646 TI - The "false" nonrecurrent inferior laryngeal nerve. AB - BACKGROUND: Communicating branches between the cervical sympathetic system and the inferior laryngeal nerve (ILN) have been described. They usually originate from the middle cervical sympathetic ganglion (MCSG). These branches (sympathetic inferior laryngeal anastomotic branch [SILAB]), usually thin, sometimes have the same diameter as the ILN. In this study we prospectively evaluated the frequency of this condition and its implications during surgical neck exploration. METHODS: From November 1998 to October 1999, 791 patients underwent surgical neck exploration, and 1253 ILNs were dissected: 656 on the right side (52.3%) and 597 on the left side (47.7%). RESULTS: On the right side, a nonrecurrent ILN was found in 3 cases (0.46%), and a large SILAB was found in 10 cases (1.5%). The SILAB originated from the superior cervical sympathetic ganglion in 2 cases and directly from the sympathetic trunk above the MCSG in 8 cases. No anomalous branch was found on the left side. CONCLUSIONS: The SILAB may originate not only from the MCSG but also from the superior cervical sympathetic ganglion or directly from the sympathetic trunk. When the SILAB is as large as the ILN, it could be mistaken for a nonrecurrent ILN. The awareness of this anatomic condition during neck dissection may help to avoid injuries of the genuine ILN running in the usual pathway. PMID- 11114647 TI - A novel, noninvasive imaging technique for intraoperative assessment of parathyroid glands: confocal reflectance microscopy. AB - BACKGROUND: Successful surgical management of primary hyperparathyroidism requires the ability to identify and distinguish normal from abnormal parathyroid tissue. Microscopic pathologic confirmation often helps with the diagnoses and decisions regarding the extent of parathyroid resection. Confocal reflectance microscopy (CRM) is an optical method of noninvasively imaging tissue without fixation, sectioning, and staining as in standard histopathology. The goal of this study was to determine if CRM imaging could be used to distinguish normal from diseased parathyroid tissue intraoperatively. METHODS: In this study, 44 parathyroid glands from 21 patients undergoing operations for primary hyperparathyroidism were imaged immediately after excision. CRM images were compared with conventional hematoxylin-and-eosin stained sections obtained from the same gland. The percentage area occupied by fat cells was calculated in images of both normal and diseased glands. RESULTS: Characteristic microscopic features of parathyroid glands were distinguishable by CRM and correlated well with histopathology. The stromal fat content of normal and diseased glands could easily be determined. The percentage area occupied by fat cells differed significantly (P <.00001) in normal glands (average, 23.0% +/- 10.9%) and adenomatous glands (average, 0.4% +/- 0.7%). CONCLUSIONS: CRM imaging rapidly revealed microscopic features that reliably differentiated normal and diseased parathyroid glands. The success of this preliminary ex vivo study promotes interest in further development of an in situ probe for in vivo clinical diagnostic use. PMID- 11114648 TI - Invited commentary: can we reliably differentiate normal from abnormal parathyroid glands? (In the operating room? Eventually?) PMID- 11114649 TI - Ultrasound and sestamibi scan as the only preoperative imaging tests in reoperation for parathyroid adenomas. AB - BACKGROUND: In an effort to determine an efficient algorithm for the evaluation of patients with parathyroid adenomas in the reoperative setting, we explored the combination of using ultrasound scans (US) and sestamibi scintigraphy as the only preoperative imaging tests. METHODS: We analyzed the outcomes of 62 consecutive patients who were treated between January 1995 and May 1999 and who were referred for persistent primary hyperparathyroidism after initial surgical exploration, at which time no abnormal parathyroid glands had been found. Although all patients underwent US, computed tomography scan, magnetic resonance imaging, and sestamibi scan, we analyzed the success of localization and reoperation using only the results of US and sestamibi scan. RESULTS: Sixty-one patients (98%) underwent curative reoperations. The sensitivity, positive predictive value, and accuracy for US were 90%, 86%, and 84%, respectively; the corresponding values for sestamibi imaging were 78%, 94%, and 74%, respectively. In 58 of 62 cases (94%) preoperative US and/or sestamibi scan accurately identified the adenoma. In 3 patients for whom combined US and sestamibi scan were inaccurate, 1 adenoma was found by intraoperative US in the strap muscle; 1 adenoma was found by blind cervical thymectomy, and 1 adenoma was found by planned sternotomy that was based on computed tomography findings. CONCLUSIONS: This study supports an algorithm of obtaining US and sestamibi scan as the initial and perhaps only preoperative localization tests for patients with primary hyperparathyroidism after failed operation, at which time no abnormal glands had been found. PMID- 11114650 TI - A continuous membrane bioreactor for ester synthesis in organic media: I. Operational characterization and stability. AB - The feasibility of continuous ester synthesis in a membrane bioreactor (MBR) by a recombinant cutinase from Fusarium solani pisi was investigated, using the optimal conditions previously attained by medium engineering. The objective was to analyze the MBR behavior as a differential or an integral reactor. The main component of the reactor was an anisotropic ceramic membrane with 15,000 NMWCO. The operating variables included the influence of substrates ratio and flow rate on the conversion degree and on the productivity. The highest conversion degree was obtained using 1M of hexanol and 0.1M of butyl acetate as acyl donor. The use of these substrate concentrations led to a conversion degree of 79.3% and a specific productivity of 41 g hexyl acetate/(d x g cutinase), when the permeate flow rate was 0.025 mL/min. The increase of flow rate to 0.4 mL/min decreased the conversion to 35.6%, although the productivity was enhanced to 294 g product/day x g enzyme. The MBR characterization involved the calculations of mass balance, recirculation rate, conversion per pass, number of cycles, and hydraulic residence time. The operational stability was also evaluated in a longterm experiment over 900 hours and the enzyme half-life was estimated to be approximately 2 years. PMID- 11114651 TI - A continuous membrane bioreactor for ester synthesis in organic media: II. Modeling Of MBR continuous operation. AB - A model was developed to describe the conversion degree in a membrane bioreactor (MBR) for the synthesis of short-chain esters as a function of the flow rate. The transesterification reaction was catalyzed by a recombinant cutinase of Fusarium solani pisi microencapsulated in reversed micelles of AOT/isooctane. The differences of product concentration in permeate and retentate together with the deactivation profiles led to an enzyme distribution evaluation that describes the experimental values attained. The model considers the bioreactor design as well as its hydrodynamics and the enzyme kinetics. The approach included the analysis of the MBR operation as a CSTR, a PFR, and a series of continuous reactors. The comparative efficiency of these reactor types is discussed. The enzyme distribution was estimated for all the cases. The best description was obtained considering a series of two CSTRs. The modeling results led to a re-evaluation of cutinase operational stability. Deactivation rates correlated very well with the hydrodynamic aspects of biocatalyst location. PMID- 11114652 TI - Model-based estimation of myeloid hematopoietic progenitor cells in ex vivo cultures for cell and gene therapies. AB - Ex vivo production of hematopoietic progenitor cells has potential applications for cell therapy to alleviate cytopenias associated with chemotherapy and for gene therapy. In both therapies, progenitor and stem cells are considered crucial factors for therapeutic success. Assays for progenitor cells, however, take 2 weeks to complete, which is similar to the length of a typical culture. Therefore, a real-time estimation of the percentage or number of progenitor cells, based on rapid measurements, would be useful for optimization of feeding and harvest decisions. In this study, metabolic activity assays and flow cytometric analysis were used to estimate the content of progenitor cells. The measured metabolic activities are a collective contribution from all types of cells. Cells in granulomonocytic cultures have been lumped into six cell types and metabolic rates have been modeled as a linear function of cell composition and growth rate and as a nonlinear function of cell density. Data from 24 experiments were utilized to determine the model parameters in a calibration step. These data include flow cytometric analysis of more mature hematopoietic cells, progenitor cell colony assays, total cell content, and metabolite concentrations, and cover a wide range of cell composition, cell density, and growth rate. After calibration, the model is able to deliver good predictions of progenitor cell content for cultures with higher percentages of progenitor cells, as well as the peak progenitor cell content, based only on parameters that can be rapidly measured. With the aid of those predictions a harvest strategy was developed that will allow optimizing the harvest time based on the culture kinetics of each patient or donor inoculum, rather than using retrospective analysis to determine a uniform harvest time. PMID- 11114653 TI - Development of a novel bioreactor system for treatment of gaseous benzene. AB - A novel, continuous bioreactor system combining a bubble column (absorption section) and a two-phase bioreactor (degradation section) has been designed to treat a gas stream containing benzene. The bubble column contained hexadecane as an absorbent for benzene, and was systemically chosen considering physical, biological, environmental, operational, and economic factors. This solvent has infinite solubility for benzene and very low volatility. After absorbing benzene in the bubble column, the hexadecane served as the organic phase of the two-phase partitioning bioreactor, transferring benzene into the aqueous phase where it was degraded by Alcaligenes xylosoxidans Y234. The hexadecane was then continuously recirculated back to the absorber section for the removal of additional benzene. All mass transfer and biodegradation characteristics in this system were investigated prior to operation of the integrated unit, and these included: the mass transfer rate of benzene in the absorption column; the mass transfer rate of benzene from the organic phase into the aqueous phase in the two-phase bioreactor; the stripping rate of benzene out of the two-phase bioreactor, etc. All of these parameters were incorporated into model equations, which were used to investigate the effects of operating conditions on the performance of the system. Finally, two experiments were conducted to show the feasibility of this system. Based on an aqueous bioreactor volume of 1 L, when the inlet gas flow and gaseous benzene concentration were 120 L/h and 4.2 mg/L, respectively, the benzene removal efficiency was 75% at steady state. This process is believed to be very practical for the treatment of high concentrations of gaseous pollutants, and represents an alternative to the use of biofilters. PMID- 11114654 TI - Classification of stability behavior of bioreactors with wall attachment and substrate-inhibited kinetics. AB - The biodegradation of pollutants in continuous operation when the microbial population exhibits wall attachment is studied. The proposed model for wall attachment assumes two morphological forms of the microbial cell connected by metamorphosis reactions with first order exchange kinetics. An analysis of the stability of the bioreactor, carried out using elementary principles of the singularity theory and continuation techniques, allows for classification in the multidimensional parameter space of the various stability behaviors exhibited by the reactor model, for both substrate-inhibited and Monod kinetics. The analysis also shows the enhanced stability behavior of the bioreactor due to wall attachment. PMID- 11114655 TI - Soya lecithin effects on the aerobic biodegradation of polychlorinated biphenyls in an artificially contaminated soil. AB - The effects of the phytogenic surfactant soya lecithin (SL) on the aerobic biodegradation of polychlorinated biphenyls (PCBs) spiked into a synthetic soil were studied. Soil was spiked with both biphenyl (4 g/kg) and Fenclor 42 (1,000 mg/kg) and treated in aerobic batch slurry-phase microcosms (17.5% w/v). Microcosms were prepared either with or without the exogenous aerobic PCB dechlorinating bacterial co-culture ECO3 (inoculum:10(8) CFU/mL). In some inoculated microcosms, SL was added at 15 or 30 g/kg. Indigenous bacteria having the capability of metabolizing biphenyl and 2-chlorobenzoic acid were found to develop in the microcosms during the experiment, and were responsible for the significant PCB biodegradation and dechlorination observed in the uninoculated controls. The addition of ECO3 bacteria resulted in only a slight PCB biodegradation increase. In the presence of SL, a higher availability of biphenyl and chlorobenzoic acid-degrading bacteria and higher PCB biodegradation and dechlorination yields were observed; the effects increased proportionally with the concentration of the applied SL. A significant decrease of soil ecotoxicity was also revealed in SL-supplemented microcosms. At both concentrations, SL was found to be a good carbon source for both the indigenous and ECO3 bacteria, as well as a product capable of enhancing the PCB bioavailability in the microcosms. PMID- 11114656 TI - Energetics of growth and penicillin production in a high-producing strain of Penicillium chrysogenum. AB - The results of a large number of carbon-limited chemostat cultures of Penicillium chrysogenum carried out on glucose, ethanol, and acetate as the growth limiting substrate have been used to obtain an estimation of the adenosine triphosphate (ATP) costs for mycelium growth, penicillin production, and maintenance and the overall stoichiometry of oxidative phosphorylation of the fungus. It was found that penicillin production was accompanied by a significant additional energy drain (73 mol of ATP per mole of penicillin-G) from primary metabolism. This finding has been confirmed in independent experiments and has been shown to result in a significantly lower estimate for the maximum theoretical yield of penicillin-G on the carbon source. PMID- 11114657 TI - Microparticles of soy lecithin formed by supercritical processes. AB - Finely divided particles of phospholipids are used to form controlled drug delivery systems called liposomes. Conventional physicochemical methods for preparing these microparticles are hampered by a major drawback-the use of organic solvents that remain at few but inhibitory concentration in the final product. This study aimed to propose an alternative method for preparing microparticles of phospholipids starting from soy lecithin-the process had to be free of solvent or at least, the solvent had to be nontoxic. Two micronization techniques based on the use of supercritical carbon dioxide were investigated: the RESS and the SAS processes. The RESS process failed to separate the particles formed from the cosolvent. Performing the SAS process with ethanol as auxiliary solvent, enabled fine particles to form with size ranging from 1 to 40 microm. Particles were spherical and partly agglomerated and seemed to be free of solvent as shown by preliminary infrared analysis. PMID- 11114658 TI - Two-dimensional model of biofilm detachment caused by internal stress from liquid flow. AB - A two-dimensional model for biofilm growth and detachment was used to evaluate the effect of detachment on biofilm structures. The detachment process is considered to be due to internal stress created by moving liquid past the biofilm. This model generated a variety of realistic biofilm-formation patterns. It was possible to model in a unified way two different biofilm detachment processes, erosion (small-particle loss), and sloughing (large-biomass-particle removal). The distribution of the fraction from total biomass detached as a function of detached particle mass, gives indications about which of the two mechanisms is dominant. Model simulations indicate that erosion makes the biofilm surface smoother. Sloughing, in contrast, leads to an increased biofilm-surface roughness. Faster growing biofilms have a faster detachment rate than slow growing biofilms, under similar hydrodynamic conditions and biofilm strength. This is in perfect accordance with the experimental evidence showing that detachment is dependent on both shear- and microbial-growth rates. High growth rates trigger instability in biofilm accumulation and abrupt biomass loss (sloughing). Massive sloughing can be avoided by high liquid shear, combined with low biomass growth rates. As the modeling results show, the causes for sloughing must be sought not only in the biofilm strength, but also in its shape. Several "mushroom-like" biofilm structures like those repeatedly reported in the literature occurred, due to a combined effect of nutrient depletion and breaking at the colony base. A rough carrier surface promotes biofilm development in hydrodynamic conditions in which the biofilm on a flat surface would not form. Although biofilm patches filled completely the cavity in which they started to grow, they were unable to spill over the carrier peaks and to fully colonize the substratum. PMID- 11114659 TI - Temperature control in a continuously mixed bioreactor for solid-state fermentation. AB - A continuously mixed, aseptic paddle mixer was used successfully for solid-state fermentation (SSF) with Aspergillus oryzae on whole wheat kernels. Continuous mixing improved temperature control and prevented inhomogeneities in the bed. Respiration rates found in this system were comparable to those in small, isothermal, unmixed beds, which showed that continuous mixing did not cause serious damage to the fungus or the wheat kernels. Continuous mixing improves heat transport to the bioreactor wall, which reduces the need for evaporative cooling and thus may help to prevent the desiccation problems that hamper large scale SSF. However, scale-up calculations for the paddle mixer indicated that wall cooling becomes insufficient at the 2-m(3) scale for a rapidly growing fungus like Aspergillus oryzae. Consequently, evaporative cooling will remain important in large-scale mixed systems. Experiments showed that water addition will be necessary when evaporative cooling is applied in order to maintain a sufficiently high water activity of the solid substrate. Mixing is necessary to ensure homogeneous water addition in SSF. Automated process control might be achieved using the enthalpy balance. The enthalpy balance for the case of evaporative cooling in the paddle mixer was validated. This work shows that continuous mixing provides promising possibilities for simultaneous control of temperature and moisture content in solid-state fermentation on a large scale. PMID- 11114660 TI - Model for on-line moisture-content control during solid-state fermentation. AB - In this study we describe a model that estimates the extracellular (nonfungal) and overall water contents of wheat grains during solid-state fermentation (SSF) with Aspergillus oryzae, using on-line measurements of oxygen, carbon dioxide, and water vapor in the gas phase. The model uses elemental balances to predict substrate dry matter losses from carbon dioxide measurements, and metabolic water production, water used in starch hydrolysis, and water incorporated in new biomass from oxygen measurements. Water losses caused by evaporation were calculated from water vapor measurements. Model parameters were determined using an experimental membrane-based model system, which mimicked the growth of A. oryzae on the wheat grains and permitted direct measurement of the fungal biomass dry weight and wet weight. The measured water content of the biomass depended heavily on the moisture content of the solid substrate and was significantly lower than the estimated values reported in the literature. The model accurately predicted the measured overall water content of fermenting solid substrate during fermentations performed in a 1.5-L scraped drum reactor and in a 35-L horizontal paddle mixer, and is therefore considered validated. The model can be used to calculate the water addition required to control the extracellular water content in a mixed solid-state bioreactor for cultivation of A. oryzae on wheat. PMID- 11114661 TI - Development of small-size tubular-flow continuous reactors for the analysis of operational stability of enzymes in low-water systems. AB - A very small-scale continuous flow reactor has been designed for use with enzymes in organic media, particularly for operational stability studies. It is constructed from fairly inexpensive components, and typically uses 5 mg of catalyst and flow rates of 1 to 5 mL/h, so only small quantities of feedstock need to be handled. The design allows control of the thermodynamic water activity of the feed, and works with temperatures up to at least 80 degrees C. The reactor has been operated with both nonpolar (octane) and polar (4-methyl-pentan-2-one) solvents, and with the more viscous solvent-free reactant mixture. It has been applied to studies of the operational stability of lipases from Chromobacterium viscosum (lyophilized powder or polypropylene-adsorbed) and Rhizomucor miehei (Lipozyme) in different experimental conditions. Transesterification of geraniol and ethylcaproate has been adopted as a model transformation. PMID- 11114662 TI - Enzyme immobilization in silica-hardened organogels. AB - In this study we describe a novel method for immobilizing enzymes in a solid nanocomposite matrix based on gelatin gels, which are subsequently hardened by in situ polymerization of tetraethoxysilane (TEOS). Chromobacterium viscosum lipase is taken as the example. This immobilization method possesses the advantages of enzyme entrapment in microemulsions, together with newly beneficial qualities, such as transparency, which permits direct spectroscopic investigation, and considerable mechanical stability in both aqueous and organic solvents, which results in the maintenance of enzymatic activity for several months. The first step is enzyme solubilization in AOT reverse micelles, followed by transformation of this solution into an organogel by the addition of gelatin. The enzyme containing gel, is then hardened by the formation of silicate polymer. A glassy nanocomposite is obtained, which is optically transparent, so that the protein can be studied directly spectroscopically. Circular dichroic spectra of cytochrome-c are shown as an example. The nanocomposite material can be dried and ground, yielding a powder that is stable in both aqueous and organic solvents. After extensive washing with water, the enzyme-containing nanocomposite showed good activity in cyclohexane. The synthesis of water-insoluble fatty acid esters was carried out in this solvent with yields close to 90%. In this case, the enzyme preparations can be used over a period of several months without loss of activity or chemical yield. PMID- 11114663 TI - Computer-based registration for digital subtraction in dental radiology. AB - OBJECTIVES: (1) To review computerized a posteriori techniques for geometry and contrast registration prior to digital subtraction in dental radiography; (2) to define a uniform notation for their methodological and technical classification and based on this key code; (3) to derive criteria for successful application of computer-based a posteriori registration for routine clinical subtraction. METHODS: All techniques are classified with respect to the (1) dimension of geometry registration; (2) origin; (3) abstraction level, and (4) linkage of features used for registration of geometry; (5) elasticity; (6) domain, and (7) parameter determination of the geometrical transform used; (8) interaction of geometrical registration; as well as (9) origin of features, (10) model of transform, and (11) interaction of procedure for contrast correction. RESULTS: With respect to clinical practicability, superior registration techniques are based on the low level abstraction of intrinsic features for both geometry and contrast registration. By approximately linking the features, a global projective transform should be generated for geometry registration by automatic methods, while automatic contrast correction should be non-parametric. This challenge is met only by one out of 36 published algorithms. Hence, although numerous computer based techniques have been published, only a few of them are applied more than once in practice. CONCLUSION: The key code proposed in this paper is useful for technical classification of a posteriori registration methods in dental radiography and allows their objective comparison. Further investigations will focus on standardization of practicable procedures to evaluate the robustness of competing methods. PMID- 11114664 TI - A comparative study of the radiological diagnosis of postoperative maxillary cyst. AB - OBJECTIVES: To compare the diagnostic accuracy for the postoperative maxillary cyst (POMC) of panoramic in combination with Waters' radiography with computed tomography (CT) and of oral and maxillofacial radiologists with non-specialists. STUDY DESIGN: Nineteen cases of POMC and 19 of postoperative changes were assessed using panoramic in combination with Waters' radiographs and CT by five oral and maxillofacial radiologists and five non-specialists on a five-point scale. The areas under the ROC curves were analysed using the Wilcoxon rank sum test to determine any differences in diagnostic accuracy between the two methods and between the two groups. RESULTS: The diagnostic accuracy of CT was higher than that of combined panoramic and Waters' radiographs for the oral and maxillofacial radiologists (P < 0.05), but not for the non-specialists (P > 0.05). The diagnostic accuracy of the oral and maxillofacial radiologists for each method was higher than that of the non-specialists group (P < 0.05). CONCLUSIONS: CT improves the evaluation of POMC. Radiological training and experience leads to more accurate diagnoses. PMID- 11114665 TI - Magnetic resonance imaging of temporomandibular disorders: classification, prevalence and interpretation of disc displacement and deformation. AB - AIM: To analyse the prevalence of disc displacements and deformations from MRI of symptomatic temporomandibular disorders (TMD). METHODS: The study was conducted retrospectively on 192 joints of 98 patients (67 females, 31 males, mean age 29 years), who had undergone bilateral MRI (except for four who had unilateral) in the sagittal (both open and closed mouth) and coronal (closed mouth only) planes. These displacements were subdivided into static (complete anterior and posterior, partial anterolateral and anteromedial, sideways lateral and medial, anterolateral and anteromedial rotational) and dynamic (with reduction, without reduction, with incomplete reduction; non-determinable). Disc deformations were subdivided into: enlargement of the posterior band, reversed biconcave shape, biplanar (flattened) and biconvex shape. RESULTS: Eighty per cent of patients had bilateral displacement, 15% unilateral and 5% a normally positioned disc. Complete anterior displacement was the commonest and sideways the rarest. Reduction was present in 58% of disc displacements, no reduction in 26%, incomplete reduction in 4%, while in the remaining 12%, it could not be determined. Rotational displacement was the most likely to feature reduction and sideways the least. Temporomandibular joints with no reduction were closely correlated with bone lesions. The most frequent deformation was biplanar and the rarest enlargement of the posterior band. CONCLUSIONS: There was a high prevalence of displacements and deformations. While they do not appear to be significant in inducing pain, they can predispose to the onset of osteoarthrosis. PMID- 11114666 TI - Intra-oral storage phosphor and conventional radiography in the assessment of alveolar bone structures. AB - OBJECTIVES: To compare storage phosphor (SP) with conventional film radiography for accuracy of linear measurements of the marginal alveolar bone and visibility of anatomical structures. METHODS: Linear measurements were made in paired SP and conventional images of dried human mandibles with a metal pin fixed 10 mm below the alveolar crest. One observer measured the distance from the alveolar crest to the reference point on the radiographs. The difference between the measured and the true distance was calculated. Two observers rated the visibility of bony structures (periodontal ligament space, periapical bone tissue, alveolar crest) in 51 paired digital and conventional images of 21 patients on a 3-point scale. Overall agreement and Kappa index were calculated. RESULTS: Accuracy of linear measurements was higher in digital radiography (mean difference 0.17 mm) than in conventional radiography (mean difference 0.59 mm). Overall, the two observers rated visibility higher in conventional radiographs. The Kappa indices for the periodontal ligament space and periapical bone indicated fair to almost perfect agreement (kappa = 0.38 and 0.5; kappa = 0.39 and 0.84) while for the alveolar crest there was only poor or moderate agreement (kappa = 0.2 and 0.5). CONCLUSIONS: The small differences in linear measurements indicate that the Digora system is suitable for clinical assessment of periodontal and peri-implant bone loss. The visibility of dental structures depends as much on the individual features assessed, as the radiographic system. PMID- 11114667 TI - A new method for the radiological investigation of residual ridge resorption in the maxilla. AB - OBJECTIVES: To develop a new method for assessing residual ridge resorption in the edentulous maxilla. METHODS: Defined experimental and reference areas in the maxilla were drawn on transparent film laid over a panoramic radiograph and digitized. Bone areas were measured with an integrated planimetry program and expressed as a ratio R. The effect of positioning errors on reliability of the method was investigated on dry skulls. The correlation between the change in ratio and actual bone loss was examined by progressively reducing the height of an artificial residual ridge on one skull. RESULTS: The coefficient of variation for the absolute ratio in different head positions was < 0.05 and its correlation coefficient of the change in R and the degree of resorption was r2 > or = 98.3%, P = 0.0001. CONCLUSIONS: Comparison of the experimental area with the reference area on serial panoramic radiographs appears suitable for the assessment of residual resorption in the maxilla. PMID- 11114668 TI - Survey of radiographic practices for periodontal disease in UK and Irish dental teaching hospitals. AB - OBJECTIVES: To assess current radiographic practices in dental teaching hospitals for the management of patients with periodontal diseases. METHODS: All 17 dental teaching hospitals in UK and Ireland were sent a questionnaire on radiographic equipment and radiograph selection currently used for assessment of patients with destructive periodontal diseases. Opinions were recorded for advantages and disadvantages of the most frequently used radiographic views. RESULTS: A 100% response rate was achieved. All hospitals used panoramic and specific periapical radiographs as one of their radiographic regimes for patients with periodontal disease. Fifty-three per cent of respondents most frequently took panoramic and selected periapical radiographs. Twenty-four per cent took full mouth periapical radiographs (FMPAs) most frequently and 18% took a panoramic radiograph alone. Twenty-four per cent of hospitals operated a protocol for selection of radiographs for periodontal patients. CONCLUSIONS: The most commonly used views taken to assess periodontal status are panoramic radiographs with selected periapicals. Few hospitals operate a protocol for prescribing radiographs. PMID- 11114669 TI - Two cases of polyostotic eosinophilic granuloma. AB - We report two cases of polyostotic eosinophilic granuloma (EG). Both plain radiographs and CT showed diffuse osteolytic lesions which suggested malignant tumors. Although EG was polyostotic, the prognosis was relatively good. Both cases developed new lesions over a follow-up period of 4-5 years and therefore further long-term review is needed. PMID- 11114670 TI - Selected Abstracts from the 7th European Congress of Dentomaxillofacial Radiology Athens, Greece. June 15 - 18 2000. PMID- 11114672 TI - Keyword Index to Volume 29. PMID- 11114673 TI - Influence of environmental and nutritional factors on salivary gland tumorigenesis with a special reference to dietary lipids. AB - Salivary gland cancer is a rare condition whose incidence varies according to different geographical regions. Several environmental factors, such as ionizing radiation and some occupational aspects, as well as habits like smoking and alcohol consumption, are related to salivary tumorigenesis. Both acinar and ductal cells may be involved in the origin of salivary gland tumours. Even though laboratory and epidemiological evidence indicates that diet and nutritional habits may modulate the tumorigenesis at different sites, little is known about this effect on salivary glands, mainly in regard to dietary lipids. However, the fact that monounsaturated fatty acids behave as protumorigenic and, on the contrary, certain polyunsaturated fatty acids exert beneficial effects, demonstrated on breast, colon and even oral cancer, gives support to our hypothesis. The suggested relationship between environmental and nutritional factors, mainly dietary lipids, and salivary gland cancer constitutes the aim of the present work. PMID- 11114674 TI - Measuring diet quality in china: the INFH-UNC-CH diet quality index. AB - OBJECTIVE: This paper describes the development and efficacy of a diet quality index (DQI) for China. DESIGN: The Dietary Guidelines for Chinese Residents motivated the selection of 10 DQI components. These components were weighted and assigned cut-offs and point schemes based on the Chinese Food Guide Pagoda, Chinese and/or international dietary reference values. The efficacy of resulting DQI scores was assessed relative to a priori expectations. SUBJECTS: The Chinese DQI was evaluated using cross-sectional 3 day diet record and anthropometric data on 7450 adults from the 1991 China Health and Nutrition Survey. METHODS: For each individual, a DQI total score was calculated as the sum of components, and DQI pattern scores calculated to reflect the underlying composition of diet quality problems. The DQI scores were compared with component scores, food and nutrient intake, weight status and sociodemographic variables. RESULTS: The total DQI score simultaneously represented all component aspects of diet quality as well as micronutrients not explicitly built into the index. The total DQI score was significantly correlated with food and nutrient intakes, BMI, urban residence and income. The DQI pattern scores correlated with DQI components and weight status as expected. CONCLUSIONS: The China DQI captures variation along several components of diet quality, appears sensitive to under- and overnutrition, as well as sociodemographic variables. The China DQI may prove useful for monitoring the nutrition transition and epidemiologic trends in China. SPONSORSHIP: National Institutes of Health (HD 38700 and R01-HD30880) and the Chinese Academy of Preventive Medicine. PMID- 11114676 TI - Frequency of fruit and vegetable consumption and blood antioxidants in the Caerphilly cohort of older men. AB - OBJECTIVE: To assess the number of portions of fruit and vegetables consumed daily by a large representative sample of older men, and to determine how blood antioxidant (vitamins E, A and carotenoids) concentrations vary with fruit and vegetable consumption. DESIGN: Cross-sectional study of free-living men. SUBJECTS: Men aged 55-69 y (dietary data, n=1957; blood data, n=1874) participating in Phase III (1989-1993) of the Caerphilly and Speedwell Collaborative Heart Disease Studies. METHODS: Dietary data were obtained by semi quantitative food-frequency questionnaire and blood samples were analysed for antioxidant vitamins. Men were subdivided into groups on the basis of portions per day of fruit and vegetables. Within these sub-groups, mean and 95% ranges of intakes and of blood antioxidant levels were obtained. Log transformations were performed where appropriate. RESULTS: Only 4.3% of the men met the recommended target of five portions, while 33.3% of the men consumed one or fewer portions of fruit and vegetables per day. Those men who consumed the poorest diets with respect to fruit and vegetable intakes were more likely to be from lower socio economic classes, drink more alcohol and be current smokers. Fruit and vegetable intake reflected plasma concentrations of antioxidants, which showed a dose response relationship to frequency of consumption. CONCLUSIONS: Older men in the UK consume much less fruit and vegetables than current recommendations. Major difficulties are likely to be encountered in trying to meet a dietary target that is clearly much higher than the fruit and vegetable consumption of large sections of the older population in the UK. SPONSORSHIP: This work was supported by the Medical Research Council. PMID- 11114675 TI - The potential of various foods to serve as a carrier for micronutrient fortification, data from remote areas in Indonesia. AB - OBJECTIVE: To estimate the potential of various industrially produced foods, to serve as a carrier for micronutrient fortification based on the frequency of their consumption in different socio-economic strata; to determine the role of fortified instant noodles as a source of micronutrients; to assess the contribution of plant foods, animal foods and fortified foods to vitamin A intake. SETTING: A survey was conducted in rural South Sulawesi and urban South Kalimantan between November 1996 and January 1997. SUBJECTS: Households (1500 in South Sulawesi; 2112 in South Kalimantan) were selected randomly by multi-stage cluster sampling. From each household, data were collected from the mother and her youngest child (0-5 y). DATA COLLECTION: Mothers were interviewed on various topics, including socio-economic status, food consumption, receipt of high-dose vitamin A capsules, health and nutritional status. RESULTS: Monosodium glutamate and salt were consumed daily in almost all households in both areas, and consumption was not associated with socio-economic status. Instant noodles were consumed in nearly all households in both areas, but consumption of fortified noodles was related to socio-economic status; it was highest among households of government employees and private investors, and lowest among farmers and share croppers. Vegetables were the most important source of vitamin A in rural South Sulawesi, while foods of animal origin were the most important source in urban South Kalimantan. CONCLUSIONS: The results support double or triple fortification of salt and/or monosodium glutamate with iodine, vitamin A and/or iron. Efforts to overcome associated technical and logistical difficulties are urgently needed. SPONSORSHIP: Opportunities for Micronutrient Interventions (OMNI); United States Agency for International Development (USAID). European Journal of Clinical Nutrition (2000) 54, 822-827 PMID- 11114677 TI - Prevalence of anemia in elderly subjects living at home: role of micronutrient deficiency and inflammation. AB - OBJECTIVE: Aging is associated with increased risk of developing anemia and micronutrient deficiencies. Wheat-based staple foods are iron fortified in Chile. We aimed to establish the prevalence and etiology of anemia in apparently healthy free-living elderly subjects. DESIGN AND SETTING: A cross-sectional study was performed in an outpatient clinic of Santiago, Chile. SUBJECTS AND METHODS: A group of 274 subjects (93 men, 181 women)>/=60 y old living at home and apparently healthy was studied. Clinical and anthropometric evaluations and dietary survey were performed. Complete blood count, and status of iron, copper, folate, vitamins B12 and A and C-reactive protein, and erythrocyte sedimentation rate were measured. RESULTS: Prevalence of anemia was 5.4% for men and 4.4% for women. Subjects with inflammatory process had a higher prevalence of anemia (22.2% men, 31.6% women). Abnormal serum retinol (<0.35 micromol/l) was found in 13.7% of men and 15.9% of women. Prevalence of folate deficiency (<7 nmol/l) was 50.5% in men and 33.1% in women. Low serum vitamin B12 (<148 pmol/l) was present in 51.1% of men and 30. 9% of women. Iron and copper deficiencies were infrequent. CONCLUSIONS: Anemia is not prevalent in free-living elderly subjects when iron intake is adequate. Inflammatory process is the main etiology of anemia in this age group. Vitamin A, folate and vitamin B12 deficiencies were found in a significant proportion of the study group. SPONSORSHIP: Sandoz Foundation for Gerontological Research. PMID- 11114678 TI - Alpha linolenic acid in cholesterol esters: a marker of alphalinolenic acid intake in newborns. AB - OBJECTIVE: To evaluate alpha-linolenic acid (ALA) (18∶3 n-3) and linolenic acid (LA) (18∶2 n-6) in cholesterol esters (CE) as markers of ALA and LA dietary intakes in preterm infants. SUBJECTS: Forty-five preterm infants: two groups fed different formulas, the third fed human milk. DESIGN: ALA and LA dietary intakes were precisely recorded in each infant to accurately determine the cumulative amount of ingested ALA and LA during two intervals: (i) between the second day after the first significant formula intake (D0) and the fifteenth day (D15); and (ii) between D0 and the first day of the 37th week of post conception age (W37). The corresponding amounts of ingested ALA and LA were related to ALA and LA levels determined by capillary column gas-liquid chromatography in plasma cholesterol esters at D15 and W37, respectively. RESULTS: ALA in CE was very significantly correlated to D0-D15 and D0-W37 ALA intakes (0.66; P=0.0001 and 0.70; P=0.0001), respectively. LA in CE was weakly correlated to D0-D15 LA intakes (0.03; P=0.01) and whatever the group (human milk or enriched formula) the correlation was lost at W37. CONCLUSION: In preterm infants, ALA in CE can be considered as representative of ALA dietary intakes, whereas LA in CE appears as a poor marker of LA intakes. PMID- 11114679 TI - Does breast cancer change patients' dietary habits? AB - PURPOSE: The results of epidemiological studies on diet and cancer are often difficult to interpret on an individual level and may influence patients' beliefs, attitudes and behaviour. This study investigated the behaviour of breast cancer patients and their attitudes to dietary changes and the need of dietary advice during their disease. PATIENTS AND METHODS: The study population consisted of breast cancer patients visiting the Department of Oncology in Turku University Hospital for treatment or follow-up in August and September 1999. A questionnaire was given to a total of 123 subjects. RESULTS: The majority, 65%, were attending the clinic for treatment, 35% for follow-up. Ninety-seven patients (86%) consumed a normal Finnish diet, six (5.3%) were vegetarians and 10 (8.1%) vegetarians consuming fish and chicken occasionally. Eleven patients (8.9%) considered diet a factor contributing to their breast cancer and 38 (31.9%) had changed their dietary habits after the diagnosis of breast cancer. The numbers were higher in younger patients with higher educational background. The main reason for change in diet was the desire to be cured of cancer (52.9% of those patients who had changed their dietary habits), in 11.8% to alleviate the symptoms of nausea and 11.8% were advised by health care professionals. The main changes reported included a reduction in the consumption of animal fat, sugar and red meat and increased consumption of fruit, berries and vegetables. Forty-nine patients (39.8%) used vitamin and mineral supplements and 27 (21.9%) consumed dietary supplements including natural products and probiotics. The source of information on how to change the diet was for 33.3% the mass media, 19.4% were advised by doctors and nurses and 11.1% by dietitians. One-third of the patients expressed a need for more information on dietary factors. CONCLUSION: Breast cancer patients' need of control over their own life prompts an interest in alternative dietary habits after diagnosis, the focus being on a healthier diet. Expert dietary information is considered important. Many patients mentioned a lack of precise dietary recommendations for their individual disease situation and depended on information from outside their treatment centre. PMID- 11114680 TI - Enhancing immunity by dietary consumption of a probiotic lactic acid bacterium (Bifidobacterium lactis HN019): optimization and definition of cellular immune responses. AB - OBJECTIVE: To define the cellular basis for immune enhancement by a probiotic lactic acid bacteria strain (Bifidobacterium lactis HN019); and to determine whether immune enhancement can be optimized by delivery in oligosaccharide enriched low-fat milk. DESIGN: A double-blind, three-stage before-and-after intervention trial. SETTING: Taipei Medical College Hospital, Taipei, Taiwan. SUBJECTS: Fifty healthy Taiwanese citizens (age range 41-81; median 60) randomly allocated to two groups. INTERVENTIONS: In stage 1 (run-in control stage) all subjects consumed reconstituted low-fat milk (LFM) for 3 weeks; in stage 2 (probiotic intervention) subjects consumed B. lactis in LFM (group A) or B. lactis in lactose-hydrolysed LFM (group B) for 3 weeks; in stage 3 all subjects returned to non-supplemented LFM for a further 3 weeks (washout stage). The innate immune functions of two different leucocyte types (polymorphonuclear (PMN) cells and natural killer (NK) cells) were assessed at four time points via in vitro analyses on peripheral blood samples. RESULTS: While consumption of LFM alone had no significant effect on immune responses, stage 2 results indicated significantly enhanced PMN cell phagocytosis and NK cell tumour killing activity following consumption of milk containing B. lactis. These increases levelled off following cessation of B. lactis consumption, but remained above the pre treatment values. Increases in PMN and NK cell activity were greatest among subjects who consumed B. lactis in lactose-hydrolysed LFM. CONCLUSIONS: Dietary consumption of the probiotic bacterium B. lactis HN019 enhanced immune function of two different types of leucocytes; the degree of enhancement was increased by consuming B. lactis in an oligosaccharide-rich substrate. SPONSORSHIP: Financial support was provided by the New Zealand Dairy Board. PMID- 11114681 TI - The effect of a probiotic milk product on plasma cholesterol: a meta-analysis of short-term intervention studies. AB - INTRODUCTION: Certain fermented dairy milk products may have beneficial effects on plasma cholesterol levels. However, a number of studies have produced conflicting results as to whether dietary supplementation by a probiotic dairy product containing the bacteria culture Causido(R) reduces plasma cholesterol. OBJECTIVE: To conduct a meta-analysis of intervention studies to evaluate the effect of the Causido(R) culture on plasma total cholesterol and low-density lipoprotein (LDL)-cholesterol. THE PROBIOTIC MILK PRODUCT: The yoghurt product Gaio(R) is fermented with Causido(R), composed of one strain of Enterococcus faecium (human species) with the proposed cholesterol-lowering effect, and two strains of Streptococcus thermophilus. STUDY INCLUSION AND DATA EXTRACTION: Six studies were identified from a literature search and from the yoghurt producer. All studies met the inclusion criteria. Summary data for plasma concentrations of total cholesterol and LDL-cholesterol were extracted from the original publications or by personal request to the authors. Data from 4-8 weeks of treatment duration was used. STATISTICAL ANALYSIS: We performed a traditional meta-analysis where mean differences between intervention and control of the pre post changes in total cholesterol and LDL-cholesterol were calculated, as well as 95% confidence intervals (CIs). RESULTS: In the six studies included in the meta analysis, the Gaio(R) interventions produced changes in total cholesterol above those of the control groups ranging from -0.02 to -1.02 mmol/l and in LDL cholesterol ranging from -0.02 to -1.15 mmol/l. After inclusion of an open-label study, the meta-analysis of the double-blind studies showed that Gaio(R) as compared to the control group changed total cholesterol by -0.22 mmol/l (95% CI: 0.35 to -0.08, P<0.01) and LDL-cholesterol by -0.20 mmol/l (95% CI: -0.33 to 0.06, P<0.005). The outcome was essentially the same if all studies were included. CONCLUSIONS: The present meta-analysis of controlled short-term intervention studies shows that the fermented yoghurt product produced a 4% decrease in total cholesterol and a 5% decrease in LDL-cholesterol when the open label study is excluded. To demonstrate sustained effects on blood lipids, long term studies are required. SPONSORSHIP: MD Foods A/S, Denmark. PMID- 11114682 TI - A definition for dietary fibre. AB - The objective of this paper is to present a definition for dietary fibre based on recent advances that have taken place not only in human nutrition but also in plant cell-wall science and animal nutrition. We propose a physiologically based framework definition but, recognizing the diversity of dietary fibre, we have proposed further classifications within this framework. We also suggest that dietary fibre be removed from the carbohydrate group of nutrients because some compounds defined as dietary fibre are not chemically carbohydrates. The definition and classification system clearly highlight areas where further studies are needed. PMID- 11114683 TI - Associations of alpha-linolenic acid and linoleic acid with risk factors for coronary heart disease. AB - BACKGROUND: Prevention of coronary heart disease (CHD) in high-risk subjects. OBJECTIVE: To investigate the associations of dietary intake of alpha-linolenic acid (ALA) and linoleic acid (LA) as assessed by food frequency questionnaire and in the plasma cholesteryl ester (CE), with CHD risk factors. DESIGN: Baseline data of a double-blind, randomized placebo-controlled trial. Subjects have hypercholesterolemia (6.0-8.0 mmol/l) and at least two other CHD risk factors (n=266). RESULTS: The reported dietary ALA and LA intakes and the LA/ALA ratio were associated with the contents in the CE (r=0.37, r=0.21, and r=0.42, respectively; P<0.01). In multivariate analysis, CE ALA was inversely associated with diastolic blood pressure (r=-0.13; P<0.05) and positively with serum triacylglycerol (r=0.13; P<0.05), and CE LA was inversely associated with serum triacylglycerol (r=-0.32; P<0.01). The CE LA/ALA ratio was strongly inversely associated with CE ALA (r=-0.95; P<0.01). In the lowest quintile of CE ALA, mean dietary intake was 0.4 energy % ALA (1.2 g/day), 8.4 energy % LA and an LA/ALA ratio of 21, and in the highest quintile 0.6 energy % ALA (1.7 g/day), 6.8 energy % LA and 12 (ratio). In the lowest quintile of CE ALA the diastolic blood pressure was 4 mm Hg lower (P trend<0.05), and the serum triacylglycerol 0.3 mmol/l higher (P trend NS) when compared with the top quintile. CONCLUSIONS: In a CHD high-risk population with LA-rich background diet, these cross-sectional data suggest that replacing LA in the diet by ALA may decrease diastolic blood pressure, and may increase serum triacylglycerol concentration. PMID- 11114684 TI - Assessment of iron status using plasma transferrin receptor in pregnant women with and without human immunodeficiency virus infection in Malawi. AB - BACKGROUND: Although anemia is highly prevalent during pregnancy and is common during human immunodeficiency virus (HIV) infection, anemia and iron status have not been well characterized in HIV-infected pregnant women. OBJECTIVE: To gain insight into iron status in HIV-infected pregnant women using plasma transferrin receptor and related indicators of anemia. STUDY DESIGN: Plasma transferrin receptor, ferritin, alpha1-acid glycoprotein, C-reactive protein and hemoglobin concentrations were measured in pregnant women, gestational age 18-28 weeks, seen in an urban antenatal clinic in Blantyre, Malawi. RESULTS: The prevalence of anemia among 662 HIV-positive and 190 HIV-negative pregnant women was 73.1% and 50.0%, respectively (P<0.0001). Among HIV-positive and HIV-negative women, median plasma transferrin receptor concentrations were 24.4 and 24.1 nmol/l (P=0.5), respectively, and median plasma ferritin concentrations were 17.8 and 20.8 microg/l (P<0.05), respectively. There was a large overlap in plasma transferrin receptor concentrations among women with and without anemia. Using the combination of hemoglobin and ferritin as a standard, the sensitivity and specificity of plasma transferrin receptor in diagnosing iron deficiency anemia was estimated at 45.9% and 68.1%, respectively. CONCLUSION: The use of plasma transferrin receptor concentrations as an indicator of iron deficiency anemia may be limited in pregnant women with chronic inflammation and infection. PMID- 11114685 TI - Triacylglycerol composition in colostrum, transitional and mature human milk. AB - OBJECTIVE: Milk triglycerides from colostrum, transitional and mature human milk, were analyzed and compared in order to determine the differences in triacylglycerol composition throughout lactation. SETTING: Department of Food and Nutrition, University of Barcelona, Spain, and Neonatology Department of the University Hospital of Granada, Spain. SUBJECTS: Twenty-two healthy lactating women aged 21-35. DESIGN AND INTERVENTIONS: The triacylglycerol profiles of 47 breast milk samples including colostrum (1-3 days), transitional milk (7-10 days) and mature milk (25-60 days) were analyzed by high-performance liquid chromatography (HPLC), with light-scattering detection (LSD). RESULTS: Significant differences regarding several triglycerides were found between three milk classes when the Kruskal-Wallis nonparametric test was applied to 47 human milk samples that had been compared using the complete chromatographic triacylglycerol profile. The ANOVAS for each equivalent carbon number (ECN) group of triglycerides revealed significant differences between colostrum, transitional milk and mature milk. By the discriminant analysis of triacylglycerol percentages, in 19 colostrum samples, 14 transitional milk samples and 14 mature milk samples, three milk types were distinguished, and three triglycerides (peak no. 4, LnOO and SOO) were found to be the most predictive variables over all the triacylglycerol profile or ECN groups. CONCLUSIONS: Each state of lactation shows a specific profile of triacylglycerol composition in human milk. However the two most abundant triacylglycerides in colostrum, POO and POL, which account for more than 49% of the total, are also dominant in transitional (34%) and mature milk (42%). PMID- 11114686 TI - A novel infant formula milk with added long-chain polyunsaturated fatty acids from single-cell sources: a study of growth, satisfaction and health. AB - OBJECTIVE: To evaluate the growth, feeding and health of babies fed a novel infant formula milk with added long chain polyunsaturated fatty acids (LCPs) produced from single-cell sources, at concentrations similar to those found in mature breast milk. DESIGN: Randomized double-blind control trial. SUBJECTS: One hundred and forty healthy, full-term infants of birth weight 2.5-4.5 kg born at the Maternity Department, East Glamorgan General Hospital, Wales, whose mothers had already decided to bottle feed. INTERVENTIONS: Subjects were randomized to two groups; one (control group) to receive a standard formula milk and the other to receive the trial milk with added LCPs. Milks were supplied in a double-blind fashion and given for the first 12 weeks of life. Anthropometric measurements were made at recruitment, 3 months, 6 months and 1 y. Feeding diaries were completed at 6 weeks and 3 months, and a parental record was kept of any ill health suffered by the subjects during the first year of life. RESULTS: Of 140 infants recruited, 31 did not complete the protocol. Small but statistically significant differences were found only in the subscapular skinfold thickness at 6 weeks and 3 months, that in the trial group being slightly higher, but unlikely to be of any clinical importance. No differences were found in the feeding patterns of the infants in the two groups. Stool patterns were similar, as were the frequencies of illness and allergy. CONCLUSIONS: This study supports the view that LCPs from single-cell sources do not have detrimental effects on the growth, feeding and general health of infants. PMID- 11114687 TI - Physical activity patterns in normal, overweight and obese individuals using minute-by-minute accelerometry. AB - OBJECTIVE: To determine the levels and patterns of daily physical activity in groups of normal-weight, overweight and obese adults using uniaxial minute-by minute accelerometry. DESIGN: Cross-sectional study of physical activity levels over a 7 day period using accelerometers programmed to collect minute-by-minute data. SETTING: Participants were recruited from large companies in Bristol and Cardiff. All meetings took place on company premises. PARTICIPANTS: One-hundred and eight participants volunteered for the study. Eighty-four (36 males, 48 females; 38.6+/-9.3 y) (mean+/-s.d. ) met the inclusion criteria for accelerometer measurements and were included in analyses. RESULTS: No significant differences in physical activity were identified between normal-weight (BMI<25) and overweight (BMI 25-29.9) participants. Obese participants (BMI>30) were significantly less active than non-obese participants (BMI< or =30) during weekdays (279.1+/-77.5 vs. 391.3+/-139.4 counts min(-1); P<0.001), weekends (222.3+/-93.9 vs. 386.2+/-177.5 counts min(-1); P<0.001) and evenings (221.1+/ 126.3 vs. 380.8+/-220.7 counts min(-1); P=0.002), but not at work (307.5+/-87.2 vs 398.7+/-163.3 counts min(-1); P=0.06). Differences in activity levels between obese and non-obese participants were greater in males than females. Hour-by-hour physical activity patterns demonstrated that obese participants were less active than the non-obese for almost every hour of the weekdays and the weekend. CONCLUSIONS: Although no differences in activity emerged between overweight and normal-weight individuals, obese participants were substantially less active than the non-obese, particularly when there was a free choice of activity or no activity. The difference in activity levels was particularly pronounced in males, and unless obese individuals are compensating by lower energy intake, these activity patterns will contribute to the maintenance of or increase in the degree of obesity. Minute-by-minute accelerometry is a valuable tool for the assessment of activity patterns and may be useful to highlight periods of inactivity for intervention design. PMID- 11114688 TI - Differences in resting energy expenditure between black and white smokers: implications for postcessation weight gain. AB - OBJECTIVE: To examine differences in resting energy expenditure (REE) between black and white smokers in order to determine whether REE might contribute to postcessation weight gain. DESIGN: Cross-sectional and prospective investigation of ethnic differences in REE. Differences in REE between black and white smokers were examined at baseline while all participants were smoking, and again during 2 weeks of abstinence from smoking. SETTING: Memphis, Tennessee, USA. SUBJECTS: Sixty-six black and 112 white smokers (age 30.4 y; cigarettes per day 21.4; weight 71.7 kg; body mass index 24.5 kg/m2). RESULTS: Black smokers had a significantly lower baseline REE after adjusting for gender and body weight. Changes in REE following smoking cessation did not differ by ethnicity. CONCLUSIONS: These results suggest that black smokers may be more energy efficient, which could contribute to ethnic differences in postcessation weight gain. PMID- 11114689 TI - Standardization of the 24-hour diet recall calibration method used in the european prospective investigation into cancer and nutrition (EPIC): general concepts and preliminary results. AB - OBJECTIVES: Despite increasing interest in the concept of calibration in dietary surveys, there is still little experience in the use and standardization of a common reference dietary method, especially in international studies. In this paper, we present the general theoretical framework and the approaches developed to standardize the computer-assisted 24 h diet recall method (EPIC-SOFT) used to collect about 37 000 24-h dietary recall measurements (24-HDR) from the 10 countries participating in the European Prospective Investigation into Cancer and Nutrition (EPIC). In addition, an analysis of variance was performed to examine the level of standardization of EPIC-SOFT across the 90 interviewers involved in the study. METHODS: The analysis of variance used a random effects model in which mean energy intake per interviewer was used as the dependent variable, while age, body mass index (BMI), energy requirement, week day, season, special diet, special day, physical activity and the EPIC-SOFT version were used as independent variables. The analysis was performed separately for men and women. RESULTS: The results show no statistical difference between interviewers in all countries for men and five out of eight countries for women, after adjustment for physical activity and the EPIC-SOFT program version used, and the exclusion of one interviewer in Germany (for men), and one in Denmark (for women). These results showed an interviewer effect in certain countries and a significant difference between gender, suggesting an underlying respondent's effect due to the higher under-reporting among women that was consistently observed in EPIC. However, the actual difference between interviewer and country mean energy intakes is about 10%. Furthermore, no statistical differences in mean energy intakes were observed across centres from the same country, except in Italy and Germany for men, and France and Spain for women, where the populations were recruited from areas scattered throughout the countries. CONCLUSION: Despite these encouraging results and the efforts to standardize the 24-HDR interview method, conscious or unconscious behaviour of respondents and/or interviewer bias cannot be prevented entirely. Further evaluation of the reliability of EPIC-SOFT measurements will be conducted through validation against independent biological markers (nitrogen, potassium). PMID- 11114690 TI - Wine and other types of alcoholic beverages and the risk of esophageal cancer. AB - OBJECTIVE: To investigate the separate and combined effect of wine-drinking and other alcoholic beverages on esophageal cancer, in a high wine-consuming population. DESIGN: Combined analysis of two hospital-based case-control studies. SETTING: Major teaching and general hospitals in the greater Milan area and in the province of Pordenone, in northern Italy. SUBJECTS: A total of 714 incident cases of esophageal cancer, and 3137 controls admitted to hospital for acute, non neoplastic conditions, unrelated to alcohol consumption. INTERVENTION: Trained interviews identified and questioned cases and controls using standardized structured questionnaires, including information on the average number of days per week each type of alcoholic beverages (wine, beer, spirits) was consumed, and the average number of drinks per day. Odds ratios (ORs) were calculated using unconditional multiple logistic regression equations. RESULTS: With reference to total alcohol drinking, as compared to non- or moderate drinkers (<3 drinks per day), the multivariate ORs were 1.98 for drinkers of 3-4 drinks per day, 4.22 for 5-7, 7.60 for 8-11, and 12.35 for > or =12 drinks per day. Higher risks were observed for wine-only drinkers and the corresponding values were 1.70, 4.21, 8.76 and 17.90. After allowance for wine intake, no association was observed between beer and spirit drinking and esophageal cancer, in a population in which 80% of alcohol came from wine. CONCLUSION: The amount of ethanol determines the risk of esophageal cancer, and the most commonly used alcoholic beverage appear to be most strongly associated. PMID- 11114691 TI - Blood pressure and aging. PMID- 11114692 TI - Seven-year follow-up of blood pressure in the Healthy Old People in Edinburgh (HOPE) cohort. AB - The relationship between blood pressure and health in old age is complex and influenced by socio-economic factors. The Healthy Old People in Edinburgh cohort were initially disease-free and untreated, providing a sample in which directionality in this relationship could be examined. Subjects' health status, medication use and blood pressure was ascertained at baseline, after 4 years, and again after 7 years. Socio-demographic and socio-economic data were also collected. A total of 603 subjects were seen at baseline, 429 at 4 years and 301 at 7 years; complete blood pressure data were available for 294. Mean blood pressures were 157/85 mm Hg, 159/87 mm Hg and 162/86 mm Hg at baseline, 4 years and 7 years respectively. When subjects with diagnosed hypertension were excluded, the presence of disease (P = 0.009) and medication use (P = 0.047) at 7 years were associated with a relative reduction in blood pressure over time. For these subjects disease was predicted by deprivation index of residential area (OR 1.24, 95% CI 1.10-1.40 per Carstairs unit) and occupational group (OR 0.85, 95% CI 0.74-0.97 per major group). In this cohort disease, excluding hypertension itself, significantly attenuated the age-related rise in systolic blood pressure; the longer disease has been present the less the increase. In addition, socio economic variables are important predictors of blood pressure change in those with disease. Deprivation index of residential area was a better predictor of disease than previous occupation in these subjects who had retired over a decade previously. Journal of Human Hypertension (2000) 14, 773-778 PMID- 11114693 TI - An epidemiological study of hypertension and its determinants in a population in transition: the THUSA study. AB - BACKGROUND: Many black persons in South Africa have been subjected to urbanisation and urbanisation has led to a significant increase in diseases of lifestyle. The determinants of hypertension in a population in transition have not been well-defined and there is a pressing need for observational epidemiological studies as well as randomised-controlled trials in populations from Africa. The aim of this study was to investigate the association between blood pressure and factors known to contribute to hypertension. METHODS: The study sample consisted mainly of Setswana speaking people, divided into different levels (strata) of urbanisation, namely stratum 1 (rural) to stratum 5 (urbanized). A total of 1821 black subjects, which included 1040 woman, were recruited and randomly selected from 37 sites from the four geographical quarters of the North West Province of South Africa. The following questionnaires were used: demographic, anthropometric, quantitative food frequency, physical activity and scales to measure psychosocial variables. Biochemical analysis (standardised methods) were done on the serum and plasma of the subjects and the blood pressure was measured with a sphygmomanometer. RESULTS: Of the total sample, 22.8% of the subjects had systolic and 20.7% diastolic blood pressures above 140/90 mm Hg. Males and females from stratum 3 showed the highest rate of hypertension (32.9% systolic and 25.1% diastolic) and stratum 5 the lowest. Blood pressure correlated positively with age, level of urbanisation, WHR (waist:hip ratio) and smoking. In the woman the diastolic blood pressure correlated the best with body mass index (BMI), serum triglycerides, total serum cholesterol, low-density lipoprotein (LDL) cholesterol and s-GGT. Coping strategies, experience of social support, cultural aspects and affect balance are related to blood pressure, especially in the case of women. CONCLUSIONS: It seems that factors associated with urbanisation are related to the manifestation of hypertension in black people of the North West Province, given the highest mean blood pressure in people living in informal settlements, where most newcomers to the urban areas live. Journal of Human Hypertension (2000) 14, 779-787 PMID- 11114695 TI - The effect of transcutaneous electric nerve stimulation in patients with therapy resistant hypertension. AB - OBJECTIVES: Afferent nerve stimulation, such as acupuncture and transcutaneous electric nerve stimulation (TENS), has shown a blood pressure reduction in both animal and man. In the present open and non-controlled study we investigated the effect on 24-h ambulatory blood pressure of low frequency TENS in a group of hypertensive subjects who do not respond properly to pharmacological treatment. METHOD: Twelve patients were investigated. The patients were treated with TENS at two acupoints on both forearms for 30 min twice daily during 4 weeks. 24-hour ambulatory blood pressure monitoring was recorded 1 week before, at start, at the end and finally 1 week after the TENS treatment. RESULTS: The blood pressure did not change significantly during the run-in period. After 4 weeks of TENS, the mean systolic blood pressure decreased by 6.3 mm Hg (P < 0.05) and the mean diastolic blood pressure decreased by 3.7 mm Hg (P < 0.05). The blood pressure reduction remained unchanged 1 week after treatment. There was no change in mean heart rate. CONCLUSION: The present study suggests that continuous TENS may have additional blood pressure-lowering properties in hypertensive patients who do not respond properly to pharmacological treatment. The effect of TENS may also have a prolonged effect. Journal of Human Hypertension (2000) 14, 795-798 PMID- 11114694 TI - Effects of the angiotensinogen gene M235T and A(-6)G variants on blood pressure and other vascular risk factors in a Spanish population. AB - Angiotensinogen (AGT) gene polymorphism has shown significant differences in the allelic frequencies between hypertensive and normotensive subjects. This allele frequency varies among ethnic groups. There are still some controversies related to the 235T-variant as a marker for essential hypertension. As part of an extensive case-control study carried out in a Spanish population, we selected the 237 subjects with a diagnosis of essential hypertension according to the established criteria. A group of 242 normotensives matched for age and gender was used as control. Smoking habits, a previous diabetes and hypertension medical history, body mass index (BMI) and blood pressure (BP) values were recorded. Glucose, plasma creatinine, lipid profile with Lp(a), homocysteine and microalbuminuria were measured. Angiotensinogen M235T-gene polymorphism was determined by polymerase chain reaction (PCR) from genomic DNA. A(-6)G polymorphism was determined by mutagenically separated PCR (MS-PCR). BP values, BMI and microalbuminuria were significantly higher in hypertensive subjects; 31.6% of hypertensives and 40.1% normotensives were active smokers. M235T genotype frequencies were not different in the hypertensive and normotensive population. Similarly, homocigotic AA predominate in the hypertensives but without statistical significance. The association of 235T-genotype or the changes in the promoter activity due to A(-6) substitution with essential hypertension was not confirmed in the multivariate regression analyses. Only a previous family history of hypertension and BMI were significantly associated with hypertension. Journal of Human Hypertension (2000) 14, 789-793 PMID- 11114696 TI - Blood pressure and metabolic profile after surgical menopause: comparison with fertile and naturally-menopausal women. AB - In 1978 a random sample (367 men and 568 women aged 18-65 years) taken from the general population of a north-eastern Italian town was screened for cardiovascular risk; 16 years later, the women were invited to a second screening. Three groups were identified at the initial screening (fertile, naturally menopausal and surgically menopausal) and four in the longitudinal study (137 remained fertile during the whole study, 205 became naturally menopausal, 56 were ovariectomised and 127 were already going through the menopause). The protocol included a questionnaire, blood pressure (BP) measurement, and blood exams. Continuous variables were adjusted for confounders. Systolic BP, prevalence of hypertension, cholesterol, glycaemia and uricaemia were similar, whereas diastolic and triglycerides (TG) were lower in surgically menopausal than in fertile women (P < 0.001). No significant difference in 16 years' variation from baseline was observed between the four groups, although women who remained fertile showed the smallest increases. In particular, neither systolic or diastolic BP increases differed between the women who were oophorectimised and those who remained fertile. 'Fertile status' was rejected from the logistic equation of incidence of hypertension, and 'age of menopause' was also rejected when this analysis was repeated in ovariectomised women. New coronary artery disease (angina pectoris or myocardial infarction) was observed in one ovariectomised woman, in three naturally menopausal, and in 13 already menopausal women which seemed to reflect the age trend. No new cases were observed in women who remained fertile. In conclusion, in Italian women surgical menopause, similarly to natural menopause, is devoid of any negative prognostic effect. Journal of Human Hypertension (2000) 14, 799-805 PMID- 11114697 TI - The potent role of increased sympathetic tone in pathogenesis of essential hypertension with neurovascular compression. AB - OBJECTIVE: To study the role of increased sympathetic tone in pathogenesis of hypertension in patients with essential hypertension with neurovascular compression. METHODS: Twenty-three patients with essential hypertension, 13 patients with secondary hypertension, and 46 normotensive subjects were investigated. Neurovascular compression was evaluated by MRT. The power spectral components of heart rate variability as indices of autonomic nerve tone were determined to investigate the possibility that sympathetic tone mediates the neurovascular compression-induced increase in blood pressure. RESULTS: Neurovascular compression of the rostral ventrolateral medulla (RVLM) was observed in 70% of essential hypertension group, none of secondary hyperension group and 16% of normotensive group (P < 0.001). The age-adjusted low-frequency power spectral density (A-PSD) (0.04 to 0.15 Hz), which is an index of sympathetic tone, was significantly higher in patients with essential hypertension (139.5 +/- 6.7%) with neurovascular compression than in essential hypertension patients without neurovascular compression (92.2 +/- 6.8%), normotensive subjects with (102.8 +/- 13.0%) and without neurovascular compression (100.1 +/- 4.1%), and patients with secondary hypertension (95.7 +/- 10.2%) (P < 0.001). There was no significant difference in the high-frequency A PSD (0.15 to 0.40 Hz), which is an index of vagal tone, among groups. CONCLUSIONS: Neurovascular compression was not always associated with an increase in sympathetic nerve tone. Hypertension was present in subjects with neurovascular compression, who had increased sympathetic tone but not in those with normal sympathetic tone. An increase in sympathetic tone may mediate the neurovascular compression-induced increase in blood pressure. Journal of Human Hypertension (2000) 14, 807-811 PMID- 11114698 TI - Absence of any significant effects of circadian blood pressure variations on carotid artery elastic properties in essential hypertensive subjects. AB - We sought in this study to examine the effects of diurnal blood pressure variations upon common carotid artery (CCA) elasticity in selected subjects with uncomplicated moderate essential hypertension. Towards this end, 174 non-smoker subjects with stage I-II essential hypertension and without diabetes mellitus, left ventricular hypertrophy and carotid atherosclerosis, were classified as dippers and non-dippers according to the diurnal variation of >10% between mean daytime and night-time systolic and diastolic blood pressure (BP) in 24-h non invasive ambulatory BP monitoring. CCA distensibility was derived by a combination of surface ultrasonographic data and simultaneous BP measurements at the brachial artery. The dippers and non-dippers were similar with respect to demographic characteristics. Non-dippers had significantly greater office systolic BP, 24-h systolic BP and ambulatory pulse pressure (PP) and significantly less (daytime-night-time) systolic and diastolic BP fall (by 16 mm Hg and 11 mm Hg respectively, P< 0.0001) compared to dippers. CCA distensibility was significantly reduced in non-dippers compared to dippers (by 0.89 dyne( 1)/cm(2/)10(-6), P < 0.05). Multiple linear regression analysis identified patient age and ambulatory PP as significant predictors of the CCA elasticity index. When patient age, 24-h systolic and diastolic BP were used as covariates in an analysis of covariance, the difference of CCA elasticity between dippers and non-dippers ceased to reach statistical significance. In contrast, when patient age, ambulatory PP, systolic (daytime-night-time) BP fall and diastolic (daytime-night-time) BP fall were used as covariates, the difference of CCA distensibility between dippers and non-dippers continued to be statistically significant. In conclusion, the excessive impairment of CCA elastic properties in non-dippers compared to dippers hypertensive seems to be ascribed to the increased of total 24-h haemodynamic load and not to the circadian pattern of BP. Journal of Human Hypertension (2000) 14, 813-818 PMID- 11114699 TI - Association between a variant in the 11 beta-hydroxysteroid dehydrogenase type 2 gene and primary hypertension. AB - The enzyme 11 beta-hydroxysteroid dehydrogenase type 2 (11BHSD2) converts cortisol to cortisone in the kidney, thereby protecting the mineralocorticoid receptor from the mineralocorticoid actions of cortisol. The syndrome of Apparent Mineralocorticoid Excess (AME), a rare monogenic form of early onset hypertension with autosomal recessive inheritance, is caused by homozygous or compound heterozygous loss of function mutations in the 11BHSD2 gene. Association has been reported between a microsatellite marker flanking the 11BHSD2 gene (D16S496) and primary hypertension. The aim of this study was to identify variants in the 11BHSD2 gene and to test if such variants or the D16S496 are associated with primary hypertension, in Swedes. To address this, the coding sequences of the 11BHSD2 gene was screened for mutations in 20 patients with primary hypertension with single strand conformation polymorphism and direct DNA sequencing techniques. A polymorphism was identified in exon 3; G534A (Glu178Glu). This polymorphism and the D16S496 microsatellite were tested for association with primary hypertension in a population consisting of 292 patients with primary hypertension and 263 normotensive control subjects. The frequency of G534G homozygotes was higher in patients with primary hypertension than in normotensive control subjects (92.8% vs 87.8%; P < 0.05). The allele frequencies of the D16S496 microsatellite did not differ between the two groups (chi(2) = 11.0, df = 10; P = 0.36). In conclusion, over-representation of individuals homozygous for the G534 allele in hypertensive patients compared with control subjects suggests that a mutation in linkage disequilibrium with the G534A polymorphism could increase susceptibility to primary hypertension. Journal of Human Hypertension (2000) 14, 819-823 PMID- 11114700 TI - Prevalence, awareness, treatment and control of hypertension in the elderly: results from a population survey. AB - Prevalence, awareness, treatment and control of hypertension were assessed in 1032 (90%) of 1147 elderly (> or = 65 years) inhabitants of three Italian villages. Blood pressure (BP) was measured at home on two separate occasions following a standardised protocol. Persons taking antihypertensive drugs or with BP values > or = 140/90 mm Hg were considered as affected by hypertension. Prevalence of hypertension was 64.8%, with higher rates in women than men, and in those aged 75-84 than in those aged 65-74. Diabetes, strokes and hypercholesterolaemia were more frequent in hypertensive than normotensive people, whereas cardiac diseases, overweight and smoking did not differ significantly between hypertensive and normotensive people. Of the 669 hypertensive patients, 439 (65.6%) were aware of their hypertension, 398 (59.5%) were being treated, and 70 (10.5%) had their hypertension controlled. Of the 230 unaware patients, 201 (87.4%) had had their BP measured in the previous year. Of these, 174 (86.6%) had stage 1 hypertension, while 27 had stage 2 hypertension with SBP values <170 mm Hg. Overall, the patients with stage 1 hypertension accounted for 68.3% of the untreated and 50.5% of the treated patients. The use of a single drug was more frequent in patients with controlled (97.1%) or stage 1 (97.0%) than with stages 2-3 (18.9%) hypertension. The drugs prescribed most were angiotensin-converting enzyme (ACE) inhibitors (45%), followed by diuretics (43%). As our findings suggest that BP values can be effectively reduced by treating or increasing drug treatment in stage 1 hypertensive patients, data on safety and effectiveness of this policy are urgently needed. Journal of Human Hypertension (2000) 14, 825-830 PMID- 11114702 TI - Annual Scientific Meeting of the British Hypertension Society. Cambridge, United Kingdom, 11-13 September 2000. Abstracts. PMID- 11114701 TI - Dissociation between microalbuminuria and common carotid thickness in essential hypertensive men. AB - BACKGROUND: The reasons why microalbuminuria (albuminuria > or = 15 microg/min), an expression of a renal microcirculatory abnormality, predicts cardiovascular disease in essential hypertension are unsettled. To test the hypothesis that microalbuminuria represents a marker of subclinical atherosclerosis, we evaluated its association with common carotid artery (CCA) intima media thickness (IMT), a measure of preclinical atherosclerosis and an independent predictor of cardiac and cerebrovascular events, in uncomplicated essential hypertensive individuals. MATERIALS AND METHODS: Albuminuria, ultrasonographic CCA IMT (the mean of six bilateral far wall measurements within 1.5 cm proximally to the flow divider), brachial blood pressure (BP), smoking habits and lipids were evaluated in 136 stage 1-3 untreated essential hypertensive men free of cardiovascular disease. RESULTS: CCA IMT did not differ between normo- (n = 99) and microalbuminuric (n = 37) patients. The correlation between CCA IMT and albuminuria was not significant, and the prevalence of microalbuminuria across IMT quartiles was not different. Microalbuminuric patients showed higher systolic BP and that parameter was the only independent correlate in a multivariate logistic regression model including also age, CCA IMT, diastolic BP, lipids and smoking habits as independent variables and microalbuminuria as the dependent one. CONCLUSION: This cross-sectional study in hypertensive subjects free of cardiovascular disease has shown a dissociation between microalbuminuria and CCA IMT, a surrogate measure of subclinical atherosclerosis, and a parameter linearly related to cardiovascular events. The data do not support the theory of microalbuminuria as a surrogate measure of subclinical atherosclerosis, while confirming the importance of systolic BP levels as an independent correlate of increased albuminuria in essential hypertension. Journal of Human Hypertension (2000) 14, 831-835 PMID- 11114704 TI - Alleviating oxidative stress in cancer immunotherapy: a role for histamine? AB - Interleukin-2 is a remarkable activator of lymphocytes with anti-neoplastic properties such as T-cells or natural killer cells, but tumor regression only rarely occurs in interleukin-2-treated cancer patients. In this review, we focus on interactions between monocytes/macrophages and T-cells/natural killer-cells, and in particular the role of such interactions for the outcome of cancer immunotherapy with interleukin-2. We propose that interleukin-2 therapy should be supplemented with compounds that alleviate toxicity inflicted by monocyte/macrophage-derived reactive oxygen metabolites within and around tumors. The hypothesis is founded on data demonstrating that (i) functions of intratumoral lymphocytes in many human malignant tumors are inhibited by reactive oxygen metabolites, generated by neighboring monocytes/macrophages, (ii) interleukin-2 only weakly activates T-cells or natural killer cells in an environment of oxidative stress, and (iii) inhibitors of the formation of reactive oxygen metabolites or scavengers of reactive oxygen metabolites synergize with interleukin-2 to activate these lymphocyte subsets. We also review the preclinical background to the use of histamine dihydrochloride, an inhibitor of reactive oxygen metabolite formation in monocytes/macrophages, as a supplement to cancer immunotherapy with interleukin-2. PMID- 11114705 TI - Generation of dendritic cells from peripheral blood of patients at different stages of chronic myeloid leukemia. AB - We report a method to generate dendritic cells (DC) from frozen leukapheresis products of patients with chronic myeloid leukemia (CML), using sterile culture bags and serum-free culture medium, ie conditions feasible for re-infusion into the patient as part of immunotherapeutic protocols. Leukapheresis products were stored from harvests performed either at diagnosis (13 patients) or after chemotherapy with subsequent granulocyte colony stimulating factor (G-CSF) administration (9 patients), for Peripheral Blood Stem Cell (PBSC) collections. In the presence of optimal concentrations of GM-CSF (50 ng/ml) and IL-4 (40 ng/ml) CML progenitors differentiated on day 7 and 14 of culture to DC, expressing CD1a,HLA-DR and CD86 surface antigens. Mature DCs exhibited on average 12-fold higher allo-stimulatory capacity for CD4+ and CD8+ cells compared to non cultured PBMC in mixed lymphocyte reaction (MLR). Only DCs obtained from CML patients at diagnosis exhibited bcr/abl fusion gene when tested by fluorescent in situ hybridization (FISH). CD34-selection on leukapheresis products from diagnosis (7 patients) resulted in later maturation of DCs (after 14-15 d), compared to the nonselected PBMC. CD34-selection significantly increased the DC growth, and improved the allo-stimulatory capacity in MLR (on average on day 14, 3.5- and 2.3-fold, respectively). Large differences were observed between individual patients and different leukapheresis products from the same patient. Our report demonstrates the possibility to generate ex vivo autologous functionally active DC in CML in a way that allows their clinical application as immunotherapeutic agents. PMID- 11114703 TI - Interferon-alpha and the pathogenesis of myeloproliferative disorders. AB - Interferon-alpha (IFN-alpha), a molecule with multiple biological actions, is widely used in the treatment of chronic myelogenous leukemia (CML) and the other myeloproliferative disorders. This glycoprotein belonging to the type I subfamily of interferons has been recombinantly manufactured and has been approved for the biotherapy of CML, now becoming the first line of treatment for CML patients in chronic phase who are not candidates for allogeneic hematopoietic stem cell or bone marrowtransplantation. Interferon-alpha action involves binding to its cell membrane receptor and initiation of an intracellular signal transduction cascade. Two major pathways mediate the biologic actions of IFN-alpha. The JAK-STAT pathway leads to phosphorylation and activation of STAT 1 and STAT 2 molecules and transcription of genes like p21 and caspase-1 resulting in cycle arrest and apoptosis. The PKR (protein kinase dsRNA-induced) kinase phosphorylates and inhibits the eukaryotic initiator of translation eIF-2alpha leading again to apoptosis. The PKR kinase cascade also leads to activation of the transcription factor NF-kappaB. The relevance of this activation is unclearand it is possiblethat NF-kappaB has not had the opportunity to transcribe its target genes as it is a substrate of effector caspases and is maybe cleaved by them before exerting any transcription activity. Through the JAK-STAT and the PKR kinase pathways IFN-alpha is able to modify the proliferative and antiapoptotic actions of the constitutively activated kinase bcr-abl, the product of the t(9;22) translocation present in CML, and has therapeutic effects in this disease. PMID- 11114706 TI - Long-term survival with metastatic cancer to the brain. AB - Metastatic cancer to the brain has a poor prognosis. The focus of this work was to determine the incidence of long-term (> or = 2y) survival for patients with brain metastases from different primary cancers and to identify prognostic variables associated with prolonged survival. A retrospective review of 740 patients with brain metastases treated over a 20 y period identified 51 that survived 2 or more years from the time of diagnosis of the brain metastasis. Prognostic variables that were examined included age, sex, histology, tumor number and location, and treatment. In the 51 patients, 35 (69%) had single lesions and 16 (31%) had multiple tumors. For all tumor types (740 patients), the actuarial survival rate was 8.1% at 2 y, 4.8% at 3 y, and 2.4% at 5 y. At 2 y, patients with ovarian carcinoma had the highest survival rate (23.9%) and patients with small cell lung cancer (SCLC) had the lowest survival rate (1.7%). At 5y, survival rates were 7.8% for ovarian carcinoma, 2.9% for non-SCLC, 2.3% for melanoma and renal cell carcinoma, 1.3% for breast carcinoma and there were no survivors with SCLC, gastrointestinal, bladder, unknown primary, or prostate cancer. Age, sex, histology, location for single tumors, systemic chemotherapy, and stereotactic radiosurgery did not significantly influence survival. The presence of a single lesion (P = 0.001, chi-square test), surgical resection (P= 0.001), and WBRT (P = 0.009) were favorable prognostic variables for extended survival. Multiple bilateral metastases was a poor prognostic indicator (P= 0.001). Multivariate analysis showed younger age (P< 0.05), single metastasis (P < 0.0001), surgical resection (P < 0.0001), whole brain radiation therapy (P < 0.0001), and chemotherapy (P = 0.0288) were associated with prolonged survival. 29 patients (57%) died of systemic disease progression, 9 (18%) died of central nervous system progression, and the cause of death was unknown in 3 (6%). Patients with a single non-SCLC, breast, melanoma, renal cell, and ovarian carcinoma brain metastasis have the best chance for long-term survival if treated with surgical resection and WBRT. PMID- 11114707 TI - Ototoxicity after high-dose chemotherapy with cyclophosphamide, thiotepa and carboplatin followed by stem cell transplantation in patients with breast cancer. AB - Our purpose was to determine the risk of ototoxicity in breast cancer patients receiving a myeloablative regimen consisting of cyclophosphamide 6000 mg/m2, thiotepa 500 mg/m2 and carboplatin 800 mg/m2 (CTCb) followed by stem cell transplantation. Fourteen consecutive patients with breast cancer were treated with high dose chemotherapy consisting of the CTCb regimen followed by stem cell transplantation. A pretransplant complete hearing study was obtained which consisted of hearing case history, audiometry and tympanometry. In addition, DPOAE (Distortion Product Otoaccoustic Emissions) was done to evaluate measurable changes in the cochlear (outer hair cell) functioning. Pre-transplant, all patients had no clinical evidence of hearing impairment and hearing studies were normal. Eleven patients had hearing studies and a telephone interview posttransplant. One patient was lost to follow-up and two patients died. One of the 11 patients tested had an abnormal post-transplant hearing study but none of them had clinically detectable hearing impairment. In our prospective study of breast cancer patients treated with the CTCb regimen, we did not observe clinically detectable hearing impairment in any of the patients tested. PMID- 11114708 TI - Pregnancy-associated breast cancer: a case-control study in a young population with a high-fertility rate. AB - Pregnancy-associated breast cancer (PABC) is not a rare event. The association frequently imposes a management challenge. We intended to review the clinical features, therapy, and outcome of patients with PABC seen at a single institution over a five-year period and to compare those with that seen in a matched control group. Data of all patients with PABC diagnosed during pregnancy were retrospectively reviewed (Group I). For each patient in Group I, three matched controls with breast cancer without pregnancy were identified (matched for age, stage, and year of diagnosis, Group II). 72 patients in Group I and 216 in Group II were identified. Their median age was similar (34 vs 35 y, respectively). The median number of prior pregnancies for patients in Groups I and II was 5. Patients had shorter duration of symptoms prior to diagnosis as compared with their controls (5.6 vs 9.4 months, P < 0.0001). 3%, 31%, 40%, and 26% of patients had Stage I to IV, respectively. A pattern that was similar to that seen in our breast cancer population. Pregnancy was terminated in 34 patients (47%), while 38 (53%) had normal spontaneous vaginal delivery. 47 patients in Group I had surgery; 37 (52%) had modified radical mastectomy and 10 (14%) had conservative surgery. In 37 patients surgery was performed after termination of pregnancy and 10 had surgery performed during pregnancy. The median number of positive lymph nodes in Group I was 4 as compared with 2 for patients in Group II. No patients in Group I had systemic chemotherapy during first trimester, while only 4 (6%) and 3 (4%) received adjuvant or neoadjuvant during second and third trimester, respectively. No congenital malformation in the newborns was diagnosed. None of the patients in Group I received radiotherapy during pregnancy. Over a median of 47.5 months, 48 (67%) patients in Group I were alive as compared to 126 (58%) in Group II, with no difference in the median survival (P= 0.79). Comparing overall survival (OS) between the two groups stage for stage also showed no significant difference. Also there was no difference in progression-free survival between the two groups. Cox proportional hazard model identified advanced stage as the only independent adverse prognostic variable that influenced OS in Group I. Despite that this series included a relatively young population with a high fertility rate, the study confirmed the lack of a survival difference between patients with PABC and their matched controls. PMID- 11114709 TI - Clinico-biological features of 30 patients with acute promyelocytic leukemia and response to combination induction chemotherapy with all-trans retinoic acid and anthracycline. AB - 30 adult patients with acute promyelocytic leukemia (APL) were seen at our institution overthe past 7 years. Their white cell count at presentation ranged from 400/microl to 54,900/microl. Cytogenetic studies were successful in 28 patients, of which 26(93%) were positive for t(15;17). Molecular analysis by reverse-transcription polymerase chain reaction demonstrated the PML-RARalpha fusion transcript in all 30 patients. The majority of patients had breakpoints at the 3' end with bcr1 products predominating. Complete remission rate of 92% was achieved using all-trans retinoic acid and anthracycline as induction chemotherapy in 26 patients. Of these, retinoic acid syndrome was observed in 4 cases, with 1 fatality. In conclusion, APL is a distinct entity with a highly specific molecular marker - t(15;17) translocation - that can be successfully induced into remission with all-trans retinoic acid and anthracycline in most patients. PMID- 11114710 TI - Additional chromosome aberrations in acute promyelocytic leukemia: characteristics and prognostic influence. AB - Patients with acute promyelocytic leukemia (APL) show other chromosome aberrations in addition to t(15;17) but their influence on the clinical outcome is still unclear. We have cytogeneticaly analyzed 43 APL patients with t(15;17)(q22;q21), treated with all-trans-retinoic acid (ATRA) according to the recommendations of the European APL 91 Group. Additional chromosome aberrations were observed in 14/43 patients (33%) studied at initial diagnosis. These patients were designed as 'complex' karyotype group and were compared to patients with t(15;17) asa sole cytogenetic abnormality ('simple' karyotype group). The 'complex' group had significantly lower platelet count and fibrinogen level and fewer cases without significant DIC at diagnosis than the 'simple' group. Comparison of 'simple' and 'complex' groups showed significant difference in complete remission rate (76% vs 35.7%, P = 0.0148) and early death rate (24% vs 64.3%, P = 0.0141). Survival analysis showed that the presence of additional chromosome abnormalities and significant DIC had an adverse effects on prognosis (P = 0.036 and P = 0.041, respectively), independent on other prognostic factors. These data indicate more aggressive biological nature of leukemic cells in patients with additional chromosome aberrations. Supplementary therapeutic strategies may be required for this subgroup of APL patients. PMID- 11114711 TI - Microvessel density in chemosensitive and chemoresistant diffuse large B-cell lymphomas. AB - Preliminary reports involving a number of different kinds of tumors have indicated that microvessel quantification may be useful in predicting disease outcome. The aim of this study was to examine the relationship between microvessel density (MVD) as a parameter of tumor angiogenesis and the response to chemotherapy in diffuse large B-cell (DLBC) lymphomas. A total of 36 DLBC lymphoma patients were evaluated, 23 of them with a chemosensitive; responsive disease (median survival 8y) and 13 with a chemoresistant, refractory disease (median survival 8 months). Microvessel quantification was performed by immunohistochemical staining, using monoclonal antibodies against factor VIII related antigen (F8RA) and against platelet/endothelial cell adhesion molecule CD31. We found that F8RA stained a significantly higher number of blood vessels (about 2.5 times more) than CD-31; 7 samples were not stained with CD-31 but were positive for F8RA. There was no significant difference between the density of microvessel staining of the two groups. In the chemosensitive DLBC lymphomas positive for F8RA, the mean number of microvessels stained was 54.5 +/- 36.1 per microscopic field (200x) examined (range 6-149) whereas in the chemoresistant group the corresponding mean number was 43.1 +/- 25.5 (range 11-94). F8RA appears to be more sensitive for staining DLBC lymphomas microvessels than CD-31. Our data demonstrate that there is no correlation between tumor MVD and response to chemotherapy in patients with DLBC lymphomas. PMID- 11114712 TI - Activities of adenosine deaminase and 5'-nucleotidase in cancerous and noncancerous human colorectal tissues. AB - In order to characterize human colorectal cancer, much attention has been paid to enzyme studies. However, little is known about the correlation between the levels of key enzymes of purine nucleotide pathway and some clinical and biological indicators of tumor invasiveness and aggressiveness. Adenosine deaminase (ADA) and 5'-nucleotidase (5'-NT) were measured in cancerous and cancer-free adjacent large bowel tissues from 38 patients with colorectal carcinoma. We have analyzed the relationship between the enzyme levels and some clinical and pathological parameters. The enzymes' activities were markedly higher in primary tumors than in corresponding normal mucosae. The ADA level in tumor tissue was significantly correlated with lymph node metastasis, histologic type, tumor location, and patient's age, whereas the 5'-NT level showed a significant correlation with tumor grade and tumor location. ADA activity in tumor tissues was significantly higher in patients whose clinical course remained stable than in those with recurrent diseases. The purine metabolism and salvage pathway activity of purine nucleotides are accelerated in the cancerous human colorectal tissue. Although our findings suggest that these enzymes' activities are most likely related to the same histomorphological architecture of the tumor, the authors believe that long-term follow-up studies are needed to evaluate the prognostic value of purine enzymes for colorectal cancer. PMID- 11114714 TI - Successful therapy of transplant-associated veno-occlusive disease with a combination of tissue plasminogen activator and defibrotide. AB - A 36-year-old man underwent matched unrelated donor bone marrow transplantation for chronic myeloid leukaemia. He developed severe hepatic veno-occlusive disease as an early post-transplant complication. Tissue plasminogen activator was initially felt to be contraindicated since the patient had concomitant pericarditis. Defibrotide was therefore commenced as treatment for veno-occlusive disease. The pericarditis improved but the veno-occlusive disease continued to worsen (peak bilirubin 353 micromol/l). Tissue plasminogen activator followed by a heparin infusion was therefore administered. However, he proceeded to develop haemorrhagic cardiac tamponade that required drainage. Thrombolysis was therefore discontinued and treatment with defibrotide resumed after an interval of 48 h. The veno-occlusive disease gradually resolved and defibrotide was discontinued once the bilirubin had plateaued. He was discharged home on day +52 post transplant. PMID- 11114713 TI - Proliferation and apoptosis in the evolution of endemic and acquired immunodeficiency syndrome-related Kaposi's sarcoma. AB - Kaposi's sarcoma (KS) is a multifocal lesion that occurs predominantly in the skin, most frequently in people infected with HIV-1, and that evolves through early stages (patch and plaque) to a tumor-like late stage (nodular). Both, endemic African (EKS) and AIDS-associated (AKS) KS expressed human herpesvirus 8 (HHV-8) as shown by PCR. By immunohistochemistry the expression of cellular Bcl-2 and c-myc was confined in early stages of both EKS and AKS to relatively few endothelial cells (EC) whereas in nodular KS most of spindle cells (SC) strongly expressed both genes. CD40 was usually strongly expressed in SC at all KS stages as well as in EC of non-involved tissue whereas CD40L (CD154) was not demonstrable. Fas (CD95) was moderately to weakly expressed by SC whereas p53 and Waf-1 were found in less than 5% of the SC. In both AKS and EKS at nodular stage almost no apoptotic SC were detected. In most AKS and EKS low levels of cell proliferation were seen but AKS showed consistently higher values compared to EKS. All clinical types and stages of KS showed a diploid cellular DNA content by flow cytometric analysis of microselected lesions. Thus, we conclude that KS during evolution represents diploid, probably reactive, cell proliferation, which progressively increases the expression of strong cellular and also viral (HHV-8) antiapoptotic factors. PMID- 11114715 TI - Pathologic rupture of the spleen during induction with ATRA in a patient with acute promyelocytic leukemia. AB - Pathological rupture of the spleen is a rare but well recognized complication in hematological malignancies. Early clinical recognition of this life-threatening complication is necessary for rapid intervention. Here, we report on the case of a 26-year-old woman with acute promyelocytic leukemia who presented rupture of the spleen on day +2 of treatment with ATRA plus idarrubicin. In patients with acute leukemia, the presence of a painful abdomen and a sudden drop in hemoglobin levels, should alert of a possible splenic rupture, even without additional symptoms. This would facilitate an early treatment intervention with no modification to the chemotherapy schedule. PMID- 11114716 TI - Overdosage of pamidronate in a patient with renal cell carcinoma. PMID- 11114717 TI - Metastatic small cell lung cancer causing biliary obstruction. PMID- 11114718 TI - Induction of p21WAF1/CIP1 and cyclin D1 expression by the Src oncoprotein in mouse fibroblasts: role of activated STAT3 signaling. AB - While the activated viral Src oncoprotein, v-Src, induces uncontrolled cell growth, the mechanisms underlying cell cycle deregulation by v-Src have not been fully defined. Previous studies demonstrated that v-Src induces constitutively active STAT3 signaling that is required for cell transformation and recent data have implicated STAT3 in the transcriptional control of critical cell cycle regulators. Here we show in mouse fibroblasts stably transformed by v-Src that mRNA and protein levels of p21 (WAF1/CIP1), cyclin D1, and cyclin E are elevated. Using reporter constructs in transient-transfection assays, the cyclin D1 and p21 promoters were both found to be transcriptionaly induced by v-Src in a STAT3 dependent manner. The kinase activities of cyclin D/CDK4, 6 and cyclin E/CDK2 complexes were only slightly elevated, consistent with the findings that coordinate increases in p21, cyclin D1 and cyclin E resulted in an increase in cyclin/CDK/p21 complexes. Similar results were obtained in NIH3T3 and BALB/c 3T3 cells stably transformed by v-Src, indicating that these regulatory events associated with STAT3 signaling represent common mechanisms independent of cell line or clonal variation. These findings suggest that STAT3 has an essential role in the regulation of critical cell cycle components in v-Src transformed mouse fibroblasts. PMID- 11114719 TI - An alternatively spliced isoform of B-Myb is a transcriptional inhibitor. AB - B-Myb is a highly conserved member of the Myb transcription factor family. The primary transcript of the B-myb gene is spliced alternatively in two mRNAs which either contain or lack a sequence corresponding to the so-called exon 9A of c myb. Recent studies showed that full-length B-Myb containing the exon 9A encoded amino acids is a cell cycle regulated transcription factor whose activity is stimulated by cyclin A/Cdk 2-dependent phosphorylation at the carboxyl-terminus of B-Myb. We have now investigated in more detail the transactivation potential of the shorter isoform of B-Myb lacking exon 9A. Here, we show that B-Myb lacking exon 9A has no transactivation activity even in the presence of cyclin A. This inactivity of the shorter isoform of B-Myb is not due an improper subcelluar localization. Our work suggests that B-Myb lacking exon 9A may act as an inhibitor for full-length B-Myb mediated transactivation. Furthermore, by analysing the transactivation potential of Gal4/B-Myb fusion proteins we have identified the amino-terminal part of the exon 9A as the principal transactivation domain of full-length B-Myb. The results presented here demonstrate that B-myb encodes both an activator and an inhibitor of transcription and, thus, reveal an additional level of regulation of B-Myb activity beside the known cyclin dependent mechanisms. PMID- 11114720 TI - Up-regulation of hypoxia-inducible factors HIF-1alpha and HIF-2alpha under normoxic conditions in renal carcinoma cells by von Hippel-Lindau tumor suppressor gene loss of function. AB - Hypoxia induces transcription of a range of physiologically important genes including erythropoietin and vascular endothelial growth factor. The transcriptional activation is mediated by the hypoxia-inducible factor-1 (HIF-1), a heterodimeric member of the basic helix-loop-helix PAS family, composed of alpha and beta subunits. HIF-1alpha shares 48 per cent identity with the recently identified HIF-2alpha protein that is also stimulated by hypoxia. In a previous study of hemangioblastomas, the most frequent manifestation of hereditary von Hippel-Lindau disease (VHL), we found elevated levels of vascular endothelial growth factor and HIF-2alpha mRNA in stromal cells of the tumors. Mutations of the VHL tumor suppressor gene are associated with a variety of tumors such as renal clear cell carcinomas (RCC). In this study, we analysed the expression of the hypoxia-inducible factors HIF-1alpha and HIF-2alpha in a range of VHL wildtype and VHL deficient RCC cell lines. In the presence of functional VHL protein, HIF-1alpha mRNA levels are elevated, whereas HIF-2alpha mRNA expression is increased only in cells lacking a functional VHL gene product. On the protein levels, however, in VHL deficient cell lines, both HIF-alpha subunits are constitutively expressed, whereas re-introduction of a functional VHL gene restores the instability of HIF-1alpha and HIF-2alpha proteins under normoxic conditions. Moreover, immunohistochemical analyses of RCCs and hemangioblastomas demonstrate up-regulation of HIF-1alpha and HIF-2alpha in the tumor cells. The data presented here provide evidence for a role of the VHL protein in regulation of angiogenesis and erythropoiesis mediated by the HIF-1alpha and HIF-2alpha proteins. PMID- 11114721 TI - The integrin linked kinase (ILK) induces an invasive phenotype via AP-1 transcription factor-dependent upregulation of matrix metalloproteinase 9 (MMP 9). AB - Overexpression of Integrin Linked Kinase (ILK) in intestinal and mammary epithelial cells results in a highly invasive phenotype, associated with increased levels of expression of the matrix metalloproteinase MMP-9. This increase was at the transcriptional level as determined by MMP-9 promoter-CAT reporter assays. Mutations in the two AP-1 binding sites within the MMP-9 promoter completely inhibited the reporter activity. We have previously shown that ILK inhibits glycogen synthase kinase-3 (GSK-3) activity. Transient transfection of wild-type GSK-3beta in ILK-overexpressing cells decreased MMP-9 promoter activity and AP-1 activity, indicating that ILK can stimulate MMP-9 expression via GSK-3beta and AP-1 transcription factor. A small molecule inhibitor of the ILK kinase reduced the in vitro invasiveness of ILK overexpressing cells as well as the invasiveness of several human brain tumor cell lines. Furthermore, both MMP-9 promoter and AP-1 activities were inhibited by the ILK inhibitor. Invasiveness of ILK-overexpressing cells was also reduced by inhibition of MMP-9. These data demonstrate that ILK can induce an invasive phenotype via AP-1-dependent upregulation of MMP-9. PMID- 11114722 TI - The C. elegans orthologue ceBNIP3 interacts with CED-9 and CED-3 but kills through a BH3- and caspase-independent mechanism. AB - We have studied ceBNIP3, the orthologue of BNIP3 in C. elegans. Sequence analysis reveals that the different domains of BNIP3 have been conserved throughout evolution. ceBNIP3 contains a C-terminal transmembrane (TM) domain, a conserved domain (CD) of 19 amino acids, a BCL-2 homology-3 (BH3)-like domain and a PEST sequence. ceBNIP3 is expressed primarily as a 25 kDa monomer and a 50 kDa homodimer. After transfection, ceBNIP3 protein is rapidly degraded through a ubiquitin-dependent pathway by the proteasome. Like BNIP3, the TM domain of ceBNIP3 mediates the localization of the protein to mitochondria and is also necessary for homodimerization and cell death in mammalian cells. Neither the putative BH3 domain nor conserved domain is necessary for killing. ceBNIP3 protein interacts with CED-9 and BCL-XL, but unlike other pro-apoptotic BCL-2 family members, the BH3-like domain does not participate in dimerization. The ceBNIP3 TM domain mediates interaction with both CED-9 and BCL-XL. ceBNIP3 interacts with CED-3 but co-expression of CED-3 and ceBNIP3 does not significantly enhance induction of cell death in the presence or absence of CED 4. ceBNIP3 kills mammalian cells by a caspase-independent mechanism. In conclusion, we find that although ceBNIP3 interacts with CED-9 and CED-3 it kills by a BH3- and caspase-independent mechanism. PMID- 11114723 TI - Interaction of the retinoblastoma protein (pRb) with the catalytic subunit of DNA polymerase delta (p125). AB - The retinoblastoma gene product (pRb) interacts with many cellular proteins to function in the control of cell division, differentiation, and apoptosis. Several pRb binding proteins complex with pRb through an amino acid sequence called the LXCXE motif. The catalytic subunit of DNA polymerase delta (p125) contains a LXCXE motif. To further study the biochemical function of this polymerase, we sought to determine if p125 interacts with pRb. Experiments using GST-pRb fusion proteins showed that p125 from breast epithelial (MCF10A) cell extracts associates with pRb. In addition, GST-p125 fusion proteins bound pRb from the same cell extracts. The pRb that associated with GST-p125 was largely unphosphorylated. Coimmunoprecipitation experiments using cell cycle synchronized cells revealed that p125 and pRb form a complex predominantly during G1 phase, the phase during which pRb is mostly unphosphorylated. In vitro phosphorylation of GST-pRb by the cyclin dependent kinases reduced the ability of p125 to associate with GST-pRh. Addition of the LXCXE containing protein SV40 large T antigen to GST-pRb blocks the ability of p125 to associate with pRb, suggesting that it may be through a LXCXE sequence by which p125 interacts with pRb. Finally, in vitro polymerase assays demonstrate that GST-pRb fusion protein stimulates DNA polymerase delta activity. PMID- 11114724 TI - Expression of the BRK tyrosine kinase in mammary epithelial cells enhances the coupling of EGF signalling to PI 3-kinase and Akt, via erbB3 phosphorylation. AB - A high proportion of human breast cancers, in contrast with normal mammary tissue, express the intracellular tyrosine kinase BRK. BRK expression enhances the mitogenic response of mammary epithelial cells to epidermal growth factor, and conferment of a proliferative advantage through this mechanism may account for the frequent elevation of BRK expression in tumours. Here we report that BRK expression in mammary epithelial cells, at pathologically relevant levels, results in an enhanced phosphorylation of the epidermal growth factor receptor related receptor erbB3 in response to epidermal growth factor. As a consequence, erbB3 recruits increased levels of phosphoinositide 3-kinase, and this is associated with a potentiated activation of Akt. This effect of BRK on the regulation of phosphoinositide 3-kinase and Akt activity may account for BRK's ability to enhance mammary cell mitogenesis, and raises the possibility that breast tumours expressing BRK may acquire a resistance to pro-apoptotic signals. PMID- 11114725 TI - Bcl-XL protects pancreatic adenocarcinoma cells against CD95- and TRAIL-receptor mediated apoptosis. AB - In this study we sought to clarify the role of the proapoptotic potential of mitochondria in the death pathway emanating from the TRAIL (APO-2L) and CD95 receptors in pancreatic carcinoma cells. We focused on the role of the Bcl-2 family member Bcl-XL, using three pancreatic carcinoma cell lines as a model system, two of which have high (Panc-1, PancTuI) and one has low (Colo357) Bcl-XL expression. In these cell lines, the expression of Bcl-XL correlated with sensitivity to apoptosis induced by TRAIL or anti-CD95. Flow cytometric analysis revealed cell surface expression of TRAIL-R1 and TRAIL-R2 on PancTuI and Colo357, and TRAIL-R2 on Panc-1 cells. In Colo357 cells retrovirally transduced with Bcl XL, caspase-8 activation in response to treatment with TRAIL or anti-CD95 antibody was not different from parental cells and EGFP-transfected controls, however, apoptosis was completely suppressed as measured by the mitochondrial transmembrane potential deltapsim, caspase-3 activity (PARP cleavage) and DNA fragmentation. Inhibition of Bcl-XL function by overexpression of Bax or administration of antisense oligonucleotides against Bcl-XL mRNA resulted in sensitization of Panc-1 cells to TRAIL and PancTuI cells to anti-CD95 antibody induced cell death. The results show that Bcl-XL can protect pancreatic cancer cells from CD95- and TRAIL-mediated apoptosis. Thus, in these epithelial tumour cells the mitochondrially mediated 'type II' pathway of apoptosis induction is not only operative regarding the CD95 system but also regarding the TRAIL system. PMID- 11114726 TI - Bcr-Abl protein tyrosine kinase activity induces a loss of p53 protein that mediates a delay in myeloid differentiation. AB - Chronic myeloid leukaemia is a haemopoietic stem cell disorder, the hallmark of which is the expression of the Bcr-Abl Protein Tyrosine Kinase (PTK). We have previously reported that activation of a temperature sensitive Bcr-Abl PTK in the multipotent haemopoietic cell line FDCP-Mix for short periods resulted in subtle changes including, a transient suppression of apoptosis and no inhibition of differentiation. In contrast, activation of the Bcr-Abl PTK for 12 weeks results in cells that display a delay in differentiation at the early granulocyte stage. Flow cytometric analysis also indicates that the expression of cell surface differentiation markers and nuclear morphology are uncoupled. Furthermore, a significant number of the mature neutrophils display abnormal morphological features. Prolonged exposure to Bcr-Abl PTK results in interleukin-3 independent growth and decreased p53 protein levels. FDCP-Mix cells expressing a dominant negative p53 and p53null FDCP-Mix cells demonstrate that the reduction in p53 is causally related to the delay in development. Returning the cells to the restrictive temperature restores the p53 protein levels, the growth factor dependence and largely relieves the effects on development. We conclude that prolonged Bcr-Abl PTK activity within multipotent cells results in a reduction of p53 that drives a delayed and abnormal differentiation. PMID- 11114727 TI - The RelA NF-kappaB subunit and the aryl hydrocarbon receptor (AhR) cooperate to transactivate the c-myc promoter in mammary cells. AB - NF-kappaB/Rel transcription factors regulate many genes involved in control of cellular proliferation, neoplastic transformation, and apoptosis, including the c myc oncogene. Recently, we have observed that levels of NF-kappaB and aryl hydrocarbon receptor (AhR), which mediates malignant transformation by environmental carcinogens, are highly elevated and appear constitutively active in breast cancer cells. Rel factors have been found to functionally interact with other transcription factors. Here we demonstrate a physical and functional association between the RelA subunit of NF-kappaB and AhR resulting in the activation of c-myc gene transcription in breast cancer cells. RelA and AhR proteins were co-immunoprecipitated from cytoplasmic and nuclear extracts of non malignant MCF-10F breast epithelial and malignant Hs578T breast cancer cells. In transient co-transfection, RelA and AhR gene products demonstrated cooperation in transactivation of the c-myc promoter, which was dependent on the NF-kappaB elements, and in induction of endogenous c-Myc protein levels. A novel AhR/RelA containing NF-kappaB element binding complex was identified by electrophoretic mobility shift analysis of nuclear extracts from RelA and AhR co-transfected Hs578T cells. Thus, the RelA and AhR proteins functionally cooperate to bind to NF-kappaB elements and induce c-myc gene expression. These findings suggest a novel signaling mechanism whereby the Ah receptor can stimulate proliferation and tumorigenesis of mammary cells. PMID- 11114728 TI - Altered Ets transcription factor activity in prostate tumor cells inhibits anchorage-independent growth, survival, and invasiveness. AB - The Ets family of transcription factors are important downstream targets in cellular transformation, as altering Ets activity has been found to reverse the transformed phenotype of Ras transformed mouse fibroblasts and of several human tumor cell lines. To determine whether Ets factors are important targets in the largely uncharacterized aberrant signaling in prostate cancer, we have altered Ets activity in the prostate tumor cell line PPC-1, by stable expression of either full-length Ets2, or a dominant inhibitor of Ets activity, the Ets2 DNA binding domain (Ets2DBD). Analysis of multiple independent clonal cell lines revealed that expression of either Ets2 or the Ets2DBD inhibited the anchorage independent growth of PPC-1 cells up to 20-fold. In contrast to our previous findings with Ras-transformed NIH3T3 cells, PPC-1 cell lines expressing either Ets2 or the EtsDBD exhibited slower attached cell growth, increased Ets-dependent gene expression, and up to a 10-fold increase in apoptotic cell death. The p21cip gene was identified as a potential target of altered Ets signaling. Interestingly, the two distinct Ets2 constructs had strikingly different effects on in vitro invasiveness. Expression of the Ets2DBD almost completely blocked PPC 1 cell invasion through Matrigel, whereas over-expression of full-length Ets2 did not inhibit invasion. Overall, these results demonstrate that the balance of Ets factor activity can regulate multiple aspects of the transformed phenotype of PPC 1 prostate tumor cells, including anchorage-independent growth, survival, and invasiveness. PMID- 11114729 TI - IGF-II/IGF-I receptor pathway up-regulates COX-2 mRNA expression and PGE2 synthesis in Caco-2 human colon carcinoma cells. AB - Nonsteroidal anti-inflammatory drugs reduce the risk of colon cancer and this effect is mediated in part through inhibition of type 2 prostaglandin endoperoxide synthase/ cyclo-oxygenase (COX-2). In the present study, we demonstrate that COX-2 expression and PGE2 synthesis are up-regulated by an IGF II/IGF-I receptor autocrine pathway in Caco-2 colon carcinoma cells. COX-2 mRNA and PGE2 levels are higher in proliferating cells compared with post-confluent differentiated cells and in cells that constitutively overexpress IGF-II. Up regulation of COX-2 expression by IGF-II is mediated through activation of IGF-I receptor because: (i) treatment of Caco-2 cells with a blocking antibody to the IGF-I receptor inhibits COX-2 mRNA expression; (ii) transfection of Caco-2 cells with a dominant negative IGF-I receptor reduces COX-2 expression and activity. Also, the blockade of the PI3-kinase, that mediates the proliferative effect of IGF-I receptor in Caco-2 cells, inhibits IGF-II-dependent COX-2 up-regulation and PGE2 synthesis. Moreover, COX-2 expression and activity inversely correlate with the increase of apoptosis in parental, IGF-II and dominant-negative IGF-I receptor transfected cells. This study suggests that induction of proliferation and tumor progression of colon cancer cells by the IGF-II/IGF-I receptor pathway may depend on the activation of COX-2-related events. PMID- 11114730 TI - Inhibition of farnesyltransferase increases TGFbeta type II receptor expression and enhances the responsiveness of human cancer cells to TGFbeta. AB - Several small GTPases of the Ras superfamily have been shown to antagonize TGFbeta signaling in human tumor cell lines. Some of these GTPases are post translationally modified by farnesylation, a lipid modification catalyzed by farnesyltransferase and required for the proteins to attach to membranes and to function. In this study, we investigated the effect of the farnesyltransferase inhibitor FTI-277 on TGFbeta-regulated cell growth and transcription. Treatment of the human pancreatic tumor cell line, Panc-1, with FTI-277 enhanced the ability of TGFbeta to inhibit both anchorage-dependent and -independent tumor cell growth. FTI-277 also enhanced the ability of TGFbeta to induce transcription, as measured by p3TP-lux reporter activity and collagen synthesis. The enhancement of TGFbeta responses by FTI-277 correlated with the stimulation of transcription and protein expression of type II TGFbeta receptor (TbetaRII). Consequently, FTI-277-treated cells exhibited a higher level of TGFbeta binding to its receptor. Thus, inhibition of protein farnesylation stimulates TbetaRII expression, which leads to increased TGFbeta receptor binding and signaling as well as inhibition of tumor cell growth and transformation. PMID- 11114731 TI - Identification of a novel human BCL-X promoter and exon. AB - BCL-XL is a key anti-apoptotic BCL-2 family protein that is widely expressed in human cancer cells and is induced in response to diverse survival signals. The translation initiation codon for BCL-XL is located in BCL-X exon II and previous analyses have indicated that BCL-XL RNAs initiate close to the start of exon II or additionally contain a non-coding first exon (exon IA) spliced to exon II. Using 5' RACE we have now identified a novel BCL-X non-coding exon (exon IB) which is spliced directly to exon II in place of exon IA. Exon IB-containing RNAs encoded BCL-XL and were detected in non-malignant lymphocytes and lymphoma cells from lymph node biopsies and were expressed at significant levels in cell lines derived from ovarian, colon and breast cancers. We identified two TATA-box sequences upstream of exon IB and demonstrated that surrounding genomic sequences contained strong promoter activity in lymphoma cells (approximately 300-fold active relative to controls). We have therefore identified a powerful new BCL-X promoter and a novel exon that contributes to BCL-XL expression. PMID- 11114732 TI - A role for FAK in the Concanavalin A-dependent secretion of matrix metalloproteinase-2 and -9. AB - To study the signaling pathway critical for the secretion of matrix metalloproteinases (MMPs), we examined the role of focal adhesion kinase (FAK) in Concanavalin A (Con A)-stimulated cells. We established a cell line in which FAK gene was conditionally inducible by use of FAK-null fibroblasts and the tetracycline repression system. In this cell line, FAK expression was undetectable in the presence of tetracycline but induced within 1 day by the removal of the drug. We found that FAK expression augmented the Con A-dependent secretion of MMP-9 and MMP-2. In contrast, proteolytic activation of MMP-2 by Con A-treatment did not require FAK expression. In addition, activation of MMP secretion and tyrosine phosphorylation of FAK by Con A, but not the proteolytic activation of MMP-2, required attachment of the cells to the extracellular matrix. Taken together, our results suggest that the FAK signaling pathway play a pivotal role in the secretion of MMPs. PMID- 11114734 TI - The protein tyrosine kinase family of the human genome. AB - As the sequencing of the human genome is completed by the Human Genome Project, the analysis of this rich source of information will illuminate many areas in medicine and biology. The protein tyrosine kinases are a large multigene family with particular relevance to many human diseases, including cancer. A search of the human genome for tyrosine kinase coding elements identified several novel genes and enabled the creation of a nonredundant catalog of tyrosine kinase genes. Ninety unique kinase genes can be identified in the human genome, along with five pseudogenes. Of the 90 tyrosine kinases, 58 are receptor type, distributed into 20 subfamilies. The 32 nonreceptor tyrosine kinases can be placed in 10 subfamilies. Additionally, mouse orthologs can be identified for nearly all the human tyrosine kinases. The completion of the human tyrosine kinase family tree provides a framework for further advances in biomedical science. PMID- 11114733 TI - Somatic mutations of fibroblast growth factor receptor 3 (FGFR3) are uncommon in carcinomas of the uterine cervix. AB - Germline mutations of the gene encoding human fibroblast growth factor receptor 3 (FGFR3) have been shown to be responsible for several related autosomal dominant forms of syndromic craniosynostosis and short limb dwarfism. Somatic activating mutations of FGFR3 were recently reported to occur in three of 12 (25%) uterine cervical carcinomas and nine of 26 (35%) bladder carcinomas, suggesting that constitutive activation of FGFR3 may be an important mechanism underlying the development and/or progression of these common epithelial malignancies. In order to investigate further a possible role for FGFR3 mutations in cervical carcinogenesis, we performed sequence-based mutational analysis of FGFR3 in 51 primary cervical carcinomas and seven cervical carcinoma-derived cell lines. The regions analysed (exons 7, 10, 13, 15, and 19) encompassed all previously described FGFR3 mutations. A single nucleotide substitution at codon 249, predicting a serine to cysteine amino acid substitution (S249C) in the FGFR3 extracellular domain, was identified in one primary tumor. Only wild type FGFR3 alleles were identified in the remaining tumors and cell lines. The S249C mutation is the only FGFR3 mutation described to date in cervical carcinomas. These findings suggest that while activating mutations of FGFR3 occur in cervical cancer, they may not be as common as initially reported. PMID- 11114735 TI - Connecting signaling and cell cycle progression in growth factor-stimulated cells. AB - A widely used model system to investigate cell proliferation is stimulation of serum-arrested cells with growth factors. Recent data suggest that there are two waves of growth factor-dependent signaling events required for a proliferative response. One is an acute burst of signaling, which occurs immediately after growth factor stimulation and lasts for 30 - 60 min. The other occurs in a different time frame (8 - 12 h post stimulation), and involves activation of cyclin dependent kinases (Cdks). In addition to a general overview of growth factor-dependent signaling, we present our 'two wave' hypothesis for how signaling and cell cycle progression are linked. PMID- 11114736 TI - Ligand discrimination by ErbB receptors: differential signaling through differential phosphorylation site usage. AB - The four members of the ErbB family of receptor tyrosine kinases (RTKs) mediate a variety of cellular responses to epidermal growth factor (EGF)-like growth factors, and serve as a model for the generation of both diversity and specificity in RTK signaling. Previous studies indicate that receptor-receptor interactions figure prominently in signaling through ErbB receptors. In addition to a role in receptor kinase activation, ligand-induced ErbB receptor homo- and heterodimerization is thought to account for the diversity of biological responses stimulated by EGF-like growth factors. Since each receptor has the potential to couple to different complements of signaling pathways, EGF-like ligands specify cellular response by dictating which pairs of receptors become activated. More recently evidence has been uncovered for ligand discrimination by individual ErbB receptor dimers; receptors appear to realize which ligand is binding and differentially respond through autophosphorylation site usage. These observations indicate that ligand stimulation of RTKs is not generic, and point to another layer in the ErbB signal diversification mechanism. The mechanistic implications of ligand discrimination are discussed. PMID- 11114737 TI - The contradictions of the insulin-like growth factor 1 receptor. AB - In recent years, the type 1 insulin-like growth factor receptor (IGF-IR) has emerged as a receptor that plays a very important role in the growth of cells, both in vivo and in vitro. The ability of the IGF-IR to induce mitogenesis and to promote survival of cells against a variety of apoptotic agents is well documented. Somewhat less known are other functions of the IGF-IR, like its ability to induce differentiation, to regulate cell size and to affect the organization of the cytoskeleton of cells. This review will focus on these lesser known functions of the IGF-IR. At the same time, we will emphasize how the IGF-IR can send contradictory signals, which depend on different domains of the receptor and the availability of downstream transducing molecules. PMID- 11114738 TI - Met receptor tyrosine kinase: enhanced signaling through adapter proteins. AB - The Met receptor tyrosine kinase is the prototypic member of a small subfamily of growth factor receptors that when activated induce mitogenic, motogenic, and morphogenic cellular responses. The ligand for Met is hepatocyte growth factor/scatter factor (HGF/SF) and while normal HGF/SF-Met signaling is required for embryonic development, abnormal Met signaling has been strongly implicated in tumorigenesis, particularly in the development of invasive and metastatic phenotypes. Following ligand binding and autophosphorylation, Met transmits intercellular signals using a unique multisubstrate docking site present within the C-terminal end of the receptor. The multisubstrate docking site mediates the binding of several adapter proteins such as Grb2, SHC, Crk/CRKL, and the large adapter protein Gab1. These adapter proteins in turn recruit several signal transducing proteins to form an intricate signaling complex. Analysis of how these adapter proteins bind to the Met receptor and what signal transducers they recruit have led to more substantial models of HGF/SF-Met signal transduction and have uncovered new potential pathways that may be involved into Met mediated tumor cell invasion and metastasis. PMID- 11114739 TI - The RET proto-oncogene in human cancers. AB - The activation of the RET proto-oncogene contributes to the development of human cancers in two different ways. Somatic rearrangements of RET with a variety of activating genes, which contribute to unscheduled expression and constitutive dimerization of the chimeric RET/PTC oncoproteins in thyroid follicular cells, are frequently found in radiation-induced papillary thyroid carcinomas. Germ-line mutations, mainly point mutations, that lead to constitutive activation of RET tyrosine kinase activity are responsible for the development of the inherited cancer syndrome, multiple endocrine neoplasia type 2. There appears to be a correlation between specific types of RET mutation and clinical phenotypes of the cancers involved. The biological effects and the signaling pathways induced by different forms of RET activation have been investigated in a variety of cultured cells as well as in genetically engineered animal models. The identification of RET mutations in most MEN 2 families (95%) has translated into improved care for MEN 2 patients. However, further investigation of the signaling pathways contributing to tumorigenesis in relevant tissues will eventually help us to develop novel strategies to prevent or to treat human papillary thyroid carcinomas, MEN 2 disease, as well as the sporadic cancers relevant to MEN 2 disease. PMID- 11114740 TI - Vascular endothelial growth factor receptors in the regulation of angiogenesis and lymphangiogenesis. AB - VEGFR-1 (Flt-1), VEGFR-2 (KDR) and VEGFR-3 (Flt4) are endothelial specific receptor tyrosine kinases, regulated by members of the vascular endothelial growth factor family. VEGFRs are indispensable for embryonic vascular development, and are involved in the regulation of many aspects of physiological and pathological angiogenesis. VEGF-C and VEGF-D, as ligands for VEGFR-3 are also capable of stimulating lymphangiogenesis and at least VEGF-C can enhance lymphatic metastasis. Recent studies have shown that missense mutations within the VEGFR-3 tyrosine kinase domain are associated with human hereditary lymphedema, suggesting an important role for this receptor in the development of the lymphatic vasculature. PMID- 11114741 TI - Focal adhesion kinase: a regulator of focal adhesion dynamics and cell movement. AB - Engagement of integrin receptors with extracellular ligands gives rise to the formation of complex multiprotein structures that link the ECM to the cytoplasmic actin cytoskeleton. These adhesive complexes are dynamic, often heterogeneous structures, varying in size and organization. In motile cells, sites of adhesion within filopodia and lamellipodia are relatively small and transient and are referred to as 'focal complexes,' whereas adhesions underlying the body of the cell and localized to the ends of actin stress fibers are referred to as 'focal adhesions'. Signal transduction through focal complexes and focal adhesions has been implicated in the regulation of a number of key cellular processes, including growth factor induced mitogenic signals, cell survival and cell locomotion. The formation and remodeling of focal contacts is a dynamic process under the regulation of protein tyrosine kinases and small GTPases of the Rho family. In this review, we consider the role of the focal complex associated protein tyrosine kinase, Focal Adhesion Kinase (FAK), in the regulation of cell movement with the emphasis on how FAK regulates the flow of signals from the ECM to the actin cytoskeleton. PMID- 11114742 TI - Eph receptors and ephrin ligands: embryogenesis to tumorigenesis. AB - Protein tyrosine kinase genes are the largest family of oncogenes. This is not surprising since tyrosine kinases are important components of signal transduction pathways that control cell shape, proliferation, differentiation, and migration. At 14 distinct members, the Eph kinases constitute the largest family of receptor tyrosine kinases. Although they have been most intensively studied for their roles in embryonic development, increasing evidence also implicates Eph family proteins in cancer. This review will address the recent progress in understanding the function of Eph receptors in normal development and how disregulation of these functions could promote tumorigenesis. PMID- 11114743 TI - Selected glimpses into the activation and function of Src kinase. AB - Since the discovery of the v-src and c-src genes and their products, much progress has been made in the elucidation of the structure, regulation, localization, and function of the Src protein. Src is a non-receptor protein tyrosine kinase that transduces signals that are involved in the control of a variety of cellular processes such as proliferation, differentiation, motility, and adhesion. Src is normally maintained in an inactive state, but can be activated transiently during cellular events such as mitosis, or constitutively by abnormal events such as mutation (i.e. v-Src and some human cancers). Activation of Src occurs as a result of disruption of the negative regulatory processes that normally suppress Src activity, and understanding the various mechanisms behind Src activation has been a target of intense study. Src associates with cellular membranes, in particular the plasma membrane, and endosomal membranes. Studies indicate that the different subcellular localizations of Src could be important for the regulation of specific cellular processes such as mitogenesis, cytoskeletal organization, and/or membrane trafficking. This review will discuss the history behind the discovery and initial characterization of Src and the regulatory mechanisms of Src activation, in particular, regulation by modification of the carboxy-terminal regulatory tyrosine by phosphatases and kinases. Its focus will then turn to the different subcellular localizations of Src and the possible roles of nuclear and perinuclear targets of Src. Finally, a brief section will review some of our present knowledge regarding Src involvement in human cancers. PMID- 11114744 TI - Role of Src expression and activation in human cancer. AB - Since the original identification of a transmissible agent responsible for the development of tumors in chickens, now known to be a retrovirus encoding the v src gene, significant progress has been made in defining the potential functions of its human homolog, SRC. The product of the human SRC gene, c-Src, is found to be over-expressed and highly activated in a wide variety of human cancers. The relationship between Src activation and cancer progression appears to be significant. Moreover, Src may have an influence on the development of the metastatic phenotype. This review discusses the data supporting a role for c-Src as a critical component of the signal transduction pathways that control cancer cell development and growth, and provides the rationale for targeting Src in drug discovery efforts. PMID- 11114745 TI - Regulation of cell death by the Abl tyrosine kinase. AB - The c-abl proto-oncogene encodes a protein tyrosine kinase that is distributed in the nucleus and the cytoplasm of proliferating cells. In the nucleus, c-Abl activity is negatively regulated by the retinoblastoma protein (RB) and positively regulated by DNA damage signals. Activation of the c-Abl kinase by DNA damage requires the function of ATM, which regulates cell cycle checkpoint, DNA repair and apoptosis in response to DNA damage. Cells lacking c-Abl can activate cell cycle checkpoints and DNA repair, but show defects in apoptosis. The apoptosis defect of c-Abl deficient cells is correlated with a defect in the induction and activation of p73, which is a functional homologue of the p53 tumor suppressor protein and has pro-apoptotic activity. The inhibition of c-Abl by RB is consistent with RB's ability to block apoptosis; while the activation of c-Abl by ATM is consistent with ATM's ability to activate cell death. The oncogenic Bcr Abl tyrosine kinase is a potent inhibitor of apoptosis, and it is retained exclusively in the cytoplasm of transformed cells. Interestingly, when Bcr-Abl is trapped inside of the nucleus through a combined disruption of its cytoplasmic retention and its nuclear export, this oncogenic Abl kinase induces apoptosis. Taken together, the current results support a role for the nuclear c-Abl tyrosine kinase in the regulation of apoptosis. Whether the cytoplasmic c-Abl kinase can actively inhibit apoptosis remains to be determined; however, a deliberate retention of c-Abl in the cytoplasm could potentially contribute to the attenuation of apoptosis response. PMID- 11114746 TI - Signaling network of the Btk family kinases. AB - The Btk family kinases represent new members of non-receptor tyrosine kinases, which include Btk/Atk, Itk/Emt/Tsk, Bmx/Etk, and Tec. They are characterized by having four structural modules: PH (pleckstrin homology) domain, SH3 (Src homology 3) domain, SH2 (Src homology 2) domain and kinase (Src homology 1) domain. Increasing evidence suggests that, like Src-family kinases, Btk family kinases play central but diverse modulatory roles in various cellular processes. They participate in signal transduction in response to virtually all types of extracellular stimuli which are transmitted by growth factor receptors, cytokine receptors, G-protein coupled receptors, antigen-receptors and integrins. They are regulated by many non-receptor tyrosine kinases such as Src, Jak, Syk and FAK family kinases. In turn, they regulate many of major signaling pathways including those of PI3K, PLCgamma and PKC. Both genetic and biochemical approaches have been used to dissect the signaling pathways and elucidate their roles in growth, differentiation and apoptosis. An emerging new role of this family of kinases is cytoskeletal reorganization and cell motility. The physiological importance of these kinases was amply demonstrated by their link to the development of immunodeficiency diseases, due to germ-line mutations. The present article attempts to review the structure and functions of Btk family kinases by summarizing our current knowledge on the interacting partners associated with the different modules of the kinases and the diverse signaling pathways in which they are involved. PMID- 11114747 TI - Janus kinases: components of multiple signaling pathways. AB - Cytoplasmic Janus protein tyrosine kinases (JAKs) are crucial components of diverse signal transduction pathways that govern cellular survival, proliferation, differentiation and apoptosis. Evidence to date, indicates that JAK kinase function may integrate components of diverse signaling cascades. While it is likely that activation of STAT proteins may be an important function attributed to the JAK kinases, it is certainly not the only function performed by this key family of cytoplasmic tyrosine kinases. Emerging evidence indicates that phosphorylation of cytokine and growth factor receptors may be the primary functional attribute of JAK kinases. The JAK-triggered receptor phosphorylation can potentially be a rate-limiting event for a successful culmination of downstream signaling events. In support of this hypothesis, it has been found that JAK kinase function is required for optimal activation of the Src-kinase cascade, the Ras-MAP kinase pathway, the PI3K-AKT pathway and STAT signaling following the interaction of cytokine/interferon receptors with their ligands. Aberrations in JAK kinase activity, that may lead to derailment of one or more of the above mentioned pathways could disrupt normal cellular responses and result in disease states. Thus, over-activation of JAK kinases has been implicated in tumorigenesis. In contrast, loss of JAK kinase function has been found to result in disease states such as severe-combined immunodeficiency. In summary, optimal JAK kinase activity is a critical determinant of normal transmission of cytokine and growth factor signals. PMID- 11114748 TI - Tyrosine kinases and gastric cancer. AB - Carcinoma of the stomach is one of the most prevalent cancer types in the world today. Two major forms of gastric cancer are distinguished according to their morphological and clinicopathological classifications (well differentiated/intestinal type and poorly differentiated/diffuse type), characteristics that could also be attributed to the altered expression of different types of oncogenes or tumor suppressor genes. Significant differences exist for gastric cancer incidence comparing people of different ethnic origins, implicating various genetic and epigenetic factors for gastric oncogenesis. There are only a limited number of molecular markers available for gastric cancer detection and prognostic evaluation, among which are tyrosine kinases. There is convincing evidence that tyrosine kinases are involved in oncogenesis and disease progression for many human cancers. Amplifications of certain tyrosine kinases (c met, k-sam and erbB2/neu) have been associated with human gastric cancer progression. Alternatively spliced transcripts and enhanced protein-expression levels for some of these tyrosine kinases are correlated with clinical outcomes for gastric cancer patients. With advent of high throughput techniques, it is now possible to detect nearly all expressed tyrosine kinases in a single screen. This increases the chance to identify additional tyrosine kinases as predictive markers for gastric cancers. In this article, we will first review the literature data concerning certain tyrosine kinases implicated in gastric carcinogenesis and then summarize more recent work which provide comprehensive tyrosine kinase profiles for gastric cancer specimens and cell lines. Two new gastric cancer molecular markers (tie-1 and mkk4) have been identified through the use of these profiles and demonstrated effective as clinical prognostic indicators. PMID- 11114749 TI - Development of inhibitors for protein tyrosine kinases. AB - In the last 5 years, through combinatorial chemistry, high-throughput screening, computational chemistry, and traditional medicinal chemistry, numerous inhibitors for various protein tyrosine kinases (PTKs) have been developed. The majority of these compounds are small molecules that compete at the ATP binding site of the catalytic domain of the enzymes. Some compounds such as pseudosubstrate-based peptide inhibitor binds to the peptide/protein substrate site of the catalytic domain. Some inhibitors, primarily monoclonal antibodies, bind to the extracellular domain of receptor tyrosine kinases. Some of these inhibitors are highly potent and selective. Several are currently undergoing clinical trials for a number of diseases such as cancer. PMID- 11114750 TI - The new NHS plan-a new direction for public health? PMID- 11114751 TI - Can health impact assessment fulfil the expectations it raises? AB - The United Kingdom and other European Governments are increasingly calling for Health Impact Assessment (HIA) of policies in order to predict how they will affect the health of populations. Approaches to HIA can be characterised as broad focus (holistic, sociological, qualitative) or tight focus (limited, epidemiological, quantitative). HIA must add value to decision making and lead to better decisions than would have otherwise been made. The quality of HIA will be judged on its utility, its predictive accuracy and its process. HIA must be closely integrated with the decision making process. HIA may be undertaken in combination with Environmental Impact Assessment (EIA) or separately. HIA does not mean that health should take primacy over other policy goals but does ensure that health is considered. PMID- 11114752 TI - Housing and health: does installing heating in their homes improve the health of children with asthma? AB - The objective of this study was to evaluate the use of NHS money to improve health by improving housing conditions. A pilot study assessing health outcomes before and after improving housing conditions was conducted, studying 72 children with previously diagnosed asthma living in 59 damp houses in Cornwall. The intervention was the installation of central heating. This improved the energy efficiency of the housing. The children's health was a symptom-based outcome measure for asthma and time lost from school. Improvements comprised installation of gas central heating in 28/59 (47%) houses, electric storage heaters in 22/59 (37%), solid fuel central heating in 7/59 (12%) and oil-fired central heating in 2/59 (4%) houses. Energy efficiency improved by a mean of 2.1 on the National Home Energy Rating scale (95% CI 1.68-2.47, P<0.001) in the 37/59 (62%) houses for which two readings were available. Initially, 69/72 (92%) children's bedrooms were unheated and 44/72 (61%) were damp; following improvements, the proportions were 10/72 (14%) and 15/72 (21%) respectively. All respiratory symptoms were significantly reduced after intervention; the greatest reduction was seen in nocturnal cough from a median score of 3 (most nights) to 1 (on one or several nights) (P<0.001) in the previous month. School-age children lost significantly less time from school for asthma in the previous 3 months (9.3 days per 100 school days before intervention and 2.1 days afterwards, P<0.01) but not for other reasons (1.4 days per 100 school days before and 3.2 after, P>0.05). In conclusion, this study provides the first evaluation of health outcomes following housing improvements. Lack of a comparison group means that effects of age, season and biased reporting cannot be eliminated. More work is needed to substantiate these results. PMID- 11114753 TI - Assessing health status and outcomes in a geriatric day hospital. AB - The study objective was to assess the feasibility and usefulness of recommended outcome measures in older people attending a geriatric day hospital for multidisciplinary assessment and rehabilitation. We used the 'Short Form 36' (SF36) questionnaire which had been proposed as a suitable outcome tool for the elderly, as well as standard assessment scales (eg Barthel index). These were administered by interviewers at the start of day hospital attendance and repeated by postal survey three and six months later. Change in overall health status was rated by the clinical team. The study took place in a geriatric day unit based in a support hospital, specialising in assessment and rehabilitation of older people. Participants were older people referred directly from the community, or following an inpatient day, whose assessment indicated a need for multidisciplinary rehabilitation. Stroke and musculo-skeletal disorders were the commonest underlying conditions. There was a high incidence of non-completion on SF36 questions relating to physical and mental function. Subsequent interviews showed that patients found some questions irrelevant. Floor effects were common. In contrast, the standard scales were invariably fully completed. Compared with local population survey data, respondents had low baseline scores on all SF36 dimensions. Differences over time were probably explained by varying methods of administration. In spite of a clinical perception of improved health status during day hospital attendance, both standard and SF36 scores showed overall deterioration. Two conclusions could be drawn from this study. 1. Measures of physical and mental disability and quality of life gave lower results than expected and continued declining over a six month period, even when the clinical team felt that the patient had improved. 2. Administration of SF36 by an interviewer is essential to obtain meaningful results in older people with poor physical health, which should be interpreted with caution. Goal-specific measures may be more useful in this group of patients. PMID- 11114754 TI - Variations in GP nursing home patient workload: results of a multivariate analysis. AB - The number of old people living in UK nursing homes has increased substantially over the past 15 y. There is evidence that such patients generate larger workloads for primary carers than do those of similar age and sex living in their own homes. Clearly, any extra workload involved in providing primary care services to nursing home patients, needs to be reflected in the resources afforded general practitioners (GPs) who are tasked with its provision. By the same token variations in workloads between patients need to be examined and explained for any insights these might provide on funding issues. To examine and explain variations in GP workload associated with nursing home patients and determine the implications of these for GP funding, a 12 month case control study of all nursing home residents over 65 y old registered with nine general practices was undertaken. A multivariate regression analysis was used to examine variations in GP workload associated with 270 nursing home patients. Multivariate regression models explaining the variation in workload cost per month in terms of the GP practice delivering care and patients age and sex had little explanatory power (R(2)=0.07). A fuller method including the patient's Barthel score and initial diagnosis as additional explanatory variables added little to the explanatory power of the model (R(2)=0.12). The ability of the multivariate models used here to explain the variation in GP workload was poor. GPs may require an allowance to compensate for differences in workload associated with nursing home patients but adjusting these payments for differences in age, sex, initial diagnosis or the other variables included in this analysis would not appear to be supported. PMID- 11114755 TI - Height and cancer mortality: results from the Glasgow University student cohort. AB - The aim of the study was to examine the association between height and cancer mortality in a socially homogeneous group of subjects. The study was based on a cohort of students, 8397 men and 2329 women, aged 16-30 y, who attended the University of Glasgow between 1948 and 1968. Mean follow-up time was 40 y. Height was measured at a medical examination performed at the student health service. The outcome measures used in the study were all-cause mortality and mortality from: all cancers, smoking and non-smoking related cancers and cancers related to sex hormones. No substantial or statistically significant associations were seen between height and all-cause or all-cancer mortality in either sex. Neither were any significant associations found between height and any of the sub-types of cancer studied (ie those related to smoking, those not related to smoking, and those related to sex hormones). Previous observations which have shown positive associations between height and cancer mortality have generally been based on populations with diverse social origins, among whom the variation in height will reflect variation in health and nutrition in childhood. The relatively low level of such variation in the present study may account for the negative findings. Public Health (2000) 114, 451-455 PMID- 11114756 TI - The anaphylaxis problem in children: community management in a UK National Health Service District. AB - From an analysis of a database of children who have been prescribed injectable adrenaline we describe our experience of the management of children with severe anaphylactic reactions to food products for the period from March 1994 to August 1999. Notifications to our department increased from 7 in the 1981-83 birth cohort to 48 in the 1990-92 birth cohort. One hundred and forty-four children living in our health district have been prescribed injectable adrenaline for emergency use. A standard protocol agreed with the Local Education Authority to support these pupils in school and to work to provide a safe environment has been in place since April 1995. Almost half the schools in the area covered by our health district have at least one affected child. Community child health staff have played a pivotal role in providing liaison between health and education professionals. PMID- 11114757 TI - Smoking habits-a question of trend or unemployment? A comparison of young men and women between boom and recession. AB - The increased unemployment rates during the 1990s were followed by decreased cigarette consumption. The aim of this study was to analyse the association between unemployment and smoking habits among young men and women during times of prosperity and recession. Two groups of final-year pupils were surveyed five years after leaving school, at the age of 21, in 1986 (boom) and 1994 (recession). The boom group included 1083 pupils; the recession group 898 pupils. The non-response rate was 2% in the boom group and 10% in the recession group. Daily tobacco use was measured through self-administered questionnaires. Daily cigarette smoking was of a lower magnitude during the recession (9.7% among men and 21.9% among women) compared to the boom (19.8% and 37.8%, respectively). A low level of education, and among women also financial problems and motherhood, were associated with more frequent smoking. Unemployment was associated with tobacco consumption, especially among women and during the boom. Thus, smoking habits were found to be a question of both unemployment and tobacco trends in society. PMID- 11114758 TI - Work stress, smoking habits and competence in supporting clients to cease smoking -a survey among Finnish nurses. AB - This paper describes results of a survey of Finnish nurses (n=882), their views of themselves as employees, their experiences of work stress and their competence to guide clients in smoking cessation. Nurses' skills to guide clients were fairly good but they had a lack of knowledge of smoking cessation centers and nicotine substitutions. The more positive the nurses' views were of themselves as employees, and the less they had experienced work stress related to their clients, the better they evaluated their skills and knowledge to guide clients to cease smoking. The results can be used to develop nurse education by providing a stronger knowledge base of smoking cessation. Nurses' view of themselves as employees could be strengthened by providing them with positive feedback from colleagues and managers. PMID- 11114759 TI - Underestimation of body mass index through perceived body image as compared to self-reported body mass index in the European Union. AB - An observational, cross-sectional study was conducted in a representative sample of the European Union (7155 men and 8077 women) to calculate the underestimation of body weight as assessed by body image among the overweight and obese population and identify the associated factors to this behavior. Participants were older than 15 years and they were living in the 15 European Union countries. Body mass index (BMI) was grouped into 4 categories using the cutpoints established by the WHO, while perceived body image (PBI) was assessed using a nine-silhouettes drawing. The degree of underestimation between PBI as compared to BMI was identified in overweight and obesity categories of BMI. A multivariable logistic regression model for each gender was used to adjust for potentially confounding variables. Men classified themselves worse than women, being more likely to underestimate their body weight (65.2% of men underestimated their weight vs 32.2% women), regardless of other socioeconomic and attitudinal variables. The greatest degree of underestimation was observed in Mediterranean individuals (68.7% of men and 37.9% of women underestimated their weight). The subjects in the 'maintenance' stage of physical activity tended more often to wrongly select their actual image (71.8% for men and 38.7% for women). PMID- 11114760 TI - Reliability of a physical activity questionnaire in middle-aged men. AB - Although some items from a questionnaire used to assess occupational and leisure activity in a cohort of male factory workers have reasonable validity, their reliability is unknown. In this study the reliability of this questionnaire was assessed in 54 middle-aged male factory employees by test-retest administration over a 4-6 week period. Kappa statistics for test-retest agreement for individual items on the questionnaire ranged between 0.24 and 0.66. Those items that demonstrated reasonable reliability and validity will subsequently be related to a range of disease endpoints in a prospective cohort study. PMID- 11114761 TI - Seroprevalence of HIV infection among pregnant women in the Veneto region (north east Italy). AB - A heterogeneous population of 4396 consecutive pregnant women (86.6% indigenous, 13.4% immigrants) attending the Department of Obstetrics and Gynecology of the University of Padua (north-east Italy) were counselled and tested for HIV infection between September 1995 and December 1997. Sociodemographic and sanitary data were collected on each case. Anti-HIV prevalence was 0.57%. Intravenous drug use and foreign birth accounted for 28% and 24%, respectively, of the anti-HIV positive cases; 44% of the HIV-positive subjects reported no risk factors. In the logistic regression HIV positivity proved independently associated with intravenous drug use (adjusted OR 76. 6), sexually transmitted diseases (adjusted OR 13.2), unmarried status (adjusted OR 4.8), birth outside the European Union (EU) (adjusted OR 3.1) and age (adjusted OR 1.1). Heterosexual HIV spread appears to be a major concern. The monitoring of trends in HIV infection among subgroups should be continued in order to control the AIDS epidemic appropriately both by promoting HIV counselling and individual care, and by watching for changes in the social background. PMID- 11114762 TI - Malaria outbreak in a malaria-free region in Oman 1998: unknown impact of civil war in Africa. AB - Beginning in April 1998, the surveillance system in Dhofar region, Oman, detected malaria cases among individuals who had no risk factors for the acquisition of malaria. An investigation was conducted to describe the outbreak and to identify its possible causes. A malaria case was defined as an unexplained fever (>38 degrees C) in a resident of the Dhofar region from April to September 1998. The investigation consisted of enhanced passive case detection, active case finding through contact screening, mass blood survey and school survey. Also an entomological survey was conducted and meteorological data was reviewed. Over a period of seven months, 1279 patients with fever were examined for malaria parasites. Sixty-five cases were positive; 60 (92%) males and 5 (8%) females. Cases occurred in all age groups (range: 2-63 years, median 25 years). Most cases were among illegal Somali immigrants (28, 43%) followed by Omanis (20, 31%). Out of the 2323 slides collected from the community and 2487 from school children, 21 slides were positive. All of them were from illegal immigrants. The entomological survey detected three vectors, previously found in the region: A. d'thali, A. sergenti and A. stephensi. Although the region is classified as a malaria-free region, it has the potential for malaria introduction. This outbreak most likely occurred due to the influx of hundreds of illegal Somali immigrants due to the civil war into the Dhofar region, providing a sufficient number of gametocyte carriers for local anopheline mosquitoes to feed on. PMID- 11114763 TI - Rabies in Israel: decades of prevention and a human case. AB - Animal rabies is endemic in Israel, with 50-80 laboratory-confirmed cases being diagnosed annually. Despite the high incidence among animals, human rabies has not occurred in Israel for almost four decades. This is likely due to the highly effective prevention policy implemented by the Ministry of Health, based on pre exposure vaccination of populations at risk, post-exposure treatment, and updated rules. Notwithstanding the previous success, a human case occurred in 1996 when a soldier was bitten, while asleep, by an unidentified small animal, which according to his description was a rat or a mouse. Since injuries by these rodents do not require antirabies treatment, no antirabies post-exposure prophylaxis was administered. Five weeks later the soldier complained of fever and nausea with interchanging periods of rage and calm, confusion, and water aversion. His condition deteriorated gradually, leading to deep coma and death. Immunofluorescence examination of a skin biopsy was positive for rabies, and PCR of saliva revealed Lyssavirus genotype 1. We review the changes in the epizootiology of rabies in Israel, the trends of human exposure to animals, and the pre- and post-exposure prophylaxis guidelines, and discuss possible measures that could have been undertaken to prevent the eventuality of this case. This case of rabies, the first after a long period without human disease, accentuates the importance of strict adherence to prevention guidelines. Considerations of geography, epidemiology, and the circumstances of exposure are crucial in the treatment decision-making process. PMID- 11114764 TI - Recommendations for water supply in arsenic mitigation: a case study from Bangladesh. AB - Arsenic problems have been observed in several countries around the world. The challenges of arsenic mitigation are more difficult for developing and poor countries due to resource and other limitations. Bangladesh is experiencing the worst arsenic problem in the world, as about 30 million people are possibly drinking arsenic contaminated water. Lack of knowledge has hampered the mitigation initiatives. This paper presents experience gained during an action research on water supply in arsenic mitigation in rural Singair, Bangladesh. The mitigation has been implemented there through integrated research and development of appropriate water supply options and its use through community participation. Political leaders and women played key roles in the success of the mitigation. More than one option for safe water has been developed and/or identified. The main recommendations include: integration of screening of tubewells and supply of safe water, research on technological and social aspects, community, women and local government participation, education and training of all stakeholders, immediate and appropriate use of the available knowledge, links between intermediate/immediate and long term investment, effective coordination and immediate attention by health, nutrition, agriculture, education, and other programs to this arsenic issue. PMID- 11114765 TI - Absolute poverty and child health in India. PMID- 11114769 TI - Ambrose J king TD KSG MB BS FRC PMID- 11114770 TI - Preserving transfer independence among individuals with spinal cord injury. AB - STUDY DESIGN: Literature review. OBJECTIVES: Upper extremity (UE) joint degeneration, particularly at the shoulder, detrimentally influences functional independence, quality of life, cardiovascular disease risk, and life expectancy of individuals following spinal cord injury (SCI). This review (1) describes UE use for transfers among individuals with SCI; (2) describes contributing factors associated with UE joint degeneration and loss of transfer independence; (3) summarizes and identifies gaps in existing research; and (4) provides suggestions for future research. RESULTS: Investigations of wheelchair transfer related UE joint and function preservation among individuals with SCI should consider factors including age and length of time from SCI onset, interface between subject-wheelchair, pain, shoulder joint range of motion (ROM) and muscle strength deficiencies or imbalances, exercise capacity and tolerance for the physical strain of activities of daily living (ADL), body mass and composition, previous UE injury or disease history, and transfer techniques. Existing studies of transfers among individuals with SCI have relied on small groups of either asymptomatic or non-impaired subjects, with minimal integration of kinematic, kinetic and electromyographic data. Descriptions of UE joint ROM, forces, and moments produced during transfers are lacking. CONCLUSIONS: Biomechanical measurement and computer modeling have provided increasingly accurate tools for acquiring the data needed to guide intervention planning to prevent UE joint degeneration and preserve function among individuals with SCI. The identification of stressful sub-components during transfers will enable intervening clinicians and engineers who design and modify assistive and adaptive devices to better serve individuals with SCI. PMID- 11114771 TI - Chronic pain after spinal cord injury: a survey of practice in spinal injury units in the USA. AB - OBJECTIVE: To determine the current practice regarding assessment and management of patients with chronic pain after spinal cord injury (SCI) in the United States of America (USA). METHODS: A postal questionnaire sent to the medical directors of 12 spinal injury units in the USA. RESULTS: A response was received from eight of the 12 units. Chronic pain was considered a significant problem amongst patients with SCI. There was inconsistency of opinion regarding prevalence estimates, investigation and management of chronic pain after SCI; but classification systems for pain were remarkably similar amongst units. Most felt that there was a need for further information, although only one unit said it was presently conducting research into the subject. CONCLUSION: Our survey has demonstrated the uncertainty that exists amongst USA specialists dealing with pain after SCI, and strengthens the case for more research into the subject with a view to developing guidelines for care. PMID- 11114772 TI - Silent hydronephrosis/pyonephrosis due to upper urinary tract calculi in spinal cord injury patients. AB - STUDY DESIGN: A study of four patients with spinal cord injury (SCI) in whom a diagnosis of hydronephrosis or pyonephrosis was delayed since these patients did not manifest the traditional signs and symptoms. OBJECTIVES: To learn from these cases as to what steps should be taken to prevent any delay in the diagnosis and treatment of hydronephrosis/pyonephrosis in SCI patients. SETTING: Regional Spinal Injuries Centre, Southport, UK. METHODS: A retrospective review of cases of hydronephrosis or pyonephrosis due to renal/ ureteric calculus in SCI patients between 1994 and 1999, in whom there was a delay in diagnosis. RESULTS: A T-5 paraplegic patient had two episodes of urinary tract infection (UTI) which were successfully treated with antibiotics. When he developed UTI again, an intravenous urography (IVU) was performed. The IVU revealed a non-visualised kidney and a renal pelvic calculus. In a T-6 paraplegic patient, the classical symptom of flank pain was absent, and the symptoms of sweating and increased spasms were attributed to a syrinx. A routine IVU showed non-visualisation of the left kidney with a stone impacted in the pelviureteric junction. In two tetraplegic patients, an obstructed kidney became infected, and there was a delay in the diagnosis of pyonephrosis. The clinician's attention was focused on a co existent, serious, infective pathology elsewhere. The primary focus of sepsis was chest infection in one patient and a deep pressure sore in the other. The former patient succumbed to chest infection and autopsy revealed pyonephrosis with an abscess between the left kidney and left hemidiaphragm and xanthogranulomatous inflammation of perinephric fatty tissue. In the latter patient, an abdominal X ray did not reveal any calculus but computerised axial tomography showed the presence of renal and ureteric calculi. CONCLUSIONS: The symptoms of hydronephrosis may be bizarre and non-specific in SCI patients. The symptoms include feeling unwell, abdominal discomfort, increased spasms, and autonomic dysreflexia. Physicians should be aware of the serious import of these symptoms in SCI patients. PMID- 11114773 TI - Late onset Pott's paraplegia. AB - BACKGROUND: Pott's disease may cause late neurological involvement due to development of sharp kyphosis. Anterior decompression and fusion is the treatment of choice for this disorder. OBJECTIVE: To determine the mid-term clinical results of patients with late onset Pott's paraplegia, who underwent anterior decompression and grafting after neurological deterioration. SETTING: A university hospital in Istanbul, Turkey. METHODS: Eight patients who developed late onset paraplegia with a mean period of 24.6 years (range, 9-46 years) after the active disease were treated with anterior decompression and grafting. The mean age at surgery was 36.1 years (range, 18-63 years) and the mean duration of neurological deterioration before surgery was 7.4 weeks (range, 2-13 weeks). The mean kyphosis angle of the patients was 105.63 degrees (range, 80 degrees- 135 degrees). No attempt to correct the curve was made in any operation. All but two patients' neurological status were evaluated according to the International Standards for Neurological and Functional Classification of Spinal Cord Injury determined by ASIA-IMSOP on admission. RESULTS: Neurological status of all patients showed progression either in Frankel scale or in motor scores in the early postoperative period. One patient needed to be reoperated on because of a deterioration of neurological status 26 months after surgery. The mean length of time since the operations is 75.9 months (range, 48 173 months) and all the patients are carrying out their lives independently with a mean motor score of 97.5 and full pin-prick and light touch scores. CONCLUSIONS: Anterior decompression and grafting is an effective procedure for the treatment of late onset paraplegia in Pott's disease. PMID- 11114774 TI - Inter-rater reliability of the 1992 international standards for neurological and functional classification of incomplete spinal cord injury. AB - AIMS: To determine the inter-rater reliability in scoring sensory and motor function and in defining sensory and motor levels in incomplete spinal cord injury, using the revised 1992 International Standards for Neurological and Functional Classification of Spinal Cord Injury (ISCSCI-92) and to determine the effect on raters agreement of one standardising assessment. METHODS: Two physicians and two physiotherapists at the Spinal Cord Injury Unit, Karolinska Hospital, classified 23 patients according to the ISCSCI-92. Kappa values were calculated. RESULTS: Kappa values varied from 0 to 0.83 (poor to very good) for the pin-prick scores, from 0 to 1 for the light touch scores and from 0 to 0.89 for motor function after the standardising assessment. Kappa values for sensory and motor levels were fair to poor after the standardising assessment. The results showed improvement in degree of agreement in 35/46 dermatomes for scoring pin-prick, in 15/42 for light touch, in 14/19 segments for motor function and for three out of four sensory and motor levels. CONCLUSION: This study indicates a weak inter-rater reliability for scoring incomplete SCI lesions using the ISCSCI 92. PMID- 11114775 TI - FES cycling may promote recovery of leg function after incomplete spinal cord injury. AB - STUDY DESIGN: Single subject pilot. OBJECTIVES: (i) To see whether strength and endurance for recreational cycling by functional electrical stimulation (FES) are possible following spinal cord injury (SCI). (ii) To develop the equipment for FES-cycling. SETTING: England. METHODS: Near-isometric or cycling exercise was performed by the incomplete SCI subject at home. RESULTS: After training for an average of 21 min per day for 16 months, the stimulated muscles increased in size and the subject was able to cycle for 12 km on the level. Surprisingly, there was a substantial increase in the measured voluntary strength of the knee extensors and the subject reports improved leg function. CONCLUSION: FES-cycling may promote recovery after incomplete spinal cord injury. If so, it offers the possibility of being a convenient method for widespread use. PMID- 11114776 TI - Three-dimensional measurement of rolling in tetraplegic patients. AB - OBJECTIVE: To investigate the differences between traumatic tetraplegic patients who can roll and those who cannot. DESIGN: Motion analysis using 3-dimensional measurement. SETTING: Rehabilitation centers in southwestern Japan. PARTICIPANTS: Nineteen male participants, all of whom had traumatic C6 complete injury. METHODS: We used an electromagnetic device to examine the degree of spinal movement in axial rotation during rolling (shifting from supine to side lying). This system (3-Space Win) measures the position and orientation of sensors in space. Two sensors were mounted on a subject over the spinous process of T1 and L5. RESULTS: The spinal rotation of patients who could not roll was significantly lower than that of patients who could roll. (The average rotation of non-rollers was 31.5+/-17.5 degrees, while the average rotation of rollers was 66.3+/ 17.3degrees). In this study, there were no statistically significant differences in the members of the two groups in terms of age, height, weight or time after injury. CONCLUSION: Rolling requires greater and adequate flexibility in the back of tetraplegic patients. PMID- 11114777 TI - Coping with spinal cord injury: personal and marital adjustment in the Hong Kong Chinese setting. AB - STUDY DESIGN: A cross-sectional retrospective study was carried out with structured questionnaires and semi-structured interviews on 66 persons with spinal cord injury (SCI) and 40 spouses. OBJECTIVES: The study aimed to explore the psychosocial adjustment of Hong Kong Chinese couples at the post SCI stage. An important study interest was the impact of care-giving in spouses of persons with SCI. SETTING: Three major regional rehabilitation centres and one community resource centre in Hong Kong. METHODS: A set of psychometric measures tapping different aspects of psychological functioning was included. These were locus of control (Levenson's Internality, Powerful Others, and Chance Scale), perceived social support (Provision of Social Relationship), coping strategies (Ways of Coping Checklist), marital adjustment (Dyadic Adjustment Scale), caregiving burden (Caregiver Burden Inventory), depression (Beck Depression Inventory), life satisfaction (Satisfaction with Life Situation), and social role adjustment (Katz Adjustment Scale - Relative Form). RESULTS: Persons with SCI with pre-injury marriage were more depressed (P<0.05) as compared with those with post-injury marriage. However, the two groups did not differ in terms of satisfaction with life situation and social role dissatisfaction. The spouses in the preinjury marriage reported a significantly higher score in time-dependent burden than those in the post-injury marriage (P<0.05). Care-giving burden was associated with locus of control, social support, and modes of coping (P<0.05). CONCLUSION: The impact of SCI is a long-lasting effect not limited to the patients but also extending to their spouses. Findings from the adjustment outcomes and coping styles of persons with SCI and their spouses indicate that they are not passive victims. A similar injury may produce different outcomes in different individuals. Rehabilitation professionals should thus be alert to both the couple's differing needs and idiosyncrasies in their helping process. PMID- 11114778 TI - Traumatic spinal cord injuries in Turkey: a nation-wide epidemiological study. AB - STUDY DESIGN: An epidemiological study conducted all over the country. OBJECTIVE: The present retrospective study was conducted to survey the new traumatic spinal cord injury (SCI) cases during 1992 in Turkey. SETTING: Intensive care units, emergency services and departments of orthopaedic surgery, neurosurgery and rehabilitation of state hospitals, rehabilitation centers, military and university hospitals. METHODS: Postal questionnaires were used for data collection and the records from medical institutes nation-wide were reviewed for the analysis of the epidemiological factors. RESULTS: Five hundred and eighty-one new traumatic SCI cases were reported in 1992. The annual incidence was found to be 12.7 per million population. Male to female ratio was 2.5:1 and the average age at injury was 35.5+/-15.1 (35.4+/-14.8 for males and 35.9+/-16.0 for females). The most common cause of injury was motor vehicle accidents (48.8%) followed by falls (36.5%), stab wounds (3.3%), gunshot injuries (1.9%) and injuries from diving (1.2%). One hundred and eighty-seven patients (32.18%) were tetraplegic and 394 patients (67.8%) were paraplegic. The most common level of injury was C5 among tetraplegics and T12 among paraplegics. The most prevalent associated injury was head trauma followed by extremity fractures. Severe head trauma resulting in death may obscure the real incidence of SCI and may cause underreporting of cases in epidemiological studies. CONCLUSION: Considering that motor vehicle accidents and falls were found to be the leading causes of traumatic SCI, it was concluded that the prevention measures should be focused mainly on these in order to reduce the frequency of SCI in Turkey. PMID- 11114779 TI - Paraplegia caused by painless acute aortic dissection. AB - OBJECTIVES: Painless acute aortic dissection in which paraplegia is the only presenting sign is rare, with limited reported cases. CASE REPORT: The authors report a patient with painless acute aortic dissection who presented with sudden onset paraplegia. Ischemic diseases of the spinal cord were suspected as the cause. MRI revealed extensive acute aortic dissection with an intramural hematoma. The patient was treated conservatively by strictly controlling his blood pressure. The treatment was successful, although the motor function of the lower extremities could not be rescued. Although 3% to 5% of patients with acute aortic dissection present with paraplegia as a result of spinal cord infarction, most of these patients experience severe pain prior to presentation. CONCLUSION: Painless acute aortic dissection in which paraplegia is the only presenting sign is very rare. However, aortic diseases, including acute aortic dissection, should always be considered as a differential diagnosis of patients with sudden onset, painless paraplegia. PMID- 11114780 TI - Brown-Sequard syndrome associated with Horner's syndrome after a penetrating trauma at the cervicomedullary junction. AB - STUDY DESIGN: Case report of a 21-year-old man that had concurrence of Brown Sequard syndrome and Horner's syndrome after a penetrating trauma in the neck. OBJECTIVES: This report analyzes the location of lesions that cause a combination of Horner's and Brown-Sequard syndrome. It is important to know the anatomic structure of spinal cord and the sympathetic nerve chain. SETTING: Spinal Cord Unit, Department of Physical Medicine and Rehabilitation, Hospital La Fe, Valencia, Instituto Oftalmologico de Alicante, Alicante, Spain. METHODS: Description of a single patient case report. RESULTS: The clinical findings and MRI showed a good correlation. The Horner's syndrome was confirmed with a 4% cocaine test. The patient received a conservative treatment with high-dose steroid therapy (NASCIS-3). CONCLUSION: The patient presented with Brown-Sequard syndrome and Horner's syndrome. Clinical examination and MRI made a quick and correct diagnosis. The patient recovered completely after the conservative treatment. PMID- 11114781 TI - A gas filled intradural cyst associated with disc degeneration. AB - A case of a lumbar intradural, extramedullary gas filled cyst is described. This was associated with degenerative disc disease and presented with radicular signs and symptoms. The radiological differential diagnosis of such a mass includes infection and tumours. The association of intradural gas with degenerative disc disease is previously reported. This case further illustrates the association of degenerative disc disease with an intradural gas filled cyst and provides an interesting radiological differential diagnosis for an extramedullary intradural mass. PMID- 11114782 TI - Sacral meningeal arteriovenous fistula supplied by branches of the hypogastric artery revealed by conus medullaris infarction. AB - OBJECTIVE: Spinal dural arteriovenous fistulas (DAVF), the most common vascular malformations of the spine, are usually supplied by branches of the intercostal or lumbar arteries. Rarely, the DAVF are fed by branches of the hypogastric artery. Only 12 such cases have been reported. CASE REPORT: A 28 year-old man presented with a 2-month history of micturition dysfunction and progressive weakness of the legs. Physical examination showed motor deficit of the lower limbs with brisk knee jerks, absent ankle reflexes and normal plantar reflexes. Cremasteric reflexes were absent. We noted hypoesthesia of the lower limbs with complete anesthesia of the perineum. MRI of the lumbo-sacral spine demonstrated an enlargement of the conus medullaris with high T2 signal intramedullary lesion. It showed also large intradural serpentine vessels. A left iliac angiogram disclosed a nidus of arteriovenous malformation (AVM) supplied by a lateral sacral artery and draining by two enlarged ascending perimedullary veins. No clinical improvement was observed after surgical removal of the AVM. CONCLUSION: The screening examination of choice for spinal DAVF remains MRI. When selective spinal arteriography is normal, we have to search for an unusual arterial supply particularly from the hypogastric artery. PMID- 11114783 TI - Inhalation of poorly soluble particles. I. Differences in inflammatory response and clearance during exposure. AB - Results from animal studies have indicated some uncertainties over the validity of a single general occupational control limit for all types of "particulates (insoluble) not otherwise classified" (PNOC) (ACGIH, 2000). Therefore, to examine the extent to which a given control limit may be valid for nontoxic dusts with different physical characteristics, this study compared the pulmonary effects in rats of inhalation exposure to two poorly soluble dusts of similar density and with relatively low toxicity: titanium dioxide and barium sulfate. The objectives were to compare the dusts in (a) their buildup and clearance in the lungs during inhalation; (b) their transfer to lymph nodes; (c) the changes, with time, in the lavageable cell population; and (d) the pathological change from histology. The exposure aerosol concentrations were selected to achieve similar mass and volume lung burdens for both dusts and to attain "overload" over the common exposure periods of about 4 mo and 7 mo. Despite obtaining similar lung burdens for both dusts, there was significantly more translocation of TiO(2) to the hilar lymph nodes than with BaSO(4). It was also found that clearance of TiO(2) was retarded whereas clearance of BaSO(4) was not. Trends in these data were clarified by the use of a simple model of particle clearance. Retardation of particle clearance and translocation to the lymph nodes are markers of the condition known as "overload" in which the alveolar macrophage-based clearance of particles from the deep lung is impaired. In addition, bronchoalveolar lavage showed that TiO(2) caused significantly more recruitment of inflammatory neutrophils to lungs than BaSO(4). These differences between the dusts were not due to differences in toxicity, solubility, or lung deposition. The explanation that the different responses are due to the different particle size distributions of the two dust types is examined in a companion paper (Tran et al., this issue). PMID- 11114784 TI - Inhalation of poorly soluble particles. II. Influence Of particle surface area on inflammation and clearance. AB - In this article the volumetric overload hypothesis, which predicts the impairment of clearance of particles deposited in the lung in terms of particle volume, is reevaluated. The degree to which simple expressions of retained lung burden explain pulmonary responses to overload was investigated using data from a series of chronic inhalation experiments on rats with two poorly soluble dusts, titanium dioxide and barium sulfate. The results indicated that the difference between the dusts in the level of inflammation and translocation to the lymph nodes could be explained most simply when the lung burden was expressed as total particle surface area. The shape of the statistical relationship for both lung responses indicated the presence of a threshold at approximately 200-300 cm(2) of lung burden. On the basis of this and other similar results, a hypothesis regarding a generic mechanism for the impairment of clearance and associated lung responses is proposed for such "low-toxicity" dusts. PMID- 11114785 TI - Estimating rock and slag wool fiber dissolution rate from composition. AB - A method was tested for calculating the dissolution rate constant in the lung for a wide variety of synthetic vitreous silicate fibers from the oxide composition in weight percent. It is based upon expressing the logarithm of the dissolution rate as a linear function of the composition and using a different set of coefficients for different types of fibers. The method was applied to 29 fiber compositions including rock and slag fibers as well as refractory ceramic and special-purpose, thin E-glass fibers and borosilicate glass fibers for which in vivo measurements have been carried out. These fibers had dissolution rates that ranged over a factor of about 400, and the calculated dissolution rates agreed with the in vivo values typically within a factor of 4. The method presented here is similar to one developed previously for borosilicate glass fibers that was accurate to a factor of 1.25. The present coefficients work over a much broader range of composition than the borosilicate ones but with less accuracy. The dissolution rate constant of a fiber may be used to estimate whether disease would occur in animal inhalation or intraperitoneal injection studies of that fiber. PMID- 11114786 TI - A retrospective review of the carcinogenicity of refractory ceramic fiber in two chronic fischer 344 rat inhalation studies: an assessment of the MTD and implications for risk assessment. AB - The purpose of this article is to review previous chronic inhalation studies in rats with refractory ceramic fiber (RCF), the mathematical modeling efforts to describe the deposition, clearance, and retention of RCF fiber in the rat and human, and the concept of "overload," and to assess the possibility that the maximum tolerated dose (MTD) was exceeded. Lastly, based on recent biopersistence and pulmonary clearance studies of several investigators with a particulate-free RCF, we examine the potential impact on the chronic RCF rat bioassay of coexposure to both RCF particulate and RCF fibers. The review concludes, inter alia, that RCF particulate coexposure probably had a major impact on the observed chronic adverse effects, that the MTD was probably exceeded at the highest exposure concentration of 30 mg/m(3) in the rat bioassay, and that inclusion of the highest dose in the risk assessment process may overstate human health risk if a linear rather than nonlinear model is used. PMID- 11114787 TI - Polycyclic aromatic hydrocarbon levels and mutagenicity of inhalable particulate matter in Santiago, Chile. AB - The air in Santiago, Chile, is among the most highly polluted in the world. Due to the high levels of pollutants and the high incidence of respiratory diseases, especially in the most susceptible groups, Santiago has been declared a saturated zone for PM(10), O(3), and CO. The aim of this work was to investigate polycyclic aromatic hydrocarbon levels and mutagenic activity of Santiago s fine and coarse fractions of inhalable particles. The levels of 16 polycyclic aromatic hydrocarbons (PAHs) were determined by high-performance liquid chromatography in organic extracts from respirable particles (OERP). Respirable particulate matter (fine and coarse) contains high levels of PAHs including six mutagenic ones classified by the IARC as carcinogenic, which represented at least 45% of the total PAH concentration. A seasonal effect was observed, with higher values in months with lower temperatures. Although a significant decline of PAH levels in OERP was observed in the last years, the levels of carcinogenic PAHs are still higher than those reported in cities of the United States, Australia, and Europe. OERP were highly mutagenic and contained direct and indirect mutagens, which produced both frameshift and base substitution mutations in Salmonella typhimurium. In addition, organic extracts from total suspended particles were also highly mutagenic at the tk locus in h1A1v2 human lymphoblasts in culture. In spite of the important decrease in PAHs in the period 1991-1996, direct mutagenic response has not changed significantly, suggesting that the levels of direct mutagenic pollutants (e.g., nitroarenes) have not decreased considerably during the last years. These results suggest a risk for Santiago s inhabitants since pollutants adsorbed in inhalable particles are highly mutagenic and can damage DNA. PMID- 11114788 TI - Trends of polycyclic aromatic hydrocarbon levels and mutagenicity in Santiago's inhalable airborne particles in the period 1992-1996. AB - Trends of polycyclic aromatic hydrocarbons (PAHs) for 1992-1996 (cold season) and their mutagenic activity were investigated in organic extracts from the Santiago, Chile, inhalable particles (PM(10)). The highest PAH concentrations were observed in 1992 and declined dramatically in the following years. During this period, total PAHs decreased 85%, carcinogenic PAHs 82%, and benzo[a]pyrene, the most potent carcinogen, 85%. In spite of this significant decrease, PAH levels in respirable particles were higher than those reported in recent studies in Australia, Europe, and the United States. PAH profiles were analyzed by principal component (PC) analysis and Pearson correlation analysis. PC1 represents 71% of the variance, suggesting that most PAHs might originate predominantly from one main generic source. Higher correlations were obtained for the major carcinogenic PAHs. Most of the samples assayed were highly mutagenic to Salmonella typhimurium both in the presence and in the absence of metabolic activation system (S9), especially in the coarse fraction, but direct mutagenicity did not decline significantly. Incubation of calf thymus DNA with organic extracts from particulate matter and xanthine oxidase allowed the detection of five nitro-PAH DNA adducts. Thus, nitroarenes might play an important role in the mutagenic activity of inhalable particles in Santiago, representing a high risk for human health. PMID- 11114789 TI - Cyclooxygenase metabolites play a different role in ozone-induced pulmonary function decline in asthmatics compared to normals. AB - Indomethacin has been used to demonstrate that cyclooxygenase (COX) metabolites of arachidonic acid play a mechanistic role in ozone-induced spirometric decline in normals (Nm). Since the weight of evidence suggests that asthmatics (Asth) do not differ substantially from Nm subjects in the magnitude of their spirometric response to ozone, we sought to determine whether COX metabolites play a similar role in the asthmatic response to ozone. Thirteen (n = 13) Asth and nine (n = 9) Nm volunteers were pretreated with indomethacin or placebo (3 days, 75 mg/day), then exposed for 2 h to 400 ppb ozone or clean air while performing mild intermittent exercise (Vi(min) = 30 L/min.). Baseline changes in spirometry (FVC, FEV(1), FEF(25), FEF(50), FEF(60p), FEF(75)) and soluble markers of COX metabolism (prostaglandin [PG] F2-alpha) were measured from induced sputum samples. Results showed similar reductions in FVC (Asth = 12%, Nm = 10%) and FEV(1) (Asth = 13%, Nm = 11%) in Asth and Nm following ozone. Variables representing small-airways function demonstrated the greatest ozone-induced decline in Asth (FEF(75) = 25%). Indomethacin pretreatment significantly attenuated ozone-induced decreases in FVC and FEV(1) in Nm, but not in Asth. Marked attenuation of ozone-induced decrements in FEF(75) and FEF(60p) was observed in Asth but not in Nm. PGF2-alpha levels were similar in both groups prior to ozone exposure with indomethacin (Asth = 65 pg/ml, Nm = 59 pg/ml), but postexposure levels in Asth were significantly elevated (118 pg/ml) compared to Nm (54 pg/ml). We conclude that COX metabolites, such as PGF2-alpha, play an important but different role in asthmatics than normals with respect to ozone induced pulmonary function decline. Specifically, COX metabolites contribute to restrictive-type changes in normals and obstructive-type changes in small airways in asthmatics. PMID- 11114790 TI - A pulmonary rat gene array for screening altered expression profiles in air pollutant-induced lung injury. AB - Pulmonary tissue injury and repair processes involve complex and coordinated cellular events such as necrosis, inflammation, cell growth/differentiation, apoptosis, and remodeling of extracellular matrix. These processes are regulated by expression of multiple mediator genes. Commercially available microarray blots and slides allow screening of hundreds to thousands of genes in a given tissue or cell preparation. However, often these blots do not contain cDNAs of one's interest and are difficult to interpret. In order to analyze the tissue expression profile of a large number of genes involved in pulmonary injury and pathology, we developed a rat gene array filter using array technology. This array consisted of 27 genes representing inflammatory and anti-inflammatory cytokines, growth factors, adhesion molecules, stress proteins, transcription factors and antioxidant enzymes; 3 negative controls, and 2 blank spots. Using rat gene-specific polymerase chain reaction (PCR) primer pairs, cDNAs for these genes were amplified and cloned into a TA vector. Plasmids with recombinant cDNA inserts were purified and blotted onto a nylon membrane. Lung total RNA was isolated at 3 or 24 h following intratracheal (IT) exposure of male Sprague Dawley rats to either saline (control), residual oil fly ash (ROFA; 3.3 mg/kg) or metals found in one instillate of ROFA: nickel (NiSO(4); 1. 3 micromol/kg) or vanadium (VSO(4); 2.2 micromol/kg). (32)P-Labeled cDNA was generated from RNA samples in a reverse transcriptase reaction and subsequently hybridized to array blots. Densitometric scans of array blots revealed a twofold induction of interleukin (IL)-6 and TIMP-1 at 24 h post ROFA or Ni exposure. The pulmonary expressions of cellular fibronectin (cFn-EIIIA), ICAM-1, IL-1beta, and iNOS genes were also increased 24 h post ROFA-, V-, or Ni-exposure. Consistent hybridization of beta-actin in all array blots and absence of hybridization signals in negative controls indicated gene specific hybridization. ROFA or metal-induced increase in the expression of IL-6 observed in array blot was validated by Northern blot hybridization. Developing a pulmonary rat gene array may provide a tool for screening the expression profile of tissue specific markers following exposure to toxic air contaminants. PMID- 11114791 TI - 10 key questions for patients to ask. PMID- 11114792 TI - Home anticoagulation monitoring is safe and effective. PMID- 11114793 TI - Keeping vessels clear after bypass surgery. PMID- 11114794 TI - Ask the doctor. For several years now, my lower legs have been painful and somewhat swollen, especially at the end of the day. My doctor says that my veins are not getting the blood back to my heart like they used to, but that it is just part of aging (I am 72 years old). He tells me to put my feet up as much as possible, but that's not very practical when you have a lot to do! Isn't there anything else I could try? PMID- 11114795 TI - Ask the doctor. I have really annoying spells of atrial fibrillation. They happen about once a month, usually with no warning. I feel flushed and lightheaded until they pass. The spells used to occur about once a week, but now I am taking amiodarone, which cuts down the frequency. Still, I hate these spells and would love to get off these pills. I have heard about "defibrillator" devices that can be implanted in your body to shock you out of fibrillation. Is this something I should look into? PMID- 11114796 TI - Ask the doctor. I am 68 years old and had a heart attack last year. I think I am doing fine and passed my last exercise test with flying colors. But about a month ago I had a fainting spell, which occurred when I went to the bathroom after a big meal. In addition, I frequently feel lightheaded for a few seconds when I stand up. I think that problem has been there for years. Do you think these spells are related to my heart? PMID- 11114797 TI - Top ten health advances of 2000. PMID- 11114798 TI - Common cold. Zinc update. PMID- 11114799 TI - The doctor-patient relationship. The gray area of honesty. PMID- 11114800 TI - Nutrition, part I. Flavonoids, the next new thing? PMID- 11114801 TI - Nutrition, part II. Should you be eating garlic for your health? PMID- 11114802 TI - By the way, doctor... I've read on the Internet that people having a heart attack can keep themselves alive by coughing. Is this true? PMID- 11114803 TI - Antisocial personality - part I. PMID- 11114804 TI - Adaptation to chronic illness. PMID- 11114805 TI - Psychotherapy vindicated? PMID- 11114806 TI - Children's behavior and schizophrenia. PMID- 11114807 TI - Coping with loss. PMID- 11114808 TI - New cardiac risk factors. Part I: Introducing risk - and the leading new culprit. PMID- 11114809 TI - Medical memo. Hiccups. PMID- 11114810 TI - On call. I learned a lot from your article on melanoma, but I'm puzzled about one point. You suggest using sunscreen, but I read in my newspaper's health column that sunscreens may cause skin cancer. Should I use a sunscreen or not? PMID- 11114811 TI - At year's end: diet, glycemic index, and the food pyramid. PMID- 11114812 TI - Breast cancer update. Part III: adjuvant therapies and recovery issues. PMID- 11114813 TI - Tai chi: meditative movement for health. PMID- 11114814 TI - Estrogen and perimenopausal depression. PMID- 11114815 TI - By the way, doctor. I'm 67 years old and have been taking regular calcium supplements for quite some time. My latest bone-density test was very positive, showing increased BMD in my spine. Yet I was told that the test results aren't reliable because my x-ray showed calcification in my spine. How can this be true? PMID- 11114816 TI - Gilbert's syndrome and jaundice in glucose-6-phosphate dehydrogenase deficient neonates. PMID- 11114817 TI - Clozapine-induced blood dyscrasias. PMID- 11114818 TI - Thrombotic thrombocytopenic purpura treatment: a point of view. PMID- 11114819 TI - Erythropoietin doping. A comment. PMID- 11114822 TI - Erythroblastopenia and Parvovirus B19 infection in a healthy child. PMID- 11114821 TI - Treatment of severe aplastic anemia using high-dose cyclophosphamide alone in China. PMID- 11114823 TI - Polycythemia in a patient with 21-hydroxylase deficiency. PMID- 11114825 TI - Fulminant late onset cold hemagglutinin disease after allogeneic bone marrow transplantation. PMID- 11114824 TI - Low dose cytarabine and Granulocyte / Macrophage-Colony Stimulating Factor induced long-term remission in a patient with acute myeloid leukemia in early relapse after intensive chemotherapy. PMID- 11114826 TI - Spontaneous superficial vein thrombosis in a young patient double heterozygous for factor V Leiden and the prothrombin G20210A mutation: a case report. PMID- 11114827 TI - Erythrocytes, haemoglobin and packed cell volume in athletes performing races in altitude environment. PMID- 11114828 TI - Altered viral fitness of HIV-1 following failure of protease inhibitor-based therapy. AB - HIV-1 isolated from patients with improved CD4+ T-cell counts despite virologic failure on a nucleoside reverse transcriptase inhibitor (NRTI) and protease inhibitor (PI)-containing regimen were characterized. Five paired virus isolates from patients before and after zidovudine, lamivudine, and ritonavir treatment were tested. Human peripheral blood leukocyte-reconstituted severe combined immunodeficient (hu-PBL-SCID) mice were infected with pre-or posttreatment isolates and plasma HIV-1 RNA levels and CD4+ T cells were measured. Two of five post-treatment isolates exhibited decreased replication in hu-PBL-SCID mice compared with the paired pretreatment isolate, and both had the V82A mutation in protease associated with resistance to PI. One additional posttreatment isolate with the M184V mutation in reverse transcriptase showed diminished replication. CD4+ T-cell depletion was similar following infection with either the pre-or posttreatment isolates. Subtle losses in the replication capacity of PI-or NRTI resistant viruses may contribute to relative preservation of CD4+ T-cell counts in persons who experience virologic failure. Cytopathic effects of viral infection for target T cells vary from patient to patient but appear not to be influenced by mutations associated with failure of therapy in this system. PMID- 11114829 TI - Comparison of T-cell subsets' reconstitution after 12 months of highly active antiretroviral therapy initiated during early versus advanced states of HIV disease. AB - This research comprised a pilot open prospective clinical study comparing T-cell subset reconstitution in antiretroviral-naive patients, after 12 months of HAART when treatment was initiated in early stages (ES; n = 18) of infection versus advanced stages (AS; n = 20). CONTROL GROUP: 10 healthy HIV-negative individuals. Immunophenotypic markers were evaluated before and after 6-and 12-months' therapy. Functional assays were performed in one subset. RESULTS: Viral load (VL) was < 200 copies/ml in all patients. Median percentages of CD4+ pretherapy were 33% and 6%, respectively, in the ES and AS groups, increment after 12 months of therapy was +15% and +13% respectively. Only the ES group achieved normal values. Declines of CD8+ percentage were significantly higher in the ES (-18%) than in the AS group (-2%). CD4+ memory and naive cells in the ES group were similar to those of controls before treatment and did not change after therapy. In contrast, CD4+ memory and naive cells in the AS group did not reach normal levels despite treatment. In the ES group, there was a significant increment in CD8+ naive cells (+8%) and a decrement in CD8+CD38+ cells (-17%), both populations reached values close to normal, whereas these subsets remained far from normal in the AS group. Improvement of lymphoproliferative response after therapy was observed in both groups. One patient in the ES group showed positive LPR against p24 after treatment. After 12 months' highly active antiretroviral therapy, only those who began such therapy in ES disease reached values within the range of healthy controls. To achieve a more complete immunologic reconstitution, early initiation of potent antiretroviral therapy should be recommended. PMID- 11114830 TI - Multicenter review of protease inhibitors in 89 pregnancies. AB - CONTEXT: Despite the success of highly active antiretroviral therapy, the optimal approach for preventing perinatal HIV-1 transmission is not known. OBJECTIVE: A retrospective survey was conducted at six centers in the United States and Puerto Rico from January 1997 to October 1998 to evaluate the effects of protease inhibitor use during pregnancy on maternal and infant safety, prematurity rate, and frequency of perinatal HIV-1 transmission. RESULTS: In the study, 91 live infants, including 3 sets of twins, and 1 neonate who died shortly after birth were born to 89 women. HIV perinatal transmission rate in this series was 0 (95% confidence interval [CI], 0%-3%). Prematurity rate was 19.1%, comparable to rates in earlier reports of HIV-1-infected women. In multiple regression analysis, only cocaine use and premature rupture of membranes were associated with prematurity (p =.03 and.008, respectively). The gestational week during which the protease inhibitors were initiated was not found to be significantly associated with prematurity. Adverse maternal, obstetric, and infant events possibly related to protease inhibitors were uncommon. CONCLUSIONS: Protease inhibitors appeared generally safe in mothers and infants in this series. No perinatal HIV-1 transmission occurred. Further prospective, controlled studies are needed to define the optimal management of HIV-1 in pregnancy. PMID- 11114831 TI - Association of severe insulin resistance with both loss of limb fat and elevated serum tumor necrosis factor receptor levels in HIV lipodystrophy. AB - HIV-lipodystrophy (HIV-LD) is characterized by the loss of body fat from the limbs and face, an increase in truncal fat, insulin resistance, and hyperlipidemia, factors placing affected patients at increased risk for vascular disease. This study evaluated insulin sensitivity and inflammatory status associated with HIV-LD and provides suggestions about its etiology. Insulin sensitivity and immune activation markers were assessed in 12 control subjects and 2 HIV-positive groups, 14 without and 15 with LD syndrome. Peripheral insulin sensitivity (mostly skeletal muscle) was determined with the hyperinsulinemic euglycemic clamp. Circulating insulin-like growth factor (IGF) binding protein-1 (IGFBP-1) and free fatty acid (FFA) levels, and their response to insulin infusion were indicative of insulin responsiveness of liver and adipose tissue, respectively. Serum levels of soluble type 2 tumor necrosis factor-alpha (TNF alpha) receptor (sTNFR2) were used as an indicator of immune activation. HIV-LD study subjects had significantly reduced (twofold) peripheral insulin sensitivity, but normal levels of FFA and reduced levels of IGFBP-1, relative to the nonlipodystrophy groups, indicating that the loss of insulin sensitivity was more pronounced in skeletal muscle than in liver or fat. The significant loss of peripheral fat in the HIV-LD group (34%; p <.05) closely correlated with the reduced peripheral insulin sensitivity (p =. 0001). Levels of sTNFR2 were elevated in all HIV-infected study subjects, but they were significantly higher in those with lipodystrophy than without, and sTNFR2 levels strongly correlated with the reduction in insulin sensitivity (p =.0001). Loss of peripheral fat, normal levels of FFA, and reduced levels of IGFBP-1 indicate that insulin resistance in HIV-LD is distinct from type 2 diabetes and obesity. The relationship between the degree of insulin resistance and sTNFR2 levels suggests an inflammatory stimulus is contributing to the development of HIV-associated lipodystrophy. PMID- 11114832 TI - Safety, tolerability, and antiretroviral effects of ritonavir-nelfinavir combination therapy administered for 48 weeks. AB - OBJECTIVE: To evaluate the safety, tolerability, and anti-HIV activity of ritonavir-nelfinavir (RTV-NFV). DESIGN: Single-site, open-label, nonrandomized, multiple-dose trial of RTV combined with two doses of NFV in protease inhibitor (PI)-naive, HIV-infected patients. METHODS: Mean baseline HIV RNA was 39,500 copies/ml; mean baseline CD4 count was 323 cells/mm3. All patients received RTV at a dosage of 400 mg twice daily. Cohorts I (N = 10) and II (N = 10) received NFV at a dosage of 500 mg and 750 mg twice daily, respectively, for the initial 12 weeks of the study before allowing intensification with reverse transcriptase inhibitors. RESULTS: The commonest effects of RTV-NFV therapy were study drug related moderate-to-severe diarrhea (9 patients in cohorts I and II) and drug related moderate-to-severe nausea (4 patients in cohorts I and II). HIV RNA was suppressed in a biphasic manner. At 48 weeks in cohort I, mean HIV RNA reduction was 2.82 log10 copies/ml (standard error [SE] =.61; p =.001; N = 4); mean CD4 cell count increase was 236 cells/mm3 (SE = 67.1; p =.006; N = 4). In cohort II, mean HIV RNA reduction at Week 48 was 2.21 log10 copies/ml (SE =.430; p =. 001; N = 8); mean CD4 cell count increase was 120 cells/mm3 (SE = 47. 5; p =.03; n = 8). In cohort I patients, 2 of 4 completing Week 48 had HIV RNA <20 copies/ml; and 3 of 4 had HIV RNA <400 copies/ml. In cohort II, 2 of 8 patients completing Week 48 had HIV RNA <20 copies/ml and 4 of 8 had HIV RNA <400 copies/ml. In addition, 3 patients in cohort I withdrew because of virologic failure not thought to be related to poor compliance. Moreover, 15 patients elected to add new reverse transcriptase inhibitors (RTIs) after week 12. CONCLUSIONS: RTV-NFV with concomitant reverse transcriptase inhibitors is a potential dual-PI option for PI naive patients. PMID- 11114833 TI - Long-term safety and antiretroviral activity of hydroxyurea and didanosine in HIV infected patients. AB - Long-term safety, immunologic effects, and antiretroviral activity of hydroxyurea and didanosine were evaluated in this retrospective study. Some 65 HIV-1-infected patients (39 of whom were antiretroviral naive) were studied (mean baseline CD4 count, 362 cells/mm3; mean plasma HIV-1 RNA viral load, 4.8 log10 copies/ml). The mean treatment duration was 20 months. Overall tolerance was good: 15 patients interrupted treatment because of clinical or biologic side effects. Four patients experienced a category B event. Patients had a mean increase of 27 CD4 cell counts after 12 months, of 112 after 24 months and of 59 after 36 months. They had a mean 1. 03 log10 fall in HIV-1 RNA after 12 months, 1.59 log10 after 24 months, and 1.27 log10 after 36 months. After 12 months, 35% developed an HIV-1 RNA viral load <200 copies/ml, 53% after 24 months, and 36% after 36 months. Those whose viral load became undetectable after 12 months have significantly lower baseline RNA values (p =.03). Fourteen patients had a viral load <3.4 log10 copies/ml after 24 months of the double therapy. A prolonged viral load suppression can be achieved using a simple combination of two drugs that are inexpensive and well tolerated. PMID- 11114834 TI - Functional health literacy is associated with health status and health-related knowledge in people living with HIV-AIDS. AB - BACKGROUND: Poor health literacy is a prevalent barrier to medical care and people with lower health literacy experience greater illness severity than people with higher health literacy. Health literacy may therefore be an important factor in the health and treatment of people living with HIV-AIDS. METHODS: A community recruited sample of 339 HIV-infected men and women completed surveys and interviews that assessed functional health literacy, health status, AIDS-related disease and treatment knowledge, and health care perceptions and experiences. Medical records were available for chart abstraction of health status for a subsample of participants. RESULTS: About 1 of 4 people living with HIV-AIDS demonstrated difficulty comprehending simple medical instructions and therefore lower health literacy. HIV-infected people with lower health literacy had lower CD4 cell counts, higher viral loads, were less likely to be taking antiretroviral medications, reported a greater number of hospitalizations, and reported poorer health than those with higher health literacy. In addition, after adjusting for years of formal education, lower health literacy was associated with poorer knowledge of one's HIV-related health status, poorer AIDS-related disease and treatment knowledge, and more negative health care perceptions and experiences. CONCLUSIONS: Health literacy is a significant factor in the health and treatment of persons living with HIV-AIDS. Interventions are needed to improve medical care and the health status of people with lower health literacy that are living with HIV-AIDS. PMID- 11114835 TI - Decreased fertility among HIV-1-infected women attending antenatal clinics in three African cities. AB - Population HIV prevalence estimates rely heavily on sentinel surveillance in antenatal clinics (ANCs), but because HIV reduces fertility, these estimates are biased. To aid interpretation of such data, we estimated HIV-associated fertility reduction among pregnant women in ANCs in Yaounde (Cameroon), Kisumu (Kenya), and Ndola (Zambia). Data collection followed existing HIV sentinel surveillance procedures as far as possible. HIV prevalence among the women was 5.5% in Yaounde, 30.6% in Kisumu, and 27.3% in Ndola. The birth interval was prolonged in HIV-positive multiparous women compared with HIV-negative multiparous women in all three sites: adjusted hazard ratios of pregnancy were 0.84 (95% confidence interval [CI]: 0.62-1.1) in Yaounde, 0.82 (95% CI: 0.70-0.96) in Kisumu, and 0.74 (95% CI: 0.61-0.90) in Ndola, implying estimated reductions in the risk of pregnancy in HIV-positive women of between 16% and 26%. For primiparous women, the interval between sexual debut and birth was longer in HIV-positive women than in HIV-negative women in all sites, although the association was lost in Ndola after adjusting for age and other factors. Consistent results in different study sites help in the development of standard methods for improving ANC-based surveillance estimates of HIV prevalence. These may be easier to devise for multiparous women than for primiparous women. PMID- 11114836 TI - HIV-1 seroprevalence, risk factors, and preventive behaviors among women in northern Thailand. AB - To study HIV-1 seroprevalence, risk factors, and preventive behaviors among reproductive-age women in northern Thailand, 804 consenting women who were identified postpartum or who were visiting family planning clinics were interviewed and tested during 1998 to 1999. Almost all women were currently married and had been pregnant more than once. Their median age was 27 years. HIV 1 seroprevalence was 3.1% overall and was higher in women aged between 25 and 29 years (5.9%), having had > or =2 lifetime sex partners (6.5%), or whose current marriage had lasted for < or =1 year (7.0%). No woman reported HIV risk factors other than heterosexual sex. Most (76%) HIV-infected women reported no casual sex partners and, therefore, had likely acquired the infection from their husbands. HIV testing and partner communications were common, but only 2% of couples used condoms consistently in the prior 6 months. Nearly half of these women perceived themselves at no or low risk for HIV infection; these women were less likely to have taken preventive actions. To prevent HIV transmission in stable partnerships in this population, additional efforts are needed to increase HIV testing and condom use, to improve women's negotiation skills, and to develop new methods that do not require partner cooperation such as vaginal microbicides or vaccines. PMID- 11114837 TI - HIV in Vietnam: the evolving epidemic and the prevention response, 1996 through 1999. AB - OBJECTIVES: To describe epidemiologic patterns and trends in HIV infection in Vietnam from 1996 through 1999, and to summarize the national response to the epidemic. METHODS: We reviewed nationwide HIV case reports, and we analyzed annual seroprevalence among different sentinel populations in 21 provinces, using the chi2 test for linear trend to assess trends in HIV prevalence. HIV prevention efforts were also reviewed. RESULTS: Through 1999, 17,046 HIV infections, including 2947 AIDS cases and 1523 deaths had been reported in Vietnam. The cumulative incidence rate for the country was 22.5 per 100,000 population. Injection drug users (IDUs) represented 89.0% of all those for whom risk was reported before 1997 and 88.0% in the period 1997 to 1999. In 1999, HIV prevalence rates among IDUs ranged by province from 0% to 89.4%. Significantly increasing HIV trends among IDUs (p <.05) were found in 14 of the 21 sentinel provinces during 1996 to 1999. HIV prevalence among commercial sex workers (CSWs) ranged from 0% to 13.2%, increased significantly in 6 of 21 provinces. In 1999, prevalence among pregnant women, blood donors, and military recruits were 0.12%, 0. 20% and 0.61%, respectively. Major prevention activities include mass information; peer education and outreach among groups at increased risk; availability of low-cost syringes and condoms through pharmacies; needle exchange pilot projects; widely available treatment for sexually transmitted diseases; antibody screening of blood for transfusion; and free medical treatment at government hospitals. DISCUSSION: The HIV epidemic continues to evolve rapidly, intensifying among IDUs and increasing among CSWs. Serosurveillance indicators of HIV in the population at large continue to indicate the relatively slow extension beyond those at highest risk. Immediate, intensive preventions in high-risk groups may decelerate expansion to the broader population. PMID- 11114838 TI - Serum complement components in HIV-infected individuals with and without fat redistribution (lipodystrophy). PMID- 11114839 TI - Sustained responses to dual nucleoside therapy in women. PMID- 11114840 TI - Aggressive daily interferon therapy in HIV-HCV coinfected patients. PMID- 11114841 TI - Phenylpropanolamine and other OTC alpha-adrenergic agonists. PMID- 11114843 TI - Automated external defibrillators. PMID- 11114842 TI - Valproate and other anticonvulsants for psychiatric disorders. PMID- 11114844 TI - A simple method for generating full length cDNA from low abundance partial genomic clones. AB - BACKGROUND: PCR amplification of target molecules involves sequence specific primers that flank the region to be amplified. While this technique is generally routine, its applicability may not be sufficient to generate a desired target molecule from two separate regions involving intron /exon boundaries. For these situations, the generation of full-length complementary DNAs from two partial genomic clones becomes necessary for the family of low abundance genes. RESULTS: The first approach we used for the isolation of full-length cDNA from two known genomic clones of Hox genes was based on fusion PCR. Here we describe a simple and efficient method of amplification for homeobox D13 (HOXD13) full length cDNA from two partial genomic clones. Specific 5' and 3' untranslated region (UTR) primer pairs and website program (primer3_www.cgv0.2) were key steps involved in this process. CONCLUSIONS: We have devised a simple, rapid and easy method for generating cDNA clone from genomic sequences. The full length HOXD13 clone (1.1 kb) generated with this technique was confirmed by sequence analysis. This simple approach can be utilized to generate full-length cDNA clones from available partial genomic sequences. PMID- 11114845 TI - Estrogen receptor alpha dinucleotide repeat polymorphism in Japanese patients with autoimmune thyroid diseases. AB - BACKGROUND: The autoimmune thyroid diseases (AITDs), comprising Graves' disease (GD) and Hashimoto's thyroiditis (HT), appear to develop as a result of complex interactions between predisposing genes and environmental triggers. Susceptibility to AITDs is conferred by genes in the human leukocyte antigen (HLA) and genes unlinked to HLA, including the CTLA-4 gene. Recently, an association to some estrogen receptor (ER)alpha genotypes with breast cancer, hypertension, osteoporosis, generalized osteoarthritis, and some autoimmune diseases such as rheumatoid arthritis has been reported. We have analyzed a dinucleotide (TA)n repeat polymorphism lying upstream of the human ERalpha gene in patients with AITDs and in normal subjects. RESULTS: Seventeen different alleles were found in 130 patients with GD, 93 patients with HT, and 190 control subjects. There was no significant difference in the distributions of ERalpha alleles between patients and controls. CONCLUSIONS: The present results do not support an association between the ERalpha gene and AITD in the Japanese population. PMID- 11114846 TI - President's address: toward optimization of the treatment of patients with breast cancer. PMID- 11114847 TI - Taking the battle against breast cancer across borders: consumers, advocates, and change. PMID- 11114848 TI - Breast conservation therapy (BCT): surgery as the cornerstone of multi-specialty care. AB - Surgical care has been the mainstay of breast cancer diagnosis and treatment. As care has evolved, increased collaborative approaches among surgeons, radiologists, radiation oncologists and medical oncologists have improved the quality of breast cancer treatment for the patient. Breast conservation therapy (BCT) exemplifies how multi-specialty care can increase cancer cure rates at the same time that the disfiguring aspects of breast cancer treatment can be minimized. New questions are being raised within clinical forums about how to do better both for the patient and for her oncologic treatment. The following questions represent three current issues in BCT: 1. What general operative approaches in BCT can minimize morbidity and optimize the cosmetic outcome from surgery? 2. What role does radiation therapy play in BCT for invasive and non invasive breast cancer to supplement surgical intervention? 3. What role can neoadjuvant chemotherapy play in improving BCT rates? PMID- 11114849 TI - Oncoplastic techniques in the conservative surgical treatment of breast cancer. PMID- 11114850 TI - Optimal management of the axilla in patients with breast cancer. AB - BACKGROUND: Although most surgeons perform some form of axillary lymph node dissection (ALND) as part of locoregional management of operable breast cancer, the extent of dissection remains controversial. PATIENTS AND METHODS: Observation of the axilla trial (protocol I) and partial dissection trial (protocol I) began in January 1996. Between January 1996 and May 2000, 45 post-menopausal and 207 women with clinically node-negative breast cancer were enrolled into protocol I and protocol II respectively. RESULTS: The 4-year cumulative incidence rate of axillary recurrence was 7% in patients with untreated the axilla. The 4-year overall survival rate was 98% in patients with untreated the axilla. The 4-year disease-free and overall survival rates were 96% and 98% respectively in patients treated with partial dissection. CONCLUSION: Total axillary dissection seems to be unnecessary in Japanese breast cancer patients with relatively small tumors. PMID- 11114851 TI - Sentinel lymph node biopsy using tin colloid RI and blue dye method. AB - Axillary dissection has been considered essential for breast cancer staging because nodal metastasis is the most powerful predictive factor for recurrence. On the other hand, morbidity, such as lymphedema and shoulder dysfunction, may occur. Sentinel node biopsy is a good way to avoid unnecessary axillary dissection. We used tin colloid as a carrier of Tc99m tracer together with the blue dye method. The detection rate of the sentinel node was 27 cases out of 29 (90%) for the blue dye method, 10 cases out of 19 (53%) for the RI method, and 27 out of 33 (82%) for the combined method. The detection rate of the RI method was improved after adding the subcutaneous injection over the tumor from 45% before adding the subcutaneous injection to 82% after adding it. The false negative rate was 11% for the blue dye method, 0% for the RI method, and 10% for the combined method. This yields a sensitivity of 89% for the blue dye method, 100% for the RI method, and 90% for the combined method. Specificity was 100% for all three methods. Accuracy was 96% for the blue dye method, 100% for the RI method, and 96% for the combined method. There were two false negative cases. The average number of sentinel lymph nodes was 2.12 for the dye method, 1.66 for the RI method, and 1.95 for the combined method. There were three of 49 cases with identified parasternal lymph nodes by RI imaging. Lymphatic mapping using tin colloid may be useful for detecting sentinel nodes. PMID- 11114852 TI - Sentinel lymph node biopsy in breast cancer using blue dye with or without isotope localization. AB - BACKGROUND: The purpose of this study was to determine the feasibility of sentinel lymph node (SLN) biopsy using blue dye with or without isotope localization to predict the presence of axillary and internal mammary lymph node (IMN) metastases in patients with breast cancer. We also investigated whether multiple sectioning of the SLN could improve the accuracy of frozen section examination. METHOD: One-hundred twenty-six patients underwent dye-guided or dye- and gamma probe-guided SLN biopsy followed by complete axillary lymph node dissection (ALND). No ALND was performed in the 14 patients with small tumors and a negative SLN. In addition, 69 patients underwent IMN biopsy. RESULTS: The axillary SLN was identified in 123 of 140 (88%) patients. An accuracy rate of 90% was obtained by frozen section examination of the SLN, which increased to 100% in patients examined with a greater number of sections. Lymphatic flow to the IMN and/or a radioactive hot spot in the IMN was found in 9 of 102 (9%) patients, while a hot node was detected using a gamma probe in only 2 of these patients. No involvement of the IMNs was found histologically in these 9 patients. IMN involvement was found in 7 of 61 (11%) patients without lymphatic flow to the IMNs or a hot spot by lymphoscintigraphy or who did not undergo lymphoscintigraphy. CONCLUSION: ALND can be avoided in patients with small breast cancers and a negative SLN. SLN biopsy guided by lymphatic mapping is unreliable for identifying metastases to IMNs. PMID- 11114853 TI - Predictive factors for response to endocrine therapy in patients with recurrent breast cancer. AB - Predictive markers and variables for response to anticancer therapy provide cancer patients with refinement of therapeutic options and a decreased likelihood of receiving an ineffective therapy. The best-established predictive marker for response to endocrine therapy for breast cancer is the status of estrogen receptors (ER) in the primary breast tumor. However, although patients with ER positive tumors have a greater than 50% objective response rate to endocrine therapy, other patients can not obtain an objective response. Therefore additional markers, such as better molecular biologic markers, are needed. Our previous study using multivariate analysis revealed that the ER status of primary tumors and the dominant site of metastasis are independent predictors for response to first-line endocrine therapy and that a response to first-line endocrine therapy is only an independent predictor for response to second-line endocrine therapy. However, all these factors are already well-established predictive markers for response to endocrine therapy. Recently, a number of new hormonal agents, such as more selective aromatase inhibitors and specific antiestrogens, have been developed and introduced. However, several questions, such as the best sequences when using hormonal agents, remain to be elucidated. On the other hand, several molecular biologic markers predicting response to endocrine therapy, such as the expression of the HER family of tyrosine kinase receptors, pS2, Bcl-2, and vascular endothelial growth factor, have been reported. To elucidate the most effective use of endocrine therapy for recurrent breast cancer, classical and new predictive factors for response to endocrine therapy are reviewed, and the clinical implications of these factors are discussed. PMID- 11114854 TI - High dose chemotherapy as adjuvant treatment in operable breast cancer with 10 or more involved axillary lymph nodes. AB - BACKGROUND: it is well known that breast cancer patients with more than 10 axillary lymph nodes involved have poor prognosis even with extensive adjuvant chemotherapy. To improve this poor outcome, high-dose adjuvant chemotherapy has been applied to the these patients. This study was intended to clarify the efficacy and usefulness of high dose adjuvant chemotherapy (HDC) for high-risk breast cancer patients and its efficacy was compared with conventional adjuvant chemotherapy (non-HDC group). PATIENTS AND METHODS: Twelve patients with breast cancer involving more than 10 axillary nodes received high-dose chemotherapy with peripheral progenitor-stem cell transplantation (PBSCT). This regimen consists of BCNU (carmustine) 130 mg/m(2) x 3, CBDCA (carboplantin) 500 mg/m(2) x 3 and CPA (chyclophosphamide) 50 mg/kgx 2 after induction chemotherapy with 3 cycles of CE (chyclophosphamide 600 mg/m(2), epirubicin 60 mg/m(2)). RESULTS: Twelve patients completed the high-dose chemotherapy regimen as planned, no patient died of chemotherapy related toxicity. After a median follow-up period of 44 months, disease-free and overall survival at 48 months after the operation for 12 patients determined by Kaplan-Meier methods was 63 % and 83 %, respectively. Disease-free survival was superior in the high-dose chemotherapy group compared with the control group but a statistical difference was not observed. CONCLUSION: High-dose chemotherapy seems to be an effective and feasible treatment for high risk breast cancer patients. However, the usefulness of high-dose adjuvant chemotherapy for high-risk breast cancer patients should be confirmed by a large scale randomized trial. PMID- 11114855 TI - Bisphosphonate therapy for bone metastases from breast cancer: clinical results and a new therapeutic approach. AB - BACKGROUND: We evaluated the usefulness of bisphosphonate (BIS) monotherapy, the safety of rapid infusion of BIS and the efficacy of BIS-sequential therapy for bone metastases from breast cancer. PATIENTS AND METHODS: Twenty-nine patients with bone metastasis or invasion were treated with BIS monotherapy. Each BIS (pamidronate 30 mg, alendronate 10 mg, or incadronate 10 mg) was infused over 30 minutes every two weeks a median of 12 times. RESULTS: With BIS therapy, five patients (17%) showed partial response of the bone lesions, and eighteen patients (64%) had pain relief. Of the nine patients treated with BIS-sequential therapy, one (11%) showed a partial response of the bone metastases, three (33%) had pain relief, and one (11%) showed a decrease in the serum tumor marker level. CONCLUSION: BIS therapy is effective against bone metastases from breast cancer, and rapid infusion of BIS is both safe and convenient for patients. BIS sequential therapy can be a unique therapeutic option in some cases. PMID- 11114856 TI - The predictive value of angiogenesis for adjuvant therapy in breast cancer. AB - Recent accumulated data have indicated that angiogenesis is a promising indicator to predict prognosis and response to treatment. In breast cancer, microvessel density (MVD), a semi-quantitative marker of neovascularization grade, has been suggested to provide independent prognostic value. In addition, the expression of both vascular endothelial growth factor (VEGF)and thymidine phosphorylase (TP), two angiogenesis regulatory molecules, were found to be closely associated with MVD, and with the effect of treatments. In particular, VEGF expression seems to be a phenotype resistant to endocrine therapy, whereas TP expression decreases sensitivity to chemotherapy containing fluorouracil. This review summarizes the current topics regarding the prognostic and predictive value of angiogenesis in primary breast cancer, and we discuss future applications of antiangiogenesis treatments in an adjuvant setting. PMID- 11114857 TI - Ductal intraepithelial neoplasia (IDH, AIDH and DCIS). PMID- 11114858 TI - A proposal for the histopathological diagnosis of ductal carcinoma in situ of the breast. AB - BACKGROUND: As the incidence of ductal carcinoma in situ (DCIS) is increasing, it is necessary to make a guideline for the pathological examination and diagnosis of DCIS, by creating criteria based on clinical and biological aspects of the disease. METHOD: We collected biopsy specimens originally diagnosed as benign lesions, from patients who subsequently developed invasive carcinoma in the ipsilateral breast. The histology of the biopsy specimens was re-evaluated principally according to the 1995 Philadelphia Consensus on DCIS. Histopathological agreement on each biopsy specimen was made by the JBCS Study Group members under a multiviewer microscope. In the course of making conclusive agreements among the pathologists, we developed a consensus for the histopathological diagnosis of DCIS, especially non-comedo types. RESULTS: DCIS is defined as a carcinoma of ductal epithelial origin, without any evidence of stromal invasion. It is necessary to note the methods of pathologic examination required to diagnose DCIS. Stromal invasion is an important prognostic factor, and should be diagnosed with caution. Classification of proliferative ductal lesions as benign or malignant (DCIS), the subtype of DCIS (nuclear grade, architecture, and necrosis), and the histological grading of DCIS are proposed and recommended. CONCLUSION: Although we have made a new proposal according to current concepts, there are still several unresolved problems. Thus further examination and modification will be necessary in the future. PMID- 11114859 TI - Atypical cystic lobule of the breast: an early stage of low-grade ductal carcinoma in-situ. AB - The authors describe the characteristics of atypical cystic lobules (ACLs), which represent a step in the formation of low-grade ductal carcinoma in-situ. The authors define ACLs as a proliferation of luminal cells showing low-grade cytological atypia without architectural atypia. ACLs were compared with conventional hyperplasia, low-grade ductal carcinoma in-situ, and lobular neoplasia. 1) In about 40% of the cases, atypical cystic lobules merged with fully established micropapillary/cribriform ductal carcinoma in-situ. 2) Immunohistochemical staining for hormone receptors, keratin nineteen, and cyclin D1 revealed that atypical cystic lobules demonstrate a consistent immunophenotype, which differs from that of normal lobules and benign lesions and matches the one of low-grade ductal carcinoma in-situ. 3) ACLs are sometimes calcified. Osteopontin-positive histiocytes infiltrated all Kossa-positive (type II microcalcification) cribriform and comedo-type carcinomas and ACLs. The similarities in cytological and immunohistochemical features, the close proximity of the two types of proliferation, and the similarities with respect to calcification suggest that atypical cystic lobules represent an early stage in the formation of certain types of low-grade ductal carcinoma in-situ. PMID- 11114860 TI - Diagnosis of ductal carcinoma in situ (DCIS) and intraductal papilloma using fluorescence in situ hybridization (FISH) analysis. AB - BACKGROUND: It is often difficult to pre-operatively diagnose ductal carcinoma in situ (DCIS)or intraductal papilloma (IDP). Current reports show that breast cancer frequently has numerical aberrations of chromosomes 1, 11 and 17. We investigated whether fluorescence in situ hybridization (FISH) analysis using three centromere-specific probes for chromosomes 1, 11 and 17 was feasible for diagnosing intraductal breast lesions. METHODS: Fine-needle aspiration specimens from 102 breast lesions including DCIS (10), invasive ductal carcinoma (IDC) (78), IDP (7), fibroadenoma (6) and mastopathy (1) were examined for numerical aberrations on chromosomes 1, 11, 17 using FISH. If over 15% of all cells showed one signal, the sample was judged monosomic. If over 20% of cells showed three or more signals, it was considered polysomic. If the specimen had an aberration of at least one chromosome, it was judged positive. RESULTS: Nine of 10 DCISs showed numerical aberrations of at least one chromosome whereas 65 of 78 IDCs and 2 of 14 benign lesions (containing 7 IDPs of which one case was positive) showed numerical aberrations on these chromosomes. The proportion of positive results was highest with DCIS. Moreover 6 out of 7 DCISs showed an aberration of all three chromosomes simultaneously and one case showed an aberration of two chromosomes. All aberrations in case of DCIS were polysomic while two benign lesions and 15 IDCs showed a monosomic pattern. CONCLUSION: FISH may enable more accurate diagnosis of intraductal breast lesions. PMID- 11114861 TI - Surgery for ductal carcinoma in situ. AB - BACKGROUND: Ductal carcinomas in situ (DCIS) are sometimes treated too aggressively by surgery. We discuss minimal invasive surgery for DCIS on the basis of our experience at the Cancer Institute Hospital in Tokyo. METHODS: We performed surgery for 667 cases of DCIS between 1987 to 1998. This twelve year period we divided into three periods; 1987-1990,1991-1994, and 1995-1998. RESULTS: DCIS comprised 10% of all breast cancers, and tended to increase in incidence over time. The number of minimally invasive procedures such as breast conserving treatment (BCT), surgery without axillary dissection, and day surgery increased in later periods. In BCT for DCIS the surgical margin status is the most important factor, the rate of negative surgical margins was higher in DCIS than invasive cancer, and especially high in cases of mammographically detected nonpalpable cancer, the incidence of which is increasing yearly. The outcome of the 667 cases was very good. No distant metastases were observed, and the incidence of ipsilateral breast cancer(including second primary cancer) in these cases was 5% CONCLUSIONS: Because small cancers, including nonpalpable cases, will be detected more frequently, minimal invasive surgery will become more common for DCIS. PMID- 11114862 TI - Treatment of noninvasive carcinoma: fifteen-year results at the National Cancer Center Hospital in Tokyo. AB - The introduction of screening mammography (MMG) will lead to increased detection of preclinical early breast cancer in Japan. It has become more important to understand the nature of these lesions. We tried to elucidate the long term prognosis and clinical and pathological characteristics of noninvasive cancers. A total of 336 (5.4%) ductal carcinoma in situ (DCIS) and 32 (0.5%) lobular carcinoma in situ (LCIS) were diagnosed in 6 277 breast carcinomas at the National Cancer Center Hospital from 1962 to 1995. Most (80%) LCIS occurred in premenopausal women. LCIS has significantly higher bilaterality than that of DCIS. Local recurrence occurred in approximately 10% of patients after breast conserving surgery for DCIS and LCIS. Four patients died of breast carcinoma, which were initially diagnosed as noninfiltrating carcinoma. The 15-year cause specific survival rates of patients with DCIS and LCIS were 98.5 % and 100 %, respectively. PMID- 11114863 TI - Endocrine options for breast cancer treatment: looking beyond tamoxifen. PMID- 11114865 TI - Adjuvant chemotherapy for early breast cancer. PMID- 11114864 TI - New molecule-targeting therapy with herceptin (trastuzumab), an anti-HER2 (c-erB 2) monoclonal antibody. PMID- 11114866 TI - Optimising treatment of bone metastases by Aredia(TM) and Zometa(TM). AB - Metastatic bone disease develops as a result of the many interactions between tumour cells and bone cells. This leads to disruption of normal bone metabolism, with the increased osteoclast activity seen in most, if not all, tumor types providing a rational target for treatment. The clinical course of metastatic bone disease in multiple myeloma, breast and prostate cancers is relatively long, with patients experiencing sequential skeletal complications over a period of several years. These include bone pain, fractures, hypercalcaemia, and spinal cord compression, all of which may profoundly impair a patient's quality of life. External beam radiotherapy and systemic endocrine and cytotoxic treatments are the mainstay of treatment in advanced cancers. However, it is now clear that the bisphosphonates provide an additional treatment strategy, which reduces both the symptoms and complications of bone involvement. Pamidronate (Aredia(TM)) is the most widely evaluated bisphosphonate and is recommended for most patients with multiple myeloma or breast cancer with bone metastases. Current research aims include the evaluation of new potent bisphosphonates such as zoledronic acid (Zometa(TM)). It is hoped that this compound is not only more convenient and easier to administer but also more effective in inhibiting skeletal morbidity. Zometa may also have some direct anticancer activity. Preclinical studies with Zometa have demonstrated its potential in malignant bone disease. Clinical studies in treatment of hypercalcemia of malignancy have been completed, as have Phase I and II trials in patients with cancer and pre-existing bone metastases. Three randomized, double-blind, controlled Phase III trials are now ongoing to establish the efficacy and safety of Zometa in treatment of bone metastases in patients with osteolytic and osteoblastic lesions. Additionally, new specific molecules such as osteoprotogerin have been developed that are based on our improved understanding of the cellular signalling mechanisms involved in cancer induced bone disease. These potent molecules are now entering clinical trials. Ongoing research is aimed at trying to define the optimum route, dose, schedule and type of bisphosphonate in metastatic bone disease and their use in the prevention and treatment of osteoporosis in cancer patients. In vitro suggestions of direct anti-cancer activity and some promising clinical data in early breast cancer have resulted in considerable interest in the possible adjuvant use of bisphosphonates to inhibit the development of bone metastases. PMID- 11114867 TI - State of the art of minimally invasive breast biopsy: principles and practice. AB - In the last two decades, the increasing use of screening mammography with the early detection of breast cancer and the newly gained understanding of the biology and changing therapy of breast cancer, emphasizing systemic therapy and minimizing extensive local surgery, has contributed to the increasing development of minimally invasive techniques for the diagnosis of breast lesions. Minimally invasive techniques provide increased patient comfort, excellent cosmetic result and minimal morbidity They are also responsible for decreased costs and better medical care by allowing an informed discussion of breast cancer therapy and planning of surgery with an emphasis on negative margins and the dissection of the sentinel node. Techniques in use include Fine-Needle Aspiration Cytology, Core-Needle biopsy, Vacuum-Assisted Core biopsy (Mammotome) and Large Core biopsy (ABBI, Site-select). We present a balanced, evidence-based approach to the diagnosis of patients with palpable or mammographic abnormalities. PMID- 11114868 TI - Current perspectives for treatment of breast cancer. PMID- 11114869 TI - Sentinel lymph node biopsy for breast cancer: combined dye-isotope technique. PMID- 11114870 TI - Waiting lists. PMID- 11114871 TI - Prostatitis syndromes: an update for urologic practice. AB - OBJECTIVE: Five learning points will review and interpret the advances in the field of prostatitis that have occurred over the last decade. LEARNING POINT #1: The clinical syndrome of prostatitis and related conditions is one of the most common clinical encounters in urology. LEARNING POINT #2: The etiology/pathogenesis of the chronic prostatitis syndromes appears to be an inter related, multifactorial cascade beginning with an initiating event and culminating with a chronic neuropathic state. LEARNING POINT #3: The NIH classification of the chronic prostatitis syndromes is appropriate for not only research studies but also routine clinical practice. LEARNING POINT #4: A careful evaluation of the lower urinary tract and a standardized assessment of symptoms is essential in the management of prostatitis patients. LEARNING POINT #5: A "best evidence-based" approach to the treatment of the prostatitis syndromes is possible. PMID- 11114872 TI - Neoadjuvant hormone therapy and radical radiotherapy for localized prostate cancer: poorer biochemical outcome using flutamide alone. AB - Since a recent meta-analysis of non-steroidal anti-androgen therapy in metastatic prostate cancer concluded that survival was worse compared with medical or surgical androgen withdrawal, we analyzed our experience with flutamide monotherapy and other forms of neoadjuvant hormone therapy (NHT) prior to radiation therapy in clinically localized prostate cancer. A total of 45 patients received flutamide and 328 patients received other NHT. Flutamide patients had higher PSA levels at diagnosis and shorter duration of treatment, which could bias the results against flutamide monotherapy. Kaplan Meier analysis of PSA -- disease free survival showed significantly poorer outcome with flutamide monotherapy. Multivariate analysis supported this conclusion. Until equivalence to other forms of NHT is shown, we do not recommend flutamide monotherapy prior to radical radiation. A prospective randomized trial would be necessary to confirm this conclusion. PMID- 11114873 TI - The value of percent free prostate specific antigen, prostate specific antigen density of the whole prostate and of the transition zone in Turkish men. AB - INTRODUCTION: This study was undertaken to evaluate the value of percent free prostate specific antigen (PSA), PSA density of the whole prostate (PSAD) and of the transition zone (TZPSAD) in reducing unnecessary biopsies in Turkish men with serum PSA levels between 2.5 and 20 ng/mL. MATERIALS AND METHODS: A total of 105 patients referred for early prostate cancer detection or lower urinary tract symptoms participated in the study. All patients had serum total PSA level, PSAD, total prostate volume, and rectal examination, 43 patients had serum free PSA level, and 65 patients had TZPSAD. Using transrectal ultrasound, sextant biopsy and two additional transitional zone biopsies were performed. The value of percent PSA, PSAD, and TZPSAD in reducing unnecessary biopsies was evaluated. RESULTS: Prostate carcinoma was detected in 25 of 105 patients (23.8%). Overall, areas under the ROC curves for percent free PSA, PSAD, and TZPSAD were 0.553, 0.595, and 0.550, respectively. In patients with a benign rectal examination, the value of percent free PSA slightly increased. On the other hand, in patients with prostate volume smaller than 50 cc, the value of percent free PSA and TZPSAD was encouraging. Areas under the ROC curves for percent free PSA, and TZPSAD were 0.615 and 0.642 respectively. CONCLUSION: In Turkish men with intermediate serum PSA concentrations, percent free PSA, PSAD, and TZPSAD are poor predictors of biopsy outcome, whereas in the prostate smaller than 50 cc, percent free PSA and TZPSAD provide additional information. This may reflect ethnic differences between this population and those reported in other series. PMID- 11114874 TI - Results of concomitant radio-chemotherapy for invasive bladder tumors. AB - INTRODUCTION: The aim of this study was to evaluate combined radiation and chemotherapy for invasive bladder tumors and to define possible biological prognostic factors. MATERIALS AND METHODS: Forty-five patients (mean age 75 years, range 68 to 86) were treated by deep bladder resection with combined radiation (5 courses of 10 Grays [Gys]) and chemotherapy (2 cycles of 100 mg of cisplatin over 7 weeks). Subsequent to the treatment, evaluations at 10 weeks, then 3 months, 6 months, and 12 months, and every year, included a clinical examination, Karnofsky evaluation, a cystoscopy with urinary cytology, and systematic deep bladder pathology biopsies after a computer tomography (CT) scan. In our series, DNA analysis and an immuno-histochemistry study of MiB1, p53, and MdR were also performed in the last 20 patients. RESULTS: At 6 months, progression of the disease was evaluated in 26% of cases; at 12 months, satisfactory local control was evaluated in 60% of cases. However, only 15 patients were in complete remission after a period of 2 years. Of these, only eight patients were in remission with more than 42 months follow-up. We observed aneuploid tumors in 30% of cases; these patients died after 6 months despite concomitant radio-chemotherapy. In our experience, the results of associated immuno-histochemistry markers was not really contributive. CONCLUSION: This therapeutic alternative may be useful in patients where a cystectomy cannot be performed. It not only permits bladder conservation, but also offers satisfactory, significant quality of life, even if survival rates remain uncertain. PMID- 11114875 TI - Pubovaginal sling technique utilizing a unique bone anchor instrumentation system. AB - INTRODUCTION: In the past, the pubovaginal sling (PVS) technique was originally delegated for the treatment of intrinsic sphincter deficiency syndrome (ISD). Today, it has not only undergone a revitalization, but is being recommended for the treatment of all forms of stress urinary incontinence (SUI) as well as the ISD syndrome. In an attempt to combine the best features of the traditional approach plus add the benefits of simplicity, reduction of costs, morbidity, and rapid return to patient normality, a new variation of the PVS has been developed. The technique utilizes pre-threaded bone anchors to which either a natural fascia or pre-prepared cadaveric fascia can be anchored. MATERIAL AND METHODS: This study consists of 78 female patients treated between September 1997 and December 1998 with our PVS procedure. The patient population spans the spectrum of pure stress incontinence, with or without associated pelvic relaxation defects, pure ISD group and lastly, those individuals who suffered from both anatomical incontinence and overactive bladder syndrome. In our 72 evaluable patients, the results as of this publication are: an overall cure rate of 86% with an additional 11% improved and 3% failure. The following text describes in detail the patient population, the surgical technique, the final results, complications, and patient satisfaction scores. Also included is a short review of the literature documenting several other techniques utilizing bone anchoring fixation devices. CONCLUSION: A simplification of the true-and-tried PVS is described which provides the surgeon with a new and exciting methodology for the treatment of all forms of hypermobility, i.e. stress incontinence, as well as the intrinsic sphincter deficiency syndrome. At the same time the surgical learning curve, patient morbidity, and hospital stay are decreased; without compromising total surgical outcome. PMID- 11114876 TI - By the way, doctor... I'm 70. I took hormone therapy for a while after menopause, but stopped taking it long ago. My memory definitely isn't as good as it used to be. If I started taking estrogen again, would my memory improve? PMID- 11114877 TI - Warning on Mellaril. PMID- 11114878 TI - Antipsychotic drugs: the weight problem. PMID- 11114879 TI - On call. I've had my PSA tested every year since 1992 and I'm happy to say it's always been normal. But now my doctor doesn't want to do the test anymore because I'm 78 years old. Should I go to another doctor for my test? PMID- 11114880 TI - Getting help for infertility. PMID- 11114881 TI - By the way, doctor. I've seen a lot in the news lately about women using birth control pills to delay their menstrual periods or avoid having them altogether. How does this work? Is it safe? PMID- 11114882 TI - Neurotrophins: key regulators of cell fate and cell shape in the vertebrate nervous system. PMID- 11114883 TI - A novel two-component hybrid molecule regulates vascular morphogenesis of the Arabidopsis root. AB - The developmental ontogeny of the vascular system (consisting of xylem, phloem and [pro]cambium) is poorly understood despite its central role in plant physiology. We show that in the Arabidopsis root meristem, xylem cell lineages are specified early, whereas phloem and procambium are established through a set of asymmetric cell divisions. These divisions require the WOODEN LEG (WOL) gene. The WOL gene encodes a novel two-component signal transducer with an unusual tandem arrangement of two receiver domains. It is expressed specifically in the vasculature from the early stages of embryogenesis on, consistent with a role as a sensor for vascular morphogenesis. PMID- 11114884 TI - Cap methyltransferase selective binding and methylation of GpppG-RNA are stimulated by importin-alpha. AB - We screened a human cDNA library for proteins that bind mRNA cap methyltransferase (MT) and isolated nuclear transporter importin-alpha (Impalpha). This direct association was confirmed by glutathione S-transferase (GST) pulldown, coimmunoprecipitation, and nuclear colocalization. In gel shift assays, MT selectively bound RNA containing 5'-terminal GpppG, and binding was inhibited by GpppG and not by m(7)GpppC. Impalpha markedly enhanced MT binding to GpppG-RNA and stimulated MT activity. MT/RNA/Impalpha complexes were dissociated by importin-beta, which also blocked the stimulation of cap methylation by Impalpha. The presence of RanGTP but not RanGDP prevented these effects of importin-beta. These findings indicate that importins play a novel role in mRNA biogenesis at the level of cap methylation. PMID- 11114885 TI - Restricted feeding uncouples circadian oscillators in peripheral tissues from the central pacemaker in the suprachiasmatic nucleus. AB - In mammals, circadian oscillators exist not only in the suprachiasmatic nucleus, which harbors the central pacemaker, but also in most peripheral tissues. It is believed that the SCN clock entrains the phase of peripheral clocks via chemical cues, such as rhythmically secreted hormones. Here we show that temporal feeding restriction under light-dark or dark-dark conditions can change the phase of circadian gene expression in peripheral cell types by up to 12 h while leaving the phase of cyclic gene expression in the SCN unaffected. Hence, changes in metabolism can lead to an uncoupling of peripheral oscillators from the central pacemaker. Sudden large changes in feeding time, similar to abrupt changes in the photoperiod, reset the phase of rhythmic gene expression gradually and are thus likely to act through a clock-dependent mechanism. Food-induced phase resetting proceeds faster in liver than in kidney, heart, or pancreas, but after 1 wk of daytime feeding, the phases of circadian gene expression are similar in all examined peripheral tissues. PMID- 11114886 TI - The unfolded protein response represses nitrogen-starvation induced developmental differentiation in yeast. AB - Diploid budding yeast exhibits two developmental programs in response to nitrogen starvation, pseudohyphal growth, and sporulation. Here we show that both programs are repressed by activation of the unfolded protein response (UPR), a stress signal transduction pathway responsible for induction of endoplasmic reticulum (ER)-resident chaperones when protein folding in the ER is impaired. Pseudohyphal growth was derepressed in ire1Delta/ire1Delta and hac1Delta/hac1Delta strains. Activation of the UPR or overexpression of the transcription factor Hac1(i)p, the product of an unconventional splicing reaction regulated by the UPR, was sufficient for repression of pseudohyphal growth and meiosis. HAC1 splicing occurred in a nitrogen-rich environment but ceased rapidly on nitrogen starvation. Further, addition of ammonium salts to nitrogen-starved cells was sufficient to rapidly reactivate HAC1 splicing. We propose that high translation rates in a nitrogen-rich environment are coupled to limited protein unfolding in the ER, thereby activating the UPR. An activated UPR then represses pseudohyphal growth and meiosis. Nitrogen starvation slows translation rates, allowing for more efficient folding of nascent polypeptide chains, down-regulation of the UPR, and subsequent derepression of pseudohyphal growth and meiosis. These findings significantly broaden the range of physiological functions of the UPR and define a role for the UPR in nitrogen sensing. PMID- 11114887 TI - FtsK functions in the processing of a Holliday junction intermediate during bacterial chromosome segregation. AB - In bacteria with circular chromosomes, homologous recombination can generate chromosome dimers that cannot be segregated to daughter cells at cell division. Xer site-specific recombination at dif, a 28-bp site located in the replication terminus region of the chromosome, converts dimers to monomers through the sequential action of the XerC and XerD recombinases. Chromosome dimer resolution requires that dif is positioned correctly in the chromosome, and the activity of FtsK, a septum-located protein that coordinates cell division with chromosome segregation. Here, we show that cycles of XerC-mediated strand exchanges form and resolve Holliday junction intermediates back to substrate irrespective of whether conditions support a complete recombination reaction. The C-terminal domain of FtsK is sufficient to activate the exchange of the second pair of strands by XerD, allowing both intra- and intermolecular recombination reactions to go to completion. Proper positioning of dif in the chromosome and of FtsK at the septum is required to sense the multimeric state of newly replicated chromosomes and restrict complete Xer reactions to dimeric chromosomes. PMID- 11114888 TI - Functional interactions between BRCA1 and the checkpoint kinase ATR during genotoxic stress. AB - The BRCA1 gene encodes a tumor suppressor that is mutated in 50% of familial breast cancers. The BRCA1 protein has been implicated in the DNA damage response, as DNA damage induces the phosphorylation of BRCA1 and causes its recruitment into nuclear foci that contain DNA repair proteins. The ataxia-telangiectasia mutated (ATM) gene product controls overall BRCA1 phosphorylation in response to gamma-irradiation (IR). In this study, we show that BRCA1 phosphorylation is only partially ATM dependent in response to IR and ATM independent in response to treatment with UV light, or the DNA replication inhibitors hydroxyurea (HU) and aphidicolin (APH). We provide evidence that the kinase responsible for this phosphorylation is the ATM-related kinase, ATR. ATR phosphorylates BRCA1 on six Ser/Thr residues, including Ser 1423, in vitro. Increased expression of ATR enhanced the phosphorylation of BRCA1 on Ser 1423 following cellular exposure to HU or UV light, whereas doxycycline-induced expression of a kinase-inactive ATR mutant protein inhibited HU- or UV light-induced Ser 1423 phosphorylation in GM847 fibroblasts, and partially suppressed the phosphorylation of this site in response to IR. Thus, ATR, like ATM, controls BRCA1 phosphorylation in vivo. Although ATR isolated from DNA-damaged cells does not show enhanced kinase activity in vitro, we found that ATR responds to DNA damage and replication blocks by forming distinct nuclear foci at the sites of stalled replication forks. Furthermore, ATR nuclear foci overlap with the nuclear foci formed by BRCA1. The dramatic relocalization of ATR in response to DNA damage points to a possible mechanism for its ability to enhance the phosphorylation of substrates in response to DNA damage. Together, these results demonstrate that ATR and BRCA1 are components of the same genotoxic stress-responsive pathway, and that ATR directly phosphorylates BRCA1 in response to damaged DNA or stalled DNA replication. PMID- 11114889 TI - Phosphorylation of histone H3 correlates with transcriptionally active loci. AB - Posttranslational modifications of the N-terminal tails of the core histones within the nucleosome particle are thought to act as signals from the chromatin to the cell for various processes. The experiments presented here show that the acetylation of histones H3 and H4 in polytene chromosomes does not change during heat shock. In contrast, the global level of phosphorylated H3 decreased dramatically during a heat shock, with an observed increase in H3 phosphorylation at the heat shock loci. Additional experiments confirm that this change in phosphorylated H3 distribution is dependent on functional heat shock transcription factor activity. These experiments suggest that H3 phosphorylation has an important role in the induction of transcription during the heat shock response. PMID- 11114890 TI - Reciprocal activation of xenobiotic response genes by nuclear receptors SXR/PXR and CAR. AB - The cytochrome P450 (CYP) gene products such as CYP3A and CYP2B are essential for the metabolism of steroid hormones and xenochemicals including prescription drugs. Nuclear receptor SXR/PXR (steroid and xenobiotic receptor/pregnenolone X receptor) has been shown both biochemically and genetically to activate CYP3A genes, while similar studies have established constitutive androstane receptor (CAR) as a CYP2B regulator. The response elements in these genes are also distinct, furthering the concept of independent regulation. Unexpectedly, we found that SXR can regulate CYP2B, both in cultured cells and in transgenic mice via adaptive recognition of the phenobarbital response element (PBRE). In a type of functional symmetry, orphan receptor CAR was also found to activate CYP3A through previously defined SXR/PXR response elements. These observations not only provide a rational explanation for the activation of multiple CYP gene classes by certain xenobiotics, but also reveal the existence of a metabolic safety net that confers a second layer of protection to the harmful effects of toxic compounds and at the same time increases the propensity for drug-drug interactions. PMID- 11114891 TI - Arabidopsis NAC1 transduces auxin signal downstream of TIR1 to promote lateral root development. AB - Auxin plays a key role in lateral root formation, but the signaling pathway for this process is poorly understood. We show here that NAC1, a new member of the NAC family, is induced by auxin and mediates auxin signaling to promote lateral root development. NAC1 is a transcription activator consisting of an N-terminal conserved NAC-domain that binds to DNA and a C-terminal activation domain. This factor activates the expression of two downstream auxin-responsive genes, DBP and AIR3. Transgenic plants expressing sense or antisense NAC1 cDNA show an increase or reduction of lateral roots, respectively. Finally, TIR1-induced lateral root development is blocked by expression of antisense NAC1 cDNA, and NAC1 overexpression can restore lateral root formation in the auxin-response mutant tir1, indicating that NAC1 acts downstream of TIR1. PMID- 11114892 TI - Targeted disruption of the three Rb-related genes leads to loss of G(1) control and immortalization. AB - The retinoblastoma protein, pRB, and the closely related proteins p107 and p130 are important regulators of the mammalian cell cycle. Biochemical and genetic studies have demonstrated overlapping as well as distinct functions for the three proteins in cell cycle control and mouse development. However, the role of the pRB family as a whole in the regulation of cell proliferation, cell death, or cell differentiation is not known. We generated embryonic stem (ES) cells and other cell types mutant for all three genes. Triple knock-out mouse embryonic fibroblasts (TKO MEFs) had a shorter cell cycle than wild-type, single, or double knock-out control cells. TKO cells were resistant to G(1) arrest following DNA damage, despite retaining functional p53 activity. They were also insensitive to G(1) arrest signals following contact inhibition or serum starvation. Finally, TKO MEFs did not undergo senescence in culture and do possess some characteristics of transformed cells. Our results confirm the essential role of the Rb family in the control of the G(1)/S transition, place the three Rb family members downstream of multiple cell cycle control pathways, and further the link between loss of cell cycle control and tumorigenesis. PMID- 11114893 TI - Ablation of the retinoblastoma gene family deregulates G(1) control causing immortalization and increased cell turnover under growth-restricting conditions. AB - The retinoblastoma suppressor pRB belongs to the family of so-called pocket proteins, which also includes p107 and p130. These proteins may functionally overlap in cell cycle control and tumor suppression. We have generated an isogenic set of embryonic stem (ES) cell lines carrying single or compound loss of-function mutations in the Rb gene family, including a cell line completely devoid of all three pocket proteins. None of the knockout combinations affected the growth characteristics of ES cells; however, concomitant ablation of all three pocket proteins strongly impaired their differentiation capacity. For the generated genotypes, primary mouse embryonic fibroblasts (MEFs) also were obtained. While inactivation of Rb alone did not alleviate the senescence response of MEFs, pRB/p107-deficient MEFs, after having adapted to in vitro culturing, continued to proliferate at modest rate. Additional ablation of p130 rendered MEFs completely insensitive to senescence-inducing signals and strongly increased their proliferation rate. Although triple-knockout MEFs retained anchorage dependence, they lacked proper G(1) control and showed increased cell turnover under growth-inhibiting conditions. PMID- 11114894 TI - Role of Bak in UV-induced apoptosis in skin cancer and abrogation by HPV E6 proteins. AB - Ultraviolet B (UVB) damage is recognized as the most important etiological factor in the development of skin cancer. Human papillomaviruses (HPV) have also been implicated in the disease, although the mechanism of action of these viruses remains unknown. We present evidence here that Bak protein is involved in signaling apoptosis in the skin in response to UVB damage, and that cutaneous HPV E6 proteins target and abrogate Bak function by promoting its proteolytic degradation both in vitro and in regenerated epithelium. Additionally, HPV positive skin cancers had undetectable levels of Bak in contrast to HPV negative cancers, which expressed Bak. This study supports a link between the virus and UVB in the induction of HPV-associated skin cancer and reveals a survival mechanism of virally infected cells. PMID- 11114895 TI - 1-Deoxy-D-xylulose 5-phosphate synthase, the gene product of open reading frame (ORF) 2816 and ORF 2895 in Rhodobacter capsulatus. AB - In eubacteria, green algae, and plant chloroplasts, isopentenyl diphosphate, a key intermediate in the biosynthesis of isoprenoids, is synthesized by the methylerythritol phosphate pathway. The five carbons of the basic isoprenoid unit are assembled by joining pyruvate and D-glyceraldehyde 3-phosphate. The reaction is catalyzed by the thiamine diphosphate-dependent enzyme 1-deoxy-D-xylulose 5 phosphate synthase. In Rhodobacter capsulatus, two open reading frames (ORFs) carry the genes that encode 1-deoxy-D-xylulose 5-phosphate synthase. ORF 2816 is located in the photosynthesis-related gene cluster, along with most of the genes required for synthesis of the photosynthetic machinery of the bacterium, whereas ORF 2895 is located elsewhere in the genome. The proteins encoded by ORF 2816 and ORF 2895, 1-deoxy-D-xylulose 5-phosphate synthase A and B, containing a His(6) tag, were synthesized in Escherichia coli and purified to greater than 95% homogeneity in two steps. 1-Deoxy-D-xylulose 5-phosphate synthase A appears to be a homodimer with 68 kDa subunits. A new assay was developed, and the following steady-state kinetic constants were determined for 1-deoxy-D-xylulose 5-phosphate synthase A and B: K(m)(pyruvate) = 0.61 and 3.0 mM, K(m)(D-glyceraldehyde 3 phosphate) = 150 and 120 microM, and V(max) = 1.9 and 1.4 micromol/min/mg in 200 mM sodium citrate (pH 7.4). The ORF encoding 1-deoxy-D-xylulose 5-phosphate synthase B complemented the disrupted essential dxs gene in E. coli strain FH11. PMID- 11114896 TI - Mutational analysis of the OprM outer membrane component of the MexA-MexB-OprM multidrug efflux system of Pseudomonas aeruginosa. AB - OprM is the outer membrane component of the MexA-MexB-OprM efflux system of Pseudomonas aeruginosa. Multiple-sequence alignment of this protein and its homologues identified several regions of high sequence conservation that were targeted for site-directed mutagenesis. Of several deletions which were stably expressed, two, spanning residues G199 to A209 and A278 to N286 of the mature protein, were unable to restore antibiotic resistance in OprM-deficient strains of P. aeruginosa. Still, mutation of several conserved residues within these regions did not adversely affect OprM function. Mutation of the highly conserved N-terminal cysteine residue, site of acylation of this presumed lipoprotein, also did not affect expression or activity of OprM. Similarly, substitution of the OprM lipoprotein signal, including consensus lipoprotein box, with the signal peptide of OprF, the major porin of this organism, failed to impact on expression or activity. Apparently, acylation is not essential for OprM function. A large deletion at the N terminus, from A12 to R98, compromised OprM expression to some extent, although the deletion derivative did retain some activity. Several deletions failed to yield an OprM protein, including one lacking an absolutely conserved LGGGW sequence near the C terminus of the protein. The pattern of permissive and nonpermissive deletions was used to test a topology model for OprM based on the recently published crystal structure of the OprM homologue, TolC (V. Koronakis, A. Sharff, E. Koronakis, B. Luisi, and C. Hughes, Nature 405:914-919, 2000). The data are consistent with OprM monomer existing as a substantially periplasmic protein with four outer membrane-spanning regions. PMID- 11114897 TI - Hin recombinase mutants functionally disrupted in interactions with Fis. AB - A previous genetic screen was designed to separate Hin recombinase mutants into distinct classes based on the stage in the recombination reaction at which they are blocked (O. Nanassy, Zoltan, and K. T. Hughes, Genetics 149:1649-1663, 1998). One class of DNA binding-proficient, recombination-deficient mutants was predicted by genetic classification to be defective in the step prior to invertasome formation. Based on the genetic criteria, mutants from this class were also inferred to be defective in interactions with Fis. In order to understand how the genetic classification relates to individual biochemical steps in the recombination reaction these mutants, R123Q, T124I, and A126T, were purified and characterized for DNA cleavage and recombination activities. Both the T124I and A126T mutants were partially active, whereas the R123Q mutant was inactive. The A126T mutant was not as defective for recombination as the T124I allele and could be partially rescued for recombination both in vivo and in vitro by increasing the concentration of Fis protein. Rescue of the A126T allele required the Fis protein to be DNA binding proficient. A model for a postsynaptic role for Fis in the inversion reaction is presented. PMID- 11114898 TI - The chvH locus of Agrobacterium encodes a homologue of an elongation factor involved in protein synthesis. AB - The virulence of Agrobacterium tumefaciens depends on both chromosome- and Ti plasmid-encoded gene products. In this study, we characterize a chromosomal locus, chvH, previously identified by TnphoA mutagenesis and shown to be required for tumor formation. Through DNA sequencing and comparison of the sequence with identified sequences in the database, we show that this locus encodes a protein similar in sequence to elongation factor P, a protein thought to be involved in peptide bond synthesis in Escherichia coli. The analysis of vir-lacZ and vir-phoA translational fusions as well as Western immunoblotting revealed that the expression of Vir proteins such as VirE2 was significantly reduced in the chvH mutant compared with the wild-type strain. The E. coli efp gene complemented detergent sensitivity, virulence, and expression of VirE2 in the chvH mutant, suggesting that chvH and efp are functionally homologous. As expected, ChvH exerts its activity at the posttranscriptional level. Southern analysis suggests that the gene encoding this elongation factor is present as a single copy in A. tumefaciens. We constructed a chvH deletion mutant in which a 445-bp fragment within its coding sequence was deleted and replaced with an omega fragment. On complex medium, this mutant grew more slowly than the wild-type strain, indicating that elongation factor P is important but not essential for the growth of Agrobacterium. PMID- 11114899 TI - Divergence of eukaryotic secretory components: the Candida albicans homolog of the Saccharomyces cerevisiae ++Sec20 protein is N terminally truncated, and its levels determine antifungal drug resistance and growth. AB - Sec20p is a component of the yeast Saccharomyces cerevisiae secretory pathway that does not have a close homolog in higher eukaryotic cells. To verify the function of Sec20p in other fungal species, we characterized the gene encoding a Sec20p homolog in the human fungal pathogen Candida albicans. The deduced protein has 27% identity with, but is missing about 100 N-terminal residues compared to S. cerevisiae Sec20p, which is part of the cytoplasmic tail interacting with the cytoplasmic protein Tip20p. Because a strain lacking both C. albicans SEC20 alleles could not be constructed, we placed SEC20 under transcriptional control of two regulatable promoters, MET3p and PCK1p. Repression of SEC20 expression in these strains prevented (MET3p-SEC20 allele) or retarded (PCK1p-SEC20 allele) growth and led to the appearance of extensive intracellular membranes, which frequently formed stacks. Reduced SEC20 expression in the PCK1p-SEC20 strain did not affect morphogenesis but led to a series of hypersensitivity phenotypes including supersensitivity to aminoglycoside antibiotics, to nystatin, to sodium dodecyl sulfate, and to cell wall inhibitors. These results demonstrate the occurrence and function of Sec20p in a fungal species other than S. cerevisiae, but the lack of the N-terminal domain and the apparent absence of a close TIP20 homolog in the C. albicans genome also indicate a considerable diversity in mechanisms of retrograde vesicle traffic in eukaryotes. PMID- 11114900 TI - Role of the Eikenella corrodens pilA locus in pilus function and phase variation. AB - The human pathogen Eikenella corrodens expresses type IV pili and exhibits a phase variation involving the irreversible transition from piliated to nonpiliated variants. On solid medium, piliated variants form small (S-phase), corroding colonies whereas nonpiliated variants form large (L-phase), noncorroding colonies. We are studying pilus structure and function in the clinical isolate E. corrodens VA1. Earlier work defined the pilA locus which includes pilA1, pilA2, pilB, and hagA. Both pilA1 and pilA2 predict a type IV pilin, whereas pilB predicts a putative pilus assembly protein. The role of hagA has not been clearly established. That work also confirmed that pilA1 encodes the major pilus protein in this strain and showed that the phase variation involves a posttranslational event in pilus formation. In this study, the function of the individual genes comprising the pilA locus was examined using a recently developed protocol for targeted interposon mutagenesis of S-phase variant VA1-S1. Different pilA mutants were compared to S-phase and L-phase variants for several distinct aspects of phase variation and type IV pilus biosynthesis and function. S-phase cells were characterized by surface pili, competence for natural transformation, and twitching motility, whereas L-phase cells lacked these features. Inactivation of pilA1 yielded a mutant that was phenotypically indistinguishable from L-phase variants, showing that native biosynthesis of the type IV pilus in strain VA1 is dependent on expression of pilA1 and proper export and assembly of PilA1. Inactivation of pilA2 yielded a mutant that was phenotypically indistinguishable from S-phase variants, indicating that pilA2 is not essential for biosynthesis of functionally normal pili. A mutant inactivated for pilB was deficient for twitching motility, suggesting a role for PilB in this pilus-related phenomenon. Inactivation of hagA, which may encode a tellurite resistance protein, had no effect on pilus structure or function. PMID- 11114901 TI - Characterization of a putative pathogenicity island from bovine Staphylococcus aureus encoding multiple superantigens. AB - Previous studies have demonstrated that a proportion of Staphylococcus aureus isolates from bovine mastitis coproduce toxic shock syndrome toxin (TSST) and staphylococcal enterotoxin C (SEC). In this study, molecular genetic analysis of one such strain, RF122, revealed the presence of a 15,891-bp putative pathogenicity island (SaPIbov) encoding the genes for TSST (tst), the SEC bovine variant (sec-bovine), and a gene (sel) which encodes an enterotoxin-like protein. The island contains 21 open reading frames specifying hypothetical proteins longer than 60 amino acids including an integrase-like gene. The element is bordered by 74-bp direct repeats at the left and right junctions, and the integration site lies adjacent to the 3' end of the GMP synthase gene (gmps) in the S. aureus chromosome. SaPIbov contains a central region of sequence identity with the previously characterized tst pathogenicity island SaPI1 (J. A. Lindsay et al., Mol. Microbiol. 29:527-543, 1998). A closely related strain, RF120, of the same multilocus enzyme electrophoretic type, random amplified polymorphic DNA type, and ribotype, does not contain the island, implying that the element is mobile and that a recent insertion/deletion event has taken place. TSST and TSST/SEC-deficient mutants of S. aureus strain RF122 were constructed by allele replacement. In vitro bovine Vbeta-specific lymphocyte expansion analysis by culture supernatants of wild-type strains and of tst and sec-bovine allele replacement mutants revealed that TSST stimulates BTB13-specific T cells whereas SEC-bovine stimulates BTB93-specific T cells. This suggests that the presence of SaPIbov may contribute to modulation of the bovine immune response. PMID- 11114902 TI - Recombinant Thermus aquaticus RNA polymerase, a new tool for structure-based analysis of transcription. AB - The three-dimensional structure of DNA-dependent RNA polymerase (RNAP) from thermophilic Thermus aquaticus has recently been determined at 3.3 A resolution. Currently, very little is known about T. aquaticus transcription and no genetic system to study T. aquaticus RNAP genes is available. To overcome these limitations, we cloned and overexpressed T. aquaticus RNAP genes in Escherichia coli. Overproduced T. aquaticus RNAP subunits assembled into functional RNAP in vitro and in vivo when coexpressed in E. coli. We used the recombinant T. aquaticus enzyme to demonstrate that transcription initiation, transcription termination, and transcription cleavage assays developed for E. coli RNAP can be adapted to study T. aquaticus transcription. However, T. aquaticus RNAP differs from the prototypical E. coli enzyme in several important ways: it terminates transcription less efficiently, has exceptionally high rate of intrinsic transcript cleavage, and is highly resistant to rifampin. Our results, together with the high-resolution structural information, should now allow a rational analysis of transcription mechanism by mutation. PMID- 11114903 TI - Identification of residues involved in catalytic activity of the inverting glycosyl transferase WbbE from Salmonella enterica serovar borreze. AB - Synthesis of the O:54 O antigen of Salmonella enterica is initiated by the nonprocessive glycosyl transferase WbbE, assigned to family 2 of the glycosyl transferase enzymes (GT2). GT2 enzymes possess a characteristic N-terminal domain, domain A. Based on structural data from the GT2 representative SpsA (S. J. Charnock and G. J. Davies, Biochemistry 38:6380-6385, 1999), this domain is responsible for nucleotide binding. It possesses two invariant Asp residues, the first forming a hydrogen bond to uracil and the second coordinating a Mn(2+) ion. Site-directed replacement of Asp41 (D41A) of WbbE, the analogue of the first Asp residue of SpsA, revealed that this is not required for activity. WbbE possesses three Asp residues near the position analogous to the second conserved residue. Whereas D95A reduced WbbE activity, activity in D93A and D96A mutants was abrogated, suggesting that either D93 or D96 may coordinate the Mn(2+) ion. Our studies also identified a C-terminal region of sequence conservation in 22 GT2 members, including WbbE. SpsA was not among these. This region is characterized by an ED(Y) motif. The Glu and Asp residues of this motif were individually replaced in WbbE. E180D in WbbE had greatly reduced activity, and an E180Q replacement completely abrogated activity; however, D181E had no effect. E180 is predicted to reside on a turn. Combined with the alignment of the motif with potential catalytic residues in the GT2 enzymes ExoM and SpsA, we speculate that E180 is the catalytic residue of WbbE. Sequence and predicted structural divergence in the catalytic region of GT2 members suggests that this is not a homogeneous family. PMID- 11114904 TI - trans-acting mutations in loci other than kdpDE that affect kdp operon regulation in Escherichia coli: effects of cytoplasmic thiol oxidation status and nucleoid protein H-NS on kdp expression. AB - Transcription of the K(+) transport operon kdp in Escherichia coli is induced during K(+)-limited growth by the action of a dual-component phosphorelay regulatory system comprised of a sensor kinase (integral membrane protein), KdpD, and a DNA-binding response regulator (cytoplasmic protein), KdpE. In this study, we screened for new dke (named dke for decreased kdp expression) mutations (in loci other than kdpDE) that led to substantially decreased kdp expression. One dke mutation was shown to be in hns, encoding the nucleoid protein H-NS. Another dke mutation was mapped to trxB (encoding thioredoxin reductase), and an equivalent reduction in kdp expression was demonstrated also for trxA mutants that are deficient in thioredoxin 1. Exogenously provided dithiothreitol rescued the kdp expression defect in trxB but not trxA mutants. Neither trxB nor trxA affected gene regulation mediated by another dual-component system tested, EnvZ OmpR. Mutations in genes dsbC and dsbD did not affect kdp expression, suggesting that the trx effects on kdp are not mediated by alterations in protein disulfide bond status in the periplasm. Reduced kdp expression was observed even in a trxB strain that harbored a variant KdpD polypeptide bearing no Cys residues. A trxB hns double mutant was even more severely affected for kdp expression than either single mutant. The dke mutations themselves had no effect on strength of the signal controlling kdp expression, and constitutive mutations in kdpDE were epistatic to hns and trxB. These results indicate that perturbations in cytoplasmic thiol oxidation status and in levels of the H-NS protein exert additive effects, direct or indirect, at a step(s) upstream of KdpD in the signal transduction pathway, which significantly influence the magnitude of KdpD kinase activity obtained for a given strength of the inducing signal for kdp transcription. PMID- 11114905 TI - Cloning of an intracellular Poly[D(-)-3-Hydroxybutyrate] depolymerase gene from Ralstonia eutropha H16 and characterization of the gene product. AB - An intracellular poly[D(-)-3-hydroxybutyrate] (PHB) depolymerase gene (phaZ) has been cloned from Ralstonia eutropha H16 by the shotgun method, sequenced, and characterized. Nucleotide sequence analysis of a 2.3-kbp DNA fragment revealed an open reading frame of 1,260 bp, encoding a protein of 419 amino acids with a predicted molecular mass of 47,316 Da. The crude extract of Escherichia coli containing the PHB depolymerase gene digested artificial amorphous PHB granules and released mainly oligomeric D(-)-3-hydroxybutyrate, with some monomer. The gene product did not hydrolyze crystalline PHB or freeze-dried artificial amorphous PHB granules. The deduced amino acid sequence lacked sequence corresponding to a classical lipase box, Gly-X-Ser-X-Gly. The gene product was expressed in R. eutropha cells concomitant with the synthesis of PHB and localized in PHB granules. Although a mutant of R. eutropha whose phaZ gene was disrupted showed a higher PHB content compared to the wild type in a nutrient rich medium, it accumulated PHB as much as the wild type did in a nitrogen-free, carbon-rich medium. These results indicate that the cloned phaZ gene encodes an intracellular PHB depolymerase in R. eutropha. PMID- 11114906 TI - Rubrerythrin and rubredoxin oxidoreductase in Desulfovibrio vulgaris: a novel oxidative stress protection system. AB - Evidence is presented for an alternative to the superoxide dismutase (SOD) catalase oxidative stress defense system in Desulfovibrio vulgaris (strain Hildenborough). This alternative system consists of the nonheme iron proteins, rubrerythrin (Rbr) and rubredoxin oxidoreductase (Rbo), the product of the rbo gene (also called desulfoferrodoxin). A Deltarbo strain of D. vulgaris was found to be more sensitive to internal superoxide exposure than was the wild type. Unlike Rbo, expression of plasmid-borne Rbr failed to restore the aerobic growth of a SOD-deficient strain of Escherichia coli. Conversely, plasmid-borne expression of two different Rbrs from D. vulgaris increased the viability of a catalase-deficient strain of E. coli that had been exposed to hydrogen peroxide whereas Rbo actually decreased the viability. A previously undescribed D. vulgaris gene was found to encode a protein having 50% sequence identity to that of E. coli Fe-SOD. This gene also encoded an extended N-terminal sequence with high homologies to export signal peptides of periplasmic redox proteins. The SOD activity of D. vulgaris is not affected by the absence of Rbo and is concentrated in the periplasmic fraction of cell extracts. These results are consistent with a superoxide reductase rather than SOD activity of Rbo and with a peroxidase activity of Rbr. A joint role for Rbo and Rbr as a novel cytoplasmic oxidative stress protection system in D. vulgaris and other anaerobic microorganisms is proposed. PMID- 11114907 TI - Characterization and evolution of anthranilate 1,2-dioxygenase from Acinetobacter sp. strain ADP1. AB - The two-component anthranilate 1,2-dioxygenase of the bacterium Acinetobacter sp. strain ADP1 was expressed in Escherichia coli and purified to homogeneity. This enzyme converts anthranilate (2-aminobenzoate) to catechol with insertion of both atoms of O(2) and consumption of one NADH. The terminal oxygenase component formed an alpha(3)beta(3) hexamer of 54- and 19-kDa subunits. Biochemical analyses demonstrated one Rieske-type [2Fe-2S] center and one mononuclear nonheme iron center in each large oxygenase subunit. The reductase component, which transfers electrons from NADH to the oxygenase component, was found to contain approximately one flavin adenine dinucleotide and one ferredoxin-type [2Fe-2S] center per 39-kDa monomer. Activities of the combined components were measured as rates and quantities of NADH oxidation, substrate disappearance, product appearance, and O(2) consumption. Anthranilate conversion to catechol was stoichiometrically coupled to NADH oxidation and O(2) consumption. The substrate analog benzoate was converted to a nonaromatic benzoate 1,2-diol with similarly tight coupling. This latter activity is identical to that of the related benzoate 1, 2-dioxygenase. A variant anthranilate 1,2-dioxygenase, previously found to convey temperature sensitivity in vivo because of a methionine-to-lysine change in the large oxygenase subunit, was purified and characterized. The purified M43K variant, however, did not hydroxylate anthranilate or benzoate at either the permissive (23 degrees C) or nonpermissive (39 degrees C) growth temperatures. The wild-type anthranilate 1,2-dioxygenase did not efficiently hydroxylate methylated or halogenated benzoates, despite its sequence similarity to broad substrate specific dioxygenases that do. Phylogenetic trees of the alpha and beta subunits of these terminal dioxygenases that act on natural and xenobiotic substrates indicated that the subunits of each terminal oxygenase evolved from a common ancestral two-subunit component. PMID- 11114908 TI - Carbon flux distribution and kinetics of cellulose fermentation in steady-state continuous cultures of Clostridium cellulolyticum on a chemically defined medium. AB - The metabolic characteristics of Clostridium cellulolyticum, a mesophilic cellulolytic nonruminal bacterium, were investigated and characterized kinetically for the fermentation of cellulose by using chemostat culture analysis. Since with C. cellulolyticum (i) the ATP/ADP ratio is lower than 1, (ii) the production of lactate at low specific growth rate (mu) is low, and (iii) there is a decrease of the NADH/NAD(+) ratio and q(NADH produced)/ q(NADH used) ratio as the dilution rate (D) increases in carbon-limited conditions, the chemostats used were cellulose-limited continuously fed cultures. Under all conditions, ethanol and acetate were the main end products of catabolism. There was no shift from an acetate-ethanol fermentation to a lactate-ethanol fermentation as previously observed on cellobiose as mu increased (E. Guedon, S. Payot, M. Desvaux, and H. Petitdemange, J. Bacteriol. 181:3262-3269, 1999). The acetate/ethanol ratio was always higher than 1 but decreased with D. On cellulose, glucose 6-phosphate and glucose 1-phosphate are important branch points since the longer the soluble beta-glucan uptake is, the more glucose 1 phosphate will be generated. The proportion of carbon flowing toward phosphoglucomutase remained constant (around 59.0%), while the carbon surplus was dissipated through exopolysaccharide and glycogen synthesis. The percentage of carbon metabolized via pyruvate-ferredoxin oxidoreductase decreased with D. Acetyl coenzyme A was mainly directed toward the acetate formation pathway, which represented a minimum of 27.1% of the carbon substrate. Yet the proportion of carbon directed through biosynthesis (i.e., biomass, extracellular proteins, and free amino acids) and ethanol increased with D, reaching 27.3 and 16.8%, respectively, at 0.083 h(-1). Lactate and extracellular pyruvate remained low, representing up to 1.5 and 0.2%, respectively, of the original carbon uptake. The true growth yield obtained on cellulose was higher, [50.5 g of cells (mol of hexose eq)(-1)] than on cellobiose, a soluble cellodextrin [36.2 g of cells (mol of hexose eq)(-1)]. The rate of cellulose utilization depended on the solid retention time and was first order, with a rate constant of 0.05 h(-1). Compared to cellobiose, substrate hydrolysis by cellulosome when bacteria are grown on cellulose fibers introduces an extra means for regulation of the entering carbon flow. This led to a lower mu, and so metabolism was not as distorted as previously observed with a soluble substrate. From these results, C. cellulolyticum appeared well adapted and even restricted to a cellulolytic lifestyle. PMID- 11114909 TI - Recombinational repair is critical for survival of Escherichia coli exposed to nitric oxide. AB - Nitric oxide (NO(.)) is critical to numerous biological processes, including signal transduction and macrophage-mediated immunity. In this study, we have explored the biological effects of NO(.)-induced DNA damage on Escherichia coli. The relative importance of base excision repair, nucleotide excision repair (NER), and recombinational repair in preventing NO(.)-induced toxicity was determined. E. coli strains lacking either NER or DNA glycosylases (including those that repair alkylation damage [alkA tag strain], oxidative damage [fpg nei nth strain], and deaminated cytosine [ung strain]) showed essentially wild-type levels of NO(.) resistance. However, apyrimidinic/apurinic (AP) endonuclease deficient cells (xth nfo strain) were very sensitive to killing by NO(.), which indicates that normal processing of abasic sites is critical for defense against NO(.). In addition, recA mutant cells were exquisitely sensitive to NO(.)-induced killing. Both SOS-deficient (lexA3) and Holliday junction resolvase-deficient (ruvC) cells were very sensitive to NO(.), indicating that both SOS and recombinational repair play important roles in defense against NO(.). Furthermore, strains specifically lacking double-strand end repair (recBCD strains) were very sensitive to NO(.), which suggests that NO(.) exposure leads to the formation of double-strand ends. One consequence of these double-strand ends is that NO(.) induces homologous recombination at a genetically engineered substrate. Taken together, it is now clear that, in addition to the known point mutagenic effects of NO(.), it is also important to consider recombination events among the spectrum of genetic changes that NO(. ) can induce. Furthermore, the importance of recombinational repair for cellular survival of NO(.) exposure reveals a potential susceptibility factor for invading microbes. PMID- 11114910 TI - Escherichia coli strains blocked in Tat-dependent protein export exhibit pleiotropic defects in the cell envelope. AB - The Tat system is a recently discovered protein export pathway that serves to translocate folded proteins, often containing redox cofactors, across the bacterial cytoplasmic membrane. Here we report that tat strains are associated with a mutant cell septation phenotype, where chains of up to 10 cells are evident. Mutant strains are also hypersensitive to hydrophobic drugs and to lysis by lysozyme in the absence of EDTA, and they leak periplasmic enzymes, characteristics that are consistent with an outer membrane defect. Both phenotypes are similar to those displayed by strains carrying point mutations in the lpxC (envA) gene. The phenotype was not replicated by mutations affecting synthesis and/or activity of all known or predicted Tat substrates. PMID- 11114911 TI - Targeted random mutagenesis to identify functionally important residues in the D2 protein of photosystem II in Synechocystis sp. strain PCC 6803. AB - To identify important residues in the D2 protein of photosystem II (PSII) in the cyanobacterium Synechocystis sp. strain PCC 6803, we randomly mutagenized a region of psbDI (coding for a 96-residue-long C-terminal part of D2) with sodium bisulfite. Mutagenized plasmids were introduced into a Synechocystis sp. strain PCC 6803 mutant that lacks both psbD genes, and mutants with impaired PSII function were selected. Nine D2 residues were identified that are important for PSII stability and/or function, as their mutation led to impairment of photoautotrophic growth. Five of these residues are likely to be involved in the formation of the Q(A)-binding niche; these are Ala249, Ser254, Gly258, Ala260, and His268. Three others (Gly278, Ser283, and Gly288) are in transmembrane alpha helix E, and their alteration leads to destabilization of PSII but not to major functional alterations of the remaining centers, indicating that they are unlikely to interact directly with cofactors. In the C-terminal lumenal tail of D2, only one residue (Arg294) was identified as functionally important for PSII. However, from the number of mutants generated it is likely that most or all of the 70 residues that are susceptible to bisulfite mutagenesis have been altered at least once. The fact that mutations in most of these residues have not been picked up by our screening method suggests that these mutations led to a normal photoautotrophic phenotype. A novel method of intragenic complementation in Synechocystis sp. strain PCC 6803 was developed to facilitate genetic analysis of psbDI mutants containing several amino acid changes in the targeted domain. Recombination between genome copies in the same cell appears to be much more prevalent in Synechocystis sp. strain PCC 6803 than was generally assumed. PMID- 11114912 TI - Renaturation of Bacillus thermoglucosidasius HrcA repressor by DNA and thermostability of the HrcA-DNA complex in vitro. AB - HrcA, a negative control repressor for chaperone expression from the obligate thermophile Bacillus thermoglucosidasius KP1006, was purified in a His-tagged form in the presence of 6 M urea but hardly renatured to an intact state due to extreme insolubility. Renaturation trials revealed that the addition of DNA to purified B. thermoglucosidasius HrcA can result in solubilization of HrcA free from the denaturing agent urea. Results from band shift and light scattering assays provided three new findings: (i) any species of DNA can serve to solubilize B. thermoglucosidasius HrcA, but DNA containing the CIRCE (controlling inverted repeat of chaperone expression) element is far more effective than other nonspecific DNA; (ii) B. thermoglucosidasius HrcA renatured with nonspecific DNA bound the CIRCE element in the molecular ratio of 2.6:1; and (iii) B. thermoglucosidasius HrcA binding to the CIRCE element was stable at below 50 degrees C whereas the complex was rapidly denatured at 70 degrees C, suggesting that the breakdown of HrcA is induced by heat stress and HrcA may act as a thermosensor to affect the expression of heat shock regulatory genes. These results will help to determine the nature of HrcA protein molecules. PMID- 11114913 TI - Control of the ferric citrate transport system of Escherichia coli: mutations in region 2.1 of the FecI extracytoplasmic-function sigma factor suppress mutations in the FecR transmembrane regulatory protein. AB - Transcription of the ferric citrate transport genes is initiated by binding of ferric citrate to the FecA protein in the outer membrane of Escherichia coli K 12. Bound ferric citrate does not have to be transported but initiates a signal that is transmitted by FecA across the outer membrane and by FecR across the cytoplasmic membrane into the cytoplasm, where the FecI extracytoplasmic-function (ECF) sigma factor becomes active. In this study, we isolated transcription initiation-negative missense mutants in the cytoplasmic region of FecR that were located at four sites, L13Q, W19R, W39R, and W50R, which are highly conserved in FecR-like open reading frames of the Pseudomonas aeruginosa, Pseudomonas putida, Bordetella pertussis, Bordetella bronchiseptica, and Caulobacter crescentus genomes. The cytoplasmic portion of the FecR mutant proteins, FecR(1-85), did not interact with wild-type FecI, in contrast to wild-type FecR(1-85), which induced FecI-mediated fecB transport gene transcription. Two missense mutations in region 2.1 of FecI, S15A and H20E, partially restored induction of ferric citrate transport gene induction of the fecR mutants by ferric citrate. Region 2.1 of sigma(70) is thought to bind RNA polymerase core enzyme; the residual activity of mutated FecI in the absence of FecR, however, was not higher than that of wild type FecI. In addition, missense mutations in the fecI promoter region resulted in a twofold increased transcription in fecR wild-type cells and a partial restoration of fec transport gene transcription in the fecR mutants. The mutations reduced binding of the Fe(2+) Fur repressor and as a consequence enhanced fecI transcription. The data reveal properties of the FecI ECF factor distinct from those of sigma(70) and further support the novel transcription initiation model in which the cytoplasmic portion of FecR is important for FecI activity. PMID- 11114914 TI - Component of the Rhodospirillum centenum photosensory apparatus with structural and functional similarity to methyl-accepting chemotaxis protein chemoreceptors. AB - Photosynthetic bacteria respond to alterations in light conditions by migrating to locations that allows optimal use of light as an energy source. Studies have indicated that photosynthesis-driven electron transport functions as an attractant signal for motility among purple photosynthetic bacteria. However, it is unclear just how the motility-based signal transduction system monitors electron flow through photosynthesis-driven electron transport. Recently, we have demonstrated that the purple photosynthetic bacterium Rhodospirillum centenum is capable of rapidly moving swarm cell colonies toward infrared light as well as away from visible light. Light-driven colony motility of R. centenum has allowed us to perform genetic dissection of the signaling pathway that affects photosynthesis-driven motility. In this study, we have undertaken sequence and mutational analyses of one of the components of a signal transduction pathway, Ptr, which appears responsible for transmitting a signal from the photosynthesis driven electron transport chain to the chemotaxis signal transduction cascade. Mutational analysis demonstrates that cells disrupted for ptr are defective in altering motility in response to light, as well as defective in light-dependent release of methanol. We present a model which proposes that Ptr senses the redox state of a component in the photosynthetic cyclic electron transport chain and that Ptr is responsible for transmitting a signal to the chemotaxis machinery to induce a photosynthesis-dependent motility response. PMID- 11114915 TI - pepA, a gene mediating pH regulation of virulence genes in Vibrio cholerae. AB - ToxT, a member of the AraC family of transcriptional regulators, controls the expression of several virulence factors in Vibrio cholerae. In the classical biotype of V. cholerae, expression of toxT is regulated by the same environmental conditions that control expression of the virulence determinants cholera toxin and the toxin coregulated pilus. Several genes that activate toxT expression have been identified. To identify genes that repress toxT expression in nonpermissive environmental conditions, a genetic screen was used to isolate mutations which alter the expression of a toxT-gusA transcriptional fusion. Several mutants were isolated, and the mutants could be divided into two classes. One class of mutants exhibited higher expression levels of toxT-gusA at both the nonpermissive pH and temperature, while the second class showed elevated toxT-gusA expression only at the nonpermissive pH. One mutant from the second class was chosen for further characterization. This mutant was found to carry a TnphoA insertion in a homolog of the Escherichia coli pepA gene. Disruption of pepA in V. cholerae resulted in elevated levels of expression of cholera toxin, tcpA, toxT, and tcpP at the noninducing pH but not at the noninducing temperature. Elevated levels of expression of toxT and tcpP at the nonpermissive pH in the pepA mutant were abolished in tcpP toxR mutant and aphB mutant backgrounds, respectively. A putative binding site for PepA was identified in the tcpPH-tcpI intergenic region, suggesting that PepA may act at the level of tcpPH transcription. Disruption of pepA caused only partial deregulation at the noninducing pH, suggesting the involvement of additional factors in the pH regulation of virulence genes in V. cholerae. PMID- 11114916 TI - Two conserved glutamates in the bacterial nitric oxide reductase are essential for activity but not assembly of the enzyme. AB - The bacterial nitric oxide reductase (NOR) is a divergent member of the family of respiratory heme-copper oxidases. It differs from other family members in that it contains an Fe(B)-heme-Fe dinuclear catalytic center rather than a Cu(B)-heme-Fe center and in that it does not pump protons. Several glutamate residues are conserved in NORs but are absent in other heme-copper oxidases. To facilitate mutagenesis-based studies of these residues in Paracoccus denitrificans NOR, we developed two expression systems that enable inactive or poorly active NOR to be expressed, characterized in vivo, and purified. These are (i) a homologous system utilizing the cycA promoter to drive aerobic expression of NOR in P. denitrificans and (ii) a heterologous system which provides the first example of the expression of an integral-membrane cytochrome bc complex in Escherichia coli. Alanine substitutions for three of the conserved glutamate residues (E125, E198, and E202) were introduced into NOR, and the proteins were expressed in P. denitrificans and E. coli. Characterization in intact cells and membranes has demonstrated that two of the glutamates are essential for normal levels of NOR activity: E125, which is predicted to be on the periplasmic surface close to helix IV, and E198, which is predicted to lie in the middle of transmembrane helix VI. The subsequent purification and spectroscopic characterization of these enzymes established that they are stable and have a wild-type cofactor composition. Possible roles for these glutamates in proton uptake and the chemistry of NO reduction at the active site are discussed. PMID- 11114917 TI - Differential responses of Escherichia coli cells expressing cytoplasmic domain mutants of penicillin-binding protein 1b after impairment of penicillin-binding proteins 1a and 3. AB - Penicillin-binding protein 1b (PBP1b) is the major high-molecular-weight PBP in Escherichia coli. Although it is coded by a single gene, it is usually found as a mixture of three isoforms which vary with regard to the length of their N terminal cytoplasmic tail. We show here that although the cytoplasmic tail seems to play no role in the dimerization of PBP1b, as was originally suspected, only the full-length protein is able to protect the cells against lysis when both PBP1a and PBP3 are inhibited by antibiotics. This suggests a specific role for the full-length PBP1b in the multienzyme peptidoglycan-synthesizing complex that cannot be fulfilled by either PBP1a or the shorter PBP1b proteins. Moreover, we have shown by alanine-stretch-scanning mutagenesis that (i) residues R(11) to G(13) are major determinants for correct translocation and folding of PBP1b and that (ii) the specific interactions involving the full-length PBP1b can be ascribed to the first six residues at the N-terminal end of the cytoplasmic domain. These results are discussed in terms of the interactions with other components of the murein-synthesizing complex. PMID- 11114918 TI - Initiation factor 2 of Myxococcus xanthus, a large version of prokaryotic translation initiation factor 2. AB - We have isolated the structural gene for translation initiation factor IF2 (infB) from the myxobacterium Myxococcus xanthus. The gene (3.22 kb) encodes a 1,070 residue protein showing extensive homology within its G domain and C terminus to the equivalent regions of IF2 from Escherichia coli. The protein cross-reacts with antibodies raised against E. coli IF2 and was able to complement an E. coli infB mutant. The M. xanthus protein is the largest IF2 known to date. This is essentially due to a longer N-terminal region made up of two characteristic domains. The first comprises a 188-amino-acid sequence consisting essentially of alanine, proline, valine, and glutamic acid residues, similar to the APE domain observed in Stigmatella aurantiaca IF2. The second is unique to M. xanthus IF2, is located between the APE sequence and the GTP binding domain, and consists exclusively of glycine, proline, and arginine residues. PMID- 11114919 TI - Methylotrophic Methylobacterium bacteria nodulate and fix nitrogen in symbiosis with legumes. AB - Rhizobia described so far belong to three distinct phylogenetic branches within the alpha-2 subclass of Proteobacteria. Here we report the discovery of a fourth rhizobial branch involving bacteria of the Methylobacterium genus. Rhizobia isolated from Crotalaria legumes were assigned to a new species, "Methylobacterium nodulans," within the Methylobacterium genus on the basis of 16S ribosomal DNA analyses. We demonstrated that these rhizobia facultatively grow on methanol, which is a characteristic of Methylobacterium spp. but a unique feature among rhizobia. Genes encoding two key enzymes of methylotrophy and nodulation, the mxaF gene, encoding the alpha subunit of the methanol dehydrogenase, and the nodA gene, encoding an acyltransferase involved in Nod factor biosynthesis, were sequenced for the type strain, ORS2060. Plant tests and nodA amplification assays showed that "M. nodulans" is the only nodulating Methylobacterium sp. identified so far. Phylogenetic sequence analysis showed that "M. nodulans" NodA is closely related to Bradyrhizobium NodA, suggesting that this gene was acquired by horizontal gene transfer. PMID- 11114920 TI - A long T. A tract in the upp initially transcribed region is required for regulation of upp expression by UTP-dependent reiterative transcription in Escherichia coli. AB - In Escherichia coli, pyrimidine-mediated regulation of upp expression occurs by UTP-sensitive selection of alternative transcriptional start sites, which produces transcripts that differ in the ability to be elongated. The upp initially transcribed region contains the sequence GATTTTTTTTG (nontemplate strand). Initiation can occur at either the first or the second base in this sequence (designated G6 and A7, with numbering from the promoter -10 region). High intracellular UTP levels favor initiation at position A7; however, the resulting transcripts are subject to reiterative transcription (i.e., repetitive UMP addition) within the 8-bp T. A tract in the initially transcribed region and are aborted. In contrast, low intracellular UTP levels favor initiation at position G6, which results in transcripts that can, in part, avoid reiterative transcription and be elongated normally. In this study, we examined the regulatory requirement for the long T. A tract in the upp initially transcribed region. We constructed upp promoter mutations that shorten the T. A tract to 7, 6, 5, 4, 3, or 2 bp and examined the effects of these mutations on upp expression and regulation. The results indicate that pyrimidine-mediated regulation is gradually reduced as the T. A tract is shortened from 7 to 3 bp; at which point regulation ceases. This reduction in regulation is due to large-percentage increases in upp expression in cells grown under conditions of pyrimidine excess. Quantitation of cellular transcripts and in vitro transcription studies indicate that the observed effects of a shortened T. A tract on upp expression and regulation are due to increases in the fraction of both G6- and A7-initiated transcripts that avoid reiterative transcription and are elongated normally. PMID- 11114921 TI - The KlGpa1 gene encodes a G-protein alpha subunit that is a positive control element in the mating pathway of the budding yeast Kluyveromyces lactis. AB - The cloning of the gene encoding the KlGpa1p subunit was achieved by standard PCR techniques and by screening a Kluyveromyces lactis genomic library using the PCR product as a probe. The full-length open reading frame spans 1,344 nucleotides including the stop codon. The deduced primary structure of the protein (447 amino acid residues) strongly resembles that of Gpa1p, the G-protein alpha subunit from Saccharomyces cerevisiae involved in the mating pheromone response pathway. Nevertheless, unlike disruption of Gpa1 from S. cerevisiae, disruption of KlGpa1 rendered viable cells with a reduced capacity to mate. Expression of a plasmidic KlGpa1 copy in a DeltaKlgpa1 mutant restores full mating competence; hence we conclude that KlGpa1p plays a positive role in the mating pathway. Overexpression of the constitutive subunit KlGpa1p(K(364)) (GTP bound) does not induce constitutive mating; instead it partially blocks wild-type mating and is unable to reverse the sterile phenotype of DeltaKlgpa1 mutant cells. K. lactis expresses a second Galpha subunit, KlGpa2p, which is involved in regulating cyclic AMP levels upon glucose stimulation. This subunit does not rescue DeltaKlgpa1 cells from sterility; instead, overproduction of KlGpa2p slightly reduces the mating of wild-type cells, suggesting cross talk within the pheromone response pathway mediated by KlGpa1p and glucose metabolism mediated by KlGpa2p. The DeltaKlgpa1 DeltaKlgpa2 double mutant, although viable, showed the mating deficiency observed in the single DeltaKlgpa1 mutant. PMID- 11114922 TI - Characterization of In53, a class 1 plasmid- and composite transposon-located integron of Escherichia coli which carries an unusual array of gene cassettes. AB - Further characterization of the genetic environment of the gene encoding the Escherichia coli extended-spectrum beta-lactamase, bla(VEB-1), revealed the presence of a plasmid-located class 1 integron, In53, which carried eight functional resistance gene cassettes in addition to bla(VEB-1). While the aadB and the arr-2 gene cassettes were identical to those previously described, the remaining cassettes were novel: (i) a novel nonenzymatic chloramphenicol resistance gene of the cmlA family, (ii) a qac allele encoding a member of the small multidrug resistance family of proteins, (iii) a cassette, aacA1b/orfG, which encodes a novel 6'-N-acetyltransferase, and (iv) a fused gene cassette, oxa10/aadA1, which is made of two cassettes previously described as single cassettes. In addition, oxa10 and aadA1 genes were expressed from their own promoter sequence present upstream of the oxa10 cassette. arr-2 coded for a protein that shared 54% amino acid identity with the rifampin ADP-ribosylating transferase encoded by the arr-1 gene from Mycobacterium smegmatis DSM43756. While in M. smegmatis, the main inactivated compound was 23-ribosyl-rifampin, the inactivated antibiotic recovered from E. coli culture was 23-O-ADP-ribosyl rifampin. The integrase gene of In53 was interrupted by an IS26 insertion sequence, which was also present in the 3' conserved segment. Thus, In53 is a truncated integron located on a composite transposon, named Tn2000, bounded by two IS26 elements in opposite orientations. Target site duplication at both ends of the transposon indicated that the integron likely was inserted into the plasmid through a transpositional process. This is the first description of an integron located on a composite transposon. PMID- 11114923 TI - Role of the dinitrogenase reductase arginine 101 residue in dinitrogenase reductase ADP-ribosyltransferase binding, NAD binding, and cleavage. AB - Dinitrogenase reductase is posttranslationally regulated by dinitrogenase reductase ADP-ribosyltransferase (DRAT) via ADP-ribosylation of the arginine 101 residue in some bacteria. Rhodospirillum rubrum strains in which the arginine 101 of dinitrogenase reductase was replaced by tyrosine, phenylalanine, or leucine were constructed by site-directed mutagenesis of the nifH gene. The strain containing the R101F form of dinitrogenase reductase retains 91%, the strain containing the R101Y form retains 72%, and the strain containing the R101L form retains only 28% of in vivo nitrogenase activity of the strain containing the dinitrogenase reductase with arginine at position 101. In vivo acetylene reduction assays, immunoblotting with anti-dinitrogenase reductase antibody, and [adenylate-(32)P]NAD labeling experiments showed that no switch-off of nitrogenase activity occurred in any of the three mutants and no ADP-ribosylation of altered dinitrogenase reductases occurred either in vivo or in vitro. Altered dinitrogenase reductases from strains UR629 (R101Y) and UR630 (R101F) were purified to homogeneity. The R101F and R101Y forms of dinitrogenase reductase were able to form a complex with DRAT that could be chemically cross-linked by 1 ethyl-3-(3-dimethylaminopropyl)-carbodiimide. The R101F form of dinitrogenase reductase and DRAT together were not able to cleave NAD. This suggests that arginine 101 is not critical for the binding of DRAT to dinitrogenase reductase but that the availability of arginine 101 is important for NAD cleavage. Both DRAT and dinitrogenase reductase can be labeled by [carbonyl-(14)C]NAD individually upon UV irradiation, but most (14)C label is incorporated into DRAT when both proteins are present. The ability of R101F dinitrogenase reductase to be labeled by [carbonyl-(14)C]NAD suggested that Arg 101 is not absolutely required for NAD binding. PMID- 11114924 TI - Identification of ccdA in Paracoccus pantotrophus GB17: disruption of ccdA causes complete deficiency in c-type cytochromes. AB - A transposon Tn5-mob insertional mutant of Paracoccus pantotrophus GB17, strain TP43, was unable to oxidize thiosulfate aerobically or to reduce nitrite anaerobically, and the cellular yields were generally decreased by 11 to 20%. Strain TP43 was unable to form functional c-type cytochromes, as determined by difference spectroscopy and heme staining. However, formation of apocytochromes and their transport to the periplasm were not affected, as seen with SoxD, a c type cytochrome associated with the periplasmic sulfite dehydrogenase homologue. The Tn5-mob-containing DNA region of strain TP43 was cloned into pSUP205 to produce pE18TP43. With the aid of pE18TP43 the corresponding wild-type gene region of 15 kb was isolated from a heterogenote recombinant to produce pEF15. Sequence analysis of 2.8 kb of the relevant region uncovered three open reading frames, designated ORFA, ccdA, and ORFB, with the latter being oriented divergently. ORFA and ccdA were constitutively cotranscribed as determined by primer extension analysis. In strain TP43 Tn5-mob was inserted into ccdA. The deduced ORFA product showed no similarity to any protein in databases. However, the ccdA gene product exhibited similarities to proteins assigned to different functions in bacteria, such as cytochrome c biogenesis. For these proteins at least six transmembrane helices are predicted with the potential to form a channel with two conserved cysteines. This structural identity suggests that these proteins transfer reducing equivalents from the cytoplasm to the periplasm and that the cysteines bring about this transfer to enable the various specific functions via specific redox mediators such as thioredoxins. CcdA of P. pantotrophus is 42% identical to a protein predicted by ORF2, and its location within the sox gene cluster coding for lithotrophic sulfur oxidation suggested a different function. PMID- 11114925 TI - Intragenic suppressors of an OmpF assembly mutant and assessment of the roles of various OmpF residues in assembly through informational suppressors. AB - We employed two separate genetic approaches to examine the roles of various OmpF residues in assembly. In one approach, intragenic suppressors of a temperature sensitive OmpF assembly mutant carrying a W214E substitution were sought at 42 degrees C, or at 37 degrees C in a genetic background lacking the periplasmic folding factor SurA. In the majority of cases (58 out of 61 revertants), the suppressors mapped either at the original site (position 214) or two residues downstream from it. In the remaining three revertants that were obtained in a surA background, an alteration of N230Y was located 16 residues away from the original site. The N230Y suppressor also corrected OmpF315 assembly at 42 degrees C in a surA(+) background, indicating that the two different physiological environments imposed similar assembly constraints. The specificity of N230Y was tested against five different residues at position 214 of mature OmpF. Clear specificity was displayed, with maximum suppression observed for the original substitution at position 214 (E214) against which the N230Y suppressor was isolated, and no negative effect on OmpF assembly was noted when the wild-type W214 residue was present. The mechanism of suppression may involve compensation for a specific conformational defect. The second approach involved the application of informational suppressors (Su-tRNA) in combination with ompF amber mutations to generate variant OmpF proteins. In this approach we targeted the Y40, Q66, W214, and Y231 residues of mature OmpF and replaced them with S, Q, L, and Y through the action of Su-tRNAs. Thus, a total of 16 variant OmpF proteins were generated, of which three were identical to the parental protein, and two variants carrying W214Q and Y231Q substitutions were similar to assembly defective proteins isolated previously (R. Misra, J. Bacteriol. 175:5049-5056, 1993). The results obtained from these analyses provided useful information regarding the compatibility of various alterations in OmpF assembly. PMID- 11114926 TI - Transcriptional organization and dynamic expression of the hbpCAD genes, which encode the first three enzymes for 2-hydroxybiphenyl degradation in Pseudomonas azelaica HBP1. AB - Pseudomonas azelaica HBP1 degrades the toxic substance 2-hydroxybiphenyl (2-HBP) by means of three enzymes that are encoded by structural genes hbpC, hbpA, and hbpD. These three genes form a small noncontiguous cluster. Their expression is activated by the product of regulatory gene hbpR, which is located directly upstream of the hbpCAD genes. The HbpR protein is a transcription activator and belongs to the so-called XylR/DmpR subclass within the NtrC family of transcriptional activators. Transcriptional fusions between the different hbp intergenic regions and the luxAB genes of Vibrio harveyi in P. azelaica and in Escherichia coli revealed the existence of two HbpR-regulated promoters; one is located in front of hbpC, and the other one is located in front of hbpD. Northern analysis confirmed that the hbpC and hbpA genes are cotranscribed, whereas the hbpD gene is transcribed separately. No transcripts comprising the entire hbpCAD cluster were detected, indicating that transcription from P(hbpC) is terminated after the hbpA gene. E. coli mutant strains lacking the structural genes for the RNA polymerase sigma(54) subunit or for the integration host factor failed to express bioluminescence from P(hbpC)- and P(hbpD)-luxAB fusions when a functional hbpR gene was provided in trans. This pointed to the active role of sigma(54) and integration host factor in transcriptional activation from these promoters. Primer extension analysis revealed that both P(hbpC) and P(hbpD) contain the typical motifs at position -24 (GG) and -12 (GC) found in sigma(54)-dependent promoters. Analysis of changes in the synthesis of the hbp mRNAs, in activities of the 2-HBP pathway enzymes, and in concentrations of 2-HBP intermediates during the first 4 h after induction of continuously grown P. azelaica cells with 2-HBP demonstrated that the specific transcriptional organization of the hbp genes ensured smooth pathway expression. PMID- 11114927 TI - Effect on heterocyst differentiation of nitrogen fixation in vegetative cells of the cyanobacterium Anabaena variabilis ATCC 29413. AB - Heterocysts are terminally differentiated cells of some filamentous cyanobacteria that fix nitrogen for the entire filament under oxic growth conditions. Anabaena variabilis ATCC 29413 is unusual in that it has two Mo-dependent nitrogenases; one, called Nif1, functions in heterocysts, while the second, Nif2, functions under anoxic conditions in vegetative cells. Both nitrogenases depended on expression of the global regulatory protein NtcA. It has long been thought that a product of nitrogen fixation in heterocysts plays a role in maintenance of the spaced pattern of heterocyst differentiation. This model assumes that each cell in a filament senses its own environment in terms of nitrogen sufficiency and responds accordingly in terms of differentiation. Expression of the Nif2 nitrogenase under anoxic conditions in vegetative cells was sufficient to support long-term growth of a nif1 mutant; however, that expression did not prevent differentiation of heterocysts and expression of the nif1 nitrogenase in either the nif1 mutant or the wild-type strain. This suggested that the nitrogen sufficiency of individual cells in the filament did not affect the signal that induces heterocyst differentiation. Perhaps there is a global mechanism by which the filament senses nitrogen sufficiency or insufficiency based on the external availability of fixed nitrogen. The filament would then respond by producing heterocyst differentiation signals that affect the entire filament. This does not preclude cell-to-cell signaling in the maintenance of heterocyst pattern but suggests that overall control of the process is not controlled by nitrogen insufficiency of individual cells. PMID- 11114928 TI - Secreted euryarchaeal microhalocins kill hyperthermophilic crenarchaea. AB - Few antibiotics targeting members of the archaeal domain are currently available for genetic studies. Since bacterial antibiotics are frequently directed against competing and related organisms, archaea by analogy might produce effective antiarchaeal antibiotics. Peptide antibiotic (halocin) preparations from euryarchaeal halophilic strains S8a, GN101, and TuA4 were found to be toxic for members of the hyperthermophilic crenarchaeal genus Sulfolobus. No toxicity was evident against representative bacteria or eukarya. Halocin S8 (strain S8a) and halocin R1 (strain GN101) preparations were cytostatic, while halocin A4 (strain TuA4) preparations were cytocidal. Subsequent studies focused on the use of halocin A4 preparations and Sulfolobus solfataricus. Strain TuA4 cell lysates were not toxic for S. solfataricus, and protease (but not nuclease) treatment of the halocin A4 preparation inactivated toxicity, indicating that the A4 toxic factor must be a secreted protein. Potassium chloride supplementation of the Sulfolobus assay medium potentiated toxicity, implicating use of a salt-dependent mechanism. The utility of halocin A4 preparations for genetic manipulation of S. solfataricus was assessed through the isolation of UV-induced resistant mutants. The mutants exhibited stable phenotypes and were placed into distinct classes based on their levels of resistance. PMID- 11114929 TI - Archaeal shikimate kinase, a new member of the GHMP-kinase family. AB - Shikimate kinase (EC 2.7.1.71) is a committed enzyme in the seven-step biosynthesis of chorismate, a major precursor of aromatic amino acids and many other aromatic compounds. Genes for all enzymes of the chorismate pathway except shikimate kinase are found in archaeal genomes by sequence homology to their bacterial counterparts. In this study, a conserved archaeal gene (gi1500322 in Methanococcus jannaschii) was identified as the best candidate for the missing shikimate kinase gene by the analysis of chromosomal clustering of chorismate biosynthetic genes. The encoded hypothetical protein, with no sequence similarity to bacterial and eukaryotic shikimate kinases, is distantly related to homoserine kinases (EC 2.7.1.39) of the GHMP-kinase superfamily. The latter functionality in M. jannaschii is assigned to another gene (gi591748), in agreement with sequence similarity and chromosomal clustering analysis. Both archaeal proteins, overexpressed in Escherichia coli and purified to homogeneity, displayed activity of the predicted type, with steady-state kinetic parameters similar to those of the corresponding bacterial kinases: K(m,shikimate) = 414 +/- 33 microM, K(m,ATP) = 48 +/- 4 microM, and k(cat) = 57 +/- 2 s(-1) for the predicted shikimate kinase and K(m,homoserine) = 188 +/- 37 microM, K(m,ATP) = 101 +/- 7 microM, and k(cat) = 28 +/- 1 s(-1) for the homoserine kinase. No overlapping activity could be detected between shikimate kinase and homoserine kinase, both revealing a >1,000 fold preference for their own specific substrates. The case of archaeal shikimate kinase illustrates the efficacy of techniques based on reconstruction of metabolism from genomic data and analysis of gene clustering on chromosomes in finding missing genes. PMID- 11114930 TI - Proteome analysis of metabolically engineered Escherichia coli producing Poly(3 hydroxybutyrate). AB - Recombinant Escherichia coli strains harboring heterologous polyhydroxyalkanoate (PHA) biosynthesis genes were shown to accumulate unusually large amounts of PHA. In the present study, integrated cellular responses of metabolically engineered E. coli to the accumulation of poly(3-hydroxybutyrate) (PHB) in the early stationary phase were analyzed at the protein level by two-dimensional gel electrophoresis. Out of 20 proteins showing altered expression levels with the accumulation of PHB, 13 proteins were identified with the aid of mass spectrometry. Three heat shock proteins, GroEL, GroES, and DnaK, were significantly up-regulated in PHB-accumulating cells. Proteins which play essential roles in protein biosynthesis were unfavorably influenced by the accumulation of PHB. Cellular demand for the large amount of acetyl coenzyme A and NADPH for the PHB biosynthesis resulted in the increased synthesis of two enzymes of the glycolytic pathway and one enzyme of the Entner-Doudoroff pathway. The expression of the yfiD gene encoding a 14.3-kDa protein, which is known to be produced at low pH, was greatly induced with the accumulation of PHB. Therefore, it could be concluded that the accumulation of PHB in E. coli acted as a stress on the cells, which reduced the cells' ability to synthesize proteins and induced the expression of various protective proteins. PMID- 11114931 TI - Vibrio fischeri genes hvnA and hvnB encode secreted NAD(+)-glycohydrolases. AB - HvnA and HvnB are proteins secreted by Vibrio fischeri ES114, an extracellular light organ symbiont of the squid Euprymna scolopes, that catalyze the transfer of ADP-ribose from NAD(+) to polyarginine. Based on this activity, HvnA and HvnB were presumptively designated mono-ADP-ribosyltransferases (ARTases), and it was hypothesized that they mediate bacterium-host signaling. We have cloned hvnA and hvnB from strain ES114. hvnA appears to be expressed as part of a four-gene operon, whereas hvnB is monocistronic. The predicted HvnA and HvnB amino acid sequences are 46% identical to one another and share 44% and 34% identity, respectively, with an open reading frame present in the Pseudomonas aeruginosa genome. Four lines of evidence indicate that HvnA and HvnB mediate polyarginine ADP-ribosylation not by ARTase activity, but indirectly through an NAD(+) glycohydrolase (NADase) activity that releases free, reactive, ADP-ribose: (i) like other NADases, and in contrast to the ARTase cholera toxin, HvnA and HvnB catalyzed ribosylation of not only polyarginine but also polylysine and polyhistidine, and ribosylation was inhibited by hydroxylamine; (ii) HvnA and HvnB cleaved 1, N(6)-etheno-NAD(+) and NAD(+); (iii) incubation of HvnA and HvnB with [(32)P]NAD(+) resulted in the production of ADP-ribose; and (iv) purified HvnA displayed an NADase V(max) of 400 mol min(-1) mol(-1), which is within the range reported for other NADases and 10(2)- to 10(4)-fold higher than the minor NADase activity reported in bacterial ARTase toxins. Construction and analysis of an hvnA hvnB mutant revealed no other NADase activity in culture supernatants of V. fischeri, and this mutant initiated the light organ symbiosis and triggered regression of the light organ ciliated epithelium in a manner similar to that for the wild type. PMID- 11114932 TI - phzO, a gene for biosynthesis of 2-hydroxylated phenazine compounds in Pseudomonas aureofaciens 30-84. AB - Certain strains of root-colonizing fluorescent Pseudomonas spp. produce phenazines, a class of antifungal metabolites that can provide protection against various soilborne root pathogens. Despite the fact that the phenazine biosynthetic locus is highly conserved among fluorescent Pseudomonas spp., individual strains differ in the range of phenazine compounds they produce. This study focuses on the ability of Pseudomonas aureofaciens 30-84 to produce 2 hydroxyphenazine-1-carboxylic acid (2-OH-PCA) and 2-hydroxyphenazine from the common phenazine metabolite phenazine-1-carboxylic acid (PCA). P. aureofaciens 30 84 contains a novel gene located downstream from the core phenazine operon that encodes a 55-kDa aromatic monooxygenase responsible for the hydroxylation of PCA to produce 2-OH-PCA. Knowledge of the genes responsible for phenazine product specificity could ultimately reveal ways to manipulate organisms to produce multiple phenazines or novel phenazines not previously described. PMID- 11114933 TI - Alternative pathways for siroheme synthesis in Klebsiella aerogenes. AB - Siroheme, the cofactor for sulfite and nitrite reductases, is formed by methylation, oxidation, and iron insertion into the tetrapyrrole uroporphyrinogen III (Uro-III). The CysG protein performs all three steps of siroheme biosynthesis in the enteric bacteria Escherichia coli and Salmonella enterica. In either taxon, cysG mutants cannot reduce sulfite to sulfide and require a source of sulfide or cysteine for growth. In addition, CysG-mediated methylation of Uro-III is required for de novo synthesis of cobalamin (coenzyme B(12)) in S. enterica. We have determined that cysG mutants of the related enteric bacterium Klebsiella aerogenes have no defect in the reduction of sulfite to sulfide. These data suggest that an alternative enzyme allows for siroheme biosynthesis in CysG deficient strains of Klebsiella. However, Klebsiella cysG mutants fail to synthesize coenzyme B(12), suggesting that the alternative siroheme biosynthetic pathway proceeds by a different route. Gene cysF, encoding an alternative siroheme synthase homologous to CysG, has been identified by genetic analysis and lies within the cysFDNC operon; the cysF gene is absent from the E. coli and S. enterica genomes. While CysG is coregulated with the siroheme-dependent nitrite reductase, the cysF gene is regulated by sulfur starvation. Models for alternative regulation of the CysF and CysG siroheme synthases in Klebsiella and for the loss of the cysF gene from the ancestor of E. coli and S. enterica are presented. PMID- 11114934 TI - Methionine-to-cysteine recycling in Klebsiella aerogenes. AB - In the enteric bacteria Escherichia coli and Salmonella enterica, sulfate is reduced to sulfide and assimilated into the amino acid cysteine; in turn, cysteine provides the sulfur atom for other sulfur-bearing molecules in the cell, including methionine. These organisms cannot use methionine as a sole source of sulfur. Here we report that this constraint is not shared by many other enteric bacteria, which can use either cysteine or methionine as the sole source of sulfur. The enteric bacterium Klebsiella aerogenes appears to use at least two pathways to allow the reduced sulfur of methionine to be recycled into cysteine. In addition, the ability to recycle methionine on solid media, where cys mutants cannot use methionine as a sulfur source, appears to be different from that in liquid media, where they can. One pathway likely uses a cystathionine intermediate to convert homocysteine to cysteine and is induced under conditions of sulfur starvation, which is likely sensed by low levels of the sulfate reduction intermediate adenosine-5'-phosphosulfate. The CysB regulatory proteins appear to control activation of this pathway. A second pathway may use a methanesulfonate intermediate to convert methionine-derived methanethiol to sulfite. While the transsulfurylation pathway may be directed to recovery of methionine, the methanethiol pathway likely represents a general salvage mechanism for recovery of alkane sulfide and alkane sulfonates. Therefore, the relatively distinct biosyntheses of cysteine and methionine in E. coli and Salmonella appear to be more intertwined in Klebsiella. PMID- 11114935 TI - Genetic and biochemical characterization of a novel umuD mutation: insights into a mechanism for UmuD self-cleavage. AB - Most translesion DNA synthesis (TLS) in Escherichia coli is dependent upon the products of the umuDC genes, which encode a DNA polymerase, DNA polymerase V, with the unique ability to replicate over a variety of DNA lesions, including cyclobutane dimers and abasic sites. The UmuD protein is activated for its role in TLS by a RecA-single-stranded DNA (ssDNA)-facilitated self-cleavage event that serves to remove its amino-terminal 24 residues to yield UmuD'. We have used site directed mutagenesis to construct derivatives of UmuD and UmuD' with glycines in place of leucine-101 and arginine-102. These residues are extremely well conserved among the UmuD-like proteins involved in mutagenesis but are poorly conserved among the structurally related LexA-like transcriptional repressor proteins. Based on both the crystal and solution structures of the UmuD' homodimer, these residues are part of a solvent-exposed loop. Our genetic and biochemical characterizations of these mutant UmuD and UmuD' proteins indicate that while leucine-101 and arginine-102 are critical for the RecA-ssDNA facilitated self-cleavage of UmuD, they serve only a minimal role in enabling TLS. These results, and others, suggest that the interaction of RecA-ssDNA with leucine-101 and arginine-102, together with numerous other contacts between UmuD(2) and the RecA-ssDNA nucleoprotein filaments, serves to realign lysine-97 relative to serine-60, thereby activating UmuD(2) for self-cleavage. PMID- 11114936 TI - Phylogeny of the major head and tail genes of the wide-ranging T4-type bacteriophages. AB - We examined a number of bacteriophages with T4-type morphology that propagate in different genera of enterobacteria, Aeromonas, Burkholderia, and Vibrio. Most of these phages had a prolate icosahedral head, a contractile tail, and a genome size that was similar to that of T4. A few of them had more elongated heads and larger genomes. All these phages are phylogenetically related, since they each had sequences homologous to the capsid gene (gene 23), tail sheath gene (gene 18), and tail tube gene (gene 19) of T4. On the basis of the sequence comparison of their virion genes, the T4-type phages can be classified into three subgroups with increasing divergence from T4: the T-evens, pseudoT-evens, and schizoT evens. In general, the phages that infect closely related host species have virion genes that are phylogenetically closer to each other than those of phages that infect distantly related hosts. However, some of the phages appear to be chimeras, indicating that, at least occasionally, some genetic shuffling has occurred between the different T4-type subgroups. The compilation of a number of gene 23 sequences reveals a pattern of conserved motifs separated by sequences that differ in the T4-type subgroups. Such variable patches in the gene 23 sequences may determine the size of the virion head and consequently the viral genome length. This sequence analysis provides molecular evidence that phages related to T4 are widespread in the biosphere and diverged from a common ancestor in acquiring the ability to infect different host bacteria and to occupy new ecological niches. PMID- 11114937 TI - Evaluation of a structural model of Pseudomonas aeruginosa outer membrane protein OprM, an efflux component involved in intrinsic antibiotic resistance. AB - The outer membrane protein OprM of Pseudomonas aeruginosa is involved in intrinsic and mutational multiple-antibiotic resistance as part of two resistance nodulation-division efflux systems. The crystal structure of TolC, a homologous protein in Escherichia coli, was recently published (V. Koronakis, A. Sharff, E. Koronakis, B. Luisl, and C. Hughes, Nature 405:914-919, 2000), demonstrating a distinctive architecture comprising outer membrane beta-barrel and periplasmic helical-barrel structures, which assemble differently from the common beta-barrel only conformation of porins. Based on their sequence similarity, a similar content of alpha-helical and beta-sheet structure determined by circular dichroism spectroscopy, and our observation that OprM, like TolC, reconstitutes channels in planar bilayer membranes, OprM and TolC were considered to be structurally homologous, and a model of OprM was constructed by threading its sequence to the TolC crystal structure. Residues thought to be important for the TolC structure were conserved in space in this OprM model. Analyses of deletion mutants and previously isolated insertion mutants of OprM in the context of this model allowed us to propose roles for different protein domains. Our data indicate that the helical barrel of the protein is critical for both the function and the integrity of the protein, while a C-terminal domain localized around the equatorial plane of this helical barrel is dispensable. Extracellular loops appear to play a lesser role in substrate specificity for this efflux protein compared to classical porins, and there appears to be a correlation between the change in antimicrobial activity for OprM mutants and the pore size. Our model and channel formation studies support the "iris" mechanism of action for TolC and permit us now to form more focused hypotheses about the functional domains of OprM and its related family of efflux proteins. PMID- 11114938 TI - Characterization of transmembrane segments 3, 4, and 5 of MalF by mutational analysis. AB - MalF and MalG are the cytoplasmic membrane components of the binding protein dependent ATP binding cassette maltose transporter in Escherichia coli. They are thought to form the transport channel and are thus of critical importance for the mechanism of transport. To study the contributions of individual transmembrane segments of MalF, we isolated 27 point mutations in membrane-spanning segments 3, 4, and 5. These data complement a previous study, which described the mutagenesis of membrane-spanning segments 6, 7, and 8. While most of the isolated mutations appear to cause assembly defects, L(323)Q in helix 5 could interfere more directly with substrate specificity. The phenotypes and locations of the mutations are consistent with a previously postulated structural model of MalF. PMID- 11114939 TI - Quorum sensing in Vibrio fischeri: analysis of the LuxR DNA binding region by alanine-scanning mutagenesis. AB - LuxR is the transcriptional activator for quorum-sensing control of luminescence in Vibrio fischeri. A series of alanine-scanning mutants spanning a predicted helix-turn-helix region in the DNA binding domain of LuxR was constructed, and the activity of each of the LuxR mutant proteins in recombinant Escherichia coli was investigated. The region covered by the mutagenesis spanned residues 190 to 224. About half of the alanine-scanning mutants showed activities similar to that of the wild-type LuxR: at least two were positive-control mutants, four appeared to be defective in DNA binding, and several others were characterized as DNA binding affinity mutants. This analysis, taken together with information about other bacterial transcription factors, provides insights into amino acid residues in LuxR that are involved in DNA binding and transcriptional activation. PMID- 11114940 TI - Amino acid residues in LuxR critical for its mechanism of transcriptional activation during quorum sensing in Vibrio fischeri. AB - PCR-based site-directed mutagenesis has been used to generate 38 alanine substitution mutations in the C-terminal 41 amino acid residues of LuxR. This region plays a critical role in the mechanism of LuxR-dependent transcriptional activation of the Vibrio fischeri lux operon during quorum sensing. The ability of the variant forms of LuxR to activate transcription of the lux operon was examined by using in vivo assays in recombinant Escherichia coli. Eight recombinant strains produced luciferase at levels less than 50% of that of a strain expressing wild-type LuxR. Western immunoblotting analysis verified that the altered forms of LuxR were expressed at levels equivalent to those of the wild type. An in vivo DNA binding-repression assay in recombinant E. coli was subsequently used to measure the ability of the variant forms of LuxR to bind to the lux box, the binding site of LuxR at the lux operon promoter. All eight LuxR variants found to affect cellular luciferase levels were unable to bind to the lux box. An additional 11 constructs that had no effect on cellular luciferase levels were also found to exhibit a defect in DNA binding. None of the alanine substitutions in LuxR affected activation of transcription of the lux operon without also affecting DNA binding. These results support the conclusion that the C-terminal 41 amino acids of LuxR are important for DNA recognition and binding of the lux box rather than positive control of the process of transcription initiation. PMID- 11114941 TI - Role for hetC in the transition to a nondividing state during heterocyst differentiation in Anabaena sp. AB - Nitrogen-deprived filaments of wild-type or hetC Anabaena sp. produce respectively, at semiregular intervals, heterocysts and weakly fluorescent cells. Unlike heterocysts, the latter cells can divide and elongate, producing a pattern of spaced series of small cells. Because a hetR::gfp fusion is expressed most strongly in the small cells, we propose that these small cells represent a very early stage of heterocyst differentiation. hetC::gfp is expressed most strongly in proheterocysts and heterocysts. PMID- 11114942 TI - Some Lactobacillus L-lactate dehydrogenases exhibit comparable catalytic activities for pyruvate and oxaloacetate. AB - The nonallosteric and allosteric L-lactate dehydrogenases of Lactobacillus pentosus and L. casei, respectively, exhibited broad substrate specificities, giving virtually the same maximal reaction velocity and substrate K(m) values for pyruvate and oxaloacetate. Replacement of Pro101 with Asn reduced the activity of the L. pentosus enzyme toward these alternative substrates to a greater extent than the activity toward pyruvate. PMID- 11114943 TI - Cloning, expression, and characterization of cis-polyprenyl diphosphate synthase from the thermoacidophilic archaeon Sulfolobus acidocaldarius. AB - cis-polyprenyl diphosphate synthases are involved in the biosynthesis of the glycosyl carrier lipid in most organisms. However, only little is known about this enzyme of archaea. In this report, we isolated the gene of cis-polyprenyl diphosphate synthase from a thermoacidophilic archaeon, Sulfolobus acidocaldarius, and characterized the recombinant enzyme. PMID- 11114944 TI - areCBA is an operon in Acinetobacter sp. strain ADP1 and Is controlled by AreR, a sigma(54)-dependent regulator. AB - The areCBA genes in Acinetobacter sp. strain ADP1, determining growth on benzyl alkanoates, are shown to be transcribed as a single operon and regulated by areR, which encodes a regulatory protein of the NtrC/XylR family. Assays of the Are enzymes and of two insertions of lacZ as a reporter gene have shown that the operon is induced by benzyl acetate, benzyl alcohol, and benzaldehyde, as well as 2- and 4-hydroxybenzyl acetates and benzyl propionate and butyrate. Two adjacent sites of transcriptional initiation were 97 and 96 bp upstream of the start codon for areC, near a sigma(54)-dependent -12, -24 promoter. Inactivation of areR and rpoN (for RNA polymerase sigma(54)) drastically reduced growth rates on the Are substrates and induction of the operon. PMID- 11114945 TI - No effect of venoconstrictive thigh cuffs on orthostatic hypotension induced by head-down bed rest. AB - Orthostatic intolerance (OI) is the most serious symptom of cardiovascular deconditioning induced by head-down bed rest or weightlessness. Wearing venoconstrictive thigh cuffs is an empirical countermeasure used by Russian cosmonauts to limit the shift of fluid from the lower part of the body to the cardio-cephalic region. Our aim was to determine whether or not thigh cuffs help to prevent orthostatic hypotension induced by head-down bed rest. We studied the effect of thigh cuffs on eight healthy men. The cuffs were worn during the day for 7 days of head-down bed rest. We measured: orthostatic tolerance (stand tests and lower body negative pressure tests), plasma volume (Evans blue dilution), autonomic influences (plasma noradrenaline) and baroreflex sensitivity (spontaneous baroreflex slope). Thigh cuffs limited the loss of plasma volume (thigh cuffs: -201 +/- 37 mL vs. control: -345 +/- 42 mL, P < 0.05), the degree of tachycardia and reduction in the spontaneous baroreflex sensitivity induced by head-down bed rest. However, the impact of thigh cuffs was not sufficient to prevent OI (thigh cuffs: 7.0 min of standing time vs. control: 7.1 min). Decrease in absolute plasma volume and in baroreflex sensitivity are known to be important factors in the aetiology of OI induced by head-down bed rest. However, dealing with these factors, using thigh cuffs for example, is not sufficient to prevent OI. Other factors such as venous compliance, microcirculatory changes, peripheral arterial vasoconstriction and vestibular afferents must also be considered. PMID- 11114946 TI - PKCalpha translocation is microtubule-dependent in passaged smooth muscle cells. AB - The translocation of protein kinase C (PKC) isozymes from their inactive cell locus to a variety of cytoskeletal, organelle, and plasmalemmal sites is thought to play an important role in their activation and substrate specificity. We have utilized confocal microscopy to compare phorbol 12, 13 dibutyrate (PDB) - stimulated translocation of PKCalpha in cultured cells derived from rat vascular smooth muscle. In enzymatically dispersed, passaged smooth muscle cells, PKCalpha was uniformly distributed throughout the unstimulated cell. PDB stimulation resulted in extensive association of the PKCalpha into filamentous strands with subsequent accumulation of the isoform in the peri-nuclear region of the cell. Dual immunostaining indicated that PKCalpha was extensively colocalized with microtubules in the interval immediately following PDB stimulation but was largely disassociated from microtubules at 10 min, at which time the translocation of PKCalpha to the peri-nucleus/nucleus was nearly complete. It was further found that the use of colchicine to disrupt the microtubules caused the loss of PKCalpha translocation to the peri-nuclear region. By comparison, cytochalasin B disruption of actin microfilaments had no significant effect on this parameter. The data suggest that PDB stimulation results in a transient association of PKCalpha with cell microtubules and that the microtubules play an important role in the translocation of PKCalpha from the cytosol in passaged cells derived from rat aortic smooth muscle. PMID- 11114947 TI - Purinergic activation of BK channels in clonal kidney cells (Vero cells). AB - We have studied the activation of a high-conductance channel in clonal kidney cells from African green monkey (Vero cells) using patch-clamp recordings and microfluorometric (fura-2) measurements of cytosolic Ca2+. The single-channel conductance in excised patches is 170 pS in symmetrical 140 mM KCl. The channel is highly selective for K+ and activated by membrane depolarization and application of Ca2+ to the cytoplasmatic side of the patch. The channel is, thus, a large-conductance Ca2+-activated K+ channel (BK channel). Cell-attached recordings revealed that the channel is inactive in unstimulated cells. Extracellular application of less than 0.1 microM ATP transiently increased the cytosolic Ca2+ concentration ([Ca2+]i) to about 550 nM, and induced membrane hyperpolarization caused by Ca2+-activated K+ currents. ATP stimulation also activated BK channels in cell-attached patches at both the normal-resting potential and during membrane hyperpolarization. The increase in [Ca2+]i was owing to Ca2+ release from internal stores, suggesting that Vero cells express G protein-coupled purinergic receptors (P2Y) mediating IP3-induced release of Ca2+. The P2Y receptors were sensitive to both uracil triphosphate (UTP) and adenosine diphosphate (ADP), and the rank of agonist potency was ATP >> UTP >/= ADP. This result indicates the presence of both P2Y1 and P2Y2 receptors or a receptor subtype with untypical agonist sensitivity. It has previously been shown that hypotonic challenge activates BK channels in both normal and clonal kidney cells. The subsequent loss of KCl may be an important factor in cellular volume regulation. Our results support the idea of an autocrine role of ATP in this process. A minute release of ATP induced by hypotonically evoked membrane stretch may activate the P2Y receptors, subsequently increasing [Ca2+]i and thus causing K+ efflux through BK channels. PMID- 11114948 TI - Exercise-induced sympathetic activation is correlated with cerebral hemisphere laterality, but not handedness. AB - To investigate whether sympathetic responses are correlated with central laterality or handedness, muscle sympathetic nerve activity (MSNA), heart rate (HR) and blood pressure (BP) were compared between right (RA) and left arm (LA) grip exercise with volitional maximum effort (MVHG) for 2 min and post-exercise arterial occlusion (PEAO) in right- and left-handed volunteers. MVHG and PEAO led to a greater increase in MSNA in RA than in LA exercise (180 vs. 150%, P=0.004; 140 vs. 85%, P=0.005). MVHG elevated HR to a significantly lesser extent in RA than in LA (35 vs. 46%, P=0.030), and the difference was maintained during PEAO. The BP rise during MVHG and PEAO was the same in RA and in LA. Muscle sympathetic nerve activity, HR and BP responses during MVHG and PEAO showed no difference between the dominant and non-dominant arm. These results suggested that the effects of central motor command and metaboreflex on sympathetic outflow to the vasculature and the heart may be selectively modulated partly by hemispherical laterality. PMID- 11114949 TI - Intramuscular pressure and electromyographic responses of the vastus lateralis muscle during repeated maximal isokinetic knee extensions. AB - The purpose of the study was to investigate changes in intramuscular pressure (IMP) (maximal during contraction - peak IMP, and between contraction, relaxation IMP - RxIMP) and surface electromyographic activity (EMG) parameters [mean frequency of the power spectrum (fmean), and signal amplitude, root mean square (RMS)] throughout 100 repetitive isokinetic contractions for six healthy subjects. Parameters were recorded simultaneously from the vastus lateralis muscle during maximal knee extension. Regression analyses revealed significant decreases for peak IMP and fmean, and an increase in RxIMP; RMS, however, did not change. All parameters demonstrated trends of change throughout the contractions that were non-linear. Details and inter relations for RxIMP and fmean were highlighted to express intramuscular fluid accumulation and fatigue development, respectively. Individual regression analyses for RxIMP revealed significant positive correlations for all subjects (range of r=0.62 to 0.89). At group level, mean RxIMP increased from 6.0 mmHg for the 1st contraction to 14.0 mmHg for the 100th contraction. For fmean, individual regressions were significantly negative for all subjects (r=-0.75 to -0.89). Fmean decreased from 89.2 Hz for the 1st contraction to 63.3 Hz for the 100th contraction. When the data were delineated between the fatigue (contractions 1-40) and endurance phases (41-100), the slopes of increase for RxIMP, and of decrease for fmean were significantly greater during the fatigue phase. RxIMP throughout the 100 contractions correlated negatively with fmean for each subject (r=-0.54 to -0.78); when delineated, the correlation between parameters was significantly greater for the fatigue as compared with the endurance phase. Relaxation IMP trends are mainly attributed to intramuscular water accumulations during repetitive contractions. In spite of consistent correlations between RxIMP and fmean a causal association could not be established. It may be suggested that a common factor occurring during the fatiguing process governs changes in RxIMP and fmean. PMID- 11114950 TI - In vivo dynamics of human medial gastrocnemius muscle-tendon complex during stretch-shortening cycle exercise. AB - The purpose of this study was to investigate the dynamics of human muscle-tendon complex (MTC) during stretch-shortening cycle exercises through in vivo observation. A total of seven male subjects performed dorsi flexion followed by plantar flexion at two different frequencies, 0.3 Hz (slow) and 1.0 Hz (fast), in a toe-standing position. The fascicle length (LF) of the medial gastrocnemius muscle during the movements was determined using a real-time ultrasonography in vivo. The LF at the switching phase from dorsi to plantar flexion was significantly shorter in the fast exercise (54.4 +/- 5.5 mm) than in the slow one (58.2 +/- 5.4 mm), suggesting that the elongation of tendon structures at that time was significantly greater in the former than in the latter. Furthermore, at the initial stage of plantar flexion during the fast movement, the LF hardly changed with a rapid shortening of tendon structures at that time. The observed relation between MTC length and force showed that the behaviour of tendon structures contributed to 20.2 and 42.5% of the total amount of work completed during plantar flexion phase in the slow and fast movements, respectively. Thus, the present results suggest that tendon structures make the dynamics of MTC more efficient during stretch-shortening cycle exercises by changing their lengths. PMID- 11114951 TI - In vivo assessment of villous microcirculation in the rat small intestine in indomethacin-induced inflammation: role of mast-cell stabilizer Ketotifen. AB - Indomethacin induces an inflammatory reaction in the small intestine in several rat strains. This animal model is widely used to study the implications of intestinal inflammation. Macroscopically, there is evidence that Indomethacin induces a hyperaemic inflammatory reaction in the mucosa, and in our previous studies, we found a significant increase in villous perfusion in the acutely inflamed small intestine. Mast-cell activation was found to take part in inflammatory reactions in several organ systems in various species. However, no data are available about the effect of mast-cell degranulation on the perfusion of the mucosa in Indomethacin-induced intestinal inflammation. Therefore, we used the mast-cell stabilizer Ketotifen to identify if mast-cell activation may contribute to changes in the perfusion of single villi in the rat ileum. We found that Ketotifen significantly reduced the Indomethacin induced increased blood flow in the main arteriole 60 min after i.v. application, indicating that mast cell degranulation effects microcirculatory disturbances in the mucosa in Indomethacin-induced intestinal inflammation. PMID- 11114952 TI - Seasonal changes in heart rate and food intake in reindeer (Rangifer tarandus tarandus). AB - This study tested the hypothesis that the annual cycle in heart rate (HR) in reindeer is, at least in part, a consequence of seasonal fluctuation in voluntary food intake. Heart rate and daily dry matter voluntary-food intake (DDMVFI) were recorded in two captive female reindeer (Rangifer tarandus tarandus) from April 1995 to August 1996. Heart rate was measured continuously in each animal for 20 24 h for 7 days each month using Polar(R) Sport Testers (PST); DDMVFI was measured in each animal daily for 17 months. Modal daily heart rate (MDHR) and DDMVFI fluctuated seasonally in close synchrony, both reaching maxima in July and minima in January. The relationship between HR and DDMVFI was investigated experimentally by manipulating the level of feeding in a stepwise manner in May, when appetite was low and in August, when DDMVFI was close to maximum. Heart rate showed stepwise changes in close synchrony with the changes in levels of feeding. These results suggest that the seasonal increase in HR in summer is a consequence of increased food intake and, likewise, decreased HR in winter is a consequence of reduced food intake. The observed relationship between food intake and HR presumably reflects changes in cardiac output and/or the rate of flow of blood to the gastrointestinal tract which are influenced by meal size. PMID- 11114953 TI - Renal tubule potassium channels: function, regulation and structure. PMID- 11114954 TI - Postprandial hypertriglyceridemia(s): time to enlarge our pathophysiologic perspective. PMID- 11114955 TI - Structural myocardial changes after coronary artery surgery. AB - BACKGROUND: Postoperative contractile dysfunction or 'myocardial stunning' has been described after coronary artery bypass grafting (CABG). In the present study we sought to determine if and to what extent clinical, structural and histochemical evidence of myocardial changes associated with stunning could be found in patients after CABG and cold crystalloid cardioplegia. MATERIALS AND METHODS: Left ventricular (LV) biopsies were obtained from CABG patients (n = 10) prior to and at the end of cardiopulmonary bypass (CPB). These biopsies were immunostained for the inducible heat-shock protein 70 (HSP-70i), intercellular adhesion molecule-1 (ICAM-1) and actin. ATP was measured by bioluminescence. RESULTS: Biopsies pre-CPB showed no evidence of myocardial damage as HSP-70i was absent and a regular actin cross-striation pattern and only constitutive ICAM-1 expression were present. After CPB we found significantly increased HSP-70i and ICAM-1 levels as well as a deranged actin cross-striation pattern with a widening of actin bands. ATP levels declined from 10 mmol L-1 pre-CPB to 4.9 mmol L-1 after CPB. Correspondingly, coronary sinus effluent showed a significant lactate production. Although, cardiac function determined by transoesophageal echocardiography did not deteriorate, significant inotropic support was necessary to maintain cardiac output. CONCLUSIONS: Our results present clinical and structural evidence of 'myocardial stunning' after CABG and cold crystalloid cardioplegia. Increased HSP-70i and ICAM-1 expression, as well as a deranged actin cross-striation pattern, might be structural markers to determine 'myocardial stunning' in clinical settings. PMID- 11114956 TI - Apolipoproteins A-I, A-II and B, lipoprotein(a) and the risk of ischaemic heart disease: the Caerphilly study. AB - Apolipoproteins B, A-I and Lp(a) have been proposed as independent predictors of subsequent ischaemic heart disease (IHD) improving on the prediction obtained by routine lipid measurements. In this report we have investigated the relative predictive ability of apolipoproteins and plasma lipids in a prospective study of middle aged men. 2398 men aged 49-65 years from the general population of Caerphilly, South Wales, UK were screened for evidence of IHD. After an overnight fast 2225 men each provided a venous blood sample on which plasma lipids, apolipoproteins B, A-I, A-II, and lipoprotein (a) (Lp(a)) were measured. Over a follow-up period of nearly 9 years, 282 (12%) men developed major IHD. Multiple logistic regression analysis showed that after adjusting for standard cardiovascular risk factors other than lipids there was a strong trend (standardised relative odds (SRO) = 1.20; P = 0.009) for incidence of IHD to increase with apolipoprotein B. However, on further adjusting for total cholesterol this trend largely disappeared (SRO = 1.05; P = 0.57). Similarly, a trend for incidence of IHD to increase with decreasing apolipoprotein A-I (SRO = 1.18; P = 0.02) disappeared when HDL cholesterol was added to the model. Levels of apolipoprotein A-II were not related to risk of subsequent IHD. Incidence of IHD was effectively constant over nearly 90% of the range of Lp(a). Only among the 5% of men with Lp(a) greater than 70 mg dL-1 was the risk of IHD significantly (P = 0.04) greater than among men with Lp(a) less than 10 mg dL-1. Apolipoproteins B and A-I do not improve on the prediction of risk of IHD provided by total and HDL cholesterol, respectively. Apolipoprotein A-II was not related to risk of IHD. Lp(a) may be independently associated with incident IHD among the 5-10% of men with the highest levels. PMID- 11114957 TI - Nitric oxide inhibition intensifies the depressant effect of cocaine on the left ventricular function in anaesthetized pigs. AB - Myocardial ischaemia and left ventricular dysfunction have been described in cocaine users. Whether nitric oxide (NO) inhibition may potentiate the effects of cocaine on coronary circulation and ventricular function is still unknown. In order to test this hypothesis, 38 pentobarbital-anaesthetized pigs were instrumented for systolic blood pressure, coronary blood flow, left ventricular dp/dt, cardiac output, left ventricular end-diastolic and end-systolic lengths and shortening fraction. The pigs were randomized into three groups: control group: i.v. saline (n = 5); group 1: i.v. cocaine, 10 mg kg-1 over 20 min (n = 17); group 2: the same doses of cocaine 30 min after i.c. L-NAME 20 microg/kg min 1 infusion (n = 16). In order to know whether the observed effects were specific of NO inhibition, in five pigs i.c. L-arginine was simultaneously infused with L NAME, in five pigs i.c. NTG, an endothelial-independent vasodilator, was simultaneously infused with L-NAME before cocaine was administered, and in nine additional pigs the proximal left anterior descending (LAD) flow was reduced to around 20% of the basal value by means of a mechanical occluder before cocaine was administered. Cocaine i.v did not change the coronary blood flow, while it induced a significant reduction in cardiac output, left ventricular dp/dt and shortening fraction (15 +/- 4-8 +/- 4%, P < 0.05). When cocaine was administered after L-NAME infused i.c. during 30 min, a significantly more severe reduction of the shortening fraction (12 +/- 3-4 +/- 2%, P < 0.0001) was induced; this effect was abolished by simultaneous perfusion of L-arginine i.c. NTG. The results when cocaine was administered after the 20% LAD flow reduction by mechanical occluder did not differ from those of cocaine alone. NO inhibition intensifies the cocaine induced left ventricular dysfunction. PMID- 11114958 TI - Diurnal triglyceride profiles in healthy normolipidemic male subjects are associated to insulin sensitivity, body composition and diet. AB - BACKGROUND: Elevated fasting and postprandial triglycerides (TG) are established risk factors for Coronary Heart Disease (CHD). Usually, fasting plasma TG are measured, although TG are mainly produced in a postprandial state. Our objective was to investigate diurnal TG profiles using serial capillary TG measurements, in normolipidemic healthy males. MATERIALS AND METHODS: Forty-eight, non-obese, non smoking males (range: 20-55 years, mean age: 32 +/- 12 years), measured diurnal capillary TG, at six fixed timepoints during the day on three different days and recorded their food intake. Insulin sensitivity was estimated by HOMA. Diurnal capillary TG profiles were calculated as integrated area under the mean capillary TG curve (TGc-AUC). RESULTS: All subjects had normal fasting plasma TG and cholesterol. The average TGc-AUC was 23.6 +/- 6.7 mmol h L-1. Significant correlations with TGc-AUC were: fasting insulin (r = 0. 40, P < 0.005), HOMA (r = 0.32, P < 0.05), relative fat mass (r = 0. 31, P < 0.05), dietary protein-(r = 0.31, P < 0.05) and saturated fat intake (r = 0.30, P < 0.05). Age was not associated to diurnal triglyceridemia. After subdividing the group into quartiles on the base of TGc-AUC, differences were found between the highest (n = 12) and lowest quartile (n = 12) for: fasting capillary TG, fasting insulin, HOMA and systolic blood pressure. Fasting plasma TG and dietary intake were not different. CONCLUSION: Diurnal TG profiles in healthy normolipidemic males are not age dependent, but are associated to insulin sensitivity, fat mass and diet. Diurnal capillary TG profiles may be a valuable additional tool in estimating a risk profile for CHD since significant differences in diurnal TG are not always reflected by elevated fasting plasma TG. PMID- 11114959 TI - Smoking or its cessation does not alter the susceptibility to in vitro LDL oxidation. AB - BACKGROUND: Enhanced induction of low density lipoprotein (LDL) oxidation may play a role in the increased cardiovascular risk in smokers. We determined LDL oxidisability in vitro in non-smokers, smokers and in subjects after smoking cessation. PATIENTS AND METHODS: Plasma lipids and copper induced LDL oxidation in vitro were measured in 31 persistent smokers, 47 smokers who tried to stop smoking and 25 non-smokers. In the smoking cessation group, blood was collected before then 1, 3, 6 and 12 months after smoking cessation, and in the persistent smoking and non-smoking groups at baseline and after 12 months. Plasma thiobarbituric acid reactive substances (TBARS) were measured 3 times (at baseline then after 1 and 3 months) in all subjects who refrained from smoking (controlled by urinary cotinine concentrations) for at least 3 months. RESULTS: At baseline, no differences in mean age, body mass index and lipid profiles between groups were present. Seventeen subjects of the smoking cessation group (36%) managed to quit during 12 months. Smoking cessation was associated with an increase in mean weight (P 90 and < 100 mmHg; plasma LDL-cholesterol > 3.9 and < 6.5 mmol L 1) were enrolled in the study. In random order, these patients received Simvastatin (20 mg day-1) or Cholestyramine (6 g three times a day) for a period of 10 months and after three months of wash-out (cross-over) the sequence was reversed for an additional 10 months. Blood pressure, lipid parameters, glycated haemoglobin and urinary albumin excretion were measured during the study. Additionally, in eight patients, urinary glycosaminoglycan excretion (GAG) was also measured during the study. RESULTS: Simvastatin and Cholestyramine were equally effective in reducing total and LDL cholesterol. Only during Simvastatin treatment a significant reduction in diastolic blood pressure and both 24 h urinary albumin and GAG excretion rates were observed, while no significant changes were seen with Cholestyramine treatment. CONCLUSIONS: Our results clearly show for the first time that the reduction of blood pressure, together with 24 h urinary albumin excretion rate - two established cardiovascular risk factors, obtained during Simvastatin therapy in hypertensive type 2 diabetic patients - is in large part independent from the reduction of LDL Cholesterol. PMID- 11114961 TI - Effect of rectal distension on gallbladder emptying and circulating gut hormones. AB - BACKGROUND: Abnormalities of upper gut motility, including a delay of gastric emptying and small bowel transit, found in patients with constipation may be secondary to factors originating in the colon or rectum as a result of faecal stasis. The aim was to determine if stimulation of mechanosensory function by rectal distension affects postprandial gallbladder emptying and release of gastrointestinal peptides participating in control of upper gut motility. MATERIALS AND METHODS: Eight healthy volunteers were studied with an electronic barostat and a plastic bag positioned in the rectum. Intrabag pressure was maintained at minimal distension pressure + 2 mmHg on one occasion and on a pressure that induced a sensation of urge on the other. Gallbladder volume and plasma concentrations of cholecystokinin (CCK), pancreatic polypeptide (PP) and peptide YY (PYY) were measured before and after ingestion of a 450-kcal mixed liquid meal. RESULTS: Rectal distension enhanced maximum gallbladder emptying from 66 +/- 7% to 78 +/- 5% (P < 0.05). Distension tended to increase integrated plasma PYY from 77 +/- 30 pM min to 128 +/- 40 pM min in the first hour after the meal (P = 0.08) and it suppressed integrated plasma PP from 1133 +/- 248 pM min to 269 +/- 284 pM min in the second hour (P < 0.05). Integrated plasma CCK concentrations were not significantly affected. CONCLUSION: Mechanosensory stimulation of the rectum enhances postprandial gallbladder emptying and influences postprandial release of gut hormones involved in the regulation of gastrointestinal motility in healthy subjects. These mechanisms may play a role in the pathogenesis of the upper gastrointestinal motor abnormalities observed in constipated patients. PMID- 11114962 TI - Cytokine secretion is impaired in women with diabetes mellitus. AB - BACKGROUND: As women with diabetes mellitus (DM) have an increased prevalence of asymptomatic bacteriuria (ASB) and it is known that a correlation exists between the increased prevalence of genitourinary tract infection and impaired cytokine production in women infected with Human Immunodeficiency Virus (HIV), we studied urinary cytokine excretion in diabetic women and compared it with that of nondiabetic controls. MATERIALS AND METHODS: To evaluate the cytokine secretion capacity of women with DM, both whole blood and isolated monocytes of women with and without DM were stimulated in vitro with lipopolysaccharide (LPS). RESULTS: Lower urinary interleukin-8 (IL-8) and interleukin-6 (IL-6) concentrations (P = 0.1 and P < 0.001, respectively) were found in diabetic women than in nondiabetic controls. A lower urinary leukocyte cell count correlated with lower urinary IL-8 and IL-6 concentrations (P < 0.05). Lower tumour necrosis factor-alpha (TNF alpha) and IL-6, but comparable interleukin-10 (IL-10) concentrations were found in whole blood (P < 0.04) and isolated monocytes (P = 0.03) of women with DM type 1 compared to women without DM. CONCLUSIONS: Diabetic women with ASB have lower urinary IL-6 concentrations than nondiabetic bacteriuric controls. In addition, monocytes of women with DM type 1 secrete lower pro-inflammatory cytokines after stimulation with LPS than monocytes of women without DM. This is not due to an inhibitory effect of the anti-inflammatory cytokine IL-10. This can have important consequences for both host defense, endothelial cell functioning and atherogenesis. PMID- 11114963 TI - Ibuprofen inhibits adhesiveness of monocytes to endothelium and reduces cellular oxidative stress in smokers and non-smokers. AB - BACKGROUND: Cigarette smoking is a major risk factor in atherosclerosis and a useful model from which to study chronic inflammation. We compared monocyte function, lipid profiles and inflammatory markers in smokers and non-smokers, before and after oral ibuprofen intake. The adhesion of freshly isolated monocytes to native and tumour necrosis factor alpha (TNFalpha) stimulated human umbilical vein endothelial cells (HUVEC), as well as superoxide anion (O2-) levels and hydrogen peroxide (H2O2) production in resting and phorbol myristate acetate (PMA) stimulated monocytes were determined. MATERIALS AND METHODS: A group of nine smokers without any other coronary risk factor was compared with an age-matched group of 9 non-smokers. Tests were performed before and after a two week course of oral ibuprofen (600 mg day-1). RESULTS: In smokers before ibuprofen, monocyte adhesion to native and TNFalpha-stimulated HUVEC was increased (P < 0001 and P < 0.01, respectively), and so were O2- levels in native and PMA-stimulated monocytes (P < 0.01 and P < 0.001, respectively). Ibuprofen reduced the adhesion of monocytes to native and stimulated HUVEC (P < 0.001) and O2- generation by resting and PMA-stimulated cells (P < 0.01) in both groups. H2O2 production by resting and PMA-stimulated monocytes was reduced in smokers and non-smokers (P < 0.01). Interestingly, ibuprofen increased HDL cholesterol levels in smokers (P < 0.01) and non-smokers (P < 0.001), and reduced the level of triglycerides in smokers (P < 0.05). CONCLUSION: Oral administration of ibuprofen reduced the adhesion of monocytes to HUVEC, suppressed oxidative stress and increased HDL cholesterol levels in smokers and non-smokers. PMID- 11114964 TI - Phospholipid composition in late infantile neuronal ceroid lipofuscinosis. AB - BACKGROUND: Neuronal ceroid lipofuscinosis (NCL) is a relatively common group of inherited neurodegenerative disorders characterised by the accumulation of autofluorescent lipopigments (ceroid) similar to lipofuscin. Because of this property, studies have concentrated on fatty acid metabolism and lipid peroxidation. METHODS: In the present study, the fatty acid composition of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) and the molecular species compositions of diacylglycerophosphocholine (diacyl GPC), diacylglycerophosphoethanolamine (diacyl GPE) and alkenylacyl GPE (plasmalogens) were investigated in cultured skin fibroblasts from three patients with a confirmed diagnosis of the late infantile form of the disease (LINCL, CLN2) and three healthy age-matched controls. RESULTS: Relatively minor differences in the fatty acid compositions of PC and PE were observed between patients and controls. However, dimethyl acetals of plasmalogens were found to be 40% higher in the patients compared to in the controls. Control and LINCL fibroblasts displayed only slight differences in the molecular compositions of diacyl GPE and diacyl GPC. In contrast, compared with normal cells, LINCL fibroblasts had higher levels of alkenylacyl GPE species containing both 18 : 1 and polyunsaturated fatty acids, but lower levels of species with 16 : 0 or 18 : 0 in the sn-1 position. CONCLUSION: The molecular composition of PC and PE subclasses in skin fibroblasts of healthy subjects and patients suffering from LINCL is here described for the first time. While few differences are noticeable in the fatty acid composition of PC and PE and the molecular species distribution of diacylGPC and diacylGPE, the alkenylacyl GPE (or ethanolamine plasmalogens) were found to differ significantly between patients and healthy controls. PMID- 11114965 TI - alpha-tocopherol improves impaired physiology of rat type II pneumocytes isolated from experimentally injured lungs. AB - BACKGROUND: Oxidant stress delivered by nitrogen dioxide (NO2) inhalation impairs the function of extracellular surfactant as well as surfactant phospholipid metabolism in type II pneumocytes. Because protection against oxidant stress is important to normal lung function, the lung contains a variety of antioxidants, including vitamin E. Whether administration of this antioxidant during NO2 inhalation attenuates NO2-induced alterations in phospholipid metabolism in type II pneumocytes has not been studied. METHODS: We exposed rats to identical NO2 body doses (720 p.p.m. x h) using continuous, intermittent, or repetitive protocols. During exposure periods, the animals received daily intramuscular injections of vitamin E (25 mg kg-1). We isolated type II pneumocytes from NO2 exposed rats and evaluated them for cell yield and viability, as well as for synthesis and secretion of phosphatidylcholine (PC) as measures of surfactant metabolism. RESULTS: The yield of type II pneumocytes was significantly elevated from animals that had been exposed continuously to NO2 whereas in intermittently and repeatedly exposed rats, cell yield was similar to yield from control animals. Viability of the isolated cells was similar in controls and all NO2 exposure protocols. Vitamin E treatment of the NO2-exposed rats neither changed cell yield nor cell viability. Phospholipid de novo synthesis, as estimated by choline incorporation into PC, was increased most after continuous NO2 inhalation whereas in the other conditions there was only a slight increase. Vitamin E administration further increased phospholipid synthesis; this difference reached statistical significance only in the case of intermittent NO2 exposure. Secretion of phosphatidylcholine from type II cells was only reduced after continuous NO2 inhalation and administration of the antioxidant reduced the impairment. CONCLUSION: Because vitamin E appears to preserve the ability of type II pneumocytes isolated from NO2-exposed rats to synthesize and secrete surfactant lipid, we conclude that administration of vitamin E may mitigate NO2-induced lung injury. PMID- 11114966 TI - Trichostrongylus colubriformis extract upregulates TNF-alpha receptor expression and enhances TNF-alpha sensitivity of L929 cells. AB - Many pathogens have developed strategies to avoid the host's immune system and hence improve their long-term survival. These strategies include antigenic variation, mimicry of host regulatory proteins and production of immunoregulatory molecules. The ruminant gastrointestinal nematode Trichostrongylus colubriformis produces several factors with homology to human immunoregulatory proteins. However, direct immunomodulation by T. colubriformis proteins has not yet been unequivocally demonstrated. Results in the present paper demonstrate that soluble T. colubriformis factors promote proliferation of the TNF-susceptible mouse fibrosarcoma cell line L929, while inhibiting proliferation of all other cell types tested. In addition, T. colubriformis homogenate enhanced the susceptibility of L929 cells to the cytotoxic action of ovine TNF-alpha. Within 1 h of exposure, T. colubriformis factors bind L929 cells in a stable fashion, yet it takes up to 24 h for the cells to become sensitised to TNF-alpha. Interestingly, the increase of both TNF-alpha sensitivity and proliferation of treated L929 cells correlated with an upregulation in expression of TNF-alpha p55 and p75 receptors. PMID- 11114967 TI - Computer-assisted analysis of molecular mimicry between human papillomavirus 16 E7 oncoprotein and human protein sequences. AB - The immunology of human papillomavirus (HPV) infections has peculiar characteristics. The long latency for cervical cancer development after primary viral infection suggests mechanisms that may aid the virus in avoiding the host immunosurveillance and establishing persistent infections. In order to understand whether molecular mimicry phenomena might explain the ability of HPV to avoid a protective immune response by the host cell, sequence similarity between HPV16 E7 oncoprotein and human self-proteins was examined by computer-assisted analysis. Data were obtained showing that the HPV16 E7 protein has high and widespread similarity to several human proteins involved in a number of critical regulatory processes. In addition, multiple identical and different E7 peptide motifs are present in the same human protein. Thus, sharing of common motifs between viral oncoproteins and molecules of normal cells may be one cause underlying the scarce immunogenicity of HPV infections. The hypothesis is advanced that synthetic peptides harbouring viral motifs not and/or scarcely represented in the host's cellular proteins may represent a valuable immunotherapeutic approach for cervical cancer treatment. PMID- 11114968 TI - HIV-1-specific cellular immune responses among HIV-1-resistant sex workers. AB - The goal of the present study was to determine whether there were HIV-1 specific cellular immune responses among a subgroup of women within a cohort of Nairobi prostitutes (n = 1800) who, despite their intense sexual exposure to HIV-1, are epidemiologically resistant to HIV-1 infection. Of the 80 women defined to be resistant, 24 were recruited for immunological evaluation. The HIV-1-specific T helper responses were determined by IL-2 production following stimulation with HIV-1 envelope peptides and soluble gp120. Cytotoxic T lymphocyte responses were determined by lysis of autologous EBV-transformed B cell lines infected with control vaccinia virus or recombinant vaccinia viruses containing the HIV-1 structural genes env, gag and pol. Resistant women had significantly increased HIV-1 specific T-helper responses, as determined by in vitro IL-2 production to HIV-1 envelope peptides and soluble glycoprotein 120, compared with low-risk seronegative and HIV-1-infected controls (P < or = 0.01, Student's t-test). Seven of the 17 (41%) resistant women showed IL-2 stimulation indices > or = 2.0. HIV-1 specific CTL responses were detected among 15/22 (68.2%) resistant women compared with 0/12 low-risk controls (Chi-squared test, P < 0.001). In the two resistant individuals tested, the CTL activity was mediated by CD8+ effectors. Many HIV-1 resistant women show evidence of HIV-1-specific T-helper and cytotoxic responses. These data support the suggestion that HIV-1-specific T-cell responses contribute to protection against HIV-1 infection. PMID- 11114969 TI - Dysregulated expression of CD69 and IL-2 receptor alpha and beta chains on CD8+ T lymphocytes in flaky skin mice. AB - T-cell activation is considered to be an important element in the pathogenesis of psoriasis, a human skin disease characterized by keratinocyte hyperproliferation, altered keratinocyte differentiation and inflammation of the dermis and epidermis. Mice homozygous for the flaky skin (fsn) mutation develop a skin disorder that has histopathological and biochemical features resembling some forms of psoriasis. It has been reported recently that peripheral lymph nodes (PLN) in fsn/fsn mice exhibit various abnormalities in T-cell development suggestive of dysregulated T- and B-cell activation. In the present study, the expression of the inducible T-cell activation antigens CD69 and IL-2 receptor alpha chain (CD25) on PLN cells from fsn/fsn mice and their phenotypically normal littermates is examined. Expression of CD69 was significantly increased on PLN cells in fsn/fsn mice (mean +/- SD, 49.9 +/- 14.7% of cells) compared with control mice (14.6 +/- 4.2%). Analysis of CD4+ and CD8+ T cell subsets revealed that expression of CD69 in fsn/fsn PLN was significantly biased toward CD8+ cells. Although expression of CD25 was preferentially associated with CD4+ rather than CD8+ cells in both fsn/fsn and control PLN, with most CD4+ CD25+ cells being CD25hi, the proportion of CD4+ cells expressing CD25 was higher in fsn/fsn than control PLN. In contrast, CD25 was expressed by 2-3% of CD8+ PLN cells in both fsn/fsn and control mice and CD25hi cells accounted for < 1% of CD8+ cells in fsn/fsn PLN. The paucity of CD25 on CD8+ cells in fsn/fsn PLN did not appear to be due to a defect in the ability of these cells to upregulate CD25, because T cell receptor stimulation in vitro induced high expression of CD25 on both CD4+ and CD8+ cells. A striking and consistent finding was that most CD8+ cells in fsn/fsn PLN expressed high levels of IL-2R beta chain (CD122). In contrast, CD122 was expressed at low levels on CD8+ cells in control mice. Analysis of PLN cells from newborn fsn/fsn mice revealed that the high expression of CD122 on CD8+ cells was established by 2 weeks of age, prior to the appearance of clinical skin disease. These data indicate that large numbers of T cells in fsn/fsn mice are activated and reinforce the view that fsn is an important regulator of lymphocyte development and function. The relationship between T-cell activation and flaky skin disease in these mice remains to be established. PMID- 11114970 TI - Production of a recombinant form of early pregnancy factor that can prolong allogeneic skin graft survival time in rats. AB - Early pregnancy factor (EPF), an extracellular chaperonin 10 homologue, has immunosuppressive and growth factor properties. In order to carry out more extensive studies on the in vivo characteristics of EPF, a recombinant form of the molecule has been prepared. Recombinant human EPF (rEPF) was expressed in Escherichia coli using the plasmid pGEX-2T expression system. Potency of rEPF in vitro in the rosette inhibition test, the bioassay for EPF, was equivalent to that of native EPF (nEPF), purified from human platelets, and synthetic EPF (sEPF). However, the half-life of activity (50% decrease in the log value) in serum, following i.p. injection, was significantly decreased (3.2 h, compared with nEPF 6.2 days, sEPF 5.8 days). This was thought to be due to modification of the N-terminus of the recombinant molecule inhibiting binding to serum carrier proteins. Because EPF can modify Th1 responses, the ability of the recombinant molecule to suppress allogeneic graft rejection was investigated. Following skin grafts from Lewis rats to DA rats and vice versa, rEPF was delivered locally at the graft site and the effect on survival time of the allografts noted. Results demonstrated that rEPF treatment significantly prolonged skin graft survival time by as much as 55% in stringent models of transplantation across major histocompatibility barriers. PMID- 11114971 TI - Human peripheral blood lymphocyte severe combined immunodeficiency (hu-PBL SCID) models of toxoplasmosis. AB - Toxoplasmosis is a potentially fatal opportunistic infection of immunocompromised hosts. Improved animal models of toxoplasmosis are needed to more nearly approximate conditions that occur in immunocompromised humans. The development of models of toxoplasmosis using human peripheral blood lymphocytes (hu-PBL) transplanted into severe combined immunodeficiency (SCID) mice is described here. Transplantation of hu-PBL into SCID mice without prior conditioning of the mice resulted in detectable differences in quantitative histological scores of brain inflammation due to Toxoplasma gondii infection, but did not alter mortality when compared to SCID mouse controls. The lack of detectable differences in survival were due to inadequate engraftment of hu-PBL, as assessed by flow cytometry. Unconditioned hu-PBL SCID mice had low titre T. gondii-specific antibody detectable after infection. When pretransplantation conditioning with irradiation and antiasialo GM 1 (n-glucolyl neuraminic acid) antibody was used, prolonged hu PBL engraftment was observed in SCID mice, which was associated with worsened histopathology and usually impaired survival when compared with SCID mouse controls. When pretransplantation conditioning with irradiation, antiasialo GM antibody and polyethylene glycol-conjugated IL-2 was used, prolonged hu-PBL engraftment was also documented, but this did not affect survival from T. gondii infection when compared with similarly conditioned SCID mouse controls. The latter conditioning protocol resulted in hu-PBL SCID mice producing high titre T. gondii-specific antibody after infection. Conditioned hu-PBL SCID mice had evidence of increased T. gondii-induced inflammatory scores when compared with conditioned SCID mice. These models show promise for the study of the pathogenesis of toxoplasmosis and conditioned hu-PBL SCID mice may have applications for the evaluation of novel therapies for toxoplasmosis in immunocompromised humans. PMID- 11114972 TI - One year in Antarctica: mucosal immunity at three Australian stations. AB - The effect of a year's isolation in Antarctica on the human mucosal immune system was assessed during the winter of 1992 at three Australian Antarctic stations: Casey, Davis and Mawson. Saliva samples were collected from each expeditioner prior to their departure from Australia and during each month in Antarctica. The concentrations of salivary immunoglobulins IgA and IgG were significantly different between the three stations, but there were no differences for salivary IgM and albumin. The mean concentrations of IgA were higher at Mawson (P < 0.008), and the mean concentrations of IgG were lower at Davis (P < 0.001) compared with the other stations. Ranges of values observed at the stations over the 12-13 months were similar. The variability of values within individuals showed station differences for salivary IgM and IgG only. The study revealed significant changes in salivary immunoglobulin values over the period in Antarctica, with similar patterns at the three Australian stations. The salivary IgA and IgM levels were lower in the first 4 months in Antarctica (January-April) and increased to maximum values in July-August, before returning to mean levels when isolation was broken in October-November. The patterns of salivary IgA and IgM suggest that stressors due to isolation may play a role in alterations of mucosal immunity in expeditioners in Antarctica. PMID- 11114973 TI - Altered superantigenic ligands demonstrate the quantitative nature of T-cell activation. AB - In a recent study, a superantigen mutated in the TCR binding site (staphylococcal enterotoxin B (SEB)delta61Y) was described, which behaved as a partial agonist for a Vbeta17-expressing T-cell clone. Evidence is now presented to demonstrate that there is distinct heterogeneity in the response of primary T cells to this protein. Some Vbeta17 T cells responded to SEBdelta61Y by modulating surface receptor expression consistent with activation, and by proliferating. Other Vbeta17 T cells did not proliferate, nor did they display a receptor expression phenotype consistent with activation. However, when repeatedly exposed to the altered superantigen, some of these non-responders entered cell cycle. This pattern of responses was not recapitulated by providing additional costimulation via CD28, although such treatment did induce some of the 'unresponsive' Vbeta17 T cells to upregulate the IL-2 receptor, indicative of partial activation. It was also found that the heterogeneous pattern could be replicated using very low doses of native SEB. The data are discussed in the context of models of T-cell activation in which differences in TCR ligand affinity and dose determine qualitatively different response phenotypes. PMID- 11114974 TI - Adherence of human monocytes and NK cells to human TNF-alpha-stimulated porcine endothelial cells. AB - Discordant xenograft models undergoing delayed rejection response are characterized by xenograft infiltration with host monocytes and NK cells, associated with the release of large quantities of pro-inflammatory cytokines, such as TNF-alpha. In the present study, human monocytes (PBMo)/NK cells (PBNK) isolated from peripheral blood and cultured porcine aortic endothelial cells (PAEC) treated with recombinant human TNF-alpha (rhTNF-alpha) were used to investigate their adhesive interactions and mAbs against porcine E-selectin, human CD11a and CD49d were used to test their relative contributions to such intercellular adhesions. The PBMo exhibited significantly greater adherence to resting (unstimulated) PAEC than PBNK. The rhTNF-alpha upregulated E-selectin and vascular cell adhesion molecule-1 (VCAM-1) expression on PAEC and augmented the adhesiveness of PAEC for PBMo and PBNK in a time- and dose-dependent manner. In mAb blocking assays, anti-E-selectin, anti-CD11a and anti-CD49d mAbs did not inhibit PBMo adherence to rhTNF-alpha-stimulated PAEC when used singly, but resulted in a maximal inhibitory effect when used in combination. Regarding PBNK, anti-E-selectin mAb had no marked influence on PBNK adherence. The combined use of anti-CD11a and anti-CD49d mAbs produced additive reduction in the PBNK binding to rhTNF-alpha-stimulated PAEC, even to far below baseline (unstimulated) levels. Therefore, it is concluded that human TNF-alpha promotes the adhesiveness of PAEC for human monocytes and NK cells and that the mechanism underlying the increased adherence differs for PBMo and PBNK. PMID- 11114975 TI - Prevention of a chronic progressive form of experimental autoimmune encephalomyelitis by an antibody against mucosal addressin cell adhesion molecule 1, given early in the course of disease progression. AB - A role for alpha4 and beta7 integrins in mediating leucocyte entry into the central nervous system in the multiple sclerosis (MS)-like disease experimental autoimmune encephalomyelitis (EAE) has been demonstrated. However, the individual contributions of their respective ligands mucosal addressin cell adhesion molecule-1 (MAdCAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-cadherin expressed on the blood-brain barrier has not been determined. In the present paper, it is shown that an antibody directed against MAdCAM-1, the preferential ligand for alpha4beta7, effectively prevented the development of a progressive, non-remitting, form of EAE, actively induced by injection of myelin oligodendrocyte glycoprotein peptide (MOG(35-55)) autoantigen. Combinational treatment with both anti-MAdCAM-1, VCAM-1, and intercellular adhesion molecule-1 (ICAM-1) (ligand for integrin lymphocyte function-associated antigen (LFA)-1) mAbs led to more rapid remission than that obtained with anti-MAdCAM-1 antibody alone. However, neither MAdCAM-1 monotherapy, nor combinational antibody blockade was preventative when administered late in the course of disease progression. In conclusion, MAdCAM-1 plays a major contributory role in the progression of chronic EAE and is a potential therapeutic target for the treatment of MS. Critically, antivascular addressin therapy must be given early in the course of disease prior to the establishment of irreversible damage if it is to be effective, as a single treatment modality. PMID- 11114977 TI - Present and future therapeutic strategies for idiopathic oligozoospermia. AB - The effectiveness of medical treatment for idiopathic oligozoospermia (IO) has been at best doubtful until now and a logical consequence of this unsatisfactory situation has been the partial displacement of this approach by assisted reproduction techniques. This state of affairs has resulted from insufficient investigation, inappropriately designed clinical trials and consistent disregard for the principles of evidence-based medicine. Protocol-related shortcomings and wrong interpretation of the data available have also been some of the all too frequent problems encountered in this therapeutic approach. In this rather misty situation, it appears that, of the therapeutic agents used so far, follicle stimulating hormone (FSH) (mainly FSH-secretagogues) may exert some beneficial effects on a number of biological endpoints related to spermatogenesis and sperm maturation. The short and medium term prospects of medical treatment for IO rest mainly with improvement of investigative procedures to a higher degree of sophistication, with emphasis placed on identifying the causes rather than the results of dysfunction so that a better selection of candidates can be made. Moreover, the introduction of prognostic indices for evaluation of the beneficial effects of a therapeutic agent may be of paramount importance. Finally, a better assessment of the preparations available and, possibly, the introduction of new more specific agents may also be an important step forward in this field. This type of large-scale effort should not be left to individual investigators or special centres working independently, but it may come under the auspices of a central regulating agency so that undisputed results from large, multicentre and uniform studies might be obtained, if medical treatment is to remain a good option. In this context, it may also be emphasized that andrology's main task should always be to treat the male with the problem rather than his healthy female partner, whenever this is possible. PMID- 11114976 TI - Differential interleukin-6 mRNA expression in Nippostrongylus brasiliensis infection of susceptible and resistant strains of mice. AB - Intestinal parasitic infection is still a major problem in humans and animals, yet host immunity against gut parasitic infection remains partially understood. Eosinophilia and mastocytosis are features of such infection that have been shown to be genetically controlled. The expression of IL-6 is detected in eosinophils, mast cells and neutrophils and may be responsible for the regulation of leucocytes at infective sites. The relationships between IL-6 expression, eosinophilia, mastocytosis and host immunity remain unclear. In the present report, a close correlation between IL-6 mRNA+ cells, eosinophilia, mastocytosis and worm expulsion is demonstrated, which may indicate a role for IL-6 in regulation of host immunity against intestinal parasite infection. PMID- 11114978 TI - Regulatory influence of germ cells on sertoli cell function in the pre-pubertal rat after acute irradiation of the testis. AB - While germ cell regulation of Sertoli cells has been extensively explored in adult rats in vivo, in contrast, very little is known about germ cell influence on Sertoli cell function at the time when spermatogenesis begins and develops. In the present study various Sertoli cell parameters (number, testicular androgen binding protein (ABP) and testin, serum inhibin-B and, indirectly, follicle stimulating hormone (FSH)) were investigated after the exposure of 19-day-old rats to a low dose of 3 Grays of gamma-rays. Differentiated spermatogonia were the primary testicular targets of the gamma-rays, which resulted in progressive maturation depletion, sequentially and reversibly affecting all germ cell classes. Testicular weight declined to a nadir when pachytene spermatocytes and spermatids were depleted from the seminiferous epithelium and complete or near complete recovery of spermatogenesis and testicular weight was observed at the end of the experiment. Blood levels of FSH and ABP were normal during the first 11 days after irradiation, when spermatogonia and early spermatocytes were depleted. While the number of Sertoli cells was not significantly affected by the irradiation, from days 11-66 after gamma-irradiation, ABP production declined and FSH levels increased when pachytene spermatocytes and spermatids were depleted and the recovery of these parameters was only observed when spermatogenesis was fully restored. Comparison of the pattern of change in serum levels of inhibin-B and testicular levels of testin and of germ cell numbers strongly suggest a relationship between the disappearance of spermatocytes and spermatids from the seminiferous epithelium and the decrease in levels of inhibin-B and increase in levels of testin from 7 to 36 days post-irradiation. Levels of testin and inhibin B were restored before spermatogenesis had totally returned to normal. In conclusion, this in vivo study shows that pre-pubertal Sertoli cell function is under the complex control of various germ cell classes. This control presents clear differences when compared with that previously observed in adult animals and depends on the Sertoli cell parameter of interest, as well as on the germ cell type. PMID- 11114979 TI - The advantages of standardized evaluation of male infertility. AB - Healthcare can be improved by standardization and by evaluation of diagnostic methods and treatments. In the field of andrology, in which large patient numbers are required for the evaluation of diagnostic procedures and treatments, structured data collection and multicentre studies are especially warranted. Concomitant with routine clinical practice, a large amount of clinical data are collected that may be used to evaluate andrological care. Structuring and electronic storage of data holds promise in terms of clarity and accessibility of the data and its use for validation studies. The aim of the present work was to study the merits of routine collection of a common dataset in a computer-based patient record (CPR) for standardization, quality of data and clinical research. It was studied whether the data were of sufficient quality and accessibility for much needed studies on aetiology, interventions and diagnostics in andrology. Data collection in a structured CPR promoted complete and comprehensive data. We describe the advantages, pitfalls and solutions with this approach. Data on the uniform examination of 1549 infertile men became readily accessible. Population characteristics, basal associations and original studies were enabled and provided insight into the efficiency of clinical practice. In 66% of men, a cause for their infertility was identified, which provides a better rationale for treatment than semen parameters alone. For more than 30% of the patients, a rational andrological treatment was available, which could be deployed before assisted reproductive technologies were resorted to. However, most treatments have not been properly validated. The thorough diagnostic evaluation identifies subgroups that require an evidence base for treatment and further study on aetiology and diagnosis. Structured collection of uniform patient data through a CPR was feasible and facilitated the evaluation of diagnostic and therapeutic modalities. The reported advantages, pitfalls and solutions with this approach may help other centres to decide on how to implement a CPR. Conscientious collection of a standard data set in infertility centres facilitates pooling of data and evidence-based multicentre research. PMID- 11114980 TI - Torsion-induced injury in rat testes does not affect mitochondrial respiration or the accumulation of mitochondrial mutations. AB - Male rats were subjected to 1 h testicular torsion of the spermatic cord or 1 h torsion followed by detorsion and recovery up to 4 weeks. The extent of tissue damage was evaluated by a testicular biopsy score count and mitochondrial function. Torsion for 1 h followed by detorsion induced significant morphological damage, which became more severe with longer periods of recovery. This morphological damage could not be correlated with mitochondrial damage as assessed by measuring the 4834 bp mitochondrial DNA 'common deletion' using a quantitative competitive polymerase chain reaction (PCR) assay. Mitochondrial respiratory chain activity, as measured by mitochondrial oxygen consumption using an oxygen electrode, did not vary between the treated animals and the controls. We conclude that the common mitochondrial DNA deletion and oxygen consumption are not good indicators of testicular damage induced by torsion. PMID- 11114981 TI - Penile prosthesis surgery under local penile block anaesthesia via the infrapubic space. AB - The aim of this study was to evaluate the effectiveness and patient tolerance to local penile block anaesthesia via the infrapubic space with penile prosthesis implantation. Local anaesthesia was administered using a 23-guage 1.5-inch needle. A 50-50 mixture of 0.5% bupivicaine (Marcaine) and 0.5% lidocaine (Xylocaine) without adrenaline was injected into the infrapubic space with additional subcutaneous penile ring infiltration at the level of the penile root. A total number of 159 patients underwent this technique, mean age 57 years (range 34-86). In 148 (93%) patients, no booster sedation was needed; eight (5%) patients needed a boost of the pre-operative sedative during crural dilatation; three (1.8%) patients required general anaesthesia owing to insufficiently effective local anaesthesia and unexpectedly difficult dilatation. It is concluded that local anaesthesia was effective and safe to produce a pain-free procedure in 93% of cases. However, as the need for booster sedation or general anaesthesia exists, the procedure should be performed under monitored anaesthetic care and pre-operative evaluation should be performed as for general anaesthesia. PMID- 11114982 TI - Constitutive production of interleukin-1alpha mRNA and protein in the developing rat testis. AB - Interleukin-1 (IL-1), a multifunctional cytokine produced mainly by activated macrophages, is also produced in the intact testis. Rat testicular IL-1 was found to be identical to IL-1alpha, judged by immunoneutralization of the bioactive protein and sequence comparison of cloned rat testicular and macrophage pro-IL 1alpha cDNA. Testicular IL-1alpha mRNA was first demonstrated on postnatal day 15, and the corresponding bioactive protein from day 20. IL-1alpha mRNA was still low on day 20, but then increased rapidly in parallel with the bioactive protein to establish a plateau level from day 25. In adult testes, IL-1alpha mRNA and immunoreactive protein were low in stage VII of the seminiferous epithelial cycle, whereas other stages showed a clearly detectable expression. In the adult testis, the concentration of IL-1alpha was 75 pg/mg testicular protein (approximately 200 pM). In conclusion, production of testicular IL-1alpha is developmentally and stage-dependently regulated, probably at the transcriptional level, emphasizing an important paracrine role in testicular function. PMID- 11114983 TI - Reproductive function in male rats after brief in utero exposure to diethylstilboestrol. AB - Long-term effects of brief in utero exposure to diethylstilboestrol (DES) during a foetal period known to be critical for gonadal development were evaluated. Rats were exposed to DES (100 microg/kg body-weight) from day 17 to 19 of pregnancy. All of the DES-treated pregnant rats (11/11) ate parts or whole of their offspring during the first day after birth (p=0.03). Surviving male offspring were examined on day 63 post-partum. DES induced a reduction in weight of the testis (p=0.06) and ventral prostate (p=0.07), even after this short exposure. DES tended to reduce the number of Sertoli cells (p=0.13). Our findings indicate that even a short in utero exposure of rats to DES during a critical period for gonadal development results in cannibalism and reduced testis and ventral prostate weight. PMID- 11114984 TI - Editorial. PMID- 11114985 TI - Recent developments in UK nurse education: horses for courses or courses for horses? PMID- 11114986 TI - This month in jan PMID- 11114987 TI - Nurses and whistleblowing: the ethical issues. AB - Whistleblowing - the public exposure of organizational wrongdoing - presents practical and ethical dilemma for nurses, and needs to be seen as part of a spectrum of increasingly confrontative actions against miscreant organizations by their employees. The ethics of whistleblowing can only be understood in relation to its moral purpose, whether that is to achieve a good outcome (a consequentialist view) or fulfil a duty (a deontological view). The consequentialist perspective is unable on its own to resolve problems arising from the balance of good and harm resulting from the act of whistleblowing (where considerable harm might be caused) or of responsibility for that harm. A deontological approach provides an analysis of these problems but raises its own problem of conflicting duties for nurses. However, a strong argument can be made for the precedence of the nurse's duty to the patient over her duty to the employer. Although both duties are based on an implicit or an explicit promise, the promise to a person (the patient) must take precedence over the promise to an organization. It can even be argued that duty to the employer may in fact justify whistleblowing by nurses in some circumstances. However, the consequences of whistleblowing are forced upon nurses in a different way by the fact that the danger of reprisals acts as a deterrent to whistleblowers, however justified their actions may be. A more robust approach to the protection of whistleblowers is needed on the part of the government and the National Health Service (NHS) to remedy this situation. PMID- 11114988 TI - Setting the context for research: exploring the philosophy and environment of a cancer clinical trials unit. AB - This paper describes a process of context setting that was undertaken prior to designing a study to assess the psychosocial impact of participation in phase I and II anticancer drug trials from the patient's perspective. The paper outlines how and why this context setting was undertaken and highlights important aspects of the culture and organization of cancer clinical trials that may influence patients' experiences in trial recruitment and participation. In this way, the context setting proved to be an invaluable tool for providing an orientation to the environment where patients received their care and treatment as well as identifying issues that would need to be taken into consideration later in the research study design. PMID- 11114989 TI - Care under threat in the modern world. AB - Despite enormous progress in the understanding and treatment of disease during the 20th century, the amount of care individuals receive from health professionals is arguably less than in previous decades. Being in the presence of caring people who practised human caring has always been the bedrock of services to individuals who were ill. With the rise of scientific positivism in the mid 19th century, traditional ways of caring for sick people, not susceptible to scientific investigation and intervention, were either abandoned or discouraged. The spread of outcome-orientated health services has led to care being redefined as the provision of the finest form of treatment that is financially viable. The spectre of a service in which the human dimension of caring is either prescribed or seen as invalid gives cause for concern. This paper argues for urgent re examination of what we understand by 'care'. PMID- 11114990 TI - A comparison between the ethics of justice and the ethics of care. AB - The parameters of the problem within which the principal aim of the present article will be addressed can be described as follows. When making ethical decisions there are different perspectives that health care professionals may use. This may lead to conflict and insufficient co-operation between the members of the health team. Two of these perspectives are the ethics of justice and the ethics of care. The ethics of justice constitutes an ethical perspective in terms of which ethical decisions are made on the basis of universal principles and rules, and in an impartial and verifiable manner with a view to ensuring the fair and equitable treatment of all people. The ethics of care, on the other hand, constitutes an ethical approach in terms of which involvement, harmonious relations and the needs of others play an important part in ethical decision making in each ethical situation. To seek some sort of way of avoiding conflict and promoting a mutual understanding about ethical decisions in the health team, there is a need to examine the ethics of justice and ethics of care. In order to understand the ethics of justice and ethics of care, the purpose of this article is to compare the two ethical perspectives. It is argued that the ethics of justice and the ethics of care represent opposite poles. If the members of the health team were to use only one of these two perspectives in their ethical decision-making, certain ethical dilemmas would almost certainly remain unresolved. Both the fair and equitable treatment of all people (from the ethics of justice) and the holistic, contextual and need-centred nature of such treatment (from the ethics of care), ought therefore to be retained in the integrated application of the ethics of justice and the ethics of care. PMID- 11114991 TI - An integrated approach to ethical decision-making in the health team. AB - When making ethical decisions there are different perspectives that health care professionals may use. This may lead to conflict and insufficient co-operation between the members of the health team. Two of these perspectives are the ethics of justice and the ethics of care. In a bid to gain a better understanding of the nature of ethical decision-making in the health team, a comparison was drawn between the ethics of justice and the ethics of care. The investigation into and comparison between the ethics of justice and the ethics of care revealed that the deficiencies in each of the two perspectives in isolation, in fact, necessitate the application of a combination of both perspectives. The aim of the article is to describe how the members of the health team can, in an integrated manner, apply both the ethics of justice and the ethics of care in their ethical decision making. The central argument of the article is based on the following premises: (1) the inadequacy of the ethics of justice and the ethics of care in isolation necessitates that both these perspectives be applied; (2) the application of both these perspectives again requires an extended rationality and discourse and (3) discourse, in its turn, requires that the emphasis falls on a specific telos and that the participants in the discourse be endowed with certain virtues in order to abide by the rules of discourse. PMID- 11114992 TI - Health promotion and the role of the school nurse: a systematic review. AB - This paper describes findings from a systematic review of the literature on the effectiveness of school nurses in promoting the health of school children. The paper gives a brief account of the background to the study and the search strategy adopted. Some key findings are presented and discussed. The brief for the review was to seek evidence of effectiveness in the practice of school nurses. The results of the review were disappointing, in that little research of acceptable quality was found and little could be said about effectiveness. The result is therefore a more diffuse review that gives a summary of descriptive research and current views and opinions, although it does also present some pointers for future research. The study was funded by Health Promotion Wales. PMID- 11114993 TI - Development and definition of the role of the operating department nurse: a review. AB - In the current cost-conscious National Health Service (NHS), the role of the nurse during anaesthesia and surgery is one that has interested health service managers keen to know what happens behind the closed doors of the operating department. It is clear that if nurses working within this specialized setting are to secure a future in providing care for surgical patients, then it is important to clarify and articulate exactly what it is that their role involves. The aim of this paper is to examine the role of the operating department nurse. First, it will illustrate how the role of the nurse has evolved alongside medical and technical advances in surgery, particularly in the last century. Second, it will highlight that while definition of the role has received attention in the North American literature, references in the British literature as to what it is that operating department nurses do, are scant. Finally, it will address the evolving role of the contemporary perioperative nurse highlighting the changes and challenges that nurses who work within this setting are currently facing. It is suggested here that nurses need to engage in role definition in order to be clear about their direction for the future, particularly within the fast changing, technologically driven environment of the operating department. PMID- 11114994 TI - Adults with terminal illness: a literature review of their needs and wishes for food. AB - Food refusal can be a source of conflict between dying people and their caregivers. This review examines: the nature and purpose of food; some reasons for and implications of anorexia in terminal illness; ethical principles underpinning responses to declining appetite and food refusal; social transactions between dying people and their caregivers in relation to needs and wishes for food; and the need for further empirical research. The nature and purpose of food in human societies has been studied extensively by anthropologists but the knowledge gained is not often imported into health care practice, where eating is seen from a medical rather than an anthropological perspective. Food refusal may be a consequence of anorexia which is the result of physiological or psychological changes or it may be a deliberate choice in acceptance of impending death. Ethical principles underpinning responses to declining appetite and food refusal have been studied extensively and clear guidance obtained about what would be appropriate behaviour in given circumstances. There is little published empirical work on social transactions between dying people and their caregivers in relation to needs and wishes for food. As the contribution made to effective care-giving by high-quality interpersonal relationships is widely recognized, further knowledge about how best to sustain such relationships in these important circumstances would be useful. Moreover, as such interpersonal relationships often occur in an institutional context, it may be that more can be learnt from close examination of social transactions about how best to structure organizational processes to maximize autonomy and comfort for patients at the end of life. Further research is indicated. PMID- 11114995 TI - Understanding beliefs and meanings in the experience of cancer: a concept analysis. AB - Although the concepts of belief and meaning are commonly used in the cancer literature, there is often an overlap in the use of the terms. Some consider the two terms as synonyms while others link them as successive elements in adjustment. Using an adaptation of the methods of concept analysis, this article differentiates belief and meaning, and also suggests that meaning exists at two levels. The defining attributes and antecedents of these closely related concepts are identified and a model case illustrating each is presented. Clarity in the conceptual definitions of beliefs and meanings can help researchers select measures that accurately reflect the phenomenon of interest. Similarly, differentiation between the concepts can help practitioners in planning focused interventions that explore clients' existing beliefs and situational and existential meanings. PMID- 11114997 TI - Concept analysis: chronic fatigue. AB - This concept analysis attempts to clarify and analyse the concept 'chronic fatigue' and does so by utilizing the framework outlined by Walker and Avant (1995). The aim is to use this work to underpin future research into the care of patients suffering from chronic obstructive pulmonary disease (COPD). The literature revealed no universal definition of fatigue and a confusion exists between fatigue and chronic fatigue. Three concept analyses are considered to assist clarification. Everyday meanings are sought as well as meanings revealed in poetry. A continuum from tired to exhausted is identified and definitions offered. Defining attributes are decided upon which will be used as an operational definition in later research. Constructed cases are created; antecedents and consequences are devised from the literature; scales, tests and descriptions of general appearance which appear within the literature are considered as empirical referents. However, whilst assisting with the understanding of the concept and future research, the complexity of this subject is still evident. PMID- 11114996 TI - Exploration of the concept of hope in the Dominican Republic. AB - An ethnonursing study explored What is the meaning of hope (esperanza/esperar) to the Dominican people? What is the importance of the concept of hope to the people of the Dominican Republic? What are the universals and diversities in the meaning of hope for this cultural group and United States' (US) mainstream culture? Hope has been considered universal and recent research has demonstrated the effect of levels of hope or hopelessness on health. Nurse researchers have explored the meaning of hope to patients in the US and have developed instruments to measure levels of hope. However, little research has explored whether hope has different attributes in various cultural groups. From data gathered in a rural Dominican village, a definition of hope and its attributes were developed: 'Hope is an essential but dynamic life force that grows out of faith in God, is supported by relationships, resources and work, and results in the energy necessary to work for a desired future. Hope gives meaning and happiness'. This definition and its attributes were compared with definitions developed from research among mainstream US populations to propose universals and diversities of hope. PMID- 11114998 TI - Fatigue in persons with renal failure who require maintenance haemodialysis. AB - Fatigue is a highly prevalent symptom experienced by persons who live with chronic illness, including those with renal failure who require maintenance haemodialysis. Fatigue, however, is a non-specific and invisible symptom and is a phenomenon that is poorly understood by health care professionals. This study examined the symptom of fatigue as experienced by a group of 39 adult haemodialysis patients. The theory of unpleasant symptoms formed the conceptual framework for the study. A descriptive correlational design was utilized to examine fatigue from an inductive approach, considering relevant physiological, psychological and situational variables based on a review of the literature. Data were collected using a structured self-report questionnaire and biochemical data from retrospective monthly blood tests. The results of the study indicated that high levels of fatigue are experienced, with correspondingly low levels of vitality, in all the areas measured - general fatigue, physical fatigue, reduced motivation, reduced activity and mental fatigue, by adult haemodialysis patients. Individual variation was noted in the dimensions of fatigue predominantly expressed. Fatigue was significantly associated with the presence of symptoms such as sleep problems, poor physical health status and depression. No associations between fatigue and the biochemical and situational variables measured were noted. Further examination of the data revealed complex relationships between the physiological and psychological factors examined. Depression was significantly associated with physical health status, sleep problems, symptoms and anxiety. Correlations were also noted between symptoms and poor physical functioning, sleep problems and depression. Based on the results, a revised version of the theory of unpleasant symptoms relating to fatigue is presented. PMID- 11114999 TI - Women's views of counselling received in connection with breast-feeding after reduction mammoplasty. AB - The aim of this study was to generate a theoretical model of the experiences of women, who had undergone reduction mammoplasty, of counselling received in connection with breast-feeding. Data were collected through interviews with 12 breast-reduced women who had given birth to a child, and the material was analysed by means of the grounded theory method. Written permission to post notices at the child welfare centres had been obtained from the primary care managers. The women themselves applied for participation in the study. The results showed that their need for counselling from somebody who listened attentively to them and gave advice in connection with breast-feeding, was considerable. The women could have a feeling of self-reliance, ambivalence, acceptance or guilt, depending on the extent to which they perceived to have received counselling from their family members. At the same time, their experiences of the encounter with the nursing staff were of crucial importance. Lack of active listening and counselling from the nursing staff can be counterbalanced by support from relatives. However, when such support is not forthcoming, the nursing staff must be able to support the women in their role of a mother, which requires that education and supervision are given to the staff. The findings of the study can be used as a basis for further research into the need for support of the families concerned. PMID- 11115000 TI - Does the duration of postnatal stay influence breast-feeding rates at one month in women giving birth for the first time? A randomized control trial. AB - The aim of the study reported in this paper was to examine the effect of postnatal stay on breast-feeding rates at one month using a randomized control trial. Participants were recruited during parent-craft classes at a large teaching hospital in the north of England. Nulliparous women in the last trimester of pregnancy were randomly allocated to a short hospital postnatal stay (6-48 hours), or a longer stay (more than 48 hours). The mothers were contacted at one month following the birth to ask about the method of feeding. The study was approved by the hospital ethical committee, and participation was voluntary. The results demonstrated no significant effect of postnatal stay on breast feeding rates at one month. The main limitation of the study was the reluctance of the mothers in the long stay group to stay in hospital for longer than three days. This resulted in only a small difference between the lengths of hospital stay of the two interventions. The overall breast-feeding rate for the study group had increased significantly when compared with local city wide rates. This increase may be as a result of a sampling bias or a Hawthorne effect. PMID- 11115001 TI - Successful breast-feeding as a result of a health education programme for mothers. AB - The present study aims to prove an improvement in breast-feeding practices in mothers who received written instructions for successful breast-feeding and had individual counselling at the time of taking this questionnaire. The instructions were based on the results of a study carried out in Slovenia in 1993 and published in the Journal of Advanced Nursing (1998), 27, 1250-1256 (Hoyer & Pokorn 1998). The variables that were then found statistically significant in the initial and continued breast-feeding became the basis of written instructions for breast-feeding. The results obtained in the new current study were compared with that basic study which included 881 mothers. This was possible because the conditions of breast-feeding between 1993 and 1995 (when the new study was launched) had not changed. The idea of baby-friendly hospital initiative (BFHI) had not yet come into force in Slovenia. The current study comprised 203 pregnant women who were first visited during their 8th month of pregnancy. In order to collect data on breast-feeding and deliver instructions, a field nurse visited each mother eight times until the completion of first year and continued to visit them every third month and for all those who were still breast-feeding after that time, until the end of the lactation period. Statistical analysis was carried out by means of SPSS statistical package and survival analysis. All mothers in the study started breast-feeding. By the end of the first month 84.7% of them were still breast-feeding, while by the end of the third month it dropped to 74.9%, and by the end of the sixth month to 45.8%. Among these, breast-feeding alone was practised in the first week by 25.7%, by the end of the first month by 16.4%, and by the end of the third month by 9.5%. All the observed parameters were better than in the basic study. The mean duration of breast-feeding was 217 days, while the longest duration was 852 days. The survival analysis showed a statistically significant difference in the duration of breast-feeding. In the current study group, mothers weaned their infants at a slower rate than in the group from the year 1993. It has been found that the written instructions as well as personal encouragement by the field nurse exerted a favourable influence on breast-feeding practices, which was taken as a guideline for our further professional work and change of standards in the field of breast-feeding promotion. PMID- 11115002 TI - Personal control in pain relief during labour. AB - Personal control is a central feature of women's involvement in their childbirth experiences. To achieve this control tacit rules and guidelines are applied to define how women and the professionals who care for them should behave. This study investigated the extent to which women exercised control in pain relief during the first stage of labour by comparing (a) the rules which they held prior to childbirth (2-3 cm cervical dilatation) with those which they afterwards felt applied to their labour and (b) the rules held by the women before and after childbirth with those held by the midwives. In a quantitative study using a repeated measures design, a questionnaire was administered to 35 midwives and to 100 women prior to and within 24 hours following their delivery. Consistency of the women's scores before and after childbirth, indicated by few statistically significant differences, tended to confirm their rules on control of pain relief. Some of the rules were held even more strongly following childbirth. A surprising finding was the even stronger agreement by midwives with some of the rules. There was a definite trend for many of the rules held by the women prior to childbirth to increase following birth towards those of the midwives. This could be the result of the experience of childbirth per se but the possibility that it was contributed to by the influence of the midwives cannot be ruled out and warrants further research. An interesting hierarchy in the rules for compliance with professional care has been highlighted. PMID- 11115003 TI - 'I don't have any other choice': spouses' experiences of placing a partner in a care home for older people in Sweden. AB - The main aim of this paper is to consider the experiences of Swedish spouses who have placed a partner in a care home for older people. Data were gathered from semi-structural interviews with 14 spouses (11 wives and 3 husbands) who had been involved in a care home placement within the previous 6 months. The results reported here are from the first component of a larger grounded theory study, the aim of which is to explore, describe and understand the experience of care home placement from a variety of perspectives and to identify the implications for policy and practice in Sweden. The focus here is on the experience of spouses, relating to the decision-making process, the move into care and subsequent contact with the care home. Four themes emerged from the data - making the decision, making the move, adjusting to the move and reorientation. The results show a lack of planning for the elder person's entry to a care home, and professional dominance of this stage of the process. The largely ambivalent emotional responses to the move that spouses experience and the difficulties in initiating and sustaining relationships with staff in the home are discussed in the light of previous research. PMID- 11115004 TI - Nursing home placement: an exploration of the experiences of family carers. AB - This study examines some of the factors leading family carers to place their older relatives in a nursing home. It also explores their thoughts and feelings about their relatives' admission to a nursing home. Analysis of in-depth interviews with relatives (n=10) found that the decision to place an older relative in a nursing home was a difficult one for families. The interviewed carers stated that admission to a nursing home was held off as long as possible but the deteriorating health of the older relative and in some cases their own health meant that there was no other option. Admission to institutional care usually followed a period of prolonged home care and occurred at a time of crisis. Family carers complained that they were given inadequate support from health care professionals and often had no choice in the decision-making process. This was particularly evident in the case of carers whose relative was transferred directly from hospital to a nursing home setting. The majority of carers in this study experienced ambiguous feelings about placing their elderly relative in a nursing home. Feelings of relief that the burden of care had been lifted, contrasted sharply with feelings of guilt that they could not continue with their 'duty of care'. Families also felt a need to justify their decision by emphasizing how friends and other family members agreed that they could not continue with home care in the interests of both their own and the older relative's health. The findings suggest that while many carers are relieved of the physical exhaustion surrounding home care, the emotional turmoil continues long after admission to a nursing home. PMID- 11115005 TI - Using restraint with nursing home residents: a qualitative study of nursing staff perceptions and decision-making. AB - The study reported in this paper applied a qualitative and interpretative approach to nursing staff perceptions of the use of restraint with elderly nursing home residents, and into nurses' decision-making on restraint use. The data were collected using unstructured interviews with a purposive sample of 20 trained and untrained nursing staff from two Swiss nursing homes. Data analysis was based on Colaizzi's phenomenological method. Three main themes were extracted from the data: (1) understanding the term restraint, (2) situations in which the decision to apply restraint is considered justified and (3) situations in which nursing staff are uncertain about the use of restraint. The underlying bases with respect to decision-making were: understanding restraint, the rights and responsibilities of both residents and staff, and the duties of staff. Staff members were ambiguous in their understanding of restraint and they showed positive as well as confused attitudes towards its use. Their behaviour was defensive and protective rather than challenging. Further research is required on what is meant by safety in care of the elderly nursing today. In nursing practice, as far as issues of restraint are concerned, greater attention should be devoted to the relationship between elderly residents' self-determination and responsibility for their actions. PMID- 11115006 TI - Dementia care mapping: an approach to quality audit of services for people with dementia in two health districts. AB - The audit reported in this paper and submitted to the Psychiatry of Old Age Management group, assessed six units within each of two health districts in the UK. Using a nonparticipatory observation method in the units selected, the aim was to measure quality and the environment of care. Dependency levels of the clients/residents were also estimated to give a clearer picture of the setting and the care requirements. This was intended to establish a baseline for the units mapped and to enable care developments to be focussed upon intended outcomes. Results led to a number of observations related to the levels of interaction between staff and clients/residents, the need for a wider range of activities to promote person-centred care, and a suggested route to the improvement in quality of life for this vulnerable group of people. Assessment of dependency levels linked to the results of the mapping showed that high dependency does not lead automatically to a lower quality of person centred care. PMID- 11115007 TI - The health-related quality of life in a Swedish sample of HIV-infected persons. AB - The purposes of the present study are (1) to assess the health-related quality of life (HRQOL) and the subjective health status in a sample of human immunodeficiency virus (HIV)-infected persons (2) to relate the results to different population groups and (3) to investigate the relationship of medical and demographic variables with HRQOL. A total of 72 HIV-infected men were included. They answered the Swedish health-related quality of life questionnaire and the health index. Demographic and medical data were obtained from the medical records. The data collection took place before entering a therapeutic HIV vaccine trial. The results showed a more negative impact on the HRQOL and subjective health status in the HIV-positive subjects, compared with male population groups. The dimensions of emotional well-being were most affected. When comparisons were made according to the medical and demographic variables for different subgroups within the HIV sample, differences in the physical-dimension scales were most prominent. Symptomatic HIV infection or acquired immunodeficiency syndrome (AIDS), anti-retroviral treatment, sick leave or disability pension, low income and basic education were associated with worse HRQOL and health status. In conclusion, it is of utmost importance to take into account, aspects of the patients' emotional well-being in nursing, as well as in medical care and interventions. Moreover, individualized caring programs are needed because the disruptions in HRQOL fluctuated within the HIV sample. PMID- 11115008 TI - The interplay between social and cultural context and perceptions of cardiovascular disease. AB - This paper seeks to explore the impact of social and cultural factors upon perceptions of the patients' cardiovascular risk and intended lifestyle changes. Qualitative and quantitative research approaches were used. The sample was purposeful; matched groups of 10 first time post myocardial infarction (MI) patients, 10 informal (spouse, blood relative or partner) and 10 formal carers (nursing staff) were selected on a convenience basis. Personal interviews were conducted with participants, during which a semistructured questionnaire was completed. A combination of descriptive statistics and qualitative analysis of these data revealed that social and cultural factors formed the basis of patients' and informal carers' perceptions of cardiovascular risks and social information networks provided the main source of information about cardiovascular risk. For the staff, known risk factors based on epidemiological evidence (lack of exercise and diet) formed the basis of their perceptions of the patients' cardiovascular risk factors. However, for all the participant groups (i.e. patients, informal carers and staff) the risk factors - diet and exercise were indicated as being the patients' intended lifestyle changes., The results illustrate potential interplay between 'social' and 'cultural' context and perceptions of cardiovascular risk and incongruency between perceptions of risk and perceptions of the patients' intended lifestyle change. These results may assist nurses in giving effective advice to patients and relatives about lifestyle change following myocardial infarction and inform future policy for cardiac rehabilitation. PMID- 11115009 TI - Heart health-associated health beliefs and behaviours of adolescents of African and African Caribbean descent in two cities in the United Kingdom. AB - The following paper presents the findings of an exploratory ethnography, the purpose of which was to identify and describe heart health associated beliefs and behaviours of year seven (Y7) and year 10 (Y10) secondary school young people of African and African Caribbean descent in two UK cities. Data were collected by the data collection technique of focus groups. However, eight focus groups were conducted involving 47 Y7 young people and 29 Y10 pupils, 76 pupils in total. The data were analysed utilizing ATLAS/ti qualitative data analysis software. This software is informed by grounded theory. Data from the study formed six themes. The findings informed the development of an interactive health promotion website which can be found at http://www.shef.ac.uk/web/uni/projects/mshhp. The paper argues that in order to provide meaningful programmes of health promotion to be developed by health care providers including school nurses and health visitors, it is essential that interventions are informed by an understanding of the health beliefs and behaviours of African and African Caribbean young people. PMID- 11115010 TI - Waiting for coronary artery bypass surgery: a qualitative analysis. AB - Waiting lists for coronary artery bypass surgery (CABS) are common in many developed countries. Yet, there is limited information available regarding patients' health care needs at this time. This paper reports on a prospective study which aimed to investigate the experience of waiting for CABS from a qualitative perspective. An inductive research approach was used to conduct interviews with 70 randomly selected patients at three intervals over the first year on the waiting list - referral for surgery, again after waiting 6 months (n=49), and finally after waiting for 1 year (n=28). Attrition was mainly caused by surgery having been performed (n=36), although death (n=4) and refusal to participate (n=2) also contributed. Domicilliary interviews were taped and transcribed verbatim. Thematic content analysis identified three central themes in this experience - uncertainty, chest pain and anxiety; with six secondary themes - powerless, dissatisfaction with treatment, anger/frustration, physical incapacity, reduced self-esteem, and altered family and social relationships. The nature and meaning inherent in each theme is described using interview quotations, and a model is proposed which summarizes this data and the relationship between themes. From this analysis, uncertainty, chest pain and anxiety emerge as important indicators of a negative outcome for these patients. This report strongly suggests that patients awaiting bypass surgery require more information regarding the waiting time for such a surgery. Nurses should also offer advice regarding pain management to help improve patients' skills and decrease the fear associated with angina. Nursing intervention and support should also be directed at reducing patients' anxiety levels. This is the first known qualitative study which specifically examines patients' perception of the waiting period prior to bypass surgery. It may therefore provide new evidence on which to base practice for nurses in both hospital and community, and may also stimulate further research in this area. PMID- 11115011 TI - Developing and testing an instrument for the measurement of individual care. AB - This paper describes a preliminary study in which an instrument was developed for the measurement of individual care in adult patients and in which the reliability and validity of the instrument were evaluated. Individual care was defined in terms of how patient individuality was taken into account and how patient participation in decision-making was facilitated. The purpose here is to describe the process by which the individual care instrument (IC) was developed, the preliminary testing of the instrument and psychometric evaluation of the instrument in a sample of adult patients discharged from a Finnish general Hospital (n=203). Item analyses showed an acceptable level of internal consistency reliability and homogeneity in each scale of the IC, and Cronbach alpha values were high in every measurement. Exploratory factor analysis supported a three-factor solution. The psychometric evaluations suggested that the instrument is worthy of further development. PMID- 11115012 TI - The critical incident technique and nursing care quality research. AB - The critical incident technique, is a highly flexible qualitative research method used in solving practical problems. Although this research method has been extensively used in the service industry to evaluate consumers' expectations and perceptions, applications to the study of health care quality are just beginning. This article describes critical incident methodology, reviews previous applications of the technique to the study of health care quality and provides illustrations from research. This practical research methodology offers the following important advantages to those interested in designing studies of care quality: identifying patients' experiences in health care settings, exploring dimensions of nurse-patient interactions and identifying patients' responses to illness and health care treatment. PMID- 11115013 TI - Nurse-sensitive outcomes of advanced practice. AB - Advanced practice nurses (APNs) in the USA are registered nurses who hold masters or doctoral degrees in a specialized area of nursing. They provide advanced clinical care to clients, manage health care systems and influence health care decision-making through expert clinical reasoning and research and theory-based action. APN impact on health care outcomes is supported by studies using physician-focused indicators, although a few studies have identified several that are sensitive to or reflective of advanced practice nursing. A modified Delphi survey was conducted during May 1997-December 1998 to determine the outcome indicators APNs recommend for use in measuring their effect on care delivery outcomes. A convenience sample of 66 APNs attending a statewide outcomes conference identified 27 potential outcome indicators. These indicators were included in a mailed survey sent to APNs working in Tennessee. Respondents were asked to rate each indicator for validity, sensitivity, feasibility, utility and cost. In the second round of the survey, they were asked whether or not they agreed with the rank ordering of indicators, which was determined by the means calculated from responses in the first round. The 10 highest ranked indicators were satisfaction with care delivery, symptom resolution/reduction, perception of being well cared for, compliance/adherence with treatment plan, knowledge of patients and families, trust of care provider, collaboration among care providers, frequency and type of procedures ordered and quality of life. APNs identified both direct and indirect measures of effect on care delivery outcomes. Some of these are currently used as indicators of advanced practice, but many are not. Additional research is needed to determine whether the indicators proposed are valid and sensitive to advanced practice care by nurses. PMID- 11115014 TI - Evaluating educational preparation for a health education role in practice: the case of medication education. AB - Current health care policy and practice contexts in the UK point to the importance of nurses' ability to make an effective contribution to educating patients about medication, as part of their role in health education and health promotion. Nurses' potential contribution to this important activity will inevitably be dependent on knowledge and skills acquired during preregistration and postregistration programmes of education. Against this backdrop, changes in pre and postregistration nurse education in the UK in the past decade highlight the importance and timeliness of evaluating the adequacy of educational preparation for a medication role. This paper reports on the findings from an evaluation of UK educational preparation for a medication education role in practice. A case study design was used to investigate current educational preparation at three education institutions. Multiple methods of data collection at each site involved focus group discussions with lecturers and practitioners, individual interviews with key personnel, nonparticipant observation of teaching sessions, postobservation interviews with students and curriculum analysis. Findings highlighted the importance of a number of dimensions of preparation for practice of such a role: the need for sufficient taught pharmacology; opportunities for application and integration of prerequisite knowledge and skills; the importance of practice-based learning; the need for an evidence-based curriculum, and the importance of clarifying outcomes and competencies required for a medication education role within pre and postregistration curricula. The paper concludes with a discussion and implications of the findings. PMID- 11115015 TI - Discourse analysis of an 'observation levels' nursing policy. AB - The practice of special observation (or constant observation) is widely used in inpatient psychiatric facilities for the care of people who are suicidal. In this study, the policy of special observation was examined using a discourse analysis method to discern prevailing ideas and practices highlighted within the policy. After reading, studying and analysing the special observation nursing policy, the authors briefly describe the document and outline the terms and phrases prevalent within the document. These recurrent ideas are then organized into five categories: professional responsibilities, suicidality, the patient's immediate context, the patient's observable behaviour and the nursing checklist. In discussion of the policy document, the invisibility of the authors, target audience and patients is noted. The authors attempt to elicit evidence for the therapeutic nurse-patient relationship in the document. In the analysis of patient, nurse and doctor roles and responsibilities, it is evident that the policy document reinforces the traditional medical hierarchy of power relations. Some assumptions that underpin the document are postulated. Questions regarding the nature of risk assessment and the evidence base for the medical prescription of special observation are raised. As well as ideas and themes evident in the document, the absence of some relevant issues is explored. While the need for succinctness and clarity in policy documents is acknowledged, the fact that patient rights, therapeutic processes and ethical dilemmas are absent is deemed significant. PMID- 11115016 TI - Clinical supervision as an emancipatory process: avoiding inappropriate intent. AB - As clinical supervision becomes more widely implemented in the United Kingdom with concerns and resistances being eroded as practitioners discover its benefits, it is important that potential limitations and perhaps failures are avoided. This paper utilizes Johns' (1996) intent-emphasis axis to explore how a technical interest, misunderstanding of expert practice, and confusion of self awareness with counselling, can detract from the supervisory process. Several of the criticisms of reflective practice will be examined to demonstrate where concerns are valid and where they may be based on misunderstandings and the need to control clinical supervision. Greater awareness of inappropriate emphasis and intent should enable even relatively inexperienced supervisors to help their supervisees move towards independence, emancipation and evolving expertise. PMID- 11115017 TI - Eating disorders in adults with intellectual disability. AB - There is an increasing focus on the nutrition of people with intellectual disability (ID), but less interest in the range of eating disorders (EDs) that they may exhibit and the bio-psycho-social impact of these conditions. Despite diagnostic and methodological difficulties, psychopathology and ED research studies suggest that 3-42% of institutionalized adults with ID and 1-19% of adults with ID in the community have diagnosable EDs. Weight surveys indicate that 2-35% of adults with ID are obese and 5-43% are significantly underweight, but the contribution of diagnosable EDs is unknown. Such data and case reports suggest that EDs are associated with considerable physical, behavioural, psychiatric and social comorbidity. Review papers have focused on the aetiology and treatment of pica, rumination, regurgitation, psychogenic vomiting and food faddiness/refusal. Emerging clinical issues are the development of appropriate diagnostic criteria, multimodal assessment and clinically effective treatment approaches. Key service issues include staff training to improve awareness, addressing comorbidity and access issues, and maintaining support for adults with ID and EDs, and their carers. Research should confirm the multifaceted aetiology and comorbidity of EDs. Then multicomponent assessment and treatment models for EDs can be developed and evaluated. PMID- 11115018 TI - Cerebral dominance and schizophrenia-spectrum disorders in adults with intellectual disability. AB - Studies of the general population without intellectual disability have suggested an association between atypical handedness and schizophrenia-spectrum disorders (SSDs). Mixed handedness is taken as an index of diminished cerebral dominance or laterality. The present study addressed the question of whether such findings extend to the neurodevelopmentally 'at risk' population of adults with intellectual disability and SSDs compared with appropriate controls. Fourteen patients with a dual diagnosis of intellectual disability and SSD were compared with 14 controls with intellectual disability alone. Assessments of self-reported hand preference and relative hand skill were completed. Self-report of hand preference revealed highly significantly greater mixed-handedness in the SSD group. Furthermore, relative hand skill performance was significantly diminished for the dominant hand. The discrepancy between dominant and non-dominant hand functioning was lower in the SSD group and this association was highly significant. The results of the present study support the usefulness of such detailed laterality assessment in this population. Mixed laterality, over and above that of the population with general intellectual disability and developmental disorder, was associated with SSD. These results are consistent with the neurodevelopmental hypothesis of schizophrenia and its cognitive neuropsychiatric/neuropsychological sequelae. PMID- 11115019 TI - Alpha2 macroglobulin elevation without an acute phase response in depressed adults with Down's syndrome: implications. AB - Studies of immune function during depression in persons without intellectual disability (ID) have revealed elevated levels of alpha2 macroglobulin (alpha2M) and an acute phase protein (APP) response. Clinical observation suggests that people with Down's syndrome (DS) may have associated genetic abnormalities in their immune systems. The APP response and alpha2M changes in depressed versus non-depressed adults with DS was the subject of the present study. The serum pan proteinase inhibitor alpha2M, and the AP proteins c-reactive protein (CRP), alpha1 antitrypsin (alpha1AT), ceruloplasmin (Cp), beta2 Macroglobulin (beta2M), transthyretin (Trans), serum amyloid protein (SAP), and albumin (Alb) were measured in 38 adults with DS, 19 of whom were diagnosed with and 19 without depression using a sandwich enzyme-linked immunosorbent assay (ELISA). The DSM-IV criteria were used for diagnoses. Medical and neurological examinations excluded medical disorders associated with APP response. Only alpha2M and CRP were significantly different in the depressed versus non-depressed groups. The alpha2M was higher, a response similar to one observed in depressed people without ID, but the CRP was lower in the depressed group, especially in those subjects not on psychotropic medications, contrary to the expected APP response to depression. The results suggest that alpha2M elevation in depressed adults with DS is independent of the APP response. An alternative explanation for its elevation is proposed linking the core symptom of depression with the mammalian dormancy/hibernation process. Further studies are needed to confirm that alpha2M elevation is specific to depression and that it might provide a helpful marker for the diagnosis of depression in people with ID. PMID- 11115020 TI - Sequence of cognitive decline in dementia in adults with Down's syndrome. AB - Adults with Down's syndrome (DS) are known to be at risk of dementia of the Alzheimer type (DAT), but because of their lifelong intellectual deficits, it is difficult to determine the earliest signs and characteristics of age-associated decline and dementia. In a longitudinal study in which all participants were healthy at the time of their entry into the study, the present authors compared the amount of decline on the subtests of the WISC-R to determine the sequence of cognitive decline associated with varying stages of dementia. Twenty-two individuals with varying degrees of cognitive decline were compared to 44 adults with DS who have remained healthy. All participants functioned in the mild or moderate range of intellectual disability at initial testing. On each subtest of the WISC-R, the amount of change experienced by the healthy participants over the study period was compared to the amount of change found for each of the groups with decline. Out of the individuals who showed declines, 10 adults with DS were classified as having 'questionable' decline based on the presence of memory impairment, and five and seven adults with DS were classified as in the 'early stage' and 'middle stage' of DAT, respectively, based on the presence of memory impairment, score on the Dementia Scale for Down Syndrome and a physician's diagnosis. It was found that participants who were identified as 'questionable', in addition to the memory loss that determined their classification, also showed significant declines on the Block Design and Coding subtests. The five adults in the early stage of dementia showed declines on these subtests, and in addition, on the Object Assembly, Picture Completion, Arithmetic and Comprehension subtests. The seven adults in the middle stage of dementia showed declines on these subtests, plus declines on Information, Vocabulary and Digit Span subtests. The Picture Arrangement and Similarities subtests were not useful in distinguishing between the groups because of baseline floor effects for a substantial proportion of participants. The present longitudinal study showed a sequence of cognitive decline associated with DAT, beginning with a possible 'pre clinical' stage, and progressing through the early and middle stages. This approach begins to define the sequence of declining cognitive capacities that contributes to the observed functional deterioration caused by Alzheimer's disease and that is likely to reflect the involvement of cortical areas as the disease progresses. PMID- 11115021 TI - Receipt of psychotropic medication by people with intellectual disability in residential settings. AB - Previous studies have reported that the rate of prescription of antipsychotic medication for people with intellectual disability is far in excess of the expected prevalence of psychoses for this population. Recent research identifying factors which predict the use of psychotropic medication suggests that challenging behaviour may play a key role in determining the receipt of antipsychotic medication. The present study reports the prevalence of psychoactive medication receipt for 500 people with intellectual disability living in different forms of residential provision in the UK. Variables which predict the receipt of psychotropic medication are also identified. The results show differences between forms of residential provision in rates of medication receipt. Analyses of predictors of psychotropic medication receipt suggest that, whilst the receipt of antidepressants is predicted by symptoms of mental ill health, the receipt of both antipsychotics and hypnotics/anxiolytics is predicted by variables related to challenging behaviour. PMID- 11115022 TI - Olanzapine for chronic, stereotypic self-injurious behaviour: a pilot study in seven adults with intellectual disability. AB - Dopamine one (D1) receptor supersensitvity in the corpus striatum is said to be the primary mechanism within the dopamine model proposed for chronic, refractory self-injurious behaviour (SIB), which may explain why conventional neuroleptics have proven largely ineffective. In common with other atypical antipsychotic agents, olanzapine has more affinity for the D1 receptor. The present study explored whether olanzapine could reduce rates of the stereotypic form of chronic SIB, a subtype where dopamine dysfunction is the most likely underlying mechanism. A clinical sample of seven patients with various levels of learning disability who displayed features of stereotypic SIB were assessed over a 6-week period of baseline measurement and a 15-week treatment phase during which olanzapine was added to existing medication. Both SIB and other aberrant behaviours were measured by daily nurse rating and the Self-Injury Trauma Scale (SITS). All measurements were unblind. Doses ranged from 5 to 15 mg. Out of the seven subjects, three showed a clear improvement, one showed a marginal improvement, one deteriorated, and the data was equivocal for the remaining two individuals. The means of the SITS Number and Severity Indices (NI and SI, respectively) reduced significantly from baseline during both the 5- and 10-mg treatment phases, and taking treatment as a whole, by 53% and 48%, respectively (NI: mean = 0.7 units reduction, P = 0.02; SI: mean = 0.9 units reduction, P = 0.04). The risk index also reduced, but did not reach significance. A modest reduction in mean nurse-rated SIB was not significant for either phase or for treatment as a whole. At doses above 5mg, mean scores deteriorated on balance, although two responders showed a marginal additional improvement. Olanzapine was well tolerated with one adverse event reported (somnolence) which was mild and transient. The present pilot study suggests that olanzapine can reduce stereotypic SIB. A larger trial is indicated. PMID- 11115023 TI - BIOMED-MEROPE project: service provision for adults with intellectual disability: a European comparison. AB - The aim of the present paper is to describe and compare services for adults with intellectual disability (ID) and mental health needs in five European countries: Austria, England, Greece, Ireland and Spain. A framework and structure for collecting information about service provision was designed. This information was collected through a mixture of interviews with service providers, questionnaires and a review of the research literature within each country. Information was collected on historical context, policy, legislation, assessment, treatment and the structure of services for people with ID and mental health problems. Overall, the needs of those with additional mental health needs have not been specifically addressed at a national level with perhaps the exception of England and Ireland, although there are still gaps in services in these nations. Normalization has been adopted in each of the five countries, and there are moves toward deinstitutionalization, integration and inclusion. Families and self-advocacy groups have grown. The pace of this change varies between and even within countries. The main findings of the study include: unclear policy, trends for legislative changes, increased prevalence of mental health problems, inadequate generic service provision, a need for specialist mental health services, a need for improved interconnections of services, and a need for training developments. Policy and legislation in the five European countries under consideration tend to separate the disability aspects of people with ID from their mental health needs. Consequently, the service needs of this group remain largely invisible. This might be a direct reflection of policy clarity and legislation, or could be the result of a failure to implement existing guidelines. This has a detrimental effect on the lives of people with ID, and their families and carers. PMID- 11115024 TI - The dementia scale for Down's syndrome. PMID- 11115025 TI - Cisapride treatment for gastro-oesophageal reflux in children: a systematic review of randomized controlled trials. AB - The aim of the systematic review was to determine the effect of cisapride compared with placebo or other non-surgical therapies for the treatment of symptoms of gastro-oesophageal reflux in children. We searched MEDLINE, EMBASE, the Cochrane Controlled Trials Register, Science Citation Index and reference lists for randomized controlled trials which compared cisapride with placebo or other non-surgical therapy in children. We included only trials which reported reflux-related symptoms as an outcome, provided that cisapride was administered orally for at least I week. Seven trials (286 children in total) compared cisapride with placebo. Two trials reported good concealment of treatment allocation. The pooled odds ratio for the 'same or worse' symptoms was 0.34 (95% CI 0.10, 1.19). There was substantial heterogeneity between studies (P < 0.00001) and the funnel plot was asymmetrical. Adverse effects (mainly diarrhoea) were not significantly increased with cisapride (pooled odds ratio (OR) 1.80: 0.87, 3.70). The reflux index was significantly reduced in children treated with cisapride (weighted mean difference -6.49: -10.13, -2.85). One study (50 children) compared cisapride with gaviscon plus carobel: the OR for the 'same or worse' symptoms was 3.26 (0.93, 11.38). There was no clear evidence that cisapride reduced symptoms of gastro-oesophageal reflux. As smaller, poorer quality studies were biased in favour of a positive treatment effect, the pooled OR overestimated the potential benefits of cisapride. There was some evidence to suggest that gaviscon plus carobel may be a more effective option than cisapride. PMID- 11115026 TI - Management of tuberculosis in children. AB - This paper provides specific guidelines on the management of tuberculosis infection and disease, covering general principles, recommended drug regimens and discuss the evidence to support these. It also covers use of corticosteroids, intermittent therapy, directly observed therapy and an approach to the management of a patient with drug-resistant tuberculosis. PMID- 11115027 TI - Home visiting for vulnerable infants in Australia. PMID- 11115028 TI - Sudden infant death syndrome in South Australia 1968-97. Part I: changes over time. AB - OBJECTIVE: To compare the epidemiology of sudden infant death syndrome (SIDS) over three consecutive decades. METHODOLOGY: The birth history, infant's developmental and health history, infant care practices for the infant, death scene investigation and autopsy findings for all infants dying suddenly and unexpectedly in South Australia (SA) between January 1968 and December 1997 were studied. RESULTS: The incidence of SIDS in SA rose through the 1970s and early 1980s with the highest incidence being in infants born in 1986 at 2.4 per 1000 live births (LB). Two factors felt to be dangerous for some infants were identified being left unobserved in the prone position and having the head covered by bed clothes. Publicity about the risk of prone sleeping has been accompanied by a fall in SIDS deaths, to an incidence of 0.5 per 1000 LB in 1997. The incidence in Aboriginal infants, and infants living in lower socio-economic conditions has always been high, but the over-representation of these groups has increased in the last 5 years. CONCLUSION: It no infant under 8 months of age was placed prone or was able to get to prone unobserved before the age when they can easily get back to supine, and no infant was able to get the head completely covered while unobserved, the incidence of SIDS in SA should fall below 0.2 per 1000 LB. PMID- 11115029 TI - Sudden infant death syndrome (SIDS) in South Australia 1968-97. Part 2: the epidemiology of non-prone and non-covered SIDS infants. AB - OBJECTIVE: To identify the risk factors for infants who die suddenly and unexpectedly, but whose deaths are not related to prone position, or having the head covered. METHODOLOGY: A case-control study was designed in which the cases were infants who had died of sudden infant death syndrome (SIDS) in South Australia between January 1974 and December 1997, who were found not prone, not bed sharing and with the head not covered. The controls were two infants for each case, born in the same year and found in the prone position (again not bed sharing and with the head not covered). RESULTS: Sudden unexpected death infancy is rare in non-prone infants with the head not covered. occurring on average twice a year in South Australia, where there are 18,000-21,000 births per year. In this group there was a higher percentage of infants with features associated with low socio-economic groups (teenage pregnancies and maternal smoking), sibling SIDS, suspicion of non-accidental injury and the presence of minor congenital anomalies, especially cardiac anomalies. CONCLUSIONS: The majority of unexpected deaths in infancy can be prevented by not allowing infants to be unobserved in prone position, and by preventing them from getting their faces covered. For the few infants not found in these positions, a careful investigation should be made for malformations or non-accidental injury. PMID- 11115030 TI - Sudden infant death syndrome in South Australia 1968-97. Part 3: is bed sharing safe for infants? AB - OBJECTIVE: To examine the risk of death for bed-sharing infants. METHODOLOGY: All unexpected infant deaths occurring in South Australia between 1970 and 1997, occurring after the infant was put to rest and diagnosed by after death scene investigation and autopsy as sudden death infant syndrome, accidental death, or 'undetermined' were studied. RESULTS: Accidents were the most likely cause of death for 5% of infants who died in designated infant containers (cots, cradles, etc), 24% of those who were sharing a bed or couch, and 72% of those who were placed alone on a bed or couch. CONCLUSIONS: While bed sharing showed an increased risk of dying accidentally, when compared with infants sleeping in designated infant containers, the risk of accidental death in this study was even greater for infants left alone on adult beds or couches. PMID- 11115031 TI - Promoting secure attachment, maternal mood and child health in a vulnerable population: a randomized controlled trial. AB - OBJECTIVE: To evaluate the efficacy of an early home-based intervention on the quality of maternal-infant attachment, maternal mood and child health parameters in a cohort of vulnerable families. METHODOLOGY: A total of 181 families were recruited to the study in the immediate postnatal period on the basis of a self report questionnaire relating to known family vulnerability factors. Families were assigned randomly to intervention (90), or control (91) groups. The intervention group received a series of home visits from a child health nurse (weekly to 6 weeks, fortnightly to 3 months), with a subgroup receiving home based short-term dynamic therapy from a social worker. Parent/family function was assessed at inception and at 4 months by the Parenting Stress Index and the Edinburgh Post Natal Depression Scale. At 4 months the quality of the home environment was assessed, utilizing the Home Observation for Measurement of the Environment Inventory, as were child and family health parameters and satisfaction with the community child health service. RESULTS: At 4 month follow up, 160 families (80 intervention, 80 control) were available for assessment. The intervention improved family functioning at 4 months. All aspects of the home environment, including the quality of maternal-infant attachment and mothers' relationship with their child, were significantly enhanced. In particular, significant and positive differences were found in parenting with the intervention group feeling less restrictions imposed by the parenting role, greater sense of competence in parenting, greater acceptability of the child, and the child being more likely to provide positive reinforcement to the parent. Early differences in maternal mood were not maintained at 4 months. Various child health parameters were enhanced including immunization status, fewer parent reported injuries and bruising, and researcher confirmed lack of smoking in the house or around the infant. The families were consistently more satisfied with their community health service. CONCLUSIONS: This form of early home based intervention targeted to vulnerable families promotes an environment conducive for infant mental and general health and hence long-term psychological and physical well-being, and is highly valued by the families who receive it. PMID- 11115032 TI - A 'chronic disorder' health-care model for children with complex developmental disorders. AB - OBJECTIVES: To conceptualize, develop and evaluate a 'chronic disorder' clinical model of health services for children with 'low-severity' developmental disorders assessed and treated within a public Child Development Unit. METHODOLOGY: Concepts of family empowerment, child resiliency and the management of clinical complexity were explicitly incorporated into the service model and the clinical strategies in order to address long-term goals of prevention and health promotion. To explore the efficacy of this model, a telephone audit survey was conducted of parents of 42 children seen sequentially through this modified service. RESULTS: Parent data indicate a high level of satisfaction with the integrated, individualized assessment processes and effective transmission of information through both written reports and dedicated discussion visits. A treatment methodology based on parent chosen treatment goals for short and long time-frames was experienced as achievable and successful. CONCLUSIONS: These data suggest that the service goals of an integrated team process, parent empowerment and effective clinical collaborations may be achievable from a health care setting. A change in emphasis from remedial treatment to adaptation, health promotion and tertiary prevention is integral to these strategies. The concepts, clinical model and evaluation are presented to stimulate discussion around the question of what child health services are attempting to achieve for children with complex developmental disorders. PMID- 11115033 TI - Outcome of primary vesicoureteric reflux detected following fetal renal pelvic dilatation. AB - OBJECTIVE: Postnatal investigation of mild degrees of fetal hydronephrosis has allowed subsequent detection of infants with vesicoureteric reflux (VUR). This study was designed to provide short to medium term information on such infants who had primary VUR, the rates of renal damage and progression over time, the risk factors for such damage and to compare the characteristics of those who had mild dilatation of the fetal renal pelvis (4-9 mm) with those who had moderate severe dilatation (> or = 10 mm). METHODOLOGY: Since June 1989, infants whose antenatal sonography had identified a fetal renal pelvis with an anteroposterior diameter of > 4 mm were investigated postnatally with renal ultrasonography and micturating cystourethrogram (MCU), and placed on antimicrobial prophylaxis. Those with VUR received 99mTc-dimercaptosuccinic acid (DMSA) scintigraphy. Infants were followed until discharge based on resolution of VUR, surgery, or low grade VUR. A 5.5 year cohort between June 1989 and December 1994 formed the study population. A review of notes and clinical review (if still under follow up) was undertaken. Vesicoureteric reflux on MCU was regraded according to the International Classification, and reflux nephropathy on DMSA scans was regraded according to criteria proposed by Goldraich. Regression analysis was used to assess risk factors for renal damage. RESULTS: There were 69 infants (37 girls, 32 boys) who were identified with primary VUR, with 37/69 having bilateral reflux. Eight had a urinary tract infection during the follow-up period. There was a broad distribution of grades of reflux detected (Grades I-3, Grades II-23, Grades III-19, Grades IV - 17, Grades V-7). 99m-Tc-dimercaptosuccinic acid scans on 57/69 (83%) demonstrated renal damage in eight infants (14%). This was predominantly global contraction of function. No progression of renal damage was seen over 2-7 years. Regression analysis showed a strong association between Grades IV, V reflux and the presence of renal damage (P < 0.001). Review of the degrees of fetal renal pelvic dilatation showed that 60/69 infants were detected because of mild (4-9 mm) dilatation. The majority (43/60) had lower grades of reflux (Grades I, II, 3), but there was no obvious cut-off between 4 and 9 mm that could predict high grade VUR (Grades IV, V). CONCLUSIONS: The use of 4 mm to define an abnormal fetal renal pelvis allows a much larger group of infants with high grade primary VUR to be detected than if a higher cut-off measurement is used. Although it also detects many more infants with low grade primary VUR, there is no obvious cut-off point at which this effect predominates. Progressive renal damage was not seen in follow up of up to 7 years of age. Renal damage on DMSA scanning in this group is almost exclusively a pattern of global contraction of function. The presence of high-grade VUR appears to be the only important factor in predicting the presence of renal damage. PMID- 11115034 TI - Ascertainment of birth defects: the effect on completeness of adding a new source of data. AB - BACKGROUND: The Western Australian (WA) Birth Defects Registry aims for complete ascertainment of birth defects in WA, but the proportions of birth defects in rural areas and in Aboriginal children are lower than in metropolitan and non Aboriginal children. The effect on ascertainment of adding data from the Rural Paediatric Service (RPS) was investigated. METHOD: A file of all cases of birth defects for children born 1980-1997 and recorded on the RPS database was linked to the Registry. RESULTS: The addition of this new data source had little effect on the overall prevalence of birth defects (an increase from 5.38 to 5.41%). There was a slightly greater effect on the prevalence of birth defects in rural residents (4.67%-4.76%) and Aboriginal children (4.55-4.78%), although the prevalence for each of these groups is still less than for metropolitan residents and non-Aboriginal infants, respectively. All major categories of birth defects were represented in the new cases and, in general, their addition made little difference to the prevalence of each category. The exception was fetal alcohol syndrome, which increased from 0.13 per 1000 to 0.18 per 1000 once the 21 new cases from the RPS were added. CONCLUSION: Complete ascertainment of birth defects is important in developing and evaluating preventive programs, and in investigating clusters of birth defects. PMID- 11115035 TI - Parental attitudes to health of children in child-care centres and options when children are ill. AB - OBJECTIVE: To determine parental attitudes regarding the health of children attending child-care centres, to explore concerns when children who normally attend child care are ill, and to investigate options in these circumstances. METHODOLOGY: Focus groups conducted with parents whose children attended child care centres. Ten focus groups were conducted. RESULTS: Many parents encountered difficulty when children who normally attend child care were ill and there was a lack of options for care. Parents were concerned about the spread of infections among children but considered that there were also many health and other advantages for their children in attending child care. Child-care centres were perceived as providing a valuable support role for families. CONCLUSIONS: Many parents lack adequate options for care when their children are ill. Parents' concerns regarding health in child care are important in policy decisions regarding the health of children in child care, and the development of alternative care services for children. PMID- 11115036 TI - A randomized trial of enteral feeding volumes in infants born before 30 weeks' gestation. AB - OBJECTIVE: To compare the effect of two volumes of enteral feeds on postnatal growth in infants born before 30 weeks gestation. METHODOLOGY: Fifty-four infants, less than 30 weeks gestational age, who reached full enteral feeds were randomized to remain on 150 mL/kg per day (150 group) or increase to 200 mL/kg per day (200 group). The primary outcome measure was growth at 35 weeks corrected gestational age (CGA). RESULTS: There were no statistically significant differences in demographic or clinical parameters between the study groups at commencement of the study, although there was a trend for infants in the 150 group to be lighter (895 g vs 1020 g, P = 0.27). Milk intakes were increased in 43% of the infants in the 150 group, whereas 54% of the infants in the 200 group required reduced intakes. Infants in the 200 group had greater daily weight gains (16.7 g/kg per day vs 15.2 g/kg per day, P = 0.047) and at 35 weeks CGA were heavier (2020 g vs 1885 g, P = 0.014) and had a greater arm fat area (282 mm2 vs 218 mm2, P = 0.009). There was no difference in length or head circumference at 35 weeks CGA, and no difference in any growth parameter at 1 year of age. Morbidity was not different between the groups. CONCLUSIONS: The individual milk volume requirements for adequate weight gain without significant adverse effects vary between 150 and 200 mL/kg per day in extremely premature infants. For many infants in both groups, the assigned target volume was not appropriate. Increased milk intakes (and therefore higher caloric and mineral intakes) are associated with increased daily weight gains and a greater weight at 35 weeks CGA. The weight gain may be due to an increase in fat deposition. PMID- 11115037 TI - Is acid base determination an accurate predictor of pyloric stenosis? AB - OBJECTIVE: To determine if acid base status predicts which vomiting patients have pyloric stenosis. DESIGN: Retrospective chart review. SETTING: Tertiary paediatric hospital. METHODOLOGY: We compared the clinical and biochemical parameters of 100 patients with a discharge diagnosis of pyloric stenosis and 84 patients of a similar age who presented to the emergency department with vomiting and who had an acid base determination. Patients were included from January 1995 to January 1997. Clinical correlates consisted of age, duration of vomiting, weight loss, gestation, and family history of pyloric stenosis. Biochemical correlates were pH, bicarbonate, base excess (BE), chloride, potassium, and sodium. RESULTS: Independent variables of significance were pH, BE, chloride, bicarbonate, potassium, weight loss (all of which had a P value < 0.0001), and sex (P = 0.006). Each variable was placed in a logistic regression equation with pyloric stenosis being the dominant variable. Variables of significance were pH (P = 0.0001), BE (P = 0.0001), and chloride (P = 0.009). A model for predicting pyloric stenosis using these variables was then created with pH > 7.45, chloride < 98, and BE > +3, with a positive predictive value of 88%. CONCLUSION: Acid base determination is a useful screening tool when considering pyloric stenosis. This model now needs to be validated on a prospective series of patients with vomiting. PMID- 11115038 TI - Non-accidental fractures in infants: risk of further abuse. AB - OBJECTIVES: To look for features of non-accidental fractures in infants aged under I year and assess the risk of subsequent morbidity and mortality. METHODOLOGY: A retrospective analysis of 99 children aged under 1 year who presented to the Mater Children's Hospital, Brisbane, between January 1990 and December 1993, and were found to have a fracture. The 99 infants were divided into non-accidental and accidental groups. Comparison was made between the two groups for age, sex and type of fracture. Deaths, subsequent injuries and child protection notifications until March 1997 were compared between groups. RESULTS: Of the 99 infants with fracture (64 males, 35 female), the skull and femur were the most prevalent sites of fracture. Twenty-six infants had fractures assessed as non-accidental. This group was younger but did not differ significantly in gender or site of fracture. Infants aged under 4 months had a significantly greater risk of their fracture being non-accidental (P = 0.0007). Subsequent substantiated child protection notifications occurred in nine of the non accidental group and in one of the accidental group (P = 0.000001). There was no significant difference in the rate of subsequent notifications between those infants with abuse who were removed from their carers and those not removed. Subsequent injuries presenting to hospital occurred in 17 of the accidental group and three of the non-accidental group (P = 0.20). There were no deaths. CONCLUSION: Infants aged under 1 year with fractures have a high prevalence of abuse. The risk of abuse as cause for the fracture is greater in those aged under 4 months. Infants with non-accidental fractures have a high risk of further abuse even with intervention. PMID- 11115039 TI - Choice of formula and human milk supplement for preterm infants in Australia. AB - Human milk (HM) is considered to be the optimal feed of choice for neonates, however, for preterm infants, HM fortifiers are often added to increase growth. If HM is unavailable, preterm formula is the next best option for preterm infants. Choosing which fortifier, if any, to use or which formula can be confusing. In this paper, the composition of milk feeds available in Australia and New Zealand is reviewed with the aim of assisting paediatricians to decide which feed is most appropriate for their patients. PMID- 11115040 TI - A taste of honey. AB - Infantile botulism classically presents with a triad of clinical features: Bulbar palsies (slow/absent pupil response) Alert Absent fever Other common features are: Constipation, ptosis and poor feeding The diagnosis is a clinical one, confirmed by EMG and by testing stool for the organism, C. botulinum, or its toxin. Parents should be advised not to dip pacifiers in honey. In future Botulinum Immune Globulin may be available and should be considered if further trials are published showing benefit. Nosocomial infections, such as RSV, are important and should be carefully avoided, particularly in patients recovering from botulism. PMID- 11115041 TI - Infantile-onset megalencephalic leucoencephalopathy in two siblings. AB - Infantile-onset megalencephalic leucoencephalopathy (IML) is a recently recognized autosomal recessive white matter disorder. Unlike other megalencephalic leucoencephalopathies, in patients with IML a mild clinical course, a slowly progressive delay in motor development and mild mental deterioration are typical. We report on two affected siblings who have typical clinical and radiological findings of IML. Cranial magnetic resonance imaging showed involvement of the capsula externa, extrema and interna, nucleus dentatus, crus cerebri, periventricular and subcortical white matter. In addition, bilateral cystic changes were determined predominantly in the temporal lobes. There were no clear biochemical or metabolic disturbances. In the present paper, we discuss the clinical and neuroimaging findings of IML. PMID- 11115042 TI - Encephalocraniocutaneous lipomatosis (Fishman syndrome): a rare neurocutaneous syndrome. AB - Encephalocraniocutaneous lipomatosis (ECCL) is a rare congenital neurocutaneous syndrome comprising unilateral cranial lipomas, lipodermoids of the eye and brain abnormalities. A 3-year-old boy who presented at birth with a scalp lipoma and an ipsilateral epibulbar lipodermoid is described. Infantile spasms developed at 9 months of age and cerebral imaging showed thickened and calcified cortex at the right occiput and hemiatrophy of the right hemisphere. These features were consistent with ECCL. Most children with ECCL have significant developmental delay, but we have found that control of seizures was associated with a significant improvement in developmental outcome. PMID- 11115043 TI - Meningococcal pericarditis in a 2-year-old child: reactive or infectious? AB - Pericarditis is an uncommon manifestation of infection of Neisseria meningitidis. Pericarditis may be caused by direct invasion or immune-complex-mediated (reactive) inflammation. We outline the case of a two-year-old girl with probable reactive pericarditis, review the cases reported in the English literature since 1966 and discuss the pathogenesis of meningococcal pericarditis. PMID- 11115044 TI - Hansenula anomala infection in a neonate. AB - We present an unusual neonatal fungal infection, Hansenula anomala in a very low birthweight infant who underwent abdominal surgery for an omphalocele. Despite treatment with adequate doses of amphotericin B, the yeast continued to grow from the blood culture, and was only eradicated with the use of oral ketoconazole. PMID- 11115045 TI - Bed wetters: not just wet blankets. PMID- 11115046 TI - How safe is hepatitis B vaccination at birth? PMID- 11115047 TI - Sudden infant death syndrome: is the winter prepoderance due to some infants having the head covered? PMID- 11115048 TI - A broader vision for contemporary paediatrics: implications for the paediatric workforce. PMID- 11115049 TI - Serum cytokine values and fatigue in chronic hepatitis C infection. AB - Patients with chronic hepatitis C infection often experience fatigue. In many clinical situations, an association between fatigue and altered serum cytokine levels has been found. Altered cytokine levels in patients with hepatitis C have not shown a correlation with the degree of serum transaminase elevation or pathological change on liver biopsy. The aim of our study was to examine whether there was an association between abnormal serum cytokine levels and fatigue in patients with compensated chronic hepatitis C. Patients referred to a tertiary care hepatology clinic who were hepatitis C antibody positive and who had elevated alanine aminotransferase (ALT) levels were eligible for entry into the study. A control group was also included. Subjects in both groups who had characteristics other than hepatitis C that were known to alter cytokine values and/or cause fatigue were excluded. Patients completed a validated questionnaire to determine their fatigue severity score (FSS). Bioassays were used to measure interleukin (IL)-1, IL-6 and tumour necrosis factor-alpha (TNF-alpha) levels in early morning serum samples taken from patients and controls. Altered cytokine values were defined as those more than two standard deviations above the mean control value. Data was analysed using SPSS, version 8.01. Of the 78 patients with chronic hepatitis C who participated in the study, 19 (24%), 24 (30%) and 45 (56%) had elevated levels of IL-1, IL-6 and TNF-alpha, respectively, compared with only two (6%) of the control group who had elevation of any of the three cytokines. No correlation was found between the FSS and serum cytokine levels, when analysed singly or in combination, in patients with chronic hepatitis C. Hence, alteration in early morning serum levels of IL-1, IL-6 and TNF-alpha in patients with chronic hepatitis C infection and elevated ALT levels bear no correlation with the symptom of fatigue. PMID- 11115050 TI - Retreatment for 24 vs 48 weeks with interferon-alpha2b plus ribavirin of chronic hepatitis C patients who relapsed or did not respond to interferon alone. AB - We assessed the efficacy of interferon (IFN) plus ribavirin over 24 or 48 weeks for the retreatment of patients with chronic hepatitis C who had relapsed or did not respond to a previous course of IFN. One-hundred and twenty patients (69 non responders and 51 relapsers) were randomly assigned to receive IFN-alpha2b (3 million units thrice weekly) plus ribavirin (1,000-1,200 mg per day) for 24 weeks (group A: 58 patients) or 48 weeks (group B: 62 patients). Treatment was discontinued at week 12 if the alanine aminotransferase (ALT) level remained elevated. The rate of sustained response was 15.5% in group A and 37.1% in group B (P = 0.013). Relapsers treated for 48 weeks had a sustained response rate of 66.6% compared with a sustained response rate of only 25% in those treated for 24 weeks (P = 0.004). Moreover, a sustained response was seen in 14.3% of non responders treated for 48 weeks and in 8.8% of those treated for 24 weeks (P = 0.71). Fifty-three per cent of patients with a normal ALT level and undetectable hepatitis C virus (HCV) RNA at week 12 had a sustained response compared with 14% of those who were HCV RNA positive at week 12 (P < 0.001). Independent predictive factors of sustained response were: therapy for 48 weeks (P = 0.0026), relapse after IFN treatment (P = 0.0006), loss of HCV RNA at week 12 (P = 0.0008) and HCV genotype non-1 (P = 0.024). Hence, in patients with chronic hepatitis C who failed to respond to a previous course of IFN monotherapy, combination therapy with IFN plus ribavirin for 48 weeks seems to be more effective than IFN plus ribavirin for 24 weeks. PMID- 11115051 TI - HCV RNA levels during therapy with amantadine in addition to interferon and ribavirin in chronic hepatitis C patients with previous nonresponse or response/relapse to interferon and ribavirin. AB - Interferon (IFN) alpha in combination with ribavirin (RIB) is standard therapy for patients with chronic hepatitis C virus (HCV) infection. However, many patients do not respond with sustained HCV clearance to this therapy. At present, no accepted treatment strategy exists for these patients. Recent preliminary data have suggested that amantadine (AMA) is effective against HCV infection. In a pilot study, we treated 13 nonresponders and 10 response/ relapsers to previous IFN/RIB therapy with AMA 200 mg per day in combination with IFN 3 MU thrice weekly, and RIB 1000 mg per day for 24 weeks, with a 24-week follow-up period after end-of-treatment. At the end-of-treatment, 1 previous nonresponder and 5 previous response/relapsers were HCV RNA negative. At the end of follow-up, only 1 previous response/relapser remained HCV RNA negative and had a sustained response. During therapy, serum HCV RNA became undetectable in 4 previous nonresponders, of whom 3 had a breakthrough at week 24. Twenty-one patients continued therapy without dose reductions. One patient discontinued therapy prematurely due to sleeping disturbances, and another patient was withdrawn from therapy due to heavy alcohol intake. We conclude that the addition of AMA to IFN and RIB was well tolerated but had little, if any, impact on HCV RNA eradication in nonresponders or response/relapsers to previous IFN/RIB combination therapy. PMID- 11115052 TI - Long-term follow-up of sustained responders to interferon therapy, in patients with chronic hepatitis C. AB - Interferon (IFN) therapy has been proven to induce the normalization of serum alanine aminotransferase (ALT) levels and to eradicate the hepatitis C virus (HCV) in some patients with chronic hepatitis C, and these patients are usually defined as 'sustained responders'. However, there have been some reports of hepatocellular carcinoma (HCC) in these patients, and the development of HCC remains life-threatening in patients who clear HCV. We analysed the long-term prognoses of patients with chronic hepatitis C in whom HCV was eradicated with IFN. We investigated 392 sustained responders to IFN therapy, from 1,277 patients with chronic HCV infection who received IFN treatment at one of our institutions between April 1989 and March 1999. We analysed the medical records and looked for the development of HCC. About 30% of the sustained responders had been lost to follow-up 3 years after the end of IFN therapy, and the follow-up rate of sustained responders was significantly lower than that of non-sustained responders (P < 0.0001). HCC were found in eight patients: in seven patients HCC developed within 5 years after completion of IFN therapy; but in one patient, a single HCC less than 3 cm in diameter was detected between 7 and 8 years after completion of IFN. Of the five patients who had regular medical follow-up, the HCC was solitary, and the patients survived without any evidence of recurrence. Of the three patients who had not been followed-up, two died from HCC and HCC recurred in the third. These results suggest that HCC can develop in sustained responders and that sustained responders should be followed-up closely after completion of IFN so that HCC may be detected at an early stage. The optimal duration of the follow-up period of the sustained responders remains unclear. Additional prospective studies are required in order to establish an appropriate follow-up protocol for sustained responders to IFN. PMID- 11115053 TI - The HCV National Register: towards informing the natural history of hepatitis C infection in the UK. AB - The aim of this paper is to describe the development of a national hepatitis C register and the completeness of the data it contains. This is a descriptive report of the structure and function of the register, including case definitions, registration and follow-up procedures, and methods used to maximize data quality and to obtain comparative data sources. The register contains data on HCV infected individuals who acquired their infections on a known date and by a known route; to date all are transfusion recipients identified during the UK lookback exercise, who tested positive or indeterminate for anti-HCV after receiving 'infected' blood issued before the introduction of routine testing of the blood supply for anti-HCV. By 31 December 1999, 871 (87%) of 996 eligible transfusion recipients had been registered, and 984 (99%) flagged in the NHS Central Registers. Registered patients had been infected for an average of 11.1 years (SEM 0.1); around half were being cared for by clinicians with a specialist interest in liver disease. Except for the information on tobacco use, current alcohol use, and hepatitis B status, data were more than 80% complete, and for most variables, more than 90% complete. The consistency of data abstraction was found to be 98% (SEM 0.5). In conclusion, the Register contains high quality anonymised data on one of the largest cohorts of individuals with HCV infections acquired on a known date and by a known route. It could serve as a model for other chronic disease registers; developers may find the structure, design, and methodological issues addressed useful. PMID- 11115054 TI - Lamivudine-high dose interferon combination therapy for chronic hepatitis B patients co-infected with the hepatitis D virus. AB - Currently, the best option for patients with hepatitis delta is interferon alpha therapy for at least one year. To evaluate the effect of the combination lamivudine-high-dose interferon alpha therapy, we first treated eight patients with chronic hepatitis delta infection with lamivudine for at least 24 weeks; then lamivudine was combined with a high dose of interferon alpha followed by a regular dose (9 MU tiw). Follow-up was 12 weeks. Virological, biochemical and histological features were evaluated for response to therapy. At baseline, all patients were HBsAg positive in serum and HDV RNA-(PCR)positive in plasma; HBV DNA was undetectable with the Digene Hybrid Capture assay (limit of detection 1.5 x 10(6) geq ml-(1)) in all cases. Transaminases were elevated in all patients; median ALT 68 (range 48-143) IU l(1). Seven of eight patients completed the course; one patient with a pre-existing sickle cell trait was withdrawn from the trial due to the development of a nephrotic syndrome. The HBsAg-concentration in serum decreased in two out of seven patients (29%). However, there was no significant decrease in the HBsAg-concentration in serum during treatment (median 3654 PEU l(-1) (range 548-7,684) to 5300 PEU l(-1) (range 168-19,639)). The drop of HDV RNA in plasma from baseline during treatment was not significant. Decrease of HDV RNA was observed in three out of seven patients (43%) (median 10(5) geq ml(-1); range 10(3)-10(6) to median 10(3) geq ml(-1); range 10(2)-10(7)). Serum ALT did not change as reflected by a median of 68 IU l(-1) (range 48-143) at start of therapy to 63 IU l(-1) (range 20-171) at the end of therapy. At the end of treatment transaminases had normalised in one patient and decreased in three other patients (improvement in 57%). However, three of these four patients showed a rebound after withdrawal of therapy. The Histology Activity Index (HAI) indicated a drop from a median score of 7 (range 5-9) at baseline to 5 (range 3 8) at the end of treatment, but an increase in fibrosis from a median grade of 2 (range 1-3) at baseline to 3 (range 1-4) at the end of treatment was observed. In conclusion, this study does not yield support for the combination of an HBV suppressor and 16 weeks of high-dose interferon for therapy aimed at eradicating the hepatitis delta virus. PMID- 11115055 TI - Haemophilic patients with hepatitis C have higher viral load compared to other well-defined patient groups. AB - Comparison of hepatitis C viral load between different patient populations has been hampered by the use of different technology in individual studies. We had the impression that haemophilic (HAEM) patients had a higher serum load of hepatitis C virus (HCV) compared to other HCV-infected patients. We therefore studied viral load and genotypes in active illicit drug users (IDU), HAEM patients and patients with post-transfusion hepatitis (PTH). The study comprises 225 HCV-RNA positive patients, 117 IDU, 60 HAEM patients and 48 PTH patients. All patients were anti-HIV negative. HCV-RNA was measured with a quantitative reverse transcription polymerase chain reaction (RT-PCR) method, HCV-genotypes were determined with genotype specific primers in RT-PCR in 221 patients. Four patients could not be genotyped with our assay and were excluded. Overall viral load was higher in genotypes 1 and 2 compared to genotype 3, median values of HCV RNA were 1,400 x 10(3) geq ml(-1), 2,700 x 10(3) geq ml(-1) and 270 x 10(3) geq ml(-1), respectively. HAEM patients had significantly higher viral load for both genotypes 1 and 3 compared to the IDU and PTH patients. In a multiple linear regression model HCV-RNA viral load was independently associated with HAEM and genotype, but not to age, gender or disease duration. In conclusion, HAEM patients have higher viral load than IDU and PTH patients. The reason for this is unknown, but it may be due to host factors or mode of transmission with multiple inoculations. PMID- 11115056 TI - Presence of TTV DNA in serum, liver and peripheral blood mononuclear cells from patients with chronic hepatitis. AB - The main site of TT virus (TTV) replication remains unknown. Therefore, we have studied the presence and titres of TTV DNA in paired serum, liver and PBMC samples from 50 patients with liver disease (32 with chronic hepatitis B or C, seven with cryptogenic hepatitis and 11 with nonviral liver disease) were included. TTV DNA was analysed by polymerase chain reaction (PCR) using primers from the open reading frame 1 (ORF 1) and from the untranslated region (UTR) and titres were semiquantified by PCR using an external standard. TTV DNA was detected in 26% of serum, 24% of liver and 14% of PBMC samples with ORF 1 primers. When UTR primers were used, 70% of serum and liver samples and 64% of PBMC were TTV DNA positive. No differences between TTV positive and negative patients were found regarding epidemiological or biochemical parameters. Trypsin treatment and fluorescent in situ hybridization confirm the intracellular location of TTV in PBMC. The mean of TTV DNA titres was statistically higher in liver than in serum or PBMC. TTV titres in serum correlated with those in PBMC but not with those in liver. In conclusion, although the liver seems to be the main site for TTV replication, this virus is also able to infect PBMC. PMID- 11115057 TI - Sequence variations of precore/core and precore promoter regions of hepatitis B virus in patients with or without viral reactivation during cytotoxic chemotherapy. AB - Reactivation of the hepatitis B virus (HBV) is a well-described complication among cancer patients undergoing cytotoxic chemotherapy. Mutations in the preC/C and the preC promoter regions of HBV have been reported in some patients who developed this condition. A G-to-A mutation at nt 1896 in the preC/C region (HBeAg negative/ anti-HBe positive) has been associated with more severe liver disease than that caused by wild type virus. In addition, it has been suggested that patients with these mutations may be more likely to reactivate than those with the wild type virus. Whether or not such mutations were present before the commencement of or developed during the course of cytotoxic chemotherapy is not known. In this study, 28 cancer patients (consisting of 14 consecutive patients who developed HBV reactivation and another 14 who had no reactivation during cytotoxic chemotherapy) are reported. The objectives were firstly, to determine the prechemotherapy HBeAg status and nucleotide sequences of the preC/C and preC promoter regions of HBV in order to determine if these parameters affected the rate of reactivation, and secondly, for those who developed reactivation, to determine whether the mutations were present before chemotherapy or developed during, possibly as a result of, cytotoxic chemotherapy. HBV DNA was amplified by PCR and nucleotide sequencing performed on samples taken prior to chemotherapy and at the time of reactivation. Results revealed that 16 of the 28 patients were HBeAg negative/anti-HBe positive. Of these 16, four (57%) of the seven patients who had nt 1896 mutation, but only one (17%) of the six who had the wild type HBV genome, developed reactivation. Three had no detectable HBV DNA. In the majority of cases, the type of virus, i.e. wild/mutant at preC/C, that was detected during the reactivation was identical to that detected in the pretreatment samples. With respect to the preC promoter region, the two commonest mutations detected were at nt 1762 (A to T) and nt 1764 (G to A). When this region was translated into amino acid sequences, stop codons leading to truncated X protein at carboxyl terminus were found in four patients, three of whom developed HBV reactivation. We conclude that chronic HBV carriers who are HBeAg negative/anti-HBe positive with nt 1896 mutation (G to A) may be more likely to develop HBV reactivation during cytotoxic chemotherapy than those with the wild type virus. Cytotoxic chemotherapy does not appear to select out mutant HBV, or to be consistently mutagenic in patients who develop HBV reactivation. The occurrence of stop codons in the amino acid sequences of the X protein in three patients who developed HBV reactivation, including one who was detected only at the time of reactivation, is of particular interest, as such mutant viruses remain replication competent. PMID- 11115058 TI - Sequence, expression and reconstitution of an HCV genome from a British isolate derived from a single blood donation. AB - Morphological analysis of hepatitis C virus and development of antiviral drugs to eradicate this agent have been seriously hampered by the low viraemias observed during natural infection and the unavailability of a cell culture system for virus propagation. Recently a low-grade hepatitis has been reported in chimpanzees after intrahepatic transfection of full-length synthetic HCV RNA and successful infections shown to be critically dependent on the integrity and genetic homogeneity of the reconstituted clone. In this study we describe and characterize a full HCV RNA sequence derived from a case of chronic sporadic hepatitis. The genotype was shown to be 1a with a low level of intraclonal sequence heterogeneity, and processing of both structural and nonstructural proteins has been documented. The assembly of the full genome has also been achieved. The low level of intraclonal variation observed may reflect infection with a single isolate and the fact that cloning was performed on virus obtained from a single blood donation makes this clone a good candidate for future in vivo and in vitro transfection studies. PMID- 11115059 TI - Retinal vein thrombosis associated with chronic hepatitis C: a case series and review of the literature. AB - The role of procoagulant autoantibodies in hepatitis C virus (HCV) infection is unclear. Three individuals with HCV infection and a unique genetic hypercoagulable state developed retinal vein thrombosis (RVT) in association with interferon-alpha (IFN-alpha) therapy. It is probable that a combination of active HCV infection in a genetically susceptible individual receiving IFN-alpha accounted for the observed RVT. PMID- 11115060 TI - Relationship between nonphenacetin combined analgesics and nephropathy: a review. Ad Hoc Committee of the International Study Group on Analgesics and Nephropathy. AB - BACKGROUND: The debate on the association between nonphenacetin-containing combined analgesics and renal disease has lasted for several years. METHOD: A peer review committee of scientists, selected jointly by the regulatory authorities of Germany, Switzerland, and Austria and the pharmaceutical industry was asked to critically review data on the relationship between nonphenacetin combined analgesics and nephropathy. RESULTS: The committee regarded epidemiologic evidence on nonphenacetin combined analgesics as inconclusive because of sparse information and substantial methodological problems. The committee also noted that a diagnosis of analgesic-associated nephropathy (AAN) in clinical practice usually depends on information about exposure before or in the early stages of the disease and is seldom accompanied by specific histologic evidence. The morphologic finding of papillary calcification can arise from other conditions and is not specific for AAN. For these reasons, the identification criteria for AAN should be reappraised with scientific methods to validate the diagnostic procedure. In the limited amount of experimental pharmacological data in humans and animals, the committee found no convincing evidence to confirm or refute the hypothesis that nonphenacetin combined analgesics are more nephrotoxic than single formulations. For caffeine taken with combined analgesics, the currently available information is not sufficient to postulate a harmful toxicological effect. CONCLUSION: The committee's two main conclusions were that sufficient evidence is absent to associate nonphenacetin combined analgesics with nephropathy and that new studies should be done to provide appropriate data for resolving the question. PMID- 11115061 TI - Importance of quantitative genetic variations in the etiology of hypertension. AB - Recent progress has been remarkable in identifying mutations which cause diseases (mostly uncommon) that are inherited simply. Unfortunately, the common diseases of humankind with a strong genetic component, such as those affecting cardiovascular function, have proved less tractable. Their etiology is complex with substantial environmental components and strong indications that multiple genes are implicated. In this article, we consider the genetic etiology of essential hypertension. After presenting the distribution of blood pressures in the population, we propose the hypothesis that essential hypertension is the consequence of different combinations of genetic variations that are individually of little consequence. The candidate gene approach to finding relevant genes is exemplified by studies that identified potentially causative variations associated with quantitative differences in the expression of the angiotensinogen gene (AGT). Experiments to test causation directly are possible in mice, and we describe their use to establish that blood pressures are indeed altered by genetic changes in AGT expression. Tests of differences in expression of the genes coding for the angiotensin-converting enzyme (ACE) and for the natriuretic peptide receptor A are also considered, and we provide a tabulation of all comparable experiments in mice. Computer simulations are presented that resolve the paradoxical finding that while ACE inhibitors are effective, genetic variations in the expression of the ACE gene do not affect blood pressure. We emphasize the usefulness of studying animals heterozygous for an inactivating mutation and a wild-type allele, and briefly discuss a way of establishing causative links between complex phenotypes and single nucleotide polymorphisms. PMID- 11115062 TI - Genetic evidence for a novel gene(s) involved in urogenital development on 10q26. AB - BACKGROUND: Although the frequent association between distal 10q monosomy and urogenital anomalies suggests the presence of a gene(s) for urogenital development on distal 10q, molecular deletion mapping has not been performed for the putative gene(s). In this study, we examined genotype-phenotype correlations in patients with distal 10q monosomy. METHODS: This study consisted of six karyotypic males (cases 1 through 6) and four karyotypic females (cases 7 through 10) with 10q26 monosomy. Cases 3 through 5 and 7 through 10 had urinary anomalies such as vesicoureteral reflux and hypoplastic kidney, and cases 1 through 6, 8, and 9 exhibited genital anomalies such as micropenis, hypospadias, cryptorchidism, and hypoplastic labia majora. Fluorescence in situ hybridization (FISH) for 10q telomere, whole chromosome 10 painting, and microsatellite analysis for 35 loci on distal 10q were performed in cases 1 through 8. RESULTS: FISH and whole chromosome painting confirmed distal 10q monosomy in cases 1 through 8. Microsatellite analysis revealed that hemizygosity for the region distal to D10S186 was shared by cases with urinary anomalies and that for the region distal to D10S1248 was common to cases with genital anomalies. Furthermore, it was indicated that PAX2, GFRA1, and EMX2 on distal 10q, in which the deletions could affect urinary and/or genital development, were present in two copies in cases 1 through 8. CONCLUSIONS: The results suggest that a novel gene(s) for urinary development and that for genital development reside in the approximately 20 cM region distal to D10S186 and in the approximately 10 cM region distal to D10S1248, respectively, although it remains to be determined whether the two types of genes are identical or different. PMID- 11115063 TI - Postnatal development and progression of renal dysplasia in cyclooxygenase-2 null mice. AB - BACKGROUND: Genetic ablation of cyclooxygenase-2 (COX-2) resulted in cystic renal dysplasia and early death in adult mice. The ontologic development of the renal pathology and the biochemical and physiological abnormalities associated with the dysplasia are unknown. METHODS: Mice homozygous for a targeted deletion of COX-2 (-/-) were compared with wild-type littermates (+/+). Somatic and kidney growth and renal histology were studied at the day of birth and at a number of postnatal ages. Systolic blood pressure, urinalysis, urine osmolality, serum and urine chemistries, and inulin clearance were evaluated in adult animals. RESULTS: Beginning at postnatal day 10 (PN10), kidney growth was suppressed in -/- animals, while somatic growth and heart growth were unaffected. By PN10, -/- kidneys had thin nephrogenic cortexes and crowded, small, subcapsular glomeruli. The pathology increased with age with progressive outer cortical dysplasia, cystic subcapsular glomeruli, loss of proximal tubular mass, and tubular atrophy and cyst formation. Adult -/- kidneys had profound diffuse tubular cyst formation, outer cortical glomerular hypoplasia and periglomerular fibrosis, inner cortical nephron hypertrophy, and diffuse interstitial fibrosis. The glomerular filtration rate was reduced by more than 50% in -/- animals (6.82 +/- 0.65 mL/min/kg) compared with wild-type controls (14.7 +/- 1.01 mL/min/kg, P < 0. 001). Plasma blood urea nitrogen and creatinine were elevated in null animals compared with controls. Blood pressure, urinalysis, urine osmolality, and other plasma chemistries were unaffected by the deletion of COX-2. CONCLUSIONS: Deficiency of COX-2 results in progressive and specific renal architectural disruption and functional deterioration beginning in the final phases of nephrogenesis. Tissue-specific and time-dependent expression of COX-2 appears necessary for normal postnatal renal development and the maintenance of normal renal architecture and function. PMID- 11115064 TI - Antibody to transforming growth factor-beta ameliorates tubular apoptosis in unilateral ureteral obstruction. AB - BACKGROUND: Unilateral ureteral obstruction (UUO) is characterized by progressive renal atrophy, renal interstitial fibrosis, an increase in renal transforming growth factor-beta (TGF-beta), and renal tubular apoptosis. The present study was undertaken to determine the effect of a monoclonal antibody to TGF-beta (1D11) in UUO. METHODS: Mechanical stretch was applied to tubular epithelial cells (NRK 52E) by a computer-assisted system. Three doses of 1D11 (either 0.5, 2, or 4 mg/rat) were administered to rats one day prior to UUO and every two days thereafter, and kidneys were harvested at day 13. Fibrosis was assessed by measuring tissue hydroxyproline and mRNA for collagen and fibronectin. Apoptosis was assessed with the terminal deoxy transferase uridine triphosphate nick end labeling assay. TGF-beta levels were determined by bioassay. Western blot and immunostaining were used to identify proliferating cell nuclear antigen (PCNA), p53, bcl-2, and inducible nitric oxide synthase (iNOS). RESULTS: Stretch significantly induced apoptosis in NRK-52E cells, which was accompanied by an increased release of TGF-beta; 1D11 (10 microg/mL) totally inhibited stretch induced apoptosis. Control obstructed kidney contained 20-fold higher TGF-beta as compared with its unobstructed kidney; 1D11 neutralized tissue TGF-beta of the obstructed kidney. Control obstructed kidney exhibited significantly more fibrosis and tubular apoptosis than its unobstructed counterpart, which was blunted by 1D11. In contrast, 1D11 significantly increased tubular proliferation. p53 immunostaining was localized to renal tubular nuclei of control obstructed kidney and was diminished by 1D11. In contrast, bcl-2 was up-regulated in the 1D11-treated obstructed kidney. Total NOS activity and iNOS activity of the obstructed kidney were increased by 1D11 treatment. CONCLUSION: The present study strongly suggests that an antibody to TGF-beta is a promising agent to prevent renal tubular fibrosis and apoptosis in UUO. PMID- 11115065 TI - Dopamine induces the expression of heme oxygenase-1 by human endothelial cells in vitro. AB - BACKGROUND: In a retrospective study of the kidney transplantations performed at our institution, we found that the administration of dopamine (DA) to the organ donors resulted in a significant improvement of long-term organ survival of the retrieved kidneys. To study the mechanisms underlying the organ protection associated with the administration of DA prior to transplantation, we questioned whether DA induces the antioxidative enzyme heme oxygenase-1 (HO-1) in cultured endothelial cells. METHODS: Human umbilical vein endothelial cells (HUVECs) in culture were incubated with varying concentrations of DA for different time periods. Cells were subsequently assessed for the expression of HO-1 by Western blot and semiquantitative reverse transcription-polymerase chain reaction (RT PCR). RESULTS: The presence of DA resulted in a dose- and time-dependent up regulation of HO-1 both on RNA and protein level, whereas HO-1 was barely detectable under basal conditions. RT-PCR indicated the increased presence of HO 1 messenger RNA after 2 hours of incubation with DA, which peaked after 24 hours. The induction of HO-1 antigen was detectable after eight hours, as visualized by Western blot analysis. The addition of the antioxidant agents ascorbic acid and N acetyl-cysteine both lead to dose-dependent inhibition of DA-mediated HO-1 induction. DA-mediated up-regulation of HO-1 was not influenced by the addition of either the D2-receptor antagonist haloperidol or the D1-receptor antagonist SCH 23390. CONCLUSION: We conclude that DA induces the expression of the protective enzyme HO-1 in cultured endothelial cells by an oxidative mechanism. These findings may explain the beneficial effect of DA administration to kidney donors and indicate the potential role of DA in organ preconditioning. PMID- 11115066 TI - Early diabetes mellitus stimulates proximal tubule renin mRNA expression in the rat. AB - BACKGROUND: Enhanced intrarenal angiotensin II (Ang II) activity may contribute to diabetic nephropathy. The proximal tubule is a proposed site of significant intrarenal Ang II production. We determined the effect of early diabetes on mRNA expression of components of the proximal tubule renin-angiotensin system. METHODS: Three groups of male Sprague-Dawley rats were studied after two weeks: (1) control (C), (2) streptozotocin-induced diabetes (STZ), and (3) STZ-induced diabetes, with normoglycemia maintained by insulin implants (STZ-I). Competitive reverse transcription-polymerase chain reaction was used to assay mRNA for renin, angiotensinogen, and angiotensin-converting enzyme in suspensions of proximal tubules; plasma and kidney levels of Ang II were measured by radioimmunoassay, and Western analysis of Ang II subtype 1 (AT1) receptors was performed. RESULTS: STZ rats tended to have increased plasma and intrarenal levels of Ang II compared with C and STZ-I rats. In proximal tubules, mRNA for renin was significantly increased in STZ rats, with reversal to control values in STZ-I rats (C, 2432 +/- 437 vs. STZ, 5688 +/- 890 fg/0.25 microg RNA, P < 0.05 vs. C, N = 9, vs. STZ-I, 1676 +/- 376 fg/0.25 microg RNA, P = NS vs. C). In STZ rats, the AT1 receptor antagonist losartan caused a further fivefold increase in proximal tubule renin mRNA, associated with proximal tubular renin immunostaining. STZ had no significant effect on mRNA expression for angiotensinogen or angiotensin converting enzyme in proximal tubules. By Western blot analysis, cortical and proximal tubule AT1 receptor protein expression was significantly decreased in STZ rats. CONCLUSIONS: These data suggest activation of the proximal tubule renin angiotensin system in early STZ diabetes, mediated at least partly by enhanced expression of renin mRNA. Increased local production of Ang II could contribute to tubulointerstitial injury in this model. PMID- 11115067 TI - Integrin expression and IgA nephropathy: in vitro modulation by IgA with altered glycosylation and macromolecular IgA. AB - BACKGROUND: Signal transduction by mesangial cell (MC) integrins regulates cell growth and survival, extracellular matrix production, and organization. The aim of the study was to investigate human MC integrin modulation by differently glycosylated IgA and macromolecular IgA, which are thought to play a pathogenetic role in IgA nephropathy (IgAN). METHODS: MCs were incubated with purified human polymeric IgA, heat-aggregated IgA, IgA glycoforms generated by enzymatic hydrolysis of saccharide residues and serum fractions from IgAN patients, and controls isolated by lectin affinity and containing IgA with peculiar glycan patterns. Integrins were quantitated by flow cytometry. RESULTS: Cultured MCs highly expressed alphavbeta3 and some alpha3beta1; alphavbeta3 was up-regulated by matrix components (P < 0.02). In vitro desialylated and degalactosylated polymeric human IgA enhanced alphavbeta3 expression on cultured MCs (P < 0.001). Serum IgA glycoforms isolated from IgAN patients with high exposure of internal sugars, GalNAc, Neu5Ac2,6GalNAc, and Man enhanced alphav expression on cultured MCs more than healthy controls. CONCLUSIONS.: These data support the hypothesis that IgA glycation plays a role in modulating the cell-matrix interaction, and that this mechanism can be operating in IgAN. PMID- 11115068 TI - Troglitazone halts diabetic glomerulosclerosis by blockade of mesangial expansion. AB - BACKGROUND: Renal complications of long-term, poorly controlled type 2 diabetes mellitus include glomerulosclerosis and interstitial fibrosis. The onset and progression of these complications are influenced by underlying pathophysiologies such as hyperglycemia, hypertriglyceridemia, and hypercholesterolemia. Troglitazone, a thiazolidinedione, has been shown to ameliorate these metabolic defects. However, it was not known whether therapeutic intervention with troglitazone would prevent the onset and progression of glomerulosclerosis. METHODS: Sixty male ZDF/Gmitrade mark rats and 30 age-matched Zucker lean rats were in the study. The ZDF/Gmitrade mark rats were divided into two groups, one in which blood glucose levels were uncontrolled (30 animals) and another (30) in which blood glucose was controlled via dietary administration of troglitazone. Ten animals from each group were sacrificed at one, three, and six months into the study. The kidneys were harvested and processed for immunostaining with BM CSPG, a marker for mesangial matrix. Images of 200 glomeruli per animal were captured using digital imaging microscopy, and the index of mesangial expansion (total area mesangium/total area of tuft) per glomerular section was measured. RESULTS: The administration of troglitazone ameliorated the metabolic defects associated with type 2 diabetes mellitus. Moreover, the glomeruli from tissue sections of animals given troglitazone showed no mesangial expansion when compared with normoglycemic control animals, whereas the uncontrolled diabetic animals showed significant mesangial expansion at all time intervals. CONCLUSIONS: Therapeutic intervention with the thiazolidinedione troglitazone halts the early onset and progression of mesangial expansion in the ZDF/Gmitrade mark rat, preventing the development of glomerulosclerosis in this animal model of type 2 diabetes mellitus. PMID- 11115069 TI - Hypoxia promotes fibrogenesis in human renal fibroblasts. AB - BACKGROUND: The mechanisms underlying progressive renal fibrosis are unknown, but the common association of fibrosis and microvascular loss suggests that hypoxia per se may be a fibrogenic stimulus. METHODS: To determine whether human renal fibroblasts (HRFs), the primary matrix-producing cells in the tubulointerstitium, possess oxygen-sensitive responses relevant to fibrogenesis, cells were exposed to 1% O2 in vitro. RESULTS: Hypoxia simultaneously stimulated extracellular matrix synthesis and suppressed turnover with increased production of collagen alpha1(I) (Coll-I), decreased expression of collagenase, and increased tissue inhibitor of metalloproteinase (TIMP)-1. These effects are time dependent, require new RNA and protein synthesis, and are specific to hypoxia. The changes in Coll-I and TIMP-1 gene expression involve a heme-protein O2 sensor and protein kinase- and tyrosine kinase-mediated signaling. Although hypoxia induced transforming growth factor-beta1 (TGF-beta1), neutralizing anti-TGF-beta1 antibody did not block hypoxia-induced Coll-I and TIMP-1 mRNA expression. Furthermore, hypoxic-cell conditioned-medium had no effect on the expression of these mRNAs in naive fibroblasts, suggesting direct effects on gene transcription. Transient transfections identified a hypoxia response element (HRE) in the TIMP-1 promoter and demonstrated HIF-1-dependent promoter activation by decreased ambient pO2. CONCLUSIONS: These data suggest that hypoxia co ordinately up-regulates matrix production and decreases turnover in renal fibroblasts. The results support a role for hypoxia in the pathogenesis of fibrosis and provide evidence for novel, direct hypoxic effects on the expression of genes involved in fibrogenesis. PMID- 11115070 TI - Renal fibrosis in mice treated with human recombinant transforming growth factor beta2. AB - BACKGROUND: The biologic responses to transforming growth factor-beta (TGF-beta) suggest many potential therapeutic applications; however, in the only clinical trial to examine the effect of the systemic administration of a TGF-beta isoform, patients experienced significant but reversible declines in renal function. We studied the effects of administering human recombinant TGF-beta2 to adult mice. METHODS: The effect of daily administration of TGF-beta2 on tissue vasoconstriction, tissue levels of endothelin and angiotensin II, tissue hypoxia, and renal fibrosis were examined. RESULTS: Daily administration of TGF-beta2 at 10 or 100 microg/kg caused apparent tissue vasoconstriction that was visualized by vascular casting, with the largest impact seen in the kidney. Tissue levels of endothelin 1 and angiotensin II were significantly elevated in kidneys of treated mice, as was urinary thromboxane beta2. Renal fibrosis was observed in the cortical tubular interstitium and vasculature, particularly at the cortical medullary junction and medullary vasa recta; however, glomerular sclerosis was not observed. Fibrosis was correlated to focal tissue hypoxia as determined by immunohistochemical detection of tissue bound pimondazole. CONCLUSION: We conclude that there are significant histopathologic consequences, focused in the kidney, resulting from the daily administration of high doses of human recombinant TGF-beta2, and we propose that selective vascular constriction with consequent tissue hypoxia is a contributing factor. PMID- 11115071 TI - Requirement of heat shock protein 90 in mesangial cell mitogenesis. AB - BACKGROUND: Hyperplasia of mesangial cells (MCs) is a frequent finding in glomerulonephritis. Heat shock protein 90 (HSP90) is a major cellular chaperone that assists protein folding under physiological and stress conditions. METHODS: To identify genes that are potentially involved in the pathogenesis of glomerulonephritis, we analyzed glomerular gene expression in mesangioproliferative rat anti-Thy1.1 nephritis by representational difference analysis (RDA). Expression of HSP90beta in anti-Thy1.1 nephritis was studied by Northern and Western blot analyses and immunohistochemistry. In cultured rat MCs, the requirement of HSP90 for mitogenic signaling steps and MC replication was studied by incubation with the specific HSP90 inhibitor geldanamycin. RESULTS: By RDA, a cDNA fragment homologous to HSP90beta was identified. Glomerular mRNA and protein expression of HSP90beta was markedly and transiently up-regulated during the course of anti-Thy1.1 nephritis, with a maximum at day 6, coinciding with the peak of MC proliferation. By immunohistochemistry, HSP90beta expression in normal glomeruli was detected in podocytes. However, in anti-Thy1.1 nephritis, glomerular HSP90beta protein expression was strongly and transiently increased in mesangial localization. In vitro, mitogenic stimulation of rat MCs led to the induction of HSP90beta mRNA and protein. Incubation of MCs with geldanamycin dose dependently inhibited DNA synthesis and replication. Moreover, geldanamycin interfered with mitogen-induced phosphorylation of extracellular signal-regulated kinase and transcription of c-fos and Egr-1, but not with transactivation of STAT1 transcription factor. Cell cycle analysis of serum-stimulated MCs revealed that geldanamycin inhibited kinase activity of cyclin D1/CDK4 complexes and blocked progression in the G0/G1 phase and at the S/G2 phase transition. CONCLUSIONS: The up-regulation of HSP90beta in anti-Thy1.1 nephritis may reflect its functional involvement in phenotypical alterations of MCs in mesangioproliferative glomerulonephritis. Our in vitro studies indicate that HSP90 governs the capacity of MCs to respond to proliferative stimuli by regulating critical mitogenic signaling steps necessary for G1 entry and S-phase progression. PMID- 11115072 TI - Vascular endothelial growth factor accelerates renal recovery in experimental thrombotic microangiopathy. AB - BACKGROUND: Renal microvascular injury characterizes thrombotic microangiopathy (TMA). The possibility that angiogenic growth factors may accelerate recovery in TMA has not been studied. METHODS: TMA was induced in rats by the selective right renal artery perfusion of antiglomerular endothelial cell IgG (30 mg/kg). Twenty four hours later, rats received vascular endothelial growth factor (VEGF121, 100 microg/kg/day) or vehicle (control) daily until day 14. To evaluate renal function, the unperfused left kidney was removed at day 14, and rats were sacrificed at day 17. RESULTS: The induction of TMA was associated with loss of glomerular and peritubular capillary endothelial cells and decreased arteriolar density at day 1. Some spontaneous capillary recovery was present by day 17; however, repair was incomplete, and severe tubulointerstitial damage occurred. The lack of complete microvascular recovery was associated with reduced VEGF immunostaining in the outer medulla. VEGF-treated rats had more glomeruli with intact endothelium, less glomerular ischemia (collapsed glomeruli), and greater peritubular capillary density with less peritubular capillary loss. This was associated with less tubulointerstitial fibrosis, less cortical atrophy, and improved renal function. CONCLUSIONS: VEGF accelerates renal recovery in this experimental model of TMA. These studies suggest that angiogenic growth factors may provide a new therapeutic strategy for diseases associated with endothelial cell injury. PMID- 11115073 TI - Heat shock protein-70 repairs proximal tubule structure after renal ischemia. AB - BACKGROUND: Recent studies have suggested a role of heat shock protein (HSP)-70 in cytoskeletal repair during cellular recovery from renal ischemia. The aim of this study was to test the hypothesis that HSP-70 interacts in vitro with cytoskeletal elements obtained from rat renal cortex during early reflow after renal ischemia. METHODS: Cellular proteins were fractionated into cytoskeletal pellets and noncytoskeletal supernatants by Triton X-100 extraction of rat renal cortex obtained after 15 minutes or 18 hours of reflow after 45 minutes of renal ischemia, or from controls. Aliquots of isolated pellets were coincubated with aliquots of isolated supernatants in different combinations. A repeat Triton extraction was performed, and differential distribution of Na, K-ATPase or HSP-70 was assessed by Western blots and densitometric analysis. RESULTS: Coincubation of cytoskeletal pellets obtained during early reflow after renal ischemia (exhibiting severe injury of the cytoskeletal anchorage of Na,K-ATPase) and noncytoskeletal supernatant obtained during later reflow (showing high HSP expression) resulted in specific translocation of HSP-70 from the supernatant into the pellet, functionally associated with dose-dependent stabilization of Na,K-ATPase within this cytoskeletal fraction. These effects could be reproduced by incubation with purified HSP-70 and were abolished by the addition of anti-HSP 70 antibodies. CONCLUSION: These data support the hypothesis that HSP-70 interacts with cytoskeletal elements during the restoration of proximal tubule cell structure and polarity after renal ischemia. This experimental approach represents a new in vitro assay to study further the role of HSP in cellular repair. PMID- 11115074 TI - Monocyte chemoattractant protein-1 and macrophage infiltration in hypertensive kidney injury. AB - BACKGROUND: We investigated whether monocyte chemoattractant protein-1 (MCP-1) is expressed in hypertensive nephrosclerosis, and tested the effect of angiotensin II type 1 receptor blockade on MCP-1 expression and macrophage (MPhi) infiltration. METHODS: Rats with two-kidney, one-clip (2K1C) hypertension with and without treatment with the angiotensin II type 1 receptor antagonist valsartan (3 mg/kg/day) were studied. In these animals as well as in spontaneously hypertensive rats (SHR), stroke-prone SHR (SHR-SP), hypertensive mRen-2 transgenic rats (TGR), and respective control strains, MCP-1 expression in the kidney was investigated by Northern and Western blots and by immunohistochemistry. Glomerular and interstitial MPhis were counted. RESULTS: In the nonclipped kidney of 2K1C rats, MCP-1 expression was elevated at 14 and 28 days when significant MPhi infiltration was present. MCP-1 was localized to glomerular endothelial and epithelial cells, interstitial and tubular cells, MPhis, and vascular smooth muscle cells. A similar pattern of MCP-1 staining was present in TGR kidneys, whereas MCP-1 expression was not increased in SHR and SHR SP. Valsartan reduced but did not normalize blood pressure, blocked the induction of MCP-1 protein in 2K1C kidneys, and decreased interstitial MPhi infiltration significantly. CONCLUSION: MCP-1 expression is increased in angiotensin II dependent models of hypertensive nephrosclerosis and is temporally and spatially related to MPhi infiltration. The angiotensin II type 1 receptor mediates the induction of MCP-1. PMID- 11115075 TI - Cyclooxygenase-2 expression is associated with the renal macula densa of patients with Bartter-like syndrome. AB - BACKGROUND: Bartter-like syndrome (BLS) is a heterogeneous set of congenital tubular disorders that is associated with significant renal salt and water loss. The syndrome is also marked by increased urinary prostaglandin E2 (PGE2) excretion. In rodents, salt and volume depletion are associated with increased renal macula densa cyclooxygenase-2 (COX-2) expression. The expression of COX-2 in human macula densa has not been demonstrated. The present studies examined whether COX-2 can be detected in macula densa from children with salt-wasting BLS versus control tissues. METHODS: The intrarenal distribution of COX-2 protein and mRNA was analyzed by immunohistochemistry and in situ hybridization in 12 patients with clinically and/or genetically confirmed BLS. Renal tissue rejected for transplantation, from six adult patients not affected by BLS, was also examined. RESULTS: The expression of COX-2 immunoreactive protein was observed in cells of the macula densa in 8 out 11 patients with BLS. In situ hybridization confirmed the expression of COX-2 mRNA in the macula densa in 6 out of 10 cases. COX-2 protein was also detected in the macula densa in a patient with congestive heart failure. The expression of COX-2 immunoreactive protein was not observed in cells associated with the macula densa in kidneys from patients without disorders associated with hyper-reninemia. CONCLUSION: These studies demonstrate that COX-2 may be detected in the macula densa of humans. Since macula densa COX-2 was detected in cases of BLS, renal COX-2 expression may be linked to volume and renin status in humans, as well as in animals. PMID- 11115076 TI - Regression of sclerosis in aging by an angiotensin inhibition-induced decrease in PAI-1. AB - BACKGROUND: Glomerular and vascular sclerosis increase with aging, and angiotensin inhibitors ameliorate progression of this injury. We investigated the potential for achieving regression of existing age-related sclerosis, and the mechanisms by which angiotensin type 1 receptor antagonist (AIIRA) may affect remodeling of this sclerosis. We focused on plasminogen activator inhibitor-1 (PAI-1) because it is directly induced by angiotensin, inhibits matrix degradation, and may thus be pivotal in remodeling. METHODS: Eighteen-month-old male Sprague-Dawley rats were treated with the AIIRA losartan (N = 8, 80 mg/L, dry weight), sacrificed at age 21 and 24 months, and compared with age-matched untreated controls (N = 15). Blood pressure and renal function were monitored, and morphological, biochemical, and molecular analyses were done on aorta and kidney. RESULTS: Body weight increased in both groups. Mean arterial pressure (MAP) and serum creatinine remained normal (24-month MAP 115 +/- 8 vs. 113 +/- 6 mm Hg, controls vs. AIIRA, P = NS). Aorta wall thickness ratio was reduced by AIIRA at 21 and 24 months vs. age-matched controls (21 months 0. 12 +/- 0.01 vs. 0.15 +/- 0.01, P = 0.006; 24 months 0.10 +/- 0.005 vs. 0.14 +/- 0.003, AIIRA vs. controls, respectively, P = 0.0027). The aorta wall thickness ratio after treatment with AIIRA for six months was even lower than that of 18-month control rats (P = 0.018). AIIRA reduced proteinuria versus age-matched control at 24 months (253 +/- 62 vs. 390 +/- 51 mg/24 h, P = 0.0017). AIIRA at 24 months decreased glomerulosclerosis versus age-matched control (sclerosis index, 0 to 4+ scale: 0.06 +/- 0.02 vs. 0.49 +/- 0.12, P = 0.0082) to levels even lower than the 18-month baseline (0.37 +/- 0.14, P = 0.014). Renal collagen content increased with aging and was decreased by AIIRA at 24 months (5.0 +/- 0.7 vs. 3.1 +/- 0.5% collagen, P < 0.05). Apoptosis, assessed by TUNEL, was increased in tubular and interstitial cells in aging and was reduced by AIIRA versus control and baseline, respectively (TUNEL scoring, AIIRA 24 months 0.33 +/- 0.16 vs. 1.06 +/- 0.23 and 0.80 +/- 0.05, P < 0.05). PAI-1 mRNA in kidney was decreased at 24 months in AIIRA versus age-matched controls (PAI-1/GAPDH density ratio: AIIRA 24 months 0. 34 +/- 0.05 vs. 24-month controls 0.99 +/- 0.05, P < 0.05). Increased glomerular PAI-1 immunostaining with aging was decreased by AIIRA at 24 months versus age matched controls, even below baseline (staining score 0 to 4+, 0.57 +/- 0.15 vs. control 0.90 +/- 0.07, P < 0.05; baseline 1.05 +/- 0.02, P < 0.01). CONCLUSION: We conclude that AIIRA not only slows the progression of glomerular and vascular sclerosis in aging, but can also induce regression of these processes. The mechanisms appear to involve modulation of cortical cell turnover and inhibition of PAI-1 expression. PMID- 11115077 TI - Angiotensin type 2 receptor is expressed in the adult rat kidney and promotes cellular proliferation and apoptosis. AB - BACKGROUND: Angiotensin II (Ang II) is associated with cell proliferation and apoptosis. The role of the angiotensin type 2 receptor (AT2R) in these processes remains controversial. Conventional radioligand binding of 125I-Sar1, Ile8 Ang II in adult kidney has failed to demonstrate the binding for the AT2R. METHODS: The presence of the AT2R was explored in adult rat kidney by in vitro and in vivo autoradiography using the selective AT2R radioligand 125I-CGP 42112B. The roles of the angiotensin type 1 receptor (AT1R) and the AT2R in mediating cellular proliferation and apoptosis were assessed using selective AT1R or AT2R antagonists in Ang II-infused Sprague-Dawley (SD) rats. RESULTS: 125I-CGP 42112B binding was demonstrated by in vitro and in vivo autoradiography techniques in the glomeruli and proximal tubules of SD rats. This binding could be displaced by Ang II and the AT2R antagonist PD123319 but not by the AT1R antagonist valsartan. Subcutaneous infusion of Ang II for 14 days in eight-week-old SD rats induced proliferation of proximal tubular epithelial cells, as assessed by a twofold increase in proliferating cell nuclear antigen (PCNA)-positive cells and apoptosis, as assessed by a threefold increase in terminal dUTP nick end labeling (TUNEL)-positive cells. The administration of the AT2R antagonist PD123319 or the AT1R antagonist valsartan was associated with attenuation of the increases in both PCNA- and TUNEL-positive cells following Ang II infusion. Ang II infusion was associated with increased osteopontin gene and protein expression, which could be reduced by treatment with either valsartan or PD123319. CONCLUSION: These findings indicate that there is significant expression of the AT2R in the adult kidney, and that the AT2R has a role in mediating Ang II-induced proliferation and apoptosis in proximal tubular epithelial cells and expression of osteopontin. PMID- 11115078 TI - F-actin fiber distribution in glomerular cells: structural and functional implications. AB - BACKGROUND: Glomerular distention is associated with cellular mechanical strain. In addition, glomerular distention/contraction is assumed to influence the filtration rate through changes in filtration surface area. A contractile cytoskeleton in podocytes and mesangial cells, formed by F-actin-containing stress fibers, maintains structural integrity and modulates glomerular expansion. In this study, the glomerular cell distribution of F-actin and vimentin filaments was studied in normal control and nine-month streptozotocin-diabetic rats. Results in situ were compared with observations in tissue culture. METHODS: Microdissected rat glomeruli were perfused to obtain a physiological 25% glomerular expansion over the basal value. Fixation was done without change in glomerular volume. Dual fluorescent labeling of F-actin and vimentin was carried out, and samples were examined by confocal laser scanning microscopy. RESULTS: The podocyte cell bodies and their cytoplasmic projections, including the foot processes, contained bundles of vimentin-containing fibers. Except for a thin layer at the base of foot processes, they did not demonstrate F-actin. While mesangial cells in culture had F-actin as long stress fibers resembling tense cables, mesangial cells stretched in situ contained a maze of short tortuous F actin fibers organized in bundles that often encircled vascular spaces. This fibrillar organization was disrupted in diabetic glomeruli. CONCLUSION: Mesangial cells, but not podocytes, contain a cytoskeleton capable of contraction that is disorganized in long-term diabetes. Together with previous observations, the distribution of this cytoskeleton suggests that mesangial cell contraction may be involved in the redistribution of glomerular capillary blood flow, but not substantially in the modulation of glomerular distention. Disorganization of stress fibers may be a cause of hyperfiltration in diabetes. PMID- 11115079 TI - Dietary potassium and magnesium supplementation in cyclosporine-induced hypertension and nephrotoxicity. AB - BACKGROUND: Cyclosporine A (CsA)-induced hypertension and nephrotoxicity are aggravated by high sodium intake. Accumulating evidence suggests that potassium and magnesium supplementation could protect against the detrimental effects of dietary salt. In the present study, we tested the hypothesis of whether concurrent supplementation with potassium and magnesium could protect against the development of CsA-induced hypertension and nephrotoxicity more effectively than supplementation with one mineral alone. METHODS: Eight-week-old spontaneously hypertensive rats (SHRs) were divided into four groups (N = 10 in each group): (1) CsA group (5 mg/kg subcutaneously) receiving high-sodium diet (Na 2.6%, K 0.8%, Mg 0.2% wt/wt); (2) CsA group receiving a high-sodium, high-potassium diet (Na 2.6%, K 2.4%, Mg 0.2%); (3) CsA group receiving high-sodium, high-magnesium diet (Na 2.6%, K 0.8%, Mg 0.6%); and (4) CsA group receiving high-sodium, high potassium, high-magnesium diet (Na 2.6%, K 2.4%, Mg 0.6%). RESULTS: CsA induced severe hypertension and deteriorated renal functions in SHRs on high-sodium diet. Histologically, the kidneys showed severe thickening of the media of the afferent artery with fibrinoid necrosis. Potassium supplementation lowered blood pressure (198 +/- 5 vs. 212 +/- 2 mm Hg, P < 0.05) and partially prevented the development of proteinuria (-25%, P < 0.05). Magnesium supplementation decreased blood pressure to the same extent but improved renal functions more effectively than potassium. The greatest protection against CsA toxicity was achieved when dietary potassium and magnesium supplementations were combined. Urinary N-acetyl-beta-D glucosaminidase (NAG) excretion, a marker for renal proximal tubular damage, increased progressively in CsA-treated SHRs on the high-sodium diet. Neither potassium nor magnesium influenced urinary NAG excretion. We also estimated the activity of the renal dopaminergic system by measuring 24-hour urinary dopamine excretion rates. CsA suppressed the renal dopaminergic system during high-sodium diet. Magnesium supplementation, alone and in combination with potassium, protected against the development of renal dopaminergic deficiency in CsA-treated SHRs on high-sodium diet. Magnesium supplementation increased plasma-free ionized magnesium (iMg) and bone magnesium by 50 and 16%, respectively. CONCLUSIONS: Our findings indicate that both potassium and magnesium supplementations showed beneficial effects against CsA-induced hypertension and nephrotoxicity. The protective effect of magnesium clearly exceeded that of potassium. The greatest protection against CsA toxicity was achieved when potassium and magnesium were combined. We also provide evidence that the development of CsA-induced glomerular, tubular, and vascular lesions are associated with renal dopaminergic deficiency. PMID- 11115080 TI - Proteinase 3 gene polymorphisms and Wegener's granulomatosis. AB - BACKGROUND: Wegener's granulomatosis (WG) is a rare systemic autoimmune disease characterized by small-vessel vasculitis leading to organ damage and the presence of antineutrophil cytoplasmic autoantibodies (ANCAs). ANCAs were shown to be involved in the pathogenesis of the disease by increasing adhesion of polymorphonuclear cells (PMNs) to endothelial cells and through activation of primed PMN. The main autoantigen of ANCA in WG is proteinase 3 (PR3), a neutrophil- and monocyte-derived neutral serine protease. The association of WG with individuals continuously expressing a high level of PR3 on the surface of PMNs suggests that PR3 variants or altered regulation of PR3 expression might be directly involved in the pathogenesis of the disease. METHODS: We screened the entire coding and promoter sequences of the PR3 gene for polymorphisms by means of polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP). Allelic, genotypic, and haplotype frequencies were compared between 79 WG patients and a cohort of 129 healthy controls. RESULTS: Seven single-nucleotide polymorphisms (SNPs), one amino acid change (Val119Ile), one 84 bp insertion/deletion, and a microsatellite were identified. An association with WG could be demonstrated for the A-564G polymorphism in the PR3 promoter affecting a putative transcription factor-binding site. CONCLUSIONS: This study excludes certain PR3 epitope variants as autoantigenic stimuli in WG, since the Val119Ile polymorphism showed no differences between patients and controls. Overexpression of PR3, however, might predispose the patient to the development of autoimmune ANCA-associated vasculitis. PMID- 11115081 TI - Pharmacodynamics and pharmacokinetics of polyethylene glycol-hirudin in patients with chronic renal failure. AB - BACKGROUND: Hirudin selectively inhibits thrombin without cofactors and is eliminated via the kidneys. Recombinant hirudin (r-hi) has a terminal elimination half-life (t1/2) of about 50 to 100 minutes. Coupling of polyethylene glycol (PEG) to r-hi, giving PEG-hirudin (PEG-Hi), prolongs its t1/2 while enhancing efficacy. We looked at the pharmacodynamic and pharmacokinetic behavior of PEG-Hi in patients with impaired renal function. METHODS: Anticoagulant activity and the pharmacokinetic parameters of a single intravenous bolus injection of 0.05 mg/kg body weight PEG-Hi were studied in 38 subjects. They were assigned to five groups: group IA, creatinine clearance (CCr) >/= 80 mL/min, 8 healthy volunteers; group IB, CCr >/= 80 mL/min, 8 patients with normal renal function); group II, CCr 79 to 50 mL/min, 7 patients with mild chronic renal failure (CRF); group III, CCr 49 to 20 mL/min, 10 patients with moderate CRF; and group IV, CCr 0.05), and 185 +/- 52 mL (P < 0.01), respectively. In 19 individual spherical cysts selected in six patients, the mean initial volume was 15.0 +/- 7.2 mL (range 1.1 to 137 mL), and the average rate of volume increase was 0.52 +/- 0.21 mL/month (P < 0.025, range 0.02 to 4.15 mL/month). In five patients who eventually required dialysis, 11.2 years after the initial CT scan, the initial cyst/kidney volume ratio (combined for both kidneys) exceeded 0.43; four patients with lower cyst/kidney volume ratios had serum creatinine levels <1.5 mg/dL, 8.7 years after the initial CT scan. CONCLUSIONS: On the basis of this preliminary survey of archival material, we conclude that conventional contrast-enhanced CT scans can be used to quantitate volume components of progression in ADPKD. The rates of individual kidney and cyst enlargement are highly variable within and between patients, but overall, the values increase over time. The volume fraction of kidneys comprised of cysts may be a useful indicator of ADPKD progression. PMID- 11115084 TI - Effect of plasma fractions from patients with focal and segmental glomerulosclerosis on rat proteinuria. AB - BACKGROUND: Patients suffering from focal and segmental glomerulosclerosis (FSGS) and in whom this disease recurs after transplantation are likely to have an active form of the disease and to have a factor(s) (such as, albuminuric factor) present in their blood that alters glomerular permeability for albumin. METHODS: We used a sequential 50 and 70% ammonium sulfate (AS) precipitation of plasma from patients with relapsing FSGS and non-FSGS nephrotic syndrome (NS), in addition to plasma from healthy individuals, to obtain both an immunoglobulin (Ig)-rich fraction (50% AS precipitate) and a non-Ig fraction (70% AS supernatant). These fractions were injected intra-arterially or intravenously/intraperitoneally into Sprague-Dawley rats, and proteinuria (g protein/mmol creatinine) was measured for 24 hours. Ig fractions eluted from immunoadsorption onto protein A were also tested. A biochemical characterization was then carried out on the 70% AS supernatants by ultrafiltration on 30 and 50 kD cut-off membranes and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Differentially stained bands were sequenced. RESULTS: The 70% AS supernatants from FSGS patients induced proteinuria when injected intra-arterially into normal rats. This effect was significantly different (P < 0.05) from that observed when similar fractions were prepared from the plasma of patients suffering from non-FSGS NS, but was not different from that observed with fractions from healthy individuals and even with an injection of saline solution. Injections of other plasma fractions did not induce a significant proteinuria in the FSGS group versus the non-FSGS NS group. SDS-PAGE of 70% AS supernatants revealed a protein of 23 kD that was more concentrated in AS supernatants from FSGS plasma than the other plasma samples and that was identified by microsequencing as apolipoprotein A1. After sequential ultrafiltration of 70% AS supernatants on 30 and 50 kD cut-off membranes, a second band of 43 kD was found at a much higher concentration in the FSGS samples than in non-FSGS NS and healthy individuals samples. This band is likely to correspond to a candidate albuminuric factor recently reported by another group [1], and was identified by microsequencing as alpha1 acid glycoprotein or orosomucoid. Consequently, purified orosomucoid from the plasma of FSGS, non-FSGS NS patients, or healthy individuals was injected intra-arterially into rats. No differences were found between the proteinuria induced in each group. CONCLUSIONS: These data strongly suggest that in vivo injection of material into the rat is not a reliable model for testing plasma fraction activity and that the 43 kD orosomucoid is not likely to be the albuminuric factor. PMID- 11115085 TI - Prealbumin is as important as albumin in the nutritional assessment of hemodialysis patients. AB - BACKGROUND: Although serum prealbumin is considered a valid indicator of nutritional status in hemodialysis patients, there is relatively little evidence that its determination is of major prognostic significance. In this study, we aimed to determine the independent association of serum prealbumin with survival in hemodialysis patients, after adjusting for serum albumin and other indicators of protein energy nutritional status. METHODS: Serum prealbumin was measured in more than 1600 maintenance hemodialysis patients. We determined the correlations among prealbumin and other indicators of nutritional status, including serum albumin, and bioimpedance-derived indicators of body composition. The relationship between serum prealbumin and survival was determined using proportional hazards regression. RESULTS: The serum albumin was directly correlated with the serum prealbumin (r = 0.47, P < 0.0001), but still explained <25% of the variability in prealbumin. Prealbumin was inversely related to mortality, with a relative risk reduction of 6% per 1 mg/dL increase in prealbumin, even after adjusting for case mix, serum albumin, and other nutritional indicators. The increase in risk with lower serum prealbumin concentrations was observed whether the serum albumin was high or low. CONCLUSION: In hemodialysis patients, the serum prealbumin provides prognostic value independent of the serum albumin and other established predictors of mortality in this population. PMID- 11115086 TI - Effect of dwell time on carbonyl stress using icodextrin and amino acid peritoneal dialysis fluids. AB - BACKGROUND: Deterioration of the peritoneal membrane limits the technical survival of peritoneal dialysis (PD). Advanced glycation of the membrane has been incriminated in this evolution. Advanced glycation end products (AGEs) develop under the influence of glucose and of its degradation products, mainly reactive carbonyl compounds (RCOs) such as glyoxal (GO), methylglyoxal (MGO), and 3 deoxyglucosone (3-DG). The present study was undertaken to evaluate the impact of recently developed glucose-free PD fluids on AGE generation. METHODS: Recently developed glucose-free PD fluids containing either icodextrin or amino acids were investigated. GO, MGO, and 3-DG [high-performance liquid chromatography (HPLC)] and total RCOs (spectrophotometry) were measured in fresh solutions and in effluents after various dwell duration. The AGE formation potential of PD fluids and effluents was assessed by incubation at 37 degrees C, for one week, with bovine serum albumin and by the eventual measurement of pentosidine (HPLC) and Nepsilon-carboxymethyllysine (CML; gas chromatography/mass spectrometry). RESULTS: GO, MGO, and 3-DG (P < 0. 001) as well as total RCOs levels (P < 0.01) were significantly lower in icodextrin and amino acid PD fluid than in commercial, heat-sterilized, 1.36% glucose PD fluid. Pentosidine and CML generation were also significantly lower (P < 0.001) in icodextrin and amino acid PD fluid than in conventional 1.36% glucose PD fluid. The levels of total RCOs, however, increased in icodextrin and amino acid PD fluid effluents with dwell time. AGE formation potential rose accordingly, as demonstrated by a parallel increase in the generation of pentosidine and CML during incubation of PD effluents. CONCLUSION: The present data demonstrate lower RCO contents and AGE formation potential in fresh icodextrin and amino acid PD fluids than in fresh heat-sterilized glucose PD fluids. However, this difference decreases progressively during dwell time, mainly as a result of the influx of total RCOs. PMID- 11115087 TI - Protection from ototoxicity of intraperitoneal gentamicin in guinea pig. AB - BACKGROUND: Aminoglycoside antibiotics are common to treat peritonitis and exit site infections in patients on peritoneal dialysis. Ototoxicity (loss of hearing or balance) is a well-documented adverse effect of aminoglycosides, and severe ototoxic reactions have been noted in patients receiving these drugs by intraperitoneal lavage. We have proposed a free-radical hypothesis for the mechanism of aminoglycoside ototoxicity and suggested a therapeutic prevention by the concomitant administration of antioxidants or iron chelators. Here we investigate whether 2, 3-dihydroxybenzoate can prevent the ototoxicity of intraperitoneal gentamicin. METHODS: Two strains of pigmented guinea pigs received daily intraperitoneal injections of gentamicin. Both strains developed ototoxicity, although different dosages were needed to produce similar auditory deficits (120 mg gentamicin base/kg body weight daily for 19 days vs. 135 mg/kg for 14 days). Dihydroxybenzoate was administered intraperitoneally once or twice daily. Auditory thresholds were measured by evoked brain stem response. Pathology was assessed as a loss of sensory cells in surface preparations of the organ of Corti. RESULTS: The auditory threshold shifts and hair cell loss were similar to the pathology observed following subcutaneous injections of gentamicin. Animals sustained almost complete loss of outer hair cells in the basal cochlea and a progressive hearing loss with threshold shifts of 60 dB at 18 kHz. The concomitant administration of dihydroxybenzoate significantly attenuated the threshold shift to less than 30 dB and reduced the loss of hair cells. The treatment with dihydroxybenzoate did not affect serum gentamicin levels. CONCLUSIONS: Antioxidant therapy is a promising approach to prevent aminoglycoside-induced hearing loss following intraperitoneal application. PMID- 11115088 TI - Acute rejection before cytomegalovirus infection enhances von Willebrand factor and soluble VCAM-1 in blood. AB - BACKGROUND: Human cytomegalovirus (HCMV) infections in transplantation patients are associated with vascular endothelial damage. This is reflected by the appearance of cytomegalic endothelial cells (CECs) and noninfected endothelial cells (ECs) in blood. To get more insight in the extent of vascular damage during HCMV infection, we investigated the levels of soluble markers during HCMV infection in relationship to EC levels and also preceding the acute rejection episodes. METHODS: Of 46 kidney transplant patients, plasma levels of von Willebrand factor (VWF), soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), and soluble E-selectin (sE sel) were analyzed during the course of HCMV infection. RESULTS: Plasma levels of VWF and sVCAM-1 increased twofold during severe HCMV infection. Moreover, the plasma levels of VWF correlated with detectable cytomegalic and noninfected ECs in blood. The kinetics of changes in VWF and ECs (CEC and EC) demonstrated the relationship with HCMV-induced vascular damage. Levels of sICAM-1 and sE-sel in plasma did not significantly change during HCMV infection. Interestingly, the combination of HCMV infection and preceding acute transplant rejection caused the highest increases of VWF and sVCAM-1 plasma levels, reflecting an enhanced susceptibility for endothelial damage at the moment of infection. CONCLUSION: CMV infection is associated with vascular damage, and the vascular damage during CMV infection is enhanced if patients experienced acute rejection before CMV infection. PMID- 11115089 TI - Risk of bacteremia from temporary hemodialysis catheters by site of insertion and duration of use: a prospective study. AB - BACKGROUND: Uncuffed, nontunneled hemodialysis catheters remain the preferred means to gain immediate access to the circulation for hemodialysis. Bacteremia is the primary complication that limits their use. The risk of bacteremia by site of insertion and duration of use has not been well studied. METHODS: Two hundred eighteen consecutive patients who required a temporary hemodialysis catheter were prospectively followed. RESULTS: Catheters were placed at 318 new insertion sites and remained in use for a total of 6235 days. The incidence of bacteremia was 5.4% after three weeks of placement in internal jugular vein and 10.7% after one week in femoral vein [relative risk for bacteremia 3.1 (95% CI, 1.8 to 5.2)]. The incidence of bacteremia was 1.9% one day after the onset of an exit site infection but increased to 13.4% by the second day if the catheter was not removed. Guidewire exchange for malfunction and patient factors did not significantly affect the risk of bacteremia. CONCLUSIONS: Internal jugular catheters may be left in place for up to three weeks without a high risk of bacteremia, but femoral catheters in bed-bound patients should be removed after one week. Catheter exchanges over a guidewire for catheter malfunction do not increase bacteremia rates. Temporary catheters should be removed immediately if an exit site infection occurs. PMID- 11115090 TI - Intragraft events preceding chronic renal allograft rejection in a modified tolerance protocol. AB - BACKGROUND: Inbred miniature swine treated for 12 days with high-dose cyclosporine A develop tolerance to histocompatibility complex (MHC) class I mismatched renal allografts. When this protocol was modified by adding thymectomy before transplant, all animals developed acute rejection. Thereafter, by day 100, one half developed chronic rejection (progression group) and the other half recovered (recovery group). This provides an excellent experimental model to identify the mechanisms of chronic rejection as well as the early changes that may predict chronic rejection. METHODS: We assessed the cellular infiltration, immune activation, humoral immunity, and cell- and antibody-mediated graft injury in the progression and the recovery groups. In addition, we also examined circulating donor reactive cytotoxic T lymphocyte (CTL) and antidonor antibody in both groups. RESULTS: From days 8 to 18 after transplantation, the two groups were indistinguishable. Both showed acute rejection with endarteritis (type II); had IgG and IgM deposition in glomeruli and small vessels; had an infiltrate with similar numbers of T cells, proliferating (PCNA+) and activated (interleukin-2 receptor+) cells; and had a similar degree of parenchymal cell apoptosis [in situ DNA nick-end labeling (TUNEL)+]. However, by days 30 to 60, the two groups could be distinguished by several intragraft features. The recovery group became tolerant and had diminished T-cell infiltration, activation and proliferation, and no detectable antibody deposition. The number of TUNEL+-injured parenchymal cells decreased. In contrast, the progression group showed persistent cell infiltration with activation and proliferation. Significantly prominent TUNEL+ apoptotic parenchymal cells in tubules, glomeruli, peritubular capillaries and arteries were seen from day 30 to day 100. Circulating donor reactive CTL and antidonor class I IgG were detected in the progression group at higher levels than in the recovery group from days 30 to 60. CONCLUSION: In tolerance-induction protocols, unstable tolerance induction is associated with the persistent immunologic activation that mediates immunologic destruction of graft parenchymal cells and chronic rejection. Certain of the described immunopathologic findings (activation, proliferation, apoptosis, and antibody deposition) may be useful in distinguishing the type of rejection, that is, whether the allograft will progress to chronic rejection or recovery. PMID- 11115091 TI - Cyclosporine toxicity in renal transplant recipients detected by nailfold capillaroscopy with Na-fluorescein. AB - BACKGROUND: Cyclosporine represented a major advance in the medical management of patients with organ transplantation, but its use is limited by the frequent occurrence of hypertension and renal toxicity diagnosed by invasive renal biopsy. Renal histology shows a specific arteriolopathy. It was hypothesized that cyclosporine may also induce subclinical microvascular changes in the skin that might be detected noninvasively by a combination of dynamic capillaroscopy [capillary blood cell velocity (CBV)] with and without intravenous Na-fluorescein (NaF) injection and laser Doppler fluxmetry (LDF). METHODS: The nailfold skin microcirculation was evaluated in 112 consecutive renal transplant recipients (54 +/- 11 years old; 70 males and 42 females) receiving cyclosporine. The investigation was made the same day as a routine renal biopsy performed in all patients more than two years after transplantation. Renal biopsies were blindly classified as positive (N = 33) when significant specific signs of cyclosporine toxicity were clearly observed (AH2-AH3) and were otherwise negative (AH0-AH1, N = 79) according to the Banff classification. RESULTS: Time to fluorescence peak after NaF injection (tpNaF) was significantly longer in patients with positive biopsies than in patients with negative biopsies (13.9 +/- 8.1 vs. 17.5 +/- 9.4 sec, P = 0.009). All patients but three with negative biopsies (93%) had a tpNaF less than 10 seconds (sensitivity 91%, negative predictive value 93%). On the other hand, CBV, LDF, plasma levels of cyclosporine, and endothelin were similar in the two groups. CONCLUSION: Nailfold fluorescence capillaroscopy is an accurate and simple mean to rule out cyclosporine toxicity in renal transplant recipients. A normal test could avoid invasive renal biopsy in about 40% of the patients. Renal biopsy would, however, still be indicated when the test is abnormal. PMID- 11115092 TI - Urea and nitrogen excretion in pediatric peritoneal dialysis patients. AB - BACKGROUND: Adequate nutrition is critical to the care of children with end-stage renal disease, and failure to reach the target dietary intake is associated with growth failure. Prospective studies of urea and nitrogen output in adults have led to the derivation of quantitative relationships, which allow assessment of dietary protein intake when only urea appearance is known. Such a clinically useful relationship has not been defined in children receiving chronic peritoneal dialysis (PD). METHODS: We studied 18 pediatric PD patients (ages 0.8 to 14.3 years) on 132 occasions and determined norms of urea nitrogen appearance (UNA), total nitrogen appearance (TNA), and nonurea nitrogen appearance (NUNA). We stratified data on UNA, TNA, NUNA, nonprotein nitrogen appearance, and the protein equivalent of nitrogen appearance by age groups (0 to 5, 6 to 10, and 11 to 15 years of age) and demonstrated significant differences. In addition, dietary protein and energy intake were measured in the outpatient setting with food scales and dietitian interviews, and the results were stratified by age, presence of residual renal function, and recombinant human growth hormone (rhGH) therapy. RESULTS: UNA (3.05 +/- 1.38 g/day, 103 +/- 42 mg/kg/day) and TNA (4.67 +/- 1.86 g/day, 159 +/- 52 mg/kg/day) varied significantly between different age groups. NUNA in pediatric subjects (56 +/- 24 mg/kg/day) was significantly greater than previously published adult norms. A linear relationship was defined between UNA and TNA that was specific to pediatric PD patients [TNA (g/day) = 1.26(UNA) + 0.83]. When the relationship was scaled to body mass, the y intercept was significantly different in the youngest subjects [TNA = 1.03 (UNA) + 0.02 (weight in kg) + 0.56 (for subjects age 0 to 5) or 0.98 (for subjects age 11 to 15 or 6 to 10), r2 = 0.91]. Dietary protein intake was significantly greater in subjects receiving rhGH therapy, although nitrogen excretion was unchanged. CONCLUSIONS: Markers of protein metabolism in pediatric PD patients are age dependent and differ from adult values. An age-specific relationship between TNA and UNA is defined for pediatric subjects; it does not vary with rhGH or the presence of residual renal function. PMID- 11115093 TI - Plasma protein thiol oxidation and carbonyl formation in chronic renal failure. AB - BACKGROUND: Myeloperoxidase-catalyzed oxidative pathways have recently been identified as an important cause of oxidant stress in uremia and hemodialysis (HD), and can lead to plasma protein oxidation. We have examined patterns of plasma protein oxidation in vitro in response to hydrogen peroxide (H2O2) and hypochlorous acid (HOCl). We measured thiol oxidation, amine oxidation, and carbonyl concentrations in patients on chronic maintenance HD compared with patients with chronic renal failure (CRF) and normal volunteers. We have also examined the effect of the dialysis procedure on plasma protein oxidation using biocompatible and bioincompatible membranes. METHODS: Plasma proteins were assayed for the level of free thiol groups using spectrophotometry, protein associated carbonyl groups by enzyme-linked immunosorbent assay, and oxidation of free amine groups using a fluorescent spectrophotometer. RESULTS: In vitro experiments demonstrate HOCl oxidation of thiol groups and increased carbonyl formation. In vivo, there are significant differences in plasma-free thiol groups between normal volunteers (279 +/- 12 micromol/L), CRF patients (202 +/- 20 micromol/L, P = 0.005) and HD patients (178 +/- 18 micromol/L, P = 0.0001). There are also significant differences in plasma protein carbonyl groups between normal volunteers (0.76 +/- 0.51 micromol/L), CRF patients (13.73 +/- 4.45 micromol/L, P = 0.015), and HD patients (16.95 +/- 2.62 micromol/L, P = 0.0001). There are no significant differences in amine group oxidation. HD with both biocompatible and bioincompatible membranes restored plasma protein thiol groups to normal levels, while minimally affecting plasma protein carbonyl expression. CONCLUSIONS: First, both CRF and HD patients have increased plasma protein oxidation manifested by oxidation of thiol groups and formation of carbonyl groups. Second, HD with biocompatible and bioincompatible membranes restored plasma protein thiol groups to normal levels. Third, these experiments suggest that there is a dialyzable low molecular weight toxin found in uremia that is responsible for plasma protein oxidation. PMID- 11115094 TI - A fractional dialysate collection method to estimate solute removal in continuous venovenous hemodialysis. AB - BACKGROUND: Fractional direct dialysis quantification (fDDQ), whereby a known proportion of dialysate effluent is sampled, can reliably estimate total solute removal in intermittent hemodialysis (IHD). Our study aimed to develop and test the technique in continuous venovenous hemodialysis (CVVHD). METHODS: Twenty dialysate collections (mean duration 23.5 hours, range 17.25 to 26.6) were performed in 12 patients on CVVHD. An infusion pump diverted 10% of the total effluent volume to the fractional collection (fc), the remainder being channeled into the bulk collection (bc). Both fc and bc were collected on ice and assayed for urea nitrogen (UN) and creatinine (Cr). Actual solute removal (ASR) was calculated from the measured effluent volume and solute concentrations of the fc and bc. Estimated solute removal (ESR) was calculated from the product of the fc solute concentration and effluent volume. All fc/bc samples in 15 out of 20 collections underwent gram stain and aerobic/anaerobic culture. RESULTS: Bland Altman analyses suggested good agreement between ASR and ESR [absolute values of percentage differences: 95% CI = 1.73, 5.17% (UN); 1.88, 4.31% (Cr)]. Favorable concordance correlation coefficients confirmed this [rc = 0.995 (UN), 0.997 (Cr)] and were apparently unaffected by heavy pseudomonal growths in 4 out of 7 culture positive collections [rc = 0.997 (UN), 0.997 (Cr); culture negative (N = 8), rc = 0.996 (UN), 0.997 Cr)]. CONCLUSION: fDDQ, using 24-hour, pump-assisted, cooled fractional dialysate sampling reliably estimates total solute removal and provides a practical alternative to total dialysate collection in assessing delivered dialysis dose. PMID- 11115095 TI - A comparison of the results of renal transplantation from non-heart-beating, conventional cadaveric, and living donors. AB - BACKGROUND: In an attempt to address the shortage of conventional kidney donors, a non-heart-beating donor (NHBD) organ retrieval program has been established. We compared the results of kidney transplants from NHBDs (N = 77) with those from heart-beating cadaveric (HBD; N = 224) and living donors (LD; N = 49), performed in the same eight-year period. METHODS: Patients dying after failed attempts at resuscitation in the accident department or after intracerebral hemorrhage/anoxia were considered as potential NHBDs. After death, in situ kidney perfusion and cooling were achieved using an intra-aortic catheter inserted via a femoral artery cut down. Kidney retrieval and transplant operations were performed using standard techniques. RESULTS: The median (range) warm ischemic time for NHBD kidneys was 25 minutes (5 to 53 min). The initial function rates for NHBD, HBD, and LD transplants were 6.5, 76.3, and 93%, respectively. Primary nonfunction occurred in 5 of 75 evaluable NHBD transplants (7%) compared with only 6 out of 224 (2.7%) HBD and 1 out of 49 (2%) LD transplants (P = NS). Eighty-four percent of NHBD kidney recipients required postoperative dialysis for a median of 19 days. The mean (SD) serum creatinine at 12 months was 179 (73) micromol/L in NHBD kidneys compared with 152 (57) micromol/L for HBD kidneys and 138 (44) micromol/L for LD kidneys. The actuarial five-year graft survival rates for NHBD, HBD, and LD transplants were 79, 75, and 78%, respectively. During the period under study, NHBD organs accounted for 22% of the total renal transplant program. CONCLUSIONS: Despite being associated with poor initial graft function, the long-term allograft survival of NHBD kidneys does not differ significantly from the results of HBD and LD transplants. PMID- 11115096 TI - Evaluation of glomerular lesions in diabetes mellitus. PMID- 11115097 TI - Repairing renal lesions: will VEGF be the builder? PMID- 11115098 TI - Laparoscopic versus open donor nephrectomy. PMID- 11115099 TI - Reply from the authors: PMID- 11115100 TI - Is smoking a risk factor for progression of chronic renal failure? PMID- 11115101 TI - The filtered complement hypothesis. PMID- 11115102 TI - The vasculopathy of autosomal dominant polycystic kidney disease: insights from animal models. PMID- 11115103 TI - Riboregulation by DsrA RNA: trans-actions for global economy. AB - DsrA is an 87 nucleotide Escherichia coli RNA with extraordinary regulatory properties. The profound impact of its actions stems from DsrA regulating translation of two global transcription regulators, H-NS and RpoS (sigmas), by sequence-specific RNA-RNA interactions. H-NS is a major nucleoid-structuring and global repressor protein, and RpoS is the stationary phase and stress response sigma factor of RNA polymerase. DsrA changes its conformation to bind to these two different mRNA targets and thereby inhibits H-NS translation, while stimulating that of RpoS in a mechanistically distinct fashion. DsrA apparently binds both the start and the stop codons of hns mRNA and sharply decreases the mRNA half-life. DsrA also binds sequences in the 5'-untranslated leader region of rpoS mRNA, enhancing rpoS mRNA stability and RpoS translation. A cohort of genes, governed by H-NS repression and RpoS activation, are thus regulated. Low temperatures increase the levels of DsrA, with differential effects on H-NS and RpoS. Additionally, the RNA chaperone protein Hfq is involved with DsrA regulation, as well as with other small RNAs that also act on RpoS to co-ordinate stress responses. We address the possible functions of this genetic regulatory mechanism, as well as the advantages of using small RNAs as global regulators to orchestrate gene expression. PMID- 11115104 TI - Transcriptional regulation of transport and utilization systems for hexuronides, hexuronates and hexonates in gamma purple bacteria. AB - The comparative approach is a powerful tool for the analysis of gene regulation in bacterial genomes. It can be applied to the analysis of regulons that have been studied experimentally as well as that of regulons for which no known regulatory sites are available. It is assumed that the set of co-regulated genes and the regulatory signal itself are conserved in related genomes. Here, we use genomic comparisons to study the regulation of transport and utilization systems for sugar acids in gamma purple bacteria Escherichia coli, Salmonella typhi, Klebsiella pneumoniae, Yersinia pestis, Erwinia chrysanthemi, Haemophilus influenzae and Vibrio cholerae. The variability of the operon structure and the location of the operator sites for the main transcription factors are demonstrated. The common metabolic map is combined with known and predicted regulatory interactions. It includes all known and predicted members of the GntR, UxuR/ExuR, KdgR, UidR and IdnR regulons. Moreover, most members of these regulons seem to be under catabolite repression mediated by CRP. The candidate UxuR/ExuR signal is proposed, the KdgR consensus is extended, and new operators for all transcription factors are identified in all studied genomes. Two new members of the KdgR regulon, a hypothetical ATP-dependent transport system OgtABCD and YjgK protein with unknown function, are detected. The former is likely to be the transport system for the products of pectin degradation, oligogalacturonides. PMID- 11115105 TI - Evidence of recent lateral gene transfer among hyperthermophilic archaea. AB - A total of 153 nucleotide differences were found over a contiguous 16 kb region between two hyperthermophilic Archaea, Pyrococcus furiosus and Thermococcus litoralis. The 16 kb region in P. furiosus is flanked by insertion sequence (IS) elements with inverted and direct repeats. Both IS elements contain a single open reading frame (ORF) encoding a putative protein of 233 amino acids identified as a transposase. This 16 kb region has the features of a typical bacterial composite transposon and represents a possible mechanism for lateral gene transfer between Archaea or possibly between Archaea and Bacteria. A total of 23 homologous IS elements was found in the genome sequence of P. furiosus, whereas no full-length IS elements were identified in the genomes of Pyrococcus abyssi and Pyrococcus horikoshii. Only one IS element was found in T. litoralis. In P. furiosus and T. litoralis, the 16 kb region contains an ABC transport system for maltose and trehalose that was characterized biochemically for T. litoralis. Regulation of expression studies showed that the malE gene, located on the transposon, and the encoded trehalose/maltose-binding protein (TMBP) are induced in the presence of maltose and trehalose in both P. furiosus and T. litoralis. The implications of transposition as a mechanism for lateral gene transfer among Archaea are discussed. PMID- 11115106 TI - Phage conversion of exfoliative toxin A production in Staphylococcus aureus. AB - The staphylococcal exfoliative toxins (ETs) are extracellular proteins that cause splitting of human skin at the epidermal layer during infection in infants. Two antigenically distinct toxins possessing identical activity have been isolated from Staphylococcus aureus, ETA and ETB. The gene for ETA (eta) is located on the chromosome, whereas that for ETB is located on a large plasmid. The observation that relatively few clinical isolates produce ETA suggests that the eta gene is acquired by horizontal gene transfer. In this study, we isolated a temperate phage (phiETA) that encodes ETA and determined the complete nucleotide sequence of the phiETA genome. phiETA has a head with a hexagonal outline and a non contractile and flexible tail. The genome of phiETA is a circularly permuted linear double-stranded DNA, and the genome size is 43 081 bp. Sixty-six open reading frames (ORFs) were identified on the phiETA genome, including eta, which was found to be located very close to a putative attachment site (attP). phiETA converted ETA non-producing strains into ETA producers. Southern blot analysis of chromosomal DNA from clinical isolates suggested that phiETA or related phages are responsible for the acquisition of eta genes in S. aureus. PMID- 11115107 TI - Apical membrane antigen 1 plays a central role in erythrocyte invasion by Plasmodium species. AB - Apical membrane antigen 1 (AMA1) is an asexual blood-stage protein expressed in the invasive merozoite form of Plasmodia species, which are the causative agent of malaria. We have complemented the function of Plasmodium falciparum AMA1 (PfAMA1) with a divergent AMA1 transgene from Plasmodium chabaudi (PcAMA1). It was not possible to disrupt the PfAMA1 gene using 'knock-out' plasmids, although we demonstrate that the PfAMA1 gene can be targeted by homologous recombination. These experiments suggest that PfAMA1 is critical, perhaps essential, for blood stage growth. Importantly, we showed that PcAMA1 expression in P. falciparum provides trans-species complementation to at least 35% of the function of endogenous PfAMA1 in human red cells. Furthermore, expression of this transgene in P. falciparum leads to more efficient invasion of murine erythrocytes. These results indicate an important role for AMA1 in the invasion of red blood cells (RBCs) across divergent Plasmodium species. PMID- 11115108 TI - Identification of Icm protein complexes that play distinct roles in the biogenesis of an organelle permissive for Legionella pneumophila intracellular growth. AB - Legionella pneumophila is a bacterial pathogen that can enter the human lung and grow inside alveolar macrophages. To grow within phagocytic host cells, the bacteria must create a specialized organelle that restricts fusion with lysosomes. Biogenesis of this replicative organelle is controlled by 24 dot and icm genes, which encode a type IV-related transport apparatus. To understand how this transporter functions, isogenic L. pneumophila dot and icm mutants were characterized, and three distinct phenotypic categories were identified. Our data show that, in addition to genes that encode the core Dot/Icm transport apparatus, subsets of genes are required for pore formation and modulation of phagosome trafficking. To understand activities required for virulence at a molecular level, we investigated protein-protein interactions. Specific interactions between different Icm proteins were detected by yeast two-hybrid and gel overlay analysis. These data support a model in which the IcmQ-IcmR complex regulates the formation of a translocation channel that delivers proteins into host cells, and the IcmS-IcmW complex is required for export of virulence determinants that modulate phagosome trafficking. PMID- 11115109 TI - Generation of a non-sporulating strain of Streptomyces coelicolor A3(2) by the manipulation of a developmentally controlled ftsZ promoter. AB - The differentiation of Streptomyces aerial hyphae into chains of unigenomic spores occurs through the synchronous formation of multiple FtsZ rings, leading to sporulation septa. We show here that developmental control of ftsZ transcription is required for sporulation in Streptomyces coelicolor A3(2). Three putative ftsZ promoters were detected in the ftsQ-ftsZ intergenic region. In addition, some readthrough from upstream promoter(s) contributed to ftsZ transcription. S1 nuclease protection assays and transcriptional fusions of the ftsZ promoter region to the egfp gene (for green fluorescent protein) provided evidence that ftsZ2p is a developmentally controlled promoter that is specifically upregulated in sporulating aerial hyphae. This upregulation required all the six early regulatory sporulation genes that were tested: whiA, B, G, H, I and J. The DNA sequence of the promoter indicated that it is not part of the developmental regulon that is controlled by the RNA polymerase sigma factor sigma(WhiG). A strain in which the ftsZ2p promoter was inactivated grew normally during vegetative growth and formed aerial mycelium, but was deficient in sporulation septation. Thus, ftsZ2p was dispensable for vegetative growth, but was required for the strain to make sufficient FtsZ to support developmentally controlled multiple cell divisions in aerial hyphae. PMID- 11115110 TI - Critical protective role of bacterial superoxide dismutase in rhizobium-legume symbiosis. AB - In nitrogen-poor soils, rhizobia elicit nodule formation on legume roots, within which they differentiate into bacteroids that fix atmospheric nitrogen. Protection against reactive oxygen species (ROS) was anticipated to play an important role in Rhizobium-legume symbiosis because nitrogenase is extremely oxygen sensitive. We deleted the sodA gene encoding the sole cytoplasmic superoxide dismutase (SOD) of Sinorhizobium meliloti. The resulting mutant, deficient in superoxide dismutase, grew almost normally and was only moderately sensitive to oxidative stress when free living. In contrast, its symbiotic properties in alfalfa were drastically affected. Nitrogen-fixing ability was severely impaired. More strikingly, most SOD-deficient bacteria did not reach the differentiation stage of nitrogen-fixing bacteroids. The SOD-deficient mutant nodulated poorly and displayed abnormal infection. After release into plant cells, a large number of bacteria failed to differentiate into bacteroids and rapidly underwent senescence. Thus, bacterial SOD plays a key protective role in the symbiotic process. PMID- 11115111 TI - The virulence plasmid pWR100 and the repertoire of proteins secreted by the type III secretion apparatus of Shigella flexneri. AB - Bacteria of Shigella spp. are the causative agents of shigellosis. The virulence traits of these pathogens include their ability to enter into epithelial cells and induce apoptosis in macrophages. Expression of these functions requires the Mxi-Spa type III secretion apparatus and the secreted IpaA-D proteins, all of which are encoded by a virulence plasmid. In wild-type strains, the activity of the secretion apparatus is tightly regulated and induced upon contact of bacteria with epithelial cells. To investigate the repertoire of proteins secreted by Shigella flexneri in conditions of active secretion, we determined the N-terminal sequence of 14 proteins that are secreted by a mutant in which secretion was deregulated. Sequencing of the virulence plasmid pWR100 of the S. flexneri strain M90T (serotype 5) has allowed us to identify the genes encoding these secreted proteins and suggests that approximately 25 proteins are secreted by the type III secretion apparatus. Analysis of the G+C content and the relative positions of genes and open reading frames carried by the plasmid, together with information concerning the localization and function of encoded proteins, suggests that pWR100 contains blocks of genes of various origins, some of which were initially carried by four different plasmids. PMID- 11115112 TI - Characterization and functional analysis of PorB, a Chlamydia porin and neutralizing target. AB - A predicted protein (CT713) with weak sequence similarity to the major outer membrane protein (20.4% identity) in Chlamydia trachomatis was identified by Chlamydia genome analysis. We show that this protein is expressed, surface accessible, localized to the chlamydial outer membrane complex and functions as a porin. This protein, PorB, was highly conserved among different serovars, with nearly identical sequences between serovars D, B, C and L2. Sequence comparison between C. trachomatis and Chlamydia pneumoniae showed less conservation between species with 59.3% identity. Immunofluorescence staining with monospecific antisera to purified PorB revealed antigen localized within chlamydial inclusions and found throughout the developmental cycle. Antibodies to PorB neutralized infectivity of C. trachomatis in an in vitro neutralization assay confirming that PorB is surface exposed. As PorB was found to be in the outer membrane, as well as having weak structural characteristics similar to major outer membrane protein (MOMP) and other porins, a liposome-swelling assay was used to determine whether this protein had pore-forming capabilities. PorB had pore-forming activity and was shown to be different from MOMP porin activity. PMID- 11115113 TI - Activation of enteropathogenic Escherichia coli (EPEC) LEE2 and LEE3 operons by Ler. AB - Enteropathogenic Escherichia coli (EPEC) produces attaching and effacing lesions (AE) on epithelial cells. The genes involved in the formation of the AE lesions are contained within a pathogenicity island named the locus of enterocyte effacement (LEE). The LEE comprises 41 open reading frames organized in five major operons: LEE1, LEE2, LEE3, LEE4 and tir. The first gene of the LEE1 operon encodes a transcription activator of the other LEE operons that is called the LEE encoded regulator (Ler). The LEE2 and LEE3 operons are divergently transcribed with overlapping -10 promoter regions, and gene fusion studies have shown that they are both activated by Ler. Deletion analysis, using lacZ reporter fusions, of the LEE2 and LEE3 promoters demonstrated that deletions extending closer to the LEE2 transcription start site than -247 bp lead to loss of activation by Ler, whereas only 70 bp upstream of the LEE3 transcription start site is required for Ler-mediated activation. We have purified Ler as a His-tagged protein and used it to perform DNA-binding assays with LEE2 and LEE3. We observed that Ler bound to a DNA fragment containing the -300 to +1 region of LEE2; however, it failed to bind to a DNA fragment containing the -300 to +1 region of LEE3, suggesting that Ler activates both operons by only binding to the regulatory region upstream of LEE2. The Ler-activatable LEE3:lacZ fusions extended to what would be -246 bp of the LEE2 operon. A lacZ fusion from the -300 to +1 region of LEE3 failed to be activated by Ler, consistent with our hypothesis that Ler activates the expression of LEE2 and LEE3 by binding to a region located downstream of the LEE3 transcription start site. DNase I footprinting revealed that Ler protected a region of 121 bp upstream of LEE2. Purified Ler mutated in the coiled-coil domain was unable to activate transcription and to bind to the LEE2 regulatory region. These data indicate that Ler may bind as a multimer to LEE2 and activate both divergent operons by a novel mechanism potentially involving changes in the DNA structure. PMID- 11115114 TI - The txtAB genes of the plant pathogen Streptomyces acidiscabies encode a peptide synthetase required for phytotoxin thaxtomin A production and pathogenicity. AB - Four Streptomyces species have been described as the causal agents of scab disease, which affects economically important root and tuber crops worldwide. These species produce a family of cyclic dipeptides, the thaxtomins, which alone mimic disease symptomatology. Structural considerations suggest that thaxtomins are synthesized non-ribosomally. Degenerate oligonucleotide primers were used to amplify conserved portions of the acyladenylation module of peptide synthetase genes from genomic DNA of representatives of the four species. Pairwise Southern hybridizations identified a peptide synthetase acyladenylation module conserved among three species. The complete nucleotide sequences of two peptide synthetase genes (txtAB) were determined from S. acidiscabies 84.104 cosmid library clones. The organization of the deduced TxtA and TxtB peptide synthetase catalytic domains is consistent with the formation of N-methylated cyclic dipeptides such as thaxtomins. Based on high-performance liquid chromatography (HPLC) analysis, thaxtomin A production was abolished in txtA gene disruption mutants. Although the growth and morphological characteristics of the mutants were identical to those of the parent strain, txtA mutants were avirulent on potato tubers. Moreover, introduction of the thaxtomin synthetase cosmid into a txtA mutant restored both pathogenicity and thaxtomin A production, demonstrating a critical role for thaxtomins in pathogenesis. PMID- 11115115 TI - SdiA, an Escherichia coli homologue of quorum-sensing regulators, controls the expression of virulence factors in enterohaemorrhagic Escherichia coli O157:H7. AB - The quorum-sensing system in bacteria is a well-known regulatory system that controls gene expression in a cell density-dependent manner. A transcriptional regulator (LuxR homologue), signal synthase (LuxI homologue) and autoinducer (acyl homoserine lactone) are indispensable for this system in most Gram-negative bacteria. In this study, we found that SdiA, an Escherichia coli LuxR homologue, is a negative regulator of the expression of virulence factors EspD and intimin in enterohaemorrhagic E. coli (EHEC) O157:H7. The expression of EspD and intimin was inhibited at the RNA level upon SdiA overexpression. SdiA has a DNA-binding motif in its C-terminal part and can bind to the promoter regions of the esp and eae genes in vitro. Extracellular factors, which accumulate in culture supernatants of O157:H7 at the stationary phase of growth and inhibit EspD and intimin synthesis, bind to the N-terminal part of SdiA in vivo and in vitro. O157:H7 overproducing the N-terminal part of SdiA exhibited hypertranscription of EspD and intimin, suggesting that the overproduced N-terminal part had inhibited the activity of intact SdiA through titration of the extracellular factors. These results indicate that a quorum-sensing system including the SdiA protein controls colonization by O157:H7. PMID- 11115116 TI - FNR-dependent activation of the class II dmsA and narG promoters of Escherichia coli requires FNR-activating regions 1 and 3. AB - In Escherichia coli, the anaerobic expression of genes encoding the nitrate (narGHJI) and dimethyl sulphoxide (dmsABC) terminal reductases is stimulated by the global anaerobic regulator FNR. The ability of FNR to activate transcription initiation has been proposed to be dependent on protein-protein interactions between RNA polymerase and two activating regions (AR) of FNR, FNR-AR1 and FNR AR3. To further our understanding of the role of FNR-AR1 and FNR-AR3 in transcription activation, we measured the effects of FNR-AR mutants on expression of the narG and dmsA promoters, PnarG and PdmsA. All the FNR-AR1 (FNR-S73F, FNR T118A, FNR-S187P), FNR-AR3 (FNR-G85A) and FNR-AR1-AR3 (FNR-G85A-S187P) mutants that were tested decreased expression from PnarG and PdmsA in vivo. Transcription assays of PdmsA also showed that the FNR-AR mutant proteins impaired transcription activation in vitro. Furthermore, DNase I footprinting analysis confirmed that this transcription defect was not a result of altered DNA-binding properties. The function of FNR-S187P and FNR-G85A was also measured in strains containing sigma70 mutants (sigma70-K593A, sigma70-R596A and sigma70-K597A) known to be impaired in FNR-dependent transcription activation. Of all of the combinations analysed, only FNR-G85 and sigma70-K597 showed a genetic interaction, supporting the notion that FNR-AR3 and sigma70 interact functionally in the process of transcription activation. Lastly, the transcription activation defect of the FNR-AR1 and FNR-AR3 mutants was greatly reduced when expression of PnarG was assayed in the presence of nitrate. As these growth conditions promote maximal activity of PnarG as a result of the combined function of NarL, IHF and FNR, these results suggest that the requirements for FNR-AR1 and FNR-AR3 are altered in the presence of additional activators. PMID- 11115117 TI - HrpB2 and HrpF from Xanthomonas are type III-secreted proteins and essential for pathogenicity and recognition by the host plant. AB - The interaction between the plant pathogen Xanthomonas campestris pv. vesicatoria and its host plants is controlled by hrp genes (hypersensitive reaction and pathogenicity), which encode a type III protein secretion system. Among type III secreted proteins are avirulence proteins, effectors involved in the induction of plant defence reactions. Using non-polar mutants, we investigated the role of 12 hrp genes in the secretion of the avirulence protein AvrBs3 from X. c. pv. vesicatoria and a heterologous protein, YopE, from Yersinia pseudotuberculosis. Genes conserved among type III secretion systems (hrcQ, hrcR, hrcS and hrcT) as well as non-conserved genes (hrpB1, hrpB2, hrpB4, hrpB5, hrpD5 and hrpD6) were shown to be required for secretion. Protein localization studies using specific antibodies showed that HrpB1 and HrpB4, as well as the putative ATPase HrcN, were mainly found in the soluble fraction of the bacterial cell. In contrast, HrpB2 and HrpF, which is related to NolX of Rhizobium fredii, are secreted into the culture medium in an hrp-dependent manner. As HrpB2, but not HrpF, is essential for type III protein secretion, there might be a hierarchy in the secretion process. We propose that HrpF, which is dispensable for protein secretion but required for AvrBs3 recognition in planta, functions as a translocator of effector proteins into the host cell. PMID- 11115118 TI - Identification of proteases with shared functions to the proprotein processing protease Krp1 in the fission yeast Schizosaccharomyces pombe. AB - Many secretory proteins are synthesized as inactive proproteins that undergo proteolytic activation as they travel through the eukaryotic secretory pathway. The best characterized family of processing enzymes are the prohormone convertases or kexins, and these are responsible for the processing of a wide variety of prohormones and other precursors. Recent work has identified other proteases that appear to be involved in proprotein processing, but characterization of these enzymes is at an early stage. Krp1 is the only kexin identified in the fission yeast Schizosaccharomyces pombe, in which it is essential for cell viability. We have used a genetic screen to identify four proteases with specificities that overlap Krp1. Two are serine proteases, one is a zinc metalloprotease (glycoprotease) and one is an aspartyl protease that belongs to the recently described yapsin family of processing enzymes. All four proteases support the growth of a yeast strain lacking Krp1, and each is able to process the P-factor precursor, the only substrate currently known to be processed by Krp1. PMID- 11115119 TI - Analysis of mRNA decay and rRNA processing in Escherichia coli in the absence of RNase E-based degradosome assembly. AB - We demonstrate here that the assembly of the RNase E-based degradosome of Escherichia coli is not required for normal mRNA decay in vivo. In contrast, deletion of the arginine-rich RNA binding site (ARRBS) from the RNase E protein slightly impairs mRNA decay. When both the degradosome scaffold region and the ARRBS are missing, mRNA decay is dramatically slowed, but 9S rRNA processing is almost normal. An extensive RNase E truncation mutation (rnedelta610) had a more pronounced mRNA decay defect at 37 degrees C than the temperature-sensitive rne-1 allele at 44 degrees C. Taken together, these data suggest that the inviability associated with inactivation of RNase E is not related to defects in either mRNA decay or rRNA processing. PMID- 11115120 TI - Cross-regulation of competence pheromone production and export in the early control of transformation in Streptococcus pneumoniae. AB - Two operons, comAB and comCDE, play a key role in the co-ordination of spontaneous competence development in cultures of Streptococcus pneumoniae. ComAB is required for export of the comC-encoded competence-stimulating peptide (CSP). Upon CSP binding, the histidine kinase ComD activates ComE, its cognate response regulator, required for autoinduction of comCDE and for induction of the late competence genes. To understand better the early control of competence development, mutants upregulating comCDE (ComCDEUP) were isolated using a comC lacZ transcriptional fusion. Mutants were generated by polymerase chain reaction mutagenesis of the comCDE region and by in vitro transposon mutagenesis of the chromosome. Both types of ComCDEUP mutants exhibited similar phenotypes. They differed from wild type in displaying trypsin-resistant transformation, competence under acid growth conditions and expression of comCDE under microaerobiosis; increased production of CSP in the mutants could account for the various phenotypes. The ComCDEUP transposon mutations included four independent insertions in the ciaR gene, which encodes the response regulator of a two component system previously found to affect competence, and two immediately upstream of the comAB operon. The latter two resulted in comAB overexpression, indicating that CSP export is rate limiting. Among comDE point mutations, a single amino acid change in ComD (T233I) conferred constitutive, CSP-independent competence and resulted in comAB overexpression, providing support for the hypothesis that ComE regulates comAB; a ComE mutant (R120S) exhibited altered kinetics of competence shut-off. Collectively, these data indicate that pheromone autoinduction, cross-regulation of the comAB and comCDE operons and, possibly, competence shut-off contribute to the early control of competence development in S. pneumoniae. They argue for a metabolic control of competence, mediated directly or indirectly by CiaR, and they suggest that both comAB and comCDE are potential targets for regulation. PMID- 11115121 TI - A Rox1-independent hypoxic pathway in yeast. Antagonistic action of the repressor Ord1 and activator Yap1 for hypoxic expression of the SRP1/TIR1 gene. AB - Hypoxic SRP1/TIR1 gene expression depends on the absence of haem but is independent of Rox1-mediated repression. We have found a new hypoxic pathway involving an antagonistic interaction between the Ixr1/Ord1 repressor and the Yap1 factor, a transcriptional activator involved in oxidative stress response. Here, we show that Ord1 repressed SRP1 gene expression under normoxia and hypoxia, whereas Yap1 activated it. Ord1 and Yap1 have been shown to bind the SRP1 promoter in a region extending from -299 to -156 bp upstream of the start codon. A typical AP-1 responsive element lying from -247 to -240 bp allows Yap1 binding. Internal deletion of sequences within the SRP1 promoter were introduced. Two regions were characterized at positions -299/-251 and -218/-156 that, once removed, resulted in a constitutive expression of SRP1 in a wild-type strain under normoxic conditions. Deletion of both these two sequences allowed the bypass of YAP1 requirement in a Deltayap1 strain, whereas these two internal deletions did not yield increased expression in a Deltaord1 strain compared with the full-length promoter. Both a single Deltaord1 mutant and a doubly disrupted Deltayap1 Deltaord1 strain yielded normoxic constitutive SRP1 expression and increased hypoxic SRP1 induction, thereby demonstrating that ord1 is epistatic to yap1. Thus, Yap1 is not directly involved in SRP1 induction by hypoxia, but is necessary to counteract the Ord1 effect. PMID- 11115122 TI - Topological analysis and role of the transmembrane domain in polar targeting of PilS, a Pseudomonas aeruginosa sensor kinase. AB - In Pseudomonas aeruginosa, synthesis of pilin, the major protein subunit of the pili, is regulated by a two-component signal transduction system in which PilS is the sensor kinase. PilS is an inner membrane protein found at the poles of the bacterial cell. It is composed of three domains: an N-terminal hydrophobic domain; a central cytoplasmic linker region; and the C-terminal transmitter region conserved among other sensor kinases. The signal that activates PilS and, consequently, pilin transcription remains unknown. The membrane topology of the hydrophobic domain was determined using the lacZ and phoA gene fusion approach. In this report, we describe a topological model for PilS in which the hydrophobic domain forms six transmembrane helices, whereas the N- and C-termini are cytoplasmic. This topology is very stable, and the cytoplasmic C-terminus cannot cross the inner membrane. We also show that two of the six transmembrane segments are sufficient for membrane anchoring and polar localization of PilS. PMID- 11115123 TI - Proton motive force drives the interaction of the inner membrane TolA and outer membrane pal proteins in Escherichia coli. AB - The Tol-Pal system of the Escherichia coli envelope is formed from the inner membrane TolQ, TolR and TolA proteins, the periplasmic TolB protein and the outer membrane Pal lipoprotein. Any defect in the Tol-Pal proteins or in the major lipoprotein (Lpp) results in the loss of outer membrane integrity giving hypersensitivity to drugs and detergents, periplasmic leakage and outer membrane vesicle formation. We found that multicopy plasmid overproduction of TolA was able to complement the membrane defects of an lpp strain but not those of a pal strain. This result indicated that overproduced TolA has an envelope-stabilizing effect when Pal is present. We demonstrate that Pal and TolA formed a complex using in vivo cross-linking and immunoprecipitation experiments. These results, together with in vitro experiments with purified Pal and TolA derivatives, allowed us to show that Pal interacts with the TolA C-terminal domain. We also demonstrate using protonophore, K+ carrier valinomycin, nigericin, arsenate and fermentative conditions that the proton motive force was coupled to this interaction. PMID- 11115124 TI - Membrane localization of a rice calcium-dependent protein kinase (CDPK) is mediated by myristoylation and palmitoylation. AB - Calcium-dependent protein kinases (CDPKs), the most abundant serine/threonine kinases in plants, are found in various subcellular localizations, which suggests that this family of kinases may be involved in multiple signal transduction pathways. A complete analysis to try to understand the molecular basis of the presence of CDPKs in various localizations in the cell has not been accomplished yet. It has been suggested that myristoylation may be responsible for membrane association of CDPKs. In this study, we used a rice CDPK, OSCPK2, which has a consensus sequence for myristoylation at the N-terminus, to address this question. We expressed wild-type OSCPK2 and various mutants in different heterologous systems to investigate the factors that affect its membrane association. The results show that OSCPK2 is myristoylated and palmitoylated and targeted to the membrane fraction. Both modifications are required, myristoylation being essential for membrane localization and palmitoylation for its full association. The fact that palmitoylation is a reversible modification may provide a mechanism for regulation of the subcellular localization. OSCPK2 is the first CDPK shown to be targeted to membranes by an src homology domain 4 (SH4) located at the N-terminus of the molecule. PMID- 11115125 TI - A novel aldose/aldehyde reductase protects transgenic plants against lipid peroxidation under chemical and drought stresses. AB - Rapid accumulation of toxic products from reactions of reactive oxygen species (ROS) with lipids and proteins significantly contributes to the damage of crop plants under biotic and abiotic stresses. Here we have identified a stress activated alfalfa gene encoding a novel plant NADPH-dependent aldose/aldehyde reductase that also exhibited characteristics of the homologous human enzyme. The recombinant alfalfa enzyme is active on 4-hydroxynon-2-enal, a known cytotoxic lipid peroxide degradation product. Ectopic synthesis of this enzyme in transgenic tobacco plants provided considerable tolerance against oxidative damage caused by paraquat and heavy metal treatment. These transformants could also resist a long period of water deficiency and exhibited improved recovery after rehydration. We found a reduced production of lipid peroxidation-derived reactive aldehydes in these transformed plants under different stresses. These studies reveal a new and efficient detoxification pathway in plants. PMID- 11115126 TI - Cloning and functional analysis of chicory root fructan1-exohydrolase I (1-FEH I): a vacuolar enzyme derivedfrom a cell-wall invertase ancestor? Mass fingerprint of the 1-FEH I enzyme. AB - This paper describes the cloning and functional analysis of chicory (Cichorium intybus L.) fructan 1-exohydrolase I cDNA (1-FEH I). To our knowledge it is the first plant FEH cloned. Full-length cDNA was obtained by a combination of RT-PCR, 5' and 3' RACE using primers based on N-terminal and conserved amino acid sequences. Electrophoretically purified 1-FEH I enzyme was further analyzed by in gel trypsin digestion followed by matrix-assisted laser desorption ionization and electrospray time-of-flight tandem mass spectrometry. Functionality of the cDNA was demonstrated by heterologous expression in potato tubers. 1-FEH I takes a new, distinct position in the phylogenetic tree of plant glycosyl hydrolases being more homologous to cell-wall invertases (44-53%) than to vacuolar invertases (38-41%) and fructosyl transferases (33-38%). The 1-FEH I enzyme could not be purified from the apoplastic fluid at significantly higher levels than can be explained by cellular leakage. These and other data suggest a vacuolar localization for 1-FEH I. Also, the pI of the enzyme (6.5) is lower than expected from a typical cell-wall invertase. Unlike plant fructosyl transferases that are believed to have evolved from a vacuolar invertase, 1-FEH I might have evolved from a cell-wall invertase-like ancestor gene that later obtained a vacuolar targeting signal. 1-FEH I mRNA quantities increase in the roots throughout autumn, and especially when roots are stored at low temperature. PMID- 11115127 TI - MADS-box gene evolution beyond flowers: expression in pollen, endosperm, guard cells, roots and trichomes. AB - MADS-box genes encode transcriptional regulators involved in diverse aspects of plant development. Here we describe the cloning and mRNA spatio-temporal expression patterns of five new MADS-box genes from Arabidopsis: AGL16, AGL18, AGL19, AGL27 and AGL31. These genes will probably become important molecular tools for both evolutionary and functional analyses of vegetative structures. We mapped our data and previous expression patterns onto a new MADS-box phylogeny. These analyses suggest that the evolution of the MADS-box family has involved a rapid and simultaneous functional diversification in vegetative as well as reproductive structures. The hypothetical ancestral genes had broader expression patterns than more derived ones, which have been co-opted for putative specialized functions as suggested by their expression patterns. AGL27 and AGL31, which are closely related to the recently described flowering-time gene FLC (previously AGL25), are expressed in most plant tissues. AGL19 is specifically expressed in the outer layers of the root meristem (lateral root cap and epidermis) and in the central cylinder cells of mature roots. AGL18, which is most similar in sequence to the embryo-expressed AGL15 gene, is expressed in the endosperm and in developing male and female gametophytes, suggesting a role for AGL18 that is distinct from previously characterized MADS-box genes. Finally, AGL16 RNA accumulates in leaf guard cells and trichomes. Our new phylogeny reveals seven new monophyletic clades of MADS-box sequences not specific to flowers, suggesting that complex regulatory networks involving several MADS-box genes, similar to those that control flower development, underlie development of vegetative structures. PMID- 11115128 TI - Fatty acid ketodienes and fatty acid ketotrienes: Michael addition acceptors that accumulate in wounded and diseased Arabidopsis leaves. AB - Physical damage and disease are known to lead to changes in the oxylipin signature of plants. We searched for oxylipins produced in response to both wounding and pathogenesis in Arabidopsis leaves. Linoleic acid 9- and 13 ketodienes (KODEs) were found to accumulate in wounded leaves as well as in leaves infected with the pathogen Pseudomonas syringae pv. tomato (Pst). Quantification of the compounds showed that they accumulated to higher levels during the hypersensitive response to Pst avrRpm1 than during infection with a Pst strain lacking an avirulence gene. KODEs are Michael addition acceptors, containing a chemically reactive alpha,beta-unsaturated carbonyl group. When infiltrated into leaves, KODEs were found to induce expression of the GST1 gene, but vital staining indicated that these compounds also damaged plant cells. Several molecules typical of lipid oxidation, including malonaldehyde, also contain the alpha,beta-unsaturated carbonyl reactivity feature, and, when delivered in a volatile form, powerfully induced the expression of GST1. The results draw attention to the potential physiological importance of naturally occurring Michael addition acceptors in plants. In particular, these compounds could act directly, or indirectly via cell damage, as powerful gene activators and might also contribute to host cell death. PMID- 11115129 TI - The gapped xylem mutant identifies a common regulatory step in secondary cell wall deposition. AB - The phenotype of the novel gapped xylem (gpx) mutant is described. gpx plants exhibit gaps in the xylem in positions where xylem elements would normally be located. These gaps are not part of the transpiration stream and result in gpx plants having fewer functional xylem elements. The gaps are due to the absence of a secondary cell wall in developing xylem elements, resulting in complete degradation of these elements during cell death, and illustrate the importance of the secondary cell wall in retaining a functional xylem element following programmed cell death. Consequently the gpx phenotype suggests that the processes of secondary cell wall formation and cell death are independently regulated in developing xylem. gpx plants also exhibit a highly irregular pattern of secondary cell wall thickening in interfascicular cells, with some cells apparently undergoing little or no secondary cell wall deposition. Secondary cell wall deposition in plants involves the co-ordinate regulation of several complex metabolic pathways. The gpx mutant identifies a key step involved in regulating the deposition of secondary cell wall material in both xylem and interfascicular cells, and suggests that a common regulatory step controls secondary cell wall formation in these diverse cell types. The gpx mutant offers a unique opportunity to elucidate the mechanism by which the complex processes involved in secondary cell wall formation are co-ordinately regulated. PMID- 11115130 TI - A member of a novel Arabidopsis thaliana gene family of candidate Mg2+ ion transporters complements a yeast mitochondrial group II intron-splicing mutant. AB - Autocatalytic activity of some group II introns has been demonstrated in vitro, but helper functions such as the yeast MRS2 protein are essential for splicing in vivo. In our search for such helper factors in plants, we pursued the cloning of two Arabidopsis thaliana homologues, atmrs2-1 and atmrs2-2. Atmrs2-1, but not atmrs2-2, complements the yeast deletion mutant of mrs2, and this is congruent with the prediction of two adjacent transmembrane stretches in AtMRS2-1 and yeast MRS2 but not in AtMRS2-2. This complementation depends on fusion of the native yeast mitochondrial import sequence to atmrs2-1. A differing, non-mitochondrial, cellular targeting in Arabidopsis is supported by the analysis of green fluorescent protein fusion constructs after transient transformation into plant protoplasts. Further members of what now appears to be a family of 10 mrs2 homologues are identified in the Arabidopsis genome. Similarity searches with the PSI-BLAST algorithm in the protein database fail to identify homologues of this novel gene family in any eukaryotes other than yeasts, but do identify its distant relatedness to the corA group of bacterial magnesium transporters. In line with this observation, intramitochondrial magnesium concentrations are indeed restored to wild-type levels in the yeast mutant on complementation with atmrs2-1. PMID- 11115131 TI - Plant sucrose-H+ symporters mediate the transport of vitamin H. AB - A cDNA coding for a vitamin H (biotin) transport protein from Arabidopsis was identified by genetic complementation of a biotin uptake-deficient yeast mutant. Vitamin H transport by this protein was sensitive to the SH-group inhibitor p chloromercuribenzene sulfonic acid (PCMBS) and to the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP), suggesting an energy-dependent biotin-H+ symport mechanism. The transport activity could contribute to the so-far uncharacterized plant sucrose-H+ symporter AtSUC5 which mediates the energy-dependent transport of biotin and sucrose, and restores growth of the biotin transport-deficient yeast mutant on medium with low biotin concentrations. Functional comparison of the AtSUC5 transporter with previously characterized plant sucrose or monosaccharide transporters revealed that biotin transport may be a general and specific property of all plant sucrose transporters (sucrose/biotin-H+ symporters). This first report on a transporter with dual substrate specificity for two structurally unrelated molecules has a major impact on general thinking concerning the specificity of membrane transporters. The physiological relevance of this finding is discussed. PMID- 11115133 TI - Chloroplast signalling in the light induction of nuclear HSP70 genes requires the accumulation of chlorophyll precursors and their accessibility to cytoplasm/nucleus. AB - Chlorophyll precursors Mg-protoporphyrin IX and its monomethylester are candidates for plastid-derived molecules involved in light signalling from the chloroplast to the nucleus. The pool sizes of these two Mg2+-containing porphyrins and of protoporphyrin IX transiently increased upon a shift of Chlamydomonas cultures from dark to light. This increase coincided with the accumulation of mRNAs encoded by the nuclear genes HSP70A and HSP70B. Analysis of a mutant (brs-1), previously shown to be defective in the light induction of these genes, revealed high levels of protoporphyrin IX but no light-induced increase in the levels of Mg2+-containing porphyrins. Inhibitors of cytoplasmic protein synthesis prevented both the light-induced rise in pool levels and induction of the HSP70 genes. Similarly, pre-gametes, intermediates of sexual differentiation, lacked both responses to light. The block in light induction of the HSP70 genes in inhibitor-treated cells and in pre-gametes could be circumvented by the exogenous addition of Mg-protoporphyrin IX in the dark. This suggests an essential role for light-induced Mg-protoporphyrin IX accumulation in this chloroplast-to-nucleus signalling pathway. However, accumulation of this porphyrin in the dark - presumably in the chloroplast - did not result in induction. A second crucial role for light in this signalling pathway is postulated which makes this plastidic compound accessible to the cytoplasm/nucleus where the downstream signalling pathway may be activated. PMID- 11115132 TI - Na+/myo-inositol symporters and Na+/H+-antiport in Mesembryanthemum crystallinum. AB - Mitr1 and Mitr2 from Mesembryanthemum crystallinum (common ice plant) are members of a family of genes homologous to H+[or Na+]/myo-inositol symporters (ITRs), not previously studied in plants. MITR1 complemented an Itr1-deficient yeast strain. Mitr1 is strongly expressed in roots, moderately in stems, and weakly in leaves. Its transcripts increased in all organs, most dramatically in roots, under salinity stress. Mitr2 constitutes a rare transcript, slightly upregulated by salt stress in leaves only. Mitr1 transcripts are present in all cells in the root tip, but become restricted to phloem-associated cells in mature roots. Peptide antibodies against the two proteins indicated the presence of MITR1 in all organs and of MITR2 in leaves. Both are located in the tonoplast. MITR1 acts in removing sodium from root vacuoles, correlated with findings of low root sodium, while leaf vacuoles accumulate sodium in the ice plant. Up-regulation in leaves and stems is also found for Na+/H+-antiporter (Nhx-type) transcripts. Under comparable stress conditions, Nhx-and Itr-like transcripts in Arabidopsis were regulated differently. In the ice plant, co-ordinate induction of Na+/H+ antiporters and Na+/myo-inositol symporters transfers sodium from vacuoles in root cells into the leaf mesophyll as a halophytic strategy that lowers the osmotic potential. The tissue-specific differential expression of Itr- and Nhx type transcripts suggests that the vacuolar sodium/inositol symporters function to reduce sodium amounts in cells of the root and vascular tissue, while sodium/proton antiporters in leaf tissues function to partition sodium into vacuoles for storage. PMID- 11115134 TI - Expression of a truncated tobacco NtCBP4 channel in transgenic plants and disruption of the homologous Arabidopsis CNGC1 gene confer Pb2+ tolerance. AB - Recently we reported on a plasma membrane tobacco protein (designated NtCBP4) that binds calmodulin. When overexpressed in transgenic plants, NtCBP4 confers Pb2+ hypersensitivity associated with enhanced accumulation of this toxic metal. To further investigate possible modulation of Pb2+ tolerance in plants, we prepared transgenic plants that express a truncated version of this protein (designated NtCBP4DeltaC) from which its C-terminal, with the calmodulin-binding domain and part of the putative cyclic nucleotide-binding domain, was removed. In contrast to the phenotype of transgenic plants expressing the full-length gene, transgenic plants expressing the truncated gene showed improved tolerance to Pb2+, in addition to attenuated accumulation of this metal. Furthermore, disruption by T-DNA insertion mutagenesis of the Arabidopsis CNGC1 gene, which encodes a homologous protein, also conferred Pb2+ tolerance. We suggest that NtCBP4 and AtCNGC1 are components of a transport pathway responsible for Pb2+ entry into plant cells. PMID- 11115135 TI - Technical advance: an aniline blue staining procedure for confocal microscopy and 3D imaging of normal and perturbed cellular phenotypes in mature Arabidopsis embryos. AB - A new method is described for fluorescent imaging of mature Arabidopsis embryos that enables their cellular architecture to be visualized without the need for histological sectioning. Mature embryos are stained with aniline blue and cleared with chloral hydrate to allow high-resolution confocal imaging of individual cells within the embryo prior to germination. The technique allows the collection of longitudinal optical sections throughout the cotyledon, hypocotyl and root of wild-type Arabidopsis C24 embryos. Every cell within the mature embryo can be visualized with sufficient clarity and resolution to allow three-dimensional analysis of cellular architecture. Optical sectioning of mutant gnom, short-root and scarecrow embryos, and through root meristems disrupted as a consequence of targeted misexpression of diphtheria toxin, demonstrate the potential of this technique for visualizing the cellular organization of mutant and perturbed embryonic phenotypes. PMID- 11115136 TI - Technical advance: application of high-performance liquid chromatography with photodiode array detection to the metabolic profiling of plant isoprenoids. AB - The application of high-performance liquid chromatography (HPLC) using a C30 reverse-phase stationary matrix has enabled the simultaneous separation of carotenes, xanthophylls, ubiquinones, tocopherols and plastoquinones in a single chromatogram. Continuous photodiode array (PDA) detection ensured identification and quantification of compounds upon elution. Applications of the method to the characterization of transgenic and mutant tomato varieties with altered isoprenoid content, biochemical screening of Arabidopsis thaliana, and elucidation of the modes of action of bleaching herbicides are described to illustrate the versatility of the procedure. PMID- 11115137 TI - Of condoms and conundrums. PMID- 11115138 TI - A synthesis of qualitative home visiting research. AB - Over the past decade, a body of qualitative research has been developed which describes the home visiting practice of public health nurses (PHNs) to maternal child clients. This article reports a synthesis of these studies. The purpose of the synthesis was to identify common elements and differences between the research reports that would lead to theory development or support of existing theories. Methods were based on Miles and Huberman's (1994) text on qualitative data analysis. Results of the synthesis indicated that building and preserving relationships with the client is the central focus of home visiting and provides a foundation for problem identification and problem solving. Clients control access to their homes as well as the information they are willing to share with the nurse. The goals of home visiting relate to empowering mothers, supporting their independence and decision making. Similarities to Peplau's theory of Interpersonal Relations and Cox's Interaction Model of Client Health Behavior (IMCHB) are noted. PMID- 11115139 TI - Prenatal risk factors among foreign-born Central American women: a comparative study. AB - The purpose of this study was to compare the incidence of empirically established prenatal risk factors for low birthweight (LBW) outcomes among two groups of low income mothers: foreign-born Central American women and nonimmigrant, non Hispanic women. Two hundred ninety-six women who were part of a larger study of maternal role sufficiency were included in the present study: 127 Central American women and 169 nonimmigrant, non-Hispanic women who identified themselves as Black (n = 59) or White (n = 110). Data were collected by public health nurses (PHNs) during home visits and by research nurses in prenatal health department clinics. Comparisons were made between the two groups in areas of demographic characteristics, prenatal health behaviors, and prenatal stressful life-events. Foreign-born Central American mothers were found to be less educated, more likely to be living with their partners, less likely to engage in prenatal health risk behaviors, and less likely to identify stressors in their lives. The initiation of prenatal PHN services by the target group was similar to the comparison group. Their rate of LBW deliveries did not reflect the protective effect often attributed to foreign-born Hispanic mothers. Findings are discussed in light of the paradox of LBW and Hispanic heritage. Recommendations for practice, clinical research, and public policy are also addressed. PMID- 11115140 TI - Gender differences in teen parents' perceptions of parental responsibilities. AB - Accurate information concerning teen parents' knowledge of their children's development and their expectations for paternal involvement becomes increasingly important as efforts increase to promote involvement of unmarried fathers with their children. The purpose of this descriptive study was to assess differences in the knowledge and perceptions of normal child development, and expectations for paternal responsibilities between unmarried, low-income African American and Mexican American adolescent mothers and their males partners. Seven unmarried adolescent mothers participated in a focus group interview held at a family service agency in the Midwest. Afterward, their male partners and reported fathers of their babies, participated in a separate focus group interview The mean age of the adolescent mother participants was 16.7 years, the mean age of their partners was 19.3 years. Data were analyzed using a tape-based analysis method. A number of differences were identified between the perceptions of the adolescent mothers and young fathers including their level of child development knowledge, context for selecting physical methods of discipline, expectations for paternal role behaviors, and feelings about child support payments and establishing legal paternity. The study findings may help health care providers develop more effective prenatal and parenting educational experiences for adolescent parents. PMID- 11115141 TI - Mothers' explanatory models of lack of child growth. AB - This qualitative study elicited the explanatory models (EMs) of child growth held by mothers of growth-deficient children. EMs are culturally constructed explanations for a specific illness and its treatment (Kleinman, 1980). The EM concept was adapted for this study to focus on a child health condition instead of an illness. The sample comprised 22 mothers of growth deficient children who were interviewed 2 years after the conclusion of an intervention study to promote child growth. Growth deficiency was defined as below the 10th percentile for weight, height, or weight for height on the National Center for Health Statistics (NCHS) growth grids (Hamill, Drzid, Johnson, Reed, & Roche, 1976). Three major domains were identified in the EMs of growth held by mothers: (1) illness or heredity (etiology); (2) keeping track of growth (course); and (3) helping my child grow (treatment). The mothers in this study were concerned about their children's size and growth patterns and they monitored their children's growth with the methods available to them. They identified illnesses and allergies as environmental factors that negatively impact their children's growth. All mothers viewed size as a function of heredity. The findings from this study suggest that an emphasis on size will not encourage mothers to focus on their children's growth. The EMs for growth and size were different. Health care providers may be more effective in enhancing children's growth by teaching parents how to deal with the day-to-day problems of children who are picky eaters, stretching limited food money, creating mealtime schedules, and dealing with illnesses before they become severe. PMID- 11115142 TI - An evaluation of interventions to decrease intimate partner violence to pregnant women. AB - Prevention of abuse to women is a national priority; however, research has focused on identification of abuse rather than evaluating interventions. To evaluate the differential effectiveness of three levels of intervention, Brief, Counseling, and Outreach, a longitudinal study with repeated evaluation interviews at 2-, 6-, 12-, and 18-months postdelivery was completed at two urban public health prenatal clinics. The participants were 329 pregnant, physically abused Hispanic women. Both physical abuse and women's use of community resources were measured. Repeated measures ANOVA showed that severity of abuse decreased significantly (p < 0.001) across time for all intervention groups. Violence scores at 2-months postdelivery were significantly lower for the Outreach group (p < 0.05) compared to the Counseling only group, but not significantly lower than the Brief intervention group. At 6-, 12-, and 18-month follow-up there were no statistically significant differences among the intervention groups. The use of lay outreach for abused pregnant women merits further research. Abuse screening by itself, however, may be the most effective intervention to prevent abuse to pregnant women. PMID- 11115143 TI - The effects of community health nurse monitoring on hypertension identification and control. AB - Identification and control of hypertension are important public health concerns. Lack of regular health care makes diagnosis and control difficult in some populations. Community health nursing services in settings where clients regularly congregate promote identification and control of hypertension. This study focused on the effects of community health nursing screening and monitoring services for hypertension provided to participants in a breadline, a senior nutrition program, an English as Second Language (ESL) class, and employees providing these services. Community health nurses (CHNs) provided 2,407 blood pressure-related service encounters. Blood pressures were elevated in 19% of encounters, and 10% of clients had elevations that warranted referral for medical assistance. At the end of the 18-month study period, 67% of all clients with elevations and 71% of those referred for medical assistance had achieved normal blood pressures. One-way analysis of variance indicated a significant relationship between the number of encounters with the nurses and a positive outcome for all clients with elevations. This relationship was not supported for those clients referred to medical assistance. The effectiveness of intervention appeared to vary somewhat among subgroups with some groups more likely than others to achieve a positive outcome. Group differences in outcome were not statistically significant. PMID- 11115144 TI - Attitudes, subjective norm, perceived behavioral control, and intentions related to adult smoking cessation after coronary artery bypass graft surgery. AB - The purpose of this study was to examine the relationships between intention, attitudes, subjective norm, and perceived behavioral control related to smoking cessation in adults after initial coronary artery bypass graft surgery (CABG). The theoretical framework for the study was derived from Ajzen's Theory of Planned Behavior. Intention, the global and belief-based measures of attitude, subjective norm, and perceived behavioral control were measured with the Determinants of Adult Smoking Cessation (DOASC) Questionnaire developed by the investigator. Thirty-two adult smokers completed the questionnaire 2 to 3 weeks following hospital discharge. Four weeks after the questionnaire completion, a follow-up telephone call was used to determine the participants' current smoking status. The study results indicated that there was a statistically significant relationship between the intention to quit smoking after CABG and the global measure of attitude, and perceived behavioral control. This study highlights some of the beliefs about the outcomes of quitting smoking permanently after CABG which may underlie attitudes, subjective norm, and perceived behavioral control in this population. Implications for theory, practice, and research are discussed. PMID- 11115145 TI - Home care for the frail elderly based on urinary incontinence level. AB - Incontinence is a common problem in the frail elderly. We conducted interviews focusing on urinary incontinence with 249 elderly clients in the home care setting, and studied differences of the needs among three (mild, moderate, and catheter) groups based on incontinence level. The mild group had the highest number of professional care needs, although their problems were not as serious as the other two groups. The moderate group required the highest amount of daily care by caregiver. A similar need pattern was shown in the moderate and catheter groups, while more diversified needs were required in the mild group. Portable toilet, rehabilitation, and short-stay services were frequently used in the mild group. The use of telephone consultation was the highest in the moderate group, and the use of doctor visit and bathing service were higher in the catheter group. The most important challenge was significantly different in each group: preventive efforts to maintain activities of daily living (ADL) in the mild group, interpersonal relationships in the moderate group, and infection control in the catheter group. Education was necessary for caregivers in all three groups. These findings help to project realistic care needs for each client based on his or her incontinence level. PMID- 11115146 TI - Factors associated with cane use among community dwelling older adults. AB - Guided by the Theory of Planned Behavior (TPB), this study examined factors associated with cane use among community dwelling older adults. Data were collected in a cross-sectional survey of a convenience sample of 106 community residing older adults in Ottawa, Canada. Using a stepwise discriminant analysis, subjective norms, attitudes, and age surfaced as the key variables associated with cane use in this sample. The discriminant function accounted for 67% of the variance in cane use and correctly classified 91% of cases (Wilks's lambda = 0.33, lambda2 = 110.12, df = 3, p < 0.0001). The findings provide evidence for the utility of the TPB in its application to understanding cane use behaviors of older persons and have important implications for the design of theory-based fall prevention interventions to enhance the acceptance and effective use of mobility aids. PMID- 11115147 TI - Classifications for wound bed preparation and stimulation of chronic wounds. PMID- 11115148 TI - Lactate elicits vascular endothelial growth factor from macrophages: a possible alternative to hypoxia. AB - Macrophages respond to various stimuli to produce angiogenic factors but few mechanistic details are known. We examined the effects of hypoxia, lactate and nicotinamide on the expression of vascular endothelial growth factor by cultured macrophages. These agents were chosen because they down-regulate polyadenosine diphosphoribose levels. Following exposure, conditioned media were analyzed for vascular endothelial growth factor protein. Nicotinamide adenine dinucleotide, polyadenosine diphosphoribose, and vascular endothelial growth factor mRNA were measured in the cellular fraction. Angiogenic capacity of the conditioned media was tested in rabbit corneas and Matrigel implants. All three agents, hypoxia, lactate and nicotinamide, elicited significantly increased levels of vascular endothelial growth factor mRNA and vascular endothelial growth factor in the conditioned media, and these levels were paralleled by their angiogenic activity. Polyadenosine diphosphoribose in the cellular fraction was correspondingly depressed. Anti-vascular endothelial growth factor antibody inhibited most of the angiogenic response whereas anti-basic fibroblast growth factor antibody had little effect. We propose that redox changes associated with the alteration of cellular nicotinamide adenine dinucleotide and polyadenosine diphosphoribose are involved in lactate-mediated VEGF expression. PMID- 11115150 TI - Outgrowth of chondrocytes from human articular cartilage explants and expression of alpha-smooth muscle actin. AB - The objectives of this study were to investigate the effect of various enzymatic treatments on the outgrowth of chondrocytes from explants of adult human articular cartilage and the expression of a specific contractile protein isoform, alpha-smooth muscle actin, known to facilitate wound closure in other connective tissues. Explants of articular cartilage were prepared from specimens obtained from patients undergoing total joint arthroplasty. The time to cell outgrowth in vitro was determined and the expression of alpha-smooth muscle actin shown by immunohistochemistry. Treatment of the explants with collagenase for 15 minutes reduced the time to outgrowth from more than 30 days to 3 days. Hyaluronidase, chondroitinase ABC, and trypsin applied for the 15-minute period had no effect on the time to cell outgrowth when compared with untreated controls. Pretreatment with hyaluronidase prior to collagenase reduced the time to outgrowth. A notable finding of this study was that the majority of chondrocytes in the adult human articular cartilage specimens and virtually all of the outgrowing cells contained alpha-smooth muscle actin. We conclude that human articular chondrocytes have the capability to migrate through enzymatically degraded matrix and express a contractile actin isoform. Collagenase treatment reduces the time required for cell outgrowth. PMID- 11115149 TI - Chemokine and chemokine receptor expression in keloid and normal fibroblasts. AB - Keloids are benign collagenous tumors that occur during dermal wound healing in genetically predisposed individuals. The lesions are characterized by over proliferation of fibroblasts, some leukocyte infiltration, and prolonged high rates of collagen synthesis. To determine whether leukocyte chemoattractants or chemokines are participating in this disease process, immunohistochemical staining for the CXC chemokine, MGSA/GROalpha, and its receptor, CXCR2, was performed on tissue from keloids, hypertrophic scars and normal skin. Immunoreactive MGSA/GROalpha was not observed in hypertrophic scars or normal dermis, but was present in some myofibroblasts and lymphocytes in nodular areas of the keloid samples. This staining positively correlated with the degree of inflammatory infiltrate in the lesions. Keloids, but not hypertrophic scars or normal dermis, also exhibited intensive immunoreactivity for the CXCR2 receptor in endothelial cells and inflammatory infiltrates with occasional staining of myofibroblasts. In contrast, cultured fibroblasts from either keloids or normal skin did not express detectable amounts of mRNA for MGSA/GRO or CXCR2, although interleukin-1 strongly induced MGSA/GRO mRNA in both cell types. Interleukin-1 induction of MGSA/GRO was inhibited by glucocorticoid in normal and keloid fibroblasts, and the effect was more pronounced in keloid fibroblasts. This event was not correlated with inhibition of nuclear activation of NF-kappaB, AP-1 or Sp1, and might therefore be mediated by another mechanism such as decreased mRNA stability or transcriptional repression through the glucocorticoid response element in the MGSA/GRO promoter. Data from in vitro wounding experiments with cultured normal and keloid fibroblasts indicate that there were no significant differences in MGSA/GRO or CXCR2 receptor levels between normal and keloid fibroblasts. We also show that cultured keloid fibroblasts exhibit a delayed wound healing response. We postulate that the inflammatory component is important in development of keloid lesions and chemotactic cytokines may participate in this process. PMID- 11115151 TI - Proteinases, their inhibitors, and cytokine profiles in acute wound fluid. AB - Wound healing is a complex process involving the interactions of many different cell types, matrix components and biological factors, including proteinases and cytokines. This study compared the levels of proteinases (matrix metalloproteinases and plasminogen activators), proteinase inhibitors (tissue inhibitors of metalloproteinases and plasminogen activator inhibitors), inflammatory cytokines and growth factors in acute wound fluid samples collected from the surgical drains of elective breast (n = 24) and colorectal (n = 26) patients on the first postoperative day. Gelatin zymography was used to determine matrix metalloproteinase-2 and -9 levels, quenched fluorescence substrate hydrolysis was applied for total MMP activity and enzyme-linked immunoassays were used to quantitate other factors. Colorectal wound fluid samples showed significantly (p < 0.05) greater levels of the matrix metalloproteinases (MMP-1, 2, 3, and 9), tissue inhibitor of metalloproteinases-1, urokinase plasminogen activator receptor and the inflammatory cytokines (interleukin-1beta, -6, and tumor necrosis factor-alpha); e.g., matrix metalloproteinase-3 colon; median 275 (range 11-2.530) ng/ml; breast; 530-400. However, tissue plasminogen activator and growth factor levels (epidermal growth factor, platelet-derived growth factor, basic fibroblast growth factor, transforming growth factor-beta1) were significantly greater in breast samples; e.g., epidermal growth factor breast 468 (103-1, 444) pg/ml; colon 57(1-573). There was no difference in the levels of urokinase type plasminogen activator, plasminogen activator inhibitor-1 and -2, tissue inhibitor of metalloproteinases -2 or vascular endothelial growth factor. Acute wound fluid from different surgical wounds showed different profiles of proteinases, proteinase inhibitors, and cytokines. This may lead to differences in the rate of tissue remodeling and therefore healing in these two wounds in vivo. PMID- 11115152 TI - A sense phosphorothioate oligodeoxynucleotide containing the transforming growth factor beta regulatory element acts as a novel local nonsteroidal antifibrotic drug. AB - Fibrosis is a potential response to tissue injury. At present, glucocorticoids with their numerous toxic side effects are the only effective treatment for fibrotic diseases. Granulomas induced by sponge implantation were treated with single-stranded phosphorothioate oligodeoxynucleotides containing the wild type or mutated transforming growth factor-beta response element designed to inhibit the rat proalpha1(I) promoter activity. Single-stranded phosphorothioate oligonucleotides resulted in antifibrotic activity based on their ability to reduce granuloma tissue formation and selectively inhibit collagen synthesis. The mutated single-standed phosphorothioate oligonucleotides or dexamethasone given at an equivalent dose to single-standed phosphorothioate oligonucleotides failed to do so. These data suggest that the phosphorothioate oligodeoxynucleotide containing the transforming growth factor-beta regulatory element has an antifibrotic effect and may be used to inhibit the development of fibrosis. PMID- 11115153 TI - Effective treatment of oral erosive lichen planus with thalidomide. PMID- 11115154 TI - What is the evidence for a sunscreen and melanoma controversy? PMID- 11115155 TI - Clinical features and contribution of virological findings to the management of Kaposi sarcoma in organ-allograft recipients. AB - OBJECTIVES: To describe the clinical features of Kaposi sarcoma (KS) in organ allograft recipients and to determine the contribution of human herpesvirus 8 (HHV-8) investigations to the management of KS. DESIGN, SETTING, AND PATIENTS: We examined 20 organ-allograft recipients with KS at Pitie-Salpetriere Hospital, Paris, France, between November 1, 1991, and May 31, 1999. METHODS: We detected HHV-8 antibodies using an indirect immunofluorescence assay and the HHV-8 DNA genome using nonnested polymerase chain reaction with KS-associated herpesvirus 330(233) primers in peripheral blood mononuclear cells collected at transplantation and KS diagnosis. We detected the HHV-8 genome in involved and uninvolved tissue specimens and in 10 patients' serum samples collected 1 month before the first manifestation of KS. We determined the HHV-8 double-strand DNA sequence and subtypes of open reading frame 26. INTERVENTION: Management of KS consisted of progressively tapering immunosuppressive therapy regardless of KS dissemination. Associated infections were treated when possible. Chemotherapy was prescribed only when a functional disability persisted, and polychemotherapy was prescribed for life-threatening disease. MAIN OUTCOME MEASURES: Percentage of recipients with KS remission and stabilization, organ-graft survival, and death rates. RESULTS: Remission of KS was obtained in 9 (45%) of the 20 patients independently of disease dissemination, with a mean follow-up of 35 months. The kidney graft survived in 12 (67%) of the 18 patients. Only 1 patient (5%) died of KS progression. All allograft recipients had anti-HHV-8 antibodies before transplantation. We detected HHV-8 DNA in all involved tissue samples but not in serum samples 1 month before KS onset. The most prevalent subtype was HHV-8 C (9 [53%] of 17 patients) and was not associated with extradermatological extension of KS compared with subtypes A and B'. CONCLUSIONS: Virological investigations of HHV-8 contribute poorly to KS management. Prospective studies are needed to determine the role of HHV-8 virological investigations and to identify associated cofactors so as to prevent KS in organ-allograft recipients. PMID- 11115156 TI - Topical treatment of cutaneous lesions of acquired immunodeficiency syndrome related Kaposi sarcoma using alitretinoin gel: results of phase 1 and 2 trials. AB - OBJECTIVE: To evaluate the efficacy and safety of topical alitretinoin gel (9-cis retinoic acid [LGD1057], Panretin gel; Ligand Pharmaceuticals, Inc, San Diego, Calif) in cutaneous Kaposi sarcoma (KS). DESIGN: Open-label, within-patient, controlled, dose-escalating phase 1 and 2 clinical trials. In all patients, 1 or more cutaneous KS lesions were treated with alitretinoin gel, and at least 2 other lesions served as untreated controls for up to 16 weeks. Alitretinoin (0.05% or 0.1% gel) was applied twice daily for the first 2 weeks and up to 4 times daily thereafter, if tolerated. SETTING: Nine academic clinical centers. PATIENTS: One hundred fifteen patients with biopsy-proven acquired immunodeficiency syndrome (AIDS)-related KS. MAIN OUTCOME MEASURES: AIDS Clinical Trials Group response criteria. RESULTS: Statistically significant clinical responses were observed in 31 (27%) of 115 patients for the group of treated index lesions compared with 13 (11%) for the group of untreated control lesions (P<.001). Responses occurred with low CD4(+) lymphocyte counts (<200 cells/microL) and in some patients with refractory response to previous systemic anti-KS therapy. The incidence of disease progression was significantly lower for treated index lesions compared with untreated control lesions (39/115 [34%] vs 53/115 [46%]; P =.02). Alitretinoin gel generally was well tolerated, with 90% of treatment-related adverse events confined to the application site and only mild or moderate in severity. CONCLUSIONS: Alitretinoin gel has significant antitumor activity as a topical treatment for AIDS-related KS lesions, substantially reduces the incidence of disease progression in treated lesions, and is generally well tolerated. PMID- 11115157 TI - Skin toxic effects of polyethylene glycol-coated liposomal doxorubicin. AB - OBJECTIVES: To record the profile of toxic effects of polyethylene glycol-coated liposomal doxorubicin hydrochloride (Doxil) to the skin, and to evaluate whether the long circulation pattern and enhanced accumulation of liposomes in specific skin sites will result in any unique presentations. DESIGN: Patients were accrued in the frame of dose-range-finding studies that examine the toxic effects and antitumor activity of Doxil therapy in metastatic breast and prostate cancers. All patients receiving Doxil were instructed to report any skin eruption or discomfort. Skin examination was performed on a regular basis at every cycle of Doxil therapy and after specific complaints. SETTING: Outpatient day care unit of the oncology institute of a secondary-referral medical center. PATIENTS: Sixty patients (45 women and 15 men). MAIN OUTCOME MEASURES: A basic severity scale of I through IV was adopted for toxic effects to the skin, based on National Cancer Institute common toxicity criteria. RESULTS: The following 4 patterns of skin eruptions were encountered: hand-foot syndrome (n = 24), diffuse follicular rash (n = 6), intertrigolike eruption (n = 5), and new formation of melanotic macules (n = 3). Another major toxic effect of Doxil was stomatitis, which was found to be the dose-limiting factor for the maximal single dose. Alopecia and extravasation injuries did not occur. CONCLUSIONS: The profile of toxic effects of Doxil to the skin reflects its unique pharmacokinetics and tissue distribution. These skin reactions vary significantly from those associated with doxorubicin in non-liposome-encapsulated form. PMID- 11115158 TI - T-Cell clonality in pityriasis lichenoides et varioliformis acuta: a heteroduplex analysis of 20 cases. AB - BACKGROUND: Cutaneous lesions of pityriasis lichenoides et varioliformis acuta (PLEVA), a T cell-mediated cutaneous inflammatory condition, are clinically similar to lymphomatoid papulosis (LyP), leading some authors to hypothesize that they are part of the same spectrum of lymphoproliferative disorders, although reports of the development of cutaneous lymphoma in patients with PLEVA are not as frequent as they are for patients with LyP. Furthermore, unlike in cases of LyP, no systematic search for a dominant T-cell clone has been carried out in cases of PLEVA, whereas clones have been detected in a few cases of PLEVA using mainly Southern blot analysis. OBJECTIVE: To investigate T-cell clonality in a series of archival PLEVA lesions. TISSUES: Archival paraffin-embedded biopsy specimens from 20 clinically and pathologically typical cases of PLEVA were selected. MAIN OUTCOME MEASURE: Identification of a dominant T-cell clone by polymerase chain reaction and heteroduplex analysis targeted on the TCRgamma gene. Peripheral blood mononuclear cells (PBMCs) and Jurkat cells were used as negative and positive controls. Serial dilutions of Jurkat T-cell lymphoma DNA in PBMC DNA were used to assess the sensitivity of the method. RESULTS: Analysis of 13 (65%) of 20 PLEVA biopsy specimens revealed the presence of a dominant T-cell clone. Positive and negative controls confirmed the specificity of the procedure. The sensitivity was determined to be between 1% and 5% of the total T-cell infiltrate. CONCLUSIONS: This study provides further evidence for the presence of a dominant T-cell clone in skin lesions of some patients with PLEVA and supports the hypothesis that PLEVA is part of the spectrum of clonal-T-cell cutaneous lymphoproliferative disorders. PMID- 11115159 TI - Tumor necrosis factor receptor-associated periodic syndrome: a novel syndrome with cutaneous manifestations. AB - BACKGROUND: Tumor necrosis factor receptor-associated periodic syndrome (TRAPS) is an inflammatory disorder characterized by prolonged episodes of periodic fever and localized inflammation and dominantly inherited mutations in TNFRSF1A, the gene encoding the 55-kDa tumor necrosis factor receptor. To our knowledge, the cutaneous pathologic characteristics of TRAPS have not been described previously. OBJECTIVES: To characterize the dermatologic manifestations of TRAPS by clinical, microscopic, and molecular methods, and to investigate its immunophenotype. DESIGN, SETTING, AND PATIENTS: At the National Institutes of Health Clinical Center, Bethesda, Md, a tertiary care referral center, 25 patients with a clinical and molecular diagnosis of TRAPS were evaluated clinically and 10 biopsy specimens of lesional skin were examined by light microscopy and immunohistochemistry. Patients were screened for mutations in TNFRSF1A, the gene coding for the p55 tumor necrosis factor receptor. MAIN OUTCOME MEASURES: Clinical, light microscopic, and immunohistochemical features. RESULTS: The skin eruption usually lasted 4 to 21 days (mean, 13 days). Of 25 patients, 21 (84%) presented with migratory erythematous macules and patches and 10 (40%) had edematous dermal plaques. Conjunctivitis, characterized by pain and redness and/or periorbital edema, was present in 11 patients (44%). Most patients had their first skin eruption during the first 2 years of life. All patients had fever associated with the skin eruption. Most patients had associated abdominal pain (22 [88%]) and myalgia (20 [80%]). Other symptoms included arthralgia (13 [52%]), pleuritic chest pain (10 [40%]), and headache (17 [68%]). Microscopic examination of 10 biopsy specimens of lesional skin showed a superficial and deep perivascular and interstitial infiltrate of lymphocytes and monocytes. None of the biopsy specimens showed multinucleated macrophages or granulomatous or leukocytoclastic vasculitis. The results of immunohistochemistry showed a perivascular infiltrate of CD3+, CD4+, CD8+, CD68+, CD79a-, and CD20- cells. All the mutations were missense mutations in exons 2 through 4 of TNFRSF1A, directly affecting the extracellular domain of the protein. CONCLUSIONS: TRAPS is characterized by a spectrum of dermatologic findings, including migratory patches, edematous plaques, periorbital edema, and/or conjunctivitis. TRAPS is characterized by a perivascular dermal infiltrate of lymphocytes and monocytes. PMID- 11115160 TI - Detection of type 1 cytokines in discoid lupus erythematosus. AB - BACKGROUND: Although multiple studies suggest a dysregulated T-cell cytokine production in systemic lupus erythematosus, the cytokine profile in discoid lupus erythematosus (DLE) lesions is unknown. OBJECTIVES: To characterize the cytokine profile in DLE by immunohistochemical and molecular methods, and to investigate the role of cytokines in the pathogenesis of DLE. DESIGN: Patients were evaluated clinically, and biopsy specimens of lesional skin were examined by light microscopy. Reverse transcriptase-polymerase chain reaction and immunohistochemical analysis were performed on 11 biopsy specimens. We investigated the presence of interleukin (IL) 2, interferon gamma (IFN-gamma), IL 4, tumor necrosis factor alpha, (TNF-alpha), and IL-1beta messenger RNA (mRNA) in 8 biopsy specimens of DLE and compared it with 3 biopsy specimens of normal skin. SETTING: Academic referral research hospital. PATIENTS: Eight consecutive patients with a clinical and histologic diagnosis of DLE. RESULTS: Localized DLE was found in 7 patients and widespread in 1. During the 4 years of the investigation, none of the patients developed systemic lupus erythematosus. We found significantly elevated levels of IL-2 and IFN-gamma mRNA in all 8 biopsy specimens of DLE; in contrast, no transcripts of IL-2 or IFN-gamma were detected in 3 biopsy specimens of normal skin (P<.01). Similarly, elevated levels of TNF alpha mRNA were detected in 8 DLE biopsy specimens, while no TNF-alpha mRNA was detected in 3 biopsy specimens of normal skin (P<.01). No IL-4 or IL-1 beta mRNA was detected in 8 biopsy specimens of DLE lesional skin and 3 biopsy specimens of normal patient skin. Immunohistochemical analysis showed increased staining for IL-2 and IFN-gamma receptors, while no detectable IL-4 receptor was found. No cytokine mRNA or cytokine receptor protein was detected in biopsy specimens of normal skin. CONCLUSIONS: These findings suggest that DLE is associated with type 1 cytokines characterized by the expression of IL-2 and IFN-gamma. Type 1 cytokines may be critical for induction, development, and maintenance of DLE. PMID- 11115161 TI - Granulomatous cheilitis and Borrelia burgdorferi: polymerase chain reaction and serologic studies in a retrospective case series of 12 patients. AB - BACKGROUND: Granulomatous cheilitis (GC) is a chronic granulomatous inflammation of the lips of unknown etiology, which may be associated with peripheral facial nerve paralysis and/or lingua plicata (Melkersson-Rosenthal syndrome [MRS]). Borrelia burgdorferi is a spirochete that causes Lyme borreliosis, a multisystemic infectious disease with frequent occurrence of facial nerve paralysis. An etiologic role of B burgdorferi in various granulomatous diseases has been suggested. The present study was performed to examine a possible causative role of B burgdorferi for GC/MRS by B burgdorferi-specific polymerase chain reaction analysis of biopsy specimens from affected lip tissue and determination of B burgdorferi IgG and IgM serum antibodies using enzyme-linked immunosorbent assay and immunoblot tests. OBSERVATIONS: We examined a retrospective case series of 12 patients with GC/MRS from a Lyme borreliosis endemic area (median duration of disease, 8 months [range, 3-348 months]). Borrelia burgdorferi-specific DNA could not be amplified by polymerase chain reaction in any of the 12 patients. One (13%) of 8 patients tested had a serum B burgdorferi IgG response on enzyme-linked immunosorbent assay, and 2 patients (25%) had an IgM response, but immunoblot testing yielded negative results in all 8 patients. CONCLUSION: The results of the present study do not indicate that B burgdorferi has an etiologic role in GC/MRS. PMID- 11115162 TI - Lupus erythematosus associated with genetically determined deficiency of the second component of the complement. AB - BACKGROUND: The gene deletion responsible for the type I human complement C2 deficiency was reported in 1992. The purpose of our study is to evaluate clinical and immunological characteristics of 11 patients with lupus erythematosus and type I C2 deficiency. OBSERVATIONS: We observed 5 patients with a homozygous C2 deficiency and 6 with a heterozygous C2 deficiency. Eight patients had systemic lupus erythematosus, 2 had subacute cutaneous lupus erythematosus, and 1 had chronic lupus erythematosus. Photosensitivity was present in 73% of the patients, and 64% tested positive for anti-Ro (SSA) antibodies. Renal involvement that required immunosuppressive therapy was present in 54% of the patients. Ninety percent of the patients tested positive for antinuclear antibodies, and 54% tested positive for anti-double-stranded DNA antibodies. Phenotyping of the fourth component of the complement was performed in 82% of the patients and showed a C4A4B2 phenotype, which is suggestive for the type I C2 deficiency. CONCLUSIONS: Most patients with lupus erythematosus associated with C2 type I deficiency are photosensitive, and this is probably related to the presence of anti-Ro (SSA) autoantibodies. The prognosis for those patients is not better than that for patients with lupus erythematosus in general. PMID- 11115163 TI - Allergic contact dermatitis caused by skin painting (pseudotattooing) with black henna, a mixture of henna and p-phenylenediamine and its derivatives. AB - BACKGROUND: Skin painting (pseudotattooing) with henna is traditionally performed mainly in Muslim or Hindu persons. Recently, transient artists have begun using black henna mixtures to temporarily paint the skin. Emergence of allergic contact dermatitis after application indicates the presence of a skin sensitizer in such preparations and poses future risks. OBSERVATIONS: Four patients developed allergic contact dermatitis after skin painting with black henna performed in France, Egypt, and the United States. The delay of symptoms suggested previous sensitization in 1 patient and active sensitization in 3 patients. Of 3 patients who underwent patch testing, the results were positive for p-phenylenediamine in 3 patients and for p-toluylenediamine in 1 patient. These sensitizers are found in hair dye preparations. CONCLUSIONS: The mixtures used by the artists possibly contained natural henna, a rare and weak skin sensitizer, and likely contained chemical coloring agents, diaminobenzenes, such as p-phenylenediamine and/or diaminotoluenes. The long duration of skin contact, the high concentrations of sensitizing materials, and the lack of a neutralizing agent dramatically increase the risk of skin sensitization, which is why such substances are prohibited for direct skin application. Because of the worldwide vogue of skin painting, future cases of sensitization to p-phenylenediamine and diaminobenzenes or diaminotoluenes are expected. PMID- 11115164 TI - Antibodies to molluscum contagiosum virus in the general population and susceptible patients. AB - BACKGROUND: Since many attempts to cultivate molluscum contagiosum virus (MCV) in vitro have been unsuccessful, it is difficult to prepare a large quantity of antigens. To assess the seroprevalence of antibodies against MCV in 508 subjects with or without clinical MCV infection, a truncated recombinant protein from open reading frame MC133L was synthesized using Sendai virus expression system and applied to enzyme-linked immunosorbent assay as an antigen. OBSERVATIONS: Antibodies to MCV were present in 7 (58%) of 12 patients with molluscum contagiosum, 7 (6%) of 108 healthy controls, 7 (9%) of 76 with atopic dermatitis, and 7 (18%) of 39 patients with systemic lupus erythematosus, although no clinical MCV infection was observed in the latter 3 groups. Of 7 human immunodeficiency virus (HIV)-positive patients with molluscum contagiosum, 1 (14%) was antibody positive, compared with 5 (2%) of 266 HIV-positive patients without molluscum contagiosum. Serum samples from patients with atopic dermatitis and systemic lupus erythematosus showed a higher reactivity (P<.001) than those from healthy controls, while serum samples from HIV-positive subjects showed a lower reactivity (P<. 001). CONCLUSION: The humoral immune response to MCV is usually confined to patients with molluscum contagiosum and may be affected by the immunological condition of the host. PMID- 11115165 TI - Risk of developing a subsequent nonmelanoma skin cancer in patients with a history of nonmelanoma skin cancer: a critical review of the literature and meta analysis. AB - OBJECTIVE: To assess the risk of developing a basal cell carcinoma (BCC), and/or a squamous cell carcinoma (SCC), and/or Bowen disease (SCC in situ) after a nonmelanoma skin cancer (NMSC) of a specific type. DATA SOURCES: Four electronic databases were searched from January 1, 1966, to October 21, 1999. STUDY SELECTION: We included all studies published in English, identified by standard search strategies, that provided original data quantifying the risk of an NMSC among persons with a previous NMSC. DATA EXTRACTION: For each study and separate histological type of index skin tumor and subsequent skin tumor (SCC, BCC, NMSC, or Bowen disease), we determined the 3-year cumulative risk and the incidence rate of second tumors per 100 000 person-years. In cases where more than 1 study was assessing the risk of one specific tumor type after another, we undertook a formal meta-analysis. We compared the incidence of a subsequent SCC after an index SCC and of a subsequent BCC after an index BCC with the incidence of the first occurrence of such tumors in the comparable general population. DATA SYNTHESIS: We identified and reviewed 17 studies that included data for 26 tumor combinations. Overall, the 3-year cumulative risk of a subsequent SCC after an index SCC is 18%, at least a 10-fold increase in incidence compared with the incidence of first tumors in a comparable general population. For BCCs, the 3 year cumulative risk is 44%, also at least a 10-fold increase in incidence compared with the rate in a comparable general population. The risk of developing a BCC in patients with a prior SCC is about equal to that risk among persons with a prior BCC, but the risk of developing an SCC in patients with a prior BCC is low (6%). CONCLUSIONS: Although these studies vary in their study type, location, and biases, their results are consistent. The risk of developing a subsequent skin cancer of a specific type depends on the type of prior NMSC and number of prior skin tumors of that type. Based on these findings, follow-up strategies for patients with BCC and SCC are suggested. PMID- 11115166 TI - Risk factors for delayed healing of neuropathic diabetic foot ulcers: a pooled analysis. AB - OBJECTIVE: To estimate the effect of various risk factors on the probability that neuropathic diabetic foot ulcers will heal within 20 weeks of care. DESIGN AND SETTING: A pooled or meta-analysis of individual patient data from the standard care arms of 5 randomized clinical trials was conducted. We analyzed 586 subjects with diabetes mellitus who had a neuropathic diabetic foot ulcer. All patients received good wound care, debridement, and "off-loading" of the wound. MAIN OUTCOME MEASURE: Multivariable logistic regression was used to calculate the magnitude of the association of each risk factor with patients having healed wounds. RESULTS: Logistic regression odds ratios (ORs; 95% confidence intervals [95% CIs]) revealed that those patients with a diabetic neuropathic foot ulcer that healed within 20 weeks using standard care were more likely to have a smaller wound (OR = 0.67; 95% CI, 0.55-0.81), a wound that existed for a shorter period (OR = 0.73; 95% CI, 0.61-0.87), and be nonwhite (OR = 0.64; 95% CI, 0.43 0.96) compared with patients whose wounds did not heal within 20 weeks. The patient's age (OR = 0.99; 95% CI, 0.89-1.01), serum level of glycosylated hemoglobin at the start of the study (OR = 1.03; 95% CI, 0.97-1.10), and sex (OR = 1. 02; 95% CI, 0.69-1.50) were unassociated with the probability of wound healing. Substantial heterogeneity was not found among the studies. CONCLUSIONS: A standard care regimen for diabetic neuropathic foot ulcers is most likely to be effective for patients who have wounds that are small and of brief duration. This information should help dermatologists decide initially whether to use standard care, to try a new treatment, or to refer the patient to a specialty center. PMID- 11115167 TI - Combination regimens of topical calcipotriene in chronic plaque psoriasis: systematic review of efficacy and tolerability. AB - OBJECTIVE: To examine the efficacy and tolerability of calcipotriene combined with phototherapy or systemic therapies compared with monotherapy for the treatment of chronic plaque psoriasis. DESIGN: Quantitative systematic review of 11 randomized controlled trials involving a total of 756 patients with plaque psoriasis. MAIN OUTCOME MEASURES: Rate ratios (RRs) for marked improvement or clearance in patient and investigator overall assessments of response; mean difference in percentage change in Psoriasis Area and Severity Index; and RRs for clearance in patient and investigator overall assessments of response. Adverse effects were estimated with the RR and the rate difference in terms of withdrawal rate, proportion of patients experiencing adverse events, and proportion of patients with cutaneous and noncutaneous adverse effects. RESULTS: Antipsoriatic effects of acitretin, cyclosporine, and psoralen-UV-A phototherapy were enhanced with the addition of topical calcipotriene using the Psoriasis Area and Severity Index as the outcome, but this is not translated into an increase in the number of patients who achieve at least marked improvement. At the end of treatment, the RRs for marked improvement or clearance in patient assessments were as follows: calcipotriene plus acitretin vs acitretin alone (12 weeks), 1.4 (95% confidence interval [CI], 1.0-1. 9); calcipotriene plus cyclosporine vs cyclosporine alone (6 weeks), 1.2 (95% CI, 0.9-1.6); and calcipotriene plus psoralen-UV-A vs psoralen-UV-A alone (12 weeks), 1.2 (95% CI, 0.9-1.6). Patients were also no more likely to obtain marked improvement or better with calcipotriene plus UV-B therapy than with UV-B therapy alone (RR, 1. 0; 95% CI, 0.8-1.1 at 8 weeks in the patient assessment). There is limited evidence that use of calcipotriene might reduce the cumulative exposure to phototherapy and systemic treatment. During the short duration of these trials, there were no significant differences in withdrawal rates or adverse effects between the combined regimens and their corresponding monotherapy control interventions. CONCLUSIONS: Overall, there is insufficient evidence to support any large effects in favor of combination treatment. In the patient assessments, the results do not show an adjuvant effect, but there is some evidence that use of calcipotriene might reduce cumulative exposure to systemic therapy to obtain clearance. There were no long term morbidity data on the effectiveness of any of the combinations studied. Given that psoriasis is a chronic recurrent disease for most patients, longer trials are needed to determine whether the addition of topical calcipotriene to systemic therapy improves the risk-benefit ratio by reducing the long-term risk of toxic effects. Equally important is the need to examine the impact of such combinations on the duration of remission after treatment. PMID- 11115168 TI - Confounding and interaction. PMID- 11115169 TI - What is the role of topical calcipotriol in the treatment of mild to moderate chronic plaque psoriasis? PMID- 11115170 TI - Sunscreens, nevi, and melanoma revisited. PMID- 11115171 TI - Is the self-counting of moles a valid method of assessing melanoma risk? PMID- 11115172 TI - "Evidence-Based Dermatology" section in the archives of dermatology PMID- 11115173 TI - AIDS-related Kaposi sarcoma: the role of local therapy for a systemic disease. PMID- 11115174 TI - Granulomatous cheilitis, Melkersson-Rosenthal syndrome, and orofacial granulomatosis. PMID- 11115175 TI - Off-center fold: red nodules on the arm of a patient with breast cancer. PMID- 11115176 TI - Off-center fold: irregular, pigmented genital macules. PMID- 11115177 TI - Off-center fold: serpiginous plaques on the leg. PMID- 11115178 TI - Off-center fold: dark-red plaques on the scalp of a newborn. PMID- 11115179 TI - High-dose botulinum toxin type A for axillary hyperhidrosis. PMID- 11115181 TI - Cell phone chondrodermatitis. PMID- 11115180 TI - Acres of skin: the underlying problem. PMID- 11115182 TI - Disseminated porokeratosis associated with chronic renal failure: A new type of disseminated porokeratosis? PMID- 11115183 TI - Diagnostic patterns and delays in pemphigus vulgaris: experience with 99 patients. PMID- 11115184 TI - Isomorphic phenomenon: adverse effect after epilation with the long-pulsed ruby laser. PMID- 11115185 TI - Targetoid hemosiderotic hemangioma occurring in a father and son. PMID- 11115190 TI - News and notes: december 2000 PMID- 11115192 TI - Dear readers PMID- 11115191 TI - Dear readers PMID- 11115193 TI - Special selection: frosted branch angiitis with ocular toxoplasmosis PMID- 11115194 TI - Alcohol consumption and compliance among inner-city minority patients with type 2 diabetes mellitus. AB - OBJECTIVE: To examine the relation between alcohol consumption and self-reported compliance with prescribed therapies for type 2 diabetes mellitus among underserved minority patients. DESIGN: Cross-sectional sampling of consecutive patients with diabetes was performed following routine visits to their primary care physicians. Interviewers measured compliance using the Summary of Diabetes Self-Care Questionnaire and alcohol use using the timeline followback method and the Alcohol Use Disorders Identification Test. SETTING: Seven inner-city medical clinics that provide primary care services to low-income residents of South Central Los Angeles, Calif. PARTICIPANTS: A total of 392 ethnic minority patients (61% Hispanic, 29% African American) with type 2 diabetes mellitus. MAIN OUTCOME MEASURES: Self-report compliance with prescribed diet, exercise, home glucose monitoring, medications, and outpatient follow-up. RESULTS: Drinking any alcohol containing beverage within 30 days was associated with poorer adherence to prescribed dietary recommendations for the consumption of fiber (t = 2.4; P<.05), fat (t = 4.2; P<.01), sweets (t = 2.7; P<.01), and energy (calories) (t = 2.0; P<.05). Drinkers were also less likely to exercise for at least 20 minutes per day (t = 2.2; P<.05), comply with oral medication regimens (t = 4.6; P<.01), or attend outpatient follow-up visits (r = -0.11; P<.05). Alcohol use did not significantly alter compliance with home glucose monitoring, insulin use, or hemoglobin A(1c) levels, although there was a trend toward higher hemoglobin A(1c) levels among drinkers (11.0 vs 10.4). Multivariate analysis of the data demonstrates that when demographic characteristics, health care utilization, and other diabetes-related variables are held constant, the relation between alcohol use and dietary compliance remained significant. CONCLUSION: Alcohol consumption may be associated with poorer compliance with recommendations for some self-care behaviors among inner-city minority patients with diabetes. Arch Fam Med. 2000;9:964-970 PMID- 11115195 TI - Using geographic information systems to understand health care access. AB - BACKGROUND: Determining a community's health care access needs and testing interventions to improve access are difficult. This challenge is compounded by the task of translating the relevant data into a format that is clear and persuasive to policymakers and funding agencies. Geographic information systems can analyze and transform complex data from various sources into maps that illustrate problems effortlessly for experts and nonexperts. OBJECTIVE: To combine the patient data of a community health center (CHC) with health care survey data to display the CHC service area, the community's health care access needs, and relationships among access, poverty, and political boundaries. DESIGN: Georeferencing, analyzing, and mapping information from 2 databases. SETTING: Boone County, Missouri. PARTICIPANTS: Community health center patients and survey respondents. MAIN OUTCOME MEASURES: Maps that define the CHC service area and patient demographics and show poor health care access in relation to the CHC service area, CHC utilization in relation to poverty, and rates of health care access by geopolitical region. RESULTS: The CHC serves a distinctly different area than originally targeted. Subpopulations with unmet health care access needs and poverty were identified by census tract. These underserved populations fell within geopolitical boundaries that were easily linked to their elected officials. CONCLUSIONS: Geographic information systems are powerful tools for combining disparate data in a visual format to illustrate complex relationships that affect health care access. These systems can help evaluate interventions, inform health services research, and guide health care policy. Arch Fam Med. 2000;9:971-978 PMID- 11115197 TI - Costs of illness due to Bordetella pertussis in families. AB - OBJECTIVE: To assess costs of pertussis morbidity among families in a community setting. DESIGN: Prospective survey. RESULTS: Sixty-nine families (87 individuals) were studied. Twelve of 14 families with household contacts included an ill adolescent or parent. This individual was the first identified pertussis case within the household in 8 families. A family member required an average of 1.6 visits before (range, 0-7 visits) and after (range, 0-9 visits) pertussis was diagnosed; children younger than 1 year needed 2.5 and 2 visits, respectively. Symptomatic improvement occurred after a mean of 31 days (range, 4-134 days) after pertussis diagnosis and full recovery after a mean of 66 days (range, 5-383 days). Adults experienced the longest recovery time (median, 93 days) compared with other age groups. The average medical costs for an infant, child, adolescent, and adult were $2822, $308, $254, and $181, respectively. Parents lost an average of 6 workdays (range, 1-35 days) to care for an ill child; for these parents, costs associated with work loss averaged $767 per family. An average of 1.7 and 0.7 lost workdays accrued to bring an ill child to a physician's office and the emergency department, respectively. A majority (58%) of parents working while family members were ill with pertussis reported decreased work productivity ranging from 25% to 99%. Work-related costs contributed more than 60% of the overall costs of pertussis. CONCLUSIONS: The adverse financial effect of pertussis on 69 families in Monroe County, New York, was $145,903 ($2115 per family) and supports the need for booster immunizations in adolescents and adults. Arch Fam Med. 2000;9:989-996 PMID- 11115196 TI - Effectiveness of pseudoephedrine plus acetaminophen for treatment of symptoms attributed to the paranasal sinuses associated with the common cold. AB - BACKGROUND: Little data exist on the cause and treatment of subfacial pain and pressure and other discomfort attributed to the paranasal sinuses that develop early during the course of the common cold. The purpose of this study was to determine the efficacy of the combination of pseudoephedrine hydrochloride with acetaminophen for the treatment of early symptoms during colds, which are attributed by the patient to the sinuses. METHODS: Four hundred thirty subjects (216, pseudoephedrine and acetaminophen recipients; 214, placebo recipients) with cold symptoms of 48 hours or less who reported overall "sinus" symptoms of at least moderate severity were enrolled in this randomized double-blind placebo controlled 2-dose study. Self-reported symptoms were scored (0 to 4, absent to severe) before and at 2 hours after the first and second doses. The 2 primary were measured 2 hours after the second dose were the overall sinus symptom assessment and a weighted composite assessment of sinus pressure, pain, and congestion (sinus symptoms). RESULTS: Compared with baseline, 2 hours after the second dose, the mean +/- SEM overall sinus symptom assessment score had decreased by 1.30 +/- 0. 06 in the pseudoephedrine and acetaminophen-treated subjects compared with 0.93 +/- 0.06 in the placebo-treated subjects (P< or = .029). The mean +/- SEM weighted average of sinus symptoms 2 hours after the second dose of study medication had decreased by 1.14 +/- 0.06 in the pseudoephedrine and acetaminophen-treated subjects compared with 0.84 +/- 0.06 in the placebo-treated subjects (P< or = .029). Reductions in similar magnitude were also observed for each of the individual sinus symptoms, and headache and rhinorrhea. Nervousness occurred in 4% of the pseudoephedrine and acetaminophen recipients compared with 0% of placebo recipients (P =.007). CONCLUSION: Our results suggest that pseudoephedrine plus acetaminophen is effective for relief of symptoms attributable to the paranasal sinuses that may develop early in the course of a cold. Arch Fam Med. 2000;9:979-985 PMID- 11115199 TI - Prescription medication costs: a study of physician familiarity. AB - BACKGROUND: Studies in the past 25 years have suggested that physicians are not familiar with the costs of common prescription medications. OBJECTIVES: To determine physician familiarity with the cost of common prescription medications and to determine the value physicians place on knowing information regarding the cost of medications. DESIGN: Survey. SETTING: Seven community-based family medicine residency teaching clinics in Iowa. PARTICIPANTS: Two hundred five practicing resident and faculty physicians. INTERVENTIONS: From a series of $10 price intervals (range, $0.01-$80.00), physicians were asked to select the interval containing the cash price of the medication to an uninsured patient for 50 medications commonly prescribed in outpatient family medicine clinics. Physicians were also questioned about the value of medication cost information to their practice. MAIN OUTCOME MEASURES: The percentage of correct responses and the mean pricing scores were calculated for each respondent and for all medications. RESULTS: One hundred seventy-eight physicians responded (86.8%). Only 22.9% of the responses correctly identified the cost of the medication. More than two thirds (68.3%) of the responses underestimated the correct price interval. Branded drugs were underestimated in 89.9% of responses, while generic drugs were overestimated in 90.2% of responses. Overall, 64.4% of physicians believed they did not receive sufficient information in their practices regarding prescription drug costs, and nearly all (93.6%) reported that regular information on prescription medication costs would help them prescribe more cost-effectively. CONCLUSIONS: Physicians are unfamiliar with the costs of medications they commonly prescribe, and they report that regular access to information on prescription medication costs would help them prescribe more cost-effectively. Arch Fam Med. 2000;9:1002-1007 PMID- 11115198 TI - Does drug treatment of patients with acute bronchitis reduce additional care seeking? Evidence from the Practice Partner Research Network. AB - BACKGROUND: Considerable discussion has focused on treatment methods for patients with acute bronchitis. OBJECTIVE: To examine whether antibiotic or bronchodilator treatment is associated with differences in follow-up visit rates for patients with acute bronchitis. METHODS: A retrospective medical chart review was conducted for patients with a new episode of acute bronchitis over a 3-year period in the Practice Partner Research Network (29,248 episodes in 24,753 patients). Primary outcomes of interest were another visit in the next 14 days (early follow-up) or 15 to 28 days after initial treatment (late follow-up). RESULTS: Antibiotics were used more commonly in younger patients (<18 years), whereas older patients (>65 years) were more likely to receive no treatment. Younger patients treated with antibiotics were less likely to return for an early follow-up visit, but no differences were seen in adults and older patients. Late follow-up rates were not affected by the initial treatment strategy. When patients did return for a follow-up visit, no new medication was prescribed to most (66% of younger patients and 78% of older adults). However, compared with patients who did not receive an antibiotic at their first visit, patients initially treated with an antibiotic were about 50% more likely to receive a new antibiotic at their second visit. CONCLUSIONS: Initial prescribing of an antibiotic reduces early follow-up for acute bronchitis in younger patients but seems to have no effect in adults. However, reductions in future follow-up visits might be outweighed by increases in antibiotic consumption because patients who return for a follow-up visit seem to receive additional antibiotic prescriptions. Arch Fam Med. 2000;9:997-1001 PMID- 11115200 TI - Prevalence and nature of orofacial and dental problems in family medicine. AB - OBJECTIVE: To determine the prevalence and nature of orofacial and dental problems in 2 family medicine practices. DESIGN: Prospective, cross-sectional analysis of consecutive patient visits. SETTING: Urban and rural family medicine practices. PATIENTS AND PARTICIPANTS: Four hundred seventy-two patients between age 10 and 86 years. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Prevalence and nature of patient visits to family medicine practices that were either initiated by problems in the region of the oral cavity or that involved questions raised by the patient concerning oral or perioral sites. RESULTS: Twenty-one patients (4.5%) of 472 met the inclusion criteria, 16 (76%) of whom had an oral problem as the primary or secondary reason for their visit. Perioral pain and mucosal ulcerations were the most common problems, and gingival tissue was the most common location. Almost two thirds of these patients had bacterial, fungal, or viral infections. Regarding treatment, 13 (62%) of these patients received advice, 10 (48%) received prescriptions, and 3 (15%) were referred to a dentist or another medical specialist. CONCLUSIONS: Oral and perioral problems are common in the practice of family medicine, which suggests the desirability for specific oral medicine topics in the training and continuing education of primary care physicians. Arch Fam Med. 2000;9:1009-1012 PMID- 11115201 TI - A survey of primary care physician practice patterns and adherence to acute low back problem guidelines. AB - OBJECTIVE: This study evaluated physicians' self-reported management of acute low back problems in adults and adherence with published guidelines. DESIGN: Self administered written survey based on the US Agency for Health Care Policy and Research (now the Agency for Healthcare Research and Quality) guideline on acute low back problems in adults. SETTING: A region of northern Illinois with a population around 250 000 and encompassing a medium-sized city. PARTICIPANTS: One hundred eighty-two primary care physicians (nonpediatric) with medical staff appointments at area hospitals. MAIN OUTCOME MEASURE: Adherence to published recommendations. RESULTS: Eighty-seven surveys were received for a 48% response rate. Overall, survey respondents recognized 5 of 7 red flags representing serious underlying abnormality 50% or less of the time. Forty percent (35/87) of physicians provided patients with written educational material, and only 25%(22/87) indicated they evaluated motor function of the fifth lumbar nerve, the most commonly affected level in intervertebral disk disease disease. About 25% (24/87) reported routine use of plain films; and 16% (14/87), routine use of computed tomography or magnetic resonance imaging. Most oral medication use was consistent with recommendations, but many also used drugs conditionally discouraged by the guideline (muscle relaxants, 91% [79/87]; opioids, 62% [54/87]) or cautioned against (oral steroids, 45% [39/87]; antidepressants, 23% [20/87]; injection therapy, 52% [45/87]). Only 22% (19/87) of respondents used or recommended manipulation. CONCLUSIONS: The management of patients with acute low back problems by primary care physicians differs significantly from Agency for Health Care Policy and Research guideline recommendations in several key areas that include awareness of red flags, use of medication, use of radiographic studies, the need for patient education, and the use of physical modalities. Future research should focus on the impact of guideline compliance on patient outcomes and cost-effectiveness. Arch Fam Med. 2000;9:1015-1021 PMID- 11115202 TI - A survey of skin cancer screening in the primary care setting: a comparison with other cancer screenings. AB - OBJECTIVE: To determine primary care physicians' perceived importance and frequency of performance of skin cancer screening in comparison with other cancer screening examinations. DESIGN: Descriptive survey study. PARTICIPANTS: Five thousand US family physicians and internal medicine specialists randomly selected from the Official American Board of Medical Specialists Directory of Board Certified Medical Specialists. MAIN OUTCOME MEASURES: Self-reported importance and performance of cancer screening examinations. RESULTS: Eligible physicians (1363 total: 814 family physicians and 549 internists) completed the survey with a response rate of 30%. Overall, 52% of respondents rated skin cancer screening as "extremely" important, compared with 79% for digital rectal examination, 88% for clinical breast examination, and 87% for Papanicolaou testing. Thirty-seven percent of physicians reported performing complete body skin examinations on 81% to 100% of patients, compared with digital rectal examination, for which 78% of physicians reported performing the examination on 81% to 100% of patients, or the clinical breast examination, for which 82% of physicians reported performing the examination on 81% to 100% of patients. A higher percentage of physicians in practice for more than 30 years ranked skin cancer screening as extremely important and reported a higher frequency of screening examinations. Physicians in a suburban practice setting reported performing skin examinations more often than those in urban or rural settings. Overall, the self-reported frequency of skin examination was strongly correlated with the physician's importance rating of skin cancer screening. CONCLUSIONS: A majority of primary care physicians rate skin cancer screening as extremely important. The reported importance of skin cancer screening and frequency of skin cancer examination among primary care physicians is significantly less than for other cancer examinations. This likely represents a multitude of factors, including logistic constraints and lack of consensus on the efficacy of skin cancer screening. Arch Fam Med. 2000;9:1022 1027 PMID- 11115203 TI - Symptom severity and perceptions in subjects with panic attacks. AB - OBJECTIVES: To (1) identify aspects that defined the self-perceived worst panic attack, (2) determine how subjects with panic attacks perceive symptoms compared with control subjects, and (3) determine the role of symptom perceptions in seeking care for the worst panic attack. DESIGN: Cross-sectional survey. SETTING: Community-based. PATIENTS OR OTHER PARTICIPANTS: Ninety-seven subjects with panic attacks as defined by the Diagnostic and Statistical Manual of Mental Disorders, Revised Third Edition (with or without panic disorder), and 97 demographically matched controls. INTERVENTION: None. MAIN OUTCOME MEASURES: Subjects and controls completed the Symptom Perception Scales, and subjects with panic attacks completed the Acute Panic Inventory and a questionnaire concerning care-seeking behavior for their self-perceived worst attack. RESULTS: Compared with controls, subjects with panic attacks perceived many symptoms as more embarrassing but differed little in their perceptions of need for treatment, threat to life, and disruption of functioning. Particular symptoms (ie, dyspnea, fear, dizziness, and faintness) tended to differ in most perceptions. However, symptom perceptions did not play a significant role in care-seeking behavior for the worst attack. CONCLUSIONS: Subjects with panic attacks perceive symptoms as more embarrassing than controls, and have different perceptions about particular symptoms. Cognitive approaches addressing negative patient perceptions may reduce anxiety, inappropriate use of health care services, and adverse outcomes. Arch Fam Med. 2000;9:1028-1035 PMID- 11115204 TI - Profile of users of real-time interactive teleconference clinical consultations. AB - BACKGROUND: Real-time interactive teleconference clinical consultations are envisioned for increasing accessibility to medical care by patients whose demographics restrict care. There are no published studies, however, describing referrals and the referring practitioners, patients, and specialists participating in these consultations. OBJECTIVE: To assess characteristics of participants of interactive teleconference clinical consultations. DESIGN: Descriptive study, February 1, 1996, through April 30, 1999. SETTING: Eastern North Carolina: Brody School of Medicine at East Carolina University and 7 rural hospitals and clinics in its telemedicine network. SUBJECTS: Rural practitioners requesting consultations (n = 76), consulting physicians (n = 40), and patients completing evaluations following consultations (n = 495). MAIN OUTCOME MEASURES: Demographic and descriptive variables for referring providers, patients, and consulting physicians relative to the population in the region and to patients and physicians at the East Carolina University School of Medicine clinics. RESULTS: The largest number of referrals (65.2%) were made to obtain a second opinion or recommend a management plan in dermatology (33.5%), allergy (21.0%), or cardiology (17.8%). Significant patient characteristics were race (56.8% minorities), age (19.6% < or = 10 years old and 26.0% > or = 59.0 years old), sex (59% females), and insurance status (10.7% no insurance, 33.7% Medicaid, 15.4% Medicare). In addition, 38.0% had household incomes below the poverty level. Only 5.2% of the patients would have been treated by the referral practitioner, making travel necessary for consultation. Demographic characteristics of the practitioners were not statistically different. CONCLUSIONS: Participants of interactive teleconference clinical consultations are patients whose access to medical care might otherwise be limited. Use of telemedicine by practitioners is not related to age or sex. Arch Fam Med. 2000;9:1036-1040 PMID- 11115205 TI - Authors' comment PMID- 11115206 TI - Health behaviors, health status, and access to and use of health care: a population-based study of lesbian, bisexual, and heterosexual women. AB - BACKGROUND: There is a dearth of validated information about lesbian and bisexual women's health. To better understand some of these issues, we used population based data to assess variations in health behaviors, health status, and access to and use of health care based on sexual orientation. METHODS: Our study population was drawn from a population-based sample of women, the 1997 Los Angeles County Health Survey. Participants reported their sexual orientation and these analyses included 4697 women: 4610 heterosexual women, 51 lesbians, and 36 bisexual women. We calculated adjusted relative risks to assess the effect of sexual orientation on important health issues. RESULTS: Lesbians and bisexual women were more likely than heterosexual women to use tobacco products and to report any alcohol consumption, but only lesbians were significantly more likely than heterosexual women to drink heavily. Lesbians and bisexual women were less likely than heterosexual women to have health insurance, more likely to have been uninsured for health care during the preceding year, and more likely to have had difficulty obtaining needed medical care. During the preceding 2 years, lesbians, but not bisexual women, were less likely than heterosexual women to have had a Papanicolaou test and a clinical breast examination. CONCLUSIONS: In this first population-based study of lesbian and bisexual women's health, we found that lesbians and bisexual women were more likely than heterosexual women to have poor health behaviors and worse access to health care. These findings support our hypothesis that sexual orientation has an independent effect on health behaviors and receipt of care, and indicate the need for the increased systematic study of the relationship between sexual orientation and various aspects of health and health care. Arch Fam Med. 2000;9:1043-1051 PMID- 11115207 TI - Can depression treatment in primary care reduce disability? A stepped care approach. AB - OBJECTIVE: To assess effects of stepped collaborative care depression intervention on disability. DESIGN: Randomized controlled trial. SETTING: Four primary care clinics of a large health maintenance organization. PATIENTS: Two hundred twenty-eight patients with either 4 or more persistent major depressive symptoms or a score of 1.5 or greater on the Hopkins Symptom Checklist. Depression items were randomized to stepped care intervention or usual care 6 to 8 weeks after initiating antidepressant medication. INTERVENTION: Augmented treatment of persistently depressed patients by an on-site psychiatrist collaborating with primary care physicians. Treatment included patient education, adjustment of pharmacotherapy, and proactive monitoring of outcomes. MAIN OUTCOME MEASURES: Baseline, 1-, 3-, and 6-month assessments of the Sheehan Disability Scale and the social function and role limitation subscales of the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36). RESULTS: Patients who received the depression intervention experienced less interference in their family, work, and social activities than patients receiving usual primary care (Sheehan Disability Scale, z = 2.23; P =.025). Patients receiving intervention also reported a trend toward more improvement in SF-36-defined social functioning than patients receiving usual care (z = 1.63, P =.10), but there was no significant difference in role performance (z = 0.07, P =.94). CONCLUSIONS: Significant disability accompanied depression in this persistently depressed group. The stepped care intervention resulted in small to moderate functional improvements for these primary care patients. Arch Fam Med. 2000;9:1052-1058 PMID- 11115209 TI - Clues to early Alzheimer dementia in the outpatient setting. AB - BACKGROUND: As the elderly population booms and the prevalence of dementia soars, it becomes imperative that primary care physicians recognize early dementia within their own practices. Early recognition and diagnosis of dementia will allow appropriate intervention and treatment to improve morbidity. OBJECTIVE: To examine the most common symptoms associated with early Alzheimer disease (AD), as presented by patients and their families, and to compare these with the recommendations of the "7-Minute Screen" by Solomon et al for the identification of AD and the recommendations of the Agency for Health Care Policy and Research (AHCPR) for the early recognition of dementia. METHODS: A retrospective medical record review was conducted in an outpatient referral population within 2 geriatric evaluation centers. Patient medical record selection was based on Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition criteria for AD, a Mini-Mental State Examination (MMSE) score of 23 or higher, a Geriatric Depression Scale score of less than 5, age above 60 years, and at least an eighth grade level of education. RESULTS: From 1025 medical records reviewed, 50 patients were chosen who fulfilled all inclusion criteria. Forty patients (80%) missed at least 2, if not all 3, recall items on the MMSE. Thirty patients (60%) had difficulty managing finances and/or balancing a checkbook; 16 (32%) frequently repeated stories and statements; 15 (30%) became lost while driving; 10 (20%) frequently forgot the names of relatives; and 10 (20%) had poor judgment. These results demonstrated a high correlation with recall as a diagnostic factor in diagnosing early AD as found in the 7-Minute Screen. Moreover, these "clues" correlated well with the AHCPR's symptoms that indicate dementia. The symptoms specifically overlapped in the areas of learning and retaining new information (repetition), handling complex tasks (calculation), reasoning ability (judgment), and spatial ability and orientation (driving). CONCLUSIONS: There may be a constellation of symptoms associated with early AD. This constellation includes missing recall items on the MMSE, difficulty in calculation, repetition, getting lost while driving, forgetting the names of relatives, and having poor judgment. Recall is the symptom most consistent with the findings of the 7-Minute Screen in diagnosing AD. However, repetition, calculation, judgment, and driving highly correlate with the AHCPR's dementia symptom checklist. Therefore, if primary care physicians keep this constellation of symptoms in mind while evaluating their geriatric population, they will have greater ability to suspect, diagnose, and treat AD at an early stage. Arch Fam Med. 2000;9:1066-1070 PMID- 11115208 TI - Competing demands from physical problems: effect on initiating and completing depression care over 6 months. AB - OBJECTIVE AND DESIGN: To evaluate a cohort of patients with major depression to examine the effect of competing demands on depression care during multiple visits over 6 months. PARTICIPANTS AND SETTING: Ninety-two patients with 5 or more symptoms of depression and no recent depression treatment were evaluated by 12 primary care physicians in 6 practices in the usual-care arm of an effectiveness trial of the Agency for Health Care Policy and Research Depression Guidelines. MAIN OUTCOME MEASURE: Treatment was considered to be initiated if the patient reported starting a guideline-concordant antidepressant medication or making a visit for specialty counseling. Treatment completion was defined as either a 3 month course of guideline-concordant antidepressant use or completion of 8 or more specialty counseling visits. RESULTS: Among the 92 patients reporting no recent treatment at study enrollment, 57% reported starting and 17% reported completing a course of guideline-concordant antidepressant medication and or specialty counseling at the 6-month interview. The severity of physical problems among patients with high enthusiasm for depression treatment decreased the odds that patients would initiate depression therapy. Severity of physical problems had no observable effect on completing depression therapy in the group of patients who initiated treatment. CONCLUSIONS: Physical problems compete with depression for attention over multiple visits in untreated patients who are enthusiastic about getting care for their emotional problems. Interventions are needed for this high-risk group, because depression treatment could potentially enhance patients' treatment of their physical problems. Arch Fam Med. 2000;9:1059 1064 PMID- 11115210 TI - Risks associated with the practice of traditional Chinese medicine: an Australian study. AB - OBJECTIVE: To investigate the nature and frequency of adverse events that occur as a result of the practice of traditional Chinese medicine (acupuncture and Chinese herbal medicine) in Australia. METHODS: Data on adverse events were obtained as part of a comprehensive survey of all occupational health groups, government-registered and unregistered, who practiced traditional Chinese medicine or 1 of its main modalities. RESULTS: Practitioners reported numerous adverse events arising from the application of acupuncture (including fainting, nausea and vomiting, and increased pain), or the consumption of Chinese herbal medicines (including direct toxic effects and allergic reactions). Practitioners experienced an average of 1 adverse event every 8 to 9 months of full-time practice or 1 adverse event for every 633 consultations. The mean adverse event rate of nonmedical practitioners was less than half the mean adverse event rate of medical practitioners. CONCLUSIONS: The practices of acupuncture and Chinese herbal medicine are not risk-free and fatalities have occurred. Variation in adverse event rates between medical and nonmedical practitioners may reflect differences in relevant education or different reporting behaviors. These data represent the first step in the evaluation of adverse event rates in traditional Chinese medicine. Arch Fam Med. 2000;9:1071-1078 PMID- 11115211 TI - Self-reported health, illness, and self-care among finnish physicians: a national survey. AB - BACKGROUND: Physicians' health problems have been discussed mainly in relation to substance abuse and psychiatric disorders. In this study, the prevalence of common chronic diseases and their treatment were determined. OBJECTIVE: To find differences in self-reported health status, amount of sick leave, and the use of health services among physicians according to sex and specialty. Data were also compared with those of the total employed population. DESIGN AND SETTING: Cross sectional postal questionnaire survey in Finland. PARTICIPANTS AND METHODS: A random sample of licensed physicians younger than 66 years (n = 4477) was randomly selected from the register of the Finnish Medical Association. A total of 3313 physicians (74%) responded. MAIN OUTCOME MEASURES: Perceived health, prevalence of diseases, self-treatment of diseases, amount of sick leave, and medical consultations. RESULTS: Female physicians assessed their health as being better than other female employees and had used health services and had been on sick leave more often than their male colleagues. Male physicians assessed their health as being equal to that of other men. Both female and male physicians had fewer sick leave than other employees. However, physicians-especially men reported many common chronic illnesses as often or more often than other employees. Physicians had consulted other medical professionals less often than other employees, and they primarily self-treated their illnesses. Of the specialties, psychiatrists had used health services and had been on sick leave more often than other physicians. CONCLUSION: This study indicates that the usual form of care of physicians' diseases is self-treatment and "working through" illnesses. Arch Fam Med. 2000;9:1079-1085 PMID- 11115212 TI - Factors associated with emergency department utilization for nonurgent pediatric problems. AB - OBJECTIVE: To identify specific caretaker and utilization characteristics predictive of the use of the emergency departments (EDs) for nonurgent reasons. Each year more than 20 million children in the United States seek medical care in EDs. Between one third and one half of these visits are for nonurgent reasons. DESIGN: A descriptive study conducted during a 6-month period. SETTING: Two urban hospital EDs. MEASURE: A questionnaire was designed to elicit information about specific caretaker characteristics and their reasons for using the ED for their child's nonurgent medical care. SUBJECTS: Two hundred caretakers and children brought to the ED for nonacute medical care. Caretakers in this study included mothers (82%) with a mean age of 30 years, single caretakers (70%), and unemployed caretakers (60%). The average age of the children was 6.2 years. RESULTS: Most caretakers (92%) reported having a continuity physician for their children. Caretakers who reported being taken to the ED when they were children (P<.002) and those with Medicaid insurance (P<.001) were more likely to view the ED as the usual site of care. Being a single parent was a predictor for nonurgent visits (P<.05). CONCLUSIONS: Predicting which caretakers are at risk for using the ED for nonurgent care when their children are sick provides the primary care physician a means of identifying specific patients who may benefit from interventions designed to promote a more cost-effective approach to using medical resources. Arch Fam Med. 2000;9:1086-1092 PMID- 11115213 TI - Who gets screened during pregnancy for partner violence? AB - CONTEXT: Despite recommendations to screen prenatal care patients for partner violence, the prevalence of such screening is unknown. OBJECTIVES: To estimate the statewide prevalence of partner violence screening during prenatal care among a representative sample of North Carolina women with newborns and to compare women screened for partner violence with women not screened. DESIGN, SETTING, AND PARTICIPANTS: This investigation examines data gathered through the North Carolina Pregnancy Risk Assessment Monitoring System, a random sample of more than 2600 recently postpartum women who were delivered of newborns between July 1997 and December 1998. MAIN OUTCOME MEASURES: Self-reports of violence, health service factors, and sociodemographic characteristics. ANALYSIS: The prevalence of screening was computed, and odds ratios and 95% confidence intervals were used to examine bivariate and multivariable associations between being screened for partner violence and other factors. RESULTS: Thirty-seven percent of women reported being screened for partner violence during prenatal care. Logistic regression analysis found that women were more likely to be screened if they received prenatal care from (1) a public provider paid by a public source; (2) a private provider paid by a public source; or (3) a public provider paid by a private source. CONCLUSIONS: These findings suggest that the majority of prenatal care patients in North Carolina are not screened for partner violence. Screening appears to be most highly associated with whether a woman is a patient in the public sector or the private sector, and with the source of payment for prenatal care. Arch Fam Med. 2000;9:1093-1099 PMID- 11115214 TI - A comparison of family medicine research in research intense and less intense institutions. AB - BACKGROUND: Family medicine is a relatively new specialty that has been trying to develop a research base for 30 years. It is unclear how institutional research success and emphasis have affected the research productivity of family medicine departments. OBJECTIVE: To examine the research infrastructure, productivity, and barriers to productivity in academic family medicine in research intense and less intense institutions. DESIGN, SETTING, AND PARTICIPANTS: A survey of 124 chairs among institutional members of the Association of Departments of Family Medicine. Departments were categorized as being associated with research intense institutions (defined as the top 40 in National Institute of Health funding) or less intense institutions. MAIN OUTCOME MEASURES: Prioritization of research as a mission, number of funded research grants, total number of research articles published, and number of faculty and staff conducting research. RESULTS: The response rate was 55% (N = 68). Of 5 potential ratings on the survey, research was the fourth highest departmental priority in both categories of institutions. Departments in research intense institutions were larger, had more faculty on investigational tracks, and employed more research support staff (P<.05). Neither category of department published a large number (median = 10 in both groups) of peer-reviewed articles per year. Controlling for the number of full-time equivalent faculty, the departments in less intense institutions published a median of 0.7 articles, while the research intense institutions published 0.5 (P =.30). Departments in research intense institutions received more grant funding (P<.005) in both unadjusted and adjusted analyses. Chairs reported a scarcity of qualified applicants for research physician faculty openings. CONCLUSION: Future initiatives should focus on prioritizing research and creating a critical mass of researchers in family medicine. Arch Fam Med. 2000;9:1100-1104 PMID- 11115215 TI - Why family practice research? PMID- 11115216 TI - Selection bias from requiring patients to give consent to examine data for health services research. AB - BACKGROUND: New rulings nationwide require health services researchers to obtain patient consent before examining personally identifiable data. A selection bias may result if consenting patients differ from those who do not give consent. OBJECTIVE: To compare patients who consent, refuse, and do not answer. DESIGN: Patients completing an in-office survey were asked for permission to be surveyed at home and for their records to be reviewed. Survey responses and practice billing data were used to compare patients by consent status. SETTING: Urban family practice center. PATIENTS: Of 2046 eligible patients, 1106 were randomly selected for the survey, were approached by staff, and agreed to participate. Approximately 87% of the nonparticipants were eliminated through a randomization process. MAIN OUTCOME MEASURE: Consent status. RESULTS: A total of 33% of patients did not give consent: 25% actively refused, and 8% did not answer. Consenting patients were older, included fewer women and African Americans, and reported poorer physical function than those who did not give consent (P<.05). Patients who did not answer the question were older, included more women and African Americans, and were less educated than those who answered (P<.02). Visits for certain reasons (eg, pelvic infections) were associated with lower consent rates. On multivariate analysis, older age, male sex, and lower functional status were significant predictors of consent. CONCLUSIONS: Patients who release personal information for health services research differ in important characteristics from those who do not. In this study, older patients and those in poorer health were more likely to grant consent. Quality and health services research restricted to patients who give consent may misrepresent outcomes for the general population. Arch Fam Med. 2000;9:1111-1118 PMID- 11115217 TI - Are no-suicide contracts effective in preventing suicide in suicidal patients seen by primary care physicians? PMID- 11115218 TI - Athletes resuming activity after infectious mononucleosis. PMID- 11115219 TI - Rofecoxib, a new cyclooxygenase 2 inhibitor, shows sustained efficacy, comparable with other nonsteroidal anti-inflammatory drugs: a 6-week and a 1-year trial in patients with osteoarthritis. Osteoarthritis Studies Group. AB - INTRODUCTION: Rofecoxib, a cyclooxygenase 2 inhibitor (sometimes known as a specific cyclooxygenase 2 inhibitor or Coxib), is used in osteoarthritis (OA). Published information indicates rofecoxib's improved gastrointestinal safety profile over nonselective nonsteroidal anti-inflammatory agents (NSAIDs). OBJECTIVE: To evaluate the efficacy and safety of rofecoxib in treating OA in 2 studies. METHODS: Two randomized, double-blind, parallel-group studies in patients with OA of the knee or hip were conducted using identical entry criteria and end points. A 6-week placebo-controlled trial in 736 patients compared 12.5 and 25 mg of rofecoxib once daily with 800 mg of ibuprofen 3 times daily, and a 1 year study compared 12.5 and 25 mg of rofecoxib once daily with 50 mg of diclofenac 3 times daily in 693 patients. RESULTS: Rofecoxib, at 12.5 and 25 mg, demonstrated efficacy clinically comparable with ibuprofen, assessed by 3 primary end points according to predefined comparability criteria. Both rofecoxib doses and ibuprofen provided significantly greater efficacy than placebo on all primary end points at 6 weeks. Both rofecoxib doses and diclofenac showed similar efficacy over 1 year. All treatments were well tolerated. CONCLUSIONS: Rofecoxib is effective in treating OA with once-daily dosing for 6 weeks and 1 year. Rofecoxib was generally safe and well-tolerated in OA patients for 6 weeks and 1 year. Arch Fam Med. 2000;9:1124-1134 PMID- 11115220 TI - New antiepileptic drugs: into the new millennium. AB - There has been an explosion of new antiepileptic drug availability for physicians to treat patients with recurrent seizures. Principal antiepileptic drugs consisted of 6 key agents for both generalized and partial epilepsy for nearly 8 decades. Since 1993, the availability of newer "second-generation" agents has nearly doubled the armamentarium available for the 2.5 million patients who have recurrent seizures. This new influx of medications has flooded the medical and lay community with choices never before appreciated. The promise of improved tolerability with different safety and efficacy profiles has been exciting for all involved in epilepsy management. While most of the newer agents have been approved for adjunctive use in medically refractory partial epilepsy with recurrent complex partial and secondarily generalized seizures, efficacy is expanding to include generalized epilepsy and children for some agents. Arch Fam Med. 2000;9:1135-1141 PMID- 11115221 TI - The neurosurgical treatment of epilepsy. AB - Despite the new advancements in antiepileptic drug development, thousands of people with epilepsy will remain intractable to medication. For a considerable proportion of these people, epilepsy surgery is a consideration for better control of their seizures. Resective surgery is now standard practice for patients with medication-refractory epilepsy. Temporal lobectomy continues to be the most common surgery performed. Once patients fail 2 to 3 optimal trials of antiepileptic medication, further drug therapy offers a minimal number of patients freedom from seizures. In contrast, temporal lobectomy in carefully selected patients may result in seizure-free outcomes in more than 70% to 90% of patients with intractable seizures. As technology and drug availability increases in the new millennium, it is important for the primary care physician to be aware of epilepsy surgery as a means to treat patients with antiepileptic drug refractory epilepsy. Arch Fam Med. 2000;9:1142-1147 PMID- 11115222 TI - Visit-specific expectations and patient-centered outcomes: a literature review. AB - BACKGROUND: Primary care patients often have certain expectations when visiting physicians, many of which may be undetected. These unmet expectations can affect outcomes such as satisfaction with care. We performed a formal literature review to examine the effect of fulfillment of patients' visit-specific expectations on their satisfaction as well as on health status and compliance. PATIENTS AND METHODS: Included studies were conducted in primary care settings, systematically recruited patients, elicited previsit and/or postvisit expectations relative to specific visits, and measured patient-centered outcomes. Two reviewers abstracted information on study characteristics; types, timing, and method of expectation ascertainment; and outcomes. Disagreements were resolved by consensus. RESULTS: Twenty-three studies were reviewed including 7 trials, 4 cohort studies, and 12 cross-sectional studies. Patients frequently expected information rather than specific physician actions, but physicians often did not accurately perceive patients' visit-specific expectations. In 19 studies that assessed postvisit patient satisfaction, a positive association between meeting patient expectations and overall satisfaction was demonstrated in 11 studies, inconclusive in 3, and not established in 5. In 2 studies assessing physician satisfaction, physicians with access to patients' expectations were more satisfied than those without access. Other outcomes (symptom or disease improvement, health status, test ordering, health care costs, psychological symptoms) were measured in only a few studies, and the results were inconclusive. CONCLUSIONS: Addressing patients' visit-specific expectations appears to affect satisfaction to a modest degree. Future studies should evaluate methods that efficiently elicit, prioritize, and provide patients' previsit expectations for physicians and should examine the longitudinal effect of expectation fulfillment on patient outcomes. Arch Fam Med. 2000;9:1148-1155 PMID- 11115223 TI - Racial and ethnic disparities in perceptions of physician style and trust. AB - CONTEXT: While pervasive racial and ethnic inequalities in access to care and health status have been documented, potential underlying causes, such as patients' perceptions of their physicians, have not been explored as thoroughly. OBJECTIVE: To assess whether a person's race or ethnicity is associated with low trust in the physician. DESIGN, SETTING, AND PARTICIPANTS: Data were obtained from the 1996 through 1997 Community Tracking Survey, a nationally representative sample. Adults who identified a physician as their regular provider and had at least 1 physician visit in the preceding 12 months were included (N = 32,929). MAIN OUTCOME MEASURE: Patients' ratings of their satisfaction with the style of their physician and their trust in physicians. The Satisfaction With Physician Style Scale measured respondents' perceptions of their physicians' listening skills, explanations, and thoroughness. The Trust in Physician Scale measured respondents' perceptions that their physicians placed the patients' needs above other considerations, referred the patient when needed, performed unnecessary tests or procedures, and were influenced by insurance rules. RESULTS: After adjustment for socioeconomic and other factors, minority group members reported less positive perceptions of physicians than whites on these 2 conceptually distinct scales. Minority group members who lacked physician continuity on repeat clinic visits reported even less positive perceptions of their physicians on these 2 scales than whites. CONCLUSIONS: Patients from racial and ethnic minority groups have less positive perceptions of their physicians on at least 2 important dimensions. The reasons for these differences should be explored and addressed. Arch Fam Med. 2000;9:1156-1163 PMID- 11115224 TI - The physician-patient relationship: three psychodynamic concepts that can be applied to primary care. AB - Psychodynamic concepts can be used to help understand and manage certain difficulties that arise within the physician-patient relationship. The concepts of transference, countertransference, and action (in the form of acting out and enactment) are discussed. A case description is included to show how these concepts apply to the day-to-day practice of primary care medicine. Arch Fam Med. 2000;9:1164-1168 PMID- 11115225 TI - Enhancing drug compliance in lipid-lowering treatment. AB - Hyperlipidemia and the atherosclerotic conditions that result from it are well recognized as major contributors to coronary heart disease (CHD). Fortunately, several large-scale clinical trials have shown that there are effective treatments that can substantially lower atherogenic lipid levels and thereby reduce the risk of CHD mortality and morbidity. However, duplication of these dramatic trial results can be negatively affected in "real life" clinical practice by an important issue: compliance. No medications will work if patients do not take them. Unfortunately, patients who need lipid-lowering therapy are likely to need it long-term, perhaps for a lifetime. Yet, many do not adhere to the prescribed medication regimen. This article reviews some major studies of compliance for lipid-lowering drugs. The reasons why patients do not take them as prescribed vary: poor education, lack of understanding, cost, provider indifference, and others. Achieving compliance requires a multifaceted approach. It can be enhanced by encouraging patients to talk openly about their medication habits and by convincing them of the long-term benefits of reaching and maintaining target low-density lipoprotein cholesterol levels. Although more studies focusing on compliance specifically regarding CHD are needed, the current literature does provide some guidance. Arch Fam Med. 2000;9:1169-1175 PMID- 11115226 TI - Gaps in end-of-life care. AB - Every year, more than 1 million Americans die of different causes. Some die easily and comfortably. Others die with a great deal of suffering and distress. This article contrasts key aspects of the way Americans die with the way they say they would like to die. It will also highlight some of the barriers to providing high-quality end-of-life care. Arch Fam Med. 2000;9:1176-1180 PMID- 11115227 TI - Advance care planning. AB - Advance care planning is the process of planning for future medical care, particularly for the event when the patient is unable to make his or her own decisions. It should be a routine part of standard medical care and, when possible, conducted with the proxy decision maker present. It is helpful to think of the process as a stepwise approach. The steps include the appropriate introduction of the topic, structured discussions covering potential scenarios, documentation of preferences, periodic review and update of the directives, and application of the wishes when needed. The steps can be integrated flexibly into routine clinical encounters by the physician and other members of the health care team. The process fosters personal resolution for the patient, preparedness for the proxy, and effective teamwork for the professionals. The process also has pitfalls of which to be aware. Arch Fam Med. 2000;9:1181-1187 PMID- 11115228 TI - Microscopic polyangiitis in a pediatric patient. AB - Microscopic polyangiitis (MPA), previously called hypersensitivity angiitis, is a systemic necrotizing vasculitis that involves many organ systems including the skin, joints, kidneys, and lungs. Microscopic polyangiitis most commonly affects adults in the fourth and fifth decades of life, with only a few cases reported in children. We describe a pediatric patient with microscopic polyangiitis. Arch Fam Med. 2000;9:1189-1192 PMID- 11115229 TI - Case description of ascariasis. AB - Ascaris lumbricoides are among the medically important worms belonging to the phylum Nematoda (roundworms) that are parasites of the human gastrointestinal tract. Despite current sanitation and hygiene standards in the United States, infection due to intestinal roundworms is not uncommon in children and adults. A high index of suspicion is warranted as patients may present anywhere along a spectrum of illness from asymptomatic to acutely ill. The following is a case presentation and discussion of Ascaris lumbricoides, the common roundworm or intestinal longworm. Arch Fam Med. 2000;9:1193-1194 PMID- 11115231 TI - Living in medicine: med school after 50 years plus 2 weeks PMID- 11115230 TI - Phytophotodermatitis: a sometimes difficult diagnosis. AB - Phytophotodermatitis may not be diagnosed when a patient is seen with erythema and vesicles on the skin. However, with the appropriate medical history, the diagnosis of phytophotodermatitis is easily made. Arch Fam Med. 2000;9:1195-1196 PMID- 11115232 TI - The dilemma of reinstituting anticoagulation for patients with cardioembolic sources and intracranial hemorrhage: how wide is the strait between Skylla and Karybdis? PMID- 11115233 TI - The insula and cerebrogenic sudden death. AB - Sudden death is an "electrical accident" caused by fatal cardiac arrhythmias. While brain-heart control has physiological advantages, cerebrogenic sudden death and nonfatal cardiovascular disturbances can complicate stroke of all types, seizures and epilepsy, head injury, other neurological conditions, neurosurgical procedures, and intense emotional states. Cerebrogenic cardiovascular and autonomic disturbances include electrocardiographic changes, elevation of cardiac enzymes, cardiac arrhythmias, disturbances of blood pressure regulation, and cerebrogenic pulmonary edema. Evidence from experimental studies and clinical observations indicates a crucial role of the insula in cerebrogenic cardiovascular disturbances and sudden death. Future studies should focus on identification of at-risk patients, confirmation of a vulnerable period of cerebrogenic sudden death in those with different neurological conditions and intense emotional states, and clarification of the neurochemical mediators. PMID- 11115234 TI - Epilepsy and the immune system. AB - OBJECTIVE: To discuss evidence that immune mechanisms are involved in the pathogenesis of some forms of epilepsy. DATA SOURCES: Computerized data sources and published indexes and articles. STUDY SELECTION: Published reports showing disorders of the immune system in patients with epilepsy and in animals with experimental epilepsy. DATA SYNTHESIS: Rasmussen encephalitis is an example of an autoimmune disorder of the central nervous system. Serum samples of patients with this disease contain antibodies to the glutamate receptor GluR3, and immunization of animals with GluR3 induces a disorder resembling the human disease. There are still few data to prove that immune mechanisms are involved in the pathogenesis of intractable childhood epilepsies other than Rasmussen encephalitis. Epilepsy is more common in patients with systemic lupus erythematosus who have antiphospholipid antibodies, and it is possible that these antibodies can lead to immune-mediated cortical damage. Immune defects in patients with epilepsy may occur as a consequence of long-term antiepileptic treatment or may represent a genetic coupling to the convulsive disorder. CONCLUSION: The finding of an immunological basis may offer new modalities for the treatment of selected cases of intractable partial epilepsies. PMID- 11115235 TI - Transgenic mouse models and human neurodegenerative disorders. PMID- 11115236 TI - Safety of discontinuation of anticoagulation in patients with intracranial hemorrhage at high thromboembolic risk. AB - BACKGROUND: Limited data are available to guide the management of anticoagulation in patients with intracranial hemorrhage (ICH) at high thromboembolic risk. OBJECTIVE: To review the management of anticoagulation in patients with ICH at high thromboembolic risk. PATIENTS AND METHODS: We reviewed the management of anticoagulation in 141 patients who have a high risk of ischemic stroke and have ICH while taking warfarin. The 30-day risk of ischemic stroke while not taking anticoagulation treatment was determined using Kaplan-Meier survival estimates. RESULTS: The indications for anticoagulation were a prosthetic heart valve (52 patients [group 1]), atrial fibrillation and cardioembolic stroke (53 patients [group 2]), and a recurrent transient ischemic attack or an ischemic stroke (36 patients [group 3]). A prior ischemic stroke occurred in 14 (27%) of group 1 patients and in 23 (43%) of group 2 patients. Death occurred in 43% of the 141 patients. The median time not taking warfarin in this cohort was 10 days. Three patients had an ischemic stroke within 30 days of warfarin therapy discontinuation. Using Kaplan-Meier survival estimates, the probability of having an ischemic stroke at 30 days following warfarin therapy cessation in groups 1, 2, and 3 was 2.9% (95% confidence interval, 0%-8.0%), 2.6% (95% confidence interval, 0%-7.6%), and 4.8% (95% confidence interval, 0%-13.6%), respectively. In the 35 patients who had warfarin therapy restarted, none had recurrence of ICH during the same hospitalization. CONCLUSIONS: Discontinuation of warfarin therapy for 1 to 2 weeks has a comparatively low probability of embolic events in patients at high embolic risk. This should be taken into consideration when deciding whether to continue or discontinue anticoagulation in these patients at high embolic risk. Early recurrence of ICH is exceedingly uncommon. PMID- 11115237 TI - Premorbid reading activity and patterns of cognitive decline in Alzheimer disease. AB - BACKGROUND: Educational and occupational attainment have been associated with progression of Alzheimer disease in some studies. One hypothesis about this association is that education and occupation are markers for lifelong participation in cognitively stimulating activities like reading. OBJECTIVE: To assess the relation of premorbid reading activity with patterns of cognitive decline in Alzheimer disease. METHODS: During a 4-year period, 410 persons with Alzheimer disease had annual clinical evaluations, which included administration of 17 cognitive function tests from which global, verbal, and nonverbal summary measures were derived. At baseline, a knowledgeable informant was questioned about the affected person's reading frequency and access to reading materials before dementia onset. RESULTS: A composite measure of premorbid reading activity was developed. It had moderately high internal consistency and was positively correlated with education and baseline level of cognitive function. In analyses that controlled for baseline cognitive function, education, and other demographic variables, higher level of premorbid reading activity was associated with more rapid decline on the global cognitive and verbal measures but not on the nonverbal measure. CONCLUSIONS: These results suggest that both the extent and nature of premorbid cognitive experiences may affect how Alzheimer disease pathology is clinically expressed. PMID- 11115238 TI - Altered mental status in patients with cancer. AB - OBJECTIVE: To identify the causes of an altered mental status in a cancer population. METHODS: We studied 140 confused patients with cancer (100 prospectively and 40 retrospectively) between January 1, 1991, and June 30, 1992, to determine clinical findings, causes, and outcome. RESULTS: All patients had non-central nervous system cancers. The most common primary cancer types were lung (20%), gastrointestinal tract (18%), leukemia and lymphoma (17%), and breast (11%). Median patient age was 73 years, and 49% were men. Disseminated systemic metastases were present in 50% of patients; 34% were confused at hospital admission and 66% developed confusion during hospitalization. Symptoms included lethargy or coma in 61% of patients, agitation in 44%, disorientation in 83%, lateralizing signs in 41%, delusions or hallucinations in 28%, and seizures in 9%. A single cause of the altered mental status was found in 33% of patients, whereas 67% had multiple causes. Drugs, especially opioids, were associated with altered mental status in 64% of patients, metabolic abnormalities in 53%, infection in 46%, and recent surgery in 32%. A structural brain lesion was the sole cause of encephalopathy in 15% of patients. Although delirium improved in 67% of patients, it was a poor prognostic factor for overall outcome. Thirty-day mortality was 25%, and 44% of patients died within 6 months, usually from progression of the underlying cancer. Prolonged delirium suggested infection or coagulopathy. Younger patients and those with hypoxemia or kidney or liver dysfunction were more likely to die (P<.05). CONCLUSION: Patients with cancer usually have multiple causes of delirium, many of which are treatable, with rapid improvement in their cognitive status. PMID- 11115239 TI - Buccal hemineglect. AB - OBJECTIVES: To determine whether the peripersonal and intrapersonal buccal space can be affected by a hemispheric stroke and to evaluate the clinical signs resulting from buccal neglect. METHODS: A prospective study comparing 2 groups of patients with hemiplegia, 1 with a right hemispheric lesion and the other with a left hemispheric lesion. Patients were selected consecutively on the basis of specific criteria at least 1 month after stroke. RESULTS: Buccal hemineglect was usually concomitant with other hemineglect phenomena resulting from lesions of the right hemisphere (10 of 12 in right lesions and 1 of 12 in left lesions). Clinical signs associated with this condition consisted of impaired swallowing (retention, defective insalivation, presence of food debris in the left hemibuccal space, loss of saliva from the left side of the mouth, and choking); loss of the ability to perceive salty, sweet, or acid tastes; and impaired buccal representation. These problems were usually incorrectly diagnosed initially. Outcome was usually favorable, but functional disorders persisted in some patients for more than 18 months. The underlying attention and representation mechanisms are discussed with reference to experimental lesions of the postarcuate (area 6) cortex in rhesus monkeys. The area around the mouth may be considered to be, as in monkeys, a peripersonal space, ie, probably of little functional importance. The lesion may involve area 6 or its projections to the thalamus or posterior parietal cortex. CONCLUSIONS: Buccal hemineglect, which is likely to cause social embarrassment, should be considered whenever the oral phase of swallowing is impaired in a context of neglect syndromes. Prophylactic measures and rehabilitation can reduce the impact and complications of the condition (food bolus). PMID- 11115240 TI - Chronic inflammatory demyelinating polyradiculoneuropathy: a study of proposed electrodiagnostic and histologic criteria. AB - OBJECTIVE: To examine the sensitivity of the 3 proposed electrodiagnostic (EDX) criteria for demyelination, the sensitivity and specificity of the proposed Ad Hoc Subcommittee of the American Academy of Neurology AIDS [Acquired Immunodeficiency Syndrome] Task Force histologic criteria (AAN criteria), the degree of agreement among these criteria, and the diagnostic value of sural nerve histologic criteria in patients with idiopathic chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). DESIGN AND METHODS: A retrospective analysis of 24 patients with idiopathic CIDP and 12 patients with diabetic polyneuropathy (DP) who underwent comparable testing of clinical, histologic, and EDX features. RESULTS: We found 42%, 50%, and 79% sensitivity of the proposed EDX, AAN teased fiber, and AAN electron microscopic (EM) criteria, respectively, for demyelination in CIDP. The specificity of the proposed AAN teased fiber and EM criteria for demyelination was greater than 80% when tested against patients with DP. There was lack of agreement between the EDX and histologic criteria. Almost two thirds of patients with CIDP who met the EM criteria but none of the EDX criteria for demyelination showed a favorable response to immunomodulatory therapy. CONCLUSIONS: Sural nerve histologic criteria offer unique sensitivity and acceptable specificity toward the diagnosis of CIDP. Sural nerve biopsy should be considered when a clinical suspicion of CIDP remains in patients who do not meet the proposed EDX criteria for demyelination. PMID- 11115241 TI - Magnetic resonance imaging in the clinical diagnosis of Creutzfeldt-Jakob disease. AB - OBJECTIVE: To evaluate the diagnostic usefulness of magnetic resonance imaging (MRI) in the clinical diagnosis of Creutzfeldt-Jakob disease (CJD). BACKGROUND: Creutzfeldt-Jakob disease is a rare neurodegenerative disease that belongs to the group of human spongiform encephalopathies and usually affects elderly people. It is clinically characterized by rapidly progressive dementia and development of neurological symptoms, such as myoclonus or ataxia. Until now, neuroradiologic investigations have only played a minor role in establishing the clinical diagnosis of CJD, and they are often performed to exclude differential diagnoses. SETTING: A university hospital, base of the German National Creutzfeldt-Jakob Disease Surveillance Study. METHODS AND PATIENTS: In this study, MRIs from suspected cases of CJD were examined by one investigator blinded to the diagnosis. Patients were classified according to the established clinical and neuropathological criteria. RESULTS: Bilateral symmetric, high signal intensities on T2-weighted MRIs were present in the basal ganglia of 109 (67%) of 162 patients with CJD. In the control group, which consisted of non-CJD dementia patients, these abnormalities on T2-weighted MRIs were found in 4 (7%) of 58 patients. This corresponds to a high specificity in the differential diagnosis of CJD. CONCLUSION: These results indicate that MRI is a useful and valuable tool with reasonable sensitivity (67%) and high specificity (93%) and should be considered as an additional cornerstone in the clinical diagnosis of CJD. PMID- 11115242 TI - Paradoxical embolism to the basilar apex associated with May-Thurner syndrome. AB - BACKGROUND: Embolic occlusion of intracranial vessels can be caused by material arising proximally, most commonly from the heart, the aorta, or the carotid or vertebral arteries, and rarely from systemic veins. May-Thurner syndrome is an uncommon condition in which there is impaired venous return because of compression of the left common iliac vein by the overlying right common iliac artery, resulting in iliofemoral deep venous thrombosis. OBJECTIVE: To describe a young patient with presumed paradoxical embolism to the basilar apex associated with a patent foramen ovale and May-Thurner syndrome. DESIGN: Single case report. RESULTS: A 16-year-old girl with a history of bulimia and oral contraceptive use had a "top of the basilar" syndrome. She was found to have a patent foramen ovale on transthoracic and transesophageal echocardiography. Magnetic resonance venography of the lower extremities revealed May-Thurner syndrome. Antiphospholipid antibodies (antiphosphatidylserine, anticardiolipin, and antiphosphatidyl-ethanolamine), factor V Leiden mutation by polymerase chain reaction, and homocyst(e)ine levels were normal. Anticoagulation with intravenous unfractionated heparin sodium followed by warfarin sodium was used, resulting in resolution of her neurologic deficits. CONCLUSIONS: Deep venous thrombosis is notorious for its variable clinical manifestations and the potential dire consequences of a missed diagnosis. Physicians caring for patients with presumed paradoxical embolism should assess for May-Thurner syndrome. PMID- 11115243 TI - Correlation of clinical and neuroimaging findings in a case of rabies encephalitis. AB - BACKGROUND: Rabies encephalitis is a feared, virtually uniformly fatal form of central nervous system infection. The incidence of rabies encephalitis in the United States is almost certainly underestimated because of the predominance of bat-borne rabies, which can be spread without traumatic exposure. Because of its rarity in developed countries, rabies encephalitis has been seldom studied with modern imaging techniques. SETTING: University-based teaching hospital. PATIENT: A case of pathologically confirmed rabies encephalitis is presented. Diagnosis of rabies was made by seroconversion testing while the patient was alive and was confirmed postmortem by the presence of rabies antigens and Negri bodies in the brain. The patient had 2 magnetic resonance studies done that showed dramatic abnormalities in the medulla and pons that correlated with features of the neurologic examination and hypothalamic-pituitary abnormalities. RESULT: The patient had a fulminant encephalitic course that ended in death. CONCLUSION: Rabies is an uncommon cause of fatal encephalitis. Anatomic imaging studies such as computed tomographic and magnetic resonance scans have generally been negative in confirmed cases of rabies. We report a case of confirmed rabies with extensive brainstem and hypothalamic-pituitary abnormalities on magnetic resonance imaging. Although these findings are nonspecific, they should raise the clinical suspicion of rabies in the setting of aggressive encephalitis of unclear cause, and appropriate diagnostic tests should be performed. PMID- 11115244 TI - Scrub typhus encephalomyelitis with prominent focal neurologic signs. AB - BACKGROUND: Encephalomyelitis with prominent focal neurologic signs and associated neuroradiologic abnormalities has not been previously described in scrub typhus. CASE DESCRIPTION: A 22-year-old woman was admitted because of fever and an altered mental state. Neurologic examination revealed bilateral sixth and seventh nerve palsies, bilateral gaze evoked nystagmus, anarthria, dysphagia, quadriparesis, and sensory level at T1. Serum and cerebrospinal fluid samples were positive for tsutsugamushi antibody. The patient's magnetic resonance images demonstrated the lesions responsible for the neurologic dysfunctions: in the lower brainstem, cerebellar peduncles, and spinal cord. It was interesting that the gray matter of the spinal cord was predominantly involved. CONCLUSIONS: The recognition of unusual manifestations and the clinical suspicion of this treatment-responsive disease may be important, particularly in the face of increasing international and intranational travel. PMID- 11115245 TI - Global aphasia without hemiparesis secondary to Kingella kingae endocarditis. PMID- 11115246 TI - Posttraumatic headache: a neurobehavioral disorder. PMID- 11115247 TI - Posttraumatic headache--a myth? PMID- 11115248 TI - Posttraumatic headache PMID- 11115249 TI - And the controversies Go On PMID- 11115250 TI - The rise and fall of bromide therapy in epilepsy. PMID- 11115251 TI - A World Health Organization perspective on neurology and neuroscience. PMID- 11115252 TI - Comment on Berry and Hachinski. Chronic whiplash syndrome as a functional disorder. PMID- 11115253 TI - Comment on berry and hachinski PMID- 11115254 TI - Visual hallucinations and dementia with Lewy bodies. PMID- 11115255 TI - Psychological stress and other potential triggers for recurrences of herpes simplex virus eye infections. Herpetic Eye Disease Study Group. AB - OBJECTIVE: To assess psychological stress and other factors as possible triggers of ocular herpes simplex virus (HSV) recurrences. DESIGN: A prospective cohort study nested in a randomized, placebo-controlled, clinical trial. SETTING: Fifty eight community-based or university sites. PARTICIPANTS: Immunocompetent adults (N = 308), aged 18 years or older, with a documented history of ocular HSV disease in the prior year and observed for up to 15 months. EXPOSURE VARIABLES: Psychological stress, systemic infection, sunlight exposure, menstrual period, contact lens wear, and eye injury recorded on a weekly log. The exposure period was considered to be the week before symptomatic onset of a recurrence. MAIN OUTCOME MEASURE: The first documented recurrence of ocular HSV disease, with exclusion of cases in which the exposure week log was completed late after the onset of symptoms. RESULTS: Thirty-three participants experienced a study outcome meeting these criteria. Higher levels of psychological stress were not associated with an increased risk of recurrence (rate ratio, 0.58; 95% confidence interval, 0.32-1.05; P =.07). No association was found between any of the other exposure variables and recurrence. When an analysis was performed including only the recurrences (n = 26) for which the exposure week log was completed late and after symptom onset, there was a clear indication of retrospective overreporting of high stress (P =.03) and systemic infection (P =.01). Not excluding these cases could have produced incorrect conclusions due to recall bias. CONCLUSIONS: Psychological stress does not appear to be a trigger of recurrences of ocular HSV disease. If not accounted for, recall bias can substantially overestimate the importance of factors that do not have a causal association with HSV infection. PMID- 11115256 TI - Papilledema and obstructive sleep apnea syndrome. AB - OBJECTIVES: To characterize the pathogenesis and clinical features of optic disc edema associated with obstructive sleep apnea syndrome (SAS). METHODS: A series of 4 patients with SAS and papilledema (PE) underwent complete neuro ophthalmologic evaluation and lumbar puncture. In 1 patient, continuous 24-hour intracranial pressure (ICP) monitoring was also performed. RESULTS: All 4 patients had bilateral PE that was asymmetric in 2. Three patients had optic nerve dysfunction, asymmetric in 1, unilateral in 2. Daytime cerebrospinal fluid pressure measurements were within normal range. Nocturnal monitoring performed in one patient, however, demonstrated repeated episodes of marked ICP elevation associated with apnea and arterial oxygen desaturation. CONCLUSIONS: We propose that PE in SAS is due to episodic nocturnal hypoxemia and hypercarbia resulting in increased ICP secondary to cerebral vasodilation. In these individuals, intermittent ICP elevation is sufficient to cause persistent disc edema. These patients may be at increased risk for developing visual loss secondary to PE compared with patients with obesity-related pseudotumor cerebri because of associated hypoxemia. The diagnosis of SAS PE may not be appreciated because daytime cerebrospinal fluid pressure measurements are normal and because patients tend to present with visual loss rather than with symptoms of increased ICP. PMID- 11115257 TI - Correlation of the Schirmer 1 and fluorescein clearance tests with the severity of corneal epithelial and eyelid disease. AB - OBJECTIVE: To evaluate the correlations of the fluorescein clearance test (FCT) and the Schirmer 1 test with the severity of corneal epithelial and eyelid disease in normal patients and patients with tear film disorders due to meibomian gland disease (MGD) and/or aqueous tear deficiency (ATD). METHODS: Nineteen normal control subjects, 16 patients with MGD associated with rosacea, 21 patients with noninflammatory atrophic MGD, and 43 patients with ATD were enrolled. There was a similar age and sex distribution in each group. Each patient completed a symptom severity questionnaire that consisted of 11 questions and then underwent a panel of diagnostic tests in the following order: assessment of corneal and conjunctiva sensation with the Cochet-Bonnet esthesiometer, FCT, assessment of corneal fluorescein staining, Schirmer 1 test (5 minutes without anesthesia), and biomicroscopic examination of the eyelid margins and meibomian glands. The FCT was performed with a fluorophotometer by measuring the fluorescein concentration in minimally stimulated tear samples collected from the inferior tear meniscus. By studying the best area under the receiver operating characteristic curves, we developed a formula that combined the FCT and Schirmer test results, which we termed the FCT corrected by Schirmer test. RESULTS: The FCT showed stronger correlation with ocular irritation symptoms (r = 0. 35, P <.001), corneal fluorescein staining (r = 0.54, P<.001), and meibomian gland and eyelid pathologic signs than the Schirmer 1 test. A correction factor that was based on the best area under the receiver operating characteristic curves, added to the FCT score, improved its correlation with ocular irritation symptoms, eyelid margin and meibomian gland pathologic signs, and ocular surface sensitivity scores. CONCLUSIONS: Corneal epithelial disease is correlated with decreased aqueous tear production and delayed tear clearance, whereas eyelid and MGD are correlated with delayed tear clearance. The FCT corrected by Schirmer 1 test improves the correlations of the FCT with ocular irritation symptoms, corneal epithelial and eyelid pathologic signs, and corneal and conjunctival sensitivity for patients with MGD and ATD. PMID- 11115258 TI - The advanced glaucoma intervention study, 6: effect of cataract on visual field and visual acuity. The AGIS Investigators. AB - OBJECTIVE: To investigate the effect of cataract on visual function and the role of cataract in explaining a race-treatment interaction in outcomes of glaucoma surgery. METHODS: The Advanced Glaucoma Intervention Study (AGIS) enrolled 332 black patients (451 eyes) and 249 white patients (325 eyes) with advanced glaucoma. Eyes were randomly assigned to an argon laser trabeculoplasty (ALT) trabeculectomy-trabeculectomy sequence or a trabeculectomy-ALT-trabeculectomy sequence. From the AGIS experience with cataract surgery during follow-up, we estimated the expected change in visual function scores from before cataract surgery to after cataract surgery. Then, for eyes with cataract not removed, we used these estimates of expected change to adjust visual function scores for the presumed effects of cataract. In turn, we used the adjusted scores to obtain cataract-adjusted main outcome measures. MAIN OUTCOME MEASURES: Average percent of eyes with decrease of visual field (APDVF) and average percent of eyes with decrease of visual acuity (APDVA). RESULTS: Within the 2 months before cataract surgery, visual acuity was better in eyes of white patients than of black patients by an average of approximately 2 lines on the visual acuity test chart. Cataract surgery improved visual acuity and visual field defect scores, with the amounts of improvement greater when preoperative visual acuity was lower. Adjustments for cataract brought about the following relative reductions: for APDVF, a relative reduction of 5% to 11% in black patients and 9% to 11% in white patients; for APDVA, a relative reduction of 45% to 49% in black patients and 31% to 38% in white patients; and for the APDVF and APDVA race-treatment interactions, relative reductions of 25% and 45%, respectively. CONCLUSIONS: On average, visual function scores improved after cataract surgery. The findings of reduced race-treatment interactions after adjustment for cataract do not alter our earlier conclusion that the AGIS 7-year results support use of the ALT trabeculectomy-trabeculectomy sequence for black patients and of the trabeculectomy-ALT-trabeculectomy sequence for white patients without life threatening health problems. The choice of treatment should take into account individual patient characteristics and needs. PMID- 11115260 TI - Retinal detachment in the endophthalmitis vitrectomy study. AB - OBJECTIVES: To assess the frequency of retinal detachment following postcataract endophthalmitis and to evaluate the results of management of these detachments. METHODS: Prospective data collected as part of the Endophthalmitis Vitrectomy Study were analyzed. The study was a randomized clinical trial testing the roles of vitrectomy and intravenous antibiotics in restoring vision in patients with endophthalmitis following cataract surgery. RESULTS: The incidence of retinal detachment was 8.3% after treatment of endophthalmitis, with no difference in frequency based on whether initial management was vitrectomy or tap biopsy. The frequency of detachment was higher with more virulent organisms, poor presenting visual acuity, an open posterior capsule at presentation, and in patients who required an early additional procedure in the management of their endophthalmitis. Retinal detachment resulted in a poor visual outcome, with only 27% of patients achieving 20/40 final visual acuity compared with 55% of patients who did not develop detachment. Patients who were able to undergo surgery for their detachment had a better result, with 38% achieving 20/40 final visual acuity. Anatomic success after surgical repair of detachment was achieved in 78% of patients. CONCLUSION: Retinal detachment is a poor prognostic indicator following endophthalmitis, but surgical repair can salvage excellent vision in a substantial number of patients. PMID- 11115259 TI - Lens opacifications detected by slitlamp biomicroscopy are associated with exposure to organic nitrate explosives. AB - CONTEXT: Unusual cataracts (flecks) have been reported to occur at very low levels of trinitrotoluene exposure, but prevalence estimates vary widely. Cataracts have not been reported among workers in the United States exposed to organic nitrate explosives. OBJECTIVES: To determine the prevalence of unusual cataracts in a population of workers in the United States exposed to organic nitrate explosives, to determine whether associations exist with reported cataract risk factors, and to determine if other eye effects (eg, retinal hemorrhage) are associated with exposure. DESIGN: Cohort prevalence study. SETTING: A university-based ophthalmologic clinic. SUBJECTS: Sixty-one workers from an explosives plant comprised the exposed group. The comparison group consisted of 56 workers using chemicals other than organic nitrate explosives. OUTCOME MEASURES: The primary outcome measure was opacifications (flecks) of the crystalline lens, graded clinically on a scale of 0 to 4 +. Additional measures included visual acuity, applanation tonometry, and clinical evaluation using standard examination techniques. RESULTS: Sixty-three percent of the workers had anterior cortical lens opacifications in a pattern of peripheral flecks. Exposed subjects were 18 times more likely to exhibit changes than those not exposed, a statistically significant association (95% confidence interval [CI], 5.0-65.0; P<.001). A statistically significant association with the duration of exposure was also found. CONCLUSIONS: Asymptomatic, low-grade cataracts (flecks) were identified in 63% of the workers exposed to pentolite. No other eye effects were found to be associated with exposure. Cataracts were not associated with other known risk factors, but were associated with the duration of exposure. Biomicroscopy is widely available and useful for detecting changes in the asymptomatic stages. PMID- 11115261 TI - Topical soluble tumor necrosis factor receptor type I suppresses ocular chemokine gene expression and rejection of allogeneic corneal transplants. AB - OBJECTIVE: To determine the effect of topical soluble tumor necrosis factor receptor type I (sTNFR-I) on survival of murine orthotopic corneal transplants and on ocular chemokine gene expression after corneal transplantation. METHODS: BALB/c mice (N = 50) were used as recipients of multiple minor H-disparate corneal transplants from B10.D2 donors. After orthotopic corneal transplantation, mice were randomized in a masked fashion to receive either topical sTNFR-I or vehicle 3 times daily, and all grafts were evaluated for signs of rejection and neovascularization by slitlamp biomicroscopy for 8 weeks. Ocular chemokine gene expression in sTNFR-I- and vehicle only-treated groups was determined using a multiprobe ribonuclease protection assay. RESULTS: Hosts treated with topical sTNFR-I experienced significantly enhanced corneal allograft survival compared with animals treated with vehicle alone (P =.01). Moreover, postoperative messenger RNA levels of RANTES and macrophage inflammatory protein-1beta in sTNFR I-treated eyes were substantially suppressed compared with vehicle-treated eyes. Vehicle-treated eyes bearing rejected allografts expressed higher levels of messenger RNA for both chemokines than control eyes bearing accepted allografts. CONCLUSIONS: Topical treatment with sTNFR-I promotes the acceptance of allogeneic corneal transplants and inhibits gene expression of 2 chemokines (RANTES and macrophage inflammatory protein-1beta) associated with corneal graft rejection. CLINICAL RELEVANCE: Our findings support the feasibility of a topical anticytokine strategy as a means of reducing corneal allograft rejection without resorting to the use of potentially toxic immunosuppressive drugs. PMID- 11115262 TI - Development of a newly designed double-fixed Seoul-type keratoprosthesis. AB - OBJECTIVE: To develop a newly designed double-fixed keratoprosthesis (Seoul-type keratoprosthesis [S-KPro]) and to assess its mechanical stability and biocompatibility. METHODS: Twenty-five rabbits were divided into 4 groups by fixation technique, amniotic membrane (AM) implantation, and skirt material. The eyes were studied with the use of slitlamp, light, and electron microscopy. Stress testing was performed. In addition, 2 human subjects underwent S-KPro implantation. Best-corrected visual acuity was checked, and ophthalmic examination was performed. RESULTS: The average retention period of the group receiving double-fixated polyurethane-S-KPro with AM was longer (>24 weeks) than that of the others. Fibroblast invasions were found in polyurethane pores but not in polytetrafluoroethylene (Gore-Tex) pores on light microscopy. The minimal pressure that induced aqueous leakage was greater than 250 mm Hg in all of the tested eyes. Two human subjects have maintained a good postoperative condition for 18 and 8 months. CONCLUSIONS: The double-fixation technique of applied S-KPro and AM appears to be helpful in improving the stability of the keratoprosthesis. Polyurethane with relatively large pore size (40 microm) may be used successfully as a material for the keratoprosthesis skirt. CLINICAL RELEVANCE: Our results may be important for improving the clinical outcome of keratoprosthesis. PMID- 11115263 TI - Vitreous surgery simulator. AB - OBJECTIVE: To reduce the surgical risks to patients and expose surgeons to surgical experience and complications, we have developed a practical system of vitreous surgery using virtual-reality technology. METHODS: The system is composed of high-resolution color stereo binoculars, haptic devices, foot switches, and a high-speed graphics computer. To simulate vitreous surgery, we created several virtual patient eyes with retinal diseases such as preretinal membranes and subretinal neovascular tissue at the fovea. RESULTS: The simulator provided the trainees with an operating environment similar to an actual one, and allowed them to learn to maneuver surgical instruments and remove proliferative tissue on the retina, under the retina, or both. This system allowed surgeons to avoid iatrogenic complications through visual signs such as retinal hemorrhage when the instrument contacted the retinal surface. CONCLUSIONS: This simulator may not only be suitable for residents to learn ocular surgical techniques but may also allow veteran surgeons to develop new surgical methods and skills. PMID- 11115264 TI - What triggers recurrences of herpes simplex keratitis. PMID- 11115265 TI - "Pseudotumor Cerebri" by any other name. PMID- 11115266 TI - Laser pointers and the human eye: a clinicopathologic study. AB - We report the absence of photic retinal injury after exposing the retina to light from class 3A laser pointers for durations of up to 15 minutes. Three patients with uveal melanomas were scheduled to have an enucleation. Each agreed to have his or her retina exposed to laser light from a class 3A laser pointer prior to enucleation. Continuous exposure was directed to the fovea for 1 minute, to the retina 5 degrees below fixation for 5 minutes, and to the retina 5 degrees above fixation for 15 minutes. Ophthalmoscopic evaluation of the cornea, lens, and retina and fluorescein angiographic studies of the retina were conducted before, 24 hours after, and 11 days after laser exposure in the first case; before and 86 hours after exposure in the second case; and before, 96 hours after, and 15 days after exposure in the third case. Other than transient afterimages that lasted only a few minutes, we were unable to document any functional, ophthalmoscopic, fluorescein angiographic, or histologic evidence of damage to any structures of the eyes. Transmission electron microscopic studies of retinal sites targeted by the laser pointers in the second and third cases revealed ultrastructural abnormalities in the outer retina and the pigment epithelium that were similar to abnormalities seen in the retina approximately 8 mm away from the targeted sites. The risk to the human eye from transient exposure to light from commercially available class 3A laser pointers having powers of 1, 2, and 5 mW seems negligible. PMID- 11115267 TI - Immunophenotypic shift in a case of mycosis fungoides with vitreous invasion. AB - The case of an 82-year-old man who developed intraocular extension from mycosis fungoides, a cutaneous T-cell lymphoma, is presented. The patient died soon after intraocular involvement occurred. Immunohistochemistry of a skin biopsy, taken early in the course of the disease, disclosed a predominance of T cells with a helper/inducer phenotype (CD4(+)). However, an intraocular infiltrate obtained 7 years later contained mostly T cells with a suppressor/cytotoxic phenotype (CD8(+)). The occurrence of ocular invasion, the change in immunophenotype, and the predominant proliferation of CD8(+) lymphocytes may have been related to the poor outcome in this patient. PMID- 11115268 TI - Retinopathy as the initial presentation of human immunodeficiency virus 2 infection. PMID- 11115269 TI - Optic neuropathy after burns. PMID- 11115270 TI - Nasopalpebral lipoma-coloboma syndrome. PMID- 11115271 TI - Penetrating orbital injury by automobile wiper-control stalk. PMID- 11115272 TI - Subretinal hemorrhage from retinal arterial macroaneurysm simulating malignancy. PMID- 11115273 TI - Submission of data sets to journals: what's the real issue? PMID- 11115274 TI - Photorefractive keratectomy and laser in situ keratomileusis: a word from the devil's advocate. PMID- 11115275 TI - Operative corneal perforations caused by laser in situ keratomileusis. PMID- 11115276 TI - Measuring quality of life in children with obstructive sleep disorders. AB - OBJECTIVE: To validate a disease-specific health-related quality of life (HRQOL) instrument for children with obstructive sleep disorders (OSDs). DESIGN: Prospective cohort study using a 6-item health-related instrument (OSD-6). SUBJECTS: One hundred caregivers of patients with OSDs secondary to adenotonsillar hypertrophy (age range, 2-12 years) from 2 tertiary care, pediatric otolaryngology practices. INTERVENTION: The OSD-6 was administered on initial presentation and 4 to 5 weeks after adenotonsillectomy. A subset of patients repeated the OSD-6 within 3 weeks after presentation to assess test retest reliability. MAIN OUTCOME MEASURES: Test-retest reliability, internal consistency, construct validity, and responsiveness to clinical change of the OSD 6 score. RESULTS: Test-retest reliability was good (intraclass correlation coefficient = 0.74). Median OSD-6 score was 4.5 (0- to 6-point scale) with higher scores indicating poorer quality of life (QOL). Construct validity was demonstrated by the moderate correlation between OSD-6 score and global adenoid and tonsil-related QOL (R = -0.62), strong correlation between the OSD-6 change score and change in global adenoid and tonsil-related QOL (R = -0.63), and the moderate correlation between the change score and parent estimate of clinical change (R = 0.40). The mean change in OSD-6 score after adenotonsillectomy was 3.0 (95% confidence interval, 2.7-3.4). The mean standardized response was 2.3 (95% confidence interval, 1.9-2.7) indicating the instrument's large responsiveness to clinical change. The change score was very reliable (R = 0.85). CONCLUSIONS: The OSD-6 is a reliable, responsive, easily administered instrument. It is valid for detecting change after adenotonsillectomy in children with OSDs. Arch Otolaryngol Head Neck Surg. 2000;126:1423-1429 PMID- 11115277 TI - Masses of the salivary gland region in children. AB - BACKGROUND: Noninflammatory masses of the salivary gland region in children are extremely rare. Therefore, very few published individual and institution-based experiences exist. DESIGN: Retrospective chart review from 1990 through 1997. SETTING: University-based children's hospital. DESIGN: Patients 18 years of age or younger with a tumor in the salivary gland region. Masses of infectious origin were excluded. Hemangiomas and lymphangiomas were tallied for relative incidences only. RESULTS: Three hundred twenty-four consecutive cases of salivary gland masses were found: 192 hemangiomas (59.2%), 89 lymphangiomas (27.5%), and 43 (13.3%) solid masses. No significant difference was found between the age at presentation of the patients with benign solid tumors and the patients with malignant solid tumors (mean + SEM age, 7.2 + 0.7 years). Sixty-one percent of the masses were found in the parotid region; 18% were localized to the submandibular gland region; and the remaining 21% were located in a minor salivary gland site. The most common benign perisalivary masses were pilomatrixomas (20.9%), followed by pleomorphic adenomas (11.6%). The most common malignant masses were mucoepidermoid carcinomas (9.3%), followed by rhabdomyosarcomas (7.0%). Treatment was individualized to the disease. Twenty-two patients had adequate data for follow-up analysis (mean + SEM follow-up, 30.0 + 8.4 months). Four patients (18.2%) experienced recurrent or residual disease and were alive with disease at last follow-up, and 100% of our population demonstrated disease-specific survival at last follow-up. CONCLUSIONS: Vascular lesions outnumber solid tumors of the salivary gland region. The most common salivary tumors were pleomorphic adenomas, followed by mucoepidermoid carcinomas. Although certain solid salivary masses may demonstrate locally aggressive behavior, the overall prognosis is favorable. Arch Otolaryngol Head Neck Surg. 2000;126:1435-1439 PMID- 11115278 TI - Methicillin-resistant Staphylococcus aureus otorrhea after tympanostomy tube placement: an emerging concern. AB - OBJECTIVES: To review the treatment of pediatric patients with methicillin resistant Staphylococcus aureus (MRSA)-positive cultures as a result of otorrhea after tympanostomy tube placement in terms of both medication and isolation strategies and to highlight an emerging problem faced by the clinician with reference to treatment options as well as to the treatment of these patients in an outpatient setting. PATIENTS: Between December 1998 and January 2000, a total of 8 children between the ages of 1 and 11 years had MRSA-positive cultures as a result of otorrhea after tympanostomy tube placement. MAIN OUTCOME MEASURES: The Department of Infectious Diseases was notified, and a variety of topical antibiotic treatments were administered. Arch Otolaryngol Head Neck Surg. 2000;126:1440-1443 PMID- 11115279 TI - The effectiveness of tonsillectomy in diagnosing lymphoproliferative disease in pediatric patients after liver transplantation. AB - OBJECTIVE: To determine the effectiveness of diagnosing forms of lymphoproliferative disease by performing tonsillectomy in pediatric patients who develop symptomatic or asymptomatic tonsillar hypertrophy during immunosuppressive therapy after liver transplantation. DESIGN: Retrospective chart and pathological review. SETTING: Urban tertiary referral children's hospital. MAIN OUTCOME MEASURES: The presence of a pathological stage of lymphoproliferative disease or Epstein-Barr virus (EBV) diagnosed using tonsillar specimens, resulting in a change in therapy. RESULTS: Of 275 pediatric patients who underwent liver transplantation, 13 had tonsillectomy performed with histopathological review of the tonsillar specimens. The specimens from 5 patients (39%) demonstrated pathological changes thought to be consistent with EBV-related changes or a form of lymphoproliferative disease. Histological changes ranged from tonsillar hyperplasia associated with EBV infection to large cell lymphoma. Immunosuppressive therapy was reduced or discontinued, and antiviral therapy was initiated. CONCLUSION: Children who have undergone liver transplantation and develop tonsillar hypertrophy should undergo a diagnostic tonsillectomy, regardless of the clinical presentation, to rule out a form of posttransplant lymphoproliferative disease. Arch Otolaryngol Head Neck Surg. 2000;126:1444-1447 PMID- 11115280 TI - Internal support of tissue-engineered cartilage. AB - BACKGROUND: Auricles previously created by tissue engineering in nude mice used a biodegradable internal scaffold to maintain the desired shape of an ear. However, the biodegradable scaffold incited a compromising inflammatory response in subsequent experiments in immunocompetent animals. OBJECTIVE: To test the hypothesis that tissue-engineered autologous cartilage can be bioincorporated with a nonreactive, permanent endoskeletal scaffold. MATERIALS AND METHODS: Auricular elastic cartilage was harvested from Yorkshire swine. The chondrocytes were isolated and suspended into a hydrogel (Pluronic F-127) at a cell concentration of 5 x 10(7) cells/mL. Nonbiodegradable endoskeletal scaffolds were formed with 1 of 5 polymers: (1) high-density polyethylene, (2) soft acrylic, (3) polymethylmethacrylate, (4) extrapurified Silastic, and (5) conventional Silastic. Three groups were studied: (1) a control group using only the 5 polymers, (2) the 5 polymers enveloped by Pluronic F-127 only, and (3) the implants coated with Pluronic F-127 seeded with chondrocytes. All constructs were implanted subdermally; implants containing cells were implanted into the same animal from which the cells had been islolated. The implants were harvested after 8 weeks of in vivo culture and histologically analyzed. RESULTS: Only implants coated by hydrogel plus cells generated healthy new cartilage. With 3 polymers (high-density polyethylene, acrylic, and extrapurified Silastic), the coverage was nearly complete by elastic cartilage, with minimal fibrocartilage and minimal to no inflammatory reaction. The Food and Drug Administration-approved conventional Silastic implants resulted in fragments of fibrous tissue mixed with elastic cartilage plus evidence of chronic inflammation. The polymethylmethacrylate implant was intermediate in the amount of cartilage formed and degree of inflammation. CONCLUSIONS: This pilot technique combining tissue engineered autologous elastic cartilage with a permanent biocompatible endoskeleton demonstrated success in limiting the inflammatory response to the scaffold, especially to high-density polyethylene, acrylic, and extrapurified Silastic. This model facilitates the potential to generate tissue of intricate shape, such as the human ear, by internal support. Arch Otolaryngol Head Neck Surg. 2000;126:1448-1452 PMID- 11115281 TI - Vestibular neuropathy accompanying auditory and peripheral neuropathies. AB - OBJECTIVE: To define the incidence of measurable vestibular disorders in patients with auditory and peripheral neuropathies. DESIGN: Descriptive study of the case features of auditory neuropathy in 14 patients, 8 of whom had concomitant peripheral neuropathies. SETTING: University referral center. PATIENTS: Fourteen patients aged from 10 to 75 years and diagnosed as having auditory neuropathy, 8 of whom had concomitant peripheral neuropathies. MAIN OUTCOME MEASURES: Incidence of abnormal vestibular caloric test results and the relationship of such incidence to clinical variables including the ages of the subjects, the presence of a concomitant peripheral neuropathy, vestibular symptoms, and audiological findings. RESULTS: Abnormal vestibular caloric test results occurred in 9 of the 14 patients. These 9 patients were on average older (35.6 years) than patients with normal caloric responses (17.8 years). Seven of the 9 patients with abnormal caloric responses had concomitant peripheral neuropathies compared with only 1 of the 5 patients with normal caloric responses. None of the 14 patients experienced symptoms of vestibular disorder. CONCLUSIONS: Asymptomatic vestibular disorders are common in patients with auditory neuropathy when a peripheral neuropathy is also present. The reason for the abnormal vestibular test results is likely a neuropathy of the vestibular nerves. Arch Otolaryngol Head Neck Surg. 2000;126:1453-1456 PMID- 11115282 TI - Diagnosis and staging of head and neck cancer: a comparison of modern imaging modalities (positron emission tomography, computed tomography, color-coded duplex sonography) with panendoscopic and histopathologic findings. AB - OBJECTIVE: To compare the clinical value of positron emission tomography (PET) using fludeoxyglucose F 18, computed tomography (CT), color-coded duplex sonography (CCDS), and panendoscopy in the detection and staging of head and neck cancer. DESIGN: Prospective nonrandomized controlled study. SETTING: Medical school. PATIENTS: Convenience sample of 50 patients with suspected primary or recurrent head and neck cancer. INTERVENTION: Biopsy, tumor surgery. MAIN OUTCOME MEASURES: Information of diagnostic procedures compared with histopathologic features. RESULTS: Both PET and panendoscopy had a sensitivity of 95% and 100% for detection of primary tumor or recurrent carcinomas, respectively. Specificity for PET and panendoscopy was 92% and 85% in primary tumors and 100% and 80% in recurrent carcinoma, respectively. Sensitivity of CCDS and CT was 74% and 68% in primary tumors and 67% and 63% in recurrent carcinomas, respectively. Specificity was 75% and 69% in primary tumors and 100% and 80% in recurrent neoplasms. When assessing neck nodes, all imaging procedures exhibited identical sensitivity (84%). Specificity was 90%, 96%, and 88% in PET, CT, and CCDS, respectively. In recurrent lymph node metastases, sensitivity was 100%, 67%, and 67% and specificity was 87%, 91%, and 87% for PET, CT, and CCDS, respectively. CONCLUSIONS: Positron emission tomography was the most reliable imaging procedure in the detection of primary tumor and recurrent carcinomas localized in the head and neck region. Owing to its limited anatomical depiction, it cannot as yet replace other diagnostic procedures in preoperative planning but does contribute valuable complementary diagnostic information. Computed tomograpy may have difficulties in identifying recurrent carcinomas. For routine diagnosis of nodal spread in the neck, CCDS is recommended. Panendoscopy is a valuable diagnostic procedure that can provide key information in cases of superficial mucosal tumor involvement. Arch Otolaryngol Head Neck Surg. 2000;126:1457-1461 PMID- 11115283 TI - Accuracy of computer navigation in ear, nose, throat surgery: the influence of matching strategy. AB - OBJECTIVE: To measure the effect of 4 different matching strategies on the accuracy of computer navigation on the face and within the nose and rhinopharynx. DESIGN: Survey. SETTING: Laboratory study. SUBJECTS: Six human cadavers studied within 24 hours of death. INTERVENTIONS: A commercially available navigation system with infrared optical tracking was used for computer navigation on the face and within the nose of the subjects after matching with external fiducials or with 3 different configurations of anatomical landmarks. Navigation errors were measured and correlated to matching strategies and compared through statistical analysis. RESULTS: Matching with external fiducials on the face results in smaller navigation error than matching with anatomical landmarks. The configuration of matching strategies with anatomical landmarks also significantly determines the accuracy of computer navigation, especially when different locations of accuracy measurement are considered. CONCLUSION: Statistically significant findings have shown that the choice of a matching strategy is a major factor in the accuracy of computer navigation for ear, nose, throat surgery. Arch Otolaryngol Head Neck Surg. 2000;126:1462-1466 PMID- 11115284 TI - The fibula osteocutaneous flap in head and neck reconstruction: a critical evaluation of donor site morbidity. AB - OBJECTIVES: To (1) compare the complications and functional outcome of primary closure vs split-thickness skin grafting of the fibula osteocutaneous flap donor site, (2) identify patient-mix or treatment factors related to donor site complications, and (3) address early detection and management of donor site complications. DESIGN: Retrospective review and questionnaire study. SETTING: Two university tertiary referral centers. PATIENTS: Fifty-three patients (31 men and 22 women, ages 20 to 80 years) who underwent fibula osteocutaneous free tissue transfer between February 1992 and January 1997. MAIN OUTCOME MEASURES: Minor complications; major complications; and postoperative function, including sensory and motor deficits, pain, swelling, temperature intolerance, and activities of daily living. RESULTS: Four patients developed major wound complications (group 1), 11 patients developed minor wound complications (group 2), and 38 patients had no wound complications (group 3). The donor site was closed primarily in 26 patients and with a split-thickness skin graft in the remaining 27 patients. Major wound complications developed in 3 patients (12%) who underwent primary closure and 1 patient (4%) who underwent split-thickness skin grafting. Minor wound complications developed in 7 (27%) of the patients who underwent primary closure and 4 patients (15%) who underwent split-thickness skin grafting. Three patients who had major complications had residual sensory or motor deficits that resulted in impaired gait or alteration in their daily activities. Comparing all patients with complications (groups 1 and 2) to patients with no complications (group 3) demonstrated an increased incidence of donor site complications in heavy smokers (P<.05) and a strong trend toward higher donor site complications in patients who underwent primary closure (P =.10). Although trends were identified, no significant differences were found in age, comorbid illnesses, alcohol use, preoperative laboratory values, operating time, tourniquet time, or skin paddle width. CONCLUSIONS: A variety of patient-mix and operative factors are likely related to the development of donor site wound complications. Width of the skin paddle alone is not a reliable criterion for determining the need to skin graft the donor site. Primary closure tended to result in a higher rate of both major and minor wound complications compared with split-thickness skin grafting. Primary closure of fibula donor site defects should be undertaken if this can be accomplished with no tension along the suture line. If tension at the suture line is present, a skin graft should be strongly considered to minimize the possibility of a wound complication. Arch Otolaryngol Head Neck Surg. 2000;126:1467-1472 PMID- 11115285 TI - Salvage surgery after failure of nonsurgical therapy for carcinoma of the larynx and hypopharynx. AB - BACKGROUND: For larynx preservation, radiotherapy is gaining popularity for primary treatment of laryngeal and hypopharyngeal cancer, reserving surgery for salvage. OBJECTIVE: To analyze the outcome of salvage surgery after failure of primary radiotherapy. DESIGN: Nine-year retrospective outcome analysis. SETTING: University referral center. PATIENTS: Fifty-four patients with squamous cell carcinoma of the larynx (n = 39) or hypopharynx (n = 15). RESULTS: For laryngeal cancer, mean interval from radiation to detection of recurrence was 14.5 months (range, 2-66 months). Twenty-three patients (59%) presented with a more advanced tumor stage after radiation than at the initial evaluation. Total laryngectomy was needed in 36 patients (92%). Disease-specific 5-year survival rate was 63%. Survival of patients with small recurrent tumors was statistically significantly better than those with advanced tumors (P =.004). For hypopharyngeal cancer, mean interval from radiation to detection of the recurrence was 10.6 months (range, 3 40 months). Total laryngopharyngectomy was needed in 8 of 9 patients with local recurrrence; neck dissection, in 6 patients with regional recurrence. Disease specific 5-year survival rate was only 20%. Recurrent tumor and node stages did not influence the outcome. Patients with regional recurrences did no better than those with local ones. CONCLUSIONS: Salvage surgery in laryngeal cancer achieves good results, especially for small recurrences. Because of tumor progression, larynx preservation is seldom possible at the time of salvage. Salvage surgery in hypopharyngeal cancer shows poor survival regardless of tumor stage and despite radical surgical procedures, and can be recommended only for carefully selected patients. Arch Otolaryngol Head Neck Surg. 2000;126:1473-1477 PMID- 11115286 TI - Accuracy of computed tomography in determining the presence or absence of metastatic retropharyngeal adenopathy. AB - OBJECTIVE: To decide the accuracy of computed tomography in determining the presence or absence of metastatic retropharygeal adenopathy in patients with squamous cell carcinoma of the head and neck. DESIGN: A comparison of the results of retrospective blinded review of preoperative computed tomographic scans with the histologic findings of retropharyngeal node dissection at the time of surgery. SETTING: Academic tertiary care center. PATIENTS: Twenty-six patients with advanced stage squamous cell carcinoma of the head and neck. MAIN OUTCOME MEASURES: Computed tomographic findings and histologic results of retropharyngeal node dissection. RESULTS: The retropharyngeal nodes were pathologically positive for metastasis in 6 (23%) of the 26 patients. The radiologist (J.M.T.) correctly read the scan in 3 of 6 patients with histologically proved metastasis, and in 14 of 20 patients with histologic features negative for metastasis. The sensitivity of the radiologist reading was 50%, and the specificity was 70%. The positive predictive value was 33%, and the negative predictive value was 82%. CONCLUSION: The presence of retropharyngeal node metastasis cannot be determined by computed tomographic imaging alone. Arch Otolaryngol Head Neck Surg. 2000;126:1478-1481 PMID- 11115287 TI - Validity of ultrasonography in diagnosis of acute maxillary sinusitis. AB - BACKGROUND: Accurate diagnosis of maxillary sinusitis is difficult on the basis of clinical examination only because the signs and symptoms of sinusitis are nonspecific. A simple, rapid, and readily available method for diagnosing maxillary sinusitis in primary care would increase the accuracy of the diagnoses and thus reduce unnecessary antibiotic treatment. OBJECTIVE: To investigate the validity of ultrasonography compared with radiography and magnetic resonance imaging (MRI) in detection of maxillary sinusitis. DESIGN: Ultrasonography and plain-film radiography of the paranasal sinuses were performed on all patients and MRI was performed on 40 randomly selected patients on day 7 of the study. SETTING: Study office at the Department of Pediatrics of Turku University Hospital, Turku, Finland. PATIENTS: One hundred ninety-seven young adults who contacted the study office within 48 hours of the onset of symptoms of the common cold. MAIN OUTCOME MEASURES: Detection rates of maxillary sinusitis by ultrasonography, radiography, and MRI. RESULTS: Acute maxillary sinusitis was diagnosed in 24% of the sinuses by radiography and in 28% by MRI. Compared with MRI findings, the sensitivity of ultrasonography for detection of maxillary sinusitis was 64% (specificity, 95%). Using a 2-step diagnostic approach in which radiological findings were additionally considered in cases of negative ultrasound findings, a sensitivity of 86% (specificity, 95%) was observed. CONCLUSIONS: The high specificity of ultrasonography indicates that a positive ultrasound finding can be regarded as evidence of maxillary sinusitis. The addition of plain-film radiography in cases of negative ultrasound findings increases the diagnostic sensitivity to clinically acceptable levels without loss in specificity. Active use of ultrasonography would substantially decrease the need for radiological imaging of the sinuses and also help reduce unnecessary antibiotic treatment in primary care. Arch Otolaryngol Head Neck Surg. 2000;126:1482-1486 PMID- 11115288 TI - Assessment of the efficacy of endoscopy in pituitary adenoma resection. AB - OBJECTIVE: To obtain objective evidence that the use of endoscopy in the surgical management of pituitary tumors improves intraoperative visualization and significantly impacts operative outcomes. DESIGN: Case series of pituitary adenomas treated surgically by endoscope-assisted microscopic resection. SETTING: University-affiliated tertiary care medical center. PATIENTS: Consecutive sample of 9 patients referred for surgical management of pituitary adenoma. INTERVENTIONS: Each patient underwent transseptal transsphenoidal microscopic tumor resection. The procedure was modified by the use of intrasellar endoscopy as an adjunctive imaging modality. Following complete microscopic resection of tumor, rigid 0 degrees and 30 degrees 4.0-mm endoscopes were used to conduct a final survey of the sellar and parasellar spaces. Residual tumor fragments identified during this endoscopic examination were removed. OUTCOME MEASURES: Endoscopes were thought to have a significant impact on surgical therapy in cases where residual tumor that was not detected microscopically was identified and removed during endoscopic examination. Analysis of each case included correlation between intraoperative findings and retrospective review of dictated operative reports and intraoperative videotape. RESULTS: Three of the patients with macroadenoma (33% of total, 43% of macroadenoma cases) had tumor fragments that were only identified and removed endoscopically. CONCLUSIONS: Endoscopy provides distinct advantages over microscopy in imaging intrasellar and parasellar structures during pituitary tumor resection. These data support the numerous anecdotal accounts of the usefulness of pituitary endoscopy and are consistent with the small amount of objective evidence offered on the subject. Arch Otolaryngol Head Neck Surg. 2000;126:1487-1490 PMID- 11115289 TI - Simultaneous vocal fold and tongue paresis secondary to Epstein-Barr virus infection. AB - Dysphonia is a common presenting symptom in cases referred for otolaryngologic evaluation. Similarly, primary care physicians frequently see adolescents or young adults with symptomatic Epstein-Barr virus infection. Some of the patients with active Epstein-Barr virus infection who have severe clinical manifestations of infectious mononucleosis will be referred for otolaryngologic evaluation. Voice abnormalities in these patients, though, are usually limited to altered resonance due to pharyngeal crowding by hyperplastic lymphoid tissue. We describe a patient with infectious mononucleosis who was referred for evaluation of dysphonia and was diagnosed with unilateral tongue and vocal fold paresis. We also discuss the patient's clinical course and review the related literature. Although uncommon, cranial nerve palsies must be considered in the patient with Epstein-Barr virus infection who presents with voice or speech disturbance. Arch Otolaryngol Head Neck Surg. 2000;126:1491-1494 PMID- 11115290 TI - Relapsing polychondritis: a course over 20 years with cerebral involvement. AB - Relapsing polychondritis is a chronic inflammatory disease associated with an autoimmune disorder in cartilaginous tissue, eyes, labyrinth, blood vessels, and central nervous system. We describe a 75-year-old woman who presented with a 20 year history of dyspnea, inspiratory stridor, and polyarthritis. She developed dysmorphism of both ears and a saddle nose approximately 10 years earlier. Subsequently, she suffered from hearing loss and a tremor. A T2-weighted magnetic resonance imaging scan of the brain revealed multiple, spotted signal intensities. Immunohistochemical analysis of a serum sample showed antibodies to cartilaginous tissue, which were further identified on immunoblotting as antibodies to type II collagen. The extremely prolonged course of disease (>20 years) until a correct diagnosis was made is remarkable. Also, cerebral involvement, which was most likely caused by cerebral angiitis, and which, to our knowledge, has never previously been reported in this form, was detected. Arch Otolaryngol Head Neck Surg. 2000;126:1495-1498 PMID- 11115291 TI - Imaging quiz case 1. Otogenic pneumocephalus with temporal bone cerebrospinal fluid (CSF) leak. PMID- 11115292 TI - Imaging quiz case 2. Lemierre syndrome: septic thrombophlebitis of the internal jugular vein, or "postanginal" sepsis. PMID- 11115293 TI - Imaging quiz case 3. Cervical tuberculous lymphadenopathy (scrofula). PMID- 11115294 TI - Imaging quiz case 4. Cervical rib. PMID- 11115295 TI - Swimming with tympanostomy tubes. PMID- 11115296 TI - Swimming with tympanostomy tubes PMID- 11115297 TI - Swimming with tympanostomy tubes: the controversy continues PMID- 11115299 TI - Anticipatory guidance: missed opportunities. PMID- 11115298 TI - Bad science and the role of institutional review boards. PMID- 11115300 TI - Bidi cigarettes: An emerging threat to adolescent health. PMID- 11115301 TI - Anticipatory guidance: what information do parents receive? What information do they want? AB - OBJECTIVE: To determine whether parents are receiving anticipatory guidance, whether they could use more information on anticipatory guidance topics, and how receipt of anticipatory guidance relates to satisfaction with care. DESIGN AND SAMPLE: Analysis of data from a telephone interview of 2017 respondents between July 1995 and January 1996. A stratified random-digit dialing design was used to obtain a nationally representative sample of parents with children between 0 and 3 years old. MAIN OUTCOME MEASURES: Discussions with a physician or nurse about 6 anticipatory guidance topics and whether parents could use more information on these topics. Willingness of parents to pay extra to discuss these topics and receive additional care. Ratings of how well clinicians provide health care. RESULTS: The percentage of parents who had not discussed each subject with a clinician varied by topic: newborn care (< 3 months old), 38%; crying, 65%; sleep patterns, 59%; encouraging learning, 77%; discipline (ages 6-36 months), 75%; and toilet training (ages 18-36 months), 66%. Thirty-seven percent of parents had not discussed any of these topics. Among parents who had not discussed a particular issue, the percentage who reported that they could use more information ranged from 22% for both newborn care and crying to 55% for encouraging learning; similar percentages who had discussed the topics could also use more information. Parents who had discussed more of these topics with a clinician were more likely to report excellent care. Parents who could use more information on a larger number of topics were much more willing to pay for additional care. CONCLUSIONS: Although anticipatory guidance is considered an important component of well-child care, the majority of parents reported that they had not discussed most standard topics with a clinician. Many parents could use more information on these topics. Effort is required to provide parents with the information they need to take good care of their children. Arch Pediatr Adolesc Med. 2000;154:1191-1198. PMID- 11115302 TI - Characterization of methylphenidate exposures reported to a regional poison control center. AB - OBJECTIVE: To investigate the frequency, risk population, symptoms, reason, and outcome surrounding human methylphenidate exposures reported to a regional poison control center. DESIGN: Retrospective case series. SETTING/PATIENTS: All human methylphenidate exposures reported to a regional poison control center during a 2 year period were included. MAIN OUTCOME MEASURES: Data collected included age, dose, reason for exposure, symptoms (type and severity), treatment, and outcome. Age data for all other exposures reported during the same time period were also compiled. RESULTS: The study included 289 patients. Overall, 31% developed symptoms, most commonly tachycardia, agitation, and lethargy. No patient developed severe symptoms, although a less favorable outcome was seen with intentional vs unintentional exposures. When compared with the age data for all other exposures reported during the same time period, a trimodal vs bimodal pattern was seen, with the novel peak occurring in 6- to 9-year-old children. Within this age group, therapeutic error was the most common reason for exposure. CONCLUSIONS: Methylphenidate exposure was associated with symptom development in 31% of cases. An unusually high incidence of therapeutic error was noted surrounding its use, most commonly in the 6- to 11-year-old age group, defining a unique population at risk for toxic exposure. This initial data may allow targeted poisoning prevention efforts for this age group. Arch Pediatr Adolesc Med. 2000;154:1199-1203. PMID- 11115303 TI - Predictive factors for short-term symptom persistence in children after emergency department evaluation for constipation. AB - BACKGROUND: Children with symptoms and signs of constipation are commonly assessed in pediatric emergency departments (EDs). Little is known about their outcome following the ED visit. OBJECTIVES: To describe the clinical characteristics of children presenting to the ED with constipation and the ED interventions; to measure short-term symptom resolution at 48 hours and 7 days after the ED visit; and to identify predictive factors associated with poor symptom resolution at 48 hours and 7 days after the ED visit. DESIGN/METHODS: Cohort study conducted between July 10, 1997, and September 10, 1997, in a tertiary care pediatric hospital ED. All children (aged 1-18 years) with idiopathic constipation were included. Constipation was diagnosed if there were at least 2 of the following: abdominal pain, infrequent bowel movements, hard feces, fecal soiling, pain on defecation, and/or clinical evidence of excessively retained feces. Data on the presenting symptoms, signs, and ED treatment plan were collected on study enrollment and then in 2 standardized 10-minute telephone interviews at 48 hours and 7 days after the ED visit. At each follow-up, patient disposition was measured and dichotomized based on symptom resolution to "improved" vs "not improved." The presenting features and ED management were compared between groups using chi(2) analyses and t tests. RESULTS: Consent and full questionnaire completion was obtained in 102 children. The mean + SD age was 6.5 + 3.8 years; 47 (46%) were male. The predominant presenting symptom was abdominal pain (83 [81%]); the most frequent sign was palpable abdominal stool (67 [66%]). A high-fiber diet (75 [74%]) and mineral oil (48 [47%]) were prescribed most frequently. Enemas were given to 64 (63%) of the children. Improvement was found in 32 (31%) of the children at 48 hours and in the majority at 7 days (77 [75%]). Risk factors for poor symptom resolution at both 48 hours and 1 week included: female sex (odds ratio [OR] = 2.6; 95% confidence interval [CI] = 1.0 6.6); history of recurrent abdominal pain (OR = 2.8; 95% CI = 1.2 6.5); duration of primary presenting symptom longer than 2 days (OR = 2.4; 95% CI = 1.0-6.4); and history of medical visits for the same symptom (OR = 2.3; 95% CI = 1.0-5.3). There was no difference in outcome based upon ED treatment (enema vs oral or no therapy) (OR = 1.0; 95% CI = 0.4-2.3). CONCLUSIONS: Most children with constipation evaluated in the ED have acute symptoms and rapid improvement, regardless of presentation characteristics or ED management. In this study, 4 risk factors for poor outcome were found consistently at 48 hours and 7 days. This subgroup of children deserve closer clinical attention. Emergency department therapy did not influence short-term symptom resolution. Further studies are warranted to examine the effects of therapy for constipation in the ED setting. Arch Pediatr Adolesc Med. 2000;154:1204-1208. PMID- 11115304 TI - Primary care relationships in pediatric hospital clinics vs private offices. AB - OBJECTIVE: To explore residents' opinions regarding various aspects of continuity clinic in a hospital clinic vs private office, with emphasis on primary care relationships. DESIGN: Cross-sectional study. Residents (N = 47) who had spent time in both a clinic and an office setting were given a 49-question survey. Five point Likert scales were used to quantify the residents' sense of quality of various aspects of training. A set of 10 statements was included to assess the degree to which residents took primary care responsibility for a cohort of patients. Residents also chose the combination of setting(s) (office, clinic, or both) that they considered optimal for their training. SETTING: Large metropolitan area. The hospital clinic, based in a freestanding children's hospital, served a primarily indigent population. The private offices (n = 20) served the surrounding suburbs and primarily middle- and upper-income families. HYPOTHESIS: Residents would perceive their clinic-based experience as significantly better than their office-based experience with respect to establishing primary care relationships with their patients. RESULTS: Residents' ratings for the primary care responsibility index were significantly higher for the clinic than for the office (mean + SD, 4.1 + 0.8 vs 2.2 + 0. 9; P<.001). Residents found many aspects of the 2 settings comparable, including overall educational value and the preceptor as a positive role model. The office was rated higher for business and managed care aspects, care of school-age children, and experience with adolescents. The clinic was rated higher for care of infants and complex psychosocial and complex medical issues. Two thirds of residents chose the combined half day in office and half day in clinic as their preferred training model. CONCLUSION: The opportunity for residents to take advantage of the unique strengths of both clinic- and office-based training may significantly improve their overall residency experience. Arch Pediatr Adolesc Med. 2000;154:1209-1213. PMID- 11115305 TI - Treatment of sialorrhea with glycopyrrolate: A double-blind, dose-ranging study. AB - OBJECTIVE: To determine the safety and efficacy of glycopyrrolate in the treatment of developmentally disabled children with sialorrhea. DESIGN: Placebo controlled, double-blind, crossover dose-ranging study. SETTING: Outpatient facilities in 2 pediatric hospitals. PATIENTS: Thirty-nine children with both developmental disabilities and excessive and bothersome sialorrhea. MAIN OUTCOME MEASURES: Parent and investigator evaluation of change in sialorrhea and adverse effects. RESULTS: Glycopyrrolate in doses of 0.10 mg/kg per dose is effective at controlling sialorrhea. Even at low doses, 20% of children may exhibit adverse effects severe enough to require discontinuation. CONCLUSIONS: Glycopyrrolate is effective in the control of excessive sialorrhea in children with developmental disabilities. Approximately 20% of children given glycopyrrolate may experience substantial adverse effects, enough to require discontinuation of medication. Arch Pediatr Adolesc Med. 2000;154:1214-1218. PMID- 11115306 TI - Clinical comparability of ventolin formulated with hydrofluoroalkane or conventional chlorofluorocarbon propellants in children with asthma. AB - BACKGROUND: Aerosolized asthma medications with chlorofluorocarbon (CFC) propellants are being phased out because of environmental concerns about the ozone layer. Medications are being reformulated with non-ozone-depleting propellants. OBJECTIVE: To evaluate the clinical comparability of albuterol sulfate formulated in a new hydrofluoroalkane-134a (HFA) propellant (Ventolin HFA Inhalation Aerosol), and conventional CFC-containing albuterol (Ventolin Inhalation Aerosol) in children with asthma. DESIGN: Randomized, double-blind, placebo-controlled 2-week clinical trial with a 1- to 2-week run-in period. During the run-in, patients took Ventolin CFC as needed. Patients (n = 135) aged 4 to 11 years with asthma then were assigned randomly to treatment with Ventolin HFA, Ventolin CFC, or placebo administered 4 times daily via metered-dose inhaler for 2 weeks. All patients were allowed rescue albuterol use in matching propellant as needed for relief of breakthrough symptoms. The main outcome measure was the mean percentage of predicted peak expiratory flow (PEF) after the morning dose of study drug on day 1 and after 2 weeks as assessed by results of 6 hour serial tests. RESULTS: At day 1, the mean (+/- SE) percentage of predicted PEF increased postdose by 14% (+/- 1%) in the Ventolin HFA group and 13% (+/- 1%) in the Ventolin CFC group compared with 6% (+/- 2%) in the placebo group (P or =5 mg/dL), beyond 1 week. RESULTS: Twenty-one patients (41%) in the test group showed a bile leak, and a median of 1 site (range, 1-6 sites) was closed during the test. Postoperative bile leakage was observed in 3 patients (6%) in the test group and in 2 (4%) in the control group (P = .99). The odds ratio of the event was 1.53 (95% confidence interval, 0.25-9.44). The median postoperative hospital stay lasted 17 (range, 13-47) and 18 (range, 12-41) days for the test and control groups, respectively (P =.98). CONCLUSION: This randomized trial suggested no advantage in using a bile leakage test during hepatic resection. PMID- 11115341 TI - Invited critique PMID- 11115340 TI - Liver transplantation with renoportal anastomosis after distal splenorenal shunt. AB - BACKGROUND: The distal splenorenal shunt (DSRS) is designed to maintain hepatopetal portal vein flow while decompressing gastroesophageal varices. However, over time, as the underlying liver disease progresses, the DSRS loses its selectivity. The most common method of addressing this issue during orthotopic liver transplantation is shunt ligation with or without splenectomy. Dismantling the shunt increases the complexity of the transplantation, and splenectomy may increase the risk of infection. HYPOTHESIS: Anastomosis of the donor portal vein to the left renal vein without dismantling the shunt is an effective method of portal vein reconstruction for patients with a patent DSRS. DESIGN: Retrospective analysis. SETTING: University-based teaching hospital, Miami, Fla. PATIENTS: Five liver transplant recipients with patent DSRS who received an orthotopic liver transplant between September 1996 and August 1999. INTERVENTIONS: The donor portal vein was anastomosed end-to-end to the left renal vein during liver transplantation. MAIN OUTCOME MEASURES: Perioperatve morbidity, portal vein flow by Doppler study, patient survival, and graft survival. RESULTS: In all patients, the graft liver reperfused promptly via flow through the left renal vein with adequate decompression of the bowel. Normal portal venous flow was demonstrated by intraoperative and postoperative Doppler ultrasound studies. At the mean follow-up of 16 months, 4 patients were alive with well-functioning grafts. CONCLUSIONS: This novel technique has the advantage of decreasing the complexity of the procedure, without requiring splenectomy, while securing adequate portal perfusion. Additionally, it can be applied without modifications in patients with portal vein thrombosis. PMID- 11115342 TI - Improvement in healing with aggressive edema reduction after debridement of foot infection in persons with diabetes. AB - BACKGROUND: Infected foot wounds in patients with diabetes are the most common reason for diabetes-related hospital admission in the United States. Nonhealing foot wounds are the major precipitant of lower-extremity amputation in the diabetic population. HYPOTHESIS: The null hypothesis was that there would be no difference in proportion of healing with or without use of a foot-level mechanical compression device. DESIGN: Twelve-week, double-blind, randomized, controlled trial. SETTING: A university teaching hospital and related clinics. PATIENTS: One hundred fifteen patients with diabetes, 74% male, with foot infections requiring incision and debridement. INTERVENTION: All patients received either a functioning or placebo (nonfunctioning) foot compression device (Kinetic Concepts Inc, San Antonio, Tex). Patients and investigators were blinded to the functionality of the device. PRIMARY OUTCOME MEASURE: Proportion of wound healing in each group. RESULTS: There was a significantly higher proportion of healing in the active group than in the placebo group (39 [75%] of 52 patients vs 23 [51%] of 45; chi(2) = 6.0; P<.02; odds ratio, 2.9; 95% confidence interval, 1. 2-6.8). In the placebo group, there was no difference in proportion of healing between those identified as compliant (>/=50 hours of use per week) vs noncompliant (P =.10). In patients receiving active units, more patients in the compliant subgroup experienced wound healing (P<.03). When compared as a whole, there was a significant trend toward an increasing proportion of healing from the placebo-noncompliant to the placebo-compliant to the active-noncompliant to the active-compliant groups (chi(2)(trend) = 8.3; P<.005). CONCLUSIONS: Edema reduction achieved in this study by way of a pump and wrap system may increase the proportion of wound healing in patients after debridement of foot infections in patients with diabetes. Furthermore, the data suggest a potential association between increased compliance with use of the device and an increased trend toward wound healing. Arch Surg. 2000;135:1405-1409 PMID- 11115343 TI - Factor V leiden and morbid obesity in fatal postoperative pulmonary embolism. AB - HYPOTHESIS: Currently, the risk for postoperative acute pulmonary embolism (APE) is assessed clinically. We hypothesize that the expensive screening for the most common genetic thrombophilic clotting defect (factor V Leiden; R(506)Q) after exclusion of established clinical risk factors does not offer additional benefit to surgical patients. DESIGN: We reviewed protocols and histories from 8249 consecutive autopsies performed at the Mayo Clinic, Rochester, Minn. All patients who died of APE after routine surgery and who lacked any other clinical risk factors for APE were included and compared with matched controls. Genomic DNA was extracted from archival tissues and examined for R(506)Q by polymerase chain reaction amplification, restriction enzyme digestion, and direct sequencing. RESULTS: Acute pulmonary embolism was the immediate cause of death in 454 patients (5.5%). Of those, 32 (7.0%) had undergone routine surgery. These patients represent less than 0.07% of all case-adjusted surgical procedures in the same period. The rate of postoperative death from APE was higher after neurosurgical procedures (0.3%) than all other procedures (0.04%). Sixteen patients (50.0%) were morbidly obese. Only 1 patient was heterozygous and none were homozygous for R(506)Q. CONCLUSIONS: (1) Fatal APE is uncommon in surgical patients lacking clinically apparent risk factors for venous thromboembolism. (2) Neurosurgical patients are at increased risk for postoperative APE. (3) Morbid obesity is a major independent risk factor in cases of sudden death from APE postoperatively. (4) Routine preoperative screening for R(506)Q in the factor V gene does not appear to offer additional benefit in surgical patients without clinically recognizable thromboembolic risk factor(s). PMID- 11115344 TI - Protective covering of surgical wounds with honey impedes tumor implantation. AB - HYPOTHESIS: Tumor implantation (TI) development at the surgical wound following cancer surgery is still an unresolved concern. Trocar site recurrence, which is likely a form of TI, has become one of the most controversial topics and, with the widespread acceptance of laparoscopic surgery, has caused renewed interest in questions about TI. Honey has positive effects on wound healing. Physiological and chemical properties of honey might prevent TI when applied locally. DESIGN, INTERVENTIONS, AND MAIN OUTCOME MEASURES: Sixty BALB/c strain mice, divided into 2 groups, were wounded in the posterior neck area. Group 1 mice formed the control group, and group 2 mice had wounds coated with honey before and after tumor inoculation. All wounds were inoculated with transplantable Ehrlich ascites tumor. The presence of TI was confirmed in the wounded area by histopathological examination on the 10th day. RESULTS: Tumor implantation was achieved in all group 1 animals and verified by palpable mass and histopathological examination. In group 2 mice, although TI could not be detected macroscopically, it was revealed by pathological examination in 8 cases. Tumor implantation was less likely in group 2 mice (8 of 30 vs 30 of 30; P<.001). CONCLUSIONS: Tumor implantation was markedly decreased by the application of honey pre- and postoperatively. It is possible that the physiological and chemical properties of honey protected wounds against TI. Honey could be used as a wound barrier against TI during pneumoperitoneum in laparoscopic oncological surgery and in other fields of oncological surgery. PMID- 11115345 TI - Invited critique PMID- 11115346 TI - Endothelin-1 levels in severe burn injuries. AB - HYPOTHESIS: Most investigators have reported high levels of endothelin (ET)-1 in patients with thermal injury. We attempted to examine the hypothesis that ET-1 levels increase in patients with severe burn injury. PATIENTS AND METHODS: Plasma from 28 adult subjects, 14 of whom had thermal injuries with a median (range) percentage of total burn surface area of 22% (20%-76%), was assessed for ET-1 and tumor necrosis factor (TNF) alpha. Samples from closely age-matched patients were obtained on admission (day 1) and 24 hours postinjury (day 2). Samples were obtained before blood transfusion or surgical treatment occurred. Enzyme immunoassay techniques suitable for the measurements of the cytokines were used. RESULTS: Median (range) of TNF-alpha was higher in patients (day 1, 10.0 ng/L [1.2-35.0 ng/L]; day 2, 12.0 ng/L [0.4-39.0 ng/L]) than controls (0. 8 ng/L [0.3 3.2 ng/L]) (P<.005) while ET-1 levels remained significantly unchanged in patients (mean [SD], day 1, 183.0 [42.2] ng/L; day 2, 204.7 [41.7] ng/L) compared with controls (170.0 [59.8] ng/L) (P>.05). CONCLUSIONS: We observed no significantly raised levels of ET-1 in patients with thermal injury within 24 hours after burn injury. We found no significant correlation between the plasma levels of TNF-alpha and ET-1. Endothelin-1 levels did not seem to reflect severity of illness. The actual evaluation of ET-1 release in patients with thermal injury could enhance the pathophysiological study of human thermal injury. PMID- 11115347 TI - DNA flow cytometry does not predict 5- or 10-year recurrence rates for T1-2 node negative breast cancer. AB - BACKGROUND: A small proportion of T1 or T2 node-negative breast cancer tumors will recur in patients by 5 years, and more by 10 years. Results of recent studies have suggested improvement in overall survival with administration of adjuvant chemotherapy to all patients. More sensitive and specific methods are needed to identify patients at highest risk for recurrence who might benefit most from adjuvant therapy, saving others from unnecessary treatment. Some investigators have suggested DNA flow cytometry as a method to discriminate patients at greatest risk for recurrence. HYPOTHESIS: DNA flow cytometry has predictive value for breast cancer recurrence in node-negative patients. METHODS: The cancer registry of a medium-sized university-affiliated hospital was used to identify patients with T1-2 N0 M0 breast cancer treated with a uniform surgical approach and no adjuvant therapy who had completed at least 5 years of follow-up or had recurrence. Flow cytometric analysis was performed on paraffin-embedded specimens. RESULTS: Of 115 patients, 92 (80%) had disease-free survival without recurrence and 23 (20%) had recurrence. Comparison of diploid and nondiploid tumors for likelihood of recurrence revealed no association (P = .79). Furthermore, the DNA index and S-phase fraction were not significantly different between recurrent and nonrecurrent groups. CONCLUSIONS: The likelihood of recurrence of small node-negative breast cancers after mastectomy cannot be accurately predicted on the basis of DNA flow cytometric analysis. Traditional methods for determining risks-such as nuclear and histological grade, lymph node status, and tumor size-seem to be more useful. Sentinel lymph node biopsy techniques may increase the detection of micrometastases. PMID- 11115348 TI - Surgical anatomy of the spinal accessory nerve and the trapezius branches of the cervical plexus. AB - BACKGROUND: A thorough understanding of the topographical anatomy of the spinal accessory nerve and the cervical plexus branches is a basic prerequisite for positive results when operating on the neck. OBJECTIVE: To give an exact description of the topographical and surgical anatomy of the spinal accessory nerve (SAN) and the trapezius branches of the cervical plexus. DESIGN: Anatomic analysis of the SAN and the trapezius branches of the cervical plexus. SETTING: The topographical anatomy of the SAN and the cervical plexus branches were studied in the anterior and posterior triangles of the necks of 46 perfusion fixed human cadavers of both sexes, which ranged in age from 55 to 97 years (mean age, 83 years). RESULTS: The SAN can be identified on the posterior border of the sternocleidomastoid (SCM) muscle, 8.2 + 1.01 cm cranial to the clavicle. In 37% of cases, the SAN enters the posterior triangle of the neck dorsal to the SCM muscle, where it passes through the muscle in 63% of these cases. In the anterior triangle of the neck, the SAN crosses the internal jugular vein ventrally in 56% of the cases and dorsally in 44%. Regarding the cervical plexus, 1 trapezius branch could be found in 9% of the specimens, 2 in 61%, and 3 in 30%. None of the branches merged with the SAN medial to the anterior border of the trapezius muscle. In most cases, a tiny additional branch could be found arising from the SAN about 2 cm medial to the trapezius muscle. This branch enters the descendant part of the muscle approximately 2 to 3 cm cranial to the main nerve. CONCLUSIONS: Surprisingly, available data on topographical as well as surgical anatomy of the SAN and the trapezius branches of the cervical plexus are confusing and often wrong. The descriptions given herein can help to minimize the risk of injuring the SAN during neck surgery and preserve the additional innervation of the trapezius muscle granted by the rami trapezii of the cervical plexus. PMID- 11115349 TI - Signaling mechanisms of altered cellular responses in trauma, burn, and sepsis: role of Ca2+. AB - Alterations in cellular responses in various organ systems contribute to trauma-, burn-, and sepsis-related multiple organ dysfunction syndrome. Such alterations in muscle contractile, hepatic metabolic, and neutrophil and T-cell inflammatory immune responses have been shown to result from cell-signaling modulations and/or impairments in the respective cell types. Altered Ca(2+) signaling would seem to play an important role in the myocardial and vascular smooth muscle contractile dysfunction in the injury conditions; Ca(2+)-linked signaling derangement also plays a crucial role in sepsis-induced altered skeletal muscle protein catabolism and resistance to insulin-mediated glucose use. The injury-related increased hepatic gluconeogenesis and acute-phase protein response could also be caused by a pathophysiologic up-regulation of hepatocyte Ca(2+)-signal generation. The increased oxidant production by neutrophil, a potentially detrimental inflammatory response in early stages after burn or septic injuries, seems to result from an up-regulation of both the Ca(2+)-dependent as well as Ca(2+) independent signaling pathways. The injury conditions would seem to cause an inappropriate up-regulation of Ca(2+)-signal generation in the skeletal myocyte, hepatocyte, and neutrophil, while they lead to a down-regulation of Ca(2+) signaling in T cells. The crucial signaling derangement that causes T-cell proliferation suppression seems to be a decrease in the activation of protein tyrosine kinases, which subsequently down-regulates Ca(2+) signaling. The delineation of cell-signaling derangements in trauma, burn, or sepsis conditions can lead to development of therapeutic interventions against the disturbed cellular responses in the vital organ systems. PMID- 11115350 TI - Subcutaneous perfusion and oxygen during acute severe isovolemic hemodilution in healthy volunteers. AB - HYPOTHESIS: Acute severe isovolemic anemia (to a hemoglobin [Hb] concentration of 50 g/L) does not decrease subcutaneous wound tissue oxygen tension (PsqO(2)). SETTING: University hospital operating room and inpatient general clinical research center ward. SUBJECTS: Twenty-five healthy, paid volunteers. METHODS: Subcutaneous oxygen tension and subcutaneous temperature (Tsq) were measured continuously during isovolemic hemodilution to an Hb level of 50 g/L. In 14 volunteers (initially well-perfused), "normal" perfusion (Tsq >34.4 degrees C) was achieved by hydration and systemic warming prior to starting isovolemic hemodilution, while in 11 volunteers (perfusion not controlled [PNC]), no attempt was made to control perfusion prior to hemodilution. MAIN OUTCOME MEASURES: Measurements of PsqO(2), Tsq, and relative subcutaneous blood flow (flow index). RESULTS: While PsqO(2), Tsq, and flow index were significantly lower in PNC vs well-perfused subjects at baseline, there was no significant difference between them at the Hb of 50 g/L (nadir). Subcutaneous PO(2) did not decrease significantly in either group. Arterial PO(2) was not different between the groups, and did not change significantly over time; Tsq and flow index increased significantly from baseline to nadir Hb in both groups. CONCLUSIONS: The level of PsqO(2) was maintained at baseline levels during hemodilution to Hb 50 g/L in healthy volunteers, whether they were initially well-perfused or mildly underperfused peripherally. Given the significant increase in Tsq and flow index, this resulted from a compensatory increase in subcutaneous blood flow sufficient to maintain oxygen delivery. Wound healing depends to a large extent on tissue oxygen delivery, and these data suggest that even severe anemia by itself would not be sufficient to impair wound healing. Thus, transfusion of autologous packed red blood cells solely to improve healing in surgical patients with no other indication for transfusion is not supported by these results. PMID- 11115351 TI - K-ras point mutation in the nerve plexuses around the superior mesenteric artery in resectable adenocarcinoma of the pancreatic head: distribution pattern and related factors. AB - BACKGROUND: Adenocarcinoma of the pancreas is likely to spread into the nerve plexuses around the superior mesenteric artery (SMA) at a microscopic level. Since there has been no detailed report on how minute cancer invasion is distributed among the peri-SMA plexuses or which cases are more vulnerable to such an event, it has long been controversial how to treat this area when resecting the pancreatic head cancer. HYPOTHESIS: The K-ras mutation assay is more sensitive than the conventional histologic diagnosis in detecting minute cancer invasion around the SMA. DESIGN: Prospective consecutive series. SETTING: Cancer center hospital. PATIENTS AND METHODS: The entire circle of the peri-SMA tissues was obtained from 24 patients who had received an extended pancreatectomy for adenocarcinoma of the pancreatic head. They were divided into right and left hemicircular samples (48 samples), and each sample was used for both histologic and genetic diagnoses. Since all patients' primary tumors were positive for point mutation at codon 12 of the K-ras gene, the presence or absence of the mutation was determined for the peri-SMA plexuses using the mutant allele specific amplification method. RESULTS: Compared with results of the histologic examination, the K-ras mutation assay was more sensitive in detecting positive findings in the peri-SMA plexuses (12 samples from 9 patients). According to the distribution of the K-ras mutation into the right- and left-half samples, 24 patients were classified into the following 4 patterns (right/left): negative/negative in 15 patients; positive/negative in 6 patients; positive/positive in 3 patients; and negative/positive in 0 patients. In 3 patients who showed a positive/positive pattern in the genetic diagnosis, their right-half samples included more cancer cells that were detectable by routine microscopy. There was no relation between K-ras mutation and lymphatic invasion, while K-ras mutation was particularly related with the invasion of portal vein (P =.04) and posterior peripancreatic tissues (P =.002). All 3 patients with K-ras mutation in bilateral plexuses were classified by the TNM staging system as T4 using Union Internationale Contre le Cancer classification. CONCLUSIONS: The K ras mutation (at codon 12) assay indicated a simple and regular pattern of cancer extension into the nerve plexuses around the SMA from adenocarcinoma of the pancreatic head: (1) The left half of the plexus was unlikely to be involved by cancer in cases in which the right half was intact. (2) Cancer extension into the peri-SMA plexuses occurred after the posterior confine of the pancreas had been involved by direct invasion from the primary pancreatic tumor. (3) The left half was not involved in cancerous tumors classified as T1 to T3 but was occasionally involved in those classified as T4 tumors. These data seem to provide a useful indicator of some additional treatments (resection, irradiation, etc) for the peri-SMA region when a locally advanced pancreatic head cancer is treated with a curative intent. PMID- 11115352 TI - Clinical significance of hepatic resection in hepatocellular carcinoma: analysis by disease-free survival curves. AB - HYPOTHESIS: The clinical significance of hepatectomy for hepatocellular carcinoma (HCC) is still controversial because of frequent intrahepatic recurrence, which results from either recurrence due to residual intrahepatic metastasis (Rim) or recurrence due to metachronous, multicentric liver carcinogenesis (Rmc). DESIGN: Retrospective review. Disease-free survival curves were obtained by the Kaplan Meier method and the rates of Rim and Rmc were analyzed using 2 regression lines, based on the evidence that Rmc occurs at a constant rate throughout follow-up, whereas Rim occurs only in the early postoperative period. SETTING: University hospital. PATIENTS: From 1980 to 1996, 241 patients with HCC who underwent curative hepatic resection. MAIN OUTCOME MEASURE: Intrahepatic recurrence. RESULTS: Disease-free survival curves for all patients in the early (within 2 years) and late (4 years after surgery) follow-up were approximated by 2 regression lines, which represent both Rim and Rmc (Y(1) = -3.4X + 48) and only Rmc (Y(2) = -23.1X + 98). Using this approximation, the annual incidence of Rim within 2 years (a(1)-a(2)) was calculated as 19.7% and that of Rmc (a(2)) was 3.4%. The ratio of Rim in tumor recurrence (b(1)-b(2)) was 50%, and that of Rmc (b(1)) was 48%. The ratios of Rmc in patients with stages I and II HCC were 60% and 64%, respectively. In contrast, the values could not be calculated in patients with stages III and IVA because all but 2 patients showed recurrence within 4 years after surgery. CONCLUSION: Tumor recurrence is estimated to result from metachronous liver carcinogenesis in 48% of hepatectomized patients with HCC. PMID- 11115353 TI - Unilateral surgery for primary hyperparathyroidism on the basis of technetium Tc 99m sestamibi and iodine 123 subtraction scanning. AB - HYPOTHESIS: Parathyroid scanning, based on simultaneous recording of technetium Tc 99m sestamibi and iodine 123 images, is able to identify patients with multiple parathyroid gland disease and is a safe imaging technique for unilateral parathyroid surgery. DESIGN: Scintigraphic criteria of eligibility for unilateral surgery were prospectively tested against findings of conventional bilateral surgery. SETTING: Patients referred to an endocrine surgeon in a university hospital. PATIENTS: Seventy consecutive patients with primary hyperparathyroidism had dual-isotope scanning before conventional surgery. Forty-one patients had scan findings compatible with unilateral surgery, with a single focus of high intensity seen on the anterior and lateral views. The remaining 29 patients had 1 or more criteria of ineligibility: (1) scan findings pointing to multiple gland disease, (2) no well-identified focus, (3) contralateral thyroid nodule requiring surgical management, or (4) family history of hyperparathyroidism or multiple endocrine disease. MAIN OUTCOME MEASURES: Number of enlarged parathyroid glands at surgical inspection and calcemia follow-up. RESULTS: None of the 41 patients, with a single well-defined focus on the scan image, showed evidence of multiple parathyroid involvement. Each parathyroid adenoma was resected from the precise site predicted by the subtraction scan. Nine patients (13%) had surgical findings of multiple parathyroid gland disease. All 9 were ineligible based on preoperative image findings. CONCLUSIONS: Unilateral surgery can be safely offered to 60% of patients with primary hyperparathyroidism, on the basis of simultaneous (99m)Tc-sestamibi and (123)I scanning. This may reduce the length of the operation, anesthesia requirements, and hospital stay, and the risks of hypoparathyroidism and injury to the recurrent laryngeal nerve. PMID- 11115354 TI - The association of HER-2/neu amplification with breast cancer recurrence. AB - HYPOTHESIS: Amplification of the HER-2/neu oncogene in 25% of breast cancers is associated with a shortened disease-free survival. DESIGN: Retrospective analysis of a patient population referred to a tertiary care facility for HER-2/neu testing. The mean follow-up was 56 months. SETTING: Large, urban, tertiary care hospital. PATIENTS: From 1995 to 1999, a consecutive sample of 190 patients with breast cancer had tissue samples tested for overexpression of the cell surface oncoprotein by immunostaining (IM) or amplification of the HER-2/neu oncogene by fluorescence in situ hybridization or both. Forty-nine subjects were excluded because they had tissue samples tested at our institution but received their treatment elsewhere. All patients tested for HER-2/neu after diagnosis with breast cancer in 1999 (n = 47) were excluded from analysis because of short follow-up time. One patient was excluded who had in situ ductal carcinoma. The remaining 93 patients were analyzed. RESULTS: Of 93 patients, 40 (43%) had gene amplification. Overall, patients with oncogene amplification had a shorter median disease-free interval (22 months) compared with controls (40 months) (P =.003). Analysis by the Cox regression model showed that the HER-2/neu status remained significantly associated with time to relapse even after adjusting for age and tumor grade (P =.002; adjusted relative risk, 2.4; 95% confidence interval, 1.4 4.4). No association was found between gene amplification and tumor grade (P =.98), estrogen/progesterone receptor status (P = .29 and P = .43, respectively), or lymph node status (P = .98). Seventy-two patients (77%) eventually had disease recurrence, with 18 (25%) of these recurring locally. CONCLUSIONS: The HER-2/neu oncogene is an independent prognostic indicator of a subset of breast cancers that are at high risk of early recurrence, regardless of tumor grade, estrogen/progesterone receptor status, and lymph node status. Patients amplifying the HER-2/neu oncogene have a shorter disease-free survival than patients without the oncogene. PMID- 11115355 TI - Is the p53 gene mutation of prognostic value in hepatocellular carcinoma? PMID- 11115356 TI - Surgical reminiscences: the qualities of a successful surgeon. PMID- 11115357 TI - Moments in surgical history: William Stewart Halsted. PMID- 11115358 TI - Role of RANTES in experimental cardiac allograft rejection. AB - The host response to alloantigen results in upregulation of Class II MHC antigens and associated cytokine production (especially IL-2 and interferon-gamma (IFN gamma)) as well as lymphocytic infiltration and cellular activation which leads to graft damage/destruction. RANTES (Regulated upon Activation, Normal T-cell Expressed and presumably Secreted) is a member of the beta subfamily (CC) of chemokines and has been shown to function as a lymphocyte chemoattractant. We now describe the requirement for RANTES in cardiac heterotopic allograft (brown Norway into Lewis rats) rejection in rats. By Northern blot analysis, mRNA could be detected in allografts at 6 and 8 days post-transplantation but not in isogenic (Lewis) grafts. RANTES protein could be demonstrated by Western blot analysis in homogenates from allografts at day 8 but not at day 0 and could not be identified in isogenic cardiac transplants. By immunostaining, RANTES protein was present in mononuclear cells of allografts at day 6 but was absent in the isogenic transplants. When rats were treated with anti-RANTES serum, there was a significant delay in rejection time (cessation of beating) of the allografts. These data demonstrate that expression of RANTES in rat cardiac allografts is linked to the rejection phenomenon. PMID- 11115359 TI - A newly characterized human endometrial adenocarcinoma cell line (CAC-1) differentiates in response to retinoic acid treatment. AB - A new cell line of poorly differentiated human endometrial adenocarcinoma cells termed "CAC-1" cells has been established. These cells are epithelial, as indicated by positive cytokeratin and negative vimentin staining. They are rounded and possess a high nuclear-to-cytoplasmic ratio, desmosomes, surface microvilli, intercelular lumens, and pleomorphic nuclei containing multiple nucleoli. These cells have been in long-term culture for 2 years. Our previous studies demonstrated that moderately differentiated (RL95-2) cells differentiated in response to retinoic acid treatment, illustrated by their reorganization of actin filaments and cell enlargement (Carter et al., 1996; Anticancer Res. 16, 17 24). CAC-1 cells exhibited a similar response because they also organized actin filaments and enlarged in response to retinoic acid treatment. Concurrently, retinoic acid treatment caused a 40% decrease in cell detachment in an in vitro detachment assay compared to controls. A slight lag in cell growth was observed when CAC-1 cells were treated with 1 microM 13-cis or all-trans retinoic acid during a 12-day growth curve. In addition, we examined the effects of retinoic acid on protein kinase C-alpha (PKC-alpha) and myristoylated alanine-rich C kinase substrate (MARCKS). Treatment with retinoic acid caused cytoplasmic PKC alpha to increase concomitant with a decrease in PKC-alpha in the membrane. In contrast, MARCKS increased in the membrane in response to retinoic acid treatment. These data indicate that retinoid treatment causes inactivation of PKC alpha, allowing MARCKS to relocalize to the membrane, where it can cross-link actin filaments. CAC-1 cells represent an ideal model for investigating the effects of retinoids on differentiation induction concomitant with actin reorganization. PMID- 11115360 TI - Immunohistochemical analysis of CCR2, CCR3, CCR5, and CXCR4 in the human brain: potential mechanisms for HIV dementia. AB - The CXC chemokine receptor CXCR4 was the first molecule identified as a coreceptor working in conjunction with CD4 to mediate cellular entry for the human immunodeficiency virus (HIV-1). Since that original discovery, 11 other seven-mtransmembrane domain molecules, many of which are chemokine receptors, have been shown to facilitate HIV entry into cells. These include CCR5, CCR3, CCR2, CCR1, CCR8, CX3CR1, STRL33 (BONZO), GPR15 (BOB), GPR1, US28, and APJ. In studies done by this and other labs, CCR3, CCR5, and CXCR4 have been identified in CNS microglia and several laboratories, including ours, have shown that CXCR4 is expressed in neurons. Neuronal expression of CCR2, CCR3, and CCR5 has been less consistent. We performed a semiquantitative immunohistochemical analysis of the expression of CCR2, CCR3, CCR5, and CXCR4 in 23 regions of the brain and in two sections of the spinal cord. Hippocampal neurons were positive for CCR2, CCR3, and CXCR4, but not for CCR5. In other regions of the brain, neurons, and glial cells reacted with anti-CCR2, anti-CCR3, and anti-CXCR4 antibodies, whereas only glial cells (primarily microglia) were positive for CCR5. The areas of highest expression, however, seem to be subcortical regions and the limbic system. The limbic system plays a key role in memory, and the presence of CXCR4 which can bind the viral envelope protein gp120-min a subset of neurons from this system may play a role in the development of HIV-related dementia. PMID- 11115361 TI - Dexamethasone enhances mallory body formation in drug-primed mouse liver. AB - In a clinical study in which patients with alcoholic hepatitis were treated with prednisone for 1 month, posttreatment liver biopsies showed diminished inflammation, but Mallory bodies were not diminished. This suggested that steroid treatment may reduce inflammation by inhibiting NFkappaB activation. Sparing of Mallory bodies suggests that NFkappaB activation may not be involved mechanistically in Mallory body formation. To test this idea, we induced Mallory body formation in drug-primed mice with or without dexamethasone treatment. As predicted, dexamethasone decreased NFkappaB activation; however, Mallory body formation was increased. Surprisingly, TNFalpha and iNOS, which normally increase as a result of NFkappaB activation, were upregulated by the dexamethasone treatment. It was concluded that NFkappaB activation is not involved in Mallory body formation. Despite this, induced increases in TNFalpha, iNOS, c-jun/API and c-myc expression indicate that oxidative stress is likely involved in Mallory body formation. PMID- 11115362 TI - Fate of immune complexes, glomerulonephritis, and cell-mediated vasculitis in lupus-prone MRL/Mp lpr/lpr mice. AB - Immune complex formation was induced by the injection of (125)I-BSA into female MRL/Mp lpr/lpr mice, which develop spontaneous systemic lupus erythematosus (SLE) like disease, and MRL/Mp +/+ mice, which do not. At designated intervals following the injection of 10 mg of (125)I-bovine serum albumin (BSA), the nonlupus mice developed sparse, small electron-dense deposits in mesangial areas and subepithelial immune deposits that underwent partial resolution. By contrast, glomeruli of the SLE-prone mouse kidneys revealed proliferation of mesangial cells and some increase in mesangial matrix material. Numerous subepithelial and mesangial electron-dense deposits were present. Some subendothelial and intramembranous deposits were also demonstrated. Capillary lumens contained massive electron-dense deposits. The resolving subepithelial deposits observed were fewer than half the number found in kidneys of the non-SLE mice. Whole body counts were also recorded daily following the injection of (125)I-BSA. Whereas, both lupus-prone and non-SLE control mice eliminated (125)I-BSA at equivalent rates through day 12 postinoculation, those with SLE-like disease showed a decreased (125)I-BSA elimination rate between days 6 and 12. Results suggest an impairment in the ability of SLE-prone mice to resolve immune complexes, whether they are nuclear-antinuclear or from an exogenous source, i.e., BSA-anti-BSA, compared to controls in this experimental model of the superimposition of exogenous immune complex formation on systemic lupus erythematosus-like disease. PMID- 11115363 TI - Ethanol-mediated CYP1A1/2 induction in rat skeletal muscle tissue. AB - The causes of non-trauma-mediated rhabdomyolysis are not well understood. It has been speculated that ethanol-associated rhabdomyolysis may be attributed to ethanol induction of skeletal muscle cytochrome P450(s), causing drugs such as acetaminophen or cocaine to be metabolized to myotoxic compounds. To examine this possibility, the hypothesis that feeding ethanol induces cytochrome P450 in skeletal muscle was tested. To this end, rats were fed an ethanol-containing diet and skeletal muscle tissue was assessed for induction of CYP2E1 and CYP1A1/2 by immunohistochemical procedures; liver was examined as a positive control tissue. Enzymatic assays and Western blot analyses were also performed on these tissues. In one feeding system, ethanol-containing diets induced CYP1A1/2 in soleus, plantaris, and diaphragm muscles, with immunohistochemical staining predominantly localized to capillaries surrounding myofibers. Antibodies to CYP2E1 did not react with skeletal muscle tissue from animals receiving a control or ethanol containing diet. However, neither skeletal muscle CYP1A1/2 nor CYP2E1 was induced when ethanol diets were administered by a different feeding system. Ethanol consumption can induce some cytochrome P450 isoforms in skeletal muscle tissue; however, the mechanism of CYP induction is apparently complex and appears to involve factors in addition to ethanol, per se. PMID- 11115364 TI - Development of a condemned mucosa syndrome and pathogenesis of human papillomavirus-associated upper aerodigestive tract and uterine cervical tumors. AB - High-risk human papillomaviruses (HPVs) have been shown to be involved in the pathogenesis of many squamous carcinomas, particularly those of the uterine cervix. A number of random studies have also reported association of high-risk HPV subtypes with cancers of the oral cavity, larynx, hypopharynx, and esophagus. The roles of other molecular factors involved during HPV infection in these tumors still remain unclear. Recent findings from our laboratories have suggested possible mechanisms associated with HPV-mediated carcinogenesis. Both p53 mutation-dependent and mutation-independent pathways may be associated with HPV mediated carcinogenesis, the former mainly in upper aerodigestive tract tumors (UADT) and the latter in cervical tumors. In cervical tumors, inactivation of the p53 tumor suppressor protein by the E6 gene product of high-risk HPVs and mutation of the p53 gene in UADT is associated with alterations in the apoptotic regulatory bcl-2 and bax genes, leading to downregulation of programmed cell death (PCD) and increased cell proliferation. HPV infection is also associated with increased tissue angiogenesis and activation of telomerase. Altered kinetics of telomere fragments is evident in HPV-infected tissue. We therefore believe that the combined manifestations of all these factors may contribute to development of a "condemned mucosa syndrome" facilitating development UADT and cervical cancers. A distinct step in the pathogenesis of both types of tumors may only be in the mode of p53 inactivation, whereas all other events appear to be strongly correlated to the presence of HPV. The development and validation of such a molecular model has significant clinical priority. It can be used to identify target populations or individuals for intervention, to monitor effects of intervention, and to determine which individuals or groups are at increased risk of developing cancer. PMID- 11115366 TI - Editorial. PMID- 11115365 TI - Memory of past exposure to the chemokine IL-8 inhibits the contraction of fibroblast-populated collagen lattices. AB - The inflammatory cytokine makeup of healing wounds helps delineate the phases of the repair process. As an example, during the lag phase (inflammatory phase) of repair, elevated levels of IL-8, a chemokine, participate in the activation and chemotaxis of neutrophils. During the normal proliferative and remodeling phases of repair, IL-8 levels are minimal. Healing burn wounds often have small, open, slow-to-heal areas, which have been shown to contain elevated levels of IL-8. Does a limited exposure of IL-8 to fibroblasts in vitro at the concentrations measured in these unhealed sites cause altered cell behavior? To study this possibility the fibroblast-populated collagen lattice (FPCL) model, an in vitro model of wound contraction, was used to investigate fibroblast response to IL-8. As previously reported, the chronic exposure of fibroblasts to IL-8 at 30 ng/ml within FPCLs significantly inhibited FPCL contraction. Fibroblasts in monolayer culture were incubated with IL-8 for 3 days. In the absence of IL-8, FPCLs were manufactured with these preexposed cells and it was found that FPCL contraction was inhibited. Fibroblasts retained their reduced capacity of reorganizing collagen fibrils when previously exposed to IL-8. Treating fibroblasts in monolayer with IL-8 for only 1 h prior to their incorporation into collagen lattices caused inhibition of FPCL contraction. The speculation is that in vivo open areas in reepithelialized healed burns are the consequence of the local population of fibroblasts having been exposed to elevated levels of IL-8. Such an earlier exposure triggers a memory in this population of fibroblasts that alters their capacity to lay down an extracellular matrix that optimizes the migration of epidermal cells. PMID- 11115367 TI - The comeback of mitochondria to calcium signalling. AB - With this overview of the role of mitochondria in the realm of calcium signalling we have tried to provide a chronological perspective, from the very early days to the present. We have briefly sketched a timeline of the research on calcium and mitochondria during the course of the century. Particular attention is paid to recent developments which have contributed to a renewed interest in calcium handling by this organelle. PMID- 11115368 TI - Mitochondrial calcium transport: mechanisms and functions. AB - Ca(2+)transport across the mitochondrial inner membrane is facilitated by transporters having four distinct sets of characteristics as well as through the Ca(2+)-induced mitochondrial permeability transition pore (PTP). There are two modes of inward transport, referred to as the Ca(2+)uniporter and the rapid mode or RaM. There are also two distinct mechanisms mediating outward transport, which are not associated with the PTP, referred to as the Na(+)-dependent and the Na(+) independent Ca(2+)efflux mechanisms. Several important functions have been proposed for these mechanisms, including control of the metabolic rate for cellular energy (ATP) production, modulation of the amplitude and shape of cytosolic Ca(2+)transients, and induction of apoptosis through release of cytochrome c from the mitochondrial inter membrane space into the cytosolic space. The goals of this review are to survey the literature describing the characteristics of the mechanisms of mitochondrial Ca(2+)transport and their proposed physiological functions, emphasizing the more recent contributions, and to consider how the observed characteristics of the mitochondrial Ca(2+)transport mechanisms affect our understanding of their functions. PMID- 11115369 TI - Mitochondria in myelinating cells: calcium signaling in oligodendrocyte precursor cells. PMID- 11115370 TI - Mitochondria as regulators of stimulus-evoked calcium signals in neurons. AB - An important challenge in the study of Ca2+ signalling is to understand the dynamics of intracellular Ca2+ levels during and after physiological stimulation. While extensive information is available regarding the structural and biophysical properties of Ca2+ channels, pumps and exchangers that control cellular Ca2+ movements, little is known about the quantitative properties of the transporters that are expressed together in intact cells or about the way they operate as a system to orchestrate stimulus-induced Ca2+ signals. This lack of information is particularly striking given that many qualitative properties of Ca2+ signals (e.g. whether the Ca2+ concentration within a particular organelle rises or falls during stimulation) depend critically on quantitative properties of the underlying Ca2+ transporters (e.g. the rates of Ca2+ uptake and release by the organelle). This monograph describes the in situ characterization of Ca2+ transport pathways in sympathetic neurons, showing how mitochondrial Ca2+ uptake and release systems define the direction and rate of net Ca2+ transport by this organelle, and how the interplay between mitochondrial Ca2+ transport and Ca+2 transport across the plasma membrane contribute to depolarization-evoked Ca2+ signals in intact cells. PMID- 11115371 TI - Pharmacological investigation of mitochondrial ca(2+) transport in central neurons: studies with CGP-37157, an inhibitor of the mitochondrial Na(+)-Ca(2+) exchanger. AB - Mitochondria buffer large changes in [Ca(2+)](i)following an excitotoxic glutamate stimulus. Mitochondrial sequestration of [Ca(2+)](i)can beneficially stimulate oxidative metabolism and ATP production. However, Ca(2+)overload may have deleterious effects on mitochondrial function and cell survival, particularly Ca(2+)-dependent production of reactive oxygen species (ROS) by the mitochondria. We recently demonstrated that the mitochondrial Na(+) Ca(2+)exchanger in neurons is selectively inhibited by CGP-37157, a benzothiazepine analogue of diltiazem. In the present series of experiments we investigated the effects of CGP-37157 on mitochondrial functions regulated by Ca(2+). Our data showed that 25 microM CGP-37157 quenches DCF fluorescence similar to 100 microM glutamate and this effect was enhanced when the two stimuli were applied together. CGP-37157 did not increase ROS generation and did not alter glutamate or 3mM hydrogen-peroxide-induced increases in ROS as measured by DHE fluorescence. CGP-37157 induces a slight decrease in intracellular pH, much less than that of glutamate. In addition, CGP-37157 does not enhance intracellular acidification induced by glutamate. Although it is possible that CGP-37157 can enhance mitochondrial respiration both by blocking Ca(2+)cycling and by elevating intramitochondrial Ca(2+), we did not observe any changes in ATP levels or toxicity either in the presence or absence of glutamate. Finally, mitochondrial Ca(2+)uptake during an excitotoxic glutamate stimulus was only slightly enhanced by inhibition of mitochondrial Ca(2+)efflux. Thus, although CGP 37157 alters mitochondrial Ca(2+)efflux in neurons, the inhibition of Na(+) Ca(2+)exchange does not profoundly alter glutamate-mediated changes in mitochondrial function or mitochondrial Ca(2+)content. PMID- 11115372 TI - Age-related structural and functional changes of brain mitochondria. AB - Normal ageing is associated with a gradual decline in the capacity of various cell types, including neurones, to respond to metabolic stress and return to the resting state. An important factor in the decrease of this 'homeostatic reserve' is the gradual, age-dependent impairment of mitochondrial function. In this article we review some of the major structural and functional changes in mitochondria associated with ageing. Apart from the increased mutations in mitochondrial DNA and the evidence for increased oxidative stress with ageing, we also discuss, in some detail, the importance of the mitochondrial membrane structure and composition (in particular lipid composition) for mitochondrial function in general and during ageing. Although some of the neurodegenerative diseases are also associated with some degree of mitochondrial dysfunction, it is not yet clear if these changes are due to the underlining process of normal, physiological ageing or due to the specific pathophysiologic agents responsible for the neurodegenerative processes. Furthermore, we are proposing that there are important differences between normal ageing and neurodegeneration. PMID- 11115373 TI - Mitochondria and Ca(2+)in cell physiology and pathophysiology. AB - There is now a consensus that mitochondria take up and accumulate Ca(2+)during physiological [Ca(2+)](c)signalling. This contribution will consider some of the functional consequences of mitochondrial Ca(2+)uptake for cell physiology and pathophysiology. The ability to remove Ca(2+)from local cytosol enables mitochondria to regulate the [Ca(2+)] in microdomains close to IP3-sensitive Ca(2+)-release channels. The [Ca(2+)] sensitivity of these channels means that, by regulating local [Ca(2+)](c), mitochondrial Ca(2+)uptake modulates the rate and extent of propagation of [Ca(2+)](c)waves in a variety of cell types. The coincidence of mitochondrial Ca(2+)uptake with oxidative stress may open the mitochondrial permeability transition pore (mPTP). This is a catastrophic event for the cell that will initiate pathways to cell death either by necrotic or apoptotic pathways. A model is presented in which illumination of an intramitochondrial fluorophore is used to generate oxygen radical species within mitochondria. This causes mitochondrial Ca(2+)loading from SR and triggers mPTP opening. In cardiomyocytes, mPTP opening leads to ATP consumption by the mitochondrial ATPase and so results in ATP depletion, rigor and necrotic cell death. In central mammalian neurons exposed to glutamate, a cellular Ca(2+)overload coincident with NO production also causes loss of mitochondrial potential and cell death, but mPTP involvement has proven more difficult to demonstrate unequivocally. PMID- 11115374 TI - Control of apoptosis by IP(3) and ryanodine receptor driven calcium signals. AB - Intracellular calcium signals mediated by IP(3)and ryanodine receptors (IP(3)R/RyR) play a central role in cell survival, but emerging evidence suggests that IP(3)R/RyR are also important in apoptotic cell death. Switch from the life program to the death program may involve coincident detection of proapoptotic stimuli and calcium signals or changes in the spatiotemporal pattern of the calcium signal or changes at the level of effectors activated by the calcium signal (e.g. calpain, calcineurin). The fate of the cell is often determined in the mitochondria, where calcium spikes may support cell survival through stimulation of ATP production or initiate apoptosis v ia opening of the permeability transition pore and release of apoptotic factors such as cytochrome c. The functional importance of these mitochondrial calcium signalling pathways has been underscored by the elucidation of a highly effective, local Ca(2+)coupling between IP(3)R/RyR and mitochondrial Ca(2+)uptake sites. This article will focus on the IP(3)R/RyR-dependent pathways to apoptosis, particularly on the mitochondrial phase of the death cascade. PMID- 11115375 TI - Electrical coupling and plasticity of the mitochondrial network. AB - Kinetic fluorescence imaging and the potentiometric probe tetramethylrhodamine methyl ester (TMRM) were used to evoke and detect changes in membrane potential (delta Psi(m)) of individual mitochondria in living cells. As a combined effect of preferential TMRM accumulation in mitochondria, and of TMRM photoactivation, individual organelles displayed sharp transient depolarizations caused by local reactive oxygen species (ROS)-mediated gatings of the mitochondrial permeability transition pore (PTP). In COS-7 cells, such directed repetitive gatings of the PTP gave rise to stochastic delta Psi(m)flickering at the level of individual organelles, but also to prominent synchronous delta Psi(m)transitions in whole subgroups of the mitochondrial population, indicative of the existence of an underlying electrically coupled mitochondrial network. In single cells, this network could comprise as much as 65% of the total mitochondrial population, a nd exhibited a high plasticity with mitochondrial units spontaneously connecting to and disconnecting from the coupled structure within seconds. These results indicate that in resting cells, the mitochondrial network is a dynamic proton conducting structure capable to commute and coordinate electrical signals generated by the PTP. PMID- 11115376 TI - Fabry disease: preclinical studies demonstrate the effectiveness of alpha galactosidase A replacement in enzyme-deficient mice. AB - Preclinical studies of enzyme-replacement therapy for Fabry disease (deficient alpha-galactosidase A [alpha-Gal A] activity) were performed in alpha-Gal A deficient mice. The pharmacokinetics and biodistributions were determined for four recombinant human alpha-Gal A glycoforms, which differed in sialic acid and mannose-6-phosphate content. The plasma half-lives of the glycoforms were approximately 2-5 min, with the more sialylated glycoforms circulating longer. After intravenous doses of 1 or 10 mg/kg body weight were administered, each glycoform was primarily recovered in the liver, with detectable activity in other tissues but not in the brain. Normal or greater activity levels were reconstituted in various tissues after repeated doses (10 mg/kg every other day for eight doses) of the highly sialylated AGA-1 glycoform; 4 d later, enzyme activity was retained in the liver and spleen at levels that were, respectively, 30% and 10% of that recovered 1 h postinjection. Importantly, the globotriaosylceramide (GL-3) substrate was depleted in various tissues and plasma in a dose-dependent manner. A single or repeated doses (every 48 h for eight doses) of AGA-1 at 0.3-10.0 mg/kg cleared hepatic GL-3, whereas higher doses were required for depletion of GL-3 in other tissues. After a single dose of 3 mg/kg, hepatic GL-3 was cleared for > or =4 wk, whereas cardiac and splenic GL-3 reaccumulated at 3 wk to approximately 30% and approximately 10% of pretreatment levels, respectively. Ultrastructural studies demonstrated reduced GL-3 storage posttreatment. These preclinical animal studies demonstrate the dose-dependent clearance of tissue and plasma GL-3 by administered alpha-Gal A, thereby providing the in vivo rationale-and the critical pharmacokinetic and pharmacodynamic data-for the design of enzyme-replacement trials in patients with Fabry disease. PMID- 11115377 TI - Mutation analysis of the entire PKD1 gene: genetic and diagnostic implications. AB - Mutation screening of the major autosomal dominant polycystic kidney disease (ADPKD) locus, PKD1, has proved difficult because of the large transcript and complex reiterated gene region. We have developed methods, employing long polymerase chain reaction (PCR) and specific reverse transcription-PCR, to amplify all of the PKD1 coding area. The gene was screened for mutations in 131 unrelated patients with ADPKD, using the protein-truncation test and direct sequencing. Mutations were identified in 57 families, and, including 24 previously characterized changes from this cohort, a detection rate of 52.3% was achieved in 155 families. Mutations were found in all areas of the gene, from exons 1 to 46, with no clear hotspot identified. There was no significant difference in mutation frequency between the single-copy and duplicated areas, but mutations were more than twice as frequent in the 3' half of the gene, compared with the 5' half. The majority of changes were predicted to truncate the protein through nonsense mutations (32%), insertions or deletions (29.6%), or splicing changes (6.2%), although the figures were biased by the methods employed, and, in sequenced areas, approximately 50% of all mutations were missense or in-frame. Studies elsewhere have suggested that gene conversion may be a significant cause of mutation at PKD1, but only 3 of 69 different mutations matched PKD1-like HG sequence. A relatively high rate of new PKD1 mutation was calculated, 1.8x10-5 mutations per generation, consistent with the many different mutations identified (69 in 81 pedigrees) and suggesting significant selection against mutant alleles. The mutation detection rate, in this study, of >50% is comparable to that achieved for other large multiexon genes and shows the feasibility of genetic diagnosis in this disorder. PMID- 11115378 TI - BMPR2 haploinsufficiency as the inherited molecular mechanism for primary pulmonary hypertension. AB - Primary pulmonary hypertension (PPH) is a potentially lethal disorder, because the elevation of the pulmonary arterial pressure may result in right-heart failure. Histologically, the disorder is characterized by proliferation of pulmonary-artery smooth muscle and endothelial cells, by intimal hyperplasia, and by in situ thrombus formation. Heterozygous mutations within the bone morphogenetic protein type II receptor (BMPR-II) gene (BMPR2), of the transforming growth factor beta (TGF-beta) cell-signaling superfamily, have been identified in familial and sporadic cases of PPH. We report the molecular spectrum of BMPR2 mutations in 47 additional families with PPH and in three patients with sporadic PPH. Among the cohort of patients, we have identified 22 novel mutations, including 4 partial deletions, distributed throughout the BMPR2 gene. The majority (58%) of mutations are predicted to lead to a premature termination codon. We have also investigated the functional impact and genotype phenotype relationships, to elucidate the mechanisms contributing to pathogenesis of this important vascular disease. In vitro expression analysis demonstrated loss of BMPR-II function for a number of the identified mutations. These data support the suggestion that haploinsufficiency represents the common molecular mechanism in PPH. Marked variability of the age at onset of disease was observed both within and between families. Taken together, these studies illustrate the considerable heterogeneity of BMPR2 mutations that cause PPH, and they strongly suggest that additional factors, genetic and/or environmental, may be required for the development of the clinical phenotype. PMID- 11115379 TI - A genome scan for renal function among hypertensives: the HyperGEN study. AB - Decreased renal function is often a complication of hypertension. Although it has been suggested that the response of the kidney to hypertension has an underlying genetic component, there is limited information suggesting that specific genetic regions or candidate genes contribute to the variability in creatinine clearance, a commonly used measure of kidney function. As part of the Hypertension Genetic Epidemiology Network (HyperGEN) study, creatinine clearance measurements were assessed in a large biracial sample of hypertensive siblings (466 African American subjects and 634 white subjects in 215 and 265 sibships, respectively). All participants were hypertensive before the age of 60 years, and the mean age of the siblings was 52 years among the African American subjects and 61 years among the white subjects. Two residual models were created for creatinine clearance: a minimally adjusted model (which included age and age(2)) and a fully adjusted model (which included age, age(2), lean body mass, pulse rate, pulse pressure, hormone-replacement therapy, educational status, and physical activity). Standardized residuals were calculated separately for men and women in both racial groups. The heritability of the residual creatinine clearance was 17% and 18% among the African American and white subjects, respectively. We conducted multipoint variance components linkage analysis using GENEHUNTER2 and 387 anonymous markers (Cooperative Human Linkage Center screening set 8). The best evidence for linkage in African American subjects was found on chromosome 3 (LOD = 3.61 at 214.6 cM, 3q27) with the fully adjusted model, and the best evidence in white subjects was found on chromosome 3 (LOD = 3.36 at 115.1 cM) with the minimally adjusted model. Positional candidate genes that are contained in and around the region on chromosome 3 (214.6 cM) that may contribute to renal function include enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase (EHHADH) and apolipoprotein D (ApoD). These findings suggest there may be genetic regions related to the variability of creatinine clearance among hypertensive individuals. PMID- 11115380 TI - Analysis of European mtDNAs for recombination. AB - The standard paradigm postulates that the human mitochondrial genome (mtDNA) is strictly maternally inherited and that, consequently, mtDNA lineages are clonal. As a result of mtDNA clonality, phylogenetic and population genetic analyses should therefore be free of the complexities imposed by biparental recombination. The use of mtDNA in analyses of human molecular evolution is contingent, in fact, on clonality, which is also a condition that is critical both for forensic studies and for understanding the transmission of pathogenic mtDNA mutations within families. This paradigm, however, has been challenged recently by Eyre Walker and colleagues. Using two different tests, they have concluded that recombination has contributed to the distribution of mtDNA polymorphisms within the human population. We have assembled a database that comprises the complete sequences of 64 European and 2 African mtDNAs. When this set of sequences was analyzed using any of three measures of linkage disequilibrium, one of the tests of Eyre-Walker and colleagues, there was no evidence for mtDNA recombination. When their test for excess homoplasies was applied to our set of sequences, only a slight excess of homoplasies was observed. We discuss possible reasons that our results differ from those of Eyre-Walker and colleagues. When we take the various results together, our conclusion is that mtDNA recombination has not been sufficiently frequent during human evolution to overturn the standard paradigm. PMID- 11115381 TI - Independent histories of human Y chromosomes from Melanesia and Australia. AB - To investigate the origins and relationships of Australian and Melanesian populations, 611 males from 18 populations from Australia, Melanesia, and eastern/southeastern Asia were typed for eight single-nucleotide polymorphism (SNP) loci and seven short tandem-repeat loci on the Y chromosome. A unique haplotype, DYS390.1del/RPS4Y711T, was found at a frequency of 53%-69% in Australian populations, whereas the major haplotypes found in Melanesian populations (M4G/M5T/M9G and DYS390.3del/RPS4Y711T) are absent from the Australian populations. The Y-chromosome data thus indicate independent histories for Australians and Melanesians, a finding that is in agreement with evidence from mtDNA but that contradicts some analyses of autosomal loci, which show a close relationship between Australian and Melanesian (specifically, highland Papua New Guinean) populations. Since the Australian and New Guinean landmasses were connected when first colonized by humans > or =50,000 years ago but separated some 8,000 years ago, a possible way to reconcile all the genetic data is to infer that the Y-chromosome and mtDNA results reflect the past 8,000 years of independent history for Australia and New Guinea, whereas the autosomal loci reflect the long preceding period of common origin and shared history. Two Y chromosome haplotypes (M119C/M9G and M122C/M9G) that originated in eastern/southeastern Asia are present in coastal and island Melanesia but are rare or absent in both Australia and highland Papua New Guinea. This distribution, along with demographic analyses indicating that population expansions for both haplotypes began approximately 4,000-6,000 years ago, suggests that these haplotypes were brought to Melanesia by the Austronesian expansion. Most of the populations in this study were previously typed for mtDNA SNPs; population differentiation is greater for the Y chromosome than for mtDNA and is significantly correlated with geographic distance, a finding in agreement with results of similar analyses of European populations. PMID- 11115382 TI - DFNA25, a novel locus for dominant nonsyndromic hereditary hearing impairment, maps to 12q21-24. AB - Using linkage analysis, we identified a novel dominant locus, DFNA25, for delayed onset, progressive, high-frequency, nonsyndromic sensorineural hearing loss in a large, multigenerational United States family of Czech descent. On the basis of recombinations in affected individuals, we determined that DFNA25 is located in a 20-cM region of chromosome 12q21-24 between D12S327 (centromeric) and D12S84 (telomeric), with a maximum two-point LOD score of 6.82, at recombination fraction.041, for D12S1030. Candidate genes in this region include ATP2A2, ATP2B1, UBE3B, and VR-OAC. DFNA25 may be the human ortholog of bronx waltzer (bv). PMID- 11115383 TI - Inadequate use of molecular hybridization to analyze DNA in Neanderthal fossils. PMID- 11115386 TI - Preparation and characterization of a new bioinorganic composite: collagen and hydroxyapatite. AB - Studies on biomineralization in bone have opened new arenas in the field of fabrication and engineering of bone-graft devices. A new bioinorganic composite was prepared by introducing processed collagenous matrices into the calcifying solutions at pH 7.8-8.0. The calcifying solutions were prepared by using a saturated dicalcium salt solution. Hydroxyapatite grew on the matrix over a period of 3-5 days in a specially designed crystallizing chamber. Changes in biochemical characters and biophysical parameters have given interesting insights into factors affecting and effecting initiation and progression of calcification on organic matrices. PMID- 11115385 TI - Stabilizing effect of chemical additives against oxidation of lactate dehydrogenase. AB - Oxidation is one of the major pathways for denaturation of proteins during storage, and also a potential problem in protein production, isolation and purification processes. In this study, a number of additives have been tested for their protective effects against oxidation in the presence of metal ions and hydrogen peroxide. Porcine muscle lactate dehydrogenase (LDH) was used as a model protein. Oxidation and denaturation of the enzyme were followed as activity loss, modification of certain amino acids, altered secondary structure and aggregation. Loss of activity during metal-catalysed oxidation was accompanied by structural damage of the enzyme, which was not the case during oxidation with peroxide. The best protectant during both modes of oxidation was found to be the polycation, poly(ethyleneimine) (PEI), followed by EDTA. Both chemicals also increased the enzyme's half-life during oxidation with a mixture of copper ions and hydrogen peroxide. Ammonium sulphate was an effective stabilizer during metal-catalysed oxidation, while sorbitol, sucrose and hydroxyectoine provided moderate stabilization. The ectoine was also stabilizing against oxidation with hydrogen peroxide, as was poly(ethylene glycol), whereas sorbitol enhanced the rate of enzyme inactivation. The stability of LDH towards denaturation by both oxidation and temperature was increased by addition of both PEI and sorbitol, as indicated by the melting-temperature profiles of the enzyme. PMID- 11115387 TI - Improved recovery of insulin-like growth factors (IGFs) from bovine colostrum using alkaline diafiltration. AB - Studies to develop a rapid, bioprocess-compatible method to recover low-molecular mass growth factors from bovine colostrum are reported. Defatted bovine colostrum was subjected to tangential-flow filtration (TFF) using two different filters [polyether sulphone (PES) and regenerated cellulose (RC)] at pH 5.8, pH 8.0 and pH 8.0+0. 01 M NaCl. Single-pass TFF at pH 5.8 using a 100 kDa RC filter resulted in the loss of approx. 90% of insulin-like growth factor I (IGF-I) to non specific filter adsorption. Comparison of 30 kDa RC and PES filters under single pass conditions showed that yields of IGF-1 and IGF-II were highest with RC filters. Yields of IGF-I and protein from both filter types were increased at pH 8.0 and were greatest for the 30 kDa RC filter. Effects of adding large diluent volumes continuously during TFF (diafiltration) were tested at pH 5. 0 and 8.0. The use of 10 diafiltrate vols. at pH 8.0 resulted in the recovery of 15-28% of colostral IGF-1 from the RC 30 kDa permeates, 2-4-fold greater than under acidic conditions. Yields of IGF-II (39.6%) were unaffected by pH and at least 97% of total protein was retained by the 30 kDa filter at pH 8.0. Denaturing SDS/PAGE analysis of the alkaline RC 30 kDa permeates demonstrated two major regions of stained proteins at 10-13 kDa and 17-19 kDa. Acidic TFF permeates contained additional stained proteins at approximately 90, 48 and 37 kDa. Isoelectric focusing of these samples demonstrated the presence of proteins with isoelectric points of 8.2 and 8.6. The current study demonstrates a one-step bioprocess compatible technique for the recovery of low-molecular-mass polypeptides from bovine colostrum. By using alkaline diafiltration with RC filters TFF provided optimal recovery of IGF-1 from colostrum. PMID- 11115388 TI - Efficiency of promoter and cell line in high-level expression of erythropoietin. AB - Efficiency of viral promoters and various cell lines in directing high-level expression of human erythropoietin (Epo) was investigated. To investigate the effects of various viral promoters and cell lines on the Epo expression level, genomic Epo with the 5' and 3' untranslated regions (UTRs) deleted was cloned next to the simian virus 40 early promoter, cytomegalovirus early promoter or SRalpha promoter. These expression vectors were transfected into COS-7, BHK-21 and Chinese hamster ovary (CHO)/dhfr(-) cells, respectively. The COS-7 cells transfected with the vector containing the SRalpha promoter showed the highest expression level ( approximately 103 IU/ml) at 72 h post-transfection. For the development of Epo-producing stable cell lines, BHK-21 and CHO/dhfr(-) cells transfected with the 5',3'-UTR-deleted genomic Epo under the control of the SRalpha promoter were cultured with media containing zeocin. Several clones of zeocin-resistant BHK-21 and CHO/dhfr(-) cells were cultured in the presence of methotrexate (MTX). A BHK-21 clone selected in the presence of 500 nM MTX expressed and secreted approximately 490 IU/ml Epo into the medium. A CHO/dhfr(-) clone selected in the presence of 20 nM MTX expressed and secreted approximately 45 IU/ml Epo into the medium. Southern-blot analysis indicated that enhancement of Epo expression in the MTX-resistant stable cells might be related to the amplification of gene copy number. PMID- 11115389 TI - Optimization of 6-aminopenicillanic acid (6-APA) production by using a new immobilized penicillin acylase. AB - A new immobilized penicillin acylase (ECPVA) was obtained by covalent binding of penicillin acylase from Streptomyces lavendulae on Eupergit C. Enzymic hydrolysis of penicillin V catalysed by ECPVA was optimized using a 2(3) factorial design of experiments, and the selected parameters for this study were pH, temperature and substrate concentration. The immobilized enzyme showed an optimal pH value of 9.5 10.5, and an optimal temperature of 60 degrees C, whereas its soluble counterpart showed the same optimal pH value and a lower optimal temperature of 50 degrees C. PMID- 11115390 TI - Characterization of recombinant invertase expressed in methylotrophic yeasts. AB - We studied, for the first time, characterization of the invertase expressed in the methylotrophic yeasts Hansenula polymorpha and Pichia pastoris in terms of enzyme conformational stability and structural behaviour induced by temperature as a function of pH using enzymic assays, differential scanning calorimetry, fluorescence and CD. The enzyme produced in both hosts was very stable over a broad range of pH values, keeping its enzymic activity and structure above 60 degrees C. Thermal denaturation, as measured by differential scanning calorimetry, was always irreversible. However, the fact that scanning rate had no effect on the calorimetric curves gave us the chance to analyse the data from a thermodynamic point of view. The conformational stabilities were essentially identical under the experimental conditions studied, but stability was always slightly higher in the enzyme expressed in H. polymorpha. This fact indicates that the greater degree of glycosylation of this enzyme form contributed to its increased global stability. Reactivation upon heating at 80 degrees C depends on protein concentration, suggesting that irreversibility could be associated with slow refolding kinetics at high protein concentration. PMID- 11115391 TI - Site-specific and random immobilization of thermolysin-like proteases reflected in the thermal inactivation kinetics. AB - Immobilization of proteins usually leads to random orientation of the molecules on the surface of the carrier material, whereby mechanistic interpretations of changes in properties, such as thermal stability, become very difficult. Recently, we have prepared several mutant enzymes of the thermolysin-like neutral protease from Bacillus stearothermophilus, containing cysteine residues in different positions on the surface of the protein molecule. These enzymes allowed site-specific immobilization to Activated Thiol-Sepharose and showed that stabilization effects strongly depend on the position of attachment [Mansfeld, Vriend, Van den Burg, Eijsink and Ulbrich-Hofmann (1999) Biochemistry 38, 8240 8245]. The greatest stabilization was achieved after immobilization of the mutant enzymes S65C and T56C/S65C within the structural region (positions 56-69) where unfolding is initiated. In this study thermal inactivation kinetics of these two mutant enzymes, as well as those of the pseudo-wild-type enzyme and thermolysin, were compared for different types of immobilization. Besides site-specific immobilization via thiol groups, the enzymes were bound randomly via their amino groups or by mixed-type binding. The basic matrix was Sepharose 4B in all carriers. Whereas the enzymes bound site-specifically to Activated Thiol Sepharose showed clear first-order inactivation kinetics like the soluble enzymes, the other immobilized enzyme preparations were characterized by distinct biphasic inactivation kinetics reflecting the heterogeneity of enzyme molecules on the carrier with respect to thermal unfolding. Site-specific binding resulted in stronger stabilization than the mixed binding type. However, immobilization to a highly functionalized carrier via amino groups increased stability further, suggesting that multiple fixation outside of the unfolding region 56-65 is able to increase stability of the enzyme molecules additionally. PMID- 11115392 TI - An Aspergillus chitosanase with potential for large-scale preparation of chitosan oligosaccharides. AB - A chitosan-degrading fungus, designated Aspergillus sp. Y2K, was isolated from soil. The micro-organism was used for producing chitosanase (EC 3.2.1.132) in a minimal medium containing chitosan as the sole carbon source. The induced chitosanase was purified to homogeneity from the culture filtrate by concentration and cationic SP-Sepharose chromatography. The purified enzyme is a monomer with an estimated molecular mass of 25 kDa by SDS/PAGE and of 22 kDa by gel-filtration chromatography. pI, optimum pH and optimum temperature values were 8.4, 6.5 and 65-70 degrees C, respectively. The chitosanase is stable in the pH range from 4 to 7.5 at 55 degrees C. Higher deacetylated chitosan is a better substrate. Chitin, xylan, 6-O-sulphated chitosan and O-carboxymethyl chitin were indigestible by the purified enzyme. By endo-splitting activity, the chitosanase hydrolysed chitosan to form chitosan oligomers with chitotriose, chitotetraose and chitopentaose as the major products. The enzyme hydrolyses chitohexaose to form chitotriose, while the chitopentaose and shorter oligomers remain intact. The N-terminal amino acid sequence of the enzyme was determined as YNLPNNLKQIYDDHK, which provides useful information for further gene cloning of this enzyme. A 275 g-scale hydrolysis of chitosan was performed. The product distribution was virtually identical to that of the small-scale reaction. Owing to the simple purification process and high stability of the enzyme, it is potentially valuable for industrial applications. PMID- 11115394 TI - Msx3 protein recruits histone deacetylase to down-regulate the Msx1 promoter. AB - Msx1 promoter is known to be repressed by Msx1 protein [Shetty, Takahashi, Matsui, Iyengar and Raghow (1999) Biochem. J. 339, 751-758]. We show that in the transiently transfected C(2)C(12) myoblasts, co-expression of Msx3 also causes potent repression of Msx1 promoter that can be relieved by exogenous expression of cAMP-response-element-binding protein-binding protein (CBP) and p300 in a dose dependent manner. Co-immunoprecipitation and Western blot analyses revealed that Msx3 interacts with CBP and p300 and this interaction significantly decreases the histone acetyltransferase (HAT) activity of both proteins. We also discovered that Msx3-mediated repression of Msx1 promoter is synergized by the exogenous co expression of histone deacetylase 1 (HDAC1). Furthermore, the repression of Msx1 promoter by Msx3 could be relieved by treating transfected cells with trichostatin A, an inhibitor of HDAC(s). Finally, we show that Msx3 and HDAC1 can be co-immunoprecipitated in a complex that does not contain CBP and that Msx3 and HDAC1 proteins are co-localized in the nucleus. Taken together, our results strongly suggest that two distinct multiprotein complexes are present within the nuclei of C(2)C(12) cells: one containing Msx3 and HDAC(s) and another containing Msx3 and CBP and/or p300. On the basis of these results, we propose a dual mechanism of repression by Msx3 protein that involves the squelching of the HAT activity of co-activators, CBP and p300, and recruitment of HDAC(s). PMID- 11115393 TI - The neuronal calcium sensor family of Ca2+-binding proteins. AB - Ca(2+) plays a central role in the function of neurons as the trigger for neurotransmitter release, and many aspects of neuronal activity, from rapid modulation to changes in gene expression, are controlled by Ca(2+). These actions of Ca(2+) must be mediated by Ca(2+)-binding proteins, including calmodulin, which is involved in Ca(2+) regulation, not only in neurons, but in most other cell types. A large number of other EF-hand-containing Ca(2+)-binding proteins are known. One family of these, the neuronal calcium sensor (NCS) proteins, has a restricted expression in retinal photoreceptors or neurons and neuroendocrine cells, suggesting that they have specialized roles in these cell types. Two members of the family (recoverin and guanylate cyclase-activating protein) have established roles in the regulation of phototransduction. Despite close sequence similarities, the NCS proteins have distinct neuronal distributions, suggesting that they have different functions. Recent work has begun to demonstrate the physiological roles of members of this protein family. These include roles in the modulation of neurotransmitter release, control of cyclic nucleotide metabolism, biosynthesis of polyphosphoinositides, regulation of gene expression and in the direct regulation of ion channels. In the present review we describe the known sequences and structures of the NCS proteins, information on their interactions with target proteins and current knowledge about their cellular and physiological functions. PMID- 11115395 TI - Mucin biosynthesis and secretion in tracheal epithelial cells in primary culture. AB - Density-gradient centrifugation of bovine tracheal epithelial cell extracts revealed a 'high-density' (1.48 g/ml) sialic-acid-rich population as well as a 'low-density' (1.42 g/ml) one that reacted more strongly with a periodate-Schiff (PAS) assay. The sialic-acid-rich mucins were oligomeric molecules containing disulphide- bond-linked subunits and large glycosylated domains, whereas the PAS reactive component seemed to be smaller and 'monomeric'. Only the 'high-density' population was secreted from cells cultured for 5 days on plastic or a collagen type 1, Matrigel or Vitrogen substrate. Release was less from cells grown on plastic than from those on a substrate and the amount was unaffected by increasing the thickness of the collagen layer. For cells grown on collagen, the amount of the sialic-acid-rich mucin increased over 10 days, whereas the PAS reactive component was largely absent after 24 h, which was consistent with an initial release of stored PAS-reactive molecules and synthesis of the sialic-acid rich mucins de novo. Both [(3)H]proline and [(35)S]sulphate were poorly incorporated into mucins detected with the chemical assays but molecules with a higher buoyant density than that of either of the previously identified species were labelled with [(35)S]sulphate. The [(35)S]sulphate-labelled material yielded large trypsin-resistant fragments and contained O-linked glycans but was not affected by digestion with chondroitin ABC lyase or heparan sulphate lyase, suggesting that it is a mucin rather than a proteoglycan. [(35)S]Sulphate is thus a poor marker for the major oligomeric mucins produced by bovine tracheal epithelial cells but the radiolabel is incorporated into a heavily labelled mucin like component. PMID- 11115396 TI - Biosynthesis and shedding of epiglycanin: a mucin-type glycoprotein of the mouse TA3Ha mammary carcinoma cell. AB - Epiglycanin is a mucin-type glycoprotein present at the surface of TA3Ha mouse mammary tumour cells. It is a long rod-like glycoprotein with a molecular mass of 500 kDa. Its function has not yet been established but its overexpression can affect cell-cell and cell-matrix adhesion. To understand better the biological function of epiglycanin, we have studied the biochemical structure and biosynthesis of epiglycanin in TA3Ha cells. Pulse-chase labelling experiments with [(3)H]threonine revealed an early precursor with a molecular mass of approx. 300 kDa containing approx. 5-10 kDa of N-linked glycans. The precursor was gradually converted into a high-molecular-mass mature form, owing mainly, if not entirely, to O-glycosylation. The mature molecule consists of two major glycoforms that differ in sialylation. Unlike secreted mucins, epiglycanin did not form cysteine-bound multimers, providing further evidence that epiglycanin belongs to the class of membrane-associated mucins. The mature form, but not the precursor form, is shed from the cell surface. The half-life of epiglycanin on the cell surface was found to be approx. 60 h. These results provide the first detailed analysis of the biochemical structure and biosynthesis of epiglycanin. PMID- 11115398 TI - Cloning and functional expression of a degradation-resistant novel isoform of p27Kip1. AB - p27(Kip1) is an inhibitor of cyclin-dependent kinases. It has been implicated as having a role in the induction of growth arrest at the G(1) phase of the cell cycle in response to anti-mitogenic signals such as cell contact and serum starvation. Proteasome-mediated degradation plays an important role in the rapid inactivation of p27(Kip1), causing quiescent cells to re-enter the cell cycle. Although the existence of a second isoform has been suggested, no such isoform was isolated. Through screening of a cDNA library derived from growth-arrested confluent porcine endothelial cells, we obtained clones for a novel isoform of p27(Kip1) in addition to the original isoform. The novel isoform differed from the original isoform at the C-terminus. The tissue-specific expression of the original and novel isoforms was demonstrated at the mRNA and protein levels. An in vitro degradation assay demonstrated this novel isoform to be resistant to proteasome-mediated destruction. The expression as a fusion protein with green fluorescent protein revealed this isoform to be targeted to the nucleus by a bipartite nuclear-localization signal with a C-terminal part different from that of the original isoform. The expression of the novel isoform caused the growth arrest of HeLa cells and an accumulation of cells in the G(0)/G(1) phase, and this effect was similar to that seen with the original isoform. The present study suggests that the novel isoform functions as a negative regulator of the cell cycle, and may play a distinct role. The novel isoform was named p27(Kip1R) because of its resistance to degradation. PMID- 11115397 TI - Identification of a novel isoform of the cyclic-nucleotide phosphodiesterase PDE3A expressed in vascular smooth-muscle myocytes. AB - We have identified a new cyclic-nucleotide phosphodiesterase isoform, PDE3A, and cloned its cDNA from cultured aortic myocytes. The nucleotide sequence of its coding region is similar to that of the previously cloned myocardial isoform except for the absence of the initial 300-400 nt that are present in the latter, as confirmed by reverse-transcriptase-mediated PCR, 5' rapid amplification of cDNA ends and a ribonuclease protection assay. Expression in Spodoptera frugiperda (Sf9) cells yields a protein with catalytic activity and inhibitor sensitivity typical of the PDE3 family. The recombinant protein's molecular mass of approx. 131 kDa is compatible with translation from an ATG sequence corresponding to nt 436-438 of the myocardial PDE3A coding region. Antibodies against residues 424-460 (nt 1270-1380) and 1125-1141 (nt 3373-3423) of the myocardial isoform react with an approx. 118 kDa band in Western blots of homogenates of human aortic myocytes, whereas antibodies against residues 29-42 (nt 85-126) do not react with any bands in these homogenates. Our results suggest that a vascular smooth-muscle isoform ('PDE3A2') is a product of the same gene as the longer myocardial ('PDE3A1') and the shorter placental ('PDE3A3') isoforms and is generated pre-translationally in a manner that results in the absence of the 145 N-terminal amino acids of PDE3A1. PMID- 11115399 TI - Escherichia coli engineered to synthesize isopentenyl diphosphate and dimethylallyl diphosphate from mevalonate: a novel system for the genetic analysis of the 2-C-methyl-d-erythritol 4-phosphate pathway for isoprenoid biosynthesis. AB - Isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP) constitute the basic building block of isoprenoids, a family of compounds that is extraordinarily diverse in structure and function. IPP and DMAPP can be synthesized by two independent pathways: the mevalonate pathway and the recently discovered 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway. Although the MEP pathway is essential in most eubacteria, algae and plants and has enormous biotechnological interest, only some of its steps have been determined. We devised a system suitable for the genetic analysis of the MEP pathway in Escherichia coli. A synthetic operon coding for yeast 5-diphosphomevalonate decarboxylase, human 5-phosphomevalonate kinase, yeast mevalonate kinase and E. coli isopentenyl diphosphate isomerase was incorporated in the chromosome of this bacterium. The expression of this operon allowed the synthesis of IPP and DMAPP from mevalonate added exogenously and complementation of lethal mutants of the MEP pathway. We used this system to show that the ygbP, ychB and ygbB genes are essential in E. coli and that the steps catalysed by the products of these genes belong to the trunk line of the MEP pathway. PMID- 11115400 TI - Novel properties of the protein kinase CK2-site-regulated nuclear- localization sequence of the interferon-induced nuclear factor IFI 16. AB - Members of the interferon-induced class of nuclear factors possess a putative CcN motif, comparable with that within proteins such as the simian virus 40 large tumour antigen (T-ag), which confers phosphorylation-mediated regulation of nuclear-localization sequence (NLS)-dependent nuclear import. Here we examine the functionality of the interferon-induced factor 16 (IFI 16) CcN motif, demonstrating its ability to target a heterologous protein to the nucleus, and to be phosphorylated specifically by the CcN-motif-phosphorylating protein kinase CK2 (CK2). The IFI 16 NLS, however, has novel properties, conferring ATP dependent nuclear import completely independent of cytosolic factors, as well as binding to nuclear components. The IFI 16 NLS is not recognized with high affinity by the NLS-binding importin heterodimer, and transport mediated by it is insensitive to non-hydrolysable GTP analogues. The IFI 16 NLS thus mediates nuclear import through a pathway completely distinct from that of conventional NLSs, such as that of T-ag, but intriguingly resembling that of the NLS of the HIV-1 transactivator protein Tat. Since the IFI 16 CK2 site enhances nuclear import through facilitating binding to nuclear components, this represents a novel mechanism by which the site regulates nuclear-protein import, and constitutes a difference between the IFI 16 and Tat NLSs that may be of importance in the immune response. PMID- 11115402 TI - Evidence for the presence of phospholipid hydroperoxide glutathione peroxidase in human platelets: implications for its involvement in the regulatory network of the 12-lipoxygenase pathway of arachidonic acid metabolism. AB - The 12-lipoxygenase pathway of arachidonic acid metabolism in platelets and other cells is bifurcated into a reduction route yielding 12-hydroxyeicosatetraenoic acid (12-HETE) and an isomerization route forming hepoxilins. Here we show for the first time the presence of phospholipid hydroperoxide glutathione peroxidase (PHGPx) protein and its activity in platelets. The ratio of the activity of PHGPx to that of cytosolic glutathione peroxidase (GPx-1) was consistently found to be approx. 1:60 in platelets and UT7 megakaryoblasts. Moreover, short-lived PHGPx mRNA was detected in megakaryocytes but not in platelets. Carboxymethylation of selenium-containing glutathione peroxidases by iodoacetate, which results in the inactivation of PHGPx and GPx-1 without inhibition of 12-lipoxygenase, markedly altered the pattern of arachidonic acid metabolism in human platelets. Whereas the formation of 12-HETE was inhibited by 80%, a concomitant accumulation of 12 hydroperoxyeicosatetraenoic acid (12-HpETE) by two orders of magnitude as well as the formation of hepoxilins A(3) and B(3) were observed. The formation of hepoxilins also occurred when 12-HpETE was added to untreated platelets. In selenium-deficient UT7 cells, which were devoid of GPx-1 but not of PHGPx, the reduction of 12-HPETE was retained, albeit with a lower rate than in control cells containing GPx-1. We therefore believe that both GPx-1 and PHGPx are involved in the regulatory network of the 12-lipoxygenase pathway in platelets and other mammalian cells. Moreover, the diminution of hydroperoxide tone in platelets incubated with arachidonic acid leads primarily to the formation of 12 HETE, whereas the increase in hydroperoxide tone (a situation found under oxidative stress or selenium deficiency or on incubation with 12-HPETE) partly diverts the 12-lipoxygenase pathway from the reduction route to the isomerization route, thus resulting in the formation of hepoxilins. PMID- 11115401 TI - Cytosolic phospholipase A2-alpha associates with plasma membrane, endoplasmic reticulum and nuclear membrane in glomerular epithelial cells. AB - Eicosanoids mediate complement-dependent glomerular epithelial injury in experimental membranous nephropathy. The release of arachidonic acid from phospholipids by cytosolic phospholipase A(2) (cPLA(2)) is the rate-limiting step in eicosanoid synthesis. The present study examines the association of cPLA(2) with membranes of organelles. Glomerular epithelial cells were disrupted by homogenization in Ca(2+)-free buffer; organelles were separated by gradient centrifugation. The distribution of cPLA(2) and organelles was analysed by immunoblotting with antibodies against cPLA(2) and organelle markers, or by enzyme assay. In cells incubated with or without the Ca(2+) ionophore ionomycin plus PMA, cPLA(2) co-localized with plasma membrane, endoplasmic reticulum and nuclei, but not with mitochondria or Golgi. A greater amount of cPLA(2) was associated with membranes in stimulated cells, but membrane-associated cPLA(2) was readily detectable under resting conditions. The pattern of association of cPLA(2) with membrane in cells treated with antibody and complement was similar to that in cells stimulated with ionomycin plus PMA; however, complement did not enhance the membrane association of cPLA(2) protein. To determine the functional role of membrane association of cPLA(2), phospholipids were labelled with [(3)H]arachidonic acid. Cells were then incubated with or without antibody and complement and were fractionated. Complement induced a loss of radioactivity from the plasma membrane, endoplasmic reticulum and nuclei, but not from the mitochondrial fraction. Thus the release of arachidonic acid by cPLA(2) is due to the hydrolysis of phospholipids at multiple subcellular membrane sites, including the endoplasmic reticulum, plasma membrane and nucleus. PMID- 11115403 TI - Apoptosis of haematopoietic cells upon thymidylate synthase inhibition is independent of p53 accumulation and CD95-CD95 ligand interaction. AB - Treatment of haematopoietic BA/F3 cells with the thymidylate synthase inhibitor 5 fluoro-2'-deoxyuridine (FUdR) activated apoptosis through a mechanism that required continuous protein synthesis and was inhibited by Bcl-2 over-expression. Analysis of p53 levels in cells treated with FUdR indicated a marked accumulation of this protein. Accumulation of p53 was also observed in cells over-expressing Bcl-2. In BA/F3 cells transfected with a cDNA coding for the human papilloma virus protein E6, p53 accumulation after FUdR treatment was inhibited markedly. However, apoptosis was induced in both control and E6 cells to a similar extent. The role of the CD95/CD95 ligand (CD95L) system in FUdR-induced apoptosis was also assessed. As determined by reverse transcriptase PCR, BA/F3 expressed a low constitutive level of CD95L mRNA, which decreased following FUdR treatment. Moreover, blocking CD95-CD95L interactions with antagonistic CD95 monoclonal antibody did not prevent drug-induced apoptosis. Furthermore, analysis of caspase involvement showed important differences in apoptosis induced by CD95-triggering or FUdR treatment. In summary, these results suggest that apoptosis induced by thymineless stress in haematopoietic BA/F3 cells occurs by a mechanism that does not require accumulation of p53 and which is independent of CD95-CD95L interactions. PMID- 11115404 TI - Interleukin 9 induces expression of three cytokine signal inhibitors: cytokine inducible SH2-containing protein, suppressor of cytokine signalling (SOCS)-2 and SOCS-3, but only SOCS-3 overexpression suppresses interleukin 9 signalling. AB - Interleukin 9 (IL-9) is a cytokine preferentially produced by T helper type 2 lymphocytes and active on various cell types such as T- and B-lymphocytes, mast cells and haemopoietic progenitors. The IL-9 receptor (IL-9R) belongs to the haemopoietic receptor superfamily and its signal transduction involves mainly the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway. Here we studied the implication of a novel family of suppressors of cytokine signalling (called CIS, for cytokine-inducible SH2-containing protein, and SOCS, for suppressor of cytokine signalling) in IL-9 signal attenuation. In BW5147 T cell lymphoma, IL-9 induced the rapid expression of CIS, SOCS-2 and SOCS-3 with a peak after 2 h of stimulation. Using IL-9R mutants, we showed that STAT activation is required for CIS/SOCS induction: CIS and SOCS-2 expression was induced either via STAT1 and/or STAT3 or via STAT5 but only STAT1 and/or STAT3 were involved in SOCS-3 expression. The effect of these three proteins on IL-9 signal transduction was assessed by transient transfection in HEK-293 cells expressing the components of the IL-9 signalling pathway and a STAT-responsive reporter construct. These experiments showed that only SOCS-3 is able to inhibit IL-9-induced signal transduction; neither CIS nor SOCS-2 exerted any effect. Stable transfection of CIS and SOCS-3 in BW5147 lymphoma cells showed that only overexpression of SOCS-3 had an inhibitory activity on STAT activation, gene induction and the anti-apoptotic activity of IL-9. By contrast, CIS failed to affect the IL-9 response. PMID- 11115405 TI - Biochemical characterization and mechanism of action of a thermostable beta glucosidase purified from Thermoascus aurantiacus. AB - An extracellular beta-glucosidase from Thermoascus aurantiacus was purified to homogeneity by DEAE-Sepharose, Ultrogel AcA 44 and Mono-P column chromatography. The enzyme was a homotrimer, with a monomer molecular mass of 120 kDa; only the trimer was optimally active at 80 degrees C and at pH 4.5. At 90 degrees C, the enzyme showed 70% of its optimal activity. It was stable at pH 5.2 and at temperatures up to 70 degrees C for 48 h, but stability decreased above 70 degrees C and at pH values above and below 5.0. The enzyme hydrolysed aryl and alkyl beta-d-glucosides and cello-oligosaccharides, and was specific for substrates with a beta-glycosidic linkage. The hydroxy groups at positions 2, 4 and 6 of a glucose residue at the non-reducing end of a disaccharide appeared to be essential for catalysis. The enzyme had the lowest K(m) towards p-nitrophenyl beta-d-glucoside (0.1137 mM) and the highest k(cat) towards cellobiose and beta,beta-trehalose (17052 min(-1)). It released one glucose unit at a time from the non-reducing end of cello-oligosaccharides, and the rate of hydrolysis decreased with an increase in chain length. Glucose and d-delta-gluconolactone inhibited the beta-glucosidase competitively, with K(i) values of 0.29 mM and 8.3 nM respectively, while methanol, ethanol and propan-2-ol activated the enzyme. The enzyme catalysed the synthesis of methyl, ethyl and propyl beta-d-glucosides in the presence of methanol, ethanol and propan-2-ol respectively with either glucose or cellobiose, although cellobiose was preferred. An acidic pH favoured hydrolysis and transglycosylation, but high concentrations of alcohols favoured the latter reaction. The stereochemistry of cellobiose hydrolysis revealed that beta-glucosidase from T. aurantiacus is a retaining glycosidase, while N-terminal amino acid sequence alignment indicated that it is a member of glycoside hydrolase family 3. PMID- 11115406 TI - Diacylglycerol kinase theta is translocated and phosphoinositide 3-kinase dependently activated by noradrenaline but not angiotensin II in intact small arteries. AB - Diacylglycerol (DG) kinase (DGK) phosphorylates the lipid second messenger DG to phosphatidic acid. We reported previously that noradrenaline (NA), but not angiotensin II (AII), increases membrane-associated DGK activity in rat small arteries [Ohanian and Heagerty (1994) Biochem. J. 300, 51-56]. Here, we have identified this DGK activity as DGKtheta, present in both smooth muscle and endothelial cells of these small vessels. Subcellular fractionation of artery homogenates revealed that DGKtheta was present in nuclear, plasma membrane (and/or Golgi) and cytosolic fractions. Upon NA stimulation, DGKtheta translocated towards the membrane and cytosol (155 and 153% increases relative to the control, respectively) at 30 s, followed by a return to near-basal levels at 5 min; AII was without effect. Translocation to the membrane was to both Triton soluble and -insoluble fractions. NA, but not AII, transiently increased DGKtheta activity in immunoprecipitates (126% at 60 s). Membrane translocation and DGKtheta activation were regulated differently: NA-induced DGKtheta activation, but not translocation, was dependent on transient activation of phosphoinositide 3-kinase (PI 3-K). In addition, DGK activity co-immunoprecipitated with protein kinase B, a downstream effector of PI 3-K, and was increased greatly by NA stimulation. The rapid and agonist-specific activation of DGKtheta suggests that this pathway may have a physiological role in vascular smooth-muscle responses. PMID- 11115407 TI - Extracellular mechanism through the Edg family of receptors might be responsible for sphingosine-1-phosphate-induced regulation of DNA synthesis and migration of rat aortic smooth-muscle cells. AB - Exogenous sphingosine 1-phosphate (S1P) increased cytosolic Ca(2+) concentration, stimulated thymidine incorporation (DNA synthesis) and inhibited cell migration in rat aortic smooth-muscle cells (AoSMCs). Although exogenous sphingosine, a substrate of sphingosine kinase or a precursor of S1P, markedly induced the intracellular accumulation of S1P, the lipid failed to mimic the S1P-induced actions. In contrast, dihydrosphingosine 1-phosphate (DHS1P), an S1P receptor agonist, duplicated these S1P actions even though DHS1P was approx. 20-50-fold less potent than S1P. The pharmacological properties of DHS1P for the S1P receptor subtypes Edg-1, Edg-3, Edg-5 and Edg-6 were compared in Chinese hamster ovary (CHO) cells that were overexpressing the respective receptor. In these S1P receptor-overexpressing cells, DHS1P was approx. 20-30-fold less potent than S1P for the displacement of [(3)H]S1P binding and inositol phosphate response in Edg 5-expressing CHO cells, as was the case for AoSMCs. However, it was slightly (not more than 3-fold) less potent than S1P in cells expressing Edg-1, Edg-3 or Edg-6. Of the above-mentioned four types of S1P receptor, Edg-5 was abundantly expressed in AoSMCs, as demonstrated by Northern blotting. These results suggest that the intracellular accumulation of S1P is not necessary for the S1P-induced Ca(2+) response, for the stimulation of DNA synthesis or for the inhibition of cell migration. Thus these S1P-induced actions might be mediated through extracellular (or cell-surface) S1P receptors in AoSMCs: Edg-5 might be a most important receptor subtype. PMID- 11115409 TI - Analysis and modification of trehalose 6-phosphate levels in the yeast Saccharomyces cerevisiae with the use of Bacillus subtilis phosphotrehalase. AB - In the yeast Saccharomyces cerevisiae, trehalose is synthesized by the trehalose synthase complex in two steps. The Tps1 subunit catalyses the formation of trehalose 6-phosphate (Tre6P), which is dephosphorylated by the Tps2 subunit. Tps1 also controls sugar influx into glycolysis; a tps1 deletion strain is therefore unable to grow on glucose. It is unclear whether this regulatory function of Tps1 is mediated solely by Tre6P or also involves the Tps1 protein. We have developed a novel sensitive and specific assay method for Tre6P. It is based on the conversion of Tre6P into glucose and glucose 6-phosphate with purified phosphotrehalase from Bacillus subtilis. The glucose formed is measured with the glucose-oxidase/peroxidase method. The Tre6P assay is linear in the physiological concentration range. The detection limit, including the entire extraction procedure, is 15 nmol, corresponding to an intracellular concentration of 100 microM. To modify Tre6P levels in vivo, we expressed B. subtilis phosphotrehalase in yeast. The enzyme is functional because it rescues the temperature-sensitive growth defect of a tps2Delta strain and drastically lowers Tre6P levels in this strain. However, phosphotrehalase expression remains without effect on Tre6P levels in wild-type strains, as opposed to overexpression of Tps2. Because Tps2 is part of the Tre6P synthase (TPS) complex and because this complex is destabilized in tps2 deletion strains, these results can be explained if Tre6P is sequestered within the TPS complex in wild-type cells. The very low levels of Tre6P in cells overexpressing Tps2 have a limited effect on sugar phosphate accumulation and do not prevent growth on glucose. Taken together, our results support a model in which the regulatory function of Tps1 on sugar influx is mediated both by the Tps1 protein and by Tre6P. PMID- 11115408 TI - Nuclear factor kappaB-dependent mechanisms coordinate the synergistic effect of PMA and cytokines on the induction of superoxide dismutase 2. AB - Manganese superoxide dismutase (MnSOD) serves a protective role under conditions of oxidative stress mediated by such diverse agents as adriamycin, radiation, chemical hypoxia and ischaemia and might act as a newly recognized type of tumour suppressor. MnSOD is an inducible enzyme; however, the signalling molecules and pathways involved in its induction have not been fully elucidated. Recently we reported the identification of a 342 bp enhancer within the second intron (I2E) of the human gene encoding MnSOD (SOD2), which contains sites for binding nuclear factor kappaB (NF-kappaB), CCAAT-enhancer-binding protein (C/EBP) and nuclear factor 1 (NF-1). Using a human fibroblast cell line transformed by simian virus 40, we have identified the I2E fragment as being responsive to PMA. Furthermore, simultaneous treatment with PMA and cytokines (tumour necrosis factor alpha and interleukin 1beta) synergistically increases MnSOD induction. The use of mutant constructs identified the NF-kappaB element within the enhancer fragment as being essential for the PMA and PMA/cytokine effect. Mutations in the C/EBP- and NF-1 binding sites revealed a potential co-operation between proteins that bind to these sites and the NF-kappaB element. Evaluation of inhibitory kappaB (IkappaB) alpha and IkappaB-beta proteins reveals agent-specific differences in their turnover kinetics. Both C/EBP and NF-kappaB DNA-binding activities were increased in cells receiving a combination of cytokine and PMA. Supershift and immunoprecipitation studies suggest a physical interaction between C/EBP and NF kappaB proteins. Taken together, these studies suggest the activation of multiple transcription factors as well as pathways leading to increased NF-kappaB activity as being the mechanisms responsible for the synergistic induction of MnSOD by PMA and cytokines. PMID- 11115410 TI - The role of c-Myb in the up-regulation of methionine adenosyltransferase 2A expression in activated Jurkat cells. AB - Methionine adenosyltransferase (MAT) is a critical cellular enzyme which catalyses the formation of S-adenosylmethionine (SAM), the principal methyl donor. In mammals, two different genes, MAT1A and MAT2A, encode liver-specific and non-liver-specific MATs, respectively. SAM level increases during T lymphocyte activation and is required for proliferation. A major mechanism for the increase in SAM level is increased MAT2A transcription. In the current work we examined the molecular mechanism of increased MAT2A expression in activated Jurkat cells. Treatment of Jurkat cells with interleukin-2 (IL-2), PMA or PMA plus phytohaemagglutinin (PHA) resulted in a 2-fold increase in MAT2A mRNA levels and a 2-fold increase in luciferase activity driven by the transfected human MAT2A promoter construct -571/+60 but not -270/+60. The region -571 to -270 of the human MAT2A contains a c-Myb consensus binding site. c-Myb is known to be induced during T-lymphocyte activation and its mRNA level was increased after treatment of Jurkat cells with IL-2, PMA or PMA plus PHA. Increased nuclear binding to the MAT2A c-Myb site was confirmed on electrophoretic mobility-shift and supershift analyses. Mutation of the MAT2A c-Myb site abolished the stimulatory effect of these agents on c-Myb nuclear binding and MAT2A promoter activities. Overexpression of c-Myb increased MAT2A promoter activity by 2-fold. Dexamethasone, a known inhibitor of lymphocyte activation, blocked the effect of these agents on MAT2A expression by preventing the increase in c-Myb expression. PMID- 11115411 TI - Role of inducible nitric oxide synthase in the regulation of leucocyte recruitment. AB - Constitutively produced nitric oxide released by endothelial cells has been shown to act as an endogenous agent which inhibits the rolling and adhesion of leucocytes in the microcirculation. However, during various types of inflammation, expression of the inducible form of nitric oxide synthase (iNOS) can dramatically increase the amount of nitric oxide present in tissues. Furthermore, as iNOS can be expressed by a wide variety of cell types, the distribution of nitric oxide is likely to be altered relative to that in unstimulated tissue. Under these conditions, it is less well understood whether iNOS-derived nitric oxide retains the anti-adhesive capabilities of constitutively produced nitric oxide. This review summarizes work done to examine this issue. Three main approaches have been used. In vitro studies have examined the role of iNOS in adhesive interactions between stimulated endothelial cells and leucocytes, providing evidence of an anti-adhesive effect of iNOS. In addition, the role of iNOS has been examined in vivo in animal models of inflammation using pharmacological iNOS inhibitors. These experiments were extended by the advent of the iNOS-deficient (iNOS(-/-)) mouse. Intravital microscopy studies of these mice have indicated that, under conditions of low dose endotoxaemia, iNOS-derived nitric oxide can inhibit leucocyte rolling and adhesion. The potential mechanisms for these effects are discussed. In contrast, several other studies have observed either no effect or an enhancing effect of iNOS on inflammatory leucocyte recruitment. Taken together, these studies suggest that the importance of iNOS in modulating leucocyte recruitment can vary according to the type of inflammatory response. PMID- 11115412 TI - Losartan, an angiotensin type I receptor antagonist, improves conduit vessel endothelial function in Type II diabetes. AB - We have demonstrated previously that inhibition of angiotensin-converting enzyme (ACE) with enalapril and angiotensin II blockade with losartan improve acetylcholine-dependent endothelial function in resistance vessels of patients with Type II diabetes. It was therefore of interest to examine the effect of losartan on conduit vessel function in this group. The influence of losartan (50 mg daily for 4 weeks) on endothelium-dependent and -independent vasodilator function was determined in 12 subjects with Type II diabetes using a randomized, double-blind, placebo-controlled crossover protocol. Conduit vessel endothelial function was assessed using high-resolution ultrasound and the brachial artery response to reactive hyperaemia (flow-mediated dilation; FMD); glyceryl trinitrate (GTN) was used as a non-endothelium-dependent dilator. Losartan administration significantly increased the FMD response from 5.2+/-0.7% (mean+/ S.E.M.) to 7.4+/-0.6% of vessel diameter (P<0.05; paired t-test). There was no effect of losartan on the endothelium-independent responses to GTN (17.8+/-1.8% to 17.6+/-1.2%). Consistent with our previous findings in resistance vessels, administration of 50 mg of losartan daily improves NO-mediated dilation in the conduit vessels of subjects with Type II diabetes. Together with the findings that both ACE inhibition and angiotensin II blockade improve resistance vessel function in this group, it is likely that at least some of the beneficial effect is mediated through the angiotensin II/type I receptor pathway. A type I receptor antagonist seems a reasonable alternative to an ACE inhibitor to maintain conduit vessel endothelial function in Type II diabetic subjects. PMID- 11115413 TI - Hydrocortisone abolishes the angiotensin II-mediated potentiation of endothelin-1 in bovine bronchi. AB - Angiotensin II potentiates methacholine-evoked bronchoconstriction both in bovine airways in vitro and in asthmatic patients in vivo. Angiotensin II also potentiates endothelin-1-evoked contractions in vitro, but fails to alter such contractions in vivo. One possible confounding factor in patients is their use of inhaled corticosteroids. Accordingly the present study examined the effects of hydrocortisone (cortisol) on contractions evoked by methacholine and endothelin-1 in the presence and absence of angiotensin II. Contractions of rings of isolated bovine airways were measured isometrically in organ baths. Concentration-response curves were obtained for endothelin-1 or methacholine in the presence and absence of angiotensin II, hydrocortisone and a combination of angiotensin II and hydrocortisone. Hydrocortisone abolished the angiotensin II-mediated potentiation of endothelin-1-evoked, but not methacholine-evoked, contractions. Hydrocortisone alone evoked the enhancement of methacholine responses, similar to the effect produced by angiotensin II. While species differences may exist, our present results suggest that the use of corticosteroids can have a profound effect on the interaction between angiotensin II and endothelin-1. Accordingly, the presence of inhaled corticosteroids might explain the differences between the results obtained in vitro and in vivo. PMID- 11115414 TI - Relationship between the concentrations of plasma phospholipid stearic acid and plasma lipoprotein lipids in healthy men. AB - This study investigated the correlation between the plasma phospholipid (PL) saturated fatty acid (SFA) concentration (as a surrogate marker of SFA intake) and plasma lipid and lipoprotein lipid concentrations in 139 healthy Australian men aged 20-55 years old with widely varying intakes of saturated fat (vegans, n=18; ovolacto vegetarians, n=43; moderate meat eaters, n=60; high meat eaters, n=18). Both the ovolacto vegetarian and vegan groups demonstrated significant decreases in plasma total cholesterol (TC), low-density-lipoprotein cholesterol (LDL-C) and triacylglycerol concentrations compared with both the high-meat-eater and moderate-meat-eater groups. Total SFA and individual SFA [palmitic acid (16:0), stearic acid (18:0) and arachidic acid (20:0)] in the plasma PL were significantly lower in both the ovolacto vegetarian and vegan groups than in both the high- and moderate-meat-eater groups, while myristic acid (14:0) was significantly lower in the vegans than in the high-meat-eaters. Bivariate analysis of the results showed that the plasma PL stearic acid concentration was strongly positively correlated with plasma TC (P<0.0001), LDL-C (P<0.0001) and triacylglycerol (P<0.0001), with r(2) values of 0.655, 0.518 and 0.43 respectively. In multiple linear regression, after controlling for potential confounding factors (such as exercise, dietary group, age, body mass index, plasma PL myristic acid, palmitic acid and arachidic acid, and dietary total fat, saturated fat, cholesterol, carbohydrate and fibre intake), the plasma PL stearic acid concentration was still strongly positively correlated with plasma TC (P<0.0001) and LDL-C (P=0.006) concentrations. Based on the present data, it would seem appropriate for the population to reduce their dietary total SFA intake rather than to replace other SFA with stearic acid. PMID- 11115415 TI - Assessment of baroreflex sensitivity in patients with preserved and impaired left ventricular function by means of the Valsalva manoeuvre and the phenylephrine test. AB - The purpose of the present study was to assess the agreement between measurements of baroreflex sensitivity (BRS) obtained by the Valsalva manoeuvre and by the phenylephrine test in patients with previous myocardial infarction and different degrees of left ventricular dysfunction. Patients with a previous myocardial infarction were enrolled consecutively into two groups according to their left ventricular ejection fraction (LVEF): 40% (n=52). All patients underwent BRS assessment by the phenylephrine technique (Phe-BRS) and by the Valsalva manoeuvre, with the latter using both the overshoot part of phase IV (Ov VM-BRS) and the whole of phase IV (IV-VM-BRS). The linear association between methods was assessed by correlation analysis and the agreement was evaluated by computing the bias and the limits of agreement. IV-VM-BRS and Ov-VM-BRS could not be computed in 26% and 39% of patients respectively. For both indices a much higher percentage of non-computable Valsalva manoeuvre slopes was found in the group of patients with LVEF 40% the results were: r=0.91 (P<0.001), bias=0.1 ms/mmHg (P=0.84) and limits of agreement from -4.8 to 5 ms/mmHg. When comparing Phe-BRS and IV-VM-BRS, we found r=0.67 (P=0.001), bias=-1.5 ms/mmHg (P=0.06) and limits of agreement from -8.8 to 5.7 ms/mmHg in the group of patients with LVEF 40%. Dichotomizing Ov-VM-BRS, the best cut-off value to identify patients with a Phe BRS of <3 ms/mmHg was found to be 7 ms/mmHg, giving 100% sensitivity and 69% specificity. In conclusion, estimation of BRS by the Valsalva manoeuvre in post myocardial infarction patients is limited by a large number of non-measurable results. When computable, measurements are well correlated with those obtained by Phe-BRS, but, because of large limits of agreement, the two methods cannot be used interchangeably. If used as a screening test for risk stratification, the Valsalva manoeuvre could reduce by about one-third the need for phenylephrine injection. PMID- 11115416 TI - Low-frequency heart rate variability: reproducibility in cardiac transplant recipients and normal subjects. AB - Heart rate variability is a measure of autonomic nervous influence on the heart. It has been suggested that it could be used to detect autonomic reinnervation to the transplanted heart, but the reproducibility of the measurement is unknown. In the present study, 21 cardiac transplant recipients and 21 normal subjects were recruited. Three measurements of heart rate variability were performed during the day: in the morning, in the early afternoon and in the late afternoon. These tests were then repeated 1 week later and then again 1 week after that, making nine tests in all. The within-subject S.D. was 0.49 log units in normal subjects and 0.79 log units in transplant recipients. In both cases, this is about 15% of the population range. There was significant variation in heart rate variability between different times of day in both groups, and from day to day in transplant recipients. It was concluded that the reproducibility of measurements of heart rate variability is low, and that differences between measurements performed at different times of day should be interpreted with caution. PMID- 11115417 TI - Allometric analysis of the association between cardiac dimensions and body size variables in 464 junior athletes. AB - Empirically derived relationships between body size variables and cardiac dimensions have not been published previously for a large sample of male and female athletes. This process would inform scaling practice and facilitate intra- and inter-group comparisons of cardiac data. Therefore we investigated the relationships of body mass (BM), height and body surface area (BS) with a range of cardiac dimensions derived by echocardiography in 464 male and female elite junior athletes (age range 14-18 years; sporting allocation included rowers, cyclists, footballers, tennis players, swimmers and a miscellaneous group). Initial linearity checks suggested that most of the relationships between the body size variables and cardiac dimensions were non-linear, thus precluding the simple ratio standard approach to scaling. Multiple log-log least-squares linear regression confirmed commonality of slopes (between males and females, across the age range and between sporting groups) for all relationships involving BM and BS. Subsequent analyses of the slope exponent (b) for left ventricular dimensions supported previous data and were dimensionally consistent (LVM-BM, b=0.91+/-0.11; LVM-BS, b=1.44+/-0.19; where LVM is left ventricular mass), except for left ventricular internal dimension in diastole (LVIDd) (LVIDd-BM, b=0.25+/-0.04). Data for the left atria internal dimension (LA) were also dimensionally consistent (LA-BM, b=0.29+/-0.09); however, this was not the case for the right ventricular internal dimension in diastole (RVIDd) (RVIDd-BM, b=0.76+/-0.14). It is possible that these results were due to a study-specific limitation in the data range (LVIDd) and the geometric peculiarities of RVIDd compared with LVIDd. The gender/age/sporting groupxbody size interaction factor for virtually all relationships between height and cardiac dimensions was significant (P<0.05), and thus whole-group b exponents could not be generated. Generally these data support previous small-sample research with athletes, and suggest that allometric scaling, as opposed to simple ratio scaling, should be adopted in studies of cardiac dimensions in athletes. This should allow, with minimal mathematical difficulty, the production of body-size-independent cardiac indices to be evaluated in laboratory or clinical work. Further research is required to develop normative 'allometrically derived' cardiac indices, and care should be taken to determine relationships in specific population groups as well as to confirm commonality of slopes in multiple group comparisons. Caution is expressed regarding the use of height as a scaling variable in future research. PMID- 11115418 TI - The mechanism of resveratrol-induced vasorelaxation differs in the mesenteric resistance arteries of lean and obese rats. AB - Resveratrol has been shown to induce vasorelaxation. In this study, we investigated the mechanism(s) of resveratrol-induced vasorelaxation in resistance mesenteric arteries from male lean and dietary-induced obese rats. Compared with lean rats, arteries from dietary-obese rats showed significant (P<0.001) endothelial dysfunction, as indicated by a decrease (>20%) in maximal acetylcholine-induced vasorelaxation. Resveratrol (5-35 micromol/l) induced concentration-dependent relaxation of mesenteric arteries preconstricted with noradrenaline (8 micromol/l) or KCl (125 mmol/l) from both lean and dietary-obese rats. There were no significant differences between the two groups, achieving a maximum relaxation of >95% at a concentration of 35 micromol/l. In noradrenaline preconstricted arteries from lean rats, N(G)-nitro-L-arginine methyl ester (L NAME; 100 and 300 micromol/l) caused a significant (P<0.01) concentration dependent rightward shift in reseveratrol activity, with no effect on maximal responses. However, L-NAME (100 and 300 micromol/l) did not alter the effects of reseveratrol on arteries from dietary-obese rats, giving superimposed concentration-responses curves. Indomethacin was also ineffective in altering resveratrol activity in arteries from both lean and dietary-obese rats. In noradrenaline-precontracted arteries from dietary-obese rats, responses to resveratrol were not attenuated by endothelial denudation, indicating an action independent of the endothelium. This study indicates that: (a) the maximal effects of resveratrol on resistance arteries from lean and dietary-obese rats are not effected by endothelial dysfunction, and (b) the effects of resveratrol in lean animals (where endothelial function is not impaired), but not in dietary obese rats, are mediated via NO. PMID- 11115420 TI - Hyperhomocysteinaemia in young adults is not associated with impaired endothelial function. AB - A mild to moderate elevation of the total homocysteine concentration (tHcy) is now recognized as a risk factor for vascular disease. It is also associated with endothelial dysfunction in middle-aged and elderly individuals without overt atherosclerotic vascular disease. This is important, as endothelial dysfunction is a well recognized early and potentially reversible marker of the atherosclerotic process. We investigated whether mild hyperhomocysteinaemia was associated with endothelial dysfunction in otherwise healthy young males. We compared endothelial function, by measuring forearm blood flow, in 17 males with mild hyperhomocysteinaemia (defined as tHcy >10 micromol/l) and 14 controls with low tHcy (defined as <5 micromol/l). Forearm blood flow was measured in response to the intra-arterial infusion of acetylcholine (endothelial-dependent response) or sodium nitroprusside (endothelial-independent response). Responses to the vasoactive substances were expressed as the area under the curve of the change in forearm blood flow from baseline. Data are given as mean (95% confidence interval). The two groups were well matched for age, body mass index, pulse rate and blood pressure. tHcy was significantly different between the groups [12.3 (10.4-14.2) micromol/l compared with 4.9 (4.6-5.1) micromol/l; P<0.001]. Concentrations of vitamin B(12) and folate were significantly higher in the control group. There was no difference in basal forearm blood flow between the group with mild hyperhomocysteinaemia and the controls, and both the endothelial dependent [37.5 (26.2-38.8) and 35.3 (26.1-44.4) arbitrary units respectively] and -independent [26.1 (22.2-29.9) and 25.9 (21.0-30.8) units respectively] responses were not significantly different between the groups. Thus the present study demonstrates that, in healthy adults, mild elevation of tHcy was not associated with impaired endothelial-dependent vasodilation. These data suggest an age effect with regard to homocysteine and endothelial dysfunction. The development of vascular disease in individuals with hyperhomocysteinaemia may only result with higher concentrations or after prolonged exposure. PMID- 11115419 TI - The active molecular form of plasma adrenomedullin is extracted in the pulmonary circulation in patients with mitral stenosis: possible role of adrenomedullin in pulmonary hypertension. AB - Adrenomedullin (AM), a novel hypotensive peptide, preferentially dilates pulmonary vessels rather than systemic vessels. This suggests the possibility that AM is a circulating hormone which participates in regulation of the pulmonary circulation. A recent study revealed that two molecular forms of AM, i.e. a mature, active form of AM (AM-m) and an intermediate, inactive, glycine extended form of AM (AM-Gly), circulate in human plasma. In the present study we investigated the production and clearance sites and pathophysiological significance of the two molecular forms of AM in the pulmonary circulation in patients with mitral stenosis. We measured the plasma levels of AM-m and total AM (AM-T; AM-m+AM-Gly) using a recently developed specific immunoradiometric assay, and thus calculated plasma AM-Gly levels, in blood samples obtained from the femoral vein, pulmonary artery, left atrium and aorta of 28 consecutive patients with mitral stenosis (20 females and eight males; age 53+/-10 years). Patients with mitral stenosis had significantly higher venous concentrations of AM-T, AM Gly and AM-m than age-matched normal controls (AM-T, 15.9+/-2.5 and 10.6+/-2.1 pmol/l respectively; AM-Gly, 14.0+/-2.1 and 9.8+/-1.9 pmol/l respectively; AM-m, 1.9+/-0.6 and 1.1+/-0.3 pmol/l respectively; each P<0.001). There was a significant decrease in the concentrations of AM-m and AM-T between the pulmonary artery and the left atrium (AM-T, 16.1+/-2.7 and 14.0+/-2.4 pmol/l respectively; AM-m, 2.0+/-0.6 and 0.7+/-0.2 pmol/l respectively; each P<0.001); however, there were no differences in plasma AM-Gly levels between the pulmonary artery and the left atrium (14.1+/-2.3 and 13.5+/-2.3 pmol/l respectively). The venous concentrations of AM-m, AM-Gly and AM-T showed similar correlations with mean pulmonary artery pressure (AM-T, r=0.67; AM-Gly, r=0.63; AM-m, r=0.59; each P<0.001) and total pulmonary vascular resistance (AM-T, r=0.77; AM-Gly, r=0.70; AM-m, r=0.75; each P<0.001). These results suggest that the plasma concentration of AM-m is increased in parallel with those of AM-Gly and AM-T, and that the main site for clearance of AM-m from the plasma is the lung; the extracted AM-m in the lungs may help to attenuate the increased pulmonary arterial resistance in secondary pulmonary hypertension due to mitral stenosis. PMID- 11115421 TI - Role of aminothiols as a component of the plasma antioxidant system and relevance to homocysteine-mediated vascular disease. AB - Hyperhomocysteinaemia is considered to be an independent risk factor for vascular disease. Elevated plasma homocysteine may pose an oxidative stress, leading to the development of vascular damage. A component of this effect may be a disturbance of the extracellular aminothiol redox state. The relative contributions of plasma total homocysteine (tHcy) and plasma total cysteine (tCys) to the total antioxidant capacity (TAOC) of plasma was established in subjects with normal and elevated plasma tHcy. A total of 10 subjects with severe hyperhomocysteinaemia (due to inherited metabolic defects), 13 of their heterozygous parents and 72 normal healthy subjects were recruited to the study. The mean plasma tHcy in the patients was 91.8 micromol/l, compared with 13.2 micromol/l in the parents and 14.7 micromol/l in healthy control subjects. Plasma tCys and plasma TAOC were significantly lower in the subjects with severe hyperhomocysteinaemia compared with the parents and healthy control subjects (P<0.05). In blood samples from subjects with a normal tHcy, a positive correlation was observed between tCys and tHcy (P=0.0001). In contrast, in blood samples with tHcy >or=20 micromol/l, plasma tCys was negatively correlated with tHcy (P=0.0001). In samples with tHcy >or=20 micromol/l, tHcy was inversely correlated with TAOC (P=0.0001), whereas tCys was positively associated with TAOC (P=0.0001). Multiple regression analysis revealed that tCys was the most important independent determinant of TAOC in the patient and control groups when the effects of tHcy and several factors known to influence TAOC, such as urate, were taken into account. Thus hyperhomocysteinaemia may pose an oxidative stress not only through the direct cytotoxicity of homocysteine, but also from an associated fall in plasma cysteine. PMID- 11115422 TI - Regional transcapillary albumin exchange in rodent endotoxaemia: effects of fluid resuscitation and inhibition of nitric oxide synthase. AB - Sepsis is characterized by increased microvascular permeability and regional variations in capillary perfusion, which may be modulated by nitric oxide (NO) and reversed by fluid resuscitation (FR). The effects of saline FR and NO synthase blockade [by N(G)-nitro-L-arginine methyl ester (L-NAME)] on microvascular albumin transport and perfused capillary density were assessed in anaesthetized Wistar rats with acute normodynamic endotoxaemia. Separate dual isotope techniques were employed to measure the permeability index (PI(A)) and the permeabilityxsurface area product index (PI(B)), which provide different and complementary information regarding blood-tissue albumin exchange. PI(A) represents the tissue/blood distribution volume ratio of albumin. PI(B) is a composite measure of endothelial permeability and the vascular surface area available for albumin exchange, and therefore takes into account the effect of altered blood volume. Capillary density was quantified by fluorescence microscopy following circulation of Evans Blue-labelled albumin. Compared with controls, PI(A) was reduced significantly in lipopolysaccharide (LPS)-treated animals in skeletal muscle and skin, probably due to blood volume redistribution rather than to changes in permeability. PI(B) was increased significantly in LPS-treated animals in the kidney, mesentery, skeletal muscle, skin and lung, and in the small bowel following FR. FR also improved the LPS-induced metabolic base deficit, but did not alter capillary density. L-NAME significantly attenuated the LPS-induced rise in PI(B) in the lung. In conclusion, acute endotoxaemia induces tissue-dependent variations in microvascular albumin exchange. FR improves acid base disturbance in endotoxaemia, through mechanisms other than microvascular recruitment. NO appears to increase microvascular permeability in endotoxaemia, an effect that may be attenuated by L-NAME, particularly in the lung. PMID- 11115423 TI - Influence of age and dietary fish oil on plasma soluble adhesion molecule concentrations. AB - Soluble forms of intercellular adhesion molecule-1, vascular cell adhesion molecule-1 and E-selectin (termed sICAM-1, sVCAM-1 and sE-selectin respectively) are found in the plasma, and are elevated during inflammatory conditions in which there is increased expression of the cellular forms of the molecules on endothelial and other cells. sICAM-1, sVCAM-1 and sE-selectin concentrations were measured in the plasma of 140 healthy Caucasian subjects aged between 18 and 75 years (100 males/40 females). sICAM-1 concentrations varied between 59.9 and 299.7 ng/ml (median 150 ng/ml), sVCAM-1 concentrations varied between 222.8 and 1672.9 ng/ml (median 662 ng/ml) and sE-selectin concentrations varied between 12.4 and 90.3 ng/ml (median 45.5 ng/ml). There were significant positive linear correlations between age and the plasma concentrations of sICAM-1 (r=0.580; P<0.001) and sVCAM-1 (r=0.392; P<0.001), which were retained when the effects of gender, body mass index and fasting plasma triacylglycerol and total cholesterol concentrations were controlled for. The significant positive linear correlation between age and the plasma concentration of sE-selectin (r=0.234; P=0.027) was lost when other variables were controlled for. Male subjects <40 years of age had significantly lower plasma concentrations of both sICAM-1 and sVCAM-1 than males >55 years of age (both P<0.001), but the difference in plasma sE-selectin concentrations between the age groups did not reach significance (P=0.073). Subgroups of 16 males aged <40 years and 12 elderly subjects (>55 years of age) participated in a doubled-blind, placebo-controlled study of fish oil supplementation over 12 weeks. The level of eicosapentaenoic acid in plasma phospholipids did not change with placebo supplementation, but was significantly increased with fish oil supplementation in both young male and elderly subjects (median increase 200%). sICAM-1, sVCAM-1 and sE-selectin concentrations were unaffected by supplementation with placebo in either young male or elderly subjects. sICAM-1 concentrations were unaffected by fish oil supplementation. sE selectin concentrations were significantly increased by fish oil supplementation in young males (P=0.043; median increase 38%), but fish oil tended to decrease plasma sE-selectin concentrations in the elderly subjects (P=0.075), with a median decrease of 11%. sVCAM-1 concentrations were unaffected by fish oil supplementation in young males. Fish oil supplementation significantly decreased plasma sVCAM-1 concentrations in the elderly subjects (P=0.043), with a median decrease of 20% (range 16-60%). These observations suggest that fish oil decreases endothelial activation in elderly subjects. PMID- 11115424 TI - Protein and energy metabolism in chronic bacterial infection: studies in melioidosis. AB - Chronic infection is often accompanied by a wasting process, the metabolic basis of which is not fully understood. The aims of the present study were to measure protein and energy metabolism in patients with melioidosis (a serious and antibiotic-refractory Gram-negative bacterial infection which is endemic in South East Asia) in order to define the metabolic abnormalities that might contribute to wasting. Whole-body protein turnover was measured using the [(13)C]leucine technique, both in the fasted state and while consuming a high-energy meal. Resting energy expenditure was measured by indirect calorimetry, and total energy expenditure by the bicarbonate/urea method. Results were normalized for fat-free mass, as estimated from skinfold thickness. Protein turnover was increased in melioidosis patients compared with healthy controls during fasting (170.9 compared with 124.1 micromol x kg(-1) x h(-1); P=0.04), but the net rate of catabolism (22.2 compared with 20.5 micromol x kg(-1) x h(-1); P=0.77) and the anabolic response to feeding were similar in the two groups. Resting energy expenditure was higher in melioidosis patients compared with controls (191.4 and 157.3 kJ x kg(-1) x day(-1) respectively; P=0.04), but total energy expenditure (measured in a separate group of eight patients with melioidosis) was low (192.1 kJ x kg(-1) x day(-1)). In conclusion, this study found no evidence of metabolic causative factors, such as accelerated net protein catabolism during fasting, a blunted anabolic response to feeding or increased daily energy expenditure, and therefore suggests that reduced energy intake is the prime cause of wasting. The observed normal response to feeding should encourage nutritional approaches to prevent wasting. PMID- 11115425 TI - Homocysteine and thiol metabolites in vitamin B12 deficiency. AB - Homocysteine metabolism is increasingly implicated in a diverse group of clinical disorders, including atheromatous vascular disease. We studied the disposition of homocysteine via the trans-sulphuration pathway, plasma glutathione peroxidase (GPx) activity and plasma levels of the sulphated hormone dehydro-epiandrosterone sulphate (DHEAS) in six vitamin B(12)-deficient human subjects before and after 2 weeks of vitamin B(12) repletion, both in the fasting state and following an oral methionine load (0.1 g/kg body weight). Fasting plasma total homocysteine concentrations fell (P=0.03) and total cysteine concentrations rose significantly (P=0.048) after treatment for 2 weeks with vitamin B(12) injections. The magnitude of the mean fall in the fasting concentration of homocysteine (38.8 micromol/l) was similar to the mean rise in cysteine levels (36.0 micromol/l) following vitamin B(12) therapy. Circulating levels of homocysteine were increased at 4 h after a methionine load when compared with fasting levels, both before and after vitamin B(12) repletion (P=0.003 for both). Total cysteinyl glycine was lower post-methionine than in the fasting state following vitamin B(12) therapy (P=0.007). Fasting plasma GPx fell significantly after 2 weeks of vitamin B(12) therapy (P=0.05). The change in plasma GPx between the fasting state and 4 h after methionine loading was significantly different pre- and post vitamin B(12) therapy (P=0.05). The present study provides indirect support to the hypothesis that defects in the trans-sulphuration and remethylation of homocysteine produce hyperhomocysteinaemia in vitamin B(12) deficiency in human subjects. Elevated homocysteine levels directly or indirectly may up-regulate GPx. Sulphation status, as measured by plasma DHEAS, was unchanged. PMID- 11115426 TI - Elevation of two molecular forms of adrenomedullin in plasma and urine in patients with acute myocardial infarction treated with early coronary angioplasty. AB - Adrenomedullin (AM) has vasodilatory, diuretic and natriuretic actions. Two molecular forms of AM circulate in human plasma: an active, mature form of AM (AM m) and an intermediate, inactive, glycine-extended form of AM (AM-Gly). In the present study we investigated the pathophysiological significance of the two molecular forms of AM in plasma and urine in patients with acute myocardial infarction. We serially measured venous and arterial plasma levels and urinary excretion of AM-m, AM-Gly and total AM (Am-T; =AM-m+AM-Gly) over 2 weeks using our recently developed immunoradiometric assay in 26 consecutive patients with acute myocardial infarction and in age-matched normal controls, and studied the relationships between AM levels and clinical parameters. Plasma AM-m, AM-Gly and AM-T levels were increased on admission in patients with acute myocardial infarction compared with age-matched normal controls. Levels of AM-m, AM-Gly and AM-T in plasma reached a peak 24 h after the onset of symptoms. Plasma AM-m, AM Gly and AM-T levels were significantly correlated with plasma levels of brain natriuretic peptide and pulmonary arterial pressure. Plasma AM-Gly levels in the vein were similar to those in the artery, whereas plasma AM-m levels were significantly lower in the artery than in the vein. Urinary excretion of AM-m, AM Gly and AM-T was also increased on admission, and reached a peak at 12 h after the onset of symptoms. Urinary excretion of AM-m and AM-Gly was significantly correlated with urinary sodium excretion. The AM-m/AM-T ratio was significantly higher in the urine than in the vein or artery. AM-m levels were significantly correlated with AM-Gly levels in both the urine and plasma; however, there were no significant correlations between plasma and urinary AM levels. The results suggest that levels of both molecular forms of AM are increased in the urine as well as in the plasma in the acute phase of myocardial infarction. Since AM exerts potent cardiovascular and renal effects, increased concentrations of AM in plasma and urine in the acute phase of myocardial infarction may be involved in the defence mechanism against further elevations of peripheral and pulmonary vascular resistance and oliguria in acute myocardial infarction. PMID- 11115427 TI - Spironolactone for heart failure: spiraling out of control. PMID- 11115428 TI - Acute exacerbation of chronic bronchitis: what is the clinical significance of pathogenic bacteria in sputum cultures? PMID- 11115429 TI - Home mechanical ventilation in COPD: do we know when and how to use it? PMID- 11115430 TI - Lightning can strike twice: second primary lung cancers. PMID- 11115431 TI - New insights into the temporal pattern of hypoxemia in COPD. PMID- 11115436 TI - Does an IV bolus of methylprednisolone relieve dyspnea in asthma exacerbations? AB - STUDY OBJECTIVES: To assess whether IV methylprednisolone exerts a specific early effect on dyspnea in patients with an exacerbation of asthma. DESIGN: Randomized, placebo-controlled, double-blind crossover trial. SETTING: Medium-sized university general hospital. PATIENTS: Twenty-five asthma patients attending the chest clinic with spontaneous complaints of increases in dyspnea and with a Borg scale dyspnea rating >/= 1 at rest. INTERVENTIONS: At 0 min, IV methylprednisolone (125 mg) vs saline solution; at 60 min, 5 x 500 microg terbutaline inhaled from an inhaler device. MEASUREMENTS AND RESULTS: Change in dyspnea was assessed with bipolar visual analog scale (VAS) (much more short of breath, -100%; much less short of breath, + 100%), FEV(1), and visual memory (using the Benton visual retention test). Eighteen subjects (mean age, 61 years) completed the study. At 5 min and 60 min, shortness of breath improved with no statistically significant difference between saline solution and methylprednisolone. The mean (SD) VAS rating at 60 min was 29% (39%) on the day that saline solution was administered and 36% (25%) on the day the steroid was administered. FEV(1) and Benton score did not significantly change from baseline on either study day. Shortness of breath and FEV(1) improved following terbutaline administration, with no significant difference between the days on which saline solution and the steroid were administered. In the seven subjects who were randomized to receive methylprednisolone on the first day, baseline dyspnea rated on the Borg scale was significantly lower on the second day (first day: median, 3; range, 3 to 4; second day: median, 2; range, 0.5 to 3; p = 0.040). CONCLUSIONS: We conclude that in patients with an exacerbation of asthma, an IV bolus of methylprednisolone does not reduce dyspnea more than saline solution after 5 min and 60 min. PMID- 11115437 TI - Mometasone furoate has minimal effects on the hypothalamic-pituitary-adrenal axis when delivered at high doses. AB - STUDY OBJECTIVES: To investigate the potential for mometasone furoate (MF) to exert systemic effects following administration by dry powder inhaler (DPI) or metered-dose inhaler (MDI). DESIGN: Three randomized, evaluator-blind, placebo controlled, parallel-group, 28-day studies. PATIENTS: Adults with mild-to moderate persistent asthma. INTERVENTIONS: Study 1 (12 patients per treatment group; MF DPI at 200 microg bid, 400 microg qd, 800 microg qd, or 1,200 microg qd). Study 2 (16 patients per treatment group; MF DPI at 400 microg bid or 800 microg bid, or oral prednisone at 10 mg qd). Study 3 (16 patients per treatment group; MF MDI at 400 microg bid or 800 microg bid, or fluticasone propionate [FP] at 880 microg bid by MDI). MEASUREMENTS AND RESULTS: Study 1. Plasma concentrations were near the lower limit of quantitation (50 pg/mL) at the MF DPI 400-microg qd dosage and approximately 250 pg/mL at the 1,200-microg qd dosage. The area under the curve for serum cortisol concentrations over 24 h (AUC(24)) was essentially unaltered at all doses. Study 2. Plasma levels over days 7 to 28 were 100.3 +/- 5.9 pg/mL (mean +/- SEM) for MF DPI 400 microg bid, and 181.0 +/- 10.9 pg/mL for 800 microg bid. Although there were relatively low levels of suppression (19 to 25%) at earlier time points for MF DPI 400 microg bid, serum cortisol AUC(24) levels at day 28 were similar to placebo. MF DPI 800 microg bid and oral prednisone both decreased serum cortisol AUC(24) levels at days 7 to 28 by 28.0 +/- 8.3% and 67.2 +/- 3.6%, respectively. The response to cosyntropin was normal in 15, 14, 11, and 1 of the patients in the placebo, MF DPI 400 microg bid, MF DPI 800 microg bid, and prednisone groups, respectively. Study 3. MF MDI caused even less systemic exposure than by DPI. MF MDI 800 microg bid (24.0 +/- 3.1%) and FP (51.7 +/- 3.8%) caused a significant decrease in serum cortisol AUC(24) on days 14 to 28. MF MDI 400 microg bid was similar to placebo treatment at all time points. CONCLUSIONS: The MF 800-microg bid dosage (1,600 microg/d), which is twice the highest projected clinical dosage, represents the lower limit for consistently detectable systemic effects of MF. PMID- 11115438 TI - Rapid-onset asthma attack: a prospective cohort study about characteristics and response to emergency department treatment. AB - STUDY OBJECTIVES: (1) To determine the frequency of rapid-onset asthma attacks (ROAAs) and slow-onset asthma attacks (SOAAs) in adult patients with acute, severe disease (18 to 50 years old), who presented to an emergency department (ED); and (2) to establish whether ROAA patients differ from SOAA patients in terms of clinical and spirometric characteristics; and (3) in terms of the response of treatment. SUBJECTS AND METHODS: Four hundred three patients (with peak expiratory flow [PEF] or FEV(1) of < 50% of predicted value) with acute exacerbations of asthma were enrolled in the trial using a prospective cohort study. Asthma attacks were classified as an ROAA (< 6 h of symptoms) or an SOAA (> or = 6 h). All patients were treated with albuterol, four puffs at 10-min intervals (100 microg per actuation), delivered by metered-dose inhaler with a spacer device during 3 h. RESULTS: On the basis of previously determined criteria, 11.3% of patients were classified as having a ROAA. Male patients comprised 53.6% of the ROAA group (p = 0.03). In ROAA patients, the exacerbation was less likely to be attributed to respiratory tract infection (p = 0.001) and more likely to have no identifiable cause (p = 0.0001). Also, ROAA patients had lower pulmonary function (FEV(1)) at presentation (mean difference, - 0. 13; 95% confidence interval [CI], - 0.22 to - 0.04 L; p = 0.04) than SOAA patients. At the end of treatment, ROAA patients had an overall 48.0 L/min (95% CI, 14.1 to 81.8 L/min) greater improvement in PEF and a 0.31 L (95% CI, 0.08 to 0.54 L) greater improvement in FEV(1) than SOAA patients. Also, ROAA patients presented with less accessory muscle use (p < 0.05) and higher oxygen saturation (p = 0. 005). Finally, SOAA patients showed an increased incidence of hospital admission (relative risk, 3.89; 95% CI, 1.01 to 15.0). CONCLUSIONS: Data from this study support the notion that ROAAs constitute a distinct but uncommon acute asthma ED presentation, with a predominance of male patients. Upper respiratory tract infection was not believed to be a significant trigger factor in these patients, and ROAA patients had rapid deterioration of their conditions followed by a more rapid response to treatment and a lower hospital admission rate than SOAA patients. Thus, we have identified a subgroup of patients who appear to have common characteristics with patients with sudden-onset near-fatal/fatal asthma. PMID- 11115439 TI - Effects of esophageal acid perfusion on airway hyperresponsiveness in patients with bronchial asthma. AB - STUDY OBJECTIVES: The effects of gastroesophageal reflux on airway hyperresponsiveness in patients with bronchial asthma have yet to be studied in significant detail. The purpose of the present study was to determine how esophageal acid perfusion could change airway responsiveness in patients with bronchial asthma. PATIENTS AND INTERVENTIONS: In seven patients with bronchial asthma (mean +/- SD age, 55.1 +/- 6.4 years; four women and three men), esophageal pH was monitored by a pH meter and airway responsiveness was evaluated by aerosol inhalation of methacholine, during esophageal perfusion through an esophageal tube filled with either saline solution or 0. 1N hydrochloric acid (HCl), the order of which was selected at random, in 1-week intervals. Spirometry was also performed during esophageal pH monitoring. RESULTS: A significant decrease in the geometric mean of airway sensitivity or the concentration of methacholine causing a 35% fall in respiratory conductance was observed during esophageal HCl perfusion compared with that of saline solution perfusion (p < 0.01 or p < 0.003), although no significant changes were observed in vital capacity, FEV(1), peak expiratory flow, respiratory resistance, or slope of respiratory conductance during the periods of saline solution and HCl perfusion. CONCLUSION: We concluded that an increase in airway hyperresponsiveness was induced when HCl stimulated the esophagus in patients with bronchial asthma. These results suggest that esophageal reflux is one of the important factors that aggravate asthmatic status. PMID- 11115440 TI - Airway inflammation and etiology of acute exacerbations of chronic bronchitis. AB - STUDY OBJECTIVES: The etiologic role of bacterial pathogens isolated from sputum culture in 40 to 50% of acute exacerbations of chronic bronchitis (AECB) is controversial. If bacterial pathogens cause these AECB, they should be associated with greater neutrophilic airway inflammation than pathogen-negative exacerbations. DESIGN: This hypothesis was tested by comparing levels of interleukin (IL)-8, tumor necrosis factor (TNF)-alpha, and neutrophil elastase (NE) in 81 sputum samples obtained from 45 patients with AECB. Four groups were compared. In the first three groups, nontypable Haemophilus influenzae (n = 20), Haemophilus parainfluenzae (n = 27), and Moraxella catarrhalis (n = 14) were isolated as sole pathogens, respectively. In the fourth group, only normal flora was isolated (n = 20). Paired samples, obtained from individual patients at different times, that differed in their culture results were also compared. SETTING: An outpatient research clinic at a Veterans Affairs Medical Center. PATIENTS: These patients were participating in a prospective, longitudinal study of the dynamics of bacterial infection in chronic bronchitis, for which they were seen in the study clinic on a monthly basis as well as when they were experiencing symptoms suggestive of AECB. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: H influenzae exacerbations were associated with significantly higher sputum IL-8, TNF-alpha, and NE. M catarrhalis exacerbations demonstrated significantly higher sputum TNF-alpha and NE when compared to pathogen-negative exacerbations. H parainfluenzae-associated exacerbations had an inflammatory profile similar to pathogen-negative exacerbations. Sputum elastase level distinguished bacterial from nonbacterial AECB and correlated with clinical severity of the AECB. CONCLUSIONS: Increased airway inflammation associated with isolation of H influenzae and M catarrhalis supports an etiologic role of these pathogens in AECB. PMID- 11115441 TI - Distribution of lung density and mass in patients with emphysema as assessed by quantitative analysis of CT. AB - STUDY OBJECTIVE: To assess the effects of emphysema on the apex-to-base gradient of lung density (D) and lung mass (M) and to explore the relationship between M and lung function. METHODS: CT scans of whole lungs were performed in 12 healthy subjects and 29 patients who were breathing at functional residual capacity, after which lung function tests were performed. Whole D and M and regional D (RLD) and M (RLM) were calculated. The degree of emphysema was scored. RESULTS: The RLM for each height did not differ significantly between patients with disease and healthy subjects, while RLD was significantly lower in the patients with disease. A less marked nonlinear, increasing, craniocaudal gradient of D was observed in the group with disease, suggesting that the distension increases progressively from the apex to the base. RLD and RLM in the 40 to 90% lung height differed significantly among patients in the emphysema group with normal, high, and low M compared to the healthy subjects. M did not differ significantly between patients with centrilobular and panlobular emphysema, which was thought to stem from the marked variations in the results. Vital capacity was lower in the patients with low M. CONCLUSIONS: The lower RLD in the group with low M was due to both lung overinflation and to tissue loss, while in the groups with high or normal M, it was due only to lung overinflation. PMID- 11115442 TI - Additive effects of salmeterol and fluticasone or theophylline in COPD. AB - BACKGROUND: ss(2)-Agonists and corticosteroids or theophylline can interact to produce beneficial effects on airway function in asthma, but this has not been established in COPD. METHODS: Eighty patients with well-controlled COPD were randomized to receive 3 months of treatment in one of four treatment groups: (1) salmeterol, 50 microg bid; (2) salmeterol, 50 microg, plus fluticasone propionate, 250 microg bid; (3) salmeterol, 50 microg, plus fluticasone propionate, 500 microg bid; and (4) salmeterol, 50 microg, plus titrated theophylline bid. At each visit, a dose-response curve to inhaled salbutamol was constructed using a total cumulative dose of 800 microg. RESULTS: A gradual increase in FEV(1) was observed with each of the four treatments. Maximum significant increases in FEV(1) over baseline values that were observed after 3 months of treatment were as follows: salmeterol, 50 microg bid, 0.163 L (95% confidence interval [CI], 0.080 to 0.245 L); salmeterol, 50 microg, plus fluticasone propionate, 250 microg bid, 0.188 L (95% CI, 0.089 to 0. 287 L); salmeterol, 50 microg, plus fluticasone propionate, 500 microg bid, 0.239 L (95% CI, 0.183 to 0.296 L); and salmeterol, 50 microg, plus titrated theophylline bid, 0.157 L (95% CI, 0.027 to 0. 288 L). Salbutamol always caused a significant dose dependent increase in FEV(1) (p < 0.001), although the 800-microg dose never induced further significant benefit when compared with the 400-microg dose. The mean differences between the highest salbutamol FEV(1) after salmeterol, 50 microg, plus fluticasone propionate, 500 microg bid, and that after salmeterol, 50 microg, plus titrated theophylline bid or salmeterol, 50 microg bid, were statistically significant (p < 0.05). CONCLUSION: These data show that both long acting ss(2)-agonists and inhaled corticosteroids have a role in COPD. The data also show that fluticasone propionate and salmeterol given together are more effective than salmeterol alone. Moreover, it suggests that the addition of fluticasone propionate to salmeterol allows a greater improvement in lung function after salbutamol, although regular salmeterol is able to improve lung function in COPD patients without development of a true subsensitivity to its bronchodilator effect. In any case, patients must be treated for at least 3 months before a real improvement in lung function is achieved. PMID- 11115443 TI - Long-term controlled trial of nocturnal nasal positive pressure ventilation in patients with severe COPD. AB - STUDY OBJECTIVES: To determine the 1-year efficacy of noninvasive positive pressure ventilation (NPPV) added to long-term oxygen therapy (LTOT) in patients with stable severe COPD. PATIENT SELECTION AND METHODS: We prospectively randomized 52 patients with severe COPD (FEV(1) < 45%) to either NPPV plus "standard care" (96% patients with LTOT) or to standard care alone (93% patients with LTOT). The outcomes measured included the following: rate of acute COPD exacerbations; hospital admissions; intubations; and mortality at 3 months, 6 months, and 12 months. The patients were also evaluated at 3 months and 6 months for dyspnea using the Medical Research Council and Borg scales, gas exchange, hematocrit, pulmonary function, cardiac function with echocardiogram, and neuropsychological performance. RESULTS: One-year survival was similar in both groups (78%). The number of acute exacerbations was similar at all time points in patients receiving NPPV, compared with control subjects. The number of hospital admissions was decreased at 3 months in the NPPV group (5% vs 15% of patients, p < 0.05), but this difference was not seen at 6 months (18% vs 19%, respectively). The only beneficial differences were observed in the Borg dyspnea rating, which dropped from 6 to 5 (p < 0.039), and in one of the neuropsychological tests (psychomotor coordination) for the NPPV group at 6 months. CONCLUSIONS: Our study indicates that over 1 year, NPPV does not affect the natural course of the disease and is of marginal benefit in outpatients with severe COPD who are in stable condition. PMID- 11115444 TI - Genotype analysis and phenotypic manifestations of children with intermediate sweat chloride test results. AB - STUDY OBJECTIVES: Cystic fibrosis (CF) is one of the most common inherited diseases among whites. Since the cloning of the CF transmembrane conductance regulator (CFTR) gene, a number of studies have focused on associations between the genotype and phenotype in CF. This had led to the progressive identification of new groups of patients, including those who have mild lung disease and those who have normal sweat chloride values (< 60 mEq/L). The aim of the present work was to provide information on the genotype and the phenotypic characteristics of children with intermediate-range sweat chloride test results. PATIENTS AND RESULTS: We focused on children referred to the pulmonary department for various types of pulmonary disease and who had several sweat chloride test results with median values in the range of 40 to 60 mEq/L. Twenty-four patients over a 10-year period were enrolled (mean age, 4.8 years). Respiratory manifestations at initial evaluation included recurrent bronchitis, wheezing, chronic cough, and pneumonia. The duration of the follow-up ranged from 0.5 to 10.5 years. Sputum cultures revealed the presence of Haemophilus influenzae (10 children), Staphylococcus aureus (4 children), and Pseudomonas aeruginosa (3 children). Pancreatic insufficiency was found in two patients. Analysis of the entire coding sequence allowed identification of 16 known mutations in CFTR gene. Fifteen chromosomes (31.2%) carried a mutation in CFTR gene and one allele carried two mutations. Three patients were homozygous or double heterozygous (DeltaF508/DeltaF508, DeltaF508/3849 + 10 kb C-->T, S1235R/G551D). The 5-thymidine allele was identified in four children. CONCLUSION: These results indicate an higher frequency of CFTR gene mutations in patients with borderline sweat chloride test results, compared to data reported in the general population. They lead to the recommendations for complete pulmonary and GI investigations in this group of patients, as well as assiduous care and medical follow-up. PMID- 11115445 TI - Complications of indwelling catheters in cystic fibrosis: a 10-year review. AB - STUDY OBJECTIVE: Patients with cystic fibrosis (CF) frequently require recurrent courses of IV antibiotics to treat acute exacerbations of their pulmonary disease. Over time, CF patients often lose peripheral access, and indwelling central venous catheters are placed. We attempted to determine the type and incidence of catheter complications so that CF patients could be fully informed of the risks prior to placement of these catheters. DESIGN: The charts of all CF patients who attended the Adult Cystic Fibrosis Clinic of the University of Washington Medical Center from January 1989 through December 1998 were reviewed. Demographic information was obtained along with the type and duration of catheter, type and number of complications, and the use of anticoagulant medication. MEASUREMENTS AND RESULTS: Of the 218 CF patients who attended the clinic, 65 patients (30%) had indwelling catheters in place at some time during the study period. A total of 87 catheters were placed into these 65 patients. The total number of catheter-days for first indwelling catheters was 68,220. The total number of catheter-days for all catheters was 75,660 (210 catheter-years). Thirty-five catheter-related complications were identified, occurring in 26 patients. Complications included thrombosis (n = 14), infections (n = 9), mechanical problems (n = 6), pneumothorax (n = 3), superior vena cava syndrome/stenosis (n = 2), and air embolism (n = 1), for an overall complication rate of 0. 463/1,000 catheter-days. CONCLUSION: We conclude that indwelling catheters are relatively safe in patients with CF. Good infection control policies appear to prevent most infectious complications. The most common complication is that of thrombosis, which may be recurrent in some patients. Consideration should be given to prophylactic warfarin therapy despite the potential risk of significant hemoptysis in this patient population. PMID- 11115446 TI - Lung cancer in women: sex-associated differences in survival of patients undergoing resection for lung cancer. AB - STUDY OBJECTIVES: The aim of this study was to analyze various characteristics and survival in female patients treated surgically for lung cancer. DESIGN: Retrospective clinical study. PATIENTS: From 1,242 consecutive cases of primary non-small cell lung cancer treated with pulmonary resection between June 1984 and December 1998, 337 female patients (27.1%) were chosen. RESULTS: Female patients had the following characteristics: a significantly younger age at onset (62.5 +/- 0.56 years vs 64.1 +/- 0.31 years for men), a higher frequency of adenocarcinoma (86.0% vs 48.3% for men), and smaller tumors (32.7 mm vs 38.3 mm in diameter for men). Peripheral tumors were significantly more common in women than men (71.8% vs 50.6%, respectively). Among 686 patients with a history of smoking, the women smoked significantly less often (12.8% vs 91.4% for men). Complete resection was achieved significantly less often in women (79.6% vs 85.2% for men); however, women having complete resection survived significantly longer than their male counterparts. Women with a postoperative negative carcinoembryonic antigen (CEA) had a significantly better prognosis than men; however, women with a postoperative positive CEA did not. Women > or = 60 years old survived significantly longer than their male counterparts, while women < 60 years old did not. CONCLUSIONS: Once the tumor was resected completely, women survived longer, partly due to the influence of life expectancy. However, the incidence of malignant effusion was higher and the rate of complete resection was lower in women. PMID- 11115447 TI - The usefulness of positron emission tomography in evaluating patients for pulmonary malignancies. AB - STUDY OBJECTIVE: Positron emission tomography (PET) can contribute to diagnosing and staging lung cancer, but it has not been determined whether this information influences patient care. DESIGN: We reviewed the effects of thoracic PET scan results during an 11-month period. For each patient, physicians ordering these scans reported how PET specifically altered management, and graded the ease of interpretation and overall usefulness of PET on a 5-point scale. In addition, to appraise general attitudes about PET, we surveyed 488 national American Thoracic Society (ATS) members and 44 physicians at our comprehensive cancer center. RESULTS: One hundred twenty-six questionnaires regarding patients were mailed to 37 ordering physicians, and 98 responses (78%) were returned, primarily by cardiothoracic surgeons (35%) and pulmonologists (47%). Respondents reported that PET provided new information in 83 patients (85%) and altered patient management in 64 cases (65%). Major effects on management included decisions regarding biopsy (n = 16), surgery (n = 16), and palliative treatment (n = 16). Chest clinicians found PET to be more helpful (4.4 vs 3.9, p = 0.007) and easier to interpret (4.2 vs 3.7, p = 0.025) than other specialists. Among 139 ATS members (28%) responding to the general survey, 51 members (39%) had access to PET. PET was more frequently available to university-based (49%) than community-based (27%) physicians (p = 0.016). The majority of physicians without current access to PET (69%) indicated that they would like to have it available. ATS members with access to PET reported that PET results generally affect decisions regarding biopsy or surgery most often, but found the procedure less helpful than physicians at our center (2.77 vs 3. 56, p = 0.003) and ordered it less often for lung cancer staging (60% vs 96%, p = 0.002). CONCLUSION: PET scanning is useful in the management of patients with suspected thoracic malignancies, but impressions about its roles vary, with PET regarded more highly where, as at our center, it is used more often. Whether PET alters patient outcomes requires investigation. PMID- 11115448 TI - Use of intraoperative hetastarch priming during coronary bypass. AB - BACKGROUND: The use of hetastarch during coronary bypass surgery has been limited due to its unresolved potential risk for hemorrhage. Therefore, the purpose of this study was to investigate the effects of using 6% hetastarch in priming cardiopulmonary bypass (CPB) circuitry on the need for blood product transfusions and outcome after coronary bypass. MATERIALS AND METHODS: This nonrandomized retrospective study involved 887 patients who underwent isolated primary coronary artery bypass grafting. Based on the type of solution used in priming the CPB circuitry, patients were stratified into the following four different groups: group 1, crystalloid (500 mL; n = 211); group 2, 25% human albumin (50 mL; n = 217); group 3, 6% hetastarch (500 mL; n = 298); and group 4, 25% human albumin (50 mL) and 6% hetastarch (500 mL; n = 161). Patient characteristics and clinical variables were compared among the groups using the Kruskal-Wallis test. Patient survival estimates were compared using log-rank test. RESULTS: Demographic patient characteristics for all groups were similar (p > 0.05). Intraoperative and perioperative variables among groups were comparable (p > 0.05). The use of hetastarch as a part of prime solution in CPB circuitry did not alter the need for banked blood, platelets, or fresh frozen plasma transfusions (p > 0.05). The length of stay in the ICU or in the hospital was unaffected in all groups. The early (ie, 30-day) mortality rate was 1.4% in group 1, 1.8% in group 2, 1.0% in group 3, and 3.1% in group 4. Long-term survival among the groups was unaffected by the type of priming solution. CONCLUSIONS: The use of hetastarch in priming CPB circuitry is devoid of any added hemorrhagic risk after coronary bypass, and the type of prime solution for CPB has no influence on the early or late survival rates of patients undergoing primary coronary bypass. PMID- 11115449 TI - Surgery for second lung cancers. AB - PURPOSE: To evaluate the outcomes of patients surgically treated for their second primary lung cancer. METHOD: In a computerized surgical registry of > 800 consecutive patients treated for primary pulmonary carcinoma since 1980, 37 patients presented with a second lung cancer. These patients were analyzed regarding their original treatment, preoperative evaluation, operative procedures, and long-term follow-up. RESULTS: Three fifths of the patients were female, and 57% were > or = 65 years old at the time of their second operation. One patient originally had two synchronous tumors; another patient had three metachronous neoplasms. The interval between surgeries ranged from 5 to 239 months. In 31 patients, treatment for their original tumor was surgical resection alone. Lobectomy was the most common operation for the original tumor, and 78% were stage I. When the second tumor was diagnosed, 25 patients (68%) were asymptomatic. Eight patients (22%) were current smokers, and 29 patients (78%) were former smokers. The most common operation for the second tumor was a lobectomy. Surgical mortality was 5.4%. Nineteen patients (51%) survived 2 years, and 9 patients (24%) survived > or = 5 years. Eleven patients (30%) were still alive at last follow-up, 3 to 198 months postoperatively, and only 13 patients (34%) had died of their cancer. CONCLUSION: Surgical treatment of second primary pulmonary neoplasms can be performed in selected patients with acceptable long term survival. PMID- 11115450 TI - Measuring pleural fluid pH: high correlation of a handheld unit to a traditional tabletop blood gas analyzer. AB - STUDY PURPOSES: To survey hospital laboratories in the United States to determine methods used for measuring pleural fluid pH, and to compare pleural fluid pH values obtained with a traditional tabletop blood gas analyzer (BGA) to those obtained with a handheld analyzer. METHODS: Hospital laboratories nationwide were contacted by telephone to survey the methods used to measure pleural fluid pH. In a second phase, pleural fluid was prospectively collected from 19 pediatric and adult patients with pleural effusions, and pleural fluid pH was measured simultaneously with a traditional tabletop BGA and with a handheld unit. RESULTS: A total of 220 hospital laboratories were contacted by telephone, and 166 responded (75%). The methods for determining pleural fluid pH for all hospital laboratories were pH meter (35%; n = 59), BGA (32%; n = 53), and litmus paper (31%: n = 51); 2% (n = 3) did not perform the test. University hospitals were more likely to use a BGA, compared to community hospitals (p < 0.014) or children's hospitals (p < 0.001). In the comparison of pleural fluid measurements, the mean pH for the traditional BGA was 7.358 +/- 0.189, and the mean pH for the handheld unit was 7.382 +/- 0.203. The absolute difference between the two machines was 0.024 U, and the two methods were correlated (p < 0.01; r = 0.993; degrees of freedom = 36). CONCLUSION: Most hospital laboratories in the United States do not measure pleural fluid pH using a traditional BGA and use alternative methods that have previously been shown to be inaccurate. Pleural fluid pH obtained by a handheld unit has a high degree of correlation to that of a traditional tabletop BGA, and it offers a satisfactory alternative for laboratories reluctant to measure pleural fluid pH with a BGA. PMID- 11115451 TI - Transbronchial needle aspiration: guidance with CT fluoroscopy. AB - BACKGROUND: Bronchoscopy with transbronchial needle aspiration (TBNA) is valuable to diagnose lesions in the mediastinum and lung, but conventional fluoroscopic guidance may be suboptimal. We describe the use of CT fluoroscopy to provide real time, transaxial TBNA localization, thus facilitating biopsy. METHODS: Patients were selected because of prior unsuccessful bronchoscopy or anticipated difficulty owing to small size or inaccessibility of the lesion. CT fluoroscopy consists of a spiral CT scanner adapted using a rapid-reconstruction algorithm and hardware that permits real-time in-room imaging. The bronchoscope was inserted on the CT scanner, which was used to guide TBNA instruments into the target lesion. RESULTS: Of 27 patients who underwent TBNA with CT fluoroscopic assistance, 15 had mediastinal nodes, and 12 had lung nodules or focal infiltrates. Mean lesion size was 1.7 cm in the mediastinum, 2. 2 cm in the lung. A correct diagnosis was established in 10 of 12 mediastinal lesions (83%) for which follow-up was available and in 8 lung lesions (67%). Diagnoses included small cell and non-small cell lung cancer and invasive aspergillosis. False negative results were caused by sampling errors or inability to reach the lesion as documented by CT fluoroscopy. Postprocedure CT fluoroscopy revealed no complications. CONCLUSION: CT fluoroscopy provides effective, real-time guidance for TBNA and may be particularly valuable in patients with small or less accessible mediastinal or lung lesions. PMID- 11115452 TI - Rise and fall of the FEV(1). AB - BACKGROUND: Most studies of the rate of decline in ventilatory capacity in normal subjects take into account a relatively restricted number of factors, such as age, smoking, and dust exposure. There is increasing evidence to suggest that such a limited approach is inadequate. OBJECTIVE: To carry out a prospective study of those factors influencing the rate of decline of the ventilatory capacity in a cohort of automobile workers. DESIGN: Prospective cohort study. SETTING: Southern Ontario, Canada. PARTICIPANTS: A cohort of 181 workers employed in assembling and spray painting the chassis of new cars, a minority of whom used paints containing isocyanates. MEASUREMENTS: All participants underwent annual anthropometric measurements. Spirometry was carried out at yearly intervals, and a questionnaire relating to respiratory symptoms and smoking habits was completed annually by all participants. Daily monitoring of the isocyanate levels was carried out. RESULTS: There was no indication of any effect from isocyanate exposure. The annual decline in the FEV(1) was similar to that found in other studies, with the respective annual decrements for smokers, ex-smokers, and nonsmokers being 0.055 L, 0.046 L, and 0.035 L, respectively. The decline of the FEV(1) in those > 35 years old and < 35 years old differed appreciably. The decrements in the FEV(1) in subjects < 35 years old were influenced as much by excessive weight gain as by cigarette smoking. Loss of weight in those significantly overweight was frequently associated with improved lung function. CONCLUSIONS: While age and smoking play an important role in determining the rate of decline in the ventilatory capacity, it is clear that body weight plays a significant role and needs to be taken into account in all epidemiologic studies of the ventilatory capacity. PMID- 11115453 TI - CT in pulmonary hydatid disease: unusual appearances. AB - OBJECTIVE: To study the CT features of pulmonary hydatid disease. PATIENTS: Thirty-two consecutive patients with surgically proven pulmonary hydatid cysts. SETTING: SheriKashmir Institute of Medical Sciences, Srinagar, Kashmir, India, a tertiary-care referral center. INTERVENTIONS: CT of the chest was obtained in all cases on Somatom DR double rotate CT scanner (Siemens; Erlangen, Germany). RESULTS: Forty cysts of different size and shapes were encountered, 34 of them being ruptured. CT density of the cysts varied from - 42 to 160 Hounsfield units (HU; median, 15.5 HU). Apart from the classically described features of pulmonary hydatid disease, a crescent-shaped rim of air at the lower end of the cyst (inverse crescent sign) was seen in three cysts, and a bleb of air in the wall of two as-yet unruptured cysts (signet ring sign). Thick wall (>10 mm) was observed in four cysts, and each of them had associated evidence of infection. CONCLUSIONS: Inverse crescent sign, signet ring sign, high CT density, and thick wall should be recognized as features of pulmonary hydatid cysts on CT. PMID- 11115454 TI - Bronchiolitis obliterans syndrome and additional costs of lung transplantation. AB - STUDY OBJECTIVES: The influence of bronchiolitis obliterans syndrome (BOS) on costs after lung transplantation was investigated by comparing the costs of patients with and without this condition. DESIGN: Follow-up costs were prospectively investigated in a medical technology assessment of the Dutch Lung Transplant Program, in relation to the development of the BOS. First, average follow-up costs per week per patient were compared between patients who did or did not develop BOS. Second, in the BOS group, these costs were compared before and after the onset of BOS. SETTING: Dutch Lung Transplant Program, University Hospital of Groningen. RESULTS: Data on 53 patients (37 patients without BOS and 16 with BOS) who underwent transplantation between November 1990 and April 1995 were available. The average follow-up time of these 53 patients was 1.5 years. The follow-up costs amounted to an average (in Dutch guilders [Dfl]) of 1,774/wk for non-BOS patients, compared to 3,072/wk for BOS patients (+ 73%; p = 0.002; one Dfl = 50 cents US currency). This difference in costs was largely accounted for by an increase in used health-care resources, in particular hospitalization and medication. For the BOS patients, the average costs per week before and after the onset of BOS were 1,941 Dfl and 2,422 Dfl, respectively. CONCLUSION: BOS is associated with substantial extra costs. These findings reemphasize the need to focus efforts on prevention of BOS to enhance the cost-effectiveness of lung transplantation. PMID- 11115455 TI - Quantification of cytomegalovirus DNA in BAL fluid: a longitudinal study in lung transplant recipients. AB - STUDY OBJECTIVES: Cytomegalovirus (CMV) infection is common in patients receiving solid organ transplants, and it is associated with increased morbidity as well as risk for development of chronic rejection. A rapid and sensitive diagnostic method would improve the therapeutic management of CMV infection, including the monitoring of treatment effects. We investigated whether longitudinal determinations of CMV DNA quantities in BAL fluid could be useful for this purpose. DESIGN: CMV DNA levels in 340 BAL samples from 35 consecutive lung transplant recipients were studied during a median of 18 months. Seventeen (49%) of the patients developed CMV disease with pneumonitis. Twenty-seven CMV disease episodes were diagnosed. RESULTS: Patients with CMV disease had a significantly higher mean level of CMV copies per milliliter BAL fluid (1,120 +/- 4,379) compared with those without (180 +/- 1,177, p < 0.01). Viral load as well as acute rejection requiring treatment (>/= A2) were independent risk factors associated with CMV disease. Differences between the groups concerning HLA-DR matching, basic immunosuppressive therapy, and CMV serologic status D/R -/+ vs D/R +/+ were not significant. A diagnostic definition of normality based on the mean level of all episodes without CMV disease +2 SD would discriminate only 9 of the 27 CMV episodes. CONCLUSIONS: Although the viral load is increased during episodes of clinical CMV disease in lung transplant recipients, the quantitative PCR assessment of CMV DNA in BAL fluid is not discriminative enough to be useful as a diagnostic tool for CMV disease. PMID- 11115456 TI - Exercise intolerance following heart transplantation: the role of pulmonary diffusing capacity impairment. AB - STUDY OBJECTIVES: Although impairment of the diffusing capacity of the lung for carbon monoxide (DLCO) in heart transplant recipients is well-documented, there are limited data on its impact on exercise capacity in these patients. The aim of this study was to determine the effect of DLCO reduction on exercise capacity in heart transplant recipients. DESIGN: Descriptive cohort study. SETTING: A regional cardiopulmonary transplant center. PARTICIPANTS: Twenty-six heart transplant recipients who were studied before and after transplantation compared with 26 healthy volunteers. MEASUREMENTS: Spirometry and static lung volumes were measured using body plethysmography, DLCO was measured using the single-breath technique, and progressive cardiopulmonary exercise was performed using a bicycle ergometer, continuous transcutaneous blood gas monitoring, and on-line analysis of minute ventilation, oxygen uptake (VO(2)), and carbon dioxide production. RESULTS: Before transplantation, the mean percent predicted for hemoglobin corrected DLCO was reduced in patients (73.2%) compared to healthy control subjects (98.8%; p < 0.001) and declined significantly after transplantation (60.1%; p < 0.05). Although the mean maximal symptom-limited VO(2) (VO(2)max) increased after transplantation (increase, 41.3 to 48.6% of predicted; p < 0.05), it remained substantially lower than normal (92.9%; p < 0.001). There was a significant correlation between DLCO and VO(2)max after transplantation (r = 0.61; p = 0.001), but not before transplantation (r = 0.09; p = 0.66). DLCO was also inversely correlated with other respiratory responses to exercise, including the following: the ventilatory response to exercise (r = -0.44; p < 0.05); dead space to tidal volume ratio (r = -43; p < 0.05); and the alveolar-arterial oxygen gradient (r = -0. 45; p < 0.05), but there was no correlation between any of these variables and DLCO before transplantation. CONCLUSION: DLCO reduction after heart transplantation appears to represent persistent gas exchange impairment and contributes to exercise limitation in heart transplant recipients. PMID- 11115457 TI - Pulmonary veno-occlusive disease: a case series and new observations. AB - STUDY OBJECTIVES: The aim of this study was to describe our experience at one institution with pulmonary veno-occlusive disease (PVOD) during the past 10 years, with particular reference to new findings and long-term outcome. SETTING: Tertiary care, academic medical center. PATIENTS AND METHODS: Eleven patients who were evaluated and treated for PVOD at our institution were retrospectively studied. Included were all available clinical, radiographic, hemodynamic, and pathologic data. RESULTS: All 11 patients in our series had at least one symptom or clinical finding that, in conjunction with known pulmonary hypertension, suggested the diagnosis of PVOD. Digital clubbing, not previously reported in PVOD, was found in 5 patients, rales in 6, and increased interstitial markings on chest radiograph in 10. Half of the 10 patients who underwent acute vasodilator testing exhibited a decrease in pulmonary artery pressure of > 20%, although one patient died shortly after receiving IV calcium-channel blockers. Three patients have demonstrated sustained clinical improvement with therapy, which includes calcium-channel blockers, epoprostenol, and lung transplantation in one patient each. However, outcome was generally poor, with a 72% mortality within 1 year of diagnosis. CONCLUSION: The diagnosis of PVOD requires a high clinical suspicion. However, both physical examination findings and radiographic studies often provide clues to the diagnosis, which may obviate the need for lung biopsy in the majority of cases. Although there may be patients who respond to medical therapy, the use of vasoactive medications in patients with PVOD should be undertaken with great caution. Long-term survival is poor, and lung transplantation remains the only proven therapy. PMID- 11115458 TI - New onset of venous thromboembolism among hospitalized patients at Brigham and Women's Hospital is caused more often by prophylaxis failure than by withholding treatment. AB - CONTEXT: Guidelines to prevent venous thromboembolism (VTE) have been widely distributed and generally have been assumed to be effective. Therefore, among hospitalized patients, the development of VTE is thought to occur in the context of omitted prophylaxis. OBJECTIVES: To describe hospitalized patients who develop VTE and to determine whether they received antecedent prophylaxis. DESIGN: Case series. SETTING: Brigham and Women's Hospital. PATIENTS: Three hundred eighty four patients who developed in-hospital deep venous thrombosis or pulmonary embolism or who developed VTE within 30 days of prior hospital discharge. MAIN OUTCOME MEASURES: The relationship of developing new-onset VTE to the use or omission of antecedent in-hospital prophylaxis. RESULTS: Of the 384 identified patients, 272 had deep venous thrombosis alone, 62 had pulmonary embolism alone, and 50 had deep venous thrombosis and pulmonary embolism. Most were medical service patients; fewer than one fourth were general or orthopedic surgery patients. Overall, 52% had received antecedent VTE prophylaxis. Thirteen deaths (3.4%) were ascribed to pulmonary embolism, and prophylaxis was omitted in only 1 of those 13 patients. CONCLUSIONS: Most deaths from pulmonary embolism among patients hospitalized for other conditions occurred in the setting of failed prophylaxis rather than omitted prophylaxis. High-risk patients, especially medical service patients, warrant intensive VTE prophylaxis and close follow-up to ensure successful outcomes. PMID- 11115459 TI - Plasma homocysteine and severity of thoracic aortic atherosclerosis. AB - STUDY OBJECTIVES: Plasma homocysteine level is a risk factor for coronary events, stroke, and peripheral atherosclerotic disease. However, few data are available concerning the relationship between homocysteine level and severity of thoracic aortic atherosclerosis. We hypothesized in this multiplane transesophageal echocardiography (TEE) study that homocysteine level is a marker of the presence and severity of thoracic aortic atherosclerosis. DESIGN: Cross-sectional study. SETTING: University hospital. PATIENTS: Risk factors, angiographic features, and TEE findings were analyzed prospectively in 82 valvular patients. MEASUREMENTS AND RESULTS: The following risk factors were recorded: age, gender, hypertension, smoking, lipid parameters, diabetes, body mass index, and family history of coronary artery disease. Plasma levels of homocysteine, vitamin B(12), and folic acid were measured for each patient. By univariate analysis, age, diabetes, hypertension, smoking, family history of coronary artery disease, and levels of homocysteine, total cholesterol, low-density lipoprotein cholesterol, and high density lipoprotein cholesterol were significant predictors of the presence of thoracic aortic plaques. There was a positive correlation between the plasma homocysteine levels and the score of severity of thoracic atherosclerosis (r = 0.48; p = 0.0001) as well as between the homocysteine levels and the grades of severity of aortic intimal changes (p = 0.0008). Multivariate regression analysis revealed that homocysteine was an independent predictor of the presence and severity of thoracic aortic atherosclerosis. CONCLUSION: This prospective study indicates that plasma homocysteine level is a marker of severity of thoracic atherosclerosis detected by multiplane TEE. These findings emphasize the role of homocysteine as a marker of atherosclerotic lesions in the major arterial locations. PMID- 11115460 TI - The preventive effect of magnesium on coronary spasm in patients with vasospastic angina. AB - STUDY OBJECTIVES: Previous studies have reported that magnesium (Mg) deficiency is associated with coronary spasm. However, little is known about the preventive effect of Mg on coronary spasm. The present study investigated whether Mg prevents coronary spasm in patients with vasospastic angina (VSA). DESIGN: Effectiveness trial. SETTING: University medical center. PATIENTS: Twenty-two patients with VSA. INTERVENTION: Coronary spasm was induced with an intracoronary infusion of acetylcholine (Ach). After spontaneous relief of the coronary spasm, Mg sulfate (0.27 mmol/kg body weight) was infused IV over 20 min in 14 patients and isotonic glucose was infused in 8 patients as control subjects. Intracoronary infusion of Ach was then repeated, and the diameter of the coronary arteries was measured quantitatively. MEASUREMENTS AND RESULTS: Mg infusion caused coronary artery dilatation at baseline in both the spastic (5. 9 +/- 2.3%) and nonspastic segments (5.5 +/- 1.5%). Mg infusion reduced the severity of chest pain and ST segment deviations during coronary spasm. After the Mg infusion, the percent change in the diameter of the spastic segments improved from - 62.8 +/- 2.6% to - 43.7 +/- 4.7% during coronary spasm. Overall, 10 of 14 patients (71%) responded favorably to Mg infusion. Isotonic glucose infusion did not elicit changes in chest pain severity, ST-segment deviations, or the diameter of the coronary arteries during spasm. CONCLUSIONS: Mg infusion produces nonsite-specific basal coronary dilatation and suppresses Ach-induced coronary spasm in patients with VSA. PMID- 11115461 TI - Prognostic significance of left ventricular aneurysms with normal global function caused by myocarditis. AB - OBJECTIVES: To evaluate the prognosis of left ventricular (LV) aneurysms with normal global function caused by myocarditis. BACKGROUND: LV aneurysms may result from idiopathic or viral myocarditis. The prognosis of inflammatory LV aneurysms when associated with a normal cardiac function is unknown. METHODS: Among 353 patients with a histologic diagnosis of myocarditis, 12 (3.3%) had single or multiple localized LV aneurysms (length, 10.6 +/- 3.1 mm; width, 7.4 +/- 4.2 mm) with normal cardiac function. Presenting symptoms were ventricular tachycardia (VT) in nine patients and unexplained chest pain in three. All patients underwent laboratory tests and noninvasive and invasive cardiac examinations, including biventricular endomyocardial biopsy. RESULTS: In all patients, LV endomyocardial biopsy specimen showed a lymphocytic myocarditis with focal intense myocytolysis or damage of intramural vessels, whereas right ventricular biopsy was diagnostic for myocarditis only in three. Serologic study suggested a viral infection in 3 patients and an immunologic disorder in 2, although it was negative in 7. Treatment included antiarrhythmics in 9 patients with VT, ss-blockers in 1 with chest pain, and immunosuppression (prednisone and azathioprine for 5 months) in 4 with active myocarditis (2 with chest pain and 2 with VT). At intermediate-term follow-up (mean, 53 months; range, 12 to 120 months), LV function was persistently normal in all patients, with an LV aneurysm occlusion being observed in two patients. All patients were asymptomatic, with no VT recurrence or major clinical events. None required implantable electrical devices or a surgical intervention. CONCLUSIONS: LV aneurysms with normal global function caused by myocarditis are an uncommon benign entity in which major therapeutic regimens are usually unnecessary. PMID- 11115462 TI - Retrograde flow in the thoracic aorta in patients with systemic emboli: a transesophageal echocardiographic evaluation of mobile plaque motion. AB - STUDY OBJECTIVES: Blood flow in the aorta is complex and incompletely characterized. Mobile aortic plaques (MAPs), moving freely with the pulsatile aortic flow, in fact represent natural tracers that reflect the flow pattern itself. Our aim was to use MAP motion on transesophageal echocardiography (TEE) in order to characterize flow patterns in the atheromatous thoracic aorta of patients with systemic emboli. DESIGN AND PATIENTS: The study group was recruited from 250 patients referred for TEE to evaluate recent embolism. Among them, 22 patients (14 men and 8 women; mean +/- SD age, 66.3 +/- 7.2 years; 16 patients with cerebrovascular and 6 patients with peripheral emboli) with MAPs of > or = 3 mm in length formed the study group. The longest amplitudes of three spatial components of mobile lesion motions were measured: x (antegrade/retrograde [A/R]), y (up/down [U/D], and z (right/left [R/L]). RESULTS: A total of 33 mobile lesions were detected: 3 in the ascending aorta (1 patient), 13 in the arch (10 patients), and 17 in the descending aorta (11 patients). The length of mobile plaque components ranged from 3 to 13 mm; amplitudes of A/R, U/d, R/L, and retrograde flow motions ranged from 3 to 26 mm, from 1 to 16 mm, from 1 to 17 mm, and from 1 to 13 mm, respectively. Systolic rotational motion was clockwise in six patients (27%), counterclockwise in five patients (23%), incomplete (semicircle) in six patients (27%), and alternate clockwise/counterclockwise in five patients (23%). Diastolic rotational motion was clockwise in 5 patients (23%), counterclockwise in 6 patients (27%), and incomplete (semicircle) in 11 patients (50%). There were 18 multiple MAPs in seven patients: in all these cases, simultaneous rotations of MAP in different directions (as a marker for the presence of multiple vortices) were found. In nine patients with cerebral embolism, MAPs on the distal part of aortic arch solely were found; in five of them, all alternative potential sources of stroke were excluded. Therefore, retrograde cerebral embolism from distal aortic plaques in these patients is highly probable. CONCLUSIONS: Retrograde and rotational blood flow in the thoracic aorta probably exists in all patients with systemic emboli and mobile protruding aortic atheromas. Therefore, retrograde cerebral embolism from distal aortic plaques is theoretically possible. PMID- 11115463 TI - Impact of noninvasive studies to distinguish volume overload from ARDS in acutely ill patients with pulmonary edema: analysis of the medical literature from 1966 to 1998. AB - STUDY OBJECTIVE: To assess the impact of substituting noninvasive diagnostic studies for Swan-Ganz catheter (SGC) placement in the evaluation of acutely ill patients. DESIGN: Modified decision analysis. METHODS: Using published studies that define effectiveness of clinical examination, echocardiography, and SGC placement to diagnose pulmonary edema, an analysis of the impact of substituting three diagnostic approaches using (1) clinical assessment (CA), (2) M-mode two dimensional transthoracic echocardiography (EC), or (3) CA then EC if necessary for SGC placement was considered. STUDY POPULATION: Patients with acute respiratory distress and radiographic findings of pulmonary edema, and ICU patients with hypotension and/or pulmonary edema without acute cardiac ischemia. INTERVENTIONS: Three approaches using noninvasive studies were substituted for placement of SGC in the initial evaluation of pulmonary edema. MEASUREMENTS AND RESULTS: The number of SGCs placed, the number of tests needed to diagnose (NTND) all cases of volume overload, and the total number of procedure-related adverse events were calculated for each diagnostic approach and compared to SGC placement. EC, and CA then EC approaches produced fewer procedure-related serious complications and deaths, compared to the SGC approach; however, these approaches also produced a higher NTND and total procedures performed than did the SGC or CA approaches. The CA approach led to reduced NTND and procedure-related adverse events. CONCLUSIONS: Substituting noninvasive studies for SGC placement in the initial evaluation of acutely ill patients may slightly reduce procedure-related adverse events, but it may also increase the number of procedures performed. Studies of SGC use are warranted and need to include a clinical assessment control group and an analysis of resource utilization. PMID- 11115464 TI - Echocardiographic evaluation of left ventricular function in critically ill patients: dynamic loading challenge using medical antishock trousers. AB - STUDY OBJECTIVE: We hypothesized that a dynamic left ventricular (LV) evaluation during a loading challenge might enhance diagnostic capabilities of routine transesophageal echocardiography in critically ill patients and selection of therapeutic options against circulatory failure, particularly the choice between volume expansion and vasoactive agent infusion. DESIGN: Prospective clinical study in a group of 26 patients requiring hemodynamic support by vasoactive infusion because of low systemic arterial pressure (< 90 mm Hg by invasive monitoring) during mechanical ventilation. SETTING: University hospital ICU. PATIENTS: Patients required respiratory support for an episode of acute respiratory failure of various causes or for an episode of coma. They were studied by transesophageal echocardiography during mechanical ventilation in the controlled mode, before and during a loading challenge made using the legs compartment of medical antishock trousers inflated at 80 mm Hg. MEASUREMENTS: A short-axis view of the left ventricle was obtained by a transgastric approach, and end-diastolic and end-systolic areas were measured. LV stroke area (LVSA) and LV fractional area contraction (LVFAC) were calculated. RESULTS: Changes in LV echocardiographic measurements permitted separation of the patients into two groups. In nine patients (group 1), LVSA, used as an index of stroke output, was significantly increased during the challenge, together with a significant increase in LV end-diastolic area, suggesting preload improvement by the challenge. Conversely, in 17 patients (group 2), LVSA was significantly reduced by the challenge, together with a significant decrease in LVFAC, suggesting a negative effect of increased afterload by the challenge. CONCLUSION: Study of the changes in LV dimensions during loading challenge in hemodynamically unstable patients was used to evaluate the balance between the adequacy of preload and the ability of the heart to pump against an increased load, and might thus guide hemodynamic support. PMID- 11115465 TI - Morbid results of prolonged intubation after coronary artery bypass surgery. AB - OBJECTIVES: This study evaluated the morbid results of prolonged intubation after coronary artery bypass grafting (CABG). METHODS: Over 30 months, 66 of 1,112 patients undergoing CABG required prolonged intubation. They were matched with 66 patients who did not require prolonged intubation. Preoperative and operative variables were evaluated to determine which would predict prolonged intubation. The postoperative courses were then compared to evaluate the effect of prolonged intubation. The study population was divided into three groups: those who underwent early extubation, but required reintubation (n = 24); those who required initial prolonged intubation, but no reintubation (n = 22); and those who required initial prolonged intubation and reintubation (n = 20). RESULTS: Univariate analysis revealed unstable angina (p = 0.037), elevated creatinine (p = 0.001), reduced FEV(1) (p = 0.019), longer cardiopulmonary bypass time (p = 0.009), and a greater positive fluid balance at 24 h (p = 0.0001) as predictors of postoperative prolonged intubation. Multivariate regression analysis revealed elevated creatinine (p = 0.011), FEV(1) (p = 0.022), and fluid balance (p = 0.001) as predictors of prolonged intubation. The study population had longer ICU and hospital stays (p = 0.0001), with more infectious complications (p = 0.0001) and higher mortality (p = 0. 001). In the subgroups of the study population, patients not requiring reintubation had shorter ICU (p = 0.001) and hospital stays (p = 0.0001), fewer infectious complications (p = 0.0001), and reduced mortality (p = 0.0001). CONCLUSIONS: Patients undergoing CABG with reduced FEV(1), renal failure, and positive fluid balance 24 h postoperatively are at risk for prolonged intubation. Prolonged intubation results in significant acute and midterm morbidity and mortality. Early extubation followed by reintubation further increases morbidity and mortality rates in these patients. PMID- 11115466 TI - Performance of APACHE III models in an Australian ICU. AB - STUDY OBJECTIVE: Evaluation of the performance of the APACHE (acute physiology and chronic health evaluation) III ICU and hospital mortality models at an Australian tertiary adult ICU. DESIGN: Noninterventional, observational study. SETTING: Metropolitan, Australian, tertiary referral medical/surgical ICU. PATIENTS: A total of 3,398 consecutive eligible admissions from January 1, 1995, to December 31, 1997. MEASUREMENTS: Prospective collection of demographic, diagnostic, physiologic, laboratory, admission, and discharge data. RESULTS: The patient sample was younger and more commonly male, with more comorbidities and a different operative and referral source mix, compared to the APACHE III development sample. Receiver operating characteristic curve areas for ICU (0.92) and hospital mortality (0.90) demonstrated excellent discrimination. Observed ICU mortality (9.9%) did not significantly differ from the prediction of the APACHE III model (8.9%) or the APACHE III model adjusted for hospital characteristics (10.5%). The hospital mortality (16.0%) was underestimated by the APACHE III model [13.6%; chi(2)(1) = 7.4; p = 0.01]. With proprietary adjustments for hospital characteristics (14.9%) or referenced to the US database (15.6%), agreement was closer. Good calibration was found with all models except the unadjusted hospital mortality model. CONCLUSION: In contrast to other non American studies, this Australian study demonstrates that the APACHE III can perform well on independent assessment. As perfect discrimination and calibration cannot coexist in a probabilistic model with dichotomous outcomes, performance of APACHE III models with proprietary adjustment for hospital characteristic provide a good compromise for use in quality surveillance. PMID- 11115467 TI - Impact of BAL in the management of pneumonia with treatment failure: positivity of BAL culture under antibiotic therapy. AB - BACKGROUND: Pneumonia is responsible for 50% of antibiotics prescribed in ICUs. Treatment failure, ie, absence of improvement or clinical deterioration under antibiotic therapy, presents a dilemma to physicians. BAL is an invasive method validated for etiologic diagnosis in pneumonia. STUDY OBJECTIVE: To evaluate in ICU patients the impact of BAL in the etiologic diagnosis, treatment, and outcome of pneumonia with treatment failure. DESIGN: Prospective clinical study. SETTING: Nonsurgical, medical ICU of a university hospital in Brazil. PATIENTS AND PARTICIPANTS: Sixty-two episodes of pneumonia treated for at least 72 h without clinical improvement in 53 patients hospitalized for diverse clinical emergencies. Mean duration of hospitalization was 14.2 days. Mean duration of previous antibiotic therapy was 11.4 days. INTERVENTIONS: Bronchoscopy and BAL were performed in each episode. BAL fluid was cultivated for aerobic and anaerobic bacteria; the cutoff considered positive was 10(4) cfu/mL; 10(3) cfu/mL was also analyzed if under treatment. Pneumocystis carinii, fungi, Legionella spp, and Mycobacterium spp were also researched. MEASUREMENTS AND RESULTS: Fifty eight of 62 BAL were performed under antibiotics. The results showed positivity in 45 of 62 (72.6%); 42 of the 45 positive episodes (93.3%) had > 10(4) cfu/mL. The three cases with between 10(3) and 10(4) cfu/mL were considered positive and were treated according to BAL cultures. The main agents were Acinetobacter baumannii (37.1%), Pseudomonas aeruginosa (17.7%), and methicillin-resistant Staphylococcus aureus (MRSA; 16.1%); 46.7% of the episodes (21 of 45) were polymicrobial. BAL results directed a change of therapy in 34 episodes (54.8%). Overall mortality was 43.5%. There was no difference in mortality among positives, negatives, and patients who changed therapy guided by BAL culture. CONCLUSIONS: (1) BAL fluid examination was positive in 45 of 62 episodes (72.6%), with 58 of 62 BAL performed under antibiotics. This suggests that BAL may be a sensitive diagnostic method for treatment failures of clinically diagnosed pneumonias, even if performed under antibiotics; (2) the main pathogens in our study were A baumannii, P aeruginosa, and MRSA, and approximately 45% of infections were polymicrobial; (3) BAL culture results directed a change of therapy in 75.6% of positive episodes (34 of 45) and in 54.8% of all episodes of treatment failure (34 of 62); and (4) there was no difference in mortality among positives, negatives, and patients who changed therapy guided by BAL culture. PMID- 11115468 TI - Vascular endothelial growth factor level correlates with transforming growth factor-beta isoform levels in pleural effusions. AB - BACKGROUND: Recent studies have demonstrated high levels of vascular endothelial growth factor (VEGF) in exudative pleural effusions and a possible etiologic role. The factors regulating VEGF accumulation in the pleural space are unknown. Transforming growth factor (TGF)-beta is a potent stimulator of VEGF expression in vitro. We hypothesized that TGF-beta induces VEGF production in pleural tissues, and, hence, the pleural fluid VEGF levels should correlate with the levels of TGF-beta in pleural fluid of different etiologies. METHODS: Seventy pleural fluid samples were analyzed. These included 20 malignant, 13 post coronary artery bypass grafting (CABG), 8 parapneumonic, 11 miscellaneous exudative, and 18 congestive heart failure (CHF) pleural effusions. RESULTS: Pleural fluid VEGF levels showed good correlation with those of TGF-beta(1) (r = 0.58; p < 0. 0001), TGF-beta(2) (r = 0.43; p < 0.001), and lactate dehydrogenase (r = 0.65; p < 0.001). The levels of TGF-beta(1) and TGF-beta(2) also were correlated (r = 0.60; p < 0.0001). The median levels of TGF-beta(1) (2,480 pg/mL) and TGF-beta(2) (266 pg/mL) in the CHF group were significantly lower than those in the malignant (TGF-beta(1), 4,902 pg/mL; TGF-beta(2), 428 pg/mL), post-CABG (TGF-beta(1), 5,456 pg/mL; TGF-beta(2), 377 pg/mL), parapneumonic (TGF-beta(1), 5,024 pg/mL; TGF-beta(2), 464 pg/mL), and miscellaneous exudate groups (TGF beta(1), 7,690 pg/mL; TGF-beta(2), 369 pg/mL). There was no significant difference in TGF-beta(1) and TGF-beta(2) levels among the four exudate groups. CONCLUSIONS: VEGF levels in pleural effusions are significantly correlated with the levels of TGF-beta(1) and beta(2) isoforms. VEGF, TGF-beta(1), and TGF beta(2) levels were all higher in exudative effusions than in effusions secondary to CHF. PMID- 11115469 TI - Elevated levels of soluble adhesion molecules in sera and BAL fluid of individuals infected with human T-cell lymphotropic virus type 1. AB - STUDY OBJECTIVE: T-lymphocytic alveolitis and increased levels of interleukin-2 receptor-alpha (CD25)-bearing T cells in the BAL fluid (BALF) of human T-cell lymphotropic virus type 1 (HTLV-1) carriers have been reported. Several chemokines and adhesion molecules may contribute to the accumulation of T lymphocytes in the lungs of HTLV-1 carriers. To clarify the correlation between adhesion molecules and HTLV-1-associated pulmonary disorders, we compared the distribution of T-lymphocyte subsets and soluble adhesion molecules, including soluble intercellular adhesion molecule (sICAM)-1, soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble L-selectin (sL-selectin), soluble E-selectin (sE selectin), and soluble P-selectin (sP-selectin), in BALF and peripheral blood, between HTLV-1 carriers and noninfected healthy subjects. DESIGN: Flow cytometric analysis with monoclonal antibodies to cell-surface antigens was used to identify T-lymphocyte subsets in BALF samples from HTLV-1 carriers (n = 13) and noninfected healthy control subjects (n = 10). The levels of various soluble adhesion molecules in serum and in BALF were estimated by enzyme-linked immunosorbent assay. RESULTS: Higher percentages of CD3+ cells, CD3-expressing human leukocyte antigen-DR antigen, and CD3+CD25+ cells were detected in the BALF of HTLV-1 carriers than in that of noninfected control subjects. The concentrations of sICAM-1, sVCAM-1, sL-selectin, SE- selectin, and sP-selectin in the sera of patients were significantly higher than those in noninfected healthy control subjects. The concentration of sICAM-1 in the BALF of patients was significantly higher than that in noninfected healthy control subjects, and the concentration of sICAM-1 correlated well with the percentage of CD3+CD25+ cells. CONCLUSION: The concentrations of adhesion molecules in the sera of and sICAM-1 in the BALF of HTLV-1 carriers were significantly higher than those in noninfected individuals, and the concentration of sICAM-1 correlated well with the percentage of CD3+CD25+ cells in BALF. Our results suggest a possible interaction between activated T cells bearing CD25 and soluble adhesion molecules, especially sICAM-1, which may contribute to the pulmonary involvement in HTLV-1 carriers. PMID- 11115470 TI - Comparison of titanium vascular closure staples with suture repair of the thoracic aorta in swine. AB - OBJECTIVE: Devices that reduce technical difficulty and anastigmatic time when repairing large vessels such as the thoracic aorta would be beneficial. The aim of this study was to determine if titanium vascular closure staples (3 mm) could be safely and quickly applied in the repair of large vessels such as the thoracic aorta. DESIGN: Through a left thoracotomy in 10 female swine (110 to 130 lb), an interposition graft (14 to 16 mm textile) was placed into the aorta distal to the left subclavian artery. Animals were randomized at the time of repair to either running sutures (n = 5; 6-0 polypropylene) or vascular closure staples (n = 5; 3 mm). The anastomosis was evaluated after 2 months with aortograms, and the aorta was harvested to evaluate healing. RESULTS: The clamp times (mean +/- SD) were 30.8 +/- 8.2 min for suture repair and 24.8 +/- 5.1 min for vascular closure staple repair (p = 0.2). Anastomosis times were 20. 0 +/- 6.2 min for the suture group and 16.4 +/- 6.4 min for the vascular closure staple group (p = 0.4). Arch aortograms at 2 months revealed no significant difference in luminal narrowing between the two groups. Gross and microscopic examination revealed no thrombosis, well-healed wounds with a continuous intimal layer, and no differences in intimal thickness or inflammation between the two groups. CONCLUSION: Vascular closure staples were equivalent to sutures in terms of durability, graft patency, and wound healing at 2 months. Vascular closure staples may offer the trauma surgeon a quick and easy alternative when repairing large vessels such as the thoracic aorta. PMID- 11115471 TI - Clinical conference on management dilemmas: a growing vascular mass in the chest. PMID- 11115472 TI - Early detection of lung cancer with laser-induced fluorescence endoscopy and spectrofluorometry. AB - STUDY OBJECTIVES: We performed a clinical trial of laser-induced fluorescence endoscopy (LIFE) for detection of precancerous lesions and cancer including carcinoma in situ (CIS), which are difficult to detect by white-light bronchoscopy. DESIGN: Results with LIFE were compared with the criterion standard, white-light bronchoscopy. The evaluation of these endoscopic results spectrofluorometrically was examined, and pixels of LIFE images composed of digital signals for the intensities of red and green were analyzed. SETTING: Tertiary-level hospital treating referrals and subjects with suspicious results in mass screening. PATIENTS: We examined 65 subjects with suspected lung cancer by both methods, and performed biopsy on 216 lesions. RESULTS: The accuracy of diagnosis by white-light bronchoscopy, with histopathologic results as the standard, was 48.6%. The accuracy by LIFE was 72.7%. The sensitivity of conventional bronchoscopy for detection of severe dysplasia (21 biopsy specimens) or cancer (28 biopsy specimens) was 61.2% and specificity was 85.0%. With results by LIFE added, these values were 89.8% and 78.4%, respectively. Of nine patients with CIS, only LIFE showed one lesion, and only LIFE showed the extent of seven of the lesions. The autofluorescence of eight lesions was measured spectrofluorometrically; normal bronchial tissue, severe dysplasia, and cancerous tissue had spectral differences. The red/green intensity of cancers on histograms of LIFE images generally was greater than the ratios for metaplasia or normal bronchial wall. CONCLUSIONS: Use of both methods should facilitate early detection. Evaluation by spectrofluorometry and analysis of digital signal intensity of results by LIFE make results more objective. PMID- 11115473 TI - Three-dimensional CT-guided bronchoscopy with a real-time electromagnetic position sensor: a comparison of two image registration methods. AB - STUDY OBJECTIVES: To compare two different image registration methods for accurately displaying the position of a flexible bronchoscope on a previously acquired three-dimensional CT scan during bronchoscopy. SETTING: Bronchoscopy suite of a university hospital. PATIENTS: Fifteen adult patients scheduled for nonemergent bronchoscopy. METHODS: A miniature electromagnetic position sensor was placed at the tip of a flexible bronchoscope. Previously acquired three dimensional CT scans were registered with the patient in the bronchoscopy suite. Registration method 1 used multiple skin fiducial markers. Registration method 2 used the inner surface of the trachea itself for registration. Method 1 was objectively assessed by measuring the error distance between the real skin marker position and the computer display position. Methods 1 and 2 were subjectively assessed by the bronchoscopist correlating visual bronchoscopic anatomic location with the computer display position on the CT image. RESULTS: The error distance (+/- SD) from known points for registration method 1 was 5.6 +/- 2.7 mm. Objective error distances were not measured for method 2 because no accurate placement of the bronchoscope sensor could be correlated with CT position. Subjectively, method 2 was judged more accurate than method 1 when compared with the fiberoptic view of the airways through the bronchoscope. Additionally, method 2 had the advantage of not requiring placement of fiducial markers before the CT scan. Respiratory motion contributed an error of 3.6 +/- 2.6 mm, which was partially compensated for by a second tracking sensor placed on the patient's chest. CONCLUSION: Image registration method 2 of surface fitting the trachea rather than method 1 of fiducial markers was subjectively judged to be superior for registering the position of a flexible bronchoscope during bronchoscopy. Method 2 was also more practical inasmuch as no special CT scanning technique was required before bronchoscopy. PMID- 11115474 TI - Jugular wave recognition breakthrough: X' descent vs the X descent and trough. AB - Irrational nomenclature and concentrating on ascents and peaks of waves have made recognition of jugular waves an occult art. By agreeing to call atrial relaxation X: and the systolic fall in atrial pressure due to the descent of the base X', we can begin to teach the easy recognition of jugular contours. Next, it is necessary to realize that the artifacts seen on electronically derived jugular pulse tracings are not to be expected when observing the neck pulsations with the naked eye. Finally, it can be shown that the easiest way to recognize jugular waves is by timing only descents as being either systolic or diastolic according to their relation to either the patient's radial pulse or heart sounds. It is almost unknown that only a single systolic descent due to the descent of the base is usual in the normal adult jugular. PMID- 11115475 TI - Cardiac transplant vasculopathy. AB - STUDY OBJECTIVE: Coronary allograft vasculopathy (CAV) remains the major factor limiting long-term survival after heart transplantation. The purpose of this article is to review for the nontransplant physician the concept of CAV as a disease entity after heart transplantation. DESIGN: A MEDLINE search from 1985 to 1999 was performed. Data on cardiac transplant vasculopathy were divided into pathology, pathophysiology, presentation, diagnosis, and treatment. RESULTS: CAV manifests as a unique and unusually aggressive form of coronary artery disease that differs from traditional atherosclerosis. It is believed to be caused by immunologic mechanisms that combine with nonimmunologic factors to cause endothelial injury, resulting in smooth muscle proliferation and intimal thickening. This intimal hyperplasia leads to coronary obstruction, which ultimately results in allograft failure. Diagnosis of CAV can be difficult because transplant recipients have denervated hearts and rarely present with chest pain. Various noninvasive screening methods have not proved reliable. Therefore, most transplant centers perform periodic coronary angiography for routine CAV surveillance. Treatment of CAV involves modification of risk factors and the use of pharmacologic agents that alter vascular physiology. Revascularization procedures continue to play a role as palliative therapy, but are limited in their effectiveness by the diffuse nature of this disease. CONCLUSION: Cardiac transplant vasculopathy continues to play a major dilemma regarding posttransplant care. Further research is needed to develop successful preventive and therapeutic strategies that may alter the course of this disease. PMID- 11115476 TI - Physiotherapy in intensive care: towards an evidence-based practice. PMID- 11115478 TI - Abnormal finding in an overnight sleep study. PMID- 11115477 TI - Upper lobe fibrocavitary disease in a patient with back pain and stiffness. PMID- 11115479 TI - Biventricular cardiac pacing: promising new therapy for congestive heart failure. PMID- 11115480 TI - A 45-year-old man with slowly progressive shortness of breath. PMID- 11115481 TI - Long-term clarithromycin decreases prednisone requirements in elderly patients with prednisone-dependent asthma. AB - Prolonged use of prednisone is associated with serious side effects, such as osteoporosis, particularly among elderly individuals. Macrolide antibiotics exhibit anti-inflammatory effects that are distinct from their antimicrobial properties. Thus, the purpose of this case report is to describe the effects of prolonged treatment with clarithromycin, 500 mg bid, in reducing prednisone requirements in three elderly patients with prednisone-dependent asthma. Three patients (one woman and two men) aged 63 to 69 years, who had been treated with 5 to 10 mg prednisone daily for at least the last 12 months, were given clarithromycin, 500 mg bid. They were followed regularly for changes in daily prednisone dose, spirometry, quality of life, and adverse events. The prednisone dose was tapered in a stepwise fashion at each clinic visit. Within 3 to 6 months of initiation of treatment with clarithromycin, and throughout the 12-month follow-up, two of three patients discontinued prednisone therapy, while the third patient displayed increased spirometry readings and noted an increasingly better quality of life. Pulmonary function tests were stable or improved over this time period, with no reported adverse events, including increased rate of infections. One patient relapsed upon discontinuation of clarithromycin therapy but has since responded to re-initiation of treatment. Long-term oral clarithromycin may have a role in reducing prednisone requirements in elderly patients with prednisone dependent asthma. PMID- 11115482 TI - Intracardiac mass complicating Malassezia furfur fungemia. AB - Malassezia furfur is a lipophilic yeast known to colonize indwelling catheters. Although progression to vasculitis and sepsis has been described, it has rarely caused fungemia in adults receiving nutrition via an indwelling catheter. Difficulty in diagnosis occurs as M furfur does not grow on routine culture media unless it is supplemented with fatty acids. We present the first case of M furfur fungemia in an adult, complicated by a pedunculated septic thrombus arising from the superior vena cava and extending into the right atrium. Removal of the catheter, amphotericin-B therapy, and surgical debridement were required for cure. PMID- 11115483 TI - Successful use of argon plasma coagulation and tranilast to treat granulation tissue obstructing the airway after tracheal anastomosis. AB - We successfully used argon plasma coagulation (APC) and tranilast to treat granulation tissue that had formed on an end-to-end tracheal anastomosis. APC had several advantages over laser in the management of exuberant granulation tissue, and we considered tranilast to be effective in our patient. To our knowledge, this is the first description of the use of APC in the management of anastomotic granulation tissue in the trachea or bronchus. PMID- 11115484 TI - Staphylococcus aureus pericarditis masquerading as anterior mediastinal mass: mediastinal mass from pericarditis. AB - Pseudomediastinal mass as a result of bacterial pericarditis is a rare clinical presentation. We report one such case in a patient with end-stage renal disease receiving hemodialysis, who presented primarily with manifestations of right heart compression due to a large encapsulated pericardial abscess and, surprisingly, with no overt signs of sepsis. Surgical drainage, pericardiectomy, and antibiotic therapy led to a successful outcome. PMID- 11115485 TI - Tension pneumothorax and contralateral presumed pneumothorax from endobronchial intubation via cricothyroidotomy. AB - Cricothyroidotomy can be a life-saving procedure for the "can't intubate, can't ventilate" patient who has upper-airway obstruction. The procedure is usually fast and easy to do; however, complications have been reported. We report two cases in which cricothyroidotomy with an endotracheal tube led to unrecognized endobronchial intubation, ipsilateral tension pneumothorax, contralateral presumed pneumothorax, and unnecessary emergency surgery. Additionally, these led to the triad of hypotension, hypoxemia, and, probably, elevated intracranial pressure, which can worsen cerebral injury. We discuss methods to avoid these complications. PMID- 11115486 TI - Cardiac tamponade following acupuncture. PMID- 11115487 TI - Clinical diagnosis of stage I sarcoidosis. PMID- 11115488 TI - Endothelial apoptosis. PMID- 11115489 TI - Fatal pneumococcal pneumonia attributed to macrolide resistance and azithromycin monotherapy. PMID- 11115490 TI - Supranormal expiratory flow rates in patients with interstitial lung disease. PMID- 11115494 TI - Aberrant signal peptide cleavage of collagen X in Schmid metaphyseal chondrodysplasia. Implications for the molecular basis of the disease. AB - Schmid metaphyseal chondrodysplasia results from mutations in the collagen X (COL10A1) gene. With the exception of two cases, the known mutations are clustered in the C-terminal nonhelical (NC1) domain of the collagen X. In vitro and cell culture studies have shown that the NC1 mutations result in impaired collagen X trimer assembly and secretion. In the two other cases, missense mutations that alter Gly(18) at the -1 position of the putative signal peptide cleavage site were identified (Ikegawa, S., Nakamura, K., Nagano, A., Haga, N., and Nakamura, Y. (1997) Hum. Mutat. 9, 131-135). To study their impact on collagen X biosynthesis using in vitro cell-free translation in the presence of microsomes, and cell transfection assays, these two mutations were created in COL10A1 by site-directed mutagenesis. The data suggest that translocation of the mutant pre-alpha1(X) chains into the microsomes is not affected, but cleavage of the signal peptide is inhibited, and the mutant chains remain anchored to the membrane of microsomes. Cell-free translation and transfection studies in cells showed that the mutant chains associate into trimers but cannot form a triple helix. The combined effect of both the lack of signal peptide cleavage and helical configuration is impaired secretion. Thus, despite the different nature of the NC1 and signal peptide mutations in collagen X, both result in impaired collagen X secretion, probably followed by intracellular retention and degradation of mutant chains, and causing the Schmid metaphyseal chondrodysplasia phenotype. PMID- 11115495 TI - Oncogenes induce and activate endogenous p73 protein. AB - The identification of upstream pathways that signal to TP73 is crucial for understanding the biological role of this gene. Since some evidence suggests that TP73 might play a role in tumorigenesis, we asked whether oncogenes can induce and activate endogenous TP73. Here, we show that endogenous p73 alpha and beta proteins are up-regulated in p53-deficient tumor cells in response to overexpressed E2F1, c-Myc, and E1A. E2F1, c-Myc, and E1A-mediated p73 up regulation leads to activation of the p73 transcription function, as shown by p73 responsive reporter activity and by induction of known endogenous p73 target gene products such as p21 and HDM2. Importantly, E2F1-, c-Myc-, and E1A-mediated activation of endogenous p73 induces apoptosis in SaOs-2 cells. Conversely, inactivation of p73 by a dominant negative p73 inhibitor (p73DD), but not by a mutant p73DD, inhibits oncogene-induced apoptosis. These data show that oncogenes can signal to TP73 in vivo. Moreover, in the absence of p53, oncogenes may enlist p73 to induce apoptosis in tumor cells. PMID- 11115496 TI - Identification of internal autoproteolytic cleavage sites within the prosegments of recombinant procathepsin B and procathepsin S. Contribution of a plausible unimolecular autoproteolytic event for the processing of zymogens belonging to the papain family. AB - The steps involved in the maturation of proenzymes belonging to the papain family of cysteine proteases have been difficult to characterize. Intermolecular processing at or near the pro/mature junction, due either to the catalytic activity of active enzyme or to exogeneous proteases, has been well documented for this family of proenzymes. In addition, kinetic studies are suggestive of a slow unimolecular mechanism of autoactivation which is independent of proenzyme concentration. However, inspection of the recently determined x-ray crystal structures does not support this evidence. This is due primarily to the extensive distances between the catalytic thiolate-imidazolium ion pair and the putative site of proteolysis near the pro/mature junction required to form mature protein. Furthermore, the prosegments for this family of precursors have been shown to bind through the substrate binding clefts in a direction opposite to that expected for natural substrates. We report, using cystatin C- and N-terminal sequencing, the identification of autoproteolytic intermediates of processing in vitro for purified recombinant procathepsin B and procathepsin S. Inspection of the x-ray crystal structures reported to date indicates that these reactions occur within a segment of the proregion which binds through the substrate binding clefts of the enzymes, thus suggesting that these reactions are occurring as unimolecular processes. PMID- 11115497 TI - Functional relationships between capacitation-dependent cell signaling and compartmentalized metabolic pathways in murine spermatozoa. AB - Spermatozoa are highly polarized cells with specific metabolic pathways compartmentalized in different regions. Previously, we hypothesized that glycolysis is organized in the fibrous sheath of the flagellum to provide ATP to dynein ATPases that generate motility and to protein kinases that regulate motility. Although a recent report suggested that glucose is not essential for murine sperm capacitation, we demonstrated that glucose (but not lactate or pyruvate) was necessary and sufficient to support the protein tyrosine phosphorylation events associated with capacitation. The effect of glucose on this signaling pathway was downstream of cAMP, and appeared to arise indirectly as a consequence of metabolism as opposed to a direct signaling effect. Moreover, the phosphorylation events were not affected by uncouplers of oxidative respiration, inhibitors of electron transfer, or by a lack of substrates for oxidative respiration in the medium. Further experiments aimed at identifying potential regulators of sperm glycolysis focused on a germ cell-specific isoform of hexokinase, HK1-SC, which localizes to the fibrous sheath. HK1-SC activity and biochemical localization did not change during sperm capacitation, suggesting that glycolysis in sperm is regulated either at the level of substrate availability or by downstream enzymes. These data support the hypothesis that ATP specifically produced by a compartmentalized glycolytic pathway in the principal piece of the flagellum, as opposed to ATP generated by mitochondria in the mid piece, is strictly required for protein tyrosine phosphorylation events that take place during sperm capacitation. The relationship between these pathways suggests that spermatozoa offer a model system for the study of integration of compartmentalized metabolic and signaling pathways. PMID- 11115498 TI - Rescue of contractile parameters and myocyte hypertrophy in calsequestrin overexpressing myocardium by phospholamban ablation. AB - Cardiac-specific overexpression of murine cardiac calsequestrin results in depressed cardiac contractile parameters, low Ca(2+)-induced Ca(2+) release from sarcoplasmic reticulum (SR) and cardiac hypertrophy in transgenic mice. To test the hypothesis that inhibition of phospholamban activity may rescue some of these phenotypic alterations, the calsequestrin overexpressing mice were cross-bred with phospholamban-knockout mice. Phospholamban ablation in calsequestrin overexpressing mice led to reversal of the depressed cardiac contractile parameters in Langendorff-perfused hearts or in vivo. This was associated with increases of SR Ca(2+) storage, assessed by caffeine-induced Na(+)-Ca(2+) exchanger currents. The inactivation time of the L-type Ca(2+) current (I(Ca)), which has an inverse correlation with Ca(2+)-induced SR Ca(2+) release, and the relation between the peak current density and half-inactivation time were also normalized, indicating a restoration in the ability of I(Ca) to trigger SR Ca(2+) release. The prolonged action potentials in calsequestrin overexpressing cardiomyocytes also reversed to normal upon phospholamban ablation. Furthermore, ablation of phospholamban restored the expression levels of atrial natriuretic factor and alpha-skeletal actin mRNA as well as ventricular myocyte size. These results indicate that attenuation of phospholamban function may prevent or overcome functional and remodeling defects in hypertrophied hearts. PMID- 11115501 TI - Hydrogen peroxide-induced structural alterations of RNAse A. AB - Proteins exposed to oxidative stress are degraded via proteolytic pathways. In the present study, we undertook a series of in vitro experiments to establish a correlation between the structural changes induced by mild oxidation of the model protein RNase A and the proteolytic rate found upon exposure of the modified protein toward the isolated 20 S proteasome. Fourier transform infrared spectroscopy was used as a structure-sensitive probe. We report here strong experimental evidence for oxidation-induced conformational rearrangements of the model protein RNase A and, at the same time, for covalent modifications of amino acid side chains. Oxidation-related conformational changes, induced by H(2)O(2) exposure of the protein may be monitored in the amide I region, which is sensitive to changes in protein secondary structure. A comparison of the time- and H(2)O(2) concentration-dependent changes in the amide I region demonstrates a high degree of similarity to spectral alterations typical for temperature-induced unfolding of RNase A. In addition, spectral parameters of amino acid side chain marker bands (Tyr, Asp) revealed evidence for covalent modifications. Proteasome digestion measurements on oxidized RNase A revealed a specific time and H(2)O(2) concentration dependence; at low initial concentration of the oxidant, the RNase A turnover rate increases with incubation time and concentration. Based on these experimental findings, a correlation between structural alterations detected upon RNase A oxidation and proteolytic rates of RNase A is established, and possible mechanisms of the proteasome recognition process of oxidatively damaged proteins are discussed. PMID- 11115499 TI - Substrate and metal complexes of 3-deoxy-D-manno-octulosonate-8-phosphate synthase from Aquifex aeolicus at 1.9-A resolution. Implications for the condensation mechanism. AB - 3-Deoxy-D-manno-octulosonate-8-phosphate synthase (KDO8PS) from the hyperthermophilic bacterium Aquifex aeolicus differs from its Escherichia coli counterpart in the requirement of a divalent metal for activity (Duewel, H. S., and Woodard, R. W. (2000) J. Biol. Chem. 275, 22824-22831). Here we report the crystal structure of the A. aeolicus enzyme, which was determined by molecular replacement using E. coli KDO8PS as a model. The structures of the metal-free and Cd(2+) forms of the enzyme were determined in the uncomplexed state and in complex with various combinations of phosphoenolpyruvate (PEP), arabinose 5 phosphate (A5P), and erythrose 4-phosphate (E4P). Like the E. coli enzyme, A. aeolicus KDO8PS is a homotetramer containing four distinct active sites at the interface between subunits. The active site cavity is open in the substrate-free enzyme or when either A5P alone or PEP alone binds, and becomes isolated from the aqueous phase when both PEP and A5P (or E4P) bind together. In the presence of metal, the enzyme is asymmetric and appears to alternate catalysis between the active sites located on one face of the tetramer and those located on the other face. In the absence of metal, the asymmetry is lost. Details of the active site that may be important for catalysis are visible at the high resolution achieved in these structures. Most notably, the shape of the PEP-binding pocket forces PEP to assume a distorted geometry at C-2, which might anticipate the conversion from sp(2) to sp(3) hybridization occurring during intermediate formation and which may modulate PEP reactivity toward A5P. Two water molecules are located in van der Waals contact with the si and re sides of C-2(PEP), respectively. Abstraction of a proton from either of these water molecules by a protein group is expected to elicit a nucleophilic attack of the resulting hydroxide ion on the nearby C 2(PEP), thus triggering the beginning of the catalytic cycle. PMID- 11115502 TI - Testing the versatility of the sarcoplasmic reticulum Ca(2+)-ATPase reaction cycle when p-nitrophenyl phosphate is the substrate. AB - A detailed characterization of p-nitrophenyl phosphate as energy-donor substrate for the sarcoplasmic reticulum Ca(2+)-ATPase was undertaken in this study. The fact that p-nitrophenyl phosphate can be hydrolyzed in the presence or absence of Ca(2+) by the purified enzyme is consistent with the observed phenomenon of intramolecular uncoupling. Under the most favorable conditions, which include neutral pH, intact microsomal vesicles, and low free Ca(2+) in the lumen, the Ca(2+)/P(i) coupling ratio was 0.6. A rise or decrease in pH, high free Ca(2+) in the lumenal space, or the addition of dimethyl sulfoxide increase the intramolecular uncoupling. Alkaline pH and/or high free Ca(2+) in the lumen potentiate the accumulation of enzyme conformations with high Ca(2+) affinity. Acidic pH and/or dimethyl sulfoxide favor the accumulation of enzyme conformations with low Ca(2+) affinity. Under standard assay conditions, two uncoupled routes, together with a coupled route, are operative during the hydrolysis of p-nitrophenyl phosphate in the presence of Ca(2+). The prevalence of any one of the uncoupled catalytic cycles is dependent on the working conditions. The proposed reaction scheme constitutes a general model for understanding the mechanism of intramolecular energy uncoupling. PMID- 11115503 TI - The molecular mechanism for the genetic disorder familial defective apolipoprotein B100. AB - Familial defective apolipoprotein B100 (FDB) is a genetic disorder in which low density lipoproteins (LDL) bind defectively to the LDL receptor, resulting in hypercholesterolemia and premature atherosclerosis. FDB is caused by a mutation (R3500Q) that changes the conformation of apolipoprotein (apo) B100 near the receptor-binding site. We previously showed that arginine, not simply a positive charge, at residue 3500 is essential for normal receptor binding and that the carboxyl terminus of apoB100 is necessary for mutations affecting arginine 3500 to disrupt LDL receptor binding. Thus, normal receptor binding involves an interaction between arginine 3500 and tryptophan 4369 in the carboxyl tail of apoB100. W4369Y LDL and R3500Q LDL isolated from transgenic mice had identically defective LDL binding and a higher affinity for the monoclonal antibody MB47, which has an epitope flanking residue 3500. We conclude that arginine 3500 interacts with tryptophan 4369 and facilitates the conformation of apoB100 required for normal receptor binding of LDL. From our findings, we developed a model that explains how the carboxyl terminus of apoB100 interacts with the backbone of apoB100 that enwraps the LDL particle. Our model also explains how all known ligand-defective mutations in apoB100, including a newly discovered R3480W mutation in apoB100, cause defective receptor binding. PMID- 11115504 TI - Isolation and characterization of a urobilinogenoidic chlorophyll catabolite from Hordeum vulgare L. AB - A new type of chlorophyll catabolite was isolated from extracts of de-greened primary leaves of barley (Hordeum vulgare cv. Lambic). Its constitution was elucidated by one-dimensional and two-dimensional [(1)H,(13)C]-homo- and heteronuclear NMR spectroscopic techniques and by high resolution mass spectroscopy. The isolated catabolite, a water-soluble, colorless, and nonfluorescent linear tetrapyrrole, resembles urobilinogen in which one of the propionic side chains forms a five membered isocylic ring system, indicating its origin from the chlorophylls. PMID- 11115505 TI - Role of multidrug resistance protein 1 (MRP1) and glutathione S-transferase A1-1 in alkylating agent resistance. Kinetics of glutathione conjugate formation and efflux govern differential cellular sensitivity to chlorambucil versus melphalan toxicity. AB - We investigated the role of phase II (conjugation) and phase III (efflux) detoxification of the anticancer drugs melphalan (MLP) and chlorambucil (CHB). Although both drugs are substrates of Alpha-class glutathione S-transferases (GST) and the monoglutathionyl conjugates formed in these enzymatic reactions are transported by MRP1, we found that GSTA1-1 and MRP1 acted in synergy to confer resistance to CHB but not to MLP (Morrow, C. S., Smitherman, P. K., Diah, S. K., Schneider, E., and Townsend, A. J. (1998) J. Biol. Chem. 273, 20114-20120). To explain this selectivity of MRP1/GST-mediated resistance, we report results of side-by-side experiments comparing the kinetics of MLP- versus CHB-glutathione conjugate: formation, product inhibition of GSTA1-1 catalysis, and transport by MRP1. The monoglutathionyl conjugate of CHB, CHB-SG, is a very strong competitive inhibitor of GSTA1-1 (K(i) 0.14 microM) that is >30-fold more potent than that of the corresponding conjugate of MLP, MLP-SG (K(i) 4.7 microM). The efficiency of GSTA1-1-mediated monoglutathionyl conjugate formation is more than 4-fold higher for CHB than MLP. Lastly, both CHB-SG and MLP-SG are efficiently transported by MRP1 with similar V(max) although the K(m) for CHB-SG (0.37 microm) is significantly lower than for MLP-SG (1.1 microM). These results indicate that MRP1 is required for GSTA1-1-mediated resistance to CHB in order to relieve potent product inhibition of the enzyme by intracellular CHB-SG formed. The kinetic properties of MRP1 are well suited to eliminate CHB-SG at pharmacologically relevant concentrations. For MLP detoxification, where product inhibition of GSTA1-1 is less important, GSTA1-1 does not confer resistance because of the relatively poorer catalytic efficiency of MLP-SG formation. Similar analyses can be useful for predicting the pharmacological and toxicological consequences of MRP and GST expression on cellular sensitivity to various other electrophilic xenobiotics. PMID- 11115506 TI - Control of CYP11B2 gene expression through differential regulation of its promoter by atypical and conventional protein kinase C isoforms. AB - We reported previously that the protein kinase C (PKC) inhibitor GF109203X stimulated the hamster CYP11B2 promoter activity in transfected NCI-H295 cells. PKCalpha, -epsilon, and -zeta were detected in hamster adrenal zona glomerulosa and NCI-H295 cells, and PKCtheta in NCI-H295 cells. 12-O-Tetradecanoylphorbol-13 acetate (TPA) inhibited basal and stimulated cytochrome P450 aldosterone synthase mRNA expression by angiotensin (AII), dibutyryl cyclic adenosine 3':5' monophosphate (Bt2cAMP), or KCl in NCI-H295 cells. Basal CYP11B2 promoter activity was inhibited in cells cotransfected with constitutively active (CA) PKCalpha, -epsilon, and -theta mutants, whereas it was increased with CA-PKCzeta. Dominant negative (DN) PKCalpha, -theta, -epsilon, and -zeta mutants stimulated the promoter activity. AII-, KCl-, and Bt2cAMP-stimulatory effects were abolished in cells cotransfected with CA-PKCalpha, -epsilon, or -theta. The effect of Bt2cAMP was abolished by CA-PKCzeta but AII and KCl were still able to enhance the promoter activity. DN-PKCalpha, -epsilon, -theta, or -zeta did not inhibit these effects. Go6976 enhanced promoter activity, providing further evidence that PKCalpha was involved. Various CYP11B2 promoter constructs were used to identify the area associated with TPA and PKC inhibition. TPA and CA-PKCalpha, -epsilon, or -theta abolished the effects of AII, KCl, and Bt2cAMP on the activity of -102 and longer constructs. In summary, our findings suggest that the hamster CYP11B2 gene is under differential control by conventional (alpha) and atypical (zeta) PKC. PMID- 11115507 TI - Dinucleotides as growth-promoting extracellular mediators. Presence of dinucleoside diphosphates Ap2A, Ap2G, and Gp2G in releasable granules of platelets. AB - Dinucleoside diphosphates, Ap(2)A, Ap(2)G, and Gp(2)G represent a new class of growth-promoting extracellular mediators, which are released from granules after activation of platelets. The presence of theses substances was shown after purification from a platelet concentrate. The substances were identified by UV spectrometry, retention time comparison with authentic substances, matrix assisted laser desorption/ionization mass spectrometry, post-source-decay matrix assisted laser desorption/ionization mass spectrometry, and enzymatic analysis. Ap(2)A, Ap(2)G, and Gp(2)G have growth-stimulating effects on vascular smooth muscle cells in nanomolar concentrations as shown by [(3)H]thymidine incorporation measurements. The calculated EC(50) (log m; mean +/- S.E.) values were -6.07 +/- 0.14 for Ap(2)A, -6.27 +/- 0.25 for Ap(2)G, and -6.91 +/- 0.44 for Gp(2)G. At least 61.5 +/- 4.3% of the dinucleoside polyphosphates are released by platelet activation. The intraplatelet concentrations suggest that, in the close environment of a platelet thrombus, similar dinucleoside polyphosphate concentrations can be found as in platelets. Intraplatelet concentration can be estimated in the range of 1/20 to 1/100 of the concentration of ATP. In conclusion, Ap(2)A, Ap(2)G, and Gp(2)G derived from releasable granules of human platelets may play a regulatory role in vascular smooth muscle growth as growth promoting mediators. PMID- 11115508 TI - Identification of c-myc as a down-stream target for pituitary tumor-transforming gene. AB - Pituitary tumor-transforming gene (PTTG) encodes a protein implicated in cellular transformation and transcriptional regulation. To identify downstream target genes, I established cell lines with tightly regulated inducible expression of PTTG. DNA arrays were used to analyze gene expression profiles after PTTG induction. I identified c-myc oncogene as a major PTTG target. Induction of PTTG resulted in increased cell proliferation through activation of c-myc. I showed that PTTG activates c-myc transcription in transfected cells. PTTG binds to c-myc promoter near the transcription initiation site in a protein complex containing the upstream stimulatory factor (USF1). I have defined the PTTG DNA-binding site and mapped PTTG DNA binding domain to a region between amino acids 61 and 118. Furthermore, I demonstrated that PTTG DNA binding is required for its transcriptional activation function. These results definitively established the role of PTTG as a transcription activator and indicate that PTTG is involved in cellular transformation and tumorigenesis through activation of c-myc oncogene. PMID- 11115509 TI - Enhanced multispecificity of arabidopsis vacuolar multidrug resistance-associated protein-type ATP-binding cassette transporter, AtMRP2. AB - Recent investigations have established that Arabidopsis thaliana contains a family of genes encoding ATP-binding cassette transporters belonging to the multidrug resistance-associated protein (MRP) family. So named because of the phenotypes conferred by their animal prototypes, many MRPs are MgATP-energized pumps active in the transport of glutathione (GS) conjugates and other bulky amphipathic anions across membranes. Here we show that Arabidopsis MRP2 (AtMRP2) localizes to the vacuolar membrane fraction from seedlings and is not only competent in the transport of GS conjugates but also glucuronate conjugates after heterologous expression in yeast. Based on the stimulatory action of the model GS conjugate 2,4-dinitrophenyl-GS (DNP-GS) on uptake of the model glucuronide 17beta estradiol 17-(beta-d-glucuronide) (E(2)17betaG) and vice versa, double-label experiments demonstrating that the two substrates are subject to simultaneous transport by AtMRP2 and preloading experiments suggesting that the effects seen result from cis, not trans, interactions, it is inferred that some GS conjugates and some glucuronides reciprocally activate each other's transport via distinct but coupled binding sites. The results of parallel experiments on AtMRP1 and representative yeast and mammalian MRPs indicate that these properties are specific to AtMRP2. The effects exerted by DNP-GS on AtMRP2 are not, however, common to all GS conjugates and not simulated by oxidized glutathione or reduced glutathione. Decyl-GS, metolachlor-GS, and oxidized glutathione, although competitive with DNP-GS, do not promote E(2)17betaG uptake by AtMRP2. Reduced glutathione, although subject to transport by AtMRP2 and able to markedly promote E(2)17betaG uptake, neither competes with DNP-GS for uptake nor is subject to E(2)17betaG-promoted uptake. A multisite model comprising three or four semi autonomous transport pathways plus distinct but tightly coupled binding sites is invoked for AtMRP2. PMID- 11115510 TI - Roles of NF-kappaB and 26 S proteasome in apoptotic cell death induced by topoisomerase I and II poisons in human nonsmall cell lung carcinoma. AB - Activation of signaling pathways after DNA damage induced by topoisomerase (topo) poisons can lead to cell death by apoptosis. Treatment of human nonsmall cell lung carcinoma (NSCLC-3 or NSCLC-5) cells with the topo I poison SN-38 or the topo II poison etoposide (VP-16) leads to activation of NF-kappaB before induction of apoptosis. Inhibiting the degradation of IkappaBalpha by pretreatment with the proteasome inhibitor MG-132 significantly inhibited NF kappaB activation and apoptosis but not DNA damage induced by SN-38 or VP-16. Transfection of NSCLC-3 or NSCLC-5 cells with dominant negative mutant IkappaBalpha (mIkappaBalpha) inhibited SN-38 or VP-16 induced transcription and DNA binding activity of NF-kappaB without altering drug-induced apoptosis. Regulation of apoptosis by mitochondrial release of cytochrome c and activation of pro-caspase 9 followed by cleavage of poly(ADP-ribose) polymerase by effector caspases 3 and 7 was similar in neo and mIkappaBalpha cells treated with SN-38 or VP-16. In contrast to pretreatment with MG-132, exposure to MG-132 after SN-38 or VP-16 treatment of neo or mIkappaBalpha cells decreased cell cycle arrest in the S/G2 + M fraction and enhanced apoptosis compared with drug alone. In summary, apoptosis induced by topoisomerase poisons in NSCLC cells is not mediated by NF kappaB but can be manipulated by proteasome inhibitors. PMID- 11115511 TI - Isoform-specific localization of voltage-gated K+ channels to distinct lipid raft populations. Targeting of Kv1.5 to caveolae. AB - The precise subcellular localization of ion channels is often necessary to ensure rapid and efficient integration of both intracellular and extracellular signaling events. Recently, we have identified lipid raft association as a novel mechanism for the subcellular sorting of specific voltage-gated K(+) channels to regions of the membrane rich in signaling complexes. Here, we demonstrate isoform-specific targeting of voltage-gated K(+) (Kv) channels to distinct lipid raft populations with the finding that Kv1.5 specifically targets to caveolae. Multiple lines of evidence indicate that Kv1.5 and Kv2.1 exist in distinct raft domains: 1) channel/raft association shows differential sensitivity to increasing concentrations of Triton X-100; 2) unlike Kv2.1, Kv1.5 colocalizes with caveolin on the cell surface and redistributes with caveolin following microtubule disruption; and 3) immunoisolation of caveolae copurifies Kv1.5 channel. Both depletion of cellular cholesterol and inhibition of sphingolipid synthesis alter Kv1.5 channel function by inducing a hyperpolarizing shift in the voltage dependence of activation and inactivation. The differential targeting of Kv channel subtypes to caveolar and noncaveolar rafts within a single membrane represents a unique mechanism of compartmentalization, which may permit isoform specific modulation of K(+) channel function. PMID- 11115512 TI - ATP modulation of ATP-sensitive potassium channel ATP sensitivity varies with the type of SUR subunit. AB - ATP-sensitive potassium (K(ATP)) channels comprise Kir and SUR subunits. Using recombinant K(ATP) channels expressed in Xenopus oocytes, we observed that MgATP (100 microm) block of Kir6.2/SUR2A currents gradually declined with time, whereas inhibition of Kir6.2/SUR1 or Kir6.2DeltaC36 currents did not change. The decline in Kir6.2/SUR2A ATP sensitivity was not observed in Mg(2+) free solution and was blocked by the phosphatidylinositol (PI) 3-kinase inhibitors LY 294002 (10 microm) and wortmannin (100 microm), and by neomycin (100 microm). These results suggest that a MgATP-dependent synthesis of membrane phospholipids produces a secondary decrease in the ATP sensitivity of Kir6.2/SUR2A. Direct application of the phospholipids PI 4,5-bisphosphate and PI 3,4,5-trisphosphate in the presence of 100 microm MgATP activated all three types of channel, but the response was faster for Kir6.2/SUR2A. Chimeric studies indicate that the different responses of Kir6.2/SUR2A and Kir6.2/SUR1 are mediated by the first six transmembrane domains of SUR. The MgATP-dependent loss of ATP sensitivity of Kir6.2/SUR2A was enhanced by the actin filament disrupter cytochalasin and blocked by phalloidin (which stabilizes the cytoskeleton). Phalloidin did not block the effect of PI 3,4,5-trisphosphate. This suggests that MgATP may cause disruption of the cytoskeleton, leading to enhanced membrane phospholipid levels (or better targeting to the K(ATP) channel) and thus to decreased channel ATP sensitivity. PMID- 11115513 TI - The neuronal adaptor protein X11alpha interacts with the copper chaperone for SOD1 and regulates SOD1 activity. AB - The neuronal adaptor protein X11alpha participates in the formation of multiprotein complexes and intracellular trafficking. It contains a series of discrete protein-protein interaction domains including two contiguous C-terminal PDZ domains. We used the yeast two-hybrid system to screen for proteins that interact with the PDZ domains of human X11alpha, and we isolated a clone encoding domains II and III of the copper chaperone for Cu,Zn-superoxide dismutase-1 (CCS). The X11alpha/CCS interaction was confirmed in coimmunoprecipitation studies plus glutathione S-transferase fusion protein pull-down assays and was shown to be mediated via PDZ2 of X11alpha and a sequence within the carboxyl terminus of domain III of CCS. CCS delivers the copper cofactor to the antioxidant superoxide dismutase-1 (SOD1) enzyme and is required for its activity. Overexpression of X11alpha inhibited SOD1 activity in transfected Chinese hamster ovary cells which suggests that X11alpha binding to CCS is inhibitory to SOD1 activation. X11alpha also interacts with another copper binding protein found in neurons, the Alzheimer's disease amyloid precursor protein. Thus, X11alpha may participate in copper homeostasis within neurons. PMID- 11115514 TI - Intracellular redistribution of nucleolin upon interaction with the CD3epsilon chain of the T cell receptor complex. AB - T cell activation through the antigen receptor (TCR) involves the cytoplasmic tails of the CD3 subunits CD3gamma, CD3delta, CD3epsilon, and CD3zeta. Whereas the biological significance of the cytoplasmic tails of these molecules is suggested, in part, by their evolutionarily conserved sequences, their interactions with signal transduction molecules are not completely understood. We used affinity chromatography columns of glutathione S-transferase fused to the CD3epsilon cytoplasmic tail to isolate proteins that specifically interact with this subunit. In this way, we identified the shuttling protein nucleolin as a specific CD3epsilon-interacting molecule. Using competition studies and affinity chromatography on peptide columns, we were able to identify a central proline rich sequence as the nucleolin-interacting sequence in CD3epsilon. Transfection in COS cells of wild type CD3epsilon, but not of nonbinding mutants of CD3epsilon, resulted in redistribution of nucleolin from the nucleus and nucleoli to the cytoplasm. This property was transferred to a CD8 protein chimera by appending the cytoplasmic tail of CD3epsilon. We also found that nucleolin associated with the TCR complex. This association was increased upon TCR engagement, suggesting that the CD3epsilon/nucleolin interaction may have a role in T cell activation. PMID- 11115516 TI - In Reply. PMID- 11115515 TI - Nuclear import and export signals enable rapid nucleocytoplasmic shuttling of the atypical protein kinase C lambda. AB - The atypical protein kinase C (PKC) isoenzymes, lambda/iota- and zetaPKC, play important roles in cellular signaling pathways regulating proliferation, differentiation, and cell survival. By using green fluorescent protein (GFP) fusion proteins, we found that wild-type lambdaPKC localized predominantly to the cytoplasm, whereas both a kinase-defective mutant and an activation loop mutant accumulated in the nucleus. We have mapped a functional nuclear localization signal (NLS) to the N-terminal part of the zinc finger domain of lambdaPKC. Leptomycin B treatment induced rapid nuclear accumulation of GFP-lambda as well as endogenous lambdaPKC suggesting the existence of a CRM1-dependent nuclear export signal (NES). Consequently, we identified a functional leucine-rich NES in the linker region between the zinc finger and the catalytic domain of lambdaPKC. The presence of both the NLS and NES enables a continuous shuttling of lambdaPKC between the cytoplasm and nucleus. Our results suggest that the exposure of the NLS in both lambda- and zetaPKC is regulated by intramolecular interactions between the N-terminal part, including the pseudosubstrate sequence, and the catalytic domain. Thus, either deletion of the N-terminal region, including the pseudosubstrate sequence, or a point mutation in this sequence leads to nuclear accumulation of lambdaPKC. The ability of the two atypical PKC isoforms to enter the nucleus in HeLa cells upon leptomycin B treatment differs substantially. Although lambdaPKC is able to enter the nucleus very rapidly, zetaPKC is much less efficiently imported into the nucleus. This difference can be explained by the different relative strengths of the NLS and NES in lambdaPKC compared with zetaPKC. PMID- 11115517 TI - In Reply. PMID- 11115519 TI - Efficacy of preoperative dual-phase sestamibi scanning in hyperparathyroidism. AB - PURPOSE: The objective of this article is to evaluate our experience with sestamibi scanning in patients with primary and secondary hyperparathyroidism. PATIENTS AND METHODS: A retrospective review of patients referred to the radiology department at the University of Kansas Medical Center for parathyroid studies between January 1, 1993, and August 1, 1998, was done. Patients included in the study were those who underwent both dual-phase technetium (Tc-99m) sestamibi scanning and subsequent parathyroidectomy at our institution (n = 34). Twenty-six patients had primary hyperparathyroidism and 8 patients had secondary hyperparathyroidism. Fifteen had previous history of neck exploration. RESULTS: Sensitivity of sestamibi scans in detection of all abnormal pathology in cases of primary hyperparathyroidism was 60% overall. Among the subset of adenoma cases, sensitivity was 82% (14/17). Among cases of primary parathyroid hyperplasia, no scan correctly localized all abnormal glands; however 60% (3/5) showed localization of at least one hyperplastic gland. Of the 2 patients with parathyroid carcinoma, in only one case was there evidence of sestamibi retention in the correct thyroid lobe. In patients with secondary hyperparathyroidism, sestamibi scanning was successful in identifying all hyperplastic tissue in only one case (sensitivity 13%). In 7 of the 8 cases of secondary hyperparathyroidism, the scan localized at least one hyperplastic gland. CONCLUSION: Sestamibi scanning is useful in the localization of abnormal pathology in cases of primary hyperparathyroidism, especially adenomas. In cases of hyperplasia, whether attributable to primary or secondary hyperparathyroidism, sestamibi imaging is less successful. PMID- 11115520 TI - Isolated cervical recurrence of squamous cell carcinoma in the previously treated neck. AB - PURPOSE: Our study goal was to identify clinical factors associated with, and that might predict, treatment outcome for patients with an isolated cervical recurrence of squamous cell carcinoma in the previously treated neck (ICR-PTN). MATERIALS AND METHODS: We reviewed all patients with noncutaneous head and neck squamous cell carcinoma treated at our tertiary care center between 1987 and 1997, and identified 17 patients (2%) who later developed an isolated recurrence in a previously treated neck. These patients made up our study group, and their charts were thoroughly reviewed. Outcome of salvage therapy (surgery, radiation, or combined therapy) for these patients was compared with pooled clinicopathologic data using the Fisher exact test (one tail). RESULTS: Fifteen such ICR-PTN patients consented to salvage therapy. Six patients were without disease at last follow-up, and 3 were successfully palliated. A statistically significant association between the side of ICR-PTN relative to the primary tumor and outcome of salvage therapy (P =.026) was noted, with ipsilateral neck recurrence being a favorable prognostic factor. Trends that did not meet the standard for statistical significance were observed between a better outcome of salvage therapy and the following parameters: age of less than 60, nonsurgical initial treatment of the neck, and lack of a history of a recurrence before they developed the ICR-PTN. CONCLUSIONS: The current study showed that only the side of the ICR-PTN relative to the primary site is associated with outcome of salvage therapy. Based on our findings and a review of the literature, we have developed a summary of factors that might predict which patients with an ICR-PTN are most likely to benefit from aggressive salvage therapy. PMID- 11115521 TI - Malignant melanoma of the sinonasal cavity: explanation of magnetic resonance signal intensities with histopathologic characteristics. AB - PURPOSE: To evaluate the magnetic resonance (MR) findings of malignant melanoma of the sinonasal cavity and compare these findings with those of the histopathological examination. MATERIALS AND METHODS: The MR images of 11 patients with malignant melanoma of the sinonasal cavity were retrospectively reviewed. Ten patients had primary malignant melanoma of the sinonasal cavity and one had a local recurrence. The imaging findings were evaluated with special attention given to the signal intensity of the tumor, internal characteristics, and growth pattern on MR. Signal intensity and enhancement patterns of the tumors were compared with the histopathological findings. RESULTS: On T1-weighted image, 6 tumors were hyperintense (5 melanotic and one amelanotic melanoma), and 5 tumors were isointense or hypointense (4 amelanotic and one melanotic). On T2 weighted images, 2 amelanotic tumors showed hyperintensity, and 5 melanotic tumors showed hypointensity. Four tumors (one melanotic and 3 amelanotic) were isointense. Four of the 7 tumors with hyperintensity on T1-weighted images showed patchy, higher-signal intensity areas. In 3 of them, patchy areas of a higher degree of pigmentation were found on histopathological examination. There were multiple linear dark signal intensities on T2-weighted images and/or linear enhancing areas on enhanced T1-weighted images within the masses in 5 of the 11 patients. These findings could be explained as intratumoral vessels in 4 tumors and fibrous septa in one tumor on histopathological examinations. CONCLUSION: Malignant melanoma of the sinonasal cavity shows characteristic MR signal intensity, which is mainly attributable to the degree and distribution of melanin pigmentation, and partly attributable to hemorrhage within the mass. The linear, low-signal intensity on T2-weighted images or enhanced lines are intratumoral vessels or fibrous septa. PMID- 11115522 TI - Changes in the avian cochlea after single high-dose gentamicin. AB - PURPOSE: Define the time course of functional and anatomical damage and subsequent recovery (by regeneration) of hair cells in the chicken inner ear after a single high-dose of gentamicin. MATERIALS AND METHODS: Broiler chicks were given a single intraperitoneal dose (200 mg/kg) of gentamicin (n = 39) or saline (n = 39). Functional status was evaluated with auditory brainstem response (ABR) thresholds before injection and before sacrifice at 2, 5, 9, 16, 21, 28, and 70 days postinjection. The cochleae were then examined with scanning electron microscopy (SEM) to assess the extent of damage along the cochlea and absolute hair cell numbers in the basal 15% of the cochlea (high-frequency region). RESULTS: Considerable variability between animals was seen for both ABR and SEM changes. Damage was maximal at 5 days postinjection with an average ABR threshold shift of 12 dB (range -10 to 50 dB) and basal cochlear damage of 28% (range 12% 57%). Hair cell counts were significantly decreased in the basal 15% of the cochlea at 5 days. Hair cell regeneration resulted in rapid anatomical and functional recovery, but evidence of hair cell disorganization persisted at 70 days despite improved thresholds. CONCLUSION: A single high dose of gentamicin produces a significant but variable anatomical and functional insult in the chick cochlea. Hair cell regeneration results in rapid but incomplete recovery. PMID- 11115523 TI - Aphthous ulcers: a difficult clinical entity. AB - Recurrent aphthous ulcers (RAU) are the most common oral ulcerative disease, affecting 10% to 20% of the population. There are 3 clinical subtypes-minor, major, and herpetiform. Minor aphthous ulcers are the most common subtype, representing 80% to 90% of all recurrent aphthous ulcers. Clinically, RAU present as extremely painful, shallow ulcerations with an erythematous halo on unattached oral mucosa. The primary differential diagnosis is oral herpes simplex. The etiology of RAU is unknown. Topical corticosteroids are the mainstay of therapy. PMID- 11115524 TI - Gamma-probe localization of a parathyroid adenoma in the reoperative neck. AB - Preoperative localization of parathyroid adenomas in patients with hyperparathyroidism currently relies on a combination of computed tomography, magnetic resonance imaging, ultrasound, (99m)Tc-sestamibi scintigraphy, and venous sampling of parathyroid hormone. No procedure is universally reliable, however, and in reoperation for missed parathyroid adenomas, development of an optimal preoperative localization strategy becomes especially problematic. We report the case of a patient with hyperparathyroidism who required reoperation for a missed parathyroid adenoma despite preoperative localization with (99m)Tc sestamibi scintigraphy. (99m)Tc-sestamibi scintigraphy was done 2.5 hours before reoperation. On reoperation, a gamma-detecting probe (C-Track; Care Wise Medical Corporation, Morgan Hill, CA) introduced through a right neck incision was used to localize a 4-cm adenoma within 45 minutes. No significant radiation hazard existed, and no special handling of the specimen was required. The patient's hyperparathyroidism resolved within 24 hours postoperatively. Therefore, this intraoperative technique may prove to be a useful adjunct to preoperative localization studies of parathyroid adenomas, particularly in patients requiring reoperation for persistent postsurgical hyperparathyroidism. PMID- 11115525 TI - Neonatal mature teratoma of the sphenoidal sinus: a case report. AB - Teratoma in the head and neck region is very rare. We treated a child with a mature congenital teratoma that arose from the right sphenoidal sinus. He is doing well after early surgical treatment with endoscopic endonasal resection of the tumor. PMID- 11115526 TI - Intranasal glomangioma. AB - Glomangiomas of the nasal cavity are a rare occurrence. Only 9 cases of this vascular tumor have been reported in the literature. A case of intranasal glomangioma is reported with a review of pertinent literature. The histogenesis, clinical presentation, diagnosis, immunohistochemical features, and treatment of these rare tumors are presented. PMID- 11115527 TI - Nasal cartilage aplasia in a family with facioaudiosymphalangism syndrome. PMID- 11115528 TI - Acute nasopharyngitis in adults: an independent clinical entity? AB - Acute adult nasopharyngitis may exist as an independent clinical entity without preceding or following symptoms and signs of the usual and common upper respiratory tract infections. Three representative cases with color photographs are presented to support this conclusion. Routine endoscopic nasopharyngoscopy allows a better understanding of this uncommon nasopharyngeal and other disorders. PMID- 11115529 TI - Unilocular cervical lymphatic cyst in an adult. AB - Unilocular cervical lymphatic lesions are rare clinical entities. These endothelial-lined, fluid-filled structures must be distinguished from true cysts (epithelial lining), and pseudocysts (no cellular lining). This article presents a case report of a unilocular cervical lymphatic cyst in an adult. The embryology and anatomy of the cervical lymphatic system is discussed. PMID- 11115530 TI - Synchronous appearance of germ cell tumor and papillary carcinoma of the thyroid. AB - Synchronous appearance of 2 different malignancies in one patient is a rare phenomenon. We describe our experience of 2 patients with synchronous malignancies of the testis and thyroid gland, and of a third patient who developed a thyroid neoplasm unrelated to recent treatment for a germ cell tumor. The medical records of 3 male patients treated for both a germ cell tumor and a thyroid cancer between 1989 and 1994 were reviewed. Two patients with nonseminomatous germ cell tumor received postoperative chemotherapy after orchiectomy and developed a papillary carcinoma of the thyroid during treatment. A third patient, who received radiation therapy for a clinical stage 1 seminoma, recurred with biopsy proven seminoma in the neck in association with a thyroid nodule 2 years later. All 3 patients had their thyroid cancer treated by surgical resection, and one received adjuvant radioactive iodine. Two of the patients are currently alive and disease-free. One patient died of pulmonary complications that stemmed from bleomycin toxicity. Synchronous appearance of germ cell tumor and papillary carcinoma of the thyroid has not been previously described. Genetic predisposition may play a role in the development of such simultaneous neoplasms. PMID- 11115531 TI - Basal cell adenocarcinoma of the parotid gland presenting as a retroauricular abscess. PMID- 11115532 TI - Tamoxifen and breast cancer risk in women harboring a BRCA1 germline mutation: computed efficacy, effectiveness and impact. AB - The management of breast cancer prone women remains a tough issue despite the release of institutional guidelines. Currently, only the anti-estrogen agent Tamoxifen and prophylactic surgery are claimed to decrease breast cancer incidence. However, efficacy of Tamoxifen, particularly in BRCA1 gene carriers, remains controversial and acceptability of prophylactic surgery is low. To evaluate the expected impact of Tamoxifen in preventing hereditary breast cancers, a modelling was made according to the efficacy of the treatment with respect to biological predictors of response: estrogen receptor (ER) and pS2 status of a series of 33 BRCA1-related breast cancers (BRCA1-BCs), and using data on BRCA1-BCs penetrance, as well as compliance and acceptability of the strategy. Although, 88% of BRCA1-BCs are ER negative in our series, 30% of cases are pS2 positive, implying a potential hormonal sensitivity of a proportion of these cancers. From our modelling, the expected impact of Tamoxifen in BRCA1 gene carriers is a reduction of breast cancer incidence of about 10% according to acceptability and compliance, close to that of 5% and of 13.5% for prophylactic mastectomy, according to acceptability rates from US and French surveys respectively. Since autonomy of choice is the root of western ethics, cancer prone women should be informed about the low but valuable expected reduction of incidence of breast cancer using Tamoxifen preventive therapy. PMID- 11115533 TI - Human osteosarcoma cells release matrix degrading enzymes in response to chemokine activation. AB - Matrix degrading enzymes released upon autocrine and/or paracrine induction exert a key role in modulating tumor cell behavior. Osteosarcoma is a highly metastatic cancer, with a redundancy of autocrine loops. Here we report that human osteosarcoma cells express a wide array of chemokine receptors and respond to chemokine activation with the release of N-acetyl-beta-D-glucosaminidase and gelatinase/collagenase activity. Of the two cell lines studied, the osteoblast like MG-63 showed a higher responsivity compared to the less differentiated HOS. This suggests that chemokine modulation of matrix degrading enzymes requires the maintaining of the osteoblastic phenotype and of signaling pathways which occur in normal tissue. PMID- 11115534 TI - Low levels of cell cycle inhibitor p27kip1 combined with high levels of Ki-67 predict shortened disease-free survival in T1 and T2 invasive breast carcinomas. AB - Breast cancer is the second leading cause of cancer death in North American women. There is considerable need for better prognostic markers to identify those subsets of patients who would benefit from more aggressive therapy because their tumors show an increased risk of poor clinical behavior. p27kip1 is an important inhibitor of the cell cycle that acts by binding and inactivating cyclin dependent kinases (CDKs). The aim of this study was to determine the prognostic value of loss of p27 protein in invasive breast cancer. We performed an immunohistochemical study of 147 patients with T1 and T2 invasive breast cancers and compared survival in the high p27 expressing group with that of the low p27 expressing group. On univariate analysis comparing tumor size, nodal status, Ki 67, c-erbB-2, p53 and estrogen receptor, low or absent p27kip1 is a strong predictor of reduced disease-free survival. Importantly, on multivariate analysis, the combined effect of low p27 with high Ki-67 is a stronger predictor of reduced disease-free survival than either marker alone. This simple, reliable and inexpensive assay, particularly when used in combination with Ki-67, improves the ability to predict disease recurrence and could become a useful adjunct of breast cancer evaluation to better identify high risk patients. PMID- 11115535 TI - In vitro studies of a prolactin antagonist, hPRL-G129R in human breast cancer cells. AB - Human prolactin (hPRL) has been shown to be one of the important survival/growth factors that promotes the proliferation of breast cancer cells in an autocrine/paracrine manner. In our recent studies, we demonstrated that a hPRL antagonist with a single amino acid substitution mutation (hPRL-G129R) was able to inhibit breast cancer cell proliferation via induction of apoptosis (1). In this study three independent yet related experiments were carried out regarding the effects of hPRL-G129R in breast cancer cells. We investigated the possible mechanism(s) of hPRL-G129R induced apoptosis in breast cancer cells. It is well documented that transforming growth factors (TGF) in conjunction with hormones such as estrogen and PRL play a major role in modulating the proliferation and apoptosis of mammary cells. We first investigated the relationships between hPRL/hPRL-G129R and TGFs. We show that hPRL is able to down-regulate TGF beta 1 (apoptotic factor) secretion and up-regulate TGF alpha (survival factor) secretion in a dose-dependent manner in T-47D cells. More importantly the hPRL antagonist up-regulates TGF beta 1 and down-regulates TGF alpha secretion. When hPRL-G129R was applied together with hPRL, it blocked the effects of hPRL. Secondly, we tested the possible involvement of caspases in hPRL-G129R induced apoptosis. We have shown that caspase-3 is activated by hPRL-G129R at a concentration of 250 ng/ml in T-47D breast cancer cells. Thirdly, we explored the additive effects of an anti-neoplastic drug, cisplatin, with the hPRL-G129R in T47D breast cancer cells. We show that cisplatin and hPRL-G129R when applied together resulted in about 40% growth inhibition in T-47D cells. PMID- 11115536 TI - Human pancreatic cancer cells express non-functional Fas receptors and counterattack lymphocytes by expressing Fas ligand; a potential mechanism for immune escape. AB - The aim of this study was to investigate the expression and functional status of Fas ligand (FasL) and its receptor (Fas) in human pancreatic cancers. Using RT PCR and Western blotting, Fas and FasL were expressed in seven surgically resected pancreatic cancer specimens and five cell lines; Capan-1, AsPC-1, BxPC 3, PANC-1, and MIA PaCa-2. In the resected specimens, pancreatic cancer cells induced apoptosis in the surrounding lymphoid cells. In coculture experiments of pancreatic cancer and Jurkat T cells, 50% of Jurkat T cells underwent apoptosis after 2 days, however, almost all pancreatic cancer cells remained viable. In addition, by testing Fas function using anti-Fas antibody (CH11), all cell lines were resistant to Fas-mediated apoptosis except Capan-1 cells which showed sensitivity similar to that of Jurkat T cells. These results suggest that pancreatic cancer cells evade immune surveillance by expression of FasL and non functioning Fas that allow them to activated T-cells. These tumor escape mechanisms may contribute to the rapid fatal course of pancreatic cancer. PMID- 11115537 TI - Augmented hepatocellular carcinoma progression and depressed Kupffer cell activity in rat cirrhotic livers. AB - To examine the property of hepatocellular carcinoma (HCC), rat HCC cells were implanted into normal and cirrhotic rat livers. Implanted HCC grew much more progressively in cirrhotic livers than in normal livers. Kupffer cells were decreased profoundly in cirrhotic livers, resulting in markedly impaired phagocytic activity. Furthermore, production of Kupffer cell-related cytokines, such as interleukin-1 beta and tumor necrosis factor-alpha, was decreased profoundly in cirrhotic livers. Our results indicate that liver cirrhosis is a prominent promoting factor in HCC progression, and that markedly depressed Kupffer cell activity may play a role in augmented HCC progression in cirrhotic livers. PMID- 11115538 TI - Relation of type II transforming growth factor-beta receptor to hepatic fibrosis and hepatocellular carcinoma. AB - Hepatocarcinogenesis is closely related to hepatic fibrosis. In this study, we investigated the relationship of type II transforming growth factor-beta receptor (T beta RII) to hepatic fibrosis and hepatocellular carcinoma (HCC). In vivo: liver tissues were obtained from 30 patients (10 chronic hepatitis, 7 cirrhosis, 13 HCC). Protein expression and immunolocalization of T beta RII were examined by Western blot analysis and immunohistochemistry. In vitro: T beta RII protein expression in hepatoma cell lines (HepG2, Hep3B, HLE, HLF and Huh7) was examined by Western blot analysis. Next, we transfected T beta RII cDNA to Huh7, and compared the change of cell number and observed the induction of apoptosis after TGF-beta1 treatment using a FACScan flow cytometer. In vivo: T beta RII immunolocalization in liver tissues was significantly decreased in patients with HCC compared with that of patients with chronic hepatitis or liver cirrhosis. In Western blot analysis, T beta RII expression in tissues attenuated in comparison with that in non-tumor tissues in some patients with HCC. In vitro: T beta RII protein expression in HLE, HLF and Huh7 cells was weaker than that in HepG2 and Hep3B cells. In Huh7 cells transfected T beta RII cDNA, cell arrest and apoptosis were obviously induced. These results indicated that human HCC has a reduced expression of T beta RII for TGF-beta1. This may provide a selective growth advantage to HCC to escape the inhibitory growth signals of TGF-beta1, and may be linked with critical steps in the growth of hepatoma cells. PMID- 11115540 TI - Determination of telomerase activity in squamous cell carcinoma of the penis. AB - Telomerase activity was studied in 51 penile carcinomas, and detected in all samples from 3 patients with verrucous carcinoma, in 85.4% (41/48) of invasive carcinomas, in 81.8% (9/11) of adjacent non-cancerous skin and in 80% (8/10) of adjacent non-cancerous corpus cavernosum. All skin and corpus cavernosum samples from patients with prostatic carcinoma were found to be telomerase negative. Our results indicate a correlation between frequency of telomerase activity and grade of penile carcinoma. The finding of telomerase activity in skin and corpus cavernosum samples adjacent to tumor suggests that unidentified local factors may modulate telomerase activity in normal tissues. PMID- 11115539 TI - Adenovirus-mediated gene transfer into tumors: evaluation of direct readministration of an adenoviral vector into subcutaneous tumors of immunocompetent mice. AB - Because systemic administration of adenoviruses appears to be limited by their immunogenicity, we examined the feasibility of intratumoral administration of adenoviruses. Direct intratumoral administration of adenoviruses resulted in efficient but transient transgene expression. When adenoviruses were readministered directly into tumors, re-expression of the transgene was achieved. Transgene expression induced by the adenoviral readministration was, however, markedly weaker than that induced by the initial administration. Furthermore, intratumoral readministration of adenoviruses elicited profound humoral and cellular immune responses to adenoviruses. These results may have important implications for efficacy considerations when adenoviral vectors are used for clinical cancer gene therapy. PMID- 11115541 TI - Regulation of the uPA gene in various grades of human glioma cells. AB - Urokinase-type plasminogen (uPA) activator regulates a variety of processes, including morphogenesis, cell differentiation, migration, and invasion. In previous studies, we demonstrated that uPA levels are significantly higher in anaplastic astrocytoma and glioblastoma than in low-grade glioma and normal brain tissue. In the present study, our goal was to determine whether the increase in uPA production in higher-grade gliomas is caused by an increase in mRNA stability or increased transcription of the gene in three human glioma cell lines of various grades (H4, SW1783, UWR3). The half-life of uPA mRNA was about 14 h in UWR3 and 8 h in SW1783 cells. In transient transfection studies of the wild-type 2109-bp human uPA promoter in the different grades of cell lines, the uPA promoter activity was increased two-fold in SW1783, anaplastic astrocytoma cells and six-fold in UWR3 glioblastoma cells, as compared with the uPA promoter activity in low-grade H4 cells. Using human uPA promoter chloramphenicol acetyl transferase (CAT) constructs with mutations of the AP-1 element at -1967 or the PEA-3 cis element at -1973, the activity of the uPA promoter was decreased 4-fold to 10-fold in all three human glioma cell lines. In transient transfection assays, the uPA promoter was stimulated 2.2-fold in UWR3 and SW1783 cells and 3.7 fold in H4 cells in response to phorbol-12-myristat-13-acetate. We further studied the activation and inhibition of uPA promoter by co-expression of a transactivation domain lacking c-jun: a dominant negative ERK1 and ERK2 mutant and a dominant negative c-raf in glioblastoma cell line showed repressed uPA promoter activity compared with the effect of the empty expression vector. We conclude from our findings that increased transcription is the more likely mechanism underlying the increase in uPA production in high-grade gliomas. PMID- 11115542 TI - Cell growth and gene modulatory activities of Yunzhi (Windsor Wunxi) from mushroom Trametes versicolor in androgen-dependent and androgen-insensitive human prostate cancer cells. AB - The incidence of prostate cancer varies greatly throughout the world; it is highest in African-Americans and lowest in the Asian populations of China, India, and Japan. Geographical differences in both prevalence of latent prostate cancer and mortality have been postulated to be influenced by diverse tumor-promoting and protective factors, both environmental and dietary. Prostate cancer is a tumor with an extremely long latency; the pattern of prostate tumorigenesis, in terms of the display and sequence of appearance of particular molecular or biochemical features, or morphological changes, characterizing different stages of the carcinogenic process, is expected to be heterogeneous. Some insights into tumor heterogeneity and progression can be obtained from studies using cell lines, particularly those derived from different anatomical sites. The present study aims to investigate whether hormone-responsive LNCaP and androgen refractory JCA-1, PC-3, and DU-145 prostate cancer cells are responsive to Yunzhi (YZ), a proprietary dietary supplement prepared from extracts of Trametes versicolor, also known as Coriolus versicolor (a mushroom consumed by Chinese for its purported health benefits), and to elucidate its mechanism of action. Ethanolic extracts (70%) of YZ significantly reduced LNCaP cell growth, down regulated the levels of secreted PSA, but had less effects on the expression of intracellular PSA and did not affect levels of the androgen receptor. In androgen unresponsive prostate cancer cells, YZ had a much less pronounced suppressive effect on proliferation of PC-3 and DU-145 cells, compared to LNCaP, and was inactive against JCA-1 cells. Western blot analyses show that the expression of Rb, a key regulatory protein in G1/S transition, and PCNA, integrally involved in mammalian cell DNA replication, were significantly reduced by treatment with YZ in PC-3 and DU-145 cells, respectively. In contradiction, none of these biochemical parameters were affected in JCA-1 cells under identical treatment conditions. Further analysis shows that YZ increased the levels of signal transducer and activator family of transcription factors STAT 1 and STAT 3 in JCA 1 and not LNCaP cells. The greater sensitivity of LNCaP cells to this polysaccharopeptide raises the possibility that YZ may be considered as an adjuvant therapy in the treatment of hormone responsive prostate cancer; additionally, it may have chemopreventive potential to restrict prostate tumorigenic progression from the hormone-dependent to the hormone-refractory state. PMID- 11115543 TI - The novel monoclonal antibody MH8-4 inhibiting cell motility recognizes integrin alpha 3: inverse of its expression withmetastases in colon cancer. AB - The molecular basis of cell motility is obviously highly complex and is considered to be controlled by a number of molecular systems including cell adhesion molecules, their receptors, cytoskeletal components, a junctional unit connecting cytoskeletal components and membrane receptors, and various peptide growth factors. The possible involvement of proteins at the cell surface in controlling cell motility has been systematically investigated. Previously, we have addressed this question using functional monoclonal antibodies (MAbs), which inhibit cell motility as probes. In order to further identify cell surface molecules involved in metastasis of gastrointestinal tumors, the present study utilized an approach based on the selection of a colon cancer cell line RPMI4788, which showed high motility out of a large number of human gastrointestinal tumor cell lines. MAb MH8-4 was established after immunization of mice with RPMI4788 and selected on the basis of inhibition of RPMI4788 cell migration in a transwell penetration assay. MH8-4 inhibited the phagokinetic tract motility of various cancer cell lines. A cDNA cloning revealed that MH8-4 recognized a specific protein structure, integrin alpha 3. In order to determine whether these experimental results are of relevance with respect to actual human gastrointestinal tumors, we investigated integrin alpha 3 expression in 40 colon cancers with distant metastases. Our immunohistochemical study showed that in almost 27.5% of the cases, the metastatic tumors had lower integrin alpha 3 levels than their corresponding primary tumors. Moreover, there were no primary tumors with lower integrin alpha 3 expression than their corresponding metastatic tumors. Our data suggest that low integrin alpha 3 expression may be associated with the metastatic potential of certain colon cancers. PMID- 11115545 TI - K-ras mutation in sputum of primary lung cancer patients does not always reflect that of cancerous cells. AB - K-ras mutation in sputum was examined using mutant-allele-specific amplification method among 100 primary lung cancer and 15 non-oncological patients. K-ras mutation was detected in 11 out of 59 adenocarcinoma cases (18.6%), 5 out of 32 squamous cell carcinoma cases (15.6%), 2 out of 4 large cell carcinoma cases (50.0%) and 3 out of 15 non-oncological disease cases (20.0%). In the 18 cases of primary lung cancer K-ras mutation was examined in both sputum and the resected specimen of the primary lesion. In 5 cases K-ras mutation in sputum was detected without K-ras mutation in primary lesion. Therefore, these findings suggested that K-ras mutation in sputum may not be directly related to that of the primary lesion. PMID- 11115544 TI - The error-prone DNA polymerase zeta catalytic subunit (Rev3) gene is ubiquitously expressed in normal and malignant human tissues. AB - Mutagenesis induced by UV light and chemical agents in yeast is largely dependent on the function of Rev3, the catalytic subunit of DNA polymerase zeta that carries out translesion DNA synthesis. Human and mouse homologues of the yeast Rev3 gene have recently been identified, and inhibition of Rev3 expression in cultured human fibroblasts by Rev3 anti-sense was shown to reduce UV-induced mutagenesis, indicating that Rev3 also plays a crucial role in mutagenesis in mammalian cells. A common variant transcript with an insertion of 128-bp between nucleotides +139 and +140 is found in both human and mouse Rev3 cDNAs, but its biological significance has not been defined. We show here that the insertion variant is not translatable either under in vitro or in vivo conditions. We also found that the translational efficiency of Rev3 gene is enhanced by the 5' untranslated region that contains a putative stem-loop structure postulated to inhibit the translation. Since the human Rev3 gene is localized to chromosome 6q21, a region previously shown to contain genes involved in tumor suppression and cellular senescence, we examined its expression in various normal and malignant tissues. Rev3 and its insertion variant transcripts were ubiquitously detected in all 27 normal human tissues studied, with an additional variant species found in tissues with relatively high levels of Rev3 expression. Levels of Rev3 transcripts were similar in lung, gastric, colon and renal tumors compared to normal tissue counterparts. The data indicate that Rev3 expression is ubiquitous and is not dysregulated in malignancies. PMID- 11115546 TI - The detection of p53 gene mutation using a microdissection technique in primary intracranial germ cell tumors. AB - Using a microdissection technique, the contribution of the p53 mutation to tumorigenesis and prognosis in each histological subtype of the intracranial germ cell tumors (GCTs) was evaluated. Nineteen patients had primary intracranial GCTs, including 4 germinomas (GEs), 4 teratomas (TEs), 1 mixed tumor of GE and TE, and 10 mixed GCTs containing non-germinomatous malignant germ cell tumors (NG MGCTs). After microdissection of specific subtypes, genomic DNA was screened for mutations in exons 5-8 of the p53 gene, using the dideoxyfingerprinting (ddF) followed by direct DNA sequencing. The direct sequencing revealed a total of six mutations in PCR products derived from the five cases (26%) which showed mobility shifts in ddF. Among the six mutations detected, four were missense mutations and two were silent. Missense mutations of the p53 gene tended to occur more frequently in the NG-MGCT component than in the GE or TE components (3/15 vs. 1/12 vs. 0/13). The incidence of missense mutations was not different between the survivors (3/13) and the deceased (1/6). This study suggests the possible role of the p53 gene in the tumori-genesis of NG-MGCT. However, p53 gene mutation did not correlate with the prognosis of NG-MGCT. PMID- 11115547 TI - Bystander effect in uracil phosphoribosyltransferase/5-fluorouracil-mediated suicide gene therapy is correlated with the level of intercellular communication. AB - We examined whether a suicide gene/prodrug system using the uracil phosphoribosyltransferase (UPRT) of E. coli origin and 5-fluorouracil (5-FU) could achieve a bystander effect in two rodent tumor cell lines, murine colon carcinoma (Colon 26) and rat gliosarcoma (9L) cells. Cytotoxicity tests of mixed populations consisting of parent and transduced cells showed that the bystander effect was not produced in Colon 26 cells in either the UPRT/5-FU system or the herpes simplex virus-thymidine kinase/ganciclovir system but a strong bystander effect was evidenced by both suicide gene systems in 9L cells. The expression level of connexin 43, a protein that constitutes gap junctions, was high in 9L but low in Colon 26 cells. A gap junction-permeable fluorescein dye could be transferred among 9L cells but hardly at all among Colon 26 cells. Taken together, the efficacy of the bystander effect in the UPRT/5-FU system can be affected by gap junction-mediated intercellular communication. PMID- 11115548 TI - Targeted systemic chemotherapy using magnetic liposomes with incorporated adriamycin for osteosarcoma in hamsters. AB - To control the growth of primary tumors effectively with systemic chemotherapy, we recently developed intravenously administered small-sized magnetic liposomes as an anticancer drug carrier. We previously reported that intravenously administered magnetic liposomes with incorporated adriamycin (magnetic ADR liposomes) effectively delivered ADR to the target site where a permanent magnet was implanted. In the present study, the therapeutic efficacy of this novel treatment approach, which involves a combination of magnet implantation at the target site and intravenous administration of magnetic liposomes, was further evaluated by comparing tumor growth rates among different administration modalities and by histological examination of treated tumors. Small-sized magnetic ADR liposomes with a mean diameter of 146 nm were prepared by the reverse-phase evaporation method. Syrian male hamsters inoculated with osteosarcoma, Os515, in the right hind limb were studied 7 days after inoculation. One day prior to the animal study, either a permanent magnet (with magnetic force) or non-magnetic alloy (without magnetic force) was implanted in the center of the tumors. Treatment with magnetic ADR liposomes under magnetic force showed significantly greater antitumor activity than intravenous administration of ADR solution or that of magnetic ADR liposomes without magnetic force. ADR administered as magnetic liposomes eliminated weight loss of hamsters, one of the side effects produced by ADR. Interestingly, magnetic liposomes (without incorporated ADR) given under magnetic force also suppressed the tumor growth. The selective accumulation of magnetite particles in the tumor blood vessels was observed by histological examination. These results suggest that this systemic chemotherapy can effectively control the primary tumor without significant side effects, due to the targeting of magnetic ADR liposomes. PMID- 11115549 TI - Overexpression of tissue factor pathway inhibitor-2 (TFPI-2), decreases the invasiveness of prostate cancer cells in vitro. AB - Human tissue factor pathway inhibitor-2 (TFPI-2) is a 32-kDa serine protease inhibitor that inhibits plasmin, trypsin, chymotrypsin, cathepsin G, and plasma kallikrein but not urokinase and tissue-type plasminogen activators or thrombin. After discovering that TFPI-2 expression is down-regulated or lost during tumor progression, we investigated the role of TFPI-2 in the invasiveness of the prostate cancer cell line (LNCaP). We stably transfected LNCaP cells with a 0.7 kb vector expressing TFPI-2 in the sense orientation and measured the expression of TFPI-2 protein and mRNA by these cells by western and northern blotting. Neither TFPI-2 protein nor mRNA was expressed by parental LNCaP cells or vector transfected controls, but levels of both protein and mRNA were significantly increased in the sense-TFPI-2 clones. The sense clones were less invasive than the control cells in Matrigel invasion and spheroid migration assays. This is the first demonstration that upregulation of TFPI-2 plays a significant role in the invasive behavior of human prostate cancer cells. PMID- 11115550 TI - p53 expression in Wilms' tumor: a possible role as prognostic factor. AB - Although a correlation between anaplasia and mutations of the p53 tumor suppressor gene has been found in Wilms' tumor (WT) a prognostic significance of p53 in WT remains largely unresolved. The goal of this study was to obtain a better understanding of the role of p53 expression in WT. Immunohistochemical analysis was performed on formalin-fixed paraffin-embedded tumor tissues from 21 patients treated in our clinic between 1984 and 1996. Eight patients presented with stage I, six with stage II, two with stage III, four with stage IV and one patient with stage V disease. According to the presence of anaplasia, four cases were categorized as of unfavorable histology based on the criteria of the National Wilms' Tumor Study Group. Seven out of 21 WTs were positive for p53. One out of the eight patients with stage I and one out of the six patients with stage II disease scored positive for p53 as were 2/2 patients with stage III and 3/4 patients with stage IV disease. Four tumors scored positive for anaplasia (one stage I, one stage II and two stage IV disease) and all four belonged to the p53 positive group. Three of these patients died of progressive disease. Immunopositivity in general was focal in the blastemal and epithelial parts of the tumors, with differences in intensity of staining ranging from moderate to strong. Positivity in the stromal components was restricted to single cells. Statistical analysis revealed significant correlations of p53 expression to anaplasia and to survival, respectively. The association of p53 expression to tumor stage was of borderline significance. To support immunohistochemistry, we performed PCR/SSCP and DNA sequence analyses on two cases, one whose immunopositivity suggested a mutated p53, and another case which was immunonegative. A CGG --> TGG base change in codon 282 of exon 8 was found in the immunopositive tumor. In conclusion, p53 may be of prognostic relevance for poor outcome being present in close association with unfavorable histology of WTs. PMID- 11115551 TI - Modulation of inducible nitric oxide synthase expression in rat intestinal cells by colon tumor promoters. AB - Metabolic epidemiological studies in humans and laboratory animal models demonstrate that consumption of diets high in fat and low in fiber excrete increased levels of 1,2-diacyl-sn-glycerol (DAG) and secondary bile acids such as deoxycholic acid (DA) and lithocholic acid that have been shown to promote colon carcinogenesis. The secondary bile acids and DAG have been shown to activate protein kinase C (PKC) and induce colonic cell proliferation. A large body of evidence indicates that iNOS, an inducible isoform of nitric oxide synthase, is over-expressed in human colon adenomas and in chemically-induced colon tumors of laboratory animals. However, the precise cascade of intracellular events that leads to the iNOS over-expression remains to be fully explored. In this study, we investigated the relationship between induction of iNOS and activation of the PKC pathway by DAG and DC, in an in vitro system using the rat intestinal cell line, RIE-1. As an initial step, we determined whether lipopolysaccharide (LPS) and phorbol 12-myristate 13-acetate (PMA) modulate iNOS protein expression in RIE-1 cells. Treatment of RIE-1 cells with LPS and PMA for 4 h significantly elevated the iNOS protein expression. The induction of iNOS by the treatment with LPS/PMA was concentration- and time-dependent. Treatment with LPS/DAG or LPS/DC also caused iNOS over-expression in a concentration- and time-dependent fashion suggesting that DAG and DC induce iNOS activity in intestinal cells. Pretreatment with specific PKC inhibitors, bisindolylmaleimide I or Go 6976, inhibited LPS/PMA, LPS/DAG, or LPS/DC-induced iNOS expression and activity. Extracts of the cells treated with LPS/PMA, LPS/DAG or LPS/DC had a high iNOS activity compared to that of control (p<0.04 to p<0.0001). Taken together, our data suggest a possible role of colon tumor promoters, DAG and DC, for iNOS over-expression through activation of the PKC pathway. PMID- 11115552 TI - Suppression of cytochrome P450 1A1 and 4A1 gene expression in renal carcinomas of TSC2 gene mutant (Eker) rats. AB - In the kidney, cytochrome P450 (CYP) is involved in arachidonic acid metabolism and the maintenance of homeostasis, but only scarce information is available as to how CYP expression is altered in rodent renal carcinomas (RCs). TSC2 gene mutant (Eker) rat RCs are an example of a Mendelian dominantly inherited predisposition to a specific cancer in an experimental animal. In the present study, the expression of CYP in Eker RCs was studied. In the normal kidney, CYP 1A1 and 4A1 mRNAs were expressed, but this expression was suppressed in spontaneously-induced Eker RCs and in cell line Lk9dL and Lk9dR. In Lk9dL and Lk9dR, Ah receptor nuclear translocator and haemoxygenase-1 mRNAs were expressed, but this expression was inconsistent in spontaneously-induced Eker RCs. The present results showed the suppression of CYP 1A1 and 4A1 mRNA expression in spontaneously-induced Eker RCs and this suppression indicates altered metabolic conditions in Eker RCs. PMID- 11115553 TI - Insufficient effect of p27(KIP1) to inhibit cyclin D1 in human esophageal cancer in vitro. AB - The cell cycle is controlled by protein complexes composed of cyclins and cyclin dependent kinases. p27KIP1 (p27) is one of the Kip/Cip family cyclin-dependent kinase inhibitory proteins which negatively regulate cell cycle progression, and have been proposed as candidate tumor suppressor genes. To examine the role of p27 in the development of human esophageal squamous cell carcinoma (ESCC), we performed Western blot and immunoprecipitation analyses of the levels of expression of p27 protein in a series of ESCC cell lines. This protein was expressed at various levels in these cell lines during exponential growth. p27 level was significantly associated with that of cyclin D1, but not of cyclin E. Further cell cycle synchronization studies demonstrated that p27 was free or bound with affinity to cyclin E-CDK2 more than to cyclin D1-CDK4 or cyclin D1 CDK6. It is known that overexpression of cyclin D1 rather than cyclin E is involved in the pathogenesis of ESCC. Our findings indicated that high expression of p27 throughout the G1 to S phase may inhibit more likely cyclin E, than cyclin D1, which promotes tumor growth of esophageal squamous cell carcinoma. PMID- 11115554 TI - Expression of G1 cell cycle markers and the effect of adenovirus-mediated overexpression of p21Waf-1 in squamous cell carcinoma of the esophagus. AB - To determine the crucial abnormality in the cell cycle regulatory proteins in human squamous cell carcinoma of the esophagus, we examined the cell growth ratio (CGR) and basal expression levels of G1 cyclins (cyclin D1, cyclin E), cyclin dependent kinase (cdk) 2, cdk4, proliferating cell nuclear antigen (PCNA), and p21Waf-1 using 9 cell lines (KE3, KE4, TE8, TE9, TE10, TE11, YES1, YES2, and YES6). Western blotting revealed an inverse linear correlation between the basal levels of p21Waf-1 expression and CGR. The protein levels of G1 cyclins, cdks, and PCNA did not coordinately reflect the CGR. There was no relationship between p21Waf-1 expression levels and mutation of the p53 gene. Next, when the cells were stimulated with serum 48 h after the starvation, stimulated levels of the above G1 cell cycle markers were variously observed among cell lines irrespective of CGR. Serum stimulation markedly induced phosphorylated Rb in TE9 (a high CGR cell line, CGR>2.0), but not in KE4 (a low CGR cell line, CGR<1.5). Furthermore, adenovirus-mediated expression of exogenous p21Waf-1 effectively reduced cell growth in KE3 and TE9 (high CGR cell lines), but not in KE4 and TE11 (low CGR cell lines). p21Waf-1-mediated growth suppression was associated with the induction of involucrin, a marker of squamous cell differentiation. Our data suggested that the basal level, but not the stimulated level, of p21Waf-1 expression play a pivotal role in abnormal growth in human squamous cell carcinoma of the esophagus. PMID- 11115555 TI - Increased activity and nuclear localisation of inositol lipid signal transduction enzymes in rat hepatoma cells. AB - To elucidate the relationship between inositol lipid signal transduction and oncogenic transformation, the activity and subcellular distribution of phospholipase C isoforms and of phosphatidylinositol 3-kinase were analysed in Morris hepatoma cells, MH(1)C(1), with respect to normal rat liver cells. The results provide evidence of a gain of function of the enzymes involved in inositide signal transduction, the amount of which increased mainly at the nuclear level. Phospholipase C and phosphatidylinositol 3-kinase activities are significantly higher in rat hepatoma than in rat liver cells. Moreover, some phospholipase C isoforms are expressed at higher levels at the nuclear level; this is particularly evident in the case of the delta 1 isoform which is not expressed at the nuclear level in rat liver cells. Therefore, the autonomous nuclear signal transduction system, formerly reported as involved in the modulation of cell proliferation and differentiation, appears also affected in oncogenic transformation. PMID- 11115556 TI - Reduced expression of INT-6/eIF3-p48 in human tumors. AB - The int-6 gene, originally identified as a common integration site for the mouse mammary tumor virus (MMTV) in mouse mammary tumors, encodes the p48 component of the eukaryotic translation initiation factor-3 (eIF3-p48). Int-6/eIF3-p48 is expressed in all adult tissues which have been tested and early in embryonic development. Int-6/eIF3-p48 has been highly conserved throughout evolution and the deduced amino acid sequence of the human gene product is identical to the mouse protein. Viral insertions at the Int-6/eIF3-p48 locus in mouse mammary tumors result in production of chimeric Int-6/eIF3-p48/MMTV products that may act as dominant negative oncoproteins. Int-6/eIF3-p48 has also been identified as a human protein that binds to the human T-cell leukemia virus type I Tax oncoprotein. The role of Int-6/eIF3-p48 in human carcinogenesis is unknown at the present time. In this study we have examined Int-6/eIF3-p48 gene status and expression in two of the most common forms of cancer in humans, breast and lung tumors. Sixty-two breast carcinomas and 78 non-small cell lung carcinomas (NSCLC) were investigated. LOH at the Int-6/eIF3-p48 locus was observed in 5 (21%) of 24 informative breast tumors and 10 (29%) of 34 informative lung tumors. A reduced expression of Int-6/eIF3-p48 was seen in 23 (37%) of breast cancer samples and 24 (31%) of NSCLC samples. An association between Int-6/eIF3-p48 expression and LOH at the Int-6/eIF3-p48 locus was observed. Int-6/eIF3-p48 expression was not related to commonly used pathological parameters in breast cancer patients, while in NSCLC patients int-6/eIF3-p48 expression was mainly seen in adenocarcinomas (P<0.0001). In conclusion, our data show for the first time a decreased expression of Int-6/eIF3-p48 in a consistent portion of human breast and lung carcinomas, frequently associated with LOH at the Int-6/eIF3-p48 locus. Additional studies on larger series of tumor specimens with long-term follow-up are needed to determine whether Int-6/eIF3-p48 expression may represent a new prognostic or predictive marker. PMID- 11115557 TI - Direct in situ PCR allows rapid and sensitive detection of high risk human papillomavirus in cytologic specimens and formalin-fixed paraffin tissues by fluorescent labelling. AB - We developed a rapid, sensitive and robust high risk human papillomavirus (HR HPV) detection protocol based on direct in situ PCR technology and fluorochrome modified nucleotides on cytologic specimens (cell smears) and on HPV infected tissues (CIN III). Reproducible results on both cytologic specimens and paraffin embedded tissues were obtained, providing a powerful tool for clinical investigation on HR HPV infection. Quantitative PCR performed on the same tissue sections adjacent to those used for in situ techniques allowed us to establish the sensitivity of our methods, able to detect rare copies (about 15 in our paraffin-embedded tissues) of HPV. PMID- 11115558 TI - Failure of Bcl-2 to block mitochondrial dysfunction during TRAIL-induced apoptosis. Tumor necrosis-related apoptosis-inducing ligand. AB - Tumor necrosis (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family of cytokines that promotes apoptosis. TRAIL induces apoptosis in a wide variety of tumor cells but not in normal cells. Oncogene Bcl-2 can protect cells from apoptosis induced by various stress stimuli. However, it is not clear whether Bcl-2 can regulate TRAIL-induced apoptosis. The objective of this study was to investigate whether Bcl-2 can regulate apoptosis induced by TRAIL. TRAIL initiates the activation of caspases, the loss of mitochondrial transmembrane potential (Delta psi(m)), and the redistribution of mitochondrial cytochrome c. TRAIL has no effect on Delta psi(m) and apoptosis in Jurkat cells deficient in either FADD or caspase-8, suggesting both FADD and caspase-8 are required for TRAIL signaling. Overexpression of Bcl-2 delays, but does not inhibit, TRAIL induced Delta psi(m), cytochrome c release from mitochondria and apoptosis, whereas etoposide-induced apoptosis is blocked by Bcl-2. XIAP, cowpox virus CrmA and baculovirus p35 inhibits TRAIL-induced apoptosis. These data suggest that TRAIL can be used to kill Bcl-2 positive cells that can not be killed by other class of chemotherapeutic drugs. PMID- 11115559 TI - Integrin alpha 5 beta 1 suppresses apoptosis triggered by serum starvation but not phorbol ester in MCF-7 breast cancer cells that overexpress protein kinase C alpha. AB - MCF-7 breast cancer cells grow as adherent cells, but following overexpression of protein kinase C-alpha these cells (MCF-7-PKC-alpha cells) become anchorage independent and exhibit increased tumorigenicity in nude mice. MCF-7-PKC-alpha cells are also sensitized to apoptosis in response to phorbol ester but not serum starvation. Flourescence-activated cell sorting revealed that several integrin subunits were down-regulated in MCF-7-PKC-alpha cells, however, the fibronectin receptor alpha 5 beta 1 was upregulated. MCF-7-PKC-alpha cells growing under non adherent conditions underwent cell death when antibodies to alpha 5 beta 1 were added to growth media lacking serum but not when serum was present. Addition of soluble fibronectin to cells incubated without serum suppressed apoptosis triggered by anti-alpha 5 beta 1 antibodies but not by phorbol esters. MCF-7-PKC alpha cells also were shown to express more fibronectin on their cell surface than MCF-7V cells (MCF-7 cells transfected with pSV(2)M(2)6 vector only). This study indicates that the survival of MCF-7-PKC-alpha cells under non-adherent conditions in the absence of serum results from the ligation of alpha 5 beta 1 with surface-bound fibronectin, which may account, in part, for the increased aggressiveness of these cells. PMID- 11115560 TI - Multiple regions with allelic loss at chromosome 3 in superficial multifocal bladder tumors. AB - We have examined six patients with multiple low grade, low stage superficial multifocal bladder tumors with surrounding tissues for loss of heterozygosity (LOH) and microsatellite instability at chromosome 3, totaling 76 samples. The majority (4/5) of the patients had LOH at or close to the fragile histidine triad (FHIT) gene (3p14.2; D3S1300), which is a candidate tumor suppressor gene for many cancer types. One patient showed a consistent LOH with four adjacent markers around FHIT region in all the tumors whereas in the corresponding surrounding tissues the heterozygosity was retained. In addition to the region near FHIT, two other regions had frequent allelic losses - one near the p telomere (3p25-26; D3S3050) and another near the q telomere (3q27; D3S2418). The largest numbers of LOH in the surrounding tissues were found at these regions (3/5 at D3S3050 and 2/5 at D3S2418). The D3S3050 marker is located at 3p26-3p25, near the Von Hippel Lindau (VHL) tumor suppressor gene locus. LOH that were more random were found at 3q21.3-25.2 (D3S1744) and at 3p12-3p11 (D3S2465). Taken together, at least three regions at chromosome 3p25-26, 3p14.2 and 3q27 seem to have frequent loss of heterozygosity in multifocal superficial bladder tumors. We also performed a phylogenetic-type analysis to find out common changes and the degree of heterogeneity. The overall heterogeneity was low within a given patient: in all cases the majority of the tumor samples arranged in a single branch with a common origin. This point of origin varied from patient to patient, which is compatible with the earlier studies demonstrating the heterogeneity of the single primary bladder tumors. However, the phylogenetic-type analysis suggests that the FHIT region contains often the very first alterations at chromosome 3. PMID- 11115561 TI - Follicular thyroid cancer cells: a model of metastatic tumor in vitro (review). AB - We used a thyroid metastatic tumor model to analyze some of the mechanisms of invasion and metastasis in culture. Chronic TSH stimulation (thyroid stimulating hormone) was associated with enhanced tumor proliferation and aggressiveness. We present a unique metastatic tumor model including three follicular thyroid cancer cell lines using a human primary tumor and two metastases of the same patient. They contain thyroglobulin, have intact thyroid functions and response to TSH. Investigating growth factor sensitivity we found that the amplitude of stimulation or inhibition of invasion was significantly smaller in both metastatic cell lines. Unstimulated cells of the lung metastasis had the highest basal invasive potential, but were only minimally affected by the stimulation of growth factors. In contrast, the parental cell line had the lowest basal invasiveness, but was considerably stimulated by growth factors. PMID- 11115563 TI - Different characteristics of carcinoma in the gastric remnant: histochemical and immunohistochemical studies. AB - Seventy cases of cancer of the gastric remnant were divided into three groups: 33 cases following surgery for benign disease (group A), and 15 cases occurring more than 10 years and 22 cases occurring within 10 years after the first gastrectomy for malignant disease (groups B and C, respectively). Then mucin histochemical and immunohistochemical studies were undertaken. Billroth-II procedure for anastomosis was most frequently performed in group A. Intestinal metaplasia within the mucosa surrounding the carcinomas was more frequently present in groups A and C with a diffuse distribution. Intestinal-type surrounding mucosa was significantly more frequent in group C. The immunohistochemical expression of p53 protein was most frequently expressed in group B. We conclude that a different mechanism of carcinogenesis exists in these three groups; i) group A: the reflux of duodenal juice especially following B-II procedures leads to progression of the carcinoma. ii) group B: some genetic factor such as p53 may be related to the metachronous multiple carcinogenesis. iii) group C: metachronous multiple carcinogenesis within the short interval may be closely associated with diffuse intestinal metaplasia in the surrounding mucosa. PMID- 11115562 TI - Differential expression of progression-related genes in the evolution of superficial to invasive transitional cell carcinoma of the bladder. AB - It is generally accepted that there are dichotomous biologic pathways that lead to the development of either: i) superficial papillary (Ta) transitional cell carcinoma (TCC) or ii) precursor lesions to muscle-invasive (CIS, T1) TCC and muscle-invasive (> or =T2) TCC. We investigated the expression of several progression-related genes to characterize the phenotype of these tumors within these divergent developmental pathways. Using a colorimetric in situ hybridization technique, we examined the expression of mRNAs of several progression-related genes in archival, pathologic specimens from 77 patients with bladder TCC. These genes included basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), interleukin (IL)-8, matrix metalloproteinase (MMP)-9, and epidermal growth factor receptor (EGFR). Relative gene expression was quantified using image analysis. Gene expression was normalized using poly (dT) and the expression of each factor in a panel of specimens of normal urothelium. Patients were stratified according to disease stage, and the level of gene expression among the stratified groups was compared. VEGF, bFGF, IL-8, and MMP-9 expression was increased in muscle-invasive compared with superficial papillary tumors, (p<0.05) and VEGF expression was increased in muscle-invasive tumors compared with CIS specimens (p<0. 05). bFGF, IL-8, and EGFR expression was increased in CIS specimens compared with superficial papillary tumors (p<0.05). The pattern of expression of bFGF, VEGF, IL-8, MMP-9, and EGFR represent the divergent developmental pathways in the pathogenesis of bladder TCC, which characterizes superficial or invasive bladder cancer. bFGF, IL-8, and EGFR appear to be upregulated in early precursor lesions (CIS), whereas VEGF appears to be upregulated at later stages in the development of muscle-invasive TCC. PMID- 11115564 TI - Prognostic results of cisplatin IP and carboplatin IV with G-CSF in patients with ovarian cancer. AB - We performed a dose-escalation study of carboplatin combined with a fixed dose of intraperitoneal cisplatin and G-CSF in patients with epithelial ovarian cancer, and analyzed the progression-free and overall survival. Six of the patients who entered the study with stage IC and II disease are still alive with no evidence of disease. The five-year survival rate was 61% for the 18 patients with stage III and IV disease; progression-free survival over 5 years was 32%. Our results show this to be an effective treatment regimen for epithelial ovarian cancer. Prognosis is good with this combined carboplatin/cisplatin/G-CSF therapy, especially for those patients with microscopic or no residual disease. PMID- 11115565 TI - Prognostic factors for recurrence in stage II and III gastric cancer patients receiving a curative resection and postoperative adjuvant chemotherapy. AB - Prognostic value of clinicopathologic factors and biologic markers was analyzed in 185 patients who received a curative resection and adjuvant chemotherapy of pathologically confirmed stage II or III gastric cancer. No difference was found between the chemotherapeutic regimens according to the frequency of recurrence, but tumor type, histology, depth of invasion, nodal metastasis, and lymphatic and venous invasion were significantly different between recurrent (n=62) and non recurrent (n=123) patients. However, the degree of lymphatic dissection and the patterns of biological markers (DNA ploidy, p53 staining and PCNA labeling) were not different. Hepatic metastasis and venous invasion were more frequent on patients recurring within one year, compared to those who recurred later. Multivariate analyses showed that depth of invasion, level 2 lymph node metastasis and tumor histology were risk factors for recurrence. Pathologic factors were more important for predicting recurrence than biological markers. PMID- 11115566 TI - Mutation analysis of hBUB1, human mitotic checkpoint gene in multiple carcinomas. AB - hBUB1 is a human homolog of yeast mitotic check point gene that plays an important role in chromosome segregation. Recently mutations of hBUB1 were reported in colorectal cancer cell lines, indicating that inactivation of this gene could be directly involved in aneuploidy in human carcinoma cells. To obtain information of the magnitude of hBUB1 inactivation in multiple carcinomas, we examined mutations in 59 multiple carcinoma cell lines showing single base alteration, however, there was no mutation of hBUB1 with amino acid change in these carcinomas. There were four silent mutations at codon 93, codon 735, codon 430 and codon 98 in KYSE190, TE8 esophageal carcinoma cells, KATOIII gastric carcinoma cells and 697 B cell leukemia cells, respectively. Two candidates of mutation were identified in TE3 esophageal carcinoma cells and 697 B cell leukemia cell line at codon 9 and codon 285, respectively. This result suggests that the inactivation of hBUB1 may be very rare in human carcinomas, or restricted to certain cell lines of colorectal carcinomas. PMID- 11115567 TI - Establishment and characterization of malignant rhabdoid tumor of the kidney. AB - Malignant rhabdoid tumor of the kidney (MRTK) is a highly aggressive tumor which occurs in childhood and which is histologically characterized by the existence of eosinophilic intracytoplasmic inclusions. We established and characterized a cell line from this tumor with histological, immunohistochemical and cytogenetical analysis. Histologically, the tumor cells demonstrate typical eosinophilic inclusions, while immunohistochemically the cells demonstrate common mesenchymal and epithelial differentiation. Although the conventional karyotyping of this tumor lacked the abnormalities of 22q chromosome, Southern blot analysis and microsatellite analysis verified abnormalities of the BCR gene and of the hSNF5/INI1 gene. Despite the variety of locations, these common genetic abnormalities appear to contribute to distinguish rhabdoid tumor from such other small round cell tumors as primitive neuroectodermal tumor, rhabdomyosarcoma, poorly differentiated synovial sarcoma and desmoplastic small round cell tumor. PMID- 11115568 TI - Histomorphometric characteristics and cellular kinetics of colorectal polyps with epithelial serrated proliferation adjacent to carcinoma. AB - Four cases of colorectal polyps with epithelial serrated proliferation (CP-ESP) with malignant transformation were studied. In CP-ESP adjacent to carcinoma, if the nuclear size in the surface layer was significantly smaller than those in the bottom and the middle layers of the crypts, the specimen was defined as zone formation positive. If there was no significant difference among the layers, the specimen was defined as zone formation negative. Cell kinetics were evaluated using Ki-67 immunostaining. The CP-ESP regions of cases 1 and 2 showed zone formation with inferior and lateral glandular branching, and were qualitatively hyperplastic on cell kinetics. Cases 3 and 4 showed inferior and lateral glandular branching with no zone formation, and were kinetically neoplastic (adenoma). The histogenesis of hyperplastic polyps with atypia (cases 1 and 2) involves the hyperplastic polyp-carcinoma sequence. In contrast, the development of tubulovillous adenoma or serrated adenoma (cases 3 and 4) may involve the tubulovillous adenoma-carcinoma or serrated adenoma-carcinoma sequence. PMID- 11115569 TI - Skin metastases in lung cancer: analysis of a 10-year experience. AB - We reviewed our 10-year experience in skin metastases from lung cancer. We identified a total of 26 patients with 49 resected skin metastases. Skin metastases were synchronous in 6 patients, 3 of whom underwent primary simultaneous resection, and solitary in 6 (6/26=23.07%). Negative prognostic factors were primary non-resectability (p=0.001), small cell lung cancer (p=0.032), simultaneous discovery of other cutaneous (p=0.048) or extracutaneous (p=0.0005, Wald test p<0.002, Odds ratio =14.37) metastases. Skin metastasis represented the unique distant localization in almost one quarter of our cases that represent the best-survivor category: in these patients skin metastasectomy is justified. PMID- 11115571 TI - Diagnosis of metastasis of ovarian clear cell carcinoma to the peritoneum of the abdominal wall by positron emission tomography with (fluorine-18)-2-deoxyglucose. AB - A 43-year old woman, operated on for ovarian clear cell carcinoma (stage IIc) four years previously was found to have a small mass under the abdominal wall in the right lower quadrant of the abdomen. Neither diagnostic imaging (ultrasonography and MRI) nor tumor markers showed any evidence of recurrence, but positron emission tomography revealed a hot spot area, and it was diagnosed as recurrence of the ovarian carcinoma. The postoperative histopathological diagnosis was metastasis of ovarian carcinoma to the peritoneal wall. PMID- 11115570 TI - Lung cancer patients with chronic obstructive pulmonary disease. AB - The aim of this study is to evaluate characteristics in lung cancer patients with chronic obstructive pulmonary disease (COPD). Among 966 lung cancer patients admitted to our division over a period of 24 years, 73 patients were diagnosed as having COPD. There were 68 (93.2%) men and 5 women; of the tumors 43 (58.9%) were squamous cell carcinomas. Although 41 (56.2%) patients had stage IA-IIIA, only 11 (15.1%) had surgery. Coexistence of COPD was proved to be a prognostic factor (p=0.0451). Adequate palliative care to provide quality survival would be the primary goal of therapy for lung cancer patients with COPD. PMID- 11115572 TI - Serum-dependent perinuclear accumulation of Cdc42 in mammalian cells. AB - Cdc42 is a member of the Rho family of GTPase. Cdc42 has been implicated to be involved in the movement, multiplication and transformation of mammalian cells by controlling the rearrangement of actin cytoskeleton and gene expression. But the mechanism of Cdc42 function has not yet been discovered. In this report we present data showing a perinuclear accumulation of Cdc42 in response to fetal bovine serum (FBS). There was no change in the amount of Cdc42 in response to FBS in the cell. It was found that protein component(s) of serum plays a major role in the perinuclear accumulation of Cdc42. Epidermal growth factor has also been found to stimulate the perinuclear accumulation of Cdc42 while NGF has no effect. Kinase inhibitors, quercetin and NDGA were found to block signals for the perinuclear accumulation of Cdc42. This suggests that phosphorylation of cellular proteins is essential for transducing signals generated from the serum component(s) to induce the perinuclear accumulation of Cdc42. These results indicate that redistribution of Cdc42 might be an important step in alteration of gene expression for controlling various functions of the cell including cell division. PMID- 11115573 TI - The circulating auto-antibodies to p53 protein in the follow-up of lymphoma patients. AB - Mutant p53 proteins may be targets of the host immune system - consequently a certain proportion of cancer patients (the percentage varies according to the type of cancer) with tumors that carry p53 missense mutations develop circulating p53 antibodies. The present study was aimed at defining the occurrence of circulating antibodies to p53 protein in patients with various types of non Hodgkin's lymphomas (NHL). Altogether, the sera of 108 cases with various histological types of NHL and of 20 healthy controls were assessed for the presence of antibodies to p53 protein with an ELISA method. In 73 cases of NHL, also the immunohistochemical staining for p53 antigen was performed to make a rough estimation of the frequency of mutational events. The development of autoantibodies to p53 protein was observed in approximately 7% of NHL patients (predominantly in the more aggressive variants of the disease, but also in one case of small lymphocytic lymphoma). This proportion represents roughly one third of the number of patients assessed (immunohistochemically) to carry a missense p53 mutation in their tumors. The autoantibodies to p53 protein can be used as a tumor marker (early appearance, high specificity) in the follow-up of a subset of NHL patients, but, unfortunately, this subset comprises only approximately 7% of NHL patients. PMID- 11115574 TI - Comparative genomic hybridization analysis suggests a gain of chromosome 7p associated with lymph node metastasis in non-small cell lung cancer. AB - We analyzed the chromosomal gains and losses that occur in 30 non-small cell lung carcinomas by comparative genomic hybridization. Their chromosomal imbalances showed histological type-specific patterns in adenocarcinomas and in squamous cell carcinomas. The genetic changes in non-small cell lung carcinoma were also strongly dependent on metastasis to lymph node. The average numbers of chromosomal alterations were increased from 6.2 to 9.1 along with the presence of metastasis, and it gave rise to the increased copy number in specific chromosomes. In particular, a novel imbalance at 7p12-21 was recognized in a half of carcinoma with metastasis, although no genetic alteration was observed in 15 non-metastasizing lung carcinoma tested here. PMID- 11115575 TI - Characterization of the mutations of the K-ras, p53, p16, and SMAD4 genes in 15 human pancreatic cancer cell lines. AB - Human pancreatic cancer is one of the most malignant diseases in the world. In order to save patients with pancreatic cancer, it is necessary to develop novel methods for treatment. For such a purpose, a series of well-characterized cancer cell lines are of great help for in vitro studies that are generally the first step in approaching the invention of novel methods for treatment. In the present study, we analyzed 15 human pancreatic cancer cell lines for genetic alterations of the K-ras, p53, p16, and SMAD4 genes, which are very frequent targets for mutation in pancreatic cancer; these cell lines are useful resources in cancer research. PMID- 11115576 TI - Platelet-derived endothelial cell growth factor in keratinizing-type squamous cell uterine cervical cancer. AB - The purpose of this study was to determine whether intratumor expression of platelet-derived endothelial cell growth factor (PD-ECGF) is a prognostic factor. We examined specimens from 61 cases of carcinoma of uterine cervix (Ca-Cx). Specimens were stained with periodic acid-Schiff preparation with diastase (d PAS) and alcian blue (pH 2.5) to identify the presence of intracellular mucin. PD ECGF expression and microvessel count (MVC) were assessed by immunohistochemical staining. Twenty specimens stained positive for mucin. MVC correlated with clinical stage (p<0.01). There was a correlation between PD-ECGF expression and survival time (p<0.05), but no correlation was seen between mucin staining positivity, MVC, and PD-ECGF expression and survival time by the Cox proportional hazard model. Intratumor PD-ECGF expression was not a prognostic factor in keratinizing-type squamous cell carcinoma of the uterine cervix. PMID- 11115577 TI - The influence of blood arterial oxygen condition on the tumor response to preoperative radiation therapy in oral cancer patients. AB - The relationship between clinicopathological factors and response of radiation therapy in oral squamous cell carcinoma has been studied. It has been suggested that factors such as tumor site, extent and tumor differentiation determine the response to radiation therapy. It is known that oxygenation is related to the therapeutic effects of radiation therapy. However, there are few reports on the relationship between oxygen condition and the response to radiation therapy. The present study was carried out to assess whether any clinicopathological factors, including an evaluation of the oxygen condition can be used to predict the effects of preoperative radiation therapy in oral squamous cell carcinomas. Forty seven patients with oral cancer treated with external radiation therapy preoperatively were evaluated. There were no significant differences in response to the radiation with respect to age, sex, tumor site, stage, macroscopic shape of tumors, and the histological factors. The hemoglobin (Hb) and arterial oxygen content (CaO(2)) levels of favorable cases (Hb: 14.4 g/dl, CaO(2) 19.1 ml/dl) were significantly higher than those of unfavorable cases (Hb: 11.0 g/dl, CaO(2): 16.1 ml/dl). These findings suggest that oxygen conditions of oral cancer patients predict tumor response to preoperative radiation therapy. PMID- 11115578 TI - Radiation therapy with uneven fractionation for malignant gliomas. AB - From 1989 to 1995, we administered a combination of large and small fraction doses (UFX, uneven fractionation) for patients with malignant gliomas. In this study, we compared the treatment outcomes of radiation therapy by uneven fractionation to that of historical control which was treated with radiation therapy by conventional fractionation (CFX). The pathologic classification was anaplastic astrocytoma (AA) in 120 and glioblastoma multiforme (GBM) in 64 patients. Of the 184 patients, 89 patients received a conventional fractionation schedule (CFX) of radiation (2 Gy/fraction, 5 times/week; total, 60 Gy/6 weeks). The other 95 patients received an uneven fractionation schedule (UFX) of radiation (5 Gy on Monday, 1 Gy from Tuesday through Friday; total, 63 Gy/7 weeks). The one-, two-, and five-year survival rates of the AA patients were 74%, 55%, and 30%, respectively. The five-year survival rates of AA patients who received CFX and UFX schedule of radiation were 24% and 38%, respectively. In the GBM patients, the five-year survival rates were 12% in patients who received CFX and 11% in patients who received UFX. UFX seemed to be more effective than CFX for treating AA patients. In multivariate analysis using the Cox regression analysis, which included various patients and treatment characteristics, age, histology and extent of surgery were the significant prognostic factors. In conclusion, UFX is an promising fractionation method for AA patients but not for GBM patients. PMID- 11115579 TI - Gastric cancer with high telomerase activity shows rapid development and invasiveness. AB - Telomerase activity was reported to be activated in most immortal cells and cancers. As the clinical significance of telomerase activity in human gastric cancer is controversial, we investigated this activity using a telomeric repeat amplification protocol. The telomerase activity was tentatively defined by strength of activity as follows: 3+, observed with 0.06 microg of protein; 2+, observed with 0.6 microg of protein; 1+, observed with 6 microg of protein; 0, not observed under these three conditions. Telomerase activity was detected in 35 of 39 (89.7%) gastric cancer specimens. Tumors with high telomerase activities (2+/3+) tended to have a deeper invasion, lymphatic and vascular invasion, lymph node metastasis, liver metastasis, and peritoneal dissemination, as compared to findings in case of low telomerase activities (-/1+). Thus, telomerase activity of gastric cancer tissue may reflect the malignant potential of the tumor and intensive postoperative care might be required for such patients. PMID- 11115580 TI - Quintuple carcinomas with metachronous triple cancer of the esophagus, kidney, and colonic conduit following synchronous double cancer of the stomach and duodenum. AB - A patient who had undergone radical gastrectomy for synchronous gastric cancer (T(1)N(0)M(0), stage I) and duodenal cancer (Tis, stage 0) in November 1987 was found to have esophageal cancer in November 1994, and underwent radical thoracolaparotomy at our hospital (T(1)N(0)M(0), stage I). After follow-up for about 3.5 years, renal cancer was detected in April 1998, and radical nephrectomy was performed (T(1)N(0)M(0), stage I). Two years later, in April 2000, the patient was found to have a polypoid lesion in the colonic conduit used for reconstruction after esophagectomy, and endoscopic mucosal resection was performed (Tis, stage 0). The patient remains under careful follow-up, including observation of the colonic conduit and the residual large intestine. PMID- 11115581 TI - Detection of tumor-associated circulating mRNA in serum, plasma and blood cells from patients with disseminated malignant melanoma. AB - Reverse transcription polymerase chain reaction (RT-PCR)-based detection of tyrosinase mRNA is a frequently used method for the identification of circulating tumor cells in melanoma patients. The significance and practical value of this procedure for the diagnosis of tumor dissemination in melanoma patients are unclear. The conflicting results may at least partially be related to very low amounts of circulating tumor cells and to our observation that melanoma cells only transiently persist in the peripheral blood. The purpose of the present study was to evaluate the relevance of detection of extracellular melanoma specific mRNA in serum and plasma samples in comparison to blood cell samples from patients with disseminated disease (stage IV). We therefore compared the presence of specific mRNA for tyrosinase, gp100, and MART-1 by RT-PCR amplification of specific cDNA from serum, plasma, and whole blood samples of 10 melanoma patients. Melanoma-specific mRNA was detectable in whole blood samples of all ten patients tested indicating the presence of circulating melanoma cells. In addition, tyrosinase mRNA could be detected in the serum and/or plasma of 6 of 10 melanoma patients whereas gp100 and MART-1 specific transcripts were not detectable in any of the samples tested. The presence and integrity of amplifiable RNA was shown in all serum and plasma samples of patients and controls by RT-PCR-specific amplification of porphobilinogen deaminase (PBDG) mRNA. We conclude that tyrosinase mRNA but not gp100 and MART-1 mRNA can be amplified from serum and/or plasma in a subset of melanoma patients showing circulating melanoma cells. Therefore, extracellular-directed assays appear to be less sensitive and efficacious in detecting melanoma-specific transcripts compared to cellular-based assays. PMID- 11115582 TI - Radiotherapy combined with transcatheter arterial infusion of cisplatin versus oral fluoropyrimidine anticancer agent for locally advanced carcinoma of the uterine cervix: a prospective follow-up study. AB - We randomized patients with locally advanced cervical cancer to receive radiotherapy combined with transcatheter arterial infusion (TAI) of cisplatin or oral fluoropyrimidine anticancer agents, and compared the prognosis by a prospective follow-up study. Sixty patients were studied who completed their planned radiation therapy with chemotherapy at the Department of Obstetrics and Gynecology of Hiroshima University Hospital between January 1991 and December 1998. Patients were randomly assigned to receive (A) radiotherapy with TAI of 120 mg/body cisplatin twice a month at the interval of 4 weeks or (B) radiotherapy with 200 mg/day oral 5-FU or UFT every day. In both groups, radiotherapy is routinely 50 Gy of external beam irradiation to the whole pelvis and 18-20 Gy (point A dose) of intracavitary irradiation using a remote after loading system (RALS). Serious adverse reactions interfering with treatment did not appear in either group. The effective histologic response was 28/32 (87.5%) in group A and 25/28 (89.3%) in group B. The median follow-up period were 28.3 months and 25.4 months in group A and B, respectively. There was no significant difference in the overall survival and disease-free survival rates for all patients, clinical stage III and squamous cell carcinoma. We could not conclude that radiotherapy with TAI of cisplatin achieved superior therapeutic efficacy in locally advanced cervical cancer. To improve the therapeutic effects, it is important to establish a new cisplatin-containing chemoradiotherapy regimen. PMID- 11115583 TI - The significance of thymidine phosphorylase expression in colorectal cancer. AB - We measured thymidine phosphorylase activity in colorectal cancer tissue and conducted immunostaining to investigate enzyme expression in the tumor tissue. The results showed a correlation between staining ratio of thymidine phosphorylase and cancer progression as well as a correlation between enzyme activity and staining ratios of cancer cells and stromal cells. Since enzyme activity levels can be judged by staining ratios, this method may be useful for assessing cancer malignancy. PMID- 11115584 TI - Osteosarcoma in blood relatives. AB - Osteosarcoma is an uncommon tumor. Family occurrence of osteosarcoma is even rarer. Four cases of osteosarcoma in two siblings and in a father and son treated at our Institute with surgery and chemotherapy are reported. These patients had no other tumors in their family history, and had negative p53 mutations in exons 5-9 by SSCP analysis. RB, CDK4, MDM2, c-myc, c-fos, and p53 gene expression, which are the major genes involved in osteosarcoma susceptibility, were studied. Our results revealed an inactive form of p53 sporadically seen in the samples, a total loss of Rb protein expression, an increased expression of Cdk4, MDM2, c fos, and c-myc proteins which literature currently reports being the principal alterations found in osteosarcoma. These findings confirm that specific genetic alterations occur in osteosarcoma pathogenesis. PMID- 11115585 TI - Surgical treatment of laryngeal carcinoma with subglottis involvement. AB - Primary cancers arising in the subglottic region are rare and are characterized by a long asymptomatic phase. More frequently the subglottis is reached by tumors arising in the glottis or even the supraglottis through invasion of the paraglottic space. Involvement of the subglottis is associated with a relatively high frequency of stomal recurrences due to a peculiar lymphatic spread to the paratracheal nodes. We analyzed a retrospective series of 68 patients with squamous cell carcinoma of the larynx extending to the subglottis region submitted to total simple laryngectomy or total laryngectomy enlarged with hemithyroidectomy and dissection of level VI nodes (HT/SPD). Overall median follow-up is 46 months. Subglottic extension was correctly diagnosed before operation in only 13/68 patients, however the resection margins, systematically determined by the pathologist, were in every case negative. Stomal relapses in laryngectomized patients without HT/SPD have been more frequent (0.55% rate per month) than in those treated with laryngectomy and HT/SPD (0.07% rate per month). It is concluded that CT should be routinely applied in preoperative staging in order to estimate the extension of the neoplasia and surgery should always include hemithyroidectomy and dissection of the homolateral paratracheal nodes when there is even minimal involvement of the subglottis. Moreover, the high incidence of second tumors in our series is noteworthy; such patients might benefit from chemopreventive therapy. PMID- 11115586 TI - Laryngeal carcinoma--epidemiological and clinical features: experience of the Rabin Medical Center in Israel. AB - We sought to compare the epidemiological and clinical features of patients with carcinoma of the larynx treated at a major Israeli tertiary facility with other series in the literature. The charts of 361 consecutive patients from 1974 to 1995 were reviewed. Our population was distinguished from other series by a low rate of alcohol abuse (12%), high incidence of second malignancies in sites other than the upper aerodigestive tract (53%) and high rate of early-stage tumors (82%). Overall 5-year survival and local control rates were 88% and 85%, respectively. Our study suggests that the low alcohol consumption and high proportion of early-stage tumors at diagnosis, characteristic of the Israeli population of patients with laryngeal carcinoma, may explain, in part, the relatively high survival and local control rates. PMID- 11115587 TI - Splice variant expression of CD44 in patients with breast and ovarian cancer. AB - CD44 is an adhesion molecule involved in many biological functions and has been described to play a role in tumor progression as well as in promotion of metastasis. It has also been suggested that expression of certain CD44 isoforms could be useful for breast and ovarian cancer screening, detection or staging. However, many other reports document no correlation between CD44 isoform expression and tumor malignancy. In light of such contradictory findings, we evaluated by exon-specific RT-PCR whether the expression of CD44 isoforms in breast and ovarian tumors correlated with any of the diagnostic criteria used to assess these diseases. We found a deregulation in the CD44 expression pattern in malignant tumors of both type of cancer compared with the one in benign tumors or normal tissue. However, we could not find a clear correlation between this deregulation or a given CD44 isoform and any diagnostic criteria evaluated, such as age, clinical data, tumor size, hormone receptor status, histological grade or aggressiveness. PMID- 11115588 TI - The role of the breast ductal system in the diagnosis of cancer (review). AB - It has been shown that early detection of breast cancer saves lives. This review summarizes the findings of the diagnostic methods involving the breast ductal system to date, including nipple fluid cytology, nipple fluid tumor markers, ductogram, and ductoscopy. PMID- 11115589 TI - Combretastatin A-4 and doxorubicin combination treatment is effective in a preclinical model of human medullary thyroid carcinoma. AB - Medullary thyroid carcinoma (MTC), both in patients and in preclinical models, is resistant to chemotherapy. In this study, we show that the anti-neovascular agent combretastatin A-4 phosphate prodrug (CA4P) in combination with doxorubicin was effective in curtailing tumor growth in a preclinical model of human MTC. This combination of combretastatin and doxorubicin extended the doubling time of established MTC tumors in nude mice to 29 days, compared to 12 days in untreated controls. This suggests that a combination of combretastatin and a cytotoxic chemotherapeutic agent may be an effective treatment for MTC. PMID- 11115590 TI - The bacterial polysaccharide tecogalan blocks growth of breast cancer cells in vivo. AB - The growth of supportive tissue during the progression of solid tumors is an absolute requirement for the nourishment of the tumor. The blockade of this proliferative response of normal tissues to the growing tumor should hence inhibit tumor progression. We have shown earlier, that the heparinoid pentosan polysulfate (PPS) can block tumor growth and neoangiogenesis induced by Kaposi's FGF as well as by other heparin-binding growth factors (HBGFs). We now report on the effects of a bacterial polysaccharide, tecogalan, on tumor xenografts of human breast cancer cells. Tecogalan inhibited FGF-dependent SW-13 cells in vitro very similarly to PPS. Growth factor-independent MDA-MB 231 cells were used in animal studies to assess the in vivo potential of tecogalan. Subcutaneous growth of tumors was inhibited by once weekly i.v. administration of tecogalan. PPS single weekly administration showed a similar effect. No gross side effects were observed. Based on our previous studies with these models, we conclude, that tecogalan acts by blocking HBGFs released from tumor cells. Interestingly, single weekly dosing of either PPS or tecogalan appears to be strikingly more efficacious than spreading the dose over several administrations. These findings with a novel compound, tecogalan, and a novel treatment regimen, PPS, suggests a different approach to planning of therapies with these types of drugs. PMID- 11115591 TI - Non-Hodgkin's tumor and Pancoast's syndrome. AB - A 60-year old man presented with Horner's syndrome, and acute right hand and lower extremity weakness. Chest X-ray and MRI revealed a right apical lung tumor (presumed to be a primary lung cancer), with brachial plexus infiltration and spinal cord compression. Emergent radiotherapy was initiated for spinal cord compression and a biopsy was obtained 24 h later. A careful review of pathology demonstrated a non-Hodgkin's lymphoma. The patient subsequently received chemotherapy, and is now in remission. This case illustrates the importance of a tissue diagnosis before initiating therapy for a Pancoast's tumor. PMID- 11115592 TI - Cisplatin, mitomycin-C, 5-fluorouracil and leucovorin combination chemotherapy (l FCM) in locally advanced unresectable or metastatic gastric carcinoma: a phase-II study. AB - Despite relevant progress, the impact of chemotherapy on advanced gastric cancer patients' survival is still unsatisfactory. Thus, the key objective of our efforts should be palliation of the symptoms and the maintenance of an adequate quality of life. In this phase-II study, we evaluated toxicity and efficacy of the combination of cisplatin, fluorofolates and mitomycin C. Thirty-one advanced gastric cancer patients received cisplatin (15 mg/m2) and 5-fluorouracil (350 mg/m2), both for 5 consecutive days every 4 weeks; 5-fluorouracil infusion was preceded by a rapid i.v. injection of 100 mg/m2 leucovorin, while mitomycin-C (10 mg/m2) was administered on day 1 of odd cycles. Cycles were repeated every 4 weeks until disease progression. We recorded 16 objective responses (51.6%, 95% confidence interval: 42.63-60.57); furthermore, such a response rate was coupled with a moderate degree of toxicity and an extremely good tolerance. In particular, alopecia, a frequent and distressing side-effect in patients, especially women, in our series occurred only in two patients. This treatment appears to be an active and tolerable therapeutic option for patients with advanced gastric carcinoma. PMID- 11115593 TI - Arglabin-DMA, a plant derived sesquiterpene, inhibits farnesyltransferase. AB - Arglabin [1(R),10(S)-epoxy-5(S),5(S),7(S)-guaia-3(4),11(13)-dien-6, 12-olide], a sesquiterpene gamma-lactone is isolated from Artemisia glabella, a species of wormwood endemic to the Karaganda region of Kazakstan. The compound has been modified to render it water-soluble through addition of a dimethylaminohydrochloride group to the C(13) carbohydride moiety to yield Arglabin-DMA. Arglabin-DMA is a registered antitumor substance in the Republic of Kazakstan. Previously, we have shown that this compound prevents protein farnesylation without altering geranylgeranylation. We now report that Arglabin DMA inhibits the incorporation of [(3)H]farnesylpyrophosphate into human H-ras protein by FTase with an IC(50) of no greater than 25 microM. Kinetic studies show that the phosphorylated form of this compound competitively inhibits the binding of farnesyl diphosphate to FTase. This mechanism of action is different from other reported peptidomimetic FTIs which lower the affinity of ras protein to FTase. Our in vitro studies confirm that Arglabin-DMA inhibits post translational modification of ras protein in cells. Arglabin-DMA inhibits anchorage-dependent proliferation of NB cells (IC50=10 microg/ml) and inhibits anchorage-independent growth of NB and KNRK cells with about the same IC(50). Soft-agar colony formation assay of H-ras and K-ras transformed cells show IC(50)s to be 2 and 5 microg/ml, respectively. In primary cultures of human tumor cells, Arglabin-DMA inhibits cell proliferation of a variety of tumor types with IC(90)s in the range of 0.85 to 5.0 microg/ml. Because of these pharmacologic properties, we propose that Arglabin-DMA is suitable for the treatment of ras related malignancies. PMID- 11115594 TI - Silicon diodes as detectors for backscatter radiation in photon fields. AB - We investigated the use of unencapsulated silicon semiconductor detectors for backscatter radiation detection. The results were compared with Monte Carlo (MC) calculations modelling the experimental set-up. A special diode was manufactured, which was designed so that it allowed the positioning of different materials in close contact with the detector surface. Polymethylmethacrylate (PMMA), Pb, Ti and Fe (stainless steel) were used as backscatter materials. The diode signal was measured by integrating the current when irradiating the diode with an equal photon fluence obtained from a medical Co-60 source. When compared to the signal with PMMA as backscatter material the increase in signal was 21%, 27% and 73% for Ti, Fe and Pb, respectively. This is in reasonable agreement with the MC calculations, when taking the effective measurement depth in the Si diode detector into account. PMID- 11115595 TI - Extended staging results from the detection of isolated tumor cells in the liver of colorectal cancer patients. AB - Liver metastasis, as well as local recurrence, are delineating factors of postoperative survival in patients suffering from colorectal cancer. We set up a PCR-RFLP assay to detect K-ras mutated cells in liver tissue as an indicator of possible isolated tumor cells (ITC) or micro-metastasis at the time of surgery. Sixty-four patients with K-ras codons 12 or 13 mutated colorectal cancer were clinically diagnosed for liver metastasis, as well as by PCR-RFLP assay of DNA from liver biopsies. Macro-metastasis was observed in the liver of 7 patients (11%), with no additional evidence of ITC. Likewise, in the liver of 14 patients (22%) only ITC, but no macro-metastasis was detected. In another 7 patients (11%) there was both, ITC and macro-metastasis. No macro-metastasis or ITC were found in 36 patients (56%). Thus, the PCR-RFLP assay added 14 cases (22%) with potential liver-metastasis to the 14 cases (22%) detected by clinical diagnostic means. T and N status were related to the refined detection and extended classification of liver involvement. We conclude that clinical and PCR-RFLP methods supplement each other and can increase the detection of cases with liver involvement, if applied together. PMID- 11115596 TI - Evolving features of lung adenocarcinoma in Rio de Janeiro, Brazil. AB - Formerly considered rare, adenocarcinoma has become the commonest form of primary lung cancer in developed countries. Its clinical presentation has changed, with central tumors becoming more frequent. We reviewed all biopsies with a diagnosis of primary lung cancer obtained from October 1988 to April 1997 in a tertiary care hospital in Rio de Janeiro. Medical records from adenocarcinomas were analysed. Adenocarcinoma was the commonest form of lung cancer (168/409, 41%). Central tumors were observed in 43% according to radiological criteria and 47% according to bronchoscopic criteria. The frequency and clinical presentation of adenocarcinoma have evolved lately in Rio de Janeiro. PMID- 11115597 TI - Expression of nm23-H1 in transitional cell carcinoma of the upper urinary tract. AB - Transitional cell carcinoma of the upper urinary tract is an uncommon neoplasm. Relatively little information is available regarding the clinical relevance of molecular markers. This study was performed to examine the importance of nm23-H1 gene expression (NM23-H1) in this type of tumors. Immunohistochemical expression of NM23-H1 was analyzed in 90 cases of upper urinary tract cancer, and was compared for its prognostic significance with conventional biological indicators. High expression of NM23-H1 was found in 7 cases (8%), intermediate expression in 32 cases (36%), and low expression in 51 cases (57%). Reduced NM23-H1 (defined as intermediate or low level of expression) was associated with a higher histological grading (p=0.002), invasive tumor growth (p=0. 002), or an increased proliferating cell nuclear antigen labeling index (p=0.004). NM23-H1 tended to inversely relate to later recurrence or long-term survival (p=0.06), but, only tumor staging was found to be significant in predicting clinical outcome (p=0.002). nm23-H1 appears to function as a tumor suppressor for upper urinary tract cancer, however, evaluation of NM23-H1 provides limited prognostic information. PMID- 11115598 TI - Mycosis fungoides and pregnancy. AB - Mycosis fungoides is a cutaneous T-cell lymphoma, a subgroup of non-Hodgkin's lymphomas, characterized by skin infiltration and occasionally systemic involvement. The association of pregnancy and mycosis fungoides has not been described previously. A case of mycosis fungoides, stage IVb, in a pregnant woman is reported. Prior to pregnancy, the patient received adriamycin, cyclophosphamide, vincristine prednisolone (CHOP) and bleomycin and total body irradiation. Around the concepcional period she presented a cutaneous relapse palliated with photon radiotherapy. No obstetrics complications were observed during gestation. At 39 week's gestation a cesarean section was performed and a healthy 2900 g boy was delivered. Mycosis fungoides did not worsen during pregnancy and postpartum period. In conclusion mycosis fungoides did not adversely affect pregnancy outcome and gestation did not worsen the malignancy course. This case report may be valuable in managing patients with mycosis fungoides who are currently pregnant or are contemplating pregnancy. PMID- 11115599 TI - The prognostic value of S-phase fraction in preoperative radiotherapy of rectal cancer. AB - The aim of this study was to analyze the flow cytometric S-phase fraction (SPF) in rectal tumors before and after preoperative radiotherapy (15x2 Gy) and to compare the findings to the clinical outcome. Archival specimens from 84 cases, treated from 1980 to 1988 with S-phase data and complete follow-up were reviewed. There was no significant correlation between SPF and clinicopathological factors. The median SPF for the 26 diploid tumors before irradiation was 6.6%+/-3.1, compared to a significantly higher median for the 58 preirradiated aneuploid tumors (20.3%+/-6.1; p<0.0001). With a median follow-up of 6 years, there was a significant difference in the number of recurrences for aneuploid tumors with a pretreatment SPF < and >20.3 (51.7% vs. 20.7%; p=0.029), which also led to a significant difference in recurrence-free survival (p=0.05). For diploid tumors, a reduction in the percentage of cells in S-phase after radiation resulted in a borderline significant lower number of recurrences (p=0.06). It is concluded that pretreatment S-phase measurements may be of predictive value especially for aneuploid tumors. An alteration in SPF after radiotherapy may also be helpful in predicting outcome and planning therapy. PMID- 11115601 TI - Matrix metalloproteinases in skin pathology (Review). AB - Human tissue is composed in part of cells and in part of amorphous matrix components. Equilibrium exists between the synthesis and degradation of connective tissue under physiological conditions, which serves both the formation and the maintenance of tissue architecture. Synthesis progresses via mesenchymal cells, degradation is controlled by the proteolytic effect of a group of enzymes, which belong to the protein family of matrix metalloproteinases. In biological development, matrix metalloproteinases play an important role in all essential configurative processes in embryo- and histogenesis. As a result of the importance of matrix metalloproteinases in creation of connective tissue, dysregulation with excessive proteolytic activity may result in tissue damage. Histological structural changes are set in relation to the molecular expression pattern of matrix metalloproteinases using matrix relevant diseases of human skin. Discussion includes fibrosing processes, skin inflammations, wound healing, blistering diseases, premature sun-induced skin aging and primary cutaneous malignomas and their metastases. PMID- 11115600 TI - Dynamics of circadian fluctuations in serum concentration of cortisol and TNF alpha soluble receptors in gastrointestinal cancer patients. AB - The objective of the study was to investigate the dynamic changes of cortisol and soluble TNF-alpha p-55 and p-75 receptors in serum of advanced cancer patients during 24 h. The examined group consisted of 42 patients suffering from advanced gastrointestinal neoplasms (colorectal, gastric and pancreatic cancer). Serum levels of cortisol, TNF-alpha and both receptors in cancer patients were measured using ELISA type kits 6 times a day (8.00 a.m., 2.00 p.m., 6.00 p.m., 10.00 p.m., 2.00 a.m. and again 8.00 a.m.) as well as in healthy controls. The levels of cortisol, TNF-alpha and its soluble receptors were substantially increased in examined group and displayed statistically significant circadian fluctuations. Cortisol peak was found typically at 8.08 a.m., the minimum value appeared at 6 p.m. The presence of circadian rhythm of the cytokine was proved (acrophase 00.36 a.m.), however no diurnal rhythm of soluble TNF receptors was observed. The concentration of p-55 receptor was distinctly lower then p-75. The peak p-55 value appeared at 10.00 p. m. while the p-75 reached its minimum level at the same time. Although there was no statistical correlation between the receptor concentrations the shapes of both curves remained in inversely proportional manner to each other. The present results may suggest the presence of complex self-regulation mechanisms in advanced gastrointestinal cancer patients. There was no correlation observed between cortisol and TNF-alpha soluble receptor concentration. PMID- 11115602 TI - Superoxide dismutase and pulmonary oxygen toxicity: lessons from transgenic and knockout mice (Review). AB - Superoxide (O2-) has been implicated in the pathogenesis of pulmonary O2 toxicity. The studies using transgenic and knockout mice of each of the three isoforms of superoxide dismutase (SOD) e.g. , CuZnSOD, MnSOD and extracellular SOD (EC-SOD), have demonstrated that O2- produced in the mitochondria from its electron transport system and extracellular O2- generated by infiltrating neutrophils, and possibly its derivatives e.g., hydroxyl radical and peroxynitrite, are important mediators of hyperoxia-induced pulmonary injury, while cytoplasmic O2- plays a limited, if any, role in the pathogenesis of pulmonary O2 toxicity. Distal airway epithelial cells including type II alveolar and non-ciliated bronchiolar epithelial cells, are important targets for O2 radicals under the hyperoxic condition. The accessibility of these distal airway epithelial cells to in vivo gene transfer through the tracheal route of administration, suggests the potential for in vivo transfer of MnSOD and EC-SOD genes as a future approach in the prevention of pulmonary O2 toxicity. PMID- 11115603 TI - Ryanodine-sensitive Ca2+ release mechanism in non-excitable cells (Review). AB - The properties of a ryanodine-sensitive Ca2+ release channel (receptor) in non excitable cells like exocrine cells or epithelial cells are described in this review. The ryanodine-sensitive Ca2+ release from the microsomal vesicles is activated by Ca2+, caffeine, ryanodine or cyclic ADP-ribose (cADPR) and is inhibited by ruthenium red or higher concentrations (> or =100 microM) of ryanodine. The properties are similar to those of excitable cells such as muscle cells or neuronal tissues. In some non-excitable cells, the Ca2+ release induced by caffeine, ryanodine or cADPR is stimulated by calmodulin (CaM) or FK506. Kd values of [3H]ryanodine binding to the receptor protein range from 6 to 17 nM and are similar to those of a high-affinity binding site in skeletal or cardiac muscle. Maximum binding capacities (Bmax) range from 40 to 620 fmol/ mg protein and are 10 approximately 200-fold lower than those for a high-affinity binding site in skeletal muscle. Caffeine, adenine nucleotide AMP-PCP, Mg2+, ruthenium red or FK506 affects the binding. In some non-excitable cells, the ryanodine receptor (RyR) isoform RyR2 or RyR3 is expressed and has been identified. However, unlike for excitable cells, information concerning the RyR proteins, including binding sites for modulators like CaM and phosphorylation sites has not yet been obtained. PMID- 11115604 TI - Neurotransmitter receptor variants and their influence on antipsychotic treatment (Review)). AB - Psychiatric treatment requires the use of drugs which are in many cases associated with severe side effects and inadequate response. In the past decade extensive research has investigated the link between gene alterations and treatment response and several strong associations have been reported. The identification of genes influencing treatment outcome will facilitate the pre treatment selection of the most beneficial drug according to an individual's pharmacogenetic profile. Recent advances indicate that this goal is achievable in the near future. The scope of this communication is to review all the recent findings in psychopharmacogenetics leading to the individualization of treatment. PMID- 11115605 TI - NFkappaB2 (p52) promoter activation via Notch signaling pathway in rheumatoid synoviocytes. AB - Rheumatoid synoviocytes produce inflammatory cytokines and exhibit strong proliferation activity, which cause severe cartilage destruction in the joints. Previously, we reported that NFkappaB, a transcriptional factor that activated by mitogenic signals, was activated in rheumatoid synoviocytes. In addition, we revealed that Notch-1, the transcriptional factor that is involved in the developmental stages, was abnormally activated in rheumatoid synoviocytes. In this study, as one of the roles of Notch-1 for the chronic inflammation in RA, we examined its implication to NFkappaB2 activation in rheumatoid synoviocytes. Western blotting analysis showed that NFkappaB2 expression was elevated in rheumatoid synoviocytes compared with patients with osteoarthritis studied as control. We then analyzed NFkappaB2 binding activity to the promoter and revealed that kappaB binding complexes to the NFkappaB2 promoter was elevated in rheumatoid synoviocytes. We analyzed implication of Notch-1 signaling pathway on NFkappaB2 activation, and found that Notch-1 made a complex with recombination binding protein Jkappa (RBPJkappa), a repressor for NFkappaB2 promoter, and blocked binding of RBPJkappa to the promoter. These results indicated that as one of the mechanisms for nuclear translocated Notch-1, complex formation with RBPJkappa induces blocking of the NFkappaB2 promoter suppression, which might cause NFkappaB2 promoter activation. PMID- 11115606 TI - In vivo gene electroporation in skeletal muscle with special reference to the duration of gene expression. AB - To determine the limits of the duration of in vivo transferred foreign gene expression, we conducted electroporation (EP), a powerful non-viral means of gene transfer for living animals, into skeletal muscle of rats and mice with a luciferase, GFP or erythropoietin (EPO)-encoding reporter plasmid. The luciferase reporter plasmid was used for optimization of EP conditions, while GFP and EPO plasmids were used for monitoring the duration of gene expression. In the rat, increased hematocrit levels were maintained for at least 9 weeks with approximately a 3-fold increase in plasma EPO protein concentration at 4 weeks post-transfection. In the mouse, the GFP plasmid transfer confirmed that the reporter gene expression lasted as long as 3 months post-transfection. By introducing the EPO gene in vivo in the mouse, increased hematocrit levels revealed that duration of reporter gene expression was at least 14.5 months after in vivo gene EP into skeletal muscle. These results implicate an excellent potential of in vivo gene EP, applicable to both experimental and therapeutic purposes. PMID- 11115607 TI - A detailed deletion map of chromosome 20 in human oral squamous cell carcinoma. AB - Loss of heterozygosity (LOH) on the long arm of chromosome 20 (20q) has been detected in several human cancers. However, little is known about LOH on chromosome 20 in oral squamous cell carcinoma (OSCC). To determine which loci of chromosome 20 were involved in OSCC tumorigenesis, 41 cases of OSCC were examined for LOH state on chromosome 20 at 17 microsatellite loci by PCR-LOH assay. LOH occurred in 41.5% of tumors in at least one locus. Among the 17 loci, D20S48 on 20p11.2 and RPN2 on 20q12-13.1 exhibited higher frequencies of LOH, 27.6% and 31.4%, respectively. The LOH incidence was significantly higher in tumors in which the primary site was on gingiva compared with other oral sites (p=0.012). Our results indicate that allelic deletions on 20q12-13.1 and 20p11.2 may play roles in OSCC carcinogenesis, and suggest that allelic deletions on 20q might have some relation with the primary site of OSCC. PMID- 11115608 TI - Effects of interaction between parvovirus minute virus of mice NS1 and coactivator CBP on NS1- and p53-transactivation. AB - The non-structural protein NS1, encoded by the parvovirus minute virus of mice (MVM), is a potent regulator of viral gene expression in addition to prominent roles in viral replication and cytopathic effects associated with parvoviral infection. Although NS1 involves the modulation of viral and cellular transcription, the primary activation mechanism of MVM NS1 remains unclear. In the present study, we show here that the coactivator CREB binding protein, CBP, could potentiate NS1-mediated transcription as measured on the P38 promoter, which drives expression of the MVM capsid genes. NS1 bound to the two related cysteine-histidine-rich regions of CBP, referred to as C/H1 and C/H3, the former of which has an antagonistic function to CBP upon the NS1-transactivation. Furthermore, NS1 inhibited the synergistic transactivation by CBP and p53. These findings suggested that CBP as a transcriptional coactivator is required for NS1 mediated viral and cellular transcription in parvovirus-infected cells, resulting in cell proliferation and differentiation to achieve its lytic cycle. PMID- 11115609 TI - Buffering action of endogenous nitric oxide on the adrenocortical secretagogue effect of endothelins in the rat. AB - The secretagogue effect of endothelins (ETs) on the rat adrenal cortex is mediated by the ETB receptor. ETB receptors are coupled with nitric oxide (NO) synthase (NOS), and NO is known to inhibit steroid-hormone secretion from adrenal cortex. We investigated whether ETB-mediated NO production interferes with the stimulatory action of ETs on rat adrenal cortex. The selective agonist of ETB receptor BQ-3020 concentration-dependently increased aldosterone secretion from dispersed zona glomerulosa (ZG) cells and corticosterone secretion from dispersed zona fasciculata-reticularis (ZF/R) cells, and the NOS inhibitor NG-nitro-L arginine methylester (L-NAME) potentiated the effect of BQ-3020 in a concentration-dependent manner. The guanylate cyclase inhibitor Ly-83583, at a concentration suppressing guanylin- and L-arginine-induced cyclic-GMP release from dispersed adrenocortical cells, did not affect the secretory response of ZG and ZF/R cells to BQ-3020. ET-1, an agonist of both ETA and ETB receptors, stimulated the release of both aldosterone and corticosterone by in situ perfused rat adrenal gland. This effect was potentiated by L-NAME and unaffected by Ly 83583. Collectively, our findings allow us to suggest that endogenous NO exerts in vivo and in vitro a cyclic-GMP-independent buffering action on the ETB receptor-mediated adrenocortical secretagogue action of ETs. PMID- 11115610 TI - In vivo gene electroporation confers nutritionally-regulated foreign gene expression in the liver. AB - Whether or not nutritionally-regulated foreign gene expression in vivo is achievable was examined in mouse liver after in vivo gene transfer by electroporation (EP). Electric pulses were applied to a left liver lobe immediately after injection of a luciferase reporter gene driven by the liver type phosphoenolpyruvate carboxykinase (PEPCK) gene promoter. Cooling treatments especially with solid carbon dioxide in the transfection site prior to the in vivo gene EP increased reporter gene expression by a factor of 100. Body bioluminescence imaging also confirmed strong expression of the in vivo transferred reporter gene in a transfected area of the liver. Fasting conferred a 13-fold increase in the reporter gene expression in vivo in the liver when driven by the liver-type PEPCK promoter, whereas virtually no induction was found either by the SV40 promoter or by the same PEPCK promoter in the muscle when the mice were fasted. The administration of cAMP mimicked the fasting-induced reporter gene expression by the PEPCK promoter in the liver of fed mice. These results implicate that nutritionally-regulated foreign gene expression in vivo is attainable at least locally in the liver by a simple and convenient non-viral gene EP method. PMID- 11115611 TI - The CD81 expression in liver in hepatocellular carcinoma. AB - The expression of CD81, a cell receptor for hepatitis C virus, was examined on cancer cells in hepatocellular carcinoma (HCC) C (n=29) to clarify its clinical role. CD81 was expressed on hepatocyte membrane in non-tumor portions in all patients, however, this was not stained on the cancer cell membranes in 6 of 29. Reverse transcriptase-PCR revealed that CD81 gene expression was not detected in the tumorous portions in 3 of 7 samples. The loss of CD81 was prominent in poorly differentiated HCC (5 of 8) and extrahepatic metastasis patients (6 of 8). The loss of CD81 is associated with differentiation and metastasis of HCC. PMID- 11115612 TI - Stimulatory effect of zinc and growth factor on bone protein component in newborn rats: enhancement with zinc and insulin-like growth factor-I. AB - The effect of zinc and growth factor on bone protein component in newborn rats was investigated. The characterization of protein component in the femoral diaphyseal and metaphyseal tissue of newborn rats (3-35 days old) was examined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The diaphyseal and metaphyseal tissues of 7 days-old rats were cultured for 24 or 48 h in a medium containing either vehicle, zinc sulfate (10-4 M) or dipicolinate (10-3 M), a chelator of zinc ion, in the presence or absence of insulin-like growth factor-I (IGF-I; 10-8 M) or transforming growth factor-beta (TFG-beta; 10 10 M) with an effective concentration. Many cellular protein molecules were present in the diaphyseal and metaphyseal tissues; potent bands were seen in protein molecules with about 66 and 46 kDa. Protein molecule of about 66 kDa was greatly released in the medium cultured with the diaphyseal and metaphyseal tissues. This protein in the medium was increased by culture with zinc, IGF-I or TGF-beta. Total protein content in the medium cultured with the diaphyseal and metaphyseal tissues was significantly increased in the presence of zinc, IGF-I or TGF-beta. The IGF-I-increased medium protein content was significantly enhanced by zinc. This enhancing effect was not seen in TGF-beta. Alkaline phosphatase activity and deoxyribonucleic acid (DNA) content in the diaphyseal and metaphyseal tissues was significantly increased by culture with zinc, IGF-I or TGF-beta. The effect of IGF-I was significantly enhanced by zinc, while it was not found in TGF-beta. The effect of IGF-I or TGF-beta in increasing the bone components was seen in the presence of dipicolinate. This study demonstrates that zinc, like IGF-I and TGF-beta, can increase protein components in the femoral diaphyseal and metaphyseal tissues of new-born rats. Zinc may especially play a role in bone growth in collaboration with IGF-I. PMID- 11115613 TI - Anti-inflammatory effect of the aqueous extract from Lonicera japonica flower is related to inhibition of NF-kappaB activation through reducing I-kappaBalpha degradation in rat liver. AB - We investigated the anti-inflammatory effects of aqueous extract from Lonicera japonica flower (AELJ), a traditional skin rash drug, in lipopolysaccharide (LPS) induced rat liver sepsis. Immunoblot analysis showed that the level of nuclear factor (NF)-kappaBp65 was rapidly up-regulated and inhibitory (I)-kappaBalpha was down-regulated by LPS challenge. However, AELJ inhibited the increase of NF kappaBp65 and degradation of I-kappaBalpha in the liver of LPS-challenged rats. Immunohistochemical analysis of rat hepatocytes showed that LPS-induced inflammatory responses, involving degradation of I-kappaBalpha and induction of NF-kappaBp65, tumor necrosis factor (TNF)-alpha and inducible nitric oxide synthase (iNOS), are partially inhibited by pretreatment with AELJ. These results suggest that AELJ may act as a therapeutic agent for inflammatory disease through a selective regulation of NF-kappaB activation. PMID- 11115614 TI - Exclusion of candidate genes and chromosomal regions in familial neuroblastoma. AB - Two families with recurrence of neuroblastoma one Italian and one British with three and two affected children respectively were genotyped using polymorphic markers on chromosome 1 spanning the p32-p36 region frequently deleted in neuroblastoma tumor cells. Linkage to this region was excluded by haplotype inspection and negative lod scores. Furthermore, the exclusion of genes involved in neurocristopathies sometimes associated with neuroblastoma was carried out by typing the Italian family with polymorphic markers located in or near the corresponding genes. Finally, linkage analysis in the two families showed negative lod scores for markers spanning the 16p12-13 chromosomal region where a locus for familial neuroblastoma has been recently mapped. Our findings indicate that different genes are involved in the pathogenesis of familial neuroblastoma. PMID- 11115615 TI - Hypoglycemia-induced VEGF expression is mediated by intracellular Ca2+ and protein kinase C signaling pathway in HepG2 human hepatoblastoma cells. AB - Vascular endothelial growth factor (VEGF) is a potent angiogenic factor that plays a central role in angiogenesis. In this study, we investigated the mechanism of VEGF expression in HepG2 human hepatoblastoma cells under hypoglycemia. The shortage of glucose significantly enhanced VEGF mRNA expression in a time-dependent manner as well as increased DNA-binding activity of AP-1 that plays an important role in VEGF transcription. In addition, treatment of a potent PKC inhibitor, H-7 in glucose-deprived HepG2 cells suppressed hypoglycemia elevated VEGF expression as well as the increased AP-1 DNA-binding activity. Moreover, we observed that Ca2+ levels remarkably increased under low glucose condition. Consistently, an intracellular Ca2+ chelator, BAPTA/AM significantly decreased hypoglycemia-induced VEGF expression and AP-1 DNA-binding activity. Therefore, these results indicate that increase of intracellular Ca2+ level induces the activation of PKC, which induce the activation of AP-1 leading to the increase of VEGF in glucose-deprived environment. Furthermore, it provides one link in regulation of VEGF with hypoglycemia as well as information to understand how hypoglycemia induces VEGF expression and subsequently leads to tumor angiogenesis. PMID- 11115616 TI - Therapy of HPV 16-associated carcinoma with dendritic cell-based vaccines: in vitro priming of the effector cell responses by DC pulsed with tumour lysates and synthetic RAHYNIVTF peptide. AB - Murine carcinoma induced by MK 16 cells expressing HPV 16 E6/E7 oncogenes was utilized to examine the therapeutic effect of dendritic cell-based tumour vaccines. Mice carrying 5-day MK 16 tumours were injected peritumorally with either dendritic cells (DC) or DC pulsed with MK 16 tumour lysate. Both the unpulsed and MK 16 lysate-pulsed DC vaccines inhibited growth of the MK 16 transplants, the pulsed DC being more efficient than the unpulsed vaccines. In vitro priming of the effector cell-mediated anti-MK 16 responses by DC pulsed with MK 16 tumour lysate and a synthetic HPV 16 E7(49-57) peptide RAHYNIVTF was compared. The priming activity of the lysate was substantially higher than that of the HPV 16 E7(49-57) peptide; the priming activity was similar to that of a standard moderately immunogenic chemically-induced sarcoma. Taken collectively, these results suggest that DC vaccines pulsed with HPV 16-associated tumour lysates represent a prospective modality for treatment of HPV 16-associated carcinomas. PMID- 11115618 TI - Expression of metalloproteinases and its inhibitor in later stage of rabbit neointima development. AB - Neointima formation after arterial de-endothelialization refers not only to smooth muscle cell (SMC) migration and proliferation, but also involves extracellular matrix (ECM) metabolism. Most studies regarding the role of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in neointima have focused on the early phase of vascular remodeling. In this study, we examined the expression of MMP and TIMP in rabbit aortic neointima at a relatively late stage of lesion development, between 4 and 12 weeks after initial de-endothelialization. Northern blot analysis revealed expression of steady-state MMP-9 mRNA was increased up to the 4th week and MMP-2 mRNA to the 12th week after de-endothelialization. In situ hybridization shown that MMP positive cells were predominantly distributed in arterial neointima. Expression of TIMP-1 mRNA was continuously up-regulated up to the 12th week and TIMP-1 positive cells, primarily SMCs, were also localized to the neointimal tissue. Alteration at mRNA level was accompanied by that at protein level, as assessed by SDS-PAGE zymography for MMPs and immunoblotting for TIMP-1. The profile of alteration at protein level correlated well with that at mRNA level. These data suggest that synthesis of MMPs and TIMP is a prolonged process and arterial SMC is a major source of MMP production in arterial neointima. Enhanced synthesis of MMPs and TIMPs at late stage of neointimal development may contribute to arterial ECM metabolism. PMID- 11115617 TI - Zaltoprofen prevents carbon tetrachloride-induced reduction of body weight in rats. AB - Previously we reported carbon tetrachloride-induced body weight loss in rats as a new model of wasting disorders. The oral administration of a low dose of carbon tetrachloride to rats reduced the body weight and food intake at 24 h with a minimal effect on plasma alanine aminotransferase activity. Tumor necrosis factor alpha, and cyclooxygenase-1 and -2 mRNA expression in the brain was not affected by carbon tetrachloride. Zaltoprofen, which is a non-steroidal anti-inflammatory drug, prevented the carbon tetrachloride-induced body weight decrease, without preventing the carbon tetrachloride-induced loss of food intake. The present results suggest the possible application of this drug for the treatment of wasting disorders. PMID- 11115619 TI - Photosynthetic water oxidation. PMID- 11115620 TI - Remembering Melvin P. Klein (1921-2000). PMID- 11115622 TI - The tetra-manganese complex of photosystem II during its redox cycle - X-ray absorption results and mechanistic implications. AB - Using X-ray absorption spectroscopy (XAS), relevant information on structure and oxidation state of the water-oxidizing Mn complex of photosystem II has been obtained for all four semi-stable intermediate states of its catalytic cycle. We summarize our recent XAS results and discuss their mechanistic implications. The following aspects are covered: (a) information content of X-ray spectra (pre-edge feature, edge position, extended X-ray absorption fine-structure (EXAFS), dichroism in the EXAFS of partially oriented samples); (b) S(1)-state structure; (c) X-ray edge results on oxidation state changes; (d) EXAFS results on structural changes during the S-state cycle; (e) a structural model for the Mn complex in its S(3)-state; (f) XAS-based working model for the S(2)-S(3) transition; (g) XAS-based working model for the S(0)-S(1) transition; (h) potential role of hydrogen atom abstraction by the Mn complex. Finally, we present a specific hypothesis on the mechanism of dioxygen formation during the S(3)-(S(4))-S(0) transition. According to this hypothesis, water oxidation is facilitated by manganese reduction that is coupled to proton transfer from a substrate water to bridging oxides. PMID- 11115621 TI - X-ray spectroscopy-based structure of the Mn cluster and mechanism of photosynthetic oxygen evolution. AB - The mechanism by which the Mn-containing oxygen evolving complex (OEC) produces oxygen from water has been of great interest for over 40 years. This review focuses on how X-ray spectroscopy has provided important information about the structure of this Mn complex and its intermediates, or S-states, in the water oxidation cycle. X-ray absorption near-edge structure spectroscopy and high resolution Mn Kbeta X-ray emission spectroscopy experiments have identified the oxidation states of the Mn in the OEC in each of the intermediate S-states, while extended X-ray absorption fine structure experiments have shown that 2.7 A Mn-Mn di-mu-oxo and 3.3 A Mn-Mn mono-mu-oxo motifs are present in the OEC. X-ray spectroscopy has also been used to probe the two essential cofactors in the OEC, Ca2+ and Cl-, and has shown that Ca2+ is an integral component of the OEC and is proximal to Mn. In addition, dichroism studies on oriented PS II membranes have provided angular information about the Mn-Mn and Mn-Ca vectors. Based on these X ray spectroscopy data, refined models for the structure of the OEC and a mechanism for oxygen evolution by the OEC are presented. PMID- 11115623 TI - Metallo-radical hypothesis for photoassembly of (Mn)4-cluster of photosynthetic oxygen evolving complex. AB - A new hypothetical mechanism is proposed for photoassembly of the (Mn)4-cluster of the photosynthetic oxygen evolving complex (OEC). In this process, a neutral radical of Y(Z) tyrosine plays a role in oxidizing Mn2+ associated with an apo OEC, and also in abstracting a proton from a water molecule bound to the Mn2+ ion, together with D1-His190. This is in a similar fashion to the metallo-radical mechanism proposed for photosynthetic water oxidation by the (Mn)4-cluster. The model insists that a common mechanism participates in the photoassembly of the (Mn)4-cluster and the photosynthetic water oxidation. PMID- 11115624 TI - The inorganic biochemistry of photosynthetic oxygen evolution/water oxidation. AB - At the request of the organizer of this special edition, we have attempted to do several things in this manuscript: (1) we present a mini-review of recent, selected, works on the light-induced inorganic biogenesis (photoactivation), composition and structure of the inorganic core responsible for photosynthetic water oxidation; (2) we summarize a new proposal for the evolutionary origin of the water oxidation catalyst which postulates a key role for bicarbonate in formation of the inorganic core; (3) we summarize published studies and present new results on what has been learned from studies of 'inorganic mutants' in which the endogenous cofactors (Mn(n+), Ca2+, Cl-) are substituted; (4) the first DeltapH changes measured during the photoactivation process are reported and used to develop a model for the stepwise photo-assembly process; (5) a comparative analysis is given of data in the literature on the kinetics of substrate water exchange and peroxide binding/dismutation which support a mechanistic model for water oxidation in general; (6) we discuss alternative interpretations of data in the literature with a view to forecast new avenues where progress is needed. PMID- 11115625 TI - Vibrational spectroscopy of the oxygen-evolving complex and of manganese model compounds. AB - A number of molecularly specific models for the oxygen-evolving complex in photosystem II (PSII) and of manganese-substrate water intermediates that may occur in this process have been proposed recently. We summarize this work briefly. Fourier transform infrared techniques have emerged as fruitful tools to study the molecular structures of Y(Z) and the manganese complex. We discuss recent work in which mid-IR (1000-2000 cm(-1)) methods have been used in this effort. The low-frequency IR region (<1000 cm(-1)) has been more difficult to access for technical reasons, but good progress has been made in overcoming these obstacles. We update recent low-frequency work on PSII and then present a detailed summary of relevant manganese model compounds that will be of importance in understanding the emerging biological data. PMID- 11115626 TI - Comparative studies of the S0 and S2 multiline electron paramagnetic resonance signals from the manganese cluster in Photosystem II. AB - Electron paramagnetic resonance (EPR) spectroscopy is one of the major techniques used to analyse the structure and function of the water oxidising complex (WOC) in Photosystem II. The discovery of an EPR signal from the S0 state has opened the way for new experiments, aiming to characterise the S0 state and elucidate the differences between the different S states. We present a review of the biochemical and biophysical characterisation of the S0 state multiline signal that has evolved since its discovery, and compare these results to previous and recent data from the S2 multiline signal. We also present some new data from the S2 state reached on the second turnover of the enzyme. PMID- 11115627 TI - EPR/ENDOR characterization of the physical and electronic structure of the OEC Mn cluster. AB - Electron paramagnetic resonance (EPR) spectroscopy has often played a crucial role in characterizing the various cofactors and processes of photosynthesis, and photosystem II and its oxygen evolving chemistry is no exception. Until recently, the application of EPR spectroscopy to the characterization of the oxygen evolving complex (OEC) has been limited to the S2-state of the Kok cycle. However, in the past few years, continuous wave-EPR signals have been obtained for both the S0- and S1-state as well as for the S2 (radical)(Z)-state of a number of inhibited systems. Furthermore, the pulsed EPR technique of electron spin echo electron nuclear double resonance spectroscopy has been used to directly probe the 55Mn nuclei of the manganese cluster. In this review, we discuss how the EPR data obtained from each of these states of the OEC Kok cycle are being used to provide insight into the physical and electronic structure of the manganese cluster and its interaction with the key tyrosine, Y(Z). PMID- 11115628 TI - EPR studies of the water oxidizing complex in the S1 and the higher S states: the manganese cluster and Y(Z) radical. AB - The parallel polarization electron paramagnetic resonance (EPR) method has been applied to investigate manganese EPR signals of native S1 and S3 states of the water oxidizing complex (WOC) in photosystem (PS) II. The EPR signals in both states were assigned to thermally excited states with S=1, from which zero-field interaction parameters D and E were derived. Three kinds of signals, the doublet signal, the singlet-like signal and g=11-15 signal, were detected in Ca2+ depleted PS II. The g=11-15 signal was observed by parallel and perpendicular modes and assigned to a higher oxidation state beyond S2 in Ca2+-depleted PS II. The singlet-like signal was associated with the g=11-15 signal but not with the Y(Z) (the tyrosine residue 161 of the D1 polypeptide in PS II) radical. The doublet signal was associated with the Y(Z) radical as proved by pulsed electron nuclear double resonance (ENDOR) and ENDOR-induced EPR. The electron transfer mechanism relevant to the role of Y(Z) radical was discussed. PMID- 11115629 TI - Probing the Mn oxidation states in the OEC. Insights from spectroscopic, computational and kinetic data. AB - Results from a variety of experimental techniques which have been used to define the oxidation levels of Mn and other components in the S states of the water oxidising complex in Photosystem II are reviewed. A self-consistent interpretation of Mn X-ray absorption near edge spectroscopy, UV-visible and near infrared spectroscopic data suggests that Mn oxidation occurs only on the S0-->S1 transition, and that all four Mn centres have formal oxidation state III thereafter. Ligand oxidation occurs on the transitions to S2 and S3. This is supported by high level quantum chemical calculations and an analysis of the kinetics of substrate water exchange, as recently determined by Wydrzynski et al. (this journal). One type of model for the catalytic site structure and water oxidation mechanism, consistent with these conclusions, is discussed. This model invokes magnetically separate oxo bridged dimers with water oxidation occurring by a concerted 2H+/2e- transfer mechanism, with one H transfer to a bridge oxygen on each dimer. PMID- 11115630 TI - Photosynthetic water oxidation: towards a mechanism. AB - This mini-review outlines the current theories on the mechanism of electron transfer from water to P680, the location and structure of the water oxidising complex and the role of the manganese cluster. We discuss how our data fit in with current theories and put forward our ideas on the location and mechanism of water oxidation. PMID- 11115631 TI - Amino acid residues involved in the coordination and assembly of the manganese cluster of photosystem II. Proton-coupled electron transport of the redox-active tyrosines and its relationship to water oxidation. AB - The combination of site-directed mutagenesis, isotopic labeling, new magnetic resonance techniques and optical spectroscopic methods have provided new insights into cofactor coordination and into the mechanism of electron transport and proton-coupled electron transport in photosystem II. Site-directed mutations in the D1 polypeptide of this photosystem have implicated a number of histidine and carboxylate residues in the coordination and assembly of the manganese cluster, responsible for photosynthetic water oxidation. Many of these are located in the carboxy-terminal region of this polypeptide close to the processing site involved in its maturation. This maturation is a required precondition for cluster assembly. Recent proposals for the mechanism of water oxidation have directly implicated redox-active tyrosine Y(Z) in this mechanism and have emphasized the importance of the coupling of proton and electron transfer in the reduction of Y(Z)(radical) by the Mn cluster. The interaction of both homologous redox-active tyrosines Y(Z) and Y(D) with their respective homologous proton acceptors is discussed in an effort to better understand the significance of such coupling. PMID- 11115632 TI - Amino acid residues that modulate the properties of tyrosine Y(Z) and the manganese cluster in the water oxidizing complex of photosystem II. AB - The catalytic site for photosynthetic water oxidation is embedded in a protein matrix consisting of nearly 30 different polypeptides. Residues from several of these polypeptides modulate the properties of the tetrameric Mn cluster and the redox-active tyrosine residue, Y(Z), that are located at the catalytic site. However, most or all of the residues that interact directly with Y(Z) and the Mn cluster appear to be contributed by the D1 polypeptide. This review summarizes our knowledge of the environments of Y(Z) and the Mn cluster as obtained from the introduction of site-directed, deletion, and other mutations into the photosystem II polypeptides of the cyanobacterium Synechocystis sp. PCC 6803 and the green alga Chlamydomonas reinhardtii. PMID- 11115633 TI - Bicarbonate requirement for the water-oxidizing complex of photosystem II. AB - It is well established that bicarbonate stimulates electron transfer between the primary and secondary electron acceptors, Q(A) and Q(B), in formate-inhibited photosystem II; the non-heme Fe between Q(A) and Q(B) plays an essential role in the bicarbonate binding. Strong evidence of a bicarbonate requirement for the water-oxidizing complex (WOC), both O2 evolving and assembling from apo-WOC and Mn2+, of photosystem II (PSII) preparations has been presented in a number of publications during the last 5 years. The following explanations for the involvement of bicarbonate in the events on the donor side of PSII are considered: (1) bicarbonate serves as an electron donor (alternative to water or as a way of involvement of water molecules in the oxidative reactions) to the Mn containing O2 center; (2) bicarbonate facilitates reassembly of the WOC from apo WOC and Mn2+ due to formation of the complexes MnHCO3+ and Mn(HCO3)2 leading to an easier oxidation of Mn2+ with PSII; (3) bicarbonate is an integral component of the WOC essential for its function and stability; it may be considered a direct ligand to the Mn cluster; (4) the WOC is stabilized by bicarbonate through its binding to other components of PSII. PMID- 11115634 TI - Oxygen ligand exchange at metal sites - implications for the O2 evolving mechanism of photosystem II. AB - The mechanism for photosynthetic O2 evolution by photosystem II is currently a topic of intense debate. Important questions remain as to what is the nature of the binding sites for the substrate water and how does the O-O bond form. Recent measurements of the 18O exchange between the solvent water and the photogenerated O2 as a function of the S-state cycle have provided some surprising insights to these questions (W. Hillier, T. Wydrzynski, Biochemistry 39 (2000) 4399-4405). The results show that one substrate water molecule is bound at the beginning of the catalytic sequence, in the S0 state, while the second substrate water molecule binds in the S3 state or possibly earlier. It may be that the second substrate water molecule only enters the catalytic sequence following the formation of the S3 state. Most importantly, comparison of the observed exchange rates with oxygen ligand exchange in various metal complexes reveal that the two substrate water molecules are most likely bound to separate Mn(III) ions, which do not undergo metal-centered oxidations through to the S3 state. The implication of this analysis is that in the S1 state, all four Mn ions are in the +3 oxidation state. This minireview summarizes the arguments for this proposal. PMID- 11115635 TI - Photosynthetic water oxidation to molecular oxygen: apparatus and mechanism. PMID- 11115636 TI - Mechanism of photosynthetic water oxidation: combining biophysical studies of photosystem II with inorganic model chemistry. AB - A mechanism for photosynthetic water oxidation is proposed based on a structural model of the oxygen-evolving complex (OEC) and its placement into the modeled structure of the D1/D2 core of photosystem II. The structural model of the OEC satisfies many of the geometrical constraints imposed by spectroscopic and biophysical results. The model includes the tetranuclear manganese cluster, calcium, chloride, tyrosine Z, H190, D170, H332 and H337 of the D1 polypeptide and is patterned after the reversible O2-binding diferric site in oxyhemerythrin. The mechanism for water oxidation readily follows from the structural model. Concerted proton-coupled electron transfer in the S2-->S3 and S3-->S4 transitions forms a terminal Mn(V)=O moiety. Nucleophilic attack on this electron-deficient Mn(V)=O by a calcium-bound water molecule results in a Mn(III)-OOH species, similar to the ferric hydroperoxide in oxyhemerythrin. Dioxygen is released in a manner analogous to that in oxyhemerythrin, concomitant with reduction of manganese and protonation of a mu-oxo bridge. PMID- 11115637 TI - Coupling of electron and proton transfer in the photosynthetic water oxidase. AB - According to current estimates, the photosynthetic water oxidase functions with a quite restricted driving force. This emphasizes the importance of the catalytic mechanisms in this enzyme. The general problem of coupling electron and proton transfer is discussed from this viewpoint and it is argued that 'weak coupling' is preferable to 'strong coupling'. Weak coupling can be achieved by facilitating deprotonation either before (proton-first path) or after (electron-first path) the oxidation step. The proton-first path is probably relevant to the oxidation of tyrosine Y(Z) by P-680. Histidine D1-190 is believed to play a key role as a proton acceptor facilitating Y(Z) deprotonation. The pK(a) of an efficient proton acceptor is submitted to conflicting requirements, since a high pK(a) favors proton transfer from the donor, but also from the medium. H-bonding between Y(Z) and His, together with the Coulombic interaction between negative tyrosinate and positive imidazolium, are suggested to play a decisive role in alleviating these constraints. Current data and concepts on the coupling of electron and proton transfer in the water oxidase are discussed. PMID- 11115638 TI - Kinetics of electron and proton transfer during O(2) reduction in cytochrome aa(3) from A. ambivalens: an enzyme lacking Glu(I-286). AB - Acidianus ambivalens is a hyperthermoacidophilic archaeon which grows optimally at approximately 80 degrees C and pH 2.5. The terminal oxidase of its respiratory system is a membrane-bound quinol oxidase (cytochrome aa(3)) which belongs to the heme-copper oxidase superfamily. One difference between this quinol oxidase and a majority of the other members of this family is that it lacks the highly conserved glutamate (Glu(I-286), E. coli ubiquinol oxidase numbering) which has been shown to play a central role in controlling the proton transfer during reaction of reduced oxidases with oxygen. In this study we have investigated the dynamics of the reaction of the reduced A. ambivalens quinol oxidase with O(2). With the purified enzyme, two kinetic phases were observed with rate constants of 1.8&z.ccirf;10(4) s(-1) (at 1 mM O(2), pH 7.8) and 3. 7x10(3) s(-1), respectively. The first phase is attributed to binding of O(2) to heme a(3) and oxidation of both hemes forming the 'peroxy' intermediate. The second phase was associated with proton uptake from solution and it is attributed to formation of the 'oxo-ferryl' state, the final state in the absence of quinol. In the presence of bound caldariella quinol (QH(2)), heme a was re-reduced by QH(2) with a rate of 670 s(-1), followed by transfer of the fourth electron to the binuclear center with a rate of 50 s(-1). Thus, the results indicate that the quinol donates electrons to heme a, followed by intramolecular transfer to the binuclear center. Moreover, the overall electron and proton-transfer kinetics in the A. ambivalens quinol oxidase are the same as those in the E. coli ubiquinol oxidase, which indicates that in the A. ambivalens enzyme a different pathway is used for proton transfer to the binuclear center and/or other protonatable groups in an equivalent pathway are involved. Potential candidates in that pathway are two glutamates at positions (I-80) and (I-83) in the A. ambivalens enzyme (corresponding to Met(I-116) and Val(I-119), respectively, in E. coli cytochrome bo(3)). PMID- 11115639 TI - Electron pathways involved in H(2)-metabolism in the green alga Scenedesmus obliquus. AB - The green alga Scenedesmus obliquus is capable of both uptake and production of H(2) after anaerobic adaptation (photoreduction of CO(2) or photohydrogen production). The essential enzyme for H(2)-metabolism is a NiFe-hydrogenase with a [2Fe-2S]-ferredoxin as its natural redox partner. Western blot analysis showed that the hydrogenase is constitutively expressed. The K(m) values were 79.5 microM and 12.5 microM, determined with ferredoxin and H(2), respectively, as electron donor for the hydrogenase. In vitro, NADP(+) was reduced by H(2) in the presence of the hydrogenase, the ferredoxin and a ferredoxin-NADP reductase. From these results and considerations on the stoichiometry we propose that this light independent electron transfer is part of the photoreduction of CO(2) in vivo. For ATP synthesis, necessary for the photoreduction of CO(2), light-dependent cyclic electron transfer around Photosystem (PS) I accompanies this 'dark reaction'. PS II fluorescence data suggest that (a) in S. obliquus H(2)-reduction might function as the anaerobic counterpart of the O(2)-dependent Mehler reaction, and (b) the presence of either a ferredoxin quinone-reductase or NAD(P)-dehydrogenase (complex I) in S. obliquus chloroplasts. PMID- 11115640 TI - A novel hydrophobic diheme c-type cytochrome. Purification from Corynebacterium glutamicum and analysis of the QcrCBA operon encoding three subunit proteins of a putative cytochrome reductase complex. AB - Electrophoresis of a Corynebacterium glutamicum membrane preparation in the presence of sodium dodecyl sulfate, followed by staining for peroxidase activity (heme staining), showed only one band at about 28 kDa. This 28 kDa protein was purified from C. glutamicum membranes by chromatography in the presence of decylglucoside using DEAE-Toyopearl and hydroxylapatite columns, as the sole c type cytochrome in the bacterium. The cytochrome showed an alpha band at 551 nm, and its E(m, 7) was about 210 mV. A QcrCAB operon encoding the subunits of a putative quinol cytochrome c reductase was found 3'-downstream of ctaE encoding subunit III of cytochrome aa(3) in the C. glutamicum genome. The deduced amino acid sequence of qcrC, composed of 283 amino acid residues, contained two heme C binding motifs and was in agreement with partial peptide sequences obtained from the 28 kDa protein after V8 protease digestion. We propose to name this protein cytochrome cc. The presence of cytochrome cc is a common feature of high G+C content Gram-positive bacteria, since we could confirm this protein by electrophoresis; homologous QcrCAB operons are also known in Mycobacterium and Streptomyces. QcrA and qcrB of C. glutamicum encode the Rieske Fe-S protein and cytochrome b, respectively, although these proteins were not co-purified with cytochrome cc. The phylogenetic tree of cytochromes b and b(6) show that C. glutamicum cytochrome b, along with those of other bacteria in the high G+C group, is rather different from the Bacillus counterparts, but highly similar to the Deinococci and Thermus cytochromes. This indicates that there is a fourth group of bacteria in addition to the three clades: proteobacterial cytochrome b, cyanobacterial b(6) and green sulfur-low G+C Gram-positive bacteria. PMID- 11115641 TI - Effect of 13-cis violaxanthin on organization of light harvesting complex II in monomolecular layers. AB - Lutein, neoxanthin and violaxanthin are the main xanthophyll pigment constituents of the largest light-harvesting pigment-protein complex of photosystem II (LHCII). High performance liquid chromatography analysis revealed photoisomerization of LHCII-bound violaxanthin from the conformation all-trans to the conformation 13-cis and 9-cis. Maximally, the conversion of 15% of all-trans violaxanthin to a cis form could be achieved owing to the light-driven reactions. The reactions were dark-reversible. The all-trans to cis isomerization was found to be driven by blue light, absorbed by chlorophylls and carotenoids, as well as by red light, absorbed exclusively by chlorophyll pigments. This suggests that the photoisomerization is a carotenoid triplet-sensitized reaction. The monomolecular layer technique was applied to study the effect of the 13-cis conformer of violaxanthin and its de-epoxidized form, zeaxanthin, on the organization of LHCII as compared to the all-trans stereoisomers. The specific molecular areas of LHCII in the two-component system composed of protein and exogenous 13-cis violaxanthin or 13-cis zeaxanthin show overadditivity, which is an indication of the xanthophyll-induced disassembly of the aggregated forms of the protein. Such an effect was not observed in the monomolecular layers of LHCII containing all-trans conformers of violaxanthin and zeaxanthin. 77 K chlorophyll a fluorescence emission spectra recorded from the Langmuir-Blodgett (L-B) films deposited to quartz from monomolecular layers formed with LHCII and LHCII in the two-component systems with all-trans and 13-cis isomers of violaxanthin and zeaxanthin revealed opposite effects of both conformers on the aggregation of the protein. The cis isomers of both xanthophylls were found to decrease the aggregation level of LHCII and the all-trans isomers increased the aggregation level. The calculated efficiency of excitation energy transfer to chlorophyll a from violaxanthin assumed to remain in two steric conformations was analyzed on the basis of the chlorophyll a fluorescence excitation spectra and the mean orientation of violaxanthin molecules in LHCII (71 degrees with respect to the normal to the membrane), determined recently in the linear dichroism experiments [Gruszecki et al., Biochim. Biophys. Acta 1412 (1999) 173-183]. The calculated efficiency of excitation energy transfer from the violaxanthin pool assumed to remain in conformation all-trans was found to be almost independent on the orientation angle within a variability range. In contrast the calculated efficiency of energy transfer from the form cis was found to be strongly dependent on the orientation and varied between 1.0 (at 67.48 degrees ) and 0 (at 70.89 degrees ). This is consistent with two essentially different, possible functions of the cis forms of violaxanthin: as a highly efficient excitation donor (and possibly energy transmitter between other chromophores) or purely as a LHCII structure modifier. PMID- 11115642 TI - Kinetic mechanism of antiports catalyzed by reconstituted ornithine/citrulline carrier from rat liver mitochondria. AB - The transport mechanism of the reconstituted ornithine/citrulline carrier purified from rat liver mitochondria was investigated kinetically. A complete set of half-saturation constants (K(m)) was established for ornithine, citrulline and H(+) on both the external and internal side of the liposomal membrane. The internal affinity for ornithine was much lower than that determined on the external surface. The exclusive presence of a single transport affinity for ornithine on each side of the membrane indicated a unidirectional insertion of the ornithine/citrulline carrier into liposomes, probably right-side-out with respect to mitochondria. Two-reactant initial velocity studies of the homologous (ornithine/ornithine) and heterologous (ornithine/citrulline) exchange reactions resulted in a kinetic pattern which is characteristic of a simultaneous antiport mechanism. This type of mechanism implies that the carrier forms a ternary complex with the substrates before the transport reaction occurs. A quantitative analysis of substrate interaction revealed that rapid-equilibrium random conditions were fulfilled, characterized by a fast and independent binding of internal and external substrates. PMID- 11115643 TI - Nucleotide effects on liver and muscle mitochondrial non-phosphorylating respiration and membrane potential. AB - Uncoupling protein-1 homologs are hypothesized to mediate mitochondrial proton leak. To test this hypothesis, we determined the effects of ATP and other nucleotides on liver and skeletal muscle mitochondrial non-phosphorylating respiration (VO(2)), membrane potential, FCCP-stimulated respiratory control ratios, and swelling. Neither ATP nor CTP affected liver or muscle proton leak, but both inhibited the respiratory chain. Unexpectedly, CMP stimulated liver proton leak (EC(50) approximately 4.4+/-0.5 mM). Using CMP chromatography, we identified two proteins (M(r)=31.2 and 32.6 kDa) from liver mitochondria that are similar in size to members of the mitochondrial carrier protein family. We conclude (a) liver and muscle mitochondrial proton leak is insensitive to ATP and CTP, and (b) CMP activates a leak in liver mitochondria. The CMP-inducible leak may be mediated by a 30-32 kDa protein. Based on the high concentrations required, CMP is unlikely to be a physiologically important leak regulator. Nonetheless, our results show that tissues other than brown fat have inducible leaks that may be protein-mediated. PMID- 11115644 TI - The calorimetric-respirometric ratio is an on-line marker of enthalpy efficiency of yeast cells growing on a non-fermentable carbon source. AB - Although on-line calorimetry has been widely used to detect transitions in global metabolic activity during the growth of microorganisms, the relationships between oxygen consumption flux and heat production are poorly documented. In this work, we developed a respirometric and calorimetric approach to determine the enthalpy efficiency of respiration-linked energy transformation of isolated yeast mitochondria and yeast cells under growing and resting conditions. On isolated mitochondria, the analysis of different phosphorylating and non-phosphorylating steady states clearly showed that the simultaneous measurements of heat production and oxygen consumption rates can lead to the determination of both the enthalpy efficiency and the ATP/O yield of oxidative phosphorylation. However, these determinations were made possible only when the net enthalpy change associated with the phosphorylating system was different from zero. On whole yeast cells, it is shown that the simultaneous steady state measurements of the heat production and oxygen consumption rates allow the enthalpy growth efficiency (i.e. the amount of energy conserved as biomass compared to the energy utilised for complete catabolism plus anabolism) to be assessed. This method is based on the comparison between the calorimetric-respirometric ratio (CR ratio) determined under growth versus resting conditions during a purely aerobic metabolism. Therefore, in contrast to the enthalpy balance approach, this method does not rely on the exhaustive and tedious determinations of the metabolites and elemental composition of biomass. Thus, experiments can be performed in the presence of non-limiting amounts of carbon substrate, an approach which has been successfully applied to slow growing cells such as yeast cells expressing wild type or a mutant rat uncoupling protein-1. PMID- 11115645 TI - Nitrogen deprivation strongly affects photosystem II but not phycoerythrin level in the divinyl-chlorophyll b-containing cyanobacterium Prochlorococcus marinus. AB - Effects of nitrogen limitation on Photosystem II (PSII) activities and on phycoerythrin were studied in batch cultures of the marine oxyphotobacterium Prochlorococcus marinus. Dramatic decreases in photochemical quantum yields (F(V)/F(M)), the amplitude of thermoluminescence (TL) B-band, and the rate of Q(A) reoxidation were observed within 12 h of growth in nitrogen-limited conditions. The decline in F(V)/F(M) paralleled changes in the TL B-band amplitude, indicative of losses in PSII activities and formation of non functional PSII centers. These changes were accompanied by a continuous reduction in D1 protein content. In contrast, nitrogen deprivation did not cause any significant reduction in phycoerythrin content. Our results refute phycoerythrin as a nitrogen storage complex in Prochlorococcus. Regulation of phycoerythrin gene expression in Prochlorococcus is different from that in typical phycobilisome-containing cyanobacteria and eukaryotic algae investigated so far. PMID- 11115646 TI - Isolation of lumenal proteins from spinach thylakoid membranes by triton X-114 phase partitioning. AB - The proteins present in the thylakoid lumen of higher plant chloroplasts have not been rigorously examined. In this communication we present a simple and rapid procedure for the isolation of the soluble proteins and extrinsic membrane proteins present in the thylakoid lumen from spinach. Our procedure involves extensive washing of the thylakoid membranes followed by Triton X-114 phase partitioning. When analyzed by one-dimensional polyacrylamide gel electrophoresis (PAGE), we obtain results which are very similar to those obtained by Kieselbach et al. using more classical methods [T. Kieselbach, A. Hagman, B. Andersson, W.P. Schroder, J. Biol. Chem. 273 (1998) 6710-6716]. About 25 major proteins are observed upon Coomassie blue staining. Upon two-dimensional isoelectric focusing sodium dodecyl sulfate-PAGE and either Coomassie blue or silver staining, however, numerous other protein components are resolved. Our findings indicate that the total number of proteins (soluble and extrinsic membrane) present in the lumen may exceed 150. PMID- 11115647 TI - Polycations induce the release of soluble intermembrane mitochondrial proteins. AB - The release of proapoptotic proteins from the intermembrane space of mitochondria is an early critical step in many pathways to apoptosis. Induction of the mitochondrial permeability transition pore (PTP) was suggested to be the mechanism of the release of soluble mitochondrial intermembrane proteins (SIMP) in apoptosis. However, several studies suggested that proapoptotic proteins (e.g. Bax and Bid) can induce the release of SIMP (e.g. cytochrome c (cyt c) and adenylate kinase 2 (AK2)) in vivo and in vitro independent of PTP. We have found that a number of structurally diverse polycations, such as aliphatic polyamines (e.g. spermine and to a lesser extent spermidine), aminoglycosides (e.g. streptomycin, gentamicin and neomycin), and cytotoxic peptides (e.g. melittin), induce the release of SIMP from liver mitochondria, in vitro. All the polycations released AK2 together with cyt c, suggesting that rupture of the outer membrane is a common mechanism of cyt c release by these polycations. Several polycations (e.g. spermine, spermidine and neomycin) induced SIMP release without inducing significant swelling, and this release was not inhibited significantly by the PTP inhibitor cyclosporin. In contrast, under the same conditions, streptomycin and melittin induced swelling and SIMP release that was inhibited strongly by cyclosporin. Gentamicin-induced swelling and release of SIMP were partially inhibited by cyclosporin. The affinity of polyamines to the anionic phospholipids of the mitochondrial membranes (spermine=neomycin>gentamicin>streptomycin=spermidine) correlated roughly with their ability to induce PTP-independent release of SIMP, which suggests that the binding of polycations to the anionic phospholipids of the outer mitochondrial membrane facilitates the rupture of this membrane. However, some polycations facilitated the induction of PTP, possibly by binding to cardiolipin on the inner membrane. This dual mechanism may be relevant to the induction of SIMP release in apoptosis. PMID- 11115648 TI - Electrogenic events associated with electron and proton transfers within the cytochrome b(6)/f complex. AB - The kinetics and amplitude of the membrane potential changes associated with electron and proton transfers within the cytochrome b(6)/f (cyt b/f) complex (phase b) are measured in vivo in Chlamydomonas reinhardtii under anaerobic conditions. Upon saturating flash excitation, fast components in the membrane potential decay superimposed on phase b lead to an underestimation of the amplitude of this phase. In the FUD50 mutant strain, which lacks the ATP synthase, the decay of the membrane potential is slowed down compared to the wild type, and the kinetics and amplitude of phase b may be accurately determined. This amplitude corresponds to the transfer of at least 1.5 charges across the membrane per positive charge transferred to photosystem I, whatever the flash energy. This value largely exceeds that predicted by a Q-cycle process. Similar conclusions are reached using the wild type strain in the presence of 9 microM dicyclohexylcarbodiimide, which specifically inhibits the ATP synthase. It is concluded that a proton pumping process is operating in parallel with the Q cycle, with a yield of approximately 0.5 proton pumped by cyt b/f complex turnover, irrespective of the flash energy. PMID- 11115649 TI - Purification of ferredoxins and their reaction with purified reaction center complex from the green sulfur bacterium Chlorobium tepidum. AB - Four ferredoxin (Fd) fractions, namely, FdA-D were purified from the green sulfur bacterium Chlorobium tepidum. Their absorption spectra are typical of 2[4Fe-4S] cluster type Fds with peaks at about 385 and 280 nm and a shoulder at about 305 nm. The A(385)/A(280) ratios of the purified Fds were 0.76-0.80. Analysis of the N-terminal amino acid sequences of these Fds (15-25 residues) revealed that those of FdA and FdB completely agree with those deduced from the genes, fdx3 and fdx2, respectively, found in this bacterium (Chung and Bryant, personal communication). The N-terminal amino acid sequences of FdC and FdD (15 residues) were identical, and agree with that deduced from the gene fdx1 (Chung and Bryant, personal communication). The A(385) values of these Fds were unchanged when they were stored for a month at -80 degrees C under aerobic conditions and decreased by 10 15% when they were stored for 6 days at 4 degrees C under aerobic conditions, indicating that they are not extremely unstable. In the presence of Fd-NADP(+) reductase from spinach, and a purified reaction center (RC) preparation from C. tepidum composed of five kinds of polypeptides, these Fds supported the photoreduction of NADP(+) at room temperature with the following K(m) and V(max) (in micromol NADP(+) micromol BChl a(-1) h(-1)): FdA, 2.0 microm and 258; FdB, 0.49 microM and 304; FdC, 1.13 microM and 226; FdD, 0.5 microM and 242; spinach Fd, 0.54 microM and 183. The V(max) value of FdB was more than twice that previously reported for purified RC preparations from green sulfur bacteria. PMID- 11115650 TI - Characterization of the stromal protease(s) degrading the cross-linked products of the D1 protein generated by photoinhibition of photosystem II. AB - When photosystem (PS) II-enriched membranes are exposed to strong light, cross linking of the intrinsic D1 protein with the surrounding polypeptides and degradation of the D1 protein take place. The cross-linking of the D1 protein with the alpha-subunit of cytochrome b(559) is suggested to be an early event of photoinduced damage to the D1 protein (Barbato et al., FEBS Lett. 309 (1992) 165 169). The relationship between the cross-linking and the degradation of the D1 protein, however, is not yet clear. In the present study, we show that the addition of stromal extract from chloroplasts degrades the 41 kDa cross-linked product of D1/cytochrome b(559) alpha-subunit and enhances the degradation of the D1 protein. Incubation of the preilluminated PS II-enriched membranes with the stromal extract at 25 degrees C causes the degradation of the cross-linked product by more than 70%. The activity of the stromal extract showed a pH optimum at 8.0, and was enhanced by the addition of ATP or GTP. Consistent with the nucleotide effect, this stromal activity was eliminated by the preincubation of the stromal extract with apyrase, which hydrolyzes nucleotides. Also, the stromal activity was nearly fully inhibited by a serine-type protease inhibitor, 3,4 dichloroisocoumarin, which suggests participation of a serine-type protease(s). PMID- 11115651 TI - Left posterior BA37 is involved in object recognition: a TMS study. AB - Functional imaging studies have proposed a role for left BA37 in phonological retrieval, semantic processing, face processing and object recognition. The present study targeted the posterior aspect of BA37 to see whether a deficit, specific to one of the above types of processing could be induced. Four conditions were investigated: word and nonword reading, colour naming and picture naming. Repetitive transcranial magnetic stimulation (rTMS) was delivered over posterior BA37 of the left and right hemispheres (lBA37 and rBA37, respectively) and over the vertex. Subjects were significantly slower to name pictures when TMS was given over lBA37 compared to vertex or rBA37. rTMS over lBA37 had no significant effect on word reading, nonword reading or colour naming. The picture naming deficit is suggested to result from a disruption to object recognition processes. This study corroborates the finding from a recent imaging study, that the most posterior part of left hemispheric BA37 has a necessary role in object recognition. PMID- 11115652 TI - Automatic activation of positive but not negative attitudes after traumatic brain injury. AB - This study investigated social judgment problems of an individual (AM) with bilateral frontal and temporal lobe damage including damage to the amygdala. We hypothesized that AM could automatically process positive, but not negative evaluative information and could process both types of evaluative information using controlled processing. In Phase 1 of Experiment 1 AM and controls were shown a series of words one at a time and were required to make good/bad judgments as quickly as possible. Results showed that AM was more likely than controls to rate words as good, and was significantly slower to make good/bad judgments of negatively, but not positively, evaluated words. In Phase 2 AM was shown a prime (positive or negative) then target (positive or negative) and instructed to evaluate whether the target word was good or bad. Results showed that AM responded more quickly when prime and target were both positive, but not when prime and target were both negative, whereas controls showed both types of priming. Experiment 2 determined whether AM's impaired processing of negative evaluative information could be abolished under controlled processing. AM was explicitly instructed to generate positive and negative connotations of a series of single words and given essentially unlimited time. Under these conditions, AM and controls did not differ significantly in their ability to generate positive versus negative connotations of words. In Experiment 3 AM and controls both showed normal semantic priming effects. The results are interpreted within the component process model of memory. PMID- 11115653 TI - Gender differences in a dichotic listening and movement task: lateralization or strategy? AB - Although the dichotic listening procedure has been used as a non-invasive neuropsychological technique for assessing laterality of speech perception, it has tended to underestimate the proportion of the right-handed population that is left-hemisphere lateralized for speech perception [Segalowitz SJ, Bryden MP. Individual differences in hemispheric representation of language. In: Segalowitz SJ (Ed.), Language Functions and Brain Organization. Toronto: Academic Press, 1983, pp. 341-72]. These underestimations may be due to traditional dichotic procedures being susceptible to attentional biases, order of report effects, and/or memory effects that obscure functional differences between the cerebral hemispheres. In the present study, we used an adaptation of the dichotic listening procedure that was designed to be less sensitive to these confounding effects. Participants were required to move as quickly as possible to one of the two color-coded targets following verbal cues presented via headphones. Conditions of cue-word presentation were monaural, (e.g. 'blue' in one ear and a blank track in the other), dichotic-same (e.g. 'blue' in both ears), and dichotic different (e.g. 'green' in one ear and 'blue' in the other). Ninety-three percent (26 of 28) of the participants demonstrated a right ear advantage (REA) for correct responses. There was also a REA for reaction time, movement time, and the total response time. The pattern of reaction time and movement errors, however, suggest that gender differences found utilizing this dichotic procedure may be due to differences in strategic approach to the task rather than to differences in cerebral laterality. Overall, results suggest that this new adaptation of the dichotic listening procedure is very sensitive to lateralization for speech perception. PMID- 11115654 TI - Within grasp but out of reach: evidence for a double dissociation between imagined hand and arm movements in the left cerebral hemisphere. AB - What roles are played by the cerebral hemispheres in planning object-oriented reaching and grasping movements? In an attempt to address this question, we compared the abilities of the left and right hemispheres of commissurotomy patient J.W. to imagine hand manipulation (i.e., grasp) or arm transportation (i.e., reach) movements. A graphically rendered manipulandum (dowel) was briefly presented to the left (LVF) or right (RVF) visual fields in a variety of different orientations. In the grasp selection task (experiment 1), J.W. was required to determine which side of a dowel his thumb would be on if he were to engage the stimulus in a power grip using either his dominant (right) or non dominant hand. In the reach selection task (experiment 3), J.W. judged which end his elbow would be on if he treated the dowel as an armrest for his dominant or non-dominant forearm. No actual movements were allowed in either task. Movements selected in the imagery tasks were compared with those chosen during actual motor control under comparable circumstances. These comparisons revealed a left hemisphere advantage for representing grasping movements involving the right hand, and reaching movements involving the left arm. The right hemisphere, by contrast, displayed moderate accuracy when representing grasping movements with the left hand, but appeared incapable of imagining reaching movements with either arm. The double dissociation between imagery for hand and arm movements in the left cerebral hemispere is consistent with the hypothesis that grasping and reaching components of prehension involve dissociable planning mechanisms. PMID- 11115655 TI - Dense amnesia in the monkey after transection of fornix, amygdala and anterior temporal stem. AB - The traditional explanation of dense amnesia after medial temporal lesions is that the amnesia is caused by damage to the hippocampus and related structures. An alternative view is that dense amnesia after medial temporal lesions is caused by the interruption of afferents to the temporal cortex from the basal forebrain. These afferents travel to the temporal cortex through three pathways, namely the anterior temporal stem, the amygdala and the fornix-fimbria, and all these three pathways are damaged in dense medial temporal amnesia. In four experiments using different memory tasks, we tested the effects on memory of sectioning some or all of these three pathways in macaque monkeys. In a test of scene-specific memory for objects, which is analogous in some ways to human episodic memory, section of fornix alone, or section of amygdala and anterior temporal stem sparing the fornix, each produced a significant but mild impairment. When fornix section was added to the section of anterior temporal stem and amygdala in this task, however, a very severe impairment resulted. In an object recognition memory task (delayed matching-to-sample) a severe impairment was seen after section of anterior temporal stem and amygdala alone, with or without the addition of fornix section; this impairment was significantly more severe than that which was seen in the same task after amygdalectomy leaving the temporal stem intact, with or without fornix section. Animals with combined section of anterior temporal stem, amygdala and fornix were also impaired in object-reward association learning. However, the retention of pre-operatively acquired object-reward associations was at a high level. These results show that the pattern of impairments after section of anterior temporal stem, amygdala and fornix in the monkey, leaving hippocampus intact, resembles human dense amnesia and is different from the effects of hippocampal lesions in the monkey. PMID- 11115656 TI - Crossed unilateral lesions of the medial forebrain bundle and either inferior temporal or frontal cortex impair object-reward association learning in Rhesus monkeys. AB - In an accompanying paper we showed that combined transection of the fornix, amygdala and temporal stem in monkeys produced dense amnesia, including an impairment in visual object-reward association learning. We proposed that this combined surgical section had its effect by isolating temporal cortex from the ascending projections of the basal forebrain and midbrain structures. To test this hypothesis, in the present experiment we disconnected the inferior temporal cortex from these basal forebrain and midbrain structures, while sparing cortical white matter, by crossed unilateral lesions of the medial forebrain bundle in one hemisphere and inferior temporal cortex in the opposite hemisphere. The aim of the medial forebrain bundle lesion was to section axons of cells, both those that project to the cortex via the medial forebrain bundle, and those which control the activity of these same structures. A single unilateral lesion alone had no effect on the ability to learn and remember visual object-reward associations, but the crossed unilateral lesions produced an impairment in this task which was equal in severity to the impairment seen earlier after bilateral section of the fornix, amygdala and temporal stem. The impairment was not an effect of interrupting fibres to the cortex from the ventromedial hypothalamus, or of unilateral sensory neglect. This supports the hypothesis that these midbrain and basal forebrain afferents to the inferior temporal cortex are important for new visual learning. Furthermore, an impairment of equal severity was demonstrated in a separate group of animals that received crossed unilateral lesions of the medial forebrain bundle in one hemisphere and of the frontal cortex in the opposite hemisphere. We propose that the frontal cortex acts to modulate basal forebrain activity which in turn reinforces object representations in the inferior temporal cortex during learning. PMID- 11115657 TI - Acquired mind-blindness following frontal lobe surgery? A single case study of impaired 'theory of mind' in a patient treated with stereotactic anterior capsulotomy. AB - Social insight, specifically the ability to represent thoughts and feelings ('theory of mind'), may have a circumscribed and dedicated neurological substrate. Evidence of deficits in 'theory of mind' following acquired lesions would support this idea. Previous studies of lesions resulting from stroke or head injury have been hampered by lack of detailed lesion information and pre lesion documentation. We report the case of a 76-year-old man who, following a standard surgical procedure to treat bipolar affective disorder, showed evidence of impaired 'theory of mind'. This case, which is the first of its type, may contribute to the search for the brain basis of social insight. PMID- 11115658 TI - Differences in brain potentials to open and closed class words: class and frequency effects. AB - Closed class (determiners, pronouns, conjunctions, prepositions etc. ) and open class (nouns, verbs, adjectives, adverbs) words have different linguistic functions and have been proposed to be processed by different neural systems. Here, event-related potentials (ERPs) were recorded in young German-speaking subjects while they read closed class and open class words flashed upon a video screen. In the first experiment closed class words were sorted into four different frequency categories and open class words into three categories. The words were presented in a list with the subjects' task to detect occasional non words. A centroparietal negativity (N400) with a peak latency of about 400 ms varied in amplitude as a function of frequency in both classes. The N400 in closed class items, however, was considerably smaller than that in open class words of similar frequency. A left anterior negativity (N280/LPN) showed some degree of frequency-sensitivity regardless of word class. Only for the very high frequency closed class words a frontal negativity with an onset of about 400 ms was obtained (N400-700). This N400-700 effect was replicated in the second study, in which medium frequency closed and open class words and very high frequency closed class words were presented at the fifth position of simple German sentences. It is suggested that neither N400 nor the left anterior negativity (N280/LPN) distinguish qualitatively between the two word classes and thus claims about different brain systems involved in the processing of open and closed class words are not substantiated electrophysiologically. The N400-700 effect is possibly related to specific grammatical functions of some closed class items, such as determiners. PMID- 11115672 TI - Cell differentiation, synaptogenesis, and influence of the retinal pigment epithelium in a rat neonatal organotypic retina culture. AB - This study was focused on the analysis of cell type differentiation and synaptogenesis as well as outer segment formation in an organotypic culture of the neonatal rat retina during a 6-14 day period of in vitro development. Moreover, the effects of the retinal pigment epithelium (RPE) on these processes were investigated. The in vitro development resulted in a retinal architecture and lamination comparable to that of in vivo retinas. The RPE influences the proper alignment of photoreceptors as well as the formation of the outer limiting membrane (OLM), but not processes of cell differentiation, synaptogenesis and inner retinal lamination. PMID- 11115673 TI - Daily rhythm of vigilance assessed by temporal resolution of the visual system. AB - To assess the daily distribution of temporal resolution in visual detection, binocular double-pulse resolution (DPR) was measured over a 24 h period in six healthy subjects. DPR showed a significant daily variation with an amplitude for the foveal stimulus of up to 60%. Like in other vigilance-dependent daily rhythms, optimal performance occurred around midday. The DPR measurements described here are an excellent method for assessment of vigilance and mental alertness (e.g. in pharmacological studies). They show strong time-of-day differences, are highly reliable across successive measurements, and can be fully automated. PMID- 11115674 TI - Illusory contour strength does not depend on the dynamics or relative phase of the inducers. AB - We use a new objective measure of illusory contour strength, threshold reduction for aspect ratio discrimination, to examine the effect of dynamics and relative phase on the Kanizsa illusion. We found no dependence of illusory contour strength on the relative phase of flickering inducers (in phase, antiphase, or in quadrature phase) either for the standard Kanizsa square, or for modifications that facilitated or interfered with amodal completion. Comparison with a vernier acuity task indicates that the distance between the inducers, rather than the nature of the task, accounts for the insensitivity to relative phase. PMID- 11115675 TI - Ambiguity in the perception of moving stimuli is resolved in favour of the cardinal axes. AB - The aim of this study was to determine whether there is a link between the statistical properties of natural scenes and our perception of moving surfaces. Accordingly, we devised an ambiguous moving stimulus that could be perceived as moving in one of three directions of motion. The stimulus was a circular patch containing three square-wave drifting gratings. One grating was always either horizontal or vertical; the other two had component directions of drift at 120 degrees to the first (and to each other), producing four possible stimulus geometries. These were presented in a pseudorandom sequence. In brief presentations, subjects always perceived two of the gratings to cohere and move as a pattern in one direction, and the third grating to move independently in the opposite direction (its component direction). Although there were three equally plausible axes (one cardinal and two oblique) along which the coherent and independent motions could occur, subjects routinely saw motion along one of the cardinal axes. Thus, the visual system preferentially combines the two oblique gratings to form a pattern that drifts in the opposite direction to the cardinal grating. It was only when the contrast of one of the oblique gratings was changed that an oblique axis of motion was perceived. This perceptual anisotropy can be related to naturally occurring bias in the visual environment, notably the predominance of horizontal and vertical contours in our visual world. PMID- 11115676 TI - No local cancellation between directionally opposed first-order and second-order motion signals. AB - Despite strong converging evidence that there are separate mechanisms for the processing of first-order and second-order motion, the issue remains controversial. Qian, Andersen and Adelson (J. Neurosci., 14 (1994), 7357-7366) have shown that first-order motion signals cancel if locally balanced. Here we show that this is also the case for second-order motion signals, but not for a mixture of first-order and second-order motion even when the visibility of the two types of stimulus is equated. Our motion sequence consisted of a dynamic binary noise carrier divided into horizontal strips of equal height, each of which was spatially modulated in either contrast or luminance by a 1.0 c/deg sinusoid. The modulation moved leftward or rightward (3.75 Hz) in alternate strips. The single-interval task was to identify the direction of motion of the central strip. Three conditions were tested: all second-order strips, all first order strips, and spatially alternated first-order and second-order strips. In the first condition, a threshold strip height for the second-order strips was obtained at a contrast modulation depth of 100%. In the second condition, this height was used for the first-order strips, and a threshold was obtained in terms of luminance contrast. These two previously-obtained threshold values were used to equate visibility of the first-order and second-order components in the third condition. Direction identification, instead of being at threshold, was near perfect for all observers. We argue that the first two conditions demonstrate local cancellation of motion signals, whereas in the third condition this does not occur. We attribute this non-cancellation to separate processing of first order and second-order motion inputs. PMID- 11115677 TI - Depth cue integration: stereopsis and image blur. AB - Depth-of-focus limitations introduce spatial blur in images of three-dimensional scenes. It is not clear how the visual system combines depth information derived from image blur with information from other depth cues. Stereoscopic disparity is the pre-eminent depth cue, so experiments were conducted to investigate interactions between image blur and stereoscopic disparity. Observers viewed two random dot stereograms (RDSs) in a 2AFC task, and were required to identify the RDS depicting the greatest depth. In control observations, all dots in both RDSs were sharply defined. In experimental observations, one RDS contained only sharply defined dots, but the other contained differential spatial blur to introduce an additional depth cue. Results showed that the addition of differential blur made only a marginal difference to apparent depth separation, and only when the blur difference was consistent with the sign of disparity. Cue combination between blur and disparity cues is thus weighted very heavily in favour of the latter. It is shown that blur and disparity cues co-vary according to geometric optics. Since the two cues are effective over different distances, the visual system is not normally called upon to integrate them, and is most likely to make use of blur cues over distances beyond the range of disparity mechanisms. PMID- 11115678 TI - Perceived speed of motion in depth is reduced in the periphery. AB - The perceived speed of motion in depth (MID) for a monocularly visible target was measured in central and peripheral vision using a 2AFC speed discrimination task. Only binocular cues to MID were available: changing disparity and interocular velocity difference (IOVD). Perceived speed for monocular lateral motion and perceived depth for static disparity were also assessed, again in both central and peripheral vision. The purpose of the experiment was to assess the relative contributions of changing disparity and IOVD cues to the perceived speed of stereomotion. Although peripheral stimuli appeared to lie at approximately the same depth as their central counterparts, their apparent speed was reduced. Monocular/lateral and binocular/MID speeds were reduced to a similar extent. It seems that reduced apparent monocular speed leads to reduced perceived MID speed, despite the fact that the disparity system appears to be unaffected. These results suggest that the IOVD cue makes a significant contribution to MID speed perception. PMID- 11115679 TI - Central performance drop in texture segmentation: the role of spatial and temporal factors. AB - Previous studies reported that performance in texture segmentation was lower near the fovea than in the periphery. However, the exact cause of this phenomenon had been unknown. Experiment 1 replicated the central performance drop (CPD). Experiments 2 and 3 demonstrated that the previously reported CPD was due to a temporal factor, i.e. slower neural processing in central vision, rather than a spatial factor. But Experiments 4 and 5 showed that certain textures can lead to a purely spatial form of CPD due to inhibition and/or interference from high spatial frequency mechanisms in central vision. This study showed that, depending on textures, CPD can arise from either temporal or spatial causes. PMID- 11115680 TI - The planning of refixation saccades in reading. AB - The planning of the refixation saccade, i.e. the second saccade on 9- and 11 letter-strings, was assessed in two reading experiments that examined the influence of a length change at different times during the first fixation on a letter string. The results showed that the saccadic system was able to modify the first motor program if the new length information was available 150-190 ms before the execution of the refixation saccade. Moreover, the amplitude of the refixation saccade was found to be planned as a constant movement relative to the length of the item, regardless of the position of the initial fixation on the item. Finally, the refixation saccade seems to be preprogrammed before the primary saccade, depending on the length integrated at that time. Overall, these results suggest that the refixation saccade is programmed on the basis of the intrinsic properties of the item, such as its length. PMID- 11115681 TI - Processing spatial information in the sensorimotor branch of the visual system. AB - We distinguish two representations of visual space: a cognitive representation drives perception, and a sensorimotor representation controls visually guided behavior. Spatial values in the two representations are separated with the Roelofs effect: a target within an off-center frame appears biased in a location opposite the direction of the frame. The effect appears for a verbal measure (cognitive) but not for a jab at the target (sensorimotor). A 2-s response delay induces a Roelofs effect in the motor measure, showing the limit of motor memory. Motor error is not correlated with reaction time. Subjects could strike one of two identical targets, a process involving choice, without intrusion of a Roelofs effect, showing that the sensorimotor system can use its own coordinates even when a cognitive choice initiates the motor processing. PMID- 11115682 TI - Airway foreign bodies (FB): a 10-year review. AB - A retrospective chart review of children who had airway foreign body removed via direct laryngoscopy and bronchoscopy (DLB) from 1987-1997 was conducted in Children's Hospital, Boston. Patient characteristics noted included age, sex, and clinical presentation. Pre-operative radiographic findings, reason for delay in evaluation, DLB findings, length of procedure, reason for repeat DLB, and types of foreign body etc. were recorded. Serious complications from aspirated foreign bodies such as severe airway obstruction and death tend to occur in infants and younger children because of their small airway size. A history compatible with foreign body aspiration dictates diagnostic endoscopy with or without radiologic confirmation. Chest and airway radiographs supplemented by fluoroscopy can increase the ratio of correct and early diagnosis. Fluoroscopy should be universally accepted as an initial diagnostic technique in airway foreign body evaluation. Fluoroscopy is not a worthwhile investigation if a preceeding chest radiograph suggests the presence of a foreign body. Long-standing airway foreign bodies are associated with considerable morbidity, and early diagnosis remains the key to successful and uncomplicated management of foreign body aspiration. Education aimed at increasing diagnostic acumen of the physicians and heightening of public awareness are the most important steps needed to reduce the morbidity and mortality. Parents should be instructed to abstain from feeding nuts and seeds to young children and to keep small, potentially ingestible objects out of their reach. PMID- 11115683 TI - Factors contributing to limited open-set speech perception in children who use a cochlear implant. AB - Cochlear implants have enabled many children with severe to profound sensorineural hearing loss to develop speech perception skills. However, some children experience few gains while others develop high levels of speech perception. We identified potential factors contributing to poor performance with an implant by studying implanted children who do not develop functional speech perception. Five children were identified as developing no open-set word recognition skills after at least 2 years of implant use. This study group was compared to a randomly selected control group (n=10) and an age-matched control group (n=5). Pre-implant factors were examined using a Graded Profile Analysis and post-implant factors were assessed in a retrospective chart review. A greater number of pre-implant concerns were raised in the study group than in randomized controls (P<0.01). Chronological age and duration of deafness were pre-implant concerns in all study group subjects. A greater number of post-implant concerns were found in the study group than in randomly selected controls (P<0.005). We conclude that while appropriate selection of candidates for cochlear implantation is important in predicting speech perception outcomes, post-implant follow-up is also essential and must include regular monitoring of equipment, monitoring of stimulation levels with use of objective measures of stimulation levels if necessary, and consistent habilitation. PMID- 11115684 TI - German registry for hearing loss in children: results after 4 years. AB - Since April 1996 the German Registry for Hearing Loss in Children has been collecting and recording various data on permanent hearing loss in children. Up to now, the data of 3882 children have been gathered nation-wide. A particularly remarkable aspect of data so far collected is the alarmingly high age at the time of diagnosis (mild hearing loss is on average only diagnosed at 6; 2 years, moderate loss at 4; 4 years, severe at 2; 5 years and profound with 1; 9 years). The German Registry for Hearing Loss in Children provides the first reliable source of epidemiological data on permanent hearing loss in children in Germany. A constantly growing number of currently 112 co-operating partners takes part in the data acquisition. The periodical reports of results to the co-operating partners and the first publications have heightened the awareness of the importance of early diagnosis and therapy. A minimum standard for patient history was established by the primary examination report. Thus, an important contribution has been made to the prevention of avoidable secondary damage. PMID- 11115685 TI - Prevention of acute mastoiditis: fact or fiction? AB - Acute mastoiditis is the most common complication of acute otitis media (AOM). In recent years routine antibiotic treatment for acute middle ear infections was questioned and even abandoned in some countries. The goal of our study was to investigate the influence of antibiotic treatment on the occurrence and clinical outcome of acute mastoiditis and to analyze the bacteriological findings. A retrospective case record study of 48 patients with 50 episodes of acute mastoiditis hospitalized at our tertiary-care center between 1992 and 1999 was performed. Twenty-three patients (48%) received antibiotic treatment before admission whereas 25 (52%) did not. The group of patients without antibiotic pretreatment were younger (mean, 6 years) than patients with antibiotics (mean, 18 years) and their referral was delayed. The most common isolated single pathogen was Streptococcus pneumoniae. All pneumococci were sensitive to penicillin. Acute mastoiditis may be the first clinical sign of a middle ear infection, especially in very young children. Adequate antibiotic pretreatment cannot invariably prevent the development of acute mastoiditis even in the absence of penicillin resistant pathogens. PMID- 11115686 TI - One-stage revision surgery for pediatric cholesteatoma: long-term results and comparison with primary surgery. AB - OBJECTIVE: Few studies report on revision surgery for pediatric cholesteatoma, even if most studies of primary surgery show high recurrence rates. We present independently evaluated long-term results of revision surgery and compare the results with those of primary surgery. METHODS: The series consisted of 42 consecutive pediatric (age <16 years) cholesteatoma revision operations in the Helsinki University ENT Department. The primary and revision surgery was non staged, all mastoids were obliterated and the bony ear canals were reconstructed. The preoperative, surgical and annual control data were recorded in a database. The last control was independently performed (J.S.) with an average follow-up of 4.3 years and 87% attendance. RESULTS: The recurrence rate for revision operations was 38%. A retraction process developed in 38% of the ears and 67% of these retractions turned into active cholesteatomas. There was a non-significant difference for these figures as compared with primary surgery. Postoperative discharge and poor middle ear ventilation were associated with recurrence. The following factors showed significant differences between primary and revision surgery: increased number of bare facial nerve in the revision group, lower recurrence rate for experienced surgeons (as for both groups together) and improved hearing results for ears without recurrence in the revision group. CONCLUSIONS: Recurrent disease after revision surgery appears either in the attic or in the mesotympanum, or develops from a retraction pocket in a similar manner as was seen in patients receiving primary surgery. Recurrence is associated with discharging and poorly ventilated ears. Pediatric cholesteatoma surgery should be done or be supervised by experienced surgeons. The present operation methods result in many recurrences after primary and revision surgery. New surgical methods are needed to create additional aeration pathways to the epitympanum in order to improve middle ear aeration and to prevent some of the retractions. PMID- 11115687 TI - Glial lesion of the infratemporal fossa presenting as a soft tissue middle ear mass - rudimentary encephalocele or neural crest remnant? AB - We report about ectopic glial tissue of the skull base and the parapharyngeal space presenting as a soft tissue mass in the middle ear. An 11-year-old boy presented with bilateral conductive hearing loss since early childhood. The history included previously removed lesions consistent with ectopic neuroglial tissue of the tongue and the parapharyngeal space soon after birth, as well as surgery for cleft palate. High resolution computed tomography of the petrous bone and magnetic resonance imaging were useful in identifying the skull base defect and in characterizing the lesion's relation to the brain. There was no clinical, radiological or surgical evidence of any associated dural defect. The lesion was removed via a modified infratemporal approach. Histology revealed neuroglial tissue with calcifications without any signs of mesodermal or entodermal origin. On the basis of this case the pathogenesis and diagnosis of ectopic brain tissue and its relation to the more commonly encountered meningoencephalic herniations are reviewed. Furthermore therapeutical implications are discussed. PMID- 11115688 TI - Long term persistence of anti-HBs protective levels in young patients with type 1 diabetes after recombinant hepatitis B vaccine. AB - The aim of the present study was to evaluate the persistence of anti-hepatitis B protective levels in young patients with type 1 diabetes, successfully immunised with a recombinant hepatitis B vaccine. We re-evaluated, after a 4 year follow up, 54 patients and 70 age and sex-matched healthy subjects. Protective antibodies levels were found in 50/54 (92%) patients and in 67/70 (96%) controls. Moreover, anti-HBs levels were similar in diabetic patients and controls (means of log-titre and (sd); 1.95 (0.88) and 2.18 (0.64) patients and controls, respectively; P=0.11). No cases of clinical hepatitis were reported and all patients and controls remained HBc negative. These data demonstrate the persistence of anti-HBs levels in children, adolescents and young patients with type 1 diabetes after recombinant hepatitis B vaccine showing evidence of longterm immunogenity and protective effect. PMID- 11115689 TI - Antigenicity and immunogenicity of the HIV-1 gp41 epitope ELDKWA inserted into permissive sites of the MalE protein. AB - The highly conserved amino acid sequence ELDKWA of HIV-1 gp41 has been inserted into Escherichia coli MalE protein which had been shown to be an adequate carrier to present foreign epitopes to the immune system. We first investigated whether eight different permissive sites of MalE are able to tolerate an insertion of 7 50 residues encoding this epitope. Secondly, antigenicity of the epitope inserted in MalE protein was estimated from monoclonal antibody 2F5 binding analysis using the BIAcore(R) technology and its immunogenicity in mice was measured as the ability of hybrid proteins to elicit antibodies against a synthetic peptide containing this epitope. This study revealed a good correlation between the antigenicity of the inserted epitope and its immunogenicity. Increasing the length of the inserted epitope, as well as inserting multicopies of this epitope increased both its antigenicity and immunogenicity. However, none of the MalE hybrid proteins tested induced anti-HIV-1 neutralizing antibodies. This study strongly suggests that the capacity of the 2F5 epitope to induce neutralizing antibodies depends on the molecular context in which it is presented. PMID- 11115690 TI - Comparison of in vitro and in vivo methods to study stability of PLGA microencapsulated tetanus toxoid vaccines. AB - The purpose of this study was to investigate the utility of various in vitro and in vivo methods to assess the stability of experimental vaccines containing tetanus toxoid (TT) within PLGA microspheres. In vitro, the breakdown of the encapsulating polymers into their acid components led to changes in the structure of TT, as determined by the physico-chemical methods, rendering it undetectable by capture ELISA and altering its structural integrity. The changes in TT were directly related to increasing acidity of the vaccine supernate. Purified toxoid (not encapsulated) exposed to low pH (2.5) underwent similar changes but re neutralisation of buffer containing free toxoid, even after one week at pH 2.5 led to some re-folding of protein as determined by fluorescence spectroscopy and gel filtration chromatography. The microencapsulated vaccines were still able to generate an antibody response in mice even after prolonged pre-incubation at 37 degrees C and the apparent absence of detectable toxoid in the vaccine supernate. Electron microscopy demonstrated differences in the amount of degradation between different formulations of microspheres. Vaccines that had retained their spherical morphology after incubation in vitro for up to 28 days were able to induce protective antibodies response equal to that of freshly prepared vaccines, which indicates that the toxoid within intact microspheres remained immunogenic. Immunochemical and physico-chemical detection methods, performed on antigen released from PLGA vaccines in vitro, are valuable in providing information on product characteristics but may not be able to predict effectiveness and should be used with in vivo methods to evaluate the stability of such formulations. PMID- 11115691 TI - Microencapsulated enterotoxigenic Escherichia coli and detached fimbriae for peroral vaccination of pigs. AB - The feasibility of peroral immunisation with microencapsulated Escherichia coli and detached fimbriae to prevent enterotoxigenic E. coli infections in pigs was examined. For this E. coli and fimbriae were microencapsulated into poly(lactide co-glycolide) microspheres by spray-drying. Various microsphere formulations designed to deliver priming and booster doses were fed to new-born and weaned pigs. The pigs were challenged 19 days after the booster dose by peroral administration of an infective dose of the homologous E. coli. Serum IgA antibody titres and excretion of challenge E. coli, as indicators for colonisation, were determined. The data showed that no significant serum antibodies were induced, and E. coli colonisation was not reduced by the peroral administration of the various antigen-loaded microspheres. These results are in contradiction to some of the previously published experiments typically in rats or rabbits, where model antigens or unpractical immunisation procedures have frequently been used. PMID- 11115692 TI - Assessment of the stability and immunogenicity of meningococcal oligosaccharide C CRM197 conjugate vaccines. AB - In this stability study, meningococcal C-CRM(197) conjugate vaccines from two different manufacturers that differ in oligosaccharide chain length, number of conjugation sites, conjugation chemistry, manufacturing process and formulation were used. Both the bulk concentrated and final fill preparations were incubated at -20, 4, 23, 37 or 55 degrees C for 5 weeks or subjected to ten cycles of freeze-thawing. The structural stability, hydrodynamic size and integrity of the treated vaccines were monitored by size exclusion chromatography (FPLC-SEC), high performance anion exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD) and fluorescence spectroscopy techniques. The data showed that the structural stability of the oligosaccharide chains and of the protein carrier varied between the two conjugates. The experimental immunogenicity was not severely affected by repeated freeze-thawing, incubation at -20 or 4 degrees C, but one developed conformational changes in the protein carrier when incubated at 23 degrees C or above, although the integrity of the oligosaccharide structure was maintained. This was not associated with any reduction in primary IgG or IgM antibody responses to meningococcal C polysaccharide. In the other conjugate vaccine, exposure to 55 degrees C resulted in the release of a substantial proportion of free saccharide that was accompanied by significant reduction in both IgG and IgM antibody responses to immunisation in the model system. In conclusion, the two meningococcal C-CRM(197) conjugate vaccines were stable when stored at the recommended temperatures, although their structural stability and subsequent immunogenicity were influenced by their conjugation chemistry and formulation. PMID- 11115693 TI - Effect of different baculovirus inactivation procedures on the integrity and immunogenicity of porcine parvovirus-like particles. AB - We have demonstrated earlier the usefulness of recombinant porcine parvovirus (PPV) virus-like particles (VLPs) as an efficient recombinant vaccine for PPV. Here, we have demonstrated that preparations of PPV VLPs could be contaminated by recombinant baculoviruses. Since these baculoviruses can be a problem for the registration and safety requirements of the recombinant vaccine, we have tested different baculovirus inactivation strategies, studying simultaneously the integrity and immunogenicity of the VLPs. These methods were pasteurization, treatment with detergents and alkylation with binary ethylenimine (BEI). The structural and functional integrity of the PPV VLPs after the inactivation treatments were analyzed by electron microscopy, hemagglutination, double antibody sandwich (DAS)-ELISA and immunogenicity studies. Binary ethylenimine and Triton X-100 inactivated particles maintained all the original structural and antigenic properties. In addition, PPV VLPs were subjected to size-exclusion chromatography to analyze the presence of VP2 monomers or any other contaminant. The resulting highly purified material was used as the standard of reference to quantify PPV VLPs in order to determine the dose of vaccine by DAS-ELISA. After immunization experiments in guinea pigs, the antibody titers obtained with all the inactivation procedures were very similar. Triton X-100 treatment was selected for further testing in animals because of the speed, simplicity and safety of the overall procedure. PMID- 11115694 TI - Potential live vaccines for HIV. AB - Potential live vaccines for HIV were developed using an Lpp-OmpA system to target an HIV antigen, reverse transcriptase, or an immunodominant epitope of this enzyme, to the outer membrane of an attenuated strain of Salmonella SL3261. These live vaccines were administered orally to mice, and fecal IgA and helper T cell responses were measured. Results indicated a fecal IgA response specific to HIV reverse transcriptase, as well as a reverse transcriptase-specific helper T cell response, as measured by proliferation assays. Additionally, tests with the epitope vaccines showed a selective cytotoxic CD8 T cell response. These results suggest that this method of antigen targeting to the outer membrane of attenuated bacterial vectors is very promising not only for HIV vaccine development, but also for antigens from other viral or bacterial pathogens, which could be inserted into the Lpp-OmpA protein construct, to elicit mucosal IgA and T cell responses. PMID- 11115696 TI - Antibody reactions after aerogenous or subcutaneous immunization of pigs with Pasteurella multocida antigens. AB - Depending upon the antigens used and the initial titres, subcutaneous immunization of weaner pigs by means of a temperature-sensitive mutant of live Pasteurella multocida (serovar A), resulted in a significant rise of the level of specific IgG antibody already present in blood serum, but not in lung lavage fluid, and a specific stimulation of lung clearance as compared to non-immunized controls. Aerogenous immunization of a total of 108 animals in 18 experiments did not influence serum antibody titres but produced a significant rise in lung antibodies and P. multocida clearance as compared to an identical number of controls. In all 12 immunization experiments using the live mutant, increased specific IgA antibodies and in seven out of 10 experiments, elevated IgG antibodies were measured. Only aerogenous immunization was found to be capable of reducing the severity of pneumonia induced by intrabronchial infection. PMID- 11115695 TI - Randomized, cross-over, controlled comparison of two inactivated hepatitis A vaccines. AB - The immunogenicity, tolerability and interchangeability of two hepatitis A vaccines, Vaqta (Merck and Co.) and Havrix (SmithKline) were studied in a randomized, crossover, controlled clinical trial. Vaccine was administered to 201 volunteers at 0 and 26 weeks in one of four vaccine regimens: Havrix-Havrix; Havrix-Vaqta; Vaqta-Havrix or Vaqta-Vaqta. Seroconversion rates (>/=10 mIU/ml) for those whose first dose was Vaqta or Havrix, respectively, were: 41/96 (43%) versus 30/95 (32%) (P=0.15) at 2 weeks and 91/98 (93%) versus 84/97 (87%) (P=0.43) at 4 weeks, and 100% at 26 weeks. Geometric mean concentrations (GMC) of total antibody to hepatitis A virus (anti-HAV) for Vaqta and Havrix were 189 and 114 mIU/ml (P=0.011) at 4 weeks and 234 and 136 mIU/ml (P<0.001) at 26 weeks. At 30 weeks, the GMC after two doses of Havrix was 2612 mIU/ml compared with 5497 after two doses of Vaqta (P<0.001). The GMC in the Havrix-Vaqta group was 5672 mIU/ml compared with 3077 mIU/ml in the Vaqta-Havrix group (P<0.001). Less than half of vaccine recipients reported tenderness or pain. In this study, seroconversion rates of the two vaccines were similar. Vaqta produces significantly higher anti-HAV antibody than Havrix. Crossover immunization is well tolerated and results in high antibody concentrations, especially when Vaqta is the booster dose. The significance of higher anti-HAV antibody concentrations in terms of long-term protection is unknown. PMID- 11115697 TI - Mapping of B-cell epitopes on human parvovirus B19 non-structural and structural proteins. AB - In the present study, the immune reactivity to Parvovirus B19 (B19) proteins and variations in antigenic reactivity in different clinical manifestations were investigated. Sera from healthy B19 IgG positive individuals were evaluated for antibody reactivity against linear peptides. Three antigenic regions (amino acid number 191-206, 271-286, 371-386) on the B19 non-structural (NS) protein 1 were identified. The highest seroreactivity against these peptides was found against amino acid number 271-286. Seroreactivity in this group of individuals was also investigated against peptides representing selected neutralising regions of the B19 capsid proteins viral protein (VP) 1/VP2. The antigenic NS1 and VP1/VP2 regions, thus defined, were further mapped by seroreactivity against peptides containing specific deletions. The frequencies of seroreactivity against the NS1 and VP1/VP2 peptides in healthy B19 IgG positive individuals were similar to those in HIV-seropositive and persistently B19 infected patients, except that the latter group showed a lower reactivity to the C-terminal end of VP1/VP2. The identification of antigenic regions and corresponding seroreactivity in asymptomatic and persistently B19-infected patients is important for the understanding of B19-pathogenesis and for the development of B19 vaccine candidates. PMID- 11115698 TI - Induction of antigen-specific CD8+ T cells, T helper cells, and protective levels of antibody in humans by particle-mediated administration of a hepatitis B virus DNA vaccine. AB - A DNA vaccine against the hepatitis B virus (HBV) was evaluated for safety and induction of immune responses in 12 healthy, hepatitis-naive human volunteers using the needle-free PowderJect system to deliver gold particles coated with DNA directly into cells of the skin. Three groups of four volunteers received three administrations of DNA encoding the surface antigen of HBV at one of the three dose levels (1, 2, or 4 microg). The vaccine was safe and well tolerated, causing only transient and mild to moderate responses at the site of administration. HBV specific antibody and both CD4+ and CD8+ T cell responses were measured before and after each immunization. All the volunteers developed protective antibody responses of at least 10 mIU/ml. In volunteers who were positive for the HLA class I A2 allele, the vaccine also induced antigen-specific CD8+ T cells that bound HLA-A2/HBsAg(335-343) tetramers, secreted IFN-gamma, and lysed target cells presenting a hepatitis B surface antigen (HBsAg) CTL epitope. Enumeration of HBsAg-specific T cells producing cytokine indicated preferential induction of a Type 1 T helper cell response. These results provide the first demonstration of a DNA vaccine inducing protective antibody titers and both humoral and cell mediated immune responses in humans. PMID- 11115699 TI - Mycobacterium bovis bacillus calmette-guerin induces protective immunity against infection by Plasmodium yoelii at blood-stage depending on shifting immunity toward Th1 type and inducing protective IgG2a after the parasite infection. AB - Bacillus calmette-guerin (BCG)-vaccination raised dramatically the survival rates of A/J mice from infection by Plasmodium yoelii 17XL at blood-stage. The analysis of the immune response of spleen cells indicated that BCG vaccination biased the immune response toward Th1 type. Neutralization of IFN-gamma and nitric oxide abrogated the protection. The kinetics of Ab production in the course of P. yoelii 17XL infection was monitored. Surprisingly, larger amounts of parasite specific Abs were produced in BCG-vaccinated mice than in the placebo control. The vast majority of the produced IgG against parasites in BCG-vaccinated mice was IgG2a, which was observed hardly in placebo controls. The peak of IgG2a production coincided with the clearance of infection. The naive mice transferred adoptively with IgG2a from self-cured mice survived the lethal challenge from the parasite. These data indicated that BCG vaccination protected A/J mouse from P. yoelii 17XL infection by biasing immunity toward Th1-type after parasite infection and enhancing production of IgG2a, which ultimately played a major role in protection. PMID- 11115700 TI - DNA immunisation against the CFA/I fimbriae of enterotoxigenic Escherichia coli (ETEC). AB - The CFA/I fimbria promotes the attachment of enterotoxigenic Escherichia coli (ETEC) to the surface of human enterocytes. The generation of a protective immune response requires the induction of antibodies able to block the CFA/I-mediated binding of ETEC to receptors located on the small intestine epithelium or on the surface of human red blood cells, in hemagglutination tests. An eukaryotic expression plasmid, pBLCFA, encoding the CFA/I gene under the control of the human cytomegalovirus major immediate-early promoter was constructed as a prototype DNA vaccine against ETEC. pBLCFA-tranfected BHK-21 cells secreted a peptide cross-reacting with a monoclonal antibody raised against CFA/I subunits. BALB/c mice immunized intramuscularly with one or two doses of purified pBLCFA developed CFA/I-specific serum antibodies for at least 52 weeks, composed predominantly of the IgG1 subclass. pBLCFA-induced antibodies bind mainly to epitopes exposed on the surface of intact CFA/I fimbriae and do not react with immune recessive epitopes found in other ETEC fimbra sharing amino acid homologies with CFA/I. Furthermore, pBLCFA-induced antibodies were able to block the adhesive properties of the CFA/I fimbriae, as evaluated by the ability to inhibit the hemagglutination promoted by CFA/I-expressing ETEC cells. These results suggest that secretion of CFA/I encoded by pBLCFA preserves important conformational epitopes required for the generation of protective antibodies against the adhesive properties of the CFA/I fimbriae and open new perspectives for the development of DNA vaccines against enteric bacterial pathogens. PMID- 11115701 TI - Efficacies of BCG and vole bacillus (Mycobacterium microti) vaccines in preventing clinically apparent pulmonary tuberculosis in rabbits: a preliminary report. AB - Tuberculosis (TB) kills more people in the world today than any other infectious disease, and the number of drug-resistant Mycobacterium tuberculosis isolates is increasing. Vaccines, better than most of the currently available strains of bacille Calmette-Guerin (BCG), are urgently needed to control this disease. TB in rabbits resembles human TB more closely than TB in any other common laboratory animal and a most pertinent method of assessing vaccine efficacy is Lurie's tubercle count method in this species. Vaccinated and control rabbits were infected by aerosol with virulent human-type tubercle bacilli (H37Rv). At necropsy 5 weeks thereafter, the grossly visible primary tubercles in the entire lung were counted. A decrease in the number of such tubercles is a quantitative measure of vaccine efficacy: An effective vaccine prevents microscopic tubercles from growing to grossly visible (clinically apparent) size. The Pasteur substrain of BCG and two substrains of Mycobacterium microti (the vole bacillus) reduced the number of visible primary tubercles an average of 75%, whereas three other substrains of BCG and three other substrains of vole bacilli only reduced the number an average of 40%. These initial studies indicate that Lurie's tubercle count method in rabbits is a precise way to choose the best available tuberculosis vaccines. PMID- 11115702 TI - Assessment of DNA vaccine potential for juvenile Japanese flounder Paralichthys olivaceus, through the introduction of reporter genes by particle bombardment and histopathology. AB - Genetic immunisation potential, following DNA bombardment for juvenile Japanese flounder, Paralichthys olivaceus was examined. GFP plasmids bombarded at two pressures, 150 and 300psi were sampled at 1, 7, 14 and 28 days, greater immunofluorescence was observed at the higher bombardment pressure. Histopathology, at 3 h post bombardment showed considerable damage to fish epithelial and dermal tissues when bombarded at pressures greater than 200 psi, with many DNA-coated gold particles present. At 150psi there was little pathology and no DNA-coated particles. Histopathology, up to 28 days again showed little pathology at 150 psi with few DNA-coated particles, whereas at 300 psi there was significant pathology observed with many DNA-coated particles seen in conjunction with the cytoplasm of inflammatory cells. By day 28 epithelial coverage was observed with tissue damage restricted to the dermal layer. Chloramphenicol acetyltransferase (CAT) assay showed long term and stable expression of the CAT protein from day 1 to day 60. The transcription activity of two promoters; pCMV CAT and pSV2-CAT showed greater activity in the former. It was concluded that DNA vaccination potential for juvenile flounder is a viable option. PMID- 11115703 TI - Influence of codon usage on the immunogenicity of a DNA vaccine against tetanus. AB - Two related DNA vaccine vector plasmids, harbouring either wild-type (pcDNA3/ntetC) or synthetic codon optimised (pcDNA3/stetC) DNA encoding fragment C (TetC) of tetanus toxin were constructed. COS-7 cells transformed with pcDNA3/stetC reproducibly expressed higher levels of TetC than similar cells transformed with pcDNA3/ntetC. BALB/c mice immunised intramuscularly with pcDNA3/stetC produced significantly higher levels of anti-TetC antibodies in their serum in the weeks following vaccination compared to mice immunised with pcDNA3/ntetC, even when differences in the CpG content between the two sequences were controlled for using non-expressing DNA. PMID- 11115704 TI - Differences in epitope recognition, isotype and titer of antisera to Plasmodium falciparum merozoite surface protein 4 raised by different modes of DNA or protein immunization. AB - Plasmodium falciparum merozoite surface protein 4 (MSP4) is being developed as a component of a subunit vaccine against asexual stages of malaria. Three DNA constructs were produced that induced expression of MSP4 either in the cytoplasm of transfected cells or secreted from cells under the control of the human tissue plasminogen activator (TPA) signal or the native P. falciparum MSP4 signal. Only the construct containing the TPA signal induced detectable antibodies in mice, although gene expression was demonstrated in all constructs and MSP4 was shown to be secreted using either signal by in vitro transient transfection of COS cells. Two recombinant MSP4 proteins that encoded the same sequence as the plasmid DNA were produced in E. coli (EcMSP4-His) and S. cerevisiae (yMSP4-His) and used to raise antibodies in mice. Comparison of the antibodies elicited by these various antigen formulations showed differences in titer, isotype and epitope recognition. The titer of antibodies induced by DNA vaccination was lower than that induced by yMSP4-His, which in turn was lower than that induced by EcMSP4 His. The isotype profiles of the antibodies were also different, the plasmid DNA induced predominantly IgG(2a) responses whereas the two proteins induced predominantly IgG(1) responses. The antibodies induced by DNA and yMSP4-His recognized predominantly the C-terminal epidermal growth factor (EGF)-like domain of the protein, whereas EcMSP4-His induced antibodies recognizing all domains of the protein equally. The antibodies induced by DNA vaccination were directed almost extensively to conformational epitopes so that reactivity with native MSP4 was abolished after disulfide bonds in the protein were disrupted. Antibodies induced by recombinant proteins recognized linear epitopes as well and reactivity to native MSP4 was preserved after reduction and alkylation of parasite proteins. PMID- 11115705 TI - A new DTPa-HBV-IPV vaccine co-administered with Hib, compared to a commercially available DTPw-IPV/Hib vaccine co-administered with HBV, given at 6, 10 and 14 weeks following HBV at birth. AB - Three hundred and twenty eligible infants were enrolled in an open randomized clinical trial and allocated to one of two groups to receive either separate concomitant injections of a candidate combined DTPa-HBV-IPV and commercial Hib vaccine (candidate administration: DTPa-HBV-IPV+Hib) or separate concomitant injections of licensed DTPw-IPV mixed in the same syringe with Hib and HBV vaccines (comparator administration: DTPw-IPV/Hib+HBV). Vaccines were administered at 6, 10 and 14 weeks of age preceded by a monovalent dose of HBV at birth. The candidate vaccine administration was shown to be at least as immunogenic (primary objective) as the candidate administration with respect to the diphtheria, tetanus, polio, HBs and PRP seroprotection rates (primary endpoints). Post vaccination, both vaccine administrations showed an equivalent level of seroprotection with nearly all subjects (>96%) acquiring seroprotective titers against diphtheria, tetanus, polioviruses, HBsAg and PRP antigens. A markedly higher anti-HBs response post dose 2 at week 14 in the group receiving the candidate vaccine, 98.6% of subjects had seroprotective titers (GMT of 505.7 mIU/ml) compared with only 88.7% (GMT of 107.5 mIU/ml) in the comparator group. There was a lower incidence of adverse events following the DTPa-based candidate administration compared with the DTPw-based comparator. Despite the early age and short interval between doses, both administrations were immunogenic, with the concomitant administration of DTPa-HBV-IPV and Hib vaccines showing an improved tolerability over the commercial vaccines DTPw-IPV/Hib and HBV. PMID- 11115706 TI - Xerovac: an ultra rapid method for the dehydration and preservation of live attenuated Rinderpest and Peste des Petits ruminants vaccines. AB - The accepted procedure for the long-term preservation of live viruses and bacteria in vaccines has been lyophilisation. We show that thermolabile viruses can be dehydrated in vitro, within 18 h, in an excipient containing trehalose. We further demonstrate that in the resulting dehydrated state, where the viruses are captive in a metastable glass composed of trehalose, they are capable of resisting 45 degrees C for a period of 14 days with minimal loss of potency. The degree of thermotolerance achieved matches that of current 'thermostable' lyophilised vaccines, but with the distinct advantage of a shorter, cheaper and simpler process. The development and utilisation of this process can make significant improvements in current live virus vaccine production. It presents a further step away from dependence on mandatory low temperature refrigerated storage and could lead to greater confidence in vaccine stability, potency and efficacy. PMID- 11115707 TI - Protection of cattle against rinderpest by intranasal immunisation with a dry powder tissue culture vaccine. AB - Dry powder tissue culture rinderpest vaccine containing 10(2.5) TCID(50) of virus per dose administered intranasally to cattle induced high titre circulating antibody responses and protection against challenge with a virulent strain of rinderpest virus. A reduction in the dose of virus to 10(1.1) TCID(50) resulted in a failure to elicit detectable antibody responses and a lack of protection. Intranasal powder vaccine offers several advantages over conventional needle administered aqueous rinderpest vaccine, including greater stability in the absence of a cold chain, reduced risk of 'needle transfer' of other microbial agents present in the vaccinated herd and lower cost. PMID- 11115708 TI - Prevalence of hepatitis A virus infection in sewage plant workers of Central Italy: is indication for vaccination justified? AB - Prevalence of antibodies to hepatitis A virus (HAV) was studied in a group of 65 sewage plant workers living in Tuscany, Central Italy. In order to evaluate the effect of several confounders (age, place of birth, income, educational degree, sea-food consumption, etc.), subjects under study were matched with 160 other workers residing in the same area. Anti-HAV was detected in about 51% of sewage workers and 44% of other employees. The difference was not statistically significant. Both univariate and multivariate analysis showed that the main variables related to previous HAV infection were increasing age (P<0.001), birth in Southern Italy (P<0.01) and lower educational degree (P<0.001). Although other studies in Northern and Central Europe showed a slightly higher risk of infection in sewage workers versus general population, lack of evidence of occupational risk in Italy might be explained by the relative importance of a higher degree of viral circulation in the past. The changing epidemiology of HAV infection in Italy with increasing numbers of susceptibles in adults and the potential occupational risk suggest that the present indication to immunize sewage plant workers against hepatitis A should be maintained. PMID- 11115709 TI - Preparation and preclinical evaluation of experimental group B streptococcus type III polysaccharide-cholera toxin B subunit conjugate vaccine for intranasal immunization. AB - Streptococcus group B (GBS) is usually carried asymptomatically in the vaginal tract of women and can be transferred to the newborn during parturition. Serum antibodies to the capsular polysaccharide (CPS) can prevent invasive diseases, whereas immunity acting at the mucosal surface may be more important to inhibit the mucosal colonization of GBS and thus the risk of infection for the newborn. We prepared different GBS type III CPS-protein conjugate vaccines and evaluated their systemic and mucosal immunogenicity in mice. GBS type III CPS was conjugated to tetanus toxoid (TT) or recombinant cholera toxin B subunit (rCTB) either directly or to rCTB indirectly via TT. The conjugation was performed by different methods: (1) CPS was coupled to TT with 1-ethyl-3 (3 dimethylaminopropyl)-carbodiimide (EDAC), using adipic acid dihydrazide (ADH) as a spacer; (2) CPS was conjugated with rCTB using reductive amination; or, (3) N succinimidyl 3-(2-pyridyldithio) propionate (SPDP) was used to bind rCTB to the TT of the CPS-TT conjugate. Mice were immunized with these conjugates or purified CPS by subcutaneous (s.c.) and intranasal (i. n.) routes. Antibodies to GBS III in serum, lungs and vagina were measured with ELISA. All of the CPS-protein conjugates were superior to unconjugated CPS in eliciting CPS-specific immune responses in serum and mucosal tissue extracts. The conjugates, when administrated s.c., induced only IgG responses in serum, lung and vagina, while i.n. vaccination also elicited IgA responses in the lungs and vagina. The CPS-TT conjugate administrated i.n. induced a strong serum IgG, but only a weak mucosal IgA response, while the CPS-rCTB conjugate elicited high IgG as well as IgA antibodies in the lungs after i.n. immunization. GBS III CPS-TT conjugated with rCTB produced a strong systemic and local anti-CPSIII response after i.n. administration. Co-administration of CT as adjuvant enhanced the anti-CPS systemic and mucosal immune responses further after i.n. administration with the CPS conjugates. These findings indicate that: (i) i.n. immunization with GBS CPS protein conjugates was more effective than s.c immunization for stimulating serum as well as mucosal immune responses; (ii) rCTB as a carrier protein for GBS III CPS could markedly improve the mucosal immune response; and (iii) the experimental GBS type III CPS conjugates containing rCTB should be investigated as mucosal vaccine to prevent GBS infection in humans. PMID- 11115710 TI - A systematic approach to vaccine complexity using an automaton model of the cellular and humoral immune system. I. Viral characteristics and polarized responses. AB - A modern approach to vaccination faces the compound complexity of microorganism behavior and immune response triggering and regulation. Since computational modeling can yield useful guidelines for biological experimentation, we have used IMMSIM(3), a cellular automaton model for simulating humoral- and cell-mediated responses, to explore a wide range of virus-host relations. Sixty-four virtual viruses were generated by an assortment of speed of growth, infectivity level and lethal load. The outcome of the infections, as influenced by the immune response and the bolstering of cures, obtained by vaccine presensitization are illustrated in this first article. The results of the in machina experiments allow us to relate the success rate of responses to certain combinations of viral parameters and by freezing one or the other branch, and to determine that some viruses are more susceptible to humoral, and others to cellular responses, depending either on single parameters or combinations thereof. This finding allows prediction of which infection may be susceptible to polarized ((Th)(1)>Th(2) and Th(1)/=200x10(6)/l CD4(+) T lymphocytes (group 2) and healthy controls. Thirty days after PPV vaccination the GMC against PPS 6B, 14, 19F and 23F in group 1, and against 6B and 19F in group 2, were significantly lower compared with healthy controls. Both in HIV-infected and in healthy individuals who received CPV and PPV the postvaccination GMC against PPS 14, 19F and 23F were higher compared with historical controls who were not previously immunized with CPV but only received PPV. We conclude that the antibody response to CPV is impaired in HIV-infected individuals. Higher antibody concentrations were achieved in HIV-infected and healthy individuals after sequential vaccination with CPV and PPV compared with PPV vaccination alone. PMID- 11115713 TI - Solid matrix-antibody-antigen complexes incorporating equine herpesvirus 1 glycoproteins C and D elicit anti-viral immune responses in BALB/c (H-2K(d)) and C3H (H-2K(k)) mice. AB - Glycoproteins C and D (gC and gD) derived from equine herpesvirus 1 (EHV-1) infected cells were incorporated into individual solid matrix-antibody-antigen (SMAA) complexes and administered to BALB/c (H-2K(d)) and C3H (H-2K(k)) mice. Antibodies against each of the glycoproteins were produced that neutralised virus infectivity and mediated the lysis of EHV-1-infected target cells in the presence of complement. Immunoglobulin (Ig)G2b was the predominant antibody isotype produced in BALB/c mice against gC, while equal amounts of IgG2a/2b were found in the serum of C3H mice (indicative of a T-helper(1) response). Glycoprotein D immunisation elicited predominantly an IgG1 response in BALB/c mice (indicative of a T-helper(2) response) and an IgG2a/2b response in C3H mice. EHV-1-specific local and systemic T-cell proliferative responses were detected in vitro following administration of SMAA complexes. Suppression of the local T-cell response was seen following virus challenge of mice immunised with SMAA gC. SMAA gD provided some protection against intranasal EHV-1 challenge. These data show that the SMAA system is an effective way of presenting subviral components to the immune system and further emphasises the importance of including glycoprotein D as a component of a subunit EHV-1 vaccine. PMID- 11115714 TI - Immunogenicity of a hexavalent combination vaccine in rhesus monkeys. AB - Preclinical immunogenicity studies were conducted in rhesus monkeys to determine whether there is immune interference in the response to one or more components of a hexavalent vaccine (Hexavac) that contains antigens from Haemophilus influenzae (Hib), hepatitis B (HB), diphtheria (D), tetanus (T), acellular pertussis (aP) and inactivated polio virus (IPV). Antibody responses were measured following co administration of the components at three separate anatomical sites or administration as a hexavalent combination in a single site. After three injections of the hexavalent vaccine, the peak antibody responses to each component of the vaccine were >100-fold above pre-immune titers and persisted at levels >10-fold above pre-immune titers at approximately 1 year. Immune interference was observed in the peak response to HB, D and pertussis toxin, but was not seen at later time points. The results indicate that the rhesus monkey model may be useful for pre-clinical evaluation of combination vaccines. PMID- 11115715 TI - A survey of physicians' vaccine risk perception and immunization practices for subjects with immunological diseases. AB - Various publications have caused concern by implying that immunization may be linked to new cases or flare-ups of immunological diseases (IDs). In view of the resulting uncertainty, we studied physicians' vaccine risk perception and immunization practices for adults with IDs. A questionnaire was mailed to three groups of physicians in France: internal medicine specialists, general practitioners, and travel clinic physicians. Thirteen vaccines currently used for adults in France were studied. Risk perception was rated on a 10 cm visual analog scale (VAS). The distribution of the answers was compared between and within groups of physicians. Potential associations between risk perception and reported practices were investigated by multivariate analysis. In the three groups of physicians (n=762), the tetanus and Salk poliomyelitis vaccines had the lowest risk perception. The yellow fever, BCG and Sabin poliomyelitis vaccines were the least well perceived. The distribution of risk perception for these three live vaccines and the hepatitis B vaccine was uniform according to VAS grading. For the other vaccines studied, the distribution was skewed to the low-risk perception side of the VAS. Risk perception was greater for physicians who stated: (1) that certain IDs carried a high risk of adverse events following immunization; (2) that they sought the advice of the referent physician before immunization; (3) warned their patients of the risk of an ID flare-up after vaccination; (4) sought information about recent immunization in patients with a flare-up; and (5) had experience of the side effects of immunization in adults with ID. Risk perception was lower for physicians who said they updated immunizations, and for the internists. The worse the vaccine risk perception by physicians, the more uniform the distribution of perception, thus reflecting the disagreement of the scientific community about the risk of using such vaccines for adults with an ID. Risk perception and immunization practices were related in adults with ID. Understanding of decisions concerning immunization may help to improve immunization updating and prevent risk amplification when evidence is lacking. PMID- 11115716 TI - The postmarketing safety profile of varicella vaccine. AB - The postmarketing safety profile of varicella vaccine was evaluated by analyzing selected adverse experience reports temporally associated with the administration of the vaccine. There were 7963 reports voluntarily submitted to Merck for an overall reporting rate of 5.0 per 10000 doses of vaccine distributed. A varicella zoster virus (VZV) identification program detected the presence of the Oka vaccine strain in three individuals with an immune deficiency - two with pneumonia and one with hepatitis - and in three instances of secondary transmission from vaccinees with vesicular lesions to susceptible household contacts. The Oka vaccine strain was present in 23 patients and wild-type VZV was present in 15 patients with herpes zoster. Vesicular rashes that occurred within 2 weeks of vaccination were more likely to contain the presence of wild-type VZV, while vesicular rashes that occurred more than 2 weeks post-vaccination were more likely to contain the Oka vaccine strain. Eleven patients were hospitalized with complications of breakthrough varicella infection. PMID- 11115717 TI - A novel ELISA for determination of polysaccharide specific immunoglobulins. AB - A novel ELISA using Nunc CovaLink microtiter plates has been developed for the determination of polysaccharide specific antibodies in mice sera. Glucuronoxylomannan (GXM), a major capsular polysaccharide of Cryptococcus neoformans serotype B, was immobilised on CovaLink plates by covalent linkage of CNBr activated hydroxyl groups of the sugar and free bnd NH(2) groups of the plates. The binding characteristics of GXM to CovaLink and to conventional polystyrene ELISA plate (Costar) were compared. The differences were observed in quality of standard curve with anti-GXM MoAb 2H1 (R(2) values were 0.9468 and 0.9872 for Costar and CovaLink plates, respectively) and in absorbance values of sera of immunised mice which were 2.5 times higher on CovaLink than on Costar plates. Negative control was low and having the same value on both the plates. Using the novel ELISA we have analysed the influence of immunomodulatory peptidoglycan monomer on humoral immune response to GXM in mice. The treatment with the conjugate of the immunomodulator and GXM resulted in the increase of GXM specific IgGs in mice sera. Finally, the method has been successfully modified for the determination of dextran-specific antibodies in mice sera, indicating that the described procedure could be applied for other polysaccharides that have &z.sbnd;OH groups available for CNBr activation. PMID- 11115718 TI - Adjuvant action of Escherichia coli enterotoxin for delayed-type hypersensitivity to Oka vaccine virus on pernasal co-administration in mice. AB - The usefulness of a mutant of Escherichia coli enterotoxin for the induction of cellular immunity to varicella-zoster virus as a mucosal adjuvant is assessed in mice. When a commercially available live varicella vaccine (the Oka strain) and toxin were once administered simultaneously via the nasal route, delayed-type hypersensitivity to Oka vaccine virus was significantly induced and detected by footpad test in mice. Moreover, when spleen cells from mice immunized with the vaccine and toxin were re-stimulated with live vaccine in vitro, they showed more thymidine uptake and produced more IL-2 than those from mice immunized with the vaccine alone. These results suggest that mutant enterotoxin has adjuvant action to induce a specific delayed-type hypersensitivity to Oka vaccine virus on nasal co-administration with live vaccine virus. PMID- 11115719 TI - A comparison of two commercial recombinant vaccines for hepatitis B in adolescents. AB - The immunogenicity and reactogenicity profiles of three doses each of Engerix B(R) (10 microg hepatitis B surface antigen) and Recombivax(R) (5 microg hepatitis B surface antigen), given on a 0, 1, 6 month schedule to healthy adolescents were compared in a single-blind, randomized clinical trial. One month following the third dose, seroprotection rates after Engerix-B and Recombivax were similar (99 and 98%, respectively). The geometric mean titre (GMT) was statistically significantly higher following vaccination with Engerix-B (3961 vs. 1001 mIU/ml; P=0.0001, Fisher's exact test). Most of the symptoms reported were mild or moderate in intensity and transient. There were no vaccine-related serious adverse events. PMID- 11115720 TI - Impact of hepatitis B immunisation as part of the EPI. AB - Since 1992, hepatitis B vaccine has been an integrated part of Thailand's expanded programme on immunisation (EPI). Based on the data from five representative provinces, we have evaluated its impact on the countrywide prevalence of HBV infection and carrier rate. The population studied comprised 400-488 healthy and immuno-competent, subjects per area. The subjects' ages ranged from 6 months to 18 years. We examined their sera for viral hepatitis markers using commercially available test kits and established the coverage rate of hepatitis B vaccination after its inclusion into the EPI to be 71.2-94.3%. The number of individuals undergoing the complete course of vaccinations had increased four-fold. Consequently, only 0.7% of the children born after the implementation of this the novel EPI strategy were HBV carriers. PMID- 11115721 TI - CpG DNA is an effective oral adjuvant to protein antigens in mice. AB - We have previously reported that synthetic oligodeoxynucleotides containing immunostimulatory CpG motifs (CpG ODN) are potent adjuvants to protein administered by intramuscular (IM) injection or intranasal (IN) inhalation to BALB/c mice. Herein, we have evaluated oral delivery of CpG ODN with purified hepatitis B surface antigen (HBsAg) or tetanus toxoid (TT) to determine its potential as an adjuvant to oral vaccines. CpG ODN augmented systemic (IgG in plasma, CTL, T-cell proliferation) and mucosal (IgA in lung, vaginal or gut washes, feces and saliva) immune responses against both antigens. CpG stimulated both T-helper type 1 (Th1) (CTL, IgG2a) and Th2 (IgG1, IgA) responses when delivered orally. Results from this study indicate that stimulatory CpG ODN may be effective as an adjuvant with oral vaccines. PMID- 11115722 TI - Immunogenicity of a foreign peptide expressed within a capsid protein of an attenuated coxsackievirus. AB - The immunogenicity of soluble peptides can be improved by expression within recombinant microorganisms. The immunogenicity of a peptide expressed within a capsid protein of an attenuated coxsackievirus B4 was evaluated. The insertion site was chosen based on its antigenic structure. A foreign peptide was inserted into a region of the VP1 capsid protein that was identified as a T helper cell epitope. A recombinant virus containing ten amino acids of ovalbumin sequence was genetically stable and retained the biological and physical characteristics of the parental virus. The recombinant was able to elicit a T helper cell response against ovalbumin sequences. This study shows, for the first time, that coxsackievirus can be used as an expression vector and that insertion of heterologous peptides into an immunogenic region is a viable strategy for inducing T helper cell responses against foreign sequences. The implications of this work are that the attenuated coxsackievirus variant may be useful as a vaccine vector for expressing T helper cell epitopes that are important in inducing protective immunity. PMID- 11115723 TI - A novel strategy for protective Actinobacillus pleuropneumoniae subunit vaccines: detergent extraction of cultures induced by iron restriction. AB - We have characterized antigens from Actinobacillus (A.) pleuropneumoniae grown under iron restriction with respect to their immunogenic and protective potential. Antigens were the cell-free culture supernatants (CFS) obtained after treatment of A. pleuropneumoniae broth cultures with sodium deoxycholate. Using the iron-repressible transferrin-binding lipoprotein TbpB and the constitutively expressed outer membrane lipoprotein OmlA as markers, we have shown that the detergent extraction enriched the CFS with lipoproteins from the outer membrane (OM). Extractions with 0.05% of sodium deoxycholate increased the lipoprotein contents in the CFS, but did not affect the integrity of the OM. This was demonstrated by the absence of the iron-repressible integral OM transferrin binding protein TbpA. Furthermore, the absence of periplasmic and cytoplasmic proteins in CFS after extraction was determined in immunoblot analyses with anti bacterial alkaline phosphatase and anti-Hsp60 antisera, demonstrating that there was no rupture of the OMs or the plasma membranes due to the extraction procedure. Antigen preparations from A. pleuropneumoniae serotype 2 and 9 grown under iron restrictive conditions were combined, emulsified, and tested for their ability to confer protection in pigs. Pigs immunized with CFS from sodium deoxycholate extracted cultures developed a strong antibody response and, upon challenge with A. pleuropneumoniae serotype 2, the immunized pigs showed no or only mild clinical signs of disease and had a significantly lower degree of lung damage than the control pigs. PMID- 11115724 TI - Rhesus rotavirus-based quadrivalent vaccine is efficacious despite age, socioeconomic conditions and seasonality in Venezuela. AB - This report describes the results of additional analyses of the trial carried out with the rhesus rotavirus-based quadrivalent vaccine in Venezuela. In the present study, we re-examined the data from this previous rotavirus vaccine trial to assess the statistical interaction between vaccine efficacy and (i) the duration of efficacy into the second year of life, (ii) socioeconomic conditions, and (iii) rotavirus seasonality. We found that among Venezuelan children, the rotavirus vaccine confers protection against severe diarrhea during the first 2 years of life independently of socioeconomic conditions and seasonality. PMID- 11115725 TI - Both immunisation with a formalin-inactivated respiratory syncytial virus (RSV) vaccine and a mock antigen vaccine induce severe lung pathology and a Th2 cytokine profile in RSV-challenged mice. AB - Respiratory syncytial virus (RSV) is the most important cause of bronchiolitis and pneumonia in infants and young children. Immunopathology may play a role in RSV-induced disease and a severe RSV infection may also be associated with an increased risk of developing asthma. Vaccination with formalin-inactivated RSV (FI-RSV) prior to infection resulted both in human and in the mouse model in extensive lung pathology. In the mouse model, it has been shown that this aggravation of disease was associated with a shift in the balance between Th1 and Th2 cytokines towards a Th2-type response. The aim of the present study was to characterise the immunological and inflammatory responses in BALB/c mice upon RSV infection with or without prior vaccination with aluminium-adjuvanted FI-RSV or control antigens (FI-Mock). As previously reported by others, we also observed that a primary RSV infection in BALB/c mice resulted in a predominant Th1-type cytokine response, which was associated with slight bronchiolitis and alveolitis. FI-RSV vaccination prior to RSV challenge prevented virus replication and was associated with an aggravation of pulmonary histopathology and a shift towards a Th2-type response. Vaccination with FI-Mock did not prevent RSV replication in the lung but resulted in an even more pronounced Th2 response after infection while these mice were not sensitised to specific viral antigens. Thus, viral replication in a Th2 responding animal (induced by aluminium-adjuvanted mock vaccine) appears to boost the Th2 response upon RSV infection. PMID- 11115726 TI - A truncated variant of the hepatitis C virus core induces a slow but potent immune response in mice following DNA immunization. AB - Vaccination of BALB/c mice with pIDKCo, a plasmid containing the coding sequence for the first 176 amino acids of the hepatitis C virus (HCV) core protein, induced both humoral and cellular specific immune responses. Particularly, the level of anti-core antibodies increased slowly with time up to a mean value above 1:8000 that was generally superior than that found in anti-HCV positive individuals. Six out of nine anti-HCV positive human sera were able to inhibit at different extent the binding of mouse anti-core sera to a recombinant capsid protein. Our results show that it is possible to elicit a potent humoral and cellular immune response against the HCV core antigen in mice following DNA immunization. PMID- 11115727 TI - Pharmacology and behavioral pharmacology of the mesocortical dopamine system. AB - The prefrontal cortex (PFC) has long been known to be involved in the mediation of complex behavioral responses. Considerable research efforts are directed towards refining the knowledge about the function of this brain area and the role it plays in cognitive performance and behavioral output. In the first part, this review provides, from a pharmacological perspective, an overview of anatomical, electrophysiological and neurochemical aspects of the function of the PFC, with an emphasis on the mesocortical dopamine system. Anatomy of the mesocortical system, basic physiological and pharmacological properties of neurotransmission within the PFC, and interactions between dopamine and glutamate as well as other transmitters within the mesocorticolimbic circuit are included. The coverage of these data is largely restricted to what is relevant for the second part of the review which focuses on behavioral studies that have examined the role of the PFC in a variety of phenomena, behaviors and paradigms. These include reward and addiction, locomotor activity and sensitization, learning, cognition, and schizophrenia. Although the focus of this review is on the mesocortical dopamine system, given the intricate interactions of dopamine with other transmitter systems within the PFC and the importance of the PFC as a source of glutamate in subcortical areas, these aspects are also covered in some detail where appropriate. Naturally, a topic as complex as this cannot be covered comprehensively in its entirety. Therefore this review is largely limited to data derived from studies using rats, and it is also specifically restricted to data concerning the medial PFC (mPFC). Since in several fields of research the findings concerning the function or role of the mPFC are relatively inconsistent, the question is addressed whether these inconsistencies might, at least in part, be related to the anatomical and functional heterogeneity of this brain area. PMID- 11115728 TI - Aquaporins in the central nervous system. AB - In this review, we have tried to summarize most available data dealing with the aquaporin (AQP) family of water channels in the CNS. Two aquaporins have been identified so far in the CNS, AQP1 and AQP4. AQP1 is restricted to the choroid plexus of the lateral ventricles, which raises a role for this aquaporin in cerebrospinal fluid formation. AQP4 is the predominant water channel in the brain and it is more widely distributed than originally believed, with a marked prevalence over periventricular areas. In the first part of this review, we examine the complete distribution pattern of AQP4 in the CNS including its rostro caudal localization to end with its subcellular location. After discussing scarce data dealing with regulation of aquaporins in the CNS, we focus in potential roles for aquaporins. Novel recent data highlights very important roles for this aquaporin in the normal and pathological brain including, among others, role in potassium buffering, body fluid homeostasis, central osmoreception and development and restoration of brain edema. PMID- 11115729 TI - Developmental mechanisms in the pathogenesis of neurodegenerative diseases. AB - Cellular genes that are mutated in neurodegenerative diseases code for proteins that are expressed throughout neural development. Genetic analysis suggests that these genes are essential for a broad range of normal neurodevelopmental processes. The proteins they code for interact with numerous other cellular proteins that are components of signaling pathways involved in patterning of the neural tube and in regional specification of neuronal subtypes. Further, pathogenetic mutations of these genes can cause progressive, sublethal alterations in the cellular homeostasis of evolving regional neuronal subpopulations, culminating in late-onset cell death. Therefore, as a consequence of the disease mutations, targeted cell populations may retain molecular traces of abnormal interactions with disease-associated proteins by exhibiting changes in a spectrum of normal cellular functions and enhanced vulnerability to a host of environmental stressors. These observations suggest that the normal functions of these disease-associated proteins are to ensure the fidelity and integration of developmental events associated with the progressive elaboration of neuronal subtypes as well as the maintenance of mature neuronal populations during adult life. The ability to identify alterations within vulnerable neuronal precursors present in pre-symptomatic individuals prior to the onset of irrevocable cellular injury may help foster the development of effective therapeutic interventions using evolving pharmacologic, gene and stem cell technologies. PMID- 11115730 TI - Decreased exploratory activity and impaired passive avoidance behaviour in mice deficient for the alpha(1b)-adrenoceptor. AB - There is growing evidence that a dysfunction of central noradrenergic neurotransmission is involved in age-related impairments of cognitive performance and the pathophysiology of Alzheimer's disease (AD). A reduction of density of central alpha(1)-adrenergic receptors (alpha(1)-AR) has been shown in aging and AD brains. Three alpha(1)-AR subtypes (alpha(1a), alpha(1b) and alpha(1d)) have been identified by molecular cloning. However, very little is known about the functional role of distinct alpha(1)-AR subtypes in the brain. This problem was specifically addressed using a model of knockout mouse deficient in alpha(1b)-AR (alpha(1B)-/-) because these animals show a 40% reduction of alpha(1)-AR density in the brain as already reported. In comparison to the wild-type mice (alpha(1B)+/+), alpha(1B)-/- mice showed significantly reduced square entries and a reduced rearing behaviour was observed over all sessions in the open field. In passive avoidance procedures, alpha(1B)-/- mice showed a tendency towards decreased short-term-latency and a significant decline in long-term-latency. The present results indicate that mutation of a single member of the alpha(1)-AR gene family creates a distinct phenotype and provide evidence that alpha(1B)-AR is possibly involved in modulation of memory consolidation and fear-motivated exploratory activity. Furthermore, this model of knockout mice may be useful in elucidating the role of alpha(1B)-AR in dementias involving deficits of the noradrenergic system. PMID- 11115731 TI - Administration of high-dose ketoconazole, an inhibitor of steroid synthesis, prevents posttraumatic anxiety in an animal model. AB - Acute psychological stress is the presumed immediate cause of post-traumatic stress disorder (PTSD), and may also contribute to other anxiety disorders. Abnormal activity of the hypothalamic-pituitary-adrenal (HPA) axis has been tentatively implicated in some of the features of these disorders. Ketoconazole (KTCZ), an imidazole derivative, is a potent inhibitor of gonadal and adrenal steroidogenesis. The aim of this study was to explore the effects of KTCZ blockade of adrenal steroidogenesis, and consequent elevation of adreno corticotropic hormone (ACTH), on a model of chronic post-traumatic anxiety in rats. Amelioration of anxious behaviors after reduction of corticosterone would suggest that corticosterone (and by implication cortisol in humans) is an important mediator of anxious symptoms: exacerbation of such behaviors would suggest that corticosterone elevations are only secondary, and possibly implicate corticotropin releasing hormone (CRH) and/or ACTH in the pathogenesis of anxious symptoms. We exposed rats for 10 min to cat scent, a prima facie valid model for acute psychological stress, with and without high dose KTCZ for 14 days. Treatment with KTCZ abolished the chronic behavioral effects of acute exposure to a cat scent. Lower levels of anxious behavior in KTCZ-treated and exposed rats were accompanied by lower plasma corticosterone, ACTH and prolactin (PRL) levels compared to untreated exposed rats. Results in this model implicate corticosterone, but not ACTH, in the pathogenesis of chronic anxiety following acute psychological stress. PMID- 11115732 TI - Effects of acute or chronic administration of substituted benzamides in experimental models of depression in rats. AB - The effects of substituted benzamides, sulpiride and raclopride on experimental models of depression were studied in male rats after acute or chronic administration in comparison to those of the classical antidepressant, clomipramine. In contrast to clomipramine (50 mg/kg), acute doses of sulpiride or raclopride (1 or 5 mg/kg) failed to change the behavioral response of animals tested in the despair (constrained swim) test or in the model of reserpine induced changes in the open field behavior. These doses also did not modify the grooming response of rats exposed to a novel environment. Sulpiride or raclopride 10 mg/kg increased the immobility time in the despair test and reduced novelty induced grooming. The repeated injection for 21 days of sulpiride or raclopride (1 or 5 mg/kg, but not 10 mg/kg) induced a reduction of the immobility period during the constrained swim test similar to that following the chronic treatment with clomipramine 50 mg/kg. This appeared to be a clear-cut reversed dose response relationship for both substituted benzamides, being the dose potency 1 mg/kg>5 mg/kg>10 mg/kg. Raclopride was more potent than sulpiride in this respect. Furthermore, like clomipramine, sulpiride (1 or 5 mg/kg) and raclopride (1 mg/kg) antagonized reserpine-induced changes in the open field behavior and enhanced novelty-induced grooming. These results indicate that, in contrast to acute injection, repeated administration of small doses of the substituted benzamides, sulpiride or raclopride induce an effect similar to that of the classical antidepressant, clomipramine. The reverse dose-response relationship suggests that these drugs in small doses act on presynaptic dopamine D(2) receptors. This may be consistent with a postsynaptic action of greater doses that exert sedative effects and increase immobility time in the despair test. PMID- 11115733 TI - Acute antipsychotic drug administration lowers body temperature in drug-free male schizophrenic patients. AB - We examined, in a controlled design, potential alterations in body temperature of male schizophrenic patients following acute antipsychotic drug (APD) administration. Fourteen drug-free (study group) and seven schizophrenic patients maintained on APDs (comparison group) initiated or received higher dose of their APD, respectively, for 27 days. Initial body temperature was 0.36 degrees C higher in the study group (P=0.01) and decreased within 24 h to values comparable to that of the comparison group (all within normal range). PMID- 11115734 TI - Influence of buprenorphine, butorphanol and nalbuphine on the initiation of intravenous cocaine self-administration in drug naive mice. AB - The influence of different mixed mu-kappa-opioid receptor agonists-antagonists on cocaine reinforcement was studied using the method of initiation of intravenous cocaine self-administration in naive mice. Self-administration of cocaine was readily initiated according to an inverted U-shaped unit dose-response curve. Buprenorphine, butorphanol and nalbuphine tested against the optimal unit dose of cocaine (0.8 microg per infusion), inhibited initiation of cocaine self administration in a dose-dependent manner. When tested against a scale of cocaine unit doses (0.2 -1.6 microg per infusion) buprenorphine (0.1 mg/kg, s.c.) and nalbuphine (2 mg/kg, s. c.) produced a shift of the optimal cocaine dose from 0.8 to 0.4 microg/inf, while butorphanol (1 mg/kg, s.c.) shifted the optimal unit dose of cocaine to the right along the cocaine unit doses axis. Co-administration of naloxone (0.1 mg/kg, s.c.) significantly reduced the effect of buprenorphine but failed to influence the effect of nalbuphine and butorphanol on cocaine intake. Taken together, these results suggest that nalbuphine is capable of affecting cocaine's reinforcing properties in the same manner as buprenorphine during the initiation phase of cocaine self-administration behavior, while butorphanol causes the opposite effect. Although the exact opioid profile of action of the mixed opioid receptor agonists-antagonists is as yet not precisely known, the present findings suggest that multiple opioid receptor systems (i.e. mu and kappa) play a role in reinforcing properties of cocaine and that a co operative interaction between mu- and kappa-opioid systems may be of importance during initiation of cocaine self-administration. PMID- 11115735 TI - Local modulation of the 5-HT release in the dorsal striatum of the rat: an in vivo microdialysis study. AB - Using in vivo microdialysis in freely moving rats, we examined the involvement of major striatal transmitters on the local modulation of the 5-HT release. Tetrodotoxin reduced the striatal 5-HT output to 15-20% of baseline. The selective 5-HT(1B) receptor agonist CP 93129 (50 microM) reduced (50%) and the 5 HT(2A/2C) receptor agonist DOI (1-100 microM) increased (220%) the 5-HT output. Neither GABA nor baclofen (100 nM-100 microM) altered the 5-HT output. The glutamate reuptake inhibitor L-trans-PDC (1-4 mM) raised 5-HT to 280% of baseline. This effect was not antagonized by the NMDA receptor antagonist MK-801 (0.5 mg/kg i.p.). Local MK-801 (10-100 microM) did not significantly alter the 5 HT output. Finally, neither carbachol (10-100 microM) nor quipirole (10 microM-1 mM) affected 5-HT. These data suggest that the striatal 5-HT release is influenced by local serotonergic and glutamatergic (but not GABAergic) inputs. PMID- 11115736 TI - Nicotine modulates nitric oxide in rat brain. AB - Nicotine exerts its central actions by regulating cationic fluxes through nicotinic acetylcholine receptors (nAChRs). By this effect, the drug likely also modifies events occurring beyond the nAChR, including the regulation of nitric oxide (NO) synthesis. The present study was undertaken to assess the effects of acute and chronic nicotine administration (0.4 mg/kg, s.c.) on levels of NO( )(2)+NO(-)(3), stable metabolites of NO, in brain regions of male and female rats. Nicotine increased levels of the metabolites, and therefore presumably of NO, with sex differences in the degree of stimulation, the brain regions affected, and the variance between the effects of acute and chronic administration. Prior inhibition of NO synthase eliminated the effect of nicotine in all regions studied. While nicotine appeared to increase NO indirectly via glutamate receptors in the cortex and hippocampus, this was not true of the corpus striatum, where blocking NMDA-type glutamate receptors with MK-801 had no effect. The findings support the view that NO is likely involved in some of the central effects of nicotine. PMID- 11115737 TI - Anti-depressant action of melatonin in chronic forced swimming-induced behavioral despair in mice, role of peripheral benzodiazepine receptor modulation. AB - The possible antidepressant effect of physiological and pharmacological doses of melatonin was investigated in the Porsolt forced swimming-induced behavioral despair test. The duration of immobility period of BALB/c and C57BL/6J mice during a 6-min swim test was measured at noon (11:00-12:00 h), early dark (20:00 21:00 h) and at midnight (1:00-2:00 h), respectively. The circadian time cycle did not alter the duration of immobility in either strains of mice. Similarly, exogenously administered melatonin (10-1000 microg/kg congruent with 50 nM to 5 microM/mouse), a dose that could act on high affinity melatonin receptors, did not modify the duration of immobility period at any of the time intervals studied in either strains of the mice. This suggested that neither circadian variation influenced the duration of immobility period of BALB/c and C57BL/6J mice nor at physiological doses melatonin showed any anti-depressant action. Acute administration of higher doses of melatonin (2.5-10 mg/kg) failed to induce any anti-depressant activity in mice which were subjected to forced swimming test for the first time. However, daily administration of melatonin (2.5-10 mg/kg) prior to swimming test significantly reversed the increase in immobility period that was observed on chronic exposure to swimming test. This effect was comparable with the effect of GABA-benzodiazepine (BZ) receptor agonists. Similarly, like GABAergic drugs, acute administration of melatonin also showed anti-depressant activity in a mice which were exposed to chronic forced swimming test. The anti depressant action of melatonin was sensitive to reversal by peripheral BZ receptor antagonist, PK11195. Whereas, flumazenil failed to reverse the anti depressant action of melatonin, thereby suggesting that central BZ receptor were not involved in its action. In conclusion the study showed that at pharmacological doses melatonin has anti-depressant action in chronic forced swimming-induced despair behavior by an action involving peripheral BZ receptors. PMID- 11115738 TI - Chemical kindling and seizure susceptibility in morphine dependent rats. AB - In the present study, we investigated whether and how chronic morphine administration changes seizure susceptibility in rats. The role of morphine dependence on the seizure susceptibility has been evaluated with models of pentylenetetrazol (PTZ)-kindling and acute convulsions induced by PTZ, N-methyl-D aspartic acid (NMDA), picrotoxin and caffeine in adult male rats. The results showed that morphine-dependence increased seizure severity only at 1-4th PTZ injections in the kindling model. In acute convulsion tests, dependent rats demonstrated a significantly lower seizure threshold only for PTZ, while they demonstrated a significantly lower tendency to show tonic-clonic convulsions only for NMDA. It is concluded that morphine-dependence may modulate PTZ-kindling and seizure susceptibility in rats with emphasis on the role of GABA and NMDA neurotransmitter systems. PMID- 11115739 TI - Differential regulation of adenosine A(1) and A(2A) receptors in serotonin transporter and monoamine oxidase A-deficient mice. AB - The serotonin (5HT) transporter (5HTT) removes 5HT from the synaptic cleft and is thus critical to the control of serotonergic neurotransmission. Mice with a targeted inactivation of the 5HTT represent a novel and unique tool to study serotonergic system functioning. Because the release of 5HT is regulated by adenosine, we investigated 5HTT-deficient mice for possible adaptive changes of adenosine A(1) and A(2A) receptors. A(1) and A(2A) receptors were studied by means of quantitative autoradiography using the radioligands [3H]8-cyclopentyl 1,3-dipropylxanthine and [3H]CGS 21680, respectively. A comparison of 5HTT knockout versus control mice revealed upregulation of A(1) receptors in the dorsal raphe nucleus (DRN, +21%), but not in any of the serotonergic projection areas, and downregulation of A(2A) receptors in basal ganglia. The adaptive changes of A(1) and A(2A) receptors in 5HTT-deficient mice are likely to represent a compensatory neuroprotective effect mediated by the adenosinergic modulatory system. For comparison, these receptors were also studied in monoamine oxidase A (MAOA) knockout mice and in 5HTT/MAOA double knockout mice. 5HTT/MAOA double knockout mice showed adaptive changes of adenosine A(1) and A(2A) receptors similar to 5HTT knockout mice, while investigation of MAOA-deficient mice revealed an upregulation of A(2A) receptors, which may relate to a role of both MAOA and adenosine A(2A) receptors in anxiety. PMID- 11115740 TI - Effects of acute and chronic electroconvulsive stimuli on cAMP and cGMP efflux in the rat striatum and hippocampus. AB - The effects of acute and chronic electroconvulsive stimuli (ECS) on extracellular concentrations of the cyclic nucleotides, cAMP and cGMP, from the striatum and hippocampus of awake rats were studied with in vivo microdialysis in conjunction with radioimmunoassay. Acute ECS, but not acute sham-ECS, significantly increased cAMP and cGMP efflux from the striatum by about 75 and 50%, respectively. Chronic ECS did not influence significantly basal efflux of cAMP or cGMP from the striatum or the hippocampus in comparison to control animals receiving chronically sham-ECS. Administration of a challenge ECS in animals treated chronically with sham-ECS resulted in an increase in cAMP and cGMP concentrations in the striatum by 20%, but it failed to affect significantly efflux of these nucleotides in animals treated chronically with ECS. Similarly, in the hippocampus, administration of a challenge ECS in animals treated chronically with sham-ECS resulted in an increase in cAMP and cGMP concentrations by about 40 and 65%, respectively, whereas it failed to affect significantly efflux of these nucleotides in animals treated chronically with ECS. Thus, acutely administered ECS increases cAMP and cGMP efflux in the striatum and hippocampus of rats, an effect that is greatly diminished in animals chronically receiving ECS. These findings suggest changes in the cAMP and cGMP signal transduction mechanisms in response to acute and chronic ECS that may be related to the therapeutic effects of this antidepressant and antipsychotic treatment. PMID- 11115741 TI - Opposing changes in serotonin and norepinephrine transporter mRNA levels after serotonin depletion. AB - We and others have earlier shown that severe serotonin depletion leads to a compensatory down-regulation in the expression of the serotonin transporter (5HTT) gene. We have now investigated the expression of both the 5HTT and the norepinephrine transporter (NET) gene to assess the possible interaction between the noradrenergic and the serotonergic neurotransmitter systems. Acute severe serotonin depletion induced by p-chlorophenylalanine (PCPA) treatment leads to enhanced NET(Long) mRNA levels and reduced 5HTT mRNA level. This change in transporter mRNA expression was paralleled by a non-significant change in protein expression. Chronic severe serotonin depletion combined with treatment with the antidepressant imipramine leads to enhanced NET(Long) mRNA levels. Acute treatment with the monoamine oxidase A inhibitor clorgyline, acute moderate NE reduction (alpha-methyl-p-tyrosine treatment) or less severe depletion for 3 weeks have no effect on the gene expression of the transporters. Taken together, the present data demonstrate that the NET gene expression is enhanced in case of severe serotonin depletion. PMID- 11115742 TI - Pindolol can be effectively given once or twice daily for augmentation in psychiatric patients. PMID- 11115743 TI - Autotransporter proteins, evolution and redefining protein secretion. PMID- 11115744 TI - Mapping of a chromosome replication origin in an archaeon. PMID- 11115746 TI - How the bacterial flora and the epithelial cell get along. PMID- 11115745 TI - Gonococcal lipooligosaccharide: an adhesin for bacterial dissemination? PMID- 11115747 TI - Unlocking the secrets of another minimal genome. PMID- 11115748 TI - Mosquito innate immunity discriminates between different pathogens. PMID- 11115749 TI - Microbial genomics. Genome sequences of an acid-loving scavenger and a ureolytic pathogen. PMID- 11115750 TI - How does Cryptococcus get its coat? AB - During the past few decades, increasing attention has focused on pathogenic fungi both as fascinating research subjects and as the agents of serious illness in diverse patient populations. In particular, opportunistic fungi such as Cryptococcus neoformans command notice as the ranks of their immunocompromised victims grow. C. neoformans is unique among fungal pathogens for its major virulence factor, a complex polysaccharide capsule. In this article, our current understanding of the structure and biosynthesis of the capsule is reviewed, as are the many questions that remain to be answered about how Cryptococcus gets its coat. PMID- 11115751 TI - What is antibiotic resistance and how can we measure it? AB - Antibiotic resistance is being found with increasing frequency in both pathogenic and commensal bacteria of humans and animals. Quantifying resistance within and between bacterial and host populations presents scientists with complex challenges in terms of laboratory methodologies and sampling design. Here, we discuss, from an epidemiological perspective, how antibiotic resistance can be defined and measured and the limitations of current approaches. PMID- 11115752 TI - Viral replication and the coactivators p300 and CBP. AB - Productive viral infection requires coordinate regulation of viral and cellular gene expression. Viruses of different classes have evolved different mechanisms to conform to, adapt to and exploit programs of cellular gene expression. Many viral gene products influence and respond to cellular signals that control differentiation and proliferation Transcriptional coactivators are central to the regulation of the expression of genes controlling these events. p300 and CBP are closely related coactivators that regulate the transcription of specific genes, modify chromatin structure and influence cell cycle progression. In this review, the different molecular interactions of proteins encoded by DNA tumor viruses and lentiviruses with these transcriptional coactivators and related cellular proteins are summarized. PMID- 11115753 TI - Why Helicobacter pylori has Lewis antigens. AB - In mimicry with human gastric epithelial cells, the lipopolysaccharide of Helicobacter pylori expresses Lewis blood group antigens. Recent data suggest that molecular mimicry does not promote immune evasion, nor does it lead to induction of autoantibodies, but that H. pylori Lewis X mediates adhesion to gastric epithelial cells and is essential for colonization. PMID- 11115754 TI - Pentapeptide scanning mutagenesis: encouraging old proteins to execute unusual tricks. AB - Pentapeptide scanning mutagenesis is a facile transposon-based procedure for the random insertion of a variable five amino acid cassette into a target protein. The analysis of a library of proteins harbouring pentapeptide insertions can provide invaluable information on the essential and inessential regions of a target protein, as well as revealing surprising aspects of target protein function and activity. PMID- 11115756 TI - Seeing size and weight. PMID- 11115755 TI - Is perception of causality modular? PMID- 11115757 TI - Monkey hears, monkey does - like infants? AB - Summaries of recently published papers of interest to cognitive scientists. Readers who would like to contribute to this section, by identifying appropriate papers and writing short summaries, should contact the Editor (tics@current trends.com). PMID- 11115758 TI - The function of dynamic grouping in vision. AB - In this review we consider the logic and the evidence relating to the issue of dynamic grouping in human vision. Dynamic grouping is required when the visual system is creating novel descriptions, either because it is dealing with novel stimuli or it is providing information for novel purposes. In such cases, dynamic grouping provides a mechanism for discovering regularities in the data. We consider a number of examples, including grouping of visual information into surface descriptions and contour descriptions. The main issues that arise concern the configurability of the process and the effects of the propagation of local configurations. We then turn to the complex issue of visual search. Visual search allows the experimenter to establish something of the nature of pre-attentive visual descriptions and how these differ from attentive descriptions. We describe the basic results of visual search experiments in terms of the type of grouping involved. Finally, we consider the hypothesis that dynamic grouping is signalled by neuronal synchrony. PMID- 11115759 TI - Timing of human cortical functions during cognition: role of MEG. AB - Understanding of sensory and cognitive brain processes requires information about activation timing within and between different brain sites. Such data can be obtained by magnetoencephalography (MEG) that tracks cortical activation sequences with a millisecond temporal accuracy. MEG is gaining a well-established role in human neuroscience, complementing with its excellent temporal resolution the spatially more focused brain imaging methods. As examples of MEG's role in cognitive neuroscience, we discuss time windows related to cortical processing of sensory and multisensory stimuli, effects of the subject's own voice on the activity of their auditory cortex, timing of brain activation in reading, and cortical dynamics of the human mirror-neuron system activated when the subject views another person's movements. PMID- 11115760 TI - Electrophysiology reveals semantic memory use in language comprehension. AB - The physical energy that we refer to as a word, whether in isolation or embedded in sentences, takes its meaning from the knowledge stored in our brains through a lifetime of experience. Much empirical evidence indicates that, although this knowledge can be used fairly flexibly, it is functionally organized in 'semantic memory' along a number of dimensions, including similarity and association. Here, we review recent findings using an electrophysiological brain component, the N400, that reveal the nature and timing of semantic memory use during language comprehension. These findings show that the organization of semantic memory has an inherent impact on sentence processing. The left hemisphere, in particular, seems to capitalize on the organization of semantic memory to pre-activate the meaning of forthcoming words, even if this strategy fails at times. In addition, these electrophysiological results support a view of memory in which world knowledge is distributed across multiple, plastic-yet-structured, largely modality-specific processing areas, and in which meaning is an emergent, temporally extended process, influenced by experience, context, and the nature of the brain itself. PMID- 11115761 TI - Extending the classical view of representation. AB - Representation has always been a central part of models in cognitive science, but this idea has come under attack. Researchers advocating the alternative approaches of perceptual symbol systems, situated action, embodied cognition, and dynamical systems have argued against central assumptions of the classical representational approach to mind. We review the core assumptions of the representational view and these four suggested alternatives. We argue that representation should remain a core part of cognitive science, but that the insights from these alternative approaches must be incorporated into models of cognitive processing. PMID- 11115762 TI - Is 'pre-eclampsia' simply a response to the side effects of a placental tachykinin? PMID- 11115763 TI - Developmental changes of the oestrogen receptor-alpha and -beta mRNAs in the female reproductive organ of the rat--an analysis by in situ hybridization. AB - This study employed an in situ hybridization technique to compare the cellular expression of oestrogen receptor (ER) subtypes, ER alpha and beta, in the female reproductive organ of the rat during prenatal and postnatal periods. Diffuse signals of ER alpha and beta mRNAs were co-expressed in the foetal ovary; they were weak and inconsistent before onset of gonadal differentiation, but increased in intensity with age. ER beta mRNA signals in the ovary sharply increased in intensity to adult levels by postnatal days 6-7, whereas those of ER alpha mRNA remained unchanged after birth. ER alpha was the sole subtype expressed during the prenatal period from the oviduct to the vagina, being localized mainly to the sub-epithelial stromal cells, and remained predominant thereafter. Signals for ER alpha mRNA in the epithelia were confined to the oviduct during prenatal and early postnatal periods; those in uterine and vaginal epithelia first appeared by postnatal days 4-5 and 6 respectively. Expressions of ER beta mRNA in the reproductive tract were absent during the prenatal period, and were weakly expressed during the postnatal period. Thus, oestrogen action in the developing ovary may be co-mediated by both ER alpha and beta, whereas ER alpha may be the primary mediator in the differentiation and growth of the female reproductive tract. PMID- 11115764 TI - Expression and localisation of vascular endothelial growth factor and basic fibroblast growth factor during the final growth of bovine ovarian follicles. AB - Locally produced growth factors may have important modulatory roles in final ovarian follicular growth. The aim of this study was to investigate the possible participation of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF2) in bovine follicles during final growth. Ovaries were collected from a slaughterhouse within 10-20 min after exsanguination. A classification of follicles into five groups (<0.5; >0.5-5; >5-20; >20-180; >180 ng/ml) was performed according to the follicular fluid (FF) oestradiol-17 beta content. For a better characterisation of classes the mRNA expressions of FSH receptor, LH receptor and aromatase cytochrome P450 in theca interna (TI) and granulosa cells (GC) were determined. Analysis of VEGF transcript by RT-PCR showed that GC and theca cells express predominantly the smallest isoforms (VEGF(121) and VEGF(165)). VEGF mRNA expression in both tissues (TI and GC) and VEGF protein concentration in total follicle tissue increased significantly (and correlated) with developmental stages of follicle growth. The expression of mRNA for VEGF receptor (VEGFR)-1 and VEGFR-2 was very weak in GC, without any regulatory change during final follicle growth. In contrast, TI showed strong expression of mRNA for both receptors in all follicle classes examined. VEGF protein concentrations in FF increased significantly and continuously to maximum levels in preovulatory follicles. As shown by immunohistochemistry, VEGF protein was clearly localised in TI and GC of preovulatory follicles. FGF2 and FGF receptor (FGFR) mRNA expression in TI increased significantly during final growth of follicles. In contrast, the FGF2 and FGFR mRNA expression in GC was very weak and without any regulatory change during follicle growth. Histological observation revealed that FGF2 protein was localised in theca tissue (cytoplasm of endothelial cells and pericytes) but not in GC. Our results suggest that VEGF and FGF families are involved in the proliferation of capillaries that accompanies the selection of the preovulatory follicle resulting in an increased supply of nutrients and precursors, and therefore supporting the growth of the dominant follicle. PMID- 11115765 TI - Autocrine signals promote osteoblast survival in culture. AB - We have studied the survival requirements of osteoblasts to test the hypothesis that osteoblasts undergo programmed cell death (PCD) or apoptosis unless they are continuously signalled by other cells not to do so. Osteoblasts survived for 6 days in culture at high cell density in the absence of other cell types, serum or exogenous proteins, but they died with the morphological features of apoptosis in these conditions at low cell density. Osteoblast survival was enhanced during the first 2 days of culture by the addition of the sulphydryl compound, cysteine to the culture medium which was converted intracellularly to the antioxidant glutathione. Catalase, an enzyme decomposing hydrogen peroxide, also protected the cells, whereas superoxide dismutase had no effect. Therefore, osteoblasts in culture are sensitive to toxic compounds derived from molecular oxygen, i.e. hydroxyl radicals or hydrogen peroxide spontaneously generated in CMRL medium containing ascorbate and ferrous ions. Conditioned medium from high density cultures prevented osteoblast apoptosis in low density cultures, as long as antioxidants were also present. The enhancing effect of conditioned medium on osteoblast survival was prevented by neutralizing antibodies to insulin-like growth factor-I (IGF-I) and IGF-II but not by antibodies to either platelet derived growth factor (PDGF) or basic fibroblast growth factor (bFGF). These results suggest that in addition to regulating cell growth and differentiation, IGF-I and IGF-II also function as survival factors for osteoblasts. Our data also indicate that antioxidants are required for osteoblast survival and that they enhance growth factor mediated osteoblast survival. PMID- 11115766 TI - The Dstpk61 locus of Drosophila produces multiple transcripts and protein isoforms, suggesting it is involved in multiple signalling pathways. AB - The Drosophila gene Dstpk61 encodes a serine threonine protein kinase homologous to human phosphoinositide-dependent protein kinase (PDK1), and also has homologues in S. cerevisiae, S. pombe, C. elegans, A. thaliana, mouse, and sheep. Where its function has been investigated, this kinase is thought to be involved in regulating cell growth and survival in response to extracellular signals such as insulin and growth factors. In Drosophila it produces multiple transcripts, some of which appear to be sex-specific. In addition to the five Dstpk61 cDNAs we have described previously we report the existence of a further 18 expressed sequence tag (EST) cDNAs, three of which we have fully sequenced. We conclude that Dstpk61 is a complex locus that utilises a combination of alternative promoters, alternative splice sites and alternative polyadenylation sites to produce a vast array of different transcripts. These cDNAs encode at least four different DSTPK61 protein isoforms with variant N-termini. In this paper, we discuss the possible functions of the distinct Dstpk61 transcripts and how they might be differentially regulated. We also discuss the roles that DSTPK61 protein isoforms might play in relation to the protein domains they contain and their potential targets in the cell. Finally, we report the putative structure of the human PDK1 gene based on computer comparisons of available mRNA and genomic sequences. The value of using sequence data from other species for experimental design in mammalian systems is discussed. PMID- 11115767 TI - Characterisation of the IGF system in a primary adult human skeletal muscle cell model, and comparison of the effects of insulin and IGF-I on protein metabolism. AB - In an attempt to address the complex and clinically challenging question of the causes of muscle wasting in patients with cachexia, we have developed a primary adult human skeletal muscle cell model. The cultured cells were characterised by immunocytochemistry using antibodies to the myofibrillar protein constituents desmin and titin. Myotube formation was confirmed biochemically by a fourfold increase in the activity of the muscle-specific enzyme creatinine kinase, and myoblast withdrawal from the cell cycle, which is essential for terminal differentiation, was associated with progressive retinoblastoma protein dephosphorylation. Having successfully confirmed the phenotype of these adult human muscle cells, we assessed their interaction with the insulin-like growth factor (IGF) system. IGF-I is known to stimulate myoblast survival, proliferation and differentiation in cell lines, and, like insulin, is a potent anabolic agent in the regulation of protein metabolism. We have shown that IGF-I stimulated both replication and differentiation of myoblasts, whilst fibroblast growth factor-2 stimulated replication but inhibited differentiation. Examining the IGF system during the process of terminal differentiation, we found that both myoblasts and myotubes expressed insulin, IGF-I and insulin-IGF-I hybrid receptors, with the levels of all three receptor types increasing on differentiation. The cells also produced a wide range of IGF binding proteins (IGFBPs) including IGFBP-2, IGFBP-4 and abundant IGFBP-3, which has not been shown to be produced by any other skeletal muscle cell line examined to date. Both insulin and IGF-I had anabolic effects on myotube protein metabolism at physiological concentrations. Insulin was more potent than IGF-I: use of the IGF analogue long R(3)IGF-I demonstrated that the effects of exogenous IGF-I on protein metabolism were not affected by the high levels of endogenous IGFBP production. In summary, we have developed and characterised a clinically relevant in vitro model with which to address the aetiology of muscle wasting associated with chronic catabolic conditions, and we anticipate that future work will enable the development of novel, effective therapeutic interventions. PMID- 11115768 TI - Myostatin and insulin-like growth factor-I and -II expression in the muscle of rats exposed to the microgravity environment of the NeuroLab space shuttle flight. AB - The mechanism of the loss of skeletal muscle mass that occurs during spaceflight is not well understood. Myostatin has been proposed as a negative modulator of muscle mass, and IGF-I and IGF-II are known positive regulators of muscle differentiation and growth. We investigated whether muscle loss associated with spaceflight is accompanied by increased levels of myostatin and a reduction in IGF-I and -II levels in the muscle, and whether these changes correlate with an increase in muscle proteolysis and apoptosis. Twelve male adult rats sent on the 17-day NASA STS-90 NeuroLab space flight were divided upon return to earth into two groups, and killed either 1 day later (R1) or after 13 days of acclimatization (R13). Ground-based control rats were maintained for the same periods in either vivarium (R3 and R15, respectively), or flight-simulated cages (R5 and R17, respectively). RNA and protein were isolated from the tibialis anterior, biceps femoris, quadriceps, and gastrocnemius muscles. Myostatin, IGF I, IGF-II and proteasome 2c mRNA concentrations were determined by reverse transcription/PCR; myostatin and ubiquitin mRNA were also measured by Northern blot analysis; myostatin protein was estimated by immunohistochemistry; the apoptotic index and the release of 3-methylhistidine were determined respectively by the TUNEL assay and by HPLC. Muscle weights were 19-24% lower in the R1 rats compared with the control R3 and R5 rats, but were not significantly different after the recovery period. The myostatin/beta-actin mRNA ratios (means+/-s.e.m. ) were higher in the muscles of the R1 rats compared with the control R5 rats: 5.0 fold in tibialis (5.35 +/- 1.85 vs 1.07 +/- 0.26), 3.0-fold in biceps (2.46+/ 0.70 vs 0.81 +/- 0.04), 1.9-fold in quadriceps (7.84 +/- 1.73 vs 4.08 +/- 0.52), and 2.2-fold in gastrocnemius (0.99 +/- 0.35 vs 0.44 +/- 0.17). These values also normalized upon acclimatization. Our antibody against a myostatin peptide was validated by detection of the recombinant human myostatin protein on Western blots, which also showed that myostatin immunostaining was increased in muscle sections from R1 rats, compared with control R3 rats, and normalized upon acclimatization. In contrast, IGF-II mRNA concentrations in the muscles from R1 rats were 64-89% lower than those in R3 animals. With the exception of the gastrocnemius, IGF-II was also decreased in R5 animals maintained in flight simulated cages, and normalized upon acclimatization. The intramuscular IGF-I mRNA levels were not significantly different between the spaceflight rats and the controls. No increase was found in the proteolysis markers 3-methyl histidine, ubiquitin mRNA, and proteasome 2C mRNA. In conclusion, the loss of skeletal muscle mass that occurs during spaceflight is associated with increased myostatin mRNA and protein levels in the skeletal muscle, and a decrease in IGF-II mRNA levels. These alterations are normalized upon restoration of normal gravity and caging conditions. These data suggest that reciprocal changes in the expression of myostatin and IGF-II may contribute to the multifactorial pathophysiology of muscle atrophy that occurs during spaceflight. PMID- 11115769 TI - Maternal nutrition alters the expression of insulin-like growth factors in fetal sheep liver and skeletal muscle. AB - We investigated the influence of maternal dietary restriction between days 28 and 80 of gestation followed by re-feeding to the intake of well-fed ewes up to 140 days of gestation (term is 147 days) in sheep, on expression of mRNA for insulin like growth factor (IGF)-I, IGF-II and growth hormone receptor (GHR) in fetal liver and skeletal muscle. Singleton bearing ewes either consumed 3.2-3.8 MJ/day of metabolisable energy (ME) (i.e. nutrient restricted - approximately 60% of ME requirements, taking into account requirements for both ewe maintenance and growth of the conceptus) or 8.7-9.9 MJ/day (i.e. well fed - approximately 150% of ME requirements) between days 28 and 80 of gestation. All ewes were then well fed (150% of ME requirements) up to day 140 of gestation and consumed 8-10.9 MJ/day. At days 80 and 140 of gestation, five ewes were sampled from each group and fetal tissues taken. There was no difference in fetal body weight or liver weights between groups at either sampling date, or skeletal muscle (quadriceps) weight at 140 days. IGF-I mRNA abundance was lower in livers of nutrient-restricted fetuses at day 80 of gestation (nutrient restricted 2.35; well fed 3.70 arbitrary units), but was higher than well-fed fetuses at day 140 of gestation, after 60 days of re feeding (restricted/re-fed 4.27; well fed 2.83;s.e.d. 0.98 arbitrary units, P=0.061 for dietxage interaction). IGF-II mRNA abundance was consistently higher in livers of nutrient-restricted fetuses (80 days: nutrient restricted 7.78; well fed 5.91; 140 days: restricted/re-fed 7.23; well fed 6.01;s.e.d. 1.09 arbitrary units, P=0.061 for diet). Nutrient restriction had no effect on hepatic GHR mRNA abundance, but re-feeding of previously nutrient-restricted fetuses increased GHR mRNA compared with continuously well-fed fetuses (80 days: nutrient restricted 70.6; well fed 75.1; 140 days: restricted/re-fed 115.7; well fed 89.4;s.e.d. 10.13 arbitrary units, P=0.047 for dietxage interaction). In fetal skeletal muscle, IGF-I mRNA abundance was not influenced by maternal nutrition and decreased with gestation age (P<0.01). IGF-II mRNA abundance was higher in skeletal muscle of nutrient-restricted fetuses compared with well-fed fetuses at day 80 of gestation (nutrient restricted 16.72; well fed 10.53 arbitrary units), but was lower than well-fed fetuses after 60 days of re-feeding (restricted/re fed 7.77; well fed 13.72;s.e.d. 1.98 arbitrary units, P<0.001 for dietxage interaction). There was no effect of maternal nutrition or gestation age on fetal skeletal muscle GHR expression. In conclusion, maternal nutrient restriction in early to mid gestation with re-feeding thereafter results in alterations in hepatic and skeletal muscle expression of IGF-I, IGF-II and/or GHR in the fetus which may subsequently relate to altered organ and tissue function. PMID- 11115770 TI - Maternal thyroid status regulates the expression of neuronal and astrocytic cytoskeletal proteins in the fetal brain. AB - Maternal thyroid hormone (TH) crosses the placenta and is postulated to regulate fetal brain development. However, TH-dependent stages of fetal brain development remain to be characterised. We have therefore compared the levels of several neuronal and glial cytoskeletal proteins in fetal brains from normal (N) and partially thyroidectomised (TX) rat dams by immunoblotting. Pregnancies were studied both before and after the onset of fetal TH secretion, which occurs at 17.5 days gestation (dg) in the rat. Maternal hypothyroidism disrupted fetal growth, so that fetal body and brain weights were reduced near term. Vimentin expression was unaffected, however, indicating normal acquisition of neuronal and glial precursor cells. Fetal brain levels of glial fibrillary acidic protein (GFAP) were reduced at 21 dg, suggesting delayed astrocytic differentiation, although regression analysis demonstrated appropriate GFAP levels for brain weight. Levels of alpha-internexin, the earliest neurofilament protein expressed in fetal brain were reduced at 16 dg in TX dams, but increased at 21 dg. The ontogeny of neurofilament-L was also perturbed in these pregnancies, with deficient levels apparent at both 16 and 21 dg. These effects on neuronal cytoskeletal proteins were unrelated to fetal brain growth retardation. These findings confirm that maternal hypothyroidism disrupts early fetal brain development. Early disturbances in neuronal differentiation are not corrected by the onset of fetal TH secretion. Such disturbances may contribute to the neurological damage observed in children born to hypothyroxinaemic mothers. PMID- 11115771 TI - Effects of estrogens and androgens on erythrocyte antioxidant superoxide dismutase, catalase and glutathione peroxidase activities during the menstrual cycle. AB - The effects of physiological changes in estrogens and androgens on the erythrocyte antioxidant superoxide dismutase, catalase and glutathione peroxidase enzyme activities during the menstrual cycle were investigated in healthy eumenorrheic women. Blood samples were taken on alternate days from twelve normally cyclic women (age range: 20 to 27 years; mean age: 24.1 years) from the first day of one menstrual cycle until the first day of the subsequent one. Plasma was analyzed for FSH, LH, estradiol, progesterone, testosterone, free testosterone and androstenedione concentrations. Erythrocyte superoxide dismutase, catalase and glutathione peroxidase activities were evaluated on the same days and cycle length was standardized on the basis of the preovulatory estradiol peak. Significant cyclic phase-related changes were observed in glutathione peroxidase (P<0.05), with higher glutathione peroxidase activity levels from the late follicular to the early luteal phase compared with those found in the early follicular phase (P<0.001 and P<0.002 respectively). A significant positive correlation was observed between mean estradiol and glutathione peroxidase cycle-related variations (r=0.80, P<0.001), whereas no significant cycle phase-dependent changes were seen in superoxide dismutase and catalase. No effect of progesterone and androgens on the erythrocyte antioxidant enzyme system was documented. The findings indicate that physiological ovarian estradiol production during the menstrual cycle may have an important role in regulating erythrocyte glutathione peroxidase activity. PMID- 11115772 TI - Replenishment of LH stores of gonadotrophs in relation to gene expression, synthesis and secretion of LH after the preovulatory phase of the sheep oestrous cycle. AB - The pattern of replenishment of LH secretory granule stores in sheep pituitary gonadotrophs, after an induced LH surge, was determined by immunogold localisation at the ultrastructural level by electron microscopy. Twenty-four Welsh Mountain ewes were initially synchronised with progestagen devices for 14 days before luteolysis was induced by a prostaglandin F(2 alpha) analogue, cloprostenol. A further 24 h later, a preovulatory LH surge was induced by intravenous injection of a GnRH agonist, buserelin. Animals were divided into four groups (n=6) and blood sampled at 2 h intervals from 4 h prior, to 18 h after, buserelin administration and then at infrequent intervals (1 to 8 h) thereafter until death. Pulse profiles of LH were also obtained by an additional collection of blood samples within a 6 h window directly preceding death. Groups of animals were killed at 24, 48, 72 or 96 h after buserelin treatment. Pituitaries were dissected and processed for transmission electron microscopy and frozen for later molecular biological analysis. A characteristic preovulatory surge of LH was observed in all animals. The cytoplasm of gonadotrophs, in animals killed 24 h after buserelin treatment, was completely empty of secretory granules. This was associated with diminutive pituitary LH content, low pituitary GnRH binding levels and an almost complete absence (one pulse in one animal) of LH pulsatile secretion. Despite the lack of apparent secretory activity, clusters of exposed LH beta label present within the cytoplasm at this time and constant LHbeta mRNA expression levels irrespective of tissue collection time, suggest that the cell is actively synthesising LHbeta. The formation of sparse numbers of small LH beta immuno-labelled electron-dense secretory granules was apparent at 48 h after buserelin treatment, and replenishment of LH beta immuno-labelled granule stores continued until total granule numbers had increased two-fold (P<0.01) by 96 h post-treatment. Affiliated with granule replenishment was a significant increase in pituitary LH content (P<0.01), pituitary GnRH binding levels (P<0.01) and the restoration of LH pulsatile secretion. Despite the replenishment of granule stores with time, cytoplasmic area did not vary. These results suggest that restoration of pulsatile LH secretion after a preovulatory LH surge is related to replenishment of LH beta secretory granule stores and an increase in GnRH binding levels. PMID- 11115773 TI - Human granulosa cells are a site of sulphatase activity and are able to utilize dehydroepiandrosterone sulphate as a precursor for oestradiol production. AB - Dehydroepiandrosterone sulphate (DHEAS) is the most abundant androgen in the circulation and in ovarian follicular fluid. A steroid sulphatase accepting DHEAS as a substrate has been identified in the follicle, but the cellular location has not been determined. As DHEAS is also a potential source of oestrogen for endocrine-dependent tumours, a potent steroid sulphatase inhibitor oestrone-3-O sulphamate (EMATE) has been developed which inhibits this activity in rat liver and mammary tumour. The aim of this study was to investigate human granulosa cells as a site of steroid sulphatase activity, to determine whether DHEAS can be utilized as a precursor for oestrogen synthesis and to investigate the inhibitory capacity of EMATE in these cells. Conversion of DHEAS to DHEA was assessed in luteinized granulosa cells by tritiated steroid assay following incubation with or without LH or insulin and steroid accumulation in the medium measured by RIA. The effects of EMATE were assessed by addition of a range of doses during the measurement of conversion of DHEAS to DHEA. Cells from three sizes of small follicles from an unstimulated ovary were also assessed for their ability to produce oestradiol from DHEAS. Sulphatase enzyme activity was present in all cells; the mean conversion of tritiated DHEAS to DHEA was 50% (range 4-65%). LH and EMATE inhibited and insulin stimulated this activity. Addition of DHEAS to granulosa cells caused a dose-dependent increase in oestradiol and androstenedione production with no change in progesterone concentration. LH increased the accumulation of oestradiol in the medium. DHEAS also stimulated oestradiol production by granulosa cells from small follicles. This is the first demonstration that granulosa cells are a site of sulphatase activity and that DHEAS can be utilized as a substrate for androstenedione and oestrogen production. This may be of physiological importance for both normal folliculogenesis and oestrogen-dependent tumour growth. PMID- 11115774 TI - Capacitative calcium entry is involved in steroidogenesis in bovine adrenocortical fasciculata cells. AB - Capacitative Ca(2+) entry into bovine adrenocortical fasciculata cells was investigated by using the mobilization of intracellular Ca(2+) concentration ([Ca(2+)](i)) and Ca(2+)-induced steroidogenesis as the indicators. Bovine adrenocortical fasciculata cells on a glass coverslip were loaded with fura-2. The [Ca(2+)](i) mobilization was detected by a change of fura-2 fluorescence intensity. In the intracellular Ca(2+) store depleted cells, the addition of Ca(2+) to the incubation medium elicited a marked and sustained increase in [Ca(2+)](i). In the intracellular Ca(2+) store non-depleted cells, the addition of thapsigargin, an endoplasmic reticulum Ca(2+)-ATPase inhibitor, in the absence of extracellular Ca(2+), induced a slight and transient increase in [Ca(2+)](i), but an extensive and sustained increase in [Ca(2+)](i) was obtained by adding Ca(2+) to the incubation medium after the thapsigargin treatment. The sustained increase induced by thapsigargin was not inhibited by nifedipine, but was inhibited by Zn(2+) and Cd(2+) in a concentration-dependent manner. The effect of Zn(2+) was more potent than that of Cd(2+). Thapsigargin stimulated steroidogenesis in the presence of extracellular Ca(2+). The steroidogenic effect of thapsigargin was inhibited by Zn(2+) and Cd(2+) but not by nifedipine. These results suggest that there is, in bovine adrenocortical fasciculata cells, a steroidogenesis-linked Ca(2+) entry process other than that involving voltage operated Ca(2+) channels and that the process might be capacitative Ca(2+) entry. PMID- 11115775 TI - Homologous and heterologous down-regulation of leptin receptor messenger ribonucleic acid in rat adrenal gland. AB - Leptin, the adipocyte-produced hormone that plays a key role in body weight homeostasis, has recently been found to be involved in the regulation of the hypothalamic-pituitary-adrenal axis. Moreover, reciprocal interactions between leptin and glucocorticoids have been described. In the present communication, two different strategies were undertaken to explore the mode of action of leptin in the direct control of rat adrenal function. First, a synthetic peptide approach demonstrated that the inhibitory effect of leptin on basal and ACTH-stimulated corticosterone secretion in vitro is, at least partially, mapped to a domain of the native protein between amino acids 116 and 130, i.e. an area of the molecule also relevant in terms of regulation of food intake and endocrine control. Secondly, semi-quantitative RT-PCR analysis indicated a complex pattern of adrenal leptin receptor (Ob-R) mRNA expression, with predominant expression of the Ob-Ra and Ob-Rb isoforms, as well as moderate levels of the Ob-Rc and Ob-Rf variants, whereas negligible signals for the Ob-Re isoform were detected. Interestingly, such an expression pattern appeared hormonally regulated as exposure to human recombinant leptin (10(-7 )M) or ACTH (10(-7 )M) significantly decreased Ob-R isoform mRNA expression. Indeed, dose-dependent ligand-induced Ob Ra and Ob-Rb mRNA down-regulation was further confirmed by adrenal stimulation with increasing concentrations (10(-9)-10(-5 )M) of the active leptin fragment, leptin 116-130 amide. Overall, our results provide evidence for a novel regulatory step at the level of Ob-R mRNA expression in the interplay between ACTH and leptin for the tuning of rat adrenal corticosterone secretion. Furthermore, our data showing down-regulation of Ob-R mRNA expression by its cognate ligand may well be relevant to leptin physiology and its alteration in various disease states. PMID- 11115776 TI - Different transporters for tri-iodothyronine (T(3)) and thyroxine (T(4)) in the human choriocarcinoma cell line, JAR. AB - We investigated transport systems for tri-iodothyronine (T(3)) and thyroxine (T(4)) in the human choriocarcinoma cell line, JAR, using a range of structurally similar compounds to determine whether these thyroid hormones are transported by common or different mechanisms. Saturable T(3) but not saturable T(4) uptake was inhibited by a wide range of aromatic compounds (nitrendipine, nifedipine, verapamil, meclofenamic acid, mefenamic acid, diazepam, phenytoin). Nitrendipine and diazepam were the most effective inhibitors of saturable thyroid hormone uptake. Nitrendipine decreased the K(m) for T(4) uptake from a control value of around 500 nM to around 300 nM (n=6). In contrast, the K(m) for T(3) uptake was increased from a control value of around 300 nM to around 750 nM (n=4). Diazepam had similar effects. This divergent shift in affinity for the uptake of T(3) and T(4) suggested that separate uptake systems exist for these two thyroid hormones. This provides evidence for at least two transporters mediating uptake of T(3) and T(4) in JAR cells: a specific T(4) transporter that does not interact with T(3) or structurally similar compounds; and a shared iodothyronine transporter that interacts with T(3), T(4), nitrendipine and diazepam. PMID- 11115777 TI - Very strong correlation between dominant negative activities of mutant thyroid hormone receptors and their binding avidity for corepressor SMRT. AB - The syndrome of resistance to thyroid hormone (RTH) is an inherited disorder involving a mutation of the thyroid hormone receptor (TR) gene. Mutant (m) TR inhibits wild-type (wt) TR functions in a dominant negative manner, and this dominant negative effect (DNE) is a crucial factor in RTH pathogenesis. The molecular mechanism of the DNE is still unclear, although several possibilities (including competition between wt- and mTRs at the T(3) response element (TRE), sequestration of TR-associated protein(s) and titration out of functional TR) have been considered. Here we report that the DNE of mTRs is strongly correlated with their binding avidity for the retinoid X receptor (RXR), and especially for corepressor SMRT (silencing mediator for retinoid and thyroid hormone receptor), but not for the nuclear receptor corepressor, NCoR. The DNE of six natural TRs and four artificially constructed mTRs was assayed using a TR reporter gene containing TRE-DR4 (DR=direct repeat), TRE-pal (pal=palindrome) or TRE-lap (lap=inverted palindrome) in CV1 cells treated with 10 nM T(3). Of the mTRs examined, F451X (with a carboxy-terminal 11-amino-acid truncation) identified in a patient with RTH exhibited the strongest DNE on all TREs. The binding affinities between mTRs and corepressors SMRT or NCoR were quantified using a two hybrid interference assay system consisting of VP16-TR(LBD) (LBD=ligand binding domain) and Gal4(DBD)-SMRT (DBD=DNA binding domain), or Gal4(DBD)-NCoR respectively, together with the Gal4 reporter gene. In this assay, VP16-TR(LBD) and Gal4(DBD)-SMRT (or Gal4 (DBD)-NCoR) interact with each other and trans activate the Gal4 reporter gene. When an equal amount of mTR is coexpressed, it reduces the transcriptional activity of the reporter gene, depending on its binding avidity for a corepressor. A very strong correlation was observed between the SMRT-binding activity and the potency of the DNE among six natural mTRs and also among all mTRs, including four artificially constructed ones. The relationship between NCoR and DNE, however, was not significant. When we assayed the binding avidity of mTRs for RXR by using a two-hybrid assay system consisting of Gal4(DBD)-RXR(LBD) and VP16-TR(LBD), a significant correlation between DNE and binding avidity for the RXR was also observed. These results suggest that a corepressor plays an important role in DNE pathogenesis. PMID- 11115778 TI - Effects of proinflammatory cytokines on anterior pituitary 5'-deiodinase type I and type II. AB - Cytokines are locally produced in the anterior pituitary and act through para /autocrine mechanisms to modulate cell growth and hormone production. 5' Deiodinases type I (D1) and type II (D2) are also expressed in the anterior pituitary and play an integrative role in the regulation of hormone production and pituitary feedback. D1 activity is known to be regulated by proinflammatory cytokines in liver and thyroid. Therefore, we examined the effects of IL-1 beta, IL-6 and TNF alpha on 5'-deiodinase activities in reaggregates of rat anterior pituitaries and rat somatomammotroph GH3 cells cultured alone, or in a bicameral culture system together with the murine folliculo-stellate (FS) cell line TtT/Gf. In reaggregate cultures of rat anterior pituitaries IL-1 beta stimulated D1 and D2 dose-dependently and D2 activity was increased by TNF alpha. When GH3 cells were cocultured with TtT/Gf cells, D2 activities were 2.3-fold lower than in GH3 cells cultured alone. TNF alpha (50 ng/ml) and IL-6 (100 U/ml) stimulated D2 in GH3 cells when the cells were cultured alone and treated with these cytokines for 24 h. When TtT/Gf cells in the coculture model were treated with IL-1 beta, TNF alpha and IL-6, no effect on D1 or D2 activities in GH3 cells was observed. In male, adult rats a single LPS injection (i.p.) stimulated D2 and D1 activities in the anterior pituitary, and decreased liver D1 activities and serum TSH levels. In vitro, LPS stimulation of the coculture model of GH3 and FS cells also increased D1 activity. Electrophoretic mobility shift assays (EMSAs) revealed that IL-1 beta and TNF alpha activate the transcription factor NF kappa B in reaggregates of rat anterior pituitaries and in TtT/Gf cells cultured alone or cocultured with GH3 cells. Taken together, these findings imply that in anterior pituitary cells 5'-deiodinase activities are stimulated by locally produced cytokines in a para-/autocrine manner but cell types other than FS cells seem to mediate some of the effects. PMID- 11115779 TI - Chronic hypoxia upregulates the expression and function of AT(1) receptor in rat carotid body. AB - In the present study, the effects of chronic hypoxia on the expression and localization of angiotensin II (Ang II) receptors are investigated by semi quantitative reverse transcription-polymerase chain reaction (RT-PCR) and by immunohistochemistry. The effect of chronic hypoxia on the carotid body chemoreceptor activity was also examined by in vitro electrophysiology. Results from RT-PCR revealed that chronic hypoxia exhibited differential effects on the gene expression of Ang II receptors, namely AT(1) and AT(2), in the carotid body. The mRNA expression for subtypes of the AT(1) receptor, AT(1a) and AT(1b), was significantly increased in the carotid body with chronic hypoxia. To further investigate the localization of the AT(1) receptor, an immunohistochemical study was performed. The results showed that AT(1) receptor immunoreactivity was found in lobules of glomus cells in the carotid body and the immunoreactivity was more intense in chronic hypoxia than in normoxic controls. In vitro electrophysiological studies consistently demonstrated that chronic hypoxia enhanced the AT(1) receptor-mediated excitation of carotid body chemoreceptor activity. These data suggest that chronic hypoxia upregulates the transcriptional and post-transcriptional expression of AT(1) receptors in the rat carotid body. The upregulation of the expression also enhances AT(1) receptor-mediated excitation of the carotid body afferent activity. This might be important in the modulation of cardiorespiratory functions as well as fluid and electrolyte homeostasis during chronic hypoxia. PMID- 11115780 TI - Possible interactions between angiotensin II and insulin: effects on glucose and lipid metabolism in vivo and in vitro. AB - Angiotensin II (ANGII) increases insulin sensitivity in diabetic and non-diabetic subjects, even at subpressor doses, and because there is 'crosstalk' between ANGII and insulin-signaling pathways the underlying mechanism may not be due solely to changes in regional blood flow. A series of experimental studies was undertaken to evaluate the effects of ANGII on glucose and lipid metabolism in vivo and in vitro. Groups of fructose-fed, insulin-resistant Sprague-Dawley (SD) rats were pre-treated with 0.3 mg/kg per day of the AT(1)-receptor antagonist L 158 809 (n=16), or vehicle (n=16), by oral gavage. This was prior to an oral glucose tolerance test (day 5) and measurement of the effects of ANGII infusion (20 ng/kg per min i.v. for 3 h) on whole-body insulin sensitivity using the insulin suppression test (day 7). The effect of ANGII infusion on total triglyceride secretion rate (TGSR) was evaluated in normal SD rats pretreated for 7 days with L-158 809 (n=12) or vehicle (n=12). AT(1)- and AT(2)- receptor mRNA expression and [(3)H]2-deoxyglucose uptake were assessed in cultured L6 myoblasts. Short-term treatment with L-158 809 had no effect on glucose tolerance or fasting triglyceride levels in fructose-fed rats. ANGII infusion had no effect on insulin sensitivity in fructose-fed rats pretreated with vehicle (steady-state plasma glucose (SSPG) values 8.1+/-1.6 vs 8. 4+/-0.4 mmol/l), but pretreatment with L-158 809 resulted in ANGII having a modest insulin antagonist effect in this insulin-resistant model (SSPG values 9.6+/-0.3 vs 7.1+/-0.6, P<0.03). ANGII infusion had no significant effect on TGSR (e.g. 24.6+/-1.4 vs 28.4+/-0.9 mg/100 g per h in vehicle-treated animals). RT-PCR analysis showed that L6 cells express both AT(1)- and AT(2)-receptor mRNA. Incubation with ANGII (10(-9) and 10(-8) M) had no significant effect on the dose-response curve for insulin-stimulated [(3)H]2-deoxyglucose uptake. For example, C(I200) values (dose of insulin required to increase glucose uptake by 200%) were 4.5 x 10(-9) M (control) vs 3.9 x 10(-9) M and 6.2 x 10(-9) M, whereas the positive control (glucagon-like peptide-1) increased insulin sensitivity. Thus, ANGII infusion may have a modest insulin antagonist effect on glucose disposal in insulin-resistant fructose-fed rats pretreated with an AT(1)-blocker, but ANGII has no effect on TGSR or in vitro glucose uptake in L6 myoblasts. These findings are relevant to recent clinical discussions about the metabolic effects of ANGII and renin-angiotensin system blockade. PMID- 11115781 TI - Mechanisms of dysregulation of 11 beta-hydroxysteroid dehydrogenase type 1 in obese Zucker rats. AB - Obesity has been associated with alterations in glucocorticoid metabolism in both man and rodents, but the underlying mechanisms remain undefined. We have previously reported tissue-specific alterations in 11 beta-hydroxysteroid dehydrogenase type 1 (11 beta-HSD1) in obese Zucker rats predicting that reactivation of corticosterone is decreased in liver but increased in omental fat. The mechanisms of dysregulation of 11 beta-HSD1 in obesity are not known, and in this study we have investigated the potential role of glucocorticoids and insulin. In one experiment lean and obese Zucker rats were adrenalectomised, and in a second experiment they were sensitised to insulin by treatment with either metformin or rosiglitazone. Adrenalectomy (ADX) of obese animals attenuated weight gain, normalised hepatic 11 beta-HSD1 kinetics by an effect on V(max) (V(max)in sham-operated animals, 6.6+/-1.1 nmol/min per mg in lean vs 3.4+/-0.6 in obese, P<0.01; in ADX animals 5.9+/-1.1 in lean vs 6.9+/-1.8 in obese, NS), and reversed the difference in omental fat 11 beta-HSD1 activity (18.9+/-4.2% in lean ADX vs 8.2+/-2.3 in obese ADX, P=0.03). Both metformin and rosiglitazone improved insulin sensitivity in obese, but not lean animals, and had no effect on 11 beta-HSD1 activity in either liver or fat. However, both treatments normalised adrenal hypertrophy in obese animals (48+/-29 mg in obese vehicle vs 37+/-1.2 in metformin and 38+/-1.8 in rosiglitazone treated, both P<0.01), and rosiglitazone tended to attenuate hypercorticosteronaemia in obese rats. Neither treatment attenuated weight gain; in fact, weight gain was enhanced by rosiglitazone in obese rats. In summary, altered 11 beta-HSD1 activity in obese Zucker rats is reversible following adrenalectomy, but the mechanism is unclear since adrenalectomy also normalises many other metabolic abnormalities. The current study suggests that hyperinsulinaemia is not responsible for tissue-specific dysregulation of 11 beta-HSD1. However, insulin sensitisation did reverse adrenal hypertrophy, suggesting that hyperinsulinaemia may be a key factor contributing to activation of the hypothalamic- pituitary-adrenal (HPA) axis in obesity independently of tissue-specific changes in 11 beta-HSD1. PMID- 11115782 TI - The hexapeptide KP-102 (D-ala-D-beta-Nal-ala-trp-D-phe-lys-NH(2)) stimulates growth hormone release in a cichlid fish (Ooreochromis mossambicus). AB - Abstract Studies in mammals have shown that synthetic Met-enkephalin derivatives, called growth hormone-releasing peptides (GHRPs), stimulate growth hormone (GH) release. The present study was conducted to determine whether the GHRP, KP-102, specifically stimulates GH release in a teleost. Tilapia (Oreochromis mossambicus) were given a single intraperitoneal injection of KP-102 (D-Ala-D beta;-Nal-Ala-Trp-D-Phe-Lys-NH(2)) or bovine GHRH(1-29)-amide or vehicle and blood was sampled at 1, 6 and 12 h after injection. KP-102 was administered at two doses of 1 ng/g and 10 ng/g body weight, whereas GHRH (positive control) was administered at a single dose of 10 ng/g body weight. Plasma levels of tilapia GH and prolactins (tPRL(177) and tPRL(188)) were determined by radioimmunoassay. As expected, GHRH injection significantly (P<0.001) elevated plasma GH levels (ng/ml) in tilapia at 6 h post-injection. KP-102 also significantly elevated GH levels (at the low dose) at 6 (P<0.05) and 12 (P<0.01) hours post-injection. There were no significant effects on plasma PRL(s) levels, although mean levels of both PRLs were elevated at 6 h post-injection. These results show for the first time that GHRPs stimulate GH release in teleosts and suggest that the GHRP receptor and possibly a "Ghrelin-like" ligand are also present in lower vertebrates. PMID- 11115783 TI - Charting the physical activity patterns of contemporary children and adolescents. AB - The impact of physical inactivity on health is well accepted throughout the medical and health service community. However, the case has largely been established through epidemiological studies with adults. Substantial attention has been paid to the activity levels of children and adolescents, largely because of changing lifestyles that have threatened the opportunity to be active and also introduced attractive sedentary alternatives such as playing computer games. The research evidence that children have become less active to the point where it is seriously damaging their current and future health has been difficult to establish. This situation is due to difficulties in establishing sensitive health risk markers, and also with the assessment of the different elements of physical activity which in children and adolescents is a complex profile of social behaviours. Self report of activity is unreliable with young children, and objective measures are required that are cheap and effective with large samples and that are capable of measuring levels, volume and patterns of physical activity. Accelerometry in combination with diaries offers the best current solution for most activity-health relationships, and for informing intervention need and design. PMID- 11115784 TI - Obesity: criteria and classification. AB - Obesity is defined as an excess accumulation of body fat. To measure fat in the body accurately is difficult, and no method is easily available for routine clinical use. Traditionally, overweight and obesity have been evaluated by anthropometric measurement of weight-for-height. More recently, BMI has been used. The normal range is 19-24.9 kg/m2, overweight is 25-29.9 kg/m2, and obesity >/= 30 kg/m2. Not only is the total amount of fat an individual carries important, but also where the fat is distributed in the body. Fat in a central or upper body (android) distribution is most related to health risk. The most accurate way to measure central obesity is by magnetic resonance imaging or computer-assisted tomography scanning, but this approach is too expensive for routine use. Simple anthropometric measurements can be used, such as waist circumference. A waist circumference of greater than 1020 mm in men and 880 mm in women is a risk factor for insulin resistance, diabetes mellitus and cardiovascular disease. There is a clear genetic predisposition for obesity. The genetic contribution to obesity is between 25 and 40 % of the individual differences in BMI. For the overwhelming majority of individuals, the genetic predisposition will not be defined by one gene, but by multiple genes. Eventually, classification of obesity may be done by genetic means, but this approach will require more knowledge. PMID- 11115785 TI - Role of substrate utilization and thermogenesis on body-weight control with particular reference to alcohol. AB - Alcohol (ethanol; EtOH) provides fuel energy to the body (29.7 kJ (7. 1 kcal)/g, 23.4 kJ (5.6 kcal)/ml), as do other macronutrients, but no associated essential nutrients. The thermogenic effect of EtOH (on average 15 % of its metabolizable value) is much greater than that of the main substrates utilized by the body, i.e. fat and carbohydrates (CHO), suggesting a lower net efficiency of energy utilization for EtOH than for fat and CHO. EtOH cannot be stored in the body and is toxic, so that there is an obligatory continuous oxidation of EtOH and it becomes the priority fuel to be metabolized. In contrast to CHO, its rate of oxidation does not depend on the dose ingested. As with CHO intake, it engenders a shift in postprandial substrate utilization (decrease in fat oxidation), but by a non-insulin-mediated mechanism. A limited amount of EtOH can be converted to fatty acids by hepatic de novo lipogenesis (as occurs with high levels of CHO feeding) from acetate production, which inhibits lipolysis in peripheral tissues. There is no evidence that EtOH consumed under normoenergetic conditions (i.e. isoenergetically replacing CHO or fat) leads to greater body fat storage than fat or CHO. However, there is still a lack of experimental studies on the influence of EtOH on the level of spontaneous physical activity in man. This effect may well depend on the dose of EtOH consumed as well as other intrinsic factors. PMID- 11115786 TI - All-trans-retinoic acid and polyriboinosinoic:polyribocytidylic acid cooperate to elevate anti-tetanus immunoglobulin G and immunoglobulin M responses in vitamin A deficient lewis rats and Balb/c mice. AB - Vitamin A (VA) deficiency compromises antibody responses to T-cell-dependent antigens such as tetanus toxoid, but this effect can be reversed through administration of retinol or retinoic acid (RA). To test whether RA and polyriboinosinioc : polyribocytidylic acid (PIC), a known inducer of several forms of interferon (IFN), can cooperate to increase specific immunoglobulin (Ig)G and IgM production during VA deficiency, rats and mice were made VA deficient, immunized with TT and treated with all-trans-RA, PIC or their combination. VA-deficient rats produced low primary and secondary anti-tetanus IgG responses (VA-deficient controls v. VA-sufficient controls P < 0.001), although total IgG was slightly elevated when compared with VA-sufficient control rats. Although RA administered alone elevated antibody production during VA deficiency to control levels, RA combined with PIC synergistically enhanced these responses (RA and PIC group v. all other groups P < 0.0001). In contrast, Balb/c mice maintained on a VA-deficient diet and immunized in a similar fashion showed no impairment in antigen-specific IgG levels, but treatment with a combination of RA and PIC still evoked an additive enhancement in antigen-specific antibody production. Additionally, RA and PIC administration to VA-sufficient mice resulted in elevated antibody responses, suggesting that this combination should be evaluated further for its immuno-stimulatory effects. PMID- 11115787 TI - Vitamin D: a natural inhibitor of multiple sclerosis. AB - Inheriting genetic risk factors for multiple sclerosis (MS) is not sufficient to cause this demyelinating disease of the central nervous system; exposure to environmental risk factors is also required. MS may be preventable if these unidentified environmental factors can be avoided. MS prevalence increases with decreasing solar radiation, suggesting that sunlight may be protective in MS. Since the vitamin D endocrine system is exquisitely responsive to sunlight, and MS prevalence is highest where environmental supplies of vitamin D are lowest, we have proposed that the hormone, 1, 25-dihydroxycholecalciferol (1,25-(OH)2D3), may protect genetically-susceptible individuals from developing MS. Evidence consistent with this hypothesis comes not only from geographic studies, but also genetic and biological studies. Over-representation of the vitamin D receptor gene b allele was found in Japanese MS patients, suggesting it may confer MS susceptibility. Fish oil is an excellent vitamin D source, and diets rich in fish may lower MS prevalence or severity. Vitamin D deficiency afflicts most MS patients, as demonstrated by their low bone mass and high fracture rates. However, the clearest evidence that vitamin D may be a natural inhibitor of MS comes from experiments with experimental autoimmune encephalomyelitis (EAE), a model of MS. Treatment of mice with 1,25-(OH)2D3 completely inhibited EAE induction and progression. The hormone stimulated the synthesis of two anti encephalitogenic cytokines, interleukin 4 and transforming growth factor beta-1, and influenced inflammatory cell trafficking or apoptosis. If vitamin D is a natural inhibitor of MS, providing supplemental vitamin D to individuals who are at risk for MS would be advisable. PMID- 11115788 TI - Cellular and molecular aspects of iron and immune function. AB - Fe plays a critical role in the immune system and defence against infection. However, many aspects of the way in which Fe influences these processes at the molecular and cellular level are unclear. The present review summarizes the role of Fe in lymphocyte activation and proliferation, and discusses how Fe is handled by macrophages. PMID- 11115789 TI - Zinc and the immune system. AB - Zn is an essential trace element for all organisms. In human subjects body growth and development is strictly dependent on Zn. The nervous, reproductive and immune systems are particularly influenced by Zn deficiency, as well as by increased levels of Zn. The relationship between Zn and the immune system is complex, since there are four different types of influence associated with Zn. (1) The dietary intake and the resorption of Zn depends on the composition of the diet and also on age and disease status. (2) Zn is a cofactor in more than 300 enzymes influencing various organ functions having a secondary effect on the immune system. (3) Direct effects of Zn on the production, maturation and function of leucocytes. (4) Zn influences the function of immunostimulants used in the experimental systems. Here we summarize all four types of influence on the immune function. Nutritional aspects of Zn, the physiology of Zn, the influence of Zn on enzymes and cellular functions, direct effects of Zn on leucocytes at the cellular and molecular level, Zn-altered function of immunostimulants and the therapeutic use of Zn will be discussed in detail. PMID- 11115790 TI - The scientific basis of immunonutrition. AB - Substrates with immune-modulating actions have been identified among both macro- and micronutrients. Currently, the modes of action of individual immune modulating substrates, and their effects on clinical outcomes, are being examined. At present, some enteral formulas are available for the clinical setting which are enriched with selected immune-modulating nutrients. The purpose of the present paper is to review the scientific rationale of enteral immunonutrition. The major aspects considered are mucosal barrier structure and function, cellular defence function and local or systemic inflammatory response. It is notable that in critical illness the mucosal barrier and cellular defence are impaired and a reinforcement with enteral immunonutrition is desirable, while local or systemic inflammatory response should be down regulated by nutritional interventions. The results available from clinical trials are conflicting. Meta analyses of recent trials show improvements such as reduced risk of infection, fewer days on a ventilator, and reduced length of intensive care unit and hospital stay. Thus, a grade A recommendation was proclaimed for the clinical use of enteral immune-modulating diets. Improvement in outcome was only seen when critical amounts of the immune-modulating formula were tolerated in patients classified as being malnourished. However, in other patients with severe sepsis, shock and organ failure, no benefit or even disadvantages from immunonutrition were reported. In such severe conditions we hypothesize that systemic inflammation might be undesirably intensified by arginine and unsaturated fatty acids, directly affecting cellular defence and inflammatory response. We therefore recommend that in patients suffering from systemic inflammatory response syndrome great caution should be exercised when immune-enhancing substrates are involved which may aggravate systemic inflammation. PMID- 11115791 TI - Role of breast-feeding in managing malnutrition and infectious disease. AB - Breast-feeding policy tends to be an emotive issue. International agencies recommend exclusive breast-feeding for 4-6 months followed by continued partial breast-feeding into the second year of life in order to promote infant and child health and minimize the damage caused by the malnutrition-infection cycle. To what extent are these recommendations supported by the experimental evidence? Are they a simplification for emotional reasons or public health purposes? Breast feeding is believed to benefit infants because breast milk contains the ideal mix of nutrients for infants, because it contains factors which promote development of the infant's gut and immune system and which prevent pathogen invasion, and because exclusive breast-feeding prevents intake of pathogens in food or water. However, some apparently contradictory evidence exists. First, in environments which are not highly contaminated breast-fed infants tend to growth falter relative to those fed formula. Second, in such environments partial breast feeding is not associated with significantly increased gut damage relative to exclusive breast-feeding, suggesting that active promotion of gut development by breast-feeding is more important than simple avoidance of pathogens from other foods. Third, many immune factors in breast milk are probably present primarily to protect the mother, not the infant. Finally, breast milk itself may contain bacteria or viruses. This problem has come to the fore with the human immunodeficiency virus epidemic, since it is clear breast-feeding is an important mode of mother-to-child transmission. The present review will examine these challenges to the basis of the international infant feeding recommendations and will suggest that the science does actually support the policy. PMID- 11115792 TI - Cow's milk and immune-mediated diabetes. AB - Cow's milk-based infant formulas and cow's milk consumption in childhood have been suggested to promote the development of type 1 diabetes mellitus and other immune-mediated or neurological diseases. Epidemiological studies in man have led to the hypothesis that introduction of cow's milk-based infant formula within the first 3 months of life is associated with increased risk of type 1 diabetes mellitus. Furthermore, in animal models of type 1 diabetes mellitus, cow's milk proteins have been proven to be 'diabetogenic'. However, the issue seems far from being resolved. Several epidemiological studies and, more importantly, the first prospective trials did not show an association between early exposure to cow's milk and type 1 diabetes mellitus. In animal models, cow's milk proteins are modestly and variably diabetogenic, wheat or soybean proteins in the diet cause higher rates of autoimmune diabetes. In both man and rodents there is increasing evidence that the gut-associated immune system plays a major role in disease development, probably because of disturbed oral tolerance mechanisms. Oral tolerance depends on immunological homeostasis and normal maturation of the gut. These factors are influenced by growth factors and cytokines from breast milk, normal bacterial colonization, infections and diet. All these factors have been proposed as risk factors for type 1 diabetes mellitus. Hence, cow's milk proteins may provide mimicry epitopes relevant in autoimmunity, as well as destabilizing oral tolerance mechanisms by biologically active peptides. The concept of dietary regulation of autoimmunity does not apply only to cow's milk protein, but also to other dietary proteins. PMID- 11115793 TI - Micronutrients and host resistance to viral infection. AB - Previous work in our laboratory demonstrated that a virus could undergo rapid mutation in a host deficient in Se, leading to a normally avirulent virus acquiring virulence due to genome changes. Once these mutations occur, even a host with adequate Se-nutriture is susceptible to the newly virulent virus. What influence does the deficiency in Se have on the immune response of the host? Infection with myocarditic strains of coxsackievirus induces an inflammatory response in the cardiac tissue. It is this immune response that induces the heart damage, rather than direct viral effects on the heart tissue. Chemokines are chemo-attractant molecules that are secreted during an infection in order to attract immune cells to the site of the injury, and have been found to be important for the development of coxsackievirus-induced myocarditis. We found that a deficiency in Se influences the expression of mRNA for the chemokine monocyte chemo-attractant protein-1, which may have implications for the development of myocarditis in the Se-deficient host. Expression of mRNA for interferon-gamma was also greatly decreased in the Se-deficient animal. Thus, a deficiency in Se can have profound effects on the host as well as on the virus itself. How the alteration of the immune response of the Se-deficient animal affects the development of the virulent genotype remains to be answered. PMID- 11115794 TI - Micronutrients and immune function in cattle. AB - Complex inter-relationships exist between certain micronutrients, immune function and disease resistance in cattle. Several micronutrients have been shown to influence immune responses. The relationship between deficiencies of some micronutrients and disease resistance is less clear. A number of studies have indicated that Cr supplementation may improve cell-mediated and humoral immune response as well as resistance to respiratory infections in stressed cattle. With respiratory-disease challenge models Cr generally does not affect disease resistance. Deficiencies of Cu, Se, vitamin E and Co in cattle reduce the ability of isolated neutrophils to kill yeast and/or bacteria. Cu deficiency reduces antibody production, but cell-mediated immunity is generally not altered. However, Cu deficiency appears to reduce production of interferon and tumour necrosis factor by mononuclear cells. Numerous studies have linked low vitamin E and/or Se status to increased susceptibility of dairy cows to intramammary infections. In contrast to findings in laboratory animals, marginal Zn deficiency does not appear to impair antibody production or lymphocyte responsiveness to mitogen stimulation in ruminants. Co deficiency has been associated with reduced resistance to parasitic infections. It is well documented that vitamin A deficient animals are more susceptible to various types of infections. beta Carotene, possibly via its antioxidant properties, may affect immune function and disease resistance independent of its role as a precursor of vitamin A. PMID- 11115795 TI - Glutathione and immune function. AB - The immune system works best if the lymphoid cells have a delicately balanced intermediate level of glutathione. Even moderate changes in the intracellular glutathione level have profound effects on lymphocyte functions. Certain functions, such as the DNA synthetic response, are exquisitely sensitive to reactive oxygen intermediates and, therefore, are favoured by high levels of the antioxidant glutathione. Certain signal pathways, in contrast, are enhanced by oxidative conditions and favoured by low intracellular glutathione levels. The available evidence suggests that the lymphocytes from healthy human subjects have, on average, an optimal glutathione level. There is no indication that immunological functions such as resistance to infection or the response to vaccination may be enhanced in healthy human subjects by administration of glutathione or its precursor amino acid cysteine. However, immunological functions in diseases that are associated with a cysteine and glutathione deficiency may be significantly enhanced and potentially restored by cysteine supplementation. This factor has been studied most extensively in the case of human immunodeficiency virus (HIV)-infected patients who were found to experience, on average, a massive loss of S equivalent to a net loss of approximately 4 g cysteine/d. Two randomized placebo-controlled trials have shown that treatment of HIV-infected patients with N-acetyl-cysteine caused in both cases a significant increase in all immunological functions under test, including an almost complete restoration of natural killer cell activity. It remains to be tested whether cysteine supplementation may be useful also in other diseases and conditions that are associated with a low mean plasma cystine level and impaired immunological functions. PMID- 11115796 TI - Dietary treatments of obesity. AB - Numerous dietary treatments that purport to promote something unique for stimulating weight loss have been published. These treatments include fad diets, diets formulated by various commercial slimming clubs, very-low-energy diets (VLCD) and conventional diets. Fad diets may possibly reduce some weight short term; however, there is no scientific basis to their long-term use. Commercial slimming clubs may be suitable for some individuals but they need to be properly assessed professionally. There are specific guidelines for the use of VLCD, which are only appropriate for short-term use. There is scientific evidence to suggest that conventional diets can produce both short- and long-term weight loss. A successful weight-loss programme depends on a multidisciplinary team approach. Management strategies should be devised for addressing issues such as goals, monitoring, follow-up, relapse and evaluation. Initial assessments should include medical, laboratory and anthropometric data, fitness level and dietary and behavioural attitudes. These results will form the basis of the treatment plan. Frequent visits to the clinic are fundamental in promoting continuing weight loss during the long-term maintenance stage of treatment. The visits should be made worthwhile for the patient. Realistic and attainable goals for diet, exercise and behaviour modification should be made. The diet should have a novel approach and be tailored to the needs of the patient. It should be adequate nutritionally, low in energy and fat. The overall aim should be to promote lifelong changes in lifestyle, improvement in quality of life and health risks. PMID- 11115797 TI - The role of food ethics in food policy. AB - Certain developments in the agricultural and food sciences have far-reaching implications for society and the environment, which suggest the need to examine their ethical acceptability as a standard component of technology assessment. Such considerations have led to the emergence of a new academic discipline, food ethics. The present paper describes how ethical theory may be applied to the analysis of the impacts of prospective food biotechnologies to assess potential effects on four 'interest groups', i.e. consumers, producers, treated organisms and the biota (fauna and flora). The principles which structure the framework used, i.e. the ethical matrix, are adapted to the field of agriculture and food from those applied in medical ethics. Use of the ethical matrix is illustrated by applying it to the specific case of bovine somatotrophin, the genetically engineered protein hormone which is injected into lactating cattle to increase their milk yields. Ethical analysis is seen to depend on a number of critical requirements, i.e. scientific data, non-scientific evidence and predictions, suitably-qualified assessors ('competent moral judges'), the 'world-views' of the assessors and application of the precautionary principle to cope with 'uncertainty'. PMID- 11115798 TI - Ethical dilemmas in choosing a healthful diet: vote with your fork! AB - Dietary guidelines for health promotion and disease prevention in the USA recommend a consumption pattern based largely on grains, fruit and vegetables, with smaller amounts of meat and dairy foods, and even smaller amounts of foods high in fat and sugar. Such diets are demonstrably health promoting, but following them raises ethical issues related to the role of nutritionists in advising the public about healthful dietary choices, as well as to the role of the food industry in food production and marketing. In the USA a shift towards a more plant-based diet would affect the economic interests of producers of food commodities, food products and meals prepared outside the home; it would also affect the environment, food prices, trade with other countries (developing as well as industrialized) and relationships among the food industry, government agencies (domestic and international) and food and nutrition professionals. In a free-market economy any dietary choice has consequences for food producers. Thus, considerations of ethical dilemmas in choosing healthful diets suggest that food choices are political acts that offer opportunities for all parties concerned to examine the consequences of such choices and 'vote with forks'. PMID- 11115799 TI - Hyperhomocyst(e)inemia, related vitamins and dementias. AB - Vitamin B12 and to a lesser extent folate deficiencies have been associated with dementias. Both these vitamins are determinants of plasma total homocysteine concentrations. In this review the frequency distributions of plasma vitamin B12, folate and homocysteine in South African males (# 51 yrs and > 51 yrs) illustrate the lower vitamin B12 levels in older subjects, and the shift toward elevated homocysteine concentrations in elderly people. Vitamin B12 deficiency appears to be associated with neuropsychiatric disorders, including dementias, but no causal relationship based on biochemical evidence has so far been established. Supplementation with vitamin B12 improves some neurological abnormalities and reverses only mild dementia of recent onset, but does not slow the progression of dementia. Elevated homocysteine levels appears to affect cognitive function, as measured by spatial copying skills and visual event-related potentials. Measurement of plasma homocysteine may help identify individuals with vitamin deficiencies and hyperhomocysteinemia. The relation between B-vitamins, homocysteine and dementia needs to explored further before vitamin supplementation is advocated to prevent or reverse neuropsychiatric disorders. PMID- 11115800 TI - Is there epidemiologic evidence that anti-oxidants protect against disorders in cognitive function? AB - Experimental studies have identified several pathways through which oxidative stress could adversely affect cognitive function and increase the risk for dementia. Anti-oxidant supplements have been proposed as interventions for secondary or primary prevention against dementia. The existing observational studies and randomized trials examining anti-oxidants and cognitive function outcomes are reviewed. A discussion is provided of methodologic limitations of existing studies and possible directions for future investigations. PMID- 11115801 TI - Fatty acid intake and the risk of dementia and cognitive decline: a review of clinical and epidemiological studies. AB - Dietary intake of fatty acids may be related to dementia and cognitive function through a number of plausible mechanisms, such as atherosclerosis and thrombosis, inflammation, via an effect on brain development and membrane functioning, or via accumulation of beta-amyloid. This review gives an overview of the few studies that have investigated the relationship between fatty acid intake (including the fatty acids from fish) and cognitive function or dementia and summarises the results from two Dutch population-based prospective studies: the Zutphen Elderly Study (n=476) and the Rotterdam Study (n=5,386). Additionally, limitations on dietary intake studies are discussed and possible mechanisms behind the investigated associations. Data from the Rotterdam Study showed that high intakes of the following nutrients were associated with an increased risk of dementia after adjustment for confounders: total fat (RR=2.4 (95%CI: 1.1-5.2)), saturated fat (RR=1.9 (95%CI: 0.9-4.0)), and cholesterol (RR=1.7 (95%CI: 0.9-3.2)). A high fish consumption, an important source of n-3 PUFAs, reduced the risk of dementia (RR=0.4 (95%CI: 0.2-0.9)). In the Zutphen Elderly Study a high linoleic acid intake was associated with cognitive impairment (OR=1.8 (95%CI: 1.0-3.0)). A high fish consumption tended to be inversely associated with cognitive impairment and decline (RR=0.5, 95%CI: 0.2-1.2). Since diet is a risk factor that is suitable for intervention these results are hopeful and potentially very important. PMID- 11115802 TI - Glucose regulation and brain aging. AB - Blood glucose regulation is not only a complex phenomenon but glucose regulatory levels also vary significantly across individuals. Thus, whereas individuals compromised with moderately elevated blood glucose levels are diagnosed as having impaired glucose tolerance, excessive blood glucose levels render a Type II diabetes diagnosis. Type II diabetes prevalence rates in the adult population have been estimated to be between 6 and 10 percent. Although Type II diabetes has been typically associated with older people, the disease has become much more common among young adults and children. It has become increasingly evident that protracted glucose tolerance impairment usually precedes a type II diabetes diagnosis, although impaired glucose tolerance will not necessarily progress to a diabetic state. Furthermore, a number of studies have shown that impaired glucose tolerance or type II diabetes is associated with impaired cognitive function in older subjects. In addition, we recently found that cognitive deficits are also associated with moderately impaired glucose regulation in young healthy volunteers. These data, although in need of confirmation and extension, suggest that impaired glucose tolerance is associated with impaired cognition, independent of age. Moreover, since impaired glucose tolerance is more prevalent than diabetes across all ages, then our finding lead to the implication that impaired cognitive function may be more prevalent in the general population than previously estimated. Finally, the dysfunction of glucoregulatory mechanisms may be an important intervening factor when studying the evolution of cognitive function through the aging process. PMID- 11115803 TI - Dysphagia in Alzheimer disease: a review. AB - Pseudobulbar dysphagia is a common feature of Alzheimer disease (AD) especially in the late stages. In the majority of cases the clinician can select the most appropriate therapeutic modalities based on a thorough history and bedside assessment. The role of videofluoroscopy in managing the dysphagia of AD has not been established. It is unclear whether the weight loss associated with advanced AD can entirely be prevented by optimizing the management of dysphagia. Pneumonia is a common cause of morbidity and death in patients with AD. The risk of pneumonia is related not only to dysphagia and aspiration but to mobility, nutritional status and host immune response. Prevention of pneumonia through appropriate management of dysphagia is not supported by empirical evidence. The potential role of enteral feeding in patients with advanced AD is small. An evidence-based approach to enteral feeding in AD patients is outlined. PMID- 11115804 TI - Blood homocysteine and vitamin B levels are not associated with cognitive skills in healthy normally ageing subjects. AB - Increased plasma total homocysteine (tHcy) levels are a known risk factor for vascular disease and have been reported in association with cognitive impairment of old age. Alternatively, however, increased tHcy levels may simply be an indicator of B vitamin deficiency. We evaluated the relationship between plasma tHcy levels, serum vitamin B12 and folate levels, and the scores at a battery of neuropsychological tests in 54 healthy cognitively normal subjects aged 65 years and over. Hyperhomocysteinemia prevalence (plasma tHcy>15 micromol/L) was about 24%. In univariate analysis, vitamin B12 levels were associated with both verbal memory and visuo-spatial skills, whereas no association was found between psychometric test scores and folate levels or tHcy levels. However, none of the univariate associations of neuropsychological test scores and serum B12 vitamin levels was confirmed when adjusting for age, education and other confounding variables. In conclusion, although a relationship between homocysteine, B vitamins and poor cognitive skills in the elderly is plausible, this study does not suggests that such relationship is biologically important. PMID- 11115805 TI - Validity of self-reported height and weight estimates in cognitively-intact and impaired elderly individuals. AB - OBJECTIVE: A high prevalence of undernutrition has been observed in the elderly, particularly in cognitively impaired or demented individuals. Self-reported height and weight were tested as simple and non-invasive methods to efficiently screen individuals at risk. DESIGN: Cross-sectional study. PARTICIPANTS: A subset of subjects (n=465) participating in the longitudinal follow-up phase of the Canadian Study of Health and Aging (CSHA) and comprising cognitively intact and impaired individuals as well as demented subjects. MEASUREMENTS: Self-reported values of height and weight were compared to direct standard measurements using Pearson's correlation coefficients and linear regressions by cognitive status. Estimation bias was determined using paired Student t-tests. Sensitivity and specificity of body mass index (BMI) derived from self-reported data were calculated. RESULTS: Self-reported and measured weights were highly correlated (r>.90) in all three categories of cognitive status. A tendency to underestimate their weight was observed in overweight women. Correlations of recalled to measured height were excellent in normal (r=.91) and good in cognitively impaired (r=.86) and demented (r=.85) subjects. A systematic overestimation of recalled height was observed, particularly among individuals of short stature. Self reported BMI showed excellent sensitivity (>93%) in detecting underweight individuals in all three categories. CONCLUSION: Self-reported height and weight data can be obtained in normal and cognitively-impaired elderly persons as well as in mild or moderate cases of dementia and can be used as a valid tool to screen for risk of undernutrition. PMID- 11115806 TI - Growth hormone and thyrotropin hormone secretion in Alzheimer's disease. AB - The present study examined the relationship among GH and TSH secretion and neuropsychological and morphological tests in patients with Alzheimer's disease (AD). We studied 28 healthy controls and 28 AD patients with psychoneurological tests, and correlated their data with morphological data obtained by computed tomography and hormone responses of GH to GHRH, and TSH to TRH. The GH response to GHRH was higher in AD patients than in controls, 12.2 +/- 2 vs. 5.2 +/- 2.1 mcg/l, p< 0.01 but there was no difference in TSH response to TRH. On neuropsychological tests, patients were classified into 3 stages, these were indistinguishable on the basis of morphological data, GH and TSH responses at all stages including early-onset. Although GH response to GHRH was higher in AD patients than in controls, our results suggest that psychometric discrimination may be more accurate in diagnosis and staging of AD, than in predicting by morphological and hormonal responses. PMID- 11115807 TI - Effects of age and dietary restriction on animal model SAMP8 mice with learning and memory impairments. AB - This study was to investigate a hypothesis that dietary restriction (DR) suppresses learning and memory impairments in dementia animal model SAMP8 mice. Four-week-old female SAMP8 mice were fed either ad libitum (AL) or fed restricted (40% of the food consumed by AL). Results showed that acetylcholine (ACh) levels in hippocampus at aged 12 months of age were 12% higher in DR than that of AL group. Dopamine (DA) and norepinephrine (NE) levels in cerebellum at 8 and 12 months of age were significantly higher (26~94% and 34~43%, respectively) in DR group than those in AL group. Serotonin (5-HT) levels in cerebellum at aged 12 months of age were markedly increased (~53%) in DR group. Homovanillic acid (HVA) and 5-hydroxyindole acetic acid (5-HIAA) levels in cerebellum at 8 and 12 months of age were significantly increased (28~41% and 24~64%, respectively) in DR group compared with AL group. In addition, neurotransmitter-related enzymes, choline acetyltransferase (ChAT) and acetylchoinesterase (AChE) activities at 8 and 12 months of age were elevated (6~8% and 5~7%, respectively) in DR group. Monoamine oxidase-B (MAO-B) that catalyzes oxidative deamination in brain were suppressed by 7~10% in DR group. At aged 12 months of age, the generation of basal and induced reactive oxygen species (ROS) in brain significantly decreased by 20% in DR group compared with AL group. These results suggest that inhibitory effect of oxidative stress by DR may play a pivotal role in attenuating the age-related changes observed in dementia animal model SAMP8. PMID- 11115808 TI - Daily physical activity, aging and body composition. AB - Increasing age is associated with changes in physical activity and loss of fat free mass. The question is whether an active life style can delay the progressive loss of fat-free mass and functional capacity. A review of available evidence from doubly labeled water studies showed that there is no association between fat free mass and the level of physical activity in subjects over 60. Physical activity does not seem to affect the age-induced loss of fat-free mass. However, an active life style has positive effects on muscle function and thus is an important determinant of independent living at old age. PMID- 11115809 TI - Reproducibility of body composition and body water spaces measurements in healthy elderly individuals. AB - BACKGROUND AND AIMS: The assessment of body composition and water spaces in elderly persons, especially in disease situations, would be a critical component of the nutritional and hydration management. Bioelectrical impedance analysis (BIA) is a validated bed-side technique for the estimate of total body (TBW) and extracellular (ECW) water. The aim of the present study was to address the reproducibility of estimates of TBW, ECW and fat-free mass (FFM) in healthy elderly individuals (n=24, 61 +/- 3.6 yrs). METHODS: Repeated measurements of weight, height, skinfold thickness, TBW and ECW (BIA) were obtained (in almost identical experimental conditions) on days 0, 14, 28 and 42. FFM and the hydration fraction of FFM (HF) were calculated with a three compartment model. RESULTS: Analysis of variance for repeated measurements showed that time did not explain a significant part of the variance in water spaces, FFM, and HF. Mean SDs for repeated measurements were well within 1 litre for water spaces, 1 kg for FFM and 1% for HF. CONCLUSIONS: We conclude that reproducible measurements of water spaces can be obtained with BIA, which could be used to monitor hydration changes. Given the small variability in HF, FFM, can be derived from BIA with good reproducibility. PMID- 11115810 TI - Quality of life and stimulation of weight gain after treatment with megestrol acetate: correlation between cytokine levels and nutritional status, appetite in geriatric patients with wasting syndrome. AB - BACKGROUND: The geriatric wasting syndrome (GWS) has been associated with proinflammatory cytokines, depression and progressive decline in quality of life (QOL). The objective of this study was to evaluate the correlation between the changes in cytokine levels and appetite, nutritional markers, and QOL in geriatric patients with GWS following a randomized clinical trial of megestrol acetate (MA) versus placebo. METHODS: This was a prospective, double-blind, placebo-controlled trial. We evaluated 69 predominantly male (3 females) nursing home residents with weight loss of > or =5% of their usual body weight over the past three months or body weight 20% below their ideal body weight. Patients were randomly assigned to receive either placebo or megestrol acetate (MA) oral suspension (O.S.) 800 mg/day for 12 weeks and were then followed for 13 weeks off treatment. Data on appetite, weight, nutritional status, QOL and cytokine levels were collected at baseline and week 12. The correlation between appetite, weight, nutritional status, sense of well being and cytokine level changes in response to MA treatment was examined at week 12. RESULTS: Appetite, sense of well being, and QOL assessed by an "enjoyment list" significantly improved in the MA arm. Rising prealbumin showed a negative correlation with decreasing IL-6 (r = -0.51), TNFR-p 55 (r = -0.49) and sIL-2R (r = -0.38). There was also an improvement in prealbumin and a decrease in IL-6 and TNFR-p55 in the MA-arm (p < 0.01). A correlation between a decrease in the IL-6 levels and improvement in depression (r = 0.50) was seen in the MA arm as well. Improvement in appetite positively correlated with increased enjoyment of life (r = -0.41), less depression (r = 0.34), improved sense of well being (r = 0.36), prealbumin gain (r = 0.30), and weight gain (r = 0.38) by 12 weeks. Also, improvement in appetite positively correlated with improvement in nutritional parameters such as prealbumin, albumin, fat free mass and weight in the MA arm. CONCLUSIONS: In a geriatric nursing home population with weight loss, reduction in cytokine levels after MA treatment correlates with improvement in appetite, prealbumin, albumin, and improvement in quality of life. PMID- 11115811 TI - Explaining differences in the severity of familial adenomatous polyposis and the search for modifier genes. PMID- 11115812 TI - A role for IL-18 in intestinal inflammation? PMID- 11115813 TI - Autoimmune hepatitis: a fertile field. PMID- 11115814 TI - HCV in healthcare workers: a lightning strike. PMID- 11115815 TI - The worm turns on Helicobacter pylori. PMID- 11115816 TI - Multivitamins, folate, and colon cancer. PMID- 11115817 TI - Impaired transit and tolerance of intestinal gas in the irritable bowel syndrome. AB - BACKGROUND: Patients with irritable bowel syndrome (IBS) frequently complain of excessive gas but their fasting volume of intestinal gas is apparently normal. We hypothesised that the pathophysiological mechanism involved may be impairment of intestinal gas transit. AIM: To investigate intestinal gas transit and tolerance in IBS patients compared with healthy subjects. METHODS: A gas mixture (N(2), O(2), and CO(2) in venous proportions) was infused into the jejunum of 20 patients with IBS and 20 healthy controls at 12 ml/min for four hours. Gas evacuation, initially flatus from the anus (two hours) and then intrarectally (two hours), was continuously recorded. Symptom perception (0-6 scale) and abdominal distension were measured at 10 minute intervals. RESULTS: After two hours of external gas (flatus) collection, 18 of 20 IBS patients had developed gas retention (>400 ml), increased gastrointestinal symptoms (score >3), or abdominal distension (>3 mm girth increment) compared with only four of 20 control subjects. During intrarectal gas collection, 13 of 17 patients still exhibited abnormal responses. CONCLUSION: A large proportion of patients with IBS can be shown to have impaired transit and tolerance of intestinal gas loads. This anomaly may represent a possible mechanism of IBS symptoms, specifically pain and bloating. PMID- 11115818 TI - An exaggerated sensory component of the gastrocolonic response in patients with irritable bowel syndrome. AB - BACKGROUND/AIMS: Visceral hypersensitivity is a feature of the irritable bowel syndrome (IBS). Postprandial symptoms are common in these patients. The effects of nutrients on colonic perception in IBS are incompletely understood. SUBJECTS: We studied 13 healthy subjects and 16 patients with IBS-eight had diarrhoea predominant (IBS-D) and eight constipation predominant (IBS-C) IBS. METHODS: Colonic perception thresholds to balloon distension and viscerosomatic referral pattern were assessed before and after duodenal infusion of lipid or saline, respectively. At the end of the infusions, plasma levels of gastrointestinal peptides were determined. RESULTS: Lipids lowered the thresholds for first sensation, gas, discomfort, and pain in the IBS group but only for gas in the control group. The percent reduction in thresholds for gas and pain after lipids was greater in the IBS and IBS-D groups but not in the IBS-C group compared with controls. IBS patients had an increased area of referred discomfort and pain after lipids compared with before infusion whereas the referral area remained unchanged in controls. No group differences in colonic tone or compliance were observed. In both groups higher levels of cholecystokinin, pancreatic polypeptide, peptide YY, vasoactive intestinal polypeptide, and neuropeptide Y were seen after lipids. Motilin levels were higher in patients and differences in the subgroups were observed. Levels of corticotrophin releasing factor were lower in the constipated group than in the diarrhoea group. CONCLUSIONS: Postprandial symptoms in IBS patients may be explained in part by a nutrient dependent exaggerated sensory component of the gastrocolonic response. PMID- 11115819 TI - Oral glutamine in the prevention of fluorouracil induced intestinal toxicity: a double blind, placebo controlled, randomised trial. AB - BACKGROUND: 5-Fluorouracil (FU) in association with folinic acid (FA) is the most frequently used chemotherapeutic agent in colorectal cancer but it often causes diarrhoea. Animal and human studies suggest that glutamine stimulates intestinal mucosal growth. AIM: To determine if oral glutamine prevents changes in intestinal absorption (IA) and permeability (IP) induced by FU/FA. METHODS: Seventy chemotherapy naive patients with colorectal cancer were randomly assigned to oral glutamine (18 g/day) or placebo before the first cycle of FU (450 mg/m(2)) and FA (100 mg/m(2)) administered intravenously for five days. Treatment was continued for 15 days, starting five days before the beginning of chemotherapy. IA (D-xylose urinary excretion) and IP (cellobiose-mannitol test) were assessed at baseline and four and five days after the end of the first cycle of chemotherapy, respectively. Patients kept a daily record of diarrhoea, scored using the classification system of the National Cancer Institute (Bethesda, Maryland, USA). Duration of diarrhoea was recorded and the area under the curve (AUC) was calculated for each patient. RESULTS: Baseline patient characteristics and basal values of IP and IA tests were similar in the two arms. After one cycle of chemotherapy, the reduction in IA (D-xylose absorption) was more marked in the placebo arm (7.1% v 3. 8%; p=0.02); reduction of IP to mannitol was higher in the placebo arm (9.2% v 4.5%; p=0.02); and urinary recovery of cellobiose was not different between the study arms (p=0.60). Accordingly, the cellobiose-mannitol ratio increased more in the placebo arm (0.037 v 0.012; p=0.04). Average AUC of diarrhoea (1.9 v 4.5; p=0.09) and average number of loperamide tablets taken (0.4 v 2.6; p=0.002) were reduced in the glutamine arm. CONCLUSIONS: Glutamine reduces changes in IA and IP induced by FU and may have a protective effect on FU induced diarrhoea. PMID- 11115820 TI - Epidermal growth factor reduces multiorgan failure induced by thioacetamide. AB - BACKGROUND: Multiorgan failure is a severe life threatening state where present therapeutic approaches are suboptimal. Epidermal growth factor (EGF) is a potent stimulant of repair in in vitro and in vivo models. We therefore examined its potential beneficial effect in reducing mortality and injury induced by the noxious agent thioacetamide (TAA). METHODS: Mice (20 per group) were fasted overnight and received a single intraperitoneal dose of human recombinant EGF at 10 or 30 microg/kg or saline (control). Either 30 minutes before or after EGF, all animals also received TAA (40 mg/kg intraperitoneally). Twenty four hours later, surviving animals were killed, tissues collected, and degree of organ injury assessed. RESULTS: Fifty per cent (10/20) of control animals died within the first 24 hour period. Mortality was almost completely prevented by the higher dose of EGF whether given before or after TAA (p<0.01) and was reduced by about 50% with the lower dose of EGF. In control animals, the entire length of the jejunum and ileum had necrosis with or without mucosal denudation. In contrast, necrosis affected only about 10-20% of the total length in EGF treated groups (both p<0.01 v control). Control animals showed marked glomerular tuft collapse, interstitial haemorrhage, and increased plasma creatinine levels. These effects were significantly reduced in animals given EGF (30 microg/kg; p<0.01). All groups showed similar changes in liver histology (centrilobular necrosis) and alanine transaminase levels (10-fold increase). CONCLUSIONS: Although EGF did not prevent the hepatotoxicity associated with TAA, it reduced mortality, renal injury, and gastrointestinal damage. These studies provide preliminary evidence that EGF may be a novel approach for the prevention and/or treatment of multiorgan failure. PMID- 11115821 TI - Colonic crypt cell proliferation state assessed by whole crypt microdissection in sporadic neoplasia and familial adenomatous polyposis. AB - BACKGROUND: It has yet to be established whether proliferative activity in the macroscopically normal colonic mucosa is causally correlated with neoplastic risk. Measurement of proliferative activity in human subjects is of necessity usually undertaken using indirect methods with inherent limitations, and relatively little has been published on the effect of normal biological variables on such indices. AIMS: To establish the validity of mitosis counts following whole crypt microdissection as an index of the crypt cell proliferative state (CCPS) and to examine the effect of normal biological variables (age, sex, and colonic site) and colonic neoplasia on the mitotic index in macroscopically normal human colon. SUBJECTS: Mucosal samples were obtained at colectomy or colonoscopy from 107 individuals (24 controls, 23 sporadic adenoma patients, 31 sporadic carcinoma patients, and 29 patients with familial adenomatous polyposis (FAP)). METHODS: Mucosal specimens were hydrated, hydrolysed, and small groups of crypts separated from the main specimen under a dissecting microscope. The total number of mitoses/crypt were counted by one observer for each of 10 complete crypts. RESULTS: Validation work established that whole crypt mitoses counts were reliable and reproducible. There was no relation between age and mean mitoses/crypt (Pearson correlation coefficient -0.1). The CCPS count was higher for males than for females (difference in means 2.8 (95% confidence interval 0.80 4.66)) among controls but there was no gender difference in the three disease groups. For all disease groups and controls, the crypt mitotic count showed a significant linear increase (p=0.004) from the rectum to the caecum. Biopsies from within 5 cm of the macroscopic margin of a carcinoma (near) gave a mean mitosis count of 12.6 while those from more than 10 cm (far) were lower but not significantly so (p=0.12) with a count of 9.0. The mean mitoses/crypt were similar for the controls and adenomas (5.6 and 4.7, respectively) but greater for the cancers and especially for FAP (8.3 and 14.2, respectively). Statistical analysis confirmed that there were significant differences (p<0.05) between controls and all disease groups together, between sporadic disease and FAP, and between adenoma and carcinoma subjects at each of the four colonic sites. Post hoc comparison by t test showed significantly greater CCPS for FAP compared with controls (p<0.001) and for sporadic cancer versus controls (p=0.04). CONCLUSIONS: Whole crypt microdissection and mitosis counting is a reliable, reproducible, and robust technique for assessing CCPS in the human colon. CCPS is unaffected by age but increases from the distal to the proximal colon. CCPS is increased if a sporadic cancer is present and markedly increased in FAP. However, the precise relation of an increased CCPS to the neoplastic process remains uncertain. PMID- 11115822 TI - Plasma levels of progastrin but not amidated gastrin or glycine extended gastrin are elevated in patients with colorectal carcinoma. AB - BACKGROUND: The relationship between plasma gastrin levels and colorectal cancer is controversial. When confounding factors which increase plasma gastrin levels are taken into account, it has been shown that gastrin levels are not elevated in patients with colorectal cancer. However, these studies only measured amidated gastrin. Total gastrin (which includes unprocessed, partially processed, and mature forms of gastrin) has been shown to be elevated in patients with colorectal cancer. AIMS: The aim of this study was to determine whether fasting plasma levels of progastrin, amidated gastrin, or glycine extended gastrin are elevated in patients with colorectal cancer or colorectal polyps compared with controls. METHODS: Progastrin, amidated gastrin, and glycine extended gastrin were estimated by radioimmunoassay using the following antibodies: L289, 109-21, and L2. Blood samples were analysed for Helicobacter pylori by an enzyme linked immunosorbent assay. RESULTS: Median progastrin levels were significantly higher in the cancer group (27.5 pmol/l) than in the polyp (< or =15 pmol/l) or control (< or =15 pmol/l) group (p=0.0001 There was no difference in median levels of amidated gastrin between groups. Median levels of amidated gastrin were significantly higher in H pylori positive patients (19 pmol/l) than in H pylori negative patients (8 pmol/l) (p=0.0022). Median plasma progastrin levels were significantly higher for moderately dysplastic polyps (38 pmol/l) compared with mildly dysplastic (15 pmol/l) and severely dysplastic (15 pmol/l) polyps (p=0.05). CONCLUSIONS: Plasma levels of progastrin, but not amidated gastrin or glycine extended gastrin, are significantly elevated in patients with colorectal cancer compared with those with colorectal polyps or controls, irrespective of their H pylori status. We conclude that measuring plasma progastrin levels in patients with colorectal cancer is warranted. PMID- 11115823 TI - Only fibres promoting a stable butyrate producing colonic ecosystem decrease the rate of aberrant crypt foci in rats. AB - BACKGROUND: Dietary fibres have been proposed as protective agents against colon cancer but results of both epidemiological and experimental studies are inconclusive. AIMS: Hypothesising that protection against colon cancer may be restricted to butyrate producing fibres, we investigated the factors needed for long term stable butyrate production and its relation to susceptibility to colon cancer. METHODS: A two part randomised blinded study in rats, mimicking a prospective study in humans, was performed using a low fibre control diet (CD) and three high fibre diets: starch free wheat bran (WB), type III resistant starch (RS), and short chain fructo-oligosaccharides (FOS). Using a randomised block design, 96 inbred rats were fed for two, 16, 30, or 44 days to determine the period of adaptation to the diets, fermentation profiles, and effects on the colon, including mucosal proliferation on day 44. Subsequently, 36 rats fed the same diets for 44 days were injected with azoxymethane and checked for aberrant crypt foci 30 days later. RESULTS: After fermentation had stabilised (44 days), only RS and FOS produced large amounts of butyrate, with a trophic effect in the large intestine. No difference in mucosal proliferation between the diets was noted at this time. In the subsequent experiment one month later, fewer aberrant crypt foci were present in rats fed high butyrate producing diets (RS, p=0.022; FOS, p=0.043). CONCLUSION: A stable butyrate producing colonic ecosystem related to selected fibres appears to be less conducive to colon carcinogenesis. PMID- 11115824 TI - Double blind, randomised, placebo controlled study of a platelet activating factor antagonist, lexipafant, in the treatment and prevention of organ failure in predicted severe acute pancreatitis. AB - BACKGROUND: Platelet activating factor (PAF) is believed to amplify the activity of key mediators of the systemic inflammatory response syndrome (SIRS) in acute pancreatitis, resulting in multiorgan dysfunction syndrome. We tested the hypothesis that a potent PAF antagonist, lexipafant, could dampen SIRS and reduce organ failure in severe acute pancreatitis. METHODS: We conducted a randomised, double blind, placebo controlled, multicentre trial of lexipafant (100 mg/24 hours intravenously for seven days commenced within 72 hours of the onset of symptoms) involving 290 patients with an APACHE II score >6. Power calculations assumed that complications would be reduced from 40% to 24%. Secondary end points studied included severity of organ failure, markers of the inflammatory response, and mortality rate. FINDINGS: Overall, 80/138 (58%) patients in the placebo group and 85/148 (57%) in the lexipafant group developed one or more organ failures. The primary hypothesis was invalidated by the unexpected finding that 44% of patients had organ failure on entry into the study; only 39 (14%) developed new organ failure. Organ failure scores were reduced in the lexipafant group only on day 3: median change -1 (range -4 to +8) versus 0 (-4 to +10) in the placebo group (p=0.04). Systemic sepsis affected fewer patients in the lexipafant group (13/138 v 4/148; p=0.023). Local complications occurred in 41/138 (30%) patients in the placebo group and in 30/148 (20%) in the lexipafant group (20%; p=0.065); pseudocysts developed in 19 (14%) and eight (5%) patients, respectively (p=0.025). Deaths attributable to acute pancreatitis were not significantly different. Interleukin 8, a marker of neutrophil activation, and E-selectin, a marker of endothelial damage, decreased more rapidly in the lexipafant group (both p<0.05); however, absolute values were not different between the two groups. INTERPRETATION: The high incidence of organ failure within 72 hours of the onset of symptoms undermined the primary hypothesis, and power calculations for future studies in severe acute pancreatitis will need to allow for this. Lexipafant had no effect on new organ failure during treatment. This adequately powered study has shown that antagonism of PAF activity on its own is not sufficient to ameliorate SIRS in severe acute pancreatitis PMID- 11115825 TI - Cystic fibrosis transmembrane regulator (CFTR) DeltaF508 mutation and 5T allele in patients with chronic pancreatitis and exocrine pancreatic cancer. PANKRAS II Study Group. AB - BACKGROUND: An increased risk of chronic pancreatitis has been described among carriers of the cystic fibrosis transmembrane regulator (CFTR) mutation. In addition, patients with cystic fibrosis may have a higher risk of exocrine pancreatic cancer. AIMS: To determine the prevalence of the DeltaF508 mutation and 5T allele, the most common CFTR disease related variants, and to assess their association with lifestyle factors in an unselected series of patients with chronic pancreatitis or pancreatic cancer. SUBJECTS: Patients recruited to the multicentre PANKRAS II study with a diagnosis of chronic pancreatitis and pancreatic cancer from whom normal DNA was available. METHODS: The DeltaF508 mutation and 5T allele were analysed using polymerase chain reaction amplified normal DNA. Information on clinical and lifestyle factors was obtained through personal interviews. RESULTS: Among patients with pancreatitis, no DeltaF508 alleles were found and the prevalence of the 5T allele was 10.5%, similar to that described in the general population. Among patients with pancreatic cancer, the prevalence of the DeltaF508 mutation and the 5T allele was 2.4% and 5.5%, respectively. 5T allele carriers with cancer consumed significantly less alcohol than non-carriers (p=0.038). CONCLUSIONS: Our findings do not support the view that the DeltaF508 mutation and 5T allele confer a higher risk of chronic pancreatitis or pancreatic cancer. Nevertheless, our data suggest that interactions between CFTR polymorphism and environmental factors may play a role in the pathogenesis of these diseases. Our study emphasises the need for a multinational study to conclusively establish the role of CFTR variants as genetic susceptibility factors for chronic pancreatitis and pancreatic cancer. PMID- 11115826 TI - Fatty acid bile acid conjugates (FABACs)--new molecules for the prevention of cholesterol crystallisation in bile. AB - BACKGROUND: Cholesterol gall stones are a frequent disease for which at present surgery is the usual therapy. Despite the importance of bile acids it has become evident that phospholipids are the main cholesterol solubilisers in bile. Even phospholipid components, such as fatty acids, have anticrystallising activity. AIM: To synthesise fatty acid bile acid conjugates (FABACs) and study their effects on cholesterol crystallisation in bile in vitro and in vivo. METHODS: FABACs were prepared by conjugation of cholic acid at position 3 with saturated fatty acids of variable chain length using an amide bond. Cholesterol crystallisation and its kinetics (crystal observation time, crystal mass) were studied in model bile, pooled enriched human bile, and fresh human bile using FABACs with saturated fatty acids of varying chain length (C-6 to C-22). Absorption of FABACs into blood and bile was tested in hamsters. Prevention of biliary cholesterol crystallisation in vivo was tested in hamsters and inbred mice. RESULTS: FABACs strongly inhibited cholesterol crystallisation in model as well as native bile. The FABACs with longer acyl chains (C-16 to C-22) were more effective. At a concentration of 5 mM, FABACs almost completely inhibited cholesterol crystallisation in fresh human bile for 21 days. FABACs were absorbed and found in both portal and heart blood of hamsters. Levels in bile were 2-3 times higher than in blood, indicating active secretion. Appreciable levels were found in the systemic circulation 24-48 hours after a single administration. Ingested FABACs completely prevented the formation of cholesterol crystals in the gall bladders of hamsters and mice fed a lithogenic diet. CONCLUSIONS: FABACs are potent inhibitors of cholesterol crystallisation in bile. They are absorbed and secreted into bile and prevent the earliest step of cholesterol gall stone formation in animals. These compounds may be of potential use in cholesterol gall stone disease in humans. PMID- 11115827 TI - Aberrant cyclooxygenase isozyme expression in human intrahepatic cholangiocarcinoma. AB - METHODS: Cellular localisation of the cyclooxygenase (COX) isozymes COX-1 and COX 2 was analysed in 24 cholangiocarcinomas, including 17 matched tissues originating from non-tumorous liver tissue adjacent to tumours and seven biopsies of normal human liver, by immunohistochemistry using isozyme selective antibodies. RESULTS: In normal liver, constitutive expression of COX-2 protein was a characteristic feature of hepatocytes whereas no COX-2 immunosignal was detectable in normal bile duct epithelium, Kupffer, and endothelial cells. In cholangiocarcinoma cells, COX-2 protein was strongly expressed at high frequency. The intensity, percentage of positive cells, and pattern of COX-2 expression were found to be independent of the stage of tumour differentiation. In hepatocytes of matched non-tumorous tissue, COX-2 expression was unaltered. In contrast, strong COX-1 expression was frequently localised to Kupffer cells, endothelial cells, and occasionally to hepatocytes, but not to bile duct epithelial cells. In approximately half of moderately and poorly differentiated but not well differentiated cholangiocarcinomas, weak to moderate COX-1 staining was found in tumour cells while COX-1 expression in Kupffer cells was much more pronounced. CONCLUSION: Aberrant COX-2 expression occurs during the early stage while COX-1 over expression seems to be related to later stages of cholangiocarcinogenesis. PMID- 11115828 TI - Dual mechanism of vascular endothelial growth factor upregulation by hypoxia in human hepatocellular carcinoma. AB - BACKGROUND/AIMS: Vascular endothelial growth factor (VEGF) plays a key role in regulation of tumour associated angiogenesis. In the current study we analysed expression of VEGF and its receptors in human hepatocellular carcinoma (HCC) and investigated the molecular mechanisms of VEGF regulation by hypoxia. METHODS: VEGF, kinase domain region (KDR)/fetal liver kinase 1 (flk-1), and flt-1 expression were examined by immunohistochemistry and in situ hybridisation in 15 human HCC tissues. Expression of VEGF and regulation by hypoxia were assessed in three human HCC cell lines using a quantitative competitive reverse transcription polymerase chain reaction, ELISA, and a series of 5' deletion reporter gene constructs of the human VEGF promoter in transient transfection assays. RESULTS: We observed over expression of VEGF mRNA and protein in HCC compared with cirrhosis or normal liver. Expression of VEGF in tumour cells was strongly increased in areas directly adjacent to necrotic/hypoxic regions. Both VEGF receptors were detected in vascular endothelia of HCC while only KDR/flk-1 receptors were detected in endothelial cells of cirrhotic livers. Expression of VEGF was observed in all human HCC cell lines examined. Hypoxia (1% oxygen) resulted in profound upregulation of VEGF mRNA and protein levels. Furthermore, hypoxia treatment resulted in a doubling of VEGF mRNA stability. Deletion analysis of the human VEGF 5' flanking region -2018 and +50 demonstrated induction of VEGF promoter activity under hypoxic conditions which was significantly decreased following deletion of the region -1286 and -789 suggesting a substantial contribution of the -975 putative hypoxia inducible factor 1 binding site to hypoxia mediated transcriptional activation of the VEGF gene. CONCLUSION: These data suggest hypoxia as a central stimulus of angiogenesis in human HCC through upregulation of VEGF gene expression by at least two distinct molecular mechanisms: activation of VEGF gene transcription and an increase in VEGF mRNA stability. PMID- 11115829 TI - Management and outcome of pregnancy in autoimmune hepatitis. AB - BACKGROUND: There is a paucity of data in the literature on the risks associated with, and optimal management of, pregnancy in patients with autoimmune hepatitis (AIH). AIMS: To assess maternal and fetal outcomes in relation to clinical management of pregnancy in a large cohort of patients with well defined AIH. METHODS: A review of all known pregnancies in 162 females with definite AIH attending our clinics between 1983 and 1998, with respect to treatment, natural history, and outcome. RESULTS: Thirty one live births (one twin) resulted from 35 pregnancies in 18 women (seven with cirrhosis). Median age at conception was 28 years (range 18-36). Two patients presented with AIH de novo during pregnancy. At conception, in 15 pregnancies patients had been receiving azathioprine alone or (in nine) with prednisolone, in seven prednisolone alone, and in one cyclosporin. Fetal loss at > or =20 weeks' gestation occurred in two instances. Flares in disease activity occurred during four pregnancies and within three months of delivery in a further four. Among the 31 children born (median follow up 10 years) only two abnormalities have been identified: Perthes' disease in one and severe mental and physical handicap in a second who was born prematurely following decompensation of the mother's liver disease. Neither mother was receiving azathioprine. CONCLUSIONS: Successful completion of pregnancy is a realistic expectation for patients with well controlled AIH. Treatment options vary, but azathioprine appears to be generally safe and without adverse outcomes for mother or baby. Vigilance is required, however, and patients need to be monitored carefully during pregnancy and for several months post partum. PMID- 11115830 TI - Hepatocellular carcinoma in Central Europe: prognostic features and survival. AB - BACKGROUND AND AIMS: We investigated the influence of baseline characteristics of patients with hepatocellular carcinoma (HCC) on prognosis and developed a multivariate Cox model predicting survival. All patients were from Central Europe. METHODS: All 245 patients seen at the Department of Gastroenterology and Hepatology at the University of Vienna, Austria, from July 1991 to March 1998 were included in this retrospective study. Nineteen different clinical characteristics and survival time from date of diagnosis were noted. Factors determining survival time were analysed by univariate and multivariate analysis using Cox proportional hazard regression models and a new classification model was constructed. The validity of this model was tested on an independent group of 89 patients, seen from April 1998 to September 1999. RESULTS: Median survival in patients with HCC was 8.0 months. In a multivariate analysis bilirubin (>2 mg/dl), portal vein thrombosis, prothrombin time (<70%), alpha fetoprotein (>180 microg/l), tumour mass >50%, and enlarged lymph nodes were independent predictors of survival. A newly constructed Cox proportional hazard model (Vienna survival model for HCC=VISUM-HCC) identified three disease stages with different durations of survival (median survival stage 1, 15.2 months; stage 2, 7.2 months; and stage 3, 2.6 months; p=0.00001). Applying the VISUM-HCC survival model to patients in Okuda stage 2 identified subgroups with an excellent and very poor prognosis for which different treatment modalities should be offered. CONCLUSIONS: Our patients with HCC had a poor median survival of eight months. Six easily measurable clinical variables were significant predictors of survival in patients with HCC. The new VISUM-HCC survival model may be useful for stratifying patients with HCC for various clinical treatment modalities. PMID- 11115832 TI - A study of hepatitis C prevalence in healthcare workers in the West of Scotland. AB - BACKGROUND AND AIMS: Whether healthcare workers have an increased prevalence of hepatitis C virus infection as a result of exposure to patient's blood and body fluids is controversial. This study assesses the prevalence of hepatitis C virus infection in healthcare workers, and its relation to the performance of exposure prone procedures and duration of occupational exposure, allowing an estimate to be made of the incidence of occupationally acquired hepatitis C infection among medical staff. METHODS: In this anonymous retrospective cohort study, we estimated the prevalence of hepatitis C infection in 10 654 healthcare workers. ELISA-3 testing was performed on pools of five sera collected during immunisation against hepatitis B. Healthcare workers were arranged into five occupational groups, according to the degree of patient exposure, and three age bands (<30 years, 30-39 years, >40 years). RESULTS: Prevalence of antibodies to hepatitis C was 0.28% (30/10 654), comparable in all occupational groups (p=0.34) and unrelated to duration of potential exposure. Assuming that all detected infections had been occupationally acquired, the maximum estimated risk of hepatitis C infection in exposure prone medical staff was low: 1.4% for surgeons and 1.0% for physicians over a 35 year professional career. CONCLUSIONS: Hepatitis C infection is infrequent in healthcare workers in Glasgow. Those conducting exposure prone procedures do not seem to be at higher risk than other healthcare staff. PMID- 11115831 TI - Evidence for an association between the aetiology of cirrhosis and pattern of hepatocellular carcinoma development. AB - BACKGROUND: Patients with liver cirrhosis are at significant risk of hepatocellular carcinoma (HCC) that may develop as well defined nodular lesions or as more aggressive infiltrating tumours. AIM: To compare prospectively risk factors associated with nodular or infiltrating HCC in cirrhotic patients. PATIENTS AND METHODS: We studied 370 patients with cirrhosis, followed prospectively by periodic ultrasound (US) of the liver, for a mean period of 74.6 (SD 32.4) months to define the incidence and patterns of HCC development. Patients who developed HCC were compared according to tumour pattern using univariate and multivariate analysis. RESULTS: Sixty one (16.5%) patients developed HCC: HCC was classified as nodular in 49 (80.3%) and infiltrating in 12 (19.7%) according to US and computerised tomography (CT) imaging. The five and 10 year cumulative probabilities were 8.1% (95% confidence interval (CI) 5. 2%-11%) and 25.2% (15.0-35.4%) for nodular HCC and 2.1% (0.5-3.7%) and 6.9% (2.1-11.7%) for infiltrating HCC. Patients with infiltrating HCC were younger than those with nodular HCC (59.5 v 66. 2 years, 95% CI 55.2-63.8 and 64.1-68.3 years; p=0.014). Using multivariate analysis, development of nodular HCC was associated with older age (p=0.0002; relative risk (RR) 3.1; 95% CI 1.6-5.2), longer duration (p=0.09; RR 2.6; 95% CI 1.8-3.4), and more advanced stage (p=0.002; RR 2.5; 95% CI 1.3 4.5) of cirrhosis but not with the aetiology of liver disease. In contrast, development of infiltrating HCC appeared to be unrelated to age or disease duration or stage, while it was associated with hepatitis B virus infection (p=0.07; RR 3.96; 95% CI 1.1-5.2) and with hepatitis B/hepatitis C virus coinfection (p=0.0007; RR 16.9; 95% CI 3.8-36.7). CONCLUSIONS: In liver cirrhosis, we identified two patterns of HCC developing with distinct risk factors. Nodular HCC was related to the cirrhotic process per se independent of aetiological factors and may depend on the proliferative activity within regenerative nodules, while the infiltrating form of HCC was linked to hepatitis B virus infection and may reflect more direct virus induced carcinogenesis. PMID- 11115833 TI - Mitochondrial enteropathy: the primary pathology may not be within the gastrointestinal tract. AB - BACKGROUND: Mitochondrial DNA (mtDNA) defects are an important cause of disease. Although gastrointestinal symptoms are common in these patients, their pathogenesis remains uncertain. AIM: To investigate the role of the mtDNA defect in the production of gastrointestinal dysfunction. PATIENT: A 20 year old woman who presented at 15 years of age with recurrent vomiting and pseudo-obstruction, who did not respond to conservative management and ultimately had subtotal gastrectomy and Roux-en-y reconstruction. She subsequently presented with status epilepticus and was found to have a mitochondrial respiratory chain disorder due to a pathogenic mtDNA point mutation (A3243G). METHODS: Resected bowel was studied using light and electron microscopy and mtDNA analysed from both mucosal and muscular layers using polymerase chain reaction generated RFLP analysis. RESULTS: Histological and electron microscopic studies revealed no morphological abnormalities in the resected stomach, and molecular genetic analysis failed to identify the genetic defect in either the mucosal or muscle layers. CONCLUSION: This study suggests that in some individuals with gastrointestinal symptoms associated with established mitochondrial DNA disease, the primary pathology of the mitochondrial enteropathy lies outside the gastrointestinal tract. PMID- 11115834 TI - Visceral hypersensitivity: facts, speculations, and challenges. PMID- 11115836 TI - Influence of allele lineage on the role of the insulin minisatellite in susceptibility to type 1 diabetes. AB - The insulin minisatellite or variable number of tandem repeats locus (INS VNTR) is the best candidate for the type 1 diabetes mellitus (T1DM) susceptibility locus IDDM2. Small class I alleles associate with predisposition to T1DM, whereas large class III alleles associate with dominant protection. We have analysed variant repeat distribution within the minisatellite and combined this with flanking haplotypes to define five new ancestral allele lineages. Class III alleles divide into two highly diverged lineages, IIIA and IIIB, which correspond perfectly to the previously defined Protective (PH) and Very Protective (VPH) haplotypes, respectively. Class I alleles are divided into three newly defined lineages, IC+, ID+ and ID-, by a combination of variant repeat distributions and flanking haplotypes. All class I alleles are equally predisposing to T1DM except for ID- alleles which are protective when transmitted from ID-/III heterozygous fathers. Similar results have been previously reported for alleles of 42 repeats in length (allele 814) which represent a subset of the ID- lineage. Division of class ID- alleles into those of 42 repeats and those of other sizes suggested that this protective effect was a feature of all ID- alleles, irrespective of size. ID- alleles are only clearly distinguished from all other alleles by an MSPI(-) variant within IGF2 downstream of the minisatellite, suggesting that the apparent role of the minisatellite in susceptibility to T1DM may be modified by neighbouring haplotype and therefore that IDDM2 could have a multi-locus aetiology. PMID- 11115835 TI - Bacteria as the cause of ulcerative colitis. PMID- 11115837 TI - Mice lacking renal chloride channel, CLC-5, are a model for Dent's disease, a nephrolithiasis disorder associated with defective receptor-mediated endocytosis. AB - Nephrolithiasis (kidney stones) affects 5-10% of adults and is most commonly associated with hypercalciuria, which may be due to monogenic renal tubular disorders. One such hypercalciuric disorder is Dent's disease, which is characterized by renal proximal tubular defects that include low molecular weight proteinuria, aminoaciduria and glycosuria, together with rickets in some patients. Dent's disease is due to inactivating mutations of the renal-specific voltage-gated chloride channel, CLC-5, which is expressed in the proximal tubule, thick ascending limb and collecting duct. The subcellular localization of CLC-5 to the proximal tubular endosomes has suggested a role in endocytosis, and to facilitate in vivo investigations of CLC-5 in Dent's disease we generated mice lacking CLC-5 by targeted gene disruption. CLC-5-deficient mice developed renal tubular defects which included low molecular weight (<70 kDa) proteinuria, generalized aminoaciduria that was more pronounced for neutral and polar amino acids, and glycosuria. They also developed hypercalciuria and renal calcium deposits and some had deformities of the spine. Furthermore, endocytosis as assessed by horseradish peroxidase uptake in the proximal tubule was severely impaired in CLC-5-deficient mice, thereby demonstrating a role for CLC-5 in endosomal uptake of low molecular weight proteins. Thus, CLC-5-deficient mice provide a model for Dent's disease and this will help in elucidating the function of this chloride channel in endocytosis and renal calcium homeostasis. PMID- 11115838 TI - Major factors influencing linkage disequilibrium by analysis of different chromosome regions in distinct populations: demography, chromosome recombination frequency and selection. AB - Linkage disequilibrium (LD) mapping of disease genes is complicated by population and chromosome-region-specific factors. We have analysed demographic factors by contrasting intermarker LD results obtained in a large cosmopolitan population (UK), a large genetic isolate (Sardinia) and a subisolate (village of Gavoi) for two regions of the X chromosome. A dramatic increase of LD was found in the subisolate. Demographic history of populations therefore influences LD. Chromosome-region-specific effects, namely the pattern and frequency of homologous recombination, were next delineated by the analysis of chromosome 6p21, including the HLA region. Patterns of global LD in this region were very similar in the UK and Sardinian populations despite their entirely distinct demographies, and correlate well with the pattern of recombinations. Nevertheless, haplotypes extend across recombination hot spots indicative of selection of certain haplotypes. Subisolate aside, chromosome-region-specific differences in LD patterns appear to be more important than the differences in intermarker LD between distinct populations. PMID- 11115839 TI - The inter-regional distribution of HLA class II haplotypes indicates the suitability of the Sardinian population for case-control association studies in complex diseases. AB - We have analysed HLA class II gene-based substructure of the Sardinian population in order to evaluate the possible influence of this parameter in the mapping of common disease loci using association methods. We first examined the distribution of the HLA-DRB1-DQA1-DQB1 haplotypes in 631 newborns from seven different regions of the island, and found that the most frequent haplotypes were uniformly distributed in all regions, but at frequencies unique to Sardinia. Other haplotypes, common in other white European populations, are consistently rare or absent across the whole island. Analysis of molecular variance (AMOVA) showed a very low degree of genetic differentiation between the coastal regions, which have suffered repeated invasions over many years, and the most internal and isolated part of the island. This suggests that there has been little genetic flow from the various populations that have invaded the island during the last 3000 years and that Sardinia is a relatively homogeneous population. The validity of these unrelated control HLA haplotype frequencies and our claim of homogeneity were established by demonstrating the near identity of the affected family-based control (AFBAC) HLA haplotype frequencies in 243 type 1 diabetes and 495 multiple sclerosis families from Sardinia and those of the unrelated controls. These results indicate that robust case-control studies can be carried out in Sardinia offering cost efficiency over certain family-based designs. PMID- 11115840 TI - Confirmation of the DRB1-DQB1 loci as the major component of IDDM1 in the isolated founder population of Sardinia. AB - There is considerable uncertainty and debate concerning the application of linkage disequilibrium (LD) mapping in common multifactorial diseases, including the choice of population and the density of the marker map. Previously, it has been shown that, in the large cosmopolitan population of the UK, the established type 1 diabetes IDDM1 locus in the HLA region could be mapped with high resolution by LD. The LD curve peaked at marker D6S2444, 85 kb from the HLA class II gene DQB1, which is known to be a major determinant of IDDM1. However, given the many unknown parameters underlying LD, a validation of the approach in a genetically distinct population is necessary. In the present report we have achieved this by the LD mapping of IDDM1 in the isolated founder population of Sardinia. Using a dense map of microsatellite markers, we determined the peak of LD to be located at marker D6S2447, which is only 6.5 kb from DQB1. Next, we typed a large number of SNPs defining allelic variation at functional candidate genes within the critical region. The association curve, with both classes of marker, peaked at the loci DRB1-DQB1. These results, while representing conclusive evidence that the class II loci DRB1-DQB1 dominate the association of the HLA region to type 1 diabetes, provide empirical support for LD mapping. PMID- 11115841 TI - Molecular evolution of multiple recurrent cancers of the bladder. AB - We describe the reconstruction of bladder tumor development in individual patients spanning periods of up to 17 years. Genomic alterations detected in the tumors were used for hierarchical cluster analysis of tumor subclones. The cluster analysis highlights the clonal relationship between tumors from each patient. Based on the cluster data we were able to reconstruct the evolution of tumors in a genetic tree, where tumors with few aberrations precede those with many genetic insults. The sequential order of the tumors in these pedigrees differs from the chronological order in which the tumors appear. Thus, a tumor with few alterations can be occult for years following removal of a more deranged derivative. Extensive genetic damage is seen to accumulate during the evolution of the tumors. To explain the type and extent of genetic damage in combination with the low stage and grade of these tumors, we hypothesize that in bladder cancer pathogenesis an increased rate of mitotic recombination is acquired early in the tumorigenic process. PMID- 11115842 TI - Gene expression profiling of primary breast carcinomas using arrays of candidate genes. AB - Breast cancer is characterized by an important histoclinical heterogeneity that currently hampers the selection of the most appropriate treatment for each case. This problem could be solved by the identification of new parameters that better predict the natural history of the disease and its sensitivity to treatment. A large-scale molecular characterization of breast cancer could help in this context. Using cDNA arrays, we studied the quantitative mRNA expression levels of 176 candidate genes in 34 primary breast carcinomas along three directions: comparison of tumor samples, correlations of molecular data with conventional histoclinical prognostic features and gene correlations. The study evidenced extensive heterogeneity of breast tumors at the transcriptional level. A hierarchical clustering algorithm identified two molecularly distinct subgroups of tumors characterized by a different clinical outcome after chemotherapy. This outcome could not have been predicted by the commonly used histoclinical parameters. No correlation was found with the age of patients, tumor size, histological type and grade. However, expression of genes was differential in tumors with lymph node metastasis and according to the estrogen receptor status; ERBB2 expression was strongly correlated with the lymph node status (P < 0.0001) and that of GATA3 with the presence of estrogen receptors (P < 0.001). Thus, our results identified new ways to group tumors according to outcome and new potential targets of carcinogenesis. They show that the systematic use of cDNA array testing holds great promise to improve the classification of breast cancer in terms of prognosis and chemosensitivity and to provide new potential therapeutic targets. PMID- 11115843 TI - Allele association studies with SSR and SNP markers at known physical distances within a 1 Mb region embracing the ALDH2 locus in the Japanese, demonstrates linkage disequilibrium extending up to 400 kb. AB - There has been considerable recent debate concerning the distances over which levels of allelic association useful for genomic quantitative trait locus (QTL) scans can be detected. We have examined simple sequence repeat (SSR) polymorphisms and two single nucleotide polymorphisms (SNPs) in the region flanking the aldehyde dehydrogenase 2 locus, ALDH2, in populations of Japanese alcoholics and controls. These groups differ significantly in the allele frequencies for the functional SNP in exon XII of this gene located on chromosome 12. The results obtained with SSR markers complement recent investigations with SNPs over similar distances at the TCR alpha/delta locus. Significant allelic association with this marker could be detected for SSRs over distances up to 400 kb and over 37 kb for the SNP thereby extending the distance over which LD at this locus could be detected by an order of magnitude. Furthermore, as a proof of principle, we show that comparisons of allele frequency differences for the SSR markers in the case (alcoholics) and control populations would have detected the ALDH2 marker as a putative QTL. Extending the tests to include alleles at two or three flanking loci suggests that the power to detect QTLs through association can be enhanced significantly. PMID- 11115844 TI - Alternative splicing at the MEFV locus involved in familial Mediterranean fever regulates translocation of the marenostrin/pyrin protein to the nucleus. AB - Mutations in MEFV, a gene encoding a protein (marenostrin/pyrin) of unknown function, are associated with familial Mediterranean fever, a genetic condition characterized by febrile episodes of serosal inflammation. Based on its primary structure, this 781 residue protein is thought to function as a nuclear effector molecule. However, recent transient expression studies indicated a perinuclear cytoplasmic localization. Here, we describe the isolation and expression of a novel human MEFV isoform, MEFV-d2, generated by in-frame alternative splicing of exon 2. This transcript, expressed in leukocytes, predicts a 570 residue protein designated marenostrin-d2. To investigate differences in subcellular localization between the full-length protein (marenostrin-fl) and marenostrin-d2, while providing against the overexpression of transiently expressed proteins, we have generated CHO cell lines stably expressing these two isoforms fused to the green fluorescent protein. The localization pattern of marenostrin-d2 differs dramatically from that of marenostrin-fl. Marenostrin-fl is homogeneously distributed over the entire cytoplasm, whereas marenostrin-d2 concentrates into the nucleus. To map the critical domain(s) specifying these differences, deletion mutants have been generated. Deletion of the putative nuclear localization signals (NLS) does not alter the nuclear localization of marenostrin-d2 whereas, despite the lack of discernible NLS in the domain encoded by the exon 1-exon 3 splice junction, deletion of this domain indeed disrupts this localization. These data, which challenge the current domain organization model of marenostrin, strongly suggest that MEFV encodes a nuclear protein and raises the possibility that MEFV alternative splicing may control functions of wild-type and mutant marenostrin proteins by regulating their translocation to the nucleus. PMID- 11115845 TI - Defective intracellular transport and processing of OA1 is a major cause of ocular albinism type 1. AB - Ocular albinism type 1 (OA1) is an X-linked disorder mainly characterized by a severe reduction of visual acuity, hypopigmentation of the retina and the presence of macromelanosomes in the skin and eyes. Various types of mutation have been identified within the OA1 gene in patients with the disorder, including several missense mutations of unknown functional significance. In order to shed light into the molecular pathogenesis of ocular albinism and possibly define critical functional domains within the OA1 protein, we characterized 19 independent missense mutations with respect to processing and subcellular distribution on expression in COS-7 cells. Our analysis indicates the presence of at least two distinct biochemical defects associated with the different missense mutations. Eleven of the nineteen OA1 mutants (approximately 60%) were retained in the endoplasmic reticulum, showing defecNStive intracellular transport and glycosylation, consistent with protein misfolding. The remaining eight of the nineteen OA1 mutants (approximately 40%) displayed sorting and processing behaviours indistinguishable from those of the wild-type protein. Consistent with our recent findings that OA1 represents a novel type of intracellular G protein coupled receptor (GPCR), we found that most of these latter mutations cluster within the second and third cytosolic loops, two regions that in canonical GPCRs are known to be critical for their downstream signaling, including G protein coupling and effector activation. The biochemical analysis of OA1 mutations performed in this study provides important insights into the structure-function relationships of the OA1 protein and implies protein misfolding as a major pathogenic mechanism in OA1. PMID- 11115846 TI - The location and type of mutation predict malformation severity in isolated lissencephaly caused by abnormalities within the LIS1 gene. AB - Lissencephaly is a cortical malformation secondary to impaired neuronal migration resulting in mental retardation, epilepsy and motor impairment. It shows a severity spectrum from agyria with a severely thickened cortex to posterior band heterotopia only. The LIS1 gene on 17p13.3 encodes a 45 kDa protein named PAFAH1B1 containing seven WD40 repeats. This protein is required for optimal neuronal migration by two proposed mechanisms: as a microtubule-associated protein and as one subunit of the enzyme platelet-activating factor acetylhydrolase. Approximately 65% of patients with isolated lissencephaly sequence (ILS) show intragenic mutations or deletions of the LIS1 gene. We analyzed 29 non-deletion ILS patients carrying a mutation of LIS1 and we report 15 novel mutations. Patients with missense mutations had a milder lissencephaly grade compared with those with mutations leading to a shortened or truncated protein (P = 0.022). Early truncation/deletion mutations in the putative microtubule-binding domain resulted in a more severe lissencephaly than later truncation/deletion mutations (P < 0.001). Our results suggest that the lissencephaly severity in ILS caused by LIS1 mutations may be predicted by the type and location of the mutation. Using a spectrum of ILS patients, we confirm the importance of specific WD40 repeats and a putative microtubule-binding domain for PAFAH1B1 function. We suggest that the small number of missense mutations identified may be due to underdiagnosis of milder phenotypes and hypothesize that the greater lissencephaly severity seen in Miller-Dieker syndrome may be secondary to the loss of another cortical development gene in the deletion of 17p13.3. PMID- 11115847 TI - Differential chromatin packaging of genomic imprinted regions between expressed and non-expressed alleles. AB - Chromosomal regions subject to genomic imprinting comprise a functional domain exhibiting parental-specific expression of genes and hence may take a unique chromatin structure. Here we have examined the chromatin packaging state of allelic sites in the Zfp127/Snrpn locus on mouse chromosome 7 and in the Igf2r locus on mouse chromosome 17 with an assay consisting of chromatin fractionation and allele-specific detection. The results showed that non-transcribed alleles of Igf2r are packaged more compactly than transcribed alleles in F(1) hybrid mice of both types of cross between C57BL/6 and MSM strains, whereas a non-imprinted gene, Sod-2, in the vicinity of Igf2r does not show such a difference. This indicates a close correlation between imprinting and the differential packaging of chromatin. On the other hand, the Zfp127/Snrpn locus showed such an allele specific fractionation pattern only in F(1) hybrid mice of a cross but not in those of the reciprocal cross. Analysis of the congenic mice produced for this locus did not provide any difference. These results suggest that chromatin of imprinted domains in different compaction levels is affected by distinct blueprints in homologous chromosomes that are heritable through the germ line. PMID- 11115848 TI - Genetic heterogeneity and spectrum of mutations of the PRKAR1A gene in patients with the carney complex. AB - Carney complex (CNC) is an autosomal dominant multiple neoplasia syndrome, which has been linked to loci on 2p16 and 17q22-24. We recently reported that PRKAR1A, which codes for the type 1A regulatory subunit of protein kinase A (PKA), is a tumor suppressor gene on chromosome 17 that is mutated in some CNC families. To evaluate the spectrum of PRKAR1A mutations, we identified its genomic structure and screened for mutations in 54 CNC kindreds (34 families and 20 patients with sporadic disease). Fourteen families were informative for linkage analysis: four of four families that mapped to 17q had PRKAR1A mutations, whereas there were no mutations found in seven families exhibiting at least one recombination with 17q. In six of the latter, CNC mapped to 2p16. PRKAR1A mutations were also found in 12 of 20 non-informative families and 7 of 20 sporadic cases. Altogether, 15 distinct PRKAR1A mutations were identified in 22 of 54 kindreds (40.7%). In 14 mutations, the sequence change was predicted to lead to a premature stop codon; one altered the initiator ATG codon. Mutant mRNAs containing a premature stop codon were unstable, as a result of nonsense-mediated mRNA decay. Accordingly, the predicted truncated PRKAR1A protein products were absent in these cells. We conclude that (i) genetic heterogeneity exists in CNC; and (ii) all of the CNC alleles on 17q are functionally null mutations of PRKAR1A. CNC is the first human disease recognized to be caused by mutations of the PKA holoenzyme, a critical component of cellular signaling. PMID- 11115849 TI - Caveolin-3 deficiency causes muscle degeneration in mice. AB - Caveolin-3 is a muscle-specific protein integrated in the caveolae, which are small invaginations of the plasma membrane. Mutations of the caveolin-3 gene, localized at 3p25, have been reported to be involved in the pathogenesis of limb girdle muscular dystrophy (LGMD1C or caveolinopathy) with mild clinical symptoms, inherited through an autosomal dominant form of genetic transmission. To elucidate the pathogenetic mechanism, we developed caveolin-3-deficient mice for use as animal models of caveolinopathy. Caveolin-3 mRNA and its protein were absent in homozygous mutant mice. In heterozygous mutant mice, both the mRNA and its protein were normal in size, but their amounts were reduced by about half. The density of caveolae in skeletal muscle plasma membrane was roughly proportional to the amount of caveolin-3. In homozygous mutant mice, muscle degeneration was recognized in soleus muscle at 8 weeks of age and in the diaphragm from 8 to 30 weeks, although there was no difference in growth and movement between wild-type and mutant mice. No apparent muscle degeneration was observed in heterozygous mutant mice, indicating that pathological changes caused by caveolin-3 gene disruption were inherited through the recessive form of genetic transmission. PMID- 11115850 TI - Schwann cells harbor the somatic NF1 mutation in neurofibromas: evidence of two different Schwann cell subpopulations. AB - Neurofibromas are one of the most characteristic features of neurofibromatosis type 1 (NF1), an inherited autosomal-dominant neurogenetic disorder affecting 1 in 3500 individuals worldwide. These benign tumors mainly consist of Schwann cells (SCs) and fibroblasts. Recent evidence demonstrates that somatic mutations at the NF1 gene are found in neurofibromas, but it has not been demonstrated whether SCs, fibroblasts and/or both cell types bear a somatic loss of NF1. We recently established a cell culture system that allows selective expansion of human SCs from neurofibromas. We cultured pure populations of SCs and fibroblasts derived from 10 neurofibromas with characterized NF1 mutations and found that SCs but not fibroblasts harbored a somatic mutation at the NF1 locus in all studied tumors. Furthermore, by culturing neurofibroma-derived SCs under different in vitro conditions we were able to obtain two genetically distinct SC subpopulations: NF1(-/-) and NF1(+/-). These data strongly support the idea that NF1 mutations in SCs, but not in fibroblasts, correlate to neurofibroma formation and demonstrate that only a portion of SCs in neurofibromas have mutations in both NF1 alleles. PMID- 11115851 TI - Functional characterization of missense mutations at codon 838 in retinal guanylate cyclase correlates with disease severity in patients with autosomal dominant cone-rod dystrophy. AB - Three different mutations in codon 838 of GUCY2D, the gene for retinal guanylate cyclase 1, have been linked to autosomal dominant cone-rod dystrophy at the CORD6 locus. To examine the relationship between enzyme activity and disease severity, the three disease-causing substitutions (R838C, R838H and R838S) and four artificial mutations (R838A, R838E, R838L and R838K) were generated. Assay of GCAP1-stimulated cyclase activity in vitro shows that, compared with wild-type, R838E, R838L and R838K possess only low activity, whereas R838A, R838C, R838H and R838S have activity equal or superior to wild-type at low Ca(2+) concentrations. These four latter mutants showed a higher apparent affinity for GCAP1 than did wild-type. The Ca(2+) sensitivity of the GCAP1 activation was also altered with marked residual activity at high Ca(2+), the effect increasing: wild-type < R838C < R838H << R838A < R838S. Within the photoreceptor, this would result in a failure to inactivate cyclase activity at high physiological Ca(2+ )concentrations. Amongst the three disease-associated mutations, the effect correlates directly with disease severity. The wild-type and R838H mutant displayed a difference in pH sensitivity, with the mutant showing a higher specific activity with pH > 6.0. Site 838 is in the dimerization domain that forms a coiled-coil in the active protein. A computer-aided structure prediction of this region indicates that R838 in the wild-type breaks the structure at four helical turns, and there is an increasing tendency for the structure to continue for further turns in the order R838C < R838H,S,K << R838E < R838A < R838L. PMID- 11115852 TI - Mutation-dependent aggregation of tau protein and its selective depletion from the soluble fraction in brain of P301L FTDP-17 patients. AB - Mutations in the gene for the microtubule-associated protein tau are associated with frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). In this study we compared the presence of the P301L mutated tau protein from brain material of patients with that of the normal 4-repeat, using polyclonal antibodies specific for the P301L point mutation and its normal counterpart. We determined the relative ratio of mutated versus normal tau protein in the sarkosyl-soluble and -insoluble protein fractions from several brain regions. Although mutated and normal tau proteins are both present in the sarkosyl insoluble deposits, quantitative analysis showed that the mutated protein is the major component. In the sarkosyl-soluble fraction of frontal and temporal cortex the overall ratio of 3-repeat versus 4-repeat tau isoforms is unchanged but there is a dramatic depletion of mutant tau protein. Furthermore, we observed an increase in tau-immunoreactive cleavage products with the P301L antibody, suggesting that the mutant protein is partly resistant to degradation and this is confirmed by pulse-chase experiments. This is the first direct evidence using patient material that shows a selective aggregation of mutant tau protein resulting in sarkosyl-insoluble deposits and the specific depletion of mutated tau protein in the soluble fraction. PMID- 11115853 TI - Neuronal expression of the fukutin gene. AB - Fukuyama-type congenital muscular dystrophy (FCMD), a relatively common autosomal recessive disorder in Japan, is characterized by severe congenital muscular dystrophy in combination with cortical dysgenesis (polymicrogyria). The gene responsible for FCMD encodes a novel protein, fukutin, which is likely to be an extracellular protein. Pathological study of brain tissue from FCMD fetuses revealed frequent breaks in the glia limitans and basement membrane complex. Disruption of the basal lamina in FCMD muscle was also seen. Thus, structural alteration of the basal lamina appears to play a key role in the pathophysiology of FCMD. To investigate the role of fukutin in brain anomalies, we examined fukutin mRNA expression in the human brain. Northern blot and RT-PCR analysis revealed that the fukutin gene is expressed at similar levels in fetal and adult brain, whereas its expression is much reduced in FCMD brains. Tissue in situ hybridization analysis revealed fukutin mRNA expression in the migrating neurons, including Cajar-Retzius cells and adult cortical neurons, as well as in hippocampal pyramidal cells and cerebellar Purkinje cells. However, we observed no expression in the glia limitans, the subpial astrocytes (which contribute to basement membrane formation) or other glial cells. In the FCMD brain, neurons in regions with no dysplasia showed fair expression, whereas transcripts were nearly undetectable in the overmigrated dysplastic region. These observations suggest that fukutin function may influence neuronal migration itself rather than formation of the basement membrane. Furthermore, differences in mRNA levels among neurons in early developmental stages may partially differentiate normal and abnormal regions. PMID- 11115854 TI - A sarcoglycan-dystroglycan complex anchors Dp116 and utrophin in the peripheral nervous system. AB - The dystrophin-associated membrane-integrated protein complex anchors dystrophin in the sarcolemma of striated muscles and is composed of two glycoprotein subcomplexes, the dystroglycan and the sarcoglycan (SG) complexes, and a small membrane protein termed sarcospan (SPN). The SG complex consists of four transmembrane glycoproteins, alpha-SG, beta-SG, gamma-SG and delta-SG. We found that beta-SG and delta-SG were co-expressed with epsilon-SG, a alpha-SG homolog, in the peripheral nerve, but not with alpha-SG or gamma-SG. SPN, which tightly links to the SG complex in the muscle cell membrane, was absent in the peripheral nerve. These peripheral nerve SGs were colocalized at the outermost layer of the myelin sheath of nerve fibers together with the dystroglycan complex, utrophin, and a short dystrophin isoform (Dp116). Immunocytochemical analysis using SG deficient animals showed that a defect in beta- or delta-SG led to a great reduction of all residual SGs, but not of the other proteins, i.e., dystroglycans, Dp116 and utrophin, in the peripheral nerve. This observation suggests that the epsilon-, beta- and delta-SG molecules form a complex behaving as a single unit similar to the SG complex in muscle cells. An immunoprecipitation study indicated that the SG complex is associated with the dystroglycan complex and Dp116 or utrophin. These results demonstrated that Dp116 and utrophin are anchored to a novel membrane protein architecture, which consists of the SG and dystroglycan complexes, but not SPN, in the Schwann cell membrane. PMID- 11115855 TI - Disruption of the mouse Necdin gene results in hypothalamic and behavioral alterations reminiscent of the human Prader-Willi syndrome. AB - Prader-Willi syndrome (PWS) is a complex neurogenetic disorder with considerable clinical variability that is thought in large part to be the result of a hypothalamic defect. PWS results from the absence of paternal expression of imprinted genes localized in the 15q11-q13 region; however, none of the characterized genes has so far been shown to be involved in the etiology of PWS. Here, we provide a detailed investigation of a mouse model deficient for NECDIN: Linked to the mutation, a neonatal lethality of variable penetrance is observed. Viable NECDIN: mutants show a reduction in both oxytocin-producing and luteinizing hormone-releasing hormone (LHRH)-producing neurons in hypothalamus. This represents the first evidence of a hypothalamic deficiency in a mouse model of PWS. NECDIN:-deficient mice also display increased skin scraping activity in the open field test and improved spatial learning and memory in the Morris water maze. The latter features are reminiscent of the skin picking and improved spatial memory that are characteristics of the PWS phenotype. These striking parallels in hypothalamic structure, emotional and cognitive-related behaviors strongly suggest that NECDIN is responsible for at least a subset of the multiple clinical manifestations of PWS. PMID- 11115856 TI - Arabidopsis genome. A milestone in plant biology. PMID- 11115857 TI - The free flow of ideas, information, and materials. PMID- 11115858 TI - Deciphering a weed. Genomic sequencing of Arabidopsis. PMID- 11115859 TI - Bioinformatic resources, challenges, and opportunities using Arabidopsis as a model organism in a post-genomic era. PMID- 11115860 TI - The Arabidopsis knockout facility at the University of Wisconsin-Madison. PMID- 11115861 TI - Arabidopsis microarray service facilities. PMID- 11115862 TI - Large-scale profiling of the Arabidopsis transcriptome. PMID- 11115863 TI - Seed and molecular resources for Arabidopsis. PMID- 11115864 TI - A simple procedure for the analysis of single nucleotide polymorphisms facilitates map-based cloning in Arabidopsis. AB - We developed a modified allele-specific PCR procedure for assaying single nucleotide polymorphisms (SNPs) and used the procedure (called SNAP for single nucleotide amplified polymorphisms) to generate 62 Arabidopsis mapping markers. SNAP primers contain a single base pair mismatch within three nucleotides from the 3' end of one allele (the specific allele) and in addition have a 3' mismatch with the nonspecific allele. A computer program called SNAPER was used to facilitate the design of primers that generate at least a 1,000-fold difference in the quantity of the amplification products from the specific and nonspecific SNP alleles. Because SNAP markers can be readily assayed by electrophoresis on standard agarose gels and because a public database of over 25,000 SNPs is available between the Arabidopsis Columbia and Landsberg erecta ecotypes, the SNAP method greatly facilitates the map-based cloning of Arabidopsis genes defined by a mutant phenotype. PMID- 11115865 TI - New techniques enable comparative analysis of microtubule orientation, wall texture, and growth rate in intact roots of Arabidopsis. AB - This article explores root epidermal cell elongation and its dependence on two structural elements of cells, cortical microtubules and cellulose microfibrils. The recent identification of Arabidopsis morphology mutants with putative cell wall or cytoskeletal defects demands a procedure for examining and comparing wall architecture and microtubule organization patterns in this species. We developed methods to examine cellulose microfibrils by field emission scanning electron microscopy and microtubules by immunofluorescence in essentially intact roots. We were able to compare cellulose microfibril and microtubule alignment patterns at equivalent stages of cell expansion. Field emission scanning electron microscopy revealed that Arabidopsis root epidermal cells have typical dicot primary cell wall structure with prominent transverse cellulose microfibrils embedded in pectic substances. Our analysis showed that microtubules and microfibrils have similar orientation only during the initial phase of elongation growth. Microtubule patterns deviate from a predominantly transverse orientation while cells are still expanding, whereas cellulose microfibrils remain transverse until well after expansion finishes. We also observed microtubule-microfibril alignment discord before cells enter their elongation phase. This study and the new technology it presents provide a starting point for further investigations on the physical properties of cell walls and their mechanisms of assembly. PMID- 11115866 TI - A novel link between ran signal transduction and nuclear envelope proteins in plants. PMID- 11115867 TI - Glutamate-gated calcium fluxes in Arabidopsis. PMID- 11115868 TI - Glycoside hydrolases and glycosyltransferases. Families, modules, and implications for genomics. PMID- 11115869 TI - The roles of photoreceptor systems and the COP1-targeted destabilization of HY5 in light control of Arabidopsis seedling development. PMID- 11115870 TI - Constructing a plant cell. The genetic control of root hair development. PMID- 11115871 TI - Plant sugar-response pathways. Part of a complex regulatory web. PMID- 11115872 TI - Arabidopsis in planta transformation. Uses, mechanisms, and prospects for transformation of other species. PMID- 11115873 TI - The cell biology of the COP/DET/FUS proteins. Regulating proteolysis in photomorphogenesis and beyond? PMID- 11115875 TI - Microarray analysis of developing Arabidopsis seeds. AB - To provide a broad analysis of gene expression in developing Arabidopsis seeds, microarrays have been produced that display approximately 2,600 seed-expressed genes. DNA for genes spotted on the arrays were selected from >10,000 clones partially sequenced from a cDNA library of developing seeds. Based on a series of controls, sensitivity of the arrays was estimated at one to two copies of mRNA per cell and cross hybridization was estimated to occur if closely related genes have >70% to 80% sequence identity. These arrays have been hybridized in a series of experiments with probes derived from seeds, leaves, and roots of Arabidopsis. Analysis of expression ratios between the different tissues has allowed the tissue-specific expression patterns of many hundreds of genes to be described for the first time. Approximately 25% of the 2, 600 genes were expressed at ratios > or =2-fold higher in seeds than leaves or roots and 10% at ratios > or =10. Included in this list are a large number of proteins of unknown function, and potential regulatory factors such as protein kinases, phosphatases, and transcription factors. The Arabidopsis arrays were also found to be useful for transcriptional profiling of mRNA isolated from developing oilseed rape (Brassica napus) seeds and expression patterns correlated well between the two species. PMID- 11115874 TI - The Arabidopsis genome. An abundance of soluble N-ethylmaleimide-sensitive factor adaptor protein receptors. AB - Many factors have been characterized as essential for vesicle trafficking, including a number of proteins commonly referred to as soluble N-ethylmaleimide sensitive factor adaptor protein receptor (SNARE) components. The Arabidopsis genome contains a remarkable number of SNAREs. In general, the vesicle fusion machinery appears highly conserved. However, whereas some classes of yeast and mammalian genes appear to be lacking in Arabidopsis, this small plant genome has gene families not found in other eukaryotes. Very little is known about the precise function of plant SNAREs. By contrast, the intracellular localization of and interactions between a large number of plant SNAREs have been determined, and these data are discussed in light of the phylogenetic analysis. PMID- 11115877 TI - Genetic analysis of seed-soluble oligosaccharides in relation to seed storability of Arabidopsis. AB - Seed oligosaccharides (OSs) and especially raffinose series OSs (RSOs) are hypothesized to play an important role in the acquisition of desiccation tolerance and consequently in seed storability. In the present work we analyzed the seed-soluble OS (sucrose, raffinose, and stachyose) content of several Arabidopsis accessions and thus identified the genotype Cape Verde Islands having a very low RSO content. By performing quantitative trait loci (QTL) mapping in a recombinant inbred line population, we found one major QTL responsible for the practically monogenic segregation of seed stachyose content. This locus also affected the content of the two other OSs, sucrose, and raffinose. Two candidate genes encoding respectively for galactinol synthase and raffinose synthase were located within the genomic region around this major QTL. In addition, three smaller-effect QTL were identified, each one specifically affecting the content of an individual OS. Seed storability was analyzed in the same recombinant inbred line population by measuring viability (germination) under two different seed aging assays: after natural aging during 4 years of dry storage at room temperature and after artificial aging induced by a controlled deterioration test. Thus, four QTL responsible for the variation of this trait were mapped. Comparison of the QTL genetic positions showed that the genomic region containing the major OS locus did not significantly affect the seed storability. We concluded that in the studied material neither RSOs nor sucrose content had a specific effect on seed storability. PMID- 11115876 TI - A new set of Arabidopsis expressed sequence tags from developing seeds. The metabolic pathway from carbohydrates to seed oil. AB - Large-scale single-pass sequencing of cDNAs from different plants has provided an extensive reservoir for the cloning of genes, the evaluation of tissue-specific gene expression, markers for map-based cloning, and the annotation of genomic sequences. Although as of January 2000 GenBank contained over 220,000 entries of expressed sequence tags (ESTs) from plants, most publicly available plant ESTs are derived from vegetative tissues and relatively few ESTs are specifically derived from developing seeds. However, important morphogenetic processes are exclusively associated with seed and embryo development and the metabolism of seeds is tailored toward the accumulation of economically valuable storage compounds such as oil. Here we describe a new set of ESTs from Arabidopsis, which has been derived from 5- to 13-d-old immature seeds. Close to 28,000 cDNAs have been screened by DNA/DNA hybridization and approximately 10,500 new Arabidopsis ESTs have been generated and analyzed using different bioinformatics tools. Approximately 40% of the ESTs currently have no match in dbEST, suggesting many represent mRNAs derived from genes that are specifically expressed in seeds. Although these data can be mined with many different biological questions in mind, this study emphasizes the import of photosynthate into developing embryos, its conversion into seed oil, and the regulation of this pathway. PMID- 11115878 TI - Arabidopsis species hybrids in the study of species differences and evolution of amphiploidy in plants. AB - It is estimated that 5 million years of evolution separate Arabidopsis thaliana from its close relative Arabidopsis lyrata. The two taxa differ by many characteristics, and together they exemplify the differentiation of angiosperms into self-fertilizing and cross-fertilizing species as well as annual and perennial species. Despite their disparate life histories, the two species can be crossed to produce viable and vigorous hybrids exhibiting heterotic effects. Although pollen sterile, the hybrids produce viable ovules and were used as female parent in backcrosses to both parental species. The resulting backcross plants exhibited transgressive variation for a number of interesting developmental and growth traits as well as negative nuclear/cytoplasmic interactions. Moreover, the genesis of a fertile amphidiploid neospecies, apparently by spontaneous somatic doubling in an interspecific hybrid, was observed in the laboratory. The mechanisms responsible for the generation of amphiploids and the subsequent evolution of amphiploid genomes can now be studied through direct observation using the large arsenal of molecular tools available for Arabidopsis. PMID- 11115879 TI - Arabidopsis mutants resistant to S(+)-beta-methyl-alpha, beta-diaminopropionic acid, a cycad-derived glutamate receptor agonist. AB - Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that are the predominant neuroreceptors in the mammalian brain. Genes with high sequence similarity to animal iGluRs have been identified in Arabidopsis. To understand the role of Arabidopsis glutamate receptor-like (AtGLR) genes in plants, we have taken a pharmacological approach by examining the effects of BMAA [S(+)-beta methyl-alpha, beta-diaminopropionic acid], a cycad-derived iGluR agonist, on Arabidopsis morphogenesis. When applied to Arabidopsis seedlings, BMAA caused a 2 to 3-fold increase in hypocotyl elongation and inhibited cotyledon opening during early seedling development. The effect of BMAA on hypocotyl elongation is light specific. Furthermore, BMAA effects on early morphogenesis of Arabidopsis can be reversed by the simultaneous application of glutamate, the native iGluR agonist in animals. To determine the targets of BMAA action in Arabidopsis, a genetic screen was devised to isolate Arabidopsis mutants with a BMAA insensitive morphology (bim). When grown in the light on BMAA, bim mutants exhibited short hypocotyls compared with wild type. bim mutants were grouped into three classes based on their morphology when grown in the dark in the absence of BMAA. Class-I bim mutants have a normal, etiolated morphology, similar to wild-type plants. Class-II bim mutants have shorter hypocotyls and closed cotyledons when grown in the dark. Class-III bim mutants have short hypocotyls and open cotyledons when grown in the dark, resembling the previously characterized constitutively photomorphogenic mutants (cop, det, fus, and shy). Further analysis of the bim mutants should help define whether plant-derived iGluR agonists target glutamate receptor signaling pathways in plants. PMID- 11115880 TI - Arabidopsis RopGAPs are a novel family of rho GTPase-activating proteins that require the Cdc42/Rac-interactive binding motif for rop-specific GTPase stimulation. AB - The plant-specific Rop subfamily of Rho GTPases, most closely related to the mammalian Cdc42 and Rac GTPases, plays an important role in the regulation of calcium-dependent pollen tube growth, H(2)O(2)-mediated cell death, and many other processes in plants. In a search for Rop interactors using the two-hybrid method, we identified a family of Rho GTPase-activating proteins (GAP) from Arabidopsis, termed RopGAPs. In addition to a GAP catalytic domain, RopGAPs contain a Cdc42/Rac-interactive binding (CRIB) motif known to allow Cdc42/Rac effector proteins to bind activated Cdc42/Rac. This novel combination of a GAP domain with a CRIB motif is widespread in higher plants and is unique to the regulation of the Rop GTPase. A critical role for CRIB in the regulation of in vitro RopGAP activity was demonstrated using point and deletion mutations. Both types of mutants have drastically reduced capacities to stimulate the intrinsic Rop GTPase activity and to bind Rop. Furthermore, RopGAPs preferentially stimulate the GTPase activity of Rop, but not Cdc42 in a CRIB-dependent manner. In vitro binding assays show that the RopGAP CRIB domain interacts with GTP- and GDP-bound forms of Rop, as well as the transitional state of Rop mimicked by aluminum fluoride. The CRIB domain also promotes the association of the GAP domain with the GDP-bound Rop, as does aluminum fluoride. These results reveal a novel CRIB-dependent mechanism for the regulation of the plant-specific family of Rho GAPs. We propose that the CRIB domain facilitates the formation of or enhanced GAP-mediated stabilization of the transitional state of the Rop GTPase. PMID- 11115881 TI - Profilin plays a role in cell elongation, cell shape maintenance, and flowering in Arabidopsis. AB - Profilin (PFN) is an ubiquitous, low-M(r), actin-binding protein involved in the organization of the cytoskeleton of eukaryotes including higher plants. PFNs are encoded by a multigene family in Arabidopsis. We have analyzed in vivo functions of Arabidopsis PFN by generating transgenic plants carrying a 35S-PFN-1 or 35S antisense PFN-1 transgene. Etiolated seedlings underexpressing PFN (PFN-U) displayed an overall dwarf phenotype with short hypocotyls whose lengths were 20% to 25% that of wild type (WT) at low temperatures. Light-grown PFN-U plants were smaller in stature and flowered early. Compared with equivalent cells in WT, most cells in PFN-U hypocotyls and roots were shorter, but more isodiametric, and microscopic observations of etiolated PFN-U hypocotyls revealed a rough epidermal surface. In contrast, light-grown seedlings overexpressing PFN had longer roots and root hair although etiolated seedlings overexpressing PFN were either the same size or slightly longer than WT seedlings. Transgenic seedlings harboring a PFN-1-GUS transgene directed expression in root and root hair and in a ring of cells at the elongating zone of the root tip. As the seedlings matured PFN-1-GUS was mainly expressed in the vascular bundles of cotyledons and leaves. Our results show that Arabidopsis PFNs play a role in cell elongation, cell shape maintenance, polarized growth of root hair, and unexpectedly, in determination of flowering time. PMID- 11115882 TI - Pericycle cell proliferation and lateral root initiation in Arabidopsis. AB - In contrast with other cells generated by the root apical meristem in Arabidopsis, pericycle cells adjacent to the protoxylem poles of the vascular cylinder continue to cycle without interruption during passage through the elongation and differentiation zones. However, only some of the dividing pericycle cells are committed to the asymmetric, formative divisions that give rise to lateral root primordia (LRPs). This was demonstrated by direct observation and mapping of mitotic figures, cell-length measurements, and the histochemical analysis of a cyclin-GUS fusion protein in pericycle cells. The estimated duration of a pericycle cell cycle in the root apical meristem was similar to the interval between cell displacement from the meristem and the initiation of LRP formation. Developmentally controlled LRP initiation occurs early, 3 to 8 mm from the root tip. Thus the first growth control point in lateral root formation is defined by the initiation of primordia in stochastic patterns by cells passing through the elongation and young differentiation zones, up to where lateral roots begin to emerge from the primary root. Therefore, the first growth control point is not restricted to a narrow developmental window. We propose that late LRP initiation is developmentally unrelated to the root apical meristem and is operated by a second growth control point that can be activated by environmental cues. The observation that pericycle cells divide and lateral root primordia form without intervening mitotic quiescence suggests that lateral organ formation in roots and shoots might not be as fundamentally different as previously thought. PMID- 11115883 TI - An enhancer trap line associated with a D-class cyclin gene in Arabidopsis. AB - In yeast and animals, cyclins have been demonstrated to be important regulators of cell cycle progression. In recent years, a large number of A-, B-, and D-class cyclins have been isolated from a variety of plant species. One class of cyclins, the D-class cyclins, is important for progression through G1 phase of the cell cycle. In Arabidopsis, four D-class cyclins have been isolated and characterized (CYCLIN-D1;1, CYCLIN-D2;1, CYCLIN-D3;1, and CYCLIN-D4;1). In this report we describe the characterization of a fifth D-class cyclin gene, CYCLIN-D3;2 (CYCD3;2), from Arabidopsis. An enhancer trap line, line 5580, contains a T-DNA insertion in CYCD3;2. Enhancer trap line 5580 exhibits expression in young vegetative and floral primordia. In line 5580, T-DNA is inserted in the first exon of the CYCD3;2 gene; in homozygous 5580 plants CYCD3;2 RNA is not detectable. Even though CYCD3;2 gene function is eliminated, homozygous 5580 plants do not exhibit an obvious growth or developmental phenotype. Via in situ hybridization we demonstrate that CYCD3;2 RNA is expressed in developing vegetative and floral primordia. In addition, CYCD3;2 is also capable of rescuing a yeast strain that is deficient in G1 cyclin activity. PMID- 11115884 TI - Chloroplast division and morphology are differentially affected by overexpression of FtsZ1 and FtsZ2 genes in Arabidopsis. AB - In higher plants, two nuclear gene families, FtsZ1 and FtsZ2, encode homologs of the bacterial protein FtsZ, a key component of the prokaryotic cell division machinery. We previously demonstrated that members of both gene families are essential for plastid division, but are functionally distinct. To further explore differences between FtsZ1 and FtsZ2 proteins we investigated the phenotypes of transgenic plants overexpressing AtFtsZ1-1 or AtFtsZ2-1, Arabidopsis members of the FtsZ1 and FtsZ2 families, respectively. Increasing the level of AtFtsZ1-1 protein as little as 3-fold inhibited chloroplast division. Plants with the most severe plastid division defects had 13- to 26-fold increases in AtFtsZ1-1 levels over wild type, and some of these also exhibited a novel chloroplast morphology. Quantitative immunoblotting revealed a correlation between the degree of plastid division inhibition and the extent to which the AtFtsZ1-1 protein level was elevated. In contrast, expression of an AtFtsZ2-1 sense transgene had no obvious effect on plastid division or morphology, though AtFtsZ2-1 protein levels were elevated only slightly over wild-type levels. This may indicate that AtFtsZ2-1 accumulation is more tightly regulated than that of AtFtsZ1-1. Plants expressing the AtFtsZ2-1 transgene did accumulate a form of the protein smaller than those detected in wild-type plants. AtFtsZ2-1 levels were unaffected by increased or decreased accumulation of AtFtsZ1-1 and vice versa, suggesting that the levels of these two plastid division proteins are regulated independently. Taken together, our results provide additional evidence for the functional divergence of the FtsZ1 and FtsZ2 plant gene families. PMID- 11115885 TI - Regulation of etioplast pigment-protein complexes, inner membrane architecture, and protochlorophyllide a chemical heterogeneity by light-dependent NADPH:protochlorophyllide oxidoreductases A and B. AB - The etioplast of dark-grown angiosperms is characterized by the prolamellar body (PLB) inner membrane, the absence of chlorophyll, and the accumulation of divinyl and monovinyl derivatives of protochlorophyll(ide) a [Pchl(ide) a]. Either of two structurally related, but differentially expressed light-dependent NADPH:Pchlide oxidoreductases (PORs), PORA and PORB, can assemble the PLB and form dark-stable ternary complexes containing enzymatically photoactive Pchlide-F655. Here we have examined in detail whether these polypeptides play redundant roles in etioplast differentiation by manipulating the total POR content and the PORA-to-PORB ratio of etiolated Arabidopsis seedlings using antisense and overexpression approaches. POR content correlates closely with PLB formation, the amounts, spectroscopic properties, and photoreduction kinetics of photoactive Pchlide, the ratio of photoactive Pchlide-F655 to non-photoactive Pchl(ide)-F632, and the ratio of divinyl- to monovinyl-Pchl(ide). This last result defines POR as the first endogenous protein factor demonstrated to influence the chemical heterogeneity of Pchl(ide) in angiosperms. It is intriguing that excitation energy transfer between different spectroscopic forms of Pchl(ide) in etiolated cotyledons remains largely independent of POR content. We therefore propose that the PLB contains a minimal structural unit with defined pigment stoichiometries, within which a small amount of non-photoactive Pchl(ide) transfers excitation energy to a large excess of photoactive Pchlide-F655. In addition, our data suggests that POR may bind not only stoichiometric amounts of photoactive Pchlide, but also substoichiometric amounts of non-photoactive Pchl(ide). We conclude that the typical characteristics of etioplasts are closely related to total POR content, but not obviously to the specific presence of PORA or PORB. PMID- 11115886 TI - Trienoic fatty acids are required to maintain chloroplast function at low temperatures. AB - The chloroplast membranes of all higher plants contain very high proportions of trienoic fatty acids. To investigate how these lipid structures are important in photosynthesis, we have generated a triple mutant line of Arabidopsis that contains negligible levels of trienoic fatty acids. For mutant plants grown at 22 degrees C, photosynthetic fluorescence parameters were indistinguishable from wild type at 25 degrees C. Lowering the measurement temperature led to a small decrease in photosynthetic quantum yield, Phi(II), in the mutant relative to wild type controls. These and other results indicate that low temperature has only a small effect on photosynthesis in the short term. However, long-term growth of plants at 4 degrees C resulted in decreases in fluorescence parameters, chlorophyll content, and thylakoid membrane content in triple-mutant plants relative to wild type. Comparisons among different mutant lines indicated that these detrimental effects of growth at 4 degrees C are strongly correlated with trienoic fatty acid content with levels of 16:3 + 18:3, approximately one-third of wild type being sufficient to sustain normal photosynthetic function. In total, our results indicate that trienoic fatty acids are important to ensure the correct biogenesis and maintenance of chloroplasts during growth of plants at low temperatures. PMID- 11115887 TI - Characterization of the response of the Arabidopsis response regulator gene family to cytokinin. AB - We examined the expression of a family of Arabidopsis response regulators (ARR) and found that the steady-state levels of RNA for most are elevated very rapidly by cytokinin. Using nuclear run-on assays we demonstrated that this increase in ARR transcript levels in response to cytokinin is due, at least in part, to increased transcription. The start site of transcription for the ARR5 gene was identified using primer extension analysis. A DNA fragment comprised of 1.6 kb upstream of the ARR5 transcript start site conferred cytokinin-inducible gene expression when fused to a beta-glucuronidase reporter, confirming that the transcription rate of ARR5 is elevated by cytokinin. This reporter construct was also used to examine the spatial pattern of ARR5 expression. The highest levels of expression were observed in the root and shoot apical meristems, at the junction of the pedicle and the silique, and in the central portion of mature roots. The expression of ARR5 in the apical meristems was confirmed by whole mount in situ analysis of seedlings and is consistent with a role for cytokinin in regulating cell division in vivo. PMID- 11115888 TI - Stunted plant 1 mediates effects of cytokinin, but not of auxin, on cell division and expansion in the root of Arabidopsis. AB - Plants control organ growth rate by adjusting the rate and duration of cell division and expansion. Surprisingly, there have been few studies where both parameters have been measured in the same material, and thus we have little understanding of how division and expansion are regulated interdependently. We have investigated this regulation in the root meristem of the stunted plant 1 (stp1) mutation of Arabidopsis, the roots of which elongate more slowly than those of the wild type and fail to accelerate. We used a kinematic method to quantify the spatial distribution of the rate and extent of cell division and expansion, and we compared stp1 with wild type and with wild type treated with exogenous cytokinin (1 microM zeatin) or auxin (30 nM 2,4-dichlorophenoxyacetic acid). All treatments reduced average cell division rates, which reduced cell production by the meristem. Auxin lowered root elongation by narrowing the elongation zone and reducing the time spent by a cell in this zone, but did not decrease maximal strain rate. In addition, auxin increased the length of the meristem. In contrast, cytokinin reduced root elongation by lowering maximal strain rate, but did not change the time spent by a cell within the elongation zone; also, cytokinin blocked the increase in length and cell number of the meristem and elongation zone. The cytokinin-treated wild type phenocopied stp1 in nearly every detail, supporting the hypothesis that cytokinin affects root growth via STP1. The opposite effects of auxin and cytokinin suggest that the balance of these hormones may control the size of the meristem. PMID- 11115889 TI - Aux/IAA proteins are phosphorylated by phytochrome in vitro. AB - Auxin/indole-3-acetic acid (Aux/IAA) genes encode short-lived transcription factors that are induced as a primary response to the plant growth hormone IAA or auxin. Gain-of-function mutations in Arabidopsis genes, SHY2/IAA3, AXR3/IAA17, and AXR2/IAA7 cause pleiotropic phenotypes consistent with enhanced auxin responses, possibly by increasing Aux/IAA protein stability. Semidominant mutations shy2-1D, shy2-2, axr3-1, and axr2-1 induce ectopic light responses in dark-grown seedlings. Because genetic studies suggest that the shy2-1D and shy2-2 mutations bypass phytochrome requirement for certain aspects of photomorphogenesis, we tested whether SHY2/IAA3 and related Aux/IAA proteins interact directly with phytochrome and whether they are substrates for its protein kinase activity. Here we show that recombinant Aux/IAA proteins from Arabidopsis and pea (Pisum sativum) interact in vitro with recombinant phytochrome A from oat (Avena sativa). We further show that recombinant SHY2/IAA3, AXR3/IAA17, IAA1, IAA9, and Ps-IAA4 are phosphorylated by recombinant oat phytochrome A in vitro. Deletion analysis of Ps-IAA4 indicates that phytochrome A phosphorylation occurs on the N-terminal half of the protein. Metabolic labeling and immunoprecipitation studies with affinity-purified antibodies to IAA3 demonstrate increased in vivo steady-state levels of mutant IAA3 in shy2-2 plants and phosphorylation of the SHY2-2 protein in vivo. Phytochrome-dependent phosphorylation of Aux/IAA proteins is proposed to provide one molecular mechanism for integrating auxin and light signaling in plant development. PMID- 11115890 TI - The rib1 mutant is resistant to indole-3-butyric acid, an endogenous auxin in Arabidopsis. AB - The presence of indole-3-butyric acid (IBA) as an endogenous auxin in Arabidopsis has been recently demonstrated. However, the in vivo role of IBA remains to be elucidated. We present the characterization of a semi-dominant mutant that is affected in its response to IBA, but shows a wild-type response to indole-3 acetic acid (IAA), the predominant and most studied form of auxin. We have named this mutant rib1 for resistant to IBA. Root elongation assays show that rib1 is specifically resistant to IBA, to the synthetic auxin 2,4-dichlorophenoxyacetic acid, and to auxin transport inhibitors. rib1 does not display increased resistance to IAA, to the synthetic auxin naphthalene acetic acid, or to other classes of plant hormones. rib1 individuals also have other root specific phenotypes including a shortened primary root, an increased number of lateral roots, and a more variable response than wild type to a change in gravitational vector. Adult rib1 plants are morphologically indistinguishable from wild-type plants. These phenotypes suggest that rib1 alters IBA activity in the root, thereby affecting root development and response to environmental stimuli. We propose models in which RIB1 has a function in either IBA transport or response. Our experiments also suggest that IBA does not use the same mechanism to exit cells as does IAA and we propose a model for IBA transport. PMID- 11115892 TI - Pathogenesis of the human opportunistic pathogen Pseudomonas aeruginosa PA14 in Arabidopsis. AB - The human opportunistic pathogen Pseudomonas aeruginosa strain PA14 is a multihost pathogen that can infect Arabidopsis. We found that PA14 pathogenesis in Arabidopsis involves the following steps: attachment to the leaf surface, congregation of bacteria at and invasion through stomata or wounds, colonization of intercellular spaces, and concomitant disruption of plant cell wall and membrane structures, basipetal movement along the vascular parenchyma, and maceration and rotting of the petiole and central bud. Distinctive features of P. aeruginosa pathogenesis are that the surface of mesophyll cell walls adopt an unusual convoluted or undulated appearance, that PA14 cells orient themselves perpendicularly to the outer surface of mesophyll cell walls, and that PA14 cells make circular perforations, approximately equal to the diameter of P. aeruginosa, in mesophyll cell walls. Taken together, our data show that P. aeruginosa strain PA14 is a facultative pathogen of Arabidopsis that is capable of causing local and systemic infection, which can result in the death of the infected plant. PMID- 11115891 TI - Regulation and function of the Arabidopsis ABA-insensitive4 gene in seed and abscisic acid response signaling networks. AB - We have characterized developmental, environmental, and genetic regulation of abscisic acid-insensitive (ABI)4 gene expression in Arabidopsis. Although expressed most strongly in seeds, ABI4 transcripts are also present at low levels in vegetative tissue; vegetative expression is not induced by abscisic acid (ABA) or stress treatments. Comparison of transcript levels in mature seeds of ABA insensitive, ABA-hypersensitive, ABA-deficient, or heterochronic mutants indicates that ABI4 expression is altered in only two of the backgrounds, the ABA insensitive mutants abi1-1 and abi3-1. To determine whether ABI4 is necessary and/or sufficient for ABA response, we assayed the effects of loss of ABI4 function and ectopic ABI4 expression on growth and gene expression. We examined genetic interactions among three ABA response loci, ABI3, ABI4, and ABI5, by comparing phenotypes of mutants, ectopic expression lines, mutants carrying an ectopically expressed transgene, and the corresponding wild-type lines. Our results indicate some cross-regulation of expression among ABI3, ABI4, and ABI5 and suggest that they function in a combinatorial network, rather than a regulatory hierarchy, controlling seed development and ABA response. PMID- 11115893 TI - Adenosine kinase of Arabidopsis. Kinetic properties and gene expression. AB - To assess the functional significance of adenosine salvage in plants, the cDNAs and genes encoding two isoforms of adenosine kinase (ADK) were isolated from Arabidopsis. The ADK1- and ADK2-coding sequences are very similar, sharing 92% and 89% amino acid and nucleotide identity, respectively. Each cDNA was overexpressed in Escherichia coli, and the catalytic activity of each isoform was determined. Both ADKs had similar catalytic properties with a K(m) and V(max)/K(m) for adenosine of 0.3 to 0.5 microM and 5.4 to 22 L min(-1) mg(-1) protein, respectively. The K(m) and V(max)/K(m) for the cytokinin riboside N(6)(isopentenyl) adenosine are 3 to 5 microM and 0.021 to 0.14 L min(-1) mg(-1) protein, respectively, suggesting that adenosine is the preferred substrate for both ADK isoforms. In Arabidopsis plants, both ADK genes are expressed constitutively, with the highest steady-state mRNA levels being found in stem and root. ADK1 transcript levels were generally higher than those of ADK2. ADK enzyme activity reflected relative ADK protein levels seen in immunoblots for leaves, flowers, and stems but only poorly so for roots, siliques, and dry seeds. The catalytic properties, tissue accumulation, and expression levels of these ADKs suggest that they play a key metabolic role in the salvage synthesis of adenylates and methyl recycling in Arabidopsis. They may also contribute to cytokinin interconversion. PMID- 11115894 TI - Analysis of phosphate acquisition efficiency in different Arabidopsis accessions. AB - The morphological and physiological characteristics of Arabidopsis accessions differing in their phosphate acquisition efficiencies (PAEs) when grown on a sparingly soluble phosphate source (hydroxylapatite) were analyzed. A set of 36 accessions was subjected to an initial PAE evaluation following cultivation on synthetic, agarose-solidified media containing potassium phosphate (soluble) or hydroxylapatite (sparingly soluble). From the five most divergent accessions identified in this way, C24, Co, and Cal exhibited high PAEs, whereas Col-0 and Te exhibited low PAEs. These five accessions were analyzed in detail. Significant differences were found in root morphology, phosphate uptake kinetics, organic acid release, rhizosphere acidification, and the ability of roots to penetrate substrates. Long root hairs at high densities, high uptake per unit root length, and high substrate penetration ability in the efficient accessions C24 and Co mediate their high PAEs. The third accession with high PAE, Cal, exhibits a high shoot-to-root ratio, long roots with long root hairs, and rhizosphere acidification. These results are consistent with previous observations and highlight the suitability of using Arabidopsis accessions to identify and isolate genes determining the PAE in plants. PMID- 11115895 TI - The isolation and characterization in yeast of a gene for Arabidopsis S adenosylmethionine:phospho-ethanolamine N-methyltransferase. AB - Saccharomyces cerevisiae opi3 mutant strains do not have the phospholipid N methyltransferase that catalyzes the two terminal methylations in the phosphatidylcholine (PC) biosynthetic pathway. This results in a build up of the intermediate phosphatidylmonomethylethanolamine, causing a temperature-sensitive growth phenotype. An Arabidopsis cDNA library was used to isolate three overlapping plasmids that complemented the temperature-sensitive phenotype. Phospholipid analysis showed that the presence of the cloned cDNA caused a 65 fold reduction in the level of phosphatidylmonomethylethanolamine and a significant, though not equivalent, increase in the production of PC. Sequence analysis established that the cDNA was not homologous to OPI3 or to CHO2, the only other yeast phospholipid N-methyltransferase, but was similar to several other classes of methyltransferases. S-adenosyl-Met:phospho-base N methyltransferase assays revealed that the cDNA catalyzed the three sequential methylations of phospho-ethanolamine to form phospho-choline. Phospho-choline is converted to PC by the CDP-choline pathway, explaining the phenotype conferred upon the yeast mutant strain by the cDNA. In accordance with this the gene has been named AtNMT1. The identification of this enzyme and the failure to isolate a plant phospholipid N-methyltransferase suggests that there are fundamental differences between the pathways utilized by yeast and by some plants for synthesis of PC. PMID- 11115896 TI - The ACA4 gene of Arabidopsis encodes a vacuolar membrane calcium pump that improves salt tolerance in yeast. AB - Several lines of evidence suggest that regulation of intracellular Ca(2+) levels is crucial for adaptation of plants to environmental stress. We have cloned and characterized Arabidopsis auto-inhibited Ca(2+)-ATPase, isoform 4 (ACA4), a calmodulin-regulated Ca(2+)-ATPase. Confocal laser scanning data of a green fluorescent protein-tagged version of ACA4 as well as western-blot analysis of microsomal fractions obtained from two-phase partitioning and Suc density gradient centrifugation suggest that ACA4 is localized to small vacuoles. The N terminus of ACA4 contains an auto-inhibitory domain with a binding site for calmodulin as demonstrated through calmodulin-binding studies and complementation experiments using the calcium transport yeast mutant K616. ACA4 and PMC1, the yeast vacuolar Ca(2+)-ATPase, conferred protection against osmotic stress such as high NaCl, KCl, and mannitol when expressed in the K616 strain. An N-terminally modified form of ACA4 specifically conferred increased NaCl tolerance, whereas full-length ATPase had less effect. PMID- 11115897 TI - The ubiquitin-specific protease family from Arabidopsis. AtUBP1 and 2 are required for the resistance to the amino acid analog canavanine. AB - Ubiquitin-specific proteases (UBPs) are a family of unique hydrolases that specifically remove polypeptides covalently linked via peptide or isopeptide bonds to the C-terminal glycine of ubiquitin. UBPs help regulate the ubiquitin/26S proteolytic pathway by generating free ubiquitin monomers from their initial translational products, recycling ubiquitins during the breakdown of ubiquitin-protein conjugates, and/or by removing ubiquitin from specific targets and thus presumably preventing target degradation. Here, we describe a family of 27 UBP genes from Arabidopsis that contain both the conserved cysteine (Cys) and histidine boxes essential for catalysis. They can be clustered into 14 subfamilies based on sequence similarity, genomic organization, and alignments with their closest relatives from other organisms, with seven subfamilies having two or more members. Recombinant AtUBP2 functions as a bona fide UBP: It can release polypeptides attached to ubiquitins via either alpha- or epsilon-amino linkages by an activity that requires the predicted active-site Cys within the Cys box. From the analysis of T-DNA insertion mutants, we demonstrate that the AtUBP1 and 2 subfamily helps confer resistance to the arginine analog canavanine. This phenotype suggests that the AtUBP1 and 2 enzymes are needed for abnormal protein turnover in Arabidopsis. PMID- 11115898 TI - Interaction specificity of Arabidopsis calcineurin B-like calcium sensors and their target kinases. AB - Calcium is a critical component in a number of plant signal transduction pathways. A new family of calcium sensors called calcineurin B-like proteins (AtCBLs) have been recently identified from Arabidopsis. These calcium sensors have been shown to interact with a family of protein kinases (CIPKs). Here we report that each individual member of AtCBL family specifically interacts with a subset of CIPKs and present structural basis for the interaction and for the specificity underlying these interactions. Although the C-terminal region of CIPKs is responsible for interaction with AtCBLs, the N-terminal region of CIPKs is also involved in determining the specificity of such interaction. We have also shown that all three EF-hand motifs in AtCBL members are required for the interaction with CIPKs. Several AtCBL members failed to interact with any of the CIPKs presented in this study, suggesting that these AtCBL members either have other CIPKs as targets or they target distinct proteins other than CIPKs. These results may provide structural basis for the functional specificity of CBL family of calcium sensors and their targets. PMID- 11115899 TI - Overexpression of the Arabidopsis CBF3 transcriptional activator mimics multiple biochemical changes associated with cold acclimation. AB - We further investigated the role of the Arabidopsis CBF regulatory genes in cold acclimation, the process whereby certain plants increase in freezing tolerance upon exposure to low temperature. The CBF genes, which are rapidly induced in response to low temperature, encode transcriptional activators that control the expression of genes containing the C-repeat/dehydration responsive element DNA regulatory element in their promoters. Constitutive expression of either CBF1 or CBF3 (also known as DREB1b and DREB1a, respectively) in transgenic Arabidopsis plants has been shown to induce the expression of target COR (cold-regulated) genes and to enhance freezing tolerance in nonacclimated plants. Here we demonstrate that overexpression of CBF3 in Arabidopsis also increases the freezing tolerance of cold-acclimated plants. Moreover, we show that it results in multiple biochemical changes associated with cold acclimation: CBF3-expressing plants had elevated levels of proline (Pro) and total soluble sugars, including sucrose, raffinose, glucose, and fructose. Plants overexpressing CBF3 also had elevated P5CS transcript levels suggesting that the increase in Pro levels resulted, at least in part, from increased expression of the key Pro biosynthetic enzyme Delta(1)-pyrroline-5-carboxylate synthase. These results lead us to propose that CBF3 integrates the activation of multiple components of the cold acclimation response. PMID- 11115900 TI - Decision analysis of prophylactic treatment for patients with high-risk esophageal varices. AB - BACKGROUND: Clinical decision analyses were conducted to quantify the uncertainty and to identify important factors in selection of prophylactic therapy for patients with esophageal varices. METHODS: A Markov model compared variceal ligation, beta-blockers, and "watchful waiting" strategies in terms of bleeding free life years. Transition probabilities were obtained from meta-analyses of published data. A hypothetical 50-year-old white man with high-risk esophageal varices and cirrhosis served as the prototypical baseline case. Traditional n-way sensitivity analyses were applied to clarify the influence of each factor, and Monte Carlo probabilistic sensitivity analyses were used to investigate clinical uncertainty. RESULTS: Probabilistic sensitivity analyses demonstrated that 77.0% of hypothetical cases had more bleeding-free life years after variceal ligation, whereas 23% had more when treated with beta-blockers. On the basis of one-way sensitivity analyses, only 2 factors (variceal bleeding rates after ligation and treatment with beta-blockers) influenced the strategy choice. CONCLUSIONS: Variceal ligation is an effective prophylactic therapy in many cases, but nearly one quarter of patients with high-risk esophageal varices and cirrhosis may benefit more from prophylactic treatment with beta-blockers. Additional clinical studies identifying key variceal bleeding risk factors may lead to more effective clinical decision making for these patients. PMID- 11115901 TI - CT or EUS for the initial staging of esophageal cancer? A cost minimization analysis. AB - BACKGROUND: Patients with advanced (T4 and/or M1) esophageal cancer are offered palliative therapy. Computed tomography (CT) is sensitive for distant metastases but is less sensitive than endosonography for T4 disease and celiac lymphadenopathy. The aim of this study was to determine whether initial CT or endosonography costs less to diagnose advanced esophageal cancer. METHODS: A decision model compared the costs of the 2 strategies. Sensitivity analysis and threshold analysis were used to identify the most important determinants of the overall cost of identifying advanced disease. RESULTS: Initial CT is the least costly strategy if the probability of finding advanced disease by initial CT is greater than 20%, if the probability of finding advanced disease by initial endoscopic ultrasound (EUS) is less than 30%, or if the cost of EUS is greater than 3.5 times the cost of CT. However, in our referral center population, endosonography found advanced disease more frequently than CT (44% vs. 13%; p < 0.0001) and the least costly strategy was initial endosonography (expected cost $804 vs. $844). CONCLUSION: CT remains as the initial staging test of choice in most clinical settings. However, in referral centers, initial EUS may be reasonable, but individualized model inputs must be obtained before reliable conclusions can be drawn. PMID- 11115902 TI - Bleeding Dieulafoy's lesions and the choice of endoscopic method: comparing the hemostatic efficacy of mechanical and injection methods. AB - BACKGROUND: Dieulafoy's lesion has unique endoscopic and histopathologic characteristics. This is a clinical trial of endoscopic therapy in 24 patients with Dieulafoy's lesions. METHODS: Patients were divided into 2 groups according to initial endoscopic treatment method. Data were analyzed with respect to clinical and endoscopic characteristics as well as outcomes. The 24 patients were evenly divided into mechanical (9 hemoclipping, 3 band ligation) and injection groups (12). RESULTS: The average number of therapeutic endoscopic sessions needed to achieve permanent hemostasis for the mechanical and injection groups were 1.17 and 1.67, respectively. Initial hemostasis was achieved in 91.7% of patients undergoing mechanical therapy and 75% of those undergoing injection therapy, with none in the former group needing subsequent surgery in comparison to 17% of the latter group. The rate of recurrent bleeding in the mechanical therapy group was significantly lower in comparison to the injection therapy group (8.3% versus 33.3%, p < 0. 05). CONCLUSIONS: Higher efficacy in terms of initial hemostasis and less recurrent bleeding was achieved by mechanical hemostatic therapy with hemoclip and band ligation compared with injection therapy. Endoscopic mechanical therapy is recommended as effective for bleeding Dieulafoy's lesions. PMID- 11115903 TI - Early attachment of anaerobic bacteria may play an important role in biliary stent blockage. AB - BACKGROUND: In vitro studies have demonstrated that ciprofloxacin suppresses Escherichia coli attachment on stents, and ciprofloxacin has been shown to prolong stent patency in cats. However, clinical studies with antibiotic prophylaxis have produced conflicting results. The aim of this study was to isolate and identify the bacteria that attach early on unblocked stents removed from patients and to study their enzyme activities. METHODS: Eighteen unblocked biliary stents were removed from 17 patients (benign obstruction in 14 and malignant obstruction in 4). All patients received antibiotic prophylaxis (mean of 6 days). Stents were in place for a mean of 33 days. The inside of stents was scraped and sludge was cultured aerobically and anaerobically. Identification of isolated bacteria and measurement of beta-glucuronidase and phospholipase C activities were performed by using standard techniques. Gastric and duodenal juice from 18 patients with no biliary diseases was used as control samples. RESULTS: All stents were patent and only 6 had visible sludge. There were 19 anaerobes isolated from 16 stents (Clostridium perfringens 13, Clostridium bifermentans 4 and Bacteroides fragilis 2). Phospholipase C was detected in all Clostridium species. beta-Glucuronidase was produced only by 12 of 13 C perfringens isolates. Sixteen aerobes including Enterococcus species and Bacillus species were isolated but none produced beta-glucuronidase or phospholipase C. There were no aerobic gram-negative bacteria isolated from stents. Clostridium species and B fragilis were not recovered from the control samples. CONCLUSIONS: In patients who had received antibiotic prophylaxis against gram-negative bacterial infection, anaerobic bacteria may play a role in initiating stent blockage. PMID- 11115904 TI - Effectiveness of endoscopic mucosal resection with submucosal saline injection technique for superficial squamous carcinomas of the esophagus. AB - BACKGROUND: Intraepithelial cancers (m1 cancer) and cancers that penetrate the basement membrane but do not approach the muscularis mucosae (m2 cancer) do not have lymph node metastasis and thus can be removed completely with mucosal resection. Therefore, in this study, the effectiveness of endoscopic mucosal resection with submucosal saline injection for removal of superficial esophageal cancers was investigated prospectively. METHODS: Twenty-five superficial esophageal cancers in 21 patients were removed with submucosal saline injection. When it was thought that a tumor had not been completely resected en bloc, it was removed completely in piecemeal fashion. Endoscopy was repeated 1, 3, 6, 12 months or more after endoscopic resection. RESULTS: All superficial esophageal cancers were completely removed: 18 (72%) en bloc and 7 (28%) by piecemeal resection. No recurrence was found during a mean observation period of 2.0 years (range 0.8 to 3.6) after resection. Bleeding occurred in 5 cases (24%) during or after resection but was successfully treated with the endoscopic alginate or thrombin spray technique. There was no perforation. CONCLUSION: Endoscopic mucosal resection with submucosal saline injection is effective for removal of superficial cancers of the esophagus. PMID- 11115905 TI - Covered metal stents for management of inoperable malignant colorectal strictures. AB - BACKGROUND: Metal stents have been reported as an effective alternative to surgery for the palliation of patients with colorectal neoplastic obstruction. Because most of the published series describe the use of uncovered stents, the purpose of our study was to prospectively evaluate the effectiveness, feasibility, safety, and outcome of covered stents for the palliative treatment of malignant colorectal strictures. METHODS: Sixteen patients with advanced distal colorectal cancer underwent placement of 10 and 12 cm long, 23 mm diameter covered stents under fluoroscopic and endoscopic control. Clinical and endoscopic follow-up was scheduled at 3- to 6-week intervals. RESULTS: Stent insertion was successful in 15 of 16 patients (93%). Perforation occurred in one patient during stent placement requiring colostomy. Relief of bowel obstruction was documented in all successfully treated patients. The median follow-up was 21 weeks (range 1 to 46). No recurrence of obstruction was observed during the follow-up period. Stent migration occurred in 2 patients, 7 and 21 days after stent placement. CONCLUSIONS: Covered stents may provide safe and effective palliation of patients with malignant rectosigmoid strictures. Prolonged luminal patency and sealing of fistulous tracts are potential advantages of covered versus uncovered stents in the palliative treatment of colorectal malignancies. ? PMID- 11115906 TI - Urgent endoscopic nasobiliary drainage without fluoroscopic guidance: A useful treatment for critically ill patients with biliary obstruction. AB - BACKGROUND: Endoscopic nasobiliary drainage (ENBD) is routinely performed under fluoroscopic control. This is a report of our experience with urgent ENBD without fluoroscopic guidance in critically ill patients. METHODS: Twenty-six critically ill patients who underwent urgent ENBD for biliary obstruction were analyzed. ENBD was performed without fluoroscopic control because of high risk of transportation or inaccessibility of the x-ray facilities. A pig-tailed nasobiliary catheter was inserted into the bile duct with the help of a guidewire under endoscopic control to bypass the site of obstruction. Successful placement was confirmed by free flow of bile on aspiration via the nasobiliary catheter. RESULTS: A nasobiliary catheter was successfully placed in 23 patients (88%). Adequate bile drainage was achieved in 20 patients with an overall success rate of 77%. There were no procedure-related complications. The mortality rate for patients with successful biliary drainage was 10% (2 of 20), in contrast to 83% (5 of 6) for the group in which drainage was unsuccessful. CONCLUSIONS: Urgent ENBD is effective for patients with biliary obstruction. With experience, this procedure may be successfully performed in critically ill patients without fluoroscopic guidance at primary care hospitals or intensive care units where fluoroscopic facilities are not readily available. PMID- 11115907 TI - 1999 ASGE endoscopic ultrasound survey. ASGE Ad Hoc Endoscopic Ultrasound Committee. AB - BACKGROUND: Gastrointestinal endoscopic ultrasound (EUS) has become an important imaging modality for the diagnosis and staging of gastrointestinal disorders. This study assessed current EUS practice, training, coding, and reimbursement in the United States. METHODS: A direct mail survey was sent to members of the American Society for Gastrointestinal Endoscopy. RESULTS: There were 115 American respondents. The median age was 39 years, 57% were in academic practice, and 84% performed endoscopic retrograde cholangiopancreatography. The median number of EUS procedures performed was 200. In the preceding year, the median number of upper EUS was 60, lower EUS 10, and EUS/fine-needle aspiration 3. The most common indication was evaluation of esophageal or gastric lesions. Forty-six (40%) trained an average of 0.4 advanced fellows in EUS during the prior year. Of endosonographers involved in training, 53% thought formal training was necessary, for a median of 6 months and 100 procedures; 82% did not know whether they were reimbursed for EUS. There was great variation in the use of current procedural terminology (CPT) codes for lower EUS and upper EUS/fine-needle aspiration. CONCLUSIONS: EUS in the United States in 1999 is performed mostly by young, academic, interventional endoscopists. Diagnostic upper EUS is most commonly performed. Few new endosonographers are being trained. There is great variability in CPT coding of lower EUS and EUS/fine-needle aspiration procedures. PMID- 11115908 TI - Isolated metastatic melanoma to the colon mimicking colon cancer. PMID- 11115909 TI - False submucosal rectal tumor. PMID- 11115910 TI - Pseudomelanosis duodena. PMID- 11115911 TI - Depressed type early cancer in ulcerative colitis. PMID- 11115912 TI - Effectiveness of the near-infrared electronic endoscope for diagnosis of the depth of involvement of gastric cancers. AB - BACKGROUND: Near-infrared light penetrates deeply into tissue but is not visible with a conventional endoscope. We developed a new infrared electronic endoscope system and evaluated its usefulness for assessing the depth of involvement of gastric cancers. METHODS: A total of 61 patients with depressed or ulcerated gastric cancers underwent infrared endoscopy after intravenous injection of indocyanine green. RESULTS: Intramucosal cancers were observed as tumors with or without a homogeneous tumor stain, whereas submucosal or deeper cancers were observed as tumors with an inhomogeneous stain or with pooling of the dye. Histologic examination showed staining properties of gastric cancers were significantly correlated with the characteristics of the vascular bed. The accuracy for depth of cancer invasion was 89% for mucosal cancers and 89% for submucosal or deeper cancers. CONCLUSION: Our new infrared electronic endoscope system was useful for distinguishing mucosal cancers from submucosal or deeper cancers either with or without ulcerative changes. PMID- 11115913 TI - Band ligation for colonic bleeding: modification of multiband ligating devices for use with a colonoscope. AB - BACKGROUND: Current multiband ligating devices can be useful in the treatment of colorectal hemorrhage but are not designed to be used with a colonoscope. This restricts the ability to treat more proximal sites of colonic bleeding with this type of device. This study describes methods of modifying multiband ligating devices for use with a colonoscope, and the application of both single band and modified multiband ligating devices in the treatment of colonic bleeding. METHODS: Five consecutive patients with colorectal hemorrhage were treated endoscopically with a single-band ligating device or a modified multiband ligating device. The lesions treated included two postpolypectomy ulcers, one arteriovenous malformation, one post-coagulation ulcer after ablation of an arteriovenous malformation, and one diverticulum. RESULTS: Colonoscopic band ligation using either device was successful in all five patients with follow-up ranging from 2 to 5 months. CONCLUSIONS: Colonoscopic band ligation appears to be a safe and effective treatment for various types of colorectal hemorrhage. Currently available multiband ligating devices can be easily modified for use with a colonoscope. Endoscopic band ligation of colonic bleeding may be particularly useful in patients with coagulopathies and those cases in which bleeding is uncontrollable with other therapeutic modalities. PMID- 11115914 TI - Access for percutaneous transhepatic cholangioscopy in patients with nondilated bile ducts using nasobiliary catheter cholangiography and oblique fluoroscopy. AB - BACKGROUND: Percutaneous transhepatic biliary drainage is required for percutaneous transhepatic cholangioscopy. However, puncture of nondilated bile ducts under ultrasonographic guidance is difficult. METHODS: In 10 patients with no ultrasonographic evidence of intrahepatic bile duct dilatation, percutaneous transhepatic biliary drainage was performed under fluoroscopic guidance using cholangiography obtained via a nasobiliary drainage catheter. Direct puncture was performed by means of a left ventral approach using oblique C-arm fluoroscopy. RESULTS: Bile duct puncture was successful in all patients. There were no procedure-related complications. Subsequent cholangioscopy was successful in all patients. CONCLUSIONS: Direct puncture using nasobiliary drainage cholangiography and oblique fluoroscopy is a useful method when cholangioscopy is necessary in patients with nondilated bile ducts. PMID- 11115915 TI - Accessory spleen: a potential cause of misdiagnosis at EUS. AB - BACKGROUND: Accessory spleen is a common congenital anomaly. There are currently no endoscopic ultrasound (EUS) criteria for the diagnosis or differentiation of this benign splenic anomaly from pathologic disorders including neoplasms. The purpose of this study was to characterize EUS features and develop criteria for the diagnosis of accessory spleen. METHODS: Ten patients undergoing EUS were found to have a possible pancreatic tail or perisplenic mass later confirmed by CT to be either an accessory spleen or a prominent lobule of the spleen. EUS criteria assessed included size, shape, location, echo texture, echo density and border demarcation. RESULTS: Ten patients (mean age 58 years) were evaluated. Indications for EUS were evaluation of pancreaticobiliary disease in 9 patients and a gastric submucosal mass in 1 patient. Eight patients had an accessory spleen and 2 had a lobulated spleen. The mean diameter of these lesions as seen on EUS was 2.70 x 3.12 cm. Nine were round and 1 was oval. Because all were located inferolateral to the pancreatic tail and medial to the spleen, 5 of 10 were initially thought to be pancreatic masses. All of these lesions had a sharp and regular outer margin and homogeneous echo texture, 4 were hypoechoic and 6 hyperechoic. There were no specific EUS features identified that differentiated splenic lobulations from accessory spleen. CONCLUSIONS: Accessory spleen and splenic lobulation can be misinterpreted as neoplasm by EUS. Although homogeneous, they can be hyperechoic or hypoechoic. Their sharp and regular outer margin and anatomic location may help to avoid misdiagnosis. Furthermore, computed tomography may be helpful in confirming their presence. PMID- 11115916 TI - Esophagomediastinal fistula and esophageal stricture as a complication of esophageal candidiasis: a case report. PMID- 11115917 TI - Features of cap polyposis by magnifying colonoscopy. PMID- 11115918 TI - Depressed type minute subserosal invasive colon cancer: report of a case. PMID- 11115919 TI - A case of multiple lymphangiomas of the colon suggesting colonic lymphangiomatosis. PMID- 11115920 TI - Small bowel obstruction due to trichobezoar: role of upper endoscopy in diagnosis. PMID- 11115921 TI - Obstructive pancreatitis due to mucus produced by metaplastic choledochal cyst epithelium. PMID- 11115922 TI - Polyethylene glycol-induced pancreatitis. PMID- 11115923 TI - Intramural duodenal hematoma complicating acute necrotizing pancreatitis. PMID- 11115925 TI - Streptococcus bovis bacteremia associated with Strongyloides stercoralis colitis. PMID- 11115924 TI - Phlegmonous gastritis: successful treatment with antibiotics and resolution documented by EUS. PMID- 11115926 TI - EUS for gastric lymphangioma. PMID- 11115927 TI - Successful endoscopic treatment of colon lymphangioma with a ligating device. PMID- 11115928 TI - Extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) of the sigmoid colon. PMID- 11115929 TI - Esophagopleural fistula after endoscopic sclerotherapy in a child. PMID- 11115930 TI - Breakage of a nasoenteral feeding tube in a 5-year-old child. PMID- 11115931 TI - Endoscopic and endoluminal techniques for the control of gastroesophageal reflux: are they ready for widespread clinical application? PMID- 11115932 TI - Making the numbers tell the truth. PMID- 11115933 TI - The development of the swallowable video capsule (M2A). PMID- 11115934 TI - Endoclips in prolonged colonic manometry. PMID- 11115935 TI - Response PMID- 11115936 TI - Diagnostic yield of colonoscopy in patients with a positive fecal occult blood test. PMID- 11115937 TI - Response PMID- 11115938 TI - The planning of clinical studies: bias and precision. PMID- 11115939 TI - Preface PMID- 11115940 TI - EUS instrumentation and accessories: a primer. PMID- 11115941 TI - EUS in the evaluation of esophageal carcinoma. PMID- 11115942 TI - EUS-guided fine-needle aspiration in the mediastinum. PMID- 11115943 TI - The role of EUS in the diagnosis of cystic lesions of the pancreas. PMID- 11115944 TI - Endosonographic-guided therapy of pancreatic pseudocysts. PMID- 11115945 TI - EUS-guided celiac plexus block for the management of pancreatic pain. PMID- 11115946 TI - EUS-guided investigational therapy for pancreatic cancer. PMID- 11115947 TI - High-frequency US probe sonography-assisted endoscopic mucosal resection. PMID- 11115948 TI - Coding for EUS. PMID- 11115949 TI - Emerging indications for EUS. PMID- 11115951 TI - Restoration of cellular immunity against tuberculosis in patients coinfected with HIV-1 and tuberculosis with effective antiretroviral therapy: assessment by determination of CD69 expression on T cells after tuberculin stimulation. AB - Whether immunity against opportunistic pathogens can be fully restored by control of HIV-1 replication remains open to question. This longitudinal study was conducted to measure anti-tuberculosis (TB) cellular immunity in 13 HIV-1/TB coinfected patients effectively treated by highly active antiretroviral therapy (HAART) in a period of 12 months. In this study, anti-TB cellular immunity was assessed by determining the frequencies of CD 69 expression on CD4+ and CD8+ T cells in response to purified protein derivative (PPD) stimulation (abbreviated as %CD4+CD69 to PPD and %CD8+CD69 to PPD). Here, we show that %CD4+CD69 to PPD correlated with the results of tuberculin skin tests and interferon-gamma (IFN gamma) production from PPD-stimulated CD4+ T cells, and %CD8+CD69 to PPD also correlated with CD8+ T cell-mediated PPD-specific cytolysis. In overall analysis for these 13 patients, both %CD4+CD69 to PPD and %CD8+CD69 to PPD increased significantly during the 12 months (p =. 003 and p <.001, respectively). However, we found %CD4+CD69 to PPD or %CD8+CD69 to PPD failed to increase substantially in some patients (i.e., immunologic nonresponders). A significantly higher proportion of patients whose baseline CD4+ count was <50 cells/mm3 were considered to be CD4+ nonresponders compared with those whose baseline CD4+ count was >50 cells/mm3. Furthermore, baseline CD4+ cell count in nonresponders is significantly lower than that in responders, although the effectiveness of HAART did not differ between them. Our results indicate that PPD-specific frequencies of CD69 expression may be used as surrogate markers of anti-TB cellular immunity. By this method, we show that full reconstitution of anti-TB cellular immunity in HIV-1/TB coinfected patients may not necessarily be achieved by "successful" HAART and may be influenced by the baseline immune status when HAART is started. These data suggest that the decision to discontinue secondary prophylaxis for opportunistic infections should be cautiously made, even when the CD4+ cell count has significantly increased. PMID- 11115950 TI - The dominant source of CD4+ and CD8+ T-cell activation in HIV infection is antigenic stimulation. AB - To distinguish between antigenic stimulation and CD4+ T-cell homeostasis as the cause of T-cell hyperactivation in HIV infection, we studied T-cell activation in 47 patients before and during highly active antiretroviral therapy (HAART). We show that expression of human leukocyte antigen (HLA)-DR, CD38, and Ki67 on T cells decreased during HAART but remained elevated over normal values until week 48 of therapy. We confirm previous reports that T-cell activation correlates positively with plasma HIV RNA levels (suggesting antigenic stimulation), and negatively with CD4 count (suggesting CD4+ T-cell homeostasis). However, these correlations may be spurious, because misleading, due to the well-established negative correlation between CD4 count and plasma HIV RNA levels. To resolve this conflict, we computed partial correlation coefficients. Correcting for CD4 counts, we show that plasma HIV RNA levels contributed to T-cell hyperactivation. Correcting for plasma HIV RNA levels, we show that CD4+ T-cell depletion contributed to T-cell activation. Correcting for both, activation of CD4+ and CD8+ T cells remained positively correlated. Because this suggests that CD4+ and CD8+ T-cell activation is caused by a common additional factor, we conclude that antigenic stimulation by HIV or other (opportunistic) infections is the most parsimonious explanation for T-cell activation in HIV infection. Persistence of HIV antigens may explain why T-cell activation fails to revert to levels found in healthy individuals after 48 weeks of therapy. PMID- 11115952 TI - Prospective randomized two-Arm controlled study to determine the efficacy of a specific intervention to improve long-term adherence to highly active antiretroviral therapy. AB - BACKGROUND: Nearly perfect compliance seems to be indispensable to obtain the maximum benefit from highly active antiretroviral therapy (HAART). Interventions to ensure a high level of adherence during a relatively long-term period of therapy are necessary. METHODS: This is a prospective, randomized, two-arm controlled study including patients starting their first-or second-line HAART who were randomized to receive psychoeducative intervention to implement adherence (experimental group [EG]) or a usual medical follow-up (control group [CG]). We aimed to study the efficacy of a psychoeducative intervention to ensure long-term adherence to HAART, its relation with the virologic efficacy of treatment, and to determine the variables related to long-term adherence. Visits were made at weeks 0, 4, 24, and 48 for data collection. Self-reported adherence was registered at each visit and its veracity was tested by randomized blood analyses performed without previous warning to 40% of patients. Appropriate adherence was defined as the consumption of >/=95% of medication prescribed. Statistical analyses were performed both by the as treated (AT) and the intention to treat missing = failure (ITT) methods. RESULTS: In all, 116 patients were included. At week 48, 94% of patients in the EG versus 69% controls achieved adherence >/=95% (p =.008); 89% of patients in the EG versus 66% controls had HIV-1 RNA levels <400 copies/ml (p =.026). Overall, 85% of patients with adherence >/=95% but only 45% of those with adherence <95% had viral load (VL) <400 copies/ml (p =. 008). In multivariate analysis, variables significantly related to adherence were having received a psychoeducative intervention (odds ratio [OR], 6.58; p =.04), poor effort to take medication (OR, 5.38; p =.03), and high self-perceived capacity to follow the regimen (OR, 13.76; p =.04). Self-reported adherence and drug plasma levels coincided in 93% of cases. However, differences in adherence did not reach statistical significance in the ITT analysis although a clear tendency toward benefit was observed in EG. CONCLUSIONS: Specific and maintained psychoeducative interventions based on excellence on clinical practice are useful to keep high levels of adherence as well as high levels of viral suppression. There is a clear relation between high adherence levels and virologic success. Assessment of certain specific variables related to adherence may be helpful to monitor patient's compliance in the clinical setting. PMID- 11115953 TI - Dose-finding study of once-daily indinavir/ritonavir plus zidovudine and lamivudine in HIV-infected patients. AB - BACKGROUND: Strategies for treatment of HIV need to be considered in terms of combining potency, safety, and convenience of dosage. However, regimens including once-daily protease inhibitors are not yet available. We have performed a pilot study to determine an indinavir/ritonavir (IND/RTV) regimen for once-daily dosing, by monitoring plasma levels. METHODS: Antiretroviral-naive HIV-infected adults were eligible. Therapy was zidovudine/lamivudine 1 pill twice daily plus IND/RIT (liquid formulation) 800/100 mg twice daily with food. At 4-week intervals, plasma levels were measured and dosage of IND/RIT switched to 1000/100 mg daily and then 800/200 mg daily. If 12-hour minimum concentrations (Cmin12h) of IND were too low (<0.1 microg/ml) with IND/RIT 1000/100 mg once daily in the first half of the patients, it was planned to switch directly to 800/200 mg once daily in the other half. RESULTS: In all, 27 patients were recruited. Mean baseline CD4+ lymphocyte count was 107 x 106/L (range, 4-623 x 106/L). Eleven patients (40%) discontinued the study medication within the first 4 weeks due to clinical progression (n = 3) or grade 1-2 RTV related side effects (n = 8). Nine patients (group A) switched from 800/100 mg twice daily to 1000/100 mg once daily and then to 800/200 mg once daily. Seven patients (group B) switched directly to 800/200 mg once daily. At week 32, viral load was <5 copies/ml in 15 of 16 patients (94%). RTV levels were always <2.1 microg/ml. The mean and 95% confidence interval for IND Cmin and Cmax in microg/ml was: using IND/RTV 800/100 mg twice daily (n = 16) 1.4 (0.5-2.3) and 6.7 (4.4-9.1), respectively; using IND/RTV 1000/100 mg once daily (n = 9) 0.18 (0-0.41) and 8.6 (3.3-14), respectively; and using 800/200 mg once daily (n = 16) 0.38 (0-0.9), and 7.5 (0.8 14.8). For all 16 patients who received IND/RTV 800/100 mg twice daily, the Cmin value for IND was >/=0.1 microg/ml. Conversely, IND Cmin was <0.1 microg/ml in 4 of 9 receiving 1000/100 mg once daily but in only 1 of 16 receiving 800/200 mg once daily. CONCLUSION: Once-daily regimen of IND/RIT is feasible and deserves further evaluation in larger randomized trials. Liquid formulation of RIT was not well tolerated by our antiretroviral-naive patients despite lower than suggested doses. PMID- 11115954 TI - Dose-finding study of a once-daily indinavir/ritonavir regimen. AB - In antiretroviral therapy, to improve compliance the need is increasing to develop regimens that combine potency and safety with convenient dosing. The objective of our study was to find a once-daily dosing regimen of a HIV-protease inhibitor, indinavir (IDV), by combining it with ritonavir (RTV). In the study, 12 healthy volunteers took a single IDV dose of 800 mg on day 1. Plasma and urine sampling was done for 12 hours. From day 2 to day 21, participants took RTV liquid 200 mg (group A) or 400 mg (group B) once daily. Repeated pharmacokinetic sampling was performed over the course of 24 hours, after single doses of indinavir 400 mg (day 15), 800 mg (day 18), and 1200 mg (day 21). The best dosage regimen in this pharmacokinetic study was selected based on efficacy and tolerability criteria. The study comprised 10 male and 2 female healthy volunteers, mean age, 25 years (range, 18-50 years), mean weight, 70 kg (range, 52-83 kg). One male participant discontinued on day 8 due to influenza. All other participants completed the study without the occurrence of serious adverse events. RTV inhibited indinavir plasma clearance by 51% to 70%, leading to increased and prolonged IDV exposure. Renal clearance was influenced by the addition of RTV and dosage increments of IDV. The efficacy criterion was best fulfilled by 1200 mg IDV/400 mg RTV, whereas this combination performed most poorly on tolerability criteria. Based on the single dose data, a once-daily regimen of IDV with a low dose of RTV is possible. The best dosage regimen to start with among those studied here appears to be 1200 mg IDV/400 mg RTV, which could be decreased at steady state to 800 IDV/400 RTV or 1200 IDV/200 RTV if toxicity occurs. Steady-state pharmacokinetic data of once-daily IDV/RTV regimens in HIV-infected patients are warranted. PMID- 11115955 TI - Synergistic inhibition of HIV-1 in activated and resting peripheral blood mononuclear cells, monocyte-derived macrophages, and selected drug-resistant isolates with nucleoside analogues combined with a natural product, resveratrol. AB - Resveratrol (trans-3,5,4;-trihydroxystilbene) is a phytoalexin present in grapes, wine, and certain plants, which has recently been reported to possess properties that may protect against atherosclerosis, certain cancers, and inflammation. We now report that resveratrol (RV) synergistically enhances the anti-HIV-1 activity of the nucleoside analogues zidovudine (AZT), zalcitabine (ddC), and didanosine (ddI). RV at 10 microM was not toxic to cells, and by itself reduced viral replication by 20% to 30%. In phytohemagglutinin (PHA)-activated peripheral blood mononuclear cells (PBMCs) infected with HTLV-IIIB, 10 microM RV reduced the 90 % inhibitory concentration (IC90) of AZT, ddC, and ddI by 3.5-, 5.5-, and 17.8 fold, respectively. Similar antiviral activity was demonstrated when ddI was combined with 5 or 10 mM RV in PBMCs infected with clinical isolates of HIV-1. The addition of RV resulted in a >10-fold augmentation of ddI-antiviral activity in infected monocyte-derived macrophages (MDMs). In a resting cell model of T lymphocytes which were infected with HTLV-IIIB, RV plus ddI in combination, but not individually, suppressed establishment of a productive viral infection. In addition, RV plus ddI markedly inhibited the replication of four ddI-resistant viral isolates, three of which presented mutations in the RT gene conferring RT multidrug resistance. Finally, when compared with hydroxyurea (HU), both 100 mM HU and 10 mM RV showed similar enhancement of ddI-antiviral suppressive activity. However, RV was shown to have less of a cellular antiproliferative effect than HU. PMID- 11115956 TI - In vitro anti-Toxoplasma gondii antibody production by peripheral blood mononuclear cells in the diagnosis and the monitoring of toxoplasmic encephalitis in AIDS-related brain lesions. AB - The spontaneous secretion in vitro of anti-Toxoplasma gondii antibodies by peripheral blood mononuclear cells was assessed in 69 patients with AIDS-related brain lesions. The sensitivity and specificity of this assay in the diagnosis of toxoplasmic encephalitis (TE) were found to be 85.4% and 92.8%, respectively. Twenty-four patients with TE were observed during 1-year follow-up after initiation of anti-Toxoplasma treatment and classified on the basis of their clinical and radiologic responses as sustained responders (SR; n = 11), incomplete responders (IR; n = 7) or transient responders (TR; n = 6). In vitro anti-T. gondii antibody secretion decreased as early as the first month after initiation of treatment and disappeared within the year in SRs, persisted in IRs, and decreased but rebounded at relapse in the TR patients. In vitro anti-T. gondii antibody, which reflects immune system activation by parasitic antigens, could be a surrogate marker of TE in AIDS patients. PMID- 11115957 TI - Nucleoside exposure in the children of HIV-infected women receiving antiretroviral drugs: absence of clear evidence for mitochondrial disease in children who died before 5 years of age in five United States cohorts. AB - BACKGROUND: Nucleoside reverse transcriptase inhibitors (NRTIs) have been associated with mitochondrial toxicity in individuals receiving treatment. A report of two deaths in Europe attributed to mitochondrial dysfunction in HIV uninfected infants with perinatal NRTI exposure prompted a review of five U.S. cohorts. METHODS: Deaths in HIV-exposed children <60 months of age and HIV uninfected or indeterminate were reviewed. Review included birth history; perinatal antiretroviral drug exposure; hospital, laboratory, and clinic records; death reports; autopsy results; and local physician queries. Deaths were classified as unrelated (Class 1), unlikely related (Class 2), possibly related (Class 3), or highly suggestive or proven relationship (Class 4), to mitochondrial dysfunction; sudden infant death syndrome (SIDS) was categorized separately. RESULTS AND CONCLUSIONS: Among over 20,000 children of HIV-infected women, over half of whom had been exposed to NRTIs, 223 died. In HIV-uninfected children, 26 deaths were attributed to Class 1, and 4 were attributed to SIDS. In HIV-indeterminate children, 141, 10, 3, and 0 were Classes 1, 2, 3, and 4, respectively; 33 were due to SIDS and 6 could not be classified. There was no indication that antiretroviral exposure was associated with Class 2 or 3 deaths, or deaths from SIDS. A search for mitochondrial dysfunction among living children in these cohorts is ongoing. PMID- 11115958 TI - Survival in children with perinatal HIV infection and very low CD4 lymphocyte counts. AB - OBJECTIVES: To evaluate clinical conditions associated with mortality in HIV infected children with CD4+ counts <100 cells/microl. METHODS: The Pediatric Spectrum of HIV Disease Project is a longitudinal medical record review study with eight study sites in the United States, which have been enrolling children since 1989. Survival time from baseline very low CD4 count (<100 cells/microl) to death was estimated using the Kaplan-Meier method. Cox proportional hazards models were used to evaluate the effect of clinical variables on mortality. RESULTS: Of 522 children (>/=1 year of age) with serial CD4+ T-lymphocyte measurements, the median age at the first very low CD4 count was 4.8 years. The estimated median survival following the first very low CD4 count was 36 months. The following factors present at the first very low CD4 count were independently associated with a higher risk of death: younger age, weight-for-age >2 standard deviations below the mean, and previously diagnosed AIDS. The subsequent development of cytomegalovirus (CMV)-associated disease, Mycobacterium avium intracellulare (MAI) infection, wasting syndrome, or esophageal candidiasis was also independently associated with a higher risk of death. CONCLUSION: Survival in HIV-infected children with very low CD4 counts before introduction of highly active antiretroviral therapy was highly variable. Poor nutritional status and the development of CMV disease or MAI infection were associated with the shortest survival times. PMID- 11115959 TI - Partner violence, partner notification, and women's decisions to have an HIV test. AB - OBJECTIVES: Reports of partner violence against HIV-positive women after they have disclosed their serostatus have led some to reassess partner notification strategies and to speculate that fear of partner violence following partner notification may influence women's HIV testing decisions. We studied whether associations exist between women's declining to have an HIV test and history of partner violence, fear of partner violence, previous experience with partner notification, or beliefs about partner notification. METHODS: In this cross sectional study, we interviewed women seen at Newark and Miami sexually transmitted disease clinics. The women were at least 18 years old, not known to be HIV positive, not tested for HIV in the previous 3 months, and offered HIV testing during the clinic visit. Women who declined testing were compared with women who accepted. RESULTS: Of 490 participants (89% of eligible women), 16% reported partner violence in the past year, and 28% declined HIV testing. Declining the test was not significantly (p >.05) associated with history or fear of partner violence, previous experience with partner notification, or beliefs about partner notification. When specifically asked, only 2 women responded that their declining the test was related to fear that their partner or partners might harm them if the women tested positive. CONCLUSIONS: Among women seen at these clinics, we did not find evidence that declining the HIV test was strongly influenced by partner violence, previous experience with partner notification, or beliefs about partner notification. However, many women reported partner violence. Therefore, providers should assess the potential for partner violence and be prepared to make appropriate referrals. PMID- 11115960 TI - Epidemiologic characteristics of Koebner's phenomenon in AIDS-related Kaposi's sarcoma. PMID- 11115961 TI - Lipoatrophy, fat accumulation, and mixed syndrome in protease inhibitor-naive HIV infected patients. PMID- 11115962 TI - Reduction of bronchoscopy performances should be included in the advantages of highly active antiretroviral therapy (HAART) PMID- 11116043 TI - Mouse model of unstable atherosclerotic plaque? PMID- 11116044 TI - Scavenger receptors in atherosclerosis: new answers, new questions. PMID- 11116045 TI - Lipoprotein receptors, macrophages, and sphingomyelinase. PMID- 11116046 TI - Thrombin activatable fibrinolysis inhibitor and an antifibrinolytic pathway. AB - Coagulation and fibrinolysis are processes that form and dissolve fibrin, respectively. These processes are exquisitely regulated and protect the organism from excessive blood loss or excessive fibrin deposition. Regulation of these cascades is accomplished by a variety of mechanisms involving cellular responses, flow, and protein-protein interactions. With respect to regulation mediated by protein-protein interaction, the coagulation cascade appears to be more complex than the fibrinolytic cascade because it has more components. Yet each cascade is regulated by initiators, cofactors, feedback reactions, and inhibitors. Coagulation is also controlled by an anticoagulant pathway composed of (minimally) thrombin, thrombomodulin, and protein C.(1) Protein C is converted by the thrombin/thrombomodulin complex to activated protein C (APC), which catalyzes the proteolytic inactivation of the essential cofactors required for thrombin formation, factors Va and VIIIa. An analogous antifibrinolytic pathway has been identified recently. This pathway provides an apparent symmetry between coagulation and fibrinolysis and is also composed of thrombin, thrombomodulin, and a zymogen that is activated to an enzyme. The enzyme proteolytically inactivates a cofactor to attenuate fibrinolysis. However, unlike APC, which is a serine protease, the antifibrinolytic enzyme is a metalloprotease that exhibits carboxypeptidase B-like activity. Within a few years of each other, 5 groups independently described a molecule that accounts for this antifibrinolytic activity. We refer to this molecule as thrombin activatable fibrinolysis inhibitor (TAFI), a name that is based on functional properties by which it was identified, assayed, and purified. (Because of the preferences of some journals "activatable" is occasionally referred to as "activable.") This review will encompass a historical account of efforts to isolate TAFI and characterize it with respect to its activation, activity, regulation, and potential function in vivo. PMID- 11116047 TI - Role of p38 mitogen-activated protein kinase in neointimal hyperplasia after vascular injury. AB - p38 mitogen-activated protein kinase (MAPK) is involved in intracellular signals that regulate a variety of cellular responses during inflammation. However, the role of p38 MAPK in atherosclerosis, a chronic inflammatory disorder, remains uncertain. The aim of the present study was to examine the role of p38 MAPK in the development of neointimal hyperplasia in balloon-injured rat carotid arteries. Immunohistochemical studies indicated that p38 MAPK was rapidly activated in the majority of medial cells in injured arterial walls. Rats treated with FR167653, a selective inhibitor of p38 MAPK, at a dosage of 10 mg x kg(-1) x d(-1), had a 29.4% lower intima-to-media ratio than the untreated controls at 14 days after balloon injury (P:<0.05). The percentage of proliferating nuclear antigen-positive cells in the media at 48 hours was significantly lower in the FR167653-treated group than in the control group. Quantitative competitive reverse transcription-polymerase chain reaction analysis revealed that interleukin-1beta mRNA expression in arteries was significantly inhibited by FR167653 (to 18.1% of control, P:<0.05) at 8 hours after balloon injury. Moreover, p38 MAPK activation and interleukin-1beta production by lipopolysaccharide-stimulated vascular smooth muscle cells were inhibited by FR167653 in a concentration-dependent manner in vitro. These results indicate that p38 MAPK is activated in vascular walls after injury and promotes neointimal formation and suggest that selective inhibition of p38 MAPK may be effective in the prevention of restenosis after percutaneous transluminal coronary angioplasty. PMID- 11116049 TI - Fc-gamma receptor cross-linking by immune complexes induces matrix metalloproteinase-1 in U937 cells via mitogen-activated protein kinase. AB - Matrix metalloproteinase-1 (MMP-1) secreted by macrophages potentially contributes to plaque rupture. Because large quantities of immunoglobulin G and ICs (ICs), including low density lipoprotein-containing ICs (LDL-ICs), are present in atherosclerotic lesions, we examined the effect of LDL-ICs on macrophage MMP-1 expression. With the use of ICs prepared with human LDL and rabbit anti-LDL antiserum, our enzyme-linked immunosorbent assays and Northern blots showed that MMP-1 secretion and expression by U937 histiocytes were induced by LDL-ICs. Furthermore, our results showed that blocking of Fc-gamma receptor I and II (FcgammaRI and FcgammaRII) inhibited 70% and 55%, respectively, of the LDL IC-induced secretion of MMP-1. Finally, our data showed that both PD98059, an inhibitor of the mitogen-activated protein kinase pathway, and Ro-31-2880, an inhibitor of protein kinase C, inhibited LDL-IC-stimulated MMP-1 secretion in a dose-dependent manner, with 96% and 95% inhibition, respectively, at the respective doses of 50 micromol/L and 80 nmol/L. In conclusion, this study demonstrated for the first time that LDL-ICs induce macrophage MMP-1 secretion by cocross-linking FcgammaRI and FcgammaRII and triggering a protein kinase C dependent mitogen-activated protein kinase pathway. These results suggest that LDL-ICs and other ICs localized in atherosclerotic plaques may be potent stimulators for macrophage MMP-1 expression and may contribute to plaque rupture and acute coronary events. PMID- 11116048 TI - Mitogen-activated protein kinases mediate matrix metalloproteinase-9 expression in vascular smooth muscle cells. AB - Expression of matrix metalloproteinase (MMP)-9 has been linked to the progression of plaque rupture and intimal formation in arterial lesions. In this study, we determined which factors and signaling pathways are involved in regulating the MMP-9 gene. Rat carotid arterial smooth muscle cells treated with tumor necrosis factor (TNF)-alpha showed a marked increase in MMP-9 activity and mRNA level, whereas platelet-derived growth factor (PDGF) showed a slight induction of the MMP-9 mRNA level. TNF-alpha treatment caused an increase in c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38 MAPK), and extracellular signal-regulated kinase (ERK) activities, whereas PDGF treatment caused an increase in ERKs and p38 MAPK activities without any effect on JNK activity. Treatment with either SB203580 (inhibitor of p38 MAPK) or U0126 (inhibitor of the ERK pathway) downregulated the TNF-alpha-induced MMP-9 expression in a dose dependent manner. Treatment of cells with TNF-alpha and PDGF together stimulated the MMP-9 expression at a level higher than that observed with either factor alone, suggesting that TNF-alpha and PDGF have a synergistic effect on MMP-9 expression in arterial smooth muscle cells. Furthermore, suboptimal inhibitory concentrations of SB203580 and U0126 together almost completely inhibited the MMP 9 expression. These results suggest that p38 MAPK and ERK pathways contribute to the transcriptional regulation of MMP-9 in arterial smooth muscle cells. PMID- 11116050 TI - In vivo activation of rat aortic platelet-derived growth factor and epidermal growth factor receptors by angiotensin II and hypertension. AB - It is unclear whether the previous in vitro evidence of a link between angiotensin II (Ang II) and growth factor receptors can apply to the in vivo situation. In this study, we examined vascular platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) receptor activation in stroke-prone spontaneously hypertensive rats (SHRSP) and the role of Ang II. Tyrosyl phosphorylation of the growth factor receptors was determined by Western blot analysis coupled with immunoprecipitation. Tyrosyl phosphorylation of the aortic PDGF beta-receptor, but not the EGF receptor, was chronically increased in SHRSP with hypertension, compared with normotensive rats, being accompanied by increased extracellular signal-regulated kinase (ERK) activity. Treatment of SHRSP with ACE inhibitors (perindopril or enalapril) significantly reduced aortic PDGF beta-receptor tyrosyl phosphorylation and ERK activity, whereas treatment with hydralazine failed to reduce these activities. Therefore, these aortic changes in SHRSP were mediated by Ang II in response to vascular ACE. Ang II was infused into rats to examine the effects on aortic growth factor receptors. Chronic Ang II infusion, via the angiotensin type 1 receptor, significantly increased activation of the aortic PDGF beta-receptor but not the EGF receptor. Thus, the aortic PDGF beta-receptor, activated by ACE-mediated Ang II, seems to be responsible for vascular remodeling in hypertensive rats. PMID- 11116051 TI - Endoglin is overexpressed after arterial injury and is required for transforming growth factor-beta-induced inhibition of smooth muscle cell migration. AB - Endoglin is a homodimeric membrane glycoprotein primarily expressed on endothelial cells. In association with transforming growth factor (TGF)-ss receptors I and II, it can bind TGF-beta1 and -beta3 and form a functional receptor complex. There is increasing evidence that endoglin can modulate the cellular response to TGF-beta, a factor implicated in vascular lesion formation in human and experimental models. The purpose of this study was to analyze the expression of endoglin in normal and balloon-injured porcine coronary arteries and in normal and atherosclerotic human coronary arteries and to determine its ability to mediate the effects of TGF-beta on the migration of vascular smooth muscle cells (SMCs). In normal porcine coronary arteries, endoglin was of low abundance and was found primarily on endothelial cells and adventitial fibroblasts, as well as on a minority of medial SMCs. On days 3, 7, and 14 after angioplasty, endoglin was present not only on endothelial cells but also on adventitial myofibroblasts and medial SMCs of porcine coronary arteries. By day 28, few or no cells expressed endoglin. In situ hybridization revealed that endoglin mRNA expression appeared to be highest in endothelial cells on days 3, 7, and 14 days after injury and absent thereafter. With a second balloon injury, a similar pattern of endoglin protein and mRNA expression was observed. In human vascular tissue, endoglin immunolabeling was higher in endarterectomy specimens removed from diseased coronary arteries than in normal internal mammary arteries. In vitro, antisense oligonucleotides to endoglin decreased its expression and antagonized the TGF-beta-mediated inhibition of human and porcine SMC migration. In summary, upregulation of endoglin occurs during arterial repair and in established atherosclerotic plaques and may be required for modulation of SMC migration by TGF-beta. PMID- 11116052 TI - Neutrophil, not macrophage, infiltration precedes neointimal thickening in balloon-injured arteries. AB - Macrophages are abundant after stent-induced arterial injury. Inhibition of macrophage recruitment blocks neointimal growth in this model. In contrast, after superficial injury from balloon endothelial denudation, macrophages are sparse. However, many anti-inflammatory therapies remain effective against neointimal growth after balloon injury. To investigate further the role of leukocytes after injury, 41 New Zealand White rabbits underwent iliac artery balloon denudation. In 18, subcutaneous pumps were placed to deliver intravenous heparin (0.3 mg/kg per hour). Arteries were harvested at 6 hours and at 3, 7, and 14 days. In 8 animals, either M1/70 (a monoclonal antibody [mAb] against adhesion molecule Mac 1) or a nonspecific IgG was given (5 mg/kg IV bolus and then 1 mg/kg SC QOD), and arteries were harvested at 6 hours and 3 days. Computer-aided morphometry was performed as was immunohistochemistry to assess smooth muscle cell (SMC) proliferation (bromodeoxyuridine-positive cells), neutrophil content (RPN357, mAb against rabbit neutrophil/thymocyte), and macrophage content (RAM-11, mAb against rabbit macrophage). Heparin inhibited neointimal growth at 7 and 14 days (64% and 32.5% reduction, respectively; P:<0.05). Neutrophils were observed in the media early after balloon injury, and heparin and M1/70 inhibited this infiltration (82% and 83% reduction, respectively; P:<0.05 each) with a coincident inhibition of medial SMC proliferation at 3 days (49% and 84% reduction, respectively; P:<0.05 each). Macrophages were absent at all time points. Neutrophil, but not macrophage, infiltration occurs early after endothelial denudation. Inhibition of this process is associated with a reduction in medial SMC proliferation. These data suggest a central role for neutrophils in restenosis and help to explain prior reports of an inhibitory effect of anti-inflammatory therapies on neointimal growth after balloon injury. PMID- 11116053 TI - Protein kinase A-dependent stimulation of rat type II secreted phospholipase A(2) gene transcription involves C/EBP-beta and -delta in vascular smooth muscle cells. AB - Type II secreted phospholipase A(2) (sPLA(2)) releases precursors of important inflammatory lipid mediators from phospholipids. Some observations have indicated that the sPLA(2), which has been implicated in chronic inflammatory conditions such as arthritis, contributes to atherosclerosis in the arterial wall. sPLA(2) was not detected in control vascular smooth muscle cells (VSMC). Treatment of VSMC with agents that increase intracellular cAMP (eg, forskolin, dibutyryl [db] cAMP) resulted in a time- and concentration-dependent increase in sPLA(2) gene expression. Semiquantitative reverse transcriptase polymerase chain reaction (RT PCR) showed a marked dose-dependent inhibition of forskolin-induced mRNA by protein kinase A inhibitor. Electrophoretic mobility shift analysis of nuclear proteins from forskolin-treated and db-cAMP-treated VSMC with C/EBP consensus oligonucleotides and C/EBP oligonucleotides from the rat promoter revealed greater binding than in control VSMC. Incubation of VSMC with H89, a specific protein kinase inhibitor, also blocked the binding of nuclear C/EBP to the C/EBP site of the rat promoter induced by db-cAMP and forskolin. Binding was unchanged with the use of CRE consensus oligonucleotides. Antibodies revealed the specific formation of C/EBP/DNA complexes, the majority of which were supershifted by C/EBP-ss and -delta antibodies. Functional activation of C/EBP was confirmed by a luciferase reporter gene assay. A construct comprising 4 tandem repeat copies of the C/EBP element from the rat sPLA(2) promoter linked to luciferase was transcriptionally activated in VSMC by cotransfection with expression vector for the protein kinase A catalytic subunit. It was also significantly activated in transfected VSMC treated by forskolin or db-cAMP. H89 inhibited this activations. We therefore conclude that the increases in sPLA(2) mRNA and enzyme activity produced by cAMP-elevating agents is controlled by a mechanism involving nuclear C/EBP-ss and -delta acting through a protein kinase A signaling pathway. PMID- 11116055 TI - Coadministration of angiopoietin-1 and vascular endothelial growth factor enhances collateral vascularization. AB - Using growth factors to induce vasculogenesis is a promising approach in the treatment of ischemic legs and myocardium. Because the vasculogenesis requires a cascade of growth factors, their receptors, and intracellular signals, such therapies may require the application of more than a single growth factor. We examined the effect of 2 endothelial cell-specific growth factors, angiopoietin-1 (Ang1) and vascular endothelial growth factor (VEGF), on primary cultured porcine coronary artery endothelial cells. VEGF, but not Ang1, increased DNA synthesis and cell number. Ang1 or VEGF induced migration and sprouting activity, increased plasmin and matrix metalloproteinase-2 secretion, and decreased tissue inhibitors of metalloproteinase type 2 secretion. A combination of the submaximal doses of Ang1 and VEGF enhanced these effects and was more potent than the maximal dose of either alone. In a rabbit ischemic hindlimb model, a combination of Ang1 and VEGF gene delivery produced an enhanced effect on resting and maximal blood flow and capillary formation that was greater than that of either factor alone. Angiographic analyses revealed that larger blood vessels were formed after gene delivery of Ang1 or Ang1 plus VEGF than after VEGF gene delivery. These results suggest that combined treatment of Ang1 and VEGF could be used to produce therapeutic vascularization. PMID- 11116054 TI - Hypercholesterolemia superimposed by experimental hypertension induces differential distribution of collagen and elastin. AB - We studied the mural distribution of collagen types I and III and tropoelastin in enhanced experimental atherogenesis induced in rabbits by hyperlipidemia superimposed by hypertension. Animals were fed a high-cholesterol diet for 5 weeks and also subjected to midthoracic aortic coarctation for 4 weeks. Serum cholesterol levels were increased and blood pressure was elevated proximal to the coarctation. Foam cell lesions developed in the aorta proximal to the coarctation. In situ hybridization and immunohistochemistry showed that gene expression of collagen types I and III and tropoelastin was upregulated, with a differential distribution across the arterial wall. New collagen type I was mainly distributed in the intima, the outer media, and the adventitia. New collagen type III was spread more uniformly across the wall, including the adventitia, whereas tropoelastin was mainly localized in intimal foam cell lesions. Morphometric data showed an increase in wall thickness. These results suggest that collagen types I and III play a role in remodeling of the aortic wall in response to hypertension. The remarkable involvement of the adventitia in this response indicates that the adventitia is an important component of the arterial wall. Tropoelastin is closely associated with foam cell lesion formation, suggesting a role for this component in atherogenesis as well. PMID- 11116056 TI - Hyperhomocysteinemia impairs angiogenesis in response to hindlimb ischemia. AB - Hyperhomocysteinemia (HH) is an independent risk factor for atherosclerosis, including peripheral arterial occlusive disease (PAOD). Because angiogenesis and collateral vessel formation are important self-salvage mechanisms for ischemic PAOD, we examined whether HH modulates angiogenesis in vivo in a rat model of hindlimb ischemia. Rats were divided into 3 groups: the control group was given tap water, the HH group was given water containing L-methionine (1 g x kg(-1) x d(-1)), and the HH+L-arg group was given water containing methionine (1 g x kg( 1) x d(-1)) and l-arginine (2.25 vol%). At day 14 of the dietary modifications, the left femoral artery and vein were excised, and the extent of angiogenesis and collateral vessels in the ischemic limb were examined for 4 weeks. Plasma homocysteine levels significantly increased (P:<0.001), and plasma and tissue contents of nitrite+nitrate as well as tissue cGMP levels significantly decreased in the HH group compared with the control group (P:<0.01). Laser Doppler blood flowmetry (LDBF) revealed a significant decrease in the ischemic/normal limb LDBF ratio in the HH group at days 7, 14, 21, and 28 (P:<0.01 versus control). Angiography revealed a significant decrease in the angiographic score in the HH group at day 14 (P:<0.001 versus control). Immunohistochemistry of ischemic tissue sections showed a significant reduction in the capillary density in the HH group (P:<0. 001 versus control). Oral l-arginine supplementation in rats with HH (HH+L-arg) restored the decreased plasma and tissue nitrite+nitrate and cGMP contents (P:<0.05) as well as angiogenesis, as assessed by LDBF (P:<0.05 versus HH), angiographic score (P:<0.01 versus HH), and capillary density (P:<0.001 versus HH). In summary, HH impaired ischemia-induced angiogenesis and collateral vessel formation in a rat model of hindlimb ischemia in vivo. The mechanism of the HH-induced impairment of angiogenesis might be mediated in part by a reduced bioactivity of endogenous NO in the HH state. PMID- 11116057 TI - Advanced atherosclerotic lesions in the innominate artery of the ApoE knockout mouse. AB - Most previous studies of atherosclerosis in hyperlipidemic mouse models have focused their investigations on lesions within the aorta or aortic sinus in young animals. None of these studies has demonstrated clinically significant advanced lesions. We previously mapped the distribution of lesions throughout the arterial tree of apolipoprotein E knockout (apoE(-/-)) mice between the ages of 24 and 60 weeks. We found that the innominate artery, a small vessel connecting the aortic arch to the right subclavian and right carotid artery, exhibits a highly consistent rate of lesion progression and develops a narrowed vessel characterized by atrophic media and perivascular inflammation. The present study reports the characteristics of advanced lesions in the innominate artery of apoE( /-) mice aged 42 to 60 weeks. In animals aged 42 to 54 weeks, there is a very high frequency of intraplaque hemorrhage and a fibrotic conversion of necrotic zones accompanied by loss of the fibrous cap. By 60 weeks of age, the lesions are characterized by the presence of collagen-rich fibrofatty nodules often flanked by lateral xanthomas. The processes underlying these changes in the innominate artery of older apoE(-/-) mice could well be a model for the critical processes leading to the breakdown and healing of the human atherosclerotic plaque. PMID- 11116058 TI - Reduced atherosclerotic lesions in mice deficient for total or macrophage specific expression of scavenger receptor-A. AB - The absence of the scavenger receptor A (SR-A)-I/II has produced variable effects on atherosclerosis in different murine models. Therefore, we examined whether SR AI/II deficiency affected atherogenesis in C57BL/6 mice, an inbred strain known to be susceptible to diet-induced atherosclerotic lesion formation, and whether the deletion of macrophage SR-AI/II expression would modulate lesion growth in C57BL/6 mice and LDL receptor (LDLR)(-/-) mice. SR-AI/II-deficient (SR-AI/II(-/ )) female and male mice on the C57BL/6 background were challenged with a butterfat diet for 30 weeks. No differences were detected in plasma lipids between SR-AI/II(-/-) and SR-AI/II(+/+) mice, whereas both female and male SR AI/II(-/-) mice had a tremendous reduction (81% to 86%) in lesion area of the proximal aorta compared with SR-AI/II(+/+) mice. Next, to analyze the effect of macrophage-specific SR-AI/II deficiency in atherogenesis, female C57BL/6 mice were lethally irradiated, transplanted with SR-AI/II(-/-) or SR-AI/II(+/+) fetal liver cells, and challenged with the butterfat diet for 16 weeks. In a separate experiment, male LDLR(-/-) mice were reconstituted with SR-AI/II(-/-) or SR AI/II(+/+) fetal liver cells and challenged with a Western diet for 10 weeks. No significant differences in plasma lipids and lipoprotein profiles were noted between the control and experimental groups in either experiment. SR-AI/II(-/-)- >C57BL/6 mice, however, had a 60% reduction in lesion area of the proximal aorta compared with SR-AI/II(+/+)-->C57BL/6 mice. A similar level of reduction (60%) in lesion area was noted in the proximal aorta and the entire aorta en face of SR AI/II(-/-)-->LDLR(-/-) mice compared with SR-AI/II(+/+)-->LDLR(-/-) mice. These results demonstrate in vivo that SR-AI/II expression has no impact on plasma lipid levels and that macrophage SR-AI/II contributes significantly to atherosclerotic lesion formation. PMID- 11116060 TI - Sphingomyelinase converts lipoproteins from apolipoprotein E knockout mice into potent inducers of macrophage foam cell formation. AB - The apoE knockout (E0) mouse is one of the most widely used animal models of atherosclerosis, and there may be similarities to chylomicron remnant-induced atherosclerosis in humans. Although the lesions of these mice contain large numbers of cholesteryl ester (CE)-laden macrophages (foam cells), E0 plasma lipoproteins are relatively weak inducers of cholesterol esterification in macrophages. Previous in vivo work has suggested that arterial wall sphingomyelinase (SMase) may promote atherogenesis in the E0 mouse, perhaps by inducing subendothelial lipoprotein aggregation and subsequent foam cell formation. The goal of the present study was to test the hypothesis that the modification of E0 lipoproteins by SMase converts these lipoproteins into potent inducers of macrophage foam cell formation. When d<1.063 E0 lipoproteins were pretreated with SMase and then incubated with E0 macrophages, cellular CE mass and stimulation of the cholesterol esterification pathway were increased approximately 5-fold compared with untreated lipoproteins. SMase-treated E0 lipoproteins were more potent stimulators of cholesterol esterification than either E0 lipoproteins in the presence of lipoprotein lipases or oxidized E0 lipoproteins. The uptake and degradation of SMase-treated E0 lipoproteins by macrophages were saturable and specific and substantially inhibited by partial proteolysis of cell-surface proteins. Uptake and degradation were diminished by an anti-apoB antibody and by competition with human S(f) 100-400 hypertriglyceridemic VLDL, raising the possibility that a receptor that recognizes apoB-48 might be involved. In conclusion, SMase-modification of E0 lipoproteins, a process previously shown to occur in lesions, may be an important mechanism for foam cell formation in this widely studied model of atherosclerosis. Moreover, the findings in this report may provide important clues regarding the atherogenicity of chylomicron remnants in humans. PMID- 11116059 TI - Effect of human scavenger receptor class A overexpression in bone marrow-derived cells on cholesterol levels and atherosclerosis in ApoE-deficient mice. AB - In the arterial wall, scavenger receptor class A (SRA) is implicated in pathological lipid deposition. In contrast, in the liver, SRA is suggested to remove modified lipoproteins from the circulation, thereby protecting the body from their pathological action. The role of SRA on bone marrow-derived cells in lipid metabolism and atherogenesis was assessed in vivo by transplantation of bone marrow cells overexpressing human SRA (MSR1) to apoE-deficient mice. In vitro studies with peritoneal macrophages from the transplanted mice showed that macrophage scavenger receptor function, as measured by cell association and degradation studies with acetylated LDL, was approximately 3-fold increased on overexpression of MSR1 in bone marrow-derived cells as compared with control mice. Despite the increased macrophage scavenger receptor function in vitro, no significant effect of MSR1 overexpression in bone marrow-derived cells on the in vivo atherosclerotic lesion development was found. In addition to arterial wall macrophages, liver sinusoidal Kupffer cells also overexpress MSR1 after bone marrow transplantation, which may scavenge atherogenic particles more efficiently from the blood compartment. Introduction of bone marrow cells overexpressing human MSR1 in apoE-deficient mice induced a significant reduction in serum cholesterol levels of approximately 20% (P:<0.001, 2-way ANOVA) as the result of a decrease in VLDL cholesterol. It is suggested that the reduction in VLDL cholesterol levels is due to increased clearance of modified lipoproteins by the overexpressed MSR1 in Kupffer cells of the liver, thereby protecting the arterial wall against the proatherogenic action of modified lipoproteins. PMID- 11116061 TI - Plasma sphingomyelin level as a risk factor for coronary artery disease. AB - Only a fraction of the clinical complications of atherosclerosis are explained by known risk factors. Animal studies have shown that plasma sphingomyelin (SM) levels are closely related to the development of atherosclerosis. SM carried into the arterial wall on atherogenic lipoproteins may be locally hydrolyzed by sphingomyelinase, promoting lipoprotein aggregation and macrophage foam cell formation. A novel, high-throughput, enzymatic method to measure plasma SM levels has been developed. Plasma SM levels were related to the presence of coronary artery disease (CAD) in a biethnic angiographic case-control study (279 cases and 277 controls). Plasma SM levels were higher in CAD patients than in control subjects (60+/-29 versus 49+/-21 mg/dL, respectively; P:<0. 0001). Moreover, the ratio of SM to SM+phosphatidylcholine (PC) was also significantly higher in cases than in controls (0.33+/-0.13 versus 0.29+/-0.10, respectively; P:<0.0001). Similar relationships were observed in African Americans and whites. Plasma SM levels showed a significant correlation with remnant cholesterol levels (r=0.51, P:<0.0001). By use of multivariate logistic regression analysis, plasma SM levels and the SM/(SM+PC) ratio were found to have independent predictive value for CAD after adjusting for other risk factors, including remnants. The odds ratio (OR) for CAD was significantly higher for the third and fourth quartiles of plasma SM levels (OR 2.81 [95% CI 1.66 to 4.80] and OR 2.33 [95% CI 1.38 to 3. 92], respectively) as well as the SM/(SM+PC) ratio (OR 1.95 [95% CI 1.10 to 3.45] and OR 2.33 [95% CI 1.34 to 4.05], respectively). The findings indicate that human plasma SM levels are positively and independently related to CAD. Plasma SM levels could be a marker for atherogenic remnant lipoprotein accumulation and may predict lipoprotein susceptibility to arterial wall sphingomyelinase. PMID- 11116062 TI - High levels of Lp(a) with a small apo(a) isoform are associated with coronary artery disease in African American and white men. AB - Elevated levels of lipoprotein(a) [Lp(a)] and the presence of small isoforms of apolipoprotein(a) [apo(a)] have been associated with coronary artery disease (CAD) in whites but not in African Americans. Because of marked race/ethnicity differences in the distribution of Lp(a) levels across apo(a) sizes, we tested the hypothesis that apo(a) isoform size determines the association between Lp(a) and CAD. We related Lp(a) levels, apo(a) isoforms, and the levels of Lp(a) associated with different apo(a) isoforms to the presence of CAD (>/=50% stenosis) in 576 white and African American men and women. Only in white men were Lp(a) levels significantly higher among patients with CAD than in those without CAD (28.4 versus 16.5 mg/dL, respectively; P:=0.004), and only in this group was the presence of small apo(a) isoforms (<22 kringle 4 repeats) associated with CAD (P:=0.043). Elevated Lp(a) levels (>/=30 mg/dL) were found in 26% of whites and 68% of African Americans, and of those, 80% of whites but only 26% of African Americans had a small apo(a) isoform. Elevated Lp(a) levels with small apo(a) isoforms were significantly associated with CAD (P:<0.01) in African American and white men but not in women. This association remained significant after adjusting for age, diabetes mellitus, smoking, hypertension, HDL cholesterol, LDL cholesterol, and triglycerides. We conclude that elevated levels of Lp(a) with small apo(a) isoforms independently predict risk for CAD in African American and white men. Our study, by determining the predictive power of Lp(a) levels combined with apo(a) isoform size, provides an explanation for the apparent lack of association of either measure alone with CAD in African Americans. Furthermore, our results suggest that small apo(a) size confers atherogenicity to Lp(a). PMID- 11116063 TI - HIV protease inhibitors stimulate hepatic triglyceride synthesis. AB - Hyperlipidemia may complicate the use of HIV protease inhibitors (PIs) in AIDS therapy. To determine the cause of hyperlipidemia, the effect of PIs on lipid metabolism was examined with HepG2 liver cells and AKR/J mice. In HepG2 cells, the PIs ABT-378, nelfinavir, ritonavir, and saquinavir stimulated triglyceride synthesis; ritonavir increased cholesterol synthesis; and amprenavir and indinavir had no effect. Moreover, nelfinavir increased mRNA expression of diacylglycerol acyltransferase and fatty acid synthase. The retinoid X receptor agonist LG100268, but not the antagonist LG100754, further increased PI stimulated triglyceride synthesis and mRNA expression of fatty acid synthase in vitro. In fed mice, nelfinavir or ritonavir did not affect serum glucose and cholesterol, whereas triglyceride and fatty acids increased 57% to 108%. In fasted mice, ritonavir increased serum glucose by 29%, cholesterol by 40%, and triglyceride by 99%, whereas nelfinavir had no effect, suggesting these PIs have different effects on metabolism. Consistent with the in vitro results, nelfinavir and ritonavir increased triglyceride 2- to 3-fold in fasted mice injected with Triton WR-1339, an inhibitor of triglyceride clearance. We propose that PI associated hyperlipidemia is due to increased hepatic triglyceride synthesis and suggest that retinoids or meal restriction influences the effects of select PIs on lipid metabolism. PMID- 11116064 TI - ICAM-1 deficiency reduces atherosclerotic lesions in double-knockout mice (ApoE( /-)/ICAM-1(-/-)) fed a fat or a chow diet. AB - Intercellular adhesion molecule (ICAM)-1, a major adhesion molecule, plays a critical role in the homing of leukocytes to sites of atherosclerotic lesions. However, very little is known on the role of ICAM-1 in initiating and perpetuating vascular lesions in ApoE(-/-) mice fed a chow or a fat diet. This study has investigated the mean aortic lesions in mice (C57BL6 background) with a single-knockout (ApoE(-/-)) or double-knockout (DKO; ApoE(-/-), ICAM-1(-/-)) fed a chow or a fat diet over a period of 3, 6, 15, and 20 weeks. A 3-fold reduction in lesion size was observed at all time points in DKO mice fed a chow diet. However, in DKO mice fed a fat diet, a marked reduction in the aortic lesion was observed at 3 and 15 weeks, which did not reach a significant level at 6 and 20 weeks. This study shows in essence that DKO mice are protected from developing significant lesions for up to 6 weeks when fed a chow diet and from 3 to 6 weeks when fed a fat diet. After 6 weeks, the lesion size of the DKO mice follows that of the single-knockout mice when fed a chow diet and gets to the same level in mice fed a fat diet. Plasma cholesterol levels were not altered as a result of ICAM-1 deficiency. These studies show that ICAM-1 is implicated in the formation and progression of atherosclerotic lesions. PMID- 11116065 TI - Regulation of acyl-coenzyme A:cholesterol acyltransferase (ACAT) synthesis, degradation, and translocation by high-density lipoprotein(2) at a low concentration. AB - (,Although plasma HDL(2) cholesterol concentration stands in inverse relation to risk for atherosclerotic disease, little is known about the mechanism of the apparent cardioprotection. In mouse P388D1 macrophages, HDL(2) at a low concentration (< or = 40 microg/mL) inhibits macrophage acyl-coenzyme A:cholesterol acyltransferase (ACAT), the enzyme that catalyzes esterification of intracellular cholesterol. The effects of HDL(2) on ACAT synthesis, degradation, and intracellular translocation were investigated in mouse P388D1 macrophages. HDL(2) at a low concentration enhanced ACAT synthesis but not total ACAT mass. Immunocytochemical studies showed that in the absence of lipoproteins, ACAT associated primarily with the perinuclear region of the cell. The addition of HDL(2), however, induced the transfer of ACAT to vesicular structures and the cell periphery adjacent to the plasma membrane. Subfractionation combined with immunoprecipitation complemented these observations and showed that HDL(2) promoted the transfer of ACAT to the plasma membrane fraction. Brefeldin A, which inhibits vesicular protein transport from the endoplasmic reticulum to the Golgi compartment in mammalian cells, blocked ACAT translocation and partially restored ACAT activity. These results suggest that HDL(2) is an initiating factor in a signal transduction pathway that leads to intracellular ACAT translocation and inactivation. PMID- 11116066 TI - Involvement of oxysterols and lysophosphatidylcholine in the oxidized LDL-induced impairment of serum albumin synthesis by HEPG2 cells. AB - Oxidized low density lipoproteins (Ox-LDLs) are increasingly thought to be a key element in atherogenesis. We have previously reported that serum albumin has important antioxidant properties and that a reduced synthesis of albumin may represent a crucial point in the overall antioxidant defense. In the present work, we aimed at determining whether Ox-LDL could modulate albumin synthesis in cultured human hepatocytes (HepG2 cells). With the use of enzyme immunoassay and radiolabeled leucine incorporation followed by specific immunoprecipitation, Ox LDL was found to lead to a dose-dependent decrease in albumin secretion. Moreover, the protein synthesis and mRNA levels were decreased in the presence of Ox-LDL, as assessed by Northern blot analysis. Because oxysterols and lysophospholipids are key components of Ox-LDL, we tested the effects of oxysterols (7-ketocholesterol and 25-hydroxycholesterol) and lysophosphatidylcholine on albumin secretion and expression. In our experimental conditions, we found that incubations with oxysterols or lysophosphatidylcholine at pathophysiological concentrations similar to those measured in Ox-LDLs reproduced the above-mentioned inhibitory effects on albumin synthesis. On the basis of our in vitro data, we propose that this newly described biological effect of Ox-LDL might partly explain the findings of epidemiological studies indicating that reduced levels of serum albumin are associated with increased mortality. PMID- 11116067 TI - A locus influencing total serum cholesterol on chromosome 19p: results from an autosomal genomic scan of serum lipid concentrations in Pima Indians. AB - A genome-wide linkage study was analyzed to identify loci that influence serum lipid concentrations in Pima Indians. Linkage analyses were conducted for total cholesterol measured in 998 siblings from 292 nuclear families, for total triglycerides in 547 siblings from 188 families, and for high density lipoprotein (HDL) cholesterol in 590 siblings from 201 families. Genotypes were generated for 516 autosomal microsatellite markers. Multipoint variance components methods were used to assess linkage. The strongest evidence for linkage with total cholesterol was on chromosome 19p (lod score 3.89), in the vicinity of the marker D19S1034, which is near the low density lipoprotein receptor gene. The strongest evidence for linkage with HDL cholesterol was on chromosome 3q (lod score 2.64) near D3S3053. For triglycerides, the strongest evidence for linkage was on chromosome 2p near D2S1788 (lod score 1.70) and on chromosome 3p near D3S2406 (lod score 1.77). This genomic scan provides evidence for a locus influencing total cholesterol concentration on chromosome 19p. It also suggests a locus influencing HDL cholesterol on chromosome 3q. PMID- 11116068 TI - Stromelysin-1 and interleukin-6 gene promoter polymorphisms are determinants of asymptomatic carotid artery atherosclerosis. AB - The functional 5A/6A polymorphism of the stromelysin-1 promoter has been implicated as a potential genetic marker for the progression of angiographically determined atherosclerosis in patients with coronary artery disease. Recently, a novel interleukin-6 (IL-6) gene functional G/C polymorphism at -174 in the promoter has also been reported. In this study, we analyzed the relation of these two polymorphisms with carotid artery atherosclerosis in 109 randomly selected, middle-aged men without exercise-induced ischemia. Atherosclerosis was quantified as intima-media thickness (IMT) by high-resolution ultrasonography. Univariately, stromelysin genotype was significantly (P:=0.015) associated with IMT, and this relation remained (P:=0.033) after adjustments for age, cardiorespiratory fitness, body mass index, smoking, LDL cholesterol, and systolic blood pressure and for sonographers. The 5A/6A polymorphism independently explained 7% of the variance in carotid bifurcation IMT. The IL-6 polymorphism was also significantly associated (P:=0. 036) with increased IMT, with men homozygous for the G allele having IMT that was 11% greater than men homozygous for the C allele. Men who were homozygous for both the 6A and G alleles had an covariate adjusted IMT that was 36% greater than men who were homozygous for neither allele (P:<0.003). These data suggest that genetic factors that predispose to reduced matrix remodeling (stromelysin 6A allele) and to increased inflammation (IL-6 G allele) combine to increase susceptibility for intima-media thickening in the carotid bifurcation, a predilection site for atherosclerosis. PMID- 11116069 TI - Contribution of apolipoprotein C-III gene variants to determination of triglyceride levels and interaction with smoking in middle-aged men. AB - Variation within and around the apolipoprotein C-III (APOC3) gene has been associated with elevated triglyceride (Tg) levels and cardiovascular disease. The associations of 4 polymorphic variants in the APOC3 gene (3238C>G in the 3' untranslated region [SST:I], 1100C>T in exon 3, -482C>T in the insulin-responsive element, and -2854T>G in the APOC3-A4 intergenic region) with plasma Tg and cholesterol levels and their interaction with smoking have been investigated in the Second Northwick Park Heart Study (NPHSII), a large cohort of healthy men (n=2745). Analyzing the variants separately showed that 3238G, 1100T, and -482T alleles were all associated with raised Tg levels. For the 3238C>G and -482C>T sites, the Tg-raising effect appeared to depend on smoking status (test for interaction, P:=0.042 and P:=0.009, respectively), but for the 1100C>T site, the effect was constant irrespective of smoking status (test for interaction, P:=0.27). The -2854T>G site was not associated with effects on Tg levels in this sample. Because all of the variants showed significant allelic association, regression modeling was used to quantify the relative size of each effect and to assess whether the effects of the separate variants were independent. The 1100C>T variant had an independent effect on Tg levels that was not influenced by smoking status (increase of 8.2% in Tg with each T1100 allele), whereas the -482C>T variant had a separate effect that was dependent on smoking (increase of 13.7% in Tg for each -482T allele in current smokers, 8.6% in exsmokers, and -7.4% in those who never smoked). The 3238C>G variant did not show a separate independent effect on Tg concentration. Thus, by use of the regression model, it was possible to estimate how mean Tg levels would vary in groups of individuals with respect to APOC3 genotype and smoking information. Analysis in this large group of healthy men has allowed the identification of a statistically robust APOC3 genotype-smoking interaction, which now warrants further molecular study. PMID- 11116070 TI - Distribution of ApoA-I-containing HDL subpopulations in patients with coronary heart disease. AB - High density lipoproteins (HDLs) and their subspecies play a role in the development of coronary heart disease (CHD). HDL subpopulations were measured by 2-dimensional nondenaturing gel electrophoresis in 79 male control subjects and 76 male CHD patients to test the hypothesis that greater differences in apolipoprotein (apo)A-I-containing HDL subpopulations would exist between these 2 groups than for traditional lipid levels. In CHD subjects, HDL cholesterol (HDL C) was lower (-14%, P<0.001), whereas total cholesterol and the low density lipoprotein cholesterol/HDL-C ratio were higher (9% [P:<0.05] and 21% [P:<0.01], respectively) compared with control levels. No significant differences were found for low density lipoprotein cholesterol, triglyceride, and apoA-I levels. In CHD subjects, there were significantly (P:<0.001) lower concentrations of the large lipoprotein (Lp)A-I alpha(1) (-35%), pre-alpha(1) (-50%), pre-alpha(2) (-33%), and pre-alpha(3) (-31%) subpopulations, whereas the concentrations of the small LpA-I/A-II alpha(3) particles were significantly (P:<0.001) higher (20%). Because alpha(1) was decreased more than HDL-C and plasma apoA-I concentrations in CHD subjects, the ratios of HDL-C to alpha(1) and of apoA-I to alpha(1) were significantly (P:<0.001) higher by 36% and 57%, respectively, compared with control values. Subjects with low HDL-C levels (35 mg/dL). Therefore, we stratified participants according to HDL-C concentrations into low and normal groups. The differences in lipid levels between controls and HDL-C-matched cases substantially decreased; however, the significant differences in HDL subspecies remained. Our research findings support the concept that compared with control subjects, CHD patients not only have HDL deficiency but also have a major rearrangement in the HDL subpopulations with significantly lower alpha(1) and pre-alpha(1-3) (LpA-I) and significantly higher alpha(3) (LpA-I/A-II) particles. PMID- 11116071 TI - Low plasma lycopene concentration is associated with increased intima-media thickness of the carotid artery wall. AB - Although a number of epidemiological studies have evaluated the association between ss-carotene and the risk of cardiovascular diseases, there has been little research on the role of lycopene, an acyclic form of ss-carotene, with regard to the risk of cardiovascular disease. We investigated the relationship between plasma concentrations of lycopene and intima-media thickness of the common carotid artery wall (CCA-IMT) in 520 middle-aged men and women (aged 45 to 69 years) in eastern Finland. They were examined from 1994 to 1995 at the baseline of the Antioxidant Supplementation in Atherosclerosis Prevention (ASAP) study, a randomized trial concerning the effect of vitamin E and C supplementation on atherosclerotic progression. The subjects were classified into 2 categories according to the median concentration of plasma lycopene (0.12 micromol/L in men and 0.15 micromol/L in women). Mean CCA-IMT of the right and left common carotid arteries was 1.18 mm in men and 0.95 mm in women with plasma lycopene levels lower than the median and 0.97 mm in men (P:<0.001 for difference) and 0.89 mm in women (P:=0.027 for difference) with higher levels of plasma lycopene. In ANCOVA adjusting for cardiovascular risk factors and intake of nutrients, in men, low levels of plasma lycopene were associated with a 17.8% increment in CCA-IMT (P:=0.003 for difference). In women, the difference did not remain significant after the adjustments. We conclude that low plasma lycopene concentrations are associated with early atherosclerosis, manifested as increased CCA-IMT, in middle-aged men living in eastern Finland. PMID- 11116072 TI - Quantification of cholesteryl esters in human and rabbit atherosclerotic plaques by magic-angle spinning (13)C-NMR. AB - Accumulation of cholesteryl esters (CEs) is a key event in the formation of atherosclerotic plaques. More recent work suggests a role for CEs in plaque rupture leading to thrombosis, which can result in an acute event such as myocardial infarction or stroke. In this study, we present nuclear magnetic resonance (NMR) protocols for quantification of CEs in plaques in situ. Total CEs quantified by (13)C magic-angle spinning (MAS) NMR in excised plaques from human carotid arteries and rabbit aortic arteries were in good agreement with the amounts determined by subsequent standard chemical assays. The latter analysis is disadvantageous because it requires that plaque lipids be extracted from the tissue, resulting in the loss of all phase information of CEs as well as other major plaque components. With our MAS-NMR protocol, the plaque components are preserved in their native phases. Combining MAS and off-MAS NMR, we were able to quantitatively distinguish isotropic (liquid) CEs from anisotropic (liquid crystalline) CEs in plaque tissues. In a recent study, we applied a different (13)C MAS-NMR protocol to quantify crystalline cholesterol monohydrate in plaques. Together, these 2 studies describe a new, noninvasive MAS-NMR strategy for the identification and quantification of the major lipid components in plaques in situ. This approach will be useful for investigation of the relationship between plaque rupture and specific lipids in their biologically relevant phases. PMID- 11116073 TI - Genetic induction of a releasable pool of factor VIII in human endothelial cells. AB - Although it is known that factor VIII (FVIII) plasma levels increase rapidly in response to a number of stimuli, the biological stimuli behind this release is less clear. Previously, we showed that FVIII can traffic together with von Willebrand factor (vWF) into storage granules in a pituitary tumor cell line, AtT 20; however, AtT-20 cells could not be used to address the release or functional activity of released FVIII. To investigate the regulated secretion of stored FVIII, endothelial cells with intact agonist-stimulated release pathways were used. Human umbilical vein endothelial cells (HUVECs) were transduced with retroviral FVIII construct [hFVIII(V)] to create a FVIII/vWF storage pool. Immunofluorescent staining of transduced cells demonstrated FVIII in Weibel Palade bodies. In contrast, the transduction of hFVIII(V) into HT-1080 and HepG2 cells displayed FVIII only in the cytoplasm. We studied the regulated release of both FVIII and vWF from endothelial cells after agonist-induced stimulation and demonstrated a parallel release of FVIII and vWF proteins. This released FVIII was functionally active. Hence, endothelial cells transduced with hFVIII(V) store FVIII together with vWF in Weibel-Palade bodies, creating a releasable storage pool of both proteins. Because FVIII secretion can be physiologically regulated by agonists in culture, this may explain the pharmacological agonist-induced release of FVIII by drugs such as desmopressin in vivo and suggests vascular endothelium as a reasonable target of gene therapy of hemophilia A. PMID- 11116074 TI - Identification and localization of a fatty acid response region in the human plasminogen activator inhibitor-1 gene. AB - The increased expression of plasminogen activator inhibitor type-1 (PAI-1) is associated with increased concentrations of fatty acids in blood and may accelerate atherogenesis in diabetes. The present study was designed to define mechanisms by which nonesterified (free) fatty acids (FFAs) augment the expression of PAI-1. FFAs increased PAI-1 protein and mRNA expression by HepG2 cells. To identify potential regulatory elements, we constructed chimeric genes by fusing 1313, 853, 610, or 328 bp of human PAI-1 5'-flanking DNA to a luciferase reporter (PAI-LUC). A 2-fold increase in luciferase activity was seen when cells were transfected with PAI-LUC 1313, 863, or 610 and exposed to FFAs. No response to FFAs was seen with PAI-LUC 328 and after deletion of a 72-bp (-599 to -528) fragment from PAI-LUC 1313. This 72-bp fragment conferred FFA responsiveness to a different (simian virus 40) promoter. Two footprinted regions were demonstrated by DNase I analysis. Gel mobility shift assays indicated specific binding of extracted proteins to an FFA response element: 5'-TG(G/C)(1 2)CTG-3'. This sequence is repeated 4 times and is similar to an Sp1-binding site. Sp1 consensus oligonucleotides inhibited binding of extracted proteins to the regulatory elements. Accordingly, FFA-induced increased expression of PAI-1 in HepG2 cells is mediated by the binding of a transcription factor or factors to the repeated fatty acid response element, 5'-TG(G/C)(1-2)CTG-3', that is highly homologous to an Sp1-binding site. PMID- 11116075 TI - Platelet-leukocyte cross talk in whole blood. AB - Thrombosis and inflammation involve complex platelet-leukocyte interaction, the details of which are not fully elucidated. Therefore, we investigated cross talk between platelets and leukocytes in whole blood, under the following physiological conditions: at 37 degrees C, with normal calcium concentrations, and with shear force. Platelet P-selectin and leukocyte CD11b expression were used to monitor platelet and leukocyte activation, respectively, and platelet leukocyte aggregation (PLA) was analyzed. The leukocyte-specific agonist N: formyl-methionyl-leucyl-phenylalanine (10(-6) mol/L) increased P-selectin positive platelets from 2.5+/-0. 1% to 5.1+/-0.6% (P:<0.05). The increase was inhibited by either the platelet-activating factor (PAF) antagonist SR27417, the superoxide anion scavenger superoxide dismutase, the 5-lipoxygenase inhibitor Zileuton, or the 5-lipoxygenase-activating protein inhibitor MK-886, suggesting the involvement of PAF, superoxide anion, and 5-lipoxygenase products in leukocyte-induced platelet activation. The platelet-specific agonist collagen (1 microg/mL) increased leukocyte CD11b expression from 2.94+/-0.52 to 3.81+/-0.58 (P:<0. 05); this was not inhibited by the thromboxane A(2) receptor antagonist ICI 192.605 or the PAF antagonist SR27417. Platelet P-selectin expression induced by N:-formyl-methionyl-leucyl-phenylalanine and leukocyte CD11b expression induced by collagen could be suppressed by glycoprotein IIb/IIIa blockade or P selectin blockade. This study documents platelet-leukocyte cross talk under conditions that mimic a physiological state and suggests that this involves multiple mediators and mechanisms. Furthermore, new evidence of integrin and selectin involvement in intracellular and intercellular signaling during platelet leukocyte cross talk is provided. PMID- 11116076 TI - Role of matrix metalloproteinases in blood flow-induced arterial enlargement: interaction with NO. AB - Tears in the internal elastic lamina (IEL) can be observed after chronic increases in arterial blood flow, suggesting a potential role for matrix metalloproteinases (MMPs) in flow-induced vascular remodeling. We undertook to study this phenomenon by constructing an arteriovenous fistula (AVF) between the left common carotid artery (CCA) and the external jugular vein in rabbits. The diameter of the flow-loaded left CCA increased by 13.6+/-1.8% by day 3 after construction of the AVF compared with the right CCA (n=4, P:<0.01) and by 40.7+/ 7.5% by day-15 (n=10, P:<0.0001). Increased CCA diameter also coincided with IEL fragmentation. Three days after construction of the AVF, gelatin zymography of protein extracts from left CCAs of untreated rabbits showed a significant increase in the 62-kDa (active MMP-2) activity and the appearance of a lytic band at 92 kDa (pro-MMP-9). In further experiments, MMP activity was inhibited by treatment with doxycycline (DOX) or BB-94, a specific MMP inhibitor. The increase in the 62-kDa gelatinolytic band was abolished in DOX- and BB-94-treated rabbits. The 92-kDa gelatinolytic band was also reduced in DOX-treated animals. Furthermore, both increased left CCA diameter and IEL fragmentation were abolished in DOX- and BB-94-treated rabbits. To evaluate whether nitric oxide was involved in blood flow-induced MMP activation, the rabbits were treated with N:(G)-nitro-L-arginine methyl ester to inhibit nitric oxide synthesis. MMP activities were significantly decreased in the left CCAs of N:(G)-nitro-L arginine methyl ester-treated animals. Hence, blood flow-induced MMP activation is critical in flow-induced vascular enlargement and IEL fragmentation, and blood flow-induced nitric oxide participates in MMP activation. PMID- 11116077 TI - Role of RhoA and Rho kinase in lysophosphatidic acid-induced endothelial barrier dysfunction. AB - In the present study, the roles of the small GTPase RhoA and its target Rho kinase in endothelial permeability were investigated in vitro. We have shown previously that, in addition to a rise in the intracellular Ca(2+) concentration ([Ca(2+)](i)), RhoA is involved in the prolonged thrombin-induced barrier dysfunction. To study the role of RhoA and Rho kinase more specifically, endothelial cells were stimulated with lysophosphatidic acid (LPA), a commonly used RhoA activator. LPA induced a 2- to 3-fold increase in the passage of horseradish peroxidase (HRP) across endothelial monolayers that lasted for several hours, whereas thrombin induced a 5- to 10-fold increase. Comparable to the thrombin-induced barrier dysfunction, the LPA-induced barrier dysfunction was accompanied by a reorganization of the F-actin cytoskeleton and the formation of focal attachment sites. LPA induced only a transient increase in myosin light chain (MLC) phosphorylation, which returned to basal level within 10 minutes. In endothelial cells, [Ca(2+)](i) was not elevated by LPA. Chelation of Ca(2+)(i) ions by 1, 2-bis(2-aminophenoxy)ethane-N:,N:,N:',N:'-tetraacetic acid did not prevent the LPA-induced passage of HRP. Apparently, a low degree of MLC kinase activation occurred, because the MLC kinase inhibitor KT5926 reduced the levels of both basal and LPA-stimulated HRP passage. Inhibition of RhoA by the C3 transferase from Clostridium botulinum inhibited the LPA-induced cytoskeletal changes and prevented the LPA-induced HRP passage. Inhibition of Rho kinase by Y 27632 completely prevented the LPA-induced increase in HRP passage without affecting basal permeability. These data indicate that LPA-induced endothelial hyperpermeability occurs without a change in [Ca(2+)](i) and requires activation of RhoA and Rho kinase. PMID- 11116078 TI - Inhibition of mast cell-dependent conversion of cultured macrophages into foam cells with antiallergic drugs. AB - Degranulation of isolated, rat peritoneal mast cells in the presence of low density lipoprotein (LDL) induces cholesteryl ester accumulation in cocultured macrophages with ensuing foam cell formation. This event occurs when the macrophages phagocytose LDL particles that have been bound to the heparin proteoglycans of exocytosed granules. In an attempt to inhibit such foam cell formation pharmacologically, rat peritoneal mast cells that had been passively sensitized with anti-ovalbumin-IgE were treated with 2 mast cell-stabilizing antianaphylactic drugs, MY-1250 or disodium cromoglycate (DSCG). Both drugs were found to inhibit antigen (ovalbumin)-triggered release of histamine from the mast cells, revealing mast cell stabilization. In cocultures of rat peritoneal macrophages and passively sensitized mast cells, addition of MY-1250 before addition of the antigen resulted in parallel reductions in histamine release from mast cells, uptake of [(14)C]sucrose-LDL, and accumulation of LDL-derived cholesteryl esters in the cocultured macrophages. Similarly, when passively sensitized mast cells were stimulated with antigen in the presence of DSCG and the preconditioned media containing all substances released from the drug-treated mast cells were collected and added to macrophages cultured in LDL-containing medium, uptake and esterification of LDL cholesterol by the macrophages were inhibited. The inhibitory effects of both drugs were mast cell-specific because neither drug inhibited the ability of macrophages to take up and esterify LDL cholesterol. Analysis of heparin proteoglycan contents of the incubation media revealed that both drugs had inhibited mast cells from expelling their granule remnants. Thus, both MY-1250 and DSCG prevent mast cells from releasing the heparin proteoglycan-containing vehicles that bind LDL and carry it into macrophages. This study suggests that antiallergic pharmacological agents could be used in animal models to prevent mast cell-dependent formation of foam cells in vivo. PMID- 11116079 TI - Keeping the serpin machine running smoothly. PMID- 11116080 TI - Tools for the population genomics of the tubercle bacilli. PMID- 11116081 TI - Doubling the rewards: testis ESTs for Drosophila gene discovery and spermatogenesis expression profile analysis. PMID- 11116082 TI - Phylogeny of the serpin superfamily: implications of patterns of amino acid conservation for structure and function. AB - We present a comprehensive alignment and phylogenetic analysis of the serpins, a superfamily of proteins with known members in higher animals, nematodes, insects, plants, and viruses. We analyze, compare, and classify 219 proteins representative of eight major and eight minor subfamilies, using a novel technique of consensus analysis. Patterns of sequence conservation characterize the family as a whole, with a clear relationship to the mechanism of function. Variations of these patterns within phylogenetically distinct groups can be correlated with the divergence of structure and function. The goals of this work are to provide a carefully curated alignment of serpin sequences, to describe patterns of conservation and divergence, and to derive a phylogenetic tree expressing the relationships among the members of this family. We extend earlier studies by Huber and Carrell as well as by Marshall, after whose publication the serpin family has grown functionally, taxonomically, and structurally. We used gene and protein sequence data, crystal structures, and chromosomal location where available. The results illuminate structure-function relationships in serpins, suggesting roles for conserved residues in the mechanism of conformational change. The phylogeny provides a rational evolutionary framework to classify serpins and enables identification of conserved amino acids. Patterns of conservation also provide an initial point of comparison for genes identified by the various genome projects. New homologs emerging from sequencing projects can either take their place within the current classification or, if necessary, extend it. PMID- 11116083 TI - Fine-resolution physical mapping of genomic diversity in Candida albicans. AB - It has been suggested that Candida albicans, a diploid asexual fungus, achieves genetic diversity by genomic rearrangement. This important human pathogen may provide a system in which to analyze alternate routes to genomic diversity. C. albicans has a highly variable karyotype; its chromosomes contain a middle repeated DNA sequence called the Major Repeat Sequence (MRS), composed of subrepeats HOK, RPS, and RB2. RPS is tandemly repeated while the other subrepeats occur once in each MRS. Chromosome 7, the smallest of the eight chromosomes, has been previously mapped. The complete physical map of this chromosome was used to analyze chromosome 7 diversity in six strains, including two well-characterized laboratory strains (1006 and WO-1) and four clinical ones. We found four types of events to explain the genomic diversity: 1) Chromosome length polymorphism (CLP) results from expansion and contraction of the RPS; 2) reciprocal translocation occurs at the MRS loci; 3) chromosomal deletion; and (4) trisomy of individual chromosomes. These four phenomena play an important role in generating genomic diversity in C. albicans. PMID- 11116084 TI - Comparative genome analysis of the mouse imprinted gene impact and its nonimprinted human homolog IMPACT: toward the structural basis for species specific imprinting. AB - Mouse Impact is a paternally expressed gene encoding an evolutionarily conserved protein of unknown function. Here we identified IMPACT, the human homolog of Impact, on chromosome 18q11. 2-12.1, a region syntenic to the mouse Impact locus. IMPACT was expressed biallelically in brain and in various tissues from two informative fetuses and in peripheral blood from an informative adult. To reveal the structural basis for the difference in allelic expression between the two species, we elucidated complete genome sequences for both mouse Impact ( approximately 38 kb) and human IMPACT ( approximately 30 kb). Sequence comparison revealed that the two genes share a well-conserved exon-intron organization but bear significantly different CpG islands. The mouse island lies in the first intron and contains characteristic tandem repeats. Furthermore, this island serves as a differentially methylated region (DMR) consisting of a hypermethylated maternal allele and an unmethylated paternal allele. Intriguingly, this intronic island is missing from the nonimprinted human IMPACT, whose sole CpG island spans the first exon, lacks any apparent repeats, and escapes methylation on both chromosomes. These results suggest that the intronic DMR plays a role in the imprinting of Impact. PMID- 11116085 TI - Zebrafish comparative genomics and the origins of vertebrate chromosomes. AB - To help understand mechanisms of vertebrate genome evolution, we have compared zebrafish and tetrapod gene maps. It has been suggested that translocations are fixed more frequently than inversions in mammals. Gene maps showed that blocks of conserved syntenies between zebrafish and humans were large, but gene orders were frequently inverted and transposed. This shows that intrachromosomal rearrangements have been fixed more frequently than translocations. Duplicated chromosome segments suggest that a genome duplication occurred in ray-fin phylogeny, and comparative studies suggest that this event happened deep in the ancestry of teleost fish. Consideration of duplicate chromosome segments shows that at least 20% of duplicated gene pairs may be retained from this event. Despite genome duplication, zebrafish and humans have about the same number of chromosomes, and zebrafish chromosomes are mosaically orthologous to several human chromosomes. Is this because of an excess of chromosome fissions in the human lineage or an excess of chromosome fusions in the zebrafish lineage? Comparative analysis suggests that an excess of chromosome fissions in the tetrapod lineage may account for chromosome numbers and provides histories for several human chromosomes. PMID- 11116086 TI - A comparative map of the zebrafish genome. AB - Zebrafish mutations define the functions of hundreds of essential genes in the vertebrate genome. To accelerate the molecular analysis of zebrafish mutations and to facilitate comparisons among the genomes of zebrafish and other vertebrates, we used a homozygous diploid meiotic mapping panel to localize polymorphisms in 691 previously unmapped genes and expressed sequence tags (ESTs). Together with earlier efforts, this work raises the total number of markers scored in the mapping panel to 2119, including 1503 genes and ESTs and 616 previously characterized simple-sequence length polymorphisms. Sequence analysis of zebrafish genes mapped in this study and in prior work identified putative human orthologs for 804 zebrafish genes and ESTs. Map comparisons revealed 139 new conserved syntenies, in which two or more genes are on the same chromosome in zebrafish and human. Although some conserved syntenies are quite large, there were changes in gene order within conserved groups, apparently reflecting the relatively frequent occurrence of inversions and other intrachromosomal rearrangements since the divergence of teleost and tetrapod ancestors. Comparative mapping also shows that there is not a one-to-one correspondence between zebrafish and human chromosomes. Mapping of duplicate gene pairs identified segments of 20 linkage groups that may have arisen during a genome duplication that occurred early in the evolution of teleosts after the divergence of teleost and mammalian ancestors. This comparative map will accelerate the molecular analysis of zebrafish mutations and enhance the understanding of the evolution of the vertebrate genome. PMID- 11116087 TI - Identification, characterization, and mapping of expressed sequence tags from an embryonic zebrafish heart cDNA library. AB - The generation of expressed sequence tags (ESTs) has proven to be a rapid and economical approach by which to identify and characterize expressed genes. We generated 5102 ESTs from a 3-d-old embryonic zebrafish heart cDNA library. Of these, 57.6% matched to known genes, 14.2% matched only to other ESTs, and 27.8% showed no match to any ESTs or known genes. Clustering of all ESTs identified 359 unique clusters comprising 1771 ESTs, whereas the remaining 3331 ESTs did not cluster. This estimates the number of unique genes identified in the data set to be approximately 3690. A total of 1242 unique known genes were used to analyze the gene expression patterns in the zebrafish embryonic heart. These were categorized into seven categories on the basis of gene function. The largest class of genes represented those involved in gene/protein expression (25.9% of known transcripts). This class was followed by genes involved in metabolism (18.7%), cell structure/motility (16.4%), cell signaling and communication (9.6%), cell/organism defense (7.1%), and cell division (4.4%). Unclassified genes constituted the remaining 17.91%. Radiation hybrid mapping was performed for 102 ESTs and comparison of map positions between zebrafish and human identified new synteny groups. Continued comparative analysis will be useful in defining the boundaries of conserved chromosome segments between zebrafish and humans, which will facilitate the transfer of genetic information between the two organisms and improve our understanding of vertebrate evolution. PMID- 11116088 TI - The mouse Clock locus: sequence and comparative analysis of 204 kb from mouse chromosome 5. AB - The Clock gene encodes a basic helix-loop-helix (bHLH)-PAS transcription factor that regulates circadian rhythms in mice. We previously cloned Clock in mouse and human using a battery of behavioral and molecular techniques, including shotgun sequencing of two bacterial artificial chromosome (BAC) clones. Here we report the finished sequence of a 204-kb region from mouse chromosome 5. This region contains the complete loci for the Clock and Tpardl (pFT27) genes, as well as the 3' partial locus of the Neuromedin U gene; sequence analysis also suggests the presence of two previously unidentified genes. In addition, we provide a comparative genomic sequence analysis with the syntenic region from human chromosome 4. Finally, a new BAC transgenic line indicates that the genomic region that is sufficient for rescue of the Clock mutant phenotype is no greater than 120 kb and tightly flanks the 3' end of the Clock gene. PMID- 11116089 TI - Single QTL effects, epistasis, and pleiotropy account for two-thirds of the phenotypic F(2) variance of growth and obesity in DU6i x DBA/2 mice. AB - Genes influencing body weight and composition and serum concentrations of leptin, insulin, and insulin-like growth factor I (IGF-I) in nonfasting animals were mapped in an intercross of the extreme high-growth mouse line DU6i and the inbred line DBA/2. Significant loci with major effects (F > 7.07) for body weight, obesity, and muscle weight were found on chromosomes 1, 4, 5, 7, 11, 12, 13, and 17, for leptin on chromosome 14, for insulin on chromosome 4, and for IGF-I on chromosome 10 at the Igf1 gene locus itself and on chromosome 18. Significant interaction between different quantitative trait loci (QTL) positions was observed (P < 0.01). Evidence was found that loci having small direct effect on growth or obesity contribute to the obese phenotype by gene-gene interaction. The effects of QTLs, epistasis, and pleiotropy account for 64% and 63% of the phenotypic variance of body weight and fat accumulation and for over 32% of muscle weight and serum concentrations of leptin, and IGF-I in the F(2) population of DU6i x DBA/2 mice. [The quantitative trait loci described in this paper have been submitted to the Mouse Genome Database.] PMID- 11116090 TI - Sequence diversity and genomic organization of vomeronasal receptor genes in the mouse. AB - The vomeronasal system of mice is thought to be specialized in the detection of pheromones. Two multigene families have been identified that encode proteins with seven putative transmembrane domains and that are expressed selectively in subsets of neurons of the vomeronasal organ. The products of these vomeronasal receptor (Vr) genes are regarded as candidate pheromone receptors. Little is known about their genomic organization and sequence diversity, and only five sequences of mouse V1r coding regions are publicly available. Here, we have begun to characterize systematically the V1r repertoire in the mouse. We isolated 107 bacterial artificial chromosomes (BACs) containing V1r genes from a 129 mouse library. Hybridization experiments indicate that at least 107 V1r-like sequences reside on these BACs. We assembled most of the BACs into six contigs, of which one major contig and one minor contig were characterized in detail. The major contig is 630-860 kb long, encompasses a cluster of 21-48 V1r genes, and contains marker D6Mit227. Sequencing of the coding regions was facilitated by the absence of introns. We determined the sequence of the coding region of 25 possibly functional V1r genes and seven pseudogenes. The functional V1rs can be arranged into three groups; V1rs of one group are novel and substantially divergent from the other V1rs. The genomic and sequence information described here should be useful in defining the biological function of these receptors. PMID- 11116091 TI - MHC-linked olfactory receptor loci exhibit polymorphism and contribute to extended HLA/OR-haplotypes. AB - Clusters of olfactory receptor (OR) genes are found on most human chromosomes. They are one of the largest mammalian multigene families. Here, we report a systematic study of polymorphism of OR genes belonging to the largest fully sequenced OR cluster. The cluster contains 36 OR genes, of which two belong to the vomeronasal 1 (V1-OR) family. The cluster is divided into a major and a minor region at the telomeric end of the HLA complex on chromosome 6. These OR genes could be involved in MHC-related mate preferences. The polymorphism screen was carried out with 13 genes from the HLA-linked OR cluster and three genes from chromosomes 7, 17, and 19 as controls. Ten human cell lines, representing 18 different chromosome 6s, were analyzed. They were from various ethnic origins and exhibited different HLA haplotypes. All OR genes tested, including those not linked to the HLA complex, were polymorphic. These polymorphisms were dispersed along the coding region and resulted in up to seven alleles for a given OR gene. Three polymorphisms resulted either in stop codons (genes hs6M1-4P, hs6M1-17) or in a 16-bp deletion (gene hs6M1-19P), possibly leading to lack of ligand recognition by the respective receptors in the cell line donors. In total, 13 HLA linked OR haplotypes could be defined. Therefore, allelic variation appears to be a general feature of human OR genes. PMID- 11116092 TI - Characterization of nonfunctional V1R-like pheromone receptor sequences in human. AB - The vomeronasal organ (VNO) or Jacobson's organ is responsible in terrestrial vertebrates for the sensory perception of pheromones, chemicals that elicit stereotyped behaviors among individuals of the same species. Pheromone-induced behaviors and a functional VNO have been described in a number of mammals, but the existence of this sensory system in human is still debated. Recently, two nonhomologous gene families, V1R and V2R, encoding pheromone receptors have been identified in rat. These receptors belong to the seven-transmembrane domain G protein-coupled receptor superfamily. We sought to characterize V1R-like genes in the human genome. We have identified seven different human sequences by PCR and library screening with rodent sequences. These human sequences exhibit characteristic features of V1R receptors and show 52%-59% of amino acid sequence identity with the rat sequences. Using PCR on a monochromosomal somatic cell hybrid panel and/or FISH, we demonstrate that these V1R-like sequences are distributed on chromosomes 7, 16, 20, 13, 14, 15, 21, and 22 and possibly on additional chromosomes. One sequence hybridizes to pericentromeric locations on all the acrocentric chromosomes (13, 14, 15, 21, and 22). All of the seven V1R like sequences analyzed show interrupted reading frames, indicating that they represent nonfunctional pseudogenes. The preponderence of pseudogenes among human V1R sequences and the striking anatomical differences between rodent and human VNO raise the possibility that humans may have lost the V1R/VNO-mediated sensory functions of rodents. PMID- 11116093 TI - Assessment of compositional heterogeneity within and between eukaryotic genomes. AB - Using large amounts of long genomic sequences, we studied the compositional patterns of eukaryotic genomes. We developed a simple measure, the compositional heterogeneity (or variability) index, to compare the differences in compositional heterogeneity between long genomic sequences. The index measures the average difference in GC content between two adjacent windows normalized by the standard error expected under the assumption of random distribution of nucleotides in a window. We report the following findings: (1) The extent of the compositional heterogeneity in a genomic sequence strongly correlates with its GC content in all multicellular eukaryotes studied regardless of genome size. (2) The human genome appears to be highly compositionally heterogeneous both within and between individual chromosomes; the heterogeneity goes much beyond the predictions of the isochore model. (3) All genomes of multicellular eukaryotes examined in this study are compositionally heterogeneous, although they also contain compositionally uniform segments, or isochores. (4) The true uniqueness of the human (or mammalian) genome is the presence of very high GC regions, which exhibit unusually high compositional heterogeneity and contain few long homogeneous segments (isochores). In general, GC-poor isochores tend to be longer than GC-rich ones. These findings indicate that the genomes of multicellular organisms are much more heterogeneous in nucleotide composition than depicted by the isochore model and so lead to a looser definition of isochores. PMID- 11116094 TI - A random sequencing approach for the analysis of the Trypanosoma cruzi genome: general structure, large gene and repetitive DNA families, and gene discovery. AB - A random sequence survey of the genome of Trypanosoma cruzi, the agent of Chagas disease, was performed and 11,459 genomic sequences were obtained, resulting in approximately 4.3 Mb of readable sequences or approximately 10% of the parasite haploid genome. The estimated total GC content was 50.9%, with a high representation of A and T di- and trinucleotide repeats. Out of the estimated 5000 parasite genes, 947 putative new genes were identified. Another 1723 sequences corresponded to genes detected previously in T. cruzi through expression sequence tag analysis. 7735 sequences had no matches in the database, but the presence of open reading frames that passed Fickett's test suggests that some might contain coding DNA. The survey was highly redundant, with approximately 35% of the sequences included in a few large sequence families. Some of them code for protein families present in dozens of copies, including proteins essential for parasite survival and retrotransposons. Other sequence families include repetitive DNA present in thousands of copies per haploid genome. Some families in the latter group are new, parasite-specific, repetitive DNAs. These results suggest that T. cruzi could constitute an interesting model to analyze gene and genome evolution due to its plasticity in terms of sequence amplification and divergence. Additional information can be found at http://www.iib.unsam.edu.ar/tcruzi.gss. html. PMID- 11116095 TI - The mouse brain transcriptome by SAGE: differences in gene expression between P30 brains of the partial trisomy 16 mouse model of Down syndrome (Ts65Dn) and normals. AB - Trisomy 21, or Down syndrome (DS), is the most common genetic cause of mental retardation. Changes in the neuropathology, neurochemistry, neurophysiology, and neuropharmacology of DS patients' brains indicate that there is probably abnormal development and maintenance of central nervous system structure and function. The segmental trisomy mouse (Ts65Dn) is a model of DS that shows analogous neurobehavioral defects. We have studied the global gene expression profiles of normal and Ts65Dn male and normal female mice brains (P30) using the serial analysis of gene expression (SAGE) technique. From the combined sample we collected a total of 152,791 RNA tags and observed 45,856 unique tags in the mouse brain transcriptome. There are 14 ribosomal protein genes (nine under expressed) among the 330 statistically significant differences between normal male and Ts65Dn male brains, which possibly implies abnormal ribosomal biogenesis in the development and maintenance of DS phenotypes. This study contributes to the establishment of a mouse brain transcriptome and provides the first overall analysis of the differences in gene expression in aneuploid versus normal mammalian brain cells. PMID- 11116096 TI - Microarray expression profiling identifies genes with altered expression in HDL deficient mice. AB - Based on the assumption that severe alterations in the expression of genes known to be involved in high-density lipoprotein (HDL) metabolism may affect the expression of other genes, we screened an array of >5000 mouse expressed sequence tags for altered gene expression in the livers of two lines of mice with dramatic decreases in HDL plasma concentrations. Labeled cDNA from livers of apolipoprotein AI (apoAI)-knockout mice, scavenger receptor BI (SR-BI) transgenic mice, and control mice were cohybridized to microarrays. Two-sample t statistics were used to identify genes with altered expression levels in the knockout or transgenic mice compared with control mice. In the SR-BI group we found nine array elements representing at least five genes that were significantly altered on the basis of an adjusted P value < 0.05. In the apoAI-knockout group, eight array elements representing four genes were altered compared with the control group (adjusted P < 0.05). Several of the genes identified in the SR-BI transgenic suggest altered sterol metabolism and oxidative processes. These studies illustrate the use of multiple-testing methods for the identification of genes with altered expression in replicated microarray experiments. PMID- 11116098 TI - Detection of deleted genomic DNA using a semiautomated computational analysis of GeneChip data. AB - Genomic diversity within and between populations is caused by single nucleotide mutations, changes in repetitive DNA systems, recombination mechanisms, and insertion and deletion events. The contribution of these sources to diversity, whether purely genetic or of phenotypic consequence, can only be investigated if we have the means to quantitate and characterize diversity in many samples. With the advent of complete sequence characterization of representative genomes of different species, the possibility of developing protocols to screen for genetic polymorphism across entire genomes is actively being pursued. The large numbers of measurements such approaches yield demand that we pay careful attention to the numerical analysis of data. In this paper we present a novel application of an Affymetrix GeneChip to perform genome-wide screens for deletion polymorphism. A high-density oligonucleotide array formatted for mRNA expression and targeted at a fully sequenced 4.4-million-base pair Mycobacterium tuberculosis standard strain genome was adapted to compare genomic DNA. Hybridization intensities to 111,000 probe pairs (perfect complement and mismatch complement) were measured for genomic DNA from a clinical strain and from a vaccine organism. Because individual probe-pair hybridization intensities exhibit limited sensitivity/specificity characteristics to detect deletions, data-analytical methodology to exploit measurements from multiple probes in tandem locations across the genome was developed. The TSTEP (Tandem Set Terminal Extreme Probability) algorithm designed specifically to analyze the tandem hybridization measurements data was applied and shown to discover genomic deletions with high sensitivity. The TSTEP algorithm provides a foundation for similar efforts to characterize deletions in many hybridization measures in similar-sized and larger genomes. Issues relating to the design of genome content screening experiments and the implications of these methods for studying population genomics and the evolution of genomes are discussed. PMID- 11116097 TI - Gene discovery using computational and microarray analysis of transcription in the Drosophila melanogaster testis. AB - Identification and annotation of all the genes in the sequenced Drosophila genome is a work in progress. Wild-type testis function requires many genes and is thus of potentially high value for the identification of transcription units. We therefore undertook a survey of the repertoire of genes expressed in the Drosophila testis by computational and microarray analysis. We generated 3141 high-quality testis expressed sequence tags (ESTs). Testis ESTs computationally collapsed into 1560 cDNA set used for further analysis. Of those, 11% correspond to named genes, and 33% provide biological evidence for a predicted gene. A surprising 47% fail to align with existing ESTs and 16% with predicted genes in the current genome release. EST frequency and microarray expression profiles indicate that the testis mRNA population is highly complex and shows an extended range of transcript abundance. Furthermore, >80% of the genes expressed in the testis showed onefold overexpression relative to ovaries, or gonadectomized flies. Additionally, >3% showed more than threefold overexpression at p <0.05. Surprisingly, 22% of the genes most highly overexpressed in testis match Drosophila genomic sequence, but not predicted genes. These data strongly support the idea that sequencing additional cDNA libraries from defined tissues, such as testis, will be important tools for refined annotation of the Drosophila genome. Additionally, these data suggest that the number of genes in Drosophila will significantly exceed the conservative estimate of 13,601. PMID- 11116099 TI - The comparison of gene expression from multiple cDNA libraries. AB - We describe a method for comparing the abundance of gene transcripts in cDNA libraries. This method allows for the comparison of gene expression in any number of libraries, in a single statistical analysis, to identify differentially expressed genes. Such genes may be of potential biological or pharmaceutical relevance. The formula that we derive is essentially the entropy of a partitioning of genes among cDNA libraries. This work goes beyond previously published analyses, which can either compare only two libraries, or identify a single outlier in a group of libraries. This work also addresses the problem of false positives associated with repeating the test on many thousands of genes. A randomization procedure is described that provides a quantitative measure of the degree of belief in the results; the results are further verified by considering a theoretically derived large deviations rate for the test statistic. As an example, the analysis is applied to four prostate cancer libraries from the Cancer Genome Anatomy Project. The analysis identifies biologically relevant genes that are differentially expressed in the different tumor cell types. PMID- 11116100 TI - A large database of chicken bursal ESTs as a resource for the analysis of vertebrate gene function. AB - Chicken B cells create their immunoglobulin repertoire within the Bursa of Fabricius by gene conversion. The high homologous recombination activity is shared by the bursal B-cell-derived DT40 cell line, which integrates transfected DNA constructs at high rates into its endogenous loci. Targeted integration in DT40 is used frequently to analyze the function of genes by gene disruption. In this paper, we describe a large database of >7000 expressed sequence tags (ESTs) from bursal lymphocytes that should be a valuable resource for the identification of gene disruption targets in DT40. ESTs of interest can be recognized easily by online or keyword searches. Because the database reflects the gene expression profile of bursal lymphocytes, it provides valuable hints as to which genes might be involved in B-cell-specific processes related to immunoglobulin repertoire formation, signal transduction, transcription, and apoptosis. This large collection of chicken ESTs will also be useful for gene expression studies and comparative gene mapping within the chicken genome project. Details of the bursal EST sequencing project and access to database search forms can be found on the DT40 web site (http://genetics.hpi.uni-hamburg.de/dt40.html). PMID- 11116101 TI - Fluvastatin upregulates inducible nitric oxide synthase expression in cytokine stimulated vascular smooth muscle cells. AB - Nitric oxide (NO) production by inducible NO synthase (iNOS) may play an important role in the pathogenesis of atherosclerosis. Although fluvastatin has been shown to reduce progression of atherosclerosis, it is not known whether it regulates iNOS expression. We investigated the effects of fluvastatin on iNOS expression and subsequent NO synthesis in vascular smooth muscle cells (VSMCs) and the mechanism by which fluvastatin exerts its effects. Fluvastatin significantly increased interleukin-1ss (IL-1ss)-induced nitrite production by VSMCs in a time-dependent (0 to 24 hours) and dose-dependent (10(-)(8) to 10( )(5) mol/L) manner. Increased nitrite production by fluvastatin was accompanied by increased iNOS mRNA and protein accumulation. IL-1ss induced nuclear factor kappaB activation in VSMCs, which was not affected by fluvastatin. Exogenous mevalonate significantly prevented the stimulatory effect of fluvastatin on nitrite production. Cotreatment with geranylgeranyl-pyrophosphate also reversed the effect of fluvastatin. Furthermore, both Rho inhibitor C3 exoenzyme and Rho kinase inhibitor Y-27632 significantly increased IL-1ss-induced nitrite accumulation in VSMCs. These results demonstrated that fluvastatin upregulates iNOS expression and subsequent NO formation in rat VSMCs through inhibition of Rho. PMID- 11116102 TI - Increased arterial intima-media thickness and in vivo LDL oxidation in young men with borderline hypertension. AB - We used borderline hypertension as a model for prehypertension to examine the early influences of elevated blood pressure on subclinical atherosclerosis, lipoprotein oxidation, and cardiac adaptation. Healthy men (age 37+/-4 years) were classified prospectively into 2 groups on the basis of having either borderline hypertension (systolic 130 to 140 mm Hg or diastolic 85 to 89 mm Hg, n=16) or normal (<130/85 mm Hg, n=22) blood pressure values during the previous 2 years. The groups were matched for age, body size, and serum cholesterol levels. High-resolution ultrasound was used to measure intima-media thickness (IMT) of the carotid and brachial arteries, cardiac dimensions, and brachial artery endothelial function. Baseline low-density lipoprotein (LDL)-diene conjugation was measured as an estimate of in vivo LDL oxidation (ox-LDL). Compared with normotensive controls, men with borderline hypertension had higher IMT of the carotid artery (0.58+/-0.06 versus 0.75+/-0.07 mm, P<0.001) and IMT of the brachial artery (0.45+/-0.05 versus 0.57+/-0.07 mm, P<0.001), and increased levels of ox-LDL (29+/-9 versus 47+/-17 mol/L, P<0.001), but similar endothelial function. Left ventricular mass was similar in both groups, but there were significant differences in left ventricular geometry. In multivariate analyses, the predictors of carotid IMT were 24-hour systolic blood pressure (P<0.001) and ox-LDL (P=0.10). The current study demonstrates evidence of increased subclinical atherosclerosis and ox-LDL in borderline hypertension. These results are consistent with the idea that enhanced ox-LDL may be one of the pathophysiological events related to development of atherosclerosis in men with borderline elevated blood pressure. PMID- 11116103 TI - Effect of enalapril on exhaled nitric oxide in normotensive and hypertensive subjects. AB - We investigated whether an angiotensin-converting enzyme (ACE) inhibitor increases the production of nitric oxide (NO) in exhaled air in normotensive and hypertensive subjects. In study 1, 8 normotensive male subjects received a single oral dose of enalapril (5 mg) or nitrendipine (10 mg) in a crossover manner. Exhaled air was collected at baseline, and at 2, 4, and 8 hours after administration of the drug. In study 2, 10 normotensive subjects (6 men and 4 women) and 10 hypertensive subjects (6 men and 4 women) received a single oral dose of enalapril (5 mg). Exhaled air was collected at baseline and at 2 and 4 hours after administration of the drug. In study 1, enalapril significantly increased the NO release rate from the lung in normotensive subjects (36.9+/-5.1 pmol/s at baseline versus 58.3+/-7.3 pmol/s at 4 hours, P<0.01). Nitrendipine did not change the NO release rate. In study 2, enalapril significantly increased the release of NO from the lung in normotensive subjects (40.4+/-6.0 pmol/s at baseline versus 70. 3+/-11.4 pmol/s at 4 hours, P<0.01) but not in hypertensive subjects. ACE inhibition increased NO production from the lung in normotensive subjects but not in hypertensive patients. The reduction of angiotensin II production and/or the accumulation of bradykinin in the pulmonary tissue may be responsible for increased NO production in components of the lung, such as the pulmonary vascular endothelium, bronchial epithelial cells, macrophages, nasopharyngeal cells, and neurons. However, the effects of ACE inhibition on NO production from the lung differ between hypertensive subjects and normotensive subjects. PMID- 11116104 TI - Transient hypertension directly impairs endothelium-dependent vasodilation of the human microvasculature. AB - Hypertension is associated with decreased endothelium-dependent vasodilation. However, whether endothelial dysfunction is a cause or a consequence of elevated blood pressure is unknown. Therefore, to determine whether hypertension can directly induce endothelial dysfunction, we investigated the effect of increases in intra-arterial pressure on endothelium-dependent vasodilation of the human microvasculature. Small arteries (internal diameter 202+/-75 micrometer) were isolated from gluteal fat biopsies in 12 healthy normotensive subjects (8 men and 4 women; age, 46+/-10 years). Arteries were cannulated and perfused in chambers oxygenated at 37 degrees C. Endothelium-dependent and -independent responses to acetylcholine (Ach; 10(-9) to 10(-4) mol/L) and sodium nitroprusside (SNP; 10(-9) to 10(-4) mol/L), respectively, were obtained after incubating the vessel with incremental intravascular pressures of 50, 80, and 120 mm Hg for 60 minutes each. The response to Ach was also obtained in different arteries after 3 consecutive incubation periods at 50 mm Hg. Arterial internal diameter was measured directly from amplified digital images. A significant reduction in the vasodilator response to Ach was observed with increases in intravascular pressure (mean vasodilation, 62%, 49%, and 26% at 50, 80, and 120 mm Hg, respectively; P<0.001). In contrast, the response to SNP showed a nonsignificant trend toward greater vasodilation with increases in pressure (mean vasodilation, 40%, 52%, and 57% at 50, 80, and 120 mm Hg, respectively; P=0.10). There was no difference in the consecutive dose-response curves to Ach obtained at the same intravascular pressure (mean vasodilation: 48%, 46%, and 49%; P=0.61). Transient increases in intravascular pressure significantly depress endothelium-dependent vasodilation in human resistance arteries. These findings suggest that elevated blood pressure per se may cause endothelial dysfunction in humans and have implications for the pathophysiology of endothelial dysfunction in hypertension. PMID- 11116105 TI - Increased vascular adrenergic vasoconstriction and decreased vasodilation in blacks. Additive mechanisms leading to enhanced vascular reactivity. AB - Blood pressure reactivity is enhanced in young black subjects through mechanisms that are poorly understood. We compared alpha-adrenergic-mediated vasoconstrictor and ss-adrenergic vasodilator sensitivity and their relation to sympathetic activity in blacks and whites. Ten healthy black (age, 29.9+/-2.4 years) and 10 white (age, 28.3+/-1.9 years) men were studied. Forearm blood flow was measured with strain-gauge plethysmography after the intrabrachial artery administration of phenylephrine (1.25 to 20 microgram/min) and isoproterenol (60 and 400 ng/min) after application of lower-body negative pressure and after a cold pressor test. Forearm and systemic norepinephrine spillover were measured with a radioisotope dilution technique. alpha-Adrenergic vasoconstriction was markedly increased (ANOVA P=0.008) and ss-adrenergic vasodilation decreased (ANOVA P=0.02) in blacks. Phenylephrine (10 microgram/min) decreased forearm blood flow by 58.0+/ 2.5% in blacks but only by 26.6+/-6.0% in whites (P<0.001). Vasoconstrictor response to endogenous norepinephrine, stimulated by a cold pressor test, resulted in a higher forearm vascular resistance in blacks than in whites (107.3+/-13 versus 64.8+/-13 mm Hg. mL(-)(1). 100 mL(-)(1), P=0.03). There were no significant ethnic differences in basal or stimulated forearm or systemic norepinephrine spillover. Increased vasoconstrictor and decreased vasodilator responses in blacks were not correlated. Increased sympathetically mediated vascular tone caused by enhanced vasoconstriction and attenuated vasodilation, effects that would be additive, and not increased sympathetic activity could enhance vascular reactivity and may play a role in the pathogenesis of hypertension in blacks. PMID- 11116106 TI - Noninvasive assessment of the digital volume pulse. Comparison with the peripheral pressure pulse. AB - The digital volume pulse can be recorded simply and noninvasively by photoplethysmography. The objective of the present study was to determine whether a generalized transfer function can be used to relate the digital volume pulse to the peripheral pressure pulse and, hence, to determine whether both volume and pressure pulse waveforms are influenced by the same mechanism. The digital volume pulse was recorded by photoplethysmography in 60 subjects (10 women, aged 24 to 80 years), including 20 subjects with previously diagnosed hypertension. Simultaneous recordings of the peripheral radial pulse and digital artery pulse were obtained by applanation tonometry and a servocontrolled pressure cuff (Finapres), respectively. In 20 normotensive subjects, measurements were obtained after the administration of nitroglycerin (NTG, 500 microgram sublingually). Transfer functions obtained by Fourier analysis of the waveforms were similar in normotensive and hypertensive subjects. In normotensive subjects, transfer functions were similar before and after NTG. By use of a single generalized transfer function for all subjects, the radial and digital artery pressure waveforms could be predicted from the volume pulse with an average root mean square error of 4.4+/-2.0 and 4.3+/-1.9 mm Hg (mean+/-SD) for radial and digital artery waveforms, respectively, similar to the error between the 2 pressure waveforms (4.4+/-1.4 mm Hg). The peripheral pressure pulse is related to the digital volume pulse by a transfer function, which is not influenced by effects of hypertension or NTG. Effects of NTG on the volume pulse and pressure pulse are likely to be determined by a similar mechanism. PMID- 11116107 TI - Effect of antioxidant therapy on blood pressure and NO synthase expression in hypertensive rats. AB - Earlier studies have demonstrated evidence for increased reactive oxygen species, enhanced NO synthase (NOS) expression, and elevated NO production in spontaneously hypertensive rats (SHR). Given the negative-feedback regulation of NOS by NO, we hypothesized that enhanced NO inactivation by ROS may contribute to compensatory upregulation of NOS in SHR. The present study was designed to test this hypothesis. Eight-week-old male SHR and Wistar-Kyoto rats were treated for 3 weeks with either a placebo or the potent antioxidant, lazaroid (desmethyltirilazad, 10 mg. kg(-1). d(-1), by gastric gavage). Tail arterial blood pressure, urinary excretion of NO metabolites (ie, nitrate and nitrite), and immunodetectable NOS isotype proteins in the vascular, renal, cardiac, and cerebral tissues were measured. The placebo-treated SHR group showed a marked elevation of blood pressure and a significant upregulation of aorta, kidney, and cardiac tissue endothelial and inducible NOS (eNOS and iNOS, respectively) proteins and of brain and renal tissue neuronal NOS. Lazaroid therapy ameliorated hypertension and mitigated the upregulation of eNOS and iNOS in vascular, renal, and cardiac tissues but had limited effect on the expression of renal and brain neuronal NOS. In contrast, lazaroid therapy had no effect on blood pressure, urinary nitrate and nitrite excretion, or tissue NOS isotype expressions in the Wistar-Kyoto group. These findings support the role of oxidative stress in the genesis and/or maintenance of hypertension and compensatory upregulation of the expression of eNOS and iNOS in SHR. PMID- 11116108 TI - Serum levels of vascular endothelial growth factor in preeclamptic and normotensive pregnancy. AB - The purpose of these studies was first to determine if vascular endothelial growth factor (VEGF), a vascular permeability agent, is increased in the serum of women with preclinical and clinical preclampsia (PE), and second to determine how these levels change after delivery. Twenty preeclamptic and 25 normotensive women at term consented to have blood taken pre- and post-delivery. Ten preeclamptic, 10 gestational hypertensive, and 28 normotensive women had blood collected respectively at 12, 20, and 30 weeks gestation and predelivery. Serum was extracted from all samples, and VEGF concentrations were determined by radioimmunoassay. Predelivery, the median serum VEGF concentration in the preeclamptic group was 51.7 ng/mL, and in the control group the concentration was 13.9 ng/mL (P<0.0001). Serum VEGF concentrations fell within 24 hours of delivery in both groups, which resulted in median values of 3.8 ng/mL and 3.2 ng/mL respectively (P<0.3). At 12 and 20 weeks, there was no significant difference between the serum VEGF concentrations in the 3 groups (P<0.3, 0.052 respectively). At 30 weeks, prior to the onset of clinical PE, the serum VEGF levels in the eventual preeclamptic group were elevated significantly compared with the gestational hypertensive and normotensive groups (P<0.001). Predelivery serum VEGF concentrations were significantly elevated in the preeclamptic group and were similar to those in the first study (P<0.0001). These findings suggest that VEGF may be important in the pathophysiology of PE and has the potential to act as a preclinical marker for the condition. PMID- 11116109 TI - Estrogen replacement reduces age-associated remodeling in rat mesenteric arteries. AB - Estrogen replacement therapy significantly decreases the incidence of cardiovascular disease in postmenopausal women. In aging, there is an increase in vascular stiffness along with a decrease in matrix metalloproteinase (MMP) activity. Our hypothesis was that estrogen replacement would increase MMPs and therefore reduce the vascular stiffness that is associated with aging. Female Sprague-Dawley rats were implanted with a placebo or 17ss-estradiol-containing pellet (0.5 mg/pellet, 60-day release) at 10 months of age (n=6, each). Six young rats (3 months old) were also studied. After a 2-month exposure to the pellet, mesenteric arteries were studied on a pressurized arteriograph system. Distensibility and wall thickness were measured in response to stepwise increases in intraluminal pressure in Ca(2+)-free physiological saline solution buffer with papaverine (10(-4) mol/L). In response to increasing pressure, aged placebo rats exhibited a significant decrease in distensibility compared with young rats (P<0.05) that was accompanied by an increase in wall thickness (P<0.05). Conversely, estrogen replacement increased distensibility and decreased wall thickness in aged rats (old estrogen-replaced versus old placebo, P<0.05). Zymography data indicated that MMP-2 activity decreased in aging but was increased by estrogen replacement. In summary, estrogen replacement in aging female rats reduces age-associated vascular remodeling. PMID- 11116110 TI - Target organ damage and the prothrombotic state in hypertension. PMID- 11116111 TI - Relationship of fibrinogen levels and hemostatic abnormalities with organ damage in hypertension. AB - Elevated plasma levels of fibrinogen and activated coagulation pathways are risk factors of cardiovascular disease in the general population. In a cross-sectional study of a case series, we investigated the relationship between fibrinogen and hemostatic markers with target-organ damage (TOD) in patients with arterial hypertension. Prothrombin time, partial thromboplastin time, fibrinogen, fibrin D dimer, prothrombin fragment 1+2 (F1+2), and antithrombin III were measured in 352 untreated patients with mild to moderate essential hypertension and 92 normotensive controls. Staging of TOD was assessed according to W.H.O. guidelines by clinical evaluation and laboratory tests including measurements of creatinine clearance, proteinuria, ophthalmoscopy, electrocardiography, echocardiography, and ultrasound examination of major arteries. F1+2 concentrations were significantly greater in hypertensive patients than normotensive controls and were positively correlated with blood pressure. Age, blood pressure levels, duration of hypertension, smoking, HDL-cholesterol, triglycerides, and plasma fibrinogen, fibrin D-dimer, and F1+2 levels were significantly related to the presence and severity of TOD in univariate analysis. Plasma fibrinogen and D dimer levels were related to organ damage independent of age, blood pressure, duration of hypertension, and smoking status. Separate analysis indicated significant association of fibrinogen and D-dimer levels with cardiac, cerebrovascular, peripheral vascular, and renal damage. In conclusion, elevated plasma levels of fibrinogen and a prothrombotic state are associated with the presence and severity of TOD in patients with essential hypertension and may contribute to the development of atherosclerotic disease in these patients. PMID- 11116112 TI - Interaction of the ACE D allele and the GNB3 825T allele in myocardial infarction. AB - In polygenetic disorders, such as ischemic heart disease, the investigation of gene-gene interactions rather than determination of single gene effects is crucial to better understand the contribution of genetic factors. The 825T allele of the G-protein ss(3)-subunit gene (GNB3) associated with enhanced G-protein signaling is a candidate to interact with the angiotensin-converting enzyme (ACE) deletion/insertion (D/I) polymorphism to increase the risk for myocardial infarction (MI). The ACE D:/I variant affects the renin-angiotensin system hormones that activate G-protein-coupled receptors. Genotyping at the ACE and GNB3 loci was performed on 585 patients with coronary artery disease with (n=270) or without (n=315) previous MI. Logistic regression analysis demonstrated a significant interaction between the ACE D: allele and the GNB3 825T allele (P<0.001). The odds ratio for MI, associated with the 825T allele, was not increased in the presence of the ACE II genotype (OR 0.5; P=0.09) but was significantly higher in 825T allele carriers with the ACE DI genotype (OR 1.9; P=0.01) and further increased in individuals with the ACE DD genotype (OR 2.4; P=0.02). The highest odds ratio was found in homozygous 825T allele carriers with the ACE DD genotype (OR 7.5; P=0.006). Our data suggest a significant interaction of the GNB3 825T allele with the ACE D allele in MI. These hypothesis-generating data may justify larger prospective studies. PMID- 11116113 TI - alpha-adducin and angiotensin I-converting enzyme polymorphisms in essential hypertension. AB - This study focused on two genes that have previously been implicated in hypertension and may influence renal sodium handling, adducin, and angiotensin I converting enzyme (ACE). We compared their polymorphic frequencies and interaction in patients with essential hypertension (n=128) and individually age- and gender-matched normotensive control subjects. The alpha-adducin G460W polymorphism was genotyped by DNA amplification and restriction digestion. The ACE I/D polymorphism was assayed by a triple-primer method, with a "nested" polymerase chain reaction primer situated completely within the insertion sequence of the I: allele. The distributions of genotypes and alleles for the two polymorphisms were not significantly different between the case and control populations, and the cross-classification of cases by alpha-adducin and ACE genotype gave a distribution similar to that of control subjects. We have previously reported that the distributions of genotypes for two linked polymorphisms in the aldosterone synthase gene (one in the steroidogenic factor-1 [SF-1] binding site and the other an intronic conversion [IC]) were significantly different between this cohort of essential hypertensives and matched control subjects. The cross-classification of cases by alpha-adducin and SF-1, alpha adducin and IC, ACE and SF-1, and ACE and IC genotype gave a distribution similar to that of control subjects. Hence, no evidence was found to suggest an association between either the alpha-adducin G460W or the ACE I/D polymorphism and hypertension in a careful case-control study. Furthermore, the alpha-adducin G460W, ACE I/D, and aldosterone synthase SF-1 and IC polymorphisms do not appear to interact in our hypertensive population. PMID- 11116114 TI - Adrenomedullin gene delivery attenuates hypertension, cardiac remodeling, and renal injury in deoxycorticosterone acetate-salt hypertensive rats. AB - Adrenomedullin (AM) is a potent vasodilator and natriuretic peptide that plays an important role in cardiorenal function. In this study, we explored the potential protective role of AM in volume-dependent hypertension by somatic gene delivery. Adenovirus containing the human AM cDNA under the control of the cytomegalovirus promoter/enhancer was administered into deoxycorticosterone acetate (DOCA)-salt hypertensive rats via tail vein injection. A single injection of the human AM gene resulted in a prolonged reduction of blood pressure with a maximal reduction of 41 mm Hg 9 days after gene delivery. Human AM gene delivery enhanced renal function, as indicated by a 3-fold increase in renal blood flow and a 2-fold increase in glomerular filtration rate (n=5, P<0.05). Histological examination of the kidney revealed a significant reduction in glomerular sclerosis, tubular injury, luminol protein cast accumulation, and interstitial fibrosis as well as urinary protein. Human AM gene delivery caused significant decreases in left ventricular weight and cardiomyocyte diameter, which were accompanied by reduced interstitial fibrosis and extracellular matrix formation within the heart. Expression of human AM mRNA was detected in the kidney, adrenal gland, heart, aorta, lung, and liver; immunoreactive human AM levels were measured in urine and plasma. Significant increases in urinary and cardiac cAMP levels were observed in DOCA-salt rats receiving the human AM gene, indicating activation of the AM receptor. These findings showed that AM gene delivery attenuates hypertension, protects against cardiac remodeling and renal damage in volume-overload hypertension, and may have significance in therapeutic applications in cardiovascular and renal diseases. PMID- 11116115 TI - Heterozygous knock-Out of ET(B) receptors induces BQ-123-sensitive hypertension in the mouse. AB - Homozygous knock-out of ET(A) or ET(B) receptor genes results in lethal developmental phenotypes in the mouse. Such deleterious phenotypes do not occur in heterozygous littermates. However, it remains to be determined whether mice partially defective in ET(A) or ET(B) receptors display significant alterations in their responses to exogenous or endogenous endothelin-1 (ET-1). Furthermore, the anesthetized ET(B) (+/-) knock-out mice showed a significantly higher mean arterial blood pressure than the ET(A) (+/-) knock-out or their wild-type littermates. The pressor response to ET-1 but not to a selective ET(B) agonist, IRL-1620, was significantly reduced in the ET(A) (+/-) knock-out mice. In ET(B) (+/-) knock-out mice, the pressor effect of IRL-1620 was more markedly altered than those induced by ET-1. In wild-type mice, both ET(A) and ET(B) receptors were found to be involved in the pressor effect of ET-1, as confirmed by the significant and specific antagonism induced by either BQ-123 (ET(A) antagonist) or BQ-788 (ET(B) antagonist). Also, ET(A)-selective or mixed ET(A)/ET(B)- but not ET(B)-selective antagonists reversed the hypertensive state of the ET(B) (+/-) knock-out mice to the level of wild-type littermates. Finally, radiolabeled ET-1 plasmatic clearance was altered in ET(B) (+/-) but not ET(A) (+/-) knock-out mice when compared with wild-type animals. Thus, heterozygous knock-out of ET(B) receptors results in a hypertensive state, suggesting an important physiological role for that particular receptorial entity in opposing the endogenous ET-1 dependent pressor effects in the mouse. PMID- 11116116 TI - Evidence for genetic factors explaining the birth weight-blood pressure relation. Analysis in twins. AB - Epidemiological studies have consistently shown an inverse association between birth weight and systolic blood pressure in later life after adjustment for current size. To examine whether this association is explained by intrauterine or genetic factors, we investigated birth weight and blood pressure data in 53 dizygotic and 61 monozygotic adolescent twin pairs. Birth weight was obtained from the mothers. Blood pressure measurements were performed 6 times at rest and during mental stress. The dizygotic but not the monozygotic twins with the lowest birth weight from each pair had a systolic blood pressure measured at rest and during the reaction time experiment that was higher compared with their cotwins with the highest birth weight (dizygotic twins: blood pressure at rest, 119. 4+/ 9.7 mm Hg versus 117.3+/-8.5 mm Hg, P=0.07, and during a reaction time task, 126.2+/-10.8 versus 123.6+/-9.5, P=0.09; monozygotic twins: blood pressure at rest, 117.4+/-6.4 versus 118. 4+/-9.0, P=0.4, and during a reaction time task, 122.9+/-8.4 versus 124.2+/-10.8, P=0.2). The differences in blood pressure between the cotwins with the lowest and the cotwins with the highest birth weight were different in dizygotic compared with monozygotic twin pairs (for blood pressure at rest, P=0.05; for blood pressure during reaction time, P=0.03). After adjustment for differences in current weight, intrapair differences in birth weight were negatively and significantly associated with differences in systolic blood pressure at rest and during the reaction time task in dizygotic twins (regression coefficient, -5.7 mm Hg/kg [95% confidence interval, -10.4 to -1.0] and -6.3 [-12.7 to 0], respectively) but not in monozygotic twins (-0.1 [-5.4 to 5.2] and +3.5 [-1.8 to 8.8], respectively). Interaction analysis indicated that the associations were different between dizygotic twins and monozygotic twins (P=0.1 and P<0.05, respectively). These data suggest that genetic factors may play an important role in the association between birth weight and blood pressure. PMID- 11116117 TI - Consistency of hemodynamic responses to cold stress in adolescents. AB - Laboratory research on hypertension often is performed with cold stress to elicit vasoconstriction and increases in blood pressure. Several studies have shown that cardiovascular responses to the cold pressor test predict the development of hypertension. We extended this research by comparing cardiovascular responses to a traditional forehead cold pressor test and a naturalistic whole-body cold exposure. We evaluated blood pressure and impedance cardiographic measures of cardiac output and total peripheral resistance in healthy black (n=69) and white (n=47) adolescents (mean age, 14.7 years) during forehead cold pressor (3 degrees C to 4 degrees C) and passive whole-body exposure to a cold chamber (8 degrees C to 10 degrees C). Both tasks elicited increases in vascular resistance and blood pressure, but forehead cold elicited an increase in cardiac output, whereas whole body cold elicited a decrease in cardiac output (P<0.05). Consistent with previous research, there was a tendency toward greater vasoconstrictive reactivity to cold stress in blacks than in whites, particularly during whole body cold exposure (P<0.05). Cardiovascular reactivity correlated significantly between tasks, but substantial intertask consistency occurred only for cardiac and vascular reactivity in male subjects (r>0.30) but not in female subjects (r<0.15). These gender differences might reflect diminished adrenergic receptor function in female subjects. We conclude that whole-body cold exposure offers a viable, relatively naturalistic alternative to traditional cold pressor tests for the assessment of cardiovascular reactivity. PMID- 11116118 TI - Exercise training increases baroreceptor gain sensitivity in normal and hypertensive rats. AB - Exercise training attenuates arterial hypertension and increases baroreflex sensitivity in spontaneous hypertension. However, no information exists regarding the portion of the baroreflex arch in which this attenuation takes place. We tested the hypothesis that exercise training increases the afferent pathway sensitivity of baroreflex control in both normotensive and spontaneously hypertensive rats (SHR). Arterial pressure and whole-nerve activity of the aortic baroreceptor (multifiber preparation) were evaluated in 30 male rats assigned to 4 groups: sedentary and exercise-trained normotensive rats and sedentary and exercise-trained SHR. Exercise training was performed on a motor treadmill, 5 d/wk for 60 minutes, gradually progressing toward a speed of 26.8 m/min. Exercise training reduced mean arterial pressure in conscious exercise-trained SHR (183+/ 4 versus 165+/-7 mm Hg). The relation between changes in aortic baroreceptor discharge and changes in systolic arterial pressure increased significantly in exercise-trained normotensive rats (2.09+/-0.1 versus 1.44+/-0.1%/mm Hg) and exercise-trained SHR (0.92+/-0.1 versus 0.71+/-0.1%/mm Hg) compared with their respective sedentary rats. Likewise, the average aortic baroreceptor gain sensitivity (calculated by logistic equation) was significantly higher in exercise-trained normotensive rats (2.25+/-0.19 versus 1.77+/-0.03%/mm Hg) and exercise-trained SHR (1.07+/-0.04 versus 0.82+/-0.05%/mm Hg) compared with their respective sedentary control rats. In conclusion, exercise training increases aortic baroreceptor gain sensitivity in normotensive and SHR, thus improving baroreceptor sensitivity, which may result in a more efficient arterial pressure regulation by the baroreflexes. PMID- 11116119 TI - Overexpression of eNOS in NTS causes hypotension and bradycardia in vivo. AB - The role of nitric oxide (NO) in the brain in the control of blood pressure and the sympathetic nervous system is debated. This study examined the effect of overexpression of endothelial NO synthase (eNOS) in the nucleus tractus solitarii (NTS) on blood pressure in conscious rats. Adenovirus vectors encoding either eNOS (AdeNOS) or ss-galactosidase were transfected into the NTS in vivo. In the AdeNOS-treated rats, the local expression of eNOS in the NTS was confirmed by immunohistochemical staining and Western blot analysis for the eNOS protein and by increased production of nitrite/nitrate in the NTS measured by in vivo microdialysis. Blood pressure and heart rate, monitored by the use of a radiotelemetry system in a conscious state, were significantly decreased in the AdeNOS-treated group at day 5 to day 10 after the gene transfer. Urinary norepinephrine excretion also was decreased at day 7 after the gene transfer in the AdeNOS-treated group. Our results indicate that overexpression of eNOS in the NTS decreases blood pressure, heart rate, and sympathetic nerve activity in conscious rats. PMID- 11116120 TI - Effects of spinal section and of positive-feedback excitatory reflex on sympathetic and heart rate variability. AB - The sympathetic outflow appears to be capable of displaying a rhythmicity synchronous with cardiovascular Mayer's waves even after spinal section. To test the hypothesis that spinal sympathetic low frequency (LF) oscillation can be enhanced during sympathetic excitation, we recorded cardiac sympathetic nerve activity (SNA), R-R interval, arterial pressure, and ventilation in 9 unanesthetized decerebrate-vagotomized cats before and after C1 spinal section. LF and high frequency (HF) components were detected in the variability of SNA, R R interval, and systolic arterial pressure both before and after spinal section. In this latter condition, a significant coherence between LF(SNA) and LF(R-R) was present in 5 animals, whereas HF(SNA) and HF(R-R) were correlated in 4 animals. During an excitatory sympathetic spinal reflex elicited by aortic constriction, the efferent sympathetic firing was markedly enhanced (from 7+/-2 to 33+/-7 spikes/s); concomitantly, the powers of both LF(SNA) and HF(SNA) were also increased. Coherence between LF(SNA) and LF(R-R) became significant in all cases, whereas HF(SNA) and HF(R-R) became correlated in 6 animals. In 3 animals, the reflex sympathetic excitation was no longer elicitable after interrupting a vast contingent of sympathetic afferents by means of thoracic dorsal root section. We report for the first time that LF and HF oscillations are detectable in SNA, R-R interval, and systolic arterial pressure variabilities of decerebrate-vagotomized spinal cats and that an excitatory spinal reflex is capable of increasing the power of both SNA spectral components. PMID- 11116121 TI - Decreased cardiopulmonary baroreflex sensitivity in Chagas' heart disease. AB - No study has been performed on reflexes originating from receptors in the heart that might be involved in the pathological lesions of Chagas' heart disease. Our study was undertaken to analyze the role of cardiopulmonary reflex on cardiovascular control in Chagas' disease. We studied 14 patients with Chagas' disease without heart failure and 12 healthy matched volunteers. Central venous pressure, arterial blood pressure, heart rate, forearm blood flow, and forearm vascular resistance were recorded during deactivation of cardiopulmonary receptors. By reducing central venous pressure by applying -10 and -15 mm Hg of negative pressure to the lower body, we observed (a) a similar decrease of central venous pressure in both groups; (b) a marked increase in forearm vascular resistance in the control group but a blunted increase in the Chagas' group; and (c) no significant changes in blood pressure and heart rate. To analyze cardiopulmonary and arterial receptors, we applied -40 mm Hg of lower-body negative pressure. As a consequence, (a) central venous pressure decreased similarly in both groups; (b) blood pressure was maintained in the control group, whereas in patients with Chagas' disease, a decrease in systolic and mean arterial pressure occurred; (c) heart rate increased in both groups; and (d) forearm vascular resistance increased significantly and similarly in both groups. Unloading of receptors with low levels of lower-body negative pressure did not increase forearm vascular resistance in patients with Chagas' disease, which suggests that the reflex mediated by cardiopulmonary receptors is impaired in patients with Chagas' disease without heart failure. Overall control of circulation appears to be compromised because patients did not maintain blood pressure under high levels of lower-body negative pressure. PMID- 11116122 TI - Central cardiovascular action of neuropeptide Y in conscious rabbits. AB - We determined the central interactions of neuropeptide Y and leptin on cardiovascular and sympathetic responses in conscious rabbits. Intracerebroventricular injections of neuropeptide Y (0.1 and 1 nmol/40 microL) elicited dose-related decreases in arterial pressure and renal sympathetic nerve activity without a significant change in heart rate. Peak depressor or sympathoinhibitory responses of mean arterial pressure and renal sympathetic nerve activity (-13.0+/-1.5 mm Hg and -27.6+/-4.9%) were observed at 25 and 20 minutes after intracerebroventricular injection of 1 nmol of neuropeptide Y, respectively. Pretreatment with intracerebroventricular injection of leptin (3 nmol) prevented the depressor and sympathoinhibitory responses elicited by intracerebroventricular neuropeptide Y. Intravenous injection of the same dose of neuropeptide Y (1 nmol) as that used in the intracerebroventricular experiment failed to cause any cardiovascular and renal sympathetic nerve responses. On the other hand, a subdepressor dose of intracerebroventricular infusion of neuropeptide Y (1 nmol/300 microL per hour) significantly attenuated the baroreflex sensitivities assessed by renal sympathetic nerve activity and heart rate compared with vehicle infusion (G(max); -7.4+/-0.7 versus -13.7+/-0.9%/mm Hg, P:<0.01, and -4.0+/-0.3 versus -6.7+/-0.8 bpm/mm Hg, P:<0.05, respectively). These results suggest that central neuropeptide Y participates in the regulations of the sympathetic nerve activity to kidney and the baroreceptor reflex and that the depressor response induced by intracerebroventricular neuropeptide Y is modulated, at least in part, by central leptin in conscious rabbits. PMID- 11116123 TI - Malignant pheochromocytoma. Chromaffin granule transmitters and response to treatment. AB - Chromaffin granule transmitters such as chromogranin A and catecholamines have been used in the diagnosis of pheochromocytoma, but the diagnostic and prognostic value of chromogranin A have not been explored in malignant pheochromocytoma. We evaluated these transmitters in patients with pheochromocytoma (n=27), both benign (n=13) and malignant (n=14). Patients with benign pheochromocytoma were studied before and after surgical excision (n=6), whereas patients with malignant pheochromocytoma were evaluated before and after combination chemotherapy with regular cycles of cyclophosphamide/dacarbazine/vincristine (nonrandomized trial in n=9). During treatment, patient responses to chemotherapy were divided according to anatomic and clinical criteria: responders (n=5) versus nonresponders (n=4). Plasma chromogranin A rose progressively (P<0.0001) from control subjects (48.0+/-3.0 ng/mL) to benign pheochromocytoma (188+/-40.5 ng/mL) to malignant pheochromocytoma (2932+/-960 ng/mL). Parallel changes were seen for plasma norepinephrine (P<0.0001), though plasma epinephrine was actually lower in malignant than benign pheochromocytoma (P=0.0182). In bivariate analyses, chromogranin A, norepinephrine, and epinephrine discriminated between pheochromocytoma and control subjects (all P<0.0001), whereas in a multivariate analyses, norepinephrine was the best discriminator (P:=0.011). Chromogranin A was significantly different in benign versus malignant pheochromocytoma on both bivariate (P=0.0003) and multivariate (P:=0.011) analyses. After excision of benign pheochromocytoma, chromogranin A (P=0.028), norepinephrine (P=0.047), and epinephrine (P=0.037) all fell to values near normal. During chemotherapy of malignant pheochromocytoma (n=9), plasma chromogranin A (P=0.047) and norepinephrine (P=0.02) fell but not epinephrine. In 5 responders to chemotherapy, there were significant declines in chromogranin A (P=0.03) and norepinephrine (P=0.03) but not epinephrine; in 4 nonresponders, none of the transmitters changed. Plasma chromogranin A varied longitudinally with tumor response and relapse. We conclude that plasma chromogranin A is an effective tool in the diagnosis of pheochromocytoma, and markedly elevated chromogranin A may point to malignant pheochromocytoma. During chemotherapy of malignant pheochromocytoma, chromogranin A can be used to gauge tumor response and relapse. PMID- 11116124 TI - Renal protein phosphatase 2A activity and spontaneous hypertension in rats. AB - The impaired renal paracrine function of dopamine in spontaneously hypertensive rats (SHR) is caused by hyperphosphorylation and desensitization of the renal D(1) dopamine receptor. Protein phosphatase 2A (PP(2A)) is critical in the regulation of G-protein-coupled receptor function. To determine whether PP(2A) expression and activity in the kidney are differentially regulated in genetic hypertension, we examined the effects of a D(1)-like agonist, fenoldopam, in renal cortical tubules and immortalized renal proximal tubule cells from normotensive Wistar-Kyoto rats (WKY) and SHR. In cortical tubules and immortalized proximal tubule cells, PP(2A) expression and activities were greater in cytosol than in membrane fractions in both WKY and SHR. Although PP(2A) expressions were similar in WKY and SHR, basal PP(2A) activity was greater in immortalized proximal tubule cells of SHR than WKY. In immortalized proximal tubule cells of WKY, fenoldopam increased membrane PP(2A) activity and expression of the regulatory subunit PP(2A)-B56alpha, effects that were blocked by the D(1) like antagonist SCH23390. Fenoldopam had no effect on cytosolic PP(2A) activity but decreased PP(2A)-B56alpha expression. In contrast, in immortalized proximal tubule cells of SHR, fenoldopam decreased PP(2A) activity in both membranes and cytosol but predominantly in the membrane fraction, without affecting PP(2A) B56alpha expression; this effect was blocked by the D(1)-like antagonist SCH23390. We conclude that renal PP(2A) activity and expression are differentially regulated in WKY and SHR by D(1)-like receptors. A failure of D(1) like agonists to increase PP(2A) activity in proximal tubule membranes may be a cause of the increased phosphorylation of the D(1) receptor in the SHR. PMID- 11116125 TI - Application of supercritical fluid chromatography to characterize a labile digitalis-like factor. AB - A sodium pump inhibitor (digitalis-like factor), isolated from the peritoneal dialysate of volume-expanded, hypertensive patients with kidney failure who were treated with this dialysis modality, was further purified and characterized by means of supercritical fluid chromatography, a separation technique whose application to very-low-concentration biomolecules is new. Previous studies suggested that after high-performance liquid chromatography (HPLC) purification, this inhibitor was the only factor correlated with volume status and blood pressure in these patients. When this same HPLC fraction was furthered purified on 2-dimensional supercritical fluid chromatography, a single peak coeluted with [Na,K]ATPase inhibitory activity. When split specimens were used, there was a strict correlation between the peak area, measured by flame ionization detection, and activity (n=10, R=0.98, P=0.00001). Inhibitory activity after supercritical fluid chromatography was still correlated with the degree of volume expansion of donor patients (P=0.01). After HPLC purification, this volume-sensitive inhibitor was chemically labile. With further purification on supercritical fluid chromatography, the active peak was still labile with comparable half-life. Supercritical fluid chromatography coupled with flame ionization detection provided an estimate of the amount of the inhibitor present. Again using split specimens, we determined that the labile digitalis-like factor was approximately 30-fold more effective than ouabain in inhibiting renal [Na,K]ATPase activity and >/=500 times more effective than ouabain in causing vascular smooth muscle contraction. The data suggest that we have purified to homogeneity a labile digitalis-like factor that is readily distinguished from ouabain or bufalin, based on chromatographic characteristics, chemical lability, and a much lower effective concentration for its biological activity. PMID- 11116126 TI - Mechanisms of increased susceptibility to angiotensin II-induced apoptosis in ventricular cardiomyocytes of spontaneously hypertensive rats. AB - Previous findings have shown that hypotensive doses of losartan prevent the excess of apoptosis present in the hypertrophied left ventricle of adult spontaneously hypertensive rats (SHR). This study was designed to determine whether angiotensin II facilitates apoptosis in cardiomyocytes of adult SHR. Primary cultures of ventricular cardiomyocytes from 30-week-old normotensive Wistar-Kyoto rats (WKY) and SHR with left ventricular hypertrophy were exposed to 10(-)(9) mol/L angiotensin II for 24 hours. Apoptotic cells were assessed by terminal deoxynucleotidyl transferase assay and confirmed by Annexin V detection. The expression of Bax-alpha, Bcl-2, p53, and caspase-3 proteins was assessed by Western blot assays. The expression of BAX gene was assessed by Northern blot. Angiotensin II increased (P<0.01) cardiomyocyte apoptosis, and this effect was higher (P<0.001) in SHR cells than in WKY cells. Whereas losartan (10(-7) mol/L) blocked the apoptotic effect of the octapeptide in cells from the two strains of rats, PD123319 (10(-7) mol/L) inhibited angiotensin II-mediated apoptosis only in SHR cells. Angiotensin II stimulated (P<0.01) Bax-alpha protein, and this effect was higher (P<0.01) in SHR cells than in WKY cells. Angiotensin II did not modify Bcl-2, p53, and BAX mRNA in cells from the two strains of rats. Angiotensin II induced a similar increase (P<0.05) in the ratio caspase-3/procaspase-3 (an index of caspase-3 activation) in cardiomyocytes from the two strains of rats. The present in vitro results indicate that SHR cardiomyocytes exhibit enhanced susceptibility to angiotensin II-induced apoptosis. Ligand binding to angiotensin II type 1 and type 2 receptors leading to changes in posttranscriptional processing of Bax-alpha and accumulation of this proapoptotic protein may be involved in the abnormal response of SHR cardiomyocytes. These data support a role for angiotensin II in apoptosis observed in the left ventricle of these rats. PMID- 11116127 TI - Relation between serum uric acid and risk of cardiovascular disease in essential hypertension. The PIUMA study. AB - The question of serum uric acid as an independent risk factor in subjects with essential hypertension remains controversial. For up to 12 years (mean, 4.0) we followed 1720 subjects with essential hypertension. At entry, all subjects were untreated and all were carefully screened for absence of cardiovascular disease, renal disease, cancer, and other important disease. Outcome measures included total cardiovascular events, fatal cardiovascular events, and all-cause mortality. During 6841 person-years of follow-up there were 184 cardiovascular events (42 fatal) and 80 deaths from all causes. In the 4 quartiles of serum uric acid (division points: 0.268, 0.309, and 0.369 mmol/L [4.5, 5.2, and 6.2 mg/dL] in men; 0.190, 0.232, and 0.274 mmol/L [3.2, 3.9, and 4.6 mg/dL] in women), the rate (per 100 person-years) of cardiovascular events was 2.51, 1.48, 2.66, and 4.27, that of fatal cardiovascular events was 0.41, 0.33, 0.38, and 1.23, and that of all-cause deaths was 1.01, 0.55, 0.93, and 2.01, respectively. The relation between uric acid and event rate was J-shaped in both genders. After adjustment for age, gender, diabetes, total cholesterol/HDL cholesterol ratio, serum creatinine, left ventricular hypertrophy, ambulatory blood pressure, and use of diuretics during follow-up, uric acid levels in the highest quartile were associated with increased risk for cardiovascular events (relative risk, 1.73; 95% CI, 1.01 to 3.00), fatal cardiovascular events (relative risk, 1.96; 95% CI, 1.02 to 3.79), and all-cause mortality (relative risk, 1.63; 95% CI, 1.02 to 2.57) in relation to the second quartile. In untreated subjects with essential hypertension, raised uric acid is a powerful risk marker for subsequent cardiovascular disease and all-cause mortality. PMID- 11116128 TI - Cognitive performance in hypertensive and normotensive older subjects. AB - Longitudinal studies suggest that hypertension in midlife is associated with cognitive impairment in later life. Cross-sectional studies are difficult to interpret because blood pressure can change with onset of dementia and the inclusion of subjects on treatment and with hypertensive end-organ damage can make analysis difficult. We examined cognitive performance in hypertensive and normotensive subjects without dementia or stroke >/=70 years of age. Cognitive performance was determined with the use of a computerized assessment battery in 107 untreated hypertensives (55 women, age 76+/-4 years, blood pressure, 164+/ 9/89+/-7; range, 138 to 179/68 to 99 mm Hg) and 116 normotensives (51 female, age 76+/-4 years, 131+/-10/74+/-7; 108 to 149/60 to 89 mm Hg). Older subjects with hypertension were significantly slower in all tests (reaction time, milliseconds; simple, 346+/-100 versus 318+/-56, P<0.05; memory scanning, 867+/-243 versus 789+/-159, P<0.01; immediate word recognition, 947+/-261 versus 886+/-192, P<0.05; and delayed word recognition, 937+/-230 versus 856+/-184, P<0.05; picture recognition, 952+/-184 versus 894+/-137, P<0.01; spatial memory, 1390+/-439 versus 1258+/-394, P<0.01; excepting choice reaction time, 510+/-75 versus 498+/ 72, P=0.08). Accuracy was also impaired in tests of number vigilance, 99.2+/-2.5% versus 99.9+/-0.9, P<0.01; delayed word recognition, 83.5+/-16 versus 87.9+/-9.8, P<0.01; and spatial memory 64+/-32 versus 79+/-20, P<0.001. Hypertension in older subjects is associated with impaired cognition in a broad range of areas in the absence of clinically evident target organ damage. PMID- 11116129 TI - Nervous kidney. Interaction between renal sympathetic nerves and the renin angiotensin system in the control of renal function. AB - Increases in renal sympathetic nerve activity regulate the functions of the nephron, the vasculature, and the renin-containing juxtaglomerular granular cells. Because increased activity of the renin-angiotensin system can also influence nephron and vascular function, it is important to understand the interactions between the renal sympathetic nerves and the renin-angiotensin system in the control of renal function. These interactions can be intrarenal, for example, the direct (by specific innervation) and indirect (by angiotensin II) contributions of increased renal sympathetic nerve activity to the regulation of renal function. The effects of increased renal sympathetic nerve activity on renal function are attenuated when the activity of the renin-angiotensin system is suppressed or antagonized with ACE inhibitors or angiotensin II-type AT(1) receptor antagonists. The effects of intrarenal administration of angiotensin II are attenuated after renal denervation. These interactions can also be extrarenal, for example, in the central nervous system, wherein renal sympathetic nerve activity and its arterial baroreflex control are modulated by changes in activity of the renin-angiotensin system. In addition to the circumventricular organs, whose permeable blood-brain barrier permits interactions with circulating angiotensin II, there are interactions at sites behind the blood-brain barrier that depend on the influence of local angiotensin II. The responses to central administration of angiotensin II-type AT(1)-receptor antagonists into the ventricular system or microinjected into the rostral ventrolateral medulla are modulated by changes in activity of the renin-angiotensin system produced by physiological changes in dietary sodium intake. Similar modulation is observed in pathophysiological models wherein activity of both the renin-angiotensin and sympathetic nervous systems is increased (eg, congestive heart failure). Thus, both renal and extrarenal sites of interaction between the renin-angiotensin system and renal sympathetic nerve activity are involved in influencing the neural control of renal function. PMID- 11116130 TI - Nitric oxide limits pressor responses to sympathetic activation in rat spinal cord. AB - N-methyl D-aspartate (NMDA) receptor stimulation is known to activate nitric oxide (NO) synthase, an enzyme present in a high proportion of sympathetic preganglionic neurons. In this study, we have examined the possibility that NO modulates the pressor responses elicited by NMDA receptor stimulation in the spinal cord. In experiments on anesthetized rats, we determined whether intrathecal administration of either 3-morpholinylsydnoneimine chloride (SIN-1), an NO donor, or N:(G)-nitro-L-arginine methyl ester (L-NAME), an NO synthase inhibitor, affected the response to stimulation of spinal NMDA receptors by NMDA (1 pmol to 1 micromol in 10-microL intrathecal administration). Intrathecal NMDA resulted in dose-dependent increases in blood pressure. SIN-1 (100 nmol) attenuated the pressor responses to NMDA (F(1,70)=12, P=0.001). Conversely, L NAME (1 nmol to 1 micromol) augmented the pressor response to NMDA in a dose dependent manner (F(3,161)=28.3, P<0.001). The effect of L-NAME to amplify the pressor response to NMDA was reversed by L-arginine but not by D-arginine. These results indicate that endogenous synthesis of NO in the spinal cord limits the pressor response to stimulation of spinal NMDA receptors. PMID- 11116131 TI - Capacity for purinergic control of renin promoter via P2Y(11) receptor and cAMP pathways. AB - Renin secretion can be stimulated by ATP via purinergic P2Y receptors. ATP is a cotransmitter with norepinephrine and is released from the cytosol during cell damage. Such release could account for the de novo renin expression seen in the proximal tubule in renal disease and in myocardial infarct borders. Whereas most P2Y purinoceptor subtypes utilize phosphoinositide signal-transduction pathways, the effector mechanisms of the subtype P2Y(11) also involve increases in cAMP, a well-known renin secretagogue and stimulus to renin production. The present study tested the effect of ATP on human renin gene (REN) promoter activity and the role of P2Y(11). By means of reverse transcriptase-polymerase chain reaction, we found that renin-expressing Calu-6 cells express P2Y(11) mRNA. Expression was also detected in the brain, kidney, testis, muscle, liver, and spleen. We made a novel cell line (Calu-6/P2Y11) in which P2Y(11) cDNA, under the control of a strong promoter, was stably integrated into genomic DNA. These cells produced P2Y(11) mRNA during culture. Treatment of Calu-6/P2Y11 cells with 1 mmol/L ATP caused a 3 fold increase in renin mRNA and protein over 36 hours. Transient transfection of Calu-6/P2Y11 cells with constructs containing 896 bp of human REN 5'-flanking DNA linked to the luciferase reporter gene led to a 5.8+/-0.6-fold increase (mean+/ SEM) in reporter activity in response to ATP (P=0.0015). In contrast, UTP produced only a 1.4+/-0.1-fold increase (P=0.016). For ADP, it was 1.7+/-0.1-fold (P=0.011). The response profile was ATP>ADP>AMP=adenosine=0, consistent with a P2Y(11) effect. Mutation of the cAMP response element (CRE) located at -222 in the REN promoter DNA abolished the effect of ATP. Furthermore, ATP induced a rapid, time-dependent increase in the phosphorylation of CRE binding protein (CREB) and activating transcription factor-1. These data implicate a cAMP pathway in mediation of the P2Y(11) effect. In conclusion, we have made a novel cell line that overexpresses the P2Y(11) purinoceptor. Stimulation of these cells by ATP activates a cAMP signal-transduction pathway that phosphorylates CREB and stimulates renin promoter activity via the CRE at -222. The data raise the possibility of a contribution of ATP/P2Y(11) effects to sympathetic stimulation of renin, as well as to responses in renin seen after tissue damage, such as in kidney disease and myocardial infarction. PMID- 11116132 TI - Retinal neovascularization is prevented by blockade of the renin-angiotensin system. AB - Both angiotensin II and vascular endothelial growth factor are angiogenic agents that have recently been implicated in the pathogenesis of proliferative diabetic retinopathy. In this study, retinal neovascularization was examined in a model of retinopathy of prematurity with the use of neonatal transgenic (mRen-2)27 rats, which overexpress renin in tissues, and Sprague-Dawley rats. Blockers of the renin-angiotensin system were administered during the neovascularization period. The ACE inhibitor lisinopril and the angiotensin type 1 receptor antagonist losartan both increased retinal renin levels and prevented inner retinal blood vessel growth. Quantitative in situ hybridization revealed that the expression of vascular endothelial growth factor and its type 2 receptor in the inner retina and proliferating blood vessels were increased in rats with retinopathy of prematurity. Lisinopril reduced both retinal vascular endothelial growth factor and its type 2 receptor mRNA in retinopathy of prematurity rats, whereas losartan had no effect. It is predicted that agents that interrupt the renin-angiotensin system may play an important role as retinoprotective agents in various forms of proliferative retinopathy. PMID- 11116133 TI - Beneficial renal and cardiac effects of vasopeptidase inhibition with S21402 in heart failure. AB - S21402 is a vasopeptidase inhibitor that simultaneously inhibits neutral endopeptidase (NEP) and angiotensin-converting enzyme (ACE). This study determined whether chronic treatment with S21402 produced different effects on sodium and water excretion, hormonal parameters, and cardiovascular structure compared with selective inhibition of ACE and NEP in a rat model of myocardial infarction-induced congestive heart failure (CHF). CHF rats received the vasopeptidase inhibitor (S21402, 100 mg. kg(-1). d(-1)), an ACE inhibitor (captopril, 50 mg. kg(-1). d(-1)), a NEP inhibitor (SCH42495, 60 mg. kg(-1). d( 1)), or vehicle for 4 weeks. S21402 alone caused a diuresis and natriuresis (P<0.01) in CHF. After 4 weeks, blood pressure was lowered by captopril but not other treatments (P<0.01). Both S21402 and captopril increased plasma renin activity (P<0.01), all treatment lowered plasma aldosterone (P<0.05) and plasma natriuretic peptide levels were unchanged. In the kidney, S21402 inhibited NEP and ACE (P<0.01), SCH42495 inhibited NEP (P<0.01), and captopril inhibited ACE (P<0.01). Heart mass was reduced by all active treatments; captopril reduced left ventricular mass (P<0.01), SCH42495 reduced right ventricular mass (P<0.01), and S21402 decreased left (P<0.05) and right ventricular mass (P<0.01), atrial mass (P<0.05), and lung mass (P<0.01). In CHF, vasopeptidase inhibition with S21402 produces effects that differ from those of selective NEP or ACE inhibition. S21402 improved sodium and water excretion, reduced pulmonary congestion, and attenuated both right and left ventricular remodeling. These effects, which occurred in the absence of any hypotensive action, suggest that S21402 may offer several advantages over ACE inhibition alone in the treatment of heart failure. PMID- 11116134 TI - Vasopressin and blood pressure in humans. PMID- 11116135 TI - gamma -glutamyltransferase and its isoform mediate an endoplasmic reticulum stress response. AB - Although use of multiple alternative first exons generates unique noncoding 5' ends for gamma-glutamyltransferase (GGT) cDNAs in several species, we show here that alternative splicing events also alter coding exons in mouse GGT to produce at least four protein isoforms. GGTDelta1 introduces CAG four bases upstream of the primary ATG codon and encodes an active GGT heterodimeric ectoenzyme identical to constitutive GGT cDNA but translational efficiency is reduced 2 fold. GGTDelta2-5 deletes the last eight nucleotides of exon 2 through most of exon 5 in-frame, selectively eliminating residues 96-231 from the amphipathic N terminal subunit, including four N-glycan consensus sites, while leaving the C terminal hydrophilic subunit intact. GGTDelta7 introduces 22 bases from intron 7 causing a frameshift and a premature stop codon so a truncated polypeptide is encoded terminating with 14 novel residues but retaining the first 339 residues of the native GGT protein. GGTDelta8-9 deletes the terminal four nucleotides of exon 8 plus all of exon 9 and inserts 24 bases from intron 9 in-frame so the C terminal subunit of the encoded polypeptide loses residues 401-444 but gains eight internal hydrophobic residues. In contrast to the product of GGTDelta1, those derived from GGTDelta2-5, Delta7, Delta8-9 all lack transferase activity and persist as single-chain glycoproteins retained largely in the endoplasmic reticulum as determined by immunofluorescence microscopy and constitutive endoglycosidase H sensitivity in metabolically labeled cells. The developmental stage plus tissue-specific regulation of the alternative splicing events at GGTDelta7 and GGTDelta8-9 implies unique roles for these GGT protein isoforms. The ability of the GGTDelta1 and GGTDelta7 to mediate the induction of C/EBP homologous protein-10, CHOP-10, and immunoglobulin heavy chain binding protein, BiP, implicates a specific role for these two GGT protein isoforms in the endoplasmic reticulum stress response. PMID- 11116136 TI - Adp-ribosyl cyclase and cyclic ADP-ribose hydrolase act as a redox sensor. a primary role for cyclic ADP-ribose in hypoxic pulmonary vasoconstriction. AB - Hypoxic pulmonary vasoconstriction is unique to pulmonary arteries and serves to match lung perfusion to ventilation. However, in disease states this process can promote hypoxic pulmonary hypertension. Hypoxic pulmonary vasoconstriction is associated with increased NADH levels in pulmonary artery smooth muscle and with intracellular Ca(2+) release from ryanodine-sensitive stores. Because cyclic ADP ribose (cADPR) regulates ryanodine receptors and is synthesized from beta-NAD(+), we investigated the regulation by beta-NADH of cADPR synthesis and metabolism and the role of cADPR in hypoxic pulmonary vasoconstriction. Significantly higher rates of cADPR synthesis occurred in smooth muscle homogenates of pulmonary arteries, compared with homogenates of systemic arteries. When the beta NAD(+):beta-NADH ratio was reduced, the net amount of cADPR accumulated increased. This was due, at least in part, to the inhibition of cADPR hydrolase by beta-NADH. Furthermore, hypoxia induced a 10-fold increase in cADPR levels in pulmonary artery smooth muscle, and a membrane-permeant cADPR antagonist, 8-bromo cADPR, abolished hypoxic pulmonary vasoconstriction in pulmonary artery rings. We propose that the cellular redox state may be coupled via an increase in beta-NADH levels to enhanced cADPR synthesis, activation of ryanodine receptors, and sarcoplasmic reticulum Ca(2+) release. This redox-sensing pathway may offer new therapeutic targets for hypoxic pulmonary hypertension. PMID- 11116137 TI - Activated cAMP-response element-binding protein regulates neuronal expression of presenilin-1. AB - Upon binding to the cAMP-response element of a gene's promoter, the transcription factor known as cAMP-response element-binding protein (CREB) facilitates transcription of many different neuronal genes including those involved with synaptic function. Based on our previous reports of gene structure (GenBank accession number AF029701 ), we now demonstrate that activated CREB binds to the proximal promoter of the human presenilin-1 (PS-1) gene to activate PS-1 transcription in rat and in human neuronal cells. Specific stimulation of the N methyl-d-aspartate subtype of neuronal glutamate receptors activates CREB and results in increased PS-1 expression. Similarly, treatment with brain-derived neurotrophic factor activates CREB and increases PS-1 expression in a dose dependent fashion. By using adenovirus vectors expressing dominant negative forms of CREB, we were able to show that induction of PS-1 expression requires the activation of CREB. Conversely, constitutive expression of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) results in activation of CREB and increased PS-1 expression that can be blocked by the addition of selective MEK inhibitors. Our findings suggest a hypothesis where stimulation of N-methyl-d-aspartate receptors signals CREB activation to enhance PS-1 gene product expression that contributes to normal neuronal functions. PMID- 11116138 TI - Constitutive signaling by Kaposi's sarcoma-associated herpesvirus G-protein coupled receptor desensitizes calcium mobilization by other receptors. AB - We coexpressed Kaposi's sarcoma-associated herpesvirus G protein-coupled receptors (KSHV-GPCRs) with thyrotropin-releasing hormone (TRH) receptors or m1 muscarinic-cholinergic receptors in Xenopus oocytes and in mammalian cells. In oocytes, KSHV-GPCR expression resulted in pronounced (81%) inhibition (heterologous desensitization) of Ca(2+)-activated chloride current responses to TRH and acetylcholine. Similar inhibitions of cytoplasmic free Ca(2+) responses to TRH were observed in human embryonic kidney HEK 293 EM cells and in mouse pituitary AtT20 cells. Further study of oocytes showed that this inhibition was partially reversed by interferon-gamma-inducible protein 10 (IP-10), an inverse agonist of KSHV-GPCR. The basal rate of (45)Ca(2+) efflux in oocytes expressing KSHV-GPCRs was 4.4 times greater than in control oocytes, and IP-10 rapidly inhibited increased (45)Ca(2+) efflux. In the absence of IP-10, growth-related oncogene alpha caused a further 2-fold increase in (45)Ca(2+) efflux. In KSHV GPCR-expressing oocytes, responses to microinjected inositol 1,4,5-trisphosphate were inhibited by 74%, and this effect was partially reversed by interferon-gamma inducible protein 10. Treatment with thapsigargin suggested that the pool of calcium available for mobilization by TRH was decreased in oocytes coexpressing KSHV-GPCRs. These results suggest that constitutive signaling by KSHV-GPCR causes heterologous desensitization of responses mediated by other receptors, which signal via the phosphoinositide/calcium pathway, which is caused by depletion of intracellular calcium pools. PMID- 11116139 TI - Functional rescue of the nephrogenic diabetes insipidus-causing vasopressin V2 receptor mutants G185C and R202C by a second site suppressor mutation. AB - Mutations in the gene of the G protein-coupled vasopressin V2 receptor (V2 receptor) cause X-linked nephrogenic diabetes insipidus (NDI). Most of the missense mutations on the extracellular face of the receptor introduce additional cysteine residues. Several groups have proposed that these residues might disrupt the conserved disulfide bond of the V2 receptor. To test this hypothesis, we first calculated a structure model of the extracellular receptor domains. The model suggests that the additional cysteine residues may form a second disulfide bond with the free, nonconserved extracellular cysteine residue Cys-195 rather than impairing the conserved bond. To address this question experimentally, we used the NDI-causing mutant receptors G185C and R202C. Their Cys-195 residues were replaced by alanine to eliminate the hypothetical second disulfide bonds. This second site mutation led to functional rescue of both NDI-causing mutant receptors, strongly suggesting that the second disulfide bonds are indeed formed. Furthermore we show that residue Cys-195, which is sensitive to "additional cysteine" mutations, is not conserved among the V2 receptors of other species and that the presence of an uneven number of extracellular cysteine residues, as in the human V2 receptor, is rare among class I G protein-coupled receptors. PMID- 11116140 TI - Further definition of the substance P (SP)/neurokinin-1 receptor complex. MET-174 is the site of photoinsertion p-benzoylphenylalanine4 SP. AB - The covalent attachment site of a substance P (SP) analogue containing the photoreactive amino acid p-benzoyl-l-phenylalanine (Bpa) in position 8 of the C terminal portion of the peptide was identified previously as Met-181 on the neurokinin-1 (NK-1) receptor. In this study, a second photoreactive SP analogue, Bpa(4)-SP, in which the Bpa residue is located in the N-terminal portion of the peptide, was used to define further the peptide-receptor interface. The NK-1 receptor expressed in Chinese hamster ovary cells was specifically and efficiently photolabeled with a radioiodinated derivative of Bpa(4)-SP. Fragmentation analysis of the photolabeled receptor restricted the site of photoincorporation of Bpa(4)-SP to an amino acid within the sequence Thr-173 to Arg-177 located on the N-terminal side of the E2 loop. To identify the specific amino acid in this sequence that serves as the covalent attachment site for Bpa(4)-SP, a small photolabeled receptor fragment was generated by chemical cleavage with cyanogen bromide. Matrix-assisted laser desorption/ionization time of flight mass spectrometric analysis of the purified fragment identified a single protonated molecular ion with a molecular mass of 1801.3 +/- 1.8, indicating that upon irradiation, the bound photoligand covalently attaches to the terminal methyl group of a methionine residue. This result, taken together with the results of the peptide mapping studies, establishes that the site of Bpa(4)-SP covalent attachment to the NK-1 receptor is Met-174. PMID- 11116141 TI - Complement factor H is a serum-binding protein for adrenomedullin, and the resulting complex modulates the bioactivities of both partners. AB - Adrenomedullin (AM) is an important regulatory peptide involved in both physiological and pathological states. We have previously demonstrated the existence of a specific AM-binding protein (AMBP-1) in human plasma. In the present study, we developed a nonradioactive ligand blotting assay, which, together with high pressure liquid chromatography/SDS-polyacrylamide gel electrophoresis purification techniques, allowed us to isolate AMBP-1 to homogeneity. The purified protein was identified as human complement factor H. We show that AM/factor H interaction interferes with the established methodology for quantification of circulating AM. Our data suggest that this routine procedure does not take into account the AM bound to its binding protein. In addition, we show that factor H affects AM in vitro functions. It enhances AM-mediated induction of cAMP in fibroblasts, augments the AM-mediated growth of a cancer cell line, and suppresses the bactericidal capability of AM on Escherichia coli. Reciprocally, AM influences the complement regulatory function of factor H by enhancing the cleavage of C3b via factor I. In summary, we report on a potentially new regulatory mechanism of AM biology, the influence of factor H on radioimmunoassay quantification of AM, and the possible involvement of AM as a regulator of the complement cascade. PMID- 11116142 TI - Roles of Meltrin beta /ADAM19 in the processing of neuregulin. AB - Meltrin beta/ADAM19 is a member of ADAMs (a disintegrin and metalloproteases), which are a family of membrane-anchored glycoproteins that play important roles in fertilization, myoblast fusion, neurogenesis, and proteolytic processing of several membrane-anchored proteins. The expression pattern of meltrin beta during mouse development coincided well with that of neuregulin-1 (NRG), a member of the epidermal growth factor family. Then we examined whether meltrin beta participates in the proteolytic processing of membrane-anchored NRGs. When NRG beta1 was expressed in mouse L929 cells, its extracellular domain was constitutively processed and released into the culture medium. This basal processing activity was remarkably potentiated by overexpression of wild-type meltrin beta, which lead to the significant decrease in the cell surface exposure of extracellular domains of NRG-beta1. Furthermore, expression of protease deficient mutants of meltrin beta exerted dominant negative effects on the basal processing of NRG-beta1. These results indicate that meltrin beta participates in the processing of NRG-beta1. Since meltrin beta affected the processing of NRG beta4 but not that of NRG-alpha2, meltrin beta was considered to have a preference for beta-type NRGs as substrate. Furthermore, the effects of the secretory pathway inhibitors suggested that meltrin beta participates in the intracellular processing of NRGs rather than the cleavage on the cell surface. PMID- 11116143 TI - Insulin and analogue effects on protein degradation in different cell types. Dissociation between binding and activity. AB - In adult animals, the major effect of insulin on protein turnover is inhibition of protein degradation. Cellular protein degradation is under the control of multiple systems, including lysosomes, proteasomes, calpains, and giant protease. Insulin has been shown to alter proteasome activity in vitro and in vivo. We examined the inhibition of protein degradation by insulin and insulin analogues (Lys(B28),Pro(B29)-insulin (LysPro), Asp(B10)-insulin (B10), and Glu(B4),Gln(B16),Phe(B17)-insulin (EQF)) in H4, HepG2, and L6 cells. These effects were compared with receptor binding. Protein degradation was examined by release of trichloroacetic acid-soluble radioactivity from cells previously labeled with [(3)H]leucine. Short- and intermediate-lived proteins were examined. H4 cells bound insulin with an EC(50) of 4.6 x 10(-9) m. LysPro was similar. The affinity of B10 was increased 2-fold; that of EQF decreased 15-fold. Protein degradation inhibition in H4 cells was highly sensitive to insulin (EC(50) = 4.2 x 10(-11) and 1.6 x 10(-10) m, short- and intermediate-lived protein degradation, respectively) and analogues. Despite similar binding, LysPro was 11- to 18-fold more potent than insulin at inhibiting protein degradation. Conversely, although EQF showed lower binding to H4 cells than insulin, its action was similar. The relative binding potencies of analogues in HepG2 cells were similar to those in H4 cells. Examination of protein degradation showed insulin, LysPro, and B10 were equivalent while EQF was less potent. L6 cells showed no difference in the binding of the analogues compared with insulin, but their effect on protein degradation was similar to that seen in HepG2 cells except B10 inhibited intermediate-lived protein degradation better than insulin. These studies illustrate the complexities of cellular protein degradation and the effects of insulin. The effect of insulin and analogues on protein degradation vary significantly in different cell types and with different experimental conditions. The differences seen in the action of the analogues cannot be attributed to binding differences. Post-receptor mechanisms, including intracellular processing and degradation, must be considered. PMID- 11116144 TI - Human glial cell line-derived neurotrophic factor receptor alpha 4 is the receptor for persephin and is predominantly expressed in normal and malignant thyroid medullary cells. AB - Glial cell line-derived neurotrophic factor (GDNF) family ligands signal through receptor complex consisting of a glycosylphosphatidylinositol-linked GDNF family receptor (GFR) alpha subunit and the transmembrane receptor tyrosine kinase RET. The inherited cancer syndrome multiple endocrine neoplasia type 2 (MEN2), associated with different mutations in RET, is characterized by medullary thyroid carcinoma. GDNF signals via GFRalpha1, neurturin via GFRalpha2, artemin via GFRalpha3, whereas the mammalian GFRalpha receptor for persephin (PSPN) is unknown. Here we characterize the human GFRalpha4 as the ligand-binding subunit required together with RET for PSPN signaling. Human and mouse GFRalpha4 lack the first Cys-rich domain characteristic of other GFRalpha receptors. Unlabeled PSPN displaces (125)I-PSPN from GFRA4-transfected cells, which express endogenous Ret. PSPN can be specifically cross-linked to mammalian GFRalpha4 and Ret, and is able to promote autophosphorylation of Ret in GFRA4-transfected cells. PSPN, but not other GDNF family ligands, promotes the survival of cultured sympathetic neurons microinjected with GFRA4. We identified different splice forms of human GFRA4 mRNA encoding for two glycosylphosphatidylinositol-linked and one putative soluble isoform that were predominantly expressed in the thyroid gland. Overlapping expression of RET and GFRA4 but not other GFRA mRNAs in normal and malignant thyroid medullary cells suggests that GFRalpha4 may restrict the MEN2 syndrome to these cells. PMID- 11116145 TI - Transepithelially transported pro-phenoloxidase in the cuticle of the silkworm, Bombyx mori. Identification of its methionyl residues oxidized to methionine sulfoxides. AB - Pro-phenoloxidase (proPO) in insects is activated through the action of a protease cascade triggered by minute amounts of microbial cell wall components. It is an important molecule for the defense against invading microorganisms and for the repair of wounds. In the accompanying paper (Asano, T., and Ashida, M. (2001) J. Biol. Chem. 276, 11100-11112), a proPO isoform, proPO-HS, in the hemolymph of the silkworm, Bombyx mori, is reported to be transported to the cuticle. The transported proPO isoform was recovered from the cuticle and named proPO-CS. The elution profiles of proPO-CS and proPO-HS in reversed-phase high performance liquid chromatography (HPLC) were found to be different, giving a basis to the inference that proPO-CS is a modified form of proPO-HS. In the present study, we investigated the nature of the modifications occurring in proPO CS, in which proteolytically and chemically cleaved fragments originating from the subunits of proPO-CS and proPO-HS were analyzed by reversed-phase HPLC, amino acid sequencing, and mass spectrometry. A subunit of the heterodimeric proPO-CS was found to contain five or six methionine sulfoxides, and another subunit was found to contain one methionine residue oxidized to the sulfoxide. All of the oxidized methionyl residues were identified. Other than oxidation of the methionyl residues, no additional modification of proPO-CS was found. In the model structure of each subunit of proPO-CS constructed by protein modeling with the known structures of the horseshoe crab, Limulus polyphemus, hemocyanin type II subunit as templates, the methionine residues identified as methionine sulfoxide had high degrees of accessibility to the solvent. The implication of the oxidation at the methionine residues is discussed in relation to the mechanism of transepithelial transport of proPO from the hemolymph to the cuticle. PMID- 11116146 TI - The epidermal growth factor receptor engages receptor interacting protein and nuclear factor-kappa B (NF-kappa B)-inducing kinase to activate NF-kappa B. Identification of a novel receptor-tyrosine kinase signalosome. AB - The transcription factor nuclear factor-kappaB (NF-kappaB) is activated by a diverse number of stimuli including tumor necrosis factor-alpha, interleukin-1, UV irradiation, viruses, as well as receptor tyrosine kinases such as the epidermal growth factor receptor (EGFR). NF-kappaB activation by the tumor necrosis factor receptor (TNFR) involves the formation of a multiprotein complex termed a signalosome. Although previous studies have shown that the activated EGFR can induce NF-kappaB, the mechanism of this activation remains unknown. In this study, we identify components of the signalosome formed by the activated EGFR required to activate NF-kappaB and show that, although the activated EGFR uses mechanisms similar to the TNFR, it recruits a distinct signalosome. We show the EGFR forms a complex with a TNFR-interacting protein (RIP), which plays a key role in TNFR-induced NF-kappaB activation, but not with TRADD, an adaptor protein which serves to recruit RIP to the TNFR. Furthermore, we show that the EGFR associates with NF-kappaB-inducing kinase (NIK) and provide evidence suggesting multiprotein complex formation between the EGFR, RIP, and NIK. Using a dominant negative NIK mutant, we show that NIK activation is required for EGFR-mediated NF kappaB induction. We also show that a S32/36 IkappaBalpha mutant blocks EGFR induced NF-kappaB activation. Our studies also suggest that a high level of EGFR expression, a frequent occurrence in human tumors, is optimal for epidermal growth factor-induced NF-kappaB activation. Finally, although protein kinase B/Akt has been implicated in tumor necrosis factor and PDGF-induced NF-kappaB activation, our studies do not support a role for this protein in EGFR-induced NF kappaB activation. PMID- 11116147 TI - A nucleoprotein complex containing CCAAT/enhancer-binding protein beta interacts with an insulin response sequence in the insulin-like growth factor-binding protein-1 gene and contributes to insulin-regulated gene expression. AB - Highly related insulin response sequences (IRSs) mediate effects of insulin on the expression of multiple genes in the liver, including insulin-like growth factor binding protein-1 (IGFBP-1) and phosphoenolpyruvate carboxykinase (PEPCK). Gel shift studies reveal that oligonucleotide probes containing an IRS from the IGFBP-1 or PEPCK gene form a similar complex with hepatic nuclear proteins. Unlabeled competitors containing the IGFBP-1 or PEPCK IRS or a binding site for C/EBP proteins inhibit the formation of this complex. Antibody against C/EBPbeta (but not other C/EBP proteins) supershifts this complex, and Western blotting of affinity purified proteins confirms that C/EBPbeta is present in this complex. Studies with affinity purified and recombinant protein indicate that C/EBPbeta does not interact directly with the IRS, but that other factors are required. Gel shift assays and reporter gene studies with constructs containing point mutations within the IRS reveal that the ability to interact with factors required for the formation of this complex correlates well with the ability of insulin to regulate promoter activity via this IRS (r = 0.849, p < 0.01). Replacing the IRS in reporter gene constructs with a C/EBP-binding site (but not an HNF-3/forkhead site or cAMP response element) maintains the effect of insulin on promoter activity. Together, these findings indicate that a nucleoprotein complex containing C/EBPbeta interacts with IRSs from the IGFBP-1 and PEPCK genes in a sequence-specific fashion and may contribute to the ability of insulin to regulate gene expression. PMID- 11116148 TI - Insulin suppresses transactivation by CAAT/enhancer-binding proteins beta (C/EBPbeta). Signaling to p300/CREB-binding protein by protein kinase B disrupts interaction with the major activation domain of C/EBPbeta. AB - CAAT/enhancer-binding proteins (C/EBPs) play an important role in the regulation of gene expression in insulin-responsive tissues. We have found that a complex containing C/EBPbeta interacts with an insulin response sequence in the insulin like growth factor-binding protein-1 (IGFBP-1) gene and that a C/EBP-binding site can mediate effects of insulin on promoter activity. Here, we examined mechanisms mediating this effect of insulin. The ability of insulin to suppress promoter activity via a C/EBP-binding site is blocked by LY294002, a phosphatidylinositol 3-kinase inhibitor, but not by rapamycin, which blocks activation of p70(S6 kinase). Dominant negative phosphatidylinositol 3-kinase and protein kinase B (PKB) block the effect of insulin, while activated PKB suppresses promoter function via a C/EBP-binding site, mimicking the effect of insulin. Coexpression studies indicate that insulin and PKB suppress transactivation by C/EBPbeta, but not C/EBPalpha, and that N-terminal transactivation domains in C/EBPbeta are required. Studies with Gal4 fusion proteins reveal that insulin and PKB suppress transactivation by the major activation domain in C/EBPbeta (AD II), located between amino acids 31 and 83. Studies with E1A protein indicate that interaction with p300/CBP is required for transactivation by AD II and the effect of insulin and PKB. Based on a consensus sequence, we identified a PKB phosphorylation site (Ser(1834)) within the region of p300/CBP known to bind C/EBPbeta. Mammalian two hybrid studies indicate that insulin and PKB disrupt interactions between this region of p300 and AD II and that Ser(1834) is critical for this effect. Signaling by PKB and phosphorylation of Ser(1834) may play an important role in modulating interactions between p300/CBP and transcription factors and mediate effects of insulin and related growth factors on gene expression. PMID- 11116149 TI - Activation of MAPKs by angiotensin II in vascular smooth muscle cells. Metalloprotease-dependent EGF receptor activation is required for activation of ERK and p38 MAPK but not for JNK. AB - In cultured vascular smooth muscle cells (VSMC), the vasculotrophic factor, angiotensin II (AngII) activates three major MAPKs via the G(q)-coupled AT1 receptor. Extracellular signal-regulated kinase (ERK) activation by AngII requires Ca(2+)-dependent "transactivation" of the EGF receptor that may involve a metalloprotease to stimulate processing of an EGF receptor ligand from its precursor. Whether EGF receptor transactivation also contributes to activation of other members of MAPKs such as p38MAPK and c-Jun N-terminal kinase (JNK) by AngII remains unclear. In the present study, we have examined the effects of a synthetic metalloprotease inhibitor BB2116, and the EGF receptor kinase inhibitor AG1478 on AngII-induced activation of MAPKs in cultured VSMC. BB2116 markedly inhibited ERK activation induced by AngII or the Ca(2+) ionophore without affecting the activation by EGF or PDGF. BB2116 as well as HB-EGF neutralizing antibody inhibited the EGF receptor transactivation by AngII, suggesting a critical role of HB-EGF in the metalloprotease-dependent EGF receptor transactivation. In addition to the ERK activation, activation of p38MAPK and JNK by AngII was inhibited by an AT1 receptor antagonist, RNH6270. and EGF markedly activate p38MAPK, whereas but not EGF markedly activates JNK, indicating the possible contribution of the EGF receptor transactivation to the p38MAPK activation. The findings that both BB2116 and AG1478 specifically inhibited activation of p38MAPK but not JNK by AngII support this hypothesis. From these data, we conclude that ERK and p38MAPK activation by AngII requires the metalloprotease-dependent EGF receptor transactivation, whereas the JNK activation is regulated without involvement of EGF receptor transactivation. PMID- 11116150 TI - GDP dissociation inhibitor domain II required for Rab GTPase recycling. AB - Rab GTPases are localized to distinct subsets of organelles within the cell, where they regulate SNARE-mediated membrane trafficking between organelles. One factor required for Rab localization and function is Rab GDP dissociation inhibitor (GDI), which is proposed to recycle Rab after vesicle fusion by extracting Rab from the membrane and loading Rab onto newly formed transport intermediates. GDI is composed of two domains; Rab binding is mediated by Domain I, and the function of Domain II is not known. In this study, Domain II of yeast GDI, encoded by the essential GDI1/SEC19 gene, was targeted in a genetic screen to obtain mutants that might lend insight into the function of this domain. In one gdi1 mutant, the cytosolic pools of all Rabs tested were depleted, and Rab accumulated on membranes, suggesting that this mutant Gdi1 protein has a general defect in extraction of Rab from membranes. In a second gdi1 mutant, the endosomal/vacuolar Rabs Vps21/Ypt51p and Ypt7p accumulated in the cytosol bound to Gdi1p, but localization of Ypt1p and Sec4p were not significantly affected. Using an in vitro assay which reconstitutes Gdi1p-mediated membrane loading of Rab, this mutant Gdi1p was found to be defective in loading of Vps21p but not Ypt1p. Loading of Vps21p by loading-defective Gdi1p was restored when acceptor membranes prepared from a deletion strain lacking Vps21p were used. These results suggest that membrane-associated Rab may regulate recruitment of GDI-Rab from the cytosol, possibly by regulating a GDI-Rab receptor. We conclude that Domain II of Gdi1p is essential for Rab loading and Rab extraction, and confirm that each of these activities is required for Gdi1p function in vivo. PMID- 11116151 TI - Polypyrimidine track-binding protein binding downstream of caspase-2 alternative exon 9 represses its inclusion. AB - We have been using the caspase-2 pre-mRNA as a model system to study the importance of alternative splicing in the regulation of programmed cell death. Inclusion or skipping of a cassette-type exon in the 3' portion of this pre-mRNA leads to the production of isoforms with antagonistic activity in apoptosis. We previously identified a negative regulatory element (In100) located in the intron downstream of alternative exon 9. The upstream portion of this element harbors a decoy 3' acceptor site that engages in nonproductive commitment complex interactions with the 5' splice site of exon 9. This in turn confers a competitive advantage to the exon-skipping splicing pattern. Further characterization of the In100 element reveals a second, functionally distinct, domain located downstream from the decoy 3' acceptor site. This downstream domain harbors several polypyrimidine track-binding protein (PTB)-binding sites. We show that PTB binding to these sites correlates with the negative effect on exon 9 inclusion. Finally, we show that both domains of the In100 element can function independently to repress exon 9 inclusion, although PTB binding in the vicinity of the decoy 3' splice site can modulate its activity. Our results thus reveal a complex composite element that regulates caspase-2 exon 9 alternative splicing through a novel mechanism. PMID- 11116152 TI - A mouse homologue of the Drosophila tumor suppressor l(2)tid gene defines a novel Ras GTPase-activating protein (RasGAP)-binding protein. AB - p120 GTPase-activating protein (GAP) down-regulates Ras by stimulating GTP hydrolysis of active Ras. In addition to its association with Ras, GAP has been shown to bind to several tyrosine-phosphorylated proteins in cells stimulated by growth factors or expressing transforming tyrosine kinase variants. Here we report the cloning and characterization of a novel GAP-binding protein, mTid-1, a DnaJ chaperone protein that represents the murine homolog of the Drosophila tumor suppressor l(2)tid gene. Three alternatively spliced variants of mTid-1 were isolated, two of which correspond to the recently identified hTid-1(L) and hTid 1(S) forms of the human TID1 gene that exhibit opposing effects on apoptosis. We demonstrate that both cytoplasmic precursor and mitochondrial mature forms of mTid-1 associate with GAP in vivo. Interestingly, although mTid-1 is found tyrosine-phosphorylated in v-src-transformed fibroblast cells, GAP selectively binds to the unphosphorylated form of mTid-1. In immunofluorescence experiments, GAP and Tid-1 were shown to colocalize at perinuclear mitochondrial membranes in response to epidermal growth factor stimulation. These findings raise the possibility that Tid chaperone proteins may play a role in governing the conformation, activity, and/or subcellular distribution of GAP, thereby influencing its biochemical and biological activity within cells. PMID- 11116153 TI - Maximal rate and nucleotide dependence of rhodopsin-catalyzed transducin activation: initial rate analysis based on a double displacement mechanism. AB - Despite the growing structural information on receptors and G proteins, the information on affinities and kinetics of protein-protein and protein-nucleotide interactions is still not complete. In this study on photoactivated rhodopsin (R*) and the rod G protein, G(t), we have used kinetic light scattering, backed by direct biochemical assays, to follow G protein activation. Our protocol includes the following: (i) to measure initial rates on the background of rapid depletion of the G(t)GDP substrate; (ii) to titrate G(t)GDP, GTP, and GDP; and (iii) to apply a double displacement reaction scheme to describe the results. All data are simultaneously fitted by one and the same set of parameters. We obtain values of K(m) = 2200 G(t)/microm(2) for G(t)GDP and K(m) = 230 microm for GTP; dissociation constants are K(d) = 530 G(t)/microm(2) for R*-G(t)GDP dissociation and K(d) = 270 microm for GDP release from R*G(t)GDP, once formed. Maximal catalytic rates per photoexcited rhodopsin are 600 G(t)/s at 22 degrees C and 1300 G(t)/s at 34 degrees C. The analysis provides a tool to allocate and quantify better the effects of chemical or mutational protein modifications to individual steps in signal transduction. PMID- 11116154 TI - Cold shock domain factors activate the granulocyte-macrophage colony-stimulating factor promoter in stimulated Jurkat T cells. AB - Cold shock domain (CSD) family members have been shown to play roles in either transcriptional activation or repression of many genes in various cell types. We have previously shown that CSD proteins dbpAv and dbpB (also known as YB-1) act to repress granulocyte-macrophage colony-stimulating factor transcription in human embryonic lung (HEL) fibroblasts via binding to single-stranded DNA regions across the promoter. Here we show that the same CSD factors are involved in granulocyte-macrophage colony-stimulating factor transcriptional activation in Jurkat T cells. Unlike the mechanisms of CSD repression in HEL fibroblasts, CSD mediated activation in Jurkat T cells is not mediated through DNA binding but presumably through protein-protein interactions via the C terminus of the CSD protein with transcription factors such as RelA/NF-kappaB p65. We demonstrate that Jurkat T cells lack truncated CSD factor subtypes present in HEL fibroblasts, which raises the possibility that the cellular content of CSD proteins may determine their final role as activators or repressors of transcription. PMID- 11116155 TI - The yeast inositol polyphosphate 5-phosphatase Inp54p localizes to the endoplasmic reticulum via a C-terminal hydrophobic anchoring tail: regulation of secretion from the endoplasmic reticulum. AB - The budding yeast Saccharomyces cerevisiae has four inositol polyphosphate 5 phosphatase (5-phosphatase) genes, INP51, INP52, INP53, and INP54, all of which hydrolyze phosphatidylinositol (4,5)-bisphosphate. INP54 encodes a protein of 44 kDa which consists of a 5-phosphatase domain and a C-terminal leucine-rich tail, but lacks the N-terminal SacI domain and proline-rich region found in the other three yeast 5-phosphatases. We report that Inp54p belongs to the family of tail anchored proteins and is localized to the endoplasmic reticulum via a C-terminal hydrophobic tail. The hydrophobic tail comprises the last 13 amino acids of the protein and is sufficient to target green fluorescent protein to the endoplasmic reticulum. Protease protection assays demonstrated that the N terminus of Inp54p is oriented toward the cytoplasm of the cell, with the C terminus of the protein also exposed to the cytosol. Null mutation of INP54 resulted in a 2-fold increase in secretion of a reporter protein, compared with wild-type yeast or cells deleted for any of the SacI domain-containing 5-phosphatases. We propose that Inp54p plays a role in regulating secretion, possibly by modulating the levels of phosphatidylinositol (4,5)-bisphosphate on the cytoplasmic surface of the endoplasmic reticulum membrane. PMID- 11116156 TI - Reduced expression of dentin sialophosphoprotein is associated with dysplastic dentin in mice overexpressing transforming growth factor-beta 1 in teeth. AB - Transforming growth factor (TGF)-beta1 is expressed in developing tooth from the initiation stage through adulthood. Odontoblast-specific expression of TGF-beta1 in the tooth continues throughout life; however, the precise biological functions of this growth factor in the odontoblasts are not clearly understood. Herein, we describe the generation of transgenic mice that overexpress active TGF-beta1 predominantly in the odontoblasts. Teeth of these mice show a significant reduction in the tooth mineralization, defective dentin formation, and a relatively high branching of dentinal tubules. Dentin extracellular matrix components such as type I and III collagens are increased and deposited abnormally in the dental pulp, similar to the hereditary human tooth disorders such as dentin dysplasia and dentinogenesis imperfecta. Calcium, one of the crucial inorganic components of mineralization, is also apparently increased in the transgenic mouse teeth. Most importantly, the expression of dentin sialophosphoprotein (dspp), a candidate gene implicated in dentinogenesis imperfecta II (MIM 125420), is significantly down-regulated in the transgenic teeth. Our results provide in vivo evidence suggesting that TGF-beta1 mediated expression of dspp is crucial for dentin mineralization. These findings also provide for the first time a direct experimental evidence indicating that decreased dspp gene expression along with the other cellular changes in odontoblasts may result in human hereditary dental disorders like dentinogenesis imperfecta II (MIM 125420) and dentin dysplasia (MIM 125400 and 125420). PMID- 11116157 TI - An internalization signal in ClC-5, an endosomal Cl-channel mutated in dent's disease. AB - The ClC-5 chloride channel resides mainly in vesicles of the endocytotic pathway and contributes to their acidification. Its disruption in mice entails a broad defect in renal endocytosis and causes secondary changes in calciotropic hormone levels. Inactivating mutations in Dent's disease lead to proteinuria and kidney stones. Possibly by recycling, a small fraction of ClC-5 also reaches the plasma membrane. Here we identify a carboxyl-terminal internalization motif in ClC-5. It resembles the PY motif, which is crucial for the endocytosis and degradation of epithelial Na(+) channels. Mutating this motif increases surface expression and currents about 2-fold. This is probably because of interactions with WW domains, because dominant negative mutants of the ubiquitin-protein ligase WWP2 increased surface expression and currents of ClC-5 only when its PY motif was intact. Stimulating endocytosis by expressing rab5 or its GTPase-deficient Q79L mutant decreased WT ClC-5 currents but did not affect channels with mutated motifs. Similarly, decreasing endocytosis by expressing the inactive S34N mutant of rab5 increased ClC-5 currents only if its PY-like motif was intact. Thus, the endocytosis of ClC-5, which itself is crucial for the endocytosis of other proteins, depends on the interaction of a carboxyl-terminal internalization signal with ubiquitin-protein ligases containing WW domains. PMID- 11116158 TI - The BBXB motif of RANTES is the principal site for heparin binding and controls receptor selectivity. AB - The chemokine RANTES (regulated on activation normal T cell expressed and secreted; CCL5) binds selectively to glycosaminoglycans (GAGs) such as heparin, chondroitin sulfate, and dermatan sulfate. The primary sequence of RANTES contains two clusters of basic residues, (44)RKNR(47) and (55)KKWVR(59). The first is a BBXB motif common in heparin-binding proteins, and the second is located in the loop directly preceding the C-terminal helix. We have mutated these residues to alanine, both as point mutations as well as triple mutations of the 40s and 50s clusters. Using a binding assay to heparin beads with radiolabeled proteins, the (44)AANA(47) mutant demonstrated an 80% reduction in its capacity to bind heparin, whereas the (55)AAWVA(59) mutant retained full binding capacity. Mutation of the (44)RKNR(47) site reduced the selectivity of RANTES binding to different GAGs. The mutants were tested for their integrity by receptor binding assays on CCR1 and CCR5 as well as their ability to induce chemotaxis in vitro. In all assays the single point mutations and the triple 50s cluster mutation caused no significant difference in activity compared with the wild type sequence. However, the triple 40s mutant showed a 80-fold reduction in affinity for CCR1, despite normal binding to CCR5. It was only able to induce monocyte chemotaxis at micromolar concentrations. The triple 40s mutant was also able to inhibit HIV-1 infectivity, but consistent with its abrogated GAG binding capacity, it no longer induced enhanced infectivity at high concentrations. PMID- 11116159 TI - Developmental compensation of imposed astigmatism is not initiated by astigmatic accommodation in chickens. AB - PURPOSE: It is not clear whether emmetropization is confined to spherical refractive errors, or whether astiqmatic errors are also corrected via visual feedback. Experimental results from the animal model of the chicken are equivocal since compensation of imposed astimatic defocus was found in some but not all studies. Astigmatism could only be compensated by changes in the geometry of the cornea or lens. One has tested whether astigmatic spectacle lenses induce astigmatic accommodation as a possible first step of long-lasting compensation. METHODS: Thirty-five chickens were treated with cylinder lenses (+3/0D or -3/0D) for 5 h. Refractions were determined at 1.38 m distance without cycloplegia in hand-held chicks before attaching the lenses, with the lenses on (0 h), and after 3 and 5 h, and after removal of the lenses. Spheres (S), cylinders (C) and axes (A) were determined using infrared photoretinocopy in three axes (the 'PowerRefractor', equipped with a 135 mm lens). RESULTS: (1) The performance of the 'PowerRefractor' was tested in the chickens with trial lenses and gave correct refractions. (2) Astigmatic trial lenses induced refractive errors as expected from their powers in the case of +3/0D lenses: (S) +3.26 +/- 0.93D, (C) 3.45 +/- 0.87D). In the case of -3/0D lenses, slightly more hyperopic spheres were induced (refractions (S) +4.5 +/- 0.48D) but the cylinders were still as expected (-3.25 +/- 0.49D). The axes of astigmatism were correctly reproduced, since rotating the lenses changed the axes of the induced cylinders as expected. (3) Neither after 3 nor after 5 h of lens wear were there significant changes in the axes or the magnitude of astigmatism. Directly after removal of the lens, the refractions did not differ from their start-up values (with +3/0D lenses: (S) +3.31 +/- 1.05D vs. +3.22 +/- 0.76D, (C) -1.19 +/- 1.77D vs. -0.65 +/- 0.94D, (A) 96 +/- 49 vs. 113 +/- 45 deg; with -3/0D lenses: (S) 2.63 +/- 1.12D vs. 2.97 +/- 0.94D, (C) -1.11 +/- 1.15D vs. -0.53 +/- 0.56D, (A) 78 +/- 24 vs. 131 +/- 35 deg). CONCLUSIONS: The most intuitive mechanism for compensation of astigmatic refractive errors, astigmatic accommodation, could not be demonstrated in chickens. In light of this finding, it seems unlikely that a visually controlled mechanism is operating during development to reduced astigmatism by changing corneal or lenticular growth. PMID- 11116160 TI - Summation of texture segregation across orientation and spatial frequency: electrophysiological and psychophysical findings. AB - Objects are usually segregated from ground by several visual dimensions. We studied texture segregation in checkerboards defined by gradients in spatial frequency, orientation or both frequency and orientation, using Gabor-filtered noise patterns. Saliency was measured electrophysiologically using the visual evoked potential (VEP) associated with texture segregation ('tsVEP') (an associated component in the visual evoked potential), and psychophysically by a 2AFC task. Spatial frequency and orientation stimuli evoked percepts of texture segregation and tsVEPs in all 11 subjects. The tsVEPs to combined stimuli were larger than those to each dimension alone, but smaller (74%) than the algebraic sum of tsVEPs to both individual dimensions. Psychophysical detection rates differed significantly between all conditions (P < 0.001), with highest rates for the combined stimuli. The findings suggest that segregation based on a combination of 'orientation' and 'spatial frequency' is more salient than that based on either of these alone. The significant deviation from full additivity in the tsVEPs suggests that simultaneous contrasts in spatial frequency and orientation have a common processing stage. PMID- 11116161 TI - Alignment of orientation-modulated textures. AB - We conducted a Vernier acuity experiment using orientation-modulated (OM) textures in which the overall shape (skewness) of the modulations was manipulated independently of their orientation content. Misalignments between OMs were consistent with the application of global positional tags, but not on the basis of a single cue (e.g. centroid, peak, or zero-crossing). Instead, modelling of our results in terms of orientation-opponent spatial filters not only led to an excellent fit, but also to estimates of the size and shape of these filters that correspond closely to those made by other researchers using a different task and different stimulus parameters and configurations. PMID- 11116162 TI - The effects of blur and size on monocular and stereoscopic localization. AB - Monocular localization of non-abutting stimuli and stereoscopic localization of the same second-order targets are performed with the same precision (Wilcox, L.M. & Hess, R.F. (1996) Is the site of non-linear filtering in stereopsis before or after binocular combination? Vision Research, 36, 391-399). Further, both tasks show a similar dependence on the scale of the stimulus. Since prior studies used Gaussian-enveloped stimuli, modifications of stimulus scale produced concurrent changes in edge blur. The experiments reported here assess the relative contributions of size and blur to the observed dependence on envelope scale for both monocular localization and stereoacuity. Stereoacuity for first-order targets was found to be an order of magnitude better than stereoacuity for second order targets and monocular acuity for both first- and second-order targets. Further, while first-order stereopsis was found to depend solely on blur, second order stereoacuity and monocular acuity were affected by both size and blur. These results suggest that while stereoacuity for first-order stimuli may be determined by a correlative process limited by early additive noise, stereoacuity for second-order stimuli and monocular acuity for non-abutting targets are more likely limited by stimulus-dependent spatial subsampling. PMID- 11116163 TI - The properties of the motion-detecting mechanisms mediating perceived direction in stochastic displays. AB - Previous studies [e.g. Baker & Hess, 1998. Vision Research, 38, 1211-1222] have shown that perceived direction in displays composed of multiple, limited lifetime, Gabor micropatterns (G) is influenced by movement both at the fine spatial scale of the internal luminance modulation (first-order motion) and the coarse spatial scale of the Gaussian, contrast window (second-order motion). However it is presently indeterminate as to whether this pattern of results is indicative of the processes by which first-order and second-order motion signals interact within the visual system per se or those by which motion information, irrespective of how it is defined, is utilised across different spatial scales. To address this issue, and more generally the properties of the mechanisms that analyse motion in such displays, we employed stochastic motion sequences composed of either G, G added to a static carrier (G + C) or G multiplied with a carrier (G*C). Crucially G*C, unlike both G and G + C, micropatterns contain no net first order motion and second-order motion only at the scale of the internal contrast modulation. For small displacements perceived direction in all cases showed a dependence on the internal sinusoidal spatial structure of the micropatterns and characteristic oscillations were typically observed, consistent with models in which first-order motion and second-order motion are encoded on the basis of similar low-level mechanisms. Importantly for larger displacements, and also when the internal spatial structure was randomised on successive exposures (so that motion at this spatial scale was unreliable), performance tended to be veridical for all types of micropattern, even though under these conditions displacements of the G*C micropatterns should have been invisible to current, low-level, motion detecting schemes. This suggests that both low-level motion sensors and mechanisms utilising a different motion-detecting strategy such as high-level, attentive, feature-tracking may mediate perceptual judgements in stochastic displays. PMID- 11116164 TI - Motion coherence detection as a function of luminance level in human central vision. AB - We studied the changes and invariances of foveal motion detection upon dark adaptation. It is well-documented that dark adaptation affects both spatial and temporal aspects of visual processing. The question we were interested in is how this alters motion coherence detection for moving random texture. To compare motion sensitivity at different adaptation levels, we adjusted the viewing distance for equal detectability of a stationary pattern. At these viewing distances we then measured velocity tuning curves for moving random pixel arrays (RPAs). Mean luminance levels ranged from 50 down to 0.005 cd m-2. Our main conclusion is that foveal velocity tuning is amazingly close to luminance invariant, down to a level of 0.05 cd m-2. Because different viewing distances, and hence, retinal image sizes were used, we performed two control experiments to assess variations of these two parameters separately. We examined the effects of retinal inhomogeneities using discs of different size and annuli filled with RPAs. Our conclusion is that the central visual field, including the near periphery is still rather homogeneous for motion detection at 0.05 cd m-2, but the fovea becomes unresponsive at the lowest luminance level. Variations in viewing distance had marked effects on velocity tuning, both at the light adapted level and the 0.05 cd m-2 level. The size and type of these changes indicated the effectiveness of distance scaling, and show that deviations from perfect invariance of motion coherence detection were not due to inaccurate distance scaling. PMID- 11116165 TI - Motion energy versus position tracking: spatial, temporal, and chromatic parameters. AB - The fundamental question in motion perception is whether motion is an interpretation imposed on an object or feature perceived at separate positions at sequential instants, or whether it is the response of direction-sensitive detectors that can extract the motion-energy in the stimulus, i.e. the orientation of spatio-temporal energy. To answer this question we constructed stimuli whose position changed in one direction while the motion energy contained in the same spatial frequency moved in the same or the opposite direction (by superimposing moving sinusoidal gratings on stationary gratings of the same spatial frequency and orientation). In every case tested (0.25-25 Hz temporal frequency; 0.25-1.0 cyc/deg spatial frequency; achromatic and equiluminant contrast), the perceived direction of motion was in the direction of motion energy, indicating the existence of neurons which compute motion direction without explicitly computing spatial position. The measurements also confirmed that motion-energy computations can be modeled as separable in spatial and temporal frequency. PMID- 11116166 TI - The contributions of slant and tilt to the detection of local surface orientation in structure from motion. AB - The contribution of slant and tilt to the detection of differences in local surface orientation was examined for structure-from-motion (SFM) displays of a complex sinusoidal surface. Observers judged whether an elliptical SFM gauge figure appeared to be lying on the surface or intersecting it. The gauge figure orientation either matched the local surface orientation or differed from it in slant, tilt, or both. Similar sensitivity was found for deviations in slant and tilt, but greater biases and variability were found when the gauge figure deviated from the local surface orientation in slant, depending on the sign of the difference between the gauge figure and local surface orientation and the position of the gauge figure. The results are consistent with Stevens' (Biological Cybernetics, 46 (1983) 183-195) discussion of the computational advantages of slant and tilt contributing independently to the detection of differences in local surface orientation. The effects of changes in perceived surface slant and tilt during rotation and of the misperception of surface depth on the detection of local orientation in dynamic images are discussed. PMID- 11116167 TI - Limits of attentive tracking reveal temporal properties of attention. AB - The maximum speed for attentive tracking of targets was measured in three types of (radial) motion displays: ambiguous motion where only attentive tracking produced an impression of direction, apparent motion, and continuous motion. The upper limit for tracking (about 50 deg s-1) was an order of magnitude lower than the maximum speed at which motion can be perceived for some of these stimuli. In all cases but one, the ultimate limit appeared to be one of temporal frequency, 4 8 Hz, not retinal speed or rotation rate. It was argued that this rate reflects the temporal resolution of attention, the maximum rate at which events can be individuated from those that precede or follow them. In one condition, evidence was also found for a speed limit to attentive tracking, a maximum rate at which attention could follow a path around the display. PMID- 11116168 TI - Does a front-end nonlinearity confound VEP acuity measures in human infants? AB - The visual evoked potential is commonly used to estimate visual acuity in infants. The stimulus used is temporally modulated in order to drive the cortical response. Here it is proposed that distortion products generated by a front-end nonlinearity may contaminate the acuity estimate. Specifically, the nonlinearity might convert temporal modulation of a high spatial frequency grating into apparent whole-field flicker. Thus, the VEP may reflect an artifactual response to a high spatial frequency that is not resolved at the cortical level. If this were the case, one could null or attenuate the flicker response by adding whole field flicker to the grating stimulus. We looked for such nulling effects in 18 infants aged 6-17 weeks. No consistent evidence was found for the nulling effect, so it was concluded that infant VEP acuity estimates are not significantly contaminated by the hypothesized distortion product. PMID- 11116169 TI - Technical advances in multi-slice spiral CT. AB - X-ray computerised tomography (CT) scanning with continuous patient transport has been established under the name Spiral CT since several years as the standard clinical examination procedure. This technique has been improved continuously with respect to scan speed, temporal response and z-axis resolution by the use of latest technical developments: Rotation times up to 0.5 s and multi-row detector array systems. Today detector systems with M + 4 simultaneously measured slices are available. We report about recent progress of spiral CT reconstruction algorithms that are based on multi-slice data. It is demonstrated that the new technology not only provides significant reduction in overall scan times and thereby of the CT scanner?s X-ray tube load; beyond that, the new technology allows CT imaging of the beating heart with high level image quality in standard clinical routine. PMID- 11116170 TI - Data explosion: the challenge of multidetector-row CT. AB - The development of multi detector-row CT has brought many exciting advancements to clinical CT scanning. While multi detector-row CT offers unparalleled speed of acquisition, spatial resolution, and anatomic coverage, a challenge presented by these advantages is the substantial increase on the number of reconstructed cross sections that are rapidly created and in need of analysis. This manuscript discusses currently available alternative visualization tecvhniques for the assessment of volumetric data acquired with multi detector-row CT. Although the current capabilities of 3-D workstations offer many possibilities for alternative analysis of MCDT data, substantial improvements both in automated processing, processing speed and user interface will be necessary to realize the vision of replacing the primary analysis of transverse reconstruction's with alternative analyses. The direction that some of these future developments might take are discussed. PMID- 11116171 TI - Colonography using multislice CT. AB - Computed tomography (CT) represents the preferred imaging modality for imaging the large bowel when virtual endoscopic reconstructions are desired. Using the spiral acquisition technique, it has become possible to scan the entire abdomen within a single breathhold, however, slice thicknesses of 5 mm or more are necessary should the breathhold not last longer than 30-40 s. With the advent of multislice CT, contiguous 1-mm slices can be obtained through the entire abdomen while even shortening the breathhold to 25-30 s. The improved speed and spatial resolution of multislice CT results in remarkably sharp virtual reconstructions allowing detection of polyps with sizes less than 3 mm. The disadvantages must still be considered including a dataset consisting of up to 800 images representing a new challenge for postprocessing hard- and software. PMID- 11116172 TI - Multislice CT angiography. AB - Multislice CT has overcome past limitations of CT angiography (CTA): Scan length and spatial resolution can be simultaneously optimized with multislice CTA, contrast medium can be saved, and the evaluation of large anatomic areas and vessels smaller than 1 mm become possible. This article describes how to optimize scanning protocols and contrast injection, and discusses the main clinical applications of this new technique. Only three main scanning protocolssuffice for all indications. A high speed / high-volume protocol (using 4*2mm or 4*2.5mm collimation) can be employed to scan the chest or abdomen in 8-10s, or to cover the whole abdominal aorta and the peripheral runoff including the feet within 40 65s. A high resolution protocol (using 4*1mm or 4*1.25mm) can be employed for the aorta and most regional vascular beds. It allows for near isotrophic imaging and depicts fine vascular structures with excellent detail. Ultra-high resolution protocols (using 2*0.5mm or 4*0.5mm collimation) yield totally isotropic data sets, and are mainly reserved for cerebrovascular imaging. Image processing techniques, and, in particular, volume rendering have made image presentation faster and easier. Multislice CTA exceeds MRA in spatial resolution and is now able to display even small vascular side branches. Its main indications will be aortic diseases, suspected pulmonary embolism but also renal artery stenoses, preoperative workup of abdominal or cerebral vessels, and acute vascular diseases. Multisplice CTA will become a strong competitor of other minimally invasive vascular imaging techniques. PMID- 11116173 TI - Current development of cardiac imaging with multidetector-row CT. AB - Multidector-row CT (MDCT) with retrospective ECG gating allows scanning the entire heart with 1.25 mm slice thickness and 250 ms effective exposure time within 35 s investigation time. The resulting images allow for an accurate high resolution assessment of morphological detail of both the coronary arteries and the cardiac chambers. Performing a contrast-enhanced MDCT angiography (MD-CTA) in addition to a non-enhanced scan for the detection and quantification of coronary calcifications may be indicated in patients with atypical chest pain and in young patients with high cardiovascular risk. This group of patients may show non calcified plaques as the first sign of their coronary artery disease. As the proximal part of the coronary arteries is well displayed by MD-CTA it also helps to delineate the course in anomalous coronary vessels. Additional information is drawn from the preoperative use of MD-CTA do determine the distance of the left internal mammarian artery to the left anterior descending coronary artery prior to minimal invasive bypass grafting. Additional indications for MD-CTA are the non-invasive follow up after venous bypass grafting, PTCA, and coronary stent interventions. MD-CTA allows following the course of the coronary vessels to the level of third generation coronary segmental arteries. A definite diagonis to rule out coronary artery disease can be reliably made in vessels with a diameter of 1.5 mm or greater. With MDCT a number of different atherosclerotic changes can be observed in diseased coronary arteries. Non-stenotic lesions may show tiny calcifications surrounded by large areas of irregularly distributed soft tissue. Calcifications in this type of atherosclerotic coronary artery wall changes appear as 'the tip of iceberg'. Heavy calcifications usually tend to be non stenotic because of vessel remodelling resulting in a widening of the coronary vessel lumen. Therefore, heavy calcifications appear to ack like an 'internal stent' for a coronary vessel segment. Humps of soft tissue either with or without calcifications are more likely to cause significant coronary artery disease and correlate with stenoses of >50% on selective coronary catheter. These humps consist of well-defined soft tissue in the coronary artery wall. The density of this soft tissue may vary between 30-70 HU. In cases where a coronary vessel is occluded by thrombotic material, a typical sign is found with enlargement of the coronary vessel, a hypodense center and a hyperdense rim. Vessel occlusion without thrombus may also appear within a collapsed and dense lumen. In addition to the investigation of the coronary arteries, CTA with MDCT is well suited to assess the presence and morphology of myocardial scars and aneurysms, intracardial tumors and thrombi. PMID- 11116174 TI - Breast varices: imaging findings of an unusual presentation of collateral pathways in superior vena caval syndrome. AB - Imaging findings are presented of an unusual pathway of collateral circulation consisting of bilateral and diffuse dilated breast veins from a patient with long standing superior vena caval syndrome. The main importance of this case is the extent of the collateral development through the breast veins, serving as the major pathway of collateral circulation. Identification of this unusual collateral development, which resembles breast varices, was performed with contrast-enhanced chest CT scans, digital subtraction venography, color Doppler ultrasonography, and mammographic studies. Collateral development was secondary to a long segment idiopathic venous occlusion involving bilateral subclavian and brachiocephalic veins as well as vena cava superior. We conclude that dilated breast veins when detected on any imaging modality should raise the suspicion of central venous obstruction. PMID- 11116175 TI - Digital selenium radiography: detection of subtle pulmonary lesions on images acquired with and without an additional antiscatter grid. AB - OBJECTIVE: the objective of this ROC-study was to evaluate the diagnostic efficacy of images acquired with a grid in digital selenium radiography compared to that on images obtained with the integrated air gap only. MATERIALS AND METHODS: seven types of simulated lesions were superimposed onto an anthropomorphic chest phantom. Selenium radiography images were obtained either with or without an additional antiscatter grid. For images acquired with a grid either a similar or increased exposure level was used. Both normal and obese patients were simulated. RESULTS: When a grid was used with an equivalent detector dose and a higher exposure, diagnostic performance was significantly improved as compared to images obtained with only the air gap. ROC curve areas for mediastinal nodules and catheters were substantially higher for images acquired with a grid and the same exposure level compared to images obtained without a grid. However, detection of linear, net-shaped and reticulonodular structures in peripheral lung regions was significantly worse when a grid was used with an equivalent exposure level. Concerning the interpretation of images obtained from the normal and obese phantom models, no substantial differences were observed. CONCLUSION: a marked improvement in diagnostic performance could be achieved by means of the use of an additional antiscatter grid and an equivalent detector dose. However, when the same exposure was used, images acquired with the grid allowed a better detection of mediastinal structures although a worse performance was evident in radiolucent lung regions. Therefore, the routine use of a grid without increased exposure is not recommended. PMID- 11116177 TI - This round's on TiBS. PMID- 11116176 TI - Coming of age with TiBS. PMID- 11116178 TI - Reflections on the flood of literature. PMID- 11116179 TI - An Editor's view. PMID- 11116180 TI - Biochemistry strikes back. PMID- 11116181 TI - Blotting at 25. PMID- 11116182 TI - Slab-gel electrophoresis. PMID- 11116183 TI - SDS polyacrylamide gel electrophoresis. PMID- 11116184 TI - Antibodies: indispensable tools for biomedical research. PMID- 11116185 TI - The regulation of protein function by multisite phosphorylation--a 25 year update. AB - The phosphorylation of a protein can alter its behaviour in almost every conceivable way. These include modulation of its intrinsic biological activity, subcellular location, half-life and docking with other proteins. 'Multisite phosphorylation can enable several such effects to operate in the same protein. It can also determine the extent and duration of a response and is the key to signal integration. PMID- 11116186 TI - Life's 24-hour clock: molecular control of circadian rhythms in animal cells. AB - Our sleep-wake cycles and many other approximately 24-hour rhythms of behavior and physiology persist in the absence of environmental cues. Genetic and biochemical studies have shown that such rhythms are controlled by internal molecular clocks. These are assembled from the cycling RNA and protein products of a small group of genes that are conserved throughout the animal kingdom. PMID- 11116187 TI - Reflections on a quarter century of research on contractile systems. AB - By the early 1970s studies of muscle contraction reached a high level and the field gave birth to a new line of investigation into the molecular basis of cellular movements. The molecular diversity in these motile systems has proven to be greater than anticipated. Actin filament assembly without direct participation of myosin is used more widely for motility than expected. Atomic structures of key proteins and important technical advances, including single-molecule methods, have enabled detailed investigation of the mechanisms of muscle contraction and cellular motility. However, much work lies ahead to understand the mechanism of force production and to elucidate the signaling pathways that control cellular motility. PMID- 11116188 TI - Protein folding: progress made and promises ahead. AB - Over the past 25 years, enormous breakthroughs have been made in understanding protein folding mechanisms. We have now reached an exciting stage, with consensuses beginning to emerge that combine both theoretical and experimental approaches. In addition, new fields have emerged and burgeoned, including in vivo folding and the study of protein misfolding diseases. In today's post-genomic world, understanding protein folding has never been more important and the topic has wide-ranging impact in fields from structural biology to materials science. PMID- 11116189 TI - 25 years after the nucleosome model: chromatin modifications. AB - Nucleosomes play a dynamic role in transcription. The key to this role is the structure of the flexible and charged histone tails that extend from the hydrophobic nucleosome core. These tails are modified by acetylation and deacetylation, phosphorylation, methylation, ubiquitination and ATP-dependent chromatin remodeling, and they regulate functions as diverse as transcription, DNA-dependent chromatin assembly, DNA repair, mitosis and silencing by heterochromatin. PMID- 11116190 TI - Macromolecular electron microscopy in the era of structural genomics. AB - Macromolecular machines carry out many cellular functions. Cryo-electron microscopy (cryo-EM) is emerging as a powerful method for studying the structure, assembly and dynamics of such macromolecules, and their interactions with substrates. With resolutions still improving, 'single-particle' analyses are already depicting secondary structure. Moreover, cryo-EM can be combined in several ways with X-ray diffraction to enhance the resolution of cryo-EM and the applicability of crystallography. Electron tomography holds promise for visualizing machines at work inside cells. PMID- 11116191 TI - Observing proteins in their natural habitat: the living cell. AB - Fluorescence microscopy has played a tremendous role in uncovering the morphological features of cells and the expression pattern of proteins by immunofluorescence. Since the discovery of green-fluorescent proteins (GFPs), this technique has undergone a revival in the life sciences as the spatial distribution of ectopically expressed fusion proteins inside living cells can now be followed more easily. By further exploiting the photophysical properties of the emitted fluorescence with microspectroscopic methods, spatial information on the biochemical parameters of intracellular processes and reactions can be obtained. This possibility will not only play an important role in the understanding of biochemical reactions in signal processing and fidelity but also help to uncover the molecular mechanisms of organelle and cell morphogenesis. PMID- 11116192 TI - Synchrotron crystallography. AB - The past decade has seen an explosive growth in atomic-level structures determined by X-ray crystallography. Synchrotron radiation and a number of technical advances related quite directly to its development have fueled this growth. With the most recent advances coming to be used collectively and new resources being built, the foundation is laid for a dramatic further expansion of synchrotron crystallography in the next decade. Both the high-throughput applications of structural genomics and also the challenging studies of macromolecular machinery are expected to flourish. PMID- 11116193 TI - Extracellular iron: a signal that can be sensed and transduced in bacteria. PMID- 11116194 TI - Reducing (oxidative) stress: structure and mechanism. PMID- 11116196 TI - Pharmaceutical science and technology today: evolving to reflect the modern industrial life-science environment. AB - "The barriers between 'silos' in pharmaceutical R&D workflows are slowly eroding and this is driving a wider exchange of information across the various R&D disciplines." PMID- 11116195 TI - Extending life--a lifestyle choice. PMID- 11116197 TI - Therapeutic needs revive arsenic compound. AB - An arsenic compound, previously used as an insecticide, is set to become the latest addition to the armoury for treating a rare form of leukaemia. On 26 September 2000, the FDA announced the approval of Trisenox (arsenic trioxide) for the treatment of acute promyelocytic leukaemia (APL), which affects approximately 2000 people each year in the USA. PMID- 11116199 TI - Angiogenesis inhibitor enters clinical trials. PMID- 11116198 TI - Polymer gel passes safety tests, and is set to deliver drugs. PMID- 11116200 TI - Applications of immobilized stationary-phase liquid chromatography: a potential in vitro technique. AB - Immobilized artificial-membrane chromatography is a potential in vitro technique for determining lipophilicity and studying drug transport and membrane interactions. It is reproducible, efficient and simple. Several other and newer applications of immobilized stationary-phase liquid chromatography have been reported, including the purification of membrane proteins, the synthesis of biomolecules and the simultaneous determination of enzyme activity and enantioselectivity. This article describes the immobilized artificial-membrane concept and provides an overview of the applications, advantages and limitations, in general, of immobilized stationary-phase chromatography. PMID- 11116201 TI - Non-invasive administration of drugs through the skin: challenges in delivery system design. AB - Vehicles designed to enhance drug delivery through the skin must incorporate specific elements that improve the ability of the delivery system to overcome the barrier posed by the stratum corneum. This review discusses several chemical penetration enhancers that have been investigated as potential tools to increase drug flux. In addition, lipid-based delivery systems offer an attractive alternative to traditional drug vehicles. The relationship between liposome composition and drug permeation is discussed, in addition to the possible mechanism of action of lipid vesicle-mediated drug delivery. PMID- 11116202 TI - Transdermal testing: practical aspects and methods. AB - Interest in transdermal drug delivery has increased in recent years owing to its many advantages over other routes of administration. In order to evaluate a transdermal product effectively, three main issues need to be addressed: (1) the kind of skin model that will be used to evaluate the drug permeation; (2) the mathematical model that will be used to characterize the permeation of the drug across the skin; and (3) the diffusion apparatus that will be used to conduct the permeation study. PMID- 11116203 TI - Monitor editorship. PMID- 11116205 TI - Progress. Process analysis. PMID- 11116204 TI - Goodbye. PMID- 11116206 TI - Cis-acting elements regulating the placenta-specific promoter of the bovine Cyp19 gene. AB - Cyp19 encodes aromatase cytochrome P450, the key enzyme of oestrogen biosynthesis. In the bovine placenta, the majority of Cyp19 transcripts include a 5' untranslated region which is encoded by exon 1.1; this suggests that its 5' flanking region is the predominant placental promoter. The aim of the present investigation was to examine the promoter activity of this region and to map cis acting regulatory elements in order to improve our understanding of the complex regulation of this gene within the placenta. As an initial approach, human JEG-3 choriocarcinoma cells were transiently transfected with reporter-gene constructs consisting of different 5'-flanking sequences of exon 1.1 fused to the luciferase gene as a reporter. To localise and further characterise functional cis-acting elements, targeted point mutations and electrophoretic mobility-shift experiments were used. The data demonstrate, for the first time, (1) that the bovine exon 1.1 5'-flanking sequence is an active promoter, (2) that 404 bp of this region are sufficient for constitutive reporter-gene expression in JEG-3 cells and (3) that the region includes at least two different enhancer elements; the data also suggest (4) that one of these elements consists of the E-box motif CATGTG and that the second enhancer element includes the half-site hexameric sequence AGGTCA and additional nucleotides flanking this element upstream. PMID- 11116207 TI - p107 is active in the nucleolus in non-dividing human granulosa lutein cells. AB - Cells are maintained in a quiescent state by members of the retinoblastoma protein family, pRb and p130. Both are phosphoproteins and hypophosphorylated forms of pRb and p130 bind and repress the activity of E2F transcription factors, thereby preventing entry into the cell cycle. Mitogenic stimulation causes activation of cyclin dependent kinases (cdk) that phosphorylate both pRb and p130, thereby releasing E2F factors which stimulate the transcription of a number of genes that are required for DNA synthesis and for regulating the cell cycle. In non-dividing cells, cdks are maintained in an inactive state by cdk inhibitor proteins such as p27(Kip1). The aim of our study was to determine how E2F complexes are regulated during the differentiation of human primary granulosa lutein cells (GLC) of the corpus luteum (CL). The CL is formed in the ovary after ovulation at the terminal stage of folliculogenesis after completion of maturation and differentiation of Graafian follicles. As shown by flow cytometry GLC are not dividing, being predominantly in the G(0)/G(1) phase of the cell cycle and, consistent with this, they contain the cdk inhibitor protein, p27(Kip1), but not E2F-1 which is normally expressed only in proliferating cells. The GLC do express E2F-4, hypophosphorylated pRb, p130 forms 1 and 2 and, surprisingly, hypophosphorylated p107. p107 is normally present only in dividing cells where it regulates E2F activity during the cell cycle. These forms of pRb, p130 as well as p107, together with E2F-4 are all active in that they can bind an E2F DNA-binding site in a pull-down assay. Immunocytochemistry shows that these proteins are expressed in almost all GLC but have different sub-cellular distribution: p107 is concentrated in nucleoli, while p130 and E2F-4 show relatively even nuclear and cytoplasmic distributions. Both pRb and p130 have been implicated previously in repressing E2F activity in many different cell types during cell cycle arrest in G(0)/G(1). We conclude that p107 is active in human primary GLC but its nucleolar localisation would suggest that it represses ribosomal RNA synthesis rather than E2F activity. PMID- 11116208 TI - Molecular and cellular actions of a structural domain of human growth hormone (AOD9401) on lipid metabolism in Zucker fatty rats. AB - A lipolytic domain (AOD9401) of human growth hormone (hGH) which resides in the carboxyl terminus of the molecule and contains the amino acid residues 177-191, has been synthesized using solid-phase peptide synthesis techniques. AOD9401 stimulated hormone-sensitive lipase and inhibited acetyl coenzyme A carboxylase (acetyl CoA carboxylase) in isolated rat adipose tissues, in a similar manner to the actions of the intact hGH molecule. The synthetic lipolytic domain mimicked the effect of the intact growth hormone on diacylglycerol release in adipocytes. Chronic treatment of obese Zucker rats with AOD9401 for 20 days reduced the body weight gain of the animals, and the average cell size of the adipocytes of the treated animals decreased from 110 to 80 microm in diameter. Unlike hGH, synthetic AOD9401 did not induce insulin resistance or glucose intolerance in the laboratory animals after chronic treatment. The results suggest that AOD9401 has the potential to be developed into a therapeutic agent for the control of obesity. PMID- 11116209 TI - Effects of a thyromimetic on apolipoprotein B-100 in rats. AB - We have studied the effects of a cardiac sparing thyromimetic, CGS 23425, on postprandial levels of triglycerides, abundance of apolipoprotein B (apo B) protein and hepatic apo B mRNA expression in rats. When compared with control rats, triglyceride clearance was significantly accelerated by treatment with CGS 23425. A full return to baseline values was achieved within 8 h after ingesting a large quantity of fat, as compared to >24 h in control animals. The abundance of apo B-100 protein in CGS 23425-treated hyperlipidemic rats decreased in a dose dependent manner, but levels of apo B-48 were not significantly affected. Like L tri-iodothyronine (L-T(3)), treatment with 30 microg/kg CGS 23425 for 6 or 9 days decreased the levels of apo B-100 protein by 80% and 40% respectively. This change was paralleled by a 27% reduction in hepatic apo B-100 mRNA. To investigate a potential mechanism of CGS 23425 action, we measured in vitro apo B mRNA editing activity in hepatocellular extract from control or CGS 23425-treated rats. Treatment with CGS 23425 increased activity of the hepatic apo B-100 editosome, apobec-1. In human hepatoma cells which lack apobec-1 activity, apo B 100 mRNA levels remained the same in cells treated with or without the agent. In summary, these observations show that CGS 23425 decreases the levels of apo B-100 in rats. This action of CGS 23425 involves apo B-100 mRNA editing activity. PMID- 11116210 TI - The transcription factors SP1 and MAZ regulate expression of the parathyroid hormone/parathyroid hormone-related peptide receptor gene. AB - The parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptor regulates extracellular calcium concentrations and is therefore important for mineral homeostasis. ROS 17/2.8 cells, a rat osteoblast-like osteosarcoma cell line, express the PTH/PTHrP receptor and provide a good model for examining the transcriptional regulation of its gene. The rat PTH/PTHrP receptor gene has two promoters, U1 and U3, which were shown to be important for its expression. Using extracts from ROS 17/2.8 cells, we have demonstrated two regions (termed FP1 and FP2) of nuclear protein/DNA interaction within promoter sequences previously shown to be important for the activity of the U3 promoter. Nuclear extracts from rat 2 fibroblasts, which do not express the PTH/PTHrP receptor, produced one site of protein/DNA interaction which was found at a position on the promoter identical to the position of FP1 produced by a ROS 17/2.8 nuclear extract. Mutation of these two sites of protein/DNA interaction resulted in reduced U3 promoter activity. Furthermore, we have demonstrated that the transcription factors SP1 and MAZ regulate U3 promoter expression and have shown their functional significance using mutational analysis. These data demonstrate that SP1 and MAZ bind to the PTH/PTHrP receptor promoter and that they are involved in cell-specific expression of its gene product. PMID- 11116211 TI - Characterization of glucagon-like peptide-1 receptor-binding determinants. AB - Glucagon-like peptide 1 (GLP-1) is a potent insulinotropic hormone currently under study as a therapeutic agent for type 2 diabetes. Since an understanding of the molecular mechanisms leading to high-affinity receptor (R) binding and activation may facilitate the development of more potent GLP-1R agonists, we have localized specific regions of GLP-1R required for binding. The purified N terminal fragment (hereafter referred to as NT) of the GLP-1R produced in either insect (Sf9) or mammalian (COS-7) cells was shown to bind GLP-1. The physical interaction of NT with GLP-1 was first demonstrated by cross-linking ((125)I-GLP 1/NT complex band at approximately 28 kDa) and secondly by attachment to Ni(2+) NTA beads. The GLP-1R NT protein attached to beads bound GLP-1, but with lower affinity (inhibitory concentration (IC(50)): 4.5 x 10(-7) M) than wild-type (WT) GLP-1R (IC(50): 5.2 x 10(-9)M). The low affinity of GLP-1R NT suggested that other receptor domains may contribute to GLP-1 binding. This was supported by studies using chimeric glucose-dependent insulinotropic polypeptide (GIP)/GLP-1 receptors. GIP(1-151)/GLP-1R, but not GIP(1-222)/GLP-1R, exhibited specific GLP-1 binding and GLP-1-induced cAMP production, suggesting that the region encompassing transmembrane (TM) domain 1 through to TM3 was required for binding. Since it was hypothesized that certain charged or polar amino acids in this region might be involved in binding, these residues (TM2-TM3) were analyzed by substitution mutagenesis. Five mutants (K197A, D198A, K202A, D215A, R227A) displayed remarkably reduced binding affinity. These studies indicate that the NT domain of the GLP-1R is able to bind GLP-1, but charged residues concentrated at the distal TM2/extracellular loop-1 (EC1) interface (K197, D198, K202) and in EC1 (D215 and R227) probably contribute to the binding determinants of the GLP-1R. PMID- 11116212 TI - Transgenic rainbow trout expressed sGnRH-antisense RNA under the control of sGnRH promoter of Atlantic salmon. AB - A recombinant vector containing antisense DNA complementary to Atlantic salmon (Salmo salar) sGnRH cDNA driven by specific promoter Pab derived from a corresponding sGnRH gene was introduced into rainbow trout (Oncorhynchus mykiss) eggs. This resulted in transgenic animals that had integrated one copy of the transgene into their genome and transmitted it through the germline. Antisense sGnRH mRNA (AS) was expressed mainly in the brain of transgenic AS(+) fish. Levels of sGnRH endogenous mRNA in the brain were lower in 11-month-old AS(+) fish compared with nontransgenic AS(-) individuals from the same F2 progeny. sGnRH levels significantly decreased in the pituitary of transgenic males and females around the maturation period and in the brain of AS(+) immature females compared with controls. No reliable statistical difference was found in the levels of FSH and LH between AS(+) and AS(-) groups either in immature or mature fish. The majority of transgenic fish reached maturity at the same time as did nontransgenic individuals, although the maturation of AS(+) animals seemed to be more asynchronous. For the first time, the influence of antisense messengers on endogenous mRNA in transgenic fish and the corresponding protein is described. PMID- 11116213 TI - Transient prenatal expression of NPY-Y1 receptor in trigeminal axons innervating the mystacial vibrissae. AB - Using immunohistochemistry in combination with confocal laser scanning microscopy, we studied the ontogeny of neuropeptide Y-Y1 receptor (Y1-R) expression in the trigeminal system of the rat. The study was limited to the nerve fibers innervating the mystacial pad and the trigeminal ganglia. In the trigeminal ganglia, Y1-R-immunoreactive (IR) neurons were first observed at E16.5. At this same stage some nerve fibers in the trigeminal ganglia also exhibited Y1-R-like immunoreactivity (LI). Strongly Y1-R-IR nerve fibers innervating the follicles of the mystacial vibrissae were first observed at E18. After double labeling, the Y1-R-LI was found to be colocalized with the neuronal marker protein gene product 9.5. At P1 only weak labeling for the Y1-R was found around the vibrissae follicles, whereas the neurons in the trigeminal ganglia were intensely labeled. The same was true for the adult rat, but at this stage no Y1-R labeling at all was observed in nerve fibers around the vibrissal follicles. These results strongly support an axonal localization of the Y1-R at this developmental stage. The transient expression of the Y1-R during prenatal mystacial pad development suggests a role for the Y1-R in the functional development of the vibrissae. PMID- 11116214 TI - The cytoarchitecture of the nucleus angularis of the barn owl (Tyto alba). AB - The cochlear nucleus angularis (NA) of the barn owl (Tyto alba) was analyzed using Golgi, Nissl, and tract tracing techniques. NA forms a column of cells in the dorsolateral brainstem that partly overlaps with, and is rostral and lateral to, the cochlear nucleus magnocellularis (NM). Highest best frequencies are mapped in lateral NA (NAl), intermediate in medial NA (NAm), and lowest in the foot region (NAf). Cell density followed the tonotopic axis and decreased with decreasing best frequency. NA contained four major cell classes: planar, radiate, vertical, and stubby. Planar and radiate classes were further subdivided into bipolar and multipolar types according to their number of primary dendrites. Planar neurons were confined to an isofrequency band, whereas radiate neurons had dendrites that could extend across an isofrequency band. Vertical cells had long dendrites oriented perpendicularly to isofrequency bands. Stubby cells were the most numerous and were confined to an isofrequency band because of their short dendrites. Neurons in each of these four classes projected to the inferior colliculus and dorsal nucleus of the lateral lemniscus. PMID- 11116215 TI - Origins of projections from the inferior colliculus to the cochlear nucleus in guinea pigs. AB - We used retrograde tracing techniques to examine the projections from the inferior colliculus to the cochlear nucleus in guinea pigs. Following injection of a retrograde tracer into one cochlear nucleus, labeled cells were found bilaterally in all subdivisions of the inferior colliculus. The majority of cells were located in the central nucleus and external cortex; relatively few cells were located in the dorsal cortex. Multipolar (stellate) cells were labeled in all subdivisions of the inferior colliculus. In the central nucleus, disk-shaped cells were also labeled. To determine whether individual collicular neurons send collateral projections to the cochlear nuclei on both sides, we injected different fluorescent tracers into left and right cochlear nuclei in the same animal. The inferior colliculi contained very few double-labeled cells, indicating that the projections to ipsilateral and contralateral cochlear nuclei originate from separate populations of cells. PMID- 11116217 TI - Immunohistochemical localization and biochemical characterization of hypocretin/orexin-related peptides in the central nervous system of the frog Rana ridibunda. AB - In the present study, we have investigated the distribution and biochemical characteristics of hypocretin (hcrt) -like immunoreactivity in the central nervous system (CNS) of the frog Rana ridibunda by using an antiserum directed against rat hcrt2. Immunoreactive cell bodies were only detected in four diencephalic nuclei, including the anterior preoptic area and the suprachiasmatic, magnocellular, and ventral hypothalamic nuclei. In contrast, hcrt2-immunoreactive fibers were widely distributed throughout the frog CNS. In particular, a high density of hcrt-positive fibers was detected in several areas of the telencephalon, including the olfactory bulb, the nucleus of the diagonal band of Broca, and the amygdala. A dense network of hcrt-containing fibers was observed in all thalamic and hypothalamic nuclei. A low to moderate density of immunoreactive fibers was also found in the mesencephalon, rhombencephalon, and spinal cord. Reversed-phase high performance liquid chromatography analysis of frog brain extracts revealed that hcrt2-immunoreactive material eluted as two peaks, the major one exhibiting the same retention time as synthetic rat hcrt2. The present data provide the first detailed mapping of the hcrt neuronal system in the CNS of a nonmammalian vertebrate. The occurrence of hcrt-containing cell bodies in the hypothalamus and the widespread distribution of hcrt-immunoreactive fibers throughout the brain and spinal cord suggest that, in amphibians, hcrts may exert neuroendocrine, neurotransmitter, and/or neuromodulator activities. PMID- 11116216 TI - Long-term neurochemical changes after visual cortical lesions in the adult cat. AB - Peripheral deafferentation alters cortical function and such alterations have been shown to affect the cortical expression of the calcium-binding proteins calbindin and parvalbumin and of the inhibitory neurotransmitter gamma aminobutyric acid (GABA). To determine whether cortical deafferentation produces similar effects, we examined the long-term consequences of cortical lesions on the neurochemistry of interconnected cortical areas. We studied the reciprocal effects of localized damage to either visual cortical areas 17 and 18, or posteromedial lateral suprasylvian (PMLS) cortex in the adult cat. These areas are strongly interconnected and play an important role in the processing of visual information. Combined lesions of areas 17 and 18 caused a marked, topographically specific decrease in the proportion of neurons expressing calbindin in supragranular layers of PMLS cortex. Similarly, lesions of PMLS cortex caused topographically restricted decreases in calbindin expression within supragranular layers of areas 17 and 18, but not in other cortical areas with which PMLS is interconnected. To categorize the calbindin-positive neurons affected by such lesions, we carried out double-labeling experiments for the inhibitory neurotransmitter GABA. This investigation showed lesions of areas 17 and 18 to affect calbindin-positive excitatory and inhibitory neurons equally, but PMLS lesions had stronger effects on inhibitory, calbindin-positive neurons. This finding may represent differential damage to feed-forward vs. feed-back projections in the two types of lesions. Finally, the expression of parvalbumin and GABA was unchanged, even in zones of decreased calbindin immunoreactivity. Our results suggest that damage to adult visual cortical areas, whether striate or extrastriate, induces neurochemical changes in the supragranular corticocortical network to which these areas belong. That changes were restricted to calbindin expression suggests cell-specific and/or biochemical pathway specific alterations in calcium homeostasis. PMID- 11116218 TI - Confocal microscopic study of glial-vascular relationships in the retinas of pigmented rats. AB - Astroglia are interposed between the cerebral vasculature and neurons, where they may mediate the transfer of substances from the circulation to neurons and couple changes in neuronal activity to changes in cerebral blood flow. The retina is a particularly advantageous model system for studying glial-vascular interactions in situ. Confocal microscopy and three-dimensional image reconstruction were used to study the anatomical relationships between glia and the surface vasculature in retinas acutely isolated from adult pigmented rats. Retinas were immunostained using antibodies directed against the basal lamina surrounding the vasculature as well as antibodies directed against glial fibrillary acidic protein. Surface vessels of all calibers were contacted by the processes of astrocytes. The vitreal surfaces of the large retinal vessels were covered by a meshwork of immunoreactive astrocyte processes of a variety of shapes, whereas the scleral surfaces of the vessels were supported by thick bundles of astrocyte processes. In addition, glial cells were filled intracellularly with the gap junction permeable tracers Lucifer yellow and Neurobiotin. Intracellular fills clearly demonstrated the presence of astrocytes with somata that were closely apposed to the large retinal vessels. Tracer-filled astrocytes displayed a variety and complexity of shapes that was not apparent in immunostained material. Gap junctional coupling was stronger between astrocytes adjacent to the same artery than between periarterial astrocytes and astrocytes located away from arteries. Significantly fewer Muller cells were labeled when Neurobiotin was injected into astrocytes associated with arteries than when Neurobiotin was injected into astrocytes that were distant from arteries. PMID- 11116219 TI - Developmental and sensory-dependent changes of group II metabotropic glutamate receptors. AB - Metabotropic glutamate receptors (mGluRs) are potential participants of sensory dependent modification of neural connections. Here, we examined the involvement of cAMP-linked mGluRs (mGluR2/3) in sensory-dependent plasticity by studying the correlation of mGluR2/3 changes with the critical period of ocular dominance plasticity, a form of sensory-dependent plasticity, and exploring the effects of dark rearing on mGluR2/3 in the primary visual cortex of cats. Immunohistochemistry showed that the laminar distribution of mGluR2/3 changed with the critical period and was sensitive to dark rearing. The mGluR2/3 immunostaining became most intense in layer IV at the beginning of the critical period and was reduced in layer IV but became intense in layers I-III at the peak of the period, then was concentrated primarily in layers I-upper III at the end of the critical period. Dark rearing delayed these pattern changes for weeks and elevated the normally declining mGluR2/3 quantity shortly after the peak of the critical period. The effects of dark rearing and the correlation of early mGluR2/3 laminar changes with geniculocortical afferent segregation indicate that mGluR2/3 circuitry in the visual cortex is influenced by visual inputs. Our data suggest that mGluR2/3 together with another sensory-influenced mGluR, mGluR5, may participate in the sensory-dependent modification of neural connections in the visual cortex. PMID- 11116220 TI - Visualization of synaptic markers in the optic neuropils of Drosophila using a new constrained deconvolution method. AB - The fruitfly Drosophila melanogaster offers compelling genetic advantages for the analysis of its nervous system, but cell size precludes immunocytochemical analysis of wild-type structure and mutant phenotypes beyond the level of neuronal arborizations. For many antibodies, especially when immunoelectron microscopy is not feasible, it would therefore be desirable to extend the resolution limit of confocal microscopy as far as possible. Because high resolution confocal microscopy suffers from considerable blurring, so-called deconvolution algorithms are needed to remove, at least partially, the blur introduced by the microscope and by the specimen itself. Here, we present the establishment and application of a new deconvolution method to visualize synaptic markers in Drosophila optic neuropils at the resolution limit of light. We ascertained all necessary parameters experimentally and verified them by deconvolving injected fluorescent microspheres in immunostained optic lobe tissue. The resulting deconvolution method was used to analyze colocalization between the synaptic vesicle marker neuronal synaptobrevin and synaptic and putative synaptic markers in photoreceptor terminals. We report differential localization of these near the resolution limit of light, which could not be distinguished without deconvolution. PMID- 11116221 TI - Innervation of supporting cells in the apical turns of the guinea pig cochlea is from type II afferent fibers. AB - The outer supporting cells in the apical turns of the guinea pig cochlea receive a dense innervation. Our previous study (Fechner et al. [1998] J. Comp. Neurol. 400:299-300) suggested that this innervation of the Deiters' and Hensen's supporting cells was not derived from efferent fibers of the olivocochlear bundle, but its origin has not been further specified. To test the hypothesis that the innervation was afferent in origin, we traced apical afferent fibers that were retrogradely labeled by extracellular injections of horseradish peroxidase. Labeled afferent fibers were of two types: type I fibers contacted inner hair cells, whereas type II fibers crossed the tunnel and contacted outer hair cells. Significantly, most of the type II fibers also formed branches to the outer supporting cells. Although a few olivocochlear efferent fibers formed such branches, counts indicated that the overwhelming majority of the branches were produced by type II afferent fibers. These branches were not produced by basal type II fibers. Apical type II fibers also differed from basal fibers by having shorter lengths, spiraling both apically and basally, and contacting all three rows of outer hair cells. These innervation differences suggest differences in the ways that information from outer hair cells is processed in the apex versus the base of the cochlea. PMID- 11116222 TI - Development of thyrotropin-releasing hormone immunoreactivity in the brain of the brown trout Salmo trutta fario. AB - The development and adult distribution of thyrotropin-releasing hormone immunoreactive (TRHir) neurons in the brain of the brown trout, Salmo trutta fario, was studied with the streptavidin-biotin immunohistochemical method. Study of embryos, alevin, and juveniles revealed groups of TRHir neurons in the mesencephalon and rhombencephalon that have not been noted previously in adult teleosts. The earliest TRHir cells observed were those of the trigeminal motor nucleus, which expressed this substance only in embryos and alevins. In the forebrain, early-arising TRH populations were observed in the supra- and postcommissural region of the ventral telencephalic area, the anterior parvocellular preoptic nucleus, the organon vasculosum laminae terminalis, the magnocellular preoptic nucleus, the suprachiasmatic nucleus, and the posterior tuberal nucleus. TRHir cells of the olfactory bulb, abundant in the adult, appeared later. A small TRHir neuronal population was transiently observed in the habenula of alevins and juveniles. The laminar nucleus of the mesencephalon contained a small population of TRH cells in alevins and juveniles. In the isthmus, TRH was observed in cells of the interpeduncular nucleus, the nucleus isthmi, the dorsolateral tegmental nucleus, the superior reticular nucleus, and the central gray, although perikarya were TRHir only in alevin and/or juvenile stages. Some vagal motoneurons were TRHir from the late embryo stage onward. TRHir fibers were abundant in several forebrain regions of alevins and juveniles, including the medial region of the dorsal telencephalic area, the ventral telencephalic area and commissural region, the preoptic neuropil, the posterior tubercle, the anterior tuberal nucleus, and the posterior hypothalamic lobe. TRHir fibers invaded the neurohypophysis in early alevins, and their number increased subsequently to adulthood. The parvocellular superficial pretectal nucleus and the optic tectum received a rich TRHir innervation from juvenile stages onward. The interpeduncular nucleus and the secondary gustatory nucleus contained many TRHir fibers. In the rhombencephalon, TRHir fibers were scarce, except in the ventrolateral regions and the inferior olive. The distribution of TRHir fibers suggests that they were mainly related to hypophysiotropic and visceral centers, although the presence of TRH in centers related to the visual system indicates that TRH also plays other roles in the brain. We discuss the possibility that the strong expression of TRH in the embryonic trigeminal motoneurons plays a role in head morphogenesis. PMID- 11116223 TI - Organization of thalamostriatal terminals from the ventral motor nuclei in the macaque. AB - This study examines the organization of thalamostriatal projections from ventral tier nuclei that relay basal ganglia output to the frontal cortex. Although previous thalamostriatal studies emphasize projections from the intralaminar nuclei, studies in primates show a substantial projection from the ventral anterior (VA) and ventral lateral (VL) nuclei. These nuclei make up the main efferent projection from the basal ganglia to frontal cortical areas, including primary motor, supplementary, premotor, and cingulate motor areas. Functionally related motor areas of the frontal cortex and VA/VL have convergent projections to specific regions of the dorsal striatum. The distribution of VA/VL terminals within the striatum is crucial to understanding their relationship to motor cortical afferents. We placed anterograde tracer injections into discrete VA/VL thalamic areas. VA/VL thalamostriatal projections terminate in broad, rostrocaudal regions of the dorsal striatum, corresponding to regions innervated by functionally related cortical motor areas. The pars oralis division of VL projects primarily to the dorsolateral, postcommissural putamen, whereas the parvicellular VA targets more medial and rostral putamen regions, and the magnocellular division of VA targets the dorsal head of the caudate nucleus. Whereas these results demonstrate a general functional topography, specific VA/VL projections overlap extensively, suggesting that functionally distinct VA/VL projections may also converge in dorsal striatal areas. Within striatal territories, VA/VL projections terminate in a patchy, nonhomogeneous manner, indicating another level of complexity. Moreover, terminal fields contain both terminal clusters and scattered, long, unbranched fibers with many varicosities. These fiber morphologies resemble those from the cortex and raise the possibility that VA/VL thalamostriatal projections neurons have divergent connectional features. PMID- 11116224 TI - gamma-Aminobutyric acid transporters in the cat periaqueductal gray: a light and electron microscopic immunocytochemical study. AB - The gamma-aminobutyric acid (GABA) plasma membrane transporters (GATs) mediate GABA uptake into presynaptic axon terminals and glial processes, thus contributing to the regulation of the magnitude and duration of the action of GABA at the synaptic cleft. The aim of the present study was to investigate the expression of three high-affinity GABA transporters (GAT-1, GAT-2, and GAT-3) in the periaqueductal gray matter (PAG) of adult cats by using immunocytochemistry with affinity-purified antibodies. Light microscopic observations revealed GAT-1 immunoreactivity in punctate structures, particularly dense in the lateral portion of the dorsolateral PAG column. Weak GAT-2-immunopositive puncta were homogeneously distributed in the PAG. GAT-3 immunoreactivity was detected in each column of the PAG but was more intense in the dorsolateral PAG column and around the aqueduct. Electron microscopic studies showed GAT-1 immunoreactivity in distal astroglial processes, in unmyelinated and small myelinated axons, and in axon terminals making symmetric synapses on both PAG neurons and dendrites. GAT-2 immunoreactivity was present mostly in the form of patches of different sizes in the cytoplasm of neuronal elements like the perikarya and dendrites of PAG neurons, in myelinated and unmyelinated axons, and in the axon terminals forming both symmetric and asymmetric synapses. Labeling was also observed in nonneuronal elements. Astrocytic cell bodies and their distal processes as well as the ependymal cells lining the wall of the aqueduct showed patches of GAT-2 immunoreactivity. Electron microscopic observation revealed GAT-3 immunoreactivity exclusively in distal astrocytic processes adjacent to the somata of PAG neurons and in axon terminals making both symmetric and asymmetric synapses. The present results suggest that three types of termination systems of GABAergic transmission are present in the cat periaqueductal gray matter. PMID- 11116225 TI - Ultrastructural evidence that hippocampal alpha estrogen receptors are located at extranuclear sites. AB - Estrogen may mediate some of its effects on hippocampal function through the alpha isoform of the estrogen receptor (ERalpha). By light microscopy, ERalpha immunoreactivity (-I) is found in the nuclei of scattered inhibitory gamma aminobutyric acid (GABA)ergic interneurons. However, several lines of evidence indicate that estrogen also may exert some of its effects through rapid nongenomic mechanisms, possibly by binding to plasma membranes. Thus, to determine whether ERalpha is found in extranuclear sites in the hippocampal formation (HF), four different antibodies to ERalpha were localized by immunoelectron microscopy in proestrous rats. Ultrastructural analysis revealed that in addition to interneuronal nuclei, ERalpha-I was affiliated with the cytoplasmic plasmalemma of select interneurons and with endosomes of a subset of principal (pyramidal and granule) cells. Moreover, ERalpha labeling was found in profiles dispersed throughout the HF, but slightly more numerous in CA1 stratum radiatum. Approximately 50% of the ERalpha-labeled profiles were unmyelinated axons and axon terminals that contained numerous small, synaptic vesicles. ERalpha-labeled terminals formed both asymmetric and symmetric synapses on dendritic shafts and spines, suggesting that ERalphas arise from sources in addition to inhibitory interneurons. About 25% of the ERalpha-I was found in dendritic spines, many originating from principal cells. Within spines, ERalpha-I often was associated with spine apparati and/or polyribosomes, suggesting that estrogen might act locally through the ERalpha to influence calcium availability, protein translation, or synaptic growth. The remaining 25% of ERalpha-labeled profiles were astrocytes, often located near the spines of principal cells. Collectively, these results suggest that ERalpha may serve as both a genomic and nongenomic transducer of estrogen action in the HF. PMID- 11116226 TI - Size gradients of barreloids in the rat thalamus. AB - The spatial organization of the anatomical structures along the trigeminal afferent pathway of the rat conserves the topographical order of the receptor sheath: The brainstem barrelettes, thalamic barreloids, and cortical barrels all reflect the arrangement of whiskers across the mystacial pad. Although both the amount of innervation in the mystacial pad and the size of cortical barrels were shown previously to exhibit increasing gradients toward the ventral and caudal whiskers, whether similar gradients existed in the brainstem and thalamus was not known. Here, the authors investigated the size gradients of the barreloids in the ventral posteromedial nucleus of the rat thalamus. Because the angles used to cut the brain were crucial to this study, the optimal cutting angles were determined first for visualization of individual barreloids and of the entire barreloid field. Individual barreloids, arcs, and rows as well as entire barreloid fields were clearly visualized using cytochrome oxidase staining of brain slices that were cut with the optimal cutting angles. For the first five arcs (including straddlers), the length of barreloids increased in the direction of dorsal-to ventral whiskers and of caudal-to-rostral whiskers. These gradients reveal an inverse relationship between the size of barreloids and whiskers (length and follicle diameter) along arcs and rows. The largest barreloids in the ventral posteromedial nucleus were those that represent whiskers C2-C4, D2-D4, and E2-E4, which are neither the largest nor the most innervated whiskers in the mystacial pad. This implies that the extended representation is not merely a reflection of peripheral innervation biases and probably serves an as yet unknown processing function. PMID- 11116227 TI - Expression of kalirin, a neuronal GDP/GTP exchange factor of the trio family, in the central nervous system of the adult rat. AB - Kalirin is a multifunctional protein identified by its interaction with peptidylglycine alpha-amidating monooxygenase, an enzyme essential for neuropeptide biosynthesis. Several forms of Kalirin exist, all containing spectrin-like repeats, a Dbl homology (DH) domain, and an adjacent pleckstrin homology (PH) domain; several different COOH-termini provide additional DH/PH domains and a putative protein kinase. Kalirin binds Rac1 and affects cytoskeletal organization, neuropeptide secretion, and iNOS activity. By in situ hybridization, the highest levels of Kalirin mRNA were found in the cerebral cortex, hippocampal formation, and Purkinje cells, with high levels also in thalamus, caudate putamen, septal nucleus, nucleus accumbens, amygdala, and anterior olfactory nucleus. Low levels of Kalirin mRNA were detected in the paraventricular, supraoptic, and reticular thalamic nuclei and in the ventromedial hypothalamic nucleus. Brain areas with high levels of Kalirin mRNA showed strong Kalirin-like immunoreactivity. Pyramidal neurons with strongly staining soma and long dendrites were observed primarily in layer 5 of the cerebral cortex. In the hippocampus, a uniform distribution of neurons with fine dendritic staining was observed in the pyramidal cell layer, in the granule cell layer, and in the hilar cells of the dentate gyrus as well as in isolated interneurons. Cerebellar Purkinje neurons exhibited intense staining in the soma and in extensive dendritic arbors extending to the surface of the molecular layer. During embryonic development, Trio, the Drosophila orthologue of Kalirin, plays an essential role in axon guidance; localization of Kalirin to the somatodendritic region of adult neurons provides the basis for future studies of regulation and function. PMID- 11116228 TI - Spinocerebellar projections in the pigeon with special reference to the neck region of the body. AB - Neck sensory information is important for control of head and body movements in all vertebrates. Neuroanatomic tracing methods were used to study the pathways of neck afferent systems. Both the projection of primary afferent fibers and of secondary afferent pathways to brainstem and cerebellum were investigated with the anterograde transport of dextran amines as tracers (biotinylated dextran amine and tetramethyl rhodamine dextran amine). For comparison, the projections of spinocerebellar systems of wing and leg were studied also. Complementary experiments using retrograde tracers (Fast Blue, tetramethyl rhodamine dextran amine, rhodamine isothiocyanate) injected into the cerebellum served to corroborate the results of the anterograde tracing experiments. Primary neck afferent fibers terminated in the spinal gray substance with dense terminal fields in laminae I to V of the dorsal horn and lamina IX of the ventral horn as well as in the marginal nuclei located at the lateral border of the spinal cord. In the brainstem, dense terminal fields were seen in deep layers of the medullary dorsal horn, in the external cuneate nucleus, and in group x. Secondary neck afferents arising from ventral horn cells showed a significant projection to the descending and medial vestibular nuclei and to the medial cerebellar nucleus. Terminals were found both in the anterior and the posterior cerebellum. A quantitative evaluation disclosed that most terminals of neck afferents distributed in lobules II-IV of the anterior cerebellum and lobule IX of the posterior cerebellum. With injections aimed at spinocerebellar neurons located into the cervical and lumbosacral enlargements, no projections were found in the vestibular or deep cerebellar nuclei. Projections from the cervical enlargement were concentrated in lobules III-V and those from the lumbosacral enlargement in lobules III-VI. This points to a rostrocaudal somatotopic representation of neck, wing, and leg in the anterior cerebellum. The results of the retrograde tracing experiments support such a somatotopic organization. PMID- 11116229 TI - Neurochemical differentiation of functionally distinct populations of autonomic neurons. AB - The coeliac ganglion of guinea pigs displays a unique topographical arrangement of neurochemically and functionally distinct populations of sympathetic neurons. The authors used multiple-labeling immunohistochemistry to investigate the neurochemical differentiation of these neurons during embryonic and fetal development. Sympathoadrenal precursors, located on either side of the abdominal aorta, were intensely immunoreactive for tyrosine hydroxylase (TH-IR), neurofilament, and the human natural killer 1 antibody at midembryonic stages (Carnegie stages 16-19). During late embryonic stages (stages 20-23), a single bilobed ganglion had formed. At this time, neuropeptide Y immunoreactivity (NPY IR) was widely expressed in sympathetic neurons (with moderate TH-IR) and chromaffin cells (with intense TH-IR). The onset of somatostatin (Som-IR) expression followed that of NPY-IR and was restricted to sympathetic neurons. However, at late embryonic stages, most TH-IR neurons with Som-IR also expressed NPY-IR (a combination of peptides not found in the mature coeliac ganglion). Between late embryonic stages and the end of the early fetal period, there was a significant increase in the proportion of neurons in lateral regions that had both NPY-IR and TH-IR. At the same time, there was an increase in the proportion of neurons in medial regions that had both Som-IR and TH-IR. Neurons expressing both Som-IR and TH-IR were rarely observed in lateral regions of the coeliac ganglion. Thus, a clear topography within the coeliac ganglion is established during late embryonic and early fetal stages of development and reflects that found in the mature animal by the end of the early fetal period. PMID- 11116230 TI - Distribution of tyrosine hydroxylase-containing neurons and fibers in the brain of the budgerigar (Melopsittacus undulatus): general patterns and labeling in vocal control nuclei. AB - The distribution of tyrosine hydroxylase (TH) was mapped out in cells and fibers of the budgerigar (Melopsittacus undulatus) brain. Special attention was given to vocal control and auditory nuclei because budgerigars are a psittacine species in which both males and females are capable of lifelong vocal learning (Farabaugh et al. [1994] J. Comp. Psychol 108:81-92). The results show that TH staining in the central nucleus of the anterior archistriatum (AAc) resembled that of surrounding archistriatal fields, except for portions of the ventral archistriatum, which exhibited substantially more TH+ fibers. Fewer fibers and fiber baskets are present in the central nucleus of the lateral neostriatum (NLc) than in surrounding fields. Both the oval nuclei of the ventral hyperstriatum (HVo) and anterior neostriatum (NAo) exhibit less fiber staining than surrounding fields whereas fiber staining in the medial NAo (NAom) and magnicellular nucleus of the parolfactory lobe (LPOm) resemble that of surrounding fields. Staining in primary telencephalic auditory nuclei was extremely low. The only sex difference observed was slightly increased TH staining in LPOm of females compared with surrounding fields on some tissue sections. These findings are in contrast to previous findings in zebra finch (Poephila guttata), a close ended vocal learning songbird in which TH staining in vocal nuclei increases during development and remains greater than surrounding fields throughout adulthood. The present results therefore support the view that catecholamines act to inhibit vocal plasticity in adult vocal learning species. Several unique features of TH-immunoreactive (ir) cell groups were observed in the brainstem including sparsely scattered TH-ir somata immediately adjacent to the third ventricle, within the tectum, basal forebrain, archistriatum, and caudal neostriatum, and in the hippocampus. These latter populations have not been described in other avian species and resemble features of the catecholamine system generally found in either reptiles or mammals. PMID- 11116231 TI - Anatomic correlates of the face and oral cavity representations in the somatosensory cortical area 3b of monkeys. AB - We determined the somatotopy of the face and the oral cavity representation in cortical area 3b of New World owl monkeys and squirrel monkeys. Area 3b is apparent as a densely myelinated strip in brain sections cut parallel to the surface of flattened cortex. A narrow myelin-light septum that we have termed the "hand-face septum" separates the hand representation from the more lateral face and mouth representation. The face and oral cavity representation is further divided into a series of myelin-dense ovals. We show that three ovals adjacent to the hand representation correspond to the upper face, upper lip, and chin plus lower lip, whereas three or four more rostral ovals successively represent the contralateral teeth, tongue, and the ipsilateral teeth and tongue. Strips of cortex lateral and medial to the area 3b ovals, possibly corresponding to area 1 and area 3a, respectively, have similar somatotopic sequences. Although previous results suggest the existence of great variability within and across primate species, we conclude that the representations of the face and mouth are highly similar across individuals of the same species, and there are extensive overall similarities across these two species of New World monkeys. PMID- 11116232 TI - Comparative distribution of mRNA encoding the growth hormone secretagogue receptor (GHS-R) in Microcebus murinus (Primate, lemurian) and rat forebrain and pituitary. AB - The forebrain and pituitary sites of synthesis of growth hormone secretagogue receptor mRNA were identified in four adult lemurs (Microcebus murinus) by in situ hybridisation performed with a radiolabeled cRNA probe transcribed from human Growth Hormone Secretagogue-Receptor cDNA. The cRNA sense and antisense probes were hybridised to cryostat sections containing structures extending from the rostral hypothalamus to its caudal limit as defined by the mammillary bodies. The pituitary gland and areas adjacent to the hypothalamus were also analyzed. For comparative purposes, sections from five adult rats containing these structures were hybridised with the same probes. The results point to a widespread distribution of Growth Hormone Secretagogue-Receptor mRNA in the hypothalamus, hippocampal formation, and cerebellar cortex of both lemurs and rats. As in the rat, specific hybridisation was particularly dense in the arcuate nucleus. Significant species differences were observed in the periventricular nucleus, the ventromedial nucleus, the lateral hypothalamic area, and the pituitary gland. In contrast to the rat, the lemur exhibited marked labelling in the infundibular nucleus, the periventricular nucleus and the pars tuberalis of the pituitary gland, whereas no labeling was detectable in the ventromedial nucleus and the lateral hypothalamic area. These results are discussed in terms of difference between the control of growth hormone secretion, feeding behaviour and seasonal rhythmicity among murine species and primates. PMID- 11116233 TI - Course of motor and associative pallidothalamic projections in monkeys. AB - As a result of the frequent performance of lesioning and electrical stimulation procedures targeting the globus pallidus internus (GPi) to treat medically intractable hypokinetic and hyperkinetic movement disorders, the course of the pallidothalamic projections originating, in particular, from the motor territory of GPi has important clinical relevancy. To assess the organization of pallidothalamic projections originating from motor and associative portions of GPi, small quantities of the anterograde/ retrograde tracer, biotinylated dextran amine (BDA) were injected into localized regions of the caudal GPi in squirrel monkeys. The localization to motor and associative territories in GPi was confirmed by examining the corresponding regions of retrograde labeling in the striatum and subthalamic nucleus (STN). The labeled pallidothalamic fibers projected principally medially across the inferior edge of the internal capsule. The fiber bundle ventral to the caudal GPi was mainly devoid of labeling. Fibers labeled along the medial and inferior borders of GPi at centrorostral levels were traceable to the medial edge of the injections. The densest fiber labeling at rostral levels was produced by those injections with the greatest extent of rostral labeling of neurons. In opposition to generally accepted schemes, the findings from this study suggest that the pallidothalamic fibers originating from the caudal portions of GPi, including the motor territory, do not course ventromedially to form the ansa lenticularis, but rather, travel predominately medially through the lenticular fasciculus en route to the thalamus. Thus, proposed surgical schemes to target fibers ventral to the caudal GPi or at the rostral pole of GPi appear to be misguided. PMID- 11116234 TI - Projections from the nucleus of the basal optic root and nucleus lentiformis mesencephali to the inferior olive in pigeons (Columba livia). AB - The nucleus of the basal optic root (nBOR) of the accessory optic system (AOS) and the pretectal nucleus lentiformis mesencephali (LM) are involved in the analysis of optic flow and the generation of the optokinetic response. Previous studies have shown that the nBOR projects bilaterally to the medial column (mc) of the inferior olive (IO) and the LM projects to the ipsilateral mc. In the present study the retrograde tracer cholera toxin subunit B was injected into either the caudal or rostral mc. From all injections, retrogradely labeled cells were seen in the ipsilateral pretectum along the border of the medial and lateral subnuclei of the LM. Cells were also seen in bilaterally in the nBOR. On the contralateral side, a discrete group of cells was labeled in the rostral margin of the nBOR. These cells were localized in the dorsal portion of the nBOR proper and some were found in the adjacent nBOR dorsalis. On the ipsilateral side, a diffuse group of cells was seen in the caudal nBOR. Most of these cells were in the nBOR dorsalis and outside the nBOR complex in the area ventralis of Tsai and the reticular formation. From the injections into the caudal mc, a greater proportion of labeled cells was found in the LM, whereas a greater proportion of cells was found in the nBOR from the injections into the rostral mc. This differential projection from LM and nBOR to the caudal and rostral mc is consistent with the optic flow preferences of neurons in the mc, and a similar pattern of connectivity has been found in mammalian species. PMID- 11116235 TI - Comparative genomic hybridization analysis of cisplatin-resistant ovarian carcinoma cells. PMID- 11116236 TI - Changes in neurological diseases in the last half century. PMID- 11116237 TI - Sequential gadolinium-enhanced magnetic resonance angiography of the aortoiliac and the femoropopliteal arteries with repetitive administration of low-dose contrast agent. AB - To obtain a wide-range contrast MR angiography in a single examination, we performed two sequential administrations of low-dose (0.08 mmol/kg) gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) with three dimensional inversion recovery prepared fast spoiled gradient recalled acquisition in the steady-state (3D IR-fast SPGR) sequence. Signal characteristics of the sequence were estimated by computed simulations and an in vitro study. A clinical study of 19 examinations was done with sequential MR angiography of the aortoiliac and femoropopliteal arteries. Great signal differences were observed between the high and low Gd concentrations. Higher Gd concentrations generated significantly stronger signals. Greater signals were produced at TIs of longer than 150 msec than at shorter than 100 msec. In the clinical study, the arteries were visualized with sufficient signals even with a small amount of contrast agent. Contrast-to-noise ratios between the arteries and surrounding skeletal muscles or fat tissues ranged from 10.5+/-9.6 to 4.7+/-2.2 and 6.6+/-2.8 to-3.1+/-11.2, respectively. No venous enhancement was found with diluted contrast agent on the second MR angiography. Two consecutive contrast MR angiographies can be obtained with repetitive administration of low-dose contrast agent. PMID- 11116238 TI - The effect of oral clonidine premedication on lumbar cerebrospinal fluid pressure in humans. AB - Alpha-2 adrenergic agonists including clonidine decrease cerebral blood flow. The specific actions of clonidine on cerebrospinal fluid (CSF) pressure in humans remain to be elucidated. We evaluated the effect of oral clonidine premedication on lumbar CSF pressure in patients without intracranial disease. Seventy-four patients undergoing subarachnoidal block were divided randomly into either a clonidine or a control group. In the clonidine group, the patients were premedicated orally with 5 microg/kg clonidine 60 min before arrival in the operating room. Subarachnoidal puncture was performed via midline approach using a 23-gauge needle at the L2-3 or L3-4 intervertebral space with the patient in the lateral decubitus position. Before the injection of local anesthetic, lumbar CSF pressure was measured. Lumbar CSF pressure was 8.1+/-2.4 mmHg in the clonidine group, which was significantly lower than that in the control group (9.4+/-2.8 mmHg, p<0.05). The cerebral perfusion pressures were 76.2+/-12.5 mmHg in the clonidine group and 91.7+/- 15.4 mmHg in the control group (p<0.001). In the clonidine group, preanesthetic mean blood pressure had a significant correlation with lumbar CSF pressure (r=0.619, p=0.019). We conclude that Lumbar CSF pressure was attenuated by oral premedication with 5 microg/kg clonidine. Clonidine also contributed to a significant correlation between preanesthetic mean blood pressure and CSF pressure. PMID- 11116239 TI - Changes in maternal lipid peroxidation levels and antioxidant enzymatic activities before and after delivery. AB - OBJECTIVE: Our goal was to characterize the changes in maternal lipid peroxidation levels and antioxidant enzymatic activities before and after delivery. METHODS: Predelivery and 1, 24, and 48 hours post-partum plasma concentrations of malondialdehyde, erythrocyte enzyme superoxide dismutase, glutathione peroxidase and catalase were measured in uncomplicated pregnancies. RESULTS: Malondialdehyde levels increased slightly from predelivery to 24 hours post-partum and then decreased significantly at 48 hours post-partum. At one hour post-partum superoxide dismutase and catalase levels increased significantly to about 125% and 170% of predelivery levels, respectively. Thereafter, these values decreased significantly from one hour to 48 hours post-partum. The relative changes in superoxide dismutase and catalase levels at one hour post-partum compared to predelivery values correlated significantly with the duration of labor. CONCLUSION: The results suggest that the uncontrolled lipid peroxidation caused by reactive oxygen species, which are produced in consequence of tissue reoxygenation, may occur during labor and that prolonged labor, may cause maternal oxidative stress. PMID- 11116240 TI - [Do patients and/or physicians have a right to control medical information]. AB - An ongoing debate is underway on whether the right to control medical information required for objective evaluations and uncertain information should belong to the physician. In this study, the opinions of practicing physicians were examined. Information appearing on medical records was divided into 10 items, and physicians were asked whether the right to control each item belonged to the patient and/or the physician. More than 60% the physicians answered that both patients and physician have the right to control information such as the diagnosis, examination findings, informed consent, and treatment. All of these items can be categorized as personal data. On the other hand, more than 70% of the physicians believed that only physicians have the right to control information such as the process to reach a diagnosis, subjective patient information, uncertain information and discussion with other physicians. Those physicians who answered that that both patients and physicians have the right to control information such as subjective patient information were more in favor of the disclosure of medical records than the physicians who were against the patient's right to control such information (p<0.05). The results of this study suggest that the right to control medical information on medical records should be considered separately for each chart item. As the network of medical information advances, the right to control medical information will become a topic of increasing importance. PMID- 11116241 TI - Surgical treatment of coarctation complex in neonates and infants. AB - BACKGROUND: There remains controversy regarding the appropriate surgical treatment of coarctation of the aorta associated with intracardiac anomalies in neonates and infants. Furthermore, the relative benefits of one versus two-stage repair, and subclavian flap aortoplasty versus end-to-end anastomosis for some of these lesions, remain controversial. The purpose of this paper is to review our experience with two-stage repair using subclavian flap aortoplasty and to seek an appropriate procedure. METHODS AND RESULT: From June 1996 to November 1999, thirteen patients underwent subclavian flap aortoplasty in our department. The age range was 16 to 101 days (mean 52 days), and the body weight range was 1.9 to 4.5 kg (mean 3.0 kg). Anatomic diagnosis was coarctation with ventricular septal defect (six patients), double outlet right ventricle (two patients), atrioventricular canal defect (one patient), tricuspid atresia (two patients), mitral atresia (one patient), and single atrium and subaortic stenosis (one patient). There was one hospital death in our series due to the progression of pulmonary hypertension 3 months after the operation. The mean follow up for remaining twelve patients was 28 months (range 7 approximately 48 months). There was one reoperation for recurrent coarctation. Three patients underwent pulmonary artery plasty in a second operation because of right pulmonary artery stenosis. We performed the definitive operation for six patients with coarctation with ventricular septal defect and two patients with double outlet right ventricle, and we performed a bidirectional cavopulmonary shunt for four univentricular hearts who are candidates for the Fontan operation. Two patients required Damus Kaye-Stansel procedure to release restrictive bulboventricular foramen. Three patients underwent a modified Fontan operation after these palliations. In our series, the intraoperative mortality rate for subclavian flap aortoplasty was 0% and the post operative mortality rate was 7.7% (1/13). Ten patients underwent the final operation successfully, and further two patients are considered good candidates for the final operation. The overall mortality was 7.7% (1/13). CONCLUSION: Two-stage repair appears to offer a good prognosis for neonates and infants with a coarctation complex. Subclavian flap aortoplasty showed the lowest rate of restenosis. However, late mortality may be associated with the progression of pulmonary vascular disease and the presence of associated severe cardiac anomalies. Although Fontan candidates need staged operations, if biventricular repair is feasible, one-stage repair would be a reasonable procedure considering the progression of the pulmonary vascular disease and the distortion of the pulmonary artery due to pulmonary artery banding. It would appear to improve the quality of life of those children if a one-stage operation can be performed with reasonable risk and good midterm outcome. PMID- 11116243 TI - Septic arthritis of the hip associated with atopic dermatitis. A case report. AB - We report a case of septic arthritis of the hip associated with atopic dermatitis. A 15-year female felt a pain in the right hip with unknown cause on May 11, 1998. The pain subsequently became aggravated, and she was admitted to our hospital on May 18. She has had atopic dermatitis since 4 years of age. She showed generalized dermatitis with desquamation and numerous scratch marks. A culture of both skin and joint fluid revealed Staphylococcus aureus. Physical examination revealed tenderness in Scarpa triangle and restricted range of motion. Immunological serology showed an increase in eosinophils and immunoglobulin E, and a decreased reaction of lymphocyte blastoid transformation. Computed tomography (CT) and MRI showed a joint effusion in the right hip. She was diagnosed as having septic arthritis of the hip. Intravenous drip of Cefazolin of 2g was started on the first day of hospitalization and joint irrigation was done on the second day. CRP became negative at 4 weeks, but joint effusion was shown on CT. Additional joint irrigation with Amicamycin (200 mg) was done. As the joint fluid culture became negative, range of motion exercises were started at 6 weeks. She was discharged with a long-leg brace applied at 8 weeks. At 13 months after onset, she had complete relief of the pain and normal activities of daily living. No destructive changes in the hip were found on X-ray examination or MRI. In the present case, an abnormal immune system associated with atopic dermatitis as well as the habit of scratching eruptions may have led to hematogenous spread of skin infection, and caused septic arthritis of the hip. PMID- 11116242 TI - Transient osteoporosis of the hip during pregnancy. AB - We report the clinical features of and MRI findings in transient osteoporosis of the hip during pregnancy. The study population consisted of 4 patients with a mean age of 33 years. The mean gestational age at onset was 31 weeks (range: 27 to 35 weeks). The main symptoms consisted of a weight-bearing pain in the hip and gait disturbance. The pain occurred suddenly and was of unknown cause and became severe within 2 to 3 weeks. X-ray examinations showed diffuse osteoporosis in the femoral head and neck. Moreover in 3 patients, similar lesions were also found in the lumbar spine or the knee. MRI obtained from 3 patients revealed a mottled low signal lesion extending from the femoral head and neck on T1-weighted images and a high-signal lesion in the bone marrow suggesting edema on T2-weighted images. Mild elevation of C- reactive protein was shown in 2 patients. Conservative treatments with the limitation of weight bearing and bed rest were performed for all patients, and nonsteroidal anti-inflammatory drugs were given to 3 patients. The hip pain began to decline from 8 to 14 weeks after the onset, and completely disappeared from 14 to 24 weeks. X-ray examinations showed that osteoporotic lesions tended to improve at 10 to 14 weeks, on MRI, a high-signal lesion suggesting bone marrow edema resolved together with relief of the pain. No recurrence was found in any patients at mean follow-up of 70.8 months. PMID- 11116244 TI - [A critical pathway in the operating room]. PMID- 11116245 TI - [A case of tuberous sclerosis with cardiac tumor and West syndrome]. PMID- 11116277 TI - Pharmacological treatment of the ion transport defect in cystic fibrosis. AB - Cystic fibrosis (CF) is a lethal monogenetic disease characterised by impaired water and ion transport over epithelia. The lung pathology is fatal and causes death in 95% of CF patients. The genetic basis of the disease is a mutation in the cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-regulated chloride channel. The most common mutation, DeltaF508, results in a protein that cannot properly be folded in the endoplasmic reticulum, is destroyed and hence does not reach the apical cell membrane. This paper will discuss those pharmacological approaches that are directed at correcting the defect in ion transport. At present, no clinically effective drug is available, although research has defined areas in which progress might be made. These are the following: (1) the drug 4-phenylbutyrate (4PBA) increases the expression of DeltaF508-CFTR in the cell membrane, probably by breaking the association between DeltaF508-CFTR and a chaperone; (2) a number of xanthines, in particular 8 cyclopentyl-1, 3-dipropylxanthine (CPX), are effective in activating CFTR, presumably by direct binding and also possibly by correcting the trafficking defect; (3) the isoflavone genistein can activate both wild-type and mutant CFTR, probably through direct binding to the channel; (4) purinergic agonists (ATP and UTP) can stimulate chloride secretion via a Ca(2+)-dependent chloride channel and in this way compensate for the defect in CFTR, but stable analogues will be required before this type of treatment has clinical significance; (5) treatment with inhaled amiloride may correct the excessive absorption of Na(+) ions and water by airway epithelial cells that appears connected to the defect in CFTR; although clinical tests have not been very successful so far, amiloride analogues with a longer half-life may give better results. The role of CFTR in bicarbonate secretion has not yet been established with certainty, but correction of the defect in bicarbonate secretion may be important in clinical treatment of the disease. Currently, major efforts are directed at developing a pharmacological treatment of the ion transport defect in CF, but much basic research remains to be done, in particular, with regard to the mechanism by which defective CFTR is removed in the endoplasmic reticulum by the ubiquitin-proteasome pathway, which is a central pathway in protein production and of significance for several other diseases apart from CF. PMID- 11116278 TI - Therapeutic advances in ankylosing spondylitis. AB - Ankylosing spondylitis (AS) is a systemic inflammatory rheumatic disease involving spinal and sacroiliac joints. This condition is responsible for back pain, stiffness, but also loss of functional capacity with socio-economic consequences. The management of AS includes patient education, rest, a programme of regular physical exercise, together with the use of NSAIDs. Second-line treatments are required in cases of severe or refractory AS, however only sulfasalazine has proven to benefit AS patients with peripheral arthritis. In spite of this management, the disease may not be adequately controlled, mainly for patients with refractory axial disease, enthesopathy or extra-articular features. Thus, new innovative treatments are needed for AS. It is likely that the new NSAIDs or COX-2 specific inhibitors will certainly take the place of the conventional NSAIDs, with regard to their superior tolerability. Methotrexate is a therapeutic option for AS treatment, but its usefulness in this disease remains to be established in adequate controlled studies. Finally, the TNF-alpha targeting drugs, namely thalidomide and the anti-TNF-alpha mAb, infliximab, have given promising results in the treatment of severe and/or refractory AS patients, however further controlled studies are required. In addition, the long-term use (efficacy and tolerability) of these two agents deserves attention. PMID- 11116279 TI - Novel therapy in the treatment of scleroderma. AB - While the biology of the pathogenesis of scleroderma is continually being better understood, there still is no single agent or therapeutic combination that has a clear impact on the disease process. Traditional medications (colchicine, potassium aminobenzoate (potaba), D-penicillamine) are disappointing in clinical practice despite anecdotal evidence of benefit. Furthermore, the most popular traditional drug, D-penicillamine, failed to clearly show benefit when tested in a well-designed clinical trial comparing conventional high dose with a very low dose (125 mg po. every other day [corrected]) [1]. Currently, most success in managing scleroderma and improving quality of life is secondary to organ-specific therapy, such as management of a renal crisis with an ACE inhibitor, treatment of Raynaud's phenomenon with calcium channel blockers, or control of serious gastrointestinal reflux disease with a proton pump inhibitor. In this review we will focus on novel therapies that are currently being tested in the treatment of scleroderma and have the potential of modifying the disease process and overall clinical outcome. We have attempted to review the rationale for each agent, recognising that its true biological effect will only be determined in clinical trials. PMID- 11116280 TI - Immunotherapy in atopic dermatitis. AB - Atopic dermatitis (AD) is a common inflammatory disease involving the skin and often other organs and systems, mainly respiratory. A definitive general consensus on the AD pathogenesis has not yet been established, however several lines of evidence suggest that T-cells play a crucial role in priming AD early stage lesions. Main topics involved in the disease pathogenesis have been reviewed, which considered the concept of local and systemic haemopoietic events as important contributors to allergic inflammation, a concept now achieving great acceptance. The recently recognised atopic nature of the skin inflammation in AD has raised increasing interest for treatment with allergen-specific immunotherapy. However, we only found eight studies using specific immunotherapy (SIT) in AD, two double-blind, placebo-controlled (DBPC) and six observational. One controlled and five observational reported favourable outcomes. The one unique study providing negative results was flawed by the ineffective oral route of extract administration. Despite being encouraging, the reported results do not allow definitive conclusions based on meta-analytic techniques because the amount and quality of information in the literature is not sufficient. The highly promising sub-lingual immunotherapy (SLIT) is discussed with its potential capability of controlling not only the skin lesion severity but also its capability of preventing the development of atopic dermatitis into asthma. PMID- 11116281 TI - Pharmacological agents for the treatment of urinary incontinence due to overactive bladder. AB - Although the exact aetiology of overactive bladder is unknown to date, pharmacological therapy has been targeted to both the central and peripheral nervous systems. Potential CNS targets include GABA, opioid, serotonin (5-HT), dopamine and glutaminergic receptors as well as the alpha-adrenoceptors. Potential PNS targets include muscarinic receptors, calcium and potassium channels and alpha- and beta-adrenergic receptors. Since acetylcholine is the primary excitatory neurotransmitter involved in bladder (detrusor) contraction and emptying, anticholinergic agents are the primary compounds used clinically to decrease involuntary detrusor contractions. Anticholinergic therapy has a stabilising effect on the bladder (detrusor muscle); increases bladder capacity; decreases frequency of involuntary detrusor contractions; and delays the initial urge to void, but does not affect warning time. However, the clinical utility of antimuscarinic therapy is limited by the lack of receptor selectivity, resulting in the classic anticholinergic side effects of dry mouth, blurred vision, constipation and potentially, CNS effects such as somnolence and impaired cognitive function. These unwanted side effects often result in premature discontinuation of therapy and poor compliance. Previous attempts to develop uroselective alpha-adrenergic receptor antagonists have not been successful and although research continues, the hope that this class of agents would be viable alternatives to the anticholinergics remains to be proven in the clinical setting. The recent demise of several potassium channel openers does not augur well for the future of this class of agent. The reasons for the discontinuation of trials with these agents have not been fully elucidated, but one must assume that they were not uroselective and the cardiovascular side effects rendered them less than useful clinically. The serotonin re-uptake inhibitors appear to be promising novel therapeutic agents aimed at controlling bladder over-activity through specific CNS pathways. The sensory side of the micturition reflex is a potential therapeutic target. Agents to desensitise afferent nerve endings involved in C-fibre afferent reflexes include capsaicin and resiniferatoxin. Their clinical applicability is currently being evaluated. Finally, the recent findings related to the role of the P2X3 receptor in the sensory aspects of bladder filling have created new interest in the future development of agents that will improve the management of this prevalent and debilitating condition. PMID- 11116282 TI - Novel approaches to female sexual dysfunction. AB - Female sexual dysfunction is age-related, progressive and highly prevalent, affecting 30 - 50% of American women. While there are emotional and relational elements to sexual function, it has become increasingly evident that female sexual dysfunction can occur secondary to medical problems and has an organic basis. A plethora of different female sexual dysfunctions exist and in order to obtain a greater understanding of the possible treatments for these problems, it is essential to have a strong knowledge base of female pelvic anatomy, the neurogenic mediators of female sexual response, the impact of hormones on female sexual function and the aetiologies of female sexual dysfunction. Currently, there are potential therapeutic options for the treatment of female sexual dysfunction and these options include both hormonal and pharmacological therapy. However, therapeutic agents may not prove to be enough and the ideal approach to female sexual dysfunction is thus a collaborative effort between therapists and physicians, which should include a complete medical and psychosocial evaluation, as well as inclusion of the partner or spouse in the evaluation and treatment process. PMID- 11116283 TI - Pharmacology and clinical experience with fedotozine. AB - Fedotozine [(1R)-1-phenyl-1-[(3,4,5-trimethoxy)benzyloxymethyl]-N,N- dimethyl-n propylamine, (2S,3S-tartrate] is derived from the arylacetamide series. As with other compounds of this series, fedotozine is more or less selective of kappa(1) opioid receptors and particularly for the kappa(1a)-receptor subtype, where it acts as an agonist. Pharmacological studies have shown that fedotozine exerts a peripheral antinociceptive action, comparable with that of other kappa-agonists. Its main effects have been demonstrated at the level of the afferent nerve pathways originating from the gut. Fedotozine alters the processing of visceral sensations along these pathways and hence, the perception of gut stimuli at the brain level. It modifies reflexes induced in various pathological conditions, like experimental inflammation of the gut, chemically-induced peritonitis or post operative ileus. Fedotozine also decreases the nociceptive reflexes triggered by noxious gut distension in animals. In humans, fedotozine decreases the perception of gut distension, both in physiological and pathological conditions. Clinical trials undertaken in patients with functional digestive disorders, non-ulcer dyspepsia and irritable bowel syndrome, have shown that fedotozine relieves abdominal pain in these patients in 6-week treatments. kappa-Opioid receptors remain an interesting area for future development of new treatments for abdominal pain in patients with functional digestive disorders. PMID- 11116284 TI - Clinical and pharmacological experience with LJP-394. AB - LJP-394 is a synthetic biological with immunomodulatory functions. Composed of four double-stranded oligodeoxynucleotides attached to a central branched platform, the drug acts as an anti-"anti-ds-DNA" B-cell toleragen by rendering specific B-lymphocytes unresponsive to immunogen so they do not produce autoantibodies. Extensive animal studies and Phase II clinical trials suggested that the effects of LJP-394 are effective and safe when used as a weekly dose of 100 mg intravenously. Analysis of a multicentre, international Phase II/III clinical trial showed that patients with lupus nephritis and high affinity IgG antibodies to LJP-394 have clinical benefits. This includes increased time to renal flares, reduced number of renal flares, time to institution of high-dose corticosteroids and/or cyclophosphamide and lower anti-ds-DNA levels. A definitive trial is in progress. LJP-394 appears to be free of serious adverse reactions. Though promising, the role of LJP-394 in patients with active, organ threatening lupus is still not known. PMID- 11116285 TI - Lobaplatin: a new antitumour platinum drug. AB - Lobaplatin (D-19466) is a diastereometric mixture of platinum(II) complexes containing a 1,2-bis(aminomethyl)cyclobutane stable ligand and lactic acid as the leaving group. Its antitumour activity results from the formation of DNA-drug adducts, mainly as GG and AG intra-strand cross-links. Lobaplatin influences the expression of the c-myc gene, which is involved in oncogenesis, apoptosis and cell proliferation. Lobaplatin has activity in a wide range of preclinical tumour models and appears to overcome tumour resistance to cisplatin and carboplatin in some of these models. In the body, lobaplatin remains largely intact until removed by glomerular filtration. Exposure of the body to lobaplatin (AUC) correlates with dose, creatinine clearance and the degree of thrombocytopoenia. Phase I clinical trials of three quite different administration schedules found the same dose-limiting toxicity (thrombocytopoenia) and similar maximum tolerated doses (60 mg/m(2) per 3 - 4 weeks). In Phase II trials, lobaplatin showed activity in patients with a variety of tumour types. Many of the patients who responded to lobaplatin may also have responded to cisplatin and carboplatin because they had had no prior chemotherapy or had a prolonged remission after earlier treatment. In conclusion, lobaplatin is a new platinum drug, which overcomes some forms of cisplatin resistance in preclinical tumour models. Several potential clinical applications remain unexplored, such as its use in relapsed testicular cancer and in combination with other cancer chemotherapeutic agents and ionising radiation. PMID- 11116286 TI - Tedisamil: master switch of nature? AB - Decreasing heart rate is potentially useful in ischaemic heart disease. Tedisamil is a bradycardic agent resulting from its ability to inhibit transient outward current (I(to)) in atria. Tedisamil inhibits I(to), potassium current (IK), K(ATP) and the protein kinase A-activated chloride channel in ventricles as well as vascular IK and Ca(2+)-activated IK (IK((Ca))). Tedisamil prolongs cardiac action potentials and the corrected QT (QTc) of the ECG and also increases cardiac refractoriness. Tedisamil is anti-arrhythmic in animal models of ventricular arrhythmias and atrial flutter. The bradycardic effect of tedisamil is associated with a reduction in myocardial oxygen demand. On isolated rat ventricle, tedisamil is a positive inotrope and on isolated rabbit atria, tedisamil reverses the negative inotropic effect of pinacidil. Tedisamil contracts the isolated rat portal vein and aorta, reduces cromakalim-induced relaxations of contracted rat aorta and increases blood pressure in animals and humans. Tedisamil is 96% bound to plasma proteins, has a plasma half-life of about 10 h and is cleared from the kidney unchanged. Clinical trials have shown that the electrophysiology of tedisamil is that of a class III anti-arrhythmic. In coronary artery disease, tedisamil has no effect on inotropism and increases the threshold for angina. Potassium channel blockade with tedisamil may have advantages over calcium channel blockers or K(ATP) channel openers as an anti ischaemic mechanism in coronary artery disease. In exercise-induced myocardial ischaemia, beta-blockers are probably favourable to tedisamil, as they will limit the increase in heart rate, contractility and blood pressure caused by sympathetic stimulation, whereas tedisamil will not. In heart failure patients, tedisamil reduces heart rate, but increases blood pressure. The usefulness of tedisamil as a bradycardic agent is limited by the increase in blood pressure. A drug that is bradycardic without increasing blood pressure would be an improvement on tedisamil as the master switch of nature for ischaemic heart disease. PMID- 11116287 TI - Vatanidipine hydrochloride: a new long-lasting antihypertensive agent. AB - Vatanidipine is a novel dihydropyridine (DHP)-type calcium channel blocker with slow-onset pharmacological actions, which are probably due to both its slow uptake into vascular tissues and resistance in its approach to the calcium channel binding site. Vatanidipine once incorporated into vascular tissues is not easily released, even by repeated washing, thus resulting in a long-lasting action of the agent. A slow-onset and long-lasting hypotensive action was observed in various experimental hypertensive models. Clinical trials using human subjects with essential hypertension indicated that vatanidipine exerts an antihypertensive effect with a slow onset and long duration. In spite of its potent hypotensive effect, the incidence of adverse effects by vatanidipine administration has been reported to be lower than that in cases of nitrendipine. In addition to its vasodilatory effects, vatanidipine efficiently suppressed noradrenaline release from sympathetic nerve endings, thus suggesting this agent exhibits a beneficial effect in the treatment of hypertensive patients, in which the reflex activation of peripheral sympathetic nerves is unfavourable to antihypertensive therapy. In a double-blind study, vatanidipine did not show reflex tachycardia, despite producing a potent and long-lasting hypotensive effect, in contrast to the administration of nitrendipine. In an animal study, vatanidipine exhibited a protective effect against cerebrovascular lesions, through a mechanism independent of its hypotensive effect. In addition, a renoprotective effect was also observed in experimental hypertensive models. In cholesterol-fed rabbits, vatanidipine exerted an anti-atherosclerotic action, which is probably attributable to the inhibitory action of the agent on low density lipoprotein oxidation. In essential hypertensive patients, the plasma levels of cholesterol and triglyceride decreased after vatanidipine treatment, thus suggesting that this agent may have a therapeutic potential in preventing such vascular diseases as atherosclerosis. Taken together, vatanidipine appears to be a novel and useful antihypertensive agent, which can both prevent target organ damage and reduce cardiovascular morbidity and mortality. PMID- 11116301 TI - Nondestructive Angle-resolved X-ray Depth Profiling: Interpretation of Angle resolved Profiles Using a Monte Carlo Approach. AB - A technique has been developed for the interpretation of composition-depth profiles from angle-resolved X-ray data using a Monte Carlo electron scattering simulation. This is a nondestructive depth profiling procedure. Software has been developed that uses a Monte Carlo scattering simulation to generate the signal intensity from a multilayer sample for any combination of primary beam angle of incidence and take-off angle to the X-ray detector. An interactive C++ application uses this simulation to interpret measured angle-resolved depth profiles. The method has been tested using a custom-made Ag/Al "staircase" sample containing two layers each of Ag and Al. Using the technique, it is possible to quantify the composition-depth profile for the two- and three-layer "steps" of the sample. Qualitative information may be gained about the four-layer area of the sample. PMID- 11116302 TI - Nonlinear Dynamics in the Progression of Atherosclerotic Fatty Streaks: Morphometric Analysis. AB - Atherosclerotic lesions are heterogeneous in terms of their cellular and lipid composition. While heterogeneity can be the result of stochastic noise, an alternate hypothesis is that the differences observed among individual lesions arise from deterministic chaos. Five New Zealand white rabbits were fed a diet containing 0.15% cholesterol for 6 months. Segments of the aorta were fixed in formalin, stained en bloc with Nile red (NR) and filipin (F), and en face fluorescence microscopy was used to map the distribution of lipids in fatty streaks (FS). The smallest lesions detected stained only with filipin. Larger lesions stained with both Nile red and filipin and two distinct regions of Nile red staining, NR-orange (rich in polar lipids) and NR-yellow (rich in neutral lipids) were observed. Digital overlays revealed a "nested" arrangement of F, NR orange, and NR-yellow. The lesions also showed marked heterogeneity in their lipid composition. Thus, although initially similar, as FS increased in size, their composition became divergent, suggesting that the ultimate composition of a FS was highly sensitive to its initial composition. Sensitivity to initial condition is one of the hallmarks of deterministic systems. To determine if FS were self-similar, another hallmark of deterministic chaos, the borders of the different regions defined by NR and F staining were subjected to fractal analysis. For each lesion, the borders of the F, NR-orange, and NR-yellow regions were found to be fractal. Return maps were constructed for the differently stained regions. Analysis of the entire 104-lesion data set showed that although the data could be described by a four-parameter logistic model, the population was not chaotic. However, return maps drawn for the maxima of the NR-orange stained regions demonstrated chaos. Taken together, the data suggest that deterministic chaos plays a role in the evolution of atheromatous disease but, in common with most biologic systems, as the lesions progress, chaotic behavior is dampened. PMID- 11116303 TI - Preparation and Certification of K-411 Glass Microspheres. AB - The production and characterization of NBS K-411 glass microspheres in the 2-40 um range for certification as NIST Standard Reference Material(R) 2066 (SRM(R)) are described. Quantitative analysis and heterogeneity testing of the microspheres were done with an electron probe microanalyzer-X-ray energy dispersive spectrometry (EPMA-EDS) automated particle analysis procedure. Results for the trimmed and normalized data produced mean compositions for the elements Mg, Si, Ca, Fe, and O (calculated from stoichiometry) that are in good agreement with the certified values for the K-411 bulk glass (NBS SRM 470 Glasses for Mineral Analysis), but with uncertainties about twice as large as those for the bulk material. Differences from the bulk are attributable to microsphere geometry as well as mass and size effects. PMID- 11116304 TI - Fluorescence Microscopy Study of Heterogeneity in Polymer-supported Luminescence based Oxygen Sensors. AB - Despite the great potential of fluorescence microscopy, its application to date has largely been in the study of biological specimens. It will be shown that conventional fluorescence microscopy provides an invaluable tool with which to study the photophysics of polymer-supported luminescence-based oxygen sensors. The design of the imaging system, the measurement methods, and the data analysis used in the investigation of sensor systems are described. Fluorescence microscopic images of sensor films in which microheterogeneous regions exhibiting enhanced luminescence intensity and poorer oxygen quenching relative to the bulk response are shown. This is the first direct evidence that sensor molecules in various domains of the polymer support can exhibit different oxygen quenching properties. It will be shown that u- and nano-crystallization of the sensor molecule are the probable source of both the observed heterogeneous microscopic responses and the microscopic and macroscopic nonlinear Stern-Volmer plots. The implications of these results in the rational design of luminescence-based oxygen sensors are discussed. PMID- 11116305 TI - News and Commentary. PMID- 11116306 TI - Survival of genetically modified Escherichia coli carrying extraneous antibiotic resistance gene through microbial interactions. PMID- 11116307 TI - Occupational exposure during removal of windows with lead-based paint and asbestos caulking. PMID- 11116308 TI - Organochlorine pesticides and metals in select botanical dietary supplements. PMID- 11116309 TI - Monitoring groundwater at landfills equipped with leachate collection systems. PMID- 11116310 TI - Organochlorine pesticide residues in Helmeted Guineafowl (Numida meleagris), South Africa. PMID- 11116311 TI - Organochlorine pesticide residue in soils from Tibet, China. PMID- 11116312 TI - Gas chromatography-chemical ionization mass spectrometry for drug screen analyses. PMID- 11116313 TI - Metal pollution and fat accumulation in the carabid beetle Pterostichus melanarius (Coleoptera, Carabidae). PMID- 11116314 TI - Chromium in a tannery wastewater irrigated area, Leon Valley, Mexico. PMID- 11116315 TI - Airborne toxic metals in air of Mumbai City, India. PMID- 11116316 TI - Heavy metals in the false mussel, Mytilopsis domingensis, from two tropical estuarine lagoons. PMID- 11116317 TI - Bioaccumulation of aluminium in Dunaliella tertiolecta in natural seawater: aluminium-metal (Cu, Pb, Se) interactions and influence of pH. PMID- 11116318 TI - Distribution of persistent lipophilic contaminants in fish from the Grand Duchy of Luxembourg. PMID- 11116319 TI - Ecological evaluation of gadolinium toxicity compared with other heavy metals using an aquatic microcosm. PMID- 11116320 TI - Bradybaena similaris (Ferrusac) shell as a biomonitor of copper, cadmium, and zinc. PMID- 11116321 TI - Effect of heavy metals on growth and extracellular enzyme activities of mycoparasitic Trichoderma strains. PMID- 11116322 TI - Acute toxicity of four drinking water disinfection by-products to Japanese medaka fish. PMID- 11116323 TI - Algal growth recovery studies after paraquat exposure. PMID- 11116324 TI - Deltamethrin, effects on volume control, and water balance in Eisenia fetida Sav. (Annelida, Lumbricidae). PMID- 11116325 TI - Social constructionism, a lost cause. PMID- 11116326 TI - Structural evidence for the evolution of pyrogenic toxin superantigens. AB - Pathogenic bacteria have evolved a wide variety of toxins to invade and attack host organisms. In particular, strains of the bacteria Staphylococcus aureus and Streptococcus pyogenes produce a family of pyrogenic toxin superantigens (PTSAgs) that can cause illness, e.g., toxic shock syndrome, or synergize with a number of other immune system disorders. The PTSAgs are all similar in size and have a conserved two-domain tertiary fold despite minimal amino acid sequence identity. The tertiary structure of PTSAg domain 1 is similar to the immunoglobulin binding motif of streptococcal proteins G and L. PTSAg domain 2 resembles members of the oligosaccharide/oligonucleotide binding fold family that includes the B subunits of the AB(5) heat-labile enterotoxins, cholera toxin, pertussis toxin, and verotoxin. The strong structural homology between the pyrogenic toxins and other bacterial proteins suggests that the PTSAgs evolved through the recombination of two smaller beta-strand motifs. PMID- 11116327 TI - Phylogenomic analysis of the alpha proteasome gene family from early-diverging eukaryotes. AB - We employed a phylogenomic approach to study the evolution of alpha subunits of the proteasome gene family from early diverging eukaryotes. BLAST similarity searches of the Giardia lamblia genome identified all seven alpha proteasome genes characteristic of eukaryotes from the crown group. In addition, a PCR strategy for the amplification of multiple alpha subunit sequences generated single alpha proteasome products for representatives of the Kinetoplastida (Leishmania major), the Parabasalia (Trichomonas vaginalis), and the Microsporidia (Vairimorpha sp., Nosema sp., Endoreticulata sp., and Spraguea lophii). The kinetoplastid Trypanosoma cruzi and the eukaryote crown group Acanthamoeba castellanii yielded two distinct alpha proteasome genes each. The presence of seven distinct alpha proteasome genes in G. lamblia, one of the earliest-diverging eukaryotes, indicates that the alpha proteasome gene family evolved rapidly from a minimum of one gene in Archaea to seven or more in Eukarya. Results from the phylogenomic analysis are consistent with the idea that the Diplomonida (as represented by G. lamblia), the Kinetoplastida, the Parabasalia, and the Microsporidia diverged after the duplication events that originated the alpha proteasome gene family. A model for the early origin and evolution of the proteasome gene family is presented. PMID- 11116328 TI - Efficiency of the neighbor-joining method in reconstructing deep and shallow evolutionary relationships in large phylogenies. AB - The neighbor-joining (NJ) method is widely used in reconstructing large phylogenies because of its computational speed and the high accuracy in phylogenetic inference as revealed in computer simulation studies. However, most computer simulation studies have quantified the overall performance of the NJ method in terms of the percentage of branches inferred correctly or the percentage of replications in which the correct tree is recovered. We have examined other aspects of its performance, such as the relative efficiency in correctly reconstructing shallow (close to the external branches of the tree) and deep branches in large phylogenies; the contribution of zero-length branches to topological errors in the inferred trees; and the influence of increasing the tree size (number of sequences), evolutionary rate, and sequence length on the efficiency of the NJ method. Results show that the correct reconstruction of deep branches is no more difficult than that of shallower branches. The presence of zero-length branches in realized trees contributes significantly to the overall error observed in the NJ tree, especially in large phylogenies or slowly evolving genes. Furthermore, the tree size does not influence the efficiency of NJ in reconstructing shallow and deep branches in our simulation study, in which the evolutionary process is assumed to be homogeneous in all lineages. PMID- 11116329 TI - Extensive amplification and transposition of a novel repetitive element, xstir, together with its terminal inverted repeat in the evolution of Xenopus. AB - A DNA fragment containing short tandem repeat sequences (approximately 86-bp repeat) was isolated from a Xenopus laevis cDNA library. Southern blot and in situ hybridization analyses revealed that the repeat was highly dispersed in the genome and was present at approximately 1 million copies per haploid genome. We named this element Xstir (Xenopus short tandemly and invertedly repeating element) after its arrangement in the genome. The majority of the genomic Xstir sequences were digested to monomer and dimer sizes with several restriction enzymes. Their sequences were found to be highly homogeneous and organized into tandem arrays in the genome. Alignment analyses of several known sequences showed that some of the Xstir-like sequences were also organized into interspersed inverted repeats. The inverted repeats consisted of an inverted pair of two differently modified Xstirs separated by a short insert. In addition, these were framed by another novel inverted repeat (Xstir-TIR). The Xstir-TIR sequence was also found at the ends of tandem Xstir arrays. Furthermore, we found that Xstir TIR was linked to a motif characterizing the T2 family which belonged to a vertebrate MITE (miniature inverted-repeat transposable element) family, suggesting the importance of Xstir-TIR for their amplification and transposition. The present study of 11 anuran and 2 urodele species revealed that Xstir or Xstir like sequences were extensively amplified in the three Xenopus species. Genomic Xstir populations of X. borealis and X. laevis were mutually indistinguishable but significantly different from that of X. tropicalis. PMID- 11116330 TI - An updated and comprehensive rRNA phylogeny of (crown) eukaryotes based on rate calibrated evolutionary distances. AB - Recent experience with molecular phylogeny has shown that all molecular markers have strengths and weaknesses. Nonetheless, despite several notable discrepancies with phylogenies obtained from protein data, the merits of the small subunit ribosomal RNA (SSU rRNA) as a molecular phylogenetic marker remain indisputable. Over the last 10 to 15 years a massive SSU rRNA database has been gathered, including more then 3000 complete sequences from eukaryotes. This creates a huge computational challenge, which is exacerbated by phenomena such as extensive rate variation among sites in the molecule. A few years ago, a fast phylogenetic method was developed that takes into account among-site rate variation in the estimation of evolutionary distances. This "substitution rate calibration" (SRC) method not only corrects for a major source of artifacts in phylogeny reconstruction but, because it is based on a distance approach, allows comprehensive trees including thousands of sequences to be constructed in a reasonable amount of time. In this study, a nucleotide variability map and a phylogenetic tree were constructed, using the SRC method, based on all available (January 2000) complete SSU rRNA sequences (2551) for species belonging to the so called eukaryotic crown. The resulting phylogeny constitutes the most complete description of overall eukaryote diversity and relationships to date. Furthermore, branch lengths estimated with the SRC method better reflect the huge differences in evolutionary rates among and within eukaryotic lineages. The ribosomal RNA tree is compared with a recent protein phylogeny obtained from concatenated actin, alpha-tubulin, beta-tubulin, and elongation factor 1-alpha amino acid sequences. A consensus phylogeny of the eukaryotic crown based on currently available molecular data is discussed, as well as specific problems encountered in analyzing sequences when large differences in substitution rate are present, either between different sequences (rate variation among lineages) or between different positions within the same sequence (among-site rate variation). PMID- 11116331 TI - Ancient and recent horizontal invasions of drosophilids by P elements. AB - P elements of two different subfamilies designated as M- and O-type are thought to have invaded host species in the Drosophila obscura group via horizontal transmission from external sources. Sequence comparisons with P elements isolated from other species suggested that the horizontal invasion by the O-type must have been a rather recent event, whereas the M-type invasion should have occurred in the more distant past. To trace the phylogenetic history of O-type elements, additional taxa were screened for the presence of O- and M-type elements using type-specific PCR primers. The phylogeny deduced from the sequence data of a 927 bp section (14 taxa) indicate that O-type elements have undergone longer periods of regular vertical transmission in the lineages of the saltans and willistoni groups of Drosophila. However, starting from a species of the D. willistoni group they were transmitted horizontally into other lineages. First the lineage of the D. affinis subgroup was infected, and finally, in a more recent wave of horizontal spread, species of three different genera were invaded by O-type elements from the D. affinis lineage: Scaptomyza, Lordiphosa, and the sibling species D. bifasciata/D. imaii of the Drosophila obscura subgroup. The O-type elements isolated from these taxa are almost identical (sequence divergence <1%). In contrast, no such striking similarities are observed among M-type elements. Nevertheless, the sequence phylogeny of M-type elements is also not in accordance with the phylogeny of their host species, suggesting earlier horizontal transfer events. The results imply that P elements cross species barriers more frequently than previously thought but require a particular genomic environment and thus seem to be confined to a rather narrow spectrum of host species. Consequently, different P element types acquired by successive horizontal transmission events often coexist within the same genome. PMID- 11116332 TI - Horizontal transfer of archaeal genes into the deinococcaceae: detection by molecular and computer-based approaches. AB - Members of the Deinococcaceae (e.g., Thermus, Meiothermus, Deinococcus) contain A/V-ATPases typically found in Archaea or Eukaryotes which were probably acquired by horizontal gene transfer. Two methods were used to quantify the extent to which archaeal or eukaryotic genes have been acquired by this lineage. Screening of a Meiothermus ruber library with probes made against Thermoplasma acidophilum DNA yielded a number of clones which hybridized more strongly than background. One of these contained the prolyl tRNA synthetase (RS) gene. Phylogenetic analysis shows the M. ruber and D. radiodurans prolyl RS to be more closely related to archaeal and eukaryal forms of this gene than to the typical bacterial type. Using a bioinformatics approach, putative open reading frames (ORFs) from the prerelease version of the D. radiodurans genome were screened for genes more closely related to archaeal or eukaryotic genes. Putative ORFs were searched against representative genomes from each of the three domains using automated BLAST. ORFs showing the highest matches against archaeal and eukaryotic genes were collected and ranked. Among the top-ranked hits were the A/V-ATPase catalytic and noncatalytic subunits and the prolyl RS genes. Using phylogenetic methods, ORFs were analyzed and trees assessed for evidence of horizontal gene transfer. Of the 45 genes examined, 20 showed topologies in which D. radiodurans homologues clearly group with eukaryotic or archaeal homologues, and 17 additional trees were found to show probable evidence of horizontal gene transfer. Compared to the total number of ORFs in the genome, those that can be identified as having been acquired from Archaea or Eukaryotes are relatively few (approximately 1%), suggesting that interdomain transfer is rare. PMID- 11116333 TI - Coding in the noncoding DNA strand: A novel mechanism of gene evolution? AB - The question whether the noncoding DNA strand had or still has the capability for encoding functional polypeptides has been addressed in several articles. The theoretical background of the views advocating this idea arose from two groups of findings. One of them was based on various observations implying that the genetic code was adapted for double-strand coding. The other group of theories arose from the observation of gene-length overlapping open reading frames (O-ORFs) on the antisense DNA strand in a number of genes. In fact, the above theories, which I term selectionist, conceive a novel conception of gene evolution, proposing that new genes can be created by the utilization of antisense DNA strand. In contrast, neutralist theory claims that the O-ORFs are mere by-products of evolutionary processes acting to create special codon usage and base distribution patterns in the coding sequences. PMID- 11116334 TI - The quality of merC, a module of the mer mosaic. AB - We examined a region of high variability in the mosaic mercury resistance (mer) operon of natural bacterial isolates from the primate intestinal microbiota. The region between the merP and merA genes of nine mer loci was sequenced and either the merC, the merF, or no gene was present. Two novel merC genes were identified. Overall nucleotide diversity, pi (per 100 sites), of the merC gene was greater (49.63) than adjacent merP (35.82) and merA (32.58) genes. However, the consequences of this variability for the predicted structure of the MerC protein are limited and putative functional elements (metal-binding ligands and transmembrane domains) are strongly conserved. Comparison of codon usage of the merTP, merC, and merA genes suggests that several merC genes are not coeval with their flanking sequences. Although evidence of homologous recombination within the very variable merC genes is not apparent, the flanking regions have higher homologies than merC, and recombination appears to be driving their overall sequence identities higher. The synonymous codon usage bias (EN(C)) values suggest greater variability in expression of the merC gene than in flanking genes in six different bacterial hosts. We propose a model for the evolution of MerC as a host-dependent, adventitious module of the mer operon. PMID- 11116335 TI - Influence of mining-related activities on concentrations of metals in water and sediment from streams of the Black Hills, South Dakota. AB - Water and sediment samples were collected from streams in Spearfish Creek, Whitewood Creek, and Bear Butte Creek watersheds in the Black Hills, SD, an area impacted by gold mining operations. Arsenic concentrations that exceeded the U.S. Environmental Protection Agency's Maximum Concentration Limit of 50 microg/L for drinking water were found in water from Annie Creek, a tributary of Spearfish Creek, and from Whitewood Creek. Gold Run, a tributary of Whitewood Creek, and Annie Creek contained Se concentrations in water that exceeded the EPA Ecotox threshold of 5 microg/L and were classified as a high hazard for Se accumulation from water into the planktonic food chain and for resultant toxicity to fish and aquatic birds. Concentrations of As, Cd, Cu, Hg, Ni, Pb, and Zn in sediment exceeded EPA Ecotox thresholds in one or more of the watersheds suggesting potential adverse ecological effects. Sediment from Rubicon Creek, a tributary of Spearfish Creek, contained Se concentrations high enough (4.0 microg/g) to be a moderate hazard for accumulation from sediments into the benthic food chain, with resultant dietary toxicity to fish and aquatic birds. These results are discussed in light of historical mining activities and recent clean-up and reclamation efforts. Based on the results and comparisons to Ecotox tresholds, further studies of ecological effects are warranted. PMID- 11116336 TI - The effect of mercury and PCBs on organisms from lower trophic levels of a Georgia salt marsh. AB - We examined several indicators of salt marsh function, focusing on primary producers, microbes, and grass shrimp, at a Superfund site (LCP) contaminated with mercury and polychlorinated biphenyls (PCBs) and a reference site (Cross River) in Georgia. Primary production of Spartina alterniflora was assessed by measuring peroxidase activity (POD), glutathione concentration (tGSH), photosynthesis (A(net)), and transpiration (E). Microbial populations were assessed by measuring living-fungal standing crop (as ergosterol) and Microtox(R). Grass shrimp (Palaemonetes pugio) reproductive potential was determined by measuring individual egg mass, average egg area, brood size, and brood mass of gravid females. Comparison of the sites suggested that P. pugio reproduction was affected at the LCP site, but we were unable to document clear negative effects on other organisms we investigated. Due to natural environmental gradients, the Cross-River site may not have been a perfect control for the LCP site. Therefore, data from just the LCP site were reanalyzed using multiple regression. Fungal biomass was related to methylmercury concentrations, but the direction of the relationship differed between wholly dead shoots (positive) and partially dead shoots (negative). S. alterniflora POD was positively related to methylmercury concentrations. S. alterniflora A(net) and E were negatively related to elevation and salinity, respectively. Despite high levels of contamination at the LCP site, our results provided only suggestive evidence for impacts on organisms at lower trophic levels. PMID- 11116337 TI - Antioxidant modulation in response to metal-induced oxidative stress in algal chloroplasts. AB - To investigate adaptive responses to metal stress at the subcellular level, the oxidative balance in isolated chloroplasts was evaluated for the first time in the unicellular alga Gonyaulax polyedra exposed to the toxic metals Hg(2+), Cd(2+), Pb(2+), and Cu(2+). Different antioxidant responses were verified according to the metal and model of stress applied. Cells chronically exposed to metals exhibited high activity of the antioxidant enzymes superoxide dismutase and ascorbate peroxidase, high glutathione content, and decrease of peridinin levels, whereas no significant changes were detected for beta-carotene levels. In contrast, cells subjected to acute metal stress displayed twice as much beta carotene but only a slight increase in superoxide dismutase and ascorbate peroxidase activities. The correlation of acute metal treatment and oxidative stress was inferred from the higher oxygen uptake and decreased reduced glutathione pool found in treated cells. In addition, increased oxidative damage to proteins and lipids occurred mainly in cells under acute stress. Pb(2+) was the most damaging toxicant, causing protein oxidation and lipid peroxidation even at chronic treatment. These results indicate that heavy metals are able to induce oxidative stress in chloroplasts of G. polyedra, particularly under acute conditions. Nevertheless, the maintenance of a high antioxidant capacity within chloroplasts seems to be an important strategy during acclimation of G. polyedra to chronic metal stress. By acting at the subcellular site, where oxidative stress is triggered, induction of such chloroplast antioxidants might be crucial for cell survival during exposure to heavy metals. PMID- 11116338 TI - The relevance of rooted vascular plants as indicators of estuarine sediment quality. AB - Toxicity assessments and numerical quality assessment guidelines for estuarine sediments are rarely based on information for aquatic plants. The effect of this lack of information on contaminated sediment toxicity evaluations is largely unknown. For this reason, the toxicities of whole sediments collected from 15 sites in three urbanized Florida bayou-estuaries were determined for the benthic invertebrates Mysidopsis bahia and Ampelisca abdita and the plants Scirpus robustus Pursh (saltmarsh bulrush) and Spartina alterniflora Loisel (saltmarsh cordgrass). The results of the bioassays, conducted for 7 to 28 days, were compared for interspecific differences and to effects-based, sediment quality assessment guidelines. A variety of inorganic and organic analytes were detected in the estuarine sediments, and concentrations of as many as 7 analytes exceeded the sediment guidelines at the 15 sampling locations. Toxicity occurred at 2 of the 15 sampling stations based on invertebrate survival. Twelve of the 15 sediments had either a significant stimulatory or inhibitory effect on early seedling growth relative to a reference sediment (p < 0.05). The phytoresponse was specific to the location, test species, and plant tissue. There was no consistent trend between the sensitivities of the plants and invertebrates exposed to the sediments collected from the same sites. Of the 12 sediments that significantly affected seedling growth, 10 were not acutely toxic to the invertebrates. Consequently, the plant test species provided information that would have been missing if only animal test species were used. For this reason, the phytotoxicity database needs to be expanded for contaminated sediments to further evaluate interspecific sensitivities and to provide perspective on the environmental relevancy of proposed sediment quality criteria and effects-based assessment guidelines for which this information is usually missing. However, additional test method development and field validation are needed to support this effort, which includes the identification of sensitive plant test species, response parameters, and the chemical and physical sediment factors that influence plant growth. PMID- 11116339 TI - Accumulative characteristics of pesticide residues in organs of bivalves (Anodonta woodiana and Corbicula leana) under natural conditions. AB - Accumulative characteristics of pesticide residues in the gill, midgut gland, gonad, and the remaining tissues of the bivalve mollusks Anodonta woodiana and Corbicula leana were examined during the rice planting seasons of 1992 and 1994. Although seven pesticides, except thiobencarb, were accumulated all at ppb levels in the midgut gland (liver) and gonad of both bivalves during their application period, thiobencarb was accumulated in C. leana at extremely high levels of 15.70 microg g(-1) in 1992 and 12.45 microg g(-1) in 1994 in the midgut gland and 15.80 microg g(-1) in 1992 and 16.40 microg g(-1) in 1994 in the gonad, respectively. These levels were about 100 times higher than those in A. woodiana. Thiobencarb and molinate in A. woodiana and chlornitrofen (CNP) and molinate in C. leana remained in the gonad and midgut gland longer than in the gill and remaining tissues, while thiobencarb in organs of C. leana remained at ppm levels until the end of the experiments. The present study suggests that these interspecies differences can be attributed to differences between the two species in their ability to eliminate pesticides. PMID- 11116340 TI - Bioavailability of metals in stream food webs and hazards to brook trout (Salvelinus fontinalis) in the upper Animas River watershed, Colorado. AB - The water quality, habitats, and biota of streams in the upper Animas River watershed of Colorado, USA, are affected by metal contamination associated with acid drainage. We determined metal concentrations in components of the food web of the Animas River and its tributaries-periphyton (aufwuchs), benthic invertebrates, and livers of brook trout (Salvelinus fontinalis)-and evaluated pathways of metal exposure and hazards of metal toxicity to stream biota. Concentrations of the toxic metals cadmium (Cd), copper (Cu), lead (Pb), and zinc (Zn) in periphyton, benthic invertebrates, and trout livers from one or more sites in the upper Animas River were significantly greater than those from reference sites. Periphyton from sites downstream from mixing zones of acid and neutral waters had elevated concentrations of aluminum (Al) and iron (Fe) reflecting deposition of colloidal Fe and Al oxides, and reduced algal biomass. Metal concentrations in benthic invertebrates reflected differences in feeding habits and body size among taxa, with greatest concentrations of Zn, Cu, and Cd in the small mayfly Rhithrogena, which feeds on periphyton, and greatest concentrations of Pb in the small stonefly Zapada, a detritivore. Concentrations of Zn and Pb decreased across each trophic linkage, whereas concentrations of Cu and Cd were similar across several trophic levels, suggesting that Cu and Cd were more efficiently transferred via dietary exposure. Concentrations of Cu in invertebrates and trout livers were more closely associated with impacts on trout populations and invertebrate communities than were concentrations of Zn, Cd, or Pb. Copper concentrations in livers of brook trout from the upper Animas River were substantially greater than background concentrations and approached levels associated with reduced brook trout populations in field studies and with toxic effects on other salmonids in laboratory studies. These results indicate that bioaccumulation and transfer of metals in stream food webs are significant components of metal exposure for stream biota of the upper Animas River watershed and suggest that chronic toxicity of Cu is an important factor limiting the distribution and abundance of brook trout populations in the watershed. PMID- 11116341 TI - Heavy metal concentrations in marine fishes collected from fish culture sites in Hong Kong. AB - The levels of six heavy metals (Cd, Cr, Cu, Ni, Pb, and Zn) in different tissues of three species of cultured marine fishes (Epinephelus areolatus, Lutjanus russelli, and Sparus sarba) collected from three fish culture sites in Hong Kong were evaluated. Metal pollution problems in the fish culture sites were serious, as reflected by the high metal concentrations recorded in sea water, sediments, and the biomonitor Perna viridis. In general, tissues of all three species contained high concentrations of Zn and Cu, but much lower concentrations of Ni, Pb, Cd, and Cr. Similar pattern of heavy metal concentrations was observed in sea water, sediment, and P. viridis. Metal concentrations in various tissues varied greatly among species and among fish culture sites. Different tissues showed different capacity for accumulating heavy metals. Gonad of all three species contained high concentrations of Zn. On the other hand, liver seemed to be the primary organ for Cu accumulation. Overall, metal concentrations in the tissues of culture marine fishes were much lower than those in P. viridis. Despite high metal levels in sea water and sediments, concentrations of Cd, Cr, and Pb in edible tissues, including muscle and skin, did not exceed permissible levels recommended by the Hong Kong Government for human consumption. PMID- 11116342 TI - Behavioral dysfunctions correlate to altered physiology in rainbow trout (Oncorynchus mykiss) exposed to cholinesterase-inhibiting chemicals. AB - We selected four metrics of swimming behavior (distance swam, speed, rate of turning, and tortuosity of path) and the commonly used biochemical marker, brain cholinesterase (ChE) activity, to assess (1) the sensitivity and reliability of behavior as a potential biomarker in monitoring work, (2) the potential for these endpoints to be used in automated monitoring, and (3) the linkage between behavior and its underlying biochemistry. Malathion-exposed fish exhibited large decreases in distance and speed and swam in a more linear path than control fish after 24 h exposure. By 96 h exposure, fish still swam slower and traveled less distance; fish fully recovered after 48 h in clean water. Diazinon-exposed fish exhibited decreases in distance, speed, and turning rate compared to controls. After 48 h recovery in clean water, fish exposed to diazinon had not recovered to control levels. The behavioral responses provided measures of neurotoxicity that were easily quantifiable by automated means, implying that the inclusion of behavior in monitoring programs can be successful. Furthermore, correlations between behavior and biochemical endpoints, such as ChE inhibition, suggest that this approach can provide a meaningful link between biochemistry and behavior and can provide useful information on toxicant impacts. PMID- 11116343 TI - Nonlethal method for forensic evaluation of aldicarb exposure in wildlife. AB - Forensic evaluation of aldicarb exposure is difficult due to the rapid hydrolysis and oxidation of the parent compound. Oxidation products-aldicarb sulfoxide and aldicarb sulfone-are commonly analyzed, but hydrolytic products-aldicarb nitrile, aldicarb nitrile sulfoxide, aldicarb nitrile sulfone-are infrequently analyzed even though they are the primary stable products of aldicarb degradation. Nitrile analyses provide an important avenue to verify aldicarb exposure or aldicarb induced mortality. Our aproach allows lethal and sublethal exposure assessment. Extraction of samples with acetonitrile:water is followed by chromatographic determination. Sublethal exposure assessment utilizes excreta samples, which is nonlethal and requires holding animals in captivity for 12 h or less. Sublethal exposures of northern bobwhite Colinus virginianus to aldicarb can be identified with greater than 80% confidence for 6 h after dosing. By analyzing GI tracts, lethal exposures of bobwhite to aldicarb can be identified with greater than 90% confidence for 4 days post mortem and with 75% confidence for 8 days post mortem. Sublethal exposures to aldicarb was identified in greater than 80% of Peromyscus maniculatis for 6 h after dosing. Aldicarb and its transformation products were detected for 8 days post mortem in all mice that received aldicarb doses at or above the LD50. PMID- 11116344 TI - Organochlorine contaminants and biomarker response in double-crested cormorants nesting in Green Bay and Lake Michigan, Wisconsin, USA. AB - Double-crested cormorant (Phalacrocorax auritus) eggs at pipping and sibling 10 day-old chicks were collected from two colonies in Green Bay, WI, one colony in Lake Michigan, WI, and reference colonies in South Dakota and Minnesota. Egg contents and chicks were analyzed for organochlorine contaminants including polychlorinated biphenyl (PCB) congeners. Livers of embryos and chicks were assayed for hepatic microsomal ethoxyresorufin-O-dealkylase (EROD) activity. Eggshell thickness and the physical dimensions of embryo brains were measured. Concentrations of organochlorines, including p,p'-DDE (p,p' dichlorodiphenyldichloroethylene), PCBs, and PCB congeners were generally an order of magnitude higher in eggs and chicks from Wisconsin than from reference locations. Total PCBs averaged 10-13 microg/g wet weight in eggs from three Wisconsin colonies compared to 0.9 microg/g PCBs from reference locations. Double crested cormorant chicks accumulated on average 33-66 microg PCBs/day and 7-12 microg p,p'-DDE/day in the Wisconsin colonies compared to 0 microg PCBs/day and 1 microg p,p'-DDE/day in the reference colonies. At pipping, EROD activity in the livers of cormorant embryos was significantly higher in the Wisconsin colonies and significantly correlated with PCBs and the toxic equivalents (TEQs) of aryl hydrocarbon-active PCB congeners relative to 2,3,7,8-tetrachlorodibenzo-p-dioxin. However, in 10-day-old chicks EROD activity was not consistently different among colonies and was not correlated with PCBs or TEQs. A significant negative relationship between embryo brain asymmetry and the size of the egg suggested that physical constraint might be an important factor influencing the response of this bioindicator. Thinner eggshells in two colonies located near Door County, Wisconsin, suggested that historic p,p'-DDE residues associated with orchards are still an important source of p,p'-DDE in the local environment. PMID- 11116345 TI - Chlorinated hydrocarbons and biomarkers of exposure in wading birds and fish of the lower Rio Grande Valley, Texas. AB - During 1997 we evaluated reproductive success in colonial water birds nesting in the Lower Rio Grande Valley (LRGV), Texas, and correlated success with concentrations of contaminants in eggs. We also measured steroid hormones and gonadosomatic index (GSI) as biomarkers of endocrine effects in common carp (Cyprinus carpio). Nest and fledging success of green herons (Butorides virescens) and great egrets (Ardea alba) were similar to those found in other parts of North America; however, nesting success of black-crowned night-herons (Nycticorax nycticorax) was lower, very likely due to flooding of the nesting area. Except for DDE and toxaphene, all chlorinated pesticides in bird eggs were low and not of concern for negative effects on any of the three species. DDE was highest in green heron eggs and seemed to increase along a geographic gradient from west to east, with eggs from Falcon Reservoir containing low concentrations, and those at Los Indios containing the highest concentrations (approx. 11,000 ng/g WW), near or above the threshold for reproductive impairment. DDE levels in great egrets and black-crowned night-herons were below those that are associated with reproductive impairment. Mean DDE levels in carp at the JAS Farms site were above the threshold level suggested for predator protection. Toxaphene was detected in about 20% of the samples with high levels observed in green heron eggs from Los Indios (mean = 4,402 ng/g WW). These are the highest toxaphene levels reported in bird eggs in the LRGV. Toxaphene levels in fish ranged between 90 and 312 ng/g WW. In general, PCBs in bird eggs and fish tissue were low and at levels not of concern for reproductive effects. The greatest concentrations of testosterone and 11-ketotestosterone were detected in fish from the JAS Farms site, which also had the greatest concentrations of DDE. Increased androgen production and gonad development in fish at this site, relative to Pharr, could be possibly associated with endocrine disrupting effects of p,p'-DDE. DDE, toxaphene, PCBs, and hormones were highest in birds and fish from the eastern edge of the study area. PMID- 11116346 TI - Cyclodiene insecticide, DDE, DDT, arsenic, and mercury contamination of big brown bats (Eptesicus fuscus) foraging at a Colorado Superfund site. AB - Rocky Mountain Arsenal (RMA) National Wildlife Area, near Denver, Colorado, is a Superfund site contaminated by past military and industrial uses, including pesticide manufacturing. From an ecosystem standpoint, the most critical contaminants at RMA are certain cyclodiene insecticides and metabolites, p,p' DDE, p,p'-DDT, arsenic, and mercury. Bats are important ecosystem components that can be impacted by persistent contaminants because of their position in the food chain and their potential longevity and thus duration of exposure. Big brown bats (Eptesicus fuscus) were captured (n = 51) while foraging at RMA in the summers of 1997 and 1998 for determination of concentrations of contaminants of concern in carcasses, brains, and stomach contents. Adult females (n = 15) were also tracked by radiotelemetry to determine locations of nearest maternity roosts for sampling of guano for contaminant analysis and inspection for potential contaminant induced mortality. Bats captured while foraging at RMA had measurable quantities of dieldrin and DDE in masticated insect samples from stomach contents and significantly higher concentrations of dieldrin, DDE, DDT, and mercury (juveniles) in carcasses than big brown bats (n = 26) sampled at a reference area 80 km to the north. Concentrations of dieldrin and DDE in brains of bats captured while foraging at RMA were also greater than in bats from the reference area, but not high enough to suggest mortality. Maximum concentrations of DDE, DDT, and cyclodienes in brains of big brown bats were found in adult males from RMA. Guano from the two closest known roosts had significantly higher concentrations of dieldrin, DDE, and mercury than guano from two roosts at the reference area. Dieldrin concentrations in carcasses of bats from RMA were highest in juveniles, followed by adult males and adult females. DDE concentrations in carcasses were lowest in adult females at both sites and highest in adult males at RMA. No contaminant-related mortality was obvious at the small maternity colonies near RMA. Big brown bats show higher contamination than most other mammals previously sampled at this site. Concentrations and proportions of samples with detectable residues of dieldrin in carcasses of big brown bats from RMA were similar to or exceeded reports for this species from elsewhere in the United States some 25 years ago, prior to or just following restrictions on use of this compound. PMID- 11116347 TI - Spatial variation in mercury concentrations in wild mink and river Otter carcasses from the James bay territory, Quebec, Canada. AB - Mercury concentrations were analyzed in different organs/tissues of wild minks (Mustela vison) and river otters (Lutra canadensis) trapped during two seasons in the James Bay territory (49 degrees N to 55 degrees N, Quebec, Canada). In 1993 94, mean total Hg concentration (microg/g, wet weight) in 39 wild mink and 12 river otter carcasses was greatest in fur/hair samples (30.1 and 20.7 microg/g, respectively) and least in brain samples (0.96 and 0.8 microg/g, respectively) with liver, kidney, and muscle samples showing intermediate values. Pooling data from the 1993-94 and 1994-95 trapping seasons revealed mean (SD) liver total Hg concentrations of 3.71 microg/g (3.91) in 316 wild mink carcasses and 4.05 microg/g (3.41) in 153 river otter carcasses. Log liver total Hg concentration increased with age in wild mink but not in river otter. Log liver total Hg concentration in each species was greatest in areas with moraine deposits and least in areas with rich clay deposits, but the effect of soil deposits could be confounded by uneven deposition of anthropogenic Hg. Controlling for type of soil deposits, log liver total Hg concentration decreased with increasing distance from local industrial centers in each species but varied little with changes in distance from hydroelectric reservoirs. In a subsample of carcasses from the moraine sector, log liver total Hg concentration was higher in wild mink than in river otter. Spatial variation in log liver total Hg concentration in relation to soil deposit type and proximity to industrial centers suggests that the two wild furbearer species could be useful indicators of environmental contamination. PMID- 11116348 TI - Oral bioavailability of lead and arsenic from a NIST standard reference soil material. AB - The oral bioavailability of soil contaminants is measured using in vitro or in vivo techniques. Current efforts in our laboratory are focused on the comparisons of in vitro methods for bioavailability estimation with the presently employed in vivo techniques, such as animal models. We present a comparison of two techniques for oral bioavailability estimation: in vitro dissolution and in vivo rat feeding using a standard reference soil. Lead (Pb) and arsenic (As) were chosen because of the range of concentration in this soil as well as the large historical database of bioavailability values for these metals. Metal solubility was measured using a sequential soil extraction in synthetic analogues of human saliva, gastric and intestinal fluids. The soluble metal was defined as the bioaccessible fraction. Oral bioavailability of Pb and As was measured in Sprague Dawley rats by determining metal levels in the major organs and urine, feces, and blood at 1-, 2-, and 3-day time points. Extractions to determine bioaccessibility yielded a gastric component of 76.1% and 69.4% for Pb and As, respectively, and intestinal components were 10.7% and 65.9%. The oral bioavailability of the standard reference soil was 0.7% and 37.8% for Pb and As, respectively. Bioaccessibility was greater than bioavailability for both metals in both gastrointestinal compartments. Although Pb had the highest soil concentration of the selected metals, it was the least bioavailable, while As was highly available in both the in vitro and in vivo method. These types of data allow for an in vitro-in vivo comparison of a soil whose metal concentrations have been certified and validated. PMID- 11116349 TI - Internal exposure to polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDDs/PCDFs) of Bavarian chimney sweeps. AB - This study was carried out to evaluate the internal exposure to polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDDs/PCDFs) of chimney sweeps in Bavaria compared to a control group without occupational exposure. The PCDD/PCDF concentrations in the blood fat of 227 chimney sweeps were compared with the concentrations in samples from 60 controls. Using an internal standard containing 17 (13)C(12)-labeled PCDD/F congeners, the samples were cleaned up after fat elution using standard methods. The statistical analysis was adjusted to account for demographic differences, dietary habits, smoking status, and both occupational and nonoccupational contact with chlorinated hydrocarbons. Detailed information on the type of heating in the households swept, the length of time the chimney sweeps had carried out the profession (min 34, med 195, max 466 months) and the protective measures employed, were used to examine the influence of the working conditions specific to chimney sweeps on the internal PCDD/PCDF exposure. The correlation between blood-fat PCB concentrations as well as urinary chlorophenol concentrations and the exposure to PCDDs/PCDFs was evaluated. The sum of PCDD/PCDF components in chimney sweeps, expressed by International Toxic Equivalents (I-TEQ), was significantly increased compared to the control group (median: 26.36 versus 20.75 pg I-TEQ/g blood fat). For 37 chimney sweeps (16.3%) the sum of PCDDs/PCDFs exceeded the 95th percentile of the control group, i.e., 38.23 pg I-TEQ/g blood fat. Multiple regression analysis revealed that in addition to occupation, the variables age, district, and proximity to a waste incineration plant seem to have an effect on the internal PCDD/PCDF exposure. An additional influence on the internal exposure could not be determined for any of the special aspects of the work. We identified no high correlations between the concentrations of PCBs and chlorophenols and PCDDs/PCDFs. This study revealed significantly higher internal exposure to PCDDs/PCDFs in chimney sweeps than in the control group. The differences are small and within the range of the internal exposure to PCDDs/PCDFs in blood found in the general population in Germany since 1989. Further investigations in to PCDD/PCDF-related diseases in these study groups were not carried out. PMID- 11116350 TI - Cholelithiasis and choledocholithiasis: diagnostic imaging. PMID- 11116351 TI - Imaging of cholelithiasis: what does the surgeon need? PMID- 11116352 TI - Choledocholithiasis: role of US and endoscopic ultrasound. PMID- 11116353 TI - Imaging of cholelithiasis: helical CT. PMID- 11116354 TI - Use of magnetic resonance cholangiography in the diagnosis of choledocholithiasis. PMID- 11116355 TI - Intraabdominal extralobar pulmonary sequestration exhibiting cystic adenomatoid malformation: prenatal diagnosis and characterization of a left suprarenal mass in the newborn. AB - An intraabdominal extrathoracic pulmonary sequestration (IEPS) was detected by prenatal ultrasound in a fetus of 19 weeks' gestation. The well-defined echogenic mass, including multiple cystic areas, was located in the left suprarenal region. Knowledge of the characteristic ultrasound appearance helped to differentiate between neuroblastoma and IEPS before surgical treatment. Histologic examination showed an association between IEPS and features of cystic adenomatoid malformation type 2. PMID- 11116356 TI - Changes in hepatic hemodynamics after orthotopic liver transplantation: color Doppler sonography. AB - BACKGROUND: Liver perfusion has an influence on therapy results in patients undergoing orthotopic liver transplantation (OLT). The objective of the present study was to investigate changes in hepatic hemodynamics in patients after OLT with color-coded Doppler sonography (CCDS). METHODS: Forty-five consecutive patients were included. The examinations were done before, on postoperative day 1, and then weekly until the patients were discharged. Mean velocity of the portal (PV-V) and splenic (SV-V) veins and the maximum velocity and resistance index of the hepatic artery were determined. RESULTS: After OLT a significant increase in PV-V and SV-V was observed. Twenty-five patients had normal perfusion of the hepatic artery, whereas 16 patients had abnormal flow patterns. In these patients prostaglandin I(2) was used until flow rates normalized. In four patients, CCDS could not detect perfusion of the hepatic artery. CONCLUSIONS: CCDS is a suitable method for evaluating hepatic hemodynamics before and after OLT. Changes in blood flow velocities in the liver-supplying vessels are detectable, but perfusion of the hepatic artery is seldom detectable. These observations are of special interest after OLT, where liver circulation has an influence on therapy results. PMID- 11116357 TI - Capsular retraction in hepatic giant hemangioma: CT and MR features. AB - We report two cases of hepatic giant hemangiomas with capsular retraction of the liver adjacent to the tumor on computed tomography and magnetic resonance images. Our cases show that the retraction of the liver capsule adjacent to the tumor is not a finding specific to malignant hepatic tumors but can also be observed in benign hepatic tumors. PMID- 11116358 TI - Hyperechoic lines as a sonographic confirmatory sign during percutaneous transhepatic biliary drainage. AB - BACKGROUND: In ultrasonically guided percutaneous transhepatic biliary drainage, we often can recognize hyperechoic lines at the tip of the needle when the duct is penetrated successfully. We evaluated the frequency of this phenomenon and analyzed whether it was a useful sign for confirming successful bile duct puncture. METHODS: In 65 patients with biliary tract diseases, 84 catheters were placed in the course of 108 attempts at puncture. Results of puncture and the presence of hyperechoic lines were investigated prospectively. RESULTS: When the ultrasonographic findings showed hyperechoic lines, successful puncture was significantly more frequent than when the findings did not show hyperechoic lines (53/55, 96%, vs. 31/59, 53%; p < 0.0001). When we judged the hyperechoic lines as the sign of successful puncture, the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 63%, 93%, 96%, 48%, and 71%, respectively. CONCLUSION: Hyperechoic lines are a useful confirmatory sign of successful puncture. However, absence of these lines was not invariably associated with unsuccessful puncture. PMID- 11116359 TI - Evaluation of a modified cholangiographic classification system for primary sclerosing cholangitis. AB - BACKGROUND: There is no uniformly accepted classification system for the range of cholangiographic abnormalities encountered in primary sclerosing cholangitis (PSC). The aims of this study were to evaluate a previously developed classification system and to test the hypothesis that the pancreatic duct can be involved in PSC. METHODS: Two observers scored 132 endoscopic retrograde cholangiopancreatographies (ERCPs) from established PSC patients. From 30 patients, subsequent ERCPs were scored and compared with the initial ERCPs. The pancreatic duct was judged with regard to morphologic abnormalities. RESULTS: The classification system was applicable in 107 patients. In 10 ERCPs (7.6%), no clear intrahepatic abnormalities were found; 15 other ERCPs (11.4%) did not show extrahepatic abnormalities. In 30 cases, a subsequent ERCP was judged. The difference in scoring between the initial and the subsequent ERCPs was statistically significant, with the subsequent ERCP having higher intrahepatic and extrahepatic scores. Sixty-four adequately filled pancreatic ducts were analyzed. In two cases (3.1%), morphologic abnormalities were found. CONCLUSIONS: The previously developed scoring system is very applicable for almost all PSC patients when supplemented with a type 0 category. Scoring increased over time, suggesting a correlation with disease severity. The pancreatic duct does not seem to be involved in PSC. PMID- 11116361 TI - Primary splenic malignant lymphoma associated with hepatitis C virus infection. AB - We present two cases of primary splenic malignant lymphoma associated with chronic hepatitis C virus infection detected early by routine follow-up imaging studies. Septumlike structures were seen on postcontrast computed tomography or magnetic resonance imaging, which presented characteristic gross findings. These findings may suggest primary splenic malignant lymphoma during the course of chronic hepatitis C. PMID- 11116360 TI - Diagnostic pitfalls in the cholangiographic diagnosis of choledochoceles: cholangiographic quality and its effect on visualization. AB - BACKGROUND: We wanted to establish reasonable cholangiographic diagnostic criteria by determining the sensitivity of cholangiography in detecting choledochoceles and those factors that could compromise visualization of choledochoceles. METHODS: Over 4 years, 21 patients (seven male, 14 female; mean age = 67 years) were confirmed as having choledochoceles on endoscopic retrograde cholangiopancreatography (ERCP). Cholangiographic diagnosis was made by following three criteria: a radiolucent halo around the distal common bile duct (CBD), bulbous dilatation of the distal CBD, and the presence of sequential morphologic changes on serial cholangiography. Any two or more combinations of these three criteria were considered enough to diagnose a choledochocele on cholangiography. We compared cholangiographic imaging findings with the ERCP results. RESULTS: Of 21 patients with choledochoceles, nine (43%) were correctly diagnosed on cholangiography. A radiolucent halo was present in six (28%) patients; four of these cases showed optimal duodenal filling, one showed faint duodenal filling, and one showed poor duodenal filling. The shapes of the distal CBD were bulbous, conelike, and blunt. Morphologic changes such as collapsing and bulging of the choledochocele could be seen in 12 (57%) patients on serial cholangiography. Waists were seen in 11 (52%), pseudowebs in four (19%), and wrinkling of the distal CBD in seven (33%). CONCLUSION: Cholangiography should be obtained with optimal timing and adequate conditions to diagnose choledochocele correctly. PMID- 11116362 TI - Nonenhanced magnetic resonance imaging of mild acute pancreatitis. AB - BACKGROUND: Computed tomography (CT) is not always effective for demonstrating mild acute pancreatitis, and the intravenous administration of iodine contrast medium is harmful to the inflamed pancreas. The goal of this study was to evaluate the usefulness of nonenhanced magnetic resonance (MR) imaging for the depiction of mild acute pancreatitis. METHODS: We performed T1-weighted imaging with a short echo time, T2-weighted imaging, and MR cholangiopancreatography (MRCP) in 12 patients with mild acute pancreatitis. Nonenhanced CT and contrast enhanced CT were always performed before the MR studies. RESULTS: T1- and T2 weighted MR images using a breath-hold or respiratory-triggered technique produced clearer images of the inflamed pancreas than did CT. Peripancreatic fat necrosis was shown by both methods. Although MRCP demonstrated no abnormalities of the pancreatic duct, it demonstrated stones in the gallbladder and common bile duct. CONCLUSIONS: Nonenhanced MR imaging was superior to CT for depiction and confirmation of mild acute pancreatitis. PMID- 11116363 TI - Pancreatic uncinate carcinoma: sonographic findings. AB - BACKGROUND: Pancreatic carcinoma arising from the uncinate process (pancreatic uncinate carcinoma) is relatively rare. We wished to define its clinical manifestations and sonographic findings. METHODS: Clinical and sonographic data of eight cases were reviewed. RESULTS: The common bile duct and the pancreatic duct were not dilated until a very late stage. The lesion mimicked a mesenteric tumor in two cases. The superior mesenteric vessels were compressed anteriorly. Computed tomography was useful, not only for confirming the pancreatic uncinate origin of the lesion but also for determining precisely the mode of mesenteric vascular involvement. CONCLUSION: Knowledge of these unusual sonographic findings can determine the diagnostic strategy in pancreatic uncinate carcinoma. PMID- 11116364 TI - Macrocystic serous cystadenoma of the pancreas: case report. AB - We report a case of macrocystic serous cystadenoma of the pancreas. The lesion consisted of a large main cyst and several small cysts, and each cyst showed high intensity on T1-weighted and very high intensity on T2-weighted magnetic resonance images. High-intensity cyst contents may be a characteristic, if not a specific, finding of macrocystic serous cystadenoma of the pancreas. PMID- 11116365 TI - Pancreatic metastasis: sonographic findings. AB - BACKGROUND: Pancreatic metastasis is a relatively rare pathologic condition. Its ultrasound (US) findings have been infrequently reported, and there has been no previous report describing its color Doppler findings. METHODS: We reviewed the US findings of 13 such cases. RESULTS: The pancreatic metastasis consisted of a single hypoechoic nodule in four cases and multiple hypoechoic nodules in nine cases. US showed a slight dilatation of the main pancreatic duct in two cases of pancreatitis, and a marked biliary dilatation in one case of jaundice. The US results prompted introduction of appropriate treatment in these cases. Color Doppler US was useful for confirming the absence of flow abnormalities in the arterial and portal systems around the pancreas. In a patients with renal carcinoma metastasis, it showed many fine blood flow signals in the metastatic lesions. CONCLUSION: The role of US in the diagnosis of pancreatic metastasis consists of associated complications such as biliary dilatation or pancreatitis and detection of isolated lesions. Additional information provided by color Doppler US may increase diagnostic confidence. PMID- 11116366 TI - Prospective assessment of interobserver agreement for endoanal MRI in fecal incontinence. AB - BACKGROUND: Endoanal magnetic resonance (MR) imaging is a new technique for the assessment of anal sphincter integrity in fecal incontinence and an alternative to anal endosonography. The present study aimed to determine interobserver variation for assessment of anal sphincter integrity using endoanal MR imaging. METHODS: Fifty-two consecutive anally incontinent patients underwent MR imaging by using a purpose-built endoanal receiver coil and static 1.0-T magnet. T2 weighted axial, coronal, and sagittal scans were independently assessed by two radiologists who noted external and internal sphincter integrity. Findings were compared and agreement was assessed with the kappa statistic. RESULTS: There was disagreement in 18 of 49 technically adequate studies (37%; kappa = 0.46), indicating "moderate" agreement. Agreement was strongest if the sphincters were either both intact or both disrupted. Observers agreed in only one diagnosis of an isolated internal sphincter defect and in no diagnosis of an isolated external sphincter defect. CONCLUSION: The overall interobserver agreement for assessment of sphincter integrity using endoanal MR imaging is "moderate." Interobserver agreement using endoanal MR imaging is less than that reported for anal endosonography. PMID- 11116367 TI - Presacral epidermoid cyst: imaging findings with histopathologic correlation. AB - BACKGROUND: The aim of this study was to determine the imaging characteristics of presacral epidermoid cysts and correlate the imaging findings with the histopathologic findings. METHODS: We retrospectively reviewed sonographic, computed tomographic, and magnetic resonance examinations in four consecutive patients with a pathologically proven presacral epidermoid cyst. Imaging findings of the presacral epidermoid cyst were correlated with the histopathologic findings. RESULTS: In all four patients, sonography showed a presacral mass with a heterogeneous low echogenicity, and computed tomography showed a discrete well defined hypodense presacral mass with a thin wall. In the three patients who underwent magnetic resonance imaging, the mass showed a heterogeneous low signal intensity on the T1-weighted image and a high signal intensity with multiple small foci of low signal intensity in the nondependent portion of the mass on the T2-weighted image. These imaging findings correlated well with the pathologic results. Aggregates of keratinous material contributed to these imaging findings. CONCLUSION: In the diagnosis of the presacral epidermoid cyst, sonographic and magnetic resonance imaging findings may be helpful. PMID- 11116368 TI - Acute torsion and necrosis of the greater omentum herniated into a foramen of Morgagni. AB - Computed tomography is mandatory in the investigation of the acute abdomen and can provide the physician with crucial information to decide whether the patient should be treated surgically or conservatively. An unusual cause of acute abdomen is presented. Computed tomography suggested the diagnosis of omental torsion and necrosis. At surgery, the greater omentum and part of the transverse colon were incarcerated in a small diaphragmatic hernia of the Morgagni type. PMID- 11116369 TI - Strangulated transomental hernia: CT findings. AB - We report a case of surgically confirmed strangulation of small bowel through a defect in the greater omentum. Computed tomography demonstrated the presence and the location of this very unusual internal abdominal hernia. Those findings are presented. PMID- 11116370 TI - Supravesical hernia: CT diagnosis. AB - We report two cases of surgically proven supravesical hernia, one an internal supravesical hernia and the other an external supravesical hernia. Abdominal computed tomography showed the relation of the incarcerated intestine anterior to and compressing the urinary bladder. Although neither case was diagnosable preoperatively, we believe that the preoperative diagnosis of supravesical hernia by abdominal computed tomography is possible. PMID- 11116371 TI - Comparative viewing modalities for CT cystography. AB - BACKGROUND: To define the speed and accuracy of two different reconstructive techniques using computed tomography (CT) cystography for the detection and measurement of urinary bladder masses and determine the overall ease of use. METHODS: Ten patients scheduled for cystoscopy for the evaluation of hematuria or bladder masses were studied by means of thin-section CT of the air-distended bladder. Two techniques were employed by two radiologists to blindly interpret the data: conventional two-dimensional data with interactive three-dimensional problem solving (2D3DPS) and surface-shaded display (SSD) three-dimensional images. The results were compared with the data from cystoscopy. RESULTS: Twenty two (100%) of 22 masses detected on cystoscopy were visualized using the reconstructive techniques. Both modalities were shown to have high accuracy, but only the 2D3DPS had a sensitivity and specificity of 100% for both observers at the patient-level diagnosis. The sensitivities for detecting individual masses for the two observers were 100% and 64% for 2D3DPS and 64% and 70% for SSD. CONCLUSION: Both methods used to display the CT data had a high sensitivity and specificity for masses, but only the 2D3DPS had a sensitivity and specificity of 100% at the patient-level diagnosis, thus making it a feasible imaging modality for cystography. It was also preferred overall for ease of use, high accuracy, and relative low cost. PMID- 11116372 TI - Myxoid leiomyoma of the uterus: CT and MRI features. AB - A myxoid leiomyoma of the uterus is radiologically described. Sonography showed a large mass extending from the renal hilum to the pelvis. Enhanced computed tomography and magnetic resonance imaging findings denoted cystic and myxoid components. Magnetic resonance imaging demonstrated flow voids between the mass and the uterus, indicating likely origin from the uterus. Exploratory laparotomy showed a huge leiomyomatous mass. PMID- 11116377 TI - The politics of participation in watershed modeling. AB - While researchers and decision-makers increasingly recognize the importance of public participation in environmental decision-making, there is less agreement about how to involve the public. One of the most controversial issues is how to involve citizens in producing scientific information. Although this question is relevant to many areas of environmental policy, it has come to the fore in watershed management. Increasingly, the public is becoming involved in the sophisticated computer modeling efforts that have been developed to inform watershed management decisions. These models typically have been treated as technical inputs to the policy process. However, model-building itself involves numerous assumptions, judgments, and decisions that are relevant to the public. This paper examines the politics of public involvement in watershed modeling efforts and proposes five guidelines for good practice for such efforts. Using these guidelines, I analyze four cases in which different approaches to public involvement in the modeling process have been attempted and make recommendations for future efforts to involve communities in watershed modeling. PMID- 11116373 TI - Polyarteritis nodosa of the epididymis. AB - We describe magnetic resonance imaging findings in a 37-year-old man with a rare entity of isolated polyarteritis nodosa of the epididymis, which correlated well with the histopathologic findings. PMID- 11116378 TI - Early development of systems analysis in natural resources management from man and nature to the miami conservancy district. AB - Contemporary approaches to natural resources and environmental decision-making typically draw on a "systems" perspective to assess and improve management strategies. This paper describes the early genesis of the systems analysis approach. It concentrates on a period between the mid-19th to early 20th centuries. During the early part of this period, George Marsh's Man and Nature and related works laid out an approach to problem-solving that recognized the relationship among physically disperse elements in the environment, the need to balance benefits against costs, the potential for using quantitative modeling to understand management options, and the importance of integrating human and natural components into solutions. In the early 20th century, the Miami Conservancy District project brought this approach to fruition with its use of complex simulation and optimization modeling, detailed cost-benefit analysis, and its linking of economics, engineering, science, and law into a far-reaching solution to a complex water resources problem. The objective of this paper is to describe the early development and application of this conceptual approach to problem-solving. An examination of the origins of natural resources systems analysis can broaden one's perspective of the contemporary field to understand its roots as a philosophy for environmental problem-solving. PMID- 11116379 TI - Threshold-based resource management: a framework for comprehensive ecosystem management. AB - The problems posed by adaptive management for improved ecosystem health are reviewed. Other kinds of science-informed ecosystem management are needed for those regions of conflict between rapid human population growth, increased resource extraction, and the rising demand for better environmental amenities, where large-scale experiments are not feasible. One new framework is threshold based resource management. Threshold-based resource management guides management choices among four major science and engineering approaches to achieve healthier ecosystems: self-sustaining ecosystem management, adaptive management, case-by case resource management, and high-reliability management. As resource conflicts increase over a landscape (i.e., as the ecosystems in the landscape move through different thresholds), management options change for the environmental decision maker in terms of what can and cannot be attained by way of ecosystem health. The major policy and management implication of the framework is that the exclusive use or recommendation of any one management regime, be it self-sustaining, adaptive, case-by-case, or high-reliability management, across all categories of ecosystems within a heterogeneous landscape that is variably populated and extractively used is not only inappropriate, it is fatal to the goals of improved ecosystem health. The article concludes with detailed proposals for environmental decision-makers to undertake "bandwidth management" in ways that blend the best of adaptive management and high-reliability management for improved ecosystem health while at the same time maintaining highly reliable flows of ecosystem services, such as water. PMID- 11116380 TI - Benefits of collaborative learning for environmental management: applying the integrated systems for knowledge management approach to support animal pest control. AB - Resource management issues continually change over time in response to coevolving social, economic, and ecological systems. Under these conditions adaptive management, or "learning by doing," offers an opportunity for more proactive and collaborative approaches to resolving environmental problems. In turn, this will require the implementation of learning-based extension approaches alongside more traditional linear technology transfer approaches within the area of environmental extension. In this paper the Integrated Systems for Knowledge Management (ISKM) approach is presented to illustrate how such learning-based approaches can be used to help communities develop, apply, and refine technical information within a larger context of shared understanding. To outline how this works in practice, we use a case study involving pest management. Particular attention is paid to the issues that emerge as a result of multiple stakeholder involvement within environmental problem situations. Finally, the potential role of the Internet in supporting and disseminating the experience gained through ongoing adaptive management processes is examined. PMID- 11116381 TI - Proactive management of air quality. AB - Traditional air resource management systems have difficulty in addressing global issues, sustainable development, direct citizen participation, and integration with broad economic interests. As reactive management systems, they tend to be compliance-driven, static, and rigid. In contrast, proactive management systems are principle-driven, innovative, and flexible. Bridge scientists play a key role in supporting the transformation of raw data into wise action. Decision-makers need to integrate social values with knowledge about emissions, atmospheric processes, and potential environmental effects using the primary tools of measurements, monitoring, and modeling. The Alberta Clean Air Strategic Alliance, a unique partnership of governments, industry, and public interest groups formed in 1994, operates a comprehensive air management system that is capable of addressing air issues of greater complexity and uncertainty. Its success is measured by the satisfaction of its diverse stakeholders and by the number and scope of its initiatives. PMID- 11116382 TI - Using decision analysis to choose phosphorus targets for Lake Erie. AB - Lake Erie water quality has improved dramatically since the degraded conditions of the 1960s. Additional gains could be made, but at the expense of further investment and reductions in fishery productivity. In facing such cross jurisdictional issues, natural resource managers in Canada and the United States must grapple with conflicting objectives and important uncertainties, while considering the priorities of the public that live in the basin. The techniques and tools of decision analysis have been used successfully to deal with such decision problems in a range of environmental settings, but infrequently in the Great Lakes. The objective of this paper is to illustrate how such techniques might be brought to bear on an important, real decision currently facing Lake Erie resource managers and stakeholders: the choice of new phosphorus loading targets for the lake. The heart of our approach is a systematic elicitation of stakeholder preferences and an investigation of the degree to which different phosphorus-loading policies might satisfy ecosystem objectives. Results show that there are potential benefits to changing the historical policy of reducing phosphorus loads in Lake Erie. PMID- 11116383 TI - Identifying influences on model uncertainty: an application using a forest carbon budget model. AB - Uncertainty is an important consideration for both developers and users of environmental simulation models. Establishing quantitative estimates of uncertainty for deterministic models can be difficult when the underlying bases for such information are scarce. We demonstrate an application of probabilistic uncertainty analysis that provides for refinements in quantifying input uncertainty even with little information. Uncertainties in forest carbon budget projections were examined with Monte Carlo analyses of the model FORCARB. We identified model sensitivity to range, shape, and covariability among model probability density functions, even under conditions of limited initial information. Distributional forms of probabilities were not as important as covariability or ranges of values. Covariability among FORCARB model parameters emerged as a very influential component of uncertainty, especially for estimates of average annual carbon flux. PMID- 11116384 TI - Use of soil and water protection practices among farmers in three midwest watersheds. AB - Data were collected from 1011 farmers in three Midwestern watersheds (Ohio, Iowa, and Minnesota) to assess factors that influence the use of conservation production systems at the farm level. The "vested interests" perspective used to guide the investigation was derived from elements of social learning and social exchange theories. Respondents were asked to indicate their frequency of use for 18 agricultural production practices that could be adopted on Midwestern farms at the time of the study. Responses to the 18 items were summed to form a composite variable, termed "conservation production index," for use as the dependent variable in multivariate analysis. Eleven independent variables were identified from the theory as likely predictors of conservation adoption, including respondents' perceptions about production costs, output and risks, and perceived importance of access to subsidies, technical assistance, and informational/educational programs. Regression analysis was used to assess the performance of the independent variables in explaining variance in the conservation production index. Explained variance in the three regression models ranged from 2% in the Minnesota watershed to 19% in the Ohio watershed. The researchers concluded that the model had limited utility in predicting adoption of conservation production systems within the three study watersheds. Findings are discussed in the context of conservation programs within the three areas. PMID- 11116385 TI - Plant community composition and biomass in Gulf Coast Chenier Plain marshes: responses to winter burning and structural marsh management. AB - Many marshes in the Gulf Coast Chenier Plain, USA, are managed through a combination of fall or winter burning and structural marsh management (i.e., levees and water control structures; hereafter SMM). The goals of winter burning and SMM include improvement of waterfowl and furbearer habitat, maintenance of historic isohaline lines, and creation and maintenance of emergent wetlands. Although management practices are intended to influence the plant community, effects of these practices on primary productivity have not been investigated. Marsh processes, such as vertical accretion and nutrient cycles, which depend on primary productivity may be affected directly or indirectly by winter burning or SMM. We compared Chenier Plain plant community characteristics (species composition and above- and belowground biomass) in experimentally burned and unburned control plots within impounded and unimpounded marshes at 7 months (1996), 19 months (1997), and 31 months (1998) after burning. Burning and SMM did not affect number of plant species or species composition in our experiment. For all three years combined, burned plots had higher live above-ground biomass than did unburned plots. Total above-ground and dead above-ground biomasses were reduced in burned plots for two and three years, respectively, compared to those in unburned control plots. During all three years, belowground biomass was lower in impounded than in unimpounded marshes but did not differ between burn treatments. Our results clearly indicate that current marsh management practices influence marsh primary productivity and may impact other marsh processes, such as vertical accretion, that are dependent on organic matter accumulation and decay. PMID- 11116386 TI - Methodological development of an index of coastal water quality: application in a tourist area. AB - With the aim of obtaining an index of coastal water quality, a methodological procedure based on numerical classification and discriminant analysis is presented. The procedure was applied to nutrient data (ammonia, nitrite, nitrate, and phosphate) analyzed along the coastal waters of a Spanish tourist area. Using numerical classification, three levels of nutrient loading were revealed, characterizing oligotrophic, mesotrophic, and potentially eutrophic waters. Discriminant analysis was shown to be an effective methodological tool in the discrimination between trophic groups. For every group, the discriminant procedure generated the centroids. The centroids representing oligotrophic and potentially eutrophic conditions were used to establish the two extremes of the continuum of mesotrophic conditions in these coastal waters: Standardizing values from -1 to 1, the centroids for oligotrophic and potentially eutrophic waters yielded an interval that defined the range of mesotrophic conditions. This interval is proposed as a water quality index. The ability of the coastal water quality index to successfully predict mesotrophic conditions was proved with random samples. PMID- 11116387 TI - Assessing health benefits of controlling air pollution from power generation: the case of a lignite-fired power plant in Thailand. AB - The impact pathway approach (IPA) is used to estimate quantitatively the level of health effects caused by particulate matter (PM10) and sulfur dioxide (SO2) emission from a lignite-fired power plant located in the Mae Moh area in northern region of Thailand. Health benefits are then assessed by comparing the levels of estimated health impacts without and with the installation of the flue gas desulfurization (FGD) equipment. The US EPA industrial source complex model is used to model air pollution dispersion at the local scale, and the sector average limited mixing meso-scale model is used to model air pollution transport at the regional scale. The quantification of the health end points in physical terms is carried out using the dose-response functions established recently for the population in Bangkok, Thailand. Monetarization of these effects is based on the benefit transfer method with appropriate adjustment. Finally, it has been found that the installation of the FGD to control SO2 emission at Mae Moh significantly reduces adverse health effects not only on the population living near the power plant but also all over the country. A FGD unit installed at the 300-MW power unit can result, on average, in 16 fewer cases of acute mortality, 12 fewer cases of respiratory and cardiac hospital admissions, and almost 354,000 fewer days with acute respiratory symptoms annually. In monetary terms this benefit is equivalent to US $18.2 million (1995 prices) per annum. This benefit is much higher than the annualized investment and operation costs of FGD (US $7.4 million/yr). PMID- 11116388 TI - Yersinia enterocolitica O9 as a possible barrier against Y. pestis in natural plague foci in Ningxia, China. AB - A survey of Yersinia spp, as related to plague control, was made in Haiyuan of Ganning loess plateau plague focus, Yanchi of Inner Mongolia plateau plague focus, and Yinchuan city, as a control area, in Ningxia, China. In Haiyuan, where the main plague reservoir was Mongolian ground squirrel (Citellus alaschanicus) living in the prairie, Y. enterocolitica O9 was frequently isolated from pigs, dogs, rodents living in and around houses, but only rarely from hare and Mongolian ground squirrel. In Yanchi, where the main plague reservoir was Mongolian gerbil (Meriones unguiculatus) living in the prairie and Y. pestis, which was isolated from rodents up to 1991, Y. enterocolitica O9 was sometimes isolated from pigs and rodents. In all areas, some strains of Y. enterocolitica O3 and Y. pseudotuberculosis serotypes 3 and 4b were also isolated from pigs, dogs, and from rodents. We propose that an epidemiological link exists between the prevalence of Y. pestis and Y. enterocolitica O9 in domestic and rodents living in these areas in China. The residential area in Haiyuan may be protected against Y. pestis by the domestic animals and rodents which acquired cross protection against Y. pestis by infection with Y. enterocolitica O9, but this is not the case in the Yanchi district. PMID- 11116389 TI - Catalase activities of Phanerochaete chrysosporium are not coordinately produced with ligninolytic metabolism: catalases from a white-rot fungus. AB - Phanerochaete chrysosporium uses hydrogen peroxide as a substrate in its ligninolytic phase. To determine how the fungus protects itself against detrimental effects of reactive oxygen species, catalase activity was examined. The fungus produced up to four different catalase isozymes, Cat A to D. Isozyme CatC was the dominant activity in all growth conditions. A minor catalase, CatD, was apparent in low-carbon culture and upon exposure of mycelium grown on high nitrogen medium with 5-50 mm of hydrogen peroxide. In low-nitrogen culture, an increase in catalase-specific activity preceded the onset of ligninolysis by 3 days. In low-carbon culture, catalase was produced at an even higher level without development of ligninolysis. Thus, catalase activity was not coordinately produced with ligninolytic metabolism. PMID- 11116390 TI - In vitro catabolism of histidine by mixed rumen bacteria and protozoa. AB - An in vitro study was conducted to examine the metabolism of histidine (His) by mixed rumen bacteria (B), mixed rumen protozoa (P), and a combination of the two (BP). Rumen microorganisms were collected from fistulated goats fed with lucerne cubes (Medicago sativa) and a concentrate mixture twice a day. Microbial suspensions were anaerobically incubated with or without 2 mm each of His, or histamine (HTM), or 1 mm urocanic acid (URA) at 39 degrees C for 12 h. His and other related compounds in both supernatant and microbial hydrolysates were analyzed by HPLC. After 6- and 12-h incubations, the net degradation of His was 26.1% and 51.7% in B, 13.5% and 20.9% in P, and 21.7% and 46.0% in BP, respectively. The rate of the net degradation of His in B (98.0 micromol/g microbial nitrogen/h) was about 2.6 times higher than that of P during a 12-h incubation period. His was found to be degraded into urocanic acid (URA), imidazolelactic acid (ImLA), imidazoleacetic acid (ImAA), and histamine (HTM). Of these degraded His was mainly converted into URA in all microbial suspensions. The production of ImLA and ImAA was higher in B than in P suspensions, whereas the production of HTM was higher in P than in B suspensions. From these results, the existence of diverse catabolic routes of His in rumen microorganisms was indicated. PMID- 11116391 TI - Kinetics characterization of taurocholic transport in Lactobacillus reuteri. AB - Taurocholic acid transport in Lactobacillus reuteri CRL 1098 was determined. The bile acid is incorporated inside the cells by an active and saturable transport showing a typical kinetics of Michaelis-Menten with values of Km and Vmax of 0.35 mm and 20 mm, respectively. PMID- 11116392 TI - A low-pH-inducible, stationary-phase acid tolerance response in Lactobacillus acidophilus CRL 639. AB - Acidity is an important environmental condition encountered by lactobacilli during food fermentation. In this report we show that triggering the stationary phase acid tolerance response (ATR) in L. acidophilus CRL 639 depends on the final growth pH. In free-pH fermentation runs (final pH = 4.5), the cells were completely resistant to acid stress, whereas cells from cultures under controlled pH (pH = 6.0) were very sensitive. The relationship between the final pH and the development of cross-resistance to different kinds of environmental stress was also evaluated. The study of protein profiles showed the overexpression of 16 proteins from 6.5 to 70.9 kDa in stationary phase cells. Seven of these proteins (26.3, 41.4, 48.7, 49.3, 54.5, 56.1, and 70.9 kDa) were expressed as result of the stationary phase itself, while nine proteins (14.1, 18.6, 21.5, 26.9, 29.3, 41.9, 42.6, 49.6, and 56.2 kDa) were exclusively induced as a result of the drop in culture pH during free fermentation runs. These results strongly suggest the involvement of these proteins in cell adaptation to environmental changes. PMID- 11116394 TI - Evaluation of nitrogenous substrates such as peptones from fish:a new method based on Gompertz modeling of microbial growth. AB - Fish peptones from tuna, cod, salmon, and unspecified fish were compared with a casein one by using a new method based on Gompertz modeling of microbial growth. Cumulative results obtained from six species of bacteria, yeasts, and fungi showed that, in most cases, these fish peptones are very effective. Nevertheless, this study raised some questions about the standardization of fish raw material, the enzymatic hydrolysis of fish proteins, and the composition of the culture medium used for testing the peptones. PMID- 11116393 TI - Numerical taxonomy of Sarothamnus scoparius rhizobia. AB - Twenty nodule isolates from Sarothamnus scoparius (broom) growing in Poland and nine strains from plants growing in Japan were studied for phenotypic properties, plasmid presence, phage sensitivity, and host plant specificity. By numerical analysis of phenotypic properties, it was found that the studied nodule bacteria, originating from geographically different countries, constitute two separate groups affiliated to the bradyrhizobium cluster. The membership of S. scoparius rhizobia in the Bradyrhizobium genus was also supported by their long generation time, alkaline reaction in YEM medium with mannitol, lack of plasmids, and wide host plant range. PMID- 11116395 TI - Lactobacillus acidophilus autolysins inhibit Helicobacter pylori in vitro. AB - Antibacterial activity of 17 strains of lactobacilli was tested against 10 strains of H. pylori. The inhibition observed was related to the acid production and the low pH attained. No relationship between CagA phenotype of H. pylori strains and tolerance to lactic acid was observed. In mixed cultures, L. acidophilus CRL 639 showed an autolytic behavior after 24 h of culture. At this moment, H. pylori CCUG17874 showed a decrease of 2 log-cycle, and no viable count was detected after 48 h. The bactericidal effect of L. acidophilus CRL 639 in mixed cultures is related to a proteinaceous compound released after cell lysis. PMID- 11116396 TI - Molecular differentiation of Lactococcus lactis subspecies lactis and cremoris strains by ribotyping and site specific-PCR. AB - Twenty-five strains of Lactococcus lactis subspecies lactis and subspecies cremoris obtained from dairy industry and environmental collections were examined by 16S RNA automated ribotyping profiles and site-specific PCR (S-PCR). By automated ribotyping, the majority of strains were classified in accordance with phenotypic characterization, with the exception of one lactis (220) and two cremoris (BO32 and 140) strains. A complete differentiation of subspecies lactis and cremoris in agreement with conventional phenotypic methods was achieved by S PCR with a set of site-specific primer pairs (PR1, RM4, and F3) designed particularly from a deletion region found in subspecies cremoris, but not in lactis. Therefore, S-PCR with primers (PR1, RM4, and F3) is a rapid and very sensitive method for the distinction of lactis and cremoris subspecies in dairy production. PMID- 11116398 TI - Poly-beta-hydroxybutyrate metabolism is affected by changes in respiratory enzymatic activities due to cold stress in two psychrotrophic strains of Rhizobium. AB - Cold stress resulted in a decrease in the poly-beta-hydroxybutyrate (PHB) content of non-cold-acclimated Rhizobium DDSS69 cultures. Analysis of the specific activity of beta-ketothiolase and beta-hydroxybutyrate dehydrogenase revealed that decrease in PHB levels was a result of the inhibition of synthesis of PHB rather than an increase in its breakdown. Rhizobium ATR1, a cold-acclimated strain, revealed the presence of a stable PHB metabolism that did not show any significant differences either in PHB levels or in the activity of enzymes of the PHB metabolism under cold stress, suggesting that PHB is not involved in cold tolerance. Analysis of specific activities of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase of the pentose phosphate pathway showed the upward regulation of alternate pathways of carbohydrate metabolism under cold stress to rapidly generate energy to overcome the stress. There is diversity in the switching mechanisms of carbon metabolism among cold-acclimated and non-cold acclimated Rhizobium isolates. Upward regulation of malate dehydrogenase in both isolates suggests that it is a critical input for cold tolerance. PMID- 11116399 TI - Survival and nodulating ability of indigenous and inoculated Rhizobium leguminosarum biovar trifolii in sterilized and unsterilized soil treated with sewage sludge. AB - Rhizobium leguminosarum biovar trifolii was detected in soil from 41 of 47 plots, within nine sewage sludge-treated sites with different soil characteristics and heavy metal contents. However, although population size varied widely, there was no consistent correlation with soil heavy metal concentration. Indigenous populations in 20 plots within four selected sites retained their ability to induce effective nodule formation after incubation of soil in the dark for 165 days. In sterilized (gamma-irradiated) soil, Rhizobium survival varied from 0.01% to 95% depending on the soil sample and strain used. Metal-resistant strains with non-mucoid colonies survived less well than mucoid metal-sensitive strains. PMID- 11116397 TI - Identification of a Gardnerella vaginalis hemoglobin-binding protein. AB - Previous studies have shown that Gardnerella vaginalis can utilize human hemoglobin as a sole source of iron. In this study, the interaction between human hemoglobin and G. vaginalis cells was investigated. With a solid phase dot blot assay, G. vaginalis cells were shown to bind digoxigenin (DIG)-labeled human hemoglobin. A human hemoglobin-binding protein with an estimated molecular weight of 124 kilodaltons (kDa) was detected by Western blot analysis of G. vaginalis proteins. The hemoglobin-binding activity of this protein was found to be heat stable and was observed in G. vaginalis cells grown under iron-restrictive and iron-replete conditions. The 124-kDa hemoglobin-binding protein was not detected from intact G. vaginalis cells treated with trypsin prior to Western blot analysis, suggesting that this protein was surface exposed. PMID- 11116400 TI - Cgr1p, a novel nucleolar protein encoded by Saccharomyces cerevisiae orf YGL0292w. AB - Saccharomyces cerevisiae open reading frame (ORF) YGL029w (CGR1) encodes a small hydrophilic protein of unknown function. To investigate the role of this gene, we have determined the intracellular localization of the encoded product and examined the effects of Cgr1p depletion on cell growth. Tagging Cgr1p with the green fluorescent protein (GFP) or the myc epitope showed focal accumulation of the fusion protein in the yeast nucleolus, and this localization overlapped with the distribution of the nucleolar protein Nop1p. Cells depleted of CGR1 mRNA were growth impaired and hypersensitive to the translational inhibitor paromomycin, and this phenotype was complemented by episomal expression of the CGR1-GFP fusion gene. These results identify Cgr1p as a novel component of the yeast nucleolus and suggest a potential role in ribosome biogenesis. PMID- 11116401 TI - Once more, with feeling: handoscopy or the rediscovery of the virtues of the surgeon's hand. PMID- 11116402 TI - Minimally invasive parathyroid surgery. AB - BACKGROUND: Minimally invasive access for the treatment of primary hyperparathyroidism is becoming widespread, but several different approaches have been proposed in the literature. METHODS: We describe the three main types of mini-invasive parathyroidectomy, with particular attention to the gasless video assisted procedure, which is now routinely performed at our institution. RESULTS: Eighty-nine patients with a preoperatively localized single adenoma were successfully treated. Operative time was 58 mins, and there were only five conversions. DISCUSSION: After comparing the different approaches described in literature, we conclude that mini-invasive parathyroidectomy is feasible and can provide additional benefits not available with traditional surgery. At present, however, this operation can be recommended only for patients with sporadic disease, localized lesions, and absence of goiter and prior neck surgery. PMID- 11116403 TI - Laparoscopic hand-assisted surgery for hepatic and pancreatic disease. AB - Herein I describe my initial experience with the use of a novel device, the Omniport, in 15 patients undergoing hand-assisted laparoscopic surgery (HALS) on the liver and pancreas. The device, which essentially consists of a hand cuff with a spiral inflatable valve, enables withdrawal and reinsertion of the hand without loss of pneumoperitoneum during the operation. The cuff's effective sealing pressure is equal to the pneumoperitoneal pressure; hence, hand comfort is maintained during the intervention. The device was effective in maintaining pneumoperitoneum in all cases. All but one operation was completed with the HALS approach. The one conversion was due to bleeding from the superior mesenteric vein during a 90% pancreaticosplenectomy. Immediate effective control of the bleeding by compression between the thumb and index finger was achieved, and the cuff of the Omniport was deflated as the incision was enlarged. There were no postoperative complications. The HALS approach has distinct advantages in terms of exposure and safety over the total laparoscopic technique for major surgery on the liver and pancreas, and it is recommended for these interventions. PMID- 11116404 TI - Hand-assisted laparoscopic splenectomy. AB - Laparoscopic splenectomy is performed routinely in patients with small and moderately enlarged spleens at specialized centers. Large spleens are difficult to handle laparoscopically and hand-assisted laparoscopic splenectomy might facilitate the procedure through enhanced vascular control, easier retraction and manipulation, manual guidance of endostaplers, and clip appliers. A technique of hand-assisted laparoscopic splenectomy is described. PMID- 11116405 TI - Hand-assisted laparoscopic colorectal surgery. AB - Hand-assisted laparoscopic surgery is a newly developed technique. It involves the intraabdominal placement of a hand or forearm through a mini-laparotomy incision while pneumoperitoneum is maintained. In this way, the hand can be used as in an open procedure to palpate organs or tumors, reflect organs atraumatically, retract structures, identify vessels, dissect bluntly along a tissue plain, and provide finger pressure to bleeding points while proximal control is achieved. Additionally, this approach is more economical than a totally laparoscopic approach, reducing both the number of laparoscopic ports and number of instruments required. Some advocates of the technique claim that it is also easier to learn and perform than totally laparoscopic approaches and that it may increase patient safety. PMID- 11116406 TI - Prospective comparison of endorectal ultrasound, three-dimensional endorectal ultrasound, and endorectal MRI in the preoperative evaluation of rectal tumors. Preliminary results. AB - BACKGROUND: The aim of this study was to compare the value of endorectal ultrasound (EUS), three-dimensional (3D) EUS, and endorectal MRI in the preoperative staging of rectal neoplasms. METHODS: Thirty consecutive patients with rectal tumors were assessed by EUS and endorectal MRI. Additionally, three dimensional ultrasound was performed in a subgroup of 25 patients. EUS data were obtained with a bifocal multiplane transducer (10 MHz) and processed on a 3D ultrasound workstation. MR imaging was carried out with a 1. 5 T superconducting unit using an endorectal surface coil. RESULTS: EUS was carried out successfully in all 30 patients, whereas endorectal MRI was not feasible in two patients. Compared with the histopathological classification, EUS and endorectal MRI correctly determined the tumor infiltration depth in 25 of 30 and 28 patients, respectively. The comparative accuracy of EUS, 3D EUS, and endorectal MRI in predicting tumor invasion was 84%, 88%, and 91%, respectively. EUS, three dimensional EUS, and endorectal MRI enabled us to assess the lymph node status correctly in 25, 25, and 24 patients, respectively. Both three-dimensional EUS and endorectal MRI combined high-resolution imaging and multiplanar display options. Assessment of additional scan planes facilitated the interpretation of the findings and improved the understanding of the three-dimensional anatomy. CONCLUSION: The accuracy of three-dimensional EUS and endorectal MRI in the assessment of the infiltration depth of rectal cancer is comparable to conventional EUS. One advantage of both methods is the ability to obtain multiplanar images, which may be helpful for the planning of surgery in the future. PMID- 11116407 TI - Laparoscopic repair of traumatic diaphragmatic hernias. AB - BACKGROUND: Traumatic diaphragmatic hernias are serious complications of blunt abdominal or thoracic trauma. In the early posttraumatic period, they are often missed, and they may be followed by a variety of subacute or chronic symptoms due to pulmonary or intestinal obstruction. METHODS: We present three cases of traumatic diaphragmatic hernias. Two of them were successfully treated by laparoscopy and direct suturing during the early posttraumatic period; the other was treated 10 years after the trauma. RESULTS: We found that laparoscopy is a safe, successful, and gentle procedure not only for diagnosis but for treatment as well. The postoperative course was uneventful in all cases. All patients remained asymptomatic during long-term follow-up (42-60 months). These results are promising. We expect the same good long-term results after laparoscopic repair as after open conventional surgery. CONCLUSION: We recommend that surgeons with sufficient experience in laparoscopy use a minimally invasive approach to treat chronic as well as acute traumatic diaphragmatic hernias in hemodynamically stable patients. PMID- 11116408 TI - Laparoscopic treatment of large paraesophageal hernias: both excision of the sac and gastropexy are imperative for adequate surgical treatment. AB - BACKGROUND: We set out to evaluate the results of the laparoscopic treatment of large paraesophageal hernias in 22 patients. METHODS: Between 1993 and 1998, we operated on 22 consecutive patients. Preoperative assessment consisted of endoscopy, barium esophagogram, 24-h pH testing, manometry, and gastric emptying times. RESULTS: In the first three patients, the sac was not excised and gastropexy was not performed. Because of recurrences, we decided to change the technique in an attempt to avoid further complications. During middle- to long term follow-up, only three recurrences were seen in the subsequent 19 patients. There were no deaths in this series. CONCLUSIONS: Laparoscopic treatment of large paraesophageal hernias is feasible. Because recurrences may occur after successful laparoscopic treatment, both resection of the sac and some form of gastropexy are imperative. PMID- 11116409 TI - Comparison of laparoscopic and open Nissen fundoplication 2 years after operation. A prospective randomized trial. AB - BACKGROUND: Laparoscopic operation has replaced the conventional open procedure in the treatment of gastroesophageal reflux disease (GERD) in spite of the fact that long-term results based on controlled clinical trials have been lacking. The objective of this study was to compare outcome, quality of life, and patient satisfaction after laparoscopic and open Nissen fundoplication in a community hospital setting with a 2-year follow-up. METHODS: Forty-two patients with GERD were randomized to either laparoscopic (LNF) or open (ONF) Nissen fundoplication. Outcome evaluation included reflux symptoms, gastrointestinal quality of life (GIQLI), and upper GI endoscopy. RESULTS: Esophagitis was cured among all patients in the LNF group and in 90% of the ONF group. There were two patients (10%) in both groups who had medicine-dependent recurrent reflux together with significant worsening in the GIQLI scores. One patient in the LNF group has been reoperated due to a suture granuloma in the left epigastric port. Two patients in the LNF group needed esophageal dilatation due to persistent dysphagia. GIQLI scores (scale, 0-144) were equally normalized in both groups. Overall, 90% in the LNF and 100% in the ONF group were either satisfied or very satisfied with the operation. There was only one patient (LNF) who would not choose to have the operation again. CONCLUSIONS: Laparoscopic and open Nissen fundoplication seem to be equally effective methods for improving reflux symptoms and quality of life, resulting in a high rate of satisfaction among patients with an intermediate follow-up period of 2 years. PMID- 11116410 TI - Laparoscopic antireflux surgery: comparative study of Nissen, Nissen-Rossetti, and Toupet fundoplication. Societe Francaise de Chirurgie Laparoscopique. AB - BACKGROUND: The aim of this retrospective study was to compare the results of Nissen, Nissen-Rossetti, and Toupet laparoscopic fundoplication in terms of gastroesophageal reflux disease (GERD). METHODS: From 1992 to 1996, 1,470 laparoscopic fundoplications were performed using one of three procedures: Nissen (n = 655), Nissen-Rossetti (n = 423), and Toupet (n = 392). Preoperative checkup included esophagogastroduodenoscopy in 1,437 patients (97. 7%), esophageal manometry in 934 patients (63.5%), and 24-h pH-metry in 799 patients (54.3%). The results were estimated at 1 month, 3 months, and 2 years. Patients unable to visit the hospital center were contacted by telephone. RESULTS: The three groups were quite similar regarding demographic data such as age, gender, preoperative clinical symptoms, and duration of GERD. One death (0.07%) occurred. At 3 months, there were no differences among the three groups concerning conversion, morbidity, dysphagia, early reintervention, or postoperative length of stay. The length of surgery was more important in the Toupet procedure. In the Nissen group, there were fewer Visick grade I patients but more Visick grade III patients. At 2 years, the recurrence and reintervention rates were similar. The overall residual severe dysphagia rate was 0.35% (n = 5). In the Nissen group, there were fewer Visick grade I patients but more in Visick grade II patients. There was no difference in Visick grade III and IV among the groups. More than 90% of the patients were satisfied (Visick I + Visick II), with no significant difference among the three groups. CONCLUSIONS: The results of this study do not differ significantly from the data reported in the literature, suggesting such surgical techniques are effective and well tolerated, and that both can be properly used in the treatment of GERD. PMID- 11116411 TI - A simple scoring system to reduce intraabdominal septic complications after laparoscopic appendectomy. AB - BACKGROUND: The development of intraabdominal abscess (IAA) following laparoscopic appendectomy (LA) is associated with significant morbidity. The aim of the present study was to validate an IAA risk score constructed from a previous review of 156 consecutive LA. METHODS: The score was tested in 250 subsequent consecutive LA and in patients with a positive risk score. Broad spectrum antibiotics were administered in order to avoid IAA. RESULTS: Factors related to IAA included clinically complicated appendicitis, leucocytosis >15,000/microl, a difference of >1 degrees C between axillary and rectal temperature, intraoperative findings such as (gangrenes and perforation), and intraoperative perforation of the appendix. In this series, broad-spectrum antibiotic therapy in patients with a positive IAA risk score reduced the incidence of IAA from 7.05% to 1.60%. CONCLUSION: This policy of identifying high risk patient via the scoring system and instituting subsequent antibiotic therapy in patients at risk reduces the incidence of IAA following LA. PMID- 11116412 TI - Laparoscopic vs open colectomy for sigmoid diverticulitis: a prospective comparative study in the elderly. AB - BACKGROUND: The aim of this prospective comparative study was to assess the outcome of laparoscopic and open colectomy for sigmoid diverticulitis in patients aged >/=75 years. METHODS: From January 1993 to December 1998, all patients 75 years of age and older undergoing an elective colectomy for sigmoid diverticulitis were included in the study. The patients were divided into the following two groups: group 1 (n = 22) consisted of patients who underwent a laparoscopic procedure; group 2 (n = 24) consisted of patients who underwent an open procedure. RESULTS: In group 1, there were 12 women and 10 men with a mean age of 77.2 years (range, 75-82); in group 2, there were 14 women and 10 men with a mean age of 78 years (range, 76-84) (p = 0.37). There was no difference between the groups in ASA classification. The operative time was shorter in group 2 (136 vs 234 mins). The postoperative period during which parenteral analgesics were required (5.4 vs 8.2 days, p = 0.001), postoperative morbidity (18% vs 50%, p = 0.02), postoperative length of hospital stay (13.1 vs 20.2 days, p = 0.003), and the inpatient rehabilitation (6 vs 15 patients, p = 0.01) were significantly shorter for group 1 than for group 2. There were no perioperative deaths. The conversion rate was 9% in group 1. CONCLUSION: The data from the present study suggest that laparoscopic colectomy for sigmoid diverticulitis can be applied safely to older patients with fewer complication, less pain, shorter hospital stay, and a more rapid return to preoperative activity levels than that seen with open colorectal resection. PMID- 11116413 TI - How to prevent port-site metastases in laparoscopic colorectal surgery. AB - Various reports concerning port-site metastasis after laparoscopic surgery for colorectal cancer have created a new concern regarding the use of this technique for the treatment of this malignancy. The real incidence is not yet known; neither are its prognostic implications. Numerous experimental studies, both in vitro and in vivo, have been published since 1994. These studies have analyzed the possible role of pneumoperitoneum and carbon dioxide (CO(2)) and pathophysiology, as well as the influence of minimally invasive techniques on tumor response and immunity. There are no definitive results yet, but there is enough evidence to presume that the etiology of this new complication might derive from surgical technique. We present our 8-year experience with laparoscopic surgery for colorectal cancer. We also review our technique for preventing port-site implants. At this writing, we have had no port metastasis in our series of 320 colorectal cancer cases with a mean follow-up period of 54 months. The steps we follow as a routine in all cases of laparoscopic colorectal cancer are (a) fixation of trocars to the abdominal wall, (b) avoidance of touching the tumor, (c) high vascular ligation, (d) intraoperative colonoscopy and intraluminal irrigation with 5% iodine povidone, (e) specimen isolation before extraction from the abdominal cavity, and (f) intraperitoneal and trocar site irrigation with a tumoricide solution. PMID- 11116414 TI - Intraperitoneal bile collections after laparoscopic cholecystectomy: causes, clinical presentation, diagnosis, and treatment. AB - BACKGROUND: Bile leakage is more common after laparoscopic cholecystectomy than after open surgery. In our department, the rate of postoperative bile collections after open surgery is 0.2% vs 0.6% after laparoscopic cholecystectomy. METHODS: We studied 13 cases of intraperitoneal bile collection without common bile duct damage drawn from a total of 5,200 laparoscopic cholecystectomies (0.23%). Clinical presentation, symptoms, method of diagnosis, causes, time of diagnosis, correlation of time of diagnosis with definitive treatment, and postoperative results were analyzed. RESULTS: The symptoms appeared between the 5th and 8th postoperative days. They were observed in patients with either chronic or acute cholecystitis. The main causes were misapplication of clips at the cystic duct and open Luschka's duct. Ultrasound failed for early recognition of bile collections. The definitive diagnosis was made by repeat ultrasonography, CAT scan, and ERCP. CONCLUSION: The ideal treatment in these cases is a minimally invasive procedure, but since the diagnosis is frequently delayed, open surgery is performed in the majority of patients. However, there were no mortalities in this group of patients. PMID- 11116415 TI - Is it safe to reuse disposable laparoscopic trocars? An in vitro testing. AB - BACKGROUND: The reuse of disposable laparoscopic instruments carries a risk of transmitting infectious diseases such as hepatitis and HIV. We evaluated the safety of reusing disposable trocars by studying the chances of their harboring infectious viruses after resterilization in an in vitro setting. METHODS: Disposable laparoscopic trocars were exposed to horse blood contaminated with high or low viral concentrations of herpes simplex virus type 1 (HSV1) and attenuated polio virus type 1 at room temperature for 2 h. HSV1 was chosen as the surrogate for lipid viruses that include hepatitis B and HIV virus; polio virus represented the nonlipid viruses that cause infections in immunocompromised patients and are more resistant to sterilization. The trocars were subsequently cleaned and resterilized by low-temperature steam and formaldehyde at 80 degrees C for 3 h. Viral cultures were then repeated after sterilization. RESULTS: A cytopathic effect (CPE) was demonstrated at both concentrations for HSV1 in all trocars before but not after sterilization. For the polio virus, CPE was evident in 50% of the trocars (two of four) exposed to high viral concentration after sterilization. CONCLUSION: Disposable trocars are difficult to resterilize and may harbor infectious viruses after their initial use. Therefore, the reuse of disposable trocars in laparoscopic surgery cannot be recommended. PMID- 11116416 TI - The driving force in trocar insertion: a comparison between disposable and reusable trocars. AB - BACKGROUND: Insertion of the first trocar during the "closed" technique of creating a pneumoperitoneum remains one of the most hazardous maneuvres in laparoscopic surgery, with complications such as major vascular and bowel injuries. The ease with which trocars are inserted through the abdominal wall may have some bearing on these complications. METHODS: A range of both disposable and reusable trocars, which were identical in point cross section and size, were compared in an abdominal wall model reconstructed with animal hide, using a hand held pressure transducer. Multiple insertions were performed, and the results were expressed in pounds per square inch (PSI). RESULTS: The disposable trocar tested required the least effort to insert (mean pressure, 2.76 PSI), followed by the new reusable (mean pressure, 3.42 PSI), with the used reusable trocar requiring the greatest force for insertion (mean pressure, 4.80 PSI). CONCLUSIONS: The effect of previous use on ease of insertion demonstrates an obvious disadvantage of reusable instruments. The excessive force required to insert some trocars may place the patient at greater risk of trocar injury. PMID- 11116417 TI - Choice of insufflating gas influences on wound metastasis. AB - BACKGROUND: Laparoscopic cancer surgery is limited by concerns about port-site metastasis. No study has definitively addressed the behavior and growth of tumor cells after the use of specific laparoscopic gases. METHODS: In athymic rats, 10,000 colon cancer cells were injected intraperitoneally. The rats received either no pneumoperitoneum (pneumo) or pneumo (8 mmHg, 10 min) with carbon dioxide (CO(2)), nitrous oxide (N(2)O), or air. Two full-thickness incisions were made and closed in the upper abdomen of each animal. After 4 weeks, implants were identified grossly at necropsy, and invasiveness was scored according to penetration through the layers of the abdominal wall. RESULTS: Rats receiving pneumo had more frequent implants (p < 0.01) with deeper penetration (p < 0.001) than rats not receiving pneumo. Implants were more common after air pneumo than after CO(2) (p < 0.05) or N(2)O (p = 0.07) pneumo, and were less penetrating after CO(2) pneumo than after air (p < 0.001) or N(2)O (p < 0.05) pneumo. CONCLUSIONS: Carbon dioxide gas may limit the viability and invasiveness of free intraperitoneal tumor cells, as compared with air or N(2)O. PMID- 11116418 TI - Laparoscopy: searching for the proper insufflation gas. AB - BACKGROUND: Although many aspects of laparoscopic surgery have been determined, the question of which insufflation gas is the best arises repeatedly. The aim of this study was to review the findings on the major gases used today in order to provide information and guidelines for the laparoscopic surgeon. METHODS: We reviewed the literature for clinical and laboratory studies on the currently used laparoscopic insufflation gases: carbon dioxide (CO(2)), nitrous oxide (N(2)O), helium (He), air, nitrogen (N(2)), and argon (Ar). The following parameters were evaluated: acid-base changes, hemodynamic and respiratory sequelae, hepatic and renal blood flow changes, increase in intracranial pressure, outcome of venous emboli, and port-site tumor growth. RESULTS: The major advantage of CO(2) is its rapid dissolution in the event of venous emboli. Hemodynamic and acid-base changes with CO(2) insufflation usually are mild and clinically negligible for most patients. Although N(2)O is advantageous for procedures requiring local/regional anesthesia, it does not suppress combustion. Findings show that Ar may have unwanted hemodynamic effects, especially on hepatic blood flow. There are almost no hemodynamic or acid-base sequelae with the use of He, air, and N(2), but they dissolve slowly and carry a potential risk of lethal venous emboli. CONCLUSIONS: Clearly, CO(2) maintains its role as the primary insufflation gas in laparoscopy, but N(2)O has a role in some cases of depressed pulmonary function or in local/regional anesthesia cases. Other gases have no significant advantage over CO(2) or N(2)O and should be used only in protocol studies. The relation of port-site metastasis to a specific type of gas requires further research. PMID- 11116419 TI - Effect of CO(2) pneumoperitoneum on early cellular markers of retinal ischemia in rabbits with alpha-chymotrypsin-induced glaucoma. AB - BACKGROUND: Increased intraperitoneal pressure in the head-down position is associated with a significant increase in intraocular pressure (IOP) in rabbits with alpha-chymotrypsin-induced glaucoma. Also, the retinal cells are weakened by the induction of increased IOP, and/or glaucoma, even when IOP is controlled by adequate therapy; therefore, these cells need to be protected from any additional aggression. Actin and vimentin are proteins of the retinal cell cytoskeleton that react readily in response to retinal injuries, including ischemia and glaucoma. Early changes in these cytoskeleton proteins determine the morphological changes observed after retinal damage. Therefore, we set out to investigate intracytoplasmic changes in vimentin and actin after a 4-h CO(2) pneumoperitoneum in the head-down position in rabbits with alpha-chymotrypsin-induced glaucoma. METHODS: Twenty-one rabbits with alpha-chymotrypsin-induced glaucoma in one eye received general anesthesia for 4 h in the head-down position and were randomly allocated to have (a) no pneumoperitoneum, (b) a 10 mmHg CO(2) pneumoperitoneum, or (c) a 20 mmHg CO(2) pneumoperitoneum. At the end of the trial, both the right glaucomatous and the left control eyes were enucleated and investigated immunocytochemically for alterations in vimentin and actin, and morphologically for retinal layer disorganization. RESULTS: Except for the preexisting morphological changes induced by glaucoma, both the control and the glaucomatous eyes in all rabbits appeared normal in terms of retinal layer organization and the distribution of intracellular vimentin and actin whatever the intraperitoneal pressure level applied. CONCLUSION: In rabbits with alpha-chymotrypsin-induced glaucoma, a 4-h CO(2) pneumoperitoneum of or = 0.8 compared to only 36% of the NOS fixed by rubber-band, face mask: 48%. FEO2 of > or = 0.8 was achieved with 52% of the hand-held NOS in 90 s, a time we consider practical for daily routine, whereas only 10% of the NOS fixed by rubberband and 14% of the face masks accomplished this threshold. A cooperative patient is an important condition when using the NOS: a strong premedication effect, absence of dentures, and patients who can not inspire via nose and expire via mouth involve impairment of the positive effects of the NOS. 21% of the anaesthesists felt disturbed by the NOS. 72% do not believe, that induction of anaesthesia will become more safe with the NOS. For 8 patients, breathing with the NOS was disagreeable (face-mask: 3 patients), 15 were disturbed by the nose part/mouth piece (face-mask: no patient). CONCLUSION: An acceptable FEO2 of > or = 0.8 can be achieved only without leakage of both the NOS and the face-mask. Therefore, routine FEO2 monitoring seems highly desirable. Efficiency of the hand-held NOS is much better than with the NOS fastened by rubberband or the face mask. However, even the hand held NOS cannot guarantee for optimal denitrogenization. Practicability in daily use was poor, because a test of airway patency by manual ventilation prior to relaxation/intubation is not possible with the NOS. Using the device as a help in apnoic oxygenation seems useful. PMID- 11116494 TI - [The treatment of severe head-brain injuries in Austria] . AB - INTRODUCTION: We performed this study in order to assess epidemiology and current practice of treatment of severe traumatic brain injury in Austria. Our survey followed the methods of a study published by J. Ghajar et al in the USA in 1995 and we compared the results to the Brain Trauma Foundation's "Guidelines For The Management Of Severe Head Injury". METHODS: The collected data represent answers to telephone interviews of 60 surgical intensive care units. We were able to evaluate data from all departments which treat severe brain traumas (Glasgow Coma Scale < or = 8) in Austria. RESULTS: At the time the treatment modalities of severe head injuries are not homogeneous and there are also big interdisciplinary management differences (trauma surgeons versus neurosurgeons). CONCLUSIONS: Results showed that there is a need for a brain trauma databank in Austria. We also recommend formation of an interdisciplinary brain trauma working group in order to control whether guidelines and standardized therapeutic modalities are being followed. PMID- 11116495 TI - [Impact of environmental factors on the incidence of posteropative nausea and vomiting. Influence of the weather and cycle of the moon]. AB - OBJECTIVE: In a survey concerning postoperative nausea and vomiting an unexpected high number of the participants stressed the impact of environmental factors, like weather and--even more surprising--the phase of the moon, on the occurrence of postoperative nausea and vomiting (PONV). Thus, the aim of this study was to determine the influence of these factors on the incidence of PONV. METHODS: On 203 days within the 19-month study period, 2488 patients were followed up for at least 24 hours postoperatively to determine the occurrence of PONV. For each day the actual incidence of PONV was compared with the mean predicted risk of PONV calculated with two risk scores for prediction of PONV (Koivuranta, 1997; Apfel, 1998). 32 days with the most significant difference between actual and predicted incidence of PONV were analysed retrospectively by two biometeorological experts, who were blind to the information whether each day was associated with a high or low incidence of PONV, evaluated the possible impact of the weather of these days. To analyse the influence of the cycle of the moon it was prospectively classified into four different phases. RESULTS: The two biometeorologists rated 22 out of the presented 32 days correctly. The likelihood p that this rating happened by chance is 0.0251, assuming that the likelihood for predicting each day correctly is 0.5 (independent Bernoulli-experiments, e.g. throwing a coin). However, days with a high or low incidence of PONV were equally distributed within the four phases of the moon (p = 0.97; chi 2-test with Yates' correction). CONCLUSION: Results from this analysis suggest that the weather may have some impact on the occurrence of PONV. However, our data do not support the hypothesis that the phases of the moon have any influence on this symptom. PMID- 11116496 TI - [Autologous transfusion--from euphoria to reason: clinical practice based on scientific knowledge.I]. PMID- 11116497 TI - Influence on the immune system of homologous blood transfusion and autologous blood donation: impact on the routine clinical practice/differences in oncological and non-tumour surgery? PMID- 11116498 TI - [Improvement of the coagulation potential in whole blood through leukocyte depletion]. PMID- 11116499 TI - [Preoperative autologous blood donation--is there really a net gain in red blood cells? The role of iron storage]. PMID- 11116500 TI - [Preoperative autlogous blood donation--is there really a net gain of red blood cells?--The role of blood donation strategy]. PMID- 11116501 TI - [Preoperative autologous blood donation: clinical epidemiologic viewpoint]. PMID- 11116502 TI - [Lethal vascular erosion after percutaneous dilatation tracheostomy]. AB - We report on a patient who underwent dilatational tracheostomy (Ciaglia technique) because of ARDS. 29 days after the procedure she died of hemorrhage from an arrosion of the bracheocephalic trunk, caused by the cuff of the tracheal cannula. This complication has, so far, been reported only after surgical tracheostomy. The fracture of tracheal cartilages is considered to be the specific cause of this fatal complication. The consequent loss of circular stability of the trachea demands increased cuff insufflation and pressure to tighten the airway. Prevention and therapy consist in control of the cuff pressure and caudal placement of the tracheal cannula. PMID- 11116503 TI - [The influence of airway pressure changes on intracranial pressure (ICP) and the blood flow velocity in the middle cerebral artery (VMCA). Ludwig HC et al. Anasthesiol Intensivmed Notfallmed Schmerzther 2000; 36: 141-145]. PMID- 11116504 TI - Surgery for stage 5 retinopathy of prematurity: the learning curve and evolving technique. AB - PURPOSE: To describe our experience with management of eyes with stage 5 retinopathy of prematurity (ROP). METHODS: Closed vitreoretinal surgery was done on 96 eyes of patients with stage 5 ROP. Lens was sacrificed in all but one eye. Surgery involved an attempt to clear all preretinal tissue and open the peripheral trough all round. In most instances bimanual surgery under viscoelastic was performed. RESULTS: At last follow up, anatomical success (defined as attached posterior pole) was achieved in 22.5% cases. Significant postoperative problems included reproliferation and secondary glaucoma. Only two infants obtained mobile vision. CONCLUSION: Late identification of disease, lack of prior treatment such as laser or cryo, and higher incidence of narrow-narrow funnel configuration were responsible for the poor surgical results noted in this series. The poor surgical and functional results reemphasize the need for prompt screening and management of infants at risk. PMID- 11116505 TI - The role of central corneal thickness in the diagnosis of glaucoma. AB - PURPOSE: To determine the effect of central corneal thickness (CCT) on applanation tonometry and any resultant misclassification of normals as ocular hypertension. METHOD: The central corneal thickness was measured using the ultrasound pachometer in 50 normals, 25 glaucoma and 23 ocular hypertensive patients. The student's "t" test was used to determine any significant difference in CCT between the three groups. RESULTS: There was a statistically significant difference in the mean CCT of the ocular hypertensives (0.574 +/- 0.033 mm) as compared to the glaucomas (0.534 +/- 0.030 mm) and normals (0.537 +/- 0.034 mm). Applying the described correction factor for corneal thickness, 39% of eyes with ocular hypertension were found to have a corrected IOP of 21 mmHg or less. CONCLUSIONS: Increased corneal thickness in ocular hypertension may lead to an overestimation of IOP in 39% of cases. Measurement of central corneal thickness is advisable when the clinical findings do not correlate with the applanation IOP. PMID- 11116506 TI - Fellow eye treatment in excimer photo refractive keratectomy. AB - PURPOSE: To describe symmetry of response in fellow eyes of patients undergoing photorefractive keratectomy (PRK) for myopia, analyse the risk factors leading to asymmetry in response and to determine if delayed treatment of the second eye increases safety and predictability of PRK. METHODS: Retrospective review of case records of 133 patients who underwent bilateral myopic PRK and had a minimum follow up of 6 months in both eyes. RESULTS: Postoperative uncorrected visual acuity, spherical equivalent (SE) refraction within +/- 1 D of emmetropia, best corrected visual acuity (BCVA) and corneal haze were not significantly different in fellow eyes of patients undergoing PRK for myopia. Of 87 eyes in group 1 (myopia < 6 D), 96.6% had uncorrected visual acuity > or = 6/12, 89.7% were within +/- 1 D of emmetropia, none lost > or = 1 line BCVA, and none had haze > or = grade 3. Similar results for 98 eyes in group II (myopia 6 to 9.9 D) were 75.6%, 55.1%, 2.0% and 2.0% respectively. For 81 eyes in group III (myopia > or = 10 D) the results were 42.7%, 33.3%, 8.6%, and 4.9% respectively. Among 84 patients with similar preoperative myopia in both eyes, 54 (64.3%) patients had a postoperative SE difference < or = 1 D in fellow eyes. Risk factors for asymmetric response among fellow eyes included increasing preoperative myopia (p < 0.001) and dissimilar treatment technique in the two eyes (p = 0.03). Corneal haze did not increase significantly after the third postoperative month. CONCLUSION: This study demonstrates that considerable symmetry of response exists in fellow eyes of patients undergoing myopic PRK. Early PRK in the fellow eye of patients with < 6 D myopia is safe and allows quick visual rehabilitation of the patient. In patients with myopia > or = 6 D, a 3-month interval before treating the second eye may improve the safety of the procedure. PMID- 11116507 TI - Intralenticular foreign bodies: report of eight cases and review of management. AB - PURPOSE: The management of intralenticular foreign bodies (ILFBs) with or without cataract has varied from time to time in the last century. We evaluated the surgical removal of the ILFBs with cataract extraction as a single-stage procedure. METHODS: Eight consecutive cases with intralenticular foreign bodies presenting to the trauma centre at our institute, were included in the study. Planned ILFB removal with cataract extraction and IOL implantation as a single stage procedure was done in all the patients. They were followed up from 2 months to 2 years after the surgery. RESULTS: ILFBs were removed with Kelman-Mcpherson forceps in seven cases and in one it was expressed with the nucleus during extra capsular cataract extraction. Co-existent posterior capsular tears were seen in two eyes, of which only one needed a localized vitrectomy. Posterior chamber intraocular lens implantation was possible without any complication in all the cases. Postoperative uveitis seen in three cases was easily controlled with periocular steroids. Best corrected visual acuity at last examination was 6/9 or better in 7 cases and 6/12 in one case with posterior capsular opacification. CONCLUSIONS: Timing and necessity of ILFB removal may be adjusted according to the foreign body characteristics and associated ocular trauma, choosing, as far as possible, the least traumatic procedure. Use of forceps rather than magnets is safer for the removal of the ILFB. Co-existent posterior capsular tears need to be anticipated and dealt with when encountered. PMID- 11116508 TI - Spectrum of aetiological agents of postoperative endophthalmitis and antibiotic susceptibility of bacterial isolates. AB - PURPOSE: To determine the spectrum of infectious agents of postoperative endophthalmitis, the relationship with the time of onset of symptoms after surgery and the antibiotic susceptibilities of the aerobic bacterial isolates. METHODS: A retrospective review of microbiological records from January 1995 to December 1998 yielded 173 isolates from intraocular specimen of 170 patients with culture-proven postoperative endophthalmitis. Antibiotic susceptibility of these isolates was determined for various ocular antibiotics using the Kirby-Bauer disk diffusion test. Based on the time of onset of illness, clinical presentation was classified into acute, delayed and chronic. RESULTS: Among 170 cases, 71 (41.7%) were attributable to gram-negative, 64 (37.6%) to gram-positive bacteria, and 37 (21.8%) to fungi. Gram-negative bacteria included P. aeruginosa (29;17.1%), other Pseudomonas spp (15;8.8%), non-fermenters (18;10.6%) and others (10;5.8%). Among these, 40 of 72 (55.5%) were sensitive to gentamicin, 47 of 72 (65.2%) to cefotaxime, 47 of 69 (68.1%) to amikacin, 52 of 71 (73.2%) to ciprofloxacin, and 25 of 40 (62.5%) to ceftazidime. The gram-positive bacteria included S. epidermidis (22;12.9%), S. aureus (13;7.6%), P. acnes (10;5.9%), Enterococcus spp (4;2.3%), Streptococcus spp (7;4.1%) and others (8;4.8%). Among these, 41 of 53 (77.3%) were sensitive to gentamicin, 47 of 53 (88.6%) to cefotaxime, 46 of 52 (88.4%) to ciprofloxacin, 38 of 41 (92.6%) to cefazolin and 27 of 37 (72.9%) to ceftazidime. All gram-positive bacteria were sensitive to vancomycin. CONCLUSION: In this large series of postoperative endophthalmitis, gram-negative bacilli followed by fungi accounted for the largest number of cases. A high degree of resistance of gram-negative bacilli to gentamicin, cefotaxime, amikacin and ceftazidime was recorded. PMID- 11116509 TI - Orbital abscess: management and outcome. AB - PURPOSE: To discuss the diagnosis, management and outcome of various types of orbital abscess. METHODS: The medical records of 13 patients diagnosed and treated for orbital abscess were reviewed. The sources of infection included: paranasal sinusitis (n = 5), odontogenic origin of infection (n = 4), one each, temporal fossa abscess, palatal abscess, furuncle on the nose, and secondary to retrobulbar injection of steroid. Computed tomographic scans revealed the presence of an abscess in all 13 cases. Associated findings on CT scan included: sinus disease (n = 8), cavernous sinus thrombosis (n = 2) and subdural empyema (n = 2). All patients were treated with intensive, multiple, intravenous antibiotics and early surgical drainage. RESULTS: Purulent material collected surgically from the orbit cultured Staphylococcus aureus (n = 3), two each Pseudomonas aeruginosa, Proteus mirabilis, Acinetobacter species and one each beta-haemolytic Streptococci, Citrobacter frundi and Enterobacter. Final visual acuity was good in 6 patients (6/12-6/6) and no light perception in 6 others. Visual acuity could not be recorded in the infant. The other complications were intracranial abscess (n = 4), cavernous sinus thrombosis (n = 2) and restricted ocular motility (n = 1). CONCLUSIONS: A high index of suspicion is necessary, along with early institution of appropriate diagnostic imaging, and aggressive medical and surgical treatment for a favourable outcome in cases of orbital abscess. PMID- 11116510 TI - Osseous choristoma of the choroid. AB - Osseous choristoma of the choroid, also called choroidal osteoma, is a very rare and unusual form of intraocular ossification. We report a case of choroidal osteoma in a young healthy male that to the best of our knowledge is the first report from India. PMID- 11116511 TI - Diffuse malignant melanoma of the choroid simulating metastatic tumour in the choroid. AB - We report a case of diffuse choroidal melanoma in a 51-year-old patient presenting as a yellow subretinal mass with secondary retinal detachment. This case highlights the diagnostic difficulty in such cases. PMID- 11116512 TI - Herpes simplex virus DNA in the lens one year after an episode of retinitis. AB - The present report describes a case where HSV was detected by polymerase chain reaction (PCR) in the lens cortical material removed during cataract surgery one year after resolution of retinal inflammation in a patient with ARN. PMID- 11116513 TI - Albendazole therapy for a recurrent orbital hydatid cyst. AB - Till recently, the treatment of a multiloculated hydatid cyst in the confines of the orbit was every ophthalmologist's nightmare. Over the last decade, two benzimidazole compounds, mebendazole and albendazole, have been tested clinically for use in the chemotherapy of hydatid disease. PMID- 11116514 TI - Towards achieving small-incision cataract surgery 99.8% of the time. AB - A surgical approach designed to reliably attain the modern goal of small incision cataract surgery 99.8% of the time is described. Phacoemulsification as well as a manual small incision technique is utilised to achieve the desired outcome as often as possible and for all types of cataracts. The logic, and required surgical steps are described and illustrated. This surgical technique allows the advantages of small incision surgery to be reliably achieved. The method is flexible and allows decisions and steps to be modified depending on the skill and comfort zone of the individual surgeon. PMID- 11116515 TI - Visual outcome following cataract surgery in rural punjab. AB - In a cluster sample survey in rural areas of Punjab visual outcome after cataract surgery was assessed. Three hundred patients (428 cataract operated eyes) were included in the study from 24 sampled villages. The mean age at cataract extraction was 61.70 +/- 9.82 years. The average interval since the cataract surgery was 7.05 +/- 5.86 years (range 0.11-32 years). Of the 428 operated eyes, 72 (16.82%) were blind (VA < 3/60), 162 (37.85%) had low visual acuity (VA 3/60-< 6/18) and 194 (45.33%) eyes gained good visual acuity (VA > or = 6/18). Cataract surgery related complications were the principal causes leading to blindness in 50 of 72 eyes; these included corneal oedema, (17/72;23.3%), retinal detachment (14/72;19.4%), and aphakic glaucoma (13/72;18.05%). This study emphasizes the need to improve the qualitative aspect of cataract surgery including long-term follow up in rural India. PMID- 11116516 TI - An outbreak of acute conjunctivitis caused by Coxsackie virus A 24. PMID- 11116517 TI - Incidence and management of posteriorly dislocated nuclear fragments following phacoemulsification. PMID- 11116518 TI - Significant impact of limbal epithelial stem cells. PMID- 11116519 TI - On the ways we do science. PMID- 11116520 TI - Limbal stem cell deficiency: concept, aetiology, clinical presentation, diagnosis and management. AB - Defects in renewal and repair of ocular surface as a result of limbal stem cell deficiency are now known to cause varying ocular surface morbidity including persistent photophobia, repeated and persistent surface breakdown and overt conjunctivalisation of the cornea. Ocular conditions with abnormalities of ocular surface repair include pterygium, limbal tumours, aniridia, severe scarring following burns, cicatricial pemphigoid and Stevens-Johnson Syndrome, sequelae of mustard gas exposure and Herpes simplex epithelial disease, radiation keratopathy, contact lens induced keratopathy, neuroparalytic keratitis and drug toxicity. Restoring ocular health in these eyes has traditionally been frustrating. An understanding of these intricate cell renewal and maintenance processes has spurred the evolution in recent years of new treatment methods for several blinding diseases of the anterior segment; many more exciting modalities are in the offing. However, there is inadequate awareness among ophthalmologists about the current principles of management of ocular surface disorders. The purpose of this article is to help elucidate the important principles and current treatment methods relevant to ocular surface disorders. PMID- 11116521 TI - Scientific literature and gospel truth. AB - We live in an age of science, an age in which science impacts practically every phase of our life. In the field of medicine, our entire understanding of diseases and their management depends on scientific knowledge. To obtain that knowledge, we rely on the published scientific literature. Therefore, the sanctity of science must be fiercely guarded. In medicine, true science leads to valid treatment--and preservation of the life, health and (for ophthalmologists) eyesight of our patients. A corrupted science results in corrupted scientific knowledge which in turn, in medicine, leads to wrong treatment and harm to the patients. PMID- 11116522 TI - Molecular signaling in pathogenicity and host recognition in smut fungi taking Karnal bunt as a model system. AB - Karnal bunt of wheat, incited by a phytopathogen Tilletia indica (Syn. Neovossia indica) is a floret infecting disease. In the floral tissues fungus proliferates and produces massive amount of black spores. In smut fungi, belonging to order Ustilaginales, communication between cells is necessary to regulate growth, differentiation and monokaryotic to dikaryotic transition during pathogenic and sexual development. Neighbouring cells are able to communicate with each other by direct cell to cell contact through plasma membrane bound signaling molecules or through formation of gap junctions and alternatively through secretion of chemical signals if cells are some distance away. Current research efforts toward understanding of pathogenic and sexual development in phytopathogenic fungi, offer a number of opportunities. These include the analysis of molecular signal(s) for direct contribution of sexual interactions to ability of smut and bunt pathogens to cause disease. These efforts will provide not only to explore the mechanisms of pathogenesis, but also to enhance knowledge of basic cellular biology of an economically important group of fungi. PMID- 11116523 TI - Animal experimentation: a rational approach towards drug development. AB - Man's observation of animals as objects of study undoubtedly began in prehistoric times. The first recorded attempt involving the use of live animals for research was by Ersistratis in Alexandria in 300 B.C. Animal investigation has clearly made possible the enormous advances in drug development in this century. A cursory review of any modern text book of pharmacology or medicine will attest the many drugs currently available to benefit mankind in the struggle to eradicate and control diseases. The main purpose of this article is to describe some of the experimental work on animals which contributed to the discovery and development of drugs benefiting human beings and other animal species. Since animal experimentation has occupied a focal position in all the research leading to useful drugs, one will appreciate that it will be necessary to limit the discussion to certain aspects of this broad and interesting topic. With this in mind, an attempt is made to relate briefly the nature of animal investigations which were instrumental in the development of major classes of drugs. Some attention has also been focused on legislation's on animal experimentation of some developed countries with emphasis on India and to views on animal experimentation. We hope this article will stimulate the minds of the scientists for a rational debate on the future of animal experimentation. PMID- 11116524 TI - Partial characterization of serum immunoglobulins of Oreochromis mossambicus. AB - Serum immunoglobulins of O. mossambicus were purified using chromatography methods--CM affinity gel blue chromatography followed by two step purification involving a combination of ion-exchange and gel filtration chromatography. Studies revealed that O. mossambicus produces only one class of high molecular weight macroglobulin as determined by molecular sieving by Sepharose CL 6-B. Immunoelectrophoresis of purified O. mossambicus serum against rabbit anti O. mossambicus serum gave only a single precipitin line. Further analysis of the immunoglobulin by SDS-PAGE showed that the IgM macroglobulin weighs about 900,000 Da, composed of mu-like heavy chain weighing about 90 kDa each and light chains weighing about 30 kDa each. PMID- 11116525 TI - Damage induction by direct electric current in tumoural target cells. AB - Damage induction to tumour target cells (P815) by direct electric current (DC) was investigated. A 6 min treatment of P815 cells with DC generated decreased levels of cell viability and proliferation. The ultrastructural analysis of DC treated cells revealed the presence of blebs, loss of cell surface filopodia, and ruptures in cell membrane. Mitochondrial alterations, swelling of cells, cytoplasmic matrix rarefaction, and cellular debri formation were also observed. The study shows that tumoural target cells can be damaged by direct electric current and this approach may provide means to understand the mechanism of tumour regression induced by electrochemical therapy. PMID- 11116526 TI - Endogenous strychnine, nicotine, and morphine--description of hypo and hyper strychninergic, nicotinergic and morphinergic state in relation to neuropsychiatric diseases. AB - Previous work from this laboratory had demonstrated the presence of endogenous morphine, strychnine and nicotine in the mammalian brain and human serum samples. Morphine is synthesised from tyrosine and strychnine and nicotine from tryptophan. This study examines the role of strychnine, nicotine and morphine in neuropsychiatric disorders. The blood levels of tyrosine, tryptophan, strychnine, nicotine and morphine were studied as also RBC membrane Na(+)-K+ ATPase activity. It was found that serum tyrosine levels were reduced and tryptophan levels elevated in all neuropsychiatric disorders studied with a reduction in RBC Na(+) K+ ATPase activity. Nicotine was present in significant amounts in serum of patients with schizophrenia, CNS glioma and syndrome X with multiple lacunar state. Morphine was present in significant amounts only in the serum of patients with multiple sclerosis and MDP. Strychnine was present in significant amounts in the serum of patients with epilepsy, Parkinson's disease and MDP. The presence of nicotine and strychnine in significant amounts could be related to elevated tryptophan levels suggesting the synthesis of these alkaloids from tryptophan. Morphine was not detected in most of the disorders owing to low tyrosine levels noted in them. Na(+)-K+ ATPase inhibition noticed in most of the disorders could be related to decreased hyperpolarising morphinergic transmission and increased depolarising nicotinergic and strychinergic transmission. The role of morphine, strychnine and nicotine in the pathogenesis of these disorders in the setting of membrane Na(+)-K+ ATPase inhibition is discussed. PMID- 11116527 TI - Regenerative ability of gastrocnemius muscle under diabetic condition with special reference to SDH & m-ATPase. AB - The present study analyzes the regeneration of skeletal muscle in streptozotocin (STZ) induced diabetic rats. Two weeks later the gastrocnemius muscle from diabetic rats were transplanted into diabetic and normal host to initiate regeneration and the normal gastrocnemius muscle was transplanted in normal and diabetic hosts for comparison. The regenerates were analyzed after 15 and 30 days of transplantation for histochemical (with respect to SDH and m-ATPase) and 7, 14, 21 and 28 days after transplantation for biochemical studies (with respect to SDH and m-ATPase). Least enzymatic activity and the poorest regenerative ability in case of normal muscle in diabetic host (NM-DbH) and comparatively higher enzymatic activity and better regenerative ability in diabetic muscle in normal host (DbM-NH) was observed. The result of this study strongly supports that the normal host environment is crucial for the muscle recovery. PMID- 11116528 TI - Effect of thyroidectomy on fast and slow muscle fibres of rat gastrocnemius muscle. AB - A combined histochemical, biochemical and electrophoretic study with respect to the enzymes succnic dehydrogenase(SDH), myofibrillar adenosine triphosphatase (m ATPase), lactate dehydrogenase (LDH) isozymes and myosin light chains was carried out to investigate the response of rat gastrocnemius muscle (medial head). Twelve weeks after thyroidectomy, the results indicated a shift from fast to slow type pattern of LDH isozymes, fibre type transformation from Type II to Type I and a decrease in SDH and m-ATPase activity. The results suggest, possible thyroidal involvement in determining the phenotypic properties of skeletal muscle. PMID- 11116529 TI - Evaluation of glutathione and ascorbic acid as suppressors of drug-induced lipid peroxidation. AB - In different sets of experiment lipid peroxidation induction capacity of two drugs, viz., ceftizoxime sodium, a third generation cephalosporin antibiotic, and acyclovir, an antiviral agent, was studied using goat whole blood as the lipid source. Ceftizoxime sodium caused significant extent of lipid peroxidation. Lipid peroxidation being a toxicity mediating process, such observation may be related to the toxic potential of the drug. Insignificant induction of lipid peroxidation was found in case of acyclovir and this is in good agreement with the safety record of the drug. Glutathione and ascorbic acid could significantly reduce ceftizoxime sodium induced lipid peroxidation, suggesting that free radical scavenging action of antioxidants may be exploited by possible antioxidant co therapy to reduce iatrogenicity of the drug in persons with impaired endogenous antioxidant defence. Glutathione and ascorbic acid appear to be promising candidates for further investigation in this regard. PMID- 11116530 TI - Inter-conversion of free sugars and accumulation of galactomannan in response to supplied metabolic mediators during pod filling in guar. AB - Detached inflorescences of guar (Cyamopsis tetragonoloba), each bearing 4 uniformly-developing pods at 42 days post anthesis (DPA), were cultured for 6 days in complete liquid medium manipulated with a fixed concentration of mannose and varying concentration of myo-inositol. Such inflorescences, but with 2 pods, were also maintained in the solutions of (i) glucose(U-14C) containing myo inositol or phytohormones, and (ii) mannose(U-14C) containing galactose for 36 hr. Effect of such exogenously supplied metabolic mediators on interconversion of free sugars in pod wall, endosperm and cotyledons and galactomannan accumulation in endosperm was studied. Myo-inositol decreased, over control, the relative proportion of invert sugars in pod wall, endosperm and cotyledons and at lower concentration (27.75 mM) it decreased the level of free sugars in pod wall and galactomannan in endosperm. In all pod tissues, 14C from both glucose and mannose got incorporated into myo-inositol as well as various sugars and maximum incorporation occurred in sucrose. High concentration of total free sugars and their 14C activity in pod wall indicated that this pod tissue was a potent accumulator of free sugars. With myoinositol, the relative proportion of 14C from glucose into raffinose sugars of pod wall and endosperm increased with a simultaneous decrease in this incorporation into galactomannan of the latter. Accompanying this, relative proportion of 14C into hexoses and myo-inositol decreased in pod tissues. Galactose increased 14C incorporation from mannose into total free sugars, sucrose and galactomannan with a concomitant decline in the labelling of hexoses. IAA and ABA enhanced 14C incorporation from glucose into total free sugars and this enhancement was much higher with IAA than ABA. The latter inhibited 14C incorporation into galactomannan. Based on these results, it was suggested that myo-inositol at lower concentration was inadequate to mediate the metabolism of sugars and, thereby, galactomannan synthesis. Galactose and mannose exhibited a mutual beneficial effect on their transportation to pods. Phytohormones stimulated the accumulation of sucrose in pod wall for its obligatory unloading into the seed. PMID- 11116531 TI - Microspore culture in Corchorus olitorius: effect of growth regulators, temperature and sucrose on callus formation. AB - Culture of isolated microspores and of anthers on media containing IAA directed free microspore development to an embryogenic pathway in C. olitorius. The first division of microspores on transfer to culture media was symmetrical in contrast to the asymmetrical division seen in normal development in vivo. Initially, 10 30% microspores divided symmetrically, but only 0.2-1% of the dividing microspores continued dividing and produced multicellular microcalli. About 30% of these microcalli produced callus but only on medium with 2.0 mg/L zeatin and 0.1 mg/L IAA. Incubation in the dark at temperatures of 35 degrees C for 1 day and then 25 degrees C was found effective for induction of first embryonic division in Corchorus. The frequency of microspore callus formation was higher on medium containing either 3% or 5% sucrose. Addition of colchicine and addition of activated charcoal to the above medium did not enhance microspore division in Corchorus olitorius. On transfer to different media most calli produced roots but regeneration of shoots and embryos was not induced. PMID- 11116532 TI - Influence of host plant on growth and reproduction of Aphis nerii and feeding and prey utilization of its predator Menochilus sexmaculatus. AB - Influence of different physiological stages (young, mature and senescent) of Calotropis gigantea leaves on growth and reproductive ability of A. nerii and feeding, prey utilization, fecundity and lipid content of its predator M. sexmaculatus were investigated. Increased reproductive period, total life span and reproduction of nymphs per female of A. nerii were observed when reared on mature leaves. This relative preference of A. nerii and maximum utilization of mature leaves as compared to other physiological aged leaves are mainly due to changes in the chemical composition such as protein, carbohydrate, lipid, amino acid, nitrogen and phenolic of C. gigantea. Further, aphids reared on mature leaves influenced its predator's (M. sexmaculatus) growth, prey utilization and reproductive performances. Fecundity and longevity were high, while developmental time of predator was shorter on mature leaves fed aphid. Maximum prey utilization and increased efficiency of ingested and digested food of predator was observed on mature leaves reared aphid. The results are interpreted and discussed in relation to plant aphid and predator interaction (tritrophic). PMID- 11116533 TI - Influence of dietary ginger (Zingiber officinales Rosc) on antioxidant defense system in rat: comparison with ascorbic acid. AB - Ginger (Z. officinale; 1% w/w) significantly lowered lipid peroxidation by maintaining the activities of the antioxidant enzymes--superoxide dismutase, catalase and glutathione peroxidase in rats. The blood glutathione content was significantly increased in ginger fed rats. Similar effects were also observed after natural antioxidant ascorbic acid (100 mg/kg, body wt) treatment. The results indicate that ginger is comparatively as effective as ascorbic acid as an antioxidant. PMID- 11116534 TI - Hypoglycemic, diuretic and hypocholesterolemic effect of winter cherry (Withania somnifera, Dunal) root. AB - Hypoglycemic, diuretic and hypocholesterolemic effects of roots of W. somnifera (ashvagandha) were assessed on human subjects. Six mild NIDDM subjects and six mild hypercholesterolemic subjects were treated with the powder of roots of W. somnifera for 30 days. Suitable parameters were studied in the blood and urine samples of the subjects along with dietary pattern before and at the end of treatment period. Decrease in blood glucose was comparable to that of an oral hypoglycemic drug. Significant increase in urine sodium, urine volume, significant decrease in serum cholesterol, triglycerides, LDL (low density lipoproteins) and VLDL (very low density lipoproteins) cholesterol were observed indicating that root of W. somnifera is a potential source of hypoglycemic, diuretic and hypocholesterolemic agents. Clinical observations revealed no adverse effects. PMID- 11116536 TI - Role of satellite RNA of an Indian isolate of cucumber mosaic virus in inducing lethal necrosis of tobacco plants. AB - Lethal necrosis or systemic stem necrosis followed by death of Nicotiana benthamiana, severe leaf deformations of N. tabacum cv. white burley and blister formations on N. tabacum cv. samsun NN symptoms were induced by experimental inoculations of CMV RNA preparations containing satellite RNA (sat-RNA). Inoculations of RNA preparations without sat-RNA did not induce that severe symptoms on these plants, only late mild mosaic was observed. It is suggested that sat-RNA of CMV isolate has a certain role for enhancing severity of symptoms in tobacco plants. Local and systemic lethal necrosis of N. benthamiana is due to sat-RNA present with genome of CMV isolate. It is the first report of lethal necrosis induced in N. benthamiana by CMV satellite. PMID- 11116535 TI - [TAME]-esterase--a new cardiovascular risk factor in smokers. AB - A mathematical model has been proposed to study the effect of [TAME] esterase on blood clotting time. Using this model, clotting time was found to decrease by 30% with increasing plasma [TAME] esterase activity in a group of smokers. It is hypothesized that [TAME] esterase through its effect on Hageman factor could affect clotting time. However mechanism of clotting by [TAME] esterase remains to be elucidated. It is concluded that [TAME] esterase is involved in the cascade of reactions leading to blood coagulation and increased [TAME] esterase activity could be an additional risk factor for possible cerebro-vascular accidents in smokers. PMID- 11116537 TI - Nitrosomethylurea induced streptomycin resistance in Lycopersicon esculentum Mill. AB - High frequency of streptomycin resistant variants of Lycopersicon esculentum were isolated on selective shoot regeneration medium supplemented with IAA (0.5 mg/L), zeatin (1.5 mg/L) and streptomycin sulphate (500 mg/L). Nonmutagenized (controls) and NMU treated cotyledons were placed on shoot regeneration medium supplemented with antibiotic streptomycin. Resistant shoots appeared at a high frequency in mutagenized cotyledons, whereas in controls morphogenesis was suppressed, accompanied by bleaching. Shoot regeneration occurred from the nodular tissues developed at the cut ends of cotyledons. Resistant shoots developed into complete plantlets on rooting medium containing selective concentration of antibiotic. Stability of streptomycin resistance was confirmed by leaf assay and reciprocal crosses between streptomycin-resistant and sensitive plants. PMID- 11116543 TI - [Drug combinations in the treatment of asthma. Are they really useful?]. PMID- 11116538 TI - Micropropagation of Sapindus mukorossi Gaertn. AB - Bud break and multiple shoots were induced in apical and axillary meristems derived from one month old seedlings of S. mukorossi on Murashige and Skoog (MS) medium supplemented with benzylamino purine (BAP) 0.4 microM or 0.8 microM alone. A combination of BAP and gibberellic acid (GA3) 0.4 microM and 2.8 microM produced elongated multiple shoots from both types of explants. Excised shoots were rooted on MS medium respectively with indole-3-butyric acid (IBA) 3.4 microM or 2.4 microM. The regenerated plantlets were successfully acclimatized and transferred to soil. PMID- 11116544 TI - [Situation in Spain of the diagnostic resources and of the treatment with continuous positive pressure respiration in sleep obstructive apnea-hypopnea syndrome]. AB - Sleep apnea-hypopnea syndrome (SAHS) is a major health problem whose estimated prevalence is 2 to 4% of the population. The Respiratory Insufficiency and Sleep Disorders Task Force estimated in 1995 that over one million people suffer SAHS with clinical repercussions in Spain while the number of patients receiving treatment was 8,000; diagnostic resources were not widely available. The aim of this study was to analyze the current situation in Spain. A survey carried out nation-wide in the period from 1995 to 1997 revealed that approximately 28,000 individuals were receiving nighttime support ventilation, signifying a prevalence of 72 per 100,000 inhabitants for this type of treatment. This situation has come about in a context of insufficient availability of diagnostic tools, with nocturnal oxygen levels having been established for some 37% of patients. We conclude that a broad plan to diagnose and treat patients with SAHS is required. The plan should include: a) greater availability of diagnostic tools; b) protocols for coordination; c) programs for continuous training and updating of knowledge of this disease, and d) revision of systems for financing support ventilation. PMID- 11116545 TI - [Development and results of a screening program for COPD in primary care. The PADOC Project(Program for the Increase in the Diagnosis of COPD in Primary Care]. AB - Several studies have shown that up to 75% of patients with chronic obstructive pulmonary disease (COPD) remain undiagnosed. Early diagnosis of such individuals will allow appropriate preventive and therapeutic measures to be prescribed. The PADOC project aimed to determine the efficacy of a COPD screening program for Spanish primary care settings. The participating primary care physicians were required to administer a spirometric test (forced expiratory volume) to all individuals who had not previously been given a diagnosis of COPD, who visited the clinic for any reason over a period of three months and who met the following enrollment criteria: a) age over 35 years and b) smoker of more than 10 cigarettes per day or ex-smoker of more than 10 packs per year. Individuals meeting the criteria for suspicion of COPD (FEV1 < 90% of predicted and FEV1/FVC < 70%) were referred to a pneumologist for confirmation of the diagnosis. One hundred ninety-four primary care physicians participated, administering 3,209 valid spirometric tests. The physicians identified 723 likely cases of COPD (22.5%). Pneumologists examined 278 patients (38.4% of the likely cases) and the final diagnosis was COPD in 153 of the 278 (55%) and asthma in 28 (10%). Therefore, 4.3% of all patients given spirometric tests received a diagnosis of COPD and 0.8% received a diagnosis of asthma. Agreement between spirometric measurements taken by the primary care physicians and those taken by pneumologists was low for FVC and FVC (%) (intra-class correlation coefficient ICC = 0.38 and 0.45, respectively) but good for FEV1 and FEV1 (%) (ICC = 0.78 and 0.67, respectively). We conclude that primary care screening for COPD is possible and would allow us to detect up to 22% of possible cases. Patient flow from one level of clinical care to another should be improved, given that most of the possible cases detected (61.6%) were not seen by the referral pneumologist. Agreement between spirometric measurements taken by primary care physicians and pneumologists was low for FVC but good for FEV1. PMID- 11116546 TI - [Residual pleural thickening in tuberculous pleuritis. Associated factors]]. AB - BACKGROUND: To study the factors related to the development of residual pleural thickening in pleural tuberculosis. PATIENTS AND METHODS: We studied 39 patients with tuberculous pleural effusion. A chest X-ray was taken of each patient at the end of treatment. The patients' medical histories, pleural fluid findings and diagnostic chest films were evaluated. Residual pleural thickening was defined as thickening that was visibly greater than 2 mm in the lower side portion of the chest film. RESULTS: Residual pleural thickening developed in 26% of patients and was found mainly in men (RR = 3.86). In no patients with Lowenstein-Jensen cultures positive for Mycobacterium tuberculosis did pleural complications develop. CONCLUSION: Residual pleural thickening is a common complication of tuberculous pleural effusion. Residual pleural thickening in tuberculous pleurisy occurs more often in men and older patients, and in cases in which pleural liquid culture is negative for M. tuberculosis. PMID- 11116547 TI - [Prognosis of T3N0M0 small-cell non-anaplastic bronchogenic carcinoma]. AB - We analyzed the survival after surgery for non-small cell lung cancer (NSCLC) classified as T3N0. Between January 1969 and 1995, 151 patients underwent surgery for NSCLC in our hospital. Survival analysis was performed using the Kaplan-Meier statistical method and the curves were compared using Mantel-Cox, Breslow and Tarone-Ware tests. The estimated five-year survival in the studied population was 44.46 +/- 4.30%. Four groups were defined based on degree of tumoral invasion of mediastinal structures, parietal pleura, chest wall or superior sulcus. Significant differences in five-year survival were observed between groups. Patients in the mediastinal group (59.98 +/- 8.71%) had the best prognosis, followed by patients with parietal pleura involvement (52.79 +/- 6.69%). Survival in the chest wall group was 27.53 +/- 7.22%. No patients with superior sulcus tumors survived over five years (median survival 1.50 +/- 1.16 years; 95% confidence interval 0.00 to 3.77 years). Prognosis is clearly determined by degree of tumoral invasion in T3N0 patients. In spite of the evident conceptual improvements achieved with the revised International Staging System, the system still fails to fully define prognosis in such cases. PMID- 11116548 TI - [Diagnostic errors related with tuberculosis in hospitalized patients]. AB - OBJECTIVE: To assess whether diagnostic error could have been avoided for patients who were mistakenly hospitalized for tuberculosis (TB) when in fact TB was not present or who were mistakenly hospitalized for another disease when they in fact had TB. METHODS: This cross-sectional, descriptive study examined the medical records of all patients admitted to the TB ward of Hospital San Juan de Dios (SJD) in Santa Cruz, Bolivia over a period of 28 consecutive months. RESULTS: Sixty-four (9.8%) of the 650 patients admitted were diagnosed incorrectly. Upon admission, all relevant information was recorded in the medical histories of 10 patients (15.6%) and at the physical examination of 28 patients (43.8%). Taking of a full medical history, performing a complete physical examination, and correctly interpreting the chest film would have led to correct diagnosis for 34 patients (53.2%) and would have been suggestive for 23 (35.9%) more. Such information plus the results of a sputum smear examination could have established or ruled out the presence of TB for 32 patients (50.0%) and suggested the appropriate diagnosis in another 27 patients (42%). CONCLUSION: The careful use of basic diagnostic tools could prevent serious clinical errors related to TB before such patients are admitted to a specialized ward. PMID- 11116549 TI - [Training the peripheral muscles in patients with COPD]. PMID- 11116550 TI - [Diagnosis and treatment of infectious exacerbation in cystic fibrosis]. PMID- 11116551 TI - [Dysfunction of the vocal cords simulating exercise-induced asthma]. AB - Vocal cord dysfunction is a respiratory condition characterized by anomalous adduction of the vocal cords during inspiration, causing significant air flow limitation in the larynx. Few such cases have been described in which dysfunction is triggered by exercise. We report the case of a young women with severe dyspnea appearing as a result of physical activity. We first deal with issues of differential diagnosis in relation to several other diseases, particularly exercise-induced asthma and then discuss therapeutic approaches. PMID- 11116552 TI - [Bronchiolitis obliterans in a patient previously working as a printer in a textile factory]. AB - We report the case of a 24-year-old man with a diagnosis of bronchiolitis obliterans, a rare clinical condition; the similarity to Ardystil syndrome was striking. Relevant occupational history included work in a textile air-brushing factory. Also noteworthy were lesions observed by CT scan in the form of cystic formations measuring less than 1 cm, a finding not previously described in the context of bronchiolitis obliterans. The patient improved immediately after starting corticoid treatment although scans failed to improve over several months of follow-up. PMID- 11116553 TI - [Catamenial hemoptysis. Report of a case]. PMID- 11116554 TI - [Small-cell bronchial carcinoma with secondary hypertrophic osteoarthropathy]. PMID- 11116555 TI - [Persistent ductus arteriosus as a cause of pulmonary inoperability]. PMID- 11116569 TI - Behavioral state-dependent thermal feedback influencing the hypothalamic thermostat. AB - The experimental evidence on the behavioral state-dependent compartmentalization of temperature in the central nervous system of three homeothermic species has been reviewed to address the question of how selective brain cooling influences hypothalamic temperature regulation. PMID- 11116570 TI - Auditory deprivation modifies biological rhythms in the golden hamster. AB - To assess to what extent auditory sensory deprivation affects biological rhythmicity, sleep/wakefulness cycle and 24 h rhythm in locomotor activity were examined in golden hamsters after bilateral cochlear lesion. An increase in total sleep time as well as a decrease in wakefulness (W) were associated to an augmented number of W episodes, as well as of slow wave sleep (SWS) and paradoxical sleep (PS) episodes in deaf hamsters. The number of episodes of the three behavioural states and the percent duration of W and SWS increased significantly during the light phase of daily photoperiod only. Lower amplitudes of locomotor activity rhythm and a different phase angle as far as light off were found in deaf hamsters kept either under light-dark photoperiod or in constant darkness. Period of locomotor activity remained unchanged after cochlear lesions. The results indicate that auditory deprivation disturbs photic synchronization of rhythms with little effect on the clock timing mechanism itself. PMID- 11116571 TI - Input-output relations of Deiters' lateral vestibulospinal neurons with different structures of the brain. AB - It has been the goal of this review to describe the functional interrelations between Deiters' vestibular nucleus and numerous brain structures. Emphasis is placed on dynamic and integrative properties of linkages between the neurons of Deiters' nucleus and many other brain structures in order to begin considering the capabilities of the loops in the light of motor control and coordination of movement. The problem of somatotopy within the loops is also considered. Putting this information together, the possible roles of Deiters' nucleus in the control of movements are described. It is suggested that Deiters' nucleus in co-operation with cerebral cortex, cerebellum, subcortical and brainstem structures are responsible for the integration and realization of different movements. PMID- 11116586 TI - Management of clusters of sexually transmitted infection. PMID- 11116587 TI - Surveillance of influenza and other respiratory infections. PMID- 11116592 TI - Management of building services. Procurement for highly|| serviced healthcare facilities. AB - The question of whether design and build is the best||| procurement system is hard to tell as other procurement alternatives||| (main-contractor and principal services contractor coupled with project||| management team, and construct management method plus good building team) can||| also give satisfactory results. Besides, it is also known that there is no||| standard solution or best buy amongst procurement systems. Indeed, there is no||| single method of procurement which can be suitable for every project for all||| time. It is also accepted that the choice and use of an inappropriate||| procurement system is not the only reason for inefficient project management,||| people and experience are far more important than strategy. Coordination of||| building services has a direct bearing on the success of a hospital project.||| Services must be fully integrated and coordinated. One thing is clear, as far||| as building services coordination is concerned, the traditional procurement||| system is not conducive to effect coordination process, however, the integrated||| nature of D&B has relieved all the burdens on the client's design team,||| and, from the face of this procurement path, all coordination problems that the||| client would normally encounter from the traditional method will no longer||| exist. Of course, the technical and managerial problems in relation to||| coordination still exist but they would all be passed to the contractor side.||| To protect the contractor's interest, all coordination problems would still||| have to be managed and controlled by D&B contractor and his specialist||| contractors expeditiously. Indeed, if design is not fully integrated, and||| construction and services are not completely coordinated, no matter how||| integrated is the team, D&B method will not work. To sum up, the author's||| discussions can be summarized as follows: Project success can be enhanced by||| engendering a team spirit and a high degree of cooperation between project||| participants. This can be achieved by the use of non-traditional procurement||| systems. Building services and hospital engineering systems must be properly||| coordinated at both design and installation stages. Coordination must be seen||| as both technical and managerial issues. A professional building team can||| manage the complex issues of services coordination. But it is known that an||| integrated building team can give better result. D&B is not a panacea,||| other procurement paths with adequate project management can provide a better||| alternative than either the traditional or the design and build||| system. PMID- 11116598 TI - Pneumatic tube systems--key to relieving the|| pressure. PMID- 11116599 TI - [Epidemiology of rubella in France, 20 years after the advent of vaccination]. PMID- 11116601 TI - [The tobacco mosaic virus from Martinus Beijerinck to James Watson]. PMID- 11116602 TI - [Valvular heart disease in adults: forward]. PMID- 11116603 TI - [Epidemiology and etiology of acquired heart valve diseases in adults]. AB - Acquired valvular heart disease remains common. Their causes have changed during the past 30 years with the increase in life expectancy and with the decrease in the incidence of rheumatic fever. Calcific aortic stenosis is currently the predominant cause, followed by left heart valvular regurgitations due to degenerative disease and infective endocarditis. Post-radiation valvular disease might become more frequent in the future. More recently, left heart valvular diseases were observed in the United States in patients treated by anorexigen treatment. PMID- 11116604 TI - [Echocardiography of heart valve diseases]. AB - Echography can be considered as the gold standard method for quantifying a valvular dysfunction, evaluating its hemodynamic impact and assessing anatomy. Echocardiography is a major step in the diagnosis and the pretherapeutic evaluation of valvular disease. PMID- 11116605 TI - [Aortic stenosis in the elderly]. AB - Aortic stenosis is observed with an increasing frequency in the elderly. Main features in the elderly are the frequency of advanced stage of the heart disease, and of associated diseases, either cardiac, in particular coronary lesions, or extracardiac. This can explain that perioperative morbidity and mortality are higher than in younger patients. However, after the operative period, long-term prognosis is good. Life expectancy becomes similar to a general population and functional results are excellent. In case of symptomatic aortic stenosis in the elderly, the decision to operate should take into account an evaluation of the individual risk to benefit ratio, and avoid to differ the decision until an advanced stage of the disease. PMID- 11116606 TI - [Dystrophic aortic insufficiency]. AB - Dystrophic aortic regurgitation is the result of 2 diseases: (1) aortic regurgitation, consequence of (2) aortic dilatation due to decreased aortic wall resistance. Marfan syndrome, which is a genetic disease, should be looked for systematically, with the help of an ophthalmologist and a rheumatologist. Aortic dilation is responsible for the increased mortality because of aortic dissection. Diagnosis is often made when the aorta is dilated wheras the aortic regurgitation is minimal or moderate; when the patient is asymptomatic. This has 2 consequences: siblings of Marfan patient should be examined by echocardiography; surgical replacement of the ascending aorta is often performed because of the aortic dilation, not because of the aortic regurgitation. PMID- 11116607 TI - [Infectious endocarditis]. AB - The incidence of infective endocarditis has not decreased over the last years. Infective endocarditis remains severe: the mortality rate during the initial hospital stay is 15%, and a surgical intervention is needed in 25% of the cases. Stricter application of prophylaxis guidelines and better diagnostic and therapeutic management should decrease the frequency and severity of infective endocarditis. PMID- 11116608 TI - [Mitral valve insufficiency and surgical repair]. AB - Management of mitral regurgitation have benefited over the last 10 years from the better understanding of its natural history and from the advent of new echocardiographic quantitative methods. The dismal prognosis displayed by patients with flail leaflet and severe mitral regurgitation medically treated in one hand and the demonstration of the dramatic consequences of impaired pre operative left ventricular function in the other, have been a strong incentive for early surgical correction of the disease. In the same time, mitral valve repair developed because of the improvement in the surgical techniques, of changes in aetiology and because of the widespread use of intra-operative transoesophageal echocardiography. Mitral repair has been shown to be an independent and beneficial predictor of overall survival, operative mortality and late survival and consequently became the support of early surgical strategies. But it is not the only factor to predict mortality and morbidity, and one must not forget the decisive and independent part played by age, preoperative symptoms and above all pre-operative left ventricular function. Therefore, mitral valve repair must not be considered as a pretext to postpone intervention but should be an other reason to intervene earlier. PMID- 11116609 TI - [Valvular stenosis: treatment by percutaneous dilatation]. AB - For more than a decade, cardiologists have now used procedures for treating valvular stenosis. The most widely used, percutaneous mitral commissurotomy, is performed through the transseptal pathway, usually with the Inoue balloon. Subsequently, the valvular surface doubles and the incidence of serious complications is low in experienced teams. Evaluation of ten years of experience shows that long-term results are good if the dilatation was efficient immediately. In the other cases, surgery must be performed. Percutaneous mitral commissurotomy is the preferred solution in young patients with good valvular anatomy; in others, its indication should be discussed on a case-by-case basis. It is contraindicated in the presence of thrombosis of the left auricle or severe valvular calcification. The results of aortic valvuloplasty for calcified stenosis in the elderly subject are poor, with elevated mortality and morbidity. Valvuloplasty little affects the natural history of a tight aortic stenosis but can offer transitory functional improvement for several months. Its indications are rare. PMID- 11116610 TI - [Valvular prostheses and their follow-up]. AB - In use since 1961, valvular prostheses allow the correction of the severe valvular diseases when conservative procedures are not possible. Current prostheses have outstanding haemodynamic features. Mechanical prostheses have a supposedly unlimited life span but require anticoagulant treatment. Bioprostheses do not need such treatment but end up to deteriorate and need reoperation. Both can be affected by valve-related complications (thromboembolic events, endocarditis deterioration, desinsertion...): an ideal prosthesis does not exist yet. All patients with valvular prosthesis require close follow-up, where echocardiography holds proeminent place. PMID- 11116611 TI - [The Flexner Report]. PMID- 11116612 TI - [Acute hearing loss. Diagnostic approach]. PMID- 11116613 TI - [Hyperadrenocorticism in the adult. Etiology, diagnosis]. PMID- 11116614 TI - [Secondary amenorrhea. Diagnostic approach]. PMID- 11116615 TI - [Ankle sprains. Diagnosis, emergency guidelines]. PMID- 11116616 TI - [Prenatal diagnosis of genetic diseases. Indications, methods, legal and ethical aspects]. PMID- 11116617 TI - [Neonatal screening for phenylketonuria and hypothyroidism]. PMID- 11116618 TI - [Mediastinal adenopathies and mediastinal tumors. Diagnostic approach]. PMID- 11116639 TI - Alarm bells. HCFA changes ring alarm bells for EMS agencies|| nationwide PMID- 11116640 TI - Fundamentals of leadership. PMID- 11116641 TI - Preventing sexual harassment in EMS. PMID- 11116642 TI - Physicians under scrutiny for fraudulent billing for "gang" visits. PMID- 11116643 TI - Monitoring fiscal intermediary payment performance. PMID- 11116644 TI - Managed care in flux: strategies for the current state. AB - Given the combination of consumer backlash, a rash of antimanaged care bills, renewed medical inflation, rising prescription drug costs and the threat of class action HMO lawsuits, managed care is in a state of flux. However, managed care in some form is here to stay. This author recommends that plan sponsors adopt strategies to reap the positives managed care offers while the industry is reshaping itself. PMID- 11116645 TI - Delivering health care benefits: staying focused on real solutions. AB - All signs point to an explosion of new approaches to financing, delivering, managing and purchasing health care. However, before employers abandon the current model, they need to verify that the alternatives will be truly advantageous and that new approaches will not place employees at greater risk than they are today. A through assessment of the advantages and risks of various approaches will equip employers to make the best business decisions and to be articulate participants in the coming debate. PMID- 11116646 TI - Self-funding health insurance for small employers: is it the right way to go? AB - Self-funding offers small employers an alternative approach to providing health care benefits for their employees. It may not be appropriate for every small employer but, in situations where it is feasible, self-funding offers employers a way to get a measure of control over health care costs while providing the benefits employees expect. PMID- 11116647 TI - Exclusions of infertility treatment in the aftermath of Bragdon v. Abbott. AB - This article analyzes the Supreme Court's decision in Bragdon v. Abbott to determine whether the Court's reading of ADA necessarily leads to the conclusion that a group health plan's or insurer's exclusion or limitation of coverage with respect to infertility is prohibited by ADA. The authors conclude that it may be advisable for plans to perform at least minimal actuarial calculations with respect to the current or anticipated costs of covering infertility treatments prior to adopting any exclusions or limitations of coverage. PMID- 11116648 TI - Small business no longer has to mean small benefits. AB - The Internet has drastically changed the benefits environment for small businesses. Formerly, employers with a small workforce were severely limited in providing benefits to their employees. Today, by utilizing the Web, many owners of small businesses are offering their employees benefit packages comparable to those offered at major corporations. This author offers suggestions for effective benefit shopping on the Internet. PMID- 11116649 TI - New study challenges the value of reducing patients' length of stay. PMID- 11116650 TI - OPPS begins with shaky start and mixed reviews. PMID- 11116651 TI - JCAHO considers new standards to reduce errors. PMID- 11116652 TI - Do patients suffer if social work role is diluted? PMID- 11116653 TI - Take off the blinders, social workers advised. PMID- 11116654 TI - Peer review: complying with JCAHO's standards. Part 1. PMID- 11116656 TI - Debate continues: is pharmacist shortage creating risks? PMID- 11116655 TI - How to win, hold support for measurement initiatives. PMID- 11116657 TI - University study identifies problems with IOM report. PMID- 11116658 TI - Countdown begins for compliance with HIPAA electronic standards. PMID- 11116659 TI - Capitation: how well is your back covered? PMID- 11116660 TI - 'New' Aetna and Kaiser face future. PMID- 11116661 TI - Mergers, acquisitions afoot in disease management industry. PMID- 11116662 TI - Predicting MCOs' future by learning from the past. Interview by Patrick Mullen. PMID- 11116663 TI - Pneumococcal conjugate vaccine for young children. AB - Pneumococcal disease is a common cause of morbidity and mortality in the pediatric population. Pneumococcal infections, which account for most serious bacterial disease in infancy and early childhood, are a major cause of acute otitis media, sinusitis, pneumonia, bacterial meningitis, and bacteremia. Streptococcus pneumoniae is the causative agent in a large percentage of these infections, although other microorganisms also play a role. The recent emergence of drug-resistant strains has provided a strong incentive for preventing pneumococcal infections by vaccination. However, the capsular polysaccharide pneumococcal vaccines used to immunize adults are neither immunogenic nor protective in young children due to poor antibody responses. Therefore, research has focused on development of additional immunogenic pneumococcal vaccines to provide long-term immunity in children < 2 years of age. The most promising approach has been the development of a protein-polysaccharide conjugate vaccine for the seven serotypes (4, 6B, 9V, 14, 18C, 19F, and 23F) that most commonly cause infections in childhood. An effective conjugate vaccine that protects against these serotypes has the potential to prevent 85 percent of bacteremia episodes, 83 percent of meningitis episodes, and 65 percent of otitis media cases in the U.S. among children younger than 6 years. The Food and Drug Administration (FDA) recently approved the first protein-polysaccharide conjugate vaccine to prevent invasive pneumococcal diseases in infants and toddlers < 2 years of age. This conjugated vaccine against pneumococcus uses the same technology as the successful vaccine against Haemophilus influenzae type b. It consists of an immunogenic but inert protein coupled covalently to the polysaccharide coat of the selected strains of pneumococci. The conjugated antigen induces a more powerful, T-cell-based immune response in infants, which is developed by the time they are 2 months of age. Some important questions regarding this vaccine for children < 2 years of age: Is the vaccine safe? Is it immunogenic? Is it efficacious in preventing invasive pneumococcal disease and controlling otitis media? FINDINGS: Results of three randomized double-blind trials designed to evaluate the safety and immunogenicity of this vaccine in healthy children < 2 years of age were reported within the last three years. The studies found that the vaccine is safe and highly immunogenic for all seven serotypes. The most recent study, involving over 37,000 young children, also evaluated the vaccine's efficacy, and reported that the vaccine is highly effective in preventing invasive disease and has had an impact on otitis media. CONCLUSIONS: The heptavalent pneumococcal conjugate vaccine is safe and highly effective in preventing pneumococcal meningitis and bacteremic pneumonia in young children < 2 years of age; it is less effective in preventing otitis media. Based on the results of three well-designed studies demonstrating the vaccine's safety, immunogenicity, and efficacy, the vaccine is safe and effective for active immunization of children < 2 years of age against invasive disease caused by seven Streptococcus pneumoniae serotypes included in the vaccine. At this time, there is no clear medical consensus regarding its safety and efficacy for control of otitis media in children < 2 years of age. This application has not been evaluated by the FDA. The pneumococcal conjugate vaccine should be considered experimental, and has not been shown to be safe or efficacious for Streptococcus pneumoniae disease other than that caused by the serotypes included in the vaccine and for invasive infection, such as bacteremia or meningitis, caused by other microorganisms. PMID- 11116664 TI - Women's health: is it the conception or the delivery? PMID- 11116665 TI - Any way you cut it, employers appear to save if Medicare adopts drug benefit. PMID- 11116666 TI - Changing work systems. AB - This paper is concerned with finding practical ways of incorporating human and organizational concerns during the development and use of new information technologies (IT). It is structured in four parts. First, we outline the work of the Institute of Work Psychology at the University of Sheffield. One major interest is with the human and organizational aspects of the new information technologies. We are especially interested in work organization and job design, the allocation of tasks between and among humans and computers, the roles of users, the roles of senior managers, the management of change, and the performance of new investments in information technology (IT). Second, we review the evidence from survey work and from detailed case studies concerning the performance of IT. This reveals that most IT investments do not meet their performance objectives, and that the reasons for this are rarely purely technical in origin. Change is too often technology-led, and too little attention is paid to human and organizational factors. Most companies fail to consider how work should be organized and how jobs should be designed to make the new technologies more effective. Usually, users have no substantial influence on system development. Senior managers are criticized for their lack of understanding and action in these areas. Third, we consider the potential in the United Kingdom for changing work systems incorporating new technology. One way forward is through the development and use of sets of theoretically derived tools that can be used by managers and others. To this end we are working with several collaborators on a portfolio of tools in the following areas: organizational design job design allocation of tasks between and among humans and computers. We then review our approach and offer views of the areas and ways in which this work could develop, including opportunities for international collaboration. PMID- 11116667 TI - Sociocultural dilemma of Japanese steeplejacks. AB - Japanese steeplejacks are good at working in high places as construction workers, and they have been called tobi for a longtime. They now play an important role in completing modern civil engineering projects and in the construction of high-rise buildings; however, their lifestyle is considered by most to be quaint but outdated. Originally, they were unskilled workmen at construction sites. In the 18th century, they were engaged in repairing houses or setting up scaffolding, helping carpenters, but they worked as firefighters whenever fires broke out. Their traditional work system did not change throughout the Meiji era, although Japanese society became greatly modernized. After World War II, the industrialization of Japanese society required highly developed technology in civil engineering and architecture. This provided an opportunity for them to establish their positions as trained professional workers. However, the number of skilled tobi professionals has continued to decrease because the younger generation does not consider this profession desirable career. Improving not only the professional skills but also the way of living to the extent as a modern high tech society demands will be the key for the tobi's work system to become attractive. PMID- 11116668 TI - Ergonomic problems of migrant workers in Australia. PMID- 11116669 TI - Sharing positive experiences in making changes in work and life in a local district in Vietnam. AB - A series of research and training programmes were undertaken in a rural district in the Mekong Delta area in Vietnam so as to facilitate positive changes in the work and life under the initiatives of the local people. As in other parts of the area, it was amid rapid changes in life-style under the shift toward a market economy. Field research relying on direct observation methods were carried out in various rice field jobs and local small enterprises for understanding the improvement potentials in the work and lives of the local people. To strengthen the local improvement initiatives, participatory training programmes were developed and practiced. In the training programmes, joint walk-through observations using an action checklist were stressed. The results were discussed in small groups and meetings. Interactive communications between the local people, while visiting their own houses and workplaces, took place. Based on these field research results, various positive changes were successfully implemented. They included a wide range of improvements such as more efficient work methods, safer use of technologies, hygienic housing conditions, and improved household plans. Our experiences demonstrated that practical approaches relying on local initiatives could facilitate the implementation of positive changes in the local socio-cultural settings. PMID- 11116670 TI - Ergonomics studies in Thailand: a future prospect. AB - Ergonomics studies in Thailand began in 1967 after Professor Kovit Satavuthi graduated from the University of Birmingham, where he worked with Professor Nigel Corlett. Ergonomics was offered as an elective in the IE curriculum at Chulalongkorn University. Professor Satavuthi was later invited to teach manufacturing process and system at the Department of Occupational Health and Safety, School of Public Health, Mahidol University. He then added the ergonomics concept to the class as part of his lectures. The first international conference was organized by the Central Coordinating Board of SEAMEO-TROPMED Project, and it was held in Bangkok from 19-23 June 1972, thus bringing ergonomics into public attention. Most participants came from medical science and public health fields, however. Ergonomics popularity has begun to gain momentum in the past 4-5 years. This could be the result of more universities paying serious consideration to provide ergonomics knowledge to their students. It is also well-accepted that ergonomics is a multidisciplined subject. It requires knowledge from medical sciences, sociology, psychology, public health, and engineering. The results of ergonomics studies will have greater chances of success if they are concluded from all disciplines of knowledge. This paper reports some previous studies in Thailand and discuss how to build up an ergonomics team and to produce reasonable results with a sound application plan. PMID- 11116671 TI - Ergological viewpoint in real-life study. AB - I think that real-life studies on human work must be done, if possible, not objectively, but subjectively from an ergological viewpoint. Essentially, I consider ergology as the biology of human living and work, along German biologist Haeckel's idea. He considered ergology a field of anthropology. The instinctive (genetic) element and the learned (environmental) element constitute human behavior. It is conceivable that the adaptation by human learnability has today brought about high-tech society, and genetic human attributes (human nature) have been adapted to the living environment of hunters and gatherers, which has been long-lasting in human history. Hunters-gatherers form a band consisting of about 50 (30-100) persons and move in a more or less vaguely defined territory (1,000 3,000 km2). The unsuitableness of human nature for modern scientific civilization has caused various problems. Always keeping human nature in mind when considering human living and work is the philosophical background of human ergology. Two real life working conditions in agriculture, the packing of leeks and promoting a system for securing employees in strawberry production, are presented. PMID- 11116672 TI - Observation of human behavior as a field study method. PMID- 11116673 TI - Aging and efficacy of work: a methodological discussion. AB - Based on a comparison of various human work systems in different cultures, the relationship between efficacy of work and aging is highlighted as a basic component. Age-dependent and age-independent individual variations in labor efficiency in subsistence or nonindustrial societies, which were disclosed in the author's field investigations, are compared with those in industrial societies. Human ergological implications for industrial workers' retirements because of their contracts are also discussed in relation to the behaviorally similar but freely organized work patterns in subsistence societies. PMID- 11116674 TI - An approach to human work systems development under the circumstances of an aging society and international business operations. AB - At least three serious aspects of problems exist as obstacles for the national economy in Japan to grow or even to maintain its present level: the lack of natural resources, the trends of a decreasing young labor force, and an increase in the shift of domestic business operations to foreign countries. Although top managers make the decisions of product or service planning, or both, work systems designers are also responsible for conserving the resources. An action against the decrease of the young work force is needed to maintain work systems in an operable condition. The business shifts to foreign countries affect all the people, who are losing job opportunities. The present paper presents an approach to reorienting human work systems within the scope of the work systems designers' roles under the circumstance of these social environments. The following discussion is based on the assumptions that work organizations be productive for themselves and the world, effective and efficient for themselves, and contributive to their communities and the world. In essence, an approach to human work systems development should be fair to managers and workers alike. Presented are cases of these work systems as developed along the perspectives mentioned. PMID- 11116675 TI - The change of productive skills and human work systems--function of work concept. AB - The techniques and skills of the age are changing, but the work situation is not corresponding to these changes. Furthermore, this causes problems in human resources development. We have investigated the contents of productive skills, vocational abilities, and work conditions, and as a result, productive skills were divided into two patterns: intellectual-management skills and sensory-motor skills. The contents of intellectual-management skills are divided into measurement, examination, design and arrangement, analysis and judgment, and understanding of technical knowledge. The contents of sensory-motor skills are divided into vision, audition and touch, visual judgment, motion of limbs, judgment by hands, and quality of the product. A sensory-motor skill is defined as one that mainly depends on judgment and memory capabilities. The former includes skills related to traditional handicrafts, processing assembling, and machine operation. The latter includes skills related to programming, designing, and controlling. A connection deeply related to the human being's essence can be found in both skills. When we examine human work systems, it is important to consider this essence. PMID- 11116676 TI - Why do older workers have problems with technology? AB - Why do older workers seem to have problems with technology? In this paper, I will review several possible reasons and illustrate them with evidence, often anecdotal, from our work as ergonomics practitioners. We find that older workers have more to "unlearn" from their accumulated experience. They may suffer from gradual or not so gradual ailing faculties of sight, hearing, dexterity, stamina, memory, and reaction time. They may exhibit a fear of making mistakes, and they may have strongly established preferences and pessimism about technological gimmicks. But the real problem is often that the designers have failed to anticipate the requirements of their users; they have failed to design for a range of abilities broader than their own; they have failed to test their designs with real people; and they have failed to learn from the experience of the market. Getting design right for older users is really only a continuation of getting design right for all. PMID- 11116677 TI - Implications of flexible work systems for work studies. AB - The relationship between changing work systems and work study methods is discussed by focusing on a recent trend toward more flexible work systems in different countries. These systems are commonly characterized by (a) uncoupling of working hours from business hours, (b) combining different atypical work forms and (c) individualized work patterns. Increased flexibility in working patterns is demanded because business hours may be covered by different workers and because special arrangements become necessary for unpopular shifts or linking separate jobs. Changes in work systems seem more successful when they are associated with (a) multiskilled work, (b) independent task implementation, (c) networked communication, (d) balanced or adjusted workload, and (e) accommodation of workers' preferences. Special attention is drawn to safety and health concerns and work-life effects. Many intervention studies done for work improvement are paying attention to multifaceted work aspects, locally available options, and participation by people. All these elements are important because prepackaged solutions do not exist. The following three views seem especially useful for action-oriented work studies: 1) Look at multiple aspects of the work, including work content and atypical work forms; 2) Know worker preferences and available options; and 3) Take into account work elements that may not be well defined, but important from local points of view. It is envisioned that these studies can provide support for a well-informed participatory process of work system changes in each local context. PMID- 11116678 TI - Different features of work systems in Indonesia and their consequent approaches. AB - Indonesia, with its ultimate development goal of "developing the people and the community as a whole," in fact is facing problems in the execution of this goal. With a population of more than 200 million persons, different in sociocultural background, educational level and environmental conditions, it is understandable that the process and results of technological choices and transfers for various target groups will be different. A wide range of work systems is found, from the simplest man-tool system to the most complex. The conditions are becoming even more complex, a phenomenon especially evident through studies of their sociocultural, psychological, and environmental factors. As a consequence, if success is to be gained in anticipating and understanding the role of Indonesia in the global competition that lies ahead, a very wise approach to the situation by using local values that are often based on traditional habits and customs in a modern context should be carried out. This approach will require an immense amount of time, dedication and effort. Improvement endeavors that have been carried out in different work systems in different types of activities and industries, showed that if the improvement to be sustained, a holistic, systemic, and interdisciplined participatory approach should be taken into consideration where the technical, economical, ergonomic, sociocultural, energy, and environmental factors will play significant roles. PMID- 11116679 TI - Rural health care takes center stage as advocacy efforts heat up. PMID- 11116680 TI - Georgia hospitals contribute nearly $25 billion to the state's economy. PMID- 11116681 TI - Hospitals finding it difficult serving as "safety nets". PMID- 11116682 TI - HIPAA standards for electronic transaction: final regulations published. PMID- 11116683 TI - GHA's evacuation task force sets goals at improving disaster plans. PMID- 11116684 TI - Health care's newest crisis: more patients, less health workers. PMID- 11116685 TI - Cataract surgery in patients with uveitis. PMID- 11116686 TI - Deep intramuscular methylprednisolone for the treatment of cystoid macular oedema in uveitis. AB - PURPOSE: To study the use of intramuscular methylprednisolone in the treatment of cystoid macular oedema in patients with uveitis. METHODS: A total of 32 patients with various types of uveitis with unilateral cystoid macular oedema were recruited. Patients received 160 mg of intramuscular methylprednisolone into the thigh and were reassessed 6-8 weeks later. In 17 patients in whom there had been no significant improvement, a further injection of methylprednisolone was given; a total of 49 injections. RESULTS: After a first injection a significant improvement in vision (an increase of > or = 2 Snellen lines) was seen in 15 of 32 eyes (47%), and 17 of 32 eyes (53%) showed no significant change. After the second injection 7 of 17 eyes (41%) showed a significant improvement but there was no change in 9 of 17 eyes (53%). Only one eye deteriorated more than 2 lines of Snellen acuity. There were minimal ocular or systemic side effects associated with this form of treatment. CONCLUSIONS: Despite limited success, deep intramuscular methylprednisolone may have a role as an alternative mode of treatment in uveitis patients with unilateral cystoid macular oedema, thus avoiding the potential hazards of periocular injections. In comparison with orally administered systemic corticosteroids, an injection ensures patient compliance and is associated with fewer side effects. PMID- 11116687 TI - Idiopathic polypoidal choroidal vasculopathy: a disease with diverse clinical spectrum and systemic associations. AB - PURPOSE: To report the clinical findings, angiographic results, clinical course, response to laser photocoagulation and systemic-associations in a group of patients with idiopathic polypoidal choroidal vasculopathy (IPCV). METHODS: All patients with IPCV attending the macular clinic underwent a complete ocular examination, and fluorescein and indocyanine green angiography. In addition, a systemic examination including blood pressure, full blood count, plasma viscosity and coagulation status of patients was carried out. RESULTS: We present a series of 5 patients (7 eyes) with clinical and angiographic evidence of IPCV with follow-up of 3-6 years. We report diverse demographic and clinical manifestations. One patient had polypoidal lesions found at the peripheral retina (anterior to equator) of both eyes. Three patients were treated with laser photocoagulation and achieved stable vision; 2 patients who had no laser treatment experienced deteriorated vision, one of whom had a vitrectomy. One patient was hypertensive, 2 patients were found to have raised plasma viscosity, and 1 patient had thrombocytopenia. CONCLUSIONS: The clinical spectrum of IPCV is wider than previously documented. It is a distinct clinical entity which should be differentiated from other forms of haemorrhagic and exudative maculopathy. The availability of indocyanine green angiography has allowed increased recognition of these cases. Early selective laser treatment on lesions affecting maculae could stabilise the disease. Its association with systemic cardiovascular disease and blood disorder may predispose to the recurrence of haemorrhagic events in this entity. PMID- 11116688 TI - Indocyanine green enhanced transpupillary thermotherapy of circumscribed choroidal haemangioma. AB - PURPOSE: To investigate the effect of indocyanine green in enhancing the effect of diode laser for treatment of circumscribed choroidal haemangioma. METHODS: Intravenous indocyanine green was used to enhance uptake of heat energy during transpupillary diode thermotherapy of circumscribed choroidal haemangioma. RESULTS: In a series of 6 patients treated in this manner, response to treatment was much improved compared with results without the use of indocyanine green. All patients retained the same or better visual acuity after treatment, and 67% of eyes improved visual acuity by more than 2 lines on Snellen testing. Ultrasonographic and angiographic evidence of improvement was seen in all patients. Treatment complications were minimal, comprising transient worsening of preexisting cystoid macular oedema in one case, and a small macular branch vein occlusion in the retina overlying the treated area. CONCLUSIONS: The use of indocyanine green as a contrast medium during transpupillary thermotherapy allows consistent uptake of diode laser energy, and shortens the duration of laser burn required. It is a cost-effective, easily performed outpatient procedure, with lower morbidity than other treatment modalities. PMID- 11116689 TI - Systemic non-Hodgkin's lymphoma presenting as a serpiginous choroidopathy: report of a case and review of the literature. AB - Intraocular lymphoma is a rare tumour, the two main types being primary central nervous system non-Hodgkin's lymphoma (CNS-NHL) and, less commonly, systemic lymphoma that has spread to involve the eye. We present a case of a systemic NHL resulting in a serpiginous choroidopathy. To our knowledge, this presentation has not previously been reported. The diagnosis of this case was made based on the temporal relationship between the clinical activity of the choroidopathy and its response to chemotherapy. Diagnosis of intraocular lymphoma on vitreous biopsy is notoriously difficult, and this case is presented along with a review of the literature regarding vitreous biopsy and diagnostic techniques. PMID- 11116690 TI - Pigmented paravenous retinochoroidal atrophy: a literature review supported by a unique case and insight. AB - Pigmented paravenous retinochoroidal atrophy (PPRCA) is a rare disorder of unknown origin characterised by bone corpuscle pigmentation accumulation along the distribution of the retinal veins. In addition there are peripapillary pigmentary changes as well as areas of chorioretinal atrophy adjacent to the perivenular pigmentary changes. The finding of PPRCA is usually incidental and does not affect vision. The literature regarding this condition is reviewed. Its natural course has been considered controversial. Observation of the initial insult leading to PPRCA has not previously been reported. A case that provides a photographic record over 20 years of the development of this condition from the initial insult, including the gradual development of the characteristic fundus appearance, is described. The initial presentation was with a sudden reduction of vision and gross diffuse macular oedema in one eye, which was rapidly followed by similar involvement of the fellow eye despite treatment. Thus it was possible to examine the patient at the stage of the initial insult, 5 years before the development of the typical and pathognomonic retinal changes of PPRCA. The clinical and electrophysiological findings are discussed, as also is the relevance of this case to the literature reviewed. PMID- 11116691 TI - Macular ischaemia in Behcet's disease. AB - PURPOSE: To report macular ischaemia and visual loss in patients with panuveitis due to Behcet's disease. METHODS: We describe macular ischaemia, a rare finding, in 3 eyes of 3 patients with panuveitis who were diagnosed and treated as having Behcet's disease. The patients underwent fundus fluorescein angiography (FFA) using a digital imaging system and were treated with topical and oral steroids and cyclosporine in 2 cases, and with added azathioprine in the third case. RESULTS: The 3 eyes showed macular ischaemia associated with peripheral retinal vasculitis on FFA and control of inflammation was achieved in all cases. After an average of 2 years follow-up, visual acuity and macular ischaemia improved in 2 eyes, while no recovery was seen in the third. CONCLUSION: The presence of macular ischaemia on FFA in Behcet's disease is a predictor of poor visual outcome. Macular ischaemia may show partial recovery with the treatment of the disease. PMID- 11116692 TI - Factor VII deficiency in a patient with retinal arteriolar tortuosity syndrome. AB - PURPOSE: To report a pedigree with hereditary retinal arteriolar tortuosity with macular haemorrhage and abnormality of the coagulation system. METHODS: Case report and literature review. RESULTS: A 49-year-old woman was referred due to macular haemorrhage in both eyes. Her 16-year-old son had recurrent retinal haemorrhages which presented at age 16 years and had mild retinal arteriolar tortuosity. Coagulation studies in the son revealed normal activated partial thromboplastin time (APTT), prolonged prothrombin time (PT) and 30% activity of factor VII. CONCLUSIONS: Factor VII deficiency may aggravate the haemorrhages in retinal arteriolar tortuosity syndrome. We therefore suggest conducting routine coagulation studies (PT, APTT) in all patients with retinal arteriolar tortuosity syndrome. Determination of factor VII activity is warranted only in patients with normal APTT and prolonged PT. PMID- 11116693 TI - High-altitude retinopathy and retinal vascular dysregulation. AB - PURPOSE: (a) To show that high-altitude retinopathy (HAR) is common at high altitudes even in well-acclimatised climbers and that it should not be regarded as part of the spectrum of benign mountain sickness but rather as a clinical sign with a separate aetiology. (b) To test the hypothesis that HAR could be interpreted as a clinical expression of 'ocular vascular dysregulation'. METHODS: Both eyes of the 8 mountaineers of the First Vienna Himalayan Expedition in May/June 1996 were examined 2 weeks before departure to and 2 weeks after descent from a high altitude. Retinal blood flow was measured in the right eyes of 7 climbers, using the Heidelberg Retina Flowmeter (HRF). RESULTS: Two of the 8 climbers had bilateral retinal haemorrhage after the expedition. In 5 climbers chronic hypoxic exposure caused an increase in retinal blood flow between +18% and +96%, and in 2 climbers a decrease in retinal blood flow between -21% and 31%. The 2 climbers (climbers 1 and 2) with bilateral retinal haemorrhage showed a significant increase in HRF parameters. CONCLUSIONS: HAR may be a clinical sign of mountaineers with a tendency towards ocular vascular dysregulation. The pronounced increase in all haemodynamic parameters in the 2 climbers with retinal haemorrhage combined with a dilated epipapillary network 2 weeks after the exposure reflects a retinal vessel configuration, as might be expected at high altitudes under acute hypoxic stress. An inadequate autoregulatory response of the retinal circulation under conditions of chronic hypoxia may play an important part in the pathogenesis of HAR. PMID- 11116694 TI - The role of oxidative stress in diabetic retinopathy. AB - PURPOSE: To investigate the role of oxidative stress in the development of diabetic retinopathy. METHODS: This study included 25 patients with diabetic retinopathy (group I), 34 patients with non-insulin-dependent diabetes mellitus without any angiopathy complications (group II) and 26 healthy subjects (group III). The serum malondialdehyde (MDA)-like metabolite levels as an index of lipid peroxidation, the erythrocyte glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and serum vitamin C levels of the patients and healthy subjects were measured. RESULTS: The mean serum concentration of MDA-like metabolites of patients in group I was 4.38 +/- 1.31 nmol/ml, in group II was 3.38 +/- 0.95 nmol/ml and in group III was 2.61 +/- 0.85 nmol/ml. There were significant differences between the groups (p = 0.001 for group I compared with group II, p = 0.0001 for group I compared with group III and p = 0.002 for group II compared with group III). There was a significant correlation between the serum lipid peroxidation concentrations and duration of the disease (r = 0.36, p = 0.047). The mean erythrocyte GSH-Px and SOD levels of group I were respectively 68.97 +/- 18.04 and 1597.78 +/- 296.46 U/g Hb, of group II were 64.30 +/- 19.26 and 1581.33 +/- 278.08 U/g Hb, and of group III were 65.52 +/- 17.58 and 1587.44 +/- 281.17 U/g Hb. There were no significant differences among the antioxidant enzyme levels in the three groups (p > 0.05). The mean serum vitamin C level in group I was 42.72 +/- 8.90 mumol/l, in group II was 49.26 +/- 11.52 mumol/l and in group III was 58.57 +/- 9.75 mumol/l. There were significant differences among the mean serum vitamin C levels of the three groups (p = 0.02 for group I versus group II p = 0.001 for group I versus group III and p = 0.002 for group II versus group III). CONCLUSIONS: Free radicals forming in diabetes mellitus and increasing over time may play a role in the development of diabetic retinopathy, which is an important complication of the disease. PMID- 11116695 TI - Silicone intraocular lens compression and double lens implants in diseased eyes. AB - PURPOSE: To assess the outcomes of double lens implants in hyperoptic eyes with associated pathology. METHOD: Double lens implants were used in 4 eyes of 4 patients each with a different ophthalmic or neuro-ophthalmic disease. Biometry was performed in the standard contact fashion and lens power formulae used included SRK/T, Holladay and Hoffer Q. RESULTS: Average spherical equivalent refraction improved from +6.875 D to +0.38 D. Absolute average prediction error was greatest for SRK/T (2.65 D) and least for Holladay (1.73 D). Refractive suprises were influenced by the underlying disease process. One patient showed central lens compression. CONCLUSION: Underlying disease can produce biometry errors. Structural ophthalmic or neurological disease is not a contraindication to the use of double lens implants. Double lens implants are useful to correct refractive error in the presence of underlying disease. PMID- 11116696 TI - Comparison between relative dispersion analysis of high-pass resolution perimetry and standard threshold perimetry. AB - PURPOSE: To evaluate the correlation of the dispersion index (DI) of relative dispersion analysis (RDA), a new high-pass resolution perimetry (HRP) index, with other HRP indices and those of the Humphrey standard threshold perimeter (STP) parameters. METHODS: Sixty-eight eyes were randomly recruited. Thirty-one eyes were classified as glaucomatous (high intraocular pressure, abnormal visual field and/or optic disc) and 37 as ocular hypertensives (high intraocular pressure, normal visual field, normal optic disc). All the subjects were examined with Humphrey Perimeter, program 30-2, and HRP. The HRP data were also analysed with the RDA program. Statistical analysis was performed with Student's t-test, Pearson's r correlation coefficient, Mann-Whitney nonparametric test and Spearman correlation coefficient when appropriate. RESULTS: Within the entire sample significant correlations were found between the RDA index (DI) and all the HRP indices (p < 0.001) and corrected pattern standard deviation (p < 0.01), pattern standard deviation (PSD) (p < 0.01), mean deviation (p < 0.05) and short-term fluctuation (p < 0.05) of STP. A stronger correlation was found in glaucomatous patients. In subjects with ocular hypertension DI was only weakly correlated with PSD, local deviation and form index. No difference in DI was found between glaucoma and ocular hypertension. CONCLUSION: The DI of HRP has the theoretical capacity to detect localised inhomogeneity of retinal sensitivity, but at present our data do not support this hypothesis. Before any clinical applications of this index further studies are needed. PMID- 11116697 TI - New perimetric threshold test algorithm with dynamic strategy and tendency oriented perimetry (TOP) in glaucomatous eyes. AB - PURPOSE: To investigate the time-wise reliability and efficiency of two new perimetric test algorithms, two computerised static threshold perimetry strategies, namely dynamic strategy (DS) and tendency oriented perimetry (TOP), were compared with the standard full-threshold strategy (normal strategy, NS). METHODS: We examined 41 eyes of 41 normal individuals without any ocular disease and 36 eyes of 36 glaucomatous patients, with the NS (4-to-2 dB), DS and TOP using an Octopus 1-2-3 perimeter. We analysed test time, stimulus time and the two global indices, mean sensitivity (MS) and loss variance (LV). Program 32X was used as test grid pattern. RESULTS: The mean test time for the NS was reduced by 52% with the DS and by 78% with the TOP strategy. Concerning the global indices, the MS value did not differ among the three strategies in the control or glaucoma group. However, the LV value was lower in the TOP strategy compared with the other two strategies in the glaucoma group. This suggested that the TOP strategy underestimated local glaucomatous visual field defects. The ability to detect early-stage glaucoma with the DS and TOP was inferior to that with the NS. CONCLUSIONS: The DS was more efficient than the TOP strategy for the detection of early glaucomatous defects, whereas the TOP strategy required less testing time. The TOP strategy may be an appropriate approach for patients in whom time consuming perimetry is not possible, or in whom the visual field defect is already advanced. PMID- 11116698 TI - Compliance and viewpoint of glaucoma patients in Greece. AB - PURPOSE: To document the prevalence of non-compliance and to investigate patients' perceptions concerning glaucoma in a Greek cohort. METHODS: We investigated 100 consecutive patients referred to our glaucoma clinic and already treated for chronic glaucoma. Compliance and patients' insight were ascertained by two independent observers by means of a predetermined questionnaire. All patients were subsequently assessed for their ability to instil their eyedrops accurately. RESULTS: Fifty one per cent of our patients were not aware of the nature of glaucoma, but 80% were afraid it might lead to blindness. Clinically significant non-compliance (more than two doses missed per week) was established in 44% of our patients. Men and those using eyedrops more than 4 times a day were more likely to default. Non-compliant patients exhibited higher mean intraocular pressure (22.9 vs 18.5 mmHg; p > 0.001) and worse visual field loss (10.8 vs 7.0 dB; p = 0.008) compared with compliant patients. Involuntary non-compliance was also common in our group, with only 53% instilling their eye drops accurately. CONCLUSION: Non-compliance is a significant limiting factor in glaucoma therapy in Greece. PMID- 11116699 TI - Randomised fellow eye comparison of the effectiveness of dorzolamide and apraclonidine on intraocular pressure following phacoemulsification cataract surgery. AB - PURPOSE: To compare the effectiveness of 2% dorzolamide and 0.5% apraclonidine on intraocular pressure (IOP) following phacoemulsification cataract surgery. METHODS: This prospective, randomised study comprised 54 eyes of 27 consecutive patients with age-related cataract scheduled for cataract surgery in both eyes. In each patient the eye with the higher degree of cataract was randomly assigned to receive one drop of either dorzolamide or apraclonidine immediately after surgery. The fellow eye was operated on later and received the other treatment. Cataract surgery was performed with a superior 6.0 mm sutureless frown incision, phacoemulsification and implantation of a three-piece PMMA intraocular lens. The IOP was measured pre-operatively as well as 6 h and 20-24 h and 1 week post operatively. RESULTS: The mean pre-operative IOP was not significantly different between the groups (dorzolamide group, 14.9 +/- 2.3 mmHg; apraclonidine group, 14.6 +/- 2.5 mmHg; p = 0.450). At 6 h post-operatively, the mean IOP was significantly lower in the dorzolamide than in the apraclonidine group (15.6 +/- 3.9 mmHg vs 18.0 +/- 4.0 mmHg; p < 0.001). An IOP increase of more than 5 mmHg at 6 h post-operatively occurred in 3 (12%) eyes in the dorzolamide group and in 9 (36%) eyes in the apraclonidine group (p = 0.034). At 20-24 h post-operatively and at 1 week post-operatively no difference was found between the groups. CONCLUSIONS: 2% Dorzolamide is more effective than 0.5% apraclonidine in preventing the early post-operative IOP increase following phacoemulsification cataract surgery. PMID- 11116700 TI - The effect of retrobulbar irradiation on exophthalmos, ductions and soft tissue signs in Graves' ophthalmopathy: a retrospective analysis of 90 cases. AB - PURPOSE: Retrospective analysis of the effect of retrobulbar irradiation on exophthalmos, ductions and soft tissue signs in patients with Graves' ophthalmopathy. METHODS: We analysed the charts of 111 consecutive patients who were treated with retrobulbar irradiation according to standardised intake criteria between 1992 and 1997. After exclusion of patients who underwent other treatment (with steroids or orbital decompression) shortly before or within 6 months after irradiation, and on whom insufficient data were available, 90 patients were included. For these 90 patients, we analysed the exophthalmometry, ductions, soft tissue signs and visual acuity shortly before irradiation and after 3 and 6 months, respectively. RESULTS: In the whole group, the Hertel value was on average 22 mm (SD 2.9) both before irradiation and after 3 and 6 months of follow-up. Separate analysis of data on 25 patients with bilateral exophthalmos of more than 24 mm also revealed no change in exophthalmos at follow-up. In the whole group, both abduction and elevation had improved by about 1 degree (SD 6.6 degrees; p = 0.05) after 3 months. This improvement has little clinical significance. In a subgroup of 14 patients who showed more than 10 degrees of restricted eye motility in one or more directions, both abduction and elevation had increased by about 4 degrees (SD 10 degrees; p = 0.02) at 3 and 6 months follow-up. Soft tissue signs had improved at 6 months after irradiation. We found no change in visual acuity after irradiation. CONCLUSION: Retrobulbar irradiation in Graves' ophthalmopathy does not seem to reduce exophthalmos. It probably improves eye motility in patients with severe restrictions. The late improvement in soft tissue signs may either be a late effect of irradiation or be related to the natural history of the disease. PMID- 11116702 TI - Non-organic visual loss in children. AB - BACKGROUND: Patients with non-organic visual loss (NOVL) can take up a disproportionate amount of clinic time and clinicians often resort to expensive and prolonged investigations to ensure the correct diagnosis. This is especially the case in children. METHODS: The case notes of 30 children (18 girls, 12 boys) were retrospectively reviewed following presentation with a primary complaint of visual impairment and a diagnosis of non-organic visual loss. This figure represents 1% of new paediatric referrals to our unit. Associated symptoms included headache, periorbital pain, diplopia, photopsia and photophobia. Visual field defects were present in 5 patients and spasm of the near reflex in 1 child. RESULTS: Treatment consisted of reassurance and was associated with recovery of normal visual function in all cases. Three children were referred to other health care professionals. All psychophysical, electrophysiological and neuroradiological investigations were negative. CONCLUSION: Our study shows that non-organic visual loss is relatively common in pre-pubertal children and that this condition can be safely diagnosed using standard clinical tests in the majority of cases. Prompt diagnosis prevents unnecessary investigations and prolonged 'disease' course. Coexisting social conflict was common and may be a contributory factor. Careful explanation and reassurance to both the child and parents remains the mainstay of management. PMID- 11116701 TI - Echographic study of extraocular muscle thickness in children and adults. AB - BACKGROUND: Echobiometric evaluation of extraocular muscles in normal subjects has been performed previously, but only in adults. We determined extraocular muscle thickness in normal subjects in three age groups. METHODS: Extraocular muscle thickness was studied in 75 normal subjects divided into three age groups (5-10, 11-15 and 28-37 years) using a Biovision B-scan-S instrument in standardized A-mode (frequency, 10 MHz; biometry resolution, 0.15 mm; depth, 40 60 mm; points on X axis, 512; levels on Y axis, 256). All measurements were performed by the same operator and repeated five times. The reproducibility of the technique was determined using the coefficient of variation. The one-way ANOVA test was used to compare the three groups, and the two-tailed unpaired t test was used to compare subjects aged 5-10 years and those aged 11-15 years, and subjects aged 11-15 years with those aged 28-37 years. RESULTS: The technique showed good reproducibility. In subjects 5-10 years old, the coefficient of variation was 8%; in subjects 11-15 years and 28-37 years old, it was 5%. Increased muscle thickness was observed with age (p < 0.001). A statistically significant difference between the medial and inferior recti muscles in subjects 11-15 years and 28-37 years old was found (p < 0.001). CONCLUSIONS: The increased thickness of all recti muscles may be influenced by growth (primarily during puberty), and the variations in thickness of the extraocular muscles may be attributable to near-vision stimulus of the inferior and medial recti muscles. PMID- 11116703 TI - Factors leading to lens implant decentration and exchange. AB - PURPOSE: To examine the intra- and post-operative factors leading to posterior chamber intraocular lens (IOL) decentration in patients requiring IOL exchange, and to identify avoidable causes of IOL decentration. METHODS: Case records of 17 patients who had undergone posterior chamber IOL exchange were examined for: (i) any complication or alteration to the original intended surgical procedure, (ii) IOL type and position at the completion of initial surgery, (iii) IOL position at the time of re-operation. RESULTS: The decentred lens implants were injected silicone plate-haptic IOLs in 10 patients, small (5.5 mm) optic diameter PMMA IOLs in 4 patients and large (7 mm) optic diameter PMMA IOLs in 3 patients. In all cases, decentration was due to IOL subluxation. Early decentration of the injected lenses was due to IOL implantation in eyes without a continuous capsulorrhexis. In contrast late decentration was due to subluxation associated with capsule fibrosis. Decentration of small optic PMMA IOLs was found to be associated with an anterior capsule tear and haptic malposition in the ciliary sulcus. Decentration of large optic PMMA IOLs was associated with posterior displacement of one haptic through a posterior capsule defect, zonule dehiscence or fixation of one haptic in the sulcus and one in the capsule bag. CONCLUSION: Clinically significant post-operative subluxation of injected silicone IOLs may be minimised by implanting only into a lens capsule bag with an intact capsulorrhexis. The risk of decentration of small optic PMMA IOLs may be minimised by positioning the haptics at 90 degrees to any capsulorrhexis tear. After cataract surgery complicated by posterior capsule rupture or zonule dehiscence, it is important to assess the remaining capsule support and, where sufficient, implant a large optic diameter posterior chamber IOL in the ciliary sulcus. PMID- 11116704 TI - Cataract surgery and the optometrist. AB - PURPOSE: To determine the outcome of discharge on the first day following cataract surgery and the feedback from patients' optometrists. METHODS: Casenotes of patients who had cataract surgery between 1 April 1997 and 30 June 1998 were analysed. Patients without complications were discharged on day 1 and advised to see their optometrist at 1 month. Patients were given a form for refraction with a pre-paid envelope for their optometrist. Completed letters from the optometrists were returned to the hospital to be analysed by the principal surgeon and acted on appropriately. A questionnaire was sent out to patients whose notes did not contain any information after the first post-operative examination. RESULTS: A total of 318 eyes from 288 patients underwent cataract surgery. Completed forms from the optometrist were received in 245 (77%) cases; no optometrist's letter was found in the remaining 73 cases (23%). Of these 73 patients, 50 (68%) had other ocular pathology requiring hospital follow-up and 9 had died. There were only 6 patients about whom post-operative information could not be obtained. There were no significant differences regarding the age and sex of those who did or did not attend the optometrist. CONCLUSION: Patients without complications can be discharged to the care of their optometrist on the first day following cataract surgery. With good communication between hospital and the optometrist, better use can be made of available resources. PMID- 11116706 TI - Sildenafil (Viagra) a cause of proliferative diabetic retinopathy? PMID- 11116705 TI - Visual and tear function improvement after superficial phototherapeutic keratectomy (PTK) for mid-stromal corneal scarring. AB - PURPOSE: To study the changes in visual and tear film function following superficial excimer laser phototherapeutic keratectomy in patients with mid stromal corneal scars. METHODS: Fourteen eyes of 14 patients with mid-stromal corneal scars seen at the Department of Ophthalmology at Kobe Kaisei Hospital underwent superficial phototherapeutic keratectomy (PTK). The subjects underwent routine ophthalmic examinations, corneal sensitivity measurements, tear film break-up time (BUT), Schrimer test and tear film lipid layer interferometry. Thirty eyes of 15 normal control subjects were also studied. The patients and the control subjects were compared for pre-PTK tear function parameters and tear film lipid layer interferometry grade. The alterations in these parameters within 6 months following PTK were also determined. RESULTS: Visual improvement was achieved in 12 eyes (86%). A hyperopic shift was observed in all eyes. The average pre-PTK corneal sensitivity and tear film BUT were lower in patients compared with control subjects before PTK. Tear film lipid layer interferometry grades were also higher in the patients than the controls before PTK. All these parameters improved gradually and significantly after PTK. Schirmer test results did not show any significant alterations after PTK. CONCLUSION: We conclude that PTK is an effective means of treating corneal scars and attaining visual improvement, even in cases with deeper corneal involvement, and may obviate the need for corneal transplantation. Simultaneous improvements in corneal sensitivity and tear film stability suggest favourable effects of PTK on the ocular surface. PMID- 11116707 TI - A new method of temporary tarsorrhaphy. PMID- 11116708 TI - Delayed onset acute retinal necrosis 20 years following herpetic encephalitis. PMID- 11116709 TI - The role of IgM isotype anticardiolipin antibodies in occlusive ocular vascular disease: report of two cases with primary antiphospholipid antibody syndrome. PMID- 11116710 TI - Corneal birth trauma: a cause for sensory exotropia. PMID- 11116711 TI - Chronic conjunctivitis due to lacrimal system blockage relieved by dacryocystorhinostomy. PMID- 11116712 TI - Recurrent lens epithelial cell proliferation with an AcrySof lens implant. PMID- 11116713 TI - Opacification of silicone intraocular implant requiring lens exchange. PMID- 11116714 TI - Bilateral retro-orbital plasmacytoma. PMID- 11116715 TI - Infectious crystalline keratopathy after stitch removal in a lamellar corneal graft. PMID- 11116716 TI - Solitary choroidal metastasis managed by transpupillary thermotherapy. PMID- 11116717 TI - Indirect delivery of argon laser to disperse premacular haemorrhage in a recumbent visually handicapped patient with Terson's syndrome. PMID- 11116718 TI - Safe management of a late-onset bleb leak with a needling technique. PMID- 11116719 TI - Early blindness due to retinopathy of infantile cystinosis. PMID- 11116720 TI - The application of ultrasonic biomicroscopy in the management of traumatic hypotony. PMID- 11116721 TI - Bilateral pre-retinal macular haemorrhage following blunt head injury. PMID- 11116722 TI - Treatment of aqueous misdirection by trans-scleral diode laser photocoagulation. PMID- 11116723 TI - Teaching junior ophthalmologists phacoemulsification under topical anaesthesia. PMID- 11116724 TI - Sixth nerve palsy following epidural spinal cord stimulation for lower limb ischaemia. PMID- 11116725 TI - Can vitreous haemorrhage indicate non-accidental injury if mild retinopathy of prematurity is present? PMID- 11116726 TI - Nightmares with topical beta-blocker. PMID- 11116727 TI - The importance of recognising Streptococcus milleri as a cause of orbital cellulitis. PMID- 11116728 TI - How to enjoy the sun safely. PMID- 11116729 TI - The elderly couple who complain about each other. PMID- 11116730 TI - Key developments in dermatology. PMID- 11116731 TI - Casebook: atopic eczema. PMID- 11116732 TI - Diagnosing bacterial skin infections. PMID- 11116733 TI - Recognising blisters: causes and treatment. PMID- 11116734 TI - Acne: when, where and how to treat. PMID- 11116735 TI - Recognising common nail conditions: a guide. PMID- 11116736 TI - Breast cancer: detection and management. PMID- 11116737 TI - Peptic ulcer after H pylori. PMID- 11116739 TI - Rosea by any other name... PMID- 11116738 TI - Passing the MRCGP: preparing the video. PMID- 11116740 TI - A stressed GP turns to drink and drugs. PMID- 11116741 TI - Recent advances in care for the elderly. PMID- 11116742 TI - Mouth conditions in the elderly: Part 1. PMID- 11116743 TI - The patient with possible dementia. PMID- 11116744 TI - Managing hip osteoarthritis. PMID- 11116745 TI - Diagnosing skin disease in the elderly. PMID- 11116746 TI - A guide to helping the patient who complains of constipation. PMID- 11116748 TI - Passing the MRCGP: the written paper. PMID- 11116747 TI - Changing to insulin in type 2 diabetes. PMID- 11116749 TI - Testicular self-examination. PMID- 11116750 TI - A nasty surprise over lunch. PMID- 11116751 TI - The effect of diflunisal on the elimination of triamterene in human volunteers. AB - A major metabolic pathway for triamterene (a potassium sparing diuretic) is aromatic hydroxylation followed by sulphate conjugation. Diflunisal (a salicylate anti-inflammatory agent) also undergoes sulphate conjugation of its phenolic group as a major pathway. We investigated the possible effect of diflunisal on the elimination of triamterene (competition for phenolic sulphonation) in six healthy volunteers by studying the disposition of single doses of triamterene (100 mg) taken alone and in the presence of steady-state levels of diflunisal. Diflunisal coadministration (500 mg b.i.d.) had no effect on the pharmacokinetics of triamterene itself. However, plasma AUC of p-hydroxytriamterene sulphate was greater (4.6 times), and its renal clearance lower (0.24 times), in the presence of diflunisal. There was no change in the formation clearance or protein binding of p-hydroxytriamterene sulphate in the presence of diflunisal. The data point to competition for renal excretory pathways rather than sulphonation capacity. This interaction could have clinical relevance since p-hydroxytriamterene sulphate is pharmacologically active. PMID- 11116752 TI - Identification of UDP-glucuronosyltransferases involved in the human hepatic metabolism of GV150526, a novel glycine antagonist. AB - The major metabolic pathway for elimination of GV150526 is by glucuronidation exerted by glucuronosyl transferases (UGTs). Potential exists for the modification of GV150526 pharmacokinetics by drugs capable of inhibiting the glucuronidation of GV150526. Using human liver microsomes, 44 compounds were screened for inhibition of GV150526 glucuronidation. These compounds were selected because they are extensively glucuronidated themselves or are used as concomitant medication in the treatment of acute stroke. For 11 compounds out of the 44, full inhibition kinetics were performed to determine their Ki-value and mechanism of inhibition. To predict possible in vivo drug-drug interactions, the theoretical percentage of inhibition (i) was determined, based on in vitro determined Ki-values, and the expected Cmax plasma levels of GV150526 and the inhibitor. Of the 11 compounds examined, only propofol had an i-value of 6.6; for all other compounds i-values were lower than 2.1. These results indicate that although in vitro inhibition is observed, the likelihood of in vivo drug-drug metabolic interactions occurring is low. The inhibition results suggest that in addition to UGT1A1, also UGT1A3, UGT1A8/9, and UGT2B4 are involved in the glucuronidation of GV150526. The involvement of UGT1A1 and UGT1A8/9 was confirmed from studies using cDNA expressed human UGT cell lines. PMID- 11116753 TI - The fate of diphenyl sulphide, diphenyl sulphoxide and diphenyl sulphone in the rat. AB - Radiolabelled [UL-14C]-diphenyl sulphide, [UL-14C]-diphenyl sulphoxide and [UL 14C]-diphenyl sulphone were administered by gavage (1.0 mmol/kg body weight) to adult male Wistar rats following an overnight fast. For all compounds, faeces were the major route of excretion of radioactivity (50%). Urinary elimination (40%) was similar during the first (19%) and second (16%) days and a small amount of radioactivity (6%) was found within the carcass after four days. From urinary and faecal data, metabolism occurred via ring hydroxylation with subsequent conjugate formation. Oxidation of the sulphur to form the sulphoxide and sulphone also took place; a small amount of sulphoxide reduction was apparent but no sulphone reduction was found. No evidence for exclusion of the sulphur was obtained, and it appeared unlikely that extensive cleavage of the ring structures occurred. PMID- 11116754 TI - Effect of clarithromycin on the pharmacokinetics of tolbutamide. AB - The aim of this 2 x 2 randomized double blind crossover study was to evaluate the effect of a single dose of clarithromycin on the pharmacokinetics of tolbutamide in nine healthy male volunteers. Each volunteer received orally 500 mg of tolbutamide, or 500 mg of tolbutamide and 250 mg of clarithromycin. The washout period between the two treatments was 7 days. Serum levels of tolbutamide were determined by HPLC. Serum profiles were analysed using a non-compartmental model. Blood glucose levels were also estimated using a glucometer (Ames) and Glucostix (Bayer). There was approximately 20% increase in mean absorption rate constant and 26% increase in mean bioavailability of tolbutamide in the presence of clarithromycin. A hypoglycemic effect was reported upon co-administration of the two drugs. PMID- 11116755 TI - beta-Lactamase activities and resistance to antibiotics of Haemophilus influenzae, H. parainfluenzae and H. aphrophilus strains identified in throat cultures from children. AB - Haemophilus bacteria are normally present in the upper respiratory tract of healthy individuals. However, these bacteria could be opportunistic pathogens especially in children. The present study was conducted to determine beta lactamase activity of Haemophilus from the throat cultures of children with upper respiratory tract infections. 154 Haemophilus strains were isolated from throat swabs of 208 children whom had upper respiratory tract infections. Among the 154 Haemophilus strains isolated, 117 H. influenzae (76%), 35 H. parainfluenzae (22.7%), and two H. aphrophilus (13%) were identified by API NH. beta-Lactamase activity was positive in 42 isolates of 117 H. influenzae isolates, while it was negative in 75 isolates. beta-Lactamase activity was positive in 20 H. parainfluenzae isolates, and negative in 15. All the H. aphrophilus isolates were beta-lactamase negative. It is known that beta-lactamase positive Haemophilus bacteria are resistant to some antibiotics. Therefore, the antibiotic resistance of Haemophilus was further investigated in relation to beta-lactamase activity. The in vitro antibacterial susceptibilities of Haemophilus strains for ampicillin, sulbactam-ampicillin, trimethoprim-sulfamethoxazole, gentamicin, chloramphenicol and ciprofloxacin were tested by disk diffusion method on chocolate agar. In 42 beta-lactamase-positive H. influenzae isolates, 32 isolates were resistant against ampicillin. In 20 beta-lactamase-positive H. parainfluenzae isolates, 16 were resistant against ampicillin. The two beta lactamase negative H. aphrophilus were sensitive to ampicillin. Biotypes and serotypes were also investigated. Biotypes of H. influenzae strains were as follows: 40 strains biotype II, 25 strains biotype I, 14 strains biotype III, and 38 strains biotypes VII, VIII, V, and IV. Biotypes of 35 H. parainfluenzae strains were: 6 strains biotype III, 5 strains biotype I, 5 strains biotype IV. Biotypes of remaining 19 isolates were II, VIII, VI and VII. The serotypes of H. influenzae strains were determined by specific antiserums. Serotypes of 117 H. influenzae found were type a, b, c, d, and f. PMID- 11116756 TI - Does carbamazepine-epoxide conjugate with glutathione? On an existing but almost forgotten possibility. AB - The anticonvulsant carbamazepine is widely used to treat affective disorders and behavioural disorders in non-epileptic children. We report an elevated plasma level of carbamazepine-10,11-epoxide in a cystinuric child after daily medication with 300 mg carbamazepine while the serum level of carbamazepine was in the therapeutic range. The concentrations of carbamazepine and its epoxide derivative were determined by HPLC. The formation of a glutathione conjugate of carbamazepine-10,11-epoxide is raised as a hypothesis. PMID- 11116757 TI - Education provision for surgical senior house officers. AB - The provision of formal education in surgical SHO training is a requirement for posts to be accredited by postgraduate deans. We carried out a questionnaire survey of surgical SHOs attending a Royal College of Surgeons of England study day to quantify the amount of time given to formal education and supervised operating. We found that 79% of SHOs at district general hospitals and 87.5% at teaching hospitals were provided with a formal education programme. Of these SHOs, 77% were able to attend the teaching on a regular basis. In an average week, 8% of SHOs received no supervised operating experience and 37% had no operations supervised by a consultant. These results suggest that many hospitals fall short of the accepted guidelines on basic surgical training. This issue must be addressed by the Royal Colleges. PMID- 11116758 TI - Calman training. PMID- 11116759 TI - Rationing of surgery in the National Health Service: the plastic surgery model. PMID- 11116760 TI - Titanic troubles? PMID- 11116761 TI - Virtual vivas. PMID- 11116762 TI - Research plans for current basic surgical trainees: still publish or perish? PMID- 11116763 TI - Teaching orthopaedic skills. PMID- 11116764 TI - Clinical governance, the profession, the 'P' words and responsibility. PMID- 11116765 TI - Organisation of acute general hospital services: a surgical synopsis of the JCC document. PMID- 11116766 TI - Care throughout the NHS? PMID- 11116767 TI - Live anatomy. PMID- 11116768 TI - A new mental health service: high quality and user-led. PMID- 11116769 TI - Predictive value of pharmacological activity for the relative efficacy of antidepressant drugs. Meta-regression analysis. AB - BACKGROUND: There is uncertainty about the contribution of specific pharmacological properties to the efficacy of antidepressants. AIMS: To assess whether specific pharmacological characteristics of alternative antidepressants resulted in altered efficacy compared to that of selective serotonin reuptake inhibitors in the treatment of major depression. METHOD: Meta-regression analysis of randomised trials that compare treatment with a selective serotonin reuptake inhibitor and an alternative antidepressant. RESULTS: One-hundred-and-five randomised trials were included. None of the factors identified a priori predicted a statistically significant improvement in outcome across the trials. CONCLUSIONS: This analysis does not provide evidence that antidepressants acting at more than one pharmacological site differ in efficacy from drugs selective for serotonin reuptake in the treatment of major depression. PMID- 11116770 TI - Violence risk prediction. Clinical and actuarial measures and the role of the Psychopathy Checklist. AB - BACKGROUND: Violence risk prediction is a priority issue for clinicians working with mentally disordered offenders. AIMS: To review the current status of violence risk prediction research. METHOD: Literature search (Medline). Key words: violence, risk prediction, mental disorder. RESULTS: Systematic/structured risk assessment approaches may enhance the accuracy of clinical prediction of violent outcomes. Data on the predictive validity of available clinical risk assessment tools are based largely on American and North American studies and further validation is required in British samples. The Psychopathy Checklist appears to be a key predictor of violent recidivism in a variety of settings. CONCLUSIONS: Violence risk prediction is an inexact science and as such will continue to provoke debate. Clinicians clearly need to be able to demonstrate the rationale behind their decisions on violence risk and much can be learned from recent developments in research on violence risk prediction. PMID- 11116771 TI - Assessing effectiveness of treatment of depression in primary care. Partially randomised preference trial. AB - BACKGROUND: There is a mismatch between the wish of a patient with depression to have counselling and the prescription of antidepressants by the doctor. AIMS: To determine whether counselling is as effective as antidepressants for depression in primary care and whether allowing patients to choose their treatment affects their response. METHOD: A partially randomised preference trial, with patients randomised to either antidepressants or counselling or given their choice of either treatment. The treatment and follow-up were identical in the randomised and patient preference arms. RESULTS: There were 103 randomised and 220 preference patients in the trial. We found: no differences in the baseline characteristics of the randomised and preference groups; that the two treatments were equally effective at 8 weeks, both for the randomised group and when the randomised and patient preference groups for a particular treatment were combined; and that expressing a preference for either treatment conferred no additional benefit on outcome. CONCLUSIONS: These data challenge several assumptions about the most appropriate treatment for depression in a primary care setting. PMID- 11116772 TI - Patient-held shared care records for individuals with mental illness. Randomised controlled evaluation. AB - BACKGROUND: Few formalized shared care schemes exist within psychiatry and the evidence base for sharing psychiatric care is weak. AIMS: To evaluate the utility of patient-held shared care records for individuals with long-term mental illness. METHOD: Cluster-randomised controlled parallel-group 12-month trial involving 90 patients with long-term mental illness drawn from 28 general practices. RESULTS: Carrying a shared care record had no significant effect on mental state or satisfaction with psychiatric services. Compared with controls, patients in the shared care group were no more likely to be admitted (relative risk 1.2, 95% CI 0.86-1.67) and attend clinic (relative risk 0.96, 95% CI 0.67 1.36) over the study period. Uptake of the shared care scheme was low by patients and professionals alike. Subjects with psychotic illness were significantly less likely to use their records (relative risk 0.51, 95% CI 0.27-0.99). CONCLUSIONS: Patient-held records may not be helpful for patients with long-term mental illness. PMID- 11116773 TI - The Australian National Survey of Mental Health and Well-Being. Common psychological symptoms and disablement. AB - BACKGROUND: The mental health of populations can be represented by case prevalence rates and by symptom scales. Scales have the advantage of identifying sub-syndromal levels of distress, which may be common and associated with considerable disability. AIMS: To examine the distribution of common psychological symptoms and associated disablement in the Australian population. METHOD: A household sample of 10,641 individuals representative of the adult population of Australia was interviewed using the Composite International Diagnostic Interview and completed scales measuring recent symptoms and disablement. RESULTS: Symptom scales showed similar associations with socio economic variables as did diagnoses, although only a small amount of variance in symptom levels was explained by these variables. Considerable disablement was associated with symptom levels indicating distress but not reaching levels for formal diagnoses of anxiety or depression. CONCLUSIONS: Symptom scales provide parsimonious measures of psychological distress and are appropriate for use in large-scale surveys of mental health and disablement. PMID- 11116774 TI - Time to recovery of an inception cohort with hitherto untreated unipolar major depressive episodes. AB - BACKGROUND: Generalisability of existing studies on the naturalistic history of major depression is undermined by overrepresentation of in-patients and tertiary care academic centres, inclusion of patients already on treatment and/or incomplete follow-up. AIMS: To report the time to recovery of an inception cohort of unipolar major depressive episodes. METHOD: A multi-centre prospective follow up study of patients with a mood disorder, who had been selected to be representative of the untreated first-visit patients at 23 psychiatric settings from all over Japan. RESULTS: The median time to recovery of the index episode after treatment commencement was 3 months (95% CI 2.5-3.6): 26% of the cohort reached asymptomatic or minimally symptomatic status by 1 month, 63% by 3 months, 85% by 12 months and 88% by 24 months. CONCLUSIONS: Our estimate of the episode length was 25-50% shorter than estimates reported in the literature. PMID- 11116775 TI - Association of depression and gender with mortality in old age. Results from the Amsterdam Study of the Elderly (AMSTEL). AB - BACKGROUND: The association between depression and increased mortality risk in older persons may depend on the severity of the depressive disorder and gender. AIMS: To investigate the association between major and mild depressive syndromes and excess mortality in community-living elderly men and women. METHOD: Depression (Geriatric Mental State AGECAT) was assessed in 4051 older persons, with a 6-year follow-up of community death registers. The mortality risk of neurotic and psychotic depression was calculated after adjustment for demographic variables, physical illness, cognitive decline and functional disabilities. RESULTS: A total of 75% of men and 41% of women with psychotic depression had diet at follow-up. Psychotic depression was associated with significant excess mortality in both men and women. Neurotic depression was associated with a 1.67 fold higher mortality risk in men only. CONCLUSIONS: In the elderly, major depressive syndromes increase the risk of death in both men and women, but mild depression increases the risk of death only in men. PMID- 11116777 TI - Nithsdale Schizophrenia Surveys. 20. Cognitive function in a catchment-area-based population of patients with schizophrenia. AB - BACKGROUND: Cognitive deficits are a core aspect of schizophrenia but there has been no study of cognitive function in a catchment-area-based population of patients with schizophrenia. AIMS: To assess cognitive function in a population of patients with schizophrenia, and relate it to community functioning. METHOD: All patients with schizophrenia in Nithsdale, south-west Scotland, were identified (n = 182). Measures of assessment were: National Adult Reading Test (NART), Mini-Mental State Examination (MMSE), Rivermead Behavioural Memory Test (RBMT), Executive Interview (EXIT), FAS Verbal Fluency and Health of the Nation Outcome Scales (HoNOS). RESULTS: We assessed 138 patients, mean age 48 years (standard deviation (s.d.) 15). Only 14% were in-patients. The mean premorbid IQ as assessed by NART was 98 (s.d. 14); 15% of patients had significant global cognitive impairment (MMSE); 81% had impaired memory (RBMT); 25% had executive dyscontrol (EXIT); and 49% had impaired verbal fluency (FAS). Scores on the functional impairment sub-scale of HoNOS correlated with all measures of cognitive impairment. CONCLUSIONS: Cognitive dysfunction is pervasive in a community-based population of patients with schizophrenia. PMID- 11116776 TI - Impact of emotion on memory. Controlled study of the influence of emotionally charged material on declarative memory in Alzheimer's disease. AB - BACKGROUND: In an earlier study we showed that a powerful emotional experience (the Kobe earthquake) reinforced memory retention in patients with Alzheimer's disease, but we could not control factors other than the emotional impact of the earthquake. AIMS: To test our previous findings in a controlled experimental study. METHOD: Recall tests consisting of two short stories were administered to 34 patients with Alzheimer's disease and 10 normal subjects. The two stories were identical except for one passage in each story: one was emotionally charged (arousing story) and the other (neutral story) was not. RESULTS: In both groups, the emotionally charged passage in the arousing story was remembered better than the counterpart in the neutral story. In addition, the extent of the memory improvement was similar in the subjects and in the controls. CONCLUSIONS: The results provide further evidence that emotional arousal enhances declarative memory in patients with Alzheimer's disease, and give a clue to the management of people with dementia. PMID- 11116778 TI - Third ventricle enlargement and developmental delay in first-episode psychosis: preliminary findings. AB - BACKGROUND: Third rather than lateral ventriculomegaly may be a more specific finding in psychosis. The relevance of ventricular abnormality remains unclear. AIMS: To investigate the developmental correlates of ventricular enlargement. METHOD: Information on childhood development and magnetic resonance images in 1.5 mm contiguous sections were collected on 21 patients experiencing a first episode of psychosis. RESULTS: Patients (n = 21) had significantly less whole brain volume and enlarged third and lateral ventricles compared to controls (n = 25). Third ventricle (r = 0.48, P < 0.03) and lateral ventricle (r = 0.65, P < 0.01) volumes correlated with developmental score. Patients with developmental delay had significantly larger third and lateral ventricles than those without. CONCLUSIONS: Enlargement of both third and lateral ventricles is found in first episode psychosis and is related to developmental delay in childhood. Insult to periventricular areas is relevant to the neurobiology of the disease. These findings support the view that schizophrenia involves disturbance of neurodevelopmental processes in some patients. PMID- 11116779 TI - Psychosocial and psychiatric risk factors for suicide. Case-control psychological autopsy study. AB - BACKGROUND: Few studies of suicide have simultaneously examined the individual and combined effects of psychosocial and psychiatric risk factors. AIMS: To do so in a representative sample of suicides. METHOD: A case-control psychological autopsy was conducted among 113 consecutive suicides and 226 living controls matched for age, gender, ethnicity and area of residence in Taiwan. RESULTS: Five major risk factors (loss event, suicidal behaviour in first-degree relatives, ICD 10 major depressive episode, emotionally unstable personality disorder and substance dependence) were found to have independent effects on suicide from multivariate conditional logistic regression analysis. CONCLUSIONS: Effective intervention and management for loss event and major depressive episode among emotionally unstable subjects with a family tendency of suicidal behaviour, frequently also comorbid with alcohol or other substance dependence, may prove to be most effective for suicide prevention in different populations. PMID- 11116780 TI - Seasonal variation in suicides: diminished or vanished. Experience from England and Wales, 1982-1996. AB - BACKGROUND: Seasonal variation in suicidal death has been observed in many countries. In particular, a cyclic variation was found for both men and women in England and Wales in the 1960s and 1970s. Men showed a single 12-month cycle whereas women showed two cycles. AIMS: To re-examine the seasonal variation in suicides in England and Wales for the period 1982-1996. METHOD: A harmonic analysis was used to detect the seasonality of the suicide data. RESULTS: The seasonal effect on suicide is greatly diminished in England and Wales. This is shown by the reduced amplitude and smaller proportion of variance accounted for by the season. CONCLUSIONS: The seasonal effect on suicide has either diminished or vanished. PMID- 11116782 TI - Implications of evolutionary theory for psychiatry. PMID- 11116781 TI - Implications of evolutionary theory for psychiatry. PMID- 11116783 TI - Comments on the UK700 case management trial. PMID- 11116784 TI - Comments on the UK700 case management trial. PMID- 11116785 TI - Lithium and mortality. PMID- 11116786 TI - Finding the evidence in forensic rehabilitation. PMID- 11116787 TI - Arachnophobia: a practical management device. PMID- 11116788 TI - Thrombocytosis due to clozapine treatment: working towards an early marker for clozapine-induced agranulocytosis. PMID- 11116789 TI - Etomidate-induced convulsion prior to electroconvulsive therapy. PMID- 11116790 TI - Focus on psychiatry in Poland. Past and present. PMID- 11116791 TI - [Evaluation of the psychometric properties of the Spanish version of 5 questionnaires for the evaluation of post-traumatic stress syndrome]. AB - AIM: To determine the psychometric properties of the Spanish versions of the following scales for assessing PTSD patients: TQ, CAPS-DX, DTS, TOP-8, and DGRP. METHODS: Data from 63 PTSD patients and 23 healthy subjects were analysed. Internal consistency and test-retest reliability (after 15 days) were calculated. Convergent validity was analysed by correlating subjects' scores with the number of symptoms (DSM-IV) and scores on the CGI scale. Discriminative capability was analysed by comparing TQ, CAPS-DX, DTS, TOP-8, and DGRP patients' scores with scores from healthy subjects, and between patients' subgroups according to the presence or absence of psychiatric comorbidity and to the degree of severity as determined by the CGI-S. RESULTS: CAPS-DX, DTS, TOP-8, and DGRP showed an adequate internal consistency (Cronbach alpha: 0.74-0.91) and all of them obtained ICC between 0.77 and 0.93. The five questionnaires were able to discriminate between patients and healty subjects, and between the patients' subgroups. CONCLUSIONS: The Spanish versions of the TQ, CAPS-DX, DTS, TOP-8, and DGRP have shown adequate reliability and validity for assessing PTSD patients in daily clinical practice. CAPS-DX seems to be more adequate a diagnostic and DTS a severity rating scale. PMID- 11116792 TI - [Symptomatology and gender in schizophrenia]. AB - OBJECTIVE: To study the possible symptomatological gender differences in a sample of schizophrenic outpatients. METHOD: A sample of 239 schizophrenic patients (DSM IV criteria) was administered a demographic questionnaire and the Positive and Negative Syndrome Scale (PANSS). The PANSS symptoms were grouped in accordance with Kay's five factors model. RESULTS: The ratio women/men was 1/2. Women had a later age at onset. Women were more likely to be married and to live independently, as well to be occupationally active. Both groups were similar in subtype of schizophrenia. No differences were found in PANSS' results; in global or grouped scores. CONCLUSION: No symptomatological differences were found between women and men with schizophrenia. PMID- 11116793 TI - [Attempted suicide versus suicidal intention: a study of differential characteristics]. AB - INTRODUCTION: Suicidal behaviours are internationally considered as a serious health problem and, consequently, considerable efforts are being made in the search for factors related with this behaviour. OBJECTIVES: To determine if there are any differences between those who attempted suicide and those with suicidal ideation. PATIENTS AND METHOD: During 1997-98, we interviewed a total of 134 patients hospitalised due to attempted suicide (n = 49), suicidal ideation (n = 38), and a control group made up of other psychiatric patients (n = 47) in the Psychiatric Unit of the Hospital de Jove (Gijon). All patients completed the "Protocol for Investigation of Suicidal Behaviour" which contains different sociodemographic and clinical variables, and the Scale for Suicide Ideation, Hopelessness Scale, Hamilton Depression Rating Scale, and Eysenck Personality Questionnaire-Adult form. RESULTS: No significant differences were found in sociodemographic or clinical variables between the suicidal ideation and attempted suicide groups. Only the intensity of hopelessness as measured on the Beck scale (greater in the suicidal ideation group) and emotional instability in females (also greater in the suicidal ideation group) are the variables which differentiate those who present suicidal ideation and those who attempt suicide. PMID- 11116794 TI - [Long-term effects of the treatment with risperidone versus conventional neuroleptics on the neuropsychological performance of euthymic bipolar patients]. AB - INTRODUCTION: An increasing number of studies point to the persistence of cognitive deficits in a subpopulation of euthymic bipolar disorders. Atypical antipsychotics represent an interesting alternative treatment compared with conventional neuroleptics, because the former might cause lesser affection of cognitive functions. The objective of this study was to establish, for euthymic bipolar patients who need long-term antipsychotic treatment, the neuropsychological and outcome (occupational functioning) differences between the patients treated with risperidone and the patients treated with conventional neuroleptics. SUBJECTS AND METHOD: A sample of twenty RDC bipolar I and II patients were assessed by means of the SADS. All of them were euthymic (HDRS < 8; YMRS < 6) for at least 6 months and treated with antipsychotics. Patients who received risperidone (N = 11) were compared with patients who received conventional neuroleptics (N = 9) with respect to their neuropsychological profile. RESULTS: There were significant differences in Trail Making Test-part B (p = 0.038) and in occupational outcome (p = 0.024), favouring patients who were treated with risperidone. There were no significant differences in other neuropsychological tests, but the patients treated with risperidone tended to score higher in most measures of cognitive performance. CONCLUSIONS: Among bipolar patients who need long-term antipsychotic treatment for preventing relapses, patients who receive risperidone show more cognitive flexibility and better occupational adaptation than patients treated with conventional neuroleptics. PMID- 11116795 TI - [Alcohol consumption in opioid-dependence patients in treatment with naltrexone]. AB - OBJECTIVES: Alcohol consumption in opiate dependence in naltrexone treatment was evaluated. MATERIAL AND METHODS: The alcohol consumption and their modifications were evaluated in two groups of heroin dependents in naltrexone treatment. A prospective study was made in two periods of time, evaluating 50 patients in the first one, and 82 in the second one. RESULTS: In both of them we found a significant increase of the amount of alcohol consumed, that was higher in the patients who had shown alcohol abuse before opiate dependence. CONCLUSIONS: The opiate dependent in naltrexone treatment increase of alcohol consumed. PMID- 11116796 TI - [Risperidone in the treatment of psychotic disorders associated with mental retardation]. AB - INTRODUCTION: The safety and effectiveness of risperidone in patients with a diagnostic of mental retardation and/or psychotic and disturbance disorder was assessed in an open and observational study. MATERIAL AND METHODS: A total of 127 patients with DSM-III-R diagnoses of mental retardation together with psychotic disturbance (psychotic and/or behaviour disturbance) was included in an open label surveillance study. The study objectives were to evaluate safety and efficiency of risperidone in these patients during a six-month period. Risperidone was assessed by the Clinical Global Impression (CGI), schizophrenia subscales of Scale for Evaluating Emotional Disorders in Severely and Profoundly Mentally Retarded Persons (DASH) and Assessment and Information Rating Profile (AIRP), UKU subscale for neurological side effects and spontaneous reports. RESULTS: Ten patients (7.9%) were excluded from the statistical analysis due to protocol violation. Seventeen patients (14.5%) dropped out. Risperidone was used at a mean dosage of 2.98 (2.37 mg daily. Risperidone produced a significant reduction, vs baseline, in the mean total scores of CGI and DASH-AIRP scales from day 15 onwards. There was a significant reduction in the total UKU subscale for neurological, autonomic side effects scores and psychotic symptoms. Risperidone was generally well tolerated. During the 6 months of study period, 93.2% of the patients did not suffer any adverse event; the resting 7.7% suffered one or more side effects. DISCUSSION: In this study, risperidone was a safety and effectiveness treatment in patients with a diagnostic of mental retardation and psychotic and/or disturbance disorder associated. PMID- 11116797 TI - [Functional neuroimaging of emotions and bipolar disorder]. AB - In this review we comment the results of functional neuroimaging works of emotions on normal population and some parallelisms with the emotional changes of bipolar disorder correlated with their functional neuroimaging. Initially we refer the emotional ontogenetical development of human brain based on regional cerebral sanguineous flow evolution (FSC). Secondly we describe the differences of FSC between the externally generated emotions versus internally; between positive versus negative emotions and the correlation between FSC and some facial expressions. When FSC of bipolar disorder is compared with normal emotions on general population, we observe that temporal cortex, the prefrontal medial and insular anterior cortex, change their perfusion with the switch or the change of emotional expression. It is possible to determine if the findings obtained in samples of healthy subjects and bipolar patients converge in a dimensional model, or if on the contrary they support the categorical hypotheses, moving the emotional aspects to a second term on bipolar disorder. PMID- 11116798 TI - [Hyperviscosity syndrome and mental disorders]. AB - The hyperviscosity syndrome has been described clinically as the triad of bleeding, visual signs and neurological manifestations associated with elevated serum viscosity. Several reports have recognised an association between hyperviscosity and altered mental status. Since to our knowledge only a case of hyperviscosity-induced delirium has been described (1), we raise the possibility of this diagnosis in the most of this reported cases, based on the nature of the symptoms, sudden onset and fluctuating course, and its resolution with plasmapheresis. In this paper we review the literature about hyperviscosity syndrome and altered mental status. In conclusion, serum hyperviscosity should be added to the large list of causes altered mental status, especially of delirium. Since plasmapheresis can reverse clinical symptoms, it early recognition and the measurement of serum viscosity is essential in patients suffering from diseases that may lead to this syndrome, and who develop psychiatric symptoms. PMID- 11116799 TI - [A pseudopsychiatric case caused by thrombotic thrombocytopenic purpura]. AB - The present article describes the case of a woman, aged 25 years, who the non specific and variable neurologic symptomatology and the confusional episodes observed at admission required psychiatric care until the diagnosis, and subsequent treatment, of thrombotic thrombocytopenic purpura (TTP) was done. The variable and unstable manifestations induce "to interpret the disease psychologically", we must not forget the importance of the somatic assessment in psychiatry to prevent premature and mistaken conclusions that delay a correct diagnosis and, therefore, an adequate treatment. PMID- 11116800 TI - [Psychotic syndrome after withdrawal of cyproheptadine: remission with olanzapine]. AB - Cyproheptadine is a drug with direct antimuscarinic, antihistaminic (H1) and antiseronergic (5HT2) effects. In spite of its use as an orexigenic agent in normal or anorectic subjects, little is known about its effect in affective, anxiety or psychotic disorders. We present the case of a 14-year patient beginning with a psychotic syndrome after the sudden withdrawal cyproheptadine, achieving a total control of the symptoms with olanzapine at low dose, besides a good family, school and social adaptation. PMID- 11116804 TI - [In Process Citation] AB - Case-reports from two female patients with an occlusion||| of the right external iliac artery and femoral artery are presented due to a||| large-vessel vasculitis. Both patients suffered from systemic lupus||| erythematosus This rare manifestation occurred within the first few years of||| the disease and was important for prognosis and further treatment. Other||| manifestations of the disease were general symptoms and polyarthritis. In one||| case the vasculitis was confirmed by histology. A fibrous thickening of the||| intima and a vasculitis of small vessels within the adventitia were the||| prominent feature. This observation supports the idea of small vessel||| vasculitis as the characteristic manifestation in lupus||| erythematosus. PMID- 11116809 TI - [Investigation of the mite fauna content of dust samples collected from pig and poultry farms. Report of the first finding in Western Europe of the house-dust mite Dermatophagoides evansi in dust from poultry houses]. AB - Mites can be important sources of airborne allergens, especially on farms. Two dust samples from pig farms and three dust samples from poultry farms were investigated for mites. House-dust mites were present in the poultry-dust samples, but not in the pig-dust samples. Furthermore, storage mites and predatory mites also were found in the poultry-dust samples. Specifically, the house-dust mite Dermatophagoides evansi was found in the dust samples from two poultry farms. Subsequently, a dust sample was collected from five other poultry farms. Again, D. evansi was present in dust from these farms. This is the first time that D. evansi is reported in dust from poultry farms in Western Europe outside Norway. If D. evansi cross-reacts with other Dermatophagoides spp., then poultry farmers and their families, but also other professionals working in the poultry industry, such as veterinarians, may be exposed to house-dust mites with potential clinical consequences, both domestic and occupational. PMID- 11116810 TI - [Dramatic increase of congenital defects in calves]. PMID- 11116811 TI - [A remarkable variety of subjects at the American Congress of Parasitology]. PMID- 11116812 TI - [Immunocastration?]. PMID- 11116813 TI - [Minister Brinkhorst opens yearly congress 2000 with heartfelt speech. "You are dependent on the cattle farmer, but he is dependent on you"]. PMID- 11116814 TI - [Veterinary antibiotics regimen in the Netherlands]. PMID- 11116833 TI - [In Process Citation] PMID- 11116834 TI - [In Process Citation] AB - The shoulder joint takes a special position among all the||| other joints of the human body because of its special requirements of stability||| and mobility. Knowledge of the biomechanics of the shoulder joint forms the||| basis for the development of modern concepts of reconstructive surgery and||| arthroplasty. Most of the biomechanical findings are the result of research||| performed on cadaver shoulders using increasingly sophisticated methods of||| measurement. These studies elucidate the interaction of the static and dynamic||| factors which contribute to the delicate balance of the glenohumeral joint.||| Recently performed research is increasingly being focussed on more detailed||| analyses of muscle forces and stress distribution in the subchondral bone and||| periarticular soft tissues. The efficiency of the computer systems now||| available has enabled the development of complex, virtual shoulder models and||| three-dimensional finite element analyses. In the future a pure mechanical||| understanding has to be modified to extend to a concept which includes more||| data obtained from living subjects, especially with regard to muscle activity||| under varying loads and neuromuscular feedback systems which currently are||| difficult to assess. PMID- 11116835 TI - [In Process Citation] AB - Painful stiffness of the shoulder is an ill-defined||| clinical entity that is difficult to assess and delicate to treat. The||| nomenclature used is broad and includes terms such as frozen shoulder, adhesive||| capsulitis, focal algodystrophy, stiff shoulder, contracted shoulder, and||| others. Apart from its idiopathic form, the disease can be initiated by trauma,||| infection, tumour, radiation, systemic and local metabolic disturbances.||| Pathoanatomically, the common denominator is an inflammatory vascular||| proliferation followed by thickening, scarring, and retraction of the joint||| capsule. The inflammatory process often starts at the rotator interval and may||| extend to the subacromial space. Clinical diagnosis is based on history and||| physical examination. Generally the onset of pain precedes the perception of a||| reduced range of motion by weeks or months. In early stages of the disease, the||| inflammatory type of pain dominates, i.e., the patient's main complaint ist||| pain at night. In the later stage, range of motion gradually decreases.||| Patients do not often complain about reduced motion, probably because of its||| slow onset. Treatment options are a combination of mobilisation exercises with||| intra-articular steroids, hydraulic distension of the joint capsule,||| manipulation under anaesthesia, arthroscopic and/or open arthrolysis. The||| appropriate choice of protocol is just as important as its correct timing. In||| the inflammatory phase, aggressive treatment protocols are probably||| contraindicated. Complications of invasive protocols are rare but deleterious||| and therefore have to be taken into consideration. New anti-anglogenetic agents||| may enhance functional results and shorten the rehabilitation||| phase. PMID- 11116836 TI - [In Process Citation] AB - Calcifying tendinitis of the rotator cuff is a common||| disorder of the shoulder which affects mainly individuals between 30 and 50||| years of age. The etiology is still a matter of speculation. The calcification||| is a reactive process actively mediated by cells in a viable environment. The||| deposit undergoes an evolution (precalcific stage--calcific stage with||| formative phase, resting period and resorption--postcalcific stage), which||| ultimately remodels normal tendon tissue. However, the evolutionary stages of||| the disease do not always follow the typical sequence. A symptomatic deposit||| may persist or postcalcific tendinitis develop. The treatment should be based||| on the knowledge of the natural history of the disease, which shows a strong||| tendency towards self healing by spontaneous resorption of the deposit. The||| stage of evolution of the disease should be judged, combining pain history,||| morphology of the deposit on plain X-rays, and ultrasound findings. The||| therapeutic approach depends on the evolution of the disease. During the||| resorption phase we favor a conservative approach. For deposits which are not||| under resorption we propose a concept which consists of three steps: a||| conservative approach, extracorporal shock wave therapy (ESWT) or needling, and||| arthroscopic surgery. The efficacy of ESWT and needling has still to be proven.||| Patients with persisting pain after steps 1 and 2 are candidates for surgical||| removal of the deposit. We prefer the arthroscopic approach. In some cases an||| additional arthroscopic subacromial decompression (ASD) is||| indicated. PMID- 11116837 TI - [In Process Citation] AB - The impingement syndrome is a common disorder of the||| shoulder girdle. The causes for this syndrome may be anatomic changes in the||| coracoacromial arch, also within the ultrastructural regions, on the one hand,||| or changes in the biomechanics which have developed for various reasons, on the||| other. Diagnosis is based on roentgenograms using the appropriate technique. In||| large-scale studies, sonography has proved to be an extremely sensitive||| screening method for differential diagnosis of rupture of the rotator cuff.||| Magnetic resonance imaging might gain in value in the diagnosis of impingement||| as regards differential diagnosis of rupture of the rotator cuff because this||| technique- when employed appropriately--allows exact viewing of the soft tissue||| and the anterior part of the acromion. In the majority of cases conservative||| treatment is the method of choice. Methods of treatment are sonography,||| galvanization, and application of heat. Physiotherapy should not be initiated||| until pain relief has been achieved by other measures. Infiltration therapy is||| of considerable value in the management of pain due to impingement. Application||| of cortisone into the subacromial space must also be considered critically. As||| regards conservative therapy, only few evidence-based publications provide||| information on the effectiveness of different treatment regimens. Surgical||| therapy is only indicated in cases of pain resistant to the conservative||| therapy for a certain period. Furthermore, only an outlet impingement can be||| treated successfully by surgical decompression. The surgeon decides on the||| surgical method--open surgery or arthroscopy. Of course, arthroscopic methods||| are less invasive; however, up to now the superiority of one of the surgical||| methods over the other could not yet be proven by mid-term clinical results.||| Other surgical methods such as wedge osteotomy in the region of the spina||| scapulae are still in the experimental stage. By surgical and conservatives||| methods, good and even excellent results can be achieved in about 80% of the||| cases. The question remains as to why 20% of the patients show unsatisfactory||| results. One explanation might be that factors such as pathologic changes of||| the muscular balance and an altered microstructure of the tendons of the||| rotator cuff are rarely taken into consideration and Neer's concept of||| decompression is overestimated. Further research will be required in the field||| of biomechanics but clinical research on treatment concepts should also be||| undertaken to develop more differentiated strategies of||| treatment. PMID- 11116838 TI - [In Process Citation] AB - The traumatic etiology of rotator cuff lesions is a||| matter of controversial discussed in legal assessments. Because of the||| relatively high prevalence of degenerative changes with increasing age,||| including partial and complete rotator cuff tears, it may be difficult to||| demonstrate the cause of an acute traumatic rotator cuff tear. This article||| presents a review of the literature concerning current knowledge of the||| anatomy, biomechanics and pathogenesis of rotator cuff pathology. According to||| this, a catalogue of potentially adequate and inadequate trauma mechanisms is||| proposed. Another focus of this work is in the post-traumatic diagnostic steps||| following persistent rotator cuff deficient shoulder function. X-ray,||| ultrasound, MRI and operative findings in rotator cuff tears underline the||| criteria for distinguishing between traumatic and degenerative lesions. From a||| legal point of view (e.g., private accident insurance, workers compensation||| claims), various minor and major arguments are defined, which could help the||| expert in judging the cause of post-traumatic rotator cuff||| deficiency. PMID- 11116839 TI - [In Process Citation] AB - Although most subacromial decompressions are performed||| arthroscopically, rotator cuff repair is still performed using an open or||| mini-open procedure. Arthroscopic techniques have improved in the last decade,||| however, so that rotator cuff repair can also be performed arthroscopically.||| The potential complications of open repair are thus reduced and the superior||| functional results of cuff repair in comparison to debridement alone are||| maintained. The proposed advantages of the arthroscopic method are that it||| provides access to the glenohumeral joint for inspection and treatment of||| intra-articular lesions. The skin incisions are smaller, detachment of the||| deltoid muscle is not necessary, and there is less soft tissue dissection. By||| inspecting the bursal and articular side of the ruptured cuff, it is possible||| to measure the size of the tear and assess the quality of the tendon and||| whether it can be repaired. We present our arthroscopic technique of rotator||| cuff repair using bioabsorbable suture anchors and demonstrate our 1- to||| 6-years results with various suture anchors. PMID- 11116840 TI - [In Process Citation] AB - The anatomy and biomechanics of the acromioclavicular||| (AC) joint have been understood for a long time; however, the importance of||| this joint in the clinical setting is often underestimated. During clinical||| examination various sensitive functional tests can document any AC pathology.||| For X-ray documentation special techniques are necessary. Other imaging||| techniques are rarely indicated. The Rockwood classification for AC joint||| separation has increased our understanding of the pathology, which, in turn,||| leads to a better understanding of conservative and surgical therapy. Within||| the last few decades surgical treatment has shifted from AC to coracoclavicular||| stabilization. In patients with clinically relevant degenerative joint disease,||| resection of the lateral clavicle has proved to be a reproducible procedure.||| This operation can be performed using the conventional, open technique or with||| a minimally invasive procedure (arthroscopic resection of the AC joint; ARAC).||| In unstable joints, resection should be combined with a stabilization||| procedure. PMID- 11116841 TI - [In Process Citation] AB - Degenerative arthrosis of the shoulder is less commonly||| diagnosed than at the joints of the lower extremity. The shoulder joint does||| not bear weight and some of the mechanical stresses are taken up by the||| subacromial space. However, anatomical studies reveal a significant incidence||| of degenerative changes at the glenohumeral joint with increasing age.||| Arthrosis is caused by mechanical loading, rotator cuff defects, and abnormal||| joint motion following surgery. Clinical symptoms are rarely focused on the||| glenohumeral joint. Clinical findings are also unspecific. Rotation of the||| elevated arm with compression of the joint is a reliable sign of arthrosis. The||| diagnosis is made with anteroposterior and axial radiographs. Ultrasonography||| should always be performed and computed tomography only in specific cases.||| Nonoperative treatment includes analgesic and antiphlogistic medication, motion||| therapy, and muscle exercises. Shoulder arthroplasty is favoured in advanced||| arthrosis. New prostheses are intended to reconstruct the normal anatomy as||| closely as possible. PMID- 11116842 TI - [In Process Citation] AB - Since their first description several years ago, superior||| glenoid labral lesions have increasingly been blamed for shoulder problems||| associated with sports. Originally merely describing arthroscopically visible||| upper labral/biceps abnormalities, the current understanding is that often||| clinical problems such as impingement pain or even rotator cuff disease can be||| secondary to these lesions, especially in overhead athletes. Impingement in||| these cases is caused by superior shoulder instability originating from an||| unstable biceps insertion that is present for example in SLAP (superior labrum||| from anterior to posterior) lesions. Additional problems such as internal or||| posterosuperior impingement that are often found simultaneously in these||| patients are pathomorphologically located in the same anatomical region and||| therefore make exact diagnosis and thus treatment more complex. Magnetic||| resonance imaging with intra-articular contrast enhancement and particularly||| arthroscopy are the primary tools for exact diagnosis and classification of||| superior labral/biceps pathology. Therapeutically, lesions with unstable biceps||| origin (SLAP types 2 and 4) require operative refixation, as we have seen in||| our 50 cases in the last 4 years, in order to reestablish the stabilising||| effect of the biceps tendon for the shoulder joint. The arthroscopic technique||| for repair of these lesions using different devices of implantable suture||| anchors is presented. Long-term pain-free shoulder function in competitive||| athletes, throwers in particular, thus requires anatomical reconstruction of||| the originally unstable biceps, which is the causal therapy for these||| lesions. PMID- 11116843 TI - [Premature skin aging caused by smoking]. PMID- 11116844 TI - [Current status of photoepilation]. AB - During the last few years the fast development of different laser and laser-like systems for photoepilation and their one-sided representation in media has led to confusion among physicians and patients. The object of this review is to give a structured report of different pulsed laser and laser-like systems (Epilight, Photoderm, long-pulsed ruby-, alexandrite- and diodelaser) that are available in Germany, including their side effects and complications. The current status of scientific investigation in this field is discussed. PMID- 11116845 TI - [Rare cutaneous manifestaions of lupus erythematosus. A clinical overview]. AB - Lupus erythematosus (LE) is a disease with a wide spectrum of cutaneous and systemic manifestations and has been the subject of many studies over several decades. Clinical features of patients with LE show a great variation, and for this reason it is difficult to develop a unifying concept of this disease. Consequently, this has led to the identification of subsets which have been defined by constellations of clinical and photobiological features, histological changes as well as laboratory abnormalities. Besides the characteristic classical forms such as systemic LE (SLE), subacute cutaneous LE (SCLE), and discoid LE (DLE), there are uncommon variants of LE which often lead to diagnostic difficulties. Bullous LE (BLE) and urticarial vasculitis are listed as characteristic but non-specific manifestations of systemic LE. LE tumidus (LET), LE hypertrophic/verrucous (LEHV), chilblain LE, and LE profundus (LEP) are uncommon subtypes of chronic cutaneous LE. Annular erythema and papulonodular mucinosis are further uncommon cutaneous manifestations of LE. This clinical review summarizes the typical features of the uncommon forms of LE in order to improve clinical diagnostic precision and to achieve a better differentiation of the subtypes. PMID- 11116846 TI - [Control of section margins without any gaps in paraffin sections of melanoma of the face]. AB - BACKGROUND AND OBJECTIVE: Malignant melanomas on sun-exposed skin are often poorly circumscribed and thus recur frequently. The aim of our clinical trial was to compare conventional to a modified micrographic surgery of primary melanoma. PATIENTS/METHODS: 28 patients with in-situ (n = 7) and invasive (n = 21) melanoma were treated with conventional surgical excision with wide margins; eight of these patients developed a local recurrence. In comparison, 20 patients with primary in-situ (n = 7) and invasive (n = 13) melanoma as well as four patients with recurrence after conventional surgery underwent modified micrographic surgery with delayed closure of the wound. RESULTS: Paraffin specimens of the margins revealed in half of the patients remnants of melanoma although excision was extended to non-lesional skin. Within a mean follow-up period of 21.3 months, none of our patients treated by modified surgery developed a recurrence. CONCLUSIONS: Local recurrencies of melanoma on sun exposed skin may be avoided by means of a modified micrographic surgery using permanent histologic sections. PMID- 11116847 TI - [Symptom of micro-orchism for fertility and endocrine testicular function]. AB - BACKGROUND AND OBJECTIVE: There are no reliable findings regarding the frequency and etiology of the spermatogenetic and endocrine functional restrictions in the small testicle (microorchidism). This information is needed to properly assess the risk when multiple testicular biopsies for assisted reproduction techniques lead to further volume reduction. PATIENTS/METHODS: 535 patients consulting our andrological clinic were included into the study. Orchidometric findings were corelated to spermatological data. Furthermore, FSH, Inhibin B, LH, testosterone and 17 estradiol were analysed. In 116 cases extended andrological studies clarified the etiology of the microorchidism. RESULTS: 26.5% of the patients had a testicular volume < or = 12 ml, 67.1% were in the normal area, 6.4% show a testicular volume > 25 ml. Patients with small testicles had azoospermia in 44.6%, OAT-syndrome in 20.2% and endocrine hypogonadism in 19.8%. The most frequent causes were Klinefelter-syndrome, maldescensus testis, varicocele, secondary atrophies and idiopathic clinical pictures. CONCLUSIONS: Microorchidism is understood as a one-sided testicular volume < or = 12 ml in adult men. We consider the risky taking of tissue out of the small testicle as avoidable, if predictive diagnostic factors for the discovering of spermatozoa via operation are analysed. An androgen-substitution in microorchidism may be carried out in selected individuals. PMID- 11116848 TI - [Penicillin allergy as a diagnostic problem. Overview and personal studies]. AB - BACKGROUND AND OBJECTIVE: Penicillin allergy is a common clinical problem. The distinction between penicillin and para-infectious exanthems is difficult. We investigated the reliability of the history, as well as the sensitivity and specificity of skin tests and specific IgE levels. PATIENTS/METHODS: 160 patients with a history of penicillin allergy were retrospectively evaluated in the outpatient department of a dermatological clinic. RESULTS: Nearly 50% were diagnosed as allergic to penicillin by detection of specific IgE or skin test. About 60% of the patients with immediate type reactions, and 72% with maculo papular erythema showed positive reactions in skin tests. Significantly more patients were diagnosed as allergic to penicillin by intradermal testing than by prick testing (p < 0.05). The sensitivity of the specific IgE RAST was 17.9%; the specifity, 89.5%. For the prick test the sensitivity was 8.2%; the specificity 90.8%. For the intradermal test the sensitivity was 26%; the specifity 69.7%. CONCLUSIONS: We suggest a step by step procedure to detect penicillin allergy making the diagnostic results as valid as possible. PMID- 11116849 TI - [Hereditary prolidase deficiency. Contribution to differential therapy refractory leg ulcer diagnosis]. AB - Leg ulcers may be caused by many different diseases. Most frequently, they are due to vasculopathies, to a lesser extent to metabolic, neuropathic or hematologic diseases. Neoplasms, connective tissue diseases, infections, trauma, and panniculitis should also be included in the differential diagnosis. A 38-year old Caucasian female patient with hereditary prolidase deficiency developed progressive and very painful leg ulcers. The ulcers first appeared in childhood and did not respond to various treatments. Additional features of prolidase deficiency included mental retardation, short stature, extensive dental caries, and multiple malar teleangiectases. Hereditary prolidase deficiency is a very rare autosomal recessive disease. It is caused by heterogeneous mutations of the prolidase gene and affects many aspects of protein metabolism. Ion exchange chromatography and high voltage electrophoresis of urine can prove the suspected diagnosis. So far, there is no efficient therapy for hereditary prolidase deficiency. All reported treatment attempts have ended in failure. PMID- 11116850 TI - [Cowpox and catpox infection. 2 Clinical case reports]. AB - In patients with painful, possibly hemorrhagic vesicles or black crusts, especially on hands/fingers or face/neck with typical history (contact to cows or feral/cats) the possibility of a cowpox/catpox infection has to be considered. The clinical diagnosis can be confirmed with the electron microscopy; cytoplasmic inclusions may be detected histologically. Further useful diagnostic tools are the identification of the cowpox/catpox virus by PCR or in cell culture as well as serological tests to detect virus specific antibodies. We report the development of typical skin lesions of a cowpox/catpox infection in two female patients who had contact with cows or cats. Recent diagnostic and therapeutic approaches are also discussed. PMID- 11116851 TI - [A hemorrhagic acral form of dyskeratosis follicularis Darier]. AB - A 76-year-old woman presented with a venous leg ulcer 7. Incidentally hemorrhagic macules on hands and fingers and dystrophy of toe and fingernails were noticed. Except for submammary erythema, no further dermatologic signs were shown. Histology of palmar lesions fit with a hemorrhagic lesion of dyskeratosis follicularis in acral skin. During following months actinic induced reddish-brown hyperkeratotic papules appeared on the forehead and in seborrheic and intertriginous areas of the trunk. Papules were most prominent during summer and almost completely resolved in winter. The hemorrhagic variant of dyskeratosis follicularis represents a rare clinical variant which has to be separated from other purpuric macules on acral sites. PMID- 11116852 TI - [Bilateral segmental neurofibromatosis simulating epidermal nevus]. AB - Neurofibromatosis is a neuroectodermal systemic disease. A rare variant of this condition is bilateral segmental neurofibromatosis. A 29-year-old man presented with bilateral papillomatous plaques in the lumbar dermatomes. Clinically, the lesions were very similar to an epidermal nevus but histologic examination revealed superficial neurofibromas. Family history, ophthalmologic and neurologic investigations were unremarkable. The unusual morphologic presentation of bilateral segmental neurofibromas in this case points to the wide clinical spectrum of the disease and the significance of histologic examination in systematic nevoid lesions. PMID- 11116853 TI - [Ectopic parotid tissue. An usual differential cervical cystic tumor diagnosis]. AB - Cystic tumors of the neck are usually cysts, inflammatory masses, lymph nodes, benign and malignant tumors or metastases. Malformations are less common. Ectopic salivary tissue is a rather rare but clinically important malformation. Accessory and aberrant tissue can be identified histologically. There may be an association with lymphatic tissue. Possible complications include inflammation as well as the development of benign and in rare cases malignant salivary gland tumors. We report on two patients who underwent surgical treatment under the tentative diagnosis of epidermal cysts which histologically turned out to be ectopic parotid tissue. PMID- 11116854 TI - [Recurrent mitigated erysipelas in severe combined immunodeficiency]. AB - A patient with congenital severe combined swiss-type immune deficiency suffered from recurrent cellulitis and at least 26 documented cases of bacterial pneumonia. Parenteral immunoglobins prevented the recurrence of the bacterial pneumonias but not of the cellulitis. This was successfully halted by benzathine penicillin injections every four weeks. PMID- 11116855 TI - [Botulinum toxin. A neurotoxin for dermatologic therapy]. PMID- 11116856 TI - [Certification of curricula for graduate education of the specialist. Netherlands system of graduate education]. PMID- 11116857 TI - [What is the place today for the new treatments for condylomata acuminata?]. PMID- 11116858 TI - [Imiquimod: immunologic-response modifier]. PMID- 11116859 TI - [Imiquimod--clinical efficacy]. PMID- 11116860 TI - [Perspectives for the immunotherapy of condylomata]. PMID- 11116861 TI - [Medico-economic analysis of therapy of external condylomata acuminata]. PMID- 11116862 TI - [Physiopathology and epidemiology of HPV anogenital diseases]. PMID- 11116863 TI - [Does industry buy university research? Cooperation is necessary, but not the dependency]. PMID- 11116864 TI - [Diffuse rules on patent protection of genes. Scientists risk retroactive payment requirements]. PMID- 11116865 TI - [The Catholic church is positive toward xenotransplantations]. PMID- 11116866 TI - [No more screws in malunited femoral neck fractures. "Total hip alloplasty prevents reoperations and suffering"]. PMID- 11116867 TI - [Work-related stress behind sudden death cases? Two young Swedes hit by a known Japanese phenomenon]. PMID- 11116868 TI - [Hopeful future for echocardiography. Progress within both the function and perfusion areas]. AB - Echocardiography is presently a feasible method for quantitative estimation of intracardiac flows, pressure levels and for hemodynamic evaluation of valvular disease. The evaluation of regional myocardial function is still based on subjective scrutiny, and no routine method for the estimation of myocardial blood flow is available. We present an overview of newly developed techniques that are beginning to gain purchase in clinical practice. The use of native second harmonic imaging to improve image quality and of tissue Doppler to provide objective measurements of regional myocardial function is discussed. This article describes the transformation of tissue Doppler information into parametric images as in strain rate imaging, and overviews the use of ultrasound contrast agents. Used together with new imaging modalities, myocardial contrast echocardiography holds promise for future quantification of myocardial blood volume and flow. Other emerging echocardiographic technologies discussed are non-invasive measurement of coronary flow reserve and three dimensional cineloop visualization, developed to increase our understanding of cardiovascular physiological and anatomical coupling. PMID- 11116869 TI - [New products improve treatment of chronic leg ulcers. Fewer dressing changes reduce both the patients' inconveniences and the costs]. PMID- 11116870 TI - [Insufficient legal rights of traffic accident victims. Insurance companies should force more active compensation claims]. AB - According to law, persons injured in road traffic accidents are entitled to compensation. In almost 92% of cases, victims accept a modified solution suggested by the insurance company involved. If an agreement cannot be reached, or the disability is 10% or more, the case is referred to The Road Traffic Injuries Commission (TSN). Insurance companies often claim that the lengthy wait for a final ruling--up to four years or more--is due to the lack of a doctor's certificate of disability. PMID- 11116871 TI - [Insurance medicine training--a cost-effective initiative]. AB - A training program in insurance medicine for younger physicians in clinical practice is presented. The program is individualized, problem-based and oriented towards clinical practice. Physicians participate in the program for two or five working days--depending on the level of clinical competence--and the following items are stressed: case reports with clinical insurance medicine aspects, knowledge of ordinary work places among the general population, training in early rehabilitation, and awareness of the process of certification for sick leave. During the past three years 73 physicians at all levels of clinical competence, mostly in the specialties general practice and occupational health, have passed the training program. There are many positive spin-off effects of the training, as well as substantial reductions in the rate of physician certification for sick leave and for early retirement pensions in the region. In 1999 the average level of sick-leave utilization was two days less than the national average, a reduction in relative terms by about 50% from 1996, when the regional level was 20% higher than the national level. In addition, during the period 1996-1999 the number of appeals of decisions of the Regional Social Insurance Office has been reduced from 33% to 6% of all decisions in which physician certificates for sick leave played a role. A nation-wide introduction of such a training program might possibly reduce the state expenditure for sick-leave insurance by 3 billion SEK on a yearly basis (about 10% of the current sick-leave related expenditure). PMID- 11116872 TI - [Teaching future physicians about gender differences. Gender of the physician does matter!]. PMID- 11116873 TI - [The foundation of "feminine" and "masculine". Useful theories for the training of future physicians concerning the importance of gender]. AB - A gender perspective on health and consultation is part of medical education today. Teaching about gender must not focus on differences between men and women as essential, biological, and unchangeable. The meaning of "feminine" and "masculine" is largely a social construction, i.e. the behavior and character of an individual are seldom determined by sex. Furthermore, women and men live under different conditions and have different positions in society. Medical students need to be aware of this and reflect upon the influence it may have on their professional role and practice. To achieve this awareness, knowledge about the construction of gender is needed. This article reviews relevant research in this field. The gender of the physician is used as a basis and illustration of this. PMID- 11116874 TI - [Superficial pain in connection with coitus is a complex pain syndrome]. PMID- 11116875 TI - [A Swedish study of women treated for cervix cancer. Gynecologic cancer often affects sexuality]. AB - Gynecological cancer and its treatments are often associated with physical, psychological and social consequences that affect the woman's and the couple's sexuality. Especially due to their impact on sexuality, physical changes distress the women considerably, and warrant consideration for women of all ages. Women with gynecological cancer ask for information about possible side-effects of the disease and treatment that can affect the sexual function. At follow-up visits these issues can be attended to through information, with specific suggestions including advice about topical estrogen and dilators, to alleviate possible long term sequelae. PMID- 11116876 TI - [Decisions made by the industry to stop trials--important to inform when and why it happens]. PMID- 11116877 TI - [Solution of current health problem: "a health check money" makes free choice possible for the patients]. PMID- 11116878 TI - [Sudden death in an adult professional with no visible sickness--some reflections]. PMID- 11116879 TI - [Competition between child health services one more time or a trustful cooperation?]. PMID- 11116880 TI - [BMJ is not on the impact factor list]. PMID- 11116881 TI - [Is allergy to electricity an electrophobia?]. PMID- 11116882 TI - [Are parents of children with neuropsychiatric diagnoses child abusers and child molesters?]. PMID- 11116883 TI - [A child's own forensic psychiatrist?]. PMID- 11116884 TI - [The Lapp physician Axel Bill was an Agorander-descendent]. PMID- 11116885 TI - [Defend vaccination against measles! A life-threatening disease for immunocompromised patients]. PMID- 11116886 TI - [Continuing professional development is the aim of a changed continuing education]. PMID- 11116887 TI - [Molecular defects may cause epilepsy. New discoveries can provide better possibilities for directional diagnostics and treatment]. AB - Specific defects in neuronal ion channel proteins have recently been identified in some forms of hereditary epilepsy. A deletion of 300 amino acids from the COOH terminal of the K+ channel reduces the electrical stability of the neuron in subjects with benign familial neonatal seizures. Defects in the protein subunits of the Na+ channel may prolong neuronal depolarization in children with generalized epilepsy with febrile convulsions. A point mutation in one of the ACh receptor subunits may reduce the function of inhibitory interneurons in subjects with autosomal dominant nocturnal frontal lobe epilepsy. Finally, several different defects in the Ca2+ channel amino acid sequence have been identified in various types of epilepsy in mice in which symptoms and EEG show similarities to those in human petit mal. This remarkable progress in the precise localization of ion channel defects in epilepsy provides a novel basis for the development of more differentiated diagnosis and pharmacological therapy. PMID- 11116888 TI - [Fetal alcohol syndrome--unnecessary suffering which has not become rarer. Eyes are affected in up to 90 per cent of cases]. AB - Fetal alcohol syndrome, FAS, is characterized by pre- and/or postnatal growth deficiency, CNS dysfunction, typical facial features and evidence of prenatal alcohol exposure. FAS is often an unrecognized diagnosis and is based on clinical symptoms alone. The cognitive and behavoral disturbances have a great influence on the children's ability to learn and on their adult life. FAS is to a great extent found in areas with less than satisfactory socio-economic conditions. Since ophthalmologic involvement is frequent, an eye examination may be helpful in making a diagnosis. Prevention is essential since the brain damage prevails, despite comprehensive medical and social supportive efforts being made during childhood and in education. PMID- 11116889 TI - [Eye muscle paralysis can increase the knowledge on autism]. AB - One third of Swedes with thalidomide embryopathy have sixth and seventh cranial nerve palsy, named Mobius syndrome/sequence, and some of them suffer from autism. Further, other patients with Mobius sequence have autism. The teratogenic effect of thalidomide on the development of different organ structures occurs early in pregnancy. By studying the birth defects, information can be obtained concerning when the damage occurred. Erroneous development of the sixth and seventh cranial nerves is induced early in pregnancy. The ensuing palsy may be associated with autism. New research is focusing on genes controlling early cranial nerve development. PMID- 11116890 TI - [Cytostatic therapy reduces the immune defense. Children treated for leukemia have impaired immunity against measles and rubella]. AB - A study is summarized analyzing the levels of serum antibodies against vaccination antigens in 43 children treated for acute lymphoblastic leukemia. Two different therapeutical regimens were used. All children had been immunized against measles and rubella before being diagnosed with leukemia. Eight of the 24 children treated 1986-1991 lacked protective levels of antibodies against measles; four of the 24 children lacked antibodies against rubella. In the second cohort of children (n = 16) treated from 1992 and onwards, nine lacked protective levels of antibodies against measles, eight lacked antibodies against rubella. PMID- 11116892 TI - [Gender perspective in medicine]. PMID- 11116891 TI - [Vaccination of the elderly against pneumococcal disease is cost-efficient. Mass vaccination of all aged 65 and over is recommended]. AB - The pneumococcal vaccine has been shown to be about 70 percent efficacious in preventing invasive pneumococcal disease in elderly persons. In a European multicenter study, pneumococcal vaccination was moderately cost-effective in preventing hospital admission due to invasive pneumococcal disease in persons 65 years of age or above. In Sweden the cost was approximately 300,000 SEK per quality adjusted life years (QALY) gained, but only about 60,000 SEK per QALY in a two-way sensitivity analysis making reasonable assumptions regarding the incidence and mortality of invasive pneumococcal disease in this age group. On the basis of these findings, pneumococcal vaccination should be recommended for all persons 65 years of age or older. PMID- 11116893 TI - [Gender in the brain. A critical scrutiny of the biological gender differences]. AB - Down through history, biological arguments have often been used to legitimize a social gender order characterized by male supremacy. In the 1990's, a lively debate on the biological grounds of gender differences once again emerged in various fields. In the present article, the biological models used for explaining cognitive and behavioral gender differences are scrutinized, and recent research is discussed in light of history. These biological models emanate from theories about sex hormones, genetics and brain anatomy. Regarding the cognitive effects of sex hormones, no consensus has been reached, indicating a need for further research. Studies of relationships between genetics on the one hand and sexual orientation and behavior on the other are theoretically obscure and have thus far failed to prove a trustworthy connection. While there is indeed a difference in total brain size--men's brains are heavier than women's--it is not known whether this difference has any import beyond the fact that men have larger bodies. The existence of differences in brain lateralization and the size of the corpus callosum have been powerfully dismissed in several recent reviews. The design and interpretation of medical research in this field are still colored by gender stereotyped preconceptions and expectations, which obstructs efforts to gain a solid understanding of the biological differences/similarities between men and women. The media's interest in publicizing research results on gender differences, irrespective of magnitude or practical significance, further alerts us to the importance of scientific reason. There exists a very real risk today that medical gender research may be reduced to research about differences. If this problem is not addressed, it might lead to the reinforcement of the gendered structures of society. PMID- 11116894 TI - [Treatment of whooping cough between heaven and earth]. PMID- 11116895 TI - [Norwegian guidelines for "sore throats" nothing for Swedish throats!]. PMID- 11116896 TI - [A reply: our guidelines are useful also for Swedish throats]. PMID- 11116897 TI - [Comments from STRAMA: Contradictory Norwegian guidelines for the treatment of tonsillitis]. PMID- 11116898 TI - [Mammography screening at the cross-roads--a proposal concerning the information to women]. PMID- 11116899 TI - [Private contra public--final reply 1: the content is more important than the form]. PMID- 11116900 TI - [An explanation: evidence-based medicine is not a precipice for insufficiently documented activities]. PMID- 11116901 TI - [Final reply on drug handling in home care services: the guidelines should have been worked out together with specialties]. PMID- 11116902 TI - [Ontogenetic changes in the photosynthetic apparatus and effect of cytokinins]. AB - Data on structural and functional changes in the photosynthetic apparatus of agricultural plants are presented. Results of studies on the regulatory effects of cytokinin-like phytohormones on photosynthesis are discussed. Cytokinins were found to be involved in the structural formation and maintenance of the photosynthetic apparatus, conditions of the stomata and the supply of CO2 to carboxylation sites through the leaf mesophyll, the syntheses of pigments and enzyme systems, and the regulation of photoreduction and carbon metabolism. Possible mechanisms mediating the regulatory effects of cytokinins are discussed. PMID- 11116903 TI - [The 25th anniversary of the State Scientific Research Center for Applied Microbiology. Achievements and prospects]. AB - State Research Center for Applied Microbiology was founded in 1974 for accelerated development of molecular biology and molecular genetics in the USSR and rapid practical application of achievements of these sciences to economy and medicine. PMID- 11116904 TI - [Microbiological degradation of asymmetrical dimethylhydrazine--a toxic component of rocket fuel]. AB - A possibility of microbiological cleaning of water and soil polluted with asymmetric dimethylhydrazine (ADMH), a highly toxic rocket fuel ingredient (RFI), was studied. Several isolates (bacteria, yeast, and micromycetes) capable of utilizing ADMH as the only source of nitrogen, carbon, and energy were isolated from RFI-polluted tundra soil. Acceleration of RFI biodegradation was achieved using a biosorbent that involved cells of the degrader strain immobilized on granulated activated carbon. Biological testing in Escherichia coli and cereals (wheat and barley) demonstrated that biodegradation significantly decreased the integral toxicity of solutions containing ADMH, suggesting its utility for microbiological cleaning of polluted territories. PMID- 11116905 TI - [Initial stages of steel biocorrosion]. AB - Initial stages of corrosion of mild steel induced by Klebsiela rhinoscleromatis BO2 were studied in various media. The effect of the microorganism was detected 8 10 h after inoculation. The number of viable cells were virtually unchanged within one month in all media, but the corrosive activity of the strain decreased. The corrosive activity of microorganisms can be determined by spectrophotometry even only after incubation for 24 h. At a low level of organic substrate, even strong colonization with microorganisms does not inevitably result in a significant damage to metals. PMID- 11116906 TI - [Microbial degraders of some organochlorine compounds]. AB - A possibility of isolation of microorganisms, potential destructors of chlorinated organics from aged Vietnamese soils polluted with dioxine-containing defoliants was demonstrated. As an example, the ability of one isolated strain to metabolize pentachlorophenol and 2,4-dichlorophenoxyacetic acid was shown under laboratory conditions. An attempt was made to identify intermediates of pentachlorophenol metabolism using HPLC. PMID- 11116907 TI - [Effect of space flight conditions on properties of hydrocarbon-oxidizing bacteria]. AB - Results of experiments on the Mir space station (EO-25 and EO-26) demonstrated that the conditions of orbital flight, primarily the cosmic radiation, was a mutagenic factor affecting both the genotype and phenotype of an oil-oxidizing bacterial strain, Mycobacterium flavescens EX-91. The emerging mutants differed from original culture by the rate of colony growth and the ability to ferment certain carbohydrates or synthesize beta-galactosidase. Changes in the rate of utilization of raw oil and individual hydrocarbon types (constituting model mixtures) suggest that cosmic radiation may serve as a means of obtaining mutant clones of microorganisms with new properties. PMID- 11116908 TI - [Plant biotests of soil and water, polluted with petroleum and petroleum products]. AB - Reactiona of higher plants (mustard, oat, rye, salad, dill and barley) and microalgae (Euglena gracilis) on the contamination of soil and water with petroleum and oil products was studied. The germination of seeds was analyzed. The length of sprouts, dry biomass and length of plant roots, as well as the optical density of micro-algal broth culture were determined. Negative effects of soil and water contamination with petroleum and oil products on plant and microalgal parameters examined was shown. After biological destruction of contaminants by an association of destructor strains (Acinetobacter sp., Mycobacterium flavescens and Rhodoccocus sp.), the toxicity of contaminated mediums decreased. The data suggest that the integral toxicity of soil and water contaminated with petroleum and oil products and toxicity change during biodestruction of these pollutants can be analyzed by using plant test organisms. PMID- 11116909 TI - [Study of the integral toxicity of aqueous media, polluted by petroleum and petroleum products using bacterial tests]. AB - A biotest kit was used to assess the integral toxicity level of aquatic medium contamination with petroleum and petroleum-based products. The integral toxicity dynamics was also monitored during biodegradation of petroleum and petroleum based products by an association of petroleum-degrading strains including Acinetobacter sp., Mycobacterium flavescens, and Rhodococcus sp. The following bacterial tests were used: the bioluminescence (BL) test based on Photobacterium leiognathi; electro-orientation (EO), optoosmotic (OO), and growth test; as well as the reducing activity (RA) test based on the Agrobacterium radiobacter culture. No significant increase in the integral toxicity level of aquatic medium was observed when diesel fuel and kerosene contamination had been subjected to biodegradation. Although express biotests (EO, OO, RA, and BL) detected a pronounced increase in the integral toxicity of aquatic medium, long-term growth biotest revealed no statistically significant increase in the toxicity level. PMID- 11116910 TI - [Development and testing of "Ekosorb" biosorbent based on association of petroleum-oxidizing bacteria for purifying petroleum-polluted soils]. AB - A biosorbent containing an association of oil-oxidizing bacteria as a main constituent was developed, in which Lessorb, a product of moss and wood thermal processing, was used as a carrier. Xeroprotectors preserving the cell viability and oil-oxidizing activity in the biosorbent on drying and after long-term storage were selected. The use of this biosorbent for cleaning oil-polluted sod podzol soils showed a two-threefold cleanup rate acceleration at different pollution levels (8 and 24 l/m2), especially in the presence of a nitrogen phosphate fertilizer. The biosorbent increased the populations of certain groups of soil microorganisms and the total soil biological activity. PMID- 11116911 TI - [Creation and use of a liquid substance based on association of petroleum oxidizing bacteria]. AB - An association of four bacterial strains with high oil-oxidizing and bioemulsifying activities, psychrophilicity, resistance to chemical pollutants, and lack of pathogenicity was selected from a collection of natural oil-oxidizing microorganisms. A new liquid preparation containing stabilizers and preservatives that maintain the cell viability and oil-oxidizing activity during long-term storage was developed. A field experiment in oil-polluted sod-podzol and clay sand soils demonstrated that this preparation accelerated the biodegradation of oil and its individual fractions, especially in the presence of mineral and organic fertilizers. Treatment of oil-polluted soil with this preparation and additives decreased the oil-induced suppression of certain groups of soil microflora. PMID- 11116912 TI - [Microorganisms as possible indicators of soil pollution by dioxin-containing defoliants]. AB - The diversity of microorganisms from soils treated in the past with various dosages of dioxin-containing defoliants was studied. Population alterations dependent on dioxin concentrations were elucidated. Soil fungi and, to a smaller extent, actinomycetes were found to be the most sensitive to dioxins. PMID- 11116913 TI - [Development of biological methods of combating steel corrosion, induced by aerobic microorganisms]. AB - Cocultivation of degrading microorganisms and their antagonists decreases the corrosion loss of carbon steel by 20 to 80%. It was found that a microorganism can either accelerate or inhibit corrosion, depending on the nutrient. The magnitude of the effect on corrosion depends on the ability of the microorganism to respond to changes in the nutrient-medium composition by releasing acidic or alkaline metabolites. PMID- 11116914 TI - [Chemical-microbiological diagnosis of stress-corrosive damage to pipelines]]. AB - Samples of soil, ground, electrolyte, corrosion products, and protective coating were taken after excavating pipelines. The depth of stress corrosion cracks of the pipe steel was mostly related to the numbers of sulfate-reducing and denitrifying bacteria. In certain types of soil, damage correlated with the number of acid-producing microorganisms and aerobic chemoorganotrophs (saprophytes). A correlation was found between the extent of stress-corrosion damage to pipelines and the contents of reduced iron, sulfides, and organic carbon in surrounding ground. PMID- 11116915 TI - [Increase in the ecological danger upon the use of biocides for fighting corrosion induced by microorganisms]. AB - Five synergistic combinations of biocides were found, among which the combination of kathon + copper sulfate was the most efficient against Serratia marcescens. Depending on the ratio of these biocides, the synergistic effect of this pair allowed 4-20-fold decreases in the effective concentrations. Combinations of biocides with salts (carbonates and phosphates) that facilitate passivation of steel were found, which considerably decreased the corrosion losses of mild steel in comparison to isolated treatment with biocides or salts. The data showed that biocides must be added to corrosion-prone systems simultaneously with the beginning of their exploitation. Otherwise, considerably excessive amounts of biocides or their synergistic compositions are needed. PMID- 11116916 TI - [Direct quantitative method for evaluating the effect of biocides on Pseudomonas fluorescens in different culture media]. AB - A method for quantitative evaluation of the effects of biocides is presented. The method was tested in experiments with Pseudomonas fluorescens grown on various agar nutrient media. The effective concentrations of biocides that decreased the maximum specific rate of the colony biomass growth (mu'm) were called S (suppressing) concentrations, and concentrations that decreased the number of colony-forming units (CFU) were taken as L (sublethal) concentrations. The efficiency of the reported approach was demonstrated in experiments with three biocides tested in four nutrient media. It was found that the biocide sensitivity of Pseudomonas fluorescens varied by a factor of 30, depending on the amount and the type of the nutrient substrate. PMID- 11116943 TI - [Cervical emphysema caused by Forestier's disease]. AB - Diffuse idiopathic skeletal hyperostosis is a rare cause of dysphagia, preferentially affecting older men. This condition is also known as Forestier's malady, being characterized by a paravertebral ossification of 4 contiguous vertebrae at least. Seldom is the last etiology evoked for high dysphagia after other possible diagnosis exclusion. The diagnosis is confirmed by standard or contrast radiography. Introductory treatment is medical, being the surgery indicated for serious or resisting cases. One case is reported with dysphagia as primary sign of Forestier's malady, and review of possible diagnostic exams and treatments. PMID- 11116944 TI - [Cordectomy in the surgical treatment of tumors T1 of the vocal cords. Review of our cases (1974-1990)]. AB - We decided to review the results of cordectomy in our surroundings over the 22 years of existence of our Service. 631 clinic histories of patients undergoing surgery for laryngeal cancer in our Department, between 1974 and 1990, were reexamined, and from those selected 56 pertaining to T1 tumors treated with cordectomy. In this reduced group were studied epidemiology, clinical and pathologic data, complications, survival and death causes as well. The 58 considered were of male sex, middle aged (58-86) excepting 9 patients under 50. Their relation with tobacco eas clear, although less than in our general series (87.93% smokers). Of the 58 patients group 4 of then disappear during the 5 years follow-up term (8.62%). Three died: one from ganglion disease, other from local recurrence and the last one from other condition. In brief, global survival are accounted for 94.34% and the adjusted survival was 96.1%. PMID- 11116945 TI - [Lingual abscess]. AB - The unilateral abscess of the anterior portion of the tongue is an extremely rare occurrence. We report a case in which we have used the ultrasonography in the diagnosis, and discuss, as etiologic hypothesis the participation of Blandin-Nuhn glands in the genesis of these abscess. PMID- 11116946 TI - [Surgery of cancer of the larynx. Analysis of the results of our cases]. AB - After 20 years of existence of this Department we decided to carry out a review of the characteristics of our series and also of the results of treatment, according to the protocol elaborated and followed in agreement with the Tumour Committee if the Hospital. 631 medical histories of patients diagnosed and operated for cancer of larynx between 1974 and 1990 were reviewed. 83 of which (13.154%) disappeared during the 5 year follow-up period. Epidemiological, clinical, location, extension and treatment fdata were considered as well as the survival results and a single descriptive statistical analysis performed. The mean age of our study was 59.02 years, showing a clear relation to exposure to tobacco (91.44% were smokers and 63.39% of more than 20 cigarettes per day). The predominant location of the growth was supraglottic (64.05% of cases) followed by glottic in 33.76%. The most frequently used surgical technique was the total laryngectomy and total and extended to either the pharynx or tongue basis in 76.28%, followed by supraglottic laryngectomy in 13m5%. The stages were rather advanced with predominance of stage III /40.60%) and stage IV (21.17%). Global survive rate of our series accounted for 68.61% while the adjusted survival rate was 71.4%. PMID- 11116947 TI - [Neurinoma of the cervical hypoglossal nerve. A clinical case and review of the literature]. AB - Neurilemmomas are relatively rare tumors derived from Schwann cells that may occur at any part of the body. Many are encapsulated and they are more frequently encountered in adults. A neurilemmoma of the hypoglossal cervical nerve, an unusual site, is reported. Submaxillary angioma was initially suspected. PMID- 11116948 TI - [Ambulatory septoplasty]. AB - The AA. admit that the septoplasty under general anaesthesia is a surgical procedure that can be realized ambulatory, without added risks to patient. With viewpoint they have done a retrospective survey of the whole group of septoplasties performed under general anaesthesia at Torrelavega Hospital (Santander) in two years, between April 1995 and August 1997. The total number of cases amounted 56 (49 men and 7 women), lasting the procedure, more or less, one hour; postanaesthesia recovery also about 86 minutes and the hospital permanence about six hours and a half. Readmitted were 14 cases, because vomiting (7), for laryngeal spasm (2), bleeding (1), retention of urine (1) and other because vertigo (1), fever (1) and headache (1). PMID- 11116949 TI - [Mucinous cystadenoma of a minor salivary gland of the nasal fossa]. AB - Tumors of minor nasal salivary glands are relatively sparse. The case reported is a monomorph adenoma of benign character, which first clinical diagnosis mimicked a naso-sinusal polyposis. Examination of the removed piece by functional endoscopic surgery firstly was considered as an adenocarcinoma of low malignity degree but afterwards resulted as a mucinous cysto-adenoma of minor salivary glands with favorable clinical behaviour. PMID- 11116950 TI - [Verrucous carcinoma: retrospective study of 8 cases]. AB - Clinical findings, histopathology, therapeutic management and evolution of 8 cases of laryngeal verrucous carcinomata, treated at Virgen de la Salud Hospital from 1981 till 1997, are discussed. With the excuse of perform a retrospective perusal of our series, the AA. review the medical literature regarding possible new questions about this uncommon growth: as radiotherapy versus surgery and aetiological link of the tumor with the human papilloma virus. PMID- 11116951 TI - [Treatment of laryngeal stenosis with laser]. AB - Laryngo-tracheal stenosis are conditions in which the treatment remain under discussion and sometimes is far from disappointing. Our results are commented and literature perusal of the subject as well. PMID- 11116952 TI - [Type I Chiari malformation as a cause of inferior vertical nystagmus]. AB - Down beat nystagmus is a central nystagmus. In the literature reviewed the two most common causes that can produce it are cerebellar degenerations and Chiari malformation. The site of lesion causing this nystagmus appears to be the brainstem or the cerebellum, although the definitive etiopathogeny remains controversial. We report the case of a 73-year-old woman with sudden unsteadiness of gait and down beating nystagmus, without any other vestibular or neurologic signs. Simple radiology and MRI evidenciated a Chiari malformation type 1 associated to plastybasia. PMID- 11116953 TI - Antigen recognition by human gamma delta T cells: pattern recognition by the adaptive immune system. PMID- 11116954 TI - Tissue distribution, antigen specificity and effector functions of gamma delta T cells in human diseases. PMID- 11116955 TI - Antigen-recognition properties of murine gamma delta T cells. PMID- 11116956 TI - Human gamma delta T lymphocytes in HIV disease: effector functions and control by natural killer cell receptors. PMID- 11116957 TI - A role for epithelial gamma delta T cells in tissue repair. PMID- 11116958 TI - Intraepithelial gamma delta T lymphocytes: sentinel cells at mucosal barriers. PMID- 11116959 TI - gamma delta cells in gut infection, immunopathology, and organogenesis. PMID- 11116960 TI - gamma delta T cells in autoimmunity. PMID- 11116961 TI - Alternative techniques in trochanteric hip fracture surgery. Clinical and biomechanical studies on the Medoff sliding plate and the Twin hook. AB - In allowing compression along the femoral shaft (uniaxial dynamization) and optional compression along the femoral neck (biaxial dynamization), the Medoff sliding plate (MSP) represents a new principle in the fixation of trochanteric hip fractures. The Twin hook with 2 apical hooks was designed as an alternative to the lag screw. In 3 prospective consecutive case series and 1 prospective randomized study together comprising 342 trochanteric fractures, these alternative techniques were investigated. 3 postoperative fixation failures occurred in the unstable intertrochanteric fractures treated with biaxial dynamization with the MSP (n = 194), and 5 in those treated with the sliding hip screw (n = 62) (p = 0.04). A mean femoral shortening of 15 mm with the MSP and 11 mm with the sliding hip screw was found (p = 0.03). More medialization of the femoral shaft occurred with the sliding hip screw (26%) than with the MSP (12%) in patients with marked femoral shortening (p = 0.03). 3 postoperative fixation failures occurred in subtrochanteric fractures treated with uniaxial dynamization (n = 29) and 2 in those treated with biaxial dynamization (n = 19). Medialization of the femoral shaft occurred in 9 of the 19 biaxially dynamized fractures. The Twin hook was used in 50 patients and appeared to provide similar fixation stability as the lag screw. Biomechanical tests confirmed improved stress transmission over the fracture area with the MSP compared to the sliding hip screw in intertrochanteric fractures, and similar fixation stability with the MSP and the Intramedullary Hip Screw in subtrochanteric fractures. In axial and torsional loading, the Twin hook demonstrated gradually increasing resistance to migration. With the lag screw, the peak load was higher, but after migration with failure of the support by the threads, the loads were similar. Biaxial dynamization with the MSP appears to control fracture impaction effectively and minimizes the rate of postoperative fixation failure in intertrochanteric fractures. In subtrochanteric fractures, uniaxial dynamization prevents medialization of the femoral shaft and is therefore preferred to biaxial dynamization. The Twin hook appears to provide adequate fixation stability, and with potential for simplified intraoperative handling and reduced dissection, the Twin hook may pose advantages compared to the lag screw. PMID- 11116997 TI - The Oswaldo Cruz Institute and its importance in the Brazilian society. Perspectives for the 21st century. PMID- 11117000 TI - The uphill battle for higher Pap smear reimbursements. How the test was won. PMID- 11117001 TI - Therapeutic plasmapheresis for autoimmune disease. Advances and outcomes. PMID- 11117002 TI - The Stark labyrinth. Navigating your way through fraud and abuse legislation. PMID- 11117003 TI - FutureScope: scanning the horizon for environmental and|| technical changes. PMID- 11117004 TI - The HIPAA clock is running: final HIPAA transactions and|| code sets standards published. PMID- 11117005 TI - Cross functional team work; the challenge of complexity and|| diversity. PMID- 11117006 TI - [Stereologic analysis of mammary glands in primiparous rats with lead poisoning]. AB - The mammary glands of primigravid Wistar rats were investigated by stereological analysis in conditions of lead acetate administration via drinking water. The experiment was made in early summer period when absorbing ability of rat intestinum for lead is highest. Morphological state of glands was observed on 7th, 14th and 21st day of pregnancy. Groups of control animals were drinking deionized water but groups of experimental animals were drinking deionized water with added lead acetate. On paraffin sections coloured by HE method, volume density (VV) of glands structural elements were determined by multipurpose test system M42. Stereological analysis shows that lead diminishes the proliferative capacity of the mammary parenchyma and changes the morphofunctional differentiation of the overall structure in mammary gland during pregnancy. At the same time lead modifies quantitative trait of the mammary gland, i.e. the composition of the volume units in organ, without changing intramammary syncornization between the epithelium and lipocytes. PMID- 11117007 TI - [Variation in the lateral plate of the pterygoid process and the lateral subzygomatic approach to the mandibular nerve]. AB - OBJECTIVES: The objective of this work is studying wariable anatomy formations at the posterior border of the pterygoid process lateral plate of the sphenoid hobe (lig. et foramen pterygospinale, lig. et foramen crotaphitico-buccinatorium), and its statistical processing considering sex, age and sides of studied sculls. MATERIAL AND METHODS: At 305 sculls of mature persons of both sexes (167 males and 138 females) we studied incompletely ossificated lig. pterygospinale, opening of the same name which has become by its full ossification, incompletely ossificated lig. crotaphitico-buccinatorium and opening made by it's complete ossification, and position of this structures in relation to foramen ovale. RESULTS: Incompletely ossificated lig. pterygospinale was found at 12 sculls on both sides or 3.93% (8 male's and 4 female's sculls). One-sidedly, on the right one, incompletely ossificated lig. pterygospinale was found at 14 sculls (4.59%), and on the left side at 19 sculls (6.22%). Incompletely ossificated lig. pterygospinale, present on both sides of sculls, was in the case of persons older than 50 (9 sculls or 75%). Pterygospinous opening which has become by complete ossification of lig. pterygospinale was found on both sides at 4 sculls (1.31%), two of them were male's and two of them female's sculls. In alll four cases persons were older than 40. Pterygospinous opening found on the left side was present at only 3 sculls (0.98%) and on the right side at 4 sculls (1.31%). Pterygospinous opening was present one-sidedly, every time, in the case of male's sculls. Incompletely ossificated lig. crotaphitico-buccinatorium, meaning incomplete opening, was found at 17 sculls on both sides (5.57%). 10 of them were male persons' sculls, and 7 of them females'. On the right side, incompletely ossificated lig. crotaphitico-buccinatorium was found at 11 sculls (3.60%)--6 male's and 5 female's, and on the left side at 16 sculls (5.24%). Foramen crotaphitico-buccinatorium was found on both sides only at 12 sculls (0.97%)--one male's and one female's. Foramen crotaphitico-buccinatorium was found on the left side at 8 sculls (3.90%) and, on the right side, this opening was found at 3 sculls (0.98%). Incompletely of completely ossificated lig. pterygospinale, located laterally from foramen ovale, was found at 5 sculls (5 male's and 1 female's) or 1.63%, and in the other case, incopletely or completely ossificated lig. crotaphitico-buccinatorium laterally from foramen ovale was found at 7 sculls (5 male's and 2 female's) or 2.29%. CONCLUSION: While applying conductive anaesthesia on mandibular nerve by lateral subzygomatic route, variable ossificated formations at lateral plate's posterior border of pterygoid process should be kept in mind. It is possible that a needle, at a depth of 35 mm, comes across incompletely or completely ossificated lig. pterygospinale et crotaphitico bucciantorium. Based on ours anatomy research we concluded that these formations are more oftenly present on males aged over 50, and that one-sided presence is more often on the left side. Ossificated formations which were the object of our research are not an obstacle to high-quality applying of conductive anaesthesia on trigeminale ganglion et mandibular nerve each time when they exist, but only when they are located laterally from foramen ovale, what appeared at 12 sculls (3.93%) in our case. PMID- 11117008 TI - [Effect of starvation on blood protein levels in the population of Dobrinja (1992 1995)]. AB - In nutritional protein deficiency, numerous studies verified utilization of amino acids generated from tissue degradation in intensive protein synthesis. Unlike liver, muscle protein synthesis is extremely dependent on external supplies of essential amino acids. Prolonged nutritional protein deficiency results in decrease of body weight as well as total protein concentration, in particular in early days of starvation. In prolonged starvation during the war, significant decrease of body weight was registered in 70 subjects while their total protein concentration remained within the expected range and did not significantly differ the values recorded in the control group. Concentration of serum albumines in the control group was lower than the concentration recorded in the tested group, while the serum globulins concentration was higher in the control group. Although the difference in body weight between the tested and the control group was statistically significant, no significant difference in the concentration of total proteins, albumines and globulines was recorded. PMID- 11117009 TI - [Typing of isolated mycobacteria]. AB - Microbiological diagnosis of causes of pulmonic or other tuberculosis localization includes microscopic analysis of disease material cause which was taken earlier, his specific elaboration and inoculation at appropriate bases. Bases are incubated until 12 weeks. In the purpose of identification of M. tuberculosis complex or some other (conditionally) potentially pathogenous king of micobacterium, depending on immunity status of the investigated one, it is necessary to apply conventional as well as modern methods of molecular biology in laboratory work. First of all this means differentiation of M. tuberculosis complex from other bacterium (MOT), that is eventually differentiation inside complex. Beside classical tests it is possible to apply some methods of molecular biology in laboratory conditions, as well as nucleus sonds or method for reproduction of one part of nucleus acid which fragments, with a help of restrictive encimes, into several parts (PCR/PRA). With this method it is possible to do complete identification of isolated kinds of micobacterium according to the size of aplicon's fragments (part of reproduces DNA). An appropriate time, which is necessary for execution of these tests, is measured in hours and days, and for the classical tests in weeks and months. PMID- 11117010 TI - [Use of antimicrobial drug prescriptions in children with a common cold or diarrhea]. AB - There are justified reasons to assume that antibicrobial drugs are too frequently prescribed in routine pediatric practice. The main aim of the study was to prove the frequency of antimicrobial administration to children. A total of 209 children of either sex and under 18 years of age with common colds and 95 children with diarrhea who were examined at the Primary Health Care Department of Hospital Konjic, Bosnia and Herzegovina for the period of Jan 1999-Apr 2000 were investigated for antimicrobial therapy. Antimicrobials were prescribed to 175 (83.73 per cent) patients with common cold and 78 (82.11 per cent) patients with diarrhea. Taking into consideration that the aetiology of these conditions was predominantly viral, the outcome of this study provides the evidence of irrational administration of antimicrobials and states the possible ones: diagnostic difficulties in routine pediatric practice, low educational level of physicians, lack of consulting skills, overworked primary health care physicians, influence of inpatient parents about prescribing antibiotics and influence of pharmaceutical industry. It is widely known that the negative effects of such superflous administration could leave numerous consequences to the patients health condition. Besides that, irrational use of the above mentioned drugs financially charges health system funds. It is therefore necessary to enhance consulting skills, to improve research and evidence based guidelines and monitoring strategies as well in order to support rational antimicrobial administration to children. The observation that irrational use of antibiotics was detected frequently with both common cold and diarrea suggests the need for continous monitoring and analyses of antibiotic administration to children from our region. PMID- 11117011 TI - [Iodine deficiency in the Federation of Bosnia and Herzegovina]. AB - Iodine deficiency which causes the wide spectrum of disorders for all ages, is one of the significant public health problem worldwide. From the ancient times different iodine deficiency disorders were noticed in Bosnia and Herzegovina and in its some areas the goiter existed in endemic form. These facts confirm that its soil bas been iodine deficient and that necessity for iodine prophylaxis is obvious on its territory. The study was based on 5,523 children, of both sex boys and girls school age from 7 to 14 years, randomly selected with the equal participate subjects in relation to the age. The sample is representative and it has been assessed based on: total number of school children aged from 7 to 14 years, anticipated prevalence of goiter 5% level of probability 95%, relative punctuality 30% and the factor called "design effect" which is 3. The study was carried out in whole ten cantons in the schools with equal representation among cities and villages. In examining of prevalence of giter we used inspection and palpation. Determination of iodine concentration in urine was carried out by the method is based on Sandel-kolthof's reaction. The technique used for determination of concentration of iodine in salt was iodinemetric titration. The prevalence of goiter was 27.6% in Federation of Bosnia and Heryegovina. The highest prevalence of goiter was in Bosnia Podrinje Canton (51.20%) while the lowest was in West Herzegovina Canton (12.90%). The urinary iodine excretion in investigated children varied from 1 to 208 *mg/L with median of 77.6 *mg/L. Iodine contetn in household salt samples was from 3 to 29.8 mg/kg, range 14.4 + 5.9 mg/kg. The results of our study show the persistence of mild to moderate iodine deficiency in Bosnia and Herzegovine Federation. Therefore according to the recommendations of the World Health Organisation, UNICEF and International Council for Control of Iodine Deficiency Disorders, the salt for human, and animal consumption as well as for food industry which is consuming on its teritory, has to be iodinated on the place of its production without looking back whether or not it is produced or imported in Bosnia and Herzegovina Federation, lodination has to be performed with 20 to 30 mg KI per one kg of salt, thereby an average the iodine content has to be 25 mg per kg. In this way it will be prevented the wide spectrum of disorders, which we often are not aware for that its are caused by iodine deficiency. In addition it will be prevented many very important socioeconomical consequences of iodine deficiency. PMID- 11117012 TI - [MRI of the spleen]. AB - PURPOSE: The aim of this study is to show the possibilities of MRI in diagnostic of the spleen affections. MATERIAL AND METHODS: During a two years period, MRI of the splen alone was performed in 13 patients, while MRI of the liver was performed in 213 patients. With the liver, we examined the spleen as well. Using 1.0T Unit (Siemens Magnetom Impact) we performed next sequences: T1W SE, T2W TSE FS, PD T2W, with and without breath hold. Rutinly we used body coil. We divided all patients in two groups: 1st group--13 patients undergoing spleen examination, and 2nd group--213 patients with liver and spleen examination with retrospective analysis. RESULTS: In the first group--10 patients, MRI of the spleen provided us with some new information compared with CT and US. In three patient there were no new diagnostic information. In second group, accidently showed cystic postraumatic lesions, there was trombotic aneurysm of the lienal artery. Also in the patients with liver disease spelnomegaly was found in 25. DISCUSSION: According to the literature and our initial experience, MRI of the spleen is not yet in wide clinical practice because of the little difference of the relaxation time of the spleen and proton density. Introducing of the paramagnetic contrast media will provide increasing of abilities in the detection of the tumor lesion. Further development advances towards diagnostic of the lymphatic infiltration, staging of metastatic carcinoma, and differential diagnostic of splenomegaly. CONCLUSION: According to our experience and literature, MRI of the spleen is not yet in wide clinical practice, for now. PMID- 11117013 TI - [Ebstein's anomaly with patent foramen ovale and supraventricular paroxysmal tachycardia]. AB - A case of asymptomatic 36 years old men with Ebstein anomaly of tricuspid valve, patent foramen ovale and paroxismal supraventricular tachycardia is presented in this paper. A routine physical examination revealed a heart systolic murmur along left and right side of sternum, with right bundle branch block on ECG. It led to the detailed non-invasive cardiologic examination (transthoracic, transeophageal and stress echocardiography, ambulatory ECG), what disclosed above mentioned Ebstein anomaly. Also he was operated on upon superior lip fissure at the first year of life. Regarding his very mild symptoms (NYHA g. 1), he was treated only medicamentosly. PMID- 11117014 TI - [Results of IVF treatment methods in Germany in 1998--the German IVF Registry]. AB - We used data in Germany IVF registar 1998 from 86 (94.6%) centers which take upon IVF procedures. The total documentary were treated 46730. The cultivated data were 45459. The visible is increase number of group which use IVF procedures and growth IVF/ICSI centers with options KRYO preservations. In Germany 1998, IVF ICSI, KRYO and GIFT procedures were treated 30,009 women, on an average 1,515 cycles per year. The endurance leaning of children was amoung 2 and 8 years for both procedures. In stimulated cycles on an average were taken 9.04 eggs (follicule punction). During IVF procedures it was 22.64% clinical pregnancy and ICSI procedures 23.53% clinical pregnancy. In most of the cases cycles were stimulated with long gonadotropine protocol, with GnRH-agonists, after suppression adenohypophysis. Considering pregnancies and abortions in addiction to chosen gonadothropine it was not possible to confirm significant difference neither for IVF of ICSI procedures. Results of therapy showed clear addiction to age of women and become worse after 39 the year of life and distintcly less after 49th year for both procedures. During transvaginal ultrasonography follicule punction reported is 253 (0.67%) complications. Severe cases such as ovarial hyperstimulation syndroms (OHSS III) were reported in 1.12% cases. PMID- 11117015 TI - [Personal experience with use of the antimicrobial agent, Neloren R, in maxillofacial surgery]. AB - During the treatment of sick as well as injured and operated patients it was used the therapy according to the clinical experience of doctors and possibility of the choice of some antibiotic. It was used the therapy according to antibiogram whenever it was possible. In the study the results of using of Neloren antibiotic, linkomicin, produced in "Bosnalijek" Sarajevo, were followed up comparing to the other antibiotics. Neloren effects on Streptococcus pyogenes, Streptococcus penumoniae, S. viridian's, Staphylococcus aureus (resistant on Penicillin), Corynebacterium diphteriae, Bacillus anthracis Bateroides fragilis, Rikecia prowaseki and some dostridiaes. In developing of the study the patients were involved who had bone fracture caused by influence of different exogene ethiological factors and the patients with benign pathological process in bone structure, the most often osteomyelitis or abscess caused by dentogene source as well as the patients who were treated by functional or corrective aesthetic operations such as osteoplastic of jaw or correction of bone deformation. The results were shown by chart and graphic representation and they will be compared to the results around the world. PMID- 11117016 TI - [Stuttering and left-handedness] . AB - The main purpose of this study was to determine the incidence of left handedness in 380 stuttering children and adolescents and also to examine the differences in variables of stuttering severity between the left- and right handed subjects. The results showed that the incidence of left-handedness in stuttering children was not significantly different from normally fluent children. The difference in variables of stuttering between the right- and left-handed subjects was determined using the t-test. The results obtained revealed that the difference between the left- and right-handed subjects was not statistically significant in variables of stuttering severity. PMID- 11117017 TI - [Post-traumatic stress reaction in children]. AB - In this work is presented most frequently traumatic experience that had children during the war, level of traumatization, and discovery most frequently post traumatic stress reactions. The works covering are school children age until 7 since 15 year with polytraumatic stress experience. PMID- 11117018 TI - [Types of changes in laboratory technics used in prosthetic technology]. AB - Because of the phenomena which were noticed in clinical and laboratory work of permanent stomatological metallographical and other researches of the consequences of free changed conditions in melting and cooling phase of dental alloy auropal were carried out. Modality of the changes during melting of this alloy in dental laboratory were systematically chosen according to noticed phenomena in praxis. Main objective was to metallographicly determine these phenomena during the process. With application of usual methods for setting of metallographical carved pie, samples were researched with metallographical microscope Epiryp 2 from Zeiss-jena, under different melting conditions and filmed with different enlargement. Even small changes of melting and cooling conditions are causing in microstructure and these are reflecting on the alloy characteristics. According to this melting procedure is a very sensitive and complex problem in termical laboratory processing which demands maximum attention an strict following fabrical instructions in order to avoid unwanted consequences in clinical application. PMID- 11117019 TI - [Anomalies in the location of the ulnar artery]. AB - In this report is described anomally of arteria ulnaris topography. By anatomical dissection of cadaver of a male new-born, we noticed the ulnar artery, on the right arm, layed superficially, under of fascia antebrachi. The ulnar artery was the branch of brachial artery, and it run directly from the brachial artery, in the level of interepicondilar line, to the madial part of the distal end of antebrachium, and it attached by ulnar nerve before they entered Guyon's canal. PMID- 11117020 TI - [Abu al-Qasim Az-Zahrawi--a great Arab surgeon] . AB - History is a witness of the great importance and influence of islamic science from the period of "Golden Age of Arabic Civilisation". A famous scientist said: "Science has no country, it is international; we all share in fruits of investigations of people from different traditions and all ages." There are many worldwide famous arabian scientists: El-Kindi, Er-Razi, Ibn Sina, El-Biruni, Ibn Hajsem, Ez-Zahravi, El-Farabi, Ibn Zuhr, Ibn Ruzd etc. These names, among several hundreds of arabian physicians, attribute "Golden Age" of islamic science. That period was characterised by movements, reprocessing of ideas. That reprocessing of ideas has gained the great minds together, and that process is continuous. That is why we have to be grateful to them. One of them, who is not enough famous within medical professionals, Az-Zahrawi, belong to the greatest surgeons in the arabian "golden age" period. PMID- 11117021 TI - [Emergency medicine in Bosnia-Herzegovina--new trends]. PMID- 11117022 TI - [Reforms and modernization in the emergency medical care system in the Federation of Bosnia and Herzegovina]. AB - This paper reports on the result of an in-depth review by the Federal Ministry of Helath (MOH)/This review was undertaken by an expert committee appointed to review the delivery of emergency healthcare services in the context of the overall organizational structure of the country's healthcare system. Within their mission to reform health care system in Bosnia and Herzegovina. It was worked out by the expert commission for the development of the health care organizational scheme proposal, in part dealing with urgent medical services. The committee's followed the universal and well-documented principle that 1) The timely delivery of particularly pre-hospital emergency healthcare reduces unnecessary loss-of life and disability 2) The timely delivery of emergency healthcare has a positive impact on effectiveness and efficiency of the overall performance of the healthcare systems. A retrospective analysis of pre-hospital emergency services based on a systems analysis, procedures and experiences revealed the need for a system approach such as documented, for instance, by the National Highway Traffic and Safety Administration (NHTSA) "Agenda for the Future" in the U.S.A. A preliminary set of recommendations was made which requires further follow up, particularly through an international dialogue at Conference like these. Of paramount importance is the development of a systems approach to the delivery of Emergency Healthcare, both in the pre-hospital and in-hospital domain. This systems approach should be developed in the context of the overall infrastructure and healthcare system of BiH. When such an organizational concept is accepted as a "doctrine", then it will be possible to establish a truly national system, with -and this is of great importance--a set of minimum quality standards attached. PMID- 11117023 TI - [Emergency medicine today--quo vadis? (developments and perspectives)]. AB - The author analyses nowday condition of emergency medicine in Federation Bosnia and Herzegovina and all its components. He shows all negativenesses in work according an unorganized and an uneffective system of "reaction and answer". In conclusion, it is given the suggest for benefit of this very important part of health care. PMID- 11117024 TI - The German approach to emergency/disaster management. AB - Disaster control and disaster relief in Germany are public tasks. But the government has shifted the responsibility of the administration of these tasks to the 16 states, the so called "Lander", because the EFG is a federal republic. The same is valid for the civil defense and the civil protection in the case of military or international risks. The 16 states are also responsible for the legislation of rescue service, fire fighting service and disaster control (natural and technical disasters). Counties and district-free cities are responsible for the organisation of these services. The German system is based on the principle of subsidiary between official and private institutions. A lot of official and private relief organisations are responsible for the execution of disaster relief tasks. In Germany the following organisations exist: Official (GO): Technisches Hilfswerk (THW/Federal Technical Support Service), Feuerwehren (Fire Brigades/professionals and volunteers) Academie of Emergency Planning and Civil Defense Private (NGO): Arbeiter-Samariter-Bund Deutschland (ASB/Workers' Samaritan Association Germany), Deutsche Gesellschaft zur Rettung Schiffbruchiger (DGzRS, German Lifesaving Association), Deutsches Rotes Kreuz (DRK/German Red Cross), Johanniter-Unfall-Hilfe (JUH/St. John's Ambulance), Malteser Hilfsdienst (MEID/Maltese-Relief-Organisation). ASB, DRK, JUH and MHD are specialised in the field of rescue, medical and welfare services and medical disaster relief. 80% of the German rescue service and 95% of the German disaster medical relief are realised by these NGO's. NGO's and GO's employ more than 1.2 million volunteers and appr. 100,000 professionals. Rescue service is carried out by professionals, disaster relief by volunteers. The German constitution allows to call the federal army in case of disaster, to support the disaster relief organisations (for example: flood Oder River 1997, train-crash "ICE" 1998). In all counties and district free cities disaster control staffs are set up by the administration. During disaster relief operations a operational command is on site. Most of the counties and district free cities, medical executives, rescue staff executives along with fire executive officers are responsible for the medical rescue organisation. All emergency physicians and medical executives have attended special training or a 520 hours-training-course (Paramedics). All volunteers of the medical service in the disaster relief organisations are trained in separate special courses (90 hours). Over the last years, civil protection, disaster relief and rescue services in the FRG have been reorganised. In 1997, the civil protection was reformed by a new federal act. Disaster relief of the "Lander" is supported by Federal Government with about 9000 vehicles and a budget for training. Emergency physicians have to take part in a (80) eighty hours lasting course on emergency medicine from an interdisciplinary point of view; they are only allowed to do rescue missions after having proved basic experience in emergency medicine as well as having completed a (18) eighteen-months postgraduate training period at least. Senior emergency physicians receive and additional (40) forty-hours-lasting theoretical and practical training-after three years practice in rescue services as a minimum. There are special training courses offered for Medical and Non-Medical Personal to cope with disaster situation by different institutions and organisations. PMID- 11117025 TI - [Problems in the development of the emergency health care system]. AB - Since February 1994, during and after 4-years supervision of American specialists, Emergency department of Zenica hospital has been trying to implement Anglo-American working system within the hospital framework (video). Principles of quality functioning of Emergency Medical Service (EMS) are based on: Population that depends on different demographic factors and prevention programme (education of population, quality functioning of health legislation). Pre hospital treatment depends on good quality communication and transport. Hospital treatment based on good functioning of Emergency department and Intensive care unit as well as proper coordination with other specialties. Proper implementation of items stated above depends on top-class teaching and compulsory periodical screening of attained knowledge and skills, properly organized communication, transport and technical equipment. Emergency medicine is not only a sum of urgencies from the existing conditions but a special medical discipline and it has special and unique approach to diagnosis and therapy of acute health disorders. Therefore, a main weakness in the development of emergency medicine is: no recognition of emergency medicine as a unique specialty in the most European countries, non-existing departments of emergency medicine at medical faculties, no unification of BiH emergency medicine system, undeveloped monitoring and development evaluation of emergency medicine etc. The World Association of Emergency Medicine should have an important role for emergency medicine recognition, and in its future development through links with health legislation and educational associations (ACLS, ATLS, APLS). PMID- 11117026 TI - [Organization of emergency medical care in Tuzla Canton]. AB - The main duty for all health institutions and individuals is an organization and emergency medical care providing. The aim of this paper is to analyse the existing staff, equipment, facilities and emergency medicine organisation in 13 municipalities of Tuzla Canton. In this paper we have analysed 13 emergency medical services in Tuzla Canton: 1. Tuzla; 2. Banovici; 3. Celic; 4. Doboj Istok; 5. Gracanica; 6. Gradacac; 7. Kalesija; 8. Kladanj; 9. Lukavac; 10. Srebrenik; 11. Teocak; 12. Sapna; 13. Zivinice. We have analysed the number of employed medical workers (doctors--specialists, general practitioners, senior medical technicians, medical technicians), ambulances and equipment (ECG, defibrillators, oxygen devices, intubation kits, aspirators) and communication system. Present situation is compared with standards and norms for every municipality, from our and heads' of units point of view, concerning equipment, staff and facilities for each unit. It is evident that the present situation is far from the needs concerning staff, equipment, facilities and emergency care organization in Tuzla Canton. PMID- 11117027 TI - [Structure and organization of emergency medical care in Tuzla Canton and possibilities of interdisciplinary cooperative projects]. AB - Aim of this paper is to provide substantial and organizational help to emergency medical service as a part of primary health care and other health care levels in the interdisciplinary cooperation. Projects of interdisciplinary cooperation, that could be implemented within BiH as pilot projects are proposed in this paper as well. Legal regulations, that serve as a base for the organisation and a scope of emergency medicine work served as a draft for elaboration. Legal base is found in the Constitution of Bosnia and Herzegovina and Federation of Bosnia and Herzegovina, Law on health care, Articles 3, 16, 20 and 59 in particular. We point out item 7 concerning temporarily standards and norms of health care. This item should be considered seriously by experts in this field and Public Health as well, and elaborate its revision and new proposals. The Law on the records in the health care regulates monitoring of the work of this service. 2 OB form should be reviewed in a way that gives us good quality monitoring, more significant data which will enable us to take better attitude toward the organization and work framework. The same should be done with individual data to facilitate a work concerning emergency medical procedures. There are 14 emergency medicine units in Tuzla Canton: 51 doctors are employed (37 general practitioners and 14 specialists). They provide emergency medical care for 537.000 inhabitants together with 114 other health workers with secondary and advanced education. In 1999, there were 216.162 check-ups by doctors, 2700 home visits and 29.705 patients were referred to specialists and laboratories for further treatment. Other health workers provided 584.432 services and 3.889 home visits These data will be especially discussed from the inhabitants' point of view concerning their needs, quality of services and record keeping forms for emergency medical service. The following projects are proposed for the future interdisciplinary co operation in emergency medical care providing: Education of inhabitants (especially certain population) for mutual help, self-help and first aid. Research about inhabitants' needs and demands concerning emergency medical care. Framework of emergency medicine organisation, required resources and having higher levels of medical care as a support. Prevention of emergencies through health promotion. Doctrinarian attitude toward the emergency medical service work, stage by stage treatment and a role of comfortable transport. Emergency situations in a mass disasters and ways of organization and scope of emergency medical service work and others in the interdisciplinary and intersectional co operation. Proposed projects are the result of our present knowledge about the medical service organisation in our country. The aim is to meet the needs of the population concerning emergency medical services in the interdisciplinary co operation in the health care system through realisation of these projects. PMID- 11117028 TI - [Emergency medical care in small urban regions] . AB - Emergency medical care is a health care segment which is essential one both in large and in small urban places. To be effective it is necessary to be well organised, and also technically equipped and properly staffed. Concerning the situation in this field, conclusions are very often made on the basis of large and very well organised units. Current legislation gave it a pretty significant place. In practice it was proven as an indispensable part of system that provides medical care for suddenly ill or injured patients, both in war and peacetime. During the war, out of total number of admissions, 5231 injured patients were treated (including cases of death), out of which 36% had just that treatment as a definite one while the rest had to be treated in hospital. In the post-war period (1996, 1997, 1998, and 1999), out of total number of 95.000 treated patients, 848 was injured in car accidents and 52% of them got a final treatment on the level of emergency medical service. However, medical amateurs but medical professionals as well put emergency medical care on a margin of evaluation. Certain number of patients is coming to emergency medical service to get prescription, be directed to specialist or simply they don't want to waist their time while waiting in the crowded surgery of general practitioner. The management does not give enough attention to staff, facilities or technical equipment of emergency medical service. Only by properly established emergency medical service we would have better and more effective subsequent stages in the treatment of suddenly ill and injured persons in large but even more in small urban places. PMID- 11117029 TI - [Problems with organization and operation of emergency medical services in the post-war period in districts with a population of up to 60,000 inhabitants]. AB - Demographic characteristics of Gracanica municipality:--Organization of the municipality on 15 local communities that are 5-20 km away from the centre of municipality.--Road conditions--Starting of industry within Gracanica municipality and social status of the employed population.--There are two main roads that are passing through the territory of municipality--These highways are passing through densely inhabited areas and traffic is heavy. Distance of Tuzla Clinical centre. Services of Gracanica Outpatient Department organize their work although lacking professional staff and equipment. Significance of Emergency Medical Service (EMS) for the municipality and broader community under these circumstances. Present EMCS organization in Gracanica is the following: EMS in Gracanica is organized as a separate service with three permanent teams and temporary engagement of 2 doctors who are on specialization in pediatrics and lungs diseases. A team comrising of a specialist of emergency medicine, three medical technicians and one driver works from 7.00 to 15.00 on regular working days and other doctors work at EMCS from 15.00 to 7.00 and on weekend as well. EMS doesn't examine patients outside the EMS rooms. EMS equipment consists of one ECG, one defibrillator, two aspirators, one oxygen concentrator and a resuscitation set. The Japanese Government donated equipment. Lack of intravenous solutions, drugs and other material for the emergency treatment is notified as permanent problem in emergency care service. Next problem is unsuitable and non functional space with difficult access and lack of ambulances because the whole rolling stock was destroyed by shelling in 1992. In any case, all these circumstances diminish a team efficiency in providing emergency care. Even under these circumstances EMS had 10,415 examinations during the first sixth months in 2000, provided 45,265 services and treated 912 injuries out of which 64 were traffic injuries. Concerning the complicity of work in EMS Gracanica, standards and norms related to emergency medical care, which have been proposed by the Federal Ministry of Health couldn't be met. By these standards one team covers 20,000 inhabitants and EMS from 19.00 one day until 7.00 the next day and Saturdays and Sundays as well. The question is what to do between 15.00 and 19.00, when every Outpatient Department is closed? For normal functioning of EMS in Gracanica municipality we should do more work on the prevention and work organization in the surgeries of General Medicine in a distant local communities. In that way, EMS wouldn't be a surgery of General Medicine after 15.00. It's necessary to ensure an adequate space, professional staff (at least 4 teams with their leaders) new equipment, ambulances, medicaments, disposable material and furniture. It is necessary:--to develop a system of communication tha could cover municipality and Clinical Centre area,--make educational plan and to respect it,- to establish a cooperation with EMS in neighboring municipalities,--to discuss EMS role in relation to family medicine organization within General hospital in Gracanica. PMID- 11117031 TI - [A single dispatch center and communication system]. AB - Dispatch centre is one of the main part of emergency medical system. The basic role's of dispatch centre are: emergency call, admission, trage, and coordination. High level of responsibility demands continuous education and training of dispatch centre personnel. PMID- 11117030 TI - [Development and organization of the Education Center at the Institute of Emergency Medical Services in Sarajevo]. AB - This paper presents an experience of the Educational Centre of EMSC Sarajevo in the postgraduate training of the various profiles of medical personnel (medical doctors and medical technicians) that are employed in the emergency services of the pre-hospital and hospital type in Bosnia and Herzegovina. A period of the last four years (1996-2000) when the Educational Centre was restructured, becoming a one of the most active services in the medical sector of the EMSC Sarajevo has been emphasised in particular. Educational Centre of EMSC was participating in a number of international projects related to the education. Forty-five (45) courses of the various levels and with different programs: BLS, ALS, ACLS, ACLS-instructor course, ATLS, EMT-course, EMT-advanced, EMT-instructor and EMT-dispatcher course were carried out in the mentioned period. Seven thousand eight hundred and twelve (7812) hours of theoretical teaching and practical training were provided for the 570 candidates who successfully completed training in various programs. First aid training for civilian population was also conducted. Twenty one first.aid courses were carried out and successfully achieved by 324 candidates. On this way, 196 hours of theoretical teaching and practical training were realised. In order to make first aid training popular, 160 children from pre-school institutions (kindergartens) of Sarajevo Canton were also introduced to first aid principles. It has been pointed out that well equipped and trained team for the urgent medical intervention with a necessary team work is a crucial factor for the successful treatment of emergency that means a patient in life threatening situation. PMID- 11117032 TI - Croatian experience with the landmines: deaths and injuries from the landmines in the area of Sisak during five-years period (1995-2000). AB - After the war in Croatia, thousands of landmines were left on the fields and because of this today we have many casualties. Area around Sisak is a region where a large number of landmines were laid and accidents are frequent. The authors of this article present results of retrospective analyses of accidents (death, injuries and type of injuries) which occurred after the military operation "Oluja" since August 1995 until March 2000. Data is collected from local hospital and police department in Sisak and compared with data obtained from Croatian Mines Action Center. PMID- 11117033 TI - [Treatment of paroxysmal supraventricular tachycardia in emergency medical care]. AB - This paper presents results of the Emergency medical service centre Sarajevo work concerning treatment of paroxysmal supraventricular tachycardia in the period from 1 January 1998 until 2 February 1998. It indicates the importance of monitoring in diagnostic procedures and therapy and a role of oxygen in this therapy as well. Efficiency of medical therapy in definite treatment of the patients is also presented. PMID- 11117034 TI - [Occurrence of atrial fibrillation and cerebrovascular insult in patients at the Emergency Center of the General Hospital in Konjic]. AB - Atrial fibrillation (AF) presents disorganized activity of atrium with irregular ventricular striking. It is observed in paroxysmal and persisting form. Paroxysmal form is more risky for embolism (about 6 times). Aim of this research was to determine frequency of atrial fibrillation as increased risk for cerebrovascular insult (CVI). We conducted our research in general hospital Konjic and all patients who were treated in the period 1998-1999 are included. The results of the research are the following: 1. Total number of CVI patients was 126 what compared with total number of hospitalized patients indicates increased prevalence of 8.8% in relation to world parameters (2-5%). 2. 55.7% CVI patients had verified persistent AF what also represents increased percentage compared to the world parameters. 3. The following cardiovascular diseases are involved in the etiology of AF where CVI was a result: *atherosclerosis of heart 84.9%, *coronary disease 15.6%, *heart malformations, 4.76%, *other diseases (hyperthyroidism 1.5%, idiopathic AF). 4. Incidence of other risk factors at the patients with CVI and AF: *hypertension 63.4%, *diabetes mellitus 23.8%, *smoking 9.5%, *hyperlipoproteinemia 5.5%, *2 and more risk factors 97.8%. 5. Patients with CVI and AF are more often women and more than 60 years old. According to the results of this research, an incidence of AF in CVI is lager in comparison to the world parameters. AF in persisting form occurs mostly at arteriosclerosis of heart. Women more than 60 years old are the most jeopardized category. Concerning that CVI is in the first place as a cause of invalidity and in the third place as a cause of death, AF as a risk for CVI has important place in the prevention and treatment together with other risk factors. PMID- 11117035 TI - [Snake bites and calcium preparations]. AB - Exact statistical data about snake bites are mostly coming from developed countries. In the United States, for example, 10 to 15 persons die because of a snake bites per annum. In the whole world, according to WHO data from 1983, 10,000 individuals die per annum. In Europe, one case of death occurs in 2 to 3 years. Epidemiological studies, carried out in different countries, showed that a half of the victims came in Emergency Medical Service (EMS) with mild signs of poisoning or without any sign at all. In Mostar, 189 patients were admitted for medical treatment in ten years period (1972-1983) and there was no death case. In the period 1998-2000, increased number of cases snake bites was notified. Two cases were with severe clinical signs and we want to present them in our paper. PMID- 11117036 TI - [Prehospital treatment of pulmonary edema at the Emergency Medical Services Center in Sarajevo]. AB - Quality of prehospital treatment of pulmonary edema depends on functioning of emergency medical service, education and level of competence of medical staff, service equipment, suitable transport and work coordination with Emergency Medical Centre of Clinical centre in Sarajevo. In 1998 in Emergency Medical Service Centre (EMSC) 54 patients with pulmonary edema were treated. In different rates pulmonary edema was associated with myocardial infarction, hypertension, hypotension, rhythm disorders, chronic obstructive pulmonary disease and heroin overdose. Out of total number of treated patients three had lethal outcome what indicates good organisation and efficiency of EMSC Sarajevo. PMID- 11117037 TI - [Use of telematics in emergency medicine]. AB - Health telematics is a composite term for health-related activities, services and systems carried out over a distance by means of information and communications technololgies, for the purposes of global health promotion, disease control and health care, as well as education, management, and research for health. The concept of health telematics encompasses the following functional areas:--tele education;--telemedicine;--telematics for health research;--telematics for health services management. Communications technologies are rapidly revolutionizing health care. For example, electronic communications support diagnosis and treatment of disease. TeleMedicine is an umbrella term for growing disciplines such as TeleRadiology, TelePathology, TeleCardiology, TelePsychiatry and TeleEducation. TeleMedicine is a component of TeleHealth, which includes the use of telecommunications technology and services for the surveillance and control of diseases and education. In this article authors describes the role of telemedicine and telematics in medical education and medical praxis. PMID- 11117038 TI - [Global MED-NET]. AB - This poster will show a concept as well as some practical solutions for the database which is intended to be used by all members and institutions of the Emergency Medical Care international institutions network. This poster shows how data is automatically processed in a rapid and good quality way, while in the same time the lives are saved and time and money are saved too. PMID- 11117039 TI - [Requirements for hospital emergency diagnosis in definitive treatment of injuries]. AB - For the purpose of prompt diagnostics, next to emergency room, we have x-ray room with a possibility of chest and skeleton x-ray filming, as well as one mobile x ray machine and ultrasound device, with a radiologist and radiographer on 24 hours shift. Subspecialists radiologists are on call, if necessary to perform special radiological procedure like angiography, CT, or MRI. We use CT like screening method for all neurological cases in emergency room. Well equipped urgent laboratory is on service 24 hours a day with a specialist biochemist and laboratory technician so all the urgent laboratory findings are available promptly and even can submit a result by phone. Blood bank has also extended department next to emergency room with full service 24 hours a day. Our experience showed that urgent diagnostic, organised in this way, provides faster definite treatment whether patient is going to Intensive Care Unit (ICU) for further reanimation or to operating room for urgent surgical procedure. PMID- 11117040 TI - [Medical treatment and air transport evacuation (initial view and project realization)]. AB - In this paper is presented modality of emergency medical transport by air. Here are a lot of examples of this way of organisation in developed countries as Europe and USA. This way of transport is very effective system of air transport, and it will be very practically in the future in countries in transition as B&H. PMID- 11117041 TI - [The Bjelnasnica-Sarajevo Medical Center--the project concept]. PMID- 11117042 TI - Carpal injuries in children. AB - Pediatric carpal injury is unusual. Because of its rarity and imaging difficulties, the diagnosis is often delayed. Open reduction is advocated for displaced injuries. Pin fixation provides temporary stabilization of displaced injuries without permanently compromising joint motion. In older children, intercarpal fusion may be elected for treatment of intercarpal instability. PMID- 11117043 TI - Articular fractures of the fingers in children. AB - Articular fractures of the fingers in children may result in severe sequelae that impair function of the hand. All displaced fractures must be reduced and fixed. Undisplaced fractures must be securely immobilized (including neighboring joints) and checked frequently for a possible displacement. Even when tested properly, articular fractures may have an unsatisfactory outcome. PMID- 11117044 TI - The cartilaginous cap fracture. AB - The pediatric cartilaginous cap fracture is a transverse fracture through the neck of the proximal or middle phalanx. Nomenclature, mechanism of injury, classification of the fracture, diagnosis, treatment, and complications are discussed. PMID- 11117045 TI - Fingertip replantation in children. AB - Despite common unfavorable mechanisms, fingertip replantation is a rewarding procedure in children. Cosmetic final results are usually better than those obtained by local or pedicled flaps. The success rate and the sensory reinnervation are also better than what can be expected in adult patients. PMID- 11117046 TI - Replantation of large segments in children. AB - If one looks at the final results obtained in children, one should conclude that replantation of large segments is more often indicated in children than in adult patients. Nevertheless, the more common components of crush or avulsion and the frequent severe associated lesions must restrain the surgeon's enthusiasm when indicating replantation of a large segment in children. The possible dramatic consequences of a late revascularization syndrome can be easily foreseen as an outcome of replantation of a large segment in children. Moreover, the problem of growth must be faced from the start, programming secondary surgery either for soft tissue assessment (skin retraction treatments, tendon lengthening, muscle sliding) or for bone lengthening. The final outcome being a functional arm, special care has to be taken in nerve repair integrated with possible secondary tendon transfers to compensate the functional deficit. With all these limitations in indications, care in emergency, and correct timing and planning for secondary surgery, the final functional results of macroreplantations in children will certainly be improved. PMID- 11117047 TI - Coverage of large skin defects of the pediatric upper extremity. AB - Reconstructive surgery of the locomotion apparatus in children has been built from several surgical advances. It is a specific activity that cannot be detached from all other orthopedic pediatric surgery. The care of problems in a child must be provided with a holistic point of view: It cannot be the addition of different techniques done by different surgeons. Bone morphology has a close relationship with the soft tissues; an injury of the envelope has consequences for growth harmony. PMID- 11117048 TI - Two-stage flexor tendon grafting in children. AB - Two-stage flexor tendon grafting is a procedure to salvage severely injured flexor tendon systems. It is a complex procedure that requires a surgeon's experience and the cooperation of the child, parents, and a therapist. The results are best in older children (10 to 15 years old) when using the Paneva Holevitch's technique with a suture in the palm of the hand. A continuing problem is always the future of the graft and the growth potential in children. PMID- 11117049 TI - Peripheral nerve repairs and their results in children. AB - It seems that lesions of nerves in children account for 10% to 15% of the total cases seen in this, as in other, specialist units. The prognosis for recovery after repair is defined by the severity of the original injury. The authors' worst results were seen in complete lesions of the supraclavicular brachial plexus and in the untidy wounds of war, in which penetrating missiles destroyed the proximal femur and the adjacent nerves. The outcome for injuries to the lumbosacral plexus, from fractures of the pelvis or from penetrating missile injury, are as bad. The most significant variable in determining prognosis is delay. Evidently, children are at least as vulnerable to the harmfulness of this as are adults. Skillful primary repair, however, is often followed by recovery to near normal levels, even in complex cases. Adequate follow-up is essential to detect and treat progressive deformity, especially at the ankle and foot. PMID- 11117050 TI - Hand and wrist injuries in young athletes. AB - Successful treatments of musculoskeletal injuries in the pediatric population demand a thorough understanding of the basic anatomy and its biomechanics, and the physiology of growth and development of the immature skeleton. In addition, good treatment outcomes rely on the treating physician being an effective teacher to the young athlete and the patient's parents, coaches, and trainers. At the same time, the physician must be a good student in learning the nature of the patient's sports and each patient's athletic ability and aspirations. Most pediatric hand and wrist injuries can be treated nonoperatively with proper immobilization techniques and activity modification, but cases requiring surgical intervention must be recognized promptly to avoid long-term complications. PMID- 11117051 TI - Vascular cell tumors of the hand in children. AB - The diagnosis of vascular cell tumors of the upper limb in children can be extremely difficult. There is considerable variation in their presentation and natural history. In most instances, the correct diagnosis can be achieved after a careful history and examination. Almost invariably, they are benign lesions and, of these, hemangiomas and vascular malformations account for more than 90%. Worrying tumors of the intermediate and malignant grade are rare. Any atypical, rapidly growing tumor with superficial ulceration and bony destruction on radiology should be regarded as malignant. Histologic differentiation of all problematic vascular swellings requires the services of an experienced pediatric vascular pathologist. In the future, specific cellular marker profiles of individual lesions may simplify the diagnosis. Conservative, symptomatic management is the first line of treatment for almost every vascular swelling. Intervention is reserved for swellings complicated by ulceration, uncontrollable rapid growth, coagulopathies, cardiovascular compromise, or the possibility of malignancy. Small lesions may generally be simply excised, whereas larger lesions often remain a problem. PMID- 11117052 TI - Fibromatoses and related tumors of the hand in children. A clinicopathologic review. AB - Benign and malignant tumors of the hand are rare in children. This article reviews some of the common tumors that affect the hand in children, with an emphasis on clinico-pathologic correlations. Illustrated case histories on some rare tumors are also included. PMID- 11117053 TI - The hand in recessive dystrophic epidermolysis bullosa. AB - Recessive dystrophic epidermolysis bullosa is still a difficult challenge for surgeons and doctors. Hand retractions are different from burns. The medical team must have a good knowledge of the disease. Surgery must be performed with the collaboration of every member of the team, as well as the family. In young children, complete correction of the retractions is possible. On the other hand, when retractions are present for a long time in children and adolescents, surgery is purely functional. Static and dynamic splints delay recurrences, but their prolonged use presents psychological obstacles in adolescents and adults. PMID- 11117054 TI - Volkmann's ischemic contracture. Prevention and treatment. AB - It may be concluded that treatment of patients with Volkmann's ischemic contracture is complicated and depends on a number of different variables. Optimal treatment of an established contracture requires a through examination of the extent of damage of the ischemia, followed by conservative therapy or operation. The most important measures concerning Volkmann's ischemic contracture, however, involve measures to prevent the contracture. It is poignant that very simple measures, such as monitoring high-risk injuries and immediate vascular repair or decompression if symptoms of a compartment syndrome are present, can prevent this disabling condition. The following summaries hopefully provide guidelines for prevention and treatment of Volkmann's ischemic contracture. PMID- 11117055 TI - Rheumatic disorders of the hand and wrist in childhood and adolescence. AB - Rheumatic diseases are common in the pediatric population. Because the hand surgeon is often the first specialist to whom children with rheumatic disease involving the upper extremity are referred, it is important they are aware of the wide variety of disorders that can present with joint complaints to facilitate prompt referral and treatment of these patients. PMID- 11117056 TI - Traumatic arthritis of the hand and wrist in children. AB - Principles, and not established rules, form the basis for treating children with painful, posttraumatic arthritis involving the hand and wrist. The authors' preference is to exhaust nonoperative measures unless the involved joints are unstable or grossly incongruent. The authors would also recommend a greater degree of cautious observation in young children because of reports of successful outcomes associated with remodeling. Once surgery is necessary, the authors prefer motion-preserving procedures, bearing in mind that arthrodesis is well tolerated in the thumb interphalangeal joint and the finger DIP joints. Some authors have presented novel treatments, including complex microvascular reconstructions, but these authors recommend these procedures only in special circumstances, and only by physicians who are experienced with the techniques. PMID- 11117057 TI - Upper limb lengthening. AB - Congenital deficiencies and developmental deformities of the upper extremity often result in complex deformities that include, to variable degrees, shortening and angulation. Because of the nonweight-bearing status of the upper extremity, these deformities are better tolerated and often of less functional significance than their counterparts in the lower extremity. The need for lengthening therefore is less common in the upper extremity than in the lower extremity. When planning a lengthening procedure to the upper limb, the surgeon must be aware of some specific indications, goals, and complications. In the final analysis, one must weigh the risk of upper limb lengthening against the benefits. In fact, there are many pitfalls, and it is not a surgery to be undertaken lightly. In the hands of experienced specialists, it can achieve excellent results. The potential complication rate is high initially, but tends to diminish with increasing experience. The authors believe that the functional, cosmetic, and psychological benefits of upper limb lengthening outweigh the risk of permanent sequelae and functional impairment in selected patients. PMID- 11117058 TI - Treatment of the upper limb in the child with arthrogryposis. AB - There are good options to maximize upper limb function for the child with arthrogryposis. Preservation of available elbow and wrist motion is essential. A comprehensive plan for upper limb treatment includes operative and nonoperative care. PMID- 11117059 TI - Madelung's deformity. Surgical correction through the anterior approach. AB - A resurgence of interest in Madelung's deformity has developed recently because of improved operations for correction of the deformity, identification of the genetic loci for the condition in certain syndromal variants, identification of an anterior ligamentous structure tethering the carpus, and preventive treatments in growing children. The process is reviewed in this article and a new surgical technique is presented. The procedure is performed by way of an anterior incision that is more cosmetically appealing. The release of an anterior ligamentous structure described by Vickers is performed simultaneously with a dome shaped osteotomy of the radius. The fragments, once alignment is corrected, are stablized with temporary pin fixation and a long arm cast until the bone has healed. PMID- 11117060 TI - Management of purpura fulminans at the upper extremity. AB - A multidisciplinary approach often is needed, especially at the acute phase. Indeed, resuscitative treatment is first indicated. [figure: see text] Optimal time for surgery depends on the patient's status. Surgical management should be more aggressive than previously thought. Debridement of necrotic tissue must be performed when the eschar has demarcated to avoid infection. Escharectomy can improve the vitality of the distal segments by releasing the compressive mechanical effect. In the same vein, early amputations are likely to reduce the need for secondary revisional surgery. Necrotic tissue over a joint should be excised early and procedures to provide coverage by flaps should be undertaken to avoid osteoarthritis. Late sequelae are dominated by skeletal growth disturbances and require specific procedures. PMID- 11117061 TI - Play doctor online. Take two aspirin and click back in the morning. PMID- 11117062 TI - A virtual checkup. Advice on health, fitness, and parenting can make the Web a lifeline. PMID- 11117064 TI - Leaders converge on issues. PMID- 11117065 TI - Can long-term care survive the staffing|| crisis? PMID- 11117070 TI - Who cares? PMID- 11117071 TI - Life after practice management: Doctors return to more traditional business styles. PMID- 11117072 TI - HCFA/PRO study a benchmark, not a crisis report: TMA, THA & MSFMC come together to "make good care better". PMID- 11117073 TI - Hazard of a new patient. PMID- 11117074 TI - Treatment of elderly women with urge incontinence in middle tennessee: a single institution practice-based study. AB - INTRODUCTION AND OBJECTIVES: Urinary urge incontinence (UUI) is a major factor in reducing quality of life in elderly women. The treatment of UUI in the elderly population is complicated by comorbidities, polypharmacy, cost, and side effects. The purpose of this study was to examine our practice pattern in Middle Tennessee for the treatment of elderly women with UUI. METHODS: We retrospectively reviewed the medical records of all women over age 65 seen at our institution between January 1, 1998 and September 1, 1999 with an initial complaint of pure UUI. Diagnosis was based on history and physical examination by a single urologist (JJF). Initial treatment in all patients was medication as well as timed and double voids. Medication chosen was based on cost factors, co-morbidities, current medications, and outcome from previous treatment. RESULTS: Of 53 women ranging in age from 65-87 years of age (avg. 74.7) included in this study, 6/53 (11.3%) had a previous CVA, and 2/53 (3.7%) had grade I-II cystoceles. Initial pharmacologic treatment included anticholinergic medication in 47 patients (88.6%), and either imipramine or topical estrogen alone in the remaining 11.4%. Of the anticholinergics, hyoscyamine time capsules were used in 29, tolterodine in 7, standard oxybutynin in 5, oxybutynin XL in 1, and a combination with imipramine in 5. Thirty-four of the 53 total patients (64.1%) discontinued their medications because of no improvement 14 (41.1%), dry mouth 9 (26.4%), other side effects 9 (26.4%), cost 1, and other reasons in the remaining 2 patients. Only 17 patients (32%) stated they were doing well on their initial medications; 11 of those (64.7%) were taking hyoscyamine time capsules. Upon subjective failure, 22/36 patients (61.1%) had their medications changed, while 14/36 (38.8%) pursued behavioral therapy without additional medications. Urodynamic studies were done in 12 patients who failed empiric medical treatment (22.6%). CONCLUSIONS: Only 32% of elderly women treated medically for UUI were satisfied and continued therapy in this patient population. One-fourth of elderly women failed empiric medical management of UUI due to lack of efficacy, and one-third due to intolerable side effects. In this practice, hyoscyamine was continued more often than any other anticholinergic because of reasonable cost, efficacy, and side effect profile. PMID- 11117075 TI - Certification of death. PMID- 11117076 TI - Severe hypertension in a young woman. PMID- 11117077 TI - Echo-enhanced ultrasound--clinical and technical aspects. PMID- 11117078 TI - The Dandy-Walker complex and fetal sonography. PMID- 11117079 TI - Characterization and natural history of ventricular septal defects in the fetus. AB - OBJECTIVE: To characterize and describe the evolution of ventricular septal defects (VSD) from intra-uterine diagnosis to infancy in a population of fetuses with isolated defects. METHODS: Sixty-eight fetuses with isolated VSD represented the study population. Of these, 28 underwent termination of pregnancy, 14 died in utero or after birth and 26 reached 1 year of age. In this population, the following variables were evaluated: presence of extracardiac or chromosomal anomalies, site and size of the defect, pregnancy outcome. These variables were assessed against closure of the VSD up to 1 year of age. Necropsies were available for all fetuses following termination of pregnancy. All surviving neonates were followed up directly or by telephone until documented echocardiographic closure of the defect or until 1 year of age. RESULTS: There was a significant correlation between type of VSD and type of aneuploidy (P < 0.001). A total of 26 surviving fetuses reached 1 year of age: 46.1% (n = 12) of all defects closed in utero, 23.1% (n = 6) closed during the first year of life and 30.8% (n = 8) remained patent. Only three (15.8%) of the 19 VSDs < 3 mm remained patent in comparison with five (71.4%) of the seven defects > 3 mm (P < 0.05). None of the malalignment VSDs closed, in comparison to 69% of the peri membranous and 60% of the muscular defects. CONCLUSION: Ventricular septal defect can undergo spontaneous closure during intra-uterine life and this process depends upon the site and the size of the defect. These data may provide useful additional information to aid prenatal counseling. PMID- 11117080 TI - A preliminary study of sonographic grading of fetal intracardiac echogenic foci: feasibility, reliability and association with aneuploidy. AB - OBJECTIVES: To prospectively and quantitatively grade intracardiac echogenic focus/foci (ICEF) using sonographic gain reduction and to determine the association of ICEF by grade with fetal aneuploidy. METHODS: Women referred for raised maternal age (> or = 35 years), or > 18 years of age and with a Down syndrome risk > or = 1/270, increased trisomy 18 risk by second trimester serum screen or a prior aneuploid offspring were included in this institutionally approved protocol. Only pregnancies of gestational age between 14 and 24 weeks were included. All women had a targeted ultrasound and were offered fetal chromosome analysis. The classification of ICEF was made from a four-chamber view of the fetal heart. The echo amplitude of the ICEF was compared to that of the thoracic spine and categorized according to the comparative gain setting at which the image of the relevant structure disappeared: Grade O = no ICEF present, Grade 1 = ICEF image lost before thoracic spine when gain was reduced, Grade 2 = ICEF image lost at same gain setting as thoracic spine, Grade 3 = thoracic spine image lost before ICEF. The primary outcome was a prenatally or post-natally detected chromosomal abnormality. RESULTS: A total of 885 eligible women were examined during the 21-month study period. ICEF were seen in 29 (3.3%) fetuses: 24(83%) in the left ventricle and five (17%) in the right ventricle. A chromosome abnormality was identified in 13/671 (1.9%) fetuses without ICEF (Grade 0) and 0/21 (0%) fetuses with Grade 1 ICEF. In contrast, two of five (40%) fetuses with Grade 2 ICEF were aneuploid (P = 0.005). No Grade 3 ICEF were observed. Additional sonographic abnormalities were seen in both aneuploid fetuses with Grade 2 ICEF. Interobserver agreement on ICEF grading was noted in 50/50 (100%) examinations (kappa = 1.0). CONCLUSIONS: Sonographic grading of ICEF is feasible and highly reliable. Grade 2 ICEF, especially when accompanied by additional sonographic markers of a chromosomal abnormality, are associated with aneuploidy significantly more frequently than Grade 1 ICEF. PMID- 11117081 TI - Echocardiographic assessment of ventricular filling pressure during the second and third trimesters of gestation. AB - AIMS: To confirm changes in the atrioventricular diastolic flow velocities (peak E, peak A, E/A ratio) with gestational age and to define whether these changes genuinely reflect variations in filling pressure in the fetal heart. METHODS: Fifty normal pregnancies were studied between the 13th and the 37th weeks of gestation. The fetal flow velocity patterns were evaluated by pulsed-wave (PW) Doppler and the annular velocities of the atrioventricular valves by PW-Doppler tissue imaging (DTI). RESULTS: All indexes evaluated (atrioventricular peak flow and annular velocities) correlated significantly with gestational age. This correlation was stronger for the early diastolic indexes (tricuspid E and EA, 0.69 and 0.78; mitral E and EA, 0.61 and 0.77, respectively) and weaker for the end-diastolic indexes (tricuspid A and AA, 0.46 and 0.37; mitral A and AA, 0.45 and 0.39, respectively). Neither mitral nor tricuspid E/Ea ratio changed significantly with gestational age. CONCLUSIONS: The lack of correlation between the Doppler-assessed ventricular filling pressures and gestational age suggests absence of significant changes of ventricular compliance during the second and third trimesters of pregnancy, and a progressive enhancement of active relaxation and/or changes in loading conditions. PMID- 11117082 TI - Trisomy 21: 91% detection rate using second-trimester ultrasound markers. AB - OBJECTIVES: To examine cardiovascular and non-cardiovascular prenatal ultrasound markers and determine which markers physicians of varying skill levels could use to identify fetuses with trisomy 21. METHODS: Eighty second-trimester fetuses with trisomy 21 and 2000 controls underwent real-time plus color Doppler examination of cardiovascular and non-cardiovascular systems followed by amniocentesis. Non-cardiac markers were central nervous system malformations (CNS); choroid plexus cysts (CPC); abnormal nuchal skin fold (NSF); hyperechoic bowel (HB); and pyelectasis (PY). Cardiac markers consisted of ventricular septal defect, right-to-left chamber disproportion (RL); tricuspid regurgitation; mitral regurgitation (MR); pericardial effusion; and outflow tract abnormalities (OFT). Multinomial logistic regression was used to identify interactivity between the markers. Logistic regression was utilized to identify which combinations of markers significantly contributed to the identification of fetuses with trisomy 21 and to compute the likelihood ratio. RESULTS: All but three markers (CPC, MR, OFT) contributed significantly to the identification of 91% of fetuses with trisomy 21 with a false-positive rate of 14%. When only non-cardiovascular markers were examined, all but CPC contributed to the identification of 60% of fetuses with trisomy 21 with a false-positive rate of 5.9%. Combining right-to left chamber disproportion with CNS, NSF, HB and PY identified 75% of fetuses with trisomy 21 with a false-positive rate of 6.4%. All markers were independent predictors of trisomy 21 except RL and NSF. CONCLUSION: Ultrasound can detect between 60 and 91% of fetuses with trisomy 21 depending upon which markers are selected for evaluation. PMID- 11117083 TI - Femur length and trisomy 21: impact of gestational age on screening efficiency. AB - OBJECTIVE: This study assesses two methods used to define relatively short femur in screening for trisomy 21 and examines changes in performance of screening with gestational age. DESIGN: Retrospective analysis of data on menstrual age, femur length (FL) and biparietal diameter (BPD) in 49 trisomy 21 pregnancies and 6069 normal controls. Reference ranges were derived for BPD/FL versus menstrual age and for FL versus BPD. Two methods of defining short femur (BPD/FL and observed to-expected FL ratio) were examined for false-positive rates and detection rates for trisomy 21 at different gestational ages. RESULTS: In the control group the BPD/FL ratio and its standard deviation decreased with menstrual age. Trisomy 21 was associated with a significantly higher BPD/FL ratio (P < 0.001) and the deviation increased significantly with menstrual age (P < 0.05). Eleven percent of 28 fetuses examined at 15-17 weeks had a BPD/FL above the 95th centile compared with 24% of 21 fetuses examined at 18-20 weeks (P = 0.40). The median observed-to-expected FL ratio in the control group was 1.0 throughout the gestational age range but the standard deviation decreased significantly with menstrual age (P < 0.01). Trisomy 21 was associated with a significantly reduced observed-to-expected FL ratio (P < 0.001) and the deviation increased significantly with menstrual age (P < 0.05). A fixed cut-off of 0.91 for observed to-expected FL ratio provided a false-positive rate of 12% at 15-17 weeks compared with 6% at 18-20 weeks of gestation (P < 0.001) with detection rates of 29 and 38%, respectively (P = 0.73). CONCLUSION: Irrespective of the definition used to define the condition, relatively short femur is a poor marker for trisomy 21 particularly when the assessment takes place before 18 weeks of gestation. PMID- 11117084 TI - Nuchal thickness evolution in trisomy 18 fetuses. AB - OBJECTIVE: To assess the natural evolution of nuchal thickness in trisomy 18 fetuses. METHODS: Serial measurements of nuchal thickness were performed over a 1 to 5-week period in 17 fetuses with trisomy 18, from the 10th to the 20th week of pregnancy. To avoid a confounding effect due to gestational age, nuchal thickness was also expressed in standard deviations (SD) for the corresponding gestational week. In addition, the changes were assessed in terms of the presence of clinically positive thickening, considered as such when its measurement was above 2.5 SD for the corresponding gestational week. RESULTS: On the initial nuchal thickness measurement, sensitivity for trisomy 18 decreased from 66% on assessment at 10-13 weeks of gestation to 38% at 14-16 weeks. In serial determinations, a mean increase of 1.3 mm [95% confidence interval (CI), 0.1-2.5] was observed for a mean period of 21 days. When corrected for gestational age, the mean increase of 0.2 SD (95% CI, -1.2 to -1.6) was found to be non significant. No clinically relevant changes were recorded on re-examination, with nuchal thickening remaining stable in 76% of cases. CONCLUSION: Nuchal thickening at re-examination was observed in a similar proportion of trisomy 18 fetuses as when initially observed. PMID- 11117085 TI - Correlation of prenatal ultrasound diagnosis and pathologic findings in fetal brain abnormalities. AB - OBJECTIVE: To determine the degree of agreement between prenatal ultrasound diagnosis of brain abnormalities and subsequent pathologic findings. METHODS: Between August 1993 and August 1999 there were 62 cases where a fetus with a prenatal ultrasound diagnosis of a brain abnormality other than neural tube defects underwent autopsy at the Regional Department of Pediatric Pathology. The cerebral diagnosis at ultrasound was compared with the findings at autopsy in all cases. RESULTS: In 47 of 61 (77%) cases the same defects were seen on ultrasound and at autopsy. The most common disparity was with the Dandy-Walker malformation or variant, where only six of the 14 (43%) cases prenatally diagnosed with this condition showed the same abnormality at autopsy. When fetuses with the Dandy Walker malformation or variant were excluded, the scan findings correlated with autopsy in 41 of 47 (87%). In the main group with discordant findings, five of the seven cases where termination of pregnancy was undertaken had other fetal anomalies on ultrasound examination which were confirmed at autopsy. In the sixth case there was autolysis of brain tissue which affected detailed autopsy. CONCLUSIONS: A very high level of agreement between prenatal ultrasound and autopsy findings was found for all abnormalities of the fetal brain, except for the Dandy-Walker malformation or variant. Potential discrepancy in findings between ultrasound and autopsy should be explained to patients who are considering termination of pregnancy for the Dandy-Walker type of abnormality. PMID- 11117086 TI - Serial fetal lung volume measurement using three-dimensional ultrasound. AB - OBJECTIVE: To establish reference intervals for fetal lung growth. DESIGN: Longitudinal observational study. SUBJECTS: Fifty-eight women with initially uncomplicated singleton pregnancies were recruited from the antenatal population of a teaching hospital. Four women were excluded from the final analysis because of complications arising in their pregnancy. METHODS: Each subject was serially scanned at monthly intervals. At each visit lung volume was measured using an ultrasound-based computerized three-dimensional imaging system. Multilevel models were used to determine conditional and unconditional reference intervals. RESULTS: Reference intervals for fetal lung growth were derived. Fetal lung volume increases in a non-linear way with gestation. CONCLUSIONS: Our computerized system has the capacity to be used in conjunction with any standard two-dimensional ultrasound scanner in order to measure volume. Lung volume measurement may be useful in predicting pulmonary hypoplasia. PMID- 11117087 TI - Prenatal ultrasound diagnosis of congenital cystic adenomatoid malformation of the lung: a report of 26 cases and review of the literature. AB - OBJECTIVES: To evaluate the sonographic appearances and prenatal natural history of congenital cystic adenomatoid malformation of the lung. METHODS: In each case a detailed examination of the thoracic lesion and a complete fetal survey was performed. The pregnancies that elected to continue were followed to term. RESULTS: A total of 26 cases were identified. The pregnancy was electively terminated in nine cases (35%). All the remaining 17 pregnancies ended in liveborn infants (100%). The lesion disappeared completely in three fetuses (18%). Of the 14 infants in whom the lesion was confirmed at birth nine required surgery in the neonatal or post-natal period. Five children did not undergo surgery. CONCLUSIONS: Conservative management appears to be an adequate medical practice in cases of isolated congenital unilateral cystic adenomatoid malformation of the lung, in the absence of hydrops and/or acute polyhydramnios. PMID- 11117088 TI - Comprehensive analysis of uterine artery flow velocity waveforms for the prediction of pre-eclampsia. AB - OBJECTIVES: To evaluate the performance of velocimetric indices of uterine artery flow velocity waveforms (FVW's) at 20 weeks' gestation, alone or in combination with qualitative analysis, and establish the optimal screening method for the prediction of pre-eclampsia. METHODS: A total of 614 primiparous women had color flow/pulsed Doppler (CFPD) imaging of both uterine arteries at 20 weeks gestation. Receiver operator characteristic (ROC) curves were created for the systolic/end-diastolic (A/B) ratio, resistance index (RI) and systolic/early diastolic (A/C) ratio for placental and non-placental uterine arteries, individually or in combination with the presence of unilateral or bilateral notches. Based on data from ROC curves, the sensitivity of each method was compared with the false-positive rate set at 17 and 11%. RESULTS: The highest sensitivity (88%) and specificity of (83%) was obtained using bilateral notches/mean RI > or = 0.55 (50th centile) and unilateral notches/mean RI > or = 0.65 (80th centile). When the false-positive rate was set at 17%, the inclusion of bilateral notches significantly improved the sensitivity of RI (P < 0.001), placental RI (P < 0.01), placental A/C ratio (P < 0.05), mean A/C ratio (P < 0.01) and mean A/B ratio (P < 0.05). Bilateral notches/mean RI or A/B cut-offs were also superior to the persistence of a notch in either artery combined with RI (P < 0.01) or A/B ratio (P < 0.05). When the false-positive rate was set at 11%, the inclusion of bilateral notches did not improve the sensitivity of the A/C (P = 1.00) or A/B ratio (P > 0.10). Placental velocimetric indices performed better than mean indices but the differences in sensitivity at the set false positive rates were not statistically significant. CONCLUSION: At 20 weeks' gestation, bilateral notches with mean RI cut-offs is the best screening method if further screening later in pregnancy is proposed. The A/C ratio is complementary to bilateral notches when the false-positive rate is set at 17% and an effective quantitative substitute when the false-positive rate is set at 11%. PMID- 11117089 TI - Predicting the risk of pre-eclampsia and a small-for-gestational-age infant by quantitative assessment of the diastolic notch in uterine artery flow velocity waveforms in unselected women. AB - OBJECTIVES: To develop a new quantitative index, the notch depth index (NDI), to evaluate its association with the risk of pre-eclampsia and a small-for gestational-age (SGA) infant and to compare its clinical usefulness with that of the uterine artery resistance index (RI) and the peak systolic to early diastolic velocity (A/C) ratio. METHODS: Uterine artery color Doppler ultrasound was performed in 288 consecutive healthy pregnant women at 20.2 +/- 2.0 (range 16.0 23.9) weeks of gestation. The NDI represents the depth of the early diastolic notch divided by the maximal diastolic velocity. RESULTS: Nine (3.1%) of the 288 women developed pre-eclampsia and 18 women (6.3%) delivered an SGA infant. The NDI was associated with subsequent onset of pre-eclampsia. The optimal cutoff value for the NDI in predicting pre-eclampsia was 0.14, giving a sensitivity, specificity and a positive predictive value (PPV) of 67, 92, and 22%, respectively. The PPV of the NDI was the largest of the three indices evaluated (12% for the RI and 16% for the A/C ratio). The relative risk for pre-eclampsia in women with values equal to or greater than the optimal cutoff values of the RI, A/C ratio and the NDI was 9.7 (95% confidence interval, 2.5-3.7), 19.2 (4.2 91), and 19.2 (5.1-71), respectively. The NDI of 0.14 improved the PPV of 18% determined by the presence of notches in bilateral uterine arteries. The optimal cutoff value of 0.14 for the NDI in predicting an SGA infant yielded a higher PPV (22%) than those for the RI (9%) and A/C ratio (12%). CONCLUSIONS: The NDI value in the second trimester is associated with the later onset of pre-eclampsia, and is clinically more useful in predicting pre-eclampsia than the two conventional indices. PMID- 11117090 TI - Maternal cerebral hemodynamics in pregnancy-related hypertension. A prospective transcranial Doppler study. AB - AIM: To compare maternal cerebral hemodynamics, as assessed by transcranial Doppler studies, with the clinical and radiological findings in different types of pregnancy-related hypertension and to determine their pathophysiology. METHODS: A prospective study of 66 consecutive pregnant women with hypertensive disorders (eclampsia, n = 3; pre-eclampsia, n = 41; isolated hemolysis, elevated liver enzymes, and low platelet count (HELLP)-syndrome, n = 12; pre-eclampsia superimposed on chronic hypertension, n = 5; chronic hypertension, n = 5) and 21 women with uncomplicated pregnancies. Mean blood flow velocities (Vmean) were assessed serially by means of transcranial Doppler in all basal arteries and correlated with changes in mean arterial blood pressure (MABP) and the clinical course. RESULTS: Patients with the pre-eclampsia/eclampsia syndrome showed significantly elevated Vmean values as compared to controls. In the course of the illness Vmean over the whole length of all insonated basal arteries rose simultaneously. The three eclamptic patients showed the highest Vmean values (156, 182, 192 cm/s, respectively), of the middle cerebral artery (MCA) while MABP was 135, 135, and 150 mmHg, respectively. In pre-eclamptic patients the maximal Vmean MCA ranged from 80 (67, 93) to 145 (114, 151) cm/s [median (25th, 75th percentile)] depending on the severity of clinical presentation. In patients with isolated HELLP-syndrome changes in Vmean were either mild (5/12 cases) or absent (7/12 cases). CONCLUSIONS: Considerable differences in cerebral hemodynamics were observed in the various types of pregnancy-related hypertensive disorders examined in this study. Our findings in patients with pre eclampsia/eclampsia syndrome suggest a breakdown of autoregulation with hyperperfusion and vasogenic edema being the most probable pathophysiological mechanism. PMID- 11117091 TI - Sonographic appearance of first trimester complete hydatidiform moles. AB - OBJECTIVE: Complete hydatidiform moles are now being diagnosed earlier in gestation, thus the clinical presentation and pathologic findings of complete molar pregnancy have changed. We studied the sonographic appearance of first trimester moles and the ability of ultrasound to detect them. METHODS: We reviewed the sonographic interpretation and sonograms, when available, from all patients with first trimester complete moles diagnosed at our institution from January 1988 to March 1996. RESULTS: Of the 24 patients in our study, the mean gestational age at time of the sonogram was 8.7 +/- 2.0 weeks (mean +/- SD) with a range of 5.7-12.3 weeks. The initial sonographic interpretation was a complete mole in 17 (71%) cases, partial mole versus failed pregnancy in two (8%), and failed pregnancy in five (21%) cases. Of the 22 patients with sonograms available for review, interpretation on review of the images was a complete mole in 18 (82%) cases, partial mole versus failed pregnancy in one (5%), and failed pregnancy in three (14%) cases. The typical sonographic appearance of a first trimester complete mole was a complex, echogenic, intra-uterine mass containing many small cystic spaces. CONCLUSION: The majority of first trimester complete moles demonstrate a typical ultrasound appearance such that the diagnosis can be made with ultrasound in most cases. PMID- 11117092 TI - Doppler evaluation of the uterine and spiral arteries from different sampling sites and phases of the menstrual cycle during controlled ovarian hyperstimulation. AB - OBJECTIVE: To compare the pulsatility index (PI) and resistance index (RI) at different sampling sites of the uterine and spiral arteries in the early and mid menstrual phases. MATERIALS AND METHODS: The uterine and spiral arteries of 110 women undergoing similar ovarian hyperstimulation and intra-uterine insemination regimes were examined using transvaginal color Doppler ultrasound. The uterine artery was sampled at five sites: (1) ascending branch; (2) descending branch; (3) proximal, near branch division; (4) mid, 0.5 cm distal to the division; (5) lateral location, 1 cm distal to the division. The spiral artery was sampled at three sites: (a) anterior; (b) fundal; (c) posterior. The uterine and spiral arteries were examined twice, on days 2-3 and 14-16, respectively, of the menstrual cycle. The women were also grouped according to age at examination, < or = 30 years and > 30 years. The PI and RI values for different sites, menstrual phase, and age were compared. RESULTS: The mean PI and RI values of the uterine artery were: (1) 2.86 +/- 1.20 and 0.92 +/- 0.13; (2) 2.66 +/- 1.15 and 0.89 +/- 0.12; (3) 2.88 +/- 1.26 and 0.90 +/- 0.15; (4) 3.03 +/- 1.02 and 0.91 +/- 0.07; (5) 3.23 +/- 1.38 and 0.89 +/- 0.12; and of the spiral artery were (a) 1.61 +/- 1.01 and 0.69 +/- 0.17; (b) 1.69 +/- 0.74 and 0.74 +/- 0.17; (c) 1.73 +/- 0.86 and 0.68 +/- 0.17. The PI values for uterine and spiral arteries at two phases of the menstrual cycle were 2.92 +/- 1.18 and 1.55 +/- 0.72 (days 2-3); 3.11 +/- 1.15 and 1.80 +/- 1.02 (days 14-16), respectively; for younger women (age < or = 30 years) these values were 2.83 +/- 1.22 and 1.6 +/- 0.85 and for older women (age > 30 years) 3.0 +/- 1.34 and 1.72 +/- 0.96, respectively. CONCLUSIONS: There were no significant differences in PI and RI values of the uterine and spiral arteries at different sampling sites, phase of the menstrual cycle or age. The higher PI values tended to occur in the lateral uterine artery and posterior spiral artery, during the mid-menstrual phase and in the older age group. The PI and RI values of the mid-uterine and fundal spiral artery sampling sites are representative of the whole uterine artery and spiral artery, respectively. PMID- 11117093 TI - Transient hyperechogenicity of the fetal pericardium: a case report. AB - An unusually echogenic fetal pericardium was visualized using ultrasound at 21 weeks' gestation. Serial prenatal examinations revealed its progressive, spontaneous resolution. Follow-up examinations of the newborn and infant failed to demonstrate any clinical or echocardiographic abnormality. The occurrence of this unusual pericardial abnormality and its transient nature should be considered during sonographic examination of the fetus. PMID- 11117094 TI - A malformed fetus in a rudimentary uterine horn pregnancy. AB - We present a case of a 13-week pregnancy with a malformed fetus in a ruptured, non-communicating rudimentary horn. The patient, a 21-year-old woman with pelvic and right-sided abdominal pain, was admitted to the gynecology clinic of our institution. A ruptured rudimentary horn pregnancy was diagnosed by ultrasonography. The fetus in the gestation sac showed evisceration of the liver and intestines and an absent left femur. There was an amniotic band extending across the body of the fetus. The ruptured horn was excised by laparotomy. The factors associated with rudimentary horn pregnancy and related fetal abnormalities are discussed. PMID- 11117095 TI - Re: Fetal aortic arch measurements between 14 and 38 weeks' gestation: in-utero ultrasonographic study. Ultrasound Obstet Gynecol 2000; 15:226-30. PMID- 11117096 TI - Picture of the month. Clinomicrodactyly. PMID- 11117097 TI - Shame, self-consciousness, and locus of control in people who stutter. AB - Stuttering is a multidimensional disorder, including psychological as well as physiological elements. This investigation of the value of 3 psychological constructs (shame, self-consciousness, and locus of control) in the prediction of 3 self-reported behavioral dimensions of stuttering (struggle, avoidance, and expectancy) revealed shame and self-consciousness to be significant psychological predictors of the selected dimensions of stuttering, whereas locus of control was found not to be. Certain demographic elements, including affiliations with others who stutter, were also determined to be predictive of the stuttering dimensions. The present findings and their implications for theory, research, and practice are discussed. PMID- 11117098 TI - Individual differences in race schematicity as predictors of African American and white children's race-relevant memories and peer preferences. AB - Race schematicity, memories for racially stereotyped portrayals, and race-based peer preferences of 70 young children (32 African American, 38 White) were assessed. Consistent with schema-based models of stereotyping, race schematicity was positively and significantly associated with memory distortions of racial stereotype-inconsistent drawings into stereotype-consistent ones. Conversely, race schematicity was negatively and significantly associated with accurate memories for racial stereotype-inconsistent content, and with memory distortions of racial stereotype-consistent portrayals into stereotype-inconsistent ones. As predicted, race schematicity was positively and significantly associated with same-race peer preference bias, as was children's age in months. Results support application of the schematicity construct and relevant social psychological research with adults to the study of young African American and White children's racial stereotyping and processing of race-relevant information. PMID- 11117099 TI - Asymmetries in infants' attention to the presence or absence of features. AB - Three experiments were performed to test whether infants show a bias for detecting the presence of a feature in a stimulus rather than its absence. In the 1st experiment, 24 16-week-old infants were given 3 paired-comparison problems, each of which included a 25-s familiarization phase followed by 2 test trials. Infants were familiarized to 1 member of a set of capital alphabetical letters (E F; Q-O; B-R). Then they were given a paired-comparison recognition test under 1 of 2 conditions. In the feature-present condition, the familiar letter (e.g., F) was paired with a novel letter containing the addition of a distinguishing element (e.g., E). In the feature-absent condition, infants were presented with a familiar letter (e.g., E) paired with a novel letter in which 1 element was removed (e.g., F). Infants showed a novelty preference to the letter in which the distinguishing feature was present, but there was no preference for novelty in the feature-absent condition. The 2nd experiment showed that infants' fixation to the letter containing the presence of the feature was not due to a simple preference for the letter with the greater number of elements. Finally, to test whether infants' failure to discriminate the absence was due to insufficient encoding time, 36 infants were tested in a 3rd experiment in which familiarization time was varied. After 20 s of familiarization, no evidence of discrimination was observed in either the feature-present or feature-absent condition. After 30 s, however, infants could discriminate the novel letter in the feature-present condition but not in the feature-absent condition. The significance of these results is discussed in terms of theoretical explanations for the development of the feature-presence bias. PMID- 11117100 TI - Differential risk perceptions for unintended pregnancy, STDs, and HIV/AIDS among urban adolescents: some preliminary findings. AB - Two pilot studies tested the hypothesis that adolescents perceive differential risks for unintended pregnancy (UP), sexually transmitted diseases (STDs), and human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS). The 1st study used a college sample consisting of 14 adolescents (21 years or younger) and 64 adults (over 21) who rated the likelihood that they and others would experience 15 health problems. The 2nd study used a community sample of 48 adolescents between 16 and 21 years of age who rated 11 health problems in a similar manner. Optimistic bias and uniqueness of risk in adolescents' perceived susceptibility to adverse sexual outcomes were examined. Optimistic bias is the difference between ratings of risk to self and risk to others, reflecting lower risk to self. Uniqueness of risk is the difference between ratings of risk and a baseline risk estimate (i.e., the mean rating for all non-sex-related health problems). Consistent with the hypothesis, adolescents perceived differential risks for UP, STDs, and HIV/AIDS. Implications for adolescent prevention programs are discussed. PMID- 11117101 TI - When knowing becomes remembering: individual differences in susceptibility to suggestion. AB - In two experiments, the authors explored factors that might influence a person's tendency to make source-monitoring errors about autobiographical memories. In the first experiment, undergraduates retrieved a memory from childhood (a) that was known about but not remembered, (b) that was remembered, or (c) for which they were unsure of their memory's source. After writing down the memory, experimental groups listened to a guided visualization tape and answered questions about the event--interventions designed to help them focus on details of their memory. Controls also retrieved and wrote down a memory; however, instead of visualizing the memory, they were instructed to conduct a visual search task. Results indicated that guided visualization led participants to rate known memories closer to remembered events. A second experiment examined individual difference variables that might be related to this know-to-remember shift. Results indicated that extraversion, external locus of control, a memory that conveyed fear, and overall affective content predicted this rating. The applicability of these findings to the psychotherapy process is discussed. PMID- 11117102 TI - Gender comparisons in the perception of self-competence among four-year-old children. AB - The purpose of this study was to determine the presence of gender differences in the perception of self-competence among 4-year-old children. Sixty-one-4-year olds (27 girls and 34 boys) from predominantly European American backgrounds participated in the study. The children's self-competence was measured using the Pictorial Scale of Perceived Competence and Social Acceptance for Young Children (Preschool version; S. Harter & R. Pike, 1984), which has 4 separate subscales: (a) cognitive competence, (b) physical competence, (c) peer acceptance, and (d) maternal acceptance. A within-subject 2-way analysis of variance with repeated measures of 4 (subscales of perception of self-competence) x 2 (gender) was performed to determine if gender differences existed in the children's perception of self-competence. Analyses of the data showed no significant gender differences in the scores on the 4 subscales for the perception of self-competence among the children. These results could be interpreted as being due to a less gender stereotyped society and androgynous environment for these preschoolers. PMID- 11117103 TI - Prevalence of anxiety and depression in Australian adolescents: comparisons with worldwide data. AB - Data from two Australian studies were combined so that the prevalence of anxiety and depression in a large, normative sample of Australian adolescents could be investigated. The combined sample comprised 1,299 adolescents randomly selected from metropolitan and country schools in Melbourne, a large Australian city. The data were examined in order to ascertain the percentages of adolescents who scored above the clinical cut-off on two self-report instruments--the Revised Children's Manifest Anxiety Scale (C. R. Reynolds & B. O. Richmond, 1985) and the Reynolds Adolescent Depression Scale (W. M. Reynolds, 1986). The results of these analyses were then compared with previously reported prevalence rates from studies worldwide. This comparison revealed striking differences in the prevalence of anxiety and depression across different countries and cultures. Limitations attributable to different self-report measures and imposed-etic approaches are discussed. Issues pertaining to social and political climate are also raised. PMID- 11117105 TI - [Epidemiology of aging]. PMID- 11117104 TI - The Adolescent Unresolved Attachment Questionnaire: the assessment of perceptions of parental abdication of caregiving behavior. AB - This article reports on the Adolescent Unresolved Attachment Questionnaire (AUAQ), a brief questionnaire that assesses the caregiving experiences of unresolved adolescents (as recipients of caregiving). The AUAQ was developed and validated in a large normative sample (n = 691) and a sample of 133 adolescents in psychiatric treatment. It is a self-report questionnaire consisting of 3 scales with Likert-type responses ranging from strongly disagree to strongly agree. The Aloneness/Failed Protection Scale assesses the adolescent's perception of the care provided by the attachment figure. The Fear Scale taps the fear generated by the adolescent's appraisal of failed attachment figure care. The Anger/Dysregulation Scale assesses negative affective responses to the perceived lack of care from the attachment figure. All scales demonstrated satisfactory internal reliability and agreement between scores for adolescents (n = 91) from the normative sample who completed the AUAQ twice. Adolescents in the clinical sample also completed the Adult Attachment Interview (AAI; C. George, N. Kaplan, & M. Main, 1984/1985/1996); the AUAQ demonstrated high convergent validity with the AAI. PMID- 11117106 TI - [Renal osteodystrophy (2): its treatment in renal insufficiency before dialysis]. AB - 1. In the patient with renal insufficiency before dialysis, the phosphocalcic disorders appear insidiously. They are dominated by hyperparathyroidism which will be diagnosed on the initially yearly determination of plasma intact PTH as soon as creatinine clearance decreases below 60 ml/min, eventhough there is still no modification in plasma concentrations of calcium and phosphate. Its diagnosis should lead to initiate the therapeutic measures in order to prevent the irreversible thining of the corticals by endosteal resorption and later the occurrence of histological and radiological osteitis fibrosa favoring fractures. 2. Hyperparathyroidism prevention relies on two main measures: prevention of phosphate retention and hypocalcemia is implemented by progressive phosphate and protein restriction (from 1 g/kg/day when Ccr < 60 ml/min to 0.6 g/kg/day when Ccr < 20 ml/min) and administration of CaCO3 (1.5 g at lunch and dinner to better complex the phosphate) as soon as PTH is above normal; optimal vitamin D repeletion will be implemented by systematic supplementation of native vitamin D or 25OH vitamin D3 in order to bring P25OHD between 30-60 ng/ml (75-150 nmol/l) or more generally around the upper limit of the epidemiologic range of the laboratory; these measures should aim at maintaining plasma intact PTH in its optimal range variable with the degree of renal insufficiency: 0.5-1; 1-2.5 and 2 3 folds the upper limit of normal for creatinine clearance respectively at 60-30; 30-10 and < 10 ml/min. 3. Because of their hyperphosphatemic and hypercalcemic effect, 1 alpha-hydroxylated vitamin D derivatives will be regularly efficient and safe only when non-calcemic non-aluminic phosphate binder will be available and proven to be without side-effects. 4. Instrumental (surgical or by alcohol injection) parathyroidectomy should be considered when plasma intact PTH is > 5 to 7 times the upper limit of normal in the presence of hypercalcemia (> 2.60 mmol/l) and/or hyperphosphatemia (> 1.70 nmol/l) in spite of the above measures, the decision being reinforced by coexistence of bone radiologic abnormalities and metastatic calcifications. 5. Adynamic bone diseases are rare before hemodialysis in the absence of aluminum exposition by the drinking water or the aluminum phosphate binders. In absence of aluminum it will be prevented by maintaining PTH in its optimal range. 6. Osteomalacia before hemodialysis is mainly due, in the absence of aluminum exposition, to vitamin D deficiency, hypocalcemia and acidosis. It is readily cured by physiological doses of native vitamin D or 25OH vitamin D3 bringing plasma 25 OHD above 16 ng/ml, in association with alkaline salts of calcium and if necessary of sodium bicarbonate. PMID- 11117107 TI - [Primary hyperparathyroidism]. AB - Primary hyperparathyroidism is the third most frequent endocrine disorder. The condition required for diagnosis is inappropriately elevated secretion of parathyroid hormone (PTH) with respect to calcemia. Most often, the disease is due to a parathyroid adenoma, i.e. a monoclonal benign parathyroid tumor, less often to a parathyroid hyperplasia. The main tumorogenic mechanisms currently proposed are a DNA rearrangement in the PTH locus (transposition of the PTH promoter upstream to Cyclin D1/PRAD 1 gene) and a mutation of the gene responsible for multiple endocrine neoplasia type I. The clinical presentation has strikingly evolved towards a milder, asymptomatic form, frequently diagnosed on systematic screenings. Though the mechanism of hypercalcemia is better understood, several hypothesis are still being considered about the regulation of tumoral PTH secretion: the role of the expression of calcium-receptor in parathyroid gland cells, vitamin D receptor and estrogen receptor polymorphisms, etc. Surgery is still advised for symptomatic forms of the disease, either because of a bone involvement, or because of an evolutive nephrolithiasis. In the near future, the new calcium-receptor agonists could be a relevant therapeutic approach. PMID- 11117108 TI - [Voluntary lithium salt poisoning; risks of slow release forms]. AB - Lithium carbonate has been an invaluable drug in the treatment of manic depressive illness. At the present time slow-release forms are available for better stability of circulating drug level with one daily take. However the better stability of lithiemia with these forms has to be balanced with a higher toxic risk in cases of lithium carbonate overdose because of continuous release. We report three cases of overdose with sustained-release lithium salts reaching life threatening complications. Two hemodialysis sessions were necessary to control plasma lithium levels. In two cases, lithiemia rebond was observed after the first hemodialysis session, and was associated with severe neurological disorders. This publication pointed out the therapeutic rule of at least two hemodialysis sessions in cases of severe slow release lithium carbonate salts intoxication. PMID- 11117109 TI - [Alport's syndrome: apropos of 2 families]. AB - Two families with Alport's syndrome are presented. In the first family, the mode of inheritance is X-linked dominant; the molecular defect involves the alpha 5 chain of type IV collagen, absent from the glomerular and epidermal basal membrane in hemizygous males. In the other family, the transmission is autosomal recessive and the defect involves alpha 3(IV), absent from the glomerular basement membrane in homozygotes but alpha 5(IV) is normally present in the epidermal basement membrane. The consequences of these data on diagnosis and genetic counseling are discussed. PMID- 11117111 TI - The journal's journey PMID- 11117110 TI - Our past is our bridge to the future PMID- 11117115 TI - Interactive health communication on the Internet:|| advantages, disadvantages and implications for health|| professionals. PMID- 11117133 TI - Facility profile. Doctor writes Rx for design. Loyola University Medical Center, Department of Neurological Surgery. PMID- 11117134 TI - The big picture. PMID- 11117135 TI - Watts up? PMID- 11117136 TI - Making waves. PMID- 11117140 TI - Federation makes pitch for BBA relief at summer|| congressional hearings. PMID- 11117137 TI - Shop smart. Fight pollution--practice EPP. PMID- 11117144 TI - Depression in the workforce. Who will insist on good|| care? PMID- 11117142 TI - U.S. Senator Spencer Abraham (R-MI). A healthcare|| legislative update: the Health Care Provider Bill of Rights (S.|| 2999). PMID- 11117149 TI - Productivity plus. TAG, you're it! PMID- 11117145 TI - How business coalitions simplify health care|| business. PMID- 11117151 TI - Differential validity of the Defense Mechanism Manual for the TAT between Asian Americans and Whites. Thematic Apperception Test. AB - Thematic Apperception Test (Murray, 1943) responses of 69 Asian American (hereafter, Asian) and 83 White students were coded for defenses according to the Defense Mechanism Manual (Cramer, 1991b) and studied for differential validity in predicting paper-and-pencil measures of relevant constructs. Three tests for differential validity were used: (a) differences between validity coefficients, (b) interactions between predictor and ethnicity in criterion prediction, and (c) differences between groups in mean prediction errors using a common regression equation. Modest differential validity was found. It was surprising that the DMM scales were slightly stronger predictors of their criteria among Asians than among Whites and when a common predictor was used, desirable criteria were overpredicted for Asians, whereas undesirable ones were overpredicted for Whites. The results were not affected by acculturation level or English vocabulary among the Asians. PMID- 11117152 TI - Prototypical profiles of the Brief Psychiatric Rating Scale. AB - Three prototypical profiles of the Brief Psychiatric Rating Scale (BPRS; Overall & Gorham, 1962) were isolated using a Q-type factor-analytic strategy with a sample of homeless men with mental illness (N=165). The 3 profiles--depressed, actively psychotic, and withdrawn--were used to study changes in BPRS profiles over time in a control group and a group that received assertive community treatment (ACT). Over2 time periods (inception to 12 months and 12-24 months), the 2 groups did not differ in terms of changes in profile shape, but they did differ in terms of changes in profile elevation. The ACT group evidenced a decrease in symptom severity during the last 12 months, whereas the control group showed an increase. Although changes in profile shape in both groups did occur, there was a significant tendency for the shape of the BPRS profiles to remain stable from the inception of the study to the 12-month assessment and from that time to the 24-month assessment. We describe the uses of these prototypical profiles and discuss the applicability of this analytical approach to other assessment instruments. PMID- 11117153 TI - MMPI-2 in the inpatient assessment of women with eating disorders. AB - The MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) reveals similar patterns across all Diagnostic and Statistical Manual of Mental Disorders (4th ed.; American Psychiatric Association, 1994) eating-disorder diagnoses. In this study, 550 women with eating disorders completed the MMPI-2. The 3 highest mean elevations for all eating-disorder diagnostic groups occurred on the same scales in the same order: 2, 7, and 3. The modal code for all groups was 2-7/7-2. However, multivariate analyses using the 16 validity and clinical scales, as well as the 27 content and supplementary scales, indicated that the MMPI-2 also distinguishes among eating disorders, especially in that patients with restricting anorexia report less psychopathology than other groups. These results are compared with the results of past eating-disorder research that used the older MMPI (Hathaway & McKinley, 1983). PMID- 11117154 TI - On assessing individual differences in rumination on sadness. AB - The Rumination on Sadness Scale (RSS), an individual-difference measure of rumination on sadness, was developed as an alternative to the Ruminative Responses scale of the Response Styles Questionnaire (RRRSQ; Nolen-Hoeksema & Morrow, 1991). Research has shown the RRRSQ to consist of multiple, not highly intercorrelated factors; only I factor explicitly addresses rumination. In Study 1, a 1-factor solution to a principal components analysis was shown to hold for responses to the RSS. The RSS was also shown to be reliable. In Study 2, convergent and discriminant validity of the RSS were assessed. In Study 3, individuals with high RSS scores exhibited more distress regarding current concerns with the introduction of a delay period (to allow them to ruminate) after a sad mood induction. PMID- 11117155 TI - A relational interpretation of the Rorschach color determinants. AB - This article presents a contemporary relational interpretation of the Rorschach that is consistent with the empirical features of the Comprehensive System. The specific focus is on color determinants and the FC:CF+C ratio. The approach follows that of Schachtel ( 1959), who argued that how one perceives others reveals the quality of relatedness between oneself and others. Schachtel identified a developmental sequence of relatedness (perceptual-relatedness modes) and linked these levels of perceiving and relating to the Rorschach color determinants. I suggest that this developmental sequence, elaborated in a contemporary context, defines the expected or normative course of relatedness, whether across a lifetime, a particular relationship, or an interpersonal encounter. I further propose that relationships emerge and develop through organized trial-and-error activity, in the sense of Piaget's (1952) trying out (assimilating) and simultaneously modifying (accommodating) one's understanding within a relationship. Relatedness levels and the developmental transitions between them are described in terms of the underlying assimilation and accommodation processes. Within this general approach, the FC:CF+C ratio is defined as reflecting styles of relating to one's interpersonal environment, with each relational style based on normative or on variant relational development. Four fundamental relational styles-healthy, egocentric, veneered egocentric, and defensive-are described and coordinated with the Comprehensive System FC:CF+C potential findings. PMID- 11117156 TI - Cross-validation of the MMPI-2 in detecting malingered posttraumatic stress disorder. AB - We attempted to cross-validate findings from a previous study (Elhai, Gold, Sellers, & Dorfman, in press) using a clinical sample of combat-related war veterans to distinguish genuine from malingered posttraumatic stress disorder (PTSD) on the MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989). The MMPI-2 scores of 124 male combat war veterans at the PTSD outpatient treatment program of a Veterans Affairs Medical Center were compared with those of 84 adult college students instructed and trained to malinger PTSD. MMPI-2 overreporting variables examined were F, F-Fb, F-K, F(p), Ds2, O-S, OT, and FBS. A stepwise discriminant analysis identified F. F-Fb, F-K, Ds2, O-S, and OT as the best malingering predictors. A predictive discriminant analysis yielded good hit rates for the model with impressive cross-validation results. We assessed cutting scores for the predictors of the model. We discuss clinical implications for using the MMPI-2 to distinguish malingered PTSD from combat-related PTSD. PMID- 11117157 TI - Assessing depression with the MMPI and MMPI-2. AB - The usefulness of the MMPI (Hathaway & McKinley, 1951 ) and MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) for diagnosing and assessing symptomatic depression has been the subject of considerable debate for a number of years. In this article, we review the relative contributions of the MMPI and MMPI-2 clinical and content scales in predicting depression. Positive predictive power, negative predictive power, and overall classification rate were computed for Scale 2 (D) of the MMPI and MMPI-2 and the Depression content scale (DEP) of the MMPI-2. Scale 2 (D) of both the MMPI and MMPI-2 appears to be moderately accurate in predicting depression. Although some studies suggest that the content scale DEP provides incremental validity over Scale 2 (D) of the MMPI-2, the results of this review indicate that the content scale DEP of the MMPI-2 does not exceed the diagnostic efficiency of Scale 2 in predicting depression. PMID- 11117158 TI - Interpersonal dependency and personality pathology: variations in Rorschach Oral Dependency scores across Axis II diagnoses. AB - Although several investigations have examined the relationship of Rorschach Oral Dependency (ROD; Masling, Rabie, & Blondheim, 1967) scores to Axis I diagnosis, there has been very little research assessing variations in ROD scores across Axis II personality disorders (PDs). In this study, ROD scores were compared in 5 PD groups (borderline PD inpatients, borderline PD outpatients, avoidant dependent PD outpatients, narcissistic PD outpatients, and antisocial PD outpatients), and 2 non-PD comparison groups (psychotic disorder inpatients and college students). Borderline PD inpatients had significantly higher ROD scores than borderline PD outpatients, antisocial PD outpatients, and college students; no other between-group differences were found. We discuss implications of these results for research on dependency and Axis II psychopathology and offer suggestions for future studies. PMID- 11117159 TI - Effect of altered instructions on the MMPI-2 profiles of college students who are not motivated to distort their responses. AB - Research has shown that when the MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) is administered with instructions designed to make people aware of the validity indexes and when people are encouraged to respond honestly rather than invalidate the test through defensiveness, their scores on validity scales suggest a more candid responding. Before such modified administration procedures can be broadly used to increase the validity of the MMPI (Hathaway & McKinley, 1943) profiles, it is necessary to have more information about the effect of the altered instructions on test performance of people in general. This study involved administering the MMPI-2 to a group of 218 "normals" (college students) with instructions that differed from traditional instructions. Specifically, the test instructions were altered to explain more about the test and the presence of validity scales to apprise participants that disingenuous responding can be detected. The responses of college students taking the test under altered instructions were compared with those of a similar sample of 150 college students who took the test under standard instructions. Although under altered conditions there was a statistically significant tendency for measures assessing defensiveness (L, K, and S) to be lower for some participants (women but not men), the differences were trivial. Modified instructions made little practical difference in test performance when people who were not motivated to deceive on the test were informed of the presence of validity checks on the test. PMID- 11117160 TI - Convergent validity of alternative MMPI-2 personality disorder scales. AB - The Morey, Waugh, and Blashfield (1985) MMPI (Hathaway et al., 1989) personality disorder scales provided a significant contribution to personality disorder research and assessment. However, the subsequent revisions to the MMPI and the multiple revisions to the diagnostic criteria sets that have since occurred may have justified comparable revisions to these scales. Somwaru and Ben-Porath (1995) selected a substantially different set of items from the MMPI-2 (Butcher, Dahlstrom, Graham, Tellegen, & Kaemmer, 1989) to assess Diagnostic and Statistical Manual of Mental Disorders (4th ed.; American Psychiatric Association, 1994) personality disorder diagnostic criteria. In our study, we compared the convergent validity of these alternative MMPI-2 personality disorder scales with respect to 3 self-report measures of personality disorder symptomatology in a sample of 82 psychiatric outpatients. The results suggested that Somwaru and Ben-Porath's scales are as valid as the original Morey et al. scales and might be even more valid for the assessment of borderline, antisocial, and schizoid personality disorder symptomatology. PMID- 11117161 TI - Looking, speaking, and acting like it. PMID- 11117198 TI - The social organization of fish shoals: a test of the predictive power of laboratory experiments for the field. AB - By contrast with a multitude of laboratory studies on the social organization of fish, relatively little is know about the size, composition and dynamics of free ranging fish shoals. We give an overview of the available information on fish shoals and assess to what degree the predictions made from laboratory studies are consistent with field data. The section on shoal choice behaviour in the laboratory is structured so that the evidence for different shoaling preferences is discussed in the context of their mechanisms and functions. Predictions based on experiments in captivity regarding preferences for conspecifics, individuals of similar body length and unparasitized fish were highly consistent with field observations on free-ranging shoals whereas preferences for familiar conspecifics and kin remain to be conclusively demonstrated in the field. In general, there is a shortage of studies in which shoaling preferences have been investigated both in the laboratory and the field, and field studies have so far been largely descriptive revealing little about the underlying mechanisms of observed patterns. Given the great importance of fish shoals both in fundamental and applied research, an advancement of our knowledge of their social organization should significantly contribute to a better understanding of a whole range of topics including reciprocal altruism, group-living and self-organization. PMID- 11117199 TI - Individual feeding specialisation in shorebirds: population consequences and conservation implications. AB - Individual feeding specialisation in shorebirds is reviewed, and the possilble mechanisms involved in such specialisations. Any specialisation can he seen as an individual strategy, and the optimum strategy for any given individual will be conditional upon its specific priorities and constraints. Some specialisations are related to social status and some to individual skills. Some are also probably frequency-dependent. However, most shorebird specialisations are constrained to a large extent by individual morphology, particularly bill morphology. For example, larger birds are able to handle larger prey, and birds with longer bills are able to feed on more deeply buried prey. Sex differences in bill length are uncommon in the Charardriidae, which are surface peckers, but are common in the Scolopacidae, which feed by probing in soft substrates. Sex differences in bill morphology are frequently associated with sex differences in feeding specialisation. There is evidence that different feeding specialisations are associated with different payoffs, in which case the probability of failing to reproduce or of dying will not be distributed equally throughout the population. I consider the population consequences of such feeding specialisations, particularly the different risks and benefits associated with different habitats or diets. I also consider the way in which individuals may differ in their response to habitat loss or change. I suggest that population models designed to predict the effect of habitat loss or change on shorebirds should have the ability to investigate the differential response of certain sections of the population, particularly different ages or sexes, that specialise in different diets or feeding methods. PMID- 11117200 TI - Thyroid hormones and their effects: a new perspective. AB - The thyroid hormones are very hydrophobic and those that exhibit biological activity are 3',5',3,5-L-tetraiodothyronine (T4), 3',5,3-L-triiodothyronine (T3), 3',5',3-L-triiodothyronine (rT3) and 3,5',-L-diiothyronine (3,5-T2). At physiological pH, dissociation of the phenolic -OH group of these iodothyronines is an important determinant of their physical chemistry that impacts on their biological effects. When non-ionized these iodothyronines are strongly amphipathic. It is proposed that iodothyronines are normal constituents of biological membranes in vertebrates. In plasma of adult vertebrates, unbound T4 and T3 are regulated in the picomolar range whilst protein-bound T4 and T3 are maintained in the nanomolar range. The function of thyroid-hormone-binding plasma proteins is to ensure an even distrubtion throughout the body. Various iodothyronines are produced by three types of membrane-bound cellular deiodinase enzyme systems in vertebrates. The distribution of deiodinases varies between tissues and each has a distinct developmental profile. Thyroid hormones. (1) the nuclear receptor mode is especially important in the thyroid hormone axis that controls plasma and cellular levels of these hormones. (2) These hormones are strongly associated with membranes in tissues and normally rigidify these membranes. (3) They also affect the acyl composition of membrane bilayers and it is suggested that this is due to the cells responding to thyroid-hormone-induced membrane rigidificataion. Both their immediate effects on the physical state of membranes and the consequent changes in membrane composition result in several other thyroid hormone effects. Effects on metabolism may be due primarily to membrane acyl changes. There are other actions of thyroid hormones involving membrane receptors and influences on cellular interactions with the extracellulara matrix. The effects of thyroid hormones are reviewed and appear to b combinations of these various modes of action. During development, vertebrates show a surge in T4 and other thyroid hormones, as well as distinctive profiles in the appearance of the deiodinase enzymes and nuclear receptors. Evidence from the use of analogues supports multiple modes of action. Re-examination of data from th early 1960s supports a membrane action. Findings from receptor 'knockout' mice supports an important role for receptors in the development of the thyroid axis. These iodothyronines may be better thought of as 'vitamone'-like molecules than traditional hormonal messengers. PMID- 11117201 TI - Stems, nodes, crown clades, and rank-free lists: is Linnaeus dead? AB - Recent radical proposals to overhaul the methods of biological classification are reviewed. The proposals of phylogenetic nomenclature are to translate cladistic phylogenies directly into classifications, and to define taxon names in terms of clades. The method has a number of radical consequences for biologists: taxon names must depend rigidly on the particular cladogram favoured at the moment, familiar names may be reassigned to unfamiliar groupings, Linnaean category terms (e.g. phylum, order, family) are abandoned, and the Linnaean binomen (e.g. Homo sapiens) is abandoned. The tenets of phylogenetic nomenclature have gained strong support among some vocal theoreticians, and rigid principles for legislative control of clade names and definitions have been outlined in the PhyloCode. The consequences of this semantic maelstrom have not been worked out. In pratice, phylogenetic nomenclature will bc disastrous, promoting confusion and instability, and it should be abandoned. It is based on a fundamental misunderstanding of the difference between a phylogeny (which is real) and a classification (which is utilitarian). Under the new view, classifications are identical to phlylogenies, and so the proponents of phylogenetic nomenclature will end up abandoning classifications altogether. PMID- 11117202 TI - The function significance of silk decorations of orb-web spiders: a critical review of the empirical evidence. AB - A number of taxonomically diverse species of araneoid spiders adorn their orb webs with conspicuous silk structures, called decorations or stabilimenta. The function of these decorations remains controversial and several explanations have been suggested. These include: (1) stabilising and strengthening the web; (2) hiding and concealing the spider from predators; (3) preventing web damage by larger animals, such as birds; (4) increasing foraging success; or (5) providing a sunshield. Additionally, they may have no specific function and are a consequence of stress or silk regulation. This review evaluates the strength of these explanations based on the evidence. The foraging function has received most supporting evidence, derived from both correlative field studies and experimental manipulations. This contrasts with the evidence provided for other functional explanations, which have not been tested as extensively. A phylogenetic analysis of the different decoration patterns suggests that the different types of decorations are as evolutionary labile as the decorations themselves: the analysis shows little homology and numerous convergences and independent gains. Therefore, it is possible that different types of decorations have different functions, and this can only be resolved by improved species phylogenies, and a combination of experimental and ultimately comparative analyses. PMID- 11117219 TI - Effects of trilinolein on superoxide dismutase activity and mrna levels in aortic smooth muscle cells. AB - 1. Atherosclerotic cardiovascular disease is still the leading cause of death in Western countries. Oxygen free radicals are considered to be intimately involved in the development of atherosclerosis. Anti-oxidants may help to protect mammalian cells from the damage induced by these reactive oxygen species. Many reports have indicated that anti-oxidants used in the treatment or prevention of disease could modify the levels of superoxide dismutase (SOD). However, the effects of long-term anti-oxidant treatment on the levels of SOD in smooth muscle cells (SMC) is still unclear. In the present study, the effects of the lipophilic anti-oxidant trilinolein on the activity and gene expression of SOD in SMC were evaluated. 2. After 2 days incubation with 0.1 micromol/L trilinolein, the activity and mRNA levels of SOD were increased in rat aortic SMC (A7r5), but there was no significant change in these parameters with a higher concentration of 1 micromol/L trilinolein. 3. In contrast, after 7 days incubation with trilinolein, both the activity and mRNA levels of SOD were lowered in a dose dependent manner. 4. These data emphasize the importance of choosing an optimal dosage for supplementation with anti-oxidants in humans for the scavenging of oxygen free radicals. PMID- 11117220 TI - Responses of regional kidney perfusion to vasoconstrictors in anaesthetized rabbits: dependence on agent and renal artery pressure. AB - 1. We tested the effects of intravenous infusions of angiotensin II (AngII; 300 ng/kg per min) and the vasopressin V1 receptor agonist [Phe2,Ile3,Orn8] vasopressin (30 ng/kg per min) on regional kidney perfusion in an extracorporeal circuit model in anaesthetized rabbits in which renal artery pressure (RAP) can be set independently of systemic mean arterial pressure. To test whether the level of RAP can influence the renal vascular response to [Phe2,Ile3,Orn8] vasopressin, we compared its effects when RAP was initially set at approximately 65 mmHg with those when RAP was set at approximately 130 mmHg. 2. When RAP was initially set at approximately 65 mmHg, a 20min infusion of AngII increased RAP (13%) and reduced renal blood flow (RBF; 50%) and cortical perfusion (CBF; 43%). Medullary perfusion (MBF) transiently increased during the first 10 min of infusion, but was not significantly different from control levels during the final 5 min of infusion. 3. When RAP was initially set at approximately 65 mmHg, a 20 min infusion of [Phe2,Ile3,Orn8]-vasopressin increased RAP (9%) and reduced RBF (21%); MBF was reduced by 57%, but CBF was reduced by only 15%. In contrast, when RAP was initially set at approximately 130 mmHg, infusion of [Phe2,Ile3,Orn8]-vasopressin reduced RAP (7%) and increased RBF (13%). In these experiments, MBF was reduced by 38%, but CBF increased by 6%. 4. Our experiments show that AngII preferentially reduces CBF, while [Phe2,Ile3,Orn8]-vasopressin preferentially reduces MBF. The renal vascular responses to [Phe2,Ile3,Orn8] vasopressin appear to be profoundly affected by the level of RAP, because increasing RAP from approximately 65 to approximately 130 mmHg transforms its cortical vasoconstrictor effect into cortical vasodilatation while leaving the response of the medullary microvasculature relatively unchanged. Whether renal vascular responses to other vasoactive agents (e.g. AngII) are similarly affected by the level of RAP remains to be determined. PMID- 11117221 TI - Proceedings of the Australian Physiological and Pharmacological Society symposium: integrative cardiovascular function. A symposium to honour Saxon White. Newcastle, Australia, September 1999. PMID- 11117222 TI - Lymphatic vasomotion. AB - 1. Experimental findings in the past decade have greatly advanced present understanding of electrical/mechanical rhythmicities in smooth muscle, including vasomotion. Lymphatic vessels show strong vasomotor activity and have provided a key experimental system to study these processes. 2. Evidence from lymphatic vessels, blood vessels and other smooth muscles indicates that rhythmical contractions arise through a Ca2+ store-controlled pacemaker mechanism, which can function to cause smooth muscle constriction. 3. Such a model fits with observations that vasomotion can be near synchronous over large vessel lengths involving many cells. 4. The alternative interpretation that smooth muscle rhythmicities are generated by a cardiac-like electrical pacemaker mechanism has not been substantiated in any smooth muscle preparation under normal physiological conditions. However, elements of this latter mechanism are likely to be present at least in some smooth muscles, serving to modulate pacemaking. PMID- 11117224 TI - Heart-lung interactions: the sigh and autonomic control in the bronchial and coronary circulations. AB - 1. The Darwin hypothesis that human and animal expressions of emotion are the product of evolution and are tied to patterns of autonomic activity specified to progress the emotion remains under challenge. 2. The sigh is a respiratory behaviour linked with emotional expression in animals and humans from birth to death. The aim of the present study was to explore Darwin's hypothesis with respect to tied autonomic activity underlying sigh-induced changes in the bronchial and coronary circulations. 3. Awake dogs were prepared using pulsed ultrasonic flow probes on the right bronchial artery, parent intercostal artery and brachial artery, or on the right, circumflex and anterior descending coronary arteries. Central venous (CVP) and arterial pressures (AP) were measured; heart rate and flow conductances were derived. Three spontaneous sighs were monitored before and during random blockade of individual and combinations of cholinoceptors, alpha-adrenoceptors and beta-adrenoceptors using methscopolamine, phentolamine and propranolol infusions. The data were subject to a 2(3) factorial analysis. 4. A spontaneous sigh is marked by a transient fall and return (< 3 s) in CVP of 18 mmHg (from 4 +/- 1 to -14 +/- 2 mmHg), usually followed by apnoea lasting 23 +/- 2 s. There is an immediate tachycardia and small rise in AP (phase 1) then, during apnoea, bradycardia and a fall in AP (phase 2). During phase 2, bronchial and coronary blood flow and conductance rise two- to three-fold over 30s (peak at 8s). The vascular changes are absent in parent intercostal and brachial beds. PMID- 11117223 TI - Quantitative analysis of vascular to cardiac selectivity of L- and T-type voltage operated calcium channel antagonists in human tissues. AB - 1. Classical L-type voltage-operated calcium channel (VOCC) antagonists dilate blood vessels, depress myocardial contractility and slow cardiac conduction. 2. We compared four L-type VOCC antagonists and a novel tetralol derivative, mibefradil, reportedly 10-fold more selective for T- (transient) over L-type VOCC in two in vitro assays of human tissue, namely isolated small arteries from the aortic vasa vasorum in a myograph and right atrial trabeculae muscle under isometric force conditions. 3. In arteries contracted with K+ (62 mmol/L), the relaxation pIC50 values for the VOCC antagonists felodipine, nifedipine, amlodipine, verapamil and mibefradil were 8.30, 7.78, 6.64, 6.26 and 6.22, respectively. In atrial trabeculae, the pIC50 values to inhibit the inotropic response to a submaximal concentration of isoprenaline (6 nmol/L) for felodipine, nifedipine, verapamil, amlodipine and mibefradil were 7.21, 6.95, 6.91, 5.94 and 4.61, respectively. 4. Taking the anti-log (pIC50 vessel-pIC50 atrium) the vascular relaxation to cardiac depression potency ratios for mibefradil, felodipine, nifedipine, amlodipine and verapamil were 41, 12, 7, 5 and 0.22, respectively. 5. We conclude that, in human tissue assays, perhaps T- over L-type VOCC selectivity confers the most favourable vascular selectivity on mibefradil. Alternatively, splice variants of L-type VOCC in the vasculature (CaV1.2b) may be more sensitive to mibefradil than the splice variants in the heart (CaV1.2a). PMID- 11117225 TI - Effect of fentanyl on baroreflex control of circumflex coronary conductance. AB - 1. Fentanyl, a synthetic mciro-opioid receptor agonist, is the preferred induction and maintenance anaesthetic agent in cardiac surgery. 2. Its actions on myocardial blood flow are poorly understood. There are reports of intra-operative myocardial ischaemia. Its reported actions on cardiorespiratory control vary widely, but do involve hypertension, bradycardia and peripheral vasoconstriction. 3. Accordingly, the postulate that fentanyl would cause coronary vasoconstriction and myocardial disadvantage was examined in awake dogs with a continuous wave Doppler flow probe mounted on the circumflex coronary artery. 4. Continuous intravenous infusion of fentanyl citrate (550 ng/kg per min) raised plasma concentrations of fentanyl to 3.37 ng/mL in a linear fashion at 20 min. There was a fall in core temperature of 0.7 degrees C and, although no apparent depression of ventilation or fall in arterial or coronary sinus PO2, there was a rise in PCO2 and H+ concentration. Some dogs salivated and panted transiently. Thus, fentanyl may reset temperature regulation in low doses but, at higher doses, is associated with metabolic acidosis. 5. In sinus rhythm, the arterial pressure of the dogs fell slightly, then rose to 115% of resting control. Circumflex flow and conductance rose early, then conductance steadily declined to 83%. Heart rate fell, then rose before returning to pre-infusion levels. The early circumflex coronary vasodilator effects, but not the later vasoconstrictor effects, were reduced in dogs with paced hearts. 6. In dogs with paced hearts, a dose-effect study using 138, 275, 550 and 1100 ng/kg per min fentanyl suggested that, at low plasma concentrations of 1-2 ng/mL, vasodilatation does occur in both coronary and systemic circulations; however, at higher doses, intense coronary and systemic vasoconstriction supervenes. 7. The dose-response effect of fentanyl on arterial baroreflex control of circumflex conductance was examined during the immediate 8 s circumflex vasodilator response to a step rise in aortic pressure caused by inflation of an intra-aortic balloon. At low plasma concentrations of fentanyl, baroreflex control of circumflex conductance appears to be enhanced but, with increasing plasma concentrations of fentanyl, appears to be depressed. 8. Therefore, the effects of fentanyl are dose dependent. At low plasma concentrations, left ventricular blood flow and its baroreflex control is enhanced but, at higher concentrations, it is depressed. PMID- 11117226 TI - Effects of anaesthesia on regional coronary control mechanisms. AB - 1. The regional coronary circulation is under the control of local metabolic and myogenic factors, but is also influenced by autonomic systems, including sympathetic and parasympathetic nerves. 2. General anaesthetic agents influence not only local control through changes in metabolic demand, but also neural control through suppression of autonomic influence. 3. Anaesthetic agents have differing effects on reflex control systems, which are dependent on coronary territory and ventricular rate. 4. Effects of anaesthesia should be taken into account when interpreting results in anaesthetized models of coronary control. PMID- 11117227 TI - Recent views on integrated coronary control: significance of non-uniform regional control of coronary flow conductance. AB - 1. Previous work from this laboratory and others has shown that powerful autonomic influences modulate coronary flow. In particular, the parasympathetic nervous system produces vasodilatation when activated by baroreceptors via the vagus nerve. 2. Differences exist in baroreflex coronary vasodilator mechanisms among the right, circumflex and anterior descending coronary vascular beds in the awake chronically instrumented dog. 3. Our hypothesis is that neurogenic acetylcholine acting from the adventitial side and endothelial nitric oxide from the luminal aspect of coronary smooth muscle compete with powerful intrinsic myogenic constrictor mechanisms to regulate regional flow conductance. 4. There is also increasing evidence that heterogeneity of control systems exists in different-sized coronary vessels within an individual coronary vascular bed. 5. It is concluded that coronary vessels in vascular beds can no longer be assumed to respond in a uniform manner to neural, myogenic, metabolic or humoral factors. 6. These new perspectives of regional control mechanisms have important implications for understanding pathophysiological mechanisms inducing and sustaining tachyarrhythmias involved in ischaemic heart disease. PMID- 11117228 TI - Bronchovascular role in pulmonary congestion. AB - 1. A postulated role for the bronchial circulation in the development of pulmonary congestion may be based on recent studies of bronchovascular control. 2. The bronchial circulation is the nutrient blood supply of the conducting airways and, therefore, plays an important role in the function of the bronchial mucosa. Mucosal swelling secondary to elevation of mucosal capillary hydrostatic pressure may decrease airway calibre, increase resistance to airflow and precipitate symptoms of pulmonary congestion. 3. Resting mucosal capillary hydrostatic pressure is relatively constant due to autoregulation of bronchial blood flow and is maintained low by nett bronchovascular constriction due to the dominance of autonomic vasoconstriction over nitric oxidedependent vasodilatation. 4. Bronchial blood flow is also regulated by cardiac afferent reflexes. Stimulation of cardiac vagal and spinal afferents produces vasodilatation and vasoconstriction, respectively. Tonic activity of cardiac spinal afferents probably contributes to the resting autonomic vasoconstriction. 5. Therefore, mild heart failure, which is associated with abnormal cardiovascular reflex function, may decrease cardiac spinal afferent-mediated bronchial vasoconstriction and produce active dilatation due to stimulation of cardiac vagal afferents by excessive myocardial stretch, leading to bronchial mucosal swelling and pulmonary congestion. PMID- 11117229 TI - What drives the tonic activity of presympathetic neurons in the rostral ventrolateral medulla? AB - 1. The present review discusses the mechanisms that maintain the tonic activity of presympathetic cardiovascular neurons in the rostral part of the ventrolateral medulla. 2. Experimental evidence is reviewed that indicates that these neurons receive both tonic excitatory and tonic inhibitory synaptic inputs. The former appear to be mediated, at least in part, by glutamate receptors and the latter appear to be mediated by GABA receptors. 3. There is also evidence that these neurons have the capacity to generate action potentials in the absence of synaptic inputs. However, at present, there is not clear evidence that such an intrinsic pacemaker-like mechanism contributes to the tonic activity of these neurons under normal resting conditions. 4. These neurons are also chemosensitive and this may contribute to their tonic activation under conditions of hypoxia or hypercapnia. PMID- 11117230 TI - Effect of plasma cholesterol on red blood cell oxygen transport. AB - 1. Oxygen (O2) transfer from the blood to tissues is a function of the red blood cell (RBC) O2 saturation (SO2), the plasma O2 content being negligible. Under conditions of increased tissue O2 demand, the SO2 of arterial blood does not change appreciably (97%); however, the SO2 of mixed venous blood, equal to that of the perfused tissues, can go as low as 20%. 2. Tissue O2 availability is limited by the exposure time to a RBC, which decreases under conditions of maximum stress (< 1 s). If the O2 unloading time was to increase significantly, because of a decrease in the RBC diffusion constant or an increase in the RBC membrane thickness, the RBC O2 unloading time would exceed tissue (e.g. cardiac) transit time and O2 transfer would be impaired. 3. Cholesterol constitutes the non-polar, hydrophobic lipid of the enveloping layer of the RBC membrane. As the cholesterol content of the RBC increases, the fluidity of the membrane decreases and the lipid shell stiffens. 4. Early studies demonstrated that high blood cholesterol concentrations were associated with reduced blood O2 transport; in essence, the haemoglobin dissociation curve was shifted to the left. 5. Current investigations have shown that the cholesterol RBC membrane barrier to O2 diffusion delayed O2 entry into the RBC during saturation and delayed O2 release from the RBC during desaturation. In an analysis of 93 patients divided by their cholesterol concentration into five groups, the percentage change in blood O2 diffusion was inversely proportional to the cholesterol concentration. 6. The RBC membrane cholesterol is in equilibrium with the plasma cholesterol concentration. It stands to reason that as the plasma cholesterol increases, the RBC membrane becomes impaired and O2 transport is reduced. 7. The implications of this new perspective on O2 transport include the ability to increase tissue oxygenation by lowering plasma cholesterol. PMID- 11117231 TI - Neural regulation of renal blood flow: a re-examination. AB - 1. The importance of renal sympathetic nerve activity (RSNA) in the regulation of renal function is well established. However, it is less clear how the renal vasculature responds to the different mean levels and patterns of RSNA. While many studies have indicated that small to moderate changes in RSNA preferentially regulate renin secretion or sodium excretion and only large changes in RSNA regulate renal blood flow (RBF), other experimental evidence suggests that small changes in RSNA can influence RBF 2. When RSNA has been directly measured in conjunction with RBF, it appears that a range of afferent stimuli can induce reflex changes in RBF. However, many studies in a variety of species have measured RBF only during stimuli designed to reflexly increase or decrease sympathetic activity, but have not recorded RSNA. While this approach can be informative, it is not definitive because the ability of the vasculature to respond to RSNA may, in part, reflect the resting level of RSNA and, therefore, the vasoconstrictive state of the vasculature under the control conditions. 3. Further understanding of the control of RBF by RSNA has come from studies that have analysed the underlying rhythms in sympathetic nerve activity and their effect on the cardiovascular system. These studies show that the frequency response characteristic of the renal vasculature is such that higher frequency oscillations in RSNA (above 0.6 Hz) contribute to setting the mean level of RBF. In comparison, lower frequency oscillations in RSNA can induce cyclic vasoconstriction and dilation in the renal vasculature, thus inducing oscillations in RBF. 4. In summary, the present review discusses the neural control of RBF, summarizing evidence in support of the hypothesis that RBF is under the influence of RSNA across the full range of RSNA. PMID- 11117232 TI - The compromised intra-uterine environment: implications for future lung health. AB - 1. Epidemiological studies of infants, children and adults indicate that prenatal compromises that restrict fetal growth and cause low birthweight increase the risk of respiratory deficiencies after birth. 2. It is apparent that the lung has a limited ability to recover from early developmental compromises and that altered development can permanently impair lung architecture. 3. Lung development in utero can be adversely affected by factors associated with fetal growth restriction, namely fetal hypoxaemia, reduced substrate supply and hypercortisolaemia. 4. We have conducted a series of studies of respiratory development in chronically catheterized ovine fetuses and postnatal lambs in which growth restriction was induced during late gestation by embolizing the umbilico-placental vascular bed, a technique that replicates key aspects of human placental insufficiency. 5. During late gestation, restricting the growth of the ovine fetus did not alter lung weight or lung liquid secretion or volume when each factor was related to bodyweight, but it did lead to increased lung DNA concentrations and an increased thickness of the air-blood barrier. Expression of pulmonary surfactant proteins A, B and C were not altered and, hence, it was unlikely that surfactant protein synthesis had been impaired by growth restriction. 6. When growth restriction continued to term, lambs were born with a low birthweight and remained small compared with controls for 8 weeks after birth. Low-birthweight lambs were mildy hypoxaemic and compliances of their lungs and chest wall were, respectively, decreased and increased relative to controls. Pulmonary surfactant proteins A, B and C were not deficient, indicating that decreased lung compliance most likely had a structural basis. PMID- 11117233 TI - Acute effects of moderate exercise on serum lipids, lipoproteins and apolipoproteins in sedentary young women. AB - 1. The aim of the present study was to examine the effects of moderate exercise on serum lipids, lipoproteins and apolipoproteins in seven sedentary young women under controlled conditions. 2. The subjects exercised on separate days for 30 or 60 min at an intensity of 60% of maximal oxygen uptake on a cycle ergometer. Total cholesterol, triglycerides, high-density lipoprotein-cholesterol (HDL-C), HDL2-C, HDL3-C, low-density lipoprotein-cholesterol, apolipoproteins A-I, A-II and B were measured in the serum at the end of the 60 min rest period before each exercise, immediately after the performance of each exercise and at 30 min and 1, 2 and 24 h after each exercise. 3. The results showed that there were no significant differences between the pre- and postexercise samples for any of the parameters tested. 4. The results of the present study suggest that a single bout of exercise designed to simulate a typical training workout has no noticeable effect on serum lipids, lipoproteins and apolipoproteins in normal sedentary young women who have normal lipid profiles, are in the follicular phase of their menstrual cycle and who consume a relatively low-fat diet. PMID- 11117234 TI - Changes in absorptive function of rat intestine injured by methotrexate. AB - 1. Methotrexate (MTX), an anticancer drug, has been shown to induce acute injury in the small intestine. The present study was designed to investigate the in vivo absorptive function of the small intestine injured by MTX using an amino-beta lactam antibiotic cephalexin (CEX). Time-dependent changes in diamine oxidase (DAO) and alkaline phosphatase (ALP) activity in the small intestine and histopathological findings were also measured in rats treated with MTX (20 mg/kg). 2. Most severe mucosal damage was observed 2 days after MTX treatment and the area under the plasma concentration-time curve of CEX (AUC(CEX)) following oral administration of 20 mg/kg tended to decrease. Thereafter, the AUC(CEX) increased significantly and the histopathological changes diminished within 5 days. 3. Both villus height and mucosal weight followed the same pattern, decreasing in the first 2 or 3 days following treatment, increasing on the 5th day and returning to control levels by the 10th day. Methotrexate-induced changes in the mucosal wet weight/whole intestinal weight ratio were significantly correlated with those of AUC(CEX), but did not correlate with mucosal DAO and ALP activity. 4. These findings provide evidence that the change in the total amount of CEX is an index of the active transport function, probably by intestinal peptide transporter (PEPT1), and is well reflected by histopathological changes in the intestinal mucosa induced by MTX. In addition, there is a possibility that this method could be applied in the clinical setting for diagnosis of intestinal status and absorptive function. PMID- 11117235 TI - Development of an experimental model of liver cirrhosis in rabbits. AB - 1. The aim of the present study was to develop an experimental model of liver cirrhosis in rabbits using CCl4 and phenobarbital. 2. Liver cirrhosis was induced in male New Zealand white rabbits (n = 10) by intragastric administration of CCl4 once weekly starting 14 days after the addition of phenobarbital to the drinking water (50 mg/day). Controls received phenobarbital only (n = 7). Alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GGT), albumin and bilirubin levels were determined throughout CCl4 treatment. The initial dose of CCl4 was 20 microg and subsequent doses were calculated to maintain AST and ALT levels between 400 and 800 IU/L for the duration of treatment (16 weeks). Indocyanine green (ICG) clearance was performed before and at the end of CCl4 treatment. Animals were killed at 16 weeks and three fragments of each liver lobe were processed for histological examination. A semiquantitative score was used to evaluate the development of fibrosis. 3. Cirrhosis developed in 80% of rabbits treated with CCl4. These animals did not gain weight compared with controls (P < 0.05). A significant reduction of ICG clearance was observed in CCl4-treated rabbits compared with controls (P < 0.05). The AST, ALT, bilirubin and gamma-GGT levels were elevated in CCl4-treated rabbits. 4. In conclusion, this model is successful in producing liver cirrhosis and may be useful in studies investigating metabolic, immunological or biochemical changes during the evolution of chronic liver disease. PMID- 11117236 TI - Glutathione metabolism in cyclosporine A-treated rats: dose- and time-related changes in liver and kidney. AB - 1. We investigated the simultaneous effects of cyclosporine A (CsA) treatment in rats on glutathione metabolism, oxidative status and their interorgan relationship in the liver and kidney. 2. Reduced and oxidized glutathione (GSH and GSSG, respectively), lipid peroxidation and the activity of several enzymes of the glutathione cycle were evaluated in adult Wistar rats treated daily (i.p.) with saline, CsA vehicle (olive oil) or CsA (10 and 20mg/kg per day) for either 1 or 4 weeks (short- and long-term treatments, respectively). 3. Cyclosporine A treatment elicited a significant depletion in liver GSH content and a decrease in the GSH/GSSG ratio that was unrelated to either the time of treatment or the dose used; these effects were already evident after 1 week of treatment. Renal GSH levels remained unaffected or increased, while those of GSSG increased markedly in all CsA-treated rats, leading to decreases in the GSH/GSSG ratio, except in rats treated in the short term with the lower dose of CsA. These changes in the GSH/GSSG ratio were time and dose dependent. Short-term CsA treatment using the higher dose and long-term treatment with both doses of CsA progressively enhanced lipid peroxidation, which was reflected by increased levels of thiobarbituric acid-reactive substances in both hepatic and renal homogenates. Hepatic gamma glutamylcysteine synthetase activity was increased after long-term treatment with both doses of CsA, whereas the activity of GSH hepatic peroxidase and GSH transferase was not significantly modified in any of the experimental groups. In contrast, renal gamma-glutamyl transpeptidase activity decreased in a progressive fashion, with the magnitude of this decrease being dose and time dependent. The plasma levels of total glutathione increased only in rats treated in the long term, regardless of the dose of CsA used, and remained unaltered in animals treated in the short term. 4. In summary, the data collected indicate that CsA treatment alters the interorgan homeostasis of glutathione and the oxidative status of the rat liver and kidney, which is associated with increases in lipid peroxidation in both organs, and also induces modifications in the activity of some enzyme related to the glutathione cycle. PMID- 11117237 TI - Ischaemic preconditioning ameliorates functional disturbance and impaired renal perfusion in rat ischaemia-reperfused kidneys. AB - 1. The effects of ischaemic preconditioning (IP) on renal function, haemodynamics and lipid peroxidation in the rat ischaemia-reperfused kidney model were examined. 2. In Wistar male rats, application of a single or three periods of 5 min bilateral renal ischaemia was performed prior to 30 min bilateral ischaemia and 90 min reperfusion (IR). The glomerular filtration rate (GFR) was estimated in terms of inulin clearance. Fractional excretion of sodium (FE(Na)) and lithium (FE(Li)), indicating total and proximal tubular sodium handling, respectively, was also measured and renal blood flow was monitored throughout the experiment. In addition, renal lipid peroxidation (LPO) levels in reperfused kidneys were evaluated. 3. A 2.8-fold increase in recovery of GFR (P < 0.005), a 50% reduction in FE(Na) (P < 0.005) and a 40% decrease in FE(Li) (P < 0.05) after IR resulted from the single period of 5 min IP. Renal blood flow was also higher than that in the control group (P < 0.01). No change of LPO levels was observed. 4. We conclude that IP may have an ability to ameliorate reperfused renal function and haemodynamics with a suitable period of preconditioned ischaemia, although this effect is independent of LPO. PMID- 11117238 TI - Comparison between DOQI and Canadian guidelines for peritoneal dialysis. AB - The recently published Canadian Society of Nephrology (CSN) Guidelines for Peritoneal Dialysis Adequacy differ in a number of ways from the United States National Kidney Foundation Dialysis Outcomes Quality Initiative (DOQI) guidelines on the same topic. The three main differences are (1) the CSN targets are the same for continuous ambulatory peritoneal dialysis (CAPD) and automated peritoneal dialysis (APD), whereas the DOQI targets are higher for APD; (2) the CSN guidelines have a lower creatinine clearance (CCr) target of 50 L/wk for low and low-average transporters compared to 60 L/wk for high and high-average transporters, but no such distinction is made by DOQI; and (3) the CSN has set lower limit targets as well as preferred targets for Kt/V and CCr. Other differences are that the CSN recommendations do not give Kt/V the same primacy over CCr that DOQI does. Also, the CSN recommendations give greater emphasis to the risks associated with high transport status. This review looks at the reasons for these differences and demonstrates that some are due to information that has become available since the publication of the DOQI guidelines. However, it is emphasized that, when recommendations are predominantly opinion- rather than evidence-based, geographic and economic factors may also account for differences. PMID- 11117239 TI - The pathophysiological puzzle of uremic pruritus--insights and speculations from therapeutic and epidemiological studies. PMID- 11117240 TI - Recommended peritoneal dialysis curriculum for nephrology trainees. The International Society for Peritoneal Dialysis (ISPD) Standards and Education Subcommittee. PMID- 11117241 TI - Role of surfactant in peritoneal dialysis. AB - Evidence is reviewed that demonstrates how the mesothelial cell in the normal peritoneum and comparable serosal cavities secretes surface-active phospholipid (SAPL) as a means of protecting itself and the membrane it forms with its neighbors. It is shown how SAPL, if adsorbed (reversibly bound) to mesothelium, can impart excellent lubricity, antiwear and release (antistick) properties, while impeding surgical adhesion formation. More-speculative benefits include acting as a deterrent to fibrosis and as a barrier to both protein leakage and pathogen invasion by spanning cell junctions. Such spanning would also "pin down" cell corners, impeding peeling as the first step in exfoliation encountered in prolonged continuous ambulatory peritoneal dialysis (CAPD). The molecular mechanism underlying each of these possible functions is adsorption. Morphological and hydrophobicity studies are discussed as validation for such an adsorbed lining and how it can be fortified by administering exogenous SAPL. Any role for SAPL in ultrafiltration is much more controversial. However, a surfactant lining can explain the very high permeability of the membrane to lipid soluble drugs, implying that it is a barrier to water-soluble solutes. The clinical and animal evidence is conflicting but would seem to be best explained by a role for the barrier in promoting semipermeability, and hence the osmotic driving force for water transmission. Thus, adsorption of exogenous SAPL in CAPD patients with low ultrafiltration seems to restore this barrier function. The future direction for surfactant in CAPD would seem to rest with the physical chemists in producing formulations that optimize adsorption, probably involving a compromise between water solubility and surface activity of the phospholipids selected. It might even warrant using the interdialytic interval for readsorbing SAPL without the problem of dilution by a large volume of dialysate. PMID- 11117242 TI - A long-term study of a bicarbonate/lactate-based peritoneal dialysis solution- clinical benefits. The Bicarbonate/Lactate Study Group. AB - OBJECTIVE: A bicarbonate/lactate peritoneal dialysis solution (Bic/Lac) has been developed based on in vitro and ex vivo data showing better preservation of cell function and correction of pain on infusion. DESIGN: This was a randomized, prospective, controlled, open-label study comparing a new 25 mmol/L bicarbonate/15 mmol/L lactate with a standard 40 mmol/L lactate-buffered peritoneal dialysis solution (Lac) over a 12-month treatment period. SETTING: 17 European nephrology centers. PATIENTS: 106 (70 Bic/Lac and 36 Lac) well-dialyzed continuous ambulatory peritoneal dialysis (CAPD) patients. INTERVENTIONS: Dialysis adequacy and peritoneal equilibration test (PET, week -4, months 3, 6, and 12); 24-hour ultrafiltration (week -4, months 1, 3, and 6); blood biochemistry (week -2, day 0, months 1, 2, 3,6, 9, and 12); and a product acceptability questionnaire (months 1 and 6). RESULTS: 88 patients completed the first 6 months, and 44 the full year. The solutions were shown to be therapeutically equivalent with respect to plasma bicarbonate and peritoneal urea and creatinine clearances. Ultrafiltration in the Bic/Lac group increased significantly from baseline by about 150 mL/day for the whole of the 6-month treatment period (p < 0.05). The biochemistry profile, adverse events, and physical examination (except body weight where there was a statistically significant increase in the Bic/Lac group) results did not differ significantly between the two groups. Reduced pain/discomfort on infusion or an increased sense of well-being was reported by 41% of patients on the Bic/Lac fluid. CONCLUSIONS: The Bic/Lac solution is safe and effective in correcting uremic acidosis, provides relief of inflow pain/discomfort, and improves ultrafiltration and body weight. PMID- 11117244 TI - Relationship between dialysis adequacy and quality of life in long-term peritoneal dialysis patients. AB - OBJECTIVE: The purpose of this study was to compare quality of life (QOL) between peritoneal dialysis (PD) patients with adequate and inadequate total solute clearance (TSC). We also tried to determine the relationship between QOL and TSC. DESIGN: A cross-sectional study design was used in which QOL was evaluated and compared between PD patients with adequate and inadequate TSC. SETTING: The PD unit of a university teaching hospital. PATIENTS: Sixty-seven patients were recruited, 38 on continuous ambulatory PD and 29 on continuous cyclerassisted PD. METHODS: Patients were divided into adequate and inadequate groups, based on the results of either total urea clearance (Kt/Vurea) or total creatinine clearance (weekly CCr). The demographic data, dialysis variables, and clinical parameters of these patients were all collected. QOL was evaluated using the SF-36 questionnaire, which contains eight domains and is a comprehensive and validated instrument for QOL evaluation. QOL of patients in adequate and inadequate groups was compared. The relationship between QOL and TSC was also examined. RESULTS: Among patients grouped by Kt/Vurea, patients in the adequate group had significantly higher scores in two domains of the SF-36, that is, physical and emotional role functioning, than did those in the inadequate group. The total SF 36 scores were positively correlated with Kt/Vurea when all patients were pooled together. However, among patients grouped by weekly CCr, there was no significant difference in any of the eight domains of the SF-36 between patients in the adequate and inadequate groups. No correlation was found between the total SF-36 scores and weekly CCr. CONCLUSION: Our study had two important findings: First, PD patients with adequate total solute clearance, based on Kt/Vurea and not on weekly CCr, had a better QOL. Second, Kt/Vurea is better correlated with QOL than weekly CCr. These findings suggest that Kt/Vurea is a better parameter for the clinical evaluation of total solute clearance from the viewpoint of QOL. PMID- 11117243 TI - Production and regulation of matrix metalloproteinases and their inhibitors by human peritoneal mesothelial cells. AB - OBJECTIVE: Human peritoneal mesothelial cells (HPMC) are likely to be involved in maintenance of the peritoneal membrane. We determined whether these cells were able to synthesize the matrix degrading enzymes, matrix metalloproteinases (MMPs), likely to be responsible for the breakdown of this membrane, and whether this secretion could be modulated by cytokines involved in the inflammatory response. DESIGN: MMP activity in conditioned medium of growth-arrested HPMC was measured by zymography. Cultures were incubated in the presence and absence of the cytokines transforming growth factor-beta (TGFbeta) and interleukin (IL) 1beta in order to determine the effects of these cytokines on this process. The mRNA for these MMPs, together with that of their specific inhibitors, tissue inhibitors of metalloproteinases (TIMPs), was also examined by reverse transcriptase polymerase chain reaction RESULTS: HPMC were shown to constitutively secrete the metalloproteinases MMP-2 and MMP-3 in vitro. In response to the proinflammatory cytokine IL-1beta , the protein and mRNA for MMP 9 was induced, while secretion of MMP-2 was unaltered. Similarly, the mRNA for MMP-3 was also increased relative to actin following the addition of IL-1beta. TGFbeta was shown to slightly induce the secretion of MMP-2 together with the mRNA for TIMP I, TIMP II, and, to a greater extent, TIMP III. Used peritoneal dialysate was also shown to induce MMP-9 secretion, and this effect was blocked by the co-incubation of IL-1 receptor antagonist. The secretion of enzyme activity was shown to be from the apical surface of the cells. CONCLUSION: HPMC have the ability to control the accumulation of extracellular matrix by secreting the matrix degrading molecules MMP-2, MMP-3, and MMP-9. In addition, the secretion of these enzymes, together with that of their inhibitors (TIMPs) is regulated by the cytokines IL-1beta and TGFbeta. This process is likely to be important in both the normal maintenance of the integrity of the peritoneal membrane and in the changes that occur following prolonged peritoneal dialysis. PMID- 11117245 TI - Peritoneal transport characteristics, comorbid diseases and survival in CAPD patients. AB - OBJECTIVE: To evaluate the influence of initial peritoneal transport rate, serum albumin concentration, and comorbid diseases on continuous ambulatory peritoneal dialysis (CAPD) patient survival. DESIGN: A prospective single-center study with a long-term follow-up. PATIENTS: A total of 213 consecutive CAPD patients, who underwent a peritoneal equilibration test (PET) at a mean of 7 days (range 3 - 30 days) after beginning CAPD, were included in this study. One hundred twenty patients were male, 116 patients had comorbid diseases, and mean age was 49.5 years (range 18 - 76 years). METHODS: A modified PET was performed using 4.25% glucose dialysis solution. Based on the dialysate-to-plasma creatinine concentration ratio at 4 hours' dwell (D4/P4 Cr, 0.62 +/- 0.14), patients were divided into high (H), high-average (HA), low-average (LA), or low (L) transporters. RESULTS: Of 213 patients, 16.9% were classified as H transporters, 30.5% as HA, 36.6% as LA, and 16.0% as L transporters. The H transporter group had a higher proportion of men, higher proportion of patients with comorbid diseases, lower initial serum albumin concentration, lower D4/D0 glucose, and lower drained volume. The initial D4/P4 Cr correlated with initial serum albumin (r= -0.35, p < 0.001). The patients with comorbid diseases had lower initial serum albumin and higher initial D4/P4 Cr. On Kaplan-Meier analysis, 2-year patient survival of group H was significantly lower compared to the other groups combined (57.1% vs 79.5%, p = 0.009). On Cox proportional hazards analysis, age, comorbid diseases, initial serum albumin concentration, and initial D4/P4 Cr were found to be independent risk factors for mortality. However, in the patients without comorbid diseases, patient survival was not different between group H and the other transport groups combined (p > 0.05), and only age was found to be an independent risk factor for mortality. CONCLUSION: These data suggest that a high peritoneal transport rate at initial PET is associated with high mortality, and that this is in part due to an increased prevalence of comorbid disease in H transporters. These H transporters with comorbid diseases represent a subset of patients with an especially poor prognosis. In patients without comorbid diseases, high transport status or low serum albumin concentration was not an independent risk factor for mortality. PMID- 11117246 TI - What is the optimal frequency of cycling in automated peritoneal dialysis? AB - OBJECTIVE: The recent increase in the use of automated peritoneal dialysis (APD) has led to concerns about the adequacy of clearances delivered by this modality. Few clinical studies looking at the effects of varying the individual components of the APD prescription on delivered clearance have been done, and most published data are derived from computer modeling. Most controversial is the optimal frequency of exchanges per APD session. Many centers prescribe 4 to 6 cycles per night but it is unclear if this is optimal. The purpose of this study was to address at what point the beneficial effect of more frequent cycles is outweighed by the concomitant increase in the proportion of the total cycling time spent draining and filling. METHODS: A comparison was made between the urea and creatinine clearances (CCrs) achieved by 4 different APD prescriptions, used for 7 days each, in 18 patients. The prescriptions were for 9 hours each and were all based on 2-L dwell volumes, but differed in the frequency of exchanges. They were 5 x 2 L, 7 x 2 L, and 9 x 2 L, as well as a 50% tidal peritoneal dialysis (TPD) prescription using 14 L. Ultrafiltration, dwell time, glucose absorption, sodium and potassium removal, protein excretion, and relative cost were also compared. Clearances due to day dwells and residual renal function were not included in the calculation. RESULTS: Mean urea clearances were 7.5, 8.6, 9.1, and 8.3 L/night for the four prescriptions respectively. Urea clearance with 9 x 2 L was significantly greater than with the other three prescriptions (p < 0 0.05). Urea clearance with 7 x 2 L and TPD were superior to 5 x 2 L (p < 0.05). Mean CCr was 5.1, 6.1, 6.4, and 5.6 L/night, respectively. Compared to 5 x 2-L, the 7 x 2-L, 9 x 2-L, and TPD prescriptions achieved greater CCr (p < 0.05). Taking both urea and CCr into account, 9 x 2 L was the optimal prescription in 12 of the 18 patients. Ultrafiltration and sodium and potassium removals were all significantly greater with the higher frequency prescriptions. CONCLUSION: The 5 x 2-L prescription significantly underutilizes the potential of APD to deliver high clearances, and 7 x 2 L is a consistently superior prescription if 2-L dwells are being used. Although more costly, 9 x 2 L should be considered if higher clearances are required. PMID- 11117247 TI - Peritoneal transport characteristics with glycerol-based dialysate in peritoneal dialysis. AB - BACKGROUND: Glycerol is a low molecular weight solute (MW 92 D) that can be used as an osmotic agent in continuous ambulatory peritoneal dialysis (CAPD). Due to its low molecular weight, the osmotic gradient disappears rapidly. Despite the higher osmolality at the beginning of a dwell, ultrafiltration has been found to be lower for glycerol compared to glucose (MW 180 D) when equimolar concentrations are used. Previous studies have shown glycerol to be safe for long term use, but some discrepancies have been reported in small solute transport and protein loss. OBJECTIVE: To assess permeability characteristics for a 1.4% glycerol dialysis solution compared to 1.36% glucose. DESIGN: Two standardized peritoneal permeability analyses (SPA), one using 1.4% glycerol and the other using 1.36% glucose, in random order, were performed within a span of 2 weeks in 10 stable CAPD patients. The length of the study dwell was 4 hours. Fluid kinetics and solute transport were calculated and signs of cell damage were compared for the two solutions. SETTING: Peritoneal dialysis unit in the Academic Medical Center, Amsterdam. RESULTS: Median values for the 1.4% glycerol SPA were as follows: net ultrafiltration 251 mL, which was higher than that for 1.36% glucose (12 mL, p < 0.01); transcapillary ultrafiltration rate 2.12 mL/min, which was higher than that for glucose (1.52 mL/min, p = 0.01); and effective lymphatic absorption rate 1.01 mL/min, which was not different from the glucose-based solution. Calculation of peritoneal reflection coefficients for glycerol and glucose showed lower values for glycerol compared to glucose (0.03 vs 0.04, calculated with both the convection and the diffusion models). A marked dip in dialysate-to-plasma ratio for sodium was seen in the 1.4% glycerol exchange, suggesting uncoupled water transport through water channels. Mass transfer area coefficients for urea, creatinine, and urate were similar for both solutions. Also, clearances of the macromolecules beta2-microglobulin, albumin, IgG, and alpha2-macroglobulin were not different for the two osmotic agents. The median absorption was higher for glycerol, 71% compared to 49% for glucose (p < 0.01), as could be expected from the lower molecular weight. The use of a 1.4% glycerol solution during a 4-hour dwell caused a small but significant median rise in plasma glycerol, from 0.22 mmol/L to 0.45 mmol/L (p = 0.02). Dialysate cancer antigen 125 and lactate dehydrogenase (LDH) concentrations during the dwell were not different for both solutions. CONCLUSIONS: These findings show that glycerol is an effective osmotic agent that can replace glucose in short dwells and show no acute mesothelial damage. The higher net ultrafiltration obtained with 1.4% glycerol can be explained by the higher initial net osmotic pressure gradient. This was seen especially in the first hour of the dwell. Thereafter, the osmotic gradient diminished as a result of absorption. The dip in dialysate-to-plasma ratio for sodium seen in the glycerol dwell can also be explained by this high initial osmotic pressure gradient, implying that the effect of glycerol as an osmotic agent is more dependent on intact water channels than is glucose. PMID- 11117248 TI - Application of mupirocin cream at the catheter exit site reduces exit-site infections and peritonitis in peritoneal dialysis patients. PMID- 11117249 TI - The effect of CAPD on the lipid profile of pediatric patients. PMID- 11117250 TI - Effect of increasing dialysate flow rates on adequacy of peritoneal dialysis. PMID- 11117251 TI - Lean body mass estimation by densitometry and creatinine kinetics in chronic peritoneal dialysis patients. PMID- 11117252 TI - Hepatitis E virus markers in a peritoneal dialysis population. PMID- 11117253 TI - Total lymphocyte count during the course of CAPD treatment. PMID- 11117255 TI - Literature. September-October 2000. PMID- 11117254 TI - A case of eosinophilic peritonitis treated with oral corticosteroids. PMID- 11117256 TI - F-box proteins and protein degradation: an emerging theme in cellular regulation. AB - Selective protein degradation by the ubiquitin-proteosome pathway has recently emerged as a powerful regulatory mechanism in a wide variety of cellular processes. Ubiquitin conjugation requires the sequential activity of three enzymes or protein complexes called the ubiquitin-activating enzyme (E1), the ubiquitin-conjugating enzyme (E2), and the ubiquitin-protein ligase (E3). In most eukaryotes, there are a small number of similar E1 isoforms without apparent functional specificity. The specific selection of target proteins is accomplished by the E2 and E3 proteins. One of the best-characterized families of E3s are the SCF complexes. The SCF is composed of a cullin (Cdc53), SKP1, RBX1 and one member of a large family of proteins called F-box proteins. The function of the F-box protein is to interact with target proteins. In some cases, the stability of the F-box protein may regulate activity of the SCF complex. In addition, post translational modification of the cullin subunit by the ubiquitin-like protein RUB/NEDD8 appears to regulate SCF function. In plants, the SCF has so far been implicated in floral development, circadian clock, and response to the plant growth regulators auxin and jasmonic acid. PMID- 11117257 TI - Plant lipoxygenase 2 is a translation initiation factor-4E-binding protein. AB - The eukaryotic initiation factor 4E (eIF4E) emerged recently as a target for different types of regulation affecting translation. In animal and yeast cells, eIF4E-binding proteins modulate the availability of eIF4E. A search for plant eIF4E-binding proteins from Arcabictopsis thaliana using the yeast genetic interaction system identified a clone encoding a lipoxygenase type 2 (AtLOX2). In vitro and in vivo biochemical assays confirm an interaction between AtLOX2 and plant eIF4E(iso) factor. A two-hybrid assay revealed that AtLOX2 is also able to interact with both wheat initiation factors 4E and 4E(iso). Deletion analysis maps the region of AtLOX2 involved in interaction with AteIF(iso)4E between amino acids 175 and 232. A sequence related to the conserved motif present in several eIF4E-binding proteins was found in this region. Furthermore, the wheat p86 subunit, a component of the plant translation eIF(iso)4F complex, was found to interfere with the AteIF(iso)4E-AtLOX2 interaction suggesting that p86 and AtLOX2 compete for the same site on eIF(iso)4E. These results may reflect a link between eIF4Es factors mediating translational control with LOX2 activity, which is probably conserved throughout the plant kingdom. PMID- 11117258 TI - Regulation of NADPH-cytochrome P450 reductase expressed during Douglas-fir germination and seedling development. AB - NADH-cytochrome P450 is a key enzyme that transfers electrons from NADPH to the cytochrome P450 family of enzymes. To begin to determine the regulation of CPR gene expression and enzyme activity in Douglas-fir a full-length cDNA was isolated from a seedling lambda ZAP cDNA library and the ORF was used to develop a synthetic CPR-peptide-based antiserum. Northern blot analysis indicated CPR expression was regulated both developmentally prior to seed maturation and during germination, and differentially in the cotyledons, radicle and megagametophyte of seed and seedling tissues. The CPR-peptide antiserum detected a single CPR in seed and seedling microsomes analyzed by western blot of two-dimensional SDS polyacrylamide gels. In microsomal extracts from whole seeds and seedlings, the amount of CPR protein remained constant while NADPH:cytochrome c reductase activity increased during stratification, germination and early seedling development. In contrast to cotyledons and megagametophyte, the level of CPR protein detected in radicles was higher than expected when compared to the amount of CPR transcript. PMID- 11117259 TI - Splicing of arabidopsis tRNA(Met) precursors in tobacco cell and wheat germ extracts. AB - Intron-containing tRNA genes are exceptional within nuclear plant genomes. It appears that merely two tRNA gene families coding for tRNA(GpsiA(Tyr)) and elongator tRNA(CmAU(Met)) contain intervening sequences. We have previously investigated the features required by wheat germ splicing endonuclease for efficient and accurate intron excision from Arabidopsis pre-tRNA(Tyr). Here we have studied the expression of an Arabidopsis elongator tRNA(Met) gene in two plant extracts of different origin. This gene was first transcribed either in HeLa or in tobacco cell nuclear extract and splicing of intron-containing tRNA(Met) precursors was then examined in wheat germ S23 extract and in the tobacco system. The results show that conversion of pre-tRNA(Met) to mature tRNA proceeds very efficiently in both plant extracts. In order to elucidate the potential role of specific nucleotides at the 3' and 5' splice sites and of a structured intron for pre-tRNA(Met) splicing in either extract, we have performed a systematic survey by mutational analyses. The results show that cytidine residues at intron-exon boundaries impair pre-tRNA(Met) splicing and that a highly structured intron is indispensable for pre-tRNA(Met) splicing. tRNA precursors with an extended anticodon stem of three to four base pairs are readily accepted as substrates by wheat and tobacco splicing endonuclease, whereas pre-tRNA molecules that can form an extended anticodon stem of only two putative base pairs are not spliced at all. An amber suppressor, generated from the intron-containing elongator tRNA(Met) gene, is efficiently processed and spliced in both plant extracts. PMID- 11117260 TI - Functional analysis of a RPD3 histone deacetylase homologue in Arabidopsis thaliana. AB - Histone acetylation is modulated through the action of histone acetyltransferase and deacetylase, which play key roles in the regulation of eukaryotic gene expression. We have screened the expressed sequence tag database with the yeast histone deacetylase RPD3 sequence and identified two Arabidopsis homologues, AtRPD3A and AtRPD3B. The deduced amino acid sequences of AtRPD3A and AtRPD3B show high overall homology (55% identity) to each other. AtRPD3A encodes a putative protein of 502 amino acids with 49% identity to the yeast RPD3. AtRPD3B encodes a putative protein of 471 amino acids and shares 55% amino acid identity with the yeast RPD3. Northern analysis indicated that AtRPD3A was highly expressed in the leaves, stems, flowers and young siliques of Arabidopsis plants, whereas the AtRPD3B transcript was not detected in these organs. An AtRPD3A fusion protein repressed transcription when directed to a promoter driving a reporter gene, indicating a role for AtRPD3A protein in gene repression. Arabidopsis plants were transformed with a gene construct comprising a truncated AtRPD3A cDNA in the antisense orientation driven by a strong constitutive promoter, -394tCUP. Antisense expression of AtRPD3A resulted in decreased endogenous AtRPD3A transcript and delayed flowering in transgenic Arabidopsis plants, suggesting that the transition from the vegetative to reproductive phase of development could be affected by histone acetylation. Our study demonstrates the important role of histone deacetylases in plant growth and development. PMID- 11117261 TI - Transient expression of a pea MAP kinase gene induced by gibberellic acid and 6 benzyladenine in unpollinated pea ovaries. AB - PsMAPK3, a new MAP kinase cDNA, was cloned from ovaries of Pisum sativum L. Expression of PsMAPK3 is at low basal levels in unpollinated ovaries but it is rapidly induced by gibberellic acid (peak at 30 min) and 6-benzyladenine (peak at 45 min). Both treatments promoted the development of a parthenocarpic fruit. In situ hybridization localized PsMAPK3 mRNA in ovules. The transcript was additionally detected in the mesocarp when it is expanding in response to the treatments. These observations suggest that gibberellins and cytokinins regulate PsMAPK3 mRNA levels in pea ovary shortly after fruit set is induced. PMID- 11117263 TI - Two dioxygenase genes, Ids3 and Ids2, from Hordeum vulgare are involved in the biosynthesis of mugineic acid family phytosiderophores. AB - A cDNA clone, Ids3 (iron deficiency-specific clone 3), was isolated from an Fe deficient-root cDNA library of Hordeum vulgare. Ids3 encodes a protein of 339 amino acids with a calculated molecular mass of 37.7 kDa, and its amino acid sequence shows a high degree of similarity with those of plant and fungal 2 oxoglutarate-dependent dioxygenases. One aspartate and two histidine residues for ferrous Fe binding (Asp-211, His-209, His-265) and arginine and serine residues for 2-oxoglutarate binding (Arg-275, Ser-277) are conserved in the predicted amino acid sequence of Ids3. Ids3 expression was rapidly induced by Fe deficiency, and was suppressed by re-supply of Fe. Among eight graminaceous species tested, Ids3 expression was observed only in Fe-deficient roots of H. vulgare and Secale cereale. which not only secrete 2'-deoxymugineic acid (DMA), but also mugineic acid (MA) and 3-epihydroxymugineic acid (epiHMA, H. vulgare), and 3-hydroxymugineic acid (HMA, S. cereale). The Ids3 gene is encoded on the long arm of chromosome 4H of H. vulgare, which also carries the hydroxylase gene that converts DMA to MA. Moreover, the Ids2 gene, which is the plant dioxygenase with the highest homology to Ids3, is encoded on the long arm of chromosome 7H of H. vulgate, which carries the hydroxylase gene that converts MA to epiHMA. The observed expression patterns of the Ids3 and Ids2 genes strongly suggest that IDS3 is an enzyme that hydroxylates the C-2' positions of DMA and epiHDMA, while IDS2 hydroxylates the C-3 positions of MA and DMA. PMID- 11117262 TI - Characterization and cDNA cloning of two glycine- and histidine-rich antimicrobial peptides from the roots of shepherd's purse, Capsella bursa pastoris. AB - Two novel antimicrobial peptides were isolated and characterized from the roots of shepherd's purse, Capsella bursa-pastoris. These antimicrobial peptides, named shepherin I and shepherin II, consist of 28 and 38 amino acids, respectively, and are glycine- and histidine-rich peptides. Shepherin I and shepherin II have 67.9% and 65.8% (mol/mol) glycine, respectively, and 28.6% and 21.1% (mol/mol) histidine, respectively. Both shepherins have a Gly-Gly-His motif. These antimicrobial peptides exhibit antimicrobial activity against Gram-negative bacteria and fungi. Circular dichroism spectra of shepherin I and shepherin II showed that shepherin I and shepherin II in 50% trifluoroethanol have 66.7% and 75% random coils, respectively, without any alpha-helices. cDNA sequence analysis revealed that shepherin I and shepherin II are produced from a single polypeptide, designated shep-GRP, consisting of 120 amino acids; shep-GRP has five distinct domains, an amino-terminal putative signal peptide, a shepherin I, a linker dipeptide, a shepherin II and a carboxy-terminal peptide. Southern blot analysis indicates that the gene encoding shepherins belongs to a low-complexity gene family. Northern blot analysis revealed that transcripts of shep-GRP are present in roots but not in leaves and stems. PMID- 11117264 TI - Defence gene expression in soybean is linked to the status of the cell death program. AB - Soybean cell cultures (cv. Williams 82) respond to Pseudomonas syringae bacteria expressing the avirulence gene AvrA with a hypersensitive reaction, a programmed cell death (PCD) of plant cells to pathogen attack. This PCD is under control of salicylic acid (SA) via an unknown mechanism. In the presence of low concentrations of SA, the cells undergo a very rapid cell death, which needs only half of the time required for the normal hypersensitive reaction (HR). Northern blot studies for defence-related genes show that the expression of many of these genes is tightly linked to the status of the cell death program rather than to pathogen-derived elicitors. Thus the expression is much faster in the SA accelerated PCD than in the normal hypersensitive reaction. In contrast, other pathogen-responsive genes are induced independently of the speed of PCD, indicating a divergent signalling mechanism. The production of reactive oxygen species during the oxidative burst of bacteria-inoculated soybean cells is slightly enhanced in the presence of SA but occurs at the same time as in untreated cells, suggesting that SA exhibits the control of the PCD downstream of the oxidative burst. Consistent with these findings a HR-specific marker gene is neither directly induced by H2O2 or SA. However, this gene shows a high expression in the regular HR and is induced much faster in the SA-accelerated PCD. PMID- 11117265 TI - A non-toxic pokeweed antiviral protein mutant inhibits pathogen infection via a novel salicylic acid-independent pathway. AB - Pokeweed antiviral protein (PAP), a ribosome-inactivating protein isolated from Phytolacca americana, is characterized by its ability to depurinate the sarcin/ricin (S/R) loop of the large rRNA of prokaryotic and eukaryotic ribosomes. In this study, we present evidence that PAP is associated with ribosomes and depurinates tobacco ribosomes in vivo by removing more than one adenine and a guanine. A mutant of pokeweed antiviral protein, PAPn, which has a single amino acid substitution (G75D), did not bind ribosomes efficiently, indicating that Gly-75 in the N-terminal domain is critical for the binding of PAP to ribosomes. PAPn did not depurinate ribosomes and was non-toxic when expressed in transgenic tobacco plants. Unlike wild-type PAP and a C-terminal deletion mutant, transgenic plants expressing PAPn did not have elevated levels of acidic pathogenesis-related (PR) proteins. PAPn, like other forms of PAP, did not trigger production of salicylic acid (SA) in transgenic plants. Expression of the basic PR proteins, the wound-inducible protein kinase and protease inhibitor II, was induced in PAPn-expressing transgenic plants and these plants were resistant to viral and fungal infection. These results demonstrate that PAPn activates a particular SA-independent, stress-associated signal transduction pathway and confers pathogen resistance in the absence of ribosome binding, rRNA depurination and acidic PR protein production. PMID- 11117266 TI - Arabidopsis ATP A2 peroxidase. Expression and high-resolution structure of a plant peroxidase with implications for lignification. AB - Lignins are phenolic biopolymers synthesized by terrestrial, vascular plants for mechanical support and in response to pathogen attack. Peroxidases have been proposed to catalyse the dehydrogenative polymerization of monolignols into lignins, although no specific isoenzyme has been shown to be involved in lignin biosynthesis. Recently we isolated an extracellular anionic peroxidase, ATP A2, from rapidly lignifying Arabidopsis cell suspension culture and cloned its cDNA. Here we show that the Atp A2 promoter directs GUS reporter gene expression in lignified tissues of transgenic plants. Moreover, an Arabidopsis mutant with increased lignin levels compared to wild type shows increased levels of ATP A2 mRNA and of a mRNA encoding an enzyme upstream in the lignin biosynthetic pathway. The substrate specificity of ATP A2 was analysed by X-ray crystallography and docking of lignin precursors. The structure of ATP A2 was solved to 1.45 A resolution at 100 K. Docking of p-coumaryl, coniferyl and sinapyl alcohol in the substrate binding site of ATP A2 were analysed on the basis of the crystal structure of a horseradish peroxidase C-CN-ferulic acid complex. The analysis indicates that the precursors p-coumaryl and coniferyl alcohols are preferred by ATP A2, while the oxidation of sinapyl alcohol will be sterically hindered in ATP A2 as well as in all other plant peroxidases due to an overlap with the conserved Pro-139. We suggest ATP A2 is involved in a complex regulation of the covalent cross-linking in the plant cell wall. PMID- 11117267 TI - Immune mechanisms in interstitial lung diseases. PMID- 11117268 TI - Eosinophil cationic protein and eosinophil protein X in the nasal lavage of children during the first 4 weeks of life. SPACE Collaborative Study Team. AB - Eosinophil cationic protein (ECP) and eosinophil protein X (EPX) are well established as markers of eosinophil activation. We analyzed ECP and EPX concentrations in nasal lavage fluids (NALF) of 378 neonates during their first 4 weeks of life. Inclusion criteria were a positive history of parental allergy and a positive skin prick test or specific IgE (RAST class > or = 2) against at least one out of a panel of common aeroallergens in one or both parents. Twenty-four infants with no history of parental allergy were used as controls. A volume of 2 ml of 0.9% saline was instilled into each nostril and immediately recovered by a suction device. ECP and EPX were analyzed by radioimmunoassay. In 65 samples of three consecutive lavages, EPX was detected in nine samples (13.8%) in the control group, whereas it was detected in 197/360 samples (54.7%) in the study population. The corresponding figures for ECP were 17/65 (26.2%) in the control group and 173/365 (47.4%) in the study group. Both proteins showed a skewed distribution (median/5-95th percentiles for ECP: 0 microg/l [0-69.4] and EPX: 6.6 microg/l [0-73.2]). The differences between the control group and the study group were statistically significant, regardless of the allergic disease of the parents. In children of allergic parents, activation proteins of the eosinophil granulocyte are released on the nasal mucosal surface in about 50% of the studied population at the age of 4 weeks. This early onset of eosinophil activation in the nasal respiratory epithelium may reflect a genetic predisposition to allergy or early exposure to allergens. PMID- 11117269 TI - The effect of Chinese herbal medicine, xiao-qing-long tang (XQLT), on allergen induced bronchial inflammation in mite-sensitized mice. AB - BACKGROUND: There are detailed descriptions of the clinical experiences and prescriptions of asthma in traditional Chinese medicine. Xiao-qing-long tang (XQLT), or sho-seiryo-to by its Japanese name, is one of the Chinese herbal medicines used to treat bronchial asthma and allergic rhinitis for centuries. However, the therapeutic mechanisms of this medication are still far from clear. In this study, a house-dust-mite (Dermatophagoides pteronyssinus [Der p]) sensitized murine model of asthma was used to evaluate the immunomodulatory effect of XQLT on the allergen-induced airway inflammation in asthma. METHODS: Three different protocols were designed to evaluate the treatment and/ or long term prophylactic effect of XQLT in Der p-sensitized mice. XQLT extracts (1 gm/kg) were administered to sensitized mice 1 h before allergen challenge (AC) (group A), 24 h after AC (group B), and every other day six times before AC (group C), respectively. Cellular infiltration and T-cell subsets in the bronchoalveolar lavage fluid (BALF) of allergen-challenged mice were analyzed. Intrapulmonary lymphocytes were also isolated to evaluate their response to allergen stimulation. RESULTS: When XQLT was administered to the sensitized mice before AC (groups A and C), it suppressed airway inflammation by decreasing the number of total cells and eosinophil infiltration in the BALF, and downregulated the allergen- or mitogen-induced intrapulmonary lymphocyte response of sensitized mice as compared to those of controls. This immunomodulatory effect of XQLT may be exerted through the regulation of T-cell subsets by elevation or activation of the CD8+ and double-negative T-cell population in the lung. However, the administration of XQLT to sensitized mice 24 h after AC (group B) did not have the same inhibitory effect on the airway inflammation as XQLT given before AC. CONCLUSIONS: The administration of XQLT before AC has the immunomodulatory effect of reducing bronchial inflammation in the allergen-sensitized mice. PMID- 11117270 TI - Cytokine production from peripheral whole blood in atopic and nonatopic asthmatics: relationship with blood and sputum eosinophilia and serum IgE levels. AB - BACKGROUND: The cytokine network is thought to be essential in orchestrating airway inflammation in asthma. Although evidence has accumulated to suggest that atopic asthma is a Th2 disease, much less is known about nonatopic asthma. METHODS: We have compared the production of IL-4, IL-6, IFN-gamma, and TNF-alpha from peripheral blood leukocytes between atopic (n=21) and nonatopic (n=22) asthmatics and healthy nonatopic subjects (n=20). Peripheral blood was incubated for 24 h either without stimulus or with LPS or PHA. Cytokines were measured by the immunotrapping technique (Dynamic Immunoassay). RESULTS: When compared to healthy nonatopic subjects, both atopic and nonatopic asthmatics showed increased blood and sputum eosinophilia associated with raised total serum IgE levels. Similarly, both asthma groups displayed spontaneous, endotoxin-induced overproduction of IL-6. Enhanced spontaneous, endotoxin-induced release of IL-4 combined with reduced spontaneous IFN-gamma production was seen only in atopic asthma. In this group of patients, the production of IL-4 was related to the extent of blood and sputum eosinophilia. In nonatopic asthmatics, serum levels of IgE were inversely related to the production of IFN-gamma. CONCLUSIONS: Both atopic and intrinsic asthma display raised blood and airway eosinophilia, raised total serum IgE, and overproduction of IL-6 from peripheral blood. Atopic asthma is also characterized by impaired spontaneous release of IFN-gamma and increased production of IL-4 that correlates with the magnitude of eosinophilic inflammation. PMID- 11117271 TI - Preseasonal local allergoid immunotherapy to grass pollen in children: a double blind, placebo-controlled, randomized trial. AB - BACKGROUND: We assessed the efficacy of preseasonal local allergoid immunotherapy in a group of children with asthma and/or rhinitis and/or rhinoconjunctivitis due to grass pollen. METHODS: We randomly assigned 24 children allergic to grass pollen to receive local allergoid immunotherapy for 3 months before the pollen season and 24 such patients to receive identically appearing placebo. The immunotherapy consisted of tablets of monomeric allergoid grass pollen allergens held in the mouth until they dissolved and then swallowed. The study was double blind. Symptoms and medications were scored on diary cards during the pollen season. Nasal eosinophil cationic protein levels were measured by the monoclonal antibodies EG1 and EG2 outside the pollen season and at low and at high pollen concentration during the pollen season. RESULTS: The active-treatment group had a statistically significant reduction of total symptoms (P<0.05), especially bronchial symptoms (P<0.05), in comparison with the placebo group. Immunotherapy was well tolerated and compliance was good. Nasal levels of EG2 and EG1 increased significantly during the pollen season, but there was no difference between groups. EG2/EG1 increased significantly only in the placebo group during natural allergen exposure (P<0.01). CONCLUSIONS: Our results suggest that this immunotherapy is effective for the treatment of asthma due to grass pollen in children. PMID- 11117273 TI - Involvement of inferior turbinate mucosa in chronic sinusitis--localization of T cell subset. AB - BACKGROUND: In chronic sinusitis (CS), different subsets of leukocytes are involved in development of persistent inflammation of the nasal mucosa. The localization and differentiation of these infiltrating lymphocytes may help us to understand the inflammatory interactions in the epithelium, lamina propria, and seromucous glands of the nasal mucosa in CS. METHODS: We examined frozen sections of inferior turbinates from 14 patients with nonallergic CS and 10 normal nonallergic controls. We used the avidin-biotin-peroxidase (ABC) technique with monoclonal antibodies against CD3 (total T cells), CD4 (T-helper/inducer cells), CD8 (T-suppressor/cytotoxic cells), CD22 (B cells), CD56 (natural killer cells), elastase (neutrophil granulocytes), eosinophil cationic protein (eosinophil granulocytes), and CD68 (macrophages). RESULTS: We found significant increases (P < 0.05) of CD3, CD4, and CD8 T cells and B cells in the nasal mucosa of patients with CS. The number of CD68 cells and eosinophils showed no significant rise. CONCLUSIONS: The different types of leukocytes play a key role in the defense of the respiratory tract. The analysis of the distribution of cells in the epithelium, mucosa, and glands of the inferior turbinate confirmed that nonallergic CS is, in fact, chronic, bacterial rhinosinusitis involving the inferior turbinates, and that the pathomechanism is therefore different from that of nasal polyposis. PMID- 11117272 TI - A new method for collecting airborne allergens. AB - BACKGROUND: Measurement of airborne allergens has hitherto been done with the use of fixed-location pumps or personal air samplers. Our objective was to find out whether ionizers could be good tools for collecting airborne allergens. As a model we have used cat allergen (Fel d l). We have compared Fel d l levels collected by the ionizer at different time periods, as well as comparing Fel d l levels obtained with the ionizer with those of low- and high-volume pumps. METHODS: Dust samples from floors and air samples collected with ionizers and pumps, obtained in 31 homes with cat, 23 homes without cats, and 28 day-care centres, were analysed for cat allergen content (Fel d I) by ELISA. RESULTS: Fel d l was present in the reservoir in all homes with cats, ranging from 660 to 375,000 ng/g (GM 75,000) and in the air collected by the ionizer from 2.0 to 204 ng/24 h (GM 19.3). The allergen in homes without cat varied from < 55 to 1,800 ng/g (GM 166). Corresponding levels in air were found in two of these homes (2.3 and 7.3 ng/24 h). There was a correlation between the number of cats and the amount of airborne cat allergen (r: 0.47; P < 0.05). The levels in day-care centres were < 55 to 3,070 ng/g in dust (GM 360) and < 1.1 to 7.9 ng/24 h in the air (GM 1.6). We obtained a moderately strong correlation between air and dust samples in homes with cats (rs: 0.64; P< 0.001) and in day-care centres (rs: 0.49; P<0.05). We found that a collection period of 24 h is preferable for the ionizer. The intrahome reliability coefficient was nearly two times higher for the ionizer (r: 0.69) than the pump (r: 0.39). CONCLUSIONS: The ionizer seems to be a good tool for monitoring the environment. It is easy to use and silent and does not disturb the airflow in the room. PMID- 11117274 TI - Prevalence of asthma and rhinitis in relation to long-term exposure to gaseous air pollutants. AB - The relationship between long-term exposure to air pollutants, especially with regard to photochemical air pollutants, and asthma prevalence in developed countries is controversial. The objective of this cross-sectional survey was to compare mean levels of the main gaseous air pollutants and prevalence rates of rhinitis, asthma, and asthmatic symptoms. It included 2,445 children from the 8th and 9th school grades who had been living for at least 3 years in an area where some communities undergo the heaviest photochemical exposure in France. Data on rhinitis, asthmatic symptoms, and asthma prevalence were gathered with the ISAAC paper and video questionnaires. The relation between level of air pollutants and asthma was assessed first by comparison of crude prevalence rates (chi-square test), and then by simple regression analysis and multiple logistic regression analysis. No consistent association between mean SO2 and NO2 levels, and prevalence of rhinitis, asthma, or asthmatic symptoms could be demonstrated. In contrast, there were statistically significant associations between prevalence of asthmatic symptoms and mean ozone O3) concentration. The interpretation of such findings is not straightforward, as these symptoms can be interpreted either as respiratory irritation due to exposure to nonspecific airway stimuli or as a true asthmatic state. Additional studies are required to clarify this important issue. In conclusion, this large cross-sectional epidemiologic survey performed in an area of high photochemical air pollution did demonstrate statistically significant associations between the prevalence of asthmatic symptoms and mean O3 concentration. PMID- 11117275 TI - Effects of human lung fibroblasts on eosinophil degranulation. AB - BACKGROUND: Although eosinophils and fibroblasts are thought to contribute to the mechanisms of chronic asthmatic inflammation, the interaction between eosinophils and fibroblasts has not been thoroughly clarified. We examined eosinophil cationic protein (ECP) release from human eosinophils cultured in the presence of human lung fibroblast HFL-1. METHODS: Eosinophils from healthy donors were cultured with or without C5a for 16 h in the presence of human fetal lung fibroblasts which had previously been incubated with or without TNF for 4 h. ECP in supernatants was measured by RIA. RESULTS: ECP release was potentiated only when both eosinophils and fibroblasts were activated by C5a and TNF, respectively, while it was not significantly potentiated when either eosinophils or fibroblasts were activated. Paraformaldehyde fixation of fibroblasts had some suppressive effect on ECP enhancement, and mAb against GM-CSF partly inhibited ECP enhancement. Coculture of eosinophils and fibroblasts with stimulus treatment resulted in the enhancement of both eosinophil adhesion and ECP release. The potentiation of ECP release was partially inhibited by anti-ICAM-1 mAb, anti-CD18 mAb, and anti-CD29 mAb, which caused partial and comparable inhibition of the enhancement of eosinophil adhesion. CONCLUSIONS: This study suggests that the activation of fibroblasts may have some role in the potentiation of ECP release from cocultured eosinophils, and that adhesion of eosinophils to fibroblasts may partly be involved in the mechanism of ECP potentiation. PMID- 11117276 TI - Expression of the American cockroach Per a 1 allergen in mammalian cells. AB - BACKGROUND: Cockroach allergens are one of the major etiologic risk factors for developing IgE-mediated allergic respiratory illness throughout the world. Per a 1 is a cross-reactive allergen of American and German cockroaches. This study aimed to investigate the expression of a recombinant American cockroach (Periplaneta americana) Per a 1, C42, allergen in mammalian COS-1 cells. METHODS: The COS-1 cells and Escherichia coli were used to express the P. americana C42 allergen. Recombinant proteins were purified with hydroxylapatite and DE52 chromatography. Biologic reactivities of recombinant proteins were examined by direct IgE binding and IgE inhibition studies with the enzyme-linked immunosorbent assay (ELISA). RESULTS: C42 was successfully expressed in the mammalian COS-1 cell as a 50-kDa secreted protein, and purified from the culture medium. The specific human IgE antibodies against recombinant C42 from either E. coli (C42-E. coli) or COS-1 (C42-COS-1) were compared by ELISA with 12 sera from Per a 1 and C42 skin-test-positive patients. All atopic sera contained specific IgE antibodies to C42 from either E. coli or COS-1. Moreover, recombinant C42-COS 1 bound higher levels of serum IgE than recombinant C42-E. coli among C42 sensitive atopic patients, and a statistically significant difference (P<0.01) was found between them. In addition, recombinant C42-COS-1 as an inhibitor revealed higher inhibition of IgE binding to natural Per a 1 than recombinant C42 E. coli. CONCLUSIONS: The biologically highly reactive recombinant C42 produced in the COS-1 cell provides an alternative expression system and will facilitate studies on the immune response of asthma patients to cockroach allergens. PMID- 11117277 TI - Reduction of exercise-induced asthma oxidative stress by lycopene, a natural antioxidant. AB - BACKGROUND: Lycopene has previously been shown to have high antioxidative activity. In view of the controversy regarding the beneficial effect of antioxidants on asthma, the acute effects of lycopene (LYC-O-MATO) on airway hyperreactivity were assessed in patients with exercise-induced asthma (EIA). METHODS: Twenty patients with EIA participated in our study to verify the antioxidative effects. The test was based on the following sequence: measurement of baseline pulmonary function, 7-min exercise session on a motorized treadmill, 8-min rest and again measurement of pulmonary function, 1-week, oral, randomly administered, double-blind supplementation of placebo or 30 mg/day of lycopene (LYC-O-MATO), measurement of pulmonary function at rest, 7-min exercise session, and 8-min rest and again measurement of pulmonary function. A 4-week washout interval was allowed between each protocol. RESULTS: All patients given placebo showed significant postexercise reduction of more than 15% in their forced expiratory volume in 1 s (FEV1). After receiving a daily dose of 30 mg of lycopene for 1 week, 11 (55%) patients were significantly protected against EIA. Serum analyses of the patients by high-pressure liquid chromatography detected in the lycopene-supplemented patients an elevated level of lycopene compared to the placebo group, with no change in retinol, tocopherols, or in the other carotenoids. CONCLUSIONS: Our results indicate that a daily dose of lycopene exerts a protective effect against EIA in some patients, most probably through an in vivo antioxidative effect. PMID- 11117279 TI - Is allergic asthma associated with delayed fetal maturation or the persistence of conserved fetal genes? PMID- 11117278 TI - Hypersensitivity pneumonitis caused by Fusarium napiforme in a home environment. AB - BACKGROUND: We report a case of hypersensitivity pneumonitis (HP) in a 17-year old male student caused by Fusarium napiforme found in his home environment. METHODS: The patient was diagnosed according to history, chest radiograph, spirometry, high-resolution chest CT, and transbronchial lung biopsy. To identify the causative agent, cultured aeromolds were collected by the open-plate method. From the main fungi cultured, fungal antigens were prepared, and immunoblot analysis with the patient's serum and each fungal antigen was performed. RESULTS: Five fungal species were isolated from the patient's home. Immunoblotting analysis with the patient's serum demonstrated more than 10 IgG-binding fractions to F. napiforme extract only, while little binding was noted with the other fungal antigens. CONCLUSIONS: We should be aware that HP may be caused by F. napiforme in the home environment. PMID- 11117280 TI - Intranasal beta-agonist in allergic rhinitis. PMID- 11117281 TI - Amiodarone-induced angioedema. PMID- 11117282 TI - Is alternative medicine acceptable in allergology? PMID- 11117283 TI - Erythema multiforme to vancomycin. PMID- 11117284 TI - Anisakis antigen interferes in IL-4 analysis. PMID- 11117285 TI - Increased END/EPX in ongoing asthma. PMID- 11117286 TI - Effects of provocholine and methacholine on airway responses. PMID- 11117287 TI - Biomechanical consequences of an isolated overload on the human vertebral body. AB - The biomechanical consequences of an isolated overload to the vertebral body may play a role in the etiology of vertebral fracture. In this context, we quantified residual strains and reductions in stiffness and ultimate load when vertebral bodies were loaded to various levels beyond the elastic regimen and related these properties to the externally applied strain and bone density. Twenty-three vertebral bodies (T11-L4, from 23 cadavers aged 20-90 years) were loaded once in compression to a randomized nominal strain level between 0.37 and 4.5%, unloaded, and then reloaded to 10% strain. Residual strains of up to 1.36% developed on unloading and depended on the applied strain (r2=0.85) but not on density (p = 0.25). Percentage reductions in stiffness and ultimate load of up to 83.7 and 52.5%, respectively, depended on both applied strain (r2 = 0.90 and r2 = 0.32, respectively) and density (r2 = 0.23 and r2 = 0.22, respectively). Development of residual strains is indicative of permanent deformations, whereas percentage reductions in stiffness are direct measures of effective mechanical damage. These results therefore demonstrate that substantial mechanical damage-which is not visible from radiographs-can develop in the vertebral body after isolated overloads, as well as subtle but significant permanent deformations. This behavior is similar to that observed previously for cylindrical cores of trabecular bone. Taken together, these findings indicate that the damage behavior of the lumbar and lower thoracic vertebral body is dominated by the trabecular bone and may be an important factor in the etiology of vertebral fracture. PMID- 11117288 TI - Efficacy of monitoring long-bone fracture healing by measurement of either bone stiffness or resonant frequency: numerical simulation. AB - Development of noninvasive mechanical tests to monitor fracture healing has been hindered because relationships between bone geometry, measurement conditions, and fracture callus strength are not well understood. Beam theory was used to analyze the effects of fracture length, fracture location, end conditions, and fracture callus stiffness on mechanical properties (resonant frequency, bending stiffness, and torsional stiffness) of healing bone. Actual bone mineral geometry from a human tibia, quantified every 1 mm, was used in the beam analysis. Geometry of the fracture callus segment was uniformly scaled from the values for intact bone. Experimental tests on multisegmented machined rods were used to verify analytical methods. Mechanical properties of the healing bone initially increased very rapidly to 30-70% of the stiffness of intact bone, depending on the configuration. The increases then tapered off dramatically. Lateral bending stiffness was sensitive to changes in callus properties for a larger portion of the healing process than was either torsional stiffness or resonant frequency. Because callus strength increases at half the rate of callus stiffness, measures of whole-bone mechanical properties can provide insight into changes in callus strength until a maximum of less than one-half the strength of intact bone is regained. The analytical method presented is proposed for clinical use to develop individualized models of bone, fracture, and fixation conditions to identify early stages of healing. Because increases in whole-bone mechanical properties are small in the later stages of fracture healing, however, such measures must be used prudently beyond the initial stages. PMID- 11117289 TI - Vascular proliferation and blood supply during distraction osteogenesis: a scanning electron microscopic observation. AB - This scanning electron microscopic study examined the spatial and temporal features of proliferating vessels of regenerating bone tissue and blood supply during distraction osteogenesis. A rat model of tibial lengthening was used with a protocol divided into a latency period of 7 days, a distraction period that lasted 14 days with a daily distraction rate of 0.5 mm in two steps, and a consolidation period of 21 days. Vascular casting was done on the hindlimbs before osteotomy and on postoperative days 7, 14, 21, 28, and 42. Scanning electron microscopic findings were correlated with radiological and histological observations. On days 7 and 14, the proliferation of periosteal vessels was pronounced and there was distinct subperiosteal bone formation on the osteotomized surfaces. On day 21, vascular branches from the medullary canal of the host bone formed a vascular network, which gave rise to multiple axial, straight vascular branches, running parallel to the direction of distraction, toward the interzone, in accordance with the progress of mineralization. On day 28, the periosteum provided vascularization to the peripheral side of the interzone whereas the center of the interzone was still relatively avascular. On day 42, the periosteal and medullary vascular channels were completely connected at the distraction site including the interzone, which was occupied by developing and mature bone trabeculae. These results suggest that vascular proliferation occurs actively during the latency and distraction periods and then gradually decreases over time. A close temporal and spatial relationship exists between formation of regenerated bone and vascular proliferation of the periosteum and medullary canal. PMID- 11117290 TI - Mechanobiology and joint conformity regulate endochondral ossification of sesamoids. AB - Sesamoid bones form by the endochondral ossification of sesamoid cartilages. This ossification process is thought to be similar to that responsible for the formation of secondary ossific nuclei in long-bone epiphyses. Sesamoids ossify much later in development than do epiphyses, however, and bone formation within sesamoids often begins by way of multiple ossific nuclei. Endochondral growth and ossification in the formation of secondary ossific nuclei have previously been correlated with distributions of the octahedral shear and hydrostatic stresses generated in vivo within cartilage anlagen. In this study, we used two dimensional finite element analysis to predict the distributions of octahedral shear and hydrostatic stresses in an idealized model of a sesamoid cartilage subjected to in vivo loading. We examined the influence of sesamoid joint conformity. The distribution of an osteogenic stimulus was calculated with an approach similar to that used to predict epiphyseal ossification. The results suggest that, compared with conforming joints, nonconformity between the sesamoid cartilage and its articulating surface, which arises during early development, produces higher contact pressures within the sesamoid and leads to a thicker articular cartilage layer. For a nonconforming joint surface, the results suggest that ossification is favored anywhere within a broad internal region of the sesamoid, whereas a layer at the articular surface will remain cartilaginous. These findings highlight the subtle differences between ossification processes in epiphyses and sesamoids, indicating that the mechanical stress environment in sesamoids produces a diffuse stimulus leading to the onset of ossification and that the degree of joint nonconformity may influence the thickness of the articular cartilage layer. PMID- 11117291 TI - Expression of cartilage oligomeric matrix protein (COMP) by embryonic and adult osteoblasts. AB - Cartilage oligomeric matrix protein has been implicated as an important component of endochondral ossification because of its direct effects on chondrocytes. The importance of this protein for skeletal development and growth has been recently illustrated by the identification of mutations in cartilage oligomeric protein genes in two types of inherited chondrodysplasias and osteoarthritic phenotypes: multiple epiphyseal dysplasia and pseudoachondroplasia. In the present study, we report the presence of cartilage oligomeric protein in embryonic and adult osteoblasts. A foot from a 21-week-old human fetus, subchondral bone obtained from knee replacement surgery in an adult patient, and a limb from a 19-day postcoital mouse embryo were analyzed with immunostaining and in situ hybridization. In the human fetal foot, cartilage oligomeric protein was localized to osteoblasts of the bone collar and at the newly formed bone at the growth plate and bone diaphyses. Immunostaining was performed on the adult subchondral bone and showed positive intracellular staining for cartilage oligomeric protein of the osteoblasts lining the trabecular bone. There was no staining of the osteocytes. Immunostaining of the mouse limb showed the most intense staining for cartilage oligomeric protein in the hypertrophic chondrocytes and in the surrounding osteoblast cells of the developing bone. Cartilage oligomeric protein mRNA and protein were detected in an osteoblast cell line (MG-63), and cartilage oligomeric protein mRNA was detected from human cancellous bone RNA. These results suggest that the altered structure of cartilage oligomeric protein by the mutations seen in pseudoachondroplasia and multiple epiphyseal dysplasia may have direct effects on osteoblasts, contributing to the pathogenesis of these genetic disorders. PMID- 11117292 TI - Inhibitory effects of the quinolone antibiotics trovafloxacin, ciprofloxacin, and levofloxacin on osteoblastic cells in vitro. AB - We studied the inhibitory effects of the fluoroquinolones levofloxacin, ciprofloxacin, and trovafloxacin on growth and extracellular matrix mineralization in MC3T3-E1 osteoblast-like cell cultures. Levofloxacin had the least inhibitory effect on cell growth, with a 50% inhibitory concentration of approximately 80 microg/ml at 48 and 72 hours. Ciprofloxacin had an intermediate degree of inhibition, with a 50% inhibitory concentration of 40 microg/ml at 48 and 72 hours. Trovafloxacin exerted a profound inhibitory effect on cell growth, with a 50% inhibitory concentration of 0.5 microg/ml, lower than clinically achievable serum levels. The decreased cell counts with up to 2.5 microg/ml of trovafloxacin and with up to 40 microg/ml of ciprofloxacin were not associated with decreased rates of 5-bromo-2'-deoxyuridine incorporation per cell. Alatrovafloxacin, the L-alanyl-l-alanine prodrug of trovafloxacin, exerted effects on proliferation and 5-bromo-2'-deoxyuridine incorporation similar to those of the parent compound. The quinolones evaluated also inhibited extracellular matrix mineralization by MC3T3-E1 cells. Treatment of confluent cultures with trovafloxacin, ciprofloxacin, or levofloxacin resulted in strong inhibition of calcium deposition, as determined on day 14 by alizarin red staining and biochemical analysis. The effect was apparent with 2.5-5 microg/ml of each of the three antibiotics tested and progressively increased to more than a 90% decline in the calcium/protein ratio with 20-40 microg/ml antibiotic concentration. Further in vivo studies are advocated to evaluate the relevance of the in vitro cytotoxicity reported here to bone healing in orthopaedic patients. PMID- 11117294 TI - Changes in cross-sectional geometry of the distal femoral metaphysis associated with inflammatory arthritis are reduced by a bisphosphonate (zoledronate). AB - An increased risk of fracture is a feature of rheumatoid arthritis and of animal models of inflammatory arthritis. We examined geometrical changes in the metaphyseal cortex of the distal femur in an animal model of inflammatory arthritis. Additionally, we examined the effect of a bisphosphonate in preventing these changes. Five groups of rabbits were studied: normal controls, those with inflammatory arthritis, and three groups with arthritis treated with bisphosphonate. To determine geometrical properties, image analysis was performed on digitized cross sections of the femoral metaphyseal cortices. The results demonstrated that the posterior cortical wall was significantly less thick in rabbits with arthritis than in normal rabbits and in the rabbits in the three bisphosphonate treatment groups (p < 0.05). Moment of inertia about the lateral medial axis was reduced in rabbits with arthritis compared with normal rabbits (p < 0.05). Cross-sectional area was not significantly different between groups. The changes suggest a mechanism of weakening of bone in arthritis; when the results are coupled with results of previous porosity studies, severe directional weakness is apparent. Bisphosphonate was effective in preserving bone integrity in inflammatory arthritis. PMID- 11117293 TI - Markers of joint tissue turnover in joint fluids from hips with osteonecrosis of the femoral head. AB - Osteonecrosis of the femoral head often results in secondary osteoarthritis of the hip joint; however, the pathologic processes underlying the destruction of articular cartilage are not fully understood. Molecular markers in the hip joint fluids were measured to examine the changes in turnover of cartilage and other joint tissues. Marker data were related to clinical, radiological, and histopathological changes in the articular cartilage of the hip. Forty-five patients (median age: 43 years) were studied. The median time between the onset of symptoms and sampling of hip synovial fluid was 6 months. Aggrecan fragments, C-propeptide of type-II collagen, matrix metalloproteinase-3, and tissue inhibitor of metalloproteinases-1 levels in joint fluid were determined by immunoassay. Osteonecrosis of the femoral head was graded by radiology as minimal collapse of the femoral head (stage 2: 26 patients), severe collapse (stage 3: 15 patients), or severe collapse with osteoarthritis (stage 4: four patients). Histological changes of the articular cartilage, consistent with early-stage osteoarthritis, were evident at stage 3 and were more advanced at stage 4. The average concentrations of proteoglycan fragments and C-propeptide of type-II collagen were 207 (SD 182) microg/ml and 19.6 (SD 19.3) ng/ml, respectively. The average concentrations of matrix metalloproteinase-3 and tissue inhibitor of metalloproteinases-1 were 177 (SD 291) nM and 23.0 (SD 9.9) nM, respectively. Measurable levels for all markers assayed were noted in the earliest stage of the disease, only a few months after the onset of symptoms and well before the appearance of radiological changes. Levels of matrix metalloproteinase-3 and molar ratios of matrix metalloproteinase-3/tissue inhibitor of metalloproteinases 1 were higher in early stage disease than in later stage disease. PMID- 11117295 TI - Comparison of biomechanical and biochemical properties of cartilage from human knee and ankle pairs. AB - Cartilage was obtained from eight matched knee (tibiofemoral and femoropatellar) and ankle (talocrural) joints of five different donors (both left and right from donors 14, 22, and 38 years of age, and left only from donors 31 and 45 years of age) within 24 hours of death. All cartilage was graded as normal by the macroscopic visual Collins' scale and the histological Mankin scale. Cylindrical disks of cartilage were harvested from 10 sites within the tibiofemoral and femoropatellar joint surfaces and four sites within the talocrural joint, and uniaxial confined compression measurements were performed to quantify a spectrum of physical properties including the equilibrium modulus, hydraulic permeability, dynamic stiffness, streaming potential, electrokinetic coupling coefficient, and electrical conductivity. Matched specimens from the same 14 sites were used for complementary measurements of biochemical composition and molecular interaction, including water content, hypotonic swelling behavior, and sulfated glycosaminoglycan and collagen contents. In comparison of the top 1-mm slices of talar cartilage with the top 1-mm of tibiofemoral cartilage, the talar cartilage appeared denser with a higher sulfated glycosaminoglycan content, lower water content, higher equilibrium modulus and dynamic stiffness, and lower hydraulic permeability. The equilibrium modulus increased with increasing sulfated glycosaminoglycans per wet weight and decreased with increasing water content for all joint surfaces. Multiple linear regression showed that greater than 80% of the variation in the equilibrium modulus could be accounted for by variations in the biochemical parameters (water content, sulfated glycosaminoglycans/wet weight, and hydroxyproline content/wet weight) for each joint surface. Nonhomogeneous depth-dependent changes in the physical properties and biochemical composition of full-thickness distal femoral cartilage were consistent with previous reports. Since the compressive deformation of cartilage during cyclic loading is confined to the more superficial regions, the differences in properties of the upper regions of the talar compared with tibiofemoral or femoropatellar cartilage may be important in the etiology of osteoarthritis. PMID- 11117296 TI - Distribution of chondrocytes containing alpha-smooth muscle actin in human articular cartilage. AB - Relatively little is known about the contractile behavior of the human articular chondrocyte. Other connective tissue cells are known to express a contractile actin isoform, alpha-smooth muscle actin, in response to injury, at selected stages of wound healing, and in certain pathological conditions. This and recent work demonstrating contractile behavior in adult canine articular chondrocytes in vitro prompted the present study of the distribution of alpha-smooth muscle actin containing chondrocytes in human articular cartilage. Approximately 75% of the chondrocytes in the superficial region of cartilage expressed alpha-smooth muscle actin as demonstrated by immunohistochemistry. In contrast, only approximately 10% of the cells in the deep region stained for this contractile actin isoform. There was no correlation of the percentage of alpha-smooth muscle actin-positive cells in either region with Mankin grade or with age. This is the first report of a contractile potential for human articular chondrocytes. The roles of alpha smooth muscle actin in these cells warrant further investigation. The question of whether it is necessary to refer to these cells as myochondrocytes is considered. PMID- 11117297 TI - Polysulphated glycosaminoglycan treatments can mitigate decreases in stiffness of articular cartilage in a traumatized animal joint. AB - A single, blunt impact to the rabbit patellofemoral joint has been shown to decrease the stiffness of retropatellar cartilage and increase the thickness of the underlying bone. Polysulphated glycosaminoglycan treatments, on the other hand, have been shown to inhibit the degradation of articular cartilage and possibly increase synthesis of collagen and glycosaminoglycans in experimental studies on diseased joints. The aim of the current study was to examine the effect of early treatments with polysulphated glycosaminoglycans on cartilage using an in vivo post-trauma animal model. The study used 24 Flemish Giant rabbits in three groups: control, impacted, and impacted with treatment. Treatment consisted of intramuscular injections the day of insult and every 4 days thereafter for 6 weeks. At 30 weeks after trauma, mechanical tests were performed on the retropatellar cartilage to determine its mechanical stiffness. The patellae were also grossly evaluated for surface lesions on the retropatellar cartilage and histologically processed to measure the thickness of the subchondral bone. The rabbits that received no treatment had a statistically significant decrease in stiffness (modulus) for the cartilage of the impacted patellae compared with that of the contralateral, unimpacted patellae and compared with the cartilage of rabbits in the control group. The degradation in mechanical stiffness, however, was not observed in patellae of rabbits in the group receiving treatment. There was also a significant increase in the underlying thickness of the subchondral plate on the impacted patellae compared with that on the contralateral, unimpacted sides for rabbits in both the treated and nontreated groups. In conclusion, the polysulphated glycosaminoglycan treatments minimized a decrease in mechanical stiffness (modulus) of retropatellar articular cartilage 30 weeks after trauma. The mechanism by which the mechanical stiffness of the cartilage was preserved is unknown. PMID- 11117298 TI - Quantitative assessment of the effect of 0.05% chlorhexidine on rat articular cartilage metabolism in vitro and in vivo. AB - Wound infection remains a problem. Syringe and needle jet lavage of chlorhexidine gluconate 0.05% removed or killed 99.8% of contaminating bacteria within 1 minute in a wound model. In clinical use, however, possible toxicity to articular cartilage is a concern. In an established intact rat patella model in vitro, 1 minute of exposure to chlorhexidine 0.05% and chlorhexidine jet lavage did not significantly alter cartilage metabolism. A 1-hour exposure decreased metabolic activity. In vivo, a 30-minute exposure with or without rinsing produced no impairment of metabolic activity 6 weeks later, suggesting that cartilage has the potential for biological recovery. However, injecting and leaving chlorhexidine 0.05% in the joints was detrimental to the metabolic activity of the articular cartilage as assessed 6 weeks later. Thus, chlorhexidine gluconate 0.05% could be used on normal articular cartilage. Any potential damage from prolonged exposure can be avoided by rinsing after 1 minute. PMID- 11117299 TI - Disc degeneration and bone density in monozygotic twins discordant for insulin dependent diabetes mellitus. AB - The effects of insulin-dependent diabetes mellitus on bone density and connective tissue degeneration have theoretical interest and practical relevance. Several experimental studies in animals have demonstrated the harmful effects of insulin deficiency on connective tissues. However, clinical studies in humans have produced somewhat contradictory results, most likely due to difficulties controlling for general degeneration and factors associated with diabetes. In nine pairs of monozygotic twins discordant for insulin-dependent diabetes mellitus, we compared femoral and lumbar bone mineral density (assessed by dual energy x-ray absorptiometry) and spinal degeneration (assessed by magnetic resonance imaging). The bone densities were, on average, 0.1-0.3% lower (p = 0.87 0.96) in diabetic patients. However, after controlling for smoking, we found that the bone density in the femoral neck was 2.5% (0.025 g/cm2) lower in diabetic individuals than in their twins (p = 0.09). The five magnetic resonance imaging parameters used to evaluate disc degeneration did not differ between diabetic patients and their twins. In conclusion, our results provide no evidence that insulin-dependent diabetes mellitus has any major effect on bone density or disc degeneration. PMID- 11117300 TI - Hyaluronan-based polymers in the treatment of osteochondral defects. AB - Articular cartilage in adults has limited ability for self-repair. Some methods devised to augment the natural healing response stimulate some regeneration, but the repair is often incomplete and lacks durability. Hyaluronan-based polymers were tested for their ability to enhance the natural healing response. It is hypothesized that hyaluronan-based polymers recreate an embryonic-like milieu where host progenitor cells can regenerate the damaged articular surface and underlying bone. Osteochondral defects were made on the femoral condyles of 4 month-old rabbits and were left empty or filled with hyaluronan-based polymers. The polymers tested were ACP sponge, made of crosslinked hyaluronan, and HYAFF-11 sponge, made of benzylated hyaluronan. The rabbits were killed 4 and 12 weeks after surgery, and the condyles were processed for histology. All 12-week defects were scored with a 29-point scale, and the scores were compared with a Kruskall Wallis analysis of variance on ranks. Untreated defects filled with bone tissue up to or beyond the tidemark, and the noncalcified surface layer varied from fibrous to hyaline-like tissue. Four weeks after surgery, defects treated with ACP exhibited bone filling to the level of the tidemark and the surface layer was composed of hyaline-like cartilage well integrated with the adjacent cartilage. At 12 weeks, the specimens had bone beyond the tidemark that was covered with a thin layer of hyaline cartilage. Four weeks after surgery, defects treated with HYAFF-11 contained a rim of chondrogenic cells at the interface of the implant and the host tissue. In general, the 12-week defects exhibited good bone fill and the surface was mainly hyaline cartilage. Treated defects received significantly higher scores than untreated defects (p < 0.05), and ACP-treated defects scored significantly higher than HYAFF-11-treated defects (p < 0.05). The introduction of these hyaluronan-based polymers into defects provides an appropriate scaffolding and favorable microenvironment for the reparative process. Further work is required to fully assess the long-term outcome of defects treated with these polymers. PMID- 11117301 TI - Healing of canine articular cartilage defects treated with microfracture, a type II collagen matrix, or cultured autologous chondrocytes. AB - The effects of three different treatments on the healing of articular cartilage defects were compared with use of a previously developed canine model. In the articular surface of the trochlear grooves of 12 adult mongrel dogs, two 4-mm diameter defects were made to the depth of the tidemark. Four dogs were assigned to each treatment group: (a) microfracture treatment, (b) microfracture with a type-II collagen matrix placed in the defect, and (c) type-II matrix seeded with cultured autologous chondrocytes. After 15 weeks, the defects were studied histologically. Data quantified on histological cross sections included areal or linear percentages of specific tissue types filling the defect, integration of reparative tissue with the calcified and the adjacent cartilage, and integrity of the subchondral plate. Total defect filling (i.e., the percentage of the cross sectional area of the original defect filled with any type of reparative tissue) averaged 56-86%, with the greatest amount found in the dogs in the microfracture group implanted with a type-II collagen matrix. The profiles of tissue types for the dogs in each treatment group were similar: the tissue filling the defect was predominantly fibrocartilage, with the balance being fibrous tissue. There were no significant differences in the percentages of the various tissue types among dogs in the three groups. PMID- 11117302 TI - Effects of harvest and selected cartilage repair procedures on the physical and biochemical properties of articular cartilage in the canine knee. AB - This study utilizes a canine model to quantify changes in articular cartilage 15 18 weeks after a knee joint is subjected to surgical treatment of isolated chondral defects. Clinical and experimental treatment of articular cartilage defects may include implantation of matrix materials or cells, or both. Three cartilage repair methods were evaluated: microfracture, microfracture and implantation of a type-II collagen matrix, and implantation of an autologous chondrocyte-seeded collagen matrix. The properties of articular cartilage in other knee joints subjected to harvest of articular cartilage from the trochlear ridge (to obtain cells for the cell-seeded procedure) were also evaluated. Physical properties (thickness, equilibrium compressive modulus, dynamic compressive stiffness, and streaming potential) and biochemical composition (hydration, glycosaminoglycan content, and DNA content) of the cartilage from sites distant to the surgical treatment were compared with values measured for site-matched controls in untreated knee joints. No significant differences were seen in joints subjected to any of the three cartilage repair procedures. However, a number of changes were induced by the harvest operation. The largest changes (displaying up to 3-fold increases) were seen in dynamic stiffness and streaming potential of patellar groove cartilage from joints subjected to the harvest procedure. Whether the changes reported will lead to osteoarthritic degeneration is unknown, but this study provides evidence that the harvest procedure associated with autologous cell transplantation for treatment of chondral defects may result in changes in the articular cartilage in the joint. PMID- 11117303 TI - Differential expression of transforming growth factor-alpha and macrophage colony stimulating factor/colony-stimulating factor-1R (c-fins) by multinucleated giant cells involved in pathological bone resorption at the site of orthopaedic implants. AB - The immunologic response to prosthetic biomaterial particles is characterized by macrophage-rich inflammatory infiltrate, formation of multinucleated giant cells, and aseptic loosening at the site of arthroplasty. We investigated the in vivo expression and tissue distribution of transforming growth factor alpha, macrophage colony-stimulating factor, and the receptor for colony-stimulating factor-1 at the site of bone erosion in patients with clinically failed orthopaedic implants (n = 30). The expression was further compared with that detected in the inflamed synovial membranes from patients with rheumatoid arthritis or osteoarthritis (n = 15) and one patient with osteoclastoma (giant cell tumour of bone). Immunostaining of the tissue demonstrated positivity for transforming growth factor alpha within the inflammatory macrophage and multinucleated giant cell infiltrate in the diseased synovial membrane and the bone-implant interface. A comparative analysis between the synovium and retrieval interface membranes (pseudosynovium) revealed a high level of expression of transforming growth factor alpha, with intense membrane staining on multinucleated giant cells in all failed arthroplasties with pseudosynovium. In addition, the frequency, antigenic phenotype, and pattern of transforming growth factor alpha expression on multinucleated giant cells in the interface were markedly similar to those observed for multinucleated giant cells in osteoclastoma. Multinucleated giant cells within the interface lacked the expression of macrophage colony-stimulating factor and colony-stimulating factor 1 receptor, whereas those at the bone surfaces exhibited strong immunoreactivity. The predominant expression of transforming growth factor alpha by multinucleated giant cells in the bone-implant interface and its similarity to osteoclastoma highlight the importance of assessing transforming growth factor alpha as a possible contributor to the development of bone-resorbing giant cells at the site of failed orthopaedic implants. PMID- 11117304 TI - Loosening of sacral screw fixation under in vitro fatigue loading. AB - Sacral screw fixation is frequently used for fusion of the lower lumbar spine, but sacral screws appear to offer less secure fixation than lumbar pedicle screws, and failure due to loosening under fatigue loading is common. The aim of this study was to examine in vitro the stability of medial and lateral bicortical and unicortical sacral screw fixation under a physiologically relevant fatigue loading pattern. Bone mineral density, screw insertion torque, and screw-fixation stiffness were measured prior to cyclic loading between 40 and 400 N compression at 2 Hz for 20,000 cycles. The screw-fixation stiffness was measured every 500 cycles, and the axial pullout strength of the screws was recorded following loading. All of the lateral insertions loosened under the applied loading, but some of the medial insertions remained stable. Medial insertions proved stiffer and stronger than lateral insertions, and bicortical fixations were stronger than unicortical fixations. Bone mineral density and insertion torque were correlated with screw stiffness and pullout strength, although better correlation was found for insertion torque than bone mineral density. Bone mineral density is a good preoperative indicator of sacral screw-fixation strength, and insertion torque is a good intraoperative indicator. An insertion torque greater than 1.5 Nm is suggested as an indicative value for a stable medial unicortical insertion, whereas an insertion torque greater than 2 Nm suggests a stable medial bicortical insertion. It appears that, apart from the choice of technique (screw orientation and depth), minimizing the load on the screws during the initial part of the fusion process is also critical to maintain stability of the fused section and to obtain a solid fusion mass. PMID- 11117305 TI - Nitric oxide as a mediator of nucleus pulposus-induced effects on spinal nerve roots. AB - Nerve root dysfunction and sciatic pain in disc herniation are considered to be caused by mechanical compression and related to the presence of nucleus pulposus in the epidural space. Autologous nucleus pulposus has been shown to induce endoneural edema and to decrease nerve-conduction velocity in spinal nerve roots in experimental disc herniation models, and inflammatory mediators have been suggested to be involved in these mechanisms. Nitric oxide, a potent inflammatory mediator, is implicated in vasoregulation, neurotransmission, and neuropathic pain. Nitric oxide synthesis can be induced by different cytokines, e.g., tumor necrosis factor-alpha, which recently was shown to be of pathophysiological importance in experimental disc herniation. The enzyme nitric oxide synthase mediates the production of nitric oxide. Three series of experiments were performed in rat and pig disc herniation models to (a) investigate nitric oxide synthase activity in spinal nerve roots after exposure to autologous nucleus pulposus and (b) evaluate the effects of systemic treatment with aminoguanidine, a nitric oxide synthase inhibitor, on vascular permeability and nerve-conduction velocity. In a disc herniation model in the rat, calcium-independent nitric oxide synthase activity was measured in nerve roots exposed to nucleus pulposus; however, no nitric oxide synthase activity was detected in nerve roots from animals that underwent a sham operation, reflecting increased inducible nitric oxide synthase activity. In nucleus pulposus-exposed spinal nerve roots in the pig, the edema was less severe after systemic aminoguanidine administration than without aminoguanidine treatment. Aminoguanidine treatment also significantly reduced the negative effect of nucleus pulposus on nerve-conduction velocity in spinal nerve roots in the pig. These results demonstrate that nucleus pulposus increases inducible nitric oxide synthase activity in spinal nerve roots and that nitric oxide synthase inhibition reduces nucleus pulposus-induced edema and prevents reduction of nerve-conduction velocity. Furthermore, the results suggest that nitric oxide is involved in the pathophysiological effects of nucleus pulposus in disc herniation. PMID- 11117306 TI - Radiographic determinants of the elbow rotation axis: experimental identification and quantitative validation. AB - This study identifies new radiographic indices to approximate the location of the elbow rotational axis. With use of electromagnetic motion tracking source data, the average rotational axis of the ulnohumeral articulation was calculated in seven cadaveric specimens. Quasi-lateral radiographs of the elbow specimens were then analyzed to identify radiographic landmarks of the elbow axis in the lateral view. The spatial relationships of these landmarks with the elbow aligned on-axis were contrasted with their relationships in eight distinct off-axis alignments. Elbow axis orientation in the transverse plane (internal/external rotation) was identified by the location of a dense intramedullary cortical line, appearing in the projection of the distal humerus in relation to the periosteal surface of the posterior cortex of the humerus. This intramedullary line corresponds to the posteromedial cortex of the distal humerus. Correct alignment occurred when this line laid 27.1+/-3.7% of the anteroposterior humeral diameter anterior from the periosteal surface of the posterior cortex. Axis orientation in the coronal plane (abduction/adduction) was identified by the concentric appearance of radiographic arcs formed by the capitellum, trochlear sulcus, and medial trochlear flange. Using these radiographic indices, three orthopaedic surgeons were able to fluoroscopically align the elbow along the axis of rotation with an accuracy of 3.7+/-1.8 degrees. These results are immediately applicable to fluoroscopic identification of the elbow axis. This technique can be used to increase the accuracy of hinge placement during application of hinged external fixation or distraction arthroplasty. PMID- 11117307 TI - Structural properties of the subscapularis tendon. AB - The subscapularis muscle is an important mover and stabilizer of the glenohumeral joint. The purpose of this study was to measure regional variations in the structural properties of the subscapularis tendon in two joint positions. Subscapularis tendons from cadaveric shoulders were divided into four sections superiorly to inferiorly and tested to failure at 0 or 60 degrees of glenohumeral abduction. Arm position had a significant influence on stiffness in the inferior and superior portions (p < 0.05). The inferior region showed a higher stiffness in the hanging-arm position (0 degrees) than at 60 degrees of abduction (27.4+/ 17.7 compared with 9.5+/-5.9 N/mm). Meanwhile, stiffness of the superior portion was higher at 60 degrees of abduction than in the hanging-arm position (208.7+/ 60.9 compared with 147.2+/-32.3 N/mm). In the hanging-arm position (0 degrees) and at 60 degrees of abduction, the superior and midsuperior portions failed at significantly higher loads (superior: 623.2+/-198.6 and 478.2+/-206.6 N at 0 and 60 degrees of abduction, respectively; midsuperior: 706.2+/-164.6 and 598.4+/ 268.4 N, respectively) than did the inferior portion (75.1+/-54.2 and 30.3+/-13.0 N, respectively). Likewise, stiffness of the superior and midsuperior portions was significantly higher than that of the inferior region in both positions. Higher stiffness and ultimate load in the superior tendon region may explain the infrequent extension of rotator cuff tears into the subscapularis tendon. Conversely, the significantly lower ultimate load and stiffness in the inferior tendon region could facilitate anterior dislocation of the humeral head when this portion stabilizes the joint in a dislocated position. Therefore, repair of torn inferior portions of the subscapularis tendon should be considered in surgery for glenohumeral instability. PMID- 11117309 TI - Equivalence of single and incremental subfailure stretches of rabbit anterior cruciate ligament. AB - Experimental models are often used in the laboratory to produce incomplete soft tissue injuries simulating those observed clinically. Single and incremental stretch protocols have been utilized. The latter has many advantages over the former. This study was designed to determine if incremental and single ligamentous stretches are biomechanically equivalent. Eleven paired fresh rabbit bone-anterior cruciate ligament-bone preparations were used. One of each pair (single-stretch protocol) was stretched to 88% of the average failure deformation and then stretched to failure. The other ligament (incremental-stretch protocol) was stretched to 55, 66, 77, and 88% of the average failure deformation and then stretched to failure. All stretches were performed at 1.2 m/sec. Stress relaxation tests were performed before and after the 88% stretch for both stretch protocols. Relaxation curves were parameterized as forces at six time points and were also fitted to a three-element model. Load-deformation curves recorded during stretch to failure were characterized by eight parameters. Each incremental stretch step produced a significant increase in deformation, indicating alteration in the mechanical properties of the ligament. Both groups of ligaments, when intact, exhibited no differences in relaxation curves (p > 0.2). The 88% stretches, produced by each of the two stretch protocols, significantly altered the viscoelastic behavior of the ligaments (p < 0.002). However, after the 88% stretch, there were no differences in either viscoelastic (p > 0.1) or load-deformation (p > 0.1) parameters of the two stretch protocols. In conclusion, the 88% subfailure stretch significantly altered the mechanical properties of ligament, and the incremental and single stretches were biomechanically equivalent. PMID- 11117308 TI - Mixture of hyaluronic acid and phospholipid prevents adhesion formation on the injured flexor tendon in rabbits. AB - Dipalmitoyl phosphatidylcholine, a highly surface-active polar lipid, has been implicated as a potential boundary lubricant for synovial joints. We examined the effects of dipalmitoyl phosphatidylcholine on the flexor tendon and its protective effect against postoperative adhesion in two experimental steps. First, the flexor digitorum fibularis and the distal pulley of rabbits were set for a friction test. The test was performed with saline solution, sodium hyaluronate, or a mixture of dipalmitoyl phosphatidylcholine and sodium hyaluronate as the lubricant. The friction coefficient was significantly lower with the mixture of dipalmitoyl phosphatidylcholine and sodium hyaluronate than with saline solution or sodium hyaluronate. We concluded that the decreased friction coefficient indicates that dipalmitoyl phosphatidylcholine could complement the boundary-lubricating ability of the tendon. In the second experiment, we used an experimental adhesion model of the flexor digitorum fibularis in the rabbit. During the operation, either saline solution, sodium hyaluronate, or a mixture of dipalmitoyl phosphatidylcholine and sodium hyaluronate was injected into the tendon sheath. The specimen was sent to another tester, and the work required to tear off the adhesion was measured. The work required was significantly greater for the tendons that had been injected with saline solution than for those given injections of dipalmitoyl phosphatidylcholine and sodium hyaluronate. Our findings suggest that dipalmitoyl phosphatidylcholine plays an important role in the boundary lubrication of the tendon and that after tendon injury, the administration of a mixture of dipalmitoyl phosphatidylcholine and sodium hyaluronate may improve tendon lubrication and prevent adhesion formation. PMID- 11117310 TI - Lactones of methyl 3-O. AB - Lactones of methyl 3-O-[(R)- and (S)-1-carboxyethyl]-alpha-D-gluco-, galacto- and manno-pyranoside were prepared by treatment of the sugar derivatives in acetic acid. The lactones were formed between the 1-carboxyethyl substituent and 2-OH or 4-OH in different proportions depending on the stereochemistry of the parent compounds. Relative formation rates in acetic acid-d4 and hydrolysis rates in buffered D2O solutions at pD 2.4, 4.6 and 7.4 were estimated. Hydrolysis of the formed lactones is relatively slow in D2O at pD 4.6, which permitted characterization of the lactones by 1H and 13C NMR spectroscopy in buffered D2O solutions. Hydrolysis of the lactones in 1 M aqueous NaOH at 80 degrees C gave no detectable isomerization of the alpha-carbon. The set of lactones formed from the 1-carboxyethyl substituted methyl glycosides used in this study showed large similarities in the NMR shifts (delta delta values). Deviations from the observed shift pattern were found for two lactones. Our findings strongly suggest that those two lactones differ from the rest by adopting a boat-like conformation, whereas the others adopt pseudo-chair conformations. PMID- 11117311 TI - Synthesis of 4-cyano- and 4-nitrophenyl 2,5-anhydro- 1,6-dithio-alpha-D-gluco- and -alpha-L-guloseptanosides carrying different substituents at C-3 and C-4. AB - Treatment of 1,6:2,5-dianhydro-3,4-di-O-methanesulfonyl-1-thio-D-glucitol in methanol with sodium hydroxide afforded 1,6:2,5:3,4-trianhydro-1-thio-allitol, 1,4:2,5-dianhydro-6-methoxy-1-thio-D-galactitol, 1,6:2,5-dianhydro-4-O-methyl-1 thio-D-glucitol, 1 ,6:2,5-dianhydro-3-O-methanesulfonyl-1 -thio-D-glucitol and 1 ,6:2,5-dianhydro-4-deoxy-1-thio-D-erythro-hex-3-ulose (14) in 5, 4, 28, 5.5 and 41% yield, respectively. Formation of these derivatives can be explained via a common sulfonium intermediate. Reduction of 14 with sodium borohydride and subsequent acetylation afforded 3-O-acetyl-1,6:2,5-dianhydro-4-deoxy-1-thio-D xylo-hexitol, the absolute configuration of which was proved by X-ray crystallography. The 1,6:2,5-dianhydro-1-thio-D-hexitol derivatives in which the free OH groups were protected by acetylation, methylation or mesylation were converted by a Pummerer reaction of their sulfoxides into the corresponding 1-O acetyl hexoseptanose derivatives which were used as donors for the glycosidation of 4-cyano- and 4-nitrobenzenethiol, respectively. The Pummerer reaction of 1,6:2,5-dianhydro-4-deoxy-3-O-methyl-1-thio-D-xylo-hexitol S-oxide gave, besides 1-O-acetyl-2,5-anhydro-3-deoxy-4-O-methyl-6-thio-alpha-L- (23) and 1-O-acetyl-2,5 anhydro-4-deoxy-3-O-methyl-6-thio-alpha-D-xylo-hexoseptanose (25), 1-O-acetyl-4 deoxy-2,6-thioanhydro-D-lyxo-hexopyranose, formed in a rearrangement reaction. The same rearrangement took place, when a mixture of 23 and 25 was used as donor in the glycosidation reaction with 4-cyanobenzenethiol, applying trimethylsilyl triflate as promoter. The oral antithrombotic activity of the obtained alpha thioglycosides was determined in rats, using Pescador's model. PMID- 11117312 TI - Synthesis and glycan priming activity of acetylated disaccharides. AB - Five disaccharides related in structure to the glycans of vertebrate mucins have been chemically synthesized using orthogonal blocking, coupling and deblocking techniques. These include 2-naphthylmethyl 3,4,6-tetra-O-acetyl-beta-D galactopyranosyl-( 1 --> 4)-2-acetamido-3,6-di-O-acetyl-2-deoxy-beta-D glucopyranoside (6), 2-naphthylmethyl 2-aceta-mido-3,4,6-tri-O-acetyl-2-deoxy beta-D-glucopyranosyl-(1 --> 3)-2,4,6-tri-O-acetyl-beta-D-galactopyranoside (14), 2-naph-thylmethyl2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl-(1 --> 3)-2 acetamido-4,6-di- O-acetyl-2-deoxy-alpha-D-galactopyranoside (20), 2 naphthylmethyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D-glucopyranosyl-(1 - > 3)-2-acetamido-4,6-di-O-acetyl-2-deoxy-alpha-D-galactopyranoside (23) and 2 naphthylmethyl 2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D-glu-copyranosyl-(1 - > 6)-2-acetamido-3,4-di-O-acetyl-2-deoxy-alpha-D-galactopyranoside (27). These per-O-acetylated compounds were fed to U-937 cells to test their ability to prime oligosaccharide synthesis, inhibit glycoprotein biosynthesis and alter adhesion to E-selectin expressed on endothelial cells. The results show that 6, 14, and 20 served as substrates for oligosaccharide synthesis. The generation of glycoside primed glycans altered the formation of glycoproteins on the cell surface and inhibited cell adhesion dependent on E-selectin. PMID- 11117313 TI - New syntheses of 1D- and 1L-1,2-anhydro-myo-inositol and assessment of their glycosidase inhibitory activities. AB - The 1D and 1L enantiomers of 1,2-anhydro-myo-inositol (conduritol B epoxide) were synthesised from 1D-pinitol and 1L-quebrachitol, respectively, and their activities were compared in selected glycosidase inhibition assays. The 1D enantiomer was found to be the active isomer, functioning as an irreversible inhibitor of sweet almond beta-D-glucosidase. Neither isomer was active against the alpha-D-glucosidase from Bacillus stearothermophilus or the beta-D galactosidase from Aspergillus oryzae. PMID- 11117314 TI - Synthesis of the Lewis b hexasaccharide and squarate acid-HSA conjugates thereof with various saccharide loadings. AB - The Lewis b hexasaccharide, alpha-L-Fucp-(1 --> 2)-beta-D-Galp-(1 --> 3)-[alpha-L Fucp-(1 --> 4)]-beta-D-GlcpNAc-(1 --> 3)-beta-D-Galp-(l --> 4)-beta-D-Glcp, has been synthesised using a convergent synthesis. Starting from ethyl 4,6-O benzylidene-2-deoxy-2-phthalimido-1-thio-beta-D-glucopyranoside, a thioglycoside tetrasaccharide donor block, was constructed through two orthogonal glycosylations with glycosyl bromide donors. First, the galactose moiety was introduced using silver triflate as a promoter and then the two fucose residues under halide-assisted conditions. Finally, this tetrasaccharide was linked to a spacer-equipped 3I,4I-diol lactose acceptor in a DMTST-promoted coupling to give, after deprotection, the Lewis b hexasaccharide as its 2-aminoethyl spacer equipped derivative. This was coupled to human serum albumin (HSA), using the squarate ester methodology, in various saccharide-protein ratios, to give neoglycoconjugates with different saccharide loadings in about 50%) efficiency. PMID- 11117315 TI - Stereoselective synthesis of 1,2-cis- and 2-deoxyglycofuranosyl azides from glycosyl halides. AB - Protected 1,2-cis glycofuranosyl azides with alpha-D-ribo, beta-D-arabino and 2 deoxy-2-fluoro-beta-D-arabino configurations were efficiently prepared from the appropriate 1,2-trans glycosyl halides bearing non-participating 0-2 substituent by inversion with sodium azide under phase transfer catalytic conditions (80-85% yields, 90-96% de). The same method failed to result in sufficiently good beta selectivity starting from 2-deoxy-3,5-di-O-(p-toluoyl)-alpha-D-ery-thro pentofuranosyl chloride (5alpha) (40% de). The selectivity in favour of the protected 2-deoxy-beta-D-erythro-pentofura-nosyl azides was greatly improved (74 80% de) by treating 5alpha and its p-chlorobenzoyl analog 6alpha with cesium or potassium azide in dimethylsulfoxide at room temperature (83-85% yields). PMID- 11117316 TI - Synthesis of kanamycin A analogs containing a 6-amino-6-deoxyglycofuranose moiety. AB - Three kanamycin A analogs containing 6-amino-6-deoxyglycofuranoses have been prepared as candidates for potential activity against resistant bacteria producing 6'-N-acetyltransferase. They are 4-O-(6-amino-3,5,6-trideoxy-alpha-D-, beta-D-, and -beta-L-erythro -hexofuranosyl)-6-O-(3-amino-3-deoxy-alpha-D glucopyranosyl)-2,5-dideoxy-5-epi-5-fluorostreptamine. Structure-activity relationships of these compounds are discussed. PMID- 11117317 TI - FAB CIDMS/MS analysis of partially methylated maltotrioses derived from methylated amylose: a study of the substituent distribution. AB - Amylose was methylated with CH3I in alkaline aqueous suspension, yielding methylated amylose (MeAl) with a degree of substitution of 1.44 (s < 0.01). Determination of the monomer composition showed that HO-6 and HO-2 were highly substituted in contrast to HO-3 (7:2:5.5, HO-2:HO-3:HO-6). By using partial acid hydrolysis, oligomers were prepared that varied both in degree of polymerisation and in methyl-content. Studies on the distribution of substituents in trimers showed large deviations from random distributions. By using CID tandem mass spectrometry, the substituent distribution in these trimers was determined in more detail. Various sets of trimers with equal amounts of methyl-groups but differing in substituted positions were quantified. From the monomer composition of MeAl, the probability of each trimer was calculated and compared to the outcome of the measured distributions. It was concluded that trimers with terminal tri- or non-substituted glucose monomers at the non-reducing end were formed preferentially during partial hydrolysis and that partial hydrolysis of MeAl yielded oligomers in a non-random way. This is the first study that describes the partial hydrolysis of MeAl in such detail. PMID- 11117318 TI - The structure of the carbohydrate backbone of the core-lipid A region of the lipopolysaccharide from Proteus mirabilis serotype O28. AB - The following structure of the lipid A-core region of the lipopolysaccharide (LPS) from Proteus mirabilis serotype O28 was determined using NMR, MS, and chemical analysis of the core oligosaccharide, obtained by mild acid hydrolysis of LPS, and of the products of alkaline deacylation of the LPS: carbohydrate sequence [see text] where S = beta-GalALys (amide of beta-D galactopyranosyluronic acid with the alpha-amino group of L-lysine) or beta GalALys4PEtN are present in the ratio of approximately 1:1. beta-GalA and Ara4N (indicated by bold italics) are present in non stoichometric amount. All sugars are present in the pyranose form and all except L-Ara4N have the D configuration. PMID- 11117319 TI - A kinetic estimate of the free aldehyde content of aldoses. AB - The relative free aldehyde content of eight hexoses and four pentoses has been estimated within about 10% from the rate constants for their reaction with urazole (1,2,4-triazole-3,5-dione). These values of the percent free aldehyde are in agreement with those estimated from CD measurements, but are more accurate. The relative free aldehyde contents for the aldoses were then correlated to various literature NMR measurements to obtain the absolute values. This procedure was also done for three deoxyaldoses, which react much more rapidly than can be accounted for by the free aldehyde content. This difference in reactivity between aldoses and deoxyaldoses is due to the inductive effect of the H versus the OH on C-2'. This may help explain why deoxyribonucleosides hydrolyze much more rapidly than ribonucleosides. PMID- 11117320 TI - Characterization of doubly substituted polysaccharide derivatives. AB - Derivatives of cellulose, amylose and chitosan, bearing simultaneously 10 undecenoyl and arylaminocarbonyl or benzoyl groups were characterized by the combined use of 1H NMR and elemental analysis. The mathematical manipulation of elemental analysis data permits the calculation of the degree of substitution for each kind of substituent. The method was validated and is applicable to other derivatives. PMID- 11117321 TI - Diffusion of short chain alcohols from amorphous maltose-water mixtures above and below their glass transition temperature. AB - The apparent diffusion coefficient for short chain alcohols in undercooled maltose-water mixtures close to the calorimetric glass transition temperature, Tg, was measured by following desorption using headspace gas chromatography. The plasticising effect of the alcohols on Tg was characterised using differential scanning calorimetry. The initial appearance of alcohol in the headspace showed a linear dependence on the square root of time, allowing it to be modelled as a Fickian diffusive process. The diffusion coefficient decreased with increasing molecular size of alcohol and proximity to Tg. Close to the glass transition the variation of diffusion coefficient with temperature and composition does not follow that of viscosity and, for ethanol, divergence was observed at Tg/T> 0.88. PMID- 11117322 TI - NMR study of the complexation of D-gulonic acid with tungsten(VI) and molybdenum(VI). AB - By using multinuclear (1H, 13C, 17O, 95Mo, 183W) magnetic resonance spectroscopy (1D and 2D), D-gulonic acid is found to form ten and seven complexes, respectively, with tungsten(VI) and molybdenum(VI), in aqueous solution, depending on pH and metal-ligand molar ratios. Two isomeric 1:2 (metal-ligand) complexes involving the carboxylate and the adjacent OH group are present in the pH range 2-9. At intermediate and high pH, molybdate forms a 2:1 tetradentate complex involving the four secondary hydroxyl groups, whereas tungstate forms one 2:1 terdentate species. At low and intermediate pH values, three 2:1 complexes are found for both metals, involving the carboxylate group and three secondary hydroxyl groups, as well as a 5:2 species involving the carboxylate group and all the secondary hydroxyl groups; the concentration of this species increases in time mainly at the expense of 2:1 and 1:2 complexes. Tungstate can also form two additional species, probably a 5:2 species involving the carboxylate group and all the hydroxyl groups, and a 2:1 pentadentate species involving the carboxylate group and all the secondary hydroxyl groups. In alkaline solutions, tungstate is able to form an additional 2:1 pentadentate complex involving all the hydroxyl groups. PMID- 11117323 TI - Viscoelastic properties of pectin--co-solute mixtures at iso-free-volume states. AB - Small-deformation oscillatory measurements were performed on pectin-sucrose glucose syrup systems at a total level of solids of 81%, with the polysaccharide content being fixed at levels of industrial use (1%). The experimental temperature range was between 50 and - 50 degrees C. Analysis of the temperature dependence of viscoelastic processes by the equation of Williams, Landel, and Ferry provides values of fractional free volume for the temperatures covering the glass transition region. The shift factors used in the conversion of mechanical spectra into master curves were normalised at suitably different temperatures so that their temperature dependence becomes coincident. The treatment implies an iso-free-volume state and relates to changes in the monomeric friction coefficient with increasing levels of intermolecular interactions in the mixture. A free-volume related glass transition temperature was defined and manipulated markedly by introducing pectin of variable degrees of esterification to the sucrose-glucose syrup system. PMID- 11117325 TI - Isothermal and non-isothermal crystallization in amorphous sucrose and lactose at low moisture contents. AB - Differential scanning calorimetry has been used in isothermal and non-isothermal modes to provide information on the crystallization of sucrose and lactose at low water contents. Using approaches previously applied to polymer crystallization an attempt has been made to combine the isothermal and non-isothermal data into a single curve. This is achieved by the use of appropriate shift factors in the time and temperature domains. This was successful for sucrose but not for lactose. It was suggested that this was because lactose crystallizes into multiple forms whereas sucrose crystallizes in a single form. PMID- 11117324 TI - Molecular recognition of carbohydrates by a resorcinarene. Selective transport of alditols through a supported liquid membrane. AB - A supported liquid membrane (SLM) containing a resorcinarene carrier has been used for the selective transport of erythritol, threitol, ribitol and xylitol from concentrated (1.0-0.01 M) aqueous solutions. The membrane is made of a microporous polytetrafluoroethylene film impregnated with a 0.01 M solution of the carrier in CCl4. The permeabilities of the SLM for all alditols were calculated. On the basis of the flux dependence on the initial concentrations of carrier and alditol, the rate-determining step in the transport mechanism is shown to be the migration of the 1:1 carrier-carbohydrate complex in the immobilized organic phase. The flux of sugar is related to the initial concentration of alditol in the feed phase by a saturation law, which allowed the determination of the apparent diffusion coefficients and the stability constants of the resorcinarene complexes of alditols formed in the liquid membrane. PMID- 11117326 TI - Synthesis of allyl 2-O-(alpha-L-arabinofuranosyl)-6-O-(alpha-D-mannopyranosyl) beta-D-mannopyranoside, a unique plant N-glycan motif containing arabinose. AB - The synthesis of the trisaccharide allyl 2-O-(alpha-L-arabinofuranosyl)-6-O (alpha-D-mannopyranosyl)-beta-D-mannopyra-noside is reported. Stereoselective glycosylation at C-6 of a non-protected allyl beta-mannoside with the acetylated alpha-D-mannosyl bromide gave the alpha-(1 --> 6)-disaccharide as the main product and the crystalline 3,6-branched trisaccharide as minor compound. Further glycosylation of the 2,3 diol (1 --> 6) disaccharide with L-arabinofuranosyl bromide furnished a mixture of 3-O- and 2-O-alpha-L-Ara-trisaccharides from which the title compound was isolated. PMID- 11117327 TI - Immunostimulant (1 --> 3)-D-glucans from the cell wall of Cryphonectria parasitica (Murr.) Barr strain 263. AB - A (1 --> 3)-beta-D-glucan with approximately 30% of the residues having a beta-D Glc-(1 --> 6) branch is the main water-soluble component of the cell wall polysaccharide of Cryphonectria parasitica (Murr.) Barr strain 263. A (1 --> 3) glucan with both alpha and beta anomeric linkages was found in the water insoluble polysaccharide fraction. Both fractions possess immunological activity, being able to induce the production of either tumour necrosis factor alpha (TNF alpha) or nitrite (NO2-). PMID- 11117328 TI - Acid-base properties of the reaction product of sialic acid with fluorogenic reagent, 1,2-diamino-4,5-methylenedioxybenzene (DMB). AB - N-Acetylneuraminic acid (Neu5Ac) forms the highly fluorophoric quinoxalinone derivative (Q) when treated with 1,2-diamino-4,5-methylenedioxybenzene (DMB). Effects of protonation and deprotonation on the fluorescence of Q were examined at room temperature. The strong fluorescence was found to be caused by the neutral form Q but not the protonated form of its excited state [Q]* and at pH below 1 the emission was completely quenched. The deprotonated singlet form [Q-]* was a less efficient fluorescer than [Q]*. PMID- 11117329 TI - Full structure of the O-specific polysaccharide of Proteus mirabilis O24 containing 3,4-O-[(S)-1-carboxyethylidene]-D-galactose. AB - The structure of a neutral polysaccharide isolated by degradation with dilute acetic acid of the lipopolysaccharide (LPS) of P. mirabilis O24 has been determined recently [E. Literacka et al., FEBS Lett., 456 (1999) 227-231]. Further studies of this LPS using alkaline degradation and hydrolysis at pH 4.5 showed that the polysaccharide chain includes an acetal-linked pyruvic acid residue, which is removed completely during delipidation with acetic acid. A revision using 1H and 13C NMR spectroscopy and methylation analysis resulted in determination of the following full structure of the repeating unit of the O specific polysaccharide: carbohydrate sequence [see text] where D-Gal3,4(S-Pyr) is 3,4-O-[(S)-1-carboxyethylidene]-D-galactose. PMID- 11117330 TI - Structure of the O-specific polysaccharide of Mesorhizobium huakuii IFO15243T. AB - The structure of the O-specific polysaccharide isolated by mild acid hydrolysis of the lipopolysaccharide of Mesorhizobium huakuii IFO15243T was studied using methylation analysis and various one- and two-dimensional 1H and 13C NMR experiments. The O-antigen polysaccharide was found to be linear polymer constituted by a trisaccharide repeating unit of the following structure: --> 2) alpha-L-6dTalp-(1 --> 3)-alpha-L-6dTalp-(1 --> 2)-alpha-L-Rhap-(1 -->. PMID- 11117331 TI - Structure of a viscous exopolysaccharide produced by Lactobacillus helveticus K16. AB - A viscous extracellular polysaccharide produced by Lactobacillus helveticus K16 has been investigated. Sugar and methylation analysis, 1H and 13C NMR spectroscopy revealed that the polysaccharide is composed of a hexasaccharide repeating unit. The sequence of sugar residues was determined by use of two dimensional nuclear Overhauser effect spectroscopy and heteronuclear multiple bond connectivity experiments. The structure of the repeating unit of the exopolysaccharide from L. helveticus K16 is as follows: carbohydrate sequence [see text]. PMID- 11117332 TI - An NMR study of the lactonization of alpha-N-acetylneuraminyl-(2 --> 3)-lactose. AB - The composition of the products formed by treatment of commercial alpha-Neu5Ac-(2 --> 3)-beta-D-Galp-(1 --> 4)-D-Glc (3'-sialyllactose) with glacial acetic acid was investigated by 1H-13C one- and two-dimensional NMR spectroscopy and fast atom bombardment-mass spectrometry. The data confirmed that the major product of the reaction was alpha-Neu5Ac-(2 --> 3)-beta-D-Galp-(1 --> 4)-D-Glc-(1c --> 2b) lactone, which reverted to the starting material on standing in aqueous solution at ambient temperature, but for which complete NMR assignments are reported. The NMR data led to the tentative conclusion that the reaction also yielded small amounts of lactose, and alpha-Neu5Ac-(2 --> 3)-beta-D-Galp-(1 --> 4)-D-Glc-(1c - > 4b)-lactone which was stable in aqueous solution. PMID- 11117333 TI - 6-Deoxy-6-fluoro-L-ascorbic acid: crystal structure and oxidative degradation. AB - Ascorbic acid and its oxidation products have been implicated in non-enzymatic modification of proteins in aging and diseases of oxidative stress. We have studied the feasibility of using 6-deoxy-6-fluoroascorbic acid (6) for identification of ascorbic acid degradation products by 19F NMR spectroscopy. Crystals of compound 6 from nitromethane belonged to the space group P2(1) with a = 5.547(2), b = 6.769(3), c = 9.302(2) A, beta = 91.80(3) degrees and Z = 2. Atomic coordinates, bond lengths and angles, hydrogen coordinates, anisotropic and isotropic displacement parameters were similar if not identical with those of native ascorbic acid. Similarly, UV properties and oxidation kinetics by CuCl2 at different pH values were essentially identical with ascorbic acid. Using 750 MHz 19F NMR spectroscopy, five to six new fluorinated products were detected after overnight oxidation of 6 with Cu2+, suggesting that 6 may be a powerful and sensitive tool for assessment of its catabolism in vivo. PMID- 11117334 TI - Quality of life in women with urinary tract infections: is benign disease a misnomer? AB - BACKGROUND: The objective of this study was to undertake an exploratory evaluation of quality-of-life indicators for women suffering from urinary tract infections. METHODS: The RAND 36-Item Health Survey 1.0 (SF-36) was administered to 47 women with a diagnosed urinary tract infection who were being cared for in the Family Medicine Center, Student Health Services, or Urology Outpatient Clinic. A control population of 71 women was obtained from the female members of an undergraduate geography class, a community basketball league, and a local women's choir. RESULTS: All subsections of the SF-36 quality-of-life indices were significantly decreased in the subject population compared with the control population (lower score indicates lower quality of life): patient general health perception (63.3 vs 78.9, P < .001) physical functioning (76.6 vs 87.6, P = .012), role limitation owing to physical health (53.8 vs 93.0, P < .001) and emotional health (67.4 vs 88.3, P < .001), vitality (43.0 vs 64.9, P < .001), emotional well-being (64.4 vs 80.2, P < .001), pain (58.7 vs 91.5, P < .001), and social functioning (60.4 vs 90.4. P < .001). CONCLUSION: Suffering from an urinary tract infection has a detrimental influence on patient quality of life. The effect of urinary tract infections on women and their perception of quality of life have not been hitherto reported in the medical literature. The significant findings in this study call into question whether acute, non-life threatening illness should be regarded as benign. PMID- 11117335 TI - Why are antibiotics prescribed for patients with acute bronchitis? A postintervention analysis. AB - BACKGROUND: Despite the findings in controlled trials that antibiotics provide limited benefit in the treatment of acute bronchitis, physicians frequently prescribe antibiotics for acute bronchitis. The aim of this study was to determine whether certain patient or provider characteristics could predict antibiotic use for acute bronchitis in a system where antibiotic use had already been substantially reduced through quality-improvement efforts. METHODS: A retrospective chart review was performed in an academic family medicine training center that had previously instituted a quality-improvement project to reduce antibiotic prescribing for acute bronchitis. Patients who had acute bronchitis diagnosed during an 18-month period and who had no other secondary diagnosis for respiratory distress or a condition that would justify antibiotics were selected from a computerized-record database and included in the study (n = 135). Charts were reviewed to document patient symptoms, physical findings, provider and patient characteristics, and treatment. RESULTS: Thirty-five (26%) patients received antibiotics for their acute bronchitis. Adults were more likely to receive antibiotics than children (34% vs 3%, P < .001). Analysis of 20 different symptoms and physical findings showed that symptoms and signs were poor predictors of antibiotic use. Likewise, no significant differences were found based on prescribing habits of individual providers or provider level of training. CONCLUSION: In a setting where antibiotic use for acute bronchitis had been decreased through an ongoing quality-improvement effort, it did not appear that providers selectively used antibiotics for patients with certain symptoms or signs. Other factors, such as nonclinical cues, might drive antibiotic prescribing even after clinical variation is suppressed. PMID- 11117336 TI - Successful withdrawal of thyroid hormone therapy in nursing home patients. AB - BACKGROUND: Studies of community-dwelling patients have indicated that substantial numbers of patients might have had thyroid hormone therapy prescribed inappropriately and that thyroid hormone therapy in some can be discontinued without adverse effects or evidence of clinical hypothyroidism. We wanted to find out whether thyroid hormone therapy in selected nursing home patients could be withdrawn without adverse effect. METHODS: Participants for the study were drawn from four skilled nursing facilities in Connecticut. All patients on thyroid hormone therapy who resided in one of the four facilities at the time the study began were eligible if they met the inclusion criteria and gave consent to participate in the study. We measured baseline thyrotropin (TSH) levels and reduced thyroid hormone therapy by approximately onehalf if baseline TSH levels were 7 mU/L or less. If at a 1-month follow-up measurement a patient's TSH level was 7 mU/L or less, we discontinued thyroid hormone therapy. If TSH levels remained 7 mU/L or less at the next follow-up measurement 1 month later, we measured the free thyroxine (T4) level. If the free T4 level was normal, the patient remained off thyroid hormone therapy, and a final TSH value was measured after a further 2 months. RESULTS: There were 915 patients residing at the four homes at the time the study began. One hundred fifteen were on thyroid hormone therapy; 40 had elevated TSH levels in their nursing home records; and 31 refused to participate in the study. Twenty-two patients were excluded because they died or were discharged before completion of the study, had an elevated baseline TSH reading, or were taking medications that could complicate the accurate measurement of TSH. Twenty-two patients began hormone withdrawal. One patient had an increase in psychiatric symptoms during the withdrawal phase. No other adverse effects were noted. Eleven patients (50%) had the thyroid hormone therapy withdrawn successfully. CONCLUSION: Thyroid hormone therapy was successfully withdrawn from one half of the nursing home residents studied. Previous studies conducted in community-dwelling patients have shown similar findings. Many older patients began taking thyroid hormone therapy when younger either for inappropriate reasons or for what turned out to be transient hypothyroidism. If the findings of this study are generalizable for other nursing home residents, there are important implications for health and health care costs. PMID- 11117338 TI - Brown recluse spider bites. AB - BACKGROUND: Brown recluse spider bites are a serious medical problem in the southeastern United States. Although most bites are asymptomatic, envenomation can result in a constellation of systemic symptoms referred to as loxoscelism. Patients can also develop necrotic skin ulcers (necrotic arachnidism). These ulcers are often difficult to heal and can require skin grafting or amputation of the bitten appendage. METHODS: A search of the literature was performed using the search words "spider envenomation," "brown recluse spider bites," and "arachnid envenomation." RESULTS AND CONCLUSIONS: Most brown recluse spider bites are asymptomatic. All bites should be thoroughly cleansed and tetanus status updated as needed. Patients who develop systemic symptoms require hospitalization. Surgical excision of skin lesions is indicated only for lesions that have stabilized and are no longer enlarging. Steroids are indicated in bites that are associated with severe skin lesions, loxoscelism, and in small children. Dapsone should be used only in adult patients who experience necrotic arachnidism and who have been screened for glucose-6-phosphate dehydrogenase deficiency. Topical nitroglycerin can be of value in decreasing the enlargement of necrotic skin ulcers. PMID- 11117337 TI - Effects of physician supply on early detection of breast cancer. AB - BACKGROUND: There are few studies examining the effects of physician supply on health-related outcomes. We hypothesized that increasing physician supply and, in particular, increasing primary care supply would be related to earlier detection of breast cancer. METHODS: Information on incident cases of breast cancer occurring in Florida in 1994 (n = 11,740) was collected from the state cancer registry. Measures of physician supply were obtained from the 1994 AMA Physician Masterfile. The effects of physician supply on the odds of late-stage diagnosis were examined using multiple logistic regression. RESULTS: There was no relation between overall physician supply and stage of breast cancer of diagnosis. Each 10th percentile increase in primary care physician supply, however, resulted in a 4% increase in the odds of early-stage diagnosis (adjusted odds ratio = 1.04, 95% confidence interval = 1.01-1.06). CONCLUSIONS: The supply of primary care physicians was significantly associated with earlier stage of breast cancer at diagnosis. This study suggests that an appropriate balance of primary care and specialty physician supply might be an important predictor of health outcomes. PMID- 11117339 TI - When you hear hoof beats: four principles for separating zebras from horses. AB - BACKGROUND: When a patient comes to the clinic with a new complaint, the often wide array of possible causes creates uncertainty about the optimal evaluation and treatment. Selecting an approach to evaluation involves values that range from ruling out all disease processes (all zebras) regardless of cost to limiting cost by looking only for those processes that are likely (assuming all hoof beats are created by horses). Neither extreme is an optimal approach. We do not want to spend money on unnecessary tests, but we also do not want to miss a rare but potentially serious and treatable disease. METHODS: We offer four principles and their accompanying corollaries that make it possible to separate more easily hoof beats for horses from those for zebras. RESULTS AND CONCLUSIONS: By applying these principles and the accompanying corollaries, a physician can more efficiently determine the most efficient and cost-effective approach to taking care of patients. PMID- 11117340 TI - First-episode psychosis: a clinical approach. AB - BACKGROUND: Psychotic illnesses, such as schizophrenia and bipolar illness, are relatively common and clearly devastating diseases. Most scientific literature focuses on research and care of patients suffering from psychotic illnesses in the middle age-group; subsequently, the first episode or early stages of psychotic illnesses have been relatively ignored, especially the issues of early diagnosis and intervention. The purpose of this article is to highlight issues of first-episode schizophrenia for the family physician and to discuss (1) diagnosis, (2) neuropsychiatry research, (3) new medications, and (4) family issues. METHODS: To approach the issues of first-episode schizophrenia, we describe a case of a young woman who suffered her first episode of psychosis. Relevant literature related to the early stages of psychosis, including new pharmacologic treatments, is addressed. RESULTS: This report of our patient, a 19 year-old woman, illustrates the problems of a long prodromal phase of her illness, the use of medications that might have worsened her condition, and the successful use of new antipsychotic medications. Her family's issues as the patient went through this phase of her illness and recovery are reviewed. CONCLUSIONS: Patients at the outset of a psychotic illness are frequently first seen by a family physician. Familiarity with current diagnostic criteria and effectiveness of new treatments can lead to improved detection and overall outcome. PMID- 11117341 TI - Compassion and the art of family medicine: from Osler to Oprah. PMID- 11117342 TI - Is repeat testing needed for Helicobacter pylori? PMID- 11117343 TI - Iatrogenic hyponatremic seizures after routine pelvic ultrasonic imaging. PMID- 11117344 TI - Primitive neuroectodermal tumor arising from the kidney. PMID- 11117345 TI - Is there evidence that primary care physician supply influences mammography use? PMID- 11117346 TI - Antibiotic prescribing for acute bronchitis: how low can we go? PMID- 11117348 TI - Training for rural emergency care. PMID- 11117349 TI - Cancer screening rates. PMID- 11117350 TI - Future of generalism. PMID- 11117351 TI - Vaginal cuff testing. PMID- 11117352 TI - Intrachromosomal distribution of telomeric repeats in Eumops glaucinus and Euntops perotis (Molossidae, Chiroptera). AB - The location of chromosomal telomeric repeats (TTAGGG)n was investigated in two species of the Molossidae family, Eumops glaucinus and Eumops perotis. The diploid chromosome number (2n) is 40 in E. glaucinus and 48 in E. perotis and the fundamental numbers (FN) are 64 and 58, respectively. It has been suggested that the E. glaucinus karyotype has evolved from the E. perotis karyotype through Robertsonian fusion events. In the present study, the telomeric sequences were detected at the termini of chromosomes in both species. In addition, E. glaucinus also displayed telomeric repeats in centromeric and pericentromeric regions in almost all biarmed chromosomes. Conversely, in E. perotis pericentromeric signals were only observed in two biarmed chromosomes. In both E. glaucinus and E. perotis, such telomeric sequences were observed as part of the heterochromatin. The interstitial sites of telomeric sequences suggest that they are remnants of telomeres of ancestral chromosomes that participated in the fusion event. PMID- 11117354 TI - Intraspecific variation in the distribution of the interstitial telomeric (TTAGGG)n sequences in Micoureus demerarae (Marsupialia: Didelphidae). AB - The distribution of the telomeric sequence (TTAGGG)n was studied in chromosomes of Micoureus demerarae (2n = 14), a South American marsupial, by fluorescence in situ hybridization (FISH). The telomeric repeat sequence was present at both ends of all chromosomes, but also various interstitial telomeric sequences (ITS) were detected in the pericentromeric heterochromatic regions. Intraspecific differences in the number of ITS (2 to 8) were observed without intraindividual variation. The presence of telomere-like sequences in the same regions of constitutive heterochromatin suggest that these segments are not necessarily remnants of true telomeres resulting from chromosome rearrangements but could be part of the satellite DNA. PMID- 11117353 TI - Organization of the X and Y chromosomes in human, chimpanzee and mouse pachytene nuclei using molecular cytogenetics and three-dimensional confocal analyses. AB - We used multicolour fluorescence in-situ hybridization on air-dried pachytene nuclei to analyse the structural and functional domains of the sex vesicle (SV) in human, chimpanzee and mouse. The same technology associated with 3-dimensional analysis was then performed on human and mouse pachytene nuclei from cytospin preparations and tissue cryosections. The human and the chimpanzee SVs were very similar, with a consistently small size and a high degree of condensation. The mouse SV was most often seen to be large and poorly condensed, although it did undergo progressive condensation during pachynema. These results suggest that the condensation of the sex chromosomes is not a prerequisite for the formation of the mouse SV, and that a different specific mechanism could be responsible for its formation. We also found that the X and Y chromosomes are organized into two separate and non-entangled chromatin domains in the SV of the three species. In each species, telomeres of the X and Y chromosomes remain clustered in a small area of the SV, even those without a pseudoautosomal region. The possible mechanisms involved in the organization of the sex chromosomes and in SV formation are discussed. PMID- 11117355 TI - Spontaneous occurrence of a Robertsonian fusion involving chromosome 19 by single whole-arm reciprocal translocation (WART) in wild-derived house mice. AB - Chromosomal races of the house mouse (Mus musculus domesticus) bear Robertsonian (Rb) fusions, which consist of centric translocations between two non-homologous acrocentric chromosomes. The high level of diversity of these fusions in house mice is generated by de-novo formation of Rb fusions and subsequent whole-arm reciprocal exchanges (WARTs). This paper describes the spontaneous occurrence of a new Rb fusion, Rb(4.19), in progeny of wild-derived house mice segregating for Rb(4.12). The chromosomal mutation was traced to a female which exhibited germline and somatic mosaicism indicating an early embryonic origin of the mutation. FISH analysis of centromerically-located ribosomal genes suggested that no modification was observed on chromosomes 12 and 19 prior to or following the occurrence of Rb(4.19). Distribution of telomeric sequences showed that both Rb fusions lacked telomeres in their centromeric regions. It is argued that this spontaneous mutation most likely originated by single whole-arm reciprocal translocation (WART) between Rb(4.12) and an acrocentric chromosome 19, resulting in Rb(4.19) and a neo-acrocentric chromosome 12. Sequences required for centromeric function and proximal telomeres would have been transferred to the neo-chromosome 12 from chromosome 19 during the translocation. The existence of such WARTs which generate derived acrocentric chromosomes has several implications for chromosomal evolution in house mice. PMID- 11117356 TI - A biodiversity approach in the neotropical erythrinidae fish, Hoplias malabaricus. Karyotypic survey, geographic distribution of cytotypes and cytotaxonomic considerations. AB - Hoplias malabaricus, a widely distributed neotropical freshwater fish, shows a conspicuous karyotypic diversification. An overview of this diversity is presented here comprising several Brazilian populations, and some others from Argentina, Uruguay and Surinam. Seven general cytotypes are clearly identified on the basis of their diploid number (2n = 39 to 2n = 42), chromosomal morphology and sex chromosome systems, which can be clustered into two major karyotypic groups. This clustering suggests that karyotype structure would be more informative than the diploid number regarding cytotype relationships in this fish group. While some cytotypes show a wide geographical distribution, some others appear to be endemic to specific hydrographic basins. Sympatric cytotypes can occur without detection of hybrid forms; this situation points to a lack of gene flow, a fact that is also reinforced by studies with genomic markers. The karyotypic data support the view that the nominal taxon H. malabaricus corresponds to a species complex comprising distinct evolutionary units, each with well-established chromosomal differences. PMID- 11117357 TI - The chromosome complement of Olea europaea L.: characterization by differential staining of the chromatin and in-situ hybridization of highly repeated DNA sequences. AB - The chromosome complement of olive (Olea europaea L.) has been characterized by differential staining of the chromatin and chromosomal localization of highly repeated DNA sequences and ribosomal cistrons. DAPI staining produces different sized positive bands in various locations on all the chromosomes. By combining this band pattern with the results obtained from cytological hybridization of OeTaq80, OeTaq178, and OeGEM86 DNA tandem repeats, most of the pairs can be distinguished from each other, in spite of the large number of chromosomes (2n = 46), their small size and similar morphology. Different tandem-repeated DNA sequences may be contained into single heterochromatic chromosome regions, even though there are regions where repeats of only one family are present. OeTaq80- and OeGEM86-related DNA sequences are rather specific to the heterochromatin at the chromosome ends, while most sequences related to the longer OeTaq178 probe are confined to interstitial heterochromatin. Some exceptions suggest that major chromosomal rearrangements occurred during genome evolution. Polymorphism, which may differentiate olive cultivars, was observed within chromosome pairs I, V, and VII. PMID- 11117358 TI - Centromeric heterochromatin in the cattle rob(1;29) translocation: alpha satellite I sequences, in-situ MspI digestion patterns, chromomycin staining and C-bands. AB - The centromeric regions and alpha-satellite I sequence were studied on chromosomes 1, 29 and the rob(1;29) translocation in a Portuguese breed of cattle, Barrosa, carrying the translocation. Rob(1;29) centromeric regions showed heterochromatic bands with propidium iodide but, unlike the acrocentric autosomes, no strong centromeric bands were revealed with chromomycin A3. An alpha-satellite I sequence was not found at the centromeres of the X, Y and rob(1;29) chromosomes in the breed, although it was present at the centromeres of all acrocentric chromosomes including 1 and 29. Restriction enzyme banding with MspI revealed polymorphisms between different rob(1;29) chromosomes in both centromeric and intercalary regions. The data show that the centromeric region of the rob(1;29) chromosome has lost the alpha-satellite I sequences, while retaining other heterochromatin, and suggest that this important and widespread translocation has occurred multiple times. PMID- 11117359 TI - Frequency increase and mitotic stabilization of a B chromosome in the fish Prochilodus lineatus. AB - Six populations of the fish Prochilodus lineatus were analysed for B chromosome frequency. A study of spermatogenesis revealed the absence of B accumulation during the stages analysed. In one of the populations, from the Mogi-Guacu river where samples have been analysed over a ten-year period, B chromosome frequency doubled between 1979-80 and 1987 89, whereas no additional changes were noticed in samples collected in 1991-92. The analysis of B chromosome mitotic instability, manifested by intraindividual variation in B chromosome number, indicated a very significant decrease during this time period. This suggests that, in the 1980s, this population was in the final stage of B chromosome invasion, and that there was a possible causal relationship between B mitotic instability and the accumulation mechanism that caused its frequency increase. Mitotic stabilization might thus be a way by which a mitotically unstable B chromosome may become neutralized. PMID- 11117360 TI - Stable methylation patterns in interspecific antelope hybrids and the characterization and localization of a satellite fraction in the Alcelaphini and Hippotragini. AB - Conflicting data has recently appeared concerning altered methylation patterns in interspecific mammalian hybrids and the potential this may hold for driving karyotypic evolution. We report no detectable methylation difference in the genomic DNA of different interspecific F1 antelope hybrids (family Bovidae) and their parent species using the methylation-sensitive enzyme HpaII and its methylation insensitive isoschizomer MspI. However, both enzymes released a tandemly repeated satellite array. Characterization of the repeat using Southern blotting and a combination of sequencing, fluorescence in-situ hybridization (FISH) and C-banding, shows some similarity in the family of repeats between the hybridizing antelope species groups, and that the satellite is localized in the centromeric C-band positive regions of the chromosomes. Moreover, although there is little meaningful sequence homology with the well characterized bovine 1.715 satellite DNA, there is 86% sequence similarity with the sheep/goat satellite I, suggesting that they are related and are likely to have originated and evolved separately from the bovine unit. PMID- 11117361 TI - Replication timing of homologous alpha-satellite DNA in Roberts syndrome. AB - Roberts syndrome (RS) is associated with a characteristic constitutive heterochromatin anomaly, namely, at metaphase the centromeres and heterochromatic segments appear split. In addition to this cytogenetic phenomenon, known as the RS effect, several other cytological features, especially affecting mitotic chromosome disjunction, are also observed. Applying FISH to interphase nuclei, we investigated the replication patterns of homologous alphoid centromeric DNA of chromosomes 9, 11, 16 and 17 in three patients showing the RS effect and in four normal individuals. A tendency for homologous centromeres to replicate asynchronously was observed in RS patients. This tendency was more evident in chromosomes 9 and 16, with large heterochromatic blocks and particularly subject to RS effect. This asynchrony could reflect a more generalized alteration in repetitive DNA replication timing that, in turn, would prevent the establishment of proper cohesion between sister chromatid heterochromatin, leading to the RS effect. PMID- 11117362 TI - Long-range analysis of the centromeric region of Drosophila melanogaster chromosome 3. PMID- 11117363 TI - Human activity-regulated cytoskeleton-associated gene (ARC) maps to chromosome 8q24. PMID- 11117364 TI - Human HRK gene maps to position 12q13.1. PMID- 11117365 TI - Brown trout 5S rDNA maps to chromosome 38. PMID- 11117366 TI - Ascorbic acid improves postischemic vasodilatation impaired by infusion of soybean oil into canine iliac artery. AB - This study was conducted to (a) assess postischemic vasodilatation by changes in the vascular cross-sectional area using simultaneous intravascular two dimensional and Doppler ultrasound before and after the infusion of Intralipid (Pharmacia & Upjohn, Peapack, NJ, U.S.A.); (b) evaluate how antioxidant ascorbic acid modifies the effects of Intralipid on postischemic vasodilatation: and (c) clarify the changes in plasma nitrite and nitrate (NOx-) levels after the infusion of Intralipid with and without ascorbic acid. Twenty-eight mongrel dogs were used to measure for vascular cross-sectional area and average instantaneous peak velocity in the iliac arteries after the 5-min occlusion of the arteries. Postischemic vasodilatation was impaired after the infusion of Intralipid (20%, 2 ml/kg) and this impaired response was reversed by the co-administration of ascorbic acid (30 mg/kg). NG-monomethyl-L-arginine completely abolished postischemic vasodilatation. Plasma NOx levels were significantly reduced after the infusion of Intralipid compared with baseline (11.6+/-0.4 vs. 12.9+/-0.3 microM, p = 0.025) and after infusion of Intralipid with ascorbic acid compared with baseline (11.8+/-0.5 vs. 13.1+/-0.4 microM, p = 0.047). We concluded that ascorbic acid reverses the endothelial dysfunction induced by Intralipid without increasing plasma NOx- levels and that deactivation of nitric oxide by oxidative stress is a primary contributor to Intralipid-induced impaired vasodilation. PMID- 11117367 TI - Cyclosporine A and cremophor EL induce contractions of human saphenous vein: involvement of thromboxane A2 receptor-dependent pathway. AB - Chronic treatment with Sandimmune (cyclosporine A [CsA] dissolved in Cremophor EL [CrEL]) is often associated with hypertension and nephrotoxicity. The aims of the present study were to assess the effect of Sandimmune and its two main components (CsA and CrEL) on human saphenous veins and to study the underlying mechanism of their contractile responses. In organ bath, concentration-response curves for Sandimmune (36 ng/ml-120 microg/ml of CsA). CsA (36 ng/ml-120 microg/ml), or CrEL (2.4 microg/ml-8 mg/ml) were elicited in the presence of a thromboxane A2 (TXA2) receptor antagonist (GR32191, 0.3 microM), a cyclooxygenase inhibitor (indomethacin, 1 microM), a 5-lipoxygenase inhibitor (AA861, 10 microM), or their respective vehicles. In addition, the production of TXA2 after CsA challenge was assessed by enzyme immunoassay. Sandimmune, CsA, and CrEL induced concentration dependent contractions on human saphenous veins. In terms of potency, CsA was a more potent vasoconstrictor agent than CrEL (EC50 values: 11.9+/-3.7 microg/ml (CsA, n = 12) vs. 1.2+/-0.4 mg/ml (CrEL, n = 16), p < 0.05). In contrast, in terms of efficacy, CrEL induced greater contractions than CsA (Emax (% of KCl 90 mM-induced contraction): 98.1+/-16.1% (CrEL, n = 16) vs. 17.0+/-4.3% (CsA, n = 12) p < 0.05). Pretreatment with GR32191 significantly reduced by 85% and 56% the contractions elicited by CsA and CrEL, respectively, whereas indomethacin had no effect. Finally, CsA (12 and 120 microg/ml) failed to stimulate TXA2 production. These in vitro data suggest that Sandimmune-induced contractions on human vascular smooth muscle appear to be mediated by CsA in the therapeutic ranges of doses and by both CsA and CrEL, which, in supratherapeutic doses, acted through a TXA2 receptor-dependent pathway. PMID- 11117368 TI - No evidence for functional involvement of 5-HT2B receptors in serotonin-induced vasodilatation in the human forearm. AB - The receptor involved in the serotonin (5-hydroxytryptamine [5-HT])-induced vasodilatation in the human forearm has not yet been identified. Experimental data point to the 5-HT2B receptor located on the endothelium. RS-127445 (2-amino 4-(4-fluoronaphthyl-1-yl)-6-isopropylpyrimidine) is a novel potent and selective 5-HT2B receptor antagonist. The effect of oral RS-127445 (500 mg) on 5-HT-induced vasodilatation was studied in a double-blind, randomized, placebo-controlled, crossover study in six healthy volunteers. On each study day 5-HT (0.5 ng/kg/min) was infused into the brachial artery for 8 min, before drug administration and at intervals of 20, 65, 110, 230, and 470 min after oral ingestion. At each infusion, plasma samples for study drug assay were taken and forearm blood flow was assessed using venous occlusion plethysmography. Although (log) drug concentrations exceeded pKi, there was no correlation between RS-127445 concentrations and 5-HT-induced vasodilatation. 5-HT-induced vasodilatation did not differ between treatments and time points. It appears that there is no functional involvement of 5-HT2B receptors in 5-HT-mediated vasodilatation in the human forearm. PMID- 11117369 TI - Sympathoinhibitory and depressor responses to long-term infusion of nifedipine in spontaneously hypertensive rats on high-salt diet. AB - Short-term (by hour) intracerebroventricular (i.c.v.) or i.v. infusion of nifedipine at low rates evokes parallel decreases in renal sympathetic nerve activity (RSNA) and blood pressure (BP) in spontaneously hypertensive rats (SHR). In the present study, effects of long-term administration of nifedipine on BP and control of sympathetic tone were examined in SHR on a high-salt (8%) diet. From 6 to 8 weeks of age, for 2 weeks concomitant with taking a high-salt diet, rats were also treated with subcutaneous infusion of nifedipine at 10, 50, or 100 microg/kg/h or vehicle solvent as control using osmotic minipumps. At the end of the 2-week treatment period, mean arterial pressure (MAP), heart rate (HR), and RSNA at rest and in response to air-jet stress, i.c.v. injection of the alpha adrenoceptor agonist guanabenz (25 microg), and i.v. injection of the ganglionic blocker hexamethonium were recorded in conscious rats. In rats on nifedipine 50 or 100 microg/kg/h, resting MAP was significantly lower (136+/-4 or 130+/-4 vs. 145+/-2 mm Hg in control rats, p < 0.05 for both), the sympathoinhibitory and depressor responses to i.c.v. guanabenz were significantly decreased, and the absolute decreases in MAP in response to i.v. injection of hexamethonium were significantly smaller. Sympathoexcitatory and pressor responses to air-jet stress, however, were not affected by nifedipine. Infusion of nifedipine at the three rates for 2 weeks caused concentrations of plasma nifedipine in a dose related manner. Nifedipine was not detected in tissues of rats treated with 10 microg/kg/h nifedipine but was present in brain and other tissues of rats treated with nifedipine at the two higher rates. Thus in SHR on high-salt intake long term treatment with nifedipine at 50 or 100 microg/kg/h decreased resting BP and the sympathetic component in BP control. In addition to possible peripheral effects, long-term administration of nifedipine may also act centrally to decrease sympathetic activity and BP, likely by increasing activity in central pathways involving sympathoinhibition, but not in pathways involving sympathoexcitation as evaluated by air-stress. PMID- 11117370 TI - Modification of stretch-induced shortening of repolarization by streptomycin in the isolated rabbit heart. AB - The exact mechanism of mechano-electrical feedback and stretch-induced arrhythmias is unknown, but the role of stretch-activated ion channels and specific calcium channels has been proposed. The aim of the present study was to test the hypothesis that stretch-activated ion channels and not calcium channels contribute to stretch-related alterations of repolarization and that these effects can be neutralized by stretch-activated channel block. We studied the interaction of acute ventricular dilatation and the stretch-activated channel blocker streptomycin and the specific calcium channel blocker verapamil in an isolated retrogradely perfused rabbit heart model in which the left ventricular size is modified by abruptly changing the volume of a fluid-filled balloon placed in the left ventricle. Acute ventricular dilatation led to a rate-dependent decrease in repolarization. The mean effective refractory period (ERP) and monophasic action potential duration (MAP90) for cycle lengths between 300 and 1,000 ms decreased from 174.2+/-9 ms and 178.9+/-7 ms to 161.6+/-11 ms and 169.7+/-5 ms, respectively. Streptomycin (80 microM) inhibited this stretch related shortening of repolarization (ERP: 175.4+/-8 ms; MAP90: 179.7+/-8 ms, p < 0.05) but had almost no effect on already dilated ventricles. Counteraction of the observed electrophysiologic changes could only be achieved by increasing the streptomycin concentration to 200 microM. Streptomycin nearly completely suppressed stretch-related ectopic ventricular complexes. In contrast, verapamil (1 microM) had no effect on stretch-related changes in repolarization and stretch induced arrhythmias. The present study indirectly implicates stretch-activated ion channels in the genesis of stretch-related changes in repolarization and arrhythmias. The electrophysiologic changes after ventricular dilatation to a degree that increases left ventricular pressure in a clinically relevant range can be influenced by the stretch-activated channel blocker streptomycin but not by specific calcium channel block. This may have clinically important implications for the development of new antiarrhythmic drugs. PMID- 11117371 TI - Intrapericardial beta-adrenergic blockade with esmolol exerts a potent antitachycardic effect without depressing contractility. AB - Hyperadrenergic states of various etiologies can contribute to tachycardias. Systemic beta-adrenergic blockade suppresses sinus tachycardia but may adversely affect arterial blood pressure and contractility, because the drug gains access to myocardial cells as well as to the sinoatrial node. We examined whether intrapericardial beta-adrenergic blockade with esmolol could suppress tachycardia without reducing contractility as a result of limited drug diffusion, which would be sufficient to penetrate the superficial sinoatrial node but not the deeper myocardial layers. In five anesthetized pigs, we provoked a reflex heart rate increase of 50 beats/min with hemorrhage. The rapidly acting beta-adrenergic blocking agent esmolol (1 mg/kg) was administered intrapericardially using a new percutaneous transatrial access method and a catheter system that can be rapidly and safely introduced. Esmolol equivalently suppressed hemorrhage-induced sinus tachycardia when administered intrapericardially (from 192 to 158 beats/min at 5 min, p < 0.05) or intravenously (from 177 to 151 beats/min at 1 min, p < 0.05). The antitachycardic effect of intrapericardial esmolol was prolonged compared with intravenous esmolol (10 min vs. 3 min, p < 0.05). Intrapericardial esmolol did not affect blood pressure or left ventricular dP/dt max, an index of contractility, whereas intravenous esmolol decreased blood pressure at 1 min for 2 min (p < 0.05) and simultaneously decreased left ventricular dP/dt max at 1 min for < 2 min (p < 0.05). Intrapericardial esmolol suppresses adrenergically induced sinus tachycardia without decreasing contractility or blood pressure. The transatrial approach for intrapericardial delivery of certain 1-adrenergic blocking agents could be employed to control tachycardias in emergency care and surgical settings in patients with impaired cardiac contractility and propensity to hypotension. PMID- 11117372 TI - Nonselective I(Kr)-blockers do not induce torsades de pointes in the anesthetized rabbit during alpha1-adrenoceptor stimulation. AB - Selective I(Kr)- (the rapid component of the delayed rectifier potassium current) blockers are known to induce torsades de pointes (TdPs) in anesthetized rabbits during alpha1-adrenoreceptor stimulation. However, effects of nonselective I(Kr) blockers, which produce TdPs in other animal models and in humans, are not known in this model. We examined two nonselective I(Kr)-blockers (quinidine, 1.25 mg/kg/min i.v [n = 7]; and terfenadine, 0.31 mg/kg/min i.v. [n = 7]) for their effects on electrocardiographic parameters and on incidence of cardiac arrhythmias in anesthetized rabbits during alpha1-adrenoceptor stimulation with methoxamine. We compared the drugs with two highly selective I(Kr)-blockers (dofetilide, 0.04 mg/kg/min i.v. [n = 7]; and clofilium, 0.08 mg/kg/min i.v. [n = 6]). Polymorphic ventricular tachycardia or TdPs were induced by dofetilide and clofilium at mean doses > or =0.33 mg/kg and 0.4 mg/kg i.v., in all animals tested (vs. none in solvent; p < 0.05). TdPs usually developed into ventricular fibrillation and developed after prolongation of QT/JT interval and of QT dispersion. Terfenadine and quinidine significantly increased PQ, QT, and QTc interval and largely increased QRS duration and QT dispersion. These compounds elicited intraventricular conduction defects and cardiac arrest, due to asystole, in all animals tested (vs. 0% in solvent; p < 0.05). Interestingly, these two nonselective I(Kr)-blockers did not produce TdPs or ventricular fibrillation in any animals tested. Our results thus indicate that selective I(Kr)-blockers elicit TdPs, whereas nonselective I(Kr)-blockers do not induce this type of arrhythmia in this rabbit model. Consequently, it should be noted that this rabbit model is not always useful to evaluate nonselective I(Kr)-blocker-induced TdPs and QT interval and QT dispersion, rather than TdPs, are also important indicators for drug-induced cardiac arrhythmias. PMID- 11117373 TI - Biphasic response to histamine in rabbit penile dorsal artery. AB - The effects of specific histamine agonists and antagonists on isolated rabbit penile dorsal artery segments were explored using in vitro isometric techniques. Histamine caused the constriction of both precontracted and resting segments. In precontracted arterial rings treated with the H1 receptor antagonist mepyramine, histamine evoked a vasodilatation, followed by contraction at higher concentrations. The vasoconstrictor effect of histamine and the H1 receptor agonist, 2-pyridylethylamine (PEA) on preparations under conditions of basal tone, was competitively antagonized by mepyramine (10(-9)-10(-8) M). The relaxant effect of histamine, unmasked by mepyramine, was abolished by cimetidine. Dimaprit, the H2 receptor agonist, provoked a relaxation of precontracted segments that was also competitively inhibited by cimetidine (10(-6)-10(-5) M). Selective H3 receptor activation with the agonist (R)alpha-methylhistamine (10( 10)-10(-4) M) produced no effect in penile dorsal artery. The biphasic response to histamine was unaffected by endothelium removal or the nitric oxide inhibitor NG-nitro-L-arginine methyl ester (L-NAME) (3 x 10(-4) M) and its precursor, L arginine (3 x 10(-4) M). Similarly, the cyclooxygenase inhibitor, indomethacin (3 x 10(-6) M) and a combination of Ca2+-activated K+ channel blockers apamin (5 x 10(-7) M) and charybdotoxin (10(-7) M) showed no effect on the histamine-induced relaxation or contraction. In conclusion, contraction, the predominant effect of histamine, is mediated by the activation of H1 receptors that mask the relaxant effect brought about by H2 receptors. Both these effects appear to be mediated by direct action on the smooth muscle, with no participation of nitric oxide or cyclooxygenase products or Ca2+-activated K+ channels. PMID- 11117374 TI - Effects of long-term treatment with verapamil on left ventricular function and myocardial blood flow in patients with dilated cardiomyopathy without overt heart failure. AB - Myocardial blood flow (MBF) abnormalities are present in early stage dilated cardiomyopathy (DCM) and have been attributed to coronary microvascular abnormalities. The favorable effects of verapamil on coronary microcirculation might indicate its use in early stage DCM. We assessed the safety of long-term combination therapy of verapamil and enalapril and its effects on both left ventricular function and myocardial perfusion compared with enalapril alone in 18 patients with DCM (15 men, 3 women; mean age, 50+/-9 years) without overt heart failure (NYHA class I-II). At baseline and after 6 months of randomized treatment with either enalapril (10-20 mg) (nine patients, group 1) or enalapril (10-20 mg) and verapamil (120-240 mg) (nine patients, group 2), left ventricular function was assessed at rest, during handgrip, and during bicycle exercise by equilibrium radionuclide angiography. Mean MBF was measured at rest and after dipyridamole by positron emission tomography (PET) and 13N-ammonia as a flow tracer. At baseline, the two groups had reduced left ventricular ejection fraction at rest, which was further impaired during isometric exercise, but increased at peak bicycle exercise. MBF was similarly reduced in the two groups at rest and during dipyridamole. During treatment, no adverse events occurred in either group. After 6 months there was no significant difference in the main study variables either between the two groups or within each group before and after treatment. Long-term combination therapy with verapamil and enalapril is safe in patients with DCM without overt heart failure. Despite no favorable effect on myocardial perfusion, combined treatment prevented deterioration of left ventricular function, similarly to enalapril alone. PMID- 11117375 TI - Decreased sensitivity to nitric oxide in the aorta of severely hypercholesterolemic apolipoprotein E-deficient mice. AB - A normal response to nitric oxide donors has been cited as evidence that impaired endothelium-dependent vasodilation during hypercholesterolemia is due to decreased synthesis of nitric oxide. This tenet was examined by determining responses to nitric oxide gas as well as to acetylcholine and sodium nitroprusside in the isolated aorta of apolipoprotein E-deficient mice fed normal or Western-type cholesterol-rich diet until 21 or 35 weeks of age. In mice fed normal chow, relaxation to all agents remained comparable to that obtained in wild-type mice. In mice fed Western diet, the relaxation to acetylcholine as well as to nitric oxide was decreased at 35 weeks of age. At 21 weeks of age, decreased sensitivity to nitric oxide was observed despite a normal response to acetylcholine. The response to sodium nitroprusside was normal in all groups. A decrease in aortic superoxide dismutase activity as well as an increase in aortic superoxide anion generated in the presence of NADH as measured by lucigenin chemiluminescence was observed in the group fed Western diet at 35 weeks. This provides evidence that altered superoxide anion could contribute to the deterioration in nitric oxide sensitivity that underlies the impaired endothelium dependent relaxation. These data indicate that decreased sensitivity to nitric oxide may contribute to the development of impaired endothelium-dependent relaxation in hypercholesterolemia. The response to sodium nitroprusside appears not to reflect the decreased sensitivity of vascular smooth muscle to authentic nitric oxide. PMID- 11117376 TI - Endothelin-1-induced contraction of mesenteric small arteries is mediated by ryanodine receptor Ca2+ channels and cyclic ADP-ribose. AB - Contraction of vascular smooth muscle by endothelin-1 is dependent on extracellular and intracellular Ca2+. However, the role of ryanodine-sensitive Ca2+ stores in endothelin-1-induced contraction is unknown. Vascular contraction was measured in mesenteric small arteries (200-300 microm intraluminal diameter) isolated from Sprague-Dawley rats and maintained at a constant intraluminal pressure of 40 mm Hg. The presence of functional ryanodine receptor Ca2+ release channels (RyRC) was demonstrated by the finding that ryanodine (10 microM), which locks the RyRC in a subconductance state, produced significant contraction of small arteries in the presence of 15 mM KCl. This effect was inhibited by dantrolene (10 microM), a RyRC inhibitor. Dantrolene significantly reduced the ET(A) receptor-mediated contraction to endothelin-1 (10(-11)-10(-9) M). The ability of dantrolene to reverse contraction induced by endothelin-1 was also determined. Dantrolene (1-10 microM) produced concentration-dependent relaxation of vessels precontracted to 38+/-3% of resting diameter with endothelin-1 but had no effect in vessels precontracted to a similar degree with phenylephrine or KCl. Because activation of RyRC may be dependent on production of cyclic ADP-ribose, the effect of nicotinamide (2 mM), an inhibitor of ADP-ribosyl cyclase, on contraction to endothelin-1 was determined. Nicotinamide had an inhibitory effect similar to that produced by dantrolene. A combination of nicotinamide and dantrolene had no greater effect than either agent alone, suggesting a common pathway for cyclic ADP-ribose and RyRC. In summary, endothelin-1 induces contraction of small mesenteric arteries through ET(A) receptor-mediated production of cyclic ADP-ribose and activation of RyRC. PMID- 11117377 TI - Vasodilator responses of coronary conduit and resistance arteries to continuous nitroglycerin infusion in humans: a Doppler guide wire study. AB - To examine the responses of coronary conduit and resistance arteries to the continuous i.v. administration of nitroglycerin in 15 patients with atypical chest pain, we measured coronary blood flow velocity in the left anterior descending coronary artery using a Doppler guide wire and the lumen diameter and cross-sectional area by quantitative coronary angiography. Systolic flow, diastolic flow, total coronary flow, and coronary vascular resistance were calculated. Stepwise increases in dose of nitroglycerin resulted in significant dose-dependent decrease in mean aortic pressure (p < 0.01) and increase in lumen diameter (p < 0.05). After nitroglycerin administration of 0.5 microg/kg/min, systolic flow decreased significantly by 89.9+/-15.7% (p < 0.01), and diastolic flow increased significantly by 74.2+/-37.1% (p < 0.05). Total coronary flow did not change significantly with the various doses of nitroglycerin. However, coronary vascular resistance decreased significantly at concentrations greater than 0.5 microg/kg/min nitroglycerin. Continuous nitroglycerin infusion did not reduce either diastolic or total coronary blood flow despite a significant reduction in coronary perfusion pressure. These results indicate that subendocardial blood flow might be maintained during continuous i.v. infusion of nitroglycerin within the clinical dose range. PMID- 11117378 TI - tPA, but not uPA, significantly affects antithrombotic therapy by a glycoprotein IIb/IIIa antagonist, but not by a factor Xa inhibitor. AB - To define the interaction of fibrinolytic components with platelets or coagulation factors on thrombus formation, we investigated mouse deficient in tissue plasminogen activator (tPA -/-) or urokinase plasminogen activator (uPA -/ ) and in their wild-type control (tPA +/+, uPA +/+). A thrombus was induced in the murine carotid artery using photochemical reaction. Blood flow was monitored and the time needed before the vessel became completely obstructed was within 12 min in all types of mice. When DX-9065a, a selective factor Xa inhibitor, or GR144053, a platelet glycoprotein (GP) complex IIb/IIIa antagonist was applied, the time required to occlusion was prolonged in a dose-dependent manner in all types of mice. When a factor Xa inhibitor was injected in tPA -/- mice, the estimated ED50 was not changed. However, when GR144053 was injected in tPA -/- mice, the most significant changes were observed: the estimated ED51 was 19.6 times higher than the one in tPA +/+ mice. Platelet aggregation, hemostasis tests, and bleeding times were not significantly different among the different types of mice. In conclusion, the antithrombotic effect of platelet inhibition by a GPIIb/IIIa antagonist, is severely affected by the absence or presence of tPA production. On the contrary, the inhibition of factor Xa shows a stable antithrombotic effect with or without tPA. Thus the lack of tPA, but not of uPA, significantly affects antithrombotic efficacy. PMID- 11117379 TI - Coronary vasomotor dysfunction in the cardiac allograft: impact of different immunosuppressive regimens. AB - Immunosuppression may have an important impact on early graft coronary endothelial injury. We investigated functional and morphologic coronary alterations, myocardial expression, and cardiac release of possible mediators of allograft vasculopathy within 6 months after cardiac transplantation with respect to different immunosuppressive regimens. Epicardial and microvascular endothelium dependent and endothelium-independent vasomotor function and epicardial intimal thickening were measured in 8 transplant recipients treated with cyclosporin A (CyA), azathioprine, and prednisone (group 1), 9 transplant recipients treated with tacrolimus (TKL), azathioprine, and prednisone (group 2), and 14 patients treated with TKL, mycophenolate mofetil (MMF), and prednisone (group 3). The gene expressions of inducible and endothelial nitric oxide synthase (iNOS and eNOS), endothelin-1, prostacyclinsynthase, and thromboxansynthase were analyzed in endomyocardial biopsy specimens using semiquantitative reverse transcription polymerase chain reaction. Transcardiac cytokine release, endothelin-1, and nitrate-release were determined from plasma samples. Epicardial endothelial dysfunction (vasoconstriction to acetylcholine > 10%) and microvascular smooth muscle cell dysfunction (flow velocity increase to adenosine and nifedipine < 2.0) were enhanced in heart transplant recipients immunosuppressed with TKL, azathioprine, and prednisone. The prevalence of epicardial dysfunction was 78% in group 2 versus 44% and 46% in group 1 and 3 (p < 0.05), respectively. The prevalence of microvascular dysfunction was 56% in group 2 versus 13% and 7% in group 1 and 3 (p < 0.02), respectively. Coronary vasomotor dysfunction was associated with increased myocardial iNOS expression (p < 0.05), decreased eNOS expression (p < 0.05), and enhanced cardiac immunoreactive interleukin-6 (p < 0.01). Coronary intimal thickening was not different between the groups. The combination of TKL and MMF appears to be superior to TKL and azathioprine (and comparable to CyA and azathioprine) concerning preservation of early coronary vasomotor function, eNOS expression, iNOS suppression as well as cardiac interleukin-6 release. This may have an important impact on subsequent development of transplant coronary atherosclerosis. PMID- 11117380 TI - Role of sodium channels in ventricular fibrillation: a study in nonischemic isolated hearts. AB - Because the role of sodium channels in the initiation and maintenance of VF is not fully elucidated, we studied the significance of sodium channel activity in VF using sodium channel blockers. In nonischemic isolated feline hearts, the following electrophysiologic parameters were measured before and after application of tetrodotoxin (5 x 10(-7) M, n = 6) or lidocaine (1 x 10(-5) M, n = 8): (a) during pacing, epicardial conduction time; refractoriness; the fastest rate for 1:1 pacing/response capture, and all tissue resistivity, indirectly reflecting intercellular electrical resistance; (b) during 8 min of electrically induced tachyarrhythmias, all tissue resistivity; peak frequency (to measure average frequency based on fast-Fourier transformation analysis); and normalized entropy (to measure the degree of arrhythmia organization). In nonischemic isolated rabbit hearts (n = 4), three-dimensional mapping was performed before and after application of lidocaine (1 x 10(-5) M). In feline hearts, lidocaine and tetrodotoxin application resulted in: (a) more spontaneous arrhythmia termination (63-67%) than in nontreated hearts (7%); (b) transformation from mainly VF into ventricular tachycardia with increased organization; and (c) prolongation of conduction time (155-248%) (p < 0.01 for all parameters). The ventricular refractory period was slightly prolonged by tetrodotoxin in the right ventricle and exhibited rate-dependent shortening in control and with lidocaine. Tetrodotoxin and lidocaine reduced the pacing rate for 1:1 pacing/response capture, and all tissue resistivity was not significantly affected. Peak frequency was decreased by tetrodotoxin and lidocaine mainly in the left ventricle (p < 0.01). In nontreated left ventricles, peak frequency was increased over time but was attenuated by lidocaine. In isolated rabbit hearts, several simultaneous wave fronts were detected during VF in nontreated hearts and were reduced to only one or two major wavefronts after application of lidocaine. Suppression of sodium channel activity that primarily slowed conduction time and had little or no effect on ventricular refractory period and all tissue resistivity resulted in less stable and more organized arrhythmias and reduced tachyarrhythmia rate compared with nontreated hearts. These results suggest an active role for sodium channels in the maintenance of ventricular fibrillation. PMID- 11117381 TI - Effect of perhexiline and oxfenicine on myocardial function and metabolism during low-flow ischemia/reperfusion in the isolated rat heart. AB - Perhexiline is a potent prophylactic anti-anginal agent that has been shown to inhibit myocardial utilization of long-chain fatty acids and to inhibit the mitochondrial enzyme carnitine palmitoyltransferase (CPT)-1. We compared the hemodynamic and biochemical effects of perhexiline (0.5 and 2.0 microM) and of another CPT-1 inhibitor, oxfenicine (0.5 mM), in Langendorff-perfused rat hearts subjected to 60 min of low-flow ischemia (95% flow reduction) followed by 30 min of reperfusion. Both perhexiline (2 microM only) and oxfenicine attenuated (p < 0.003, p < 0.0002, respectively) increases in diastolic tension during ischemia, without significant effects on developed tension, or on cardiac function during reperfusion. Myocardial concentrations of long-chain acylcarnitines (LCAC), products of CPT-1 action, were decreased (p < 0.05) by oxfenicine, unaffected by 2 microM perhexiline, and increased slightly by 0.5 microM perhexiline. Perhexiline, but not the active metabolite of oxfenicine, also inhibited cardiac CPT-2 with similar IC50 and Emax, although lower Hill slope, compared with CPT-1. Oxfenicine, but not perhexiline, reduced concentrations of the endogenous CPT-1 inhibitor, malonyl-CoA. Perhexiline, but not oxfenicine, inhibited myocardial release of lactate during normal flow. We conclude that (a) perhexiline protects against diastolic dysfunction during ischemia in this model, independent of major changes in LCAC accumulation and (b) this may result from simultaneous effects of perhexiline on myocardial CPT-1 and CPT-2. PMID- 11117382 TI - Inhibitory effects of dronedarone on muscarinic K+ current in guinea pig atrial cells. AB - Dronedarone (SR33589), an amiodarone-like noniodinated antiarrhythmic agent, is undergoing clinical trials in atrial fibrillation. Because vagal activation plays a role in the pathophysiology of supraventricular arrhythmias, we have assessed the ability of dronedarone (0.01, 0.1, and 1 microM), compared with amiodarone (0.1, 1, and 10 microM) to inhibit the muscarinic acetylcholine receptor-operated K+ current (I(K(ACh))) in single cells isolated from guinea pig atria (patch clamp technique). I(K(ACh)) was activated by extracellular application of carbachol (10 microM) or by intracellular loading with GTP-gamma-S (100 microM). Dronedarone and amiodarone reduced the carbachol-induced I(K(ACh)) with an IC50 (concentration required for 50% inhibition) slightly above 10 nM and 1 microM, respectively. Dronedarone also inhibited the GTP-gamma-S induced K+ current by 28% and 58% at 0.01 and 0.1 microM, respectively. These data suggest that dronedarone inhibits I(K(ACh)) by depressing the function of K(ACh) channel itself or associated GTP-binding proteins. Compared with amiodarone, dronedarone is approximately 100 times more potent on I(K(ACh)) and seems more selective in inhibiting I(K(ACh)) with respect to its antagonism of other inward and outward currents reported in the literature. This relative high potency of dronedarone to reduce I(K(ACh)) may be involved, at least in part, in the antiarrhythmic action of dronedarone against atrial fibrillation. PMID- 11117383 TI - Effect of genistein on cardiovascular responses to angiotensin II in conscious unrestrained rats. AB - Soybeans contain genistein, a phytoestrogen that may have beneficial effects in coronary artery disease, osteoporosis, and breast cancer. In vitro studies demonstrated that genistein reduced vascular smooth muscle contractions to angiotensin II. We tested the hypothesis that genistein attenuates the overall cardiovascular responses to angiotensin II via nongenomic mechanisms in conscious rats. Mean arterial blood pressure (MAP) was recorded from conscious unrestrained Sprague-Dawley rats (n = 26) approximately 48 hours after surgery. Cumulative dose response curves to angiotensin II (10-200 ng/kg/min) were constructed before and after i.v. treatment with genistein given as a single bolus dose of 160 microg/kg or 1500 microg/kg, or as a loading dose of 160 microg/kg followed by an infusion at a rate of 20 microg/kg. Angiotensin II infusions were associated with graded increases in arterial pressure ranging between 0+/-1 and 35+/-4 mm Hg. These pressor responses were accompanied by significant dose-dependent decreases in heart rate. None of the genistein treatment regimens significantly affected the pressor responses to angiotensin II. Accordingly, we conclude that short-term i.v. treatment with genistein does not depress pressor responsiveness to angiotensin II. PMID- 11117384 TI - The changing and complex relationship between paediatric cardiologists and life. PMID- 11117385 TI - Self-efficacy: understanding a psychological concept which can improve the quality of health care. PMID- 11117386 TI - Cardiac surgery in the setting of trisomy 13. PMID- 11117387 TI - Self-efficacy and physical activity in adolescents with trivial, mild, or moderate congenital cardiac malformations. AB - Our purpose was to examine the cognitive processes that influence involvement in physical activity among 100 adolescents, 55 boys and 45 girls, ranging in age from 12 to 18 years, with trivial, mild, or moderate forms of congenital cardiac disease. We hypothesized, first, that the severity of the congenital cardiac malformation itself has an indirect effect on self-efficacy regarding physical activity, and that the relationship between the two is mediated by the recommendations of the cardiologist and the attitude of the mother. Second, we argued that self-efficacy serves as a mediating variable between the recommendations of the cardiologist and the attitude of the mother, on the one hand, and involvement in physical activity, on the other. The results confirmed both hypotheses. In a population of adolescents with trivial to moderate congenital cardiac malformations, beliefs in self-efficacy, rather than severity of the disease, were the most influential factors in determining whether or not adolescents will engage in sports or other physical activities. We also demonstrated the importance of the role played by the recommendations of the cardiologist in determining both the attitudes of the mother and the belief in self-efficacy of the adolescents. PMID- 11117388 TI - Twisted atrioventricular connections in double inlet right ventricle: evaluation by magnetic resonance imaging. AB - Twisted atrioventricular connections occur almost exclusively in the hearts with biventricular atrioventricular connections. Only one example of double inlet left ventricle has been illustrated in which the axes of the two atrioventricular valves crossed each other. We describe herein three patients, and one autopsied specimen, with double inlet right ventricle in which magnetic resonance imaging clearly demonstrated twisted atrioventricular connections. PMID- 11117389 TI - Impact on parents of feeding young children with congenital or acquired cardiac disease. AB - Feeding an infant is an interactive process that facilitates social, emotional and culturally based skills. Children with congenital or acquired cardiac disease frequently require supportive regimes with regard to feeding so as to maintain weight, resulting in altered experiences for both the child and family. This study evaluated the practical, emotional and social ramifications for parents, of having a child with cardiac disease who also experienced difficulties with oral feeding. The study sampled three groups of parents who had children less than 3 years of age: those with cardiac disease who had difficulty in feeding, those with cardiac disease and no such difficulty, and those with no medical diagnosis. Parents completed a questionnaire about feeding, a time diary of activities involved in feeding, and Tuckman's Mood Thermometers, which measure anger and 'poorness-of-mood' associated with feeding the identified child. Parents of children with cardiac disease and a feeding difficulty reported a significantly more negative mood-state, and significantly longer time associated with feeding, than parents of children in the other two groups. Emerging themes from qualitative analysis of the data suggested that having a child with congenital cardiac disease producing difficulty in feeding had a strong negative impact on the whole family. PMID- 11117390 TI - Severe tortuosity and stenosis of the systemic, pulmonary and coronary vessels in 12 patients with similar phenotypic features: a new syndrome? AB - We describe what is, to the best of our knowledge, a previously unreported association in patients with similar facial features, skin and joint laxity, of lengthening and tortuosity of systemic, pulmonary and coronary vessels. We evaluated 12 patients with similar phenotypes, from eight different families. Detailed echocardiographic and angiographic evaluations were performed in all, and biopsies of the skin in seven. All patients have elongated facies, prominent ears, micrognathia and laxity of their joints. Angiographic pictures showed a varying degree of lengthening and tortuosity of systemic, pulmonary, and coronary arteries. Pulsatile carotid arteries formed cervical masses in 2 patients, and three had severe renal arterial stenoses. All showed varying degrees of branch and peripheral pulmonary arterial stenosis, necessitating placement of stents in six. Biopsy of the skin proved normal in all seven patients studied, thus excluding cutis laxa, Ehlers-Danlos and Marfan syndromes. The constellation of abnormalities suggests a genetic syndrome of connective tissue etiology. Further genetic studies, and gene mapping, are underway. PMID- 11117391 TI - Long-term results after valvotomy for congenital aortic valvar stenosis in children. AB - As interest increase in the Ross procedure performed as a therapeutic option for children with congenital aortic valvar stenosis, it becomes increasinly important to know the late results of aortic valvotomy in this population. We have therefore examined retrospectively the medical records of 121 consecutives survivors undergoing aortic valvotomy before 10 years of age between 1974 and 1992. The mean age at the first valvotomy was 29 months, with a range from 3 days to 10 years. The mean duration of follow up was 9.4 years, with a range from 1.6 to 22 years. Fifteen patients (12.3%; 70% CL: 10-16) died: 9 following reoperation, and 6 late after surgery. Death was related to the hearts in 86% of cases. The actuarial survival rate was 79% (70% CL: 72/84) at 10 years. Young age at the first valvotomy, and the number of procedures, emerged as risk factors of secondary mortality. Reoperations on the aortic valve, 73 in all, were required in 56 patients. The second procedure was done after a mean interval of 6 years, with a range from 1 day to 18 years. This was for restenosis in three-quarters of the cases. The aortic valve was replaced in 30 patients, at a mean of 9 years, and with a range from 9 months to 18 years, after the first procedure. The survival without replacement at 20 years was 29% (70% CL: 15-49). No factor was identified with a relationship either to reoperation or valvar replacement. Long term results after aortic valvotomy, therefore, show a high late mortality, frequent reinterventions, and an almost inescapable eventual need for valvar replacement. The ongoing use of the Ross operation is justified, even if longterm studies in children are still needed to validate its use. PMID- 11117392 TI - The middle aortic syndrome: an important feature of Williams' syndrome. AB - The middle aortic syndrome, with diffuse narrowing of the thoracic and abdominal aorta, was present in 10 of 18 patients with Williams' syndrome (55%). There were 3 thoracic coarctations, and 2 abdominal coarctations, with gradients greater than 20 mmHg across the zone of narrowing. Seven patients had mild renal arterial stenosis, and 6 had visceral arterial stenoses. Ten were hypertensive. Measured dimensions of the aortic lumen failed to increase with age in 3 males who had serial angiographic studies. One developed mesenteric arterial stenosis, with mild bilateral renal arterial stenoses, between the ages of 9 and 19 years. Aortic intravascular ultrasound performed in 2 patients confirmed abnormally thick vessel walls with small lumens. Diffusely narrowed and thick-walled stiff arteries, lacking elastin, are a feature of Williams' syndrome. The arteriopathy tends to progress with age, and systemic hypertension is common in teenagers and beyond. The middle aortic syndrome was present in more than half our patients, and does not necessarily reflect a bias because of cardiologic referral. Aortography with measurement of aortic diameters and delineation of the visceral branches is an important requirement for complete evaluation of patients with Williams' syndrome. PMID- 11117393 TI - Transthoracic three-dimensional echocardiography for the assessment of straddling tricuspid or mitral valves. AB - BACKGROUND: The advent of 3D echocardiography has provided a technique which, potentially, could afford significant additional information over conventional cross-sectional echocardiography in the assessment of patients with straddling atrioventricular valves prior to surgical correction. METHODS: Eight patients, aged from 1 month to 9.2 years, were examined with 3D echocardiography. All but three had discordant ventriculoarterial connections or double outlet right ventricle. Data suitable for reconstruction was acquired with transthoracic scanning. Right and left ventricular volumes were calculated in the 3D dataset. RESULTS: 3D echocardiography proved capable of defining the exact degree of straddling by imaging the proportion of tension apparatus attached to either side of the ventricular septum. It was able also to display the atrioventricular junction "en face", thus permitting identification of the precise site of insertion of the muscular ventricular septum relative to the atrioventricular junction. This made it possible first, to calculate the degree of valvar override, and second, to predict the location of the penetrating atrioventricular bundle. End-diastolic volume of the right ventricle in those with straddling tricuspid valves was 73 (61-83)% of normal, and, of the left ventricle in those with mitral valvar straddling 71 (40-97)% of normal. CONCLUSIONS: 3D echocardiography can aid in planning the optimal surgical procedure in patients with straddling or overrriding atrioventricular valves, as it provides diagnostic information superior to standard cross-sectional techniques. It also allows for exact measurement of the volumes of the respective ventricles. PMID- 11117394 TI - Myocardial high-energy phosphate metabolism is altered after treatment with anthracycline in childhood. AB - OBJECTIVES: We aimed to investigate whether changes in high-energy phosphate metabolism after treatment of children and young adults with anthracycline can be demonstrated non-invasively by 31P magnetic resonance spectroscopy. BACKGROUND: Abnormal myocardial energy metabolism has been suggested as a mechanism for anthracycline-induced cardiotoxicity. Deterioration in such has been shown in animal studies by resonance spectroscopy. METHODS: We studied 62 patients, with a mean age of 13.5+/-5 years, 3.7+/-4.3 years after a cumulative anthracycline dose of 270+/-137 mg/m2. Normal echocardiographic findings had been elicited in 54 patients. The control group consisted of 28 healthy subjects aged 20+/-7 years. Resonance spectrums of the anterior left ventricular myocardium were obtained at 1.5 Tesla using an image-selected in vivo spectroscopy localization technique. RESULTS: The ratio of phosphocreatine to adenosine triphosphate after blood correction was 1.09+/-0.43 for the patients, and 1.36+/-0.36 (mean+/-SD) for controls (p=0.005), with a significantly reduced mean ratio even in the subgroup of patients with normal echocardiographic results (1.11+/-0.44 versus 1.36+/ 0.36, p=0.01). The ratio did not correlate with the cumulative dose of anthracycline. The ratio of phosphodiester to adenosine triphosphate was similar in patients and controls (0.90+/-0.56 versus 0.88+/-0.62). CONCLUSIONS: In patients treated with anthracyclines in childhood, myocardial high-energy phosphate metabolism may be impaired even in the absence of cardiomyopathy. Our data support the concept that anthracycline-induced cardiotoxicity is not clearly dose dependent. PMID- 11117395 TI - Familial absent pulmonary valve syndrome without deletions of chromosome 22q11. AB - Deletions of chromosome 22q11 are common in patients with tetralogy of Fallot, and in those with absent pulmonary valve syndrome. In this report, we describe a pair of siblings with absent pulmonary valve syndrome, neither of whom had deletions of chromosome 22q11. The finding of familial absent pulmonary valve syndrome without deletion of 22q11 in our patients suggests an alternative genetic basis for this rare condition. PMID- 11117396 TI - Fetal diagnosis of lethal dysfunction of the right heart in three siblings. AB - A woman, having already delivered one child, underwent fetal echocardiography during three subsequent pregnancies. All three showed enlargement and poor function of the right-sided chambers. The first was still-born, the second died as a neonate, while the third pregnancy was terminated. Pathological examination revealed the same findings in each fetus, possibly representing a variation of Uhl's anomaly, or alternatively a hitherto unrecognised cardiomyopathic process. PMID- 11117397 TI - What should be provided by a service for fetal cardiology? PMID- 11117398 TI - Early extubation after surgical repair of tetralogy of Fallot. AB - In recent years, post-operative intensive care of the child with congenital cardiac disease has placed an emphasis on earlier weaning from mechanical ventilation. We describe our experience of postoperative fast-tracking of children undergoing cardiac surgery during a charitable mission in Venezuela, where resources and equipment were severely limited. During our stay, 11 children, with a median age of 2 years, underwent total correction of tetralogy of Fallot. The median duration of ventilation was 2.5 hours, and all patients were extubated within 12 hours of surgery. Effective analgesia was achieved without the need for continuous intravenous infusions of opiates. This experience shows that early extubation can safely be carried out in well-selected patients after surgery to correct congenital cardiac malformations. This allows faster throughput of patients, and helps provide an efficient and cost-effective service. PMID- 11117399 TI - Cardiac surgery in a girl with trisomy 13. AB - We describe a profoundly retarded infant girl with multiple anomalies caused by trisomy 13. Due to heart failure, which was resistant to medical treatment, we closed successfully a ventricular septal defect at three months of age. She died at 10 months of age. Despite the short survival, we believe that the patient benefitted significantly from the surgical repair of her cardiac defect. PMID- 11117400 TI - Recanalization of an occluded modified Blalock-Taussig shunt by balloon angioplasty within 12 hours of its construction. AB - An infant developed severe desaturation within a few hours of construction of a modified Blalock-Taussig shunt. Echocardiography revealed that the shunt had become occluded, and this was confirmed angiographically. At catheterisation, therefore, we passed a 0.014" percutaneous transluminal coronary angioplasty wire through the occluded shunt into the right pulmonary artery and then dilated the shunt successfully using a 5 mm coronary angioplasty balloon. Six weeks later, the shunt remained patent. PMID- 11117401 TI - Endovascular stenting as an emergency treatment for neonatal coarctation. AB - We describe a neonate with critical coarctation of the aorta. Since treatment with Prostaglandin E1 had failed to reopen the arterial duct, and surgery was deemed to be associated with an unacceptably high risk in this unstable neonate, it was decided to perform balloon dilation of the coarctation as a palliative procedure to stabilize the patient. Balloon angioplasty failed to reduce the pressure gradient across the coarctation, so a stent was implanted retrogradely as an emergency procedure. Subsequently, the stented segment was resected surgically and end-to-end anastomosis created without complications at seven months of age. PMID- 11117402 TI - Images in congenital heart disease. Kawasaki disease--on matters of the heart. PMID- 11117403 TI - Recommendations of the British Paediatric Cardiac Association for therapeutic cardiac catheterisation in congenital cardiac disease. AB - The aims of these recommendations are to improve the outcome for patients after, and to provide acceptable standards of practice of therapeutic cardiac catheterisation performed to treat congenital cardiac disease. The scope of the recommendations includes all interventional procedures, recognising that for some congenital malformations, surgical treatment is equally as effective as, or occasionally preferable to, interventional treatment. The limitations of the recommendations are that, at present, no data are available which compare the results of interventional treatment with surgery, and certainly none which evaluate the numbers and types of procedures that need to be performed for the maintenance of skills. Thus, there is a recognised need to collect comprehensive data with which these recommendations could be reviewed in the future, and re written as evidence-based guidelines. Such a review will have to take into account the methods of collection of data, their effectiveness, and the latest developments in technology. The present recommendations should, therefore, be considered as consensus statements, and as describing accepted practice, which could be used as a basis for ensuring and improving the quality of future care. PMID- 11117404 TI - Isomeric arrangement of the left atrial appendages and visceral heterotaxy. PMID- 11117405 TI - Anomalous origin of the main stem of the coronary artery associated with sudden death in a young athlete. PMID- 11117406 TI - Incidence of endophthalmitis after cataract surgery in the Netherlands: several surgical techniques compared. AB - In order to investigate the incidence of endophthalmitis following various types of cataract surgery, we sent a questionnaire to the members of the Dutch Intraocular Implant Club (NIOIC). Retrospectively, information was obtained about the number of performed cataract extractions, the techniques used, and the number of patients with postoperative endophthalmitis. To estimate bias by underreporting, we calculated the nationwide incidence of endophthalmitis after cataract surgery in the same period of time. The response rate to the questionnaire was 51.2%. In the reporting group the incidence of endophthalmitis was 0.11%. This incidence was comparable with the calculated nationwide incidence (0.15%). Comparison between the incidences after phacoemulsification (0.10%) and after other techniques (0.16%) showed no significant difference in the questionnaire group. A complicated cataract extraction preceded 12 out of the 38 reported cases with endophthalmitis. In conclusion, the incidence of endophthalmitis after cataract surgery in the Netherlands is comparable with the incidence reported in literature. The occurrence of complications during surgery rather than the technique used affects on the development of endophthalmitis. PMID- 11117407 TI - Results of scleral buckling operations in primary rhegmatogenous retinal detachment. AB - Within the scope of a Retinal Fellowship of one year, we evaluated the anatomic and functional results of scleral buckling operations in primary rhegmatogenous retinal detachments. Eighty Consecutive non-selected patients with a primary retinal detachment were operated by one surgeon (Retinal Fellow-ELH). In 55 eyes an encircling band and radial buckle(s) were placed, the other 25 eyes were treated with a segmental buckle or a combination of both. In 62 eyes subretinal fluid was drained, and in 57 eyes air or SF-6 gas was injected. The anatomic success rate after one operation was 81% (65/80 eyes) and the final success rate was 99%. 38/65 (58%) of the eyes obtained a best corrected post-operative visual acuity of > or = 0.4. The most important cause of re-detachment was Proliferative vitreoretinopathy (PVR; 11%). Pre-operative variables that yielded an unfavourable outcome in this study were: PVR, pseudophakic eye, larger breaks, more than one break, longer duration of the detachment, and 3 or more quadrants of detachment. Our anatomic success rate and risk factors are in agreement with findings described in the literature, yet we had a high rate of PVR and many patients with a low visual acuity (58% < or = 0.3) pre-operatively. PMID- 11117408 TI - Intrastromal corneal ring segments (ICRs): three- and six months results. AB - In the period from May 1997 to November 1998 twenty-one eyes of fifteen patients with sighted eyes received Intrastromal Corneal Ring Segments (ICRS) for the surgical treatment of low myopia. The mean pre-operative refraction was -3.0 diopters (D) ranging from -1.5 D to -4.1 D. Three months results showed that 44% of the treated eyes had an uncorrected visual acuity (UCVA) of 1.0 or better while 94% had an UCVA of 0.5 or better. In 94% the postoperative refraction was within one diopter (D) of emmetropia and in 61% within 0.5 D of emmetropia. Mean induced astigmatism was 1.03 (std 0.75). The best spectacle corrected visual acuity (BSC VA) after 3 months was 1.0 or better in all patients. After six months the UCVA was 1.0 or better in 43% of the treated eyes while in 100% the UCVA was 0.5 or better. The post-operative refraction was within 1 D of emmetropia in 100% and within 0.5 D of emmetropia in 81%. Mean induced astigmatism was 1.00 (std 0.50). BSCVA was in 88% 1.0 or better and in 100% 0.9 or better. In one patient vascular ingrowth occurred due to a ring segment located under the incision. Three patients experienced a lower UCVA due to postoperative astigmatism. In only one patient in this series the ICRS were removed because of induced astigmatism. These preliminary results show that the ICRS seem to offer a safe, stable and reproducible method to correct low myopia. The operation carries a low risk but care must been taken to insert the ICRS as central as possible to prevent postoperative astigmatism. PMID- 11117409 TI - Formation of proliferative vitreoretinopathy in primary rhegmatogenous retinal detachment. AB - Proliferative vitreoretinopathy (PVR) is the only cause of ultimate failure following retinal detachment surgery. This study aimed to review the rate of postoperative PVR in a series of 186 consecutive patients with primary rhegmatogenous retinal detachments. All 186 detachments were repaired with a scleral buckling procedure combined with cryotherapy. Drainage of subretinal fluid was done at the discretion of the surgeon. The mean follow-up was 12 months. In this series 152 (82%) of primary detachments were repaired succesfully with a single operation. Sixty-eight percent of patients regained 0.3 or better visual acuity, and 3% of patients were left with visual acuity of 1/60 or less. After two or more operations the retina was attached in 96% of the cases. In 12 (6%) eyes PVR was responsible for the initial surgical failure. In 4 cases PVR (grade B and limited C) was present prior to surgery. In 3 cases PVR developed within 2 days postoperatively, in 3 cases after 3-6 weeks and in another 2 cases after 8-10 months. Eight out of 12 (66%) PVR patients had undergone cataract surgery. One PVR case had preoperative intraocular inflammation. An association between the duration of retinal detachment, or drainage of subretinal fluid and the development of PVR could not be demonstrated. In conclusion, the rate of postoperative PVR in primary rhegmatogenous retinal detachments was low. PVR preoperatively present and pseudophakia may be risk factors. PMID- 11117410 TI - Phacotrabeculectomy. AB - The outcome of combined same-site phacoemulsification, posterior chamber lens implantation and trabeculectomy was retrospectively studied in patients with cataract and moderately controlled glaucoma, with a follow-up of at least 6 months. Primary phacotrabeculectomy without antimetabolites was performed in 74 patients. Mean IOP decreased from 22.8 to 14.3 mm Hg (35.3%). A maximum IOP of 19 mm Hg without glaucoma medication was reached in 66.2%. Mean logMAR visual acuity increased from 0.58 to 0.30. Primary phacotrabeculectomy has been shown to be a safe and effective procedure with good IOP control and rapid visual rehabilitation. PMID- 11117411 TI - Acute bacterial endophthalmitis after cataract extraction: results of treatment. AB - In order to evaluate the results, we reviewed all 34 patients treated in our hospital for endophthalmitis after cataract surgery between January 1994 and January 1998. After cultures were taken, all patients received intraocular, subconjunctival and topical vancomycin and ceftazidime. Additionally, twelve patients received the same antibiotics systemically. Besides steroids were administered in all patients. In 79% of the patients the bacterial culture was positive. Coagulase negative Staphylococcus was the most frequently isolated microorganism (48%). After treatment a visual acuity of 0.1 or more was achieved in 62% of the patients. The best final results were achieved in the patients with an initial visual acuity of 1/300 or more, and in the patients from whom a coagulase negative Staphylococcus was isolated. PMID- 11117412 TI - Retention properties of the fluorinated bonded phase on liquid chromatography of aromatic hydrocarbons. AB - The unique characteristics of a bonded-fluoroalkylsilane column, Fluofix, are described by comparison with those of a bonded octylsilane column, C8, in the reversed-phase chromatography of aromatic hydrocarbons. The selectivity of homologous aromatics on the fluorinated column varied with the methanol concentration in the eluent. At a larger proportion of methanol than 80:20 (v/v) the larger aromatics eluted faster than the smaller ones. However, with less methanol, the aromatics were eluted in order of their molecular size, as usually reported for a conventional hydrocarbonaceous column. A dependence of the retention mechanism on the composition of the eluent was suggested by the decrease in the entropy-enthalpy compensation temperature, beta, with increase of methanol concentration in the eluent. In order to explain the concentration dependence of the retention, the molecular interaction energy in the retention process was calculated by computer simulation. The interaction energy between aromatics and the stationary ligand on the Fluofix column was smaller than that on the C8 column and comparable to the interaction energy between the aromatics and methanol. At higher methanol concentrations, solute-fluorinated ligand complex formation was obstructed by the methanol molecules solvating the solute, reducing the retention of the larger aromatics. PMID- 11117413 TI - Degradation pathway of homoserine lactone bacterial signal molecules by halogen antimicrobials identified by liquid chromatography with photodiode array and mass spectrometric detection. AB - The degradation pathway of acylated homoserine lactone bacterial signaling molecules by oxidizing hypochlorite and stabilized hypobromite antimicrobials has been characterized. A reversed-phase HPLC separation using a cyano column was developed to detect the parent lactones, lactone-hydrolysis products, and halogenation products. Elucidation of the structures of the reaction products was done with the aid of online photodiode array UV spectroscopy and atmospheric pressure chemical ionization mass spectrometry. Quantitative output of the HPLC method was also used to estimate the kinetics of the degradation pathway. The results of this work found that only beta-keto-amide signal molecules are halogenated, where normal amide signals are not, and may represent one possible mechanism for control of industrial biofilms. PMID- 11117414 TI - DNA adsorption onto polyethylenimine-attached poly(p-chloromethylstyrene) beads. AB - In this study, DNA adsorption properties of polyethylenimine (PEI)-attached poly(p-chloromethylstyrene) (PCMS) beads were investigated. Spherical beads with an average size of 186 microm were obtained by the suspension polymerization of p chloromethylstyrene conducted in an aqueous dispersion medium. Owing to the reasonably rough character of the bead surface, PCMS beads had a specific surface area of 14.1 m2/g. PEI chains could be covalently attached onto the PCMS beads with equilibrium binding capacities up to 208 mg PEI/g beads, via a direct chemical reaction between the amine and chloromethyl groups. After PEI adsorption with 10% (w/w) initial PEI concentration, free amino content of PEI-attached PCMS beads was determined as 0.91 mequiv./g. PEI-attached PCMS beads were utilized as sorbents in DNA adsorption experiments conducted at +4 degrees C in a phosphate buffer medium of pH 7.4. DNA immobilization capacities up to 290 mg DNA/g beads could be achieved with the tried sorbents. This value was approximately 50-times higher relative to the adsorption capacities of previously examined sorbents. PMID- 11117415 TI - Improved method for rapid determination of vitamin A in small samples of breast milk by high-performance liquid chromatography. AB - Employing a short, potassium hydroxide (KOH) saponification (25 min) and rapid extraction with n-hexane-toluene (1:1) and a <4-min retention time, a convenient, rapid, accurate and precise determination of vitamin A in breast milk was developed. The recovery rate was 102.3 +/- 2.5% and the small relative standard deviation (intra-assay 1.9%, inter-assay 2.3%) resulted in high precision. The amount of breast milk needed is very low (1 ml) which makes the method suitable for the use in free field studies with large sample sizes. The simple extraction and separation steps allow a high throughput screening under routine laboratory conditions. PMID- 11117416 TI - Simplified determining procedure for routine residue monitoring of sulphamethazine and sulphadimethoxine in milk. AB - A simplified determining/identifying method for residual sulphamethazine (SMZ) and sulphadimethoxine (SDM) in milk by using a high-performance liquid chromatography (HPLC) with a photo-diode array detector was presented. Both sulphonamides in cow's milk samples were extracted by only stirring with ethanol followed by an Ultrafree-MC/Biomax as a centrifugal ultra-filtration unit. For determination/identification of SMZ and SDM, a Mightysil RP-18 GP Aqua column and a mobile phase of 25% (v/v) ethanol solution (in water) with a photo-diode array detector was used. Average recoveries from spiked SMZ and SDM (10-1000 ng/ml each drug) were > or = 83% with the relative standard deviations between 1.4 and 3.7%. The limit of quantitation (LOQ) were calculated to be 5 ng/ml for SMZ and 10 ng/ml for SDM, respectively. The values were below the MRL/tolerance (SMZ, 25 ng/ml; SDM, 10 ng/ml). The total time and solvent required for the analysis of one sample were <35 min and <2 ml of only ethanol, respectively. No toxic solvents were used. The developed procedure was harmless to the human and environment. PMID- 11117417 TI - Determination of perchlorate at parts-per-billion levels in plants by ion chromatography. AB - A method for the analysis of perchlorate in plants was developed, based on dry weight, and applied to the analysis of plant organs, foodstuffs, and plant products. The method reduced greatly the ionic interferences in water extracts of plant materials. The high background conductivity, due to the plant matrix, was reduced sufficiently to allow quantitation of perchlorate with little or no matrix interference. Ion chromatography (IC) on a microbore AS16 anion-exchange column and a conductivity detector was used for separation and detection of perchlorate from the ionic plant extract. The extract was heated to precipitate proteins, centrifuged, exposed to alumina, and filtered through a cartridge filled with divinylbenzene to yield a water clear extract for IC analysis, even from highly colored solutions. Heating the extract and treatment with alumina reduced substantially the ionic content of the extracts without loss of perchlorate. PMID- 11117418 TI - Optimization and evaluation of atomic emission gas chromatographic detection for nitrogen using the 388 nm molecular emission spectral band. AB - Nitrogen determination by gas chromatography with atomic emission (microwave induced plasma) detection (GC-AED) has been accomplished using the 174 nm atomic emission line, but with very limited selectivity and sensitivity. Nitrogen can also be detected using the cyanogen (CN) molecular band at 388 nm. A commercial GC-AED system was modified to allow the use of the 388 nm line for nitrogen detection, giving an improvement of 100-fold in sensitivity and selectivity, when compared with the 174 nm mode. Limits of detection of 10 pg/s were common, representing a 10-fold improvement. Compound-independent behavior was found for the system, working with optimum operating conditions, while instrumental problems were clearly reflected by a drastic compound dependent behavior. Response factors showed an important dependency upon the concentration of the element present. This dependency affected the accuracy of the determination of empirical formulae using GC-AED. PMID- 11117419 TI - Selectivity equivalence of poly(ethylene glycol) stationary phases for gas chromatography. AB - The solvation parameter model is used to study differences in selectivity for poly(ethylene glycol) stationary phases for packed column (Carbowax 20M) and fused-silica, open-tubular column (HP-20M, AT-Wax, HP-INNOWax and DB-FFAP) gas chromatography. All phases are dipolar, strongly hydrogen-bond basic with no hydrogen-bond acidity and of moderate cohesion. No two phases are exactly alike, however, and selectivity differences identified with cavity formation and dispersion interactions, n- and pi-electron pair interactions, dipole-type interactions and hydrogen-bond interactions are quantified by differences in the system constants at a fixed temperature where retention occurs solely by gas liquid partitioning. The system constants vary linearly with temperature over the range 60-140 degrees C (except for n- and pi-electron pair interactions which are temperature invariant) facilitating a general comparison of the importance of temperature on selectivity differences for compared phases. From a mechanistic point of view it is demonstrated that selectivity differences can result from chemical differences between the poly(ethylene glycol) stationary phases and from differences in the relative contribution of interfacial adsorption to the retention mechanism. The latter depends on both system properties and solute characteristics. PMID- 11117420 TI - Determination of ergosterol on mouldy building materials using isotope dilution and gas chromatography-tandem mass spectrometry. AB - Ergosterol content of building materials was quantified using gas chromatography tandem mass spectrometry (GC-MS-MS) in an ion trap with external ionisation. Hydrolysing the samples by classic extraction at 85 degrees C for 90 min in vials was faster, more precise and safer than microwave assisted extraction. [4 2H2]ergosterol was synthesised and used as internal standard, giving method standard deviation of 5-10% from 10 to 30 ng to 10-15 microg ergosterol in the sample. The use of GC-MS-MS meant that no solid-phase extraction clean-up was needed, so one person could easily prepare 40-80 samples per day. PMID- 11117421 TI - Gas-liquid chromatographic determination of major tobacco alkaloids. AB - Polar and apolar columns were tested for the chromatographic analysis of tobacco alkaloids. Disadvantages of polar-based polyethylene glycol phases are reported, such as the adsorption of the nornicotine alkaloid and the occurrence of errors on quantitation of alkaloids due to the presence of interfering compounds in tobacco extracts. A chromatographic method using a new apolar base-modified phase was evaluated for the analysis of alkaloids in tobacco products. The nicotine and other major alkaloids were quantitated using two internal standards simultaneously. PMID- 11117422 TI - Solid-phase microextraction for organochlorine pesticide residues analysis in Chinese herbal formulations. AB - Solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS) was used to determine pesticide residues in Chinese herbal formulations. Fibers coated with a 100-microm film thickness of poly(dimethylsiloxane) was used to extract 19 organochlorine pesticides (OCPs). The pesticides in the study consisted of alpha-, beta-, gamma-and delta hexachlorocyclohexane, p,p'-DDD, p,p'-DDE, p,p'-DDT, o,p'-DDT, aldrin, dieldrin, endrin, endrin aldehyde, endrin ketone, endosulfan (I, II and sulfate), heptachlor, heptachlor epoxide, and methoxychlor. The optimal experimental procedures for the adsorption and desorption of pesticides were evaluated. The linearity was obtained with a precision below 11% RSD for the studied pesticides expect endosulfan sulfate (21%) in a wide range from 1 to 200 ng/g. Detection limits were reached at below ng/g levels. Heptachlor epoxide was determined at a calculated limit of 0.03 ng/g. Comparison between SPME and Soxhlet extraction showed that SPME has a less than one order detection limit for residue pesticide determination. The proposed method was tested by analyzing herbal formulations from a local market for OCP multiresidues. Some residues studied were detected in the analyzed samples. The results demonstrate the suitability of the SPME-GC-MS approach for the analysis of multi-residue OCPs in Chinese herbal formulations. PMID- 11117423 TI - Separation of unmodified polystyrene nanosphere standards by capillary zone electrophoresis. AB - Size separation of five unmodified polystyrene nanosphere standards with diameters between 50 and 600 nm has been achieved in phosphate buffer solutions as carrier electrolyte. The electrophoretic mobility increases with particle diameter. Optical spectra was shown to be different for particles of different size. Effects of injection time, applied voltage, pH, and phosphate concentration in carrier electrolyte on particle separation were studied. Under optimal conditions the peak efficiency ranged from 600 to 10,500 theoretical plate numbers depending on nanosphere diameter was achieved. PMID- 11117424 TI - Fibre optical scanning with high resolution in thin-layer chromatography. AB - In this paper a high-performance thin-layer chromatography (HPTLC) scanner is presented in which a special fibre arrangement is used as HPTLC plate scanning interface. Measurements are taken with a set of 50 fibres at a distance of 400 to 500 microm above the HPTLC plate. Spatial resolutions on the HPTLC plate of better than 160 microm are possible. It takes less than 2 min to scan 450 spectra simultaneously in a range of 198 to 610 nm. The basic improvement of the item is the use of highly transparent glass fibres which provide excellent transmission at 200 nm and the use of a special fibre arrangement for plate illumination and detection. PMID- 11117425 TI - Determination of dyes in foodstuffs by capillary zone electrophoresis. AB - A rapid method based on capillary zone electrophoresis coupled with photodiode array detection has been developed to determine the dyes Tartrazine E-102, Sunset Yellow FCF E110, Amaranth E-123, New Coccine E-124, Patent Blue V calcium salt E 131 and Allura Red AC E-129 in foodstuffs. Separation was done by using a Bare CElect-FS75 CE column, using a 10 mM phosphate buffer at pH 11.0. Hydrodynamic injections at 0.5 p.s.i. for 4 s (21 nl of sample) and 20 kV separation voltage were used. The quantitation limits for the six dyes ranged from 3 to 6 microg/ml. A linear relationship between 3 to 95 microg/ml, with correlation coefficient better than 0.995 was obtained. This method has been applied to the determination of the studied dyes in beverages, jellies and syrups. PMID- 11117427 TI - Influence of dose and age on the response of the allopurinol test for ornithine carbamoyltransferase deficiency in control infants. AB - Measurement of urinary orotidine and orotic acid after an oral allopurinol challenge is an important diagnostic test for ornithine carbamoyltransferase deficiency that is sometimes used in infants (< 1 year of age), although there is little information on normal test results in this age group. We found higher orotidine excretion in normal infants than in older children given the test, whereas orotate excretion was similar in both groups. The increased orotidine excretion appears to be due to the use in the infants of higher allopurinol doses per kilogram of body weight than in the children. The normalized-dose dependency of the orotidine response extends even to adult age. Thus, dose-normalized responses should be used in the test and there is no need for careful age matching of the controls. PMID- 11117426 TI - Erythropoietic and hepatic porphyrias. AB - Porphyrias are divided into erythropoietic and hepatic manifestations. Erythropoietic porphyrias are characterized by cutaneous symptoms and appear in early childhood. Erythropoietic protoporphyria is complicated by cholestatic liver cirrhosis and progressive hepatic failure in 10%, of patients. Acute hepatic porphyrias (delta-aminolaevulinic acid dehydratase deficiency porphyria, acute intermittent porphyria, hereditary coproporphyria and variegate porphyria) are characterized by variable extrahepatic gastrointestinal, neurological psychiatric and cardiovascular manifestations requiring early diagnosis to avoid life-threatening complications. Acute hepatic porphyrias are pharmacogenetic and molecular regulatory diseases (without porphyrin accumulation) mainly induced by drugs, sex hormones, fasting or alcohol. The disease process depends on the derepression of hepatic delta-aminolaevulinic acid synthase following haem depletion. In contrast to the acute porphyrias, nonacute, chronic hepatic porphyrias such as porphyria cutanea tarda are porphyrin accumulation disorders leading to cutaneous symptoms associated with liver disease, especially caused by alcohol or viral hepatitis. Alcohol, oestrogens, haemodialysis, hepatitis C and AIDS are triggering factors. Porphyria cutanea tarda is the most common porphyria, followed by acute intermittent porphyria and erythropoietic protoporphyria. The molecular genetics of the porphyrias is very heterogenous. Nearly every family has its own mutation. The mutations identified account for the corresponding enzymatic deficiencies, which may remain clinically silent throughout life. Thus, the recognition of the overt disorder with extrahepatic manifestations depends on the demonstration of biochemical abnormalities due to these primary defects and compensatory hepatic overexpression of hepatic delta aminolaevulinic acid synthase in the acute porphyrias. Consequently, haem precursors are synthesized in excess. The increased metabolites upstream of the enzymatic defect are excreted into urine and faeces. The diagnosis is based on their evaluation. Primary enzymatic or molecular analyses are noncontributary and may be misleading. Acute polysymptomatic exacerbations accompany a high excretory constellation of porphyrin precursors delta-aminolaevulinic acid and porphobilinogen. Homozygous or compound heterozygous variants of acute hepatic porphyrias may already manifest in childhood. PMID- 11117428 TI - Mutation detection in 65 families with a possible diagnosis of ornithine carbamoyltransferase deficiency including 14 novel mutations. AB - The high new mutation rate and the wide spectrum of mutations found in patients with ornithine carbamoyltransferase (OCT) deficiency means that direct mutation analysis is essential for providing accurate carrier detection and prenatal diagnosis in affected families. We present our strategy for mutation detection in the OCT gene and summarize the results from 31 families with a confirmed diagnosis and 34 families with a suspected diagnosis of OCT deficiency, and describe 14 previously unreported mutations. PMID- 11117429 TI - Phenylalanine supplementation improves the phenylalanine profile in tyrosinaemia. AB - Tyrosinaemia types I and II are caused by enzyme deficiencies in the tyrosine catabolism pathway. Successful treatment is possible with the novel enzyme inhibitor NTBC in tyrosinaemia type I and with dietary tyrosine and phenylalanine restriction in both conditions. This is achieved with a low natural protein intake and a supplementary amino acid formula that is phenylalanine- and tyrosine free. Patients on this regimen had been noted, periodically, to have very low plasma phenylalanine concentrations (<20 micromol/L). The tyrosine and phenylalanine profiles in six patients were measured. Five of the six patients had very low concentrations of phenylalanine during the later half of the day. The response to phenylalanine supplementation was assessed and supplementing the diet with phenylalanine 30-40 mg/kg per day resulted in normal concentrations throughout the day. Possible complications of hypophenylalaninaemia and potential preventive treatment strategies are discussed. Further studies are needed to investigate the longer-term clinical and biochemical consequences of phenylalanine supplementation. PMID- 11117430 TI - Effects of ethanol and of alcohol dehydrogenase inhibitors on the reduction of N acetylaspartate levels of brain in mice in vivo: a search for substances that may have therapeutic value in the treatment of Canavan disease. AB - N-Acetylaspartate (NAA) is an important osmolyte in the vertebrate brain that participates in an intercompartmental metabolic cycle. It is synthesized primarily in neurons from L-aspartate (Asp) and acetyl-CoA and, after its regulated release, it is hydrolysed by aspartoacylase in an oligodendrocyte compartment to produce Asp and acetate. NAA also gives a strong 1H magnetic resonance spectroscopic signal, which has led to its widespread use as a neuronal marker. Utilizing this noninvasive technique, the NAA concentrations in normal brain and in brains exhibiting a variety of CNS disease syndromes have been studied. In normal individuals, the concentration of NAA has been observed to be relatively stable over long periods. However, in many CNS disease processes there are long-term changes in the level of NAA that have been considered to signal changes in neuron density or function. We report that the concentration of NAA in brain is malleable and that, in addition to normal endogenous variation or changes due to disease processes, it can be modified by a variety of exogenous drugs and other substances. As a result of this investigation, we have also been able to identify a new class of NAA-active compounds--pyrazole and pyrazole derivatives--that have the ability to reduce brain NAA concentrations in normal mice. The importance of these findings in understanding the NAA intercompartmental cycle, and its role in Canavan disease, a genetic aspartoacylase deficiency disease, are discussed. PMID- 11117431 TI - Differentiation of hair growth cycle from scalp hair roots for the diagnosis of glucose-6-phosphate dehydrogenase deficiency in neonates. AB - Hair analysis can be used as a screening tool in the diagnosis of genetic diseases. The scalp hair roots of 67 normal neonates and 39 neonates with glucose 6-phosphate dehydrogenase (G6PD) deficiency were analysed using Fourier transform infrared (FT-IR) microspectroscopy to differentiate the stages of the hair growth cycle and to diagnose the genetic disorder on the basis of spectral differences. We have demonstrated that FT-IR microspectroscopy is a rapid and effective noninvasive diagnostic method to differentiate scalp hair roots of normal neonates into the anagen, catagen or telogen phases of the hair growth cycle, using IR spectral differences within the 3000-2800 cm(-1) region and the IR peak area ratio of 2854 cm(-1)/2873 cm(-1) or 1084 cm(-1)/amide II band (p<0.001). Moreover, G6PD-deficient neonates could be accurately diagnosed from telogen phase hair roots owing to significant differences in IR peak area ratios of 2854 cm(-1)/2873(-1) or 1084 cm(-1)/amide II band compared to normal values in healthy neonates. The result suggests that the application of FT-IR microspectroscopy may be capable not only of differentiating the hair growth cycle into anagen, categen or telogen phases but also of detecting G6PD deficiency. Hair root analysis promises to be a useful complement to serum and urine analysis in the diagnosis of genetic diseases. PMID- 11117433 TI - Studies of the V94M-substituted human UDPgalactose-4-epimerase enzyme associated with generalized epimerase-deficiency galactosaemia. AB - Impairment of the human enzyme UDPgalactose 4-epimerase (hGALE) results in epimerase-deficiency galactosaemia, an inborn error of metabolism with variable biochemical presentation and clinical outcomes reported to range from benign to severe. Molecular studies of the hGALE loci from patients with epimerase deficiency reveal significant allelic heterogeneity, raising the possibility that variable genotypes may constitute at least one factor contributing to the biochemical and clinical heterogeneity observed. Previously we have identified a single substitution mutation, V94M, present in the homozygous state in all patients genotyped with the severe, generalized form of epimerase-deficiency galactosaemia. We report here further studies of the V94M-hGALE enzyme, overexpressed and purified from a null-background yeast expression system. Our results demonstrate that the mutant protein is impaired relative to the wild-type enzyme predominantly at the level of Vmax rather than of Km. Studies using UDP-N acetylgalactosamine as a competitor of UDPgalactose further demonstrate that the Km values for these two substrates vary by less than a factor of 3 for both the wild-type and mutant proteins. Finally, we have explored the impact of the V94M substitution on susceptibility of yeast expressing human GALE to galactose toxicity, including changes in the levels of galactose 1-phosphate (gal-1-P) accumulated in these cells at different times following exposure to galactose. We have observed an inverse correlation between the level of GALE activity expressed in a given culture and the degree of galactose toxicity observed. We have further observed an inverse correlation between the level of GALE activity expressed in a culture and the concentration of gal-1-P accumulated in the cells. These data support the hypothesis that elevated levels of gal-1-P may underlie the observed toxicity. They further raise the intriguing possibility that yeast may provide a valuable model not only for assessing the impact of given patient mutations on hGALE function, but also for exploring the metabolic imbalance resulting from impaired activity of GALE in living cells. PMID- 11117434 TI - Microphotometric analysis of NADH-tetrazolium reductase deficiency in fibroblasts of patients with Leber hereditary optic neuropathy. AB - We employed a microphotometric approach to examine whether a defect in the mitochondrial respiratory complex I expected in Leber hereditary optic neuropathy (LHON) as the consequence of a mtDNA (11778G>A) mutation in the ND4 gene coding for a subunit of the respiratory complex I can be detected at the single-cell level. Genetically stable fibroblast cell lines were established from skin biopsies of two members of a Chinese Indonesian family with LHON. The fibroblasts were homoplasmic for the 11778G>A mutation. The activity of the respiratory complex I was examined histochemically by staining for NADH-tetrazolium reductase. The histochemical staining showed a typical pattern with an apparent concentration of the activity around the nucleus, suggested as the reflection of the gradient in the thickness of the unsectioned fibroblast cells. Microphotometric quantification of the staining intensity showed that the activity is linear for at least 60 min. The activity shows a discontinuity in its Arrhenius kinetics with a break point at 13.0-13.5 degrees C (activation energy at 50-58 J/mol and 209-238 J/mol above and below the break temperature, respectively), indicating the membrane association of the NADH-tetrazolium reductase activity. Both patients showed lower fibroblast NADH-tetrazolium reductase activity, with a reduction of degrees 30%. Our results demonstrate the utility of microphotometric analysis in the study of biochemical defects associated with mutations in the mtDNA. PMID- 11117432 TI - A serine/alanine polymorphism in the nucleotide-binding fold-2 of the sulphonylurea receptor-1 (S1369A) is associated with enhanced glucose-induced insulin secretion during pregnancy. AB - The sulphonylurea receptor-1 (SUR-1) regulates glucose-induced insulin secretion by controlling K+-ATP channel activity of the pancreatic beta-cell membrane. In this study, we investigated the putative role of a T/G-polymorphism (exon 33, codon 1369; S1369A) in the adenosine diphosphate-sensing nucleotide-binding fold 2 (NBF-2) of the SUR-1 on glucose-induced insulin secretion during an oral glucose tolerance test in pregnant women (PW; n=182). Compared to PW with the T/T genotype, statistically significant elevated C-peptide concentrations were found 60 min after glucose intake in PW with the T/G and G/G genotype (T/T 9.0+/-0.4 ng/ml vs T/G 10.8+/-0.4 ng/ml or G/G 10.8+/-0.7 ng/ml, p=0.01). Furthermore, compared to PW with T/T genotype the deltaC-peptide (60/0 min) was significantly enhanced in PW with T/G or G/G genotype (T/T 6.7+/-0.3 vs T/G 8.9+/-0.4 or G/G 8.9+/-0.7, p=0.0009). A significant correlation of C-peptide concentrations with blood glucose (BG) 60 min after glucose intake was only found in PW with the T/T genotype (r=0.6, p<0.0004). Similarly, a significant correlation of insulin concentrations with BG 60 min after glucose intake was observed in PW with T/T genotype (r=0.5, p<0.0001) and T/G genotype (r=0.24, p<0.03) but not in PW with G/G genotype (r=0.01, p=0.9). From our data we conclude that in PW with the alanine substitution in the NBF-2 region, the insulin response of the pancreatic beta-cell after glucose intake is enhanced and does not correlate with actual BG levels. PMID- 11117435 TI - Quantitative measurement of total and free 3-hydroxy fatty acids in serum or plasma samples: short-chain 3-hydroxy fatty acids are not esterified. AB - Diagnostic protocols for disorders of mitochondrial fatty acid oxidation (FAO) generally include the measurement of plasma acylcarnitines. Many biochemical intermediates of FAO resulting from a metabolic block require carnitine conjugation for transport out of the mitochondria, and so occur as fatty acid carnitine conjugates in the blood. Both short- and long-chain acylcarnitines are generally determined, and this procedure has a critical role to play in the diagnosis of disorders of the very long-chain, medium-chain and short-chain acyl CoA dehydrogenase defects. Less is known about the utility of acylcarnitines for the measurement of the various chain length intermediates of the 3-hydroxyacyl CoA dehydrogenase steps of beta-oxidation. This study utilizes stable-isotope dilution gas chromatography-mass spectrometry to determine the serum or plasma concentrations of free 3-hydroxy fatty acids (3-OHFAs) of chain lengths C6 to C16. The 3-OHFA concentrations are determined in samples from normal individuals, hyperketotic individuals and patients with long-chain L-3-hydroxyacyl-CoA dehydrogenase and short-chain L-3-hydroxyacyl-CoA dehydrogenase deficiencies, both before and after hydrolysis. The results of the study indicate the relative amounts of conjugated intermediates of all chain lengths. Long-chain 3-OHFAs (C14 and C16) are found in elevated concentrations after hydrolysis, whereas short chain and medium-chain 3-OHFAs (C6 to C12) show no difference in concentrations between the two samples in all subjects tested, suggesting that only long-chain 3 hydroxy species form conjugates. This finding has important implications for the use of the acylcarnitine assay for the diagnosis of defects involving short-chain and medium-chain 3-hydroxy fatty acids. PMID- 11117436 TI - A new case of 2-methylacetoacetyl-CoA thiolase deficiency? PMID- 11117437 TI - Early onset of complete heart block in Pearson syndrome. PMID- 11117438 TI - Fatal neonatal outcome in a case of muscular mitochondrial DNA depletion. PMID- 11117439 TI - Fumarate hydratase deficiency: increased fumaric acid in amniotic fluid of two affected pregnancies. PMID- 11117440 TI - Hyperketonaemia in glycerol kinase deficiency. PMID- 11117444 TI - Continuous monitoring of the stimulated area in multifocal ERG. AB - INTRODUCTION: Multifocal electroretinography (MF-ERG) is widely used in the detection of local retinal dysfunction. However, the position of the stimulus on the retina and the stability of fixation are usually not directly accessible. Thus, devices have been designed for a continuous fundus visualization during recording. METHODS: MF-ERGs were recorded with a RetiScan system connected to two different Scanning-laser ophthalmoscopes (SLOs) that use either a red (633 nm) or green (415 nm) laser for stimulation, and a VERIS 4 system connected to a piggyback stimulator prototype that added the stimulus to the optical pathway of the SLO by means of a wavelength-sensitive mirror. Fundus visualization was achieved with the infrared lasers of the SLOs (780 and 835 nm). RESULTS: The most extensive study so far with a green laser stimulus in a cat model of retinal degeneration demonstrated the capability of the device to detect retinal landmarks and the different stages of degeneration. Also, the advantages of exactly reproducible stimulus positioning for averaging within and comparison between disease groups became apparent. The results with the same setup in transgenic mice suggest a pure cone origin of the responses. In humans, recordings show that fixation is sufficiently good in most subjects. It is not clear yet whether red or green laser stimulation (or both) is preferable. The results with the prototype were very similar to the MF-ERGs obtained with a standard CRT screen. CONCLUSIONS: All three devices allowed us to record MF-ERGs with continuous fundus monitoring. Although further refinements are necessary, it is obvious that fundus controlled methods will improve the reliability of MF-ERG in future research on glaucoma, transplantation studies, and evaluation of gene therapy. PMID- 11117446 TI - Multifocal, pattern and full field electroretinograms in cats with unilateral optic nerve section. AB - AIM: To look for a subcomponent of the mFERG generated at the optic nerve head and increasing in latency with distance from it. To compare multifocal electroretinogram (mFERG, mPERG) changes to those in full field ERGs and transient and steady state pattern and focal ERGs (PERGs, FERGs) in cats with total unilateral optic nerve section. METHOD: We recorded multifocal flash ERGs (mFERGs) at three levels of intensity and multifocal pattern ERGs (mPERGs) within 61 equal areas after total unilateral optic nerve section in five long term (> 18 month) survival cats, as part of a long term serial study of full field flash and pattern ERG changes to many stimuli, in a larger population. Cats were anaesthetised with Ketamine/Xylazine and wore Henkes electrodes with 6mm artificial pupils. Intact retinal circulation was verified by fluorescein angiography and optic nerve section by retinal photography and histology. We compared the mean and mean summed multifocal responses, from the normal and denervated eyes. We also compared the mean interocular difference around the area centralis and as a function of distance from the optic nerve head, across the horizontal meridian for the mFERG to the most intense stimulus. The degree of change was compared to that in other types of ERG, in the larger set of cats. RESULTS: mFERGs were similar across cats. Response density was flat with no prominence at the area centralis. Average summed mFERGs were similar in the normal and denervated eye. In the interocular differences a component near OP2 was reduced in the first kernel to the most intense stimulus, near OP1 and OP3 in the second kernel and locally, there was a hint of a component near OP2, which varied in latency in a ring around the disk and at the area centralis. Nevertheless, no component could be seen, varying in latency with distance from the optic nerve head, across the horizontal meridian. No mPERG was recordable in these conditions. Full field PERGs and FERGs were very reduced. Full field flash ERG amplitude changes were small (4-20%) and slower to appear than PERG changes. Degree of ERG reduction and correlation with PERG losses was greatest for the mesopic OPs, low for the scotopic tests (STR, ERG and OPs) and near zero for the mesopic ERG. The mFERG and mesopic ERG both lost OPs without overall amplitudes. CONCLUSIONS: The cat mFERG does not have a component, varying in latency with distance from the optic nerve head. The only change was qualitiatively similar to that in the light adapted ERG. With intense stimuli there were local changes around the time of OP2 near the area centralis which might be explained by local variations in ganglion cell density. PMID- 11117451 TI - The effects of change in lead stimulus modality on the modulation of acoustic blink startle. AB - In two experiments we investigated the effect of generalized orienting induced by changing the modality of the lead stimulus on the modulation of blink reflexes elicited by acoustic stimuli. In Experiment 1 (n = 32), participants were presented with acoustic or visual change stimuli after habituation training with tactile lead stimuli. In Experiment 2 (n = 64), modality of the lead stimulus (acoustic vs. visual) was crossed with experimental condition (change vs. no change). Lead stimulus change resulted in increased electrodermal orienting in both experiments. Blink latency shortening and blink magnitude facilitation increased from habituation to change trials regardless of whether the change stimulus was presented in the same or in a different modality as the reflex eliciting stimulus. These results are not consistent with modality-specific accounts of attentional startle modulation. PMID- 11117452 TI - Heritability of different measures of smooth pursuit eye tracking dysfunction: a study of normal twins. AB - Research studies have found that smooth pursuit eye movement dysfunction may serve as an index of genetic liability to develop schizophrenia. The heritability of various measures of smooth pursuit eye tracking proficiency and the saccades that occur during smooth pursuit was examined in 64 monozygotic (MZ) and 48 dizygotic (DZ) twin pairs. Two age cohorts were assessed (11-12 and 17-18 years of age). Intraclass correlations indicated significant similarity in the MZ twins for almost all measures in both age cohorts, whereas few of the DZ twin correlations attained significance. Biometrical modeling indicated that genetic mechanisms influence performance on both global and specific eye tracking measures, accounting for about 40% to 60% of the variance. These findings suggest that the underlying brain systems responsible for smooth pursuit and saccade generation during pursuit are under partial genetic control. PMID- 11117453 TI - Motor and nonmotor event-related potentials during a complex processing task. AB - Identification of the necessary stimulus properties to elicit the stimulus preceding negativity (SPN) has been the impetus for numerous research studies. The current study was conducted to explore the possibility that the SPN is an index of cognitive resource allocation. An auditory warning stimulus (S1) indicated whether an easy or difficult discrimination would occur at S2. The SPN was collected before a nonmotor discrimination task (S2) that consisted of identifying the higher of two bars. To eliminate the influence of motor processing prior to S2, a button press on the side of the higher bar was held until perception of a response cue (S3). Additionally, P3, contingent negative variation (CNV), and behavioral measures were collected to assist in assessing the SPN. Results indicated that although the SPN exhibited increased negativity, no differences were observed based on task difficulty. However, task difficulty did affect P3 data for both the warning tone and the discrimination task, an effect not observed for the CNV. Overall, the data did not support that hypothesis that the SPN provides an index of cognitive demand. PMID- 11117454 TI - An electrophysiological index of stimulus unfamiliarity. AB - This study investigated the functional significance of the N2 response to novel stimuli. In one condition, background, target, and deviant stimuli were simple geometric figures. In a second condition, all stimulus types were unfamiliar/unusual figures. In a third condition, background and target stimuli were unusual figures and deviant stimuli were simple shapes. Unusual figures, whether they were deviant, target, or background stimuli, evoked larger N2 responses than their simple, familiar counterparts. N2 elicited by an unusual background stimulus was larger than that evoked by simple, deviant stimuli, a pattern opposite that exhibited by the subsequent P3. Deviance from immediate context had limited influence over N2 amplitude. The results suggest that novelty N2 and novelty P3 reflect the processing of different aspects of "novel" visual stimuli. The novelty P3 is particularly sensitive to deviation from immediate context. In contrast, the novelty N2 is sensitive to deviation from long-term context that renders a stimulus unfamiliar and difficult to encode. PMID- 11117455 TI - Cardiovascular reactivity and adaptation to recurrent psychological stress: the moderating effects of evaluative observation. AB - The impact of evaluative observation on cardiovascular reactivity and adaptation to recurrent psychological stress was evaluated in 162 undergraduate men and women. All participants performed three mental arithmetic tasks with or without evaluative observation. Impedance cardiographic, blood pressure, task performance, and stress appraisal measures were recorded for each task. Evaluative observation moderated the effects of task repetition on cardiac reactivity but not vascular reactivity. The introduction of evaluative observation disrupted cardiac adaptation, resulting in a resurgence of beta adrenergic cardiac reactivity (p < .005), whereas the removal of evaluative observation promoted cardiac adaptation. Evaluative observation also increased stress appraisals and slowed task performance. The results support the dual process theory of habituation, rather than stimulus comparator theory, but only partially support cognitive appraisal theory. PMID- 11117456 TI - Rate of force development and the lateralized readiness potential. AB - We examined the relationship between force and rate of force development aspects of movement dynamics and electroencephalogram motor components as reflected in the lateralized readiness potential (LRP). Using self-paced tasks, in Studies 1 and 3 we investigated whether differential speed and accuracy constraints in discrete and repetitive finger force production tasks influenced the LRP. These studies showed that speed tasks produced larger LRP than accuracy tasks regardless of whether the movement type was discrete or repetitive. In Studies 2 and 4 we studied four conditions with two levels of force and two levels of rate of force development. The largest LRPs were found with the greatest rate of force development. Overall, the four studies demonstrated that preparation for differential rates of force development is a major component reflected in the LRP. PMID- 11117457 TI - Scalp recorded direct current potential shifts associated with quenching thirst in humans. AB - As an indicator of cortical excitability, direct current (DC) potentials were recorded from thirsted subjects before, during and after drinking 400 ml of water. Self-rated thirst was distinctly reduced after drinking. Compared with control conditions in which the subjects remained thirsty, during drinking a widespread negative potential shift occurred averaging over -70 microV at Cz. At the transition from the consumatory phase to the postconsumption phase, a slow positive potential shift commenced that was most pronounced over the anterior cortex (averaging over +40 microV at Fz) and persisted for more than 3 min after drinking. Control conditions excluded muscle activity, ocular movements, and changes in body fluid and serum osmolality as possible non-neuronal sources of the DC-potential changes. The sequence of negative and positive potential shifts associated with drinking indicates a coordinate regulation of cortical excitability that may facilitate consumatory behavior and its context-dependent encoding into memory. PMID- 11117458 TI - Electrodermal activity and temperament in preschool children. AB - This study had two objectives: To examine poorly understood patterns of young children's electrodermal reactivity and to test the hypothesis that this reactivity reflects individual differences in the behavioral inhibition system (BIS). We recorded skin conductance responses (SCRs) from 92 4-year-old children during a laboratory session that encompassed physiological and psychological stimuli. Physiological stimuli (breaths), moderately loud to loud sounds (expected and unexpected) and, to a lesser extent, stimuli with psychological significance elicited clear SCRs. Induction of psychological conflict and exposure to emotional film clips for the most part did not elicit increases in skin conductance (SC). Children's temperament dimensions of fearfulness and effortful (or inhibitory) control--two components of the BIS--were assessed using robust observational batteries at age 2 and 4 years. The theoretically expected correlations between overall SC lability (reflecting SC levels) and both dimensions of temperament were significant, albeit modest and limited to the contemporaneous measures at age 4. PMID- 11117459 TI - Temperament as a moderator of pathways to conscience in children: the contribution of electrodermal activity. AB - A longitudinal study (G. K. Kochanska, 1997) showed that temperamental fearfulness, assessed at toddler age via observational data and maternal ratings, moderated pathways to internalized conscience at age 4. For fearful children, maternal gentle discipline deemphasizing power predicted conscience development; for fearless children, attachment security predicted conscience development. Electrodermal reactivity assessed at age 4 on the same children was used as a physiological reflection of fearful temperament and was substituted for the earlier fearfulness measure to test the theoretical model. As expected, for electrodermally reactive children, maternal gentle discipline predicted conscience, whereas for nonreactive children, attachment security predicted conscience. The findings support the notions of (a) electrodermal reactivity at an early age as a correlate of temperament, (b) temperament as a moderator of socialization in early moral development, and (c) lovelessness in psychopathic individuals as an index of the failure of the alternative pathway (via attachment) to conscience in fearless children. PMID- 11117460 TI - Right N170 modulation in a face discrimination task: an account for categorical perception of familiar faces. AB - Behavioral studies have shown that two different morphed faces belonging to the same identity are harder to discriminate than two faces stemming from two different identities. The temporal course of this categorical perception effect has been explored through event-related potentials. Three kinds of pairs were presented in a matching task: (1) two different morphed faces representing the same identity (within), (2) two other faces representing two different identities (between), and (3) two identical morphed faces (same). Following the second face onset in the pair, the amplitude of the right occipitotemporal negativity (N170) was reduced for within and same pairs as compared with between pairs, suggesting an identity priming effect. We also observed a modulation of the P3b wave, as the amplitude of the responses for within pairs was higher than for between and same pairs, suggesting a higher complexity of the task for within pairs. These results indicate that categorical perception of human faces has a perceptual origin in the right occipitotemporal hemisphere. PMID- 11117461 TI - Mismatch negativity in children and adults, and effects of an attended task. AB - Attention has been shown to modulate the amplitude of the mismatch negativity (MMN) elicited by a small deviation in auditory stimuli in adults. The present study examined the effects of attention and deviant size on MMN amplitude in children. Children and adults were presented with sequences of tones containing standards (1000 Hz) and three deviants varying in degree of deviance from the standard (1050, 1200, and 1500 Hz). Tones were presented in three conditions: (1) while participants ignored them; (2) while participants listened to them and responded to all three deviants; and (3) while participants again ignored them. We found that the MMNs elicited by the hard deviant (1050 Hz) were larger when the children were actively discriminating the stimuli than when they were ignoring them. However, the MMNs elicited by the easy and medium deviants (1500 and 1200 Hz, respectively) in the children and by all three deviants in the adults were not affected by attention. PMID- 11117462 TI - Hierarchical processing and level-repetition effect as indexed by early brain potentials. AB - Event-related potentials were recorded to investigate the mechanisms of hierarchical processing and level-repetition effect. Participants identified targets that appeared at global, local, or both levels of hierarchical patterns. Reaction times showed global precedence and level-repetition effects. An occipital P1 wave was enhanced to local relative to global targets. The P1 to local targets was also larger when preceded by global than local targets. Global and both-level target selections were indexed by two posterior negativities peaking at 130 and 190 ms poststimulus, whereas local target selection was indexed by a broad occipitotemporal negativity. A late selection positivity was observed over the left occipitotemporal site for global targets but over the central site for local targets. The findings suggest that sensory-perceptual mechanisms contribute to global precedence and level-repetition effects in hierarchical processing. PMID- 11117463 TI - The relationship between respiratory-related evoked potentials and the perception of inspiratory resistive loads. AB - This study investigated the relationship between resistive load magnitude, load magnitude estimation, and the respiratory-related evoked potential. In Part 1, 10 healthy subjects estimated the magnitude of five inspiratory resistive loads. Two subjects were shown to have a markedly reduced slope of the magnitude estimation resistive load relationship and were suggested to be "poor perceivers" of respiratory stimuli. In Part 2, evoked potentials were recorded from the same 10 subjects using the same resistive loads as Part 1. A log-log plot of the group averaged P1 amplitudes showed a linear relationship with resistive load. Aberrant P3 components were seen in the 2 poor perceiving subjects and one of the 2 showed no late response. In the other 8 subjects, P3 varied as a function of resistive load, being augmented to larger loads. These results provide evidence that P3 may be a key index of the perception of respiratory sensitivity and effort. PMID- 11117464 TI - Patterns of sympathetic and parasympathetic reactivity in a sample of children and adolescents. AB - We hypothesized that patterns of sympathetic and parasympathetic reactivity observed in adults would be apparent in a sample of children and adolescents and that these patterns would be consistent across tasks. We explored the relationship between these patterns and psychosocial risk factors for cardiovascular disease. We measured preejection period (PEP) and an index of respiratory sinus arrhythmia (mean successive difference [MSD] statistic) during three reactivity tasks. We classified participants into four groups based on increases or decreases in PEP and MSD. Ninety percent of the sample exhibited the same pattern during at least two of the tasks. PEP and MSD demonstrated consistency, suggesting individual response stereotypy. Exhibiting a consistent pattern of decreased PEP and increased MSD was associated with less child- and parent-reported family conflict. These results are discussed in the context of vagal regulation of environmental demands. PMID- 11117465 TI - Immediate and delayed stimulus repetitions evoke different ERPs in a serial-probe recognition task. AB - In this study we examined whether event-related potentials (ERPs) associated with stimulus repetition and recognition in a serial-probe recognition task were comparable to ERPs in other tasks that are more typically used to investigate old/new ERP effects. The experiment consisted of 320 trials in which a recognition probe followed a four-item memory set; 160 trials consisted of images depicting common objects that were easy to label (EL task), and 160 trials consisted of images depicting abstract patterns that were difficult to label (DL task). Nineteen participants indicated whether a probe that followed each memory set was or was not presented in the memory set. Half of the probes matched, and half did not match, an item in the preceding memory set. ERPs appeared to reflect two processes--one that differentiated between recently presented stimuli and other stimuli and another that distinguished between repeated stimuli and new stimuli. ERPs to recent probes were more positive than ERPs to other probes in the EL and DL tasks. ERPs to match (old) probes were more positive than ERPs to nonmatch (new) probes only in the EL task. PMID- 11117466 TI - Time adaptive denoising of single trial event-related potentials in the wavelet domain. AB - We present a new wavelet-based method for single trial analysis of transient and time variant event-related potentials (ERPs). Expecting more accurate filter settings than achieved by other techniques (low-pass filter, a posteriori Wiener filter, time invariant wavelet filter), ERPs were initially balanced in time. By simulation, better filter performance could be established for test signals contaminated with either white noise or isospectral noise. To provide an example of real application, the method was applied to limbic P300 potentials (MTL-P300). As a result, variance of single trial MTL-P300s decreased, without restricting the corresponding mean. The proposed method can be regarded as an alternative for single-trial ERP analysis. PMID- 11117467 TI - Endomorphin-2 but not Leu-enkephalin modulates spatial learning when microinjected in the CA3 region of the rat hippocampus. AB - The recently discovered endogenous mu-selective opioid peptide, endomorphin-2, and the endogenous delta-selective opioid peptide, Leu-enkephalin, were tested for their ability to affect spatial learning in the Morris water task. It was found that microinjection of 10 nmol endomorphin-2 into the CA3 region of the rat hippocampus significantly impaired spatial learning. However, the two lower doses tested, 3.3 and 1 nmol, had no effect in this test. Leu-enkephalin did not have any effect on spatial learning at the two doses tested (10 and 3.3 nmol). Neither peptide had any effect on motor performance as measured by swim speed. The results indicate that mu-receptors in the CA3 region of the rat hippocampus are more relevant than delta-receptors for spatial learning. PMID- 11117468 TI - Distribution of the 4F2 light chain, LAT1, in the mouse brain. AB - LAT1 (the 4F2-light chain 1) is a component of the CD98 surface antigen which participates in multiple functions including amino acid transport, cell survival, cell fusion and integrin activation. We examined the distribution of LAT1 in the mouse brain using immunohistochemistry. LAT1 was expressed on the microvessels, the subfornical organ, the subcommissural organ, ventromedial nucleus of the hypothalamus, subgranular zone of the dentate gyrus, ependymal layer of the lateral ventricles, and the olfactory bulb. LAT1-positive cells incorporated bromodeoxyuridine in the latter three regions where neurogenesis occurs in adult mouse brain, indicating that LAT1 is involved in neuronal cell proliferation, as well as in amino acid transport in these regions. PMID- 11117469 TI - EEG spectral activity during paradoxical sleep: further evidence for cognitive processing. AB - Paradoxical sleep (PS), in which periods with (phasic) and without (tonic) rapid eye movements are intermingled, is hypothesized to be related to cognitive processing and dreaming. Based on polysomnographic data from 12 healthy subjects, this study focuses on the spectral differentiation between phasic and tonic periods. Phasic PS periods exhibited decreased theta and alpha power in the posterior brain areas suggesting the interference of visual processing related to dream imagery. Phasic PS periods were also characterized by a shift from beta to gamma activity in frontal, central and occipital areas reflecting specific phasic related activation. Together, these findings bring new evidence for the existence of visual imagery and cognitive processing during phasic PS. PMID- 11117470 TI - Neuregulin is associated with nerve regeneration in axonal neuropathies. AB - Neuregulins (NRG), the only trophic factors known to support Schwann cell survival, have a potential for use in treatment of patients with chronic neuropathies. Expression of one NRG isoform, NRG-beta1, was increased in axons in the early phase of axonal degeneration in nerve biopsy specimens from patients with neuropathy. Regenerating nerve fibers contained abundant NRG-beta1 in axons, but nerves from patients with chronic axonal neuropathy showed less. NRG-beta1 may act as an axonally derived signal having a trophic effect on denervated Schwann cells, facilitating their supportive role in axonal regeneration. PMID- 11117471 TI - Hypertrophic A10 dopamine neurones in a rat model of attention-deficit hyperactivity disorder (ADHD). AB - To clarify whether a hypo or hyperfunctioning mesocorticolimbic system is the neural substrate of Attention-Deficit Hyperactivity Disorder (ADHD), we carried out a morphometric analysis on an animal model, the Naples high excitability rat (NHE). Male adult NHE and control rats were used for tyrosine hydroxylase (TH) immunocytochemistry in the ventral tegmental area and substantia nigra in coronal cryostat sections. PC-assisted image analysis showed larger DA neurones in the ventral tegmental area but not in the substantia nigra of NHE rats than in controls, associated with a higher expression of TH in the neuropil. Thus, the increased activity and impaired attention of NHE rats are associated with a hyperfunctioning mesocorticolimbic system in this ADHD model. PMID- 11117472 TI - Reduced reward processing in the brains of Parkinsonian patients. AB - Regional cerebral blood flow (rCBF) in healthy controls and non-demented, non depressed Parkinsonian patients was measured using H2(15)O PET while subjects performed a prelearned pattern recognition task with delayed response. To investigate differences between the two groups in response to reward, the experimental design consisted of three reinforcement conditions: no reinforcement consisting of nonsense feedback, positive symbolic reinforcement and monetary reward. In the controls, monetary reward activated bilaterally the striatum and anterior cingulate gyrus, as well as unilaterally the left cerebellum, midbrain and medial frontal gyrus. Symbolic reinforcement revealed a similar pattern of activation, except that the striatum and left midbrain showed no activation. The Parkinsonian patients responded to monetary reward with increased activation bilaterally in the cerebellum, medial frontal gyrus, and anterior cingulate gyrus as well as unilaterally in the right fusiform gyrus and midbrain and left caudate nucleus and precentral gyrus. Symbolic reinforcement induced significantly increased rCBF in the right cerebellum only. Compared with symbolic reinforcement, monetary reward produced extended activation of temporoparietal association cortex. The pattern observed in the controls demonstrates the role in reward processing of dopaminergic mesolimbic pathways in the healthy human brain, whereas the pattern in the Parkinsonian patients suggests the involvement of compensatory cortical loops in the diseased brain. PMID- 11117473 TI - Nociceptin/orphanin FQ inhibits ischaemia-induced glutamate efflux from rat cerebrocortical slices. AB - Nociceptin/orphanin FQ (NC), the endogenous ligand for the G-protein coupled nociceptin receptor (NCR), has a modulatory role in various physiological processes including neurotransmitter release. We have examined the effects of NC, the analogues NC(1-13)NH2 and [F/G]NC(1-13)NH2 and the competitive antagonist [Nphe1]NC(1-13)NH2 (Nphe1) on glutamate efflux during an acute simulated ischaemic challenge in rat cerebrocortical slices. The increase in glutamate efflux seen with ischaemia was inhibited by NC (EC50 250 nM). At micromolar concentrations, the analogues were found to have a similar effect on glutamate efflux compared to NC. In all cases, inhibition of glutamate efflux was abolished by Nphe1 (30 microM). These results suggest a neuroprotective action for NC. PMID- 11117474 TI - Nicotinic receptor subunit mRNA in the thalamus of the rat: relevance to schizophrenia? AB - Recent evidence suggests that aberrant nicotinic receptor (nAChR) expression plays an important role in schizophrenia. The present study sought to examine the distribution of nAChRs within the thalamus and associated cholinergic structures by examining nAChR subunit mRNA expression using in situ hybridization histochemistry. Transcripts for alpha4 and beta2 subunits were found in high levels in all thalamic nuclei and at lower levels in cholinergic nuclei (PPTg and MS/VDB). Distribution of mRNA encoding for additional subunits was restricted; lower alpha3 subunit transcript levels were detected in the anterior thalamic nuclei and portions of the lateral and posterior thalamic nuclei, with alpha7 transcripts being detected in cholinergic nuclei, the IMD and very low levels in the RTN. Low levels of alpha6 and beta3 transcript were found only within the RTN. PMID- 11117475 TI - Evidence for dual effects of nitric oxide in the forced swimming test and in the tail suspension test in mice. AB - L-Arginine (L-Arg), a substrate for nitric oxide synthase (NOS) at a dose of 250 500 mg/kg, i.p., significantly reduced the duration of immobility both in the forced swimming test (FST) and in the tail suspension test (TST), two models of depression in mice, without changing locomotion in an open field. Paradoxically, a similar effect was observed with the administration of N(G)-nitro-L-arginine (L NNA) (0.3-10 mg/kg, i.p.), an inhibitor of NOS. However, higher doses of L-Arg (750-1000 mg/kg) and L-NNA (30 mg/kg) did not produce any anti-immobility effect in FST and TST. The inactive isomers D-Arg (100-1000 mg/kg, i.p.) and D-NNA (0.3 30 mg/kg, i.p.) did not affect immobility duration in either the FST and TST. Preadministration of L-NNA (30 mg/kg, i.p.), but not of D-NNA completely blocked the anti-immobility effect of L-Arg (500 mg/kg, i.p.) in the FST. Similarly, L Arg (750 mg/kg, i.p.), but not D-Arg blocked the anti-immobility effect of L-NNA (3 mg/kg, i.p.) in the FST. The results indicate that either the synthesis of NO or the inhibition of its synthesis may produce antidepressant-like effects when assessed in the FST and TST. The physiological meaning of this finding is still obscure, but it could indicate that NO has a dual role in the modulation of depression. PMID- 11117476 TI - The effect of deviant stimulus probability on the human mismatch process. AB - The present study addresses the separate activities of frontal and temporal MMN generators which might be differentially affected by a change in the probability of standard stimuli. As the probability of standard stimuli was increased, the frontal MMN component significantly increased in amplitude, while the temporal one was not affected. Correspondingly, the scalp current density (SCD) maps showed that the temporal MMN generator was activated even at low probability of standard stimuli, suggesting that even the weak memory trace could start the automatic mismatch process, whereas the frontal MMN generator was activated only with increased probabilities of standard stimuli, suggesting that the stronger the memory trace is, the easier it might trigger the involuntary switching of attention to stimulus change. PMID- 11117477 TI - A readily releasable pool of single inhibitory boutons in culture. AB - The number of presynaptic vesicles that are immediately available for release, the readily releasable pool (RRP), is a strong determinant of synaptic strength and plasticity. The properties of the RRP in individual GABAergic synapses were examined in superior colliculus cultures. The RRP was depleted by high frequency trains and cumulative evoked IPSC amplitudes (CA) were calculated. The amplitude of monoquantal responses (q) was determined on the basis of mIPSC histograms. On average, the RRP, defined as CA/q, comprised about 10 vesicles. About 60% of the RRP could be released by a single stimulus. After depletion, the RRP was replenished with a time constant of about 14 s. These data provide information for further studies on the capacity of individual inhibitory synapses to modulate sensory information transfer. PMID- 11117478 TI - Deviant neurophysiological asymmetry in children with language impairment. AB - Deviant anatomical asymmetry of perisylvian cortex is argued to be linked to specific language impairment (SLI). However, no studies have examined whether deviant functional asymmetry underlies the processing of spoken language. In the current study, brain-electrical activity was recorded from 31 scalp sites to the function word 'the' embedded in auditorally presented stories and nonsense contexts. The SLI children showed reversed asymmetry at electrode sites over temporal cortex compared to control children in processing this word in all contexts. They also appear to lack some contribution from a deep neural generator in processing 'the' in the story. This investigation is the first to demonstrate a direct link between deviant neurophysiological asymmetry and the processing of spoken language in children with SLI. PMID- 11117479 TI - Expression of PEP-19 inhibits apoptosis in PC12 cells. AB - PEP-19 is a calmodulin-regulatory protein found specifically within neurons, though cellular functions of this protein have not been determined. In an effort to define potential effects of PEP-19, PC12 cell lines expressing this protein were generated and subjected to apoptotic stimuli. As measured by LDH release, cell death in PEP-19 expressing cells was 2- to 5-fold less following u.v. irradiation, and 2- to 4-fold less following staurosporine treatment than controls. Additionally, PEP-19-expressing cells displayed decreased DNA ladder formation, chromatin and condensation, caspase activation following staurosporine treatment. Overall, these results demonstrate that PEP-19 can inhibit apoptotic processes in PC12 cells, suggesting a potential regulatory mechanism for pathways leading to cell death. PMID- 11117480 TI - Neuronal ageing in long-term cultures: alterations of Ca2+ homeostasis. AB - Deficiencies of Ca2+ homeostasis are proposed to play an important role in neuronal ageing and/or neurodegeneration. The aim of this study was to investigate, in a defined neuronal population, primary cerebellar granule neuron culture, the time-dependent changes in Ca2+ homeostasis and compare them with data obtained in cerebellar brain slices from aged rats. In neurons aged in culture (DIV 23), a small decrease in the resting [Ca2+]i was associated with a decrease in the maximal rate of [Ca2+]i increase upon KCl-induced depolarization and in the amplitude of the [Ca2+]i response, when compared with mature neurons (DIV 9). The most significant change of [Ca2+]i signal parameters was a 50% decrease in the rate of [Ca2+]i recovery after the stimulation. These results were similar to those obtained in aged brain slices, and indicate that primary neuronal cultures could serve as a model for studying the age-related changes in Ca2+ homeostasis. PMID- 11117481 TI - Fast temporal interactions in human auditory cortex. AB - The temporal resolution of the human primary auditory cortex (AC) was studied using middle-latency evoked fields. Paired sounds with either the same or different spectral characteristics were presented with gaps between the sounds of 1, 4, 8 and 14 ms. Spatio-temporal modelling showed (1) that the response to the second sound was recognizable with gaps of 1 ms and rapidly increased in amplitude with increasing gap durations, (2) an enhanced N40m amplitude at gaps > 4 ms, (3) delayed N19m-P30m latencies when the stimuli were different. The median psychoacoustical thresholds were 1.6 ms for the same stimuli and 2.5 ms for different stimuli, confirming the electrophysiological evidence for rapid pattern specific temporal processing in human primary auditory cortex. PMID- 11117482 TI - Lewy body variant of Alzheimer's disease: alpha-synuclein in dystrophic neurites of A beta plaques. AB - The contribution of alpha-synuclein accumulation in Alzheimer's disease (AD) plaques is currently a matter of scientific debate. In the present study antisera against the N- and C-terminus, the full-length protein and the central so-called non-amyloid component (NAC) domain of the alpha-synuclein protein were used to address this question in brains of cases with typical AD and of cases with the Lewy body (LB) variant of AD. In typical AD cases, none of the antisera revealed evidence for co-accumulation of alpha-synuclein with extracellular A beta peptides in plaques or in dystrophic neurites decorating the plaque core. Interestingly, cases with mixed pathology of the LB variant of AD revealed accumulation of alpha-synuclein in LBs and in dystrophic neurites of A beta plaques. PMID- 11117483 TI - Mitogen-activated protein kinase phosphatase-1 in olfactory neural regeneration. AB - Uniquely, olfactory neurons continuously replace themselves. Olfactory bulb ablation induces coordinated degeneration and regeneration in olfactory neuroepithelium; up-regulated growth factors bind to their receptors, initiating a phosphorylation cascade activating mitogen-activated protein kinases (MAPK). MAPK then relay proliferation signals to the nucleus. Mitogen-activated protein kinase phosphatase-1 (MKP-1) inactivates MAPK. We examined MKP-1 expression in adult mouse olfactory epithelium following bulbectomy using a reverse transcription-polymerase chain reaction (RT-PCR) as well as Western analysis. While MKP-1 expression was high in olfactory epithelium from control mice, it decreased greatly 2-3 weeks after bulbectomy in temporal association with increased tyrosine phosphorylation of MAPK. Thus, reduced MKP-1 expression may contribute to regeneration of olfactory neuroepithelium after bulbectomy via decreased dephosphorylation of MAPK. PMID- 11117484 TI - Tachykinin-induced stimulation of neuropeptide Y gene expression in the rat arcuate nucleus. AB - Previous neurocytochemical data indicate the presence of synaptic contacts between tachykinergic terminals and neuropeptide Y (NPY) neurons in the arcuate nucleus of the rat suggesting that tachykinins may regulate NPY neuronal activity. To examine the functional signification of such regulation, the effect of intracerebroventricular administration of neurokinin A on NPY mRNA levels was studied using in situ hybridization. Repeated treatment with NKA (40 microg/day for 3 days) induced a 44% increase in NPPY mRNA expression compared with saline injected control animals. These results demonstrate a positive effect of tachykinins on NPY gene expression and suggest either a direct or indirect control of arcuate NPY neurons by endogenous tachykinins. PMID- 11117485 TI - N-Acetylaspartate and DARPP-32 levels decrease in the corpus striatum of Huntington's disease mice. AB - Huntington's disease (HD) is an autosomal dominant condition involving progressive neurodegeneration, primarily the corpus striatum and cerebral cortex. We have used in vivo magnetic resonance spectroscopy (MRS) to assess specific neuronal markers in transgenic mice (R6/1 line) expressing exon I of the human huntingtin gene with an expanded CAG repeat. Levels of N-acetylaspartate (NAA), an indicator of healthy neuronal function, were significantly reduced (26%) in the corpus striatum of HD mice relative to wild-type littermates at 5 months of age. However, levels of cholines and creatine-phosphocreatine were not altered in the HD mice. Expression of dopamine- and cAMP-regulated phosphoprotein, 32 kDa (DARPP-32), was assessed by immunohistochemistry in the striatum of HD mice and found to be downregulated by 5 months and, even more dramatically, at 11 months of age. In contrast, expression of calbindin was not significantly decreased in HD mice. Our results suggest that the observed decreases in DARPP-32 and NAA may contribute to aberrant receptor signalling and neuronal dysfunction in HD. PMID- 11117486 TI - Optical detection of embryogenetic expression of vagal excitability in the rat brain stem. AB - We traced and identified the ontogenetic expression of neural excitability related to the vagus nerve in the embryonic rat brain stem. Multiple-site optical recordings of neural activities revealed two response areas in the E12 rat brain stem: one corresponding to the dorsal motor nucleus of the vagus nerve, and the other reflecting the activities of sensory nerve fibers. In embryos younger than E11, no optical response was identified, suggesting that excitability of the motoneurons and/or sensory nerve fibers is first generated no later than E12. A contour line map of the neural responses suggested that, in contrast to older embryos, the functional organization of the vagal nucleus is not orderly at the time of the initial expression of neural excitability. PMID- 11117487 TI - The masking effect in foreign speech sounds perception revealed by neuromagnetic responses. AB - The backward masking effect on non-native consonants by a following vowel was examined using neuromagnetic responses to synthesized speech sounds. Native speakers of Japanese were presented with sequences of frequent (85%) and infrequent (15%) speech sounds (/ra/ and /la/ respectively, no /l/ /r/ contrast in Japanese language). The duration of the stimuli was 110 ms in a short session and 150 ms in a long session. In the short session, the stimuli were terminated in the course of the transition from the consonant to the vowel to diminish the masking effect from the vowel part. A distinct magnetic counterpart of mismatch negativity (MMNm) was observed for the short session, whereas a smaller MMNm was observed for the long session. PMID- 11117488 TI - Do perceived loudness cues contribute to duration mismatch negativity (MMN)? AB - This study explored the contribution of perceived loudness cues to mismatch negativity produced in response to a 125 ms duration deviant tone among a regular sequence of 50 ms standard tones. Each individual was required to adjust the intensity of a 125 ms tone until it matched the perceived loudness of a 50 ms tone. The mismatch negativity produced to the duration deviant presented at the same intensity as the standard was contrasted with that produced to the same deviant at each individual's perceived loudness equivalence. Despite detectable difference perceived loudness (approximately 1.3 dB), adjusting the intensity of the deviant tone did not significantly reduce mismatch negativity amplitude to the duration deviant. Results are discussed with reference to temporal window of integration. PMID- 11117489 TI - Increased hippocampal supragranular Timm staining in subjects with bipolar disorder. AB - Biochemical and structural abnormalities have been reported in hippocampus of subjects with mood disorders. This study examined the organization of mossy fibers in anterior hippocampus of subjects obtained from the Stanley Neuropathology Consortium. Frozen postmortem hippocampal sections from subjects with major depression, bipolar disorder, schizophrenia and non-psychiatric controls were stained using the Neo-Timm procedure, which selectively stains mossy fibers. Increased Timm staining in the supragranular layer was found in subjects with bipolar disorder relative to control subjects. These results are suggestive of neuronal sprouting in hippocampus of subjects with bipolar disorder. There were no significant associations between supragranular Timm staining and suicide, length illness or drug treatment at the time of death. PMID- 11117490 TI - Cl(-)-dependent excitotoxicity is associated with 3H2O influx in chick embryonic retina. AB - The aim of this study was to show that Cl(-)-dependent excitotoxicity, with its characteristic cell swelling, involves actual water influx into the intracellular compartment. Taking advantage of the Ca2+ omission paradigm of Cl(-)-dependent excitotoxicity, in the chick embryonic neural retina ex vivo, which is associated with toxicity levels (lactate dehydrogenase (LDH) release) considerably higher than those seen after simple exposure of the retinas to glutamate agonists, we have demonstrated that an intracellular water intake of 4.2 microl into retinal cells is associated with 13.3% total retinal LDH release. The fact that mannitol blocks both water inflow and LDH release appears to link both events from a pathogenic point of view. PMID- 11117491 TI - The CNS updates its context estimate in the absence of feedback. AB - Human motor behaviour is remarkably accurate and appropriate even though our bodies and the objects we interact with change over time. To achieve such performance the motor system has to tailor the motor commands to the current context, that is the properties of objects in the world and the prevailing environmental conditions. The current context can be estimated by integrating two sources of information, sensory feedback and knowledge about how the context is likely to have changed from the previous estimate. Here we show that in the absence of sensory feedback about the context the second process is able to extrapolate the likely evolution of the context without requiring awareness that the context is changing. PMID- 11117492 TI - Different coupling for the reach and grasp components in bimanual prehension movements. AB - In this study on functional coupling in bimanual grasping movements, nine normal subjects had to reach and grasp two different objects simultaneously. Both objects could be either small or large, resulting in four different conditions of bimanual grasping. The main dependent variables were the coupling coefficients calculated either between the hand displacements or between the grip apertures of both hands, serving as variables for the coupling of the reach and the grasp component respectively. The correlation was significantly higher for the reach component than for the grasp component, with only the latter one changing significantly with variation of object size. These findings suggest different temporo-spatial coupling modes for the reach and the grasp components of bimanual grasping movements. PMID- 11117493 TI - Congenital helpless rats as a genetic model for cortex metabolism in depression. AB - The validity of congenital helplessness as a genetic rat model for human depression was investigated in cortical regions of the rat brain thought to be analogous to those showing abnormalities in human neuroimaging studies. Cortex metabolism was analyzed using quantitative cytochrome oxidase histochemistry. Congenital helpless rats showed changes in frontal and cingulate regions comparable to those that have demonstrated metabolic differences in human depression. Significant metabolic decreases were found in dorsal frontal, medial orbital, and anterior cingulate, whereas a significant increase was found in infraradiata (subgenual) cingulate. The direction of these changes were the same as those seen in human studies. These findings support the validity of congenital helplessness as a model for human depression. PMID- 11117494 TI - Immunohistochemical localization of the P2Y1 purinergic receptor in Alzheimer's disease. AB - The biological actions of extracellular nucleotides are mediated by two distinct classes of P2 receptor, P2X and P2Y. The G protein-coupled P2Y receptors comprise five mammalian subtypes, P2Y(1-11). The P2Y1 subtype is expressed abundantly throughout the human brain and is specifically localized to neuronal structures. In the present study, the distribution of the P2Y1 receptor was investigated in Alzheimer's disease (AD) brains. In contrast to control human brain, the P2Y1 receptor was localized to a number of characteristic AD structures such as neurofibrillary tangles, neuritic plaques and neuropil threads. Immunoblot analysis showed that this specific immunostaining observed over tangles was not a result of cross-reactivity between the anti-P2Y1 antiserum and abnormal tau protein, the major constituent of tangles. The significance of this altered P2Y1 cellular distribution in AD brains is at present unclear. PMID- 11117495 TI - Neuroma: a donor-age independent source of human Schwann cells for tissue engineered nerve grafts. AB - Schwann cells are used in combination with biological matrices as tissue engineered nerve grafts in animal models offering a new therapeutic approach for treatment of lesions of the peripheral nervous system. A high yield of human Schwann cells from adult donors is only achieved by pharmacological stimulation, which should, however, be avoided in clinical therapy. Here, we establish cultures of activated human Schwann cells which were isolated from peripheral nerve neuroma which developed after a median nerve lesion. To allow nerve reconstruction neuroma have to be resected. Such neuroma tissue is virtually predegenerated and shows activation of Schwann cells, implying good adherence and high mitotic activity. This allows, irrespective of donor age, growing within a short time period and without any pharmacological treatment. PMID- 11117496 TI - Brain and CSF entry of the novel non-nucleoside reverse transcriptase inhibitor, GW420867X. AB - (S)-2-ethyl-7-fluoro-3-oxo-3, 4-dihydro-2H-quinoxaline-carboxylic acid isopropylester (GW420867X) inhibits HIV-1 reverse transcriptase and could be used for the treatment of HIV infection. This study quantified the movement of [14C]GW420867X into the CNS by means of a guinea-pig brain perfusion technique. Results indicated that [14C]GW420867X can enter the brain (Kin: 38.4+/-7.7 microl min(-1) g(-1)) and cerebrospinal fluid (CSF; Kin: 1.2+/-0.1 microl min(-1) g( 1)). Self-inhibition studies also suggested the presence of a saturable transport system for [14C]GW420867X at the blood-brain barrier (BBB). Thus [14C]GW420867X can enter the brain via the BBB and, compared with the blood-CSF barrier, this route is the predominant pathway for the brain entry of this drug. This would suggest that GW420867X is a promising drug for the treatment of HIV infection within the brain. PMID- 11117497 TI - GABA(B(1)) splice variant mRNAs are differentially affected by electroshock induced seizure in rats. AB - Receptor autoradiography with the high affinity antagonist radioligand [3H]CGP62349 and in situ hybridization with radiolabelled oligonucleotides were used to investigate GABA(B) receptor protein expression, and GABA(B(1)) mRNA splice variant (GABA(B(1a)) and GABA(B(1b)) levels, in brain sections from rats 4 h following a single electroshock-induced generalized seizure. Densitometric analysis indicated that GABA(B(1a)) mRNA levels were not significantly altered by an acute electroshock seizure, but that GABA(B(1b)) mRNA levels were significantly increased throughout the brain. GABA(B) receptor expression at this time point was unaffected by the seizure. The observed up-regulation of GABA(B(1b)) mRNA levels may imply increased importance of this splice variant in the regulation of further seizure activity. PMID- 11117498 TI - Cx36 is dynamically expressed during early development of mouse brain and nervous system. AB - Connexins are structural proteins that are part of the gap junctional channels which couple cells in different tissues. Connexin36 (Cx36) is a new member of the connexin gene family, found to be expressed essentially if not exclusively in neuronal cells in adult CNS of mouse, rat and man. Here we have studied Cx36 expression during murine embryonic development. Cx36 shows a highly dynamic pattern of expression. It is first (E9.5) evident in the forebrain and later its expression expand caudally in the midbrain. At E12.5 its expression correlates with major morphogenetic boundaries in the developing mouse brain, specifically with the dorsoventral telencephalic boundary and the Zona Limitans Intrathalamica. Starting at midgestation (E12.5), it is also expressed in both sympathetic and spinal ganglia, and in two longitudinal stripes along the spinal cord. PMID- 11117499 TI - The neural basis of romantic love. AB - The neural correlates of many emotional states have been studied, most recently through the technique of fMRI. However, nothing is known about the neural substrates involved in evoking one of the most overwhelming of all affective states, that of romantic love, about which we report here. The activity in the brains of 17 subjects who were deeply in love was scanned using fMRI, while they viewed pictures of their partners, and compared with the activity produced by viewing pictures of three friends of similar age, sex and duration of friendship as their partners. The activity was restricted to foci in the medial insula and the anterior cingulate cortex and, subcortically, in the caudate nucleus and the putamen, all bilaterally. Deactivations were observed in the posterior cingulate gyrus and in the amygdala and were right-lateralized in the prefrontal, parietal and middle temporal cortices. The combination of these sites differs from those in previous studies of emotion, suggesting that a unique network of areas is responsible for evoking this affective state. This leads us to postulate that the principle of functional specialization in the cortex applies to affective states as well. PMID- 11117500 TI - Acute marijuana effects on rCBF and cognition: a PET study. AB - The effects of smoking marijuana on cognition and brain function were assessed with PET using H2(15)O. Regional cerebral blood flow (rCBF) was measured in five recreational users before and after smoking a marijuana cigarette, as they repeatedly performed an auditory attention task. Blood flow increased following smoking in a number of paralimbic brain regions (e.g. orbital frontal lobes, insula, temporal poles) and in anterior cingulate and cerebellum. Large reductions in rCBF were observed in temporal lobe regions that are sensitive to auditory attention effects. Brain regions showing increased rCBF may mediate the intoxicating and mood-related effects of smoking marijuana, whereas reduction of task-related rCBF in temporal lobe cortices may account for the impaired cognitive functions associated with acute intoxication. PMID- 11117501 TI - The variability of serial fMRI data: correlation between a visual and a motor task. AB - We investigated whether the intersession variability of serial fMRI studies correlates between two activation modalities, i.e. a standardized visual and a standardized motor task. Six volunteers were scanned in at least weekly intervals. The number of pixels activated as well as the activation amplitude varied widely. The maximal difference of the number of pixels activated was 1150%, of the activation amplitude 250%. In three volunteers, the variability was highly correlated between the two tasks. Three other volunteers showed one outlier each. We conclude that the intersession variability is due to global factors affecting the whole brain, but that due to unpredictable outliers, using a standardized task to normalize the data of interest is of limited value. PMID- 11117502 TI - Both sides of human cerebellum involved in preparation and execution of sequential movements. AB - Using an event-related fMRI procedure, we investigated the role of the human cerebellum in sequential finger movements. Subjects performed a delayed sequential finger movement task in which an instructive cue preceded the imperative signal by 16.5 s. Bilateral activation was observed in the cerebellum following both the cue and imperative signals. The activated regions overlapped within the cerebellum, extending across intermediate and lateral regions corresponding to lobules HV-HVII. In contrast, activation in primary motor cortex was primarily restricted to the execution phase and most prominent in the contralateral hemisphere. These results indicate that the cerebellum is bilaterally recruited for the preparation and execution of sequential movements. PMID- 11117503 TI - Ca2+-dependent kainate excitotoxicity in the chick embryonic neural retina ex vivo. AB - The chick embryonic neural retina ex vivo has been singled out as a unique example of Cl(-)-dependent/Ca2+-independent excitotoxicity. However, after continuous incubation with 100 microM kainate, we have demonstrated the susceptibility of the chick retina to Ca2+-mediated damage, which becomes apparent after 12 h of exposure to the agonist in the absence of Cl-. Of the 20.8% lactate dehydrogenase released after 24 h incubation with kainate, some 11% is Cl(-)-dependent and the rest (9.8%) is presumably Ca2+-dependent. Upon omission of both Cl- and Ca2+, a 5% residual toxicity can still be detected after 24 h. This can be overcome by inclusion of EGTA in the incubation medium to neutralize Ca2+ released during incubation. A Ca2+-dependent toxicity mechanism is then operative in the embryonic chick retina ex vivo. PMID- 11117504 TI - Different regulatory elements are necessary for alpha1 tubulin induction during CNS development and regeneration. AB - Developing and regenerating neurons induce genes whose products are necessary for axonal growth, such as that encoding alpha1 tubulin. To determine whether alpha1 tubulin gene induction uses similar mechanisms during CNS development and regeneration, we compared wild-type and mutant alpha1 tubulin promoter activity in the developing and regenerating CNS of transgenic zebrafish. Wild-type alpha1 tubulin promoter activity increased dramatically in the developing and regenerating CNS. In contrast, we generated a mutation in the alpha1 tubulin promoter that prevented its increase during development but retained regeneration dependent induction in the adult. These results suggest that at least some of the signaling mechanisms used to activate alpha1 tubulin promoter activity during CNS regeneration are different from those used to activate this promoter during development. PMID- 11117505 TI - Effect of age on burst firing characteristics of rat hippocampal pyramidal cells. AB - During behavior, hippocampal pyramidal cells emit high frequency bursts, modulated by the animal's location and the 7 Hz theta rhythm. During rest, CA1 EEG exhibits large irregular activity (LIA), containing sharp-wave/ripple complexes, during which pyramidal cells exhibit burst discharge. Aging results in altered intracellular calcium homeostasis, increased electrical coupling and reduced cholinergic modulation within CA1, all of which might affect burst discharge characteristics. During LIA, old rats exhibited more short (3-7 ms) inter-spike intervals, with no change in mean firing rate. During behavior induced theta rhythm, however, interval distributions were not affected by age. Thus, different mechanisms must underlie burst discharge in theta and LIA states. Moreover, age related changes in the cholinergic system appear not to play a major role in shaping the temporal discharge characteristics of CA1 pyramidal cells. The mechanism and significance of the higher frequency bursting in old rats during LIA remains to be determined. PMID- 11117506 TI - Effects of brain-derived neurotrophic factor and neurotrophin-3 on expression of mRNAs encoding c-Fos, neuropeptides and glutamic acid decarboxylase in cultured spinal neurons. AB - There is growing evidence suggesting that neurotrophins have modulating effects on the pain signaling system at spinal levels. In order to determine whether neurotransmitter expression is modulated in response to the elevation of neurotrophins, the changes in c-fos, neuropeptide and glutamic acid decarboxylase (GAD) mRNAs expression was evaluated after BDNF or NT-3 was applied to cultured spinal neurons. Reverse transcription polymerase chain reaction analysis revealed that BDNF induced a significant increase in the expression of preprodynorphin (pDYN), preproenkephalin (pENK), neuropeptide Y (NPY) and GAD mRNAs. In contrast, the pENK, not the pDYN, NPY and GAD, mRNA levels increased after the treatment of NT-3. Both BDNF and NT-3 produced a rapid increase in c-fos mRNA. These results suggest that BDNF and NT-3 have differential neuronal effects on the synthesis of spinal cord neurotransmitters that are involved in the modulation of nociceptive information. PMID- 11117507 TI - Up-regulation of glial cell line-derived neurotrophic factor (GDNF) following traumatic spinal cord injury. AB - We investigated the temporal and spatial expression patterns of the GDNF gene after subjecting rats to an acute contusion injury of the spinal cord using the weight drop technique. Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that GDNF transcription in the spinal cord began to increase within 30 min after injury and peaked within 3 h. Immunohistochemical analysis showed GDNF immunoreactivity to be present mainly in microglia and macrophages 1 day after injury, but not in neurons or astrocytes. This immediate upregulation of GDNF gene expression may be a component of an inflammatory process and probably exerts a protective effect on neurons following spinal cord injury (SCI). PMID- 11117508 TI - A novel nitrate ester reverses the cognitive impairment caused by scopolamine in the Morris water maze. AB - The objective of this study was to test the hypothesis that activation of soluble guanylyl cyclase and increased cGMP formation in the brain would improve task acquisition in cognitively impaired animals. We evaluated the effects of a novel nitrate ester, GT 715 (2,3-dinitrooxy-(2,3-bis-nitrooxypropyldisulfanyl) propane), in scopolamine-induced impairment of task acquisition in the Morris water maze. GT 715 improved task acquisition in scopolamine-pretreated animals in a time- and dose-dependent manner, whereas the prototypical nitrate ester, glyceryl trinitrate (GTN), was ineffective. GT 715 also was more effective and more potent than GTN for activation of hippocampal guanylyl cyclase. The results of this study therefore suggest that stimulation of cerebral soluble guanylyl cyclase activity may be an effective strategy to improve learning and memory performance in individuals in whom cognitive abilities are impaired by injury, disease, or ageing. PMID- 11117509 TI - Do Ca2+ channels share NMDA receptors in plasticity of synaptic transmission in the rat visual cortex? AB - We examined the involvement of Ca2+ channels in LTP of responses in rat visual cortex slices. Stimulating layer IV, field potentials including EPSP1 and EPSP2 from layer II/III were recorded. L-type Ca2+ channel blocker nifedipine did not have a considerable effect on LTP of the responses. T-type Ca2+ channel blocker Ni2+ decreased potentiation of EPSP1 and almost blocked that of EPSP2. Effect of visual experience on the function of the channels is also considered. These results indicate that T-type Ca2+ channels play a real role in stable LTP of EPSP2. Also the function of the channels was almost the same in dark and light reared visual cortices. PMID- 11117510 TI - Up-regulated CNTF plays a protective role for retrograde degeneration in the axotomized rat retina. AB - Using reverse transcription-polymerase chain reaction and in situ hybridization, we investigated the expression and cellular localization of ciliary neurotrophic factor receptor alpha (CNTFRalpha) in the rat retina following optic nerve transection (ONT). Following ONT, a signal for CNTFRalpha mRNA appeared in a layer-specific and time-dependent manner. In the ganglion cell layer, the signal showed a peak value 1 day after ONT, and then gradually decreased. In the inner nuclear layer the signal reached a peak value at 14 days of about 500% of control level, but then decreased at 4 weeks. Our findings suggest that CNTF might play a protective role for the retrograde degeneration of retinal cells induced by ganglion cell death in the rat retina following ONT. PMID- 11117511 TI - The effects of galvanic stimulation on the human vestibulo-ocular reflex. AB - We studied the effects of 5 mA bilateral or unilateral, bipolar or monopolar, galvanic stimulation on the horizontal vestibulo-ocular reflex (hVOR) in six normal subjects during 0.01, 0.05, 0.1, 0.5 and 1 Hz yaw rotations and in two subjects during high-acceleration, low-amplitude yaw head rotations (head impulses). Bipolar galvanic stimulation induced horizontal nystagmus in all subjects and an asymmetry of the hVOR only during rotations below 0.1 Hz. Monopolar stimulation had no significant effect. The findings suggest that in humans galvanic stimulation affects those primary horizontal semicircular canal neurons that mediate the hVOR via indirect pathways through the velocity storage mechanism. PMID- 11117512 TI - Acute effects of corticosterone on LiCl-induced rapid gustatory conditioning in rats: a taste reactivity analysis. AB - Previous research has shown that acute corticosterone treatment can have rapid effects on learning and memory. Using the taste reactivity test (TRT), the present study examined the effect of acute administration of corticosterone on sucrose palatability and the development of LiCl-induced rapid gustatory conditioning. On each of two conditioning days rats were injected with either a low dose of lithium chloride (LiCl; 0.75 mEq, i.p.) or saline (NaCl; 0.9%, i.p.) and 10 min later, received a second injection of either corticosterone (5 mg/kg, i.p.) or cyclodextrin vehicle. Rats were then placed in the TRT chamber, where 1 min intraoral infusions of sucrose (0.3 M) were delivered every 10 min. Taste reactivity responses were videotaped and later analyzed for frequency of occurrence. Rats treated with both LiCl and corticosterone showed enhanced aversive responding and reduced ingestive responding relative to control rats treated with LiCl and vehicle. The implication that corticosterone may have a rapid enhancing effect on gustatory conditioning is discussed. PMID- 11117513 TI - Galanin-like peptide (GALP) is expressed in rat hypothalamus and pituitary, but not in DRG. AB - Galanin-like peptide (GALP) was recently purified on the basis of its preferential activation of galanin receptor subtype 2 (GALR2) compared with galanin receptor subtype 1 (GALR1). Using in situ hybridization of adult rat brain, pituitary and dorsal root ganglia (DRG) we demonstrate that GALP mRNA expression is restricted to the arcuate nucleus and median eminence of the hypothalamus, and to the posterior lobe of the pituitary. No expression was detected elsewhere in brain, or in the DRG. In adult mouse, no expression was detected in brain or in DRG either before or after axotomy, suggesting that GALP has no apparent role in the axotomy response of DRG. PMID- 11117514 TI - Limited auditory memory for conspecific songs in a non-territorial songbird. AB - Males of territorial songbird species have to remember a large number of conspecific songs to defend their territories, while non-territorial songbirds do not need to. A study of a territorial species suggested seemingly unlimited auditory memory size. We measured auditory memory in Bengalese finches, a non territorial songbird species, to examine whether the auditory memory size for conspecific songs depends on the ecological requirements for song use. Five birds were trained by operant techniques to classify song stimuli into two arbitrary categories. The learning curve reached an asymptote within approximately 100 sessions in all five birds and only eight songs were concurrently remembered on average. Results suggest that ecological requirements for song use are correlated with the auditory song memory capacity. PMID- 11117515 TI - NT-4/5 reduces cell death in inner nuclear as well as ganglion cell layers in neonatal rat retina. AB - Using the TUNEL method, we examined the effect of intraocular NT-4/5 injections on cell death in ganglion and non-ganglion cell layers in 5-day-old rat retinas. NT-4/5 reduced the level of naturally occurring cell death in all retinal layers. Twenty-four hours after superior colliculus (SC) lesions there was a significant increase in the density of TUNEL+ profiles in the RGC layer (6.43/mm2 in normal vs (8.89/mm2 after lesions) which was ameliorated by intraocular NT-4/5 injections (8.79/mm2). Surprisingly, after SC ablation a significant increase in TUNEL+ profiles was also seen in non-ganglion cell layers (52.25/mm2 in normal vs 89.35/mm2 after lesions), mostly in the developing inner nuclear layer. Death in non-ganglion cell layers was also significantly reduced (43.09/mm2) after NT-4/5 eye injections. PMID- 11117516 TI - Alteration of the somatosensory cortical map in peripheral mononeuropathy due to carpal tunnel syndrome. AB - Substantial plasticity of the mature mammalian somatosensory cortex was demonstrated after deprivation of sensory input produced by amputation or somatosensory deafferentation. Following transection of the median nerve, adult owl and squirrel monkeys exhibit extensive reorganization in the cortical representation of the hand in areas 3b and 1. In the present study we investigated the possible effect of incomplete median nerve damage on sensory cortex somatotopy in a patient with unilateral carpal tunnel syndrome. We assessed interhemispheric differences of the hand representation in SI by means of magnetic source imaging. Additional intersubject data comparison was performed for specific results on the basis of available normal data from the literature and from own investigations in five healthy volunteers. Our results demonstrated a decreased extension of the cortical zone representing the injured median nerve and suggested invasion of the deprived area by cortical sectors receiving inputs from the little finger (supplied by the ulnar nerve) and from the dorsum of the thumb (innervated by the radial nerve). The study indicates topographic rearrangement of the hand representational zone in the human primary somatosensory cortex in a case of chronic median nerve injury. PMID- 11117517 TI - PrP fragment 106-126 is toxic to cerebral endothelial cells expressing PrP(C). AB - A hydrophobic, fibrillogenic peptide fragment of human prion protein (PrP106-126) had in vitro toxicity to neurons expressing cellular prion protein (PrP(C)). In this study, we proved that primary cultures of mouse cerebral endothelial cells (MCEC) express PrP(C). Incubation of MCEC with PrP106-126 (25-200 microM) caused a dose-dependent toxicity assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay, lactate dehydrogenase release, bis benzimide staining for nuclear morphology, and trypan blue exclusion test. Pentosan polysulphate (50-100 microg/ml), a drug effective in scrapie prophylaxis, dose-dependently attenuated the injury. MCEC cultures from mice homogenous for the disrupted PrP gene were resistant to the toxicity of PrP106 126. In conclusion, cerebral endothelium expressing PrP(C) may be directly damaged during spongiform encephalopathies. PMID- 11117518 TI - Three neuroscientists to share Nobel Prize in physiology and medicine. PMID- 11117519 TI - Mandela speaks up for research into HIV in Africa. PMID- 11117520 TI - Creutzfeldt-Jakob disease: analysis of the MRI signal. PMID- 11117521 TI - Developmental evolutionary biology of the vertebrate ear: conserving mechanoelectric transduction and developmental pathways in diverging morphologies. AB - This brief overview shows that a start has been made to molecularly dissect vertebrate ear development and its evolutionary conservation to the development of the insect hearing organ. However, neither the patterning process of the ear nor the patterning process of insect sensory organs is sufficiently known at the moment to provide more than a first glimpse. Moreover, hardly anything is known about otocyst development of the cephalopod molluscs, another triploblast lineage that evolved complex 'ears'. We hope that the apparent conserved functional and cellular components present in the ciliated sensory neurons/hair cells will also be found in the genes required for vertebrate ear and insect sensory organ morphogenesis (Fig. 3). Likewise, we expect that homologous pre-patterning genes will soon be identified for the non-sensory cell development, which is more than a blocking of neuronal development through the Delta/Notch signaling system. Generation of the apparently unique ear could thus represent a multiplication of non-sensory cells by asymmetric and symmetric divisions as well as modification of existing patterning process by implementing novel developmental modules. In the final analysis, the vertebrate ear may come about by increasing the level of gene interactions in an already existing and highly conserved interactive cascade of bHLH genes. Since this was apparently achieved in all three lineages of triploblasts independently (Fig. 3), we now need to understand how much of the morphogenetic cascades are equally conserved across phyla to generate complex ears. The existing mutations in humans and mice may be able to point the direction of future research to understand the development of specific cell types and morphologies in the formation of complex arthropod, cephalopod, and vertebrate 'ears'. PMID- 11117522 TI - The PDX-1 activation domain provides specific functions necessary for transcriptional stimulation in pancreatic beta-cells. AB - PDX-1 is a homeodomain transcription factor whose targeted disruption results in a failure of the pancreas to develop. Mutations in the human pdx-1 gene are linked to an early onset form of non-insulin-dependent diabetes mellitus. PDX-1 binds to and transactivates the promoters of several physiologically relevant genes within the beta-cell, including insulin, glucose transporter 2, glucokinase, and islet amyloid polypeptide. This study focuses on the mechanisms by which PDX-1 activates insulin gene transcription. To evaluate the role of PDX 1 in transcription of the insulin gene, a chloramphenicol acetyltransferase reporter construct was designed with a single yeast GAL4-DNA binding site in place of the A3/PDX-1 binding element in the rat insulin II enhancer. In the presence of GAL4:PDX chimeras containing N-terminal transactivation domain sequences, this GAL4-substituted insulin construct was active in PDX-1-expressing beta-cells and not non-beta-cells. PDX-1 activation was mediated through three highly conserved segments of the transactivation domain. In addition, when cotransfected together with the GAL4-substituted insulin enhancer reporter gene in glucose-responsive MIN-6 beta-cells, glucose-induced activation is observed with GAL4:PDX-1 but not with fusions of the heterologous activation domains from herpes virus VP16 or adenovirus-5 E1A proteins. Using A3 element-substituted GAL4 insulin enhancer reporter constructs containing mutations in two additional key control elements, E1 and C1, we also show that full activation requires cooperative interactions between other enhancer-bound factors, particularly the E1 element activators. In contrast, the activity of the VP16 activation factor was not dependent on the activators of either the E1 or C1 sites. These results suggest that the PDX-1 transactivation domain is specifically required for appropriate regulation of insulin enhancer function in beta-cells. PMID- 11117523 TI - COUP-TFI (chicken ovalbumin upstream promoter-transcription factor I) regulates cell migration and axogenesis in differentiating P19 embryonal carcinoma cells. AB - The developmental expression patterns of the nuclear orphan receptors COUP-TFs (chicken ovalbumin upstream promoter-transcription factors) have been correlated to neurogenesis in several animal species. Nevertheless, the role of COUP-TFs in neurogenesis remains unknown. We have studied the functional involvement of COUP TFI in retinoic acid (RA)-induced neuronal differentiation of P19 embryonal carcinoma cells through two complementary approaches: 1) deregulated expression of COUP-TFI, and 2) inactivation of endogenous COUP-TFs by means of a dominant negative COUP-TFI mutant. Low levels of wild-type (wt)COUP-TFI transgene expression did not inhibit neural cell fate and primarily enhanced neuron outgrowth from RA-treated P19 aggregates. In contrast, high COUP-TFI expression impeded the neuronal differentiation of P19 cells induced with RA, resulting in cell cultures lacking neurons. This morphological effect was correlated to an elevated level of E-cadherin mRNA. The dominant-negative COUP-TFI mutant induced cell packing after RA treatment and inhibited neurite extension and neuron outgrowth from aggregates. A RGD peptide interference assay indicated that endogenous COUP-TFs could favor migration of neurons through an integrin dependent mechanism. Accordingly, vitronectin mRNA levels were shown to be up regulated by COUP-TFI by RT-PCR analysis, and COUP-TFI stimulated the mouse vitronectin promoter activity in transient transfection assays. Taken together, these data indicate that COUP-TFI is not simply a global repressor of retinoid functions, but shows a high selectivity for regulating genes involved in cellular adhesion and migration processes that are particularly important for neuronal differentiation. PMID- 11117524 TI - Inhibition of tumor growth by the antiangiogenic placental hormone, proliferin related protein. AB - Proliferin-related protein (PRP) is a potent placental antiangiogenic hormone. To test the antiangiogenic potential of PRP to block tumor growth, we engineered tumor cells to express this hormone. Both SV40-transformed BALB/c mouse 3T3 fibroblasts and rat C6 glioma cells have markedly reduced growth rates as tumors in mice if they express high levels of PRP. In both models, the small tumors that form are largely avascular, whereas control tumors are rich in blood vessels, consistent with PRP limiting tumor growth by preventing neovascularization of the tumors. The antiangiogenic effects of PRP are also detected on human endothelial cells, suggesting that the receptor and signaling pathway of this mouse hormone are conserved between mouse and human and may represent useful targets for the development of antiangiogenic therapeutics. That signaling pathway appears to involve an inhibition of arachidonic acid release, based on the ability of arachidonic acid to overcome the antiangiogenic effects of PRP. PMID- 11117525 TI - A missense mutation G2320R in the thyroglobulin gene causes non-goitrous congenital primary hypothyroidism in the WIC-rdw rat. AB - A convincing line of evidence is being developed that the congenital nongoitrous hypothyroidism and dwarfism observed in the WIC-rdw rat may indeed be caused by a primary defect in thyroid hormonogenesis. In support of this hypothesis, several recent reports have shown the presence of elevated molecular chaperone levels in the WIC-rdw thyrocytes, the endoplasmic reticulum of which was markedly dilated, suggesting a defect in intracellular protein transport. Here the studies were undertaken to identify the precise molecular defect in the WIC-rdw rat. First, the genetic linkage analysis revealed that the rdw locus was on rat chromosome 7 and was identical to the thyroglobulin (Tg) gene locus. Moreover, the Tg protein level was reduced in the WIC-rdw thyroid despite a similar level of the Tg gene transcripts that were indistinguishable in their size from the normal. Next, the complete sequencing of the rdw and the normal rat Tg cDNAs revealed a single nucleotide change, G6958C, resulting in a G2320R missense mutation in a highly conserved region of the Tg molecule. Finally, transient expression of the intact Tg cDNA containing the rdw mutation in the COS-7 cells showed no detectable Tg in the secreted media, indicating a severe defect in the export of the mutant Tg. Together, our observations suggest that a missense mutation, G2320R, in the Tg gene is responsible for the rdw mutation in the WIC-rdw rat. PMID- 11117526 TI - Ligand-activated progesterone receptor isoform hPR-A is a stronger transactivator than hPR-B for the expression of IGFBP-1 (insulin-like growth factor binding protein-1) in human endometrial stromal cells. AB - In human endometrium, the levels of progesterone receptor (PR) isoforms hPR-A and hPR-B are differentially regulated during the reproductive cycle. Progesterone significantly increases the content of hPR-A, the predominant isoform in decidualized stromal cells (1). The purpose of this study was to determine the capacity of hPR-A and hPR-B to transactivate the progestin-dependent target gene in human endometrial stromal cells. We examined the effect of cotransfection of hPR-A or hPR-B on the expression of the human insulin-like growth factor binding protein-1 (IGFBP-1) in endometrial stromal cells. The primary culture of human endometrial stromal cells was transfected with the hPR-A or hPR-B expression vector and the IGFBP-1 promoter construct p275CAT, which contains two functional progesterone response elements (PRE1 and PRE2) in decidualized stromal cells. Medroxyprogesterone acetate (MPA) increased the promoter activities ranging from 1.2- to 27-fold in cells cotransfected with hPR-A or hPR-B in eight endometrial specimens. The promoter activity increased by the hPR-A was significantly higher than hPR-B (15 +/- 8 vs. 4 +/- 2, mean +/- SD; n = 8, P < 0.005). Site-specific mutation showed that the induced activity by hPR-A was mediated through the PRE1 and PRE2 sites. Addition of hPR-B reduced the effect of hPR-A. The high transactivation capacity of hPR-A was also activated by other ligands, progesterone, Org 2058, and norethindrone. These observations indicate that hPR-A is a stronger transactivator than hPR-B for the IGFBP-1 promoter in endometrial stromal cells. Previous studies have shown the progestin-dependent production of IGFBP-1 correlates with its mRNA levels and transcription rate. Thus, we have determined the effect of hPR-A and hPR-B on the production of IGFBP-1 in stromal cells treated with MPA. The production rate in cells uniformly infected with AdPRA (recombinant Ad5-directed PR expression system) was significantly higher (P < 0.001) than the rate in uninfected cells and in cells infected with AdPRB or AdCMV (the Ad5 viral expression vector). This result, in concert with the promoter analysis, provides evidence that hPR-A is a strong inducer for the chromosomal IGFBP-1 gene in endometrial stromal cells. PMID- 11117527 TI - Activation of peroxisome proliferator-activated receptors (PPARs) by their ligands and protein kinase A activators. AB - The nuclear peroxisome proliferator-activated receptors (PPARs) alpha, beta, and gamma activate the transcription of multiple genes involved in lipid metabolism. Several natural and synthetic ligands have been identified for each PPAR isotype but little is known about the phosphorylation state of these receptors. We show here that activators of protein kinase A (PKA) can enhance mouse PPAR activity in the absence and the presence of exogenous ligands in transient transfection experiments. Activation function 1 (AF-1) of PPARs was dispensable for transcriptional enhancement, whereas activation function 2 (AF-2) was required for this effect. We also show that several domains of PPAR can be phosphorylated by PKA in vitro. Moreover, gel retardation experiments suggest that PKA stabilizes binding of the liganded PPAR to DNA. PKA inhibitors decreased not only the kinase-dependent induction of PPARs but also their ligand-dependent induction, suggesting an interaction between both pathways that leads to maximal transcriptional induction by PPARs. Moreover, comparing PPAR alpha knockout (KO) with PPAR alpha WT mice, we show that the expression of the acyl CoA oxidase (ACO) gene can be regulated by PKA-activated PPAR alpha in liver. These data demonstrate that the PKA pathway is an important modulator of PPAR activity, and we propose a model associating this pathway in the control of fatty acid beta oxidation under conditions of fasting, stress, and exercise. PMID- 11117528 TI - The nuclear receptor corepressor (N-CoR) contains three isoleucine motifs (I/LXXII) that serve as receptor interaction domains (IDs). AB - Unliganded thyroid hormone receptors (TRs) repress transcription through recruitment of corepressors, including nuclear receptor corepressor (N-CoR). We find that N-CoR contains three interaction domains (IDs) that bind to TR, rather than the previously reported two. The hitherto unrecognized ID (ID3) serves as a fully functional TR binding site, both in vivo and in vitro, and may be the most important for TR binding. Each ID motif contains a conserved hydrophobic core (I/LXXII) that resembles the hydrophobic core of nuclear receptor boxes (LXXLL), which mediates p160 coactivator binding to liganded nuclear receptors. Although the integrity of the I/LXXII motif is required for ID function, substitution of ID isoleucines with leucines did not allow ID peptides to bind to liganded TR, and substitution of NR box leucines with isoleucines did not allow NR box peptides to bind unliganded TR. This indicates that the binding preferences of N CoR for unliganded TR and p160s for liganded TR are not dictated solely by the identity of conserved hydrophobic residues within their TR binding motifs. Examination of sequence conservation between IDs, and mutational analysis of individual IDs, suggests that they are comprised of the central hydrophobic core and distinct adjacent sequences that may make unique contacts with the TR surface. Accordingly, a hybrid peptide that contains distinct adjacent sequences from ID3 and ID1 shows enhanced binding to TR. PMID- 11117529 TI - ARIP3 (androgen receptor-interacting protein 3) and other PIAS (protein inhibitor of activated STAT) proteins differ in their ability to modulate steroid receptor dependent transcriptional activation. AB - Steroid receptors mediate their actions by using various coregulatory proteins. We have recently characterized ARIP3/PIASx alpha as an androgen receptor (AR) interacting protein (ARIP) that belongs to the PIAS [protein inhibitor of activated STAT (signal transducer and activator of transcription)] protein family implicated in the inhibition of cytokine signaling. We have analyzed herein the roles that four different PIAS proteins (ARIP3/PIASx alpha, Miz1/PIASx beta, GBP/PIAS1, and PIAS3) play in the regulation of steroid receptor- or STAT mediated transcriptional activation. All PIAS proteins are able to coactivate steroid receptor-dependent transcription but to a differential degree, depending on the receptor, the promoter, and the cell type. Miz1 and PIAS1 are more potent than ARIP3 in activating AR function on minimal promoters. With the natural probasin promoter, PIAS proteins influence AR function more divergently, in that ARIP3 represses, but Miz1 and PIAS1 activate it. Miz1 and PIAS1 possess inherent transcription activating function, whereas ARIP3 and PIAS3 are devoid of this feature. ARIP3 enhances glucocorticoid receptor-dependent transcription more efficiently than Miz1 or PIAS1, and all PIAS proteins also activate estrogen receptor- and progesterone receptor-dependent transcription but to a dissimilar degree. The same amounts of PIAS proteins that modulate steroid receptor dependent transcription influence only marginally transactivation mediated by various STAT proteins. It remains to be established whether the PIAS proteins play a more significant physiological role in steroid receptor than in cytokine signaling. PMID- 11117530 TI - Temporal formation of distinct thyroid hormone receptor coactivator complexes in HeLa cells. AB - Thyroid hormone receptors (TRs) regulate transcription by recruiting distinct coregulatory complexes to target gene promoters. Coactivators implicated in ligand-dependent activation by TR include p300, the CREB-binding protein (CBP), members of the p160/SRC family, and the multisubunit TR-associated protein (TRAP) complex. Using a stable TR-expressing HeLa cell line, we show that interaction of TR with members of the p160/SRC family, CBP, and the p300/CBP-associated factor (PCAF) occurs rapidly (approximately 10 min) following addition of thyroid hormone (T3). In close agreement with these observations, we find that TR is associated with potent histone acetyltransferase activity rapidly following T3 treatment. By contrast, we observe that formation of TR-TRAP complexes occurs significantly later (approximately 3 h) post T3 treatment. An examination of the kinetics of T3-induced gene expression in HeLa cells reveals bimodal or delayed activation on specific T3-responsive promoters. Taken together, our data are consistent with the hypothesis that T3-dependent activation at specific target promoters may involve the regulated action of multiple TR-coactivator complexes. PMID- 11117531 TI - Development of peptide antagonists that target estrogen receptor beta-coactivator interactions. AB - The biological actions of estrogen are manifest through two genetically distinct estrogen receptors (ER alpha and ER beta) that display nonidentical expression patterns in target tissues. The phenotypic alterations in response to estrogens in mice disrupted for either or both of these receptors are not identical, suggesting that each subtype plays a unique role in ER-action. However, the lack of subtype-specific agonists and antagonists has made it difficult to define the processes that are regulated by ER alpha and/or ER beta. Previously, we have reported the identification and characterization of a series of LXXLL-containing peptide antagonists that block estrogen signaling by preventing the association of ER alpha with required coactivators. As expected, given the similarity of the coactivator binding pockets among nuclear receptors, most of the peptide antagonists identified inhibited the activity of multiple receptors. However, by altering sequences flanking the core LXXLL motif, some receptor selectivity was afforded. Building on this observation, we have screened combinatorial phage libraries, expressing peptides in the format X7LXXLLX7, for peptides that interact in a specific manner with ER beta. Using this approach, a series of highly specific, potent peptide antagonists have been identified that efficiently inhibit ER beta-mediated estrogen signaling when introduced into target cells. Interestingly, in cells where both ER subtypes were expressed, these ER beta antagonists were capable of attenuating ER action, suggesting that ER alpha and ER beta do indeed form functional heterodimeric complexes. We believe that suitably formulated versions of these peptides can be used to study ER beta action in vitro and in vivo. In addition, the unanticipated specificity of the peptides identified should serve as an impetus to investigate the use of this approach to develop peptide antagonists of other nuclear receptors and unrelated transcription factors. PMID- 11117532 TI - Temporally distinct and ligand-specific recruitment of nuclear receptor interacting peptides and cofactors to subnuclear domains containing the estrogen receptor. AB - Ligand binding to estrogen receptor (ER) is presumed to regulate the type and timing of ER interactions with different cofactors. Using fluorescence microscopy in living cells, we characterized the recruitment of five different green fluorescent protein (GFP)-labeled ER-interacting peptides to the distinct subnuclear compartment occupied by blue fluorescent protein (BFP)-labeled ER alpha. Different ligands promoted the recruitment of different peptides. One peptide was recruited in response to estradiol (E2), tamoxifen, raloxifene, or ICI 182,780 incubation whereas other peptides were recruited specifically by E2 or tamoxifen. Peptides containing different sequences surrounding the ER interacting motif LXXLL were recruited with different time courses after E2 addition. Complex temporal kinetics also were observed for recruitment of the full-length, ER cofactor glucocorticoid receptor-interacting protein 1 (GRIP1); rapid, E2-dependent recruitment of GRIP1 was blocked by mutation of the GRIP1 LXXLL motifs to LXXAA whereas slower E2 recruitment persisted for the GRIP1 LXXAA mutant. This suggested the presence of multiple, temporally distinct GRIP 1 recruitment mechanisms. E2 recruitment of GRIP1 and LXXLL peptides was blocked by coincubation with excess ICI 182,780. In contrast, preformed E2/ER/GRIP1 and E2/ER/LXXLL complexes were resistant to subsequent ICI 182,780 addition whereas ICI 182,780 dispersed preformed complexes containing the GRIP1 LXXAA mutant. This suggested that E2-induced LXXLL binding altered subsequent ligand/ER interactions. Thus, alternative, ligand-selective recruitment and dissociation mechanisms with distinct temporal sequences are available for ER alpha action in vivo. PMID- 11117533 TI - Receptor/beta-arrestin complex formation and the differential trafficking and resensitization of beta2-adrenergic and angiotensin II type 1A receptors. AB - Beta-arrestins target G protein-coupled receptors (GPCRs) for endocytosis via clathrin-coated vesicles. Beta-arrestins also become detectable on endocytic vesicles in response to angiotensin II type 1A receptor (AT1AR), but not beta2 adrenergic receptor (beta2AR), activation. The carboxyl-terminal tails of these receptors contribute directly to this phenotype, since a beta2AR bearing the AT1AR tail acquired the capacity to stimulate beta-arrestin redistribution to endosomes, whereas this property was lost for an AT1AR bearing the beta2AR tail. Using beta2AR/AT1AR chimeras, we tested whether the beta2AR and AT1AR carboxyl terminal tails, in part via their association with beta-arrestins, might regulate differences in the intracellular trafficking and resensitization patterns of these receptors. In the present study, we find that beta-arrestin formed a stable complex with the AT1AR tail in endocytic vesicles and that the internalization of this complex was dynamin dependent. Internalization of the beta2AR chimera bearing the AT1AR tail was observed in the absence of agonist and was inhibited by a dominant-negative beta-arrestin1 mutant. Agonist-independent AT1AR internalization was also observed after beta-arrestin2 overexpression. After internalization, the beta2AR, but not the AT1AR, was dephosphorylated and recycled back to the cell surface. However, the AT1AR tail prevented beta2AR dephosphorylation and recycling. In contrast, although the beta2AR-tail promoted AT1AR recycling, the chimeric receptor remained both phosphorylated and desensitized, suggesting that receptor dephosphorylation is not a property common to all receptors. In summary, we show that the carboxyl-terminal tails of GPCRs not only contribute to regulating the patterns of receptor desensitization, but also modulate receptor intracellular trafficking and resensitization patterns. PMID- 11117534 TI - Janus kinase 2 (JAK2) regulates prolactin-mediated chloride transport in mouse mammary epithelial cells through tyrosine phosphorylation of Na+-K+-2Cl- cotransporter. AB - Epithelial chloride (Cl-) transport is achieved by the coordinated action of symporters such as the Na+-K+-2Cl- cotransporter (NKCC1) and chloride channels such as the cystic fibrosis transmembrane conductance regulator (CFTR). As a secretory tissue, mammary epithelial cells are obvious candidates for such mechanisms, but Cl- transport and its hormonal regulation have been poorly delineated in mammary epithelial cells. We determined whether the mammary epithelial cell line, HC11, transports chloride and whether this was regulated by PRL, a hormone known to stimulate ion transport. HC11 cells express both CFTR and NKCC1. Exposure to PRL or PGE1 increased Cl- transport in HC11 cells. This was inhibited by the NKCC1 blocker, furosemide, and by the Cl- channel inhibitor, diphenylamine 2-carboxylate. Dose and time course of PRL action indicate that PRL had maximal effect on Cl- transport at 1 microg/ml and at 10 min of stimulation. Examination of the signaling pathways suggests that the PRL effect on Cl- transport does not involve an increase in [Ca2+]i or MAP kinase activity. RT-PCR analyses indicate that HC11 cells express mRNA for Janus kinase 1 (JAK1), JAK2, and signal transducer and activator of transcription 5 (STAT5) but not for JAK3. PRL treatment of HC11 cells increased phosphorylation of STAT5. The JAK2 inhibitor AG490 blocked phosphorylation of STAT5 and PRL-induced, but not PGE1 induced, Cl- transport. NKCC1, but not CFTR, is tyrosine phosphorylated in HC11 cells. PRL enhanced tyrosine phosphorylation of NKCC1, and this effect was attenuated by the JAK2 inhibitor AG490. These results are the first demonstrations of a role for tyrosine phosphorylation of NKCC1 and of the PRL JAK2 cascade in the regulation of Cl- transport. PMID- 11117535 TI - Truncated activin type I receptor Alk4 isoforms are dominant negative receptors inhibiting activin signaling. AB - Activin, a member of the transforming growth factor beta (TGFbeta) superfamily of cytokines, inhibits cell proliferation in a variety of cell types. The functions of activin are mediated by type I and type II serine/threonine kinase receptors. The main type I receptor mediating activin signaling in human cells is ActRIB, also called Alk4. We have previously reported that several truncated Alk4 receptor isoforms are exclusively expressed in human pituitary tumors, and that the majority of such tumors did not exhibit activin-induced growth arrest in culture. We therefore studied the function of these truncated receptor isoforms. Transient expression of these truncated receptors inhibited activin-activated transcription from an activin-responsive reporter construct, 3TPLux. When each of these truncated Alk4 receptors was stably transfected into K562 cells, activin induced expression of an endogenous gene, junB, was blocked, indicating that inhibition of gene expression also occurred at the chromosomal level. Furthermore, activin administration failed to cause growth inhibition and an increase of the G1 population in these cells. Coimmunoprecipitation experiments showed that the truncated Alk4 receptors formed complexes with type II activin receptors, but were not phosphorylated. These data indicate that the truncated activin type I receptors, predominantly expressed in human pituitary adenomas, function as dominant negative receptors to interfere with wild-type receptor function and block the antiproliferative effect of activin. This may contribute to uncontrolled pituitary cell growth and the development of human pituitary tumors. PMID- 11117536 TI - Urocortin, but not corticotropin-releasing hormone (CRH), activates the mitogen activated protein kinase signal transduction pathway in human pregnant myometrium: an effect mediated via R1alpha and R2beta CRH receptor subtypes and stimulation of Gq-proteins. AB - CRH and CRH-related peptides such as urocortin mediate their actions in the human myometrium via activation of two distinct classes of CRH receptors, R1 and R2. These heptahelical receptors are able to stimulate a number of different intracellular signals; one key mediator of G protein-activated intracellular signaling is the cascade of p42/p44, mitogen-activated protein kinase (MAPK). We therefore hypothesized that activation of MAPK might mediate CRH and or/urocortin actions in the myometrium. In cultured human pregnant myometrial cells, urocortin but not CRH was able to induce MAPK phosphorylation and activation, suggesting that in the human myometrium these two peptides have distinct actions and biological roles. To identify the particular receptor subtypes mediating this phenomenon, all known CRH receptors present in the human myometrial cells were stably expressed individually in HEK293 and CHO cells, and their ability to activate MAPK was tested. The R1alpha and R2beta, but not the R1beta, R1c, or R1d, receptor subtypes were able to mediate urocortin-induced MAPK activation. The signaling components were further investigated; activation of Gs, Go, or Gi proteins did not appear to be involved, but activation of Gq with subsequent production of inositol triphosphates (IP3) and protein kinase C (PKC) activation correlated with MAPK phosphorylation. Studies on Gq protein activation using [alpha-32P]-GTP-gamma-azidoanilide and IP3 production in cells expressing the R1alpha or R2beta CRH receptors demonstrated that urocortin was 10 times more potent than CRH. Moreover, urocortin (UCN) generated peak responses that were 50 70% greater than CRH in activating the Gq protein and stimulating IP3 production. In conclusion, UCN acting thought multiple receptor subtypes can stimulate myometrial MAPK via induction of the Gq/phospholipase C/IP3/PKC pathway, whereas CRH-induced activation of this pathway appears to be insufficient to achieve MAPK activation. PMID- 11117537 TI - From telomeres to T-antigens: many roads...multiple pathways...novel associations in the search for the origins of human gliomas. PMID- 11117538 TI - Activity-dependent hyperpolarization and conduction block in chronic inflammatory demyelinating polyneuropathy. AB - Voluntary activity produces activity-dependent hyperpolarization of the active motor axons. The present study investigated whether this hyperpolarization produces conduction block in chronic inflammatory demyelinating polyneuropathy (CIDP). Studies were performed in 10 healthy control subjects, 7 patients with CIDP, and 3 patients with multifocal motor neuropathy. The compound muscle action potential (CMAP) of the abductor pollicis brevis was recorded in response to supramaximal stimuli to the median nerve at the wrist, alternating with measurements of axonal excitability. After a maximal voluntary contraction for 60 seconds, the amplitude of the maximal CMAP was significantly reduced in symptomatic CIDP patients by 40%, but there were only slight changes in the CMAPs of healthy controls, asymptomatic CIDP patients, and multifocal motor neuropathy patients. In symptomatic CIDP patients, the activity-dependent conduction block paralleled the activity-dependent hyperpolarization and was presumably precipitated by it. In these patients, the safety margin for impulse conduction was estimated to be about 12%. Activity-dependent conduction block may be clinically important in chronic demyelinating diseases and can be demonstrated electrophysiologically if testing occurs across pathological sites. PMID- 11117539 TI - Factors predicting prognosis of epilepsy after presentation with seizures. AB - The objective of this study was to identify the factors, at the time of diagnosis, that determine the prognosis for remission of epilepsy. A prospective community-based cohort study of 792 patients recruited at the time of their first diagnosis of epileptic seizures was undertaken; in those classified 6 months after presentation, the median follow-up period was 7.2 years (quartiles at 6.2 and 8.2 years) after presentation. We analyzed data from 6 months after the first identified seizure, which prompted the diagnosis of epilepsy, to allow us to factor in those aspects contingent on a diagnostic assessment Baseline clinical and demographic data were analyzed using the Cox proportional hazards regression model with remission of epilepsy for 1, 2, 3, and 5 years as outcome measures. The dominant clinical feature predicting remission was the number of seizures in the 6-month diagnostic assessment period. Thus, the chance of entering 1 year of remission by 6 years for a patient who had 2 seizures during this initial 6 months was 95%; for 5 years of remission, it was 47% as opposed to 75% for 1 year of remission and 24% for 5 years of remission if there had been 10 or more seizures during this period. The number of seizures in the early phase of epilepsy (here, taken as the first 6 months after presentation) is the single most important predictive factor for both early and long-term remission of seizures. PMID- 11117540 TI - Phantom limb pain in the human brain: unraveling neural circuitries of phantom limb sensations using positron emission tomography. AB - Pain and other phantom limb (PL) sensations have been proposed to be generated in the brain and to be reflected in activation of specific neural circuits. To test this hypothesis, hypnosis was used as a cognitive tool to alternate between the sensation of PL movement and pain in 8 amputees. Brain activity was measured using positron emission tomography. PL movement and pain were represented by a propagation of neuronal activity within the corresponding sensorimotor and pain processing networks. The sensation of movement was significantly (corrected for multiple comparisons) related to activity in the supplementary motor area and the primary sensorimotor cortex. The sensation of a painful PL posture activated the same brain areas but was weaker and less extended in the supplementary motor area. In contrast to the sensation of movement, pain was significantly related to activity in the thalamus, anterior cingulate, and lateral prefrontal cortex. Subjectively rated PL pain sensation correlated positively to activations in the anterior and posterior cingulate. These findings provide evidence that PL sensations are produced by the same central nervous processes that underlie the experience of the body when it is intact and that the corporeal awareness of PL pain is encoded in a thalamocortical network. PMID- 11117541 TI - Frontotemporal dementia with novel tau pathology and a Glu342Val tau mutation. AB - It is unclear how tau gene mutations cause frontotemporal dementia (FTD) with parkinsonism linked to chromosome 17 (FTDP-17), but those in exon 10 (E10) or the following intron may be pathogenic by altering E10 splicing, perturbing the normal 1:1 ratio of four versus three microtubule-binding repeat tau (4R:3R tau ratio) and forming tau inclusions. We report on a 55-year old woman with frontotemporal dementia and a family history of FTDP-17 in whom we found a novel E12 (Glu342Val) tau gene mutation, prominent frontotemporal neuron loss, intracytoplasmic tau aggregates, paired helical tau filaments, increased 4R tau messenger RNA, increased 4R tau without E2 or E3 inserts, decreased 4R tau with these inserts, and a 4R:3R tau ratio greater than 1 in gray and white matter. Thus, this novel Glu342Val mutation may cause FTDP-17 by unprecedented mechanisms that alter splicing of E2, E3, and E10 to preferentially increase 4R tau without amino terminal inserts and promote aggregation of tau filaments into cytopathic inclusions. PMID- 11117542 TI - Pick's disease is associated with mutations in the tau gene. AB - Recently, mutations within the tau gene have been associated with some familial forms of frontotemporal dementia. To investigate whether tau gene mutations are also associated with Pick's disease, we analyzed the tau gene in 30 cases of pathologically confirmed Pick's disease. Two coding mutations were identified in separate cases of Pick's disease. A glycine-to-arginine mutation at codon 389 was detected in 1 case and a lysine-to-threonine mutation at codon 257 was identified in another. Analysis of dephosphorylated tau from the brain of the patient with the codon 389 mutation revealed a prominent band representing tau, with four microtubule-binding domains and no amino terminal inserts. This is in contrast to Pick's disease without any tau gene mutations, which consist of tau with mainly three microtubule-binding domains and only a trace of tau, with four microtubule binding domains. Functional analysis of tau with these two mutations demonstrated a reduced ability of tau to promote microtubule assembly. Surprisingly, these mutations increased tau's susceptibility to calpain I digestion, suggesting that this feature may be related to the formation of a Pick type of histology. Moreover, these data suggest that Pick's disease is not a separate entity but part of the frontotemporal dementia disease spectrum. PMID- 11117543 TI - Delusions associated with elevated muscarinic binding in dementia with Lewy bodies. AB - The relation between disturbances of cholinergic neurotransmission and delusions (DELs) has not been investigated in degenerative dementias such as dementia with Lewy bodies (DLB). A cohort of dementia patients were assessed with standardized clinical evaluations (including the Columbia University Scale for Psychopathology in Alzheimer's Disease), which were repeated annually until death. DLB was confirmed neuropathologically in 21 patients. Neurochemical evaluation included M1 receptor autoradiography (pirenzepine binding), biochemical measurement of choline acetyltransferase (ChAT), and acetylcholinesterase (AChE) histochemistry in brain regions hypothesized to be involved in the genesis of psychosis. Compared with 11 age-matched controls, CHAT and pirenzepine levels were most extensively reduced in the temporal and parietal neocortex of DLB patients. In Brodmann area 36, DELs were significantly associated with elevated pirenzepine binding (131.0 vs 93.5, t = 2.7), whereas visual hallucinations were associated with significant reductions in ChAT (1.7 vs 2.5, t = 2.5). There were no significant associations with other areas or with cholinesterase. Although DELs and visual hallucinations were both linked with disturbances in cholinergic neurotransmission, the nature of the associations was different. Upregulation of the postsynaptic muscarinic receptor may be central in the genesis of DELs, with important treatment implications. PMID- 11117544 TI - A novel form of distal hereditary motor neuronopathy maps to chromosome 9p21.1 p12. AB - Distal hereditary motor neuronopathies (dHMNs) form a heterogeneous group of rare disorders characterized by distal weakness and wasting in the limbs with no significant sensory involvement. Harding has classified dHMNs into seven categories based on clinical and genetic criteria. We report a novel form of autosomal recessive dHMN in 7 consanguineous families located in the Jerash region of Jordan. Onset of the disease is between 6 and 10 years of age and is characterized by weakness and atrophy of the lower limbs associated with pyramidal features. Within 2 years, symptoms progress to the upper limbs. Neurophysiological studies typically show normal conduction velocities, reduced compound motor action potential amplitudes, normal sensory nerve action potentials, and chronic neurogenic changes on needle electromyography. No significant abnormalities are seen on sural nerve biopsy. We call this novel form of dHMN Jerash hereditary motor neuronopathy. We studied the families at the molecular genetic level and mapped the Jerash hereditary motor neuronopathy gene to an approximately 0.54-cM region on chromosome 9p21.1-p12, flanked by microsatellite polymorphic marker loci D9S1845 and D9S1791. A maximum LOD score of 19.80 at theta = 0.001 was obtained between the disease and locus D9S1878. PMID- 11117545 TI - Neuropsychological effects of interferon beta-1a in relapsing multiple sclerosis. Multiple Sclerosis Collaborative Research Group. AB - Cognitive dysfunction is common in multiple sclerosis (MS), yet few studies have examined effects of treatment on neuropsychological (NP) performance. To evaluate the effects of interferon beta-1a (IFNbeta-1a, 30 microg administered intramuscularly once weekly [Avonex]) on cognitive function, a Comprehensive NP Battery was administered at baseline and week 104 to relapsing MS patients in the phase III study, 166 of whom completed both assessments. A Brief NP Battery was also administered at 6-month intervals. The primary NP outcome measure was 2-year change on the Comprehensive NP Battery, grouped into domains of information processing and learning/memory (set A), visuospatial abilities and problem solving (set B), and verbal abilities and attention span (set C). NP effects were most pronounced in cognitive domains vulnerable to MS: IFNbeta-1a had a significant beneficial effect on the set A composite, with a favorable trend evident on set B. Secondary outcome analyses revealed significant between-group differences in slopes for Brief NP Battery performance and time to sustained deterioration in a Paced Auditory Serial Addition Test processing rate, favoring the IFNbeta-1a group. These results support and extend previous observations of significant beneficial effects of IFNbeta-1a for relapsing MS. PMID- 11117546 TI - Neurological disability correlates with spinal cord axonal loss and reduced N acetyl aspartate in chronic multiple sclerosis patients. AB - Axonal degeneration has been proposed as a cause of irreversible neurological disability in multiple sclerosis (MS) patients. The purpose of this study was to quantify axonal loss in spinal cord lesions from 5 paralyzed (Expanded Disability Status Scale score > or =7.5) MS patients and to determine if axonal number or volume correlated with levels of the neuronal marker N-acetyl aspartate (NAA). Axonal loss in MS lesions ranged from 45 to 84% and averaged 68%. NAA levels were significantly reduced (>50%) in cross sections of spinal cords containing MS lesions. Reduced NAA correlated with reduced axonal numbers within lesion areas. In addition, NAA levels per axonal volume were significantly reduced in demyelinated axons (42%) and in myelinated axons in normal-appearing white matter (30%). The data support axonal loss as a major cause of irreversible neurological disability in paralyzed MS patients and indicate that reduced NAA as measured by magnetic resonance spectroscopy can reflect axonal loss and reduced NAA levels in demyelinated and myelinated axons. PMID- 11117547 TI - Intracellular accumulation and reduced sarcolemmal expression of dysferlin in limb--girdle muscular dystrophies. AB - Dysferlin has recently been identified as a novel gene involved in limb-girdle muscular dystrophy type 2B (LGMD2B) and its allelic disease, Miyoshi myopathy. The predicted structure of dysferlin suggests that it is a transmembrane protein possibly involved in membrane fusion. Thus, unlike previously identified structural proteins in muscular dystrophy, dysferlin is likely involved in a novel pathogenic mechanism for this disease. In this study, we have analyzed the expression of dysferlin in skeletal muscle of patients with disruptions in the dystrophin-glycoprotein complex and patients with a clinical diagnosis of LGMD2B or Miyoshi myopathy. We show expression of dysferlin at the sarcolemma in normal muscle and reduced sarcolemmal expression along with accumulation of intracellular staining in dystrophic muscle. Electron microscopy in Miyoshi myopathy biopsies suggests that the cytoplasmic staining could be a result of the abundance of intracellular vesicles. Our results indicate that dysferlin expression is perturbed in LGMD and that both mutations in the dysferlin gene and disruption of the dystrophin-glycoprotein complex can lead to the accumulation of dysferlin within the cytoplasm. PMID- 11117548 TI - The selective muscarinic M1 agonist AF102B decreases levels of total Abeta in cerebrospinal fluid of patients with Alzheimer's disease. AB - beta-Amyloid (Abeta) deposits in diffuse and compact senile plaques in the brain are one of the defining histopathological features of Alzheimer's disease (AD). Preventing Abeta deposition is a goal of drug therapy for AD, because excessive amounts of Abeta may be toxic to neurons. In preclinical studies, activation of the muscarinic M1 receptor subtype inhibited Abeta secretion from cultured cells. To determine whether a similar sequence occurs in human beings, we administered the selective M1 agonist AF102B to 19 AD patients and measured total Abeta (Abeta(total)) levels in cerebrospinal fluid (CSF) before and during treatment. Abeta(total) levels in CSF decreased in 14 patients by 22%, increased in 3 patients, and were unchanged in 2 patients; the overall decrease in the group as a whole was statistically significant. To test the specificity of the M1 effect, we also measured the relative changes in Abeta(total) levels in CSF during treatments in separate sets of AD patients with the acetylcholinesterase inhibitor physostigmine or the anti-inflammatory drug hydroxychloroquine. CSF Abeta(total) levels did not change significantly in the 9 AD patients in the physostigmine protocol or in the 10 AD patients in the hydroxychloroquine study. These data provide evidence that the activation of M1 receptors reduces Abeta levels in the CSF of AD patients. If this effect also occurs in brain, M1 agonists may have long-term therapeutic benefits by lowering amyloid in AD. PMID- 11117549 TI - Multifocal motor neuropathy: diagnostic criteria that predict the response to immunoglobulin treatment. AB - As multifocal motor neuropathy (MMN) is a potentially treatable disorder, its differentiation from lower motor neuron disease is important. Evidence of conduction block (CB) is considered one of the relevant criteria for the diagnosis of MMN. Strict criteria for CB may lead to underdiagnosis of MMN, however. Using a standardized examination, we studied the clinical, laboratory, and electrophysiological characteristics of 37 patients presenting with a lower motor neuron disorder and electrophysiological features compatible with segmental demyelination. We propose a set of clinical, laboratory, and electrophysiological criteria for the diagnosis of MMN, which has been verified by follow-up and response to treatment with intravenous immunoglobulins. Based on the clinical, laboratory, and electrodiagnostic features, 21 patients were diagnosed with definite MMN (17 responders), 7 were diagnosed with probable MMN (5 responders), and 9 were diagnosed with possible MMN (1 responder). Age at onset, the number of affected limb regions, and the number of patients with a creatine kinase level greater than 180 U/L were significantly lower in responders than in nonresponders. Elevated anti-GM1 antibodies and definite CB were found significantly more often in responders. The proposed diagnostic criteria may be useful in clinical practice and therapeutic trials. PMID- 11117550 TI - Conjugal multiple sclerosis: population-based prevalence and recurrence risks in offspring. Canadian Collaborative Study Group. AB - From a population-based sample of 15,504 patients attending Canadian multiple sclerosis (MS) clinics, we have determined the frequency of conjugal MS and have estimated the recurrence risk in offspring of such matings. Twenty-three MS cases were found among 13,550 spouses of study probands for a crude conjugal rate of 0.17% (95% CI of 0.10%-0.24%). Despite ascertainment bias that expectedly inflates this number, this is a frequency intermediate between the point prevalence (0.1%) and lifetime risk (0.2%) for the general population and close to an order of magnitude less than reported for half siblings reared apart (1.06%) from the same population. Six of the 49 offspring of conjugal pairs also had MS, and age conversion gives a rate similar to the concordance rate for Canadian monozygotic twins. However, this correction may not be appropriate in this special case. Despite an ascertainment bias in favor of recognizing affected spouses and a large population sample, the common environment in adulthood shared by spousal pairs could not be shown to increase the risk of conjugal MS. Although the high recurrence rate in offspring is similarly subject to an upward bias, the low risk for MS spouses and the high risk for offspring support other data indicating that familial risk is genetically determined. Furthermore, these results imply that susceptibility alleles are shared by unrelated individuals with the disease. PMID- 11117551 TI - Reactivation of human neurotropic JC virus expressing oncogenic protein in a recurrent glioblastoma multiforme. AB - Examination of the primary tumor of glioblastoma multiforme and its recurrence for their association with JC virus revealed that, while the viral genome is present in both initial and recurrent tumors, expression of the viral oncoprotein T-antigen occurs only in the recurrent tumor cells. Accordingly, the level of inducible cellular transcription factors, including the p65 subunit of NF-kappaB and YB-1, which have the ability to stimulate JCV gene expression, was found to be higher in the recurrent tumor cells. These observations suggest that induction of the regulatory factors after resection of the primary tumor may have reactivated JC virus gene expression and led to redevelopment of the tumor in brain. PMID- 11117552 TI - Herpes simplex virus type 1 (HSV-1)--induced retinitis following herpes simplex encephalitis: indications for brain-to-eye transmission of HSV-1. AB - Herpes simplex encephalitis is a severe neurological disease with high mortality and morbidity rates. Reactivated herpes simplex virus type 1 (HSV-1) can cause relapses and might even spread to the retina, where it can induce a potentially blinding eye disease, known as acute retinal necrosis. In the present study, the HSV-1 strains in the brain and eye of 2 patients with acute retinal necrosis following an episode of herpes simplex encephalitis were genotyped. The HSV-1 strains in both the brain and eye were identical in each patient, but they differed interindividually. The data suggest brain-to-eye transmission of HSV-1 in these patients. PMID- 11117553 TI - A novel tau mutation (N296N) in familial dementia with swollen achromatic neurons and corticobasal inclusion bodies. AB - Familial dementia with swollen achromatic neurons and corticobasal inclusion bodies is a neurodegenerative disease that resembles corticobasal degeneration. It is characterized by the presence of abundant neuronal and glial tau protein deposits. Here we describe a novel silent mutation in exon 10 of tau (N296N) in this familial dementia. By exon trapping, the mutation produced an increase in the splicing in of exon 10, indicating that it probably causes disease through an overproduction of four-repeat tau. PMID- 11117554 TI - Therapeutic benefit of polyamine-modified catalase as a scavenger of hydrogen peroxide and nitric oxide in familial amyotrophic lateral sclerosis transgenics. AB - Continuous subcutaneous administration of polyamine-modified catalase that has increased permeability at the blood-brain barrier showed both a highly significant delay in onset and an increase in survival in a transgenic mouse model of familial amyotrophic lateral sclerosis having a point mutation in the gene encoding copper/zinc superoxide dismutase. These results suggest that hydrogen peroxide-mediated oxidative stress with subsequent free radical damage involving nitric oxide and possibly hydroxyl radicals in motor neurons may be the culprit in familial amyotrophic lateral sclerosis. PMID- 11117555 TI - Demyelinating antibodies in multiple sclerosis. PMID- 11117556 TI - Interleukin-1beta gene polymorphism and susceptibility to temporal lobe epilepsy with hippocampal sclerosis. PMID- 11117557 TI - No difference in plasma or urinary F2-isoprostanes among patients with Huntington's disease or Alzheimer's disease and controls. PMID- 11117558 TI - Changes of in vivo gastrointestinal motor pattern in pacemaker-deficient (WsRC Ws/Ws) rats. AB - In vitro studies on pacemaker-deficient W-mutants have revealed a disappearance of rhythmic contraction in their gastrointestinal tracts. Their contractile force has not been diminished, however. In contrast, W-mutants often present dysmoility like symptoms with distension of the gastrointestinal tract in vivo. Gastrointestinal motility of W-mutant rats was examined in vivo by an extraluminal strain-gauge force transducer method. We examined a normal gastrointestinal motor pattern in the rats with two distinct motor phases, digestive and interdigestive. Moreover, we detected a failure to form an interdigestive contractile complex in pacemaker-deficient rats. The interdigestive motor activity of the gastrointestinal tract is important for cleaning gastrointestinal tract in preparation for the next meal. The impairment of the interdigestive contractile complex may be related to the dysmoility-like symptoms of W-mutant rats in vivo. PMID- 11117559 TI - Long-term postoperative functional evaluation of pylorus preservation in Imanaga pancreatoduodenectomy. AB - Our purpose was to determine whether pylorus-preservation in the Imanaga (PpPDI) method minimizes postoperative impairment of gastrointestinal function. Nine patients who had undergone PpPDI (postoperative years: 5.7 +/- 2.6) and nine patients who had undergone conventional Imanaga pancreatoduodenectomy (PDI) (postoperative years: 6.8 +/- 2.0) were evaluated for symptoms, nutritional parameters, and physiologic function of the biliary tract and residual stomach using gastric emptying and hepatobiliary scintigraphy. The body weight recovered to 99.3% +/- 3.8% of pre-illness body weight in PpPDI, showing a significantly better recovery than in patients after the PDI procedure (91.0% +/- 6.4%, P < 0.05). The mean gastric emptying half-time (GET1/2) in the upright position after PDI was significantly shorter (42.3 min) than after PpPDI (80.8 min, P < 0.05). Mixture of food with bile was conserved better in the PpPDI group than in the PDI group. In the long term, the pylorus-preserving Imanaga-type procedure minimizes disruption of gastric function and asynchrony between ingested food and bile. PMID- 11117560 TI - Hepatic expression of hepatocyte growth factor gene mRNA in acute liver failure. AB - Hepatocyte growth factor plays a key role in liver regeneration but the role of liver in its synthesis in acute liver failure is unclear. We therefore measured hepatic expression of hepatocyte growth factor mRNA in this condition in comparison to H3 histone mRNA, a marker of cellular proliferation. Hepatocyte growth factor mRNA levels were quantified by specific RNase protection assay in nine patients with acute liver failure and found to be similar to those in six normal controls. Hepatocyte proliferation, as assessed by H3 histone mRNA expression, was not detected in normal liver but was present in six of nine patients with acute liver failure (P < 0.05) and was not correlated with expression of hepatocyte growth factor mRNA (rs = -0.28). Liver is unlikely to be the source of the high serum hepatocyte growth factor levels observed in acute liver failure. PMID- 11117561 TI - Ursodeoxycholic acid suppresses extent of lipid peroxidation in diseased liver in experimental cholestatic liver disease. AB - The therapeutic benefit of ursodeoxycholic acid (UDCA) in treating cholestatic liver disease is globally recognized. It is generally accepted that the mechanism of action of UDCA can be attributed to several diverse processes that appear to be uniformly targeted towards minimizing the deleterious actions of accumulated hydrophobic bile acids in the cholestatic liver. Since hydrophobic bile acids are prooxidants, emerging in vitro evidence suggests that UDCA may have an antioxidant mechanism of action. We hypothesize that UDCA suppresses the extent of lipid peroxidation in the cholestatic liver. This hypothesis was tested by assessing the extent of lipid peroxidation in livers harvested from chronic bile duct ligated (CBDL) rats dosed daily for 24 days with 5, 10, or 15 mg/kg UDCA. The extent of lipid peroxidation was evaluated by determining the hepatic content of conjugated dienes, lipid peroxides, and malondialdehyde. The data were compared with identical data collected from unoperated control and 24-day bile duct manipulated (SO) rats. In the two groups of control rats, UDCA has no effect on the serum indices of liver function. In CBDL rats, UDCA suppressed the increased extent of lipid peroxidation in the liver in a dose-dependent manner in the absence of improvement of laboratory parameters of liver function and hepatic architecture. In conclusion, UDCA suppresses the augmented extent of lipid peroxidation in the diseased liver of CBDL rats. PMID- 11117562 TI - Fatty infiltration of liver in hyperlipidemic patients. AB - Hyperlipidemia is a known risk factor for fatty infiltration of the liver, a condition that can progress to cirrhosis and liver failure. The objectives of this study were to document the prevalence of fatty infiltration in the livers of hyperlipidemic patients and to identify the predictor variables associated with this condition. Over an 18-month recruitment period, clinical, biochemical, and radiologic assessments were performed in a cross-sectional manner in 95 adult patients referred to an urban hospital-based lipid clinic for evaluation and management of hyperlipidemia. The mean (+/-SD) age of the patients was 55 +/- 13 years. Forty-eight (51%) were male. Fifty-two patients (55%) had hypercholesterolemia, 25 (26%) severe hypertriglyceridemia, 14 (15%) mixed hyperlipidemia, and 4 (4%) moderate hypertriglyceridemia. Obesity and diabetes were present in 36 (38%) and 12 (12%) of cases, respectively. A total of 61 (64%) patients had elevated liver enzyme tests. The most common enzyme abnormalities were an elevated serum ALT in 45 (47%) and GGT in 43 (45%) of patients. Ultrasound findings revealed diffuse fatty liver in 47 patients (50%), of which 21 cases (22%) were mild, 18 (19%) moderate, and 8 (9%) severe. The majority of patients with hypercholesterolemia [35/52 (67%)] had normal ultrasounds, whereas severe hypertriglyceridemia and mixed hyperlipidemia were frequently associated with radiologic evidence of fatty liver (odds ratios 5.9 and 5.1 respectively, P < 0.01). Independent predictors of fatty liver were; AST (P = 0.001), hyperglycemia (P = 0.02), and age (P = 0.04). In a model incorporating known risk factors for fatty liver, diabetes was the only risk factor other than hypertriglyceridemia that was significantly associated with fatty infiltration. No such effect was seen with age, gender, obesity, or alcohol consumption. In conclusions, the results of this study indicate that ultrasonographic evidence of fatty infiltration of the liver is evident in approximately 50% of patients with hyperlipidemia. Hypertriglyceridemia is the lipid profile most often associated with this condition. Serum AST values, hyperglycemia, and age independently predict the presence of fatty infiltration, while hypertriglyceridemia and diabetes are the only risk factors that significantly increase the risk of fatty infiltration in hyperlipidemic patients. PMID- 11117563 TI - Correlation between stellate cell activation and serum fibrosis markers in choline-deficient L-amino acid-defined diet-induced rat liver fibrosis. AB - The aim of this study was to investigate the association of stellate cell activation with serum fibrosis markers in a rat model of hepatic fibrosis prepared using a choline-deficient L-amino acid (CDAA) defined diet. CDAA diet administration resulted in increased liver hydroxyproline contents in a time dependent manner with activated stellate cells, expressing alpha-smooth muscle actin (alpha-SMA) as well as increased serum concentrations of amino-terminal procollagen type III peptide (PIIIP) and the 7S fragment of type IV collagen. Hydroxyproline content of the liver showed a closer correlation with the serum 7S (r = 0.75, P < 0.01) concentration than with the serum PIIIP (r = 0.51, P < 0.01) concentration. The percent area of alpha-SMA-positive cells showed stronger correlation with the serum PIIIP concentration (r = 0.85, P < 0.01) than with the 7S concentration (r = 0.50, P < 0.01). These results indicate that the serum PIIIP concentration reflects the activity of fibrogenesis, while the serum 7S concentration reflects the accumulation of collagen fibers in the liver. PMID- 11117564 TI - Hepatocellular carcinoma in autoimmune hepatitis. AB - To determine if hepatocellular carcinoma can develop in autoimmune hepatitis in the absence of viral infection and to assess its frequency, liver tissue removed at hepatectomy was tested for HBV DNA and HCV RNA in one patient and the frequency of hepatocellular carcinoma was determined in 212 other uniformly followed individuals. The liver tissue from the propositus was uninfected and only one patient (0.5%) in the cohort undergoing routine follow-up developed malignancy during 1,732 patient-years of observation. Only one of 88 patients with cirrhosis (1%) developed hepatocellular carcinoma during 1,002 patient-years of observation after cirrhosis (mean, 123 +/- 9 months) and of the 65 patients with histological cirrhosis for at least five years, only one developed carcinoma during 162 +/- 8 months (incidence, 1 per 965 patient-years). We conclude that hepatocellular carcinoma can develop in autoimmune hepatitis in the absence of viral infection. Its occurrence is rare and only in long-standing cirrhosis. PMID- 11117565 TI - Serum alanine aminotransferase levels in relation to hepatitis B and C virus infections among drug abusers in an area hyperendemic for hepatitis B. AB - Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections are the major agents responsible for hepatitis in Taiwan. The purpose of this study was to assess the serum alanine aminotransferase (ALT) activity in relation to HBV and HCV infection among drug abusers. This survey included 769 male drug abusers aged 14-59 years, from the Kaohsiung Narcotic Abstention Institute and Kaohsiung Prision. The prevalence of HBsAg seropositivity was 21.5%, and anti-HCV seropositivity was 27.2%, respectively. Drug abusers with HBsAg or anti-HCV had higher serum AST and ALT levels than those without HBsAg and anti-HCV. The prevalence of raised ALT and AST (> or =45 IU/liter) in the HCV-positive group was more significant than in the negative group, while that of the HBsAg-positive group did not reach statistical significance. Among the HCV-positive group, ALT levels are more closely associated with HCV infection than AST levels. Our results indicated that HCV infection plays an important role in the etiology of raised ALT activity among drug abusers, while HBV infection plays a minor role. ALT screening still remains a simple and valuable method in the early recognition of HCV infection. PMID- 11117566 TI - Effectiveness of interferon treatment for patients with chronic hepatitis C virus infection and normal aminotransferase levels. AB - To determine the effects of interferon treatment, we studied 77 Japanese patients with hepatitis C virus (HCV) infection and normal alanine aminotransferase (ALT). Of 77 patients, 37 were given natural interferon-alpha for 24 weeks, and 40 not given interferon acted as controls. Serum samples were tested for HCV RNA and genotypes by polymerase chain reaction (PCR). HCV RNA levels were measured by competitive PCR. Of 37 treated patients, 11 (29.7%) had sustained elimination throughout a six-month follow-up, while HCV RNA was not eliminated in any untreated patients. At 24 months, the number of patients with elevated ALT was not significantly different between treated (13.5%) and untreated patients (15%). Interferon eliminates HCV RNA in patients with normal ALT without severe side effects. The natural history of HCV infection should be clarified so that the interferon treatment regimen can be tailored to the needs of each patient. PMID- 11117567 TI - Marked transaminase elevation in anorexia nervosa. PMID- 11117568 TI - Thrombocytemia as a predictor of portal hypertension in schistosomiasis. AB - Sufferers of schistosomiasis mansoni can evolve a clinical form of the disease associated with portal hypertension. To differentiate this form, routine clinical tests and biological indices were evaluated. In all, 54 HBsAg- and HCV-negative patients were studied, 42 with schistosomiasis and 12 normal volunteers. Using clinical criteria, ultrasonography, and endoscopy, the schistosomiasis patients were classified into two groups: mild chronic form (MS, N = 14) and chronic form associated with portal hypertension (PH, N = 28). The laboratory parameters of the MS group did not differ from the controls. The PH group differed from the others in prothrombin index, thrombocytemia, gamma-glutamyltransferase, serum alpha2-macroglobulin, and the calculated indices. ROC plot cutoff levels verified that isolated thrombocytemia was the most efficient marker for discrimination of the PH and MS forms. Thrombocytemia of 130 x 10(9) platelets/liter discriminated the groups with an 86% accuracy when all patients were analyzed and 96% when only schistosomiasis patients who did not consume alcohol were included. PMID- 11117569 TI - Effect of transjugular intrahepatic portosystemic shunt on thrombocytopenia associated with cirrhosis. AB - Thrombocytopenia is a frequent complication of cirrhosis. Its pathogenesis is not well known, but it has been suggested that splenic congestion induced by portal hypertension may be a major contributory factor. However, the available data regarding the effect of portal decompression either by surgical shunts or transjugular intrahepatic portosystemic shunt (TIPS) on peripheral platelet count in cirrhotics is conflicting. We studied the effects of TIPS on platelet count and mean platelet volume, following a successful TIPS placement. The platelet count had a tendency to decrease but was not statistically significant (120,100 +/- 72,100/mm3 before TIPS vs 99,800 +/- 51,400/mm3 after TIPS). The mean platelet volume remained essentially unchanged (9.8 +/- 1.5 fL before TIPS and 9.9 +/- 1.5 fL after TIPS). These results confirm that TIPS has an unpredictable effect on platelet count in cirrhotic patients with thrombocytopenia. The lack of a consistent increase in the peripheral mean platelet volume following TIPS placement suggests that TIPS is unable to significantly enhance the release of platelets sequestered in the splenic compartment in portal hypertension. PMID- 11117570 TI - Infected hepatic cyst masquerading as abdominal aortic aneurysm. PMID- 11117571 TI - Effects of ischemia-reperfusion on hepatic glutathione and plasmatic markers of graft function during in situ split-liver transplantation in adult recipients. AB - In situ split-liver transplantation is a new surgical technique where the bipartition of a single liver allows procurement of a right graft (segments I, IV, V-VIII) for an adult recipient (75% of the total liver volume), and a left graft (segments II and III) for a child recipient. The present study was designed to assess the effects of ischemia-reperfusion on right grafts obtained by in situ split-liver transplantation. To this aim, hepatic glutathione and conventional plasmatic markers of allograft function (alanine and aspartate aminotransferase, total bilirubin, prothrombin time, lactate dehydrogenase, gamma glutamyltranspeptidase, and alkaline phosphatase) were evaluated in four adult recipients. At the time of reperfusion, a marked glutathione decrease was found in the segment VI in three cases, whereas the amount of glutathione in segment IV was related to the duration of cold ischemia in all cases. Upon reperfusion, a marked increase in plasmatic alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase was found. A recovery in prothrombin time was observed from the first day in three cases. An increasing trend in total bilirubin, gamma-glutamyltranspeptidase, and alkaline phosphatase was noted from the second day after transplant. This preliminary study suggests a possible relationship between the duration of cold ischemia, amount of glutathione in segment IV of the right graft, and the trend in plasmatic markers of allograft damage during in situ split-liver transplantation in adult recipients. PMID- 11117572 TI - Comparative study of perioperative management of hepatic resection. AB - In an attempt to identify the factors that influence outcome after hepatic resection, patient background was reviewed and operative morbidity and mortality rates were assessed during two periods: 1985-1988 (group I: N = 96) and 1995-1998 (group II: N = 109). There were no differences in patient background factors between the two groups, but intraoperative blood loss, operative morbidity, and mortality were significantly reduced in group II compared to group I. There has been a significant reduction in postoperative complications, even in cases complicated by liver cirrhosis or obstructive jaundice. As a result of appropriate surgical procedures, postoperative complications in cirrhosis have been markedly decreased. After preoperative percutaneous biliary drainage in obstructive jaundice, attempts have been made to reduce the volume of blood loss even in extensive hepatectomy, the extent of liver resection in poor risk cases has been reduced without sacrificing radicality, and, by minimizing surgical stress, perioperative management has been greatly improved. PMID- 11117573 TI - Preventive effect of FK 506 (tacrolimus hydrate) on experimentally induced acute liver injury in rats. AB - The aim of the study was to investigate the effect of the immunosuppressant FK 506 (tacrolimus hydrate) on acute liver injury induced by Propionibacterium acnes and lipopolysaccharide (LPS). Acute liver injury was induced in male Wistar rats by injecting the animals with P. acnes (10 mg/rat), and administering LPS (10 microg/rat) seven days later. One group was given FK 506 (1 mg/kg) 24 and 2 hr before administration of LPS, and the other group was given the same dose of saline. The 24-hr survival rate, serum alanine aminotransferase (ALT) concentration, and tumor necrosis factor (TNF) -alpha mRNA and protein concentrations in the liver and spleen were then compared. Hepatic macrophages were also isolated from rats seven days after P. acnes injection, LPS, and FK 506 or saline were added to the culture supernatant, and TNF-alpha production was studied. The 24-hr survival rate was 100% in the FK 506-treated group, in contrast with 16.6% in the saline group. Four hours after LPS injection, the serum ALT concentration was 755 +/- 401 in the saline group versus 119 +/- 42 units/ml (P < 0.01) in the FK 506-treated group. The serum TNF-alpha concentration was lower in the FK 506-treated group (1,419 +/- 957 pg/ml) than in the saline group (9205 +/- 2215) (P < 0.01). The mRNA and protein concentrations in the liver and spleen in the two groups did not differ significantly 1 hr after LPS injection but were significantly lower in the FK 506-treated group after 4 hr. FK 506 did not directly inhibit TNF-alpha production by isolated cultured hepatic macrophages. FK 506 is unable to inhibit initial TNF-alpha production by hepatic macrophages (or probably that by splenic macrophages either) stimulated by injection of LPS in P. acnes + LPS-induced acute liver injury. However, the immunosuppressant does limit hepatic damage by inhibiting subsequent aggravation of inflammation by the cytokine network. PMID- 11117574 TI - Association of obesity and type II diabetes mellitus as a risk factor for gallstones. AB - Age, female sex, and obesity are well-known risk factors for gallstones; in contrast the possible role of type 2 diabetes mellitus (type-2 DM) is controversial. One reason for this discrepancy might be that type 2 DM is often accompanied by obesity. Therefore, the aim of this study was to evaluate the importance of obesity and of type 2 DM, separately and together, as risk factors for gallstones. In all, 203 obese patients with normal glucose tolerance (obese NGT), 446 obese patients with type 2 DM (obese type 2 DM), 269 lean patients with type 2 DM (lean type 2 DM) and 250 lean subjects with a normal glucose tolerance (lean NGT) were evaluated by ultrasonography for the presence of gallstones. At univariate analysis patients with gallstones (177) were older and were more frequently affected by both obesity and type 2 DM, and had higher triglycerides and fasting blood glucose levels. At multiple logistic regression analysis, only age and obesity, both in the presence or in absence of type 2 DM, were strongly associated with gallstones (P < 0.001); diabetes alone had a lower level of statistical significance (P = 0.07). These data suggest that obesity is a stronger risk factor for gallstones than type 2 DM. PMID- 11117575 TI - Polymorphism of cystic fibrosis gene in Japanese patients with chronic pancreatitis. AB - Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene and the 5T genotype of the polythymidine tract at the exon 9 splice branch/acceptor site are shown to be associated with chronic pancreatitis in Caucasian patients. In contrast to Western countries, cystic fibrosis is extremely rare in Japan. In this study, we investigated the association of mutations or polymorphisms of the CFTR gene with chronic pancreatitis in Japanese patients. Forty-seven patients with chronic pancreatitis (alcohol-related in 31, idiopathic in 14, and familial in 2) were examined for the deltaF508 and R117H mutations and polymorphisms of intron 8. DNA was extracted from leukocytes. Mutations and polymorphisms were examined by the allele-specific polymerase chain reactions and confirmed by direct sequencing. None of the patients had deltaF508 or R117H mutations in the CFTR gene. All of 47 healthy Japanese showed the homozygous 7T/7T genotype, whereas the frequencies of 5T, 7T, and 9T alleles were 0.043, 0.894, and 0.064 in the patients, respectively. The difference in allele frequency is statistically significant. Therefore, the present study indicates the association of polymorphism of the polythymidine tract in intron 8 of the CFTR gene with chronic pancreatitis in Japanese patients. PMID- 11117576 TI - Alcohol and aldehyde dehydrogenase polymorphisms in Japanese patients with alcohol-induced chronic pancreatitis. AB - In order to clarify the genetic factors in alcohol-related chronic pancreatitis among Japanese, we determined the genotype of two major alcohol-metabolizing enzymes, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH). The restriction fragment-length polymorphisms of the ADH2 and the ALDH2 genes were analyzed in 47 normal subjects and 31 patients with alcoholic pancreatitis. No significant difference between the patient and control groups was found in the ADH2 genotypes. A significant genetic difference between the two groups was found in the ALDH2 locus. The frequency of the ALDH2*1 allele was found to be 0.681 and that of the ALDH2*2 allele was 0.319 in the controls, while these values were 0.935 and 0.065 in the patients, respectively. Most of the patients (27 of 31) were ALDH2*1/2*1, only four were ALDH2*1/2*2, and none of the patients were ALDH2*2/2*2. These results indicate that genetic polymorphism of the ALDH2 gene influences the risk of developing alcoholic pancreatitis in Japanese. PMID- 11117577 TI - Are antibodies to carbonic anhydrase II specific for anti-mitochondrial antibody negative primary biliary cirrhosis? AB - Antibodies to carbonic anhydrase II (CAII) have been reported to be specific to anti-mitochondrial antibody (AMA)-negative primary biliary cirrhosis (PBC). We examined whether antibodies to CAII are specific for AMA-negative PBC or a nonspecific response in autoimmune liver disease. Antibody assays to CAII, by western immunoblot (dilution 1:200), were performed on sera from 16 AMA-negative PBC patients, 21 AMA-positive PBC patients, 21 autoimmune hepatitis type 1 (AIH) patients, and 18 alcoholic liver disease (ALD) patients. CAII antibody activity was found in 8 of 16 (50%) of the AMA-negative PBC patients, 9 of 21 (43%) of the AMA-positive PBC group, 10 of 21 (48%) of the AIH group, and in 3 of 18 (17%) of the ALD control group. There was no difference in the prevalence of CAII antibody reactivity between the AMA-negative PBC, AMA-positive PBC, and AIH groups. In conclusion, we determined that CAII antibodies are detected with equal frequency in AMA-positive PBC and AIH. Given that CAII antibodies have been reported in other nonhepatic autoimmune diseases, we conclude that CAII antibodies are likely a nonspecific marker of autoimmunity rather than specific for AMA-negative PBC. PMID- 11117578 TI - Combination analysis of genetic alterations and cell proliferation in small intestinal carcinomas. AB - We analyzed K-ras and p53 mutations, microsatellite instability (MSI), and loss of heterozygosity (LOH) using the polymerase chain reaction (PCR) methods, as well as p53 expression and the Ki-67 labeling index (LI) using immunohistochemistry in 10 duodenal and 10 jejunal/ileal carcinomas. K-ras mutations were detected in two duodenal (20%) and three jejunal/ileal (30%) carcinomas and p53 mutations in one (10%) and three (30%), respectively. LOH of 17p was detected in two duodenal (20%) and two jejunal/ileal (20%) carcinomas and p53 expression in four duodenal (40%) and four jejunal/ileal (40%). One duodenal (10%) and two jejunal/ileal (20%) carcinomas demonstrated MSI. LOH of APC was detected in three jejunal/ileal (30%), but none of the duodenal carcinomas. The Ki-67 LI was 44% in duodenal and 52.6% in jejunal/ileal carcinomas. A subset of small intestinal carcinomas showed involvement of K-ras and p53 mutations, LOH of APC, and MSI. A difference was also apparent for LOH of APC between duodenal and jejunal/ileal carcinomas. PMID- 11117579 TI - Effector Th-1 cells with cytotoxic function in the intestinal lamina propria of patients with Crohn's disease. AB - A large body of evidence points to a pivotal relationship between Th-1 cells and mucosal inflammation in Crohn's disease (CD). The aim of the present study was to assess whether CD is associated with specific functional activity of lamina propria T lymphocytes (LPT), particularly purified CD4, such as cytotoxic activity and specific cytokine-secreted profile. The results showed that CD4 LPT in patients displayed a chronically activated memory-like surface phenotype and, when compared to controls, had a significantly enhanced antibody-redirected cytotoxicity. Interestingly, the ratio of perforin expression in CD4 LPT was higher compared to controls, and a redirected lysis of human RBC mediated by a CD4 subset of intestinal lamina propria was evident, suggesting a cytolytic pore forming mechanism. Moreover, a unique Th-1 cytokine profile pattern in the CD4 cells from CD was defined. These effector cells produced 12 times more IFN-gamma, two times more TNF-alpha, and three times less IL-4 than controls. In contrast, no increase in IL-2 was detected, while IL-5 was undetectable. Our studies suggest that these preexisting in vivo activated CD4 LPT may play an important role in the inflammatory process in CD, thus directly contributing to the intestinal lesions. PMID- 11117580 TI - Increased production of IL-4 by gut T-cell lines from patients with dermatitis herpetiformis compared to patients with isolated gluten-sensitive enteropathy. AB - Dermatitis herpetiformis (DH) and isolated gluten-sensitive enteropathy (GSE) are gluten-sensitive diseases in which ingestion of dietary gluten results in the development of clinical disease. Patients with DH develop cutaneous IgA deposits and a severe skin disease, but rarely develop gastrointestinal symptoms. Patients with isolated GSE develop clinically significant gastrointestinal symptoms, but not skin disease or cutaneous IgA deposits. The aim of this study was to investigate the mechanism by which a mucosal immune response to the same dietary antigen can result in two distinct clinical phenotypes. T-cell lines were derived from activated T-cells in the small bowel mucosa of five patients with DH and 14 patients with isolated GSE and analyzed for T-cell markers and cytokine production in vitro. T-cell lines from DH and isolated GSE patients produced IFN gamma after stimulation (mean: DH = 2,619 pg/ml; isolated GSE = 1,993 pg/ml; NS). T-cell lines from patients with DH, however, produced significantly more IL-4 than the T-cell lines from patients with isolated GSE (IL-4: DH = 2,010 pg/ml; isolated GSE = 235 pg/ml; P < 0.05). Analysis of intracytoplasmic cytokine production by the T-cell lines showed that T-cell lines from patients with DH were CD4+ predominant, with a greater proportion of CD4+/IL4+ cells than CD4+/IFN gamma+ cells. In contrast, isolated GSE T-cell lines were predominantly CD8+, with an equal proportion of IL-4- and IFN-gamma-positive cells. These studies demonstrate that T cell lines from patients with DH produce significantly more IL 4 than T-cell lines from patients with isolated GSE, while producing similar amounts of IFN-gamma. This difference in cytokine pattern may play an important role in the different clinical manifestations of these two forms of gluten sensitivity. PMID- 11117581 TI - Chronic diarrhea impairs intestinal antioxidant defense system in rats at weaning. AB - The aim of the present study was to evaluate the influence of severe protein energy malnutrition on the antioxidant defense system in the small and large intestine in rats at weaning. Chronic diarrhea and the subsequent malnutrition were induced by oral intake of a lactose-enriched diet. Twenty rats were weaned at 21 days of age, and the control group was fed a semipurified synthetic diet for two weeks. The malnourished group was fed the same diet but carbohydrates were replaced by lactose, and they developed diarrhea one day after. Rats were killed, and macroscopic and histological features were analyzed, DNA content was measured, and alkaline phosphatase, myeloperoxidase, and gamma glutamyltranspeptidase activities were determined to assess the degree of intestinal injury. Glutathione levels as well as the activities of intestinal glutathione transferase, glutathione reductase, total glutathione peroxidase, selenium-dependent glutathione peroxidase, superoxide dismutase, and catalase were measured to study the antioxidant defense system. Malnourished rats showed loss of body weight and an increase in length and weight in jejunum and ileum, while no significant changes were observed in colon. Epithelial cells showed fewer and shorter microvilli, larger mitochondria with low inner density and loss of cristae, dilated endoplasmic reticulum, and Golgi apparatus. The protein-to DNA ratio was higher in the jejunum, ileum, and colon of malnourished rats. Glutathione levels decreased 40% in jejunum and 50% in colon of malnourished rats. A 40-50% decrease in the activity of all the enzymes of the antioxidant defense system was observed in the jejunum and ileum of malnourished rats, while only catalase and glutathione transferase activities decreased 50% in colon. These results suggest that early chronic diarrhea and severe protein-energy malnutrition impair the antioxidant defense system in both the small and large intestine, which may have a role in the pathogenesis and maintenance of the vicious circle of malabsorption-diarrhea-malnutrition in infancy. PMID- 11117582 TI - Neuromuscular and vascular hamartoma of the small bowel: case report and review of the literature. PMID- 11117583 TI - Association of fasting breath nitrous oxide concentration with gastric juice nitrate and nitrite concentrations and Helicobacter pylori infection. AB - Fasting breath nitrous oxide (N2O) concentration was determined in relation to pH, nitrate (NO3-) and nitrite (NO2-) concentrations in gastric juice, and Helicobacter pylori infection in 86 successive patients. N2O was measured with an infrared-photoacoustic analyzer. NO3- and NO2- were measured using the Griess reaction. Fasting breath N2O concentration in controls (87 +/- 21 ppb) was not significantly different from that in patients with gastric ulcer or other gastric lesions. Breath N2O was significantly correlated with gastric NO3- (P < 0.01) and was higher in patients with elevated gastric NO2- (246 +/- 87 ppb) than in patients without NO2- (75 +/- 13 ppb). Breath N2O did not differ significantly between subjects who were positive or negative for H. pylori. In conclusion, fasting breath N2O concentration is in some manner related to intragastric NO3- and NO2- concentrations. The possible use of measuring breath N2O as predictors of cancer needs further research. PMID- 11117584 TI - Expression of progastrin-derived peptides and somatostatin in fundus and antrum of nonulcer dyspepsia subjects with and without Helicobacter pylori infection. AB - The hypergastrinemia and hyperacidity associated with Helicobacter pylori infection has been explained by either a primary excess of gastrin or a lack of inhibitory influence by somatostatin (SOM). The objective of the present study was to compare the concentrations of fundic and antral SOM- and antral progastrin derived peptides in nonulcer dyspepsia (NUD) subjects with and without H. pylori infection. Antral and fundic mucosal biopsies were extracted and assayed for SOM and gastrin amide, glycine-extended gastrin (gastrin gly), progastrin, and total gastrin. There was a significant sixfold reduction in antral SOM but no change in fundic SOM content in H. pylori-infected subjects compared to uninfected subjects. Antral gastrin amide concentrations were significantly higher in infected subjects. However, the concentrations of the nonamidated gastrin forms (progastrin and glycine-extended gastrin) were significantly lower in the infected subjects, indicating an increased conversion of the precursor forms of gastrin to amidated gastrin, the type known to stimulate gastric acidity. The present study demonstrates that the elevated gastrin concentrations associated with H. pylori infection may be due to a reduction in the paracrine inhibitory effect of SOM on antral gastrin release. In addition, the posttranslational processing of gastrin to the amidated forms is increased in infected subjects, explaining why the elevation in antral gastrin is confined to the amidated form. PMID- 11117585 TI - Early initiation of post-PEG feeding: do published recommendations affect clinical practice? AB - Since its introduction in 1980, percutaneous endoscopic gastrostomy (PEG) has become the procedure of choice for long-term enteral feeding in patients who are unable to take adequate oral nutrition. Traditionally, gastroenterologists have waited for 24 hr after PEG tube placement before initiating feedings. However, recently published data have described the safety and efficacy of initiating PEG tube feedings as early as 3 hr after PEG tube placement. The objectives of this study were to determine how many hours after PEG tube placement practicing gastroenterologists initiate feedings and to determine whether they were aware of the pertinent literature. A four-page questionnaire was mailed to 35 practicing gastroenterologists, and the data gathered were analyzed using the SAS statistical package. A majority (81.5%) were aware of articles published in the last six years regarding the safety and efficacy of early initiation of feeding after PEG; of interest, however, only 10.7% were initiating feedings less than 3 hr after PEG tube placement. In conclusion, there exists a discrepancy between published recommendations and clinical practice regarding early initiation of feedings after PEG tube placement in this group of gastroenterologists who were surveyed. PMID- 11117586 TI - Effect of L-NMMA on postprandial transient lower esophageal sphincter relaxations in healthy volunteers. AB - In a previous study we showed that nitric oxide (NO) synthesis inhibition by NG monomethyl-L-arginine (L-NMMA) reduced the number of transient lower esophageal sphincter relaxations (TLESRs) triggered by gastric balloon distention. The role of NO in postprandial TLESRs and gastroesophageal reflux, however, is unknown. Therefore, we studied the effect of L-NMMA on meal-induced TLESRs and reflux episodes with simultaneous recording of esophageal peristalsis, intraesophageal and intragastric pH, and gastric emptying in healthy volunteers. Ingestion of a solid meal resulted in an increase in TLESRs [8.5 (6.3-11.0) 60 min] which was significantly inhibited by L-NMMA [6.0 (4.0-8.8) 60 min, P < 0.05]. In addition, the total number of reflux episodes was reduced. L-NMMA had no effect on intragastric meal distribution and gastric emptying, but attenuated the postprandial increase in intragastric pH. These results confirm the involvement of NO in the neurocircuitry underlying the triggering of TLESRs. The reduction in reflux by L-NMMA has to be confirmed in patients with gastroesophageal reflux disease. NO may be involved in the regulation of gastric acid secretion. PMID- 11117587 TI - Gastrointestinal bleeding in a woman with pulmonary tuberculosis. PMID- 11117588 TI - Achalasia presenting as acute airway obstruction. AB - Achalasia presenting as acute airway obstruction is an uncommon complication. We report the case of an elderly woman with previously undiagnosed achalasia who presented with acute respiratory distress due to megaesophagus. Emergency endotracheal intubation and insertion of a catheter into the esophagus, with continuous aspiration was required. Upon introduction of the esophageal catheter an abruptand audible air decompression occurred, with marked improvement of the clinical picture. Endoscopic injection of botulinum toxin was chosen as the definitive treatment with good clinical result. The pathophysiology of the phenomenon of esophageal blowing in achalasia is unclear, but different hypothetical mechanisms have been suggested. One postulated mechanism is an increase in upper esophageal sphincter (UES) residual pressure or abnormal UES relaxation with swallowing in achalasia patients. We reviewed the UES manometric findings in 50 achalasia patients and compared it with measurement performed in 45 healthy controls. We did not find any abnormalities in UES function in any of our achalasia patients group, or in the case under study. An alternative hypothesis postulates that airway compromise in patients with achalasia results from the loss UES belch reflex (abnormal UES relaxation during esophageal air distension), and in fact, an abnormal UES belch reflex was evidenced in our case. PMID- 11117590 TI - Parallel reliability of the functional independence measure and the Barthel ADL index. AB - PURPOSE: The aim of this study was to evaluate the concordance between the assessments of ADL according to the Functional Independence Measure (FIM) and the Barthel ADL index (BI) by means of a rank-invariant statistical method. METHOD: The construct validity, also called parallel reliability, of FIM and BI was assessed on the item level. Two different approaches to condensing the FIM assessments into 2-4 scale steps on the item level in order to calibrate the two ADL instruments were compared. One was determined by the theoretical operational definitions and the other defined by the empirical definitions. The 204 assessments of elderly persons were made three months after stroke by means of interviews. RESULTS: The parallel reliability of the FIM and the BI on the item level was strong, both according to the theoretical cut-off levels defined by the operational definitions and the empirical cut-off levels defined by the marginal distributions. CONCLUSIONS: The concordance between FIM and BI was high. There was a slight difference in favour of the operational definitions. The clinical key elements are to have a critical attitude towards ADL instruments; how they are constructed and operationalized. These are important elements in the quality assurance in the everyday work. PMID- 11117589 TI - Health care utilization by people with multiple sclerosis in The Netherlands: results of two separate studies. AB - PURPOSE: For chronically ill persons it is assumed that they make heavy demands on health care services. In the literature one hardly finds any publications to substantiate or refute this assumption. The main purpose of our study is to describe the health care utilization of people with multiple sclerosis (MS) in the Netherlands and its relationships with severity and duration of the disease. METHODS: Two different samples of persons with MS were used. In the first sample (University Hospital Groningen) severity of MS was based on medical judgement, while in the second sample (Dutch Multiple Sclerosis Society) severity was self reported. In both samples, use of health care facilities was assessed with a mail questionnaire. RESULTS: The methods for determining severity resulted in different distributions for severity of MS. However, the results were quite similar with respect to health care utilization. It appeared that the severity of MS was related to the number of professional caretakers MS-patients had contact with during one year. Duration of MS seemed not to be related to the number of caretakers. Only for specific caretakers, most notably GP, physiotherapist, home help and ergotherapist, the contact frequency increased with severity of MS. No such relationship was found between the frequency of contact with the neurologist and severity of MS. CONCLUSIONS: People with MS do not make a heavy demand on health care facilities in general but only on certain health care provisions. This is in contrast with the general notion that all chronically ill make a heavy demand on health care facilities in general. PMID- 11117591 TI - Prevalence and identification of problems in daily functioning in a primary medicine clinic. AB - PURPOSE: This pilot study compares scores on a health status/functional assessment measure to clinician identification of problems in functioning and referrals for these problems, based on examination of information in the patient's medical chart. METHOD: A sample of 194 participants at a primary medicine clinic in an urban general hospital completed a measure of health status and functioning, the Medical Outcomes Trust Short Form 36 (SF-36). Chart reviews were conducted to assess whether problems in functioning were addressed by the primary care clinician. RESULTS: Overall, levels of functioning on the scales of the SF-36 were well below norms for the general US population from the Medical Outcomes Study. Older adults showed lower physical functioning and higher emotional functioning than younger adults. Participants with 1, 2, or 3 chronic conditions showed increasingly lower levels of physical functioning. For participants with functional assessment scale scores in the lowest quartile, problems in functioning noted in the chart ranged from 13%-28%. Only 6% 20% of participants with marked problems in functioning were referred for further assessment or treatment. CONCLUSIONS: Functional problems are frequently important indicators of risk of development of secondary complications and need for referral. Questionnaire screening may increase identification and referral for problems in functioning in primary care settings. PMID- 11117592 TI - Self-body split: issues of identity in physical recovery following a stroke. AB - PURPOSE: To explore the perceived life and identity changes described by individuals following a single stroke using a life narrative approach. METHOD: Individuals admitted to hospital with a stroke, no previous disability, returning home; took part in life narrative interviews in hospital, and six months and one year post-discharge. The Gross Motor subscale of the Rivermead Motor Assessment and Nottingham 10 point Activities of Daily Living Scale were completed. RESULTS: Eight stroke respondents (five male, three female; mean age 67 years (range 56 82). The one year mean motor score was 9 (range 7-11) and self-care score was 9 (range 7-10). All respondents described a fundamental change in their lives and identity. The main issue was a split between themselves and their body. In hospital their body appeared to become separate, precarious and perplexing. By one year the majority still found their body unreliable, and their physical ability influenced by the social setting. CONCLUSION: The new experience of a split between self and body appears to be the focus of life for at least a year. This study suggests that rehabilitation professionals should consider longer-term (although not necessarily intensive) physical activity programmes that address these psychological as well as neuromuscular changes. PMID- 11117593 TI - Rehabilitation considerations of prosthetic fittings for Kaposi's sarcoma amputees. AB - PURPOSE: We report on cutaneous limb manifestations of Kaposi's sarcoma and the secondary infection of these lesions that necessitated five lower-limb amputations. METHOD: The cases are briefly described and prosthetic adaptations in respect to pressure, traction and sweating on the skin are considered. RESULTS: All four patients ambulated initially; one lady died, the double amputee stopped walking owing to the excessive physical demand, and two patients ambulate freely. CONCLUSION: Special considerations to the cutaneous/prosthesis interface are necessary in order to provide these patients with optimal ambulatory ability. PMID- 11117594 TI - Met and unmet needs reported by severely disabled people in southern England. AB - PURPOSE: To examine the met and unmet needs for rehabilitation of disabled people living in the community in Southern England. METHOD: A cross sectional interview study of people with a primary physical disability, aged 1665. Disabled people were randomly selected from two existing disability registers, which comprised disabled people who had been identified by community rehabilitation services as being in need of regular surveillance by formal assessment of their care needs. A new semi-structured needs assessment questionnaire was developed and validated for the study (the Southampton Needs Assessment Questionnaire, SNAQ). Level of disability was examined with the OPCS Disability and Severity Scales. RESULTS: Ninety three disabled people participated. Their median (IQR) OPCS score was 8 (6 10). Participants reported a median (IQR) of three unmet needs (2-7). The most prevalent unmet needs were for adaptations, equipment, physiotherapy and wheelchairs, rather than unmet needs for intellectual and social fulfilment. CONCLUSIONS: Disabled people who were already in touch with community rehabilitation services continued to express unmet needs for further services. Meeting the more basic needs relating to people's housing, equipment, physiotherapy and wheelchairs may enable them to be more independent and fulfilled in other areas of their lives. PMID- 11117595 TI - The active urine collection device: a novel continence management system focusing particularly on the needs of disabled women. AB - PURPOSE: This paper introduces a novel incontinence management system which applies space technology to the problem of urinary incontinence. The purpose of this paper is to invite comments and suggestions from professionals in the disability and rehabilitation field. METHOD: Severe urinary incontinence is a distressing condition for which there are few satisfactory management solutions. We have been able to develop a compact liquid handling system that can cope with the initial surge of urine which characteristically reaches a high flow rate of around 25 ml/sec. By using a sophisticated system of filters, the device can cope with mixed streams of urine and air which are inevitable when a non-invasive patient/device interface is used. The next phase of the project is to develop prototype devices with the aid of users to produce an effective appliance that users and carers find practical and acceptable to use. CONCLUSION: With the aid of users, carers and rehabilitation experts we hope to develop an excellent user friendly product. We believe this device can make a positive contribution to the quality of life of disabled people with continence difficulties. PMID- 11117602 TI - State of the environment in the arrangement area of the enterprises for repairing and utilization of nuclear-powered submarines. AB - The influence of nuclear-powered utilization (disjunction) upon the state of health of the soil, vegetation and atmospheric air was studied. It was stated that the concentration of hazardous metals in the air of an industrial site did not exceed the permissible levels. In the residential area the cases of increased concentrations of manganese and chromium were noted. The major pollutants of vegetation are manganese, titanium, copper and nickel. The authors propose a complex of anthropogenic factors to be the cause of the environmental contamination by hard metals. The volume activity of radioactive aerosols in the studied site is confined to the local hum. PMID- 11117608 TI - Genetic epidemiologic studies on age-specified traits. NIA Aging and Genetic Epidemiology Working Group. AB - This commentary calls attention to the value of combining genetic and epidemiologic methods in studies to understand the determinants of two crucial aspects of aging: ages at which certain outcomes (e.g., disease, mortality) occur and rates of change with age of individual's characteristics (e.g., physiologic functions, disease risk factors). Inclusion of age in the specification of traits in genetic epidemiologic studies could lead to improved strategies to increase healthy life expectancy and evaluate individuals' risk for age-related morbidity. Special issues that make genetic epidemiologic approaches important for studies of age-specified phenomena as well as opportunities and challenges for such studies are discussed, including study designs, sampling frames, databases, analytic tools, and related methodological issues. This commentary is based on a report prepared by the Aging and Genetic Epidemiology Working Group, convened by the National Institute on Aging to review opportunities for research on the genetic epidemiology of aging-related outcomes. The report, which contains more extensive discussion, literature review, and references, is available on the World Wide Web at http://www.nih.gov/nia/conferences/GeneticReport111199.htm. PMID- 11117609 TI - Impact of prenatal glucose screening on the diagnosis of gestational diabetes and on pregnancy outcomes. AB - The authors examined the impact of universal screening on the diagnosis of gestational diabetes and its complications. All mothers and newborns registered by the Canadian Institute for Health Information from 1984 to 1996 (even-numbered fiscal years only) were included in the analysis. Over this time period, the proportion of women with gestational diabetes increased ninefold (from 0.3% to 2.7%) while the proportion with prepregnancy diabetes fell from 0.7% to 0.4%. As rates of gestational diabetes increased, a corresponding reduction in the risks of complications (polyhydramnios, amniotic cavity infection, cesarean delivery, and preeclampsia) occurred for women with gestational diabetes. The incidence of gestational diabetes fell in Metro-Hamilton (where screening was discontinued in 1989) but remained high in the rest of Ontario (where screening continued in most areas). No related temporal trends for fetal macrosomia, cesarean delivery, or other diabetes-related complications were observed, regardless of screening policy. The authors concluded that the substantial increase in gestational diabetes in Canada is an artifact caused by universal screening, with no evidence of beneficial effects on pregnancy outcomes. PMID- 11117610 TI - Neural tube defects among Mexican Americans living on the US-Mexico border: effects of folic acid and dietary folate. AB - Populations of Mexican descent have high occurrences of neural tube defects (NTDs). A recent study suggested that folic acid supplements may not protect these populations from NTDs. In a case-control study, the authors investigated the role of folic acid and dietary folate intake in NTD risk among Mexican Americans living along the Texas-Mexico border. From January 1995 to February 1999, 148 Mexican-American women with NTD-affected pregnancies and 158 women with normal live births were interviewed in person about use of vitamin supplements and dietary intakes during a 6-month periconceptional period (from 3 months before conception to 3 months after conception). Daily preconceptional consumption of vitamin supplements containing folic acid was 2.5% in control women and 2.0% in case women (odds ratio = 0.77; 95% confidence interval (CI): 0.19, 3.22). With adjustment for maternal age, education, obesity, and previous stillbirth or miscarriage, the risk estimate was essentially null (odds ratio = 1.12; 95% CI: 0.22, 5.78). Combined folic acid intake from diet and supplements showed only a modest risk reduction for intakes of > or = 1.0 mg per day (adjusted odds ratio = 0.73; 95% CI: 0.31, 1.72). The fact that the primary folic acid exposure was in the form of dietary polyglutamates rather than the more easily absorbed supplemental monoglutamates may explain an apparent decreased effect in this population. PMID- 11117611 TI - Prenatal and postnatal risk factors for mental retardation among children in Bangladesh. AB - This study evaluated the contribution of prenatal, perinatal, neonatal, and postnatal factors to the prevalence of cognitive disabilities among children aged 2-9 years in Bangladesh. A two-phase survey was implemented in 1987-1988 in which 10,299 children were screened for disability. In multivariate analyses, significant independent predictors of serious mental retardation in rural and urban areas included maternal goiter (rural odds ratio (OR) = 5.14, 95% confidence interval (CI): 1.23, 21.57; urban OR = 4.82, 95% CI: 2.73, 8.50) and postnatal brain infections (rural OR = 29.24, 95% CI: 7.17, 119.18; urban OR = 13.65, 95% CI: 4.69, 39.76). In rural areas, consanguinity (OR = 15.13, 95% CI: 3.08, 74.30) and landless agriculture (OR = 6.02, 95% CI: 1.16, 31.19) were also independently associated with the prevalence of serious mental retardation. In both rural and urban areas, independent risk factors for mild cognitive disabilities included maternal illiteracy (OR = 2.48, 95% CI: 0.86, 7.12), landlessness (OR = 4.27, 95% CI: 1.77, 10.29), maternal history of pregnancy loss (OR = 2.61, 95% CI: 0.95, 7.12), and small for gestational age at birth (OR = 3.86, 95% CI: 1.56, 9.55). Interventions likely to have the greatest impact on preventing cognitive disabilities among children in Bangladesh include expansion of existing iodine supplementation, maternal literacy, and poverty alleviation programs as well as prevention of intracranial infections and their consequences. Further population-based studies are needed to confirm and understand the association between consanguinity and serious cognitive disability. PMID- 11117612 TI - Association of coffee consumption with gallbladder disease. AB - Coffee consumption was recently shown to protect against symptomatic gallbladder disease in men. The authors examined the relation of ultrasound-documented gallbladder disease with coffee drinking in 13,938 adult participants in the Third National Health and Nutrition Examination Survey, 1988-1994. The prevalence of total gallbladder disease was unrelated to coffee consumption in either men or women. However, among women a decreased prevalence of previously diagnosed gallbladder disease was found with increasing coffee drinking (p = 0.027). These findings do not support a protective effect of coffee consumption on total gallbladder disease, although coffee may decrease the risk of symptomatic gallstones in women. PMID- 11117613 TI - Psychiatric and sociodemographic predictors of attrition in a longitudinal study: The Netherlands Mental Health Survey and Incidence Study (NEMESIS). AB - This article discusses the effects of sociodemographics and the presence of psychiatric disorders diagnosed in the 12 months before the first interview by using the Diagnostic and Statistical Manual of Mental Disorders: DSM-III-R, third edition, revised, on three types of attrition (failure to locate, refusal to participate, morbidity/mortality) in the second wave (1997-1998) of the Netherlands Mental Health Survey and Incidence Study, a longitudinal, general population survey of psychopathology among 7,076 subjects aged 18-64 years. Compared with those reinterviewed successfully, persons not located at the 1-year follow-up (n = 219) were more often younger, poorly educated, urban, not cohabiting with a steady partner, and born outside the Netherlands. Refusers (n = 923) had a lower educational level. Morbidity/mortality (n = 72) was associated with higher age, lower educational level, not being employed, and somatic disorders. After adjustment for sociodemographics, none of the disorders was positively associated with refusal. Failure to locate was linked to agoraphobia, alcohol abuse, and the categories of mood, substance use, and eating disorders. Morbidity/mortality was linked to dysthymia, agoraphobia, simple phobia, obsessive-compulsive disorder, and the category of anxiety disorders. Overall attrition was only slightly higher among respondents with one or more disorders (odds ratio = 1.20, 95% confidence interval: 1.04, 1.38). Thus, psychopathology has only weak-to-moderate effects on attrition and is mainly related to failure to locate and morbidity/mortality but not to refusal. PMID- 11117614 TI - Depressive symptoms in Hispanic and non-Hispanic White rural elderly: the San Luis Valley Health and Aging Study. AB - Literature on depression in rural and Hispanic elderly adults is sparse. This report describes the prevalence of depressive symptoms in 1,151 community dwelling, Hispanic and non-Hispanic White participants in the San Luis Valley Health and Aging Study, conducted in rural Colorado during 1993-1995. The prevalence and odds ratios of high depressive symptoms, defined as a Center for Epidemiologic Studies Depression Scale score of > or = 16, were calculated. The crude prevalence of high depressive symptoms was 11.4% (95% confidence interval: 9.6, 13.6). Female gender, chronic diseases, dissatisfaction with social support, living alone, and lower income and education were associated with depressive symptoms. There were no ethnic differences in the men. The age-adjusted odds ratio of depressive symptoms in Hispanic women compared with that of non-Hispanic White women was 2.11 (95% confidence interval: 1.32, 3.38). After adjustment for multiple sociodemographic and health risk factors, the odds ratio in Hispanic women was 2.12 (95% confidence interval: 1.19, 3.80). Higher depressive symptoms in Hispanic women varied by acculturation level. The odds ratio in the high acculturation stratum was 1.56 (95% confidence interval: 0.75, 3.27) and in the low acculturation stratum was 2.51 (95% confidence interval: 1.11, 5.70). A lower acculturation level may increase the risk for depression in older Hispanic women. PMID- 11117615 TI - Dietary fat in relation to risk of multiple sclerosis among two large cohorts of women. AB - Ecologic correlations suggest that higher intake of saturated fat and lower intake of polyunsaturated fat might increase the risk of multiple sclerosis (MS), but the results of case-control studies have been inconsistent. Because no prospective data are available, the authors examined these associations in two large cohorts, the Nurses' Health Study, which consisted of 92,422 women with 14 years of follow-up (1980-1994) and the Nurses' Health Study II, which consisted of 95,389 women with 4 years of follow-up (1991-1995). They documented 195 new cases of MS. The pooled multivariate relative risks comparing women in the highest quintile with those in the lowest were 1.1 (95% confidence interval: 0.7, 1.7) for total fat, 0.7 (95% confidence interval: 0.5, 1.2) for animal fat, 1.2 (95% confidence interval: 0.7, 2.1) for vegetable fat, 0.8 (95% confidence interval: 0.5, 1.3) for saturated fat, 1.1 (95% confidence interval: 0.7, 1.7) for monounsaturated fat, 1.7 (95% confidence interval 1.0, 2.8) for n-6 polyunsaturated fat, 1.3 (95% confidence interval: 0.8, 2.0) for trans unsaturated fat, and 0.7 (95% confidence interval: 0.4, 1.1) for cholesterol. Omega-3 fatty acids from fish were also unrelated to risk. However, the authors observed a nonsignificantly lower risk of MS for a higher intake of linolenic acid. These findings do not support relations between intakes of total fat or major specific types of fat and the risk of MS. PMID- 11117616 TI - Serum carotenoids and markers of inflammation in nonsmokers. AB - One explanation for discrepant results between epidemiologic studies and randomized trials of beta-carotene and cardiovascular disease may be a failure to consider inflammation as a confounder. To evaluate the potential for such confounding, the authors relate the serum concentrations of five carotenoids (alpha-carotene, beta-carotene, beta-cryptoxanthin, lycopene, and lutein/zeaxanthin) to levels of three inflammatory markers (C-reactive protein, fibrinogen, and white blood cell count) measured during the Third National Health and Nutrition Survey, 1988-1994. The analysis included 4,557 nonsmoking participants aged 25-55 years. Adjusted concentrations of all five carotenoids were significantly lower in those with C-reactive protein levels above 0.88 mg/dl (p = 0.001). There was a trend toward lower adjusted beta-cryptoxanthin concentrations with increasing level of fibrinogen (p value test for trend = 0.01), but other carotenoids were not related. Many of the carotenoid concentrations were lower among participants with high white blood cell counts. After log transformation, only adjusted mean beta-carotene levels were significantly lower in those with white blood cell counts above 7.85 x 10(9)/liter (p < 0.01). These cross-sectional data do not clarify the biologic relation between carotenoids and C-reactive protein but, to the extent that the carotenoids are associated with C-reactive protein levels, a carotenoid-heart disease association may be, in part, an inflammation-heart disease association. PMID- 11117617 TI - Validation of three food frequency questionnaires and 24-hour recalls with serum carotenoid levels in a sample of African-American adults. AB - The validity of self-reported fruit and vegetable intake in minority populations has not been adequately established. In this study, the authors examined the association of three food frequency questionnaires (FFQs) and 24-hour dietary recalls with serum carotenoid levels. Approximately 1,000 African-American adults recruited from 15 churches in Atlanta, Georgia (1997-1998) completed three fruit and vegetable FFQs: a seven-item instrument assessing intake during the past month; a two-item measure assessing usual intake; and a 36-item measure adapted from the Health Habits and History Questionnaire. A total of 414 participants received a 24-hour recall by telephone, and 105 of them received two additional recalls. Serum levels of lycopene, lutein, cryptoxanthin, alpha-carotene, and beta-carotene were assessed in 813 participants and used as the validity criterion. The correlations of fruit and vegetable servings with specific and total serum carotenoid levels were generally higher for the 36-item FFQ than for the two-item and seven-item instruments. The strongest correlation of fruit and vegetable servings with total carotenoid levels was observed for the three recalls (r = 0.42), with the 36-item FFQ and the single 24-hour recall yielding comparable correlations (r = 0.35 and r = 0.37, respectively). The validity of the 36-item fruit and vegetable FFQ was generally as strong as the validity of both 1 and 3 days of recalls. Given the lower cost and time needed for administration relative to recalls, it appears that the 36-item FFQ has merit for evaluating fruit and vegetable health interventions. PMID- 11117618 TI - Vegetable and fruit consumption and risks of colon and rectal cancer in a prospective cohort study: The Netherlands Cohort Study on Diet and Cancer. AB - The relation between vegetable and fruit consumption and colorectal cancer risk was comprehensively assessed in the Netherlands Cohort Study on Diet and Cancer using a validated 150-item food frequency questionnaire. After 6.3 years of follow-up (1986-1992), over 1,000 incident cases of colorectal cancer were registered. Using case-cohort analysis, the authors calculated rate ratios and 95% confidence intervals adjusted for age, alcohol intake, and family history of colorectal cancer. For colon cancer, no statistically significant associations with total vegetable intake or total fruit intake were found. However, among women, an inverse association was observed with vegetables and fruits combined (for the highest quintile vs. the lowest, the rate ratio was 0.66 (95% confidence interval: 0.44, 1.01)). Brassica vegetables and cooked leafy vegetables showed inverse associations for both men and women. Among women and, to a lesser extent, among men, inverse associations were stronger for distal colonic tumors than for proximal colonic tumors. For rectal cancer, no statistically significant associations were found for vegetable consumption or fruit consumption or for specific groups of vegetables and fruits; only Brassica vegetables showed a positive association in women. As in other cohort studies, the observed inverse relation between vegetable and fruit consumption and occurrence of colorectal cancer was less strong than relations reported in case-control studies. PMID- 11117619 TI - Use of twins as mutual proxy respondents in a case-control study of breast cancer: effect of item nonresponse and misclassification. AB - A case-control study of breast cancer in twins diagnosed before 1988 was used to characterize the effects on odds ratios when proxy responses from co-twins are used. North American disease-discordant pairs were ascertained through advertisements, and mailed questionnaires were returned from both members of 671 pairs and from one member of 391 pairs. Biases from the proxy response were attributed to nonresponse or misclassification. Nonresponse varied according to type of exposure variable, depth of detail requested, joint exposure status of the pair, respondent's case-control status, zygosity, and social closeness of the pair. Misclassification was minimal, generally nondifferential, and a high degree of reliability between the proxy and self-report was indicated by the kappa statistic and the intraclass correlation coefficient. By using double-respondent pairs, a method was developed to adjust proxy responses for both sources of bias. These adjustments resulted in minor changes to the odds ratios for the variables studied (age at menarche, reproductive factors, and hormone use). A larger difference was observed between the odds ratios based on all pairs and those based on double-respondent pairs only. These findings demonstrate that, for these variables in this population, twins are reliable proxies for each other and that results from single-respondent pairs should be included. PMID- 11117620 TI - Re: "antidepressant medication use and breast cancer risk. PMID- 11117621 TI - Re: "antidepressant medication use and breast cancer risk". PMID- 11117622 TI - The anatomy of language: contributions from functional neuroimaging. AB - This article illustrates how functional neuroimaging can be used to test the validity of neurological and cognitive models of language. Three models of language are described: the 19th Century neurological model which describes both the anatomy and cognitive components of auditory and visual word processing, and 2 20th Century cognitive models that are not constrained by anatomy but emphasise 2 different routes to reading that are not present in the neurological model. A series of functional imaging studies are then presented which show that, as predicted by the 19th Century neurologists, auditory and visual word repetition engage the left posterior superior temporal and posterior inferior frontal cortices. More specifically, the roles Wernicke and Broca assigned to these regions lie respectively in the posterior superior temporal sulcus and the anterior insula. In addition, a region in the left posterior inferior temporal cortex is activated for word retrieval, thereby providing a second route to reading, as predicted by the 20th Century cognitive models. This region and its function may have been missed by the 19th Century neurologists because selective damage is rare. The angular gyrus, previously linked to the visual word form system, is shown to be part of a distributed semantic system that can be accessed by objects and faces as well as speech. Other components of the semantic system include several regions in the inferior and middle temporal lobes. From these functional imaging results, a new anatomically constrained model of word processing is proposed which reconciles the anatomical ambitions of the 19th Century neurologists and the cognitive finesse of the 20th Century cognitive models. The review focuses on single word processing and does not attempt to discuss how words are combined to generate sentences or how several languages are learned and interchanged. Progress in unravelling these and other related issues will depend on the integration of behavioural, computational and neurophysiological approaches, including neuroimaging. PMID- 11117623 TI - A comparative study of mammalian tracheal mucous glands. AB - We have compared the distribution, numbers and volume of mucous glands in the tracheas of 11 mammalian species. No glands were present in the rabbit. The mouse only contained glands at the border between the trachea and larynx. In the rat, glands were commonest in the cephalad third of the trachea, but on average were much scarcer than in the larger species. Between species, there was a significant correlation between airway diameter and gland volume per unit surface area, suggesting that the rate of deposition of inhaled particles may increase in large airways. In the ventral portion of the trachea of about half the species, the glands were concentrated between the cartilaginous rings; in others they were evenly distributed over and between the rings. In most species in which the trachealis muscle attached to the internal surface of the cartilaginous rings, the glands were external to the muscle. In all species in which the muscle attached to the external surface of the cartilaginous rings, the glands were internal to the muscle. In the ox, goat, dog and sheep, the volume of glands per unit tracheal surface area was markedly greater in the ventral than the dorsal aspect of the trachea. The reverse was true of the pig. In humans, gland density in the 2 regions was similar. The frequency of gland openings was determined in the ox, goat, pig, dog and sheep tracheas, and ranged from 0.3 per mm2 in the dorsal portion of the sheep trachea to 1.5 per mm2 in the ventral portion of the ox trachea. For these 5 species, the volume of gland acini per unit luminal surface area varied linearly with the numbers of gland openings, with the volume of individual glands being constant at approximately 120 nl. PMID- 11117624 TI - Curving and looping of the internal carotid artery in relation to the pharynx: frequency, embryology and clinical implications. AB - Variations of the course of the internal carotid artery in the parapharyngeal space and their frequency were studied in order to determine possible risks for acute haemorrhage during pharyngeal surgery and traumatic events, as well as their possible relevance to cerebrovascular disease. The course of the internal carotid artery showed no curvature in 191 cases, but in 74 cases it had a medial, lateral or ventrocaudal curve, and 17 preparations showed kinking (12) or coiling (5) out of a total of 265 dissected carotid sheaths and 17 corrosion vascular casts. In 6 cases of kinking and 2 of coiling, the internal carotid artery was located in direct contact with the tonsillar fossa. No significant sex differences were found. Variations of the internal carotid artery leading to direct contact with the pharyngeal wall are likely to be of great clinical relevance in view of the large number of routine procedures performed. Whereas coiling is ascribed to embryological causes, curving is related to ageing and kinking is thought to be exacerbated by arteriosclerosis or fibromuscular dysplasia with advancing age and may therefore be of significance in relation to the occurrence of cerebrovascular symptoms. PMID- 11117625 TI - Intact myelinated fibres in biopsies of ventral spinal roots after preganglionic traction injury to the brachial plexus. A proof that Sherrington's 'wrong way afferents' exist in man? AB - Bell-Magendie's law of separation of spinal function states that afferent and efferent fibres join the spinal cord separately in ventral and dorsal spinal nerve roots. For over 100 years there have been reports that challenge the exclusiveness of this law in mammals; very few studies have referred to man. We conducted a prospective morphological study in patients with preganglionic traction injuries of the brachial plexus to address this question. Avulsed ventral and dorsal roots were examined after variable intervals from the injury for histological and ultrastructural evidence for myelinated afferent fibres entering the cord via the ventral roots. Intact myelinated fibres were found in all ventral root specimens, but the majority of fibres in later biopsies are regenerative. A small number of fibres could be demonstrated that are likely to be 'wrong way ventral afferents'. Their number is falsely low due to wallerian degeneration of dorsal and ventral afferents following the mechanical and ischaemic effects of traction injury. Our findings are the first morphological evidence in human material that Bell-Magendie's law might not entirely be correct and they underline the difficulties in comparing traumatic with experimental rhizotomy. PMID- 11117626 TI - The thymus in the mouse changes its activity during pregnancy: a study of the microenvironment. AB - The mouse thymus changes dramatically during pregnancy. It shrinks in size, and the cortex is extensively reduced from midpregnancy onwards. Despite this, there is surprisingly little evidence for any increase in apoptosis, and considerable evidence that mitosis of thymocytes continues throughout pregnancy. In spite of overall involution the thymic medulla actually expands in midpregnancy due to a combination of mitosis of epithelial cells and an accumulation of lymphocytes. The extent and nature of these changes are examined in this study at the ultrastructural level. The epithelial cells of the subcapsular cortex (type 1 cells) become wrinkled and exhibit powers of phagocytosis, whilst the other cortical epithelial cells are relatively unchanged, although the formation of epithelial/thymocyte rosettes and thymic nurse cells is more clearly seen in midpregnancy than usual. Other changes associated with pregnancy involve the medullary epithelial cells that undergo an increased level of mitosis. Their greater numbers surround accumulations of lymphocytes to form the characteristic medullary epithelial rings. Cell movement through blood vessel walls was clearly observed in midpregnancy, but not at other times. Interdigitating cells in the medulla become more conspicuous as pregnancy proceeds and the cells become phagocytic. The endoplasmic reticulum in plasma cells becomes expanded, indicating increased secretory activity. These results highlight the active nature of the thymus in pregnancy in spite of its involution. This picture contradicts the conventional notion that an involuted thymus is inactive. PMID- 11117627 TI - Visualisation of three-dimensional microcracks in compact bone. AB - Microdamage in bone contributes to the loss of bone quality in osteoporosis and is thought to play a major role in both fragility and stress fractures (Schaffler et al. 1995). In this study, in vivo microcracks in human ribs were bulk-stained in basic fuchsin and viewed in longitudinal section and in 3 dimensions using 2 different computer-based methods of reconstruction: (1) serial sectioning of methylmethacrylate embedded sections using a sledge macrotome and identification of microcracks using UV epifluorescence followed by computerised reconstruction of microcracks using software and (2) laser scanning confocal microscopy of thick sections followed by reconstruction of microcracks into a 3-D image. The size and shape of microcracks were found to be similar using both techniques. Both techniques of reconstruction showed microcracks to be approximately elliptical in shape. From the serial sectioning reconstructions (n = 9), microcracks were found to have a mean length of 404 +/- 145 microm (mean +/- S.D.) (in the longitudinal direction) and mean width of 97 +/- 38 microm (in the transverse direction). Using epifluorescence microscopy, 92 microcracks were identified; mean microcrack length was 349 +/- 100 microm in the longitudinal direction. This was consistent with other results (Burr & Martin, 1993) and with the theoretical prediction of an elliptical crack shape with aspect ratio (longitudinal: transverse) of 5:1 deduced from analysis of random 2-D sections (Taylor & Lee, 1998). The results obtained provide new data on the nature of microcracks in bone and the method has the potential to become a useful tool in the calculation of stress intensity values which indicate the probability of an individual microcrack propagating to cause a stress or fragility fracture. PMID- 11117628 TI - The human vomeronasal organ. Part II: prenatal development. AB - During the 20th century, the human vomeronasal organ (VNO) has been controversial regarding its structure, function, and even identity. Despite reports that provide evidence for its presence throughout prenatal and postnatal ontogeny, some studies and numerous textbooks declare its absence in late fetal and postnatal humans. To that end, the present study was designed to establish firmly whether the human VNO is homologous with that of other mammals and whether it degenerates (partially or completely) or persists throughout prenatal development. Fifty human embryos and fetuses (33 d to 32 wk fertilisation age) and 2 neonates were examined by light microscopy. Four embryonic primates (mouse lemurs) were examined for a comparison of VNO embryogenesis. The presence or absence and structural characteristics of the VNO and supporting tissues are described. The first appearance of the VNO was in the form of bilateral epithelial thickenings of the nasal septum, the vomeronasal primordium. The primordia invaginated between 37 and 43 d of age and formed the tubular VNO. The tubular VNO was located dorsally at a variable distance from, but was always spatially separated from the paraseptal cartilages. The mouse lemurs examined in this study and other reports from the literature indicate that the human VNO resembles that of primates having functional VNOs until just after a tubular VNO is formed. Examination of the VNO and adjacent tissues suggested that the VNO may lose receptor cells and corresponding vomeronasal nerves and become a ciliated, pseudostratified epithelium between approximately 12 and 14 wk of age. Our findings indicate the prenatal human VNO goes through 3 successive stages: early morphogenesis, transformation (of the epithelium), and growth. These observations indicated that (1) all embryonic humans develop a vomeronasal organ which is homologous with the VNOs of other mammals, but which has become displaced and highly variable in relative location during embryogenesis; (2) the human vomeronasal organ does not degenerate prenatally, but very likely loses the functional components of the vomeronasal complex of other mammals; and (3) the remnant of the human VNO persists until birth and beyond. PMID- 11117629 TI - Morphology of human intracardiac nerves: an electron microscope study. AB - Since many human heart diseases involve both the intrinsic cardiac neurons and nerves, their detailed normal ultrastructure was examined in material from autopsy cases without cardiac complications obtained no more than 8 h after death. Many intracardiac nerves were covered by epineurium, the thickness of which was related to nerve diameter. The perineurial sheath varied from nerve to nerve and, depending on nerve diameter, contained up to 12 layers of perineurial cells. The sheaths of the intracardiac nerves therefore become progressively attenuated during their course in the heart. The intraneural capillaries of the human heart differ from those in animals in possessing an increased number of endothelial cells. A proportion of the intraneural capillaries were fenestrated. The number of unmyelinated axons within unmyelinated nerve fibres was related to nerve diameter, thin cardiac nerves possessing fewer axons. The most distinctive feature was the presence of stacks of laminated Schwann cell processes unassociated with axons that were more frequent in older subjects. Most unmyelinated and myelinated nerve fibres showed normal ultrastructure, although a number of profiles displayed a variety of different axoplasmic contents. Collectively, the data provide baseline information on the normal structure of intracardiac nerves in healthy humans which may be useful for assessing the degree of nerve damage both in autonomic and sensory neuropathies in the human heart. PMID- 11117630 TI - Ultrastructural localisation of NADPH-d/nNOS expression in the superior cervical ganglion of the hamster. AB - This study examined NADPH-d and nNOS expression in the SCG of hamsters. By light microscopy, numerous NADPH-d/NOS positive processes were widely distributed in the ganglion. Ultrastructurally, the NADPH-d reaction product was associated with the membranous organelles of neuronal soma, dendrites, myelinated fibres, small granular cells, and axon profiles bearing agranular vesicles. The NOS immunoreaction product, on the other hand, was localised in the cytoplasm of principal neurons and dendrites. Some of the NADPH-d/NOS labelled processes formed junctional contacts including synapses or zonulae adherentia. Compared with the neurons, the nonneuronal cells in the ganglion, namely, macrophages, satellite cells and endothelial cells were labelled by NADPH-d but devoid of nNOS immunoreaction product. The results suggest that the NADPH-d/NOS positive fibres in the SCG originate not only from the projecting fibres of the lateral horns of thoracic spinal cord, but also from the principal neurons and small granular cells; some may represent visceral afferent fibres. Electron microscopic morphometry has shown that about 67% of the principal neurons contain NADPH-d reaction product, and that the majority were small to medium sized neurons based on cross-sectional areas in image analysis. On the basis of the present morphological study, it is concluded NO is produced by some local neurons and possibly some nonneuronal cells in the SCG as well as some fibres of extrinsic origin. In this connection, NO may serve either as a neurotransmitter or neuromodulator. PMID- 11117632 TI - Anatomical variations in the pattern of the right hepatic veins: possibilities for type classification. AB - A morphological study of the right hepatic veins (RHVv) was conducted based on the shape and the confluence pattern of the superior right hepatic vein (SRHV) and the presence of accessory right hepatic veins. The study was performed in 110 undamaged, randomly selected, cadaveric human livers prepared using the corrosion cast methodology. The principles for classifying the RHVv into types were as follows: the length of the vein trunk, the confluence of 2 or 3 main tributaries that form a trunk, and the accessory right hepatic veins that modify the venous drainage of the right side of the liver. Four types of SRHV were identified. Type 1 (20 %), type 2 (40 %) and type 3 (25 %) were the most common, while type 4 (15 %) was linked to the accessory right hepatic veins in cases where they drain a surgically important part of the liver. Accessory right hepatic veins were found in a total of 31 casts (28 %). The hepatocaval confluence was studied and the tributary-free part of the SRHV trunk before it entered the inferior vena cava was measured. The tributary-free part of the SRHV was longer than 1 cm in 77 % of the casts. Anastomoses between the terminal tributaries of the veins involved in the drainage of the right side of the liver were also investigated. PMID- 11117631 TI - Exposure of rats to a high but not low dose of ethanol during early postnatal life increases the rate of loss of optic nerve axons and decreases the rate of myelination. AB - Visual system abnormalities are commonly encountered in the fetal alcohol syndrome although the level of exposure at which they become manifest is uncertain. In this study we have examined the effects of either low (ETLD) or high dose (ETHD) ethanol, given between postnatal days 4-9, on the axons of the rat optic nerve. Rats were exposed to ethanol vapour in a special chamber for a period of 3 h per day during the treatment period. The blood alcohol concentration in the ETLD animals averaged approximately 171 mg/dl and in the ETHD animals approximately 430 mg/dl at the end of the treatment on any given day. Groups of 10 and 30-d-old mother-reared control (MRC), separation control (SC), ETLD and ETHD rats were anaesthetised with an intraperitoneal injection of ketamine and xylazine, and killed by intracardiac perfusion with phosphate buffered glutaraldehyde. In the 10-d-old rat optic nerves there was a total of approximately 145,000-165,000 axons in MRC, SC and ETLD animals. About 4% of these fibres were myelinated. The differences between these groups were not statistically significant. However, the 10-d-old ETHD animals had only about 75,000 optic nerve axons (P < 0.05) of which about 2.8 % were myelinated. By 30 d of age there was a total of between 75,000-90,000 optic nerve axons, irrespective of the group examined. The proportion of axons which were myelinated at this age was still significantly lower (P < 0.001) in the ETHD animals (approximately 77 %) than in the other groups (about 98 %). It is concluded that the normal stages of development and maturation of the rat optic nerve axons, as assessed in this study, can be severely compromised by exposure to a relatively high (but not low) dose of ethanol between postnatal d 4 and 9. PMID- 11117633 TI - Mouse granulated metrial gland cells require contact with stromal cells to maintain viability. AB - Granulated metrial gland (GMG) cells differentiate in the uterine wall in pregnancy in mice but the mechanisms which control their differentiation and maintenance are unknown. In vivo, GMG cells share an intimate association with fibroblast-like stromal cells. The importance of this association has been assessed by examining the effects of withdrawal of stromal cell contact on GMG cell maintenance in vitro. When single cell suspensions of cells were prepared from mouse metrial glands there was a steady decline in numbers with days of culture but usually some remained at 7 d of culture. The ability of metrial gland cells to kill Wehi 164 target cells in 51Cr-release cytotoxicity assays was retained by cells cultured for at least 3 d. When explants of metrial gland were maintained in culture to allow GMG cell migration onto the culture flask, the attached GMG cells were lost by 1 d later. Overall, these results suggest that a juxtacrine regulatory mechanism maintains GMG cells. The rapid loss of unsupported GMG cells in culture has major implications in the design of assays to examine GMG cell function. PMID- 11117634 TI - Anatomy of the cystic artery arising from the gastroduodenal artery and its choledochal branch--a case report. AB - Variations in the branching pattern of the common hepatic artery often occur and may be encountered during cholecystectomy. Variants of the cystic artery, its branches and relations with the biliary structures and blood vessels emphasise the importance of arterial dissection in biliary surgery. In this study, a rare variant of the cystic artery and its choledochal branch is described. The cystic artery arose from the gastroduodenal artery, passed anterior to structures in the free margin of lesser omentum and travelled a long distance before supplying the gall bladder. A long choledochal branch was noted accompanying the common bile duct. Surgical implications of this variation of the cystic and choledochal arteries are discussed. PMID- 11117635 TI - Variations in the normal anatomy of the collateral ligaments of the human elbow joint. AB - The variations which occur in the medial and lateral ligament complexes of the elbow were investigated. These occurred frequently with the standard appearances occurring in no more than half the specimens on the medial side and one quarter of those on the lateral side. Surgeons who regularly perform elbow arthroplasty must be aware of these considerations, especially with the introduction of unconstrained prostheses which rely upon the ligament complex for their postoperative stability. PMID- 11117636 TI - Prognostic factors for pneumococcal bacteremia in a university hospital. AB - The records of adult patients with pneumococcal bacteremia who were seen over an 8-year-period at an 1,100-bed university teaching hospital were reviewed in order to revise the clinical and laboratory findings and to identify the risk factors associated with mortality. A total of 156 patients were studied, 101 men and 55 women. The mean age of the patients was 65 years. Eighty-seven percent of the patients had community-acquired bacteremia and 13% had nosocomial pneumococcal bacteremia. The overall mortality was 33.9% and the related mortality was 20.5%. The following factors were associated with an increased risk of adverse outcome in the univariate analysis: mechanical ventilation (risk ratio [RR]=3.40; 95% confidence interval [95% CI]=1.44-8.05), administration of parenteral nutrition (RR=3.40; 95% CI =1.44-8.05), and the presence of an intravenous catheter (RR=2.33; 95% CI=1.27-4.24). In the multivariate analysis, the independent prognostic factors for mortality were as follows: development of clinical complications during the episode of bacteremia, rapidly fatal illness, advanced age and administration of parenteral nutrition. The results suggest that the overall mortality due to pneumococcal bacteremia continues to be high. Four independent risk factors associated with increased mortality were identified. Prevention and immunization with polyvalent pneumococcal polysaccharide vaccine should be practiced more widely. PMID- 11117637 TI - Epidemiological, microbiological, clinical, and prognostic factors of bacteremia caused by high-level vancomycin-resistant Enterococcus species. AB - A case-control study was performed between 1994 and 1996 in order to study the epidemiological, microbiological, clinical, and prognostic features of high-level vancomycin-resistant enterococcal bacteremia. Seventeen consecutive patients who had clinically significant bacteremia due to vancomycin-resistant enterococci (vanA genotype: 16 Enterococcus faecalis, 1 Enterococcus faecium) were compared with 169 who had vancomycin-susceptible enterococcal bacteremia. The following were selected by multivariate analysis as independent risk factors that influenced the development of high-level vancomycin-resistant enterococcal bacteremia: prior glycopeptide therapy (P=0.049); inclusion in a hemodialysis program (P=0.046); prior therapy with corticosteroids or antineoplastic agents (P=0.029); and prior surgical treatment (P=0.022). The following other factors were selected by univariate analysis: tracheostomy (P=0.002); prolonged hospitalization (P=0.01); and any kind of puncture (P=0.02). The crude associated mortality rate was 13.4%. Gene amplification of vanA was positive for 17 strains of enterococci. Pulsed-field gel electrophoresis of genomic DNA after SmaI digestion of vanA isolates revealed that one strain predominated (10 isolates), though at least four similar banding patterns were identified (6 isolates). The 16 strains closely related to the outbreak were investigated further. The surgical intensive care unit was the first and most involved service. The hospital outbreak of vanA vancomycin-resistant enterococcal bacteremia occurred between 1994 and 1995 and was caused by Enterococcus faecalis. This is believed to be the first and only such outbreak described in a Spanish hospital thus far. PMID- 11117638 TI - Comparative study of three culture systems for optimal recovery of mycobacteria from different clinical specimens. AB - A new broth-based nonradioactive culture system, MB-Redox (Heipha Diagnostika Biotest, Germany), was compared with the liquid radiometric Bactec 460 TB system and the solid Lowenstein-Jensen (L-J) medium for recovery rate and time to detection of mycobacteria. Of the 605 clinical specimens studied, 100 grew acid fast bacilli (AFB). The isolation rate for all AFB was 84% for Bactec, 69% for MB Redox, and 48% for L-J. Eighteen percent of the 100 isolates grew only in Bactec, 10% only in MB-Redox, and 5% only in L-J. The average times to detection of the 100 isolates were 13.2, 17.8, and 28.3 days, respectively, and the mean growth times for the 34 AFB detected by all three media were 13.7, 15.2, and 26.8 days, respectively. Mycobacterium tuberculosis was isolated from 15 clinical samples, with detection times of 16.8 days for Bactec, 18.3 for MB-Redox, and 22.8 days for L-J medium; 10 of these isolates grew in all media, with detection times of 14.2, 16.4, and 18.7 days, respectively. Seven were also positive on direct smear, with detection times of 12.4, 13.7, and 17.7 days. Two of the Mycobacterium tuberculosis isolates were recovered only in Bactec and another one only in L-J medium. Mycobacterium haemophilum grew only in the liquid systems, which provided the special growth factors this bacterium requires. Only the combination of all three systems yielded optimal recovery. MB-Redox gave reliable results, offering the advantages of ready-to-use tubes in which the antibiotic supplement is already incorporated and easy and immediate reading of the results. Since this system does not contain any radioactive substance, results can be confirmed with acid-fast staining and conventional and molecular tests. PMID- 11117639 TI - Macrolide resistance phenotypes and genotypes in French clinical isolates of Streptococcus pneumoniae. Observatoire de Normandie du Pneumocoque. AB - The aim of this study was to analyze the mechanisms of macrolide resistance in French clinical isolates of Streptococcus pneumoniae. A total of 838 strains of pneumococci were isolated in 1997 in Normandy, a region of western France, by 19 microbiology laboratories. Fifty-three percent had displayed diminished susceptibility to penicillin G and 50% were resistant to erythromycin. From this collection, 92 penicillin-intermediate or -resistant and 18 penicillin susceptible strains resistant to erythromycin were studied. The presence of erm genes coding for ribosomal methylases and of mefE-like genes responsible for macrolide efflux was screened by a multiplex polymerase chain reaction and confirmed by DNA/DNA hybridization. Of the 110 strains studied, 108 were cross resistant to erythromycin, spiramycin and clindamycin, including 105 strains containing ermB-related genes and three strains that contained a combination of ermB- and mefE-related genes. Two strains apparently susceptible to clindamycin but resistant to spiramycin also contained ermB-related genes. No strain was resistant to erythromycin alone or contained only a mef-like gene. Therefore, resistance to erythromycin is mostly related to ribosomal methylation in this region of France. PMID- 11117640 TI - Genetic diversity among strains of Moraxella catarrhalis cultured from the nasopharynx of young and healthy Brazilian, Angolan and Dutch children. AB - The present study describes the carriage patterns and genetic variability of Moraxella catarrhalis strains isolated from children living in different countries. Moraxella catarrhalis is genetically heterogeneous, but little is known about its geographic distribution and phenotypic and genetic diversity in warm-climate countries. A collection of 99 isolates from 30 Brazilian, 19 Angolan and 50 Dutch healthy children, all less than 5 years of age, was investigated for phenotypic and genotypic relatedness. The isolates from the three countries were similar where biochemical reactivity was concerned: 89 strains were beta lactamase-producing and 87 were complement-resistant as determined by phenotype. There was no geographical difference in the prevalence of beta-lactamase producing isolates, but the carriage rate of complement-resistant strains was significantly higher in Dutch than in Angolan children (P=0.004). Complement resistance of 66 randomly selected strains was genetically confirmed in a Southern hybridization assay by a novel DNA probe that is specific for complement resistant strains and that demonstrated a sensitivity of 97% and a specificity of 100%. PCR amplification based on the probe sequence had a sensitivity of 98% and a specificity of 57% when compared to the outcome of a conventional culture spot test. PCR restriction fragment length polymorphism analysis of the MU 46 locus and pulsed-field gel electrophoresis of SpeI DNA macrorestriction fragments revealed genetic heterogeneity of strains from within and between the three countries, and no geographical clustering could be established. In conclusion, similar phenotypic characteristics but genotypic heterogeneity was found among Moraxella catarrhalis strains colonizing children in three different continents. PMID- 11117641 TI - Extended-spectrum TEM- and SHV-type beta-lactamase-producing Klebsiella pneumoniae strains causing outbreaks in intensive care units in Italy. AB - The aim of the present study was to investigate the production of extended spectrum beta-lactamases (ESbetaLs) and the epidemiological correlations in a total of 107 Klebsiella pneumoniae strains resistant to third- and fourth generation cephalosporins. The strains were collected from patients in four intensive care units (3 neonatal and 1 general) in three hospitals in Italy between March 1996 and July 1997. All strains were found to produce ESbetaLs. Phenotypic (antibiotyping and ESbetaL patterns) and genotypic (plasmid profile and pulsed-field gel electrophoresis) analyses showed that a single strain had been responsible for each outbreak in each of the four intensive care units. Isoelectric focusing, activity on substrates and gene sequencing showed that the strains produced SHV-5, SHV-2a, SHV-12 and TEM-52 beta-lactamases. This is not only the first time that ESbetaL-producing Klebsiella pneumoniae strains have been reported as causing epidemics in Italian hospitals, it is also, to the best of our knowledge, the first time that an outbreak caused by a TEM-52 ESbetaL producing Klebsiella pneumoniae strain has been reported. The data presented here illustrate the complexity of determining the epidemiological pattern of ESbetaL producers in large hospitals that do not have an ESbetaL-monitoring program. PMID- 11117642 TI - Assessment of intercentre reproducibility and epidemiological concordance of Legionella pneumophila serogroup 1 genotyping by amplified fragment length polymorphism analysis. AB - The aims of this work were to assess (i) the intercentre reproducibility and epidemiological concordance of amplified fragment length polymorphism analysis for epidemiological typing of Legionella pneumophila serogroup 1, and (ii) the suitability of the method for standardisation and implementation by members of the European Working Group on Legionella Infections. Fifty coded isolates comprising two panels of well-characterised strains, a "reproducibility" panel (n=20) and an "epidemiologically related" panel (n=30), were sent to 13 centres in 12 European countries. Analysis was undertaken in each centre following a previously determined standard protocol. Results were analysed by the participants, using gel analysis software where available, and submitted to the coordinating centre. The coordinating centre reanalysed all results visually and selected data-sets with gel analysis software. Data analysis by participants yielded reproducibility (R) values of 0.20-1.00 and epidemiological concordance (E) values of 0.11-1.00, with 6 to 34 types. Following visual analysis by the coordinating centre, R=0.78-1.00, and E=0.67-1.00, with 10-20 types. Analysis of three data-sets by the coordinating centre using gel analysis software yielded R=1.00 and E=1.00, with 12, 13 or 14 types. This method can be used as a simple, rapid screening tool for epidemiological typing of isolates of Legionella pneumophila serogroup 1. Results demonstrate that the method can be highly reproducible (R=1.00) and epidemiologically concordant (E=1.00), with good discrimination. The electropherograms generated are amenable to computer-aided analysis, but strict adherence to a previously defined laboratory protocol is required. Following designation of representative type strains and patterns, this method will be adopted by the European Working Group on Legionella Infections as the first internationally standardised typing method for use in the investigation of travel-associated Legionella infections. PMID- 11117643 TI - Osteomyelitis of the hip joint associated with systemic cat-scratch disease in an adult. AB - Reported here is the case of a 29-year-old male with cervical lymphadenopathy, fever and weight loss, followed by acute painful osteomyelitis of the left hip joint due to cat-scratch disease. The diagnosis was established by detection of IgG antibodies to Bartonella henselae in serum and histologic examination of a lymph node including a positive polymerase chain reaction test. Treatment consisted of clarithromycin and cefotiam for 2 weeks. Four weeks after discharge, all of the patient's symptoms had completely resolved. Magnetic resonance imaging of the left hip joint showed marked regression of bone inflammation 4 months later and normalization after 8 months. Cat-scratch disease should be considered in the differential diagnosis of osteomyelitis in an adult, especially when lymphadenitis is present. PMID- 11117644 TI - Changing epidemiology of dengue fever in travelers to Thailand. AB - A study was conducted to evaluate the incidence and seasonal pattern of dengue fever occurring among Israeli travelers to Southeast Asia. The illness was diagnosed by Israeli physicians stationed abroad and by serological methods carried out in Israel. Between 1994 and 1998 dengue fever was confirmed in 103 travelers, 80 of whom were diagnosed in Israel. A sharp increase in the incidence was noted in 1998 as compared with the previous 4 years. The attack rate during 1998 in a defined group of travelers was 3.4/1,000 and reached a peak of 5/1,000 during the dry season of 1998. PMID- 11117645 TI - Evaluation of a Helicobacter pylori stool antigen test for the diagnosis and follow-up of infections in children. AB - The aim of this study was to evaluate the performance of a newly developed enzyme immunoassay kit (HpSA) for detecting Helicobacter pylori antigens in the stool of children. This study was comprised of 58 children referred to various endoscopy units for evaluation of gastrointestinal symptoms and upper gastroduodenal endoscopy and 11 children for post-therapy follow-up. In the first group, 23 children were diagnosed as positive for Helicobacter pylori using bacteriological and/or histological methods. Stool antigens were detected in 20 of these positive patients, for a sensitivity of 86.9% and a negative predictive value of 91.9%. Since only one false-positive reaction was observed with the HpSA kit, the specificity was 97.1% and the positive predictive value 95.2%. Results obtained for post-therapy follow-up were also promising. The HpSA assays were negative for the eight children whose infections were eradicated after therapy, and a positive result was obtained for two of three patients who had a persistent infection. PMID- 11117646 TI - cagA, vacA and iceA virulence genes of Helicobacter pylori isolates of children in Finland. AB - cagA, vacA s and m genotypes and iceA alleles were analyzed from Helicobacter pylori strains isolated from 17 Finnish children and 32 children of non-Finnish origin living in Finland. Twelve children in the latter group were eastern European and 15 were of African origin. Only three children of non-Finnish origin were born in Finland. The vacA sla subtype was more prevalent in the isolates from Finnish children than African children (76% vs. 7%, P<0.001); vacA s1b frequencies were 5% and 67%, respectively (P<0.001). The iceA1 allele was significantly more prevalent in African than Finnish isolates (93% vs. 35%, P< 0.01). Considerable variation was noted in the frequency of vacA s1 subtypes and iceA alleles in children originating from different geographic regions, but the geographic variation of s1 subtypes resembled that described in other reports. PMID- 11117647 TI - Evaluation of a new commercial immunoassay for rapid detection of Campylobacter jejuni in stool samples. AB - In order to evaluate a new commercial enzyme immunoassay (ProspecT Campylobacter Microplate Assay; Alexon-Trend, USA) for the detection of Campylobacter jejuni and Campylobacter coli in stool samples, 30 faecal specimens known to be culture positive for Campylobacter jejuni were tested with the new assay. The detection limit was approximately 3 x 10(6)/ml in faecal suspensions. The sensitivity relative to culture was 80% (24/30). All of the 24 positive samples, except for one, remained positive after being stored at -20 degrees C for 60 days. The specificity of the test was 100%. Interestingly, 6 of 11 additional Campylobacter jejuni culture-positive samples that had been obtained from patients with Guillain-Barre syndrome and stored at -20 degrees C for periods of up to 5 years tested positive in the assay. The performance of the assay indicates that it has potential value for use in future early intervention studies. PMID- 11117648 TI - Prospective evaluation and follow-up of European patients with visceral leishmaniasis and HIV-1 coinfection in the era of highly active antiretroviral therapy. AB - A prospective descriptive study was designed to determine the impact of highly active antiretroviral therapy (HAART) in the evolution of visceral leishmaniasis (VL) in HIV-1 infected patients. Thirty-two patients were treated with meglumine antimoniate or amphotericin B in lipid formulations. Patients who had undergone previous HAART at study entry (n=17) continued with therapy while receiving treatment for VL. Patients who had never undergone HAART started it after VL treatment finished (n=15). Ten patients were lost to follow-up. All of the remaining patients (n=20) continued to receive HAART and were followed for an average of 441 days. Relapses were observed in 5 of 20 patients. These results indicate that HAART neither prevents the incidence of VL relapse nor modifies the clinical picture described in the pre-HAART era. PMID- 11117649 TI - Hantavirus infections among mammalogists studied by focus reduction neutralisation test. PMID- 11117650 TI - Value of terminal subculture of automated blood cultures in patients with candidaemia. PMID- 11117651 TI - Clinical comparison of immunoblot and antibody index for detection of intrathecal synthesis of specific antibodies in Lyme neuroborreliosis. PMID- 11117652 TI - Zaleplon: a pyrazolopyrimidine sedative-hypnotic agent for the treatment of insomnia. AB - BACKGROUND: Insomnia is the subjective complaint of poor sleep or an inadequate amount of sleep that adversely affects daily functioning. For the past 4 decades, treatment of insomnia has shifted away from the use of barbiturates toward the use of hypnotic agents of the benzodiazepine class. However, problems associated with the latter (eg, next-day sedation, rebound insomnia, dependence, and tolerance) have prompted development of other agents. OBJECTIVE: This review describes the recently approved nonbenzodiazepine agent, zaleplon. METHODS: Studies of zaleplon were identified through a search of English-language articles listed in MEDLINE and International Pharmaceutical Abstracts, with no limitation on year. These were supplemented by educational materials from conferences. RESULTS: The efficacy and tolerability of zaleplon have been documented in the literature. Zaleplon has been shown to improve sleep variables in comparison with placebo. Like most hypnotic agents, zaleplon can be used for problems of sleep initiation at the beginning of the night, but its short duration of clinical effect may also allow patients to take it later in the night without residual effects the next morning. Zaleplon can be taken < or = 2 hours before awakening without "hangover" effects. It is generally well tolerated, with headache being the most commonly reported adverse event in clinical trials (15%-18%). Compared with flurazepam, a long-acting benzodiazepine sedative-hypnotic agent, zaleplon causes significantly less psychomotor and cognitive impairment (P < 0.001). Zaleplon has not been studied in pregnant women or children. The dose of zaleplon should be individualized; the recommended daily dose for most adults is 10 mg. CONCLUSIONS: Insomnia has a substantial impact on daily functioning. If pharmacologic treatment is indicated for insomnia, the choice of an agent should be guided by individual patient characteristics. PMID- 11117653 TI - Pantoprazole: a new proton pump inhibitor. AB - OBJECTIVE: This paper reviews the pharmacology, clinical efficacy, and tolerability of pantoprazole in comparison with those of other available proton pump inhibitors (PPIs). METHODS: Relevant English-language research and review articles were identified by database searches of MEDLINE, International Pharmaceutical Abstracts, and UnCover, and by examining the reference lists of the articles so identified. In selecting data for inclusion, the author gave preference to full-length articles published in peer-reviewed journals. RESULTS: Like other PPIs, pantoprazole exerts its pharmacodynamic actions by binding to the proton pump (H+,K+ -adenosine triphosphatase) in the parietal cells, but, compared with other PPIs, its binding may be more specific for the proton pump. Pantoprazole is well absorbed when administered as an enteric-coated, delayed release tablet, with an oral bioavailability of approximately 77%. It is hepatically metabolized via cytochrome P2C19 to hydroxypantoprazole, an inactive metabolite that subsequently undergoes sulfate conjugation. The elimination half life ranges from 0.9 to 1.9 hours and is independent of dose. Pantoprazole has similar efficacy to other PPIs in the healing of gastric and duodenal ulcers, as well as erosive esophagitis, and as part of triple-drug regimens for the eradication of Helicobacter pylori from the gastric mucosa. It is well tolerated, with the most common adverse effects being headache, diarrhea, flatulence, and abdominal pain. In clinical studies, it has been shown to have no interactions with various other agents, including carbamazepine, cisapride, cyclosporine, digoxin, phenytoin, theophylline, and warfarin. CONCLUSIONS: Pantoprazole appears to be as effective as other PPIs. Its low potential for drug interactions may give it an advantage in patients taking other drugs. PMID- 11117654 TI - Effect of zanamivir on duration and resolution of influenza symptoms. AB - BACKGROUND: Zanamivir is a neuraminidase inhibitor, the first of a new class of drugs with potent, specific antiviral activity against influenza A and B. Administration by inhalation results in direct delivery to the respiratory tract, the principal site of viral replication. OBJECTIVE: This study was undertaken to determine the effectiveness of zanamivir on duration and resolution of influenza symptoms. METHODS: Using a method similar to that employed in amantadine treatment studies to obtain supporting evidence of efficacy for US Food and Drug Administration consideration, pooled data from 6 zanamivir phase II and III clinical trials involving primarily previously healthy adults were analyzed to categorize patients as accelerated resolvers (temperature <37.8 degrees C < or = 24 hours after dosing, plus a > or = 50% reduction in symptom score by 36 hours), early resolvers (temperature <37.8 degrees C < or = 36 hours after dosing), and nonaccelerated resolvers (febrile >36 hours after dosing). Patients in the accelerated and early categories were termed rapid resolvers; the others were slow resolvers. Patients recorded their symptom severity, temperature, ability to perform normal daily activities, and use of relief medication on a diary card. The primary end point of median time to alleviation of symptoms was also reexamined, with the additional requirement that patients were not taking relief medication when their symptoms were alleviated. This analysis was intended to control for the possible effect of relief medication on the primary end point. RESULTS: In the influenza-positive population (n = 1572), significantly more zanamivir-treated patients were rapid resolvers compared with those receiving placebo (807 [72%] vs 765 [64%], P < 0.001). Significant benefits of zanamivir treatment were observed in patients with a baseline temperature of > or = 37.8 degrees C (630 [68%] vs 595 [57%], P < 0.001) or > or = 38.3 degrees C (382 [67%] vs 365 [52%], P < 0.001) and in patients considered by their physi- cian to have severe symptoms at the start of therapy (252 [70%] vs 222 [63%], P = 0.02). Differences were even more apparent in patients with high-risk conditions (74% vs 53%, P = 0.014) and in those aged > or = 50 years (70% vs 54%, P = 0.005). Zanamivir treatment was also associated with a significant reduction in time to alleviation of symptoms with no use of relief medication. This was particularly noticeable in those aged > or = 50 years; time to alleviation was 13 days in patients receiving placebo and 6 days in patients receiving zanamivir (P < 0.001). In these trials, adverse events were reported at a similar frequency in patients receiving zanamivir and those receiving placebo. CONCLUSIONS: Zanamivir is more effective than placebo in patients with influenza at providing early symptom relief and a reduced duration of illness at a time when use of relief medication has ended. These benefits are seen across different patient groups but appear to be particularly marked in patients who are aged > or = 50 years, who have underlying illnesses, who are considered high risk, or who are more severely ill at the beginning of therapy. PMID- 11117655 TI - Ibuprofen liquigel for oral surgery pain. AB - BACKGROUND: Ibuprofen liquigel is a solubilized potassium ibuprofen 200-mg gelatin capsule formulation that was approved for over-the-counter use in 1995. OBJECTIVE: This study compared the analgesic efficacy and tolerability of ibuprofen liquigel 200 mg, ibuprofen liquigel 400 mg, acetaminophen caplets 1000 mg, and placebo in patients experiencing moderate or severe pain after surgical removal of impacted third molars. METHODS: This randomized, double-blind, parallel-group, 6-hour study was conducted in 210 patients experiencing moderate or severe postoperative pain. Ratings of pain intensity and pain relief were recorded every 15 minutes for the first hour, at 90 and 120 minutes, and then hourly through hour 6. The onsets of first perceptible relief and meaningful relief were recorded using 2 stopwatches. An analysis of variance model was employed to test for significant differences (P < or = 0.05) between treatment groups with respect to pain relief, pain intensity difference, total pain relief (TOTPAR), and summed pain intensity difference (SPID). Stopwatch measures were analyzed using the Cox proportional hazards model. Drug tolerability was assessed by monitoring the occurrence of adverse events. RESULTS: During the first 2 hours of the study (TOTPAR 2 and SPID 2), all active treatments were significantly more efficacious than placebo (P < 0.001), with ibuprofen liquigel 200 and 400 mg significantly more efficacious than acetaminophen 1000 mg (P < 0.05 and P < 0.01, respectively). For the entire duration of the study (TOTPAR 6 and SPID 6), only the 2 doses of ibuprofen liquigel were significantly more efficacious than placebo (P < 0.001). Ibuprofen liquigel 200 and 400 mg were also significantly more efficacious than acetaminophen 1000 mg on the summary measures TOTPAR 6 and SPID 6 (P < 0.01 and P < 0.001, respectively). Analysis of the stopwatch data revealed that all active treatments displayed significantly more rapid onsets to confirmed first perceptible relief (P < 0.001 to < 0.05) and meaningful relief (P < 0.001 to < 0.01) than did placebo, with ibuprofen liquigel 400 mg displaying a significantly more rapid onset to meaningful relief than acetaminophen 1000 mg (P < 0.05) and a significantly more rapid onset to confirmed first perceptible relief than acetaminophen 1000 mg (P < 0.001) and ibuprofen liquigel 200 mg (P < 0.01). All adverse events were considered mild or moderate, with an overall incidence of 11.5% in the ibuprofen liquigel 200-mg group, 6.8% in the ibuprofen liquigel 400-mg group, 19.0% in the acetaminophen 1000-mg group, and 25.9% in the placebo group. CONCLUSIONS: Ibuprofen liquigel provided greater peak and overall analgesic effects and a more rapid onset to analgesia than did acetaminophen 1000 mg. PMID- 11117656 TI - Adverse events and treatment discontinuations in clinical trials of fluoxetine in major depressive disorder: an updated meta-analysis. AB - BACKGROUND: A 1993 meta-analysis of US Investigational New Drug clinical trials of fluoxetine reinforced this agent's more favorable adverse-event profile compared with tricyclic antidepressants (TCAs). OBJECTIVES: The present meta analysis sought to provide a reanalysis of updated adverse-event and discontinuation data for fluoxetine 20 to 80 mg/d compared with TCAs and placebo in the treatment of major depressive disorder (MDD) in adults. A subanalysis to assess the safety profile of the most commonly used effective dose of fluoxetine in MDD (20 mg) was also conducted. METHODS: Data were obtained from 25 double blind clinical trials involving 4016 patients with MDD randomized to treatment with fluoxetine 20 to 80 mg/d, TCAs, or placebo. The subanalysis included data from 6 trials involving 1258 patients treated with fixed 20-mg doses of fluoxetine or placebo. Spontaneously reported treatment-emergent adverse events, reasons for discontinuation, and events leading to discontinuation were compared between groups. RESULTS: The age of the 4016 randomized patients ranged from 12 to 90 years, with a mean age of 46 years. Most patients were white (92%), and 62% were female. The age of the 1258 patients in the 20-mg fixed-dose population ranged from 18 to 90 years, with a mean age of 54 years; as in the total population, most of these patients were white (92%), and 57% were female. The adverse-event profiles of fluoxetine and TCAs in these trials were consistent with the typical profiles of selective serotonin reuptake inhibitors and TCAs. At a dose of 20 mg/d, fluoxetine-treated patients had a discontinuation rate due to adverse events that was not statistically significantly different from that in placebo recipients. Discontinuation rates due to lack of efficacy were not significantly different between fluoxetine and TCAs. However, significantly more TCA-treated patients discontinued therapy because of adverse events and significantly fewer completed treatment compared with fluoxetine-treated patients (both, P < 0.001). The most common events (> or = 2%) leading to discontinuation were asthenia, dizziness, insomnia, nausea, nervousness, somnolence, and tremor in fluoxetine-treated patients and abnormal vision, agitation, constipation, dizziness, dry mouth, headache, nausea, nervousness, rash, somnolence, sweating, and tremor in TCA-treated patients. CONCLUSIONS: Data from this large series of clinical trials confirm that fluoxetine is well tolerated in the acute treatment of MDD in adults, especially at a dosage of 20 mg/d, and is better tolerated than the recommended doses of TCAs. PMID- 11117657 TI - Reimportation of prescription drugs: a rush to judgment? PMID- 11117658 TI - Indirect cost of HIV infection in England. AB - BACKGROUND: Few studies have estimated the indirect costs of care for HIV infection in England by stage of infection at a population level. OBJECTIVE: This study estimated annual indirect costs of the HIV epidemic in England in 1997-1998 from both a public-sector and societal perspective. METHODS: Service costs for HIV-infected individuals were indexed to 1997-1998 English prices. Average annual indirect costs included the costs of statutory, community, and informal services; disability payments; and lost economic productivity by stage of HIV infection. Disability payments were excluded from the societal perspective, whereas the degree of lost economic productivity was varied for the sensitivity analyses. Total average annual indirect costs by stage of HIV infection were calculated, as were population-based costs by stage of HIV infection and overall population costs. RESULTS: Annual indirect costs from the public-sector and societal perspectives, respectively, ranged from pound sterling 3169 (dollars 5252) to pound sterling 3931 (dollars 6515) per person-year for asymptomatic individuals, pound sterling 5302 (dollars 8787) to pound sterling 7929 (dollars 13,140) for patients with symptomatic non-AIDS, and pound sterling 9956 (dollars 16,499) to pound sterling 21,014 (dollars 34,825) for patients with AIDS. Estimated population-based indirect costs from the public-sector perspective varied between pound sterling 109 million (dollars 181 million) and pound sterling 145 million (dollars 241 million) for 1997-1998, respectively, comprising between 58% and 124% of direct treatment costs for triple drug therapy in England during 1997. From the societal perspective, estimated population-based costs varied between pound sterling 84 million (dollars 138 million) and pound sterling 119 million (dollars 198 million) in 1997-1998, comprising between 45% and 102% of direct treatment costs and cost of care, respectively, during 1997. CONCLUSIONS: Average indirect costs increase as HIV-infected individuals' illness progresses. Whether one takes a public-sector or societal perspective, indirect costs add a considerable amount to the cost of delivering health care to HIV-infected individuals. Both direct and indirect costs, when obtainable, should be used to assess the economic consequences of HIV infection and treatment interventions. PMID- 11117659 TI - Health care utilization by migraine patients: a 1998 Medicaid population study. AB - BACKGROUND: In the last decade, a number of studies have documented the economic impact of migraine headaches on society. Although previous research has shown that patients with migraine headache consume a greater amount of health care resources than those without migraine, the economic impact of this condition on a Medicaid population has not been assessed. OBJECTIVE: The purpose of this study was to compare the health care resource utilization of-patients with and without migraine headache in the Idaho Medicaid population. METHODS: Idaho Medicaid claims from 1998 were reviewed to identify cases and controls. Four controls, matched for age, sex, race, and residence, were obtained for each case. Physician services, hospital services, emergency room services, and prescription use were compared between the 2 groups. Multivariate analyses were performed to determine differences between the 2 groups after controlling for potential confounders. RESULTS: Eighty percent of the cases were female, and 94% of the patients were white. Patients with migraine headache had statistically significantly higher health care resource consumption than matched controls (P < 0.05). Total log costs for prescription use, physician services, and hospital services were significantly higher (P < 0.001) in the migraine group even after controlling for migraine-associated comorbid conditions and demographic variables. CONCLUSIONS: Total health care costs for migraine patients were 1.6 times higher than for matched controls. The results of this study suggest that migraine is a significant economic burden to the Medicaid program. PMID- 11117660 TI - Palivizumab for respiratory syncytial virus prophylaxis in high-risk infants: a cost-effectiveness analysis. AB - BACKGROUND: Prophylactic therapy with palivizumab, a humanized monoclonal antibody, has been shown to reduce the number of respiratory syncytial virus (RSV)-related hospitalizations in preterm infants. The cost-effectiveness of this therapy has not been evaluated from the provider's perspective using cost data. OBJECTIVES: The objectives of this study were to determine the cost per RSV infection episode avoided by using prophylactic palivizumab therapy in a high risk infant population and to determine whether certain subgroups of infants derived greater benefit from prophylactic therapy. METHODS: A decision-analytic model simulating an RSV infection episode was developed to evaluate the cost effectiveness of palivizumab prophylaxis from the perspective of the health care system (provider). Data to populate the model were gathered from the medical literature (identified through a MEDLINE search of studies on the incidence of RSV infection) and the IMpact-RSV clinical trial. Data included incidence of RSV infection and the associated health care resource use and costs. Costs to the provider were determined using a university-affiliated hospital cost-accounting system. Cost-effectiveness ratios were calculated over a range of RSV infection incidence rates in a control population. Sensitivity analyses were performed for the cost of palivizumab therapy, the cost of RSV-related hospitalization, and the number of emergency department, physician office, and home health care visits. For the subgroup analysis, infants were classified by gestational age (<32 and > or = 32 weeks) and stratified by severity of chronic lung disease. RESULTS: The cost per additional RSV infection episode avoided ranged from dollars 0 (cost savings) to dollars 39,591 for palivizumab prophylaxis costs of dollars 2500 and from dollars 2702 to dollars 79,706 for palivizumab prophylaxis costs of dollars 4500. The model was insensitive to changes in the number of emergency department, physician office, and home health care visits. The difference in RSV incidence between the treatment and control groups was greater among infants > or = 32 weeks' gestational age than among infants <32 weeks' gestational age. onclusions: The incremental cost-effectiveness of palivizumab compared with no prophylactic therapy was sensitive to changes in the incidence of RSV infection in control infants, the average cost of RSV hospitalization, and the cost of palivizumab. Clinicians may use this information along with additional factors to determine whether palivizumab is cost-effective in their clinical setting and geographic area. PMID- 11117661 TI - Efficacy of omeprazole plus two antimicrobials for the eradication of Helicobacter pylori in a Turkish population. PMID- 11117662 TI - Doses of olanzapine, risperidone, and haloperidol used in clinical practice: results of a prospective pharmacoepidemiologic study. PMID- 11117663 TI - The natural history of premature adrenarche. AB - The growth and pubertal development of patients with premature adrenarche are reported to be normal, but the effects of this condition on pubertal growth are not well documented. In the present study the growth kinetics of a group of 38 female patients with isolated premature adrenarche followed in our institution for a period of 5.77 (SD=1.7) yr were evaluated to assess whether and how premature adrenarche affects pubertal growth and final height. Birth weight and length, height, skeletal maturation, onset of puberty, age at menarche, height prognosis and final height were documented. To examine the shape of growth kinetics, growth profiles of each girl were fitted with Preece-Baines nonlinear function (PB1) and mean constant curves were obtained by a 2-stage linear model. The biokinetic constants of the patients were compared both with those obtained by Preece and Baines in normal British girls participating in the Harpenden study and by Milani et al. in normal Italian girls. Birth weight and length of premature adrenarche patients were within the normal range for a northern Italian population [+0.29 (1.57) and -0.40 (1.49) SDS, respectively]. Analysis of the biokinetic constants showed that in PA girls the prepubertal component of height velocity, ie. the one preceding the diagnosis of PA, was consistently higher than that of both normal Italian and Harpenden girls, accounting for the increased height of the patients at the beginning of puberty. In contrast, the pubertal component of height velocity was reduced with respect to control groups, leading to a final height similar to the one estimated by the PB1 model and to the target height. In conclusion, the transient acceleration of growth and bone maturation observed in girls with premature adrenarche did not negatively influence the onset and progression of puberty but modified the growth pattern of these patients. Prepubertal growth was enhanced with respect to normal controls, and this enhancement was compensated for by a reduction of the pubertal growth component leading to a final height in accordance with the target height. This abnormal growth shape is not due to an altered tempo of growth but it is rather a consequence of premature adrenarche. PMID- 11117664 TI - Adrenarche and nutritional status. AB - The prepubertal fat spurt seen in mid-childhood coincides with the beginning of adrenarche and is associated with rising serum levels of insulin and insulin-like growth factor-I. As the adrenal cortex expresses receptors for these anabolic peptides, implying that the nutritional status is communicated to the adrenal gland, we hypothesized that nutritional status may be causally involved in the regulation of adrenal androgen secretion. To test this hypothesis, anthropometric indices of the nutritional status and 24-h urinary excretion rates of dehydroepiandrosterone sulfate (DHEAS) were studied longitudinally (during observation periods of at least 4 years) in healthy normal-weight prepubertal and pubertal children. Increases in urinary DHEAS excretion proved to be significantly elevated during periods of individual highest rises in body mass index. These findings provide the first in vivo evidence that a change in nutritional status is an important physiological regulator of adrenarche. PMID- 11117665 TI - Syndrome X: a consequence of intra-uterine malnutrition? AB - The long-term consequences of intra-uterine growth retardation (IUGR) on postnatal growth and development are well known. More recently, several epidemiological and clinical studies in adults have provided evidence that low birth weight is an independent risk factor for the syndrome of insulin resistance in mid to late adulthood. In the present study, we demonstrated that individuals born small for gestational age showed isolated insulin resistance in early adulthood with no impairment of serum lipid concentrations and blood pressure at this age. Angiotensin-I converting enzyme gene polymorphism has also been shown to modulate the effect on insulin resistance of being small for gestational age. Fetal malnutrition may be the critical factor to explain the relationship between IUGR and insulin resistance which might be modulated by genetic factors. PMID- 11117666 TI - Precocious pubarche in girls and the development of androgen excess. AB - The prevalence of ovarian hyperandrogenism, hyperinsulinism and dyslipidemia is increased among adolescent girls with a history of premature pubarche (defined as the appearance of pubic hair before the age of 8 yr). The ovarian hyperandrogenism is characterized by clinical signs of androgen excess and by an exaggerated ovarian 17-hydroxyprogesterone response to GnRH agonist stimulation. The hyperinsulinism and dyslipidemia are detectable before and during pubertal development, and are commonly accompanied by low serum levels of insulin-like growth factor binding-protein 1 (IGFBP-1) and sex hormone-binding globulin (SHBG), and by an increased prevalence of anovulation from late adolescence onwards, even in the absence of clinical signs of androgen excess. In girls, premature pubarche, hyperinsulinism, low IGFBP-1, dyslipidemia, anovulation and hyperandrogenism--and some combinations of these--have been related to reduced fetal growth, indicating that these constellations or sequences may have a prenatal origin. Together, these findings suggest that premature pubarche in girls should no longer be merely regarded as a normal variant of development, but rather as a childhood marker pointing to an increased risk for a polyendocrine metabolic disorder of prenatal origin. PMID- 11117667 TI - Pre- and postpuberal findings in premature adrenarche. AB - The complications of hyperinsulinism and insulin resistance are becoming more common in pediatrics (including type 2 diabetes mellitus, dyslipidemia and polycystic ovary syndrome) because of the increased occurrence of obesity in children. We report the occurrence of insulin resistance and marked hyperandrogenism in prepubertal minority group girls (African-American and Caribbean Hispanic) with premature adrenarche. Approximately one-third of our prepubertal patients with premature adrenarche evaluated have been noted to have marked hyperandrogenism with ACTH stimulated levels of 17-hydroxypregnenolone and the ratio of 17-hydroxypregnenolone/17-hydroxyprogesterone more than two standard deviations above the mean of normal early pubertal girls (Tanner II-III). Furthermore, those girls with the more marked hyperandrogenism have been noted to have insulin resistance as assessed by the frequently sampled intravenous glucose tolerance test. A preliminary evaluation of adolescent girls previously evaluated for premature adrenarche has revealed that those girls with hyperandrogenism and insulin resistance when evaluated in the prepubertal period continue to have obesity, insulin resistance, hyperandrogenism and symptoms of hyperandrogenism (irregular menses, hirsutism and acne). Hence, the early identification of children at risk for insulin resistance should permit early intervention in order to avoid complications. PMID- 11117668 TI - The function and roles of P450c17 in androgen excess states. AB - Cytochrome P450c17 is a multifunctional enzyme that converts C21 steroids to the C19 sex steroid precursor DHEA. Intuitively, increased expression of P450c17 in the adrenals and ovaries might be expected to accompany androgen excess states, but the function of P450c17 in androgen biosynthesis is not so simple. The abundance of cofactor proteins and the coexistence of other enzymes contribute to the flux of precursor along pathways to different classes of active steroids. This paper explores the function of P450c17 in its proper biological context, using biochemical studies of patients with isolated 17,20-lyase activity to demonstrate key concepts in P450c17 enzymology. PMID- 11117669 TI - Sex hormone-binding globulin during puberty in normal and hyperandrogenic girls. AB - Human sex hormone-binding globulin (SHBG) regulates the cellular bioavailability of SHBG-bound steroid hormones. Subtle decreases in plasma SHBG levels during puberty have a perceptible effect on the androgen-estrogen balance. This SHBG decrease is more pronounced in girls with premature pubarche who are at risk to develop functional ovarian hyperandrogenism as well as insulin resistance syndrome. Insulin is a potent inhibitor of SHBG production in the liver, and there is now evidence that SHBG is a marker of hyperinsulinemia and insulin resistance that can be associated in both obese and non-obese patients with polycystic ovary syndrome. Therefore, low SHBG could be a useful tool for identifying presymptomatic individuals with diabetes mellitus type 2 including those with androgen disorders. PMID- 11117670 TI - The gonadotropic axis in hyperandrogenic adolescents. AB - Polycystic ovary syndrome (PCOS) is a common reproductive disorder that is first clinically diagnosable approximately 3 years after menarche. Women with PCOS have exaggerated gonadotropin secretion, with an elevated LH/FSH ratio, as well as an increased frequency and amplitude of LH pulsations. Since the elevated pulse frequency is a marker of unusually rapid hypothalamic GnRH secretion, these results imply a defect at the level of the hypothalamus. Such an abnormality could theoretically be a primary defect that could cause PCOS, and if so, would be expected to be detectable at the onset of puberty. How these abnormalities of hypothalamic and pituitary function relate to other, seemingly disparate, organ defects has not yet been delineated. We studied 99 PCOS patients and 42 normal controls with 8 to 12 hours of q10 min frequent blood sampling. Women with PCOS had significantly elevated mean LH levels, LH pulse amplitude, LH pulse frequency, and LH/FSH ratios compared to normal women. However, these abnormalities were not randomly distributed. All gonadotropin secretion parameters tended to be normal in PCOS patients when measured during the luteal phase or first 7 days after an ovulatory menses. There was a strong inverse relationship of mean LH, LH/FSH ratio, and LH pulse amplitude, but not frequency, with BMI. We hypothesize that this effect of body size occurs at the pituitary level. Several investigators have performed similar investigations in adolescent girls with regular and irregular menstrual cycles. Taken together, this evidence suggests that the gonadotropin defects appear as a very early finding in the development of PCOS, but the presence of abnormalities in both gonadotropin secretion and androgen secretion does not allow one to conclude that the gonadotropin defects are causal. We conclude that gonadotropin defects, particularly excess of serum LH, is a predominant finding in hyperandrogenic women, whether they are young peripubertal adolescents, or older perimenopausal women. Whether there is a primary neuroendocrine abnormality driving excess gonadotropin secretion that causes the hyperandrogenic condition, or whether the excess gonadotropin secretion reflects abnormal feedback of another factor, still remains unknown. PMID- 11117671 TI - Diagnosis of the polycystic ovary syndrome in adolescence: comparison of adolescent and adult hyperandrogenism. AB - We performed gonadotropin releasing hormone agonist (GnRHag) tests on 23 consecutive hyperandrogenic girls 9.9-17.5 years of age who were referred to our pediatric endocrinology clinic with symptoms suggestive of PCOS. They were compared to contemporaneously studied groups of adult normal and hyperandrogenic women. We found that hyperandrogenic adolescents had clinical and endocrine features similar to those of hyperandrogenic adults. However, there were some noteworthy unique features of adolescent hyperandrogenism, such as presentation in mid-childhood with premature pubarche and the occasional diagnosis before the age of 10 years. Some differences between adolescents and adults were statistically significant, for example, pelvic ultrasonography was not as helpful in the diagnosis of FOH as it is in adults. Nevertheless, a number of questions about the development of the ovarian dysfunction remain to be answered. For example, we are unable to diagnose ovarian dysfunction before puberty or in early puberty, and the relationship of "physiologic adolescent anovulation" to PCOS remains to be defined. PMID- 11117672 TI - Molecular defects of insulin action in the polycystic ovary syndrome: possible tissue specificity. AB - Insulin resistance is a highly common occurrence in PCOS. To determine the cellular and molecular mechanisms of this insulin resistance we studied subcutaneous abdominal adipocytes isolated from age and weight matched normal cycling (NC) and PCOS subjects. Insulin resistance in PCOS adipocytes was manifested as a reduction in insulin sensitivities for stimulation of glucose transport and suppression of catecholamine-activated lipolysis with no impact on final hormone responsiveness. Insulin sensitivity in adipocytes could be normalized by the addition of an adenosine analog. A number of key steps in insulin signaling pathways receptor autophosphorylation, IRS-1 phosphorylation, PI3-kinase activation and Akt phosphorylation--were found to display normal sensitivity and responsiveness in PCOS adipocytes. Insulin action measured in cultured fibroblasts was similar in NC and PCOS cells. Insulin resistance in PCOS involves tissue and response-specific defects in insulin signal transduction that remain to be identified. Interventions that can increase insulin sensitivity, such as adenosine, may have therapeutic potential in treating the unique insulin resistance of PCOS. PMID- 11117673 TI - Role of inositolphosphoglycan mediators of insulin action in the polycystic ovary syndrome. AB - Evidence suggests that some actions of insulin are mediated by putative inositolphosphoglycan (IPG) mediators, also known as second messengers. We review studies indicating that the IPG signaling system transduces insulin's stimulation of human thecal androgen biosynthesis, thus offering a mechanism by which insulin can stimulate ovarian androgen production even in women with PCOS whose tissues are resistant to insulin's stimulation of glucose metabolism. Furthermore, a deficiency in a specific D-chiro-inositol-containing IPG may contribute to insulin resistance in women with PCOS. In support of this idea, administration of D-chiro-inositol has been demonstrated to improve glucose tolerance, decrease serum androgens and improve ovulation in PCOS. The hypothesis is advanced that PCOS may be characterized by a defect in the conversion of myo-inositol to D chiro-inositol, and that such a defect would contribute to both insulin resistance and hyperandrogenism in the syndrome. PMID- 11117674 TI - Glucose intolerance in the polycystic ovary syndrome: role of the pancreatic beta cell. AB - Women with polycystic ovary syndrome (PCOS) are at increased risk of developing diabetes mellitus type 2. Insulin resistance plays a key role in the predisposition to diabetes in PCOS; however, defects in insulin secretion also appear to contribute to its development. Since diabetes mellitus is not a universal consequence in PCOS, however, it is important to develop means to identify those women who are at highest risk. In this way, it may become possible to delay or even prevent the onset of diabetes mellitus in later life. Identification of genetic factors and use of pharmacological agents may allow early identification of those women with PCOS who are at greatest risk for development of diabetes mellitus type 2. PMID- 11117675 TI - Family history as a risk factor for the polycystic ovary syndrome. AB - We have prospectively studied, by interview, clinical examination and biochemical evaluation, the relatives of 195 consecutive PCOS patients in order to: 1) accurately determine the prevalence of PCOS, as defined by current endocrinological criteria, among first-degree relatives of affected patients, and 2) determine the overall accuracy of proband-only and family member (self report) interview for the detection of clinically evident PCOS within families. We noted that 35% of mothers and 40% of sisters of patients with PCOS will be affected by PCOS themselves. Overall, the interview using a standardized form, whether of the proband or the family relative directly, appears to be a reliable predictor of affected status in mothers. Alternatively, approximately 50% of sisters will be missed using the proband interview, although self-reporting appears to be a reasonably reliable predictor of affected status for these relatives. While we are unable to exclude an autosomal or X-linked dominant mode of inheritance, the heritability of PCOS is probably more complex, similar to that of diabetes mellitus type 2 and cardiovascular disease. In absence of molecular diagnostic markers, a positive family history appears to be the most informative risk factor for the development PCOS. PMID- 11117676 TI - Is there a male phenotype in polycystic ovary syndrome families? AB - The male phenotype in PCOS is still uncertain. Multiple possible phenotypes have been proposed including such abnormalities in male hair distribution as premature balding and metabolic abnormalities such as insulin resistance. Problems with phenotyping to date are reviewed and potential new paths for familial phenotyping are discussed. Identifying a male phenotype would increase the sample size for genetic analyses of the causative gene. It may also improve the phenotyping of female family members by identifying familial traits that are gender independent. PMID- 11117677 TI - Searching for the polycystic ovary syndrome genes. AB - PCOS is a common disorder of unknown etiology. Studies of first-degree relatives of women diagnosed with PCOS suggest familial clustering of the disease. A prospective study of first-degree female relatives of women with PCOS conducted by NCPIR found that 46% of ascertainable sisters of women with PCOS were hyperandrogenemic. NCPIR has conducted linkage and association studies using affected sibling-pair analysis and the transmission/disequilibrium test to explore candidate PCOS genes. These studies point a finger at a region 1 MB centromeric to the insulin receptor gene on chromosome 19. PMID- 11117678 TI - The role of heterozygosity for CYP21 in the polycystic ovary syndrome. AB - The phenotypic heterogeneity recognized in congenital adrenal hyperplasia secondary to 21-hydroxylase deficiency appears to extend to 21hydroxylase (CYP21) mutation carriers. To begin the search for modifying loci responsible for this phenotypic heterogeneity, we performed CYP21 genotype analysis and assays for three candidate modifier loci on genomic DNA samples obtained from 30 adolescent girls with hyperandrogenism, 14 healthy control women, and 15 female obligate CYP21 mutation carriers. The frequency of heterozygosity for CYP21 mutations was increased in women with symptomatic hyperandrogenism (10/30) compared to healthy controls (1/14). There were no significant differences in the frequencies of the modifier variants among the three groups. Although the small sample size precludes strong conclusions, CYP21 nonsense mutation carriers tend to be asymptomatic while missense mutation carriers, i.e. V281L, appear to manifest a PCOS phenotype. Evaluation of additional modifying loci in larger series of patients will help identify new genetic markers associated with PCOS. PMID- 11117679 TI - Watching your health go up in smoke. PMID- 11117680 TI - Fiberoptic nasopharyngolaryngoscopy for airway monitoring after obstructive sleep apnea surgery. AB - PURPOSE: This study evaluated the upper airway characteristics in the early postoperative period after reconstructive surgery for obstructive sleep apnea (OSA). METHODS: During a 24-month period, the upper airway of patients who underwent uvulopalatopharyngoplasty (UPPP) with genioglossus advancement (GA) or hyoid myotomy (HM) or maxillomandibular advancement (MMA) were evaluated with fiberoptic nasopharyngolaryngoscopy (NPG) preoperatively and 24 to 72 hours postoperatively. RESULTS: NPG was performed on 271 patients. One hundred seventy three patients had UPPP with GA or HM, and the remainder had MMA. All of the patients who underwent UPPP with GA or HM were found to have varying degrees of soft tissue edema involving the soft palate and the tongue base. The patients who underwent tonsillectomies and UPPP with GA or HM had greater soft palate/pharyngeal wall edema. In contrast, patients who underwent MMA had minimal edema involving the soft palate and the base of tongue, but diffuse lateral pharyngeal wall edema throughout the upper airway was identified. Eighteen of the MMA patients had ecchymosis and edema involving the pyriform sinus and aryepiglottic fold; 4 of these patients also had a hypopharyngeal hematoma involving the pyriform sinus, aryepiglottic fold, arytenoid, and false vocal cord, which partially obstructed the airway. These 4 patients were closely monitored for 1 to 2 additional days, and all were discharged without problems. None of the patients in the study had postoperative airway obstruction. CONCLUSION: NPG may be useful in postoperative airway monitoring and assist in discharge planning after upper airway reconstruction in the OSA patients. PMID- 11117681 TI - Cervical necrotizing fasciitis of odontogenic origin: a report of 11 cases. AB - PURPOSE: Although most cases of cervical necrotizing fasciitis (CNF) are odontogenic in origin, reports of this disease in the dental literature are sparse. The purpose of this study was to review the cases treated on our service, and to analyze the features of this disease and the responses to management, to supplement the understanding of this relatively rare and life-threatening disease. PATIENTS AND METHODS: All cases of infection admitted to the OMS service in a period of 10.5 years were studied retrospectively. The diagnosis of CNF was established by the findings on surgical exploration and histologic examination. The patients' age, sex, medical status, causes of the infection, bacteriology, computed tomography scan findings, surgical interventions, complications, survival, and other clinical parameters were reviewed. RESULTS: A total of 422 cases of infection were admitted, and 11 cases of cervical necrotizing fasciitis were found. The incidence of CNF was 2.6% among the infections hospitalized on the OMS service. There were 7 male and 4 female patients. Eight patients were older than 60 years of age. Seven patients had immunocompromising conditions, including diabetes mellitus in 4, concurrent administration of steroid in 2, uremia in 1, and a thymus carcinoma in 1. All patients showed parapharyngeal space involvement; four also showed retropharyngeal space involvement. Gas was found in the computed tomography scan in 6 patients, extending to cranial base in 3 of them. Anaerobes were isolated in 73% of the infections, whereas Streptococcus species were uniformly present. All patients received 1 or more debridements. Major complications occurred in 4 patients, including mediastinitis in 4, septic shock in 2, lung empyema in 1, pleural effusion in 2, and pericardial effusion in 1. All major complications developed in the immunocompromised patients, leading to 2 deaths. CONCLUSION: The mortality rate in this study was 18%. Early surgical debridement, intensive medical care, and a multidisciplinary approach are advocated in the management of CNF. PMID- 11117682 TI - Neurosensory differences after wire and rigid fixation in patients with mandibular advancement. AB - PURPOSE: The purpose of this analysis was to compare the frequency and severity of nerve damage with rigid and wire fixation in patients participating in a prospective, randomized clinical trial. PATIENTS AND METHODS: One hundred twenty six patients who required a bilateral sagittal split osteotomy and mandibular advancement were randomly assigned to receive either rigid or wire fixation. Tactile sensation in the mental nerve region bilaterally was determined presurgically and throughout the subsequent 2 years by using monofilament detection and brush stroke direction. Neurosensory levels were compared between the types of fixation over time. RESULTS: Evaluation with monofilament detection showed no significant difference between types of fixation throughout the 2-year follow-up. However, brush stroke determination showed significantly greater hypesthesia with rigid compared with wire fixation from 8 weeks through 2 years postoperatively. CONCLUSION: Rigid fixation resulted in more anesthesia in the mental nerve distribution than wire fixation when tested with brush stroke direction. However, increased anesthesia was not present when measured with monofilament determination. PMID- 11117683 TI - Reconstruction of orbital fractures with dehydrated human dura mater. AB - PURPOSE: The best method for reconstructing the fractured orbital floor remains controversial. This article evaluates the usefulness of dehydrated human dura mater for orbital floor reconstruction after facial trauma. PATIENTS AND METHODS: A retrospective analysis of 55 patients who had undergone surgical repair of orbital fractures was performed. The dura mater was used when the disruption was less than 2 cm in diameter. Fractures were divided into 3 types: type I (blow out), type II (orbitozygomatic fracture), and type III (midfacial fracture). The patients were followed-up at least 1 year after surgery, and the cosmetic and functional results were reviewed. RESULTS: A 7% complication rate was noted. No implant migration or infection resulted. One year postsurgery, all patients showed a complete resolution of their diplopia. CONCLUSION: The safety and biocompatibility of dehydrated human dura mater support its use in orbital defects less than 2 cm in diameter. PMID- 11117684 TI - The mandibular symphysis as a donor site in maxillofacial bone grafting: a quantitative anatomic study. AB - PURPOSE: The goal of this study was to quantify the amount of bone graft material present in the mandibular symphysis as well as to determine the maximal size of the corticocancellous bone block that can be harvested while avoiding mental nerve injury, tooth injury, and simultaneously preserving the preoperative facial contour. MATERIALS AND METHODS: Sixteen fixed dentate cadaver mandibles were studied. Osteotomies were performed in a monocortical fashion, 5 mm anterior to the mental foramen, cephalad to the inferior border of the mandible, caudal to the expected position of the apices of the anterior teeth, and at the midline. The size of the corticocancellous block was then measured. Bone volume, using 2 techniques, was measured by displacement volumetry. RESULTS: The average volumes obtained were 4.84 mL and 4.71 mL (range, 3.25 to 6.50 mL), respectively, for the 2 techniques of volumetry. The average block size was 20.9 x 9.9 x 6.9 mm; the smallest block was 21.0 x 6.5 x 6.0 mm, and the largest was 25.0 x 13.0 x 9.0 mm. CONCLUSION: Based on the results of this study, it is apparent that the mandibular symphysis can be reliably selected as the harvest site in a variety of oral and maxillofacial reconstructive procedures. PMID- 11117685 TI - Survival of hydroxyapatite-coated implants: a meta-analytic review. AB - PURPOSE: Some reports show a benefit of coating dental implants with hydroxyapatite (HA), and others assert that resorption of the HA coating compromises long-term implant survival. This study examined this controversy by systematically reviewing all the current literature that reports the outcomes of HA-coated implants in human clinical trials. MATERIALS AND METHODS: A systematic Medline computer search of the English literature yielded 45 human clinical trials that reported on the outcome of HA-coated implants. Eleven studies that met specific inclusion criteria were selected for detailed analysis. The studies were divided into 2 groups. One group of 5 studies reported implant survival using overall percentage, and another group of 6 studies reported implant survival using life-table analysis. RESULTS: The overall percentage survival rates ranged from 93.2% to 98.5%, with 4 to 8 years of follow-up. The cumulative survival rates for studies that used life-table analysis ranged from 79.2% to 98.5%, with 5 to 8 years of follow-up. The yearly interval survival rates reported for the studies using life-tables were variable but remained above 90% and did not show a progressive or precipitous decrease with increasing years of follow-up. CONCLUSIONS: The survival rates reported for HA-coated implants were similar to the survival rates reported for uncoated titanium implants. If resorption of the HA coating causes late failure of implants, the yearly interval survival rates should have decreased with increased years of follow-up. This decrease was not observed in the longitudinal human clinical trials that met the selection criteria for this study. Detailed analysis of these clinical trials did not show that HA-coating compromises the long-term survival of dental implants. PMID- 11117686 TI - Computerized 3-dimensional study of the embryologic development of the human masticatory muscles and temporomandibular joint. AB - PURPOSE: In this study, the development of human embryonic temporomandibular joint (TMJ) and masticatory muscles were investigated by using computed 3 dimensional reconstructions. MATERIALS AND METHODS: Sixteen human embryos and fetuses, ranging from 6.5 to 107 mm crown-rump length, were examined. RESULTS: At 10 weeks, a band of mesenchyme extending from the attachment of the lateral pterygoid muscle to the condylar process was observed to pass through the medial side of the condylar process to attach to the malleus. The temporal, masseter, and pterygoid muscles develop from the so called "temporal muscle" primordium, and the temporal muscle was in continuity with the masseter muscle until 14 weeks of fetal life. CONCLUSIONS: The study shows that the muscles of mastication arise from a single primordium. It also confirms the presence of a ligamentous attachment between the lateral pterygoid muscle and the malleus. PMID- 11117687 TI - Vaporization of melanin in oral tissues and skin with a carbon dioxide laser: a canine study. AB - PURPOSE: This study tested the efficacy of carbon dioxide (CO2) laser vaporization in ablating gingival, oral mucosal, and cutaneous melanin in dogs. MATERIALS AND METHODS: Three mongrel dogs with pigmentation of the oral mucosa, gingivae, and skin were used. Biopsy specimens from the melanin sites were obtained from all 3 dogs before laser application. Removal of the melanin sites was performed by using a 3 W continuous-wave CO2 laser. Biopsies of the treated areas were performed 1, 2, 4, and 6 weeks after laser treatment in all 3 dogs and also, at 11 weeks in 1 dog. The histologic sections were stained with hematoxylin and eosin and Masson-Fontana dye. A computerized morphometric program calculated the average percentage of the melanin layer in the Masson-Fontana-stained sections. RESULTS: Clinical and histologic examination showed the CO2 laser to be effective in eliminating the pigmented areas in all tissues treated. No recurrence of melanin was detected in either the oral mucosa or gingiva at any of the follow-up times. In the skin, however, a small amount of melanin repigmentation was noticeable. CONCLUSIONS: CO2 laser surgery proved an effective tool for obliterating superficial melanin discoloration. To prevent recurrence of the pigmentation, the area must be cleared completely of melanin, directing the laser beam carefully along the visible margins of the area. PMID- 11117688 TI - Expansile intraosseous lesion of the maxilla. PMID- 11117689 TI - A comparison of performance on the OMSITE and ABOMS written qualifying examination. Oral and Maxillofacial Surgery In-Training Examination. American Board of Oral and Maxillofacial Surgery. AB - PURPOSE: The purpose of this study was to see whether there was a correlation between the performance of persons on the Oral and Maxillofacial Surgery In Training Examination (OMSITE) and their performance on the American Board of Oral and Maxillofacial Surgery Written Qualifying Examination (ABOMS WQE). METHODS: All OMSITE scores for residents in their last years of training during the years 1992 to 1998 and their scores on the ABOMS WQE were tabulated by American College Testing (ACT) (Iowa City, IA) and submitted for analysis. The data were analyzed using Pearson's correlation coefficients to determine any relationship between the 2 scores. Likelihood ratios were calculated to show the probability of candidates passing the ABOMS WQE based on their OMSITE score. RESULTS: There were 765 scores provided by ACT for the years 1992 to 1998. A significantly positive correlation existed between the OMSITE and ABOMS WQE raw scores for those taking the ABOMS WQE for the first and second times, but not for subsequent attempts. Persons who scored a raw score of over 211 on the OMSITE all passed the ABOMS WQE on their first attempt. Those with a raw score of less than 134 on their OMSITE all failed the ABOMS WQE on their first attempt. CONCLUSION: A highly positive correlation exists between candidate performance on the OMSITE and the ABOMS WQE. PMID- 11117690 TI - Squamous cell carcinoma of the tongue in a 13-year-old boy. PMID- 11117692 TI - Correction of micrognathia attributable to ankylosis of the temporomandibular joint using a gradual distraction technique: case report. PMID- 11117693 TI - Necrotizing sialometaplasia: report of a case after lower lip mucocele excision. PMID- 11117691 TI - Treatment of Frey's syndrome with type A botulinum toxin: case report. PMID- 11117694 TI - Malignant transformation of a gigantic pleomorphic adenoma of the submandibular gland: a case report. PMID- 11117695 TI - Reconstruction of a large mandibular defect by distraction osteogenesis: a case report. PMID- 11117696 TI - Intraoral mandibular distraction osteogenesis in a patient with severe micrognathia secondary to TMJ ankylosis using a tooth and bone-anchored device (PIT device): a case report. PMID- 11117697 TI - Distraction osteogenesis of a fibular revascularized flap for improvement of oral implant positioning in a tumor patient: a case report. PMID- 11117698 TI - Familial florid cemento-osseous dysplasia: a case report. PMID- 11117699 TI - Nodular fascitis of the maxilla in a child. PMID- 11117700 TI - Mandibular supernumerary tooth causing neurosensory changes: a case report. PMID- 11117701 TI - Extra-articular ankylosis of the mandible after failed orthognathic surgery: report of a case. PMID- 11117702 TI - Squamous cell carcinoma of the tongue after bone marrow transplantation in a patient with Fanconi's anemia. PMID- 11117703 TI - Disagreement over degrees. PMID- 11117704 TI - Disagreement over degrees. PMID- 11117705 TI - Missing the point. PMID- 11117706 TI - Critical politics of carbon sinks. PMID- 11117707 TI - Wellcome Trust funds bid to unravel zebrafish genome. PMID- 11117708 TI - Deadlock in The Hague, but hopes remain for spring climate deal. PMID- 11117710 TI - German universities used forced labour. PMID- 11117709 TI - Argentinian researchers fight government plans for reform. PMID- 11117711 TI - NASA U-turns enrage planetary scientists. PMID- 11117712 TI - Dispute over rock dating settled out of court. PMID- 11117714 TI - Chinese science goes to Earth. PMID- 11117713 TI - Germany rues 'complacency' over BSE testing strategy. PMID- 11117715 TI - Britain plumps for support of big project...but ESO membership comes at a price. PMID- 11117716 TI - To the planets on a shoestring. PMID- 11117717 TI - Important differences between sources of embryonic stem cells. PMID- 11117718 TI - Weak euro hits PhDs too. PMID- 11117719 TI - Bernoulli was ahead of modern epidemiology. PMID- 11117720 TI - Aquaculture: part of the problem, not a solution. PMID- 11117721 TI - Funding would prevent waste of research time. PMID- 11117722 TI - What dreams may come? PMID- 11117723 TI - Too many memories. PMID- 11117724 TI - Fighting the Ebola virus. PMID- 11117725 TI - Superconductivity. C60--the hole story. PMID- 11117726 TI - Evolutionary biology. The problem of variation. PMID- 11117727 TI - Biogeography. Chile refuges. PMID- 11117728 TI - Earthquake science. Shaking faults loose. PMID- 11117729 TI - Bone versus immune system. PMID- 11117730 TI - Cultural revolution in whale songs. PMID- 11117731 TI - A stable binary quasicrystal. PMID- 11117733 TI - Numismatic gyrations. PMID- 11117732 TI - DNA methylation in Drosophila melanogaster. PMID- 11117734 TI - Electronics using hybrid-molecular and mono-molecular devices. AB - The semiconductor industry has seen a remarkable miniaturization trend, driven by many scientific and technological innovations. But if this trend is to continue, and provide ever faster and cheaper computers, the size of microelectronic circuit components will soon need to reach the scale of atoms or molecules--a goal that will require conceptually new device structures. The idea that a few molecules, or even a single molecule, could be embedded between electrodes and perform the basic functions of digital electronics--rectification, amplification and storage--was first put forward in the mid-1970s. The concept is now realized for individual components, but the economic fabrication of complete circuits at the molecular level remains challenging because of the difficulty of connecting molecules to one another. A possible solution to this problem is 'mono-molecular' electronics, in which a single molecule will integrate the elementary functions and interconnections required for computation. PMID- 11117735 TI - Superconductivity at 52 K in hole-doped C60. AB - Superconductivity in electron-doped C60 was first observed almost ten years ago. The metallic state and superconductivity result from the transfer of electrons from alkaline or alkaline-earth ions to the C60 molecule, which is known to be a strong electron acceptor. For this reason, it is very difficult to remove electrons from C60--yet one might expect to see superconductivity at higher temperatures in hole-doped than in electron-doped C60, because of the higher density of electronic states in the valence band than in the conduction band. We have used the technique of gate-induced doping in a field-effect transistor configuration to introduce significant densities of holes into C60. We observe superconductivity over an extended range of hole density, with a smoothly varying transition temperature Tc that peaks at 52 K. By comparison with the well established dependence of Tc on the lattice parameter in electron-doped C60, we anticipate that Tc values significantly in excess of 100 K should be achievable in a suitably expanded, hole-doped C60 lattice. PMID- 11117736 TI - Genetic control and evolution of sexually dimorphic characters in Drosophila. AB - Sexually dimorphic abdominal pigmentation and segment morphology evolved recently in the melanogaster species group of the fruitfly Drosophila. Here we show that these traits are controlled by the bric a brac [corrected] (bab) gene, which integrates regulatory inputs from the homeotic and sex-determination pathways. bab expression is modulated segment- and sex-specifically in sexually dimorphic species, but is uniform in sexually monomorphic species. We suggest that bab has an ancestral homeotic function, and that regulatory changes at the bab locus played a key role in the evolution of sexual dimorphism. Pigmentation patterns specified by bab affect mating preferences, suggesting that sexual selection has contributed to the evolution of bab regulation. PMID- 11117737 TI - Evidence against a redshift z > 6 for the galaxy STIS123627+621755. AB - The identification of galaxies at extreme distances provides the most direct information about the earliest phases of galaxy formation. But at redshifts z > 5 even the most luminous galaxies appear faint; the interpretation of low signal-to noise ratio data is difficult and misidentifications do occur. Here we report optical and near-infrared observations of the source STIS123627+621755, which was previously suggested to be at a redshift of 6.68 (ref. 1). At that redshift, and with the reported spectral energy distribution, the galaxy should be essentially invisible at wavelengths less than 9,300 A, because the intervening intergalactic medium absorbs almost all light energetic enough to ionize neutral hydrogen--that is, with wavelengths less than the redshifted Lyman limit of lambda = (1 + z) x 912A. At near-infrared wavelengths, however, the galaxy should be relatively bright. Here we report a detection of the galaxy at 6,700 A and a non-detection at a wavelength of 1.2 microm, contrary to expectations for z approximately 6.68. The data conservatively require that STIS123627+621755 has a redshift z < 6. PMID- 11117738 TI - Unusual spectral energy distribution of a galaxy previously reported to be at redshift 6.68. AB - Observations of distant galaxies are important both for understanding how galaxies form and for probing the physical conditions of the Universe at early times. It is, however, very difficult to identify galaxies at redshifts z > 5, because they are so faint and have few spectral characteristics. We previously reported the probable identification of a galaxy at z = 6.68, based on one line and an apparent break in the spectrum just shortwards of that, which we interpreted as Lyman alpha emission and the Lyman alpha break, where photons with shorter wavelengths are absorbed by the intervening neutral hydrogen gas. Here we present optical photometry that shows moderate detections of light in the B- and V-band images, which are inconsistent with the expected absence of flux shortwards of the Lyman alpha break for alpha galaxy at z > 5, and inconsistent with the previous flux measurement. Moreover, the spectral energy distribution for this object cannot readily be fitted by any known galaxy spectral template at any redshift, so the redshift is undetermined. PMID- 11117739 TI - First-order transition in confined water between high-density liquid and low density amorphous phases. AB - Supercooled water and amorphous ice have a rich metastable phase behaviour. In addition to transitions between high- and low-density amorphous solids, and between high- and low-density liquids, a fragile-to-strong liquid transition has recently been proposed, and supported by evidence from the behaviour of deeply supercooled bilayer water confined in hydrophilic slit pores. Here we report evidence from molecular dynamics simulations for another type of first-order phase transition--a liquid-to-bilayer amorphous transition--above the freezing temperature of bulk water at atmospheric pressure. This transition occurs only when water is confined in a hydrophobic slit pore with a width of less than one nanometre. On cooling, the confined water, which has an imperfect random hydrogen bonded network, transforms into a bilayer amorphous phase with a perfect network (owing to the formation of various hydrogen-bonded polygons) but no long-range order. The transition shares some characteristics with those observed in tetrahedrally coordinated substances such as liquid silicon, liquid carbon and liquid phosphorus. PMID- 11117740 TI - Changes in deep-water formation during the Younger Dryas event inferred from 10Be and 14C records. AB - Variations in atmospheric radiocarbon (14C) concentrations can be attributed either to changes in the carbon cycle--through the rate of radiocarbon removal from the atmosphere--or to variations in the production rate of 14C due to changes in solar activity or the Earth's magnetic field. The production rates of 10Be and 14C vary in the same way, but whereas atmospheric radiocarbon concentrations are additionally affected by the carbon cycle, 10Be concentrations reflect production rates more directly. A record of the 10Be production-rate variations can therefore be used to separate the two influences--production rates and the carbon cycle--on radiocarbon concentrations. Here we present such an analysis of the large fluctuations in atmospheric 14C concentrations, of unclear origin, that occurred during the Younger Dryas cold period. We use the 10Be record from the GISP2 ice core to model past production rates of radionuclides, and find that the largest part of the fluctuations in atmospheric radiocarbon concentrations can be attributed to variations in production rate. The residual difference between measured 14C concentrations and those modelled using the 10Be record can be explained with an additional change in the carbon cycle, most probably in the amount of deep-water formation. PMID- 11117741 TI - Triggering of earthquake aftershocks by dynamic stresses. AB - It is thought that small 'static' stress changes due to permanent fault displacement can alter the likelihood of, or trigger, earthquakes on nearby faults. Many studies of triggering in the near-field, particularly of aftershocks, rely on these static changes as the triggering agent and consider them only in terms of equivalent changes in the applied load on the fault. Here we report a comparison of the aftershock pattern of the moment magnitude Mw = 7.3 Landers earthquake, not only with static stress changes but also with transient, oscillatory stress changes transmitted as seismic waves (that is, 'dynamic' stresses). Dynamic stresses do not permanently change the applied load and thus can trigger earthquakes only by altering the mechanical state or properties of the fault zone. These dynamically weakened faults may fail after the seismic waves have passed by, and might even cause earthquakes that would not otherwise have occurred. We find similar asymmetries in the aftershock and dynamic stress patterns, the latter being due to rupture propagation, whereas the static stress changes lack this asymmetry. Previous studies have shown that dynamic stresses can promote failure at remote distances, but here we show that they can also do so nearby. PMID- 11117742 TI - Geochemical evidence for terrestrial ecosystems 2.6 billion years ago. AB - Microorganisms have flourished in the oceans since at least 3.8 billion years (3.8 Gyr) ago, but it is not at present clear when they first colonized the land. Organic matter in some Au/U-rich conglomerates and ancient soils of 2.3-2.7 Gyr age has been suggested as remnants of terrestrial organisms. Some 2.7-Gyr-old stromatolites have also been suggested as structures created by terrestrial organisms. However, it has been disputed whether this organic matter is indigenous or exogenic, and whether these stromatolites formed in marine or fresh water. Consequently, the oldest undisputed remnants of terrestrial organisms are currently the 1.2-Gyr-old microfossils from Arizona, USA. Unusually carbonaceous ancient soils--palaeosols--have been found in the Mpumalanga Province (Eastern Transvaal) of South Africa. Here we report the occurrences, elemental ratios (C, H, N, P) and isotopic compositions of this organic matter and its host rocks. These data show that the organic matter very probably represents remnants of microbial mats that developed on the soil surface between 2.6 and 2.7 Gyr ago. This places the development of terrestrial biomass more than 1.4 billion years earlier than previously reported. PMID- 11117743 TI - Nutritional constraints in terrestrial and freshwater food webs. AB - Biological and environmental contrasts between aquatic and terrestrial systems have hindered analyses of community and ecosystem structure across Earth's diverse habitats. Ecological stoichiometry provides an integrative approach for such analyses, as all organisms are composed of the same major elements (C, N, P) whose balance affects production, nutrient cycling, and food-web dynamics. Here we show both similarities and differences in the C:N:P ratios of primary producers (autotrophs) and invertebrate primary consumers (herbivores) across habitats. Terrestrial food webs are built on an extremely nutrient-poor autotroph base with C:P and C:N ratios higher than in lake particulate matter, although the N:P ratios are nearly identical. Terrestrial herbivores (insects) and their freshwater counterparts (zooplankton) are nutrient-rich and indistinguishable in C:N:P stoichiometry. In both lakes and terrestrial systems, herbivores should have low growth efficiencies (10-30%) when consuming autotrophs with typical carbon-to-nutrient ratios. These stoichiometric constraints on herbivore growth appear to be qualitatively similar and widespread in both environments. PMID- 11117744 TI - Bacterial dehalorespiration with chlorinated benzenes. AB - Chlorobenzenes are toxic, highly persistent and ubiquitously distributed environmental contaminants that accumulate in the food chain. The only known microbial transformation of 1,2,3,5-tetrachlorobenzene (TeCB) and higher chlorinated benzenes is the reductive dechlorination to lower chlorinated benzenes under anaerobic conditions observed with mixed bacterial cultures. The lower chlorinated benzenes can subsequently be mineralized by aerobic bacteria. Here we describe the isolation of the oxygen-sensitive strain CBDB1, a pure culture capable of reductive dechlorination of chlorobenzenes. Strain CBDB1 is a highly specialized bacterium that stoichiometrically dechlorinates 1,2,3 trichlorobenzene (TCB), 1,2,4-TCB, 1,2,3,4-TeCB, 1,2,3,5-TeCB and 1,2,4,5-TeCB to dichlorobenzenes or 1,3,5-TCB. The presence of chlorobenzene as an electron acceptor and hydrogen as an electron donor is essential for growth, and indicates that strain CBDB1 meets its energy needs by a dehalorespiratory process. According to their 16S rRNA gene sequences, strain CBDB1, Dehalococcoides ethenogenes and several uncultivated bacteria form a new bacterial cluster, of which strain CBDB1 is the first, so far, to thrive on a purely synthetic medium. PMID- 11117745 TI - Calcium stores regulate the polarity and input specificity of synaptic modification. AB - Activity-induced synaptic modification is essential for the development and plasticity of the nervous system. Repetitive correlated activation of pre- and postsynaptic neurons can induce persistent enhancement or decrement of synaptic efficacy, commonly referred to as long-term potentiation or depression (LTP or LTD). An important unresolved issue is whether and to what extent LTP and LTD are restricted to the activated synapses. Here we show that, in the CA1 region of the hippocampus, reduction of postsynaptic calcium influx by partial blockade of NMDA (N-methyl-D-aspartate) receptors results in a conversion of LTP to LTD and a loss of input specificity normally associated with LTP, with LTD appearing at heterosynaptic inputs. The induction of LTD at homo- and heterosynaptic sites requires functional ryanodine receptors and inositol triphosphate (InsP3) receptors, respectively. Functional blockade or genetic deletion of type 1 InsP3 receptors led to a conversion of LTD to LTP and elimination of heterosynaptic LTD, whereas blocking ryanodine receptors eliminated only homosynaptic LTD. Thus, postsynaptic Ca2+, deriving from Ca2+ influx and differential release of Ca2+ from internal stores through ryanodine and InsP3 receptors, regulates both the polarity and input specificity of activity-induced synaptic modification. PMID- 11117746 TI - Analysis of calcium channels in single spines using optical fluctuation analysis. AB - Most synapses form on small, specialized postsynaptic structures known as dendritic spines. The influx of Ca2+ ions into such spines--through synaptic receptors and voltage-sensitive Ca2+ channels (VSCCs)--triggers diverse processes that underlie synaptic plasticity. Using two-photon laser scanning microscopy, we imaged action-potential-induced transient changes in Ca2+ concentration in spines and dendrites of CA1 pyramidal neurons in rat hippocampal slices. Through analysis of the large trial-to-trial fluctuations in these transients, we have determined the number and properties of VSCCs in single spines. Here we report that each spine contains 1-20 VSCCs, and that this number increases with spine volume. We are able to detect the opening of a single VSCC on a spine. In spines located on the proximal dendritic tree, VSCCs normally open with high probability (approximately 0.5) following dendritic action potentials. Activation of GABA(B) receptors reduced this probability in apical spines to approximately 0.3 but had no effect on VSCCs in dendrites or basal spines. Our studies show that the spatial distribution of VSCC subtypes and their modulatory potential is regulated with submicrometre precision. PMID- 11117747 TI - Role of cortical tumour-suppressor proteins in asymmetric division of Drosophila neuroblast. AB - Cellular diversity during development arises in part from asymmetric divisions, which generate two distinct cells by transmitting localized determinants from a progenitor cell into one daughter cell. In Drosophila, neuroblasts undergo typical asymmetric divisions to produce another neuroblast and a ganglion mother cell. At mitosis, neural fate determinants, including Prospero and Numb, localize to the basal cortex, from which the ganglion mother cell buds off; Inscuteable and Bazooka, which regulate spindle orientation, localize apically. Here we show that a tumour-suppressor protein, Lethal giant larvae (Lgl), is essential for asymmetric cortical localization of all basal determinants in mitotic neuroblasts, and is therefore indispensable for neural fate decisions. Lgl, which itself is uniformly cortical, interacts with several types of Myosin to localize the determinants. Another tumour-suppressor protein, Lethal discs large (Dlg), participates in this process by regulating the localization of Lgl. The localization of the apical components is unaffected in lgl or dlg mutants. Thus, Lgl and Dlg act in a common process that differentially mediates cortical protein targeting in mitotic neuroblasts, and that creates intrinsic differences between daughter cells. PMID- 11117748 TI - The tumour-suppressor genes lgl and dlg regulate basal protein targeting in Drosophila neuroblasts. AB - Drosophila neuroblasts are a model system for studying asymmetric cell division: they divide unequally to produce an apical neuroblast and a basal ganglion mother cell that differ in size, mitotic activity and developmental potential. During neuroblast mitosis, an apical protein complex orients the mitotic spindle and targets determinants of cell fate to the basal cortex, but the mechanism of each process is unknown. Here we show that the tumour-suppressor genes lethal giant larvae (lgl) and discs large (dlg) regulate basal protein targeting, but not apical complex formation or spindle orientation, in both embryonic and larval neuroblasts. Dlg protein is apically enriched and is required for maintaining cortical localization of Lgl protein. Basal protein targeting requires microfilament and myosin function, yet the lgl phenotype is strongly suppressed by reducing levels of myosin II. We conclude that Dlg and Lgl promote, and myosin II inhibits, actomyosin-dependent basal protein targeting in neuroblasts. PMID- 11117750 TI - Development of a preventive vaccine for Ebola virus infection in primates. AB - Outbreaks of haemorrhagic fever caused by the Ebola virus are associated with high mortality rates that are a distinguishing feature of this human pathogen. The highest lethality is associated with the Zaire subtype, one of four strains identified to date. Its rapid progression allows little opportunity to develop natural immunity, and there is currently no effective anti-viral therapy. Therefore, vaccination offers a promising intervention to prevent infection and limit spread. Here we describe a highly effective vaccine strategy for Ebola virus infection in non-human primates. A combination of DNA immunization and boosting with adenoviral vectors that encode viral proteins generated cellular and humoral immunity in cynomolgus macaques. Challenge with a lethal dose of the highly pathogenic, wild-type, 1976 Mayinga strain of Ebola Zaire virus resulted in uniform infection in controls, who progressed to a moribund state and death in less than one week. In contrast, all vaccinated animals were asymptomatic for more than six months, with no detectable virus after the initial challenge. These findings demonstrate that it is possible to develop a preventive vaccine against Ebola virus infection in primates. PMID- 11117749 TI - T-cell-mediated regulation of osteoclastogenesis by signalling cross-talk between RANKL and IFN-gamma. AB - Bone resorption is regulated by the immune system, where T-cell expression of RANKL (receptor activator of nuclear factor (NF)-kappaB ligand), a member of the tumour-necrosis factor family that is essential for osteoclastogenesis, may contribute to pathological conditions, such as autoimmune arthritis. However, whether activated T cells maintain bone homeostasis by counterbalancing the action of RANKL remains unknown. Here we show that T-cell production of interferon (IFN)-gamma strongly suppresses osteoclastogenesis by interfering with the RANKL-RANK signalling pathway. IFN-gamma induces rapid degradation of the RANK adapter protein, TRAF6 (tumour necrosis factor receptor-associated factor 6), which results in strong inhibition of the RANKL-induced activation of the transcription factor NF-kappaB and JNK. This inhibition of osteoclastogenesis is rescued by overexpressing TRAF6 in precursor cells, which indicates that TRAF6 is the target critical for the IFN-gamma action. Furthermore, we provide evidence that the accelerated degradation of TRAF6 requires both its ubiquitination, which is initiated by RANKL, and IFN-gamma-induced activation of the ubiquitin proteasome system. Our study shows that there is cross-talk between the tumour necrosis factor and IFN families of cytokines, through which IFN-gamma provides a negative link between T-cell activation and bone resorption. Our results may offer a therapeutic approach to treat the inflammation-induced tissue breakdown. PMID- 11117751 TI - Coenzyme Q is an obligatory cofactor for uncoupling protein function. AB - Uncoupling proteins (UCPs) are thought to be intricately controlled uncouplers that are responsible for the futile dissipation of mitochondrial chemiosmotic gradients, producing heat rather than ATP. They occur in many animal and plant cells and form a subfamily of the mitochondrial carrier family. Physiological uncoupling of oxidative phosphorylation must be strongly regulated to avoid deterioration of the energy supply and cell death, which is caused by toxic uncouplers. However, an H+ transporting uncoupling function is well established only for UCP1 from brown adipose tissue, and the regulation of UCP1 by fatty acids, nucleotides and pH remains controversial. The failure of UCP1 expressed in Escherichia coli inclusion bodies to carry out fatty-acid-dependent H+ transport activity inclusion bodies made us seek a native UCP cofactor. Here we report the identification of coenzyme Q (ubiquinone) as such a cofactor. On addition of CoQ10 to reconstituted UCP1 from inclusion bodies, fatty-acid-dependent H+ transport reached the same rate as with native UCP1. The H+ transport was highly sensitive to purine nucleotides, and activated only by oxidized but not reduced CoQ. H+ transport of native UCP1 correlated with the endogenous CoQ content. PMID- 11117752 TI - The PSI-H subunit of photosystem I is essential for state transitions in plant photosynthesis. AB - Photosynthesis in plants involves two photosystems responsible for converting light energy into redox processes. The photosystems, PSI and PSII, operate largely in series, and therefore their excitation must be balanced in order to optimize photosynthetic performance. When plants are exposed to illumination favouring either PSII or PSI they can redistribute excitation towards the light limited photosystem. Long-term changes in illumination lead to changes in photosystem stoichiometry. In contrast, state transition is a dynamic mechanism that enables plants to respond rapidly to changes in illumination. When PSII is favoured (state 2), the redox conditions in the thylakoids change and result in activation of a protein kinase. The kinase phosphorylates the main light harvesting complex (LHCII) and the mobile antenna complex is detached from PSII. It has not been clear if attachment of LHCII to PSI in state 2 is important in state transitions. Here we show that in the absence of a specific PSI subunit, PSI-H, LHCII cannot transfer energy to PSI, and state transitions are impaired. PMID- 11117753 TI - Governments prime basic nanotech research, applied activity yet to soar. PMID- 11117754 TI - Britain turns its attention to nanotech transfer. PMID- 11117755 TI - US industry starts to think big by acting small. PMID- 11117756 TI - Canada tries to limit nanotech brain drain. PMID- 11117757 TI - Japan sets sights on success in nanotechnology. PMID- 11117758 TI - The long term health consequences of polycystic ovary syndrome. AB - Women with polycystic ovary syndrome have both insulin resistance and beta cell dysfunction. Consequently, they are at increased risk of developing diabetes and cardiovascular disease. Women with polycystic ovary syndrome present to clinicians at a young age and as such offer a unique opportunity to identify insulin resistant patients at an early stage. This enables the modification of risk factors and diagnosis of diabetes before the onset of macro- and micro vascular symptoms. Increased emphasis should thus be placed on long term risk management and diabetic screening with advice on smoking, exercise and, if appropriate, weight loss. Where possible drugs that exacerbate insulin resistance should be avoided and consideration should be given to the use of insulin sensitising agents, particularly in the obese. PMID- 11117759 TI - The aetiology of gastroschisis. PMID- 11117760 TI - Prospective evaluation of three different models for the pre-operative diagnosis of ovarian cancer. AB - OBJECTIVE: To test the accuracy of the risk of malignancy index, the revised risk of malignancy index and Tailor's regression model to diagnose malignancy in women with known adnexal masses. DESIGN: Prospective collaborative study. SETTING: Gynaecology Assessment Unit, Department of Obstetrics and Gynaecology, King's College Hospital, London. SAMPLE: Sixty-one women with known adnexal masses were examined pre-operatively. Women were recruited from three South London hospitals. METHODS: The demographic, biochemical and sonographic data recorded for each patient included: age; menopausal status; CA125 levels; tumour volume; ultrasound characteristics; and Doppler blood flow analysis (peak and mean blood velocities, the pulsatility and resistance indices). The diagnosis of malignancy was made for each woman using all three models and the results compared with the final histopathological diagnosis. RESULTS: Thirty-eight women had benign tumours and 23 had ovarian cancer. Women with malignant tumours were significantly older than those with benign masses. There were also significant differences in CA125 levels, locularity, presence of papillary proliferations and ascites between the two groups. Tailor's regression model achieved a 43% sensitivity and 92% specificity in the diagnosis of malignancy. This compared with a 74% sensitivity and 92% specificity with the risk of malignancy model, and a 74% sensitivity and 89% specificity with the revised risk of malignancy model. CONCLUSION: When applied prospectively all three diagnostic models performed less accurately than originally reported, despite clinical signs of malignancy being present in many cases. It is likely that their accuracy would be even less in a population of women in whom there was a substantial clinical uncertainty. Intra-tumoral blood velocity and CA125 levels were the best individual parameters for discrimination between benign and malignant tumours. PMID- 11117761 TI - Pregnancy and delivery: a urodynamic viewpoint. AB - OBJECTIVE: The aims of this study were to establish prospectively the prevalence of objective bladder dysfunction before and after delivery by means of urodynamic investigations and to assess the effect of obstetric variables on bladder function. DESIGN: Prospective longitudinal study. Twin channel subtracted cystometry was performed in the standing and sitting position, with a cough stress test at the end of filling. The investigations were repeated three months postpartum. PARTICIPANTS: Two hundred and eighty-six nulliparae with singleton pregnancies who were delivered between April 1996 and November 1997 attended for antenatal assessment after 34 weeks of gestation and 161 who returned postpartum. SETTING: Department of Obstetrics and Gynaecology in a London teaching hospital. RESULTS: The mean urodynamic values both in pregnancy and postpartum lower than values defined in a non-pregnant population. The prevalence of genuine stress incontinence and detrusor instability were antenatally 9% and 8%, respectively, and postpartum 5% and 7%, respectively. Obstetric and neonatal factors were not related to urodynamic variables. CONCLUSIONS: Despite the reported high prevalence of urinary incontinence related to pregnancy and childbirth, neither pregnancy nor delivery resulted in any consistent effects on objective bladder function. Postpartum urodynamic measurements were not related to either obstetric or neonatal variables, but were dependent on antenatal values. PMID- 11117762 TI - Birth trauma: short and long term effects of forceps delivery compared with spontaneous delivery on various pelvic floor parameters. AB - OBJECTIVE: To compare the effects of forceps delivery and spontaneous delivery on pelvic floor functions in nulliparous women. DESIGN: A longitudinal prospective study with investigations during the first pregnancy, 10 weeks and 10 months after delivery. SETTING: Antenatal clinic in a teaching hospital. POPULATION: One hundred and seven patients aged 28 +/- 4 years, divided into those with forceps (n = 25) or spontaneous (n = 82) delivery. METHODS: Investigations with a questionnaire, clinical examination, assessment of bladder neck behaviour, urethral sphincter function, intra-vaginal/intra-anal pressures during pelvic floor contractions. RESULTS: The incidence of stress urinary incontinence was similar in both groups at 9 weeks (32% vs 21%, P = 0.3) and 10 months (20% vs 15%, P = 0.6) after delivery, as was the incidence of faecal incontinence (9 weeks: 8% vs 4%, P = 0.9; 10 months: 4% vs 5%, P = 1) and the decreased sexual response at 10 months (12% vs 18%, P = 0.6). Bladder neck behaviour, urethral sphincter function and intra-vaginal and intra-anal pressures were also similar in the two groups. However, 10 months after delivery, the incidence of a weak pelvic floor (20% vs 6%, P = 0.05) and the decrease in intra-anal pressure between the pre- and post-delivery values (-17 +/- 28 cm H2O vs 3 +/- 31 cm H2O, P = 0.04) were significantly greater in the forceps-delivered women. CONCLUSIONS: Forceps delivery is not responsible for a higher incidence of pelvic floor complaints or greater changes in bladder neck behaviour or urethral sphincter functions. However, patients with forceps delivery have a significantly greater decrease in intra-anal pressure and a greater incidence of a weak pelvic floor. PMID- 11117763 TI - A comparison of bladder neck movement and elevation after tension-free vaginal tape and colposuspension. AB - OBJECTIVE: To compare elevation and movement of the bladder neck after tension free vaginal tape and open Burch colposuspension, using transperineal ultrasound. DESIGN: Pospective, non-randomised study using pre- and post-operative transperineal ultrasound of the bladder neck. SETTING: Tertiary referral urogynaecology unit at a London teaching hospital. SAMPLE: Thirty consecutive women who underwent tension-free vaginal tape or colposuspension for primary genuine stress incontinence between March 1998 and June 1999. METHODS: Women underwent transperineal ultrasound of the bladder neck prior to and three to four weeks after surgery. MAIN OUTCOME MEASURES: Bladder neck elevation, angle and movement in relation to the pubic symphysis. RESULTS: For both tension-free vaginal tape and colposuspension the post-operative bladder neck angles at rest and valsalva were more acute than pre-operatively. The post-operative linear movement on valsalva was less than pre-operatively. For colposuspension the rotational movement on valsalva was significantly less post-operatively, but for tension-free vaginal tape there was only a trend towards less post-operative rotational movement. Post-operative angles and movement were significantly less for colposuspension. The resting bladder neck position was elevated significantly more by colposuspension. CONCLUSION: Both tension-free vaginal tape and colposuspension decrease bladder neck angles at rest and valsalva, linear movement on valsalva and elevate the bladder neck. The colposuspension causes significantly more change than the tension-free vaginal tape. This suggests the mechanism of continence for the tension-free vaginal tape is less dependent on bladder neck change than the colposuspension. PMID- 11117764 TI - Vault prolapse and rectocele: assessment of repair using sacrocolpopexy with mesh interposition. AB - OBJECTIVE: To assess the sacrocolpopexy with mesh interposition in women with pelvic organ prolapse. DESIGN: A prospective study. SETTING: Tertiary referral urogynaecology and pelvic floor reconstruction unit. POPULATION: Twenty-nine consecutive women with symptomatic vault prolapse and rectocele. MAIN OUTCOME MEASURES: Subjective and objective success rates and complications. RESULTS: The mean age was 57 years. The mean number of past prolapse operations was 2.6 which included two past sacrospinous ligament fixations and 17 past posterior repairs. The mean follow up was 14 months. There was an increase in constipation from 41% to 50%, a decrease in faecal soiling from 21% to 10%, and an increase in incomplete defecation from 24% to 36% . Dyspareunia decreased from 38% to 17%, and there was some improvement in the stress and urge incontinence. There was a significant reduction of vault prolapse and rectocele (P < 0.001). All women with Stage II and Stage III vault prolapse were corrected, with an increase in Stage I prolapse from 20% to 27%. All women with Stage II and Stage III rectocele were corrected with a decrease in Stage I prolapse from 36% to 7% . The only significant interoperative complication was a cystotomy. One mesh became infected post-operatively which required removal. CONCLUSIONS: Sacrocolpopexy and mesh interposition is a safe and reliable operation for the correction of vault prolapse and rectocele. A long term follow up is necessary to detect any late complications. PMID- 11117765 TI - Subtotal abdominal hysterectomy: a surgical advance or a backward step? AB - OBJECTIVE To review the short and medium term outcomes of subtotal abdominal hysterectomy. We also describe the management of cervical stump complications by vaginal trachelectomy or large loop excision of the transformation zone. DESIGN: Retrospective analysis. SETTING: Warwick General Hospital, Warwickshire, UK. SAMPLE: One hundred and fifty women underwent subtotal abdominal hysterectomy between 1993 and 1999. Five women had vaginal trachelectomy and another five had large loop excision of the transformation zone for complications relating to the cervical stump. RESULTS: The prevalence of intra-operative and early post operative complications was 4% and 7.3%, respectively. Twenty women (13.3%) had late complications, of whom 17 (11%) presented with symptoms directly related to the stump (two had also genuine stress incontinence). Three presented with genuine stress incontinence alone. The commonest problem was regular menstruation, which occurred in 12 women (8%). Ten of these women underwent vaginal trachelectomy or large loop excision of the transformation zone. None had intra-operative or post-operative complications. CONCLUSIONS: The high prevalences of cervical stump problems should be taken into account before a change in surgical procedure from total to subtotal hysterectomy is recommended. Further prospective studies with prolonged follow up are needed to evaluate the risks and benefits of retaining the cervix at hysterectomy. Total hysterectomy, preferably by the vaginal route, remains the procedure of choice for most women. Should a problem develop, vaginal trachelectomy or large loop excision of the transformation zone by an experienced surgeon are the best options for these women. PMID- 11117766 TI - Three methods for hysterectomy: a randomised, prospective study of short term outcome. AB - OBJECTIVE: To detect differences in clinical short term outcome between total abdominal hysterectomy, vaginal hysterectomy and laparoscopic assisted vaginal hysterectomy. DESIGN: Randomised controlled trial. SETTING: Department of Obstetrics and Gynaecology, Hospital of Helsingborg, Sweden. SAMPLE: One hundred twenty women scheduled for hysterectomy for various indications. METHODS: Randomisation into three treatment arms: total abdominal hysterectomy (n = 40); vaginal hysterectomy (n = 40) and laparoscopic assisted vaginal hysterectomy (n = 40). During traditional abdominal and vaginal surgery, laparoscopic assistance was kept to a minimum. Substantial number of cases needed volume-reducing manoeuvres due to uterine size. MAIN OUTCOME MEASURES: Duration of surgery, anaesthesia, time in hospital and recovery time. RESULTS: Mean duration (range) of surgery was significantly longer for laparoscopic assisted vaginal hysterectomy compared with vaginal hysterectomy and total abdominal hysterectomy, 102 min (50-175), 81 min (35-135) and 68 min (28-125), respectively. Mean stay in hospital and mean time to recovery was significantly longer for total abdominal hysterectomy compared with vaginal hysterectomy and laparoscopic assisted vaginal hysterectomy. The difference between vaginal hysterectomy and laparoscopic assisted vaginal hysterectomy was not significant. It was possible to remove uteri under 600 g with all three methods. Four laparoscopic assisted vaginal hysterectomies and one vaginal hysterectomy were converted to open surgery. Reoperation and blood transfusion were required after two vaginal hysterectomies and one laparoscopic assisted vaginal hysterectomy. One woman needed blood transfusion after total abdominal hysterectomy. CONCLUSIONS: Traditional vaginal hysterectomy proved to be feasible and the faster operative technique compared with vaginal hysterectomy with laparoscopic assistance. The abdominal technique was somewhat faster, but time spent in theatre was not significantly shorter. Abdominal hysterectomy required on average a longer hospital stay of one day and one additional week of convalescence compared with traditional vaginal hysterectomy. Vaginal hysterectomy should be a primary method for uterine removal. PMID- 11117767 TI - A randomised comparison and economic evaluation of laparoscopic-assisted hysterectomy and abdominal hysterectomy. AB - OBJECTIVES: To determine the safety, cost effectiveness and effect on quality of life of laparoscopic-assisted vaginal hysterectomy (LAVH) compared with total abdominal hysterectomy (TAH) in the management of benign gynaecological disease. DESIGN: Randomised controlled trial and economic evaluation. SETTING: Three hospitals in the West of Scotland. PARTICIPANTS: Two hundred women scheduled for an abdominal hysterectomy for benign gynaecological disease. MAIN OUTCOME MEASURES: Conversion rate of LAVH to TAH, complication rates, NHS resource use and costs, quality of life using EuroQol 5 D visual analogue scale, and achievement of milestones. RESULTS: The overall incidence of operative complications was 14% in the TAH group and 8% in the LAVH group, with an 8% conversion rate. Length of operation was significantly greater in the women having LAVH at 81 +/- 30 min vs 47 +/- 16 min (P < 0.001). There was no difference in analgesic requirements between the groups although there was a significantly shorter hospital stay for those having LAVH. The rate of post surgery recovery, satisfaction with operation and quality of life at four weeks post-operative were similar in the two groups of women. LAVH was significantly more expensive than TAH and remained more expensive for all but the most extreme scenario. CONCLUSIONS: This study demonstrates that despite the decreased length of hospital stay, LAVH is more expensive than TAH. In addition, recovery following operation and patient satisfaction were not affected by the route chosen. It is unlikely that LAVH represents an efficient use of NHS resources. PMID- 11117768 TI - The endometrial response to sequential and continuous combined oestrogen progestogen replacement therapy. AB - OBJECTIVES: 1. To determine the prevalence of endometrial hyperplasia in postmenopausal women taking standard proprietary regimens of sequential oestrogen/progestogen; 2. to determine the effects of nine months treatment with an oral continuous combined regimen of 2 mg 17beta-oestradiol and 1 mg norethisterone acetate (Kliofem [Kliogest outside the UK]; Novo Nordisk, Denmark) on endometrial histology in postmenopausal women. DESIGN: An open, prospective study in postmenopausal women. SETTING: Fifty-four menopause clinics in the UK. PARTICIPANTS: 2028 postmenopausal women: 1312 (Group A) taking sequential oestrogen-progestogen hormone replacement therapy (HRT), and 716 (Group B) not taking HRT, were recruited. In Group A, 388 women took preparations containing 10 days of progestogen, 921 had 12 days, and 3 had 13 days per cycle. METHODS: Endometrial aspiration biopsies were taken towards the end of a three-month run in period (Group A) or at study entry (Group B), before administration of the continuous combined HRT regimen. Biopsies were repeated at the end of the nine month treatment period. MAIN OUTCOME MEASURE: Endometrial histology. RESULTS: Initial endometrial biopsy data were available for 1106 women in Group A, who by the time of endometrial investigation had been taking HRT for a median duration of 2.56 years (5th to 95th centiles: 0.77 to 8.49 years). Data were available for 661 untreated women, who had no bleeding and had not taken HRT within the last year (Group B). Complex hyperplasia was found in 59 women (5.3%), and atypical hyperplasia in a further eight (0.7%) in Group A. In Group B there were no cases with complex hyperplasia, but one woman showed atypical hyperplasia (0.2%). At the end of the nine months of continuous combined therapy there was no case of hyperplasia among 1196 biopsies (upper 95% confidence limit of risk 0.31%) in women completing the study. Within this Group all of the women with complex hyperplasia arising during previous sequential HRT and who completed the study (n = 38) reverted to normal endometrial patterns. There was no case of endometrial carcinoma during the study. CONCLUSIONS: Despite taking standard regimens of sequential HRT containing at least 10 days of progestogen, there was a 5.3% prevalence of complex endometrial hyperplasia, and a 0.7% prevalence of atypical hyperplasia. However, continuous combined HRT (Kliofem) containing daily progestogen is not associated with an increased risk of hyperplasia and will convert the endometrium to normal in those with complex hyperplasia arising during previous sequential HRT. PMID- 11117770 TI - Pregnancy testing prior to sterilisation. AB - OBJECTIVE: To determine the incidence of positive pregnancy test on the day of laparoscopic sterilisation. DESIGN: Prospective longitudinal observational study. SETTING: Gynaecology unit in a UK teaching hospital. SAMPLE: Between 1 January 1997 and 31 December 1998, eight hundred and two consecutive women were admitted for laparoscopic sterilisation after assessment in the gynaecology clinic. On the day of planned surgery, all women had a pregnancy test performed on a urine sample taken that morning following overnight fasting, immediately prior to operation. MAIN OUTCOME MEASURES: A positive pregnancy test on the day of planned surgery. RESULTS: Of 802 women tested, 21 (2.6%) were pregnant. A careful medical history taken before surgery revealed evidence of amenorrhoea and menstrual irregularity in 17 of the pregnant women. Of the 21 pregnant women, 11 underwent termination of pregnancy, six continued the pregnancy, four had a miscarriage and one had an ectopic pregnancy. CONCLUSION: The routine practice of pregnancy testing on the day of laparoscopic sterilisation introduced in our hospital should continue to be part of a thorough clinical assessment before surgery. This may help to reduce the considerable level of existing litigation in a high risk area of gynaecological practice. PMID- 11117769 TI - Combined oral oestradiol valerate-norethisterone treatment over three years in postmenopausal women: correlation between oestrogen levels and bone mineral density sites. AB - OBJECTIVE: To compare trabecular and compact bone response and relationship to oestrogen status using continuous oestradiol valerate 2 mg and norethisterone 0.7 mg daily as hormone replacement and to determine the therapeutic range of 17 beta oestradiol. DESIGN: Open label trial. SETTING: Independent endocrine clinic SAMPLE: One hundred and thirty-one patients were compared at point of entry and at 36 months. METHODS: Postmenopausal women were assessed using a Lunar dual photon and single photon bone scanner, and bone mineral density of the lumbar spine, right hip and left forearm were annually correlated with 17 beta oestradiol and oestrone levels over three years. Total alkaline phosphatase was compared between improvers and decliners of bone mineral density. RESULTS: Significant improvement in bone mineral density (P < 0.0001) occurred at all sites except the left forearm, where bone loss was prevented. There was no correlation between oestrogen levels and bone mineral density improvements at hip sites. However, in the lumbar spine larger improvements in bone mineral density occurred in women with 17 beta-oestradiol levels > 185 pmol/L compared with those below, which were statistically significant for those with 17 beta-oestradiol levels > 248 pmol/L. Bone turnover, as quanitifed by total alkaline phosphatase measurements, was suppressed in most patients, but there were no differences in the mean alkaline phosphatase levels between the best improvers and worst decliners for lumbar spine bone mineral density. Improvers had an age mean of 5.21 years greater than decliners (P = 0.01) and a mean duration difference since the menopause of 5 1 years compared with decliners (P = 0.007). CONCLUSION: This combined continuous preparation of hormone replacement therapy improves not only trabecular bone but prevents compact bone loss, and the data suggest that the therapeutic range of 17 beta-oestradiol is between 200 pmol/L and 350 pmol/L. PMID- 11117771 TI - Risk factors and clinical manifestations of pre-eclampsia. AB - OBJECTIVE: To study associations between established risk factors for pre eclampsia and different clinical manifestations of the disease. DESIGN: A population-based, nested case-control study. SETTING: Information from 12,804 consecutive deliveries that took place over three years at a birth clinic, which alone serves a population of nearly 240,000 in Rogaland county, Norway. SUBJECTS: Cases of pre-eclampsia (n = 323) and healthy controls (n = 650) were selected. Pre-eclampsia was defined as increase in diastolic blood pressure (> or = 25 mmHg to > or = 90 mmHg) and proteinuria (> or = 1+ by dipstick testing) after 20 weeks of pregnancy. MAIN STUDY MEASURES: Parity, previous pre-eclampsia, blood pressure, maternal weight, and maternal smoking were included as study variables. Women with pre-eclampsia were grouped according to clinical manifestations of the disease (i.e. severity [mild, moderate or severe]) and time of onset (early or late gestation). Associations with the study factors were estimated as relative risks (odds ratio, OR). RESULTS: Both nulliparity and hypertension increased pre eclampsia risk, with no clear preference for any clinical subtype. High maternal weight was related to a higher risk of mild and moderate, but not severe, pre eclampsia. Previous pre-eclampsia strongly increased the risk for pre-eclampsia in the current pregnancy, and the risk of early onset disease was especially high (OR 42.4; 95% CI 11.9-151.6). Overall, smoking was associated with a reduced risk for pre-eclampsia (OR 0.6; 95% CI 0.4-0.9). However, no effect of smoking was observed in the early onset disease group and among women with repeated pre eclampsia. CONCLUSION: Nulliparity and hypertension increased the risk for each subgroup of pre-eclampsia, but high maternal weight, previous pre-eclampsia and smoking were not consistently associated with each clinical subtype. This observation may suggest that heterogeneous clinical manifestations of pre eclampsia may be preceded by different pathological mechanisms. PMID- 11117772 TI - Cardiac troponin I in pre-eclampsia and gestational hypertension. AB - OBJECTIVE: To investigate serum cardiac troponin I, a sensitive marker of cardiac myocyte damage, in normal pregnancy and pregnancies complicated by hypertension with and without significant proteinuria. DESIGN: Prospective cross sectional study. SETTING: University hospital delivery suite. SAMPLE: Serum samples obtained from women in normal pregnancy and in pregnancies complicated by hypertension with and without significant proteinuria. METHOD: Women with hypertension in pregnancy (at least two readings of systolic blood pressure > 140 mmHg and diastolic blood pressure > 90 mmHg) (n = 26) and normotensive women (n = 43) were included in the study. Serum cardiac troponin I was measured using Beckman Access immunoassay. MAIN OUTCOME MEASURE: Serum cardiac troponin I level in the pregnancies complicated by hypertension (with and without significant proteinuria) compared with the levels measured in normotensive women. RESULTS: The median serum cardiac troponin I level in pregnancies complicated by hypertension was 0.118 ng/mL (n = 26) which was significantly greater than that measured in samples obtained from normotensive women in pregnancy (0.03 ng/mL; n = 43) (P < 0.0001). There were higher median serum cardiac troponin I levels in hypertensive women with significant proteinuria (0.155 ng/mL; n = 6), compared with those without proteinuria (0.089 ng/mL; n = 20; P = 0.03). CONCLUSION: Serum cardiac troponin I is elevated in women with hypertensive disorders of pregnancy indicating some degree of cardiac myofibrillary damage in these disorders. PMID- 11117773 TI - Factors influencing postnatal liver function tests. AB - OBJECTIVE: To investigate liver function tests (LFTs) changes in the puerperium and the influence of specific obstetric events on these changes. DESIGN: A longitudinal observational study. SETTING: West Middlesex University Hospital, Twickenham. POPULATION: Ninety-four women with uncomplicated pregnancy who delivered at term. METHODS: Aspartate transaminase (AST), alanine transaminase (ALT), gamma glutamyl transferase (GGT), total bilirubin (Bilirubin) and alkaline phosphatase (ALP) were measured in early labour and on day 1, day 2, day 5 and day 10 postnatal. MAIN OUTCOME MEASURES: Peak enzyme concentration, time of peak enzyme concentration, the area under the curve for each enzyme and the rate of change of enzyme level from predelivery to peak concentration. RESULTS: All LFTs were affected by delivery (P < 0.001), increasing by 88% (0-500%) on day 2 or day 5 for AST, 147% (0-1140%) on day 5 for ALT and 63% (0-450%) on day 5 or day 10 for GGT. Multiple linear regression showed that caesarean section and opioid administration was associated with a faster rise in AST (P = 0.001, P = 0.033 respectively). The mean peak GGT concentration was 39% higher in women having caesarean section compared with vaginal delivery (P = 0.015). Univariate analysis showed that perineal trauma, use of Entonox, maternal age at delivery and breastfeeding also influenced LFT concentration significantly. CONCLUSION: Liver enzyme levels change significantly in the puerperium and are affected by common obstetric events, particularly caesarean section. This study aids clinical interpretation of postnatal LFTs in women recovering from liver-related illnesses, by facilitating the differentiation of physiological and pathological processes. PMID- 11117774 TI - A randomised controlled trial of vaginal clindamycin for early pregnancy bacterial vaginosis. AB - OBJECTIVE: To determine whether treatment of bacterial vaginosis (BV) with vaginal clindamycin affects pregnancy outcome. MATERIALS AND METHODS: Mothers with singleton pregnancies and without previous preterm delivery in 17 health centres in Oulu from March 1996 Until March 1998, in whom BV was diagnosed by Gram stain of a vaginal swab at the first antenatal visit (at the 12th gestational week) were randomised at Oulu University Hospital to have a one-week course of vaginal clindamycin, or placebo. A follow up sample of Gram stain was taken two weeks after randomisation and at the 30th gestational weeks. Pregnancy outcome data was obtained from hospital records. Primary outcome was preterm birth, and puerperal infectious morbidity the other outcome measure. RESULTS: During the study period 1956 women were screened, of whom 143 (7.3%) were BV- positive. One hundred and one were randomised. The total preterm birth rate of BV+ women randomised was 9.9% (10/101). Preterm birth occurred in 20.7% (6/29) vs 0% (0/26) according to whether BV persisted or not (P < 0.01). The preterm birth rate was 13.7% (7/51) in the clindamycin group vs 6.0% (3/50) in the placebo group (OR 2.5, 95% CI 0.6-10). BV was cured just after treatment in 17 out of 51 (33%) of the clindamycin- treated patients vs 17 out of 50 (34%) of the placebo- treated patients (OR 1.0, 95% CI 0.4-2.2). There was a difference in puerperal infectious morbidity in patients where BV persisted (31%, 9/29) compared with those in which BV did not persist (7.7%, 1/26) (OR 5.4, 95% CI 1.04-28). Infections were seen in 4/51 (8%) of the clindamycin treated vs 10/50 (20%) of the placebo treated cases, (OR 0.3, 95% CI 0.1-1.2). CONCLUSION: The prevalence of BV was lower than expected in this low risk population, but nevertheless it increased the risk of preterm birth and puerperal infectious morbidity, the risk being highest in cases where BV persisted during pregnancy. Vaginal clindamycin treatment for BV in the first trimester of pregnancy did not appear to reduce the risk of preterm birth or puerperal infections. PMID- 11117775 TI - Naegele's rule: a reappraisal. AB - The origin of Naegele's rule to calculate the expected date of confinement is reviewed. It is possible that the rule has been misinterpreted, resulting in an earlier estimated date of delivery with implications for induction of labour for post dates pregnancy. PMID- 11117776 TI - Anti-E in pregnancy. AB - Since the introduction of anti-Rhesus (Rh) D prophylaxis for RhD-negative women, other Rh and non-Rh red cell alloantibodies have become relatively more important and are now responsible for the greater proportion of haemolytic disease of the newborn. Anti-C and anti-E are the most commonly implicated non-D Rh antibodies in the pathogenesis of haemolytic disease of the newborn'. In 1977 Pepperell et al. reported the outcome of 44 women with anti-E. This is the only published series that investigates the implications of anti-E during pregnancy. The present report presents a retrospective study of the outcome of 122 pregnancies in which anti-E was the sole alloantibody detected. PMID- 11117777 TI - Triplet pregnancy with a coexisting complete hydatidiform mole of monospermic origin in a spontaneous conception. PMID- 11117778 TI - Splenic marginal zone lymphoma with or without plasmacytic differentiation. AB - We report a series of 31 cases of splenic marginal zone lymphomas with an enlarged spleen and a multimicronodular macroscopic pattern. Two groups, A and B, were distinguished based on the presence (A) or absence (B) of a lymphoplasmacytic component with monoclonal immunoglobulin expression in the cytoplasm. There were no differences between the groups as far as age, sex, spleen weight, and progression. The only difference was the presence in group A of a monoclonal serum component and autoimmune disorders, particularly autoimmune hemolytic anemia. In most cases in which a liver and/or bone marrow biopsy was performed, lymphomatous infiltration was detected. Seven cases had a seric monoclonal IgM of 5 g/L or more and liver or bone marrow infiltration, corresponding to the definition of Waldenstrom's macroglobulinemia. Lymphoma cells had a monocytoid, centrocytoid and, in group A, lymphoplasmacytic morphology. The lymphomatous cells were positive for CD20, CD45 RA, and bcl-2. They expressed IgD in 9 cases, partially in 6, and were negative for IgD in 9 of the 24 cases studied. Progression seems to be slow, with a long survival. Three patients presented with transformation into a large B-cell lymphoma, which was responsible for death in two patients. PMID- 11117779 TI - Keratin immunohistochemistry detects clinically significant metastases in bone marrow biopsy specimens in women with lobular breast carcinoma. AB - Some patients with breast cancer currently undergo bone marrow biopsy to make clinical decisions regarding therapy; however, lobular carcinoma can be difficult to detect in routine histologic sections. The authors reviewed retrospectively all available bone marrow biopsies from patients with lobular carcinoma diagnosed between January, 1, 1989, and September, 25, 1997, at the City of Hope National Medical Center to identify useful morphologic features and to determine the utility of pan-keratin immunohistochemical (IHC) staining. A total of 65 biopsies from 54 patients were reviewed. Thirteen of the 65 biopsies were classified initially as containing metastatic tumor based on histologic features alone. With the addition of keratin IHC, seven additional cases of metastatic disease were detected. Forty of the 54 patients received stem cell replacement or autologous bone marrow transplantation. Disease-free survival after high-dose chemotherapy with stem cell replacement or autologous bone marrow transplantation was stratified into three groups based on hematoxylin and eosin (H&E) staining and IHC results. Two-year disease-free survival was 33% for the H&E-/IHC+ group versus 90% for the H&E-/IHC- group (p = 0.005) among patients clinically free of disease at the time of stem cell replacement or autologous bone marrow transplantation. Two-year disease-free survival was 0% in the H&E+/IHC+ group (p = 0.04, compared with the H&E-/ IHC+ group). The authors conclude that routine morphologic examination without the aid of keratin IHC is unreliable in detecting clinically relevant metastatic lobular carcinoma in bone marrow biopsies. These findings suggest that pan-keratin immunostaining may be indicated on bone marrow biopsy specimens from lobular carcinoma patients if the biopsy appears histologically negative for metastatic tumor on H&E sections. PMID- 11117780 TI - Malignant melanoma with paradoxical maturation. AB - Typically, melanocytic nevi "mature" (i.e., exhibit a morphologic shift to smaller or spindle cells with progressive depth in the dermis). In contrast, most malignant melanomas (conventional MMs) lack maturation, and are composed of large pleomorphic cells throughout. The authors describe a series of melanomas with paradoxical maturation mimicking the pattern of nevi. Seventeen primary invasive melanomas with paradoxical maturation (IMPs), two epidermotropic metastatic melanomas with maturation (EMMMs), 13 compound nevi (CN), and 14 conventional MMs without apparent maturation were analyzed by histologic, cytomorphometric, and immunohistochemical techniques. With increasing dermal depth, both CN and IMPs had smaller nuclear and cellular areas, and decreased expression of Ki-67, glycoprotein (gp)100 (with HMB-45), and tyrosinase. IMPs had significant differences from conventional MMs; namely, smaller nuclear and cytoplasmic areas (deep), and decreased expression of Ki-67 (superficial and deep), gp100 (deep), and tyrosinase (deep). IMPs also had notable differences from CN: namely, larger nuclear and cellular areas, more confluence, more mitotic figures, increased Ki 67 and gp100 expression in both the superficial and deep portions, and more melanin (deep). The two EMMMs exhibited histologic and immunohistochemical features similar to the primary IMPs. IMP, because of its mimicry of nevus, can present a diagnostic hazard. The authors propose histologic, morphometric, and immunohistochemical criteria that facilitate recognition and accurate diagnosis of this unusual variant of melanoma. PMID- 11117781 TI - RET/PTC activation in hyalinizing trabecular tumors of the thyroid. AB - Hyalinizing trabecular tumor (HTT) of the thyroid is a neoplasm of follicular derivation with a histogenesis that is still the subject of debate. Morphologic affinities between HTT and papillary carcinoma, including nuclear pseudoinclusions and grooves, suggest that these tumors may be of similar origin. The authors investigated the relationship between these two types of tumors by assessing HTT for the presence of rearrangements of the proto-oncogene rearranged during transfection (RET) that, in thyroid tumors, are specific for papillary carcinoma. A series of 14 HTTs, including two cases associated with classic papillary carcinoma, was studied by means of immunohistochemistry and reverse transcription-polymerase chain reaction. Seven follicular adenomas with focal hyalinized trabecular areas served as control cases. Three of the 14 HTT cases under consideration displayed rearrangements of RET generating the RET/papillary thyroid carcinoma type 1 (PTC1) oncogene. In another case, RET expression was detected focally by immunohistochemistry alone. Finally, in one mixed HTT papillary carcinoma sample, RET/PTC1 expression was detected, but only in the papillary component. None of the control follicular adenomas contained rearrangements of RET/PTC. These findings demonstrate that a comparable percentage (28.6%) of HTTs and papillary carcinomas exhibit the same RET proto oncogene alterations. Thus, HTT may represent the "hyalinizing trabecular" variant of papillary carcinoma rather than a separate entity. PMID- 11117782 TI - Hyalinizing trabecular tumor of the thyroid: a variant of papillary carcinoma proved by molecular genetics. AB - Hyalinizing trabecular tumors of the thyroid are interesting but uncommon neoplasms. They have been classified as benign hyalinizing trabecular adenomas or malignant hyalinizing trabecular carcinomas. They share both epidemiologic and morphologic features with papillary carcinoma, and there has been much speculation about the relationship between these two entities. Because RET/PTC gene rearrangements are specific to papillary thyroid carcinoma, the authors examined the presence of RET/PTC-1, -2, and -3 in eight hyalinizing trabecular tumors using reverse transcription-polymerase chain reaction with Southern hybridization and immunohistochemistry. They detected the presence of a RET/PTC gene rearrangement in six of the eight hyalinizing trabecular tumors. This confirms the long-standing suspicion that hyalinizing trabecular tumors do indeed represent a morphologic variant of papillary carcinoma. PMID- 11117783 TI - Lennert's lymphoma: a variant of cytotoxic T-cell lymphoma? AB - We studied 10 cases of Lennert's lymphoma (lymphoepithelioid lymphoma) to evaluate the cellular origin of the neoplastic cells. There were six men and four women, aged 38 to 75 years (median, 56 yrs; mean, 59 yrs). The lymphoma cells tended to remain confined to lymph nodes, and extranodal involvement was rare. The mean overall survival was 42.2 months, which is relatively good compared with other peripheral T-cell lymphomas. Morphologically, the lymph node was occupied by small to large clusters of epithelioid cells interspersed with medium to large atypical lymphoid cells. In seven cases, large atypical lymphoid cells resembling Hodgkin's or Reed-Sternberg cells were observed. The phenotypes of these neoplastic cells were CD3+ CD4- CD8+ in five cases, CD3+ CD4+ CD8- in four cases, and CD3+ CD4- CD8- in one case. TIA-1 was positive by immunohistochemical staining in seven cases, whereas four cases were positive for granzyme B. Clonal rearrangement of the T-cell receptor gene was confirmed in all cases by either Southern blot hybridization or a polymerase chain reaction-based denature gradient gel electrophoresis method. Epstein-Barr virus was negative by in situ hybridization in all but one case. Lennert's lymphoma was formerly known as a CD4+ helper T-cell neoplasm. Our results suggest that, at least in some cases, the neoplastic cells are of cytotoxic T-cell origin. PMID- 11117784 TI - Should a Gleason score be assigned to a minute focus of carcinoma on prostate biopsy? AB - The grading system for prostate carcinoma devised by Gleason is a strong prognostic indicator. The primary and secondary patterns are combined to give a tumor score, referred to as Gleason score or sum. Gleason scores on biopsy correlate with the prostatectomy Gleason scores, and in combination with pretreatment serum prostate-specific antigen and digital rectal examination results, predict tumor stage and lymph node status. However, when only a minute focus of tumor is present on biopsy, the Gleason score is assigned by doubling the Gleason pattern. The goal of this study was to determine if a Gleason score assigned to a minimal focus of adenocarcinoma had predictive value. Paired biopsies and prostatectomy specimens from 963 cases of men with clinically localized prostate cancer were examined. Minimal tumor on biopsy was defined as less than 1 mm or 5% involvement of one biopsy core; excluded from this definition were biopsies where two Gleason patterns could be identified and/or tumor was seen on more than one biopsy core. Terms often used to describe these lesions include "single minute focus of carcinoma" or "adenocarcinoma, too small to give a Gleason grade." One hundred five cases (10.9%) met the above criteria for minimal carcinoma. The correlation of Gleason scores between biopsies and prostatectomy specimens overall was good with exact agreement for 57% of cases and a difference of +/-1 unit in 92% of cases. The correlation for the minimal tumors on biopsy and prostatectomy was slightly worse with exact agreement in 52.4% (55 of 105) and a difference of +/-1 unit in 87.6% (92 of 105). The majority of minimal tumors (83.8% or 88 of 105) were assigned a Gleason score of 6. A total of 31.8% of these 88 cases were upgraded and 5.7% were downgraded. Multivariate analysis on all cases looking for predictors of tumor stage found biopsy Gleason score, perineural invasion, pretreatment prostatic-specific antigen, and digital rectal examination all predicted higher tumor stage with odds ratios of 1.86 (95% confidence interval [CI], 1.53-2.27; p = 0.0001), 2.06 (95% CI, 1.43-2.95; p = 0.0001), 1.08 (95% CI, 1.05-1.11; p = 0.0001), and 1.41 (95% CI, 1.04-1.91; p = 0.0289), respectively. In a model restricted to the 105 cases with minimal carcinoma, pretreatment prostatic-specific antigen was the only independent predictor of higher tumor stage with an odds ratio of 1.15 (95% CI, 1.01-1.31; p = 0.0380); Gleason score was not found to significantly predict higher tumor stage (odds ratio, 1.156; p = 0.6680). The results of this study confirm that biopsy Gleason score in most cases predicts prostatectomy Gleason score and tumor stage. However, for cases with minimal tumor on biopsy, the assigned Gleason score did not predict tumor stage. To properly convey this uncertainty to clinicians, a cautionary note should accompany Gleason scores derived from a minimal focus of carcinoma. PMID- 11117785 TI - Histologic and immunohistologic findings and prognosis of 40 cases of gastric large B-cell lymphoma. AB - It has been considered that gastric large B cell lymphoma mainly consists of mucosa-associated lymphoid tissue lymphoma (MALToma) with large cell transformation. However, debate continues about the cell lineage. We analyzed 61 operated cases of gastric B cell lymphoma, mainly focusing on 40 cases of diffuse large cell lymphoma (DLCL). Immunohistologically, two cases were classified as CD10-positive follicular lymphoma, 19 cases were low-grade MALToma, 11 CD10 negative DLCL with a component of low-grade MALToma (high-grade MALToma), 12 CD10 positive DLCL, and 17 CD10-negative DLCL without MALToma (pure DLCL). Lymphoepithelial lesion (LEL) was found in all -cases of high-grade MALToma, and in eight of these its invasion was confined to the mucosa and submucosa. Expression of Bcl-6 was detected in two cases of high-grade MALToma. Only two cases of CD10-positive DLCL had large cell LEL, and seven cases showed tumor invasion beyond the submucosa. All 12 cases were positive for Bcl-6, and a delicate meshwork of CD35 (Ber-MAC-DRC)-positive follicular dendritic cells was detected in eight cases. Pure DLCL of all 17 cases reached the proper muscle layer or more, and expression of Bcl-6 was detected in 10 cases. For patients with pure DLCL, overall survival was significantly (p <0.05) worse than those of high-grade MALToma and CD10-positive DLCL by Kaplan-Meier and log-rank methods. Clinical staging and Bcl-6 expression were also good prognostic factors in patients with DLCL. Three groups of gastric DLCL each had unique histologic findings, immunohistologic characteristics, and prognosis. PMID- 11117786 TI - Pleomorphic lobular carcinoma: morphology, immunohistochemistry, and molecular analysis. AB - Infiltrating pleomorphic lobular carcinoma (PLC) is an aggressive variant of infiltrating lobular carcinoma. Recently, in situ changes identical to PLC (PLCIS) have been described. The role of prognostic markers and their correlation with therapeutics, clinical outcome, and genetic changes is not well established in PLC. The authors examined 38 cases of this entity to understand better this tumor's biology. Immunohistochemical (IHC) analysis was performed in 21 specimens for estrogen and progesterone steroid receptors, p53, Her 2 (p185), and GCDFP-15. Genomic deoxyribonucleic acid was obtained from microdissected tumor as well as normal control cells, and loss of heterozygosity was investigated at the ESR (16q24), p53 (TP53 17p), Her 2 (17q 11-12), and BRCA 1 (17q12-25) loci. In this series, the average patient age was 57.5 years (age range, 24-92 years). Twenty seven women were postmenopausal. Tumor size ranged from 1.2 to 25 cm. Six patients were a pathologic stage I; 19, stage II; 12, stage III; and one, stage IV. Histologically, multifocal nodular aggregates of discohesive pleomorphic tumor cells were seen interspersed in dense and fibrotic breast parenchyma. Twenty-nine percent of the specimens demonstrated associated signet ring cells. The remainder had dishesive, globoid, plasmacytoid cells with high-grade nuclear features. PLCIS was identified in 17 of 38 patients (45%), and lobular carcinoma in situ (LCIS) was noted in 8 patients (21%). IHC analysis showed estrogen immunoreactivity in 81%, progesterone in 67%, GCDFP-15 in 71%, and Her 2 in 81% (2+ to 3+ membranous staining) of specimens. Antibodies to p53 stained the tumor cell nuclei in 48% of the tumors. Loss of heterozygosity was identified in 52% of the specimens at the p53 locus, 18% at the ESR locus, 19% to 24% at the Her 2 loci, and 27% to 32% at the BRCA 1 locus. Follow-up was available in 19 patients and ranged from 12 months to 15 years (mean, 73 months). Seven patients had no evidence of disease at last examination (range, 1-15 years), three patients were alive with disease (range, 2-14 years), and nine patients were dead of disease (range, 2 months-9 years). Six patients had subsequent diagnoses of tumor in the contralateral breast. Analysis shows that PLC tends to appear in older postmenopausal women who present with locally advanced disease. PLCIS was found to be associated with PLC 45% of the time. The aggressive clinical course of patients with PLC is supported by tumor immunoreactivity with unfavorable markers Her 2 and p53. Overexpression of Her 2 in PLC may be therapeutically relevant, enabling the use of novel chemotherapeutic drugs like Herceptin. Interestingly, tumors that were Her 2 immunoreactive also maintained estrogen hormone immunoreactivity. PMID- 11117787 TI - Immunohistochemical detection of FLI-1 protein expression: a study of 132 round cell tumors with emphasis on CD99-positive mimics of Ewing's sarcoma/primitive neuroectodermal tumor. AB - The histologic and immunohistochemical differentiation of Ewing' s sarcoma/primitive neuroectodermal tumor (ES/PNET) from other small, blue, round cell tumors may be difficult. Despite initial promise, CD99 (MIC2) has not proven to be a specific marker. Approximately 90% of ES/PNET have a specific t(11; 22)(q24;q12) that results in fusion of the EWS and FLI-1 genes, and overexpression of FLI-1 protein. A recent study has shown immunohistochemical FLI 1 expression in five of seven of the ES/PNET cases tested. We evaluated FLI-1 expression in 132 well-characterized small, blue, round cell tumors. All tumors were immunostained for FLI-1 (1:40, Sc 356 polyclonal, Santa Cruz Biotechnology) using steam heat for epitope retrieval. Only nuclear staining was accepted as positive. Endothelial cells were strongly positive in all cases and served as an internal control. In many cases, a subset of lymphocytes also stained positive. No staining was seen in any other normal tissue. FLI-1 expression was seen in 29 of 41 (71%) ES/PNET, 7 of 8 (88%) lymphoblastic lymphomas, 0 of 8 poorly differentiated synovial sarcomas (PDSS), 0 of 32 rhabdomyosarcoma (RMS), 0 of 30 neuroblastomas, 0 of 8 esthesioneuroblastomas, 0 of 3 Wilms' tumors, 0 of 1 mesenchymal chondrosarcoma, and in 1 of 1 desmoplastic round cell tumor. This last case was known to have an EWS/WT-1 fusion. Although the EWS/FLI-1 fusion gene is specific for ES/PNET, FLI-1 protein expression is not. Significantly, the great majority of lymphoblastic lymphomas (also CD99-positive) are strongly FLI-1 positive. Immunohistochemical detection of FLI-1 may be valuable in confirming the diagnosis of ES/ PNET in cases in which molecular genetic evaluation is not feasible. FLI-1 protein expression is also helpful in distinguishing ES/PNET from other tumors that may be CD99-positive, such as PDSS and RMS. It is not surprising that some ES/ PNET are FLI-1-negative, because not all ES/PNET have the classic EWS/FLI-1, and some cases of ES/PNET may produce either low levels of protein or idiotypically different protein. PMID- 11117788 TI - Isochromosome 7q in adult Wilms' tumors: diagnostic and pathogenetic implications. AB - Wilms' tumors affecting adults are rare and are thought to have a worse prognosis than similar stage tumors in the pediatric population. To understand these tumors better, the authors reviewed their multi-institutional experience in a series of nine lesions diagnosed as Wilms' tumors in adults. In addition to histologic and immunohistochemical examination, they performed cytogenetic analysis and fluorescence in situ hybridization. On review, four cases were reclassified: two "blastema only" as Ewing's sarcoma/primitive neuroectodermal tumor and the other two as clear cell sarcoma of soft parts and sarcoma not otherwise specified (NOS). Of the remaining five cases, three exhibited biphasic histology and two were triphasic. In this group, there were three women and two men, and patient age ranged from 17 to 37 years (median age, 26 years). Tumor size was large and ranged from 10 to 31 cm (median tumor size, 12.5 cm). Histologically, the tumors showed the typical features of Wilms' tumors with varying amounts of blastema (n = 5), epithelium (n = 5), and stroma (n = 2). No tumors contained anaplasia, and persistent renal blastema was not identified in the non-neoplastic kidney in any specimen. All tumors were positive for cytokeratins (CK7, n = 3; pankeratin, n = 5), and one tumor was weakly positive for CD99 (0-13). Molecular analysis including dual color fluorescence in situ hybridization (all tumors), and cytogenetic analysis (n = 2) disclosed the presence of isochromosome 7q in three of five tumors whereas all tumors were diploid with respect to chromosome 12. Follow-up data ranged from 6 to 133 months (median follow-up, 82 months) with progression in only one patient who had stage IV disease with lymph node and lung metastases at presentation. The authors conclude that adult Wilms' tumor has been overdiagnosed. Most "blastema-only" tumors in adults are not Wilms' tumors, and in an adult, biphasic morphology should be the minimum criteria for their diagnosis. Using strict diagnostic criteria, adult Wilms' tumors have a relatively favorable prognosis. The characteristic findings of isochromosome 7q, lack of trisomy or tetrasomy for chromosome 12, and absence of persistent renal blastema suggest that the pathogenesis of Wilms' tumors in adults may be different than in the pediatric population. These genetic features may be helpful in distinguishing adult Wilms' tumors from other primary renal tumors. PMID- 11117789 TI - Pregnancy-like (pseudolactational) hyperplasia: a primary diagnosis in mammographically detected lesions of the breast and its relationship to cystic hypersecretory hyperplasia. AB - Pregnancy-like (pseudolactational) hyperplasia (PLH) has long been recognized as an incidental finding in breast biopsies performed for various clinically detected benign and malignant conditions. The histologic features of PLH have been well described, including some instances exhibiting cytologic and structural atypia. The presence of calcifications in these lesions was rarely mentioned and was considered to be of little consequence. More recently, however, calcifications in PLH have become the target of needle localization and needle core biopsies. The authors report 12 instances in which PLH was the primary diagnosis in biopsy specimens obtained for radiographic abnormalities, usually calcifications. Six of 12 procedures (50.0%) were performed for mammographically detected calcifications, four cases for a mass, one for an "abnormal mammogram," and one for galactorrhea. Calcifications were present in PLH in 10 biopsies, in benign terminal ducts in one specimen, and were not identified histologically in the remaining specimen. In most instances, calcifications associated with PLH had smooth round or lobulated contours and distinctive, internal, unevenly spaced laminations. Cystic hypersecretory hyperplasia (CHH) was present in five specimens. In four of the five specimens, CHH merged with PLH (PLH/CHH). Four of 12 specimens (33.3%) showed atypia within foci of PLH/CHH. PLH should be recognized as a primary diagnosis in breast biopsies for mammographically detected abnormalities such as calcifications. Some calcifications associated with PLH have a distinctive histologic appearance, and their recognition can aid in the diagnosis of PLH. Additional cases of PLH/CHH must be studied to ascertain the clinical significance, if any, of this previously undescribed entity. The precancerous significance of PLH/CHH and of PLH with atypia has not been determined. In most instances, surgical excision would be prudent if PLH/ CHH or PLH with atypia is present in a needle core biopsy specimen. PMID- 11117790 TI - The reliability of frozen-section diagnosis in the pathologic evaluation of Hirschsprung's disease. AB - There is a trend toward primary resection without prior colostomy in the surgical treatment of Hirschsprung's disease (HD) that renders evaluation of the initial pathologic specimen of utmost importance. To determine the accuracy of diagnostic methods for HD in routine practice, biopsies and resection specimens of all patients being evaluated for possible HD during a 3-year period were reviewed and correlated. Eighty patients underwent a total of 132 procedures related to the diagnosis or treatment of HD, including 93 intraoperative frozen-section (FS) evaluations. FS analysis was performed on 12 of the initial pathologic specimens, with concordance between the FS and permanent-section diagnoses in 67% of specimens. The concordance rate for all FS evaluations in patients with HD was 89%, which is substantially lower than the institutional rate for all patients. Pathologic diagnoses resulted in two patients receiving suboptimal surgical treatment. Both of these errors resulted from incorrect FS diagnoses on initial diagnostic specimens. In conclusion, there is a high rate of incorrect FS diagnoses in HD. It is obvious that if FS is being used as the initial diagnostic method, and primary reanastomosis is to be performed in the same procedure, an incorrect FS diagnosis has severe implications. FS as an initial diagnostic procedure for HD is not recommended. PMID- 11117791 TI - Esophageal lichen planus: case report and review of the literature. AB - Involvement of the esophagus by lichen planus is a rarely reported condition. The histologic features of esophageal lichen planus, which may differ from those of cutaneous disease, have only rarely been illustrated. We describe a 58-year-old woman with skin and oral lichen planus who presented with dysphagia and an esophageal stricture that were ultimately diagnosed as esophageal lichen planus. Multiple esophageal biopsies demonstrated a lichenoid, T cell-rich lymphocytic infiltrate, along with degeneration of the basal epithelium and Civatte bodies. Correct diagnosis of esophageal lichen planus is critical because of its prognostic and therapeutic distinction from other more common causes of esophagitis and stricture formation. PMID- 11117792 TI - Hyalinizing trabecular tumor of the thyroid: adenoma, carcinoma, or neoplasm of uncertain malignant potential? PMID- 11117842 TI - Fluorescence-guided resection of glioblastoma multiforme by using 5 aminolevulinic acid-induced porphyrins: a prospective study in 52 consecutive patients. AB - OBJECT: It has been established that 5-aminolevulinic acid (5-ALA) induces the accumulation of fluorescent porphyrins in glioblastoma multiforme (GBM), a phenomenon potentially exploitable to guide tumor resection. In this study the authors analyze the influence of fluorescence-guided resection on postoperative magnetic resonance (MR) imaging and survival in a series of patients who underwent surgery in the authors' department. METHODS: Fifty-two consecutive patients with GBM received oral doses of 5-ALA (20 mg/kg body weight) 3 hours before induction of anesthesia. Intraoperatively, tumor fluorescence was visualized using a modified operating microscope. Fluorescing tissue was removed whenever it was considered safely possible. Residual enhancement on early postoperative MR imaging was quantified and related to each patient's characteristics to determine which factors influenced resection. Survival was analyzed using the Kaplan-Meier method and multivariate analysis was performed in which the Karnofsky Performance Scale (KPS) score, residual fluorescence, patient age, and residual enhancement on MR images were considered. Intraoperatively, two fluorescence qualities were perceived: solid fluorescence generally reflected coalescent tumor, whereas vague fluorescence mostly corresponded to infiltrative tumor. Complete resection of contrast-enhancing tumor was accomplished in 33 patients (63%). Residual intraoperative tissue fluorescence left unresected for safety reasons predicted residual enhancement on MR images in 18 of the 19 remaining patients. Age, residual solid fluorescence, and absence of contrast enhancement in MR imaging were independent explanatory factors for survival, whereas the KPS score was significant only in univariate analysis. No perioperative deaths and one case of permanent morbidity were encountered. CONCLUSIONS: The observations in this study indicate the usefulness of 5-ALA induced tumor fluorescence for guiding tumor resection. The completeness of resection, as determined intraoperatively from residual tissue fluorescence, was related to postoperative MR imaging findings and to survival in patients suffering from GBM. PMID- 11117843 TI - Cerebral circulation and metabolism in the acute stage of subarachnoid hemorrhage. AB - OBJECT: The mechanism of reduction of cerebral circulation and metabolism in patients in the acute stage of aneurysmal subarachnoid hemorrhage (SAH) has not yet been fully clarified. The goal of this study was to elucidate this mechanism further. METHODS: The authors estimated cerebral blood flow (CBF), cerebral metabolic rate of oxygen (CMRO2), O2 extraction fraction (OEF), and cerebral blood volume (CBV) preoperatively in eight patients with aneurysmal SAH (one man and seven women, mean age 63.5 years) within 40 hours of onset by using positron emission tomography (PET). The patients' CBF, CMRO2, and CBF/CBV were significantly lower than those in normal control volunteers. However, OEF and CBV did not differ significantly from those in control volunteers. The significant decrease in CBF/CBV, which indicates reduced cerebral perfusion pressure, was believed to be caused by impaired cerebral circulation due to elevated intracranial pressure (ICP) after rupture of the aneurysm. In two of the eight patients, uncoupling between CBF and CMRO2 was shown, strongly suggesting the presence of cerebral ischemia. CONCLUSIONS: The initial reduction in CBF due to elevated ICP, followed by reduction in CMRO, at the time of aneurysm rupture may play a role in the disturbance of CBF and cerebral metabolism in the acute stage of aneurysmal SAH. PMID- 11117844 TI - Volume reduction in the caudate nucleus following stereotactic placement of lesions in the anterior cingulate cortex in humans: a morphometric magnetic resonance imaging study. AB - OBJECT: The goal of this study was to test hypotheses regarding changes in volume in subcortical structures following anterior cingulotomy. METHODS: Morphometric magnetic resonance (MR) imaging methods were used to assess volume reductions in subcortical regions following anterior cingulate lesioning in nine patients. Magnetic resonance imaging data obtained before and 9 +/- 6 months following anterior cingulotomy were subjected to segmentation and subcortical parcellation. Significant volume reductions were predicted and found bilaterally within the caudate nucleus, but not in the amygdala, thalamus, lenticular nuclei, or hippocampus. Subcortical parcellation revealed that the volume reduction in the caudate nucleus was principally referrable to the body, rather than the head. Furthermore, the magnitude of volume reduction in the caudate body was significantly correlated with total lesion volume. CONCLUSIONS: Taken together, these findings implicate significant connectivity between a region of anterior cingulate cortex (ACC) lesioned during cingulotomy and the caudate body. This unique data set complements published findings in nonhuman primates, and advances our knowledge regarding patterns of cortical-subcortical connectivity involving the ACC in humans. Moreover, these findings indicate changes distant from the site of anterior cingulotomy lesions that may play a role in the clinical response to this neurosurgical procedure. PMID- 11117845 TI - Differentiation of green fluorescent protein-labeled embryonic stem cell-derived neural precursor cells into Thy-1-positive neurons and glia after transplantation into adult rat striatum. AB - OBJECT: The aim of this investigation was to assess new information concerning the capacity of transplanted embryonic stem cell (ESC)-derived neuronal cells to migrate into host brain and to evaluate these cells as a possible source for cell replacement therapy in neurodegenerative disorders such as Parkinson's disease (PD). METHODS: The authors investigated the ability of ESC-derived neural precursor cells to migrate and differentiate in a host striatum by using a D3 derived ESC clone that was transfected stably with a chicken beta-actin cytomegalovirus enhancer-driven green fluorescent protein (GFP)-labeled construct. This procedure allowed easy monitoring of all transplanted cells because of the green fluorescent labeling of donor cells. This approach also afforded easy estimation of cell integration and simultaneous observation of the entire transplanted cell population in relation to immunocytochemically identified neuronal and glial differentiation. After selection of nestin-positive neural precursor cells in a synthetic medium, they were implanted into the striatum of male adult Wistar rats. Their integration was analyzed on morphological studies performed 3 days to 4 weeks posttransplantation. CONCLUSIONS: The investigators found that after transplantation, a subpopulation of GFP-labeled cells differentiated into various neural morphological types that were positive for the mouse-specific Thy-1 antigen, which is known be expressed on neurons, as well as being positive for the astroglial marker glial fibrillary acidic protein. Moreover, GFP-expressing cells that were negative for either of these markers remained close to the injection site, presumably representing other derivatives of the neural lineage. Together, these findings contribute to basic research regarding future transplantation strategies in neurodegenerative diseases such as PD. PMID- 11117846 TI - Subnecrotic stereotactic radiosurgery controlling epilepsy produced by kainic acid injection in rats. AB - OBJECT: Any analysis of the potential role of stereotactic radiosurgery for epilepsy requires the experimental study of its potential antiepileptogenic, behavioral, and histological effects. The authors hypothesized that radiosurgery performed using subnecrotic tissue doses would reduce or abolish epilepsy without causing demonstrable behavioral side effects. The kainic acid model in rats was chosen to test this hypothesis. METHODS: Chronic epilepsy was successfully created by stereotactic injection of kainic acid (8 microg) into the rat hippocampus. Epileptic rats were divided into three groups: high-dose radiosurgery (60 Gy, 16 animals), low-dose (30 Gy, 15 animals), and controls. After chronic epilepsy was confirmed by observation of the seizure pattern and by using electroencephalography (EEG), radiosurgery was performed on Day 10 postinjection. Serial seizure and behavior observation was supplemented by weekly EEG sessions performed for the next 11 weeks. To detect behavioral deficits, the Morris water maze test was performed during Week 12 to study spatial learning and memory. Tasks involved a hidden platform, a visible platform, and a probe trial. After radiosurgery, the incidence of observed and EEG-defined seizures was markedly reduced in rats from either radiosurgically treated group. A significant reduction was noted after high-dose (60 Gy) radiosurgery in Weeks 5 to 9 (p < 0.003). After low-dose (30 Gy) radiosurgery, a significant reduction was found after 7 to 9 weeks (p < 0.04). During the task involving the hidden platform, kainic acid-injected rats displayed significantly prolonged latencies compared with those of control animals (p < 0.05). Hippocampal radiosurgery did not worsen this performance. The probe trial showed that kainic acid-injected rats that did not undergo radiosurgery spent significantly less time than control rats in the target quadrant (p = 0.03). Rats that had undergone radiosurgery displayed no difference compared with control rats and demonstrated better performance than rats that received kainic acid alone (p = 0.04). Radiosurgery caused no adverse histological effects. CONCLUSIONS: In a rat model, radiosurgery performed with subnecrotic tissue doses controlled epilepsy without causing subsequent behavioral impairment. PMID- 11117847 TI - Effects of mitogen-activated protein kinase inhibitors on cerebral vasospasm in a double-hemorrhage model in dogs. AB - OBJECT: Mitogen-activated protein kinase (MAPK) may be involved in the pathogenesis of cerebral vasospasm after subarachnoid hemorrhage. This study was conducted to investigate the ability of the MAPK inhibitors PD-98059 and U-0126 to reverse vasospasm in a double-hemorrhage model in dogs. METHODS: Twenty-two adult mongrel dogs of either sex, each weighing 18 to 24 kg, were divided randomly into four groups: control SAH (four dogs), vehicle- (dimethyl sulfoxide, six dogs), PD-98059- (six dogs), and U-0126-treated groups (six dogs). The double hemorrhage model was created by an autologous blood injection into the cisterna magna on Days 0 and 2. An intracisternal injection of MAPK inhibitors was administered once per day on Days 3 through 6. Cerebral angiography was performed on Days 0 and 7 before the animals were killed. Western blot analysis was used to study the effects of hemorrhage and drug treatment on the MAPK immunoprecipitation. Severe vasospasm developed in the dogs in the control and vehicle-treated groups (basilar artery [BA] diameter reduction 46.6 +/- 5.5% and 49.3 +/- 4.6%, respectively). In the PD-98059-treated group, most of the dogs developed mild vasospasm (18.9 +/- 6.2%). In the U-0126-treated group, severe vasospasm was observed despite treatment (39.6 +/- 6.4%). The PD-98059 but not the U-0126 abolished MAPK immunoprecipitation in the spastic BAs. However, treatment with either PD-98059 or U-0126 improved the clinical scores of the dogs. CONCLUSIONS: The present study is the first in which the effects of MAPK inhibitors on vasospasm have been investigated in vivo. The authors demonstrate that MAPK may play a role in vasospasm and that PD-98059 is a potential candidate for the treatment of cerebral vasospasm. PMID- 11117848 TI - Effects of potassium channel inhibitors on the relaxation induced by the nitric oxide donor diethylamine nitric oxide in isolated human cerebral arteries. AB - OBJECT: The goal of this study was to investigate whether K+ channels are involved in nitric oxide (NO)-induced relaxation of isolated human cerebral arteries. METHODS: Successive concentration-response curves relating to the use of the NO donor diethylamine NO (DEA/NO) were established in the absence and presence of different K+ channel inhibitors after mounting human cerebral arteries onto a wire myograph. The arteries were obtained from macroscopically intact tissue that had been removed during brain tumor operations. A high K+ concentration partially inhibited the relaxant effects of DEA/NO. Different K+ channel inhibitors (tetraethylammonium [TEA], 10(-3) M; charybdotoxin, 10(-7) M; glibenclamide, 10(-6) M; 4-aminopyridine [4-AP], 10(-3) M; BaCl2, 5 x 10(-5) M; and apamin, 10(-6) M) alone failed to affect the responses to DEA/NO. However, a combination of TEA, glibenclamide, 4-AP, and BaCl2 partially blocked the relaxant effects of DEA/NO. In addition, the effects of DEA/NO were inhibited by the thromboxane A2 analog U46619 (3 x 10(-7) M). CONCLUSIONS: Inhibitors of the large conductance or small-conductance Ca++-activated K+ channels, the adenosine triphosphate-sensitive K+ channels, and the delayed-rectifier or inward-rectifier K+ channels failed to alter the effects of DEA/NO when only one K+ channel blocker was used. However, a regimen of a combination of K+ channel blockers that possess selectivity for different channels demonstrated that different K+ channel types are involved; these channels may function in a redundant manner and compensate for each other. Selective thromboxane A2 agonists are capable of inhibiting the relaxant response to the NO donor. PMID- 11117849 TI - Angiographically verified progression of moyamoya disease in an adult. Case report. AB - The authors present the case of a 37-year-old man with definite moyamoya disease in whom angiographic findings drastically changed. The patient presented with left hemiparesis due to lacunar infarction. Angiography initially disclosed a narrow right carotid artery (CA) siphon and severe stenosis of the horizontal segment of the left middle cerebral artery. Four years later, the patient experienced right-central facial paresis, which developed because of a small putaminal hematoma. Angiography results demonstrated occlusion of the internal CA siphons bilaterally, with moyamoya vessels. It therefore appears that in some adults, moyamoya disease is accompanied by very progressive vascular changes. PMID- 11117850 TI - Development of a cerebral arteriovenous malformation documented in an adult by serial angiography. Case report. AB - This 61-year-old man with a right-sided tentorial dural arteriovenous fistula (DAVF) was initially treated with staged stereotactic radiosurgery and transarterial embolization. Results of follow-up cerebral angiography performed 4 years later demonstrated complete obliteration of the dAVF and development of a previously undetected cerebellar arteriovenous malformation (AVM). The newly diagnosed AVM was treated with repeated stereotactic radiosurgery. This represents the first reported case of the development of a cerebral AVM documented in an adult by serial angiography. PMID- 11117851 TI - De novo formation and rupture of an azygos pericallosal artery aneurysm. Case report. AB - An azygos pericallosal artery (APCA) aneurysm is a rare anomaly that is closely associated with saccular aneurysms. This is the earliest report to document de novo formation and rupture of an aneurysm at the bifurcation of an unpaired pericallosal trunk. The authors report the case of a woman who presented at the age of 52 years with subarachnoid hemorrhage (SAH) from the rupture of a newly formed APCA bifurcation aneurysm, 7 years after she had undergone surgery to clip a ruptured anterior cerebral artery aneurysm. De novo formation of aneurysms after SAH rarely occurs and certain risk factors like multiple and familial aneurysms, arterial hypertension, or smoking have been postulated. Late follow-up examination with angiography to detect de novo aneurysms should be considered in patients with this vascular anomaly after SAH. PMID- 11117852 TI - Paraparesis induced by inflammatory contents of a pneumonectomy cavity. Case report. AB - The authors report on a patient who developed acute-onset paraparesis after underoing a thoracotomy 40 years earlier for a carcinoid adenoma. No infectious or neoplastic origin could be found to explain the patient's current clinical course and radiographic findings. The postoperative events in this case are discussed, as well as the literature regarding postthoracotomy complications. PMID- 11117853 TI - Traumatically induced lymphangioma of the ulnar nerve. Case report. AB - Lymphangiomas, benign hamartomatous lesions involving lymphatic tissue, result from a failure of lymphatic channels to communicate with the venous system or normal lymphatic channels. The authors describe a case in which a lymphangioma arising within the ulnar nerve developed after trauma to the same area. This is the second reported case of a lymphangioma that originated from a peripheral nerve and the first case in which the lesion was associated with trauma. The authors propose that a lymphangioma involving the peripheral nerve may be the result of trauma. PMID- 11117854 TI - Experimental evaluation of the Spiegelberg intracranial pressure and intracranial compliance monitor. Technical note. AB - The goal of this study was to compare the Spiegelberg intraventricular intracranial pressure (ICP)/intracranial compliance monitoring device, which features an air-pouch balloon catheter, with existing gold-standard methods of measuring ICP and intracranial compliance. A Spiegelberg intraventricular catheter, a standard intraventricular catheter, and a Codman intraparenchymal ICP microsensor were placed in five sheep, which previously had been given anesthetic and paralytic agents, to allow comparative measurement of ICP at incremental levels (range 5-50 mm Hg). Intracranial pressure measured using the Spiegelberg intraventricular air-pouch balloon catheter displayed a linear correlation with ICP measured using the standard intraventricular fluid-filled catheter (r2 = 0.9846, p < 0.001; average bias -0.74 mm Hg), as well as with ICP measured using the Codman intraparenchymal strain-gauge sensor (r2 = 0.9778, p < 0.001; average bias 0.01 mm Hg). Automated measurements of intraventricular compliance obtained using the Spiegelberg compliance device were compared with compliance measurements that were made using the gold-standard manual cerebrospinal fluid bolus injection technique at ICPs ranging from 5 to 50 mm Hg, and a linear correlation was demonstrated between the two methods (r2 = 0.7752, p < 0.001; average bias -0.019 ml/mm Hg). The Spiegelberg air-pouch ICP/compliance monitor provides ICP and compliance data that are very similar to those obtained using both gold-standard methods and an intraparenchymal ICP monitor over a range of pathophysiological ICPs. The automated closed Spiegelberg system offers practical advantages for the measurement of intraventricular compliance. Assessment of the clinical utility and robustness of the Spiegelberg system, together with the development of an intraparenchymal device, would enhance the clinical utility of automated compliance measurement and expand the range of its applications. PMID- 11117855 TI - Intratumoral injection of plastic adhesive material for removal of cavernous sinus hemangioma. Technical note. AB - The authors present a case in which a cavernous sinus (CS) hemangioma was totally removed following intratumoral injection of a plastic fixation material. This unique method is extremely useful for the removal of CS hemangiomas, which often feature massive intraoperative bleeding as an unsolved problem. PMID- 11117856 TI - Recurrent arteriovenous malformation in an adult. Case illustration. PMID- 11117857 TI - Thalamic venous angiomas draining into a vein of Galen varix. Case illustration. PMID- 11117858 TI - Recurrent artery of Heubner: Otto Heubner's description of the artery and his influence on pediatrics in Germany. AB - Although the recurrent artery of Heubner is one of the best known cerebral arteries, little has been written in the neurosurgical or anatomical literature about its discovery. The artery is of primary importance to cerebrovascular surgeons, who identify it during clipping of anterior communicating artery aneurysms. Johann Otto Leonhardt Heubner (1843-1926), who described this artery in 1872, is better known as the father of German pediatrics. He was appointed to the first professorship in Germany exclusively devoted to pediatrics at the Charite Children's Clinic of Berlin University. Although he initially studied internal medicine in Leipzig under Carl Reinhold August Wunderlich and Ernst Leberecht Wagner, his early research involved anatomical studies of the circulation of the brain, from which he described syphilitic endarteritis (Heubner's disease). Finding morphological studies inconclusive, he turned to more physiological experiments. Together with the physiologist Max Rubner, Heubner performed important studies on energy metabolism in infancy, creating the notion of the nutrition quotient. In this article the authors review Heubner's life and scientific discoveries. PMID- 11117859 TI - Subthalamic nucleus. PMID- 11117860 TI - Nasal speculum. PMID- 11117862 TI - Ventriculostomy. PMID- 11117861 TI - Growth hormone-releasing peptide and growth hormone secretion. PMID- 11117863 TI - Quality of life in patients with glioblastoma multiforme participating in a randomized study of brachytherapy as a boost treatment. AB - OBJECT: Until recently the assessment of outcome in patients treated for glioma has emphasized length of survival with the evaluation of quality of life (QOL) limited to unidimensional, mostly physical, measures. The authors report the multidimensional assessment of QOL as part of a randomized clinical trial of brachytherapy as a boost in the initial treatment of patients with glioblastoma multiforme. METHODS: A questionnaire previously developed by the senior authors and psychometrically validated was completed by patients on randomized entry into the study and at follow-up review every 3 months thereafter. The questionnaire was presented in a linear-analog self-assessment format. Karnofsky Performance Scale (KPS) scores were also recorded on each occasion. No differences were found between patients in either arm of the study (conventional radiation therapy consisting of 50 Gy in 25 fractions or conventional radiation plus a brachytherapy boost of a minimum peripheral tumor dose of 60 Gy) in KPS and QOL scores during the 1st year of follow-up review. However, there was a statistically significant deterioration in patients' overall KPS scores during the 1st year of follow up compared with baseline scores. Of QOL items evaluated, statistically significant deteriorations were found in self care, speech, and concentration, and on subscale analyses, cognitive functioning and physical experience (symptoms) deteriorated significantly during the 1st year of follow up, compared with baseline values. The correlation between QOL and KPS scores was low. CONCLUSIONS: Future studies in patients harboring malignant gliomas must incorporate measures assessing QOL because traditional measures focusing on physical or neurological functioning give an incomplete assessment of the patient's experience. PMID- 11117865 TI - The impact of age and sex on the incidence of glial tumors in New York state from 1976 to 1995. AB - OBJECT: In this study the authors describe secular trends in the incidence of three glial tumors--glioblastoma multiforme (GBM), astrocytoma not otherwise specified (ANOS), and anaplastic astrocytoma (AA)--in New York state from 1976 through 1995. They also describe the effect of age and sex on the relative risk (RR) for these tumors, specifically GBM. METHODS: Crude, age-, and sex-specific incidence rates were calculated for each tumor type from 1976 to 1995 by using data from the New York State Cancer Registry. Age-adjusted incidence rates were calculated by the direct standardization procedure, in which the 1970 United States Census Population Standard Million is used. The RR of GBM for the female population was calculated and plotted. Statistical comparisons were made using Pearson's correlation coefficient and regression analysis with the coefficient of variation. CONCLUSIONS: The age-adjusted incidence of these three glial tumors increased during the study period. Increases in age-specific incidence of GBM were primarily limited to patients 60 years of age or older. The reasons for these increases cannot be fully explained with the data. Those in the female population had a lower risk of developing these tumors than those in the male. For GBM, the protective effect of sex was first evident at the approximate age of menarche, was greatest at the approximate age of menopause, and decreased in postmenopausal age strata. The overall protective effect of female sex and the described trend in RR for GBM in the female population suggests that sex hormones and/or genetic differences between males and females may play a role in the pathogenesis of this tumor. PMID- 11117864 TI - Survival of patients with synchronous brain metastases: an epidemiological study in southeastern Michigan. AB - OBJECT: It has been suggested that synchronous brain metastases (that is, those occurring within 2 months of primary cancer diagnosis) are associated with a shorter survival time compared with metachronous lesions (those occurring more than 2 months after primary cancer diagnosis). In this study the authors used data obtained from the National Cancer Institute's Surveillance, Epidemiology, and End Results program to determine the incidence of synchronous brain metastases and length of survival of patients in a defined population of southeastern Michigan residents. METHODS: Data obtained in 2682 patients with synchronous brain metastases treated between 1973 and 1995 were reviewed. Study criteria included patients in whom at least one brain metastasis was diagnosed within 2 months of the diagnosis of primary cancer and those with an unknown primary source. The incidence per 100,000 population increased fivefold, from 0.69 in 1973 to 3.83 in 1995. The most frequent site for the primary cancer was the lung (75.4%). The second largest group (10.7%) consisted of patients in whom the primary site was unknown. The median length of survival was 3.2 months. There was no significant difference in the median survival of patients with primary lung/bronchus and those with an unknown primary site (3.3 months and 3.2 months, respectively). CONCLUSIONS: Patients who present with synchronous lesions have a poor prognosis, and the predominant cause of death, in more than 90% of cases, is related to systemic disease; however, despite poor median survival times, certain patients will experience prolonged survival. PMID- 11117866 TI - Primary extradural meningiomas: a report on nine cases and review of the literature from the era of computerized tomography scanning. AB - OBJECT: Primary meningiomas arising outside the intracranial compartment (primary extradural meningiomas [PEMs]) are rare tumors. To develop a better understanding of these tumors and to establish a comprehensive classification scheme for them, the authors analyzed a series of patients treated at the M. D. Anderson Cancer Center (MDACC) and reviewed all cases reported in the English-language literature since the inception of the use of computerized tomography (CT) scanning. METHODS: Clinical records, results of radiographic studies, and histological slides were reviewed for all cases of PEM at MDACC. Demographic features, symptoms, tumor location, histological grade, and patient outcome were assessed in all cases. A comprehensive literature search identified 168 PEMs in 142 patients reported during the CT era. These reports were also analyzed for common features. Tumors for both data sets were classified as purely extracalvarial (Type I), purely calvarial (Type II), and calvarial with extracalvarial extension (Type III). Type II and Type III tumors were further categorized as convexity (C) or skull base (B) lesions. The incidence of PEMs at MDACC was 1.6%, which was consistent with the rate reported in the literature. In both data sets, the male/female ratio was nearly 1:1. The most common presenting symptom was a gradually expanding mass. The age of patients at diagnosis of PEM was bimodal, peaking during the second decade and during the fifth to seventh decades. In all MDACC cases and in 90% of those reported in the literature the PEMs were located in the head and neck. The majority of tumors originated in the skull (70%). In the MDACC series and in the literature review, the majority (67% and 89%, respectively) of tumors were histologically benign. Although fewer PEMs were malignant or atypical (33% at MDACC and 11% in the literature), their incidence was higher than that observed for primary intracranial meningiomas. Distant metastasis was not a common feature reported for patients with PEMs (6% in the literature). Outcome data were available in 96 of the cases culled from the CT-era literature. The combination of the MDACC data and the data obtained from the literature demonstrated that patients with benign Type IIB or Type IIIB lesions were more likely to experience recurrence than patients with benign Type IIC or Type IIIC tumors (26% compared with 0%, p < 0.05). The more aggressive atypical and malignant tumors were associated with a statistically significant higher death rate (29%) relative to benign tumors (4.8% death rate, p < 0.004). CONCLUSIONS: Defining a tumor as a PEM is dependent on the tumor's relation to the dura mater and the extent and direction of its growth. Classification of PEMs as calvarial or extracalvarial and as convexity or skull base lesions correlates well with clinical outcome. PMID- 11117867 TI - Combined magnetic resonance imaging- and positron emission tomography-guided stereotactic biopsy in brainstem mass lesions: diagnostic yield in a series of 30 patients. AB - OBJECT: In the management of brainstem lesions, the place of stereotactic biopsy sampling remains debatable. The authors compared the results of magnetic resonance (MR) imaging, positron emission tomography (PET) scanning, and histological studies obtained in 30 patients who underwent MR imaging- and PET guided stereotactic biopsy procedures for a brainstem mass lesion. METHODS: Between July 1991 and December 1998, 30 patients harboring brainstem mass lesions underwent a stereotactic procedure in which combined MR imaging and PET scanning guidance were used. Positron emission tomography scanning was performed using [18F]fluorodeoxyglucose in 16 patients, methionine in two patients, and both tracers in 12 patients. Definite diagnosis was established on histological examination of the biopsy samples. Interpretation of MR imaging findings only or PET findings only was in agreement with the histological diagnosis in 63% and 73% of cases, respectively. Magnetic resonance imaging and PET findings were concordant in 19 of the 30 cases; in those cases, imaging data correlated with histological findings in 79%. Treatment based on information derived from MR imaging was concordant with therapy based on histological findings in only 17 patients (57%). Combining MR imaging and PET scanning data, the concordance between the neuroimaging-based treatment and treatments based on histological findings increased to 19 patients (63%). In seven patients who underwent biopsy procedures with one PET-defined and one MR imaging-defined trajectory, at histological examination the PET-guided samples were more representative of the tumor's nature and grade than the MR imaging-guided samples in four cases (57%). In 18 patients PET scanning was used to define a biopsy target and provided a diagnostic yield in 100% of the cases. CONCLUSIONS: Although the use of combined PET and MR imaging improves radiological interpretation of a mass lesion in the brainstem, it does not accurately replace histological diagnosis that is provided by a stereotactically obtained biopsy sample. Combining information provided by MR imaging and PET scanning in stereotactic conditions improves the accuracy of targeting and the diagnostic yield of the biopsy sample; an MR imaging- and PET guided stereotactic biopsy procedure is a safe and efficient modality for the management of mass lesions of the brainstem. PMID- 11117868 TI - Primary intracerebral and aneurysmal subarachnoid hemorrhage in Izumo City, Japan. Part I: incidence and seasonal and diurnal variations. AB - OBJECT: The purpose of this community-based study was first to estimate the incidence rates of primary intracerebral hemorrhage (ICH) and aneurysmal subarachnoid hemorrhage (SAH) in Izumo City, Japan, and second to investigate whether there were seasonal and diurnal periodicities in their onset. METHODS: During 1991 through 1996, 267 patients with primary ICH and 123 with aneurysmal SAH were treated in Izumo City. The crude and the age- and sex-adjusted annual incidence rates per 100,000 population for all ages were 52 and 48 for ICH and 24 and 23 for SAH, respectively. These incidence rates were higher than those previously published for any other geographical region. The incidence rates of both ICH and SAH increased almost linearly with age. For ICH, a significant seasonal pattern was observed in men and in patients younger than 65 years, with a peak in winter and a trough in summer. However, no significant seasonal fluctuation was found in women or in individuals aged 65 years or older. There was no significant seasonal periodicity for SAH, even when patients were analyzed according to sex and age. Diurnal variations in the onset of both ICH and SAH were significant (except in men with SAH), with a nadir between midnight and 6:00 a.m. CONCLUSIONS: The actual incidence rates of both primary ICH and aneurysmal SAH seem to be much higher than have been reported so far. In addition, the data indicate the existence of seasonal periodicity for men and younger patients with ICH, and that the risk of both ICH and SAH is lower during nighttime. PMID- 11117869 TI - Primary intracerebral and aneurysmal subarachnoid hemorrhage in Izumo City, Japan. Part II: management and surgical outcome. AB - OBJECT: The purpose of this study was to assess the overall management and surgical outcome of primary intracerebral hemorrhage (ICH) and aneurysmal subarachnoid hemorrhage (SAH) among the 85,000 residents of Izumo City, Japan. METHODS: During 1991 through 1996, 267 patients with ICH and 123 with SAH were treated in Izumo. Of the 267 patients with ICH, 25 underwent hematoma removal by open craniotomy or suboccipital craniectomy and 34 underwent stereotactic evacuation of the hematoma, whereas aneurysm clipping was performed in 71 of the 123 patients with SAH; operability rates were thus 22% for ICH and 58% for SAH (p < 0.0001). The overall 30-day survival rates were 86% for ICH and 66% for SAH (p < 0.0001) and the 2-year survival rates were 73% and 62% (p = 0.0207), respectively. In patients who underwent surgery, 30-day and 2-year survival rates were 93% for ICH and 100% for SAH (p = 0.0262), and 75% for ICH and 97% for SAH (p = 0.0002), respectively. In patients with ICH, the most important predictors of 30-day case-fatality rates were the volume of the hematoma, the Glasgow Coma Scale (GCS) score, rebleeding, and midline shifting, whereas those for 2-year survival were the GCS score, age, rebleeding, and hematoma volume. In patients with SAH, the most important determinants of 30-day case-fatality rates were the GCS score and age, whereas only the GCS score had a significant impact on 2-year survival. CONCLUSIONS: The overall survival rates for patients with ICH or SAH in Izumo were more favorable than those in previously published epidemiological studies. However, despite improved surgical results, the overall management of ICH and SAH still produced an unsatisfactory outcome, mainly because of primary brain damage. PMID- 11117871 TI - Dynamic nature of cavernous malformations: a prospective magnetic resonance imaging study with volumetric analysis. AB - OBJECT: Although cavernous malformations (CMs) are not detected in angiographic studies, they have a characteristic appearance on magnetic resonance (MR) images. A number of reports published in the last decade have focused on the behavior of these lesions within the clinical environment. However, little has been published about the evolution of CMs over time, as observed in imaging studies. To understand imaging-documented changes in CMs over time, we analyzed MR images of 114 cavernous malformations in 68 patients who were followed prospectively. METHODS: For each CM the location, volume, and MR imaging signal characteristics were recorded. Volume data were available for 107 lesions from initial images. The mean volume of these 107 CMs was 2779 mm3. The lesions ranged in size from 0.5 to 46,533 mm3 (46.5 cm3). Volume data from a second set of images were available for 76 CMs (mean interval from first imaging session 26 months), and from a third set of images for 24 lesions (mean interval from second imaging session 18 months). Over the first follow-up interval, the mean volume change was -991 mm3 (a decrease of approximately 1 cm3) and over the second interval the mean volume change was -642 mm3. Although these mean volume changes appear modest, volume changes in single lesions during follow-up intervals were more dramatic, with decreases as large as 45,629 mm3 (45.6 cm3) and increases as large as 6,074 mm3 (6 cm3). Serial examinations of the MR imaging signal characteristics of these CMs demonstrate a trend for maturation of blood products from a subacute, to a mixed, and finally to a chronic appearance. Three lesions appeared de novo during the follow-up period. CONCLUSIONS: On the basis of their analysis, the authors conclude that CMs exhibit a range of dynamic behaviors including enlargement, regression, and de novo formation, as well as progression through a series of characteristic MR imaging appearances. PMID- 11117870 TI - Long-term natural history of hemorrhagic moyamoya disease in 42 patients. AB - OBJECT: The purpose of this study was to delineate the long-term natural history of hemorrhagic moyamoya disease (MMD). METHODS: A retrospective review was conducted among 42 patients suffering from hemorrhagic MMD who had been treated conservatively without bypass surgery. The group included four patients who had undergone indirect bypass surgery after an episode of rebleeding. The follow-up period averaged 80.6 months. The clinical features of the first bleeding episode and repeated bleeding episodes were analyzed to determine the risk factors of rebleeding and poor outcome. Intraventricular hemorrhage with or without intracerebral hemorrhage was a dominant finding on computerized tomography scans during the first bleeding episode in 29 cases (69%). During the follow-up period, 14 patients experienced a second episode of bleeding, which occurred 10 years or longer after the original hemorrhage in five cases (35.7%). The annual rebleeding rate was 7.09%/person/year. The second bleeding episode was characterized by a change in which hemisphere bleeding occurred in three cases (21.4%) and by the type of bleeding in seven cases (50%). After rebleeding the rate of good recovery fell from 45.5% to 21.4% and the mortality rate rose from 6.8% to 28.6%. Rebleeding and patient age were statistically significant risk factors of poor outcome. All four patients in whom there was indirect revascularization after the second bleeding episode experienced a repeated bleeding episode within 8 years. CONCLUSIONS: The occurrence of rebleeding a long time after the first hemorrhagic episode was not uncommon. Furthermore, the change in which hemisphere and the type of bleeding that occurred after the first episode suggested the difficulty encountered in the prevention of repeated hemorrhage. PMID- 11117872 TI - Stereotactic radiosurgery for cavernous malformations. AB - OBJECT: The use of stereotactic radiosurgery to treat cerebral cavernous malformations (CMs) is controversial. To evaluate the efficacy and safety of CM radiosurgery, the authors reviewed the experience at the Mayo Clinic during the past 10 years. METHODS: Seventeen patients underwent radiosurgery for high surgical-risk CMs in the following sites: thalamus/basal ganglia (four patients), brainstem (12 patients), and corpus callosum (one patient). All patients had experienced at least two documented hemorrhages before undergoing radiosurgery. Stereotactic magnetic resonance (MR) imaging was used for target localization in all cases. The median margin radiation dose was 18 Gy and the median maximum dose was 32 Gy. The median length of follow-up review following radiosurgery was 51 months. The annual hemorrhage rate during the 51 months preceding radiosurgery was 40.1%, compared with 8.8% in the first 2 years following radiosurgery and 2.9% thereafter. In 10 patients (59%) new neurological deficits developed that were associated with regions of increased signal on long-repetition time MR imaging performed a median of 8 months (range 5-16 months) after radiosurgery. Three patients recovered, giving the group a permanent radiation-related morbidity rate of 41%. Compared with 31 patients harboring arteriovenous malformations (AVMs) of sizes and in locations similar to those of the aforementioned CMs, who underwent radiosurgery during the same time period, the patients with CMs were more likely to experience radiation-related complications (any complication, 59% compared with 10%; p < 0.001; permanent complication, 41% compared with 10%; p = 0.02). CONCLUSIONS: It is impossible to conclude that radiosurgery protects patients with CMs against future hemorrhage risk based on the available data, although it appears that some reduction in the bleeding rate occurs after a latency interval of several years. The risk of radiation-related complications after radiosurgery to treat CMs is greater than that found after radiosurgery in AVMs, even when adjusting for lesion size and location and for radiation dose. PMID- 11117873 TI - Efficacy and safety of the endothelin, receptor antagonist TAK-044 in treating subarachnoid hemorrhage: a report by the Steering Committee on behalf of the UK/Netherlands/Eire TAK-044 Subarachnoid Haemorrhage Study Group. AB - OBJECT: Delayed cerebral ischemia remains an important cause of death and disability in patients who have suffered subarachnoid hemorrhage (SAH). Endothelin (ET) has a potent contractile effect on cerebral arteries and arterioles and has been implicated in vasospasm. The authors administered ET(A/B) receptor antagonist (TAK-044) to patients suffering from aneurysmal SAH. They then assessed whether this agent reduced the occurrence of delayed cerebral ischemic events and examined its safety profile in this group of patients. METHODS: Four hundred twenty patients who had suffered an SAH were recruited into a multicenter, randomized, double-blind, placebo-controlled, parallel-group phase II trial. The primary end point was whether a delayed ischemic event occurred within 3 months after the first dose of the study drug and the secondary end points included determining whether a delayed ischemic event occurred by 10 days after the first dose of the study drug, whether a new cerebral infarct was demonstrated on a computerized tomography scan or at postmortem examination by 3 months after administration of the initial dose, the patient's Glasgow Outcome Scale scores at 3 months after the initial dose, and adverse events. There was a lower incidence of delayed ischemic events at 3 months in the TAK-044-treated group: 29.5% compared with 36.6% in a group of patients receiving placebo. The estimated relative risk was 0.8 with a 95% confidence interval of 0.61 to 1.06. There were no significant differences in the secondary end points, including clinical outcomes in the placebo-treated and TAK-044-treated groups. CONCLUSIONS: The TAK-044 was well tolerated by patients who had suffered an SAH, even though hypotension and headache--side effects compatible with the drug's vasodilatory properties--occurred. It would be valuable to proceed to a fully powered phase III trial of an ET receptor antagonist in treating aneurysmal SAH. PMID- 11117874 TI - Ruptured irregularly shaped aneurysms: pseudoaneurysm formation in a thrombus located at the rupture site. AB - OBJECT: The authors describe the clinical, radiological, and pathological findings of ruptured cerebral aneurysms with irregular configurations. METHODS: Eight patients with subarachnoid hemorrhage due to ruptured irregularly shaped aneurysms were examined. The preoperative radiological findings in these cases were compared with the pathological and operative findings of endovascular or open surgery. All of the aneurysms exhibited delayed opacification and delayed washout of contrast medium from the irregularly shaped portion of the aneurysm on digital subtraction angiography and/or helical computerized tomography scanning. Endovascular embolization with platinum coils was attempted in the first four patients who underwent treatment. In three of these patients the aneurysm ruptured again during the endovascular procedure. In the fourth patient an intraaneurysm thrombus was observed during the procedure and clipping was performed. In the subsequent four patients, three underwent clipping without complication and one underwent partial aneurysm embolization because of poor general status. A thrombus adjacent to the aneurysm dome was observed in the patients who underwent open surgery. Pathological examination of the operative specimens revealed a pseudoaneurysm-like cavity in the thrombus that was adherent to the aneurysm. CONCLUSIONS: Ruptured irregularly shaped aneurysms may be accompanied by fragile pseudoaneurysm-like cavities located at the rupture point. Because these aneurysms have a high risk of repeated rupture during an endovascular procedure, advancing microinstruments to the weaker portion of the aneurysm should be avoided. PMID- 11117875 TI - Characterization of polyphenol oxidase in coffee. AB - Polyphenol oxidase (PPO) was characterized in partially purified extracts of leaves (PPO-L) and fruit endosperm (PPO-E) of coffee (Coffea arabica L.). PPO activity was higher in early developmental stages of both leaves and endosperm of fruits. Wounding or exposure of coffee leaves to methyl jasmonate increased PPO activity 1.5-4-fold. PPO was not latent and was not activated by protease treatment. PPO activity was stimulated 10-15% with sodium dodecyl sulphate (SDS) at 0.35-1.75 mM, but at higher concentrations activities were similar to the control samples, without detergent. Prolonged incubation of extracts with trypsin or proteinase K inhibited PPO activity but pepsin had no effect. Inhibition of PPO with proteinase K was increased in the presence of SDS. PPO activity from both tissues was optimal at pH 6-7 and at an assay temperature of 30 degrees C. Activity was highest with chlorogenic acid as substrate with a Km of 0.882 mM (PPO-L) and 2.27 mM (PPO-E). Hexadecyl trimethyl-ammonium bromide, polyvinylpyrrolidone 40. cinnamic acid and salicylhydroxamic acid inhibited PPO from both tissues. Both enzymes were inactivated by heat but the activity in endosperm extracts was more heat labile than that from leaves. The apparent Mr determined by gel filtration was 46 (PPO-L) and 50 kDa (PPO-E). Activity-stained SDS polyacrylamide gel electrophoresis (PAGE) gels and western blots probed with PPO antibodies suggested the existence of a 67 kDa PPO which is susceptible to proteolytic cleavage that generates a 45 kDa active form. PMID- 11117876 TI - A 38 kDa allylic alcohol dehydrogenase from the cultured cells of Nicotiana tabacum. AB - An NADP+-dependent alcohol dehydrogenase (allyl-ADH) was isolated from the cultured cells of Nicotiana tabacum. The allyl-ADH was found to be efficient for the dehydrogenation of secondary allylic alcohols rather than saturated secondary alcohols and it was specific for the S-stereoisomer of the alcohols. The enzyme catalyzed the reversible reaction whereby the carbonyl group of enones is reduced to the corresponding allylic alcohol or vice versa. Two possible primary structures of the allyl-ADH were deduced by the sequence analyses of full-length cDNAs (allyl-ADH1 and ally-ADH2), which were cloned by the PCR method. These analyses indicated that the allyl-ADHs are composed of 343 amino acids having the molecular weights 38083 and 37994, respectively, and they showed approximately 70% homology to the NADP+-dependent oxidoreductases belonging to a plant zeta crystallin family. PMID- 11117877 TI - Cloning and expression of homospermidine synthase from Senecio vulgaris: a revision. AB - Homospermidine synthase. which catalyses the first pathway-specific reaction in pyrrolizidine alkaloid biosynthesis, was cloned from root cultures of Senecio vulgaris and expressed in E. coli. The open reading frame encodes a protein of 370 amino acids with a molecular mass of 40,740 Da. The enzyme is strictly dependent on spermidine as aminobutyl donor since it cannot be substituted by putrescine. The homospermidine synthase from S. vulgaris showed 97.9 and 99.3% nucleic acid identity with two HSS sequences from the closely related species Senecio vernalis. This report also revises data from a previous publication (Kaiser, A., 1999. Cloning and expression of a cDNA encoding homospermidine synthase from Senecio vulgaris (Asteraceae) in Escherichia coli. Plant J. 19. 195 201.) that is incorrect. PMID- 11117878 TI - UV mutagenesis and enzyme inhibitors as tools to elucidate the late biosynthesis of the spirobisnaphthalenes. AB - The metabolite pattern of UV mutants of the spirobisnaphthalene producing fungus F-24'707 by TLC and HPLC analysis has been investigated. Mutants with differences in colony morphology or colour compared to the parent strain were isolated. Cultivation in shaking flasks and P flasks showed differences in the metabolite pattern of some of the strains. Furthermore, enzyme inhibitors were used to block the spirobisnaphthalene biosynthesis of the parent strain at different steps. Feeding of precursors and intermediates of cladospirone bisepoxide (15) led to a two-fold increase of the production of 15. From these data and preceding biosynthetic studies we deduced a general pathway for the biosynthesis of all spirobisnaphthalenes of the fungus F-24'707. This enables us to present the hypothesis that all bisnaphthalenes described so far are produced using a common pathway with only a few intermediates as central branching points. PMID- 11117879 TI - Gibberellin biosynthesis: metabolic evidence for three steps in the early 13 hydroxylation pathway of rice. AB - [14C4]GA53, [14C4]GA44, and [2H2/14C4]GA19 were injected separately into seedlings of rice (Oryza sativa) using a dwarf mutant (d35) that has low levels of endogenous gibberellins (GAs). After 8 h incubation, the shoots were extracted and the labeled metabolites were identified by full-scan gas chromatography mass spectrometry (GC-MS) and Kovats retention indices (KRIs). Our results document the metabolic sequence, GA53-->GA44-->GA19-->GA20 and the presence of endogenous GA53, GA44, GA19, GA20 and GA1. Previous metabolic studies have shown the presence of the step, GA20-->GA1 in rice. Taken together, the data establish in vegetative shoots of rice the presence of the early 13-hydroxylation pathway, a pathway that originates from GA12 and leads to bioactive GA1. PMID- 11117881 TI - Anthocyanic vacuolar inclusions--their nature and significance in flower colouration. AB - The petals of a number of flowers are shown to contain similar intensely coloured intravacuolar bodies referred to herein as anthocyanic vacuolar inclusions (AVIs). The AVIs in a blue-grey carnation and in purple lisianthus have been studied in detail. AVIs occur predominantly in the adaxial epidermal cells and their presence is shown to have a major influence on flower colour by enhancing both intensity and blueness. The latter effect is especially dramatic in the carnation where the normally pink pelargonidin pigments produce a blue-grey colouration. In lisianthus, the presence of large AVIs produces marked colour intensification in the inner zone of the petal by concentrating anthocyanins above levels that would be possible in vacuolar solution. Electron microscopy studies on lisianthus epidermal tissue failed to detect a membrane boundary in AVI bodies. AVIs isolated from lisianthus cells are shown to have a protein matrix. Bound to this matrix are four cyanidin and delphinidin acylated 3,5 diglycosides (three, new to lisianthus), which are relatively minor anthocyanins in whole petal extracts where acylated delphinidin triglycosides predominate. Flavonol glycosides were not bound. A high level of anthocyanin structural specificity in this association is thus implied. The specificity and effectiveness of this anthocyanin "trapping" is confirmed by the presence in the surrounding vacuolar solution of only delphinidin triglycosides, accompanied by the full range of flavonol glycosides. "Trapped" anthocyanins are shown to differ from solution anthocyanins only in that they lack a terminal rhamnose on the 3 linked galactose. The results of this study define for the first time the substantial effect AVIs have on flower colour, and provide insights into their nature and their specificity as vacuolar anthocyanin traps. PMID- 11117880 TI - Biosynthesis of the dimethylallyl moiety of glabrol in Glycyrrhiza glabra hairy root cultures via a non-mevalonate pathway. AB - Incorporation of [1-13C]glucose indicates that the biosynthesis of the hemiterpene moiety of glabrol, the main prenylated flavanone in the hairy root cultures of Glycyrrhiza glabra, proceeds via a glyceraldehyde/pyruvate non mevalonate pathway. PMID- 11117882 TI - Chemotaxonomy of Plantago. Iridoid glucosides and caffeoyl phenylethanoid glycosides. AB - Data for 34 species of Plantago (Plantaginaceae), including subgen. Littorella (= Littorella uniflora), have been collected with regard to their content of iridoid glucosides and caffeoyl phenylethanoid glycosides (CPGs). In the present work, 21 species were investigated for the first time and many known compounds were found together with three new iridoid glucosides. Of these, arborescoside and arborescosidic acid, both of the uncommon type with an 8,9-double bond, were present in several species, while 6-deoxymelittoside was found only in P. subulata. The known compounds deoxyloganic acid, caryoptoside and rehmannioside D were isolated from the genus for the first time. The earlier reported occurrence of sorbitol in the family was confirmed, and this compound was shown by NMR spectroscopy to be the main sugar in the three species investigated for this. The combined data show that CPGs are present in all species investigated. With regard to the iridoids, the distribution patterns showed a good correlation with the classification of Rahn. Thus, aucubin is typical for the whole genus, while bartsioside and catalpol as well as 5-substituted iridoids are each characteristic for a subgenus in the family. Finally, the close relationship between Plantago and Veronica suggested by chloroplast DNA sequence analysis. could be corroborated by the common occurrence of the rare 8,9-unsaturated iridoids in these two genera. PMID- 11117883 TI - Antimalarial preracemosols A and B, possible biogenetic precursors of racemosol from Bauhinia malabarica Roxb. AB - Racemosol and demethylracemosol, together with their possible biogenetic precursors, preracemosol A and preracemosol B, were isolated from the roots of Bauhinia malabarica Roxb. While only racemosol and demethylracemosol exhibited cytotoxicity against KB and BC cell lines, all four compounds exhibited moderate antimalarial activity. PMID- 11117884 TI - Iridoid and megastigmane glycosides from Phlomis aurea. AB - From the leaves of Phlomis aurea, two new iridoids of unique structures named 3 epiphlomurin (1) and phlomurin (2), a new megastigmane glucoside phlomuroside (3) and a new benzyl alcohol glycoside having the structure benzyl alcohol-O-beta xylopyranosyl-(1-->2)-beta-glucopyranoside (4) have been isolated together with four known iridoids auroside, lamiide, 8-epiloganin and ipolamiide, two known phenolic glycosides acteoside (verbascoside) and syringin, one known phenylethanoid glycoside 2-phenylethyl-O-beta-xylopyranosyl-(1-->2)-beta glucopyranoside, one known lignan liriodendrin and three known flavonoids chrysoeriol-7-O-beta-glucopyranoside, acacetin-7-O-beta-glucopyranoside and luteolin-7-O-beta-glucopyranoside. The structures of the isolated compounds were verified by means of mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectral analyses. PMID- 11117885 TI - Eleganoside-A, B and C from Pseudocalymma elegans, a native of Brazil. AB - The toxic methanol-soluble part of Pseudocalymma elegans (leaves), a native of Brazil, yielded three new iridoidglucosides (1a-3a) as their acetate-derivatives (1-3) named eleganoside-A (1a), B (2a) and C (3a) which have been characterized with the aid of spectroscopic techniques, including 2D NMR. PMID- 11117886 TI - Biotransformation of geraniol, nerol and citral by sporulated surface cultures of Aspergillus niger and Penicillium sp. AB - The biotransformation of geraniol, nerol and citral by Aspergillus niger was studied. A comparison was made between submerged liquid, sporulated surface cultures and spore suspensions. This bioconversion was also carried out with surface cultures of Penicillium sp. The main bioconversion products obtained from geraniol and nerol by liquid cultures of A. niger were linalool and alpha terpineol. Linalool, alpha-terpineol and limonene were the main products obtained from nerol and citral by sporulated surface cultures, whereas geraniol was converted predominantly to linalool, also resulting in higher yields. Bioconversion of nerol with Penicillium chrysogenum yielded mainly alpha terpineol and some unidentified compounds. With P. rugulosum the major bioconversion product from nerol and citral was linalool. The bioconversion of nerol to alpha-terpineol and linalool by spore suspensions of A. niger was also investigated. Finally the biotransformation with sporulated surface cultures was also monitored by solid phase microextraction (SPME). It was found that SPME is a very fast and efficient screening technique for biotransformation experiments. PMID- 11117891 TI - An aphid repellent glycoside from Solanum laxum. AB - A spirostanic saponin was isolated from the ethanolic extract of the aerial parts of Solanum laxum Steud. The compound, named luciamin, was characterised by NMR spectroscopy, mass spectrometry and chemical methods, as (22R, 25S)-spirost-5-en 3 beta, 15 alpha-diol 3-O-|beta-D-glucopyranosyl (1 --> 2)-beta-D-glucopyranosyl ( 1 --> 4)-[alpha-L-rhamnopyranosyl-( 1 --> 2)]-beta-D-galactopyranoside|. The compound was tested against the aphid Schizaphis graminum by incorporation in artificial diets. It showed a deterrent (toxic) activity against the insect and is the first spirostane glycoside reported to have this activity. PMID- 11117901 TI - Secoiridoid glucosides from Fraxinus americana. AB - Investigation of the leaves of Fraxinus americana led to the isolation of five secoiridoid glucosides, demethylligstroside, (2"R)- and (2"S)-2" hydroxyoleuropeins, fraxamoside and frameroside, together with 18 known compounds. Their structures were determined on the basis of spectroscopic studies and chemical evidence. PMID- 11117902 TI - Back to the laws of thermodynamics. PMID- 11117903 TI - Mobile phones and the illusory pursuit of safety. PMID- 11117904 TI - Calprotectin, a faecal marker of organic gastrointestinal abnormality. PMID- 11117905 TI - Increasing demand while decreasing costs of generic medicines. PMID- 11117906 TI - Helium/oxygen and severe COPD. PMID- 11117907 TI - Oiling of health messages in marketing of food. PMID- 11117908 TI - Influence of potty training habits on dysfunctional bladder in children. PMID- 11117909 TI - What is mentoring? PMID- 11117910 TI - Rhythm or rate control in atrial fibrillation--Pharmacological Intervention in Atrial Fibrillation (PIAF): a randomised trial. AB - BACKGROUND: Atrial fibrillation is the most commonly encountered sustained cardiac arrhythmia. Restoration and maintenance of sinus rhythm is believed by many physicians to be superior to rate control only. However, there are no prospective data that compare both therapeutic strategies. METHODS: The Pharmacological Intervention in Atrial Fibrillation (PIAF) trial was a randomised trial in 252 patients with atrial fibrillation of between 7 days and 360 days duration, which compared rate (group A, 125 patients) with rhythm control (group B, 127 patients). In group A, diltiazem was used as first-line therapy and amiodarone was used in group B. The primary study endpoint was improvement in symptoms related to atrial fibrillation. FINDINGS: Over the entire observation period of 1 year, a similar proportion of patients reported improvement in symptoms in both groups (76 responders at 12 months in group A vs 70 responders in group B, p=0.317). Amiodarone administration resulted in pharmacological restoration of sinus rhythm in 23% of patients. Walking distance in a 6 min walk test was better in group B compared with group A, but assessment of quality of life showed no differences between groups. The incidence of hospital admission was higher in group B (87 [69%] out of 127 vs 30 [24%] out of 125 in group A, p=0.001). Adverse drug effects more frequently led to a change in therapy in group B (31 [25%] patients compared with 17 [14%] in group A, p=0.036). INTERPRETATION: With respect to symptomatic improvement in patients with atrial fibrillation, the therapeutic strategies of rate versus rhythm control yielded similar clinical results overall. However, exercise tolerance is better with rhythm control, although hospital admission is more frequent. These data may serve as a basis to select therapy in individual patients. PMID- 11117911 TI - Natural cytotoxic activity of peripheral-blood lymphocytes and cancer incidence: an 11-year follow-up study of a general population. AB - BACKGROUND: One of the most critical questions in immunosurveillance is whether differences between individuals with regards to natural immunological host defence can predict future development of cancer. Although this question has so far remained open, there are clear indications of significant roles of several naturally cytotoxic lymphocytes in preventing the development of cancer. We began a prospective cohort study among a Japanese general population in 1986, using various immunological and biochemical markers. METHODS: Natural cytotoxic activity of peripheral-blood mononuclear cells was assessed by isotope-release assay in 3625 residents of a Japanese population mostly older than 40 years of age, between 1986 and 1990. Immunological and biochemical markers were also measured, and participants were given a questionnaire on lifestyle. We did an 11 year follow-up survey of the cohort members looking at cancer incidence and death from all causes, and analysed the association between cytotoxic activity of peripheral-blood lymphocytes assessed at baseline and cancer incidence found in the subsequent follow-up. FINDINGS: 154 cancer cases were used in the analysis. When we categorised the cytotoxic activity of peripheral-blood lymphocytes by tertiles, age-adjusted relative risk of cancer incidence (all sites) was 0.72 (95% CI 0.45-1.16) for men with high cytotoxic activity, and 0.62 (0.38-1.03) for men with medium cytotoxic activity, taking the risk of those with low cytotoxic activity as reference. For women with high cytotoxic activity relative risk was 0.52 (0.28-0.95), and for those with medium cytotoxic activity 0.56 (0.31-1.01). For both sexes with high and medium cytotoxic activity risk was 0.63 (0.43-0.92) and 0.59 (0.40-0.87), respectively. INTERPRETATION: Our results indicate that medium and high cytotoxic activity of peripheral-blood lymphocytes is associated with reduced cancer risk, whereas low activity is associated with increased cancer risk suggesting a role for natural immunological host defence mechanisms against cancer. PMID- 11117912 TI - Clinical progression, survival, and immune recovery during antiretroviral therapy in patients with HIV-1 and hepatitis C virus coinfection: the Swiss HIV Cohort Study. AB - BACKGROUND: Hepatitis C virus (HCV) infection is highly prevalent among HIV-1 infected individuals, but its contribution to the morbidity and mortality of coinfected patients who receive potent antiretroviral therapy is controversial. We used data from the ongoing Swiss HIV Cohort Study to analyse clinical progression of HIV-1, and the virological and immunological response to potent antiretroviral therapy in HIV-1-infected patients with or without concurrent HCV infection. METHODS: We analysed prospective data on survival, clinical disease progression, suppression of HIV-1 replication, CD4-cell recovery, and frequency of changes in antiretroviral therapy according to HCV status in 3111 patients starting potent antiretroviral therapy. RESULTS: 1157 patients (37.2%) were coinfected with HCV, 1015 of whom (87.7%) had a history of intravenous drug use. In multivariate Cox's regression, the probability of progression to a new AIDS defining clinical event or to death was independently associated with HCV seropositivity (hazard ratio 1.7 [95% CI 1.26-2.30]), and with active intravenous drug use (1.38 [1.02-1.88]). Virological response to antiretroviral therapy and the probability of treatment change were not associated with HCV serostatus. In contrast, HCV seropositivity was associated with a smaller CD4-cell recovery (hazard ratio for a CD4-cell count increase of at least 50 cells/microL=0.79 [0.72-0.87]). INTERPRETATION: HCV and active intravenous drug use could be important factors in the morbidity and mortality among HIV-1-infected patients, possibly through impaired CD4-cell recovery in HCV seropositive patients receiving potent antiretroviral therapy. These findings are relevant for decisions about optimum timing for HCV treatment in the setting of HIV infection. PMID- 11117913 TI - End-of-life decisions in medical practice in Flanders, Belgium: a nationwide survey. AB - BACKGROUND: Our study is a repeat of the Dutch death-certificate study on end-of life decisions (ELDs). The main objective was to estimate the frequency of euthanasia (the administration of lethal drugs with the explicit intention of shortening the patient's life at the patient's explicit request), physician assisted suicide (PAS), and other ELDs in medical practice in Flanders, Belgium. METHODS: A 20% random sample of 3999 deaths was selected from all deaths recorded between Jan 1 and April 30, 1998. The physicians who signed the corresponding death certificates received one questionnaire by post per death. FINDINGS: The physicians' response rate was 1355 (52%). 1925 deaths were described. The results were corrected for non-response bias, and extrapolated to estimated annual rates after seasonal adjustment for death causes, and we estimate that 705 (1.3%, 95% CI 1.0-1.6) deaths resulted from euthanasia or PAS. In 1796 (3.2%, 2.7-3.8) cases, lethal drugs were given without the explicit request of the patient. Alleviation of pain and symptoms with opioids in doses with a potential life shortening effect preceded death in 10,416 (18.5%, 17.3-19.7) cases and non treatment decisions in 9218 (16.4%, 15.3-17.5) cases, of which 3261 (5.8%, 5.1 6.5) with the explicit intention of ending the patient's life. INTERPRETATION: ELDs are prominent in medical practice in Flanders. The frequency of deaths preceded by an ELD is similar to that in the Netherlands, but lower than that in Australia. However, in Flanders the rate of administration of lethal drugs to patients without their explicit request is similar to Australia, and significantly higher than that in the Netherlands. PMID- 11117914 TI - A prospective social and molecular investigation of gonococcal transmission. AB - BACKGROUND: Gonorrhoea is a common infectious disease, poorly controlled despite effective treatments. Tracing chains of transmission is difficult, because sexual partners are commonly difficult or impossible to identify. We assess the use of gonococcal opa-typing in identifying transmission links not revealed through interview. METHODS: Epidemiological data and gonococcal isolates were collected prospectively from patients at two UK clinics in London and Sheffield. Social and epidemiological data were combined with molecular typing of gonococcal isolates by a new methodology based on the polymorphisms of the opa gene. FINDINGS: In London, interview data and opa-typing on samples from 215 cases showed a diverse population with few links. In Sheffield, interview data identified links between 51 (43%) of 120 cases, whereas opa-typing suggested a more connected population: 95 (79%) of cases had shared profiles. There was a highly significant correlation between the two distributions with epidemiological clusters appearing as a subset of the opa clusters. Two large opa clusters, of 18 and 43 cases, accounted for 50% of local cases of gonorrhoea. Discordance between epidemiological and opa typing data was observed at highly connected points in the sexual network. INTERPRETATION: Opa-typing is a more powerful tool for epidemiological investigation of gonorrhoea transmission than earlier methods. Opa-typing can link infections that would otherwise remain unlinked, and may aid interventions to control endemic disease. PMID- 11117915 TI - Clinical picture. Mucocutaneous pigmentation due to zinc deposition. PMID- 11117916 TI - Subarachnoid haemorrhage treated with anticoagulants. PMID- 11117917 TI - Prenatal detection of fetal Down's syndrome from maternal plasma. AB - Fetal DNA is present in maternal plasma, and a proportion of such DNA is seen in intact fetal cells. We investigated the use of fluorescence in-situ hybridisation (FISH) techniques on maternal plasma samples. In plasma samples obtained from three women carrying fetuses affected by trisomy 21 (Down's syndrome), we identified fetal cells with three chromosome-21 signals. These results show the feasibility of non-invasive detection of fetal chromosomal aneuploidy by maternal plasma analysis. PMID- 11117918 TI - Hereditary catalase deficiencies and increased risk of diabetes. AB - Partial or near-total lack of erythrocyte catalase activity is a rare condition, generally thought to be benign. However, little is known of the frequency of common diseases of adult onset in human beings with catalase deficiency. We report that, in a series of Hungarian patients with catalase deficiency, there is a higher frequency of diabetes than in unaffected first-degree relatives and the general Hungarian population. We speculate that quantitative deficiency of catalase might predispose to cumulative oxidant damage of pancreatic beta-cells and diabetes. PMID- 11117919 TI - Crohn's disease associated with spondyloarthropathy: effect of TNF-alpha blockade with infliximab on articular symptoms. AB - Four patients with Crohn's disease and spondyloarthropathy were treated with infliximab for treatment-resistant gut inflammation. A substantial improvement in gastrointestinal signs and symptoms was noted, which was accompanied by a rapid reduction in C-reactive protein concentrations. Moreover, all four patients had a significant improvement of axial manifestations and/or peripheral arthritis, related to their spondyloarthropathy. This fast and substantial improvement of the articular manifestations of Crohn's disease after infliximab treatment warrants further investigation of the therapeutic potential of TNF-alpha blockade in other subtypes of spondyloarthropathies. PMID- 11117920 TI - Effects of posture on sympathetic nervous modulation in patients with chronic heart failure. AB - We investigated which recumbent position is preferred by patients with chronic heart failure (CHF) and whether sympathetic nervous modulation differs in three recumbent positions. We assessed 12 patients with CHF by spectral analysis of heart-rate variability and measurement of plasma norepinephrine concentrations. The right lateral decubitus position was preferred for significantly longer periods than the left lateral decubitus and supine positions. Sympathetic nervous modulation was most attenuated in the right lateral decubitus position. The right lateral decubitus position preferred by patients with CHF may be a self protective mechanism to control increased sympathetic nervous modulation. PMID- 11117921 TI - Health-seeking behaviour of individuals with a cough of more than 3 weeks. AB - Sex inequalities can lead to poorer access to health care and delays to diagnosis of tuberculosis in women. In a population-based survey we assessed health-seeking behaviour in adults with long-term cough. The prevalence of cough was 1% (213) and 2% (279) in men and women, respectively. Women took more health-care actions than men, but chose less qualified providers and reported lower health expenditure per visit. Delay before seeking hospital treatment was longer for women (41 days) than men (19 days; p=0.04), and more men (27; 36%) than women (14; 14%; p=0.0006) reported giving a sputum sample at hospital. Sex-sensitive strategies for tuberculosis control are needed and should take into account sex differences in health-care seeking behaviour as well as a possible sex bias among health-care providers. PMID- 11117922 TI - UK government looks to expand research on embryos. PMID- 11117923 TI - New risk factor for HIV-1 might be blessing in disguise. PMID- 11117924 TI - Low health literacy prevents equal access to care. PMID- 11117925 TI - US election deadlock portends little change in research. PMID- 11117926 TI - India health survey finds too many women and children in poor health. PMID- 11117927 TI - Physics and biology of mobile telephony. AB - Although safety guidelines--to which mobile telephones and their base-stations conform--do protect against excessive microwave heating, there is evidence that the low intensity, pulsed radiation currently used can exert subtle non-thermal influences. If these influences entail adverse health consequences, current guidelines would be inadequate. This review will focus on this possibility. The radiation used is indeed of very low intensity, but an oscillatory similitude between this pulsed microwave radiation and certain electrochemical activities of the living human being should prompt concern. However, being so inherently dependent on aliveness, non-thermal effects cannot be expected to be as robust as thermal ones, as is indeed found; nor can everyone be expected to be affected in the same way by exposure to the same radiation. Notwithstanding uncertainty about whether the non-thermal influences reported do adversely affect health, there are consistencies between some of these effects and the neurological problems reported by some mobile-telephone users and people exposed longterm to base station radiation. These should be pointers for future research. PMID- 11117928 TI - Epidemiological evidence on health risks of cellular telephones. AB - It is too soon for a verdict on the health risks from cellular telephones, especially in view of changing technology. From the Interphone project and some other large studies in progress, better information may emerge. Based on the epidemiological evidence available now, the main public-health concern is clearly motor vehicle collisions, a behavioural effect rather than an effect of radiofrequency exposure as such. Neither the several studies of occupational exposure to radiofrequencies nor the few of cellular telephone users offer any clear evidence of an association with brain tumours or other malignancies. Even if the studies in progress were to find large relative effects for brain cancer, the absolute increase in risk would probably be much smaller than the risk stemming from motor vehicle collisions. Cellular telephones affect the quality of our lives in myriad ways, for good and ill; the health risk is just one part of a picture that is slowly coming into focus. PMID- 11117929 TI - Oscar Wilde's terminal illness: reappraisal after a century. PMID- 11117930 TI - Randomised comparisons of medical tests: sometimes invalid, not always efficient. PMID- 11117931 TI - Intracranial haemorrhage with bolus thrombolytic agents. PMID- 11117932 TI - Intracranial haemorrhage with bolus thrombolytic agents. PMID- 11117933 TI - Intracranial haemorrhage with bolus thrombolytic agents. PMID- 11117934 TI - Intracranial haemorrhage with bolus thrombolytic agents. PMID- 11117935 TI - HIV-1 and scrub-typhus. PMID- 11117936 TI - Bone density in cosmonauts. PMID- 11117937 TI - Breast cancer and dieldrin. PMID- 11117938 TI - Cardiovascular disease in South Asians. PMID- 11117939 TI - Non-heart-beating donors for renal transplantation. PMID- 11117940 TI - Non-heart-beating donors for renal transplantation. PMID- 11117941 TI - Roll Back Malaria. PMID- 11117942 TI - Roll Back Malaria. PMID- 11117943 TI - Spread of HIV and AIDS in China. PMID- 11117944 TI - Prader-Willi syndrome. PMID- 11117945 TI - Jenner's cowskin. PMID- 11117946 TI - Host adapted serotypes of Salmonella enterica. PMID- 11117947 TI - Salmonella in sub-Antarctica: low heterogeneity in Salmonella serotypes in South Georgian seals and birds. AB - The number of human visitors to Antarctica is increasing rapidly, and with it a risk of introducing infectious organisms to native animals. To study the occurrence of salmonella serotypes in sub-Antarctic wildlife, faecal samples were collected from gentoo penguins, macaroni penguins, gray-headed albatrosses, black browed albatrosses and Antarctic fur seals on Bird Island in the South Georgian archipelago during the austral summer of 1996 and 1998. In 1996, S. havana, S. typhimurium and S. enteritidis were isolated from 7% of gentoo penguins and 4% of fur seals. In 1998, however, 22% of fur seals were found to be infected with S. havana, S. enteritidis and S. newport. All isolates, except one, showed identical pulsed-field gel electrophoresis-patterns within each serotype, irrespective of sampling year and animal reservoir. No significant antibiotic resistance was found. The very low heterogeneity in the salmonella isolates found could either indicate a high genetic adaptation of the bacteria to the environment or a recent introduction of salmonella into the area. PMID- 11117948 TI - Evaluation of a serological Salmonella mix-ELISA for poultry used in a national surveillance programme. AB - A Mix-ELISA using lipopolysaccharide antigens from Salmonella enterica serotype Enteritidis and Typhimurium was evaluated using samples collected over time in the Danish salmonella surveillance programme for poultry. Serological samples (n = 42,813) taken from broiler-breeder flocks after a year of bacteriological monitoring with negative results were used for calculating the flock and individual test specificities, which were 0.997 and 0.999, respectively. Layer flocks from the table egg sector were used for calculation of positive predictive values. In the survey, flocks were examined for salmonella by Mix-ELISA and by faecal culture, and in case of a positive result in either of these a repeated, serological testing was performed, and 60 animals were organ-cultured. If one of these samplings was positive, the flock was declared salmonella infected. In a period of 3 months, 35 flocks were found to be positive in the routine samples. Of these, 32 were serologically positive, 2 both serologically and faecally positive and 1 flock only faecally positive. For flocks serologically positive in the surveillance programme, a positive-predictive value of 0.62 for organ culture positivity was found, and while considering serological follow-up samples, the value was 0.95. PMID- 11117949 TI - Risk factors for indigenous campylobacter infection: a Swedish case-control study. AB - A case-control study was conducted in western Sweden (Alvsborg County). The aim of the study was to identify any special food items or behaviours associated with an increased risk of contracting campylobacter infection. A total of 101 cases and 198 controls were matched for age, sex and district of residence. The following risk factors or risk behaviours were associated with campylobacter infection: drinking unpasteurized milk (OR 3.56, 95% CI 1.46-8.94), eating chicken (OR 2.29, 95% CI 1.29-4.23), or eating pork with bones (chops OR 2.02, 95% CI 1.17-3.64; loin of pork OR 1.83, 95% CI 1.07-3.12), barbecuing (OR 1.98, 95% CI 1.10-4.34), and living or working on a farm (farm OR 3.06, 95% CI 1.58 6.62, hen/chicken-breeder OR 3.32, 95% CI 1.56-6.78), daily contact with chickens or hens (OR 11.83, 95% CI 3.41-62.03). PMID- 11117950 TI - Improved methods of detection of meningococcal DNA from oropharyngeal swabs from cases and contacts of meningococcal disease. AB - In the UK the increasing use of pre-admission parenteral antibiotic therapy in meningococcal disease has lessened the value of routine cultures as a tool to confirm diagnosis, and laboratory confirmation of invasive meningococcal infection is achieved increasingly by non-culture, nucleic acid amplification methods. The purpose of this study was to evaluate a DNA extraction and meningococcal-specific DNA amplification methodology for detection of meningococci from oropharyngeal swabs. One hundred and six swabs from suspected or confirmed cases of meningococcal disease, and 94 swabs from contacts of meningococcal disease cases were examined. Of laboratory-confirmed cases, 38/65 (58.5%) yielded a positive oropharyngeal swab PCR result and 5/24 (20.8%) swabs from suspected but laboratory-unconfirmed cases were PCR positive. No significant differences in PCR positivity rates were found between the types of swab transport systems utilized, but transport time to the testing laboratory was found to affect PCR positivity (P < 0.05). Application of meningococcus-specific PCR to oropharyngeal swabs, in addition to routine culture of swabs, can provide valuable epidemiological information as well as case confirmation for contact management. PCR amplification of meningococcal PCR from oropharyngeal swabs will also increase the ascertainment in swabbing surveys carried out as part of meningococcal disease outbreak investigation and management. PMID- 11117951 TI - Genetic characterization of a new variant within the ET-37 complex of Neisseria meningitidis associated with outbreaks in various parts of the world. AB - A new variant within the electrophoretic type (ET)-37 complex of Neisseria meningitidis, ET-15, first detected in Canada in 1986, has been associated with severe clinical infections and high mortality rates in several European countries, Israel and Australia. To ascertain the genetic and epidemiological relationships of ET-15 strains from different geographical areas, 72 ET-15 isolates from 10 countries were compared to 13 isolates representing other clones of the ET-37 complex. The 85 strains were analysed by pulsed-field gel electrophoresis (PFGE) using 2 restriction endonucleases and Southern hybridization with 10 genetic markers. Four ET-15 strains and 4 other strains of the ET-37 complex were further examined using an additional restriction enzyme and a total of 18 genetic markers. PFGE fingerprints of the ET-15 strains were closely related. Strains within each country were even more closely related, suggesting single introductions of the clone. Physical mapping of genes in ET-15 and other strains of the ET-37 complex demonstrated that large genetic rearrangements of the genome have occurred in association with the appearance of the ET-15 variant. PMID- 11117952 TI - Genetic relatedness of group A streptococci of the newly designated serotype M90 causing a food-borne outbreak and sporadic infections. AB - Twenty-six isolates of the newly designated M90 serotype group A Streptococcus (GAS) from a large food-borne outbreak of pharyngitis in Greece and six M90 sporadic isolates from UK, were typed by pulsed-field gel electrophoresis (PFGE). Twenty-four outbreak isolates were identical and two closely related. The Greek isolates were possibly related with one UK isolate, while other sporadic isolates exhibited distinct PFGE profiles from the former isolates. PMID- 11117953 TI - Comparative investigations of Listeria monocytogenes isolated from a turkey processing plant, turkey products, and from human cases of listeriosis in Denmark. AB - Listeria monocytogenes was isolated from critical control points in a Danish turkey processing plant, from turkey products and from cases of human listeriosis. During processing in the plant the prevalence of L. monocytogenes ranged from 25.9 to 41.4%. Cleaning and disinfection decreased the prevalence to 6.4%. Isolates of L. monocytogenes were characterized by pulsed-field gel electrophoresis (PFGE) using restriction endonuclease ApaI. Identical DNA types were obtained from turkey products and the processing line even after cleaning and disinfection. Two identical DNA types were demonstrated among isolates from turkey products and human cases of listeriosis. The prevalence of L. monocytogenes in turkey products ranged from 7.3 to 17.4% for ready-to-eat products and raw products, respectively. Since none of the 27 flocks examined before slaughter sampled positive for L. monocytogenes and the prevalence increased during processing, the potential risk from turkey meat was apparently due to factory hygiene rather than intrinsic contamination of the turkeys. PMID- 11117954 TI - Risk factors for human brucellosis in Yemen: a case control study. AB - Brucellosis is known to occur in Yemen but its epidemiology has not been extensively studied. The present investigation examined risk factors for human brucellosis in Yemen using a hospital-based case-control study. A total of 235 consecutive patients with brucellosis attending the Central Health Laboratory in Sana'a, Yemen, were matched in respect of age, sex, and place of residence, rural or urban, with 234 controls selected from individuals attending the Central Health Laboratory for unrelated health problems. Clinical information on patients and controls was supplemented with occupational and socio-economic data obtained by interview of cases and controls using a standard questionnaire. After controlling for confounding factors significant risk factors for infection related to occupation as a farmer (OR 2.5 (95% CI 1.4-4.5, P < 0.0001)), shepherd (OR 7.8 (95% CI 1.0-61, P 0.05)) or microbiologist (OR 24.5 (95% CI 2.9-204, P 0.003)); and drinking fresh milk (OR 2.0 (95% CI 1.3-4.3, P 0.001)) and laban (OR 22.7 (95% CI 1.7-4.2 P < 0.0001)). Taking other milk products and offal were not risk factors. Socio-economic and educational factors were also independent risk factors. Occupational, food and socio-economic risk factors significantly confounded one another. Yemen shares some but not all of the risk factors of neighbouring countries. The interrelation between the various factors is complex and studying any one in isolation may give a false impression of its public health significance. Control through education of the population to minimize exposure to, and contact with, animals and their milk and milk products and to boil milk before drinking it or using it to make buttermilk, would be difficult as these would represent such fundamental changes to established patterns of behaviour of this society. Ideally there would be a campaign to control the infection by animal vaccination but the costs and logistic difficulty would be great. Presently there is a clear need for doctors in Yemen to be made aware of the frequency of this infection, the means available for clinical and laboratory diagnosis and effective treatment, while strategies to control the disease in Yemen are formulated and field tested. PMID- 11117955 TI - Tuberculosis in the veterans healthcare system: a six-year review and evaluation of programme effectiveness. AB - The Department of Veterans Affairs operates a large, centrally administered health care system consisting of 173 hospitals and 4 free standing outpatient clinics nationwide with approximately 945,115 hospital discharges, 24.2 million outpatient visits, and 2.86 million persons served annually over the time frame of the review. The purpose of the study was to define whether such a system could effect timely change in the incidence of tuberculosis (TB) using centralized programme planning and flexible field implementation. A retrospective review of the number of newly diagnosed cases of active TB treated at veterans health care facilities between 1 October 1990 and 30 September 1997 was determined by using a standardized annual case census. Intervention included implementation of the most current guidelines for the prevention of transmission of TB in the community and hospital setting, including administrative and engineering controls and a change in personal protective equipment. Centrally directed programme guidance, education, and funding were provided for field use in health care facilities of widely varying size and complexity. The numbers of total reported cases of TB decreased significantly (P < 0.001) throughout the veterans health care system (nationally and regionally), with the case rate decreasing at a rate significantly greater than that seen in the USA as a whole (P < 0.0001). TB associated with multi-drug resistance (isoniazid and rifampin) and HIV coinfection also significantly decreased over the study period. Therefore, a large, centrally administered health care system can effectively combat a re emerging infectious disease and may also demonstrate a successful outcome greater than seen in other, perhaps less organized health care settings. PMID- 11117956 TI - A waterborne outbreak of small round structured virus, campylobacter and shigella co-infections in La Neuveville, Switzerland, 1998. AB - An outbreak of gastro-enteritis occurred in La Neuveville, township with 3358 inhabitants. A retrospective cohort study of 1915 participants showed that 1607 (84%) had been ill. Campylobacter jejuni was isolated from 28 patient faecal samples, Shigella sonnei from 21 patients and small round structured viruses (SRSV) from 6 patients. More than one pathogen was identified in eight persons. The epidemic curve was characteristic of a point-source outbreak. The risk for illness was significantly higher among persons who had drank unboiled drinking water than among those who had not (1290 [80.3%] of 1607 vs. 86 [27.9%] of 308; RR = 2.87; 95% CI 2.40-3.45). Risk increased significantly with the quantity of water consumed (P < 0.00 x 10(-6)). An SRSV isolate from water and one human faeces had an identical DNA sequence. The outbreak was due to a pump failure producing a spill of sewage into the groundwater. We conclude that transmission was waterborne and that measures including early warning, basic hygiene and sanitation improvements controlled this epidemic. PMID- 11117957 TI - The epidemiology of adenovirus infections in Greater Manchester, UK 1982-96. AB - Data relating to 3313 adenovirus isolates from patients in Greater Manchester, UK between 1982 and 1996 were analysed using chi2 tests and 95% confidence intervals. Of the 3098 isolates that were typed, 18.6% were serotype 2, 14.9% serotype 3, 12.1% serotype 1 and 10.9% serotype 41. There was evidence of a seasonal occurrence of serotype 7 (March-August), serotype 2 (January-April), serotype 4 (June-August) and subgenus F (September-November). Children less than 5 years old were the most common group of patients with adenovirus infection (61.3%) compared to 24.2% for adults and only 5.6% for school children (5-15 years). Gastric symptoms were the most common amongst infants (47.6%) followed by respiratory (27.5%) and general symptoms (12.9%). In adults, the overwhelming clinical condition was conjunctivitis (88.9%). Despite the traditional association with adenoviruses, remarkably few cases of pharyngoconjunctival fever were recorded (1.7%). PMID- 11117958 TI - The seroepidemiology of rubella in western Europe. AB - Most of the countries in western Europe have now implemented mass infant rubella immunization programmes, instead of or in addition to selective vaccination in order to achieve the elimination of congenital rubella syndrome. The European countries Denmark, England and Wales, Finland, France, Germany, Italy and the Netherlands undertook large, national serological surveys collecting several thousand serum specimens during 1994-8. Antibodies against rubella virus were detected by a variety of enzyme immuno-assays. Comparability of the assay results was achieved by a standardized methodology. The age- and sex-stratified serological results were related to the schedules, coverage of rubella vaccination and the incidence in these countries. The results show widely differing levels of immunity to rubella both in the general population and in the specific age groups of males and females. A low rate (< 5%) of susceptibles in childhood and adolescents of both sexes was obtained only in Finland and the Netherlands. Countries such as Italy with only moderate coverage for the infant immunization programme currently have both high susceptibility levels in the general population and in the at-risk population. The likelihood is of continued epidemics of rubella with cases of congenital rubella syndrome. The continued implementation of selective vaccination will help to offset the impact of this ongoing transmission and to protect women on reaching childbearing age. PMID- 11117959 TI - Rubella in the Russian Federation: epidemiological features and control measures to prevent the congenital rubella syndrome. AB - A review of the epidemiology of clinical rubella in the Perm region of the Russian Federation from 1979-97 showed that the incidence was about 220 cases per 100,000 population. Congenital rubella syndrome (CRS) accounted for 15% of birth defects and for about 3.5 cases of CRS per 1000 live births per year. Surveys of the seroepidemiology of rubella infection revealed that the susceptibility rate among pregnant women (i.e. rubella virus antibody haemagglutination-inhibition (HAI) assay titres < 10) was 16.5%. As serum rubella antibody HAI titres > or = 10 both prevented infection in pregnant women and protected their foetuses, serological testing has been introduced into the routine antenatal services. Pre existing rubella antibodies were found not to interfere with the immune response to vaccination, so selective immunization was provided to girls approaching puberty and to women of childbearing age. A programme of epidemiological surveillance is being developed to define tactics for the widescale introduction of rubella vaccination. PMID- 11117960 TI - Community studies on hepatitis B in Rajahmundry town of Andhra Pradesh, India, 1997-8: unnecessary therapeutic injections are a major risk factor. AB - In Rajahmundry town in India, 234 community cases of jaundice were interviewed for risk factors of viral hepatitis B and tested for markers of hepatitis A-E. About 41% and 1.7% of them were positive for anti-HBc and anti-HCV respectively. Of 83 cases who were tested within 3 months of onset of jaundice, 5 (6%), 11 (13.3%), 1 (1.2%), 5 (6%) and 16 (19.3%) were found to have acute viral hepatitis A-E, respectively. The aetiology of the remaining 60% (50/83) of cases of jaundice could not be established. Thirty-one percent (26/83) were already positive for anti-HBc before they developed jaundice. History of therapeutic injections before the onset of jaundice was significantly higher in cases of hepatitis B (P = 0.01) or B-D (P = 0.04) than in cases of hepatitis A and E together. Other potential risk factors of hepatitis B transmission were equally prevalent in two groups. Subsequent studies showed that the majority of injections given were unnecessary (74%, 95% CI 66-82%) and were administered by both qualified and unqualified doctors. PMID- 11117961 TI - UK measles outbreak in non-immune anthroposophic communities: the implications for the elimination of measles from Europe. AB - We describe the epidemiology of the first nationwide outbreak of measles infection in the UK since the implementation of a mass vaccination campaign. Notifications of infectious diseases, interview and postal questionnaire identified 293 clinical cases, 138 of which were confirmed by salivary IgM, measles virus isolation and PCR. Twelve were epidemiologically linked to confirmed cases. The outbreak began in London, after contact with measles infection probably imported from Italy. Measles genotyping determined by sequence analysis confirmed spread to other unimmunized anthroposophic communities in the north, south west and south coast of England. Only two cases had been vaccinated against measles infection, and 90% of cases were aged under 15 years. Measles virus can selectively target non-immune groups in countries with high vaccine uptake and broader herd immunity. Without harmonization of vaccination policies and uniform high coverage across Europe, the importation and spread of measles virus amongst non-immune groups may prevent the elimination of measles. PMID- 11117962 TI - The epidemiology of measles in Poland: prevalence of measles virus antibodies in the population. AB - WHO has adopted a goal of eliminating indigenous measles from the European Region by the year 2007. The strategy focuses on reducing the proportion of susceptible individuals in the population to low levels and maintaining these low levels of susceptibility. Routine vaccination against measles for children aged 13-15 months was introduced in Poland in 1975, and a second dose added in 1991. High coverage (> 95%) is achieved with both doses. In order to assess progress towards measles elimination in Poland, a serological survey was performed to evaluate the impact of vaccination on the susceptibility profile of population. Three thousand residual serum samples from individuals aged 1-30 years were collected from hospitals in six selected voivodeships (administration units) in Poland. These were tested for measles-specific IgG using a commercial ELISA. Overall 4% (120/3000) were negative for measles virus antibody. The highest proportion of negatives (8.2%) occurred among cohorts born 1977-81--the only cohorts in which susceptibility exceeded the WHO targets. 'Catch-up' vaccination strategies should target these cohorts. PMID- 11117963 TI - Vaccine effectiveness for influenza in the elderly in welfare nursing homes during an influenza A (H3N2) epidemic. AB - Influenza vaccine effect on the occurrence and severity of influenza virus infection in a population residing in nursing homes for the elderly was studied as a cohort study during an influenza A (H3N2) epidemic in Japan. Of 22,462 individuals living in 301 welfare nursing homes, 10,739 voluntarily received inactivated, sub-unit trivalent influenza vaccine in a programme supported by the Osaka Prefectural Government. There were statistically significantly fewer cases of influenza, hospital admissions due to severe infection, and deaths due to influenza in the vaccinated cohort compared to the unvaccinated controls. No serious adverse reactions to vaccination were recorded. Thus influenza vaccination is effective for preventing influenza disease in persons aged 65 years and over, and should be an integral part of the care of this population residing in nursing homes. PMID- 11117964 TI - HCV prevalence in pregnant women in the UK. AB - The objective of the study was to assess the prevalence and epidemiology of hepatitis C virus (HCV) infection in pregnant women in the North Thames region, and in the UK in general. Demographic data were linked to neonatal samples prior to anonymization and testing by anti-HCV EIA, and with RIBA 3 confirmation. Risk factors for maternal infection were explored. Area-specific seroprevalence rates were multiplied into population sizes to estimate HCV prevalence in pregnant women in the UK. A total of 241/126,009 samples were confirmed anti-HCV positive, and a further 40 were indeterminate, representing a seroprevalence of 0.19-0.22%; 51% of maternal HCV infections were in UK-born women (71% of the population), and 22% in women from continental Europe (5% of the population). Among European-born women, HCV prevalence was associated with birth in continental Europe, partner not being notified at birth registration, partner born in a different region to the mother, and inner city residence. Four of the 241 anti-HCV positive samples (1.7%) were also anti-HIV-1 positive. It was estimated that each year an estimated 1150 out of 730,000 pregnancies in the UK would involve a woman infected with HCV (uncertainty range 660-1850), a prevalence of 0.16% (0.09 0.25%). On the basis of reported rates of mother-to-child transmission of HCV, this would represent approximately 70 paediatric HCV infections per year. PMID- 11117965 TI - Hepatitis B and C among Berlin dental personnel: incidence, risk factors, and effectiveness of barrier prevention measures. AB - A study of 215 Berlin dentists and 108 dental assistants recruited at the 1997 Berlin Dental Society meeting assessed their occupational risk of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection. HBV vaccine coverage, and barrier prevention methods used. Among dentists, 7% (95% CI 4-11) and 0.5% (95% CI 0-3) had serological evidence of previous HBV and HCV infection, respectively. Similar figures for dental assistants were 1% (95% CI 0-5) and 0% (95% CI 0-4). Only 74% of dentists and 63% of dental assistants reported HBV vaccination. Approximately half always used gloves, eye glasses, or face masks. HBV unvaccinated dentists whose patients had HBV risk factors had a greater risk of HBV infection; those who always wore face masks were at lower risk (OR 0.2, 95% CI 0.02-0.98). These data indicate that among Berlin dentists, the HCV risk was lower than that of HBV and that face masks may have lowered the risk of HBV. The use of eye glasses or gloves did not appear to lower the risk of HBV acquisition in this population. PMID- 11117966 TI - Is cytomegalovirus infection a co-factor in HIV-1 disease progression? AB - The influence of cytomegalovirus (CMV) infection as a co-factor in HIV-1 disease progression has mainly been studied in haemophiliacs and remains controversial. Based on the files of 1683 HIV-1-infected patients in the Seropositive Cohort (SEROCO) and Haemophiliacs Cohort (HEMOCO) cohorts, we studied the role of CMV infection in progression to CD4+ cell counts of less than 200 microl, AIDS onset and death, in various HIV exposure groups. Adjusted relative risk (aRR) of progression to AIDS and to death was respectively 1.30 (P = 0.05) and 1.58 (P = 0.007). In the sexual exposure group the influence of CMV infection on the risk of progression to AIDS was of borderline significance (aRR = 1.50; P = 0.07) and was more marked on the risk of death (aRR = 2.00; P = 0.03). No such influence of CMV infection was observed in the transfusion and intravenous drug use exposure groups. When we studied the influence of CMV infection according to the stage of HIV disease, the main effect was on progression from AIDS to death, probably because CMV disease is a late event. Sexual CMV transmission and frequent re exposure to CMV may explain why CMV infection emerged as an important co-factor for HIV progression only in the sexual exposure group. PMID- 11117967 TI - Prevalence of Eperythrozoon spp. infection and congenital eperythrozoonosis in humans in Inner Mongolia, China. AB - Eperythrozoon is an obligate parasitic bacteria found in many species of animals. A large scale investigation of the prevalence of Eperythrozoon spp. in humans, was conducted in a developing country using light, electron microscope and animal inoculation. Samples were collected in undeveloped areas of Inner Mongolia in China over a 2-year period of 1994-6. Of the 1529 investigated samples, 35.3% were found to be Eperythrozoon spp. positive. The prevalence of infection was associated with occupation and seasonal variations. The infections were mainly mild, in 89.6% of cases (excluding pregnant women and their children). Of 74 pregnant women tested in the areas of high prevalence, 44 were confirmed Eperythrozoon spp. positive. Similarly, eperythrozoa were found in all 44 umbilical cords tested and in the neonatal peripheral blood samples taken at birth. These data suggest that eperythrozoa can be transmitted via the placenta. PMID- 11117968 TI - PCR-based detection of chlamydial infection in swine and subsequent PCR-coupled genotyping of chlamydial omp1-gene amplicons by DNA-hybridization, RFLP-analysis, and nucleotide sequence analysis. AB - Lung and intestine of 49 pigs with respiratory diseases and endocervical swabs from 205 sows with reproductive disorders were investigated for chlamydial infection by polymerase chain reaction. PCR primers targeted DNA sequences on the chlamydial omp1 or omp2 genes. PCR amplicons were generated from 49.0% of pigs with respiratory disease, from 60.0% of sows with reproductive disorders, from 24.5% of respiratory healthy controls, but from no endocervical swabs from fertile sows. By DNA hybridization, a high prevalence of mixed infections with Chlamydophila abortus and Chlamydia suis in the porcine lung and intestine was found and confirmed by RFLP and nucleotide analysis. Of the omp1-PCR amplicons from endocervical swabs 81.3% were identified as Chlamydophila abortus, indicating an association of this chlamydial species with reproductive disorders in sows. Nucleotide sequence analysis of omp1-amplicons identified as deriving from Chlamydia suis shared a maximum of 82.7% homology with the reference strain S45. PMID- 11117969 TI - A long-term study of Rattus norvegicus in the London borough of Enfield using baiting returns as an indicator of sewer population levels. AB - This is a long-term study that investigates the dynamics of a population of Rattus norvegicus (Berk) inhabiting a sewerage system in London. Thirteen years (1986/7-1998/9) of data from sewer baiting records were analysed (a total of 35,478 records). Manholes were baited with the anticoagulant Brodifacoum (0.005%) on a pinhead oatmeal bait base. Time series analysis was conducted on the data set to determine the underlying trend of the data and the population fluctuations about this trend. An exponential curve was found to give an accurate and realistic fit to the data and indicated that the rat population had decreased over the study period. Decomposition analysis indicated a 5-year cycle best described fluctuations around this trend. PMID- 11117970 TI - Prevalence of antibodies to Brucella spp. in cattle, sheep, goats, horses and camels in the State of Eritrea; influence of husbandry systems. AB - Samples from 2427 cattle, 661 goats, 104 sheep, 98 camels and 82 horses were screened for brucella infections by the Rose Bengal Test and positive reactors confirmed by the complement fixation test. In cattle, the highest individual seroprevalence was in dairy herds kept under the intensive husbandry system, with an individual prevalence of 8.2% and unit (herd) seroprevalence of 35.9%. This was followed by the pastoral husbandry system in the Western Lowlands with 5.0% individual but a higher unit (vaccination site) prevalence of 46.1%. The lowest was in the mixed crop-livestock system in the Southern Highlands with individual 0.3% and unit (village) prevalence of 2.4%. In sheep and goats, no positive animals were detected in the mixed crop-livestock areas. In the Eastern Lowlands individual prevalences of 3.8% (goats) and 1.4% (sheep) and unit prevalence of 33.3% (goats) and 16.7% were found, while 14.3% of individual goats and 56.3% of the units in the Western Lowlands were positive. No positive horses were found. The present study documents the first serological evidence of Brucella spp. infection in camels (3.1%) in Eritrea. PMID- 11117971 TI - Presence of enteroviruses and reoviruses in the waters of the Italian coast of the Adriatic Sea. AB - EEC directive 76/160 requires member states to apply microbiological and chemico physical standards for the quality of recreational waters. In observation of this regulation, in the present study 144 samples of seawater were taken over a 12 month period and tested to determine viral contamination. The samples were collected from the coastal waters of the Italian town of Pesaro, which is located on the Adriatic Sea. Using cell culture techniques, 32.6% of the seawater samples were found to be contaminated with enteroviruses. Isolation of these viruses was most frequent in the summer months. Thus, our results indicate the need to increase the frequency of monitoring of these waters and to eliminate the sources of contamination. PMID- 11117972 TI - Tuberculosis drug resistance in England and Wales. How much is 'home-grown'? AB - The fact that a substantial proportion of tuberculosis drug resistance (especially multidrug resistance) is due to transmission of resistant strains or treatment failure in the UK underlines the need to strengthen control in this country. Early identification and effective treatment of cases (including measures to support adherence and appropriate use of initial treatment regimes involving at least four drugs in those at risk of isoniazid resistance) would help to control the problem [1]. PMID- 11117973 TI - Phenylpropanolamine and the risk of hemorrhagic stroke. AB - BACKGROUND: Phenylpropanolamine is commonly found in appetite suppressants and cough or cold remedies. Case reports have linked the use of products containing phenylpropanolamine to hemorrhagic stroke, often after the first use of these products. To study the association, we designed a case-control study. METHODS: Men and women 18 to 49 years of age were recruited from 43 U.S. hospitals. Eligibility criteria included the occurrence of a subarachnoid or intracerebral hemorrhage within 30 days before enrollment and the absence of a previously diagnosed brain lesion. Random-digit dialing identified two matched control subjects per patient. RESULTS: There were 702 patients and 1376 control subjects. For women, the adjusted odds ratio was 16.58 (95 percent confidence interval, 1.51 to 182.21; P=0.02) for the association between the use of appetite suppressants containing phenylpropanolamine and the risk of a hemorrhagic stroke and 3.13 (95 percent confidence interval, 0.86 to 11.46; P=0.08) for the association with the first use of a product containing phenylpropanolamine. All first uses of phenylpropanolamine involved cough or cold remedies. For men and women combined, the adjusted odds ratio was 1.49 (95 percent confidence interval, 0.84 to 2.64; P=0.17) for the association between the use of a product containing phenylpropanolamine and the risk of a hemorrhagic stroke, 1.23 (95 percent confidence interval, 0.68 to 2.24; P=0.49) for the association with the use of cough or cold remedies that contained phenylpropanolamine, and 15.92 (95 percent confidence interval, 1.38 to 184.13; P=0.03) for the association with the use of appetite suppressants that contained phenylpropanolamine. An analysis in men showed no increased risk of a hemorrhagic stroke in association with the use of cough or cold remedies containing phenylpropanolamine. No men reported the use of appetite suppressants. CONCLUSIONS: The results suggest that phenylpropanolamine in appetite suppressants, and possibly in cough and cold remedies, is an independent risk factor for hemorrhagic stroke in women. PMID- 11117974 TI - Adverse cardiovascular and central nervous system events associated with dietary supplements containing ephedra alkaloids. AB - BACKGROUND: Dietary supplements that contain ephedra alkaloids (sometimes called ma huang) are widely promoted and used in the United States as a means of losing weight and increasing energy. In the light of recently reported adverse events related to use of these products, the Food and Drug Administration (FDA) has proposed limits on the dose and duration of use of such supplements. The FDA requested an independent review of reports of adverse events related to the use of supplements that contained ephedra alkaloids to assess causation and to estimate the level of risk the use of these supplements poses to consumers. METHODS: We reviewed 140 reports of adverse events related to the use of dietary supplements containing ephedra alkaloids that were submitted to the FDA between June 1, 1997, and March 31, 1999. A standardized rating system for assessing causation was applied to each adverse event. RESULTS: Thirty-one percent of cases were considered to be definitely or probably related to the use of supplements containing ephedra alkaloids, and 31 percent were deemed to be possibly related. Among the adverse events that were deemed definitely, probably, or possibly related to the use of supplements containing ephedra alkaloids, 47 percent involved cardiovascular symptoms and 18 percent involved the central nervous system. Hypertension was the single most frequent adverse effect (17 reports), followed by palpitations, tachycardia, or both (13); stroke (10); and seizures (7). Ten events resulted in death, and 13 events produced permanent disability, representing 26 percent of the definite, probable, and possible cases. CONCLUSIONS: The use of dietary supplements that contain ephedra alkaloids may pose a health risk to some persons. These findings indicate the need for a better understanding of individual susceptibility to the adverse effects of such dietary supplements. PMID- 11117975 TI - Caffeine intake and the risk of first-trimester spontaneous abortion. AB - BACKGROUND: Some epidemiologic studies have suggested that the ingestion of caffeine increases the risk of spontaneous abortion, but the results have been inconsistent. METHODS: We performed a population-based, case-control study of early spontaneous abortion in Uppsala County, Sweden. The subjects were 562 women who had spontaneous abortion at 6 to 12 completed weeks of gestation (the case patients) and 953 women who did not have spontaneous abortion and were matched to the case patients according to the week of gestation (controls). Information on the ingestion of caffeine was obtained from in-person interviews. Plasma cotinine was measured as an indicator of cigarette smoking, and fetal karyotypes were determined from tissue samples. Multivariate analysis was used to estimate the relative risks associated with caffeine ingestion after adjustment for smoking and symptoms of pregnancy such as nausea, vomiting, and tiredness. RESULTS: Among nonsmokers, more spontaneous abortions occurred in women who ingested at least 100 mg of caffeine per day than in women who ingested less than 100 mg per day, with the increase in risk related to the amount ingested (100 to 299 mg per day: odds ratio, 1.3; 95 percent confidence interval, 0.9 to 1.8; 300 to 499 mg per day: odds ratio, 1.4; 95 percent confidence interval, 0.9 to 2.0; and 500 mg or more per day: odds ratio, 2.2; 95 percent confidence interval, 1.3 to 3.8). Among smokers, caffeine ingestion was not associated with an excess risk of spontaneous abortion. When the analyses were stratified according to the results of karyotyping, the ingestion of moderate or high levels of caffeine was found to be associated with an excess risk of spontaneous abortion when the fetus had a normal or unknown karyotype but not when the fetal karyotype was abnormal. CONCLUSIONS: The ingestion of caffeine may increase the risk of an early spontaneous abortion among non-smoking women carrying fetuses with normal karyotypes. PMID- 11117976 TI - Prognosis of cancers associated with venous thromboembolism. AB - BACKGROUND: Little is known about the prognosis of cancer discovered during or after an episode of venous thromboembolism. METHODS: We linked the Danish National Registry of Patients, the Danish Cancer Registry, and the Danish Mortality Files to obtain data on the survival of patients who received a diagnosis of cancer at the same time as or after an episode of venous thromboembolism. Their survival was compared with that of patients with cancer who did not have venous thromboembolism (control patients), who were matched in terms of type of cancer, age, sex, and year of diagnosis. RESULTS: Of 668 patients who had cancer at the time of an episode of deep venous thromboembolism, 44.0 percent of those with data on the spread of disease (563 patients) had distant metastasis, as compared with 35.1 percent of 5371 control patients with data on spread (prevalence ratio, 1.26; 95 percent confidence interval, 1.13 to 1.40). In the group with cancer at the time of venous thromboembolism, the one year survival rate was 12 percent, as compared with 36 percent in the control group (P<0.001), and the mortality ratio for the entire follow-up period was 2.20 (95 percent confidence interval, 2.05 to 2.40). Patients in whom cancer was diagnosed within one year after an episode of venous thromboembolism had a slightly increased risk of distant metastasis at the time of the diagnosis (prevalence ratio, 1.23 [95 percent confidence interval, 1.08 to 1.40]) and a relatively low rate of survival at one year (38 percent, vs. 47 percent in the control group; P<0.001). CONCLUSIONS: Cancer diagnosed at the same time as or within one year after an episode of venous thromboembolism is associated with an advanced stage of cancer and a poor prognosis. PMID- 11117977 TI - Transmission of hepatitis C virus from a patient to an anesthesiology assistant to five patients. PMID- 11117978 TI - Images in clinical medicine. Trousseau's sign. PMID- 11117979 TI - Syncope. PMID- 11117980 TI - Hyperparathyroid and hypoparathyroid disorders. PMID- 11117981 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 39-2000. A 47-year-old woman with multilobar pulmonary consolidation. PMID- 11117982 TI - The FDA, regulation, and the risk of stroke. PMID- 11117983 TI - Inhibition of food intake by inhibitors of fatty acid synthase. PMID- 11117984 TI - Coronary heart disease in women. PMID- 11117985 TI - Coronary heart disease in women. PMID- 11117986 TI - Coronary heart disease in women. PMID- 11117987 TI - Coronary heart disease in women. PMID- 11117988 TI - Pravastatin therapy and the risk of stroke. PMID- 11117989 TI - Pravastatin therapy and the risk of stroke. PMID- 11117990 TI - Pravastatin therapy and the risk of stroke. PMID- 11117991 TI - Prolongation of the QT interval and SIDS. PMID- 11117992 TI - Prolongation of the QT interval and SIDS. PMID- 11117993 TI - Prolongation of the QT interval and SIDS. PMID- 11117995 TI - Proceedings of the 20th Annual Conference on Peritoneal Dialysis. San Francisco, California, USA. February 2000. PMID- 11117994 TI - Secondary acute myeloid leukemia after treatment of acute monoblastic leukemia. PMID- 11117996 TI - Organizing glucose disposal: emerging roles of the glycogen targeting subunits of protein phosphatase-1. AB - Glucose is stored in mammalian tissues in the form of glycogen. Glycogen levels are markedly reduced in liver or muscle cells of patients with insulin-resistant or insulin-deficient forms of diabetes, suggesting that impaired glycogen synthesis may contribute to development of hyperglycemia. Recently, interest in this area has been further stimulated by new insights into the spatial organization of metabolic enzymes within cells and the importance of such organization in regulation of glycogen metabolism. It is now clear that a four member family of glycogen targeting subunits of protein phosphatase-1 (PP1) plays a major role in coordinating these events. These proteins target PP1 to the glycogen particle and also bind differentially to glycogen synthase, glycogen phosphorylase, and phosphorylase kinase, thereby serving as molecular scaffolds. Moreover, the various glycogen-targeting subunits have distinct tissue expression patterns and can influence regulation of glycogen metabolism in response to glycogenic and glycogenolytic signals. The purpose of this article is to summarize new insights into the structure, function, regulation, and metabolic effects of the glycogen-targeting subunits of PP1 and to evaluate the possibility that these molecules could serve as therapeutic targets for lowering of blood glucose in diabetes. PMID- 11117998 TI - Baculovirus-mediated gene transfer into pancreatic islet cells. AB - Baculovirus transduction is a gene transfer method that uses a moth cell virus for mammalian cells in culture, which results in a high-level prolonged expression. Here we demonstrate that recombinant baculoviruses can serve as efficient gene transfer vehicles for delivering foreign genes driven by mammalian promoters into human and mouse pancreatic islet cells. Existing methods, such as various transfection and electroporation techniques, either suffer from low efficiency or cause extensive membrane damage. Viral vectors have emerged as an important tool for gene delivery and expression in mammalian cells but suffer from several drawbacks, such as lengthy construction time and expression of viral genes. The baculovirus Autographa californica multiple nuclear polyhedrosis virus is widely used as a vector for expression of foreign genes in insect cells and, more recently, in some mammalian cells. Using several green fluorescent protein- and LacZ-expressing constructs in a cytomegalovirus promoter cassette, we obtained efficient gene expression in primary human and mouse islet cells. There was no impairment of glucose-stimulated intracellular free calcium responses after baculovirus infection. The safety and the relative ease of construction and propagation of the virus makes the baculovirus system a useful tool for facilitating the transfer of foreign genes. PMID- 11117997 TI - Activation of glucose transport by AMP-activated protein kinase via stimulation of nitric oxide synthase. AB - Glucose transport in skeletal muscle is stimulated by two distinct stimuli, insulin and exercise. The mechanism by which exercise stimulates glucose transport is not known, although it is distinct from the insulin-mediated pathway. Recently, it has been shown that AMP-activated protein kinase (AMPK) is activated by exercise in skeletal muscle, whereas pharmacological activation of AMPK by 5-amino-4-imidazolecarboxamide riboside (AICAR) leads to increased glucose transport. It has been postulated, therefore, that AMPK may be the link between exercise and glucose transport. To address this, we have examined the signaling pathway involved in the stimulation of glucose uptake after activation of AMPK. Here we show that activation of AMPK by AICAR in rat muscle and mouse H 2Kb muscle cells activates glucose transport approximately twofold. AMPK in H-2Kb cells is also activated by hyperosmotic stress and the mitochondrial uncoupling agent, dinitrophenol, both of which lead to increased glucose transport. In contrast, insulin, which activates glucose transport two- to-threefold in both rat muscle and H-2Kb cells, has no effect on AMPK activity. A previous study has shown that AMPK phosphorylates and activates endothelial nitric oxide synthase (NOS). We show here that NOS activity in H-2Kb cells is activated after stimulation of AMPK by AICAR. Treatment of H-2Kb cells or rat muscle with NOS inhibitors completely blocks the increase in glucose transport after activation of AMPK. In addition, an inhibitor of guanylate cyclase also blocks activation of glucose transport by AICAR in H-2Kb cells. These results indicate that activation of AMPK in muscle cells stimulates glucose transport by activation of NOS coupled to downstream signaling components, including cyclic GMP. PMID- 11117999 TI - Early graft failure of xenogeneic islets in NOD mice is accompanied by high levels of interleukin-1 and low levels of transforming growth factor-beta mRNA in the grafts. AB - Early graft failure, graft rejection, and autoimmune recurrence remain unresolved issues in islet xenotransplantation in type 1 diabetes. The first aim of this study was to examine the existence of early graft failure in spontaneously diabetic autoimmune NOD mice after rat islet transplantation under technically controlled circumstances. The second aim was to examine the mediators of this early xenograft dysfunction. First, we demonstrated a higher percentage of early xenograft failure (48%) in spontaneously diabetic NOD mice as compared with chemically diabetic old NOD (13%, P < 0.05) and C57Bl/6 (7%, P < 0.01) mice. In addition, in spontaneously diabetic NOD mice, xenogeneic islets displayed early graft failure more frequently than allogeneic (23%, P < or = 0.05) or isogeneic islets (7%, P < 0.01). No early graft failure was observed in allotransplantation or isotransplantation in chemically diabetic mice. Reverse transcriptase polymerase chain reaction analysis of cytokine mRNA in islet xenografts 8 h after transplantation showed higher levels of interleukin (IL)-1 mRNA in autoimmune diabetic mice compared with chemically diabetic old NOD mice (1.40 +/- 0.32 vs. 0.90 +/- 0.14 IL-1 copies/beta-actin copies, P < 0.05). In contrast, mRNA levels of transforming growth factor (TGF)-beta were lower in spontaneously diabetic NOD mice than in chemically diabetic old NOD mice (0.67 +/- 0.16 vs. 1.36 +/- 0.50 TGF-beta copies/beta-actin copies, P < 0.05). No differences in tumor necrosis factor-alpha, IL-6, and inducible nitric oxide synthase were seen between autoimmune and nonautoimmune diabetic mice. T-cell cytokines (IL-2, IL-4, IL-10, and gamma-interferon) were absent in all mice until 48 h after transplantation. These data suggest that early islet xenograft failure is more common in spontaneously diabetic NOD mice and could be due to a nonspecific inflammatory reaction locally in the grafts. PMID- 11118000 TI - Suppression and acceleration of autoimmune diabetes by neutralization of endogenous interleukin-12 in NOD mice. AB - A corpus of evidence suggests that T-helper type 1 (Th1)-dependent cellular immunity plays a pivotal role in the pathogenesis of autoimmune diabetes. This study was intended to find ways to prevent the development of NOD diabetes using a neutralizing anti-interleukin (IL)-12 antibody (C17.8) that inhibits Thl cell differentiation. When C17.8 was administered from 5 to 30 weeks of age, NOD mice exhibited suppression of both insulitis and diabetes. However, when C17.8 administration ceased at 15 weeks of age, 8 of 13 recipients showed diabetes at 30 weeks of age. These results suggest that IL-12 plays an important role not only in the development of effector cells but also in their activation. In contrast, when C17.8 was injected into 2-week-old female NOD mice for 6 consecutive days, all 16 recipients showed diabetes at 30 weeks of age, whereas 12 of 20 control mice became diabetic. This result suggests that depletion of endogenous IL-12 at a young age results in the enhancement of diabetes. Flow cytometric analysis indicated that activated memory T-cells were present in higher numbers after C17.8 treatment. Transfer of spleen cells from 15-week-old C17.8-treated NOD mice to NOD-scid mice resulted in an earlier onset and a higher incidence of diabetes. Furthermore, administration of C17.8 to 2-week-old NOD mice also resulted in a much earlier onset of diabetes. These results suggest that short-term treatment with anti-IL-12 antibody prohibits IL-2 production at a young age, which may influence the expansion and apoptosis of pathogenic T-cells, resulting in the acceleration of autoimmune diabetes. PMID- 11118001 TI - Interferon-gamma receptor signaling is dispensable in the development of autoimmune type 1 diabetes in NOD mice. AB - There have been two previous conflicting reports that the development of T-cell mediated autoimmune diabetes (type 1 diabetes) was respectively unaffected or inhibited in NOD mice genetically deficient in the T-helper (Th) 1 cytokine interferon (IFN)-gamma or the alpha-chain subunit of its receptor. Our goal was to resolve this conundrum by congenically transferring, from a 129 donor strain to the NOD background, a functionally inactivated gene for the beta-chain signaling (located on chromosome 16) rather than the alpha-chain ligand binding domain (located on chromosome 10) of the IFN-gamma receptor. These NOD.IFNgammaRBnull mice were characterized by normal patterns of leukocyte development and T-cells that produced greatly enhanced levels of the putatively type 1 diabetes-protective Th2 cytokine interleukin (IL)-4. However, despite being unable to respond to the primary Thl cytokine IFN-gamma and having T-cells that produce greatly enhanced levels of IL-4, NOD.IFNgammaRBnull mice remained highly susceptible to type 1 diabetes. This result indicated that the previously reported inhibition of type 1 diabetes in NOD mice carrying a functionally inactivated IFN-gamma receptor alpha-chain gene may have been due to a closely linked and previously unidentified diabetes resistance allele. Furthermore, our results indicate that the pathogenicity of diabetogenic T-cells in NOD mice is not dampened by an inability to respond to IFN-gamma and a concurrent shift to greatly enhanced Th2 cytokine production. This finding calls into question whether clinical protocols designed to shift beta-cell autoreactive T-cells from a Thl to Th2 cytokine production profile will truly be safe and efficacious in blocking the development of type 1 diabetes in humans. PMID- 11118002 TI - Dehydroepiandrosterone sulfate and beta-cell function: enhanced glucose-induced insulin secretion and altered gene expression in rodent pancreatic beta-cells. AB - Administration of dehydroepiandrosterone (DHEA), or its sulfated form (DHEAS), controls hyperglycemia in diabetic rodents without directly altering insulin sensitivity. We show that DHEAS enhanced glucose-stimulated insulin secretion when administered in vivo to rats or in vitro to beta-cell lines, without changing cellular insulin content. Insulin secretion increased from 3 days of steroid exposure in vitro, suggesting that DHEAS did not directly activate the secretory processes. DHEAS selectively increased the beta-cell mRNA expression of acyl CoA synthetase-2 and peroxisomal acyl CoA oxidase in a time-dependent manner. Although DHEAS is a peroxisomal proliferator, it did not alter the mRNA expression of peroxisomal proliferator-activated receptor (PPAR) alpha or beta, or enhance the activity of transfected PPAR alpha, beta, or gamma in vitro. Thus, DHEAS directly affected the beta-cell to enhance glucose-stimulated insulin secretion and increased the mRNA expression of specific beta-cell mitochondrial and peroxisomal lipid metabolic enzymes. This effect of DHEAS on insulin secretion may contribute to the amelioration of hyperglycemia seen in various rodent models of diabetes. PMID- 11118003 TI - Recombinant human betacellulin promotes the neogenesis of beta-cells and ameliorates glucose intolerance in mice with diabetes induced by selective alloxan perfusion. AB - Betacellulin (BTC), a member of the epidermal growth factor family, is expressed predominantly in the human pancreas and induces the differentiation of a pancreatic acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC has a physiologically important role in the endocrine pancreas. In this study, we examined the in vivo effect of recombinant human BTC (rhBTC) on glucose intolerance and pancreatic morphology using a new mouse model with glucose intolerance induced by selective alloxan perfusion. RhBTC (1 microg/g body wt) or saline was injected subcutaneously every day from the day after alloxan treatment. The intraperitoneal glucose tolerance test revealed no difference between rhBTC-treated and rhBTC-untreated glucose-intolerant mice at 2-4 weeks. However, glucose tolerance was significantly improved and body weight was significantly increased in rhBTC-treated mice compared with untreated mice at 8 weeks. Islet-like cell clusters, consisting mainly of beta-cells, were increased in the pancreas and were localized in contact with the ductal lining cells and sometimes with acinar cells. In conclusion, administration of rhBTC improved glucose tolerance in this mouse model by increasing beta-cell volume, primarily through accelerated neogenesis from ductal lining cells. PMID- 11118004 TI - Differential patterns of glucose-induced electrical activity and intracellular calcium responses in single mouse and rat pancreatic islets. AB - Although isolated rat islets are widely used to study in vitro insulin secretion and the underlying metabolic and ionic processes, knowledge on the properties of glucose-induced electrical activity (GIEA), a key step in glucose-response coupling, has been gathered almost exclusively from microdissected mouse islets. Using a modified intracellular recording technique, we have now compared the patterns of GIEA in collagenase-isolated rat and mouse islets. Resting membrane potentials of rat and mouse beta-cells were approximately -50 and -60 mV, respectively. Both rat and mouse beta-cells displayed prompt membrane depolarizations in response to glucose. However, whereas the latter exhibited a bursting pattern consisting of alternating hyperpolarized and depolarized active phases, rat beta-cells fired action potentials from a nonoscillating membrane potential at all glucose concentrations (8.4-22.0 mmol/l). This was mirrored by changes in the intracellular Ca2+ concentration ([Ca2+]i), which was oscillatory in mouse and nonoscillatory in rat islets. Stimulated rat beta-cells were strongly hyperpolarized by diazoxide, an activator of ATP-dependent K+ channels. Glucose evoked dose-dependent depolarizations and [Ca2+]i increases in both rat (EC50 5.9-6.9 mmol/l) and mouse islets (EC50 8.3-9.5 mmol/l), although it did not affect the burst plateau potential in the latter case. We conclude that there are important differences between beta-cells from both species with respect to early steps in the stimulus-secretion coupling cascade based on the following findings: 1) mouse beta-cells have a larger resting K+ conductance in 2 mmol/l glucose, 2) rat beta-cells lack the compensatory mechanism responsible for generating membrane potential oscillations and holding the depolarized plateau potential in mouse beta-cells, and 3) the electrical and [Ca2+]i dose-response curves in rat beta-cells are shifted toward lower glucose concentrations. Exploring the molecular basis of these differences may clarify several a priori assumptions on the electrophysiological properties of rat beta-cells, which could foster the development of new working models of pancreatic beta-cell function. PMID- 11118005 TI - Hedgehog signaling regulation of insulin production by pancreatic beta-cells. AB - Hedgehogs (Hhs) are intercellular signaling molecules that regulate tissue patterning in mammalian development. Mammalian Hhs include Sonic hedgehog (Shh), Indian hedgehog (Ihh), and Desert hedgehog (Dhh). The absence of Shh expression is required for the early development of the endocrine and exocrine pancreas, but whether Hh signaling functions in the fully developed adult endocrine pancreas is unknown. Here we report that Hhs Ihh and Dhh and their receptors patched (Ptc) and smoothened are expressed in the endocrine islets of Langerhans of the fully developed rat pancreas and in the clonal gamma-cell line INS-1. We demonstrate the coexpression of Ptc with insulin in beta-cells of mouse pancreatic islets, indicating that beta-cells are targets of active Hh signaling. The administration of cyclopamine, a Hh signaling inhibitor, decreases both insulin secretion from and insulin content of INS-1 cells. The effects of Hh signaling on insulin production occur at the transcriptional level because activation of Hh signal transduction by ectopic expression of Shh increases rat insulin I promoter activation in a dose-dependent manner in transient transfections of INS-1 and MIN6 beta-cell lines. In contrast, inhibition of Hh signaling with increasing concentrations of cyclopamine progressively reduces insulin promoter activity. Furthermore, the treatment of INS-1 cells with cyclopamine diminishes endogenous insulin mRNA expression. We propose that Hh signaling is not restricted to patterning in early pancreas development but also continues to signal in differentiated beta-cells of the endocrine pancreas in regulating insulin production. Thus, defective Hh signaling in the pancreas should be considered as a potential factor in the pathogenesis of type 2 diabetes. PMID- 11118006 TI - Subcellular localization, mobility, and kinetic activity of glucokinase in glucose-responsive insulin-secreting cells. AB - We investigated the subcellular localization, mobility, and activity of glucokinase in MIN6 cells, a glucose-responsive insulin-secreting beta-cell line. Glucokinase is present in the cytoplasm and a vesicular/granule compartment that is partially colocalized with insulin granules. The granular staining of glucokinase is preserved after permeabilization of the cells with digitonin. There was no evidence for changes in distribution of glucokinase between the cytoplasm and the granule compartment during incubation of the cells with glucose. The rate of release of glucokinase and of phosphoglucoisomerase from digitonin-permeabilized cells was slower when cells were incubated at an elevated glucose concentration (S0.5 approximately 15 mmol/l). This effect of glucose was counteracted by competitive inhibitors of glucokinase (5-thioglucose and mannoheptulose) but was unaffected by fructose analogs and may be due to changes in cell shape or conformation of the cytoskeleton that are secondary to glucose metabolism. Based on the similar release of glucokinase and phosphoglucoisomerase, we found no evidence for specific binding of cytoplasmic digitonin-extractable glucokinase. The affinity of beta-cells for glucose is slightly lower than that in cell extracts and, unlike that in hepatocytes, is unaffected by fructose, tagatose, or a high-K+ medium, which is consistent with the lack of change in glucokinase distribution or release. We conclude that glucokinase is present in two locations, cytoplasm and the granular compartment, and that it does not translocate between them. This conclusion is consistent with the lack of adaptive changes in the glucose phosphorylation affinity. The glucokinase activity associated with the insulin granules may have a role in either direct or indirect coupling between glucose phosphorylation and insulin secretion. PMID- 11118007 TI - beta-cell glucokinase deficiency and hyperglycemia are associated with reduced islet amyloid deposition in a mouse model of type 2 diabetes. AB - Type 2 diabetes is characterized by impaired beta-cell function, hyperglycemia, and islet amyloid deposition. The primary constituent of islet amyloid is the 37 amino acid beta-cell product called islet amyloid polypeptide (IAPP) or amylin. To study mechanisms of islet amyloid formation, we developed a transgenic mouse model that produces and secretes the amyloidogenic human IAPP (hIAPP) molecule and have shown that 81% of male transgenic mice develop islet amyloid after 14 months on a high-fat diet. To test whether impaired beta-cell function and hyperglycemia could enhance islet amyloid formation, we cross-bred our hIAPP transgenic mice with beta-cell glucokinase-knockout mice (GKKO) that have impaired glucose-mediated insulin secretion and fasting hyperglycemia. The resulting new (hIAPPxGKKO) line of mice had higher basal plasma glucose concentrations than the hIAPP transgenic mice at 3, 6, and 12 months of age (P < 0.05), as did GKKO mice compared with hIAPP transgenic mice at 6 and 12 months of age (P < 0.05). Basal plasma immunoreactive insulin (IRI) levels were lower in hIAPP x GKKO mice than in hIAPP transgenic mice at 6 months of age (P < 0.05). The area under the glucose curve in response to an intraperitoneal glucose challenge (1 g/kg body weight) was larger in hIAPPxGKKO mice than in hIAPP transgenic mice at 3, 6, and 12 months of age (P < 0.005) and in GKKO mice compared with hIAPP transgenic mice at 6 and 12 months of age (P < 0.005). The area under the IRI curve was lower in hIAPPxGKKO mice at 6 and 12 months of age (P < 0.05) than in hIAPP transgenic mice and in GKKO mice compared with hIAPP transgenic mice at 12 months of age (P < 0.05). Despite the presence of hyperglycemia, hIAPPxGKKO mice had a lower incidence (4 of 17 vs. 12 of 19, P < 0.05) and amount (0.40 +/- 0.24 vs. 1.2 +/- 0.3 arbitrary units, P < 0.05) of islet amyloid than hIAPP transgenic mice had. As expected, no islet amyloid was observed in GKKO mice lacking the hIAPP transgene (0 of 13). There was no difference in pancreatic content of IRI and hIAPP among the three groups of mice. Thus, despite the presence of impaired islet function and hyperglycemia, hIAPPxGKKO mice had a decreased incidence and quantity of islet amyloid. Therefore, our data suggest that impaired beta-cell glucose metabolism or hyperglycemia are not likely to contribute to islet amyloid formation in diabetes. Furthermore, this finding may explain the lack of progression of glycemia in patients with maturity-onset diabetes of the young. PMID- 11118008 TI - Mechanism by which metformin reduces glucose production in type 2 diabetes. AB - To examine the mechanism by which metformin lowers endogenous glucose production in type 2 diabetic patients, we studied seven type 2 diabetic subjects, with fasting hyperglycemia (15.5 +/- 1.3 mmol/l), before and after 3 months of metformin treatment. Seven healthy subjects, matched for sex, age, and BMI, served as control subjects. Rates of net hepatic glycogenolysis, estimated by 13C nuclear magnetic resonance spectroscopy, were combined with estimates of contributions to glucose production of gluconeogenesis and glycogenolysis, measured by labeling of blood glucose by 2H from ingested 2H2O. Glucose production was measured using [6,6-2H2]glucose. The rate of glucose production was twice as high in the diabetic subjects as in control subjects (0.70 +/- 0.05 vs. 0.36 +/- 0.03 mmol x m(-2) min(-1), P < 0.0001). Metformin reduced that rate by 24% (to 0.53 +/- 0.03 mmol x m(-2) x min(-1), P = 0.0009) and fasting plasma glucose concentration by 30% (to 10.8 +/- 0.9 mmol/l, P = 0.0002). The rate of gluconeogenesis was three times higher in the diabetic subjects than in the control subjects (0.59 +/- 0.03 vs. 0.18 +/- 0.03 mmol x m(-2) min(-1) and metformin reduced that rate by 36% (to 0.38 +/- 0.03 mmol x m(-2) x min(-1), P = 0.01). By the 2H2O method, there was a twofold increase in rates of gluconeogenesis in diabetic subjects (0.42 +/- 0.04 mmol m(-2) x min(-1), which decreased by 33% after metformin treatment (0.28 +/- 0.03 mmol x m(-2) x min(-1), P = 0.0002). There was no glycogen cycling in the control subjects, but in the diabetic subjects, glycogen cycling contributed to 25% of glucose production and explains the differences between the two methods used. In conclusion, patients with poorly controlled type 2 diabetes have increased rates of endogenous glucose production, which can be attributed to increased rates of gluconeogenesis. Metformin lowered the rate of glucose production in these patients through a reduction in gluconeogenesis. PMID- 11118009 TI - Overexpression of glutamine: fructose-6-phosphate amidotransferase in the liver of transgenic mice results in enhanced glycogen storage, hyperlipidemia, obesity, and impaired glucose tolerance. AB - To examine the effect of increased hexosamine flux in liver, the rate-limiting enzyme in hexosamine biosynthesis (glutamine:fructose-6-phosphate amidotransferase [GFA]) was overexpressed in transgenic mice using the PEPCK promoter. Liver from random-fed transgenic mice had 1.6-fold higher GFA activity compared with nontransgenic control littermates (276 +/- 24 pmol x mg(-1) x min( 1) in transgenic mice vs. 176 +/- 18 pmol x mg(-1) x min(-1) in controls, P < 0.05) and higher levels of the hexosamine end product UDP-N-acetyl glucosamine (288 +/- 11 pmol/g in transgenic mice vs. 233 +/- 10 pmol/g in controls, P < 0.001). Younger transgenic mice compared with control mice had lower fasting serum glucose (4.8 +/- 0.5 mmol/l in transgenic mice vs. 6.5 +/- 0.8 mmol/l in controls, P < 0.05) without higher insulin levels (48.0 +/- 7.8 pmol/l in transgenic mice vs. 56.4 +/- 5.4 pmol/l in controls, P = NS); insulin levels were significantly lower in transgenic males (P < 0.05). At 6 months of age, transgenic animals had normal insulin sensitivity by the hyperinsulinemic clamp technique. Hepatic glycogen content was higher in the transgenic mice (108.6 +/- 5.2 pmol/g in transgenic mice vs. 32.8 +/- 1.3 micromol/g in controls, P < 0.01), associated with an inappropriate activation of glycogen synthase. Serum levels of free fatty acids (FFAs) and triglycerides were also elevated (FFAs, 0.67 +/- 0.03 mmol/l in transgenic mice vs. 0.14 +/- 0.01 in controls; triglycerides, 1.34 +/- 0.15 mmol/l in transgenic mice vs. 0.38 +/- 0.01 in controls, P < 0.01). Older transgenic mice became heavier than control mice and exhibited relative glucose intolerance and insulin resistance. The glucose disposal rate at 8 months of age was 154 +/- 5 mg x kg(-1) x min(-1) in transgenic mice vs. 191 +/- 6 mg x kg(-1) x min(-1) in controls (P < 0.05). We conclude that hexosamines are mediators of glucose sensing for the regulation of hepatic glycogen and lipid metabolism. Increased hexosamine flux in the liver signals a shift toward fuel storage, resulting ultimately in obesity and insulin resistance. PMID- 11118010 TI - Effects of skyrin, a receptor-selective glucagon antagonist, in rat and human hepatocytes. AB - Peptidic glucagon antagonists have been shown to lower blood glucose levels in diabetic models (1-3), but attempts to identify small molecular weight glucagon receptor-binding antagonists have met with little success. Skyrin, a fungal bisanthroquinone, exhibits functional glucagon antagonism by uncoupling the glucagon receptor from adenylate cyclase activation in rat liver membranes (1). We have examined the effects of skyrin on cells transfected with the human glucagon receptor and on isolated rat and human hepatocytes. The skyrin used was isolated from Talaromyces wortmanni American Type Culture Collection 10517. In rat hepatocytes, skyrin (30 micromol/l) inhibited glucagon-stimulated cAMP production (53%) and glucose output (IC50 56 micromol/l). There was no detectable effect on epinephrine or glucagon-like peptide 1 (GLP-1) stimulation of these parameters, which demonstrates skyrin's selective activity. Skyrin was also evaluated in primary cultures of human hepatocytes. Unlike cell lines, which are largely unresponsive to glucagon, primary human hepatocytes exhibited glucagon dependent cAMP production for 14 days in culture (EC50 10 nmol/l). Skyrin (10 micromol/l) markedly reduced glucagon-stimulated cAMP production (55%) and glycogenolysis (27%) in human hepatocytes. The inhibition of glucagon stimulation was a specific property displayed by skyrin and oxyskyrin but not shared by other bisanthroquinones. Skyrin is the first small molecular weight nonpeptidic agent demonstrated to interfere with the coupling of glucagon to adenylate cyclase independent of binding to the glucagon receptor. The data presented in this study indicate that functional uncoupling of the human glucagon receptor from cAMP production results in metabolic effects that could reduce hepatocyte glucose production and hence alleviate diabetic hyperglycemia. PMID- 11118011 TI - Paradoxical effect of troglitazone in normal animals: enhancement of adipocyte but reduction of liver insulin sensitivity. AB - Troglitazone is an antidiabetic agent that improves the ability of adipocytes to store triglycerides by enhancing their insulin sensitivity. Although potent in insulin-resistant states, the effect of troglitazone on lipid and glucose turnover in normal animals has not been assessed. Euglycemic clamps were performed as an insulin dose response in normal mongrel dogs (n = 6). Somatostatin was infused without hormone replacement (zero insulin) for 90 min. Insulin was then either portally replaced (1.8 pmol x min(-1) x kg(-1), overreplaced (5.4 pmol x min(-1) x kg(-1)), or overreplaced peripherally to match the systemic levels of the portal overreplacement dose (2.3 pmol x min(-1) x kg( 1)) for 180 min. A total of 600 mg troglitazone was then given orally each day for 3 weeks and continued throughout a second experimental phase, at which point the euglycemic clamps were repeated. In concordance with previous studies, endogenous glucose production (EGP) was similar whether insulin was delivered portally or peripherally, both before and during troglitazone treatment. Although free fatty acids (FFAs) at zero insulin were not affected, there was a leftward shift of the insulin-FFA dose response curve secondary to a suppression of FFA release into plasma. EGP was paradoxically elevated by troglitazone treatment because of an elevation of both gluconeogenesis and glycogenolysis. In conclusion, troglitazone reduced hepatic sensitivity to FFAs. Because EGP is a primary determinant of fasting blood glucose, we hypothesize that a protective mechanism exists in normal animals, preventing hypoglycemia during insulin sensitization with troglitazone. PMID- 11118012 TI - A high fasting plasma insulin concentration predicts type 2 diabetes independent of insulin resistance: evidence for a pathogenic role of relative hyperinsulinemia. AB - Fasting hyperinsulinemia is a widely used surrogate measure of insulin resistance and predicts type 2 diabetes in various populations. Whether fasting hyperinsulinemia predicts diabetes independent of insulin resistance is unknown. In 319 Pima Indians with normal glucose tolerance, fasting plasma insulin concentration and insulin-stimulated glucose disposal (M) (hyperinsulinemic clamp) were inversely related, but at any given M, there was substantial variation, with some subjects being hyperinsulinemic and others being hypoinsulinemic relative to their degree of insulin sensitivity. In 262 of the 319 subjects followed prospectively over 6.4 +/- 3.9 years, a high fasting plasma insulin concentration was a significant independent predictor of diabetes, in addition to low M and low acute insulin response (AIR) (intravenous glucose challenge). In 161 of the 319 subjects with follow-up measurements of M and AIR (5.1 +/- 3.9 years), a high relative fasting plasma insulin concentration predicted a decline in AIR but not in M before the onset of diabetes. The adjusted fasting plasma insulin concentration was a familial trait (heritability of 0.52) and in a genome-wide scan, there was suggestive evidence of linkage (logarithm of odds score 1.77) to a region on chromosome 3q, which harbors the gene encoding GLUT2. These results provide the first prospective evidence in humans that fasting hyperinsulinemia itself has a primary role in the pathogenesis of diabetes, independent of insulin resistance. Whether amelioration of basal insulin hypersecretion will prevent diabetes remains to be elucidated. PMID- 11118013 TI - Impaired oxidation of plasma-derived fatty acids in type 2 diabetic subjects during moderate-intensity exercise. AB - The present study was intended to investigate the different components of fatty acid utilization during a 60-min period of moderate-intensity cycling exercise (50% of VO2max) in eight male type 2 diabetic subjects (aged 52.6 +/- 3.1 years, body fat 35.8 +/- 1.3%) and eight male obese control subjects (aged 45.1 +/- 1.4 years, body fat 34.2 +/- 1.3%) matched for age, body composition, and maximal aerobic capacity. To quantitate the different components of fatty acid metabolism, an isotope infusion of [U-13C]-palmitate was used in combination with indirect calorimetry. In separate experiments, the 13C label recovery in expired air was determined during infusion of [1,2-13C]-acetate (acetate recovery factor). There were no differences in energy expenditure or carbohydrate and total fat oxidation between the groups. The rate of appearance (Ra) of free fatty acid (FFA) (P < 0.05) and the exercise-induced increase in Ra of FFA were significantly lower (P < 0.05) in type 2 diabetic subjects compared with control subjects (baseline vs. exercise [40-60 min]; type 2 diabetes 11.9 +/- 0.9 vs. 19.6 +/- 2.2 micromol x kg(-1) fat-free mass [FFM] x min(-1) and control 15.8 +/- 1.8 vs. 28.6 +/- 2.1 micromol x kg(-1) FFM x min(-1)). The oxidation of plasma derived fatty acids was significantly lower in type 2 diabetic subjects during both conditions (P < 0.05, baseline vs. exercise [40-60 min]; type 2 diabetes 4.2 +/- 0.5 vs. 14.1 +/- 1.9 micromol x kg(-1) FFM x min(-1) and control 6.2 +/- 0.6 vs. 20.4 +/- 1.9 micromol x kg(-1) FFM x min(-1)), whereas the oxidation of triglyceride-derived fatty acids was higher (P < 0.05). It is hypothesized that these impairments in fatty acid utilization may play a role in the etiology of skeletal muscle and hepatic insulin resistance. PMID- 11118014 TI - Thiazolidinediones inhibit the expression of beta3-adrenergic receptors at a transcriptional level. AB - The effect of the thiazolidinediones (TZDs) darglitazone and troglitazone on beta3-adrenergic receptor (AR) expression was studied in cultured cell lines representing several tissues. After 24 h of exposing HIB-1B brown adipocytes to 30 micromol/l darglitazone or 20 micromol/l troglitazone, beta3-AR mRNA levels were reduced by 75%. This effect was significant within 1 h of exposure to a maximal dose of these drugs, with the full effect obtained within 10 h. The darglitazone ID50 was approximately 10 nmol/l, similar to the Kd of TZDs binding to peroxisome proliferator-activated receptor-gamma (PPAR-gamma). These drugs also decreased beta3-AR mRNA in 3T3-F442A white adipocytes, but not in SK-N-MC cells, which lack PPAR-gamma2. A luciferase reporter gene containing 1.4 kb of 5' flanking sequence of the mouse beta3-AR was transiently transfected, with or without PPAR-gamma2, in SK-N-MC cells. The vigorous expression of luciferase driven by the beta3-AR gene sequence was inhibited by TZDs in a PPAR-gamma2 dependent manner. The half-lives of gamma3-AR precursor RNA and mRNA were short, approximately 40 and approximately 100 min, respectively, and remained unaffected by TZD treatment. Exposure of HIB-1B cells to 30 micromol/l darglitazone was associated with reduced beta3-AR mRNA levels, as well as decreased response of uncoupling protein 1 to norepinephrine + propranolol (a beta1 beta2-AR antagonist) or the specific beta3-AR agonist CL 316, 243. Both the beta3-AR mRNA level and response to these stimuli fully recovered by 24 h of removing the drug, indicating that the beta3-AR protein and its coupling to adenylyl cyclase rapidly followed the changes in mRNA. Thus, TZDs can rapidly reduce beta3-AR expression at the transcriptional level, acting through PPAR-gamma2. The rapid turnover and responses of beta3-AR to perturbations, along with numerous other factors reported to regulate its expression, suggest a tight control of beta3-AR and function. Lastly, leptin being the only other known gene suppressed by TZDs, the present studies support a concerted lipogenic effect of these drugs. PMID- 11118015 TI - Longitudinal compensation for fat-induced insulin resistance includes reduced insulin clearance and enhanced beta-cell response. AB - Central adiposity is highly correlated with insulin resistance, which is an important risk factor for type 2 diabetes and other chronic diseases. However, in normal individuals, central adiposity can be tolerated for many years without development of impaired glucose tolerance or diabetes. Here we examine longitudinally the mechanisms by which glucose tolerance can be maintained in the face of substantial insulin resistance. Normal dogs were fed a diet enriched with moderate amounts of fat (2 g x kg(-1) x day(-1)), similar to that seen in modern "cafeteria" diets, and the time course of metabolic changes in these animals was examined over 12 weeks. Trunk adiposity as assessed by magnetic resonance imaging increased from 12 to 19%, but body weight remained unchanged. Insulin sensitivity (SI) as determined by frequently sampled intravenous glucose tolerance tests was measured over a 12-week period. SI decreased 35% by week 1 and remained impaired for the entire 12 weeks. Intravenous glucose tolerance was reduced transiently for 1 week, recovered to baseline, and then again began to decline after 8 weeks. First-phase insulin response began to increase after week 2, peaked by week 6 (190% of basal), and then declined. The increase in insulin response was due partially to enhanced beta-cell function (22%) but due also to an approximately 50% reduction in insulin clearance. This compensation by insulin clearance was also confirmed with insulin clamps performed in fat-fed versus control dogs. The present study confirms the ability of the normal individual to compensate for fat induced insulin resistance by enhanced insulin response, such that the product of insulin sensitivity x secretion is little changed. However, the compensation is due as much to reduced insulin clearance as increased beta-cell sensitivity to glucose. Reduced hepatic extraction of insulin may be the first line of defense providing a higher proportion of secreted insulin to the periphery and sparing the beta-cells during compensation for the insulin-resistant state. PMID- 11118016 TI - A model to explore the interaction between muscle insulin resistance and beta cell dysfunction in the development of type 2 diabetes. AB - Type 2 diabetes is a polygenic disease characterized by defects in both insulin secretion and insulin action. We have previously reported that isolated insulin resistance in muscle by a tissue-specific insulin receptor knockout (MIRKO mouse) is not sufficient to alter glucose homeostasis, whereas beta-cell-specific insulin receptor knockout (betaIRKO) mice manifest severe progressive glucose intolerance due to loss of glucose-stimulated acute-phase insulin release. To explore the interaction between insulin resistance in muscle and altered insulin secretion, we created a double tissue-specific insulin receptor knockout in these tissues. Surprisingly, betaIRKO-MIRKO mice show an improvement rather than a deterioration of glucose tolerance when compared to betaIRKO mice. This is due to improved glucose-stimulated acute insulin release and redistribution of substrates with increased glucose uptake in adipose tissue and liver in vivo, without a significant decrease in muscle glucose uptake. Thus, insulin resistance in muscle leads to improved glucose-stimulated first-phase insulin secretion from beta-cells and shunting of substrates to nonmuscle tissues, collectively leading to improved glucose tolerance. These data suggest that muscle, either via changes in substrate availability or by acting as an endocrine tissue, communicates with and regulates insulin sensitivity in other tissues. PMID- 11118017 TI - Prandial glucose effectiveness and fasting gluconeogenesis in insulin-resistant first-degree relatives of patients with type 2 diabetes. AB - Impaired glucose effectiveness (i.e., a diminished ability of glucose per se to facilitate its own metabolism), increased gluconeogenesis, and endogenous glucose release are, together with insulin resistance and beta-cell abnormalities, established features of type 2 diabetes. To explore aspects of the pathophysiology behind type 2 diabetes, we assessed in a group of healthy people prone to develop type 2 diabetes (n = 23), namely first-degree relatives of type 2 diabetic patients (FDR), 1) endogenous glucose release and fasting gluconeogenesis measured using the 2H2O technique and 2) glucose effectiveness. The FDR group was insulin resistant when compared with an age-, sex-, and BMI matched control group without a family history of type 2 diabetes (n = 14) (M value, clamp: 6.07 +/- 0.48 vs. 8.06 +/- 0.69 mg x kg(-1) lean body weight (lbw) x min(-1); P = 0.02). Fasting rates of gluconeogenesis (1.28 +/- 0.06 vs. 1.41 +/ 0.07 mg x kg(-1) lbw x min(-1); FDR vs. control subjects, P = 0.18) did not differ in the two groups and accounted for 53 +/- 2 and 60 +/- 3% of total endogenous glucose release. Glucose effectiveness was examined using a combined somatostatin and insulin infusion (0.17 vs. 0.14 mU x kg(-1) x min(-1), FDR vs. control subjects), the latter replacing serum insulin at near baseline levels. In addition, a 360-min labeled glucose infusion was given to simulate a prandial glucose profile. After glucose infusion, the integrated plasma glucose response above baseline (1,817 +/- 94 vs. 1,789 +/- 141 mmol/l per 6 h), the ability of glucose to simulate its own uptake (1.50 +/- 0.13 vs. 1.32 +/- 0.16 ml x kg(-1) lbw x min(-1)), and the ability of glucose per se to suppress endogenous glucose release did not differ between the FDR and control group. In conclusion, in contrast to overt type 2 diabetic patients, healthy people at high risk of developing type 2 diabetes are characterized by normal glucose effectiveness at near-basal insulinemia and normal fasting rates of gluconeogenesis. PMID- 11118018 TI - Pharmacokinetics and pharmacodynamics of subcutaneous injection of long-acting human insulin analog glargine, NPH insulin, and ultralente human insulin and continuous subcutaneous infusion of insulin lispro. AB - To compare the pharmacokinetics/dynamics of the long-acting insulin analog glargine with NPH, ultralente, and continuous subcutaneous (SC) infusion of insulin lispro (continuous subcutaneous insulin infusion [CSII]), 20 C-peptide negative type 1 diabetic patients were studied on four occasions during an isoglycemic 24-h clamp. Patients received SC injection of either 0.3 U/kg glargine or NPH insulin (random sequence, crossover design). On two subsequent occasions, they received either an SC injection of ultralente (0.3 U/kg) or CSII (0.3 U x kg(-1) x 24 h(-1)) (random sequence, crossover design). After SC insulin injection or CSII, intravenous (IV) insulin was tapered, and glucose was infused to clamp plasma glucose at 130 mg/dl for 24 h. Onset of action (defined as reduction of IV insulin >50%) was earlier with NPH (0.8 +/- 0.2 h), CSII (0.5 +/- 0.1 h), and ultralente (1 +/- 0.2 h) versus glargine (1.5 +/- 0.3 h) (P < 0.05) (mean +/- SE). End of action (defined as an increase in plasma glucose >150 mg/dl) occurred later with glargine (22 +/- 4 h) than with NPH (14 +/- 3 h) (P < 0.05) but was similar with ultralente (20 +/- 6 h). NPH and ultralente exhibited a peak concentration and action (at 4.5 +/- 0.5 and 10.1 +/- 1 h, respectively) followed by waning, whereas glargine had no peak but had a flat concentration/action profile mimicking CSII. Interindividual variability (calculated as differences in SD of plasma insulin concentrations and glucose infusion rates in different treatments) was lower with glargine than with NPH and ultralente (P < 0.05) but was similar with glargine and CSII (NS). In conclusion, NPH and ultralente are both peak insulins. Duration of action of ultralente is greater, but intersubject variability is also greater than that of NPH. Glargine is a peakless insulin, it lasts nearly 24 h, it has lower intersubject variability than NPH and ultralente, and it closely mimics CSII, the gold standard of basal insulin replacement. PMID- 11118019 TI - Impaired in vivo stimulation of lipolysis in adipose tissue by selective beta2 adrenergic agonist in obese adolescent girls. AB - Studies performed in adults with long-standing obesity suggest a reduced lipolytic sensitivity to catecholamines in subcutaneous abdominal adipose tissue (AT). We used microdialysis to study the in situ lipolytic effects of dobutamine (selective beta1-agonist) and terbutaline (selective beta2-agonist) on glycerol release (lipolytic index) in abdominal subcutaneous AT in 10 obese girls aged 13 17 years, BMI 38 +/- 2.1 kg/m2, and in 7 lean girls aged 11-17 years, BMI 21 +/- 1.1 kg/m2, and compared them with 10 obese women aged 21-39 years, BMI 36 +/- 1.6 kg/m2, and 10 lean women aged 18-42 years, BMI 21 +/- 0.4 kg/m2. Terbutaline at 10(-6) mol/l stimulated glycerol release more efficiently in lean girls than in obese girls (peak response approximately 350 vs. 150% of control, P < 0.01). At the lower concentration of agonist, no significant difference was seen. In women, terbutaline was more effective in lean than in obese women in stimulating glycerol release at both 10(-8) mol/l (peak response lean approximately 175% vs. obese 125% of control) and 10(-6) mol/l (approximately 300 vs. 150% of control, P < 0.05). No significant difference in glycerol release between obese and lean girls or women was detected with selective beta1-stimulation. Our data demonstrate a specific impairment in the capacity of beta2-adrenergic agonists to promote lipolysis in subcutaneous abdominal adipose tissue of obese adolescent girls and women. Thus, decreased mobilization of fat during activation of the adrenergic system might be present early in the development of adolescent obesity. PMID- 11118020 TI - Exercise and thiazolidinedione therapy normalize insulin action in the obese Zucker fatty rat. AB - Thiazolidinediones and exercise are both known to improve insulin action independently. Therefore, we determined whether combined therapy could normalize insulin action in the Zucker fatty (ZF) rat. Rats were fed troglitazone as a 0.2% food admixture over a 3-week exercise training period (treadmill running 5 days/week, 20 m/min, 0% grade, 60 min/day). Subsequent to drug and/or exercise therapy, animals were chronically cannulated in the carotid artery (sampling) and jugular vein (infusion). After a 4-day recovery from surgery, animals were exposed to a hyperinsulinemic (40 mU x kg(-1) x min(-1)) euglycemic clamp (8.5 +/ 0.12 mmol/l; P = 0.45 between groups). Independently, exercise (n = 7) and troglitazone (n = 7) improved the glucose disposal rate 20% (P = 0.04) and 76% (P = 0.001), respectively, when compared with untreated ZF controls (n = 11). In combination, exercise and troglitazone therapy (n = 6) produced significant increments in the following: tracer-determined glucose disposal rate (combined therapy, 52.4 +/- 2.9 mg x kg(-1) x min(-1), vs. untreated ZF, 25.8 +/- 0.8 mg x kg(-1) x min(-1); P = 0.0001), total GLUT4 protein (twofold increase; P = 0.001), insulin receptor substrate (IRS)-1 protein (fourfold increase; P = 0.0001), and Akt phosphorylation (2.9-fold increase; P = 0.002). In conclusion, 1) exercise and troglitazone therapy each improved insulin action in the ZF rat, whereas the combination of the two led to complete normalization of insulin sensitivity, and 2) combination treatment also resulted in normalization of GLUT4 total protein, IRS-1 protein, and Akt phosphorylation compared with lean littermates. PMID- 11118021 TI - Native and modified LDL activate extracellular signal-regulated kinases in mesangial cells. AB - Glycation and/or oxidation of LDL may promote diabetic nephropathy. The mitogen activated protein kinase (MAPK) cascade, which includes extracellular signal regulated protein kinases (ERKs), modulates cell function. Therefore, we examined the effects of LDL on ERK phosphorylation in cultured rat mesangial cells. In cells exposed to 100 microg/ml native LDL or LDL modified by glycation, and/or mild or marked (copper-mediated) oxidation, ERK activation peaked at 5 min. Five minutes of exposure to 10-100 microg/ml native or modified LDL produced a concentration-dependent (up to sevenfold) increase in ERK activity. Also, 10 microg/ml native LDL and mildly modified LDL (glycated and/or mildly oxidized) produced significantly greater ERK activation than that induced by copper oxidized LDL +/- glycation (P < 0.05). Pretreatment of cells with Src kinase and MAPK kinase inhibitors blocked ERK activation by 50-80% (P < 0.05). Native and mildly modified LDL, which are recognized by the native LDL receptor, induced a transient spike of intracellular calcium. Copper-oxidized (+/- glycation) LDL, recognized by the scavenger receptor, induced a sustained rise in intracellular calcium. The intracellular calcium chelator (EGTA/AM) further increased ERK activation by native and mildly modified LDL (P < 0.05). These findings demonstrate that native and modified LDL activate ERKs 1 and 2, an early mitogenic signal, in mesangial cells and provide evidence for a potential link between modified LDL and the development of glomerular injury in diabetes. PMID- 11118022 TI - Defective intracellular antioxidant enzyme production in type 1 diabetic patients with nephropathy. AB - There is an individual susceptibility to diabetic nephropathy, and oxidative stress is believed to play an important role in the pathogenesis of diabetic complications. Active oxygen species induce antioxidant enzyme expression in tissues, an effect considered to be a defensive mechanism. To test whether altered intracellular antioxidant enzyme production might explain the predisposition to diabetic nephropathy, we studied the effect of long-term (12 weeks) exposure to normal (5 mmol/l) or high (22 mmol/l) glucose concentrations on fibroblast antioxidant enzyme gene expression and protein activity in type 1 diabetic patients with and without nephropathy, nondiabetic nephropathic patients, and nondiabetic control subjects. Under conditions of normal glucose concentration in the culture media, CuZnSuperoxide-dismutase, MnSuperoxide dismutase, catalase, and glutathione-peroxidase activity and mRNA expression were not different among the four groups. Under high-glucose conditions, CuZnSuperoxide-dismutase mRNA and activity increased similarly in all groups (P < 0.001 vs. basal), whereas MnSuperoxide-dismutase did not change. In contrast, catalase mRNA and activity as well as glutathione-peroxidase mRNA and activity increased in fibroblasts from type 1 diabetic patients without nephropathy (P < 0.001), in fibroblasts from nondiabetic nephropathic patients (P < 0.001), and in fibroblasts from nondiabetic control subjects (P < 0.001), but not in fibroblasts from type 1 diabetic patients with nephropathy. Exposure to high glucose concentrations significantly increased lipid peroxidation in cells, higher levels being found in cells from diabetic patients with nephropathy (P < 0.001). These data, while confirming that exposure to high glucose concentrations induces an antioxidant defense in skin fibroblasts from normal subjects, demonstrate a failure of this defensive mechanism in cells from type 1 diabetic patients with nephropathy, whereas skin fibroblasts from diabetic patients without complications or from nondiabetic nephropathic patients have an intact antioxidant response to glucose-induced oxidative stress. PMID- 11118023 TI - Diabetes in the Goto-Kakizaki rat is accompanied by impaired insulin-mediated myosin-bound phosphatase activation and vascular smooth muscle cell relaxation. AB - Our laboratory has demonstrated that insulin rapidly stimulates myosin-bound phosphatase (MBP) activity in vascular smooth muscle cells (VSMCs). In this study, we examined whether diabetes is accompanied by alterations in MBP activation and elucidated the components of the signaling pathway that mediate the effects of diabetes. VSMCs isolated from Goto-Kakizaki (GK) diabetic rats (a model for type 2 diabetes) exhibited marked impairment in MBP activation by insulin that was accompanied by failure of insulin to decrease the phosphorylation of a regulatory myosin-bound subunit (MBS) of MBP and inhibit Rho kinase activity resulting in increased myosin light-chain (MLC)20 phosphorylation and VSMC contraction. In VSMCs isolated from control rats, insulin inactivates Rho kinase and decreases MBS phosphorylation, leading to MBP activation. In addition to this pathway, insulin also appears to activate MBP by stimulating the phosphatidylinositol (PI) 3-kinase/nitric oxide (NO)/cGMP signaling pathway because treatment with the synthetic inhibitors of PI 3-kinase, NO synthase (NOS), and cGMP all blocked insulin's effect on MBP activation, whereas cGMP agonists and sodium nitroprusside (SNP) mimicked insulin's effect on MBP activation. VSMCs from diabetic GK rats exhibit reductions in insulin-mediated induction of inducible NOS protein expression and cGMP generation but normal MBP activation in response to SNP and cGMP agonist. This observation led us to examine the effect of diabetes on the activation status of the upstream insulin signaling components. Although GK diabetes did not affect insulin-stimulated tyrosine phosphorylation of the insulin receptor or its content, insulin stimulated insulin receptor substrate (IRS)-1 tyrosine phosphorylation was severely impaired. This was accompanied by marked reductions in IRS-1-associated PI 3-kinase activity. We conclude that insulin stimulates MBP via its regulatory subunit, MBS, by inactivating Rho kinase and stimulating NO/cGMP signaling via PI 3-kinase as part of a complex signaling network that controls MLC20 phosphorylation and VSMC contraction. Defective signaling via Rho kinase and the IRS-1/PI 3-kinase/NOS/cGMP pathway may mediate the inhibitory effects of hyperglycemia and diabetes on MBP activation in this experimental model. PMID- 11118024 TI - APOE polymorphisms and the development of diabetic nephropathy in type 1 diabetes: results of case-control and family-based studies. AB - The goal of this study was to examine the association between known polymorphisms in the apolipoprotein E gene (APOE) and diabetic nephropathy (DN) in type 1 diabetes. We used both a case-control comparison and a family-based study design known as the transmission/disequilibrium test (TDT). For the case-control comparison, we collected DNA from 223 subjects with clinically diagnosed DN and 196 control subjects with normoalbuminuria and long-duration type 1 diabetes (> or = 15 years). For the family-based study, we obtained DNA from both parents of 154 DN subjects and 81 control subjects. The frequency of the epsilon2 allele of exon 4 of APOE was significantly higher in DN subjects than in control subjects. The risk of DN was 3.1 times higher (95% CI 1.6-5.9) in carriers of this allele than in noncarriers. In the family study, heterozygous parents for the E2 allele preferentially transmitted epsilon2 to DN offspring (64 vs. 36%, P < 0.03). Four additional polymorphisms (i.e., -491 A/T, -219 G/T, IE1 G/C, and APOCI insertion/deletion [I/D]) that flank the APOE locus were not associated with DN in either the case-control comparison or in the family-based study. In conclusion, the results of the case-control as well as the family-based study provide evidence that the epsilon2 allele of APOE increases the risk of DN in type 1 diabetes. The molecular mechanisms underlying this risk are unclear at present. PMID- 11118025 TI - A common promoter variant of the leptin gene is associated with changes in the relationship between serum leptin and fat mass in obese girls. AB - Mutations in the leptin gene lead to rare obese syndromes of Mendelian inheritance in humans and rodents. However, no relevant mutations are found in the coding region of leptin gene DNA in patients with common multifactorial obesity. These obese patients tend to have an elevation of serum leptin proportional to their adiposity but with a rather wide dispersion of leptin levels for a given body fat content, which in part is attributable to sexual dimorphism. The current study, performed in two independent Caucasian cohorts of obese girls, shows that a frequent promoter variant of the leptin gene is associated with changes in the relationship between serum leptin and body fatness. Girls of comparable adiposity have different circulating leptin levels, depending on their genotype at this locus. Girls with the -/- Lep -2,549 genotype have 25% lower mean leptin levels than the girls with other genotypes, as reflected by differences in the regression slopes of leptin-to-fat mass. Therefore, genetic factors related to the leptin gene may be important in defining the set point of obese individuals (i.e., the circulating leptin level for a given degree of body fatness). This definition may be of both physiological and therapeutic relevance, although a phenotypic association with an individual single-nucleotide polymorphism is not sufficient to assign function to this particular nucleotide site. PMID- 11118026 TI - Parental transmission of type 2 diabetes: the Framingham Offspring Study. AB - Study of parental transmission of diabetes provides insight into the relative contributions of underlying maternal and paternal influences. We estimated risk for type 2 diabetes and milder degrees of glucose intolerance associated with parental diabetes among subjects of the population-based Framingham Offspring Study, in which participants are primarily Caucasian and at relatively low risk for diabetes and for which both parental and offspring phenotypes were ascertained by direct examination. Parental diabetes, assessed over 40 years of biennial follow-up, was defined by use of hypoglycemic drug therapy or a casual plasma glucose level > or = 11.1 mmol/l at any examination. Offspring glucose tolerance, assessed over 20 years of quadrennial follow-up, was defined by fasting plasma glucose levels > or = 7.8 mmol/l at any two examinations, use of hypoglycemic drug therapy at any examination, or with a 75-g oral glucose tolerance test (1980 World Health Organization criteria) at the most recent examination. We calculated odds ratios (ORs) and 95% CIs for offspring glucose tolerance status using generalized estimating equations to account for differential correlations within and between families. The 2,527 offspring came from 1,303 nuclear families, of which 77.6% had two or more siblings per family and in which the prevalence of parental diabetes was 24.6%. The mean offspring age was 54 years (range 26-82), 53% were women, 8.6% had diabetes, 11.4% had impaired glucose tolerance, 76.3% had no parental diabetes, 10.5% had maternal diabetes, 11.5% had paternal diabetes, and 1.7% had bilineal diabetes. Relative to individuals without parental diabetes, the age-adjusted ORs (95% CI) for offspring type 2 diabetes or abnormal glucose tolerance (fasting plasma glucose > or = 6.1 mmol/l or 2-h postchallenge glucose tolerance > or = 7.8 mmol/l) among individuals with maternal diabetes were 3.4 (2.3-4.9) and 2.7 (2.0-3.7), respectively; among individuals with paternal diabetes were 3.5 (2.3-5.2) and 1.7 (1.2-2.4), respectively; and among individuals with bilineal diabetes were 6.1 (2.9-13.0) and 5.2 (2.6-10.5), respectively. Although maternal and paternal diabetes conferred equivalent risk for offspring type 2 diabetes, offspring with maternal diabetes were slightly more likely to have abnormal glucose tolerance compared with those with paternal diabetes (OR 1.6, 95% CI 1.1-2.4). Offspring with maternal diabetes and an age of onset of <50 years had marked increased risk for both type 2 diabetes (9.7, 4.3-22.0) and abnormal glucose tolerance (9.0, 4.2 19.7). We conclude that risk ratios for offspring type 2 diabetes are consistent with a simple additive risk model, where risk when both parents are affected equals the sum of risk when either parent is affected. For maternal diabetes to confer excess risk for mild but not overt glucose intolerance, offspring of diabetic fathers may transit abnormal to impaired glucose tolerance relatively quickly, or diabetic mothers may transmit risk for a mild slowly progressive form of abnormal glucose tolerance in addition to overt diabetes. Very high risk for abnormal glucose homeostasis among offspring with young age-of-onset maternal diabetes is consistent with hypotheses that perinatal exposures increase diabetes risk. Given equivalent risk ratios for type 2 diabetes, fathers may transmit unique paternal genetic factors of similar strength to maternal environmental factors. PMID- 11118027 TI - Intrauterine exposure to diabetes conveys risks for type 2 diabetes and obesity: a study of discordant sibships. AB - Intrauterine exposure to diabetes is associated with an excess of diabetes and obesity in the offspring, but the effects of intrauterine exposure are confounded by genetic factors. To determine the role of the intrauterine diabetic environment per se, the prevalence of diabetes and the mean BMI were compared in siblings born before and after their mother was recognized as having diabetes. Nuclear families in which at least one sibling was born before and one after the mother was diagnosed with type 2 diabetes were selected. Consequently, the siblings born before and after differed in their exposure to diabetes in utero. A total of 58 siblings from 19 families in which at least one sibling had diabetes were examined at similar ages (within 3 years). The risk of diabetes was significantly higher in siblings born after the mother developed diabetes than in those born before the mother's diagnosis of diabetes (odds ratio 3.7, P = 0.02). In 52 families, among 183 siblings without diabetes, the mean BMI was 2.6 kg/m2 higher in offspring of diabetic than in offspring of nondiabetic pregnancies (P = 0.003). In contrast, there were no significant differences in risk of diabetes or BMI between offspring born before and after the father was diagnosed with diabetes. Intrauterine exposure to diabetes per se conveys a high risk for the development of diabetes and obesity in offspring in excess of risk attributable to genetic factors alone. PMID- 11118028 TI - Further evidence for a susceptibility locus for type 2 diabetes on chromosome 20q13.1-q13.2. AB - We previously reported suggestive linkage between type 2 diabetes and markers in a region on chromosome 20q using data from a collection of 29 Caucasian families in which type 2 diabetes with middle-age-onset was segregated as an autosomal dominant disorder. To map more precisely the susceptibility locus (or loci) within this broad region, we increased the family collection and genotyped all families for additional markers, both within the critical region and spaced over the rest of chromosome 20. Altogether 526 individuals (including 241 with diabetes) from the total collection of 43 families were included in the study. All individuals were genotyped for 23 highly polymorphic markers. Positive evidence for linkage was found for a 10-cM region on the long arm of chromosome 20q13.1-q13.2 between markers D20S119 and D20S428. The strongest evidence in two point as well as multipoint linkage analysis (P = 1.8 x 10(-5)) occurred at the position corresponding to marker D20S196. The individuals with diabetes in the seven most strongly linked families had high serum insulin levels during fasting and 2-h post-glucose load periods. We did not find any evidence for linkage between type 2 diabetes and any other region on chromosome 20. In conclusion, our larger and more comprehensive study showed very strong evidence for a susceptibility gene for insulin-resistant type 2 diabetes located on the long arm of chromosome 20 around marker D20S196. PMID- 11118029 TI - Non-class II HLA gene associated with type 1 diabetes maps to the 240-kb region near HLA-B. AB - Several studies provide evidence that in addition to the DQ-DR genes, HLA contains another uncharacterized gene or genes associated with type 1 diabetes. Our aim was to investigate the effect of this gene independently of the DQ-DR genes and to localize it with a matched case-control study. More than 1,400 patients and 30,000 control individuals from Finland were studied. They were first genotyped for the selected alleles of the HLA-DQB1, -DQA1, and -DRB1 genes. For the DR3/4(0404) genotype, 75 patients and 181 control subjects were stratified, and 241 patients and 354 controls were stratified for the DR3/4(0401) genotype. Ten microsatellite markers in the HLA class III and I regions (D6S273, TNFa, C12A, STR MICA, MIB, C125, C143, C245, C3211, and MOGc) and selected alleles of the HLA-A and HLA-B genes were studied. In the DR3/4(0404)-stratified group, we found that markers located between C12A and C143 near the HLA-B gene confer a strong additional diabetes association. This was confirmed by the population differentiation test in both DR3/4(0404)- and DR3/4(0401)-stratified groups. Our data indicate that an additional gene associated with type 1 diabetes is located in the 240-kb region near HLA-B. We excluded STR MICA polymorphism as a mutation responsible for diabetes association. PMID- 11118030 TI - Cd36 and molecular mechanisms of insulin resistance in the stroke-prone spontaneously hypertensive rat. AB - Insulin resistance is of pathogenic importance in several common human disorders including type 2 diabetes, hypertension, obesity and hyperlipidemia, but the underlying mechanisms are unknown. The spontaneously hypertensive rat (SHR) is a model of these human insulin resistance syndromes. Quantitative trait loci (QTLs) for SHR defects in glucose and fatty acid metabolism, hypertriglyceridemia, and hypertension map to a single region on rat chromosome 4. Genetic analysis of an SHR derived from a National Institutes of Health colony led to the identification of a causative mutation in the SHR Cd36. We have investigated glucose and fatty acid metabolism in the stroke-prone SHR (SHRSP). We demonstrate defects in insulin action on 2-deoxy-D-glucose transport (SHRSP 3.3 +/- 1.5 vs. 21.0 +/- 7.4 pmol x min(-1) x [20 microl packed cells](-1), SHRSP vs. WKY, respectively, P = 0.01) and inhibition of catecholamine-stimulated lipolysis (P < 0.05 at all concentrations of insulin) in adipocytes isolated from SHRSP. In contrast, basal levels of catecholamine-stimulated nonesterified free fatty acid (NEFA) release and plasma levels of NEFA are similar in SHRSP and WKY. These results are in agreement with the data on the SHR.4 congenic strain, which suggested that the QTL containing Cd36 mutations accounted for the entire defect in basal catecholamine action but only for approximately 40% of the SHR defect in insulin action. In the SHR, both abnormalities appear consequent of defective Cd36 expression. Because Cd36 sequence and expression are apparently normal in SHRSP, it is likely that the molecular mechanism for defective insulin action in this strain is caused by a gene(s) different than Cd36. PMID- 11118031 TI - Isolation of a peptide for targeted drug delivery into human head and neck solid tumors. AB - Lack of tumor specificity remains a major problem with chemotherapies in that side effects prevent the delivery of dosages of drugs that are required to eliminate tumors. In this report, we describe the isolation of a 12-mer peptide (HN-1), with approximately 1% of the mass of typical antibodies, that meets several criteria for targeted drug delivery into a solid tumor. First, internalization of HN-1 by human head and neck squamous cell cancer (HNSCC) cells suggests that HN-1 is capable of translocating drugs across cell membranes. Second, HN-1 appears to be HNSCC-specific, given its reduced uptake by nonmalignant human oral keratinocytes and other types of human cells, its preferential binding to primary HNSCC, and its localization to HNSCC-derived xenografts. Third, the presence of HN-1 within HNSCC xenografts suggests that it is capable of penetrating tumor tissues. Our results establish the utility of tumor-specific peptides for targeted drug delivery into solid tumors. PMID- 11118032 TI - Tumor metastasis and the reciprocal regulation of prometastatic and antimetastatic factors by nuclear factor kappaB. AB - To investigate the role of the transcription factor nuclear factor kappaB (NFkappaB) in tumor metastasis, we generated a murine lung alveolar carcinoma cell line (Line 1) defective in NFkappaB-signaling by retroviral delivery of a dominant negative inhibitor of NFkappaB. The NFkappaB signal blockade resulted in the down-regulation of prometastatic matrix metalloproteinase 9, a urokinase-like plasminogen activator, and heparanase and reciprocal up-regulation of antimetastatic tissue inhibitors of matrix metalloproteinases 1 and 2 and plasminogen activator inhibitor 2. NFkappaB signal blockade did not affect tumor cell proliferation in vitro or in vivo but prevented intravasation of tumor cells in an in vivo chick chorioallantoic membrane model of metastasis as well as spontaneous metastasis in a murine model. These findings suggest that NFkappaB plays a central and specific role in the regulation of tumor metastasis and may be an important therapeutic target for development of antimetastatic cancer treatments. PMID- 11118033 TI - WISH-PC2: a unique xenograft model of human prostatic small cell carcinoma. AB - Prostatic small cell carcinoma is an aggressive subtype of prostate cancer that usually appears as a progression of the original adenocarcinoma. We describe here the WISH-PC2, a novel neuroendocrine xenograft of small cell carcinoma of the prostate. This xenograft was established from a poorly differentiated prostate adenocarcinoma and is serially transplanted in immune-compromised mice where it grows within the prostate, liver, and bone, inducing osteolytic lesions with foci of osteoblastic activity. It secretes to the mouse Chromogranin A and expresses prostate plasma carcinoma tumor antigen-1, six-transmembrane epithelial antigen of the prostate, and members of the Erb-B receptor family. It does not express prostate-specific antigen, prostate stem cell antigen, prostate-specific membrane antigen, and androgen receptor, and it grows independently of androgen. Altogether, WISH-PC2 provides an unlimited source in which to study the involvement of neuroendocrine cells in the progression of prostatic adenocarcinoma and can serve as a novel model for the testing of new therapeutic strategies for prostatic small cell carcinoma. PMID- 11118034 TI - PSGR, a novel prostate-specific gene with homology to a G protein-coupled receptor, is overexpressed in prostate cancer. AB - PSGR, a new prostate tissue-specific gene with homology to the G protein-coupled odorant receptor gene family, has been identified. Here we report the characteristics of the predicted protein sequence of PSGR and its prostate tissue specificity and expression profile in human prostate cancer and matched normal tissues. Using multiple tissue Northern blots from over 50 different tissues, PSGR expression was restricted to human prostate tissues. Paired normal and tumor specimens from 52 primary prostate cancers, obtained by laser capture microdissection or manual microdissection, were analyzed for PSGR expression by semiquantitative and real-time PCR assays. The differential expression of PSGR between normal and tumor tissues was highly significant (P < 0.001), and 32 of 52 (62%) matched prostate specimens exhibited tumor-associated overexpression of PSGR. Of note, there was very little or no expression of PSGR in many normal specimens in comparison with the generally high expression of PSGR seen in matched tumor specimens. In situ hybridization assays showed restricted PSGR expression in the epithelial cells of the normal and tumor tissue sections. Restricted expression of PSGR in prostatic epithelial cells, overexpression of the PSGR in a significant percentage of prostate cancers, and the predicted protein sequence of PSGR with seven transmembrane domains provide a foundation for future studies evaluating the potential of PSGR as a prostate cancer gene expression marker and the utility of PSGR protein as a novel target for developing immunotherapeutic strategies for prostate cancer. PMID- 11118035 TI - In vitro sensitivity of T-cell lymphoblastic leukemia to UCN-01 (7 hydroxystaurosporine) is dependent on p16 protein status: a Pediatric Oncology Group study. AB - p16 regulates the cell cycle pathway by inhibiting the cyclin Ds-cyclin-dependent kinase (CDK) 4/6-mediated phosphorylation of retinoblastoma protein (pRb). Previously, we reported that most primary T-cell acute lymphoblastic leukemia (T ALL) harbored p16 inactivation and hyperphosphorylated pRb without cyclin Ds or CDK4/6 alterations. Therefore, inhibiting CDK4/6 may be an ideal therapeutic approach for p16 (-) T-ALL. UCN-01 (7-hydroxystaurosporine) is a potent antitumor agent that exerts its effects through the inhibition of CDKs. We now report that p16 protein expression status of T-ALL cells influences their sensitivity to UCN 01. In 36 primary T-ALL cells, the IC50s of UCN-01 in the 27 p16 (-) cells (43+/ 52 nM) was significantly lower than that in the 9 p16 (+) cells (258+/-260 nM). Our results suggest that agents like UCN-01 may be useful as a p16-selective therapy for T-ALL. PMID- 11118036 TI - Activity of a novel camptothecin analogue, homocamptothecin, in camptothecin resistant cell lines with topoisomerase I alterations. AB - Homocamptothecin (hCPT), which differs from camptothecin (CPT) by the presence of an additional methylene group in the E-ring, was evaluated in CPT-resistant cell lines. Topoisomerase I (top1)-deficient leukemia P388/CPT45 cells were highly resistant to hCPT, which demonstrates that top1 is the primary target of hCPT. Three CPT-resistant cell lines with top1 point mutations (Chinese hamster lung fibroblast DC3F/C10, human prostate carcinoma DU-145/RC1, and human leukemia CEM/C2) and their top1 enzymes were cross-resistant to hCPT. The antiproliferative activity of hCPT was greater than that of CPT in both parental and CPT-resistant cell lines, particularly in the prostate cell lines. The top1 cleavage complexes formed in the presence of hCPT appear to be more stable than those induced by CPT. Together, these data indicate that hCPT is a specific top1 inhibitor, which shares a common binding site with CPT in the topl-DNA cleavage complexes. Because of its potency, hCPT might overcome resistance to CPT in some cancer cells. PMID- 11118037 TI - DLC-1 is deleted in primary hepatocellular carcinoma and exerts inhibitory effects on the proliferation of hepatoma cell lines with deleted DLC-1. AB - We investigated the expression and deletion of DLC-1 (frequently deleted in liver cancer gene), first reported in 1998 and having a high homology with rat p122RhoGAP in hepatocellular carcinoma (HCC). Six (20%) of 30 human HCC samples and 2 (40%) of 5 HCC cell lines were found to have no detectable DLC-1 expression by reverse transcription-PCR. Homozygous DLC-1 deletion was detected by Southern blotting in two of six HCC samples and in both HCC cell lines with no DLC-1 expression. Transfection of DLC-1 into 5 HCC cell lines (two with DLC-1 deletion and three with intact DLC-1) showed significant growth inhibition in these two HCC cell lines with deleted DLC-1 with both 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide and colony formation assays but not in three other HCC cell lines with intact DLC-1. Our findings suggest that DLC-1 may play an important role in hepatocarcinogenesis. PMID- 11118038 TI - RB18A, whose gene is localized on chromosome 17q12-q21.1, regulates in vivo p53 transactivating activity. AB - Among the different cellular factors that regulated p53 functions, we previously identified (P. Drane et al., Oncogene, 15: 3013-3024, 1997) RB18A, a new gene whose encoded Mr 205,000 protein interacted in vitro, through its COOH-terminal domain, with p53. Therefore, we analyzed the in vivo role of RB18A by measuring its effect on the transactivating activity of p53 on physiological promoters. We herein demonstrated that RB18A, which interacted also in vivo with p53, activated Bax promoter and inhibited p21Waf1 or IGF-BP3 promoters. In addition, fluorescence in situ hybridization mapping led to localizing the RB18A gene on chromosome 17q12-q21.1, loci associated with human cancers. This is the first demonstration that in vivo RB18A, in a protein-protein interaction, regulates p53 transactivating activity. PMID- 11118039 TI - Neutral endopeptidase promotes phorbol ester-induced apoptosis in prostate cancer cells by inhibiting neuropeptide-induced protein kinase C delta degradation. AB - Phorbol esters induce apoptosis in androgen-sensitive LNCaP cells, which express neutral endopeptidase (NEP), but not in androgen-independent prostate cancer (PC) cells, which lack NEP expression. We investigated the role of NEP in PC cell susceptibility to 12-O-tetradecanoylphorbol-13-acetate (TPA). Western analysis showed that expression of NEP and protein kinase Cdelta (PKCdelta) correlated with PC cell sensitivity to TPA-induced growth arrest and apoptosis in LNCaP cells and in TSU-Prl cells expressing an inducible wild-type NEP protein. Inhibition of NEP enzyme activity using the specific NEP inhibitor CGS24592, or inhibition of PKCdelta using Rottlerin at concentrations that inhibit PKCdelta but not PKCalpha, significantly inhibited TPA-induced growth inhibition and cell death. Furthermore, pulse-chase experiments showed PKCdelta is stabilized in LNCaP cells and in TSU-Pr1 cells overexpressing wild-type NEP compared with PC cells lacking NEP expression. This results from NEP inactivation of its neuropeptide substrates (bombesin and endothelin-1), which in the absence of NEP stimulate cSrc kinase activity and induce rapid degradation of PKCdelta protein. These results indicate that expression of enzymatically active NEP by PC cells is necessary for TPA-induced apoptosis, and that NEP inhibits neuropeptide-induced, cSrc-mediated PKCdelta degradation. PMID- 11118040 TI - Direct evidence for the contribution of activated N-ras and K-ras oncogenes to increased intrinsic radiation resistance in human tumor cell lines. AB - Transformation with ras oncogenes results in increased radiation sur vival in many but not all cells. In addition, prenyltransferase inhibitors which inhibit ras proteins by blocking posttranslational modification radiosensitize cells with oncogenic ras. These findings suggest that oncogenic ras contributes to intrinsic radiation resistance. However, because introduction of ras oncogenes does not increase radiation survival in all cells and because prenyltransferase inhibitors target molecules other than ras, these studies left the conclusion that ras increases the intrinsic radi ation resistance of tumor cells in doubt. Here we show that genetic inactivation of K- or N-ras oncogenes in human tumor cells (DLD 1 and HT1080, respectively) leads to increased radiosensitivity. Reintroduction of the activated N-ras gene into the HT1080 line, having lost its mutant allele, resulted in increased radiation resistance. This study lends further support to the hypothesis that expression of activated ras can contribute to intrinsic radiation resistance in human tumor cells and extends this finding to the K- and N- members of the ras family. These findings support the development of strategies that target ras for inactivation in the treatment of cancer. PMID- 11118041 TI - Pharmacodynamics of tamoxifen and its 4-hydroxy and N-desmethyl metabolites: activation of caspases and induction of apoptosis in rat mammary tumors and in human breast cancer cell lines. AB - The antiestrogen tamoxifen (TAM) is extensively metabolized by cytochrome P-450 in humans and rodents. The active, estrogen receptor-binding metabolites, 4 hydroxy TAM (OHT) and N-desmethyl TAM (DMT) have been well characterized. We showed that the s.c. injection of 1 mg/kg TAM in adult female Sprague Dawley rats bearing carcinogen-induced mammary tumors resulted in rapid serum decline of parent TAM but higher exposure of the metabolites, OHT and DMT. We found for the first time that the administration of TAM for a short time resulted in a delayed induction of caspase activity and apoptosis within the mammary tumors. When TAM, OHT, or DMT was added to human breast cancer cell lines in culture, each elicited a time- and dose-dependent induction of caspase activity, preceding apoptosis. Importantly, pretreatment of the cells with a pharmacological inhibitor of caspases [benzyloxy Val-Ala-Asp-fluoromethyl ketone (z-VAD-fmk)] blocked apoptosis induced by all three of the compounds, implicating a critical role of caspases in TAM-, OHT-, or DMT-induced apoptosis. The results obtained from these studies suggest that one possible mechanism of inhibition of mammary carcinogenesis and tumor growth in vivo may be the induction of caspase-dependent apoptosis, and that the metabolites OHT and DMT may contribute to the antitumor effect of TAM. PMID- 11118042 TI - Sulindac sulfone inhibits K-ras-dependent cyclooxygenase-2 expression in human colon cancer cells. AB - Both the sulfide and sulfone metabolites of sulindac, a nonsteroidal anti inflammatory drug, display anticarcinogenic effects in experimental models. Sulindac sulfide inhibits cyclooxygenase (COX) enzyme activities and has been reported to suppress ras-dependent signaling. However, the mechanisms by which sulindac sulfone suppresses cancer growth are not as defined. We studied the effects of these sulindac metabolites in human colon cancer-derived Caco-2 cells that have been transfected with an activated K-ras oncogene. Stable transfected clones expressed high levels of COX-2 mRNA and protein, compared with parental cells. K-ras-transfected cells formed tumors more quickly when injected into severe combined immunodeficiency disease mice than parental cells, and this tumorigenesis was suppressed by treatment with sulindac. Sulindac sulfone inhibited COX-2 protein expression, which resulted in a decrease in prostaglandin synthase E2 production. Sulindac sulfide had little effect on COX-2 in this model, but did suppress prostaglandin synthase E2 production, presumably by inhibiting COX enzyme activity. These data indicate that the sulfide and sulfone derivatives of sulindac exert COX-dependent effects by distinct mechanisms. PMID- 11118043 TI - Mutations of adenomatous polyposis coli and beta-catenin genes during progression of lung tumors induced by N-nitrosobis(2-hydroxypropyl)amine in rats. AB - In the present study, we investigated mutations of the adenomatous polyposis coli (APC) and beta-catenin genes to clarify possible molecular mechanisms underlying development of lung tumors induced by N-nitrosobis(2-hydroxypropyl)amine (BHP) in rats. Male Wistar rats, 6 weeks of age, were given 2000 ppm BHP in drinking water for 12 weeks and then maintained without further treatment until sacrifice at week 25 DNA was extracted from paraffin-embedded tissues, and PCR-single-strand conformation polymorphism analysis, followed by nucleotide sequencing, was performed. No APC mutations were detected in 17 hyperplasias, but 2 of 15 adenomas (13.3%) and 8 of 20 adenocarcinomas (40.0%) showed changes within exon 1 to the mutation cluster region in exon 15. For beta-catenin, no mutations were detected in 17 hyperplasias, but 3 of 15 adenomas (20.0%) and 5 of 20 adenocarcinomas (25.0%) had alterations within or flanking codons corresponding to important phosphorylation sites. Immunohistochemical staining showed beta catenin protein localized in the cell membranes in the surrounding normal appearing lung and 216 hyperplasias and localized mainly in the cytoplasm and/or nucleus in 10 of 37 adenomas (27.0%) and 21 of 40 adenocarcinomas (52.5%). These results suggest that the APC-beta-catenin-T-cell factor signaling pathway is involved in the acquisition of growth advantage from adenomas to adenocarcinomas in BHP-induced rat lung carcinogenesis. PMID- 11118044 TI - Identification of differentially expressed genes in human gliomas by DNA microarray and tissue chip techniques. AB - New genomic large-scale screening techniques have made the task of establishing an accurate molecular fingerprint of cancer cells feasible. Here, we have used a two-phase strategy for identification of molecular alterations in gliomas. First, cDNA microarrays (Clontech Laboratories, Inc., Research Genetics) were used to pinpoint differentially expressed genes between normal brain and diffuse astrocytomas (grades II-IV), and between a primary tumor and a later tumor reoccurrence in the same patient. More than 200 gene expression alterations were detected from glioblastomas, whereas relatively few changes were seen in grade II and grade III tumors. The most distinct progression-related expression change was the up-regulation of the insulin-like growth factor binding protein 2 (IGFBP2) gene. Second, a high-density tissue microarray of 418 brain tumors was constructed and used for clinical validation of gene expression changes. Strong expression of IGFBP2 was associated with progression and poor patient survival in diffuse astrocytomas (P < 0.0001). Third, comparisons of the data between (a) multiple spots retrieved from one predefined tumor region (IGFBP2 and vimentin immunohistochemistry, 20 tumors) or between (b) standard slides and arrayed tissues (p53 immunohistochemistry, 42 tumors) revealed very little variation. In conclusion, the combined use of DNA microarrays and tissue microarrays offers a powerful strategy for rapid identification and thorough characterization of differentially expressed genes in gliomas. PMID- 11118045 TI - Allelic deletions of cell growth regulators during progression of bladder cancer. AB - Cell growth regulators include proteins of the p53 pathway encoded by the genes CDKN2A (p16, p14arf), MDM2, TP53, and CDKN1A (p21) as well as proteins encoded by genes like RB1, E2F, and MYCL. In the present study we investigated allelic deletions of all these genes in each recurrent bladder tumor from well-defined clinical material with more than 3 years of follow-up. We followed three groups (22 or 23 patients/group) of patients with: (a) recurrent noninvasive tumors (Ta); (b) primary muscle-invasive tumors (T2-T4); and (c) progressing tumors (Ta/T1 --> T2/T4). We found a significant difference in the numbers of gene loci hit by deletions muscle-invasive versus noninvasive tumors (P = 0.0000002), with the genes most often hit by deletions in muscle-invasive tumors being TP53, RB1, and MYCL. A number of novel findings were made. Losses of MYCL and RB1 alleles were more pronounced in patients having concomitant field disease because 11 of 14 informative cases showed losses compared with 3 of 8 cases without field disease. A more pronounced deletion of TP53 (P = 0.002) and RB1 (P = 0.02) was found in the progressing tumor group compared with the recurrent noninvasive group, and, finally, the combined loss of TP53 and RB1 was present only in the progressing tumor or muscle-invasive groups. Deletion of two or more loci in TP53, MYCL, RB1, and CDKN2A was found in 10 patients in the progressing tumor group and in only 1 patient in the recurrent noninvasive group (P = 0.004). The data demonstrate that a characteristic difference between recurrent noninvasive and recurrent progressing bladder tumors is loss of cell cycle-regulatory genes in the latter group. PMID- 11118046 TI - Antiandrogenic effects of novel androgen synthesis inhibitors on hormone dependent prostate cancer. AB - We have found that in addition to being potent inhibitors of 17alpha hydroxylase/C17,20-lyase and/or 5alpha-reductase, some of our novel androgen synthesis inhibitors also interact with the mutated androgen receptor (AR) expressed in LNCaP prostate cancer cells and the wild-type AR expressed in hormone-dependent prostatic carcinomas. The effects of these compounds on the proliferation of hormone-dependent human prostatic cancer cells were determined in vitro and in vivo. L-2 and L-10 are delta4-3-one-pregnane derivatives. L-35 and L-37 are delta5-3beta-ol-androstane derivatives, and L-36 and L-39 are delta4 3-one-androstane-derived compounds. L-2, L-10, and L-36 (L-36 at low concentrations) stimulated the growth of LNCaP cells, indicating that they were interacting agonistically with the mutated AR expressed in LNCaP cells. L-35, L 37, and L-39 acted as LNCaP AR antagonists. To determine whether the growth modulatory effects of our novel compounds were specific for the mutated LNCaP AR, competitive binding studies were performed with LNCaP cells and PC-3 cells stably transfected with the wild-type AR (designated PC-3AR). Regardless of AR receptor type, all of our novel compounds were effective at preventing binding of the synthetic androgen methyl-trienolone[17alpha-methyl-(3H)-R1881 to both the LNCaP AR and the wildtype AR. L-36, L-37, and L-39 (5.0 microM) prevented binding by >90%, whereas L-35 inhibited binding by 30%. To determine whether the compounds were acting as agonists or antagonists, LNCaP cells and PC-3AR cells were transfected with the pMAMneoLUC reporter gene. When luciferase activity was induced by dihydrotestosterone, all of the compounds were found to be potent inhibitors of transcriptional activity, and the pattern of inhibition was similar for both receptor types. However, L-2, L-10, and L-36 were determined to be AR agonists, and L-35, L-37, and L-39 were wild-type AR antagonists. When tested in vivo, L-39 was the only AR antagonist that proved to be effective at inhibiting the growth of LNCaP prostate tumor growth. L-39 slowed tumor growth rate in LNCaP tumors grown in male SCID mice to the same level as orchidectomy, significantly reduced tumor weights (P < 0.05), significantly lowered serum levels of prostate specific antigen (P < 0.02), and significanty lowered serum levels of testosterone (P < 0.05). L-39 also proved to be effective when tested against the PC-82 prostate cancer xenograft that expresses wild-type AR. These results show that some of our compounds initially developed to be inhibitors of androgen synthesis also interact with the human AR and modulate the proliferation of hormone-dependent prostatic cancer cells. Therefore, compounds such as L-39, which have multifunctional activities, hold promise for the treatment of androgen dependent prostate tumors. PMID- 11118047 TI - Locoregional cancer treatment with magnetic drug targeting. AB - The specific delivery of chemotherapeutic agents to their desired targets with a minimum of systemic side effects is an important, ongoing challenge of chemotherapy. One approach, developed in the past to address this problem, is the i.v. injection of magnetic particles [ferrofluids (FFs)] bound to anticancer agents that are then concentrated in the desired area (e.g., the tumor) by an external magnetic field. In the present study, we treated squamous cell carcinoma in rabbits with FFs bound to mitoxantrone (FF-MTX) that was concentrated with a magnetic field. Experimental VX-2 squamous cell carcinoma was implanted in the median portion of the hind limb of New Zealand White rabbits (n = 26). When the tumor had reached a volume of approximately 3500 mm3, FF-MTX was injected intraarterially (i.a.; femoral artery) or i.v. (ear vein), whereas an external magnetic field was focused on the tumor. FF-MTX i.a. application with the external magnetic field resulted in a significant (P < 0.05), complete, and permanent remission of the squamous cell carcinoma compared with the control group (no treatment) and the i.v. FF-MTX group, with no signs of toxicity. The intratumoral accumulation of FFs was visualized both histologically and by magnetic resonance imaging. Thus, our data show that i.a. application of FF-MTX is successful in treating experimental squamous cell carcinoma. This "magnetic drug targeting" offers a unique opportunity to treat malignant tumors locoregionally without systemic toxicity. Furthermore, it may be possible to use these magnetic particles as a "carrier system" for a variety of anticancer agents, e.g., radionuclides, cancer-specific antibodies, and genes. PMID- 11118048 TI - Yeast cytosine deaminase improves radiosensitization and bystander effect by 5 fluorocytosine of human colorectal cancer xenografts. AB - The efficacy of cancer gene therapy using bacterial cytosine deaminase (bCD)/5 fluorocytosine (5-FC) enzyme/prodrug strategy is limited by the inefficiency of cytosine deaminase (CD)-catalyzed conversion of 5-FC into 5-fluorouracil (5-FU). We have shown previously that yeast CD (yCD) is more efficient at the conversion of 5-FC than bCD. In the current study, we hypothesized that the increased production of 5-FU by yCD would enhance the efficacy of the CD/5-FC treatment strategy by increasing the bystander effect as well as the efficacy of radiotherapy because of the radiosensitizing capacity of 5-FU. To test this hypothesis, we generated stable HT29 human colon cancer cell lines expressing either bCD (HT29/bCD) or yCD (HT29/yCD). The amount of 5-FU produced in HT29/yCD tumors after a single injection of 5-FC (1000 mg/kg, i.p.) was 15-fold higher than that produced in HT29/bCD tumors. In tumor-bearing nude mice, the average minimum relative tumor size (compared with pretreatment values) of HT29/bCD tumors treated with 5-FC and radiation (500 mg/kg i.p. and 3 Gy, 5 days a week for 2 weeks) was 0.55+/-0.1, compared with 0.01+/-0.01 in HT29/yCD tumors (P = 0.002). Moreover, an increased cytotoxic and radiosensitizing effect of 5-FC on bystander cells was observed in vitro and in vivo when yCD was expressed in HT29 cells instead of bCD. In mice bearing HT29 tumors containing 10% HT29/yCD cells, the combined treatment resulted in a minimum tumor size of 0.20+/-0.07 compared with 0.60+/-0.1 in 10% HT29/bCD cells (P < 0.001). These results demonstrate that the use of yCD in the CD/5-FC strategy has a high potential to improve the therapeutic outcome of combined gene therapy and radiotherapy in cancer patients. PMID- 11118049 TI - Tyrosinase mutants are capable of prodrug activation in transfected nonmelanotic cells. AB - Tyrosinase has been suggested as a prodrug-converting enzyme for the treatment of melanoma. We hypothesized that tyrosinase expression in transfected nonmelanotic cells can be used in a gene therapy paradigm of prodrug activation. To verify our hypothesis, we used the following tyrosinase variants: (a) a full-length human tyrosinase clone (T); (b) a mutant lacking the COOH-terminal cytoplasmic domain (TdeltaC); (c) a mutant lacking the COOH-terminal transmembrane and cytoplasmic domains (TdeltaTC); and (d) a fusion with the eight COOH-terminal amino acids of lysosome-associated membrane protein-1 (TL). Expression of mutant and wild-type tyrosinases was induced by transfection in nontumorigenic human cells of epithelial origin (293HEK, MCF-10A adenoma, and NHDF-Ad human dermal fibroblasts) as well as in tumor cells (9L gliosarcoma, MCF7 adenocarcinoma, and HT-1080 fibrosarcoma). When compared with the wild-type tyrosinase transfectants, truncated mutant expression resulted in higher mRNA levels that paralleled higher enzyme activity of the truncated mutants. Two model tyrosinase prodrugs, hydroxyphenyl-propanol (HPP) and N-acetyl-4-S-cysteaminylphenol (NAcSCAP) inhibited proliferation and caused cell death of transfected cells in a dose dependent manner. Effects of prodrug treatment were compared for tumorigenic cells and their nontumorigenic counterparts. Two truncated mutants (TdeltaC and TdeltaTC) showed low endogenous cytotoxicity and efficiently suppressed proliferation and induced cytotoxicity in transfected tumor cells in the presence of NAcSCAP. Overall, these results indicate that the developed tyrosinase mutants hold promise as prodrug activation systems for tumoral gene therapy. PMID- 11118050 TI - A tetravalent single-chain antibody-streptavidin fusion protein for pretargeted lymphoma therapy. AB - Single-chain Fv antibody fragments from the CD20-specific murine monoclonal antibody B9E9 were genetically engineered as streptavidin fusions [single-chain Fv-streptavidin (scFvSA) fusion protein] for use in pretargeted radioimmunotherapy. The scFvSA constructs were expressed as soluble, tetrameric species in the periplasm of Escherichia coli. Expression levels were affected by the order of the variable regions and the length and composition of the single chain Fv linker. The best expressor was obtained with the variable regions in the heavy chain-light chain configuration separated by a 25-mer Gly4Ser linker. This construct produced 250-300 mg of soluble, tetrameric fusion protein per liter of fermentor culture. The fusion protein (Mr 173,600) was purified from crude lysates by iminobiotin affinity chromatography with an overall yield of about 50% and was analyzed for functionality both in vitro and in vivo. Immunoreactivity of the scFvSA fusion protein and its nanomolar affinity to CD20-positive Ramos cells were comparable with the B9E9 monoclonal antibody. The fusion protein had a biotin dissociation rate identical to recombinant streptavidin and bound an average of 3.6 biotins/molecule of a possible 4 biotins/molecule. Labeled fusion protein cleared from the blood of BALB/c mice with a P half-life of about 16 h. In nude mice bearing Ramos xenografts, the fusion protein demonstrated sufficient tumor localization of functional streptavidin to enable efficient, tumor-specific targeting of a subsequently administered radionuclide-chelate/biotin molecule. These results suggest that large quantities of functional scFvSA can be produced for clinical testing as a therapy for non-Hodgkin's lymphoma. PMID- 11118051 TI - Suppression of ganglioside GD3 expression in a rat F-11 tumor cell line reduces tumor growth, angiogenesis, and vascular endothelial growth factor production. AB - Ganglioside GD3 is overexpressed in many types of tumors and may be associated with tumor progression and the development of metastatic potential. In our previous study (G. Zeng et al., Biochemistry, 38: 8762-8769, 1999), we established a subclone of the rat dorsal root ganglion-derived F-11 cells in which the expression of ganglioside GD3 was inhibited by stable transfection of the antisense vector against CMP-NeuAc: GM3 alpha2-8 sialyltransferase (GD3 synthase) gene. This cell line exhibits markedly reduced rate of tumor growth in vivo. Here, we further characterized the antisense-transfected cell line, and the results showed that these cells formed small, minimally vascularized tumors exhibiting extensive necrosis. In vivo Matrigel assay revealed reduced vascularization and low hemoglobin content in the antisense xenografts. Significantly fewer new vessels were found on the antisense xenografts and the skin around them than those on/around the xenografts formed by the sense transfected and untransfected F-11 cells. The hemoglobin content of the antisense xenografts was much lower than that of the xenografts formed by the control cells. The reduced angiogenesis in the antisense xenografts was correlated with a decrease in vascular endothelial growth factor (VEGF) production. The expression of VEGF was suppressed in the antisense xenografts and the conditioned culture media of the antisense-transfected F-11 cells as determined by Western blotting analysis. This was further confirmed by immunohistochemistry of the tumors using antibodies against VEGF and platelet/endothelial cell adhesion molecule (PECAM 1). Therefore, our results demonstrate that reduced tumor growth in nude mice by suppression of GD3-synthase expression in F-11 cells results from minimal angiogenesis of the tumors through down-regulation of the VEGF expression, which indicates an important role for GD3 in tumor angiogenesis. PMID- 11118052 TI - Identification of a 6-cM minimal deletion at 11q23.1-23.2 and exclusion of PPP2R1B gene as a deletion target in cervical cancer. AB - Previous functional and deletion mapping studies on cervical cancer (CC) have implicated one or more tumor suppressor genes (TSGs) on chromosome 11 at q13 and q22-24 regions. Of these, the 11q22-24 region exhibits frequent allelic deletions in a variety of solid tumor types, suggesting the presence of critical genes for tumor suppression in this region. However, the precise region of deletion on 11q is not clearly defined in CC. In an attempt to accurately map the deleted region, we performed an extensive loss of heterozygosity (LOH) mapping in 58 tumors using 25 polymorphic loci on both the short and long arms. The pattern of LOH identified three sites of deletions, two on 11p (p15.11-p15.3 and p12-13), and one on 11q (q23.1-q23.2). The 11q23.1-q23.2 exhibited highest frequency (60.6%) of deletions, suggesting that this could be the site of a candidate TSG in CC. The minimal deletion at 11q23.1-23.2 was restricted to a 6-cM region between 123.5 and 129.5 cM genetic distance on chromosome 11, identifying the site of a potential TSG important in the pathogenesis of CC. At least five known genes and 28 UniGene clusters were mapped to the present commonly deleted region. In addition, we have excluded a previously known TSG PPP2R1B at 11q23 as a deletion target in CC. The definition of the minimal deletion and the availability of expressed sequence resources should facilitate the identification of the candidate TSG. PMID- 11118053 TI - Down-regulation of monocyte chemotactic protein-3 by activated beta-catenin. AB - Accumulation of intracellular beta-catenin, as a result of inactivation of the adenomatous polyposis coli (APC) gene or by mutation of the beta-catenin gene (CTNNB1) itself, is involved in a wide range of human cancers. By means of fluorescent differential display using a murine fibroblast cell line (L-MT), which expresses an activated form of beta-catenin that accumulates in the cells, we found that expression of murine monocyte chemotactic protein-3 (mMCP-3) was suppressed by activated beta-catenin. Inversely, expression of MCP-3 in human colon cancer cells was induced by depletion of beta-catenin after adenovirus mediated transfer of wild-type APC genes into the cells. A reporter-gene assay indicated that the accumulation of beta-catenin in the nucleus suppressed activity of the MCP-3 promoter through a putative T-cell factor/lymphocyte enhancer factor (Tcf/LEF)-binding site, ATCAAAG; but when the promoter sequence contained a two-base substitution in the binding site, it failed to suppress reporter-gene (luciferase) activity. An electrophoretic mobility-shift assay using the putative Tcf/LEF-binding sequence revealed interaction of the candidate sequence with the beta-catenin complex. Furthermore, induction of MCP-3 cDNA into HT-29 colon cancer cells increased expression of two markers of differentiation: alkaline phosphatase and carcinoembryonic antigen. Our results implied that activation of beta-catenin through the Tcf/LEF signaling pathway may participate in colonic carcinogenesis by inhibiting MCP-3-induced differentiation of colorectal epithelial cells. PMID- 11118054 TI - Thioredoxin nuclear translocation and interaction with redox factor-1 activates the activator protein-1 transcription factor in response to ionizing radiation. AB - Thioredoxin (TRX) is a cytoplasmic, redox-sensitive signaling factor believed to participate in the regulation of nuclear transcription factors mediating cellular responses to environmental stress. Activation of the activator protein (AP)-1 transcription factor is thought to be mediated in part by redox-sensitive interactions between the nuclear signaling protein redox factor-1 (Ref-1) and TRX. In this study, the role of TRX and Ref-1 in the activation of the AP-1 complex was examined in HeLa and Jurkat cell lines exposed to ionizing radiation (IR). After exposure to IR, nuclear levels of immunoreactive TRX increased, accompanied by an increase in AP-1 DNA binding activity. It was shown that a physical interaction between Ref-1 and TRX occurs within the nucleus and is enhanced after exposure to IR. Furthermore, TRX immunoprecipitated from irradiated cells was capable of activating AP-1 DNA binding activity in nonirradiated nuclear extracts. In addition, immunodepletion of Ref-1 from nuclear extracts demonstrated that the increase in AP-1 DNA binding activity after IR was also dependent upon the presence of Ref-1 from irradiated cells. Finally, the ability of both TRX and Ref-1 from irradiated cells to stimulate AP 1 DNA binding in nonirradiated nuclear extracts was abolished by chemical oxidation and restored by chemical reduction. These results indicate that, in response to IR, TRX and Ref-1 undergo changes in redox state that contribute to the activation of AP-1 DNA binding activity. These experiments suggest that a redox-sensitive signaling pathway leading from TRX to Ref-1 to the AP-1 complex participates in the up-regulation of DNA binding activity in response to ionizing radiation. PMID- 11118055 TI - Mice vaccination with interleukin 12-transduced colon cancer cells potentiates rejection of syngeneic non-organ-related tumor cells. AB - Cell-based gene therapy after cytokine gene transfer is being investigated for autologous and allogeneic vaccination in cancer therapy. Here we show that mice vaccinated with 3-5 x 10(6) interleukin 12 (IL-12) gene-transduced CT26 colon cancer cells developed a long-lasting antitumor immune memory able to reject not only parental cells but also syngeneic, LM3 mammary, and MCE fibrosarcoma tumorigenic cells. In contrast, mice vaccinated with 0.5-1 x 10(6) CT26 cells transduced with pBabe neo IL-12 retrovirus cells (CT26-IL12) were only able to reject parental cells. An increase in the total circulating levels of IgG2a and a clear shift toward a systemic Th1 response developed, regardless of the amount of injected CT26-IL12 cells. On the contrary, a strong increase in anti-CT26 specific IgG2a levels was observed only when 3-5 x 10(6) CT26-IL12 cells were injected. Immunocompetent mice vaccinated with 3-5 x 10(6) CT26-IL12 cells developed local nodules for a few days, which then ceased growing. These nodules comprised mainly blood vessels, suggesting that an angiogenic process was taking place. CD8+ T cells were responsible for the anti-LM3 tumor cell memory, whereas CD4+ T cells were not involved. Splenocytes and lymphocytes obtained from mice immunized against CT26 cells were able to kill LM3 cells in vitro. Adoptive transfer of lymphocytes obtained from animals immunized against CT26 colon cancer cells suppressed LM3 mammary tumor growth in tumor-bearing mice. The present studies raised the possibility of isolating CTL clones and identifying CTL epitopes shared by different tumor cell types, which can be a target for cancer therapy. PMID- 11118056 TI - Dissociation of staurosporine-induced apoptosis from G2-M arrest in SW620 human colonic carcinoma cells: initiation of the apoptotic cascade is associated with elevation of the mitochondrial membrane potential (deltapsim). AB - We have identified an alternative apoptotic cascade induced in SW620 human colonic carcinoma cells by the protein kinase antagonist staurosporine (stsp). Consistent with its effect in other colonic epithelial cells, stsp induced G2-M arrest and apoptosis of SW620 cells. However, despite the paradigm that growth arrest triggers apoptotic cascades, apoptosis was detected before G2-M arrest. Reports have linked dissipation of the mitochondrial membrane potential (deltapsim) to the initiation of apoptosis and have linked elevation of the deltapsim to the escape from apoptosis However, neither apoptosis nor cell cycle arrest were altered by the collapse of the deltapsim, and increased deltapsim enhanced the initiation of apoptosis but blocked G2-M arrest. Although reactive oxygen species (ROS) have been implicated in some colonic epithelial cell and stsp-induced cascades, neither antioxidants nor the inhibition of RNA or protein synthesis altered apoptosis of SW620 cells. Finally, cytosolic cytochrome c has been linked to activation of caspase-3 and dissipation of the deltapsim. However, caspase-3 activation preceded the accumulation of cytochrome c in the cytosol and was accompanied by transient elevations in both the deltapsim and mitochondria associated cytochrome c. Therefore, we have identified a distinct apoptotic cascade in SW620 cells that was induced independently of growth arrest, dissipation of the deltapsim, ROS production, or synthesis of de novo RNA or protein, and we have linked its efficient initiation to early elevation of the deltapsim. PMID- 11118057 TI - The CD24/P-selectin binding pathway initiates lung arrest of human A125 adenocarcinoma cells. AB - Carbohydrates on tumor cells have been shown to play an important role in tumor metastasis. We demonstrated before that CD24, a Mr 35,000-60,000 mucine-type glycosylphosphatidylinositol-linked cell surface molecule, can function as ligand for P-selectin and that the sialylLex carbohydrate is essential for CD24-mediated rolling of tumor cells on P-selectin. To investigate the role of both antigens more closely, we transfected human A125 adenocarcinoma cells with CD24 and/or fucosyltransferase VII (Fuc TVII) cDNAs. Stable transfectants expressed CD24 and/or sialylLex. Biochemical analysis confirmed that in A125-CD24/FucTVII double transfectants, CD24 was modified with sialylLex. Only double transfectants showed rolling on P-selectin in vivo. When injected into mice, double transfectants arrested in the lungs, and this step was P-selectin dependent because it was strongly enhanced in lipopolysaccharide (LPS) pretreated wild-type mice but not in P-selectin knockout mice. CD24 modified by sialylLex was required on the tumor cells because the LPS-induced lung arrest was abolished by removal of CD24 from the cell surface by phosphatidylinositol-specific phospholipase C. A125-FucTVII single transfectants expressing sialylLex but not CD24 did not show P-selectin mediated lung arrest. The sialylLex epitope is abundantly expressed on human carcinomas, and significant correlations between sialylLex expression and clinical prognosis exist. Our data suggest an important role for sialylLex modified CD24 in the lung colonization of human tumors. PMID- 11118058 TI - Antiangiogenic effects of latent antithrombin through perturbed cell-matrix interactions and apoptosis of endothelial cells. AB - Antithrombin is a plasma protein of the serpin superfamily that may occur as several conformational variants. The native form of antithrombin is a major regulator of blood clotting. In the present study, we have identified the mechanism underlying the antiangiogenic action of a heat-denatured form, denoted latent antithrombin. Fibroblast growth factor (FGF)-induced angiogenesis in the chick embryo and angiogenesis in mouse fibrosarcoma tumors were inhibited by treatment with latent antithrombin at 1 mg/kg/day. Thermolysin-cleaved and native antithrombin were less efficient in these respects. Treatment with latent antithrombin induced apoptosis of cultured endothelial cells and inhibited cell migration toward FGF-2. Under these conditions, FGF-2-stimulated FGF receptor kinase activity was unaffected. However, actin reorganization, activation of focal adhesion kinase, and focal adhesion formation were disturbed by latent antithrombin treatment of FGF-2-stimulated endothelial cells. These data indicate that latent antithrombin induces apoptosis of endothelial cells by disrupting cell-matrix interactions through uncoupling of focal adhesion kinase. PMID- 11118059 TI - The effect of fibroblast growth factor 8, isoform b, on the biology of prostate carcinoma cells and their interaction with stromal cells. AB - Fibroblast growth factor 8, isoform b (FGF8b), has been implicated in the oncogenesis of the prostate and mammary epithelia. We examined whether overexpression of FGF8b in a weakly tumorigenic prostate carcinoma cell line, LNCaP, could alter the growth and tumorigenic properties of these cells. LNCaP cells were infected with a lentivirus vector carrying FGF8b cDNA and the green fluorescent protein (GFP) cDNA in the same construct, and the infected cell population was sorted on the basis of GFP protein expression. It was demonstrated that, in comparison with the cells transduced with GFP-vector alone, LNCaP cells with FGF8b-GFP expression manifested an increased growth rate, higher soft agar clonogenic efficiency, enhanced in vitro invasion, and increased in vivo tumorigenesis. Most strikingly, whereas parental or vector-control LNCaP cells failed to grow at all in an in vivo tumorigenesis/diaphragm invasion assay in nude mice, the cells overexpressing FGF8b proliferated as deposits of tumor cells on the diaphragm, frequently with indications of tumor cell invasion into the diaphragm. Coculturing of primary prostatic or non-prostatic stromal cells with the infected LNCaP cells led us to observe that: (a) stromal cells, irrespective of tissue origin, strongly suppressed LNCaP cell growth; (b) FGF8b producing LNCaP cells could partially evade the stromal inhibition, perhaps from the autocrine stimulatory effect of FGF8b; and (c) production of FGF8b in the coculture had a stimulatory effect on the proliferation of the stromal cells, prostatic or non-prostatic. This stimulation was not attributable to the direct action of FGF8b on stromal cells. Instead, it appears that epithelial-stromal cell-cell contact and some unknown soluble factors secreted by LNCaP cells upon stimulation of FGF8b are required for the maximal effect. Together, these results suggest that the growth rate and biological behavior of prostatic cancer cells can be altered to a more aggressive phenotype by up-regulation of FGF8b expression. These changes in phenotype also influence the interaction of the affected cells with stromal cells. The data obtained may have direct relevance to the progression of prostate cancer, recognizing that FGF8b is naturally overexpressed in advanced disease. PMID- 11118061 TI - Differential gene expression between normal and tumor-derived ovarian epithelial cells. AB - The majority of ovarian tumors arise from the transformation of the ovarian surface epithelial cells, a single layer of cells surrounding the ovary. To identify genes that may contribute to the malignant phenotype of ovarian cancers, cDNA representational difference analysis was used to compare expressed genes in primary cultures of normal human ovarian surface epithelium (HOSE) and ovarian tumor-derived epithelial cells from the Cedars-Sinai Ovarian Cancer (CSOC) repository. A total of 255 differentially expressed genes were identified, of which 160 and 95 were specifically expressed in HOSE and CSOC cells, respectively. Using cDNA array hybridization, the expression profiles of the genes identified by cDNA-representational difference analysis were examined in an additional 5 HOSE and 10 CSOC lines. The comparison of average signal of each gene revealed 44 HOSE-specific and 16 CSOC-specific genes that exhibited at least a 2.5-fold difference in expression. A large number of genes identified in this study encode membrane-associated or secreted proteins and, hence, may be useful as targets in the development of serum-based diagnostic markers for ovarian cancer. Very few genes associated with protein synthesis or metabolism were identified in this study, reflecting the lack of observable differences in phenotypic or growth characteristics between HOSE and CSOC cells. Northern blot analysis on a subset of these genes demonstrated comparable levels of gene expression as observed in the cDNA array hybridization. PMID- 11118060 TI - HGF/NK4, a four-kringle antagonist of hepatocyte growth factor, is an angiogenesis inhibitor that suppresses tumor growth and metastasis in mice. AB - We reported that NK4, composed of the N-terminal hairpin and subsequent four kringle domains of hepatocyte growth factor (HGF), acts as the competitive antagonist for HGF. We now provide the first evidence that NK4 inhibits tumor growth and metastasis as an angiogenesis inhibitor as well as an HGF antagonist. Administration of NK4 suppressed primary tumor growth and lung metastasis of Lewis lung carcinoma and Jyg-MC(A) mammary carcinoma s.c. implanted into mice, although neither HGF nor NK4 affected proliferation and survival of these tumor cells in vitro. NK4 treatment resulted in a remarkable decrease in microvessel density and an increase of apoptotic tumor cells in primary tumors, which suggests that the inhibition of primary tumor growth by NK4 may be achieved by suppression of tumor angiogenesis. In vivo, NK4 inhibited angiogenesis in chick chorioallantoic membranes and in rabbit corneal neovascularization induced by basic fibroblast growth factor (bFGF). In vitro, NK4 inhibited growth and migration of human microvascular endothelial cells induced by bFGF and vascular endothelial growth factor (VEGF) as well as by HGF. HGF and VEGF activated the Met/HGF receptor and the KDR/VEGF receptor, respectively, whereas NK4 inhibited HGF-induced Met tyrosine phosphorylation but not VEGF-induced KDR phosphorylation. NK4 inhibited HGF-induced ERK1/2 (p44/42 mitogen-activated protein kinase) activation, but allowed for bFGF- and VEGF-induced ERK1/2 activation. These results indicate that NK4 is an angiogenesis inhibitor as well as an HGF antagonist, and that the antiangiogenic action of NK4 is independent of its activity as HGF antagonist. The bifunctional properties of NK4 to act as an angiogenesis inhibitor and as an HGF antagonist raises the possibility that NK4 may prove therapeutic for cancer patients. PMID- 11118062 TI - K-ras codon 12 mutation induces higher level of resistance to apoptosis and predisposition to anchorage-independent growth than codon 13 mutation or proto oncogene overexpression. AB - The position of the point mutation in the c-K-ras gene appears associated with different degrees of aggressiveness in human colorectal tumors. In addition, colon tumors carrying K-ras codon 12 mutations associate with lower levels of apoptosis than tumors lacking this mutation. To test the hypothesis of a distinct transforming capacity of different K-ras forms in an in vitro system, we generated stable transfectants of NIH3T3 cells expressing a plasmid containing K ras mutated at codon 12 (K12) or at codon 13 (K13), or overexpressing the K-ras proto-oncogene (Kwt-oe). We evaluated changes in morphology, proliferative capacity, contact inhibition, and predisposition to apoptosis and anchorage independent growth in K12, K13, and Kwt-oe transformants. In addition, we studied alterations in expression and/or activation of proteins that participate in signal transduction downstream of Ras or are involved in the regulation of apoptosis and cell-cell (E-cadherin and beta-catenin) and cell-substrate (focal adhesion kinase) interactions. We observed that K13 or Kwt-oe transformants died synchronically 24-48 h after reaching confluency. Their death was apoptotic. In contrast, K12 grew, forming bigger colonies with higher cell densities; and before reaching confluency, spontaneously formed spheroids and showed no sign of apoptosis. The enhanced resistance to apoptosis, loss of contact inhibition, and predisposition to anchorage-independent growth in the K12 transformants were associated with higher AKT/protein kinase B activation, bcl-2, E-cadherin, beta catenin, and focal adhesion kinase overexpression, and RhoA underexpression, whereas the increased sensitivity of K13 or Kwt-oe transformants to apoptosis was associated with increased activation of the c-Jun-NH2-terminal kinase 1 pathway. All transformants showed a similar overactivation of mitogen-activated protein kinases and levels of bax expression similar to the endogenous level. Therefore, in our in vitro model, the localization of the mutation in the K-ras gene predisposes to a different level of aggressiveness in the transforming phenotype. K12 may increase aggressiveness not by altering proliferative pathways, but by the differential regulation of K-Ras downstream pathways that lead to inhibition of apoptosis, enhanced loss of contact inhibition, and increased predisposition to anchorage-independent growth. These results offer a molecular explanation for the increased aggressiveness of the tumors with K-ras codon 12 mutations observed in the clinical setting. PMID- 11118063 TI - Modulation of tumor angiogenesis by stem cell factor. AB - Mast cells accumulate within solid tumors and can release many angiogenic factors, suggesting that they may modulate vascularization of tumors. Stem cell factor (SCF) stimulates mast cell migration, proliferation, and degranulation and therefore may influence mast cell behavior within tumors. We investigated the contribution of SCF to tumor angiogenesis by manipulating its level in mammary tumors. Sense or antisense cDNA fragments of rat SCF were ligated into an episomal expression vector. Ethylnitrosourea-induced rat mammary tumor cell lines were transfected with vector containing either control (no insert, C-P), sense (S P), or antisense (AS-P) SCF DNA. The functional nature of the transfectants was confirmed by measuring SCF in cell lysates and conditioned media. Immunohistochemical analysis of the tumors induced in Berlin-Druckrey rats by these transfected cells demonstrated that mast cell number and microvascular density were significantly higher in S-P tumors and significantly lower in AS-P tumors, compared with C-P tumors. The expression of von Willebrand factor, an endothelial cell marker, showed a similar pattern. AS-P tumors were significantly smaller than either C-P or S-P tumors. These data suggest that SCF modulates tumor growth and angiogenesis via the involvement of mast cells. PMID- 11118064 TI - The phosphatidylinositol 3-kinase/AKT kinase pathway in multiple myeloma plasma cells: roles in cytokine-dependent survival and proliferative responses. AB - Interleukin 6 (IL-6) and insulin-like growth factor I (IGF-I) induce proliferative and antiapoptotic responses in multiple myeloma (MM) plasma cells. Because these cytokines may activate the phosphatidylinositol 3-kinase (PI 3 K)/AKT kinase pathway in other cell types, we investigated the role of PI 3-K/AKT in MM cell responses. IGF-I effectively activated PI 3-K in 8226 and OCI-My5 MM cells, but IL-6 was ineffective. However, IL-6 successfully activated PI 3-K in AF-10 MM cells and IL-6-dependent MH.60 plasmacytoma/hybridoma cells. IGF-I also successfully activated PI 3-K in four of four MM patient specimens, and IL-6 activated PI 3-K in three of four specimens. Inhibition of PI 3-K activity with wortmannin or Ly294002 blocked the antiapoptotic effect of IGF-I and the proliferative effect of IL-6 in the myeloma cell lines. Furthermore, a dominant negative PI 3-K construct, expressed in AF-10 cells by adenoviral infection, also significantly inhibited the IL-6 proliferative response in MM cells. In correlation with activation of PI 3-K, IGF-I also effectively activated the AKT kinase in 8226 and OCI-My5 cells, and IL-6 activated AKT in AF-10 and MH.60 cells. However, although incapable of activating PI 3-K in 8226 and OCI-My5 cells, IL-6 successfully activated AKT in these MM lines, suggesting PI 3-K independent mechanisms of AKT activation. The prevention of a myeloma cell proliferative response resulting from inhibition of PI 3-K activity was not associated with an inhibition of IL-6-dependent extracellular signal-regulated kinase (ERK) activation. These results support a role for the PI 3-K/AKT pathway in cytokine-dependent responses in myeloma cells, which is independent of any activation of the ERK pathway. PMID- 11118065 TI - Design of a peptide inhibitor that blocks the cell fusion mediated by glycoprotein 41 of human immunodeficiency virus type 1. AB - Fusion between the envelope of HIV and the plasma membrane of target cells is mediated by gp41, the envelope glycoprotein of HIV. Peptides derived from the membrane-proximal helical motif of the extracellular domain of gp41 effectively inhibit the infection of HIV, and their inhibitory activities are known to be correlated with the helical propensity of the peptides. We have designed small peptides that can form a stable alpha helix and thereby inhibit gp120/41-mediated cell fusion. A 19-mer peptide from the membrane-proximal helical motif of gp41 had no secondary structure in solution, and failed to block gp41-mediated cell fusion. When amino acids with low helical propensity were substituted, and helix capping sequences were introduced at both ends of the peptides, the modified peptides formed a stable helical structure. They also bound to the coiled-coil motif of gp41 presented at the C terminus of thioredoxin and blocked the cell fusion mediated by gp120/41. These results implied that such modification was enough to change a short peptide derived from gp41 into a potent inhibitor against the infection of HIV. PMID- 11118066 TI - Naive CD4+ T lymphocytes express high levels of Bcl-2 after highly active antiretroviral therapy for HIV infection. AB - The mechanism causing the increasing number of peripheral T cells after highly active antiretroviral therapy (HAART) is still unclear. The bcl-2 oncogene prevents spontaneous apoptosis (SA) in lymphocytes. Spontaneous apoptosis could be a determinant of HIV immunodeficiency and can be reversed by HAART including protease inhibitors (PI-HAART). The aims of our study were to measure Bcl-2 protein expression in memory (CD45RO+) and naive (CD45RO-) CD4+ and CD8+ T lymphocytes of HIV+ patients and to correlate it with efficacy of PI-HAART. Forty nine HIV+ patients (cases) and 26 HIV- individuals (controls) were evaluated. Patients receiving PI-HAART, and who had undetectable HIV plasma viral load (VL-, n = 21), had higher levels of Bcl-2 than did VL+ patients (n = 28), both in CD4+ cells (p < 0.0001) and in CD8+ cells (p < 0.001). VL+ patients had lower Bcl-2 levels than did controls in CD8+ cells (p = 0.02), but not in CD4+ cells (p > 0.05). Interestingly, VL- patients had higher Bcl-2 expression than did controls both in CD4+ cells (p < 0.0001) and in CD8+ cells (p = 0.03). In a subcohort of the same patients, Bcl-2 was significantly higher in VL- patients (n = 10) than in controls (n = 12), both in naive CD4+ cells (p < 0.0001) and in naive CD8+ cells (p = 0.01). Naive CD4+ cells had higher Bcl-2 expression in VL- than in VL+ patients (p = 0.01). In a subsequent longitudinal study of nine HIV patients, naive CD4+ cells increased after effective PI-HAART (p = 0.03), which paralleled an increase in Bcl-2 expression in the same cells (p = 0.02). In conclusion, upregulation of bcl-2 could be a mechanism of immune reconstitution of naive CD4+ T cells induced by PI-HAART. PMID- 11118067 TI - A randomized trial comparing the introduction of ritonavir or indinavir in 1251 nucleoside-experienced patients with advanced HIV infection. AB - ISS-IP1, a multicenter, randomized, 48-week open trial, was designed to compare the introduction of ritonavir or indinavir in patients with previous nucleoside experience and CD4+ cell counts below 50/mm3. Concomitant antiretroviral treatment with nucleoside analogs was allowed. Primary efficacy measures were survival and time to a new AIDS-defining event or death, analyzed through the whole period of observation by the intention-to-treat approach. Primary toxicity measures were time to treatment discontinuation and adverse events, grade at least 3/serious, analyzed by an on-treatment approach. Evaluation-of efficacy also included CD4+ cell and RNA response. The trial enrolled 1251 patients in 5 months. At baseline, mean CD4+ cell count was about 20 cells/mm3 and mean HIV RNA copy number was 4.9 log10/ml in both groups. Overall, 402 patients in the ritonavir group and 250 patients in the indinavir group permanently discontinued the assigned treatment (relative risk, 1.96; 95% CI, 1.68-2.30; p = 0.0001), with most of this difference dependent on a higher number of discontinuation for adverse events in the ritonavir group. After a mean follow-up of 307 days (ritonavir, 304; indinavir, 309), 124 deaths (ritonavir, 61; indinavir, 63; relative risk, 0.96; 95% CI, 0.67-1.36; p = 0.80) and 330 new AIDS-defining events (ritonavir, 170; indinavir, 160; relative risk, 1.05; 95% CI, 0.85-1.31; p = 0.60) were observed. CD4+ cell counts increased in both groups in patients still receiving treatment, with about 100 cells gained by week 24 and 150 cells gained by week 48. Body weight also increased over time in both groups. Analysis of RNA response showed a decrease of 1.5 log10 or higher in both treatment groups. Overall, 400 patients in the ritonavir group and 338 patients in the indinavir group developed at least one grade 3/serious new adverse event during follow-up (relative risk, 1.48; 95% CI, 1.28-1.72; p = 0.0001). Favorable CD4+ cell and RNA responses at 24 and 48 weeks were observed in both groups of patients remaining on treatment. Indinavir showed slightly better effects in sustaining RNA, CD4+ cell, and body weight responses. Ritonavir and indinavir results were comparable in terms of clinical outcome (survival and AIDS-defining events). PMID- 11118068 TI - Interaction between HIV type 1 glycoprotein 120 and CXCR4 coreceptor involves a highly conserved arginine residue in hypervariable region 3. AB - Several seven-transmembrane chemokine receptors are known to function as entry coreceptors for human immunodeficiency virus type 1. CCR5 and CXCR4 are the major coreceptors for non-syncytium-inducing (NSI) and syncytium-inducing (SI) viruses, respectively. During the natural course of infection, the emergence of variants with a phenotypic transition from NSI to SI and rapid disease progression is associated with expanded coreceptor usage to CXCR4. Characteristic amino acids at several positions in the hypervariable region 3 (V3) of gp120 have been linked to CXCR4 utilization. Previously, we reported that a highly conserved arginine residue of V3 played an important role in CCR5 utilization. In this study, the possible involvement of the same arginine residue in CXCR4 utilization was investigated. Amino acid substitutions introduced to this arginine on R5X4 viruses were found to have a significant effect on their utilization of CXCR4. These results, taken together with those reported previously, suggest that this highly conserved arginine may contribute to the functional convergence of chemokine coreceptor utilization by human immunodeficiency viruses and may represent a unique target for future antiviral design. PMID- 11118069 TI - Emergence of new forms of human immunodeficiency virus type 1 intersubtype recombinants in central Myanmar. AB - We have previously shown that HIV-1 env subtypes B' (a Thai-B cluster within subtype B) and E (CRF01_AE) are distributed in Yangon, the capital city of Myanmar. However, HIV strains from the rest of country have not yet been genetically characterized. In the present study, we determined env (C2/V3) and gag (p17) subtypes of 25 specimens from central Myanmar (Mandalay). Phylogenetic analyses identified 5 subtype C (20%), in addition to 10 CRF01_AE (40%) and 4 subtype B' (16%). Interestingly, the remaining six specimens (24%) showed discordance between gag and env subtypes; three gag subtype B'/env subtype C, one gag subtype B'/env subtype E, one gag subtype C/env subtype B', and one gag subtype C/env subtype E. These discordant specimens were found frequently among injecting drug users (4 of 12, 33%) and female commercial sex workers (2 of 8, 25%) engaging in high-risk behaviors. The recombinant nature of these HIV-1 strains was verified in three specimens, indicating the presence of new forms of HIV-1 intersubtype C/B' and C/B'/E recombinants with different recombination breakpoints. The data suggest that multiple subtypes of B', C, and CRF01_AE are cocirculating in central Myanmar, leading to the evolution of new forms of intersubtype recombinants among the risk populations exhibiting one of the highest HIV infection rates in the region. PMID- 11118070 TI - Molecular function of the CD4 D1 domain in coreceptor-mediated entry by HIV type 1. AB - The surface molecule CD4 plays a key role in initiating cellular entry by the human immunodeficiency virus type 1 (HIV-1), and it is now recognized as acting synergistically with select chemokine receptors (coreceptors) in the infection process. The present study was undertaken to determine whether the extracellular region of CD4 is sufficient to induce fusion of HIV-1 virions with target cells in the absence of its anchoring function. Using pseudotype reporter viruses to quantitate infection, soluble CD4 (sCD4) was tested for its ability to induce fusion by viruses utilizing CCR5 as their coreceptor. We found that sCD4 was competent to replace membrane-bound CD4 to trigger infection mediated by several HIV-1 envelopes. Furthermore, in a comparison of the envelopes of HIV-1 NL4-3 and a chimera containing the gp120 V3 loop of Ba-L, the V3 region was found to be one factor affecting susceptibility to induction by sCD4. In addition, using truncated and mutant derivatives of sCD4, the amino-terminal D1 domain of CD4 was found to be necessary and sufficient for induction of fusion and to require an intact gp120-binding site for this activity. These results delineate determinants on CD4 and gp120 required for fusion induction in collaboration with a coreceptor, and suggest a mechanism whereby CD4 may contribute to viral infection in trans. PMID- 11118071 TI - Functional and structural defects in HIV type 1 nef genes derived from pediatric long-term survivors. AB - DNA sequences and three distinct in vitro functions of Nef were evaluated in a group of seven perinatally infected children. nef gene sequences obtained before and after virus culture showed that one of the five non-/slow progressors harbored a virus with large deletions. nef genes from the remaining four children were full length but contained discrete changes at a higher frequency than the rapid progressors. In functional studies, 40 of 44 Nef proteins derived from the whole study group were capable of binding the cellular serine kinase p62, indicating that this function is well conserved among naturally occurring viruses. In contrast, representative Nef proteins derived from the long-term non /slow progressors were found to be defective or far less capable of enhancing viral replication and/or viral infectivity in herpesvirus saimiri-transformed human T cells and peripheral blood mononuclear cells. On reversion of highly prevalent point mutations in the defective proteins, viral replication could be restored to wild-type levels. Our results suggest that nef genes derived from pediatric long-term nonprogressors have gross deletions in isolated cases but a higher prevalence of discrete changes that may impair Nef function in primary T cell assays, but not all functions reported for Nef. PMID- 11118072 TI - Frequency of cytokine-producing T cells in HIV-infected patients treated with stavudine, didanosine, and ritonavir. AB - To assess prospectively the influence of the control of viral replication on the frequency of cytokine-producing T cells, and to correlate these changes with immune activation, we conducted a 15-month follow-up study of IFN-gamma- and IL-2 producing CD4+ and CD8+ T cells at a single-cell level in 12 previously untreated patients receiving highly active antiretroviral therapy (HAART). At baseline we observed a strikingly high proportion of IFN-gamma-producing CD8+ T cells. The treatment-induced decrease in the proportion of IFN-gamma-producing CD8+ T cells ran parallel to the decrease in HLA-DR+ and CD38+CD8+ T cell subsets and was associated with the reduction in HIV RNA level. IL-2-producing cells were mainly CD4+. As a consequence of CD4+ T cell loss, the number of IL-2-producing CD4+ T cells was lower in patients than in control subjects (52 vs. 171 cells/microl), but the proportion of these cells was unchanged (22.4 vs. 19.3). During therapy the proportion of CD4+ IL-2-producing cells was initially stable and then fell markedly at month 5, followed by a gradual return to previous values. The reduction in viral load was associated with the fall in the proportion of CD4+ activated subsets. Intracellular cytokine assays are a new approach to the assessment of T cell function in HIV infection. Our results suggest that the functional capacity of CD4+ T cells is probably less severely altered than previously thought on the basis of conventional assays. CD8+ T cells exhibit an increased capacity to produce IFN-gamma that is associated with an increase in activation marker expression. These alterations decrease partially and in parallel under treatment. PMID- 11118073 TI - Longitudinal changes in CD4+ T cell antigen receptor diversity and naive/memory cell phenotype during 9 to 26 months of antiretroviral therapy of HIV-infected patients. AB - Although skewing of the CD4+ TCR repertoire in advanced HIV infection is well documented, increases in polyclonality during antiretroviral therapy have been less consistently observed. Ten patients, each with documented abnormalities within the CD4+ TCR repertoire, were studied by CDR3 spectratyping, semiquantitative PCR, and SSCP during 9-26 months of therapy. Naive and memory cell phenotypes were analyzed by flow cytometry. Six of 10 patients showed increased polyclonality of their TCR repertoires, 1 showed no change, and 3 showed increased TCR skewing, despite suppressed viral replication. Overall, there was no significant change in the percentage of abnormal BV subfamilies (from a mean of 25.5 to 17.1%) or the percentage of naive CD4+ T cells (from a mean of 18 to 25%). Further, progression of TCR repertoire disruptions was observed in some patients even with suppression of plasma viral RNA below 500 copies/ml. Although a spectrum of changes may be seen within the CD4+ TCR repertoire in the setting of antiretroviral therapy, increases in polyclonality are observed in some patients. PMID- 11118074 TI - T cell responses to recall antigens, alloantigen, and mitogen of HIV-infected patients receiving long-term combined antiretroviral therapy. AB - The effect of highly active antiretroviral therapy (HAART) on T cell responses in 30 HIV-infected patients was studied. Lymphocyte proliferation in response to influenza A virus, HIV-1 p24, gp160, allogeneic leukocytes, and mitogen, as well as influenza-specific cytotoxic T lymphocyte (CTL) responses, were measured. AIDS patients had decreased T cell-proliferative responses to influenza and alloantigen compared with asymptomatic patients. Absence of positive proliferative responses of HIV-infected patients to HIV-1 antigens was not associated with increased interleukin 10 production. Correlation was observed between influenza-specific CTL response and T cell proliferation, as well as CD4+ T lymphocyte counts, indicating the importance of CD4+ helper T cells for generating antiviral CTL responses. Finally, these results show that HAART treated asymptomatic patients, but not AIDS patients, have T cell responses comparable to those of control individuals. It remains to be determined whether immune-based therapy will contribute any additional benefit to patients who received HAART. PMID- 11118075 TI - Effect of mycobacterial infection on virus loads and disease progression in simian immunodeficiency virus-infected rhesus monkeys. AB - The effect of a mycobacterial infection on AIDS disease was studied in the simian model. Monkeys were infected with the primary virulent isolate SIV/DeltaB670 and inoculated 90 days later with BCG, an attenuated strain of Mycobacterium bovis. All monkeys experienced a dramatic transient increase in plasma viremia and CCR5 expression on T lymphocytes after BCG inoculation. Only two of the four SIV+ animals had substantial proliferative responses to PPD, with poor responders developing disseminated BCG during the course of the experiment. BCG inoculation of SIV-infected long-term nonprogressor (LTNP) monkeys was also performed. Similar to the acutely infected animals, two of three LTNPs experienced increases in plasma viral levels and CCR5 expression. In the majority of animals studied, there was no accelerated progression to AIDS despite the concomitant transient stimulation of virus replication and CCR5 expression on T lymphocytes. PMID- 11118076 TI - Identification of a genetic subcluster of HIV type 1 subtype C (C') widespread in Ethiopia. AB - Others and we have previously shown that subtype C is the predominant HIV-1 subtype and the major cause of AIDS in Ethiopia. The present study shows that subtype C in Ethiopia has a genetic subcluster, designated C', has not increased in frequency, or spread geographically, over the period 1988 (%C' = 23/53) to 1996-1997 (%C' = 26/50). There is no association of the HIV-1 subtype C or subcluster C' with geographic location, time of sample collection, or risk group in Ethiopia. Of 105 randomly collected samples representing 7 different towns in Ethiopia, all but 2 (1 subtype A from Addis Ababa, 1997 and 1 subtype D from Dessie, 1996) belong to subtype C. PMID- 11118077 TI - Study of HIV type 1 gag/env variability in The Gambia, using a multiplex DNA polymerase chain reaction. AB - A multiplex DNA PCR assay was developed for the simultaneous first-round amplification of HIV-1 gag and env fragments for the heteroduplex mobility assay (HMA). This assay was compared with the conventional amplification assay, using DNA extracted from PBMC samples from 30 HIV-1-seropositive individuals from The Gambia, who were enrolled between 1992 and 1997. From 27 of 30 (90%) samples both gag and env HMA fragments were amplified simultaneously. In one sample only the gag HMA fragment could be amplified by multiplex DNA PCR, and in two samples amplification was negative for both gag and env HMA in multiplex as well as the mono-DNA PCR. Of the 28 Gambian isolates subtyped by gag/env HMA or by sequencing and phylogenetic analysis, the majority (19 of 28; 68%) were intersubtype recombinant. Fifteen of 28 (53%) samples were circulating recombinant form (CRF) CRF02.AG variants. Two isolates clustering with the previously documented Gambian isolate GM4 (previously described as an env GC recombinant) are classified as gag A/env J recombinants. PMID- 11118078 TI - Molecular characterization of human immunodeficiency virus type 1-infected individuals from bolivia reveals the presence of two distinct genetic subtypes B and F. AB - Thirty HIV-1-positive samples from Bolivia were genetically characterized on the basis of HMA and DNA sequencing, revealing the presence of B and F subtypes, in accordance with the molecular epidemiology pattern already described for other South American countries such as Brazil and Argentina. The interpatient divergence of subtype B Bolivian specimens was on average 14.2% (4.3-19.8%) at the nucleotide level, whereas the two unlinked subtype F samples (BO23 and BO29) were only 8.2% divergent, suggesting a more recent introduction of this subtype in the country. In our study group, which represents 13% of the HIV/AIDS cases already described in Bolivia as of May 1996, the transmission occurred more frequently through heterosexual exposures (46.7%), followed by homosexual (23.3%), bisexual (10%), intravenous drug use (3.3%), and vertical (3.3%); in one case the potential exposure category could not be defined (3.3%). No association could be established between exposure categories, gender, or clinical classification and subtype distribution in the Bolivian HIV/AIDS patients. PMID- 11118079 TI - Low mannose-binding lectin serum concentrations in HIV long-term nonprogressors? PMID- 11118080 TI - Finding a place for public preferences in liver allocation decisions. AB - Over the last decade there have been major advances in all aspects of liver transplantation with the consequence that the number of patients who could benefit from the procedure is increasing. As a result, the number of patients listed for liver transplantation is growing while the donor pool is remaining constant or even falling. The effect of this donor shortage is seen clearly both in Europe and in North America. For example, in North America data from UNOS shows that between 1988 and 1997 the number of cadaveric donor liver transplants rose from 1,713 to 4,100. The number of patients waiting for transplant rose over the same time from 616 to 9,647. This shortage of organs has tragic consequences. Although the proportion of patients dying on the waiting list is falling, the number of patients dying on the liver transplant waiting list increased from 196 to 1,129 over this same period of time. PMID- 11118081 TI - Selection of recipients for renal transplantation in European transplant centres. PMID- 11118082 TI - The selling of live donor nephrectomy: the old economy meets the new economy. PMID- 11118083 TI - Influence of long-term preservation with endobronchially administered perfluorodecalin on pulmonary graft function. AB - BACKGROUND: Experimental studies demonstrated a suppression of oxygen-derived free radicals, reduced adhesion of activated neutrophils on the endothelium and an increase of de novo synthesis of surfactant during liquid ventilation with perflurocarbon. The purpose of this study was to assess the pulmonary graft function after preservation with endobronchially administered perfluorocarbon as an alternative to flush perfusion. METHODS: Native bred pigs underwent orthotopic left lung transplantation. Donor lungs were flushed in situ with either a low potassium dextran solution (LPD, n=6) or a perfluorochemical was administered endobronchially (PFC, n=6) and were then stored after removal for 18 hr at 4 degrees C. Pulmonary graft function was assessed after reperfusion for 5 hr by measuring pulmonary gas exchange and hemodynamics during isolated ventilation and perfusion. Tissue specimens were taken for analysis of morphology and wet/dry ratio. All values were compared to a sham-operated group (n=6). RESULTS: Pulmonary gas exchange of the graft revealed reduced paO2 values and elevated paCO2 values in the PFC group throughout the observation period as compared with the LPD group and sham group. Endothelial alterations and fibrin exudate in the PFC group were significantly more pronounced. Lungs in the LPD group showed functional and morphological recovery close to sham group. CONCLUSIONS: Long-term preservation with endobronchially administered perfuorocarbon is possible. Impaired pulmonary graft function and pronounced morphological alterations indicate an aggravation of the ischemic reperfusion injury after lung transplantation compared to LPD preserved lungs. PMID- 11118084 TI - Mechanism and prevention of cold storage-induced human renal tubular cell injury. AB - BACKGROUND: The recent observation that cold storage of kidneys and tubular cells causes marked increase in free radical-catalyzed F2-isoprostanes suggests that radicals might be formed during cold storage. As cold temperature is associated with reduced metabolic and enzymic activity, the notion that cold temperature causes free radical production appeared less tenable. The objective was, therefore, to seek direct evidence for the free radical production during the cold storage of human renal tubular cells, and to define the roles of extrinsic and intrinsic antioxidants in cold-induced cell injury. METHODS: Human renal tubular cells were cold-stored at 4 degrees C for varying duration in University of Wisconsin solution and subjected to mRNA analysis, biochemical measurements, and cytoprotective studies. RESULTS: Cold storage caused a time-dependent reduction in glutathione levels, and an increase in the formation superoxide, hydrogen peroxide, and hydroxyl radicals. Cold-induced lactate dehydrogenase (LDH) release, ATP depletion, DNA damage, and membrane degradation were suppressed with the inclusion of antioxidant 2-methyl aminochroman or deferroxamine. The cells that were structurally protected with antioxidants were also intact functionally, as they had significantly improved cell proliferation. To examine the effect of cold on intrinsic antioxidant gene expression, antioxidant mRNA levels were analyzed using reverse transcription-polymerase chain reaction. The gene expression of mitochondrial Mn-superoxide dismutase (SOD), but not of cytosolic Cu,Zn-SOD or of glutathione peroxidase expression increased with cold exposure. The oxidant-sensitive gene heme oxygenase I increased slightly with 48-hr cold storage. CONCLUSIONS: Cold storage of human tubular cells causes marked increase in free radicals. These are likely of mitochondrial origin as there is a differential inducement of Mn-SOD gene, and are causal to cold-induced cell injury as extrinsic antioxidants abrogated the injury. Our findings support the strategy of adding antioxidants to preservation solutions or the strategy of pre-conditioning the organs to oxidative stress to minimize cold storage-induced organ damage. PMID- 11118085 TI - Nitric oxide diminishes apoptosis and p53 gene expression after renal ischemia and reperfusion injury. AB - BACKGROUND: The role of nitric oxide in the ischemic injury of the kidney is still controversial. The aim of this study was to reevaluate the beneficial effect of exogenous nitric oxide and define its effects as regulator of gene p53 expression and apoptosis in the ischemic renal injury. METHODS: Sprague-Dawley rats were subjected to 75 min of renal warm ischemia and contralateral nephrectomy. The animals were divided into six groups (n=6 per group): Two sham groups at 4 and 24 hr, two ischemic control (IC) at same times and two treated groups (Na-NP), studied at same intervals, where sodium nitroprusside (5 mg/kg) was given 15 min before reperfusion. The parameters evaluated included: serum creatinine, blood urea nitrogen, neutrophil infiltration determined by myeloperoxidase, gene p53 expression determined by reverse transcriptase polymerase chain reaction, apoptosis determined by peroxidase in situ technique and light histology. RESULTS: There were significant improvements in serum creatinine and blood urea nitrogen at 24 hr in the NA-NP group when compared with the IC group (P<0.05). Myeloperoxidase levels were higher in the IC when evaluated against the Na-NP groups. Na-NP exhibited a downregulating effect in the expression of gene p53 when compared to the IC group. Apoptosis was more evident in the IC group and had moderately increased histological damage when compared to the Na-NP group. CONCLUSIONS: Nitric oxide demonstrated a protective effect in the ischemic injury of the kidney and exerted an antiapoptotic action dowregulating the expression of gene p53. PMID- 11118086 TI - Ex vivo expansion of canine dendritic cells from CD34+ bone marrow progenitor cells. AB - BACKGROUND: The aims of this study were to ex vivo expand canine dendritic cells and determine their phenotype and functional characteristics. METHODS: CD34+ selected cells and CD34+-depleted canine bone marrow (BM) cells were cultured in Iscove's modified medium for 14 days. Cytokines added to the cultures included human granylocyte/macrophage colony-stimulating factor 5 ng/ml, hFlt3 ligand 200 ng/ml, and human tumor necrosis factor-alpha 10 ng/ml. Cultured cells and purified subpopulations were assessed for cell surface antigen expression, morphology, and function by flow cytometric analysis, electron microscopy, and an allogeneic mixed lymphocyte reaction at day 14. RESULTS: Two main cell populations were identified, DR++(bright)/CD14- and DR+(dim)/CD14+. Ex vivo expanded CD34+-selected cells showed increased allostimulatory activity compared to both cultured CD34+-depleted cells and mononuclear cells. In contrast, ex vivo expansion from CD34+-depleted cells was unsuccessful. After sorting cells from the ex vivo expanded CD34+-selected bone marrow to enrich for DR++/CD14- cells, a 42-fold increase (median) of allostimulatory activity was observed as compared with sorted DR+/CD14+ cells (P=0.02). CONCLUSIONS: Cells with dentric cell-like phenotypes and functions can be cultured from canine CD34+-selected bone marrow cells. Future studies will address the roles of these cells in engraftment, graft versus host reactions and graft-host tolerance in a canine hematogoietic stem cell transplantaton model. PMID- 11118087 TI - Prevention of graft-versus-host disease by a novel immunosuppressant, PG490-88, through inhibition of alloreactive T cell expansion. AB - BACKGROUND: PG490-88 is a water soluble, semisynthetic derivative of a novel compound PG490 (triptolide) purified from the Chinese herb Tripterygium Wilfordii Hook F. METHODS: PG490-88 was administrated into recipient mice in a model (B10.D2-->BALB/c) of lethal graft-versus-host disease (GVHD) to study the effects of PG490-88 on GVHD and on the various steps involved in the pathological course of GVHD. RESULTS: Injection of PG490-88 i.p. at a dose of 0.535 mg/kg/day for the first 3 weeks after transplantation protected all the recipients from developing GVHD up to 100 days after transplantation. PG490-88 inhibited in vivo both CD4+Vbeta3+ and CD8+Vbeta3+ T cell (alloreactive T cells in this model) expansion in the spleen by 64.09 and 34.02%, respectively, at the time when Vbeta3+ cell expansion was in the logarithmic phase (day 3 after transplantation). Intracellular cytokine staining without further in vitro activation demonstrated 47.42% inhibition of IL-2 production among CD4+ spleen cells in PG490-88-treated mice as compared to GVHD control on day 3 after transplantation. In contrast, CD25 (alpha chain of interleukin-2 receptor) expression did not differ. CONCLUSIONS: PG490-88 is highly effective in prevention of murine GVHD. The immunosuppressive effect of PG490-88 is mediated by inhibition of alloreactive T cell expansion through interleukin-2 production. PMID- 11118088 TI - Prognostic factors for long-term actual patient survival after orthotopic liver transplantation in children. AB - BACKGROUND: Orthotopic liver transplantation has become the treatment of choice for children with end-stage liver disease. Although results have improved the last decades, still a considerable number of children die after transplantation. The aim of this study was to analyze long-term actual survival and to identify prognostic factors for such survival rates. METHODS: A consecutive series of 66 children receiving transplants who had or could have had a follow-up of at least 5 years was retrospectively analyzed. Actual survival and prognostic factors in relation to patient, donor, and operation related variables were assessed after multivariate analysis. RESULTS: Actual 1-, 3-, and 5-year patient survival was 86%, 79%, and 73%, respectively. A high Child-Pugh (C-P) score or C-P class C, high donor age, high blood loss index, and retransplantation were predictive factors for actual patient survival. A high blood loss index was correlated with biliary atresia, low recipient age and weight, and with previous upper abdominal operations. The duration of stay of the donor at the intensive care unit (ICU) was a predictive factor for retransplantation. CONCLUSIONS: Children with diseases eligible for liver transplantation should be seen early in the course of their disease in a transplantation center. All possible measures should be taken during the transplantation procedure to keep the blood loss at a minimum. Children with biliary atresia deserve special attention in this respect. The choice of donors has implications for survival. PMID- 11118089 TI - The influence of cytomegalovirus viraemia on the outcome of recurrent hepatitis C after liver transplantation. AB - BACKGROUND: Several interrelated host and hepatitis C virus (HCV) associated factors have been proposed to explain the variable outcomes in HCV recurrence. Recent evidence suggests that cytomegalovirus (CMV) infection not only is co factor in progression of HCV recurrence but may precipitate allograft rejection. We investigated whether short-term CMV viremia influences HCV recurrence, the number and grade of acute rejection episodes, and the histological course of HCV recurrence during the first year after orthotopic liver transplantation (OLT) for HCV-related cirrhosis. METHODS: A cohort of 39 patients transplanted for cirrhosis HCV-related was analyzed. Patients were evaluated twice weekly for CMV infection by a blood polymerase chain reaction (PCR) assay. Triple therapy with cyclosporine or tacrolimus, azathioprine and prednisolone was the initial immunosuppressive regimen. Preemptive treatment with ganciclovir was started when two consecutive PCRs for CMV were positive. Liver biopsies were performed on day 7 after OLT or when indicated. A 3-day IV 1 g methilprednisolone was given to patients with moderate or severe rejection. Ishak's score was used to grade inflammation and to stage fibrosis. RESULTS: Neither CMV viremia nor CMV disease after OLT for HCV-related cirrhosis adversely influenced the incidence and grade of acute rejection episodes nor the histological outcome of post transplant HCV recurrence, during the first year after liver transplantation. CONCLUSION: CMV viremia as detected by PCR does not affect the progression of HCV recurrence in liver grafts. PMID- 11118090 TI - Profile of anemia in children after liver transplantation. AB - BACKGROUND: Clinical and hematological profile of chronic anemia in children after orthotopic liver transplantation (OLT) is unknown. METHODS: We prospectively studied children after orthotopic liver transplantation (OLT) with hemoglobin levels < 2 standard deviation of age appropriate mean for > 6 months. Investigations included hemogram, reticulocyte count, peripheral blood smear, serum vitamin B-12, folic acid levels, iron studies, Coomb's tests, serum erythropoietin (EPO) levels, and stool and urine tests for occult blood. RESULTS: Fifty-six participants (22 male and 34 female, mean age 82.9 months, range 20 232, mean post-OLT duration 48.8 months, range 6-132) were studied. The causes of anemia were idiopathic (32), iron deficiency (4), viral infections (2, HIV=1, parvovirus=1), and lymphoproliferative disease (2). Fifteen participants showed spontaneous recovery within 1-6 months. Thirty-one children with idiopathic anemia had low or normal EPO levels (mean 7.33 mmicro/L, range <2.5 to 15.9, normal 4-24). When outliers (iron deficiency=4, HIV disease= 1) were excluded, there was no statistical correlation between hematocrits and EPO levels. Serum vitamin B-12 levels (n=52) were elevated (normal 110-930 pg/ml) (mean=1,186 pg/ml) in 32 (61.5%) and were significantly higher in those with abnormal liver function tests. CONCLUSION: Anemia is a common problem in children after OLT. More than half the participants had anemia of unknown etiology with an inappropriate EPO response for the degree of anemia. The normal negative correlation between hematocrit and EPO was lost in these children. The observation regarding serum vitamin B-12 levels requires further study. PMID- 11118091 TI - Valaciclovir prophylaxis of cytomegalovirus infection and disease in renal transplantation: an economic evaluation. AB - BACKGROUND: Cytomegalovirus (CMV) disease is a major cause of morbidity and mortality in solid organ transplant patients and is associated with large additional healthcare expenditures. An economic evaluation of valaciclovir CMV prophylaxis in a renal transplant population is reported. METHODS: Medical resource use data were collected alongside a multicenter multinational randomized, placebo-controlled, double-blind trial of valaciclovir CMV prophylaxis in renal transplantation. Patients were stratified into donor seropositive/recipient sero-negative (D+R-) and recipient seropositive (R+) groups. Patients were followed-up 6 months posttransplant. A cost-effectiveness analysis from the perspective of the French health care system was performed using the number of cases of CMV disease avoided at 6 months as the clinical endpoint. RESULTS: Resource use was significantly increased among patients who developed CMV disease compared to those who did not develop disease. In the high risk D+R- group, valaciclovir prophylaxis was associated with an average of 5.5 fewer inpatient hospital days (P < OR =0.05) and with significantly lower use of other healthcare resources. In the R+ group, valaciclovir prophylaxis prevented cases of CMV disease at a marginally greater mean cost per patient compared with placebo. For D+R- patients valaciclovir prophylaxis was therefore an economically superior strategy, resulting in fewer cases of CMV disease and lower total mean healthcare expenditures. CONCLUSIONS: Valaciclovir CMV prophylaxis in renal transplantation is a more cost-effective therapy compared with placebo, in the high-risk D+R- patient population. For the R+ group, the incremental cost per case of CMV disease was modest. PMID- 11118092 TI - Combined liver-pancreas transplantation in a patient with primary sclerosing cholangitis and insulin-dependent diabetes mellitus. AB - Combined liver-pancreas transplantation is a relatively uncommon procedure. We report successful combined liver-pancreas transplantation in a patient with primary sclerosing cholangitis and insulin-dependent diabetes mellitus and review the literature on this topic. PMID- 11118093 TI - Extracorporeal liver perfusion using human and pig livers for acute liver failure. AB - BACKGROUND: Patients with fulminant hepatic failure (FHF) often die awaiting liver transplantation. Extracorporeal liver perfusion (ECLP) has been proposed as a method of "bridging" such patients to transplantation. We report the largest experience to date of ECLP using human and porcine livers in patients with acute liver failure. METHODS: Patients with FHF unlikely to survive without liver transplantation were identified. ECLP was performed with human or porcine livers. Patients underwent continuous perfusion until liver transplantation or withdrawal of support. Two perfusion circuits were used: direct perfusion of patient blood through the extracorporeal liver and indirect perfusion with a plasma filter between the patient and the liver. FINDINGS: Fourteen patients were treated with 16 livers in 18 perfusion circuits. Nine patients were successfully "bridged" to transplantation. ECLP stabilized intracranial pressure (ICP) and cerebral perfusion pressure (CPP). Arterial ammonia levels fell from a median of 146 to 83 micromol/liter within 12 hr and this reduction was maintained at least 48 hr. Pig and human ECLP lowered ammonia levels equally. Serum bilirubin levels also fell from a median of 385 to 198 micromol/liter over the first 12 hr but the response was not sustained as well with porcine livers. There was no immunological benefit to using the the filtered perfusion circuit. INTERPRETATION: These data demonstrate that ECLP is safe and can provide metabolic support for comatose patients with fulminant hepatic failure for up to 5 days. While labor and resource intensive, this technology is available to centers caring for patients with acute liver failure and deserves wider evaluation and application. PMID- 11118094 TI - Risk of nonmelanoma skin cancer in Italian organ transplant recipients. A registry-based study. AB - BACKGROUND: Organ transplant recipients are at an increased risk of nonmelanoma skin cancer. Few data concern heart transplantation and populations from southern Europe. METHODS: A total of 1,329 patients who received their first kidney (1,062 subjects) or heart allograft (267 subjects) were included in a partly retrospective cohort study to evaluate the risk of skin cancer. The incidence rate per 1,000 person-years and the cumulative incidence were computed. Standardized morbidity ratio was estimated by comparison with Italian cancer registry data. To analyze the role of potential prognostic factors, Cox's regression method was used. RESULTS: The overall incidence rate of nonmelanoma skin cancer was 10.0 cases per 1,000 posttransplant person-years (95% confidence interval 8.2-11.7). This estimate was far higher than expected in the general population. The overall risk of developing skin cancer increased from a cumulative incidence of 5.8% after 5 posttransplant years to an incidence of 10.8% after 10 years of graft survival. In a Cox proportional hazard risk model, the most important factors that appeared to favor the development of skin cancer were age at transplantation and sex. After adjustment for age at transplantation and sex, no definite increased risk was documented among heart as compared with kindney transplant recipients. CONCLUSIONS: Our study confirms the increased risk of nonmelanoma skin cancer among organ transplant recipients in a southern European population. PMID- 11118095 TI - The impact of recipient cytokine genotype on acute rejection after renal transplantation. AB - BACKGROUND: Acute allograft rejection remains an important cause of morbidity after kidney transplantation, and has been shown to be a crucial determinant of long-term graft function. As cytokines are major regulators of the immune system, genetic variation in cytokine production or activity may influence susceptibility to acute rejection. This study sought to determine the impact of recipient cytokine and cytokine receptor polymorphisms on acute rejection after renal transplantation. METHODS: A total of 209 cadaveric renal transplant recipients were selected for analysis according to the presence or absence of graft rejection in the first 30 days after transplantation. DNA was genotyped for 22 polymorphisms in 11 cytokine and cytokine receptor genes using the polymerase chain reaction with sequence specific primers. Results were stratified by incidence and severity of rejection, and by HLA-DR mismatching. RESULTS: No association between any polymorphism and the incidence or severity of acute rejection was detected. In particular, no association was seen with tumor necrosis factor or interleukin-10 genotype, either alone or in combination. CONCLUSIONS: We have failed to demonstrate any association between recipient cytokine genotype and acute rejection after cadaveric renal transplantation. Although more extensive studies may disprove these findings, it would seem premature to use recipient cytokine genotyping to predict transplant outcome, or to guide immunosuppressive therapy after transplantation. PMID- 11118096 TI - Practice variations in the evaluation of adult candidates for cadaveric kidney transplantation: a survey of the European Transplant Centers. AB - BACKGROUND: This survey was conducted to investigate similarities and differences in the diagnostic evaluation of adult candidates for cadaveric renal transplantation and the criteria for acceptance to the cadaveric renal transplant waiting-list in the European transplant centers. METHODS: A questionnaire listing 45 diagnostic procedures (consultations of 9 specialties, 18 imaging techniques and 18 laboratory investigations), 45 medical conditions constituting possible reasons for exclusion from renal transplantation, and 10 properties characterizing the responding transplant center was sent to 214 European transplant centers. RESULTS: A completed questionnaire was returned by 154 of 214 centers (72%). Significant disagreement (P<0.001) exists about the necessity of 28 of the 45 surveyed diagnostic procedures and about the acceptability of transplant candidates for 15 of the 45 surveyed medical conditions. The influence of center characteristics on the observed practice variations was examined by multinomial logistic regression (factors: Center size, waiting-list pressure, responsibility for organizing the diagnostic work-up, status of transplant center, responsibility for decision about acceptance of candidates and geographic location of center): In 13 of 28 controversial diagnostic procedures, geographic location of the centers turned out to be the only significant determining factor (P<0.001), whereas the dissent about medical conditions is not influenced significantly by the analyzed factors. CONCLUSION: The detected significant practice variations in the evaluation of renal transplant candidates may either indicate where scientific evidence is missing and more clinical research is needed or where the existing evidence has not been adequately disseminated and convincing guidelines should be established. PMID- 11118097 TI - Activation of apoptotic and inflammatory pathways in dysfunctional donor hearts. AB - BACKGROUND: Myocardial dysfunction is common after brain death, but the mechanisms remain unclear. Apoptosis is tightly regulated by enzymes termed the caspases. We have investigated the caspases involved in the terminal part of the apoptotic pathway in dysfunctional (nontransplanted) donor hearts and their relation to inflammatory markers and compared them to hearts with good ventricular function (transplanted donors). METHODS: Thirty-one donor hearts assessed for transplantation were examined. Western blotting was used to measure pro-caspase-9, caspase-3, DFF45, the activated nuclease CPAN and poly (ADP ribose) polymerase, a DNA repair enzyme inactivated by caspase-3. Caspase-3 activity was also measured. Histologic and immunocytochemical analysis for HLA Class II and Real Time polymerase chain reaction for tumor necrosis factor-alpha and interleukin 6 were performed to detect inflammatory activation. RESULTS: Cleaved caspase-9 was higher (5.53+/-0.6 vs. 3.64+/-0.4 O.D. units, P<0.01) in nontransplanted compared with transplanted donors and there was a trend for higher pro-caspase-9 (5.20+/-1.0 vs. 4.22+/-0.4 O.D. units, P=NS). Levels of pro caspase-3 were higher in nontransplanted (9.66+/-0.5 vs. 5.15+/-0.5 O.D. units, P<0.00001) donors and cleavage products of caspase-3 were elevated in 14 of 14 nontransplanted and 2 of 17 transplanted donors. Intact DFF-45 (8.94+/-0.36 vs. 6.14+/-0.30 O.D. units, P<0.000005), its spliced product (2.38+/-0.35 vs. 0.4+/ 0.21 O.D. units, P=0.0001) and the nuclease caspase-activated nuclease (2.01+/ 0.3 vs. 0.66+/-0.16 OD units, P=0.001) were higher in nontransplanted donors. The caspase-3 substrate poly (ADP-ribose) polymerase was higher in nontransplanted (1.16+/-0.13 vs. 0.61+/-0.22 O.D. units, P=0.57) donors. CONCLUSIONS: The caspases are elevated in dysfunctional donor hearts compared with hearts with good ventricular function with a possible link to inflammatory activation supporting the concept that brain death causes inflammatory activation which can lead to apoptosis with a possible important effect on function. PMID- 11118098 TI - Ex vivo generation of effective Epstein-Barr virus (EBV)-specific CD8+ cytotoxic T lymphocytes from the peripheral blood of immunocompetent Epstein Barr virus seronegative individuals. AB - BACKGROUND: Although readily accomplished from immunocompetent Epstein-Barr virus (EBV) seropositive individuals, the effective ex vivo generation of EBV-specific cytotoxic T lymphocytes (CTL) from the peripheral blood mononuclear cells (PBMC) of EBV-seronegative subjects has proven to be a challenge. The focus of our study was to ascertain optimized culture conditions required for the ex vivo generation of EBV-reactive autologous CTL from the PBMC of EBV-seronegative volunteers. METHOD: Freshly isolated PBMC obtained from immunocompetent EBV-seronegative and seropositive individuals were used to generate EBV-specific autologous CTL lines using both conventional and a novel, modified ex vivo culture technique. RESULTS: In contrast to responses observed in EBV-seropositives after two to three rounds of ex vivo stimulation, gamma-irradiated autologous lymphoblastoid cell lines (LCL) were incapable of eliciting an effective anti-EBV cytotoxic response when freshly-isolated PBMC from EBV-seronegative individuals were used as responders. Under these culture conditions, CD4+ T cells with preferential expression of the Th2-type cytokine IL-4 were predominantly expanded in the PBMC obtained from EBV seronegative individuals. However, the addition of recombinant human (rh) IL-12 during the primary phase of ex vivo stimulation resulted in augmentation of EBV specific cytolysis of autologous LCL by CD8+ T cells. Furthermore, there was down regulation in the secretion of IL-4 and up-regulation in that of the Th1-type cytokine IFN-gamma by responder CD4+ and CD8+ T cells. CONCLUSIONS: Taken together these data suggest that the addition of rhIL-12 during the primary phase of ex vivo stimulation of freshly isolated PBMC from EBV-seronegative individuals results in skewing of the immune response predominantly towards a CD4+ Th1-type (IFN-gamma) with the generation of an efficacious CTL-mediated anti-EBV reactivity. This novel ex vivo approach for generating effective autologous EBV specific CTL could be adopted to treat refractory post-transplant lymphoproliferative disorders, which may be encountered in EBV-seropositive-->EBV seronegative organ transplant recipients. Additionally, these ex vivo generated anti-EBV T cells could also be infused perioperatively to enhance prophylactically immunity against EBV infection in high-risk EBV-seronegative organ allograft recipients. PMID- 11118099 TI - Heterogeneity of T cell clones specific for a single indirect alloantigenic epitope (I-Ab/H-2Kd54-68) that mediate transplant rejection. AB - BACKGROUND: One of the complexities of solid organ allograft rejection is the inherent diversity of the specific T cell antigenic epitopes that participate in this response, including the role of direct alloantigen recognition and indirect recognition of donor-derived peptides in recipient antigen-presenting cells. To probe the role of distinct T cell receptor (TCR) avidity differences and the role of cytokine expression patterns of different effector T cells that may participate in allograft rejections, we have identified a dominant allopeptide derived from the H-2Kd molecule, recognized by H-2b CD4 T cells in the context of syngeneic I-Ab. METHODS: To identify a stimulatory peptide derived from the H-2Kd molecule, a panel of synthetic overlapping peptides was screened for immunogenicity and a panel of T cell clones established. These clones were characterized for TCR Vbeta usage by mAb staining and/or reverse transcribed polymerase chain reaction analysis, peptide dose sensitivity as a marker of TCR avidity, cytokine expression phenotype in vitro, and their ability to mediate rejection of a vascularized cardiac allograft after adoptive transfer to immunodeficient mice. RESULTS: The H-2Kd54-68 peptide was identified as a dominant stimulatory peptide by the ability of T cells from C57BL/6 (H-2b) mice primed by a combination of allogeneic spleen cell injection and mixed peptide immunization to mount an in vitro proliferative response and interferon-gamma production by peptide stimulation. Furthermore, direct immunization with synthetic H-2Kd54-68 peptide of normal C57BL/6 mice resulted in accelerated rejection of both skin and cardiac allografts from B10.D2 (H-2d) mice, but not 3rd party B10.BR (H-2k) grafts. A panel of 15 distinct CD4+ clones specific for H 2Kd54-68 peptide were established and shown to utilize a variety of TCR Vbeta and different apparent TCR avidities to H-2Kd54-68 peptide when stimulated in vitro. To characterize these clones further, two clones were chosen based on the difference of avidity to H-2Kd54-68 peptide. The cytokine expression pattern was determined and indirect alloantigen specificity confirmed by analysis of responses to purified peptide and B10.D2 spleen cells using normal H.2b and I Abeta chain knockout mice as APC donors. Both of these T cell clones were able to mediate rejection of B10.D2, but not B10.BR hearts, in immunodeficient mice, but the morphological pattern of T cell infiltration was distinct. CONCLUSIONS: These results demonstrate the potential importance of fine dissection of the alloantigeneic response to solid organ transplants and provide unique insights into the role of TCR avidity and cytokine expression patterns in different morphological patterns of transplant rejection. PMID- 11118100 TI - Nondepleting anti-CD4 monoclonal antibody enhances the ability of oral alloantigen delivery to induce indefinite survival of cardiac allografts: oral tolerance to alloantigen. AB - BACKGROUND: We examined whether oral administration of alloantigen could induce the prolonged survival of cardiac allografts. METHODS: Hearts from CBK (H2k+Kb) transgenic or (C57BL/10xCBA)F1 (H2bxH2k) mice were transplanted into CBA (H2k) recipients pretreated orally with 1 x 10(7) donor splenocytes in the presence or absence of a nondepleting anti-CD4 (YTS 177, 200 microg/dose). RESULTS: Modest prolongation of CBK cardiac grafts was induced in CBA mice fed with multiple doses of CBK splenocytes (MST 42 days compared with controls fed with syngeneic CBA splenocytes, 12 days). When the CD4 monoclonal antibody, YTS177, was administered for 2 days before the first oral delivery of CBK splenocytes, all mice accepted their grafts indefinitely (MST > 100 days versus mice treated with anti-CD4 alone, 11.5 days). To determine if feeding multiple doses of alloantigen was essential, CBA mice were given CBK splenocytes orally on a single occasion in combination with the anti-CD4. The majority of the grafts survived indefinitely (MST >100 days). This oral treatment regimen also induced indefinite prolongation of (C57BL/10xCBA)F1 cardiac grafts. CONCLUSION: The induction of unresponsiveness by oral administration of alloantigen can be augmented by a nondepleting anti CD4, YTS177, when given before the first oral delivery of allogeneic cells. PMID- 11118101 TI - Embolization of a mesenteric arteriovenous fistula following pancreatic allograft: the steal effect. AB - The first pancreatic transplant was performed in 1966 by Kelly and Lillehei at the University of Minnesota (1, 2). Nearly 30 years later, Ozaki et al., published the first occurrence of an arteriovenous fistula (AVF) in a transplanted allograft bundle (3). From its early days as a radical and experimental procedure in the treatment of insulin-dependent diabetes mellitus, this operation has progressed to become routine in many major medical centers. However, the incidence of technical complications remains relatively high, ranging from 10 to 40% (1). The list of potential complications includes allograft pancreatitis, vascular thrombosis, hemorrhage, pseudoaneurysm formation, anastomotic leaks, intra-abdominal infections, and, al. though rare, AVF. Lowell et al., in 1993, reported this last complication in 3 of 90 consecutive pancreatic recipients (4). These same authors promoted the theory that AVF formation was directly related to procurement technique: a nonspecific "blind ligation" of mesenteric vessels along the inferior pancreatic border. However, this approach continues to be used commonly. We have identified the occurrence of an allograft superior mesenteric artery-superior mesenteric vein (SMA-SMV) AVF in a pancreas-after-kidney (PAK) transplant recipient. PMID- 11118102 TI - A high panel-reactive antibody rescue protocol for cross-match-positive live donor kidney transplants. AB - BACKGROUND: Alloimmunization can present a virtually insurmountable barrier to kidney transplantation. Past protocols to desensitize patients using plasmapheresis and cyclophosphamide have not been broadly applied because of the fear of complications, including high rates of immunologic failure. METHODS: Fifteen patients with a positive donor-recipient cross-match were desensitized with plasmapheresis to permit live donor (LD) transplantation under newer maintenance immunosuppressants. Pretransplant the patients received plasmapheresis three times weekly for a planned maximum of six treatments, plus intravenous hyperimmune globulin, tacrolimus, mycophenolate mofetil, and prednisone. Patients who were successfully desensitized and received transplants were given 10 days of OKT3 postoperatively. RESULTS: Eleven of the 15 patients became anti-human globulin cross-match-negative after one to five plasmapheresis treatments and underwent LD transplantation. Relatively low initial titers of donor-specific antibody were predictive of successful attainment of a negative cross-match. Few side effects and rejection episodes were observed. All transplant patients remain dialysis-free after 3-26 months of follow-up. CONCLUSION: A positive cross-match is not necessarily a contraindication to LD transplantation, especially for patients with low donor-specific alloantibody titers. PMID- 11118103 TI - Postshunt encephalopathy in liver transplanted children with portal vein thrombosis. AB - BACKGROUND: Surgical portosystemic shunting has been reported to alleviate successfully portal hypertension in liver transplanted recipients with portal vein thrombosis. METHODS: We report two liver transplanted children with portal vein thrombosis who developed post-shunt acute encephalopathy. In one child, a mesocaval H-type shunt was created surgically because of bleeding related to Roux en-Y loop varices at 3 months posttransplantation; in the other, a large spontaneous splenorenal shunt was discovered at the time of diagnosis of portal vein thrombosis on day 34 posttransplantation and was preserved. RESULTS: Post shunt encephalopathy developed 6 months and 2.7 years after transplantation, causing death in one child. CONCLUSIONS: This report illustrates the risk and the possible dismal outcome of post-shunt encephalopathy in liver transplanted children. Therapeutic procedures other than portosystemic shunting that will restore an hepatopetal portal flow to the liver graft should be considered in liver-transplanted children with portal vein thrombosis. PMID- 11118104 TI - Orlistat decreases the plasma level of cyclosporine and may be responsible for the development of acute rejection episodes. PMID- 11118105 TI - Co-administration of orlistat and cyclosporine in a heart transplant recipient. PMID- 11118106 TI - Too many confounding variables. PMID- 11118107 TI - Prevention of bone loss with alendronate in kidney transplant recipients. PMID- 11118108 TI - Laparoscopic donor nephrectomy: pro. PMID- 11118109 TI - Laparoscopic donor nephrectomy: con. PMID- 11118110 TI - Cation-pi effects in the complexation of Na+ and K+ with Phe, Tyr, and Trp in the gas phase. AB - Na+ and K+ gas-phase affinities of the three aromatic amino acids Phe, Tyr, and Trp were measured by the kinetic method. Na+ binds these amino acids much more strongly than K+, and for both metal ions the binding strength was found to follow the order Phe < or = Tyr < Trp. Quantum chemical calculations by density functional theory (DFT) gave the same qualitative ordering, but suggested a somewhat larger Phe/Trp increment. These results are in acceptable agreement with predictions based on the binding of Na+ and K+ to the side chain model molecules benzene, phenol, and indole, and are also in reasonable agreement with the predictions from purely electrostatic calculations of the side-chain binding effects. The binding energies were compared with those to the aliphatic amino acids glycine and alanine. Binding to the aromatic amino acids was found to be stronger both experimentally and computationally, but the DFT calculations indicate substantially larger increments relative to alanine than shown by the experiments. Possible reasons for this difference are discussed. The metal ion binding energies show the same trends as the proton affinities. PMID- 11118112 TI - Observation of amide anions in solution by electrospray ionization mass spectrometry. AB - Negative quasimolecular ions of aromatic carboxylic acid amides have been observed unexpectedly under electrospray ionization conditions. Hypothetically, deprotonation of either carboxamide or carboximidic acid tautomers can produce anions with equivalent resonance structures, the stability of which is affected by conjugated aromatic substituents. In this study, a series of meta and para substituted benzamides were analyzed using electrospray ionization mass spectrometry in aqueous methanolic solutions. The degree of ionization was found to be pH dependent and was enhanced by electron-withdrawing substituents and suppressed by electron-donating groups. The observed effect on apparent acidity can be accounted for by resonance stabilization. PMID- 11118111 TI - Leaving group and gas phase neighboring group effects in the side chain losses from protonated serine and its derivatives. AB - The gas phase fragmentation reactions of protonated serine and its YNHCH(CH2X)CO2H derivatives, beta-chloroalanine, S-methyl cysteine, O-methyl serine, and O-phosphoserine, as well as the corresponding N-acetyl model peptides have been examined via electrospray ionization tandem mass spectrometry (MS/MS). In particular, the competition between losses from the side chain and the combined loss of H2O and CO from the C-terminal carboxyl group of the amino acids or H2O or CH2CO from the N-acetyl model peptides are compared. In this manner the effect of the leaving group (Y = H or CH3CO, vary X) or of the neighboring group can be examined. It was found that the amount of HX lost from the side chain increases with the proton affinity of X [OP(O)(OH)2 > OCH3 approximately equals OH > Cl]. The ion due to the side chain loss of H2O from the model peptide N acetyl serine is more abundant than that from protonated serine, suggesting that the N-acetyl group is a better neighboring group than the amino group. Ab initio calculations at the MP2(FC)/6-31G*//HF/6-31G* level of theory suggest that this effect is due to the transition state barrier for water loss from protonated N acetyl serine being lower than that for protonated serine. The mechanism for side chain loss has been examined using MS3 tandem mass spectrometry, independent synthesis of proposed product ion structures combined with MS/MS, and hydrogen/deuterium exchange. Neighboring group rather than cis 1,2 elimination processes dominate in all cases. In particular, the loss of H3PO4 from O phosphoserine and N-acetyl O-phosphoserine is shown to yield a 3-membered aziridine ring and 5-membered oxazoline ring, respectively, and not the dehydroalanine moiety. This is in contrast to results presented by DeGnore and Qin (J. Am. Soc. Mass Spectrom. 1998, 9, 1175-1188) for the loss of H3PO4 from larger peptides, where dehydroalanine was observed. Alternate mechanisms to cis 1,2 elimination, for the formation of dehydroalanine in larger phosphoserine or phosphothreonine containing peptides, are proposed. PMID- 11118113 TI - Proton affinity of uracil. A computational study of protonation sites. AB - Relative stabilities of uracil tautomers and cations formed by gas-phase protonation were studied computationally with the B3LYP, MP2, QCISD, and QCISD(T) methods and with basis sets expanding from 6-31G(d,p) to 6-311+G(3df,2p). In accordance with a previous density functional theory study, the dioxo tautomer 1a was the most stable uracil isomer in the gas phase. Gibbs free energy calculations using effective QCISD(T)/6-311+G(3df,2p) energies suggested >99.9% of 1a at equilibrium at 523 K. The most stable ion isomer corresponded to N-1 protonated 2,4-dihydroxypyrimidine, which however is not formed by direct protonation of 1a. The topical proton affinities in 1a followed the order O-8 > O 7 > C-5 > N-3 > N-1. The thermodynamic proton affinity of 1a was calculated as 858 kJ mol(-1) at 298 K. A revision is suggested for the current estimate included in the ion thermochemistry database. PMID- 11118114 TI - Measurement of collision-induced dissociation rates for tantalum oxide ions in a quadrupole ion trap. AB - A study of factors influencing the collision-induced dissociation (CID) rate of strongly bound diatomic ions effected via resonance excitation in a quadrupole ion trap is presented. From these studies, an approach to measuring the CID rates is described wherein product ion recovery is optimized and the effect of competitive processes (e.g., parent ion ejection and product ion reactions) on rate measurements are prevented from influencing rate measurements. Tantalum oxide ions (dissociation energy = 8.2 eV), used as a model system, were formed via reactions of glow discharge generated Ta+ ions with residual gases in the ion trap. Neon (0.5 mtorr) was found to be a more favorable target gas for the dissociation of TaO+ than He and Ar, but collisional activation of TaO+ ions in neon during ion isolation by mass selective instability necessitated ion cooling prior to dissociation. A 25 ms delay time at qz = 0.2 allowed for kinetic cooling of stored TaO+ ions and enabled precise dissociation rate measurements to be made. CID of TaO+ was determined to be most efficient at qz = 0.67 (226 kHz for m/z 197). Suitable resonance excitation voltages and times ranged from 0.56 to 1.2 V(p-p) and 1 to 68 ms, respectively. Under these conditions, measurement of rates approaching 80 s(-1) for the dissociation of TaO+ could be made without significant complications associated with competing processes, such as ion ejection. PMID- 11118115 TI - Simultaneous analysis of butene isomer mixtures using process mass spectrometry. AB - The feasibility of simultaneous analysis of mixtures containing two to four butene isomers and up to six total components using process mass spectrometry is assessed. As for typical (nonisomeric) applications of process mass spectrometry, simultaneous analysis is based on the assumption that the electron ionization mass spectra of mixtures are linear combinations of the spectra of the individual constituents. Limits of detection for binary isomer mixtures are on the order of 0.1% to 10%, limited by the ability to distinguish small differences between similar spectra. As spectral and mixture complexity increase, both accuracy and precision decrease. Not surprisingly the similarity of the spectra of stereoisomers cis- and trans-2-butene is greater than that of the other (nonstereoisomeric) isomer pairs, and mixtures containing both cis- and trans-2 butene are the most difficult to quantitate. However, even for mixtures of all four butenes, accuracy (root-mean-square error = 2.43%), precision (average coefficient of variation = 6.72%), and linearity (correlation coefficient of a plot of measured versus actual concentration r2 = 0.985 +/- 0.002) are reasonably good. PMID- 11118116 TI - Utilization of MS3 spectra for the multicomponent quantification of diastereomeric N-acetylhexosamines. AB - A rapid and accurate means of quantifying mixtures of diastereomeric N acetylhexosamine monosaccharides using MS3 product ions is introduced. The method involves derivatizing the monosaccharides with [Co(DAP)2Cl2]Cl (where DAP is diaminopropane), and subjecting the derivatized products to collision-induced dissociation (CID) in a quadrupole ion trap mass spectrometer. Each diastereomer provides unique MS3 product ion abundances. The abundances for the pure monosaccharide standards are used in a system of equations in order to quantify mixtures of these diastereomers. Using the system of equations is quite advantageous, as it is the only mass spectrometric method that has been shown to successfully quantify mixtures of more than two isomers. The utility of the method is demonstrated by successfully quantifying various two and three component mixtures of the diastereomeric monosaccharides. Furthermore, the method is used to quantify the recovery of a single diastereomeric monosaccharide from an acidic resin. Although the multicomponent quantification method described herein is used to quantify mixtures of N-acetylhexosamine diastereomers, it could be applied to any group of isomers, provided distinguishing CID spectra are obtained. This is the first known report of utilizing MS3 product ions for quantification of structural isomeric mixtures. PMID- 11118117 TI - Post-source decay in the analysis of polystyrene by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - Various secondary series are observed in matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectra of polystyrene. The number and positions of the series depend on the choice of matrix and added cation. For a given treatment, series observed in linear mode are not necessarily observed in reflectron mode, and vice versa. Post-source decay analysis was used to determine that the secondary series arise at least in part from formation and decay of adducts of polystyrene with matrix species. There is some treatment-to treatment variation, but adduct formation and decay were observed for all tested treatments. The multiplicity of secondary series makes it unclear whether post source decay occurs for the main series (polystyrene + cation)+ ions under the conditions normally used for polystyrene analysis. Such ions do undergo post source decay at laser fluences greater than normally used. Although only polystyrene was investigated in this work, other polymers may also produce adduct and decay series in MALDI analysis. Their presence can mask the presence of minor components in a sample, but at least as observed here, do not have a strong influence on molecular mass determinations. PMID- 11118118 TI - A new approach for effecting surface-induced dissociation in an ion cyclotron resonance mass spectrometer: a modeling study. AB - With the increasing use of ion cyclotron resonance (ICR) for tandem mass spectrometry (MS/MS) analysis of biomolecules, surface-induced dissociation (SID) should be given serious consideration as an ion activation technique. There are at least two compelling reasons to consider SID: it can deposit significant amounts of internal energy into large ions, and no collision gas is required. These potential advantages have led us to undertake a modeling study of the SID process in an ICR using the ion optics program SIMION. The various methods previously used to obtain SID spectra are compared to a new approach for effecting SID in an ICR. Through simulations, many different parameters present in the experiment are correlated to the kinetic energy of the parent ion upon impact and the overall product ion collection efficiency (and hence the signal intensity) expected. The modeling results suggest this new approach allows larger, more precise, and controllable impact energies to be used, as well as providing higher collection efficiencies. The validity of the modeling results is supported by good qualitative agreement with previously reported experimental results. PMID- 11118119 TI - Use of a kinetic energy orifice as a probe of metastable dissociation in Fourier transform ion cyclotron resonance mass spectrometry. AB - Although Fourier transform ion cyclotron resonance mass spectrometry is a powerful tool in the qualitative observation of gas phase reactions, ion detection is on the millisecond time scale, orders of magnitude longer than typically found when using a sector instrument. Observations of short-lived species such as chemically activated adduct ions can be accomplished using selective ion excitation as a probe of intermediate lifetime. Whereas ion elimination has been shown to be effective in monitoring ion lifetimes on the microsecond time scale, problems associated with detecting ions produced with high kinetic energies limits the technique. Use of a kinetic energy orifice as an ion skimmer effectively eliminates ions near the center of the ion cell at relatively low kinetic energies. By modifying a single section cell to include a kinetic energy orifice, the lifetimes of chemically activated adduct ions have been investigated. PMID- 11118120 TI - Evaluation of carrier gases for use in high-field asymmetric waveform ion mobility spectrometry. AB - Effects of carrier gas type (N2, O2, CO2, N2O, and SF6) on changes in the ratio of high- to low-field ion mobility, Kh/K, of cesium, gramicidin S, tetrahexylammonium, heptadecanoic acid, and aspartic acid in fields of up to 67 Td are presented. The theory of the mobility of ions at high E/N in different gases is discussed. Plots of Kh/K as a function of the ionic energy parameter, E/N, for the five ions in each of the gases were derived from experimental data collected using a high-field asymmetric waveform ion mobility spectrometer. The change in the ratio of high- to low-field ion mobility of cesium in carrier gases of O2 and N2 showed excellent agreement with literature values. The behavior of cesium in O2 and N2 is used to illustrate that the ratio Kh/K as a function of effective temperature is invariant with gas type as long as the well depth of the interaction potential significantly exceeds thermal energy. From these results, it appears that the well depth of the interaction potential of the heavier ions studied here, including gramicidin S, tetrahexylammonium, and heptadecanoic acid, with bath gases such as N2 and O2, is shallow relative to thermal energy. PMID- 11118121 TI - Simultaneous quantitation of the 5'-triphosphate metabolites of zidovudine, lamivudine, and stavudine in peripheral mononuclear blood cells of HIV infected patients by high-performance liquid chromatography tandem mass spectrometry. AB - A high-performance liquid chromatography (HPLC) method utilizing triple quadrupole mass spectrometry (MS) detection was developed and validated for the simultaneous measurement of the intracellular nucleoside 5'-triphosphate anabolites of zidovudine (ZDV-TP), lamivudine (3TC-TP), and stavudine (d4T-TP). These compounds were extracted from patient peripheral blood mononuclear cells (PBMCs) which are the sites of HIV replication and drug action. Ion-exchange solid phase extraction (SPE) followed by enzymatic digestion with alkaline phosphatase was utilized to yield the measurable nucleoside forms of the nucleotides. Reversed phase C-18 SPE with addition of a nucleoside internal standard, 3'-azido-2',3'-dideoxyuridine (AzdU) allowed for the indirect measurement of the original 5'-triphosphate concentration by HPLC/MS/MS. Quantitation was performed from calibration curves generated from authentic 5' triphosphate standards spiked in PBMCs from healthy volunteers. Analytical range for the three 5'-triphosphates was equivalent to 50-45,000 pg. Mean interassay accuracies for 3TC-TP, d4T-TP, and ZDV-TP (n > 90) were 99.4%, 100.1%, and 108.0%, respectively. Mean interassay precisions (%C.V.) for 3TC-TP, d4T-TP, and ZDV-TP (n > 90) were 8.8%, 10.4%, and 8.2%, respectively. Recovery of the extraction method was 79.2%, 83.1%, and 98.3% for 3TC-TP, d4T-TP, and ZDV-TP, respectively. This method can be utilized to measure the intracellular 5' triphosphate levels in HIV infected patients receiving antiretroviral therapy containing the nucleoside reverse transcriptase inhibitors 3TC, d4T, or ZDV. PMID- 11118130 TI - Proposed nomenclature for mating type genes of filamentous ascomycetes. PMID- 11118131 TI - Molecular organization of mating type loci in heterothallic, homothallic, and asexual Gibberella/Fusarium species. AB - Mating type (MAT) genes were cloned from three members of the Gibberella/Fusarium complex that differ in reproductive mode: heterothallic G. fujikuroi, homothallic G. zeae, and asexual F. oxysporum. The G. fujikuroi MAT locus organization is typical of other heterothallic pyrenomycetes characterized to date; i.e., there are three genes at MAT1-1 and one at MAT1-2. G. zeae has homologues of all four genes encoded by the two G. fujikuroi MAT idiomorphs, tightly linked on the same chromosome, interspersed with sequences unique to G. zeae. Field isolates of F. oxysporum, although asexual, have either the MAT1-1 or the MAT1-2 genes found in sexual species and these genes are highly similar to those of heterothallic G. fujikuroi. RT-PCR analysis proved that the F. oxysporum MAT genes are expressed and that all putative introns found in each of the four MAT genes in G. fujikuroi and F. oxysporum are removed. Apparent failure of F. oxysporum to reproduce sexually could not be attributed to mutations in the MAT genes themselves. PMID- 11118132 TI - Phylogenetic species recognition and species concepts in fungi. AB - The operational species concept, i.e., the one used to recognize species, is contrasted to the theoretical species concept. A phylogenetic approach to recognize fungal species based on concordance of multiple gene genealogies is compared to those based on morphology and reproductive behavior. Examples where Phylogenetic Species Recognition has been applied to fungi are reviewed and concerns regarding Phylogenetic Species Recognition are discussed. PMID- 11118133 TI - The control of meiosis progression in the fungus Coprinus cinereus by light/dark cycles. AB - Meiosis progression in Coprinus cinereus is controlled by light/dark cycles. Light is essential to propel basidia into karyogamy and light intensity determines the timing of meiotic events. The higher the light intensities, the faster the fruiting bodies enter karyogamy. The critical period when light has this influence is between 16 and 6 h before karyogamy. The control is highly stage specific. A 3-h dark period is essential for a Java dikaryon and the Japanese A(mut)B(mut) homokaryon to enter meiotic metaphase; without it the fruit body is permanently arrested at diffused diplotene. This arrest is light intensity-dependent (>20 hlx) and temperature-dependent (e.g., 27 degrees C). The placement of the dark period is very stage specific; it has no effect when placed before karyogamy stage. A dikaryon of London origin is light blind and able to complete meiosis under continuous high light regime. Fruiting bodies arrested under a continuous high light can be rescued by a 3-h dark treatment, but there is always an 8-h lag time to enter meiotic metaphase. It is possible that the dark effect signals cellular processes leading to division events. Cytological studies of arrested fruiting bodies showed that chromosomes are normal in meiotic prophase through pachytene and diplotene, but are unable to undergo chromosome condensation. Genetic crosses between a monokaryon of Java stock J6;5.4 and a monokaryon BL55 or H5 of London stock showed that light-blindness is dominant, and is controlled by a single Mendelian gene. PMID- 11118134 TI - Ca2+ regulation of Phyllosticta ampelicida pycnidiospore germination and appressorium Formation. AB - Phyllosticta ampelicida conidia germinate only after making contact with and attaching to a substratum. Previous studies suggested a role for Ca2+ in this process. A Ca2+ buffering system was used to control the external free Ca2+ concentration. Both germination and appressorium formation were reduced or abolished with low Ca2+ (less than or equal to nanomolar levels) but were nearly 100% at millimolar levels of Ca2+. Germination initiation required Ca2+ within 10 25 min after the spore made contact with the substratum. Appressorium initiation required Ca2+ 90-120 min following initial contact. Ca2+ channel blockers nicardipine and lanthanum abated spore development. TMB-8, a blocker of internal Ca2+ channels, reduced both developmental events. Gadolinium, a putative stretch activated Ca2+ channel blocker, abolished both developmental events at nanomolar levels. Calmodulin antagonists, compounds R-24751 and 48/80, abated spore development at micromolar levels. Together, these results suggest that Ca2+ signaling is involved in both germination and appressorium formation in P. ampelicida pycnidiospores. PMID- 11118135 TI - The isolation of FOS-1, a gene encoding a putative two-component histidine kinase from Aspergillus fumigatus. AB - In fungi, two-component histidine kinases are involved in response mechanisms to extracellular changes in osmolarity, resistance to dicarboximide fungicides, and cell-wall assembly. In the human opportunistic fungus, Candida albicans, each of the three histidine kinases plays a role in virulence. Here, we identify, for the first time, a gene, FOS-1, from the human pathogenic fungus Aspergillus fumigatus that predicts a protein with homology to two-component histidine kinases. The predicted FOS-1 protein is highly homologous to bacterial and other fungal histidine kinases in several functional domains, but is divergent at the amino- and carboxy-termini. A mutant lacking the FOS-1 locus, DeltaFOS-1, did not exhibit a detectable defect in either hyphal growth or morphology when grown on solid or liquid medium. However, in liquid medium, conidiophore development of the DeltaFOS-1 mutant was delayed. Compared to wild type, the DeltaFOS-1 strain was neither osmotically sensitive nor sensitive or resistant to a number of nondicarboximide antifungal drugs, but was highly resistant to dicarboximide fungicides and resistant to novozym 234, suggesting that FOS-1p may play a role in the regulation of cell-wall assembly. PMID- 11118136 TI - The ecological risks and benefits of genetically engineered plants. AB - Discussions of the environmental risks and benefits of adopting genetically engineered organisms are highly polarized between pro- and anti-biotechnology groups, but the current state of our knowledge is frequently overlooked in this debate. A review of existing scientific literature reveals that key experiments on both the environmental risks and benefits are lacking. The complexity of ecological systems presents considerable challenges for experiments to assess the risks and benefits and inevitable uncertainties of genetically engineered plants. Collectively, existing studies emphasize that these can vary spatially, temporally, and according to the trait and cultivar modified. PMID- 11118137 TI - Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes. AB - The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription. PMID- 11118138 TI - Orchestrated transcription of key pathways in Arabidopsis by the circadian clock. AB - Like most organisms, plants have endogenous biological clocks that coordinate internal events with the external environment. We used high-density oligonucleotide microarrays to examine gene expression in Arabidopsis and found that 6% of the more than 8000 genes on the array exhibited circadian changes in steady-state messenger RNA levels. Clusters of circadian-regulated genes were found in pathways involved in plant responses to light and other key metabolic pathways. Computational analysis of cycling genes allowed the identification of a highly conserved promoter motif that we found to be required for circadian control of gene expression. Our study presents a comprehensive view of the temporal compartmentalization of physiological pathways by the circadian clock in a eukaryote. PMID- 11118139 TI - The origins of genomic duplications in Arabidopsis. AB - Large segmental duplications cover much of the Arabidopsis thaliana genome. Little is known about their origins. We show that they are primarily due to at least four different large-scale duplication events that occurred 100 to 200 million years ago, a formative period in the diversification of the angiosperms. A better understanding of the complex structural history of angiosperm genomes is necessary to make full use of Arabidopsis as a genetic model for other plant species. PMID- 11118140 TI - Strange magnetism and the anapole structure of the proton. AB - The violation of mirror symmetry in the weak force provides a powerful tool to study the internal structure of the proton. Experimental results have been obtained that address the role of strange quarks in generating nuclear magnetism. The measurement reported here provides an unambiguous constraint on strange quark contributions to the proton's magnetic moment through the electron-proton weak interaction. We also report evidence for the existence of a parity-violating electromagnetic effect known as the anapole moment of the proton. The proton's anapole moment is not yet well understood theoretically, but it could have important implications for precision weak interaction studies in atomic systems such as cesium. PMID- 11118141 TI - Molybdenum nanowires by electrodeposition. AB - Metallic molybdenum (Mo(o)) wires with diameters ranging from 15 nanometers to 1.0 micrometers and lengths of up to 500 micrometers (0.5 millimeters) were prepared in a two-step procedure. Molybdenum oxide wires were electrodeposited selectively at step edges and then reduced in hydrogen gas at 500 degrees C to yield Mo(o). The hemicylindrical wires prepared by this technique were self uniform, and the wires prepared in a particular electrodeposition (in batches of 10(5) to 10(7)) were narrowly distributed in diameter. Wires were obtained size selectively because the mean wire diameter was directly proportional to the square root of the electrolysis time. The metal nanowires could be embedded in a polystyrene film and lifted off the graphite electrode surface. The conductivity and mechanical resiliency of individual embedded wires were similar to those of bulk molybdenum. PMID- 11118142 TI - High-resolution inkjet printing of all-polymer transistor circuits. AB - Direct printing of functional electronic materials may provide a new route to low cost fabrication of integrated circuits. However, to be useful it must allow continuous manufacturing of all circuit components by successive solution deposition and printing steps in the same environment. We demonstrate direct inkjet printing of complete transistor circuits, including via-hole interconnections based on solution-processed polymer conductors, insulators, and self-organizing semiconductors. We show that the use of substrate surface energy patterning to direct the flow of water-based conducting polymer inkjet droplets enables high-resolution definition of practical channel lengths of 5 micrometers. High mobilities of 0.02 square centimeters per volt second and on-off current switching ratios of 10(5) were achieved. PMID- 11118143 TI - Ultrahigh-density nanowire arrays grown in self-assembled diblock copolymer templates. AB - We show a simple, robust, chemical route to the fabrication of ultrahigh-density arrays of nanopores with high aspect ratios using the equilibrium self-assembled morphology of asymmetric diblock copolymers. The dimensions and lateral density of the array are determined by segmental interactions and the copolymer molecular weight. Through direct current electrodeposition, we fabricated vertical arrays of nanowires with densities in excess of 1.9 x 10(11) wires per square centimeter. We found markedly enhanced coercivities with ferromagnetic cobalt nanowires that point toward a route to ultrahigh-density storage media. The copolymer approach described is practical, parallel, compatible with current lithographic processes, and amenable to multilayered device fabrication. PMID- 11118144 TI - Creating long-lived superhydrophobic polymer surfaces through mechanically assembled monolayers. AB - We show that elastomeric surfaces can be tailored using "mechanically assembled monolayers" (MAMs), structures that are fabricated by combining self-assembly of surface grafting molecules with mechanical manipulation of the grafting points in the underlying elastic surface. The versatility of this surface modification method is demonstrated by fabricating MAMs with semifluorinated (SF) molecules. These SF-MAMs have superior nonwetting and barrier properties in that they are "superhydrophobic" and nonpermeable. We also establish that these material characteristics do not deteriorate even after prolonged exposure to water, which usually causes surface reconstruction in conventionally prepared SF self assembled monolayers. PMID- 11118145 TI - External control of 20th century temperature by natural and anthropogenic forcings. AB - A comparison of observations with simulations of a coupled ocean-atmosphere general circulation model shows that both natural and anthropogenic factors have contributed significantly to 20th century temperature changes. The model successfully simulates global mean and large-scale land temperature variations, indicating that the climate response on these scales is strongly influenced by external factors. More than 80% of observed multidecadal-scale global mean temperature variations and more than 60% of 10- to 50-year land temperature variations are due to changes in external forcings. Anthropogenic global warming under a standard emissions scenario is predicted to continue at a rate similar to that observed in recent decades. PMID- 11118146 TI - Extended life-span conferred by cotransporter gene mutations in Drosophila. AB - Aging is genetically determined and environmentally modulated. In a study of longevity in the adult fruit fly, Drosophila melanogaster, we found that five independent P-element insertional mutations in a single gene resulted in a near doubling of the average adult life-span without a decline in fertility or physical activity. Sequence analysis revealed that the product of this gene, named Indy (for I'm not dead yet), is most closely related to a mammalian sodium dicarboxylate cotransporter-a membrane protein that transports Krebs cycle intermediates. Indy was most abundantly expressed in the fat body, midgut, and oenocytes: the principal sites of intermediary metabolism in the fly. Excision of the P element resulted in a reversion to normal life-span. These mutations may create a metabolic state that mimics caloric restriction, which has been shown to extend life-span. PMID- 11118147 TI - Docosahexaenoic acid, a ligand for the retinoid X receptor in mouse brain. AB - The retinoid X receptor (RXR) is a nuclear receptor that functions as a ligand activated transcription factor. Little is known about the ligands that activate RXR in vivo. Here, we identified a factor in brain tissue from adult mice that activates RXR in cell-based assays. Purification and analysis of the factor by mass spectrometry revealed that it is docosahexaenoic acid (DHA), a long-chain polyunsaturated fatty acid that is highly enriched in the adult mammalian brain. Previous work has shown that DHA is essential for brain maturation, and deficiency of DHA in both rodents and humans leads to impaired spatial learning and other abnormalities. These data suggest that DHA may influence neural function through activation of an RXR signaling pathway. PMID- 11118148 TI - Global analysis of the genetic network controlling a bacterial cell cycle. AB - This report presents full-genome evidence that bacterial cells use discrete transcription patterns to control cell cycle progression. Global transcription analysis of synchronized Caulobacter crescentus cells was used to identify 553 genes (19% of the genome) whose messenger RNA levels varied as a function of the cell cycle. We conclude that in bacteria, as in yeast, (i) genes involved in a given cell function are activated at the time of execution of that function, (ii) genes encoding proteins that function in complexes are coexpressed, and (iii) temporal cascades of gene expression control multiprotein structure biogenesis. A single regulatory factor, the CtrA member of the two-component signal transduction family, is directly or indirectly involved in the control of 26% of the cell cycle-regulated genes. PMID- 11118149 TI - The bacterial flagellar cap as the rotary promoter of flagellin self-assembly. AB - The growth of the bacterial flagellar filament occurs at its distal end by self assembly of flagellin transported from the cytoplasm through the narrow central channel. The cap at the growing end is essential for its growth, remaining stably attached while permitting the flagellin insertion. In order to understand the assembly mechanism, we used electron microscopy to study the structures of the cap-filament complex and isolated cap dimer. Five leg-like anchor domains of the pentameric cap flexibly adjusted their conformations to keep just one flagellin binding site open, indicating a cap rotation mechanism to promote the flagellin self-assembly. This represents one of the most dynamic movements in protein structures. PMID- 11118150 TI - Development of CD8alpha-positive dendritic cells from a common myeloid progenitor. AB - Dendritic cells (DCs) are critical in both initiating adaptive immune responses and maintaining tolerance to self antigens. These apparently contradictory roles have been suggested to depend on different subsets of DCs that arise from either myeloid or lymphoid hematopoietic origins, respectively. Although DC expression of CD8alpha is attributed to a lymphoid origin, here we show that both CD8alpha+ and CD8alpha- DCs can arise from clonogenic common myeloid progenitors in both thymus and spleen. Thus, expression of CD8alpha is not indicative of a lymphoid origin, and phenotypic and functional differences among DC subsets are likely to reflect maturation status rather than ontogeny. PMID- 11118152 TI - Introduction: new perspectives in chronic renal insufficiency. PMID- 11118153 TI - Chronic renal insufficiency: current understandings and their implications. AB - Chronic renal insufficiency (CRI) is underrecognized and undertreated even in sophisticated health care systems. This is particularly distressing in light of the growing number of effective interventions available to slow the progression of kidney disease and ameliorate many of its comorbid conditions. Progress, to a certain extent, is impeded by the lack of generally accepted definitions, clear diagnostic criteria, and practical screening tests. Available epidemiologic data suggest that 800,000 Americans have creatinine levels >/=2.0 mg/dL and over 6 million have levels >/=1.5 mg/dL. Population-based surveys show that age, male gender, and black race are predictors of kidney disease. For reasons that are not well understood, the incidence of kidney failure has nearly doubled over the last 15 years, indicating a parallel increase in CRI. This trend has significant economic implications. Other implications of CRI related to the use and availability of health care resources are appropriate referrals to nephrologists and early intervention to optimize care of patients with CRI. A multipronged approach to providing optimal care involves interventions that may delay the progression of renal dysfunction, proactive prevention of uremic complications, measures to forestall the progress of comorbid conditions, and timely preparation of patients for renal replacement therapy. Early recognition of CRI, expeditious referral for specialty care, and the utilization of a comprehensive program of care optimization will help meet the nation's goals as articulated in the National Institutes of Health's Healthy People 2010: Chronic Kidney Disease. PMID- 11118151 TI - Functional requirement for class I MHC in CNS development and plasticity. AB - Class I major histocompatibility complex (class I MHC) molecules, known to be important for immune responses to antigen, are expressed also by neurons that undergo activity-dependent, long-term structural and synaptic modifications. Here, we show that in mice genetically deficient for cell surface class I MHC or for a class I MHC receptor component, CD3zeta, refinement of connections between retina and central targets during development is incomplete. In the hippocampus of adult mutants, N-methyl-D-aspartate receptor-dependent long-term potentiation (LTP) is enhanced, and long-term depression (LTD) is absent. Specific class I MHC messenger RNAs are expressed by distinct mosaics of neurons, reflecting a potential for diverse neuronal functions. These results demonstrate an important role for these molecules in the activity-dependent remodeling and plasticity of connections in the developing and mature mammalian central nervous system (CNS). PMID- 11118154 TI - Defining a renal anemia management period. AB - Estimates of the prevalence of anemia during chronic renal insufficiency (CRI) vary depending on how anemia is defined. An analysis of patients beginning dialysis in the United States found that 67% had a hematocrit of less than 30% and 51% had a hematocrit of less than 28%. The anemia of CRI, therefore, appears to be prevalent even as it is underrecognized and undertreated-despite the widespread realization that there is much to be gained by treatment with recombinant human erythropoietin, with little risk of accelerating the progression of kidney disease. It is difficult to separate the effects of anemia in CRI from those of other comorbid conditions, but it is clear that anemia is a strong predictor of mortality and cardiac morbidity. Correction of anemia would be expected to negate the contribution of anemia to the mortality and cardiac morbidity associated with CRI. While this hypothesis is well-validated in hemodialysis patients, data in the population with CRI are preliminary but encouraging. Recent small prospective studies have established that treatment of anemia with recombinant human erythropoietin can reverse some degree of the cardiac morphological changes seen in CRI. While awaiting the results of large long-term clinical trials, the concept of the renal anemia management period (RAMP) draws attention and focus to the need for proactive and aggressive treatment of anemia among patients with CRI. The RAMP is defined as the period of time after the onset of CRI during which anemia develops, requiring diagnosis and treatment. Treatment of anemia during the RAMP has the potential to ameliorate, or even prevent, significant future morbidity in patients with CRI. PMID- 11118155 TI - Cardiovascular disease in chronic renal insufficiency. AB - Cardiovascular illness is an important contributor to the morbidity of kidney disease. The spectrum of cardiovascular disease (CVD) in patients with chronic renal insufficiency (CRI) includes left ventricular hypertrophy (LVH) and dilatation, ischemic heart disease, and peripheral vascular disease. Both "traditional" and "uremia-specific" factors contribute to the occurrence and progression of cardiac disease in renal patients. A growing body of recent evidence indicates that the processes contributing to CVD commence early in CRI, leading to concentric LVH, left ventricular dilatation, congestive heart failure, and ischemic heart disease. Many of the coexisting conditions that have been identified consistently as contributing to the burden of cardiovascular illness in renal populations can be modified through medical interventions. Specific therapies exist for hypertension, anemia, hyperparathyroidism, and dyslipidemia. Studies to date have demonstrated that treatment of many of these factors-such as anemia and hypertension during end-stage renal disease-appear to benefit the cardiovascular system. Earlier intervention may offer the best opportunity to reduce the burden of illness in all groups of CRI patients. Identification of patients at the onset of kidney disease and attention to the known traditional and "uremic" risk factors are emerging as promising strategies. Long-term interventional studies are needed to determine costs, benefits, and risks of such strategies. PMID- 11118156 TI - Complications of chronic renal insufficiency: beyond cardiovascular disease. AB - The less rigorous attention to the management of the complications of chronic renal insufficiency (CRI) and its comorbid conditions has potentially tragic consequences. In fact, with early recognition and intervention, many of the complications of CRI and its comorbid conditions can be ameliorated or prevented. We review here the most prevalent, troublesome, and potentially preventable complications and comorbidities of CRI with a view toward developing high quality, cost-effective strategies for delivering early interventional care. Complications of CRI include malnutrition, anemia, disorders of divalent ion metabolism and osteodystrophy, metabolic acidosis, and dyslipidemia. Important comorbid conditions of CRI are hypertension, diabetes mellitus, and cardiovascular disease. Clinical intuition suggests that early intervention will avert morbidity related to the hypoalbuminemia and other nutritional disorders of CRI, the metabolic acidosis, and the dyslipidemias, but prospective data are lacking at present. Correction of anemia, usually with recombinant human erythropoietin, may be key to the prevention of cardiac disease and other comorbidities of CRI. Incipient disorders of bone and mineral metabolism are managed prospectively using such measures as protein restriction to reduce phosphorus intake, phosphate binders, calcium supplementation, and vitamin D analogues. Hypertension, whatever its original etiology, is clearly an important risk factor for the progression of kidney failure and for the development of diffuse vascular disease; appropriate and aggressive treatment is essential. In patients with diabetic nephropathy, the principles of both primary and secondary prevention have been validated in several large trials of glycemic and blood pressure control. The seeds of these insidious, challenging, and costly comorbid conditions are sown very early in CRI, at a time when they are-in theory-most amenable to intervention. We therefore must be as proactive as possible in the timely implementation of relatively simple therapies that have the potential to prevent some of these adverse outcomes of CRI. PMID- 11118157 TI - Anemia management in patients with chronic renal insufficiency. AB - The introduction of recombinant human erythropoietin (rHuEPO) more than a decade ago provided the first effective treatment for the anemia of chronic renal insufficiency (CRI). The use of rHuEPO in the treatment of anemia has been associated with partial regression of left ventricular hypertrophy among both dialysis and nondialysis patients, and has been shown to reduce the frequency of cardiac complications such as congestive heart failure and number of days of hospitalization among dialysis patients. Despite this evidence, the anemia of CRI remains highly prevalent, underrecognized, and undertreated. A number of considerations arise regarding the management of anemia among patients with CRI. In this article, we review the rationale for treatment of anemia, current management practices, proposed treatment strategies, and the economic implications of improved anemia treatment. PMID- 11118158 TI - Research directions: new clinical frontiers. AB - One of the greatest remaining challenges facing nephrology research is obtaining data with detail and precision for the three large, yet "forgotten," populations that span the spectrum of kidney disease: patients with chronic renal insufficiency (CRI), peritoneal dialysis patients, and kidney transplant patients. Studies of these populations, particularly the CRI group, are hampered by the relative mobility of these patients, the lack of stringent epidemiologic or clinical definitions, and the tendency to extrapolate data from hemodialysis populations into other clinical settings. This article suggests a two-pronged approach to a research agenda: first, by recognizing the need for better data regarding the natural history of these kidney failure subsets and their comorbidities; and second, by directing greater effort at identifying rational, efficacious, and cost-effective interventions to influence their natural history positively. Specific efforts are suggested in all three populations. For patients with CRI, studies should be directed at (1) identifying high-risk patients; (2) determining methods for making optimal referrals to the nephrologist; (3) identifying and managing CRI, its complications, and its comorbid conditions; and (4) establishing processes for the smooth transition to dialysis. The peritoneal dialysis population will benefit from studies addressing the treatment of anemia and its ability to modify cardiovascular illness and quality of life. Kidney transplant studies should also focus on the identification and management of comorbid conditions, as well as the effects of various interventions on quality of life. Rational evidence-based care of these conditions, which are critically important to patients, their families, and the health care system in general, must await the conduct of well-designed prospective observational and interventional trials. PMID- 11118159 TI - Using injury data for violence prevention. Government proposal is an important step towards safer communities. PMID- 11118160 TI - Self management in asthma care. Professionals must rethink their role if they are to guide patients successfully. PMID- 11118161 TI - Screening for familial hypercholesterolaemia. Effective, safe treatments and dna testing make screening attractive. PMID- 11118162 TI - Emerging arboviral encephalitis. Newsworthy in the West but much more common in the East. PMID- 11118164 TI - Managing chronic disease-round 2 PMID- 11118163 TI - Research misconduct: Britain's failure to act. PMID- 11118165 TI - Plans for tackling research fraud may not go far enough. PMID- 11118166 TI - University must tell patients that they were research "guinea pigs" PMID- 11118167 TI - In brief PMID- 11118168 TI - Public's satisfaction with the NHS declines. PMID- 11118169 TI - Gut cells engineered to produce insulin PMID- 11118170 TI - Trusts must use, not just collect, clinical data. PMID- 11118171 TI - Early x ray for low back pain confers little benefit. PMID- 11118172 TI - US doctors consider access to health care for all. PMID- 11118173 TI - Untying the strings. PMID- 11118175 TI - Outcome of case finding among relatives of patients with known heterozygous familial hypercholesterolaemia. AB - OBJECTIVES: To assess the feasibility of detecting new cases of heterozygous familial hypercholesterolaemia by using a nurse led genetic register. DESIGN: Case finding among relatives of patients with familial hypercholesterolaemia. SETTING: Two lipid clinics in central and south Manchester. SUBJECTS: 259 (137 men and 122 women) probands and 285 first degree relatives. RESULTS: Of the 200 first degree relatives tested, 121 (60%) had inherited familial hypercholesterolaemia. The newly diagnosed patients were younger than the probands and were generally detected before they had clinically overt atherosclerosis. Concentrations of serum cholesterol were, respectively, 8.4 (1.7 SD) mmol/l and 8.1 (1.9 SD) mmol/l in affected men and women and 5.6 (1.0 SD) mmol/l and 5.6 (1.1 SD) mmol/l in unaffected men and women. Screening for risk factors as recommended in recent guidelines for coronary heart disease prevention would have failed to identify most of the affected relatives in whom hypertension, diabetes mellitus, cigarette smoking, and obesity were uncommon. CONCLUSIONS: By performing cholesterol tests on 200 relatives, 121 new patients with familial hypercholesterolaemia were discovered. Because 1 in 500 people in the United Kingdom are affected by this condition, to detect a similar number by population screening over 60 000 tests would be required, and only a few of these patients would have been detected had cholesterol testing been restricted to those with other risk factors for coronary heart disease. A case exists for organising a genetic register approach, linking lipid clinics nationally. PMID- 11118176 TI - Relation between private health insurance and high rates of caesarean section in Chile: qualitative and quantitative study. AB - OBJECTIVES: To explore the circumstances and factors that explain the association between private health insurance cover and a high rate of caesarean sections in Chile. DESIGN: Qualitative analysis of audiotaped in-depth interviews with obstetricians and pregnant women; quantitative analysis of data from face to face semistructured interview survey conducted postnatally (with women who had given birth in the previous 24-72 hours), and of a review of medical notes at a public hospital, a university hospital, and a private clinic. SETTING: Santiago, Chile. PARTICIPANTS: Qualitative arm: 22 obstetricians, 21 pregnant women; quantitative arm: 540 postnatal women. MAIN OUTCOME MEASURES: Rates of caesarean section in different types of institutions; consultants' views on private practice; work patterns in private practice; women's reasons for choosing private care; women's preferences on method of delivery. RESULTS: Private health insurance cover requires the primary maternity care provider to be an obstetrician. In the postnatal survey, women with private obstetricians showed consistently higher rates of caesarean section (range 57-83%) than those cared for by midwives or doctors on duty in public or university hospitals (range 27-28%). Only a minority of women receiving private care reported that they had wanted this method of delivery (range 6-32%). With the diversification in the healthcare market, most obstetricians now have demanding peripatetic work schedules. Private maternity patients are a lucrative source of income. The obstetrician is committed to attend these private births in person, and the "programming" (or scheduling) of births is a common time management strategy. The rate of elective caesarean sections was 30-68% in women with private obstetricians and 12-14% in women not attended by private obstetricians. CONCLUSIONS: Policies on healthcare financing can influence maternity care management and outcomes in unforeseen ways. The prevailing business ethos in health care encourages such pragmatism among those doctors who do not have a moral objection to non-medical caesarean section. PMID- 11118177 TI - Presence of relatives during testing for brain stem death: questionnaire study. PMID- 11118178 TI - Choosing a collaborator PMID- 11118174 TI - Reduction of postoperative mortality and morbidity with epidural or spinal anaesthesia: results from overview of randomised trials. AB - OBJECTIVES: To obtain reliable estimates of the effects of neuraxial blockade with epidural or spinal anaesthesia on postoperative morbidity and mortality. DESIGN: Systematic review of all trials with randomisation to intraoperative neuraxial blockade or not. STUDIES: 141 trials including 9559 patients for which data were available before 1 January 1997. Trials were eligible irrespective of their primary aims, concomitant use of general anaesthesia, publication status, or language. Trials were identified by extensive search methods, and substantial amounts of data were obtained or confirmed by correspondence with trialists. MAIN OUTCOME MEASURES: All cause mortality, deep vein thrombosis, pulmonary embolism, myocardial infarction, transfusion requirements, pneumonia, other infections, respiratory depression, and renal failure. RESULTS: Overall mortality was reduced by about a third in patients allocated to neuraxial blockade (103 deaths/4871 patients versus 144/4688 patients, odds ratio=0.70, 95% confidence interval 0.54 to 0.90, P=0. 006). Neuraxial blockade reduced the odds of deep vein thrombosis by 44%, pulmonary embolism by 55%, transfusion requirements by 50%, pneumonia by 39%, and respiratory depression by 59% (all P<0.001). There were also reductions in myocardial infarction and renal failure. Although there was limited power to assess subgroup effects, the proportional reductions in mortality did not clearly differ by surgical group, type of blockade (epidural or spinal), or in those trials in which neuraxial blockade was combined with general anaesthesia compared with trials in which neuraxial blockade was used alone. CONCLUSIONS: Neuraxial blockade reduces postoperative mortality and other serious complications. The size of some of these benefits remains uncertain, and further research is required to determine whether these effects are due solely to benefits of neuraxial blockade or partly to avoidance of general anaesthesia. Nevertheless, these findings support more widespread use of neuraxial blockade. PMID- 11118179 TI - Qualitative study of views of health professionals and patients on guided self management plans for asthma. AB - OBJECTIVES: To explore the views held by general practitioners, practice nurses, and patients about the role of guided self management plans in asthma care. DESIGN: Qualitative study using nine focus groups that each met on two occasions. SETTING: South Wales. SUBJECTS: 13 asthma nurses, 11 general practitioners (six with an interest in asthma), and 32 patients (13 adults compliant with treatment, 12 non-compliant adults, and seven teenagers). RESULTS: Neither health professionals nor patients were enthusiastic about guided self management plans, and, although for different reasons, almost all participants were ambivalent about their usefulness or relevance. Most professionals opposed their use. Few patients reported sustained use, and most felt that plans were largely irrelevant to them. The attitudes associated with these views reflect the gulf between the professionals' concept of the "responsible asthma patient" and the patients' view. CONCLUSIONS: Attempts to introduce self guided management plans in primary care are unlikely to be successful. A more patient centred, patient negotiated plan is needed for asthma care in the community. PMID- 11118180 TI - Always double check PMID- 11118181 TI - Quality of web based information on treatment of depression: cross sectional survey. AB - OBJECTIVES: To evaluate quality of web based information on treatment of depression, to identify potential indicators of content quality, and to establish if accountability criteria are indicators of quality. DESIGN: Cross sectional survey. DATA SOURCES: 21 frequently accessed websites about depression. MAIN OUTCOME MEASURES: (i) Site characteristics; (ii) quality of content-concordance with evidence based depression guidelines (guideline score), appropriateness of other relevant site information (issues score), and subjective rating of site quality (global score); and (iii) accountability-conformity with core accountability standards (Silberg score) and quality of evidence cited in support of conclusions (level of evidence score). RESULTS: Although the sites contained useful information, their overall quality was poor: the mean guideline, issues, and global scores were only 4.7 (range 0-13) out of 43, 9.8 (6-14) out of 17, and 3 (0.5-7. 5) out of 10 respectively. Sites typically did not cite scientific evidence in support of their conclusions. The guideline score correlated with the two other quality of content measures, but none of the content measures correlated with the Silberg accountability score. Content quality was superior for sites owned by organisations and sites with an editorial board. CONCLUSIONS: There is a need for better evidence based information about depression on the web, and a need to reconsider the role of accountability criteria as indicators of site quality and to develop simple valid indicators of quality. Ownership by an organisation and the involvement of a professional editorial board may be useful indicators. The study methodology may be useful for exploring these issues in other health related subjects. PMID- 11118182 TI - "Extracts from "Clinical evidence": Menopausal symptoms. AB - DEFINITION: Menopause begins one year after the last menstrual period. Symptoms often begin in the perimenopausal years. INCIDENCE/PREVALENCE: In the United Kingdom the mean age for the menopause is 50 years 9 months. The median onset of the perimenopause is between 45.5 and 47.5 years. One Scottish survey (of 6096 women aged 45 to 54 years) found that 84% had experienced at least one of the classic menopausal symptoms, with 45% finding one or more symptoms a problem. AETIOLOGY/RISK FACTORS: Urogenital symptoms of menopause are caused by decreased oestrogen concentrations, but the cause of vasomotor symptoms and psychological effects is complex and remains unclear. PROGNOSIS: Menopause is a physiological event. Its timing may be genetically determined. Although endocrine changes are permanent, menopausal symptoms such as hot flushes, which are experienced by about 70% of women, usually resolve with time. However, some symptoms, such as genital atrophy, may remain the same or worsen. AIMS: To reduce or prevent menopausal symptoms, and to improve quality of life with minimum adverse effects. OUTCOMES: Frequency and severity of vasomotor, urogenital, and psychological symptoms; quality of life. METHODS: Clinical Evidence search and appraisal December 1999. We included only randomised controlled trials (RCTs) and systematic reviews that met Clinical Evidence quality criteria. PMID- 11118183 TI - For and against: doctors should advise adolescents to abstain from sex. PMID- 11118184 TI - An integrated national pharmaceutical policy for the United Kingdom? PMID- 11118185 TI - Obituaries PMID- 11118186 TI - Referral and diagnostic process in suspected colorectal cancer needs to be improved to achieve two week target. PMID- 11118187 TI - Effect of screening programme on mortality from breast cancer. Benefit of 30% may be substantial overestimate. PMID- 11118188 TI - Effect of screening programme on mortality from breast cancer. Women might not accept mammography if benefit is lower than is currently thought. PMID- 11118189 TI - Effect of screening programme on mortality from breast cancer. Investment in treatment would be more cost effective. PMID- 11118190 TI - Giving medicine a fair trial. Patients' altruism should be appreciated. PMID- 11118191 TI - Giving medicine a fair trial. Patients' perspective must be acknowledged. PMID- 11118192 TI - Giving medicine a fair trial. Blanket enthusiasm for trials won't help. PMID- 11118193 TI - Giving medicine a fair trial. It needs to be established whether patients really fare better in trials. PMID- 11118194 TI - Increased high risk sexual behaviour in homosexual men. There is no evidence for a decreased incidence of HIV infection. PMID- 11118195 TI - Increased high risk sexual behaviour in homosexual men. Findings are similar in Manchester. PMID- 11118196 TI - Increased high risk sexual behaviour in homosexual men. Clarity may have been lost through including too much information. PMID- 11118197 TI - Income inequality and mortality in Canada and the United States. Third explanation is plausible. PMID- 11118198 TI - Evolutionary dynamics of full genome content in Escherichia coli. AB - The evolutionary history of the entire Escherichia coli chromosome was traced by examining the distribution of the approximately 4300 open reading frames (ORFs) from E.coli MG1655 among strains of known genealogical relationships. Using this framework to deduce the incidence of gene transfer and gene loss, a total of 67 events-37 additions and 30 deletions-were required to account for the distribution of all genes now present in the MG1655 chromosome. Nearly 90% of the ORFs were common to all strains examined, but, given the variation in gene content and chromosome size, strains can contain well over a megabase of unique DNA, conferring traits that distinguish them from other members of the species. Moreover, strains vary widely in their frequencies of deletions, which probably accounts for the variation in genome size within the species. PMID- 11118199 TI - The molecular basis of the specificity of action of K(ATP) channel openers. AB - K(ATP) channels incorporate a regulatory subunit of the ATP-binding cassette (ABC) transporter family, the sulfonylurea receptor (SUR), which defines their pharmacology. The therapeutically important K(+) channel openers (e.g. pinacidil, cromakalim, nicorandil) act specifically on the SUR2 muscle isoforms but, except for diazoxide, remain ineffective on the SUR1 neuronal/pancreatic isoform. This SUR1/2 dichotomy underpinned a chimeric strategy designed to identify the structural determinants of opener action, which led to a minimal set of two residues within the last transmembrane helix of SUR. Transfer of either residue from SUR2A to SUR1 conferred opener sensitivity to SUR1, while the reverse operation abolished SUR2A sensitivity. It is therefore likely that these residues form part of the site of interaction of openers with the channel. Thus, openers would target a region that, in other ABC transporters, is known to be tightly involved with the binding of substrates and other ligands. This first glimpse of the site of action of pharmacological openers should permit rapid progress towards understanding the structural determinants of their affinity and specificity. PMID- 11118200 TI - The structural basis of the catalytic mechanism and regulation of glucose-1 phosphate thymidylyltransferase (RmlA). AB - The synthesis of deoxy-thymidine di-phosphate (dTDP)-L-rhamnose, an important component of the cell wall of many microorganisms, is a target for therapeutic intervention. The first enzyme in the dTDP-L-rhamnose biosynthetic pathway is glucose-1-phosphate thymidylyltransferase (RmlA). RmlA is inhibited by dTDP-L rhamnose thereby regulating L-rhamnose production in bacteria. The structure of Pseudomonas aeruginosa RmlA has been solved to 1.66 A resolution. RmlA is a homotetramer, with the monomer consisting of three functional subdomains. The sugar binding and dimerization subdomains are unique to RmlA-like enzymes. The sequence of the core subdomain is found not only in sugar nucleotidyltransferases but also in other nucleotidyltransferases. The structures of five distinct enzyme substrate- product complexes reveal the enzyme mechanism that involves precise positioning of the nucleophile and activation of the electrophile. All the key residues are within the core subdomain, suggesting that the basic mechanism is found in many nucleotidyltransferases. The dTDP-L-rhamnose complex identifies how the protein is controlled by its natural inhibitor. This work provides a platform for the design of novel drugs against pathogenic bacteria. PMID- 11118201 TI - Inhibition of Bmp signaling affects growth and differentiation in the anagen hair follicle. AB - Growth and differentiation of postnatal hair follicles are controlled by reciprocal interactions between the dermal papilla and the surrounding epidermal hair precursors. The molecular nature of these interactions is largely unknown, but they are likely to involve several families of signaling molecules, including Fgfs, Wnts and Bmps. To analyze the function of Bmp signaling in postnatal hair development, we have generated transgenic mice expressing the Bmp inhibitor, Noggin, under the control of the proximal Msx2 promoter, which drives expression in proliferating hair matrix cells and differentiating hair precursor cells. Differentiation of the hair shaft but not the inner root sheath is severely impaired in Msx2-Noggin transgenic mice. In addition to hair keratins, the expression of several transcription factors implicated in hair development, including Foxn1 and Hoxc13, is severely reduced in the transgenic hair follicles. Proliferating cells, which are normally restricted to the hair matrix surrounding the dermal papilla, are found in the precortex and hair shaft region. These results identify Bmps as key regulators of the genetic program controlling hair shaft differentiation in postnatal hair follicles. PMID- 11118202 TI - Xrcc2 is required for genetic stability, embryonic neurogenesis and viability in mice. AB - Repair of DNA damage by homologous recombination has only recently been established as an important mechanism in maintaining genetic stability in mammalian cells. The recently cloned Xrcc2 gene is a member of the mammalian Rad51 gene family, thought to be central to homologous recombination repair. To understand its function in mammals, we have disrupted Xrcc2 in mice. No Xrcc2(-/ ) animals were found alive, with embryonic lethality occurring from mid gestation. Xrcc2(-/-) embryos surviving until later stages of embryogenesis commonly showed developmental abnormalities and died at birth. Neonatal lethality, apparently due to respiratory failure, was associated with a high frequency of apoptotic death of post- mitotic neurons in the developing brain, leading to abnormal cortical structure. Embryonic cells showed genetic instability, revealed by a high level of chromosomal aberrations, and were sensitive to gamma-rays. Our findings demonstrate that homologous recombination has an important role in endogenous damage repair in the developing embryo. Xrcc2 disruption identifies a range of defects that arise from malfunction of this repair pathway, and establishes a previously unidentified role for homologous recombination repair in correct neuronal development. PMID- 11118203 TI - Asymmetrically localized Bud8p and Bud9p proteins control yeast cell polarity and development. AB - Diploid strains of the budding yeast Saccharomyces cerevisiae change the pattern of cell division from bipolar to unipolar when switching growth from the unicellular yeast form (YF) to filamentous, pseudohyphal (PH) cells in response to nitrogen starvation. The functions of two transmembrane proteins, Bud8p and Bud9p, in regulating YF and PH cell polarity were investigated. Bud8p is highly concentrated at the distal pole of both YF and PH cells, where it directs initiation of cell division. Asymmetric localization of Bud8p is independent of the Rsr1p/Bud1p GTPase. rsr1/bud1 mutations are epistatic to bud8 mutations, placing Rsr1p/Bud1p downstream of Bud8p. In YF cells, Bud9p is also localized at the distal pole, yet deletion of BUD9 favours distal bud initiation. In PH cells, nutritional starvation for nitrogen efficiently prevents distal localization of Bud9p. Because Bud8p and Bud9p proteins associate in vivo, we propose Bud8p as a landmark for bud initiation at the distal cell pole, where Bud9p acts as inhibitor. In response to nitrogen starvation, asymmetric localization of Bud9p is averted, favouring Bud8p-mediated cell division at the distal pole. PMID- 11118204 TI - Expression of the endogenous type II secretion pathway in Escherichia coli leads to chitinase secretion. AB - Escherichia coli K-12, the most widely used laboratory bacterium, does not secrete proteins into the extracellular medium under standard growth conditions, despite possessing chromosomal genes encoding a putative type II secretion machinery (secreton). We show that in wild-type E.coli K-12, divergent transcription of the two operons in the main chromosomal gsp locus, encoding the majority of the secreton components, is silenced by the nucleoid-structuring protein H-NS. In mutants lacking H-NS, the secreton genes cloned on a moderate copy-number plasmid are expressed and promote efficient secretion of the endogenous, co-regulated endochitinase ChiA. This is the first time that secretion of an endogenous extracellular protein has been demonstrated in E.coli K-12. PMID- 11118205 TI - In vivo kinetics of protein targeting to the endoplasmic reticulum determined by site-specific phosphorylation. AB - We have developed a novel assay to detect the cytosolic localization of protein domains by inserting a short consensus sequence for phosphorylation by protein kinase A. In transfected COS-1 cells, this sequence was labeled efficiently with [(32)P]phosphate only when exposed to the cytosol and not when translocated into the lumen of the endoplasmic reticulum. The phosphorylation state of this sequence can therefore be used to determine the topology of membrane proteins. This assay is sufficiently sensitive to detect even the transient cytosolic exposure of the N-terminal domain of a membrane protein with a reverse signal anchor sequence. The extent of phosphorylation per newly synthesized polypeptide was shown to reflect the time of exposure to the cytosol, which depends on translation, targeting and translocation of the N-terminus. By altering the length of the N-terminal domain or manipulating the translation rate, it was determined that protein targeting is rapid and requires only a few seconds. The rate of N-terminal translocation was estimated to be approximately 1.6 times the rate of translation. PMID- 11118206 TI - Sequential action of two GTPases to promote vacuole docking and fusion. AB - Homotypic vacuole fusion occurs by sequential priming, docking and fusion reactions. Priming frees the HOPS complex (Vps 11, 16, 18, 33, 39 and 41) to activate Ypt7p for docking. Here we explore the roles of the GDP and GTP states of Ypt7p using Gdi1p (which extracts Ypt7:GDP), Gyp7p (a GTPase-activating protein for Ypt7p:GTP), GTPgammaS or GppNHp (non-hydrolyzable nucleotides), and mutant forms of Ypt7p that favor either GTP or GDP states. GDP-bound Ypt7p on isolated vacuoles can be extracted by Gdi1p, although only the GTP-bound state allows docking. Ypt7p is converted to the GTP-bound state after priming and stably associates with HOPS. Gyp7p can cause Ypt7p to hydrolyze bound GTP to GDP, driving HOPS release and accelerating Gdi1p-mediated release of Ypt7p. Ypt7p extraction does not inhibit the Ca(2+)-triggered cascade that leads to fusion. However, in the absence of Ypt7p, fusion is still sensitive to GTPgammaS and GppNHp, indicating that there is a second specific GTPase that regulates the calcium flux and hence fusion. Thus, two GTPases sequentially govern vacuole docking and fusion. PMID- 11118207 TI - SHPS-1 regulates integrin-mediated cytoskeletal reorganization and cell motility. AB - The transmembrane glycoprotein SHPS-1 binds the protein tyrosine phosphatase SHP 2 and serves as its substrate. Although SHPS-1 has been implicated in growth factor- and cell adhesion-induced signaling, its biological role has remained unknown. Fibroblasts homozygous for expression of an SHPS-1 mutant lacking most of the cytoplasmic region of this protein exhibited increased formation of actin stress fibers and focal adhesions. They spread more quickly on fibronectin than did wild-type cells, but they were defective in subsequent polarized extension and migration. The extent of adhesion-induced activation of Rho, but not that of Rac, was also markedly reduced in the mutant cells. Activation of the Ras extracellular signal-regulated kinase signaling pathway and of c-Jun N-terminal kinases by growth factors was either unaffected or enhanced in the mutant fibroblasts. These results demonstrate that SHPS-1 plays crucial roles in integrin-mediated cytoskeletal reorganization, cell motility and the regulation of Rho, and that it also negatively modulates growth factor-induced activation of mitogen-activated protein kinases. PMID- 11118208 TI - Membrane association induces a conformational change in the Ebola virus matrix protein. AB - The matrix protein VP40 from Ebola virus is targeted to the plasma membrane, where it is thought to induce assembly and budding of virions through its association with the lipid bilayer. Ebola virus VP40 is expressed as a monomeric molecule in solution, consisting of two loosely associated domains. Here we show that a C-terminal truncation of seven residues destabilizes the monomeric closed conformation and induces spontaneous hexamerization in solution, as indicated by chemical cross-linking and electron microscopy. Three-dimensional reconstruction of electron microscopy images shows ring-like structures consisting of the N terminal domain along with evidence for flexibly attached C-terminal domains. In vitro destabilization of the monomer by urea treatment results in similar hexameric molecules in solution. In addition, we demonstrate that membrane association of wild-type VP40 also induces the conformational switch from monomeric to hexameric molecules that may form the building blocks for initiation of virus assembly and budding. Such a conformational change induced by bilayer targeting may be a common feature of many viral matrix proteins and its potential inhibition may result in new anti-viral therapies. PMID- 11118209 TI - Epstein-Barr virus-encoded poly(A)(-) RNA supports Burkitt's lymphoma growth through interleukin-10 induction. AB - Akata and Mutu cell lines are derived from Burkitt's lymphoma (BL) and retain the in vivo phenotype of Epstein-Barr virus (EBV) expression that is characterized by expression of EBV-determined nuclear antigen 1 (EBNA1), EBV-encoded RNAs (EBERs) and transcripts from the BAM:HI A region (BARF0). We found that EBV-positive Akata and Mutu cell clones expressed higher levels of interleukin (IL)-10 than their EBV-negative subclones at the transcriptional level. Transfection of an individual EBV latent gene into EBV-negative Akata cells revealed that EBERs were responsible for IL-10 induction. Recombinant IL-10 enabled EBV-negative Akata cells to grow in low (0.1%) serum conditions. On the other hand, growth of EBV positive Akata cells was blocked by treatment either with an anti-IL-10 antibody or antisense oligonucleotide against IL-10. EBV-positive BL biopsies consistently expressed IL-10, but EBV-negative BL biopsies did not. These results suggest that IL-10 induced by EBERs acts as an autocrine growth factor for BL. EBERs, EBER1 and EBER2, are non-polyadenylated RNAs and are 166 and 172 nucleotides long, respectively. The present findings indicate that RNA molecules could regulate cell growth. PMID- 11118210 TI - Influenza A virus NS2 protein mediates vRNP nuclear export through NES independent interaction with hCRM1. AB - For nuclear export of proteins, the formation of a ternary export complex composed of the export substrate, a cellular export factor and Ran-GTP is crucial. CRM1 is a cellular export factor for proteins containing leucine-rich nuclear export signals (NESs). Although the NES sequence is crucial for nuclear export, its exact role in the formation of the ternary export complex is controversial. Here we demonstrate an interaction between human CRM1 (hCRM1) and influenza A virus NS2 protein, which contains an NES motif in its N-terminal region. Replacement of the hydrophobic amino acids in the NES motif did not abolish NS2's interaction with hCRM1. Using our recently established systems for the generation of influenza virus or virus-like particles from cloned cDNAs, we found that NS2 is essential for nuclear export of influenza virus ribonucleoprotein (RNP) complexes, and that alteration of the NS2-NES abrogated this event and influenza virus generation. These findings suggest that the NS2 NES is not crucial for the interaction of this protein with hCRM1, but is for the formation of the ternary export complex with Ran-GTP. PMID- 11118211 TI - A novel ability of Smad3 to regulate proteasomal degradation of a Cas family member HEF1. AB - Smad3 is a key signal transducer of transforming growth factor-ss (TGF-ss) and activin, and is known to be a DNA-binding transcriptional regulator. Here we report a novel property of Smad3 in regulating the proteasomal degradation of the human enhancer of filamentation 1 (HEF1), which is a member of the Cas family of cytoplasmic docking proteins. Our studies revealed that Smad3 interacts with HEF1 and triggers the proteasomal degradation of HEF1 in overexpression systems. In addition, TGF-ss stimulation induces rapid proteasomal degradation of endogenous HEF1 in different TGF-ss-responsive cell lines. Interestingly, the degradation of HEF1 protein in epithelial cells is followed closely by an increase in HEF1 mRNA, resulting in a time-dependent increase in HEF1 protein level in TGF-ss-treated cells. Furthermore, we observed that an elevated HEF1 protein level inhibits TGF ss-induced Smad3-mediated gene responses. These data provide the first evidence for a novel cytoplasmic activity of Smad3 in regulating proteasomal degradation of HEF1 and also suggest a role for HEF1 in a negative feedback mechanism of the TGF-ss signaling pathway. PMID- 11118212 TI - Stress induces peroxisome biogenesis genes. AB - Peroxisomes are the cellular location of many antioxidants and are themselves significant producers of reactive oxygen species. In this report we demonstrate the induction of peroxisome biogenesis genes in both plant and animal cells by the universal stress signal molecule hydrogen peroxide. Using PEX1-LUC transgenic plants, rapid local and systemic induction of PEX1-luciferase could be demonstrated in vivo in response to physiological levels of hydrogen peroxide. PEX1-luciferase was also induced in response to wounding and to infection with an avirulent pathogen. We propose a model in which various stress situations that lead to the production of hydrogen peroxide can be ameliorated by elaboration of the peroxisome compartment to assist in restoration of the cellular redox balance. PMID- 11118213 TI - TAZ: a novel transcriptional co-activator regulated by interactions with 14-3-3 and PDZ domain proteins. AB - The highly conserved and ubiquitously expressed 14-3-3 proteins regulate differentiation, cell cycle progression and apoptosis by binding intracellular phosphoproteins involved in signal transduction. By screening in vitro translated cDNA pools for the ability to bind 14-3-3, we identified a novel transcriptional co-activator, TAZ (transcriptional co-activator with PDZ-binding motif) as a 14-3 3-binding molecule. TAZ shares homology with Yes-associated protein (YAP), contains a WW domain and functions as a transcriptional co-activator by binding to the PPXY motif present on transcription factors. 14-3-3 binding requires TAZ phosphorylation on a single serine residue, resulting in the inhibition of TAZ transcriptional co-activation through 14-3-3-mediated nuclear export. The C terminus of TAZ contains a highly conserved PDZ-binding motif that localizes TAZ into discrete nuclear foci and is essential for TAZ-stimulated gene transcription. TAZ uses this same motif to bind the PDZ domain-containing protein NHERF-2, a molecule that tethers plasma membrane ion channels and receptors to cytoskeletal actin. TAZ may link events at the plasma membrane and cytoskeleton to nuclear transcription in a manner that can be regulated by 14-3-3. PMID- 11118215 TI - Chromatin-mediated transcriptional regulation by the yeast architectural factors NHP6A and NHP6B. AB - The Saccharomyces cerevisiae NHP6A and NHP6B proteins are chromatin architectural factors, functionally and structurally related to the mammalian high mobility group (HMG)-1 and -2 proteins, a family of non-sequence-specific DNA binding proteins. nhp6a nhp6b mutants have various morphological defects and are defective in the induced expression of several RNA polymerase II-transcribed genes. We found that NHP6A/B proteins are also required for full induction of the yeast CHA1 gene. Importantly, CHA1 basal level expression is increased 10-fold in an nhp6a nhp6b double deletion mutant. Micrococcal nuclease and DNase I analysis of the CHA1 gene in this strain showed an open promoter structure, characteristic of the activated state of this promoter, even under non-inducing conditions. To address the possible function of the NHP6A/B proteins in chromatin-mediated gene regulation, we performed whole-genome transcriptional profiling of a Deltanhp6a Deltanhp6b yeast strain. Our results suggest that NHP6A/B proteins play an important regulatory role, repressing as well as potentiating expression of genes involved in several cellular processes, and that NHP6A/B control is exerted at the level of the individual gene. PMID- 11118214 TI - P/CAF-mediated acetylation regulates the function of the basic helix-loop-helix transcription factor TAL1/SCL. AB - The basic helix-loop-helix transcription factor TAL1 (or SCL) is a critical regulator of hematopoietic and vascular development and is misexpressed in the majority of patients with T-cell acute lymphoblastic leukemia. We found previously that TAL1 could interact with transcriptional co-activator and co repressor complexes possessing histone acetyltransferase and deacetylase activities, respectively. Here, we report that TAL1 is subject to acetylation in vivo and can be acetylated by p300 and the p300/CBP-associated factor P/CAF in vitro. P/CAF-mediated acetylation, which mapped to a lysine-rich motif in the loop region, increased TAL1 binding to DNA while selectively inhibiting its interaction with the transcriptional co-repressor mSin3A. Furthermore, P/CAF protein, TAL1-P/CAF interaction and TAL1 acetylation increased significantly in murine erythroleukemia cells induced to differentiate in culture, while enforced expression of an acetylation-defective P/CAF mutant inhibited endogenous TAL1 acetylation, TAL1 DNA-binding activity, TAL1-directed transcription and terminal differentiation of these cells. These results reveal a novel mechanism by which TAL1 activity is regulated and implicate acetylation of this transcription factor in promotion of erythroid differentiation. PMID- 11118216 TI - Patterns of histone acetylation suggest dual pathways for gene activation by a bifunctional locus control region. AB - The five genes of the human growth hormone (hGH) cluster are expressed in either the pituitary or placenta. Activation of the cluster is dependent on a locus control region (LCR) comprising pituitary- specific (HSI,II, -15 kb), placenta specific (HSIV, -30 kb) and shared (HSIII, -28 kb; HSV, -32 kb) DNase I hypersensitive sites. Gene activation in the pituitary is paralleled by acetylation of a 32 kb chromatin domain 5' to the cluster centered at HSI,II. In the present study we observed that acetylation of this region in placental chromatin was discretely limited to shared HSIII and HSV. Transgenic studies revealed placenta-specific activation of linked genes by a determinant (P element) located 2 kb 5' to each of the four placentally expressed genes. A localized peak of histone acetylation was observed at these P-elements in placenta but not pituitary. These data support a model for bifunctional action of the hGH LCR in which separate positive determinants, HSI,II and the P-elements, activate their respective target genes by tissue-specific recruitment of distinctly regulated histone acetyl transferase activities. PMID- 11118217 TI - Nuclear factor 1 (NF1) affects accurate termination and multiple-round transcription by human RNA polymerase III. AB - We have shown previously that the TFIIIC1/TFIIIC1' fraction interacts specifically with the VA1 terminator regions to affect both termination and initiation/reinitiation of transcription by human RNA polymerase III. Here, we further purified the VA1 terminator-binding factor to apparent homogeneity and found, by peptide sequence analysis, that it belongs to the NF1 protein family. NF1 interacts specifically with the NF1-binding sites within the terminator regions of the VA1 gene and with two subunits (TFIIIC220 and TFIIIC110) of human TFIIIC2. Immunodepletion with anti-NF1 antibodies dramatically decreases transcription from the VA1 template in nuclear extract, and mutation at the NF1 binding site in the terminator region of the VA1 gene selectively affects multiple-round transcription (reinitiation of transcription) and termination. In addition, NF1 acts in conjunction with TFIIIC to promote accurate termination by RNA polymerase III on a C-tailed VA1 template. PMID- 11118218 TI - Escherichia coli RNA polymerase core and holoenzyme structures. AB - Multisubunit RNA polymerase is an essential enzyme for regulated gene expression. Here we report two Escherichia coli RNA polymerase structures: an 11.0 A structure of the core RNA polymerase and a 9.5 A structure of the sigma(70) holoenzyme. Both structures were obtained by cryo-electron microscopy and angular reconstitution. Core RNA polymerase exists in an open conformation. Extensive conformational changes occur between the core and the holoenzyme forms of the RNA polymerase, which are largely associated with movements in ss'. All common RNA polymerase subunits (alpha(2), ss, ss') could be localized in both structures, thus suggesting the position of sigma(70) in the holoenzyme. PMID- 11118219 TI - Srb7p is a physical and physiological target of Tup1p. AB - The holoenzyme of transcription integrates the positive and negative signals from the promoters of eukaryotic genes. We demonstrate that the essential holoenzyme component Srb7p is a physiologically relevant target of the global repressor Tup1p in Saccharomyces cerevisiae. Tup1p binds Srb7p in vivo and in vitro, and all genes tested that are repressed by Tup1p are derepressed when wild-type Srb7p is replaced by a mutant derivative of Srb7p that is no longer capable of interacting with Tup1p. Therefore, Srb7p is the first holoenzyme component essential for repression by Tup1p for which a physical interaction with Tup1p has been demonstrated. Furthermore, we find that Srb7p also binds Med6p and that this interaction is necessary for full transcriptional activation by different activators. Our finding that Med6p and Tup1p compete for the interaction with Srb7p suggests a model for Tup1p-mediated repression. PMID- 11118220 TI - GreA and GreB proteins revive backtracked RNA polymerase in vivo by promoting transcript trimming. AB - The GreA and GreB proteins of Escherichia coli show a multitude of effects on transcription elongation in vitro, yet their physiological functions are poorly understood. Here, we investigated whether and how these factors influence lateral oscillations of RNA polymerase (RNAP) in vivo, observed at a protein readblock. When RNAP is stalled within an (ATC/TAG)(n) sequence, it appears to oscillate between an upstream and a downstream position on the template, 3 bp apart, with concomitant trimming of the transcript 3' terminus and its re-synthesis. Using a set of mutant E.coli strains, we show that the presence of GreA or GreB in the cell is essential to induce this trimming. We show further that in contrast to a ternary complex that is stabilized at the downstream position, the oscillating complex relies heavily on the GreA/GreB-induced 'cleavage-and-restart' process to become catalytically competent. Clearly, by promoting transcript shortening and re-alignment of the catalytic register, the Gre factors function in vivo to rescue RNAP from being arrested at template positions where the lateral stability of the ternary complex is impaired. PMID- 11118221 TI - The spliceosome deposits multiple proteins 20-24 nucleotides upstream of mRNA exon-exon junctions. AB - Eukaryotic mRNAs exist in vivo as ribonucleoprotein particles (mRNPs). The protein components of mRNPs have important functions in mRNA metabolism, including effects on subcellular localization, translational efficiency and mRNA half-life. There is accumulating evidence that pre-mRNA splicing can alter mRNP structure and thereby affect downstream mRNA metabolism. Here, we report that the spliceosome stably deposits several proteins on mRNAs, probably as a single complex of approximately 335 kDa. This complex protects 8 nucleotides of mRNA from complete RNase digestion at a conserved position 20-24 nucleotides upstream of exon-exon junctions. Splicing-dependent RNase protection of this region was observed in both HeLa cell nuclear extracts and Xenopus laevis oocyte nuclei. Immunoprecipitations revealed that five components of the complex are the splicing-associated factors SRm160, DEK and RNPS1, the mRNA-associated shuttling protein Y14 and the mRNA export factor REF. Possible functions for this complex in nucleocytoplasmic transport of spliced mRNA, as well as the nonsense-mediated mRNA decay pathway, are discussed. PMID- 11118223 TI - SECIS-SBP2 interactions dictate selenocysteine incorporation efficiency and selenoprotein hierarchy. AB - Selenocysteine incorporation at UGA codons requires cis-acting mRNA secondary structures and several specialized trans-acting factors. The latter include a selenocysteine-specific tRNA, an elongation factor specific for this tRNA and a SECIS-binding protein, SBP2, which recruits the elongation factor to the selenoprotein mRNA. Overexpression of selenoprotein mRNAs in transfected cells results in inefficient selenocysteine incorporation due to limitation of one or more of these factors. Using a transfection-based competition assay employing overexpression of selenoprotein mRNAs to compete for selenoprotein synthesis, we investigated the ability of the trans-acting factors to overcome competition and restore selenocysteine incorporation. We report that co-expression of SBP2 overcomes the limitation produced by selenoprotein mRNA overexpression, whereas selenocysteyl-tRNA and the selenocysteine-specific elongation factor do not. Competition studies indicate that once bound to SECIS elements, SBP2 does not readily exchange between them. Finally, we show that SBP2 preferentially stimulates incorporation directed by the seleno protein P and phospholipid hydroperoxide glutathione peroxidase SECIS elements over those of other selenoproteins. The mechanistic implications of these findings for the hierarchy of selenoprotein synthesis and nonsense-mediated decay are discussed. PMID- 11118222 TI - Molecular basis of sequence-specific recognition of pre-ribosomal RNA by nucleolin. AB - The structure of the 28 kDa complex of the first two RNA binding domains (RBDs) of nucleolin (RBD12) with an RNA stem-loop that includes the nucleolin recognition element UCCCGA in the loop was determined by NMR spectroscopy. The structure of nucleolin RBD12 with the nucleolin recognition element (NRE) reveals that the two RBDs bind on opposite sides of the RNA loop, forming a molecular clamp that brings the 5' and 3' ends of the recognition sequence close together and stabilizing the stem-loop. The specific interactions observed in the structure explain the sequence specificity for the NRE sequence. Binding studies of mutant proteins and analysis of conserved residues support the proposed interactions. The mode of interaction of the protein with the RNA and the location of the putative NRE sites suggest that nucleolin may function as an RNA chaperone to prevent improper folding of the nascent pre-rRNA. PMID- 11118224 TI - A new pathway of translational regulation mediated by eukaryotic initiation factor 3. AB - We report a new pathway of translation regulation that may operate in interferon treated or virus-infected mammalian cells. This pathway is activated by P56, a protein whose synthesis is strongly induced by interferons or double-stranded RNA. Using a yeast two-hybrid screen, we identified the P48 subunit of the mammalian translation initiation factor eIF-3 as a protein that interacts with P56. The P56-P48 interaction was confirmed in human cells by co immunoprecipitation assays and confocal microscopy. Gel filtration assays revealed that P56 binds to the large eIF-3 complex that contains P48. Purified recombinant P56 inhibited in vitro translation of reporter mRNAs in a dose dependent fashion, and that inhibition was reversed by the addition of purified eIF-3. In vivo, expression of transfected P56 or induction of the endogenous P56 by interferon caused an inhibition of overall cellular protein synthesis and the synthesis of a transfected reporter protein. As expected, a P56 mutant that does not interact with P48 and eIF-3 failed to inhibit protein synthesis in vitro and in vivo. PMID- 11118225 TI - A post-translational modification in the GGQ motif of RF2 from Escherichia coli stimulates termination of translation. AB - A post-translational modification affecting the translation termination rate was identified in the universally conserved GGQ sequence of release factor 2 (RF2) from Escherichia coli, which is thought to mimic the CCA end of the tRNA molecule. It was shown by mass spectrometry and Edman degradation that glutamine in position 252 is N:(5)-methylated. Overexpression of RF2 yields protein lacking the methylation. RF2 from E.coli K12 is unique in having Thr246 near the GGQ motif, where all other sequenced bacterial class 1 RFs have alanine or serine. Sequencing the prfB gene from E.coli B and MRE600 strains showed that residue 246 is coded as alanine, in contrast to K12 RF2. Thr246 decreases RF2-dependent termination efficiency compared with Ala246, especially for short peptidyl-tRNAs. Methylation of Gln252 increases the termination efficiency of RF2, irrespective of the identity of the amino acid in position 246. We propose that the previously observed lethal effect of overproducing E.coli K12 RF2 arises through accumulating the defects due to lack of Gln252 methylation and Thr246 in place of alanine. PMID- 11118226 TI - A recurrent general RNA binding domain appended to plant methionyl-tRNA synthetase acts as a cis-acting cofactor for aminoacylation. AB - The cDNA encoding rice methionyl-tRNA synthetase was isolated. The protein exhibited a C-terminal polypeptide appended to a classical MetRS domain. This supplementary domain is related to endothelial monocyte activating polypeptide II (EMAPII), a cytokine produced in mammals after cleavage of p43, a component of the multisynthetase complex. It is also related to Arc1p and Trbp111, two tRNA binding proteins. We expressed rice MetRS and a derivative with a deletion of its EMAPII-like domain. Band-shift analysis showed that this extra-domain provides MetRS with non-specific tRNA binding properties. The EMAPII-like domain contributed a 10-fold decrease in K:(M) for tRNA in the aminoacylation reaction catalyzed by the native enzyme, as compared with the C-terminally truncated MetRS. Consequently, the EMAPII domain provides MetRS with a better catalytic efficiency at the free tRNA concentration prevailing in vivo. This domain binds the acceptor minihelix of tRNA(Met) and facilitates its aminoacylation. These results suggest that the EMAPII module could be a relic of an ancient tRNA binding domain that was incorporated into primordial synthetases for aminoacylation of RNA minihelices taken as the ancestor of modern tRNA. PMID- 11118227 TI - DNA of Drosophila melanogaster contains 5-methylcytosine. AB - It is commonly accepted that the DNA of Drosophila melanogaster does not contain 5-methylcytosine, which is essential in the development of most eukaryotes. We have developed a new, highly specific and sensitive assay to detect the presence of 5-methylcytosine in genomic DNA. The DNA is degraded to nucleosides, 5 methylcytosine purified by HPLC and, for detection by 1D- and 2D-TLC, radiolabeled using deoxynucleoside kinase and [gamma-(32)P]ATP. Using this assay, we show here that 5-methylcytosine occurs in the DNA of D. melanogaster at a level of approximately 1 in 1000-2000 cytosine residues in adult flies. DNA methylation is detectable in all stages of D.melanogaster development. PMID- 11118229 TI - Remembering dr. Robert dorwart PMID- 11118228 TI - In memoriam PMID- 11118230 TI - Bob Dorwart's legacy. PMID- 11118231 TI - Some distinctive features of the impact of managed care on psychiatry. AB - Past research suggests that the spread of managed care is affecting the treatment of mental and physical illnesses differently. This article develops six hypotheses that could explain the differential effects of managed behavioral health care, based on characteristics of mental disorders, professional norms of treatment, and the broader societal consequences of untreated mental illness. Using data from the 1998 Socioeconomic Monitoring System fielded by the American Medical Association, we tested these hypotheses by comparing the experiences of psychiatrists under managed care with those of primary care providers and medical specialists. We found the following: (1) psychiatrists face substantially more aggressive external review than do primary care providers and are less successful in overturning denials; (2) psychiatrists feel significantly more at risk for disaffiliation from health plans; (3) psychiatrists report facing review protocols that are more confusing than those for primary care physicians, but psychiatrists' staff spend less time on external review; (4) psychiatrists are more likely than other physicians to report that their patients have difficulty making informed choices about managed care; and (5) psychiatrists evidence greater time commitment to advocacy on behalf of their patients with respect to managed care. PMID- 11118232 TI - Medicaid behavioral health carve-outs: a new generation of privatization decisions. AB - This article addresses issues related to the privatization of various functions within the mental health system. It acknowledges the contributions of Robert Dorwart, who explored trends with regard to the privatization of inpatient psychiatric services. The authors then highlight changes in the division of labor between the public and private sectors regarding the financing and delivery of mental health services and the management of the system. Responsibility for funding the mental health system has remained largely a public responsibility while responsibility for production or delivery of services in the mental health system is typically held by private, for-profit, and not-for-profit organizations. The roles of managing the mental health system and setting policy are now shared between the private and public sectors in a number of states that have implemented Medicaid behavioral health carve-out programs. This article explores the impact of such privatization on cost, access, and quality of services by examining the experiences of three states with carve-outs. The authors suggest that while organizational form is an important issue, concerns about privatization should be tempered by attention to the contracting decisions made by purchasers, the level of resources devoted to services, and the adequacy of administration of the system. PMID- 11118233 TI - Review of programs for persons who are homeless and mentally ill. AB - Despite recent prosperity in the U.S., homelessness is still a widespread social problem. It is estimated that 25% of homeless persons have a serious mental illness. This article will review the literature evaluating prevention services and specialized outreach, treatment, and housing programs designed to reduce homelessness for individuals who are mentally ill. Although these interventions have been helpful in addressing the complex needs of the homeless mentally ill, it is difficult to measure how they have improved outcomes. It is even more challenging to determine whether the programs are cost-effective. Since public resources are used to maintain services for the homeless mentally ill, policy makers must be informed about whether the best outcomes are achieved at the lowest possible cost. Following a discussion of the successes of the individual programs and the challenges they confront, several important questions are identified related to improving the efficiency of these programs. Although the establishment of such programs indicates that progress has been made toward alleviating the burdens facing people who are homeless and mentally ill, collaboration among all stakeholders-especially between the mental health community and consumer advocates-needs to be further enhanced. New research can be conducted in a way that improves how information is evaluated and used. PMID- 11118234 TI - Developing a quality management system for behavioral health care: the Cambridge Health Alliance Experience. AB - The rapid pace of change in the health care system presents tremendous challenges for clinicians and managers charged with the delivery of mental health and substance abuse services. Declining reimbursement, new incentive structures, and increasing competition are placing unprecedented pressure on providers to deliver care efficiently. Regulatory scrutiny, consumer dissatisfaction, and a growing awareness of gaps between actual and ideal practice have led to intensifying pressure to improve quality. Yet system change has also presented new opportunities for managing cost and quality of care. Consolidation of facilities and practices into integrated networks, developments in information systems technology, and the emergence of models to facilitate change have led to the rise of "quality management," a framework for assessing and improving clinical, operational, and financial performance within a health care organization. This article reviews some of the precipitating factors and theoretical structures underlying quality management and then, through a case study of one organization's experience, describes the implementation of a quality management program in a behavioral health care delivery system. The case study emphasizes how theoretical frameworks were operationalized and how organizational structure and process were shaped to address challenges well known in quality management, such as authority, accountability, and follow-through. A multiphase model of quality management program development is formulated and used to provide context for this program's development. PMID- 11118235 TI - Seclusion and restraint: a review of recent literature. AB - An October 1998 Hartford Courant investigative series highlighted alleged cases of brutality and death suffered by involuntarily secluded, restrained, and/or emergently medicated patients. The resulting public and professional furor prompted a spate of new federal regulations and legislative initiatives setting national standards for reporting and clinical oversight. These events provide stimulus for this literature review. Rates, duration, and methods of seclusion and restraint still vary widely. Little evidence is available to guide clinical practice regarding relative benefits and risks of various methods to control acute adult patient aggression; even less evidence exists in child and adolescent populations. Further efficacy and effectiveness studies are needed to address this issue. Various programmatic efforts successfully reduce seclusion and restraint-at times dramatically-and can be used as examples of systematic quality improvement so "best practices" may evolve and spread throughout psychiatric inpatient settings. PMID- 11118236 TI - Synapses, sea slugs, and psychiatry. PMID- 11118237 TI - Prevalence of cerebral white matter lesions in elderly people: a population based magnetic resonance imaging study: the Rotterdam Scan Study. PMID- 11118238 TI - Vascular depression: new light on an established idea? PMID- 11118239 TI - Neurology of ciguatera. AB - Ciguatera is a widespread ichthyosarcotoxaemia with dramatic and clinically important neurological features. This severe form of fish poisoning may present with either acute or chronic intoxication syndromes and constitutes a global health problem. Ciguatera poisoning is little known in temperate countries as a potentially global problem associated with human ingestion of large carnivorous fish that harbour the bioaccumulated ciguatoxins of the photosynthetic dinoflagellate Gambierdiscus toxicus. This neurotoxin is stored in the viscera of fish that have eaten the dinoflagellate and concentrated it upwards throughout the food chain towards progressively larger species, including humans. Ciguatoxin accumulates in all fish tissues, especially the liver and viscera, of "at risk" species. Both Pacific (P-CTX-1) and Caribbean (C-CTX-1) ciguatoxins are heat stable polyether toxins and pose a health risk at concentrations above 0.1 ppb. The presenting signs of ciguatera are primarily neurotoxic in more than 80% of cases. Such include the pathognomonic features of postingestion paraesthesiae, dysaesthesiae, and heightened nociperception. Other sensory abnormalities include the subjective features of metallic taste, pruritus, arthralgia, myalgia, and dental pain. Cerebellar dysfunction, sometimes diphasic, and weakness due to both neuropathy and polymyositis may be encountered. Autonomic dysfunction leads to hypotension, bradycardia, and hypersalivation in severe cases. Ciguatoxins are potent, lipophilic sodium channel activator toxins which bind to the voltage sensitive (site 5) sodium channel on the cell membranes of all excitable tissues. Treatment depends on early diagnosis and the early administration of intravenous mannitol. The early identification of the neurological features in sentinel patients has the potential to reduce the number of secondary cases in cluster outbreaks. PMID- 11118240 TI - Prevalence of cerebral white matter lesions in elderly people: a population based magnetic resonance imaging study. The Rotterdam Scan Study. AB - OBJECTIVE: White matter lesions are often seen on MR scans of elderly non demented and demented people. They are attributed to degenerative changes of small vessels and are implicated in the pathogenesis of cognitive decline and dementia. There is evidence that especially periventricular white matter lesions are related to cognitive decline, whereas subcortical white matter lesions may be related to late onset depression. The frequency distribution of subcortical and periventricular white matter lesions according to age and sex reported. METHODS: A total of 1077 subjects aged between 60-90 years were randomly sampled from the general population. All subjects underwent 1.5T MR scanning; white matter lesions were rated separately for the subcortical region and the periventricular region. RESULTS: Of all subjects 8% were completely free of subcortical white matter lesions, 20% had no periventricular white matter lesions, and 5% had no white matter lesions in either of these locations. The proportion with white matter lesions increased with age, similarly for men and women. Women tended to have more subcortical white matter lesions than men (total volume 1.45 ml v 1. 29 ml; p=0.33), mainly caused by marked differences in the frontal white matter lesion volume (0.89 ml v 0.70 ml; p=0.08). Periventricular white matter lesions were also more frequent among women than men (mean grade 2.5 v 2.3; p=0.07). Also severe degrees of subcortical white matter lesions were more common in women than in men (OR 1.1; 95% confidence interval (95% CI) 0.8-1.5) and periventricular white matter lesions (OR 1.2; 95% CI 0.9-1.7), albeit that none of these findings were statistically significant. CONCLUSIONS: The prevalence and the degree of cerebral white matter lesions increased with age. Women tended to have a higher degree of white matter lesions than men. This may underlie the finding of a higher incidence of dementia in women than in men, particularly at later age. PMID- 11118241 TI - Saint vitus and his dance. PMID- 11118242 TI - Additional educational needs in children born to mothers with epilepsy. AB - OBJECTIVES: To examine the relative risks of additional educational needs (AENs) in children exposed to antiepileptic drug (AED) monotherapy and polytherapy regimes in utero. METHODS: A retrospective survey of women between the ages of 16 to 40 registered at the Mersey Regional Epilepsy Clinic, who received a postal questionnaire concerning their experience of pregnancy and the subsequent schooling of live-born children. RESULTS: 721 (57%) women of the 1267 approached returned an adequately completed questionnaire; 330 (46%) had given birth to at least one live-born child. Information was collected on 594 children, 400 of whom were of school age (4-18). 150 (37.5%) had been exposed to monotherapy in utero, 74 (18.5%) were exposed to polytherapy, and 176 were not exposed to any AEDs. The odds ratio of AENs for all children exposed to AEDs in utero compared with those unexposed was 1.49 (95% confidence interval (95% CI) 0.83 -2.67). Odds ratios for AENs for each therapy subgroup compared with those unexposed were also calculated for all children. Those exposed to valproate monotherapy had an odds ratio of 3.4 (95% CI 1.63-7.10) by contrast with an odds ratio of 0.26 (95% CI 0.06- 1.15) for carbamazepine. Polytherapy including valproate had similarly high odds ratios for AENs compared with those unexposed of 2.51 ( 95% CI 1.04-6.07) versus the odds ratio of 1.51 ( 95% CI 0.56-4.07) for polytherapy excluding valproate. CONCLUSIONS: Although the findings should be treated with caution, they suggest that monotherapy or polytherapy with valproate during pregnancy carries particular risks for the development of children exposed in utero. PMID- 11118243 TI - Brain perfusion abnormalities in Alzheimer's disease: comparison between patients with focal temporal lobe dysfunction and patients with diffuse cognitive impairment. AB - OBJECTIVES: Patients with Alzheimer's disease (AD) showing a selective impairment of episodic and semantic memory have recently been classified as affected by focal temporal lobe dysfunction (FTLD) and considered as a distinct subgroup of patients affected by a particular form of AD. The aim was to compare the cerebral perfusion of patients with AD with FTLD and patients with AD with the more typical profile of diffuse cognitive impairment (dAD). METHODS: Ten patients with AD with FTLD, 14 patients with AD with dAD, and 12 normal controls were studied. All the 24 patients with AD underwent a complete neuropsychological assessment. SPECT examination with [(99m)Tc]-HMPAO, using a four head brain dedicated tomograph, was performed in patients and controls. Tracer uptake was quantified in 27 regions of interest (ROIs), including lateral and mesial temporal areas. Mean counts in the 27 ROIs of controls, patients with FTLD and those with dAD were compared using an ANOVA for repeated measures with Bonferroni's correction. A logistic regression analysis, followed by a receiver operating characteristic (ROC) analysis, was also applied to select SPECT patterns which significantly differentiated patients with FTLD and those with dAD. RESULTS: Two scintigraphic patterns of abnormalities, shaping a double dissociation between the FTLD and dAD groups, emerged: a bilateral mesial temporal hypoperfusion, characteristic of FTLD and a posterior parietal (and temporal parietal) hypoperfusion characteristic of patients with dAD. CONCLUSIONS: These scintigraphic findings provide further support to the hypothesis that FTLD is not a mere stage but a distinct anatomoclinical form of AD. The combination of neuropsychological tests and [(99m)Tc]-HMPAO SPECT may be very useful in identifying patients with FTLD from the wider group of patients with dAD. This issue is particularly worthwhile, as there is increasing evidence that patients with FTLD have a slower rate of cognitive decline. PMID- 11118244 TI - Variation of visual evoked potential delay to stimulation of central, nasal, and temporal regions of the macula in optic neuritis. AB - OBJECTIVES: To compare the degree of visual evoked potential (VEP) delay to stimulation of central, nasal, and temporal regions of the macula in optic neuritis, to determine whether the differential involvement of parvocellular and magnocellular fibre types suggested by other studies is governed by retinotopic factors. METHODS: VEPs were recorded to reversal of 40' checks in the central (4 degrees radius) and the left and right surrounding regions of the visual field (as far as 10 degrees vertical and 14 degrees horizontal) in 30 patients recently recovered from the acute stage of optic neuritis, and in 17 age matched controls. RESULTS: In the control group, VEP latencies were similar to stimulation of the central and temporal regions of the macula, marginally shorter from the nasal region. In the patients with optic neuritis, VEPs were significantly more delayed from the central region, on average by about twice as much as from the nasal and temporal regions. Delays seen in some of the VEPs from the patients' fellow eyes tended to be more uniformly distributed. CONCLUSIONS: Although the central region of the macula is where the density of parvocellular innervation is greatest, there is no reason to suppose that the VEPs to stimulation of the nasal and temporal regions (almost all P100 activity arising from within the central 10 degrees ) are mediated by fibres of another type. Consequently it is suggested that the central fibres were most affected by demyelination, not on account of their belonging to the parvocellular type but because of their particular situation in the optic nerve. Centrally located fibres may experience greater exposure to factors causing demyelination, or fibres located closer to the edge of the plaque may undergo more effective remyelination in the first few weeks after the acute episode. PMID- 11118245 TI - Facilitation of transmission in heteronymous group II pathways in spastic hemiplegic patients. AB - OBJECTIVE: A potent heteronymous group II excitation of quadriceps motor neurons has been recently demonstrated in normal subjects. The present study was undertaken to investigate whether this heteronymous group II excitation also contributes to spasticity in hemiplegic patients. METHOD: The early and late facilitations of the quadriceps H reflex elicited by a conditioning volley to the common peroneal nerve at three times motor threshold, attributed to non monosynaptic group I and group II excitations respectively, were investigated. The comparison was drawn between results obtained in 20 patients after stroke, with hemiplegia due to a vascular lesion in the territory of the middle cerebral artery, and 20 age and sex matched normal subjects. RESULTS: A significant increase in the group I as well as in the group II common peroneal nerve induced facilitation of the quadriceps H reflex was seen on the spastic side of the patients (group I: 159 (SEM 10)% of control H reflex; group II: 165 (SEM 8)%) compared with their unaffected side (group I: 126 (SEM 4)%; group II: 128 (SEM 5)%) (Wilcoxon signed rank test, p<0. 01), or to the right (group I: 132 (SEM 4)%; group II: 131 (SEM 5)%) or left (group I: 130 (SEM 3)%; group II: 135 (SEM 6)%) side of controls (Mann-Whitney U test, p<0.01). No significant correlation (Spearman rank test) was found between the degree of group I and group II induced facilitations on the spastic side of the patients and the degree of clinically assessed spasticity (Ashworth scale). CONCLUSION: These results reflect a facilitation of the transmission in the interneuronal pathway coactivated by group I and group II afferents, probably resulting from a change in their descending control in spastic hemiplegic patients. PMID- 11118246 TI - Pattern of localisation error in patients with stroke to sound processed by a binaural sound space processor. AB - OBJECTIVE: The ability of 46 patients with supratentorial stroke and 15 healthy subjects to localise sounds was tested using an apparatus with headphone and sound space processor. METHODS: With a binaural sound space processor, sounds were randomly presented from seven directions in the 180 degree frontal area of the subject at intervals of 30 degrees. The subject was asked to imagine a clock face through the horizontal plane passing through the subject's ears with 12 o'clock denoting a sound from directly in front of the subject. After each sound, the subject indicated the direction from which he or she thought the sound came by mentioning the corresponding hour hand on the clock face; therefore, the answer directions were also separated by 30 degrees. A total of 21 sounds with three sounds from each direction, were presented in random order. The error between the presented direction and the answered direction of each sound was calculated. RESULTS: The mean absolute error which does not distinguish whether an error was in the counterclockwise or clockwise direction, was larger in the patients with stroke than in the healthy subjects. Overall, the patients with stroke who had right brain damage (n=29) had a larger mean absolute error than those who had left brain damage (n=17). The patients with right brain damage did not show any systematic deviation such as a rightward error or leftward error. CONCLUSION: A right brain lesion or left brain lesion can cause a patient to have error in sound localisation, and patients with right brain damage generally have a larger mean absolute error of sound localisation. The difference in the mean absolute error of sound localisation between patients with stroke with right brain damage and those with stroke with left brain damage may be explained by the inattention theory of hemispatial neglect. PMID- 11118248 TI - Excitability properties of motor axons in patients with spontaneous motor unit activity. AB - OBJECTIVES: Measures of nerve excitability provide information about biophysical properties of peripheral axons in disease states. One measure, the strength duration time constant (tau(SD)), was previously reported to be prolonged in motor axons of patients with acquired neuromyotonia. The present study used a new protocol that applies a more comprehensive and sensitive panel of measures of axonal excitability, to determine firstly whether changes in tau(SD) were present in a group of patients with evidence of spontaneous motor unit activity; and secondly, if such changes in tau(SD) were present, whether other parameters of axonal excitability were affected, to clarify the mechanism of the change in tau(SD). METHODS: Eleven patients with both symptoms and EMG evidence of spontaneous motor unit activity were studied. Eight patients had autoimmune associated acquired neuromyotonia (aNMT) and three had the cramp fasciculation syndrome. The protocol first measured stimulus-response behaviour using two stimulus durations (from which the distribution of strength-duration time constants was estimated), and then threshold tracking was used to determine threshold electrotonus to 100 ms polarising currents, a current-threshold relation (indicating inward and outward rectification), and the recovery of excitability after supramaximal activation. RESULTS: The results were compared with previously published normal data. The value for tau(SD) of motor axons in the patient group was 0.43 (0. 02) ms (mean (SEM)), identical with the control value. Most other indices of axonal excitability, including those dependent on fast potassium channels, were also found to be normal. When compared with age matched controls however, the patients with acquired neuromyotonia had significantly greater late subexcitability after an impulse, greater excitability overshoots after depolarisation or hyperpolarisation, and more accommodation. CONCLUSIONS: No clear evidence for the mechanism of ectopic discharge in these patients was obtained, probably because the activity was generated focally, and most often at the motor nerve terminals. The unexpected finding of increased excitability overshoots and accommodation compared with age matched controls, suggests a relative up regulation of slow potassium conductance, possibly as a consequence of the continuous motor unit activity. PMID- 11118247 TI - Anti-GQ1b IgG antibody syndrome: clinical and immunological range. AB - OBJECTIVES: To clarify the nosological relation among Miller Fisher syndrome (MFS), Guillain-Barre syndrome (GBS) with ophthalmoplegia, Bickerstaff's brain stem encephalitis (BBE), and acute ophthalmoparesis without ataxia. Serum samples from patients with each condition often have anti-GQ1b IgG antibody. METHODS: Information on antecedent illness, initial symptoms, neurological signs during the illness, and CSF findings were reviewed in 194 patients with anti-GQ1b IgG. It was determined whether overlapping MFS and GBS (MFS/GBS), as well as overlapping BBE and GBS (BBE/GBS), is explained by the combined action of anti GQ1b IgG and anti-GM1 or anti-GD1a IgG, serological markers of GBS. RESULTS: Based on the diagnostic criteria, all the patients with acute ophthalmoparesis, MFS, MFS/GBS, BBE/GBS, and BBE had external ophthalmoplegia; all the patients with MFS, MFS/GBS, or GBS had hyporeflexia or areflexia; and all those with MFS and BBE showed ataxia. Tendon reflexes were decreased or absent in 91% of those with BBE/GBS, 67% of those with BBE, and 53% of those with acute ophthalmoparesis. Ataxia was present in 68% of the patients with MFS/GBS and 45% of those with BBE/GBS. Antecedent illness caused by upper respiratory tract infection had occurred in 60% to 80% of these patients, and CSF albuminocytological dissociation in 25% to 75%. Anti-GM1 or anti-GD1a IgG was present in 50% of those with GBS, 35% of those with MFS/GBS, 27% of those with BBE/GBS, 16% of those with MFS, and 8% of those with BBE. CONCLUSIONS: These findings together with the common autoantibody (anti-GQ1b IgG) suggest that a common autoimmune mechanism functions in the pathogenesis of these illnesses. In a larger study, it was confirmed clinically that MFS, GBS, BBE, and acute ophthalmoparesis are closely related, forming a continuous range. This is supported by the immunological findings. The term "anti-GQ1b IgG antibody syndrome" is not intended to be used as a clinical diagnosis, but recognition of this syndrome is useful for understanding the aetiological relation among the various illnesses and for introducing the established treatments of GBS for use with other conditions. PMID- 11118249 TI - X-linked sideroblastic anaemia with ataxia: another mitochondrial disease? AB - OBJECTIVES: The syndrome of X-linked sideroblastic anaemia with ataxia is rare, described only twice in the literature. The aim was to obtain clinical neurological and haematological data about this rare syndrome throughout adult life. METHODS: A family is described with two affected brothers and two affected maternal uncles. The family was evaluated clinically. Haematological investigations included full blood count, blood film, iron studies, free erythrocyte protoporphyrin (FEP) concentrations and a bone marrow examination where possible. RESULTS: Core neurological features included motor delay, ataxia evident from early childhood, and dysarthria. Neurological features were non progressive until the fifth decade when slow progression became evident. Some family members showed mild spasticity. Patients usually have a mild asymptomatic anaemia or a borderline decreased mean corpuscular volume. Blood film examination showed Pappenheimer bodies. Bone marrow examination showed ring sideroblasts, indicating raised erythrocyte iron. Free erythrocyte protoporphyrin (FEP) concentrations were raised. CONCLUSIONS: Haematological features are subtle and can be easily overlooked, and individual patients may not display all the abnormal features. X-linked ataxias are rare and incorrect genetic advice may be given if the diagnostic haematological features of X-linked sideroblastic anaemia are overlooked. Males with early onset ataxia should have a haematological evaluation including a blood film, with a bone marrow examination if abnormal blood count indices and measurement of FEP concentrations raise suspicion. The condition has parallels with Pearson's syndrome and Friedreich's ataxia. All three conditions are associated with mitochondrial iron handling defects and ataxia. The human ATP binding cassette gene (hABC7) is a candidate gene and requires further investigation. PMID- 11118250 TI - Reduced item set for the amyotrophic lateral sclerosis assessment questionnaire: development and validation of the ALSAQ-5. AB - OBJECTIVE: The 40 item amyotrophic lateral sclerosis assessment questionnaire (ALSAQ-40) is a subjective health measure designed specifically to assess areas of importance to patients with ALS. It was designed for use in surveys and clinical trials of this patient group, and has been assessed for reliability and validity. Despite its relative brevity there are situations where an even shorter form of the instrument would be desirable. Consequently, this paper reports a process of item reduction which results in a brief five item version of the instrument. METHODS: Data from two surveys of patients with ALS who completed the ALSAQ-40 were analysed to develop a short form ALSAQ. Questionnaire items were correlated with their dimension total scores. Highly correlated items were transformed onto a scale from 0 to 100 and results compared with the parent dimension. RESULTS: Five items were selected that produced results that closely resembled those of the five dimension scores of the ALSAQ-40. CONCLUSIONS: Results on the new measure compared with the parent ("gold standard") ALSAQ-40 suggest that the measure can produce similar results to the longer form but with considerable economy. PMID- 11118251 TI - Diagnosis and surgical management of intraspinal synovial cysts: report of 19 cases. AB - OBJECTIVE: Synovial cysts of the vertebral facet joints are a source of nerve root compression. Different surgical procedures are in use, but no consensus has been formed so far as to which method should be used in synovial cysts. To clarify the role of surgical management, the efficacy of operative procedures and factors influencing the outcome in our own series of 19 patients treated between 1994 and 1998 were analysed. METHODS: Nineteen patients with a mean age of 65 years underwent surgery for medically intractable radicular pain or neurological deficits caused by synovial cysts. The patients' records were retrospectively analysed for neurological deficits, cysts diameter, operative approach, segmental hypermobility, and clinical outcome; CT and MRI were analysed for additional degenerative changes. RESULTS: In 17 patients an excellent result and in two patients a good postoperative result was achieved. Twelve patients were found to have hypermobility of the facet joints and six had spondylolisthesis. There was no correlation between cyst diameter, operative approach, and outcome. No intraoperative or postoperative complications occurred. CONCLUSIONS: Age and hypermobility may play a part in the aetiology of facet joint synovial cysts. As all operative strategies showed equally good clinical outcome, total excision via a small flavectomy as the least invasive approach should be considered therapy of choice in patients with cysts causing neurological deficits. PMID- 11118252 TI - Amitriptyline enhances the central component of physiological tremor. AB - OBJECTIVES: Postural tremor is a regularly encountered side effect of amitriptyline which can be strong enough to cause discontinuation of therapy. The aim was to characterise amitriptyline induced tremor and to assess if the central or reflex component of physiological tremor was modulated by this drug. METHODS: The postural hand tremor was measured in 15 patients on a clinical rating scale, by power spectral analysis of accelerometer, forearm flexor, and extensor EMG before and after the beginning of amitriptyline treatment for major depression or chronic pain syndrome. A coherence analysis between flexor and extensor muscles on the same side was performed. RESULTS: There was a clinically visible increase in postural tremor in a third of these patients. The tremor amplitude measured by accelerometer total power increased in every patient under amitriptyline. The EMG synchronisation as reflected by significant peaks in the flexor or extensor spectrum generally occurring at higher frequencies (8-18 Hz) than the accelerometric tremor frequencies (6-11 Hz) did not change. The number of patients with a significant flexor-extensor coherence in the 7-15 Hz range increased significantly under amitriptyline, the frequency bands of significant coherence corresponded with the EMG frequencies, and both were independent of changes to the hand's resonant frequency by added inertia. CONCLUSIONS: An enhancement of postural tremor under amitriptyline is a common phenomenon although not always clinically apparent. The increase in EMG-EMG coherence indicates an increased common central drive to the motor units as its frequency is not influenced by peripheral resonance or reflex mechanisms. This is the first account of a drug induced enhancement of the central component of physiological tremor. PMID- 11118253 TI - A neuropathological study of vascular factors in late-life depression. AB - OBJECTIVES: Depression is a common psychiatric disorder in late life and it may be associated with vascular disease processes. Although there are clinical and neuroimaging studies lending support to such a "vascular depression" hypothesis there have been no neuropathological studies to directly test this. Postmortem tissue was investigated to determine whether late life depression was associated with atheromatous change in large and medium vessels and microvascular disease in the brain. METHODS: Postmortem tissue was obtained from 20 patients with a history of at least one episode of DSM-IV major depression and 20 control subjects. Standard procedures were carried out to analyze and quantify Alzheimer type pathology (plaques, tangles, Braak staging) and cortical Lewy bodies. Coronary arteries, cerebral vessels, and aorta were rated for atheromatous disease on a 0-3 scale and the four neocortical areas were rated for microvascular disease. RESULTS: The two groups showed no significant differences in age, sex, or postmortem delay. There was a significant increase in atheromatous disease in the depressed group (p=0.023). No differences were found for microvascular disease, either in the brain generally or locally in the frontal lobes. No subject had any significant Alzheimer type or Lewy body pathology. CONCLUSIONS: Neuropathological evidence was found for an excess of atheromatous disease, related to the aortic and cerebral vessels, in late life depression. However, there was no evidence of an increase in microvascular disease. The findings broadly support the vascular depression hypothesis. PMID- 11118254 TI - Abnormal peripheral auditory asymmetry in schizophrenia. AB - OBJECTIVE: Auditory processing difficulties have been reported in schizophrenia. This study explores peripheral auditory function in patients with schizophrenia in whom certain early disturbances of auditory message filtering have been found and may be associated with certain abnormalities which are particularly localised in the left temporal lobe. METHODS: Otoacoustic emissions, including click evoked and spontaneous emissions and measurements of functioning of the medial olivocochlear efferent system were obtained from 12 chronic schizophrenic patients and compared with normative data recorded from 12 normal controls. RESULTS: Otoacoustic emission amplitudes and medial olivocochlear functioning were similar between the normal controls and schizophrenic patients; the schizophrenic patients did, however, differ from the normal controls in otoacoustic emission intensity and in medial olivocochlear asymmetry. A tendency to a higher number of spontaneous peaks, and a significantly higher click evoked otoacoustic emission response amplitude were found in the right ear compared with the left ear of schizophrenic patients. For the medial olivocochlear system, whereas normal controls showed greater attenuation in the right than in the left ear, schizophrenic patients lacked such an asymmetry. CONCLUSION: In the absence of any attention task, the findings show disturbed peripheral lateralisation in schizophrenia of mechanisms involved in auditory information filtering. Such a lack of right ear advantage in medial olivocochlear functioning may thus be a peripheral reflection of central lateralisation anomalies. PMID- 11118255 TI - Release of biochemical markers of damage to neuronal and glial brain tissue is associated with short and long term neuropsychological outcome after traumatic brain injury. AB - OBJECTIVES: The present study aimed at the analysis of release patterns of neurobiochemical markers of brain damage (neuron specific enolase (NSE) and protein S-100B) in patients with traumatic brain injury and their predictive value with respect to the short and long term neuropsychological outcome. METHODS: Serial NSE and S-100B concentrations were analysed in blood samples taken at the first, second, and third day after traumatic brain injury. In 69 patients who fulfilled the inclusion criteria (no history of neurological or psychiatric disorder or alcohol or drug dependency, blood sampling according to the scheduled time scale, aged between 16 and 65 years) standardised neurological examinations and qualitative and quantitative evaluation of CT were performed. Comprehensive neuropsychological assessment was performed in 39 subjects 2 weeks after admission and in 29 subjects at a 6 month follow up examination. RESULTS: Most patients presented with minor head injuries (GCS>/=13) at the time of admission. Six months later most patients were fully independent in activities of daily living. Two thirds of the patients, however, still had neuropsychological dysfunction. Patients with short and long term neuropsychological disorders had significantly higher NSE and S-100B serum concentrations and a significantly longer lasting release of both markers. A comparative analysis of the predictive value of clinical, neuroradiological, and biochemical data showed initial S-100B values above 140 ng/l to have the highest predictive power. CONCLUSIONS: The analysis of post-traumatic release patterns of neurobiochemical markers of brain damage might help to identify patients with traumatic brain injury who run a risk of long term neuropsychological dysfunction. PMID- 11118256 TI - Increased jugular bulb saturation is associated with poor outcome in traumatic brain injury. AB - The objective was to compare secondary insults, particularly decreases in jugular bulb oxyhaemoglobin saturation (SjO(2)), during intensive care in patients with "poor" and "good" outcomes 12 months after traumatic brain injury. A prospective observational study of patients' physiological data collected each minute from multimodality monitoring was carried out. Patients had duration of physiological insults quantified as a percentage of their validated monitoring time (once invalid data due to technical reasons were removed). Treatment protocols were designed to minimise secondary insults by maintaining intracranial pressure (ICP) less than 20 mm Hg, and cerebral perfusion pressure (CPP) greater than 70 mm Hg, with prompt correction of hypoxia and pyrexia. Twelve months after injury patients' neurological function was assessed using the Glasgow outcome scale (GOS). A poor outcome was defined as GOS 1 to 3 (group 1) and a good outcome as GOS 4 and 5 (group 2). Seventy five patients (64 male), median age of 34 years (range 15 to 70), were studied. At 12 months 33 patients had a poor outcome (group 1), and 42 a good outcome (group 2). Group 1 spent proportionately more time with SjO(2) greater than 75% compared with group 2 (p<0.05), and more time with SjO(2) below 54% (p<0.04). Group 1 patients also spent proportionately more time with CPP less than 70 mm Hg than group 2 (p<0.04). Patients in group 1 were older (p<0.04) and had a lower postresuscitation Glasgow coma score (p<0.002). There was no difference between the groups for ICP, injury severity score, peripheral pulse saturation, and pyrexia. This study confirms that secondary insults, including an increased SjO(2), occur significantly more in patients with poor outcomes. More research into strategies to reduce the impact of secondary insults, including management of increased SjO(2), is required. PMID- 11118257 TI - Clinical features and prognostic factors of cerebral venous sinus thrombosis in a prospective series of 59 patients. For The Cerebral Venous Sinus Thrombosis Study Group. AB - The prognosis of cerebral venous sinus thrombosis (CVST) is variable, and outcome may range from complete recovery to death. Prognostic factors to predict outcome in the acute phase of CVST have not been analysed in a prospective study. Prognostic factors in patients enrolled in a clinical treatment trial were prospectively investigated. Poor outcome after 12 weeks, defined as death or dependency (Oxford handicap score > or =3), was used as the principle outcome measure. Univariate relations between possible prognostic factors and outcome at 12 weeks were analyzed with chi(2) tests. Treatment and all factors associated with prognosis (p< or = 0.25) were forced into a logistic regression model with a forward selection procedure. Fifty nine patients (50 women, nine men) were studied, with a mean age of 37 years (range 18 to 80 years). After 12 weeks 10 patients (17%) had a poor outcome. The univariate identified factors related to poor outcome were papilloedema, altered consciousness, coma, age older than 33 years, diagnostic delay < or =10 days, intracerebral haemorrhage, and involvement of the straight sinus. Isolated intracranial hypertension and a delta sign on CT were associated with good outcome. In the multivariate analysis coma and cerebral haemorrhage were significantly associated with a poor outcome, with odds ratios of 8.2 (95% confidence interval (95% CI) 1. 3-50.1) and 20.7 (95% CI 1.6-264.3) respectively. Involvement of the straight sinus was also weakly, but not significantly, associated with poor outcome. In conclusion, coma and intracerebral haemorrhage are independent predictors for poor outcome of CVST. PMID- 11118258 TI - Cognitive dysfunction as a major determinant of disability in patients with heart failure: results from a multicentre survey. On behalf of the GIFA (SIGG-ONLUS) Investigators. AB - Cognitive dysfunction is a frequent finding among older patients with left ventricular systolic dysfunction; however, the clinical outcomes of such a finding are unknown. Also, disability is a common condition in heart failure, poorly responding to commonly used cardiovascular medications. The association between cognitive dysfunction and disability was assessed in 1583 patients with heart failure, but without cerebrovascular disease, previous stroke, or Alzheimer's disease, who were enrolled during 2 years of a multicentre pharmacoepidemiology survey. The association between groups of variables (demographics, comorbid conditions, medications, and objective tests, including the Hodkinson abbreviated mental test) and functional disability (as indicated by need for intensive assistance in at least one of Katz' activities of daily living) was first analysed using separate age and sex adjusted logistic regression models. Those variables, significant at a p<0.1 level in these models, were simultaneously entered into an age and sex adjusted summary regression model. Among 1583 patients suitable for analysis, cognitive dysfunction (as detected by abbreviated mental test score <7) was detected in 265/461 disabled patients, and in 150/1122 independent subjects (p<0.0001). According to logistic regression analysis, cognitive dysfunction was associated with disability (OR=6.49; 95% CI=4.39-9.59) after adjusting for potential confounders.Thus, cognitive dysfunction in patients with heart failure is independently associated with disability, which currently represents an overwhelming medical and financial problem to patients, caregivers, and public health services. As early recognition and treatment of low cardiac output states might reverse cognitive dysfunction, cost effective treatment for heart failure should include systematic diagnostic and therapeutic approaches to cognitive dysfunction. PMID- 11118259 TI - Encephalomyelitis due to Cryptococcus neoformans var gattii presenting as spinal tumour: case report and review of the literature. AB - A 24 year old immunocompetent German resident is described who developed multifocal encephalomyelitis due to infection with Cryptococcus neoformans var gatti, commonly considered a disease of tropical regions. In the light of current knowledge on the epidemiology of C neoformans var gatti and the travel history of the patient it is assumed that the infection was acquired outside Europe. As exclusive intramedullary involvement is an outstandingly rare manifestation in spinal cryptococcosis, the particular diagnostic procedure and the therapeutic strategies are discussed PMID- 11118260 TI - Herpes simplex encephalitis: involvement of apolipoprotein E genotype. AB - It was previously found that herpes simplex type 1 virus (HSV1) when present in the brain, is a risk factor for Alzheimer's disease in carriers of the type 4 allele of the gene for apolipoprotein E (apoE epsilon4), and apoE epsilon4 is a risk factor for herpes labialis. Whether a specific allele of the gene is involved in susceptibility to another disorder caused by HSV1-herpes simplex encephalitis (HSE)-has now been investigated. DNA was prepared from formalin fixed, paraffin-embedded blocks of specimens from the brain or spleen of 14 United Kingdom patients with HSE, confirmed by necropsy, and from the CSF of seven United Kingdom clinical patients with HSV1 in their CSF detected by polymerase chain reaction (PCR). ApoE genotype of the DNA from blocks was determined by seminested PCR, and of the DNA from CSF by one step PCR, followed by restriction endonuclease digestion. The apoE allele frequencies were compared with values previously obtained for 238 normal people from the United Kingdom. The apoE epsilon2 allele frequency of the patients with HSE was 26%, significantly higher than the value of 7% for the normal subjects (OR=4.6, 95% confidence interval (95% CI) 2. 0-10.8). The apoE epsilon3 and epsilon4 allele frequencies did not differ significantly between the two groups. Thus, it seems that apoE epsilon2 is a risk factor for HSE. PMID- 11118261 TI - Primary diffuse leptomeningeal gliomatosis simulating tuberculous meningitis. AB - Three patients are reported on who presented with communicating hydrocephalus due to presumed tuberculous meningitis. Subsequent clinical deterioration despite antituberculous chemotherapy prompted reassessment with FDG-PET scanning and meningeal biopsy in one case and repeat CSF cytology with special staining in the second. The third patient died and postmortem confirmed a diagnosis of primary diffuse leptomeningeal gliomatosis. In the first two patients, MRI of the entire neuraxis showed no evidence of a primary intraparenchymal tumour. These cases emphasise the need for repeated reassessment in patients with culture negative lymphocytic meningitis. In addition, this is the first report of FDG-PET scanning in leptomeningeal gliomatosis. PMID- 11118262 TI - PMP22 related congenital hypomyelination neuropathy. AB - The peripheral myelin protein 22 (PMP22) is a tetraspan membrane protein which is localised in the compact myelin of the peripheral nerves. In fibroblasts, where it was originally identified as growth arrest related factor 3 (Gas3), PMP22 has been shown to modulate cell proliferation; in the peripheral nervous system its roles are still debated. The duplication of PMP22 is the most common cause of the demyelinating form of the autosomal dominant Charcot-Marie-Tooth neuropathy (CMT1A); rarer missense mutations of PMP22 also cause CMT1A or severe dehypomyelinating neuropathies of infancy grouped under the heading of Dejerine Sottas syndrome (DSS). Here, a sporadic patient affected with DSS is described; nerve biopsy disclosed a picture of hypomyelination/amyelination with basal laminae onion bulbs and no florid demyelination and it was consistent with congenital hypomyelination neuropathy (CHN); molecular analysis disclosed a novel point mutation of PMP22 that causes a non-conservative arginine for cysteine substitution at codon 109, in the third transmembrane domain. CHN is the rarest and severest form of DSS and it is thought to reflect dysmyelination rather than demyelination. The reported case suggests that missense point mutations may alter a putative role of PMP22 in modulating Schwann cell growth and differentiation. PMID- 11118263 TI - Wilhelm Conrad Von Rontgen (1845-1923). PMID- 11118264 TI - Isolated medulla oblongata function after severe traumatic brain injury. AB - The objective was to report the first pathologically confirmed case of partly functionally preserved medulla oblongata in a patient with catastrophic traumatic brain injury.A patient is described with epidural haematoma with normal breathing and blood pressure and a retained coughing reflex brought on only by catheter suctioning of the carina. Multiple contusions in the thalami and pons were found but the medulla oblongata was spared at necropsy. In conclusion, medulla oblongata function may persist despite rostrocaudal deterioration. This comatose state ("medulla man") closely mimics brain death. PMID- 11118265 TI - Medial medullary syndrome due to vertebral artery dissection. PMID- 11118266 TI - Hashimoto's encephalopathy responding to plasmapheresis. PMID- 11118267 TI - Meningoencephalitis after streptokinase treatment. PMID- 11118268 TI - Atypical course of neuropathic Gaucher's disease: follow up from early infancy until adulthood. PMID- 11118269 TI - Creutzfeldt-Jakob disease in a young person with valine homozygosity at codon 129: sporadic or variant? PMID- 11118270 TI - Multiple sclerosis treatment trial precipitates divorce. PMID- 11118271 TI - Allodynia: a sensory analogue of motor mirror neurons in a hyperaesthetic patient reporting instantaneous discomfort to another's perceived sudden minor injury? PMID- 11118272 TI - Acquired hepatocerebral degeneration: full recovery after liver transplantation. PMID- 11118273 TI - Unexpected sudden death after lateral medullary infarction. PMID- 11118274 TI - Postictal psychosis related regional cerebral hyperfusion. PMID- 11118276 TI - Quaternary structure of the lactose transport protein of streptococcus thermophilus in the detergent-solubilized and membrane-reconstituted state PMID- 11118275 TI - HTLV-I and HIV infection of the CNS in tropical areas. PMID- 11118278 TI - Progress in renal transplantation. AB - PURPOSE: The improvements in renal transplantation over the last 10 years have been one of the great success stories in medicine. We reviewed these successes with a focus on the following: changes in demographics of donors and recipients in Canada, the benefits of new immunosuppressive regimes and the efforts to minimize their toxicity and finally, our understanding of measures to circumvent chronic rejection. MATERIALS AND METHODS: A review of current transplantation literature was performed and pertinent data presented. As well, information from the Canadian Organ Replacement Register was selected to provide an overview of changes in renal transplantation in Canada. RESULTS: Despite the stable rate of transplantation in Canada, the number of new patients starting dialysis each year roughly equals the entire national renal transplant waiting list. These patients are older and have more complex co-morbidities mandating prudent use of immunosuppression so as to minimize toxicity. Standard triple therapy consists of a calcineurin inhibitor, an antimetabolite and corticosteroids. Antibody therapy is indicated in sensitized recipients and newer monoclonal humanized antibodies offer less toxicity. Nonspecific therapies are less favorable due to unwanted side effects and we can now identify subsets of patients who are most likely to benefit from specific therapy. Newer non-nephrotoxic agents hold promise for future regimens. However a paucity of large, multicenter, randomized trials, tested against standard protocols, limits their current indications. Many immunologic and non-immunologic factors influence the outcome of renal transplantation and play a role in the development in acute and chronic rejection. CONCLUSIONS: The challenges of renal transplantation over the next 10 years are: 1) in the development of specific therapies that can be altered according to patient co-morbidities and other factors influencing outcome; 2) minimizing toxicity; 3) preventing chronic rejection; and 4) improving our national organ donation network. PMID- 11118279 TI - FPSUND: a new clinical classification of urinary incontinence. AB - Urinary incontinence is a frequent condition that is usually clinically classified into three main subgroups: urge, stress and mixed. The latter, which can account for up to 50% of the patients, is notoriously heterogeneous. It is one of the reasons why the reports of therapeutic approaches to treat incontinence vary in the medical literature and it also explains the difficulty to compare results between studies. In an attempt to address this problem and to clarify the field of urinary incontinence, we have developed new clinical classification of urinary incontinence (FPSUND) where each symptom related to incontinence is rated from 0 (no symptoms) to 3 (severe symptoms). In this acronym, "F" stands for frequency of micturition, "P" for the use of protection, "S" for the stress component of incontinence, "U" for urgency, "N" for the number of nocturnal micturition and "D" for the number of diurnal micturition. Urologists from nine different centers across Canada were asked to evaluate female patients suffering from urinary incontinence using the FPSUND classification. A total of 148 women, aged 18 to 70, suffering from urinary incontinence were thus enrolled in the study. A second, independent evaluation of the same patients was performed by registered nurses or by urodynamic technicians. The reproducibility of the classification between two observers, as measured by the Weighted Kappa score was excellent, with kappa scores between 0.47 and 0.74 (p<0.05). Overall, the users of the classification found it very easy to use in a clinical setting. We would like to propose the FPSUND classification of urinary incontinence as a useful mean to evaluate patients suffering from incontinence and as a way to assess treatment outcome. PMID- 11118280 TI - Should pathological T3 and/or margin positive prostate cancer receive adjuvant therapy? A radiation oncologist's view. AB - Patients with pathological stage T3 and/or margin positive prostate cancer after radical prostatectomy have a high risk of tumor recurrence, usually heralded by rising PSA. Adjuvant therapy such as radiotherapy and/or hormone therapy needs to be explored to provide a better outcome. To date the exact role and result of adjuvant therapy remains unclear. However there has been increasing suggestion that adjuvant radiotherapy improves local control and disease free survival. Also adjuvant hormone therapy may play a role in this group of patients with a high metastatic potential. This review article addresses the clinical significance of PT3 and/or margin positive prostate cancer and explores the rationale behind considering adjuvant therapy including postoperative radiotherapy and/or hormone therapy. PMID- 11118281 TI - Selective arterial embolization for post-traumatic high flow priapism. AB - We report on a 23 year old patient with high flow priapism following blunt perineal trauma in which arterial-cavernosal fistula was missed by penile Doppler ultrasonography but was successfully localized by arteriography and embolized using Gelfoam pledgets. Detumescence was complete in 2 days and sexual function returned to the premorbid state after 4 weeks. The diagnosis, pathophysiology, and treatment of high flow priapism and review of the literature are discussed. PMID- 11118282 TI - Prostate cancer in a hypogonadal male receiving androgen supplementation. AB - Although androgen supplementation is clearly indicated in most hypogonadal males, it is also gaining popularity in the health management of the aging male without clearly low testosterone levels as the concept of male menopause becomes more accepted and recognized. The benefits include improved energy, sexual function and bone strength. A potential downside of exogenous androgen is the provocation of an underlying prostate cancer, which now represents the most common type of cancer and the second most common cause of cancer death among men in Canada. This problem is illustrated in our case of prostate cancer in a hypogonadal male on androgen supplementation. We recommend that serum PSA and digital rectal examination (DRE) be performed prior to the initiation of and regularly throughout the course of androgen supplementation. Changes in either should be investigated appropriately and termination of androgen supplementation should be considered an important component of management of prostate cancer in these patients. PMID- 11118283 TI - Glucocorticoid-induced apoptosis in lymphocytes. AB - Although the effects of glucocorticoids on lymphocytes have been scrutinized for years, researchers have yet to determine how these hormones induce apoptosis in susceptible cells. Compelling evidence indicates that DNA binding of the GR and subsequent transcriptional regulation of specific genes is required for this process. However, it is not clear whether the activation or repression of responsive genes is essential and more importantly, which of these genes, if any, are responsible for the induction of apoptosis. This review will focus on how glucocorticoid-induced apoptosisin lymphocytes is mediated by the glucocorticoid receptor, including a discussion of GR structure, function, and recent data implicating its role in the apoptotic process. PMID- 11118284 TI - Novel oxidatively stable subtilisin-like serine proteases from alkaliphilic Bacillus spp.: enzymatic properties, sequences, and evolutionary relationships. AB - The genes for five subtilisin-like serine proteases from alkaliphilic strains of Bacillus exhibiting resistance to oxidative inactivation were cloned and sequenced. The deduced amino acid sequences of the enzymes were highly homologous (greater than 88% identity). They were composed of 638 or 639 amino acids, including a possible approximately 200-amino acid prepro-peptide, and unique stretches of approximately 160 amino acids were found in the C-terminal regions. The molecular masses of mature enzymes (433 or 434 amino acids) were approximately 45 kDa for all. Amino acid sequence comparison and phylogenetic analysis indicated that these enzymes are far removed from other known subtilisins in the line of molecular evolution. We propose that these novel proteases be categorized as a new class of subtilisins, named oxidatively stable, alkaline protease. PMID- 11118285 TI - The heart is a source of circulating cardiotrophin-1 in humans. AB - Cardiotrophin-1 (CT-1) is a new member of the interleukin (IL)-6 family of cytokines and one of the endogenous ligands for gp130 signaling pathways in the heart, which has potent hypertrophic and survival effects on cardiac myocytes. However, the clinical significance of CT-1 is poorly understood, mainly because there is no widely applicable specific and sensitive assay system for measuring plasma levels of circulating CT-1. We therefore developed a competitive radioimmunoassay (RIA) for human CT-1 with rabbit antiserum recognizing the N terminus region of human CT-1 and using recombinant human CT-1 as a calibrator. The assay displays no cross-reactivities with any of the IL-6 family of cytokines including IL-11, leukemia inhibitory factor, ciliary neurotrophic factor, and oncostatin M. The lower detection limit in buffer was found to be 43 fmol/ml, and the working range was 120-8300 fmol/ml (CV < 15%). This RIA directly recognizes CT-1-like immunoreactivity in human plasma with a mean value of 571 +/- 75 fmol/ml (mean +/- SD) in healthy volunteers. The RIA coupled with gel filtration chromatographic analyses showed that the major molecular form of circulating CT-1 corresponds to recombinant full-length human CT-1. Moreover, there is a significant increase in the plasma CT-1 concentration from the aorta and coronary sinus, which clearly indicates that the heart secretes CT-1 via the coronary sinus into the peripheral circulation. This RIA should serve as a powerful tool for investigating the clinical significance of CT-1. PMID- 11118286 TI - Knockout of the mouse glutamate cysteine ligase catalytic subunit (Gclc) gene: embryonic lethal when homozygous, and proposed model for moderate glutathione deficiency when heterozygous. AB - The biosynthesis of reduced glutathione (GSH) is carried out by the enzymes gamma glutamylcysteine synthetase (GCL) and GSH synthetase. GCL is the rate-limiting step and represents a heterodimeric enzyme comprised of a catalytic subunit (GCLC) and a ("regulatory"), or modifier, subunit (GCLM). The nonhomologous Gclc and Gclm genes are located on mouse chromosomes 9 and 3, respectively. GCLC owns the catalytic activity, whereas GCLM enhances the enzyme activity by lowering the K(m) for glutamate and increasing the K(i) to GSH inhibition. Humans have been identified with one or two defective GCLC alleles and show low GSH levels. As an initial first step toward understanding the role of GSH in cellular redox homeostasis, we have targeted a disruption of the mouse Gclc gene. The Gclc(-/-) homozygous knockout animal dies before gestational day 13, whereas the Gclc(+/-) heterozygote is viable and fertile. The Gclc(+/-) mouse exhibits a gene-dose decrease in the GCLC protein and GCL activity, but only about a 20% diminution in GSH levels and a compensatory increase of approximately 30% in ascorbate-as compared with that in Gclc(+/+) wild-type littermates. These data show a reciprocal action between falling GSH concentrations and rising ascorbate levels. Therefore, the Gclc(+/-) mouse may be a useful genetic model for mild endogenous oxidative stress. PMID- 11118287 TI - PPARalpha agonists reduce 11beta-hydroxysteroid dehydrogenase type 1 in the liver. AB - 11beta-hydroxysteroid dehydrogenase type 1 (11betaHSD1) is an enzyme that converts cortisone to the active glucocorticoid, cortisol. Cortisol-cortisone interconversion plays a key role in the regulation of glucose metabolism, since mice deficient in 11betaHSD1 are resistant to diet-induced hyperglycemia. Peroxisome proliferator activator receptors (PPAR) are key regulators of glucose and lipid homeostasis. We observed a striking downregulation of murine hepatic 11betaHSD1 expression and activity after chronic treatment of wild-type mice with PPARalpha agonists, while 11betaHSD1 in the livers of PPARalpha knockout mice, or in mice treated for only 7 h with PPARalpha agonists, was unaltered. Our results are the first to show PPARalpha agonists can affect glucocorticoid metabolism in the liver by altering 11betaHSD1 expression after chronic treatment. Regulation of active glucocorticoid levels in the liver by PPARalpha agonists may in turn affect glucose metabolism, consistent with reports of their antidiabetic effects. PMID- 11118288 TI - Nonuniform distribution of Ca(2+) uptake sites within human neutrophils. AB - The rate at which Ca(2+) returns towards the basal concentration is controlled by the action of Ca(2+) pumps, both on the plasma membrane and on organelles within the cytosol. The distribution of Ca(2+) uptake sites within the cytosol was investigated using rapid confocal imaging (55 ms/frame) of fluo3-loaded human neutrophils. In some zones within the cell, the uptake of Ca(2+) from the cytosol followed a single exponential time course, whereas in others, there was accelerated kinetics after about 3 s. Using the full array of data, to produce a cell-map of Ca(2+) uptake rates a clear nonuniformity of Ca(2+) uptake sites throughout the neutrophil cytosol was observed. The location of the Ca(2+) uptake sites did not correlate with the granules or the main body of the nucleus, but Ca(2+) uptake was highest near the vestigial Golgi/ER, the edges of the nuclear lobes and at the leading cell edge. The possibility exists that the nonuniform distribution of Ca(2+) uptake sites plays a role in restricting Ca(2+) signals with the neutrophil cytosol. PMID- 11118289 TI - The P(174)L mutation in the human hSCO1 gene affects the assembly of cytochrome c oxidase. AB - Mutations of the yeast SCO1 gene result in impaired COX assembly. Recently, heterozygous mutations in the human homologue hSCO1 have been reported in infants suffering from neonatal ketoacidotic coma and isolated COX deficiency (Valnot et al., 2000). One of the hSCO1 alleles harboured a frame shift mutation resulting in a premature stop codon, the other a missense mutation leading to a substitution of proline(174) by leucine. This position is next to the essential CXXXC motif, which is conserved in all Sco1p homologues. We used chimeric proteins with the amino-terminal portion derived from yeast Sco1p and carboxy terminal portion including the CXXXC motif from the human hSco1p to provide experimental evidence for the pathogenic nature of the P(174)L mutation. These chimeras are able to complement yeast sco1 null mutants. Introduction of the P(174)L mutation affects the function of these chimeric proteins severely, as shown by impaired COX assembly and loss of COX activity. PMID- 11118290 TI - Evidence for the presence of two homer 1 transcripts in skeletal and cardiac muscles. AB - A family of proteins, Homers 1, 2 and 3, involved in activity-dependent control of signal transduction has been recently described in neurons [Xiao, B., Tu, C. J., Petralia, R. S., Yuan, J. P., Doan, A., Breder, C. D., Ruggiero, A., Lanahan, A. A., Wenthold, R. J., and Worley, P. F. (1998) Homer regulates the association of group 1 metabotropic glutamate receptors with multivalent complexes of Homer related, synaptic proteins. Neuron 21, 707-716]. By RT-PCR and RNasePA, mRNAs transcripts for Homer 1a and Homer 1c, but not Homer 1b, are detected in both skeletal and cardiac muscles of the rat. Full-length cloning of Homer 1a and Homer 1c cDNAs has been accomplished: There is no tissue specificity, by comparing skeletal muscle, cardiac muscle and cerebellum, and there are a few species-specific base substitutions, by comparing rat and mouse sequences. The regulatory mechanism exerted via transition of Homer 1 isoform composition may be operative in striated muscles. PMID- 11118291 TI - KB-R7943 inhibits store-operated Ca(2+) entry in cultured neurons and astrocytes. AB - We have studied cyclopiazonic acid (CPA)-sensitive store-operated Ca(2+) entry (SOCE) in cultured neurons and astrocytes and examined the effect of 2-[2-[4-(4 nitrobenzyloxy)phenyl]]isothiourea (KB-R7943), which is often used as a selective inhibitor of the Na(+)-Ca(2+) exchanger (NCX), on the SOCE. CPA increased transiently intracellular Ca(2+) concentration ([Ca(2+)](i)) followed by a sustained increase in [Ca(2+)](i) in neurons and astrocytes. The sustained increase in [Ca(2+)](i) depended on the presence of extracellular Ca(2+) and inhibited by SOCE inhibitors, but not by a Ca(2+) channel inhibitor. CPA also caused quenching of fura-2 fluorescence when the cells were incubated in Mn(2+) containing medium. KB-R7943 at 10 microM inhibited significantly CPA-induced sustained increase in [Ca(2+)](i) in neurons and astrocytes. KB-R7943 also inhibited CPA-induced quenching of fura-2 fluorescence in the presence of extracellular Mn(2+). These results indicate that cultured neurons and astrocytes possess SOCE and that KB-R7943 inhibits not only NCX but also SOCE. PMID- 11118292 TI - Roles of agonist-binding sites in nicotinic acetylcholine receptor function. AB - Under equilibrium conditions, the nicotinic acetylcholine receptor from Torpedo electroplax carries two high affinity-binding sites for agonists. It is generally assumed that these are the only agonist sites on the receptor and that their occupancy results in rapid channel activation followed by slower conformational transitions that lead to the high affinity equilibrium state. These slow transitions are thought to reflect the physiological process of desensitization. Here we show that preequilibration of the high affinity sites with saturating concentrations of carbamylcholine does not diminish the ion flux response to subsequent exposure to higher (activating) concentrations of this agonist. This finding has profound implications with respect to receptor function: (1) occupancy of the high affinity sites per se does not desensitize the receptor and (2) these sites cannot be directly involved in receptor activation. It is thus necessary to invoke the presence of additional binding sites in channel opening. PMID- 11118293 TI - Activation of p42/44 and p38 mitogen-activated protein kinases by extracellular calcium-sensing receptor agonists induces mitogenic responses in the mouse osteoblastic MC3T3-E1 cell line. AB - Recently, substantial evidence has accumulated that the G-protein-coupled, extracellular calcium (Ca(2+)(o))-sensing receptor (CaR) is expressed in bone marrow-derived cells, including osteoblasts, stromal cells, monocytes macrophages, and osteoclast precursor cells. Our previous studies have shown that the mouse osteoblastic MC3T3-E1 cell line also expresses the CaR and exhibits mitogenic responses when exposed to various CaR agonists. In this study, in order to understand the signaling pathway(s) mediating this response, we studied the effects of CaR agonists on the phosphorylation of p42/44 mitogen-activated protein kinase (MAPK) (Erk1/2), p38 MAPK, and c-Jun N-terminal kinase (JNK) in MC3T3-E1 cells. Raising the level of Ca(2+)(o) (4.5 mM) or addition of the polycationic CaR agonists, gadolinium (Gd(3+)) (25 microM), neomycin (300 microM) or spermine (1 mM), each stimulated phosphorylation of both p42/44 and p38 MAPKs, but not JNK, as assessed using phospho-specific antibodies to the respective MAPKs. Furthermore, phosphorylation of p42/44 and p38 MAPK were markedly inhibited by their selective and potent inhibitors, PD98059 (50 microM) and SB203580 (10 microM), respectively. Finally, the two inhibitors suppressed [(3)H]thymidine incorporation into DNA in MC3T3-E1 cells at a normal level of Ca(2+)(o) (1.8 mM) as well as when stimulated by high (4.5 mM) Ca(2+)(o), Gd(3+), or neomycin. Thus, in mouse osteoblastic MC3T3-E1 cells, both the p42/44 and p38 MAPK cascades play pivotal roles in CaR-stimulated mitogenic responses. PMID- 11118294 TI - Multidrug resistance P-glycoprotein is not involved in cholesterol esterification. AB - The aim of the present paper is to reinvestigate the role of multidrug resistance P-glycoprotein MDR1 and MDR-associated protein (MRP1) in cholesterol esterification using well-characterized inhibitors. Using specific substrate efflux assay, we show that GF120918 (0.2 microM) and probenecid (5 mM) were specific inhibitors of MDR1 and MRP1, respectively. In HepG2 cells, neither of them affect the esterification of cholesterol derived from the uptake of cholesterol-rich lipoprotein, while both verapamil (100 microM) and progesterone (100 microM) were able to inhibit cholesterol esterification. Similar results were obtained with verapamil, progesterone, and GF120918 in the MDR1 overexpressing cells MCF7/ADR. The capacity of progesterone to reduce cholesterol esterification is not correlated with its ability to inhibit MDR1 but is rather due to direct inhibition of acyl-CoA:cholesterol acyltransferase (ACAT). We conclude that the esterification of cholesterol is not correlated with MDR1 or MRP1 activity, thus excluding their role in the intracellular transport of endocytosis-derived cholesterol. PMID- 11118295 TI - Defective remodeling of cardiolipin and phosphatidylglycerol in Barth syndrome. AB - Cardiolipin (CL) and phosphatidylglycerol (PG) are the major polyglycerophospholipids observed in mammalian tissues. CL is exclusively found in the inner mitochondrial membrane and is required for optimal function of many of the respiratory and ATP-synthesizing enzymes. The role of CL in oxidative phosphorylation is, however, not fully understood and although reduced CL content leads to aberrant cell function, no human disorders with a primary defect in cardiolipin metabolism have been described. In this paper we present evidence that patients with the rare disorder X-linked cardioskeletal myopathy and neutropenia (Barth syndrome, MIM 302060) have a primary defect in CL and PG remodeling. We investigated phospholipid metabolism in cultured skin fibroblasts of patients and show that the biosynthesis rate of PG and CL is normal but that the CL pool size is 75% reduced, indicating accelerated degradation. Moreover, the incorporation of linoleic acid, which is the characteristic acyl side chain found in mammalian CL, into both PG and CL is significantly reduced, whereas the incorporation of other fatty acids into these phospholipids is normal. We show that this defect was only observed in Barth syndrome patients' cells and not in cells obtained from patients with primary defects in the respiratory chain, demonstrating that the observed defect is not secondary to respiratory chain dysfunction. These results imply that the G4.5 gene product, which is mutated in Barth syndrome patients, is specifically involved in the remodeling of PG and CL and for the first time identify an essential factor in this important cellular process. PMID- 11118296 TI - Mutant p53 forms a complex with Sp1 on HIV-LTR DNA. AB - Many mutants of p53 activate HIV-LTR driven transcription and promote HIV replication. The region of the HIV-LTR containing Sp1-binding sites is important for this effect. In this study we test the hypothesis that mutant p53 interacts with DNA-bound Sp1 and in this way can increase transcription from Sp1-dependent promoters. We have used the breast cancer cell line MDA-MB-468 that expresses endogenous mutant p53(His273) as our source of p53 protein. First, we demonstrated that this mutant p53 participates in activating transcription from the HIV-LTR by showing that HIV-LTR-directed transcription in MDA-MB-468 cells is inhibited in a dominant-negative manner by p53(Val135). Using HIV-LTR DNA affinity chromatography, we detected coelution of p53(His273) and Sp1. We also demonstrated that this mutant p53 binds sequence specifically to the super consensus sequence (SCS) and that Sp1 coeluted with p53(His273) from a column containing this site. These data indicate that p53(His273) can associate with DNA bound Sp1 suggesting that activated HIV-LTR transcription associated with mutant p53 occurs through a DNA driven multi-protein complex. PMID- 11118297 TI - Modulation of the effects of osteoprotegerin (OPG) ligand in a human leukemic cell line by OPG and calcitonin. AB - The discovery of osteoprotegerin (OPG), osteoprotegerin ligand (OPGL), and RANK has elucidated the mechanism by which osteoblasts and stromal cells regulate osteoclastic differentiation and function and mediate the effects exerted by other hormones and cytokines. We have investigated the effects of these novel cytokines on the preosteoclastic cell line FLG 29.1. We show that OPGL alone and in combination with macrophage colony-stimulating factor (CSF-1) dramatically reduced replication and increased tartrate-resistant acid phosphatase activity. However, although FLG29.1 cells appear to adhere to the bone surface, they are not able to form resorption lacunae. OPG and calcitonin completely abolished the differentiation induced by OPGL. RANK was detectable in FLG 29.1 and the number of positive cells was increased by OPGL/CSF-1 treatment while reduced by calcitonin. We propose that calcitonin could interact with the OPG/OPGL, and its effects on RANK may explain in part the action of this hormone in suppressing bone resorption. PMID- 11118298 TI - COL9A2 allelotypes in intervertebral disc disease. AB - An allelic variation of the COL9A2 gene encoding the alpha(2)-chain of collagen IX has recently been identified as a genetic risk factor for intervertebral disc prolapse, resulting in a tryptophane (Trp) substitution at position 326 of the protein. To enable quick screening of a large population we established a single enzyme (BsmFI) restriction assay which was validated by screening disc tissue samples of 250 patients (age, 47.1 +/- 13.7 years). Positive results were confirmed by nucleotide sequencing. The Trp allele was found in three patients (1.2%) who suffered from their first prolapse and were significantly older (70.7 +/- 8.5 years) than the other 247 patients. Since the substitution affects a domain covalently linked to collagen II fibrils, we conclude that this allelotype may contribute to reduced collagen crosslinking, disc instability and eventually prolapse in the elderly. PMID- 11118299 TI - EWS fli-1 antisense nanocapsules inhibits ewing sarcoma-related tumor in mice. AB - EWS Fli-1, a fusion gene resulting from a t(11;22) translocation is found in 90% of both Ewing's sarcoma and primitive neuroectodermal tumor (PNET). In the present study, we show that recently developed polyisobutylcyanoacrylate nanocapsules with an aqueous core were able to encapsulate efficiently high amounts of phosphorothioate oligonucleotides (ODN) directed against EWS Fli-1 chimeric RNA. Release of these ODN in serum medium was shown to be biphasic which was explained by the presence of two types of nanocapsules able to release ODN with different kinetics. In addition, nanocapsules were found to provide protection of these oligonucleotides from the degradation in serum. These ODN nanocapsules permitted to obtain inhibition of Ewing sarcoma-related tumor in mice after intratumoral injection of a cumulative dose as low as 14.4 nanomoles. This new type of non viral vector shows great potential for in vivo administration of oligonucleotides. PMID- 11118300 TI - Ginkbilobin, a novel antifungal protein from Ginkgo biloba seeds with sequence similarity to embryo-abundant protein. AB - A novel single-chained antifungal protein with a molecular weight of 13 kDa displaying an N-terminal sequence with marked similarity to embryo-abundant protein from the white spruce was isolated from the seeds of Ginkgo biloba using ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi gel blue gel, ion exchange chromatography on SP-Sepharose, and then gel filtration on Superdex 75. The protein, designated ginkbilobin, exerted potent antifungal activity against a variety of fungi, including Botrytis cinerea, Mycosphaerella arachidicola, Fusarium oxysporum, Rhizoctonia solani, and Coprinus comatus. Ginkbilobin exhibited a moderate antibacterial action against Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. It suppressed the activity of HIV-1 reverse transcriptase and the proliferation of murine splenocytes. PMID- 11118301 TI - Inhibition of human platelet aggregation by nitric oxide donor drugs: relative contribution of cGMP-independent mechanisms. AB - Inhibition of platelet activation by nitric oxide (NO) is not exclusively cGMP dependent. Here, we tested whether inhibition of platelet aggregation by structurally distinct NO donors is mediated by different mechanisms, partly determined by the site of NO release. Glyceryl trinitrate (GTN), sodium nitroprusside (SNP), S-nitrosoglutathione (GSNO), diethylamine diazeniumdiolate (DEA/NO), and a novel S-nitrosothiol, RIG200, were examined in ADP (8 microM)- and collagen (2.5 microgram/ml)-activated human platelet rich plasma. GTN was a poor inhibitor of aggregation whilst the other NO donors inhibited aggregation, irrespective of agonist. These effects were abolished by the NO scavenger, hemoglobin (Hb; 10 microM, P < 0.05, n = 6), except with high concentrations of DEA/NO, when NO concentrations exceeded the capacity of Hb. However, experiments with the soluble guanylate cyclase inhibitor, ODQ (100 microM), indicated that only SNP-mediated inhibition was exclusively cGMP-dependent. Furthermore, the cGMP-independent effects of S-nitrosothiols were distinct from those of DEA/NO, suggesting that different NO-related mediators (e.g., nitrosonium and peroxynitrite, respectively) are responsible for their actions. PMID- 11118302 TI - Cloning and characterization of thermostable endoglucanase (Cel8Y) from the hyperthermophilic Aquifex aeolicus VF5. AB - Aquifex aeolicus is the hyperthermophilic bacterium known, with growth temperature maxima near 95 degrees C. The cel8Y gene, encoding a thermostable endoglucanase (Cel8Y) from Aquifex aeolicus VF5, was cloned into a vector for expression and expressed in Escherichia coli XL1-Blue. A clone of 1.7 kb fragment containing endoglucanase activity, designated pKYCY100, was sequenced and found to contain an ORF of 978 bp encoding a protein of 325 amino acid residues, with a calculated molecular mass of 38,831 Da. This endoglucanase was designated cel8Y gene. The endoglucanase has an 18-amino-acid signal peptide but not cellulose binding domain. The endoglucanase of A. aeolicus VF5 had significant amino acid sequence similarities with endoglucanases from glycosyl hydrolase family 8. The predicted amino acid sequence of the Cel8Y protein was similar to that of CMCase of Cellulomonas uda, BcsC of Escherichia coli, CelY of Erwinia chrysanthemi, and CMCase of Acetobacter xylinum. The molecular mass of Cel8Y was calculated to be 36,750 Da, which is consistent with the value obtained from result of CMC-SDS PAGE of the purified enzyme. Cel8Y was thermostable, exhibiting maximal activity at 80 degrees C and pH optima of 7.0 and with half-lives of 2 h at 100 degrees C, 4 h at 90 degrees C. PMID- 11118303 TI - Characterization of spontaneous mutation in the oxyR strain of Escherichia coli. AB - Escherichia coli K-12 strain EY5, deficient in oxyR, was constructed to assess the role of oxyR and oxyR-regulated regulon in spontaneous mutagenesis. Mutagenesis was monitored by selecting two forward mutations of colicin B sensitive to resistance and valine-sensitive to resistance, one base substitution mutation of rifampicin-sensitive to resistance and one reversion of argE3 his-4 to Arg(+) His(+). Deficiency of oxyR did not lead to the enhancement of spontaneous mutation frequencies of the four markers tested. By DNA sequence analysis, we determined 49 colicin B-resistant mutants derived from EY5 and found that 37% were base substitutions, 29% IS element insertions, 20% deletions, and 14% single base frameshifts. Among the base substitutions, G:C-->T:A transversions predominated followed by G:C-->A:T transitions and A:T-->T:A transversions. These spectra were essentially the same as those from oxyR(+) strains. The results indicate that oxyR and oxyR-regulated genes do not play a significant role in the defense against spontaneous mutagenesis. PMID- 11118304 TI - Evidence for multiple active states of Plasmodium falciparum hypoxanthine-guanine xanthine phosphoribosyltransferase. AB - The lack of de novo purine biosynthesis in the malaria parasite Plasmodium falciparum makes the purine salvage enzyme hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGXPRT) a drug target. However, high activities for the purified recombinant enzyme have not been achieved, indicating that the P. falciparum enzyme requires very precise conditions for its maximal activity. In this report we have standardized the activation conditions necessary for high levels of activity, which is critically dependent on the ratios of enzyme, phosphoribosylpyrophosphate (PRPP), hypoxanthine, and buffer conditions. We demonstrate that excess substrates will push the enzyme to a less active state. We also present evidence for the existence of different kinetic states of the enzyme during activation and storage. Our kinetic data show that hypoxanthine is the substrate with highest affinity for the enzyme with a K(m) well below 1 microM. The activated enzyme has a maximum activity of 8.370 micromol/min/mg for hypoxanthine which is 10.8 times more than the previous reports. We discuss the biological relevance and implications of these results on drug design efforts. PMID- 11118305 TI - A novel endogenous PP2C-like phosphatase dephosphorylates casein kinase II phosphorylated Physarum fragmin. AB - Plasmodial fragmin, a Physarum polycephalum F-actin severing and capping protein, is phosphorylated by casein kinase II at Ser(266) (De Corte, V., Gettemans, J., De Ville, Y., Waelkens, E., and Vandekerckchove, J. (1996), Biochemistry 35, 5472 5480). In this study, we report the purification and characterization of the corresponding fragmin phosphatases. One of the enzymes was purified to near homogeneity from a cytosolic extract; it dephosphorylates CKII-phosphorylated fragmin, a peptide encompassing the CKII phosphorylation site of fragmin as well as histone 2A, CKII-phosphorylated casein and the CKII model-peptide substrate: R(3)E(3)S(P)E(3). Its activity was highly stimulated by Mn(2+) and Mg(2+), and based on its lack of sensitivity toward phosphatase effectors we could exclude similarities with PP1, PP2A and PP2B phosphatases. All biochemical properties of the phosphatase point to a PP2C-like enzyme. A second phosphatase dephosphorylating fragmin was identified as a Physarum alkaline phosphatase. PMID- 11118306 TI - Dissociation of Kar2p/BiP from an ER sensory molecule, Ire1p, triggers the unfolded protein response in yeast. AB - The unfolded protein response (UPR) is a signal transduction pathway induced by a variety of endoplasmic reticulum (ER) stresses and functions to maintain homeostasis of the cellular membrane in eukaryotes. Various ER stresses result in the accumulation of unfolded proteins in the ER, which is sensed by the transmembrane protein kinase/ribonuclease Ire1p that transmits a signal from the ER to the nucleus in Saccharomyces cerevisiae. Here we report that the yeast ER chaperone Kar2p/BiP, a member of the HSP70 family found in the ER, directly regulates the UPR by the interaction with Ire1p. In the absence of ER stress, Kar2p binds the lumenal domain of Ire1p and keeps Ire1p in an inactive unphosphorylated state. Upon exposure of cells to ER stresses, Kar2p is released from Ire1p, resulting in activation of Ire1p and signal transduction to the nucleus. Subsequently, KAR2 mRNA is induced and Kar2p accumulates in the ER in a time-dependent manner, restoring the system to the basal state. This negative autoregulation is similar to the regulation of mammalian cytosolic chaperone Hsp70 via its interaction with heat shock factor 1. PMID- 11118307 TI - Cloning of 17beta-hydroxysteroid dehydrogenase-I cDNAs from Japanese eel ovary. AB - 17beta-hydroxysteroid dehydrogenase-I (17beta-HSD-I) is a key steroidogenic enzyme for estradiol-17beta (E(2)) production. cDNAs encoding 17beta-HSD-I were cloned for the first time in lower vertebrates from the ovary of a teleost, the Japanese eel. The deduced amino acid sequence from these cDNAs was approximately 50% identical to mammalian 17beta-HSD-Is. 17beta-HSD-I mRNA was not detected in previtellogenic ovaries by Northern blotting. However, transcript abundance increased in early vitellogenic ovaries obtained from fish artificially matured by gonadotropic treatment, but thereafter did not appear to change further. Recombinant 17beta-HSD-I expressed in human kidney 293 cells selectively converted estrone to E(2), but androstenedione, testosterone, or E(2) were not converted to any other steroids. Although it is widely accepted that E(2) is produced from testosterone in other species of teleosts, the substrate specificity of eel 17beta-HSD-I suggests that a steroidogenic pathway for production of E(2) from androstenedione via estrone exists in the Japanese eel ovary. PMID- 11118308 TI - Bioluminescent Mycobacterium aurum expressing firefly luciferase for rapid and high throughput screening of antimycobacterial drugs in vitro and in infected macrophages. AB - The slow growth and highly infectious nature of Mycobacterium tuberculosis is a limiting factor in its use as test organism in high throughput screening for inhibitory compounds. To overcome these problems, use of surrogate strains and reporter genes have been considered. In this study, we have investigated the application of a fast growing nonpathogenic M. aurum expressing firefly luciferase in rapid screening of antituberculosis compounds in vitro and in infected macrophages using bioluminescence assay. The assay is based on luminescence determination using luciferin as substrate. Inhibition of bioluminescence was obtained with frontline antimycobacterial drugs like streptomycin, rifampicin, isoniazid, ethambutol, ofloxacin, and sparfloxacin at their reported MICs. Inhibition could be observed as early as 2 h in vitro and within 24 h in infected macrophages. The system can reliably be used in high throughput screening. PMID- 11118309 TI - Amino acid residues conferring herbicide tolerance in tobacco acetolactate synthase. AB - Acetolactate synthase (ALS) is the common enzyme in the biosynthetic pathways leading to valine, leucine, and isoleucine in plants and microorganisms. ALS is the target site of several classes of structurally unrelated herbicides including sulfonylureas, imidazolinones, and triazolopyrimidines. To identify the residues conferring herbicide tolerance in tobacco ALS, site-directed mutagenesis for three residues, Ala121, Pro187 and Ser652, was performed. Mutant A121T showed strong resistance to Londax (a sulfonylurea) and Cadre (an imidazolinone), while mutant S652T was resistant only to Cadre. The S652N mutation abolished the binding affinity of FAD, and inactivated the enzyme. Double mutation of Ala121 and Ser652 with Thr yielded a mutant highly tolerant to Londax, Cadre, and TP (a triazolopyrimidine sulfonamide), but has enzymatic properties similar to those of wild-type. Substitution of Pro187 with Ser resulted in the enzyme highly susceptible to oxidation and fragmentation. These results suggest that two residues Ala121 and Ser652 are potent residues conferring herbicide resistance in tobacco ALS, and that double mutation of Ala121 and Ser652 by Thr can confer stronger tolerance to Londax, Cadre, and TP. PMID- 11118310 TI - Functional complementation by wheat eIF2alpha in the yeast GCN2-mediated pathway. AB - Translational control by specific eIF2alpha phosphorylation on serine 51 has been characterized in all eukaryotes with the significant exception of plants. In order to evaluate the capability of plant eIF2alpha to functionally control translation, the wild type (51S) and a nonphosphorylatable mutant (51A) of wheat eIF2alpha were expressed in a yeast genetic system. Expression of either wheat protein did not handicap growth under conditions that repress the eIF2alpha phosphorylation pathway. However, under conditions that induce specific eIF2alpha phosphorylation only strains expressing wheat 51S were able to grow between 2 and 4 days. Growth was dependent upon activity of yeast eIF2alpha kinase GCN2 and resulted in the increased translation of GCN4. The association between plant eIF2alpha and yeast eIF2B is supported by their specific coimmunoprecipitation from transgenic yeast cells. These data support the similarity among eukaryotic translational initiation processes and strengthen the concept that plants may contain an eIF2alpha phosphorylation pathway. PMID- 11118311 TI - Recognition of 2'-deoxy-l-ribonucleoside 5'-triphosphates by human telomerase. AB - Telomerase is classified as one of the reverse transcriptases (RTs). To clarify whether l-enantiomers of natural 2'-deoxyribonucleoside 5'-triphosphates (dNTPs) are recognized by human telomerase, a quantitative telomerase assay based on the "stretch PCR" method was developed and used for kinetic analysis of the inhibitory effects of these compounds on the enzyme. Among the four l-enantiomers of dNTPs, l-dTTP and l-dGTP inhibited telomerase activity and the others showed slight or no inhibitory effect. Lineweaver-Burk plot analysis showed that the inhibition modes of l-dTTP and l-dGTP were partially competitive (mixed type) and competitive with the corresponding substrate dNTP, respectively. However, the K(i) values of l-dTTP and l-dGTP (21 and 15 microM) were several times larger than the K(m) values (3-6 microM). These results suggest that the active site of telomerase is not able to discriminate strictly the chirality of dNTPs, although it is more discriminatory than HIV-1 RT. PMID- 11118312 TI - A novel RING finger-B box-coiled-coil protein, GERP. AB - The RING finger domain occurs in a wide variety of proteins involved in cellular regulation. The polymerase chain reaction was used to search for novel RING finger proteins, using primers derived from expressed sequence tags (ests). A cDNA encoding a novel RING finger protein expressed in brain, lung, breast, placenta, kidney, muscle, and germinal center B cells is described. The human gene is expressed in a variety of tumors, including anaplastic oligodendroglioma and maps to chromosome 10q24.3, a region showing frequent deletion or loss of heterozygosity in glioblastomas. It was therefore designated glioblastoma expressed RING finger protein (GERP). GERP contains an N-terminal RING finger, followed by two B-boxes and a coiled-coil, and thus belongs to the RBCC subfamily of RING finger proteins. The structure of this protein and its mapping to a locus thought to harbor tumor suppressor genes indicates that it may be a new tumor suppressor gene important in gliomas and other malignancies. PMID- 11118313 TI - Identification and characterization of a novel gene KE04 differentially expressed by activated human dendritic cells. AB - To better understand the molecular mechanisms of dendritic cells (DC) function, we isolated differentially expressed genes in Ag-activated DC by a PCR-based subtractive hybridization technique. A novel full-length cDNA, KE04, was thus isolated from KLH-activated human PBMC-derived DC. KE04 cDNA encoded a 346-aa protein devoid of functionally indicative motifs. KE04 protein showed 64% identity with a Caenorhabditis elegans protein and 83% identity with a human putative protein. Distant relationship was also found with other prokaryotic and eukaryotic proteins. Differential expression of KE04 in activated DC other than nonactivated DC was confirmed at both mRNA and protein levels. KE04 mRNA expression was detectable in various human tissues and cell lines by Northern blot and RT-PCR. Western blot and confocal microscopy analysis indicated that its cytolocalization was intracellular. Our data suggest the potential involvement of KE04 in DC activation and will facilitate the research of molecular mechanism of DC function. PMID- 11118314 TI - Reciprocal modulation of transcriptional activities between HIV-1 Tat and MHC class II transactivator CIITA. AB - HIV-1 is the etiologic agent of acquired immune deficiency syndrome (AIDS). Functional loss of antigen-presenting cells (APC) in HIV-1 infection is considered to be involved in AIDS pathogenesis. We found that actions of the viral transactivator Tat and the transactivator of MHC class II genes, CIITA, are mutually inhibitory. While Tat inhibited expression of MHC class II genes in APC, overexpression of CIITA inhibited Tat and subsequently HIV-1 replication. This action of Tat appears to be mediated by sequestering the common cofactor, cyclin T1, but not p300 and CBP. These reciprocal actions between Tat and CIITA not only explains the functional impairment of APC in HIV-1 infection but also rationalizes the suppression of HIV-1 virus load by induction of CIITA such as IFN-gamma. PMID- 11118315 TI - Hepatocyte growth factor induces differentiation of adult rat bone marrow cells into a hepatocyte lineage in vitro. AB - Bone marrow (BM) cells originally include alpha-fetoprotein (AFP)- and c-Met [a receptor for hepatocyte growth factor (HGF)]-expressing cells. In vitro treatment of BM cells with HGF induced albumin-expressing hepatocyte-like cells. Furthermore, those hepatocyte-like cells expressed cytokeratins 8 and 18, which are typically expressed in normal adult hepatocytes. These findings demonstrate that BM cells include AFP-expressing hepatic progenitor cells that can be differentiated into hepatocytes by HGF in culture, indicating that such cultures are useful resources for cell transplantation therapy for liver diseases. PMID- 11118317 TI - Proline alleviates salt-stress-induced enhancement in ribulose-1, 5-bisphosphate oxygenase activity. AB - Sodium chloride enhanced oxygenase activity while curtailing carboxylase activity of Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase; EC 4.1.1.39) purified to electrophoretic homogeneity. Exposure to 200 mM NaCl brought about an increase in the potential of Rubisco to oxygenate RuBP by over 50%. On the other hand, proline suppressed both oxygenase as well as carboxylase activities of Rubisco. Interestingly, proline-induced suppression in oxygenase activity was significantly higher than that of carboxylase activity. Most amazingly, salt stress-induced enhancement in oxygenase activity was fully alleviated by proline even when present at a concentration as low as 50 mM. The findings presented in this communication clearly demonstrate for the first time that stress-induced proline accumulation might have a critical role in lowering the loss in fixed carbon by curtailing salt-stress-promoted enhancement in oxygenase activity of Rubisco. PMID- 11118316 TI - Deficiencies of hippocampal Zn and ZnT3 accelerate brain aging of Rat. AB - We examined the link of hippocampal Zn to the functional impairments with aging using senescence-accelerated mouse prone 10 (SAMP10) with deficits in learning and memory. Zn in hippocampal mossy fiber pathway was less distributed in aged SAMP10 than that in the age-matched control. Furthermore, expression of Zn transporter 3, ZnT3, which plays to accumulate Zn in synaptic vesicles in the mossy fiber pathway, was markedly reduced in the hippocampal region even in young SAMP10. Moreover, excessive presynaptic release of glutamate as well as glycine and expression of glial fibrillary acidic protein, a marker of neuronal cell injury, were observed in the hippocampus of aged SAMP10 compared to the control. The present results suggest that age-dependent deficiencies of Zn in synaptic vesicles of the mossy fiber pathway induced by low expression of ZnT3 cause glutamatergic excitotoxicity in the hippocampal neurons and the deterioration of learning and memory in SAMP10. PMID- 11118318 TI - Mutagenesis of organophosphorus hydrolase to enhance hydrolysis of the nerve agent VX. AB - Organophosphorus hydrolase (OPH) is capable of hydrolyzing a wide variety of organophosphorus pesticides and chemical warfare agents. However, the hydrolytic activity of OPH against the warfare agent VX is less than 0.1% relative to its activity against parathion and paraoxon. Based on the crystal structure of OPH and the similarities it shares with acetylcholinesterase, eight OPH mutants were constructed with the goal of increasing OPH activity toward VX. The activities of crude extracts from these mutants were measured using VX, demeton-S methyl, diisopropylfluoro-phosphate, ethyl parathion, paraoxon, and EPN as substrates. One mutant (L136Y) displayed a 33% increase in the relative VX hydrolysis rate compared to wild type enzyme. The other seven mutations resulted in 55-76% decreases in the relative rates of VX hydrolysis. There was no apparent relationship between the hydrolysis rates of VX and the rates of the other organophosphorus compounds tested. PMID- 11118319 TI - First-generation monoclonal antibodies identifying the human leukotriene B(4) receptor-1. AB - The leukotriene B(4) receptor (BLTR) is a seven-transmembrane chemoattractant receptor that is important in pro-inflammatory responses. We have produced the first widely applicable monoclonal antibodies against the human BLTR and confirmed the antibody specificity using flow cytometric analysis of three different cell lines stably expressing the recombinant receptor. The antibodies did not cross-react with the recently cloned second LTB(4) receptor, BLTR2, or the Cys LT1 and Cys LT2 receptors. Functional analysis in combination with two color flow cytometry showed that the BLTR antibodies bind to cells that are activated by LTB(4). The antibodies were shown to recognize BLTR in cell ELISA and immunocytochemistry. Endogenous expression of BLTR in CD15-positive blood leukocytes and in differentiated HL-60 cells was also demonstrated with the antibodies. PMID- 11118320 TI - Two novel genes, human neugrin and mouse m-neugrin, are upregulated with neuronal differentiation in neuroblastoma cells. AB - We herein report two new genes, human neugrin and mouse homologue m-neugrin, found by screening the cDNA library for the human spinal cord. The neugrin mRNA encodes 219 amino acids and its deduced amino acid sequence contains an NLS-like domain. No previously known motif is found in it. m-neugrin mRNA encodes 233 amino acids. Neugrin and m-Neugrin are 70% homologous in amino acid sequence. Northern analysis revealed that neugrin was strongly expressed in the heart, brain, and skeletal muscle, and m-neugrin in the liver, kidney, and brain. A transfection study indicated that these proteins are localized in the nucleus. Although the expression of neugrin was found to be ubiquitous in the nervous system, in situ hybridization showed that both neugrin and m-neugrin were expressed mainly in the neurons rather than the glial cells. Their expression was highly upregulated with the neurite outgrowth associated with neuronal differentiation in neuroblastoma cell lines. These results indicate that neugrin and m-neugrin are mainly expressed in neurons in the nervous system, and play an important role in the process of neuronal differentiation. PMID- 11118321 TI - Altered subunit communication in subfamilies of trimeric dUTPases. AB - The enzyme dUTPase is essential in preventing uracil incorporation into DNA. Design of antagonists against this novel chemotherapeutic target requires identification of species-specific differences in the structure and mechanism of the enzyme. This task is now approached via comparisons of available crystallographic structures of dUTPases from Homo sapiens, Escherichia coli, and retroviruses. The eukaryotic protein uniquely displays polar and charged amino acid residues participating in threefold intersubunit interactions. In bacterial and retroviral dUTPases, threefold interactions are mainly hydrophobic. The residues responsible for this contrast are mapped in multiple sequence alignment to positions differently and characteristically conserved in distinct evolutionary branches. The general feature of this contrast is further strengthened by a second eukaryotic model structure constructed using comparative modeling. The dUTPase cDNA from Drosophila melanogaster was identified, sequenced, and the model structure of the encoded polypeptide displayed a polar hydrogen-bonding network of threefold interactions, identically to the human structure. Results allow clear distinction between two subfamilies of trimeric dUTPases where altered subunit communication may account for a functional difference in the catalytic cycle. PMID- 11118322 TI - Structure of the VPATPD gene encoding subunit D of the human vacuolar proton ATPase. AB - The HSD11B2 and VPATPD genes encoding the human kidney isozyme of 11beta hydroxysteroid dehydrogenase (11-HSD2) and subunit D of the vacuolar proton ATPase, respectively, are located on chromosome 16q22. They are transcribed from complementary DNA strands and their 3' ends are only 0.5 kilobase apart. Because polymorphisms in HSD11B2 have been associated with hypertension and salt sensitivity, we characterized the human VPATPD gene. It spans 19 kb and consists of 8 exons. The encoded protein is 99.5% identical to mouse subunit D at the amino acid level. An alternating purine-pyrimidine tract is located in the 3' untranslated region of VPATPD. On genotyping 17 hypertensive subjects, no length polymorphism was found. Although VPATPD and HSD11B2 are both expressed in kidney and placenta, they are regulated differently; forskolin upregulates HSD11B2 but not VPATPD in human choriocarcinoma JEG3 cells. The functional significance of the proximity of these two genes remains to be established. PMID- 11118323 TI - A family of compact plasmid vectors for DNA immunization in humans. AB - DNA immunization technology is based on the availability of adequate vectors for cloning and expression of heterologous immunoactive proteins in mammalian cells. We have developed a family of DNA plasmid vectors suitable to manipulate antigen expression and location. Their in vitro and in vivo functionality and application are also reported. The developed immune response, the aspects considered for vector design, and the possible independent manipulation of both blocks for the generation of bicistronic constructs, make of the pAEC family of plasmid vectors a source for DNA vaccine candidate's development for further evaluation in human clinical trials, and for potential use in the gene therapy approach. PMID- 11118324 TI - N-Acetylcysteine counteracts erythrocyte alterations occurring in chronic obstructive pulmonary disease. AB - A key role has been proposed for reactive oxygen species (ROS) in chronic obstructive pulmonary disease (COPD). Aim of the present work was to evaluate possible implications of ROS in the integrity and function of the cell type mainly involved in oxygen uptake and delivery to the peripheral tissues: the erythrocyte. Red blood cells (RBCs) were thus collected from blood samples from COPD patients. Furthermore, blood samples from the same patients treated with the antioxidizing drug of widespread use in such disease i.e., N-acetylcysteine (NAC), were also considered. Morphometric and analytical cytology studies were then conducted. We report herein that: (i) alterations of RBC ultrastructure were detectable in RBCs from COPD patients, that (ii) relevant changes of spectrin cytoskeleton and glycophorin expression were also found and that (iii) NAC treatment was capable of significantly counteracting these changes. These results are consistent with a reappraisal of the role of RBCs in this disease. PMID- 11118325 TI - The RNA aptamer-binding site of hepatitis C virus NS3 protease. AB - Nonstructural protein 3 (NS3) of hepatitis C virus (HCV) is a trypsin-like protease and is essential for processing of viral polyprotein. Accordingly, it is a potential target for anti-HCV drugs. Recently we could isolate RNA aptamers (G9 I, II, and III) which bind and inhibit NS3 protease using in vitro selection strategy. In addition, G9-I aptamer showed noncompetitive inhibition. In order to elucidate the binding site of G9-I aptamer in NS3 protease domain (deltaNS3), we carried out alanine scanning mutagenesis at positive charged residues on the surface of deltaNS3. The result of binding analysis by surface plasmon resonance measurements and protease inhibition assay clarified that Arg161 as well as Arg130 of deltaNS3 are essential for interaction with G9-I aptamer. This region appears to be a potential targeting site for anti-HCV drugs. PMID- 11118326 TI - Nonredundant roles of the elongation factor MEN in postimplantation development. AB - The MEN/ELL gene was cloned as a fusion partner of the MLL gene in the t(11;19)(q23;p13.1) translocation, which is found in adult myeloid leukemia. MEN belongs to a family of RNA polymerase II elongation factors and dysregulated production of MEN through the MLL promoter could cause malignant transformation of myeloid cells. To pursue the physiological role and determine the requirement of the MEN gene product in mouse development, we generated knockout mice (MEN-/-) by gene targeting in embryonic stem cells. After intercrossing heterozygous mice to generate homozygous mutants, we identified no homozygotes (MEN-/-) even at E9.5, as well as after birth, by Southern analysis. Moreover, histological examinations revealed degenerative changes in nearly one-fourth of E6.5 embryos, which were gradually resorbed by E8.5. Our findings demonstrated that MEN-/- mice are embryonic lethal, and die before E6.5 and after implantation. MEN should play a nonredundant role in postimplantation development of mice. PMID- 11118327 TI - Tissue and cell distribution of a mammalian proteasomal ATPase, MSS1, and its complex formation with the basal transcription factors. AB - The proteasome is an eukaryotic multi-subunit protease complex composed of one 20S core component and two 19S regulatory complexes. The regulatory complex contains 6 putative ATPases. We investigated tissue and cell distribution of one of these ATPases, MSS1 (mammalian suppressor of sgv1). MSS1 was ubiquitously present in rat tissues as was the 20S core component of proteasome. However, the ratio of MSS1 to 20S varied greatly among tissues and MSS1 was concentrated in the thymus. Glycerol gradient sedimentation analysis revealed that MSS1 is included in protein complexes whose density is lighter than that of the proteasome. MSS1 was distributed in mammalian cells ubiquitously, while proteasome was rather concentrated in the nuclei. Hence, a novel molecular status of MSS1 distinct from proteasome is implicated. Interestingly, multiple basal transcription factors for RNA polymerase II, including TBP, TFIIB, TFIIH, and TFIIF, were found to be associated with MSS1. These results suggest that MSS1, in addition to proteolysis, plays a role in DNA metabolism including transcriptional regulation. PMID- 11118329 TI - Induction of heme oxygenase-1 can act protectively against cardiac ischemia/reperfusion in vivo. AB - Enhanced production of reactive oxygen species plays a role in myocardial injury following ischemia/reperfusion. Heme oxygenase-1 (HO-1) is a heme-catabolizing enzyme that is induced by and acts against oxidant-induced tissue injury. We examined whether HO-1 expression was regulated following ischemia and reperfusion in the rat heart. HO-1 expression increased as early as 24 h after reperfusion. Strong HO-1 expression was seen in monocytes/macrophages and myofibroblasts. Next, we examined whether the induction of HO-1 could ameliorate cardiac injury following ischemia/reperfusion. Intraperitoneal hemin injection (30 mg/kg/day) for 2 days prior to the operation resulted in an about 2.8-fold increase in HO-1 expression in the rat heart. Hemin treatment significantly decreased infarct area (6 +/- 2%) compared to the control (21 +/- 2%), which was reversed by the coadministration of an HO inhibitor in a dose-dependent manner. Our data suggest that induction of HO-1 can reduce the cardiac injury in vivo following ischemia/reperfusion. PMID- 11118328 TI - Expression and evolution of the Drosophila attacin/diptericin gene family. AB - We describe the genes for three new glycine-rich antimicrobial peptides in Drosophila, two attacins (AttC and AttD) and one diptericin (DptB). Their structures support the proposal that these glycine-rich antimicrobial peptides evolved from a common ancestor and are probably also related to proline-rich peptides such as drosocin. AttC is similar to the nearby AttA and AttB genes. AttD is more divergent and located on a different chromosome. Intriguingly, AttD may encode an intracellular attacin. DptB is linked in tandem to the closely related Diptericin. However, the DptB gene product contains a furin-like cleavage site and may be processed in an attacin-like fashion. All attacin and diptericin genes are induced after bacterial challenge. This induction is reduced in imd mutants, and unexpectedly also in Tl(-) mutants. The 18w mutation particularly affects the induction of AttC, which may be a useful marker for 18w signaling. PMID- 11118330 TI - Human neuroblastoma cell differentiation requires protein kinase C-theta. AB - Neuroblastoma LAN-5 cells exposed to retinoic acid cease to multiply and extend neurite outgrowths acquiring a neuronal phenotype. We now report that protein kinase C-theta; (PKC-theta;) isozyme is involved in this differentiation process due to the following findings: (i) PKC-theta; is expressed by LAN-5 cells as a nuclear and perinuclear protein; (ii) cell stimulation with retinoic acid promotes in a large increase in the expression level of the kinase and its intracellular redistribution; and (iii) a PKC-theta; antisense oligonucleotide reduces at the same time the expression level of the kinase and the cell response to retinoic acid. Altogether these data are consistent with a specific role played by PKC-theta; in the differentiation program of neuronal cells. PMID- 11118331 TI - Multiple deletions of mtDNA remove the light strand origin of replication. AB - Idiopathic inflammatory myopathies are progressive, debilitating muscle diseases. The pathogenesis of these disorders is multifactorial and appears to include mutations of the mitochondrial genome, which are usually indicated by morphological changes of mitochondria. The vast majority of all mitochondrial DNA deletions found are located between the origins of replication in the "major region" between nt5760-nt190. Using long distance PCR and sequencing techniques, we detected deletions which were unusually large (ca. 10500-12800 bp) and show uncommon 5'-breakpoints between nt800 and nt3326. Unlike most other deletions, their breakpoints are far upstream of the "major region." The atypical location of these deletions suggests a different pathomechanism. The impact of the mitochondrial DNA deletions in the pathogenetic cascade remains uncertain. PMID- 11118332 TI - Transcriptional inactivation of p53 by deletions and single amino acid changes in mouse mot-1 protein. AB - Mouse mortalin proteins, mot-1 and mot-2, differ by only two amino acid residues in their C-terminus. In previous studies we showed that they differ in their subcellular distributions and interactions with the tumor suppressor protein, p53. By using mot-1 deletion mutants and amino acid substitution constructs, we report here that inability of mot-1 to affect p53 activity in vivo is dependent on the presence of both of the unique mot-1 amino acids and all three of the predicted hsp70, EF hand, and leucine zipper motif regions. The two proteins and their single amino acid mutants showed different mobilities on SDS-polyacrylamide gel presenting an evidence for their different secondary structures. Taken together, the data suggest that each of the two differing amino acids between mot 1 and mot-2 is an important determinant of their secondary structures and in vivo activities. PMID- 11118333 TI - High levels of zinc ions induce loss of mitochondrial potential and degradation of antiapoptotic Bcl-2 protein in in vitro cultivated human prostate epithelial cells. AB - Prostate epithelial cells contain the highest levels of zinc among all organs and tissues in the human body. Zinc is accumulated primarily in the mitochondria, where it is responsible for inhibition of mitochondrial aconitase activity, thereby increasing citrate production. The present study was designed to clarify the role of zinc for human prostate epithelial cell growth and apoptosis. Apoptosis of in vitro cultivated human prostate epithelial cells exposed to ZnCl(2) was analyzed by determination of phospholipid membrane asymmetry, nuclear fragmentation, DNA strand breaks, changes of mitochondrial potential and cellular pro/antiapoptotic proteins. Zinc induced apoptosis without involvement of p53 by decreasing mitochondrial transmembrane potential (DeltaPsi(m)) and Bcl-2 protein levels in proliferating epithelial cells. Thus, the high local concentrations of zinc ions in the prostatic lumen seem to be necessary to regulate proliferative activities and to enforce epithelial differentiation processes. PMID- 11118334 TI - Molecular cloning and characterization of a new human histamine receptor, HH4R. AB - A new histamine receptor, HH4R, was cloned from human leukocyte cDNA. The deduced amino acid sequence showed about 40% identity to that of the human histamine H3 receptor, HH3R. HH4R-expressing cells responded to histamine, inhibiting forskolin-induced cAMP accumulation. An H3 agonist, N-alpha-methylhistamine (NAMHA), bound specifically to HH4R, while another H3 agonist, R(-)-alpha methylhistamine (RAMHA), and the H3 antagonist, thioperamide, competed with this binding. RAMHA, NAMHA, and imetit inhibited forskolin-induced cAMP accumulation in HH4R-expressing cells. However, the binding affinities and agonistic activities of H3 agonists to HH4R were weaker than those to HH3R. Low expression of HH4R was detected in a wide variety of peripheral tissues by RT-PCR; however, in contrast with HH3R, expression was not detected in the brain. These observations indicate that the clone is a distinct histamine receptor from HH3R, and thus is named HH4R. PMID- 11118335 TI - Morphine upregulates mu opioid receptors of human and monkey lymphocytes. AB - Opioid receptors of subtypes delta, kappa, and mu similar to those found in brain cells have been identified in immune cells. The current study demonstrates by competitive polymerase chain reaction the treatment of human lymphocytic cells with morphine resulting in an increased amount of gene expression of mu opioid receptors. Antibodies against the MOR-1, the neuronal mu opioid receptor, were used in Western blot analysis of mu proteins and the results revealed a single band of approximately 50 kDa, the intensity of which was increased by morphine treatment. Similar results of mu opioid receptor activation were observed when monkey lymphocytes were treated with morphine. These studies suggest that in addition to causing an immune effect through communication with the neuroendocrine system, the psychoactive drug morphine may modulate immune functions by acting directly on the mu opioid receptors expressed on lymphocytes. PMID- 11118336 TI - Bioflavonoid quercetin inhibits mitosis and apoptosis of glomerular cells in vitro and in vivo. AB - Bioflavonoids have been regarded as therapeutic agents for a wide range of disease including inflammation. In this report, we investigated effects of bioflavonoid quercetin on mitosis and apoptosis of glomerular cells in vitro and in vivo. Serum-stimulated rat mesangial cells were treated with or without quercetin, and total cell number, percentages of mitotic cells, and incorporation of [(3)H]-thymidine were evaluated. All three assays showed that mitogenic activity of mesangial cells was markedly attenuated by quercetin. To examine the effect of quercetin on apoptosis, mesangial cells were pretreated with or without quercetin and stimulated by hydrogen peroxide or tumor necrosis factor-alpha. Hoechst staining and DNA ladder assay showed that both apoptotic responses were dramatically inhibited by quercetin. We further investigated effects of quercetin on in vivo mitosis and apoptosis of glomerular cells. Rats were administered with or without quercetin intraperitoneally, and nephrotoxic serum nephritis was induced. Immunohistochemical analyses showed that treatment with quercetin significantly reduced the number of proliferating cell nuclear antigen (+) cells and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (+) cells in the glomerulus. These data suggested that quercetin has the potential for inhibiting mitosis and apoptosis of glomerular cells both in vitro and in vivo. PMID- 11118337 TI - Activation of prostacyclin synthesis by mechanical stimulation in the ciliated protozoan Tetrahymena thermophila. AB - We detected a HPLC peak corresponding to 6-keto-PGF(1alpha), a stable metabolite of prostacyclin (PGI(2)), in [1-(14)C]arachidonate metabolites from the ciliated protozoan Tetrahymena thermophila. Quantitative analysis of 6-keto-PGF(1alpha) by enzyme immunoassay revealed that the synthesis and release were rapidly activated by the mechanical stimulation of a short centrifugation. The activation was suppressed significantly by a cyclooxygenase inhibitor, ibuprofen, and was independent of the extracellular Ca(2+). External addition of PGI(2) and its stable analogue, beraprost, caused a transient increase in the tumbling frequency of swimming. Other prostanoids, PGE(2) and PGF(2alpha), have no effect on the swimming. These results indicate that a free-living ciliate, T. thermophila, synthesizes and has a specific sensitivity to PGI(2). PMID- 11118338 TI - Relation between phosphatidylserine exposure and store-operated Ca(2+) entry in stimulated cells. AB - A significant increase in intracellular Ca(2+) is required to trigger the remodeling of the cell plasma membrane. Scott syndrome is an extremely rare inherited disorder of the transmembrane migration of phosphatidylserine toward the exoplasmic leaflet in blood cells. We have recently reported a reduced capacitative Ca(2+) entry in Scott cells [Martinez et al. (1999) Biochemistry 38, 10092-10098]. We have investigated here the links between defective phosphatidylserine exposure and Ca(2+) signaling in Scott cells by focusing on the Ca(2+) entry following the emptying of intracellular stores. After depletion of caffeine- or thapsigargin-sensitive stores, Ca(2+) entry was lower in Scott compared to control lymphoblasts. However, the simultaneous depletion of both types of stores restored a normal Ca(2+) influx across the plasma membrane in Scott cells and phosphatidylserine externalization ability was improved concomitantly with capacitative Ca(2+) entry. These observations point to the essential role of capacitative Ca(2+) entry in the control of phosphatidylserine exposure of stimulated cells. PMID- 11118339 TI - YC-1, a benzyl indazole derivative, stimulates vascular cGMP and inhibits neointima formation. AB - The pathobiologic process of arterial stenosis following balloon angioplasty continues to be an enigmatic problem in clinical settings. This research project investigates the ability of YC-1, a benzyl indazole derivative that sensitizes sGC/cGMP, to stimulate endogenous cGMP and attenuate balloon injury-induced neointima (NI) formation in the rat carotid artery. Northern and Western blot analyses revealed enhanced acute expression of iNOS and inducible heme oxygenase (HO-1) mRNA and protein in the injured artery. The contralateral uninjured artery also demonstrated acute HO-1 mRNA and protein induction without detectable iNOS expression. Perivascular application of YC-1 immediately following injury significantly stimulated acute vessel wall cGMP compared to untreated controls. YC-1 treated sections demonstrated significant reduction in NI area (-74%), NI area/medial wall area (-72%), and NI thickness (-76%) 2 weeks post-injury. These results directly implicate YC-1 as a potent new therapeutic agent capable of reducing post-angioplasty stenosis through endogenous CO- and/or NO-mediated, cGMP-dependent processes. PMID- 11118340 TI - Recombinant Cry3Aa has insecticidal activity against the Andean potato weevil, Premnotrypes vorax. AB - The Andean potato weevil, Premnotrypes vorax, an insect of the order Coleoptera, is a major cause of damage to potato crops in the Andean regions of South America. The insecticidal Cry proteins from Bacillus thuringiensis are useful biological pesticides, and some are toxic to Coleopteran insects. We overexpressed recombinant, histidine-tagged Cry3Aa protein in Escherichia coli host cells. The recombinant protein was solubilized at high pH with urea, purified using Ni(2+)-nitrilo-triacetic acid affinity resin, and dialysed to lower pH and remove urea. Bioassays were performed with an insect media whose surface was spread with 70 microgram/mL purified native or recombinant toxins. First instar larvae exposed to toxin treated media for 5 days exhibited mortalities from 57% (native Cry3Aa) to 52% (recombinant Cry3Aa). Purified native and recombinant Cry3Aa proteins appeared to be equally toxic to the Andean potato weevil. PMID- 11118341 TI - Myeloperoxidase-catalyzed oxidation of melatonin by activated neutrophils. AB - In the presence of hydrogen peroxide, horseradish peroxidase (HRP) catalyzes the production of N(1)-acetyl-N(2)-formyl-5-methoxykynuramine from melatonin. This reaction consumes oxygen and exhibits chemiluminescence in the 440-540 nm region. The excited cleavage product derived from the thermolysis of an intermediate dioxetane is suggested to be the emitting species. Chemiluminescence and the indole ring cleavage product were also observed when HRP/H(2)O(2) was replaced by phorbol myristate acetate or opsonized zymosan-activated neutrophils. Azide, a myeloperoxidase inhibitor, strongly suppressed melatonin oxidation. Superoxide dismutase has a strong inhibitory effect on light emission but catalase and uric acid are without effect on the emission. The oxidation of melatonin by activated neutrophils may be relevant to the in vivo functions of myeloperoxidase and melatonin. The possible biological implication of melatonin oxidation by neutrophils, especially in inflammatory conditions, is discussed. PMID- 11118342 TI - Sensitivity of trypanosomatid protozoa to DFMO and metabolic turnover of ornithine decarboxylase. AB - alpha-Difluoromethylornithine (DFMO), the specific and irreversible inhibitor of ornithine decarboxylase (ODC), was able to induce the arrest of proliferation in Leishmania mexicana and ODC-transformed Trypanosoma cruzi cultures grown in a semi-defined medium essentially free of polyamines. Conversely, Crithidia fasciculata and Phytomonas 274 were not affected by the inhibitor. The drug resistance of Crithidia and Phytomonas was neither caused by an impairment of DFMO uptake nor by a decrease of the enzyme affinity for the inhibitor. We were also able to rule out the possibility of ODC overexpression in the drug-tolerant parasites. The measurements of ODC metabolic turnover indicated that the enzymes from Crithidia and Phytomonas have a short half-life of 20-40 min, while ODC from Leishmania and transgenic Trypanosoma cruzi are rather stable with a half-life longer than 6 hours. Analyses of polyamine internal pools under different growth conditions have shown that DFMO was able to markedly decrease the levels of putrescine and spermidine in all parasites, but the depletion of spermidine was higher in trypanosomatids containing an ODC with slow turnover. Our results suggest that in these parasites cultivated in the presence of the drug, spermidine might decrease below critical levels needed to maintain trypanothione concentrations or other conditions essential for normal proliferation. PMID- 11118343 TI - Potent heterologous antifungal proteins from cheeseweed (Malva parviflora). AB - Two novel antifungal proteins were purified and characterized from cheeseweed (Malva parviflora). Both proteins, designated CW-1 and CW-2, are composed of two different subunits of 5000 and 3000 Da, respectively. These proteins possess very potent antifungal activities, and more interestingly the inhibition is fungicidal instead of fungistatic. At low salt condition, the IC(50) of CW-1 and CW-2 against Fusarium graminearum (Fg) is 2.5 ppm. At high salt condition which diminishes the antifungal activity of many antifungal proteins, both CW-1 and CW 2 still maintain potent activity against Fg with IC(50) of 10 ppm. The two subunits could be separated by gel filtration in the presence of 6 M urea, but their antifungal activity cannot be recovered after the removal of urea. Amino acid sequence analysis indicates that both subunits of CW-1 show homology to 2S albumin, whereas the two subunits of CW-2 have homology to vicilin protein from cotton. To our knowledge, this is the first report of isolation and characterization of heterologous antifungal proteins from any source. PMID- 11118344 TI - A novel conformer of oxidized Paracoccus pantotrophus cytochrome cd(1) observed by freeze-quench NIR-MCD spectroscopy. AB - Paracoccus pantotrophus cytochrome cd(1) is a physiological nitrite reductase and an in vitro hydroxylamine reductase. The oxidised "as isolated" form of the enzyme has bis-histidinyl coordinated c-heme and upon reduction its coordination changes to histidine/methionine. Following treatment of reduced enzyme with hydroxylamine, a novel, oxidised, conformer of the enzyme is obtained. We have devised protocols for freeze-quench near-ir-MCD spectroscopy that have allowed us to establish unequivocally the c-heme coordination of this species as His/Met. Thus it is shown that the catalytically competent, hydroxylamine reoxidised, form of P. pantotrophus cytochrome cd(1) has different axial ligands to the c-heme than "as isolated" enzyme. PMID- 11118345 TI - The BRCT regions of tumor suppressor BRCA1 and of XRCC1 show DNA end binding activity with a multimerizing feature. AB - The BRCT regions are two repeating structures in BRCA1 at the carboxyl-terminus and are ubiquitous in some proteins involved in cell cycle checkpoint and in DNA repair. Here, using electron microscopy, we show direct evidence that the BRCT regions of BRCA1 bound double-strand breaks of DNA. The BRCT regions could multimerize thus forming large protein particles. Smeared patterns of DNA fragments were consistently shown in the gel retardation assay. A single BRCT was sufficient for DNA binding. The smeared patterns were also observed in BRCTs of TopBP1, suggesting that multimerization may be an important feature of BRCTs. The recombinant second BRCT of XRCC1 (X-ray repair cross-complementing group 1), whose folding was determined by X-ray crystallography, also showed similar DNA end binding images. It is possible that some BRCTs are fundamental structures that detect DNA damages. PMID- 11118346 TI - APC protein is required for initiation of neuronal differentiation in rat pheochromocytoma PC12 cells. AB - The adenomatous polyposis (APC) gene product is highly expressed in the central nervous system. To elucidate the contribution of the APC protein to neuronal differentiation, we used an inducible antisense mRNA vector to suppress APC protein expression and examined neuronal differentiation of PC12 cells induced by nerve growth factor (NGF). When antisense mRNA was induced, APC protein expression was suppressed to 20% of the noninduced level. In those cells, neurite extension induced by NGF and expression of microtubule-associated protein 2 (MAP2) was completely inhibited. However, once cells had differentiated, antisense APC mRNA expression and subsequent suppression of APC protein expression had no effect on either cell morphology or MAP2 protein expression. These results suggest that the wild type APC is critically involved only in the initiation of neuronal differentiation, but not in the maintenance of the differentiated phenotype, or that the neuronal phenotype could be maintained at lower level of APC protein. PMID- 11118347 TI - A reevaluation of the peroxynitrite scavenging activity of some dietary phenolics. AB - Peroxynitrite is implicated in many diseases. Hence, there is considerable interest in potential therapeutic peroxynitrite scavengers. Diet-derived phenolics have been claimed to be powerful peroxynitrite scavengers. However, the reactivity of peroxynitrite can be significantly modified by bicarbonate and this has not been considered in evaluations of the scavenging activity of phenols. Bicarbonate (25 mM) significantly decreased the ability of several phenolic compounds (caffeic acid, o- and p-coumaric acid, gallic acid, ferulic acid) but not others (catechin and epicatechin) to inhibit peroxynitrite-mediated tyrosine nitration. Bicarbonate (25 mM) also decreased the ability of catechin, epicatechin, quercetin and ferulic acid but not chlorogenic acid, gallic acid, caffeic acid and o-coumaric acid to inhibit peroxynitrite-mediated alpha(1) antiproteinase inactivation. These results show that physiological concentrations of bicarbonate substantially modify the ability of dietary phenolics to prevent peroxynitrite-mediated reactions. When assessing compounds for peroxynitrite scavenging, experiments should be conducted in the presence of bicarbonate to avoid misleading results. PMID- 11118348 TI - TT virus mRNAs detected in the bone marrow cells from an infected individual. AB - Although replicative forms of TT virus (TTV) DNA have been found in the liver and bone marrow cells, mRNAs of TTV have not yet been detected in these tissues. The entire nucleotide sequence of a TTV clone [TYM9 (3759 bases)] isolated from a patient with high TTV viremia (10(6) copies/ml) was determined, and the poly(A)(+) RNAs from bone marrow cells were subjected to reverse transcription polymerase chain reaction with primers specific for the TYM9 sequence. Sequence analysis of the amplified products revealed the presence of three distinct species of spliced TTV mRNAs [2.9, 1.2, and 1.0 kilobases (kb)] with common 5' and 3' termini as well as splicing to bind nucleotide (nt) 181 to nt 283. The shorter mRNAs of 1.2 kb and 1.0 kb possessed another splicing to join nt 681 with nt 2341 or nt 2579. The transcription profile of TTV found in an infected human corroborates that observed in vitro. PMID- 11118349 TI - Cofactor recycling with immobilized heterologous cytochrome P450 105D1 (CYP105D1). AB - Immobilisation of cells and enzymes can be a convenient and rapid way for testing and transforming substances. Cytochromes P450 may be useful in numerous biotransformations of varied lipophilic substrates, performing both regio- and stereo-specific monooxygenation reactions. However, one limitation of their use in vitro is the requirement of cofactor for the supply of electrons in the catalytic cycle. Here we report CYP105D1 from Streptomyces griseus expressed in Escherichia coli can be immobilised from cell-free extracts using DE52, that the immobilised protein is active in bioconversions and that a requirement for cofactor can be sustained by a recycling system for NADH regeneration. PMID- 11118350 TI - Proteasome-dependent degradation of cytosolic chaperonin CCT. AB - The chaperonin containing t-complex polypeptide 1 (CCT) is a heterooligomeric molecular chaperone that assists in the folding of actin, tubulin, and other cytosolic proteins. We show here that degradation of CCT in mammalian cells is inhibited by a proteasome-specific inhibitor, lactacystin. When CCT synthesis was inhibited by growth arrest of cells, the decrease in CCT levels was much slower in the presence of lactacystin than in its absence. Pulse-chase experiments indicated that degradation of CCT is inhibited 2- to 2.5-fold by addition of lactacystin. In addition, CCT degradation rate in ts85 cells that produce thermolabile ubiquitin-activating enzyme E1 was reduced 3-fold at the nonpermissive temperature compared to the degradation at the permissive temperature. These results indicate that the ubiquitin-proteasome system is involved in CCT degradation. PMID- 11118351 TI - Visualization of TT virus particles recovered from the sera and feces of infected humans. AB - TT virus (TTV) has not yet been cultured or visualized. We attempted to recover and visualize TTV-associated particles from the serum samples and feces of infected humans. Serum samples were obtained from 7 human immunodeficiency virus (HIV)-infected patients. Three patients had a high TTV DNA titer (10(8) copies/ml), three had a low TTV DNA titer (10(2) copies/ml), and one was negative for TTV DNA. Fecal supernatant was obtained from a different TTV-infected subject. The serum samples were fractionated by high-performance liquid chromatography, and TTV DNA-rich fractions were subjected to floatation ultracentrifugation in cesium chloride. Virus-like particles, 30-32 nm in diameter, were found in the 1.31-1.33 g/cm(3) fractions from each of the three serum samples with high TTV DNA titer, but not in any fraction from the four serum samples that either were negative for TTV DNA or had low TTV DNA titer. The TTV particles formed aggregates of various sizes, and immunogold electron microscopy showed that they were bound to human immunoglobulin G. Similar virus like particles with a diameter of 30-32 nm banding at 1.34-1.35 g/cm(3) were visualized in fecal supernatant with TTV genotype 1a by immune electron microscopy using human plasma containing TTV genotype 1a-specific antibody. PMID- 11118352 TI - Cloning and characterization of a novel nuclear Bcl-2 family protein, zfMcl-1a, in zebrafish embryo. AB - The importance of the Bcl-2 family proteins in normal vertebrate embryogenesis is being recognized; however, their regulatory mechanism is poorly understood. To elucidate the embryonic roles of Bcl-2 family proteins, we cloned and characterized the first zebrafish Bcl-2 family protein, zfMcl-1a. Zebrafish Mcl 1a shows the highest homology to rat Mcl-1 and contains several conserved BH domains of the Bcl-2 family proteins. It also contains a nuclear localization signal (NLS). Using EGFP reporter analysis, we verified the nuclear localization of zfMcl-1a. Deletion of the NLS resulted in distribution of the fusion protein in the cytoplasm. Northern blot analysis indicated that zfMcl-1a mRNA is 1.5 kb and was expressed in oocytes and throughout embryonic development. Notably, the expression of zfMcl-1a transcript was significantly downregulated during gastrulation. These results suggest that zfMcl-1a is a novel nuclear Bcl-2 family protein and is likely to play an important role in zebrafish oogenesis and embryogenesis. PMID- 11118353 TI - p29, a novel GCIP-interacting protein, localizes in the nucleus. AB - GCIP, a newly identified cyclin D-interacting protein, was found to reduce the phosphorylation of retinoblastoma protein and inhibit E2F1-mediated transcriptional activity. To explore more GCIP interacting proteins, the yeast two-hybrid screening using GCIP as a bait protein was performed. One novel gene, p29, was demonstrated to associate with GCIP in the yeast two-hybrid method and in vitro GST pull-down assay. Multiple tissue Northern blot analysis showed that p29 was abundantly expressed in the heart, skeletal muscle, and kidney relative to other tissues. The transient expression of HA-tagged p29 in HeLa cells localized in the nucleus. Taken together, we have isolated a novel protein, p29, which may be involved in the functional regulation of GCIP. PMID- 11118354 TI - Building a better vector: the manipulation of AAV virions. PMID- 11118355 TI - Herpes simplex virus type 1 latency in the murine nervous system is associated with oxidative damage to neurons. AB - The pathological consequences of herpes simplex virus type 1 (HSV-1) latency in the nervous system are not well understood. To determine whether acute and latent HSV-1 infections of the nervous system are associated with oxidative damage, mice were inoculated with HSV-1 by the corneal route, and the extent of viral infection and oxidative damage in trigeminal ganglia and brain was assessed at 7, 90, and 220 days after inoculation. Abundant HSV-1 protein expression in the nervous system was observed in neurons and non-neuronal cells at 7 days after inoculation, consistent with viral replication and spread through the trigeminal and olfactory systems. Acute HSV-1 infection was associated with focal, neuronal and non-neuronal 4-hydroxy-2-nonenal- and 8-hydroxyguanosine-specific immunoreactivity, indicating oxidative damage. Rare HSV-1 antigen-positive cells were observed at 90 and 220 days after inoculation; however, widespread HSV-1 latency-associated transcript expression was detected, consistent with latent HSV 1 infection in the nervous system. HSV-1 latency was detected predominantly in the trigeminal ganglia, brainstem, olfactory bulbs, and temporal cortex. Latent HSV-1 infection was associated with focal chronic inflammation and consistently detectable evidence of oxidative damage involving primarily neurons. These results indicate that both acute and latent HSV-1 infections in the murine nervous system are associated with oxidative damage. PMID- 11118356 TI - Different architectures in the assembly of infectious bursal disease virus capsid proteins expressed in insect cells. AB - Infectious bursal disease virus (IBDV) capsid is formed by the processing of a large polyprotein and subsequent assembly of VPX/VP2 and VP3. To learn more about the processing of the polyprotein and factors affecting the correct assembly of the viral capsid in vitro, different constructs were made using two baculovirus transfer vectors, pFastBac and pAcYM1. Surprisingly, the expression of the capsid proteins gave rise to different types of particles in each system, as observed by electron microscopy and immunofluorescence. FastBac expression led to the production of only rigid tubular structures, similar to those described as type I in viral infection. Western blot analysis revealed that these rigid tubules are formed exclusively by VPX. These tubules revealed a hexagonal arrangement of units that are trimer clustered, similar to those observed in IBDV virions. In contrast, pAcYM1 expression led to the assembly of virus-like particles (VLPs), flexible tubules, and intermediate assembly products formed by icosahedral caps elongated in tubes, suggesting an aberrant morphogenesis. Processing of VPX to VP2 seems to be a crucial requirement for the proper morphogenesis and assembly of IBDV particles. After immunoelectron microscopy, VPX/VP2 was detected on the surface of tubules and VLPs. We also demonstrated that VP3 is found only on the inner surfaces of VLPs and caps of the tubular structures. In summary, assembly of VLPs requires the internal scaffolding of VP3, which seems to induce the closing of the tubular architecture into VLPs and, thereafter, the subsequent processing of VPX to VP2. PMID- 11118357 TI - Genetic diversity of hantaviruses isolated in china and characterization of novel hantaviruses isolated from Niviventer confucianus and Rattus rattus. AB - The antigenic and genetic properties of 46 hantaviruses from China, 13 from patients, 23 from rodents, and 10 from unknown hosts, were compared with those of other hantaviruses. The viruses were classified as either Hantaan (HTN) or Seoul (SEO) viruses. A phylogenetic analysis of the partial M (300 bp) and S (around 485 bp) genomes of HTN viruses identified nine distinct genetic subtypes, one consisting of isolates from Korea. The SEO viruses were divided into five genetic subtypes, although they had less variability than the HTN subtypes. There was a correlation between the subtype and province of origin for four subtypes of HTN viruses, confirming geographical clustering. Hantaan virus NC167 isolated from Niviventer confucianus and SEO virus Gou3 isolated from Rattus rattus were the basal clades in each virus. The phylogenetic trees constructed from the entire S and M segments suggested that NC167 was introduced to N. confucianus in a host switching event. The reactivity of a panel of 35 monoclonal antibodies was almost exactly the same in NC167 and a representative HTN virus and in Gou3 and a representative SEO virus. However, there was a one-way cross-neutralization between them. These results confirm the varied nature of Murinae-associated hantaviruses in China. PMID- 11118358 TI - Differential expression of TGF-beta, IL-2, and other cytokines in the CNS of Theiler's murine encephalomyelitis virus-infected susceptible and resistant strains of mice. AB - Intracranial inoculation of susceptible SJL mice with Theiler's murine encephalomyelitis virus (TMEV) results in biphasic disease consisting of early acute disease, followed by late chronic demyelinating disease, associated with mononuclear infiltrates and demyelinating lesions. In contrast, resistant C57BL/6 (B6) mice develop only early acute disease. We employed cytokine-specific RT-PCR to determine the expression of cytokine transcripts in the CNS of TMEV-infected SJL and B6 mice. During early acute disease, we have found a strong proinflammatory (Th1) cytokine response in the CNS of both TMEV-infected SJL and B6 mice, demonstrated by the expression of transcripts for IFN-gamma, IL-1, IL-6, IL-12p40, and TNF-alpha. At 8 days postinfection (p.i.), TGF-beta1 and TNF-alpha transcripts were present at significantly higher levels (P < 0.01) in the CNS of SJL susceptible mice in comparison to those found in the CNS of B6 mice. Immunohistochemical staining revealed that TGF-beta protein was expressed in leptomeningeal mononuclear inflammatory cell infiltrates in the brain of SJL mice but not in B6 mice, at 8 days p.i. TGF-beta may be responsible for the failure of SJL mice to develop an effective anti-TMEV CTL response. During late chronic demyelinating disease, high levels of proinflammatory Th1 cytokines were found in the CNS of SJL mice, but not B6 mice. Significantly higher levels (P < 0.01) of anti-inflammatory cytokine transcripts (IL-4, IL-5, and IL-10 (Th2 cytokines) and TGF-beta) were found in the spinal cord of TMEV-infected SJL mice with chronic demyelinating disease than in the spinal cord of B6 mice during the same time period (39 or 60 days p.i.). These anti-inflammatory cytokines may contribute to the downregulation of the proinflammatory response in SJL mice. High levels of IL 2 transcripts and protein appeared transiently in the spinal cord of TMEV infected SJL mice before the onset of demyelinating disease and coincided with an influx of new T cells into the CNS and/or expansion of remaining T cells that have not been eliminated after viral clearance. PMID- 11118359 TI - Different patterns of restriction to B19 parvovirus replication in human blast cell lines. AB - B19 parvovirus can replicate in erythroid progenitor cells and in a small number of human blast cell lines. To better understand and analyze the B19 virus replicative cycle, we performed and compared the infection of bone marrow cells and of different blast cell lines with erythroblastoid and megakaryoblastoid phenotypic characteristics (UT-7, TF-1, M-07, and B1647). Following in vitro infection, B19-specific nucleic acids were characterized with regard to the genome-replicative intermediates, the transcription pattern, and the localization of virus-specific nucleic acids inside infected cells. While all cell lines tested proved to be susceptible to B19 virus infection, two different patterns of restriction to replication of B19 virus were observed. In the first restriction pattern, observed in UT-7 cells, the single-stranded viral DNA was converted to double-stranded replicative intermediates, identical to those found in bone marrow cells, and a full set of viral transcripts were observed. However, replication and transcription were restricted to a small subset of cells, and production of capsid proteins was not detected. In the second restriction pattern, observed in TF-1, M-07, and B1647 cells, the single-stranded viral DNA was not converted to double-stranded replicative intermediates. PMID- 11118360 TI - Alleviation of murine leukemia virus repression in embryonic carcinoma cells by genetically engineered primer binding sites and artificial tRNA primers. AB - The primer binding site (PBS) plays pivotal roles during reverse transcription of retroviruses and also is the target of a cellular host defense impeding the transcription of murine leukemia virus (MLV) harboring a proline (pro) PBS in embryonic cells. Both the PBS and the tRNA primer are copied during reverse transcription and anneal as complementary DNA sequences creating the PBS of the integrated provirus. The pro PBS of MLV can be exchanged by PBS sequences matching endogenous or engineered tRNAs to allow replication of Akv MLV-derived vectors in fibroblasts. Here we use the PBS escape mutant B2 to demonstrate the capacity of the synthetic tRNA(B2) to function in reverse transcription in competition with endogenous tRNAs in fibroblasts and embryonic carcinoma (EC) cells. We further show symmetry between PBS and the primer by the ability of the synthetic tRNA(B2) to confer escape from EC repression of a PBS-Pro vector. Of a panel of vectors with the repressed pro PBS substituted for other natural or artificial PBS sequences, all except one efficiently expressed the neo marker gene when transferred to NIH/3T3 and EC cells, hence avoiding PBS-mediated silencing in EC cells. A non-natural PBS matching an artificially designed tRNA molecule conferred no further relief from repression than that attained with the B2 escape mutant or the natural alternative PBSs. Interestingly, a vector harboring a PBS matching tRNA(Lys1.2) suffered repression similar to the wild type PBS-Pro but was partially rescued by a single point mutation of the PBS. PMID- 11118361 TI - Expression of a foreign epitope by porcine reproductive and respiratory syndrome virus. AB - The potential of porcine reproductive and respiratory syndrome virus (PRRSV) as a viral vector was explored by the insertion of a sequence encoding a foreign antigen into the infectious cDNA clone of the Lelystad virus isolate. An epitope of the hemagglutinin (HA) protein of human influenza A virus was introduced at the 5' end and at the 3' end of ORF7, in each case resulting in a fusion protein between the HA epitope and the nucleocapsid (N) protein. Furthermore, in the construct carrying the HA sequences at the 5' end of ORF7, additional in-frame insertions encoding the autoprotease 2A of foot-and-mouth disease virus were made between the HA and ORF7 sequences to ensure the generation of a functional N protein from its hybrid precursor. When RNA transcripts from these full-length cDNA clones were transfected into BHK-21 cells, they were each found to replicate, to express the HA epitope, and to produce progeny virus. However, fusion of the HA epitope directly to the nucleocapsid protein either at the N terminus or at the C terminus adversely affected both the viability and the genetic stability of the recombinant PRRS viruses. Serial passage of the recombinant viruses on porcine alveolar macrophages demonstrated that these viruses had lost (part of) the HA epitope at passage four. In contrast, in the PRRS viruses expressing the HA epitope from a precursor cleavable by the autoprotease 2A peptide, the HA epitope was still intact after four passages, and no effect on the viability of these viruses was observed. Immunoprecipitation and pulse chase experiments revealed the efficient and presumably cotranslational cleavage of the HA epitope from the N protein by the 2A protease. Our results demonstrate the feasibility of using PRRSV as a viral vector that might be suitable for the delivery of antigens from other pathogens to the immune system of the pig. PMID- 11118362 TI - Molecular cloning and biological characterization of full-length HIV-1 subtype C from Botswana. AB - Human immunodeficiency virus type 1 (HIV-1) subtype C is now responsible for more than half of all HIV-1 infections in the global epidemic and for the high levels of HIV-1 prevalence in southern Africa. To facilitate studies of the biological nature and the underlying molecular determinants of this virus, we constructed eight full-length proviral clones from two asymptomatic and three AIDS patients infected with HIV-1 subtype C from Botswana. Analysis of viral lysates showed that Gag, Pol, and Env structural proteins were present in the virions. In four clones, the analysis suggested inefficient envelope glycoprotein processing. Nucleotide sequence analysis of the eight clones did not reveal frameshifts, deletions, premature truncations, or translational stop codons in any structural, regulatory, or accessory genes. None of the subtype C clones were replication competent in donor peripheral blood mononuclear cells (PBMCs), macrophages, Jurkat(tat) cells, or U87. CD4.CCR5 cells. However, infection by two clones could be rescued by complementation with a functional subtype C envelope clone, resulting in a productive infection of PBMCs, macrophages, and U87. CD4.CCR5 cells. PMID- 11118363 TI - Induction of immune responses and break of tolerance by DNA against the HIV-1 coreceptor CCR5 but no protection from SIVsm challenge. AB - An inactivating mutation in the human CCR5 gene reduces the risk of HIV-1 infection in individuals with homozygous alleles. We explored whether genetic immunization would induce an immune response directed to CCR5 structures and if immunological tolerance toward endogenous CCR5 could be broken. We also studied whether this immunization approach could protect cynomolgus monkeys from an infection, with SIVsm, which primarily uses CCR5 as a coreceptor. Epidermal but not intramuscular delivery of the CCR5 gene to mice elicited strong IgG antibody binding responses to CCR5. Intramucosal immunization of cynomolgus macaques with CCR5 DNA followed by boosts with CCR5 peptides induced prominent IgG and IgA antibody responses in serum and vaginal washings. The CCR5-specific antibodies neutralized the infectivity of primary human R5 HIV-1 strains, and the macaque SIVsm but not that of a tissue culture-adapted X4 HIV-1 strain. The consecutive CCR5 gene and CCR5 peptide immunizations induced B- and T-cell responses to peptides representing both human and macaque amino acid sequences of the respective CCR5 proteins. This indicates that tolerance was broken against endogenous macaque CCR5, which has a 98% homology to the human CCR5 gene. After the final boost, the vaccinated monkeys together with two control monkeys were challenged with SIVsm. Neither protection against nor enhancement of SIVsm infection was achieved. PMID- 11118364 TI - Soluble glycosaminoglycans Do not potentiate RANTES antiviral activity on the infection of primary macrophages by human immunodeficiency virus type 1. AB - Macrophages play an important role in human immunodeficiency virus (HIV)-1 infection. They exist in various differentiation and activation states in vivo, a heterogeneity that may affect their interactions with HIV-1 and susceptibility to drugs. Here, we found that RANTES and MIP-1beta, heparin, or soluble chondroitin sulfate B, but not chondroitin sulfate A, inhibited HIV-1(BaL) infection of macrophages obtained as the adherent cells of 5-day cultures of blood mononuclear cells (PBMC), followed by 2 days without either nonadherent PBMC or added cytokines (MDM-5d), whereas they did not affect infection of macrophages obtained as the adherent cells recovered from 1-h incubation of PBMC and subsequent 7-day culture with macrophage colony-stimulating factor (MDM-MCSF). Such different behavior was not related to differences in HIV-1 binding but rather to postbinding steps, as HIV-1(BaL) attached similarly to MDM-5d and MDM-MCSF, a binding that was affected by soluble glycosaminoglycans but not by RANTES. Of note, CCR5 expression on both types of MDM was comparable, and it was not downregulated by RANTES on either. Mixing RANTES with each of the glycosaminoglycans did not restore inhibition of MDM-MCSF infection by HIV-1; however, heparin at concentrations that had low antiviral activity for MDM-5d counteracted RANTES anti-HIV-1 activity for these cells, whereas chondroitin sulfate B had no additive effect on that of RANTES. Both glycosaminoglycans affected RANTES binding to MDM. Thus, in contrast to cell surface proteoglycans that contribute to the attachment of RANTES to macrophages and enhance its anti HIV-1 activity, soluble glycosaminoglycans do not facilitate, and may even offset, the anti-HIV-1 activity of RANTES. PMID- 11118365 TI - Two influenza A virus-specific Fabs neutralize by inhibiting virus attachment to target cells, while neutralization by their IgGs is complex and occurs simultaneously through fusion inhibition and attachment inhibition. AB - Mabs H36 (IgG2a) and H37 (IgG3) recognize epitopes in antigenic sites Sb and Ca2, respectively, in the HA1 subunit of influenza virus A/PR/8/34 (H1N1). Their neutralization was complex. Our aim here was to investigate the mechanism of neutralization by the IgGs and their Fabs. In MDCK and BHK cells, both IgGs neutralized primarily by inhibiting virus-cell fusion, although at higher IgG concentrations virus attachment to target cells was also inhibited. In contrast, the Fabs neutralized entirely by inhibiting virus attachment, although a higher concentration of Fab than IgG was required to bring this about. Both H36 and H37 exerted a concentration-dependent spectrum of neutralization activity, with virus cell fusion inhibition and virus-cell attachment inhibition being the predominant mechanisms at low- and high-antibody concentration, respectively, and both mechanisms occurring simultaneously at intermediate concentrations. However, it may be that attachment inhibition was a secondary event, occurring to virus that had already been neutralized through inhibition of its fusion activity. Neutralization by H36 and H37 Fabs was a simple process. Both inhibited virus attachment but required much higher (>100-fold) molar concentrations for activity than did IgG. The functional affinities of the IgGs were high (0.4-0.6 nM) and differences between these and the affinity of their Fabs (H36, nil; H37, 23-fold) were not sufficient to explain the differences observed in neutralization. Similar neutralization data were obtained in two different cell lines. The dose response curve for neutralization by H36 F(ab')(2) resembled that for IgG, although eightfold more F(ab')(2) was required for 50% neutralization. Overall, neutralization mechanisms of H36 and H37 antibodies were similar, and thus independent of antigenic site, antibody isotype, and target cell. PMID- 11118366 TI - Molecular characterization of serotype G9 rotavirus strains from a global collection. AB - Between 1992 and 1998, serotype G9 human rotavirus (RV) strains have been detected in 10 countries, including Thailand, India, Brazil, Bangladesh, Malawi, Italy, France, the United States, the United Kingdom, and Australia, suggesting the possible emergence of the fifth common serotype worldwide. Unlike the previously characterized reference G9 strains (i.e., WI61 and F45), the recent G9 isolates had a variety of gene combinations, raising questions concerning their origin and evolution. To identify the progenitor strain and examine the on-going evolution of the recent G9 strains, we characterized by genetic and antigenic analyses 16 isolates obtained from children with diarrhea in India, Bangladesh, the United States, and Malawi. Specifically, we sequenced their VP7 and NSP4 genes and compared the nucleotide (nt) and deduced amino acid sequences with the reference G9 strains. To identify reassortment, we examined the products of five gene segments; VP4, VP7, and NSP4 genotypes (genes 4, 9, and 10); subgroups (gene 6); electropherotypes (gene 11); and the genogroup profiles of all of the recent G9 isolates. Sequence analysis of the VP7 gene indicated that the recent U.S. P[6],G9 strains were closely related to the Malawian G9 strains (>99% nt identity) but distinct from G9 strains of India ( approximately 97% nt identity), Bangladesh ( approximately 98% nt identity), and the reference strains ( approximately 97% nt identity). Phylogenetic analysis identified a single cluster for the U.S. P[6],G9 strains that may have common progenitors with Malawian P[6],G9 strains whereas separate lineages were defined for the Indian, Bangladeshi, and reference G9 strains. Northern hybridization results indicated that all 11 gene segments of the Malawian P[6],G9 strains hybridized with a probe derived from a U.S. strain of the same genotype and may have the same progenitor, different from the Indian G9 strains, whereas the Bangladesh strains may have evolved from the U.S. G9 progenitors. Overall, our findings suggest that much greater diversity among the newly identified G9 strains has been generated by reassortment between gene segments than through the accumulation of mutations in a single gene. PMID- 11118367 TI - Herpesvirus saimiri pathogenicity enhanced by thymidine kinase of herpes simplex virus. AB - Herpesvirus saimiri can be used as an efficient gene expression vector for human T lymphocytes and thus may allow applications in experimental leukemia therapy. We constructed recombinant viruses for the functional expression of the thymidine kinase (TK) of herpes simplex virus type 1 (HSV) as a suicide gene. These viruses reliably allowed the targeted elimination of transduced nonpermissive human T cells in vitro after the administration of ganciclovir. To test the reliability of this function under the most stringent permissive conditions, in this study we analyzed the influence of the prodrugs ganciclovir and acyclovir in common marmosets on the acute leukemogenesis induced by either wild-type herpesvirus saimiri C488 or by a recombinant derivative expressing TK of HSV. Antiviral drug treatment did not influence the rapid development of acute disease. In contrast, the presence of the HSV tk gene resulted in a faster disease progression. In addition, HSV TK-expressing viruses showed faster replication than wild-type virus in culture at low serum concentrations. Thus, HSV TK accelerates the replication of herpesvirus saimiri and enhances its pathogenicity. This should be generally considered when HSV TK is applied as a transgene in replication competent DNA virus vectors for gene therapy. PMID- 11118368 TI - Insertion of cellular NEDD8 coding sequences in a pestivirus. AB - For the cytopathogenic (cp) bovine viral diarrhea virus (BVDV) strain CP 821, a duplication of the genomic region encoding part of NS2, NS3, NS4A, and part of NS4B together with a nonviral insertion was detected. Further analyses including molecular cloning and sequencing of the putative cellular recombination partner showed that the insertion in CP 821 originated from a bovine mRNA encoding the cellular protein NEDD8, which is 58% identical to ubiquitin. To our knowledge the genome of CP 821 represents the first viral RNA with a NEDD8 coding insertion. Remarkably, the insertion site differs from that described for insertions of ubiquitin. The NEDD8 sequence allows an additional cleavage of the viral polyprotein, whereby an NS3 with an unusual N-terminus is generated. Furthermore, the CP 821-specific genomic alterations were introduced into an infectious noncytopathogenic (noncp) BVDV cDNA clone. After transfection of bovine cells with the respective RNA, a cp virus was recovered. This showed that the NEDD8 coding insertion together with the duplicated viral sequences represents the genetic basis for cytopathogenicity of CP 821. In addition to the recovered cp virus, noncp BVDV rapidly evolved after transfection. This is the first time that a change from the cp to the noncp phenotype was demonstrated in the course of replication in tissue culture cells. PMID- 11118369 TI - Latency-associated sense transcripts are expressed during in vitro human cytomegalovirus productive infection. AB - Published studies have identified novel sense transcripts in latent human cytomegalovirus (HCMV) infection. These cytomegalovirus latency-associated transcripts (CLTs) have start sites upstream from the MIE gene productive start site (PSS). We evaluated the expression of the sense CLTs in an in vitro HCMV productive infection. Transcripts with initiation sites upstream from the PSS were detected in infected human fibroblasts using reverse transcription polymerase chain reaction and 5' rapid amplification of cDNA ends (RACE) analysis. DNA sequencing of 5' RACE PCR products confirmed that the start sites were consistent with sense CLT expression. Furthermore, ribonuclease protection assay analysis showed that transcripts initiating at the latent start site-1 were most abundant at late times postinfection after transcription from the PSS had decreased. In addition, transcription consistent with sense CLT expression was identified in HCMV-infected dTHP-1 cells, monocyte-derived macrophages, and endothelial cells, as well as in clinical isolate-infected human fibroblasts. These findings clearly demonstrate the expression of sense CLTs during in vitro HCMV infection. PMID- 11118370 TI - Equine herpesvirus 1 (EHV-1) glycoprotein M: effect of deletions of transmembrane domains. AB - Equine herpesvirus 1 (EHV-1) recombinants that carry either a deletion of glycoprotein M (gM) or express mutant forms of gM were constructed. The recombinants were derived from strain Kentucky A (KyA), which also lacks genes encoding gE and gI. Plaques on RK13 cells induced by the gM-negative KyA were reduced in size by 80%, but plaque sizes were restored to wild-type levels on gM expressing cells. Electron microscopic studies revealed a massive defect in virus release after the deletion of gM in the gE- and gI-negative KyA, which was caused by a block in secondary envelopment of virions at Golgi vesicles. Recombinant KyA expressing mutant gM with deletions of predicted transmembrane domains was generated and characterized. It was shown that mutant gM was expressed and formed dimeric and oligomeric structures. However, subcellular localization of mutant gM proteins differed from that of wild-type gM. Mutant glycoproteins were not transported to the Golgi network and consequently were not incorporated into the envelope of extracellular virions. Also, a small plaque phenotype of mutant viruses that was indistinguishable from that of the gM-negative KyA was observed. Plaque sizes of mutant viruses were restored to wild-type levels by plating onto RK13 cells constitutively expressing full-length EHV-1 gM, indicating that mutant proteins did not exert a transdominant negative effect on wild-type gM. PMID- 11118371 TI - Complete nucleotide sequence of the chiba virus genome and functional expression of the 3C-like protease in Escherichia coli. AB - We cloned the genome RNA of the Chiba virus (ChV; Hu/NLV/Chiba 407/1987/JP) and determined its complete nucleotide sequence. The genome is predicted to be a positive-sense, single-stranded RNA of 7697 bases, excluding a poly(A) tract. Comparison of the nucleotide and amino acid sequences with those of other members of the species Norwalk virus (NV) revealed that ChV belongs to genogroup I NV. The ChV genome contains three open reading frames (ORFs). A large 5'-terminal ORF (ORF1) encodes a polyprotein with 1785 amino acids that are likely processed into functional proteins, including RNA helicase, VPg, protease, and RNA-dependent RNA polymerase. ORF2 encodes the capsid protein with 544 amino acids, and a small 3' terminal ORF (ORF3) encodes a basic protein with 208 amino acids. The amino acid sequences of five cleavage sites in ORF1 are highly conserved compared with those of other members of NV. When expressed in Escherichia coli, the glutathione-S transferase (GST) fusion protein of the ChV protease connected via a short peptide containing a human rhinovirus 3C protease cleavage site was cleaved into GST and the protease; however, this cleavage did not occur when the Cys mutation was introduced into the putative active site of the protease. Moreover, the ChV protease recognized and cleaved the predicted proteolytic sites between VPg and protease and between protease and RNA polymerase. Therefore, the ChV protease expressed in E. coli retained an enzymatic activity and a substrate specificity similar to that of the human rhinovirus 3C protease. PMID- 11118372 TI - Ser(2194) is a highly conserved major phosphorylation site of the hepatitis C virus nonstructural protein NS5A. AB - Phosphorylation of the nonstructural NS5A protein is highly conserved among hepatitis C virus (HCV) genotypes. However, the precise site or sites of phosphorylation of NS5A have not been determined, and the functional significance of phosphorylation remains unknown. Here, we showed by two-dimensional phosphopeptide mapping that a protein kinase or kinases present in yeast, insect, and mammalian cells phosphorylated a highly purified HCV genotype 1b NS5A from insect cells on identical serine residues. We identified a major phosphopeptide (corresponding to amino acids 2193-2212 of the HCV 1b polyprotein) by using negative-ion electrospray ionization-microcapillary high performance liquid chromatography-mass spectrometry. The elution time of the phosphopeptide determined by negative-ion electrospray ionization-mass spectrometry corresponded with the elution time of the majority of (32)P-label that was incorporated into the phosphopeptide by an in vitro kinase reaction. Subsequent analysis of the peak fraction by automated positive-ion electrospray ionization-tandem mass spectrometry revealed that Ser(2194) was the major phosphorylated residue on the phosphopeptide GpSPPSLASSSASQLSAPSLK. Substitution for Ser(2194) with Ala resulted in the concomitant disappearance of major in vivo phosphorylated peptides. Ser(2194) and surrounding amino acids are highly conserved in all HCV genotypes, suggesting NS5A phosphorylation at Ser(2194) may be an important mechanism for modulating NS5A biological functions. PMID- 11118373 TI - In vitro induction of HIV-1 replication in resting CD4(+) T cells derived from individuals with undetectable plasma viremia upon stimulation with human T-cell leukemia virus type I. AB - Microbial coinfections have been associated with transient bursts of human immunodeficiency virus (HIV) viremia in patients. In this study we investigated whether human T-cell leukemia virus type I (HTLV-I), another human retrovirus that is prevalent among certain HIV-infected populations, can induce HIV-1 replication in patients who had been successfully treated with highly active antiretroviral therapy. We demonstrate that supernatants from HTLV-I-producing MT 2 cells can induce in vitro replication of HIV-1 from highly purified, resting CD4(+) T cells obtained from individuals with undetectable plasma viremia. Depletion of proinflammatory cytokines from the supernatants reduced, but did not abrogate, the ability to induce HIV-1 replication, indicating that other factors such as HTLV-I Tax or Env also have a role. The HTLV-I-mediated effect does not require productive infection: exposure to heat-inactivated HTLV-I virions, purified Tax protein, or HTLV-I Env glycoprotein also induced expression of HIV 1. Furthermore, we demonstrate that coculture of resting CD4(+) T cells with autologous CD8(+) T cells markedly inhibits the HTLV-I-induced virus replication. Our results suggest that coinfection with HTLV-I may induce viral replication in the latent viral reservoirs; however, CD8(+) T cells may play an important role in controlling the spread of virus upon microbial stimulation. PMID- 11118374 TI - Role of M protein aggregation in defective assembly of temperature-sensitive M protein mutants of vesicular stomatitis virus. AB - The goal of these experiments was to determine the steps in virus assembly that are defective at the nonpermissive temperature in temperature-sensitive (ts) matrix (M) protein mutants of vesicular stomatitis virus. It has been proposed that mutations in M protein either reduce the binding affinity for nucleocapsids or lead to aggregation, reducing the amount of M protein available for virus assembly. Cytosolic or membrane-derived M proteins from wild-type VSV and two ts M protein mutant viruses, tsM301 and tsO23, as well as a revertant of tsO23 virus, O23R1, were analyzed for binding to nucleocapsid-M protein (NCM) complexes and for M protein aggregation. The experiments presented here showed that ts M proteins synthesized at the nonpermissive temperature were capable of binding to nucleocapsids and that aggregation of ts M proteins did not reduce the amount of soluble M protein below the amount required for assembly of the O23R1 virus. Instead, the most pronounced defect in ts M proteins was in the ability of membrane-derived M proteins to be solubilized in the presence of the detergent Triton X-100. It is proposed that this detergent-insoluble form of M protein interferes with a step necessary to initiate assembly of NCM complexes. A similar detergent, Triton X-114, caused aggregation of membrane-derived wild-type M protein, disproving an earlier proposal that membrane-derived M protein behaves like an integral membrane protein in the presence of Triton X-114. Aggregation of wild-type M protein in the presence of Triton X-100 could be induced by incubation at 37 degrees C with a high-molecular-weight fraction isolated from uninfected cells by sucrose gradient centrifugation. These results implicate host components in inducing M protein aggregation. PMID- 11118375 TI - Herpes simplex virus infection induces replication of human immunodeficiency virus type 1. AB - Genital herpes has been associated with increased efficiency of the sexual transmission and enhanced replication of human immunodeficiency virus type 1 (HIV 1). In this study we demonstrate that exposure to infectious or heat-inactivated herpes simplex virus (HSV) type 1 or 2 virions increases HIV-1 expression in macrophages at least in part by inducing NF-kappaB activity. Neutralizing antibodies to the HSV glycoprotein gB or gD markedly attenuated these virion mediated effects on HIV-1 expression in macrophages. Thus HSV infection of macrophages that reside in genital mucosal tissue induces HIV-1 replication in these cells. Our study may have implications for the management of patients who are coinfected with the two viruses. PMID- 11118376 TI - Expression and folding of human very-low-density lipoprotein receptor fragments: neutralization capacity toward human rhinovirus HRV2. AB - Minor group human rhinoviruses (HRVs) use members of the low-density lipoprotein receptor family for cell entry. To investigate the utility of receptor fragments as viral inhibitors, various polypeptide segments derived from the ligand binding domain of human very-low-density lipoprotein receptor (VLDLR) were expressed in a soluble form in bacteria. Whereas none of the fragments was active in virus binding immediately after recovery from the cell lysates, constructs encompassing complement type repeats 1-3, 1-6, and 1-8 spontaneously acquired virus binding activity by incubation at 4 degrees C in buffer containing Ca(2+) ions and lacking any redox system. When immobilized receptor-associated protein (RAP), a specific chaperone for VLDLR, was present during the incubation, the yield of protein active in ligand binding was substantially increased. A VLDLR fragment with repeats 4-6 failed to bind virus; however, it bound RAP. Bacterial expression of truncated VLDLR 1-3 at high yield, easy purification, and folding together with high inhibitory activity toward HRV2 makes this protein a promising starting point for the development of an oligopeptide-based antiviral agent. Using sucrose density gradient centrifugation, we demonstrate the formation of virus-receptor complexes. The recombinant receptors can thus be used for structure determination by electron cryo-microscopy. PMID- 11118377 TI - Temporal loss of Nef-epitope CTL recognition following macaque lipopeptide immunization and SIV challenge. AB - To address the subtle interactions between antiviral cytotoxic T-cell (CTL) immune responses and the evolution of viral quasispecies variants in vivo, we performed a longitudinal study in a simian immunodeficiency virus (SIV)-infected rhesus macaque that had a long experimental SIV infection before developing simian AIDS. Before being infected with SIV, this animal was immunized with a mixture of seven lipopeptides derived from SIV Nef and Gag proteins and showed a bispecific antiviral CTL response directed toward Nef 169-178 and 211-225 peptides. After SIV infection, CTL activity against the Nef 169-178 epitope was no longer detectable, as assessed from peripheral blood mononuclear cells stimulated by autologous SIV. CTL activity against the 211-225 epitope was lost after 3 months, and an additional CTL response to the amino acids 112-119 Nef epitope emerged. Analysis of the Nef proviral sequence revealed the presence of immune escape variants first in the 211-225 epitope and much later in the 112-119 epitope. In contrast, epitope 169-178 showed only two mutations among all viral sequencing performed. We conclude that in this macaque, bispecific CTL exerted a strong selective pressure and escape virus mutants finally emerged. We identified CTL recognizing a conserved Nef epitope 112-119 (SYKLAIDM), essential for viral replication, which could be associated with a prolonged AIDS-free period. These results stress the importance of the induction of broader multispecific CTLs directed against highly conserved and functional T-cell epitopes by vaccination, with the aim of keeping HIV infection in check. PMID- 11118378 TI - In vitro concatemer formation catalyzed by vaccinia virus DNA polymerase. AB - During poxvirus infection, both viral genomes and transfected DNAs are converted into high-molecular-weight concatemers by the replicative machinery. However, aside from the fact that concatemer formation coincides with viral replication, the mechanism and protein(s) catalyzing the reaction are unknown. Here we show that vaccinia virus DNA polymerase can catalyze single-stranded annealing reactions in vitro, converting linear duplex substrates into linear or circular concatemers, in a manner directed by sequences located at the DNA ends. The reaction required > or =12 bp of shared sequence and was stimulated by vaccinia single-stranded DNA-binding protein (gpI3L). Varying the structures at the cleaved ends of the molecules had no effect on efficiency. These duplex-joining reactions are dependent on nucleolytic processing of the molecules by the 3'-to 5' proofreading exonuclease, as judged by the fact that only a 5'-(32)P-end label is retained in the joint molecules and the reaction is inhibited by dNTPs. The resulting concatemers are joined only through noncovalent bonds, but can be processed into stable molecules in E. coli, if the homologies permit formation of circular molecules. This reaction provides a starting point for investigating the mechanism of viral concatemer formation and can be used to clone PCR-amplified DNA. PMID- 11118379 TI - Neutralization of human papillomavirus (HPV) pseudovirions: a novel and efficient approach to detect and characterize HPV neutralizing antibodies. AB - The development of vaccines against human papillomaviruses (HPVs) has long been hampered by the inability to grow HPVs in tissue culture and the lack of an efficient neutralization assay. To date, less than 10% of more than 100 different HPV types can be grown in athymic and "SCID" mouse xenograft systems or raft culture systems. Recently, the in vitro generation of HPV pseudovirions and their use in neutralization assays were demonstrated. The major shortcomings of the current approaches to HPV neutralization are the lack of HPV virions for most types for the xenograft methods and the time-consuming and inefficient generation of infective pseudovirions for the latter methods, which precludes their use in large-scale HPV clinical trials or epidemiological studies. We describe here a novel and efficient approach to generating pseudovirions in which HPV virus-like particles (VLPs) are coupled to the beta-lactamase gene as a reporter. We show that it is not necessary to encapsidate the reporter gene constructs into the pseudovirions. Using sera from human volunteers immunized with HPV-11 VLPs expressed in yeast, we demonstrate that our novel neutralization assay compares favorably with the athymic mouse neutralization assay. Furthermore, our assay was used to define neutralizing monoclonal antibodies to HPV-6, which were previously unknown. PMID- 11118380 TI - Recombinant hepatitis delta antigen from E. coli promotes hepatitis delta virus RNA replication only from the genomic strand but not the antigenomic strand. AB - Hepatitis delta antigen (HDAg) of hepatitis delta virus (HDV) typically consists of two related protein species. The small HDAg (S-HDAg) is a 24-kDa protein of 195 amino acids and the large HDAg (L-HDAg) is a 27-kDa protein with an additional 19 amino acids at its C-terminus. These two proteins have distinct functions in the HDV life cycle. We have developed conditions for expressing S HDAg and L-HDAg in E. coli as soluble proteins to facilitate large-scale purification. These proteins were purified to homogeneity and shown to be biologically active. Transfection of the purified recombinant S-HDAg together with HDV genomic RNA resulted in viral RNA replication. Surprisingly, the purified S-HDAg could not initiate replication from the antigenomic-sense HDV RNA, even though the latter led to RNA replication when transfected with an mRNA encoding the S-HDAg. These results suggest that initiation of HDV RNA synthesis from the antigenomic RNA may require a form of HDAg that is modified in mammalian cells; in contrast, RNA synthesis from the genomic RNA could be initiated by the recombinant S-HDAg from E. coli. Interestingly, the purified L-HDAg appeared as multiple protein species, including one corresponding to S-HDAg, probably as a result of degradation. The partially proteolyzed L-HDAg also initiated HDV RNA replication under the same conditions. These results add to the mounting evidence that genomic- and antigenomic-strand HDV RNA syntheses are carried out by different mechanisms. PMID- 11118381 TI - Change in receptor-binding specificity of recent human influenza A viruses (H3N2): a single amino acid change in hemagglutinin altered its recognition of sialyloligosaccharides. AB - Human H3N2 influenza A viruses were known to preferentially bind to sialic acid (SA) in alpha2,6Gal linkage on red blood cells (RBC). However, H3N2 viruses isolated in MDCK cells after 1992 did not agglutinate chicken RBC (CRBC). Experiments with point-mutated hemagglutinin (HA) of A/Aichi/51/92, one of these viruses, revealed that an amino acid change from Glu to Asp at position 190 (E190D) was responsible for the loss of ability to bind to CRBC. A/Aichi/51/92 did not agglutinate CRBC treated with alpha2, 3-sialidase, suggesting that SAalpha2,3Gal on CRBC might not inhibit the binding of the virus to SAalpha2,6Gal on CRBC. However, the virus agglutinated derivatized CRBC resialylated with SAalpha2, 6Galbeta1,4GlcNAc. These findings suggested that the E190D change might have rendered the HA able to distinguish sialyloligosaccharides on the derivatized CRBC containing the SAalpha2,6Galbeta1,4GlcNAc sequence from those on the native CRBC. PMID- 11118382 TI - Differential budding efficiencies of human T-cell leukemia virus type I (HTLV-I) Gag and Gag-Pro polyproteins from insect and mammalian cells. AB - In this study, we examined the ability of human T-cell leukemia virus type I (HTLV-I) Gag and Gag-Pro to assemble immature virus-like particles (VLPs) and bud from insect and mammalian cells. Transmission electron microscopy of insect cells infected with a recombinant baculovirus carrying the entire gag gene revealed that Pr53(Gag) is targeted to the plasma membrane, where it extensively accumulates and forms electron-dense evaginations. However, no particles could be detected either inside the cells or in the culture supernatants. With the Gag-Pro expressing construct, we observed HTLV-I-specific cytoplasmic proteolysis of the Gag precursor, but again no particle released in the culture supernatants. Transmission electron microscopic analysis of insect cells expressing Gag-Pro polyprotein revealed large vacuoles in the cytoplasm and no budding particles at the plasma membrane. In contrast, human immunodeficiency virus type 1 Gag polyprotein expressed in insect cells is able to release VLPs. These data showed that unlike other retroviruses, Pr53(Gag) is unable to be released as immature VLPs from insect cells. To determine whether the block in particle budding and release is due to an intrinsic property of Pr53(Gag) or the absence of essential cellular factors in insect cells, we expressed Gag and Gag-Pro polyproteins in human 293 cells. The results indicate that Pr53(Gag) and p24 capsid are released within particles into the culture supernatants of human 293 cells. We found that the myristylation of the N-terminal glycine residue is essential for Gag release. Altogether, these results strongly suggest that the proper assembly of HTLV-I particles is dependent on mammalian host cell factors. PMID- 11118383 TI - Where Have All the Case Reports Gone? PMID- 11118384 TI - Antibodies to Haemophilus influenzae serotype b in the Netherlands a few years after the introduction of routine vaccination. AB - We assessed antibodies to the capsular polysaccharide of Haemophilus influenzae type b (HibPS) in the Dutch population a few years after a mass vaccination against H. influenzae (Hib) was begun. We observed sharp declines in the geometric mean titer (GMT) and the prevalence of HibPS antibodies at levels of < or =0.15 microg/mL in children who had received 4 doses of vaccine: from 8.65 microg/mL (prevalence, 99.4%) after 0-2 months to 0.8 microg/mL (prevalence, 83.3%) after 27-29 months. In adult groups, both the prevalence of HibPS antibodies and the GMT declined significantly with increasing age but remained high (prevalence, > or =83.7%; GMT, 0.73 > or = microg/mL). We conclude that the overall immunity in the Dutch population seems satisfactory. We draw our conclusions from the current serosurveillance study and from the sharp decline in invasive Hib disease noted after the introduction of vaccination. The key questions for the future are (1) whether Hib and cross-reacting organisms will circulate sufficiently to provide natural reexposure, and (2) how long memory immunity will persist after vaccination without reexposure. PMID- 11118385 TI - Adjunctive salvage therapy with inhaled aminoglycosides for patients with persistent smear-positive pulmonary tuberculosis. AB - A proportion of patients with drug-resistant and drug-susceptible tuberculosis (TB) have sputum that is smear and culture positive for Mycobacterium tuberculosis for a prolonged period of time, despite conventional therapy. Among such patients with refractory TB, an unblinded, observational study was undertaken that used conventional TB therapy and adjunctive aerosol aminoglycosides. Patients with persistent smear- and culture-positive sputum for M. tuberculosis (despite > or =2 months of optimal systemic therapy) were selected for adjunctive treatment via inhalation with aminoglycosides, and microbiological responses were monitored. Thirteen of 19 patients converted to smear negativity during the study: 6 of 7 with drug-susceptible TB and 7 of 12 with drug-resistant TB. Among patients with drug-susceptible TB, the median time to sputum conversion was 23 days, a shorter time than for a population of historical control patients. Recurrent infection was not observed. Adjunctive aerosol aminoglycosides may expedite sterilization of sputum among certain patients with refractory TB and diminish the risk of transmission. PMID- 11118386 TI - Fungal endocarditis: evidence in the world literature, 1965-1995. AB - We analyzed 270 cases of fungal endocarditis (FE) that occurred over 30 years. Vascular lines, non-cardiac surgery, immunocompromise and injection drug abuse are increasing risk factors. Delayed or mistaken diagnosis (82% of patients), long duration of symptoms before hospitalization (mean +/- standard deviation, 32+/-39 days) and extracardiac manifestations were characteristic. From 1988 onwards, 72% of patients were diagnosed preoperatively, compared with 43% before 1988 (P=.0001). The fungi most commonly isolated were Candida albicans (24% of patients), non-albicans species of Candida (24%), Apergillus species (24%), and Histoplasma species (6%); recently-emerged fungi accounted for 25% of cases. The mortality rate was 72%. Survival rates were better among patients who received combined surgical-antifungal treatment, were infected with Candida, and had univalvular involvement. Improvement in the survival rate (from <20% before 1974 to 41% currently) coincided with the introduction of echocardiography and with improved diagnostic acumen. Fungal endocarditis recurs in 30% of survivors. It is recommended that fungal endocarditis be diagnosed early through heightened diagnostic acumen; that patients be treated with combined lipid-based amphotericin B and early surgery; and that patients be followed up for > or =4 years while on prophylactic antifungal therapy. PMID- 11118387 TI - Type 1/Type 2 immunity in infectious diseases. AB - T helper type 1 (Th1) lymphocytes secrete secrete interleukin (IL)-2, interferon gamma, and lymphotoxin-alpha and stimulate type 1 immunity, which is characterized by intense phagocytic activity. Conversely, Th2 cells secrete IL-4, IL-5, IL-9, IL-10, and IL-13 and stimulate type 2 immunity, which is characterized by high antibody titers. Type 1 and type 2 immunity are not strictly synonymous with cell-mediated and humoral immunity, because Th1 cells also stimulate moderate levels of antibody production, whereas Th2 cells actively suppress phagocytosis. For most infections, save those caused by large eukaryotic pathogens, type 1 immunity is protective, whereas type 2 responses assist with the resolution of cell-mediated inflammation. Severe systemic stress, immunosuppression, or overwhelming microbial inoculation causes the immune system to mount a type 2 response to an infection normally controlled by type 1 immunity. In such cases, administration of antimicrobial chemotherapy and exogenous cytokines restores systemic balance, which allows successful immune responses to clear the infection. PMID- 11118388 TI - Concerns regarding the Centers for Disease Control's published guidelines for pelvic inflammatory disease. AB - The International-Infectious Disease Society for Obstetrics and Gynecology-USA (I IDSOG-USA) has concerns about the most recently published Centers for Disease Control and Prevention (CDC) guidelines for pelvic inflammatory disease (PID). I IDSOG-USA advocates the following changes when the guidelines are revised. We recommend the use of the term "upper genital tract infection" (UGTI), followed by the designation of the etiologic agent, instead of the currently employed term, "pelvic inflammatory disease," or PID. In diagnoses, there should be greater emphasis on signs and symptoms related to subclinical or occult UGTI. Therapeutic recommendation for the treatment of UGTI should be documented for various stages of this diverse disease entity. There should be greater emphasis on hospitalization for infected nulligravida teenagers. This permits monitoring of antibiotic treatment and provides a site for medical educational efforts to teach this medically underserved segment of our society how to protect their future fertility, their health, and their lives. PMID- 11118389 TI - Vancomycin-intermediate and -resistant Staphylococcus aureus: what the infectious disease specialist needs to know. AB - Ever since the first strain of Staphylococcus aureus with reduced susceptibility to vancomycin and teicoplanin was reported from Japan, there has been a lot of confusion regarding the laboratory and clinical approach to patients with infections due to S. aureus with reduced susceptibility to vancomycin. To date, 6 clinical infections with vancomycin-intermediate S. aureus (VISA) have been reported in the United States. Intermediate resistance appears to develop from preexisting strains of methicillin-resistant S. aureus in the presence of vancomycin, and all but 1 infection occurred in patients with exposure to dialysis for renal insufficiency. Detection of VISA is difficult in the laboratory, and special inquiries about susceptibility testing methods may be needed. These VISA-infected patients had underlying illnesses, and their infections did not appear to respond well to conventional treatment. Prevention strategies have been outlined. Without continued vigilance in enforcing infection control measures, improved use of antimicrobials, and coordination of efforts among public health authorities, increasing levels of vancomycin resistance in S. aureus are likely to be encountered. PMID- 11118390 TI - Determinants of antimicrobial prophylaxis use and treatment for wasting among patients with advanced human immunodeficiency virus disease in the United States, 1995-1998. AB - Despite US Public Health Service (USPHS) recommendations for antimicrobial prophylaxis for patients with advanced human immunodeficiency virus (HIV) disease, the proportion of patients who receive prophylaxis is not known. We measured the prevalence of antimicrobial prophylaxis use, and treatment for HIV wasting at baseline among 531 patients with advanced HIV disease enrolled in a multicenter randomized trial of red blood cell transfusion. Use of antimicrobial prophylaxis and treatment for wasting in the 30 days before enrollment was ascertained in patients eligible for primary prophylaxis, secondary prophylaxis, or both, according to USPHS guidelines. There was high utilization of primary and secondary Pneumocystis carinii pneumonia prophylaxis, variability in primary Mycobacterium avium complex prophylaxis by center, and low use of primary cytomegalovirus prophylaxis. Treatment of wasting was more common in white than nonwhite patients and in patients with HIV disease who lived in the region west of the Mississippi River of the United States versus those whose lived in the eastern region. PMID- 11118391 TI - Sex differences in nevirapine rash. AB - Nevirapine is a nonnucleoside reverse transcriptase inhibitor (NNRTI) that has the most common treatment limiting side effect of rash. Severe rash has been observed in 3% of patients taking nevirapine in clinical trials, 85% of whom were men. In a multicenter, retrospective cohort study of all patients who received nevirapine over a 5-year period, severe rash was noted in 9 of 95 women and 3 of 263 men (risk ratio [RR], 8.31; 95% confidence interval [CI], 2.3-30.0; P=.005). Women were more likely to discontinue nevirapine therapy because of rash (RR, 4.5; 95% CI, 1. 9-10.5; P=.0005). After adjusting for age and baseline CD4 cell count in multivariate analysis, women had a 7-fold increase in risk for severe rash and were 3.5 times more likely to discontinue nevirapine therapy. In women of reproductive age for whom contraception may occur, nevirapine remains the NNRTI of choice. Recognition of sex differences in this severe adverse event will be important in prescribing nevirapine. PMID- 11118392 TI - Metabolic abnormalities and cardiovascular disease risk factors in adults with human immunodeficiency virus infection and lipodystrophy. AB - We evaluated metabolic and clinical features of 71 HIV-infected patients with lipodystrophy by comparing them with 213 healthy control subjects, matched for age and body mass index, from the Framingham Offspring Study. Thirty HIV-infected patients without fat redistribution were compared separately with 90 matched control subjects from the Framingham Offspring Study. Fasting glucose, insulin, and lipid levels; glucose and insulin response to standard oral glucose challenge; and anthropometric measurements were determined. HIV-infected patients with lipodystrophy demonstrated significantly increased waist-to-hip ratios, fasting insulin levels, and diastolic blood pressure compared with controls. Patients with lipodystrophy were more likely to have impaired glucose tolerance, diabetes, hypertriglyceridemia, and reduced levels of high-density lipoprotein (HDL) cholesterol than were controls. With the exception of HDL cholesterol level, these risk factors for cardiovascular disease (CVD) were markedly attenuated in patients without lipodystrophy and were not significantly different in comparison with controls. These data demonstrate a metabolic syndrome characterized by profound insulin resistance and hyperlipidemia. CVD risk factors are markedly elevated in HIV-infected patients with fat redistribution. PMID- 11118393 TI - Paronychia in association with indinavir treatment. AB - To assess a possible association between antiretroviral treatment and paronychia, we conducted a retrospective cohort study of 288 human immunodeficiency virus positive protease inhibitor recipients. Indinavir treatment-adjusted for age, sex, CD4 count, diabetes status and other antiretroviral drug exposures-was significantly associated with paronychia of the great toe (hazard ratio 4.7; 95% confidence interval 1.6-13.9). PMID- 11118394 TI - Reactivation of hepatitis B virus replication accompanied by acute hepatitis in patients receiving highly active antiretroviral therapy. AB - We describe 2 patients who were initially positive for antibodies to hepatitis B surface antigen and who experienced a strong and sudden increase of hepatitis B virus (HBV) replication during highly active antiretroviral therapy (HAART). We found that reactivation of HBV replication during HAART can occur independently of lamivudine resistance or withdrawal of lamivudine, and in spite of increasing CD4(+) cell counts. PMID- 11118396 TI - Errata. PMID- 11118395 TI - Choosing a route of administration for quadrivalent meningococcal polysaccharide vaccine: intramuscular versus subcutaneous. AB - A clinical trial was conducted to compare intramuscular (im) with subcutaneous (sc) routes for administration of quadrivalent meningococcal polysaccharide vaccine in 141 adults. Safety assessment showed the im route had reduced erythema (P<.01) and reduced headache on days 1 and 2 (P<.05). Serological testing for serum bactericidal antibody titers against capsular groups A and C did not detect significant differences. PMID- 11118397 TI - Toxoplasmosis, a severe complication in allogeneic hematopoietic stem cell transplantation: successful treatment strategies during a 5-year single-center experience. AB - Toxoplasmosis is a rare but often fatal complication that occurs after patients undergo allogeneic hematopoietic stem cell transplant. At our institution, toxoplasmosis was diagnosed in 8 of 301 patients who received stem cell transplants. Disseminated toxoplasmosis with a rapid fatal course was observed in 2 patients. Six patients had cerebral toxoplasmosis diagnosed on the basis of neurological signs and observation of the patients' mental confusion, seizures, and typical lesions (which were assessed by computed tomography, magnetic resonance imaging, or both). Seroconversion of antitoxoplasma immunoglobulin and a discovery of toxoplasma deoxyribonucleic acid in the cerebrospinal fluid (confirmed by use of polymerase chain reaction) were documented in all patients. Treatment consisted of clindamycin therapy (for 2 patients) and of pyrimethamine clindamycin therapy, sulfadiazine therapy, or both (for 5 patients). Patients showed improvement after therapy, as assessed by clinical and radiological means. Three of 8 patients survive-1 without any residual neurological symptoms and 2 with minimal neurological symptoms. PMID- 11118398 TI - Fluid management in severe preeclampsia (VESPA): survey of members of ISSHP. AB - OBJECTIVE: To determine international expert practice of fluid management and monitoring in severe preeclampsia. METHODS: The 447 members of the ISSHP (International Society for the Study of Hypertension in Pregnancy) were issued a postal questionnaire to determine their views and practices of fluid management in severe preeclampsia. RESULTS: One hundred sixty-six (37%) completed questionnaires were received. Responses indicated that there is no consensus regarding most aspects of management of severe preeclamptic patients. In particular, there is no agreement about which fluid type to administer and how to assess circulatory status in these patients. There were also wide variations in the use of plasma volume expansion as a treatment modality. Statistical comparison of the use of Swan-Ganz catheters in "theoretical" and "actual" practice showed highly significant differences (p < 0.001). The majority of respondents were interested in participating in future research. CONCLUSION: The results reflect genuine uncertainty generated by a lack of evidence from randomized trials addressing the acute management of severe preeclamptic patients. Even where clinicians are confident "in theory" that a particular form of treatment is the best, they do not appear to have the resources or commitment to match this with "practice." The majority of respondents were very keen to develop the questions raised further in the context of multicenter clinical trials. PMID- 11118399 TI - Lack of agreement between central venous pressure and pulmonary capillary wedge pressure in preeclampsia. AB - OBJECTIVE: To establish if agreement exists between central venous pressure (CVP) and pulmonary capillary wedge pressure (PCWP) measurements in severe hypertension in pregnancy as analyzed by tests of bias, precision, and 95% limits of agreement. METHODS: In a prospective study, CVP and PCWP data in 30 patients were collected by means of a pulmonary artery catheter from initiation of therapy until delivery. Patients with a diastolic blood pressure of more than 110 mm Hg were included. Correlation and agreement between CVP and PCWP before and after treatment were evaluated. RESULTS: The correlation coefficient (r) for CVP-PCWP data in 30 untreated patients was r = 0.64 (p = 0.0002) and for 256 pairs of posttreatment data, it was r = 0.53 (p < 0.0001). Linear regression and correlation for each individual patient in 29 patients with more than 3 measurements showed a significant correlation (p < 0.05) in 19 patients (66%). Correlation was poor (p > 0.05) in 10 patients (34%). The mean difference between PCWP and CVP was 3.5 +/- 2.6 mm Hg (limits of agreement: -1.6 to 8.7) in untreated patients. The mean difference between PCWP and CVP for 256 pairs of data derived posttreatment was 4.9 +/- 3.8 mm Hg (limits of agreement: -2.7 to 12. 5). CONCLUSION: Invasive measurements of CVP and PCWP were found to agree poorly. Until a reliable noninvasive method is available to measure left ventricular preload, PCWP is the measurement of choice when invasive hemodynamic monitoring is necessary in patients with severe preeclampsia. PMID- 11118400 TI - Clinical usefulness of maternal body mass index in twin pregnancies. AB - OBJECTIVE: We examined whether maternal body mass index (BMI) during prepregnancy is useful for prediction of maternal preeclampsia in twin pregnancies. METHODS: We studied 250 dichorionic twin pregnancies and 3196 singleton pregnancies. Maternal BMI was calculated during prepregnancy in both twin and singleton pregnancies. The incidence of maternal preeclampsia was compared among three groups, low-BMI [< -1.5 standard deviations (SD)], normal-BMI, and high-BMI (> + 1.5 SD) groups, in both singleton and twin pregnancies. RESULTS: In singleton pregnancies, the incidence of maternal preeclampsia in the high-BMI group was significantly higher than that in the normal-BMI group (p < 0.05). The relative risk by high BMI was 8.5 (95% confidence interval: 5.6-12.0). However, in twin pregnancies, no significant differences were observed in these values. CONCLUSIONS: Body mass index during prepregnancy was not useful for the prediction of preeclampsia in twin pregnancies. Mechanisms other than maternal weight may be associated with the beginning of preeclampsia in twin pregnancies. PMID- 11118401 TI - Doppler ultrasound screening predicts recurrence of poor pregnancy outcome in subsequent pregnancies, but not the recurrence of PIH or preeclampsia. AB - OBJECTIVE: To assess the role of Doppler uterine artery screening in the prediction of recurring hypertensive disorders in a high-risk population. METHODS: Ninety-four women with a history of hypertensive disorders in previous pregnancies underwent ultrasound color Doppler to analyze blood flow in the uterine arteries at 21-22 weeks of gestation. We evaluated the performance of the Pulsatility Index (PI) as well as the diastolic notch to predict recurring hypertensive disorders. Outcome measures were the recurrence of hypertensive disorders, and poor pregnancy outcome due to intrauterine death growth retardation, intrauterine death, placental abruption, hemolysis, elevated liver enzymes, and low platelets (HELLP) syndrome, eclampsia, or premature birth. Onset of symptoms was before 35 weeks in all cases of poor pregnancy outcome. RESULTS: Doppler flow recordings were obtained from a well-defined location in both uterine arteries. The predictive value of the uterine artery PI for recurring hypertensive disease was poor and not significant; interestingly, however, the predictive values for poor pregnancy outcome were good (sensitivity 83%, specificity 71%, p < 0.001). The PI also provides a good test for intrauterine growth retardation (sensitivity 80%, specificity 69%, p < 0.01). The "diastolic notch" did not perform as well as the PI. CONCLUSIONS: Uterine artery screening did significantly predict the recurrence of poor pregnancy outcome due to hypertensive complications in this high-risk group. In contrast, gestational hypertension and preeclampsia with normal pregnancy outcome were not significantly predicted by uterine artery screening. PMID- 11118402 TI - Angiotensin sensitivity test revisited. AB - OBJECTIVE: To determine the validity of a single angiotensin sensitivity test as predictor of pregnancy-induced hypertension with special reference to the dietary sodium intake at the time of testing. METHODS: The angiotensin sensitivity test was successfully performed at 32 weeks' gestation in 104 women. In 90 of these women, the 24-h urinary sodium-creatinine ratio was known. Using an effective pressure dose of 10 ng/kg/min as the cutoff level, test characteristics were assessed in both the total population and after subdivision into a sodium restricted (n = 23) and an unrestricted diet group (n = 67). RESULTS: The incidence of pregnancy-induced hypertension was 13.4%. The number of positive angiotensin sensitivity tests was 7.5%. Test characteristics showed poor sensitivity (22.2%) and high specificity (94.8%); positive and negative predictive values were 40.0% and 88.7%, respectively. None of the sodium restricted women was angiotensin sensitive. Sodium restriction did not have a significant influence on sensitivity, specificity, and predictive values of the test. CONCLUSION: The angiotensin sensitivity test is not an appropriate screening test to predict hypertensive disorders of pregnancy. No significant effect of dietary sodium restriction was found. PMID- 11118403 TI - Methylenetetrahydrofolate reductase polymorphisms in preeclampsia and the HELLP syndrome. AB - OBJECTIVE: To investigate the prevalence of the 677 (C --> T) and 1298 (A --> C) polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene in our preeclamptic population. For a summary estimation of the risk of the 677 (C --> T) polymorphism for preeclampsia, we also performed a meta-analysis on four previously published case-control studies to which our results were added. METHODS: Genotypes were analyzed by polymerase chain reaction followed by restriction enzyme analysis. The results of 176 nonpregnant women, previously hospitalized for preeclampsia in a tertiary care center, were compared with 403 Dutch population-based controls. Results were statistically analyzed with a chi square test. MEAN OUTCOME MEASURES: The incidence of the 677 (C --> T) and 1298 (A --> C) polymorphisms in the MTHFR gene. RESULTS: The incidence of both MTHFR missense polymorphisms was not significantly different between cases and controls. We found an odds ratio (OR) of 1.5 [95% confidence interval (CI) 0.8 2.6, p = 0.17] and an OR of 1.0 (95% CI 0.6-1.9, p = 0.23) for the 677 (C --> T) and the 1298 (A --> C) polymorphism, respectively, in cases comparing the prevalence of the homozygous genotype versus the other two genotypes. The meta analysis resulted in a significant OR of 2.0 (95% CI 1.4-2.9). CONCLUSIONS: In contrast to four previous studies, we were neither able to confirm an increased risk for preeclampsia to the 677 (C --> T) polymorphism nor did we find an increased risk for preeclampsia to the 1298 (A --> C) polymorphism. From the meta analysis, however, we conclude that it cannot be ruled out that the homozygous 677TT genotype is a modest but significant risk factor for preeclampsia. PMID- 11118404 TI - Apolipoprotein E polymorphism in South African Zulu women with preeclampsia. AB - OBJECTIVES: Apolipoprotein E may contribute to the hypertiglyceridemia and consequent endothelial dysfunction of preeclampsia. We carried out a study to determine whether the apolipoprotein E genotype plays any role as a risk factor for preeclampsia in a black South African population with a high incidence of preeclampsia. DESIGN: A descriptive, prospective study design was used. SETTING: King Edward VIII Hospital, a tertiary care, referral academic hospital in Durban, South Africa. PATIENTS AND PARTICIPANTS: One hundred three South African Zulu women with preeclampsia and 110 healthy normotensive women attending the antenatal clinic were recruited. MAIN OUTCOME MEASURES: The relationship between the apolipoprotein E allele and genotype frequencies to preeclampsia as well as adverse perinatal outcome. RESULTS: The frequencies of varepsilon2 and varepsilon4 alleles (0. 19 and 0.25, respectively) were much higher than those reported in other population groups. However, there was no significant difference in the apolipoprotein E genotype and allele frequencies between the study and the control groups. The varepsilon2/2 genotype was associated with increased risk of perinatal death (p = 0.047). CONCLUSION: The study suggests that, despite the high incidence of both preeclampsia and the varepsilon2 and varepsilon4 alleles in South African Zulu women, apolipoprotein E genotype does not appear to be a risk factor for preeclampsia in this population. PMID- 11118405 TI - Inhibin A serum levels in proteinuric and nonproteinuric pregnancy-induced hypertension: evidence for placental involvement in gestational hypertension? AB - OBJECTIVES: To evaluate the serum levels of inhibin A in pregnant women with different types of hypertension. METHODS: A case-control study, including 60 cases (20 women with preeclampsia, 20 with mild gestational hypertension, and 20 with chronic hypertension), and 60 gestational-age- and parity-matched controls. Inhibin A was measured in duplicate by enzyme-linked immunosorbent assay in serum samples frozen at -80 degrees C. RESULTS: As compared to controls, inhibin A levels were significantly elevated in women with preeclampsia ?2.32 standard deviation (SD) 1.4? versus 0.50 (0.29) ng/mL, p < 0.001) and gestational hypertension [1.09 (0.73) versus 0.55 (0.29) ng/mL, p < 0.05], but not in the group of chronic hypertension [0.88 (0.69) versus 0.54 (0.39) ng/mL, p = 0.08]. Overlap in inhibin A values between cases and controls was observed in 20% (4/20) of women with preeclampsia and 55% (11/20) with gestational hypertension. CONCLUSIONS: Increased serum inhibin A may indicate that a proportion of mild nonproteinuric hypertension cases are associated with placental involvement. PMID- 11118406 TI - Platelet responsiveness to L-arginine in hypertensive disorders of pregnancy. AB - OBJECTIVE: In chronically hypertensive (CH), preeclamptic (PE), and normotensive pregnant women (N), we investigated ex vivo platelet aggregation in response to L arginine (L-Arg) and sodium nitroprusside (SN), which are respectively the substrate and donor of nitric oxide (NO). METHODS: Platelet aggregation was determined with a dual-channel aggregometer by measuring transmittance of light through the sample in comparison to platelet poor plasma, as a reference. Aggregation induced by adenosine diphosphate was continuously recorded for 3 min and measured before and after preincubation with L-Arg and SN. RESULTS: Preincubation with L-Arg significantly reduced platelet aggregation in N and CH patients (p < 0.05) but not in PE women. Preincubation with SN affected aggregation in PE women also (p < 0.001). No correlation was found between platelet response to L-Arg or SN stimuli and the severity of hypertensive disorders expressed as week of gestation at delivery or birth weight. CONCLUSIONS: The present study demonstrates that a decreased platelet sensitivity to L-Arg characterizes PE women, whereas SN maintains its antithrombotic power. This impairment seems to be specific for PE, because platelets of CH patients utilize L-Arg normally. This finding supports the involvement of the L-Arg-NO pathway in the pathogenesis of the procoagulative features of PE and probably in the onset of the disease. The maintained response to SN in PE patients suggests a possible therapeutical use of NO donors in the disease. PMID- 11118407 TI - Evaluation of a noninvasive transcranial Doppler and blood pressure-based method for the assessment of cerebral perfusion pressure in pregnant women. AB - OBJECTIVE: We have developed a Doppler method for the estimation of cerebral perfusion pressure (CPP) using noninvasive techniques. Our objective was to evaluate our new method in pregnant women. METHODS AND MATERIALS: Laboring women with a lumbar epidural in situ had transcranial Doppler interrogation of the maternal middle cerebral artery (MCA) to measure systolic, diastolic, and mean velocities. A pressure transducer was connected to the epidural catheter and pressure was recorded. Systolic (SBP), diastolic (DBP), and mean (MAP) blood pressure were taken with a Dinamap monitor. Doppler estimated CPP (mm Hg) = [V(mean)/(V(mean) - V(diastolic)](MAP - DBP) and directly measured CPP = MAP - Epidural pressure data were plotted on a Bland-Altman graph with limits of agreement. The mean difference (the mean of the sum of both positive and negative differences) and absolute difference (the mean of the sum of the absolute differences) were calculated. In addition, linear and polynomial regression analyses were performed. RESULTS: Twenty laboring women were studied. All had normal pregnancies. The mean maternal age was 28 +/- 7 years and the mean gestational age was 39 +/- 2 weeks. The mean maternal MAP was 77 +/- 12 mm Hg. The Bland-Altman plot showed a mean difference of 2.2 mm Hg at a mean CPP of 65 +/- 12 mm Hg; with a standard deviation of 4.8 mm Hg, the absolute difference was 3.9 +/- 3.0 mm Hg at a mean CPP of 65 +/- 12 mm Hg. The regression analysis showed an r = 0.92, r(2) = 0.86, and p < 0.0001. CONCLUSIONS: Our formula allows the estimation of CPP using a simple calculation and noninvasively acquired data. This method may be of use for frequent, easy, and accurate CPP and intracranial pressure estimation and may, as such, have significant research and clinical applications. PMID- 11118408 TI - Reproductive tissue renin gene expression in preeclampsia. AB - OBJECTIVE: Using an analogy with renin gene overexpression, low-renin hypertension animal models, we wished to test the hypothesis that renin gene expression is increased in decidua basalis in human gestation with preeclampsia. METHODS: Human placentas were obtained immediately after delivery from 11 control (C) and 11 preeclamptics (PE). Tissue samples were microdissected and renin gene expression in decidua basalis (DB), chorionic villi (CV), and decidua vera (DV) was measured using dot-blot hybridization. RESULTS: Overall renin gene expression is highest in decidua basalis (mean +/- SEM, 2.66 +/- 0.69 densitometry area units) compared to chorionic villi (mean +/- SEM, 1.85 +/- 0.5) or compared to decidua vera (mean +/- SEM, 1.63 +/- 0.9) (both t-tests p = 0.001 two-tailed and analysis of variance p = 0.0001). Renin gene expression in DB and in CV was similar in both preeclamptic and normal pregnancies (DB mean +/- SEM C 2.79 +/- 0.96 versus PE 2.54 +/- 1.04, and CV mean +/- SEM C 2.11 +/- 0.91 versus PE, 1.59 +/- 0. 44). Renin gene expression in DV was approximately threefold higher in tissues from preeclamptics compared to control (mean +/- SEM PE 2. 44 +/- 1.76 versus C 0.82 +/- 0.42). Using the median value of 0.5 units for DV as cutoff, the preeclamptics displayed higher renin gene expression (chi square p = 0.033, two tailed). CONCLUSION: Our data suggest that renin gene expression is increased in preeclampsia in decidua vera. This may explain previously reported increased renin secretion in uterine circulation in preeclampsia. PMID- 11118409 TI - The effects of hybridization in plants on secondary chemistry: implications for the ecology and evolution of plant-herbivore interactions. AB - Natural hybridization is a frequent phenomenon in plants. It can lead to the formation of new species, facilitate introgression of plant traits, and affect the interactions between plants and their biotic and abiotic environments. An important consequence of hybridization is the generation of qualitative and quantitative variation in secondary chemistry. Using the literature and my own results, I review the effects of hybridization on plant secondary chemistry, the mechanisms that generate patterns of chemical variation, and the possible consequences of this variation for plants and herbivores. Hybrids are immensely variable. Qualitatively, hybrids may express all of the secondary chemicals of the parental taxa, may fail to express certain parental chemicals, or may express novel chemicals that are absent in each parent. Quantitatively, concentrations of parental chemicals may vary markedly among hybrids. There are five primary factors that contribute to variation: parental taxa, hybrid class (F(1), F(2), etc.), ploidy level, chemical class, and the genetics of expression (dominance, recessive vs. additive inheritance). This variation is likely to affect the process of chemical diversification, the potential for introgression, the likelihood that hybrids will facilitate host shifts by herbivores, and the conditions that might lead to enhanced hybrid susceptibility and lower fitness. PMID- 11118410 TI - Distribution and sequence analysis of the centromere-associated repetitive element CEN38 of Sorghum bicolor (Poaceae). AB - Fluorescence in situ hybridization (FISH) of a large-insert genomic clone, BAC 22B2, previously suggested that Sorghum bicolor (2n = 20) has the tetraploid architecture A(b)A(b)B(b)B(b). Here, we report on BAC 22B2 subclone pCEN38 (1047 bp insert) as related to sorghum and sugarcane. Mitotic FISH of six different subclones of BAC 22B2 showed that pCEN38 produced the strongest specificity to the A(b) subgenome and signal occurred primarily near centromeres. Southern blots of pCEN38 to 21 crop plants revealed a narrow taxonomic distribution. Meiotic metaphase I FISH positioned pCEN38 sequences near active centromeres. Pachytene FISH revealed that the distributions are trimodal in several B(b) and possibly all sorghum chromosomes. DNA sequencing revealed that the pCEN38 fragment contains three tandemly repeated dimers (<280 bp) of the same sequence family found in sorghum clone pSau3A10, and that each dimer consists of two divergent monomers (<140 bp). Sequence comparisons revealed homology between the pCEN38 monomers and the SCEN 140 bp tandem repeat family of sugarcane. FISH of pCEN38 yielded signal in centromere regions of most but not all sugarcane chromosomes. Results suggest that sugarcane and sorghum share at least one ancestor harboring elements similar to pCEN38 and SCEN and that each species had an ancestor in which the repetitive element was weakly present or lacking. PMID- 11118411 TI - Unfertilized ovules of Epilobium obcordatum (Onagraceae) continue to grow in developing fruits. AB - To determine whether unfertilized ovules continue to grow when in an ovary containing fertilized ovules, we measured ovule lengths in developing fruits of Epilobium obcordatum that were harvested 4, 5, 8, and 10 d post pollination. We found that unfertilized ovules that were in the presence of fertilized ovules continued to grow and that there was a broad range of overlap in their sizes at all sampling times. This effect was found for two types of unfertilized ovules that occur throughout the length of the ovary: normal, unfertilized ovules, apparently bypassed by pollen tubes; and sterile ovules lacking an embryo sac. In addition, there is a position effect within developing fruits. Both fertilized and unfertilized ovules are larger at the stylar end. In six samples resulting from pollination with a single pollen tetrad, a total of 18 embryos were found, and the effect on unfertilized ovules, greatest at the stylar end, diminished with distance from the ovules with embryos. Our results are consistent with the interpretation that diffusible hormones produced by developing seeds cause nearby unfertilized ovules to grow. We conclude that caution is necessary when attempting to infer ovule fertilization histories from the appearances of ovules in developing and mature fruits. What are often inferred to be aborted seeds, in many cases, may not be seeds at all. They may be enlarged, unfertilized ovules. PMID- 11118412 TI - Genetic variation and evolutionary trade-offs for sexual and asexual reproductive modes in Allium vineale (Liliaceae). AB - Populations of Allium vineale commonly include individuals with very different allocation patterns to three modes of reproduction: sexual flowers, aerially produced asexual bulbils, and belowground asexual offsets. If selection is currently acting to maintain these different allocation patterns there must be a genetic basis for variation in allocation to these three reproductive modes. In addition, negative genetic correlations between reproductive traits would imply evolutionary trade-offs among reproductive strategies. We evaluated the heritability of these allocation patterns by growing 16 clones from a single population in the greenhouse at two levels of fertilization. Bulb mass and the mass and number of bulbils, offsets, and flowers were used as response variables, in addition to the proportion allocated to each reproductive mode. We found evidence of substantial heritable variation in allocation to sexual reproduction and in allocation within the two modes of asexual reproduction, indicating high sensitivity of these allocation patterns to natural selection. We also found evidence of strong negative genetic correlations between bulbil and flower traits, as well as between bulbil and offset traits, with one group of genotypes allocating greater resources to aerial asexual bulbils and the second group allocating more resources to belowground asexual offsets and aerial flowers. Phenotypic plasticity in allocation to above- vs. belowground asexual reproduction and sexual vs. asexual aerial reproduction was limited, indicating that plants are unlikely to change reproductive mode in response to nutrient availability. Together, then, we have demonstrated strong heritability for, and trade-offs in, the reproductive allocation patterns within this plant population. PMID- 11118413 TI - Regional specialization of Sarcodes sanguinea (Ericaceae) on a single fungal symbiont from the Rhizopogon ellenae (Rhizopogonaceae) species complex. AB - We have sampled the mycorrhizal roots of 76 snow plants (Sarcodes sanguinea, Monotropoideae, Ericaceae) in two areas of the Sierra Nevada of California that are ~180 km apart. To identify the fungal symbionts associated with these plants, we first analyzed restriction fragment length polymorphisms (RFLPs) of the internal transcribed spacer region (ITS) of the fungal nuclear ribosomal repeat. Fungal ITS-RFLPs were successfully produced from 57 of the 76 plants sampled, and all symbionts shared the same DNA fragment pattern. The morphology of S. sanguinea mycorrhizae was consistent with that expected from a Rhizopogon species in section Amylopogon. To confirm and refine this identification, a total of six fungal ITS sequences were determined from S. sanguinea mycorrhizae. These sequences were analyzed together with eight existing and eight newly determined ITS sequences from Rhizopogon section Amylopogon. The newly determined sequences include an ITS sequence from the fungal symbiont of pine drops (Pterospora andromedea, Monotropoideae, Ericaceae), a plant that was previously reported to be exclusively associated with the Rhizopogon subcaerulescens group. When these sequences were analyzed together, the Sarcodes symbionts grouped tightly with several collections of R. ellenae including the holotype, one collection of R. idahoensis, and one collection of R. semireticulatus. A different lineage comprised collections of R. subgelatinosus, R. subcaerulescens, another collection of R. semireticulatus, and the Pterospora symbiont. We conclude that S. sanguinea associates exclusively with a single species in the R. ellenae species complex throughout our sampling range. These results indicate a much higher level of specificity in S. sanguinea than was previously reported and confirm the emerging pattern that nonphotosynthetic, monotropoid plants generally associate very specifically with a narrow range of ectomycorrhizal fungi. PMID- 11118414 TI - High root concentration and uneven ectomycorrhizal diversity near Sarcodes sanguinea (Ericaceae): a cheater that stimulates its victims? AB - Sarcodes sanguinea is a nonphotosynthetic mycoheterotrophic plant that obtains all of its fixed carbon from neighboring trees through a shared ectomycorrhizal fungus. We studied the spatial structuring of this tripartite symbiosis in a forest where Sarcodes is abundant, and its only fungal and photosynthetic plant associates are Rhizopogon ellenae and Abies magnifica, respectively. We found disproportionately high concentrations of Abies roots adjacent to Sarcodes roots compared to the surrounding soil. Rhizopogon ellenae colonizes the vast majority of those Abies roots (86-98%), and its abundance tends to decrease with increasing distance from Sarcodes plants. At 500 cm from Sarcodes plants we did not detect R. ellenae, and the ectomycorrhizal community instead was dominated by members of the Russulaceae and Thelephoraceae, which are commonly dominant in other California pinaceous forests. The highly clumped distribution of Abies-R. ellenae ectomycorrhizas indicates that Sarcodes plants either establish within pre-existing clumps, or they stimulate clump formation. Several lines of evidence favor the latter interpretation, suggesting an unexpected mutualistic aspect to the symbiosis. However, the mechanism involved remains unknown. PMID- 11118415 TI - A test of the reserve meristem hypothesis using Verbascum thapsus (Scrophulariaceae). AB - The reserve meristem hypothesis predicts that latent meristems may act as a bet hedging strategy given high-cost, predictable herbivory. Under this hypothesis, damage to a plant should elicit greater branching. This prediction was tested in Verbascum thapsus with three experiments manipulating the intensity and type of damage to reproductive tissue. In the first experiment, seed set was prevented in the treatment group by stigma excision and lanolin application to 80% of the flowers of each plant. In the second experiment, a minimum of two mating pairs of weevils were added to treated plants prior to the onset of flowering. In the third experiment, all fruits were sliced lengthwise twice. All three treatments significantly reduced seed set. In the first two experiments, treated plants significantly increased degree of branching (branch number and total branch length). This supports the reserve meristem hypothesis as an explanation for greater branching in larger plants of V. thapsus. Interestingly, the fruit destruction experiment failed to elicit a branching response, which suggests that the timing of damage is important. PMID- 11118416 TI - Fitness consequences of branching in Verbascum thapsus (Scrophulariaceae). AB - The reserve meristem hypothesis proposes that strong apical dominance suppresses lateral meristems and branches to escape from predictable damage (herbivory). This hypothesis was tested for Verbascum thapsus and its seed predator the weevil Gynmnetron tetrum by two mensurative experiments. The following predictions were made under this hypothesis: the proportion of individuals branched within a population will increase with increased damage, the main stalk of branched plants will be more damaged, and branching increases net seed production. Fifty populations of V. thapsus were extensively surveyed, and one pair of similar sized individuals (branched vs. unbranched) were selected from each population to determine damage patterns and measure seed production. Two of the predictions of the reserve meristem hypothesis were clearly supported. The proportion of fruits damaged on the main stalk of branched plants was significantly greater than unbranched plants, and branched plants produced significantly more seeds. Hence, the reserve meristem hypothesis is supported as an adaptive interpretation of apical dominance in this species. This study is a potential example of overcompensation following granivory in the field. PMID- 11118417 TI - Morphological variation of Pinus flexilis (Pinaceae), a bird-dispersed pine, across a range of elevations. AB - Limber pine (Pinus flexilis James) grows across a wider range of elevations than any other tree species in the central Rockies, from ~1600 m at Pawnee Buttes to >3300 m at Rollins Pass. In this study we investigated two possible explanations for limber pine's success across a broad range of elevations: (1) the sites on which it is found, although separated by >1000 m elevation, may not be very different with respect to environmental factors that affect tree growth, and (2) limber pine growth is insensitive to environmental factors that change with elevation. We compared site characteristics of 12 limber pine stands at elevations ranging from 1630 to 3328 m as well as the growth and morphology of trees in each of these stands. Mean daily air temperature in July decreased linearly with the elevation of the site from 22.8 degrees to 12.6 degrees C. The growth and morphology of limber pine leaves, shoots, and trees were, in general, not related to the elevation or July mean air temperature of the sites. There was, however, a significant decrease in stomatal density with increasing elevation, which may be an acclimational response to restrict water loss at high elevations. Our data suggest that the fundamental and realized niche of limber pine is broad with respect to air temperature. In light of the high gene flow and only slight genetic differentiation among populations of species with bird dispersed seeds, such as limber pine, it is especially unusual to see similar growth throughout an environmental gradient. Physiological and anatomical plasticity or wide physiological tolerance ranges may enable limber pine to uncouple its growth from its environment. PMID- 11118418 TI - Seedbed and moisture availability determine safe sites for early Thuja occidentalis (Cupressaceae) regeneration. AB - Regeneration of many late-successional tree species depends on specialized safe sites. The primary objective was to investigate the roles of seedbed and moisture retention as dimensions of safe sites for the early regeneration of drought sensitive northern white cedar (Thuja occidentalis). We hypothesized that rates of germination, survival, and growth of T. occidentalis are unlikely to differ among seedbed types under conditions of abundant water, but that differences are likely to emerge as water becomes more limited. In a 67-d greenhouse experiment, cedar seeds were sown on logs, leaf litter, and soil of cedar and paper birch (Betula papyrifera) canopy origin. Seedbeds were subjected to three water treatments. Among the water treatments, highest germination rates occurred within the high water treatment, although germination on cedar litter was comparable to that of the low water treatment. Higher germination and survival rates occurred on decayed logs than other natural seedbeds for medium (P = 0.001) and low (P < 0.0001) water treatments. Germination on birch logs occurred at higher rates than on cedar logs within the low water treatment (P = 0.04). Seedling growth for the medium water treatment was lower on leaf litter than any other type of seedbed (P < 0.01). Results generally demonstrated that the interplay between seedbed and moisture retention is a component of safe sites for T. occidentalis regeneration. PMID- 11118419 TI - Post-fire recolonization of dominant epiphytic lichen species on Quercus hypoleucoides (Fagaceae). AB - Following a forest fire (27 500 ha) in 1994, post-fire recolonization of Quercus hypoeleucoides by epiphytic lichens was documented as changes in lichen cover, number of small thalli, specific factors that affected reestablishment of lichens, and modes of dispersal. Three sites in the Chiricahua Mountains (Arizona, USA) were chosen according to the severity of fire damage-unburned, moderately burned, and severely burned. From 1994 through 1997, the amount of dead lichen cover significantly increased at the moderately burned site. For the same time period, the amount of live lichen cover significantly increased at the severely burned site. Numbers of new thalli increased significantly at the severely burned site each year but only in the last year (1996-1997) for the moderately burned site. Bark texture and proximity to trees with lichens were among the most important physical factors for recolonization. The most important means of dispersal for Flavopunctelia praesignis was fragmentation. For Punctelia hypoleucites, the primary means of dispersal was spores. Increases in live lichen cover and numbers of new thalli occur faster in severely burned areas probably due to the loss of lichens on tree trunks, which provides space and a lack of competition. PMID- 11118420 TI - Additive and nonadditive effects of herbivory and competition on tree seedling mortality, growth, and allocation. AB - The interaction between simulated cotyledon herbivory and interspecific competition was studied in a greenhouse experiment using two species of trees, Acer rubrum and Quercus palustris, which commonly invade abandoned agricultural fields. Herbivory treatments were applied as a gradient of cotyledon removal for A. rubrum with 0, 25, 50, 75, and 100% of cotyledon tissue removed. Cotyledons from Q. palustris were clipped and removed (control, early, and late removal) to create a gradient of seed reserve availability. The competition treatment consisted of plugs of old-field vegetation that filled the pots with perennial cover. Mortality of seedlings was higher with competition. There was a significant interaction between herbivory and competition with the highest mortality occurring with competition at the highest intensity of herbivory in both species. Herbivory reduced biomass for Q. palustris only, while competition reduced biomass in both species. Neither species showed an interaction between herbivory and competition for growth. There was a significant interaction between herbivory and competition on allocation patterns for both species, with greater allocation to roots with competition at the highest intensity of herbivory. This study demonstrates the potential for cotyledon herbivory and competition to interact, altering the invasion of tree seedlings into abandoned agricultural land. PMID- 11118421 TI - Phylogeny and classification of Oleaceae based on rps16 and trnL-F sequence data. AB - Phylogenetic relationships among 76 species of Oleaceae, representing all 25 recognized genera of the family, were assessed by a cladistic analysis of DNA sequences from two noncoding chloroplast loci, the rps16 intron and the trnL-F region. Consensus trees from separate and combined analyses are congruent and agree well with nonmolecular data (chromosome numbers, fruit and wood anatomy, leaf glycosides, and iridoids). The two debated genera Dimetra and Nyctanthes, previously suggested to belong to Verbenaceae (sensu lato) or Nyctanthaceae, are shown to belong to Oleaceae, sister to the hitherto genus incertae sedis Myxopyrum. This clade is also supported by anatomical and chemical data. The subfamily Jasminoideae is paraphyletic, and a new classification is presented. The subfamily level is abandoned, and the former Jasminoideae is split into four tribes: Myxopyreae (Myxopyrum, Nyctanthes, and Dimetra), Fontanesieae (Fontanesia), Forsythieae (Abeliophyllum and Forsythia), and Jasmineae (Jasminum and Menodora). The tribe Oleeae (previous subfamily Oleoideae) is clearly monophyletic, comprising the subtribes Ligustrinae (Syringa and Ligustrum), Schreberinae status novus (Schrebera and Comoranthus), Fraxininae status novus (Fraxinus), and Oleinae (12 drupaceous genera). An rps16 sequence obtained from Hesperelaea, known only from the type specimen collected in 1875, confirmed the placement of this extinct taxon in the subtribe Oleinae. PMID- 11118422 TI - Subtribal and generic relationships of Maxillarieae (Orchidaceae) with emphasis on Stanhopeinae: combined molecular evidence. AB - The monophyly of and phylogenetic relationships within the orchid tribe Maxillarieae Pfitzer were evaluated using parsimony analyses of combined nuclear ribosomal and plastid DNA sequence data of ITS 1 and 2, matK, and the trnL intron and the trnL-F intergene spacer. Each of the separate analyses produced highly congruent but weakly supported patterns (by the bootstrap), so these were combined in a single analysis. Analysis of 90 ingroup taxa (representing ~35% of currently recognized genera) and four outgroup taxa produced resolved and highly supported cladograms. Based on the cladograms, we recognize six subtribes: Eriopsidinae, Oncidiinae (including Pachyphyllinae, Ornithocephalinae, and Telipogoninae), Stanhopeinae, Coeliopsidinae, Maxillariinae (including Lycastinae and Bifrenariinae), and Zygopetalinae (including Cryptarrheninae, Dichaeinae, Huntleyinae, and Warreinae). Stanhopeinae were sampled most intensively; their generic relationships were highly resolved in the analysis and largely agree with currently accepted generic concepts based on morphology. Coeliopsidinae (Coeliopsis, Lycomormium, Peristeria) are sister to Stanhopeinae. Correlations are drawn among phylogeny, pollination mechanisms, and life history traits. PMID- 11118423 TI - Molecular phylogeny and biogeography of Linanthus (Polemoniaceae). AB - To better understand the evolutionary history of Linanthus (Polemoniaceae) and its relatives, molecular phylogenies based on DNA sequence data from the internal transcribed spacer (ITS) region of nrDNA and the chloroplast gene matK were estimated using several methods. Our data suggest two separate and well-supported lineages of Linanthus in close association with two other genera-Leptodactylon and Phlox. These results agree with previous molecular systematic work on the Polemoniaceae, but do not support the traditional classification of the genus as a natural group, nor do they support the sectional classification within the genus. With a distribution centered primarily in western North America and a high degree of endemism in the California Floristic Province, it has been suggested by Raven and Axelrod that the origin and diversification of Linanthus and its relatives were tied to the development of a summer-dry climate in western North America, which began around 13-15 million years ago (mya). Increased drying during the Pliocene (1.2-5 mya) has also been hypothesized by Axelrod to have led to an increase in plant speciation in California and adjacent areas. Divergence times within the Linanthus lineages were estimated from the ITS and matK gene trees. A log-likelihood ratio test could not reject clock-like evolution for the matK data; however, the clock was strongly rejected for the ITS data set. Although ITS molecular evolution was not clock-like, the estimated times of divergence were similar to those of the matK data set. Within both lineages of Linanthus there seems to have been considerable diversification that has occurred since the Pliocene. PMID- 11118424 TI - Tribal delimitation and phylogenetic relationships of Loteae and Coronilleae (Faboideae: Fabaceae) with special reference to Lotus: evidence from nuclear ribosomal ITS sequences. AB - The temperate herbaceous tribes Loteae and Coronilleae have traditionally been regarded as taxonomically distinct entities. More recent morphological assessments, however, have challenged this view and suggest combining the two tribes under Loteae. Two key features used to distinguish the Coronilleae from Loteae include jointed fruits and branched root nodules. We evaluate the taxonomic utility of these characters using information derived from phylogenetic analyses of the internal transcribed spacers ITS1 + 2, and the intervening 5.8S region of nuclear ribosomal DNA. Results from this study show that neither the Loteae nor Coronilleae form individual monophyletic groups, and that key fruit and root nodule characters used to distinguish the Coronilleae are homoplastic. Given these data, we support the recognition of a single tribe, Loteae. We also find that Lotus, the largest and most morphologically complex genus in either tribe, is not monophyletic. Rather, it consists of two geographically distinct lineages, Old and New World, each of which are more closely related to other Loteae genera: Old World Lotus are more closely related to Old World Anthyllis, while New World Lotus show closer affinities to Old World Coronilla. These data also have important implications for the biogeography of New World Lotus: equally most parsimonious reconstructions suggest a complex scenario of intercontinental dispersals that involve not only Old World Lotus but Coronilla as well. PMID- 11118425 TI - Allozyme evidence for genetic autopolyploidy and high genetic diversity in tetraploid cranberry, Vaccinium oxycoccos (Ericaceae). AB - Polyploidy has been important in the evolution of angiosperms and may significantly affect population genetic diversity and structure. Nineteen isoenzyme loci were studied in diploid and tetraploid populations of Vaccinium oxycoccos (Ericaceae), and the results are compared with data previously reported for the related V. macrocarpon. Diploid V. oxycoccos and V. macrocarpon were readily discriminated based on their allozymic variation. No evidence for fixed heterozygosity was found in tetraploid V. oxycoccos. In contrast, all polymorphic loci exhibited both balanced and unbalanced heterozygotes, with some individuals exhibiting a pattern consistent with the presence of three alleles. These results support an autopolyploid origin for tetraploid V. oxycoccos. However, tetraploid V. oxycoccos possessed a suite of alleles not found in diploid V. oxycoccos; half of these alleles were shared with V. macrocarpon. This suggests that autotetraploid V. oxycoccos may have undergone hybridization with V. macrocarpon or that the autotetraploid retained the genetic variation present in an ancestral diploid species. Following theoretical expectations, proportion of polymorphic loci, mean number of alleles, and observed heterozygosity were significantly higher for the autotetraploid than for the diploid. Mean inbreeding (F(IS)) was similar for diploid and tetraploid V. oxycoccos. The latter exhibited population differentiation (F(ST)) exceeding both diploid species. PMID- 11118426 TI - Origin and relationships of the tarweed-silversword lineage (Compositae Madiinae). AB - Based on results from phylogenetic analyses of nuclear 18S-26S rDNA internal transcribed spacer (ITS) region sequences, we suggest that the monophyletic tarweed and silversword subtribe (Madiinae) is phylogenetically nested among epaleate, x = 19 species of helenioid Heliantheae. Strong bootstrap support (100%) was obtained for a sister-group relationship between Madiinae and Arnica (including Mallotopus and Whitneya) in an analysis including representatives of recognized genera in a principally Californian clade (Madieae sensu Baldwin) identified from a phylogenetic investigation of Heliantheae s.l. (sensu lato) and Eupatorieae. In all minimum-length trees, the robust lineage comprising Madiinae and Arnica (x = 19) is part of a larger clade that also comprises Eatonella s.s. (sensu stricto), Hulsea, and Venegasia, all with x = 19. The phylogenetic position of Madiinae within a group of genera based uniformly on x = 19 leads us to conclude that the modal numbers of n = 7 and n = 8 (and other numbers, as low as n = 4) in Madiinae are the results of extreme dysploidy. Among the x = 19 "arnicoid" taxa, the near-universal characteristics of perenniality (except in the monotypic Eatonella s.s. and a minority of hulseas) and montane or high latitudinal occurrence (except in the monotypic Venegasia) lead us to suggest that the most recent common ancestor of the tarweeds (a principally annual group of seasonally dry, low-elevation habitats) was probably a montane, herbaceous perennial resembling the unusual subalpine and alpine tarweeds constituting Raillardella s.s. (x = 17), an arnica-like genus. In Madiinae, Raillardella s.s. may be plesiomorphic in habit, capitular and ecological characteristics, and high base chromosome number. Shifts to an annual habit and to low chromosome numbers in Madiinae have been followed by subsequent episodes of polyploidy and descending dysploidy. We conclude that genome evolution in Madiinae has been marked by wide swings in chromosome number that confuse identification of diploids and polyploids. PMID- 11118427 TI - Flowers, fruits, seeds, and pollen of Landeenia gen. nov., an extinct sapindalean genus from the Eocene of Wyoming. AB - The new genus Landeenia is recognized on the basis of flowers, pollen, infructescences, fruits, and seeds from the middle Eocene of southwestern and northwestern Wyoming. Landeenia aralioides (MacGinitie) comb.nov. has cymose inflorescences with actinomorphic, bisexual flowers, a pentamerous calyx, about ten stamens, and a superior gynoecium of ~18 carpels sharing a single style. The fruits are globose to oblate, loculicidally dehiscent capsules, with a persistent calyx, and contain flat, elliptical seeds that are surrounded by a small wing. Pollen removed from the anthers is tricolpate with finely striate sculpture. Although clearly dicotyledonous, the combination of characters found in Landeenia is not known in any modern genus. The familial affinities of the plant, though certainly not with the Araliaceae as previously thought, remain uncertain. However, the combination of characters is consistent with treatment as a member of the Sapindales. The fossil material is thus assigned to the rank of Sapindales Incertae sedis. PMID- 11118428 TI - Floral ontogeny, pattern formation, and evolution in Hibbertia and Adrastaea (Dilleniaceae). AB - Floral development was compared with scanning electron microscopy in 12 Australian species of Hibbertia representing most of its morphological variation, and in the related Adrastaea (Dilleniaceae). Calyx and corolla arise in quincuncial helices in radially symmetrical species, while the petals initiate unidirectionally from one side in zygomorphic species. Stamen number (3-200+) proliferates by centrifugal addition of individual primordia or by innovations of common primordia and ring meristems. Common primordia arise in single-stamen positions alternately with petals, and each produces one to several stamens centrifugally that remain attached to a shared base and form a stamen fascicle. A ring meristem in Adrastaea initiates a whorl of five stamens, alternate with the first stamens but outside their whorl. In radially symmetrical species of Hibbertia, a first ring of stamens is supplemented centrifugally by additional stamens on a meristem ring. The first stamens in zygomorphic species of Hibbertia initiate as a terminal ridge on the floral apex, with subsequent stamens added centrifugally on one side and two carpels initiated on the opposite side. The carpels arise as a simultaneous ring in radially symmetrical flowers, or as a simultaneous pair in zygomorphic species. Staminodial presence is viewed as of minor significance. Four pollinator syndromes are proposed for Hibbertia, related to differing floral architecture. PMID- 11118429 TI - The cellular uptake of anandamide is coupled to its breakdown by fatty-acid amide hydrolase. AB - Anandamide is an endogenous compound that acts as an agonist at cannabinoid receptors. It is inactivated via intracellular degradation after its uptake into cells by a carrier-mediated process that depends upon a concentration gradient. The fate of anandamide in those cells containing an amidase called fatty-acid amide hydrolase (FAAH) is hydrolysis to arachidonic acid and ethanolamine. The active site nucleophilic serine of FAAH is inactivated by a variety of inhibitors including methylarachidonylfluorophosphonate (MAFP) and palmitylsulfonyl fluoride. In the current report, the net uptake of anandamide in cultured neuroblastoma (N18) and glioma (C6) cells, which contain FAAH, was decreased by nearly 50% after 6 min of incubation in the presence of MAFP. Uptake in laryngeal carcinoma (Hep2) cells, which lack FAAH, is not inhibited by MAFP. Free anandamide was found in all MAFP-treated cells and in control Hep2 cells, whereas phospholipid was the main product in N18 and C6 control cells when analyzed by TLC. The intracellular concentration of anandamide in N18, C6, and Hep2 cells was up to 18-fold greater than the extracellular concentration of 100 nm, which strongly suggests that it is sequestered within the cell by binding to membranes or proteins. The accumulation of anandamide and/or its breakdown products was found to vary among the different cell types, and this correlated approximately with the amount of FAAH activity, suggesting that the breakdown of anandamide is in part a driving force for uptake. This was shown most clearly in Hep2 cells transfected with FAAH. The uptake in these cells was 2-fold greater than in vector-transfected or untransfected Hep2 cells. Therefore, it appears that FAAH inhibitors reduce anandamide uptake by cells by shifting the anandamide concentration gradient in a direction that favors equilibrium. Because inhibition of FAAH increases the levels of extracellular anandamide, it may be a useful target for the design of therapeutic agents. PMID- 11118430 TI - Mechanism of human group V phospholipase A2 (PLA2)-induced leukotriene biosynthesis in human neutrophils. A potential role of heparan sulfate binding in PLA2 internalization and degradation. AB - Human group V phospholipase A(2) (hVPLA(2)) has been shown to have high activity to elicit leukotriene production in human neutrophils (Han, S. K., Kim, K. P., Koduri, R., Bittova, L., Munoz, N. M., Leff, A. R., Wilton, D. C., Gelb, M. H., and Cho, W. (1999) J. Biol. Chem. 274, 11881-11888). To determine the mechanism by which hVPLA(2) interacts with cell membranes to induce leukotriene formation, we mutated surface cationic residues and a catalytic residue of hVPLA(2) and measured the interactions of mutants with model membranes, immobilized heparin, and human neutrophils. These studies showed that cationic residues, Lys(7), Lys(11), and Arg(34), constitute a part of the interfacial binding surface of hVPLA(2), which accounts for its moderate preference for anionic membranes. Additionally, hVPLA(2) binds heparin with high affinity and has a well defined heparin-binding site. The site is composed of Arg(100), Lys(101), Lys(107), Arg(108), and Arg(111), and is spatially distinct from its interfacial binding surface. Importantly, the activities of the mutants to hydrolyze cell membrane phospholipids and induce leukotriene biosynthesis, when enzymes were added exogenously to neutrophils, correlated with their activities on phosphatidylcholine membranes but not with their affinities for anionic membranes and heparin. These results indicate that hVPLA(2) acts directly on the outer plasma membranes of neutrophils to release fatty acids and lysophospholipids. Further studies suggest that products of hVPLA(2) hydrolysis trigger the cellular leukotriene production by activating cellular enzymes involved in leukotriene formation. Finally, the temporal and spatial resolution of exogenously added hVPLA(2) and mutants suggests that binding to cell surface heparan sulfate proteoglycans is important for the internalization and clearance of cell surface bound hVPLA(2). PMID- 11118431 TI - Agonist-induced conformational changes at the cytoplasmic side of transmembrane segment 6 in the beta 2 adrenergic receptor mapped by site-selective fluorescent labeling. AB - The environmentally sensitive, sulfhydryl-reactive, fluorescent probe N,N' dimethyl-N-(iodoacetyl)-N'-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) ethylene-diamine (IANBD) was used as a molecular reporter of agonist-induced conformational changes in the beta(2) adrenergic receptor, a prototype hormone-activated G protein-coupled receptor. In the background of a mutant beta(2) adrenergic receptor, with a minimal number of endogenous cysteine residues, new cysteines were introduced in positions 269(6.31), 270(6.32), 271(6.33), and 272(6.34) at the cytoplasmic side of transmembrane segment (TM) 6. The resulting mutant receptors were fully functional and bound both agonists and antagonist with high affinities also upon IANBD labeling. Fluorescence spectroscopy analysis of the purified and site-selectively IANBD-labeled mutants suggested that the covalently attached fluorophore was exposed to a less polar environment at all four positions upon agonist binding. Whereas evidence for only a minor change in the molecular environment was obtained for positions 269(6.31) and 270(6.32), the full agonist isoproterenol caused clear dose-dependent and reversible increases in fluorescence emission at positions 271(6.33) and 272(6.34). The data suggest that activation of G protein-coupled receptors, which are activated by "diffusible" ligands, involves a structural rearrangement corresponding to the cytoplasmic part of TM 6. The preferred conformations of the IANBD moiety attached to the inserted cysteines were predicted by employing a computational method that incorporated the complex hydrophobic/hydrophilic environment in which the cysteines reside. Based on these preferred conformations, it is suggested that the spectral changes reflect an agonist-promoted movement of the cytoplasmic part of TM 6 away from the receptor core and upwards toward the membrane bilayer. PMID- 11118432 TI - A conserved cis-acting element in the parathyroid hormone 3'-untranslated region is sufficient for regulation of RNA stability by calcium and phosphate. AB - Calcium and phosphate regulate parathyroid hormone (PTH) gene expression post transcriptionally by changes in protein-PTH mRNA 3'-untranslated region (UTR) interactions, which determine PTH mRNA stability. We have identified the protein binding sequence in the PTH mRNA 3'-UTR and determined its functionality. The protein-binding element was identified by binding, competition, and antisense oligonucleotide interference. The sequence was preserved among species suggesting its importance. To study its functionality in the context of another RNA, a 63 base pair cDNA PTH sequence was fused to the growth hormone (GH) gene. There is no parathyroid (PT) cell line and therefore an in vitro degradation assay was used to determine the stability of transcripts for PTH, GH, and a chimeric GH-PTH 63 nucleotides with PT cytosolic proteins. The full-length PTH transcript was stabilized by PT proteins from rats fed a low calcium diet and destabilized by proteins from rats fed a low phosphate diet, correlating with PTH mRNA levels in vivo. These PT proteins did not affect the native GH transcript. However, the chimeric GH transcript was stabilized by low calcium PT proteins and destabilized by low phosphate PT proteins, similar to the PTH full-length transcript. Therefore, we have identified a PTH RNA-protein binding region and shown that it is sufficient to confer responsiveness to calcium and phosphate in a reporter gene. This defined element in the PTH mRNA 3'-UTR is necessary and sufficient for the regulation of PTH mRNA stability by calcium and phosphate. PMID- 11118433 TI - Carbohydrate-carbohydrate binding of ganglioside to integrin alpha(5) modulates alpha(5)beta(1) function. AB - Gangliosides GT1b and GD3, components of keratinocyte membranes, inhibit keratinocyte adhesion to fibronectin. Although ganglioside sialylation is known to be important, the mechanism of inhibition is unknown. Using purified insect recombinant alpha(5) and beta(1) proteins and alpha(5)beta(1) integrin from lysed keratinocyte-derived SCC12 cells, we have shown that GT1b and GD3 inhibit the binding of alpha(5)beta(1) to fibronectin. Co-immunoprecipitation of GT1b and alpha(5)beta(1) from SCC12 cells and direct binding of GT1b and GD3 to affinity purified alpha(5)beta(1) from SCC12 cells and insect recombinant alpha(5)beta(1), particularly the alpha(5) subunit, further suggest interaction between ganglioside and alpha(5)beta(1). The carbohydrate moieties of integrin appear to be critical since gangliosides are unable to bind deglycosylated forms of alpha(5)beta(1) from SCC12 and insect cells or poorly glycosylated recombinant alpha(5)beta(1) from Escherichia coli cells. The GT1b-alpha(5)beta(1) interaction is inhibited by concanavalin A, suggesting that GT1b binds to mannose structures in alpha(5)beta(1). The preferential binding of GT1b to high mannose rather than reduced mannose ovalbumin further implicates the binding of GT1b to mannose structures. These data provide evidence that highly sialylated gangliosides regulate alpha(5)beta(1)-mediated adhesion of epithelial cells to fibronectin through carbohydrate-carbohydrate interactions between GT1b and the alpha(5) subunit of alpha(5)beta(1) integrin. PMID- 11118434 TI - The relative activities of the C2GnT1 and ST3Gal-I glycosyltransferases determine O-glycan structure and expression of a tumor-associated epitope on MUC1. AB - In breast cancer, the O-glycans added to the MUC1 mucin are core 1- rather than core 2-based. We have analyzed whether competition by the glycosyltransferase, ST3Gal-I, which transfers sialic acid to galactose in the core 1 substrate, is key to this switch in MUC1 glycosylation that results in the expression of the cancer-associated SM3 epitope. Of the three enzymes known to convert core 1 to core 2, by the addition of GlcNAc to GalNAc in core1 C2GnT1 is the dominant enzyme expressed in normal breast tissue. Expression of C2GnT1 is low or absent in around 50% of breast cancers, whereas expression of ST3Gal-I is consistently increased. Mapping of ST3Gal-I and C2GnT1 within the Golgi pathway showed some overlap. To examine functional competition, the enzymes were overexpressed in T47D cells, which normally make core 1-based structures, have no detectable C2GnT1 activity and express the SM3 epitope. Overexpression of C2GnT1 resulted in loss of binding of SM3 to MUC1, accompanied by a decrease in the GalNAc/GlcNAc ratio, indicative of a switch to core 2 structures. Transfection of a C2GnT1 expressing line with ST3Gal-I restored SM3 binding and reduced GlcNAc incorporation into MUC1 O-glycans. Thus, even when C2GnT1 is expressed, the O glycans added to MUC1 become core 1-dominated structures, provided expression of ST3Gal-I is increased as it is in breast cancer. PMID- 11118435 TI - The STAR/GSG family protein rSLM-2 regulates the selection of alternative splice sites. AB - We identified the rat Sam68-like mammalian protein (rSLM-2), a member of the STAR (signal transduction and activation of RNA) protein family as a novel splicing regulatory protein. Using the yeast two-hybrid system, coimmunoprecipitations, and pull-down assays, we demonstrate that rSLM-2 interacts with various proteins involved in the regulation of alternative splicing, among them the serine/arginine-rich protein SRp30c, the splicing-associated factor YT521-B and the scaffold attachment factor B. rSLM-2 can influence the splicing pattern of the CD44v5, human transformer-2beta and tau minigenes in cotransfection experiments. This effect can be reversed by rSLM-2-interacting proteins. Employing rSLM-2 deletion variants, gel mobility shift assays, and linker scan mutations of the CD44 minigene, we show that the rSLM-2-dependent inclusion of exon v5 of the CD44 pre-mRNA is dependent on a short purine-rich sequence. Because the related protein of rSLM-2, Sam68, is believed to play a role as an adapter protein during signal transduction, we postulate that rSLM-2 is a link between signal transduction pathways and pre-mRNA processing. PMID- 11118436 TI - A novel heat-labile phospholipid-binding protein, SVS VII, in mouse seminal vesicle as a sperm motility enhancer. AB - SVS VII, one of seven major proteins in mouse seminal vesicle secretion, was purified to homogeneity. Neither glycoconjugate nor free thiol group was detected in the protein. The primary structure deduced from the corresponding cDNA was confirmed using amino acid sequence determination, which supported the finding that SVS VII consists of 76 amino acid residues with five disulfide bridges. Accordingly, it has a theoretical molecular mass of 8538, which was proven using the mass spectrum of SVS VII. The CD spectrum of SVS VII in 50 mm phosphate buffer at pH 7.4 appeared as one negative band arising from the beta form at 217 nm and several fine structures due to nonpeptide chromophores including a prominent band for the disulfide bond at 250 nm. This, together with the predicted secondary structures, indicated no helices but a mixture of beta form, beta turn, and unordered form in SVS VII. A cytochemical study illustrated the presence of the SVS VII-binding region on the entire surface of mouse sperm. The SVS VII-sperm binding was inhibited by the dispersed sperm lipids. The results of TLC overlay assay for the binding of (125)I-SVS VII to phospholipids and the interaction between SVS VII and phospholipid liposomes demonstrated a specific binding of this protein to both phosphatidylethanolamine and phosphatidylserine. The SVS VII-sperm binding greatly enhanced sperm motility but did not induce sperm capacitation. Heating the protein solution for 10 min at 90 degrees C unfolded the protein molecule, and the unfolded SVS VII immobilized the sperm. PMID- 11118437 TI - Functional replacement of the essential ESS1 in yeast by the plant parvulin DlPar13. AB - A functionally Pin1-like peptidyl-prolyl cis/trans isomerase (PPIase(1)) was isolated from proembryogenic masses (PEMs) of Digitalis lanata according to its enzymatic activity. Partial sequence analysis of the purified enzyme (DlPar13) revealed sequence homology to members of the parvulin family of PPIases. Similar to human Pin1 and yeast Ess1, it exhibits catalytic activity toward substrates containing (Thr(P)/Ser(P))-Pro peptide bonds and comparable inhibition kinetics with juglone. Unlike Pin1-type enzymes it lacks the phosphoserine or phosphothreonine binding WW domain. Western blotting with anti-DlPar13 serum recognized the endogenous form in nucleic and cytosolic fractions of the plant cells. Since the PIN1 homologue ESS1 is an essential gene, complementation experiments in yeast were performed. When overexpressed in Saccharomyces cerevisiae DlPar13 is almost as effective as hPin1 in rescuing the temperature sensitive phenotype caused by a mutation in ESS1. In contrast, the human parvulin hPar14 is not able to rescue the lethal phenotype of this yeast strain at nonpermissive temperatures. These results suggest a function for DlPar13 rather similar to parvulins of the Pin1-type. PMID- 11118438 TI - Functional conservation of phosphorylation-specific prolyl isomerases in plants. AB - The phosphorylation-specific peptidyl prolyl cis/trans isomerase (PPIase) Pin1 in humans and its homologues in yeast and animal species play an important role in cell cycle regulation. These PPIases consist of an NH(2)-terminal WW domain that binds to specific phosphoserine- or phosphothreonine-proline motifs present in a subset of phosphoproteins and a COOH-terminal PPIase domain that specifically isomerizes the phosphorylated serine/threonine-proline peptide bonds. Here, we describe the isolation of MdPin1, a Pin1 homologue from the plant species apple (Malus domestica) and show that it has the same phosphorylation-specific substrate specificity and can be inhibited by juglone in vitro, as is the case for Pin1. A search in the plant expressed sequence tag data bases reveals that the Pin1-type PPIases are present in various plants, and there are multiple genes in one organism, such as soybean (Glycine max) and tomato (Lycopersicon esculentum). Furthermore, all these plant Pin1-type PPIases, including AtPin1 in Arabidopsis thaliana, do not have a WW domain, but all contain a four-amino acid insertion next to the phospho-specific recognition site of the active site. Interestingly, like Pin1, both MdPin1 and AtPin1 are able to rescue the lethal mitotic phenotype of a temperature-sensitive mutation in the Pin1 homologue ESS1/PTF1 gene in Saccharomyces cerevisiae. However, deleting the extra four amino acid residues abolished the ability of AtPin1 to rescue the yeast mutation under non-overexpression conditions, indicating that these extra amino acids may be important for mediating the substrate interaction of plant enzymes. Finally, expression of MdPin1 is tightly associated with cell division both during apple fruit development in vivo and during cell cultures in vitro. These results have demonstrated that phosphorylation-specific PPIases are highly conserved functionally in yeast, animal, and plant species. Furthermore, the experiments suggest that although plant Pin1-type enzymes do not have a WW domain, they may fulfill the same functions as Pin1 and its homologues do in other organisms. PMID- 11118439 TI - Interaction between YY1 and the retinoblastoma protein. Regulation of cell cycle progression in differentiated cells. AB - Overexpression of the transcription factor YY1 activates DNA synthesis in differentiated primary human coronary artery smooth muscle cells. Overexpression of the retinoblastoma protein together with YY1 blocked this effect. In growth arrested cells, YY1 resides in a complex with the retinoblastoma protein, but the complex is not detected in serum-stimulated S phase cultures, indicating that the interaction of the retinoblastoma protein and YY1 is cell cycle-regulated. Recombinant retinoblastoma protein directly interacts with YY1, destabilizing the interaction of YY1 with DNA and inhibiting its transcription initiator function in vitro. We conclude that in differentiated cells elevation of the nuclear level of YY1 protein favors progression into the S phase, and we propose that this activity is regulated by its interaction with the retinoblastoma protein. PMID- 11118440 TI - Differential association of products of alternative transcripts of the candidate tumor suppressor ING1 with the mSin3/HDAC1 transcriptional corepressor complex. AB - The candidate tumor suppressor ING1 was identified in a genetic screen aimed at isolation of human genes whose expression is suppressed in cancer cells. It may function as a negative growth regulator in the p53 signal transduction pathway. However, its molecular mechanism is not clear. The ING1 locus encodes alternative transcripts of p47(ING1a), p33(ING1b), and p24(ING1c). Here we report differential association of protein products of ING1 with the mSin3 transcriptional corepressor complex. p33(ING1b) associates with Sin3, SAP30, HDAC1, RbAp48, and other proteins, to form large protein complexes, whereas p24(ING1c) does not. The ING1 immune complexes are active in deacetylating core histones in vitro, and p33(ING1b) is functionally associated with HDAC1-mediated transcriptional repression in transfected cells. Our data provide basis for a p33(ING1b)-specific molecular mechanism for the function of the ING1 locus. PMID- 11118441 TI - Cuticular pro-phenoloxidase of the silkworm, Bombyx mori. Purification and demonstration of its transport from hemolymph. AB - Pro-phenoloxidase (proPO) in insects is implicated in the defense against microbes and wounding. The presence of proPO in the cuticle was suggested more than 30 years ago, but it has not been purified. The extract of cuticles of the silkworm, Bombyx mori, was shown to contain two proPO isoforms (F-type and S-type proPOs, which have slightly different mobilities in polyacrylamide gel electrophoresis under nondenaturing conditions). The two isoforms were purified to homogeneity. From hemolymph of the same insect, two types of proPO with the same electrophoretic mobilities as those of cuticular isoforms were separated and were shown to be different at five amino acid residues in one of their subunits. The isoforms in the hemolymph and cuticle were activated by a specific activating enzyme. The resulting active phenoloxidases exhibited almost the same substrate specificities and specific activities toward o-diphenols. The substrate specificities and the susceptibilities to inhibitors, including carbon monoxide, indicated that the purified proPO isoforms were not zymogens of laccase-type phenoloxidase. The proPO in hemolymph was shown to be transported to the cuticle. This demonstration was corroborated by the failure to detect proPO transcripts by Northern analysis of total RNA from epidermal cells. In reversed-phase column chromatography, cuticular and hemolymph proPOs gave distinct elution profiles, indicating that some yet to be identified modification occurs in hemolymph proPO and results in the formation of cuticular proPO. There was little transportation of cuticular proPO to the cuticle when it was injected into the hemocoel. The nature of the modification is described in the accompanying paper (Asano, T., and Ashida, M. (2001) J. Biol. Chem. 276, 11113-11125). PMID- 11118442 TI - Nitric oxide disrupts H2O2-dependent activation of nuclear factor kappa B. Role in sensitization of human tumor cells to tumor necrosis factor-alpha -induced cytotoxicity. AB - Tumor necrosis factor alpha (TNF-alpha) exerts its effect by two distinct signaling pathways. It can trigger cytotoxicity in sensitive target cells. TNF alpha can also promote nuclear factor kappaB (NF-kappaB) activity and regulate the expression of genes that interfere with apoptosis and thus conferring resistance to several apoptotic stimuli. We have observed that interferon-gamma (IFN-gamma) sensitizes human ovarian carcinoma cell lines to TNF-alpha-mediated apoptosis and further, IFN-gamma induces the expression of the inducible nitric oxide synthase (iNOS) and the generation of nitric oxide (NO). This study examines the role of NO in the sensitization of the ovarian carcinoma cell line AD10 to TNF-alpha-mediated cytotoxicity. Treatment of AD10 cells with the NOS inhibitor l-NMA blocked the IFN-gamma-dependent sensitization whereas NO donors (S-nitroso-N-acetylpenicillamine) sensitized these cells to TNF-alpha cytotoxicity. Analysis of the activation status of NF-kappaB upon treatment with NO donors confirmed the inhibitory role of NO on both the NF-kappaB DNA-binding property and its activation. Moreover, the inhibition of NF-kappaB nuclear translocation by NO donors directly correlated with the intracellular concentration of H(2)O(2) and was reversed by the addition of exogenous H(2)O(2). These findings show that NO might interfere with TNF-alpha-dependent NF-kappaB activation by interacting with O(2) and reducing the generation of H(2)O(2), a potent NF-kappaB activator. Therefore, NO-mediated disruption of NF-kappaB activation results in the removal of anti-apoptotic/resistance signals and sensitizes tumor cells to cytotoxic cytokines like TNF-alpha. PMID- 11118444 TI - Two mild cystic fibrosis-associated mutations result in severe cystic fibrosis when combined in cis and reveal a residue important for cystic fibrosis transmembrane conductance regulator processing and function. AB - The number of complex cystic fibrosis transmembrane conductance regulator (CFTR) genotypes identified as having double-mutant alleles with two mutations inherited in cis has been growing. We investigated the structure-function relationships of a severe cystic fibrosis (CF)-associated double mutant (R347H-D979A) to evaluate the contribution of each mild mutation to the phenotype. CFTR mutants expressed in HeLa cells were analyzed for protein biosynthesis and Cl(-) channel activity. Our data show that R347H is associated with mild defective Cl(-) channel activity and that the D979A defect leads to misprocessing. The mutant R347H-D979A combines both defects for a dramatic decrease in Cl(-) current. To decipher the molecular mechanism of this phenotype, single and double mutants with different charge combinations at residues 347 and 979 were constructed as charged residues were involved in this complex genotype. These studies revealed that residue 979, located in the third cytoplasmic loop, is critical for CFTR processing and Cl(-) channel activity highlighting the role of charged residues. These results have also important implications for CF, as they show that two mutations in cis can act in concert to alter dramatically CFTR function contributing to the wide phenotypic variability of CF disease. PMID- 11118443 TI - Nucleocytoplasmic shuttling of heterodimeric splicing factor U2AF. AB - The U2 small nuclear ribonucleoprotein auxiliary factor (U2AF) is a heterodimeric splicing factor composed of 65-kDa (U2AF(65)) and 35-kDa (U2AF(35)) subunits. The large subunit of U2AF recognizes the intronic polypyrimidine tract, a sequence located adjacent to the 3' splice site that serves as an important signal for both constitutive and regulated pre-mRNA splicing. The small subunit U2AF(35) interacts with the 3' splice site dinucleotide AG and is essential for regulated splicing. Like several other proteins involved in constitutive and regulated splicing, both U2AF(65) and U2AF(35) contain an arginine/serine-rich (RS) domain. In the present study we determined the role of RS domains in the subcellular localization of U2AF. Both U2AF(65) and U2AF(35) are shown to shuttle continuously between the nucleus and the cytoplasm by a mechanism that involves carrier receptors and is independent from binding to mRNA. The RS domain on either U2AF(65) or U2AF(35) acts as a nuclear localization signal and is sufficient to target a heterologous protein to the nuclear speckles. Furthermore, the results suggest that the presence of an RS domain in either U2AF subunit is sufficient to trigger the nucleocytoplasmic import of the heterodimeric complex. Shuttling of U2AF between nucleus and cytoplasm possibly represents a means to control the availability of this factor to initiate spliceosome assembly and therefore contribute to regulate splicing. PMID- 11118445 TI - CD45 is required for CD40-induced inhibition of DNA synthesis and regulation of c Jun NH2-terminal kinase and p38 in BAL-17 B cells. AB - Stimulation of B cell antigen receptor (BCR) may induce proliferation, differentiation, or apoptosis, depending upon the maturational stage of the cell and the presence or absence of signals transmitted via coreceptors. One such signal is delivered via CD40; for instance, ligation of CD40 rescues B cells from BCR-induced apoptosis. Here we show that, in contrast to WEHI-231 cells, CD40 ligation did not reverse BCR-induced growth inhibition in the BAL-17 mature B cell line and CD40 ligation itself inhibited proliferation. This inhibitory signaling was not observed in CD45-deficient cells. Further analyses demonstrate that transfection of dominant-negative form of SEK1 or treatment with SB203580 strongly reduced CD40-induced inhibition of BAL-17 proliferation, suggesting a requirement for c-Jun NH2-terminal kinase and p38 in CD40-induced inhibition of proliferation. Interestingly, CD40-initiated activation of c-Jun NH2-terminal kinase and p38 was enhanced and sustained in CD45-deficient cells, and these phenotypes were reversed by transfecting CD45 gene. However, CD40-mediated induction of cell surface molecules was not affected in CD45-deficient cells. Taken collectively, these results suggest that CD45 exerts a decisive effect on selective sets of CD40-mediated signaling pathways, dictating B cell fate. PMID- 11118446 TI - Molecular rulers: an assessment of distance and spatial relationships of Src tyrosine kinase Sh2 and active site regions. AB - The three-dimensional structures of the inactive conformations of Hck and Src, members of the Src protein-tyrosine kinase family, have recently been described. In both cases, the catalytic domain lies on the opposite face of the enzyme from the SH2 and SH3 domains. The active conformation of these enzymes has not yet been described. Given the known role of the SH2 and SH3 domains in promoting substrate binding, enzyme activation likely reorients the relative spatial arrangement between the SH2/SH3 domains and the active site region. We describe herein a series of "molecular rulers" and their use in assessing the topological and spatial relationships of the SH2 and active site regions of the Src protein tyrosine kinase. These synthetic compounds contain sequences that are active site directed (-Glu-Glu-Ile-Ile-(F(5))Phe-, where (F(5))Phe is pentafluorophenylalanine) and SH2-directed (-Tyr(P)-Glu-Glu-Ile-Glu-), separated by a sequence of variable length. The most potent bivalent compound, acetyl-Glu Glu-Leu-Leu-(F(5))Phe-(GABA)(3)-Tyr(P)-Glu-Glu-Ile-Glu-amide (where GABA is gamma aminobutyric acid), displays a >120-fold enhancement in inhibitory potency relative to the simple monovalent active site-directed species, acetyl-Glu-Glu Leu-Leu-(F(5))Phe-amide. The short linker length (3 GABA residues) between the active site- and SH2-directed peptide fragments suggests that the corresponding domains on the Src kinase can assume a nearly contiguous spatial arrangement in the active form of the enzyme. PMID- 11118447 TI - Self-association of the H3 region of syntaxin 1A. Implications for intermediates in SNARE complex assembly. AB - Intracellular membrane fusion requires SNARE proteins found on the vesicle and target membranes. SNAREs associate by formation of a parallel four-helix bundle, and it has been suggested that formation of this complex promotes membrane fusion. The membrane proximal region of the cytoplasmic domain of the SNARE syntaxin 1A, designated H3, contributes one of the four helices to the SNARE complex. In the crystal structure of syntaxin 1A H3, four molecules associate as a homotetramer composed of two pairs of parallel helices that are anti-parallel to each other. The H3 oligomer observed in the crystals is also found in solution, as assessed by gel filtration and chemical cross-linking studies. The crystal structure reveals that the highly conserved Phe-216 packs against conserved Gln-226 residues present on the anti-parallel pair of helices. Modeling indicates that Phe-216 prevents parallel tetramer formation. Mutation of Phe-216 to Leu appears to allow formation of parallel tetramers, whereas mutation to Ala destabilizes the protein. These results indicate that Phe-216 has a role in preventing formation of stable parallel helical bundles, thus favoring the interaction of the H3 region of syntaxin 1a with other proteins involved in membrane fusion. PMID- 11118448 TI - ERK5 and ERK2 cooperate to regulate NF-kappaB and cell transformation. AB - We have previously demonstrated an involvement of MEK5 and ERK5 in RafBXB stimulated focus formation in NIH3T3 cells. We find here that MEK5 and ERK5 cooperate with the RafBXB effectors MEK1/2 and ERK1/2 to induce foci. To further understand MEK5-ERK5-dependent signaling, we examined potential MEK5-ERK5 effectors that might influence focus-forming activity. Consistent with results from our focus-formation assays, constitutively active variants of MEK5 and MEK1 synergize to activate NF-kappaB, and MEK5 and ERK5 are required for activation of NF-kappaB by RafBXB. The MEK5-ERK5 pathway is also sufficient to activate both NF kappaB and p90 ribosomal S6 kinase. Our results support the hypothesis that NF kappaB and p90 ribosomal S6 kinase are involved in MEK5-ERK5-dependent focus formation and may serve as integration points for ERK5 and ERK1/2 signaling. PMID- 11118449 TI - Roles of glucitol in the GutR-mediated transcription activation process in Bacillus subtilis: tight binding of GutR to tis binding site. AB - Glucitol induction in Bacillus subtilis requires a transcription activator, GutR, and a sequence located upstream of the gut promoter. To understand the initial steps involved in the GutR-mediated transcription activation process and the physiological roles of glucitol, GutR was overproduced and purified. In the absence of glucitol, GutR exists as a monomer and binds directly to its binding site in the gut regulatory region. This binding site was mapped to a 29-base pair imperfect inverted repeat located between -78 and -50, and there is only one GutR binding site within the regulatory region. The kinetic parameters of the interaction between GutR and its binding site were monitored in real time using surface plasmon resonance. The half-life of the GutR-DNA complex in the absence of glucitol was estimated to be 6.8 min. In contrast, in the presence of glucitol, the half-life of the complex was extended to longer than 19 h by affecting only the off-rate but not the on-rate. This effect is glucitol specific. These data indicate that glucitol binds to GutR and induces GutR to have an extremely tight binding at its binding site. The physiological relevance of this process in transcription activation is discussed. PMID- 11118450 TI - Nam1p, a protein involved in RNA processing and translation, is coupled to transcription through an interaction with yeast mitochondrial RNA polymerase. AB - Alignment of three fungal mtRNA polymerases revealed conserved amino acid sequences in an amino-terminal region of the Saccharomyces cerevisiae enzyme implicated previously as harboring an important functional domain. Phenotypic analysis of deletion and point mutations, in conjunction with a yeast two-hybrid assay, revealed that Nam1p, a protein involved in RNA processing and translation in mitochondria, binds specifically to this domain. The significance of this interaction in vivo was demonstrated by the fact that the temperature-sensitive phenotype of a deletion mutation (rpo41Delta2), which impinges on this amino terminal domain, is suppressed by overproducing Nam1p. In addition, mutations in the amino-terminal domain result specifically in decreased steady-state levels of mature mitochondrial CYTB and COXI transcripts, which is a primary defect observed in NAM1 null mutant yeast strains. Finally, one point mutation (R129D) did not abolish Nam1p binding, yet displayed an obvious COX1/CYTB transcript defect. This mutation exhibited the most severe mitochondrial phenotype, suggesting that mutations in the amino-terminal domain can perturb other critical interactions, in addition to Nam1p binding, that contribute to the observed phenotypes. These results implicate the amino-terminal domain of mtRNA polymerases in coupling additional factors and activities involved in mitochondrial gene expression directly to the transcription machinery. PMID- 11118451 TI - Differential MAPK pathways utilized for HGF- and EGF-dependent renal epithelial morphogenesis. AB - Cells derived from the inner medullary collecting duct undergo in vitro branching tubulogenesis to both the c-met receptor ligand hepatocyte growth factor (HGF) as well as epidermal growth factor (EGF) receptor ligands. In contrast, many other cultured renal epithelial cells respond in this manner only to HGF, suggesting that these two receptors may use independent signaling pathways during morphogenesis. We have therefore compared the signaling pathways for mIMCD-3 cell morphogenesis in response to EGF and HGF. Inhibition of the p42/44 mitogen activated protein kinase (MAPK) pathway with the mitogen-activated protein kinase kinase (MKK1) inhibitor PD98059 (50 microm) markedly inhibits HGF-induced cell migration with only partial inhibition of EGF-induced cell motility. Similarly, HGF-dependent, but not EGF-dependent, branching morphogenesis was more greatly inhibited by the MKK1 inhibitor. Examination of EGF-stimulated cells demonstrated that extracellular-regulated kinase 5 (ERK5) was activated in response to EGF but not HGF, and that activation of ERK5 was only 60% inhibited by 50 microm PD98059. In contrast, the MKK inhibitor U0126 markedly inhibited both ERK1/2 and ERK5 activation and completely prevented HGF- and EGF-dependent migration and branching process formation. Expression of dominant negative ERK5 (dnBMK1) likewise inhibited EGF-dependent branching process formation, but did not affect HGF-dependent branching process formation. Our results indicate that activation of the ERK1/ERK2 signaling pathway is critical for HGF-induced cell motility/morphogenesis in mIMCD-3 cells, whereas ERK5 appears to be required for EGF-dependent morphogenesis. PMID- 11118452 TI - Entropic stabilization of the tryptophan synthase alpha-subunit from a hyperthermophile, Pyrococcus furiosus. X-ray analysis and calorimetry. AB - The structure of the tryptophan synthase alpha-subunit from Pyrococcus furiosus was determined by x-ray analysis at 2.0-A resolution, and its stability was examined by differential scanning calorimetry. Although the structure of the tryptophan synthase alpha(2)beta(2) complex from Salmonella typhimurium has been already determined, this is the first report of the structure of the alpha subunit alone. The alpha-subunit from P. furiosus (Pf-alpha-subunit) lacked 12 and 6 residues at the N and C termini, respectively, and one residue each in two loop regions as compared with that from S. typhimurium (St-alpha-subunit), resulting in the absence of an N-terminal helix and the shortening of a C terminal helix. The structure of the Pf-alpha-subunit was essentially similar to that of the St-alpha-subunit in the alpha(2)beta(2) complex. The differences between both structures were discussed in connection with the higher stability of the Pf-alpha-subunit and the complex formation of the alpha- and beta-subunits. Calorimetric results indicated that the Pf-alpha-subunit has extremely high thermostability and that its higher stability is caused by an entropic effect. On the basis of structural information of both proteins, we analyzed the contributions of each stabilization factor and could conclude that hydrophobic interactions in the protein interior do not contribute to the higher stability of the Pf-alpha-subunit. Rather, the increase in ion pairs, decrease in cavity volume, and entropic effects due to shortening of the polypeptide chain play important roles in extremely high stability in Pf-alpha-subunit. PMID- 11118453 TI - Activation of the cyclin-dependent kinase CTDK-I requires the heterodimerization of two unstable subunits. AB - RNA polymerase II CTD kinases are key elements in the control of mRNA synthesis. They constitute a family of cyclin-dependent kinases activated by C-type cyclins. Unlike most cyclin-dependent kinase complexes, which are composed of a catalytic and a regulatory subunit, the yeast CTD kinase I complex contains three specific subunits: a kinase subunit (Ctk1), a cyclin subunit (Ctk2), and a third subunit (Ctk3) of unknown function that does not exhibit any similarity to known proteins. Like the Ctk2 cyclin that is regulated at the level of protein turnover, Ctk3 is an unstable protein processed through a ubiquitin-proteasome pathway. Interestingly, Ctk2 and Ctk3 physical interaction is required to protect both subunits from degradation, pointing to a new mechanism for cyclin turnover regulation. We also show that Ctk2 and Ctk3 can each interact independently with the kinase. However, despite the formation of CDK/cyclin complexes in vitro, the Ctk2 cyclin is unable to activate its CDK: both Ctk2 and Ctk3 are required for Ctk1 CTD kinase activation. The different specific features governing CTDK-I regulation probably reflect requirement for the transcriptional response to multiple growth conditions. PMID- 11118454 TI - Identification of the calmodulin-binding domain of recombinant calcium independent phospholipase A2beta. implications for structure and function. AB - Calcium-independent phospholipase A(2) (iPLA(2)) is the major phospholipase A(2) activity in many cell types, and at least one isoform of this enzyme class is physically and functionally coupled to calmodulin (CaM) in a reversible calcium dependent fashion. To identify the domain in recombinant iPLA(2)beta (riPLA(2)beta) underlying this interaction, multiple techniques were employed. First, we identified calcium-activated CaM induced alterations in the kinetics of proteolytic fragment generation during limited trypsinolysis (i.e. CaM footprinting). Tryptic digests of riPLA(2)beta (83 kDa) in the presence of EGTA alone, Ca(+2) alone, or EGTA and CaM together resulted in the production of a major 68-kDa protein whose kinetic rate of formation was specifically attenuated in incubations containing CaM and Ca(+2) together. Western blotting utilizing antibodies directed against either the N- or C-terminal regions of riPLA(2)beta indicated the specific protection of riPLA(2)beta by calcium-activated CaM at a cleavage site approximately 15 kDa from the C terminus. Moreover, calcium activated calmodulin increased the kinetic rate of tryptic cleavage near the active site of riPLA(2)beta. Second, functional characterization of products from these partial tryptic digests demonstrated that approximately 90% of the 68-kDa riPLA(2)beta tryptic product (i.e. lacking the 15-kDa C-terminus) did not bind to a CaM affinity matrix in the presence of Ca(2+), although >95% of the noncleaved riPLA(2)beta as well as a 40-kDa C-terminal peptide bound tightly under these conditions. Third, when purified riPLA(2)beta was subjected to exhaustive trypsinolysis followed by ternary complex CaM affinity chromatography, a unique tryptic peptide ((694)AWSEMVGIQYFR(705)) within the 15-kDa C-terminal fragment was identified by RP-HPLC, which bound to CaM-agarose in the presence but not the absence of calcium ion. Fourth, fluorescence energy transfer experiments demonstrated that this peptide (694) bound to dansyl-calmodulin in a calcium dependent fashion. Collectively, these results identify multiple contact points in the 15-kDa C terminus as being the major but not necessarily the only binding site responsible for the calcium-dependent regulation of iPLA(2)beta by CaM. PMID- 11118455 TI - Mutations of the second extracellular loop of the human lutropin receptor emphasize the importance of receptor activation and de-emphasize the importance of receptor phosphorylation in agonist-induced internalization. AB - Alanine scanning mutagenesis of the second extracellular loop of the human lutropin receptor (hLHR) showed that mutation of most of the residues present in this region either enhance or impair the internalization of agonist. A more complete analysis of four mutants, two that enhanced internalization (F515A and T521A) and two that impaired internalization (S512A and V519A), showed that the two mutants that impaired internalization also show a decrease in the sensitivity for agonist-induced cAMP accumulation, whereas the two mutants that enhanced internalization show an increase in the sensitivity for agonist-induced cAMP accumulation. None of these mutants had an effect on the agonist-induced phosphorylation of the hLHR, however. We conclude that, in contrast to the prevailing view of the relative importance of receptor phosphorylation in the internalization of G protein-coupled receptors, the phosphorylation of the hLHR is less important than the agonist-induced activation of the hLHR in the process of internalization. PMID- 11118456 TI - Infrared spectroscopic characterization of the interaction of cationic lipids with plasmid DNA. AB - Fourier transform infrared spectroscopy was used to characterize the interaction of the cationic lipids 1,2-dioleoyl-3-trimethylammonium-propane and dioctadecyldimethylammonium bromide with plasmid DNA. The effect of incorporating the neutral colipids cholesterol and dioleoylphosphatidylethanolamine on this interaction was also examined. Additionally, dynamic and phase analysis light scattering were used to monitor the size and zeta potential of the resulting complexes under conditions similar to the Fourier transform infrared measurements. Results suggest that upon interaction of cationic lipids with DNA, the DNA remains in the B form. Distinct changes in the frequency of several infrared bands arising from the DNA bases, however, suggest perturbation of their hydration upon interaction with cationic lipids. A direct interaction of the lipid ammonium headgroup with and dehydration of the DNA phosphate is observed when DNA is complexed with these lipids. Changes in the apolar regions of the lipid bilayer are minimal, whereas the interfacial regions of the membrane show changes in hydration or molecular packing. Incorporation of helper lipids into the cationic membranes results in increased conformational disorder of the apolar region and further dehydration of the interfacial region. Changes in the hydration of the DNA bases were also observed as the molar ratio of helper lipid in the membranes was increased. PMID- 11118457 TI - Role of the substrate conformation and of the S1 protein in the cleavage efficiency of the T4 endoribonuclease RegB. AB - The T4 endoribonuclease RegB is involved in the inactivation of the phage early messengers. It cuts specifically in the middle of GGAG sequences found in early messenger intergenic regions but not GGAG sequences located in coding sequences or in late messengers. In vitro RegB activity is very low but is enhanced by a factor up to 100 by the ribosomal protein S1. In the absence of clear sequence motif distinguishing substrate and non-substrate GGAG-containing RNAs, we postulated the existence of a structural determinant. To test this hypothesis, we correlated the structure, probed by NMR spectroscopy, with the cleavage propensity of short RNA molecules derived from an artificial substrate. A kinetic analysis of the cleavage was performed in the presence and absence of S1. In the absence of S1, RegB efficiently hydrolyses substrates in which the last G of the GGAG motif is located in a short stem between two loops. Both strengthening and weakening of this structure strongly decrease the cleavage rate, indicating that this structure constitutes a positive cleavage determinant. Based on our results and those of others, we speculate that S1 favors the formation of the structure recognized by RegB and can thus be considered a "presentation protein." PMID- 11118458 TI - The structure of denatured alpha-lactalbumin elucidated by the technique of disulfide scrambling: fractionation of conformational isomers of alpha lactalbumin. AB - The structure of denatured alpha-lactalbumin (alpha-LA) has been characterized using the method of disulfide scrambling. Under denaturing conditions (urea, guanidine hydrochloride, guanidine thiocyanate, organic solvent or elevated temperature) and in the presence of thiol initiator, alpha-LA denatures by shuffling its four native disulfide bonds and converts to a mixture of fully oxidized scrambled structures. Analysis by reversed-phase HPLC reveals that the denatured alpha-LA comprises a minimum of 45 fractions of scrambled isomers. Among them, six well populated isomers have been isolated and structurally characterized. Their relative concentrations, which represent the fingerprinting of the denatured alpha-LA, vary substantially under different denaturing conditions. These results permit independent plotting of the denaturation and unfolding curves of alpha-LA. Most importantly, unique isomers of partially unfolded alpha-LA were shown to populate at mild and selected denaturing conditions. Organic solvent disrupts preferentially the hydrophobic alpha-helical domain, generating a predominant isomer containing two native disulfide bonds at the beta-sheet domain and two scrambled disulfide bonds at the alpha-helical region. Thermal denaturation selectively unfolds the beta-sheet domain of alpha LA, producing a prevalent isomer that exhibits structural characteristics of the molten globule state of alpha-LA. PMID- 11118459 TI - Crystal structure of Streptococcus pneumoniae N-acetylglucosamine-1-phosphate uridyltransferase bound to acetyl-coenzyme A reveals a novel active site architecture. AB - The bifunctional bacterial enzyme N-acetyl-glucosamine-1-phosphate uridyltransferase (GlmU) catalyzes the two-step formation of UDP-GlcNAc, a fundamental precursor in bacterial cell wall biosynthesis. With the emergence of new resistance mechanisms against beta-lactam and glycopeptide antibiotics, the biosynthetic pathway of UDP-GlcNAc represents an attractive target for drug design of new antibacterial agents. The crystal structures of Streptococcus pneumoniae GlmU in unbound form, in complex with acetyl-coenzyme A (AcCoA) and in complex with both AcCoA and the end product UDP-GlcNAc, have been determined and refined to 2.3, 2.5, and 1.75 A, respectively. The S. pneumoniae GlmU molecule is organized in two separate domains connected via a long alpha-helical linker and associates as a trimer, with the 50-A-long left-handed beta-helix (LbetaH) C terminal domains packed against each other in a parallel fashion and the C terminal region extended far away from the LbetaH core and exchanged with the beta-helix from a neighboring subunit in the trimer. AcCoA binding induces the formation of a long and narrow tunnel, enclosed between two adjacent LbetaH domains and the interchanged C-terminal region of the third subunit, giving rise to an original active site architecture at the junction of three subunits. PMID- 11118460 TI - Interim analyses in clinical trials: why do we plan them? PMID- 11118461 TI - Treatment of patients with metastatic renal carcinoma with a combination of subcutaneous interleukin-2 and interferon alfa with or without fluorouracil. Groupe Francais d'Immunotherapie, Federation Nationale des Centres de Lutte Contre le Cancer. AB - PURPOSE: Subcutaneous recombinant interleukin-2 (rIL-2) and recombinant interferon alfa-2a (rIFNalpha-2a) have been used extensively in the treatment of metastatic renal cancer. Most results, coming from noncontrolled phase II trials, showed inconsistent rates of response. More recently, the addition of fluorouracil (FU) was proposed to improve the efficacy of these regimens. PATIENTS AND METHODS: The role of a subcutaneous combination of rIL-2 and rIFNalpha-2a with or without FU was investigated. Patients were randomly assigned to receive a combination of rIL-2 and rIFNalpha-2a at weeks 1, 3, 5, and 7 or the same combination together with a continuous infusion of FU at weeks 1 and 5. The major end points of this multicenter, randomized trial were progression-free survival, response rate, and toxicity. Overall survival was a secondary end point. Tumor responses were reviewed by an independent committee. Analysis of the results was performed on an intention-to-treat basis. RESULTS: One hundred thirty one patients were enrolled. There was no difference in toxicity between the arms, and no toxic death was observed. One partial response was observed in arm A and five in arm B. Progression-free survival did not differ between the arms, and rates at 1 year were 12% and 15% in arms A and B, respectively. No statistically significant differences were detected in any end point. CONCLUSION: The subcutaneous combination of rIL-2 and rIFNalpha-2a with or without FU does not benefit patients with metastatic renal carcinoma. Neither of these regimens can be recommended as standard treatment. The results of the subcutaneous cytokine regimen seem disappointing. PMID- 11118463 TI - Received dose and dose-intensity of chemotherapy and outcome in nonmetastatic extremity osteosarcoma. European Osteosarcoma Intergroup. AB - PURPOSE: To examine the relationship between received dose, received dose intensity (RDI), and survival in patients with osteosarcoma. PATIENTS AND METHODS: Between 1983 and 1993, the European Osteosarcoma Intergroup (EOI) conducted two randomized trials involving patients with high-grade, nonmetastatic, biopsy-proven osteosarcoma of the extremity. These trials shared a common treatment arm of doxorubicin (DOX) 75 mg/m(2) and cisplatin (CDDP) 100 mg/m(2) planned for six cycles at 3-week intervals. Definitive surgery was scheduled at week 9, after three cycles. Survival time was calculated from 122 days, the scheduled end of chemotherapy. RESULTS: A total of 287 patients randomized to DOX/CDDP received at least one cycle of chemotherapy, and 232 (81%) received all six cycles. On average, 79% of the intended dose of DOX and 80% of the intended dose of CDDP was given. Mean time to completion of chemotherapy was 1.27 times that specified by the protocol. Mean RDI was 0.64 for DOX (SD = 0.19) and 0.65 for CDDP (SD = 0.18). Progression-free survival was lower for those who received one to five cycles compared with those who completed all six cycles (hazards ratio, 1.69; 95% confidence interval, 1.03 to 2.78). Survival and progression-free survival were lowest for patients with RDI less than 0.6, although these differences were not statistically significant at the 5% level. There was no clear evidence of preoperative dose or dose-intensity influencing histologic response. CONCLUSION: This analysis did not establish a clear survival benefit for increasing received dose or dose-intensity in the context of this two drug regimen. The hypothesis that increasing dose-intensity may improve survival in osteosarcoma requires prospective evaluation. PMID- 11118462 TI - Long-term outcome for patients with nonmetastatic osteosarcoma of the extremity treated at the istituto ortopedico rizzoli according to the istituto ortopedico rizzoli/osteosarcoma-2 protocol: an updated report. AB - PURPOSE: To provide an estimate of long-term prognosis for patients with osteosarcoma of the extremity treated in a single institution with neoadjuvant chemotherapy and observed for at least 10 years. PATIENTS AND METHODS: Patients with nonmetastatic osteosarcoma of the extremity were preoperatively treated with high-dose methotrexate, cisplatin, and doxorubicin (ADM). Postoperatively, good responders (90% or more tumor necrosis) received the same three drugs used before surgery, whereas poor responders (less than 90% tumor necrosis) received ifosfamide and etoposide in addition to those three drugs. RESULTS: For the 164 patients who entered the study between September 1986 and December 1989, surgery was a limb salvage in 136 cases (82%) and a good histologic response was observed in 117 patients (71%). At a follow-up ranging from 10 to 13 years (median, 11.5 years), 101 patients (61%) remained continuously free of disease, 61 relapsed, and two died of ADM-induced cardiotoxicity. There were no differences in prognosis between good and poor responding patients. ADM-induced cardiotoxicity (six patients), male infertility (10 of the 12 assessable patients), and second malignancies (seven patients) were the major complications of chemotherapy. Despite the large number of limb salvages performed, only four local recurrences (2.4%) were registered. CONCLUSION: With an aggressive neoadjuvant chemotherapy, it is possible to cure more than 60% of patients with nonmetastatic osteosarcoma of the extremity and amputation may be avoided in more than 80% of them. Because local or systemic relapses, myocardiopathies, and second malignancies are possible even 5 years or more after the beginning of treatment, a long-term follow-up is recommended for these patients. PMID- 11118464 TI - Phase II feasibility study of sequential couplets of Cisplatin/Topotecan followed by paclitaxel/cisplatin as primary treatment for advanced epithelial ovarian cancer: a National Cancer Institute of Canada Clinical Trials Group Study. AB - PURPOSE: Despite the improved results in advanced ovarian cancer achieved with the addition of paclitaxel to frontline therapy, there remains room for improvement. One approach is to add new agents such as topotecan. Because myelosuppression limits the delivery of topotecan with paclitaxel/cisplatin in a three-drug combination, we explored giving sequential couplets of cisplatin/topotecan followed by paclitaxel/cisplatin. PATIENTS AND METHODS: Forty four patients with residual epithelial ovarian carcinoma after primary surgery were studied. Cisplatin 50 mg/m(2) on day 1 and topotecan 0.75 mg/m(2) on days 1 through 5 were administered at 21-day intervals for four cycles, followed by interval debulking surgery (if optimal debulking was not achieved with primary surgery), and then paclitaxel 135 mg/m(2) over 24 hours on day 1 and cisplatin 75 mg/m(2) on day 2 at 21-day intervals for four cycles. RESULTS: Such sequential couplets are feasible. Myelotoxicity was the major toxic effect, but it was of short duration. The granulocyte nadir with topotecan/cisplatin occurred late (median, day 18), so retreatment on day 21 was not always possible. There was no unexpected nonhematologic toxicity. The regimen was active in this group of patients who had undergone largely suboptimal debulking surgery. In 34 patients with clinically measurable disease, the overall response rate was 78%, and 30 (77%) of the 39 patients with elevated CA 125 levels at baseline had normalization of CA 125 levels by the end of therapy. CONCLUSION: Sequential couplets of cisplatin/topotecan followed by paclitaxel/cisplatin are feasible. The efficacy data in this suboptimal group of patients has encouraged us to proceed with a randomized study based on this approach. PMID- 11118465 TI - Germline BRCA1/2 mutations and p27(Kip1) protein levels independently predict outcome after breast cancer. AB - PURPOSE: Decreased levels of the cyclin-dependent kinase inhibitor p27(Kip1) in breast cancer are associated with a poor outcome. The prognostic significance of BRCA1/2 mutations is less clear, and the relationship between BRCA1/2 mutation status, p27(Kip1) protein levels, and outcome has not been studied. PATIENTS AND METHODS: Pathology blocks from 202 consecutive Ashkenazi Jewish women with primary invasive breast cancer were studied. Tumor DNA was tested for the three common BRCA1/2 founder mutations present in Ashkenazi Jews, and p27(Kip1) expression was evaluated by immunohistochemistry. The median follow-up was 6.4 years. RESULTS: Thirty-two tumors (16%) were positive for a BRCA1/2 mutation. Low p27(Kip1) expression was seen in 110 tumors (63%) and was significantly associated with BRCA1/2 mutations (odds ratio, 4.0; 95% confidence interval [CI], 1.4 to 11.1; P =.009). BRCA1/2 mutation carriers had a significantly worse 5-year distant disease-free survival (DDFS) compared with women without BRCA1/2 mutations (58% v 82%; P =.003). Similar results were seen for women whose tumors expressed low levels of p27(Kip1), compared with those with high levels (5-year DDFS, 68% v 93%; P<.0001). In a multivariate analysis, both BRCA1/2 mutation and low p27(Kip1) expression were associated with a shorter DDFS (relative risk [RR], 2.1; 95% CI, 1.0 to 4.3; P =.05; and RR, 3.9; 95% CI, 1.4 to 11.1; P =.01, respectively). CONCLUSION: In this study, we showed that BRCA1/2 mutations were associated with low levels of p27(Kip1) in breast cancer. Both BRCA1/2 and p27(Kip1) status were identified as independent prognostic factors. PMID- 11118466 TI - Familial invasive breast cancers: worse outcome related to BRCA1 mutations. AB - PURPOSE: Although all studies confirm that BRCA1 tumors are highly proliferative and poorly differentiated, their outcomes remain controversial. We propose to examine, through a cohort study, the pathologic characteristics, overall survival, local recurrence, and metastasis-free intervals of 40 patients with BRCA1 breast cancer. PATIENTS AND METHODS: A cohort of 183 patients with invasive breast cancer, treated at the Institut Curie and presenting with a familial history of breast and/or ovarian cancer, were tested for BRCA1 germ-line mutation. Tumor characteristics and clinical events were extracted from our prospectively registered database. RESULTS: Forty BRCA1 mutations were found among the 183 patients (22%). Median follow-up was 58 months. BRCA1 tumors were larger in size (P =.03), had a higher rate of grade 3 histoprognostic factors (P =.002), and had a higher frequency of negative estrogen (P =.003) and progesterone receptors (P =.002) compared with non-BRCA1 tumors. Overall survival was poorer for carriers than for noncarriers (5-year rate, 80% v 91%, P =.002). Because a long time interval between cancer diagnosis and genetic counseling artificially increases survival time due to unrecorded deaths, the analysis was limited to the 110 patients whose diagnosis-to-counseling interval was less than 36 months (19 BRCA1 patients and 91 non-BRCA1 patients). The differences between the BRCA1 and non-BRCA1 groups regarding overall survival and metastasis-free interval were dramatically increased (49% v 85% and 18% v 84%, respectively). Multivariate analysis showed that BRCA1 mutation was an independent prognostic factor. CONCLUSION: Our results strongly support that among patients with familial breast cancer, those who have a BRCA1 mutation have a worse outcome than those who do not. PMID- 11118467 TI - Symptoms of posttraumatic stress in young adult survivors of childhood cancer. AB - PURPOSE: This study assessed the prevalence of posttraumatic stress symptoms in young adult survivors of childhood cancer and the association of posttraumatic stress with anxiety, adjustment, perceptions of illness and treatment, and medical data extracted from oncology records. PATIENTS AND METHODS: Seventy-eight young adults (ages 18 to 40 years) who had been treated for childhood cancer completed questionnaires and psychiatric interviews assessing posttraumatic stress, anxiety, perceptions of their illness and treatment, and symptoms of psychologic distress. Data on treatment intensity and severity of medical late effects were collected via chart review. RESULTS: Of the patient sample, 20.5% met American Psychiatric Association Diagnostic and Statistical Manual criteria for posttraumatic stress disorder (PTSD) at some point since the end of their treatment. Clinically significant levels of intrusive (9%) and avoidant (16.7%) symptoms were reported. Participants also reported elevated state and trait anxiety. Participants with PTSD reported higher perceived current life threat, more intense treatment histories, and higher (and clinically significant) levels of psychologic distress than those who did not have PTSD. CONCLUSION: One-fifth of this sample of young adult survivors of childhood cancer met criteria for a diagnosis of PTSD, with clinically significant symptoms of intrusion and avoidance reported. As in other samples, PTSD in young adult survivors was associated with anxiety and other psychologic distress. Survivors' perceptions of treatment and its effects were more highly associated with posttraumatic stress than were more objective medical data. The data suggest that cancer-related posttraumatic stress may emerge in young adulthood and may affect the achievement of developmental milestones and orientation toward health care. PMID- 11118468 TI - Quantitative tumor cell content of bone marrow and blood as a predictor of outcome in stage IV neuroblastoma: a Children's Cancer Group Study. AB - PURPOSE: This study investigated the prognostic value of quantifying tumor cells in bone marrow and blood by immunocytology in children with high-risk, metastatic neuroblastoma. PATIENTS AND METHODS: Patients with stage IV neuroblastoma (N = 466) registered on Children's Cancer Group study 3891 received five cycles of induction chemotherapy and were randomized either to myeloablative chemoradiotherapy with autologous purged bone marrow rescue or to nonmyeloablative chemotherapy. Subsequently, they were randomized to 13-cis retinoic acid or no further treatment. Immunocytologic analyses of bone marrow and blood were performed at diagnosis, week 4, week 12, bone marrow collection, and end induction and were correlated with tumor biology, clinical variables, treatment regimen, and event-free survival (EFS). RESULTS: Immunocytology identified neuroblastoma cells in bone marrow of 81% at diagnosis, 55% at 4 weeks, 27% at 12 weeks, 19% at bone marrow collection, and 14% at end induction. Tumor cells were detected in blood of 58% at diagnosis and 5% at collection. There was an adverse effect on EFS of increasing tumor cell concentration in bone marrow at diagnosis (P =.04), at 12 weeks (P =.006), at bone marrow collection (P <.001), and at end induction (P =.07). Positive blood immunocytology at diagnosis was associated with decreased EFS (P: =.003). The prognostic impact of immunocytology was independent of morphologically detected bone marrow disease, MYCN status, and serum ferritin level in bivariate Cox analyses. CONCLUSION: Immunocytologic quantification of neuroblastoma cells in bone marrow and blood at diagnosis and in bone marrow during induction chemotherapy provides prognostic information that can identify patients with very high-risk disease who should be considered for experimental therapy that might improve outcome. PMID- 11118469 TI - Phase I study of chimeric human/murine anti-ganglioside G(D2) monoclonal antibody (ch14.18) with granulocyte-macrophage colony-stimulating factor in children with neuroblastoma immediately after hematopoietic stem-cell transplantation: a Children's Cancer Group Study. AB - PURPOSE: Ganglioside G(D2) is strongly expressed on the surface of human neuroblastoma cells. It has been shown that the chimeric human/murine anti-G(D2) monoclonal antibody (ch14.18) can induce lysis of neuroblastoma cells by antibody dependent cellular cytotoxicity and complement-dependent cytotoxicity. The purposes of the study were (1) to determine the maximum-tolerated dose (MTD) of ch14.18 in combination with standard dose granulocyte-macrophage colony stimulating factor (GM-CSF) for patients with neuroblastoma who recently completed hematopoietic stem-cell transplantation (HSCT), and (2) to determine the toxicities of ch14.18 with GM-CSF in this setting. PATIENTS AND METHODS: Patients became eligible when the total absolute phagocyte count (APC) was greater than 1, 000/microL after HSCT. ch14.18 was infused intravenously over 5 hours daily for 4 consecutive days. Patients received GM-CSF 250 microg/m(2)/d starting at least 3 days before ch14.18 and continued for 3 days after the completion of ch14.18. The ch14.18 dose levels were 20, 30, 40, and 50 mg/m(2)/d. In the absence of progressive disease, patients were allowed to receive up to six 4-day courses of ch14.18 therapy with GM-CSF. Nineteen patients with neuroblastoma were treated. RESULTS: A total of 79 courses were administered. No toxic deaths occurred. The main toxicities were severe neuropathic pain, fever, nausea/vomiting, urticaria, hypotension, mild to moderate capillary leak syndrome, and neurotoxicity. Three dose-limiting toxicities were observed among six patients at 50 mg/m(2)/d: intractable neuropathic pain, grade 3 recurrent urticaria, and grade 4 vomiting. Human antichimeric antibody developed in 28% of patients. CONCLUSION: ch14.18 can be administered with GM-CSF after HSCT in patients with neuroblastoma with manageable toxicities. The MTD is 40 mg/m(2)/d for 4 days when given in this schedule with GM-CSF. PMID- 11118470 TI - Phase I and pharmacokinetic study of irofulven, a novel mushroom-derived cytotoxin, administered for five consecutive days every four weeks in patients with advanced solid malignancies. AB - PURPOSE: To evaluate the toxicity and pharmacologic behavior of the novel mushroom-derived cytotoxin irofulven administered as a 5-minute intravenous (IV) infusion daily for 5 days every 4 weeks to patients with advanced solid malignancies. PATIENTS AND METHODS: In this phase I trial, 46 patients were treated with irofulven doses ranging from 1.0 to 17.69 mg/m(2) as a 5-minute IV infusion (two patients received a 1-hour infusion) daily for 5 days every 4 weeks. The modified continual reassessment method was used for dose escalation. Pharmacokinetic studies were performed on days 1 and 5 to characterize the plasma disposition of irofulven. RESULTS: Forty-six patients were treated with 92 courses of irofulven. The dose-limiting toxicities on this schedule were myelosuppression and renal dysfunction. At the 14.15-mg/m(2) dose level, renal dysfunction resembling renal tubular acidosis occurred in four of 10 patients and was ameliorated by prophylactic IV hydration. The 17.69-mg/m(2) dose level was not tolerated because of grade 4 neutropenia and renal toxicity, whereas the 14.15-mg/m(2) dose level was not tolerable with repetitive dosing because of persistent thrombocytopenia. Other common toxicities included mild to moderate nausea, vomiting, facial erythema, and fatigue. One partial response occurred in a patient with advanced, refractory metastatic pancreatic cancer lasting 7 months. Pharmacokinetic studies of irofulven revealed dose-proportional increases in both maximum plasma concentrations and area under the concentration-time curve, while the agent exhibited a rapid elimination half-life of 2 to 10 minutes. CONCLUSION: Given the results of this study, the recommended dose of irofulven is 10.64 mg/m(2) as a 5-minute IV infusion daily for 5 days every 4 weeks. The preliminary antitumor activity documented in a patient with advanced pancreatic cancer and the striking preclinical antitumor effects of irofulven observed on intermittent dosing schedules support further disease-directed evaluations of this agent on the schedule evaluated in this study. PMID- 11118471 TI - Phase I dose-finding study of a new taxane, RPR 109881A, administered as a one hour intravenous infusion days 1 and 8 to patients with advanced solid tumors. AB - PURPOSE: To define the maximum-tolerated dose, recommended phase II dose (RD), dose-limiting toxicity (DLT), and pharmacokinetics of a novel taxane, RPR 109881A, administered on days 1 and 8 of a 21-day cycle. PATIENTS AND METHODS: Twenty-nine patients were enrolled and treated according to a modified continual reassessment method from a starting dose of 7.5 mg/m(2) to 52.5 mg/m(2). Detailed pharmacokinetic analyses of blood and urine were performed on days 1 and 8 of the first cycle. Toxicity was monitored weekly. RESULTS: DLT consisting of grade 3 or 4 diarrhea was seen in three of six patients at 52.5 mg/m(2). Grade 3 or 4 granulocytopenia was also seen in five of six patients treated at this dose (four of six in the first cycle). At the next lower dose level, 45 mg/m(2) toxicity was moderate, with only one of 12 patients experiencing severe diarrhea and grade 4 granulocytopenia with associated infection. Drug concentrations were consistent with a three-compartment open model. The total-body clearance suggests a linear dose-concentration relationship. RPR 109881A has a high clearance (mean, 42.6 L/h/m(2)), a large volume of distribution (mean, 952 L/m(2)), and a long terminal half-life (mean, 24 hours). There was no drug accumulation between days 1 and 8. One partial response was seen in a patient with renal cell carcinoma. CONCLUSION: The RD of RPR 109881A given as a 1-hour infusion on days 1 and 8 of a 21-day cycle is 45 mg/m(2). At this dose the drug is well tolerated and should be further studied. PMID- 11118472 TI - Clarification of anastrozole/megestrol acetate trial program design. PMID- 11118473 TI - Sensitive nonradioactive detection of mRNA in tissue sections: novel application of the whole-mount in situ hybridization protocol. AB - The relative insensitivity of nonradioactive mRNA detection in tissue sections compared to the sensitive nonradioactive detection of single-copy DNA sequences in chromosome spreads, or of mRNA sequences in whole-mount samples, has remained a puzzling issue. Because of the biological significance of sensitive in situ mRNA detection in conjunction with high spatial resolution, we developed a nonradioactive in situ hybridization (ISH) protocol for detection of mRNA sequences in sections. The procedure is essentially based on the whole-mount ISH procedure and is at least equally sensitive. Increase of the hybridization temperature to 70C while maintaining stringency of hybridization by adaptation of the salt concentration significantly improved the sensitivity and made the procedure more sensitive than the conventional radioactive procedure. Thicker sections, which were no improvement using conventional radioactive ISH protocols, further enhanced signal. Higher hybridization temperatures apparently permit better tissue penetration of the probe. Application of this highly reliable protocol permitted the identification and localization of the cells in the developing heart that express low-abundance mRNAs of different members of the Iroquois homeobox gene family that are supposedly involved in cardiac patterning. The radioactive ISH procedure scarcely permitted detection of these sequences, underscoring the value of this novel method. PMID- 11118474 TI - Cell-specific processing of chromogranin A in endocrine cells of the rat stomach. AB - The rat stomach is rich in endocrine cells. The acid-producing (oxyntic) mucosa contains ECL cells, A-like cells, and somatostatin (D) cells, and the antrum harbours gastrin (G) cells, enterochromaffin (EC) cells and D cells. Although chromogranin A (CgA) occurs in all these cells, its processing appears to differ from one cell type to another. Eleven antisera generated to different regions of rat CgA, two antisera generated to a human (h) CgA sequences, and one to a bovine (b) CgA sequence, respectively, were employed together with antisera directed towards cell-specific markers such as gastrin (G cells), serotonin (EC cells), histidine decarboxylase (ECL cells) and somatostatin (D cells) to characterize the expression of CgA and CgA-derived peptides in the various endocrine cell populations of the rat stomach. In the oxyntic mucosa, antisera raised against CgA(291-319) and CGA(316-321) immunostained D cells exclusively, whereas antisera raised against bCgA(82-91) and CgA(121-128) immunostained A-like cells and D cells. Antisera raised against CgA(318-349) and CgA(437-448) immunostained ECL cells and A-like cells, but not D cells. In the antrum, antisera against CgA(291 319) immunostained D cells, and antisera against CgA(351-356) immunostained G cells. Our observations suggest that each individual endocrine cell type in the rat stomach generates a unique mixture of CgA-derived peptides, probably reflecting cell-specific differences in the post-translational processing of CgA and its peptide products. A panel of antisera that recognize specific domains of CgA may help to identify individual endocrine cell populations. PMID- 11118475 TI - Lectin and proteoglycan histochemistry of feline pacinian corpuscles. AB - We studied carbohydrate residues of glycoproteins and proteoglycans (PGs) in peritoneal Pacinian corpuscles of five adult cats. Terminal monosaccharides of glycoproteins and related polysaccharides were identified by lectin histochemistry and the PGs and glycosaminoglycans (GAGs) by specific antibodies. The most intensive lectin staining reactions indicated an abundance of glycoconjugates with terminal mannose (Man) or sialic acid residues, but no complex-type oligosaccharides were detected within the corpuscles. Terminal fucose (Fuc) and galactose (Gal) residues typical for O-linked mucin-type glycoproteins generally associated with high water binding capacity were also absent. Antibodies against unsulfated chondroitin (C-0-S), chondroitin-4-sulfate (C-4-S), and decorin showed positive reactions in the interfibrillar spaces between the lamellae, around collagen fibers, and around the lamellae of the perineural capsule, especially in the outer parts known to contain Type II collagen. Biglycan showed a preference for the innermost part of the perineural capsule (intermediate layer), known to contain Type V collagen. Collagen V and biglycan are both linked to growth processes. Hyaluronic acid (HA), chondroitin-6 sulfate (C-6-S) chains, and a chondroitin sulfate proteoglycan (CSPG) were co localized in the terminal glia. The study of carbohydrates with high water binding capacity may contribute to our understanding of the high viscoelasticity of Pacinian corpuscles. PMID- 11118476 TI - Demonstration of acetylcholinesterase molecular forms in a continuous tubular lysosomal system of rat pancreatic acinar cells. AB - By applying the highly sensitive cytochemical Gautron's technique, we were able to reveal AChE activity in rat pancreatic acinar cells, particularly at the level of a complex membrane-bound network formed by tubules with varicosities located around the nuclei and close to the basolateral membrane. The Golgi apparatus was devoid of cytochemical reaction beside the trans-Golgi network cisternae, which showed a positive reaction. The RER of some acinar cells also presented a signal, demonstrating their capability of synthesizing AChE. Immunogold using a specific anti-AChE antibody yielded similar results. Double-labeling experiments corroborated the presence of enzyme cytochemical and immunocytochemical signals in the same lysosomal tubular network. Biochemical sedimentation assays confirmed the presence of AChE in acinar cells, which exists as two globular molecular forms, G(1) and G(4). These results were obtained with pancreatic tissue in situ as well as with isolated acinar cells maintained in culture and devoid of neural elements. The existence of a continuous tubular lysosomal network containing AChE is in agreement with previous reports on acinar and other cell types, and supports a more general hypothesis on dynamic continuities among cell structures. Whether AChE is being secreted by the acinar cells or internalized through this endo-lysosomal system was not defined. However, the capability of the acinar cells to synthesize AChE and to channel it through a tubular system is a good indication that the cells can modulate their cholinergic stimulation for optimal secretion of digestive enzymes. PMID- 11118477 TI - Expression of the beta6 integrin subunit is associated with sites of neutrophil influx in lung epithelium. AB - Inhalation of ozone by Rhesus monkeys results in epithelial injury and granulocyte influx in both conducting airways and respiratory bronchioles. We have reported that ozone-induced neutrophil recruitment and subsequent epithelial repair can be inhibited in vivo with a CD18 antibody. The antibody-mediated effect is abrogated by local instillation of C5a (a CD18-independent neutrophil chemoattractant), thereby demonstrating a role for neutrophils in lung epithelial repair processes. As an extension of this study, we examined the effect of ozone and neutrophil influx on epithelial expression of the beta6 integrin, an adhesion molecule associated with proliferation and repair. Expression of beta6 integrin was determined by immunohistochemistry for ozone-exposed monkeys treated with either control immunoglobulins or a CD18 antibody. The tracheal epithelium of ozone-exposed monkeys treated with control immunglobulins expressed the beta6 integrin. In contrast, the tracheal epithelium of ozone-exposed monkeys treated with CD18 antibody exhibited very low to undetectable expression of beta6 integrin. In association with C5a instillation and neutrophil influx, beta6 integrin was also observed in respiratory bronchiolar epithelium from both control and ozone-exposed animals. These findings cumulatively suggest that lung epithelial cell expression of beta6 integrin is associated with sites of neutrophil recruitment. PMID- 11118478 TI - Associations of PKC isoforms with the cytoskeleton of B16F10 melanoma cells. AB - Although PKC plays a major role in regulating the morphology and function of the cytoskeleton, little is known about in situ associations of specific isoforms with the cytoskeleton. We demonstrate that seven PKC isoforms are expressed in B16F10 melanoma cells and show different levels of induction by serum. Using cell cytoskeleton preparations (CSKs), confocal microscopy, and immunocytochemistry, all isoforms show specific patterns of localization to focal contact-like structures (alpha, delta), very small cytoplasmic granules/vesicles (all isoforms), dense ordered arrays of small granules in the perinuclear region (alpha, delta), granules/vesicles associated with a homogeneous framework in the cytoplasm adjacent to the nucleus (gamma), or irregular-shaped patches of granules at or near the nuclear perimeter (eta, theta). In addition, several isoforms are present as cytoplasmic granules/ vesicles in linear or curvilinear arrays (alpha, delta, epsilon, theta). When isoform localization is examined using 3.7% formaldehyde or methanol:acetone, the patterns of localization in CSKs are often difficult or impossible to detect, and many are described here for the first time. Double-labeling experiments with CSK demonstrate that PKC actin co localizes with punctate alpha-rich particles above the nucleus, granules of epsilon throughout the cytoplasm, and with theta in irregular-shaped aggregates associated with the nucleus. Vimentin co-localizes with perinuclear granules of delta and beta(2), and alpha-tubulin co-localizes with theta in structures at or near the nuclear surface and in microtubules associated with the microtubule organizing center (MTOC). In summary, the present study demonstrates that seven PKC isoforms are endogenously expressed in B16F10 melanoma cells. These isoforms show various levels of induction by serum and specific patterns of association with various components of the detergent-resistant cell cytoskeleton. PMID- 11118479 TI - Mucin MUC1 is seen in cell surface protrusions together with ezrin in immunoelectron tomography and is concentrated at tips of filopodial protrusions in MCF-7 breast carcinoma cells. AB - MUC1, a transmembrane member of the mucin family, is believed to have anti adhesive properties because of its highly sialylated, extended, and rigid rod like conformation. The ERM proteins (ezrin, radixin, and moesin) function as membrane-cytoskeletal linkers. MUC1 and ezrin are enriched in microvilli in MCF 7az breast carcinoma cells. Similar localization was also found in peripheral membrane areas and in filopodium-like protrusions. Whereas ezrin was consistently detected in the cell-cell contact region, MUC1 was less frequently found there. MUC1 was distinctly expressed in long filopodial protrusions and was highly concentrated at their tips, which also contained ezrin, whereas F-actin was found along the stalk. This localization of MUC1 suggests a role for MUC1 in transient cell structures of migrating cells and transient cell adhesion. No direct association has yet been found between MUC1 and ezrin. However, both MUC1 and ezrin had a similar overall distribution pattern in microvilli and filopodium like protrusions in immunoelectron tomography. In addition, MUC1 and ezrin showed spatial association, because several 10-nm gold particles used to decorate ezrin were seen in the vicinity close to the clusters of 5-nm gold particles decorating MUC1. Therefore, MUC1 appears to be associated with ezrin, but the nature of this association requires further study. PMID- 11118480 TI - Time course of osteopontin, osteocalcin, and osteonectin accumulation and calcification after acute vessel wall injury. AB - Although mineral deposits have long been described to be a prominent feature of atherosclerosis, the mechanisms of arterial calcification are not well understood. However, accumulation of the non-collagenous matrix bone-associated proteins, osteopontin, osteocalcin, and osteonectin, has been demonstrated in atheromatous plaques. The aim of this study was to evaluate the role of these proteins in arterial calcification and, more precisely, during the initiation of this process. A model of rapid aortic calcification was developed in rabbits by an oversized balloon angioplasty. Calcification was followed using von Kossa staining and osteopontin, osteocalcin, and osteonectin were identified using immunohistochemistry. The aortic injury was rapidly followed by calcified deposits that appeared in the media as soon as 2 days after injury and then accumulated in zipper-like structures. Osteonectin was not detected in calcified deposits at any time after injury. In contrast, osteopontin and osteocalcin were detected in 8- and 14-day calcified structures, respectively, but not in the very early 2-day mineral deposits. These results suggest that these matrix proteins, osteopontin, osteocalcin, and osteonectin, are not involved in the initiation step of the aortic calcification process and that the former two might play a role in the regulation of arterial calcification. PMID- 11118481 TI - Differential subcellular localization of zinc in the rat retina. AB - In the retina, zinc is believed to be a modulator of synaptic transmission and a constituent of metalloenzymes. To determine whether the intracellular localization of zinc correlates with function, we examined the localization of endogenous zinc in the rat retina using the silver amplification method. By light microscopy, reaction products were detected in the pigment epithelial cells (PE), the inner segment of photoreceptors (IS), the outer nuclear layer (ONL) and the inner nuclear layer (INL), the outer plexiform layer (OPL) and the inner plexiform layer (IPL), and the ganglion cell layer (GC). The heaviest accumulation of precipitate was observed in PE and IS, whereas only a little precipitate was found in GC. When the intracellular zinc was chelated with diethyldithiocarbamate, a small amount of precipitate was observed only in ONL. By electron microscopy, zinc was associated with three compartments. In OPL and IPL, zinc was associated with neural processes, while in PE, IS, INL, and GC it was associated with the Golgi apparatus. In ONL, zinc was associated with the nucleus. Zinc in the neural processes is believed to act as a modulator of synaptic transmission, and zinc associated with the Golgi apparatus is assumed to catalyze metalloenzyme reactions. PMID- 11118482 TI - Utilizing the peptidyl-prolyl cis-trans isomerase pin1 as a probe of its phosphorylated target proteins. Examples of binding to nuclear proteins in a human kidney cell line and to tau in Alzheimer's diseased brain. AB - The human parvulin Pin1 is a member of the peptidyl-prolyl cis-trans isomerase group of proteins, which modulate the assembly, folding, activity, and transport of essential cellular proteins. Pin1 is a mitotic regulator interacting with a range of proteins that are phosphorylated before cell division. In addition, an involvement of Pin1 in the tau-related neurodegenerative brain disorders has recently been shown. In this context, Pin1 becomes depleted from the nucleus in Alzheimer's disease (AD) neurons when it is redirected to the large amounts of hyperphosphorylated tau associated with the neurofibrillary tangles. This depletion from the nucleus may ultimately contribute to neuron cell death. Recently we have devised a novel methodology in which exogenous Pin1 is used as a TEM probe for its target proteins. Here we extend this methodology to provide further evidence that Pin1 binds at enhanced levels to mitotic nuclear proteins and to hyperphosphorylated tau in AD brain. We suggest that exogenous Pin1 labeling can be used to elucidate the phosphorylation status of its target proteins in general and could specifically provide important insights into the development of tau-related neurodegenerative brain disorders. PMID- 11118483 TI - In vivo reactivation of DNases in implanted human prostate tumors after administration of a vitamin C/K(3) combination. AB - Human prostate cancer cells (DU145) implanted into nude mice are deficient in DNase activity. After administration of a vitamin C/vitamin K(3) combination, both alkaline DNase (DNase I) and acid DNase (DNase II) activities were detected in cryosections with a histochemical lead nitrate technique. Alkaline DNase activity appeared 1 hr after vitamin administration, decreased slightly until 2 hr, and disappeared by 8 hr after treatment. Acid DNase activity appeared 2 hr after vitamin administration, reached its highest levels between 4 and 8 hr, and maintained its activity 24 hr after treatment. Methyl green staining indicated that DNase expression was accompanied by a decrease in DNA content of the tumor cells. Microscopic examination of 1-microm sections of the tumors indicated that DNase reactivation and the subsequent degradation of DNA induced multiple forms of tumor cell death, including apoptosis and necrosis. The primary form of vitamin-induced tumor cell death was autoschizis, which is characterized by membrane damage and the progressive loss of cytoplasm through a series of self excisions. These self-excisions typically continue until the perikaryon consists of an apparently intact nucleus surrounded by a thin rim of cytoplasm that contains damaged organelles. PMID- 11118484 TI - Expression and role of vascular endothelial growth factor in liver regeneration after partial hepatectomy in rats. AB - Vascular endothelial growth factor (VEGF) plays a major role in angiogenesis, which is essential for both healing of injured tissue and proliferation of carcinoma cells. In this study we elucidated the expression and role of VEGF in rat liver regeneration after partial hepatectomy. VEGF expression was mainly detected in periportal hepatocytes and reached a maximal level 48-72 hr after partial hepatectomy by both immunohistochemistry and in situ hybridization. Similarly, immunohistochemistry for Ki-67 showed that the proliferative activity of sinusoidal endothelial cells was highest in the periportal area and reached a maximal level 72 hr after partial hepatectomy. Moreover, neutralization of VEGF significantly inhibited proliferative activity of hepatocytes (p<0. 0001), as well as sinusoidal endothelial cells (p<0.001), at 48 and 96 hr after partial hepatectomy. Conversely, injection of VEGF significantly promoted proliferative activity of hepatocytes (p<0. 0001) as well as sinusoidal endothelial cells (p<0.0005) at 48 hr after partial hepatectomy. These results suggest that VEGF promotes proliferation of hepatocytes through reconstruction of liver sinusoids by proliferation of sinusoidal endothelial cells. Furthermore, these data point to a new therapeutic strategy, the use of VEGF and other hepatocyte growth factors in fulminant or severe acute hepatitis. PMID- 11118485 TI - Development and characterization of 1C6-203, a new monoclonal antibody specific to human thymidine phosphorylase. AB - Thymidine phosphorylase (dThdPase) is an essential enzyme for activation of the oral cytostatic drug capecitabine and its intermediate metabolite, doxifluridine, to 5-fluorouracil in tumors. Methods to estimate dThdPase expression in tumor tissue might be useful to predict the efficacy of capecitabine and doxifluridine in cancer patients. We established a new monoclonal antibody (MAb), 1C6-203, applicable for dThdPase immunohistochemistry and compared its staining characteristics with those of a previously established MAb, 654-1. In 4% paraformaldehyde-fixed colorectal carcinoma, 1C6-203 and 654-1 stained cancer cells in 19/30 and 9/30 patients, respectively. In 10% formalin-fixed colorectal carcinoma, 1C6-203 and 654-1 stained cancer cells in 18/30 and 6/30 patients, respectively. In negative 10% formalin-fixed tissues, microwave treatment improved the positivity of 654-1-stained cancer cells. These results suggest that an epitope recognized by 1C6-203 is resistant to epitope masking by formaldehyde fixation, whereas that for MAb 654-1 is sensitive. Therefore, MAb 1C6-203 might be more suitable than MAb 654-1 for evaluating dThdPase expression in colorectal carcinoma. PMID- 11118486 TI - Calponin--knocked out but not down! PMID- 11118487 TI - Actions of hypoxia on catecholamine synthetic enzyme mRNA expression before and after development of adrenal innervation in the sheep fetus. AB - We have investigated adrenal mRNA expression of the catecholamine synthetic enzymes tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT) following acute hypoxia in fetal sheep before (< 105 days gestation, n = 20) and after (> 125 days gestation, n = 20) the development of adrenal innervation and following pretreatment with the nicotinic receptor anatgonist hexamethonium (n = 12). Total RNA was extracted from fetal adrenal glands collected at specific time points at 3-20 h after the onset of either hypoxia ( approximately 50% reduction in fetal arterial oxygen saturation (SO2) for 30 min), or normoxia. Before 105 days, there was a decrease in adrenal TH mRNA expression at 20 h after hypoxia and adrenal TH mRNA expression was directly related to the changes in arterial PO2 measured during normoxia and hypoxia. After 125 days, adrenal TH mRNA levels were suppressed for up to 12 h following hypoxia. In both age groups, adrenal PNMT mRNA expression increased at 3-5 h after hypoxia and was inversely related to the changes in fetal arterial PO2 during normoxia or hypoxia. After 125 days, the administration of hexamethonium (25 mg kg(-1), I.V.) reduced TH mRNA but not PNMT mRNA expression after normoxia. After hexamethonium pretreatment, there was no significant change in either adrenal TH or PNMT mRNA expression following hypoxia. We conclude that acute hypoxia differentially regulates adrenal TH and PNMT mRNA expression in the fetal sheep both before and after the development of adrenal innervation. After the development of adrenal innervation, however, the effect of acute hypoxia upon adrenal TH and PNMT mRNA expression is dependent upon neurogenic input acting via nicotinic receptors. PMID- 11118488 TI - A sodium channel mutation causing epilepsy in man exhibits subtle defects in fast inactivation and activation in vitro. AB - Generalized epilepsy with febrile seizures plus (GEFS+) is a benign epileptic syndrome of humans. It is characterized by febrile and afebrile generalized seizures that occur predominantly in childhood and respond well to standard antiepileptic therapy. A mutation in the b1-subunit of the voltage-gated sodium channel, linked to chromosome 19q13 (GEFS+ type 1) has been found in one family. For four other families, linkage was found to chromosome 2q21-33 (GEFS+ type 2) where three genes encoding neuronal sodium channel a-subunits are located (SCN1 3A). Recently, the first two mutations were identified in SCN1A. We introduced one of these mutations, which is highly conserved to SCN1A, into the cDNA of the gene SCN4A encoding the a-subunit of the human skeletal muscle sodium channel (hSkm1). The mutation is located in the S4 voltage sensor of domain IV, predicting substitution of histidine for the fifth of eight arginines (R1460H in hSkm1). Functional studies were performed by expressing the a-subunit alone in the mammalian tsA201 cell line using the whole-cell patch clamp technique. Compared to wild-type (WT), mutant R1460H channels showed small defects in fast inactivation. The time course of inactivation was slightly (1.5-fold) slowed and its voltage dependence reduced, and recovery from inactivation was accelerated 3 fold. However, there was no increase in persistent sodium current as observed for SCN4A mutations causing myotonia or periodic paralysis. The activation time course of R1460H channels was slightly accelerated. Slow inactivation was slightly but significantly stabilized, confirming the importance of this region for slow inactivation. The combination of activation and fast inactivation defects can explain the occurrence of epileptic seizures, but the effects were much more subtle than the inactivation defects described previously for mutations in SCN4A causing disease in skeletal muscle. Hence, with regard to pathological excitability, our results suggest a greater vulnerability of the central nervous system compared to muscle tissue. PMID- 11118489 TI - Quantification of calcium signal transmission from sarco-endoplasmic reticulum to the mitochondria. AB - Recent studies have shown that ryanodine and IP3 receptor (RyR/IP3R)-mediated cytosolic Ca2+ signals propagate to the mitochondria, initiating chains of events vital in the regulation of different cellular functions. However, the fraction of released Ca2+ utilized by the mitochondria during these processes has not been quantified. To measure the amount of Ca2+ taken up by the mitochondria, we used a novel approach that involves simultaneous fluorescence imaging of mitochondrial and cytosolic [Ca2+] in permeabilized H9c2 myotubes and RBL-2H3 mast cells. Communication between sarco-endoplasmic reticulum (SR/ER) and mitochondria is maintained in these permeabilized cells, as evidenced by the large RyR/IP3R driven mitochondrial matrix [Ca2+] and NAD(P)H signals and also by preservation of the morphology of the SR/ER-mitochondrial junctions. Ca2+ was released from the SR/ER by addition of saturating caffeine or IP3 and subsequently thapsigargin (Tg), an inhibitor of SR/ER Ca2+ pumps. The amount of Ca2+ transmitted to the mitochondria was determined by measuring increases of global [Ca2+] in the incubation medium (cytosolic [Ca2+] ([Ca2+]c)). Mitochondrial Ca2+ uptake was calculated from the difference between [Ca2+]c responses recorded in the absence and presence of uncoupler or from [Ca2+]c elevations evoked by uncoupler or ionophore applied after complete Ca2+ mobilization from the SR/ER. [Ca2+]c increases were calibrated by adding Ca2+ pulses to the permeabilized cells. In H9c2 cells, caffeine induced partial mobilization of SR Ca2+ and mitochondria accumulated 26% of the released Ca2+. Sequential application of caffeine and Tg elicited complete discharge of SR Ca2+ without further increase in mitochondrial Ca2+ uptake. In RBL-2H3 mast cells, IP3 by itself elicited complete discharge of the ER Ca2+ store and the increase of the ionophore-releasable mitochondrial Ca2+ content reached 50% of the Ca2+ amount mobilized by IP3 + Tg. Thus, RyR/IP3R direct a substantial fraction of released Ca2+ to the mitochondria. PMID- 11118490 TI - Stoichiometry of human recombinant neuronal nicotinic receptors containing the b3 subunit expressed in Xenopus oocytes. AB - The neuronal nicotinic subunit beta3 forms functional receptors when co-expressed with both an alpha and a beta subunit, such as alpha3 and beta4. We examined the subunit stoichiometry of these 'triplet' alpha3beta4beta3 receptors by expression in Xenopus oocytes of the alpha3, beta4 and beta3 subunits, either in wild-type form or after insertion of a reporter mutation. The mutation chosen was the substitution of a conserved hydrophobic residue in the second transmembrane domain of the subunits (leucine or valine 9THORN ) with a hydrophilic threonine. In other ion channels within the nicotinic superfamily, this mutation type consistently increases the potency of agonists. In muscle-type nicotinic receptors, the magnitude of this effect is approximately constant for each mutant subunit incorporated. In alpha3beta4beta3 receptors, the ACh EC50 was decreased by approximately 17-fold when this mutation was in alpha3 alone and only by fourfold when beta3 alone was mutated. Mutating beta4 was equivalent to mutating alpha3, suggesting that the 'triplet' receptor contains one copy of beta3 and two copies each of alpha3 and beta4. Mutating beta3 and alpha3 or beta3 and beta4 reduced the ACh EC50 further, to values two- to threefold lower than those seen when only alpha3 or beta4 carried the mutation. In 'pair' alpha3beta4 receptors (known to contain two alpha and three beta subunits), mutating beta4 had a greater effect on the ACh EC50 than mutating alpha3, in agreement with an alpha:beta ratio of 2:3 and a constant and independent effect of each copy of the mutation. Our results suggest that alpha3beta4beta3 neuronal nicotinic receptors contain one copy of beta3 and two copies each of alpha3 and beta4 and confirm that in pair alpha3beta4 receptors the alpha/beta subunits are present in a 2:3 ratio. PMID- 11118491 TI - Regulation of the sensitivity of acetylcholine receptors to nicotine in rat habenula neurons. AB - Time-dependent changes in nicotinic acetylcholine receptor (nAChR) function were studied in acutely isolated medial habenula neurons during whole-cell perfusion. The peak amplitude of inward currents induced by 1 s pulses of nicotinic agonists, applied at 30 s intervals, gradually increased over the first several minutes of whole-cell recording. The ratio of response amplitudes at 1 and 15 min (t15/t1) was 1.9. Run-up of responses occurred independently of channel activation and was specific to nAChRs. The channel blocker chlorisondamine (30 microM), co-applied with nicotine, was used to irreversibly block the majority (91 %) of the nAChRs that opened in the first 2 min of recording. Run-up in the remaining 9 % unblocked channels assessed at 15 min (t15/t2 = 3.4) was similar to that in control cells not exposed to nicotine and chlorisondamine simultaneously, implying that run-up is not due to the incorporation of new receptors. A marked alteration in the sensitivity of nAChRs to extracellular Ca2+ was also observed during whole-cell perfusion. The ratio of current amplitudes obtained in 0.2 and 4.0 mM Ca2+ changed from 0.54 (t = 5 min) to 0.82 (t = 30 min). Inward rectification of nicotine-induced responses was reduced during internal dialysis. Voltages for half-maximal conductance were -23.0 and -13.8 mV at 2 and 15 min, respectively. Inclusion of either free Mg2+ ( approximately 2 mM) or spermine (100 microM) in the internal solution counteracted the change in rectification, but did not prevent run-up. The period of run-up was followed by a use-dependent run-down phase. Little run-down in peak current amplitude was induced provided that agonist was applied infrequently (5 min intervals), whereas applications at 30 s intervals produced a loss of channel function after approximately 15 min whole-cell perfusion. The time at which run-down began ( approximately 5-30 min) was correlated with the initial rate of nAChR desensitization ( approximately 200 4000 ms); slowly desensitizing nicotinic currents demonstrated delayed run-down. We suggest that run-up of nAChR-mediated responses does not require receptor activation and may result from a change in channel open probability. We also hypothesize that channel run-down reflects accumulation of nAChRs in long-lived desensitized/inactivated states. PMID- 11118492 TI - Functional differences between cardiac and renal isoforms of the rat Na+-Ca2+ exchanger NCX1 expressed in Xenopus oocytes. AB - The transcript of the Na+-Ca2+ exchanger gene NCX1 undergoes alternative splicing to produce tissue-specific isoforms. The cloned NCX1 isoforms were expressed in Xenopus oocytes and studied using a two-electrode voltage clamp method to measure Na+-Ca2+ exchanger activity. The cardiac isoform (NCX1.1) expressed in oocytes was less sensitive to depolarizing voltages and to activation by [Ca2+]i than the renal isoform (NCX1.3). The cardiac isoform of NCX1 is more sensitive to activation by protein kinase A (PKA) than the renal isoform which may be explained by preferential phosphorylation. The cardiac isoform of NCX1 is phosphorylated to a greater extent than the renal isoform. The action of PKA phosphorylation which increases the activity of the cardiac isoform of the Na+ Ca2+ exchanger in oocytes was confirmed in adult rat ventricular cardiomyocytes by measuring Na+-dependent Ca2+ flux. We conclude that alternative splicing of NCX1 confers distinct functional characteristics to tissue-specific isoforms of the Na+-Ca2+ exchanger. PMID- 11118493 TI - Regulation kinetics of Na+-Ca2+ exchange current in guinea-pig ventricular myocytes. AB - To investigate the regulation of native cardiac Na+-Ca2+ exchange by cytoplasmic Na+ (Na+i) and Ca2+ (Ca2+i), we recorded the Na+-Ca2+ exchange current (INa-Ca) from inside-out 'macro patches' excised from intact guinea-pig ventricular cells. The half-maximal concentration (Kh) of Ca2+i required to induce an inward INa-Ca was 7 uM. The Kh of Na+i required to induce an outward INa-Ca was 21 mM, and tended to decrease at the steady state of Na+-dependent inactivation. The time constant (tau) of Na+-dependent inactivation was ~1.5 s at 100 mM Na+i and 1 uM Ca2+i. The Kh for Na+i was 14 mM. Ca2+i augmented the peak outward INa-Ca (Kh = 0.2 uM) and attenuated Na+-dependent inactivation (Kh = 2.2 uM). The outward INa Ca was activated by 5 uM Ca2+i with a half-time to reach steady state (t1/2) of ~0.4 s. This activation was composed of two exponential processes. Deactivation of the current upon Ca2+i removal also consisted of two exponential processes and had a t1/2 of ~0.5 s. A Na+-Ca2+ exchange model, consisting of one consecutive 4Na+:1Ca2+ exchange cycle and two inactive states, well mimicked the experimental data with regard to ion dependencies and regulation kinetics. These data provide detailed information on the kinetics of the Na+i- and Ca2+i-dependent regulation of native Na+-Ca2+ exchange. They also indicate that the regulation kinetics operate faster in macro patches than in the giant membrane patch from cardiac 'blebs', or in Xenopus oocytes expressing a cloned exchanger (NCX1.1). PMID- 11118494 TI - Calcium dynamics associated with action potentials in single nerve terminals of pyramidal cells in layer 2/3 of the young rat neocortex. AB - Calcium dynamics associated with a single action potential (AP) were studied in single boutons of the axonal arbor of layer 2/3 pyramidal cells in the neocortex of young (P14-16) rats. We used fluorescence imaging with two-photon excitation and Ca2+-selective fluorescence indicators to measure volume-averaged Ca2+ signals. These rapidly reached a peak (in about 1 ms) and then decayed more slowly (tens to hundreds of milliseconds). Single APs and trains of APs reliably evoked Ca2+ transients in en passant boutons located on axon collaterals in cortical layers 2/3, 4 and 5, indicating that APs propagate actively and reliably throughout the axonal arbor. Branch point failures are unlikely to contribute to differences in synaptic efficacy and reliability in the connections made by layer 2/3 pyramidal cells. AP-evoked Ca2+ transients in boutons were mediated by voltage-dependent Ca2+ channels (VDCCs), predominantly by the P/Q- and N subtypes. Ca2+ transients were, on average, of significantly larger amplitude in boutons than in the flanking segments of the axon collateral. Large amplitude Ca2+ transients in boutons were spatially restricted to within <= 3 m of axonal length. Single AP-evoked Ca2+ transients varied up to 10-fold across different boutons even if they were located on the same axon collateral. In contrast, variation of Ca2+ transients evoked by successive APs in a given single bouton was small (coefficient of variation, c.v. <= 0.21). Amplitudes of AP-evoked Ca2+ signals did not correlate with the distance of boutons from the soma. In contrast, AP-evoked Ca2+ signals in spines of basal dendrites decreased slightly (correlation coefficient, r2 = -0.27) with distance from the soma. Measurements with the low-affinity Ca2+ indicator Magnesium Green suggest that the volume averaged residual free [Ca2+]i in a bouton increases on average by 500 nM following a single AP. Higher concentrations of indicator caused, on average, a decrease in the amplitude and an increase in the decay time constant of Ca2+ transients. Assuming a single-compartment model the concentration dependence of decay time constants suggests a low endogenous Ca2+ binding ratio close to 140, indicating that of the total Ca2+ influx ( approximately 2 fC) less than 1% remained free. The indicator concentration dependence of decay time constants further suggests that the residual free Delta[Ca2+]i associated with an AP decays with a time constant of about 60 ms (35 C) reflecting a high Ca2+ extrusion rate of about 2600 s(-1). The results show that AP-evoked volume-averaged Ca2+ transients in single boutons are evoked reliably and, on average, have larger amplitudes than Ca2+ transients in other subcellular compartments of layer 2/3 pyramidal cells. A major functional signature is the large variation in the amplitude of Ca2+ transients between different boutons. This could indicate that local interactions between boutons and different target cells modify the spatiotemporal Ca2+ dynamics in boutons and cause target cell-specific differences in their transmitter release properties. PMID- 11118495 TI - Prolonged depolarization promotes fast gating kinetics of L-type Ca2+ channels in mouse skeletal myotubes. AB - The effects of prolonged conditioning depolarizations on the activation kinetics of skeletal L-type calcium currents (L-currents) were characterized in mouse myotubes using the whole-cell patch clamp technique. The sum of two exponentials was required to adequately fit L-current activation and enabled determination of both the amplitudes (A(fast) and A(slow)) and time constants (tau(fast) and tau(slow)) of each component comprising the macroscopic current. Prepulses sufficient to activate (200 ms) or inactivate (10 s) L-channels did not alter tau(fast), tau(slow), or the fractional contribution of either the fast (A(fast)/(A(fast) + A(slow)) or slow (A(slow)/(A(fast) + A(slow))) amplitudes of subsequently activated L-currents. Prolonged depolarizations (60 s to +40 mV) resulted in the conversion of skeletal L-current to a fast gating mode following brief repriming intervals (3-10 s at -80 mV). Longer repriming intervals (30-60 s at -80 mV) restored L-channels to a predominantly slow gating mode. Accelerated L currents originated from L-type calcium channels since they were completely blocked by a dihydropyridine antagonist (3 microM nifedipine) and exhibited a voltage dependence of activation similar to that observed in the absence of conditioning prepulses. The degree of L-current acceleration produced following prolonged depolarization was voltage dependent. For test potentials between +10 and +50 mV, the fractional contribution of Afast to the total current decreased exponentially with the test voltage (e-fold approximately 38 mV). Thus, L-current acceleration was most significant at more negative test potentials (e.g. +10 mV). Prolonged depolarization also accelerated L-currents recorded from myotubes derived from RyR1-knockout (dyspedic) mice. These results indicate that L-channel acceleration occurs even in the absence of RyR1, and is therefore likely to represent an intrinsic property of skeletal L-channels. The results describe a novel experimental protocol used to demonstrate that slowly activating mammalian skeletal muscle L-channels are capable of undergoing rapid, voltage-dependent transitions during channel activation. The transitions underlying rapid L-channel activation may reflect rapid transitions of the voltage sensor used to trigger the release of calcium from the sarcoplasmic reticulum during excitation contraction coupling. PMID- 11118496 TI - The effect of acidosis on systolic Ca2+ and sarcoplasmic reticulum calcium content in isolated rat ventricular myocytes. AB - We have investigated the mechanisms responsible for the changes of systolic Ca2+ that occur in voltage-clamped rat ventricular myocytes during acidosis produced by application of the weak acid butyrate (30 mM). Intracellular pH regulation was inhibited with dimethylamiloride (bicarbonate-free solution). The application of butyrate produced an intracellular acidification of 0.33 pH units. This was accompanied by a decrease in systolic Ca2+ to about 50% of control. However, within 2 min, systolic Ca2+ returned to control levels. The decrease in systolic Ca2+ was accompanied by a decrease in the Na+-Ca2+ exchange current observed on repolarisation so that the calculated Ca2+ efflux on Na+-Ca2+ exchange was less than the entry on the L-type Ca2+ current. The magnitude of the Na+-Ca2+ exchange current recovered along with systolic Ca2+ until it equalled the Ca2+ entry on the L-type Ca2+ current. From the measurement of Ca2+ fluxes, it was calculated that, during acidosis, the cell gains 121.6+/-16.2 micromol l(-1) of Ca2+. This is equal to the measured increase of sarcoplasmic reticulum (SR) calcium content obtained by applying caffeine (20 mM) and integrating the resulting Na+-Ca2+ exchange current. We conclude that the recovery of the amplitude of the systolic Ca2+ transient is due to decreased SR calcium release, resulting in reduced Ca2+ efflux from the cell leading to increased SR calcium content. PMID- 11118497 TI - Preferential role of intracellular Ca2+ stores in regulation of isometric force in NIH 3T3 fibroblast fibres. AB - Fibroblast contraction plays a major role in wound repair, but the regulatory mechanisms are not well known. We investigated the relations between isometric force and intracellular calcium concentration ([Ca2+]i) in fibroblast fibres. These fibres were made with mouse NIH 3T3 fibroblasts cultured with native collagen in a three-dimensional matrix. Calf serum (CS; 30%) elicited a monotonic increase in force that attained a maximum within 15 min and could be sustained indefinitely. In contrast, [Ca2+]i increased to a peak at 3 min after CS stimulation, then returned to baseline levels by 10 min. Pretreatment with Ca2+ free medium or the Ca2+-channel antagonist nicardipine (10 microM) blocked the CS induced [Ca2+]i increase, but force was not affected. KCl (50 mM) stimulation on the other hand, elicited a prolonged increase in [Ca2+]i but did not increase force. Inhibition of the endoplasmic reticulum Ca2+ release with Ca2+-ATPase inhibitors cyclopiazonic acid (5 microM) or thapsigargin (5 microM) nearly abolished (<20% control) the increase in [Ca2+]i and force response to CS. Treatment with ryanodine (10 microM) and caffeine (20 mM) had a similar effect. The phospholipase C inhibitor U73122 (3 microM) reduced the CS-induced increases in [Ca2+]i and force by 70 and 40%, respectively. We conclude that fibroblast isometric force is not coupled to Ca2+ arising from transmembrane influx but is correlated with the transient [Ca2+]i increase due to release from intracellular stores. Store-released Ca2+ may initiate activation pathways for fibroblast force development, but is not required for force maintenance. PMID- 11118499 TI - Tetraethylammonium potentiates the activity of muscarinic potassium channels in guinea-pig atrial myocytes. AB - The modulation of native muscarinic potassium channels (KACh) by tetraethylammonium (TEA) was studied at 35 degrees C in cell-free patches from acutely dissociated guinea-pig atrial myocytes. The channels were identified unambiguously by their conductance, inward rectification, rapid gating kinetics and pharmacological responses to muscarinic agonists and GTPgammaS. Addition of 5 mM TEA to the cytoplasmic side of the patches almost doubled the open probability of KACh channels that had been activated maximally by GTPgammaS. In contrast even 30 mM TEA did not significantly potentiate the response to carbachol in whole cell recordings. Unlike GTPgammaS, TEA alone did not activate KACh channels de novo, but in patches that showed spontaneous KACh activity, 5 mM TEA increased channel open probability fourfold in the absence of added sodium, ATP or guanine nucleotides. Furthermore, the effect of TEA was not blocked by 10 uM atropine or by 1 mM GDPbetaS, and subsequent addition of 0.1 mM GTPgammaS did not stimulate channel activity further in the presence of TEA. Phosphatidylinositol 4,5 bisphosphate (PIP2) also stimulates KACh channels under these conditions, but the kinetics of gating differ from channels stimulated by either TEA or GTP, which are very similar to one another. The effects of TEA were not mimicked by tetramethyl- or tetrapentylammonium or by sodium or spermine, and TEA did not potentiate the activity of other inwardly rectifying potassium (KATP) channels in patches from cardiac myocytes. We consider the possibility that TEA is mimicking the effect of an unidentified cellular factor, not sodium or PIP2, which normally occupies the TEA site on KACh channel proteins but which diffuses away when the patch is excised. PMID- 11118498 TI - Characterisation of inhibitory and excitatory postsynaptic currents of the rat medial superior olive. AB - The medial superior olive (MSO) is part of the binaural auditory pathway, receiving excitatory projections from both cochlear nuclei and an inhibitory input from the ipsilateral medial nucleus of the trapezoid body (MNTB). We characterised the excitatory and inhibitory synaptic currents of MSO neurones in 3- to 14-day-old rats using whole-cell patch-clamp methods in a brain slice preparation.A dual component EPSC was mediated by AMPA and NMDA receptors. The AMPA receptor-mediated EPSC decayed with a time constant of 1.99+/-0.16 ms (n = 8). Following blockade of glutamate receptors, a monosynaptic strychnine sensitive response was evoked on stimulation of the MNTB, indicative of a glycine receptor-mediated IPSC. GABAA receptors contributed to IPSCs in rats under 6 days old (bicuculline blocked 30% of the IPSC). In older rats little or no bicuculline sensitive component was detectable, except in the presence of flunitrazepam. These glycinergic IPSCs showed a reversal potential that varied with changes in [Cl-]i, as predicted by the Nernst equation. The IPSC exhibited two developmentally relevant changes. (i) At around postnatal day 6, the GABAA receptor-mediated component declined, leaving a predominant glycine-mediated IPSC. The isolated glycinergic IPSC decayed with time constants of 7.8+/-0.3 and 38.3+/-1.7 ms, with the slower component contributing 7.8+/-0.6% of the peak amplitude (n = 121, 3-11 days old, -70 mV, 25 deg C). (ii) After day 11 the IPSC fast decay accelerated to 3.9+/-0.3 ms (n = 12) and the magnitude of the slow component declined to less than 1%. Spontaneous miniature glycinergic IPSCs (mIPSCs) were variable in amplitude and were of large conductance (1.83+/-0.19 nS, n = 8). The amplitude was unchanged on lowering [Ca2+]o. The time course of evoked and spontaneous miniature glycinergic IPSCs were compared. The 10-90% rise times were 0.7 and 0.6 ms, respectively. The evoked IPSC decayed with a fast time constant of 7.2+/-0.7 ms, while the mIPSC decayed with a fast time constant of 5.3+/-0.4 ms in the same seven cells.The glycinergic IPSC decay was voltage dependent with an e-fold change over 118 mV. The temperature dependence of the IPSC decay indicated a Q10 value of 2. Picrotoxin and cyanotriphenylborate had little or no effect on IPSCs from 6- to 14-day-old animals, implying homomeric channels are rare. We conclude that the MSO receives excitatory inputs mediated by AMPA and NMDA receptors and a strong glycinergic IPSC which has a significant contribution from GABAA receptors in neonatal rats. Functionally, the IPSC could increase membrane conductance during the decay of binaural glutamatergic EPSCs, thus refining coincidence detection and interaural timing differences. PMID- 11118500 TI - Susceptibility of ATP-sensitive K+ channels to cell stress through mediation of phosphoinositides as examined by photoirradiation. AB - Cell stress is implicated in a number of pathological states of metabolism, such as ischaemia, reperfusion and apoptosis in heart, neurons and other tissues. While it is known that the ATP-sensitive K+ (KATP) channel plays a role during metabolic abnormality, little information is available about the direct response of this channel to cell stress. Using photoirradiation stimulation, we studied the effects of cell stress on both native and cloned KATP channels. Single KATP channel currents were recorded from cell-attached and inside-out patches of rat ventricular myocytes and COS-1 cells coexpressing SUR2 and Kir6.2. KATP channel activity increased within < 1 min upon irradiation. The activity resulted from increased maximal open probability and decreased ATP inhibition. The effects remained after the irradiation was stopped. Irradiation also affected the channels formed only by Kir6.2DeltaC35. The irradiation-induced activation was comparable to that induced by phosphoinositides. Analysis of phosphatidylinositol composition revealed an elevated phosphatidylinositol bisphosphate level with irradiation. Wortmannin, an inhibitor of phosphatidylinositol kinases, decreased both the irradiation-induced channel activity and the production of phosphatidylinositol bisphosphates. Radical scavengers also reduced the irradiation-induced activation, suggesting a role for free radicals, an immediate product of photoirradiation. We conclude that photoirradiation can modify the single-channel properties of KATP, which appears to be mediated by phosphoinositides. Our study suggests that cellular stress may be linked with KATP channels, and we offer a putative mechanism for such a linkage. PMID- 11118501 TI - Zinc-induced changes in ionic currents of clonal rat pancreatic -cells: activation of ATP-sensitive K+ channels. AB - The effects of zinc (Zn2+) on excitability and ionic conductances were analysed on RINm5F insulinoma cells under whole-cell and outside-out patch-clamp recording conditions. We found that extracellular application of 10-20 microM Zn2+ induced a reversible abolition of Ca2+ action potential firing, which was accompanied by an hyperpolarisation of the resting membrane potential. Higher concentrations of Zn2+, in the tens to hundreds micromolar range, induced a reversible reduction of voltage-gated Ca2+ and, to a lesser extent, K+ currents. Low-voltage-activated Ca2+ currents were more sensitive to Zn2+ block than high voltage-activated Ca2+ currents. The Zn2+-induced hyperpolarisation arose from a dose-dependent increase in a voltage-independent K+ conductance that was pharmacologically identified as an ATP-sensitive K+ (KATP) conductance. The effect was rapid in onset, readily reversible, voltage independent, and related to intracellular ATP concentration. In the presence of 1 mM intracellular ATP, half-maximal activation of KATP channels was obtained with extracellular application of 1.7 microM Zn2+. Single channel analysis revealed that extracellular Zn2+ increased the KATP channel open state probability with no change in the single channel conductance. Our data support the hypothesis that Zn2+ binding to KATP protein subunits results in an activation of the channels, therefore regulating the resting membrane potential and decreasing the excitability of RINm5F cells. Taken together, our results suggest that Zn2+ can influence insulin secretion in pancreatic beta-cells through a negative feedback loop, involving both KATP and voltage-gated conductances. PMID- 11118502 TI - Novel activation of non-selective cationic channels by dinitrosyl iron thiosulfate in PC12 cells. AB - Low molecular mass dinitrosyl iron complexes (DNICs) are nitrosating agents and it is known that the dinitrosyl iron moiety can be transferred to proteins. The aim of the present study was to determine if the formation of protein-bound dinitrosyl iron can modulate ionic channel activity. In PC12 cells, dinitrosyl iron-thiosulfate (50 microM) caused irreversible activation of a depolarizing inward current (IDNIC). IDNIC was partially inhibited by the metal chelator diethyldithiocarbamate (DETC, 1 mM), but not by the reducing/denitrosylating agent dithiothreitol (DTT, 5 mM). The activation of IDNIC was not reproduced by application of nitric oxide (NO., 100 microM), S-nitrocysteine (200 microM) or ferrous iron-thiosulfate (50 microM), and was not prevented by the irreversible guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo-[4, 3-a]quinoxalin-1-one (ODQ, 1 microM). Similarly, intracellular perfusion of dinitrosyl iron-thiosulfate (100 microM) did not result in activation of IDNIC. Ion replacement experiments show that the DETC-sensitive component of IDNIC is a non-selective cationic current. In accordance, IDNIC was blocked by antagonists of receptor-operated calcium entry, gadolinium (25 microM) and SK&F 96365 (25 microM). Single-channel measurements from outside-out patches reveal that the DETC-sensitive component of IDNIC is an inward current carried by a cationic channel having a conductance of 50 pS. The present observations suggest that the formation of ion channel-bound dinitrosyl iron represents another mechanism of regulation of ion channel activity by NO.-related species, which may be particularly important in pathophysiological processes where NO. is overproduced. PMID- 11118503 TI - Coding and adaptation during mechanical stimulation in the leech nervous system. AB - The experiments described here were designed to characterise sensory coding and adaptation during mechanical stimulation in the leech (Hirudo medicinalis). A chain of three ganglia and a segment of the body wall connected to the central ganglion were used. Eight extracellular suction pipettes and one or two intracellular electrodes were used to record action potentials from all mechanosensory neurones of the three ganglia. When the skin of the body wall was briefly touched with a filament exerting a force of about 2 mN, touch (T) cells in the central ganglion, but also those in adjacent ganglia (i.e. anterior and posterior), fired one or two action potentials. However, the threshold for action potential initiation was lower for T cells in the central ganglion than for those in adjacent ganglia. The timing of the first evoked action potential in a T cell was very reproducible with a jitter often lower than 100 us. Action potentials in T cells were not significantly correlated. When the force exerted by the filament was increased above 20 mN, pressure (P) cells in the central and neighbouring ganglia fired action potentials. Action potentials in P cells usually followed those evoked in T cells with a delay of about 20 ms and had a larger jitter of 0.5-10 ms. With stronger stimulations exceeding 50 mN, noxious (N) cells also fired action potentials. With such stimulations the majority of mechanosensory neurones in the three ganglia fired action potentials. The spatial properties of the whole receptive field of the mechanosensory neurones were explored by touching different parts of the skin. When the mechanical stimulation was applied for a longer time, i.e. 1 s, only P cells in the central ganglion continued to fire action potentials. P cells in neighbouring ganglia fully adapted after firing two or three action potentials.P cells in adjacent ganglia, having fully adapted to a steady mechanical stimulation of one part of the skin, fired action potentials following stimulation of a different region of the skin. These results indicate that a brief and localised stimulation of the skin can activate more than a dozen different mechanosensory neurones in the three ganglia and after 100 ms of steady stimulation many of these mechanosensory neurones stop firing action potentials and fully adapt. Adaptation occurs primarily at the nerve endings and mechanosensory neurones can quickly respond to mechanical stimulation at a different location on the skin. PMID- 11118504 TI - Re-evaluation of phorbol ester-induced potentiation of transmitter release from mossy fibre terminals of the mouse hippocampus. AB - To investigate the mechanisms by which phorbol esters potentiate transmitter release from mossy fibre terminals we used fura dextran to measure the intraterminal Ca2+ concentration in mouse hippocampal slices. A phorbol ester, phorbol 12,13-diacetate (PDAc), potentiated the field excitatory postsynaptic potential (fEPSP) slope. PDAc also enhanced the stimulation-dependent increase of [Ca2+]i in the mossy fibre terminal (Delta[Ca2+]pre). The magnitude of the PDAc induced fEPSP potentiation (463+/-57% at 10 microM) was larger than that expected from the enhancement of Delta[Ca2+]pre (153+/-5%). The Delta[Ca2+]pre was suppressed by omega-agatoxin IVA (omega-AgTxIVA, 200 nM), a P/Q-type Ca2+ channel specific blocker, by 31%. The effect of PDAc did not select between omega-AgTxIVA sensitive and -resistant components. The PDAc-induced potentiation of the fEPSP slope was partially antagonized by the protein kinase C (PKC) inhibitor bisindolylmaleimide I (BIS-I, 10 microM), whereas the Delta[Ca2+]pre was completely blocked by BIS-I. Although the BIS-I-sensitive fEPSP potentiation was accompanied by a reduction of the paired-pulse ratio (PPR), the BIS-I-resistant component was not. Whole-cell patch clamp recording from a CA3 pyramidal neuron in a BIS-I-treated slice demonstrated that PDAc (10 microM) increased the frequency of miniature excitatory postsynaptic currents (mEPSCs, 259+/-33% of control) without a noticeable change in their amplitude (102+/-5% of control). These results suggest that PKC potentiates transmitter release by at least two distinct mechanisms, one Delta[Ca2+]pre dependent and the other Delta[Ca2+]pre independent. In addition, some phorbol ester-mediated potentiation of synaptic transmission appears to occur without activating PKC. PMID- 11118505 TI - Regulation of adenosine transport by D-glucose in human fetal endothelial cells: involvement of nitric oxide, protein kinase C and mitogen-activated protein kinase. AB - The effects of elevated D-glucose on adenosine transport were investigated in human cultured umbilical vein endothelial cells isolated from normal pregnancies. Elevated D-glucose resulted in a time- (8-12 h) and concentration-dependent (half maximal at 10+/-2 mM) inhibition of adenosine transport, which was associated with a reduction in the Vmax for nitrobenzylthioinosine (NBMPR)-sensitive (es) saturable nucleoside with no significant change in Km. d-Fructose (25 mM), 2 deoxy-D-glucose (25 mM) or D-mannitol (20 mM) had no effect on adenosine transport. Adenosine transport was inhibited following incubation of cells with the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA; 100 nM, 30 min to 24 h). D-Glucose-induced inhibition of transport was abolished by calphostin C (100 nM, an inhibitor of PKC), and was not further reduced by PMA. Increased PKC activity in the membrane (particulate) fraction of endothelial cells exposed to D-glucose or PMA was blocked by calphostin C but was unaffected by NG-nitro-L-arginine methyl ester (L-NAME; 100 microM, an inhibitor of nitric oxide synthase (NOS)) or PD-98059 (10 microM, an inhibitor of mitogen-activated protein kinase kinase 1). D-Glucose and PMA increased endothelial NOS (eNOS) activity, which was prevented by calphostin C or omission of extracellular Ca2+ and unaffected by PD-98059. Adenosine transport was inhibited by S-nitroso-N acetyl-l, d-penicillamine (SNAP; 100 microM, an NO donor) but was increased in cells incubated with L-NAME. The effect of SNAP on adenosine transport was abolished by PD-98059. Phosphorylation of mitogen-activated protein kinases p44mapk (ERK1) and p42mapk (ERK2) was increased in endothelial cells exposed to elevated D-glucose (25 mM for 30 min to 24 h) and the NO donor SNAP (100 microM, 30 min). The effect of D-glucose was blocked by PD-98059 or L-NAME, which also prevented the inhibition of adenosine transport mediated by elevated D-glucose. Our findings provide evidence that D-glucose inhibits adenosine transport in human fetal endothelial cells by a mechanism that involves activation of PKC, leading to increased NO levels and p42-p44mapk phosphorylation. Thus, the biological actions of adenosine appear to be altered under conditions of sustained hyperglycaemia. PMID- 11118506 TI - Displacement of the contents of dentinal tubules and sensory transduction in intradental nerves of the cat. AB - Experiments were performed on anaesthetized cats to test the hypothesis that fluid flow through dentinal tubules is part of the mechanism involved in the transduction of pain-producing stimuli in teeth. In 11 animals, fluid flow through dentine and single- and multi-unit activity in intradental nerves were recorded simultaneously during the application of changes in hydrostatic pressure (-500 to +500 mm Hg) to exposed dentine. Seventeen A-fibres (conduction velocity (CV), 10.6-55.1 m s(-1)) were isolated that were pressure sensitive. The thresholds of these units in terms of dentinal fluid flow were in the range 0.3 2.1 nl s(-1) mm(-2) during outward flow from the pulp and 2.0-3.5 nl s(-1) mm(-2) during inward flow. All the units were more sensitive to outward than inward flow. Twenty-eight units (CV, 0.6-48.8 m s-1) were not pressure sensitive, and 12 of these had conduction velocities in the C-fibre range (< 2.5 m s(-1)). The velocities of the tubular contents were calculated by estimating the number and diameters of dentinal tubules exposed. At the threshold of single-fibre responses these velocities were in the range 31.7-222.9 microm s(-1) during outward flow 211.4-369.6 microm s-1 during inward flow. Repetitive pressure stimulation of dentine resulted in a progressive reduction in the evoked discharge, which was probably due to pulp damage. In seven animals, 10 single intradental nerve fibres were selected that responded to hydrostatic pressure stimuli and their responses to the application of hot, cold, osmotic, mechanical and drying stimuli to exposed dentine were investigated. With these stimuli dentinal fluid flow could not be recorded in vivo for technical reasons and was therefore recorded in vitro after completion of the electrophysiological recordings. With each form of stimulus, the discharge evoked in vivo was closely related to the flow predicted from the in vitro measurements. The results were therefore consistent with the hypothesis that the stimuli act through a common transduction mechanism that involves fluid flow through dentine. PMID- 11118507 TI - Electrically evoked neuropeptide release and neurogenic inflammation differ between rat and human skin. AB - Protein extravasation and vasodilatation can be induced by neuropeptides released from nociceptive afferents (neurogenic inflammation). We measured electrically evoked neuropeptide release and concomitant protein extravasation in human and rat skin using intradermal microdialysis. Plasmapheresis capillaries were inserted intradermally at a length of 1.5 cm in the volar forearm of human subjects or abdominal skin of rats. Capillaries were perfused with Ringer solution at a flow rate of 2.5 or 1.6 microl min(-1). After a baseline period of 60 min capillaries were stimulated electrically (1 Hz, 80 mA, 0.5 ms or 4 Hz, 30 mA, 0.5 ms) for 30 min using a surface electrode directly above the capillaries and a stainless-steel wire inserted in the capillaries. Total protein concentration was assessed photometrically and calcitonin gene-related peptide (CGRP) and substance P (SP) concentrations were measured by enzyme-linked immunosorbent assay (ELISA). In rat skin, electrical stimulation increased CGRP and total protein concentration in the dialysate. SP measurements showed a larger variance but only for the 1 Hz stimulation was the increased release significant. In human skin, electrical stimulation provoked a large flare reaction and at a frequency of 4 Hz both CGRP and SP concentrations increased significantly. In spite of the large flare reactions no protein extravasation was induced, which suggests major species differences. It will be of interest to investigate whether the lack of neurogenic protein extravasation is also valid under pathophysiological conditions. PMID- 11118508 TI - Contractile properties and proteins of smooth muscles of a calponin knockout mouse. AB - The role of h1-calponin in regulating the contractile properties of smooth muscle was investigated in bladder and vas deferens of mice carrying a targeted mutation in both alleles designed to inactivate the basic calponin gene. These calponin knockout (KO) mice displayed no detectable h1-calponin in their smooth muscles. The amplitudes of Ca2+ sensitization, force and Ca2+ sensitivity were not significantly different in permeabilized smooth muscle of KO compared with wild type (WT) mice, nor were the delays in onset and half-times of Ca2+ sensitization, initiated by flash photolysis of caged GTPgammaS, different. The unloaded shortening velocity (Vus) of thiophosphorylated fibres was significantly (P<0.05) faster in the smooth muscle of KO than WT animals, but could be slowed by exogenous calponin to approximate WT levels; the concentration dependence of exogenous calponin slowing of Vus was proportional to its actomyosin binding in situ. Actin expression was reduced by 25-50%, relative to that of myosin heavy chain, in smooth muscle of KO mice, without any change in the relative distribution of the actin isoforms. We conclude that the faster Vus of smooth muscle of the KO mouse is consistent with, but does not prove without further study, physiological regulation of the crossbridge cycle by calponin. Our results show no detectable role of calponin in the signal transduction of the Ca2+ sensitization pathways in smooth muscle. PMID- 11118509 TI - Distinctive patterns of static and dynamic gamma motor activity during locomotion in the decerebrate cat. AB - Simultaneous recordings were made from gamma (gamma) motor axons and from muscle spindle afferents of the medial gastrocnemius (MG) muscle during locomotion in decerebrate cats. The gamma-neurons were identified as static or dynamic (gammas or gammad) by correlating their behaviour during midbrain stimulation with changes in muscle spindle afferent responses to muscle stretch. On the basis of their behaviour during locomotion, gammas neurons could be divided into two groups. One group (type-1) showed strongly and smoothly modulated discharge increasing in parallel with the active muscle shortening in ankle extension, but with phase advance. The other group (type-2) also showed a modulated pattern, but with increased firing centred on the flexion phase. The proportions of the two were 13 type-1 and 7 type-2. The type-1 firing pattern accurately predicted the difference in firing frequency for secondary afferents obtained by subtracting from the recordings made during active movements the response of the same units to the movements repeated passively in the absence of fusimotor activity. The type-2 pattern also became consistent with the difference signal, when operated on by a phase lag appropriate to the effects of bag2 intrafusal fibres. These results suggest that there may be some degree of separate control of chain and bag2 intrafusal fibres. The discharge of gammad axons was also found to fluctuate with the locomotor cycle, with a pattern very distinct from that of the gammas records. The gammad firing frequency rose very suddenly from zero to a maximum at the onset of muscle shortening and continued into the beginning of lengthening. The term 'interrupted' discharge is suggested as a useful description. The timing of this discharge was shown to be appropriate for sensitising the primary afferents to detect the onset of stretch. PMID- 11118510 TI - Caffeine ingestion does not alter carbohydrate or fat metabolism in human skeletal muscle during exercise. AB - This study examined the effect of ingesting caffeine (6 mg kg-1) on muscle carbohydrate and fat metabolism during steady-state exercise in humans. Young male subjects (n = 10) performed 1 h of exercise (70% maximal oxygen consumption (VO2,max)) on two occasions (after ingestion of placebo and caffeine) and leg metabolism was quantified by the combination of direct Fick measures and muscle biopsies. Following caffeine ingestion serum fatty acid and glycerol concentration increased (P< or =0.05) at rest, suggesting enhanced adipose tissue lipolysis. In addition circulating adrenaline concentration was increased (P< or =0.05) at rest following caffeine ingestion and this, as well as leg noradrenaline spillover, was elevated (P< or =0.05) above placebo values during exercise. Caffeine resulted in a modest increase (P< or =0.05) in leg vascular resistance, but no difference was found in leg blood flow. Arterial lactate and glucose concentrations were increased (P< or =0.05) by caffeine, while the rise in plasma potassium was dampened (P< or =0.05). There were no differences in respiratory exchange ratio or in leg glucose uptake, net muscle glycogenolysis, leg lactate release or muscle lactate, or glucose 6-phosphate concentration. Similarly there were no differences between treatments in leg fatty acid uptake, glycerol release or muscle acetyl CoA concentration. These findings indicate that caffeine ingestion stimulated the sympathetic nervous system but did not alter the carbohydrate or fat metabolism in the monitored leg. Other tissues must have been involved in the changes in circulating potassium, fatty acids, glucose and lactate. PMID- 11118511 TI - Interstitial and arterial-venous [K+] in human calf muscle during dynamic exercise: effect of ischaemia and relation to muscle pain. AB - Changes in the concentration of interstitial K+ surrounding skeletal muscle fibres ([K+]I) probably play some role in the regulation of cardiovascular adjustments to muscular activity, as well as in the aetiology of muscle pain and fatigue during high-intensity exercise. However, there is very little information on the response of [K+]I to exercise in human skeletal muscle. Five young healthy subjects performed plantar flexion exercise for four 5 min periods at increasing power outputs ( approximately 1-6 W) with 10 min intervening recovery periods, as well as for two 5 min periods with ischaemia at approximately 1 and approximately 3 W. Microdialysis probes were inserted into the gastrocnemius medialis muscle of the right leg to measure [K+]I, and K+ release from the plantar flexors during and after incremental exercise was calculated from plasma flow and arterial venous differences for K+. Calf muscle pain was assessed using a visual analogue scale. On average, [K+]I was 4.4 mmol l(-1) at rest and increased during minutes 3-5 of incremental exercise by approximately 1-7 mmol l(-1) as a positive function of power output. K+ release also increased as a function of exercise intensity, although there was a progressive increase by approximately 1-6 mmol l 1 in the [K+] gradient between the interstitium and arterial-venous plasma. [K+]I was lower during ischaemic exercise than control exercise. In contrast to this effect of ischaemia on [K+]I, muscle pain was relatively higher during ischaemic exercise, which demonstrates that factors other than changes in [K+]I are responsible for ischaemic muscle pain. In conclusion, this study has demonstrated that during 5 min of dynamic exercise, [K+]I increases during the later period of exercise as a positive function of exercise intensity, ischaemia reduces [K+]I during rest and exercise, and the increase in [K+]I is not responsible for muscle pain during ischaemic exercise. PMID- 11118512 TI - Contribution of skeletal muscle 'ergoreceptors' in the human leg to respiratory control in chronic heart failure. AB - The role of skeletal muscle ergoreceptors (afferents sensitive to muscle contraction, differentiated into metaboreceptors, sensitive to metabolic changes, and mechanoreceptors, sensitive to mechanical changes) in the genesis of the increased ventilatory drive in chronic heart failure is controversial. We have aimed to clarify the contribution of muscle metaboreceptors in the leg to ventilation and to compare this with the contribution of mechanoreceptors. Eighteen heart failure patients and 12 controls were studied. Metaboreceptor and mechanoreceptor responses were measured in the leg by bicycle exercise with and without regional circulatory occlusion during recovery, and by active and equivalent passive limb movement, respectively.Patients, in comparison with controls, had a lower peak VO2 (Oxygen uptake) (18.1+/-1.6 vs. 24.5+/-2.5 ml min( 1) kg(-1), P< 0.05), and an evident metaboreceptor contribution to the ventilatory response (3.5+/-1.6 vs. -4.0+/-1.3 l min(-1), P<0.001). Passive limb movement increased ventilation in both patients and controls (+3.7+/-0.4 and +2.9+/-0.5 l min(-1) from baseline, P<0.003), but this was associated with an increase in VO2 (+0.1+/-0.01 and +0.1+/-0.02 l min(-1) from baseline, P<0.001). The ratio of the increase in ventilation to the increase in VO2 during passive movement was not significantly higher than that during active exercise for either patients or controls, suggesting a limited contribution from the mechanoreceptors. In chronic heart failure the presence of a muscle metaboreceptor reflex is also demonstrated in the leg, while mechanoreceptors exhibited a non-significant contribution in both patients and controls. The hypothesis of a peripheral origin of symptoms of exertional intolerance in this syndrome is confirmed as being mainly due to metabolic stimulation of the muscle metaboreceptors. PMID- 11118513 TI - A model of calcium channels. AB - We propose a model of calcium channels that can explain most of their observed properties, including the anomalous mole fraction effect and mutation of the glutamate residues. The structure grossly resembles that of the KcsA potassium channel except for the presence of an extracellular vestibule and a shorter selectivity filter containing four glutamate residues. Using this model in electrostatic calculations and Brownian dynamics simulations, we study mechanisms of ion permeation and selectivity in the channel. Potential energy profiles calculated for multiple ions in the channel provide explanations of ion permeation, the block of Na(+) currents by Ca(2+) ions, and many other observed properties. Brownian dynamics simulations provide quantitative predictions for the channel currents which reproduce available experimental data. PMID- 11118514 TI - Structure and function of ATA3, a new subtype of amino acid transport system A, primarily expressed in the liver and skeletal muscle. AB - To date, two different transporters that are capable of transporting alpha (methylamino)isobutyric acid, the specific substrate for amino acid transport system A, have been cloned. These two transporters are known as ATA1 and ATA2. We have cloned a third transporter that is able to transport the system A-specific substrate. This new transporter, cloned from rat skeletal muscle and designated rATA3, consists of 547 amino acids and has a high degree of homology to rat ATA1 (47% identity) and rat ATA2 (57% identity). rATA3 mRNA is present only in the liver and skeletal muscle. When expressed in Xenopus laevis oocytes, rATA3 mediates the transport of alpha-[(14)C](methylamino)isobutyric acid and [(3)H]alanine. With the two-microelectrode voltage clamp technique, we have shown that exposure of rATA3-expressing oocytes to neutral, short-chain aliphatic amino acids induces inward currents. The amino acid-induced current is Na(+)-dependent and pH-dependent. Analysis of the currents with alanine as the substrate has shown that the K(0. 5) for alanine (i.e., concentration of the amino acid yielding half-maximal current) is 4.2+/-0.1 mM and that the Na(+):alanine stoichiometry is 1:1. PMID- 11118515 TI - Advances in the characterization of supported lipid films with the atomic force microscope. AB - During the past decade, the atomic force microscope (AFM) has become a key technique in biochemistry and biophysics to characterize supported lipid films, as testified by the continuous growth in the number of papers published in the field. The unique capabilities of AFM are: (i) capacity to probe, in real time and in aqueous environment, the surface structure of lipid films; (ii) ability to directly measure physical properties at high spatial resolution; (iii) possibility to modify the film structure and biophysical processes in a controlled way. Such experiments, published up to June 2000, are the focus of the present review. First, we provide a general introduction on the preparation and characterization of supported lipid films as well as on the principles of AFM. The section 'Structural properties' focuses on the various applications of AFM for characterizing the structure of supported lipid films: visualization of molecular structure, formation of structural defects, effect of external agents, formation of supported films, organization of phase-separated films (coexistence region, mixed films) and, finally, the use of supported lipid bilayers for anchoring biomolecules such as DNA, enzymes and crystalline protein arrays. The section 'Physical properties' introduces the principles of force measurements by AFM, interpretation of these measurements and their recent application to supported lipid films and related structures. Finally, we highlight the major achievements brought by the technique and some of the current limitations. PMID- 11118516 TI - Characterisation of thapsigargin-releasable Ca(2+) from the Ca(2+)-ATPase of sarcoplasmic reticulum at limiting [Ca(2+)]. AB - The Ca(2+) binding sites of the Ca(2+)-ATPase of skeletal muscle sarcoplasmic reticulum (SR) have been identified as two high-affinity sites orientated towards the cytoplasm, two sites of low affinity facing the lumen, and a transient occluded species that is isolated from both membrane surfaces. Binding and release studies, using (45)Ca(2+), have invoked models with sequential binding and release from high- and low-affinity sites in a channel-like structure. We have characterised turnover conditions in isolated SR vesicles with oxalate in a Ca(2+)-limited state, [Ca(2)](lim), where both high- and low-affinity sites are vacant in the absence of chelators (Biochim. Biophys. Acta 1418 (1999) 48-60). Thapsigargin (TG), a high-affinity specific inhibitor of the Ca(2+)-ATPase, released a fraction of total Ca(2+) at [Ca(2+)](lim) that accumulated during active transport. Maximal Ca(2+) release was at 2:1 TG/ATPase. Ionophore, A23187, and Triton X-100 released the rest of Ca(2+) resistant to TG. The amount of Ca(2+) released depended on the incubation time at [Ca(2+)](lim), being 3.0 nmol/mg at 20 s and 0.42 nmol/mg at 1000 s. Rate constants for release declined from 0. 13 to 0.03 s(-1). The rapidly released early fraction declined with time and k=0.13 min(-1). Release was not due to reversal of the pump cycle since ADP had no effect; neither was release impaired with substrates acetyl phosphate or GTP. A phase of reuptake of Ca(2+) followed release, being greater with shorter delay (up to 200 s) following active transport. Reuptake was minimal with GTP, with delays more than 300 s, and was abolished by vanadate and at higher [TG], >5 microM. Ruthenium red had no effect on efflux, indicating that ryanodine sensitive efflux channels in terminal cisternal membranes are not involved in the Ca(2+) release mechanism. It is concluded that the Ca(2+) released by TG is from the occluded Ca(2+) fraction. The Ca(2+) occlusion sites appear to be independent of both high-affinity cytoplasmic and low-affinity lumenal sites, supporting a multisite 'in line' sequential binding mechanism for Ca(2+) transport. PMID- 11118517 TI - Selective NMR observation of inhibitor and sugar binding to the galactose-H(+) symport protein GalP, of Escherichia coli. AB - The binding of the transport inhibitor forskolin, synthetically labelled with (13)C, to the galactose-H(+) symport protein GalP, overexpressed in its native inner membranes from Escherichia coli, was studied using cross-polarization magic angle spinning (13)C NMR. (13)C-Labelled D-galactose and D-glucose were displaced from GalP with the singly labelled [7-OCO(13)CH(3)]forskolin and were not bound to any alternative site within the protein, demonstrating that any multiple sugar binding sites are not simultaneously accessible to these sugars and the inhibitor within GalP. The observation of singly (13)C-labelled forskolin was hampered by interference from natural abundance (13)C in the membranes and so the effectiveness of double-quantum filtration was assessed for the exclusive detection of (13)C spin pairs in sugar (D-[1,2-(13)C(2)]glucose) and inhibitor ([7-O(13)CO(13)CH(3)]forskolin) bound to the GalP protein. The solid state NMR methodology was not effective in creating double-quantum selection of ligand bound with membranes in the 'fluid' state (approx. 2 degrees C) but could be applied in a straightforward way to systems that were kept frozen. At -35 degrees C, double-quantum filtration detected unbound sugar that was incorporated into ice structure within the sample, and was not distinguished from protein-bound sugar. However, the method detected doubly labelled forskolin that is selectively bound only to the transport system under these conditions and provided very effective suppression of interference from natural abundance (13)C background. These results indicate that solid state NMR methods can be used to resolve selectively the interactions of more hydrophobic ligands in the binding sites of target proteins. PMID- 11118518 TI - Effects of the antibiotic peptide microcin J25 on liposomes: role of acyl chain length and negatively charged phospholipid. AB - This paper reports the effects of microcin J25 (MccJ25) on the microviscosity and permeability of phospholipid vesicles of different compositions. The results obtained indicate that MccJ25 interacts with egg L-alpha-phosphatidylcholine (PC) vesicles as demonstrated by peptide intrinsic fluorescence determinations. The interaction depends on the lipid composition of the vesicles. MccJ25 interaction induces a significant fluidity increase of egg PC vesicles. This effect is time and concentration dependent. Both trimethyl ammonium 1,6-diphenyl-1,3,5 hexatriene and 1,6-diphenyl-1, 3,5-hexatriene gave the same results. The microviscosity of L-alpha-phosphatidylcholine dipalmitoyl small unilamellar vesicles (SUVs) was affected while that of L-alpha-phosphatidylcholine dimyristoyl vesicles was not, indicating that the effect was strongly dependent on the chain length of fatty acids. On the other hand, negatively charged L-alpha phosphatidyl-DL-glycerol (PG) vesicles remarkably inhibited the peptide effect. Nevertheless vesicles composed of L-alpha-phosphatidylethanolamine:PG:cardiolipin (7:2:1), a composition resembling bacterial membrane, were sensitive to the MccJ25 effect. MccJ25 effectively dissipated the valinomycin-induced membrane potential, but induced only a modest leakage (5%) of the trapped Tb(+3) dipicolinic acid complex. These results indicate that the peptides interact and perturb the bilayer of SUVs. The relationships between this effect and bactericidal action remain to be elucidated. PMID- 11118519 TI - pH and external Ca(2+) regulation of a small conductance Cl(-) channel in kidney distal tubule. AB - A single channel characterization of the Cl(-) channels in distal nephron was undertaken using vesicles prepared from plasma membranes of isolated rabbit distal tubules. The presence in this vesicle preparation of ClC-K type Cl(-) channels was first established by immunodetection using an antibody raised against ClC-K isoforms. A ClC-K1 based functional characterization was next performed by investigating the pH and external Ca(2+) regulation of a small conductance Cl(-) channel which we identified previously by channel incorporation experiments. Acidification of the cis (external) solution from pH 7.4 to 6.5 led to a dose-dependent inhibition of the channel open probability P(O). Similarly, changing the trans pH from 7.4 to 6.8 resulted in a 4-fold decrease of the channel P(O) with no effect on the channel conductance. Channel activity also appeared to be regulated by cis (external) Ca(2+) concentration, with a dose dependent increase in channel activity as a function of the cis Ca(2+) concentration. It is concluded on the basis of these results that the small conductance Cl(-) channel present in rabbit distal tubules is functionally equivalent to the ClC-K1 channel in the rat. In addition, the present work constitutes the first single channel evidence for a chloride channel regulated by external Ca(2+). PMID- 11118520 TI - Involvement of the TOM complex in external NADH transport into yeast mitochondria depleted of mitochondrial porin1. AB - The protein(s) responsible for metabolite transport through the outer membrane of the yeast Saccharomyces cerevisiae mitochondria depleted of mitochondrial porin (also known as voltage-dependent anion selective channel), termed here porin1, is (are) still unidentified. It is postulated that the transport may be supported by the protein import machinery of the outer membrane, the TOM complex (translocase of the outer membrane). We demonstrate here that in the absence of functional porin1, the blockage of the TOM complex by the fusion protein termed pb(2)-DHFR (consisting of the first 167 amino acids of yeast cytochrome b(2) preprotein connected to mouse dihydrofolate reductase) limits the access of external NADH to mitochondria. It was measured by the ability of the blockage to inhibit external NADH oxidation by the proper dehydrogenase located at the outer surface of the inner membrane. The inhibition depends on external NADH concentration and increases with decreasing amounts of the substrate. In the presence of 1 microg of pb(2)-DHFR per 50 microg of mitochondrial protein almost quantitative inhibition was observed when external NADH was applied at the concentration of 70 nmol per mg of mitochondrial protein. On the other hand, external NADH decreases the levels of pb(2)-DHFR binding at the trans site of the TOM complex in porin1 depleted mitochondria in a concentration-dependent fashion. Our data define an important role of the TOM complex in the transport of external NADH across the outer membrane of porin1-depleted mitochondria. PMID- 11118521 TI - Characteristics of L-glutamine transport during Caco-2 cell differentiation. AB - Glutamine is the main fuel of intestinal epithelial cells, as well as a precursor for the intense nucleotide biosynthesis which arises with the rapid turnover of enterocytes. In order to determine whether glutamine uptake may vary as a function of metabolic demand, glutamine transport across the brush-border membrane of differentiating Caco-2 cells has been investigated. The uptake of L [(3)H]glutamine was measured between day 7 and day 21 post-seeding. Kinetic analysis with glutamine concentrations ranging from 6.25 microM to 12.8 mM revealed the involvement of high affinity Na(+)-dependent (K(t)=110 microM) and low affinity Na(+)-independent (K(t)=900 microM) transport components at day 7. Both components were partially inhibited by L-lysine in a competitive fashion, suggesting that four different systems were responsible for glutamine uptake: B(0), B(0,+), b(0,+) and L. All four systems were present during the differentiation process, with systems L and B(0) being responsible for up to 80% of glutamine uptake. Caco-2 cell differentiation was associated with a marked decrease in L-glutamine uptake, which affected both the Na(+)-dependent and the Na(+)-independent components. In contrast to glucose uptake, the development of L glutamine uptake across the brush-border membrane of Caco-2 cells may reflect an adjustment to cell metabolic demand rather than the progressive appearance of a vectorial transport process. PMID- 11118522 TI - Sequencing and heterologous expression in Saccharomyces cerevisiae of a Cryptococcus neoformans cDNA encoding a plasma membrane H(+)-ATPase. AB - A cDNA containing an open reading frame encoding a putative plasma membrane H(+) ATPase in the human pathogenic basidiomycetous yeast Cryptococcus neoformans was cloned and sequenced by means of PCR and cDNA library hybridization. The cloned cDNA is 3475 bp in length, containing a 2994 bp open reading frame encoding a polypeptide of 997 amino acids. As in the case of another basidiomycetous fungus (Uromyces fabae), the deduced amino acid sequence of CnPMA1 was found to be more homologous to those of P-type H(+)-ATPases from higher plants than to those from ascomycetous fungi. In order to prove the sequenced cDNA corresponds to a H(+) ATPase, it was expressed in Saccharomyces cerevisiae and found to functionally replace its own H(+)-ATPase. Kinetic studies of CnPMA1 compared to ScPMA1 show differences in V(max) values and H(+)-pumping in reconstituted vesicles. The pH optimum and K(m) values are similar in both enzymes. PMID- 11118523 TI - Effects of normal alcohols and isoflurane on lipid headgroup dynamics in nicotinic acetylcholine receptor-rich lipid vesicles. AB - The trend of evidence suggests that general anesthetics act directly on proteins in the neural membrane. However, the fact that the functions of nicotinic acetylcholine receptor (sodium permeability, desensitization rate) are modulated by the composition of the membrane in which it is reconstituted has been thought to be a result of the variation of interactions between acetylcholine receptor and membrane. In this study, protein-lipid interaction at the level of the lipid headgroup was investigated using electron paramagnetic resonance (EPR) and headgroup spin label. Lipid headgroup mobility was evaluated with rotational correlation time from the EPR spectrum. Protein-lipid interaction at headgroup depth was demonstrated from the motionally restricted component of the spectrum. Rotational correlation time increased to 13 ns from 7 ns due to protein-lipid interaction. The effect of anesthetic (ethanol, 1-hexanol, and isoflurane) on protein-lipid interaction was investigated, and the correlation time was 13 ns. It is concluded that the anesthetics used in this study did not alter protein lipid interaction at the level of the lipid headgroup, so far as observed by rotational correlation time, without excluding the possibility that anesthetics that perturb protein-lipid interactions modulate receptor functions via this mechanism. PMID- 11118524 TI - Sugar recognition mutants of the melibiose carrier of Escherichia coli: possible structural information concerning the arrangement of membrane-bound helices and sugar/cation recognition site. AB - Melibiose carrier mutants, isolated by growing cells on melibiose plus the non metabolizable competitive inhibitor thiomethyl-beta-galactoside (TMG), were studied to determine sugar and cation recognition abnormalities. Most of the mutants show good transport of melibiose but have lost the recognition of TMG. In addition, most mutants show little or no transport of lactose. Cation recognition is also affected as all of these mutants have lost the ability to transport protons with melibiose. The amino acids causing these mutations were determined by sequencing the melB gene on the plasmid. The mutations were located on helices I, IV, VII, X and XI. We propose that these five helices are in proximity with each other and that they line the sugar/cation transport channel. PMID- 11118525 TI - Supported planar bilayer formation by vesicle fusion: the interaction of phospholipid vesicles with surfaces and the effect of gramicidin on bilayer properties using atomic force microscopy. AB - We have used magnetic alternating current mode atomic force microscopy (MAC-AFM) to investigate the formation of supported phospholipid bilayers (SPB) by the method of vesicle fusion. The systems studied were dioleoylphosphatidylcholine (DOPC) on mica and mica modified with 3-aminopropyl-triethoxy-silane (APTES), and DOPC vesicles with gramicidin incorporated on mica and APTES-modified mica. The AFM images reveal three stages of bilayer formation: localized disklike features that are single bilayer footprints of the vesicles, partial continuous coverage, and finally complete bilayer formation. The mechanism of supported phospholipid bilayers formation is the fusion of proximal vesicles, rather than surface disk migration. This mechanism does not appear to be affected by incorporation of gramicidin or by surface modification. Once formed, the bilayer develops circular defects one bilayer deep. These defects grow in size and number until a dynamic equilibrium is reached. PMID- 11118526 TI - The optimisation of the helix/helix interaction of a transmembrane dimer is improved by the IMPALA restraint field. AB - A continuous membrane model (IMPALA) was previously developed to predict how hydrophobic spans of proteins insert in membranes (Mol. Mod. 2 (1996) 27). Using that membrane model, we looked for the interactions between several hydrophobic spans. We used the glycophorin A dimer as an archetype of polytopic protein to validate the approach. We find that the native complex do not dislocate when it is submitted to a 10(5) steps optimisation whereas separated spans converge back to a native-like complex in the same conditions. We also observe that IMPALA restraints are not strictly mandatory but do increase the efficiency of the procedure. PMID- 11118527 TI - The current produced by the E779A mutant rat Na(+)/K(+) pump alpha1-subunit expressed in HEK 293 cells. AB - The current (I(p)) generated by the wild-type or the glutamate (E) 779 alanine (A) mutant of the rat Na(+)/K(+) pump alpha1-subunit expressed in HEK 293 cells was studied at 35 degrees C by means of whole-cell recording in Na(+)-free and Na(+)-containing solution. Glutamate 779 is located in the fifth transmembrane domain of the alpha-subunit of the Na(+)/K(+)-ATPase. Compared with the wild type, the E779A mutant exhibited an apparent K(+)(o)-affinity decreased by a factor of 3-4 both in Na(+)-free and in Na(+)-containing media. The competition of Na(+)(o) and K(+)(o) for cation binding sites of the pump remained unchanged. Similarly, in Na(+)-free solution the shape of the I(p)-V curves for various external K(+)-concentrations ([K(+)](o)) was essentially the same. However, in Na(+)-containing solutions the shape of I(p)-V curves from cells expressing the mutant of the rat alpha1-subunit clearly differed from the shape observed in cells expressing the wild-type, but voltage dependence of the pump current persisted. A prominent Na(+)(o)-activated, electrogenic Na(+)-transport mediated by the pump, displaying little voltage dependence in the potential range tested ( 80 to +60 mV), was present in the cells expressing the E779A mutant pump. The data suggest that exchanging E779 for A in the rat Na(+)/K(+) pump alpha1-subunit causes a modest decrease in the apparent K(+)(o) affinity and a profound, Na(+)(o)-dependent alteration in the electrogenicity of the mutant pump expressed in HEK 293 cells. PMID- 11118528 TI - A fluorescence study of the interaction and location of (+)-totarol, a diterpenoid bioactive molecule, in model membranes. AB - (+)-Totarol, a diterpene extracted from Podocarpus totara, has been reported as a potent antioxidant and antibacterial agent. Although the molecular mechanism of action of this hydrophobic molecule remains unknown, recent work made in our laboratory strongly suggests that it could be lipid-mediated. Since (+)-totarol contains a phenolic ring, we have studied the intrinsic fluorescent properties of this molecule, i.e., quantum yield, lifetime, steady-state anisotropy and emission spectra, both in aqueous and in phospholipid phases, in order to obtain information on the interaction and location of (+)-totarol in biomembrane model systems. The phospholipid/water partition coefficient of (+)-totarol was found to be very high (K(p)=1.8x10(4)), suggesting that it incorporates very efficiently into membranes. In order to estimate the transverse location (degree of penetration) of the molecule in the fluid phase of DMPC model membranes, the spin labelled fatty acids 5-NS and 16-NS were used in differential quenching experiments. The results obtained show that (+)-totarol is located in the inner region of the membrane, far away from the phospholipid/water interface. Since (+) totarol protects against oxidative stress, its interaction with an unsaturated fatty acid, trans-parinaric acid, was studied using fluorescence resonance energy transfer. No significant interactions were observed, molecules of trans-parinaric acid distributing themselves randomly amongst those of (+)-totarol in the phospholipid membrane. PMID- 11118529 TI - DOTAP cationic liposomes prefer relaxed over supercoiled plasmids. AB - Cationic liposomes and DNA interact electrostatically to form complexes called lipoplexes. The amounts of unbound (free) DNA in a mixture of cationic liposomes and DNA at different cationic lipid:DNA molar ratios can be used to describe DNA binding isotherms; these provide a measure of the binding efficiency of DNA to different cationic lipid formulations at various medium conditions. In order to quantify the ratio between the various forms of naked DNA and supercoiled, relaxed and single-stranded DNA, and the ratio between cationic lipid bound and unbound DNA of various forms we developed a simple, sensitive quantitative assay using agarose gel electrophoresis, followed by staining with the fluorescent cyanine DNA dyes SYBR Green I or SYBR Gold. This assay was compared with that based on the use of ethidium bromide (the most commonly used nucleic acid stain). Unlike ethidium bromide, SYBR Green I DNA sensitivity and concentration-dependent fluorescence intensity were identical for supercoiled and nicked-relaxed forms. DNA detection by SYBR Green I in solution is approximately 40-fold more sensitive than by ethidium bromide for double-stranded DNA and approximately 10-fold for single-stranded DNA, and in agarose gel it is 16-fold more sensitive for double stranded DNA compared with ethidium bromide. SYBR Gold performs similarly to SYBR Green I. This study shows that: (a) there is no significant difference in DNA binding isotherms to the monocationic DOTAP (DOTAP/DOPE) liposomes and to the polycationic DOSPA (DOSPA/DOPE) liposomes, even when four DOSPA positive charges are involved in the electrostatic interaction with DNA; (b) the helper lipids affect DNA binding, as DOTAP/DOPE liposomes bind more DNA than DOTAP/cholesterol; (c) in the process of lipoplex formation, when the DNA is a mixture of two forms, supercoiled and nicked-relaxed (open circular), there is a preference for the binding to the cationic liposomes of plasmid DNA in the nicked-relaxed over the supercoiled form. This preference is much more pronounced when the cationic liposome formulation is based on the monocationic lipid DOTAP than on the polycationic lipid DOSPA. The preference of DOTAP formulations to bind to the relaxed DNA plasmid suggests that the binding of supercoiled DNA is weaker and easier to dissociate from the complex. PMID- 11118530 TI - The cell wall integrity/remodeling MAPK cascade is involved in glucose activation of the yeast plasma membrane H(+)-ATPase. AB - Glucose triggers transcriptional and post-transcriptional mechanisms that increase the amount and the activity of Saccharomyces cerevisiae plasma membrane H(+)-ATPase. In a previous study, we found that a mutation in the Rsp5 ubiquitin protein ligase enzyme affected the post-transcriptional activation of the enzyme by glucose. Mutations at the RSP5 locus alter the glucose-triggered K(m) decrease. In a genetic screening for multicopy suppressors of the rsp5 mutation, we identified the WSC2/YNL283c gene. Deletion of the WSC2 gene disturbs ATPase activation by glucose, abolishing the K(m) decrease that occurs during this process. Wsc2 is a component of the PKC1-MPK1 mitogen-activated protein kinase (MAPK) signaling pathway that controls the cell wall integrity. Deletion of the MPK1/SLT2 gene disturbs the glucose-triggered K(m) decrease in ATPase. PMID- 11118531 TI - Desensitization, surface expression, and glycosylation of a functional, epitope tagged type I PACAP (PAC(1)) receptor. AB - To study desensitization and glycosylation of the type I pituitary adenylate cyclase-activating polypeptide (PACAP) receptor (PAC(1)R), a hemagglutinin (HA) epitope was inserted within the N-terminal extracellular domain, allowing immunological detection of PAC(1)R both in intact and permeabilized cells. PAC(1)R was tagged without loss of functions in ligand binding and ligand stimulated cAMP production. In transiently transfected COS-7 cells, PAC(1)R was localized both in the plasma membrane and the cytoplasm around the nucleus. By immunoblot analysis, the immunoreactive bands with relative molecular masses ranging from 45 to 70 kDa were detected in the membrane fractions of PAC(1)R expressing COS-7 cells. Digestion of the membranes with endoglycosidase F or treatment of the cells with tunicamycin decreased the size of the receptor to major bands of smaller size (approximately 45 and 48 kDa), suggesting that these two forms of PAC(1)R represent core proteins. Flow cytometric analysis indicated that the agonist promoted a disappearance of cell surface receptor. In accordance with this observation, preexposure of cells to PACAP38 induced a desensitization of PAC(1)R to the agonist response, although it did not cause a reduction in PAC(1)R mRNA or protein level and even slightly elevated them. These results suggest that agonist-induced desensitization of PAC(1)R involves the receptor sequestration. PMID- 11118532 TI - Synthesis and thermotropic characterization of a homologous series of racemic beta-D-glucosyl dialkylglycerols. AB - The phase behaviour of aqueous dispersions of a series of synthetic 1,2-di-O alkyl-3-O-(beta-D-glucosyl)-rac-glycerols with both odd and even hydrocarbon chain lengths was studied by differential scanning calorimetry and low angle X ray diffraction (XRD). Thermograms of these lipids show a single, strongly energetic phase transition, which was shown to correspond to either a lamellar gel/liquid crystalline (L(beta)/L(alpha)) phase transition (short chain compounds, n < or =14 carbon atoms) or a lamellar gel/inverted hexagonal (L(beta)/H(II)) phase transition (longer chain compounds, n > or =15 carbon atoms) by XRD. The shorter chain compounds may exhibit additional transitions at higher temperatures, which have been identified as lamellar/nonlamellar phase transitions by XRD. The nature of these nonlamellar phases and the number of associated intermediate transitions can be seen to vary with chain length. The thermotropic phase properties of these lipids are generally similar to those reported for the corresponding 1,2-sn-diacyl alpha- and beta-D-glucosyl counterparts, as well as the recently published 1, 2-dialkyl-3-O-(beta-D glycosyl)-sn-glycerols. However, the racemic lipids studied here show no evidence of the complex patterns of gel phase polymorphism exhibited by the above mentioned compounds. This suggests that the chirality of the glycerol molecule, by virtue of its position in the interfacial region, may significantly alter the phase properties of a lipid, perhaps by controlling the relative positions of hydrogen bond donors and acceptors in the polar region of the membrane. PMID- 11118533 TI - Dipalmitoylphosphatidylcholine membranes modified with zeaxanthin: numeric study of membrane organisation. AB - The model of a dipalmitoylphosphatidylcholine (DPPC) bilayer containing a xanthophyll pigment zeaxanthin (ZEA) is proposed. The model is based on the ten state Pink-Green-Chapman model of a lipid monolayer. The Monte Carlo method of computer simulation has been applied. Our model of the lipid membrane consists of two lipid monolayers with ZEA molecules spanning the lipid bilayer. The concentration of ZEA molecules is assumed to be conserved. Within the model, the interactions between lipid monolayers in a bilayer exist through ZEA molecules only. The experimental data concerning the aggregation of ZEA in DPPC from the literature and from our research were applied as a criterion to fit the model parameters. The model gives the dependences of the main phase transition temperature on ZEA/DPPC molar ratio, the percentage of ZEA in a monomeric form on ZEA/DPPC molar ratio and on temperature. The dependences obtained within the model and the experimental ones are in qualitative agreement. The influence of intermolecular interaction parameters on ZEA aggregation has been discussed. The differences between the model and the experimental results concerning mainly the pattern of ZEA aggregation have been discussed. Analyses of the lipid microconfiguration allow to advance the hypothesis concerning the influence of ZEA on the membrane permeability. PMID- 11118534 TI - Membrane impermeant antioestrogens discriminate between ligand- and voltage-gated cation channels in NG108-15 cells. AB - Native 5-HT(3) and AChR ligand-gated cation channels can be inhibited (blocked) by the non-steroidal antioestrogen tamoxifen. However, the exact site and mechanism of inhibition by tamoxifen on these channels remain unclear. We have investigated the action of the membrane impermeant quaternary derivative, ethylbromide tamoxifen (EBT), on native ligand-gated 5-HT(3) receptor channels and voltage-gated K(+) channels in NG108-15 cells using whole cell patch clamp. Extracellular EBT inhibited whole cell cationic currents of 5-HT(3) receptors with IC(50) of 0.22+/-0.4 microM (n(H)=1.05+/-0.2). The channel block was characterised by voltage independent and use independent behaviour (similar to that of tamoxifen). EBT was unable to inhibit voltage-gated K(+) currents in NG108-15 cells. This was in contrast to the inhibition by tamoxifen which, at similar concentrations, accelerated the apparent inactivation of these outward K(+) currents. The inhibition of 5-HT(3) receptors by a membrane impermeant derivative of tamoxifen supports the view that the binding site for antioestrogens is extracellular and the inhibition is not mediated through genomic/transcriptional activity. PMID- 11118535 TI - Involvement of the C-terminal part of Pseudomonas fluorescens OprF in the modulation of its pore-forming properties. AB - The major outer-membrane protein, OprF, from the psychrotrophic bacterium Pseudomonas fluorescens undergoes a reduction of its conductance value (from 250 pS to 80 pS) when the growth temperature is shifted from 28 degrees C to 8 degrees C. The involvement of changes in tertiary or quaternary structure in this behaviour, was implied by enzymatic digestion experiments in which OprFs purified from 8 degrees C and 28 degrees C cultures showed different accessibility to pronase. Resistant proteolytic fragments of 19 kDa, obtained from both OprF preparations, were identified as the N-terminal half of the native protein. These 19 kDa fragments induced ion channels in planar lipid bilayers with similar conductance values of 65-75 pS in 1 M NaCl, in contrast to the native proteins. Thus, the C-terminal part of the protein is required for the growth temperature dependent modulation of OprF channel-forming properties. LPS was not detected on the proteolytic fragments while it was found in similar amounts on the native OprFs. These results suggest the LPS/porin association occurs through the C terminal part of the porin. Radiolabelling experiments showed different phosphorylation levels of LPS for 8 degrees C and 28 degrees C cultures. Thus, in response to growth temperature, the structural modification of the LPS could be associated to the modulation of OprF pore size. PMID- 11118536 TI - Effect of low external calcium on the ERG current of NG108-15 cells. AB - K(+) currents through ERG (ether-a-go-go related gene) channels were recorded in whole-cell voltage clamped NG108-15 neuroblastomaxglioma hybrid cells. The channels were fully activated by low holding potential (V(H)=-20 mV) and long depolarizing prepulses. Hyperpolarizing pulses elicited inward currents which deactivated after reaching a peak. Lowering [Ca(2+)](o) from 5 to 1. 5 or 0.5 mM decreased tau(-1), the rate constant of deactivation. The effect can be explained by a shift of the tau(-1)(V) curve to more negative potentials caused by an increase in surface charge density. Plotting tau(-1) against [Ca(2+)](o) for different potentials yielded straight lines; their slope was independent of potential at -140 to -120 mV and decreased at more positive potentials. The time to peak curve and the maximum of the steady-state inward current were also shifted to more negative potentials. In addition, peak ERG inward current increased. Raising [Ca(2+)](o) from 5 to 10 mM accelerated deactivation and decreased the peak current. 5 mM Ba(2+) affected tau(-1) similarly and inhibited peak current more strongly whereas 5 mM Mg(2+) was less potent. As found by Faravelli et al. (J. Physiol. 496 (1996) 13), bath solutions devoid of divalent cations (0 Ca(2+), 0 Mg(2+), 0.1 or 1.1 mM EGTA) abolished deactivation almost completely. The phenomenon was seen with bath containing either 40 or 6.5 mM K(+). Its occurrence was favored by raising the temperature to 34 degrees C. It suggests a particular requirement of channel closing for Ca(2+). PMID- 11118537 TI - Organisation of xanthophyll pigments lutein and zeaxanthin in lipid membranes formed with dipalmitoylphosphatidylcholine. AB - Carotenoid pigments and in particular xanthophylls play several physiological functions in plant and animal membranes. Xanthophylls are present in biological membranes in the form of pigment-protein complexes but also as direct components of lipid phase. The biological activity of carotenoids in membranes depends on a molecular organisation of pigments in lipid bilayers, in particular the localisation, orientation and aggregational state. In the present work the organisation of lutein- and zeaxanthin-containing lipid membranes was analysed with the application of electronic absorption spectroscopy. Both xanthophyll pigments incorporated to the dipalmitoylphosphatidylcholine (DPPC) unilamellar liposomes form H-type molecular aggregates, manifested by the hypsochromic shift of the main absorption band of carotenoids. The aggregation of lutein and zeaxanthin in DPPC membranes was observed even at relatively low concentrations of a pigment in the lipid phase (1-5 mol%). Gaussian analysis of the absorption spectra of lutein and zeaxanthin in DPPC membranes in terms of the exciton splitting theory revealed the formation of different molecular structures of pigments interpreted as dimers, trimers, tetramers and large aggregates. The fraction of lutein and zeaxanthin in the monomeric form was found to depend on the physical state of the lipid phase. Pronounced monomerisation of lutein and zeaxanthin was observed as accompanying the transition from the P(beta)' phase to the L(alpha) phase of DPPC, mostly at the expense of the trimeric and tetrameric forms. The fraction of monomers of lutein is always lower by 10-30% than that of zeaxanthin under the same experimental conditions. Different organisational forms of lutein and zeaxanthin in the model system studied are discussed in terms of possible physiological functions of these pigments in the membranes of the retina: zeaxanthin in the protection of the lipid phase against oxidative damage and lutein in absorbing short wavelength radiation penetrating retina membranes. PMID- 11118538 TI - Vibrio cholerae cytolysin: assembly and membrane insertion of the oligomeric pore are tightly linked and are not detectably restricted by membrane fluidity. AB - Hemolytic strains of Vibrio cholerae secrete a cytolysin that, upon binding as a monomer, forms pentameric pores in animal cell membranes. Pore formation is inhibited at low temperature and in the absence of cholesterol. We here posed the following questions: firstly, can oligomerization be observed in the absence of pore formation? Secondly, is membrane fluidity responsible for the effect of temperature or of cholesterol upon pore formation? The first issue was approached by chemical cross-linking, by electrophoretic heteromer analysis, and by electron microscopy. None of these methods yielded any evidence of a non-lytic pre-pore oligomer. The second question was addressed by the use of two susceptible liposome models, consisting of cholesterol admixed to bovine brain lipids and to asolectin, respectively. The two liposome species clearly differed in membrane fluidity as judged by diphenylhexatriene fluorescence polarization. Nevertheless, their permeabilization by the cytolysin decreased with temperature in a closely parallel fashion, virtually vanishing at 5 degrees C. Omission of cholesterol from the liposomes uniformly led to an increase in membrane fluidity but prevented permeabilization by the cytolysin. The effects of temperature and of cholesterol upon cytolysin activity are thus not mediated by fluidization of the target membrane. The findings of our study distinguish V. cholerae cytolysin from several previously characterized pore-forming toxins. PMID- 11118539 TI - Contributions of charged residues in a cytoplasmic linking region to Na channel gating. AB - Na channels inactivate quickly after opening, and the very highly positively charged cytoplasmic linking region between homologous domains III and IV of the channel molecule acts as the inactivation gate. To test the hypothesis that the charged residues in the domain III to domain IV linker have a role in channel function, we measured currents through wild-type and two mutant skeletal muscle Na channels expressed in Xenopus oocytes, each lacking two or three charged residues in the inactivation gate. Microscopic current measures showed that removing charges hastened activation and inactivation. Macroscopic current measures showed that removing charges altered the voltage dependence of inactivation, suggesting less coupling of the inactivation and activation processes. Reduced intracellular ionic strength shifted the midpoint of equilibrium activation gating to a greater extent, and shifted the midpoint of equilibrium inactivation gating to a lesser extent in the mutant channels. The results allow the possibility that an electrostatic mechanism contributes to the role of charged residues in Na channel inactivation gating. PMID- 11118540 TI - Heterogeneous distribution of plasma membrane glycoconjugates in pancreatic acinar cells. AB - Flow-cytometric studies of lectin binding to individual acinar cells have been carried out in order to analyse the distribution of membrane glycoconjugates in cells from different areas of the pancreas: duodenal lobule (head) and splenic lobule (body and tail). The following fluoresceinated lectins were used: wheat germ agglutinin (WGA), Tetragonolobus purpureus agglutinin (TP) and concanavalin A (Con A), which specifically bind to N-acetyl D-glucosamine and sialic acid, L fucose and D-mannose, respectively. In both pancreatic areas, two cell populations (R1 and R2) were identified according to the forward scatter (size). On the basis of their glycoconjugate pattern, R1 cells displayed higher density of WGA and TP receptors than R2 cells throughout the pancreas. Although no difference in size was found between the cells from duodenal and splenic lobules, N-acetyl D-glucosamine and/or sialic acid and L-fucose residues were more abundant in plasma membrane cell glycoconjugates from the duodenal lobule. The results provide evidence for biochemical heterogeneity among individual pancreatic cells according to the distribution of plasma membrane glycoconjugates. PMID- 11118541 TI - Preparation of immunoliposomes bearing poly(ethylene glycol)-coupled monoclonal antibody linked via a cleavable disulfide bond for ex vivo applications. AB - Several methods for the preparation of sterically stabilized immunoliposomes (SIL) have recently been described. This report examines an established method for coupling anti-CD34 My10 mAb to poly(ethylene glycol)-liposomes (PEG liposomes) containing the anchor pyridyldithiopropionylamino-PEG phosphatidylethanolamine (PDP-PEG-PE) via a cleavable disulfide bond. Efficient attachment of pyridyldithio-derivatized mAb took place (equivalent to coupling ca. 70% of total input protein) at 2 mol percent of the functionalized PEG-lipid. The My10-SIL bound specifically to CD34+ cells (human leukemic KG-1a and hematopoietic progenitor cells) and the extent of binding was a function of liposomal lipid concentration, the mAb density in the liposome surface and the CD34 cell expression. In mixtures with CD34- cells (CHO or Jurkat), CD34+KG-1a cells were determined by flow cytometry at percentages (1-4%) similar to those reported in clinical samples (such as cord blood, mobilized peripheral blood and bone marrow) using a direct immunostaining with My10-SIL. The disulfide bond was stable in cell culture medium (10% of fetal calf serum) during 8 h and cell-bound SIL can be released from cells by treatment with dithiothreitol as reducing agent under mild conditions (1 h of incubation with 50 mM DTT at 20 degrees C). SIL binding and subsequent dithiothreitol treatment did not influence the cell viability. Our approach should contribute to the development of targetable liposomal vehicles to CD34+ cells for use in ex vivo conditions as sorting of hematopoietic stem cells. PMID- 11118542 TI - Localization and rearrangement modulation of the N-terminal arm of the membrane bound major coat protein of bacteriophage M13. AB - During infection the major coat protein of the filamentous bacteriophage M13 is in the cytoplasmic membrane of the host Escherichia coli. This study focuses on the configurational properties of the N-terminal part of the coat protein in the membrane-bound state. For this purpose X-Cys substitutions are generated at coat protein positions 3, 7, 9, 10, 11, 12, 13, 14, 15, 17, 19, 21, 22, 23 and 24, covering the N-terminal protein part. All coat protein mutants used are successfully produced in mg quantities by overexpression in E. coli. Mutant coat proteins are labeled and reconstituted into mixed bilayers of phospholipids. Information about the polarity of the local environment around the labeled sites is deduced from the wavelength of maximum emission using AEDANS attached to the SH groups of the cysteines as a fluorescent probe. Additional information is obtained by determining the accessibility of the fluorescence quenchers acrylamide and 5-doxyl stearic acid. By employing uniform coat protein surroundings provided by TFE and SDS, local effects of the backbone of the coat proteins or polarity of the residues could be excluded. Our data suggest that at a lipid to protein ratio around 100, the N-terminal arm of the protein gradually enters the membrane from residue 3 towards residue 19. The hinge region (residues 17-24), connecting the helical parts of the coat protein, is found to be more embedded in the membrane. Substitution of one or more of the membrane-anchoring amino acid residues lysine 8, phenylalanine 11 and leucine 14, results in a rearrangement of the N-terminal protein part into a more extended conformation. The N-terminal arm can also be forced in this conformation by allowing less space per coat protein at the membrane surface by decreasing the lipid to protein ratio. The influence of the phospholipid headgroup composition on the rearrangement of the N-terminal part of the protein is found to be negligible within the range thought to be relevant in vivo. From our experiments we conclude that membrane-anchoring and space-limiting effects are key factors for the structural rearrangement of the N-terminal protein part of the coat protein in the membrane. PMID- 11118543 TI - Molecular characterization of taurine transport in bovine aortic endothelial cells. AB - Cultured bovine aortic endothelial (BAE) cells expressed a Na(+)/Cl(-)-dependent taurine uptake activity that saturated with an apparent K(0.5) of approximately 4.9 microM for taurine and was inhibited by beta-alanine, guanidinoethane sulfonate, and homotaurine. We isolated a taurine transporter clone from a BAE cell cDNA library that revealed >91% sequence identity at the amino acid level to the previously cloned high-affinity mammalian taurine transporters. The biochemical and pharmacological properties of the bovine taurine transporter cDNA expressed in Xenopus oocyte was similar to those of the high-affinity taurine transporter. Surprisingly, F(-) blocked taurine uptake in BAE cells with an IC(50) of approximately 17.5 mM. The endogenous taurine uptake was also inhibited by the protein kinase C activator phorbol 12-myristate 13-acetate, but not by its inactive analog, 4 alpha-phorbol 12,13-didecanoate. The endogenous uptake was stimulated, however, by hypertonic stress and the increase was due to an increase in the V(max) of taurine uptake. Our results provide the first description of a molecular mechanism that may be responsible for maintaining the intracellular taurine content in the endothelial cells. PMID- 11118544 TI - Influence of transmembrane peptides on bilayers of phosphatidylcholines with different acyl chain lengths studied by solid-state NMR. AB - The molecular orientation in a lipid membrane of the peptide fragment VEYAGIALFFVAAVLTLWSMLQYLSAAR (phosphatidylglycerophosphate synthase (Pgs) peptide E) of an integral membrane protein, Pgs, in Escherichia coli has been investigated by solid-state 15N nuclear magnetic resonance (NMR) on macroscopically aligned lipid bilayers. The secondary structure of the peptide in lipid vesicles was determined by circular dichroism spectroscopy. Furthermore, the phase behaviour of the Pgs peptide E/dierucoylphosphatidylcholine (DEruPC)/water system was determined by (2)H, (31)P and 15N solid-state NMR spectroscopy. The phase behaviour obtained was then compared to that of the Pgs peptide E solubilised in dioleoylphosphatidylcholine and water that was previously studied by Morein et al. [Biophys. J. 73 (1997) 3078-3088]. This was aimed to answer the question whether a difference in the length of the hydrophobic part of this peptide and the hydrophobic thickness of the lipid bilayer (hydrophobic mismatch) will affect the phase behaviour. The peptide mostly has a transmembrane orientation and is in an alpha-helical conformation. An isotropic phase is formed in DEruPC with high peptide content (peptide/lipid molar ratio (p/l) > or =1:15) and high water content (> or =50%, w/w) at 35 degrees C. At 55 and 65 degrees C an isotropic phase is induced at high water content (> or =50%, w/w) at all peptide contents studied (no isotropic phase forms in the lipid/water system under the conditions in this study). At high peptide contents (p/l> or =1:15) an isotropic phase forms at 20 and 40% (w/w) of water at 55 and 65 degrees C. A comparison of the phase behaviour of the two homologous lipid systems reveals striking similarities, although the thicknesses of the two lipid bilayers differ by 7 A. This suggests that the rationalisation of the phase behaviour in terms of the hydrophobic mismatch is not applicable to these systems. The C-terminus of Pgs peptide E is amphiphilic and a considerable part of the peptide is situated outside the hydrophobic part of the bilayer, a property of the peptide that to a large extent will affect the lipid/peptide phase behaviour. PMID- 11118545 TI - Effect of angiotensin II non-peptide AT(1) antagonist losartan on phosphatidylethanolamine membranes. AB - Losartan was found to affect both the thermotropic behavior and molecular mobility of dimyristoyl- and dipalmitoyl-phosphatidylcholine membranes (Theodoropoulou and Marsh, Biochim. Biophys. Acta 1461 (1999) 135-146). At low concentrations, the antagonist is located close to the interfacial region of the phosphatidylcholine bilayer while at high mole fractions it inserts deeper in the bilayers. In the present study, we investigated the interactions of losartan with phosphatidylethanolamine membranes using differential scanning calorimetry (DSC), electron spin resonance (ESR) and 31P nuclear magnetic resonance (NMR) spectroscopy. DSC showed that the antagonist affected the thermotropic transitions of dimyristoyl-, dipalmitoyl- and dielaidoyl-phosphatidylethanolamine membranes (DMPE, DPPE and DEPE, respectively). ESR spectroscopy showed that the interaction of losartan with phosphatidylethanolamine membranes is more superficial than in the case of phosphatidylcholine bilayers. Additionally, losartan increased the spin-spin broadening of 12-PESL spin labels in the gel phase of DMPE and DPPE membranes, while in the case of DEPE membranes the opposite effect was observed. (31)P-NMR showed that the antagonist stabilizes the fluid lamellar phase of DEPE membranes relative to the hexagonal H(II) phase. Our results show that losartan affects the thermotropic behavior of phosphatidylethanolamine membranes, while the molecular mobility of the membranes is not affected greatly. Furthermore, its interactions with phosphatidylethanolamine membranes are more superficial than with phosphatidylcholine bilayers. PMID- 11118546 TI - The distribution of alpha-tocopherol in mixed aqueous dispersions of phosphatidylcholine and phosphatidylethanolamine. AB - The effect of alpha-tocopherol on the structure and phase behaviour of mixed aqueous dispersions of phosphatidylcholine and phosphatidylethanolamine has been examined by synchrotron X-ray diffraction. Equimolar mixtures of dioleoylphosphatidylethanolamine:dioleoylphosphatidylcholine and dimyristoylphosphatidylcholine:dioleoylphosphatidylethanolamine did not show evidence of phase separation of an inverted hexagonal structure typical of alpha tocopherol and phosphatidylethanolamine from lamellar phase. Mixed dispersions of dioleoyl derivatives of phosphatidylethanolamine:phosphatidylcholine (3:1) form a typical miscible gel phase at low temperatures but which phase separates into lamellar liquid-crystal and inverted hexagonal phases at temperatures greater than 65 degrees C. The presence of 1, 2 or 5 mol% alpha-tocopherol caused a decrease in the temperature at which the inverted hexagonal phase appears. Phase separation of non-lamellar phase from lamellar gel phase can be detected in the presence of 7.5 and 10 mol% alpha-tocopherol, indicating a limited capacity of the phosphatidylcholine to incorporate alpha-tocopherol into the lamellar domain. A partial phase diagram of the ternary mixture has been constructed from the X ray scattering data. It was concluded that there is no preferential interaction of alpha-tocopherol with phosphatidylethanolamine in mixed aqueous dispersions containing phosphatidylcholines. PMID- 11118547 TI - Stabilization of O-pyromellitylgramicidin channels in bilayer lipid membranes through electrostatic interaction with polylysines of different chain lengths. AB - Functioning of membrane proteins, in particular ionic channels, can be modulated by alteration of their arrangement in membranes. We addressed this issue by studying the effect of different chain length polylysines on the kinetics of ionic channels formed in a bilayer lipid membrane (BLM) by O pyromellitylgramicidin carrying three negative charges at the C-terminus. The method of sensitized photoinactivation was applied to the analysis of the channel association-dissociation kinetics (characterized by the exponential factor of the curve describing the time course of the flash-induced decrease in the transmembrane current, tau). Addition of polylysine to the bathing solutions of BLM led to the deceleration of the photoinactivation kinetics, i.e. to the increase in tau. It was shown here that for a series of polylysines differing in their chain lengths, the value of tau grew as their concentration increased above a threshold level until at a certain concentration of each polylysine tau reached maximum. At higher polylysine concentrations tau began to decrease and finally became close to the control level observed in the absence of polylysine. With lengthening of the polylysine chain the maximum value of tau increased, the concentration dependence became steeper, and the threshold concentration decreased. The increase in the ionic strength of the medium shifted the concentration dependence of tau to higher polylysine concentrations and decreased the maximum value of tau. It was concluded that the increase in tau was caused by the formation of domains of O-pyromellitylgramicidin molecules induced by binding of polylysines. This can be related to functional aspects of polycation-induced sequestering of negatively charged transmembrane peptides in neutral membranes. PMID- 11118548 TI - The structure and thermotropic phase behaviour of dipalmitoylphosphatidylcholine codispersed with a branched-chain phosphatidylcholine. AB - The structure and thermotropic phase behaviour of a fully hydrated binary mixture of dipalmitoylphosphatidylcholine and a branched-chain phosphatidylcholine, 1, 2 di(4-dodecyl-palmitoyl)-sn-glycero-3-phosphocholine, were examined using differential scanning calorimetry, synchrotron X-ray diffraction and freeze fracture electron microscopy. The branched-chain lipid forms a nonlamellar phase when dispersed alone in aqueous medium. Mixed aqueous dispersions of the two phospholipids containing less than 33 mol% of the branched-chain lipid form lamellar phases over the whole temperature range were studied (4 degrees C to 60 degrees C). When present in proportions greater than 33 mol% it induces a hexagonal phase in mixed aqueous dispersions with dipalmitoylphosphatidylcholine at temperatures above the fluid phase transition. At temperatures below 35 degrees C a hexagonal phase coexists with a gel bilayer phase. The lamellar<- >nonlamellar transition can be explained satisfactorily on the basis of the shape of the molecule expressed in terms of headgroup and chain cross-sectional areas. At temperatures below 35 degrees C macroscopic phase separation of two gel phases takes place. Freeze-fracture electron microscopy revealed that one gel phase consists of bilayers with a highly regular, periodic superstructure (macro ripples) whereas the other phase forms flat, planar bilayers. The macro-ripple phase appears to represent a relaxation structure required to adapt to the packing constraints imposed by the incorporation of the branched-chain lipid into the dipalmitoylphosphatidylcholine host bilayer. The data suggest that structural changes that take place on cooling the mixed dispersion below the lamellar<- >nonlamellar phase transition temperature cannot be adequately described using the molecular form concept. Instead it is necessary to take into account the detailed molecular form of the guest lipid as well as its physical properties. PMID- 11118549 TI - Acceleration of phospholipid flip-flop in the erythrocyte membrane by detergents differing in polar head group and alkyl chain length. AB - The detergents, alkyltrimethylammonium bromide, N-alkyl-N, N-dimethyl-3-ammonio-1 propanesulfonate (zwittergent), alkane sulfonate, alkylsulfate, alkyl-beta-D glucopyranoside, alkyl-beta-D-maltoside, dodecanoyl-N-methylglucamide, polyethylene glycol monoalkyl ether and Triton X-100, all produce a concentration dependent acceleration of the slow passive transbilayer movement of NBD-labeled phosphatidylcholine in the human erythrocyte membrane. Above a threshold concentration, which was well below the CMC and characteristic for each detergent, the flip rate increases exponentially upon an increase of the detergent concentration in the medium. The detergent-induced flip correlates with reported membrane-expanding effects of the detergents at antihemolytic concentrations. From the dependence of the detergent concentration required for a defined flip acceleration on the estimated membrane volume, membrane/water partition coefficients for the detergents could be determined and effective detergent concentrations in the membrane calculated. The effective membrane concentrations are similar for most types of detergents but are 10-fold lower for octaethylene glycol monoalkyl ether and Triton X-100. The effectiveness of a given type of detergent is rather independent of its alkyl chain length. Since detergents do not reduce the high temperature dependence of the flip process the detergent-induced flip is proposed to be due to an enhanced probability of formation of transient hydrophobic structural defects in the membrane barrier which may result from perturbation of the interfacial region of the bilayer by inserted detergent molecules. PMID- 11118550 TI - Membrane dynamics as seen by fourier transform infrared spectroscopy in a cyanobacterium, Synechocystis PCC 6803. The effects of lipid unsaturation and the protein-to-lipid ratio. AB - The roles of lipid unsaturation and lipid-protein interactions in maintaining the physiologically required membrane dynamics were investigated in a cyanobacterium strain, Synechocystis PCC 6803. The specific effects of lipid unsaturation on the membrane structure were addressed by the use of desaturase-deficient (desA( )/desD(-)) mutant cells (which contain only oleic acid as unsaturated fatty acid species) of Synechocystis PCC 6803. The dynamic properties of the membranes were determined from the temperature dependence of the symmetric CH(2) stretching vibration frequency, which is indicative of the lipid fatty acyl chain disorder. It was found that a similar membrane dynamics is maintained at any growth temperature, in both the wild-type and the mutant cell membranes, with the exception of mutant cells grown at the lower physiological temperature limit. It seems that in the physiological temperature range the desaturase system of the cells can modulate the level of lipid desaturation sufficiently to maintain similar membrane dynamics. Below the range of normal growth temperatures, however, the extent of lipid disorder was always higher in the thylakoid than in the cytoplasmic membranes prepared from the same cells. This difference was attributed to the considerable difference in protein-to-lipid ratio in the two kinds of membranes, as determined from the ratio of the intensities of the protein amide I band and the lipid ester C&z.dbnd6;O vibration. The contributions to the membrane dynamics of an ab ovo present 'structural' lipid disorder due to the protein-lipid interactions and of a thermally induced 'dynamic' lipid disorder could be distinguished. PMID- 11118551 TI - Volume-activated K(+)(Rb(+)) efflux in lactating rat mammary tissue. AB - The effect of cell swelling, induced by a hyposmotic shock, on K(+)(Rb(+)) efflux from lactating rat mammary tissue explants has been studied. A hyposmotic challenge increased the fractional release of K(+)(Rb(+)) from mammary tissue in the absence and presence of the loop-diuretic bumetanide (100 microM). However, the volume-sensitive moiety of K(+)(Rb(+)) efflux was proportionately larger when bumetanide was present in the incubation medium. On the other hand, a hyposmotic shock appeared to reduce the bumetanide-sensitive component of K(+)(Rb(+)) efflux. The increase in K(+)(Rb(+)) efflux, induced by cell swelling, was dependent upon the extent of the hyposmotic challenge. In the presence of bumetanide, substituting Cl(-) with NO(3)(-) reduced the initial increase in volume-sensitive K(+)(Rb(+)) efflux. However, volume-sensitive K(+)(Rb(+)) release was prolonged in the presence of NO(3)(-). Volume-activated K(+)(Rb(+)) efflux from rat mammary tissue explants was inhibited by quinine. Cell swelling increased the intracellular concentration of Ca(2+) in a fashion which depended on the presence of extracellular Ca(2+). However, removing extracellular Ca(2+) did not inhibit volume-activated K(+)(Rb(+)) efflux from rat mammary tissue explants. The results are consistent with the presence of volume-activated K(+) channels in lactating rat mammary tissue. Volume-activated K(+) efflux may play a central role in mammary cell volume regulation. PMID- 11118552 TI - Functional characterization of purified zinc transporter from renal brush border membrane of rat. AB - Major zinc binding protein purified from renal brush border membrane (BBM) (R. Kumar, R. Prasad, Biochim. Biophys. Acta 1419 (1999) 23) was reconstituted into liposomes and its functional characteristics were investigated. Physical incorporation of the major zinc binding protein into the proteoliposomes was checked by SDS-PAGE, which showed a single band on silver staining. The structural integrity of the proteoliposomes was assessed by phase contrast microscopy, which revealed the proteoliposomes as globular structures and intact boundaries. Further structural integrity/leakiness of the proteoliposomes was checked by monitoring efflux of Zn(2+) from the pre-loaded proteoliposomes in the presence of either 2 mM Ca(2+) or Cd(2+) or Zn(2+). It was observed that even after 2 h of the initiation of efflux, 85-95% of Zn(2+) was retained in the proteoliposomes, thereby indicating that proteoliposomes were not leaky and maintained structural integrity during the uptake study. Zinc uptake into the proteoliposomes followed Michaelis-Menten kinetics with affinity constant (K(m)) of 1.03 mM and maximal velocity (V(max)) of 1333 nmol/mg protein per min. The uptake process followed first-order kinetics with a rate constant (k) of 1. 09x10(-3) s(-1). The specificity of zinc transport system was determined by studying the interaction of divalent cations viz. Ca(2+) and Cd(2+) with the zinc uptake. It was observed that Cd(2+) competitively inhibited the zinc uptake process with inhibitory concentration (K(i)) of 2.9 mM. Kinetic analysis of inhibitory effect of Cd(2+) on zinc uptake revealed an increase in K(m) to 1.74 mM without influencing V(max). Zn(2+) uptake into the proteoliposomes was found to be temperature sensitive and Arrhenius plot showed a breakpoint at 27 degrees C. The apparent energies of activation (E(a)) were found to be 7.09 and 2.74 kcal/mol below and above the breakpoint, respectively. The initial velocity of Zn(2+) uptake increased with the increase in outwardly directed proton gradient ([H](i) greater than [H](o)). The Zn(2+) uptake was inhibited by DCCD, thereby suggesting the involvement of -COOH groups in the translocation of Zn(2+) across the lipid bilayer. The ratio of acidic to basic amino acids (1.26) strongly indicates that it is an acidic protein. The cysteine content in this protein was insignificant, which further corroborates the possibility that the acidic amino acids might be prominent candidates for binding to zinc. The findings of the present study confirms that 40 kDa major zinc binding glycoprotein purified from renal BBM is a zinc transporter involved in the influx of Zn(2+) into the epithelial cells of the renal tubular system. PMID- 11118553 TI - The effect of dimethyl sulphoxide on the structure and phase behaviour of palmitoleoylphosphatidylethanolamine. AB - The thermotropic phase behaviour and structure of a nonbilayer-forming lipid, 1 palmitoyl-2-oleoyl-phosphatidylethanolamine, dispersed in water and in aqueous solutions of up to 50 wt% dimethyl sulphoxide (DMSO) have been characterised using synchrotron X-ray diffraction methods. It was found that the presence of DMSO in the solvent induced an increase in the temperature of lamellar-gel to lamellar-liquid-crystal phase transition and a decrease in the temperature of the lamellar-liquid-crystal to inverted-hexagonal phase transition of the phospholipid. The presence of DMSO also caused a decrease in the X-ray repeat spacings of all the phases studied. Electron density profiles of the phospholipid dispersed in water and 50 wt% DMSO in the bilayer gel state were calculated. The presence of 50 wt% DMSO caused the apparent disappearance of the solvent layer separating phospholipid bilayers in the gel state. The results suggest that DMSO contributes to the bilayer electron density profile and that the amphiphilic solvent molecules partition into the interfacial region. PMID- 11118555 TI - ATPase activity and transport by a cGMP transporter in human erythrocyte ghosts and proteoliposome-reconstituted membrane extracts. AB - We previously described the [(3)H]cGMP-binding characteristics of a CHAPS solubilized protein that we proposed to be a cGMP transporter. We now report the ATPase activity of the membrane-bound, solubilized and reconstituted form of a cGMP transporter. The membrane-bound protein of unsealed ghosts had a linear ATPase activity over a 120 min incubation period with optimal activity of about 400 pmol/mg/min. The apparent K(m) and V(max) for ATP were about 0.5 mM and 300 pmol/mg/min, respectively. When solubilized with CHAPS the specific activity of the protein was reduced to about 70 pmol/mg/min. Reconstitution of the CHAPS preparation into phospholipid bilayer using rapid detergent removal by Extracti gel column resulted in proteoliposomes which had ATPase activity similar to that found in the erythrocyte membranes. The proteoliposomes displayed a linear ATP dependent uptake of [(3)H]cGMP with an apparent K(m) value of 1. 0 microM. This low K(m)-uptake of [(3)H]cGMP in proteoliposomes was not affected by 10 microM of AMP, cAMP and GMP, but was completely abolished in the presence of the non hydrolyzable ATP analogue, ATP-gamma-S. Some ATPase activation was also observed in the presence of 2 microM cAMP, but it is unclear whether this activity was coupled to the cGMP transporter. Our results show that the membrane protein responsible for cGMP transport has an ATPase activity and transports the cyclic nucleotide in the presence of ATP. PMID- 11118554 TI - Novel cationic amphiphilic 1,4-dihydropyridine derivatives for DNA delivery. AB - In order to find new efficient and safe agents for gene delivery, we have designed and synthesized nine novel single- and double-charged amphiphiles on the base of 1,4-dihydropyridine (1,4-DHP) ring. Some biophysical properties of the amphiphilic dihydropyridines and their complexes with DNA were examined. We investigated the transfer of beta-galactosidase gene into fibroblasts (CV1-P) and retinal pigment epithelial (D 4O7) cell lines in vitro. The structure-property relationships of the compounds were investigated in various ways. The net surface charges of 1,4-DHP liposomes were highly positive (25-49 mV). The double-charged compounds condensed DNA more efficiently than single-charged and the condensation increases with the increasing +/- charge ratio between the carrier and DNA. Double-charged compounds showed also buffering properties at endosomal pH and these compounds were more efficient in transfecting the cells, but transfection efficiency of amphiphiles was cell type-dependent. The length of alkyl chains in double-charged compounds affected the transfection efficacy. The most active amphiphile (compound VI) was double-charged and had two C(12) alkyl chains. At optimal charge ratio (+/- 4), it was 2.5 times more effective than PEI 25 and 10 times better than DOTAP, known efficient polymeric and liposomal transfection agents. Formulation of amphiphiles with DOPE did not change their activities. Our data demonstrate some important effects of amphiphile structure on biophysics and activity. The data also suggest that cationic amphiphilic 1,4-DHP derivatives may find use as DNA delivery system. PMID- 11118556 TI - beta(2)-glycoprotein I-dependent alterations in membrane properties. AB - beta(2)-Glycoprotein I (beta(2)GP1), a 50 kDa serum glycoprotein, binds anionic phospholipids and plays a role in phosphatidylserine (PS)-dependent coagulation and apoptotic processes. To characterize the molecular consequences that occur to target membranes upon binding of beta(2)GP1, the interaction between beta(2)GP1 and PS-containing vesicles was investigated by fluorescent spectroscopy. Membranes containing pyrene-labeled lipid showed that binding of beta(2)GP1 induced a decrease in excimer/monomor ratios (E/M) of the target membrane. Although these membrane alterations occurred in isotonic buffer, the effects were greater in low ionic strength buffer and were coincident to membrane precipitation. In contrast, increases in membrane polarization were only seen in low ionic strength buffer. Analysis of beta(2)GP1 binding kinetics by resonance energy transfer between fluorescein-labeled beta(2)GP1 and rhodamine-containing PS vesicles revealed a two-component process: (1) a primary and rapid binding via the C-terminus that occurred <2 s in both isotonic and low ionic strength buffers, and (2) a sequential binding of the N-terminus that was approximately 100-fold slower in low ionic strength solution. Taken together, these data suggest that beta(2)GP1 alters the fluidity and membrane polarization of its target membrane, which in low ionic strength buffer is of sufficient magnitude to induce precipitation. PMID- 11118557 TI - Discovery of a novel channel-forming protein in the cell wall of the non pathogenic Nocardia corynebacteroides. AB - Detergent extracts of whole cells of the Gram-positive, non-pathogenic, strictly aerobic bacterium Nocardia corynebacteroides contain channel-forming activity. The protein responsible for channel formation was identified using lipid bilayer experiments. It was purified to homogeneity and had an apparent molecular mass of about 134 kDa on SDS-PAGE when it was solubilized at 40 degrees C. When the 134 kDa protein was heated to 100 degrees C for 10 min in sample buffer, it dissociated into subunits with a molecular mass of about 23 kDa and focused at pI of 4.5 during isoelectric focusing. The pure 134 kDa protein was able to increase the specific conductance of artificial lipid bilayer membranes from phosphatidylcholine-phosphatidylserine mixtures by the formation of ion-permeable channels. The channels had an average single-channel conductance of 5.5 nS in 1 M KCl and were found to be cation-selective. Asymmetric addition of the 134 kDa protein to lipid bilayer membranes resulted in an asymmetric voltage-dependence. The analysis of the single-channel conductance as a function of cation radii using the Renkin correction factor and the effect of negative charges on channel conductance suggested that the diameter of the cell wall porin is about 1.0 nm. The channel characteristics of the cell wall channel of N. corynebacteroides were compared with those of other members of the mycolata. They share common features because they are composed of small molecular mass subunits and form large and water-filled channels. PMID- 11118558 TI - Extracellular pH modulates kinetics of the Na(+),K(+)-ATPase. AB - To investigate effects of pH on the Na(+),K(+)-ATPase, we used the Xenopus oocytes to measure transient charge movements in the absence of extracellular K(+), and steady-state currents mediated by the pump as well as ATPase activity. The activity of purified Na(+), K(+)-ATPase strongly depends on pH, which has been attributed to protonation of intracellular sites. The steady-state current reflects pump activity, the transient charge movement voltage-dependent interaction of external Na(+) ions with the pump molecule and/or conformational changes during Na(+)/Na(+) exchange. The steady-state current exhibits a characteristic voltage dependence with maximum at about 0 mV at low external K(+) (< or =2 mM) and with 50 Na(+). This dependency is not significantly affected by changes in external pH in the range from pH 9 to pH 6. Only below pH 6, the voltage dependence of pump current becomes less steep, and may be attributed to a pH-dependent inhibition of the forward pump cycle by external Na(+). External stimulation of the pump by K(+) in the absence of Na(+) can be described by a voltage-dependent K(m) value with an apparent valency z(K). At higher external pH the z(K) value is reduced. The transient current signal in the absence of external K(+) can be described by the sum of three exponentials with voltage dependent time constants of about 50 ms, 700 micros and less than 100 micros during pulses to 0 mV. The charge distribution was calculated by integration of the transient current signals. The slowest component and the associated charge distributions do not significantly depend on external pH changes. The intermediate component of the transients is represented by a voltage-dependent rate constant which shows a minimum at about -120 mV and increases with decreasing pH. Nevertheless, the contribution to the charge movement is not altered by pH changes due to a simultaneous increase of the amplitude of this component. We conclude that reduction of external pH counteracts external K(+) and Na(+) binding. PMID- 11118560 TI - Sertaconazole in the treatment of mycoses: from dermatology to gynecology. AB - Sertaconazole is a pharmaceutical product in the form of a cream, gel, powder and solution for dermatological use and vaginal cream, tablets and ovules for gynecological use. It is marketed in 24 countries and registered in a further 22. The active ingredient is 2% sertaconazole nitrate. Sertaconazole nitrate is an azole antifungal agent, with notable antifungal activity. Its molecule has a highly lipophilic fragment. This is a review of the efficacy and safety of all pharmaceutical forms of sertaconazole in order to provide data on vaginal sertaconazole for marketing purposes. PMID- 11118561 TI - Vaginal candidosis: epidemiological and etiological factors. AB - A review is presented of the emerging problem of candidal colonization regarding epidemiological and etiological factors. In recent years a change in epidemiological trends has been observed. Vaginal candidosis seems to show a higher frequency to recur and a significant increase in infections caused by non albicans species of candidas has been stated. The three stage mechanisms of adhesion, blastopore germination and epithelium invasion are emphasized. There is a balance between candidal organisms and vaginal defense factors (lactobacilli, cellular and humoral immunity) controlling and limiting fungal growth. Vaginitis appears because of an increased number or an enhanced virulence of candidas. In some other patients, a decreased vaginal defense mechanism is a determinant factor. There are still a number of factors involved in clinical candidal vulvovaginitis that need to be clarified. PMID- 11118562 TI - Recurrent vaginal candidiasis changes in etiopathogenical patterns. PMID- 11118563 TI - Sertaconazole: pharmacology of a gynecological antifungal agent. AB - Sertaconazole is a broad spectrum antifungal agent with excellent activity against yeasts, dermatophytes and opportunistic fungi. In addition to this antifungal efficacy, it has a good safety profile, sustained cutaneous retention, and low systemic absorption, all of which make it ideal for topical applications. In this study, the pharmacological properties of sertaconazole related to the treatment of vaginal fungi, in particular vulvovaginal candidiasis, are reviewed. As with all other infectious processes, the interacting components are infectious microorganism, host and drug. The following properties of sertaconazole have been investigated in pre-clinical studies: its in vitro spectrum of activity and potency against causative agents and accompanying factors in vaginal infection; its mechanism of action, whether it acts on the pathogenic properties of the microorganism; if it affects host defense mechanisms and how its antifungal activity is manifested in vivo in experimental candidiasis in the mouse. PMID- 11118565 TI - In this issue ellipsis PMID- 11118564 TI - Topical treatment of vaginal candidosis with sertaconazole and econazole sustained-release suppositories. AB - OBJECTIVE: To compare the efficacy and safety of sertaconazole and econazole sustained-release suppositories in the treatment of vulvovaginal candidosis. STUDY DESIGN: 369 women with symptoms and signs of vulvovaginitis were enrolled in this multicenter, randomized, double-blind study. After clinical examination and vaginal sampling, 183 women were treated with a 300-mg sertaconazole suppository and the other 186 with a 150-mg econazole suppository. They were evaluated 1 week after treatment and those who were clinically uncured received a second suppository and were re-assessed 1 week later. All women were evaluated 1 month after the last administration. At each follow-up visit, clinical efficacy was assessed and a vaginal sampling was performed for microscopic examination and culture. RESULTS: Of the 369 women included, only the 310 who had a positive culture for a strain of Candida were taken into account for efficacy analysis: 150 in the sertaconazole group and 160 in the econazole group. One hundred and five women (49 in the sertaconazole group and 56 in the econazole group) were not clinically cured after 1 week and received a second suppository. There were no differences between the two groups for the rates of clinical recovery (disappearance of signs and symptoms) and mycological recovery (negative culture), 1 week after the first application (62.1 and 67.7%, respectively), 1 week after the second application for women treated twice (72.3 and 80.6%, respectively) and for all patients 1 month after the last application (65.3 and 62.0%, respectively). Among the patients cured 1 week after the last application, the mycological recurrence rate after 1 month was significantly higher in the econazole group (32.7 vs. 19.8%, P=0.035). There was a trend towards better efficacy of sertaconazole after the first application, whereas the two treatments had similar efficacy in women treated twice. There were no serious adverse events and only local irritation was reported, without any statistically significant difference between the two groups. CONCLUSION: Single topical administration of sertaconazole and econazole had similar efficacy and safety but the former is associated with a lower rate of mycological recurrence one month after achieving a negative culture. PMID- 11118566 TI - Differential treatment in acute upper cervical spine injuries: a critical review of a single-institution series. AB - BACKGROUND: A single-institution series of injuries of the upper cervical spine are analyzed retrospectively and the literature relevant to the topic is reviewed. METHODS: Seventy patients (34 female, 36 male, mean age 47 years) were admitted during a 5-year period for injuries of the upper cervical spine. Sixty five were followed for a mean time of 18 months. Three isolated ligamentous instabilities, 6 isolated C1 fractures, 3 complex C2 fractures, 10 combined C1/C2, and 48 C2 fractures (17 hangman's, 31 odontoid) were diagnosed. Twenty nine patients were treated conservatively and for 41 patients surgery was the primary treatment. Twenty-three ventral odontoid screw fixations, 8 ventral platings and 10 dorsal stabilizations were performed. Stability was evaluated using flexion-extension radiography. Pain levels and neurological outcome were assessed. RESULTS: Operative mortality and neurological morbidity were 0%. Two wound infections and 3 instabilities (17%) in odontoid Type II fractures primarily treated with ventral odontoid screw fixation needed dorsal restabilization. During follow-up examinations the neurological status of three patients was improved. In 62 patients preoperative status was attained. Six patients evaluated their pain as severe, two as disabling. CONCLUSIONS: Candidates for surgery as the primary treatment include those with isolated ligamentous instabilities, Type III hangman's fractures and Type II odontoid fractures with dislocation more than 5 mm. In combined C1/C2 fractures the axis fracture dictates the treatment strategy. Patients who undergo dorsal procedures and have involvement of C1 have a greater chance of developing persistent pain. PMID- 11118567 TI - Morphological changes of cerebral penetrating arteries in a canine double hemorrhage model. AB - BACKGROUND: Morphological presentations of cerebral vasospasm, such as dystrophy and desquamation of endothelial cells, corrugation of the internal elastic layer, and necrotic changes in smooth muscle cells, are well defined in large cerebral arteries. This study was undertaken to examine pathological changes in cerebral penetrating arteries in a canine double hemorrhage model. METHODS: Eighteen mongrel dogs were subjected to an autologous arterial blood (0.4 mL/kg) injection into the cisterna magna on day 0 and day 2 after withdrawal of an equivalent amount of cerebrospinal fluid. Angiogram was performed on day 0 before the blood injection and on the day the dogs were sacrificed. The dogs were divided into four groups: control (day 0) (n = 4), hemorrhage and sacrificed on day 3 (n = 4), day 5 (n = 5), and day 7 (n = 5). The penetrating arteries were removed and found to be spastic on days 3, 5, and 7, but not in the control group. RESULTS: Endothelial dystrophy and partial desquamation were recorded in all dogs sacrificed on days 5 and 7. Condensation of chromatin, blebbing of the membrane, and condensation of cytoplasm were identified in many endothelial cells, features that are consistent with apoptosis. The morphological changes in the penetrating arteries were more pronounced on days 5 and 7. CONCLUSIONS: Vasospasm occurred in cerebral penetrating arteries in a canine double hemorrhage model. The morphological change in penetrating arteries, especially apoptosis in endothelial cells, is consistent with an early phase of vasospasm. Vasospasm in a penetrating artery may contribute to the cerebral ischemia that occurs during vasospasm. PMID- 11118569 TI - Prognostic significance of clinical and angiogenesis-related factors in low-grade oligodendrogliomas. AB - BACKGROUND: Keeping in mind that oligodendrogliomas have unpredictable biological behavior, the aim of this study is to investigate the prognostic significance of VEGF expression and microvessel density in a homogeneous series of low-grade oligodendrogliomas. METHODS: For this study 36 patients with a low-grade oligodendroglioma treated by surgical resection and radiotherapy were selected. At the time of surgery, in all cases the Karnofsky Performance Scale (KPS) score was more than 70, and the study of the resected tumor disclosed a Ki-67/MIB-1 labeling index (MBI-1 LI) less than 1%. In this homogeneous series, immunohistochemical studies were performed using monoclonal antibodies against VEGF in order to study the expression of this cytokine, and against vascular endothelial CD-34 antigen, in order to identify microvessels. RESULTS: Our results show that in contrast to low-grade astrocytomas, low-grade oligodendrogliomas lacked immunoreactive VEGF. Oligodendrogliomas with low vascular density (less than 20 microvessels per microscopical field, at 200 x) or high vascular density (more than 100 microvessels per field, at 200 x) were identified, but this factor had no influence on the survival rate of patients. On the other hand, analysis of the present series showed that clinical factors, such as age or extent of surgical resection, were not significantly associated with survival. CONCLUSIONS: In contrast to low-grade astrocytomas, the angiogenesis score of low-grade oligodendrogliomas (counting the number of microvessels in tumor tissue) adds little information to help predict tumor behavior. PMID- 11118568 TI - Prevention of vasospasm in penetrating arteries with MAPK inhibitors in dog double-hemorrhage model. AB - BACKGROUND: Vasospasm in the penetrating arteries contributes to ischemic neurological deficit. It may be as important as angiographic vasospasm because it would explain the discrepancies between angiographic vasospasm and clinical symptoms in some patients. It may also underlie the different effects of vasodilators. The present study examined this hypothesis by looking at the effect of the inhibitors of mitogen-activated protein kinase (MAPK) on vasospasm of the penetrating arteries. METHODS: Twenty-two adult mongrel dogs of either sex were used for the dog double-hemorrhage model. The dogs were randomly divided into four groups: control-hemorrhage, vehicle-treated, PD98059-treated, and U0126 treated groups. The drug injections were started on Day 3 after the first subarachnoid hemorrhage (SAH). The clinical status of the dogs was studied, based on their activity, appetite, and focal neurological symptoms. On Day 7, all the dogs were sacrificed, and the penetrating arteries from the brain stem were prepared for transmission electron microscopy. RESULTS: (1) Severe vasospasm developed in the basilar arteries in the SAH-without-treatment group (control), in the DMSO-treated group (DMSO), and in the U0126 treatment group with mean reduction of the basilar artery diameter of 46.57%, 49.3%, and 39.6%, respectively. In the PD98059-treatment group only a mild vasospasm was observed and the mean reduction of the basilar artery diameter was 18.9%. (2) All the dogs in the control SAH and vehicle-treated groups developed severe angiographic and clinical vasospasm. The penetrating arteries were contracted, and the endothelial and smooth muscle cells were dystrophic. (3) The dogs in the U0126-treated group developed severe angiographic, but not clinical, vasospasm. The penetrating arteries were not contracted, and the endothelial and smooth muscle cells were not dystrophic. (4) The dogs in the PD98059 group developed mild angiographic vasospasm. No dog developed clinical symptoms that could be attributed to vasospasm. In morphological studies, the penetrating arteries were slightly contracted, but the cells were not dystrophic. CONCLUSIONS: Vasospasm of the penetrating arteries, but not angiographic vasospasm, is consistent with the clinical symptoms and signs of vasospasm. MAPK may be important in maintaining vasospasm of both major and penetrating cerebral arteries. The correlation of the improvement in the clinical score with the reduction of vasospasm in the penetrating arteries demonstrated an important role of penetrating arteries in the morbidity and mortality caused by SAH. PMID- 11118570 TI - Expression of tenascin-C in astrocytic tumors: its relevance to proliferation and angiogenesis. AB - BACKGROUND: The expression and distribution of the extracellular matrix protein tenascin-C (TN-C) may be enhanced in human astrocytomas. The purpose of this study is to evaluate the expression of TN-C according to histological malignancy of tumor cells and its relevance to neoplastic angiogenesis in human astrocytic tumors. METHODS: Between 1994 and 1998, 52 astrocytic tumor specimens including 4 pilocytic astrocytomas, 13 astrocytomas, 3 anaplastic astrocytomas, and 32 glioblastomas were used in this study. A retrospective analysis was performed to evaluate a statistical correlation between TN-C expression and proliferative indices. We characterized the expression of TN-C in neoplastic vessels, around individual tumor cells as a tumor network, and in tumor cells by immunohistochemistry using antibodies against human TN-C. The proliferative indices were also investigated by immunostaining with the MIB-1 antibody against the Ki-67 proliferation antigen. RESULTS: TN-C immunoreactivity was found to be enhanced in tumor vessels and tumor networks of high-grade astrocytic tumors. The vascular TN-C deposition was greater in high-grade than in low-grade astrocytic tumors (p < 0.05). Its expression was the most intense in glioblastomas. Proliferation indices increased with tumor grade and MIB-1 labeling index (LI) was highest in glioblastomas. Moreover, expression of TN-C in tumor vessels was correlated with proliferative indices. CONCLUSIONS: Our data show that TN-C in human astrocytic tumors may be identified as a factor contributing to malignant progression. And also, enhanced expression of TN-C in tumor vessels having a high proliferative index indicates that TN-C could be involved in neoplastic angiogenesis. PMID- 11118571 TI - Proliferative activity in craniopharyngiomas: clinicopathological correlations in adults and children. AB - BACKGROUND: Craniopharyngiomas are slow-growing, locally invasive intracranial tumors that can generate considerable morbidity, and recurrences are often difficult to manage. Because reliable morphologic criteria for accurately predicting the clinical outcome of these tumors are lacking, we evaluated the growth potential of craniopharyngiomas by measuring their proliferative activity based on MIB-1 immunostaining for the Ki-67 antigen, which is expressed during all phases of the cell cycle except G(0.) METHODS: Paraffin sections from 37 cases of craniopharyngiomas were immunostained with the monoclonal antibody MIB 1, and a labeling index was derived in each case from an the with the highest labeling. RESULTS: MIB-1 immunoreactivity was mainly confined to the peripheral palisaded epithelium of craniopharyngiomas. In adult craniopharyngiomas, MIB-1 labeling indices (MIB-LI) varied from 0.1% to 34.6% (mean 8.9%; SD 9. 8), and in pediatric tumors the indices ranged from 1.8% to 15.0% (mean 6.3%; SD 3.7). MIB LI was not found to be an independent predictor of recurrence, although in all the pediatric cases that recurred, MIB-LI in the second specimen was greater. CONCLUSIONS: The actively proliferating compartment in craniopharyngiomas seems to be the peripheral palisaded epithelium. Low MIB-LI observed in the majority of tumors is in concordance with the slow growth and low-grade invasiveness associated with craniopharyngiomas. However, unlike other intracranial neoplasms, where Ki-67 labeling indices have been useful in predicting tumor behavior, a clear relationship could not be demonstrated between MIB-1 immunoreactivity, morphological features and clinical outcomes in adults or children with craniopharyngiomas. PMID- 11118572 TI - Spontaneous intraventricular rupture of craniopharyngioma cyst. AB - BACKGROUND: Rupture of a cystic craniopharyngioma is a rare phenomenon. The rupture of the cyst causes decompression of the adjacent neural structures resulting in spontaneous improvement of the visual symptoms or level of sensorium. The leakage of its contents into the subarachnoid space gives rise to meningismus. We report an extremely rare phenomenon of an intraventricular rupture of a cystic craniopharyngioma, which resulted in acute neurological deterioration and chemical ventriculitis. CASE DESCRIPTION: A 38-year-old lady presented with a 1-year history of frontal lobe dysfunction and bilateral primary optic atrophy. The CT scan showed a multi-loculated, hyperdense lesion in the region of the third ventricle and suprasellar cistern. She suffered acute deterioration of neurological status; computed tomography (CT) scan showed a hypodense lesion in the suprasellar cistern with persistent hydrocephalus. She was treated with ventricular drainage, steroids and anticonvulsants. Ventricular fluid showed high cholesterol and LDH levels. The diagnosis of craniopharyngioma was subsequently verified histologically. CONCLUSIONS The intraventricular rupture of a cystic craniopharyngioma can result in acute clinical deterioration and morbidity because of chemical ventriculitis. This is unlike the rupture in the subarachnoid space or sphenoid sinus which usually results in symptomatic improvement, although chemical meningitis may occur. This rare phenomenon should be recognized, and prompt ventricular drainage is advised. The literature is reviewed, and management of this condition is discussed. PMID- 11118574 TI - Factor XIII deficiency and postoperative hemorrhage after neurosurgical procedures. AB - BACKGROUND: Factor XIII is of physiological importance for hemostasis, especially in patients undergoing surgery. It catalyzes the enzymatic cross-linking of fibrin monomers into stable polymers and protects polymers from plasmatic and nonspecific degradation. Postoperative hemorrhage in patients with congenital and acquired Factor XIII deficiencies has been described in various surgical fields. However, there are no data about the incidence and clinical relevance of decreased Factor XIII after neurosurgical procedures. The objective of our study was to investigate the association between Factor XIII deficiency and postoperative hemorrhage after intracranial surgery. METHODS: A total of 1264 patients who underwent intracranial operations were reviewed retrospectively. Standard coagulation parameters were monitored during the perioperative course in all patients. Factor XIII testing was performed postoperatively in 34 patients in whom coagulopathies were suspected despite normal platelets, fibrinogen, prothrombin, and partial thromboplastin time. Data were analyzed to evaluate the association of Factor XIII deficiency and major postoperative hemorrhage. RESULTS: In this series of 1264 patients, a total of 20 patients (1. 6%) suffered from a major postoperative hemorrhage. Of the 34 patients with suspected coagulopathies and postoperative Factor XIII testing, 11 had a major postoperative hemorrhage. Normal levels of Factor XIII, defined as more than 60%, were found in 26 of the 34 patients. Factor XIII deficiency, defined as less than 60%, was found in eight patients. All patients with Factor XIII deficiency (n = 8) had a major postoperative hemorrhage. Of the remaining 26 patients with normal Factor XIII levels only three had a postoperative hemorrhage (p < 0.00001, Fisher's exact test). CONCLUSIONS: Decreased Factor XIII activity may be associated with an increased risk of postoperative hemorrhage after intracranial surgery. PMID- 11118573 TI - Spontaneous peritumoral haemorrhage associated with sinus confluence meningioma: case report. AB - BACKGROUND: Torcular or sinus confluence meningioma is rare and surgically formidable. This reported sinus confluence meningioma was associated with peritumoral intracerebral hemorrhage. The surgical strategy and the mechanism of peritumoral hemorrhage are discussed. CASE DESCRIPTION: A 42-year-old woman presented with a history of headache, vomiting, and cerebellar dysfunction for 2 months. Plain computed tomography (CT) scan and magnetic resonance imaging (MRI) demonstrated a high-density mass in the torcular region involving both lateral sinuses. MR angiography demonstrated complete occlusion of the left lateral sinus and straight sinus and stenosis of the right lateral sinus. Two years after her first operation she experienced sudden headache and left upper quadrant hemianopsia. Plain CT scan and MRI showed a hyperintense tumor in the torcular region with an intracerebral hematoma in the right occipital lobe. An angiogram demonstrated occlusion of the caudal part of the superior sagittal sinus, bilateral transverse sinuses, and straight sinus. Gross total removal of the tumor was done along with the left lateral sinus through a suboccipital and a supratentorial occipital craniotomy in the first operation. The patient underwent total resection of the tumor at second operation through a bilateral occipital and suboccipital craniotomy along with resection of the dura including the confluence, the caudal part of the superior sagittal sinus, the right lateral sinus, and the straight sinus. The postoperative course was uneventful and postoperative MRI showed total removal of the tumor. CONCLUSION: Sinus confluence meningioma may present with peritumoral hemorrhage. Radical removal may be possible when the sinus confluence is completely occluded and there is good collateral drainage. PMID- 11118575 TI - Spontaneous movement of a metallic intracranial foreign body: case report. PMID- 11118576 TI - Parietal sinus pericranii: case report and technical note. AB - BACKGROUND: Sinus pericranii is a rare vascular anomaly that is defined as a group of abnormal communications between the extracranial and intracranial venous systems, usually involving the superior sagittal sinus. Different surgical techniques have been used to manage this anomaly. Surgical technique and radiologic findings are discussed. CLINICAL PRESENTATION: A case of parietal sinus pericranii is presented that was developed spontaneously. This 33-year-old woman presented with a soft fluctuant mass in the right parietal region adjacent to the midline. She complained of headache and dizziness. The preoperative radiologic findings of simple skull X-ray, computed tomography, bone scan, and direct venogram are presented. The lesion was removed completely and then multiple small vascular channels through the underlying skull were obliterated by air-powered diamond drilling. The presence of vascular endothelium in the pathologic specimen suggested a congenital origin. CONCLUSION: We think this method is very easy and useful for managing the multiple small fenestrations of the sinus pericranii without recurrence. PMID- 11118578 TI - Introduction. PMID- 11118577 TI - A response to Dr. Dunsker, President of the AANS. PMID- 11118579 TI - Rare sugars and sugar-based synthons by chemo-enzymatic synthesis. AB - The unique catalytic potential of the fungal enzyme pyranose oxidase was demonstrated by preparative conversions of a variety of carbohydrates, and by extensive chemical characterization of the reaction products with NMR spectroscopy. The studies revealed that POx not only oxidizes most substrates very efficiently but also that POx possesses a glycosyl-transfer potential, producing disaccharides from beta-glycosides of higher alcohols. Although most substrates are oxidized by POx at the C-2 position, several substrates are converted into the 3-keto-derivatives. On the basis of these products, strategies are developed for the convenient production of sugar-derived synthons, rare sugars and fine chemicals by combining biotechnical and chemical methods. PMID- 11118580 TI - Dynamics of proteolysis and its influence on the accumulation of intracellular recombinant proteins. AB - A method to quantify the impact of proteolysis on accumulation of recombinant proteins in E. coli is described. A much smaller intracellular concentration of staphylococcal protein A (SpA) (14.7 mg. g(-1)) compared to the fusion protein SpA-betagalactosidase (138 mg. g(-1)) is explained by a very high proteolysis rate constant of SpA. The SpA synthesis rate reached a maximum one hour after induction and gradually decreased to half of this value at the end of the cultivation. The decrease of the synthesis rate and the 1st order kinetics of proteolysis lead to an equilibrium between synthesis and degradation of SpA from 2 h after induction. This resulted in no further SpA accumulation in cells, though synthesis continued for at least 10 h. Similar experiments with recombinant protein ZZT2 also revealed that most of the synthesized product was degraded. The order of proteolysis kinetics depended on the concentration of the recombinant protein: at low concentrations both SpA and ZZT2 were degraded according to first order kinetics, while at high concentrations ZZT2 was degraded according to zero order kinetics. In a protease Clp mutant the degradation rate decreased and intracellular concentration of ZZT2 increased from 50 mg. g(-1) to 120 mg. g(-1). The measurements of proteolysis rate throughout the cultivation enabled calculation of a hypothetical accumulation of the product assuming complete stabilization. In this case the concentration would have increased from 50 to 280 mg. g(-1) in 11 h. Thus, this method reveals the potential to increase the productivity by eliminating proteolysis. PMID- 11118581 TI - Surfactin and iturin A effects on Bacillus subtilis surface hydrophobicity. AB - The synthesis of extracellular molecules such as biosurfactants should have major consequences on bacterial adhesion. These molecules may be adsorbed on surfaces and modify their hydrophobicities. Certain strains of Bacillus subtilis synthesize the lipopeptides, which exhibit antibiotic and surface active properties. In this study the high-performance liquid chromatography (HPLC) analysis of the culture supernatants of the seven B. subtilis strains, showed that the lipopeptide profile varied greatly according to the strain. Among the three lipopeptide types, only iturin A was produced by all B. subtilis strains. Bacterial hydrophobicity, evaluated by the water contact angle measurements and the hydrophobic interaction chromatography, varied according to the strain. Two strains (ATCC 15476 and ATCC 15811) showing extreme behaviors in term of hydrophobicity were selected to study surfactin and iturin A effects on bacterial hydrophobicity. The two lipopeptides modified the B. subtilis surface hydrophobicity. Their effects varied according to the bacterial surface hydrophobic character, the lipopeptide type and the concentration. Lipopeptide adsorption increased the hydrophobicity of the hydrophilic strain but decreased that of the hydrophobic. Comparison of lipopeptide effects on B. subtilis surface hydrophobicity showed that surfactin was more effective than iturin A for the two strains tested. PMID- 11118582 TI - Construction and expression of antibody targeted plasminogen activator* AB - It has been known that antibody-mediated plasminogen activator will be much more specific than its parent molecular. To get a cheaper and more effective medicine for thrombolytic therapy, we used SZ51, a GMP140 specific monoclonal antibody, and a truncated single-chain urokinase to construct a novel targeted plasminogen activator. PCR was used to amplify the region of VL and VH chains from Fab of SZ51, GMP140 specific monoclonal antibody, and scu-PA-32KD(leu144-leu411) from urokinase gene, respectively. Through suitable linker and appropriate restriction sites, these fragments were joined together and inserted into the expression vector, pET-5a, via NdeI site. The recombinant protein was expressed in BL21 (DE3) plyS, a kind of E. coli. It was shown in Western-blotting and ELISA that the protein could interact with the multiple cloned antibody of urokinase. After partial purification: dialysis, Sephadex G-100, dialysis and Phenyl-Sepharose fast flow, the product had a strong fibrinolytic activity through activating plasminogen on fibrin plate. The specific activity was about 47,000 IU/mg, corresponding to 80,000 IU/mg for the part of rscu-PA-32k, and the activity could be inhibited specifically by urokinase specific antibody. Activation of plasminogen by the chimera followed Michaelis-Menten kinetics, and the K(m) was 1.08 uM. PMID- 11118583 TI - Secretion of biologically active murine interleukin-10 by Lactococcus lactis. AB - We investigated the ability of Lactococcus lactis to secrete biologically active, murine interleukin-10 (mIL-10). mIL-10 was synthesized as a fusion protein, consisting of the mature part of the eukaryotic protein fused to the secretion signal of the lactococcal Usp45 protein. The secreted protein was analyzed by PAGE, ELISA and bioassay.We show that L. lactis can efficiently secrete biologically active, murine IL-10. Determination of the N-terminal amino acid sequence confirmed correct processing of the fusion polypeptide by the lactococcal signal peptidase. The amount of mIL-10, accumulating in the medium, could be increased by a factor of ten by growing the cells in an optimized medium, buffered at near-neutral pH. Under these conditions, up to 30 mg of mIL 10 was obtained from a 10-litre fermentation. PMID- 11118584 TI - Genetic strategies to enhance penicillin acylase production in Escherichia coli. AB - We demonstrated the improvement of penicillin acylase (PAC) production by optimization of the host/vector system using genetic engineering strategies. Several expression plasmids with improved efficiency for the transcription of the pac gene and/or translation of the pac mRNA were constructed. Mutant strains, isolated by a novel screening method, were effective for use as the expression host to produce PAC. The feasibility of using the mutant strains harboring a selection of expression plasmids for the production of PAC was evaluated. The effect of the mutation(s) resulting in the improved PAC producing ability was characterized. While the production of PAC was significantly enhanced using the optimized host/vector system, the formation of PAC inclusion bodies was shown to be another step limiting the production of recombinant PAC. PMID- 11118585 TI - Production of poly(3-hydroxybutyrate) from inexpensive substrates. AB - Two inexpensive substrates, starch and whey were used to produce poly(3 hydroxybutyrate) (PHB) in fed-batch cultures of Azotobacter chroococcum and recombinant Escherichia coli, respectively. Oxygen limitation increased PHB contents in both fermentations. In fed-batch culture of A. chroococcum, cell concentration of 54 g l(-1) with 46% PHB was obtained with oxygen limitation, whereas 71 g l(-1) of cell with 20% PHB was obtained without oxygen limitation. The timing of PHB biosynthesis in recombinant E. coli was controlled using the agitation speed of a stirred tank fermentor. A PHB content of 80% could be obtained with oxygen limitation by increasing the agitation speed up to only 500 rpm. PMID- 11118586 TI - Performance of a miniaturized bioreactor in space flight: microtechnology at the service of space biology. AB - We describe here the performance and the use of microtechnology in a miniaturized bioreactor developed for the continuous cultivation of yeast cells, Saccharomyces cerevisiae, in microgravity. This bioreactor has been used on two Shuttle missions, where its functionality was successfully demonstrated. In the future, bioreactors will become a key element for long-term experiments, and would also be applied in the cultivation of mammalian cells or tissues for medical applications. PMID- 11118587 TI - Data and knowledge based experimental design for fermentation process optimization. AB - A novel method for the sequential experimental design in order to optimize fed batch fermentations was applied to a hyaluronidase fermentation by Streptococcus agalactiae. A Lambda-optimal design was introduced to minimize the model parameter estimation error and to maximize the performance of the fermentation process. The method employs hybrid models that contain mechanistic, fuzzy and neural network components. PMID- 11118588 TI - Superoxide dismutase biosynthesis by two thermophilic bacteria. AB - Some high-molecular weight antioxidant defense system components of two thermophilic bacteria isolated from spa waters of Serbia (Yugoslavia) and identified as Bacillus stearothermophilus and Thermothrix sp. were studied. In addition to superoxide dismutase (SOD; EC 1.15.1.1), qualitative analyses demonstrated the presence of catalase (EC 1.11.1.6), peroxidases and oxidases in both bacterial strains. Cell-free extracts were subjected to nondenaturing polyacrylamide gel electrophoresis (PAGE) and SOD activity in the eluates of the corresponding bands was examined in the presence of several specific inhibitors. A slight decrease of SOD activity observed in the presence of 0.3 M potassium cyanide and its complete insensitivity to hydrogen peroxide (5 mM) and sodium azide (20 mM) action suggest that the enzyme occurring in the two thermophiles represents Mn SOD. A high SOD activity recorded in cell-free extracts strongly recommends these two bacterial strains as potential producers of this important antioxidant defense system component at industrial scale. PMID- 11118589 TI - Metabolic flux analyses for serine alkaline protease production. AB - The intracellular metabolic fluxes through the central carbon pathways in Bacillus licheniformis in serine alkaline protease (SAP) production were calculated to predict the potential strategies for increasing the performance of the bacilli, by using two optimization approaches, i.e. the theoretical data based (TDA) and the theoretical data-based capacity (TDC) analyses based on respectively minimum in-vivo SAP accumulation rate and maximum SAP synthesis rate assumptions, at low-, medium-, and high-oxygen transfer conditions. At all periods of low-oxygen transfer condition, in lag and early exponential periods of medium-oxygen transfer (MOT) condition, and SAP synthesis period of high-oxygen transfer (HOT) condition, the TDA and TDC analyses revealed that SAP overproduction capacity is almost equal to the observed SAP production due to the regulation effect of the oxygen transfer. In the growth and early SAP synthesis period and in SAP synthesis period at MOT condition the calculated results of the two analyses reveal that SAP synthesis rate of the microorganism can be increased 7.2 and 16.7 folds, respectively; whereas, in the growth and early SAP synthesis period at HOT condition it can be increased 12.6 folds. The diversions in the biochemical reaction network and the influence of the oxygen transfer on the performance of the bacilli were also presented. The results encourage the application of metabolic engineering for lifting the rate limitations and for improving the genetic regulations in order to increase the SAP production. PMID- 11118590 TI - Preparation of high activity whole cell biocatalyst by permeabilization of recombinant flocculent yeast with alcohol. AB - Flocculent yeast Saccharomyces cerevisiae YF234 (MATa ura3-52 trp1Delta2 his ade 2-1 can1-100 sta1 FLO8) cells overexpressing glyoxalase I and having strong flocculation ability were permeabilized with isopropyl alcohol and ethanol under various conditions. The treatment with 40% isopropyl alcohol significantly improves the initial reaction rates of recombinant flocculent yeast cells. Moreover, the reactivity of permeabilized flocculent yeast cells was similar to that of dispersed cells with EDTA. On the other hand, the flocculation ability of yeast cells was not affected by the treatment with alcohol solutions of various concentrations and treatment time length. Therefore, the recombinant flocculent yeast cells permeabilized with alcohol are very effective whole cell biocatalysts. PMID- 11118591 TI - Halorespiring bacteria-molecular characterization and detection. AB - Recently, a rapidly increasing number of bacteria has been isolated that is able to couple the reductive dehalogenation of various halogenated aromatic and aliphatic compounds like chlorophenols and tetrachloroethene to energy conservation by electron-transport-coupled phosphorylation. The potential of these halorespiring bacteria for innovative clean-up strategies of polluted anoxic environments has greatly stimulated efforts to unravel the molecular basis of the novel respiratory chains they possess. The thorough characterization of halorespiratory key components at the physiological, biochemical and molecular genetic level has revealed both structural and functional similarity of chloroaryl- and chloroalkyl-respiratory chains from different phylogenetically distinct microorganisms. The reductive dehalogenases from halorespiring bacteria were found to comprise a novel class of corrinoid-containing Fe/S-proteins. Sensitive molecular methods for monitoring both presence and fate of halorespiring bacteria have been developed, which will be instrumental for the design and maintenance of optimised in situ bioremediation processes. PMID- 11118592 TI - A model of interfacial inactivation for papain in aqueous organic biphasic systems. AB - A model was proposed to describe the effects of the main factors in aqueous organic two-liquid-phase media on the stability of papain. The relationships between the half-life of papain activity and these factors including interfacial tension, stirring rate, phase volume ratio and temperature were investigated. The results showed that these factors had notable effects on papain stability except temperature. The correlation coefficient between the model and the experimental data were 0.829, which indicated the model is practicable. PMID- 11118593 TI - Capacity of thermomonospora alba XylA to impart thermostability in family F/10 chimeric xylanases. AB - To reveal structure-function relationships of family F/10 glycanases, an in vitro molecular level shuffling experiment was conducted to accumulate useful amino acid residues from two homologous F/10 xylanases, FXYN of Streptomyces olivaceoviridis E-86 and XylA of Thermomonospora alba ULJB1, into a single chimeric xylanase. The parent genes were shuffled by crossovers at selected module borders using self-priming Polymerase Chain Reaction (PCR)s. The shuffled constructs, designated as FXYN-M3/4-XylA, FXYN-M9/10-XylA, and FXYN-M14/15-XylA were cloned and their nucleotide sequences were confirmed. Two chimera, FXYN-M3/4 XylA and FXYN-M14/15-XylA, demonstrated activity against RBB-xylan and were over expressed as His-tag fusion proteins under control of T5 promoter of pQE60. The homogeneously pure chimeric proteins, FXYN-M3/4-XylA and FXYN-M14/15-XylA showed improved thermal and pH profiles compared to those of one of the parents, FXYN. This was apparently due to the influence of amino acids inherited from thermophilic XylA. Measured K(m) and kcat values were closer to those of the other parent, XylA. Interestingly, a significant level of heat tolerance up to 60 degrees C, was recorded for FXYN-M3/4-XylA in comparison to only 40 degrees C for FXYN-M14/15-XylA though their temperature optima did not correlates with their thermal stability. These results indicated that the amino acid residues of the larger T. alba XylA DNA fragment present in FXYN-M3/4-XylA were responsible for inducing its thermal stability. PMID- 11118594 TI - Expression of E. coli araBAD operon encoding enzymes for metabolizing L-arabinose in Saccharomyces cerevisiae. AB - The Escherichia coli araBAD operon consists of three genes encoding three enzymes that convert L-arabinose to D-xylulose-5 phosphate. In this paper we report that the genes of the E. coli araBAD operon have been expressed in Saccharomyces cerevisiae using strong promoters from genes encoding S. cerevisiae glycolytic enzymes (pyruvate kinase, phosphoglucose isomerase, and phosphoglycerol kinase). The expression of these cloned genes in yeast was demonstrated by the presence of the active enzymes encoded by these cloned genes and by the presence of the corresponding mRNAs in the new host. The level of expression of L-ribulokinase (araB) and L-ribulose-5-phosphate 4-epimerase (araD) in S. cerevisiae was relatively high, with greater than 70% of the activity of the enzymes in wild type E. coli. On the other hand, the expression of L-arabinose isomerase (araA) reached only 10% of the activity of the same enzyme in wild type E. coli. Nevertheless, S. cerevisiae, bearing the cloned L-arabinose isomerase gene, converted L-arabinose to detectable levels of L-ribulose during fermentation. However, S. cerevisiae bearing all three genes (araA, araB, and araD) was not able to produce detectable amount of ethanol from L-arabinose. We speculate that factors such as pH, temperature, and competitive inhibition could reduce the activity of these enzymes to a lower level during fermentation compared to their activity measured in vitro. Thus, the ethanol produced from L-arabinose by recombinant yeast containing the expressed BAD genes is most likely totally consumed by the cell to maintain viability. PMID- 11118595 TI - Display of green fluorescent protein on Escherichia coli cell surface. AB - In this study, expression of green fluorescence protein (GFP) on the external surface of Escherichia coli was achieved by construction of a fusion protein between Lpp-OmpA hybrid and a GFP variant, GFPmut2. The GFP was fused in frame to the carboxyl-terminus of Lpp-OmpA fusion previously shown to direct various other heterologous proteins to E. coli cell surface. Western blot analysis of membrane fractions identified the Lpp-OmpA-GFP fusion protein with the expected size (43 kDa). Immunofluorescence microscopy, immunoelectron microscopy, protease and extracellular pH sensitivity assays further confirmed that GFP is anchored on the outer membrane. The GFP displayed on the E. coli outer surface retained its fluorescence and was not susceptible to the indigenous outer membrane protease OmpT even though there are two putative OmpT proteolytic sites present in GFP. Optimization of the expression conditions was conducted using fluorometry, eliminating cumbersome immuno-labeling procedures. Surface-displayed GFP could be used in a variety of applications including screening of polypeptide libraries, development of live vaccines, construction of whole cell allosteric biosensors, and signal transduction studies. PMID- 11118596 TI - Controlled malolactic fermentation in cider using Oenococcus oeni immobilized in alginate beads and comparison with free cell fermentation. AB - Cells of Oenococcus oeni (formerly Leuconostoc oenos) immobilized in alginate beads were used as starter culture to conduct malolactic fermentation in cider production. Concentrations of major organic acids and volatile compounds were monitored during the process, and results were compared to those obtained when using free cells in the same conditions. The rates of malic acid consumption were similar but lower ethanoic acid content and higher concentration of alcohols were detected with immobilized cells. These features have beneficial effects on the organoleptic properties of cider. A comparison between the kinetic behavior in immobilized and free cells, based on the data obtained for the malic acid consumption, has been developed solving the homogeneous diffusion model when it is applied to the system with immobilized cells. PMID- 11118597 TI - Candida Rugosa lipase reactions in nonionic w/o-microemulsion with a technical surfactant. AB - In enzyme catalysis there is a great interest in finding suitable organic media for less water soluble substrates in order to increase the substrate concentration and therefore the reaction rates. These requirements are fulfilled by using w/o-microemulsions as reaction media. The influences of pH, temperature, water concentration and the kinetic parameters of Candida Rugosa Lipase in a nonionic w/o-microemulsion with a surfactant of technical grade, Marlipal O13-60, are presented. In an example the enantiospecific esterification of racemic menthol with propionic anhydride using this nonionic microemulsion likely to be affordable in large scale applications is shown. For a continuous process an ultrafiltration unit is attached to a reactor within a loop. In this way, the reverse micelles containing the enzymes can be separated from the oil, containing the product, and reused afterwards. PMID- 11118598 TI - Biodesulphurization of coal: effect of pulse feeding and leachate recycle. AB - Biodesulphurization of coal was carried out under four modes of operation namely: conventional batch, constant volume pulse feeding (CVPF), increasing volume pulse feeding (IVPF) and leachate recycle. The effects of different pulse feeding strategies and leachate (product) recycle on biological performance were studied and compared with a conventional batch process. The sulphur removal rates for each of the four processes were 0.04 g/day (batch), 0.09 g/day (CVPF), 0.19 g/day (IVPF) and 0.05 g/day (leachate recycle). The values of iron solubilization rate (batch-83 ug/ml/day; CVPF-136 ug/ml/day; IVPF-198 ug/ml/day; leachate recycle-133 ug/ml/day) also followed the same trend. The percentage sulphur removal on the 30th day using batch, CVPF, IVPF and leachate recycle processes was 72%, 93%, 97% and 90%, respectively. IVPF was found to be the best operational strategy for biodesulphurization process at enhanced rates for longer duration. PMID- 11118599 TI - Influence of environmental conditions on the activity of the recombinant mannuronan C-5-epimerase AlgE2. AB - The mannuronan C-5-epimerase AlgE2 is one of a family of Ca(2+)-dependent epimerases secreted by Azotobacter vinelandii. These enzymes catalyze the conversion of beta-D-mannuronic acid residues (M) to alpha-L-guluronic acid residues (G) in alginate. AlgE2 had a pH optimum between 6.5 and 7 and a temperature optimum around 55 degrees C. Addition of low molecular weight organic compounds, including buffers, amino acids and osmoprotective compounds, affected the activity of the enzyme. The charge, size and stereochemistry of the added compounds were important. The activity of AlgE2, dissolved in various buffers (same pH), decreased with increasing fraction of positively charged buffer ions. Mono- and divalent metal ions also influenced the activity. When Ca(2+) was omitted only Sr(2+), of the metal ions tested, supported some activity of AlgE2. At high concentration of Ca(2+) (3.3 mM) these ions had a negative effect on the activity, whereas at low Ca(2+) concentration (0.58 mM) the activity was enhanced by addition of Sr(2+), and to some degree also by addition of Mg(2+) and Mn(2+). During epimerization AlgE2 occasionally causes cleavage of the alginate chain. These chain breaks could not be prevented by changes in the conditions during the epimerization. The composition and sequential structure of epimerized alginate was not altered by changes in the epimerization conditions. PMID- 11118600 TI - Aspergillus niger I-1472 and Pycnoporus cinnabarinus MUCL39533, selected for the biotransformation of ferulic acid to vanillin, are also able to produce cell wall polysaccharide-degrading enzymes and feruloyl esterases. AB - The filamentous fungal strains Aspergillus niger I-1472 and Pycnoporus cinnabarinus MUCL39533, previously selected for the bioconversion of ferulic acid to vanillic acid and vanillin respectively, were grown on sugar beet pulp. A large spectrum of polysaccharide-degrading enzymes was produced by A. niger and very few levels of feruloyl esterases were found. In contrast, P. cinnabarinus culture filtrate contained low amount of polysaccharide-degrading enzymes and no feruloyl esterases. In order to enhance feruloyl esterases in A. niger cultures, feruloylated oligosaccharide-rich fractions were prepared from sugar beet pulp or cereal bran and used as carbon sources. Number of polysaccharide-degrading enzymes were induced. Feruloyl esterases were much higher in maize bran-based medium than in sugar beet pulp-based medium, demonstrating the ability of carbon sources originating from maize to induce the synthesis of feruloyl esterases. Thus, A. niger I-1472 could be interesting to release ferulic acid from sugar beet pulp or maize bran. PMID- 11118601 TI - The effects of aeration and veratryl alcohol on the production of two laccases by the ascomycete Botryosphaeria sp. AB - The ascomycete, Botryosphaeria sp, produced two extracellular constitutive laccases (PPO-I and PPO-II) active toward the substrates: 2, 2(1)-azino-bis(3 ethyl-benzthiazoline-6-sulfonic acid) [ABTS], and 2,6-dimethoxyphenol (DMP), respectively. The production of both laccases increased when the fungal isolate was grown in the presence of veratryl alcohol, and resulted in optimal laccase production (100- and 25- fold, respectively) at 40 mM. The effect of aeration on growth and laccase production was studied in baffled flasks, and showed that aeration of the cultures increased the production of both enzymes 4-5 fold in the presence of veratryl alcohol. Both laccases were susceptible to inhibition by azide, acetate and chloride anions. Veratryl alcohol inhibited the laccase catalyzed polymerization of DMP. Growing cultures of Botryosphaeria sp. produced an exopolysaccharide of the beta-glucan type whose synthesis was depressed when grown in the presence of veratryl alcohol. PMID- 11118602 TI - Two-stage cultivation of recombinant Saccharomyces cerevisiae to enhance plasmid stability under non-selective conditions: experimental study and modeling. AB - A leucine auxotroph strain of Saccharomyces cerevisiae was used to study plasmid stability and expression using a recombinant plasmid, which contained a foreign gene for firefly luciferase (luc). This recombinant yeast was tested in a series of continuous cultures in semi-defined media with varying concentrations of yeast extract in order to study its effect on stability. While the biomass concentration and luciferase activity increased with increasing concentrations of yeast extract, the plasmid stability declined. An analysis of the growth rates showed that the recombinants enjoyed a growth rate advantage over the plasmid free cells at critically low yeast extract concentrations, possibly due to leucine starvation in the media. A two-stage cultivation strategy was designed in order to create a yeast extract limited environment so that plasmid-free cells could not grow and overtake the recombinant cells. The cells were cultivated in selective media in the first stage, and then transferred continuously to the second stage where the media was enriched by feeding yeast extract. The feed rate was kept low in order to ensure yeast extract and hence leucine starvation, thereby selecting against the plasmid-free cells. This strategy resulted in a stable existence of recombinant cells, which stabilized around 60% at steady state during the tested period of cultivation. The complex nitrogen feed helped in increasing the cell density and volumetric activity by approximately 9 and 18 fold respectively with respect to that achieved in minimal medium. The experimental data was used to formulate a mathematical model to predict cell growth and plasmid stability in two-stage cultivation, which correctly explained the experimental data. PMID- 11118603 TI - The effect of yeast elicitor on the growth and secondary metabolism of hairy root cultures of Salvia miltiorrhiza. AB - Hairy root cultures of Salvia miltiorrhiza transformed with Agrobacterium rhizogenes ATCC 15834 produced a tiny amount of tanshinones and a constituent level of phenolic acids under normal growth conditions. Upon elicitation with yeast elicitor, the production of both phenolic acids and tanshinones was enhanced. For example, the contents of two phenolic acids, rosmarinic acid and lithospermic acid B were elevated from 1.24% and 2.59% to 2.89% and 2.98% of dry wt, respectively while the intracellular content of cryptotanshinone increased from 0.001% to as much as 0.096% of dry wt. Yeast elicitor also improved the growth of hairy roots (from 3.9 g/l to 7.3 g/l on a dry wt basis). Liquid chromatography-mass spectrometry (LC-MS) was developed for simultaneous detection and identification of phenolic acids and tanshinones in the extracts of S. miltiorrhiza. Rosmarinic acid, lithospermic acid B, cryptotanshinone, tanshinone I, tanshinone IIA and tanshinone IIB were identified by comparison with standards available. Dihydrotanshinone I and methylenetanshiquinone were tentatively identified by the molecular weights and the elution comparable with the literature. An unknown compound with a molecular weight of 280 was found in yeast elicitor treated hairy root cultures, which was one of the major tanshinones induced. PMID- 11118604 TI - Rice salT promoter is activated in Papaver somniferum and Nicotiana tabacum transgenic cells in the absence of exogenous ABA. AB - With the aim of modifying secondary metabolism in Opium poppy (Papaver somniferum) and tobacco (Nicotiana tabacum) cells, gene transfer was performed using the sam1 gene from Arabidopsis thaliana under the control of the salT promoter. This promoter is induced by ABA in rice and in tobacco and we have shown that it is also induced in poppy cells (gus gene). Putatively transformed poppy and tobacco cell lines with the sam1 gene were obtained. In the absence of exogenous inducer we noticed the expression of the transgene resulting in a significant increase of SAM-S activity in all tested transformants of poppy and in half the transgenic tobacco cell lines tested. Addition of ABA to the culture medium failed to enhance the expression of the transgene in both species and resulted in a decrease of the sam1 gene expression in some cell lines. Since the salT promoter is induced by exogenous ABA in both species (gus reporter gene), we suggest a partial sam1 transgene inactivation in certain cell lines. These results show that the efficiency of a regulatory sequence may be different when fused with a reporter gene (gus) compared to fusion with a gene belonging to the housekeeping family (sam1). PMID- 11118605 TI - Proteolytic anaerobic bacteria from lake sediments of Antarctica. AB - Amongst twenty five proteolytic bacteria isolated from lake sediment samples of Antarctica, six isolates were selected based on SDS PAGE protein profile and zone of hydrolysis on casein agar at 10 degrees C. Most of the cultures were rod shaped and motile with two showing terminal bulging spores. Isolates grew between 5 degrees C to 37 degrees C and protease was induced in the late log, stationary or death phase. Isolate SPA-3 grew maximally at 10 degrees C and SPA-6 at 37 degrees C while others preferred 20 degrees C-30 degrees C for growth. The growth and protease production on casein, skimmed milk, bovine serum albumin and gelatin varied with the isolates. Acetate was the dominant volatile fatty acid (24-66% of total VFA) produced during hydrolysis of protein substrate. PMID- 11118606 TI - REM sleep - by default? AB - Elements of three old, overlapping theories of REM sleep (REM) function, the Ontogenetic, Homeostatic and Phylogenetic hypotheses, together still provide a plausible framework - that REM (i) is directed towards early cortical development, (ii) "tones up" the sleeping cortex, (iii) can substitute for wakefulness, (iv) has a calming effect. This framework is developed in the light of recent findings. It is argued that the "primitiveness" of REM and its similarity to wakefulness liken it to a default state of "non-wakefulness" or a waking antagonist, anteceding "true" (non-REM) sleep. The "toning up" is reflected by inhibition of motor, sensory and (importantly) emotional systems, together pointing to integrated "flight or fight" activity, that preoccupies/distracts the organism when non-REM is absent and wakefulness unnecessary. Dreaming facilitates this distraction. In rodents, REM can provide stress coping and calming, but REM deprivation procedures incorporating immobility may further enhance stress and confound outcomes. REM "pressure" (e.g. REM rebounds) may be a default from a loss of inhibition of REM by non-REM. REM can be reduced and/or replaced by wakefulness, without adverse effects. REM has little advantage over wakefulness in providing positive cerebral recovery or memory consolidation. PMID- 11118607 TI - The prisoners of despair: right hemisphere deficiency and suicide. AB - This paper presents an integrative approach to understanding of the inner experience of suicidal persons in terms of hemispheric asymmetry. The right hemisphere is involved in formation of polysemantic context. Polysemantic context is determined by multiple interconnections among its elements, while each concrete element bears the stamp of the whole context. Left hemisphere functioning leads to formation of monosemantic context. It is suggested that due to functional insufficiency of the right hemisphere the suicidal person demonstrates a compensatory shift to left hemisphere functioning. This shift manifests itself in reversed asymmetry of neurotransmitters, tendency to dissociation, alienated and negative perception of the body, lower sensitivity to pain, disintegration of self-representation, cognitive constriction, overly general nature of personal memories, difficulties in affect regulation as well as such personality traits as low openness to experience and personal constriction. This hypothesis raises a number of suggestions for future research. PMID- 11118608 TI - Behavioral, neurophysiological and evolutionary perspectives on unihemispheric sleep. AB - Several animals mitigate the fundamental conflict between sleep and wakefulness by engaging in unihemispheric sleep, a unique state during which one cerebral hemisphere sleeps while the other remains awake. Among mammals, unihemispheric sleep is restricted to aquatic species (Cetaceans, eared seals and manatees). In contrast to mammals, unihemispheric sleep is widespread in birds, and may even occur in reptiles. Unihemispheric sleep allows surfacing to breathe in aquatic mammals and predator detection in birds. Despite the apparent utility in being able to sleep unihemispherically, very few mammals sleep in this manner. This is particularly interesting since the reptilian ancestors to mammals may have slept unihemispherically. The relative absence of unihemispheric sleep in mammals suggests that a trade off exists between unihemispheric sleep and other adaptive brain functions occurring during sleep or wakefulness. Presumably, the benefits of sleeping unihemispherically only outweigh the costs under extreme circumstances such as sleeping at sea. Ultimately, a greater understanding of the reasons for little unihemispheric sleep in mammals promises to provide insight into the functions of sleep, in general. PMID- 11118609 TI - Age differences in nociception and pain behaviours in the rat. AB - Much remains to be learned about the effects of ageing on pain. Studies of life span changes in nociception and pain behaviours in the rat are equivocal making it difficult to draw firm conclusions. This paper reviews the available data and finds that age differences in nociception may be dependent on the pain test employed. Specifically, reflexive responses to nociceptive stimuli do not change with age while there may be no change or a linear decrease with age on more highly organized tests of nociception. Interestingly, age differences in pain behaviours on models of tissue injury and inflammation may not be linear. It is shown that important changes that begin at mid-life in neuroanatomy, neurochemistry and endogenous pain inhibition may be associated with alterations in pain sensitivity. Several testable hypotheses which might encourage future research in this domain are developed throughout this paper. PMID- 11118610 TI - Insulin receptors and insulin action in the brain: review and clinical implications. AB - Insulin receptors are known to be located on nerve cells in mammalian brain. The binding of insulin to dimerized receptors stimulates specialized transporter proteins that mediate the facilitated influx of glucose. However, neurons possess other mechanisms by which they obtain glucose, including transporters that are not insulin-dependent. Further, insulin receptors are unevenly distributed throughout the brain (with particularly high density in choroid plexus, olfactory bulb and regions of the striatum and cerebral cortex). Such factors imply that insulin, and insulin receptors, might have functions within the central nervous system in addition to those related to the supply of glucose. Indeed, invertebrate insulin-related peptides are synthesized in brain and serve as neurotransmitters or neuromodulators. The present review summarizes the structure, distribution and function of mammalian brain insulin receptors and the possible implications for central nervous system disorders. It is proposed that this is an under-studied subject of investigation. PMID- 11118611 TI - Polymer chemical structure is a key determinant of physicochemical and colloidal properties of polymer-DNA complexes for gene delivery. AB - Polyplexes are now emerging as potentially useful vectors for gene therapy. To improve our understanding of how the chemical structure of the polymer affects the properties of these systems, a series of structurally related polymers, the linear poly(amidoamine)s (PAAs), have been examined for their abilities to form complexes with DNA. Structure-dependent differences in DNA binding are shown by gel electrophoretic retardation of DNA and thermal transition analyses. Two PAAs, NG28 and NG30, stand out as having high affinity DNA binding characteristics, similar to the model homopolypeptide, poly-L-lysine. In addition, differences in complex formation, particle size and surface charge are displayed for the different polymer-DNA systems. Electron microscopy studies showed that the polymers condensed DNA into similar unit structures but only complexes with NG30 did not undergo agglomeration. This was attributed to an excess of complexed polymer forming a shell of uncomplexed polymer chain segments around a condensed DNA-polymer core. The transfection activities of these polymer complexes differ greatly, and some of these differences can be explained in a multifactorial way by the physicochemical and colloidal properties. It is concluded that polymer chemical structure dictates the apparent affinity of DNA binding, and also several of the important colloidal characteristics of the resulting complexes. PMID- 11118612 TI - Impaired expression of glutathione synthetic enzyme genes in mice with targeted deletion of the Nrf2 basic-leucine zipper protein. AB - Transcriptional activation of genes that play a role in detoxification of xenobiotics and defense against oxidative stress is mediated in part by the antioxidant response element (ARE). For example, it has been shown that the promoters for both the heavy and light chain gamma-glutamylcysteine synthetase (GCS(H) and GCS(L)) genes require the ARE. CNC-bZIP factors, together with small Maf proteins, have been shown to bind as heterodimers to the NF-E2/AP-1 element, which is similar to the consensus sequence for the ARE. Nrf1 and Nrf2, two widely expressed CNC-bZIP factors, have been implicated in the regulation of genes involved in oxidative stress response. In this study, we examined the effect of nrf2 mutation on the expression of genes involved in glutathione synthesis. We observed that transcripts for gcs(H) and gcs(L) genes were decreased in nrf2(-/-) fibroblasts and livers. Correspondingly, glutathione levels were decreased in Nrf2 deficient livers and fibroblasts. By transient transfection studies in nrf2( /-) fibroblasts, we show that transcriptional activation of reporter constructs bearing the human GCS(L) promoter, as well as the functional ARE of GCS(H) promoter, required the activator protein Nrf2. By electrophoretic mobility shift assay, recombinant Nrf2 binds the ARE of the GCS(L) and GCS(H) promoters. Overexpression of Nrf2 cDNA restored glutathione (GSH) levels in nrf2(-/-) fibroblasts, which correlated with increased steady state levels of gcs(H) and gcs(L) transcripts. These results establish a link between Nrf2 transcription factor and GSH biosynthesis. PMID- 11118613 TI - The COUP-adjacent repressor (CAR) element participates in the tissue-specific expression of the ovalbumin gene. AB - The ovalbumin (Ov) gene is an excellent model for the study of tissue-specific gene regulation as it is only active in the estrogen-stimulated oviduct. Previous studies have demonstrated that the negative regulatory element (NRE) in the Ov gene 5'-flanking region is responsible for silencing the gene in oviduct in the absence of steroids. Linker scanning analysis defined an element within the NRE designated the COUP-adjacent repressor (CAR) element as a repressor of Ov gene expression. However, the role of the CAR element in non-oviduct tissues has not been addressed. Using transient transfection analysis of various Ov 5'-flanking region constructs into the estrogen-responsive chicken hepatocyte cell line LMH/2A, we demonstrate that Ov gene expression is not induced by estrogen and that an active repressor element exists in the NRE. Deletion analysis indicates that the region from -134 to -87, which includes the CAR element, mediates this repression. Mutation of the CAR element relieves repression, leading to high levels of gene expression. These data support a model where the inhibition of Ov gene expression in non-oviduct cells is a combination of the lack of essential positive factors and the presence of an active repressor, which binds to the CAR element. PMID- 11118614 TI - Size distribution of the urokinase mRNA decay intermediates in different tissues and cell lines. AB - Many genes, particularly those encoding the products participating in the regulation of transcription, replication and tissue remodeling, produce short lived mRNA. It has been commonly accepted that once mRNA is disintegrated, the degradation process is so rapid that the decay intermediates cannot be detected. In the present study we verified this postulate and focused our attention on the quantification of the decay products of the urokinase-type plasminogen activator (uPA) mRNA that belongs to short-lived mRNAs. Using a previously described modified quantitative RT-PCR method, we have shown that intact uPA mRNA coexists in normal human tissues, Jurkat and 5637 cells with a great abundance of its degradation products. The uPA mRNA decay products were not detected in T24P cells. The content of intact uPA mRNA in normal tissues was as low as 5% of the total amount of its poly(A)(+) fraction. The size distribution of the mRNA decay products suggests that the mRNA is digested by exonucleases or/and non-specific endonuclease with cut sites evenly distributed along the mRNA chain. Different decay degrees were demonstrated for subpopulation of the uPA mRNA molecules with intact 3' and 5' ends. PMID- 11118615 TI - Alternative splicing and gene structure of the transforming growth factor beta activated kinase 1. AB - We have identified a fourth splice variant of the TGF beta-activated kinase (TAK1), called TAK1-d, and identified an error in the previously published TAK1-c sequence. Our data shows that the c and d variants encode proteins whose carboxyl ends differ markedly from those of variants a and b. Analysis of the human TAK1 gene sequence, located at 6q16.1-q16.3, shows that the coding sequence is organised in 17 exons. The four splice variants result from alternative splicing of exons 12 and 16, the reading frame of exon 17 being determined by the presence or absence of exon 16. Study of the relative levels of expression of the four splice variants showed significant variations between tissues. Our evidence suggests that the alternative splicing of the TAK1 mRNA may have important functional implications. PMID- 11118616 TI - Molecular and immunological characterisation of a polymorphic cytosolic fatty acid binding protein from the human blood fluke of humans, Schistosoma japonicum. AB - Most organisms obtain their fatty acids through their diet or by de novo synthesis, but human blood flukes belonging to the genus Schistosoma lack the oxygen-dependent pathways required for the synthesis of sterols and fatty acids so they are entirely dependent on their hosts for these and other complex lipids. Fatty acid binding proteins (FABPs) of the FABP/P2/CRABP/CRBP family of beta barrel cytosolic lipid binding proteins (cLBP) appear to be particularly important to schistosomes in the uptake, transport and compartmentalisation of host-derived fatty acids and may provide important targets for immuno- and chemotherapy. Here we describe the isolation of a set of cDNAs prepared from the Asiatic schistosome, Schistosoma japonicum, which encode two groups of cLBPs based on sequence homology and unique cDNA restriction sites. Representative clones from the two groups, one encoding a complete Sj-FABP (F10), and the other encoding a deletion mutant (F25) were characterised at the nucleic acid level by Southern and Northern hybridisation analysis, and at the protein level by immunoblotting. The presence and size of introns in the genes encoding F10 and F25 were determined and, because of the interest in the Schistosoma mansoni FABP homologue (Sm14) as a putative vaccine candidate, the immunogenicity and protective efficacy of the two proteins were also evaluated. A particularly interesting finding was the degree of Sj-FABP amino acid sequence polymorphism found to occur within the S. japonicum worm population, which appears to be greater than that described from cLBPs from vertebrates or, indeed, any other group of organisms investigated to date. PMID- 11118617 TI - Cloning and characterization of the human phosphoinositide-specific phospholipase C-beta 1 (PLC beta 1). AB - Phospholipase C-beta (PLC beta) catalyses the generation of inositol 1,4,5 trisphosphate (IP(3)) and diacylglycerol (DAG) from phosphatidylinositol 4,5 bisphosphate (IP(2)), a key step in the intracellular transduction of a large number of extracellular signals, including neurotransmitters and hormones modulating diverse developmental and functional aspects of the mammalian central nervous system. Four mammalian isozymes are known (PLC beta 1-4), which differ in their function and expression patterns in vivo. We have characterized the human PLC beta 1 genomic locus (PLC beta 1), cloned two distinct PLC beta 1 cDNAs (PLC beta 1a and b) and analysed their respective expression patterns in a comprehensive panel of human tissues using quantitative TaqMan technology. The two cDNAs derive from transcripts generated through alternative splicing at their 3' end, and are predicted to encode for PLC beta 1 isoforms differing at their carboxy-terminus. The human PLC beta 1 isoforms are co-expressed in the same tissues with a distinctly CNS-specific profile of expression. Quantitative differences in PLC beta 1 isoform expression levels are observed in some tissues. Transient expression of epitope-tagged versions of the two isoforms followed by immunofluorescence revealed localization of the proteins to the cytoplasm and the inner side of the cell membrane. Finally, we characterized the structure of the PLC beta 1 locus and confirmed its mapping to human chromosome 20. PMID- 11118618 TI - Upstream stimulatory factor is involved in the regulation of the human calcyclin (S100A6) gene. AB - Calcyclin (S100A6) is a calcium-binding protein overexpressed in several tumor cell lines including melanoma with high metastatic activity. The calcyclin gene promoter fragment -361/-167 activates transcription several fold when compared to the basal -167/+134 promoter fragment indicating the presence of enhancer element within -361/-167 bp region. By means of the electrophoretic mobility shift assay (EMSA) we found that this region contains a protein-binding site and mapped it to an E-box sequence at position -283/-278. Using antibodies against USF1 we identified the upstream stimulatory factor as the transcription factor bound to the E-box sequence in EMSA. This factor was also enriched in protein fractions obtained from Ehrlich ascites tumor cells nuclear extract by affinity chromatography using the E-box sequence as a ligand. Cotransfection of the USF1 expression vector with a plasmid carrying the luciferase gene under control of the -361/+134 calcyclin gene promoter fragment resulted in several fold activation of luciferase activity. On the other hand, mutations within the E-box led to a marked decrease in the efficiency of calcyclin gene promoter fragment. The results indicate that USF1 binds to an E-box sequence of the calcyclin gene promoter and enhances its transcription activity. This mechanism might be responsible for the upregulation of calcyclin gene expression in response to various stimuli and in tumors. PMID- 11118619 TI - Tumor cell splice variants of the transcription factor TEF-1 induced by SV40 T antigen transformation. AB - The large tumor antigen (TAg) of simian virus 40 is able to transform cells through interactions with cellular proteins, notably p53 and Rb. Among the other proteins that form complexes with TAg is TEF-1, a transcription factor utilized by the viral enhancer to activate expression of the early gene which encodes TAg. We show that fibroblasts contain several alternately spliced TEF-1 mRNAs, the most abundant of which encodes a protein with an additional four amino acid exon compared to the database entry for Hela cell TEF-1. Transformation by TAg induces alternate splicing, producing a more abundant form lacking this exon and matching the published sequence. Splicing variants lacking this exon were detected in mouse pancreatic tumors and in cell lines derived from human pancreatic cancers, in contrast to a single isoform with the exon in normal mouse pancreas. A total of eight splice variants were identified, with the loss of the four amino acid exon typical of transformed cells. These and other data presented suggest that TAg 're-models' host cell transcription factors that are used early in viral infection, and thereby mimics an event that naturally occurs during transformation. The data indicate that TEF-1 alterations may be a hallmark feature of tumorigenesis. PMID- 11118620 TI - Thyroid hormone, glucagon, and medium-chain fatty acids regulate transcription initiated from promoter 1 and promoter 2 of the acetyl-CoA carboxylase-alpha gene in chick embryo hepatocytes. AB - High-carbohydrate feeding and triiodothyronine (T3) increase the abundance of acetyl-CoA carboxylase-alpha (ACC alpha) mRNA in avian hepatocytes, whereas starvation, glucagon, and medium-chain fatty acids decrease the abundance of ACC alpha mRNA. These changes in ACC alpha mRNA levels are mediated by alterations in the rate of transcription of the ACC alpha gene. In liver, ACC alpha transcription is initiated from two promoters, promoter 1 and promoter 2, resulting in transcripts that contain heterogeneity in their 5'-untranslated regions. Here, we investigated the role of promoter 1 and promoter 2 in mediating nutrient- and hormone-induced changes in ACC alpha mRNA abundance by measuring the level of transcripts expressed from promoter 1 and promoter 2 using a ribonuclease protection assay. The results indicated that both promoter 1 and promoter 2 were regulated by starvation/refeeding in livers of intact chicks and by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocyte cultures and that alterations in the activity of promoter 2 accounted for a greater proportion of the changes in total ACC alpha mRNA abundance caused by nutrient and hormone treatment. Five DNase-hypersensitive sites were also identified between -500 and +1 bp relative to the transcription start site of promoter 2 in livers of intact chicks and in chick embryo hepatocyte cultures. In transient transfection analyses, this region of DNase hypersensitivity conferred regulation of transcription by T3, glucagon, and medium-chain fatty acids in chick embryo hepatocytes. Data from this study demonstrate that diet-induced changes in the activities of promoter 1 and promoter 2 in livers of intact chicks are mimicked in chick embryo hepatocyte cultures by manipulating the concentrations of T3, glucagon and medium-chain fatty acids in the culture medium and that cis-acting sequences mediating the effects of nutrients and hormones on promoter 2 activity are located immediately upstream of the transcription start site of this promoter. PMID- 11118621 TI - Japanese encephalitis virus up-regulates expression of macrophage migration inhibitory factor (MIF) mRNA in the mouse brain. AB - Macrophage migration inhibitory factor (MIF) is known as a proinflammatory cytokine, glucocorticoid-induced immunomodulator, and pituitary hormone, and contributes to broad-spectrum immune and inflammatory response. To investigate the expression of MIF in the central nervous system in an event of viral infection, we evaluated MIF mRNA expression in the mouse brain infected with Japanese encephalitis virus (JEV). In situ hybridization revealed that MIF mRNA expression was significantly up-regulated in the whole brain by intracranial JEV inoculation at 2 days post-inoculation (d.p.i.). Neurons as well as glial cells expressed MIF transcripts in which some of these cells were co-labeled by double staining for JEV antigens and MIF mRNA. At 4 d.p.i., when typical symptoms of encephalitis were observed, JEV antigen-positive cells were much increased in parallel with enhanced MIF mRNA, consistent with the results of Northern blot analysis. Reverse transcription-polymerase chain reaction showed that MIF mRNA was minimally changed at 1 d.p.i. in comparison with that at 0 d.p.i., but markedly up-regulated after 2 d.p.i. and sustained up to 4 d.p.i. On the other hand, a significant increase of tumor necrosis factor (TNF)-alpha mRNA was observed after only 3 d.p.i. These data suggest the possibility that MIF is involved in virus-induced encephalitis with regard to not only immune responses in the early stage, but also the exacerbation of inflammation in concert with TNF alpha in the late stages. This is the first evidence demonstrating that MIF is up regulated in the case of virus-induced encephalitis, which should contribute to the further understanding of the pathological mechanism of JEV-induced encephalitis. PMID- 11118622 TI - Functional characterization of a water channel of the nematode Caenorhabditis elegans. AB - A genome project for the species Caenorhabditis elegans has demonstrated the presence of eight cDNAs belonging to the major intrinsic protein (MIP) family. We previously characterized one of these cDNAs known as C01G6.1. C01G6.1 was confirmed to be a water channel and newly designated as AQP-CE1 [Am. J. Physiol. 275 (1998) C1459-C1464]. In this paper, we examined the function of another MIP protein encoded by F40F9.9. This cDNA encodes a 274-amino acid protein showing a high sequence identity with mammalian aquaporin-8 (AQP8) water channel (35%) and d-TIP (34%), an AQP of Arabidopsis. The expression of F40F9.9 in Xenopus oocytes increased the osmotic water permeability (P(f)) 10.4-fold, and the activation energy for P(f) from Arrhenius plot was 4.7 kcal/mol, suggesting that F40F9.9 is a water channel (AQP-CE2). AQP-CE2 was not permeable to glycerol or urea. Oocyte P(f) was reversibly inhibited by 58% after an incubation with 0.3 mM HgCl(2). To identify the mercury-sensitive site, four individual cysteine residues in AQP-CE2 (at positions 47, 132, 149, 259) were altered to serine by site-directed mutagenesis. Of these mutants, only C132S had a P(f) similar to that of the wild type together with an acquired mercury resistance, suggesting that Cys-132 is the mercury-sensitive site. Similar results were obtained by the mutation of Cys-132 to alanine (C132A). Replacement of Cys-132 with tryptophan decreased P(f) by 64%, but P(f) was still 2.5 times higher than that of the control. Cys-132 is located in the transmembrane helix 3, close to the transition to the extracellular loop C. These results suggest that the transmembrane helix 3, including Cys-132, might participate in the aqueous pore formation, or, alternatively, that Cys-132 might contribute to the construction of the AQP protein. PMID- 11118623 TI - UCP3 gene expression does not correlate with muscle oxidation rates in troglitazone-treated Zucker fatty rats. AB - Uncoupling protein-3 (UCP3), a mitochondrial carrier protein predominantly expressed in muscle, has been suggested to release stored energy as heat. The insulin-sensitizing thiazolidinediones enhance glucose disposal in skeletal muscle and have been reported to increase the expression of uncoupling proteins in various experimental systems. We therefore studied the effect of troglitazone treatment on UCP3 gene expression in muscles from lean and obese Zucker rats. In comparison with obese littermates, basal UCP3 mRNA levels in lean Zucker rats tended to be higher in white and red gastrocnemius muscles, but were lower in soleus (P<0.001) muscle and heart (P<0.01). In lean rats, troglitazone significantly increased UCP3 gene expression in white and red gastrocnemius and heart muscles (all P<0.01). In contrast, the drug reduced UCP3 mRNA expression in red gastrocnemius and soleus muscles of obese littermates (all P<0.001). The troglitazone-dependent decrease in UCP3 gene expression was accompanied by an increased weight gain in obese rats, while no such effect was observed in lean rats. In obese rats, improvement of insulin resistance by troglitazone was associated with increased rates of basal and insulin-stimulated CO(2) production from glucose measured in soleus muscle. These studies demonstrate that effects of troglitazone on UCP3 gene expression depend on the phenotype of Zucker rats and that troglitazone-induced metabolic improvements are not related to increased uncoupling resulting from upregulation of UCP3 mRNA expression in muscle. PMID- 11118624 TI - Upstream stimulatory factor 2 stimulates transcription through an initiator element in the mouse cytochrome c oxidase subunit Vb promoter. AB - Upstream stimulatory factor (USF) is a basic helix-loop-helix-leucine zipper transcription factor that plays an important role in transcriptional activation and cell proliferation. In this article, we demonstrate that the mouse cytochrome c oxidase subunit Vb gene (Cox5b) can be transactivated by ectopic expression of USF2 through an initiator (Inr) element in the core promoter. Importantly, using a dominant-negative mutant of USF2, we demonstrate the role of endogenous USF2 proteins in the transcriptional activation of the Cox5b Inr. Domains of USF2 encoded by exon 4, exon 5 and the USF-specific region are important for maximum activation of the Cox5b Inr. Using the adenovirus E1A oncoprotein, we show that p300/CBP acts as a coactivator in the USF2-dependent activation of the Cox5b Inr. We also demonstrate that although expression of multifunctional regulatory factor, Yin Yang 1 (YY1), can stimulate transcription of the Cox5b Inr to a modest extent, expression of YY1 together with USF2 greatly reduces the level of activation of the Cox5b Inr. Furthermore, we show that the transcription factor, Sp1, represses both the YY1- and the USF2-dependent activation of the Cox5b Inr, indicating competition among Sp1, YY1, and USF2. PMID- 11118625 TI - Isolation and characterisation of a novel human gene (C9orf11) on chromosome 9p21, a region frequently deleted in human cancer. AB - The chromosome 9p21 region has been described to be frequently deleted in several neoplasias. The cyclin dependent kinase inhibitor 2A (CDKN2A or P16) gene was cloned in this region and identified as a tumour suppressor gene. However, much evidence indicates the existence of another tumour suppressor gene located proximal to the CDKN2A gene, which could be involved in cutaneous malignant melanoma (CMM) initiation. In the present report we have further investigated this 9p21 chromosomal region and cloned and characterised a novel gene within it (C9orf11). This gene shares no similarities to any known gene or predicted protein representing a novel human gene. Nevertheless, a putative leucine zipper pattern is located at the C-terminal end of the predicted protein, suggesting that it could dimerise. C9orf11 encodes for a protein of 294 amino acids with a predicted molecular mass of 32.8 kDa. C9orf11 is organised in eight exons that encompass a region of approx. 13 kb. Expression analysis demonstrates that C9orf11 is highly expressed in testis, although minor expression was seen in other tissues. Mutations in the C9orf11 gene were not detected in CMM families that were negative for CDKN2A mutations. Two SNPs for the C9orf11 gene have been identified, which could be used in segregation or association studies for other disorders. PMID- 11118626 TI - Genetic analysis of the gene cluster for the synthesis of serotype a-specific polysaccharide antigen in Aactinobacillus actinomycetemcomitans. AB - The serotype a-specific polysaccharide antigen (SPA) of Actinobacillus actinomycetemcomitans consists of 6-deoxy-D-talose. A gene cluster associated with the biosynthesis of SPA was cloned and sequenced from the chromosomal DNA of A. actinomycetemcomitans SUNYaB 75 (serotype a). This cluster consisted of 14 open reading frames. Insertional inactivation of eight genes in this cluster resulted in loss of the ability of A. actinomycetemcomitans SUNYaB 75 cells to produce the polysaccharide. A protein database search revealed that the 11 sequential genes containing these eight genes were not found in SPA-associated gene clusters of the other serotypes of A. actinomycetemcomitans. These results suggest that the gene cluster is unique to serotype a and is essential to the synthesis of the SPA. PMID- 11118627 TI - A Phaseolus vulgaris lipoxygenase gene expressed in nodules and in Rhizobium tropici inoculated roots. AB - A genomic clone encoding a common bean lipoxygenase (PvLOX5) was isolated from a Phaseolus vulgaris library. Reverse transcription-polymerase chain reaction analysis revealed that PvLOX5 is expressed during nodule development and in Rhizobium tropici inoculated roots. There was no detectable expression of PvLOX5 in non-inoculated roots, healthy leaves, leaves after Pseudomonas syringae pv. tabaci infection, floral buds or dry seeds. PMID- 11118628 TI - Candidate osmosensors from Candida utilis and Kluyveromyces lactis: structural and functional homology to the Sho1p putative osmosensor from Saccharomyces cerevisiae. AB - In Saccharomyces cerevisiae, increases in external osmolarity evoke osmostress induced signalling via the HOG MAP kinase pathway. One of the upstream components of this signal transduction route is the putative osmosensor, Sho1p. With the aim to elucidate the molecular basis of osmosensing in budding yeast, we have cloned SHO1 homologues from Candida utilis and Kluyveromyces lactis which allowed determination of conserved domains of Sho1p. Results obtained from sequence comparisons, confirmed the importance of the transmembrane domains and the SH3 domain for Sho1p function. The K. lactis and S. cerevisiae Sho1p show the highest degree of homology, the isoform from C. utilis is a shorter protein. SHO1 from C. utilis, however, did complement the osmosensitivity of the sho1ssk2ssk22 strain by restoring HOG pathway function, since Hog1p dual phosphorylation after high osmotic challenge was restored in this strain after transformation with a plasmid bearing this SHO1 homologue. PMID- 11118629 TI - Cloning and preliminary characterization of a 105 kDa protein with an N-terminal kinase-like domain. AB - In the course of searching for proteins that interact with protein kinase B in 3T3-L1 adipocytes, we isolated a 105 kDa protein from 3T3-L1 adipocytes. Peptides sequenced from the protein were found to be present in several expressed sequence tags. A cDNA containing one of these expressed sequence tags was sequenced and appears to contain the entire coding region. Computer analysis revealed a potential protein kinase domain at the N-terminus; however, the first subdomain and several invariant residues characteristic of protein kinases are absent. An antibody was raised against a peptide from the 105 kDa protein. By immunoblotting, it was found that the protein was widely expressed in mouse tissues, and concentrated in the cytosol and low density microsome fractions of 3T3-L1 adipocytes. PMID- 11118630 TI - Sequence analysis of the first complete cDNA clone encoding an American cockroach Per a 1 allergen. AB - C3, designated as Per a 1.0103 according to WHO/IUIS nomenclature, encoding a novel American cockroach allergen of 395 aa (44.6 kDa), is the first complete cDNA clone with a translatable immunoreactive protein among the reported group 1 cockroach allergens. Its deduced amino acid sequence possesses a signal sequence, phosphorylation sites, mitochondrial energy transfer protein signatures, and four repeats each sharing striking similarity with the corresponding repeats of the other cockroach allergens. The latter similarity suggests that the group 1 cockroach allergens might have evolved from a primordial mitochondrial sequence by exon duplication. PMID- 11118631 TI - Cloning and expression of cDNAs encoding two enzymes of the MEP pathway in Catharanthus roseus. AB - Two periwinkle cDNAs (crdxr and crmecs) encoding enzymes of the non-mevalonate terpenoid pathway were characterized using reverse transcription-PCR strategy based on the design of degenerated oligonucleotides. The deduced amino acid sequence of crdxr is homologue to 1-deoxy-D-xylulose 5-phosphate reductoisomerases. Crmecs represents the first plant cDNA encoding a protein similar to the 2C-methyl-D-erythritol 2,4-cyclodiphosphate synthase from Escherichia coli. Expression of crdxr and crmecs genes was up-regulated in periwinkle cells producing monoterpenoid indole alkaloids. Involvement of the 2C methyl-D-erythritol 4-phosphate pathway in alkaloid biosynthesis is discussed. PMID- 11118632 TI - Presence of new alternative exons in human and mouse Fli-1 genes. AB - The mouse Fli-1 proto-oncogene is activated by proviral integration of four murine leukemia retroviruses and its human counterpart is translocated (11,22) in Ewing tumors. We have identified two alternative exons 1 by RACE analysis from a human neuroectodermal tumor. Exons 1a and 1b are located respectively 1.3 and 2.5 kb upstream from the published exon 1. Translation of these alternative messengers is predicted to generate very similar proteins. The sequence upstream from exon 1b showed functional promoter activity. Exon 1b was not conserved in the mouse but was detected in every analyzed human cell, whereas exon 1a was present only in a subset of them and also in various mouse cell lines. These results suggest that both mouse and human Fli-1 gene expression might be under the control of several independent promoter regions. PMID- 11118633 TI - Cloning, expression and regulation of Schizosaccharomyces pombe gene encoding thioltransferase. AB - The genomic DNA encoding thioltransferase was isolated from Schizosaccharomyces pombe using the polymerase chain reaction. The amplified DNA fragment was confirmed by Southern hybridization, completely digested with HindIII and BamHI, and then ligated into the yeast-Escherichia coli shuttle vector pRS316, which resulted in plasmid pEH1. The insert of plasmid pEH1 was transferred into the multi-copy vector YEp357 to generate plasmid pYEH1. The determined nucleotide sequence harbors an open reading frame consisting of four exons and three introns, which encodes a polypeptide of 101 amino acids with a molecular mass of 11261 Da. Thioltransferase activity was increased 1.6-fold in Saccharomyces cerevisiae containing plasmid pYEH1, and 1.8- and 2.7-fold in S. pombe containing plasmid pEH1 and pYEH1, respectively. The upstream sequence and the region encoding the N-terminal six amino acids were fused into promoterless beta galactosidase gene of the shuttle vector YEp357R to generate the fusion plasmid pYEHR1. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by zinc and NO-generating S-nitroso-N-acetylpenicillamine. PMID- 11118634 TI - Influence of pentoxifylline, A-802710, propentofylline and A-802715 (Hoechst) on the expression of cell cycle blocks and S-phase content after irradiation damage. AB - The toxicity of the five methylxanthine derivatives, caffeine, pentoxifylline, A802710, propentofylline and A802715, was determined against the two human melanoma lines, Be11 and MeWo, and against the two human squamous cell carcinoma lines, 4197 and 4451, by vital dye staining assay. Pentoxifylline and A802710 emerge as the least toxic showing TD(50) (toxic dose of 50%) levels of 3.0-4.0 mM. Propentofylline and caffeine take an intermediate position. A802715 has a TD(50) of 0.9-1.1 mM and is the most toxic. Subtoxic concentrations (MMP-9) and of native B cells (MMP 9>TIMP-1) is suggestive of a new function for TIMP-1. We propose that TIMP-1 acts as a survival factor controlling B-cell growth and apoptosis through an autocrine regulation process involving IL-10 secreted by EBV-B lymphocytes. PMID- 11118637 TI - Identification of two focal adhesion targeting sequences in the adapter molecule p130(Cas). AB - The adapter molecule CAS is localized primarily within focal adhesions in fibroblasts. Because many of the cellular functions attributed to CAS are likely to be dependent on its presence in focal adhesions, this study was undertaken to identify regions of the protein that are involved in its localization. The SH3 domain of CAS, when expressed in isolation from the rest of the protein, was able to target to focal adhesions, whereas a variant containing a point mutation that rendered the SH3 domain unable to associate with FAK remained cytoplasmic. However, in the context of full-length CAS, this mutation did not prevent CAS localization to focal adhesions. Two other variants of CAS that contained deletions of either the SH3 domain alone, or the SH3 domain together with an adjoining proline-rich region, also retained the capacity to localize to focal adhesions. A second focal adhesion targeting region was mapped to the extreme carboxy terminus of CAS. The identification of this second focal adhesion targeting domain in CAS ascribes a previously unknown function to the highly conserved C terminus of CAS. The regulated targeting of CAS to focal adhesions by two independent domains may reflect the important role of CAS within this subcellular compartment. PMID- 11118638 TI - kappa-Opioid receptor potentiates apoptosis via a phospholipase C pathway in the CNE2 human epithelial tumor cell line. AB - The mechanism by which kappa-opioid receptor (kappaor) modulated apoptosis was investigated in CNE2 human epithelial tumor cells. Induction of these cells to undergo apoptosis with staurosporine was associated with a massive increase in intracellular cAMP level. The inhibition of the increase in cAMP partially inhibited apoptosis as evidenced by a reduction of PARP and caspase-3 cleavage. Accordingly, a low but significant level of apoptosis is induced in these cells by the elevation of cAMP through the addition of forskolin and isobutylmethylxanthine. The existence of a cAMP-dependent and a cAMP-independent apoptotic pathway is therefore suggested. Receptor binding studies, RT-PCR experiments and Western blot analysis demonstrated the presence of type 1 kappaor in the CNE2 cells. Stimulation of kappaor in these cells resulted in the production of inositol (1,4,5)-trisphosphate, reduction of cAMP level and a marked enhancement of staurosporine-induced apoptosis. The potentiation of apoptosis by kappaor was prevented by inhibition of phospholipase C but was slightly enhanced by the presence of the active cAMP analogues, 8-CPT-cAMP and dibutyryl-cAMP. These data demonstrate for the first time that the phospholipase C pathway activated by type 1 kappaor expressed by cancer cells is involved in the potentiation of apoptosis. PMID- 11118639 TI - Oligomerization properties of the acidic ribosomal P-proteins from Saccharomyces cerevisiae: effect of P1A protein phosphorylation on the formation of the P1A-P2B hetero-complex. AB - Acidic ribosomal P-proteins form, in all eukaryotic cells, a lateral protuberance, the so-called 'stalk', which is directly involved in translational activity of the ribosomes. In Saccharomyces cerevisiae cells, there are four distinct P-proteins: P1A, P1B, P2A and P2B. In spite of the high level of their structural homology, they are not completely equivalent and may perform different functions. As yet, the protein-protein interactions between yeast P-proteins have not been fully defined. In this paper, the interplay between yeast P-proteins has been investigated by means of a two-hybrid system, chemical cross-linking and gel filtration. The data presented herein show that all P-proteins are able to form homo-oligomeric complexes. By analyzing hetero-interactions, we were able to detect strong interactions between P1A and P2B proteins. Additionally, the pair of P1B and P2A proteins is also able to form a hetero-complex, though at a very low efficiency. All P-proteins are phosphorylated by numerous protein kinases. Using the multifunctional protein kinase CK II, we have shown that incorporation of phosphate into P1A protein can exert its effect on the hetero-oligomerization process, namely by preventing the formation of the hetero-oligomer P1A-P/P2B. These findings are the first to show differences in the oligomerization behavior of the yeast P-proteins; moreover, they emphasize a significant impact of the phosphorylation on the formations of P-protein complex. PMID- 11118640 TI - Alternative glycosylation of the insulin receptor prevents oligomerization and acquisition of insulin-dependent tyrosine kinase activity. AB - Glucose deprivation leads to the synthesis of an aberrantly glycosylated ('alternative') and inefficiently processed form of the insulin proreceptor in 3T3-L1 adipocytes. To further explore the effect of aberrant (rather than absent) N-linked glycosylation of the insulin receptor, we examined the relationship of processing to function. Our studies show that the alternative form of the proreceptor does not oligomerize nor does it acquire the ability to undergo insulin-sensitive autophosphorylation. This along with an interaction with the glucose-regulated stress protein GRP78/BiP implies inappropriate folding/dimerization and retention in the ER. Glucose refeeding causes the post translational modification of the alternative form of the proreceptor to a novel 'intermediate' form which is independent of new protein synthesis. As little as 100 microM glucose (or mannose) can induce this modification. In vitro digestion of the alternative and intermediate proreceptors with SPC1/furin shows that both the alpha- and beta-subunit domains are glycosylated, albeit aberrantly. This implies that the aberrantly glycosylated proreceptor could serve as a substrate for SPC1 in a physiological setting if the receptor was able to interact with the enzyme in the appropriate compartment (i.e., the trans-Golgi network). Based on inhibitor studies, however, both the alternative and intermediate forms of the proreceptor appear to be primarily targeted to the proteasome for degradation. PMID- 11118641 TI - Mutations in SPC110, encoding the yeast spindle pole body calmodulin-binding protein, cause defects in cell integrity as well as spindle formation. AB - The 110 kDa spindle pole body component, Spc110p, is an essential target of calmodulin in budding yeast. Cells with mutations which reduce calmodulin binding to Spc110p are unable to form a mitotic spindle and die. Here we show that these effects can be overcome either directly by increasing extracellular calcium or calmodulin expression, which reverse the primary spindle defect, or indirectly through increased extracellular osmolarity or high dosage of MID2 or SLG1/HCS77/WSC1 which preserve viability. We propose that overcoming a cell integrity defect associated with the mitotic arrest enables the defective spindle pole bodies to provide sufficient function for proliferation of a large proportion of mutant cells. Our findings demonstrate a role for calcium in the Spc110p-calmodulin interaction in vivo and have important general implications for the interpretation of genetic interactions involving cell integrity genes. PMID- 11118642 TI - The effect of pulsed electromagnetic fields on the physiologic behaviour of a human astrocytoma cell line. AB - We evaluated the effects of 50 Hz pulsed electromagnetic fields (EMFs) with a peak magnetic field of 3 mT on human astrocytoma cells. Our results clearly demonstrate that, after the cells were exposed to EMFs for 24 h, the basal [Ca(2+)](i) levels increased significantly from 124+/-51 nM to 200+/-79 nM. Pretreatment of the cells with 1.2 microM substance P increased the [Ca(2+)](i) to 555+/-278 nM, while EMF exposure caused a significant drop in [Ca(2+)](i) to 327+/-146 nM. The overall effect of EMFs probably depends on the prevailing Ca(2+) conditions of the cells. After exposure, the proliferative responses of both normal and substance P-pretreated cells increased slightly from 1.03 to 1.07 and 1.04 to 1.06, respectively. U-373 MG cells spontaneously released about 10 pg/ml of interleukin-6 which was significantly increased after the addition of substance P. Moreover, immediately after EMF exposure and 24 h thereafter, the interleukin-6 levels were more elevated (about 40%) than in controls. On the whole, our data suggest that, by changing the properties of cell membranes, EMFs can influence Ca(2+) transport processes and hence Ca(2+) homeostasis. The increased levels of interleukin-6 after 24 h of EMF exposure may confirm the complex connection between Ca(2+) levels, substance P and the cytokine network. PMID- 11118643 TI - Differential effects of progesterone and 17beta-estradiol on the Ca(2+) entry induced by thapsigargin and endothelin-1 in in situ endothelial cells. AB - The effects of progesterone and 17beta-estradiol on Ca(2+) signaling in in situ endothelial cells were investigated using front-surface fluorometry of fura-2 loaded strips of porcine aortic valve. Progesterone inhibited the thapsigargin induced sustained [Ca(2+)](i) elevation (IC(50)=33.9 microM, n=4), while 17beta estradiol added a transient [Ca(2+)](i) elevation. Progesterone and 17beta estradiol had no significant effect on the thapsigargin-induced [Ca(2+)](i) elevations in the absence of extracellular Ca(2+). A Mn(2+)-induced decline of fluorescent intensity at 360 nm excitation was accelerated by thapsigargin. This acceleration was completely reversed by progesterone, but not by 17beta estradiol. Progesterone inhibited, and 17beta-estradiol enhanced the endothelin-1 (ET-1)-induced [Ca(2+)](i) elevation, while both had no effect on the ET-1 induced Ca(2+) release observed in the absence of extracellular Ca(2+) or in the pertussis toxin-treated strips. Progesterone and 17beta-estradiol thus had different effects on Ca(2+) signaling, especially on Ca(2+) influx, in endothelial cells. PMID- 11118644 TI - Transrepression of NF-kappaB is not required for glucocorticoid-mediated protection of TNF-alpha-induced apoptosis on fibroblasts. AB - The cellular resistance to tumor necrosis factor (TNF) of most cell types has been attributed to both a protective pathway induced by this cytokine and the preexistence of protective factors in the target cell. NF-kappaB has been postulated as one of the principal factors involved in antiapoptotic gene expression control on TNF-resistant cells. We have previously shown that glucocorticoids protect the naturally TNF-sensitive L-929 cells from apoptosis. Here we analyze the role of NF-kappaB and glucocorticoids on TNF-induced apoptosis in L-929 cells. We found that inhibition of NF-kappaB enhanced the sensitivity to TNF-induced apoptosis. Glucocorticoids inhibited NF-kappaB transactivation via IkappaB induction. Moreover, glucocorticoids protected from TNF-induced apoptosis even when NF-kappaB activity was inhibited by stable or transient expression of the superrepressor IkappaB. These results demonstrate that although glucocorticoids inhibit NF-kappaB transactivation in these cells, this is not required for their protection from TNF-induced apoptosis. PMID- 11118645 TI - Expression and intracellular localization of leptin receptor long isoform-GFP chimera. AB - The leptin receptor (OBR) and its ligand leptin (OB) are key players in the regulation of body weight. The OBR is a member of the class I cytokine receptor family and is alternatively spliced into at least six different isoforms. The multiple forms are identical in their extracellular and transmembrane regions but differ in lengths. The two predominant isoforms include a long form (OBR(l)) with an intracellular domain of 303 amino acids and a shorter form (OBR(s)) with an intracellular domain of 34 amino acids. We have constructed a recombinant OBR(l) chimera with the green fluorescent protein (GFP) by fusing GFP to the C-terminus of the OBR(l). The OBR(l)-GFP chimera was transiently transfected and expressed in SHSY5Y and HEK293 cells. In a STAT-Luciferase assay we show that the GFP moiety in this chimera did not affect the signalling capacity of OBR(l)-GFP. In both SHSY5Y and HEK293 cells transfected with OBR(l)-GFP, a predominant intracellular green OBR(l)-GFP fluorescence was detected in vesicles also positive for internalized fluorophore conjugated leptin. We also found that treatment with the lysosomotropic reagent monensin did not relocalize OBR(l)-GFP together with the human transferrin receptor in recycling endosomes, indicating OBR(l)-GFP not to participate in this pathway. In biotinylation-streptavidin pulse chase experiments, using antibodies raised against GFP and OBR, we observed that the rate of early appearance of OBR(s) at the cell surface, upon leptin stimulation, was faster than that found for OBR(l)-GFP. Taken together, our results provide novel data concerning the intracellular trafficking of the two different isoforms of the leptin receptor. PMID- 11118646 TI - Efficient transformation of Dictyostelium discoideum with a particle inflow gun. AB - We report experiments to transform Dictyostelium discoideum using a simple home made particle gun. Stable transformants were obtained at frequencies of up to 2500 clones/microg DNA. This is five times more than we achieve with the same vector using electroporation protocols. We also show that the particle inflow gun can be used for analysis of developmentally regulated gene expression in a transient assay. PMID- 11118647 TI - S-Allylmercaptoglutathione: the reaction product of allicin with glutathione possesses SH-modifying and antioxidant properties. AB - The reaction between allicin (diallylthiosulfinate), the active component of garlic and reduced glutathione was investigated. The product of this reaction, mixed disulfide S-allylmercaptoglutathione (GSSA) was separated by high performance liquid chromatography and identified by 1H and (13)C nuclear magnetic resonance and mass spectroscopy. The reaction is fast (with an apparent bimolecular reaction rate constant of 3.0 M(-1) s(-1)). It is pH-dependent, which reveals a direct correlation to the actual concentration of mercaptide ion (GS( )). Both GSSA and S-allylmercaptocysteine (prepared from allicin and cysteine) reacted with SH-containing enzymes, papain and alcohol dehydrogenase from Thermoanaerobium brockii yielding the corresponding S-allylmercapto proteins, and caused inactivation of the enzymes. The activity was restored with dithiothreitol or 2-mercaptoethanol. In addition, GSSA also exhibited high antioxidant properties. It showed significant inhibition of the reaction between OH radicals and the spin trap 5,5'-dimethyl-1-pyroline N-oxide in the Fenton system as well as in the UV photolysis of H2O2. In ex vivo experiments done with fetal brain slices under iron-induced oxidative stress, GSSA significantly lowered the production levels of lipid peroxides. The similar activity of GSSA and allicin as SH-modifiers and antioxidants suggests that the thioallyl moiety has a key role in the biological activity of allicin and its derivatives. PMID- 11118648 TI - A genetic interaction between a ubiquitin-like protein and ubiquitin-mediated proteolysis in Dictyostelium discoideum(1). AB - A ubiquitination factor, NosA, is essential for cellular differentiation in Dictyostelium discoideum. In the absence of nosA, development is blocked, resulting in a developmental arrest at the tight-aggregate stage, when cells differentiate into two precursor cell types, prespore and prestalk cells. Development is restored when a second gene, encoding the ubiquitin-like protein SonA, is inactivated in nosA-mutant cells. SonA has homology over its entire length to Dsk2 from Saccharomyces cerevisiae, a ubiquitin-like protein that is involved in the assembly of the spindle pole body. Dsk2 and SonA are both stable proteins that do not seem to be subjected to degradation via the ubiquitin pathway. SonA does not become ubiquitinated and the intracellular levels of SonA are not affected by the absence of NosA. The high degree of suppression suggests that SonA rescues most or all of the defects caused by the absence of nosA. We propose that NosA and SonA act in concert to control the activity of a developmental regulator that must be deactivated for cells to cross a developmental boundary. PMID- 11118649 TI - Molecular cloning and expression profile of Xenopus calcineurin A subunit(1). AB - We have cloned a cDNA encoding a catalytic subunit of calcineurin (CnA) expressed in Xenopus oocytes. The deduced amino acid sequence indicates 96.3% and 96.8% identities with the mouse and human CnAalpha isoforms, respectively. Xenopus CnA (XCnA) RNA and protein are expressed as maternal and throughout development. Recombinant XCnA protein interacted with calmodulin in the presence of Ca(2+). Deletion of calmodulin binding domain and auto-inhibitory domain revealed calcium independent phosphatase activity, thereby showing that XCnA is likely to be modulated by both calmodulin and calcium. PMID- 11118650 TI - Status of methods for assessing bacterial cell surface charge properties based on zeta potential measurements. AB - Surface interfacial physiology is particularly important to unicellular organisms with regard to maintenance of optimal cell function. Bacterial cell surfaces possess net negative electrostatic charge by virtue of ionized phosphoryl and carboxylate substituents on outer cell envelope macromolecules which are exposed to the extracellular environment. The degree of peripheral electronegativity influences overall cell surface polarity and can be assessed on the basis of zeta potential which is most often determined by estimating the electrophoretic mobility of cells in an electric field. The purpose of this review is to provide bacteriologists with assistance as they seek to better understand available instrumentation and fundamental principles concerning the estimation of zeta potential as it relates to bacterial surface physiology. PMID- 11118651 TI - A convenient fluorometric method for the detection of extracellular N acetylglucosaminidase production by filamentous fungi. AB - A rapid and convenient method for the detection of chitinases accumulating in filamentous fungal cultures was developed. The assay is performed on cultures growing in microtiter plates, with a fluorogenic substrate: 4-methylumbelliferyl N-acetyl-D-glucosaminide (4-MeUNAG). The fluorescence of the product, 4 methylumbelliferone, was detected. This method was successfully used to follow induction and repression of extracellular exochitinase activity in the biocontrol fungus Trichoderma harzianum. PMID- 11118652 TI - Lactate dehydrogenase release assay from Vero cells to distinguish verotoxin producing Escherichia coli from non-verotoxin producing strains. AB - The Vero cell assay presently used for virulence testing of verotoxigenic Escherichia coli (VTEC) requires at least 48-96 h where cytotoxicity effects are examined under a microscope. Here, a complimentary rapid assay was developed that measures endogenous lactate dehydrogenase (LDH) release from Vero or HEp-2 cells as an indicator of cytotoxicity. Toxin preparations from 24 VTEC strains induced 36-89% LDH from Vero cells and 15-62% LDH from HEp-2 cells in 12-16 h. A verotoxin-positive but enterohemolysin negative strain also showed a similar cytotoxicity effect. In contrast, three VT-negative strains caused only 13-16% LDH from Vero cells and 1-7% LDH from HEp-2 cells. Five presumptive E. coli isolates from naturally contaminated food and clinical sources did not induce significant LDH release from either cell lines. PCR analysis confirmed the presence of vt1 or vt2 genes in E. coli showing positive LDH values. Similarly, RiboPrinter analysis confirmed and identified the test strains as E. coli except for two meat isolates, which were identified as Hafnia alvei. Cytopathic effects of toxin preparations from VTEC revealed severe lysis, vacuole formation and death in Vero cells and multiple vacuoles and cell elongation in HEp-2 cells. The colorimetric cytotoxicity assay described here can provide quantitative data for determining the virulence potential of verotoxigenic E. coli in 12-16 h. PMID- 11118653 TI - Comparison of maximum specific growth rates and lag times estimated from absorbance and viable count data by different mathematical models. AB - Maximum specific growth rate (mu(max)) and lag time (lambda) were estimated from viable count and absorbance data and compared for different microorganisms, incubation systems and growth conditions. Data from 176 growth curves and 120 absorbance detection times of serially diluted cultures were evaluated using different mathematical growth models. Accurate estimates of mu(max) and lambda were obtained from individual absorbance growth curves by using the Richard model, with values of the parameter m fixed to 0.5, 1.0 or 2.0 to describing different degrees of growth dampening, as well as from absorbance detection times of serially diluted cultures. It is suggested to apply the two techniques complementarily for accurate, rapid and inexpensive estimation of microbial growth parameter values from absorbance data. In contrast, considerable limitations were demonstrated for the ability of the Exponential, the Gompertz and the Logistic models to estimate mu(max) and lambda values accurately from absorbance data. Limitations of these models were revealed due the wide range of growth conditions studies. PMID- 11118654 TI - Structural diversity of microorganisms in chemically perturbed soil assessed by molecular and cytochemical approaches. AB - Until recently, our understanding of microbial community development in soil ecosystems exposed to different inorganic and organic pollutants has been limited to culturable microorganisms because of the techniques available. The discovery that most soil microorganisms are non-culturable but potentially viable and metabolically active accelerated the application of different culture-independent methods for structural diversity assessments of the microbial community. This review examines the results of recent studies on the impact of heavy metals and organic pollutants on the diversity of the microflora obtained with methods based on analyses of signature biomarkers such as nucleic acids and fatty acids. The application of these techniques allowed researchers to pinpoint reduction of microbial diversity in contaminated soil, and significant shifts in the community structure, leading to the dominance of only a few populations (species) and the disappearance of others, some of which were never isolated by conventional methods (e.g. an increase in Acidobacterium or a decrease in terrestrial non thermophilic Crenarchaeota). Although the new techniques are not free from limitations, they allow the monitoring of the virtual impact of stressors on soil microorganisms and the direction of resuscitation of the microbial community during natural or induced bioremediation, especially when using combined approaches. PMID- 11118655 TI - Method for enumeration of 5-cyano-2,3-ditoyl tetrazolium chloride (CTC)-active cells and cell-specific CTC activity of benthic bacteria in riverine, estuarine and coastal sediments. AB - Bacteria are the most abundant and active organisms in marine sediments and are critical for nutrient cycling and as a food source to many benthic and pelagic organisms. Bacteria are found both as free-living cells and as particle associated cells, which can make investigations of these communities difficult. We found that common procedures for extracting bacteria from sediments leave the bacteria clay particle-associated and the clay particles clump, which reduce the reproducibility of direct counts. We optimized a sonication/surfactant method that produces a homogeneous suspension of bacterial cells against a uniform background of clay particles, which results in reproducible samples for epifluorescence microscopy. We developed a method to estimate CTC-positive cells and cell-specific CTC content in intact cores of surficial sediment communities from riverine, estuarine and coastal sites. Benthic bacterial abundances averaged 4.9x10(8) cells/g dry wt sediments in Apalachicola River, Florida sediments, 4.9 13.8x10(9) cells/g dry wt sediments in a variety of Apalachicola Bay sediments and 3.6x10(8) cells/g dry weight in shallow, anoxic Gulf of Mexico sediments. Percent CTC-positive cells ranged from low values of 9-10% CTC-positive cells in Apalachicola River and Apalachicola Bay sediments to high values of 25% CTC positive cells in anoxic Gulf of Mexico sediments. After correction for abiotic CTC reduction and chlorophyll interference, estimates of cell-specific CTC reduction ranged from 0.15 to 0.55 fmol CTC(red)/active cell in the Apalachicola Bay sediments to 1.6 to 3.8 fmol CTC(red)/active cell in anoxic Gulf of Mexico sediments. PMID- 11118656 TI - Infection of Vero cells with Coxiella burnetii phase II: relative intracellular bacterial load and distribution estimated by confocal laser scanning microscopy and morphometry. AB - Coxiella burnetii, the agent of Q fever in man and of coxiellosis in other species, is an intracellular pathogen not yet grown axenically. Confocal laser fluorescence microscopy and morphometry were used to measure relative C. burnetii phase II loads and their intracellular distribution in aldehyde fixed and DAPI stained Vero cell monolayers. The fluorescence of single horizontal optical sections provided useful information on relative loads of bacteria in cells and vacuoles. The relative density of the bacteria in the vacuoles was inferred from ratios of fluorescence to vacuolar section areas. Relative bacterial loads, bacterial densities and section areas of large vacuoles increased exponentially between days 2 and 4 of the infection of gamma-irradiated host cells, stabilized between days 4 and 6, and decreased thereafter. Estimated minimum doubling times were higher for the overall complement of the intracellular organisms (about 12 h) than for bacteria that were confined to larger vacuoles (about 10 h). PMID- 11118657 TI - Development of the Specific and Random Amplification (SARA)-PCR for both species identification of enterococci and detection of the vanA gene. AB - A rapid and reliable polymerase chain reaction (Specific and Random Amplification (SARA)-PCR) has been developed to identify enterococcal species and to detect the vanA gene in one single reaction. This technique was based on the use of the primer set previously designed to amplify a part of the vanA gene (Dutka-Malen et al., J. Clin. Microbiol., 33 (1995) 24-27). In the chosen low stringency conditions complex patterns were obtained, with a sharp band of approximately 700 bp in cases where the vanA gene was present. Discrimination at the species and strain level was achieved by applying the SARA-PCR assay to a collection of 55 enterococcal isolates and type strains. Simultaneously the vanA gene was detected in all strains showing high resistance to vancomycin. PMID- 11118658 TI - Can the API (RAPID) Coryne system be used for identification of rapidly growing mycobacteria? AB - Fifty-six strains of rapidly growing mycobacteria (RGM) and 14 strains of aerobic actinomycetes as quality controls (QC) were tested in the API (RAPID) Coryne system version 2. Both groups yielded codes with low identification scores, considerable overlaps, and similar diagnoses. No species-specific codes were observed. Thus, the system would not be useful for the identification of RGM. PMID- 11118659 TI - Effects of milking frequency on lying down and getting up behaviour in dairy cows. AB - The objective of this study was to investigate if cows milked twice per day have more difficulty lying down and getting up and spend less time lying than cows milked three times per day. Seventeen cows of the Swedish Red and White Cattle Breed were studied, seven of which were milked twice daily (2M) and ten were milked three times (3M) daily. They were kept in individual cubicles, that were closed in the rear end with a rope. They had free access to a mixture of silage, hay and concentrate. The individual cows were video-recorded for 24h every fourth week, starting four weeks after calving for four times. The 2M cows stood significantly longer, 128.11min, than the 3M cows, 64.88min, (P<0.01) during the 4h before morning milking. The 2M cows also had a tendency for longer duration of standing rumination (P=0.059) as well as significantly more bouts of standing rumination (P<0.01) during these hours than the 3M cows. The cows in the 3M group spent less time on the getting up movement (P<0.05) during the 4h before morning milking. The distribution of the lying bouts during these 4h differed significantly between the groups, where the 3M cows had fewer lying bouts shorter than 15min and more lying bouts longer than 90min. The results indicate that milking three times a day contributes to increased comfort in high-producing dairy cows. PMID- 11118660 TI - Consistency of side choice in the milking parlour by Holstein-Friesian cows and its relationship with their reactivity and milk yield. AB - Dairy cows often have to choose which of two sides to enter in the milking parlour. Some cows are very consistent in this choice, and it is common to assume that when these cows are more disturbed are being milked in their non-preferred side. Such disturbance might involve significantly poor welfare. In order to assess this assumption, we decided to study the behaviour and milk yield of dairy cows and their relationships with side preference in the milking parlour. The study was carried out at Cambridge University Farm, in a two-sided tandem milking parlour. The data collection followed the daily management routine. We recorded the side chosen by each cow (left or right) during 40 milking sessions. Data from 70 cows, which were present in at least 25 milking sessions (mode=36), were included in the statistical analysis. Cows' reactivity (CR) during premilking udder preparation, time spent fitting the milking cluster (FT), milk yield (MY) and duration of milking (DM) were measured. There was evident individual variation in the consistency of side choice. Individual differences (ANOVA, P<0.001) were also found in CR, FT, MY and DM; although these variables were not significantly affected by the side or the interaction animalxside (ANOVA, P>0.05). The comparison between left and right side means (paired t-test) of these variables did not show significant differences (P>0.05). We concluded that there is no evidence that the cows were discomforted or stressed when milked in the non-preferred side of the milking parlour. PMID- 11118661 TI - Reactions of calves to handling depend on housing condition and previous experience with humans. AB - This study investigated the influence of stockperson's behaviour and housing conditions on calves' behavioural reactions to people, and behavioural and physiological reactions to handling and short transport. Sixty-four Finnish Ayrshire male calves were used; half of them were housed in individual pens, the other half were housed in group pens of two calves. In both housing conditions half of the calves received minimal contact from the stockperson, while the other half were stroked on their necks and shoulders for 90s a day, after milk meals. The effects of housing and contact with the stockperson on the responses of calves to people, either entering or approaching the pen, were studied. Furthermore, calves' behavioural and physiological (cortisol, heart rate) reactions to being loaded onto a truck, transported for 30min and unloaded were observed. When a person entered the home pen, calves housed by pairs took significantly more time to interact and interacted less frequently with the person than individually housed calves did (p<0.01). Calves that received additional contact interacted for longer time with the unfamiliar person than calves with minimal contact (p=0.02). When a person approached the front of the calves' pens, less withdrawal responses were shown by calves that had received additional contact (p<0.05) than those that had received minimal contact. When the calves were loaded onto the truck, it took more time and effort to load pair housed calves than individually housed calves (p<0.01) and less effort to load calves that had received additional contact (p<0.01) compared to those that had received minimal contact. During loading additional contact calves had lower heart rates (p<0.05) than those that had received minimal contact, while during transport pair housed calves had lower heart rates compared to individually housed ones (p<0.05). For all the observations performed, no interactions were found between housing conditions and human contact.It is concluded that, compared to calves housed individually, calves housed in pairs are less ready to approach humans and less easy to handle. Providing calves with regular positive contacts makes them less fearful of people and improves handling. Due to the greater difficulty in handling calves housed in groups, it is concluded that these animals need to have regular contact with humans. PMID- 11118662 TI - Behavioural and physiological consequences of acute social defeat in growing gilts: effects of the social environment. AB - Endocrine, behavioural and immunologic processes, together with body growth, were evaluated in gilts that were defeated at 10 weeks of age in resident-intruder tests. Immediately after defeat, gilts were either separated from or reunited with a familiar conspecific (litter-mate; always a barrow). Gilts were assigned to one of four treatments: (a) DI: defeat, followed by isolation (separation from original litter-mate; n=8); (b) I: no defeat, isolation (control group; n=9); (c) DP; defeat, followed by pair-housing (reunion with original litter-mate; n=8); and (d) P: no defeat, pair-housing (control group; n=8). The following general conclusions were derived: (1) social defeat caused pronounced short-term elevations in hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal medullary activities, and of prolactin levels. Moreover, as soon as 1h after defeat, percentages of blood lymphocytes and neutrophilic granulocytes were, respectively, decreased and increased; (2) social defeat had some long-lasting influence on behaviour and physiology, but isolation predominantly determined responses in the longer term. Defeat, as well as isolation, resulted in increased cardiovascular activities compared to P controls, as observed in a novel object test (NOT: +7 days) and an aversion test (AVT: +14 days). Moreover, defeated as well as isolated gilts did not habituate to a repeated novel environment test (NET: -7, +2 and +7 days) in terms of frequencies of vocalising, whereas P controls did. Isolation, through the separation from any other pig, was responsible for the other observed long-term characteristics, which developed progressively. Isolated gilts showed high mobilities and high cortisol responses in the repeated NET (+7 days), not being habituated. This contrasted the reactions of pair-housed gilts, which were much reduced. In addition to their high cardiovascular activities in the NOT and the AVT, isolated gilts also displayed higher heart rates in the repeated NET and during human presence following the NOT, compared to pair-housed gilts. Finally, isolated gilts were more inhibited to approach a novel object (in the NOT) than pair-housed pigs; and (3) stress responses of defeated gilts were modulated by the subsequent social environment. Stimulation of the HPA-axis (plasma- and salivary cortisol) was prolonged in those defeated gilts which were isolated (observed in the first hour). Changes in leucocyte subsets were still observed after 3 days in DI, but were 'normalised' within 1 day in DP gilts. Two days after defeat, habituation to the repeated NET in terms of mobility and salivary cortisol responses occurred in control and DP gilts, but not in DI gilts. We argue that these effects of the social environment shortly after defeat were related to a stress-reducing effect of a stable social relationship, i.e. social support. PMID- 11118663 TI - Variation in behavioural responses of ewes towards predator-related stimuli. AB - Thirty-two groups of six sheep, classified into three breed categories according to their weight class (L: light, one breed (n=7); M: medium light, two breeds (n=10); H: heavy, three breeds (n=15)) were tested for antipredatory behaviour towards seven stimulus regimes. Tests were performed on 2-5-years-old ewes with lambs inside standardised and fenced home pastures. Stimulus regimes were: man in rain poncho, trolley, ball on trolley (blind stimuli), stuffed wolverine on trolley, stuffed lynx on trolley, stuffed bear on trolley, and man in rain poncho with a dog on leash (carnivore stimuli). The L breed showed the longest recovery time, the longest flight distance and the tightest flocking behaviour. Significant regressions indicate that there were negative linear relationships between sheep weight and recovery time and between sheep weight and flight distance. Carnivore stimuli caused longer recovery times (P<0.001) and longer flight distances (P<0.001) than the blind stimuli. Our results confirm the hypothesis that lighter sheep breeds display stronger antipredatory reactions than heavier breeds. PMID- 11118664 TI - Note on a method for individual recognition in feed pecking in free running groups of hens. AB - A system for automatic registration and individual recognition of feed pecking (activity and quantity) in groups of free running hens was tested. A PIT (Passive Integrated Transponder)-tag system was used to separate and register individuals when they were feeding. An electronic balance system placed under the feeder registered the amount eaten by each individual on a PC. A test with two different feed stuffs; oat and wheat was performed on three hens during a total of 6 weeks. PMID- 11118665 TI - A rapid hemolysis assay for the detection of sodium channel-specific marine toxins. AB - Current methods of detection for fish and shellfish biotoxins in monitoring and research purposes are either labor intensive, expensive, require specialized techniques or all of the above. This paper reports on the development of a fairly sensitive, rapid, and inexpensive assay which detects the presence of compounds that affect the sodium channel. It is based on the principles of the mouse neuroblastoma tissue culture assay for sodium channel specific-biotoxins using red blood cells (RBCs) from the red tilapia (Sarotherodon mossambicus). This assay has the potential to complement the use of live animal bioassay testing for marine toxins. Veratridine, a sodium channel activator and ouabain, an inhibitor of Na(+)/K(+) ATPase, both react with the tilapia RBCs by affecting the permeability of the cell's membrane. Saxitoxin (STX), its analogs, and tetrodotoxin (TTX) can inhibit the action of veratridine and ouabain leaving the cell morphologically normal. By sequencing the addition of veratridine and ouabain, with either the extracted samples, saxitoxin, tetrodotoxin, or ciguatoxin (CTX-a sodium channel activator) to the RBCs a sodium channel antagonist or activator can be detected. Results using pure concentrations of a sodium channel-specific toxin could be detected to inhibit hemolysis at a concentration of 0.3 microg/ml STX, 3.5 microg/ml for neo-STX, 3.0 microg/ml for GTX, and 5.0 microgl for TTX in the presence of ouabain and veratridine. CTX was detected at a concentration of 50 microg/ml. The RBCs from the red tilapia was used due to the fish's ability to osmoregulate its internal environment to survive in both fresh and saltwater. In addition, with growing opposition to live animal testing, this assay has been designed as a non-lethal means of testing for sodium channel affecting marine toxins. No test animals are sacrificed and blood may be drawn from the same fish for continued sample testing. PMID- 11118666 TI - Proteoglycans synthesized by cultured bovine aortic smooth muscle cells after exposure to lead: lead selectively inhibits the synthesis of versican, a large chondroitin sulfate proteoglycan. AB - To investigate the effects of lead on the formation of extracellular matrix in the vascular wall, we characterized proteoglycans synthesized by cultured vascular smooth muscle cells after exposure to the metal by biochemical techniques. Confluent cultures of bovine aortic smooth muscle cells were metabolically labeled with [(35)S]sulfate or [(35)S]methionine/cysteine in the presence of lead nitrate. The amount of glycosaminoglycans (GAGs) was evaluated by the incorporation of [(35)S]sulfate into GAGs by the cetylpyridinium chloride precipitation method. The labeled proteoglycans were characterized by DEAE Sephacel ion exchange chromatography and Sepharose CL-2B molecular sieve chromatography. The GAG M(r) and composition were analyzed by Sepharose CL-6B chromatography, and the core protein M(r) was analyzed by SDS-polyacrylamide gel electrophoresis, before and after digestion with papain or chondroitin ABC lyase. Lead significantly decreased the [(35)S]sulfate incorporation into GAGs accumulated in the cell layer and the conditioned medium. [(35)S]Sulfate-labeled proteoglycans obtained from the cell layer and the conditioned medium were separated into three peaks on DEAE-Sephacel chromatography and only the peak with the highest charge density was decreased by lead. The highly charged peak was eluted near the void volume on Sepharose CL-2B molecular sieve chromatography and sensitive to chondroitin ABC lyase on Sepharose CL-6B chromatography, indicating that lead selectively inhibits the synthesis of large and highly charged chondroitin/dermatan sulfate proteoglycans (CS/DSPGs). However, the size of chondroitin/dermatan sulfate chains of the CS/DSPGs was M(r) approximately 47000 in both the control and lead-treated cultures. On the other hand, lead decreased the accumulation of a large CS/DSPG with a core protein of approximately 450 kDa in the cell layer and the conditioned medium; the core protein was identified as versican core by Western blot analysis. It is therefore suggested that lead inhibits the synthesis of the versican core protein in vascular smooth muscle cells without a change in length of chondroitin/dermatan sulfate side chains. As a result, versican-poor extracellular matrix would be induced by lead in vascular smooth muscle cells. PMID- 11118667 TI - The protective effect of flavonol quercetin against ultraviolet a induced oxidative stress in rats. AB - Ultraviolet A (UVA) light exposed cells can induce the production of reactive oxygen species (ROS) which can damage the cellular elements. Antioxidants can interfere with the production of ROS. In this study, malondialdehyde (MDA), reduced glutathione (GSH), glutathione reductase (GSSGR), glutathione peroxidase (GPx), catalase (CAT) and superoxide dismutase (SOD) levels were measured in the liver of rats exposed to UVA light in various doses. The effects of quercetin were determined as antioxidant on those parameters. Rats were divided into three groups as control, ultraviolet (UV), and ultraviolet+quercetin (UV+Q). UV and UV+Q group rats were irradiated 4 h per day with UVA light (1.25 mW/cm(2)) during periods of 0,3,6,9 days. Thus, on days 0,3,6 and 9, the rats have received 0,54,108,162 W/cm(2) light, respectively. Quercetin (50 mg/kg body wt.) was administered intraperitoneally before each irradiation period in the UV+Q group rats. MDA level in the UV group increased significantly on day-9 when compared to the control group (P<0.05). The MDA levels in the UV+Q group decreased significantly on day-6 and 9 in comparison with the UV group (P<0.05, P<0.001, respectively). GSH levels in all groups were not significantly different. GSSGR and GPx activities in the UV group were significantly lower on day-6 and 9 than in the control group (P<0.001). On all days these enzyme activities in the UV+Q group were significantly higher than in the UV group and higher than in the control group (P<0.001). SOD and CAT activities in the UV group decreased significantly on day-3, 6, and 9 in comparison with the control group (P<0.001). These enzyme activities also increased significantly in the UV+Q group on all days when compared to the UV group (P<0.001). This study demonstrated that the exposure of rats to UVA led to oxidative stress as reflected by increased MDA levels and reduced enzymic antioxidant levels, quercetin may be useful by reducing or preventing photobiologic damage. PMID- 11118668 TI - In vitro toxicology in hepatocyte bioreactors-extracellular acidification rate (EAR) in a target cell line indicates hepato-activated transformation of substrates. AB - In this article we introduce an in vitro model for hepato-mediated toxicity testing consisting of a Hepatocyte-Bioreactor connected to a microphysiometer system for monitoring of the extracellular acidification rate (EAR) of cells. The EAR in this system represented the metabolic activity of a tested cell line under the influence of bioreactor supernatant. Cyclophosphamide (CYCL), a well-known hepato-activated cytostatic drug was used as a model substrate because of its widespread clinical use. The predrug CYCL needed CYP 450 dependent activation to its active cytotoxic metabolite 4-OH cyclophosphamide. Primary pig hepatocytes from slaughterhouse organs were cultured in a collagen sandwich configuration in specially designed flasks and after 3 days introduced into a 50 ml recirculating perfusion system including 30 microg/ml CYCL. In a parallel open circuit, this bioreactor was connected to three perfusion chambers of a microphysiometer system housing 1.5 x 10(5) ZR 751 cells (breast tumor cell line). Bioreactor supernatant including CYCL was pumped at 150 microl/min into the microphysiometer. The recorded EARs under CYCL influence were correlated to controls, which were set to be 100%. After 1 and 7 h of bioreactor supernatant perfusion, including activated CYCL, the ZR 751 cell line showed an EAR of 98.99%+/-3.15 (mean+/-SD) and 81. 32%+/-10.18 (P<0.05), respectively, as compared to controls (bioreactor supernatant from the identical set-up without CYCL). The inactivated predrug CYCL showed no effect on the EAR: Perfusion of medium with 30 microg/ml CYCL alone, excluding the bioreactor activation, resulted in an EAR of 100. 11%+/-4.74 (mean+/-SD) after 7 h. Thus the presented model of hepato-activated toxicity showed an EAR decrease in the ZR 751 cell line that reflected the toxic activation of the predrug by the bioreactor. PMID- 11118669 TI - Changes in ceramide and sphingomyelin following fludarabine treatment of human chronic B-cell leukemia cells. AB - Fludarabine is used to treat chronic lymphocytic leukemia. Both in vitro and in vivo studies have indicated that apoptosis is an important mode of fludarabine induced cell death. However, the apoptotic pathways activated are not known. The effects of apoptotic doses of fludarabine on sphingomyelin, ceramide and the production of reactive oxygen species were investigated in the chronic B-cell leukemia lines WSU and JVM-2. Apoptosis, as assessed by an increase in phosphatidylserine externalization, internucleosomal DNA fragmentation and caspase-3-like activity, was evident by 18 h after fludarabine in both cell lines. The general caspase inhibitor t-butoxycarbonyl-Asp(OMe)-fluoromethyl ketone (OMe, methyl ester) significantly inhibited apoptosis supporting a role for caspases in fludarabine-induced cell death. A 2.5- to threefold elevation in ceramide levels was observed 6 h after fludarabine treatment. Concomitantly, a decrease in sphingomyelin levels was observed. Fumonisin B1 (an inhibitor of ceramide synthase) pretreatment significantly prevented fludarabine-induced ceramide generation and apoptosis. Conversely, C6-ceramide induced apoptosis in both cell lines. No effect of fludarabine on indices of oxidative stress (dichlorofluorescin oxidation and glutathione disulfide formation) were detected, although partial protection from apoptosis, and prevention of ceramide generation and caspase-3 activation, were achieved with N-acetylcysteine. These findings are consistent with the involvement of caspases and ceramide in fludarabine-induced apoptosis in WSU and JVM-2 cells. Oxidative stress does not appear to be induced by fludarabine, although the protective effects of N-acetylcysteine suggest that thiol redox balance may play a role in the apoptotic pathway. PMID- 11118670 TI - Purification and characterization of diquat (1,1'-ethylene-2, 2'-dipyridylium)- metabolizing enzyme from paraquat-resistant rat liver cytosol. AB - To establish a paraquat-resistant Wistar rat strain, we carried out continuous sister-brother mating among rats that survived high-dose intraperitoneal administration of paraquat dichloride (360 mg/kg). The percentages of paraquat resistant rats among wild rats and among the fifth-generations were 7.1% and 20.6%, respectively. After high-dose paraquat administration, the serum paraquat concentration in sensitive rats was much higher than that in paraquat-resistant rats. The cytosol fraction of liver from paraquat-resistant rats had higher paraquat- and diquat-metabolizing activities than that of liver from paraquat sensitive rats. By contrast, microsomal fractions from livers of paraquat resistant and paraquat-sensitive rats had no paraquat- or diquat-metabolizing activity. This paraquat/diquat-metabolizing enzyme was partially purified from paraquat-resistant rat liver cytosol using affinity chromatography for diquat. At the end of the purification procedure, rat liver diquat-metabolizing enzyme was purified 1154-fold to a final specific activity of 32.32 mol/h/mg protein, and an overall recovery of about 0.46% was obtained. This enzyme oxidized diquat to diquat-dipyridone during overnight incubation at 37 degrees C, but only metabolized traces of paraquat. The molecular mass of the enzyme was estimated as 190 kDa, and its isoelectric point of it was 4.6-4.7. Kinetic study revealed the values of K(m) and V(max) to be 35.0 micromol/l and 0.81 micromol/h/ml, respectively. PMID- 11118671 TI - The influence of anticholinergic drug selection on the efficacy of antidotal treatment of soman-poisoned rats. AB - The influence of some anticholinergic drugs (atropine, benactyzine, biperiden, scopolamine) on the efficacy of antidotal treatment to eliminate soman (O pinacolyl methylphosphonofluoridate)-induced disturbance of respiration and circulation and to protect experimental animals poisoned with supralethal dose of soman (1.5 x LD(50)) was investigated in a rat model with on-line monitoring of respiratory and circulatory parameters. While the oxime HI-6 in combination with atropine prevented soman-induced changes in monitored physiological parameters insufficiently and very shortly, the combination of HI-6 with benactyzine or biperiden is able to prevent soman-induced alteration of respiration and circulation much more longer. Nevertheless, only rats treated with HI-6 in combination with scopolamine were fully protected against the lethal toxic effects of soman within 2 h following soman challenge. Our findings confirm that anticholinergic drugs with the strong central antimuscarinic activity, such as benactyzine, biperiden and especially scopolamine, seem to be more effective adjuncts to HI-6 treatment of severe acute soman-induced poisoning than atropine. PMID- 11118672 TI - The roles of glutathione and antioxidant enzymes in menadione-induced oxidative stress. AB - We investigated the role of glutathione (GSH) and antioxidant enzymes in menadione-resistance by using K300 cells (menadione-resistant cells) and parental P19 cells (menadione-sensitive cells). We found that acquisition of resistance was associated with elevations in glutathione content and DT-diaphorase activity. The activity of glutathione S-transferase (GST) was significantly decreased, while the activities of glutathione peroxidase, glutathione reductase, catalase, and superoxide dismutase in K300 cells were maintained at the same levels as compared to the parental P19 cells. Using reactive oxygen species (ROS)-sensitive fluorescence dye 2,7- dichlorodihydrofluorescein diacetate (DCFH/DA), we demonstrated that K300 cells are characterized by reduced cellular ROS as compared to the parental P19 cells during menadione's action. Menadione depleted glutathione to a small extent in the K300 cells, but a rapid depletion was observed in P19 cells. Pretreatment of K300 cells with dicumarol, a DT-diaphorase inhibitor, or buthionine sulfoximine (BSO), an inhibitor of gamma-glutamyl cysteine synthase, sensitized the cells to menadione. BSO treatment was less effective than dicumarol treatment in reversing menadione resistance in K300 cells. These results strongly support the belief that DT-diaphorase plays a central role in protecting cells against menadione-induced oxidative stress by decreasing the ROS formation. PMID- 11118673 TI - Carbon tetrachloride is immunosuppressive and decreases host resistance to Listeria monocytogenes and Streptococcus pneumoniae in female B6C3F1 mice. AB - Carbon tetrachloride (CCl(4)) is an environmental contaminant that has been detected in ambient air, seawater, surface-water and snow. The immunotoxic potential of CCl(4) was evaluated in female B6C3F1 mice. The animals were administered with CCl(4) daily for 14 days at doses of 50, 100, 500 or 1000 mg/kg body weight by gavage with corn oil as a vehicle. Exposure to CCl(4) resulted in an increase of liver weight but not the body weight and the weights of brain, spleen, lungs, thymus and kidneys. Exposure to CCl(4) produced minimal effect on differential hematological parameters; however, it produced a significant increase in serum glutamic-pyruvic transaminase (SGPT) levels in all dose groups while other serum chemistries showed sporadic increases, primarily at the dose level of 1000 mg/kg. Exposure to CCl(4) produced a decreased humoral immune response; the IgM antibody forming cell (AFC) response to sheep red blood cells (sRBC) was suppressed with the maximal decrease (45%) observed at the dose level of 1000 mg/kg. The IgM serum titer to sRBC was also reduced with a maximal decrease (54%) observed at the dose level of 500 mg/kg. Although exposure to CCl(4) had no effects on the mixed leukocyte response (MLR), cytotoxic T lymphocyte activity and natural killer (NK) cell activity, a decrease in both the absolute number and the percentage of CD4(+)CD8(-) at the dose level of 500 mg/kg was observed. The functional activity of the mononuclear phagocyte system was compromised as reflected by a decrease in the vascular clearance of (51)Cr-sRBC and a decrease in the uptake of (51)Cr-sRBC by the liver. Finally, in the two host resistance models evaluated, exposure to CCl(4) decreased host resistance to both Streptococcus pneumoniae and Listeria monocytogenes with greater susceptibility to the latter. Overall, these studies demonstrate that CCl(4) was immunosuppressive in female B6C3F1 mice. PMID- 11118674 TI - Changes in endogenous Zn and Cu distribution in different cytosolic protein fractions in mouse liver after administration of a single sublethal dose of CdCl(2). AB - The time course of change in tissue Cd, Cu and Zn contents, their distribution in cellular protein fractions as well as the profile of MT gene expression in mouse liver was described over a 7 days period following a single intraperitoneal injection of 2 mg/kg of CdCl(2). The result showed that Cd accumulated rapidly in mouse liver. Between 1 h and 7 days after administration, over 18% of the total Cd administered were found in the liver. Cd administration was also associated with the overexpression of the MT-mRNA. However, the time course of induction was not parallel to the change in tissue Cd content. When separated on a Sephadex G 75 column, majority of Cd was found to bind to the fractions known to contain the metal-binding protein, metallothionein (MT). From day 2 after Cd administration, a small amount of the metal was also found associated with the high molecular weight (HMW) proteins. In addition to Cd, tissue Zn content was affected most during the entire study. There was a significant decrease in tissue Zn content during the initial 8 h but tissue Zn content increased significantly throughout the following 6 days. At 1-7 days, majority of Zn was associated with the HMW protein fraction. Although there was no significant change in total tissue Cu content, distribution of Cu in different protein fractions was detected. While in control animals, Cu was mainly associated with the HMW proteins, some was found in the MT fraction on the second day. On the 7th day, Cu distribution had deteriorated. Together with changes seen in Cd, the results might suggest that injury had occurred in the tissue at this time. The results of the present study showed that Cd caused a change in subcellular distribution of tissue endogenous metals, which might reflect alteration of cellular functional activities. PMID- 11118675 TI - Cytochrome P450 reductase-mediated anaerobic biotransformation of ethanol to 1 hydroxyethyl-free radicals and acetaldehyde. AB - The ability of cytochrome P450 reductase to metabolize ethanol (EtOH) to acetaldehyde (AC) and 1-hydroxyethyl free radicals (1HEt) in anaerobic media was studied. Determination of AC was made by GC-FID analysis of the head space of incubation mixtures. The formation of 1HEt was established by GC-MS analysis of the adduct formed between the radical and the spin trap PBN. Results showed that pure human P450 reductase is able to biotransform EtOH to AC and 1HEt in a NADPH dependent process under an oxygen-free nitrogen atmosphere. Pure FAD in the presence of NADPH was also able to generate AC and 1HEt from the alcohol. Anaerobic incubation mixtures containing either rat liver microsomes or pure nuclei were also able to biotransform EtOH to AC and 1HEt in the presence of NADPH. These processes were inhibited by antibody against rat liver microsomal P450 reductase. Results suggest that semiquinone forms of the flavin in P450 reductase may biotransform EtOH. These reactions might be of some significance in tissues where the P450 reductase is present in the absence of specific forms of cytochrome P450 known to be involved in EtOH metabolism (e.g. CYP2E1). However the toxicological significance of this enzymatic process remains to be established. PMID- 11118676 TI - The effects of perinatal exposure to low doses of 2,3,7,8-tetrachlorodibenzo-p dioxin on immune organs in rats. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is revealed to exert diverse biological effects including immunotoxicity, mainly by inadvertently activating the transcription factor arylhydrocarbon receptor (AhR). In the present study, the developmental effects of perinatal exposure to low doses of TCDD on the major immune organs of offspring, thymus and spleen, were investigated focusing on weaning time (postnatal day (PND) 21), puberty (PND 49) and adulthood (PND 120) in male rats. Concurrently, TCDD contents in those organs were measured with a high-resolution gas chromatography-mass spectrometry (GC/MS). In the thymus and spleen, CYP1A1 mRNA induction, the sensitive reaction caused by activation of AhR, was also measured in order to examine whether perinatally administered TCDD can elicit gene expressions in these organs. When pregnant dams were administered a single oral dose of 12.5-800 ng TCDD/kg body weight on gestation day (GD) 15, the weights of the thymus and spleen of the offspring did not differ from those of control animals throughout the experiments. The thymus and spleen maternally exposed to 800 ng TCDD/kg contained 102.0 and 62.7 pg TCDD/g tissue on PND 21, respectively, and the amounts decreased thereafter. In the thymus, dose-dependent CYP1A1 mRNA induction was clearly observed by maternal exposure to 50-800 ng TCDD/kg on PND 5. The induction was gradually decreased on PND 21 and 49. On the other hand, CYP1A1 mRNA induction in the spleen was very weak. In these thymi, no reproducible change was observed by TCDD exposure in cell number and cellular population defined by CD4 and CD8 molecules at any time. In contrast, splenocyte number was shown to decrease by maternal exposure to 12.5-800 ng TCDD/kg in a dose-dependent manner on PND 49. The alteration in spleen cellularity by TCDD was not detected on PND 21 or 120. These results clarified that perinatal exposure to low doses of TCDD affects the immune organs, which is apparent in spleen around puberty and likely to be hardly relevant to AhR-dependent gene expressions. PMID- 11118677 TI - Effects of attention and semantic relation on event-related potentials in a picture-word naming task. AB - The interactive effects of attention and semantic relation on event-related potential (ERP) waveforms were examined during a naming task in which the physical stimulus array was the same across conditions. Superimposed picture-word pairs were presented in which the meaning of the words and pictures was congruent, semantically associated, or incongruent. In separate conditions, participants named words or pictures while ignoring the other stimulus. When words were attended, the superimposed pictures modulated the amplitude of P240 waves at posterior sites compared with when words were presented individually, but had no effect on N450 waves. In contrast, when pictures were attended to, the superimposed incongruent words elicited larger amplitude N450 waves at anterior sites than did congruent words or individually presented pictures. These effects affirm the independent processing of words and pictures during attention and are consistent with automatic and controlled processing of words and pictures, respectively. They also illustrate the endogenous nature of these late ERP waves. PMID- 11118678 TI - Costs or benefits of emotional conditioning on cognitive processing? AB - We report on two experiments using a transfer-of-control procedure in evaluating costs or benefits of emotionally relevant stimuli on the processing of a cognitive task. In differential conditioning the impact of Pavlovian conditioning usually is examined by contrasting instrumental responses in the presence of an excitatory conditioned stimulus (CS+) and an inhibitory conditioned stimulus (CS ). We expanded this comparison by introducing a neutral third condition. This additional neutral condition served as a within subject control for evaluating whether the CS+ increased responding and/or the CS- decreased responding. Both experiments yielded the same result; the CS- slowed down cognitive processing while the CS+ had no impact. Thus, the proposed transfer-of-control procedure may serve as a reliable and valid research tool in the evaluation of motivation and emotion in humans. PMID- 11118679 TI - Increases in total respiratory resistance during forehead temperature stimulation. AB - We investigated the effects of forehead temperature stimulation on total respiratory resistance in healthy individuals. In two experiments involving a total of 38 participants we studied the time course and stability of the response at moderate temperature (20-23 degrees C). Small plastic bags filled with water were positioned on the forehead for a duration of 60 s. Oscillatory resistance (R(os)), heart period (HP), respiratory sinus arrhythmia (RSA), and ventilatory parameters were measured continuously. Experiment 1 showed similar phasic increases in R(os) during the first 20 s of stimulation with moderate (20-23 degrees C) as compared to cold (8-11 degrees C) temperature. Phasic increases by moderate temperature were replicated in Experiment 2 over five successive stimulation trials. Within-session stability of individual differences in response was only modest. Ventilatory adjustments did not facilitate the phasic R(os) increases. As increases were mainly due to the inspiratory component of R(os), a substantial contribution of upper airway artifacts was less likely. Increases in HP were the most pronounced responses to all stimulation conditions, while RSA did not increase significantly. PMID- 11118680 TI - Correlations between ERP parameters and intelligence: a reconsideration. AB - The present study investigated differences in ERP parameters related to intelligence. For that purpose 74 individuals (Intelligence: M=107; S.D.=12; range 73-135), of average creativity passively listened to two tones and performed two auditory, and two visual oddball tasks while their EEG was recorded. The approximate entropy parameters, peak latencies and amplitudes were determined. The correlation coefficients indicated that in the attended conditions, the more intelligent individuals showed more regular ERP waveforms than less intelligent individuals. It was further found that less intelligent individuals showed increased P300 latencies and reduced amplitudes. The differences were explained with a more specific engagement of neural networks in more intelligent individuals. PMID- 11118681 TI - REACTPOOL: a code implementing a new multi-compound pool model that accounts for chemical reactions and changing composition for spills of water reactive chemicals. AB - All chemicals that react violently with water or in contact with water liberate toxic gas are included in the list of substances covered by the majority of the international legislation on major hazards. This category includes a large number of chemicals that are used widely in the process industries. A survey of accidents that occurred in the last 10 years in the USA shows numerous major incidents that involved spillages of these substances. Even so, there are almost no experimental data on the behaviour of these chemicals on release. Furthermore, there are very few published studies on modelling the behaviour of such spillages, except in the case of hydrogen fluoride. In previous work we reported a new theoretical model [J. Haz. Mat. 62 (1998) 101-129, J. Haz. Mat. 62 (1998) 131-142, J. Haz. Mat. A67 (1999) 9-40], that describes accidental spills of SO(3) and oleum, which are substances with very complex behaviour that belong to this category. It describes both the pool [J. Haz. Mat. 62 (1998) 101-129, J. Haz. Mat. 62 (1998) 131-142] and the cloud behaviour [J. Haz. Mat. A67 (1999) 9-40]. In the work reported here the pool model was modified in a generic form in order to include other water reactive chemicals. REACTPOOL is a new code that can be used for both instantaneous and continuous liquid releases under a wide range of input parameters (steady or varying). It can be used for all liquids irrespective of their volatility and reactivity, and it describes pools consisting of more than one liquid that can have changing composition and properties. The purpose of this paper is to present the general procedure followed in REACTPOOL and to show how the new model has been modified and implemented for substances other than SO(3) and oleum. The modelling procedure has been implemented in a computer code written in Visual Basic, and results of the model have been generated using this code. It should be noted that this model requires validation data, but that the availability of such data awaits the performance of suitable experimental investigations. PMID- 11118682 TI - Spill behaviour using REACTPOOL. Part I. Results for accidental releases of chlorosulphonic acid (HSO(3)Cl). AB - Chlorosulphonic acid is a toxic, highly reactive and corrosive substance that exists in its liquid form at ambient conditions. Its major hazardous potential comes from the clouds of hydrogen chloride and sulphuric acid mist produced whenever this chemical escapes from containment and is exposed to moisture. It decomposes violently and sometimes explosively in the presence of water, liberating heat. On spillage it creates liquid pools that can either boil or evaporate. There are three sources of water available for reaction: free ground water, substrate water and atmospheric moisture. Hydrogen chloride gas or aqueous solution and sulphuric acid liquid are produced by the hydrolysis reaction. This paper describes the dangers involved in cases of accidental releases of chlorosulphonic acid, referring to its properties, toxicity data and mitigation tests. It also reports results of pool behaviour using REACTPOOL [T. Kapias, R.F. Griffiths, C. Stefanidis, J. Haz. Mat., submitted for publication]. These results indicate that the pool behaviour is governed mainly by the amount of water available for reaction. Surface roughness and wind speed also have a significant effect on the results. A discussion of the results in comparison with those for other water reactive substances is presented in Part III of this series of papers [T. Kapias, R.F. Griffiths, J. Haz. Mat., submitted for publication]. The generated cloud will initially contain chlorosulphonic acid, hydrogen chloride and sulphuric acid with numerous processes taking place. Initially, it is usually denser than air. Although chlorosulphonic acid has been involved in major hazard incidents, there are no experimental data relevant to the modelling requirements. Use of REACTPOOL provides insights into the major hazard role of this substance. PMID- 11118683 TI - Effective design of greenbelts using mathematical models. AB - Trees, shrubs, and other vegetation can absorb and assimilate certain air pollutants if the pollutants are present within tolerable levels. This concept is being increasingly used in developing strips of vegetation, often called 'greenbelts' around sources of pollution. But several intricacies are associated with the exercise of effective and optimal designing of greenbelts. The pattern of dispersion of air pollutants, as effected by the density of the gaseous plume and the meteorology of the area, must be studied with great precision because these aspects would determine the location and the geometry of the greenbelt. The species composition in the greenbelt should confirm to the pollutants to be attenuated as to the geoclimatic conditions of the region. Decisions on the tree heights, and the sequence of plantation of trees and other vegetation also similarly require complex inputs. In this paper, the authors have addressed these issues and have presented a set of mathematical models, which may help in the rational and optimal design of greenbelts. PMID- 11118684 TI - Studies on energetic compounds. Part 16. Chemistry and decomposition mechanisms of 5-nitro-2,4-dihydro-3H-1,2,4-triazole-3-one (NTO). AB - The present review deals with the chemistry and thermolysis of NTO and plausible decomposition pathways have been described. The decomposition of 5-nitro-2,4 dihydro-3H-1,2,4-triazole-3-one (NTO) induced by X-ray, UV, laser, photochemical irradiation has also been discussed. High-speed photographic studies of the impact responses of NTO are also included. The thermal decomposition of labelled NTO has also been described here. Methods of detection as well as safe disposal of NTO have also been mentioned. PMID- 11118685 TI - Removal of heavy metals from sandy soil using CEHIXM process. AB - This paper presents the results of a new process of soil decontamination termed as couple electric-hydraulic gradient assisted by ion exchange medium (CEHIXM) in which a hydraulic gradient is coupled with electric gradient in a non conventional arrangement for the purpose of electrolysis and ion transport. A suitable ion exchange medium was used to capture and subsequently recover the cations. In this process, heavy metals no longer precipitated in the treated soil near the pretreatment zone. The cations were transported out of the soil and captured in the ion exchange medium. Several laboratory tests were conducted to examine the removal of Pb, Cd, Zn and Mn from sandy soils. The experimental results demonstrated the removal efficiencies more than 97% for all the four heavy metals at an energy expenditure of 345 kWh/m(3) of soil. PMID- 11118686 TI - Catalytic hydrodechlorination of tetrachloroethylene over red mud. AB - Hydrodechlorination of tetrachloroethylene was investigated using red mud (RM, a by-product in the production of alumina by the Bayer process) as the catalyst. Use of RM as a hydrodechlorination catalyst is of interest from an industrial point of view because its cost is much lower than that of commercial catalysts. Hydrodechlorination reactions were carried out in a continuous fixed bed reactor. The influence of catalyst sulfiding, temperature (50-350 degrees C), pressure (2 10MPa), hydrogen flow rate and the presence of solvents (hexane, heptane, benzene and toluene) on the reaction was studied. Sulfided red mud is active as a hydrodechlorination catalyst, conversion of tetrachloroethylene increases as the pressure and temperature increase. The solvents did not influence the conversion, nor were side reactions involving the solvent observed. The kinetics of the reaction was studied at 350 degrees C and 10MPa, conditions for which mass transfer limitations were negligible. A good fit of a Langmuir-Hinselwood model to the experimental data was obtained. PMID- 11118687 TI - Pressure-drops control strategy in a fixed-bed reactor. AB - This paper presents a strategy to control pressure-drops (head loss) in a biofilter designed according to the "Mist-Foam" concept. This concept is based on the mixing of the gaseous substrate and a liquid nutrient solution with an atomization nozzle to generate a mist passing subsequently through a synthetic polyurethane foam. In this type of bioreactor, the microbial growth reduces progressively the empty bed volume of the biofilter and causes an increase in the pressure-drops. This phenomenon can result in a complete clogging of the biofilter. The strategy of pressure-drops control presented here consists of successive interruption of the liquid flow, automatically controlled, resulting in a drying effect of the biomass. Tested during a 160 days experiment, this system has permitted to reduce and stabilize the pressure-drops in a biofilter in which the carrier exhibited a high likelihood of clogging. PMID- 11118688 TI - Evaluation of the models available for the prediction of pressure drop in venturi scrubbers. AB - The major running cost derived from the operation of venturi scrubbers is pressure drop. In the present study, the predictions of different models are compared to experimental data from venturi scrubbers of different sizes (throat diameter from 1.9 to 16cm), geometries, operating variables and liquid injection arrangements. As a result, it is concluded that most of the models must be used with caution. Much attention must be paid to the validity of the assumptions employed in the mathematical models. The equations proposed by Calvert [Scrubbing, Air Pollution, 3rd Edition, Vol. IV, Academic Press, New York, 1982], Yung et al. [JAPCA 27 (1977) 348] or Hesketh [Atomization and cloud behaviour in wet scrubbers, in: Proceedings of the US-USSR Symposium Control Fine Particulate Emissions 1974, San Francisco, 15-18 January 1974] produce good results only in very specific situations. The model proposed by Boll [Ind. Eng. Chem. Fundam. 12 (1973) 40] is simple, easy to compute and agrees reasonably well with the experimental data. Unfortunately, it cannot predict the effect of different liquid injection arrangements. The model by Azzopardi and coworkers [Filtr. Sep. 21 (1984) 196; Trans. IchemE. 69B (1991) 237; Chem Eng. J. 67 (1997) 9] was the only one to give good predictions for all the range of variables studied. On the other hand, this model is not simple and requires from the engineer an additional effort in terms of computation. In order to apply this model to the rectangular geometry, the concept of hydraulic equivalent diameter was used. PMID- 11118689 TI - Field demonstration of pervaporation for the separation of volatile organic compounds from a surfactant-based soil remediation fluid. AB - As part of a Department of Defense project, the US Environmental Protection Agency was responsible for designing, building and field operating a pilot-scale pervaporation unit. The field site was an active dry cleaning facility on the grounds of Marine Corps Base Camp Lejeune in Jacksonville, NC. The overall goal of the project was to remove tetrachloroethylene (PCE) from the soil beneath the dry cleaning shop using a surfactant-based soil remediation fluid and to recycle/reuse the surfactant. In order to reinject the recovered surfactant, the pervaporation unit was required to achieve an average 95% removal of contaminants from the extracted fluid over the duration of the test period. PCE removal averaged 95.8% during peak surfactant levels and exceeded 99.9% in the absence of surfactant, thereby meeting the reinjection requirement. Removal of a group of secondary contaminants at the site, termed Varsol compounds, was monitored via concentrations of three Varsol marker compounds: decane, undecane and 1,3,5 trimethylbenzene. The pervaporation system processed 100,000 gal of groundwater and surfactant solution over a period of 70 days. In order to evaluate and validate process performance, a variety of process variables and properties were monitored over the course of the demonstration. Pervaporation costs are projected to be on the order of $20 per 1000 gal of surfactant solution treated for a moderate size system (10 gpm). PMID- 11118690 TI - Enhanced abilities of highly swollen chitosan beads for color removal and tyrosinase immobilization. AB - The enhancement of abilities for the removal of reactive dyes and immobilization of tyrosinase onto highly swollen chitosan beads was demonstrated compared to the use of common chitosan flakes. Chitosan was prepared from natural cuttlebone wastes. It was shown that the adsorption capacity of dyes at 30 degrees C using swollen chitosan beads was around five times greater than that using common chitosan flakes. The adsorption of dyes using swollen beads was faster by 10-40% depending on the types of dyes. Finally, the capacity of tyrosinase immobilization onto swollen beads was about 14 times greater than chitosan flakes, which was reflected by the higher yield of 3,4-dihydroxyphenylalanine from tyrosine and ascorbic acid in the heterogeneous catalytic system. PMID- 11118691 TI - The effect of flow-through leaching on the diffusivity of heavy metals in stabilized/solidified wastes. AB - The flow-through leaching test is a test method employed to study the leaching behavior of monolithic stabilized/solidified (S/S) hazardous wastes under the condition that the leachate flows through the sample. This method simulates the leaching process of the S/S hazardous waste disposed under a particular landfill condition when the S/S waste is more permeable than its surrounding materials or when the deterioration of the solidified waste form has reached a state that ground water can flow-through the waste via the porosity system of the S/S waste matrix. This paper describes a study on the long-term performance of the cement based S/S heavy metal wastes using a flow-through leaching test method. Two series of leaching tests with different synthetic heavy metal waste samples were carried out. The S/S samples were made from five types of heavy metals with two kinds of binders. The metals were Pb(2+), Zn(2+), Cu(2+), Ni(2+) (positive ions as nitrates), and Cr(6+) (a negative ion as potassium dichromate), and the binders were type I ordinary portland cement (OPC) and pulverised fuel ash (PFA). The model developed by Godbee and Joy for simulating the leaching behavior was modified to estimate the diffusivity parameter in this study. The results obtained indicate that since the matrix of the solidified waste in a flow-through leaching tests is always being degraded, the values of diffusivities increase continuously during the leaching period. The diffusivity variation range was from 10(-13) to 10(-3)cm(2)/s, and were normally higher than those obtained from other test methods such as ANS 16.1 test and other dynamic leaching tests. PMID- 11118692 TI - The influence of different soil constituents on the reaction kinetics of wet oxidation of the creosote compound quinoline. AB - Creosote contaminated sites have become a widespread problem in industrialized countries. Recently, wet oxidation using high temperature, pressure, water and oxygen followed by activated sludge treatment proved to be an efficient method for removing a wide selection of creosote compounds in contaminated soils. Wet oxidation of the creosote compound quinoline was carried out in the presence of montmorillionite, quartz and humic acid. The products derived from wet oxidation were identified and treated biologically by activated sludge testing their biodegradability. The influence on the oxidation kinetics of quinoline during wet oxidation was pH dependent. Humic acid supported the oxidation of quinoline, whereas the addition of montmorillionite and quartz had either an inhibiting effect or led only to a slight increase in oxidation. In mixtures of soil constituents, especially at low contents of humic acid, the adsorption of quinoline on montmorillionite prevented oxidation at neutral pH. Thus, alkaline extraction of both quinoline and humic acid was needed for an efficient oxidation. A proposed reaction mechanism suggests that quinoline was oxidized by hydroxyl radicals formed during the oxidation of the humic acid. A wide selection of reaction products (mainly carboxylic acids, benzene and pyridine derivatives) derived from the wet oxidation of humic acid and quinoline. The reaction products from humic acid degradation had a rate limiting effect on the wet oxidation of quinoline leaving small residues of quinoline after the treatment. On the contrary, these reaction products also improved the biodegradation of products from the quinoline oxidation due to co-digestion of carboxylic acids. Therefore, the presence of soil components (mainly humic acid) improved the combined wet oxidation and biological activated sludge treatment of quinoline. PMID- 11118708 TI - Introduction. PMID- 11118709 TI - Epidemiology of paratuberculosis in wild ruminants studied by restriction fragment length polymorphism in the Czech Republic during the period 1995-1998. AB - In two studies carried out during the period 1995-1998, paratuberculosis was diagnosed in domestic and wild ruminants in the Czech Republic. The isolated Mycobacterium avium subspecies paratuberculosis strains were analysed by standardised restriction fragment length polymorphism (RFLP) [Pavlik, I., Horvathova, A., Dvorska, L., Bartl, J., Svastova, P., du Maine, R., Rychlik, I., 1999. J. Microbiol. Methods 38, 155-167]. In December 1992, 19 late pregnant Charolais heifers were imported to the Czech Republic from Hungary (original import from France to Hungary). One 11-month-old heifer roamed in the wild in a range of approximately 15-20km for 7 months from November 1993 to May 1994. Upon capture, the animal showed clinical signs of paratuberculosis (emaciation and diarrhoea). Seven other animals from the same herd were infected with the identical RFLP type B-C1 of M. paratuberculosis. During the period 1995-1996, samples were taken and examined from the small intestine and corresponding lymph nodes of 84 wild ruminants: 19 red deers (Cervus elaphus) and 65 roe-deers (Capreolus capreolus). These wild ruminants originated from 44 different locations within the same district from as the infected escaped heifer. Five M. paratuberculosis strains were isolated: one strain of RFLP type B-C1 from a stag and three strains of RFLP type B-C1 and one strain of RFLP type B-C9 from roe deer. The three wild ruminants (one stag and two roe-deer) infected with the same RFLP type B-C1 were detected in the same area as the heifer, suggesting that this was the likely infection source. However, the infection source of the roe-deer infected with strain of RFLP type B-C9 was obviously different, and the stags that escaped from the farm were purchased from an area infected with this RFLP type. In the second study carried out during 1997-1998 in the whole Czech Republic (divided into 76 districts), 718 wild ruminants were examined from 90% of the districts. M. paratuberculosis was isolated from 25 (3.5%) animals from the wild, from farms and from game parks: 7.1% of 132 red deers, 1.5% of 336 roe deers, 3.9% of 178 fallow deers (Dama dama), and 4.2% of 48 moufflons (Ovis musimon). This study discovered three RFLP types (B-C1, D-C12 and M-C16). A surprising finding was that of M. paratuberculosis (RFLP type B-C1) infection in roe-deer and a fallow deer in their natural habitat. The infection source was determined to have originated from two imported Holstein and Limousine cattle herds infected with the same strain. In the case of a mother and daughter roe deer infected with RFLP type M-C16 and a fallow deer infected with RFLP type D C12, all roaming in their natural habitat, the infection source was not discovered. The highest incidence of clinically ill wild ruminants was found in farmed red deer, and no relationship was found between the RFLP type or ruminant species and clinical status of animal. PMID- 11118710 TI - Prevalence and regional distribution of paratuberculosis in dairy herds in The Netherlands. AB - In the Netherlands a survey was conducted to estimate the prevalence of paratuberculosis in dairy herds. In total 15822 cows of at least 3 years of age, belonging to 378 herds were tested using an absorbed ELISA. Of these herds, 55% (n=207) had one or more serologically positive cows. Of the positive non vaccinated herds, most had one (n=98) or two (n=49) positive cows. The percentage positive cows per herd was 2.5+/-3.2%.The true prevalences on cow and herd levels, based on a test sensitivity that ranged from 0.3 to 0.4 and a specificity that ranged from 0.985 and 0.995, were estimated at 2. 7-6.9% and 31-71%. Seven herds had been vaccinated against paratuberculosis and these herds had a significantly higher percentage of serologically positive cows (23%) than the non vaccinated herds (2.5%). In conclusion, a small percentage of the dairy cows and a high percentage of the dairy herds in the Netherlands is serologically positive. The percentages true infected cows and herds are difficult to estimate precisely due to uncertainties in test sensitivity and specificity. PMID- 11118711 TI - Paratuberculosis in Iceland: epidemiology and control measures, past and present. AB - Paratuberculosis as well as the slow virus infections maedi/visna and jaagsiekte came to Iceland in 1933 when 20 sheep of the Karakul breed were imported from Halle, Germany. At least five of these sheep were subclinical carriers of paratuberculosis. Within 16 years paratuberculosis together with the other Karakul diseases (maedi/visna and jaagsiekte) almost ruined sheep farming, the main agricultural industry in Iceland. The first clinical case of paratuberculosis in sheep was confirmed in 1938, and in cattle in 1944. The first cattle cases of paratuberculosis appeared on farms where the disease had been prevalent in sheep for years. The virulence in cattle appeared to be considerably lower than in sheep. Extensive measures were used to control the spread of paratuberculosis in sheep. Hundreds of kilometres of fences were put up and used together with natural geographic borders to restrict the movement of sheep from infected areas. Serological and other immunological tests were also used to detect and dispose of infected individuals. These measures proved inadequate and the disease could not be eradicated. Culling and restocking of uninfected sheep in endemic areas eradicated maedi/visna and jaagsiekte but not paratuberculosis. Experiments showed that vaccination against paratuberculosis could reduce mortality in sheep by 94%. Vaccination of sheep in endemic areas has been compulsory in Iceland since 1966 and as a result losses have been reduced considerably. Today, serology is used to detect and control infection in cattle herds. Furthermore, serology is used to control vaccination of sheep and screen for infection in non-endemic areas. The complement fixation (CF) test for paratuberculosis has been used until now, but recently we have started comparing the CF test with the CSL absorbed ELISA test. PMID- 11118712 TI - Prevalence of paratuberculosis (Johne's disease) in the Belgian cattle population. AB - The national bovine paratuberculosis (PTB) seroprevalence (apparent prevalence) in the Belgian cattle population was determined by a serological survey that was conducted from December 1997 to March 1998. In a random sample of herds (N=556, 9.5%), all adult cattle of 24 months of age or older (N=13,317, 0.4%) were tested for the presence of antibodies using a commercially available absorbed ELISA test kit. The PTB median within-herd seroprevalence (proportion of detected animals within the seropositive herds) and the PTB individual-animal seroprevalence (proportion of detected animals) were, respectively, 2.9% (quartiles=1.6-5.6) and 0.87% (95% confidence interval (CI)=0.71-1.03). The PTB herd seroprevalence (proportion of detected herds) was 18% (95% CI=14-21). Assuming a test sensitivity and specificity of 45 and 99% [Sweeney et al., 1995. J. Vet. Diagn. Invest. 7 (4), 488; Sockett et al., 1992. J. Clin. Microbiol. 30 (5), 1134], respectively, the median true within-herd prevalence and the true individual animal were estimated to be 7 and 2%, respectively. The true herd prevalence of Mycobacterium paratuberculosis infection was first estimated according to currently accepted methodology. This calculation revealed that the specificity of the used test has a dramatic effect on the estimation; assuming a test sensitivity of 45% and a true within-herd prevalence of 7%, the true herd prevalence estimation decreased from 36 to 0.8% if the test specificity decreased from 99. 9 to 99%, respectively. This sensitivity analysis showed that the practical limits of the accuracy of the used screening test jeopardize the estimation of the true herd prevalence within reasonable confidence limits, because the within-herd PTB true prevalence was low. For this reason we augmented the herd specificity for herds with larger adult herd size (>5). This was done by increasing the cut-off number of positive cattle required (>/=2) to classify a herd truly positive and including herds with one positive test result if there was historical evidence of PTB (previous diagnosis and/or clinical signs). This approach resulted in an estimated true herd prevalence of M. paratuberculosis infection of 6%. The true herd prevalence for dairy, mixed and beef herds was, respectively, 10, 11 and 3%. PMID- 11118713 TI - First evidence of Johne's disease in farmed red deer (Cervus elaphus) in Belgium. AB - In a deer farm, chronic diarrhoea was seen in a 4-year-old hind. This animal died in poor condition on the farm and Johne's disease was suspected. Ziehl-Neelsen staining of the faeces of this hind were positive for the presence of clumps of small acid-fast bacilli, but faecal cultures remained negative. Direct and indirect tests were performed on 24 hinds and stags (yearlings, 2- and 4-year-old animals). The indirect tests performed were serology (Mycobacterium paratuberculosis antibody ELISA, HerdChek, Idexx), comparative cervical skin test (CCT) and lymphoproliferation test (LT) using Mycobacterium bovis purified protein derivative (PPD) and Mycobacterium avium PPD as antigens. Three positive serological results, three positive CCT and eight positive LT were observed in hinds and stags older than 2 years. No positive serological results were observed in the yearling group, whereas some sensitisation was observed in the CCT as well as in the LT for the same group of animals. The degree of concordance between these indirect tests was poor. The three seropositive animals were slaughtered and subjected to post-mortem examination. Histopathology was performed on mesenteric lymph nodes and on the terminal ileum. Visual changes in some mesenteric lymph nodes were observed, no gross lesion was seen in the intestine. Although Ziehl-Neelsen staining yielded no positive results, a catarrhal focal necrotic enteritis associated with a granulomatous lymphadenitis compatible with Johne's disease was evidenced. The mycobacterial cultures on organ samples from slaughtered animals were positive after 2 months for M. avium subspecies paratuberculosis and negative for M. bovis and M. avium. This is the first description of Johne's disease in a deer farm in Belgium. PMID- 11118714 TI - Detection of Mycobacterium avium subsp. paratuberculosis in milk from clinically affected cows by PCR and culture. AB - Milk and faeces samples from cows with clinical symptoms of paratuberculosis were examined for the presence of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) by culture and PCR. M. paratuberculosis was cultivated in variable numbers from faeces or intestinal mucosa in eight of 11 animals. In milk from five cows (all faeces culture positive), we cultivated a few colonies of M. paratuberculosis (<100 CFU per ml). Milk samples from two cows were PCR positive (both animals were faeces culture positive, and one cow was milk culture positive). One cow was culture negative on intestinal mucosa, but culture positive in milk, and two cows were negative in culture and PCR from both faeces and milk. In conclusion, the presence of M. paratuberculosis could be detected in raw milk by PCR, but cultivation of milk was more sensitive. PMID- 11118715 TI - Paratuberculosis in sheep: an emerging disease in South Africa. AB - During a serological survey for ovine paratuberculosis a total of 145934 ovine serum samples from 2019 farms throughout South Africa were tested by means of the AGID assay. Fifty-two infected farms were identified in the Western Cape and Eastern Cape provinces. Links between infected farms in the two provinces were established. Examination of the distribution of infected farms in the Western Cape indicated a positive correlation between acid soils and occurrence of infection. In an attempt to increase the sensitivity and facilitate screening of large numbers of sera two commercial ELISA systems were evaluated for their potential use in a future monitoring program. Sera from histologically positive sheep and known negative sheep flocks were used. The highest sensitivity (50. 9%) was found if both ELISA systems were run concurrently and the results of both systems combined. PMID- 11118716 TI - Parallel faecal and organ Mycobacterium avium subsp. paratuberculosis culture of different productivity types of cattle. AB - Faecal (at least 3 months before slaughtering) and organ examinations were carried out in 611 animals (497 dairy, 69 dual-purpose and 44 beef cattle) originating from eight paratuberculosis infected cattle herds. The diagnosis in cattle was established by routine intestinal culture (ileum and the adjacent lymph nodes) after slaughter. In selected 132 animals, post-mortem intensive culture was performed on tissue samples collected from the gastrointestinal tract (duodenum, jejunum, ileum, ileocecal valve, caecum, rectum) and the corresponding lymph nodes, submandibular, retropharyngeal, tracheobronchial, liver and supramammary lymph nodes, kidney, liver and spleen. In 251 (41.1%) of all 611 animals, Mycobacterium avium subspecies paratuberculosis could be isolated from the faeces; in 164 (65.7%) out of 251 shedding animals the infection was detected in the ileum and adjacent lymph nodes. The detection of M. paratuberculosis by routine intestinal culture of faecal culture positive animals varied from 46.0% in animals shedding 1 CFU (colony forming unit), to 94.7% in massive shedders. On the contrary, M. paratuberculosis was detected by routine intestinal culture in 92 (25.5%) of the 360 faecal culture negative animals. Shedding animals had significantly higher (P<0.01) number of organisms in their organs than non shedding animals. During the intensive tissue cultivation from selected 132 animals, 72 (54.5%) of them were positive. For the negative animals, no significant difference was found between the detection rate in organs examined after slaughter with routine and intensive method. However, in the subgroup of tissue culture positive animals a highly significant difference (P<0.01) was found by intensive examination (83.0%) compared with the routine examination (60.4%). Out of 72 tissue culture positive animals 73.6% of them harboured M. paratuberculosis in the gastrointestinal tract, 16.7% in the gastrointestinal tract and the parenchymatous organs, tracheobronchial and mandibular lymph nodes. The rest of the 9.7% of the infection was detected in the lymph nodes of head and lungs. Our study concerning the distribution of M. paratuberculosis by intensive examinations revealed a minimum effect of breed and production type on localisation of the agent. Thus, the results suggest that in case of an active infection, M. paratuberculosis can be localised in different organs of animals irrespective of their breed or production type. PMID- 11118717 TI - The histopathologic diagnosis of subclinical Johne's disease in North American bison (Bison bison). AB - The morphologic changes of subclinical Johne's disease in North American Bison (Bison bison) are characterized by microgranulomas composed of epithelioid macrophages and individual multinucleate giant cells of Langhans'-type occasionally containing individual cytoplasmic acid-fast bacilli compatible with Mycobacterium avium paratuberculosis. The microgranulomas are best visualized in the mesenteric lymph nodes of infected subclinical animals. Macrophages that can be confused with infection-associated epithelioid macrophages in the mesenteric lymph nodes are pigment-carrying cells from the intestinal tract. Mesenteric lymph node biopsy may be a useful diagnostic tool for detection of mild subclinical infection in individual ruminants from herds of unknown infection status. The biopsy may also be useful for Johne's disease surveillance during test-and-cull programs. PMID- 11118718 TI - Modification of a bovine ELISA to detect camelid antibodies to Mycobacterium paratuberculosis. AB - Mycobacterium avium subsp. paratuberculosis infection, or Johne's disease, reportedly has a low prevalence in South American camelid populations. Recently, however, single cases in the United States as well as an outbreak of the disease in Australian alpacas (Lama pacos) have been described. To provide a rapid and cost-effective method of diagnosing Johne's disease in this species, the bovine Parachek((R)) Johne's Absorbed EIA (CSL, Vic., Australia) was modified to create a camelid-specific serum antibody assay. An anti-llama IgG conjugated to horseradish peroxidase replaced the anti-bovine immunoglobulin. Checkerboard titration of principal reagents was performed using serum from nine tissue and/or fecal culture-positive camelids. Optimal dilutions of key components were determined in order to provide clear discrimination between positive and negative controls. Completion of a kinetic assay determined the optical density at which the enzyme-substrate reaction should be stopped. A herd of 100 camelids with no history of disease or exposure to M. a. paratuberculosis, a subset of which were tissue and/or fecal culture-negative, was tested to establish a cut-off value. Sample results were expressed as a percentage of the results for control sera by transforming optical density values to ELISA values (EV%). A preliminary EV% cut off of 20 was established. Using this prototype assay, culture-positive animals showed significantly different antibody responses from culture-negative animals. These results indicate that this camelid-specific ELISA, once refined, may be a useful tool for screening camelid herds for M. a. paratuberculosis infection. PMID- 11118719 TI - Mycobacterium avium subsp. paratuberculosis infection in cattle in Austria, diagnosis with culture, PCR and ELISA. AB - Serum samples from healthy, infected (n=11) and diseased (n=2) cattle as well as positive (n=17) and negative (n=41) reference sera were tested for antibodies to Mycobacterium avium subsp. paratuberculosis with two ELISA-methods (A-ELISA, Allied Monitors, Fayette, USA; H-ELISA, Institute of Microbiology and Animal Diseases, Veterinary University Hannover). Fecal samples of these animals were examined by PCR and culture. Also field serum samples found to be positive (n=664) or inconclusive (n=1589) by A-ELISA during a survey done on 11028 cattle of 2757 farms at different districts in Austria were retested with H-ELISA (Gasteiner et al., 1999). In both ELISA-methods total agreement between antibody detection and shedding of M. avium subsp. paratuberculosis (PCR, culture) in cases of diseased animals during the testing period was found. In subclinically infected animals H-ELISA showed a better correlation with the results of PCR and fecal culture. Reference serum samples of culturally negative cattle were negative in 98% by H-ELISA and in 82% by A-ELISA, and those of positive animals were positive in 59% by H-ELISA and in 82% by A-ELISA. The 664 A-ELISA positive field serum samples were positive in 20.5%, inconclusive in 32.5% and negative in 47% by H-ELISA. A-ELISA inconclusive sera gave positive reactions by H-ELISA in 5.2%, negative in 74.8% and inconclusive results in 20%. The highest prevalence of antibodies (7.9% by A- and 2.2% by H-ELISA) against M. avium subsp. paratuberculosis were found in cattle at the age of six and seven years. Seropositive animals were found at all tested ages. The A-ELISA gave two to three times more positive reactors than the H-ELISA. Also both tests showed the highest prevalence of reagents among Holstein Friesian (6.2% by A-ELISA, 2.5% by H-ELISA) followed by other cattle breeds. Seropositive cattle were observed in all districts of Austria in 3. 3-7.1% and in 0.5-1.8% of herds according to A- and H ELISA, respectively. PMID- 11118720 TI - Automation of an absorbed enzyme immunoassay for the detection of Mycobacterium paratuberculosis antibodies for an eradication program. AB - As part of an ongoing Johne's disease eradication program by the state of Victoria, Australia, the Victorian Veterinary Pathology Services (VVPS) has been involved in testing over 150000 cattle samples per year for the presence of Johne's-specific antibodies. The Parachek kit (CSL, Victoria, Australia) has been used throughout the project. This method was automated using a Rosys 3300 and a Plato 7 (Dade/Behring Diagnostics) to pipette samples and the Behring ELISA Processor 3 (Dade/Behring Diagnostics) to wash EIA plates, add reagents, reads absorbances and perform data reduction of the results. Automation saved about 15h of labour per day, allowing one staff member to analyse up to 2000 samples per 8h shift compared to 800 samples using a manual protocol. Problems that were encountered include pipetting issues, the reading of the Paracheck endpoint, and excessive packaging of the kit. Maintenance, calibration and control of the analysers were an integral part of the process. VVPS, working in conjunction with CSL, suggested modification to the Parachek kit to make automation of the product more convenient. The approach resulted in a change in reagent volume and a reduction in packaging that reduced the cost of the test by 25%. PMID- 11118721 TI - Comparative sensitivity of various faecal culture methods and ELISA in dairy cattle herds with endemic Johne's disease. AB - In three New South Wales dairy cattle herds with endemic Johne's disease, prevalence rates by faecal culture were determined to be 12, 18 and 22%, respectively. Whole herd faecal culture was shown to detect markedly more infected cattle than whole herd testing by the EMAI absorbed ELISA, particularly in the two herds with greatest prevalence. In the three study herds, five methods for whole herd faecal culture were compared in each. These included two methods based on primary culture on Herrold's egg yolk medium with mycobactin J (HEYM): (1) conventional decontamination with sedimentation and primary culture on HEYM; (2) Whitlock decontamination and culture on HEYM. The remaining three methods were based on radiometric (BACTEC) culture: (3) decontamination and filtration to BACTEC medium; (4) modified Whitlock decontamination to BACTEC medium and (5) Whitlock decontamination to BACTEC medium. For BACTEC cultures, two methods were compared as confirmatory tests for Mycobacterium paratuberculosis: mycobactin dependence on conventional subculture to HEYM and IS900 PCR analysis of radiometric media. Among 179 cattle tested simultaneously by all five culture methods, 38 cattle were confirmed to be shedding M. paratuberculosis. In identifying shedder cattle, method 5 was the most sensitive, followed by methods 2, 4, 1, and 3 was the least sensitive. The number of BACTEC cultures confirmed by mycobactin dependence or PCR was similar. PMID- 11118722 TI - Improved detection of Mycobacterium avium subsp. paratuberculosis In milk by immunomagnetic PCR. AB - The potential use of a novel immunomagnetic PCR (IMS-PCR) technique as a rapid method to screen milk samples for the presence of Mycobacterium avium subsp. paratuberculosis (M. ptb) was assessed. Immunomagnetic separation (IMS) for M. ptb, developed at Queen's University, Belfast, was applied to milk samples prior to IS900 PCR in order to selectively concentrate any M. ptb cells present and, at the same time, separate the cells from constituents of milk likely to inhibit subsequent PCR. This increased the sensitivity of IS900 PCR. IMS-PCR sensitivity could be further increased by initial centrifugation (2500 g for 20 min) of larger volumes of milk (10 and 50 ml), and resuspension of the sediment into a 1 ml volume appropriate for IMS treatment. Following IMS, template DNA for IS900 PCR was obtained by heating the bead-cell suspension in a thermal cycler at 100 degrees C for 15 min. It was estimated that the IMS-PCR assay could detect approximately 10(3)CFU of M. ptb per 50 ml of milk (equivalent to 20 CFU/ml), whereas the minimum detection limit of direct IS900 PCR was estimated at 10(5)CFU of M. ptb per 50 ml (equivalent to 2000 CFU/ml). A blind trial was carried out in which a total of 40 spiked (10(6)CFU M. ptb) and unspiked, raw and laboratory pasteurised milk samples were independently tested by IMS-PCR and conventional IS900 PCR. IMS-PCR correctly identified 97. 5% of milk samples (sensitivity 100%, specificity 95%), including spiked milk samples before and after laboratory pasteurisation. One false positive result was obtained which may have resulted from carryover between samples during the IMS procedure. Conventional IS900 PCR correctly identified only 72.5% of the same 40 milk samples (sensitivity 23%, specificity 100%). IMS-PCR was also shown to be capable of detecting natural M. ptb infection in raw sheep's milk, and raw and commercially pasteurised cows' milk. PMID- 11118723 TI - Use of a PCR method on fecal samples for diagnosis of sheep paratuberculosis. AB - The high sensitivity of PCR compared to the difficulties of fecal culture in sheep prompted the development of PCR protocols for detection of Mycobacterium avium subsp. paratuberculosis DNA in sheep feces. Although the PCR itself is well developed, and does not pose large technical problems, concentrating the bacteria from samples that may contain low numbers of bacilli using practical methods is still the main difficulty for the use of this technique. In this study, we describe an extraction protocol for the concentration and purification of M. avium subsp. paratuberculosis DNA from fecal samples and we compare it with other methods. The diagnostic performance of the freeze-boiling method was evaluated using a reference method [Vet. Rec. 134 (1994) 95] on fecal samples from a group of selected sheep from different flocks of known individual serological, pathological, and cultural paratuberculosis status. Using, as a reference, a combination of results in those conventional methods, the freeze-boiling PCR protocol showed a sensitivity of 94.1%, and a specificity of 92.3%. PMID- 11118724 TI - ELISA and fecal culture for paratuberculosis (Johne's disease): sensitivity and specificity of each method. AB - The sensitivity and specificity of the ELISA and fecal culture tests for paratuberculosis in dairy cattle are examined. ELISA and fecal culture data from seven dairy herds where both fecal cultures and ELISA testing was done concurrently are included. A cohort of 954 cattle including 697 parturient adults, cultured every 6 months from 10 herds followed over 4 years served as the basis to determine fecal culture sensitivity. The fecal culture technique utilized a 2g sample with centrifugation and double incubation. Of the 954 cattle cohort of all ages (calf to adult) that were fecal sampled on the first herd visit, 79 were culture positive. An additional 131 animals were detected as culture positive over the next seven tests at 6-month intervals. The sensitivity of fecal culture to detect infected cattle on the first sampling was 38%. Of the 697 parturient cattle cohort, 67 were positive on the first fecal culture, while an additional 91 adult cattle were culture positive over the next seven tests, resulting in a sensitivity of 42% on the first culture of the total animals identified as culture positive. Animals culled from the herds prior to being detected as infected and animals always fecal culture negative with culture positive tissues at slaughter are not included in the calculations. Both groups of infected cattle will lower the apparent sensitivity of fecal culture. Infected dairy herds tested concurrently with both fecal culture and ELISA usually resulted in more than twofold positive animals by culture compared to ELISA. The classification of infected cattle by the extent of shedding of Mycobacterium paratuberculosis in the feces helps define the relative proportion of cattle in each group and therefore the likelihood of detection by the ELISA test. ELISA has a higher sensitivity in animals with a heavier bacterial load, i.e. high shedders (75%) compared to low shedders (15%). Repeated testing of infected herds identifies a higher proportion of low shedders which are more likely to be ELISA negative. Thus, the sensitivity of the ELISA test decreases with repeated herd testing over time, since heavy shedders will be culled first from the herds. PMID- 11118725 TI - Dutch paratuberculosis programme history, principles and development. AB - Organised disease control started in the Netherlands in the 18th century with governmental attempts to eradicate cattle plague. At the beginning of the 20th century, the dairy industry and cattle breeding organisations initiated a programme to control infectious diseases by means of a complex system of rewards and penalties. This was also the reason for establishing the Animal Health Service in Friesland in 1919. The history of programmes to control paratuberculosis in sheep, goats, and cattle in various countries is described. The vaccination of young animals seems to be an effective measure in the prevention of clinical paratuberculosis, although changes in management and hygiene practices are also important. A control programme for infectious cattle diseases has a number of phases (a lifecycle) and different components. Two components are essential for success, namely: open and regular communication with farmers, veterinary practitioners, and other people involved and a good registration and identification system for cattle, herds, and veterinary practitioners. The Dutch paratuberculosis programme has 10 herd status levels: 5 10 for non-suspect herds and 1-4 for infected herds or herds of unknown status. The higher the status, the greater the chance that a herd is free of paratuberculosis. An outline is given of the Dutch paratuberculosis programme including its objectives, basic principles for eradication, communication plan, legal action, logistic considerations, and complementary research programme. PMID- 11118726 TI - Progress in national control and assurance programs for ovine Johne's disease in Australia. AB - Since the detection of ovine Johne's disease in Australia in 1980, 578 flocks have been diagnosed as infected, with 442 of these still infected. The disease was initially believed to be confined to the central tablelands area of NSW, but has subsequently been shown to be more widely distributed. Sheep strains of M. paratuberculosis are known to infect sheep and goats in south-eastern Australia. Although sheep strains have recently been identified in some cattle in Australia, epidemiological evidence to date supports the distinction between ovine Johne's disease, caused by sheep strains in sheep and goats, and bovine Johne's disease, caused by cattle strains in cattle, goats and alpaca, as a basis for control and eradication strategies. Four national initiatives to control and better understand OJD are outlined. The Australian Johne's Disease Market Assurance Program for sheep was launched in May 1997. By December 1998, 548 flocks had achieved an assessed negative status. Three flocks assigned a flock status have subsequently been found to be infected. National standards for State control of Johne's disease through zoning, movement controls and procedures in infected and suspect flocks have also been developed. In addition, a $40.1 m National Ovine Johne's Disease Control and Evaluation Program was agreed to in August 1998, and is currently being implemented. It is jointly funded by National and State industries, and Commonwealth and State governments. Its objectives are to deliver, through research and surveillance, a solid basis for a future decision on the most appropriate course for dealing with OJD and to maintain control of OJD nationally. PMID- 11118727 TI - Herd testing to control bovine Johne's disease. AB - In 1996, the cattle industries and government in the Australian state of Victoria established a Johne's disease (JD) test and control program under which participating farmers are provided with an annual ELISA test of their adult herd and advice on disease control that is tailored to their farm. The program is delivered through private veterinarians under contract with the government. There are over 600 herds enrolled in the program and about one third of these have had three or more whole herd tests. This paper provides a review of the program to date. It describes changes in ELISA reactor rates and numbers of clinical cases, and provides evidence for progress in the program. PMID- 11118728 TI - Compliance of Victorian dairy farmers with current calf rearing recommendations for control of Johne's disease. AB - Questionnaires were posted to 800 randomly selected registered Victorian dairy farmers in 1996. Five hundred and thirty-four responses were received and analysed. Johne's disease (JD) had been diagnosed on the farm of 13.2% of respondents in the last 5 years. JD was rated second only to neonatal diarrhoea in importance as a disease of calves, even though other diseases occurred more frequently. However, there was a low level of compliance with JD control recommendations by the respondents. There was no significant difference in the number of JD control recommendations adopted by farmers between the three major Victorian regions. There was a significant difference in compliance between farms having had a diagnosed case of JD and those that had not. Although there is awareness among dairy farmers of the importance of JD, there appears to be a poor implementation of measures by farmers to prevent the spread of the disease. Current JD control recommendations and the method of information transfer to Victorian dairy farms should be reassessed to ensure that dairy heifers are reared with minimal risk of transmission of JD. PMID- 11118729 TI - Progress in national control and assurance programs for bovine Johne's disease in Australia. AB - Cattle strains of Mycobacterium paratuberculosis are known to infect cattle, goats and alpaca in southeastern Australia, where there are also significant numbers of farmed deer. Although sheep strains have recently been identified in some cattle in Australia, epidemiological evidence to date supports the distinction (between bovine Johne's disease (JD), caused by cattle strains in cattle, goats and alpaca, and ovine JD, caused by sheep strains in sheep and goats) for the purposes of control and assurance programs. The National Johne's Disease Control Program is coordinated by the Australian Animal Health Council, working with the livestock industries and with the Commonwealth, state and territory governments. The council also brokers industry and government funding for the program. The National Johne's Disease Market Assurance Program for Cattle was launched in 1996 as the first of a suite of voluntary national market assurance programs (MAPs) to assess and certify herds as negative for JD. By December 1998, over 550 herds had achieved an assessed negative status. A MAP was also launched for alpaca in 1998 and a program for goats should be finalized in early 1999. National standards for state control of JD through zoning, movement controls and procedures in infected and suspect herds have also been developed. The paper covers factors affecting development and implementation, uptake of and improvements to national control and assurance programs for bovine JD in Australia. PMID- 11118730 TI - Further studies on the GS element. A novel mycobacterial insertion sequence (IS1612), inserted into an acetylase gene (mpa) in Mycobacterium avium subsp. silvaticum but not in Mycobacterium avium subsp. paratuberculosis. AB - We have recently described the GS element, found in Mycobacterium avium subsp. paratuberculosis (MAP), Mycobacterium avium subsp. silvaticum (MAS) and some isolates of Mycobacterium avium subsp. avium serotype 2 (MAAs2), which contains a set of genes of low GC% content, putatively associated with the biosynthesis, modification and transference of fucose to cell wall glycopeptidolipids. Here we describe a further gene of low GC% content (mpa), within the GS element in MAP. mpa is a putative acetyltransferase with homology to genes directly responsible for host specificity and virulence in Salmonella typhimurium and Shigella flexneri. Unlike other GS genes, strong homologues of mpa have not been found in related species, including Mycobacterium tuberculosis (MTB). In MAP, mpa encodes an ORF of 445aa, however, in MAS and MAAs2 mpa contains a single inserted copy of a novel insertion sequence. This element (IS1612) has two sets of inverted repeats at each terminus and encodes two ORFs with good homologies to transposase and helper proteins of IS21 (E. coli) and IS1415 (R. erythropolis). Sequence comparisons between mpa in MAP and MAS indicate the target site for IS1612 is duplicated on insertion to give a direct repeat at each end of the element. Immediately, downstream of the mpa gene in both MAP and MAS are a group of three genes with good homology to the daunorubicin resistance cluster. This cluster has a high GC% content which suggests a 'border' for the GS element. A short motif present at the beginning of this cluster matches with an inverted repeat of this motif at the beginning of the first gene in the GS element. This encapsulates the whole of this group of low GC% genes in MAP and further suggests its cassette like nature. Homologues of the GS element in MTB show a marked similarity of organisation, suggesting a parallel role for these genes in both pathogens. PMID- 11118731 TI - Transitions in immune responses to Mycobacterium paratuberculosis. AB - The host immune response to infection with Mycobacterium paratuberculosis is paradoxical, with strong cell-mediated immune responses during the early, subclinical stages of infection and strong humoral responses during the late clinical stages of the disease. Cell-mediated immune responses modulated by various T cell subsets are essential to provide protective immunity and prevent progression of the disease. Secretion of cytokines by T cell populations serves to activate macrophages to kill ingested M. paratuberculosis as well as activate other T cell subsets to contain the infection. This paper reviews the current knowledge of T cell immune responses in M. paratuberculosis infection based upon clinical studies and research using mouse models. PMID- 11118732 TI - Differences in the immune responses in lambs and kids vaccinated against paratuberculosis, according to the age of vaccination. AB - In order to evaluate and compare the peripheral immune responses induced by the vaccination against paratuberculosis in relation with the age of immunization, two groups of lambs and goat kids were vaccinated at 15 days and 5 months old, respectively. A heat-killed commercial vaccine was inoculated subcutaneously and humoral and cellular immune responses were measured by an ELISA and IFN-gamma assay, respectively, at 0, 30, 90, 180, 270 and 360 dpv in the lambs and 0, 30, 90 and 180 dpv in the caprine. IFN-gamma values did not show statistically significant differences between both groups, but when compared to the unvaccinated controls, this cytokine response tend to disappear earlier in animals vaccinated at 15 days old. The antibody response was always higher and more persistent in animals vaccinated at 5 months. The possibility of the incomplete degree of maturation of the immune system in 15 days old animals as the cause of the differences in the immune response to vaccination is suggested. PMID- 11118733 TI - Lewis rats are not susceptible to oral infection with Mycobacterium avium subsp. paratuberculosis. AB - Pathogenesis studies of Mycobacterium avium subsp. paratuberculosis infection in ruminants are hampered by the long incubation time of the disease. A laboratory animal model with a shorter incubation time would facilitate research in this field. Although small rodents are usually considered to be resistant to M.a. paratuberculosis infection, several susceptible murine strains have been found. To our knowledge, there are no detailed reports with regard to susceptibility in rats. The Lewis rat is a valuable model for inflammatory bowel disease studies as well as autoimmune diseases involving mycobacteria as inducing agents. In this study Lewis rats were used to investigate their potential as a small laboratory animal model for paratuberculosis. In total 28 female Lewis rats were orally inoculated with M.a. paratuberculosis. The rats were first inoculated at 3 weeks of age, and 12 more inoculations followed in increasing intervals during the 3 months to follow. Eight control rats received a sham inoculation. Over 9 months, two rats from each group were sacrificed at regular intervals and immunological and histopathological examinations were performed on the gastrointestinal tract, the liver and the spleen. None of the rats developed lesions which were indicative of mycobacterial infection as determined by histology with HE and Ziehl-Neelsen staining. The bacteria could not be recultured from samples taken from the gut, the liver or the spleen. The immunological tests however, showed that bacteria had entered via the intestinal tract. From this study it appears that Lewis rats are resistant to oral inoculation with M. a. paratuberculosis, and not suitable as a model to study the immunopathogenesis of paratuberculosis as it occurs in ruminants. PMID- 11118734 TI - Specific seroreactivity of Crohn's disease patients against p35 and p36 antigens of M. avium subsp. paratuberculosis. AB - Crohn's disease (CD) is a chronic inflammatory bowel disease that is similar to Johne's disease in ruminants. Recent data have strengthened the association of M. avium subsp. paratuberculosis (M. paratuberculosis) with CD. To provide more evidence of an etiological association, antibody reactivities from CD patients were tested by immunoblotting against recombinant antigens that were identified previously from our M. paratuberculosis genomic library. Two clones (designated pMptb#40 (3.2-kb insert) and #48 (1.4-kb insert) expressing a 35K (p35)- and 36K(p36)-antigens showed specific reactivities with serum samples from CD patients. Serum samples from 75% of 53 CD patients, 14% of 35 normal individuals and 10% of 10 ulcerative colitis patients reacted to p35 antigen. Reactivities were also observed with serum samples from 89% of 89 CD patients, 14% of 50 normal controls and 15% of 29 ulcerative colitis patients reacted with p36 antigen. When the reactivity results from p35 and p36 were combined, the background from the controls was eliminated, i.e. only the CD patients reacted to both p35 and p36. The positive predictive value was 98% with specificity of 98% and the negative predictive value was 76% with sensitivity of 74% (39 positive out of 53). A statistical significance (p<0.0001) was observed when the results from CD serum samples reacting with either or both antigens were compared to the controls. The reactivity of anti-M. paratuberculosis (specifically against p35 and p36 antigens) antibodies in a significant proportion of CD patients would suggest a causal role for the organism in CD. PMID- 11118735 TI - Pathogenesis and therapeutic aspects of Crohn's disease. AB - Crohn's disease is a chronic, relapsing inflammatory condition affecting any part of the human gastrointestinal tract. It is characterised by transmural inflammation with deep ulceration, thickening of the bowel wall and fistula formation. The hallmark is the non-caseating granuloma. Clinical presentation depends upon the site of the inflammation. Pain and diarrhoea are frequent. Extraintestinal manifestations develop in up to 25% of patients and perianal disease is also frequent. The aetiology of Crohn's disease remains unknown. On the host side, genetic factors are important and the immune system is central to the development of the inflammation. Several environmental factors also play a role, in particular smoking. The presence of intestinal luminal contents appears to be essential for the development of Crohn's disease. A number of specific infectious causes have also been proposed, most recently measles virus and M. avium subsp. paratuberculosis. The latter has been considered because of the similarity between BJD and Crohn's disease, although there are also important differences. Evidence suggesting that this agent plays a role includes isolation of the organism from some patients, clustering, and identification by PCR of M.a. paratuberculosis DNA in tissue. However, many other workers have been unable to reproduce these findings.Treatment of Crohn's disease is generally with 5 aminosalicylic acid (5-ASA) compounds, corticosteroids, immunosuppressive agents and antibiotics. The majority of patients with active disease will respond to one or a combination of the therapies. Recently, broad-spectrum antibiotics have been shown to be as effective as oral corticosteroids. The challenge in Crohn's disease remains the prevention of relapse once remission has been achieved. Oral 5-ASA preparations can be beneficial, particularly after surgery. Immunosuppression, particularly with azathioprine or 6-mercaptopurine, is helpful in patients with resistant inflammation. Antibiotics may delay the time to relapse when used for active disease. The use of antimycobacterial therapy directed against M.a. paratuberculosis shows promising results but needs further evaluation.Up to 80% of patients with Crohn's disease will require surgery at some stage in the course of their illness. The challenge remains to try and prevent resection of inflamed intestine and to improve the quality of life of those affected by this disorder. PMID- 11118736 TI - In situ hybridization method for studies of cell wall deficient M. paratuberculosis in tissue samples. AB - Cell wall deficient forms of mycobacteria may be important in the pathogenesis of Crohn's disease and sarcoidosis. However, no method has been available to localize this type of organisms in tissue sections. We developed an in situ hybridization method for the demonstration of Mycobacterium paratuberculosis spheroplasts (cell wall deficient forms) in paraffin embedded tissue sections.M. paratuberculosis spheroplasts were prepared by treatment with glycine and lysozyme. Pieces of beef were injected with the prepared spheroplasts. The samples were fixed in buffered formalin and paraffin embedded. A M. paratuberculosis-specific probe derived from the IS900 gene was used. Specificity was controlled by using an irrelevant probe and by hybridizing sections with spheroplasts from other bacteria. Beef samples injected with M. paratuberculosis spheroplasts were the only samples that hybridized with the probe. Beef samples containing acid-fast or spheroplast forms of M. smegmatis and M. tuberculosis as well as the acid-fast forms of M. paratuberculosis did not hybridize with the probe. Unrelated bacterial controls, i.e. Helicobacter pylori and Escherichia coli were also negative in the assay. In situ hybridization with the IS900 probe provides a specific way to localize M. paratuberculosis spheroplasts in tissue sections and may be useful for studies of the connection between M. paratuberculosis and Crohn's disease and sarcoidosis. The assay may also be valuable for studies on Johne's diseased animals. PMID- 11118737 TI - Nucleotide sequence of L1 and part of P1 of hexon gene of fowl adenovirus associated with hydropericardium hepatitis syndrome differs with the corresponding region of other fowl adenoviruses. AB - In order to study the serotypic variations in hydropericardium hepatitis syndrome (HHS) causing virus, the DNA was extracted from the purified virus, a 0.7 kb variable region of hexon gene encoding L1 and part of P1 amplified and sequenced. Both nucleotide and derived amino acid sequences, corresponding to the variable region, were compared with the published fowl adenovirus sequences (FAV serotypes 10, 1 and 8). As expected the 0.7 kb sequence showed single open reading frame (ORF). There was a nucleotide sequence variation of 8.2, 28.1 and 40.3%, respectively, with FAV serotypes 10, 1 and 8. The dendrogram constructed with the nucleotide sequences showed that HHS virus and FAV10 are closer to each other and are away to FAV1 and FAV8. However, the derived amino acid sequence showed variations as high as 28.8, 38 and 45.1% with FAV serotypes 10, 1 and 8, respectively. Such high degree variation has been found due to the shift in the reading frame caused by deletions indicating that the FAV4 associated with HHS is unique and different from FAV10. To the best of our knowledge this is the first report on nucleotide sequence analysis of hexon gene fragment of FAV4 associated with HHS. PMID- 11118738 TI - Genetic diversity and evolution of Mycoplasma capricolum subsp. capripneumoniae strains from eastern Africa assessed by 16S rDNA sequence analysis. AB - Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae), the causal agent of contagious caprine pleuropneumonia (CCPP), is a member of the so-called Mycoplasma mycoides cluster. These mycoplasmas have two rRNA operons in which intraspecific variations have been demonstrated. The sequences of the 16S rRNA genes of both operons from 13 field strains of M. capripneumoniae from three neighbouring African countries (Kenya, Ethiopia, and Tanzania) were determined. Four new and unique polymorphism patterns reflecting the intraspecific variations were found. Two of these patterns included length differences between the rrnA and rrnB operons. The length difference in one of the patterns was caused by a two-nucleotide insert (TG) in the rrnB operon and the length difference in the other pattern was due to a three-nucleotide deletion, also in the rrnB operon. Another pattern was characterised by a polymorphic position caused by a mutation that is known to cause streptomycin resistance in other bacterial species. The strain with this pattern was also found to be resistant to streptomycin. Streptomycin resistant clones were selected from four M. capripneumoniae strains to further investigate the correlation of this mutation to streptomycin resistance. Mutations in the 16S rRNA genes had occurred in two of these strains. The fourth pattern included a new polymorphism in position 1059. The results show that polymorphisms in M. capripneumoniae strains can be used as epidemiological markers for CCPP in smaller geographical areas and to study the molecular evolution of this species. PMID- 11118739 TI - Influence of ampicillin, ceftiofur, attenuated live PRRSV vaccine, and reduced dose Streptococcus suis exposure on disease associated with PRRSV and S. suis coinfection. AB - The objective of this research was to evaluate the efficacy of two antimicrobials (ampicillin and ceftiofur), a modified-live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine, and low dose exposure to Streptococcus suis on disease associated with PRRSV/S. suis coinfection. Fifty-six, crossbred, PRRSV free pigs were weaned at 10-12 days of age and randomly assigned to five treatment groups. All pigs were inoculated with 2ml of 10(6.4) TCID50/ml of high virulence PRRSV isolate VR-2385 intranasally at 29-31 days of age (day 0 of the study) followed 7 days later by intranasal inoculation with 2ml of 10(8.9)colony forming units(CFU)/ml S. suis type 2 isolate ISU VDL #40634/94. Pigs in group 1 (n=10) served as untreated infected positive controls. Pigs in group 2 (n=12) were treated with 5.0 mg/kg ceftiofur hydrochloride intramuscularly (IM) on days 8, 11, and 14. Pigs in group 3 (n=11) were treated with 11 mg/kg ampicillin IM on days 8-10. Pigs in group 4 (n=12) were vaccinated 14 days prior to PRRSV challenge with a commercial modified-live PRRSV vaccine. Pigs in group 5 (n=11) were exposed to a 1:100 dilution of the S. suis challenge inoculum 19 days prior to S. suis challenge. Mortality was 80, 25, 82, 83, and 36% in groups 1-5, respectively. The reduced dose S. suis exposure had some residual virulence, evidenced by S. suis induced meningitis in two pigs after exposure. Treatment with ceftiofur hydrochloride and reduced dose exposure to S. suis were the only treatments which significantly (P<0.05) reduced mortality associated with PRRSV/S. suis coinfection, significantly (P<0.05) reduced recovery of S. suis from tissues at necropsy, and significantly (P<0.05) reduced the severity of gross lung lesions. PMID- 11118740 TI - Genetic analysis of exfoliative toxin A-producing Staphylococcus aureus isolated from mastitic cow's milk. AB - Exfoliative toxin A (ETA), produced by Staphylococcus aureus, is the causative agent of staphylococcal scalded-skin syndrome (SSSS) in children. Recently, we reported that ETA was detected by reverse passive latex agglutination in three isolates of S. aureus from cow's milk, but that these ETA-positive isolates did not cause the so-called Nikolsky sign in neonatal mice. In this study, therefore, the eta gene encoding ETA and regulatory genes of these bovine isolates were analyzed by the polymerase chain reaction (PCR) and sequencing. The eta gene was amplified from three bovine isolates by PCR and their resulting nucleotide sequences found to correspond to the eta gene from the human isolate, except for three nucleotides in the upstream region of the eta open reading frame (ORF). An accessory gene regulator (agr), which is a global regulatory locus, was detected in these bovine isolates by PCR amplification. In addition, the ORF (J-4), located 120 bp upstream from the eta ORF of the human isolate, was also amplified from these bovine isolates, with their nucleotide sequences differing at 32 positions from the human isolate. Bovine and human ORF J-4 equally enhanced production of ETA in the recombinants of the eta gene, suggesting that the variation in bovine ORF J-4 may be not be the cause of the difference in amount of ETA produced by bovine and human isolates. PMID- 11118741 TI - Study of antigenic heterogeneity among Actinobacillus pleuropneumoniae serotype 7 strains. AB - Actinobacillus pleuropneumoniae serotype 7 strains were studied for their antigenic heterogeneity using rabbit polyclonal hyperimmune sera against all the known twelve reference strains of A. pleuropneumoniae and a battery of different serological tests such as coagglutination (COA), immunodiffusion (ID), indirect hemagglutination (IHA), counterimmunoelectrophoresis (CIE), rapid dot-ELISA (RDE), serum soft-agar (SSA) and growth agglutination (GA). Reference serotype 7 strain (WF83) showed cross-reactivity with reference serotype 1B strain but not with other serotypes. Field serotype 7 strains showed cross-reactivities with serotypes 1A, 1B, 4, 9, 10, and 11 in COA, ID, and CIE tests, but not in IHA test. Two field strains of serotype 7 (90-3182 and 86-1411) which appeared to be different from the typical serotype 7 strains were selected for further antigenic characterization by SDS-PAGE, Western blot, and Tricine SDS-PAGE assays, and identified as serotypes 1 and 7, respectively. For serotyping atypical strains, it is suggested to use Western blot assay as a confirmatory test to identify serotype-specific capsular and somatic antigens. PMID- 11118742 TI - Analysis of expression of flagella by Salmonella enterica serotype typhimurium by monoclonal antibodies recognising both phase specific and common epitopes. AB - Monoclonal antibodies specific for phase 1 ("i" antigen), phase 2 ("1,2" antigen) and common epitopes of the flagellins of Salmonella enterica serotype Typhimurium were raised. Having confirmed their specificity, the monoclonal antibodies were used to develop semi-quantitative ELISAs in order to assess the relative expression of the two phases by strains of Typhimurium. The majority of Typhimurium strains representative of a wide cross-section of definitive types from animal and environmental sources preferentially expressed phase 1 antigen in vitro. DT40 strains were unique in expressing phase 2 preferentially. The ratio of phase 1 to phase 2 expressed by strains tended to be constant for any one strain when strains were grown on a number of conventional laboratory media. However, the ratio of phases was shown to be modulated by incubation at 42 degrees C and buffering media at pH values, notably 4.5, other than neutral. Selenite broth and Rambach media repressed flagellation. PMID- 11118743 TI - Antibodies against bovine herpesvirus (BHV) 5 may be differentiated from antibodies against BHV1 in a BHV1 glycoprotein E blocking ELISA. AB - We examined whether antibodies against bovine herpesvirus (BHV) 5 cross-react with BHV1 antigens and whether they could interfere with BHV1 eradication programmes. Six calves were experimentally infected with different doses of BHV5 strain N569; homologous antibodies were first detectable on day 11 post infection; they cross-reacted in a BHV1 virus neutralisation test, in a BHV1 glycoprotein (g)-B blocking ELISA and in a BHV1-gE ELISA, but not in a BHV1-gE blocking ELISA. This study indicates that, in ongoing BHV1 eradication programmes, based on vaccines that lack gE, BHV5 infections may not lead to false positive serological reactions in case cattle are tested for BHV1-gE antibodies by the BHV1-gE blocking ELISA; antibodies against BHV5 may be differentiated from antibodies against BHV1. The BHV1-gE blocking ELISA may, therefore, offer opportunities for the serological differentiation between BHV1 and BHV5 infections. PMID- 11118744 TI - Chitin synthase 1 (Chs1) gene sequences of Microsporum equinum and Trichophyton equinum. AB - Chitin synthase 1 (Chs1) genes from Microsporum equinum and Trichophyton equinum were compared with those of the other dermatophytes. The Chs1 nucleotide sequences of these dermatophytes from horses showed more than 80% similarity to those of Arthroderma benhamiae, A. fulvum, A. grubyi, A. gypseum, A. incruvatum, A. otae, A. simii, A. vanbreuseghemii, Epidermophyton floccosum, T. mentagrophytes var. interdigitale (T. interdigitale), T. rubrum and T. violaceum. Especially high degree of nucleotide sequence similarity of more than 99% was noted between the Chs1 gene fragments of M. equinum and A. otae, and those of T. equinum, T. interdigitale and A. vanbreuseghemii, respectively. The phylogenetic analysis of their sequences revealed that M. equinum was genetically very close to A. otae and T. equinum to A. vanbreuseghemii. A molecular analysis of Chs1 genes will provide useful information for the genetic relatedness of M. equinum and T. equinum and confirm the value of DNA sequencing in identification of these two dermatophytes. PMID- 11118745 TI - Erratum to "Pathogenic characteristics of Escherichia coli strain isolated from newborn piglets with diarrhoea in Brazil". PMID- 11118746 TI - Analysis of microsatellite polymorphism in red deer, roe deer, and fallow deer -- possible employment in forensic applications. AB - DNA microsatellites play a major role in population genetics, linkage mapping, and parentage studies of mammals. In addition, they may be used for forensic purposes, if an individual identification of a specific animal is necessary. Therefore, we tested a variety of microsatellite polymorphism derived from reindeer (Rangifer tarandus) by PCR and sequencing analysis for use in red deer (Cervus elaphus), roe deer (Capreolus capreolus) and fallow deer (Dama dama). Twelve of these microsatellites were selected for further analysis. In all these microsatellite polymorphism short tandem repeats could be detected for one or all three species as shown by sequencing analysis. In red deer, more than two alleles were found in eight microsatellites, in roe deer more than two alleles could be demonstrated in seven microsatellites, whereas in fallow deer more than two alleles were found in only two microsatellite polymorphism. A comparison of sequences of PCR products from the three deer species with the sequences of reindeer revealed several differences between the four species. In six microsatellites -- selected because or their reliability in PCR and because of their polymorphic character -- we established a sequenced allelic ladder and give population data of all three species from 82 deer of the Northeast region of Germany (Vorpommern). Our results show the possibility to use microsatellite polymorphism in the identification of deer in forensic applications like poaching. PMID- 11118747 TI - Blood concentrations of tetracaine and its metabolite following spinal anesthesia. AB - Blood concentrations of tetracaine and its metabolite, p-butylaminobenzoic acid, were measured after spinal anesthesia with tetracaine which had been administered to patients under going orthopedic surgery. Tetracaine, an ester anesthetic, was given to 10 patients, the dose was 8-14mg, and blood samples were collected 1, 2 and 6h after the injection of tetracaine. We used gas chromatography/mass spectrometry for purposes of analysis. Tetracaine was not detected in any blood sample, but the metabolite was detected in each sample with the mean concentrations of 126.5, 97.9 and 43.3ng/ml at 1, 2 and 6h, respectively. This data will be useful in determination of the cause of death after spinal anesthesia with tetracaine. PMID- 11118748 TI - Highly sensitive analysis of methamphetamine and amphetamine in human whole blood using headspace solid-phase microextraction and gas chromatography-mass spectrometry. AB - A simple and highly sensitive method for analysis of derivatized methamphetamine (MA) and amphetamine (AM) in whole blood was developed using headspace solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry electron impact ionization selected ion monitoring (GC-MS-EI-SIM). A whole blood sample, deuterated-MA (d(5)-MA), as an internal standard (IS), tri-n-propylamine and pentafluorobenzyl bromide were placed in a vial. The vial was heated and stirred at 90 degrees C for 30min. Then the extraction fiber of the SPME was exposed at 90 degrees C for 30min in the headspace of the vial while being stirred. The derivatives adsorbed on the fiber were desorbed by exposing the fiber in the injection port of a GC-MS. The calibration curves showed linearity in the range of 0.5-1000ng/g for both MA and AM. The time for analysis was about 80min per sample. In addition, this proposed method was applied to two autopsy cases where MA ingestion was suspected. In one case, MA and AM concentrations in the mixed left and right heart blood were 165 and 36.9ng/g, respectively. In the other case, MA and AM concentrations were 1.79 and 0.119 microg/g in the left heart blood, and 1.27 and 0.074 microg/g in the right heart blood, respectively. PMID- 11118749 TI - A report of the 1997, 1998 and 1999 Paternity Testing Workshops of the English Speaking Working Group of the International Society for Forensic Genetics. AB - We present the results of the 1997, 1998 and 1999 Paternity Testing Workshops of the English Speaking Working Group of the International Society for Forensic Genetics. The numbers of participating laboratories were 24 (1997), 31 (1998) and 32 (1999). In 1997, all laboratories drew the correct conclusion that the alleged father was the biological father of the child. In 1998, the alleged father was the biological brother of the child and all laboratories excluded him. The scenario in 1999 was a deficiency case consisting of mother, child and the parents of the alleged father and all but one laboratory drew the correct conclusion.The percentage of laboratories routinely performing variable number of tandem repeat (VNTR) investigations using single locus probes (SLPs) and restriction fragment length polymorphism (RFLP) decreased from 83% in 1997 to 66% in 1999. In the three workshops, more than 90% of the laboratories used polymerase chain reaction (PCR) based systems. In 1999, 80% of the laboratories performing PCR, used commercially available short tandem repeat (STR) kits. Other commonly used systems were HLA and PolyMarker investigated with PCR. Conventional systems and RFLP investigations of VNTRs with multi loci probes (MLPs) were used routinely by approximately 20% of the participating laboratories. All laboratories submitting results in the three workshops used RFLP-based VNTRs or/and PCR based VNTRs/STRs. Inter-laboratory comparisons of the results showed a very high concordance. The overall coefficients of variation between the laboratories of the results of RFLP typing of the commonly used VNTRs D2S44, D7S21, D7S22 and D12S11 were 1.2-1.3%. Consistent results were obtained in the great majority of the systems investigated by PCR and typing errors counted for less than 0.3% of the PCR based results. PMID- 11118750 TI - Genetic variation at eight STR loci in the Korean population. AB - The frequency distributions of eight STR loci were surveyed in 510 unrelated individuals from the Korean population PMID- 11118751 TI - Data for nine autosomal STRs markers from Valencia (East Mediterranean coast of the Iberian Peninsula). AB - Nine STRs loci have been typed in a sample from Valencia, a population from the East Mediterranean coast of the Iberian Peninsula. PMID- 11118752 TI - STR data for 21 loci in northwestern Africa. PMID- 11118753 TI - STR data from S. Tome e Principe (Gulf of Guinea, West Africa). AB - Allele frequencies for eight STRs (CD4, FES/FPS, MBPB, TH01, TP53, TPO, F13A1, VWA) were estimated from samples (sized between 279 and 328) of unrelated individuals born in S. Tome e Principe (Gulf of Guinea, West Africa). PMID- 11118754 TI - Sequence analysis and population data on the 'new' short tandem repeat locus D5S2360. AB - We have studied the sequence structure and population genetics of a 'new' short tandem repeat polymorphism at locus D5S2360 in German Caucasians. Sequencing at this locus revealed a considerable variation, which is characterized by a tetranucleotide (AGAT)(n) repeat pattern with (GAT), (AGATT), and (AG) repeats dispersed throughout the alleles. These microvariations do not necessarily alter the size of the alleles. They may vary by one or two pairs or they may remain unchanged in size. At locus D5S2360 we observed 33 allelic lengths comprising at least 36 different alleles. Population data revealed a high polymorphism with a heterozygosity rate of approximately 92.5%. PMID- 11118755 TI - Blood carbofuran concentrations in suicidal ingestion cases. AB - We describe four fatal cases due to ingestion of carbofuran, a carbamate insecticide. Carbofuran was detected in the gastric contents using thin layer chromatography (TLC) and gas chromatography/mass spectrophotometry (GC/MS), and quantified in the blood using a gas chromatograph equipped with nitrogen phosphorus detector (NPD). Fatal concentrations of carbofuran in blood ranged from 0.32 to 11.6 microg/ml. PMID- 11118756 TI - DNA profiling of disputed chilli samples (Capsicum annum) using ISSR-PCR and FISSR-PCR marker assays. AB - A case of marketing of spurious seeds of chilli, Capsicum annum in the brand name of an elite variety referred to us from an Indian court of law, for identification is described here. The highly reproducible molecular marker assays, inter simple sequence repeat polymerase chain reaction [ISSR-PCR] and FISSR-PCR (for fluorescent ISSR-PCR) were used for differentiating the four disputed chilli samples. A total number of 17 ISSR anchored primers, which included nine di-, and eight tri-nucleotide primers were used for the analysis. The ISSR-PCR products were separated on a 2% agarose gel. A total of 212 and 288 bands were resolved by seven di- and eight tri-nucleotide primers, respectively, with an average of 30 bands per primer. Five out of nine dinucleotide primers and four out of eight trinucleotide primers could unambiguously differentiate all the four disputed chilli samples. The sensitivity and informativeness of the ISSR-PCR assay were further enhanced by the use of FISSR-PCR technique. The FISSR-PCR assay revealed a total number of 566 bands using three tri- and one di-nucleotide primers with an average of 141 bands per primer. These four primers could reliably distinguish all the four disputed samples unambiguously. In developing countries like India, violation of Plant Breeder's Rights is a major concern of law. The present report is, therefore, a step to protect the Plant Breeder's Rights by making use of reliable and modern DNA technologies. PMID- 11118757 TI - Fatal retroperitoneal haemorrhage: an unusual complication of percutaneous endoscopic gastrostomy. AB - A 93-year-old lady with dementia, neurological dysphagia and aspiration pneumonia, died from massive retroperitoneal haemorrhage which developed as a rare and, it is believed, hitherto unreported, complication of percutaneous endoscopic gastrostomy (PEG), which was performed for feeding purposes. It is postulated that the initial, unsuccessful attempt at needle puncture of the stomach, under endoscopic guidance, had resulted in iatrogenic perforation and laceration of the splenic and superior mesenteric veins close to their confluence with the portal vein. It would also appear that dense fibrous adhesions between the pyloro-antral region of the stomach and the posterior hepatic surface had altered the immediate anatomical relations of the stomach in such a manner as to have predisposed to these events. PMID- 11118758 TI - Fatal traumatic rupture of an aortic aneurysm of the sinus of Valsalva: an autopsy case. AB - This report describes an autopsy case of a rare type of aortic sinus of Valsalva aneurysm, which caused fatal rupture from a blunt chest impact. A 51-year-old male was hit in the chest with a fist, lost consciousness after about 15min and died after approximately 7h. The postmortem examination revealed a large saccular aneurysm of the right coronary sinus bulging on the right atrium, which had a full laceration causing pericardial hematoma (cardiac tamponade). A related chest wall injury was observed in the right outer mammary region. A rare type of bulging onto the right atrium and subsequent sclerosis of the right coronary artery appeared to have greatly contributed to the fatal rupture. PMID- 11118759 TI - Mapping of the cerebral response to hypoxia measured using graded asymmetric spin echo EPI. AB - Graded asymmetric spin echo-echo planar imaging (ASE-EPI) was used to measure transient alterations in cerebral oxygenation resulting from 60 seconds of anoxia in alpha-chloralose anaesthetised rats. The anoxic period induced a transient fall ( approximately 1 min) in signal intensity followed by a prolonged signal overshoot consistent with an autoregulatory response to oxygen deprivation. The magnitude of signal response, integrated over the entire brain, increased linearly with the echo asymmetry (t(ge)). However, that increase in sensitivity was offset by a reduced signal to noise ratio and quality of the image data. The responses of four regions of interest within the brain to the anoxic stimulus, and the effect of increasing the echo asymmetry, were compared. A comparable magnitude of signal decrease was observed in all brain regions except the superficial cortex that included pial vessels. As t(ge) was incremented differences in signal attenuation between regions became more pronounced. The signal overshoot observed upon restoration of normal breathing gases showed similar trends, producing similar normalised vascular responses for all regions of interest studied. Different regions of interest showed comparable time courses of the signal overshoot suggesting that similar autoregulatory vascular mechanisms operate in all brain regions. These findings additionally show that the use of graded ASE-EPI produced a characteristic profile of maximum signal change measured during and following the anoxic period for each brain region. They suggest that the shape of this profile was determined by the local vasculature within each region of interest; this feature could be exploited in activation studies to eliminate regions with significant signal changes originating from large draining vessels. Finally, the consistent physiological response observed, when the overshoot was compared to the magnitude of the signal drop, demonstrated that modification of the spin echo offset parameter did not mask or detrimentally alter the signal change resulting from the underlying physiological perturbation. PMID- 11118760 TI - Compensation of susceptibility-induced signal loss in echo-planar imaging for functional applications. AB - Functional magnetic resonance imaging favors the use of multi-slice gradient recalled echo-planar imaging due to its short image acquisition times, whole brain coverage and sensitivity to BOLD contrast. However, despite its advantages, gradient-recalled echo-planar imaging also is sensitive to static magnetic field gradients arising primarily from air-tissue interfaces. This can lead to image artifacts such as voxel shifts and complete signal loss. A method to recover signal loss by adjusting the refocusing gradient amplitude in the slice-select direction, preferably axially, is proposed. This method is implemented as an automated computer algorithm that partitions echo-planar images into regions of recoverable signal intensities using a histogram analysis and determines each region's proper refocusing gradient amplitude. As an example, different refocusing gradient amplitudes are interleaved in a fMRI acquisition to maximize the signal to noise ratio and obtain functional activation in normal and dropout regions. The effectiveness of this method is demonstrated by recovering signal voids in the orbitofrontal cortex, parahippocampal/amygdala region, and inferior visual association cortex near the cerebellum. PMID- 11118761 TI - Conventional high resolution versus fast T(2)-weighted MR imaging of the heart: assessment of reperfusion induced myocardial injury in an animal model. AB - Cardiac image quality in terms of spatial resolution and signal contrast was assessed for conventional and newly developed T(2)-weighted fast spin-echo imaging with high k-space segmentation. The capability in revealing regional myocardial edema and cellular damage was examined by a porcine model using histopathologic correlation. Twelve porcine hearts were excised from slaughtered animals and instantly perfused with 1000 mL cold cardioplegic solution. After 4 h of cold ischemia the hearts were reperfused for one hour using a "Langendorff" perfusion model followed by MR imaging at 1.5 Tesla. Three additional pig hearts served as controls and were studied by MR directly after harvesting. Histopathological analysis of regional tissue changes was performed macro- and microscopically. Short axis T(2)-weighted (3000/45 and 90) high quality fast spin echo (FSE) images were recorded without cardiac action and signal intensity was correlated with histology. These images also served as gold standard for evaluation of newly developed faster sequences allowing measuring times shorter than 20 s. Fast T(2)-weighted imaging comprised single-slice fast spin echo (moderate echo train length of 23 echoes, FSE(m)), and multi-slice single-shot half-Fourier fast spin-echo (71 echoes, FSE(HASTE)) sequences, supplemented by versions with inversion recovery preparation (FSE(m)IR and FSE(HASTE)IR). Systolic function after reperfusion was restored in 10 porcine hearts. Tissue alterations included myocardial edema and contraction band necrosis which was found to be most severe in myocardium with maximum T(2) SI. Especially FSE(m) and FSE(m)IR sequences allowed differentiation of all categories of tissue damage on a high level of significance. In contrast, single-shot FSE(HASTE) and FSE(HASTE)IR sequences did not provide sufficient image quality to discriminate moderate and severe myocardial damage (p > 0.05). Different degrees of myocardial injury after ischemia and reperfusion can be staged by MR imaging, especially using conventional high resolution T(2)-weighted FSE sequences. The animal study indicates that fast T(2)-weighted FSE(m) and FSE(m)IR sequences lead to superior image quality and diagnostic accuracy compared to FSE(HASTE) and FSE(HASTE)IR imaging. PMID- 11118762 TI - Superparamagnetic iron oxide-enhanced MR imaging for early and late radiation induced hepatic injuries. AB - Superparamagnetic iron oxide (SPIO)-enhanced MRI was performed in twenty-one patients undergoing proton-beam radiotherapy for hepatocellular carcinomas. Patients were divided into two groups: early and late phase hepatic injuries. Each group was investigated 3 to 9 weeks and 4 to 65 months after the start of irradiation, respectively. T(1)-weighted, T(2)-weighted, and T(2)*-weighted images were obtained before and after SPIO administration. In all postcontrast sequences in the early phase, irradiated livers demonstrated relatively higher intensity than nonirradiated livers and the radiation-to-liver contrast-to-noise ratio (C/N) was improved. Postcontrast T(2)*-weighted images showed the highest C/N. In the late phase, the irradiated areas showed high intensity on T(2) weighted images and low intensity on T(1)-weighted images without SPIO, while high intensity on T(1)-weighted images with SPIO. The C/N increased with SPIO in all sequences and postcontrast T(2)-weighted images showed the highest C/N in the late phase. SPIO-enhanced MRI is useful to evaluate this entity both in the early and late phase of clinical studies. PMID- 11118763 TI - MR imaging and MR angiography in vascular Behcet's disease. AB - The aims of this study are to demonstrate the ability of magnetic resonance imaging (MRI) and magnetic resonance angiography (MRA) in the diagnosis and evaluation of vascular involvement in Behcet's disease. Twelve patients with vascular involvement due to Behcet's disease were included in this study. We believe that MRI and MRA are safe and noninvasive methods that can be used to confirm and monitor vascular Behcet's disease. PMID- 11118764 TI - Four dimensional bolus tagging imaging of pulsatile flow. AB - Inversion bolus tagging MR methods were used to provide a graphic depiction of the axial velocity in three spatial dimensions for pulsatile flow through complex geometries. Visualization of the flow field was readily apparent, and a train of tagged boli were depicted providing an immediate overview of the displacement of flowing fluid over the entire pulsatile cycle. Tagging efficiency obtained using adiabatic inversion pulses was improved compared to that with a windowed sinc pulse. Results from phantom experiments on steady flow were correlated with computational fluid dynamic (CFD) simulations. The use of 3D methods reduced spatial partial volume effects, and the displacement of boli in a steady flow experiment correlated well with CFD simulations. The use of adiabatic inversion pulses resulted in sharp edged inversion regions with good retention of longitudinal magnetization. However in order to keep the pulse duration short, of the order of 2-5 ms, a rather large RF amplitude had to be used. The inversion bolus tagging method is useful in visualizing the flow field in multiple levels for pulsatile fluid flowing through complex geometries, and may be useful in fluid dynamic applications. PMID- 11118765 TI - Arterial spin tagging perfusion imaging of rat brain: dependency on magnetic field strength. AB - Perfusion-weighted imaging (PWI), using the method of arterial spin tagging, is strongly T(1)-dependent. This translates into a high field dependency of the perfusion signal intensity. In order to determine the expected signal improvement at higher magnetic fields we compared perfusion-weighted images in rat brain at 4.7 T and 7 T. Application of PWI to focal ischemia and functional activation of the brain and the use of two different anesthetics allowed the observation of a wide range of flow values. For all these (patho-)physiological conditions switching from 4.7 T to 7 T resulted in a significant increase of mean perfusion signal intensity by a factor of 2.96. The ratio of signal intensities of homotopic regions in the ipsi- and contralateral hemisphere was field independent. The relative contribution of a) T(1) relaxation time, b) net magnetization, c) the Q-value of the receiver coils and d) the degree of adiabatic inversion to the signal improvement at higher field strength were discussed. It was shown that the main parameters contributing to the higher signal intensity are the lengthening of T(1) and the higher magnetization at the higher magnetic field. PMID- 11118766 TI - On the accuracy of EPI-based phase contrast velocimetry. AB - For over a decade, echo-planar imaging (EPI) has been used in both the medical and applied sciences to capture velocity fields of fluid flows. However, previous studies have not rigorously confirmed the accuracy of the measurements or sought to understand the limitations of the technique. In this study, a bipolar gradient was added to a flow-compensated EPI pulse sequence to obtain rapid phase contrast images of steady and unsteady flows through two step stenoses. For steady Re = 100 and 258 flows, accuracy was measured through systematic comparisons with CFD simulations, mass flow rate measurements, and spin echo phase contrast images. On average, the EPI image data exhibited velocity errors of 5 to 10 percent, while mass was conserved to within 5.6 percent at each axial position. Compared to spin echo phase contrast images, the EPI images have 50 percent lower signal-to-noise ratio, larger local velocity errors, and similar mass conservation characteristics. An unsteady flow was then examined by starting a pump and allowing it to reach a steady Re = 100 flow. Accuracy in this case was measured by the consistency between mass flow rate measurements at different axial positions. Images taken at 0.3 s intervals captured the velocity field evolution and showed that 50 to 100 percent errors occur when the flow changes on a time scale faster than the image acquisition time. PMID- 11118767 TI - Detection of susceptibility effects using simultaneous T(2)* and magnetic field mapping. AB - Tracking susceptibility effects is a convenient way to detect small inclusions in a bulk tissue matrix by MRI. We propose a quantitative assessment of these susceptibility effects by simultaneously mapping T(2)* and magnetic field from the time course of magnitude and phase using a multiple GE sequence at 4.7 T. A high-pass scheme is also introduced to highlight the mesoscopic magnetic field variations due to local susceptibility differences specifically in the magnetic field map. Applying this method to muscle tissue, we demonstrate that connective tissue generates detectable susceptibility effects through concomitant local magnetic field variation and T(2)* shortening. PMID- 11118768 TI - Wavelet-based enhancement for detection of left ventricular myocardial boundaries in magnetic resonance images. AB - MRI is noninvasive and generates clear images, giving it great potential as a diagnostic instrument. However, current methods of image analysis are too time consuming for dynamic systems such as the cardiovascular system. Since dynamic imagery generate a huge number of images, a computer aided machine vision diagnostic tool is essential for implementing MRI-based measurement. In this paper, a wavelet-based image technique is applied to enhance left ventricular endocardial and epicardial profiles as the preprocessor for a dynamic programming based automatic border detection algorithm. Statistical tests are conducted to verify the performance of the enhancement technique by comparing borders manually drawn with 1. borders generated from the enhanced images, and 2. borders generated for the original images. PMID- 11118769 TI - 3D image registration using a fast noniterative algorithm. AB - This note describes the implementation of a three-dimensional (3D) registration algorithm, generalizing a previous 2D version [Alexander, Int J Imaging Systems and Technology 1999;10:242-57]. The algorithm solves an integrated form of linearized image matching equation over a set of 3D rectangular sub-volumes ('patches') in the image domain. This integrated form avoids numerical instabilities due to differentiation of a noisy image over a lattice, and in addition renders the algorithm robustness to noise. Registration is implemented by first convolving the unregistered images with a set of computationally fast [O(N)] filters, providing four bandpass images for each input image, and integrating the image matching equation over the given patch. Each filter and each patch together provide an independent set of constraints on the displacement field derived by solving a set of linear regression equations. Furthermore, the filters are implemented at a variety of spatial scales, enabling registration parameters at one scale to be used as an input approximation for deriving refined values of those parameters at a finer scale of resolution. This hierarchical procedure is necessary to avoid false matches occurring. Both downsampled and oversampled (undecimating) filtering is implemented. Although the former is computationally fast, it lacks the translation invariance of the latter. Oversampling is required for accurate interpolation that is used in intermediate stages of the algorithm to reconstruct the partially registered from the unregistered image. However, downsampling is useful, and computationally efficient, for preliminary stages of registration when large mismatches are present. The 3D registration algorithm was implemented using a 12-parameter affine model for the displacement: u(x) = Ax + b. Linear interpolation was used throughout. Accuracy and timing results for registering various multislice images, obtained by scanning a melon and human volunteers in various stationary positions, is described. The algorithm may be generalized to more general models of the displacement field, and is also well suited to parallel processing. PMID- 11118770 TI - Evaluation of algorithms for analysis of NMR relaxation decay curves. AB - Quantitative processing of NMR relaxation images depends on the characteristics of the used fitting algorithm. Therefore several common fitting algorithms are compared for decay curves with low signal-to-noise ratios. The use of magnitude data yields a non-zero base line, and is shown to result in an overestimation of the decay time. A simple base line correction is no solution since this yields an equally large underestimation due to overcorrection of the first part of the curve. The use of squared data does yield reliable results, but only in the case of monoexponential decays. The best fitting algorithm under all experimentally occurring conditions turns out to be using real data after phase correction. A phase correction scheme is proposed, which applies to all imaging experiments for which the phase of the pixels is constant over the echo train. This scheme is validated for a phantom and for a tulip bulb. PMID- 11118771 TI - Short TE in vivo (1)H MR spectroscopic imaging at 1.5 T: acquisition and automated spectral analysis. AB - Spectral analysis of short TE in vivo proton magnetic resonance spectroscopic imaging (MRSI) data are complicated by the presence of spectral overlap, low signal to noise and uncharacterized signal contributions. In this study, it is shown that an automated data analysis method can be used to generate metabolite images from MRSI data obtained from human brain at TE = 25 ms and 1.5 T when optimized pulse sequences and a priori metabolite knowledge are used. The analysis approach made use of computer simulation methods to obtain a priori spectral information of the metabolites of interest and utilized a combination of parametric spectral modeling and non-parametric signal characterization for baseline fitting. This approach was applied to data from optimized PRESS-SI and multi-slice spin-echo SI acquisitions, for which sample spectra and metabolite images are shown. PMID- 11118772 TI - Correlation between proton magnetic resonance spectroscopic lactate measurements and vascular reactivity in chronic occlusive cerebrovascular disease: a comparison with positron emission tomography. AB - The purpose of this study is to investigate the correlation between lactate levels and cerebral vascular reactivity (VR) in regions outside an area of chronic cerebral infarction. Multivoxel proton magnetic resonance spectroscopy ((1)H-MRS) and positron emission tomography (PET) were performed in 11 patients who suffered chronic cerebral infarction. Of these 11 patients, 4 were examined before and after bypass surgery. Two regions-of-interests (ROIs) were placed outside the area of chronic infarction. One ROI was placed within a control region on the contralateral side. A lactate peak area was obtained in all ROIs. An N-acetyl aspartate (NAA) peak area was obtained in the ROI within the control region. The ratio of the lactate peak area and NAA peak area (Lct/NAA) was calculated for normalization of the lactate level, and was found to be 0.13 +/- 0. 10 (range, 0 to 0.43). The VR was recorded at 13.3 +/- 20.7% (range, - 44.3 to 68.9%), utilizing PET and administering acetazolamide. A significant negative correlation was observed between the Lct/NAA ratio and VR (r = - 0.709, p < 0.0001). These results suggest that lactate levels and VR are closely related in regions outside areas of chronic cerebral infarction. PMID- 11118773 TI - Mealiness assessment in apples and peaches using MRI techniques. AB - Mealiness (woolliness in peaches) is a negative attribute of sensory texture that combines the sensation of a desegregated tissue with the sensation of lack of juiciness. In this study, 24 apples cv. Top Red and 8 peaches cv. Maycrest, submitted to 3 and 2 different storage conditions respectively have been tested by mechanical and MRI techniques to assess mealiness. With this study, the results obtained on apples in a previous work have been validated using mathematical features from the histograms of the T2 maps: more skewed and the presence of a tail in mealy apples, similar to internal breakdown. In peaches, MRI techniques can also be used to identify woolly fruits. Not all the changes found in the histograms of woolly peaches are similar from those observed in mealy apples pointing to a different underlying physiological change in both disorders. PMID- 11118774 TI - Serial gadolinium-DTPA of spinal cord MRI in multiple sclerosis: triple vs. single dose. AB - We compared the sensitivity of single and triple dose Gd-DTPA magnetic resonance imaging (MRI) in detecting enhancing lesions in the spinal cord (SC) from 15 patients with multiple sclerosis (MS). The patients were examined monthly on four occasions. We detected two enhancing lesions in two of 15 (13%) patients when a single dose of Gd-DTPA was used. No additional lesions were detected when a triple dose of Gd-DTPA was used. These results 1) confirm that enhanced spinal cord imaging does not significantly increase the detection of active lesions in MS, 2) they do not support the general application of triple dose Gd-DTPA when examining the SC but 3) suggest that further studies taking into account SC symptoms are necessary. PMID- 11118775 TI - Dental health education: from education to informed decision making. PMID- 11118776 TI - The evolving concept of health in nursing research: 1988-1998. AB - This article concerns the evolution of the concept of health, as reported in selected nursing journals, over the last 10 years. It builds on an analysis reported by Reynolds [Reynolds CL. The measurement of health in nursing research. Adv Nurs Sci 1988;10(4):23-31.] who initially investigated the concept of health and the means by which it was measured during the period 1977-1987. Using the same journals as Reynolds, the methodology of systematic review is used to analyse the way in which health is defined, the frequency with which it is investigated and the means by which it is measured, and these data are compared with Reynolds findings. The results indicate that; three times as many studies have been conducted in the last 10 years, a more holistic concept of health has emerged, and the instruments used are becoming more sophisticated. These findings are discussed and the implications for patient education and health care professionals considered. PMID- 11118777 TI - The diffusion process of patient education in Dutch community pharmacy: an exploration. AB - Patient education activities in pharmacies are receiving much attention. These activities are relatively new and implementation requires individual and organisational change in pharmacies. The aim of this study is to identify barriers and facilitators to the implementation of patient education in community pharmacies and to classify these barriers and facilitators into the diffusion stages of Rogers' 'Innovations in Organisations' model [Rogers, EM. Diffusion of innovations. 4th ed. New York: The Free Press, 1995]. Six focus group interviews, three with pharmacists and three with pharmacy technicians (total n = 38) were carried out. The initiation phase has been dealt with by community pharmacies, whereas the implementation phase has not. The barriers and facilitators in the redefining/restructuring stage were mainly related to the organisation of patient education. In the clarifying and routinizing stages, barriers were related to repetition and knowledge transfer. The facilitators in these stages relate to performing and talking about patient education. Interventions for implementation of patient education should aim at these barriers and facilitating factors. PMID- 11118778 TI - Psychosocial and quality of life correlates of glycemic control during intensive treatment of type 1 diabetes. AB - To identify emotional and attitudinal barriers to improved glycemic control (HbA1c) during intensive diabetes treatment, 55 patients attending a 4-5 month intensive diabetes medical/education clinic were followed. Subjects completed a battery of psychological surveys, had HbA1c and body mass index measured, and rated their attitude toward weight gain and the extent of problems with specific self-management behaviors before and after the medical intervention. Although HbA1c improved on average, 29% had only modest improvement and 16% showed no improvement. The number of diabetes-related annoyances, worry about hypoglycemia, and diabetes-related emotional distress diminished. Only the satisfaction subscale of the Diabetes Quality of Life survey, diabetes-related emotional distress, and problems with self-management behaviors correlated with HbA1c. Treatment-related frustration and emotional distress may initially act as motivators to improve glycemia but can later become barriers to that goal. Interventions designed to help patients overcome attitudinal barriers should be incorporated into medical programs geared toward improving glycemia. PMID- 11118779 TI - Substitute decision-making: measuring individually mediated sources of uncertainty. AB - One aspect of O'Connor's Decisional Conflict Scale [O'Connor, A.M., Validation of a decisional conflict scale, Med. Decis. Making 15 (1995) 25-30] is the assessment of selected factors (perceived lack of information, undue social pressure, lack of support from others, and lack of clarity about personal values) that are believed to contribute to decisional uncertainty. This study explored the appropriateness of this uncertainty measure in the substitute decision-making context. Forty-nine mothers deciding on gastrostomy tube insertion for their children completed the scale, and also provided verbal reports about the contributory factors. For each of the four factors, relatively high-, moderate-, and low-scoring sub-groups were identified; then the associated verbal reports were examined for across-sub-group differences. Differences in verbal reports about information (chi 2 = 6.990, P = 0.0082), perceived pressure (chi 2 = 8.377, P = 0.0038), social support (chi 2 = 5.573, P = 0.0182), and perceived gains and losses (chi 2 = 3.85, P = 0.0499; chi 2 = 5.76, P = 0.0164) were observed, implying consistency between quantitative scores and verbal reports. This quantitative/qualitative hybrid approach may be clinically useful for assessing individually mediated factors contributing to decision uncertainty, and for evaluating therapeutic interventions in other substitute decision-making contexts. PMID- 11118780 TI - A qualitative study on detecting cancer symptoms and seeking medical help; an application of Andersen's model of total patient delay. AB - Patient delay is the interval between the day someone first becomes aware of an unexplained symptom and the day they seek medical consultation. This pre diagnostic period is comprised of several stages which may involve delay on the part of the individual. This study investigated factors influencing the process of detecting cancer symptoms and consulting a general practitioner (GP). Twenty three patients were interviewed about their experiences during this process. Among factors stimulating the process of detection and consultation were associating symptoms with cancer, and discussing symptoms with others. Being ashamed or embarrassed about the symptoms and attributing symptoms to common ailments were among the impeding factors. The findings of the present study suggest that future health education on early detection of cancer should focus on increasing knowledge and providing positive information about early detection of cancer. It is recommended that educational materials be disseminated to the general public via more channels, including non-medical channels. PMID- 11118781 TI - Difficulties of diabetic patients in learning about their illness. AB - The aim of this report is to shed light on the difficulties experienced by diabetic patients in learning about their illness. One hundred and thirty-eight diabetic people (97 IDD and 41 NIDD) were questioned at two survey locations, one national (63) and one regional (75), by means of a closed answer questionnaire. One hundred and four (75%) had attended a formal programme of diabetes education. They were asked which points in their diabetes education they had best understood and which they had least understood. The main results show that, globally, they easily acquire the manual skills. Conversely, numerous learning difficulties are associated with the skills required to solve problems and make decisions, such as adaptation of doses of insulin. These results are comparable to those obtained in a previous study in which professional carers were asked about their difficulties in educating their patients. PMID- 11118782 TI - The effects on knowledge of the systematic education of patients with joint diseases treated with NSAIDs and diuretics. AB - In a randomised, controlled trial, patients with joint diseases and concomitant treatment with NSAIDs and diuretics received systematic education. The intervention group was given information by a self-conducted, interactive Kodak Photo-CD program in addition to personal drug information and non-commercial drug leaflets. Awareness of drug interactions and encouragement of self-adjustment of treatment was focused on. Control patients received conventional information. Three months after randomisation, knowledge was tested by means of a questionnaire. At 3 months there was a significant difference in attained score between the intervention group and the control group. Greater knowledge was achieved, especially on drug interaction, in the intervention group. In conclusion, less than 1 h of systematic education significantly improved patients' knowledge on essential issues of concomitant treatment with NSAIDs and diuretics. Knowledge of effects, side-effects and interactions of drugs is essential for self-adjustment of treatment. The method employed, which is standardised and produces a reproducible quantity of education, might be applicable to several other medical conditions. PMID- 11118783 TI - Readiness to adopt adequate postural habits: an application of the Transtheoretical Model in the context of back pain prevention. AB - Based on a biomechanical model, an adequate body posture can contribute to the prevention of back pain and back pain chronicity. This study examines the explanatory value of the Transtheoretical Model (TTM) for the adoption of adequate postural habits in a cross-sectional sample of 149 employees of a German administration unit (mean age 40.2 years, 50% female). Using newly developed instruments with satisfactory psychometric properties, basic assumptions of the TTM could be confirmed: self-efficacy and the perceived pros for maintaining a good body posture increased significantly across the stages, while the perceived cons decreased. Additionally, the use of preventive strategies for back pain prevention increased linearly and significantly across the stages of change. The study supports the applicability of the TTM for postural behavior. Considering stages of change as an intervening variable may contribute to clarifying the relationship between participation in low back schools and prevention of back pain chronicity. Longitudinal and intervention study data are needed to support these assumptions. PMID- 11118784 TI - Informed consent in Dutch dental practice: knowledge, attitudes and self-efficacy of dentists. AB - The introduction of the 'Medical Treatment Contract Act' in the Netherlands 5 years ago, established some major rights and duties of both patients and members of the medical profession. The aim of this study was to assess Dutch dentists' knowledge, attitudes and self-efficacy with regard to several topics of this act. A questionnaire was sent to 806 dental practitioners; 41.6% of them responded. Results show that dentists are well informed about some of the most important topics of this law, such as the requirement to obtain the patient's consent to major dental treatments. The results concerning their attitude and self-efficacy, however, are less positive. A majority of the respondents believe that the principle of informed consent will lead to a commercialization of the dentist patient relation. Also, fear for legal procedures and difficulty with what patients must be informed about appears present. It is concluded that improvement of the implementation of the requirements of this legislation in dental practice requires both a change in attitude and an enhancement of the communicative skills of dentists. PMID- 11118785 TI - Asthma leaflets for patients: what do asthma nurses use? AB - This study investigated patient information leaflets used by trained asthma nurses, and nurses' satisfaction with these. Main outcome measures were frequency of use, sources of material, and rating of reliability and readability. A total of 775 practice nurses with a diploma in asthma working in the general practice setting were surveyed using a postal questionnaire. Forty two percent of questionnaires (326) were returned. The provision of asthma information is an integral part of patient care by the trained asthma nurse. Most nurses (260, 83%) gave out between one and ten leaflets per week. An abundance of diverse information is available from a variety of sources, main sources used were Glaxo Wellcome (cited by 47% of respondents), National Asthma Campaign (NAC) (19% of respondents) and Astra (19%). Pharmaceutical company material was considered more easily available, free of charge and more attractively presented. NAC material was viewed as more accurate. Assessment of NAC, Glaxo-Wellcome and Astra booklets revealed that they conformed to British Thoracic Society Guidelines for acute asthma. Thus, nurses stated a preference for charity information but in practice used a predominance of pharmaceutical company information. This appears to be because pharmaceutical company information was more readily available and free of charge. Nurses felt unease with self-perceived over-reliance on pharmaceutical company information. PMID- 11118786 TI - Feed intake and serum GH, LH and cortisol in gilts after intracerebroventricular or intravenous injection of urocortin. AB - In three experiments (Exp), ovariectomized gilts received intracerebroventricular (ICV; Exp 1 - with restraint, Exp 2 - without restraint) or intravenous (i.v.; Exp 3) injections of urocortin or saline to assess effects on feed intake and serum GH, LH, and cortisol. Following a 20-hr fast, feed was presented at 1 hr (Exp 1) or 30 min (Exp 2 and 3) after injection (time = 0 hr) of saline or 5 (U5) or 50 (U50) microg/pig (Exp 1 and 2) or 5 microg/kg BW (Exp 3) of urocortin. Blood samples were collected every 15 min from -2 to 6 hr relative to injection and hormone data pooled 2 hr before and hourly after treatment. Treatment with U50 decreased feed intake, relative to saline (treatment x time interaction; P < 0.05), when delivered ICV but not i.v. A treatment by time interaction was detected for GH (Exp 1, 2, 3) and LH (Exp 1 and 2; P < 0.01). Serum GH increased over time (relative to -2 hr; P < 0.05) following treatment with urocortin but not saline regardless of route of administration. Conversely, in Exp 1 (U5 and U50) and Exp 2 (U50), LH decreased relative to -2 hr with a delayed decrease during Exp 1. Serum cortisol was not affected by treatment in Exp 1, but increased following urocortin in Exp 2 and 3 (treatment by time interaction, P < 0.01). These data provide evidence that urocortin modulates GH and LH concentrations and suppresses feed intake in gilts via mechanisms which may be independent of cortisol and may depend upon dose and route of administration. PMID- 11118787 TI - Tumor necrosis factor-alpha and nitrite/nitrate responses during acute mastitis induced by Escherichia coli infection and endotoxin in dairy cows. AB - Concentrations of tumor necrosis factor-alpha (TNF-alpha) and of NO(x) (sum of nitrite and nitrate as indicators of endogenous nitric oxide production) in milk and blood plasma were measured in three mastitis models in dairy cows in early lactation. Escherichia coli P4:O37 bacteria or endotoxin O111:B4 were administered into both left quarters of 12 and 6 cows, respectively. Six of the E. coli-infected cows were treated with a bactericidal antibiotic (Enrofloxacin; Bayer AG, Leverkusen, Germany) i.v. at 10 hr and subcutaneously (sc) at 30 hr after infection. NO(x) concentrations transiently increased maximally 10- to 11 fold in milk of E. coli-infected quarters with or without antibiotic treatment at 24 hr and after endotoxin administration. NO(x) concentrations did not change in milk of unchallenged quarters and in blood plasma. Increases of NO(x) were proceeded by a transient (96- to 149-fold) rise of milk TNF-alpha concentrations, which in endotoxin-administered quarters was maximal at 6 hr and in infected quarters without or with Enrofloxacin treatment at 10 and 14 hr. In blood plasma TNF-alpha concentrations only moderately increased to peaks in endotoxin administered cows at 6 hr and in E. coli-infected cows at 14 hr postchallenge. In one severely sick, nontreated E. coli-infected cow milk, TNF-alpha response at 14 hr was excessive and followed by a spectacular rise of NO(x) concentration in milk between 48 and 72 hr. In conclusion, a possible clinical relevance of nitric oxide production associated with a rise of intramammary and systemic TNF-alpha during acute mastitis by E. coli infection and endotoxin in lactating dairy cows is indicated, but could not be inhibited by antibiotic treatment. PMID- 11118788 TI - Effect of oxytocin on expression of cytosolic phospholipase A2 mRNA and protein in ovine endometrial tissue in vivo. AB - The induction of endometrial prostaglandin (PG) F2alpha synthesis by oxytocin is dependent upon activation of phospholipase (PL) A2 and mobilization of arachidonic acid. The objective of this study was to determine if oxytocin stimulates PGF2alpha synthesis by inducing synthesis of cytosolic PLA2 (cPLA2). In Experiment 1, 15 ovariectomized ewes were given progesterone and estradiol to simulate an estrous cycle. Ewes were then given an injection of oxytocin on Day 14 of the simulated estrous cycle. Jugular blood samples were collected and assayed for 13,14-dihydro-15-keto-prostaglandin F2alpha (PGFM). Uteri were collected at 0, 7.5, 25, 90, or 240 min postinjection (n = 3 ewes/time point). Total RNA was isolated from caruncular endometrium and subjected to dot-blot analysis. Oxytocin induced a rapid and transient increase in serum PGFM (P < 0.01). However, endometrial concentrations of cPLA2 mRNA did not change following oxytocin administration (P > 0.10). In Experiment 2, 11 ovary-intact ewes were given oxytocin (n = 5) or saline (n = 6) on Day 15 after estrus. Jugular blood samples were collected and assayed for serum concentrations of PGFM. Uteri were collected at 15 min postinjection. Homogenates were prepared from caruncular endometrium and subjected to Western blot analysis. Concentrations of PGFM were higher in oxytocin treated ewes compared to saline treated ewes at 15 min postinjection (P < 0.01). Endometrial concentrations of cPLA2 protein were greater in the cytosolic than in the microsomal fraction (P < 0.01). Oxytocin did not affect the amount of cPLA2 protein in either fraction (P > 0.10). In conclusion, oxytocin did not effect expression of either cPLA2 mRNA or protein in ovine endometrium. Oxytocin may stimulate PGF2alpha synthesis by activating cPLA2 protein that is already present in an inactive form. PMID- 11118790 TI - Influence of light environment and photoperiod on plasma melatonin and cortisol profiles in young domestic boars, comparing two commercial melatonin assays. AB - Eighteen crossbred boars, three siblings from each of six litters, were distributed randomly among three groups after weaning. The 'Natural long-day' group was housed in a standard room with windows, whereas the 'Artificial long day' and 'Artificial short-day' groups were housed in light-sealed rooms and under an artificial light regimen (1400 lx). In spring (16-17 hr of light) plasma levels of melatonin and cortisol were measured in samples collected hourly for 24 hr. Two commercial melatonin radioimmunoassays with preassay diethyl ether extraction were compared. Only the assay from Buhlmann Laboratories AG showed low to undetectable melatonin levels during the light-phase and was used for further analysis. Dark-phase melatonin concentrations were higher than light-phase melatonin concentrations (P < 0.001). Dark-phase melatonin concentrations were higher in the 'Natural long-day' group than in the 'Artificial long-day' and the 'Artificial short-day' groups (P < 0.001). Sire had a significant effect on dark phase melatonin concentrations (P < 0.01), but not on light-phase levels. Cortisol concentrations were higher during the light-phase than during the dark phase, and light-phase cortisol concentrations were higher in the 'Natural long day' group than in the 'Artificial long-day' and the 'Artificial short-day' groups (P < 0.01). This study showed that peripubertal boars express a typical circadian melatonin rhythm under both artificial light regimens and in standard pig stable environment. Natural photoperiod and indoor lighting seem to interact in shaping the melatonin profile in standard stable environment. The great individual variation in the amplitude of the dark-phase melatonin levels could in this study be explained by the different sires. PMID- 11118789 TI - Effects of weaning on somatotrophic gene expression and circulating levels of insulin-like growth factor-1 (IGF-1) and IGF-2 in pigs. AB - The present study evaluated somatotrophic gene expression in liver, muscle and adipose tissue 4 d after weaning, a time point corresponding to greatly reduced serum concentrations of insulin-like growth factor (IGF)-1 and IGF-2 in pigs. Two week-old barrows were either cross-fostered to a sow (SOW, n = 8) or weaned and fed a phase 1 diet containing either 0 or 7% spray-dried plasma (NP, n = 8 and SDP, n = 8; respectively). Piglets were allocated such that two size groups were equivalently represented in each experimental group (small, 3.5-4.3 kg and large, 4.6-5.7 kg). Animals were weighed daily and sacrificed 4 d after weaning for blood and tissue collection. Daily gains of the SOW piglets were significantly greater than those of the weaned pigs for the first 3 d of the experiment (P < 0.0001). Weight gains in the SOW and SDP pigs between d 3 and 4 were equivalently elevated relative to the NP pigs (P < 0.0001). Serum IGF-1 and IGF-2 concentrations were decreased in both NP and SDP compared to SOW (P < 0.0001). Serum IGF-2 levels were significantly lower in small piglets (P = 0.006). A Weaning Group X Size interaction was noted for liver IGF-2 mRNA (P < 0.03), reflecting a higher level of expression in large SOW piglets relative to small SOW piglets. Weaning did not affect IGF-1, IGF-2, or growth hormone (GH) receptor mRNA levels in liver, muscle, or fat (P > 0.05). Liver IGF-binding protein (IGFBP)-3 and acid-labile subunit (ALS) mRNA levels also were unaffected by weaning. Small pigs had lower levels of liver ALS (P = 0.0003), muscle IGF-2 (P = 0.02), and muscle GH receptor (P = 0.006) mRNAs. In contrast, adipose tissue IGF 1 and IGF-2 mRNA levels were greatest in the small piglets (P = 0.001 and 0.029, respectively). PMID- 11118791 TI - Concentrations of leptin in serum and milk collected from lactating sows differing in body condition. AB - Leptin concentrations in the circulation and milk were determined in sows that differed in body condition at farrowing, and in feed consumption during lactation. Serum concentrations of leptin at farrowing and weaning were highest in sows exhibiting the greatest amount of backfat. Leptin was detected in both skim and whole milk throughout lactation, but levels were not correlated with backfat thickness or circulating leptin concentrations. This report provides the first evidence for the presence of leptin in sow milk; its function in the physiology of suckling pigs remains to be determined. PMID- 11118792 TI - MRI of disseminated developmental dysmyelination in Fukuyama type of CMD. AB - Whether the pathologic origin of white matter lesions in Fukuyama type of congenital muscular dystrophy (FCMD) is delayed myelination or dysmyelination is a controversial issue. This study investigated pathologic distribution in white matter with heavily T(2)-weighted images using fluid-attenuated inversion recovery (FLAIR) pulse sequence. For detection of abnormal white matter lesions, FLAIR images were approximately twice as sensitive as T(2)-weighted images and five times as sensitive as T(1)-weighted images of spin echo sequence. The distribution of the white matter lesions was disseminated and not correlated with cortical disarrangement. The distribution was not consistent with delayed myelination. These findings support the evidence found using in vitro proton-NMR spectroscopy that the pathologic origin of white matter lesions is dysmyelination. When conventional magnetic resonance imaging is used, masked white matter lesions are easy to misidentify as delayed myelination instead of disseminated developmental dysmyelination. The lesions in the white matter of FCMD are masked because of brain development. PMID- 11118793 TI - Cognitive and behavioral problems in children with centrotemporal spikes. AB - Atypical features in benign epilepsy of childhood with centrotemporal spikes (BECTS) are not uncommon. There are children with BECTS who do not have a benign outcome in terms of neuropsychologic functioning. BECTS have been linked with Landau-Kleffner syndrome (LKS) and continuous spikes and waves during slow sleep (CSWS). At the Medical College of Georgia from January 1988 to June 1999, 78 children, ages 2-16 years, were identified to have electroencephalogram evidence of centrotemporal spikes. Their medical records were reviewed for developmental history, behavioral problems, and school performance. Children with structural lesions/other epileptic syndromes were excluded. Fifty-six demonstrated a history of clinical seizures compatible with BECTS and 22 demonstrated centrotemporal spikes without clinical seizures. Among all children with centrotemporal spikes, 9% (n = 7) were diagnosed with mild intellectual disability (intelligence quotient < 70), 10% (n = 8) with borderline functioning, 31% (n = 24) with behavioral problems, and 17% (n = 13) with specific learning disabilities. Three children with BECTS experienced language delay and regression. Seizure control for BECTS usually is achieved without much difficulty, with excellent long-term prognosis. However, the data presented indicate that a large number of BECTS patients exhibit learning or behavior problems that require intervention. A small number may demonstrate language outcome similar to children with LKS and CSWS. PMID- 11118794 TI - Effects of antiepileptic drugs on evoked potentials in epileptic children. AB - To evaluate the visual and auditory function in children and adolescents who are undergoing monotherapy with sodium valproate, carbamazepine, and phenobarbital visual-evoked potentials and brainstem auditory-evoked potentials were measured in 58 epileptic patients (30 males and 28 females), ages 13.7 +/- 6.9 years. Fifty healthy sex- and age-matched children served as controls. The measurements were performed before the beginning of therapy and after 12 months. Before the beginning of therapy, there were no significant differences in visual- and auditory-evoked potentials between the control group and the three groups of epileptic children. After 12 months of therapy, patients treated with carbamazepine demonstrated a significant (P < 0.001) increase of P100 latencies when compared with baseline data and control values; moreover, these patients exhibited a significant increase of peak latencies of waves I-III-V and interpeak interval I-V at auditory second evaluation. The patients treated with sodium valproate manifested a significant (P < 0.05) increase in VEP P100 latencies. On the contrary, children receiving phenobarbital did not manifest any significant abnormality at visual- and auditory-evoked potentials measurements. Our study demonstrates that for patients treated with carbamazepine and sodium valproate, an electrophysiologic dysfunction of visual and auditory sensory pathways can be present after 12 months of treatment. PMID- 11118795 TI - Steroids or vigabatrin in the treatment of infantile spasms? AB - In many countries, vigabatrin is now recommended as the first choice of treatment for infantile spasms instead of steroids. The aim of this study was to review the efficacy and side effects of the two drugs, steroids and vigabatrin, by using data from published series. Results suggest that vigabatrin certainly is efficacious in the treatment of the disorder but, on the whole, it does not seem to be any more effective than steroids, especially corticotrophin, even in children with tuberous sclerosis. The possible benefits of vigabatrin do not justify the risks of the possible irreversible visual changes associated with vigabatrin. PMID- 11118796 TI - Cerebral cortical dysplasia: assessment by MRI and SPECT. AB - The objective of this study was to establish correlations between image findings and pathologic deficits in patients with cerebral cortical malformations. The results of magnetic resonance imaging (MRI) and single-photon emission computed tomography (SPECT) in addition to clinical data for 15 patients with cerebral cortical malformations were reviewed retrospectively. MRI led to the diagnoses of bilateral perisylvian syndrome, hemimegalencephaly, focal polymicrogyria, band heterotopia, and focal cortical dysplasia (FCD). Interictal SPECT did not reveal hypoperfusion in any case of polymicrogyria. Ictal SPECT images revealed hyperperfusion of the lesion in three patients with polymicrogyria, with accompanying hyperperfusion of the basal ganglia in two of the three patients. On the other hand, interictal SPECT images demonstrated hypoperfusion of the lesion in four patients with FCD. Ictal SPECT images revealed hypoperfusion of the lesion in two patients, hyperperfusion of the lesion in one patient, and hypoperfusion of the basal ganglia in two patients with FCD. This difference in perfusion between polymicrogyria and FCD observed in this study may reflect histologically different characteristics. This relative hyperperfusion of the cortex and the basal ganglia observed on ictal SPECT, which was found in two polymicrogyria patients with complex partial seizures and partial seizures evolving to secondary generalized seizures, respectively, suggests that the cortical-subcortical interaction is related to the mechanism of loss of consciousness or seizure generation. PMID- 11118797 TI - Single-photon emission computed tomography of the brain in autism: effect of the developmental level. AB - Brain single-photon emission computed tomography was performed in 22 autistic and 10 nonautistic disabled patients. The regional cerebral blood flow in both laterotemporal and dorso-medio-lateral frontal areas decreased significantly in the autistic group compared with in nonautistic group. In the autistic group, the regional cerebral blood flow was significantly higher in the right temporal and right parietal lobes than that in the left ones. Inversely, the regional cerebral blood flow in the frontal and occipital lobes was significantly higher on the left side than on the right side. In the nonautistic group, except for in the dorso-medio-lateral frontal lobes (left > right), there was no difference in the regional cerebral blood flow in either cerebrum or cerebellum. A positive correlationship between regional cerebral flow and development quotient (intelligence quotient) was observed in the left laterotemporal and both dorso medio-lateral frontal areas, and a negative one was observed in the cerebellar vermis area. These results suggest that the regional cerebral blood flow decrease in the temporal and frontal areas relates to not only the brain mechanism of autism reported previously but also intelligence levels. PMID- 11118798 TI - Shuddering attacks-report of four children. AB - Shuddering attacks are recognized as an uncommon benign disorder occurring during infancy or early childhood. It is necessary to distinguish these episodes from epileptic seizures. The attacks seem to involve shivering movements occurring daily for several seconds without impairment of consciousness. According to the criteria for benign myoclonus of early infancy, both shuddering attacks and benign myoclonus of early infancy should be regarded as having the same nosologic entity. I studied the pathophysiology of shuddering attacks in four children between 8 and 14 months of age using a video-electroencephalographic monitoring system. In one patient the frequency of shuddering movements, which was read as contamination of the electromyography on electroencephalography during attacks, seemed to be almost the same as that as of essential tremor. Shuddering attacks have decreased in number or disappeared in all four patients, but one exhibited mild abnormalities on magnetic resonance imaging and had relatives with epilepsy, and another had a flattened sella turcica. Although previous reports suggest that these movements are benign and needless investigations should be avoided, a problem related to the development of the nervous system may be present in children with shuddering attacks. PMID- 11118799 TI - Cerebral infarction in Menkes' disease. AB - Menkes' disease is an X-linked disorder caused by impaired intracellular transport of copper. Currently, no therapy effectively arrests the relentless neurodegeneration of Menkes' disease. Previous neuroimaging reports of patients with Menkes' disease describe a range of abnormalities, including intracranial vessel tortuosity and cerebral white matter changes. We report two infants with Menkes' disease who developed ischemic cerebrovascular disease early in infancy. Magnetic resonance studies, including diffusion-weighted imaging and proton magnetic resonance spectroscopy, demonstrated bilateral infarctions of deep gray matter nuclei, a finding not previously described in Menkes' disease. Potential mechanisms for these cerebrovascular lesions in Menkes' disease include the susceptibility to free radical attack and inadequate energy supply from oxidative phosphorylation. These infarctions may play an unrecognized but important role in the neurodegeneration of children with Menkes' disease. The development of effective therapeutic agents against this disease will require a more detailed understanding of such underlying mechanisms. PMID- 11118800 TI - SPECT in focal enterovirus encephalitis: evidence for local cerebral vasculitis. AB - We report a 4-year-old, left-handed male with focal coxsackievirus A3 encephalitis who presented with seizures and acquired aphasia. Electroencephalography exhibited focal spike discharges over the right frontal regions, but cranial magnetic resonance imaging did not reveal any structural abnormalities. However, brain single-photon emission computed tomography performed during the acute phase disclosed focal hypoperfusion in the right frontal lobe, consistent with decreased regional cerebral blood flow in the territory of some branches of the right cerebral anterior artery. Without specific treatment, the patient recovered completely within 1 month, when brain single-photon emission computed tomography images returned to normal and cranial magnetic resonance imaging still demonstrated no abnormalities. The present case suggests the possible role of transient local cerebral vasculitis in the pathogenesis of focal enterovirus encephalitis. PMID- 11118801 TI - Periodic alternating nystagmus in two children with a similar, unusual phenotype. AB - The report describes two unrelated male children, aged 6 and 8 years, respectively, with congenital periodic alternating nystagmus, congenital strabismus, microcephaly with cortical and cerebellar hypoplasia, mental retardation, low stature, and bat ears. Karyotypes were normal. Neuropediatric and ophthalmologic examinations, radiologic imaging of the brain, and laboratory analyses were performed to exclude other causes of periodic alternating nystagmus, such as ataxia-telangiectasia, acquired disease of the caudal brainstem or the cerebellum, albinism, or loss of vision resulting from cataract or vitreous hemorrhage. The similar morphologic and clinical features of both patients raise the possibility that they have an identical syndrome. PMID- 11118802 TI - Motor-evoked potentials in a child recovering from transverse myelitis. AB - We describe a 13-year-old female, with an inability to walk because of transverse myelitis, who demonstrated progressive recovery of both motor function and motor- evoked potentials (MEP). At 4 weeks after onset, amplitudes of MEP were decreased, latencies were prolonged, and cortical somatosensory-evoked potentials (SEP) were absent. At 6 and 12 weeks, MEP revealed progressively higher amplitudes and shorter latencies. SEP also recovered. MEP and SEP recovery paralleled clinical recovery. MEP in response to transcranial magnetic stimulation may provide guidance regarding recovery from spinal cord disorders in children. PMID- 11118803 TI - Antiepileptic drug-induced visual hallucinations in a child. AB - The neurologic signs and symptoms of carbamazepine and phenytoin toxicity, such as ataxia, dysarthria, and nystagmus, are well known. The psychiatric manifestations of toxicity, such as psychosis and hallucinations, however, are less widely recognized. This study reports the case of a 9-year-old male with seizures who developed intermittent complex visual hallucinations after therapy with antiepileptic drugs was begun. This study considered seizures, migraine, underlying psychiatric diathesis, and drug toxicity as possible etiologies but after extensive investigation concluded that his symptoms were most likely a drug side effect. PMID- 11118804 TI - A case of 3-methylglutaconic aciduria misdiagnosed as cerebral palsy. AB - 3-Methylglutaconic aciduria is a rare hereditary metabolic disorder characterized by increased urinary excretion of 3-methylglutaconic and 3-methylglutaric acids. Four clinical forms are recognized. This study presents the case of a 5-year-old male with type IV 3-methylglutaconic aciduria, initially diagnosed as "static encephalopathy." The slow evolution and other clinical characteristics, together with cerebral magnetic resonance imaging (MRI) findings, eventually directed the diagnosis to organic aciduria that was confirmed by urine test. This study proposes that the clinical criteria for childhood cerebral palsy should be rigorously respected; neuroimaging studies, particularly MRI, should be conducted to confirm the diagnosis, especially in atypical cases. PMID- 11118805 TI - Familial moyamoya disease in Caucasians. AB - There are few reports of moyamoya disease (MMD) in the Caucasian population and even fewer descriptions of the natural history of the disease. The study reports a 12-year follow-up of two white male siblings with MMD. Although both brothers had a persistently abnormal electroencephalogram and a learning disorder, the siblings recovered on aspirin and antiepileptic therapy with minimal neurologic residua. The occurrence of MMD in these siblings contributes to the evidence that MMD has a hereditary basis in Caucasians, as well as the Japanese. Even with the use of conservative measures, children may still have persistent yet minor cerebrovascular insults. In addition to the two patients discussed, this study reviewed the literature on all cases of MMD reported within families. Future follow-up studies are required to determine the natural history and the appropriate medical and surgical management of MMD. PMID- 11118806 TI - The distribution of neuropeptide Y-immunoreactive neurons and nerve fibers in the forebrain of the carp Cyprinus carpio L. AB - The present study reports the distribution of Neuropeptide Y (NPY)-immunoreactive neurons and fibers in the forebrain of the adult carp Cyprinus carpio L. Serial Nissl-stained sections were used for cytoarchitecture and identification of anatomical structures. Immunostaining of NPY-containing neurons and fibers was used as neurochemical marker and tool for comparison with other species, including the goldfish. The general outline of the cytoarchitecture of the carp forebrain is similar to that of other Cypriniformes. However, using NPY immunohistochemistry, we found several specific differences with the goldfish, especially in the diencephalon. In the hypothalamus of the carp NPY immunoreactive (NPYir) neurons were identified in the n. dorsolateralis thalami, and in the n. ventralis lateralis thalami. In the same location, we observed the n. anterior hypothalami and the n. preglomerulosus pars lateralis, described in the goldfish, as parts of n. prerotundus. However, in the carp we were not able to identify a n. preglomerulosus pars medialis, a n. preglomerulosus pars medialis commissuralis and a n. glomerulosus. We describe a n. rotundus, in which we did not find substructures typical of the goldfish. Further differences with the goldfish, trout and salmon were also noted. PMID- 11118807 TI - Distribution of P2X1, P2X2, and P2X3 receptor subunits in rat primary afferents: relation to population markers and specific cell types. AB - We determined the co-expression of immunoreactivity (IR) for ATP-receptor subunits (P2X1, P2X2, and P2X3), neuropeptides, neurofilament (NF), and binding of the isolectin B(4) from Griffonia simplicifolia type one (GS-I-B(4)) in adult dorsal root ganglion neurons. P2X1-IR was expressed primarily in small DRG neurons. Most P2X1-IR neurons expressed neuropeptides and/or GS-I-B(4)-binding, but lacked NF-IR. P2X1-IR overlapped with P2X3-IR, though each was also found alone. P2X2-IR was expressed in many P2X3-IR small neurons, as well as a group of medium to large neurons that lacked either P2X3-IR or GS-I-B(4)-binding. A novel visible four-channel fluorescence technique revealed a unique population of P2X2/3-IR neurons that lacked GS-I-B(4)-binding but expressed NF-IR. Co expression of P2X1, and P2X3 in individual neurons was also demonstrated. We examined P2X subunit-IR on individual recorded neurons that had been classified by current signature in vitro. Types 1, 2, 4 5, and 7 expressed distinct patterns of P2X-IR that corresponded to patterns identified in DRG sections, and had distinct responses to ATP. Types with rapid ATP currents (types 2, 5, and 7) displayed P2X3-IR and/or P2X1-IR. Types with slow ATP currents (types 1 and 4) displayed P2X2/3-IR. Type 1 neurons also displayed P2X1-IR. This study demonstrates that the correlation between physiological responses to ATP and the expression of particular P2X receptor subunits derived from expression systems is also present in native neurons, and also suggests that novel functional subunit combinations likely exist. PMID- 11118808 TI - Unilateral upregulation of cyclooxygenase-2 following cerebral, cortical photothrombosis in the rat: suppression by MK-801 and co-distribution with enzymes involved in the oxidative stress cascade. AB - Cyclooxygenase-2 (COX-2) is an essential enzyme for prostaglandin synthesis from arachidonic acid, during which considerable amounts of superoxide are produced. During pathological conditions, superoxide and nitric oxide (NO) rapidly form peroxynitrite, a potent cytotoxin, causing symptoms referred to as oxidative stress response. Superoxide is controlled by enzymes such as manganese- or copper zinc-dependent superoxide dismutase (Mn-SOD, CuZn-SOD), glutathione peroxidase (GPx) and antioxidants derived from heme oxygenase (HO) activity such as biliverdin and bilirubin. NO derives from 3 NO-synthases (NOS I-III) from which the calcium-dependent NOS-I and III are activated rapidly due to hyperexcitation. We studied the induction of COX-2 by immunohistochemistry at days 1, 2 and 5 following cortical photothrombosis in normal and MK-801 treated rats. The results showed a weak constitutive, neuronal expression of COX-2 in cortex and amygdala. Layers II+III contained considerably more COX-2 than infragranular layers. One and 2 days following injury COX-2 was highly upregulated in the supragranular layers of the whole injured hemisphere compared with sham-operated animals and compared to the contralateral unlesioned hemisphere, whereas at day 5 COX-2 levels had returned to baseline. MK-801 treatment caused a reduction in COX-2 upregulation at day one and by day 2 no significant differences between injured and contralateral hemisphere were measurable. COX-2 positive neurons were found in close association with NOS-I containing neurons and their fibers but were not colocalized. In addition, codistribution of COX-2 was found with HO-1, CuZn-SOD and GPx containing cells, whereas COX-2 was colocalized with HO-2 and/or MnSOD in cortical neurons. PMID- 11118809 TI - Low and infrequent expression of nitric oxide synthase/NADPH-diaphorase in neurons of the human supraoptic nucleus: a histochemical study. AB - The gas nitric oxide is a messenger in brain signaling. In the hypothalamo hypophyseal system nitric oxide is involved in the control of the expression and/or release of peptide hormones (corticotropin-releasing hormone, gonadotropin releasing hormone, vasopressin and oxytocin). Nitric oxide synthase (NOS), the enzyme generating nitric oxide, is abundantly present in the magnocellular nuclei of the rat hypothalamus. Its localization in the human hypothalamus is less well studied. Hence, we investigated the anatomical distribution of neuronal nitric oxide synthase in the human supraoptic nucleus by use of immunohistochemical and enzyme histochemical techniques. The immunohistochemical localization of NOS was studied in 31 matched human hypothalami (13 control cases, eight depressed patients and ten schizophrenics). NADPH-diaphorase studies were carried out on seven additional hypothalami (three normal brains, four schizophrenics). Apparent inter-individual differences exist with regard to the occurrence of the enzyme in supraoptic neurons. In a majority of cases no immunostaining or histochemical reaction for the enzyme was observed. In seven cases (three controls, two schizophrenics, two depressives) a population of nitrergic nerve cells was seen in the dorsomedial part of the nucleus. This group of cells also stained for NADPH-diaphorase. Also, there were a few NOS-immunopositive neurons scattered throughout the nucleus. Additionally, thin NADPH-diaphorase positive fibers were observed to cross the nucleus. Our data show that, unlike the rat, the human supraoptic nucleus contains only a small number of nitrergic neurons. No correlation was found between the expression of the enzyme in supraoptic neurons and the psychiatric status of the patients. PMID- 11118810 TI - Nitric oxide producing neurones in the rat medulla oblongata that project to nucleus tractus solitarii. AB - The production of nitric oxide in neurones of the rat medulla oblongata that project to the nucleus tractus solitarii (NTS) was examined by simultaneous immunohistochemical detection of nitric oxide synthase (NOS) and of cholera toxin B-subunit (CTb), which was injected into the caudal zone of the NTS. Neurones immunoreactive for CTb and neurones immunoreactive for NOS were widely co distributed and found in almost all the anatomical divisions of the medulla. Dual labelled cells, containing both CTb and NOS immunoreactivities were more numerous ipsilaterally to the injection sites. They were concentrated principally in the more rostral zone of the NTS, raphe nuclei, dorsal, intermediate and lateral reticular areas, spinal trigeminal and paratrigeminal nuclei and the external cuneate and medial vestibular nuclei. Isolated dual-labelled neurones were also scattered throughout most of the divisions of the reticular formation. These observations indicate that many areas of the medulla that are known to relay somatosensory and viscerosensory inputs contain NOS immunoreactive neurones that project to the NTS, and may, therefore, contribute to the dense NOS immunoreactive innervation of the NTS. The release of nitric oxide from the axon terminals of these neurones may modulate autonomic responses generated by NTS neurones in relation to peripheral sensory stimuli, and thus ultimately regulate sympathetic and/or parasympathetic outflow. PMID- 11118811 TI - Nitric oxide and postangioplasty restenosis: pathological correlates and therapeutic potential. AB - Balloon angioplasty revolutionized interventional cardiology as a nonsurgical procedure to clear a diseased artery of atherosclerotic blockage. Despite its procedural reliability, angioplasty's long-term outcome can be compromised by restenosis, the recurrence of arterial blockage in response to balloon-induced vascular trauma. Restenosis constitutes an important unmet medical need whose pathogenesis has yet to be understood fully and remains to be solved therapeutically. The radical biomediator, nitric oxide (NO), is a natural modulator of several processes contributing to postangioplasty restenosis. An arterial NO deficiency has been implicated in the establishment and progression of restenosis. Efforts to address the restenosis problem have included trials evaluating a wide range of NO-based interventions for their potential to inhibit balloon-induced arterial occlusion. All types of NO-based interventions yet investigated benefit at least one aspect of balloon injury to a naive vessel in a laboratory animal without inducing significant side effects. The extent to which this positive, albeit largely descriptive, body of experimental data can be translated into the clinic remains to be determined. Further insight into the pathogenesis of restenosis and the molecular mechanisms by which NO regulates vascular homeostasis would help bridge this gap. At present, NO supplementation represents a unique and potentially powerful approach to help control restenosis, either alone or as a pharmaceutical adjunct to a vascular device. PMID- 11118812 TI - Phenolic antioxidants attenuate neuronal cell death following uptake of oxidized low-density lipoprotein. AB - Oxidative stress is implicated in neuronal loss associated with neurodegeneration such as in Parkinson's disease, Alzheimer's disease and age-related cognitive decline. Recent reports indicate that the consumption of flavonoid-rich fruits partly reverses the age-related neuronal and cognitive decline. In this study, cultured striatal neurons were exposed to oxidized lipids in the form of low density lipoprotein (oxLDL) as a model for the induction of oxidative injury, and the abilities of phenolic antioxidants, flavonoids and hydroxycinnamic acid derivatives, to attenuate this neuronal damage were examined. OxLDL was demonstrated to enter neuronal cells and to be capable of eliciting neurotoxicity in a dose- and time-dependent manner, inducing DNA fragmentation and cell lysis. Flavonoids exert protective effects, which appear to be related to specific structural characteristics, particularly relevant being those defining their reduction potentials and partition coefficients. In summary, these data suggest a possible role for flavonoids in reducing neurodegeneration associated with chronic disorders in which oxidative stress is implicated. PMID- 11118813 TI - Conjugation position of quercetin glucuronides and effect on biological activity. AB - Quercetin glycosides are common dietary antioxidants. In general, however, potential biological effects of the circulating plasma metabolites (e.g., glucuronide conjugates) have not been measured. We have determined the rate of glucuronidation of quercetin at each position on the polyphenol ring by human liver cell-free extracts containing UDP-glucuronosyltransferases. The apparent affinity of UDP-glucuronosyltransferase followed the order 4'- > 3'- > 7- > 3, although the apparent maximum rate of formation was for the 7-position. The 5 position did not appear to be a site for conjugation. After isolation of individual glucuronides, the inhibition of xanthine oxidase and lipoxygenase were assessed. The K(i) for the inhibition of xanthine oxidase by quercetin glucuronides followed the order 4'- > 3'- > 7- > 3-, with quercetin-4' glucuronide a particularly potent inhibitor (K(i) = 0. 25 microM). The glucuronides, with the exception of quercetin-3-glucuronide, were also inhibitors of lipoxygenase. Quercetin glucuronides are metabolites of quercetin in humans, and these compounds can retain some biological activity depending on conjugation position at expected plasma concentrations. PMID- 11118814 TI - A thrombocyte-induced myocardial dysfunction in the ischemic and reperfused guinea pig heart is mediated by reactive oxygen species. AB - In recent investigations, we could demonstrate that thrombocytes are able to contribute to ischemia- and reperfusion-induced injury of the heart. The aim of the current study was to investigate whether reactive oxygen species are responsible for induction of myocardial dysfunction under these conditions. Isolated, perfused, and pressure-volume work-performing guinea pig hearts were exposed to a 30-min low-flow ischemia (1 ml/min) and were reperfused (5 ml/min). Washed, homologous blood platelets were administered as a 1-min bolus (20,000 per microliter of perfusion buffer), either during the 15th minute of ischemia or in the first or fifth minute of reperfusion in the presence of thrombin (0.3 U/ml perfusion buffer)). The radical scavengers superoxide dismutase (SOD; 10 U/ml perfusate) and catalase (30 U/ml perfusate) were added during ischemia or in the first or fifth minute of reperfusion, respectively. Intracoronary platelet retention (in percentage of platelets applied) and recovery of EHW (postischemic EHW in percentage of preischemic EHW) were quantified. Ischemic and reperfused hearts with time-matched application of platelets but without administration of SOD or catalase served as controls. Interestingly, both administration of SOD during ischemia and in reperfusion significantly improved recovery of EHW (88.4 +/- 2%, 82. 6 +/- 1%, and 90 +/- 3%, respectively) as compared with the case of controls (56.2 +/- 3%, 42 +/- 2%, and 75 +/- 2%, respectively). Platelet retention, however, was not significantly influenced by administration of SOD during ischemia or reperfusion (26 +/- 2%, 31 +/- 2%, and 26 +/- 2%) compared with controls (30.5 +/- 3%, 33 +/- 2%, and 22 +/- 3%, respectively). Coadministration of catalase, on the other hand, exhibited some cardioprotective potential only in the first minute of reperfusion (recovery, 61% +/- 4%) as compared with the case of control (42 +/- 2%). We conclude that thrombocytes under conditions of ischemia and reperfusion are able to induce a myocardial dysfunction mediated by reactive oxygen species. Superoxide seems to play a major role in this respect. PMID- 11118815 TI - Specific oxidative alterations in vastus lateralis muscle of patients with the diagnosis of chronic fatigue syndrome. AB - Chronic fatigue syndrome (CFS) is a poorly understood disease characterized by mental and physical fatigue, most often observed in young white females. Muscle pain at rest, exacerbated by exercise, is a common symptom. Although a specific defect in muscle metabolism has not been clearly defined, yet several studies report altered oxidative metabolism. In this study, we detected oxidative damage to DNA and lipids in muscle specimens of CFS patients as compared to age-matched controls, as well as increased activity of the antioxidant enzymes catalase, glutathione peroxidase, and transferase, and increases in total glutathione plasma levels. From these results we hypothesize that in CFS there is oxidative stress in muscle, which results in an increase in antioxidant defenses. Furthermore, in muscle membranes, fluidity and fatty acid composition are significantly different in specimens from CFS patients as compared to controls and to patients suffering from fibromyalgia. These data support an organic origin of CFS, in which muscle suffers oxidative damage. PMID- 11118816 TI - Target cell-derived superoxide anions cause efficiency and selectivity of intercellular induction of apoptosis. AB - Transformed fibroblasts are specifically eliminated by their nontransformed neighbors through intercellular induction of apoptosis. This process depends on the number of nontransformed effector cells and on the local density of transformed target cells. Intercellular signalling is inhibited by SOD (a scavenger of superoxide anions), taurine (a scavenger of HOCl), 4-aminobenzoyl hydrazide (a mechanism-based inhibitor of peroxidase), DMSO (a hydroxyl radical scavenger), and two inhibitors of NO synthase. Therefore, selective apoptosis induction seems to be based on superoxide anion production by transformed cells, their spontaneous dismutation to hydrogen peroxide, and HOCl generation by a novel effector cell-derived peroxidase. HOCl then interacts with target cell derived superoxide anions to yield hydroxyl radicals. Due to the short diffusion pathway of superoxide anions, hydroxyl radical generation is confined to the intimate vicinity of transformed cells. In parallel, NO derived from effector cells interacts with superoxide anions of target cells to yield the apoptosis inducer peroxynitrite. Reconstitution experiments using transformed or nontransformed cells in conjunction with myeloperoxidase, HOCl, or an NO donor demonstrated that superoxide anions generated extracellularly by transformed cells participate in intercellular signalling and at the same time determine transformed cells as selective targets for intercellular induction of apoptosis. PMID- 11118817 TI - Roles of phosphate and an enoyl radical in ferritin iron mobilization by 5 aminolevulinic acid. AB - 5-Aminolevulinic acid (ALA), a heme precursor that accumulates in acute intermittent porphyria (AIP) and lead poisoning, undergoes enolization and subsequent iron-catalyzed oxidation at neutral pH. Iron is released from horse spleen ferritin (HoSF) by both ALA-generated O(2)(.-) and enoyl radical (ALA(z.rad)), which amplifies the chain of ALA oxidation (autocatalysis). Iron chelators such as EDTA, ATP, but not citrate, and phosphate accelerate this process and ALA-promoted iron release from HoSF is faster in horse spleen isoferritins containing larger amounts of phosphate in the core. ALA (+0.377 V versus standard hydrogen electrode) is less effective in releasing iron from ferritin than are thioglycollic acid, 6-hydroxydopamine, and N,N,N', N' tetramethyl-p-phenylenediamine. During electrochemical one electron oxidation of ALA in a nitrogen atmosphere, spin trapping experiments with 3,5-dibromo-4 nitrosobenzenesulfonic acid demonstrated the formation of a spin adduct characterized by a six line signal, indicating a secondary carbon-centered radical and attributed to a resonant ALA&z.rad; radical. Iron is also released in such anaerobic electrochemical oxidations of ALA in the presence of ferritin, suggesting that, in addition to O(2)(*-), ALA&z.rad; can promote iron mobilization from ferritin. Hence, ALA&z.rad; may amplify the metal-catalyzed oxidation of ALA, damaging ALA-accumulating cells and possibly contributing to the symptoms of porphyria. PMID- 11118818 TI - Glutathione depletion induces apoptosis of rat hepatocytes through activation of protein kinase C novel isoforms and dependent increase in AP-1 nuclear binding. AB - Treatment of isolated rat hepatocytes with the glutathione depleting agents L buthionine-S,R-sulfoximine or diethylmaleate reproduced various cellular conditions of glutathione depletion, from moderate to severe, similar to those occurring in a wide spectrum of human liver diseases. To evaluate molecular changes and possible cellular dysfunction and damage consequent to a pathophysiologic level of GSH depletion, the effects of this condition on protein kinase C (PKC) isoforms were investigated, since these are involved in the intracellular specific regulatory processes and are potentially sensitive to redox changes. Moreover, a moderate perturbation of cellular redox state was found to activate novel PKC isoforms, and a clear relationship was shown between novel kinase activation and nuclear binding of the redox-sensitive transcription factor, activator protein-1 (AP-1). Apoptotic death of a significant number of cells, confirmed in terms of internucleosomal DNA fragmentation was a possible effect of these molecular reactions, and was triggered by a condition of glutathione depletion usually detected in human liver diseases. Finally, the inhibition of novel PKC enzymatic activity in cells co-treated with rottlerin, a selective novel kinase inhibitor, prevented glutathione-dependent novel PKC up regulation, markedly moderated AP-1 activation, and protected cells against apoptotic death. Taken together, these findings indicate the existence of an apoptotic pathway dependent on glutathione depletion, which occurs through the up regulation of novel PKCs and AP-1. PMID- 11118819 TI - Free radical production and changes in superoxide dismutases associated with hypoxia/reoxygenation-induced apoptosis of embryonic rat forebrain neurons in culture. AB - Following hypoxia/reoxygenation (6h/96h), cultured neurons from the embryonic rat forebrain undergo delayed apoptosis. To evaluate the participation of oxidative stress and defense mechanisms, temporal evolution of intraneuronal free radical generation was monitored by flow cytometry using dihydrorhodamine 123, in parallel with the study of transcriptional, translational, and activity changes of the detoxifying enzymes Cu/Zn-SOD and Mn-SOD. Two distinct peaks of radical generation were depicted, at the time of reoxygenation (+ 27%) and 48 h later (+ 25%), respectively. Radical production was unaffected by caspase inhibitors YVAD CHO or DEVD-CHO, which prevented neuronal damage, suggesting that caspase activation is not an upstream initiator of radicals in this model. Cell treatment by vitamin E (100 microM) displayed significant neuroprotection, whereas the superoxide generating system xanthine/xanthine oxidase induced apoptosis. Transcript and protein levels of both SODs were reduced 1 h after the onset of hypoxia, but activities were transiently stimulated. Reoxygenation was associated with an increased expression (139%), but a decreased activity (21%) of the inducible Mn-SOD, whereas Cu/Zn-SOD protein and activity were low and progressively increased until 48 h post-hypoxia, when the second rise in radicals occurred. In spite of a temporal regulation of SODs, which parallels radical formation, oxidative stress might account for neurotoxicity induced by hypoxia. PMID- 11118820 TI - Hyperinsulinemia: the missing link among oxidative stress and age-related diseases? AB - Mounting evidence supports Harman's hypothesis that aging is caused by free radicals and oxidative stress. Although it is known that oxidant species are produced during metabolic reactions, it is largely unknown which factor(s), of physiological or pathophysiological significance, modulate their production in vivo. In this hypothesis paper, it is postulated that hyperinsulinemia may have such function and therefore promote aging, independently of elevations of glycemia. Hyperinsulinemia is secondary to impaired insulin stimulated glucose metabolism at the level of skeletal muscle (insulin resistance) and is seen in about one third of glucose tolerant humans following dietary carbohydrate intake. If other insulin-stimulated (or inhibited) pathways retain normal sensitivity to the hormone, hyperinsulinemia could, by its effects on antioxidative enzymes and on free radical generators, enhance oxidative stress. Other proaging effects of insulin involve the inhibition of proteasome and the stimulation of polyunsaturated fatty acid (PUFA) synthesis and of nitric oxide (NO). The hypothesis that hyperinsulinemia accelerates aging also offers a metabolic explanation for the life-prolonging effect of calorie restriction and of mutations decreasing the overall activity of insulin-like receptors in the nematode Caenorhabditis elegans. PMID- 11118821 TI - Plasma glutamine response to enteral administration of glutamine in human volunteers (free glutamine versus protein-bound glutamine). AB - The goal of the present work was to compare the plasma glutamine response to exogenous glutamine administration in human volunteers; glutamine was provided as a free amino acid, bound to proteins, or in the form of peptides. Plasma glutamine concentrations were measured in eight human volunteers at 30, 60, 90, 120, and 240 min after receiving a drink containing 30 g of protein from one of the five different proteins tested (sodium caseinate, sodium caseinate + free glutamine, carob germ flour, carob protein concentrate, and carob protein hydrolysate). Peak plasma glutamine concentrations were 42% higher than postabsorptive basal values when exogenous glutamine was administered in the form of free glutamine added to caseinate (925.9 +/- 67.7 versus 651.3 +/- 44.0 micromol/L, respectively). In contrast, when glutamine was offered 100% bound to proteins (carob proteins), peak plasma glutamine concentration increased only between 18% and 23% from basal values, possibly because of the lower digestibility of carob proteins versus that of caseinate + free glutamine, to a different glutamine utilization at the gut level, or to a different response in endogenous glutamine kinetics to enteral administration of glutamine, depending on the molecular form of the glutamine source (free or protein bound). PMID- 11118822 TI - Effects of exercise training and amino-acid supplementation on body composition and physical performance in untrained women. AB - The purpose of this study was to determine the effects of 6 wk of essential amino acid (EAA) supplementation on body composition and exercise performance in untrained women (n = 21). Subjects were randomly assigned to a placebo (cellulose) or an EAA (average daily dose of 18.3 g of EAAs in pill form) group. Each subject participated in aerobic and heavy-resistance training three times per week. Body composition was assessed via dual x-ray absorptiometry analysis. Muscular endurance was determined via treadmill time to exhaustion, and strength was assessed by the total amount of weight lifted for one set to exhaustion at an estimated 12 repetitions maximum. No changes occurred in either group for body weight, lean body mass, fat mass, or bone mineral content. Treadmill time to exhaustion (TTE) improved significantly (P < 0.05) in the EAA group (mean +/- SD; pre-TTE = 13.15 +/- 3.67 min, post-TTE = 14. 73 +/- 4.26 min), whereas the placebo group did not change significantly. The total weight lifted at the subject's maximum 12 repetitions did not significantly change in either group. In previously untrained individuals, the ingestion of EAAs combined with aerobic and heavy-resistance training for 6 wk did not have a significant effect on body composition or muscular strength; however, aerobic muscular endurance increased significantly. PMID- 11118823 TI - Loss of zinc and selenium does not occur through peritoneal dialysis. AB - A prospective study was done to determine whether zinc (Zn) and selenium (Se) loss occurs in patients on continuous ambulatory peritoneal dialysis through their dialysate effluent. Fifty pairs of aliquots of dialysis fluid were collected from 29 patients. Each paired set of specimens consisted of the dialysate fluid before instillation into the peritoneal cavity and a specimen of the spent effluent after dialysis for comparison. Zn and Se concentrations were measured using inductively coupled plasma atomic emission spectrophotometry. The range of dialysate fluid Zn concentration before instillation was 0 to 1.75 microg/mL and that for Se was 0 to 0.33 microg/mL. Ranges for Zn and Se in the postdialysis effluent were 0 to 0.60 and 0 to 0.56 microg/mL, respectively. The concentration differences in 50 pairs of samples were analyzed with Wilcoxon's test. The difference in Zn levels between the predialysis specimen and the effluent was 0.009 +/- 0.036 microg/mL (P = 0.154). The difference for Se was 0.018 +/- 0.21 microg/mL (P = 0.118). In conclusion, no significant loss or gain of Zn or Se occurs in patients undergoing continuous ambulatory peritoneal dialysis. PMID- 11118824 TI - Osteoporosis with vertebral fractures associated with pregnancy and lactation. AB - Three cases of young women who developed severe vertebral osteoporosis after pregnancy and during lactation are described. These patients shared several features: a low-calcium diet during most of their lives, very-low body weight in two patients, and a positive family history of osteoporosis in two patients. Initial studies disclosed vertebral fractures, severely diminished bone mineral density of the spine (Z score = -3.3 to -4.1), and a less severely affected bone mineral density of the hip (Z score = -1.6 to -2.3). During the prolonged follow up of these patients, treated with oral biphosphonates, vitamin D, and calcium, an improved clinical response with a marked recovery of spine bone mineral density was observed. Poor general nutrition, low calcium intake, and a positive family history of osteoporosis appear to be strong risk factors for pregnancy- and lactation-associated osteoporosis. Although the mechanism of action is uncertain, calcium, vitamin D, and antiresorptive agents may have been beneficial in the treatment of this severe disorder. PMID- 11118825 TI - Long-term effects of severe undernutrition during the first year of life on brain development and learning in Chilean high-school graduates. AB - The objective of this study was to assess the relative impact of undernutrition during the first year of life on brain development, intellectual quotient (IQ), and scholastic achievement (SA) of poor Chilean high-school graduates (mean age = 18.3 +/- 0.9 y). A comparative study of two groups of high-school graduates from a low socioeconomic stratum was carried out. The undernourished group (n = 16), who had suffered from severe undernutrition during the first year of life, was compared with the non-undernourished group (n = 16). The final sample consisted of 32 right-handed high-school graduate students born at term who had no history of alcoholism or symptoms of brain damage, epilepsy, or heart disease and whose mothers had no history of smoking, alcoholism, or drug intake before and during pregnancy. Socioeconomic status was measured by using Graffar's modified method. Birth weight was used as the prenatal nutritional status index, and postnatal nutritional status was assessed by the body mass index, Z score for head circumference, and brachial anthropometry. IQ was determined with the Wechsler Intelligence Scale for Adults, and SA was determined with test in language and mathematics with the academic aptitude test. Brain development was evaluated by magnetic resonance imaging. Statistical analysis included variance tests, Scheffe's test for comparison of means, correlation, and multiple regression. Maternal schooling, brain volume, and undernutrition were the independent variables, with the greatest explanatory power in IQ variance (r(2) = 0.714). Only IQ explained SA variance (r(2) = 0.860); IQ, corpus callosum length, anteroposterior diameter, and maternal schooling were the independent variables, with the greatest explanatory power in the academic aptitude test variance (r(2) = 0.949). Results show that the long-term effects of malnutrition at an early age may affect brain development, IQ, and SA in school-age children. These findings are useful for nutrition and educational planning. PMID- 11118826 TI - Body composition in HIV-infected women. AB - Although loss of lean body mass is a common complication of human immunodeficiency virus (HIV) infection that can occur across the disease trajectory, few studies have characterized the body composition of HIV-infected women. We used bioelectrical impedance analysis to characterize the body composition of HIV-infected (n = 56) and uninfected (n = 12) women who were matched on percentage of ideal body weight. The HIV-infected women did not differ from the uninfected women by height-adjusted fat mass or body cell mass. Intergroup comparisons among the HIV-infected women showed that underweight women had significantly less fat mass than did normal-weight women but did not significantly differ with respect to body cell mass. Among all HIV-infected women, CD4(+) lymphocyte count was positively correlated with fat mass (r = 0.32, P = 0.01) but not with body cell mass. No significant correlations were found between any body-composition parameter and plasma viral load. Our findings suggest that, unlike men, HIV-infected underweight women show a preferential loss of fat mass and a relative preservation of body cell mass. This altered pattern of weight loss may relate to higher premorbid fat stores in women and/or hormonal differences. PMID- 11118827 TI - Trace-element status in milk and plasma of Kuwaiti and non-Kuwaiti lactating mothers. AB - There is a wide variation in the reported data on the concentrations of trace elements in human milk from different countries, but such data are not available for Kuwait. The objective of this study was to analyze the concentration of zinc, copper, manganese, and iron in milk and plasma of Kuwaiti and non-Kuwaiti mothers during prolonged lactation. Milk samples (from 34 donors) were collected early in the morning before feeding the infant. Trace elements were analyzed using atomic absorption spectrophotometry. Protein content and activity of superoxide dismutase were assayed spectrophotometrically. Concentration of zinc, copper, iron, and total protein and activity of superoxide dismutase in milk and of only zinc in plasma of Kuwaiti mothers were significantly higher than those of non Kuwaitis. Concentration of zinc, copper, manganese, and total protein in milk of both groups decreased as lactation continued but that of milk iron and plasma trace elements remained unchanged. The data of Kuwaiti mothers are consistent with those of previous reports on hyperuricemia, and the prevalence of obesity was found to be higher in the Kuwaiti population than in other countries. High protein content in association with high concentration of trace elements in milk of Kuwaiti versus non-Kuwaiti mothers may indicate that protein content in milk is an important determining factor for the concentration and bioavailability of these elements. PMID- 11118828 TI - Low dosages of exogenous growth hormone and its effect on growth in an animal model of suboptimal nutrition. AB - In a previous study, weight gain, insulin growth factor-1 (IGF-1) and insulin growth factor binding protein-3 (IGFBP-3) were increased in rats fed suboptimal levels of energy and administered 0.1 mg/100 g of body weight of recombinant human growth hormone (rhGH). Our objective was to determine whether these anabolic effects were still obtained with lower dosages of rhGH in similarly fed rats. Three groups of male, prepubertal Sprague-Dawley rats were administered rhGH and three groups of similar rats were given normal saline solution daily (0.05 mg/100 g of body weight subcutaneously). All rats were fed a balanced 1:1 carbohydrate:fat ratio diet for 4 wk. Restricted rats within each treatment were pair fed 80% and 60% ad libitum. Daily body weight, food intake, and efficiency were recorded. After 4 wk, serum IGF-1 and IGFBP-3, body fat, fat-free mass, and total body water were determined. Total weight gain and serum IGFBP-3 were significantly higher, with a tendency for increased body fat, in rats treated with rhGH and fed at 60% ad libitum. However, within each treatment, energy restriction caused decreased body fat and total body water. These results suggest that lower dosages of rhGH provide anabolic effects during suboptimal energy intake. PMID- 11118829 TI - Effect of select antioxidants on malondialdehyde modification of proteins. AB - To determine whether commonly used antioxidants alter malondialdehyde (MDA) modification of proteins, a known mechanism of free radical-related tissue injury, we studied the effect of adding 1 mg/mL of pycnogenol, 5 mM of alpha tocopherol, 5 mM of ascorbate, and 0.2 mg/mL of an ethanol equivalent of red and white wine on MDA-protein content of endothelial cells in culture. The addition of pycnogenol but not of the other antioxidants was associated with significant reduction in MDA-protein content compared with controls (0.521 +/- 0.041 in arbritrary units versus 1.011 +/- 0.021, P < 0. 001). To determine whether the observed effect occurs distal to MDA generation, the effect of these antioxidants on the modification of bovine serum albumin with MDA generated in a cell-free system was studied. In this cell-free assay, pycnogenol but not the other antioxidants reduced MDA-BSA generation by approximately 50%. It is concluded that pycnogenol may reduce MDA modification of proteins at a step distal to MDA generation. This may be an additional mechanism of protective effects of pycnogenol against oxidative stress. PMID- 11118830 TI - Potential markers of the nutritional status in an experimental model. AB - The activity of adenine deaminase (ADA) and purine nucleoside phosphorylase (PNP) as potential nutritional markers was analyzed in an experimental model. Weanling Wistar rats were fed a protein-free diet ad libitum to obtain a severe degree of wasting. An age-matched control group received a stock diet. At the end of the experiment, body weight (BW) and thymus weight (TW) were determined. Activity of ADA and PNP was determined on thymocytes of protein-deprived and control rats; the results, expressed as micromoles of uric acid x 10(-1)/W (W = TW/BW(0.75)), were 17.0 +/- 2.6 versus 9.1 +/- 3.0 for ADA and 11.5 +/- 4.2 versus 3.9 +/- 1.0 for PNP (P < 0.01). These results suggest that the nutritional stress provoked by the administration of a protein-free diet from weaning onward affects the development of thymocytes. Moreover, the increase in the activity of ADA and PNP would be an alternative mechanism to avoid the accumulation of high levels of deoxynucleotides, which would be toxic for T lymphocytes. However, some investigators have observed an increase of ADA activity in human serum under some adverse conditions; for this reason and taking into account the present findings, it would be interesting to determine the relation between the activity of ADA and PNP in thymocytes and serum in experimental models to analyze and propose these biochemical parameters as potential and useful markers of nutritional status; it also would be interesting to test this relation in human studies. PMID- 11118831 TI - Recent advances in the use of vitamin A (retinoids) in the prevention and treatment of cancer. AB - Vitamin A, its physiologic metabolites, and synthetic derivatives (retinoids) have been shown to have protective effects against the development of certain types of cancer. In addition, pharmacologic amounts of retinoids have been used with some success in the treatment of a few human tumors. The chemoprevention effect of retinoids is most likely exerted at the tumor-promotion phase of carcinogenesis. Retinoids block tumor promotion by inhibiting proliferation, inducing apoptosis, inducing differentiation, or a combination of these actions. Clinically, isotretinoin (13-cis-retinoic acid) significantly decreases the incidence of second primary tumors in patients with head-and-neck cancer and reduces appearance of non-melanoma skin cancer in patients with xeroderma pigmentosum. Retinoic acid has proved to be an effective treatment for promyelocytic leukemia. However, retinoid resistance limits its use as a single agent. Clinical trials are in progress to determine the efficacy of retinoids in treating other types of cancer such as neuroblastoma and breast carcinoma. The development of receptor-selective retinoids and selective inhibitors of retinoid metabolism may lead to further use of retinoids in both chemoprevention and treatment of cancer. PMID- 11118832 TI - Abnormal liver functions as a result of total parenteral nutrition in a patient with short-bowel syndrome. AB - The pathogenesis of total parenteral nutrition (TPN)-induced liver cholestasis is poorly understood. Cholestasis generally occurs late in TPN therapy in association with elevated serum alkaline phosphatase and total bilirubin concentrations. Such factors as preexisting medical conditions, excessive nutrient infusion, amino-acid deficiency, absence of enteral stimulation, protracted duration of therapy, continuous infusion schedule, and hypoalbuminemia have all been suggested as possible etiologies. Various treatments have been proposed for the correction of TPN-induced cholestasis including administration of bile salt and antimicrobial therapies. To avoid potential hepatic complications associated with TPN, certain preventive measures can be considered. Administration of energy substrates should not be excessive. A mixed-fuel system that includes lipids should be implemented. TPN should be cycled if it will be used long term, and initiation of enteral nutrition should begin as soon as possible. PMID- 11118833 TI - Micronutrients and infections: an African perspective. PMID- 11118834 TI - Dietary fatty acids and cardiovascular health. PMID- 11118835 TI - A cocktail approach to antioxidant therapy. PMID- 11118836 TI - Retinoids and cancer prevention: crossing the line between food and drug. PMID- 11118837 TI - Does the fate of enterally administered glutamine depend on its molecular form? Bound versus free amino acid. PMID- 11118838 TI - Measurement and clinical significance of body-composition changes in HIV disease. PMID- 11118839 TI - Role of S-adenosylmethionine in hyperhomocysteinemia and in the treatment of alcoholic liver disease. PMID- 11118840 TI - Caloric intake and longevity. PMID- 11118841 TI - The antiinfective vitamin arises once more. PMID- 11118843 TI - The odds ratio: impact of study design. PMID- 11118842 TI - The changing smoking and health scene in the Czech Republic. PMID- 11118844 TI - Eicosapentaenoic acid (EPA): an antiinflammatory omega-3 fat with potential clinical applications. PMID- 11118845 TI - Annual Symposium, American College of Nutrition, Washington, DC, October 1-4, 1999. PMID- 11118846 TI - Methylene blue derivatives--suitable photoantimicrobials for blood product disinfection? AB - Photodynamic antimicrobial agents based on the well-established phenothiazinium biological stain methylene blue offer a simple method for the inactivation or destruction of pathogens contained in donated blood and blood products. The technique is currently concentrated on viruses and the disinfective procedure can be carried out in blood bags using basic low-power light sources. Pathogens of the bacterial, yeast and protozoal classes are also susceptible to phenothiaziniums. The photoantimicrobial mode of action is usually via oxidative damage to cellular components, either due to redox reactions between the agent and a biomolecular target or by the action of reactive oxygen species generated in situ by photodynamic action. The targeting of various microbial species is discussed in relation to the physicochemical make-up of the photosensitizers, and future directions are suggested. PMID- 11118847 TI - beta-Lactam susceptibility patterns and investigation of cephalosporin hydrolysing beta-lactamases of Gram-negative extraintestinal clinical isolates. AB - Of more than 3500 isolates of enterobacteriaceae, 48-69% were resistant to aminopenicillins and 11-45% to amoxycillin+clavulanic acid. Resistance to second and third generation cephalosporins was present in 11-17 and 3-8% of Escherichia coli, 47-56 and 15-52% of Klebsiella-Enterobacter, 36-57 and 16-27% of Proteus, Providencia and Morganella isolates. Pseudomonas aeruginosa strains varied in their resistance to antipseudomonal beta-lactams. Isoelectric points, inhibitor profiles and substrate profiles of beta-lactamases extracted from representatives of the resistant strains indicated that the resistance was mainly due to the hyperproduction of chromosomally encoded AmpC beta-lactamases. This was confirmed by plasmid profile and PCR investigations. Extended-spectrum beta-lactamase and metallo-penicillinase producing strains were not found. One Pseudomonas maltophilia strain produced an oxacillinase. PMID- 11118848 TI - Susceptibility of bacterial isolates to gatifloxacin and ciprofloxacin from clinical trials 1997-1998. AB - MICs of gatifloxacin and ciprofloxacin against 3482 pre-treatment, clinical trial isolates collected during 1997-1998 are reported. These data suggested that gatifloxacin was four- to eight-fold more active than ciprofloxacin against Gram positive bacteria, with gatifloxacin MIC(90)s < or = 0.33 mg/l against Staphylococcus aureus and Streptococcus pneumoniae, and < or = 1.0 mg/l versus viridans streptococci and Enterococcus faecalis. Both quinolones had similar MIC(90)s versus Enterobacteriaceae (generally < or = 0.38 mg/l, except 0. 7-0.8 mg/l for Citrobacter freundii) and Pseudomonas aeruginosa ( approximately 8 mg/l). A total of 78% P. aeruginosa had gatifloxacin MICs < or = 2 mg/l. Gatifloxacin was more active than ciprofloxacin against Acinetobacter species and non-P. aeruginosa pseudomonads. Both had exceptional activity versus Haemophilus spp, Moraxella catarrhalis and Neisseria gonorrhoeae. In summary, compared to ciprofloxacin, gatifloxacin had improved activity against Gram-positive bacteria and comparable activity against Gram-negative bacteria. PMID- 11118849 TI - Gemifloxacin and ciprofloxacin pharmacodynamics in an in-vitro dynamic model: prediction of the equivalent AUC/MIC breakpoints and doses. AB - To compare the antimicrobial effects (AMEs) of gemifloxacin (GEM) and ciprofloxacin (CIP) on Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa, a series of pharmacokinetic profiles of GEM (a single dose with the half-life (T(1/2)) of 7.4 h and CIP (two 12 h doses with T(1/2) of 4 h) were simulated in vitro over eight-fold ranges of the AUC/MIC ratio. Species- and strain-independent linear relationships observed between the intensity of AME (I(E)) and log AUC/MIC were not superimposed for GEM and CIP (r(2)=0.99 and 0.98, respectively). The predicted ratio for GEM that might be equivalent to a clinically established breakpoint value of AUC/MIC=125 (mg h/l)/(mg/l) for CIP was estimated at 110 (mg h/l)/(mg/l). It was calculated, that a daily dose of CIP that might provide the same AME as a clinical dose of GEM (320 mg) on a hypothetical strain of S. aureus with MICs=MIC(50)s would be as high as 2 x 3200 mg. PMID- 11118850 TI - Studies on the genotoxicity of turmeric extracts in bacterial system. PMID- 11118851 TI - New concepts for future control of HIV derived from studies of pathogenesis. PMID- 11118852 TI - Clinical impact of anti retroviral resistance testing: current problems and future directions. PMID- 11118854 TI - Genotyping in HIV drug resistance mutations: epidemiology in 145 patients. AB - The epidemiology of HIV-1 genomic mutations causing antiretroviral drug resistance, in 145 HIV-1 infected patients were obtained using a new sequencing technique. All patients were in a follow up treatment for at least 4 months with all drugs available in clinical practice in accordance to international guidelines. The percentage of the mutations were calculated both on the number of all participants and on the number of patients who received the drugs selecting for that specific resistance. It was possible then to evaluate patients who showed the mutation without receiving the drug. We consider this new sequencing method reliable and hopeful in clinical practice, giving the opportunity for a guided therapy for the single patient and in detecting and monitoring the antiretroviral drug resistance mutations in the pertinent geographic area following a periodic surveillance program. PMID- 11118853 TI - Resistance to antiretroviral drugs in patients with primary HIV-1 infection. Investigators of the Quebec Primary Infection Study. AB - The widespread use of antiretroviral agents (ARVs) and the growing occurrence of HIV strains resistant to these drugs have given rise to serious concerns regarding the transmission of resistant viruses to newly infected persons. Plasma viral RNA from 80 individuals newly infected between 1997 and 1999 was genotyped by automated sequencing to analyze the profile of viruses resistant to nucleoside and non-nucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs) and to protease inhibitors (PIs). The prevalence of mutations that conferred primary resistance to PIs (L10I, D30Y, V82A, L90M) was 15% of the cohort. RT genotypic variants, associated with high-level resistance to ARVs, were observed in 21% of individuals, including NRTI, NNRTI and multidrug (MDR) resistance in 6, 5, and 10% of cases, respectively. The phenotypic susceptibility of viral isolates to ARVs was also assayed and showed transmission of high-level resistance to ZDV, 3TC, and PIs in those individuals with MDR. The transmission of drug-resistant HIV genotypic variants is a serious problem that merits further attention by public health officials, virologists, and clinicians. PMID- 11118856 TI - Laboratory diagnosis of human cytomegalovirus (HCMV) central nervous system disease in AIDS patients. PMID- 11118855 TI - Use of HIV-1 p24 as a sensitive, precise and inexpensive marker for infection, disease progression and treatment failure. AB - HIV RNA is an acknowledged marker of disease activity and predictive of progression, while the p24 antigen is considered unsuitable. This is at odds with the fact that viral pathogenesis is usually mediated by proteins. One might expect that p24, if analyzed properly, might even be superior to RNA. This hypothesis was investigated in clinical studies using a sensitive and precise p24 test (heat-mediated immune complex dissociation with signal amplification-boosted ELISA). This test was as sensitive and specific as the polymerase chain reaction (PCR) for viral RNA (200-400 copy detection), an overall better predictor of CD4 decline and survival, while RNA prevailed in predicting AIDS. The lower costs of p24 testing also permit a closer monitoring of patients with an earlier detection of anti-retroviral treatment failures. PMID- 11118857 TI - Use of CMV transcripts for monitoring of CMV infections in transplant recipients. AB - The development of the nucleic acid sequence-based amplification (NASBA) technology has allowed qualitative determination of human cytomegalovirus (HCMV) immediate-early (IE) and late (pp67) transcripts for monitoring of HCMV infections in the post transplantation period. pp67-mRNA NASBA was shown to be less sensitive than pp65 antigenemia and leukoDNAemia, yet more sensitive than viremia in (i) detecting HCMV infection in both patients and blood samples and (ii) anticipating diagnosis of HCMV infection in solid organ (heart, lung) transplant recipients (SOTR). Use of pp67-mRNA NASBA, as a parameter for initiation of pre-emptive therapy, could be employed as an alternative to detecting antigenemia or DNAemia in SOTR, whereas in bone marrow transplant recipients (BMTR) its use would be too risky because of the delayed detection of HCMV infection. On the other hand, IE-mRNA NASBA was shown to be largely superior to the other assays both in detecting HCMV infection in patients and blood samples and in anticipating diagnosis of HCMV infection. This appears particularly useful in BMTR, where early initiation of antiviral treatment is mandatory in order to prevent the appearance of HCMV interstitial pneumonia. Pre emptive therapy in BMTR, however, if based upon IE-mRNA NASBA would imply treatment of a greater number of patients as compared with antigenemia- or DNAemia-guided treatment. The clinical usefulness of this approach should be evaluated in prospective trials in the near future, pp67-mRNA NASBA in SOTR with reactivated HCMV infections and IE-mRNA NASBA in BMTR could represent two new virologic parameters to be used as a cutoff for pre-emptive therapy control of HCMV infections in the post-transplant period. Viral transcripts are more direct markers of viral replication in vivo and their disappearance indicates block of the replication process. PMID- 11118858 TI - Cytomegalovirus pp65 antigenemia in HIV patients: retrospective on 3 years study. PMID- 11118859 TI - Epidemiology of HIV-1 and emerging problems. AB - Broad use of antiretroviral drugs is becoming a factor that is important to consider for understanding the HIV-1 epidemiology. Since 1993, we observe that a proportion of new infections within major risk groups in Amsterdam is caused by azidothymidine (AZT)-resistant viruses. After the introduction of combination therapy in The Netherlands in 1997, new infections with drug-resistant viruses have not been documented. Large-scale monitoring of anti-HIV-1 therapy failures revealed that antiretroviral drugs may yield previously undescribed resistant viruses, which contain a two amino acid insertion (68SS/V69) within their reverse transcriptase genes in combination with mutations at codons 67 and 215. These viruses are highly resistant to AZT, 3TC, and d4T, and moderately resistant to ddI and ddC. PMID- 11118860 TI - Bacterial agents of lower respiratory tract infections (LRTIs), beta-lactamase production, and resistance to antibiotics in elderly people. DEDALO Study Group. AB - This study determined the etiology of lower respiratory tract infections in the elderly and assessed whether the growth of beta-lactamase producing bacteria is particularly favoured in these patients. Between December 1998 and May 1999, 187 patients with community-acquired pneumonia (CAP), and 887 patients with acute exacerbations of chronic obstructive pulmonary disease (AECOPD) were enrolled. The mean age was 74 years (range of 65-94 year). Sputum and bronchial aspirate for microbiological investigation were obtained. Besides organisms commonly involved in bacterial infections of the lower respiratory tract (i.e. Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis), Enterobacteriaceae and Pseudomonas spp. were also found. A high percentage of these bacteria were beta-lactamase producers. These data along with the clinical presentation, severity of infection, and epidemiological knowledge, might represent a guide for the choice of empiric antimicrobial treatment. PMID- 11118861 TI - Antibiotic resistance in Gram-positive cocci. AB - Gram-positive cocci still predominate as a cause of nosocomial- and community acquired infections. These organisms frequently reveal a high natural, intrinsic resistance to antimicrobials. Additionally, these bacteria are able to acquire resistance to frequently used drugs rapidly through selective pressure of the environment and via the genetic evolution of bacteria. The wide application of antimicrobials in medical and veterinary practice, usage of antibiotics in agriculture and common usage of antiseptics and disinfectants result in selective pressure. The use of antibiotics directly selects resistant variants to different antibiotics or disinfectants. The same genetic element (e.g. qac or smr) conferring resistance to some disinfectants are often present on the same plasmid conferring resistance to antibiotics. Selection of resistant variants occurs most frequently in the hospital environment. Staphylococcus aureus and enterococci are the most commonly isolated bacteria causing nosocomial infections. Among those giving therapeutic problems are methicillin-resistant staphylococci and vancomycin-resistant enterococci. Resistance to high levels of aminoglycosides or penicillins among hospital enterococcal strains can completely abolish synergism of the drugs. In these cases glycopeptides will be the drugs of choice in the treatment of serious infections. Recently S. aureus strains with decreased susceptibility to vancomycin has appeared. A mechanism for this elevated resistance, although intensively investigated, still remains unknown. PMID- 11118862 TI - Epidemiology of Gram-negative antibiotic resistance in outpatients: a year of surveillance. PMID- 11118863 TI - Resistance of bacteria in urinary tract infections. AB - Bacterial infection of the urinary tract is a common health problem in young women but also the most common nosocomial infection (>33%) contributing to the mortality of patients, and increasing the duration and cost of hospitalization. Escherichia coli is the most predominant organism and its prevalence varies in different studies. The high consumption of inappropriately prescribed antibiotics, combined with multiple pathology and frequent use of invasive devices, is a major factor contributing to high levels of resistance. There is a serious decrease in susceptibility of E. coli strains to amoxycillin, due to the presence of R-TEM enzymes, to cotrimoxazole and trimethoprim. Nitrofurantoin and fosfomycin-trometamol remain highly active against urinary Enterobacteriaceae, with over 90% of E. coli being susceptible. Knowledge of the most likely causative organisms and the prevalence of resistance pathogens to antimicrobial agents is essential to select antibiotics and to establish guidelines for the empirical treatment of urinary tract infections. PMID- 11118864 TI - The impact of resistance on the management of urinary tract infections. AB - Urinary tract infections requiring treatment are extremely common. It is estimated that between 20 and 50% of adult women will have had at least one symptomatic urinary tract infection. When considering the optimal therapy of any infection, patient factors, organism factors, drug factors (e.g. pharmacokinetics), side-effects and cost as well as antimicrobial resistance all need to be considered. This paper deals with the impact of increasing antibiotic resistance on the management of urinary tract infections. PMID- 11118865 TI - Changes in the resistance patterns among upper respiratory tract infection pathogens. PMID- 11118866 TI - Short-course treatment for acute tonsillopharyngitis. PMID- 11118867 TI - New technologies and applications of PCR in clinical diagnosis. PMID- 11118868 TI - Pharmacokinetics and pharmacodynamics of oral cephalosporins as critical factors in choice of antibiotics. PMID- 11118869 TI - Strategies to treat patients with antibiotic resistant Helicobacter pylori. AB - Increased resistance to clarithomycin and metronidazole, the two main antibiotics used to treat Helicobacter pylori infection, has led to a search for alternatives to the proton pump inhibitor based triple therapies commonly used. The main rescuse therapy is a bismuth-based quadruple therapy. However, triple therapies with tetracycline and metronidazole or amoxicillin and metronidazole can be considered in the case of clarithomycin resistance. They can also be used in the case of metronidazole resistance by increasing the dose and duration of metronidazole. The only therapy without clarithomycin and metronidazole includes rifabutin and amoxicillin. Dual therapies with amoxicillin and a proton pump inhibitor at high dose can also be used. PMID- 11118870 TI - The intensive care physician and control of antimicrobial resistance. PMID- 11118871 TI - Iron deficiency anaemia and Helicobacter pylori infection. AB - Iron deficiency anaemia (IDA) is the most common form of anaemia world-wide. IDA is the simple result of an imbalance between iron loss and absorption. Gastric function with hydrochloric and ascorbic acid is essential for iron absorption. Some strains of Helicobacter pylori are able to acquire iron, competing with the host. A large percentage of patients with atrophic body gastritis (ABG) develop IDA and 61% of them are H. pylori positive. Recent evidence suggests that H. pylori infection could cause IDA in the absence of peptic ulcer or other upper gastrointestinal (GI) tract bleeding lesions. Gastritis extending to the corpus and a high bacterial load are features of these patients. About 70% of IDA patients with ABG or H. pylori gastritis are premenopausal women. Both ABG and H. pylori gastritis should be considered when evaluating the GI tract of patients with iron deficiency anaemia. PMID- 11118872 TI - Treatment and prevention of antibiotic associated diarrhea. AB - Mild or severe episodes of antibiotic-associated diarrhea (AAD) are common side effects of antibiotic therapy. The incidence of AAD differs with the antibiotic and varies from 5 to 25%. The major form of intestinal disorders is the pseudomembranous colitis associated with Clostridium difficile which occurs in 10 20% of all AAD. In most cases of AAD discontinuation or replacement of the inciting antibiotic by another drug with lower AAD risk can be effective. For more severe cases involving C. difficile, the treatment of diarrhea requires an antibiotic treatment, with glycopeptides (vancomycin) or metronidazole. Another approach to AAD treatment or prevention is based on the use of non-pathogenic living organisms, capable of re-establishing the equilibrium of the intestinal ecosystem. Several organisms have been used in treatment or prophylaxis of AAD such as selected strains of Lactobacillus acidophilus, L. bulgaricus, Bifidobacterium longum, and Enterococcus faecium. Another biotherapeutic agent, a non-pathogenic yeast, Saccharomyces boulardii has been used. In animal models of C. difficile colitis initiated by clindamycin, animals treated with S. boulardii (at end of vancomycin therapy) had a significant decrease in C. difficile colony forming units, and of toxin B production. In several clinical randomised trials (versus placebo), S. boulardii has demonstrated its effectiveness by decreasing significantly the occurrence of C. difficile colitis and preventing the pathogenic effects of toxins A and B of C. difficile. It has been shown to be a safe and effective therapy in relapses of C. difficile colitis. A good response has been seen in children with AAD, treated by S. boulardii only. In ICUs prevention of AAD remains based on limitation of antibiotic overuse and spread of C. difficile or other agents of AAD should be prevented by improved hygiene measures (single rooms, private bathrooms for patients, use of gloves and hand washing for personnel). In addition the increasing use of biotherapeutic agents such as S. boulardii should permit the prevention of the major side effect of antibiotics, i.e. AAD in at risk patients. PMID- 11118873 TI - Clinical and microbiologic efficacy and safety profile of linezolid, a new oxazolidinone antibiotic. AB - Gram-positive cocci are important causes of infection both in the community and in the hospital, with repercussions on mortality and increased economic costs. Treatment of these infections is made difficult by the increasing emergence of multi-resistant organisms, primarily among Gram-positive cocci, such as vancomycin-resistant enterococci, methicillin-resistant Staphylococcus aureus and coagulase-negative staphylococci, and penicillin-resistant pneumococci. Linezolid, a member of the new class of synthetic antimicrobials named oxazolidinones, has several favourable characteristics including high activity against multiresistant Gram-positive cocci. In a number of clinical trials, linezolid showed good clinical and microbiologic efficacy in the therapy of infections caused by these organisms. It can be considered a valid option for treating both community- and hospital-acquired infections due to multiresistant Gram-positive cocci. PMID- 11118874 TI - The use of probiotics in medical practice. AB - Probiotics are defined as living organisms, beneficial to health when ingested. Different species of microorganisms such as lactic acid bacteria or yeasts have been proposed for human use. These microorganisms differ from each other and it is, therefore, unlikely that they will act in the same way. Probiotics could be used for several conditions such as diarrhoea, candidal vaginitis, urinary tract infections, immune disorders, lactose intolerance, hypercholesterolaemia and food allergy. The effects of probiotics in some of these conditions have been directly observed, in others it has been only suggested on the basis of in vitro studies and from experimental animal models. Controlled trials are needed to determine the scientific basis for their use, the correct formulation and ways of administration in different clinical situations. PMID- 11118875 TI - Is there in vivo-in vitro correlation between antifungal susceptibility, species of Candida spp. and clinical outcome? PMID- 11118876 TI - Antifungal therapy in thoracic cancer patients with low dose liposomal amphotericin B (AmBisome). PMID- 11118877 TI - Candida albicans cellular internalization: a new pathogenic factor? AB - The preliminary results of a study to show the possibility that Candida albicans can internalize into epithelial cells are reported. The study was performed on buccal, vaginal and HeLa cells. Buccal and vaginal cells, at a concentration of 5 x 10(4) cells/ml and HeLa monolayers were incubated for 2, 3 and 4 h with 10(5) colony forming units of a Candida albicans isolate. After incubation, non internalised yeasts were eliminated and samples were processed for examination by Scanning Electron Microscopy. Results suggest that receptor-mediated endocytosis could be involved in the pathogenesis of recurrent oral and vaginal infections. This phenomenon could represent an interesting experimental model to testing drug interference in the development of therapeutic strategies against C. albicans infections. PMID- 11118878 TI - Pneumocystis carinii diagnosis: an update. AB - From 1994 to date we have been using the internal transcribed spacers (ITSs) nested polymerase chain reaction (PCR) to investigate the possibility of diagnosing Pneumocystis carinii pneumonia on non-invasive samples collected from HIV-positive patients with pulmonary involvement. The objectives were: (1) to test the sensitivity, specificity and prognostic value of PCR in diagnosis and follow up of PCP; (2) to investigate the eventual occurrence and role of asymptomatic carriers of P. carinii; (3) to evaluate the prognostic significance of blood PCR positivity versus respiratory samples; (4) to verify the occurrence of exogenous infections or endogenous reactivations in cases of recurrent P. carinii pneumonia; and (5) to study the possible correlation between P. carinii genotype identified and capability of blood dissemination, prior prophylactic treatments, clinical parameters and outcome of the patients. PMID- 11118879 TI - Cell-autonomous and somatic signals control sex-specific gene expression in XY germ cells of Drosophila. AB - When XX germ cells develop in a testis they become spermatogenic. Thus, somatic signals determine the sex of genetically female germ cells. In contrast, XY germ cells experimentally transferred to an ovary do not differentiate oogenic cells. Because such cells show some male characteristics when analyzed in adults, it was assumed that XY germ cells autonomously become spermatogenic. Recently, however, evidence showing that a female soma feminizes XY germ cells was reported. The conclusion was drawn that the sex determination of XY germ cells is dictated by the sex of the soma. We monitored the fate of XY germ cells placed in a female environment throughout development. Here we report that such germ cells respond to both cell-autonomous and somatic sex-determining signals, depending on the developmental stage. Analyzing the expression of sex-specific molecular markers, we first detected autonomous male-specific gene expression in XY germ cells embedded in female embryos and larvae. At later stages, however, we found that sex-specific regulation of gene expression within XY germ cells is influenced by somatic gonadal cells. After metamorphosis, XY germ cells developing in a female soma start expressing female-specific and male-specific markers. Transcription of female-specific genes is maintained, while that of male-specific genes is later repressed. We show that in such XY germ cells, the female-specific gene Sex lethal (Sxl) is activated. Within the germline, Sxl expression is required for the activation of a further female-specific gene and the repression of male specific genes. We thus report for the first time the existence of downstream targets of the gene Sxl in the germline. PMID- 11118880 TI - Control of endoreduplication domains in the Drosophila gut by the knirps and knirps-related genes. AB - Endoreduplication cycles that lead to an increase of DNA ploidy and cell size occur in distinct spatial and temporal patterns during Drosophila development. Only little is known about the regulation of these modified cell cycles. We have investigated fore- and hindgut development and we present evidence that the Drosophila knirps and knirps-related genes are key components to spatially restrict endoreduplication domains. Our lack and gain-of-function experiments show that knirps and knirps-related which encode nuclear orphan receptors transcriptionally repress S-phase genes of the cell cycle required for DNA replication and that this down-regulation is crucial for gut morphogenesis. Furthermore, we demonstrate that both genes are activated in overlapping expression domains in the fore- and hindgut in response to Wingless and Hedgehog activities emanating from epithelial signaling centers that control the regionalization of the gut tube. Our results provide a novel link between morphogen-dependent positional information and the spatio-temporal regulation of cell cycle activity in the gut. PMID- 11118881 TI - Functional analysis of regulatory elements controlling the expression of the ecdysone-regulated Drosophila ng-1 gene. AB - The steroid hormone ecdysone controls multiple aspects of insect development, including larval moults and metamorphosis, and can induce specific genetic responses in different tissues. The definition of the molecular mechanisms able to mediate this tissue-specific responsiveness may greatly contribute to understanding how such an accurate genetic response is achieved. In this work we have identified, by transgenic analysis, the regulatory elements directing the expression of ng-1, an ecdysone-regulated Drosophila gene showing a highly specific developmental expression profile. Our results show that an ecdysone responsive element located within the ng-1 coding region is necessary for high level gene expression, whereas the gene's spatial and temporal expression profile is fully controlled by a distinct upstream regulatory region. This region binds a set of transcriptional factors, including the FKH regulatory protein, which can potentially modulate the ecdysone genetic regulated response. PMID- 11118882 TI - Pax4 regulatory elements mediate beta cell specific expression in the pancreas. AB - Pax4 is a member of the Pax (Pax, paired box) family of transcription factors with restricted expression and essential functions in the developing pancreas. Pax4 deficient mice do not develop pancreatic beta and delta cells and thus die after birth due to diabetes. In this study using transgenic mouse technology we report the identification and characterization of a 0.9 kb DNA fragment in the 5' region of the gene, which by itself is able to direct Pax4 expression in the endocrine pancreas, recapitulating the beta-cell-specific in vivo expression pattern of Pax4. The 0.9 kb DNA fragment contains an area spanning 407 base pairs that is highly conserved between human and mouse showing 88% identity. This promoter region contains sequence motifs that have been shown to be involved in beta-cell-specific expression of insulin, Pdx1 and islet amyloid polypeptide (IAPP). In addition, we determined a previously undescribed 5'intron. PMID- 11118883 TI - Some distal limb structures develop in mice lacking Sonic hedgehog signaling. AB - Patterning of the limb is coordinated by the complex interplay of three signaling regions: the apical ectodermal ridge (AER), the zone of polarizing activity (ZPA), and the non-ridge limb ectoderm. Complex feedback loops exist between Shh in the ZPA, Bmps and their antagonists in the adjacent mesenchyme, Wnt7a in the dorsal ectoderm and Fgfs in the AER. In contrast to the previously reported complete absence of digits in Shh(-/-) mice, we show that one morphologically distinct digit, with a well-delineated nail and phalanges, forms in Shh(-/-) hindlimbs, while intermediate structures are severely truncated and fused. The presence of distal autopod elements is consistent with weak expression of Hoxd13 in Shh(-/-) hindlimbs. Shh(-/-) forelimbs in contrast have one distal cartilage element, a less-well differentiated nail and fused intermediate bones. Interestingly, Ihh is expressed at the tip of Shh mutant limbs and could account for formation of distal structures. In contrast to previous studies we also demonstrate that Shh signaling is required for maintenance of normal Fgf8 expression, since expression of Fgf8, unlike some other AER marker genes, is rapidly lost from anterior to posterior after E10.5, with only a small domain of Fgf8 expression remaining posteriorly. Furthermore, loss of expanded Fgf8 expression is paralleled by a collapse of the handplate. Our data show that development of most intermediate elements of the hindlimb skeleton are Shh dependent, and that Shh signaling is required for anterior-posterior expansion of the AER in both limbs and for the subsequent branching of zeugopod and autopod elements. Finally, we show that Shh is also required for outgrowth of the limb ectoderm and thus for the formation of a distinct limb compartment. PMID- 11118884 TI - 3D confocal reconstruction of gene expression in mouse. AB - Three-dimensional computer reconstructions of gene expression data will become a valuable tool in biomedical research in the near future. However, at present the process of converting in situ expression data into 3D models is a highly specialized and time-consuming procedure. Here we present a method which allows rapid reconstruction of whole-mount in situ data from mouse embryos. Mid gestation embryos were stained with the alkaline phosphotase substrate Fast Red, which can be detected using confocal laser scanning microscopy (CLSM), and cut into 70 microm sections. Each section was then scanned and digitally reconstructed. Using this method it took two days to section, digitize and reconstruct the full expression pattern of Shh in an E9.5 embryo (a 3D model of this embryo can be seen at genex.hgu.mrc.ac.uk). Additionally we demonstrate that this technique allows gene expression to be studied at the single cell level in intact tissue. PMID- 11118885 TI - A reporter transgene based on a human keratin 6 gene promoter is specifically expressed in the periderm of mouse embryos. AB - We report the developmental regulation of a lacZ reporter transgene fused to the promoter region of the human keratin 6a gene. In mouse embryos, the transgene is expressed in the periderm (the outermost layer of embryonic epidermis), as are the endogenous keratin 6 alpha and beta genes. A subset of periderm cells, localized to temporary epithelial fusions, is known to contain keratin 6 protein, and we find that these cells also harbor LacZ enzymatic activity. PMID- 11118886 TI - Expression pattern of the Sox31 gene during zebrafish embryonic development. AB - We have identified a novel Sox gene in zebrafish (Danio rerio), Sox31, closely related to mammalian group B Sox genes. The gene is maternally expressed. Zygotic transcription starts at gastrulation, in the presumptive neuroectoderm. Later, expression is restricted to the developing central nervous system, including forebrain, midbrain, hindbrain and spinal cord. PMID- 11118887 TI - Variant hepatocyte nuclear factor 1 expression in the mouse genital tract. AB - Variant Hepatocyte Nuclear Factor 1 (vHNF1/HNF1beta) is a homeodomain-containing transcription factor first expressed in the primitive endoderm and its derivatives, the visceral and parietal endoderm. It is subsequently expressed in epithelial cells of different organs, including the primitive gut and derivatives (liver, pancreas, lung), the kidney, and transiently, in the neural tube. We report here new data concerning vHnf1 expression in the mouse genital tract, using both RNA analyses and our vHnf1 heterozygous mutant mouse line, in which the first coding exon of the vHnf1 gene is replaced by the NLSLacZ reporter gene. Both beta-galactosidase activity and vHnf1 transcripts are detected in epididymus, vas deferens, seminal vesicle, prostate, uterus and oviduct. RNA analysis and in situ hybridization studies demonstrate that vHnf1 transcripts are restricted to the germinal cells of the testis. Unexpectedly, no beta galactosidase activity is detected in the testis. We further show that, in addition to the somatic transcript, two more abundant vHnf1 transcript variants, which lack exons 1-4, appear in this organ after meiosis. PMID- 11118888 TI - The spatial and temporal expression pattern of Nesp and its antisense Nespas, in mid-gestation mouse embryos. AB - We describe the spatiotemporal expression pattern of Nesp, and its antisense transcript, Nespas. We found non-complementary expression of these two oppositely imprinted transcripts during mouse embryogenesis, in a number of forming embryonic structures. Nesp expression was primarily seen in the somites and vasculature, whereas Nespas was mainly detected in the progress zone, mesenchyme and ectoderm of the limb, and the neural tube. PMID- 11118889 TI - Cloning and expression analysis of the mouse T-box gene Tbx18. AB - T-box genes encode transcription factors that regulate a variety of developmental processes. In this report, we describe the cloning and expression analysis of the novel mouse T-box gene Tbx18. During development expression is most prominent in the proepicardial organ and in the epicardium of the heart. Other sites of expression include the cranial paraxial mesoderm, the presomitic mesoderm, the anterior somite half, the genital ridge, and the developing limb buds. PMID- 11118890 TI - Cloning and expression analysis of the mouse T-box gene tbx20. AB - T-box genes constitute a conserved multi-gene family with important roles in many developmental processes. In this report, we describe the cloning and expression analysis of a novel mouse T-box gene, Tbx20. Expression is prominent in the extraembryonic mesoderm, in the developing heart, the eye anlage and motor neurons of hindbrain and spinal cord. PMID- 11118891 TI - Overlapping expression of zebrafish T-brain-1 and eomesodermin during forebrain development. AB - T-box transcription factors are important determinants of embryonic cell fate and behaviour. Two T-box genes are expressed in the developing telencephalon of several vertebrate species, including amphibia, birds and mammals. Here we report the cloning of zebrafish T-brain-1 (tbr1) and eomesodermin (eom). As a prelude to genetic studies of neuro-ectodermal fate determination we studied their expression pattern during embryogenesis and early larval development. Eom is expressed in the presumptive telencephalon from around the 4-5 somite stage in bilaterally symmetric groups of cells; the number of positive cells increases dramatically with time and encompasses the entire dorsal telencephalon by the 22 somite stage. Tbr1 is expressed from the 18 somite stage in a subset of eom expressing cells. By 24 hpf eom and tbr1 are expressed in largely overlapping domains in the dorsal telencephalon, tbr1 is expressed in postmitotic cells whereas eomes is also expressed in proliferative ventricular zone cells. Both genes are also found in a small domain of the diencephalon bordering the telencephalon. A detailed analysis of the expression of tbr1 and eom in the brain of 4 day old larvae shows that the two T-box genes are differentially expressed in various cell populations of the developing brain. PMID- 11118892 TI - alpha4 integrin is expressed in a subset of cranial neural crest cells and in epicardial progenitor cells during early mouse development. AB - By RNA in situ hybridization and immunohistochemical analyses of early stage mouse embryos, we find that alpha 4 integrin gene is expressed in migratory cranial neural crest cells originating from the presumptive forebrain, midbrain, and rhombomeres 1 and 2 of the presumptive hindbrain. alpha 4 is also expressed in epicardial progenitor cells in the septum transversum that migrate to the heart. PMID- 11118893 TI - Cloning and expression of the chick p63 gene. AB - We have isolated a chick cDNA for p63, a member of the p53 transcription factor family. This cDNA encodes a protein of 582 amino acids for an alpha isoform in the C-terminal region, while lacking the N-terminal transactivation domain. The chick p63 gene is first expressed in the prospective cutaneous ectoderm at stage 6 and later in the developing epithelia. The p63 expression is intense in specialized epithelial structures, such as apical ectodermal ridge of the limb bud, epithelia of branchial arches and feather buds. Furthermore, we have found that the transcripts are detected in the interdigital epithelium, intersomite epithelium, and epaxial dermamyotome. PMID- 11118894 TI - Dynamic expression of Drosophila TRAF1 during embryogenesis and larval development. AB - TNF-receptor associated factors (TRAFs) comprise a family of adaptor proteins that act as downstream signal transducers of the TNF receptor superfamily and the Toll/interleukin-1 receptor family. The mammalian TRAFs 2, 5 and 6 are known to activate JNK- and NF-kappaB signaling pathways, whereas the function of the other three mammalian family members, TRAF 1, 3 and 4 is less well characterized. Vertebrate TRAFs have a very similar structure with the exception of TRAF1: aside the characteristic C-terminal TRAF domain, they share a N-terminal RING finger followed by five or, in the case of TRAF4, seven regularly spaced zinc fingers. Two TRAF homologues are present in the genome of Drosophila melanogaster, DTRAF1 and DTRAF2 (also known as DTRAF6) and both have been implicated in the Toll receptor pathways leading to the activation of NF-kappa B and JNK. DTRAF1 is most closely related to mammalian TRAF4 which is predominantly expressed during nervous system development and in ephitelial progenitor cells. In order to gain insight into possible roles of DTRAF1 during development, we have performed a detailed transcriptional analysis of the gene at various embryonic and larval stages. PMID- 11118895 TI - Expression of the Xvax2 gene demarcates presumptive ventral telencephalon and specific visual structures in Xenopus laevis. AB - vax2 is a recently isolated homeobox gene, that plays an important role in controlling the dorso-ventral patterning of the retina. In this paper we present a thorough description of the Xvax2 expression pattern all along Xenopus embryogenesis, and compare this pattern in detail to that shown by Xvax1b and Xpax2, two genes also involved in ventral eye development. At early neurula stages, while Xpax2 starts to be expressed within the eye field, both Xvax2 and Xvax1b are exclusively activated in the presumptive ventral telencephalon. Since midneurula stages, Xvax2 and Xvax1b are also transcribed in the medial aspect of the eye field. At tailbud and tadpole stages, Xvax2, Xvax1b and Xpax2 expression overlaps in the optic stalk and nerve and in the optic disk, while Xvax2 and Xvax1b also display specific activation domains in the ventral retina as well as in the ventral telencephalon and diencephalon. Finally, during metamorphosis a high level of both Xvax2 and Xvax1b transcription is maintained in the optic chiasm. In addition, Xvax1b is transcribed in the ventral hypothalamus and in the hypophysis, whereas a strong Xvax2 expression is retained in the ventral portion of the mature retina. PMID- 11118896 TI - Neuralin-1 is a novel Chordin-related molecule expressed in the mouse neural plate. AB - Cysteine-rich repeats (CRs) of the type described in Chordin constitute conserved domains present in an expanding family of secreted molecules. These motifs were shown to mediate directly the antagonism of BMP signaling by Chordin and play a major role during development. Here we report the cloning and expression pattern of neuralin-1, a new member of the chordin family. The mouse cDNA was cloned by homology with a human genomic sequence encoding putative CRs. In the human genome, neuralin-1 transcripts are encoded by 8 exons that span a region of at least 80 kilobases located on chromosome Xq22.1-23. Neuralin-1 is a 333 amino acid protein containing three CRs, two of them highly similar to the Chordin CRs that bind BMP. Like chordin, neuralin-1 is able to induce secondary axes after mRNA injection in Xenopus embryos. Interestingly, during late gastrulation, neuralin-1 and chordin present distinct and complementary expression patterns in the mouse: neuralin-1 expression starts in the neural plate at mid-gastrulation, whereas chordin expression at that stage is restricted to the node and midline mesendoderm. Later on, neuralin-1 expression becomes restricted to discrete regions of the central nervous system and to derivatives of the neural crest cells. During organogenesis, neuralin-1 presents a broad expression pattern in many tissues such as dorsal root ganglia, gut, condensing cartilages of the skeleton and developing hair follicles. PMID- 11118897 TI - Expression of a novel mouse gene 'mbFZb' in distinct regions of the developing nervous system and the adult brain. AB - Using an induction gene trap approach with bFGF, we identified a novel mouse gene mbFZb (mouse basic FGF repressed, putative Zic binding protein). It shows high homology to a putative Zic3 binding protein in Xenopus (AF129131). We used the in situ hybridization technique on sections to analyze the expression pattern of mbFZb during embryonic development in the mouse. From E9.5 onwards it is expressed mainly in the developing nervous system, with high levels of expression in the proliferating neuroepithel of the brain and the neural tube, the ganglia of the cranial nerves V, VII, VIII, IX and X and the dorsal root ganglia. MbFZbtranscript can also be detected in the developing eye and the brown fat. In the adult brain strong expression is restricted to the olfactory bulb, the hippocampus, the piriform cortex and the Purkinje cells of the cerebellum. On the cellular level the mbFZb/GFP fusion protein is localized in vesicular structures in the cytoplasm. PMID- 11118898 TI - The extraneuronal monoamine transporter Slc22a3/Orct3 co-localizes with the Maoa metabolizing enzyme in mouse placenta. AB - Monoamine clearance is a combined function of uptake mechanisms in the plasma membrane with intracellular metabolizing enzymes. Two different uptake mechanisms have been described. Uptake(1) is located in presynaptic neurones, whereas uptake(2) is extraneuronal. Recently, the Slc22a3/Orct3 gene was identified as the extraneuronal monoamine transporter. In mouse embryonic development Orct3 expression is restricted to the placenta, which is also a site of expression of neuronal transporters. We have used RNA blots and in situ hybridization to examine the expression of Orct3 and other members of the monoamine uptake and metabolizing pathways in mouse placenta. The results show that Orct3 expression overlaps that of the monoamine metabolizing enzyme Maoa in the labyrinth layer of the placenta with an expression pattern distinct from that of the neuronal transporters Slc6a2/Net and Slc6a4/Sert. PMID- 11118899 TI - Expression of Pbx1b during mammalian organogenesis. AB - Mammalian Pbx genes (Pbx1-3) encode a family of TALE homeodomain proteins that function as transcriptional regulators in numerous cell types (Curr. Opin. Genet. Dev. 8 (1998) 423). The present study highlights distinctive features of Pbx1b expression during mouse embryonic development as a framework to understand its biological functions. Immunohistochemical analyses demonstrate extensive expression of Pbx1b throughout post-implantation development, with highest levels observed during early to mid-gestation. Its initial distribution is predominantly associated with condensing mesoderm, however, Pbx1b displays dynamic expression patterns in derivatives of all principal germ layers. In particular, Pbx1b localizes to sites of mesenchymal-epithelial interactions during periods of active morphogenesis in tissues such as the lung, kidney, tooth buds and vibrissae follicles. Furthermore, BrdU labeling studies reveal that Pbx1b expression domains partially overlap with regions of cellular proliferation. Taken together, these data suggest that Pbx1b contributes to multiple cellular processes during embryogenesis, which may include roles in cell-autonomous regulation as well as in the mediation of tissue interactions. PMID- 11118900 TI - The vegetally localized mRNA fatvg is associated with the germ plasm in the early embryo and is later expressed in the fat body. AB - Vegetally localized RNAs in Xenopus oocytes have been implicated in the establishment of the primary germ layers and the formation and development of the primordial germ cells. fatvg mRNA is localized through the late pathway to the vegetal cortex. Like Vg1 mRNA fatvg is distributed throughout the entire cortex; however, unlike Vg1 there is a small fraction of the fatvg mRNA that is associated with the mitochondrial cloud. In early cleavage stage embryos, fatvg mRNA is associated with the germ plasm located at the tips of the vegetal blastomeres of the embryo. While several localized RNAs that follow the Message Transport Organizer (METRO) pathway have been found in the germ plasm in embryos, fatvg is a late pathway RNA that is associated with the germ plasm. In tadpoles, fatvg mRNA shows a novel pattern of expression which is distinct from the germ cell lineage and is detected at the dorso-anterior margin of the endodermal mass along the midline in two clusters of cells. fatvg mRNA expression is also detected later in the developing fat bodies, the major adipose tissues of the frog. PMID- 11118901 TI - Dynamic expression patterns of the pudgy/spondylocostal dysostosis gene Dll3 in the developing nervous system. AB - Defects in the Notch pathway ligand Dll3 have been identified in the mouse pudgy (Dll3(pu)) and human spondylocostal dysostosis (SD, MIM 277300) mutations. Although these mutations are primarily associated with segmental defects in the axial skeleton and somitic patterning, they also exhibit cranial neurological defects. Therefore we have looked at the expression of Dll3 in the developing mouse nervous system. The expression of Notch ligands and receptors shares common features at 10.75 dpc in the rhombic lips and dorsal hindbrain. Temporal analysis of Dll3 expression from 9.0 to 11.0 dpc reveals that it is strongly expressed in laminar columns linked with regions of neuronal differentiation and hindbrain segmentation. Transverse sections show that Dll3 is expressed in territories where commissural neurons are formed. We have also looked at neuronal patterning in the mid-hindbrain region in Dll3(pu) mutants. PMID- 11118902 TI - Culturability of Mycobacterium tuberculosis cells isolated from murine macrophages: a bacterial growth factor promotes recovery. AB - Very little is known about the culturability and viability of mycobacteria following their phagocytosis by macrophages. We therefore studied populations of the avirulent 'Academia' strain of Mycobacterium tuberculosis isolated from murine peritoneal macrophage lysates several days post-infection in vivo. The resulting bacterial suspensions contained a range of morphological types including rods, ovoid forms and coccoid forms. Bacterial viability measured using the MPN method (dilution to extinction in liquid medium) was often much higher than that measured by CFU (plating on solid medium). Viability in the MPN assay was further enhanced when the Micrococcus luteus protein, Rpf, was incorporated into the liquid culture medium at picomolar concentrations. Rpf is an example of a family of autocrine growth factors found throughout the high G+C cohort of Gram positive bacteria including M. tuberculosis. M. tuberculosis cells obtained from macrophages had altered surface properties, as compared with bacteria grown in vitro. This was indicated by loss of the ability to adsorb bacteriophage DS6A, a reduced tendency to form clumps, acquisition of ethidium bromide stainability following heat treatment, and loss of Rpf-mediated resuscitation following freezing and thawing. These results indicate that a proportion of 'unculturable' M. tuberculosis cells obtained from macrophages is either injured or dormant and that these cells may be recovered or resuscitated using Rpf in liquid medium. PMID- 11118903 TI - Amino acid- or protein-dependent growth of Trichophyton mentagrophytes and Trichophyton rubrum. AB - Culture conditions were examined for Trichophyton mentagrophytes and Trichophyton rubrum, which are major pathogens involved in dermatophytosis. They grew well in Sabouraud's dextrose broth or RPMI 1640. Growth in phosphate-buffered yeast nitrogen base supplemented with glucose was very slow, although growth improved significantly with the addition of amino acids or proteins to the medium. The fungi could also grow using human nail fragments as the only source of nutrition. Examination of proteases by substrate gel electrophoresis indicated that distinct sets of proteases are secreted from the dermatophytes in two different media, Sabouraud's dextrose broth and nail fragments. A protease inhibitor, phenylmethanesulfonyl fluoride, inhibited the growth of the fungi on nail fragments, but it did not inhibit their growth in Sabouraud's dextrose broth. PMID- 11118904 TI - Protection against respiratory syncytial virus (RSV) elicited in mice by plasmid DNA immunisation encoding a secreted RSV G protein-derived antigen. AB - Plasmid vectors encoding two different variants, one cytoplasmic and one secreted version, of a candidate vaccine BBG2Na to respiratory syncytial virus (RSV), were constructed and evaluated in a nucleic acid vaccination study. The two different vectors, which employed the Semliki Forest virus gene amplification system, were found to express BBG2Na appropriately in in vitro cell cultures. Immunisation of mice with the plasmid vectors elicited significant serum anti-BBG2Na IgG responses only in the mice receiving the plasmid encoding the secreted version of BBG2Na. Consistent with antibody induction data, sterilising lung protection against RSV-A challenge was also only observed in this group. These results indicate that the targeting of antigen expression (intracellular versus secreted) would be an important factor to consider in the design of nucleic acid vaccines. PMID- 11118905 TI - Transcutaneous immunisation with herpes simplex virus stimulates immunity in mice. AB - Herpes simplex virus (HSV) is common throughout the world and is a target for vaccine development. Transcutaneous immunisation is a novel technique that uses the application of vaccine antigens in solution on the skin in the presence of cholera toxin (CT) as an adjuvant. This study investigated the potential of transcutaneous immunisation in C3H mice, using CT co-administered with whole inactivated HSV-1 (CT+HSVi) or HSV-1 antigens extracted from infected Vero cells (CT+HSVag) or a control protein (CT+BSA). The application of any of the three vaccines on to bare mouse skin resulted in the migration of Langerhans cells from the epidermis and in the production of serum antibodies to CT. Both HSV preparations generated serum and mucosal (faecal) antibodies to HSV, with the CT+HSVi vaccine being a more potent stimulator of humoral immunity. The CT+HSVag vaccine, however, was the more potent stimulator of cell-mediated immunity, giving rise to a strong delayed type hypersensitivity response and lymphocyte proliferation in vitro. When the mice were challenged by epidermal inoculation of HSV, the CT+HSVag vaccine induced a higher level of protection than the CT+HSVi vaccine, a result which may indicate that the efficacy of HSV vaccines depends on stimulation of cell-mediated rather than humoral responses. The success of topical vaccination suggests that the transcutaneous route may offer a promising potential for novel vaccine delivery which merits further investigation. PMID- 11118906 TI - The relationship between O-chain expression and colonisation ability of Helicobacter pylori in a mouse model. AB - The influence of lipopolysaccharide (LPS) O-polysaccharide chain production on the colonisation ability of Helicobacter pylori in four mouse models (NMRI, C57BL/6, CBA/Ca, and BALB/cA mice) was studied. H. pylori strains that produced smooth-form LPS (S-LPS) detectable in silver-stained electrophoretic gels colonised mice. In contrast, a laboratory-passaged strain G50 and the culture collection strain CCUG 17874 did not colonise mice; the former strain produced low amounts of O-chains only detectable in immunoblotting but not in silver stained gels, whereas the latter produced rough-form LPS (R-LPS) without O chains. Furthermore, a galE isogenic mutant, which produced R-LPS, did not colonise mice. However, after repeated broth culture, strains G50 and CCUG 17874 produced S-LPS detectable in silver-stained gels and were capable of colonising mice. Consistent with the production of O-chains, all colonising strains produced Lewis (Le) antigens, Le(x) and/or Le(y). Except for low expression of Le(y) by non-colonising G50, reflecting low production of O-chains, all other non colonising strains and the galE mutant lacked expression of Le antigens consistent with their production of R-LPS. Lectin typing of strains supported these findings, and also showed that lectin types did not differ before and after colonisation. The low level of O-chain production and Le antigen expression by the non-colonising G50 may not be sufficient to aid colonisation. Examination of protein profiles of H. pylori strains before inoculation showed that protein expression was not significantly different between colonising and non-colonising strains. These results show that S-LPS production with O-chain expression is required by H. pylori for colonisation in a number of mouse models and that care should be taken with inoculating H. pylori strains that loss of O-chains does not occur during subculturing. PMID- 11118907 TI - Quantitative assessment of IgG antibodies to Helicobacter pylori and outcome of ischaemic heart disease. AB - Criticisms of serological studies on Helicobacter pylori and ischaemic heart disease (IHD) include: undiagnosed heart disease in live controls; no assessment of severity or outcome of IHD; and qualitative not quantitative measurements of IgG to the bacteria. The aim was to assess quantitatively IgG levels specific for H. pylori (ng ml(-1)) among patients who survived a myocardial infarction (MI) with those who died of IHD. Sera were from four groups: (1) men who survived one MI; (2) men matched for age and socioeconomic background to group 1; (3) individuals who died suddenly of IHD; (4) accidental deaths matched for age and sex to group 3. Levels of IgG to H. pylori increased with age (P<0.005) but were not associated with smoking or socioeconomic groups. There was a correlation between IgG to the bacteria and decreasing socioeconomic levels only among group 1 (P<0.01). IgG levels were higher for subjects who died of heart disease (median=151 ng ml(-1)) compared with survivors (median=88 ng ml(-1)) (P=0.034) and higher for survivors compared with their controls (median=58 ng ml(-1)) (P=0.039). Future serological studies of H. pylori in relation to IHD should be quantitative and severity of disease considered in analyses. PMID- 11118908 TI - Caspase-3 activation and apoptosis induction coupled with the retrograde transport of shiga toxin: inhibition by brefeldin A. AB - Caspase proteolytic activities, such as caspase-3, -2 and -6, of THP-1 human monocytic cells were markedly increased in a time- and dose-dependent manner by treatment with purified Shiga toxin 1 (Stx1) or Stx2. Caspase-3 activation was strictly correlated with internucleosomal DNA fragmentation and chromatin condensation of the cells. In addition, the specific caspase-3 inhibitor, Ac-DEVD CHO, decreased the percentage of apoptotic cells. The purified B-subunit of Stx1 did not induce apoptosis in THP-1 cells. Caspase-3 activation, DNA fragmentation and chromatin condensation caused by Stx were completely blocked by pretreatment of cells with brefeldin A, an inhibitor of Golgi functions. The findings suggest that Stx1 as well as Stx2 activate caspase-3, which plays a critical role in apoptosis, and that the apoptotic signals rise after Stx is transported to the Golgi apparatus. PMID- 11118909 TI - Detection of PCR products of Escherichia coli O157:H7 in human stool samples using surface plasmon resonance (SPR). AB - A method for the rapid detection of verotoxin-producing Escherichia coli O157:H7 in stools was evaluated. Strains possessing Shiga toxin-2 (stx-2) genes were isolated from stool samples and amplified using oligonucleotide primers. Stools spiked with cultured E. coli O157:H7 (strain 298 or strain 1646) were detected to be polymerase chain reaction (PCR) positive at 10(2) cfu per 0.1 g of stool. Stool samples from patients and healthy carriers showed a high correlation between positive results for a PCR and the presence of verotoxin-producing E. coli O157:H7, confirmed by isolation of serotype O157:H7 on sorbitol MacConkey medium (10 of 10 stool samples). These PCR products could be detected using a BIAcore 2000 surface plasmon resonance device using peptide nucleic acid as a sensor probe. In this report we use this method for the rapid detection of DNA from significant pathogenic organisms. PMID- 11118910 TI - Streptococcus pneumoniae heat shock protein 70 does not induce human antibody responses during infection. AB - Mouse monoclonal antibodies (mAbs) were developed against Streptococcus pneumoniae in search for potential common pneumococcal proteins as vaccine antigens. mAb 230,B-9 (IgG1) reacted by immunoblotting with a 70-kDa protein which was isolated by immunoaffinity chromatography and subsequent preparative electrophoresis. N-terminal amino acid sequencing showed homology to that of heat shock protein 70 (hsp70). The hsp70 epitope reactive with mAb 230,B-9 was found in all the pneumococci examined as well as in other streptococci and enterococci. The epitope was not expressed in several other examined Gram-positive or negative bacteria. Pneumococcal hsp70 has by other investigators been proposed to be a vaccine candidate. Binding experiments using flow cytometry showed that the epitope was not surface-exposed on live exponential phase grown S. pneumoniae. Human patient sera did not react with affinity-purified pneumococcal hsp70. Therefore the pneumococcal hsp70 does not seem to be of special interest in a vaccine formulation. The human sera contained antibodies to high molecular proteins co-purified with hsp70. Some of these proteins could be the pneumococcal surface protein A. PMID- 11118911 TI - A high molecular mass constituent of cranberry juice inhibits helicobacter pylori adhesion to human gastric mucus. AB - Because previous studies have shown that a high molecular mass constituent of cranberry juice inhibited adhesion of Escherichia coli to epithelial cells and coaggregation of oral bacteria, we have examined its effect on the adhesion of Helicobacter pylori to immobilized human mucus and to erythrocytes. We employed three strains of H. pylori all of which bound to the mucus and agglutinated human erythrocytes via a sialic acid-specific adhesin. The results showed that a high molecular mass constituent derived from cranberry juice inhibits the sialic acid specific adhesion of H. pylori to human gastric mucus and to human erythrocytes. PMID- 11118912 TI - Culture supernatants of patient-derived Aspergillus isolates have toxic and lytic activity towards neurons and glial cells. AB - Infection of the central nervous system by the ubiquitous fungi Aspergillus spp. is a life-threatening disease. Therefore we investigated the mechanism of brain damage by fungal infection. To examine whether secretory factors of Aspergillus isolates derived from patients can induce death of different brain cells, culture supernatants of Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus and Aspergillus niger were added to different astrocytes as well as to the neuroblastoma cell line SK-N-SH, and to the microglial cell line CHME. All four fungal species were shown to secrete toxic factors with neurons being most sensitive against these factors. Very low amounts and short incubation times are sufficient to induce irreversible cell damage, indicating that secreted factors might also affect distant brain regions. Further characterization of the toxic factors revealed that A. fumigatus and A. terreus produced small, heat-stable components whereas the toxic activity of A. niger filtrates was triggered by a high molecular mass factor which could be inactivated by heat. The active component of A. flavus had a molecular mass similar to that of A. niger but was heat-stable and had a significantly lower activity. Taken together these results indicate that secretion of different necrotizing factors might contribute to brain lesions in patients with cerebral aspergillosis. PMID- 11118913 TI - Identification of the secreted macromolecular immunogens of Staphylococcus aureus by analysis of serum. AB - The ability of sera to recognise secreted macromolecules of Staphylococcus aureus was examined by ELISA and Western immunoblotting. Individual secreted proteins were also studied using both human sera and sera from rabbits immunised with secreted macromolecules. Patients sera showed a wide range of IgG antibody titres to secreted macromolecules and whole bacteria. Controls showed a significantly lower IgG response. Western immunoblotting revealed that a significant number of secreted proteins were recognised by circulating IgG antibodies. Surprisingly, both the sera from controls and from patients recognised similar macromolecules including a number of potential virulence factors. The major difference was in the IgG binding to a 16-kDa component, which was recognised by the majority of the sera from infected individuals, but only by a small number of sera from healthy controls. The higher incidence of antibodies recognising the 16 kDa component may be related to our earlier finding that the major bone resorbing component of S. aureus is a heterodimeric protein containing a 16-kDa subunit, the activity of which could be blocked by sera. PMID- 11118914 TI - Immune cell functions, lipids and host natural resistance. AB - Nutritional status may exert a profound effect on immune system functions. Hence, several parameters of immune system are modified by dietary lipid administration, as lymphocyte proliferation, cytokine production, natural killer activity, antigen presentation, etc. Thus, numerous studies have indicated the key role of lipids as immune response modulators. These properties have been applied in the treatment of autoimmune and inflammatory diseases. As a result, the reduction or suppression of immune status due to lipid incorporation promotes an impairment in the ability of host natural response to eliminate infectious microorganisms as bacteria or parasites. In the present review, we analyze the current status about the relationship among dietary lipids, reduction of immune parameters and reduction of host natural response against infectious diseases. Many discrepancies are discussed, although several studies indicate a close association between dietary lipid manipulation and impairment in the elimination of bacteria, viruses or parasites. On the other hand, other studies point out a beneficial effect of dietary lipid manipulation on the host natural response. Future investigations will determine the events involved in the regulation of immune response by fatty acids and their role in the elimination of pathogenic agents. PMID- 11118915 TI - The effects of Cryptococcus neoformans-secreted antigens on tumor necrosis factor alpha-induced intercellular adhesion molecule-1 expression on human lung epithelial cells. AB - Since primary infection with Cryptococcus neoformans usually occurs in the lungs, and since pulmonary cryptococcosis involves interactions between yeasts and alveolar epithelial cells, we have begun to study the effects of C. neoformans and its secreted antigens (SA) on epithelial reactions potentially associated with localized inflammation. We report here that SAs from encapsulated and acapsular strains of C. neoformans caused significant reductions in tumor necrosis factor-alpha (TNF-alpha)-induced intercellular adhesion molecule-1 (ICAM 1) expression on A549 lung epithelial cells in culture. We also present evidence that the reduction in ICAM-1 expression was not associated with SA-induced shedding of this adhesion molecule. PMID- 11118916 TI - Effect of cisapride on portal haemodynamics in patients with liver cirrhosis using duplex Doppler ultrasonography. AB - PURPOSE: Cisapride, a benzimide derivative, is a gastrointestinal prokinetic agent without dopamine-antagonistic or cholinomimetic effects. This study aims at assessing the effect of cisapride oral administration on portal flow in patients with advanced post hepatitic cirrhosis using duplex Doppler ultrasound (US). METHODS: A total of 12 patients with post-hepatitic liver cirrhosis were included in the study. Duplex Doppler sonographic examinations were performed before and after treatment. The subjects received 10 mg cisapride before starting the measurement procedure and then three times a day for 2 days. Portal haemodynamics including vessel diameters (mm), mean flow velocities (cm/s), blood flows (ml/min) were investigated. RESULTS: Mean portal vein diameters, mean portal flow velocity and portal blood flow volume showed decreases of 18.6, 22.1 and 43.6% (P<0.001), respectively. After cisapride administration the portal vein diameter did not change in two patients and the portal vein velocity did not change in three patients. No significant change was found in systolic blood pressure, diastolic blood pressure or pulse rate after the administration of cisapride. CONCLUSION: In this study, it was demonstrated that oral administration of cisapride results in a significant reduction of portal blood flow but there were no changes in heart rate or systolic pressure in patients with cirrhosis of the liver. PMID- 11118917 TI - Correlations among prostatic biopsy results, transrectal ultrasound findings and PSA levels in diagnosing prostate adenocarcinoma. AB - OBJECTIVE: The objective of this study was to evaluate transrectal ultrasound (TRUS) findings and prostate-specific antigen (PSA) levels in relation to prostatic biopsy results and to analyze their individual and combined performances in diagnosing prostate adenocarcinoma (PAC). METHODS: Men (n=143) with PSA levels above 4 ng/ml underwent TRUS and randomized ultrasound-guided prostatic biopsy through the peripheral zone, including additional hypoechoic nodules biopsies, if they were noted on TRUS. Data related to TRUS, biopsy, and PSA level results were then correlated. RESULTS: A significant correlation between TRUS images suspicious for PAC and a biopsy-confirmed diagnosis of PAC, or between the lack of such images and a negative biopsy result, was not found. However, a significant correlation was found between positive biopsy results and PSA levels greater or equal to 10 ng/ml. The sensitivity of transrectal ultrasound in making a diagnosis of PAC was 63%, whereas its specificity was 73%. CONCLUSION: We conclude that while the separate performances of these examinations were not effective in diagnosing PAC, the integrated use of these methods was more adequate for making the diagnosis. PMID- 11118918 TI - Ultrasound-guided surgery of deep seated brain lesions. AB - OBJECTIVE: Computer aided navigation systems have been introduced to optimize the neurosurgical strategies minimizing the damage to the healthy brain tissue. As the loss of cerebrospinal fluid and surgical manipulation alter the position of the lesion in the external reference system, there is a risk of being misguided to deep seated brain tumors. In this context we present our experiences with intraoperative ultrasound which represents a real-time navigation system. PATIENTS AND METHODS: 45 patients with subcortical intracerebral lesions were operated on with the support of ultrasound imaging. Tumor depth from the surface measured 5 (superficial) to 68 mm. The minimum size of a cavernoma was 8 mm. Histopathological diagnoses included 17 cavernomas, 12 metastases and 16 gliomas. Ultrasound localization was achieved by two perpendicular projections and the surgical trajectory of the lesion secured by catheter placement. RESULTS: Intraoperative ultrasound allowed an easy identification of all brain lesions. Additionally, a reliable localization of the tumor margins, including gliomas and metastases, was reached in 23 of 28 (82%) cases. The remaining 17 cavernomas were located all by intraoperative ultrasound, even the deep seated brain lesions of less than 10 mm in diameter could be easily detected. The real-time mode enabled a control of the surgical procedure. Technical problems arose from irregular and vaulted cortex surface, head positioning and surgical manipulation causing air artefacts with distal reduction of the signal intensity. CONCLUSION: Intraoperative ultrasound is a reliable intraoperative guidance tool in the localization of deep seated brain tumors because it operates in real-time, thus increasing the safety of the surgical procedure and of the tumor resection. PMID- 11118919 TI - A new Doppler signal enhancing agent for flow assessment in breast lesions. AB - OBJECTIVE: To determine the diagnostic performance of SonoVue (Bracco) in the enhancement of Doppler signals in breast lesions and in the improvement of diagnostic accuracy. METHODS: This multicenter study included 220 patients undergoing investigations of parenchymal lesions, 40 of which had breast tumors. After a baseline Doppler examination, intravenous doses of 0.3, 0.6, 1.2 and 2.4 ml SonoVue were injected. Doppler signal quality before and after injection was compared. Off-site assessment of the global quality of Doppler signal and duration of clinical useful enhancement, as well as off-site and on-site evaluation of quality of color and spectral Doppler, were performed. On-site evaluation of diagnostic accuracy was also carried out. Safety assessments included monitoring of adverse events up to 24 h following the last injection of SonoVue. RESULTS: On-site evaluations: baseline Doppler was conclusive in only 4/21 carcinomas and in 2/17 benign lesions. Enhanced Doppler improved differential diagnosis in 20/21 carcinomas and in 9/12 benign lesions. Time to color enhancement was 0.55 min for the lowest and 0.35 min for the highest dose. The total duration of enhancement was 3.47 min for the lowest and 5.62 min for the highest dose, respectively. Off-site assessment: SonoVue improved the quality of Doppler blood flow information both in parenchymal and focal lesions. Statistically significant changes from baseline in global quality of Doppler investigations were observed at all four SonoVue doses (P<0.05). The duration of clinically useful signal enhancement increased with doses and a significant dose relationship was obtained (P<0.001). Mild adverse events were observed in two patients only. CONCLUSION: The results obtained from this study, following both off-site and on-site assessment, demonstrate that the administration of SonoVue to patients with focal breast lesions provides significant improvement over the baseline of Doppler signal quality and a clinically useful duration of signal enhancement, related to the dose. SonoVue was shown to be a safe and well tolerated compound. PMID- 11118920 TI - Color and pulsed Doppler ultrasound findings in normally functioning transjugular intrahepatic portosystemic shunts. AB - OBJECTIVE: To evaluate blood flow changes inside normally functioning transjugular intrahepatic portosystemic shunts (TIPS), using color Doppler ultrasound (CDUS) and pulsed Doppler ultrasound (PDUS). METHODS: A total of 72 patients (mean age 54, range 36-78 years) underwent TIPS placement, portal angiography, CDUS and PDUS examinations. Measurements inside the stent were taken at the portal side, at the central part and at the venous end of the TIPS. RESULTS: In well functioning TIPS the mean peak velocity (PV) on the portal side was 37 m/s (range 22-65 cm/s), in the area of the incoming intrahepatic portal branch the mean PV was 59 cm/s (range 40-95 cm/s) and at the side of the incoming hepatic vein the mean PV was 135 cm/s (range 88-220 cm/s). In the punctured hepatic and portal veins the mean PV was 25 cm/s (range 15-30 cm/s) and 18 cm/s (10-22 cm/s), respectively. The flow increase from the portal to the mid part (P less than or = 0.001) and to the venous side was statistically significant (P less than or = 0.001). CONCLUSION: A velocity gradient between the portal and the venous side of a TIPS is a normal finding caused by branches of the portal and hepatic vein joining the TIPS from the side and it is characteristic of a normally functioning TIPS. PMID- 11118921 TI - Variability in Doppler ultrasound influences referral of patients for carotid surgery. AB - OBJECTIVE: Colour Doppler ultrasound is operator dependent, but it is unclear how much clinical impact this might have on patient referral for carotid endarterectomy. Our aim was to quantify the interobserver variability of Doppler ultrasound. METHODS: Consecutive patients attending for carotid Doppler ultrasound underwent two examinations on the same day, in random order, by two of three observers blind to each other's results. Severity of stenosis was assessed using standard velocity criteria and lesion appearance. RESULTS: A total of 189 patients were scanned (378 ICAs). Of the 134 ICAs scanned by observers 1 and 2, observer 1 classified 11 as 80-99% stenosis (operable), compared with nine by observer 2. Of the 206 ICAs scanned by observers 1 and 3, observer 1 classified 11 as 80-99% stenosis, compared with only five by observer 3. Of the 38 ICAs scanned by observers 2 and 3, observer 2 classified 2 as 80-99% stenosis compared with none by observer 3. Overall, clinical management would differ in 10/378 (3%) of ICAs, but in 10/22 (45%) of those considered operable by one of the three observers. CONCLUSION: There was clinically important interobserver variability in the assessment of ICA disease, which could result in serious errors if endarterectomy were performed on the basis of a single Doppler ultrasound. PMID- 11118922 TI - Ultrasound-guided percutaneous ethanol injection under general anesthesia for the treatment of hepatocellular carcinoma on cirrhosis: long-term results in 268 patients. AB - OBJECTIVE: Percutaneous ethanol injection (PEI) under general anesthesia (One shot PEI) is a therapy for large and multiple hepatocellular carcinoma (HCC) by the injection of a large amount of ethanol into the tumor. We report our results with 5-year survival rates in patients with HCC on cirrhosis treated with One shot PEI. PATIENTS AND METHODS: From October 1992 to March 1998, 268 cirrhotic patients (age 42-82 years; 191 males; 95 Child-Pugh's A class, 150 B and 23 C class of cirrhosis) with 515 HCC nodules underwent One-shot PEI. Diameter of HCC nodules ranged from 0.6 to 14 cm (mean 5.02 +/- 2.2 cm; median: 4 cm). One hundred and thirty-eight patients had a single nodule (range 3.2-14 cm; mean 5.6 +/- 2.1 cm), 130 had multiple nodules, up to six nodules (mean 2.9 nodules) (range 0.6-11 cm; mean 4.8 +/- 2.1 cm) RESULTS: CT showed complete necrosis in 357/506 nodules (70%). Five patients (1.8%) with nine nodules died as a result of the procedure (variceal bleeding in three cases, liver failure in one and hemoperitoneum in one). The overall survival rates were 93, 83, 74, 65 and 59% at 1, 2, 3, 4 and 5 years, respectively. Survival rates were 90, 84, 82 and 82% at 12, 24, 36 and 48 months, respectively, in patients with a single nodule less than or = 5 cm, and 97, 71, 59, 59 and 59% at 12, 24, 36, 48 and 60 months, respectively, in patients with single nodule >5 cm. Patients with multiple nodules had survival rates of 97, 89, 75, 60 and 60% at 12, 24, 36, 48 and 60 months, respectively. CONCLUSION: PEI of large and multiple HCC showed survivals similar to conventional PEI for patients with smaller tumors. PMID- 11118923 TI - Doppler ultrasound diagnosis of a superficial ulnar artery. AB - The ulnar artery may lie in a superficial position in the forearm as a normal anatomical variant. We report a case where this variant was clinically mistaken for phlebitis but was diagnosed correctly with Doppler ultrasound. In this report, we discuss the anatomy and clinical relevance of the superficial ulnar artery. PMID- 11118924 TI - Clinical report: contrast enhancement of tumor perfusion as a guidance for biopsy. AB - We describe three cases where biopsies from various tumors were guided by contrast enhancement of tumor perfusion. After i.v. administration of Levovist (Schering AG, Berlin, Germany), the tumors showed both hyper- and hypovascular areas. Biopsies from the latter showed marked necrosis or fibrosis. This may cause biopsies not being conclusive. To ensure fully diagnostic biopsies from irregular tumors we propose the biopsy to be performed from the most vascular part demonstrated by contrast enhancement. PMID- 11118925 TI - Diagnosis of acute rupture of the anterior cruciate ligament of the knee by sonography. AB - The accuracy of sonography in the diagnosis of acute rupture of the anterior cruciate ligament (ACL) was tested. Sixty-two patients with a recent traumatic haemarthrosis were examined. A haematoma at the origin of the ACL in the femoral intercondylar notch was interpreted as evidence of ligament injury. The standard of reference was arthroscopy or clinical follow-up. The sonographic findings were confirmed in 59 of 62 cases. The sensitivity was 88%, the specificity 98%, and the positive and negative predictive values 93 and 96%. PMID- 11118926 TI - Laparoscopic ultrasound of the liver. AB - OBJECTIVE: Despite recent advances in medical imaging, pre-operative evaluation of liver tumors, whether benign or malignant, is often lacking in accuracy and precision. With the development of surgical laparoscopy, the benefits of diagnostic laparoscopy have been combined with those of operative ultrasound. This article aims to describe the technique of laparoscopic ultrasound of the liver, and to define its applications and the role of its association with diagnostic laparoscopy in the localization and assessment for resectability of liver tumors. METHODS: After an initial visual inspection with the laparoscope, laparoscopic ultrasound is utilized to further examine the liver. This relies largely on recognition of branches of the portal vein and tributaries of the hepatic veins. During this procedure, the hepatic parenchyma is also examined. Minimal displacement of the transducer, using clockwise and anti-clockwise rotatory movements, allows a full exploration of the liver. RESULTS: The combination of visual with sonographic laparoscopy allows accurate localization of benign and malignant hepatic tumors, as well as ultrasound-guided biopsies of these. Laparoscopic ultrasound can detect small lesions previously unseen by pre operative imaging techniques. The relationship of tumors to adjacent blood vessels can be defined. Portal vein thrombosis can be diagnosed. CONCLUSION: Curability and liver tumor resectability can be determined and the appropriate surgical treatment thus planned. PMID- 11118927 TI - American College of Epidemiology Ethics Guidelines: filling a critical gap in the profession. PMID- 11118928 TI - Reliability of Chalmers' scale to assess quality in meta-analyses on pharmacological treatments for osteoporosis. AB - PURPOSE: This study estimates the inter-rater and test-retest reliability of Chalmers' quality score scale in the context of bone mass loss and fracture rate in postmenopausal women. METHODS: An exhaustive literature search was performed on Medline to locate clinical trials studying the effect of medication use on bone mass loss and fracture rate in postmenopausal women. Twenty articles were randomly selected and four raters independently assessed the quality of each article with Chalmers' scale. Among the 20 articles, 10 were blinded on authors' names, journal, year of publication and source of funding. Raters were also asked to assess all 20 articles one more time, two months after the first evaluation. Intraclass (ICC) and test-retest correlation coefficients were calculated. RESULTS: The overall inter-rater ICC was 0.66 [0.55, 0.79](95%). The overall test retest reliability of Chalmers' scale was 0.81 [0.67, 0. 98](95%). When ratings were stratified according to articles' blinding status, blinded assessments generated a smaller inter-rater ICC than non-blinded assessments: 0.30 [0.17, 0.53](95%) vs. 0.80 [0. 71, 0.90](95%). In addition, analyzing sub-scales separately generated different estimates of reliability. CONCLUSIONS: This study shows that the reliability of the quality scale developed by Chalmers substantially varies between sub-scales, and is highly dependent on articles' blinding status. The possibility of bias in rating non-blinded articles can not be ruled out. The reliability of the scale can also be dependent on the outcome studied. PMID- 11118929 TI - Electronic telephone directory listings: preferred sampling frame for a population-based study of childhood cancer in Australia. AB - PURPOSE: We report our telephone-based system for selecting community control series appropriate for a complete Australia-wide series of Ewing's sarcoma cases. METHODS: We used electronic directory random sampling to select age-matched controls. The sampling has all listed telephone numbers on an up-dated CD-Rom. RESULTS: 95% of 2245 telephone numbers selected were successfully contacted. The mean number of attempts needed was 1.94, 58% answering at the first attempt. On average, we needed 4.5 contacts per control selected. Calls were more likely to be successful (reach a respondent) when made in the evening (except Saturdays). The overall response rate among contacted telephone numbers was 92.8%. Participation rates among female and male respondents were practically the same. The exclusion of unlisted numbers (13.5% of connected households) and unconnected households (3.7%) led to potential selection bias. However, restricting the case series to listed cases only, plus having external information on the direction of potential bias allow meaningful interpretation of our data. CONCLUSION: Sampling from an electronic directory is convenient, economical and simple, and gives a very good yield of eligible subjects compared to other methods. PMID- 11118930 TI - Urinary tract infection: self-reported incidence and associated costs. AB - PURPOSE: To estimate the annual incidence, cumulative probability of presumed urinary tract infection (UTI) by age, and the social costs. METHODS: Analysis of a random digit dialing survey of 2000 women in the United States. RESULTS: 10.8 percent (95% CI: 9.4, 12.1%) of women aged 18 and older reported at least one presumed UTI during the past 12 months, with the majority of the cases occurring among women with a history of two or more UTI episodes in their life. We estimate that by age 24, one-third of women will have at least one physician-diagnosed UTI that was treated with prescription medication. Overall, an estimated 11.3 million women in the United States had at least one presumed UTI treated with antibiotics in 1995. We estimate the annual cost of UTI cases with prescriptions to be $1.6 billion in 1995. If the costs occurring after 1995 are discounted at 5% annually, the total cost over 20 years has a present value of $25.5 billion. CONCLUSION: If a vaccine were developed that would prevent either initial or recurrent UTI the net benefits to society would be substantial, even at a developmental cost of one billion dollars. PMID- 11118931 TI - The perinatal advantage of Mexican-origin Latina women. AB - PURPOSE: To determine if there is a perinatal advantage for birth outcomes among Mexican-origin Latina (Latina) women compared to white non-Hispanic (white) women, after adjusting for maternal, paternal, and infant factors. METHODS: 1,439,583 births from the 1990-1993 California linked birth and infant death certificate data sets were analyzed for the risk of low birth weight infants and infant mortality. RESULTS: Latina women had a statistically higher unadjusted risk of low birth weight infants and infant mortality compared to white women. After adjusting for potential confounders, Latina women had a similar risk of low birth weight infants and a lower risk of infant mortality relative to white women. In multivariate analyses, the most significant risk factor for low infant birth weight was young gestational age (OR = 82.91 for gestational age 1-230 days and OR = 10.62 for gestational age 231-258 days) and the most significant risk factor for infant mortality was low birth weight (OR = 53.99 for infant birth weight <500 grams and OR = 9.27 for infant birth weight 500-2499 grams). CONCLUSION: There was some evidence of a perinatal advantage for Latina women, when compared to white women and after adjusting for numerous potential confounders. To further reduce the risk of low birth weight infants and infant mortality, additional research is needed for etiologic clues beyond race/ethnicity and other traditional risk factors. PMID- 11118933 TI - The differential effect of education and occupation on body mass and overweight in a sample of working people of the general population. AB - PURPOSE: To assess whether two indicators of social class, education and occupation, have independent and/or synergistic effects in determining the body mass and overweight. METHODS: Body mass index (BMI), education, and occupation were assessed in a survey of 1767 men and 1268 women from a representative sample of currently working people of the general population of Geneva, Switzerland. Education and occupation were categorized as low, medium, and high. Overweight was defined as BMI > or = 25 kg/m(2). RESULTS: The prevalence of overweight was 52.1% in men and 28.7% in women. Men with overweight were more likely to have low education while women with overweight had lower education and lower occupation. Education and occupation were inversely related to BMI in both genders and, in women, had a synergistic effect (p-value for the interaction = 0.03). CONCLUSIONS: Education and occupation have independent and, in women, synergistic effects on BMI. The two indicators may express different mechanisms through which low social class is related to high body mass. PMID- 11118932 TI - Effects of the ApoE polymorphism on plasma lipoproteins in Mexican Americans. AB - PURPOSE: To evaluate the impact of apolipoprotein E (apoE) genotypes on lipoprotein measurements relative to that of other known cardiovascular risk factors in participants of a large population-based family study. METHODS: We measured concentrations of apoE, the major constituents of HDL (cholesterol, apoAI), LDL-C (cholesterol and apoB), and fraction of apoE in lipoprotein size classes in 859 participants of the San Antonio Family Heart Study, and then tested the association between the three common apoE genotypes (epsilon2epsilon3, epsilon3epsilon3, and epsilon3epsilon4) and lipoprotein traits using the measured genotype approach to account for residual familial correlations. RESULTS: Allele frequencies in this population for epsilon2, epsilon3, and epsilon4 were 3.5%, 89.6%, and 6.9%, respectively. As expected, adjusted apoE concentrations were highest in those with epsilon2epsilon3, intermediate in those with epsilon3epsilon3, and lowest in those with epsilon3epsilon4. The concentrations of total cholesterol, LDL-C and apoB were lowest in those with epsilon2epsilon3, intermediate in those with epsilon3epsilon3, and highest in those with epsilon3epsilon4. There was no significant effect of apoE genotypes on triglycerides, HDL-C, or apoAI levels. Compared to subjects with epsilon3epsilon4, subjects with epsilon2epsilon3 had relatively less apoE in LDL and HDL(1), and relatively more in HDL(2) and HDL(3) size fractions. The effect of apoE genotypes was significantly greater on apoB in women than in men. ApoE genotypes accounted for 4.5%, 12.3%, and 4.7% of the total genetic variation in apoB, apoE, and LDL-C, respectively. CONCLUSION: ApoE genotypes account for a modest, albeit significant, proportion of phenotypic variation in concentrations of LDL-C, apoB, and apoE, and distributions of apoE among lipoproteins in this population; these genotypes have a greater effect on apoB levels in women than in men. PMID- 11118934 TI - Short-term storage of blood samples and DNA isolation in serum separator tubes for application in epidemiological studies and clinical research. AB - PURPOSE: To investigate the use of a simple DNA isolation technique for application in epidemiologic studies. To analyze systematically the potential impact of lag time between blood drawing and DNA isolation and the condition of storage of blood samples on the quantity and quality of isolated DNA in large scale epidemiologic studies. METHODS: A modified single tube DNA isolation technique was used. DNA was isolated from samples collected from six participants and processed in triplicate: a) without delay after blood drawing; b) after blood cells were stored at 4 degrees C for 7 days; c) after blood cells were stored at 70 degrees C for 7 days; and d) after storage for 7 days at -70 degrees C with addition of lysis/digestion buffer. Polymerase chain reaction (PCR) and Southern blot analysis were performed to analyze the quality of the isolated DNA. RESULTS: The average amount of DNA isolated ranged from 27.0 to 71.1 microg/4.5 ml whole blood. Storage at 4 degrees C yielded, on the average, 20% less DNA than the samples processed without delay or after storage at -70 degrees C, although this difference was not statistically significant. All four conditions studied allowed isolation of highly pure DNA suitable for genetic analyses by Southern blot analysis and polymerase chain reaction. CONCLUSIONS: This pilot study suggests that storage for 7 days and at different temperatures allows isolation of high quality DNA. Using the described technique, storage of up to 7 days permits processing of large numbers of samples (50-70) in a single day, allowing for a reliable and cost-efficient way of processing in various settings. Further studies are needed to investigate the influence of long-term storage of biological specimens on DNA isolation and quality. PMID- 11118935 TI - Effects of intra-abomasal infusion of beta-casomorphins on circulating concentrations of hyperglycaemic insulin and glucose in dairy cows. AB - The effects of intra-abomasal infusion of a mixture of -casomorphins on circulating concentrations of insulin and glucose prestimulated by either abomasal (experiment 1) or intravenous (experiment 2) glucose were studied using non-lactating dairy cows. In both experiments, bolus infusion of 240 mg of a mixture of three beta-casomorphins (beta-casomorphin-4-amide, -5 and -7) was given via an abomasal infusion line. The beta-casomorphins significantly lowered the responses of serum insulin to both abomasal and intravenous glucose infusions (P<0.05). However, the beta-casomorphins did not significantly affect circulating glucose concentrations. The insulinopenic action of the beta-casomorphins is consistent with the action of somatostatin-28 (SS-28) as judged from the effects of SS-28 on the insulin secretion when administered intravenously in experiment 1. PMID- 11118936 TI - Nitrogen excretion and changes in blood components during emersion of the subtidal spider crab Maia squinado (L.). AB - Survival ability of Maia squinado to emersion and subsequent reimmersion was determined in winter and summer conditions. Male spider crabs were less tolerant of emersion than females. Emersion (up to 24 h in summer and to 48 h in winter) induced a marked reduction of nitrogen excretion, especially ammonia excretion. Increase in blood ammonia content was rapid and very high in summer (1750 micromol l(-1)), but non-lethal levels. Estimation of the body ammonia overload showed that only 30% of unexcreted ammonia accumulated in blood. The ammonia release at reimmersion indicated that ammonia also accumulated in other body compartments. Increase in blood urate content, which indirectly reduces ammonia production, was similar at both seasons. Emersed M. squinado was rapidly resorting to anaerobic metabolism, especially in summer when its blood haemocyanin content is low. A strong hyperglycemia was developed in the first 12 h of emersion at both seasons. Mortality occurring beyond 24 h of reimmersion, when the body ammonia overload is cancelled and the recovery of most of blood components is achieved, remains unexplained. PMID- 11118937 TI - Stable lead isotope ratios from distinct anthropogenic sources in fish otoliths: a potential nursery ground stock marker. AB - Variations measured in the lead (Pb) stable isotope ratios in otoliths of juvenile tropical reef fish Scarus perspiculatus, Abudefduf abdominalis and Dascyllus albisella reflect mixing of anthropogenic lead from the Kaneohe Bay watershed and 'background' lead characteristic of the adjacent ocean. The otoliths and water samples collected in a transect across the bay demonstrated nearly identical Pb isotopic trends. The Pb isotopic composition of the watershed has a low 206Pb/204Pb signature primarily reflecting past combustion of tetra ethyl Pb additive in fuels. Ocean water not contaminated by this watershed signature has a different, high 206Pb/204Pb isotopic composition, similar to previously measured Asian anthropogenic aerosols and natural eolian dusts, where the anthropogenic signal dominates. Where a history of past anthropogenic Pb contamination exists, it may be possible to use the ratios of Pb stable isotopes in fish otoliths to reconstruct the nursery grounds of fish. PMID- 11118938 TI - Aerial ventilatory responses of the mudskipper, Periophthalmodon schlosseri, to altered aerial and aquatic respiratory gas concentrations. AB - Periophthalmodon schlosseri is a mudskipper which uses the vascularized buccopharyngeal cavity as a respiratory organ. The fish construct mud burrows that contain hypoxic water, but store air inside the burrows. Because the burrow gas is frequently hypoxic and hypercapnic, the effects of altered respiratory gas concentrations on the aerial ventilation frequency (V(F)), inspiratory tidal volume (V(T)) and minute volume (V(M)=V(F)xV(T)) of P. schlosseri were studied by pneumotachography. Both total buccopharyngeal gas volume (V(BP)) and V(T) scaled significantly with body mass (mass exponents=1.10 and 1.03, respectively), and V(T)/V(BP) was 0.54+/-0. 05 (S.E.M., n=6). V(BP), expressed as a percentage of body volume, was much higher (16%) than in other air-breathing gobies (2-4%). When fish respired in normoxic air and water, V(F) was 0.25+/-0.04 breaths min( 1), V(T) 7.6+/-0.6 ml 100 g(-1), and V(M) 1.80+/-0.18 ml 100 g(-1) min(-1). Aquatic hypoxia did not significantly affect V(F), V(T), or V(M). In both moderate (P(O(2))=10 kPa) and severe (P(O(2))=5 kPa) aerial hypoxia, V(F) and V(M) increased significantly. V(T) increased significantly only during severe aerial hypoxia. In aerial hypercapnia, V(F) and V(M) increased significantly. PMID- 11118939 TI - Carotenoid pigments and trophic behaviour of deep-sea shrimps (Crustacea, decapoda, alvinocarididae) from a hydrothermal area of the mid-atlantic ridge. AB - Pigments and trophic behaviour of three species of Alvinocarididae from a Mid Atlantic hydrothermal site were analysed. Carotenoid pigments are responsible for the more or less marked colouration of these animals. The carotenoid content of whole animals and different tissues were evaluated. Rimicaris exoculata exhibits an increased carotenoid level at the juvenile stage, while Chorocaris chacei and Alvinocaris markensis contain only few traces of pigment. Free and esterified astaxanthin, reported for most pelagic crustaceans, are present in these deep-sea shrimps. The origin of carotenoids of crustaceans living in the aphotic zone is discussed. PMID- 11118940 TI - GABA, 5-HT and amino acids in the rotifers Brachionus plicatilis and Brachionus rotundiformis. AB - gamma-Aminobutyric acid (GABA) and 5-hydroxytryptamine (5-HT) have been shown to increase the reproduction of the Brachionus plicatilis (NH3L strain). In the present study, the endogenous presence of GABA and 5-HT in the rotifers B. plicatilis (NH3L and Kamiura strains) and Brachionus rotundiformis (Langkawi strain) were confirmed by dot blot immunoassay and high-performance liquid chromatography (HPLC). HPLC showed that GABA and 5-HT concentrations in the three rotifer strains range from 71 to 188 pmol/mg and from 12 to 64 pmol/mg, respectively. A total of 33 amino acids were also detected in B. plicatilis and B. rotundiformis, with glutamic acid, serine, glycine, taurine, threonine, alanine, arginine, proline, valine and isoleucine in high concentrations relative to other amino acids. PMID- 11118941 TI - The influence of body mass, climate, and distribution on the energetics of South Pacific pigeons. AB - Rate of metabolism and temperature regulation were studied in 16 species of South Pacific pigeons, which constitute 13 fruit-eaters, 1 seed-eater, 1 fruit/nut eater, and 1 fruit/leaf-eater; 14 tropical and two temperate species; and ten mainland and six intermediate- or small-island species. The data presented here and those from 11 additional columbids indicate in an analysis of covariance that log(10) basal rate of metabolism is correlated with log(10) body mass (P< or =0.0001), distribution (P=0.0023), and climate (P=0.0016). These factors account for 94.3% of the variation in log(10) basal rate of metabolism. In this analysis the lowest basal rates, corrected for body mass, are found in tropical pigeons living on small oceanic islands, whereas the highest basal rates are found in temperate species living on continents. The reduction of basal rate in large columbids facilitates their long-term persistence on small islands characterized by a limited resource base and unstable weather. Some small-island specialists have a smaller mass than their continental relatives, which further reduces resource requirements. The question whether a reduction in basal rate occurs in small columbids on small islands is unresolved. Log(10) minimal thermal conductance is apparently correlated only with log(10) body mass (P< or =0.0001); r(2)=89.4%. The mean nocturnal body temperature of columbids is 39.7 degrees C. PMID- 11118942 TI - Extracellular norepinephrine in the medial hypothalamus increases during feeding in chicks: a microdialysis study. AB - Norepinephrinergic function in the medial hypothalamus is important for the regulation of feeding behavior in chicks as well as in rats. This study was conducted to clarify the variation of extracellular norepinephrine (NE) in the medial hypothalamus, including the paraventricular nucleus (PVN) and the ventromedial hypothalamic nucleus (VMN), during feeding behavior of layer-type chicks. To measure extracellular NE and 4-hydroxy-3-methoxyphenylglycol (MHPG), a major metabolite of NE, we used microdialysis and high-pressure liquid chromatography (HPLC) with electrochemical detection. After the collection of baseline samples, food-deprived animals were allowed access to the food for 3 h. Extracellular NE significantly increased during the first hour of access to food, and then returned to baseline levels. MHPG also increased during the feeding, but its increase continued throughout the remainder of the experiment. This study suggests that the variation of NE in the medial hypothalamus may be involved in the control of feeding in layer-type chicks. PMID- 11118943 TI - Detection of plasmalogen from plasma low density lipoprotein and high density lipoprotein in carp, Cyprinus carpio, and rainbow trout, Oncorhynchus mykiss. AB - The study revealed the presence of plasmalogens in the low density lipoprotein (LDL) and high density lipoprotein (HDL) of the fish. The composition of the plasmalogen in the carp plasma LDL phospholipids was 0.94 and 0.23% in the HDL; the LDL phospholipids in the rainbow trout were 0.44% and the HDL was 0.18%. Aldehydes from the plasmalogen were derivatized with dansylhydrazides and separated by high performance liquid chromatography (HPLC). Their presence was detected using a fluorescence detector. Hexadecanal (C16: 0), octadecanal (C18: 0) and octadecenal (C18: 1) were determined to be the major components in the carp and rainbow trout. PMID- 11118944 TI - Haemolysis of human and sheep red blood cells in glycerol media: the effect of pH and the role of band 3. AB - Haemolysis of red blood cells (RBC) in glycerol media may be measured spectrophotometrically. The haemolytic process in a rapid phase obeys a first order rate law. The rate constant expresses the rate of haemolysis. To gain a better understanding of the mechanism of haemolysis in glycerol media, the effects of pH and band 3 inhibitors on the rate of haemolysis in human and sheep RBC were observed. Over the pH range used (pH 5.8-10.0), the rate of haemolysis decreased with increase in pH in sheep RBC. By contrast, the rate of haemolysis increased from pH 5.8 to 6.4 and decreased above pH 6.4 in human RBC. The different effects of pH on the rate of haemolysis are due to inhibition of glycerol permeability by H(+) in human RBC but not in sheep RBC. This is supported by the different effects of temperature and Cu(2+) on the rate of haemolysis in human and sheep RBC. We did not observe complete inhibition of haemolysis by the classical band 3 inhibitor, 4, 4'-diisothiocyanatostilbene-2,2' disulfonic acid (DIDS). Another band 3 inhibitor 4,4'-dinitrostilbene-2,2' disulfonic acid (DNDS) showed only weak inhibition. Phenylgloxal (PG), another band 3 inhibitor, had no effect whatsoever on the rate of haemolysis. These results indicate that the anion pathway of band 3 is not the preferred route of transport of glycerol in mammalian RBC. PMID- 11118945 TI - Dopaminergic innervation of the rainbow trout pituitary and stimulatory effect of dopamine on growth hormone secretion in vitro. AB - To elucidate which factors regulate growth hormone (GH) secretion in rainbow trout, dopaminergic innervation of the rainbow trout pituitary along with the action of dopamine in vitro, were studied. Brains with attached pituitaries were double-labeled for putative dopaminergic neuronal fibers and somatotropes, using fluorescence immunohistochemistry. A direct dopaminergic innervation to the proximal pars distalis (PPD) with dopaminergic fibers terminating adjacent to somatotropes was demonstrated. Growth hormone secretion from whole pituitaries was measured in perifusate using a homologous GH-RIA. Dopamine (DA; 10(-7)-2x10( 6) g ml(-1)) increased basal GH secretion, with the GH secretion normalizing again after the DA exposure was halted. When pituitaries were pre-treated with somatostatin-14 (SRIF-14; 10(-12)-10(-9) g ml(-1)), before being exposed to different doses of DA, there was an inhibition of GH secretion which was not reversed after treatment of SRIF-14 was halted, unless DA was added. It is concluded that dopamine can function as a GH secretagogue in the rainbow trout pituitary gland. PMID- 11118962 TI - Q & A about this issue PMID- 11118963 TI - The Cochrane collaboration--why is it important to the future of clinical pharmacology? PMID- 11118964 TI - Could conditional release of new drugs provide the information required to study drug effectiveness? - A discussion paper. AB - Conditional release is the approval of a new drug onto the market, subject to specific conditions relating to effectiveness and safety that, if achieved, will lead to full approval. Conditional approval of a new drug, during which time it is used in normal clinical practice, should allow the collection of data on effectiveness and safety to provide a genuine cost effectiveness evaluation. Proposals put forward in 1977 in the United Kingdom for approving a drug on a conditional basis while monitoring for adverse drug reactions are examined, and issues that would affect a present day conditional release scheme are identified. These issues are: who would do the evaluation and who would pay for it; how would patients be identified and registered; would all new drugs be monitored and for how long; what data would be reported and evaluated; and who would do the reporting? How a conditional release scheme would work in Canada in light of these questions is considered and a method based on pharmacists registering patients and on physicians and/or patients reporting data to an independent organization funded by governments and the pharmaceutical industry is outlined. Under certain conditions, conditional release would provide the information to allow true cost effectiveness and safety assessments instead of the current inadequate predictions based on efficacy and safety data from clinical trials. It is important that academics and drug approval and monitoring agencies work together to develop active systems to improve the postapproval evaluation of effectiveness, safety and cost effectiveness of new drugs in Canada. PMID- 11118965 TI - Is there an indication for the use of barbiturate-containing analgesic agents in the treatment of pain? Guidelines for their safe use and withdrawal management. Canadian Pharmacists Association. AB - OBJECTIVE: To provide medical and pharmaceutical practitioners with information on the effectiveness, safety and risks associated with barbiturate-containing analgesic (BCA) agents and an approach to management of withdrawal from BCAs. METHODS: The benefits of using BCAs in the treatment of pain (compared with using non-BCAs) were weighed against potential risks. The procedures for discontinuation of BCAs in patients with pain were considered, and evidence showing that BCAs constitute a public health risk was reviewed. The evidence serving as the basis for the clinical guidelines was compiled from a number of sources, including key papers in the field, published and unpublished papers, and reports by Addiction Research Foundation researchers, a MEDLINE search from 1967 to November 1996, and communication with Canadian manufacturers of BCAs. RESULTS: There is no evidence that there is a clinically important enhancement of analgesic efficacy of BCAs due to the barbiturate constituent. No epidemiological studies on the relative frequency of abuse and dependence, or studies that have analyzed combination product use from a public health and social benefit to risk perspective, have been published to clarify the issues about the nature, extent and seriousness of these problems. RECOMMENDATIONS: No evidence exists to show a clinically important enhancement of analgesic efficacy of BCAs due to the barbiturate constituents. Because BCAs do not have a therapeutic advantage, there is no clinical reason to choose such a combination product when a simpler and often less expensive analgesic formulation (eg, acetaminophen, acetylsalicylic acid, nonsteroidal anti-inflammatory drug or narcotic) or a more specific anti migraine drug (eg, dihydroergotamine or sumatriptan) is available. BCAs should be avoided in elderly people and should not be used in children. Extrapolation from published reports on abuse and withdrawal syndrome with these drugs suggests that BCAs have the potential to produce drug dependence and addictive behaviour, especially with regular use. In BCA overdose, the barbiturate component is only one of the clinically significant contributors to any morbidity, but its presence can complicate the management of additive or synergistic toxicities. Therefore, there is no reason to choose a combination product when a simpler product may be a safer alternative by minimizing the potential for addiction and the occurrence of additive side effects or toxicities. It is further recommended that prescribers re-evaluate treatment for patients using BCAs. Recommendations for withdrawal are provided, based on estimated consumption. PMID- 11118966 TI - Antidote availability in Quebec hospital pharmacies: impact of N-acetylcysteine and naloxone consumption. AB - OBJECTIVES: To study the availability of 13 specific antidotes in hospitals and correlate the availability of those antidotes to the number of poisonings seen in hospitals using N-acetylcysteine and naloxone consumption as a surrogate. METHODS: Pharmacy directors of hospitals with an emergency department were surveyed for number of adequately stocked antidotes (N-acetylcysteine, ethanol, cyanide antidote kit or hydroxycobalamine, deferoxamine, digoxin-immune FAB, dimercaprol, flumazenil, glucagon, methylene blue, naloxone, physostigmine, pralidoxime and pyridoxine). RESULTS: Data were obtained from 96 of 112 (86%) of the pharmacies surveyed. Number of adequately stocked antidotes per hospital ranged from zero to nine of 13. There was a correlation between all hospital characteristics evaluated and the number of adequately stocked antidotes (P<0.05). Correlations between the number of adequately stocked antidotes and the amount of N-acetylcysteine and naloxone consumed were significant (rs=0.58, P<0.001; r(s)=0.53, P<0.001). The amount of N-acetylcysteine consumed, the number of annual visits to the emergency department and the number of hours of pharmacy coverage on weekends independently predicted the presence of adequately stocked antidotes. CONCLUSIONS: Larger hospitals are more likely to have adequate stocks of antidotes. Adequate stocking of antidotes is significantly correlated with the amount of N-acetyl- cysteine and naloxone consumed. This suggests that hospitals more likely to see serious acetaminophen and opiate poisonings are more likely to maintain adequate stocks of antidotes. PMID- 11118967 TI - Longitudinal change in the treatment of nausea and vomiting of pregnancy in Ontario. AB - BACKGROUND: Health problems associated with untreated nausea and vomiting of pregnancy (NVP) include maternal weight loss, dehydration, and electrolyte and acid-base disturbances. Negative social consequences include the deterioration of domestic, social and occupational function of the affected women. In 1994 it was documented that most physicians in Ontario tended not to treat women with NVP pharmacologically. PURPOSE: To investigate the longitudinal change in therapeutic practice of physicians, with respect to the treatment of NVP. SUBJECTS AND METHODS: A questionnaire was distributed to a randomly selected sample of physicians that included community-based family physicians, hospital-based family physicians, obstetricians and physicians who called the Motherisk Program for information. The participants were questioned about their choices in the pharmacological treatment of NVP. The data from the survey were compared with those from a previously published survey conducted in 1994. RESULTS: In 1999, 90.2% of physicians surveyed reported to have used pharmacological means to treat NVP. In 1999, 95% of the physicians surveyed reported to have prescribed doxylamine succinate-pyridoxine hydrochloric acid (Diclectin), and 11% prescribed dimenhydrinate (Gravol) to treat NVP. These results are significantly different from those found in 1994 (90% prescribed Gravol as the first choice). CONCLUSIONS: In 1999, temporally related to various educational efforts, physicians offered treatment for NVP more readily, including the drug recommended by the regulatory agency. These changes may explain in part the recently documented decrease in hospitalizations due to NVP in Canada. PMID- 11118968 TI - (NH4)3 AB - The title compound, triammonium cis-diaqua-cis-dioxo-trans-disulfatovanadate 1.5 hydrate, was obtained by oxidizing V(IV) to V(V) in a 2 M sulfuric acid solution of vanadyl sulfate and adding ammonium sulfate. Here, the V atom is sandwiched by two sulfate groups by corner-sharing to form a discrete [VO(2)(SO(4))(2)(OH(2))(2)](3-) anion. The water molecules occupy cis positions in the equatorial plane of the vanadium octahedron. PMID- 11118969 TI - Lithium potassium borate and lithium rubidium borate: new non-linear optical crystals AB - The title compounds, LiKB(4)O(7) and LiRbB(4)O(7), are newly developed non-linear optical crystals containing two kinds of anionic groups, namely (B(3)O(8))(7-) and (B(5)O(10))(5-). The (B(3)O(8))(7-) groups form infinite spiral chains parallel to the [100] direction, which are interconnected by sharing O atoms with (B(5)O(10))(5-) groups. PMID- 11118970 TI - Bis(mu-pyridine-2,6-carboxylato-O,N, O':O)bis[triaquamanganese(II)]-pyridine-2,6 dicarboxylic acid (1/2). AB - In the structure of the title compound, [Mn(2)(C(7)H(3)NO(4))(2)(H(2)O)(6)].2C(7)H(5)NO(4), a centrosymmetric dinuclear complex, hexaaaquabis(pyridine-2, 6-dicarboxylato)dimanganese(II) and free pyridine-2,6-dicarboxylic acid are present in a 1:2 ratio. In the complex, each Mn(2+) ion is coordinated by three O atoms and one N atom from the pyridine-2, 6 dicarboxylate ligands and by three water O atoms, resulting in a distorted pentagonal bipyramidal coordination. Within the centrosymmetric dinuclear complex, two Mn(2+) ions are bridged by two carboxylate O atoms. The crystal structure is stabilized by hydrogen bonds involving all the H atoms of the water ligands. PMID- 11118971 TI - catena-Poly[[diaqualithium(I)]-mu-[9H-purine-2,6(1H, 3H)-dionato-O2:N7]]. AB - In the title compound, [Li(C(5)H(3)N(4)O(2))(H(2)O)(2)](n), the coordinate geometry about the Li(+) ion is distorted tetrahedral and the Li(+) ion is bonded to N and O atoms of adjacent ligand molecules forming an infinite polymeric chain with Li-O and Li-N bond lengths of 1.901 (5) and 2.043 (6) A, respectively. Tetrahedral coordination at the Li(+) ion is completed by two cis water molecules [Li-O 1.985 (6) and 1.946 (6) A]. The crystal structure is stabilized both by the polymeric structure and by a hydrogen-bond network involving N-H.O, O-H.O and O H.N hydrogen bonds. PMID- 11118972 TI - A cyanide-bridged bimetallic complex, AB - The crystal structure of the bimetallic cyanide-bridged title complex, triaqua 1kappa(3)O-&mgr;-cyano-1:2kappa(2)N:C-pentacyano-2kappa(5)C-t etrakis(N, N dimethylformamide)-1kappa(4)O-chromium(III)praseodymium(III) monohydrate, was obtained by single-crystal X-ray diffraction. The central praseodymium(III) ion is eight-coordinate, arranged in a square antiprism, while the chromium(III) ion is six-coordinate, oriented octahedrally. Molecules in the crystal lattice are held together by a network of hydrogen bonds. PMID- 11118973 TI - [1,1,2,2-] AB - The title compound, 1,1,2,2-tetracarbonyl-1,2-&mgr;-carbonyl-4, 11 dimethylsulfido-closo-1,2-dicobaltadodecaborane, [Co(2)(C(4)H(20)B(10)S(2))(CO)(5)], has a closo 12-vertex 1, 2-Co(2)B(10)H(8) structure with SMe(2) ligands at the exo-4- and 11-positions. The cluster displays close structural similarities to the SEt(2) analogue. PMID- 11118974 TI - Bis2,6-bis AB - The title compound, [Zn(C(29)H(29)N(5))(2)](ClO(4))(2).2CH(3)NO(2), contains a Zn(II) ion showing only small deviations from local D(2d) symmetry. The lower rhombicity exhibited by this complex compared with that of its Cu(II) congener suggests that the highly rhombic stereochemistry exhibited by the latter is largely imposed by the stereoelectronic preferences of the Cu(II) ion. PMID- 11118975 TI - Tetraaquabis(3,5-dicarboxybenzoato-O)cobalt(II) AB - The hydrothermal reaction of cobalt(II) chloride with trimesate (3, 5 dicarboxybenzoate) ions in aqueous solution gives the novel title complex, [Co(C(9)H(5)O(6))(2)(H(2)O)(4)]. The Co(II) ion lies on an inversion centre and is octahedrally coordinated to two trimesate anions and four water molecules. Hydrogen bonds ensure the three-dimensional architecture of the structure. PMID- 11118976 TI - A donor-acceptor compound composed of a dinuclear zinc(II) complex of a robson macrocycle with 8-methylquinoline AB - A donor-acceptor compound, diaqua-1kappaO,2kappaO-[&mgr;-11, 23-dimethyl 3,7,15,19-tetraazatricyclo[19.3.1.1(9,13)]hexacosa-1(25), 2,7,9,11,13(26),14,19,21,23-decaene-25,26-diolato-1kappa(4)N(3),N(7), O(25),O(26):2kappa(4)N(15),N(19),O(25),O(26)]dizinc(II) diperchlorate bis(8 methylquinoline) ethanol disolvate, [Zn(2)(C(24)H(26)N(4)O(2))(H(2)O)(2)](ClO(4))(2).2C(10)H(9)N. 2C(2)H(6)O, obtained by the reaction of a dinuclear zinc(II) complex of a Robson macrocycle (acceptor) and 8-methylquinoline (donor), lies about an inversion centre and the coordination about the unique Zn atom is a distorted square pyramid. The fifth coordination site is occupied by the water molecule, Zn-O = 2.016 (2) A, and the average macrocyclic Zn-O and Zn-N distances are 2.059 (6) and 2.059 (3) A, respectively. PMID- 11118977 TI - Hexakis(1H-imidazole-kappaN3)nickel(II) bis[O,O'-diisopropyl dithiophosphate(1 )]. AB - In the title complex, [Ni(Im)(6)]((i)Pr-dtp)(2) or [Ni(C(3)H(4)N(2))(6)](C(6)H(14)O(2)PS(2))(2), the coordination around the Ni atom, located on an inversion centre, is octhahedral with all positions being occupied by tertiary N atoms of the imidazole moieties. Hydrogen bonds link the anions and cations into a two-dimensional network in the bc plane. PMID- 11118978 TI - Solvent-free AB - The title compound, [Au(2)Cl(2)Fe(C(17)H(14)P)(2)], (I), contains the expected linear gold centres. The ferrocene moiety acts as a P, P'-bridging ligand, wherein the Fe atom lies on an inversion centre. The P-Au-Cl angle is 177.56 (8) degrees and bond distances Au-P and Au-Cl are 2.2261 (18) and 2.2781 (18) A, respectively. The structure is almost identical to that of the metal complex in (I).2CH(2)Cl(2) [Canales, Gimeno, Jones, Laguna & Sarroca (1997). Inorg. Chem. 36, 5206-5211], but differs considerably from that in 3(I).2CHCl(3) [Hill, Girard, McCabe, Johnson, Stupik, Zhang, Reiff & Eggleston (1989). Inorg. Chem. 28, 3529-3533], in that in the latter, the two independent molecules are linked by a short Au.Au contact. PMID- 11118979 TI - cis-Tetracarbonylbis(tricyclohexylphosphine)molybdenum(0) and pentacarbonyl(tricyclohexylphosphine)molybdenum(0) AB - In the present redetermination of the complex cis tetracarbonylbis(tricyclohexylphosphine)molybdenum(0), (I), [Mo(C(18)H(33)P)(2)(CO)(4)] or cis-eta(1)-[P(C(6)H(11))(3)](2)Mo(CO)(4), the Mo atom has a distorted octahedral geometry with a large P-Mo-P angle of 104.8 (1) degrees. A strong trans influence on the carbonyls in (I) is seen in a shortening of the Mo-C and a lengthening of the C-O distances opposite the phosphines compared with those that are cis. This influence is greatly diminished in the complex pentacarbonyl(tricyclohexylphosphine)molybdenum(0), (II), [Mo(C(18)H(33)P)(CO)(5)] or eta(1)-[P(C(6)H(11))(3)]Mo(CO)(5), the core of which has a slightly distorted C(4v) geometry. PMID- 11118980 TI - Di-&mgr;-hydroxo-bis(diisopropylgallium)-1,4,8,11-tetramethyl-1,4,8, 11 tetraazacyclotetradecane (1/1) AB - The hydrolysis product [Ga(2)(C(3)H(7))(4)(OH)(2)].C(14)H(32)N(4), derived from the tetrakis(triisopropylgallium)-1,4,8, 11-tetramethyl-1,4,8,11 tetraazacyclotetradecane (1/1) adduct, consists of a centrosymmetric [(i)Pr(2)Ga(&mgr;-OH)](2) unit hydrogen bonded through the hydroxyl group to a nitrogen on an adjacent centrosymmetric 1,4,8,11-tetramethyl-1,4,8, 11 tetraazacyclotetradecane molecule, resulting in the generation of a molecular chain through the crystal. PMID- 11118981 TI - Cyclometallated compounds. XV. A tetranuclear acetate-bridged cyclopalladated molecular box AB - The title complex, tetra-&mgr;-acetato-O:O'-bis[&mgr;-1, 4-bis(2 pyridyloxy)phenylene-N,C(2):N',C(6)]dipalladium(II) bis(trichloromethane) dihydrate, [Pd(4)(C(16)H(10)N(2)O(2))(2)(C(2)H(3)O(2))(4)].2CHCl(3).2H(2)O, the product of the reaction of 1,4-bis(2-pyridyloxy)benzene with palladium acetate, is shown to be a tetranuclear, rather than a polymeric, species. It crystallizes about a centre of inversion and has two doubly cyclopalladated ligands bridged by four acetate groups. The cyclopalladated ligand is far from planar in the complex and has the central benzene rings pi-stacked. The chelate rings exist in shallow boat conformations. PMID- 11118982 TI - N-(3-Chlorophenyl)-alpha-phenylnitrone: association with benzoic acid through hydrogen bonding AB - The identity of the title complex, C(13)H(10)ClNO.C(7)H(6)O(2), is confirmed to be a hydrogen-bonded adduct of benzoic acid and N-(3-chlorophenyl)-alpha phenylnitrone. The two aromatic rings in the nitrone are trans about the C=N bond. PMID- 11118983 TI - 1,2-Diphenyl-2-(phenyldiazenyl)-2-tosylethanone, an example of a fungicide containing a sulfonyl group. AB - The molecule of the title compound, C(27)H(22)N(2)O(3)S, adopts an irregular propeller shape with the tetrahedral C1 atom pivotal. The alpha-azophenyl and alpha-phenyl moieties are approximately coplanar. Electrostatic attraction of the oppositely charged atoms generates several short intramolecular contacts involving the sulfonyl, azo and carbonyl groups. Characteristic bond-length distribution of the central part of the molecule indicates that the Coulombic charge transfer is supplemented by hyperconjugation involving donation of electron density from the azo moiety towards the sulfonyl and carbonyl groups. PMID- 11118984 TI - Racemic dipeptide glycyl-DL-leucine at 120 K. AB - The structure of glycyl-DL-leucine, C(8)H(16)N(2)O(3), has been determined at 120 K by single-crystal X-ray diffraction. In addition to three N-H.O-type hydrogen bonds of the positively charged RNH(3)(+) group of the zwitterionic molecule, an intermolecular N-H. O contact exists between the peptide bond and the carboxylate group. Four hydrogen-bond cycles were identified, giving a complex pattern. PMID- 11118985 TI - 13,13a-Dihydro-13-methoxy-9-methyl-1-benzopyrano[4,3-i]dinaphtho[2, 1-c;1',2'-f] 2,8-dioxabicyclo[3.3.1]nonane. AB - The crystal structure of the title compound, C(32)H(24)O(4), contains three fused dihydropyran rings (A, B and C); ring A is fused with a benzene ring while the other two rings, B and C, are fused with naphthalene rings. Ring A adopts a half chair conformation with an equatorial methoxy group, whereas ring B assumes a distorted half-chair conformation, the A/B ring junction being trans. Ring C adopts a distorted half-boat conformation and is nearly orthogonal to ring B. Ring C is inclined to the best plane of ring A at an angle of 112.1 (1) degrees. PMID- 11118986 TI - A silaproline-containing dipeptide. AB - The silaproline-containing dipeptide N-(3, 3-dimethyl-1-pivaloyl-1-aza-3-sila-5 cyclopentylcarbonyl)-L- alanine isopropylamide, C(17)H(33)N(3)O(3)Si, has two independent molecules in the asymmetric unit and each adopts a beta-II folded conformation, where the amide on the terminal C interacts intramolecularly with the pivaloyl O atom. The five-membered silaproline ring is C(beta)-puckered, an infrequent conformation for the homologous proline ring. PMID- 11118987 TI - 2-Hydroxyphenyl 2-methylpyrazolo[1,5-a]pyrimidin-6-yl ketone. AB - The molecules of the title compound, C(14)H(11)N(3)O(2), form a three-dimensional soft hydrogen-bonded network involving C-H...N hydrogen bonds. PMID- 11118988 TI - 5,5-Dimethyl-3-(5-methyl-1H-pyrazol-3-ylamino)cyclohex-3-en-1-one AB - The molecules of the title compound, C(12)H(17)N(3)O, are linked by two N-H.O hydrogen bonds to form a three-dimensional network. The N. O distances are 2.804 (3) and 2.766 (3) A, both involving a common acceptor O atom. PMID- 11118989 TI - Weak and strong hydrogen-bonding patterns in the structure of 2, 4-bis(2 hydroxybenzoyl)-2,3-dihydro-1H-pyrido AB - The title compound, C(25)H(19)NO(4)S, (IV), was produced by a cyclocondensation reaction similar to that which had previously produced an unexpected product, thus giving a novel route for such reactions. The structure of (IV) contains two S(6) motifs formed by strong intramolecular O-H.O hydrogen bonds. Weak C-H.O hydrogen bonds form primary C(11), R(1)(2)(7) and R(2)(2)(8) motifs which combine to form a complex three-dimensional network. PMID- 11118990 TI - Hydrogen-bonded dimers in N-(2-nitrophenylthio)saccharin AB - In the title compound, 2-(2-nitrophenylthio)-1, 2-benzothiazol-3(2H)-one 1,1 dioxide, 2-O(2)NC(6)H(4)S(C(7)H(4)NO(3)S) or C(13)H(8)N(2)O(5)S(2), the planes of the saccharin and nitrophenylthiolate portions are almost orthogonal. The molecules are linked by C-H.O=S hydrogen bonds [C.O 3.308 (3) A, H.O 2.44 A and C H.O 155 degrees ] into cyclic centrosymmetric R(2)(2)(16) dimers, reinforced by aromatic pi.pi stacking interactions between the nitrated aryl rings. PMID- 11118991 TI - Conformational preferences and supramolecular aggregation in 2 nitrophenylthiolates: 2-nitrophenyl-beta-D-thiogalactopyranoside AB - In the title compound, C(12)H(15)NO(7)S, the molecular conformation shows a concerted disrotatory twist of the nitro group and the galactose fragment out of the plane of the aryl ring. The molecules are linked by O-H.O hydrogen bonds [O.O range 2.725 (2)-3.024 (2) A and O-H.O range 155-175 degrees ] to form a three dimensional framework. PMID- 11118992 TI - The surprising sp rotameric structure of 9-methyl-9-pivaloylfluorene AB - Methylation of 9-lithiated ap-9-pivaloylfluorene, (I), as well as pivaloylation of 9-lithiated 9-methylfluorene provided rotationally stable sp-9-methyl-9 pivaloylfluorene, (III), C(19)H(20)O, which lies about a crystallographic mirror plane. Fluorene (I) exists exclusively in the ap configuration in solution (NMR) as well as in the crystalline state, reflecting the unfavorable interaction between the tert-butyl and fluorene-ring pi electrons in the sp configuration. The existence of (III) exclusively in the sp configuration indicates that, in this case, the interaction between the tert-butyl group and the fluorene-ring pi electrons provides relatively more thermodynamic stability than the steric interaction between the tert-butyl and 9-methyl groups (ap configuration). PMID- 11118993 TI - cis,cis,cis-tetramethyl 1,2,4,5-cyclohexanetetracarboxylate AB - The title compound, C(14)H(20)O(8), was synthesized from the hydrogenation of tetramethyl 1,4-cyclohexadiene-1,2,4, 5-tetracarboxylate with a catalytic amount of palladium/carbon. All four carbonyl moieties of the methyl ester groups are on the same face of the chair-conformed ring. The substantial ring distortion associated with the 1,3-diaxial methoxycarbonyl substituents is reflected in the large difference between bond angles as well as torsion angles, respectively, that in undistorted cyclohexanes would be approximately the same. PMID- 11118994 TI - 2,2'-Bipyridinium bis(perchlorate) AB - The title compound, [H(2)bipy](ClO(4))(2) or C(10)H(10)N(2)(2+). 2ClO(4)(-), was obtained at the interface between an organic (2, 2'-bipyridine in methanol) and an aqueous phase (perchloric acid in water). The compound crystallizes in space group P-1 and comprises discrete diprotonated trans-bipyridinium cations, [H(2)bipy](2+), and ClO(4)(-) anions. The cations and anions are connected through N-H.O and C-H.O hydrogen bonds [distances N.O 2.817 (4) and 2.852 (4) A, and C.O 3.225 (6)-3.412 (5)A]. The C-C bond distance between the two rings is 1.452 (5) A. The bipyridinium cation has a trans conformation and the N-C-C-N torsion angle is 152.0 (3) degrees. PMID- 11118995 TI - alpha-Onocerin chloroform hemisolvate. AB - The triterpenoid natural product alpha-onocerin [8, 14-secogammacera-8(26),14(27) diene-3,21-diol], determined here as the chloroform hemisolvate, C(30)H(50)O(2).0.5CHCl(3), consists of two independent symmetric trans-decalin C(15) building blocks. Hydrogen bonds between the hydroxyl groups form an infinite two-dimensional network perpendicular to the c axis. PMID- 11118996 TI - 2-Amino-5-chloro-1,3-benzoxazol-3-ium 2-(3,4-dichlorophenoxy)acetate. AB - The 1:1 organic salt of the title compound, C(7)H(6)ClN(2)O(+). C(8)H(5)Cl(2)O(3)(-) or [(2-ABOX)(3,4-D)], comprises the two constituent molecules associated by an R(2)(2)(8) graph-set interaction through the carboxylate group of 3,4-D across the protonated N/N sites of 2-ABOX [N.O 2.546 (3) and 2.795 (3) A]. Cation/anion pairs associate across an inversion centre forming discrete tetramers via an additional three-centre hydrogen-bonding association from the latter N amino proton to a phenoxy O atom [N.O 3.176 (3) A] and a carboxylate O atom [N.O 2.841 (3) A]. This formation differs from the polymeric hydrogen-bonded chains previously observed for adduct structures of 2 ABOX with carboxylic acids. PMID- 11118997 TI - (1R,2s,3S,6R,9S)-5,5,10,10-tetrachloro-2-methyltricyclo AB - The title compound, C(12)H(10)Cl(4)O(2), has a pseudoasymmetric centre at the methyl-substituted carbon and, in the solid state, a boat-like conformation. PMID- 11118998 TI - Ethyl 2-[N-(tert-butyloxycarbonyl)-L-alanylamino]-4-methyl-1, 3-thiazole-5 carboxylate reveals a trans orientation of the preceding amide N--H with respect to the thiazole-ring sulfur. AB - The title molecule, C(15)H(23)N(3)O(5)S, was prepared as a synthetic precursor to 4-methylthiazole-based DNA minor groove binders which would bear chiral amino acids in the sequence. The crystallographic evidence presented herein shows that the aromatic amide NH group preceding the thiazole ring points away from the direction of sulfur. The molecule is biplanar, with the dihedral angle between the N-terminus peptide moiety and the thiazole-containing plane being 49.7 (5) degrees, with a bend at the Calpha carbon. PMID- 11118999 TI - Two N-ortho-nitrobenzenesulfonyl alpha,alpha-disubstituted amino acid adducts. AB - The carboxy group of 2-methyl-N-[(2-nitrophenyl)sulfonyl]alanine, C(10)H(12)N(2)O(6)S, forms centrosymmetric hydrogen-bonded dimers with an O.O distance of 2.629 (2) A and an intramolecular N-H. O(nitro) hydrogen bond N.O distance of 2.823 (2) A. 1-[(2-Nitrophenyl)sulfonylamino]cyclohexanecarboxylic acid, C(13)H(16)N(2)O(6)S, has Z' = 2 and forms similar interactions. PMID- 11119000 TI - trans-4- AB - The crystal structure of the dipolar chromophoric title compound, C(20)H(20)N(3)(+).PF(6)(-), is described. The phenylene and pyridyl rings are almost coplanar [dihedral angle 7.5 (2) degrees ], but the phenyl substituent forms a dihedral angle of 56.6 (1) degrees with the pyridyl ring. The compound crystallizes in the non-centrosymmetric space group Cc and is a likely candidate for the display of quadratic non-linear optical effects. PMID- 11119001 TI - 4-Methyl-N- AB - The title compound, C(20)H(19)NO(2)S(2), is formed by a palladium-copper catalyzed reaction between 4-methyl-N-[2-(prop-2 ynylsulfanyl)phenyl]benzenesulfonamide and p-iodotoluene. The molecules contain three essentially planar parts, namely an aminothiophenol moiety (A), a toluenesulfone moiety excluding the oxo ligands (B) and a tolyl group (C), approximately orthogonal to each other; the dihedral angles A/B, A/C and B/C are 111.6 (1), 89.3 (1) and 101.4 (1) degrees, respectively. Intermolecular N-H.O hydrogen bonds link the molecules into infinite one-dimensional chains. PMID- 11119002 TI - 5-Methyl-2'-deoxy-4'-thio-alpha-uridine (R)-S-oxide. AB - The pyrimidine ring of the title compound, C(10)H(14)N(2)O(5)S, is planar to within 0.024 (1) A and makes an angle of 75.46 (10) degrees with the mean plane of the thiosugar ring. In terms of standard nucleoside nomenclature, this ring has the C3'-endo conformation. The O5'-C5'-C4'-C3' torsion angle is 166.5 (3) degrees and the glycosidic torsion angle S4'-C1'-N1-C2 is -52.1 (2) degrees (syn). PMID- 11119003 TI - A spiro-linked pyrene-naphthoquinone. AB - The title molecule, 2'-pyrenylspiro[2, 3-dihydro-1H-cyclopenta[b]naphthalene-2,5' 1',3'-dioxane]-4,9-dione, C(32)H(22)O(4), contains an electron-donating pyrene group spiro-linked to an electron-accepting naphthoquinone. The molecules are V shaped in profile and stack to form columns along b with alternating, approximately coplanar, pyrene and naphthoquinone fragments. Intermolecular contacts within a column are consistent with some degree of pi contact and possible long-range delocalization. Individual columns form a herringbone pattern when the crystal is viewed along b. PMID- 11119004 TI - An NH3+...phenyl interaction in L-phenylalanyl-L-valine. AB - One of the amino H atoms of L-phenylalanyl-L-valine, C(14)H(20)N(2)O(3), participates in a rare secondary interaction in being accepted by the aromatic ring of the phenylalanine side chain. The phenyl group is also a donor in a weak hydrogen bond to the peptide carbonyl group. PMID- 11119005 TI - 2,3-Bis(4-nitrobenzylthio)maleonitrile AB - The maleonitrile moiety of the title compound, (2Z)-2, 3-bis[(4 nitrobenzyl)sulfanyl]but-2-enedinitrile, C(18)H(12)N(4)O(4)S(2), is almost planar. The two benzene rings are nearly parallel to each other and perpendicular to the maleonitrile plane. Intermolecular S.S and pi-pi interactions are observed in the crystal structure. PMID- 11119006 TI - 2,4-Dibromo-3-methoxy-6-nitrobenzaldeyde: effect of substituents on ring geometry AB - In the title compound, C(8)H(5)Br(2)NO(4), the endocyclic angles of the ring deviate significantly from the ideal value of 120 degrees. The substituents deviate from the plane of the ring, with large twist angles for the aldehyde, nitro and methoxy groups. The geometry of the molecule in the crystal is compared with that of the isolated molecule, as given by a self-consistent field molecular orbital Hartree-Fock calculation. Only weak hydrogen bonds of the C-H.Br and C H.O types are present in the crystal structure. PMID- 11119007 TI - 7-Nitro-1H-indazole, an inhibitor of nitric oxide synthase. AB - The crystal structure of 7-nitro-1H-indazole, C(7)H(5)N(3)O(2), an inhibitor of nitric oxide synthase, shows the existence of an intramolecular hydrogen bond between an O atom of the nitro group and the NH group of the indazole ring. The crystal packing consists of intermolecular hydrogen bonding and indazole.indazole interactions. PMID- 11119008 TI - The 1:1 adduct of Kemp's triacid with 2-aminopyridine AB - The crystal structure determination of the molecular proton-transfer adduct of Kemp's triacid (cis-cis-1,3,5-trimethylcyclohexane-1,3, 5-tricarboxylic acid, KTA) with 2-aminopyridine (2-APY), namely 2-aminopyridinium 3,5-dicarboxy-1,3, 5 trimethylcyclohexanecarboxylate, 2-APY(+).KTA(-) or C(5)H(7)N(2)(+).C(12)H(17)O(6)(-), has revealed a centrosymmetric hydrogen-bonded cyclic KTA homodimer repeating unit [O.O 2.524 (4) A] linked into a polymer structure through the pyridinium and amino groups of the 2-APY molecule [O.N 2.736 (4), 2.989 (4) and 2.999 (4) A]. PMID- 11119009 TI - (+)-Gibberellin C: hydrogen-bonding pattern of the monohydrate of a non-racemic pentacyclic diterpenoid. AB - In the monohydrate of the title compound, (+)-2beta, 4aalpha-dihydroxy-1,7 dimethyl-8-oxo-4bbeta,7alpha- gibbane-1alpha, 10beta-dicarboxylic acid-1,4a lactone, C(19)H(24)O(6).H(2)O, intermolecular hydrogen bonding progresses helically along b from carboxyl to ketone [O...O = 2.694 (5) A]. The carboxyl and lactone carbonyl groups in translationally related molecules within a helix both accept hydrogen bonds from the same water of hydration. The oxygen of this water in turn accepts a hydrogen bond from the hydroxyl group of a third screw-related molecule in an adjacent counterdirectionally oriented helix, yielding a complex three-dimensional hydrogen-bonding array. Intermolecular O...H-C close contacts were found to the carboxyl and lactone carbonyls, the hydroxyl, and the water. PMID- 11119010 TI - 2,2,4,4-Tetramethyl-1,5-diphenyl-6,7,8-trioxa-3-thiabicyclo AB - The structure of the title ozonide, C(20)H(22)O(3)S, produced without the use of ozone, has been defined at 123 (1) K. In the triclinic crystal, the molecule has symmetry close to C(s), and its ozonide and 1,4-oxathiane rings have envelope and chair conformations, respectively. The ozonide unit has an O-O bond length of 1.4721 (12) A and a C-O-O-C torsion angle of -1.45 (12) degrees. PMID- 11119011 TI - Pyrazine-2,3-dicarboxamide AB - In the crystal structure of the title diamide, C(6)H(6)N(4)O(2), linear tapes of carboxamide N-H.O and pyrazine C-H.N hydrogen-bond dimers are connected by N-H.O bonds to form a staircase-like pattern. PMID- 11119012 TI - [Alfred Fournier: "Contribution to the study of gonorrheal rheumatism". 1869]. PMID- 11119013 TI - Postprandial reactive hypoglycemia. AB - Postprandial reactive hypoglycemia (PRH) can be diagnosed if sympathetic and neuroglucopenic symptoms develop concurrently with low blood sugar (<3.3 mmol). Neither the oral glucose tolerance test (OGTT) nor mixed meals are suitable for this diagnosis, due to respectively false positive and false negative results. They should be replaced by ambulatory glycemic control or, as recently proposed, an hyperglucidic breakfast test. PRH patients often suffer from an associated adrenergic hormone postprandial syndrome, with potential pathologic consequences such as cardiac arrhythmia. PRH could result from (a) an exaggerated insulin response, either related to insulin resistance or to increased glucagon-like peptide 1; (b) renal glycosuria; (c) defects in glucagon response; (d) high insulin sensitivity, probably the most frequent cause (50-70%), which is not adequately compensated by hypoinsulinemia and thus cannot be measured by indices of insulin sensitivity such as the homeostatic model assessment. Such situations are frequent in very lean people, or after massive weight reduction, or in women with moderate lower body overweight. PRH is influenced by patient's alimentary habits (high carbohydrate-low fat diet, alcohol intake). Thus, diet remains the main treatment, although alpha-glucosidase inhibitors and some other drugs may be helpful. PMID- 11119014 TI - Autoimmune markers in slow type 1 diabetes: confrontation to type 1 diabetes. AB - Slow onset type 1 diabetes is an heterogeneous entity. Its clinical features may mimick type 2 diabetes but its pathophysiological mechanisms are close to type 1 diabetes. AIM OF THE STUDY: To find out the frequencies, levels and associations of ICA, GADab and IA-2ab in type 2 diabetic patients with atypical phenotype. To compare it to type 1 diabetes. PATIENTS AND METHODS: ICA, GADab and IA-2ab were determined in: - 61 patients (age at diagnosis 48.2 +/- 10, range 36-73 years) with an initial diagnosis of type 2 diabetes but having at least one symptom suggesting a slow type 1 diabetes (loss of weight, absence of obesity at diagnosis or secondary failure of oral hypoglycaemic agents). - 70 patients with type 1 diabetes (age 18 +/- 8.9, range 2-35 years). Clinical data evaluated in slow type 1 were maximal BMI, BMI and loss of weight at diagnosis and autoimmune disease. Fasting C-peptide and insulinemia were also assessed. RESULTS: (Slow type 1 diabetes versus type 1 diabetes). ICA (43% vs 70%; p <0.01) and IA-2ab (16% vs 75%; p <0.01) were more frequent in type 1. GADab were as frequent (62% vs 74%). Association of the three antibodies (15.7% vs 58.5%; p <0.05) were more frequent in type 1. Prevalence of GADab alone (27.5% vs 7.5%; p <0.05) was higher in slow type 1 diabetes and with higher levels (median 55.5 UI/ml vs 17 UI/ml; p <0.01). There was no difference for levels of ICA (25.5 UJDF/ml vs 28 UJDF/ml) or IA-2ab (11.5 UI/ml vs 38.5 UI/ml). BMI of GADab positive patients was lower. Delay of insulinotherapy was shorter in GADab or ICA positive patients. We did not find any relationship between antibodies presence and fasting C-peptide or insulinemia. CONCLUSION: Slow type 1 diabetes should be evoked in atypical type 2 diabetes. Slow onset type 1 diabetic patients have different autoimmune patterns suggesting a different pathophysiological process. GADab and ICA are useful markers to predict future insulinopenia. PMID- 11119015 TI - Managing type 2 diabetes in France: the ECODIA survey. AB - In order to describe the profile and medical management of type 2 diabetes patients in France, a descriptive cross-sectional survey was conducted in 1999 among a national random sample of 311 general practitioners and 51 specialists. A practitioner questionnaire was designed to collect information on a representative sample of 4,119 patients presenting with type 2 diabetes. Data collected included demographic and clinical information and a full description of diabetes management over a 6-month retrospective period. Over 50% of the patients were more than 67 years old; 54% were male. Diabetes had been diagnosed 8.9 years earlier on average, most frequently (73%) during a visit not related to diabetes' symptoms or complications. 42% of patients had a BMI > or =30 kg/m(2), 46% were hypertensive (BP > 140-80 mmHg), 53% had a LDL-Cholesterol over 1.3 g/l. Overall, 33% of patients had at least one diabetic complication. 60% of patients had had at least one HbA1c dosage in the last 6 months. Among them, 31% had a HbA1c level over 8% and 35% between 6. 5% and 8%. 85% of patients were treated with oral anti diabetic drugs, 9.5% with diet and exercise only and 5% with insulin. Sulfonylureas were the most commonly prescribed anti-diabetic agent, either alone or in combination. This survey confirms that the management of patients with type 2 diabetes is still often inappropriate in France despite recent progress. Improved disease management and monitoring is required in France as in other developed countries. PMID- 11119016 TI - Optimized transient insulin infusion in uncontrolled type 2 diabetes: evaluation of a pragmatic attitude. AB - Glucotoxicity generated by hyperglycemia creates a vicious circle worsening the imbalance of diabetes mellitus. A pump-optimized transient insulin treatment can be used to break this fate and restore some degree of insulin sensitivity in uncontrolled type 2 diabetes. The aim of this retrospective study was to evaluate type 2 diabetics with a secondary failure to a maximal oral antidiabetic therapy, treated with a transient subcutaneous insulin therapy during 3 days. The following criteria were analysed: delay before permanent insulin treatment, prognosis factors of evolution, weight evolution and glucose control in patients maintained under oral treatment. We studied 250 type 2 diabetics, and 515 insulin infusions. The average follow-up was 3.5 years. At the end of the follow-up 63 patients required insulin from the inception of the study (Group 1), 76 secondarily resumed insulin (Group 2), and 111 remained with oral treatment (Group 3). Patients in Group 1 were significantly older, with higher HbA1c and a lower body mass index (BMI). On average, the patients in Group 3 were submitted to less than 2 insulin infusions, their BMI from the beginning to the end of the follow-up remained stable, while HbA1c improved. We conclude that transient optimized insulin treatment during 3 consecutive days is effective. Thus, 45% of the initial global population remain under oral therapy after 3.5 years with a better glucose control and a stable weight. PMID- 11119017 TI - Counterregulatory responses to hypoglycemia in patients with glucokinase gene mutations. AB - The glucokinase gene is expressed not only in pancreatic B cells and in the liver, but also in pancreatic alpha cells, and in some cells of the central nervous system. A decreased glucokinase activity in the latter cell types may interfere with counterregulatory responses to hypoglycemia. In order to assess functional consequences of glucokinase mutations, counterregulatory hormones secretion and glucose production (6,6(- 2) H glucose) were monitored during an hyperinsulinemic clamp at about 2.4 pmol.kg(- 1).min(- 1) insulin with progressive hypoglycemia in 7 maturity onset diabetes of the young (MODY) type 2 patients, 5 patients with type 2 diabetes, and 13 healthy subjects. Basal glucose concentrations were significantly higher in MODY2 patients (7.6 +/- 0.4 mmol.l(- 1) ) and type 2 diabetic patients (12.4 +/- 2.3 mmol.l(- 1) ) than in healthy subjects (5.3 +/- 0.1 mmol.l(- 1), p<0.01) but counterregulatory hormones concentrations were identical. Insulin-mediated glucose disposal and suppression of endogenous glucose production at euglycemia were unchanged in MODY2 patients, but were blunted in type 2 diabetes. During progressive hypoglycemia, the glycemic thresholds of MODY2 patients for increasing glucose production (5.0 +/- 0.4 mmol.l(- 1) ) and for glucagon stimulation (4.5 +/- 0.4 mmol. l(- 1) ) were higher than those of healthy subjects and type 2 diabetic patients (3.9 +/- 0.1 and 4.1 +/- 0.1 mmol.l(- 1) respectively for glucose production and 3.7 +/- 0.1 and 3.5 +/- 0.1 mmol.l(- 1) for glucagon stimulation, p <0.02 in both cases). These results indicate that counterregulatory responses to hypoglycemia are activated at a higher plasma glucose concentration in MODY2 patients. This may be secondary to decreased glucokinase activity in hypothalamic neuronal cells, or to alterations of glucose sensing in pancreatic alpha cells and liver cells. PMID- 11119018 TI - Lipid peroxidation and antioxidant enzyme levels in type 2 diabetics with microvascular complications. AB - Serum levels of total cholesterol, triglycerides, lipoproteins, lipid peroxides (TBARS) and erythrocyte antioxidant enzyme activities were measured in 105 non insulin dependent diabetic patients, among whom 38 had microvascular complications (MVC) of diabetes. All the diabetic patients had higher concentrations of glycated hemoglobin (HbA1) compared to controls (10.51 +/- 2.42% vs 6.31 +/- 0.85% P <0.001). Significant increase of serum triglycerides (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL-C) and very low density lipoprotein cholesterol (VLDL-C) and a significant decrease of high density lipoprotein cholesterol (HDL-C) were observed in the diabetic patients compared to controls (TG: 2.31 +/- 0.9 mmol/l vs 1.53 +/- 0.48 mmol/l P <0. 001; TC: 5.94 +/- 1.4 mmol/l vs 4.3 +/- 0.85 mmol/l P <0.001; LDL-C: 3.96 +/- 1.33 mmol/l vs 2.39 +/- 0.8 mmol/l P <0.001; VLDL-C: 0.46 +/- 0.2 mmol/l vs 0.3 +/- 0.09 mmol/l P <0.001; HDL-C: 0.81 +/- 0.24 mmol/l vs 1.04 +/- 0.18 mmol/l P <0.001). Significantly increased levels of serum TBARS were observed in diabetic patients compared to those in controls (TBARS: 6.7 +/- 1.5 mmol/l vs 5.14 +/- 0.61 mmol/l P <0.001). Erythrocyte catalase (CAT) activity was increased and Glutathione peroxidase (GPx) activity was decreased in diabetic patients compared to controls, but no significant change in Superoxide dismutase (SOD) activity was observed in diabetic patients (CAT: 104.94 +/- 37.1 KU/g Hb vs 85.8 +/- 23.6 KU/g Hb, P <0.01; GPx: 30 +/- 9.7 U/g Hb/min vs 40.84 +/- 12.3 U/g Hb/min, P <0. 001; SOD: 2.4 +/- 1.2 U/mg Hb/min vs 2.55 +/- 0.84 U/mg Hb/min, P=NS). In comparison with the diabetic group without MVC, the diabetic group with MVC had decreased GPx and SOD activities, while no difference was observed between these two groups regarding CAT activity (GPx: 25.32 +/- 8.4 U/g Hb/min vs 34.5 +/- 8.8 U/g Hb/min, P <0.001; SOD: 1.83 +/- 0.53 U/mg Hb/min vs 2.84 +/- 1.4 U/mg Hb/min, P<0.001; CAT: 106.3 +/- 39.9 KU/g Hb vs 103 +/- 34.9 KU/g Hb, P =NS). TBARS concentrations were significantly increased in the group with MVC compared to the group without these complications, indicating a positive relationship between TBARS and MVC of diabetes (7.05 +/- 1.23 mmol/l vs 6.3 +/- 1.02 mmol/l, P <0.001). Serum triglycerides, LDL and VLDL cholesterol concentration were significantly higher in diabetics with MVC than in diabetics without the complications (TG: 2.7 +/- 0.98 mmol/l vs 2.13 +/- 0.82 mmol/l, P<0.01; LDL - C: 4.45 +/- 1.3 mmol/l vs 3.67 +/- 1.3 mmol/l, P <0. 02; VLDL-C: 0.53 +/- 0.19 mmol/l vs 0.43 +/- 0.16 mmol/l, P <0.01), and the serum levels of TC in the group with MVC showed a positive correlation with their lipid peroxide levels (r =0.368, P <0.001). The increase in TBARS and the decreased GPx and SOD activities in diabetics with MVC in this study indicate that these factors may contribute to the occurrence of micro vascular complications in NIDDM patients. PMID- 11119019 TI - Mutation screening of the PPARalpha gene in type 2 diabetes associated with coronary heart disease. AB - The peroxisome proliferator-activated receptor alpha (PPARalpha) is a ligand activated transcription factor belonging to the nuclear hormone receptor superfamily. PPARalpha plays a key role in lipid and glucose metabolism, inflammatory response and energy homeostasis. The aim of our study was to screen the PPARalpha gene for mutations, and to test the genetic contribution of PPARalpha in diabetes and its vascular complications. The first two non coding exons and the coding region of the PPARalpha gene were screened by single strand conformation polymorphism (SSCP) and sequencing in 74 unrelated Type 2 diabetic patients with history of coronary heart disease (CHD) (18 Caucasian and 56 Indian subjects). A total of 7 nucleotide variants were detected: two single amino acid substitutions, a silent mutation, four intron base changes. Association studies were undertaken in two populations of Type 2 diabetic patients from Pondichery and from France, to test the distribution of allelic frequencies for L162V (exon 5) and A268V (exon 7) polymorphisms. No association was found between these PPARalpha variants and diabetes or CHD. However, in the Caucasian diabetic male population with CHD, the Val162 allele carriers showed higher concentrations of total cholesterol and Apo B when compared to non-carriers (p =0.01 and p =0.005, respectively). A trend toward elevated concentrations of total cholesterol and Apo B was also observed in the Caucasian diabetic male patients without CHD carrying Val162 allele. In conclusion, it is likely that PPARalpha gene does not have a major role in diabetes and CHD in our populations, although we can not exclude a minor contribution of the PPARalpha gene to the risk of CHD associated with Type 2 diabetes through a modulation of atherogenic plasma lipids. PMID- 11119020 TI - Hyperosmolar nonketotic syndrome with hypernatremia: how can we monitor treatment? AB - We report the case of a 62 year-old symptomatic patient with severe hyperglycemic hyperosmolality associated with hypernatremia. During treatment, the progressive decrease in serum tonicity, which resulted in the amelioration of the neurological symptoms, followed the decrease in serum glucose and mainly the corrected serum sodium levels rather than the decrease in the uncorrected serum sodium levels. The case illustrates the usefulness of glucose - corrected serum sodium levels to monitor treatment in such conditions in order to avoid neurological consequences caused by the decrease in serum osmolality. PMID- 11119021 TI - Ketoacidosis accompanied by epileptic seizures in a patient with diabetes mellitus and mitochondrial myopathy, encephalopathy, lactic acidosis and stroke like episodes (MELAS). AB - We herein report a rare case of MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes) and diabetes mellitus with ketoacidosis. An 18-year-old female patient was diagnosed to have diabetes mellitus and insulin therapy was thereafter initiated. At 26 years of age, she was hospitalized for diabetic ketoacidosis, soon followed by a loss of consciousness, left-sided dysmetria, and ataxic speech. MELAS was diagnosed because of the presence of ragged red fibers in a muscle biopsy. At 33 years of age, she was admitted to our hospital because of ketoacidosis and partial status epilepticus. A blood gas examination revealed as follows; arterial pH, 6.88; bicarbonate, 2.1 mmol/l; base excess - 29.8 mmol/l. The serum level of glucose had also increased to 30 mmol/l. The serum levels of lactate and B hydroxybutyrate were elevated to 11.4 mmol/l and 1,990 micromol/l, respectively. Ketoacidosis improved by fluid replacement and continuous intravenous insulin infusion. A brain MRI demonstrated hyperintensity areas on FLAIR images in the bilateral temporal lobes and the cerebellum. A proton MRS demonstrated the abnormal lactate accumulation in the bilateral temporal and occipital lobes. Since epileptic seizures are rare in patients with diabetic ketoacidosis, such seizures may indicate the existence of MELAS syndrome. PMID- 11119022 TI - [Is scuba diving allowed in diabetic patients treated with insulin?]. AB - Scuba diving is usually prohibited in diabetic patients at risk of hypoglycemic attacks (i.e. IDDM and NIDDM treated with sulfonylureas) due to the particular severity of these episodes in a hostile environment. In fact, diabetic subjects can perform this popular leisure activity without extra-risk, provided strict requisites in aptitude, practice and environment are enforced. These limitations lead to the establishment of practical guidelines, as proposed in this paper. PMID- 11119024 TI - [Le Journal de Chirurgie: a key tool for authors and readers] PMID- 11119023 TI - [How to prescribing Viagra in practice...]. AB - Diabetes can induce sexual disorders by different mechanisms. These troubles are more frequent in diabetics subjects. Thus, management of sexual impotence is an important aspect of diabetes care. However, most diabetologists are not trained to treat sexual disorders. The recent availability of oral drugs, i. e. Sildenafil (Viagra), has partly simplified the treatment of sexual impotence, particularly in diabetic patients. However, Viagra is efficient in only 60% cases in diabetic subjects. In the remaining cases, intracavernosal injections or vacuum can be used. Since Viagra has been available, more diabetic patients complained with sexual disorders, and ask for treatment of impotence. Cardio vascular diseases must retain more attention in diabetic patients who are exposed to silencious myocardial ischemia. In such subjects, Viagra is not contra indicated, but must be used after myocardial explorations, especially if the patients have cardio-vascular risk factors. However, patients and their doctors have been threatened by death cases reported with Viagra in United States. The lack of detailed informations has restrained Viagra's prescription. The following explains how to manage sexual disorders as part of diabetes care, and suggests rules for Viagra's prescription in diabetic patients. PMID- 11119025 TI - [Gene therapy of liver tumors]. AB - Gene therapy is a new therapeutic option for liver tumors which has been evaluated in animal models. The liver has characteristic features particularly interesting for gene transfer. Because of the rapid tumor growth within a quiescent parenchyma, retroviral transfer could express selective tropism. Temporary anatomic vascular exclusion would allow selective perfusion of the liver and avoid extrahepatic diffusion of the gene therapy. The early results have been promising in the animal. Clinical studies are currently under way in patients with liver metastasis from colorectal cancer. PMID- 11119026 TI - [Pretherapeutic assessment of esophageal carcinoma]. AB - The prognosis of esophageal carcinoma is closely related to tumor stage and treatment modalities are applied according to the TNM stage of the disease and the performance status of the patient. Selecting the appropriate treatment based on accurate staging is of paramount importance. The subject of this review concerns advances in pretherapeutic assessment of esophageal carcinoma. Advantages and limits of pretherapeutic examinations are exposed with special interest to endoscopic and radiological investigations (sonography, computed tomography, magnetic resonance imaging, PET-scan, endoscopic ultrasonography, bronchoscopy.). Impact on therapeutic management is clarified. PMID- 11119028 TI - [Delorme procedure for rectal prolapse]. PMID- 11119027 TI - [Guidelines of the SFAR (Societe Francaise d'Anesthesie et de Reanimation). Antibiotic combinations or monotherapy in surgery and surgical intensive care. Extracts relating to the "visceral surgery" conference of experts]. PMID- 11119029 TI - [Surgical treatment of vaginal hydrocele]. PMID- 11119030 TI - [Y loop]. PMID- 11119032 TI - [Toothpicks: be careful!]. PMID- 11119031 TI - [Surgical management of community-acquired peritonitis in children. Analysis of a survey]. AB - The purpose of this work was to determine current practices of pediatric surgeons in the management of community peritonitis, excepting antibiotic use and resuscitation car. A questionnaire was sent to 63 French pediatric surgeons in France. Forty-six answers concerning appendicular peritonitis were analyzed. Laparoscopy was a good indication for 40 surgeons. A sample for bacteriological examination was never ordered. Conditions for washing and drainage were detailed. Treatment to prevent postoperative pain was used by all. PMID- 11119033 TI - [A 73-year-old woman with a voluminous epigastric mass of the left hypochondrium]. PMID- 11119034 TI - [Hormone replacement therapy in menopause and colorectal disease]. AB - Colorectal disease is frequent. Accumulating evidence indicates that postmenopausal hormone replacement therapy may reduce the risk of this pathology in women. Through MEDLINE computer searches, we identified English-language articles with quantitative data on the relation of postmenopausal hormone replacement therapy to colorectal cancers and adenoma. Twenty-four studies about colorectal cancer and five studies about colorectal adenoma were reviewed. Epidemiological data suggest a 20 to 30% reduced risk of colorectal cancer and adenoma among women taking postmenopausal hormones. There is biological evidence to support this association. PMID- 11119035 TI - [Arcus tendineus fascia pelvis: anatomical study]. AB - OBJECTIVE: To reveal the anatomy of arcus tendineus fascia pelvis. MATERIAL: and methods. 2 fixed female cadaver pelvises (88 and 66 years old) were dissected. RESULTS: The arcus tendineus fascia pelvis is a fibrous recess of the pelvic fascia which is 10 cm long, laterally to the obturator internus muscle and medially to the peritoneum. It reaches the ischial spine lower and anteriorly to the pubo-vesical ligament. The third posterior part of the arcus tendineus of the pelvic fascia is commun with the posterior part of the arcus tendineus of the levator ani. This third posterior part is like a curve concave anteriorly. This curve is thick and easy to recognize by the palpation. This third posterior curve is 1cm in front of the ischial spine and 2cm in front of the pudendal vessel which took place behind the ischial spine. The vessels of the obturator internus muscle originated from the internal iliac vessel and crossed laterally the median part of the arcus tendineus of the pelvic fascia. CONCLUSION: Sutures must be placed through the anterior and median part of the arcus tendineus fascia pelvis, in front of the posterior part of the arcus tendineus fascia pelvis to avoid any injury to the pudendal vessels. PMID- 11119036 TI - [Laparoscopic retropubic colposuspension (Burch procedure). Techniques and continence results]. AB - INTRODUCTION: Laparoscopy allows to perform Burch colposuspension in case of genuine urinary stress incontinence but this approach must not lead to reduced success rates. OBJECTIVE: The aim of this study was to describe laparoscopic Burch techniques and evaluate urinary continence and satisfaction rates. MATERIALS AND METHODS: Clinical and urodynamic data were reviewed for 48 consecutive patients operated on from May 1993 to June 1999. The methods for entering the space of Retzius and performing the colposuspension were analyzed. The continence and satisfaction rates were evaluated with a questionnaire. RESULTS: The operative technique we are now using is the extraperitoneal approach with an umbilical trocar site. A non-absorbable mesh is fixed to the Cooper's ligament and to the vaginal fascia. With a mean follow-up of 41 months, the satisfaction rate is 76.8% with a cure rate of 37.3% and an improvement rate of 41.8%. CONCLUSION: Long term results with laparoscopic Burch colposuspension are relatively good but a bit lower than those published with traditional open technique. The effects of the learning curve with an evolving technique are to be considered when analyzing the results. PMID- 11119037 TI - [Is mammography useful in Paget's disease of the breast?]. AB - PURPOSE: Interest of the mammogram in Paget's disease of the breast, especially for a therapeutic decision in otherwise asymptomatic women with Paget's disease, who would be candidates for conservative treatment. MATERIALS AND METHODS: 61 women with histological Paget's disease of the nipple, treated by mastectomy, were retrospectively analyzed with clinical, radiological and pathological correlations. RESULTS: An underlying carcinoma was found in 60 cases (98.4%), atypical epithelial hyperplasia in one. In the 24 women without breast palpable mass, 17 (71%) had a normal mammogram, 12 (50%) had carcinoma with an invasive component, 14 (58%) had a cancer at a distance from the nipple, 17 (71%) had a multifocal carcinoma. All 37 women with a palpable mass had a pathological mammogram, 36 of them had carcinoma with an invasive component, 35 (95%) a cancer at a distance from the nipple, 31 (84%) a multifocal carcinoma. CONCLUSION: Mammogram is of limited value in management of Paget's disease of the breast for women without breast palpable mass; it can not predict the site of malignancy, nor the invasive component. PMID- 11119038 TI - [Psychosocial counseling in assisted reproduction technologies: a pilot study of a new approach: the "ressources evaluation"]. AB - OBJECTIVES: During these last 15 years, our comprehension of what infertile couples go through has deepened, allowing us to target our offer of psychological assistance in the framework of medically assisted reproduction. From intervening in critical situations, we have passed onto a preventative model, integrated in the first stages of medical investigation. This preventative concept is called "resource counseling". The couples' capacity to cope with the ups and downs of treatment (stress, failure, parenthood) is evaluated in order to define the most adequate type of psychological assistance. MATERIAL: and method. Follow-up of 60 couples, 12 months after counseling by means of an open half-hour interview and a semi structured questionnaire: pilot study. RESULTS: 85% of the couples accepted to participate. Of these couples, 73% felt reassured by the counseling and 50% uncover personal resources enabling them to better cope with their situation. 89% agreed that counseling should take place before treatment. CONCLUSION: "Counseling" gives most not demanding couples reassurance, enabling them to become partners of the team for questions of decision-making and compliance. However, couples have difficulty in asking for help which underlines the importance of envisaging to systematically fix another appointment after 6 to 12 months of treatment. This would also allow further prospective detection of the psychosocial risks of the treatments and to optimize the prevention of their side effects. PMID- 11119039 TI - [Prenatal outcome of sex chromosome anomalies diagnosed during pregnancy: a retrospective study of 47 cases]. AB - OBJECTIVE: To study the circumstances of discovery and the prenatal outcome of sex hormone anomalies diagnosed by invasive prenatal techniques during pregnancy and analyze which factors could be implicated in the parents' choice to terminate or carry on with pregnancy. METHODS: We reviewed retrospectively 47 cases of sex chromosome anomalies diagnosed and managed in our prenatal diagnosis unit over a 9-year period between January 1, 1990 and December 31, 1998. Only cases karyotyped in our laboratory and with a complete follow-up were considered. RESULTS: Cytogenic findings were mainly turner syndrome (n=25) and Klinefelter syndrome (n=12). The other karyotypes were the following: 47, XXX (n=6), 47, XYY (n=2), and 49, XXXXY (n=2). Among the 47 pregnancies, 11 (23.4%) were carried to term. The rate of pregnancy termination (68.1%) was high. The decision to terminate varied depending on the abnormal karyotype: 88% for Turner syndrome, 42% for Klinefelter syndrome, 33% for 47, XXX, 50% for 47, XYY and 100% for 49, XXXXY. The pregnancy termination rate was significantly higher when one or more abnormal ultrasound findings was present (92.3% vs 41.2%, p<0.01). CONCLUSION: Our study confirms that termination rates remain high in case of sex hormone anomalies. Associated ultrasonographic findings play a major role in the parents' choice to terminate or carry on with the pregnancy. It would appear that the development of consensual guidelines in pluridisciplinary fetal medicine centers can help reduce the disparities currently observed among French centers in the management of fetuses with sex chromosome anomalies. PMID- 11119040 TI - [Late rejection of Mersylene mesh used for promontory fixation with colonic obstruction]. AB - We describe a case of colonic obstruction in a 48-year-old woman, 3. 5 years after surgery for genito-urinary prolapse. The causal agent was the Mersylene mesh used for promontory fixation. Late rejection of this kind of prosthesis has been described previously, but such a digestive complication is exceptional. PMID- 11119041 TI - [Complete remission after abdominal wall metastasis from uterine cervical cancer on a laparoscopic trochar site: a case report]. AB - Abdominal wall metastasis to laparoscopic trochar sites after preoperative staging procedure is rare for uterine cervix cancer. Prognosis is unfavorable. We report a case of metastasis to a laparoscopic trochar site in a patient with a stage IIB cervical cancer with no nodal involvement who is alive four and a half years after radical surgery and radiotherapy. PMID- 11119042 TI - [Sonographic findings of acardiac twin in the first trimester of pregnancy]. AB - Twin Reversed Arterial Perfusion (TRAP) sequence, or the acardiac anomaly, is a rare complication of monozygotic twins, that occurs in approximately 1 in 35,000 births. In the first trimester, once the diagnosis of monochorionic twins is made, particularly if one embryo shows abnormal development or generalized edema, the TRAP sequence must be considered. Moreover, malformations reported in the perfused twin are often severe. Diagnosis is confirmed by using Doppler ultrasound which demonstrated the presence of retrograde perfusion in the umbilical cord of the abnormal twin. The more complete the body form is, the more similarity the vascular relationships have to the pattern found in a full-term fetus. Final outcome and treatment decisions can be determined based on hemodynamic criteria. We report a case of a misdiagnosed TRAP sequence with no structural malformations detected by ultrasonography at 12 weeks of gestation. PMID- 11119043 TI - [Re: "Gestational trophoblastic diseases." Potential mechanism of formation of complete diploid moles]. PMID- 11119045 TI - [In Process Citation] PMID- 11119044 TI - [Should the intensive care anesthesiologist call in the obstetrician before performing epidural analgesia during labor? SFAR (Societe Francaise d'Anesthesie Reanimation). CNGOF (College National des Gynecologues et Obstetricien Francais]. PMID- 11119046 TI - [Vigilance disorders and driving]. PMID- 11119047 TI - [Myotubular myopathy]. AB - The myotubular myopathy is an X-linked centronuclear myopathy characterized by severe neonatal hypotonia and generalized muscle weakness which most frequently results in the premature death of the newborn infants by respiratory failure. The characteristic muscle histopathology consists in centrally positioned nuclei in most muscle fibers. In 1996, the gene responsible for the disease, MTM1, was identificated in Xq28 region. Since then, more than hundred mutations have been isolated. Genotype - phenotype correlation is complex because mutations are found along the entire coding sequence of the MTM1 gene. Most mutations are associated with very severe phenotype with death before the first year of life, however a milder phenotype has been individualized. It is important to be aware of the existence of such milder MTM phenotype because in those patients a very mild expression permit normal life into adulthood. PMID- 11119048 TI - [Clinical manifestations and therapeutic approach in neurosarcoidosis]. AB - Neurological impairment is a frequent cause of morbidity and mortality in patients with sarcoidosis. The aim of this study was to evaluate the clinical manifestations of the disease, the response to corticosteroids and alternative treatments. During a 5 year period, diagnosis of neurosarcoidosis was performed in 40 patients. We retrospectively analyzed clinical, laboratory data and response to treatments. Mean age was 41.3 years (range 17-72). Mean time of follow-up was 46 months. Neurologic signs were the first symptom in 50 p. 100 of cases and an isolated manifestation in 12.5 p. 100. Central nervous system impairment was seen in 60.7 p. 100, meningitis in 27 p. 100. Other clinical manifestations were cranial nerve palsies (27 p. 100), peripheral neuropathy (33 p. 100), myopathy (16 p. 100). Eighty percent of the patients were treated by corticosteroids. Because of a lack of efficacy 40 p. 100 of patient required alternative treatment (including methotrexate, cyclophosphamide, azathioprin, cyclosporin). Complete recovery was observed in only 27.5 p. 100 of cases confirming the severity of neurosarcoidosis. Forty percent of patients were clinically stable and 10 p. 100 worsened. No patient died. This study confirms that intensive initial treatment is often necessary to prevent irreversible lesions. Alternative treatment should be rapidly initiated in resistant forms. PMID- 11119049 TI - [Incidence of grasping and its relationship to cerebral lesions]. AB - Grasping is associated with frontal lobe pathology. Nevertheless, there is lack of precise anatomical correlations and very few studies are published. The aim of the study was to determine the incidence of the grasping and its relationship to cerebral lesions. We studied 236 patients admitted to the Neurology Department (108 women and 128 men; mean age 65.3), and tested with a standardized procedure (De Renzi and Barbieri, 1992). A score of grasping was determined for each patient. The locations of the cerebral lesions were assessed by two neurologists using the method of Damasio and Damasio (1989). Grasping was found in 38 patients (16.1p.100) with dementia or cerebral damage. In all cases, the lesion affected the frontal lobe. The patients with grasping showed a significant higher number of lesioned areas particularly for the frontal and the parietal regions. The score of right grasping was significantly higher with a lesion in the right paraventricular frontal and in the left parietal paraventricular areas. The score of left grasping was significantly higher with a lesion in the left frontal paraventricular area. These results are discussed in relationship with motor control. PMID- 11119050 TI - [Interobserver reliability of a new horizontal pointing manoeuvre: comparison with conventional tests]. AB - Finger-to-nose test, fine finger movements, to maintain arms against gravity, alternate movements of hands, Barany's test, muscle tone evaluation, Stewart Holmes test and handwriting are the conventional clinical tests most frequently used in daily practice to evaluate voluntary movements of the upper limb. We describe a new clinical manoeuvre consisting of a horizontal pointing movement of one upper limb (the moving limb) towards the contralateral motionless limb. We have analyzed the reliability of our new test and of the conventional tests between 2 observers using kappa statistics in a group of 34 right-handed neurological patients. Agreement is very good for our test, since k has a value of 0.63 for left upper limb and a value of 0.64 for right upper limb. The only conventional test characterized by a higher interobserver reliability is handwriting. Furthermore, our manoeuvre is original, as attested by partial association coefficients analysis between our test and conventional tests. This might be due to the fact that our manoeuvre is the sole test investigating rapid proximal movements towards a fixed area in space. In conclusion, our horizontal pointing manoeuvre has a very good reliability between 2 observers and appears original. PMID- 11119051 TI - [Topiramate in clinical practice (part 1). Multicentric retrospective analysis of the efficacy of topiramate as add-on therapy according to the topographic form of focal epilepsy]. AB - Randomized, controlled studies of new antiepileptic drugs do not always highlight their best utilization in clinical practice. The authors gathered 361 cases of focal epilepsies treated with topiramate (TPM) as an add-on to other anti epileptic drugs prior to marketing. Among these, only 237 were treated for at least 3 months and analyzed here. These patients were treated by neurologists in a clinical setting, with free choice of associated drugs, titration and final daily doses. Compared with controlled studies, TPM was titrated slowly (mean rate: 43 mg/week, vs 100 to 200), and was given at a lower final dose (346 mg/d, vs 200 to 1000). This analysis confirmed the efficacy of TPM as add-on therapy in focal epilepsies (9.3p.100 totally controlled, 19 p.100 with reduction of seizures=90 p. 100, 52.7 p.100 responders at=50 p.100). It showed that there was a striking response in epilepsies originating from the central areas, which are often drug-resistant (19 p.100 totally controlled, 33.33 p. 100 with reduction of seizures=90 p.100). There were responders in all topographic groups. There was however no specific response according to etiology. PMID- 11119052 TI - [Contribution of magnetic resonance imaging in sclerotic combined degeneration of the spinal cord due to vitamin B12 deficiency]. AB - Subacute combined degeneration (SCD) of the spinal cord is known to present histopathologically degenerative lesions in the spinal cord, but few studies on the neuroradiological findings have so far been reported. We present the interest of initial and follow-up MR findings in three cases of SCD. In the three cases, a causal event precipitated the onset of neurological symptoms: general anesthesia for the first and the third one and folic acid treatment for the second one. Clinical evolution was favorable after specific treatment with nearly total recovery. The initial MR study disclosed lesions predominantly involving the posterior columns of the spinal cord: high intensity on T2 weighted image was seen in the initial MR study and disappeared three months after treatment in correlation with good recovery, but with a delay. The recognition of this MR pattern suggests that MRI may be used in conjunction with clinical assessment to confirm the diagnosis and to monitor the efficacity of treatment in SCD. PMID- 11119053 TI - [Lysosome enzyme pseudodeficiency]. AB - Inherited deficiency of lysosomal hydrolase often displays different clinical features. However, a greatly reduced enzyme activity may be observed in healthy individuals. This pseudo-deficiency concerns at least nine lysosomal hydrolases. When a deficiency has been proved, the presence of mutations known to cause pseudodeficiencies must be searched, above all in Tay-Sachs disease and metachromatic leukodystrophy. PMID- 11119054 TI - [Progressive multifocal leukoencephalopathy and pulmonary sarcoidosis]. AB - An association of progressive multifocal leukoencephalopathy (PML) and sarcoidosis in a 47 year-old-woman is reported. This is the third case in which PML has been diagnosed by PCR. Clinical, biological and radiological features were in agreement with previous findings in immunologically suppressed patients. JC virus should be systematically detected by PCR in blood and cerebrospinal fluid (CSF) in patients with sarcoidosis presenting neurological and radiological PML manifestations. PMID- 11119055 TI - [Frontal pseudo-tumoral form of adrenoleukodystrophy]. AB - A case of adrenoleukodystrophy in a 9-year old boy revealed by a predominant frontal syndrome is reported. Brain MRI showed an unusual pseudo-tumoral frontal lesion. The diagnosis was confirmed by increased plasma levels of very long chain fatty acids. His young brother had an isolated adrenal insufficiency with normal brain MRI. The frontal predominance of the lesion and the clinical polymorphism of the disease in this family are discussed. PMID- 11119057 TI - [Relapsing polyneuropathy in the post-partum period]. AB - A patient had five relapses of polyneuropathy: four developed during post-partum. The rapid onset of symptoms with subsequent complete recovery are in favor of a recurrent Guillain-Barre syndrome rather than a chronic relapsing inflammatory polyneuropathy. PMID- 11119056 TI - [Primary lateral sclerosis with breast cancer, a potential paraneoplastic neurological syndrome]. AB - Few reports indicate that motor neuron diseases may have paraneoplastic origin. A 70 year-old woman suffering from progressive upper motor neuron disease is presented. Laboratory, radiological and neurophysiologic studies were compatible with primary lateral sclerosis. Six years later a routine screening led to the discovery of a breast cancer, suggesting that the upper motor neuron syndrome could be paraneoplastic. So, in female patients with primary lateral sclerosis, a mammography should be recommended to search for breast cancer. PMID- 11119058 TI - [ [In Process Citation] PMID- 11119059 TI - [History of German neuropathology]. PMID- 11119060 TI - [E.D.S.S. Expanded Disability Status Scale]. PMID- 11119061 TI - [Immunological mechanisms of fatigue in patients with multiple sclerosis]. PMID- 11119062 TI - [Relationship between fatigue in multiple sclerosis and the chronic fatigue syndrome]. PMID- 11119063 TI - [Perspectives on managing patients with fatigue]. PMID- 11119064 TI - [Analysis of subjective complaints of fatigue in patients with multiple sclerosis]. PMID- 11119065 TI - [Approach to cognitive disorders in multiple sclerosis. Relationships between handicap and MRI findings]. PMID- 11119066 TI - [Cognition assessment in multiple sclerosis: difficulties in methodology and assessment protocols]. PMID- 11119067 TI - [Workshop on fatigue and multiple sclerosis]. PMID- 11119068 TI - [Workshop on cognitive functions and multiple sclerosis]. PMID- 11119069 TI - Trace Metals' abnormalities in hemodialysis patients: relationship with medications. AB - A multicenter collaborative study was performed to investigate the prevalence of abnormal blood contents of 6 trace metals, copper (Cu), zinc (Zn), aluminum (Al), lead (Pb), cadmium (Cd), and mercury (Hg), in hemodialysis (HD) patients and to analyze their relationship with the medications, such as CaCO3, Ca acetate, Al containing phosphate-binding agents, 1,25-dihydroxy vitD3, 1-hydroxy vitD3, and erythropoietin (EPO), as well as hematocrit level, by chi-square statistics. From 6 medical centers in Taiwan, we included 456 patients in maintenance HD for more than 4 months for this study, and they had continued the previously mentioned medications for at least 3 months. Blood samples were collected before initiating HD, and atomic absorption spectrophotometry was used to measure plasma levels of Cu, Zn, and Al as well as whole blood levels of Pb, Cd, and Hg. Three hundred seventy-five (78%) of the HD patients had low plasma Zn levels, that is, <800 microg/L, and the mean (+/-SD) concentration was 705.8 (+/-128.23) microg/L in all subjects. One hundred forty-one (31%) of the HD patients had high plasma Al, that is, >50 microg/L, and the mean (+/-SD) was 44.30 (+/-28.28) microg/L in all subjects. Three hundred thirty-three (73%) of the dialysis patients had high Cd levels, that is, >2.5 microg/L, and the mean (+/-SD) was 3.32 (+/-1.49) microg/L in all subjects. The majority of HD patients had normal blood levels of Cu, PB, and Hg. Only 21 (4. 6%), 5 (1.1%), and 3 (0.06%) patients had elevated blood levels of Cu, Pb, and Hg, respectively. Their mean (+/-SD) blood concentration of Cu, Pb, and Hg were 1,049.78 (+/-233.25) microg/L, 7.45 (+/-3.95) microg/dL, and 3.17 (+/-25.56) microg/L, respectively. Three patients had elevated plasma Hg concentrations, that is, 546, 12.6, and 24.0 microg/L, respectively. In the 152 normal healthy age and sex matched control group, the blood levels of Al, Cd, and Pb were all significantly lower than the HD patients. However, the levels of Cu and Zn were higher in the control group. The Hg level was not significantly different in both groups. There was no statistical difference between patients with normal and abnormal blood levels of trace metals in various medications except Al containing phosphate binder. The Al containing phosphate binder users had significantly higher plasma Al levels (54.71 +/- 26.70 versus 41.15 +/- 28.03 microg/L, p < 0.001) and hematocrit levels (29.61 +/- 4.61 versus 27. 81 +/- 3.91, p < 0.0005). There was no statistical correlation between erythropoietin (EPO) dose and hematocrit level in these patients. In conclusion, the blood level of trace metals of these HD patients except Al was not related to their medications. However, caution must be exercised in interpreting this result as dose and duration of medication; efficiency of HD and water treatment may play an important role. Otherwise, environmental factors, diet, and the aging process may contribute to the trace metal burden in uremia. Thus, Zn and Cu are abundant in seafood, and Cd is abundant in contaminated plants such as rice. PMID- 11119070 TI - Development of a DNA immunoadsorbent: coupling DNA on sepharose 4FF by an efficient activation method. AB - To remove anti-DNA antibodies from a patient's plasma with systemic lupus erythematosus (SLE), a DNA immunoadsorbent was developed by covalently coupling calf thymus DNA on activated Sepharose 4FF. Sepharose 4FF was activated with 5 norbornene-2,3-dicarboximido carbonochloridate (Cl-CO-ONB), which was proven to be a very effective method for preparation of affinity chromatographic adsorbents. The activation was carried out in dry acetone using 4 (dimethylamine)pyridine (DMAP) and triethylamine (TEA) as catalysts at 4 degrees C or at room temperature. The coupling of DNA to the activated support was investigated as a function of pH, temperature, time, concentration of DNA, and activation level. It was found that the pH for optimal coupling is 3.0, and the amount of coupled DNA increases with an increase either in the concentration of DNA or the activation level. The maximum amount of coupled DNA could reach 1.0 mg DNA/ml support. The incubation of 5 to 20 ml of SLE plasma with 1.0 ml of adsorbent resulted in an 80 to 90% decline in the anti-DNA antibody level. Nonspecific adsorption for normal IgG and total protein is less than 15%. PMID- 11119071 TI - Cyclosporin A and therapeutic plasma exchange in the treatment of severe systemic lupus erythematosus. AB - Despite treatment with intensive immunosuppressive drug regimens, often the prognosis of patients suffering from systemic lupus erythematosus (SLE) is poor. Side effects such as infections and malignancies often occur. It was the aim of this trial to assess the effect of immunosuppression, in particular with cyclosporin, and the efficacy, safety, and clinical utility of intermittent treatment with therapeutic plasma exchange (TPE) in comparison to previous intensive therapy strategies using corticosteroids, azathioprine, and/or cyclophosphamide. In this prospective trial, 28 patients (24 women, 4 men, aged 36.3 +/- 11.8 years at the diagnosis of SLE) were treated for up to 10 years with drug regimens out of corticosteroids, azathioprine, and/or cyclophosphamide. Then, over a period of up to 8 years, in addition to conventional therapies, especially in active stages of the disease with extremely high concentrations of anti-DNA, anti-nuclear antibodies, and circulating immunocomplexes, TPE sessions were carried out depending on symptomatology. In addition, the patients received cyclosporin. Compared with previous treatment modalities, clinical symptoms improved more quickly and more effectively. During the study period of a mean of 5 years, corticosteroids, azathioprine, and cyclophosphamide were reduced by 40 to 100%. No severe side effects were seen. In acute stages of SLE and in forms with persistently high antibody levels, the addition of TPE sessions and cyclosporin as the basic immunosuppressive drug is mostly very effective with regard to improving clinical symptomatology. PMID- 11119072 TI - Transperitoneal transport of glucose in vitro. AB - The effect of fluid mixing intensification, damage of mesothelial cells, gentamicin, and icodextrin on the diffusive glucose transport across the peritoneal membrane were evaluated in in vitro studies. A mathematical model of mass transport was used to calculate the diffusive permeability, expressed as a diffusive permeability coefficient (P). In the control conditions, the rate of glucose transfer from the interstitial to the mesothelial side of membrane (I- >M) and in the opposite direction (M-->I) remained constant, and the P value at mean was 2,731 +/- 1,493 x 10-4 (cm x s-1). The change of the stirring rate from 5.5 to 11 ml/min increased P values by about 74% for transport direction I-->M and 58% for M-->I, and the change from 11 to 22 ml/min enhanced P at mean by about 42% for both directions. The damage of the mesothelial layer, using sodium deoxycholate (2.5 mmol/L; 103.6 mg%), increased the glucose transfer from the interstitial to the mesothelial side of the peritoneum by 41% and to the opposite direction by 70%. Addition of icodextrin to the glucose solution increased glucose bidirectional transport at mean by about 14% for I-->M and 24% for M-->I. Furthermore, gentamicin did not change the I-->M transfer, but diminished M-->I transport by about 12%. In conclusion, the reduction of unstirred fluid layers at the mesothelium and the interstitium-fluid interfaces, removal of mesothelium, and addition of icodextrin increased the diffusive glucose transport in vitro; unstirred fluid layers restricted glucose transfer (I-->M) more than the mesothelium; and peritoneal glucose transport, directed from the mesothelial to the interstitial side of the peritoneum, decreased slightly after the addition of gentamicin. PMID- 11119073 TI - Supplemental systemic oxygen support using an intestinal intraluminal membrane oxygenator. AB - An intraluminal membrane oxygenator (IMO) prototype was surgically inserted in the ileum and evaluated as a method of supporting systemic oxygenation in an acutely hypoxemic porcine model. Animals were assigned randomly to the test (n = 12) or the control (n = 8) groups, which underwent identical protocols with the exception of the O2 flow in the IMO device, which was shut off in the control group. In each case, hypoxia was induced by a reduction in the inspired oxygen fraction (FiO2) to 0.14. A highly significant improvement (p < 0.005) in arterial and venous O2 content and lower arteriovenous O2 difference (p < 0.05), cardiac output, and hemoglobin (p < 0.005) were found in the test group during hypoxia. The results show that it is possible to meet a physiologically significant portion of the body's O2 demands via the intestine during respiratory hypoxia and suggests that similar devices may be of significant potential value as a supplemental oxygenation device in cases of respiratory distress. PMID- 11119074 TI - Low power laser protects human erythrocytes In an In vitro model of artificial heart-lung machines. AB - The protective effect of the low power helium-neon (He-Ne) laser against the damage of human erythrocytes in whole blood was examined in a perfusion model using an artificial heart-lung machine. Preserved human whole blood was diluted and perfused in 2 closed circuits with a double roller pump. The laser irradiated one of the circuits (laser group), and none the other (control group). In the laser group, erythrocyte deformability and erythrocyte adenosine triphosphate (ATP) levels were significantly higher, and free hemoglobin levels were significantly lower than those in the control group. Subsequent morphological findings by means of scanning electron microscope were consistent with these results. Low power He-Ne laser protected human erythrocytes in the preserved diluted whole blood from the damage caused by experimental artificial heart-lung machines. The clinical application of low power laser treatment for extracorporeal circulation is suggested. PMID- 11119075 TI - Scanning electron microscopic analysis of arterial line filters used in cardiopulmonary bypass. AB - The clinical value of arterial line filters is still a controversial issue. Proponents of arterial line filtration argue that filters remove particulate matter and undissolved gas from circulation while opponents argue the absence of conclusive clinical data. We conducted scanning electron microscope (SEM) studies of arterial line filters used clinically in the cardiopulmonary bypass circuits during adult cardiac surgery and analyzed the types and characteristics of materials entrapped in the arterial line filters. Twelve arterial line filters were obtained during routine hypothermic cardiopulmonary bypass in 12 adult cardiac patients. The arterial line filter was a screen type with a pore size of 40 microm (Baxter Health Care Corporation, Bentley Division, Irvine, CA, U.S.A. ). After opening the housing, the woven polyester strands were examined with SEM. All segments examined (120 segments, each 2.5 x 2. 5 cm) contained no embolic particles larger in their cross-sectional area than the pore size of the filter (40 microm). The origins of embolic particulates were mostly from environmental foreign bodies. This may suggest a possible need for more aggressive filtration of smaller particulates than is generally carried out at the present time. PMID- 11119076 TI - Selection of a polyurethane membrane for the manufacture of ventricles for a totally implantable artificial heart: blood compatibility and biocompatibility studies. AB - Membranes made from 4 commercial poly(carbonate urethanes): Carbothane (CB), Chronoflex (CF), Corethane 80A (CT80), and Corethane 55D (CT55), and from 2 poly(ether urethanes): Tecoflex (TF) and Tecothane (TT) were prepared by solution casting and sterilized by either ethylene oxide (EO) or gamma radiation. Their biocompatibility was evaluated in vitro in terms of proliferation, cell viability, and adhesion characteristics of human umbilical veins (HUVEC), monocytes (THP-1), and skin fibroblasts, and by measuring complement activation through the generation of the C3a complex. Their hemocompatibility was determined by measuring the level of radiolabeled platelet, neutrophil, and fibrin adhesion in an ex vivo arteriovenous circuit study in piglets as well as via an in vitro hemolysis test. The results of this study showed no endothelial cell proliferation on any of the materials. The cell viability study revealed that the CB, CF, and TF membranes sterilized by EO maintained the highest percentage of monocyte viability after 72 h of incubation (>70%) while none of the gamma sterilized membranes displayed any cell viability. The fibroblast adhesion and C3a generation assays revealed that none of the materials supported any cell adhesion or activated complement, regardless of the sterilization method. The hemolysis test also confirmed that the 4 poly(carbonate urethanes) were hemolytic while none of the poly(ether urethanes) were. Finally, the ex vivo study revealed that significantly more platelets adhered to the CB and CT55 membranes while the levels of neutrophil and fibrin deposition were observed to be similar for all 6 materials. In conclusion, the study identified the CF and TF membranes as having superior biocompatibility and hemocompatibility compared to the other polyurethanes. PMID- 11119077 TI - Platelet activation in the gyro C1E3 centrifugal pump: comparison with the terumo capiox and the Nikkiso HPM-15. AB - The Gyro C1E3 was developed as a cardiopulmonary bypass pump incorporating the sealless double pivot bearing system. In this study, we evaluated platelet activation induced by the Gyro C1E3 in vitro and in comparison to that of other centrifugal pumps. Rates of increase (RI) for beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) in the Gyro C1E3 were calculated from in vitro data and compared with the rate of increase in the Capiox (Terumo) and HPM-15 (Nikkiso) pumps. Fresh human blood was used, and a flow of 5.0 L/min with a DeltaP (pressure difference between the outlet and inlet of the pump) of 100 mm Hg employed. RI = Deltabeta-TG/DeltaN and DeltaPF4/DeltaN were used where Deltabeta TG is the increase in beta-TG, DeltaPF4 is the increase in PF4, and DeltaN is the increase in the passing number and where N = Qt/V (t = time, V = priming volume, and Q = flow rate). The mean RI for beta-TG was 0.26 +/- 0. 05 in the Gyro C1E3, 0.20 +/- 0.07 in the Capiox, and 0.15 +/- 0.02 in the HPM-15. The mean RI for PF4 was 0.15 +/- 0.03 in the Gyro C1E3, 0.12 +/- 0.05 in the Capiox, and 0.09 +/- 0.04 in the HPM-15. While there was no difference in RI for beta-TG and PF4 between the Gyro C1E3 and Capiox, RI for beta-TG and PF4 were significantly higher in the Gyro C1E3 than in the HPM-15 (p = 0.006 and 0.029). In vitro evaluation using RI for beta-TG and PF4 showed platelet damage caused by the Gyro C1E3 and the Capiox to be nearly equal while the HPM-15 was less traumatic to platelets than the Gyro C1E3. PMID- 11119078 TI - The effect of sudden failure of a rotary blood pump on left ventricular performance in normal and failing hearts. AB - We investigated the hemodynamic effect of regurgitation (or back-flow) due to sudden failure of a rotary blood pump (diagonal pump). Seven healthy sheep (Group C) and 7 with chronic heart failure (Group F) were studied. Chronic heart failure was obtained by intracoronary injection of microspheres several weeks earlier. Left ventricular function and ventricular efficacy were assessed by the pressure volume relationship. The back-flow through the stopped pump was significantly lower in Group F (2.3 +/- 0.34 L/min) than in Group C (2.8 +/- 0.33 L/min). Mean aortic blood pressure dropped significantly from 68.3 +/- 9.65 to 61.9 +/- 9.75 mm Hg in Group C and from 62.5 +/- 9.12 to 51.5 +/- 9.08 in Group F but remained stable during the 15 min period of pump stop. Parameters of left ventricular contractility (preload recruitable stroke work) dropped significantly in both groups, remained stable during the pump stop, and returned to baseline values 30 min after the end of back-flow. The ventricular efficacy (in terms of energy transfer) was tolerant against this acute volume overload even in the failing hearts. Sudden pump failure of a rotary blood pump leads to an acute depression of the hemodynamic state and myocardial contractility. However, this depression remained stable over 15 min, did not lead to further deterioration of the animals, and was completely reversible. PMID- 11119079 TI - Perfusate lactate dehydrogenase level and intrarenal resistance could not be adequate markers of perfusion quality during isolated kidney perfusion. AB - The main goal of this work was to study the influence of perfusion pressure and flow waveform during kidney perfusion, and the relationship between renal vascular resistance (RVR) and lactate dehydrogenase (LDH) concentration in the perfusate. Simultaneous constant pressure kidney perfusions were performed with either pulsatile or continuous flow at either 30 or 80 mm Hg of constant perfusion pressure. Mean flow, pressure, and RVR were displayed online during perfusion. Perfusate samples for LDH, creatine phosphatase kinase (CPK), and alkaline phosphatase (AP) determinations were taken. At the end of the perfusion, 2 ml of Evans blue was injected into the circuit to obtain images of perfusate distribution, and the kidneys were weighed. Also, hematoxylin/eosine studies were performed, showing more Bowman's space and tubular dilation in kidneys perfused with high pressure. We did not find differences in RVR between kidneys perfused at 30 and 80 mm Hg; nevertheless, perfusate distribution was better in the 80 mm Hg perfusions. We did not find any correlation between enzyme release and RVR in kidneys perfused with different mean pressures. These findings suggest that vascular resistance and LDH concentration cannot be independently considered as adequate markers of perfusate distribution. PMID- 11119080 TI - Combination of inhaled nitric oxide therapy and inverse ratio ventilation in patients with sepsis-associated acute respiratory distress syndrome. AB - Inverse ratio ventilation (IRV) is a ventilatory technique that uses an inspiratory to expiratory ratio (I:E) greater than 1:1. We studied the effects of mechanical ventilation with an I:E of 1:3, 1:1, and 2:1 on arterial oxygenation in 10 patients with sepsis-associated acute respiratory distress syndrome (ARDS). At each I:E, patients received 0 and 4 ppm of inhaled nitric oxide (INO) in random order for 30 min. Respiratory and cardiovascular parameters were measured. Of the 10 patients studied, 7 responded to IRV and 3 did not. An increase in the I:E and the addition of INO significantly improved arterial oxygenation in the responders (p < 0. 0001 and p < 0.006, respectively). The combination of an increase in the I:E and INO had an additive effect on arterial oxygenation. The combined use of IRV and INO is a more effective method of avoiding hypoxemia than either INO or IRV alone. PMID- 11119081 TI - Comparative analysis of alpha-stat and pH-stat strategies with a membrane oxygenator during deep hypothermic circulatory arrest in young pigs. AB - Using young pigs, this study compared the strategies of alpha-stat and pH-stat during deep hypothermic circulatory arrest (DHCA) for the cooling time of brains during the induction of hypothermia and rewarming time with cardiopulmonary bypass (CPB); the cerebral perfusion rate and metabolism rate, and the ratio of these 2 rates; and the extent of the cerebral edema development after circulatory arrest. Fourteen young pigs were assigned to 1 of 2 strategies of gas management. Cerebral blood flow was measured with a cerebral venous outflow technique. With CPB, core cooling was initiated and continued until the nasopharyngeal temperature fell below 20 degrees C. The flow rate was set at 2,500 ml/min. Once the temperature reached below 20 degrees C, the animals were subjected to DHCA for 40 min. During the cooling period, the acid-base balance was maintained using either alpha-stat or pH-stat strategy. After DHCA, the body was rewarmed to the normal body temperature. The animals then were sacrificed, and we measured the brain water content. The cerebral perfusion and metabolism rates were measured before the onset of CPB, before cooling, before DHCA, 15 min after rewarming, and upon the completion of rewarming. The cooling time was significantly shorter with alpha-stat than with pH-stat strategy while no significant differences were observed in the rewarming time between groups. Also, no significant differences were found in cerebral blood flow volume, metabolic rate, or flow/metabolic rate ratio between groups. In each group, the cerebral blood flow volume, metabolic rate, and flow/metabolic rate ratio showed significant differences in body temperature. Brain water content showed no significant differences between the 2 groups. In summary, this study found no significant differences between alpha stat and pH-stat strategies, except in the cooling time. The cooling time was rather shorter with the alpha-stat than with the pH-stat strategy. PMID- 11119082 TI - Experimental use of a compact centrifugal pump and membrane oxygenator as a cardiopulmonary support system. AB - Compactness and high performance are the most important requirements for a cardiopulmonary support system. The Nikkiso (HPM-15) centrifugal pump is the smallest (priming volume; 25 ml, impeller diameter; 50 mm) in clinically available centrifugal pumps. The Kuraray Menox (AL-2000) membrane oxygenator, made of double-layer polyolefin hollow fiber, has a minimum priming volume (80 ml) and a low pressure loss (65 mm Hg at 2.0 L/min of blood flow) compared with other oxygenators. The aim of this study was to evaluate the performance of the most compact cardiopulmonary support system (total priming volume: 125 ml) in animal experiments. The cardiopulmonary bypass was constructed in a canine model with the Nikkiso pump and Menox oxygenator in comparison with a conventional cardiopulmonary support system. The partial cardiopulmonary bypass was performed for 4 h to evaluate the gas exchange ability, blood trauma, serum leakage, hemodynamics, and blood coagulative parameters. The postoperative plasma free hemoglobin level of the compact cardiopulmonary system was 29.5 +/- 10.21 mg/dl (mean +/- SD), which was lower than that of the conventional cardiopulmonary system, 48.75 +/- 27.39 mg/dl (mean +/- SD). This compact cardiopulmonary system provided the advantage in terms of reduction of the priming volume and less blood damage. These results suggested the possibility of miniaturization for the cardiopulmonary bypass support system in open-heart surgery in the near future. PMID- 11119083 TI - Development of an affordable diaphragmatic pump for cardiopulmonary bypass: an in vivo evaluation. AB - A new diaphragmatic pump (L-Y pump) and its drive unit were developed in our institute. The pump has a priming volume of 80 ml. The pump housing is 72 mm in diameter and 42 mm in height. Its total weight is 139 g. To assess and confirm the function and controllability of this pump, comparative studies of cardiopulmonary bypass (CPB) with L-Y pump (group A) and conventional roller pump (Group B) were performed using dogs. Both pumps provided pump flow of 90 to 100 ml/kg/min. The hemodynamics of both groups were stable and within the normal range. No leakage or thrombus formation was observed in the L-Y pump. All biochemistry data showed no significant differences between the 2 groups. This data demonstrated low plasma-free hemoglobin levels in the L-Y pump group; after 120 min of CPB, mean plasma free hemoglobin levels were 48.7 +/- 8.6 mg/dl in the roller pump group and 21.4 +/- 7.1 mg/dl in the L-Y pump group, and minimal hemolysis was indicated. In conclusion, this L-Y pump and its controller system might be useful for CPB in terms of its low hemolysis and good pump quality. This pump demonstrated easy manipulation, good controllability, and provided a sufficient pulsatile flow. This pump is suitable not only for CPB, but also as a long-term circulatory support system. PMID- 11119084 TI - The DALI system. PMID- 11119085 TI - The management of intractable haematuria. PMID- 11119086 TI - Magnetic resonance urography enhanced by gadolinium and diuretics: a comparison with conventional urography in diagnosing the cause of ureteric obstruction. AB - OBJECTIVE: To compare the ability of magnetic resonance urography (MRU), enhanced using gadolinium and frusemide diuresis, and conventional intravenous urography (IVU) to diagnose the cause of ureteric obstruction. PATIENTS AND METHODS: The study included 82 patients in whom IVU showed or suggested obstruction and who also underwent MRU. The images from both methods were interpreted by various investigators independently; two evaluated the IVU and two others the MRU, the latter being unaware of the diagnosis after IVU. If the diagnosis remained unclear, further investigations (e.g. computed tomography, retrograde pyelography or ureteroscopy) were conducted. RESULTS: The diagnoses were ureteric calculi in 72 patients, ureteric tumours in eight and extra-ureteric tumours in two. In those with urolithiasis, the diagnosis was correct with IVU in 49 patients and with MRU in 64. The diagnosis in this group was incorrect with MRU in only two patients. The main reason for the failure of IVU was absent contrast medium excretion. Three of eight patients with ureteric tumours were correctly diagnosed by IVU but in three patients the diagnosis was incorrect. MRU correctly diagnosed seven of the eight patients in this group, with no false diagnosis. CONCLUSION: IVU is currently likely to remain the standard procedure for imaging the upper urinary tract, but this study shows the potential of MRU when enhanced with gadolinium and frusemide. MRU may be helpful if there is a dilated system with no excretory function, in pregnant women, in children and in those with contrast medium allergy. PMID- 11119087 TI - Bacterial adherence to ofloxacin-blended polylactone-coated self-reinforced L lactic acid polymer urological stents. AB - OBJECTIVE: To determine whether ofloxacin coating has any effect on bacterial adherence to bioresorbable self-reinforced L-lactic acid polymer (SR-PLLA) urological stents. MATERIALS AND METHODS: SR-PLLA stents were coated with epsilon caprolactone/L-lactide copolymer blended with ofloxacin at three different concentrations of ofloxacin (0.5, 2 and 5% w/w). The adherence of five bacterial strains (Pseudomonas aeruginosa, Enterococcus faecalis, Proteus mirabilis and two strains of Escherichia coli) to the coated SR-PLLA stents was analysed. Uncoated stent pieces were used as controls. The effect of ofloxacin coating on bacterial growth in the microenvironment of the stent pieces was also analysed. RESULTS: Ofloxacin coating prevented bacterial adherence to SR-PLLA stent material; this effect correlated significantly with the ofloxacin concentration of the caprolactone coating. Ofloxacin coating reduced the amount of bacteria in the microenvironment of the stent, but because of natural resistance, ofloxacin coating had little effect on E. faecalis. CONCLUSION: Except for E. faecalis, ofloxacin coating may reduce stent-associated infections. However, further studies are needed to confirm its biocompatibility and efficacy in clinical use. PMID- 11119088 TI - A prospective evaluation of the pathogenesis of detrusor instability in women, using electron microscopy and immunohistochemistry. AB - OBJECTIVE: To characterize the types of detrusor smooth muscle junctions in the bladders of women with detrusor instability and in a control group without, and to assess whether there are differences in the cell junctions between these groups. PATIENTS AND METHODS: The study included 13 women with detrusor instability (median age 57 years, range 32-86) and 11 control women (median age 50 years, range 33-62). Bladder biopsies were taken from each participant, processed for electron microscopy and immunohistochemistry (using a labelled antibody to vinculin) and analysed by investigators who were unaware of the patients' diagnoses. RESULTS: Adherens (intermediate) junctions in classic and rudimentary forms were present in all biopsies from patients and controls. Adherens junctions and dense plaques occupied almost the complete cell border in most samples. Complete immunohistochemistry was possible in seven patients and five controls. In almost every detrusor smooth muscle cell studied, there was staining of the entire cell border with labelled antibody to vinculin in all biopsies. CONCLUSIONS: This study provides evidence against an ultrastructural basis for idiopathic detrusor instability based on possible differences in detrusor smooth muscle intercellular junctions. Virtually the entire cell membrane of detrusor smooth muscle fibres is occupied by adherens junctions in classic and rudimentary forms, and with dense plaques present in samples from women with an unstable bladder and from controls. There was no junction detected in those with instability that was not present in the control group. The adherens junctions in the bladder facilitate mechanical coupling between cells. PMID- 11119089 TI - Minimally invasive non-laser thermal techniques for prostatectomy: a systematic review. The ASERNIP-S review group. PMID- 11119090 TI - The control of haemolysis during transurethral resection of the prostate when water is used for irrigation: monitoring absorption by the ethanol method. AB - OBJECTIVE: To determine whether the addition of ethanol to water for irrigation during transurethral resection of the prostate (TURP) and monitoring breath ethanol could be used to detect irrigant absorption and to limit free plasma haemoglobin in cases of absorption. PATIENTS AND METHODS: One hundred patients (46 in Pitea, Sweden and 54 in Uong bi, Vietnam) underwent surgery for benign prostatic hyperplasia (BPH) under an intermittent irrigation technique using water containing 2% ethanol. An expired breath alcohol meter was used to monitor ethanol in the patients' breath every 5 min. Blood samples taken after TURP were assessed for free haemoglobin in 99 patients, and other markers of haemolysis were also evaluated in the Swedish group. RESULTS: Thirty-two patients had detectable ethanol in their breath. There was a close correlation between the maximum ethanol reading during surgery and the level of free plasma haemoglobin after TURP (r = 0.90, P < 0.001). There was no correlation between the duration of TURP and the free haemoglobin level. CONCLUSION: Monitoring breath ethanol during TURP assesses absorption and so can help to keep control of haemolysis. It is suggested that the value on the alcohol meter should not be allowed to exceed 0.15 (corresponding to a blood ethanol level of 0.15 per thousand), which should maintain the free plasma haemoglobin level at < 1.0 g/L after TURP. Restricting the operative duration per se is not a reliable safety measure. PMID- 11119091 TI - A randomized trial comparing bladder infusion with standard catheter removal after transurethral resection of the prostate. AB - OBJECTIVE: To examine the effect of bladder infusion before catheter removal on patients' readiness for discharge and the day of discharge after transurethral resection of the prostate (TURP). PATIENTS AND METHODS: The study comprised 75 consecutive patients undergoing TURP who were randomized to either have their catheter removed in the standard manner (38 patients), or to undergo bladder infusion before a trial of voiding (ToV) on the second day after TURP (37 patients). RESULTS: In those undergoing bladder infusion, seven (19%) patients were discharged on the same day as their ToV, compared with five (13%) in the standard group. Of the 75 patients, 15 (68%) were discharged by the third day after TURP whether or not the bladder had been filled. In the infusion group, 23 (62%) were ready for discharge on the same day as their TOV, compared with only 14 (37%) in the standard group (P < 0.05). CONCLUSION: Bladder infusion before a ToV after TURP significantly increases the rate of readiness for discharge, allowing an early decision to discharge on the second day in a large proportion of patients. PMID- 11119092 TI - Urological cancers: do early detection strategies exist? AB - OBJECTIVE: To determine the perceptions of healthcare professionals and the general public about the symptoms, diagnosis and available treatment of urological cancers, and thus the perceived value of screening for their early detection. SUBJECTS AND METHODS: Two questionnaires were developed, based on semi structured interviews, and distributed to 288 healthcare professionals, comprising 182 general practitioners (GPs), 56 practice nurses and 50 urology nurses, and to 250 members of the general public in three different socio economic groups. The questionnaires asked about the symptoms, diagnosis and treatment of prostate, bladder and testicular cancer, and whether the respondents considered that screening for these cancers would be useful in ameliorating morbidity or death from these diseases. RESULTS: The response rate was very poor (13 of the GPs, 7%; 34 of the nurses, 32%; and 58 members of the general public, 23%). This severely limited the interpretation of the results, but the responses highlighted areas which need addressing. Obvious symptoms were well understood by all the groups but less well-known symptoms could be missed. No GP supported screening for prostate cancer and only seven of the GPs believed in teaching testicular self-examination, but practice nurses were considerably more active in all aspects of patient education. The general public felt that they needed more information to make decisions about urological cancers, although there was a general feeling that 'screening saved lives'. CONCLUSION: This survey showed that no healthcare professional seems to have a clearly defined role in informing potential patients about screening. The general public, who seem to perceive from the media that early detection is beneficial, are confused by the lack of clarity about policies for urological cancers. PMID- 11119093 TI - Androgen receptor signalling in the prostate. PMID- 11119094 TI - Treatment options after failure of local curative treatments in prostate cancer: a controversial issue. PMID- 11119095 TI - Detecting circulating prostate cells in patients with clinically localized prostate cancer: clinical implications for molecular staging. AB - OBJECTIVE: To evaluate the clinical utility of using the reverse transcriptase polymerase chain reaction (RT-PCR) to detect prostate-specific antigen (PSA) mRNA in peripheral blood samples from patients with prostate cancer, as a predictor of extraprostatic disease, and to assess any correlations with known predictive markers of this condition. PATIENTS AND METHODS: Immediately before radical prostatectomy, peripheral blood samples were taken from 25 men with clinically localized prostate cancer and analysed for PSA mRNA using RT-PCR (in 'hot-start' conditions and confirmed using ClaI restriction enzyme). The relationships between PSA mRNA positivity, pathological and clinical features were analysed; PSA mRNA positivity, PSA level and biopsy Gleason score were then compared as predictors of extraprostatic disease. RESULTS: There was no relationship between PSA mRNA positivity and pathological stage (pT2 or pT3), and no association between PSA mRNA positivity and serum PSA level, PSA density, the findings on a digital rectal examination or transrectal ultrasonography, and perineural invasion in the prostatic biopsy. However, there was a significant correlation between the Gleason score of the preoperative biopsy and PSA mRNA positivity. The best predictors of extraprostatic disease were the biopsy Gleason score and the PSA level. CONCLUSION: There was no significant advantage in using the RT-PCR assay of PSA mRNA before surgery to stage prostate cancer and to discriminate between organ-confined and extraprostatic neoplasms. PMID- 11119096 TI - Penile lengthening. AB - OBJECTIVE: To describe a technique for penile lengthening and the results achieved. PATIENTS AND METHODS: The penis is completely disassembled into its anatomical parts; the glans cap remains attached dorsally to the neurovascular bundle and ventrally to the urethra and corporal bodies. A space is created between glans cap and the tip of corpora cavernosa; this space is used to insert autologous cartilage previously harvested from the rib, the space being measured beforehand when the corpora cavernosa are erect. The anatomical entities and inserted cartilage are joined together to form a longer penis. The increased length of the penis depends directly on the elasticity of the urethra and especially of the neurovascular bundle. From June 1995 to March 1999 the technique was applied in 19 patients aged 18-52 years, who were followed for a mean (range) of 3.3 (1-4.5) years. RESULTS: The increase in penile length was moderate, at 2-4 cm; there were no injuries of the neurovascular bundle or urethra, and no erectile dysfunction. Fifteen patients reported painless sexual intercourse, the remaining four patients providing no data. During the follow-up the cartilage insert remained at about the same size as that at initial implantation. CONCLUSION: The penile disassembly technique combined with the interposition of rib cartilage in the space between the glans cap and tips of the corpora cavernosa provides a genuine increase in penile length, with satisfactory results. PMID- 11119097 TI - The correction of penile curvature with the Essed-Schroder technique: a long-term follow-up assessing functional aspects and quality of life. AB - OBJECTIVES: To investigate, in a retrospective analysis using a detailed questionnaire, the long-term functional results in and quality of life (QoL) of patients after undergoing the Essed-Schroder procedure, a standard technique for correcting penile curvature. PATIENTS AND METHODS: From 1994 to 1999, 40 patients (median age 24 years) had their penile curvature corrected using a modified Essed Schroder technique. Assessments by the investigators and a self-completed questionnaire were used to evaluate the functional and cosmetic aspects of the procedure, and QoL issues. RESULTS: Complete follow-up data were available in 31 of the 40 (78%) patients (19 with congenital curvature and 12 with Peyronie's disease). The median follow-up was 22 months. The degree of penile angulation before surgery was estimated as < 45 degrees in five patients, 45-90 degrees in 22 and > 90 degrees in four. In 21 patients (68%) sexual intercourse was uncomfortable or impossible; 26 (84%) reported an impaired QoL because of the penile curvature. After surgery the cosmetic and functional result was good or sufficient in 25 patients (81%); all 25 were able to have sexual intercourse with no problems. Penile shortening (> 2 cm) was reported by six patients. A significant improvement in QoL was reported by 15 patients (48%), but of the 12 patients with Peyronie's disease before surgery, six reported impaired rigidity and two recurrence of their penile curvature afterward. Whereas only seven of 12 patients with Peyronie's disease reported good functional results, 18 of the 19 with congenital curvature reported good or excellent results after surgery. CONCLUSION: The Essed-Schroder method is a simple operation which provides good functional and cosmetic results. Patients with congenital curvature of the penis have better results than those with Peyronie's disease. PMID- 11119098 TI - Q-flap reconstruction of panurethral strictures. AB - OBJECTIVE: To review our experience with an extended Q-shaped penile skin flap for the reconstruction of panurethral strictures. PATIENTS AND METHODS: Between 1991 and 1999, 15 men with extensive strictures underwent a single-stage urethral reconstruction with a distal circumferential penile skin flap incorporating a ventral midline extension (Q-flap). None had undergone previous urethroplasty nor had any been circumcised. RESULTS: The Q-flap provided a pedicled strip of penile skin with a mean (range) length of 17 (15-24) cm; no additional graft materials were necessary. Excellent results were obtained in 10 patients; in the remainder, complications included recurrent stricture (in two) and (in one patient each) a cerebral vascular accident, urethrocutaneous fistula, meatal stenosis, femoral neuropathy and prolonged catheterization for focal extravasation. CONCLUSION: The Q-flap provides an abundant hairless penile skin flap that enables single-stage panurethral reconstruction while eliminating the additional time and morbidity of harvesting further grafts. PMID- 11119099 TI - Population-based outcomes after 28,246 in-hospital vasectomies and 1,902 vasovasostomies in Western Australia. AB - OBJECTIVES: To examine trends in vasectomy and vasovasostomy, and the surgical complications and factors associated with reversal after vasectomy, and paternity after vasovasostomy. PATIENTS AND METHODS: Procedure rates were estimated from 1980 to 1996 in the population of Western Australia. Linked hospital morbidity records were used in the follow-up of men after vasectomy to estimate the risks of complications and reversals. Records of vasovasostomies were linked to the paternity field on birth registrations. Independent effects of the study factors were examined using Cox regression. RESULTS: There was little net change in vasectomy rates, whereas vasovasostomy rates increased in men aged 30-49 years. Risks of surgical complications were low and decreased for vasovasostomy. At 12 15 years after vasectomy, the risk of reversal levelled at 2. 4% in the total cohort and at 11.1% in men aged 20-24 years. The risk of vasovasostomy was 69% greater after vasectomy performed in 1994-96 than in 1980-84 (P = 0.011). The factors strongly associated with reversal were age < 30 years and being single, divorced or separated at the time of vasectomy. Paternity was achieved after an estimated 53% of vasovasostomies. Successful reversal was more likely if the man was younger at vasectomy and the time elapsed was comparatively short. Compared with vasovasostomies performed in 1980-84, the success rate of those in 1994-96 was almost four times higher. CONCLUSION: Population rates of vasectomy are stable but the risk of seeking a reversal has increased. Outcomes after vasovasostomy have improved. Care should be taken during the counselling of men before vasectomy, and especially in those aged <30 years. PMID- 11119100 TI - Tubularized incised plate urethroplasty for proximal hypospadias. AB - OBJECTIVE: To review our experience of using the tubularized incised plate (TIP) urethroplasty (useful in the treatment of distal hypospadias) to treat proximal hypospadias. MATERIALS AND METHODS: From March 1997 to March 2000 primary repairs were carried out on 40 boys (mean age 4.5 years) with proximal hypospadias. After degloving the penile skin the meatus was at the mid-shaft in 10 boys, at the proximal penile shaft in 11, at the penoscrotal junction in 16, at the scrotum in two and at the perineum in one. The 21 patients with a mid or proximal shaft meatus were categorized as having mid-shaft and the other 19 as having posterior hypospadias. Tunica albuginea plication (TAP) was used to correct residual ventral curvature. The method of urethroplasty was adapted from that described by Snodgrass. The key step of the TIP repair is a midline incision of the urethral plate; a subcutaneous tissue flap dissected from the inner prepuce is used to cover the neourethra. An 8 or 10 F nasogastric tube is used as a urethral stent and removed 7 or 8 days after surgery. Follow-up endoscopy and urethral sounding were carried out in 17 of the patients aged < 6 years; the mean follow-up was 12.5 months. RESULTS: TAP was used to correct penile curvature in nine (23%) of the patients. Excluding stenosis, the TIP repair was successful in 20 (90%) of those with mid-shaft and in 16 of the 19 with posterior hypospadias; for all complications the respective rates were 19 of 22 and 15 of 19. The overall success rate was 88% for all 40 patients with proximal hypospadias; a urethrocutaneous fistula occurred in two of those with mid-shaft and three of those with posterior hypospadias. Urethral meatal stenosis occurred in four (12%) of the patients (two in each group); two were associated with a fistula and the other two had only mild meatal stenosis. The overall complication rate was 17.5% (three and four in the mid and the posterior hypospadias groups, respectively). The meatal stenosis was managed by simple dilatation in three and meatoplasty in one patient. Endoscopically, the mucosa of neourethra was pink and smooth in all 17 patients assessed. The calibre of all 17 neourethra was > or = 8 F and in 13 was > or = 10 F. CONCLUSION: TIP repair is a reliable method for treating both mid-shaft and posterior hypospadias. PMID- 11119101 TI - The development of a pig model to study fetal vesico-ureteric reflux. AB - OBJECTIVE: To develop a surgical protocol to induce vesico-ureteric reflux (VUR) in utero by ablating the ureteric tunnel in a fetal pig model. MATERIALS AND METHODS: Fetal surgery was conducted on nine sows, which were divided into three groups according to changes in the surgical protocol. Sows in groups 2 and 3 received different anaesthetics and antibiotics, and the operating theatre temperature was increased. In all cases, the intramural part of the ureter was unroofed in the fetuses, which were then returned to the uterus. Upon delivery, cystograms were taken in the male piglets, and the urinary tracts removed for anatomical and histological examination. RESULTS: All three sows in group 1 delivered healthy piglets, but the fetuses that had undergone surgery were mummified. In group 2 the animals survived the fetal intervention, as shown by ultrasonography after surgery, but the four sows aborted spontaneously within a week. In group 3, both sows delivered normally developed piglets, three of which had undergone ablation of the ureteric tunnel. VUR was present only in those renal units in which the ureteric tunnel was ablated, and this was associated with hydronephrosis, dilatation of the ureters and thinning of the renal parenchyma on gross pathological examination. CONCLUSIONS: The fetal pig model of VUR not only appears to be feasible, but with similarities in renal anatomy and physiology also seems to be ideal for investigating fetal VUR. PMID- 11119102 TI - Permeability of intestinal mucosa from urinary reservoirs in man and rat. AB - OBJECTIVE: To evaluate the barrier properties of intestinal mucosa chronically exposed to urine and to evaluate possible differences between ileal and colonic segments used in the reconstruction of the urinary tract. MATERIALS AND METHODS: Mucosal specimens from patients with continent reservoirs with an abdominal stoma, or orthotopic neobladders constructed from colonic segments, were obtained at revisional surgery. Control segments were obtained during right-sided hemicolectomy. In addition, ileal and colonic segments from enterocystoplasties in rats were assessed. The mucosa-to-serosa passage of marker molecules, i.e. (14)C-mannitol, (3)H-glucose, fluorescein isothiocyanate-dextran 4400 and ovalbumin, was measured using modified Ussing diffusion chambers. RESULTS: In man, there were no permeability differences between segments exposed to urine and control segments for any of the marker molecules. In rats, there was less passage of markers in ileal and colonic transplanted segments than in intestinal segments from sham-operated animals. CONCLUSIONS: Intestinal mucosa that has been in chronic contact with urine maintains its barrier function; in the rat model the permeability was even decreased. In addition, there were no detectable differences between ileal and colonic segments in this model. PMID- 11119103 TI - A novel antireflux uretero-ileal anastomosis for urinary diversion: an experimental study. AB - OBJECTIVES: To assess a new antireflux valve technique in a dog model of urinary diversion, and thus provide a reliable and easily constructed antireflux system for an ileal reservoir. MATERIALS AND METHODS: In five female beagle dogs, 3 cm of ileum were intussuscepted into the reservoir formed using the adjacent 10 cm of ileum. The intussuscepted ileum was sutured to the reservoir wall after stripping the mucosa of the reservoir in a trapezoidal zone opposite the similarly stripped mucosa of the intussuscepted ileum, to avoid dessusception. After the distal 4 cm of the ureters was united and pulled through the intussuscepted segment of ileum, the combined distal ureter was sutured to the labial edge of the intussuscepted segment. Finally, the reservoir was anastomosed to the bladder as an enterocystoplasty. Dogs were evaluated by ascending cystography and intravenous pyelography at 1 and 6 months. The valve and upper urinary tract were evaluated histopathologically at 6 months. RESULTS: In all dogs the antireflux system remained intact and prevented reflux. The intussuscepted ileum was firmly attached to the reservoir wall and the submucosa of each segment was united. The upper urinary tract was normal with no ureteric stenosis. Histopathologically, the ureter was surrounded by intact ileal serosa and showed no inflammation or scarring. CONCLUSIONS: The very short ileal segment required and the firm attachment of the constructed valve to the reservoir were the advantages of this practical and reliable new antireflux system. PMID- 11119104 TI - Electrophysiological recordings from the rat prostate gland in vitro: identified single-cell and transepithelial (lumen) potentials. AB - OBJECTIVE: To develop a preparation for the in vitro maintenance of the rat prostate gland and thus allow intracellular and transepithelial voltage measurements. MATERIALS AND METHODS: Ventral prostate glands from male rats were dissected free of connective tissue, separated into smaller lobes and maintained in vitro at 30 degrees C. Voltages were recorded with sharp micropipettes in identified cellular and luminal compartments, differentiated by several electrophysiological and histological parameters, including intracellular staining. RESULTS: Intracellular epithelial membrane potentials (median -40 mV) and transepithelial or luminal potentials (mean -4.2 mV) were recorded successfully. Luminal epithelial cells were dye-coupled. Prostate tissue could be maintained in vitro with no apparent electrophysiological or structural deterioration for up to approximately 7 h. CONCLUSION: Rat prostate tissue can be successfully maintained in vitro and electrophysiological recordings made from identified cellular compartments. PMID- 11119105 TI - Biomarker distribution after injection into the canine prostate: implications for gene therapy. AB - OBJECTIVE: To evaluate the distribution of biomarkers after transrectal injection into the canine prostate and to report a method for enhancing the distribution of gene expression. MATERIALS AND METHODS: Carbon black was first used to evaluate the histopathological distribution in canine prostate of single or multiple injections via the transurethral, transperineal and transrectal routes. The distribution of canarypox virus (ALVAC) vector-delivered gene expression was then compared using both fluid-phase injection techniques and delivery in a solid carrier composed of a gelatine sponge matrix. RESULTS: After transurethral administration, carbon black was detected as scattered particles in ducts and acini, mostly in the periphery of the gland. Direct transrectal injection of carbon black resulted in a localized collection at the site of injection, with only a minimal peri-acinar distribution. Transrectal injection of the fluid-phase (virus suspended in diluent) ALVAC vector encoding the beta-galactosidase gene resulted in a similar distribution, with limited gene expression at the site of injection and in the needle track. Delivery of the same number of virus particles in the gelatine sponge matrix resulted in qualitatively greater gene expression. CONCLUSIONS: Direct injection of the canine prostate with biomarkers, including viral vectors, in the fluid-phase results in very localized gene expression, while the distribution was more widespread after delivery in a gelatine sponge matrix. PMID- 11119106 TI - A comparison of the histopathological findings after open and percutaneous needle testicular biopsy in adult male rats. AB - OBJECTIVE: To define the histopathological changes occurring as a result of open and percutaneous needle testicular biopsy in adult rats. MATERIALS AND METHODS: Percutaneous needle and open testicular biopsies were taken from 35 male albino rats (120-140 days old). Nine of the rats were killed 10 days after the biopsy, eight after 30 days and the other eight 60 days after the biopsy. A control group of six rats underwent orchidectomy with no preceding testicular biopsy. RESULTS: Significant tubulitis and obstructive findings were detected 10 days after needle biopsy (P < 0.05); epididymo-orchitis was frequent after open biopsy during this period. At 10-30 days after needle biopsy the mean seminiferous tubule diameters were significantly greater than in either the control group or after open biopsy (P < 0.05). The histopathological damage recovered 60 days after open and needle biopsy. CONCLUSION: Although percutaneous, a needle biopsy (as an alternative to open biopsy) causes tubulitis and frequent obstructive findings in the early period; therefore, repeat testicular procedures should be planned after these changes have resolved. PMID- 11119107 TI - Laparoscopically-assisted percutaneous nephrolithotomy for the treatment of anterior calyceal diverticula. PMID- 11119108 TI - Spread of silicone to inguinal lymph nodes from a leaking testicular prosthesis: a cause for chronic fatigue? PMID- 11119109 TI - Prostatic small-cell neuroendocrine carcinoma with disease progression monitored by measurement of serum progastrin-releasing peptide. PMID- 11119110 TI - Re: Is only meatoplasty a legitimate surgical solution for extremedistal hypospadias? A long-term follow-up after adolescence. PMID- 11119112 TI - Re: Extravesical transitional cell carcinoma as a result of implantation after perforation of the bladder. PMID- 11119115 TI - Re: Are dedicated bladder films necessary as part of intravenous urography for haematuria. PMID- 11119116 TI - Re: A prospective pilot evaluation of urinary and immunohistochemical markers as predictors of clinical stage of urothelial carcinoma of the bladder. PMID- 11119117 TI - Re: Ethanol-glycine irrigating fluid for transurethral resection of the prostate in practice. PMID- 11119119 TI - Recent advances in the aetiology of cryptogenic fibrosing alveolitis. AB - Crytogenic fibrosing alveolitis is the commonest intersititial lung disease but, until recently, very little has been known about its aetiology. The histopathologist usually sees this disease at transbronchial biopsy or at autopsy. This article reviews the current knowledge of the aetiology of cryptogenic fibrosing alveolitis looking at possible infective, occupational, drug-related, smoking-associated, genetic and dietary factors. Knowledge of the possible roles of these factors in the disease process informs histopathologists when they are reporting these biopsies and enables them to make a larger contribution to defining the pathogenetic mechanisms. PMID- 11119120 TI - Architectural remodelling in acute and chronic interstitial lung disease: fibrosis or fibroelastosis? AB - AIMS: Pulmonary fibrosis in acute and chronic lung disease has been much investigated, but little attention has been directed at the elastic tissue in these situations. Our aim was to verify whether elastic deposition accompanies collagen deposition in the repairing process of acute and chronic lung injury. METHODS AND RESULTS: We measured, by image analysis, the content of fibres of the collagenous and elastic systems of the alveolar septum in histological slides sampled from autopsied lungs, using the picrosirius-polarization method and Weigert's resorcin-fuchsin stain, respectively. Five groups were studied: I, 10 normal patients; II, 10 patients with cardiogenic pulmonary oedema; III, 23 adult respiratory distress syndrome (ARDS) patients in the early phase; IV, 14 ARDS patients in the late fibroproliferative phase; and V, 10 idiopathic pulmonary fibrosis patients. The first two groups were used as controls. The content of fibres of the collagenous and elastic systems was significantly increased in groups IV and V as compared to the other groups. CONCLUSIONS: Our results indicate that deposition of elastic system fibres is present in the fibroproliferative phase of ARDS and in usual interstitial pneumonia and suggest that this event may contribute to the alveolar mechanical dysfunction and remodelling that occur in acute and chronic interstitial lung disease. PMID- 11119121 TI - Sauropus androgynus-constrictive obliterative bronchitis/bronchiolitis- histopathological study of pneumonectomy and biopsy specimens with emphasis on the inflammatory process and disease progression. AB - AIMS: The histopathology of the Sauropus androgynus (SA)-constrictive bronchiolitis obliterans (BO) is still controversial. A recent report using pneumonectomy specimens showed that the major histopathology was obliterative arteriopathy with segmental necrosis of small bronchi instead of constrictive BO as previously described. METHODS AND RESULTS: We analysed semiquantitatively and immunohistochemically the histopathology of one pneumonectomy and four biopsies specimens of SA-associated lung disease. We found a significant number of constrictive and obliterative bronchioles 1 mm or less in diameter and segmental inflammatory destruction with complete luminal obliteration of the bronchi less than 3 mm in diameter in the pneumonectomy specimen (37% and 25%, respectively). Fibromuscular intimal sclerosis of the bronchial arteries was identified in 15% of the bronchi 4 mm or less in diameter. The inflammation in these airways was composed predominantly of T-lymphocytes, macrophages, mast cells and eosinophils. They were present throughout the evolutionary stages of the bronchiolitis ranging from early oedematous to the late fibrotic obliterative stage. Double immunohistochemical stains revealed negative proliferative cell nuclear antigen for most of the T-lymphocytes and macrophages but positive for fibroblasts. CONCLUSIONS: A more accurate histopathological designation of the SA-associated lung disease should be constrictive obliterative bronchitis/bronchiolitis, with the participation of T-lymphocytes, macrophages, mast cells, eosinophils and fibroblasts in its morphogenesis. The persistent accumulation of inflammatory cells was mediated predominantly by continued recruitment to the site of injury from the bloodstream, resulting eventually in the irreversible fibrosis of the bronchioles and the bronchi less than 3 mm in diameter. Obliterative arteriopathy is suspected of being only an indirect contributing factor. PMID- 11119122 TI - Comparative methodological analysis of erbB-2/HER-2 gene dosage, chromosomal copy number and protein overexpression in breast carcinoma tissues for diagnostic use. AB - AIMS: This study was performed to test the validity of different methods for determining the status of the erbB-2/HER-2 oncogene in breast cancer tissues for diagnostic use. METHODS AND RESULTS: Forty formalin-fixed, paraffin-embedded breast carcinomas were investigated by fluorescence in situ and comparative genomic hybridization (FISH, CGH) as well as by immunohistochemistry (IHC) using Dako-HercepTest and CB11 antibody (Ventana). Additionally, competitive differential polymerase chain reaction (cdPCR) was performed on frozen samples to estimate gene dosage alterations of erbB-2/HER-2. Amplification was detected in 12-23% and protein overexpression in 16-68% of the cases, depending on the methodology and/or the reagent used. Perfect concordance (100%) was found between the results of cdPCR and CB11-IHC, and a 97% concordance between FISH and CB11 IHC. The concordance between Dako-HercepTest and CB11-IHC was 78%: seven of eight 2+ carcinomas with the Dako-HercepTest were classified as nonamplified using FISH. CONCLUSIONS: Our results indicate that high-level expression as well as normal erbB-2/HER-2 status of breast carcinomas can be detected reliably both by IHC and gene dosage assessment in paraffin material for diagnostic use. However, borderline results, especially those with 2 + immunopositivity, should be interpreted with caution and increased emphasis should be given to other clinical and prognostic information available. PMID- 11119123 TI - Clinical drug-resistant nodular sclerosing Hodgkin's lymphoma is associated with decreased bcl-2 expression in the surrounding lymphocytes and with increased bcl 2 expression in the Reed-Sternberg cells. AB - Identification of patients with Hodgkin's lymphoma who are primary refractory to chemotherapy will have great impact on treatment planning. Several studies have indicated that up-regulation of the bcl-2 proto-oncogene expression in non Hodgkin's lymphoma can cause resistance to chemotherapeutic drugs. For this reason we investigated the relationship between expression of bcl-2, the pro apoptotic protein Bax and MIB-1 with clinical drug-resistance in Hodgkin's lymphoma. METHODS AND RESULTS: Seven patients with nodular sclerosis Hodgkin's lymphoma under 40 years of age, who failed to achieve complete remission upon primary chemotherapy and 10 matched patients who did achieve complete response were selected from the Eindhoven Cancer Registry. Tissue sections from both groups of patients were stained for bcl-2, Bax and MIB-1. In the non-responding patients Reed-Sternberg cells expressed bcl-2 more frequently using a cut-off point of <25% to indicate negative cells (P = 0.04), whereas no differences were observed in the expression of Bax and MIB-1. In non-responding patients the lymphocytes surrounding Reed-Sternberg cells expressed bcl-2 less frequently (P = 0.002), using a cut-off point of < 25%, whereas no difference was observed in the expression of Bax and MIB-1. CONCLUSION: A high percentage of Reed-Sternberg cells expressing bcl-2 protein and a low expression of bcl-2 proteifi in the surrounding lymphocytes is associated with treatment-failure, and subsequent poor survival in young patients with nodular sclerosing Hodgkin's lymphoma. These results need further validation in larger studies. PMID- 11119124 TI - Prognostic factors in ovarian carcinosarcoma: a clinicopathological and immunohistochemical analysis of 23 cases. AB - Carcinosarcoma of the ovary is a rare, highly aggressive neoplasm comprising histologically of both epithelial and mesenchymal components. The aim of this study was to evaluate the clinicopathological prognostic factors in ovarian carcinosarcoma, including the immunohistochemical expression of p53 protein and Ki67. METHODS AND RESULTS: Twenty-three cases of carcinosarcoma of the ovary were studied retrospectively. The clinicopathological and immunohistochemical parameters including p53 and Ki67 staining were statistically analysed to investigate the prognostic significance of this tumour. The overall 5-year survival rate was 27.1%; 100% for stage I, 31.3% for stage II, 10.9% for stage III and 0% for stage IV. The low-stage group (stages I and II) was found to be a significant prognostic factor for patient survival (P = 0.0113). None of the other factors (tumour size, histological type of carcinomatous and sarcomatous components, mitotic count, vascular space invasion and immunoreactivity for p53 protein and Ki6 7) was found to be a statistically significant prognostic indicator. CONCLUSIONS: Ovarian carcinosarcoma is a rare malignancy with poor prognosis. In this study, advanced stage appears to be poor prognostic indicator of survival in patients with ovarian carcinosarcoma. PMID- 11119125 TI - Pilocytic astrocytomas do not show most of the genetic changes commonly seen in diffuse astrocytomas. AB - AIMS: While it is well known that pilocytic astrocytomas are clinically distinct from diffuse astrocytomas, few comprehensive studies have focused on their genetic differences. The aim of this study was to examine pilocytic astrocytomas for genetic alterations that are commonly seen in diffuse astrocytomas. METHODS AND RESULTS: By using molecular genetic and immunohistochemical techniques, we evaluated p16, p53, CDK4 and PTEN genes in 29 pilocytic astrocytomas. Mutation screening of p53 and PTEN was performed by single strand conformation polymorphism analysis followed by direct sequencing. Loss of heterozygosity (LOH) of p53, p16 and 10q23-25 loci was performed with microsatellite markers and genomic microsatellite instability (MSI) was also screened. Protein expression of p16, p53, CDK4 and PTEN was examined by immunohistochemistry. Five tumours were found to have single genetic alterations, which included a p53 mutation, a PTEN mutation, MSI at a single microsatellite marker of the p16 locus, and one single LOH at each p16 and 10q23 loci. Protein expressions of p16, CDK4 and PTEN were detected in 73%, 61% and 38% of tumours, respectively. Significantly and in sharp contrast to diffuse astrocytomas, no pilocytic astrocytoma in our series stained for p53 protein. CONCLUSION: Pilocytic astrocytomas have neither MSI phenotype nor recurrent alterations of the p53 and p116 genes. However, altered expression of PTEN may be important in the genesis of pilocytic astrocytomas. We conclude that pilocytic astrocytomas are genetically distinct from diffuse astrocytomas. Lack of p53 mutation/immunostaining may serve as a diagnostic adjunct for differentiating pilocytic astrocytomas from diffuse astrocytomas in small neurosurgical biopsies. PMID- 11119126 TI - Trisomy 6 in Merkel cell carcinoma: a recurrent chromosomal aberration. AB - We retrospectively investigated 17 cases of primary and metastasizing Merkel cell carcinomas (MCC) from 14 patients using chromosomal in-situ hybridization (CISH) to study the occurrence of trisomy 6 in these lesions. METHODS AND RESULTS: Histological diagnosis on all tumour samples was obtained on haematoxylin and eosin stained sections. Immunohistochemistry was performed with antibodies against pancytokeratin (CAM 5.2), cytokeratin 20 (CK20), MIC2 antigen (CD99), neuron-specific enolase (NSE), and chromogranin A (chrA). Sections (4 microm) of the paraffin-embedded tumours were analysed with alpha-satellite centromeric probes for chromosome 6 or 17 using CISH. The signal was amplified by the Tyramide Signal Amplification (TSA) assay. Immunohistochemically, the tumours showed the same general epithelial neuro-endocrine pattern: 11/13 expressed cytokeratin 20, and 47% exhibited trisomy 6, with no significant difference between primary and metastatic lesions. Incomplete follow-up data did not allow us to establish a prognostic value of trisomy 6, however, this aberration might be an additional diagnostic tool in distinguishing MCC from other small round blue cell tumours. CONCLUSIONS: CISH seems to be a promising adjunctive method to diagnose Merkel cell carcinoma. Trisomy 6 should be investigated more closely in these cases, as has been done for chromosomes 1 and 11. Of particular interest would be identification of modifications in proto-oncogene(s) located on chromosome 6. PMID- 11119127 TI - Palisading and verocay body-prominent dermatofibrosarcoma protuberans: a report of three cases. AB - The aim of this report is to draw attention to nuclear palisading and Verocay body formation as peculiar, previously undescribed histological findings in rare instances of dermatofibrosarcoma protuberans (DFSP). METHODS AND RESULTS: Three indurated, nodular or plaque skin lesions were diagnosed as DFSP on the basis of their storiform proliferation of spindle-shaped cells diffusely infiltrating the dermis and subcutaneous tissue. Sclerosing and giant cell areas were also identified. Unexpectedly, conspicuous nuclear palisading was also noted in all cases and Verocay body formation was present in two. Immunostains were positive for CD34 and negative for S100 protein in every instance. Proliferating cells were seen to display fibroblast-like features by ultrastructural study of one case. CONCLUSIONS: DFSP may rarely show a schwannoid histological appearance as the result of nuclear palisading and even Verocay body formation. In this setting, both the search for DFSP characteristic morphologic features and the performance of CD34 and S100 protein immunohistochemistry will facilitate the correct diagnosis. PMID- 11119128 TI - Mammary carcinoma with prominent cytoplasmic lipofuscin granules mimicking melanocytic differentiation. AB - We report a case of mammary carcinoma with striking cytoplasmic pigmentation in a 60-year-old Japanese woman who presented with a self-evident nodular lesion in the left breast. METHODS AND RESULTS: After a fine needle aspiration revealing atypical clusters of cells, an excisional biopsy was performed. Histologically, a partially cystic 18 mm lesion containing a 5-mm mural nodule was present. The mural nodule and adjacent thickened epithelium were comprised of atypical cells focally invading into the cyst wall. Striking abundant granular brown pigment resembling melanin was present in some of the neoplastic cells. The differential diagnosis included metastatic melanoma and mammary carcinoma with melanocytic differentiation. After a series of special stains and immunohistochemical studies, the diagnosis of mammary carcinoma with extensive cytoplasmic lipofuscin pigment was rendered. CONCLUSION: Mammary carcinoma with lipofuscin pigment to the degree seen in this case which mimics melanocytic differentiation has not, to our knowledge, previously been documented. PMID- 11119129 TI - Consistency of histopathological reporting of laryngeal dysplasia. The Scottish Pathology Consistency Group. AB - AIMS: Clinical management of premalignant and malignant lesions of the larynx is dependent on histopathological evaluation. The Scottish Pathology Consistency Group assessed interobserver variation in the evaluation of laryngeal dysplasia. METHODS AND RESULTS: One hundred laryngeal biopsies ranging from normal to invasive carcinoma were assessed. The overall Kappa result of 0.32 was disappointing. However, agreement on those categories which dictate significantly different management was more favourable. The Kappa figure for mild dysplasia versus severe dysplasia/CIS was 0.7, the Kappa figure for mild dysplasia versus severe dysplasia/CIS and invasive carcinoma was 0.77. The Kappa figure for mild and moderate dysplasia versus severe dysplasia/ CIS and invasive carcinoma was 0.57. An attempt to use a two grade system gave a Kappa figure of 0.52. CONCLUSIONS: Our group had a satisfactory agreement on the distinction of mild from severe dysplasia and on microinvasive carcinoma without any discussion as to histopathological criteria to be used. Clinical management--review endoscopy, repeat cord stripping, radiotherapy and laryngectomy--is in general dependent on histological assessment. Thus the agreement on categories which underpin clinical management is reassuring. However, assessment of moderate dysplasia remains problematic. An attempt to utilize a two grade system--low grade from high grade dysplasia/CIS--may have merit. The implications of the terminology used must be agreed among pathologists and clinicians working closely within clinicopathological cancer groups. PMID- 11119130 TI - What criteria reliably distinguish melanoma from benign melanocytic lesions? AB - The differential diagnosis of melanocytic lesions is fraught with difficulty and a common source of litigation either if a lesion misreported as 'benign' recurs locally or re-presents with nodal metastases or if an atypical naevus is called 'malignant' leading to a cosmetically unsatisfactory wider resection, unwarranted anxiety about prognosis and adverse life insurance prospects. Several authors have claimed that there are valid morphological criteria which, alone or in combination, enable reliable distinction between benign and malignant melanocytic lesions. Others question these criteria and, doubting the extent to which unequivocal diagnoses can be rendered in all cases, believe that the diagnosis is purely subjective and that most diagnostic errors are non-negligent. To address these issues, expert opinions were commissioned from three sets of authors. Okun, Edelstein & Kasznica emphasize that a significant minority of melanocytic lesions are so borderline morphologically that diagnostic uncertainty is allowable and that such uncertainty can be handled responsibly. Kirkham, in favouring the methodical use of criteria, concedes that they are 'largely opinion-based rather than evidence-based, but do go beyond mere subjective pattern analysis'. In agreement with Okun and his colleagues. Slater emphasises that no single feature is reliable by itself and that all aspects, including clinical details, should be interpreted together; he has no hesitation in reporting the diagnosis as 'uncertain' in doubtful cases. In the absence of a specific marker pathognomonic of melanocytic malignancy, the diagnosis will continue to rely on the judicious application of morphological criteria with a small proportion of elusive cases in which diagnostic uncertainty should not be concealed. PMID- 11119131 TI - Optimal handling and criteria for melanoma diagnosis. PMID- 11119132 TI - Doubt and uncertainty in the diagnosis of melanoma. PMID- 11119133 TI - Author's reply. PMID- 11119135 TI - Authors' reply. PMID- 11119134 TI - Is mucinous differentiation of colorectal cancer a prognostic feature? PMID- 11119136 TI - Leiomyoma with atypical cells (atypical leiomyoma) in the larynx. PMID- 11119137 TI - Authors' reply. PMID- 11119138 TI - Immunohistochemical evidence for mesothelial origin of paratesticular adenomatoid tumour. PMID- 11119139 TI - Microinvasive lobular carcinoma of the breast. PMID- 11119140 TI - Author's reply. PMID- 11119141 TI - Tuberculoid granulomas and cervical granulomatous lymphadenitis. PMID- 11119142 TI - Simple self-contained desk-top image analysis for the non-specialist. PMID- 11119143 TI - Authors' reply. PMID- 11119144 TI - From this Month's Histopathology. PMID- 11119145 TI - Antimicrobial resistance profiling and DNA Amplification Fingerprinting (DAF) of thermophilic Campylobacter spp. in human, poultry and porcine samples from the Cork region of Ireland. AB - Antimicrobial resistance (R) typing and DNA Amplification Fingerprinting (DAF) of a random collection of 84 Irish thermophilic Campylobacter isolates is described. The collection included human, veterinary (porcine) and poultry isolates cultured between 1996 and 1998 in the Cork region of Ireland. Biochemical and molecular methods were used to identify Campylobacter jejuni and Camp. coli. Many of these isolates were simultaneously resistant to several common antimicrobial agents. In particular, resistance to ampicillin, spectinomycin, sulphafurazole and tetracycline was common. A total of 74 DAF profiles was identified among the study collection, showing a high degree of diversity. Dendrogram analysis of the DNA patterns identified three main clusters at the 50% similarity level, which included two clusters of Camp. coli and a third containing a mixture of Camp. jejuni and Camp. coli. PMID- 11119146 TI - Feather keratin hydrolysis by a Vibrio sp. strain kr2. AB - The aim of the study was to characterize feather-degrading bacteria isolated from poultry industry waste. A Vibrio sp. strain kr2 producing a high keratinolytic activity when cultured on native feather-containing broth was isolated. The bacterium grew with an optimum at pH 6.0 and 30 degrees C, where maximum featherdegrading activity was also observed. Keratinase production was similar at both 25 and 30 degrees C, while the maximum concentration of soluble protein was reached at 30 degrees C. Reduction of disulphide bridges was also observed, increasing with cultivation time. The keratinase of strain kr2 was active on azokeratin, azocasein, benzoyl-arginine-p-nitroanilide and Ala-Ala-p-nitroanilide as substrates. The amino acid composition of the feather hydrolysate was determined, presenting similarities with that reported for feather lysate, feather meal and raw feathers. A novel feather-degrading bacterium was isolated and characterized, showing high keratinolytic activity. Complete feather degradation was achieved during cultivation. Strain kr2 shows potential for use for biotechnological processes involving keratin hydrolysis. PMID- 11119147 TI - Diversity of phytoplasmas isolated from insects, determined by a DNA heteroduplex mobility assay and a length polymorphism of the 16S-23S rDNA spacer region analysis. AB - Two techniques were developed for the analysis of non-cultivable mollicutes in insects. The first was aimed at detecting organisms belonging to undiscovered groups within the phytoplasma clade. After prescreening by polymerase chain reaction with phytoplasma-specific primers, nucleic acids from 54 positive samples were amplified using phytoplasma-specific fluorescein-labelled primers flanking the 16S-23S rDNA spacer region, which is variable in length among the phytoplasmas. The sizes of all the detected products were only those expected for already-described phytoplasma subclades. It was also shown that a single leafhopper might carry different phytoplasmas, at similar or very different relative concentrations. The second technique, based on the heteroduplex mobility assay, was designed for the detection of organisms phylogenetically similar to phytoplasmas but not recognized by the specific primer pair. As a result, signals generated by ribosomal DNA of organisms which appear to be closely related but not identical to phytoplasmas were detected. PMID- 11119148 TI - Microbial differentiation and changes in susceptibility to antimicrobial agents. PMID- 11119149 TI - The inactivation of foot and mouth disease, Aujeszky's disease and classical swine fever viruses in pig slurry. AB - The aim of the study was to investigate the decontamination of pig slurry containing exotic viruses of pigs, foot AND mouth disease virus (FMDV), Aujeszky's disease virus (ADV) AND classical swine fever virus (CSFV). Laboratory scale decontamination experiments showed that FMDV, ADV and CSFV were heat inactivated in slurry within 3 min at 67 degrees C, 3 min at 62 degrees C and 3 min at 60 degrees C and in Glasgow Eagles medium within 5 min at 67 degrees C, 4 min at 65 degrees C and 2 min at 65 degrees C, respectively. At pilot scale, FMDV was heat inactivated at 66 degrees C in water and 61 degrees C in slurry, ADV at 61 degrees C in water or slurry and CSFV at 62 degrees C in water and 50 degrees C in slurry. Treatment of pig slurry for the inactivation of exotic viruses may be achieved through the use of a thermal pilot plant operating in continuous mode. The work demonstrates the suitability of thermal treatment in ensuring the safety of pig slurry following a disease outbreak. PMID- 11119150 TI - Microbial succession during a composting process as evaluated by denaturing gradient gel electrophoresis analysis. AB - Microbial succession during a laboratory-scale composting process of garbage was analysed by denaturing gradient gel electrophoresis (DGGE) combined with measurement of physicochemical parameters such as temperature, pH, organic acids, total dissolved organic carbon and water-soluble humic substance. From the temperature changes, a rapid increase from 25 to 58 degrees C and then a gradual decrease, four phases were recognized in the process as follows; mesophilic (S), thermophilic (T), cooling (C) and maturing (M). The polymerase chain reaction amplified 16S rDNA fragments with universal (907R) and eubacterial (341F with GC clamp) primers were subjected to DGGE analysis. Consequently, the DGGE band pattern changed during the composting process. The direct sequences from DGGE bands were related to those of known genera in the DNA database. The microbial succession determined by DGGE was summarized as follows: in the S phase some fermenting bacteria, such as lactobacillus, were present with the existing organic acids; in the T phase thermophilic bacillus appeared and, after the C phase, bacterial populations were more complex than in previous phases and the phylogenetic positions of those populations were relatively distant from strains so far in the DNA database. Thus, the DGGE method is useful to reveal microbial succession during a composting process. PMID- 11119151 TI - Effect of antibiotics on viability staining of Escherichia coli in solid phase cytometry. AB - Solid phase cytometry (SPC) has been investigated as a tool to assess the effect of antibiotics on the viability of Escherichia coli. After exposure of the cells to the antibiotic, they are retained on a polyester membrane filter and labelled using a fluorescein derivative as a substrate for intracellular esterases. The number of fluorescent bacteria is automatically counted in an Ar laser scanning device. In the presence of nutrients, all antibiotics tested in concentrations exceeding the MIC inhibited the multiplication of cells but not the labelling per se. However, when no nutrients were added, the cells did not multiply, and inhibition of the fluorescent staining was only observed for membrane permeabilizing antibiotics, even at sub-MIC concentrations. The selective detection by SPC of membrane-permeabilizing antibiotics corroborates the requirement of membrane integrity for viability labelling of bacteria. This selectivity has been exploited to develop a method for the detection of colistin residues in milk. PMID- 11119153 TI - Effect of protective agents, rehydration media and initial cell concentration on viability of Pantoea agglomerans strain CPA-2 subjected to freeze-drying. AB - The effect of initial cell density, protective agents and rehydration media on the viability of biocontrol agent Pantoea agglomerans CPA-2 when subjected to freeze-drying was studied. Several additives were tested as protective agents against freeze-drying injury. Maximum viability of the bacterial cells was obtained with disaccharides (survival levels > 60%). Freeze-dried samples were rehydrated with several media; the highest percentage viability was obtained with 10% non-fat skim milk (100%+). The effect of initial bacterial load on the final recovery was dependent on protectant but not on rehydration media. Sucrose was an effective protectant when a high initial concentration (10(10) cfu ml(-1) was used; the opposite occurred with non-fat skim milk. The use of 10(10) cfu ml(-1) as an initial concentration, sucrose as a protectant and non-fat skim milk as a rehydration medium enabled 100% of P. agglomerans viability to be conserved after freeze-drying. Results suggest the possibility of achieving a good formulation system for the studied biocontrol agent with a high number of viable cells to be used toward pathogens, which is desirable for the industrial development of the product. PMID- 11119152 TI - Release of cell-free ice nuclei from Halomonas elongata expressing the ice nucleation gene inaZ of Pseudomonas syringae. AB - Release of ice nuclei in the growth medium of recombinant Halomonas elongata cells expressing the inaZ gene of Pseudomonas syringae was studied in an attempt to produce cell-free active ice nuclei for biotechnological applications. Cell free ice nuclei were not retained by cellulose acetate filters of 0.2 microm pore size. Highest activity of cell-free ice nuclei was obtained when cells were grown in low salinity (0.5-5% NaCl, w/v). Freezing temperature threshold, estimated to be below -7 degrees C indicating class C nuclei, was not affected by medium salinity. Their density, as estimated by Percoll density centrifugation, was 1.018 +/- 0.002 gml(-1) and they were found to be free of lipids. Ice nuclei are released in the growth medium of recombinant H. elongata cells probably because of inefficient anchoring of the ice-nucleation protein aggregates in the outer membrane. The ice+ recombinant H. elongata cells could be useful for future use as a source of active cell-free ice nucleation protein. PMID- 11119154 TI - Heat resistance of Paenibacillus polymyxa in relation to pH and acidulants. AB - The efficacy of different organic acids in decreasing the heat resistance of Paenibacillus polymyxa spores was assessed. The relationship between concentration of the undissociated form of different organic acids and decrease in heat resistance was also investigated. The heat resistance of P. polymyxa spores was tested in distilled water at 85, 90 and 95 degrees C, at pH4 and in the presence of 50, 100 and 200 mmol l(-1) of the undissociated form of lactic, citric or acetic acid and sodium citrate or acetate. The undissociated form of organic acids was responsible for increasing the heat sensitivity of spores. The most effective acid was lactic acid. The D values of the spores decreased rapidly (between 74 and 43%) in the presence of 50 mmol l(-1) of the undissociated form of organic acid, and increasing concentrations of these forms affected the heat resistance of spores less than proportionally. The heat resistance of the spores in milk was approximately threefold lower than in distilled water. This work has shown that the undissociated fraction of organic acids increases, albeit non linearly, the sensitivity of spores to heat, even in complex substrates such as milk. By knowing the amount of organic acids added to a given substrate, their dissociation constants and the final pH, it could be possible to estimate the concentration of undissociated forms and the corresponding increase in lethality of heat treatments. This would help the food industry to maximize the lethality achieved by heat processes and/or safely reduce the heat treatments already in use. PMID- 11119155 TI - Changes in the Lactobacillus community during Ricotta forte cheese natural fermentation. AB - The loss of microbial biodiversity due to the increase in large-scale industrial processes led to the study of the natural microflora present in a typical little known dairy product. The community of lactobacilli was studied in order to understand the natural fermentation of Ricotta forte cheese. The combined use of RAPD analysis, 16S rDNA sequencing and physiological tests allowed 33 different strains belonging to 10 species of Lactobacillus to be characterized. RAPD analysis revealed the heterogeneity of both the Lact. kefiri and Lact. paracasei species. The sequence analysis of the large 16S/23S rRNA spacer region enabled Lact. plantarum to be distinguished from Lact. paraplantarum, two closely related species belonging to the Lact. plantarum group. The recovery of strains endowed with interesting physiological characteristics, such as strong stress resistance, could improve technological and/or organoleptic characteristics of Ricotta forte cheese and other fermented foods. PMID- 11119156 TI - Exclusion of vanA, vanB and vanC type glycopeptide resistance in strains of Lactobacillus reuteri and Lactobacillus rhamnosus used as probiotics by polymerase chain reaction and hybridization methods. AB - Strains of Lactobacillus reuteri and Lact. rhamnosus are used as probiotics in man and animal. The aim of this study was to determine whether the glycopeptide resistance in these lactobacilli has a similar genetic basis as in enterococci. Five Lact. reuteri strains and one Lact. rhamnosus, as well as four Enterococcus control strains, were probed for the vanA gene cluster, the vanB gene and the vanC gene by PCR and Southern hybridization, and DNA/DNA hybridization. Their resistance and plasmid patterns were also investigated. All Lactobacillus strains were resistant to vancomycin but susceptible to a broad range of antibiotics. Four of the Lactobacillus strains (including the Lact. rhamnosus strain) did not harbour any plasmid and two of them contained five and 6 plasmid bands respectively. None of the Lactobacillus strains possessed the vanA, vanB or vanC gene. These findings indicate that the glycopeptide resistance of the Lactobacillus strains analysed is different from the enterococcal type. The study provides reassurance on the safety of the Lactobacillus strains used as probiotics with regard to their vancomycin resistance. PMID- 11119157 TI - Detection and differentiation between mycotoxigenic and non-mycotoxigenic strains of two Fusarium spp. using volatile production profiles and hydrolytic enzymes. AB - Volatile profiles and hydrolytic enzyme production by one non-mycotoxigenic and three mycotoxigenic strains of Fusarium moniliforme and F. proliferatum, grown in vitro for up to 96 h on a grain medium at 25 degrees C/0.95 water activity, were examined for differentiation of isolates. After spore lawn inoculation, measurements were made after 48, 72 and 96 h by sampling the head space above cultures with an electronic nose system using a 14 sensor surface polymer array, and by extraction and quantification of hydrolytic enzymes. There was good reproducibility of volatile patterns between replicates of the same treatment. Principal component analysis indicated that discrimination could be achieved between the uninoculated controls, the non-mycotoxigenic strain and the mycotoxin producing strains for both species after 48 h. The total and specific activity of three out of seven enzymes (beta-D-glucosidase, alpha-D-galactosidase and N acetyl-beta-D-glucosaminidase) were found to increase significantly in the non mycotoxigenic when compared with the toxigenic strains of both species after 72 h. Activities of the others (beta-D-fucosidase, alpha-D-mannosidase, beta-D xylosidase and N-acetyl-alpha-D-glucosaminidase) were not significantly different between strains. The study has shown for the first time that it is possible to differentiate between mycotoxigenic and non-mycotoxigenic strains of such spoilage fungi based on their volatile production patterns using an electronic nose system. These results have significance in the development of methods for the early detection of toxin-producing spoilage moulds in the food industry. PMID- 11119158 TI - Changes in major components of tea fungus metabolites during prolonged fermentation. AB - Changes in major components and microbes in tea fungus broth (or kombucha; teakwass) prepared from nine different sources during a prolonged fermentation of up to 60 days were investigated. Cell concentrations of both yeasts and acetic acid bacteria in broth were generally higher than those in the cellulosic pellicles. The residual sucrose concentration decreased linearly with time, although the rate fell after the first month. Metabolic fates of glucose and fructose produced as a result of the hydrolysis of sucrose were different. Glucose was not produced in parallel with fructose (0.085 g 100 ml(-1) d(-1)) but was produced with a lower initial rate (0.041 g 100 ml(-1) d(-1)). Both titratable acidity and gluconic acid increased steadily with time for all samples, although gluconic acid was not generated for 6 days until the fermentation had begun. Acetic acid increased slowly to a maximum value of 1.1 g 100 ml(-1) after 30 days; thereafter, it decreased gradually. Gluconic acid contributed to the titratable acidity and thus, the taste of tea fungus broth, during the final stage of fermentation. It is concluded that the desired quality or composition of kombucha can be obtained through the proper control of fermentation time. PMID- 11119159 TI - Isolation of Bacillus strains from the rhizosphere of cereals and in vitro screening for antagonism against phytopathogenic, food-borne pathogenic and spoilage micro-organisms. AB - Bacillus strains were isolated from the rhizosphere of cereals in order to be used as natural biocontrol agents (BCAs). They were screened for antagonism in vitro against various test micro-organisms. The isolates showing antagonism were identified to species level. A combination of techniques was employed for the isolation of Bacillus species. Using the direct method, only one of the 25 isolates screened showed antagonistic properties. This strain (IFS-01) was identified by means of API test strips and the ATB Plus computer programme. It proved to be Bacillus subtilis and consequently has been designated as Bacillus subtilis IFS-01. This strain produced either a broad spectrum antimicrobial compound or several compounds with different activities. The fungi and Gram positive bacteria were more sensitive to the antagonistic isolate than the Gram negative bacteria. A Bacillus strain producing BCAs which can be used as biopesticides or organic preservatives has been isolated and identified. PMID- 11119160 TI - Surface hydrophobicity, viability and efficacy in biological control of Penicillium oxalicum spores produced in aerial and submerged culture. AB - The surface hydrophobicity, viability and biocontrol ability of Penicillium oxalicum spores, produced either in aerial or submerged culture, were characterized. A phase distribution test showed that spores produced in both methods of culture were highly hydrophobic, but those produced in aerial culture were more hydrophobic. Spores stored fresh at either 4 or 25 degrees C retained a high viability (80%) after 27 weeks of storage, although aerial spores survived better. Freeze-drying severely affected viability, especially of submerged spores. Biocontrol ability against Fusarium oxysporum f. sp. lycopersici was studied in the growth chamber. Aerially- produced spores were more effective than submerged ones. Aerially-produced P. oxalicum spores appeared to have more advantages than those produced by submerged culture, in relation to both viability and efficacy. These results demonstrate that physiological changes occur depending on production conditions which significantly influences quality of the biocontrol agent. PMID- 11119161 TI - Inactivation of Mycobacterium bovis in cattle slurry by five volatile chemicals. AB - This research was undertaken to evaluate volatile chemicals which retained mycobactericidal activity in cattle slurry. Mycobacterium bovis, suspended in sterilized cattle slurry, was treated with different concentrations of five volatile chemicals with mycobactericidal activity. Following treatment of the slurry for specified time intervals, the reaction mixture was lyophilized to remove the volatile chemicals, and samples of the reconstituted slurry were used to inoculate flasks of Lowenstein-Jensen medium to determine survival or inactivation of the mycobacteria. Acetone, at a concentration of 22.5%, inactivated M. bovis in less than 24 h. Ammonium hydroxide, at a concentration of 1%, was mycobactericidal after 36 h. Chloroform at a concentration of 0.5%, ethyl alcohol at a concentration of 17.5% and xylene at a concentration of 3% inactivated the mycobacteria within 48 h. Some of the volatile chemicals with mycobactericidal activity are potentially useful at farm level. PMID- 11119162 TI - Autolysis of dairy leuconostocs and detection of peptidoglycan hydrolases by renaturing SDS-PAGE. AB - The autolysis of lactic acid bacteria plays a major role during cheese ripening. The aim of this study was to evaluate the autolytic properties and peptidoglycan hydrolase content of dairy leuconostocs. Autolysis of 59 strains of dairy Leuconostoc was examined under starvation conditions in potassium phosphate buffer. The ability of dairy leuconostocs to lyse is strain dependant and not related to the species. The peptidoglycan hydrolase profile of Leuc. mesenteroides subsp. mesenteroides 10L was analysed by renaturing gel electrophoresis. Two major activity bands migrating at 41 and 52 kDa were observed. According to the specificity analysis, strain 10L seems to contain a glycosidase and an N-acetyl-muramyl-L-alanine amidase, or an endopeptidase. The peptidoglycan hydrolase profiles of various Leuconostoc species were also compared. Several peptidoglycan hydrolase activities could be detected in the different Leuconostoc species. Further characterization of the peptidoglycan hydrolases will help to control autolysis of leuconostocs in cheese. PMID- 11119163 TI - Cell-bound and extracellular carboxylesterases from Streptomyces: hydrolytic and synthetic activities. AB - AIM: The distribution of cell-bound and extracellular carboxylesterases was investigated among the genus Streptomyces using 420 strains. METHODS AND RESULTS: Primary screening was carried out on solid media using tributyrin, triolein and Tween 60 as current substrates. Eleven representative strains were selected and grown in submerged cultures for evaluating their cell-bound and extracellular hydrolytic activity independently on various naphthyl and aliphatic esters. The best lipolytic strain was lyophilized and used as dry mycelium for catalysing the synthesis of various aliphatic esters in heptane, with molar conversions ranging from 28 to 78% after 3 days. CONCLUSIONS: Carboxylesterase activities can easily be found among the Streptomyces, often being cell-bound and also employable for catalysing esterification in organic solvent. SIGNIFICANCE AND IMPACT OF THE STUDY: A wide screening among Streptomyces, a genus poorly studied for the production of carboxylesterases, has allowed the selection of several strains with interesting enzymatic activities to be used in commercially valuable biotransformation. PMID- 11119165 TI - Comparison of three enrichment media for the isolation of Campylobacter spp. from foods. AB - AIM: This study compared the performance of three Campylobacter enrichment broths: Bolton broth (BB), Campylobacter Enrichment broth (CEB) and Preston broth (PB). METHODS AND RESULTS: Pure cultures of target and competitor organisms, and naturally-contaminated food samples, were used to establish the performance of these media. In pure culture the PB supported the growth of the greatest number of strains of Campylobacter spp. but failed to inhibit some competitor organisms. The CEB showed the opposite result, inhibiting all 15 competitor organisms used but failing to support the growth of five Campylobacter strains. By comparison, BB showed the best compromise between inhibition of competitors and growth of Campylobacter. CONCLUSIONS: Plates inoculated with BB and CEB food enrichments resulted in more Campylobacter growth than those inoculated with PB, which supported significantly less typical growth (P < or = 0.001). The most common competitor organism isolated from PB was Escherichia coli, and Pseudomonas spp. were frequently isolated from BB and CEB. Both BB and CEB were better than PB for the isolation of Campylobacter from naturally-contaminated foods, although BB yielded more confirmed Campylobacter growth than CEB. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlighted differences in performance of media used to isolate Campylobacter spp. from foods. PMID- 11119164 TI - Use of a PGU1 recombinant Saccharomyces cerevisiae strain in oenological fermentations. AB - AIM: The aim of this work was the construction of an oenological Saccharomyces cerevisiae strain able to overexpress the PGU1 gene in order to be used in trial fermentations. METHODS AND RESULTS: The recombinant strain is able to secrete an active endopolygalacturonase into the medium leaving its fermentation ability essentially unchanged. Wines obtained with the recombinant strain and the untransformed counterpart did not differ in their physicochemical parameters or major sensory characteristics. The time needed for wine filtration was dramatically reduced in wines elaborated with the PGU1 recombinant strain, and was comparable to the filtration time shown by wines elaborated from must supplemented with fungal pectolytic enzymes. CONCLUSIONS: The oenological strain constructed in this work secretes an endopolygalacturonase into the wine in an efficient manner, resulting in an improvement in wine filtration but preserving wine typicality and keeping the methanol levels unchanged. SIGNIFICANCE AND IMPACT OF THE STUDY: The PGU1 recombinant strains could be used in oenological fermentations as an alternative to commercial pectolytic enzymes of fungal origin. PMID- 11119166 TI - The effect of thermal stress on Campylobacter coli. AB - AIM: Enteropathogenic Campylobacter jejuni, Camp. coli and Camp. lari are currently the most common cause of acute infectious diarrhoeal illness in the UK. Many domestic animals, including pigs, act as natural reservoirs for these organisms and infection may occur through the ingestion of contaminated foodstuffs. The safety of locally produced porcine liver was assessed in relation to the heat susceptibility of Campylobacter spp. present in eviscerated product. METHODS AND RESULTS: Heat susceptibility (D10) studies were performed on a wild type strain of Camp. coli [NI39] isolated from porcine liver under standardized conditions. In addition, the effect of culture age and heating menstruum was determined. Thermal stress studies in phosphate-buffered saline showed Camp. coli NI39 to be heat sensitive (D10 = 8.-0, 30.8, 15.6, 10.3 s at 55.4, 57.4, 59.7, 61.2 degrees C, respectively; z = 6.10 degrees C). However, non-logarithmic biphasic survivor curves were observed at higher temperatures ( > 56 degrees C), indicating the presence of a heat-resistant subpopulation (10(4)-10(5) cfu) which was not demonstrated when examining either Salmonella typhimurium or Listeria monocytogenes. CONCLUSIONS: The use of D10 values may be limited. Therefore, porcine liver, under processing, must be treated as a potential source of Campylobacter spp., and clearly defined F-values should be quantified through the use of empirically 'spiked' samples to ensure the eradication of campylobacters from the product, for each individual process being evaluated. SIGNIFICANCE AND IMPACT OF THE STUDY: It is important to define safe processing parameters in the manufacture of products which receive mild thermal processes in order to eliminate the risk of disease to man. PMID- 11119167 TI - Effect of physician and patient gender concordance on patient satisfaction and preventive care practices. AB - OBJECTIVE: To explore the role of the gender of the patient and the gender of the physician in explaining differences in patient satisfaction and patient-reported primary care practice. DESIGN: Crosssectional mailed survey [response rate of 71%]. SETTING: A large group-model Health Maintenance Organization (HMO) in northern California. PATIENTS/PARTICIPANTS: Random sample of HMO members aged 35 to 85 years with a primary care physician. The respondents (N = 10,205) were divided into four dyads: female patients of female doctors; male patients of female doctors; female patients of male doctors; and male patients of male doctors. Patients were also stratified on the basis of whether they had chosen their physician or had been assigned. MEASUREMENTS AND MAIN RESULTS: Among patients who chose their physician, females who chose female doctors were the least satisfied of the four groups of patients for four of five measures of satisfaction. Male patients of female physicians were the most satisfied. Preventive care and health promotion practices were comparable for male and female physicians. Female patients were more likely to have chosen their physician than males, and were much more likely to have chosen female physicians. These differences were not seen among patients who had been assigned to their physicians and were not due to differences in any of the measured aspects of health values or beliefs. CONCLUSIONS: Our study revealed differences in patient satisfaction related to the gender of the patient and of the physician. While our study cannot determine the reasons for these differences, the results suggest that patients who choose their physician may have different expectations, and the difficulty of fulfi11ing these expectations may present particular challenges for female physicians. PMID- 11119168 TI - Physicians' experiences with patients who transgress boundaries. AB - BACKGROUND: Boundary violations have been discussed in the literature, but most studies report on physician transgressions of boundaries or sexual transgressions by patients. We studied the incidence of all types of boundary transgressions by patients and physicians' responses to these transgressions. METHODS: We surveyed 1,000 members of the Society of General Internal Medicine (SGIM) for the number of patient transgressions of boundaries which had occurred in the previous year. Categories were created by the investigators based on the literature. Physicians picked the most important transgression, and then were asked about their response to the transgression and its effect on the patient-physician relationship. Attitudinal questions addressed the likelihood of discharging patients who transgressed boundaries. The impact of demographic variables on the incidence of transgressions was analyzed using analysis of variance. RESULTS: Three hundred thirty (37.5%) randomly selected SGIM members responded to the survey. Almost three quarters of the respondents had patients who used their first name, while 43% encountered verbal abuse, 39% had patients who asked personal questions, 31% had patients who were overly affectionate, and 27% encountered patients who attempted to socialize. All other transgressions, including physical abuse and attempts at sexual contact, were uncommon. Only gender affected the incidence of transgressions; female physicians encountered more personal questions (P = .001), inappropriate affection (P < .005), and sexually explicit language (P < .05) than male physicians and responded more negatively to boundary transgressions. Respondents dealt with transgressions by discussion with the patient or colleagues or by ignoring the incident, but such transgressions generally had a negative impact on the relationship. Most physicians would discharge patients who engaged in physical abuse or attempts at sexual contact, but were more tolerant of verbal abuse and overly affectionate patients. CONCLUSIONS: Boundary transgressions by patients is common, but usually involves more minor infractions. Female physicians are more likely to encounter certain types of transgressions. The incidence and outcomes of such transgressions are important in assisting physicians to deal effectively with this issue. PMID- 11119169 TI - Adherence to guidelines for oral anticoagulation after venous thrombosis and pulmonary embolism. AB - OBJECTIVE: Guidelines for oral anticoagulation after deep venous thrombosis (DVT) or pulmonary embolism (PE) have recommended that patients be anticoagulated for at least 3 months after hospital discharge. We sought to determine whether this recommendation was being followed and what patient characteristics predict a shorter than recommended duration of therapy. DESIGN: Retrospective cohort study using linked health care claims data. SETTING: Routine clinical practice. PATIENTS: Five hundred seventy-three members of New Jersey's Medicaid or Pharmacy Assistance for the Aged and Disabled programs aged 65 years and older who were hospitalized for DVT or PE between January 1, 1991 and June 30, 1994. RESULTS: Of the 573 patients, 129 (23%) filled prescriptions covering less than 90 days of oral anticoagulant therapy. In multivariate models, African-American race was associated with an increased risk of a shorter than recommended duration of therapy (odds ratio [OR], 1.87; 95% confidence interval [CI], 1.14 to 3.08), but age and gender were not. Patients who used anticoagulants in the year prior to admission were less likely to have a short duration of therapy (OR, 0.30; 95% CI, 0.12 to 0.78), than were patients with PE (OR, 0.58; 95% CI, 0.38 to 0.88). CONCLUSIONS: Nearly a quarter of those anticoagulated following DVT or PE received therapy for less than the recommended length of time after hospital discharge, with African Americans more likely to have a shorter than recommended course of treatment. Further research is needed to evaluate the causes of shorter than recommended duration of therapy and racial disparities in anticoagulant use. PMID- 11119170 TI - Measuring compliance with preventive care guidelines: standardized patients, clinical vignettes, and the medical record. AB - OBJECTIVE: To determine how accurately preventive care reported in the medical record reflects actual physician practice or competence. DESIGN: Scoring criteria based on national guidelines were developed for 7 separate items of preventive care. The preventive care provided by randomly selected physicians was measured prospectively for each of the 7 items. Three measurement methods were used for comparison: (1) the abstracted medical record from a standardized patient (SP) visit; (2) explicit reports of physician practice during those visits from the SPs, who were actors trained to present undetected as patients; and (3) physician responses to written case scenarios (vignettes) identical to the SP presentations. SETTING: The general medicine primary care clinics of two university-afflliated VA medical centers. PARTICIPANTS: Twenty randomly selected physicians (10 at each site) from among eligible second- and third-year general internal medicine residents and attending physicians. MEASUREMENTS AND MAIN RESULTS: Physicians saw 160 SPs (8 cases x 20 physicians). We calculated the percentage of visits in which each prevention item was recorded in the chart, determined the marginal percentage improvement of SP checklists and vignettes over chart abstraction alone, and compared the three methods using an analysis-of variance model. We found that chart abstraction underestimated overall prevention compliance by 16% (P < .01) compared with SP checklists. Chart abstraction scores were lower than SP checklists for all seven items and lower than vignettes for four items. The marginal percentage improvement of SP checklists and vignettes to performance as measured by chart abstraction was significant for all seven prevention items and raised the overall prevention scores from 46% to 72% (P < .0001). CONCLUSIONS: These data indicate that physicians perform more preventive care than they report in the medical record. Thus, benchmarks of preventive care by individual physicians and institutions that rely solely on the medical record may be misleading, at best. PMID- 11119171 TI - Impact of obesity on health-related quality of life in patients with chronic illness. AB - OBJECTIVE: To determine the association between overweight and obesity and health related quality of life (HRQOL) in patients with chronic conditions typical of those seen in general medical practice, after accounting for the effects of depression and medical comorbidities. DESIGN: Cross-sectional analysis of data from the Medical Outcomes Study. SETTING: Offices of physicians practicing family medicine, internal medicine, endocrinology, cardiology, and psychiatry in three U.S. cities. PATIENTS: We surveyed 2,931 patients with chronic medical and psychiatric conditions. The patients completed a self-administered questionnaire at enrollment and had complete data on height and weight. MEASUREMENTS AND MAIN RESULTS: Body mass index (BMI), chronic medical conditions, and depression were obtained by structured interview. Health-related quality of life was measured by the SF-36 Health Survey. Patients who were overweight (BMI 25.0-29.9 kg/m2), patients with class I obesity (BMI 30.0-34.9 kg/m2), and patients with class II III obesity (BMI > or = 35 kg/m2) had significantly lower adjusted physical function scores (by 3.4, 7.8, and 13.8 points, respectively) compared with nonoverweight patients. Patients with class I and class II-III obesity also had significantly lower adjusted general health perceptions scores (by 2.8 and 4.4 points, respectively) and lower adjusted vitality scores (by 4.0 and 7.1 points, respectively), compared with nonoverweight patients. No significant differences between nonoverweight, overweight, and obese patients were observed for the mental health scale. Women with elevated BMI had significantly lower HRQOL scores compared with the scores of obese men in several domains. Additionally, blacks with elevated BMI had significantly lower scores than whites in several domains of HRQOL. CONCLUSIONS: Overweight and obesity have the largest association with physical function measures. Recent national standards, which have lowered the threshold for defining overweight, identify patients who are more likely to have clinically significant reductions in HRQOL and functional impairment. PMID- 11119172 TI - Perceived access problems among patients with diabetes in two public systems of care. AB - OBJECTIVE: We examined the prevalence of access problems among public clinic patients after participating in trials of automated telephone disease management with nurse follow-up. DESIGN: Randomized trial. SETTING: General medicine clinics of a county health care system and a Veterans Affairs (VA) health care system. PARTICIPANTS: Five hundred seventy adults with diabetes using hypoglycemic medication were enrolled and randomized; 520 (91%) provided outcome data at 12 months. INTERVENTION: Biweekly automated telephone assessments with telephone follow-up by diabetes nurse educators. MEASUREMENTS AND MAIN RESULTS: At follow up, patients reported whether in the prior 6 months they had failed to obtain each of six types of health services because of a financial or nonfinancial access problem. Patients receiving the intervention were significantly less likely than patients receiving usual care to report access problems (adjusted odds ratio [AOR], 0.61; 95% confidence interval [CI], 0.43 to 0.97). The risk of reporting access problems was greater among county clinic patients than VA patients even when adjusting for their experimental condition, and socioeconomic and clinical risk factors (AOR, 1.61; 95% CI, 1.02 to 2.53). County patients were especially more likely to avoid seeking care because of a worry about the cost (AOR, 2.82; 95% CI, 1.48 to 5.37). CONCLUSIONS: Many of these public sector patients with diabetes reported that they failed to obtain health services because they perceived financial and nonfinancial access problems. Automated telephone disease management calls with telephone nurse follow-up improved patients' access to care. Despite the impact of the intervention, county clinic patients were more likely than VA patients to report access problems in several areas. PMID- 11119173 TI - A picture is worth a thousand words: practical use of videotape in teaching. AB - Videotapes, through vividly displayed clinical images and teaching interactions, are valuable tools for both learners and teachers. Visual images in combination with verbal instruction have been shown to significantly increase recall and retention. Many clinicians and medical teachers are aware of videotape resources, but have not had a chance to develop their use in medical education. In this paper, we discuss creative applications of videotapes in three major categories: presenting information, triggering discussion, and as a tool for direct self observation and feedback. Videotapes may be valuable for presenting information in settings of didactic instruction; for triggering discussion during teaching workshops; and for self-observation of patient-doctor interactions and learner teacher encounters. The article presents learner-centered approaches to review a videotaped clinical encounter in order to enhance value and comfort for the learner and teacher. Sources of tapes include on-site videotaping, published educational tapes and commercial tapes shown in accordance with fair use guidelines, examples of which are referenced. Videotapes add another dimension to traditional sources of physician education. PMID- 11119174 TI - Prevalence and determinants of intimate partner abuse among public hospital primary care patients. AB - OBJECTIVE: To determine the prevalence, sociodemographic determinants, and depression correlates of intimate partner abuse among an ethnically diverse population of women patients. DESIGN: Cross-sectional telephone survey in English and Spanish of a random sample of women patients aged 18 to 46 years. SETTING: Three public hospital primary care clinics (general internal medicine, family medicine, and obstetrics/gynecology) in San Francisco, Calif. PARTICIPANTS: We interviewed 734 (74%) of the 992 eligible participants. Thirty-one percent were non-Latina white, 31% African American, and 36% Latina. MEASUREMENTS AND MAIN RESULTS: Using questions adapted from the Abuse Assessment Screen, we determined recent and lifetime history of physical, sexual, and psychological abuse. Overall, 15% reported recent abuse by an intimate partner (in the preceding 12 months); lifetime prevalence was 51%. Recent abuse was more common among women aged 18 to 29 years (adjusted odds ratio [OR] 2.1; 95% confidence interval [CI], 1.2 to 3.7), non-Latinas (adjusted OR, 1.7; 95% CI, 1.0 to 2.9), and unmarried women (adjusted OR, 1.65; 95% CI, 1.0 to 2.7). The prevalence of abuse did not differ by education, employment, or medical insurance status of the women. Compared with women with no history of abuse, a greater proportion of recently abused women reported symptoms of depression (adjusted OR, 3.5; 95% CI, 2.2 to 5.5). CONCLUSIONS: Because a substantial proportion of women patients in primary care settings are abused, screening for partner abuse and depression is indicated. In contrast to other studies, lower socioeconomic status was not associated with partner abuse history. PMID- 11119175 TI - Nutrition management of type 2 diabetes by primary care physicians: reported use and barriers. AB - A survey was mailed to a probability sample of primary care physicians in Indiana to assess their use of and barriers to nutritional therapy for patients with type 2 diabetes. Most (62%) primary care physicians reported referring their type 2 diabetes patients for nutrition counseling, while 38% reported providing counseling themselves. Patient-centered barriers were most frequently cited as reasons for poor effectiveness of nutrition therapy. This differs from previous research that cites system-level factors as barriers. PMID- 11119176 TI - Outpatient morning report: a new conference for internal medicine residency programs. AB - To clarify the use of outpatient morning report in internal medicine residency programs, we conducted a national survey of internal medicine residency directors and a local survey of a cohort of residents at a large teaching hospital. The program directors reported a 24% prevalence of outpatient morning report. The cohort of residents reported that the conference contributed much to their education by meeting specific learning needs and covering topics not covered elsewhere in their residency training. PMID- 11119177 TI - Interpersonal expectations in the patient-physician relationship. PMID- 11119178 TI - The challenge of obesity-related chronic diseases. PMID- 11119179 TI - Propensity of HIV patients to seek urgent and emergent care. HIV Cost and Services Utilization Study Consortium. AB - OBJECTIVE: To assess the propensity of HIV-infected adults to seek care for common symptoms, and to determine whether they would seek care in the emergency department (ED) or with their primary care provider. DESIGN: Cross-sectional interview study. SETTING: Patients in care in the 48 contiguous United States. PARTICIPANTS: A nationally representative group of HIV- infected adults selected using multistage probability sampling. MEASUREMENTS: Subjects were interviewed between January 1996 and April 1997. Patients with advanced disease (past AIDS diagnosis and/or CD4 cell count <200/microL) and early disease were asked how they would seek care for key HIV-associated symptom complexes. Three advanced disease and 3 early disease symptom scenarios were used. MAIN RESULTS: Most advanced disease patients (78% to 87%) would seek care right away from the ED or primary care provider for the symptoms asked. Most early disease patients (82%) would seek care right away for new respiratory symptoms; fewer would do so for headache (46%) or oral white patches (62%). In a multivariate model, independent predictors of propensity to use the ED for advanced disease symptoms included African-American ethnicity (adjusted odds ratio [OR], 2.5; 95% confidence interval [95% CI], 1.8 to 3.4); less education (adjusted OR, 1.4; 95% CI, 1.1 to 1.7); drug dependence (adjusted OR, 1.4; 95% CI, 1.1 to 1.7); annual income less than $5,000 (adjusted OR, 1.5; 95% CI, 1.0 to 2.3); and lower psychological well being (adjusted OR, 0.9; 95% CI, 0.9 to 1.0). In early disease, the following independently predicted ED use: African American (adjusted OR, 4.7; 95% CI, 3.1 to 7.1) or Hispanic ethnicity (adjusted OR 2.4; 95% CI, 1.4 to 4.3), female gender (adjusted OR, 1.6; 95% CI, 1.2 to 2.2), annual income less than $5,000 (adjusted OR, 1.8; 95% CI, 1.1 to 3. 0), and lower psychological well-being (adjusted OR, 0.9; 95% CI, 0. 8 to 1.0). CONCLUSIONS: Many patients would use the ED instead of same-day primary care for several common symptoms of HIV disease. African Americans, the poor, and patients with psychological symptoms had a higher propensity to use the ED. PMID- 11119180 TI - Cue-dose training with monetary reinforcement: pilot study of an antiretroviral adherence intervention. AB - OBJECTIVE: To assess the feasibility and efficacy of two interventions for improving adherence to antiretroviral therapy regimens in HIV-infected subjects compared with a control intervention. DESIGN: Randomized, controlled, pilot study. SETTING: Department of Veterans Affairs HIV clinic and community-based HIV clinical trials site. PARTICIPANTS: Fifty-five HIV-infected subjects on stable antiretroviral therapy regimens. Subjects were predominantly male (89%) and African American (69%), and had histories of heroin or cocaine use (80%). INTERVENTIONS: Four weekly sessions of either nondirective inquiries about adherence (control group, C), cue-dose training, which consisted of the use of personalized cues for remembering particular dose times, and feedback about medication taking using Medication Event Monitoring System (MEMS) pill bottle caps, which record time of bottle opening (CD group), or cue-dose training combined with cash reinforcement for correctly timed bottle opening (CD+CR). MEASUREMENTS: Opening of the pill bottle within 2 hours before or after a predetermined time was measured by MEMS. RESULTS: Adherence to the medication as documented by MEMS was significantly enhanced during the 4-week training period in the CD+CR group, but not in the CD group, compared with the control group. Improvement was also seen in adherence to antiretroviral drugs that were not the object of training and reinforcement. Eight weeks after training and reinforcement were discontinued, adherence in the cash-reinforced group returned to near-baseline levels. CONCLUSIONS: Cue-dose training with cash reinforcement led to transient improvement in adherence to antiretroviral therapy in a population including mostly African Americans and subjects with histories of drug abuse. However, we were not able to detect any sustained improvement beyond the active training period, and questions concerning the timing and duration of such an intervention require further study. Randomized, controlled clinical studies with objective measures of adherence can be conducted in HIV-infected subjects and should be employed for further evaluation of this and other adherence interventions. PMID- 11119181 TI - Taking antiretroviral therapy for HIV infection: learning from patients' stories. AB - OBJECTIVE: To describe how people with HIV understand and experience the problem of adhering to antiretroviral medication regimens. DESIGN: We performed a qualitative study based on interviews with HIV-infected patients, including 46 clients of AIDS service organizations, who were sampled according to age, ethnicity, and injection drug use history, and a convenience sample of 15 patients. Interviews were conducted in English or Spanish and were audiotaped and transcribed. PARTICIPANTS: Of 52 respondents who had prescriptions for antiretroviral therapy, 25 were randomly selected for in-depth analysis. Of these, 5 reported having an AIDS diagnosis, 15 reported symptoms they attributed to HIV, and 5 reported having no symptoms of HIV disease. MEASUREMENTS AND MAIN RESULTS: Investigators prepared structured abstracts of interviews to extract adherence-related data. One investigator compared the abstracts with the original transcripts to confirm the interpretations, and used the abstracts to organize and classify the findings. Most subjects (84%) reported recent nonadherent behavior, including ceasing treatment, medication "holidays," sleeping through doses, forgetting doses, skipping doses due to side effects, and following highly asymmetric schedules. Initially, most reported that they were not significantly nonadherent, and many did not consider their behavior nonadherent. Only a minority clearly understood the possible consequences of missing doses. Most said they had not discussed their nonadherence with their physicians. CONCLUSIONS: Many people rationalize their difficulty in adhering to HIV treatment by deciding that the standard of adherence they can readily achieve is appropriate. Physicians should inquire about adherence-related behavior in specific detail, and ensure that patients understand the consequences of not meeting an appropriate standard. PMID- 11119182 TI - Predictors of outcome in a primary care depression trial. AB - OBJECTIVE: Previous treatment trials have found that approximately one third of depressed patients have persistent symptoms. We examined whether depression severity, comorbid psychiatric illness, and personality factors might play a role in this lack of response. DESIGN: Randomized trial of a stepped collaborative care intervention versus usual care. SETTING: HMO in Seattle, Wash. PATIENTS: Patients with major depression were stratified into severe (N = 149) and mild to moderate depression (N = 79) groups prior to randomization. INTERVENTIONS: A multifaceted intervention targeting patient, physician, and process of care, using collaborative management by a psychiatrist and primary care physician. MEASUREMENTS AND MAIN RESULTS: Patients with more severe depression had a higher risk for panic disorder (odds ratio [OR], 5.8), loneliness (OR, 2.6), and childhood emotional abuse (OR, 2.1). Among those with less severe depression, intervention patients showed significantly improved depression outcomes over time compared with those in usual care (z = -3.06, P<.002); however, this difference was not present in the more severely depressed groups (z = 0.61, NS). Although the group with severe depression showed differences between the intervention and control groups from baseline to 3 months that were similar to the group with less severe depression (during the acute phase of the intervention), these differences disappeared by 6 months. CONCLUSIONS: Initial depression severity, comorbid panic disorder, and other psychosocial vulnerabilities were associated with a decreased response to the collaborative care intervention. Although the intervention was appropriate for patients with moderate depression, individuals with higher levels of depression may require a longer continuation phase of therapy in order to achieve optimal depression outcomes. PMID- 11119183 TI - Quality improvement for depression enhances long-term treatment knowledge for primary care clinicians. AB - OBJECTIVE: We evaluated the effect of implementing quality improvement (QI) programs for depression, relative to usual care, on primary care clinicians' knowledge about treatment. DESIGN AND METHODS: Matched primary care clinics (46) from seven managed care organizations were randomized to usual care (mailed written guidelines only) versus one of two QI interventions. Self-report surveys assessed clinicians' knowledge of depression treatments prior to full implementation (June 1996 to March 1997) and 18 months later. We used an intent to-treat analysis to examine intervention effects on change in knowledge, controlling for clinician and practice characteristics, and the nested design. PARTICIPANTS: One hundred eighty-one primary care clinicians. INTERVENTIONS: The interventions included institutional commitment to QI, training local experts, clinician education, and training nurses for patient assessment and education. One intervention had resources for nurse follow-up on medication use (QI-meds) and the other had reduced copayment for therapy from trained, local therapists (QI-therapy). RESULTS: Clinicians in the intervention group had greater increases compared with clinicians in the usual care group over 18 months in knowledge of psychotherapy (by 20% for QI-meds, P =.04 and by 33% for QI-therapy, P =.004), but there were no significant increases in medication knowledge. Significant increases in knowledge scores (P =.01) were demonstrated by QI-therapy clinicians but not clinicians in the QI-meds group. Clinicians were exposed to multiple intervention components. CONCLUSIONS: Dissemination of QI programs for depression in managed, primary care practices improved clinicians' treatment knowledge over 18 months, but breadth of learning was somewhat greater for a program that also included active collaboration with local therapists. PMID- 11119184 TI - Use of an orientation clinic to reduce failed new patient appointments in primary care. AB - Patients who fail to attend initial appointments reduce clinic efficiency. To maximize attendance by newly referred outpatients, we introduced a mandatory group orientation clinic for all new patients and determined its effects on no show rates. Orientation clinic also provided health care screening and opportunities for patient feedback. The new patient no-show rate for initial provider visits decreased significantly from 45% before institution of orientation clinic to 18% afterwards (P<.0001). The total no-show (patients who failed to attend orientation clinic or an initial provider visit) rate of the postintervention group was 51% (P = .28, compared with before the intervention). This intervention improved the efficiency and minimized the wasted time of our clinicians. PMID- 11119185 TI - Journal reading habits of internists. AB - We assessed the reading habits of internists with and without epidemiological training because such information may help guide medical journals as they make changes in how articles are edited and formatted. In a 1998 national self administered mailed survey of 143 internists with fellowship training in epidemiology and study design and a random sample of 121 internists from the American Medical Association physician master file, we asked about the number of hours spent reading medical journals per week and the percentage of articles for which only the abstract is read. Respondents also were asked which of nine medical journals they subscribe to and read regularly. Of the 399 eligible participants, 264 returned surveys (response rate 66%). Respondents reported spending 4.4 hours per week reading medical journal articles and reported reading only the abstract for 63% of the articles; these findings were similar for internists with and without epidemiology training. Respondents admitted to a reliance on journal editors to provide rigorous and useful information, given the limited time available for critical reading. We conclude that internists, regardless of training in epidemiology, rely heavily on abstracts and prescreening of articles by editors. PMID- 11119186 TI - Serum potassium and cardiovascular mortality. AB - BACKGROUND: The impact of serum potassium on mortality is inadequately defined. OBJECTIVE: To determine the association of serum potassium with mortality. METHODS: We analyzed NHANES I Epidemiological Follow-up Study data from 1974 1992. Of 2,992 subjects with baseline serum potassium, 156 were excluded because their vital status was not known. A total of 2,836 subjects with serum potassium within 2.7-5.4 mmol/L were studied. All-cause and cardiovascular mortality were assessed controlling for sociodemographic status, smoking, medical history, and clinical characteristics. RESULTS: At baseline, mean age was 46.6 years, and mean serum potassium was 4.07 mmol/L. Subjects were stratified into three groups by mean +/-1 standard deviation of serum potassium: low, 2.7-3.7 mmol/L (N = 477); middle, 3.8-4.4 mmol/L (N = 1,982); and high, 4.5-5.4 mmol/L (N = 377). The cardiovascular mortality rate per 1,000 person-years adjusted for age, gender, and race for the high serum potassium group (8.1) was significantly higher than the middle (5.3) and low (6.5) serum potassium groups. Further analysis, controlling for age, gender, race, smoking status, cholesterol, and history of diabetes, renal disease, and cardiovascular disease, revealed that the increased cardiovascular mortality among subjects with moderately increased serum potassium was most prominent in those reporting use of diuretics (hazard ratio, 2.65; 95% confidence interval [95% CI], 1.20 to 5.85) and those with abnormal renal function (hazard ratio, 1.89; 95% CI, 1.05 to 3.41). CONCLUSION: In this general population sample with mostly normal serum potassium, higher serum potassium was independently associated with increased cardiovascular mortality. PMID- 11119187 TI - Adherence and health care utilization in HIV/AIDS-rational or rationalizing? PMID- 11119188 TI - Interventions that improve the quality of depression care: where do we go from here? PMID- 11119189 TI - Latex allergy: where are we? PMID- 11119190 TI - Anaesthesia, perioperative management and outcome of correction of extrahepatic biliary atresia in the infant: a review of 50 cases in the King's College Hospital series. AB - Extrahepatic biliary atresia (EHBA) is an uncommon condition presenting in the first few weeks of life. It has an incidence of 0. 5-1 per 10 000 live births and is the end result of a destructive inflammatory process involving the extrahepatic biliary system of unknown aetiology occurring in utero. The net result is neonatal jaundice due to bile stasis, with subsequent hepatocellular damage and cirrhosis. In the untreated, patient death is inevitable within 2 years. Precise diagnosis (or exclusion) of EHBA in the persistently jaundiced infant must be made urgently and major surgery (hepatic portoenterostomy: Kasai procedure) carried out as soon as possible, preferably before 6-8 weeks of age. This review is concerned with anaesthesia for correction of EHBA in 50 consecutive patients and also outlines the experience gained in the largest European centre for correction of EHBA where the number of cases now approaches 500. PMID- 11119191 TI - Noninvasive blood pressure measurement in the upper and lower limbs of anaesthetized children. AB - A group of 50 children, aged 5 months to 15 years, and who were undergoing routine surgery under general anaesthesia, were studied to investigate the difference in noninvasive blood pressure readings obtained from inflatable cuffs placed on the upper arm and the lower leg. In contrast to adult data, it was found that the blood pressure measured from the leg in children aged 8 years and under, was significantly lower than that measured from the arm. The leg cuff measurements could not, however, be reliably used to predict arm blood pressure. PMID- 11119192 TI - Effect of different anaesthetic regimes on the oculocardiac reflex during paediatric strabismus surgery. AB - The oculocardiac reflex (OCR) is induced by mechanical stimulation and therefore is frequently encountered during strabismus surgery. This study was designed to determine how various anaesthetic regimes modulate the haemodynamic effects of the OCR during paediatric strabismus surgery. Thirty-nine patients (4-14 years, ASA I) were randomized to one of four anaesthetic regimes: group P: propofol (12 mg.kg(-1).h(-1)) and alfentanil (0.04 mg.kg(-1).h(-1)); group S: sevoflurane 1 1.2 MAC in 30% O(2)/70% N(2)O; group K: ketamine racemate (10-12 mg. kg(-1).h( 1)) and midazolam (0.3-0.6 mg.kg(-1).h(-1); group H: halothane 1-1. 2 MAC in 30% O(2)/70% N(2)O. Electrocardiogram (ECG), beat-to-beat heart rate (HR) and blood pressure (BP) changes were measured during and after a standardized traction was applied to an external eye muscle (4-6 Newton, 90 s). OCR was defined as a 10% change in HR induced by traction. OCR occurred in 77% of patients. Whereas virtually all patients in the P, H and S groups developed OCR, only 22% developed it in group K. Median HR change in group P (-37 bpm) was significantly greater (P < 0.05) than in group H (-17 bpm) or group K (-7 bpm). Median BP change in group K (+10 mmHg) was significantly different (P < 0.05) from group H (-5 mmHg), group S (-3 mmHg) and group P (-8 mmHg). Atrioventricular rhythm disorders were significantly more frequent in group P compared with group K (P < 0.02). Respiration-induced sinus dysrhythmia was significantly less frequent (P < 0.001) in group K (0%), compared with group P (100%), group H (56%) and group S (55%). Of the anaesthetic techniques studied, ketamine anaesthesia is associated with the least haemodynamic changes induced by OCR during strabismus surgery in paediatric patients. PMID- 11119193 TI - Epidural sufentanil during paediatric cardiac surgery: effects on metabolic response and postoperative outcome. AB - The metabolic and neuroendocrine effects of caudal epidural analgesia were studied during paediatric cardiac surgery. Combined epidural and general anaesthesia (EPI group; n=12) was compared with deep opioid anaesthesia (DOA group; n=12). During anaesthesia and surgery, haemodynamic stability was similar in the two groups. There was no significant difference between groups concerning the metabolic response to surgery but circulating catecholamines were significantly lower in the EPI group during and after surgery. Perioperative release of IL-6 was higher in the EPI group possibly reflecting a longer aortic clamp time. Incidence of postoperative life-threatening dysrhythmias was very low in the two groups. No significant reduction of postoperative mechanical ventilation, intensive care unit or hospital stays was reported with epidural analgesia. The incidence of postoperative infections was higher than expected in the two groups because of the poor properative clinical status of most of the children included in the study. PMID- 11119194 TI - Plasma concentrations of bupivacaine after combined spinal epidural anaesthesia in infants and neonates. AB - The unbound and bound plasma concentration of bupivacaine in 50 infants less than 55 weeks postconceptual age was determined following combined spinal and epidural anaesthesia (csea). Plasma concentrations were determined at 15-min intervals up to 60 min postspinal anaesthesia. Maximum plasma bupivacaine levels were recorded between 45 and 60 min post CseA. Total plasma concentrations above a toxic threshold level of 4 microg.ml(-1) were recorded in 4% of patients and above 2.5 microg.ml(-1) in 10% of patients. Unbound bupivacaine levels were greater than a presumed toxic level of 0.25 microg.ml(-1) in 16% of cases and above 0.3 microg. ml(-1) in 14% of cases. A wide range of protein binding was measured (varying from 53.8-98.2%) and could not be correlated with standard indicators of local anaesthetic binding. Two neonates had brief apnoeas in the immediate perioperative phase but no adverse cardiac or central nervous system events attributable to the performance of Csea were demonstrated. PMID- 11119195 TI - Analgesia following paediatric day-surgical orchidopexy and herniotomy. AB - We surveyed 90 boys, aged 1-13 years, who had undergone either orchidopexy or herniotomy, in a cohort study. Their pain and vomiting were assessed using a simple 4-point score in the Recovery Unit by the nursing staff, and at home by the parents. There were no significant differences in pain or vomiting scores between the two groups in the immediate postoperative period. However, children having orchidopexy experienced more pain at home during the first night and the following day than those having herniotomy. Nearly one-third of the former group had moderate to severe pain at home, in contrast to less than one-tenth of children having herniotomy, who are also more likely to be painfree on the next day. We concluded that children having herniotomy can be treated adequately at home with paracetamol alone, whereas children having orchidopexy may require supplementation with stronger analgesics. PMID- 11119196 TI - Investigations using logistic regression models on the effect of the LMA on morphine induced vomiting after tonsillectomy. AB - The effect of intraoperative airway management on postoperative vomiting after tonsillectomy is unknown. Logistic regression was used in a retrospective study to investigate the effect of the laryngeal mask airway (LMA) on a morphine dose vomiting response curve. Charts were reviewed in 351 children in whom the airway was managed with either a LMA (n=177) or a tracheal tube (n=174). A mean perioperative morphine dose of 0.10 mg.kg(-1) (SD 0.09) was used in 248 children and a further 103 children were given no opioid. One hundred and eighteen of these 248 children vomited (47.6%) compared to 14 of 103 children given no morphine (13.6%). The probability of vomiting was related to morphine dose using logistic regression with both a linear and an E(max) model. Both the calibration (Hosmer-Lemishow goodness of fit chi-squared test lambda(2), P=0.81) and discrimination (area under the receiver operating characteristic plot, AUC ROC=0.67) of the E(max) model were better than the linear model (lambda(2), P=0.49; AUC ROC=0.64). Pharmacodynamic parameter estimates for the Emax model were P(0) (the baseline probability of vomiting) 0.139, P(max) (the maximal probability of vomiting due to morphine) 0.96, ED(50) (morphine dose that induces an effect equivalent to 50% of the logit P(max)) 0.09 mg.kg-1. The probability of vomiting was 50% after morphine 0.125 mg.kg-1. The use of the LMA had no effect on this dose-response curve. A covariate analysis investigating propofol for induction or isoflurane for the intraoperative maintenance of anaesthesia, however, showed that both drugs shifted the curve to the right. The probability of vomiting was 50% after morphine 0.17 mg.kg(-1) and 0.21 mg.kg(-1) for the isoflurane and propofol use curves, respectively. The concomitant use of propofol and isoflurane, but not the use of the LMA, decreases the probability of vomiting due to morphine. PMID- 11119197 TI - Vomiting and common paediatric surgery. AB - Postoperative vomiting is a common and unpleasant complication. The purpose of the present study was to verify if dexamethasone reduces the incidence of vomiting when injected IV in children anaesthetized with halothane for common paediatric operations. We also studied the incidence of vomiting when sevoflurane was used instead. Five hundred and 69 boys, aged 2-12 years (ASA physical status I, II), scheduled for inguinal field surgery were randomly assigned to receive halothane, halothane and dexamethasone and sevoflurane in three groups: halothane (n=180), halothane and IV dexamethasone (n=188) and sevoflurane (n=201). Anaesthesia was induced by inhalation of halothane or sevoflurane in oxygen and nitrous oxide and was maintained at minimum alveolar concentration of each agent throughout the surgery. For intra- and postoperative pain control iliac crest block was used in all the boys. Vomiting was defined as any expulsion of liquid gastric contents. The incidence of postoperative vomiting was 23% in the halothane group, which was significantly greater than that in the other groups (halothane and dexamethasone group, 9%; sevoflurane group, 13%). In conclusion, dexamethasone reduces the incidence and frequency of multiple emetic episodes when administered intravenously after halothane anaesthesia; sevoflurane reduces the overall incidence of vomiting, but not multiple emetic episodes. PMID- 11119199 TI - Assessment of autonomic cardiovascular changes associated with recovery from anaesthesia in children: a study using spectral analysis of blood pressure and heart rate variability. AB - Recovery from anaesthesia is associated with large changes in cardiovascular autonomic activity, which are poorly documented in children. This study was undertaken to investigate the cardiovascular autonomic activity in anaesthetized and recovering children, using a noninvasive approach based on spectral analysis of heart rate (HR) and blood pressure (BP) variability. Ten children (aged 5-13 years) undergoing major surgery were studied. Continuous HR and BP were recorded using a noninvasive device during deep anaesthesia and recovery. Spectral analysis was used to determine the main oscillatory components of HR and BP signals. For each power spectrum, the frequency components were identified as follows (i): the low frequency (LF) component (0.04-0.14 Hz) both parasympathetically and sympathetically mediated for HR and corresponding to vasomotor sympathetic modulation for BP; and (ii) the high frequency (HF) component (0.2-0.6 Hz) parasympathetically mediated for HR, and reflecting mechanical influence of ventilation on cardiac output for BP. In addition, the LF : HF ratio for HR, reflecting the cardiac sympathovagal balance, was calculated. Under deep anaesthesia, HR variability and BP variability were very low and mainly due to mechanical influence of intermittent positive pressure ventilation. Conversely, the recovery period was associated with a marked increase of HR and BP overall variability. Compared to anaesthesia, spectral analysis of HR and BP revealed that the LF component of BP and HR spectra increased 40-fold during recovery; the LF : HF ratio of HR was also increased during recovery (0.1 +/- 0.1 versus 1.3 +/- 1.2, P=0.008). The results of this study demonstrate that the recovery period is associated with an increase of cardiovascular sympathetic drive in children after major surgery. PMID- 11119198 TI - Diclofenac and flurbiprofen with or without clonidine for postoperative analgesia in children undergoing elective ophthalmological surgery. AB - We conducted a prospective, randomized study to compare the efficacy of preoperative diclofenac, flurbiprofen, and clonidine, given alone, as well as the combination of diclofenac and clonidine, and flurbiprofen and clonidine in controlling postoperative pain in 125 children. The patients (ASA I, 2-12 years) undergoing elective ophthalmological surgery were allocated to one of five groups: rectal diclofenac 2 mg.kg(-1) following oral placebo premedication, i. v. flurbiprofen 1 mg.kg(-1) following placebo premedication, oral clonidine premedication, rectal diclofenac 2 mg.kg(-1) following clonidine, and i.v. flurbiprofen 1 mg.kg(-1) following clonidine. The children received clonidine (4 microg.kg(-1)) or placebo 105 min before anaesthesia. Diclofenac or flurbiprofen was given immediately after induction of anaesthesia. Anaesthesia was induced and maintained with sevoflurane and nitrous oxide in oxygen. Postoperative pain was assessed by a blinded observer using a modified objective pain scale (OPS). No opioids were administered throughout the study. Rectal diclofenac 2 mg.kg(-1) i.v. flurbiprofen 1 mg.kg(-1), oral clonidine 4 microg.kg(-1) provided similar OPS scores and requirement for supplementary analgesics during 12 h after surgery. Combination of oral clonidine and one of these nonsteroidal analgesics minimized postoperative pain. Our findings suggest that this combined regimen may be a promising prophylactic approach to postoperative pain control in children undergoing ophthalmological surgery. PMID- 11119200 TI - Fatal complication from central venous cannulation in a paediatric liver transplant patient. AB - We report a fatal complication from central venous cannulation in a child undergoing heterotropic liver transplantation. Following the attempted placement of a cannula in the left internal jugular vein, extravasation of blood products via the cannula resulted in haemothorax, hypotension and eventual brain death. Possible causes and strategies for prevention of this complication are discussed. PMID- 11119201 TI - An unusual complication of a central venous catheter in a neonate. AB - The use of central venous catheters in neonates is associated with early and late complications. It is recognized that catheter tip migration and perforation of a viscus can occur at any time with a potentially fatal outcome. We present a case in which the successful placement of a central line was followed 2 weeks later by a sudden respiratory deterioration necessitating intubation and ventilation. The catheter tip had eroded through the wall of a pulmonary artery and a bronchus into the bronchial tree. The report highlights the serious morbidity arising from the use of central venous lines in neonates and stresses the importance of X-rays in establishing the correct position of all catheters. A sudden change in the condition of a patient should raise the suspicion of a catheter-related problem. PMID- 11119202 TI - Perioperative management of infants with the linear naevus sebaceous syndrome of Jadassohn: a report of two cases. AB - Neurofibromatosis and tuberous sclerosis are the most well-recognized of the congential phakomatoses, a group of six hereditary neuro-oculo-cutaneous disorders. Although easily diagnosed at birth by a parasagittal line of facial sebaceous naevi, the linear naevus sebaceous syndrome (LNSS) of Jadassohn is the rarest phakomatosis, one often characterized by airway and anaesthetic considerations that do not apply to the other phakomatoses. In addition to its obvious cutaneous manifestations, LNSS is characterized by hemifacial asymmetry, an anatomic predictor of difficult trachael intubation, and intractable seizure activity, a condition that limits selection of anaesthetics. The perioperative management challenges of LNSS are depicted in the presentation of two cases of LNSS with different outcomes and contrasted with the major anaesthetic considerations in the perioperative management of other, more common phakomatoses. PMID- 11119203 TI - An alternative for avoidance of general anaesthesia for infants when bilateral inguinal herniorrhaphy outlasts subarachnoid blockade. AB - Former premature infants represent a high risk surgical population. In order to minimize the risk of postoperative apnoea, subarachnoid blockade without sedation is known to be preferable to general anaesthesia for former premature infants undergoing bilateral inguinal herniorrhaphy. However, subarachnoid blockade affords only a limited duration of reliable anaesthesia. Nonroutine surgical delays and technical difficulties cannot always be anticipated by the anaesthesiologist. When bilateral inguinal herniorrhaphy outlasts the anticipated duration of subarachnoid blockade, the anaesthesiologist is confronted with a dilemma. Infants are unable to complain verbally, so the extent of subarachnoid blockade may be difficult to assess intraoperatively. Introduction of sedation or general anaesthesia under these circumstances increases the risk of postoperative apnoea, thereby defeating the purpose of the original choice of anaesthesia. Several alternatives have been proposed, but all involve disadvantages. In this report of two cases, a new solution to this clinical dilemma is presented. PMID- 11119204 TI - Anaesthesiological considerations in patients with Sneddon's syndrome. AB - Sneddon's syndrome is a rare disease with strong gender prevalence of females. This syndrome is characterized by livedo racemosa and cerebrovascular lesions. Since no specific test is available, the clinical differentiation from other disorders with similar symptomatology may raise difficulties. The cerebral involvement includes strokes with cases of more than one ischaemic event having been reported. Associations with convulsions, heart valve disease, systemic hypertension, and renal impairment have been described. We report the case of a 5 year-old boy who was anaesthesized for dental surgery. Due to the fact that 50% of Sneddon's syndrome patients develop mental retardation, even minor procedures require general anaesthesia. A review of the literature is added and specific anaesthesiological aspects of the perioperative care of Sneddon's syndrome are discussed. PMID- 11119205 TI - Subcutaneous tunnelled epidural catheters in children undergoing hip surgery. PMID- 11119206 TI - Authors' reply PMID- 11119207 TI - Fibreoptic intubation for massive gingival hyperplasia in juvenile hyaline fibromatosis. PMID- 11119208 TI - A dose of 1 mg.kg(-1) meperidine causes muscle rigidity in infants? PMID- 11119209 TI - Low gastric luminal pCO(2) due to a malmanufactured tonometer catheter. PMID- 11119211 TI - Acknowledgements PMID- 11119210 TI - The developing role of play preparation in paediatric anaesthesia. PMID- 11119212 TI - The analgesic efficacy of preoperative high dose (40 mg.kg-1) oral paracetamol after bilateral myringotomy and grommet insertion in toddlers PMID- 11119213 TI - What is the incidence of leg weakness after ilio-inguinal block in children? PMID- 11119215 TI - Delegation in paeditric anaesthesia; a postal survey of APA members PMID- 11119214 TI - The size 1(1/2) laryngeal mask airway in paediatric anaesthetic practice PMID- 11119216 TI - An evaluation of propofol combined with remifentanil: a new intravenous anaesthetic technique for short painful procedures in children PMID- 11119217 TI - A comparison of the respiratory effects of high concentrations of halothane and sevoflurane in children PMID- 11119218 TI - Metabolic stress responses in postoperative children aged 0-3 years PMID- 11119219 TI - A study to assess the feasibility of using glucometer measurement of blood glucose on fresh blood sampled from the surgical field PMID- 11119220 TI - A survey of the usage of caudal catheters amongst paediatric anaesthetists practising in the UK PMID- 11119221 TI - The effect of varying resistance or compliance on the movement of liquid during high frequency oscillation PMID- 11119222 TI - Changes in pulmonary mechanics in anaesthetized infants: treatment by a volume recruitment manoeuvre PMID- 11119223 TI - The APLS guidelines for paediatric endotracheal tube selection - how accurate are they and do they need updating? PMID- 11119224 TI - Pharmacokinetic profile of rectally administered diclofenac sodium in children undergoing adenotonsillectomy PMID- 11119225 TI - Cisatracurium pharmacokinetics and pharmacodynamics during hypothermic CPB in infants PMID- 11119226 TI - Is polysomnography predictive of respiratory complications post adenotonsillectomy in children? PMID- 11119228 TI - Review of paediatric trauma admissions at the royal london hospital (February 98 to february 99) PMID- 11119227 TI - Post sigh apnoea represents the majority of central apnoea in infants at risk for postoperative apnoea PMID- 11119229 TI - A comparison of oral transmucosal fentanyl and oral midazolam for premedication in children PMID- 11119230 TI - Intraoperative fentanyl reduces early vomiting after paediatric tonsillectomy compared with morphine PMID- 11119231 TI - An audit of serious complications during neurosurgery in children using the sitting position PMID- 11119232 TI - Infusions of local anaesthetic via caudal catheters in neonates and small infants for analgesia after major surgery PMID- 11119233 TI - Anaesthesia for young children: an audit of practices at poole general hospital, january 1997 to january 1999 PMID- 11119234 TI - The effect of induced hypothermia on the offset time of atracurium when given by infusion to critically ill children PMID- 11119236 TI - Caudal ropivacaine in infants: population pharmacokinetics and plasma concentrations PMID- 11119235 TI - Postoperative behavioural changes in children: the influence of sevoflurane PMID- 11119237 TI - Paediatric trauma in a district hospital: three cases of small bowel injury PMID- 11119238 TI - Sedation and restraint practices in UK paediatric intensive cares: a telephone survey PMID- 11119239 TI - An audit of the last seven years of paediatric anaesthetic airway problems in patients on the birmingham bone anchored hearing aid (BAHA) programme PMID- 11119240 TI - Thoraco-abdominal asynchrony in anaesthetized children affected by airway manoeuvres PMID- 11119241 TI - Recall following paediatric intensive care PMID- 11119242 TI - Caudal additives to ropivacaine in children: preservative free S-ketamine versus clonidine PMID- 11119243 TI - Serial lactate measurements during surgery involving deep hypothermic circulatory arrest PMID- 11119244 TI - Pax-5 and EBF are expressed in committed B-cell progenitors prior to the colonization of the embryonic bursa of fabricius. AB - The committed B-cell precursors developing from hemopoietic stem cells have been considered to differentiate through a common lymphoid progenitor stage in the mouse. In the chicken B-cell system, however, the committed B-cell progenitors burst as a single wave prior to the bursal colonization and most likely as direct descendants of hemopoietic stem cells. In the present report we show that prebursally committed B-cell progenitors specifically express early B-cell factor (EBF) and Pax-5 transcription factors. In addition we show that the expression of these and other B-lineage-associated transcription factors starts early in the chicken ontogeny. Altogether our findings strongly support the model of early delineation of B- and T-cell development and do not support the existence of common lymphoid stem cells. PMID- 11119245 TI - Role of pre-T cells and chemoattractants on stress-associated thymus involution. AB - Male C57BL/6 mice were stressed by immobilization for 1, 2, 3, or 5 h per day for 14 days, with subsequent assessment of (a) thymic involution, (b) in vitro migration of stressed mice bone marrow cells toward thymocyte culture supernatants from neonates and from control or stressed mice, (c) composition of the bone marrow cell population, and (d) in vitro migration of normal bone marrow cells toward stressed mice thymocyte culture supernatants. The results obtained support the view that the reduced repopulation of thymus by precursor T cells contributes to thymus involution associated with stress. It is further shown that this effect could be owing to a reduction in the number of precursor T cells in the bone marrow, and/or to a diminished production of precursor T-cell chemoattractants. PMID- 11119246 TI - Longevity of immune complexes and abnormal germinal centre formation in NZB mice. AB - Rheumatoid factor (RF)-like (antibody-antibody) immune complexes induce a selective and intensive immunoglobulin (Ig)G1-RF response after a single injection in mice. However, the longevity of the response differs between mouse strains: serum IgG1-RF antibody titres decline 40 days after injection in C57Bl/6 mice whereas levels are maintained for more than 100 days in NZB mice. In order to elucidate whether this difference was owing to a lower ability of NZB mice to clear the injected immune complexes, sections of kidney, spleen, liver and mesenteric lymph nodes were harvested at different time points after injection with RF-like immune complexes. Immunohistochemical staining revealed that NZB mice have a delayed clearance of the injected immune complexes, because the immune complexes are retained for more than 40 days in their spleens and 100 days in their kidneys, compared to only 14 days in C57Bl/6 mice. Germinal centres were also present for a longer period in the spleens of the NZB mice, accompanying the presence of the immune complexes, and were abnormally large compared to C57Bl/6 mice. The clearance of immune complexes from the spleen coincided with the decline in serum levels of IgG1-RF, indicating that prolonged retention of immune complexes is responsible for the sustained IgG1-RF response. PMID- 11119247 TI - Interaction between human complement and a pectin type polysaccharide fraction, PMII, from the leaves of Plantago major L. AB - The interaction between a pectin type polysaccharide fraction, PMII, isolated from the leaves of Plantago major, and human complement was tested in two different hemolytic complement-fixation tests and in addition by two ELISA methods detecting complement-activation products. Sera were used as a complement source of 10 arbitrary human volunteers, individually and as a pool. The complement-fixation tests were designed to measure the concentration of the pectin necessary to inhibit 50% of the hemolysis (ICH(50)). The ELISA tests for complement-activation products were measured in AU/mg using a fully activated serum as a standard. We observed a more than 200-fold difference in ICH(50) activity of the PMII pectin in one of the hemolytic tests by varying the individual sera used as complement-source. On the other hand, the ELISA complement-activation tests showed no significant variation in activity of the PMII depending on the complement-serum used. The level of antibodies against PMII detected in the complement-sera did not correlate with the ICH(50) activity of PMII. The results show that PMII is a potent complement activator with an activity of the same order of magnitude on a weight basis as that of aggregated human immunoglobulin (Ig)G. This activation leads to a complement consumption probably explaining the PMII's effect in the complement-fixation tests. PMII seems to be an activator both on the classical and the alternative pathway of activation. The results might be related to the reported wound-healing effect of the leaves of Plantago major. PMID- 11119248 TI - Garlic induces a shift in cytokine pattern in Leishmania major-infected BALB/c mice. AB - The regulation of T helper (Th)1- and Th2-type cytokine patterns is important in the final outcome of leishmaniasis in human and murine models. We examined the efficacy of garlic therapy or a combination of garlic and an antimonial drug (glucantime) in promoting healing and regulation of Th1/Th2 cytokine patterns in highly susceptible BALB/c mice infected with Leishmania major. Separate groups of infected mice received 20 mg/kg/day garlic, 60 mg/kg/day glucantime or a combination of the two, from day 30 after infection for 2 weeks. An enzyme-linked immunosorbant assay (ELISA) was performed on spleen cell culture supernatants for interferon(IFN)-gamma interleukin(IL)-2, IL-4 and IL-10. The results indicate that garlic therapy is more effective than the usual antileishmanial drug in curing the infection. Garlic-treated mice developed Th1-type cytokine responses. In contrast, glucantime therapy led to a Th2-type response in the control group with a lower level of IL-2. However, a combination of garlic and glucantime treatment was more effective than either treatment alone, and resulted in a Th1 type response similar to that which developed with garlic treatment. These results suggest that garlic extract in combination with an antimonial drug, may provide effective therapy against L. major. The immunomodulatory properties of garlic were elucidated in terms of shifting the cytokine response to a Th1-type pattern and therefore causing the protective response. PMID- 11119249 TI - Spontaneous apoptosis of neutrophils in whole blood and its relation to apoptosis gene proteins. AB - Apoptosis of neutrophils limits their pro-inflammatory potential. We tested the ability of fresh and cultured whole blood neutrophils to undergo spontaneous apoptosis and expression of p53, Fas/Apo-1, bcl-2 protein in the cells using flow cytometry. Neutrophil apoptosis was estimated using Annexin V and propidium iodide binding and verified under light microscopy. The percentage of early and late apoptotic neutrophils in the blood samples increased significantly after 20 h culture from 12.3 +/- 14.2% and 4.3 +/- 4.2% to 39.5 +/- 14% and 15.3 +/- 9.6%, respectively. The majority of late apoptotic neutrophils had altered morphology in FSC/SSC dot plot compared to alive or early apoptotic neutrophils. Cultured neutrophils presented markedly lower expression of bcl-2 protein compared to fresh blood cells: 211 +/- 321 median of fluorescence intensity (MFI) and 787 +/- 1152 MFI, respectively. The increased percentage of late apoptotic cells after culture paralleled the increase in the Fas/Apo-1 expression and negatively correlated with bcl-2 expression. We noted intracellular expression of p53 protein in neutrophils, although the expression did not correlate neither to the percentage of the apoptotic neutrophils, nor to the Fas/Apo-1 or bcl-2 expression. Our results suggested that neutrophil apoptosis is gene regulated, moreover, we present a possibility to assess the neutrophil apoptosis and cellular expression of the proteins of apoptosis related genes in whole blood samples. PMID- 11119250 TI - Prethymic progenitors from the avian para-aortic mesoderm express GATA-3 and distinct chTcf isoforms but still lack T-cell receptor-gamma rearrangements. AB - Haematopoietic precursors first colonizing the avian embryonic thymus are derived from the intraembryonic sites located around the dorsal aortae. These intraembryonic precursors have previously been demonstrated to include cells that harbour T-cell progenitor capacity and express the Ikaros transcription factor, known to be a prerequisite for lymphocyte development. In this study, we further evaluated the properties of these prethymic cells. We show that early intraembryonic cells and prethymic progenitors already express the GATA-3 transcription factor. The chicken homologue of T-cell factor-1, chTcf, is also detected in cells isolated from the avian para-aortic region. However, these intraembryonic cells retain their T-cell receptor gamma loci in germline configuration. Interestingly, chTcf was found to express different alternatively spliced isoforms during early ontogeny and thymic T-cell development, which indicates developmentally regulated expression of chTcf variants. Taken together, these results demonstrate that, although the avian prethymic progenitor cells express T-lineage-associated transcription factors, they have not yet undergone TCR rearrangements. It is therefore suggested that activation of lineage associated genes is an early event in the generation of haematopoietic progenitor cells during ontogeny. PMID- 11119251 TI - No evidence for generation of Th-2-like MBP-specific T-cell lines by blockade of the costimulatory molecule B7-1. AB - The T helper-1 (Th-1)/T helper-2 (Th-2) paradigm is relevant for the pathogenesis and therapy of multiple sclerosis. In experimental autoimmune encephalomyelitis, a shift towards a Th-2 immune response serves as treatment of the disease. In the human immune system, the factors which determine and modulate the differentiation of CD4+ T cells into the Th-1 or Th-2 phenotype have yet to be elucidated completely. Here, the split-well approach was used to analyse costimulatory requirements for the generation of myelin basic protein-specific T-cell subsets considered to play a major role in the pathogenesis of multiple sclerosis. Myelin basic protein-specific T-cell lines were isolated from peripheral blood cells of healthy individuals in the presence or absence of a blockade of the costimulatory molecule B7-1, previously reported to be involved in the development of Th-1 cells. T-helper type was determined by the interferon/interleukin ratio. Blockade of B7-1 did not increase the number of Th-2-like myelin basic protein-specific T cell lines. Thus, these data show no evidence for an influence of B7-1 blockade on the development of human myelin basic protein-specific T-cell subsets. These results have to be taken into account when discussing whether antibody-mediated B7-1 blockade might be a suitable therapy in multiple sclerosis, as demonstrated in experimental autoimmune encephalomyelitis. PMID- 11119252 TI - Th1-specific bystander costimulation imparts resistance against Mycobacterium tuberculosis infection. AB - The protection against Mycobacterium tuberculosis infection is mediated by T helper type-1 (Th1) cells. Infection of BALB/c mice with M. tuberculosis downregulates expression of a Th1-specific costimulatory molecule, M150, on the surface of infected macrophages. The proliferation of Th cells and Th1-cytokine production by these cells are higher in case of M. tuberculosis antigen presentation by uninfected macrophages than by infected macrophages. The difference in inducing interleukin(IL)-2 and interferon (IFN)-gamma secretion is abolished by providing bystander costimulation through M150 on liposomes. PMID- 11119254 TI - Editorial PMID- 11119253 TI - Interleukin-10 is an unequivocal Th2 parameter in the rat, whereas interleukin-4 is not. AB - Exposure of Wistar rats to the immunotoxic compounds hexachlorobenzene (HCB), bis(tri-n-butyltin)oxide, and benzo(a)pyrene was previously found to affect mRNA expression of interleukin (IL)-2, IL-2R alpha-chain, and interferon (IFN)-gamma, the prototypic Th1 cytokine. In contrast, the mRNA expression of IL-4, the prototypic Th2 cytokine, was unaffected. This latter finding suggested that the IL-4 mRNA expression may not be an unequivocal parameter for Th2 responses in the rat. In order to obtain such a parameter the present study was performed, consisting of two types of experiments. Expression and production of IL-4 as well as IL-10, a second Th2 cytokine, were measured. First, Lewis (Th1 prone) and Brown Norway (BN; Th2 prone) rats were exposed to HCB. Exposure was previously found to increase the serum immunoglobulin (Ig)E levels, an IL-4-dependent response, in BN but not Lewis rats, and in Lewis rats to aggravate experimental allergic encephalomyelitis (EAE), severity being inversely related to IL-10 levels. Secondly, BN rats were infected with Trichinella spiralis, an infection previously found to induce IL-4 production. HCB exposure did not affect IL-4 mRNA expression in either strain, while IL-4 production was decreased in Lewis and unaffected in BN rats. In Lewis rats both the mRNA expression and the production of IL-10 were decreased. The T. spiralis infection induced IL-4 and IL-10 mRNA expression, as well as IL-10 production. In contrast, the IL-4 production was strongly reduced. Thus, both the IL-10 mRNA expression and production correlated with the EAE development and T. spiralis infection. In HCB exposed Lewis rats and T. spiralis infected BN rats the IL-4 mRNA expression correlated with IgE levels and T. spiralis infection, respectively, whereas the IL-4 production lacked correlation in all cases. Collectively, these results suggest that IL-10 is an unequivocal Th2 parameter in the rat, whereas IL-4 is not. PMID- 11119255 TI - Genomic organization and regulation of the human interleukin-18 gene. AB - The human interleukin(IL)-18 is a key regulator of interferon(IFN)-gamma production and T-cell differentiation. Here we report the complete genomic structure and characterization of the 5'untranslated promoter region of the human IL-18 gene. The gene is composed of six exons and five introns, spanning approximately 19. 5kb. Promoter activity of the 5'-flanking region was investigated with a luciferase reporter gene assay. Transient transfection studies demonstrate a constitutive expression of the IL-18 gene in monocytic U937 and THP-1 cells. For this constitutive expression at least 92 base pairs of the promoter region are essential as shown by consecutive 5' promoter deletions in both cell types. DNA protein binding experiments revealed specific binding of activated signal transducer and activator of transcription factor-5 (STAT5) but not of STAT3 to three consensus sequences upstream in the promoter region. Cotransfection of STAT5 resulted in increased induction of the IL-18 promoter in the U937 and THP-1 cells. PMID- 11119256 TI - Effects of interleukin-12 in the long-term protection conferred by a Mycobacterium avium subunit vaccine. AB - The effects of the addition of recombinant interleukin (IL)-12 to a mycobacterial subunit vaccine were analyzed in terms of the longevity of the protective immunity generated. BALB/c mice were immunized with culture filtrate proteins from Mycobacterium avium with dimethyl-dioctadecilammonium bromide (DDA) as an adjuvant. This subunit vaccine induced protection against a challenge by M. avium which lasted for at least 6 months while waning with time until 1 year postvaccination. Whereas the addition of IL-12 enhanced the initial protective efficacy of this subunit vaccine during the first 6 months, it accelerated the loss of protective efficacy observed at 1 year postvaccination. These data confirm the adjuvant properties of IL-12 in vaccines against mycobacteria and raise the possibility of late counter-protective untoward effects. PMID- 11119257 TI - Cytokines involved in Toxoplasmic encephalitis. AB - Reactivation of cerebral toxoplasmosis occurs in approximately 30% of acquired immune deficiency syndrome (AIDS) patients who are seropositive for Toxoplasma gondii and a change in the levels of cytokines during this relapse is observed. Several cytokines are able to initiate meningeal inflammation and may play a role in the immunopathogenesis of the disease. The induction of a type 1 inflammatory cytokine response is a key event in the initiation of immunity to T. gondii. Interleukin (IL)-10 production in infected brain facilitates the persistence of parasites by down-regulating the intracerebral immune response. The in vivo and in vitro models are very numerous and this may explain the heterogeneity of the results. The role of gamma interferon is important because it is the principal mediator inducing a host resistance against T. gondii. Several cytokines stimulating or decreasing interferon (IFN)-gamma levels are reported. The particular case of AIDS patients whose humoral response is altered, is studied. PMID- 11119258 TI - Functional characterization of CD8(+) antigen-specific cytotoxic T lymphocytes after enrichment based on cytokine secretion: comparison with the MHC-tetramer technology. AB - Cell therapy with antigen-specific T cells holds promise for various diseases including cancer and viral infections. The powerful enrichment procedure based on major histocompatibility complex (MHC)-tetramers, however, is of limited applicability so far. Therefore, the recently developed cell surface affinity matrix technology that allows direct identification and enrichment of life antigen-specific T cells based on cytokine secretion was evaluated in this respect. To this end, CD8(+) T cells directed against the HLA-A(*)0201-restricted melanoma-associated peptide Melan-A (aa26-35) were generated by combining stimulation of peptide-pulsed autologous dendritic cells (DC) with antigen independent expansion with anti-CD3/anti-CD28 monoclonal antibodies (MoAb). Antigen-specific cytotoxic T lymphocyte (CTL) were detected based on stimulation induced interferon (IFN)-gamma and interleukin (IL)-4 secretion and enriched > 100-fold using the cell surface affinity matrix technology. The resulting IFN gamma- and IL-4-secreting CTL lines contained > 80% and > 70% cytokine positive T cells, respectively. They exhibited a cytotoxic activity against Melan-A expressing target cells that was significantly higher as compared to nonpurified CTL. Direct staining of enriched CTL with HLA-A2-Melan-A-tetramers revealed a high correlation between the results obtained from the cell surface affinity matrix technology and those obtained from tetrameric complexes. Altogether, our study demonstrates that cytokine-driven enrichment based on the cell surface affinity matrix technology enables selective isolation of functionally active antigen-specific CTL that may be used for an adoptive T cell transfer in immunotherapy. PMID- 11119259 TI - Synchronous deletion of Mtv-superantigen-reactive thymocytes in the CD3(medium/high) CD4(+)CD8(+) subset. AB - Multiple model systems have demonstrated that negatively selected thymocytes can be deleted during the immature CD4(+)CD8(+) CD3(low) stage after high affinity interaction of T-cell receptors (TCRs) with antigen:major histocompatibility complex (MHC) complexes. Superantigens (SAGs) derived from endogenous mammary tumour viruses (Mtv) induce negative selection of Mtv-SAG-reactive thymocytes regardless of which peptide antigen is presented by MHC molecules. In this study, the timing of deletion of multiple subsets of Mtv-SAG-reactive CD4(+)CD8(+) thymocytes was investigated by a 4 colour flow cytometry in SJL x CBA/J cross bred mice. Deletion of V beta 3(+), V beta 5(+), V beta 11(+), and V beta 17(+) Mtv-SAG-reactive thymocytes was found to occur synchronously in the most mature CD3(medium) and early CD3(high) subsets of CD4(+)CD8(+) thymocytes, in contrast with reports showing that the deletion of Mtv-SAG-reactive thymocytes can occur at different stages in particular model systems. PMID- 11119260 TI - Alterations during positive selection in the thymus of nackt CD4-deficient mice. AB - The T-cell repertoire is shaped by the positive and negative selection of immature CD4(+) CD8(+) double positive (DP) thymocytes. Positive selection of DP T cells to the CD4(+) CD8(-) and CD4(-) CD8(+) simple positive (SP) lineages is a multistep process which involves cellular interactions between thymocytes and stromal cells. Mutant nackt (nkt/nkt) mice have been shown to have a deficiency in the CD4(+) CD8(-) T-cell subset both in the thymus and in the periphery. The present report suggests that nkt/nkt mice present alterations in early steps of positive selection because they show decreases in the percentages of CD69(+) and CD5(+) cells within the DP subset. Experiments involving bone marrow transfer and thymic chimeras demonstrate that the thymic epithelium of nkt/nkt mice is involved in the alterations registered during positive selection and dictates the ultimate fate of CD4(+) SP cells. PMID- 11119261 TI - Immune complex-mediated enhancement of antibody responses without induction of delayed-type hypersensitivity. AB - Immunoglobulin (Ig)G and IgE antibodies enhance the humoral response in vivo to soluble antigens with which they form complexes. In vitro, antigen is targeted to B cells by IgE antibodies and to macrophages and dendritic cells (DCs) by IgG, thus leading to increased antigen presentation to specific T cells. Possibly these mechanisms are also responsible for antibody-mediated enhancement in vivo. We now address the question of whether IgG- and/or IgE-antigen complexes can prime for delayed-type hypersensitivity (DTH), a reaction known to require primed T helper (Th)1 cells. Mice were immunized with IgG-anti-2,4,6-trinitrophenyl (TNP)/BSA-TNP or IgE-anti-TNP/BSA-TNP. Mice given BSA-TNP alone or BSA-TNP in complete Freund's adjuvans (CFA) were used as controls. DTH and IgG-anti-BSA levels were measured after subsequent challenge with BSA. A potent BSA-specific antibody response was induced by IgE- or IgG-complexed antigen as well as by CFA/antigen but DTH-reactions were only observed in mice immunized with CFA/antigen. Both IgE and IgG enhanced the production of BSA-specific IgG1, IgG2a and IgG2b, although the most pronounced enhancement was seen in the production of IgG1. These findings suggest that Th2 cells rather than Th1 cells are involved in the immune response to IgG- and IgE-immune complexes. PMID- 11119262 TI - Enprostil, a prostaglandin-E(2) analogue, inhibits interleukin-8 production of human colonic epithelial cell lines. AB - We previously reported that intracolonic administration of enprostil, a prostaglandin-E(2) (PGE(2)) analogue, had therapeutic effects on acute colitis induced in rodents by dextran sulfate sodium (DSS). In addition, production of growth-regulated gene product/cytokine-induced neutrophil chemoattractant-1 [GRO/CINC-1; an interleukin(IL)-8 like cytokine] was suppressed in the inflamed tissues. In the present study we used a human colon cancer cell line (HT-29) to investigate enprostil effects on the IL-8 production of intestinal epithelial cells stimulated by various stimulants. In a MTT assay, concentrations of enprostil >10(-5)M had cytotoxitic effects on HT-29 cells. Furthermore, 10(-6) M enprostil suppressed IL-8 production in HT-29 cells, SW620 and CaCo2 stimulated with interleukin-1 beta (IL-1 beta) or lipopolysaccharide (LPS), but did not suppress this response when cells were stimulated with tumour necrosis factor (TNF)-alpha. These results suggest that enprostil affects a point in the pathway between the IL-1 receptor or LPS receptor and nuclear factor-kappa B(NF-kappa B), without affecting the pathway between the TNF receptor and NF-kappa B, with the latter factor being required for the IL-8 gene transcription. The therapeutic effect of exogenous enprostil on DSS colitis may involve the inhibition of IL-8 production in colonic epithelial cells stimulated by IL-1 beta or LPS. PMID- 11119263 TI - Characterization of proteoglycans and glycosaminoglycans in splenic AA amyloid induced in mink. AB - Amyloidosis of the protein AA type is readily induced in mink using repeated injections of bacterial lipopolysaccharide (LPS). We have characterized splenic proteoglycans/glycosaminoglycans (PGs/GAGs) in mink during amyloidogenesis. Moderate to rich amounts of amyloid exhibiting green birefringence was demonstrated by polarization microscopy of the splenic section stained with Congo red in seven out of eight minks after 10 weeks of LPS-treatment, and a significant increase in the total amount of PGs and GAGs in AA amyloid spleens was observed (two to eight times that in unstimulated animals). Intact PGs as well as free GAGs were extracted, and heparan sulfate (HS) was the most abundant GAG in the amyloid as well as in the control spleens. The GAGs showing the most pronounced increase in the amyloid spleens was of the chondroitin sulfate/dermatan sulfate (CS/DS) type and these were extracted in the form of free GAG chains. We conclude that there is a selective enrichment of PGs/GAGs in extracted splenic amyloid in the mink, which confirms to previous observations in human amyloid as well as in other animal species, supporting their pathogenic significance in the formation of AA amyloid. PMID- 11119264 TI - Molecular weight dependency on the production of the TNF stimulated by fractions of rye (1-->3),(1-->4)-beta-D-glucan. AB - Mixed-linkage (1-->3),(1-->4)-beta-D-glucan with a weight average molecular weight varying between 79,800 and 13,900 was purified from rye. These fractions were used for stimulation of human monocytes to produce tumour necrosis factor (TNF). A mixed-linkage beta-glucan with a weight average molecular weight of 18,900 was found to be the most potent immunostimulator. PMID- 11119265 TI - Translocation of ribosomal immunostimulant through an in vitro-reconstituted digestive barrier containing M-like cells. AB - Ribosomal preparations of pathogenic micro-organisms of the upper respiratory tract can be delivered orally for the prevention of recurrent infectious episodes, because they induce mucosal and protective immune responses. The mechanism of mucosal barrier translocation is difficult to study in animal models, little is therefore known about this process. In order to circumvent these problems, we have examined the uptake of ribosomal preparations in three experimental systems that model human intestinal cells. We found that M-like cells displayed a 8.7-fold increase in the uptake of a ribosomal immunostimulant when compared to absorptive or crypt enterocyte-like cells. The product was taken up, translocated, and delivered in the basolateral compartment by cultured M-like cells. No translocation was observed across monolayers of T84 cells (model of crypt cells). Only minimal translocation occured through monolayers of Caco-2 cells (model of absorptive enterocytes). This suggests that, in vivo, colyophilisat is delivered mainly through the M cells overlying lymphoid follicles (Peyer's patches) or nodules of the gut-associated mucosal lymphoid tissue, which are the major inductor sites of mucosal responses. Use of the M like cell cultured model could be a key step for the development of even more efficient immunostimulators in animals and human. PMID- 11119266 TI - Crosslinking of CD30 on activated human Th clones enhances their cytokine production and downregulates the CD30 expression. AB - Signalling through CD30 has been shown to mediate pleiotropic effects, depending on the type of target cell. In the present study, we have used the agonistic anti CD30 monoclonal antibodies (MoAb) M44 to study phenotypic changes in human T-cell clones of Th1 and Th2 type. Alterations in the surface expression of CD30, CD28 and CD40L following CD30 stimulation were analyzed after 24 h, and the cytokine production after CD30 crosslinking was measured at 48 h. We observed a clear reduction of surface expression of CD30 after treatment with the M44 MoAb. Our results also indicate that CD28 is significantly down modulated in the Th2 clones after CD30 crosslinking (P < 0.05, n = 5) whereas no apparent alteration was observed in the expression of CD40L. When the concentration of cytokines was measured in the supernatants after CD30 stimulation, elevated levels of interleukin (IL)-4 and IL-5 were observed in the Th2 clones, and elevated levels of interferon (IFN)-gamma in the Th1 clones. The enhanced cytokine production after CD30 crosslinking supports the presumption of CD30 functions as a positive regulator in activated T cells. PMID- 11119267 TI - Increase in T-cell subsets of oral mucosa: a late immune response in patients with treated coeliac disease? AB - BACKGROUND AND AIMS: In coeliac disease, the gut involvement is gluten-dependent. Following the introduction of a gluten-free diet, inflammatory cell infiltration decreases in the small intestinal mucosa. Our hypothesis was that the oral mucosa might mirror the changes found in coeliac disease similarly to the mucosa of the small intestine. Thus, the number of inflammatory cells in the oral mucosa would decrease in patients with coeliac disease on a gluten-free diet. METHODS: The distribution CD45RO+ and CD3(+) T cells, T-cell subpopulations (CD4(+), CD8(+), T cell receptor (TCR)alpha beta+ and TCR gamma delta+ cells) and HLA DR expression were studied in the buccal mucosa of 15 untreated and 44 gluten-free diet treated coeliac disease patients, and of 19 controls. All 15 patients with untreated coeliac disease were immunglobulin (Ig)A endomysial antibody positive and all 44 patients on gluten-free diet except one were endomysial antibody negative, as were all control subjects. RESULTS: Untreated coeliac disease patients did not differ from controls in the densities of CD45RO+ cells, CD3(+) cells or of T-cell subsets. In contrast, in treated coeliac disease patients, a significant increase in the numbers of mast cells, CD3(+) and CD4(+) lymphocytes was found in the lamina propria of oral mucosa as compared with patients with untreated coeliac disease and controls. The increase in CD3(+) T cells was in part owing to an increase in lymphocytes expressing no TCR. No differences were found in the expression of human leucocyte antigen (HLA) DR in the epithelium or in the lamina propria in the patient groups studied or in the controls. In treated coeliac disease patients only a few TCR gamma delta+ T cells were found intraepithelially and in the lamina propria, but these cells were not detected in the lamina propria of oral mucosa of patients with untreated coeliac disease or in the controls. CONCLUSIONS: The infiltration of T cells into oral mucosa was increased in treated coeliac disease patients in spite of adherence to a gluten-free diet. Because the CD3(+) T cell count was higher than those of the TCR alpha beta+ and TCR gamma delta+ T cells, there must be other cells involved, probably natural killer (NK) cells. The increase in T-cell subsets in the treated coeliac disease patients seems not to result from poor dietary compliance, but might occur as a late immune response in coeliac disease and reflect chronic immunologic stimulation followed by regeneration of memory T cells. PMID- 11119269 TI - Connectivity and HIV-1 infection: role of CD4(+) T-cell counts and HIV-1 RNA copy number. AB - Following primary infection with human immunodeficiency virus (HIV)-1, antibodies against specific HIV-1 epitopes are elicited. However, non-HIV-1 specific antibodies, including autoantibodies, also arise. In fact, it has been proposed that such autoantibodies have an important role in the pathogenesis of HIV-1 infection. Because an imbalance in connectivity has been associated with autoimmune processes, we investigated the connectivity status of HIV-1-infected individuals. Moreover, we tested the possible role of viral load and CD4(+) T cell counts, in connectivity, because these parameters appear to be important in the prognosis of HIV-1 infection. Results show that indeed, there is an alteration in connectivity in these patients, both for immunoglobulin (Ig)G and IgM, which is an immune alteration not previously identified in HIV-1 infection. In addition, our results show that viral load and CD4(+) T-cell counts are both equally important in defining the characteristic pattern of connectivity in HIV-1 infected individuals, and that neither is independently responsible for alterations in patient connectivity status. PMID- 11119268 TI - Expression of CD1d in the duodenum of patients with cow's milk hypersensitivity. AB - CD1 cell surface glycoproteins represent a family of non-major histocompatibility complex (MHC) encoded antigen-presenting molecules. All members of the CD1 family appear to mediate the recognition of microbial or endogenous lipid and glycolipid antigens. The recognition of CD1d by a unique subset of natural killer (NK) T cells that leads to rapid production of large amounts of both type 1 and type 2 cytokines can be augmented by some synthetic glycolipids. Because of the proposed role of such CD1d-restricted T cells in immunoregulation, we hypothesized that CD1d molecules participate in mucosal immune responses in patients with gastrointestinal symptoms owing to food hypersensitivity. Patients of that category represent a heterogeneous group in which poorly defined immunological mechanisms are believed to contribute to disease pathogenesis. The expression of CD1 in duodenal biopsy samples from six patients with verified intolerance to cow's milk and six healthy controls was studied by immunoperoxidase staining of cryostat sections using a panel of mouse monoclonal antibodies (MoAbs) specific for CD1a, b, c, and d. Large numbers of CD1d positive cells were found in the lamina propria of all the patients, both during the symptomatic and the asymptomatic periods, whereas healthy controls were virtually devoid of CD1d expression in the duodenum. The localization of CD1d positive cells corresponded to areas where B cells, plasma cells and dendritic cells (DC) were present. A positive correlation was found between the numbers of CD1d(+) and CD19(+) cells in the lamina propria. In contrast to previous reports, no CD1d expression was found on the epithelial cells. Although less numerous than CD1d(+) the CD1c(+) cells were also present in all the patients and in five out of six controls. No staining for CD1a or CD1b was detected in the duodenal biopsy samples from any of the subjects. The exclusive presence of CD1d in the duodenal lamina propria of the patients with cow's milk hypersensitivity might suggest the participation of these molecules in the pathogenesis of allergic reactions to food. PMID- 11119270 TI - Prognostic value of soluble intercellular adhesion molecule-1 (s-ICAM-1) in HIV infected children. AB - Central events in the host defence system and immune-mediated damage are tightly regulated by cell adhesion molecules. Sera from 28 human immunodeficiency virus (HIV)-1 infected children divided into groups according to disease severity, six seroreverting (SR) children and 25 healthy controls were studied to detect the presence of soluble intercellular adhesion molecule-1 (s-ICAM-1). Soluble ICAM-1 levels were found to be significantly increased in HIV-infected children in comparison with SR children or healthy controls. Levels of soluble ICAM-1 were higher in patients with severe forms of HIV-infection than in those with a milder form of the disease. Significant differences in titers of s-ICAM-1 were recorded between SR children and HIV-infected children with mild disease or healthy controls. There was a significant correlation between s-ICAM-1 levels and the concentrations of beta 2 microglobulin (beta 2m) and, to a lesser extend, immunoglobulin A levels (IgA). Soluble ICAM-1 levels didn't change considerably in HIV-infected children in stable clinical conditions, independently of their clinical stage of the disease, during a follow-up period of 9-12 months. Conversely, s-ICAM-1 levels increased simultaneously with the appearance of new well-defined clinical disorders or decreased during the improvement of clinical conditions. A significant negative correlation was recorded between the titers of the s-ICAM-1 and the CD4(+) T-cell levels. These results suggest that the s-ICAM 1 might be another useful tool to evaluate disease progression. PMID- 11119271 TI - Clinical relevance of cytokine production in HIV-1 infection in children on antiretroviral therapy. AB - In order to investigate the correlation among cytokine production and antiretroviral therapy (ART), viral load, CD4(+) and CD8(+) T lymphocytes, 55 human immunodeficiency virus (HIV)-1-infected children on ART or not, and 16 uninfected controls were studied. Peripheral blood mononuclear cells (PBMCs) of HIV-1-infected children and controls were cultured and spontaneous and mitogen stimulated cytokines production was quantified in the supernatants. Viral load was quantified using standard molecular assay. CD4 and CD8 T-lymphocyte counts were determined by flow cytometry. Cytokine production by mitogen-stimulated PBMCs showed different profiles in HIV-1 children whether treated or not. The tumour necrosis factor (TNF)-alpha production was higher and the interleukin (IL) 10 production was lower in the HIV-1-untreated group than in the HIV-1-treated children and controls. The IL-2 production was reduced and the RANTES production was higher in both HIV-1 groups compared with the controls. The interferon (IFN) gamma and the IL-5 production was significantly reduced in the HIV-1-treated children compared to the controls. Interestingly, the analysis of the correlation of HIV-1 phenotype with cytokine production indicated an increased RANTES production in relation to nonsyncytium-inducing viral phenotype with slow/low replication profile, whereas decreased IL-10 levels was associated to syncytium inducing (SI) strains and rapid/high replication. Our findings suggest that AVT changes on the cytokine and chemokine production play an important role in the HIV pathogenesis. PMID- 11119272 TI - Strain rate imaging by ultrasonography in the diagnosis of coronary artery disease. AB - Regional strain rate in the left ventricle can be assessed in real time and color mapped. The method is termed strain rate imaging (SRI), and findings correspond well with 2-dimensional echocardiography. This study addresses SRI as a method for localizing coronary lesions, compared with standard echocardiography. Twenty patients with acute myocardial infarction who underwent coronary angiography for clinical reasons were examined with SRI and standard echocardiography. Wall motion was graded by SRI color and separately by wall thickening. Strain rate imaging and 2-dimensional echocardiography results agreed well. An infarct related artery was identified from angiograms combined with electrocardiograms. Both methods identified an infarct-related artery in 19 possible cases and had equal sensitivity and specificity for wall segments affected by lesion. Combining the information from both methods did not change accuracy. The study validates SRI as a method for assessing regional wall function in coronary artery disease. The advantages of SRI are discussed and measurements of strain rates are given. PMID- 11119273 TI - Use of angiotensin II stress pulsed tissue Doppler imaging to evaluate regional left ventricular contractility in patients with hypertrophic cardiomyopathy. AB - There is controversy concerning whether contract-ility in the nonhypertrophied region of the left ventricular (LV) wall is impaired or normal in patients with hypertrophic cardiomyopathy (HCM). Global LV systolic function decreases with increases in afterload in this disease. This study was performed to identify abnormalities in regional LV contractility along the long and short axes in the setting of HCM with the use of angiotensin II (AT-II) stress pulsed tissue Doppler imaging (PTDI). Angiotensin II was administered intravenously to patients with asymmetric septal hypertrophy (HCM group, n = 21) and age-matched normal volunteers (N group, n = 12). We then measured the percent LV fractional shortening (%FS) and end-systolic circumferential LV wall stress by M-mode echocardiography, LV ejection fraction (LVEF) by 2-dimensional echocardiography, and time-velocity integral (TVI) of LV outflow velocity by pulsed Doppler echocardiography. The peak first and second systolic LV wall motion velocities along the long (L-Sw(1) and L-Sw(2)) and short (S-Sw(1) and S-Sw(2)) axes were measured in the LV posterior wall and ventricular septum with the use of PTDI. The end-systolic circumferential LV wall stress at baseline was significantly lower in the HCM group. The L-Sw(1) and L-Sw(2) for the posterior wall were significantly lower in the HCM group, but the S-Sw(1) and S-Sw(2) for the posterior wall and ventricular septum were similar in the two groups. The %FS, LVEF, TVI, and systolic PTDI variables along both axes for the posterior wall decreased significantly, and end-systolic circumferential LV wall stress increased significantly at AT-II doses of 0.005 or 0.010 microg/kg per minute in the HCM group. No significant changes were found in either group in the systolic PTDI variables (except for L-Sw(1)) for the ventricular septum with AT-II infusion. Contractility along the long and short axes of the nonhypertrophied LV wall is easily impaired with increases in afterload in patients with HCM, resulting in a decrease in global LV systolic function. We found AT-II stress PTDI to be a safe and useful technique for evaluating the regional LV systolic function in this disease. PMID- 11119274 TI - Targeting percutaneous transluminal septal ablation for hypertrophic obstructive cardiomyopathy by intraprocedural echocardiographic monitoring. AB - BACKGROUND: Percutaneous septal ablation has evolved as an alternative to surgery for reducing symptoms and outflow gradients in patients with hypertrophic obstructive cardiomyopathy. Intraprocedural echo-cardiographic imaging can improve clinical and hemodynamic results. Growing experience with this method has additionally shown that threatening necrosis of the myocardium distant from the septal target region can be detected. METHODS AND RESULTS: Percutaneous septal ablation was performed in 162 patients (80 women, 82 men; aged 54.1 +/- 15.5 years); 131 of whom were targeted by intraprocedural myocardial contrast echocardiography. In 11 patients (7%), an atypical target vessel or a perfusion area distant from the expected septal target region was detected, leading to a target vessel change. Permanent pacing was necessary in 14 patients (9%). Three patients (2%) died. After 3 months, the mean New York Heart Association functional class was reduced in the returning 159 patients from 2.8 +/- 0.5 to 1.3 +/- 1.0 (P <.0001) along with a gradient reduction from 77 +/- 35 to 12 +/- 22 mm Hg at rest, and from 147 +/- 43 to 44 +/- 45 mm Hg with provocation (P < .0001 each). The main reason for unsatisfactory gradient reduction was suboptimal scar placement in the patients treated before the introduction of intraprocedural myocardial contrast echocardiography. CONCLUSIONS: Percutaneous septal ablation is an effective nonsurgical technique for reducing symptoms and outflow gradients in hypertrophic obstructive cardio-myopathy. Echocardiographic guidance adds substantially to safety and efficacy of the procedure and should therefore be considered routinely. PMID- 11119275 TI - The role of intraoperative transesophageal echocardiography in patients undergoing cardiac mass removal. AB - To evaluate the role of intraoperative transesophageal echocardiography (IOTEE) during surgical removal of cardiac masses, we studied 75 consecutive patients (34 men, aged 56 +/- 16 years, range 17 to 82 years) who underwent surgery primarily for cardiac mass removal with the adjunct of IOTEE for the years 1993 through 1998. The IOTEE provided new information before bypass in 6 patients (8%), altering the planned surgical procedure in all. A newly discovered patent foramen ovale was closed in 2 patients, a second myxoma discovered in one patient, a mitral valve repaired in one patient, inferior vena caval cannulation site clarified in one patient, and in one patient the mass was no longer present and the surgery canceled. In 10 patients (13%), new post-bypass information was found, which prompted return to bypass for valve repair in 3 patients, altered nonsurgical management in 3 patients, and did not necessitate specific measures in 4 patients. Thus, in 75 patients coming to surgery for mass removal, IOTEE affected intraoperative management in 12 (16%). PMID- 11119276 TI - Dobutamine stress echocardiography and exercise electrocardiography for risk stratification in medically treated unstable angina. AB - Previous reports have demonstrated the superiority of exercise echocardiography over exercise electro-cardiography (ex-ECG) for risk stratification in patients with medically stabilized unstable angina (UA). We sought to analyze the prognostic value of dobutamine stress echocardiography (DSE) compared with ex-ECG for risk stratification in patients with UA. METHODS: Ninety-two patients with medically treated UA were studied (mean age 65 +/- 11 years, 24 women, 42% of patients had electrocardiographic abnormalities on admission). Dobutamine stress echocardiography and treadmill ex-ECG were performed on the third day after hospital admission. End points were recurrent UA, myocardial infarction (MI), or cardiac death. RESULTS: Mean follow-up was 24 +/- 7 months. During follow-up, 22 patients had cardiac events (18 recurrent UA, 2 MI, 2 cardiac deaths). The event free survival rate was 80% for patients with negative DSE results for ischemia and 52% for those with positive DSE results (log rank 9.57; P =.002), compared with an event-free survival rate of 79% for patients with negative ex-ECG results and 66% for those with positive ex-ECG results (log rank 2.06; P = not significant). Left ventricular dysfunction (P =.01) and a positive dobutamine stress echocardiogram (P =.03), but not a positive exercise electrocardiogram, were independent predictors of cardiac events during follow-up. CONCLUSIONS: Dobutamine stress echocardiography performed early in medically treated patients with UA predicts cardiac events during follow-up more accurately and with more specificity than ex-ECG does in this population. PMID- 11119277 TI - Value of transthoracic echocardiography for the detection of high-grade coronary artery stenosis: prospective evaluation in 50 consecutive patients scheduled for coronary angiography. AB - We prospectively evaluated the feasibility of direct, transthoracic evaluation of coronary arteries to diagnose flow-limiting lesions. Second harmonic mode in B mode and fundamental mode for Doppler examinations was used. A stenosis was diagnosed when maximal flow velocity at least doubled in comparison with that of the adjacent segment or when local velocity was at least 2 m/s. Of the left anterior descending coronary artery segments assessed, 34 were proximal, 35 middle, and 34 distal segments. The corresponding figures for circumflex coronary artery segments were 17 proximal and 11 middle segments and for the right coronary artery, 14 proximal and 15 distal segments. No distal circumflex and only 1 mid right coronary artery segment was visualized. Twenty-eight stenoses were diagnosed. Specificity for stenosis detection was 96% to 100% and sensitivity was 62% to 66%. Echo-cardiography was unable to document occlusions. Transthoracic echocardiography allows for coronary artery assessment in a significant portion of patients scheduled for coronary angiography. It may be used to document the presence of coronary artery stenosis. With further technologic improvements, transthoracic echocardiography could enable the monitoring of the restenosis process after percutaneous transluminal coronary angioplasty/stent intervention and coronary artery luminal narrowing after heart transplantation. PMID- 11119278 TI - Sublingual nitroglycerin delays arterial wave reflections despite increased aortic "stiffness" in patients with hypertension: a Doppler echocardiography study. AB - Venodilatation with consequent reduction in left ventricular filling and end diastolic wall stress is an important mechanism for the beneficial effects of nitroglycerin in ischemic heart disease and in left ventricular failure. The effects of sublingual nitroglycerin on arterial pulsatile hemodynamics are less well defined. Doppler echocardiography and the calibrated subclavian artery pulse tracing were used to assess hemodynamics in subjects with sustained arterial hypertension (n = 25) before and 5 to 10 minutes after sublingual deposition of 0.5 mg glyceryl trinitrate. Aortic characteristic impedance was calculated by averaging the modulus of the input impedance (ratio of pressure to flow) at high frequencies and by calculating the ratio of pressure and flow increments during upstroke. The pressure wave was split into forward and backward components, and the reflection coefficient (the ratio of backward to forward pressures) was calculated. Parameters of the arterial bed were estimated by using 2- and 3 element Windkessel models. Nitroglycerin delayed the return of arterial wave reflections by 17% (P =.02) and increased aortic characteristic impedance by 20% (P =. 01), but it did not influence total arterial compliance. Mean arterial pressure decreased 7% (P =.0001), but pulse pressure did not change. Stroke volume and the acceleration time of aortic root flow decreased by 13% (P =.0001) and 8% (P =.01), respectively. Cardiac output decreased 7% (P =.01), despite an increase in heart rate of 10% (P =.0001). Peripheral resistance tended to decrease (4%, P =.06). Thus, in subjects with sustained hypertension, sublingual nitroglycerin dilates peripheral, predominantly muscular arteries with a subsequent delayed return of reflected pressure waves. Reflex activation of the sympathetic nervous system with consequent increased acceleration of left ventricular ejection seems to counteract the effect of reduced mean arterial pressure (distending pressure) with respect to the "stiffness" of the aorta. PMID- 11119279 TI - Transition to an all-digital echocardiography laboratory: a large, multi-site private cardiology practice experience. AB - Acquisition, interpretation, and storage of digital echocardiographic images has many advantages over the standard videotape-based method. Archival, transmission, and comparative interpretation are all optimized with digital echocardiography. A study performed at one site can be immediately available for viewing and analysis at another site by means of standard data transfer technology. Echocardiograms can be interpreted in the context of prior studies, which are readily available for side-by-side comparison. The transition to an all-digital laboratory involves the commitment of persons at multiple levels in the cardiology practice, including administrators, information technology specialists, sonographers, and physicians. Quality of patient care, use of physicians' and sonographers' time, and long-term financial benefit are all areas where improvement may be realized with the use of digital echocardiography. We present our experience in the development of an all-digital echocardiography laboratory, and we conclude that digital echo-cardiography is practical and can be implemented readily in a clinical setting. We performed several correlative analyses during this transition to validate the consistency and accuracy of digital interpretation compared with those of analog methods. The transition process from analog (videotape) to digital, including full wide area network exchange, took approximately 8 months. As technology advances, issues surrounding storage, comparison, and acquisition formats will continue to develop. We hope that our experience will help others make the transition to the digital environment and benefit from the ease of image access, the ability to comparatively interpret echocardiograms, and the superior image quality afforded by this advancement. PMID- 11119280 TI - Aberrant origin of the left coronary artery from the right aortic sinus: surgical intervention based on echocardiographic diagnosis. AB - An athletic 15-year-old girl with aberrant left coronary artery from the right coronary sinus, presented with syncope during exercise. Trans-thoracic echocardiography was the only imaging technique that clearly demonstrated her anomaly. The results of magnetic resonance and selective coronary angiographic imaging were inconclusive. Surgical intervention was successfully performed on the basis of the echocardiographic diagnosis. PMID- 11119281 TI - Entrapment of subvalvular mitral tissue causing intermittent failure of a St Jude mitral prosthesis. AB - Frequently portions of the mitral valve and sub-valvular apparatus are left intact during mitral valve replacement to help preserve left ventricular function. We describe a patient with paroxysmal congestive heart failure caused by intermittent entrapment of the subvalvular apparatus in the prosthesis, preventing complete valve closure. PMID- 11119282 TI - The value of repeat transesophageal echocardiography in the evaluation of embolism from the aorta. AB - Transesophageal echocardiography (TEE) is now widely used in the evaluation of patients with unexplained stroke or transient ischemic attack, in part to exclude the presence of protruding aortic arch atheromas. We report two cases in which repeated TEE revealed an aortic clot not seen on the earlier transesophageal echocardiogram performed immediately after embolization. These cases illustrate the dynamic nature of aortic thrombus and the role of TEE in its diagnosis. PMID- 11119283 TI - A case of left atrial appendage thrombus with embolic stroke in association with aortic stenosis in sinus rhythm. AB - Although left atrial appendage (LAA) thrombus formation in the presence of sinus rhythm may potentially be the source for embolic events in various types of heart disease, no cases of LAA thrombus with embolic stroke in association with aortic stenosis in sinus rhythm have been reported. We present a case of valvular aortic stenosis with cerebral embolism in a person who was in sinus rhythm and had an LAA thrombus diagnosed by transesophageal echocardiography. PMID- 11119284 TI - Localized aortic dissection: unusual features by transesophageal echocardiography. AB - Transesophageal echocardiography relies on the presence of an undulating intimal flap for the diagnosis of aortic dissection. Furthermore, to distinguish true dissection from echo artifacts, the flap has to be identified in more than one view, and it must have a motion independent of the aortic wall. We describe the transesophageal echocardiography appearance of a localized aortic dissection with atypical features for an intimal flap. Awareness of this unusual echocardiographic appearance of an intimal flap will avoid misdiagnosis of the potentially serious acute aortic dissection. PMID- 11119285 TI - A human skull cast model for training of intracranial microneurosurgical skills. AB - Skillful microsurgical techniques such as microvascular anastomosis and repair of nerves require dedicated and extensive laboratory training. Human microneurosurgery poses several additional technical difficulties, as the intracranial procedures are often performed through a narrow operative space at a considerable length, using knee-bend instruments. This article presents a laboratory model to simulate microneurosurgical procedures in humans. A human skull cast model made from plaster is cut such that, when placed on a operating table, it represents a standard position of a pterional approach and that the point of operation is at the same depth as the hypothetical circle of Willis. A standard pterional opening is made in de skull cast and the model is placed over the rat. Subsequently, all surgical procedures starting from tissue preparation to performing an arterial, venous, and/or nerve repair are performed with the cast over the rat using microneurosurgical knee-bent instruments and a surgical microscope. Microsurgical procedures such as end-to-end vessel anastomosis and nerve repair are technically much more challenging and difficult to execute when performed through the pterional opening of the human skull cast than without the cast model. Moreover, the cast model useful in training microsurgical techniques performed with long knee-bend instruments. It is concluded that the skull cast model represents a useful method to train intracranial microneurosurgical blood vessel anastomosis and nerve repair. PMID- 11119286 TI - Safe injection of cultured schwann cells into peripheral nerve allografts. AB - The effects of cultured host Schwann cells on axonal regeneration in peripheral nerve allografts were studied. Fischer rats served as recipient animals and Buffalo rats provided nerve allografts. Animals were randomized into 9 groups. Rats receiving tibial nerve isografts were left untreated (group I), or injected with isogeneic Fischer Schwann cells (group II) or placebo suspension (group III). Allografts obtained from Buffalo rats were left untreated (group IV), or received isogeneic Fischer Schwann cells (group V), 2 mg/kg Cyclosporin A and Fischer Schwann cells (group VI), 5 mg/kg Cyclosporin A (group VII), or 5 mg/kg Cyclosporin A with Schwann cells (group VIII). No Schwann cell tumors were identified 4 or 8 weeks postoperatively. Group IX animals, harvested 3 days postoperatively, demonstrated no evidence of injection injury. Schwann cells modestly improved axonal regeneration in both isografts and allografts and may have a clinical role in the treatment of peripheral nerve allografts. PMID- 11119287 TI - Microvascular repair with 1-mm polytetrafluoroethylene (PTFE) grafts: effect of recombinant tissue-type plasminogen activator (rt-PA) on the patency rate and healing process. AB - The present study assesses the effect of recombinant tissue-type plasminogen activator (rt-PA) on the patency rate and healing process of microvascular polytetrafluoroethylene (PTFE) grafts. Wistar rats were used, divided into four groups of 25 animals each. After dissection of the carotid artery a segment of the vessel, 1 cm long, was resected and replaced by equal length graft. Two different type fibril length (30- or 60-microm) grafts of the same wall thickness (0.18 mm) were used. Normal saline or 3 mg/kg of body weight of rt-PA was applied locally in each group of different fibril length grafts. Patency tests were performed at 15 min and 4 weeks after blood flow was reestablished. All grafts were harvested and examined histologically. The results showed that local application of rt-PA improves patency statistically significantly in both types of fibril length grafts. Patency in 60-microm fibril length grafts was statistically significantly higher than that of 30-microm fibril length grafts, whether rt-PA was used or not. The use of rt-PA had no influence on the healing process of either type of graft. PMID- 11119288 TI - Effect of vascular freezing on the histopathology of dissected small vessels in the rat: vascular freezing does induce intimal hyperplasia in arteries and veins. AB - Intimal hyperplasia is the primary response of a vessel wall after injury. It may be the single most significant factor affecting long-term patency. The purpose of this study was to find out whether freeze injury, inflicted on rat microvessels, would be followed by intimal hyperplasia. Toward this aim, we exposed the superficial femoral vessels in the rat. On one side they were frozen using liquid nitrogen spray. The other side was dissected as the control. Vessel segments, harvested immediately and after 1, 2, 3, and 5 months, were studied light microscopically for the occurrence and content of intimal hyperplasia. In the arteries a considerable intimal hyperplasia was found within a 4-week interval, persisting for at least 5 months, as a result of the freeze injury. In the veins, the intimal hyperplasia was much less marked but was nevertheless demonstrable. These findings are not in agreement with earlier studies, in which freezing of injured rat microarteries with liquid nitrogen spray was followed by complete regeneration of the vessel wall, without intimal hyperplasia taking place. The factors contributing to these differences are discussed. It is concluded that freezing of a vessel wall is followed by intimal hyperplasia, which is part of the normal healing process. PMID- 11119289 TI - Multiple telescoping anastomosis on an artery. AB - The telescoping anastomotic technique was carried out four times on the rat femoral artery in order to examine whether the technique could safely be used three to four times for repair of an artery that was injured at two different sites. In addition, the technique was repeated five and six times to determine the maximum number of anastomosis that could be performed on an artery and to clarify problems of the telescoping anastomosis. In the rats in which the anastomosis was carried out four and five times, 100% patency was obtained, while the first occlusion occurred in a graft on which the anastomosis was repeated six times. The telescoping technique appeared to safely be used three or four times for repair of an artery that had been divided at two sites if tension of the repaired vessel was kept low by the grafting, and torsion of the graft and deformities of the inserted vessel were minimal. PMID- 11119290 TI - Cold storage of rat skeletal muscle free flaps and pre-ischemic perfusion with modified UW solution. AB - We used the rat medial gastrocnemius free flap, based on a pedicle of the femoral artery and vein, in order to test the tolerance of skeletal muscle to cold ischemia-reperfusion (IR) injury, and to determine whether tolerance can be enhanced by pre-ischemic perfusion with tissue/organ preservation solutions. Muscle flaps (n = 6 per group) were subjected to variable periods of cold storage (0, 1, 2, 3, or 4 days) and 24-h normothermic reperfusion. Muscle viability, as determined by nitroblue tetrazolium (NBT) histochemical staining of viable mitochondria and supported by histological examination, was 100%, 26%, 11%, 4%, and 1%, respectively. Using 24-h cold storage/24-h reperfusion as the experimental conditions, groups of muscle flaps (n = 5 per group) were perfused immediately before cold storage with either modified, colloid-free University of Wisconsin (UW) solution, a solution described by Kohout et al. (Br J Plast Surg 1995;48:132-144) or normal saline. Perfusion with modified UW solution or Kohout's solution increased survival to 33% (P < 0.05) and 28% (not statistically significant), respectively, compared with the 19% viability of separate groups of nonperfused or saline-perfused controls. These findings indicate that cold-stored skeletal muscle is highly susceptible to cold IR injury and that the viability can be increased by prior perfusion with a tissue preservation solution such as modified UW solution. PMID- 11119291 TI - Ischemic preconditioning: lack of delayed protection against skeletal muscle ischemia-reperfusion. AB - We investigated the ability of ischemic preconditioning to induce expression of heat shock protein 70 (Hsp 70) and/or to increase muscle survival after ischemia reperfusion in the rat hind limb. Ischemic preconditioning regimens tested were; 1 x 5 min of ischemia, 4 x 5 min of ischemia interrupted by 10 min of reperfusion, 1 x 10 min of ischemia or 2 x 10 min of ischemia interrupted by 15 min of reperfusion. Western blot analysis revealed only a modest induction of Hsp 70 at 24 h after preconditioning using the latter two protocols of 1 x 10 min of ischemia or 2 x 10 min. Used at 24 h prior to prolonged ischemia, neither protocol improved muscle survival measured at 24 h after reperfusion. In conclusion, ischemic preconditioning did not produce delayed protection from ischemia-reperfusion in this model and the study suggests that ischemic preconditioning is not a useful protective strategy against skeletal muscle necrosis in the long-term. PMID- 11119292 TI - Genetic Epidemiology. PMID- 11119293 TI - Detecting association in a case-control study while correcting for population stratification. AB - Case-control studies are subject to the problem of population stratification, which can occur in ethnically mixed populations and can lead to significant associations being detected at loci that have nothing to do with disease. Here, we describe a way to measure and correct for stratification by genotyping a moderate number of unlinked genetic markers in the same set of cases and controls in which a candidate association was found. The average of association statistics across the markers directly measures stratification. By dividing the candidate association statistic by this average, a P-value can be obtained that corrects for stratification. PMID- 11119294 TI - A Bayesian hierarchical model for allele frequencies. AB - Genetic epidemiological methodologies, such as linkage analysis, often require accurate estimates of allele frequencies. When studies involve multiple sub populations with different evolutionary histories, accurate estimates can be difficult to obtain because the number of subjects per sub-population tends to be limited. Given allele counts for a collection of loci and sub-populations, we propose a Bayesian hierarchical model that extends existing empirical Bayesian approaches by allowing for explicit inclusion of prior information about both allele frequencies and inter-population divergence. We describe how such information can be derived from published data and then incorporated into the model via prior distributions for model parameters. By analysis of simulated data, we highlight how the hierarchical model, as implemented in the publicly available program AllDist, combines prior information with the observed data to refine allele frequency estimates. PMID- 11119295 TI - A weighted test using both extreme discordant and concordant sib pairs for detecting linkage. AB - Statistical procedures using extremely discordant and concordant sib-pairs have been developed for mapping quantitative trait loci in humans. To improve the power of the existing methods, test statistics placing greater weight on the more discordant or more concordant pairs are proposed. Because the optimum choice of weights would depend on the underlying genetic model, which is not usually known, a test with simple weights is suggested. This test is shown to have greater power than the currently available ones for a variety of genetic models. PMID- 11119296 TI - Likelihood-ratio affected sib-pair tests applied to multiply affected sibships: issues of power and type I error rate. AB - "All-pairs" likelihood-ratio analyses, such as those performed by MAPMAKER/SIBS [Kruglyak and Lander, 1995], require that a sibship containing N affected siblings be split into N(N - 1)/2 sibships, each containing a different pair of affected sibs, before analysis. Each of these N(N - 1)/2 sibships may also contain the other affected sibs from the original sibship, coded as unaffected, to infer missing parental genotypes, as is done automatically in MAPMAKER/SIBS. Then, the use of the same individuals both as affecteds to test for linkage and, elsewhere, as unaffecteds to infer missing parental genotypes leads to negative correlations in the estimated identity by descent sharing among affected pairs from the same original multiplex sibship. This gives a conservative test of linkage, even when no downweighting is applied. Conversely, if the other affected sibs from the original sibship are omitted, the correlations are positive and the linkage test is anticonservative in the absence of weighting. True type I error probability also depends on marker informativity, typed parents, number of affected sibs included in the analysis, and the weighting scheme. This suggests the use of simulation, rather than asymptotic theory, to assess significance levels. The power of multiplex sibships relative to affected pairs increases with increasing phenocopy percentage, but the presence of typed unaffected sibs improves the relative power of multiplex sibships greatly only when penetrance is high. It was found that the 2/N weighting proposed by Suarez and Hodge [1979] increased power over an unweighted analysis in many situations, provided significance levels were adjusted appropriately by simulation. PMID- 11119297 TI - Transmission/disequilibrium tests for quantitative traits. AB - Spielman et al. [1993] proposed a transmission-disequilibrium test (TDT), based on marker data collected on affected offspring and their parents, to test for linkage between a genetic marker and a binary trait provided there is allelic association. It has been shown that this TDT is powerful and is not affected by allelic association due to population stratification in the absence of linkage. For quantitative traits, George and Elston [1987] proposed a likelihood method to detect the effect of a candidate gene in pedigree data when familial correlations are present. This test will detect allelic association but will do so in the absence of linkage. In this paper, we investigate two new likelihood-ratio test statistics for multi-generational quantitative traits to test either for linkage in the presence of allelic association or for allelic association in the presence of linkage, such as may be due to linkage disequilibrium. We compare these two tests analytically and by simulation with respect to 1) the sample size required for the asymptotic null distributions to be valid and 2) their power to detect association in those cases in which they are not sensitive to population stratification unless linkage is present. In general, 80 nuclear families with two children each and at least one heterozygous parent, or the equivalent number of children in large pedigrees, are enough for the asymptotic null distribution of the proposed conditional and TDT methods to be valid. The theoretical power is close to the simulated power except for the case of a recessive allele with low frequency. A sampling strategy is proposed that dramatically improves power. PMID- 11119298 TI - Neoplasms in neurofibromatosis 1 are related to gender but not to family history of cancer. AB - The risk of malignancies among persons with neurofibromatosis 1 (NF1) is higher than in the general population, but the excess risk has not been precisely estimated. The effects of gender and inheritance pattern on cancer risk are unclear. Therefore, we conducted a historical cohort study to determine cancer risk factors by contacting 138 Caucasian NF1 patients originally seen at Baylor College of Medicine (BCM) in Houston between 1978 and 1984. A total of 304 patients of all ethnic groups were evaluated at BCM during this period. We successfully located 173 patients, 138 of who were Caucasian. We computed standardized incidence ratios (SIRs) with the age-, gender-, and time period specific rates from the Connecticut Tumor Registry for 2,094 person-years of observation (median follow-up = 16 years). Eleven incident tumors were reported. Females were at much higher risk of cancer than males (SIR = 5.6, 95% confidence interval (CI) 2.7-10.3 and SIR = 0.6; 95% CI, 0.0-3.0, respectively). We found no elevated cancer risk in unaffected first-degree relatives, regardless of whether the proband had cancer or not (SIR = 1.1 95% CI, 0.6-1.8 and SIR = 1.0, 95% CI, 0.6-1.5, respectively). Our results suggest that malignancy in the proband is not the result of a modifying gene that has a significant impact on general cancer risk. PMID- 11119299 TI - Ramifications of HLA class I polymorphism and population genetics for vaccine development. AB - HLA polymorphism can complicate the design and development of vaccines, especially those that contain a selected number of epitopes and are directed at pathogens prevalent worldwide. Because of HLA class I restricted antigen recognition and ethnic variation in HLA distribution, such vaccines may not be uniformly effective across populations. We, therefore, considered whether it is possible to assemble a panel of HLA-A and/or HLA-B alleles that would allow the formulation of a single vaccine for a set of Caucasian, Black, or Asian populations. In applying an algorithm to predict levels of favorable response, we identified predominant alleles in 15 representative populations. Approximately 80% of the individuals in one African Black population and five Asian populations were positive for at least one of three HLA-A alleles. Eighty percent coverage was also theoretically possible in five Caucasian populations with only five HLA A alleles. Four of five Black populations analyzed also required five alleles, but the allelic combinations differed. Our findings suggest that HLA-A alleles may be preferred targets because of the increased heterogeneity at HLA-B, although addition of a single HLA-B allele to a set of HLA-A alleles improved coverage. This approach provides for the identification of combinations of alleles that represent a desired percentage of a population and that could be targeted in designing vaccines. For vaccines with known HLA-restricted epitopes, it allows a prediction of theoretical levels of "responder" and "non-responder" status. Finally, these results might be used in the analysis of protein sequences to identify potential CD8+ T-cell epitopes in populations of interest. Biologic variables that may have further relevance are discussed. PMID- 11119300 TI - Estimation of genetic and environmental components in colorectal and lung cancer and melanoma. AB - Cancer has predominant environmental and somatic causes but the assessment of hereditary (genetic) causes is difficult, except for highly penetrant single-gene causes. Family studies are only partially informative in this regard because family members share diet and life-styles. Twin studies have been classically used to disentangle the effects of heredity and environment on disease etiology. We estimate the genetic and environmental components in colorectal and lung cancer and melanoma by comparing cancer risks in family members. The Swedish Family-Cancer Database, comprising more than 6 million individuals, was used as the source of family and cancer data. Tetrachoric correlations were used to describe similarity in cancer liability among family members. Structural equation modelling was used to derive estimates of the importance of genetic and environmental effects. The estimated genetic component ranged from 10% in colon and colorectal cancer to 18% in melanoma. For lung cancer, the share was 14%. If assortative mating were important for liability to cancer, these heritability estimates may be an underestimation of the true genetic effects. Non-shared environmental effect was 67-68% in colorectal cancer and melanoma, and 71% in lung cancer. Shared and childhood environments were equally important in colorectal cancer and melanoma, whereas no childhood effect was observed for lung cancer. PMID- 11119301 TI - Gene-diet interactions and plasma lipoproteins: role of apolipoprotein E and habitual saturated fat intake. AB - To test whether plasma lipoprotein levels and low density lipoprotein (LDL) particle size are modulated by an interaction between habitual saturated fat intake and apolipoprotein E (APOE) genotype, we studied 420 randomly selected free-living Costa Ricans. The APOE allele frequencies were 0.03 for APOE2, 0.91 for APOE3, and 0.06 for APOE4. The median saturated fat intake, 11% of energy, was used to divide the population into two groups, LOW-SAT (mean intake 8.6% energy) represents those below median intake, and HIGH-SAT (mean intake 13.5%) represents those above median intake. Significant interactions between APOE genotype and diet were found for VLDL (P = 0.03) and HDL cholesterol (P = 0.02). Higher saturated fat intake was associated with higher VLDL cholesterol (+29%) and lower HDL cholesterol (-22%) in APOE2 carriers, while the opposite association was observed in APOE4 carriers (-31% for VLDL cholesterol and +10% for HDL cholesterol). Higher saturated fat intake was associated with smaller LDL particles (-2%, P < 0.05) in APOE2 carriers, and larger LDL particles (+2%, P < 0.05) in APOE4 carriers, but the gene-diet interaction was not statistically significant (P = 0.09). Higher saturated fat intake was associated with higher LDL cholesterol in all genotypes (mean +/- SEM, LOW-SAT 2.61 +/- 0.05 vs. HIGH SAT 2.84 +/- 0.05 mmol/L, P = 0.009). These data suggest that the APOE2 allele could modulate the effect of habitual saturated fat on VLDL cholesterol and HDL cholesterol in a population with an average habitual total fat intake of less than 30%. PMID- 11119302 TI - Expansion mutation frequency and CGG/GCC repeat polymorphism in FMR1 and FMR2 genes in an Indian population. AB - Based on molecular screening, we estimated the frequencies of fragile X syndrome and FRAXE syndrome in an institutionalized population (n = 130) in New Delhi, India. Eligibility criteria for inclusion of subjects in the study were mild/moderate mental retardation, with/without family history, and the fragile X clinical phenotype. Screening by Southern hybridization revealed an overall frequency of 0.077 of the syndrome in the sample population. The disorder was observed with a high frequency (0.1) among males as compared to females (0.027). No expansions of FMR2 allele were observed in the same study sample. CGG/GCC allelic polymorphism of FMR1 and FMR2 were established from a total of 392 X chromosomes, using the radioactive polymerase chain reaction-polyacrylamide gel electrophoresis method. Distinct repeat sizes, repeat ranges, and repeat modes characterised the FMR1 and FMR2 alleles. In the X chromosomes of both MR individuals and unaffected controls, unimodal values of 29 and 15 repeats in FMR1 and FMR2 genes, respectively, were observed. Allele frequency distribution was symmetrical at the FMR1 locus whereas a significant positive skew was observed for the FMR2 alleles. The observed heterozygosity of the FMR1 gene was 0.772 compared to 0.839 of FMR2. Correlation of CGG/GCC repeats of FMR1 and FMR2 did not show any association of repeat sizes at these two loci (correlation coefficient, rho = 0.09). CGG/GCC repeat variation at FMR1 and FMR2 loci observed in this study sample are different from that reported for the other Caucasian and Asian populations. PMID- 11119303 TI - Xylella genomics and bacterial pathogenicity to plants. AB - Xylella fastidiosa, a pathogen of citrus, is the first plant pathogenic bacterium for which the complete genome sequence has been published. Inspection of the sequence reveals high relatedness to many genes of other pathogens, notably Xanthomonas campestris. Based on this, we suggest that Xylella possesses certain easily testable properties that contribute to pathogenicity. We also present some general considerations for deriving information on pathogenicity from bacterial genomics. PMID- 11119304 TI - Comparison of Mycobacterium tuberculosis genomes reveals frequent deletions in a 20 kb variable region in clinical isolates. AB - The Mycobacterium tuberculosis complex is associated with a remarkably low level of structural gene polymorphism. As part of a search for alternative forms of genetic variation that may act as a source of biological diversity in M. tuberculosis, we have identified a region of the genome that is highly variable amongst a panel of unrelated clinical isolates. Fifteen of 24 isolates examined contained one or more copies of the M. tuberculosis-specific IS6110 insertion element within this 20 kb variable region. In nine of the isolates, including the laboratory-passaged strain H37Rv, genomic deletions were identified, resulting in loss of between two and 13 genes. In each case, deletions were associated with the presence of a copy of the IS6110 element. Absence of flanking tri- or tetra nucleotide repeats identified homologous recombination between adjacent IS6110 elements as the most likely mechanism of the deletion events. IS6110 insertion into hot-spots within the genome of M. tuberculosis provides a mechanism for generation of genetic diversity involving a high frequency of insertions and deletions. PMID- 11119305 TI - Accurate prediction of protein functional class from sequence in the Mycobacterium tuberculosis and Escherichia coli genomes using data mining. AB - The analysis of genomics data needs to become as automated as its generation. Here we present a novel data-mining approach to predicting protein functional class from sequence. This method is based on a combination of inductive logic programming clustering and rule learning. We demonstrate the effectiveness of this approach on the M. tuberculosis and E. coli genomes, and identify biologically interpretable rules which predict protein functional class from information only available from the sequence. These rules predict 65% of the ORFs with no assigned function in M. tuberculosis and 24% of those in E. coli, with an estimated accuracy of 60-80% (depending on the level of functional assignment). The rules are founded on a combination of detection of remote homology, convergent evolution and horizontal gene transfer. We identify rules that predict protein functional class even in the absence of detectable sequence or structural homology. These rules give insight into the evolutionary history of M. tuberculosis and E. coli. PMID- 11119306 TI - Distinct requirements for zebrafish angiogenesis revealed by a VEGF-A morphant. AB - Angiogenesis is a fundamental vertebrate developmental process that requires signalling by the secreted protein vascular endothelial growth factor-A (VEGF-A). VEGF-A functions in the development of embryonic structures, during tissue remodelling and for the growth of tumour-induced vasculature. The study of the role of VEGF-A during normal development has been significantly complicated by the dominant, haplo-insufficient nature of VEGF-A-targeted mutations in mice. We have used morpholino-based targeted gene knock-down technology to generate a zebrafish VEGF-A morphant loss of function model. Zebrafish VEGF-A morphant embryos develop with an enlarged pericardium and with major blood vessel deficiencies. Morphological assessment at 2 days of development indicates a nearly complete absence of both axial and intersegmental vasculature, with no or reduced numbers of circulating red blood cells. Molecular analysis using the endothelial markers fli-1 and flk-1 at 1 day of development demonstrates a fundamental distinction between VEGF-A requirements for axial and intersegmental vascular structure specification. VEGF-A is not required for the initial establishment of axial vasculature patterning, whereas all development of intersegmental vasculature is dependent on VEGF-A signalling. The zebrafish thus serves as a quality model for the study of conserved vertebrate angiogenesis processes during embryonic development. PMID- 11119307 TI - Morphants: a new systematic vertebrate functional genomics approach. AB - The vertebrate genome contains a predicted 50 000-100 000 genes, many of unknown function. The recent development of morpholino-based gene knock-down technology in zebrafish has opened the door to the genome-wide assignment of function based on sequence in a model vertebrate. This review describes technical aspects of morpholino use for functional genomics applications, including the potential for multigene targeting and known methodological limitations. The result of successful gene inactivation by this agent is proposed to yield embryos with a 'morphant' phenotypic designation. The establishment of a morphant database opens the door to true functional genomics using the vertebrate, Danio rerio. PMID- 11119308 TI - Featured organism: pathogen special: Vibrio cholerae, Pseudomonas aeruginosa and Xylella fastidiosa. AB - One could almost say that it is the latest fashion to sequence a bacterial genome. However, this would belittle the efforts of those working on these important organisms, whose data will greatly help those working on the prevention of disease in the fields of medicine and agriculture. In this feature we present a guided tour of the latest additions to the 'sequenced microbes' club. Vibrio cholerae is the causative agent of cholera, which is still a threat in countries with poor sanitation and unsafe drinking water. Pseudomonas aeruginosa is responsible for a large proportion of opportunistic human infections, typically infecting those with compromised immune systems, particularly cystic fibrosis patients, those patients on respirators and burn victims. Xylella fastidiosa is a plant pathogen that attacks citrus fruits by blocking the xylem, resulting in juiceless fruits of no commercial value. PMID- 11119309 TI - Meeting highlights: beyond the genome 2000: the 18th International Congress of Biochemistry and Molecular Biology. AB - The meeting was held on 16-20 July 2000 at the International Convention Centre in Birmingham, UK, and was co-organized by the International Union of Biochemistry and Molecular Biology (IUBMB) and the Federation of European Biochemical Societies (FEBS). Although the meeting had a broad subject area, the emphasis was firmly placed on post-genomic studies, and hence several sessions should be of interest to our readers. We provide highlights of these sessions, bringing you a report on the most exciting and informative presentations. PMID- 11119310 TI - Meeting Review: The 48th ASMS conference on mass spectrometry and allied topics. AB - The general theme of the meeting was the application of mass spectrometry to pharmaceutical and biotechnological research. The majority of the oral presentations and posters were concerned with the development and application of all mass spectrometric techniques related to proteomics. PMID- 11119311 TI - Website review: interPro (the integrated resource of protein domains and functional sites). AB - The family and motif databases, PROSITE, PRINTS, Pfam and ProDom, have been integrated into a powerful resource for protein secondary annotation. As of June 2000, InterPro had processed 384 572 proteins in SWISS-PROT and TrEMBL. Because the contributing databases have different clustering principles and scoring sensitivities, the combined assignments compliment each other for grouping protein families and delineating domains. The graphic displays of all matches above the scoring thresholds enables judgements to be made on the concordances or differences between the assignments. The website links can be used to analyse novel sequences and for queries across the proteomes of 32 organisms, including the partial human set, by domain and/or protein family. An analysis of selected HtrA/DegQ proteases demonstrates the utility of this website for detailed comparative genomics. Further information on the project can be found at the European Bioinformatics Institute at http://www.ebi.ac.uk/interpro/ PMID- 11119312 TI - Website update: The UK Crop Plant Bioinformatics Network (UK CropNet). AB - UK CropNet currently provides a range of databases (and database-mining tools) to the plant community that are all freely accessible through our website (http://ukcrop.net/). Recent upgrades have meant that we can now expand the range of available facilities (e.g. addition of new databases) whilst also strengthening and improving access to existing services (e.g. providing a BLAST search facility against sequences in our databases). This article will briefly outline these and other new developments in our service. PMID- 11119315 TI - [In Process Citation] PMID- 11119316 TI - [Chagas disease in Brazil]. AB - This article presents the current situation for Chagas disease vectors in Brazil, based on data from the Brazilian National Health Foundation (FNS). Over the course of the last 20 years, continuous chemical control has resulted in a clear reduction of triatomine densities and Trypanosoma cruzi in Brazilian dwellings. Results have been particularly promising in relation to Triatoma infestans and Panstrongylus megistus, considered the most important species in the past. In parallel, data from school serological surveys, hospitalized patients, and mortality records show an important decrease in the disease. Nevertheless, some areas of the Brazilian Northeast and some residual foci of Triatoma infestans and Panstrongylus megistus remain as major challenges for public health authorities, requiring effective epidemiological surveillance. States and municipalities are required to assume this task at present, as the traditional Brazilian National Health Foundation is undergoing decentralization. PMID- 11119317 TI - [General situation and perspectives of chagas disease in Northeastern Region, Brazil]. AB - Primary and secondary data show the importance and distribution of human Chagas disease (HCD) in Northeast Brazil. Among the 27 detected vector species, Triatoma infestans, Panstrongylus megistus, Triatoma brasiliensis and Triatoma pseudomaculata are epidemiologically important. Major medical impact is attributed to T. infestans and P. megistus, the most domiciliated and vulnerable species, while the other two are native and more difficult to control. Regional differences in transmission and medical impact of HCD exist in the Northeast, where in general the disease appears to be less harmful than in other Brazilian regions like the Southeast and State of Goias. There is a downward trend in HCD transmission and morbidity in the Northeast, its control in the region is a cause of concern because of the decommissioning of the National Health Foundation without a corresponding assimilation of its routine activities by regional and municipal institutions. PMID- 11119313 TI - Current awareness on comparative and functional genomics. PMID- 11119318 TI - [Current situation with chagas disease vector control in the Americas]. AB - This article identifies and describes various epidemiological aspects in the natural transmission of Chagas disease in the Americas. It also examines the relative importance of the principal vector species in the disease's transmission and the control levels that are feasible in each instance. Estimations of the population at risk, number of infected cases, and number of chronic cases are presented. Prospects for control are discussed on the basis of past results to predict the expected results with introduced species like Triatoma infestans in the Southern Cone and Rhodnius prolixus in Central America and with the other autochthonous species in areas where they are found. Finally, the article discusses the role of other transmission mechanisms in the maintenance of endemic Chagas disease. PMID- 11119319 TI - [Epidemiological surveillance of Chagas disease]. AB - Chagas disease still constitutes an important medical problem in affected countries. In some, the extent of the disease is still unknown and control programs have not been implemented. In others the disease has been reduced due to regular control programs and other economic and social factors. Epidemiological surveillance with community participation to guard against disease transmission is now the basic challenge. Applied research and in-depth reformulation of health systems are required to establish efficient and sustainable Chagas disease surveillance programs, considering low density of peridomiciliary vectors as the most relevant factor. In addition, a large population of already infected, poor individuals require specific medical attention and social security. As a consequence of health care decentralization, Federal institutions such as the Brazilian National Health Foundation (FNS) are being progressively decommissioned, and new participants must be engaged in the process. Communities themselves, together with regional and local institutions, must take charge of surveillance in order to guarantee its efficiency and sustainability. PMID- 11119320 TI - [Operational aspects of Triatoma brasiliensis control]. AB - Vector control strategies against indigenous species is not easy, due to their capacity to reinvade treated premises from sylvatic ecotopes. Between August 1996 and December 1997 we conducted a study on reinfestation of houses after spraying in a county in the State of Ceara. Of 277 houses examined, 113 (40.8%) were infested (21.7% intradomiciliary and 35.4% peridomiciliary). Of the 433 Triatominae collected, 207 were Triatoma brasiliensis (49% of which intradomiciliary, with a mean of 1.8 insects/house) and 226 were Triatoma pseudomaculata (97% peridomiciliary). The age structure of the two indicated a univoltine development cycle for T. brasiliensis and two cycles per year for T. pseudomaculata. Four months after spraying with deltamethrin SC 25mg ia/m2, 9.7% of the houses were still positive, mainly with peridomestic infestations. Intradomiciliary wall bioassays showed persistence of the insecticide up to 9 months after spraying. Considering the high potential for recolonization of treated premises from sylvatic foci, we propose an operational strategy combining traditional evaluations and community-based surveillance with increased selective interventions and community education. PMID- 11119321 TI - [Microclimatic properties of the Triatoma brasiliensis habitat]. AB - Vector-borne transmission of Chagas disease in Northeast Brazil is basically by Triatoma brasiliensis. It is thus crucial to determine this species' microclimatic preferences as limiting factors for its distribution and ability to infest domestic environments. We analyze the microclimatic properties of the shelters in which these insects are found in wild, domestic, and peridomiciliary environments in the State of Ceara, at Brazil. We measure temperature and relative humidity (RH) every 15 minutes for 3 days. Thermal variation was greatly dampened inside both domiciliary refuges and the more protected internal places in wild stony sites. For RH, we observed a similar dampening pattern, but mean RH was lower in both domiciliary refuges and wild ones inside stony sites as compared to reference levels in the surrounding environment. The results are discussed with regard to this species' microclimatic preferences in the laboratory and its potential as determinants of its geographical distribution. PMID- 11119322 TI - [Peridomiciliary changes and implications for Triatoma brasiliensis control]. AB - A total of 9,906 annexes from 1,541 rural dwellings of Boa Viagem County, Ceara, Brazil, infested by Triatoma brasiliensis and Triatoma pseudomaculata were investigated and immediately sprayed with pyrethroid insecticide, followed by revisions at 6, 12, and 18 months. The initial infestation rate of annexes was 4.0%, with predominant infestation in animal shelters (7.0%). Mean insect density was low, regardless of triatomine species or type of annex. A progressive decrease in the number of initial annexes was observed (66% of remaining annexes), mainly those classified as "piles of materials". Only 3% of the annexes were modified by the population. New constructed annexes were important as new foci of infestation. Some 25% were infested at the end of observation period, significantly more than the "old" annexes (4.0%), a difference attributed to insecticide spraying at the beginning of the intervention. Reinfestation occurred slowly and was more frequent in animal shelters No differences were observed between traditional pyrethroid and slow-release organophosphate formulations. Selective spraying of "new" annexes is recommended. PMID- 11119323 TI - Chromosome homogeneity in populations of Triatoma brasiliensis Neiva 1911 (Hemiptera - reduviidae - triatominae). AB - Triatoma brasiliensis is the most important vector of Chagas disease in the semiarid zone of the Northeast of Brazil. Several authors have reported the occurrence of four chromatic patterns with morphological, ecological, and genetic differences. In order to determine the existence of cytogenetic differentiation between these chromatic forms, we analyzed their karyotypes and the chromosome behavior during the male meiotic process. Triatoma brasiliensis shows distinct and specific chromosome characteristics, which differ from those observed in all other triatomine species. However, no cytogenetic differences were observed between the four chromatic forms of T. brasiliensis. The lack of chromosome differentiation among them could indicate that the populations of this species are in a process of differentiation that does not involve their chromosomal organization. PMID- 11119324 TI - Biosystematics and evolution of the Triatominae. AB - In this paper we summarize the systematics of the 130 currently recognized species of Triatominae and the key features of their evolutionary background. There is increasing evidence that the subfamily has polyphyletic origins, with the various tribes and species groups probably arising from different reduviid lineages in relatively recent times. PMID- 11119325 TI - [Distribution and characterization of different populations of Triatoma brasiliensis Neiva, 1911 (Hemiptera, Reduviidae, Tritominae)]. AB - Triatoma brasiliensis Neiva, 1911 is now considered the most important Chagas disease vector in the semiarid zones of northeastern Brazil. Four distinct populations of T. brasiliensis have been identified by multidisciplinary studies: brasiliensis (Caico, RN), melanica (Espinosa, MG), macromelasoma (Petrolina, PE), and juazeiro (Juazeiro, BA). By scanning electron microscopy of egg exochorion, each population displayed a distinct ornamentation pattern. The brasiliensis, macromelasoma, and juazeiro populations were found in both artificial ecotopes and the wild, while the mel anica population was collected only in the wild. Isoenzymatic analysis detected phenotypic and genetic differences among them, with the melanica population being the most distinct. The brasiliensis population is the most important one from an epidemiological point of view, with the widest geographic distribution, the highest Trypanosoma cruzi infection rate, and occupying a wide variety of ecotopes. PMID- 11119326 TI - [Use of random amplified polymorphic DNA (RAPD) in the populational study of Triatoma brasiliensis Neiva, 1911]. AB - We evaluated the genetic variability of Triatoma brasiliensis, the main vector of Chagas disease in Northeast Brazil, using specimens from three populations. Regions of genomic DNA were amplified by RAPD (Random Amplified Polimorphic DNA), using two primers. The products were visualized after polyacrylamide gel electrophoresis followed by silver staining. A dendrogram constructed through the Dice similarity coefficient allowed for separation of the tested specimens into three distinct groups. The populations captured in areas from Ceara State showed similar profiles, but different from that captured in Piaui State. Our results indicate that RAPD can be used successfully in triatomine studies and suggest the presence of genetic variability between different populations of T. brasiliensis. PMID- 11119327 TI - [Biological potential of Triatoma brasiliensis]. AB - Biological and physiological parameters of Triatoma brasiliensis, Triatoma infestans, and Triatoma pseudomaculata were studied and compared. T. brasiliensis, reared on mice, showed a faster evolutionary cycle from first stage to adult and higher fecundity, when compared to the other species. T. infestans showed the fastest blood intake in both hosts tested, followed by T. brasiliensis and T. pseudomaculata. Clotting tests using salivary gland extracts of T. brasiliensis presented intermediary values of anti-clotting activity when compared to T. infestans and T. pseudomaculata. PMID- 11119328 TI - [Focal and total residual insecticide spraying to control Triatoma brasiliensis and Triatoma pseudomaculata in Northeast Brazil]. AB - To efficiently control the triatomines Triatoma brasiliensis and Triatoma pseudomaculata, a field trial was performed to compare conventional versus focal spraying of deltamethrin 5% SC at 25 mg a. i./m2 and the slow-release organophosphate malathion 8.3% SR at 2g a. i./m2. The assay took place in the county of Boa Viagem, Ceara State, with 1541 households, randomly separated into 4 groups. Two of them received focal spraying: PT, treated with deltamethrin indoors and in the peridomicile, and PL, which received slow-release malathion in the same circumstances. The other groups received conventional, i.e., total application: PT with deltamethrin in the intra- and peridomicile, and PL, which was treated with deltamethrin indoors and slow-release malathion in the peridomicile. Entomological surveys at 6 and 12 months post-treatment showed better results for mixed treatment, the PL group, probably due to good indoor performance for the pyrethroid and better performance of the slow-release formulation under the hostile peridomiciliary conditions. Domestic animal shelters underwent numerous modifications over the course of the year, fostering reduced insecticide performance in the peridomicile. PMID- 11119329 TI - [Epidemiological surveillance in the Chagas disease control program in Minas Gerais State, Brazil (1984-1998)]. AB - This paper describes the surveillance phase of the Chagas Disease Control Program in Minas Gerais State. Surveillance was conducted by the County Health Services with community participation in the planning, decision-making, and maintenance processes and is intended to be sustainable in the context of Brazil's Unified National Health System (SUS). PMID- 11119330 TI - [Health care reform, decentralization, prevention and control of vector-borne diseases]. AB - Economic policies are changing Latin American health programs, particularly promoting decentralization. Numerous difficulties thus arise for the control of endemic diseases, since such activities traditionally depend on vertical, and centralized structures. Theoretical arguments in favor of decentralization notwithstanding, no such tradition exists at the county level. The lack of program expertise at peripheral levels, intensive staff turnover, and even corruption are additional difficulties. Hence, the simple bureaucratic transfer of activities from the Federal to county level is often irresponsible. The loss of priority for control of endemic diseases in Latin America may mean the inexorable extinction of traditional control services. Malaria, dengue fever, and Chagas disease programs are examples of the loss of expertise and effectiveness in Latin America. A better strategy for responsible decentralization is required. In particular, a shared transition involving all governmental levels is desirable to effectively modernize programs. Maintenance of regional reference centers to ensure supervision, surveillance, and training is suggested. PMID- 11119331 TI - Neonatal screening for inborn errors of metabolism. PMID- 11119332 TI - Transcatheter closure of patent ductus arteriosus. AB - OBJECTIVE: To present short and intermediate results of catheter closure of patent ductus artereiosus using spring coils and Amplatzer duct occluder. SETTING: Tertiary care referral hospital in New Delhi. METHOD: 121 patients were diagnosed to be having patency of the arterial duct between October 1996 and December 1999. Their ages ranged between 4 mo and 480 mo (mean 80.5 mo). Before August 1998 only spring coils were used as an alternative to surgical closure of the duct, whereas between August 1998 and December 1999 both coils and device were used. RESULTS: Spring coils were attempted in 48, Amplatzer duct occluder in 44 and 29 patients were referred for surgery. Coils were successfully deployed in 42/48 (87.5%) and device in 42/44 (95.5%) patients. Complete closure of the arterial duct was seen in 96% patients in the coil closure group at the end of one year and 100% closure was achieved in the device closure group at the end of 3 months in all the patients followed. Coil embolised in 11 patients, significant stenosis of left pulmonary occurred in one patient and severe hemolysis requiring blood transfusion occurred in one patient after coil closure. Six patients required a second procedure to close the residual shunt within 6 months of the initial coil deployment. One patient had loss of femoral artery pulses in right leg after device closure and required heparin infusion for 48 hours. There were 8 failed attempts. All these patients have been operated successfully. CONCLUSION: Percutaneous closure of the patent ductus arteriosus using Coils or the Amplatzer device is an acceptable method. Small residual shunts are common after the initial procedure especially after coil deployment but most close spontaneously. PMID- 11119333 TI - Mortality and morbidity in high risk infants during a six year follow-up. AB - OBJECTIVE: To study the mortality and morbidity in high risk infants after discharge from the hospital. DESIGN: Prospective, 6 year follow up. SETTING: High Risk Clinic (HRC). METHODS: Infants discharged from a Neonatal Special Care Unit were identified for follow up using predetermined risk criteria. Home visits were made by the social worker, if appointments to HRC were missed. Verbal autopsy was performed in case of home deaths. Intercurrent illnesses and rehospitalizations were recorded. At six years, the children who had come for the final assessment at 30 months on the Bayley Scales, were recalled for assessment of the intelligence quotient, by Stanford-Binet Scale. RESULTS: Four hundred and four high risk infants and eighty six controls were enrolled. There were 40 deaths in the study period, out of which 38 occurred in the first year of life, sixty per cent of these occurring in the first three months. The mortality was significantly higher in the VLBW group. Out of the 22 hospital deaths, 72.7% were due to infection. There was a significant difference (p < 0.001) in the mortality rate between the group which attended the HRC regularly (6.4%) as compared to that of the defaulters (27.6%). Ninety five children had rehospitalization in the High Risk group as compared to two in the control group (p < 0.001). In the 286 children who were assessed at 6 years, the incidence of borderline intelligence was 14.6% as compared to 5.6% in the controls (p < 0.05). CONCLUSION: Mortality and rehospitalization rate is high in high risk infants, after discharge from the hospital. Children who appear to have normal development in the third year, may show a high incidence (14.6%) of borderline intelligence at six years. PMID- 11119334 TI - Impact of the integrated child development services (ICDS) on maternal nutrition and birth weight in rural Varanasi. AB - OBJECTIVE: To evaluate the impact of ICDS on maternal nutrition and birth weight. SETTING: 28 ICDS and 21 non-ICDS villages in two adjoining blocks of Varanasi. METHODS: 5289 pregnancies were registered during 1987-1993 in these two blocks. In the ICDS block 916 and 1453 nutrition supplemented and unsupplemented, respectively and 1748 of the non-ICDS live births with weight recorded within 48 h formed the study subjects. RESULTS: The ICDS supplemented mothers gained 100g more in pregnancy and birth weight was higher by 58 g (p < 0.05) as compared to unsupplemented ICDS mothers. Birth weight in unsupplemented ICDS areas was 25g higher as compared to non ICDS area. ICDS supplemented women had a significantly smaller proportion of low birth weight babies (14.4%) compared to ICDS unsupplemented (20.4%) and non-ICDS women (26.3%). The corresponding prevalence of preterm births was 2.0, 2.4 and 4.3%, respectively (P < 0.001). Multiple regression analysis showed that increased wight gain in pregnancy, length of gestation, caloric intake and term hemoglobin were significantly associated with birth weight. However, the length of gestation was not influenced by factors improving the birth weight. CONCLUSION: Undernourished pregnant women are benefitted by late pregnancy nutrition supplement PMID- 11119335 TI - Human immunodeficiency virus infection. PMID- 11119336 TI - Evaluation of safety of oral vitamin 'A' megadose co-administered with measles vaccination. PMID- 11119337 TI - Electrolyte abnormalities in children admitted to pediatric intensive care unit. PMID- 11119338 TI - Colorectal carcinoma in Indian children. PMID- 11119339 TI - Relative risk and prevalence of illness related to child labor in a rural block. PMID- 11119340 TI - Preliminary studies on IL-6 levels in healthy and septic Indian neonates. PMID- 11119341 TI - Pseudo-gonococcal ophthalmia neonatorum. PMID- 11119342 TI - Bilateral adrenal cysts in a newborn. PMID- 11119343 TI - Engelmann's disease with cardiomyopathy. PMID- 11119344 TI - Xeroderma pigmentosum with multiple malignancies. PMID- 11119345 TI - Basal artery occlusion with hyperlipoproteinemia type IIb causing ischemic stroke in an infant. PMID- 11119346 TI - Asymmetric crying facies. PMID- 11119347 TI - Hypomelanosis of Ito. PMID- 11119348 TI - Testing vaccine potency. PMID- 11119350 TI - Jaundice in hepatitis B immunized. PMID- 11119352 TI - Factor x deficiency: an unusual cause of spontaneous intracranial bleeding. PMID- 11119353 TI - Effect of antenatal steroids on lung maturity and brain development. PMID- 11119355 TI - Diphtheria--certainly not out. PMID- 11119357 TI - Polio eradication strategy: need for re-appraisal. PMID- 11119359 TI - Issues related to the psychological aspects of recurrent abdominal pain. PMID- 11119361 TI - Use of cisapride. PMID- 11119363 TI - Non-invasive estimation of total serum bilirubin. PMID- 11119394 TI - A prospective, observational study of postmenopausal hormone therapy and primary prevention of cardiovascular disease. AB - BACKGROUND: Most primary prevention studies have found that long-term users of postmenopausal hormone therapy are at lower risk for coronary events, but numerous questions remain. An adverse influence of hormone therapy on cardiovascular risk has been suggested during the initial year of use; however, few data are available on short-term hormone therapy. In addition, the cardiovascular effects of daily doses of oral conjugated estrogen lower than 0.625 mg are unknown, and few studies have examined estrogen plus progestin in this regard. OBJECTIVE: To investigate duration, dose, and type of postmenopausal hormone therapy and primary prevention of cardiovascular disease. DESIGN: Prospective, observational cohort study. SETTING: Nurses' Health Study, with follow-up from 1976 to 1996. PATIENTS: 70 533 postmenopausal women, in whom 1258 major coronary events (nonfatal myocardial infarction or fatal coronary disease) and 767 strokes were identified. MEASUREMENTS: Details of postmenopausal hormone use were ascertained by using biennial questionnaires. Cardiovascular disease was established by using a questionnaire and was confirmed by medical record review. Logistic regression models were used to calculate relative risks and 95% CIs, adjusted for confounders. RESULTS: When all cardiovascular risk factors were considered, the risk for major coronary events was lower among current users of hormone therapy, including short-term users, compared with never-users (relative risk, 0.61 [95% CI, 0.52 to 0.71]). Among women taking oral conjugated estrogen, the risk for coronary events was similarly reduced in those currently taking 0.625 mg daily (relative risk, 0.54 [CI, 0.44 to 0.67]) and those taking 0.3 mg daily (relative risk, 0.58 [CI, 0. 37 to 0.92]) compared with never-users. However, the risk for stroke was statistically significantly increased among women taking 0.625 mg or more of oral conjugated estrogen daily (relative risk, 1.35 [CI, 1.08 to 1.68] for 0.625 mg/d and 1.63 [CI, 1.18 to 2.26] for >/=1.25 mg/d) and those taking estrogen plus progestin (relative risk, 1.45 [CI, 1.10 to 1.92]). Overall, little relation was observed between combination hormone therapy and risk for cardiovascular disease (major coronary heart disease plus stroke) (relative risk, 0.91 [CI, 0.75 to 1.11]). CONCLUSIONS: Postmenopausal hormone use appears to decrease risk for major coronary events in women without previous heart disease. Furthermore, 0.3 mg of oral conjugated estrogen daily is associated with a reduction similar to that seen with the standard dose of 0.625 mg. However, estrogen at daily doses of 0.625 mg or greater and in combination with progestin may increase risk for stroke. PMID- 11119395 TI - The positive predictive value of cervical smears in previously screened postmenopausal women: the Heart and Estrogen/progestin Replacement Study (HERS). AB - BACKGROUND: The benefits and risks of performing annual cervical smears on postmenopausal women are not well defined. The independent effect of hormone replacement therapy on development of cytologic abnormalities is unknown. OBJECTIVE: To determine the positive predictive value of cervical smears in previously screened postmenopausal women and to determine the effect of oral estrogen plus progestin on incident cervical cytologic abnormalities. DESIGN: Prospective cohort study (incidence) and randomized, double-blind, placebo controlled trial (hormone therapy). SETTING: 20 U.S. outpatient and community clinical centers. PARTICIPANTS: 2561 women with a uterus and normal cytologic characteristics at baseline. INTERVENTIONS: Annual smears; oral conjugated equine estrogens, 0. 625 mg/d, plus medroxyprogesterone acetate, 2.5 mg/d, or identical placebo. MEASUREMENTS: Incident cytologic abnormalities (atypical squamous cells of undetermined significance, atypical glandular cells of undetermined significance, low-grade squamous epithelial lesion, and high-grade squamous epithelial lesion) and final histologic diagnoses. RESULTS: The incidence of new cytologic abnormalities in the 2 years following a normal smear was 110 per 4895 person-years (23 per 1000 person-years [95% CI, 18 to 27 per 1000 person-years]). Among the 103 women with known histologic diagnoses, one had mild to moderate dysplasia. The positive predictive value of any smear abnormality identified 1 year after a normal smear, therefore, was 0% (CI, 0% to 5.0%) (0 of 78 women); the positive predictive value of abnormalities found within 2 years was 0.9% (CI, 0.0% to 3.0%) (1 of 110 women). In hormone-treated compared with non-hormone treated women, the incidence of cytologic abnormalities was nonsignificantly higher (relative hazard, 1.36 [CI, 0.93 to 1.99]), largely because of a nonsignificant 58% greater incidence of atypical squamous cells of undetermined significance (relative hazard, 1.58 [CI, 0.99 to 2.52]). CONCLUSIONS: Because of a poor positive predictive value, cervical smears should not be performed within 2 years of normal cytologic results in postmenopausal women. Therapy with oral estrogen plus progestin does not significantly affect the incidence of cytologic abnormalities. PMID- 11119396 TI - Association of hypogonadism and estradiol levels with bone mineral density in elderly men from the Framingham study. AB - BACKGROUND: Both hypogonadism and low estrogen levels adversely affect bone health in young men. In elderly men, who are at greatest risk for osteoporotic fracture, the influence of hypogonadism on bone mineral density remains unclear, as does the relative effect of estrogen status compared to hypogonadism. OBJECTIVE: To examine the relation of hypogonadism and estrogen status to bone mineral density in elderly men. DESIGN: Community-based, prospective cohort study. SETTING: Framingham, Massachusetts. PATIENTS: Male participants of the Framingham Study. MEASUREMENTS: Total testosterone, total estradiol, and luteinizing hormone were measured in participants at all four biennial examinations from 1981 to 1989. Values from at least three of four examinations were averaged. Hypogonadism was defined as a mean testosterone level less than 10.4 nmol/L (<3.0 ng/mL) or a mean luteinizing hormone level of 20 IU/L or greater. An alternate definition of hypogonadism based only on a mean testosterone level less than 10.4 nmol/L (<3.0 ng/mL) was also used. In 1988 1989, bone mineral density was measured at the proximal femur (femoral neck, Ward triangle, and trochanter) and lumbar spine by using dual-photon absorptiometry and at the radial shaft by using single-photon absorptiometry. The association of hypogonadism with bone mineral density was examined with adjustment for confounders, including estradiol levels. A similar model that adjusted for hypogonadism was used to examine the association of estradiol level (ranked as quartiles) with bone mineral density. RESULTS: Of 448 men with bone mineral density measurements, 405 had evaluable hormone levels (mean age, 75.7 years [range, 68 to 96 years]); 71 (17.5%) of the 405 men were hypogonadal. Bone mineral density at any site did not significantly differ in hypogonadal men compared with eugonadal men (for example, bone mineral density at the femoral neck was 0.89 g/cm(2) vs. 0.87 g/cm(2), respectively; P > 0.2), even when alternate definitions of hypogonadism were used. In contrast, compared with the lowest estradiol quartile, men with higher estradiol levels had greater mean bone mineral density at all sites (for example, bone mineral density at the femoral neck was 0.84 g/cm(2), 0.88 g/cm(2), 0.86 g/cm(2), and 0.91 g/cm(2) from the lowest to the highest estradiol quartile; P for trend = 0.002). The difference in mean bone mineral density between men in the lowest and those in the highest estradiol quartile levels was similar to the effect of 10 years of aging on bone mineral density. CONCLUSIONS: In elderly men, hypogonadism related to aging has little influence on bone mineral density, but serum estradiol levels have a strong and positive association with bone mineral density. PMID- 11119397 TI - Severe liver injury after treatment with the leukotriene receptor antagonist zafirlukast. AB - BACKGROUND: In registration trials, zafirlukast, an asthma medication, caused asymptomatic elevated aminotransferase levels in up to 5% of participants. Until now, however, no cases of severe hepatitis attributed to zafirlukast have been reported. OBJECTIVE: To report the clinical characteristics of three patients with severe hepatitis due to zafirlukast. DESIGN: Case report. SETTING: One community hospital and two university hospitals. PATIENTS: Three middle-aged women taking zafirlukast, 20 mg twice per day. INTERVENTION: Discontinuation of zafirlukast therapy in three patients, steroid therapy in two patients, and orthotopic liver transplantation in one patient. MEASUREMENTS: Serum aminotransferase and bilirubin levels, standard blood tests for causes of hepatitis other than drug toxicity, and liver biopsy in two patients. RESULTS: Patient 1 recovered spontaneously, had a severe relapse after inadvertent rechallenge with the medication, and ultimately made a complete recovery. Patient 2 developed subfulminant hepatic failure and required liver transplantation. Patient 3 developed severe hepatitis that improved after treatment with corticosteroids. Liver tissue was available from two patients and showed histologic changes commonly associated with drug reactions. CONCLUSION: Patients receiving zafirlukast may develop severe liver injury and should be observed for signs and symptoms of hepatitis. PMID- 11119399 TI - Update in critical care medicine. PMID- 11119398 TI - Cardiovascular effects of 3,4-methylenedioxymethamphetamine. A double-blind, placebo-controlled trial. AB - BACKGROUND: The psychoactive stimulant 3, 4-methylenedioxymethamphetamine (MDMA), also known as "ecstasy," is widely used in nonmedical settings. Little is known about its cardiovascular effects. OBJECTIVE: To evaluate the acute cardiovascular effects of MDMA by using transthoracic two-dimensional and Doppler echocardiography. DESIGN: Four-session, ascending-dose, double-blind, placebo controlled trial. SETTING: Urban hospital. PATIENTS: Eight healthy adults who self-reported MDMA use. INTERVENTION: Echocardiographic effects of dobutamine (5, 20, and 40 microg/kg of body weight per minute) were measured in a preliminary session. Oral MDMA (0.5 and 1.5 mg/kg of body weight) or placebo was administered 1 hour before echocardiographic measurements in three weekly sessions. MEASUREMENTS: Heart rate and blood pressure were measured at regular intervals before and after MDMA administration. Echocardiographic measures of stroke volume, ejection fraction, cardiac output, and meridional wall stress were obtained 1 hour after MDMA administration and during dobutamine infusions. RESULTS: At a dose of 1.5 mg/kg, MDMA increased mean heart rate (by 28 beats/min), systolic blood pressure (by 25 mm Hg), diastolic blood pressure (by 7 mm Hg), and cardiac output (by 2 L/min). The effects of MDMA were similar to those of dobutamine, 20 and 40 microg/kg per minute. Inotropism, measured by using meridional wall stress corrected for ejection fraction, decreased after administration of dobutamine, 40 microg/kg per minute, but did not change after either dose of MDMA. CONCLUSIONS: Modest oral doses of MDMA increase heart rate, blood pressure, and myocardial oxygen consumption in a magnitude similar to dobutamine, 20 to 40 microg/kg per minute. In contrast to dobutamine, MDMA has no measurable inotropic effects. PMID- 11119400 TI - Uncomplicated acute bronchitis. AB - Acute bronchitis is an acute cough illness in otherwise healthy adults that usually lasts 1 to 3 weeks. This review describes the pathophysiology of the condition and provides a practical approach to the evaluation and treatment of adults with uncomplicated acute bronchitis. Practical points to be made are:1. Respiratory viruses appear to cause the large majority of cases of uncomplicated acute bronchitis.2. Pertussis infection is present in up to 10% to 20% of adults with cough illness of more than 2 to 3 weeks' duration. No clinical features distinguish pertussis from nonpertussis infection in adults who were immunized against pertussis as children.3. Transient bronchial hyperresponsiveness appears to be the predominant mechanism of the bothersome cough of acute bronchitis.4. Ruling out pneumonia is the primary objective in evaluating adults with acute cough illness in whom comorbid conditions and occult asthma are absent or unlikely. In the absence of abnormalities in vital signs (heart rate > 100 beats/min, respiratory rate > 24 breaths/min, and oral body temperature > 38 degrees C), the likelihood of pneumonia is very low.5. Randomized, placebo controlled trials do not support routine antibiotic treatment of uncomplicated acute bronchitis.6. Randomized, placebo-controlled trials have shown that inhaled albuterol decreases the duration of cough in adults with uncomplicated acute bronchitis.7. Intervention studies suggest that antibiotic treatment of acute bronchitis can be reduced by using a combination of patient and physician education. Decreased rates of antibiotic treatment are not associated with increased utilization, return visits, or dissatisfaction with care. PMID- 11119401 TI - Influenza: prospects for control. AB - Influenza is a disease of antiquity that annually imposes a major burden of morbidity and mortality. The available inactivated vaccine is effective for preventing influenza and the serious disease and death that can accompany it. However, annual recommendations for vaccination among persons at risk have never been adequately implemented. This remains the most pressing current need for control of influenza. Amantadine, rimantadine, and the newly available drugs zanamivir and oseltamivir are effective for influenza prevention and treatment (the former two for influenza A only). The availability of four antiviral agents that effectively prevent and treat influenza provides the physician with considerable flexibility for their use in influenza control. Optimal application of the currently available vaccine and antiviral agents should substantially reduce the impact of influenza. Other methods for influenza treatment and control are under development, and a live attenuated vaccine with substantial potential for control is nearing approval. However, better inactivated vaccines, better rapid diagnostic tests, and an increased understanding of options for use of antiviral agents are still needed. When all of these things are available and optimally applied, effective control of influenza should result. The prospect is compelling. Full participation by the practicing physician will be necessary to achieve this goal. PMID- 11119402 TI - Hormones to prevent coronary disease in women: when are observational studies adequate evidence? PMID- 11119403 TI - Androgens, estrogens, and bone in men. PMID- 11119404 TI - The department of card tricks and close magic. PMID- 11119405 TI - Management of suspected ventilator-associated pneumonia. PMID- 11119406 TI - Management of suspected ventilator-associated pneumonia. PMID- 11119407 TI - Management of Suspected Ventilator-Associated Pneumonia. PMID- 11119408 TI - Liver disease and home parenteral nutrition. PMID- 11119409 TI - Noncardiogenic pulmonary edema in marathon runners. PMID- 11119411 TI - Noncardiogenic Pulmonary Edema in Marathon Runners. PMID- 11119410 TI - Noncardiogenic pulmonary edema in marathon runners. PMID- 11119412 TI - Gastrointestinal Disease in Primary Care. PMID- 11119413 TI - Primary Care Provider's Guide to Cardiology. PMID- 11119415 TI - Developments in cervical and ovarian cancer screening: implications for current practice. PMID- 11119416 TI - Generation of gene-modified T cells reactive against the angiogenic kinase insert domain-containing receptor (KDR) found on tumor vasculature. AB - The destruction of newly forming tumor vasculature is a promising approach to inhibit tumor growth. The goal of the present study was to investigate whether human lymphocytes gene modified to express a chimeric receptor specific for the angiogenic endothelial cell receptor, KDR, could react against KDR(+) cells. Gene modified lymphocytes specifically lysed KDR(+) cells and secreted cytokines in response to KDR(+) target cells including human umbilical vein endothelial cells (HUVECs). Anti-KDR lymphocytes induced HUVECs to secrete the chemokine interleukin 8 and upregulate the adhesion molecules VCAM and E-selectin, which may be important in the recruitment of further immune effector cells to tumor. These KDR-specific lymphocytes may be useful in the adoptive immunotherapy of a broad range of cancers by inducing immune-mediated destruction of tumor neovasculature. PMID- 11119417 TI - Eradication of breast cancer xenografts by hyperthermic suicide gene therapy under the control of the heat shock protein promoter. AB - To investigate the usefulness of heat shock protein (HSP) promoter for breast cancer gene therapy, hyperthermia and HSV thymidine kinase (tk) suicide gene combination therapy was examined with mouse mammary cancer cell line FM3A. HSP promoter activity was markedly increased after heat shock (41-45 degrees C), with maximum activation (about 400-fold) at 3 hr. An in vitro cytotoxic assay showed that HSP-tk-transduced FM3A cells became more sensitive (more than 50,000 times) to ganciclovir (GCV) with heat shock, but untreated cells showed no increased cytotoxic sensitivity to GCV compared with control FM3A cells. In addition to promoter-oriented selective cell killing, a "chemosensitization effect" as a bystander effect was demonstrated by hyperthermia and suicide gene combination therapy, using a non-heat-inducible promoter. Immunohistochemical analysis revealed that this synergistic killing effect was dependent on apoptotic cell death with upregulation of both Fas and FasL (Fas ligand) expression. We also examined the efficacy of HSP-tk gene therapy in vivo by implanting breast cancer in subcutaneous and intraperitoneal models of BALB/c nude mice targeted by the HVJ-anionic liposome method. Significant tumor regression was observed in HSP-tk transduced tumors followed by hyperthermia therapy, but no such inhibition was noted in either the mock vector transfection or hyperthermia group compared with control tumor-bearing mice. Our results demonstrate that this combination system is synergistically effective in mediating Fas-dependent apoptosis for a specific gene therapy targeting HSP-expressing mammary carcinomas, even in advanced and heat-resistant breast cancer. PMID- 11119418 TI - Effects of preexisting immunity on the response to herpes simplex-based oncolytic viral therapy. AB - Herpes simplex viruses (HSV) type 1 are the basis of a number of anticancer strategies that have proven efficacious in animal models. They are natural human pathogens and the majority of adults have anti-HSV immunity. The current study examined the effect of preexisting immunity on the response to herpes-based oncolytic viral treatment of hepatic metastatic cancer in a murine model designed to simulate a clinical approach likely to be utilized for nonneurological tumors. Specifically, the anticancer effects of NV1020 or G207, two multimutated HSV-1 oncolytic viruses, were tested in immunocompetent mice previously immunized with a wild-type herpes simplex type 1 virus. Mice were documented to have humoral as well as cell-mediated immunity to HSV-1. Tumor response to oncolytic therapy was not measurably abrogated by immunity to HSV at the doses tested. The influence of route of viral administration was also tested in models of regional hepatic arterial and intravenous therapy. Route of viral administration influenced efficacy, as virus delivered intravenously produced some detectable attenuation while hepatic arterial therapy remained unaffected. These results demonstrate that when given at appropriate doses and in reasonable proximity to tumor targets, HSV-based oncolytic therapy can still be expected to be effective treatment for patients with hepatic malignancies. PMID- 11119419 TI - Preservation of graft-versus-infection effects after suicide gene therapy for prevention of graft-versus-host disease. AB - The main complications following allogeneic hematopoietic stem cell transplantation are graft-versus-host disease and poor immune reconstitution leading to severe infections. Mature donor T cells present in the transplant facilitate T cell reconstitution in adults, but also induce graft-versus-host disease, which itself impairs immune reconstitution. Thus, infusing a large number of donor T cells with a diverse repertoire should accelerate functional immune reconstitution after transplantation, only if graft-versus-host disease can be controlled. We previously demonstrated that preventive treatment with ganciclovir could control graft-versus-host disease in mice if donor T cells are made to express viral thymidine kinase as a "suicide" gene. Here we evaluated the recovery of functional antiviral immune responses in such mice. Irradiated mice received an allogeneic hematopoietic stem cell transplantation with thymidine kinase-expressing T cells and were protected from graft-versus-host disease by ganciclovir treatment, and then challenged with lymphocytic choriomeningitis virus. Grafted mice could mount efficient antilymphocytic choriomeningitis virus immune responses leading to viral elimination. Furthermore, when transplanted cells were obtained from mice previously immunized against lymphocytic choriomeningitis virus, grafted mice developed memory-type accelerated responses against the virus. We conclude that efficient graft-versus-infection effects can be mediated by naive T cells and memory donor T cells that persist after suicide gene therapy for prevention of graft-versus-host disease. PMID- 11119421 TI - Comprehensive analysis of the acute toxicities induced by systemic administration of cationic lipid:plasmid DNA complexes in mice. AB - A major limitation associated with systemic administration of cationic lipid:plasmid DNA (pDNA) complexes is the vector toxicity at the doses necessary to produce therapeutically relevant levels of transgene expression. Systematic evaluation of these toxicities has revealed that mice injected intravenously with cationic lipid:pDNA complexes develop significant, dose-dependent hematologic and serologic changes typified by profound leukopenia, thrombocytopenia, and elevated levels of serum transaminases indicative of hepatocellular necrosis. Vector administration also induced a potent inflammatory response characterized by complement activation and the induction of the cytokines IFN-gamma, TNF-alpha, IL 6, and IL-12. These toxicities were found to be transient, resolving with different kinetics to pretreatment levels by 14 days posttreatment. The toxic syndrome observed was independent of the cationic lipid:pDNA ratio, the cationic lipid species, and the level of transgene expression attained. Mechanistic studies determined that neither the complement cascade nor TNF-alpha were key mediators in the development of these characteristic toxicities. Administration of equivalent doses of the individual vector components revealed that cationic liposomes or pDNA alone did not generate the toxic responses observed with cationic lipid:pDNA complexes. Only moderate leukopenia was associated with administration of cationic liposomes or pDNA alone, while only mild thrombocytopenia was noted in pDNA-treated animals. These results establish a panel of objective parameters that can be used to quantify the acute toxicities resulting from systemic administration of cationic lipid:pDNA complexes, which in turn provides a means to compare the therapeutic indices of these vectors. PMID- 11119420 TI - Differentiation and expansion of lentivirus vector-marked dendritic cells derived from human CD34(+) cells. AB - The in vitro genetic manipulation of dendritic cells (DCs) for the expression of foreign proteins or peptides will assist in the development of immunotherapeutic approaches to treat cancer, immunological disorders, and/or infectious diseases. Reports have shown the expansion and differentiation of CD34(+) progenitor cells into mature DCs. In this article we describe the differentiation and expansion of lentivirus vector-marked DCs from umbilical cord blood, bone marrow, and cytokine mobilized peripheral blood CD34(+) cells in the presence of GM-CSF, TNF-alpha, SCF, Flt-3, and IL-4. Lentivirus-marked DCs expressed high levels of enhanced green fluorescent protein and the characteristic DC surface markers CD1a, CD83, HLA-DR, and CD80. Transduced DCs activated allogeneic CD3(+) T cells as efficiently as control (nontransduced) DCs in mixed lymphocyte reactions. These results demonstrate the potential utility of lentivirus-transduced DCs in future immunotherapy protocols. PMID- 11119422 TI - Preincubation with endothelial cell monolayers increases gene transfer efficiency into human bone marrow CD34(+)CD38(-) progenitor cells. AB - Retroviral gene transfer studies targeting bone marrow CD34(+)CD38(-) stem cells have been disappointing because of the rarity of these cells, their G(0) cell cycle status, and their low or absent expression of surface retroviral receptors. In this study, we examined whether preincubation of bone marrow CD34(+)CD38(-) stem cells with a hematopoietically supportive porcine microvascular endothelial cell line (PMVECs) could impact the cell cycle status and expression of retroviral receptors in pluripotent CD34+CD38- cells and the efficiency of gene transfer into these primitive target cells. PMVEC coculture supplemented with GM CSF + IL-3 + IL-6 + SCF + Flt-3 ligand induced >93% of the CD34(+)CD38(-) population to enter the G(1) or G(2)/S/M phase while increasing this population from 1.4% on day 0 to 6.5% of the total population by day 5. Liquid cultures supplemented with the identical cytokines induced 73% of the CD34(+)CD38(-) population into cell cycle but did not maintain cells with the CD34(+)CD38(-) phenotype over time. We found no significant increase in the levels of AmphoR or GaLVR mRNA in PMVEC-expanded CD34(+)CD38(-) cells after coculture. Despite this, the efficiency of gene transfer using either amphotropic vector (PA317) or GaLV vector (PG13) was significantly greater in PMVEC-expanded CD34(+)CD38(-) cells (11.4 +/- 5.6 and 10.9 +/- 5.2%, respectively) than in either steady state bone marrow CD34(+)CD38(-) cells (0.6 +/- 1.7 and 0.2 +/- 0.6%, respectively; p < 0.01 and p < 0.01) or liquid culture-expanded CD34(+)CD38(-) cells (1.4 +/- 3.5 and 0.0%, respectively; p < 0.01 and p < 0.01). Since PMVEC coculture induces a high level of cell cycling in human bone marrow CD34(+)CD38(-) cells and expands hematopoietic cells capable of in vivo repopulation, this system offers potential advantages for application in clinical gene therapy protocols. PMID- 11119423 TI - Analysis of testes and semen from rabbits treated by intravenous injection with a retroviral vector encoding the human factor VIII gene: no evidence of germ line transduction. AB - In a phase 1 clinical trial, we are evaluating a murine leukemia virus (MuLV) based retroviral vector encoding the human factor VIII gene [hFVIII(V)], administered intravenously, as a therapy for hemophilia A. Preclinical biolocalization studies in adult rabbits revealed vector-specific PCR signals in testis tissue at low levels. In follow-up animal studies we used PCR to (1) estimate the frequency with which a given cell in testis tissue is transduced, and (2) determine whether a positive PCR signal could be detected in semen samples from animals treated with hFVIII(V). Using the 99% confidence bound, results indicate that the probability that a given cell within the testis was transduced is less than 1/709,000 (97 days after treatment). This probability decreased with time after hFVIII(V) administration. Moreover, the rate of provector sequence detection in semen samples collected weekly throughout two cycles of spermatogenesis was 3/4281 reactions (0.07%), which is lower than the rate of false positives (1/800, 0.125%) observed for control animals. Using PCR assays with single-copy sensitivity, we have shown that the small number of transduced cells present in testis tissue does not give rise to detectable transduced cells in semen. PMID- 11119424 TI - Future meeting dates and protocol submission deadlines for public recombinant DNA advisory committee (RAC) review PMID- 11119425 TI - Human gene marker/therapy clinical protocols (complete updated listings). PMID- 11119426 TI - Human gene transfer protocols: summary table. PMID- 11119427 TI - Africa matters. PMID- 11119428 TI - Nocardia asteroides pericarditis in association with HIV. AB - This case report describes Nocardia pericarditis in a newly diagnosed human immunodeficiency virus (HIV) patient as an initial manifestation. Previously, two cases of Nocardia pericarditis were reported in patients with established HIV infection. To our knowledge this is the first case of Nocardia pericarditis as an initial manifestation of HIV infection. This case substantiates and emphasizes the importance of identifying Nocardia as an infectious cause of pericarditis in patients with acquired immunodeficiency. Long-term survival may be achieved with a combined medical and surgical approach. PMID- 11119429 TI - Oral lesions in HIV/AIDS patients undergoing highly active antiretroviral treatment including protease inhibitors: a new face of oral AIDS? AB - The objective of this work was to assess the prevalence of human immunodeficiency virus-related oral lesions (HIV-ROL) in HIV-positive/acquired immunodeficiency syndrome (AIDS) patients receiving highly active antiretroviral therapy (HAART) including HIV-protease inhibitors. One hundred fifty-five (154) AIDS patients (69 intravenous drug users [IDU], 53 heterosexuals, 29 males who have sex with males, 1 transfused, and 2 of unknown contagious source) receiving HAART, were examined. We found the following prevalences: HIV-ROL 53.2%; oral candidiasis 34.4%; hairy leucoplakia 26.6%; xerostomia 15.5%; herpes simplex labialis 1.9%; HIV/periodontitis-gingivitis 0.6%. No cases of Kaposi's sarcoma were observed. The highest prevalence of HIV-ROL was found in the IDU group, and in patients with viral load more than 10,000 copies and CD4(+) cell count less than 200. Using our historical controls, this suggests that the prevalence of all oral lesions, particularly oral candidiasis, herpes simplex labiali, Kaposi's sarcoma, and periodontal disease has decreased more than 30% after the institution of HAART. PMID- 11119430 TI - HIV-associated nephropathy: case study and review of the literature. AB - Human immunodeficiency virus type 1 (HIV-1)-seropositive patients are at risk for the development of a variety of acute and chronic renal diseases. The most common cause of chronic renal failure in HIV-1-seropositive patients is HIV-associated nephropathy (HIVAN). HIVAN occurs almost exclusively in black patients and the majority of published cases are of patients who present with acquired immunodeficiency syndrome (AIDS). This disease is currently the third leading cause of end-stage renal disease in blacks aged 20-64. Because HIV-1-seropositive patients may develop a wide variety of acute and chronic renal diseases, definitive diagnosis requires renal biopsy. Emerging data suggest a direct role of HIV-1 infection of kidney cells in the pathogenesis of HIVAN. There have been no well-controlled clinical trials in the treatment of HIVAN. The therapeutic agents with the most promise are angiotensin-converting enzyme inhibitors and antiretroviral medications. Long-term renal prognosis may be changing in the setting of improved aggressive antiretroviral therapy. Patient survival is determined primarily by the stage of HIV-1 infection. In this article, we present the case history of a patient who developed HIVAN. We then review the current literature concerning the epidemiology, differential diagnosis, etiology, and treatment of HIVAN. PMID- 11119431 TI - Characteristics of HIV-1-infected patients with CD4:CD8 lymphocyte ratio normalization on antiretroviral therapy. AB - Antiretroviral therapy for human immunodeficiency virus (HIV)-infection results in normalization of CD4:CD8 T-lymphocyte ratio in about 6% of cases. The T-cell ratio normalization on therapy was associated with a baseline CD4 of over 350 cells per microliter and a T-cell ratio of over 0.5 (for each, p < 0.01), but not with the current level of viral load suppression or compliance with clinic appointments (for each, p > 0.05). The patients with T-cell ratio normalization had a baseline median CD4 count of 428 cells per microliter (range, 353-883 cells per microliter) and a median T-cell ratio of 0.75 (range, 0.54-0.87). We could not address the effect of baseline viral load on T-cell ratio normalization, but there was no association with age, gender, race, HIV risk factor, or the length of antiretroviral therapy. PMID- 11119432 TI - Serologic examination of hepatitis B infection and immunization in HIV-positive youth and associated risks. The Pediatric AIDS Clinical Trials Group Protocol 220 Team. AB - This seroprevalence report examines serologic evidence of hepatitis B immunization or infection and associated demographic/behavioral factors in adolescent (aged 12-20) subjects enrolled in a nontherapeutic clinical trial at 43 Pediatric AIDS Clinical Trials Group (PACTG) clinical centers. Subjects (n = 94) infected with the human immunodeficiency virus (HIV) through sexual activity were categorized as hepatitis B virus (HBV)-immunized, HBV-infected, or nonimmune by hepatitis B serology performed on specimens collected within the subject's first 48 weeks on study (1993-1995). Sixteen percent of the 94 serologically classified subjects were immunized; 19% HBV-infected; 65% nonimmune. Of the three risk factor scores examined (sociodemographic, sexual, and substance abuse), substance use alone demonstrated a significant difference among groups (despite virtually no reported injecting drug behavior), with the sexual risk score exhibiting marginally significant differences. Logistic regression analysis (restricted to nonimmunized subjects) showed that male-male sexual activity raised the odds of HBV infection by a factor of 5.14 (95% confidence interval [CI]: 1.45-18. 23) relative to heterosexual activity; and that for every one point increase on the substance abuse risk scale the odds of infection increased 5% (95% CI: 0.99-1.10). The HBV infection rate in PACTG 220 HIV-positive females is twice United States population-based rates; the rate in PACTG 220 HIV-positive males is nearly seven times higher. Past immunization efforts in this population appear to have been based on sexual activity volume without regard to injecting drug use in sex partners. PMID- 11119434 TI - HIV/AIDS case histories: iron deficiency anemia and altered iron metabolism in HIV infection. PMID- 11119433 TI - Variations in perceived pain associated with emotional distress and social identity in AIDS. AB - This article examines associations between self-perceptions of pain and associated pain distress to gender, ethnicity and religion, health care, health status, and emotional distress. Data were collected through interviews collected in participants' homes. Participants were 151 adults with diagnoses of advanced human immunodeficiency virus (HIV) or acquired immunodeficiency syndrome (AIDS). Time since diagnosis, health status, health care, ethnicity, gender, religion, and emotional distress were examined as mediators of pain symptoms, pain distress, and anticipatory pain. Almost all participants (83%) reported AIDS related pain in the last 3 months. Unexpectedly, pain was negatively associated with time since diagnosis with AIDS. Pain symptoms and pain distress tended to vary by ethnicity, with Latinos expressing more symptoms and pain distress than African Americans. Anticipatory pain varied significantly by gender and religion, with women, Catholics, and Protestants anticipating pain more than men and non Christians. Anxiety, depression, and general emotional distress were significantly associated with pain symptoms (r = 0.44, 0.33, 0.47) and pain distress (r = 0.34, 0.31, 0.34). Health status and health care were unrelated to pain symptoms, pain distress, or anticipatory pain. Pain is a common problem for people living with HIV/AIDS. Self-reported pain is associated with cultural factors and changes in illness status. Clinicians' attention to patients' emotional distress, depression, and anxiety may assist in interventions for pain management. PMID- 11119435 TI - Four-drug regimens may be better. PMID- 11119436 TI - Pap smear intervals safe. PMID- 11119437 TI - Urology as practiced in the Canadian Health Care System. PMID- 11119438 TI - Advances in the management of localized renal cell cancer. AB - Notwithstanding recent advances in our understanding of the genetics and biology of renal cell carcinoma (RCC), surgery remains the mainstay of curative treatment for this disease. Nevertheless, the role of surgery is changing with respect to both localized RCC and patients with metastatic disease. In this article, we review recent advances in the evolution and management of patients with localized RCC. PMID- 11119439 TI - Long term outcome and cost in the management of stage I testicular seminoma. AB - PURPOSE: To validate the use of surveillance as an alternative to adjuvant RT in clinical stage I seminoma, we analyzed our experience with the two approaches in terms of long term outcome and cost. PATIENTS AND METHODS: Between January 1981 and December 1994, 471 patients with stage I testicular seminoma were treated at our institution. Of these, 245 patients received post-operative RT (25 Gy) to the retroperitoneal lymph nodes, and 226 have been managed with surveillance following orchidectomy. Two patients were included in this series twice; both had RT previously for seminoma, were placed on surveillance for a contralateral seminoma and were analyzed for outcome of both primary tumors. The costs associated with both approaches were estimated in 1994 Canadian dollars (C$). RESULTS: With a median follow-up of 7.7 years in the surveillance patients, and 9.7 years in the adjuvant RT cohort, the 5 year actuarial survival for all patients was 97% and the cause-specific survival (CSS) was 99.8%. Of the 226 patients on surveillance 37 patients have relapsed to date; five of those developed a second relapse. One patient has died of disease. Of the 245 patients treated with adjuvant RT, 14 patients have relapsed and none had a second relapse. The CSS was 100%. Thirteen patients on surveillance (5.7%) and 10 patients treated with post-operative RT (4.1%) have received chemotherapy as part of their management. One hundred and eighty-nine patients on surveillance have received no post-orchidectomy treatment to date. Surveillance was more expensive with an average additional cost per patient per year of Can$2620 over 10 years. CONCLUSIONS: Both adjuvant RT and surveillance give excellent results in stage I seminoma. The documented increased risk of second malignant tumors following RT must be taken into account when considering the additional cost of surveillance. The routine use of post-operative RT in stage I seminoma should be reconsidered and a surveillance program offered to all patients as an alternative management option. PMID- 11119441 TI - Economic evaluation of NMP22 in the management of bladder cancer. AB - The purpose of this study was to evaluate, from the perspective of the Quebec health system, the cost and cost-efficacy of using NMP22 compared with the currently recommended monitoring procedure following a transurethral resection of a bladder tumor (TURBT). This evaluation was based on results from the study by Soloway, et al. It was performed using a decision analysis technique which compared a follow-up modality using NMP22 with the conventional follow-up monitoring procedure for the first 6 months after an initial transurethral resection of a bladder tumor. Each routine cystoscopy and cytology costs a total of $155. For the 6 months following initial TURBT, the cost for standard follow up monitoring totaled $311, while the cost for the NMP22 monitoring modality was $257. On average, using NMP22 would have saved $55 per patient during the first 6 months of follow-up, resulting in a cost saving of approximately 18%. As well, 64.3% of patients would have undergone only one cystoscopy during the 6 month period, instead of the two done using the conventional modality. The trade-off for using NMP22 is that some patients would have a 3 month delay before diagnosis of a recurrence. This would occur in 8.9% of patients. NMP22 is less expensive than conventional monitoring follow-up of bladder cancer, and it can decrease patient discomfort by reducing the need for cystoscopy. Implementing it as routine follow-up monitoring should be considered, particularly for patients with low-grade tumors. PMID- 11119442 TI - Cerebrospinal fluid pseudocyst: a postoperative complication of augmentation ileocystoplasty in myelodysplastic children. AB - Peritoneal cerebrospinal fluid pseudocyst (CSFoma) formation is a rare postoperative complication of augmentation cystoplasty in children, with only three other cases being reported in the world literature. We describe two patients with CSFoma formation following augmentation ileocystoplasty. The management of these cases and a brief review of the pathogenesis and management of this condition are presented. PMID- 11119443 TI - Perinephric abscess presenting as chronic diarrhea. AB - Perinephric abscess is an uncommon diagnosis with a variable presentation and high mortality. We report an unusual case of a patient with a perinephric abscess who presented with chronic diarrhea and weight loss. PMID- 11119444 TI - Outcome of hypospadias repair using the tubularized, incised plate urethroplasty. AB - OBJECTIVE: We reviewed our results using the tubularized incised plate urethroplasty (Snodgrass procedure) for repair of penile hypospadias. MATERIALS AND METHODS: A total of 37 patients (aged 7-72 months, mean 17.5 months) underwent repair by three pediatric urologists at two institutions. The pre-op meatal position was distal in 28, mid-shaft in five, and penoscrotal in three patients. One patient, who did not have hypospadias, had a distal urethral fistula secondary to a previous circumcision. Twenty-six patients had ventral chordee and 12 required a dorsal tunica albuginea plication for correction. Urethroplasty was performed using 6-0 synthetic absorbable suture (PDS, Maxon, Dexon, or Monocryl). Urethroplasty coverage consisted of de-epithelialized dorsal preputial skin flap (32 patients), internal spermatic fascia flap (1 patient), tunica vaginalis flap (2 patients), or no coverage (2 patients). All patients were stented (8, 10 or 12 F silastic) for a mean duration of 9.8 days (range 4-12 days). Either a foam dressing (12 patients) or a Tegaderm sandwich dressing (25 patients) was used. RESULTS: Average length of hospital stay at one institution was 3.1 days (range 1-5 days). Mean follow-up was 8.8 months (range 1.5-20 months). The post-operative results were satisfactory with the meatus in a glanular position in 35 patients and a coronal position in two patients. All had a vertical orientation of the meatus. Complications included urethrocutaneous fistula in six patients, skin dehiscence in two patients, and meatal stenosis in two patients. One of the fistulas healed spontaneously. Urethral strictures have not been encountered thus far. CONCLUSIONS: The tubularized incised plate urethroplasty achieves satisfactory results with acceptable complications. It can be used for both distal and proximal hypospadias, and in the rare situation of fistula post-circumcision. Long term follow-up is needed to ensure that urethral strictures do not result from this technique. PMID- 11119445 TI - The Batista procedure. PMID- 11119446 TI - Blood pressure measurement is changing! PMID- 11119447 TI - Images in Cardiology: Aortic valve sparing operations. PMID- 11119448 TI - Managing out-of-hospital cardiac arrest survivors: 1. Neurological perspective. PMID- 11119449 TI - Should we give antibiotic prophylaxis against infective endocarditis in all cardiac patients, whatever the type of dental treatment? PMID- 11119450 TI - Images in Cardiology: "Late-late" reocclusion after coronary stenting and brachytherapy. PMID- 11119451 TI - Bacterial endocarditis following repeated tattooing. AB - Body decoration in the form of tattooing is becoming increasingly popular, especially among younger age groups. Although serious infections following tattooing are rare they are well documented. The first reported case of endocarditis caused by repeated tattooing in an individual with known valvar heart disease is presented. PMID- 11119452 TI - Images in Cardiology: Angiographic features of endomyocardial fibrosis. PMID- 11119453 TI - Vascular and valvar calcification: recent advances. PMID- 11119454 TI - Cognitive impairment in heart failure with Cheyne-Stokes respiration. AB - OBJECTIVES: To document the degree of cognitive impairment in stable heart failure, and to determine its relation to the presence of Cheyne-Stokes respiration during sleep. SUBJECTS: 104 heart failure patients and 21 healthy normal volunteers. METHODS: Overnight oximetry was used (previously validated as a screening tool for Cheyne-Stokes respiration in heart failure). Cognitive function was assessed using a battery of neuropsychological tests. Left ventricular function was assessed by echocardiography. RESULTS: Heart failure patients performed worse than the healthy volunteers in tests that measured vigilance. Reaction times were 48% slower (0.89 (0.03) s v 0.60 (0.05) s p < 0.005) and they hit twice as many obstacles on the Steer Clear simulator (75 (6.4) v 33 (4.6); p < 0.005). Cognitive impairment within the heart failure group was unrelated to either the presence of Cheyne-Stokes respiration, the degree of left ventricular dysfunction, or indices of nocturnal oxygenation. CONCLUSIONS: Vigilance was impaired in heart failure but this did not appear to be related to the presence of Cheyne-Stokes respiration during sleep. Impaired vigilance as measured on the Steer Clear test has been associated with an increased risk of motor vehicle accidents. The issue of fitness to drive in heart failure requires further attention. PMID- 11119455 TI - Images in cardiology: Complete angiographic view of the coronary-subclavian steal syndrome. PMID- 11119456 TI - Pulmonary venous flow velocity patterns in 404 individuals without cardiovascular disease. AB - OBJECTIVE: To determine the pulmonary venous flow velocity (PVFV) values in a large normal population. DESIGN: Prospective study in consecutive individuals. SETTING: University hospital. METHODS: Among 404 normal individuals, the flow velocity pattern in the right upper pulmonary vein was recorded in 315 subjects using transthoracic echocardiography, and in both upper pulmonary veins in 100 subjects using transoesophageal echocardiography. Subjects were divided into five age groups. The PVFV values were compared between transthoracic and transoesophageal echocardiography within the age groups, and intraindividually between the right and left upper pulmonary veins in transoesophageal echocardiography. RESULTS: Normal PVFV values for the right upper pulmonary vein in transthoracic and transoesophageal echocardiography are presented. The duration of flow reversal at atrial contraction was overestimated using transthoracic echocardiography (mean (SD): 96 (21) ms in transoesophageal echocardiography, 120 (28) ms in transthoracic echocardiography, p < 0.0001). Systolic to diastolic peak flow velocity ratio (S:D) increased earlier with advancing age with transoesophageal echocardiography than with transthoracic echocardiography. Similar results were found for the corresponding time-velocity integrals. Data from the left and right upper pulmonary veins differed with respect to onset and deceleration of flow velocities, but not for flow durations or peak velocities. CONCLUSIONS: Normal PVFV values generally show a wide range. The data presented will be of value in assessing left ventricular diastolic function and mitral regurgitation using the PVFV pattern. PMID- 11119457 TI - Assessment of left ventricular long axis contraction can detect early myocardial dysfunction in asymptomatic patients with severe aortic regurgitation. AB - OBJECTIVE: To identify variables that could be applied at rest to diagnose subclinical ventricular dysfunction in asymptomatic patients with severe aortic regurgitation. DESIGN: Cross sectional study. PATIENTS: Left ventricular long axis contraction was studied using tissue Doppler and M mode echocardiography in 21 patients with no symptoms (New York Heart Association (NYHA) functional class 40%). MAIN OUTCOME MEASURES: Left ventricular ejection fraction (LVEF) at baseline and peak exercise (Weber protocol), cardiopulmonary function, and left ventricular long axis function at rest (peak systolic velocity and excursion of the mitral annulus). RESULTS: In 11 patients, ejection fraction increased or did not change (from mean (SD) 55 (5)% to 58 (4)%, p < 0.05) (group I); in 10 patients it decreased by > 5% (from 54 (4)% to 42 (5)%, p < 0.001) (group II). Exercise ejection fraction was < 50% in all patients in group II. At rest, there were no differences between the groups in ejection fraction, left ventricular diameter indices, wall stress, and short axis contraction. However, patients in group II had reduced long axis contraction compared with group I: peak systolic velocity 8.6 (0.6) v 11.9 (2.2) cm/s (p < 0.001); excursion 11 (2) v 14 (2) mm (p < 0.01). A resting velocity of < 9.5 cm/s was the best indicator of poor exercise tolerance (sensitivity 90%, specificity 100%). CONCLUSIONS: Markers of reduced long axis contraction may provide simple and reliable indices of subclinical left ventricular dysfunction in asymptomatic patients with severe aortic regurgitation. PMID- 11119458 TI - Comparative accuracy of cardiovascular risk prediction methods in primary care patients. AB - OBJECTIVE: To compare the relative accuracy of cardiovascular disease risk prediction methods based on equations derived from the Framingham heart study. DESIGN: Risk factor data were collected prospectively from subjects being evaluated by their primary care physicians for prevention of cardiovascular disease. Projected cardiovascular risks were calculated for each patient with the Framingham equations, and also estimated from the risk tables and charts based on the same equations. SETTING: 12 primary care practices (46 doctors) in Birmingham. PATIENTS: 691 subjects aged 30-70 years. MAIN OUTCOME MEASURES: Sensitivity, specificity, and positive and negative predictive values of the Framingham based risk tables and charts for treatment thresholds based on projected cardiovascular disease or coronary heart disease risk. RESULTS: 59 subjects (8.5%) had projected 10 year coronary heart disease risks >/= 30%, and 291 (42.1%) had risks >/= 15%. At equivalent projected risk levels (10 year coronary heart disease >/= 30% and five year cardiovascular disease >/= 20%), the original Sheffield tables and those from New Zealand have the same sensitivities (40.0%, 95% confidence interval (CI) 26.6% to 57.8% v 41.2%, 95% CI 28.7% to 57. 3%) and specificities (98.6%, 95% CI 97.2% to 99.3% v 99.7%, 95% CI 98.8% to 100%). Modifications to the Sheffield tables improve sensitivity (91.4%, 95% CI 81.3% to 96.9%) but reduce specificity (95.8%, 95% CI 93.9% to 97.3%). The revised joint British recommendations' charts have high specificity (98.7%, 95% CI 97.5% to 99.5%) and good sensitivity (84.7%, 95% CI 71.0% to 93.0%). CONCLUSIONS: The revised joint British recommendations charts appear to have the best combination of sensitivity and specificity for use in primary care patients. PMID- 11119459 TI - Images in cardiology: Late thoracic and abdominal aortic aneurysm following discreet coarctation of the aorta repair. PMID- 11119460 TI - Slow pathway modification for atrioventricular node re-entrant tachycardia: fast junctional tachycardia predicts adverse prognosis. AB - OBJECTIVE: To examine the cycle length of the junctional tachycardia often seen during successful slow pathway ablation for atrioventricular (AV) node re-entrant tachycardia, to determine whether shorter cycle lengths predict imminent atrioventricular block. DESIGN: Retrospective analysis of consecutive patients undergoing slow pathway modification. Intracardiac recordings were analysed after digital storage to determine the development of junctional tachycardia, its duration and maximum, minimum, and mean cycle length, occurrence of heart block, persistent slow pathway conduction, or later confirmed recurrence of AV node re entrant tachycardia. SETTING: Regional cardiac centre. PATIENTS: 136 consecutive patients undergoing electrophysiological study found to have typical "slow-fast" AV node re-entrant tachycardia and subject to 137 slow pathway modification procedures. RESULTS: During successful temperature feedback controlled radiofrequency energy application, junctional tachycardia developed in 133 of 137 procedures. During ablation, 10 patients had evidence of AV block (first degree in seven patients and third degree in three), and 17 others had retrograde junctional atrial (JA) block. In these 27 patients, the junctional tachycardia was rapid, with a minimum (SD) cycle length 291 (47) ms. Conduction recovered quickly in all but two patients, one of whom required permanent pacing. Junctional tachycardia with normal AV and JA conduction in the other 111 patients was of a significantly slower minimum cycle length (537 (123) ms; p < 0.0001). CONCLUSIONS: Fast junctional tachycardia with cycle lengths under 350 ms seen during slow pathway modification is a predictor of conduction block, suggesting proximity to the compact node. Radiofrequency energy application should be terminated immediately to prevent development of AV block. An "auto cut off" facility for cycle lengths shorter than 350 ms could be built into radiofrequency ablation systems to increase safety. PMID- 11119461 TI - No incremental benefit of multisite atrial pacing compared with right atrial pacing in patients with drug refractory paroxysmal atrial fibrillation. AB - OBJECTIVE: To evaluate the incremental antifibrillatory effect of multisite atrial pacing compared with right atrial pacing in patients with drug refractory paroxysmal atrial fibrillation paced for arrhythmia prevention alone. METHODS: In 20 of these patients (mean (SD) age 64 (8) years; 14 female, six male), a single blinded randomised crossover study was performed to investigate the incremental benefit of one month of multisite atrial pacing compared with one month of right atrial pacing. Outcomes included the number of episodes of paroxysmal atrial fibrillation, their total duration obtained from pacemaker Holter memory, and quality of life using a cardiac specific questionnaire (the modified Karolinska questionnaire). RESULTS: Comparing right atrial with multisite atrial pacing, there was no significant change in either the number of paroxysmal atrial fibrillation episodes (mean (SD): right atrial pacing 77 (98) episodes v multisite pacing 52 (78) episodes, NS) or their total duration (right atrial, 4.8 (5.4) days v multisite, 6.3 (9.8) days, NS). Quality of life scores compared with baseline status were equally improved by either pacing strategy (mean percentage improvement: right atrial, 38%, p = 0.003; multisite, 44%, p = 0.003). There was no significant difference in life scores comparing the two pacing modes. CONCLUSIONS: Multisite atrial pacing has no incremental antiarrhythmic effect compared with right atrial pacing in patients paced for drug refractory paroxysmal atrial fibrillation. Quality of life is equally improved with either pacing strategy, with no differences between them. PMID- 11119462 TI - Images in cardiology: Cardiac arrest caused by spontaneous left coronary artery dissection in a young woman. PMID- 11119463 TI - Long term results of cardioverter-defibrillator implantation in patients with right ventricular dysplasia and malignant ventricular tachyarrhythmias. AB - OBJECTIVE: To study the outcome of patients with arrhythmogenic right ventricular dysplasia treated with an implantable cardioverter-defibrillator (ICD) for ventricular tachyarrhythmias complicated by haemodynamic collapse. DESIGN: Observational study. SETTING: University hospital. PATIENTS: Nine consecutive patients (eight male, one female; mean (SD) age, 36 (18) years) with arrhythmogenic right ventricular dysplasia presenting with ventricular tachycardia and haemodynamic collapse (n = 6) or ventricular fibrillation (n = 3), treated with an ICD. MAIN OUTCOME MEASURES: Survival; numbers of and reasons for appropriate and inappropriate ICD interventions. RESULTS: After a mean (SD) follow up of 32 (24) months, all patients were alive. Six patients received a median of 19 (range 2-306) appropriate ICD interventions for events detected in the ventricular tachycardia window; four received a median of 2 (range 1-19) appropriate ICD interventions for events detected in the ventricular fibrillation window. Inappropriate interventions were seen for sinus tachycardia (18 episodes in three patients), atrial fibrillation (three episodes in one patient), and for non-sustained polymorphic ventricular tachycardia (one episode in one patient). CONCLUSIONS: Patients with arrhythmogenic right ventricular dysplasia and malignant ventricular arrhythmias have a high recurrence rate requiring appropriate ICD interventions, but they also often have inappropriate interventions. Programming the device is difficult because this population develops supraventricular and ventricular tachyarrhythmias with similar rates. PMID- 11119464 TI - Echocardiographic and signal averaged ECG indices associated with non-sustained ventricular tachycardia after repair of tetralogy of fallot. AB - OBJECTIVE: To identify any possible association between different readily available non-invasive indices and potential malignant ventricular arrhythmias in patients with repaired tetralogy of Fallot. DESIGN: 27 consecutive patients, mean (SD) age 27.3 (11.7) years, were studied 15.7 (6.7) years after corrective surgery for tetralogy of Fallot, using 12 lead ECG, 24 hour Holter recordings, signal averaged ECG, and echocardiography. The following variables were measured: standard QRS duration, filtered QRS duration (fltQRS), low amplitude signal duration, and root mean square voltage of the last 40 ms of the fltQRS (RMS-40), as well as right ventricular systolic pressure, right ventricular ejection fraction, and the ratio of the maximum short axis diameters of the right and left ventricles (RD:LD). RESULTS: All patients had right bundle branch block, with a mean QRS duration of 137.1 (14.9) ms. There were no patients with sustained arrhythmia. Five patients had runs of non-sustained ventricular tachycardia (group A) and the other 22 patients did not (group B). Univariate analysis showed that fltQRS and RD:LD ratio were significantly associated with non-sustained ventricular tachycardia. In addition, a fltQRS >/= 148 ms, low amplitude signal >/= 32.5 ms, RMS-40 /= 1.05 were cut off points with a high sensitivity for detecting patients with non-sustained ventricular tachycardia. CONCLUSIONS: Abnormal signal averaged ECG and echocardiographic variables are associated with potentially malignant ventricular arrhythmias on the Holter recordings in asymptomatic patients with repaired tetralogy of Fallot. PMID- 11119465 TI - Increased platelet reactivity and significant changes in coagulation markers after cavopulmonary connection. AB - OBJECTIVE: To evaluate platelet reactivity and coagulation markers after surgical palliation of univentricular hearts. DESIGN AND PATIENTS: Cross sectional survey of 24 patients, median age 11 (range 4-22) years, at 2 (range 0.5-6) years after a total cavopulmonary connection (TCPC; n = 14) or a bidirectional Glenn anastomosis (Glenn; n = 10). MAIN OUTCOME MEASURES: Platelet reactivity and/or coagulation markers were measured in 20 patients (four excluded because of anticoagulant treatment) and compared with 33 healthy controls, median age 12 (range 6-16) years. RESULTS: None of the patients had clinically apparent thromboembolic events. However, increased platelet reactivity was observed ex vivo both after collagen induced platelet aggregation (median 73% (interquartile range 61-84%) in patients, and 61% (47-69%) in controls; p < 0.01), and after ADP induced platelet aggregation (69% (53-77%) in patients, and 56% (40-66%) in controls; p < 0.05). Concentrations of protein S antigen, antithrombin III, and protein C activity were reduced after both TCPC and Glenn. A concomitant decrease was seen in coagulation factor II, VII, X, and factor VII clot activity. CONCLUSIONS: Several abnormalities in the coagulation system were observed after bidirectional Glenn anastomosis, similar to alterations previously described in Fontan operated and TCPC patients. Antithrombotic treatment in these patients is still an unresolved issue, but aspirin is often recommended. This study shows that such a strategy is rational and the results suggest that antiplatelet treatment may be advantageous, either alone or in combination with oral anticoagulant treatment. PMID- 11119466 TI - Dental procedures in children with severe congenital heart disease: a theoretical analysis of prophylaxis and non-prophylaxis procedures. AB - OBJECTIVE: To estimate the cumulative exposure to bacteraemia from dental procedures currently recommended for antibiotic prophylaxis and compare this with cumulative exposure from dental procedures not recommended for prophylaxis. DESIGN: Retrospective analysis. SETTING: University and teaching hospital maxillofacial and dental department. PATIENTS: 136 children with severe congenital cardiac disease attending for dental treatment between 1993 and 1998 and for whom full records were available. Each dental procedure was tallied. MAIN OUTCOME MEASURES: Cumulative exposure per annum to "non-prophylaxis procedures"; cumulative exposure per annum to "prophylaxis procedures". RESULTS: Cumulative exposure to bacteraemia from prophylaxis procedures was not significantly greater than from non-prophylaxis procedures. CONCLUSIONS: The data raise important questions about the appropriateness of current guidelines for antibiotic prophylaxis of bacterial endocarditis. PMID- 11119467 TI - Development and validation of a Bayesian index for predicting major adverse cardiac events with percutaneous transluminal coronary angioplasty. AB - OBJECTIVE: To create a risk model for predicting major adverse complicating events of percutaneous transluminal coronary angioplasty (PTCA), and to test the accuracy of the model on a prospective cohort of patients SETTING: Tertiary cardiac centre METHODS: Available software can predict probabilities of events using Bayes's theorem. To establish the accuracy of these predictive tools, a Bayes table was created to evaluate major adverse complicating events (MACE) death, emergency coronary artery bypass grafting (CABG), or Q wave infarct occurring during the in-patient episode-on the first 1500 patients in the department PTCA database (development group); the predictive value of this model was then tested with the subsequent 1000 patients (evaluation group). The following probabilities were assessed to determine their association with MACE: age, sex, left ventricular function, American Heart Association lesion morphology classification, cardiogenic shock, previous CABG, diabetes, hypertension, multivessel PTCA. MAIN OUTCOME MEASURES: To establish the discriminatory ability of the predictive index, calibration plots and receiver operating characteristic (ROC) curves were obtained to compare the development and evaluation groups. RESULTS: The ROC curve plotted to determine the discriminatory value of the Bayesian table created from the development group (n = 1500) in predicting MACE in the evaluation group (n = 1000) showed a moderately predictive area under the curve of 0.76 (SEM 0.07). This predictive accuracy was confirmed with separately constructed calibration plots. CONCLUSIONS: Accurate predictions of MACE can be identified in populations undergoing percutaneous intervention. The database used allows operators to obtain consent from patients appropriately from their own experience rather than from other published data. If a national PTCA database existed along similar lines, individual operators and interventional centres could compare themselves with nationally available data. PMID- 11119469 TI - Images in Cardiology: Serial follow up of intramural haematoma associated with lumen compromise after intracoronary intervention. PMID- 11119468 TI - Three dimensional intravascular ultrasonic assessment of the local mechanism of restenosis after balloon angioplasty. AB - OBJECTIVE: To assess the mechanism of restenosis after balloon angioplasty. DESIGN: Prospective study. PATIENTS: 13 patients treated with balloon angioplasty. INTERVENTIONS: 111 coronary subsegments (2 mm each) were analysed after balloon angioplasty and at a six month follow up using three dimensional intravascular ultrasound (IVUS). MAIN OUTCOME MEASURES: Qualitative and quantitative IVUS analysis. Total vessel (external elastic membrane), plaque, and lumen volume were measured in each 2 mm subsegment. Delta values were calculated (follow up - postprocedure). Remodelling was defined as any (positive or negative) change in total vessel volume. RESULTS: Positive remodelling was observed in 52 subsegments while negative remodelling occurred in 44. Remodelling, plaque type, and dissection were heterogeneously distributed along the coronary segments. Plaque composition was not associated with changes in IVUS indices, whereas dissected subsegments had a greater increase in total vessel volume than those without dissection (1.7 mm(3) v -0.33 mm(3), p = 0.04). Change in total vessel volume was correlated with changes in lumen (p < 0.05, r = 0.56) and plaque volumes (p < 0.05, r = 0.64). The site with maximum lumen loss was not the same site as the minimum lumen area at follow up in the majority (n = 10) of the vessels. In the multivariate model, residual plaque burden had an influence on negative remodelling (p = 0.001, 95% confidence interval (CI) -0.391 to 0.108), whereas dissection had an effect on total vessel increase (p = 0.002, 95% CI 1.168 to 4.969). CONCLUSIONS: The mechanism of lumen renarrowing after balloon angioplasty appears to be determined by unfavourable remodelling. However, different patterns of remodelling may occur in individual injured coronary segments, which highlights the complexity and influence of local factors in the restenotic process. PMID- 11119470 TI - The influence of general health status and social support on symptomatic outcome following coronary artery bypass grafting. AB - OBJECTIVES: To assess health status, level of social support, and presence of coronary artery disease risk factors before and after coronary artery bypass grafting (CABG); to assess symptomatic relief approximately 12 months postoperatively; and to examine the association between preoperative health status and recurrence of symptoms. DESIGN: Observational study. SETTING: Preoperatively, in hospital outpatient department (1995-1996); postoperatively, at home (1996-97). SUBJECTS AND METHODS: Patients awaiting elective CABG were recruited one month before the expected date of operation. Preoperative assessment included severity of symptoms, coronary artery disease risk factors, short form 36 (SF-36) questionnaire, and social activities questionnaire. The presence and severity of angina and breathlessness were reported postoperatively (mean 16.4 months). Multiple regression analysis was used to identify factors associated with improved outcome following CABG. MAIN OUTCOME MEASURE: Patient reported presence and severity of angina and breathlessness. RESULTS: 183 patients were followed for a mean of 16. 4 months after CABG. Angina and breathlessness were completely relieved in 55% and 36% of patients, respectively. In patients with residual symptoms, the severity was significantly reduced (angina p < 0.001; breathlessness, p = 0.02). Patients with low SF-36 scores and low social network scores preoperatively were less likely to be relieved of symptoms (p < 0.001). Health status and social support levels preoperatively were lower than in other reported coronary artery disease patients groups. Preoperatively, coronary artery disease risk factors were higher than recommended in current guidelines: 67.4% had raised plasma cholesterol, 39.0% were hypertensive, 80% were moderately obese, and 22.9% were smokers. CONCLUSIONS: Recurrence of symptoms exceeded other published studies. Patients' perception of general health, symptoms, and social support influences outcome. PMID- 11119471 TI - Images in Cardiology: Asymptomatic right atrial myxoma in acromegalic man: a case of Carney complex. PMID- 11119472 TI - Increased serum concentrations of advanced glycation end products: a marker of coronary artery disease activity in type 2 diabetic patients. AB - OBJECTIVE: To assess whether the concentrations of serum advanced glycation end products (AGE) in diabetic patients with obstructive coronary artery disease differ from those in type 2 diabetic patients without obstructive coronary artery disease. DESIGN: Serum AGE concentrations were measured in type 2 diabetic patients and in non-diabetic patients, both with and without obstructive coronary artery disease, and the relation between these values and coronary disease severity was evaluated. RESULTS: Mean (SD) serum AGE concentrations were higher (p < 0.0125) in type 2 diabetic patients with obstructive coronary artery disease (5.5 (2.5) mU/ml, n = 30) than in patients without obstructive coronary artery disease (2.8 (0. 5) mU/ml, n = 12), and higher than in non-diabetic patients with (3. 4 (1.0) mU/ml, n = 28) and without (3.2 (0.4) mU/ml, n = 13) obstructive coronary artery disease. Serum AGE was associated with the degree of coronary arteriosclerosis in type 2 diabetic patients with obstructive coronary artery disease (single vessel: n = 13, 3.4 (0.9) mU/m; two vessel: n = 6, 5.7 (1.6) mU/m; three vessel: n = 11, 7.2 (2.5) mU/ml). Serum AGE was positively correlated with serum mean four year HbA(1C) (r = 0.46, p < 0.01), but not with recent serum HbA(1C) (r = 0.24). The four groups did not differ in the other coronary risk factors. CONCLUSIONS: Serum AGE concentrations may be associated with long term poor glycaemic control and reflect the severity of coronary arteriosclerosis in type 2 diabetic patients. PMID- 11119473 TI - Images in Cardiology: Left ventricular pacing via the great cardiac vein in a patient with tricuspid and pulmonary valve replacement. PMID- 11119474 TI - Effect of a high loading dose of clopidogrel on platelet function in patients undergoing coronary stent placement. PMID- 11119475 TI - Management of severe heart failure by specialist palliative care. PMID- 11119476 TI - Heart Failure: Treatment strategies for heart failure: beta blockers andantiarrhythmics. PMID- 11119477 TI - Coronary Disease: Acute myocardial infarction: failed thrombolysis. PMID- 11119478 TI - Haemodynamic calculations in the catheter laboratory. PMID- 11119479 TI - Left main coronary artery aneurysm with chronic total occlusion of both left coronary arteries in a young athlete. AB - Aneurysms of the left main coronary arteries are found in 0.1% of angiograms. This case involves an athlete with a left main coronary artery aneurysm, which was combined with chronic total occlusion of the proximal left anterior descending and proximal left circumflex coronary arteries. The extraordinary clinical presentation in this patient may be associated with good coronary collaterals, which may have developed in the patient in response to chronic total occlusion of the coronary artery by the aneurysm, and repeat myocardial hypoxia during high levels of performance as a soccer player. PMID- 11119480 TI - Varicella myocarditis in an adult. AB - A 24 year old male with varicella myocarditis was admitted with chest pain and fever up to 39 degrees C. The ECG showed J point and ST elevation in leads V2-V4, and inverted T waves in leads V5 and V6. Creatine kinase (CK) was raised to 435 U/l (CK-MB 36 U/l), troponin I was 63.4 microgram/l, and lactate dehydrogenase was 359 U/l, suggesting cardiac involvement of varicella infection. The left ventricle was dilated (58 mm) and left ventricular ejection fraction was globally reduced (ejection fraction 45%). Myocarditis was confirmed by endomyocardial biopsy. The patient was treated with specific varicella hyperimmunoglobulins, aciclovir, and a non-steroidal anti-inflammatory drug. During two months follow up the patient recovered completely. This case report is a reminder that a varicella infection can cause myocarditis in adults. Early diagnosis and appropriate treatment of this rare form of myocarditis may lead to complete recovery. PMID- 11119481 TI - ST segment elevation in the right precordial leads following administration of class Ic antiarrhythmic drugs. AB - Electrocardiographic changes were evaluated retrospectively in five patients without previous episodes of syncope or ventricular fibrillation who developed abnormal ST segment elevation mimicking the Brugada syndrome in leads V1-V3 after the administration of class Ic antiarrhythmic drugs. Pilsicainide (four patients) or flecainide (one patient) were administered orally for the treatment of symptomatic paroxysmal atrial fibrillation or premature atrial contractions. The QRS duration, QTc, and JT intervals on 12 lead surface ECG before administration of these drugs were all within normal range. After administration of the drugs, coved-type ST segment elevation in the right precordial leads was observed with mild QRS prolongation, but there were no apparent changes in JT intervals. No serious arrhythmias were observed during the follow up periods. Since ST segment elevation with mild QRS prolongation was observed with both pilsicainide and flecainide, strong sodium channel blocking effects in the depolarisation may have been the main factors responsible for the ECG changes. As the relation between ST segment elevation and the incidence of serious arrhythmias has not yet been sufficiently clarified, electrocardiographic changes should be closely monitored whenever class Ic drugs are given. PMID- 11119483 TI - Mutator natural Escherichia coli isolates have an unusual virulence phenotype. AB - A small percentage of natural Escherichia coli isolates (both commensal and pathogenic) have a mutator phenotype related to defects in methyl-directed mismatch repair (MR) genes. We investigated whether there was a direct link between the mutator phenotype and virulence by (i) studying the relationships between mutation rate and virulence in a mouse model of extraintestinal virulence for 88 commensal and extraintestinal pathogenic E. coli isolates and (ii) comparing the virulence in mice of MR-deficient and MR-proficient strains that were otherwise isogenic. The results provide no support for the hypothesis that the mutator phenotype has a direct role in virulence or is associated with increased virulence. Most of the natural mutator strains studied displayed an unusual virulence phenotype with (i) a lack of correspondence between the number of virulence determinants and pathogenicity in mice and (ii) an intermediate level of virulence. On a large evolutionary scale, the mutator phenotype may help parasites to achieve an intermediate rate of virulence which mathematical models predict to be selected for during long-term parasite-host interactions. PMID- 11119482 TI - Ups and downs of mucosal cellular immunity against protozoan parasites. PMID- 11119484 TI - Shigella flexneri LuxS quorum-sensing system modulates virB expression but is not essential for virulence. AB - Quorum-sensing systems regulate the expression of virulence factors in a wide variety of plant and animal pathogens, including members of the Enterobacteriaceae. Studies of Shigella virulence gene expression have demonstrated that maximal expression of genes encoding the type III secretion system and its substrates and maximal activity of this virulence organelle occur at high cell density. In these studies, we demonstrate that the expression of ipa, mxi, and spa invasion operons is maximal in stationary-phase bacteria and that conditioned media derived from stationary-phase cultures enhance the expression of these loci. In contrast, expression of virB, a transcription factor essential for the expression of invasion loci, peaks in late log phase; accordingly, virB expression is enhanced by a signal(s) present in conditioned media derived from late-log-phase cultures. Autoinducer 2 (AI-2), a quorum signaling molecule active in late log phase, was synthesized by Shigella species and enteroinvasive Escherichia coli and shown to be responsible for the observed peak of virB expression. However, AI-2 does not influence invasion operon expression and is not required for Shigella virulence, as mutants deficient in AI 2 synthesis are fully virulent. The implications of these findings with regard to both virB and invasion operon expression and the evolution of circuitries governing virulence gene expression are discussed. PMID- 11119485 TI - Surfactant protein D enhances phagocytosis and killing of unencapsulated phase variants of Klebsiella pneumoniae. AB - Pulmonary surfactant protein D (SP-D) is a collagenous C-type lectin (collectin) that is secreted into the alveoli and distal airways of the lung. We have studied the interactions of SP-D and alveolar macrophages with Klebsiella pneumoniae, a common cause of nosocomial pneumonia. SP-D does not agglutinate encapsulated K. pneumoniae but selectively agglutinates spontaneous, unencapsulated phase variants, such as Klebsiella strain K50-3OF, through interactions with their lipopolysaccharides (LPS). These effects are calcium dependent and inhibited with maltose but not lactose, consistent with involvement of the SP-D carbohydrate recognition domain. Precoating of K50-3OF with SP-D enhances the phagocytosis and killing of these organisms by rat alveolar macrophages in cell culture and stimulates the production of nitric oxide by the NR-8383 rat alveolar macrophage cell line. SP-D similarly enhances the NO response to K50-3OF LPS adsorbed to Latex beads under conditions where soluble LPS or SP-D, or soluble complexes of SP-D and LPS, do not stimulate NO production. Our studies demonstrate that interactions of SP-D with exposed arrays of Klebsiella LPS on a particulate surface can enhance the host defense activities of alveolar macrophages and suggest that activation of macrophages by SP-D requires binding to microorganisms or other particulate ligands. Because unencapsulated phase variants are likely to be responsible for the initial stages of tissue invasion and infection, we speculate that SP-D-mediated agglutination and/or opsonization of K. pneumoniae is an important defense mechanism for this respiratory pathogen in otherwise healthy individuals. PMID- 11119486 TI - Detection of bacterial virulence genes by subtractive hybridization: identification of capsular polysaccharide of Burkholderia pseudomallei as a major virulence determinant. AB - Burkholderia pseudomallei, the etiologic agent of melioidosis, is responsible for a broad spectrum of illnesses in humans and animals particularly in Southeast Asia and northern Australia, where it is endemic. Burkholderia thailandensis is a nonpathogenic environmental organism closely related to B. pseudomallei. Subtractive hybridization was carried out between these two species to identify genes encoding virulence determinants in B. pseudomallei. Screening of the subtraction library revealed A-T-rich DNA sequences unique to B. pseudomallei, suggesting they may have been acquired by horizontal transfer. One of the subtraction clones, pDD1015, encoded a protein with homology to a glycosyltransferase from Pseudomonas aeruginosa. This gene was insertionally inactivated in wild-type B. pseudomallei to create SR1015. It was determined by enzyme-linked immunosorbent assay and immunoelectron microscopy that the inactivated gene was involved in the production of a major surface polysaccharide. The 50% lethal dose (LD(50)) for wild-type B. pseudomallei is <10 CFU; the LD(50) for SR1015 was determined to be 3.5 x 10(5) CFU, similar to that of B. thailandensis (6.8 x 10(5) CFU). DNA sequencing of the region flanking the glycosyltransferase gene revealed open reading frames similar to capsular polysaccharide genes in Haemophilus influenzae, Escherichia coli, and Neisseria meningitidis. In addition, DNA from Burkholderia mallei and Burkholderia stabilis hybridized to a glycosyltransferase fragment probe, and a capsular structure was identified on the surface of B. stabilis via immunoelectron microscopy. Thus, the combination of PCR-based subtractive hybridization, insertional inactivation, and animal virulence studies has facilitated the identification of an important virulence determinant in B. pseudomallei. PMID- 11119487 TI - Staphylococcus aureus agr genotypes with enterotoxin production capabilities can resist neutrophil bactericidal activity. AB - Staphylococcus aureus pathogenicity is mainly due to the production of a number of secreted and cell surface-associated proteins under the regulation of the agr gene. A region of the agr gene was used to subgroup S. aureus strains according to restriction fragment length polymorphisms. Additionally, strains were subtyped according to the coagulase gene in order to strengthen discriminatory power. Virulence capabilities of agr genotype subgroups were evaluated using an in vitro neutrophil bactericidal assay, which showed that prevalent genotypes were significantly better at evading this primary host defense. Multiplex PCR was then used to detect enterotoxin genes among the genotype subgroups in order to determine possible virulence candidates that enable strains to combat neutrophil killing. The prevalent genotype strains were found to possess higher production capabilities for enterotoxin A than did low-prevalence strains. The significance of enterotoxin A production capabilities in affecting pathogenicity of S. aureus strains was evaluated and found to have a profound effect on neutrophil killing abilities. The use of a large epidemiological database as a tool for subgrouping strains with varying degrees of pathogenicity has allowed the identification of relevant and previously undefined virulence factors that affect a pathogen's capability to overcome host immune defenses. PMID- 11119488 TI - Enterotoxigenic Escherichia coli TibA glycoprotein adheres to human intestine epithelial cells. AB - Enterotoxigenic Escherichia coli (ETEC) is capable of invading epithelial cell lines derived from the human ileum and colon. Two separate invasion loci (tia and tib) that direct noninvasive E. coli strains to adhere to and invade cultured human intestine epithelial cells have previously been isolated from the classical ETEC strain H10407. The tib locus directs the synthesis of TibA, a 104-kDa outer membrane glycoprotein. Synthesis of TibA is directly correlated with the adherence and invasion phenotypes of the tib locus, suggesting that this protein is an adhesin and invasin. Here we report the purification of TibA and characterization of its biological activity. TibA was purified by continuous elution preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Purified TibA was biotin labeled and then shown to bind to HCT8 human ileocecal epithelial cells in a specific and saturable manner. Unlabeled TibA competed with biotin-labeled TibA, suggesting the presence of a specific TibA receptor in HCT8 cells. These results show that TibA acts as an adhesin. Polyclonal anti-TibA antiserum inhibited invasion of ETEC strain H10407 and of recombinant E. coli bearing tib locus clones, suggesting that TibA also acts as an invasin. The ability of TibA to direct epithelial cell adhesion suggests a role for this protein in ETEC pathogenesis. PMID- 11119489 TI - Characterization of a streptococcal endopeptidase with homology to human endothelin-converting enzyme. AB - A gene encoding an endopeptidase from Streptococcus parasanguis FW213 has been cloned and shown to have high sequence homology to genes encoding mammalian metalloendopeptidases. The gene, designated S. parasanguis pepO, was cloned into the pET28a expression vector, resulting in a fusion of vector sequences encoding a hexahistidine tag at the carboxyl terminus. The recombinant PepO (rPepO) was expressed in Escherichia coli and purified using an Ni(2+) affinity column. Polyclonal antiserum to rPepO was raised in rabbits and used to localize FW213 PepO to the cytosol. Southern hybridization and immunoblot analysis revealed that other oral streptococci contain regions of DNA with homology to pepO and produce a protein with antigenic properties similar to that of FW213 PepO. Enzymatic activity assays indicated that only S. parasanguis species possess the ability to cleave metenkephalin, the natural substrate of the human neutral endopeptidase (NEP). Inhibition assays revealed that S. parasanguis PepO is a member of the M13 category of metalloendopeptidases, which includes NEP and endothelin-converting enzyme 1 (ECE-1), an enzyme involved in the maintenance of vascular tone. Thiorphan and phosphoramidon, two specific inhibitors of this category of endopeptidases, were used to determine that S. parasanguis PepO is more similar to ECE-1 than to NEP. PMID- 11119490 TI - Role of flm locus in mesophilic Aeromonas species adherence. AB - The adherence mechanism of Aeromonas caviae Sch3N to HEp-2 cells was initially investigated through four mini-Tn5 mutants that showed a 10-fold decrease in adherence. These mutants lost motility, flagella, and their lipopolysaccharide (LPS) O antigen (O-Ag). Three genes, flmB-neuA-flmD, were found to be interrupted by the transposon insertions; additionally, two other genes, one lying upstream (flmA) and one downstream (neuB), were found to be clustered in the same operon. While the flmA and flmB genes were present in all mesophilic Aeromonas spp. (A. hydrophila, A. caviae, A. veronii bv. veronii, and A. veronii bv. sobria) tested, this was not the case for the neuA-flmD-neuB genes. Construction and characterization of flmB insertion mutants in five other mesophilic Aeromonas strains revealed the loss of motility, flagella, and adherence but did not alter the LPS composition of these strains. Taking the above findings into consideration, we conclude (i) that flagella and possibly the LPS O-Ag are involved in the adherence of the mesophilic Aeromonas to human epithelial cells; (ii) flmA and flmB are genes widely distributed in the mesophilic Aeromonas and are involved in flagella assembly, and thus adherence; and (iii) in A. caviae Sch3N the flmA and flmB genes are found in a putative operon together with neuA, flmD, and neuB and are involved in LPS O-Ag biosynthesis and probably have a role in flagellum assembly. PMID- 11119491 TI - Inactivation of the srtA gene in Streptococcus gordonii inhibits cell wall anchoring of surface proteins and decreases in vitro and in vivo adhesion. AB - The srtA gene product, SrtA, has been shown to be required for cell wall anchoring of protein A as well as virulence in the pathogenic bacterium Staphylococcus aureus. There are five major mechanisms for displaying proteins at the surface of gram-positive bacteria (P. Cossart and R. Jonquieres, Proc. Natl. Acad. Sci. USA 97:5013-5015, 2000). However, since many of the known surface proteins of gram-positive bacteria are believed to be exported and anchored via the sortase pathway, it was of interest to determine if srtA plays a similar role in other gram-positive bacteria. To that end, the srtA gene in the human oral commensal organism Streptococcus gordonii was insertionally inactivated. The srtA mutant S. gordonii exhibited a marked reduction in quantity of a specific anchored surface protein. Furthermore, the srtA mutant had reduced binding to immobilized human fibronectin and had a decreased ability to colonize the oral mucosa of mice. Taken together, these results suggest that the activity of SrtA plays an important role in the biology of nonpathogenic as well as pathogenic gram-positive cocci. PMID- 11119492 TI - Comparison of CXC chemokines ENA-78 and interleukin-8 expression in Helicobacter pylori-associated gastritis. AB - Colonization of the gastric mucosa with Helicobacter pylori is associated with a dense infiltration of granulocytes into the lamina propria in the active phase of gastritis. In this study, we investigated the involvement of epithelial cell derived neutrophil-activating protein 78 (ENA-78) in development of H. pylori associated gastritis. Antral biopsies from 27 patients with H. pylori-associated gastritis and 25 from H. pylori-negative individuals were first analyzed for ENA 78 and interleukin-8 (IL-8) mRNA by semiquantitative reverse transcription (RT) PCR. In H. pylori-positive patients, significantly elevated levels were found for both chemokines (P<0.05). Only IL-8 mRNA levels differed significantly (P<0.05) in H. pylori-infected individuals who had serum antibodies for cytotoxin associated protein CagA versus H. pylori-infected CagA-negative persons. Quantification of ENA-78 transcript levels by competitive RT-PCR yielded a significant 45-fold upregulation for ENA-78 transcripts in biopsies of H. pylori positive versus H. pylori-negative patients (P<0.05). In contrast to earlier findings with IL-8, the degree of ENA-78 mRNA upregulation was independent of the grade of activity of gastritis. Immunofluorescence studies on tissues of antral biopsies localized ENA-78 protein expression mainly to the gastric epithelium of H. pylori-positive patients, while control tissues were negative. Upregulation of ENA-78 and IL-8 mRNA and protein expression was also observed in an in vitro system using a gastric adenocarcinoma cell line. Only viable H. pylori yielded a strong ENA-78 and IL-8 induction, while H. pylori outer membrane proteins or water-soluble proteins had no significant effect. These data provide evidence for the importance of both IL-8 and ENA-78 in the development and perpetuation of H. pylori-associated gastritis. PMID- 11119493 TI - In vitro responsiveness of gammadelta T cells from Mycobacterium bovis-infected cattle to mycobacterial antigens: predominant involvement of WC1(+) cells. AB - It is generally accepted that protective immunity against tuberculosis is generated through the cell-mediated immune (CMI) system, and a greater understanding of such responses is required if better vaccines and diagnostic tests are to be developed. gammadelta T cells form a major proportion of the peripheral blood mononuclear cells (PBMC) in the ruminant system and, considering data from other species, may have a significant role in CMI responses in bovine tuberculosis. This study compared the in vitro responses of alphabeta and gammadelta T cells from Mycobacterium bovis-infected and uninfected cattle. The results showed that, following 24 h of culture of PBMC with M. bovis-derived antigens, the majority of gammadelta T cells from infected animals became highly activated (upregulation of interleukin-2R), while a lower proportion of the alphabeta T-cell population showed activation. Similar responses were evident to a lesser degree in uninfected animals. Study of the kinetics of this response showed that gammadelta T cells remained significantly activated for at least 7 days in culture, while activation of alphabeta T cells declined during that period. Subsequent analysis revealed that the majority of activated gammadelta T cells expressed WC1, a 215-kDa surface molecule which is not expressed on human or murine gammadelta T cells. Furthermore, in comparison with what was found for CD4(+) T cells, M. bovis antigen was found to induce strong cellular proliferation but relatively little gamma interferon release by purified WC1(+) gammadelta T cells. Overall, while the role of these cells in protective immunity remains unclear, their highly activated status in response to M. bovis suggests an important role in antimycobacterial immunity, and the ability of gammadelta T cells to influence other immune cell functions remains to be elucidated, particularly in relation to CMI-based diagnostic tests. PMID- 11119494 TI - Time between inoculations and karyotype forms of Pneumocystis carinii f. sp. Carinii influence outcome of experimental coinfections in rats. AB - The prevalence of Pneumocystis carinii pneumonia (PCP) in humans caused by more than a single genotype has been reported to range from 10 to 67%, depending on the method used for detection (3, 19). Most coinfections were associated with primary rather than recurrent disease. To better understand the factors influencing the development of coinfections, the time periods between inoculations and the genotype of the infecting organisms were evaluated in the chronically immunosuppressed-inoculated rat model of PCP. P. carinii f. sp. carinii infecting rats differentiated by karyotypic profiles exhibit the same low level of genetic divergence manifested by organisms infecting humans. P. carinii f. sp. carinii karyotype forms 1, 2, and 6 were inoculated into immunosuppressed rats, individually and in dual combinations, spaced 0, 10, and 20 days apart. Infections comprised of both organism forms resulted from admixtures inoculated at the same time. In contrast, coinfections did not develop in most rats, where a 10- or 20-day gap was inserted between inoculations; only the first organism form inoculated was detected by pulsed-field gel electrophoresis in the resultant infection. Organism burdens were reduced with combinations of forms 1 and 2 spaced 20 days apart but not in rats inoculated with forms 1 and 6. A role for the host response in the elimination of the second population and in reduction of the organism burden was suggested by the lack of direct killing of forms 1 and 2 in an in vitro ATP assay, by reduction of the burden by autoclaved organisms, and by the specific reactions of forms 1 and 2 but not forms 1 and 6. These studies showed that the time between inoculations was critical in establishing coinfections and P. carinii f. sp. carinii karyotype profiles were associated with differences in biological responses. This model provides a useful method for the study of P. carinii coinfections and their transmission in humans. PMID- 11119495 TI - Single-copy IMH3 allele is sufficient to confer resistance to mycophenolic acid in Candida albicans and to mediate transformation of clinical Candida species. AB - Parasexual genetic analysis of Candida albicans utilized the dominant selectable marker that conferred resistance to mycophenolic acid. We cloned and sequenced the IMH3(r) gene from C. albicans strain 1006, which was previously identified as resistant to mycophenolic acid (MPA) (A. K. Goshorn and S. Scherer, Genetics 123:213-218, 1989). MPA is an inhibitor of IMP dehydrogenase, an enzyme necessary for the de novo biosynthesis of GMP. G. A. Kohler et al. (J. Bacteriol. 179:2331 2338, 1997) have shown that the wild-type IMH3 gene, when expressed in high copy number, will confer resistance to this antibiotic. We demonstrate that the IMH3(r) gene from strain 1006 has three amino acid changes, two of which are nonconservative, and demonstrate that at least two of the three mutations are required to confer resistance to MPA. We used this gene as a dominant selectable marker in clinical isolates of C. albicans and Candida tropicalis. We also identified the presence of autonomously replicating sequence elements that permit autonomous replication in the promoter region of this gene. Finally, we found the excision of a phi-type long terminal repeat element outside the IMH3 open reading frame of the gene in some strains. We used the IMH3(r) allele to disrupt one allele of ARG4 in two clinical isolates, WO-1 and FC18, thus demonstrating that a single ectopic integration of this dominant selectable marker is sufficient to confer resistance to MPA. PMID- 11119496 TI - Serotype AD strains of Cryptococcus neoformans are diploid or aneuploid and are heterozygous at the mating-type locus. AB - Cryptococcus neoformans is a pathogenic basidiomycete with a defined sexual cycle involving mating between haploid yeast cells with a transient diploid state. C. neoformans occurs in four predominant serotypes (A, B, C, and D), which represent different varieties or species. Rare clinical and environmental isolates with an unusual AD serotype have been reported and suggested to be diploid. We found by fluorescence-activated cell sorter analysis that serotype AD strains are aneuploid or diploid. PCR analysis with primers specific for serotype A or D alleles of the CNA1, CLA4, and GPA1 genes revealed that both alleles are often present in serotype AD strains. PCR analysis with primers specific for genes in the MATa or MATalpha mating-type loci revealed that serotype AD strains are heterozygous for the mating-type locus. Interestingly, in several serotype AD strains, the MATalpha locus was derived from the serotype D parent and the MATa locus was inherited from a serotype A parent that has been thought to be extinct. Basidiospores from a self-fertile serotype AD strain bearing the putative serotype A MATa locus showed a very low viability ( approximately 5%), and no fertile serotype A MATa strain could be recovered. Serotype AD strains were virulent in a murine model. Hybrid AD strains could readily be isolated following a laboratory cross between a serotype A strain and a serotype D strain. In summary, serotype AD strains of C. neoformans are unusual aneuploid or diploid strains that result from matings between serotype A and D strains. Self-fertile isolates fail to undergo normal meiosis because of genetic divergence. Our findings further suggest that serotype A MATa strains may exist in nature. PMID- 11119497 TI - Human resistance to Plasmodium falciparum increases during puberty and is predicted by dehydroepiandrosterone sulfate levels. AB - Immunity to Plasmodium falciparum develops slowly in areas of endemicity, and this is often ascribed to poorly immunogenic or highly variant parasite antigens. However, among populations newly exposed to malaria, adults acquire immunity more rapidly than children. We examined the relationship between pubertal development and resistance to P. falciparum. During two transmission seasons in western Kenya, we treated the same cohort of young males to eradicate P. falciparum and then obtained blood smears each week for 4 months. We determined pubertal development by Tanner staging and by levels of dehydroepiandrosterone sulfate (DHEAS) and testosterone in plasma. In multivariate and age-stratified analyses, we examined the effect of pubertal development on resistance to malaria. In both seasons (n = 248 and 144 volunteers, respectively), older males were less susceptible than younger males. Age-related decreases in the frequency and density of parasitemia were greatest during puberty (15- to 20-year-olds). DHEAS and testosterone were significant independent predictors of resistance to P. falciparum parasitemia, even after accounting for the effect of age. Fifteen- to 20-year-old males with high DHEAS levels had a 72% lower mean parasite density (P<0.01) than individuals with low DHEAS levels. Similarly, 21- to 35-year-old males with high DHEAS levels had a 92% lower mean parasite density (P<0.001) and 48% lower frequency of parasitemia (P<0.05) than individuals with low DHEAS levels. These data suggest that the long period needed to attain full immunity could be explained as a consequence of host development rather than as the requirement to recognize variant or poorly immunogenic parasite antigens. PMID- 11119498 TI - Granulocyte-macrophage colony-stimulating factor-deficient mice have impaired resistance to blood-stage malaria. AB - The contribution of granulocyte-macrophage colony-stimulating factor (GM-CSF), a hematopoietic and immunoregulatory cytokine, to resistance to blood-stage malaria was investigated by infecting GM-CSF-deficient (knockout [KO]) mice with Plasmodium chabaudi AS. KO mice were more susceptible to infection than wild-type (WT) mice, as evidenced by higher peak parasitemia, recurrent recrudescent parasitemia, and high mortality. P. chabaudi AS-infected KO mice had impaired splenomegaly and lower leukocytosis but equivalent levels of anemia compared to infected WT mice. Both bone marrow and splenic erythropoiesis were normal in infected KO mice. However, granulocyte-macrophage colony formation was significantly decreased in these tissues of uninfected and infected KO mice, and the numbers of macrophages in the spleen and peritoneal cavity were significantly lower than in infected WT mice. Serum levels of gamma interferon (IFN-gamma) were found to be significantly higher in uninfected KO mice, and the level of this cytokine was not increased during infection. In contrast, IFN-gamma levels were significantly above normal levels in infected WT mice. During infection, tumor necrosis factor alpha (TNF-alpha) levels were significantly increased in KO mice and were significantly higher than TNF-alpha levels in infected WT mice. Our results indicate that GM-CSF contributes to resistance to P. chabaudi AS infection and that it is involved in the development of splenomegaly, leukocytosis, and granulocyte-macrophage hematopoiesis. GM-CSF may also regulate IFN-gamma and TNF-alpha production and activity in response to infection. The abnormal responses seen in infected KO mice may be due to the lack of GM-CSF during development, to the lack of GM-CSF in the infected mature mice, or to both. PMID- 11119499 TI - Phenotypic analysis and virulence of Candida albicans LIG4 mutants. AB - In previous studies, we reported the isolation and preliminary characterization of a DNA ligase-encoding gene of Candida albicans. This gene (LIG4) is the structural and functional homologue of both yeast and human ligase IV, which is involved in nonhomologous end joining (NHEJ) of DNA double-strand breaks. In the present study, we have shown that there are no other LIG4 homologues in C. albicans. In order to study the function of LIG4 in morphogenesis and virulence, we constructed gene deletions. LIG4 transcript levels were reduced in the heterozygote and were completely absent in null strains. Concomitantly, the heterozygote showed a pronounced defect in myceliation, which was slightly greater in the null strain. This was true with several solid and liquid media, such as Spider medium, medium 199, and 2% glucose-1% yeast extract-2% Bacto Peptone, at several pHs. Reintroduction of the wild-type allele into the null mutant partially restored the ability of cells to form hyphae. In agreement with the positive role of LIG4 in morphogenesis, we detected a significant rise in mRNA levels during the morphological transition. LIG4 is not essential for DNA replication or for the repair of DNA damage induced by ionizing radiation or UV light, indicating that these lesions are repaired primarily by homologous recombination. However, our data show that the NHEJ apparatus of C. albicans may control morphogenesis in this diploid organism. In addition, deletion of one or both copies of LIG4 resulted in attenuation of virulence in a murine model of candidiasis. PMID- 11119500 TI - Cyclic nucleotide signaling in Toxoplasma gondii bradyzoite differentiation. AB - The ability of Toxoplasma gondii tachyzoites to differentiate into latent bradyzoite forms is essential for pathogenesis of clinical disease. We examined the effects of cyclic nucleotides on T. gondii bradyzoite differentiation in vitro. Differentiation of tachyzoites to bradyzoites was measured in an immunofluorescence assay using ME49 or its clonal derivative PLK, two well characterized T. gondii strains. Treatment of human fibroblast cultures infected with T. gondii with 8-(4-chlorophenylthio)-cyclic GMP (CPT-cGMP), a membrane permeable, nonhydrolyzable analogue of cGMP, resulted in an increased percentage of bradyzoite-positive vacuoles. Cyclic AMP (cAMP) also induced in vitro conversion of PLK, but the method of cAMP elevation was critical. Forskolin raises cAMP levels transiently and induced bradyzoites, whereas agents predicted to cause sustained elevation of cAMP were inhibitory to parasite conversion. Levels of cAMP were measured in host cells and extracellular tachyzoites. Forskolin, CPT-cGMP, and agents known to induce bradyzoite formation elevated cAMP in host cells and PLK parasites. These data suggest cyclic nucleotide signaling pathways are important in the stress-induced conversion of T. gondii tachyzoites to bradyzoites. Furthermore, because cAMP elevation was seen in PLK but not RH, a T. gondii strain that did not differentiate well in our assay, cAMP signaling within the parasite is likely to be critical. PMID- 11119501 TI - Cloning and expression of the Actinobacillus actinomycetemcomitans thioredoxin (trx) gene and assessment of cytokine inhibitory activity. AB - Thioredoxin is a ubiquitous redox control and cell stress protein. Unexpectedly, in recent years, thioredoxins have been found to exhibit both cytokine and chemokine activities, and there is increasing evidence that this class of protein plays a role in the pathogenesis of inflammatory diseases. In spite of this evidence, it has been reported that the oral bacterium and periodontopathogen Actinobacillus actinomycetemcomitans secretes an immunosuppressive factor (termed suppressive factor 1 [SF1] [T. Kurita-Ochiai and K. Ochiai, Infect. Immun. 64:50 54, 1996]) whose N-terminal sequence, we have determined, identifies it as thioredoxin. We have cloned and expressed the gene encoding the thioredoxin of A. actinomycetemcomitans and have purified the protein to homogeneity. The A. actinomycetemcomitans trx gene has 52 and 76% identities, respectively, to the trx genes of Escherichia coli and Haemophilus influenzae. Enzymatic analysis revealed that the recombinant protein had the expected redox activity. When the recombinant thioredoxin was tested for its capacity to inhibit the production of cytokines by human peripheral blood mononuclear cells, it showed no significant inhibitory capacity. We therefore conclude that the thioredoxin of A. actinomycetemcomitans does not act as an immunosuppressive factor, at least with human leukocytes in cultures, and that the identity of SF1 remains to be elucidated. PMID- 11119502 TI - Description of staphylococcus serine protease (ssp) operon in Staphylococcus aureus and nonpolar inactivation of sspA-encoded serine protease. AB - Signature tagged mutagenesis has recently revealed that the Ssp serine protease (V8 protease) contributes to in vivo growth and survival of Staphylococcus aureus in different infection models, and our previous work indicated that Ssp could play a role in controlling microbial adhesion. In this study, we describe an operon structure within the ssp locus of S. aureus RN6390. The ssp gene encoding V8 protease is designated as sspA, and is followed by sspB, which encodes a 40.6 kDa cysteine protease, and sspC, which encodes a 12.9-kDa protein of unknown function. S. aureus SP6391 is an isogenic derivative of RN6390, in which specific loss of SspA function was achieved through a nonpolar allelic replacement mutation. In addition to losing SspA, the culture supernatant of SP6391 showed a loss of 22- to 23-kDa proteins and the appearance of a 40-kDa protein corresponding to SspB. Although the 40-kDa SspB protein could degrade denatured collagen, our data establish that this is a precursor form which is normally processed by SspA to form a mature cysteine protease. Culture supernatant of SP6391 also showed a new 42-kDa glucosaminidase and enhanced glucosaminidase activity in the 29 to 32 kDa range. Although nonpolar inactivation of sspA exerted a pleiotropic effect, S. aureus SP6391 exhibited enhanced virulence in a tissue abscess infection model relative to RN6390. Therefore, we conclude that SspA is required for maturation of SspB and plays a role in controlling autolytic activity but does not by itself exert a significant contribution to the development of tissue abscess infections. PMID- 11119503 TI - Silencing and reactivation of urease in Yersinia pestis is determined by one G residue at a specific position in the ureD gene. AB - Yersinia pestis, the plague agent, is a naturally nonureolytic microorganism, while all other Yersinia species display a potent urease activity. In this report we demonstrate that Y. pestis harbors a complete urease locus composed of three structural (ureABC) and four accessory (ureEFGD) genes. Absence of ureolytic activity is due to the presence of one additional G residue in a poly(G) stretch, which introduces a premature stop codon in ureD. The presence of the same additional G in eight other Y. pestis isolates indicates that this mutation is species specific. Spontaneous excision of the extra G occurs at a frequency of 10(-4) to 10(-5) and restores a ureolytic phenotype to Y. pestis. The virulence of two independent ureolytic clones of Y. pestis injected either intravenously, subcutaneously, or intragastrically did not differ from that of the parental strain in the mouse infection model. Coinfection experiments with an equal number of ureolytic and nonureolytic bacteria did not evidence any difference in the ability of the two variants to multiply in vivo and to cause a lethal infection. Altogether our results demonstrate that variation of one extra G residue in ureD determines the ureolytic activity of Y. pestis but does not affect its virulence for mice or its ability to multiply and disseminate. PMID- 11119504 TI - Legionella pneumophila major acid phosphatase and its role in intracellular infection. AB - Legionella pneumophila is an intracellular pathogen of protozoa and alveolar macrophages. This bacterium contains a gene (pilD) that is involved in both type IV pilus biogenesis and type II protein secretion. We previously demonstrated that the PilD prepilin peptidase is crucial for intracellular infection by L. pneumophila and that the secreted pilD-dependent proteins include a metalloprotease, an acid phosphatase, an esterase/lipase, a phospholipase A, and a p-nitrophenyl phosphorylcholine hydrolase. Since mutants lacking type IV pili, the protease, or the phosphorylcholine hydrolase are not defective for intracellular infection, we sought to determine the significance of the secreted acid phosphatase activity. Three mutants defective in acid phosphatase activity were isolated from a population of mini-Tn10-mutagenized L. pneumophila. Supernatants as well as cell lysates from these mutants contained minimal acid phosphatase activity while possessing normal levels of other pilD-dependent exoproteins. Genetic studies indicated that the gene affected by the transposon insertions encoded a novel bacterial histidine acid phosphatase, which we designated Map for major acid phosphatase. Subsequent inhibitor studies indicated that Map, like its eukaryotic homologs, is a tartrate-sensitive acid phosphatase. The map mutants grew within macrophage-like U937 cells and Hartmannella amoebae to the same degree as did wild-type legionellae, indicating that this acid phosphatase is not essential for L. pneumophila intracellular infection. However, in the course of characterizing our new mutants, we gained evidence for a second pilD-dependent acid phosphatase activity that, unlike Map, is tartrate resistant. PMID- 11119505 TI - Novel group of virulence activators within the AraC family that are not restricted to upstream binding sites. AB - Several regulators within the AraC family control the expression of genes known or thought to be required for virulence of bacterial pathogens. One of these, Rns, activates transcription from an unprecedented variety of binding-site locations. Although nearly all prokaryotic activators bind within a small region upstream and adjacent to the promoter that they regulate, Rns does not bind within this region to activate its own promoter, Prns. Instead, to activate Prns, Rns requires one binding site 224.5 bp upstream and one downstream of the transcription start site. We show in this study that several other AraC family activators recognize the same binding sites as Rns and share with it the ability to utilize a downstream binding site. Like Rns, other members of this group of activators positively regulate the expression of virulence factors in pathogenic bacteria. These regulators are also able to activate transcription from promoter proximal upstream binding sites since they are able to substitute for Rns at Pcoo, a promoter with only upstream binding sites. Thus, Rns is the prototype for a group of regulators, which include CfaR, VirF, AggR, and CsvR and which activate transcription from locations that are more diverse than those of any other known activator. PMID- 11119506 TI - Susceptibility to secondary Francisella tularensis live vaccine strain infection in B-cell-deficient mice is associated with neutrophilia but not with defects in specific T-cell-mediated immunity. AB - Previous studies have demonstrated a role for B cells, not associated with antibody production, in protection against lethal secondary infection of mice with Francisella tularensis live vaccine strain (LVS). However, the mechanism by which B cells contribute to this protection is not known. To study the specific role of B cells during secondary LVS infection, we developed an in vitro culture system that mimics many of the same characteristics of in vivo infection. Using this culture system, we showed that B cells do not directly control LVS infection but that control of LVS growth is mediated primarily by LVS-primed T cells. Importantly, B cells were not required for the generation of effective memory T cells since LVS-primed, B-cell-deficient (BKO) mice generated CD4(+) and CD8(+) T cells that controlled LVS infection similarly to LVS-primed CD4(+) and CD8(+) T cells from wild-type mice. The control of LVS growth appeared to depend primarily on gamma interferon and nitric oxide and was similar in wild-type and BKO mice. Rather, the inability of BKO mice to survive secondary LVS infection was associated with marked neutrophil influx into the spleen very early after challenge. The neutrophilia was directly associated with B cells, since BKO mice reconstituted with naive B cells prior to a secondary challenge with LVS had decreased bacterial loads and neutrophils in the spleen and survived. PMID- 11119507 TI - Salmonella enterica serotype typhimurium elicits cross-immunity against a Salmonella enterica serotype enteritidis strain expressing LP fimbriae from the lac promoter. AB - The biological significance of fimbrial phase variation in Salmonella serotypes is currently unknown. Exposure to long polar (LP) fimbriae of Salmonella enterica serotype Typhimurium results in selection against lpf phase ON cells of serotype Enteritidis during a subsequent challenge, suggesting that fimbrial phase variation may be a mechanism to evade cross-immunity between Salmonella serotypes. This notion was tested by assessing the effect of an immune response against serotype Typhimurium LP fimbriae on colonization of mice with a serotype Enteritidis mutant in which the lpf promoter region was replaced with the Escherichia coli lac promoter. During a challenge with a serotype Enteritidis mutant carrying the lac promoter in front of the lpf operon, significantly lower numbers were recovered from organs and feces of mice previously immunized with an lpf phase ON culture of serotype Typhimurium than from mice not previously exposed to LP fimbriae. Immunization with the lpf phase ON culture of serotype Typhimurium elicited antibodies that cross-reacted with a purified gluthathione-S transferase-LpfA fusion protein of serotype Enteritidis. These data suggested that cross-immunity against LP fimbrial proteins cannot be evaded if phase variation on the transcriptional level is prevented by expressing the lpf operon from the lac promoter. These data hence support the idea that phase variation of LP fimbriae is a mechanism to evade cross-immunity between serotypes Enteritidis and Typhimurium. PMID- 11119508 TI - Activation of Valpha14(+) natural killer T cells by alpha-galactosylceramide results in development of Th1 response and local host resistance in mice infected with Cryptococcus neoformans. AB - We examined the effect of alpha-galactosylceramide (alpha-GalCer) on the synthesis of gamma interferon (IFN-gamma) and local resistance in mice infected intravenously with Cryptococcus neoformans. The level of IFN-gamma in serum increased on day 3, reached a peak level on day 7, and decreased to the basal level on day 14 postinfection in mice treated with alpha-GalCer, while in vehicle treated mice, no increase was detected at any time points except for a small increase on day 7. Such effects were not observed in NKT-KO mice. In CD4KO mice, minor synthesis of IFN-gamma was detected on day 3 in sera but was completely abolished by day 7. The alpha-GalCer-induced IFN-gamma production on day 3 was partially reduced in mice depleted of NK cells by treatment with anti-asialo GM(1) antibody (Ab). Spleen cells obtained from infected and alpha-GalCer-treated mice on day 7 produced a large amount of IFN-gamma upon restimulation with live organisms, while only a marginal level of production was detected in splenocytes from infected and vehicle-treated mice. Such effects were abolished in CD4KO and NKT-KO mice. Finally, the fungal loads in the lungs and spleen on days 7 and 14 were significantly reduced in alpha-GalCer-treated mice compared to those in control mice. In NKT-KO mice, local resistance elicited by alpha-GalCer was completely abolished, although no obvious exacerbation of infection was detected. Furthermore, treatment with anti-IFN-gamma monoclonal Ab mostly abrogated the protective effect of this agent. Thus, our results indicated that activation of Valpha14(+) NKT cells resulted in an increased Th1 response and local resistance to C. neoformans through production of IFN-gamma. PMID- 11119509 TI - Haemophilus influenzae porin contributes to signaling of the inflammatory cascade in rat brain. AB - In the present study we observed that the Haemophilus influenzae type b (Hib) porin, among the different surface bacterial components, is involved in the pathophysiology of bacterial meningitis. This study demonstrates that inoculation of Hib porin into the fourth cerebral ventricle causes the simultaneous expression of interleukin-1alpha (IL-1alpha), tumor necrosis factor alpha (TNF alpha), and macrophage inflammatory protein 2 (MIP-2) at 6 h after inoculation. At 24 h, the expression of MIP-2 decreases while the expression of IL-1alpha and TNF-alpha increases. The mRNA expression of IL-1alpha, TNF-alpha, and MIP-2 is correlated with injury to the blood-brain barrier as demonstrated by the appearance of serum proteins and leukocytes in cerebrospinal fluid and by the increase in brain water content. PMID- 11119510 TI - Signaling via interleukin-4 receptor alpha chain is required for successful vaccination against schistosomiasis in BALB/c mice. AB - Although protective immunity in C57BL/6 mice induced by a single dose of the radiation-attenuated schistosome vaccine is believed to be mediated by Th1-type immune responses, we here report that in BALB/c mice protection can also depend upon signaling via the interleukin-4 (IL-4) receptor which conventionally governs the development of Th2-type immune responses. We show that in BALB/c mice deficient for the IL-4 receptor alpha chain (IL-4Ralpha(-/-)), which are unresponsive to IL-4 and IL-13, vaccine-induced protection is abrogated compared with that in wild-type (WT) mice. In vaccinated IL-4Ralpha(-/-) mice, IL-12p40 production by cells from the skin exposure site was elevated, although gamma interferon (IFN-gamma) production in draining lymphoid tissues was similar in WT and IL-4Ralpha(-/-) mice. Nevertheless, the effector response in IL-4Ralpha(-/-) mice was Th1 biased with elevated IFN-gamma in the lungs and higher immunoglobulin G2a (IgG2a) and IgG2b titers but negligible quantities of Th2 associated IgG1 and IgE. Interestingly, levels of IL-4 were equivalent in WT and IL-4Ralpha(-/-) mice, indicating that Th2 responses were not dependent upon signaling by IL-4 or IL-13. No differences in the phenotype and composition of the pulmonary effector mechanism that might explain the failure to induce protection in IL-4Ralpha(-/-) mice were detected. However, passive transfer of partial protection to naive IL-4Ralpha(-/-) mice, using serum from vaccinated WT mice, indicates that Th2-associated antibodies such as IgG1 have a role in parasite elimination in BALB/c strain mice and that signaling via IL-4R can be an important factor in the generation of protection. PMID- 11119511 TI - Human T- and B-cell responses to Schistosoma mansoni recombinant glyceraldehyde 3 phosphate dehydrogenase correlate with resistance to reinfection with S. mansoni or Schistosoma haematobium after chemotherapy. AB - Recently we reported that human T- and B-cell recognition of a 42-kDa protein (p42) in soluble extracts of adult Schistosoma mansoni worms correlates with resistance to reinfection with S. mansoni or S. haematobium. Amino acid microsequencing of p42 revealed that it consists predominantly of schistosome glyceraldehyde 3-phosphate dehydrogenase (SG3PDH). We have expressed SG3PDH in Escherichia coli and purified the recombinant protein in a soluble and enzymatically active form. Recombinant SG3PDH (rSG3PDH) reacted with human monospecific antibodies to p42. Lymphoproliferation and production of interleukin 4 and gamma interferon (IFN-gamma) after in vitro stimulation with rSG3PDH and serum isotype responses to rSG3PDH were examined in individuals with extremes of resistance and susceptibility to reinfection after treatment of previous S. mansoni or S. haematobium infection. Lymphoproliferation and IFN-gamma production in response to rSG3PDH and the presence of serum immunoglobulin G1 (IgG1), IgG3, and IgA antibodies to rSG3PDH generally characterized individuals who are resistant to reinfection after chemotherapy. The data indicate that T- and B-cell immune reactivity to rSG3PDH correlates with resistance to reinfection, confirming previous studies identifying SG3PDH as a target of protective immunity in humans, and suggest that SG3PDH should be investigated as a possible vaccine for human schistosomiasis. PMID- 11119512 TI - Vervet monkeys vaccinated with killed Leishmania major parasites and interleukin 12 develop a type 1 immune response but are not protected against challenge infection. AB - Leishmania major is a protozoan parasite that causes chronic cutaneous lesions that often leave disfiguring scars. Infections in mice have demonstrated that leishmanial vaccines that include interleukin-12 (IL-12) as an adjuvant are able to induce protective immunity. In this study, we assessed the safety, immunopotency, and adjuvant potential of two doses of IL-12 when used with a killed L. major vaccine in vervet monkeys. The induction of cell-mediated immunity following vaccination was determined by measuring delayed-type hypersensitivity, in vitro lymphocyte proliferation, and gamma interferon (IFN gamma) production. Protection was assessed by challenging the animals with L. major parasites and monitoring the course of infection. At low doses of IL-12 (10 microg), a small increase in the parameters of cell-mediated immunity was observed, relative to those in animals that received antigen without IL-12. However, none of these animals were protected against a challenge infection. At higher doses of IL-12 (30 microg), a substantial increase in Leishmania-specific immune responses was observed, and monkeys immunized with antigen and IL-12 exhibited an IFN-gamma response that was as great as that in animals that had resolved a primary infection and were immune. Nevertheless, despite the presence of correlates of protection, the disease course was only slightly altered, and protection was low compared to that in self-cured monkeys. These data suggest that protection against leishmaniasis may require more than the activation of Leishmania-specific IFN-gamma-producing T cells, which has important implications for designing a vaccine against leishmaniasis. PMID- 11119513 TI - Recombinant antigen-enterotoxin A2/B chimeric mucosal immunogens differentially enhance antibody responses and B7-dependent costimulation of CD4(+) T cells. AB - The ADP-ribosylating enterotoxins, cholera toxin (CT) and the Escherichia coli heat-labile toxin (LT-IIa), have been shown to enhance mucosal and systemic antibody (Ab) responses to coadministered antigens. The purpose of the present study was to compare the ability of the nontoxic A2/B subunits of these toxins, which have distinct targeting properties, to augment the immunogenicity of a genetically coupled protein antigen. Structurally similar chimeric proteins were generated by genetically replacing the toxic A1 subunit of CT or LT-IIa with the saliva-binding region (SBR) from the streptococcal adhesin AgI/II. Intranasal immunization of BALB/c mice with either chimeric protein induced significantly higher plasma and mucosal anti-SBR immunoglobulin A (IgA) and IgG Ab responses than SBR alone. Moreover, compared to SBR-LT-IIaA2/B, SBR-CTA2/B elicited significantly higher levels of plasma IgG1 and salivary IgA anti-SBR Ab responses. Ex vivo and in vitro experiments revealed that SBR-CTA2/B selectively up-regulated B7-2 expression on murine B cells isolated from both the nasal associated lymphoid tissue, cervical lymph nodes, and spleen. In contrast, SBR-LT IIaA2/B had little effect on B7-1 or B7-2 expression on B220(+), CD11b(+), or CD11c(+) cells. Analysis of the functional costimulatory activity of SBR-CTA2/B treated B cells revealed a significant enhancement in anti-CD3-stimulated CD4(+) T-cell proliferative responses, and this proliferation was significantly reduced by treatment with anti-B7-2 but not with anti-B7-1 or isotype control Abs. Thus, SBR-CTA2/B and SBR-LT-IIaA2/B exhibit distinct patterns of antibody responses associated with differential effects on B7-2 expression and subsequent costimulatory effects on CD4(+) T cells. PMID- 11119514 TI - Development of a recombinant antigen vaccine against infection with the filarial worm Onchocerca volvulus. AB - Efforts to control Onchocerca volvulus, the etiologic agent of river blindness, have been limited to vector control and drug treatment to eliminate microfilariae, with no means available to prevent infection. The goal of this study was to develop a vaccine against this infection using recombinant antigens that are expressed in the early larval stages of the parasite. Five recombinant antigens, Ov7, Ov64, OvB8, Ov9M, and Ov73k, were identified by screening adult and larval cDNA libraries with antibodies from immune humans, chimpanzees, or rabbits. When mice were immunized with the five individual recombinant antigens, statistically significant reductions in parasite survival were induced in mice immunized with Ov7, OvB8, or Ov64, when administered in alum but not when injected in Freund's complete adjuvant (FCA). Live larvae recovered from control and immunized mice were analyzed to determine their developmental stages. A decrease in the percentage of larvae molting from the third stage to the fourth stage was observed with mice immunized with Ov7, Ov64, or OvB8 in alum but not with mice immunized with Ov9 and Ov73k or with mice immunized with any of the five antigens in FCA. Mice immunized with a cocktail of the three protective antigens developed protective immunity equal to that seen with mice immunized with individual antigens. This study has identified, for the first time, three recombinant antigens capable of inducing protective immunity to O. volvulus. Furthermore, since the antigens functioned with alum as the adjuvant, this vaccine could potentially be used clinically to prevent river blindness in humans. PMID- 11119515 TI - Soluble egg antigen-stimulated T helper lymphocyte apoptosis and evidence for cell death mediated by FasL(+) T and B cells during murine Schistosoma mansoni infection. AB - Granuloma formation around schistosomal eggs is induced by soluble egg antigens (SEA) and mediated by the activity of CD4(+) Th lymphocytes and their cytokines. Regulation of the inflammatory Th cell response during infection is still insufficiently understood. The hypothesis of this study was that activation induced cell death (AICD) of CD4(+) T cells is involved in the immune inflammatory response. This study investigated the dynamics of splenic and granuloma CD4(+) Th cell apoptosis and Fas ligand (FasL) expression during the acute and chronic stages of murine schistosomal infection. Enhanced apoptosis of freshly isolated CD4(+) Th lymphocytes commenced after egg deposition and persisted during the peak and modulated phases of granuloma formation. After oviposition, CD4(+), CD8(+), and CD19(+) splenocytes and granuloma cells expressed elevated levels of FasL but FasL expression declined during the downmodulated stage of infection. In culture, SEA induced splenic and granuloma CD4(+) T-cell apoptosis and stimulated expression of FasL on splenic but not granuloma CD4(+) T cells, CD8(+) T cells, and CD19(+) B cells. SEA-stimulated splenocytes and granuloma cells preferentially lysed a Fas-transfected target cell line. Depletion of B cells from SEA-stimulated splenic cultures decreased CD4(+) T cell apoptosis. Coculture of purified splenic B cells with CD4(+) T cells and adoptive transfer of purified B cells indicated that antigen-stimulated B cells can kill CD4(+) Th cells. However, CD4(+) T cells were the dominant mediators of apoptosis in the granuloma. This study indicates that AICD is involved in the apoptosis of CD4(+) T cells during schistosomal infection. PMID- 11119516 TI - Invasion of human epithelial cells by Pseudomonas aeruginosa involves src-like tyrosine kinases p60Src and p59Fyn. AB - Pseudomonas aeruginosa plays a major role in respiratory tract infections or sepsis in patients with cystic fibrosis or upon suppression of the immune system. Several P. aeruginosa strains have been shown to be internalized by human epithelial cells; however, the molecular mechanisms of the invasion process are poorly characterized. Here, we show that the internalization of P. aeruginosa into human epithelial cells results in and requires activation of the Src-like tyrosine kinases p59Fyn and p60Src and the consequent tyrosine phosphorylation of several eukaryotic proteins. The significance of Src-like tyrosine kinase activation is shown by an almost complete blockade of P. aeruginosa internalization, but not adhesion, upon inhibition of Src-like tyrosine kinases. Likewise, inhibition of P. aeruginosa binding to CFTR, which has been shown to block P. aeruginosa internalization, prevents Src and Fyn activation, supporting a pivotal role of Src-like tyrosine kinases for invasion by P. aeruginosa. PMID- 11119517 TI - Modulation of human immune response by Echinococcus granulosus antigen B and its possible role in evading host defenses. AB - By directly suppressing the function of certain immune cell subsets and by stimulating other cell populations related to immunopathology, parasite-derived substances play an important role in the chronic establishment of parasitic disease. Our objective was twofold: (i) to investigate further the role of Echinococcus granulosus antigen B (AgB) in the human early inflammatory response by determining its effect on polymorphonuclear cell (PMN) random migration, chemotaxis, and oxidative metabolism and (ii) to determine its action in acquired immunity by evaluating AgB and sheep hydatid fluid (SHF)-driven Th1 (gamma interferon [IFN-gamma] and interleukin 12 [IL-12]) and Th2 (IL-4 and IL-13) cytokine production by peripheral blood mononuclear cells (PBMC) from 40 patients who had cured or stable or progressive cystic echinococcosis. AgB significantly inhibited PMN recruitment but left their random migration and oxidative metabolism unchanged. Patients' PBMC stimulated with AgB produced IL-4 and IL-13 but did not produce IL-12. They also produced significantly lower IFN-gamma concentrations than did PBMC stimulated with SHF (P = 10(-5)). AgB skewed the Th1/Th2 cytokine ratios towards a preferentially immunopathology-associated Th2 polarization, predominantly in patients with progressive disease. AgB-stimulated patients' PBMC also proliferated less than SHF-stimulated PBMC (P = 9 x 10(-3)). In vitro Th2 cytokine production was reflected in vivo by elevated specific immunoglobulin E (IgE) and IgG4 antibodies binding to AgB. These findings confirm that AgB plays a role in the escape from early immunity by inhibiting PMN chemotaxis. They also add new information on the host-parasite relationship, suggesting that AgB exploits the activation of T helper cells by eliciting a nonprotective Th2 cell response. PMID- 11119518 TI - Group B Streptococcus capsular polysaccharide-cholera toxin B subunit conjugate vaccines prepared by different methods for intranasal immunization. AB - Group B Streptococcus (GBS) type III capsular polysaccharide (CPS III) was conjugated to recombinant cholera toxin B subunit (rCTB) using three different methods which employed (i) cystamine and N-succinimidyl-3-(2 pyridyldithio)propionate (SPDP), (ii) carbodiimide with adipic acid dihydrazide (ADH) as a spacer, or (iii) reductive amination (RA). The CPS III-rCTB conjugates were divided into large- and small-molecular-weight (M(r)) fractions, and the immunogenicities of the different preparations after intranasal (i.n.) immunization were studied in mice. Both large- and small-M(r) conjugates of CPS III-rCTB(RA) or CPS III-rCTB(ADH) induced high, almost comparable levels of CPS specific immunoglobulin G (IgG) in serum, lungs, and vagina that were generally superior to those obtained with CPS III-rCTB(SPDP) conjugates or a CPS III and rCTB mixture. However, the smaller-M(r) conjugates of CPS III-rCTB(RA) or CPS III rCTB(ADH) in most cases elicited a lower anti-CPS IgA immune response than the large-M(r) conjugates, and the highest anti-CPS IgA titers in both tissues and serum were obtained with the large-M(r) CPS III-rCTB(RA) conjugate. Serum IgG anti-CPS titers induced by the CPS III-rCTB(RA) conjugate had high levels of specific IgG1, IgG2a, IgG2b, and IgG3 antibodies. Based on the effectiveness of RA for coupling CPS III to rCTB, RA was also tested for conjugating GBS CPS Ia with rCTB. As for the CPS III-rCTB conjugates, the immunogenicity of CPS Ia was greatly increased by conjugation to rCTB. Intranasal immunization with a combination of CPS Ia-rCTB and CPS III-rCTB conjugates was shown to induce anti CPS Ia and III immune responses in serum and lungs that were fully comparable with the responses to immunization with the monovalent CPS Ia-rCTB or CPS III rCTB conjugates. These results suggest that the GBS CPS III-rCTB and CPS Ia-rCTB conjugates prepared by the RA method may be used in bivalent and possibly also in multivalent mucosal GBS conjugate vaccines. PMID- 11119519 TI - Mapping of binding domains of nontypeable Haemophilus influenzae HMW1 and HMW2 adhesins. AB - Nontypeable Haemophilus influenzae is an important cause of localized respiratory tract disease, which begins with colonization of the upper respiratory mucosa. In previous work we reported that the nontypeable H. influenzae HMW1 and HMW2 proteins are high-molecular-weight nonpilus adhesins responsible for attachment to human epithelial cells, an essential step in the process of colonization. Interestingly, although HMW1 and HMW2 share significant sequence similarity, they display distinct cellular binding specificities. In order to map the HMW1 and HMW2 binding domains, we generated a series of complementary HMW1-HMW2 chimeric proteins and examined the ability of these proteins to promote in vitro adherence by Escherichia coli DH5alpha. Using this approach, we localized the HMW1 and HMW2 binding domains to an approximately 360-amino-acid region near the N terminus of the mature HMW1 and HMW2 proteins. Experiments with maltose-binding protein fusion proteins containing segments of either HMW1 or HMW2 confirmed these results and suggested that the fully functional binding domains may be conformational structures that require relatively long stretches of sequence. Of note, the HMW1 and HMW2 binding domains correspond to areas of maximal sequence dissimilarity, suggesting that selective advantage associated with broader adhesive potential has been a major driving force during H. influenzae evolution. These findings should facilitate efforts to develop a subcomponent vaccine effective against nontypeable H. influenzae disease. PMID- 11119520 TI - espC pathogenicity island of enteropathogenic Escherichia coli encodes an enterotoxin. AB - At least five proteins are secreted extracellularly by enteropathogenic Escherichia coli (EPEC), a leading cause of infant diarrhea in developing countries. However only one, EspC, is known to be secreted independently of the type III secretion apparatus encoded by genes located within the 35.6-kb locus of enterocyte effacement pathogenicity island. EspC is a member of the autotransporter family of proteins, and the secreted portion of the molecule is 110 kDa. Here we determine that the espC gene is located within a second EPEC pathogenicity island at 60 min on the chromosome of E. coli. We also show that EspC is an enterotoxin, indicated by rises in short-circuit current and potential difference in rat jejunal tissue mounted in Ussing chambers. In addition, preincubation with antiserum against the homologous Pet enterotoxin of enteroaggregative E. coli eliminated EspC enterotoxin activity. Like the EAF plasmid, the espC pathogenicity island was found only in a subset of EPEC, suggesting that EspC may play a role as an accessory virulence factor in some but not all EPEC strains. PMID- 11119521 TI - vimA gene downstream of recA is involved in virulence modulation in Porphyromonas gingivalis W83. AB - A 0.9-kb open reading frame encoding a unique 32-kDa protein was identified downstream of the recA gene of Porphyromonas gingivalis. Reverse transcription PCR and Northern blot analysis showed that both the recA gene and this open reading frame are part of the same transcriptional unit. This cloned fragment was insertionally inactivated using the ermF-ermAM antibiotic resistance cassette to create a defective mutant by allelic exchange. When plated on Brucella blood agar, the mutant strain, designated P. gingivalis FLL92, was non-black pigmented and showed significant reduction in beta-hemolysis compared with the parent strain, P. gingivalis W83. Arginine- and lysine-specific cysteine protease activities, which were mostly soluble, were approximately 90% lower than that of the parent strain. Expression of the rgpA, rgpB, and kgp protease genes was the same in P. gingivalis FLL92 as in the wild-type strain. In contrast to the parent strain, P. gingivalis FLL92 showed increased autoaggregration in addition to a significant reduction in hemagglutinating and hemolysin activities. In in vivo experiments using a mouse model, P. gingivalis FLL92 was dramatically less virulent than the parent strain. A molecular survey of this mutant and the parent strain using all known P. gingivalis insertion sequence elements as probes suggested that no intragenomic changes due to the movement of these elements have occurred in P. gingivalis FLL92. Taken together, these results suggest that the recA downstream gene, designated vimA (virulence-modulating gene), plays an important role in virulence modulation in P. gingivalis W83, possibly representing a novel posttranscriptional or translational regulation of virulence factors in P. gingivalis. PMID- 11119522 TI - Avidity, potency, and cross-reactivity of monoclonal antibodies to pneumococcal capsular polysaccharide serotype 6B. AB - Many pneumococcal capsular polysaccharides (PSs) are similar in structure, and a pneumococcal antibody often binds to all of the PSs with a similar structure. Yet, these cross-reactive antibodies may bind to the structurally related pneumococcal capsular PSs with an avidity too low to be effective. If memory B cells producing such weakly cross-reactive antibodies are elicited with pneumococcal conjugate vaccines, the memory cells for low-avidity antibodies could compromise the subsequent immune responses to the cross-reactive PS (original antigenic sin). To investigate these issues, we produced 14 hybridomas secreting monoclonal antibodies (MAbs) to the capsular PS of Streptococcus pneumoniae serotype 6B by immunizing BALB/c mice with antigens containing 6B PS and studied their epitope, avidity, in vitro opsonizing capacity, in vivo protective capacity, and "antigen binding titer" by enzyme-linked immunosorbent assay (ELISA) of 6A and 6B capsular PSs. Six MAbs bound to the non-cross-reactive 6B-specific epitope, and seven MAbs bound to the cross-reactive epitope present in both 6A and 6B PSs One MAb (Hyp6BM6) revealed a novel epitope. This epitope was found on 6A PS in solution, but not on 6A PS adsorbed onto the plastic surface of the ELISA plates. The avidity of the MAb for 6A or 6B PS ranged from 7.8 x 10(6) M(-1) to 4.1 x 10(11) M(-1). No MAbs were weakly cross-reactive, since none of the cross-reactive MAbs showed any tendency toward having less avidity to 6A PS (the cross-reactive PS) than to 6B PS. Avidity influenced the results of several antibody assays. When all of the hybridomas were examined, avidity strongly correlated with the titer of a unit amount of MAb to bind antigen-coated ELISA plates (r = 0.91) or to opsonize pneumococci in vitro (r = 0.85). Because both assay results are avidity dependent, the ELISA and the opsonization assay results were strongly correlated (r = 0.91), regardless of avidity. Avidity also correlated with the potency of a MAb to passively protect mice against pneumococcal infections. When only the immunoglobulin G hybridomas were examined, little increase in opsonizing capacity and in vivo protective potency was observed above 10(9) M(-1). Taken together, an ELISA measuring antigen binding titer may be an adequate measure of the protective immunity induced with pneumococcal vaccines, and the absence of a partially cross-reactive MAb suggests that antigenic sin may not be significant in responses to vaccines against the S. pneumoniae 6B serotype. PMID- 11119523 TI - Clonal associations among Staphylococcus aureus isolates from various sites of infection. AB - A molecular epidemiological analysis was undertaken to identify lineages of Staphylococcus aureus that may be disproportionately associated with infection. Pulsed-field gel electrophoresis analysis of 405 S. aureus clinical isolates collected from various infection types and geographic locations was performed. Five distinct S. aureus lineages (SALs 1, 2, 4, 5, and 6) were identified, which accounted for 19.01, 9.14, 22.72, 10.12, and 4.69% of isolates, respectively. In addition, 85 lineages which occurred with frequencies of <2.5% were identified and were termed "sporadic." The most prevalent lineage was methicillin-resistant S. aureus (SAL 4). The second most prevalent lineage, SAL 1, was also isolated at a high frequency from the anterior nares of healthy volunteers, suggesting that its prevalence among clinical isolates may be a consequence of high carriage rates in humans. Gene-specific PCR was carried out to detect genes for a number of staphylococcal virulence traits. tst and cna were found to be significantly associated with prevalent lineages compared to sporadic lineages. When specific infection sites were examined, SAL 4 was significantly associated with respiratory tract infection, while SAL 2 was enriched among blood isolates. SAL 1 and SAL 5 were clonally related to SALs shown by others to be widespread in the clinical isolate population. We conclude from this study that at least five phylogenetic lineages of S. aureus are highly prevalent and widely distributed among clinical isolates. The traits that confer on these lineages a propensity to infect may suggest novel approaches to antistaphylococcal therapy. PMID- 11119524 TI - Targeted reduction in expression of Trypanosoma cruzi surface glycoprotein gp90 increases parasite infectivity. AB - A previous study had shown that the expression of gp90, a stage-specific surface glycoprotein of Trypanosoma cruzi metacyclic trypomastigotes, is inversely correlated with the parasite's ability to invade mammalian cells. By using antisense oligonucleotides complementary to a region of the gp90 gene implicated in host cell adhesion, we investigated whether the selective inhibition of gp90 synthesis affected the capacity of metacyclic forms to enter target cells. Parasites were incubated for 24 h with 20 microM PS1, a phosphorothioate oligonucleotide based on a sequence of the gp90 coding strand; PS2, the antisense counterpart of PS1; or PO2, the unmodified version of PS2 containing phosphodiester linkages, and the expression of surface molecules was analyzed by flow cytometry and immunoblotting using specific monoclonal antibodies. PS2 but not PS1 or PO2 inhibited the expression of gp90. Inhibition by PS2 was dose dependent. Northern blot analysis revealed that steady-state gp90 mRNA levels were diminished in PS2-treated parasites compared to untreated controls. Treatment with PS2 did not affect the expression of other metacyclic stage surface glycoproteins involved in parasite-host cell interaction, such as gp82 and the mucin-like gp35/50. Expression of gp90 was also inhibited by other phosphorothioate oligonucleotides targeted to the gp90 gene (PS4, PS5, PS6, and PS7) but not by PS3, with the same base composition as PS2 but a mismatched sequence. Parasites treated with PS2, PS4, or PS5 entered HeLa cells in significantly higher numbers than untreated controls, whereas the invasive capacity of PS1- and PS3-treated parasites was unchanged, confirming the inverse association between infectivity and gp90 expression. PMID- 11119525 TI - Biochemical and biological properties of Staphylococcal enterotoxin K. AB - Staphylococcus aureus is an important human pathogen which is implicated in a wide variety of diseases. Major determinants of the virulence of this organism include extracellular virulence factors. Staphylococcal enterotoxins (SEs) are important causative agents in staphylococcal toxic shock syndrome and food poisoning. Our study identified a novel enterotoxin, SEK, and examined its biochemical and biological properties. SEK had a molecular weight of 26,000 and an experimentally determined pI of between 7.0 and 7.5. SEK was secreted by clinical isolates of S. aureus. We demonstrated that SEK had many of the biological activities associated with the SEs, including superantigenicity, pyrogenicity, the ability to enhance the lethal effect of endotoxin, and lethality in a rabbit model when administered by subcutaneous miniosmotic pump. Recombinant SEK was shown to stimulate human CD4(+) and CD8(+) T cells in a Vbeta specific manner; T-cells bearing Vbeta 5.1, 5.2, and 6.7 were significantly stimulated to proliferate. PMID- 11119526 TI - Salmonella pathogenicity island 2 influences both systemic salmonellosis and Salmonella-induced enteritis in calves. AB - We have used signature-tagged mutagenesis to identify mutants of the host specific Salmonella enterica serotype Dublin which were avirulent in calves and/or BALB/c mice. A mutant with a transposon insertion in the sseD gene of Salmonella pathogenicity island 2 (SPI-2), which encodes a putative secreted effector protein, was identified. This mutant was recovered from the bovine host but not from the murine host following infection with a pool of serotype Dublin mutants. However, a pure inoculum of the sseD mutant was subsequently shown to be attenuated in calves following infection either by the intravenous route or by the oral route. The sseD mutant was fully invasive for bovine intestinal mucosa but was subsequently unable to proliferate to the same numbers as the parental strain in vivo. Both the sseD mutant and a second SPI-2 mutant, with a transposon insertion in the ssaT gene, induced significantly weaker secretory and inflammatory responses in bovine ligated ileal loops than did the parental strain. These results demonstrate that SPI-2 is required by serotype Dublin for the induction of both systemic and enteric salmonellosis in calves. PMID- 11119527 TI - Role of plasma, lipopolysaccharide-binding protein, and CD14 in response of mouse peritoneal exudate macrophages to endotoxin. AB - Plasma lipopolysaccharide (LPS)-binding protein (LBP) and membrane CD14 function to enhance the responses of monocytes to low concentrations of endotoxin. Surprisingly, recent reports have suggested that LBP or CD14 may be dispensable for macrophage responses to low concentrations of LPS or may even exert an inhibitory effect in the case of LBP. We therefore investigated whether LBP and CD14 participated in the response of mouse peritoneal exudate macrophages (PEM) to LPS stimulation. In the presence of a low amount of plasma (<1%) or of recombinant mouse or human LBP, PEM were found to respond to low concentrations of LPS (<5 to 10 ng/ml) in an LBP- and CD14-dependent manner. However, tumor necrosis factor production (not interleukin-6 production) by LPS-stimulated PEM was reduced when cells were stimulated in the presence of higher concentrations of plasma or serum (5 or 10%). Yet, the inhibitory effect of plasma or serum was not mediated by LBP. Taken together with previous results obtained with LBP and CD14 knockout mice in models of experimental endotoxemia, the present data confirm a critical part for LBP and CD14 in innate immune responses of both blood monocytes and tissue macrophages to endotoxins. PMID- 11119528 TI - Vaccination with calpain induces a Th1-biased protective immune response against Schistosoma japonicum. AB - A large subunit of calpain, a calcium-activated neutral proteinase, from Schistosoma japonicum was cloned and expressed in Escherichia coli. When BALB/c mice were immunized with purified recombinant calpain (r-calpain) emulsified in complete Freund's adjuvant, a significant reduction in the number of recovered worms and also in egg production per female worm was observed (P<0.01). Spleen cells of the immunized mice showed enhanced production of gamma interferon (IFN gamma) by activated CD4(+) T cells. Considering our observation of elevated expression of inducible nitric oxide synthase mRNA in immunized mice, r-calpain induced IFN-gamma seemed to upregulate the production of nitric oxide by macrophages and subsequently mediated the killing of schistosomulae in the lung. On the other hand, spleen cells of immunized mice showed only faint interleukin-4 production in response to r-calpain in vitro, suggesting that immunization with r calpain alters the Th1-Th2 balance in murine hosts even during a Th2-promoting S. japonicum infection. Furthermore, histopathological study of the livers of immunized mice showed that granulomas formed around eggs were diminished in both size and number. Egg production by female worms was clearly decreased in immunized mice, suggesting that r-calpain also has antifecundity effects. Taken together, these results point to S. japonicum calpain as a potential vaccine candidate for both worm killing and disease prevention, possibly through the induction of a strong Th1-dominant environment in immunized mice. PMID- 11119529 TI - Identification and disruption of two discrete loci encoding hyaluronic acid capsule biosynthesis genes hasA, hasB, and hasC in Streptococcus uberis. AB - The hyaluronic acid capsule of Streptococcus uberis has been implicated in conferring resistance to phagocytosis by bovine neutrophils. Construction of a bank of random insertion mutants of S. uberis (strain 0140J) was achieved using the pGh9::ISS1 mutagenesis system (22). Phenotypic screening of approximately 5,000 clones enabled the isolation of 11 acapsular mutants. Southern hybridization indicated that two mutants carried a lesion within a group of genes similar to those involved in the assembly of the hyaluronic acid capsule found in the group A Streptococcus (GAS) has operon. The DNA sequence flanking the points of insertion confirmed the presence of homologues of GAS hasA and hasB in S. uberis. The DNA sequence flanking the ISS1 insertion in another mutant identified a homologue of hasC in S. uberis. The GAS hasABC operon structure was not conserved in S. uberis, and two discrete loci comprising homologues of either hasAB or hasC were identified. Disruption of S. uberis hasA or hasC resulted in the complete cessation of hyaluronic acid capsule production. Correspondingly, these mutants were found to have lost their resistance to phagocytosis by bovine neutrophils. The bactericidal action of bovine neutrophils on S. uberis 0140J was shown unequivocally to depend upon the capsule status of the bacterium. PMID- 11119530 TI - Temperature-regulated protein synthesis by Leptospira interrogans. AB - Leptospira interrogans is an important mammalian pathogen. Transmission from an environmental source requires adaptations to a range of new environmental conditions in the organs and tissues of the infected host. Since many pathogenic bacteria utilize temperature to discern their environment and regulate the synthesis of appropriate proteins, we investigated the effects of temperature on protein synthesis in L. interrogans. Bacteria were grown for several days after culture temperatures were shifted from 30 to 37 degrees C. Triton X-114 cellular fractionation identified several proteins of the cytoplasm, periplasm, and outer membrane for which synthesis was dependent on the culture temperature. Synthesis of a cytoplasmic protein of 20 kDa was switched off at 37 degrees C, whereas synthesis of a 66-kDa periplasmic protein was increased at the higher temperature. Increased synthesis of a 25-kDa outer membrane protein was observed when the organisms were shifted from 30 to 37 degrees C. A 36-kDa protein synthesized at 30 but not at 37 degrees C was identified as LipL36, an outer membrane lipoprotein. In contrast, expression of another lipoprotein, LipL41, was the same at either temperature. Immunoblotting with convalescent equine sera revealed that some proteins exhibiting thermoregulation of synthesis elicited antibody responses during infection. Our results show that sera from horses which aborted as a result of naturally acquired infection with L. interrogans serovar pomona type kennewicki recognize periplasmic and outer membrane proteins which are differentially synthesized in response to temperature and which therefore may be important in the host-pathogen interaction during infection. PMID- 11119531 TI - Secreted aspartic proteinase family of Candida tropicalis. AB - Medically important yeasts of the genus Candida secrete aspartic proteinases (Saps), which are of particular interest as virulence factors. Like Candida albicans, Candida tropicalis secretes in vitro one dominant Sap (Sapt1p) in a medium containing bovine serum albumin (BSA) as the sole source of nitrogen. Using the gene SAPT1 as a probe and under low-stringency hybridization conditions, three new closely related gene sequences, SAPT2 to SAPT4, encoding secreted proteinases were cloned from a C. tropicalis lambdaEMBL3 genomic library. All bands identified by Southern blotting of EcoRI-digested C. tropicalis genomic DNA with SAPT1 could be assigned to a specific SAP gene. Therefore, the SAPT gene family of C. tropicalis is likely to contain only four members. Interestingly, the SAPT2 and SAPT3 gene products, Sapt2p and Sapt3p, which have not yet been detected in C. tropicalis cultures in vitro, were produced as active recombinant enzymes with the methylotrophic yeast Pichia pastoris as an expression system. As expected, reverse transcriptase PCR experiments revealed a strong SAPT1 signal with RNA extracted from cells grown in BSA medium. However, a weak signal was obtained with all other SAPT genes under several conditions tested, showing that these SAPT genes could be expressed at a basic level. Together, these experiments suggest that the gene products Sapt2p, Sapt3p, and Sapt4p could be produced under conditions yet to be described in vitro or during infection. PMID- 11119533 TI - Inhibition of adhesion of Plasmodium falciparum-infected erythrocytes by structurally defined hyaluronic acid dodecasaccharides. AB - We recently reported that Plasmodium falciparum-infected erythrocytes (IRBCs) can adhere to hyaluronic acid (HA), which appears to be a receptor, in addition to chondroitin sulfate A (CSA), for parasite sequestration in the placenta. Further investigations of the nature and specificity of this interaction indicate that HA oligosaccharide fragments competitively inhibit parasite adhesion to immobilized purified HA in a size-dependent manner, with dodecasaccharides being the minimum size for maximum inhibition. Rigorously purified and structurally defined HA dodecasaccharides, free of contamination by CSA or other glycosaminoglycans, effectively inhibited IRBC adhesion to HA but not CSA, providing compelling evidence of a specific interaction between IRBCs and HA. PMID- 11119532 TI - Role of Fc gamma receptors in triggering host cell activation and cytokine release by Borrelia burgdorferi. AB - Borrelia burgdorferi, the spirochetal bacterium that causes human Lyme disease, encodes numerous lipoproteins which have the capacity to trigger the release of proinflammatory cytokines from a variety of host cell types, and it is generally believed that these cytokines contribute to the disease process in vivo. We previously reported that low-passage-number infectious B. burgdorferi spirochetes express a novel lipidation-independent activity which induces secretion of the proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) by the mouse MC/9 mast cell line. Using RNase protection assays, we determined that mast cells exposed in vitro to low-passage-number, but not high-passage-number, B. burgdorferi spirochetes show increased expression of additional mRNAs representing several chemokines, including macrophage-inflammatory protein 1alpha (MIP-1alpha), MIP-1beta, and TCA3, as well as the proinflammatory cytokine interleukin-6. Furthermore, mast cell TNF-alpha secretion can be inhibited by the phosphatidylinositol 3-kinase inhibitor wortmannin and also by preincubation with purified mouse immunoglobulin G1 (IgG1) and IgG2a, but not mouse IgG3, and by a mouse Fc gamma receptor II and III (FcgammaRII/III)-specific rat monoclonal antibody, suggesting the likely involvement of host FcgammaRIII in B. burgdorferi mediated signaling. A role for passively adsorbed rabbit or bovine IgG or serum components in B. burgdorferi-mediated FcgammaR signaling was excluded in control experiments. These studies confirm that low-passage-number B. burgdorferi spirochetes express a novel activity which upregulates the expression of a variety of host cell chemokine and cytokine genes, and they also establish a novel antibody-independent role for FcgammaRs in transduction of activation signals by bacterial products. PMID- 11119534 TI - Role of genetic resistance in invasive pneumococcal infection: identification and study of susceptibility and resistance in inbred mouse strains. AB - From a panel of nine inbred mice strains intranasally infected with Streptococcus pneumoniae type 2 strain, BALB/c mice were resistant and CBA/Ca and SJL mice were susceptible to infection. Further investigation revealed that BALB/c mice were able to prevent proliferation of pneumococci in the lungs and blood, whereas CBA/Ca mice showed no bacterial clearance. Rapidly increasing numbers of bacteria in the blood was a feature of CBA/Ca but not BALB/c mice. In the lungs, BALB/c mice recruited significantly more neutrophils than CBA/Ca mice at 12 and 24 h postinfection. Inflammatory lesions in BALB/c mice were visible much earlier than in CBA/Ca mice, and there was a greater cellular infiltration into the lung tissue of BALB/c mice at the earlier time points. Our data suggest that resistance or susceptibility to intranasal pneumococci may have an association with recruitment and/or function of neutrophils. PMID- 11119536 TI - Decreased infectivity in Borrelia burgdorferi strain B31 is associated with loss of linear plasmid 25 or 28-1. AB - Previous reports indicated a correlation between loss of plasmids and decreased infectivity of Borrelia burgdorferi strain B31, suggesting that plasmids may encode proteins that are required for pathogenesis. In this study, we expand on this correlation. Using the B. burgdorferi genomic sequence, we designed primers specific for each plasmid, and by using PCR we catalogued 11 linear and 2 circular plasmids from 49 clonal isolates of a mid-passage B. burgdorferi strain B31, initially derived from infected mouse skin, and 20 clones obtained from mouse skin infected with a low-passage isolate of B. burgdorferi strain B31. Among the 69 clones analyzed, nine distinct genotypes were identified relative to wild-type B. burgdorferi strain B31. Among the nine clonal genotypes obtained, only the 9-kb circular plasmid (cp9), the 25-kb linear plasmid (lp25), and either the 28-kb linear plasmid 1 or 4 (lp28-1 and lp28-4, respectively) were missing, in different combinations. We compared the infectivity of the wild-type strain, containing all known B. burgdorferi plasmids, with those of single mutants lacking either lp28-1, lp28-4, or lp25 and a double mutant missing both cp9 and lp28-1. The infectivity data indicated that B. burgdorferi strain B31 cells lacking lp28-4 were modestly attenuated in all tissues analyzed, whereas samples missing lp25 were completely attenuated in all tissues, even at the highest inoculum tested. Isolates without lp28-1 infected the joint tissue yet were not able to infect other tissues as effectively. In addition, we have observed a selection in vivo in the skin, bladder, and joint for cells containing lp25 and in the skin and bladder for cells containing lp28-1, indicating that lp25 and lp28-1 encode proteins required for colonization and short-term maintenance in these mammalian tissues. In contrast, there was no selection in the joint for cells containing lp28-1, suggesting that genes on lp28-1 are not required for colonization of B. burgdorferi within the joint. These observations imply that the dynamic nature of the B. burgdorferi genome may provide the genetic heterogeneity necessary for survival in the diverse milieus that this pathogen occupies in nature and may contribute to tropism in certain mammalian host tissues. PMID- 11119535 TI - Characterization of Vibrio cholerae O1 El tor galU and galE mutants: influence on lipopolysaccharide structure, colonization, and biofilm formation. AB - Recently we described the isolation of spontaneous bacteriophage K139-resistant Vibrio cholerae O1 El Tor mutants. In this study, we identified phage-resistant isolates with intact O antigen but altered core oligosaccharide which were also affected in galactose catabolism; this strains have mutations in the galU gene. We inactivated another gal gene, galE, and the mutant was also found to be defective in the catabolism of exogenous galactose but synthesized an apparently normal lipopolysaccharide (LPS). Both gal mutants as well as a rough LPS (R-LPS) mutant were investigated for the ability to colonize the mouse small intestine. The galU and R-LPS mutants, but not the galE mutant, were defective in colonization, a phenotype also associated with O-antigen-negative mutants. By investigating several parameters in vitro, we could show that galU and R-LPS mutants were more sensitive to short-chain organic acids, cationic antimicrobial peptides, the complement system, and bile salts as well as other hydrophobic agents, indicating that their outer membrane no longer provides an effective barrier function. O-antigen-negative strains were found to be sensitive to complement and cationic peptides, but they displayed significant resistance to bile salts and short-chain organic acids. Furthermore, we found that galU and galE are essential for the formation of a biofilm in a spontaneous phage resistant rugose variant, suggesting that the synthesis of UDP-galactose via UDP glucose is necessary for biosynthesis of the exopolysaccharide. In addition, we provide evidence that the production of exopolysaccharide limits the access of phage K139 to its receptor, the O antigen. In conclusion, our results indicate involvement of galU in V. cholerae virulence, correlated with the observed change in LPS structure, and a role for galU and galE in environmental survival of V. cholerae. PMID- 11119537 TI - Borrelia spirochetes upregulate release and activation of matrix metalloproteinase gelatinase B (MMP-9) and collagenase 1 (MMP-1) in human cells. AB - Borrelia burgdorferi, the spirochetal agent of Lyme disease, stimulated human peripheral blood monocytes to release pro-matrix metalloproteinase-9 (gelatinase B; pro-MMP-9) and active matrix metalloproteinase-1 (collagenase-1; MMP-1). Human neutrophils also released pro-MMP-9 and a 130-kDa protein with gelatinolytic activity in response to live B. burgdorferi. In addition, U937 cells and human keratinocyte cells were also stimulated to release pro-MMP-9 under the same conditions. However, human umbilical vein endothelial cells (HUVECs) released pro MMP-9 and pro-MMP-2 in a constitutive manner and were not influenced by live spirochetes. MMPs produced by human monocytes also enhanced the penetration of B. burgdorferi through extracellular matrix component barriers in vitro. Plasmin stabilized on the surface of the Lyme disease spirochete was shown to activate pro-MMP-9 to its active form. This active form was also observed in the plasma of mice infected with a relapsing fever borrelia. These results suggest that borreliae can upregulate MMPs and possibly mediate an activation cascade initiated by plasmin bound to the microbial surface. MMPs may play a role in dissemination of the Lyme disease spirochete and in the pathogenesis of Borrelia infection. PMID- 11119538 TI - Improved innate immunity of endotoxin-tolerant mice increases resistance to Salmonella enterica serovar typhimurium infection despite attenuated cytokine response. AB - During infection with gram-negative bacteria, exposure of immune cells to lipopolysaccharide (LPS) from the bacterial cell membrane induces a rapid cytokine response which is essential for the activation of host defenses against the invading pathogens. Administration of LPS to mice induces a state of hyporesponsiveness, or tolerance, characterized by reduced cytokine production upon subsequent LPS challenge. In the model of experimental Salmonella enterica serovar Typhimurium infection of mice, we assessed the question of whether complete LPS tolerance induced by repetitive doses of LPS interfered with cytokine production and host defense against gram-negative bacteria. Although production of various cytokines in response to serovar Typhimurium was attenuated by LPS pretreatment, LPS-tolerant mice showed improved antibacterial activity, evidenced by a prolongation of survival and a continuously lower bacterial load. We attribute this protective effect to three independent mechanisms. (i) Peritoneal accumulation of leukocytes in the course of LPS pretreatment accounted for enhanced defense against serovar Typhimurium during the first 6 h of infection but not for decreased bacterial load in late-stage infection. (ii) LPS tolerant mice had an increased capacity to recruit neutrophilic granulocytes during infection. (iii) LPS-tolerant mice showed threefold-increased Kupffer cell numbers, enhanced phagocytic activity of the liver, and strongly improved clearance of blood-borne serovar Typhimurium. These results demonstrate that despite attenuated cytokine response, acquired LPS tolerance is associated with enhanced resistance to infections by gram-negative bacteria and that this effect is mainly mediated by improved effector functions of the innate immune system. PMID- 11119539 TI - Actinobacillus pleuropneumoniae iron transport and urease activity: effects on bacterial virulence and host immune response. AB - Actinobacillus pleuropneumoniae, a porcine respiratory tract pathogen, has been shown to express transferrin-binding proteins and urease during infection. Both activities have been associated with virulence; however, their functional role for infection has not yet been elucidated. We used two isogenic A. pleuropneumoniae single mutants (DeltaexbB and DeltaureC) and a newly constructed A. pleuropneumoniae double (DeltaureC DeltaexbB) mutant in aerosol infection experiments. Neither the A. pleuropneumoniae DeltaexbB mutant nor the double DeltaureC DeltaexbB mutant was able to colonize sufficiently long to initiate a detectable humoral immune response. These results imply that the ability to utilize transferrin-bound iron is required for multiplication and persistence of A. pleuropneumoniae in the porcine respiratory tract. The A. pleuropneumoniae DeltaureC mutant and the parent strain both caused infections that were indistinguishable from one another in the acute phase of disease; however, 3 weeks postinfection the A. pleuropneumoniae DeltaureC mutant, in contrast to the parent strain, could not be isolated from healthy lung tissue. In addition, the local immune response-as assessed by fluorescence-activated cell sorter and enzyme-linked immunosorbent spot analyses-revealed a significantly higher number of A. pleuropneumoniae-specific B cells in the bronchoalveolar lavage fluid (BALF) of pigs infected with the A. pleuropneumoniae DeltaureC mutant than in the BALF of those infected with the parent strain. These results imply that A. pleuropneumoniae urease activity may cause sufficient impairment of the local immune response to slightly improve the persistence of the urease-positive A. pleuropneumoniae parent strain. PMID- 11119540 TI - Synthesis and surface expression of CD14 by human endothelial cells. AB - Previous studies have reported that human vascular endothelial cells lack the membrane-bound lipopolysaccharide (LPS) receptor, CD14 (mCD14). By optimizing assay conditions, including the selection of anti-CD14 monoclonal antibody, we now demonstrate that human umbilical vein endothelial cells (HUVEC) express CD14 on the cell surface. Single-passage HUVEC showed approximately 20 times less expression of CD14 than monocytes. Interestingly, there was significant loss of surface CD14 expression with increasing numbers of culture passages. Evidence for synthesis of CD14 by HUVEC was provided by the finding that L-[(35)S]methionine was incorporated into CD14. In addition, the expression of CD14 on HUVEC was upregulated by LPS, lysophosphatidic acid, and tissue culture supplements, and this upregulation was dependent on protein synthesis. Furthermore, the results imply that mCD14 is required for LPS-induced activation of endothelial cells in the absence of serum and that it acts in concert with serum factors (soluble CD14). Our results provide evidence that CD14 is expressed by endothelial cells and suggest that the previous inability to observe expression of this molecule has been due to culture and staining conditions. This finding has important implications for the understanding of the mechanisms by which LPS stimulates endothelial cells and the management of sepsis caused by gram-negative bacteria. PMID- 11119541 TI - Brucella suis-impaired specific recognition of phagosomes by lysosomes due to phagosomal membrane modifications. AB - Brucella species are gram-negative, facultatively intracellular bacteria that infect humans and animals. These organisms can survive and replicate within a membrane-bound compartment in phagocytic and nonprofessional phagocytic cells. Inhibition of phagosome-lysosome fusion has been proposed as a mechanism for intracellular survival in both types of cells. However, the biochemical mechanisms and microbial factors implicated in Brucella maturation are still completely unknown. We developed two different approaches in an attempt to gain further insight into these mechanisms: (i) a fluorescence microscopy analysis of general intracellular trafficking on whole cells in the presence of Brucella and (ii) a flow cytometry analysis of in vitro reconstitution assays showing the interaction between Brucella suis-containing phagosomes and lysosomes. The fluorescence microscopy results revealed that fusion properties of latex bead containing phagosomes with lysosomes were not modified in the presence of live Brucella suis in the cells. We concluded that fusion inhibition was restricted to the pathogen phagosome and that the host cell fusion machinery was not altered by the presence of live Brucella in the cell. By in vitro reconstitution experiments, we observed a specific association between killed B. suis-containing phagosomes and lysosomes, which was dependent on exogenously supplied cytosol, energy, and temperature. This association was observed with killed bacteria but not with live bacteria. Hence, this specific recognition inhibition seemed to be restricted to the pathogen phagosomal membrane, as noted in the in vivo experiments. PMID- 11119542 TI - Microtubule- and dynein-mediated movement of Orientia tsutsugamushi to the microtubule organizing center. AB - The host cell microfilaments and microtubules (MTs) are known to play a critical role in the life cycles of several pathogenic intracellular microbes by providing for successful invasion and promoting movement of the pathogen once inside the host cell cytoplasm. Orientia tsutsugamushi, an obligate intracellular bacterium, enters host cells by induced phagocytosis, escapes to the cytosol, and then replicates in the cytosol. ECV304 cells infected with O. tsutsugamushi revealed the colocalization of the MT organizing center (MTOC) and cytosolic orientiae by indirect immunofluorescence assay. Using immunofluorescence microscopy in the presence and absence of MT-depolymerizing agents (colchicine and nocodazole), it was shown that the cytosolic oriential movement was mediated by MTs. By transfection study (overexpression of dynamitin [also called p50], which is known to associate with dynein-dependent movement), the movement of O. tsutsugamushi to the MTOC was also mediated by dynein, the minus-end-directed MT-related motor. Although the significance of this movement in the life cycle of O. tsutsugamushi was not proven, we propose that the cytosolic O. tsutsugamushi bacteria use MTs and dyneins to propel themselves from the cell periphery to the MTOC. PMID- 11119543 TI - Initial characterization of CST1, a Toxoplasma gondii cyst wall glycoprotein. AB - Toxoplasma gondii is an important protozoan pathogen of humans that can cause encephalitis in immunocompromised individuals such as those with AIDS. This encephalitis is due to reactivation of latent infection in T. gondii-seropositive patients. Latent organisms survive within tissue cysts, which are specialized parasitophorous vacuoles containing bradyzoites. The cyst wall of this structure is produced by modification of the parasitophorous vacuole by the parasite and is important in cyst survival. The components of the cyst wall have been poorly characterized. By using immunofluorescence and immunoelectron microscopy, we have identified a monoclonal antibody (MAb 93.18) that reacts with the cyst wall. This antibody recognizes a 116-kDa glycoprotein, which we have termed CST1, containing sugar residues that bind Dolichos biflorans lectin (DBA). CST1 is distinct from T. gondii antigen labeled with succinyl Triticum vulgare lectin (S-WGA) and represents the major DBA-binding component in T. gondii. The carbohydrate components of the tissue cyst, such as CST1, are probably important in both providing stability and facilitating persistence in its host. As is seen in the carbohydrate capsules of fungi, glycoproteins in the T. gondii cyst wall may protect cysts from the immune response of the host. Further characterization of the formation of the cyst wall and its components should lead to insights into the mechanism of tissue cyst persistence and may suggest novel therapeutic approaches to eliminate tissue cysts of this organism. PMID- 11119544 TI - Legionella pneumophila entry gene rtxA is involved in virulence. AB - Successful parasitism of host cells by intracellular pathogens involves adherence, entry, survival, intracellular replication, and cell-to-cell spread. Our laboratory has been examining the role of early events, adherence and entry, in the pathogenesis of the facultative intracellular pathogen Legionella pneumophila. Currently, the mechanisms used by L. pneumophila to gain access to the intracellular environment are not well understood. We have recently isolated three loci, designated enh1, enh2, and enh3, that are involved in the ability of L. pneumophila to enter host cells. One of the genes present in the enh1 locus, rtxA, is homologous to repeats in structural toxin genes (RTX) found in many bacterial pathogens. RTX proteins from other bacterial species are commonly cytotoxic, and some of them have been shown to bind to beta(2) integrin receptors. In the current study, we demonstrate that the L. pneumophila rtxA gene is involved in adherence, cytotoxicity, and pore formation in addition to its role in entry. Furthermore, an rtxA mutant does not replicate as well as wild type L. pneumophila in monocytes and is less virulent in mice. Thus, we conclude that the entry gene rtxA is an important virulence determinant in L. pneumophila and is likely to be critical for the production of Legionnaires' disease in humans. PMID- 11119545 TI - Dendritic cells phagocytose and are activated by Treponema pallidum. AB - Cell-mediated immune processes play a prominent role in the clinical manifestations of syphilis, a sexually transmitted disease of humans caused by spirochetal bacterium Treponema pallidum. The immune cell type that initiates the early immune response to T. pallidum thus far has not been identified. However, dendritic cells (DCs) are the first immune-competent cells to encounter antigens within skin or mucous membranes, the principal sites of early syphilitic infection. In the present study, immature DC line XS52, derived from murine skin, was utilized to examine T. pallidum-DC interactions and subsequent DC activation (maturation). Electron microscopy revealed that T. pallidum was engulfed by DCs via both coiling and conventional phagocytosis and was delivered to membrane bound vacuoles. The XS52 DC line expressed surface CD14 and mRNA for Toll-like receptors 2 and 4, molecules comprising important signaling components for immune cell activation by bacterial modulins. Both T. pallidum and a synthetic lipopeptide (corresponding to the 47-kDa major membrane lipoprotein) activated the XS52 DC line, as indicated by the secretion of interleukin-12 (IL-12), IL 1beta, tumor necrosis factor alpha, and IL-6 and elevated surface expression of CD54. The combined data support the contention that DCs stimulated by T. pallidum and/or its proinflammatory membrane lipoproteins are involved in driving the cellular immune processes that typify syphilis. PMID- 11119546 TI - Role of Mycobacterium tuberculosis copper-zinc superoxide dismutase. AB - Superoxide dismutases (SODs) play an important role in protection against oxidative stress and have been shown to contribute to the pathogenicity of many bacterial species. To determine the function of the mycobacterial copper and zinc cofactored SOD (CuZnSOD), we constructed and characterized Mycobacterium tuberculosis and Mycobacterium bovis BCG CuZnSOD null mutants. Both strains were more sensitive to superoxides and hydrogen peroxide than were their respective parental strains. The survival of M. bovis BCG in unstimulated as well as activated mouse bone marrow-derived macrophages was not affected by the loss of CuZnSOD. The survival of CuZnSOD deficient-M. tuberculosis in guinea pig tissues was comparable to that of its parental strain. These results indicate that the mycobacterial CuZnSOD is not essential for intracellular growth within macrophages and does not detectably contribute to the pathogenicity of M. tuberculosis. PMID- 11119547 TI - Group A streptococcal rofA gene is involved in the control of several virulence genes and eukaryotic cell attachment and internalization. AB - The serotype M6 group A streptococcal RofA regulator was previously shown to exert a direct positive control of protein F1 expression and, concomitantly, fibronectin binding. Using a serotype M6 rofA mutant, we demonstrate here that this regulator has a potentially indirect negative influence on the expression of the mga, emm6, pel-sagA, and speA virulence genes. Additionally, the rofA mutant exhibited reduced eukaryotic cell internalization rates in combination with decreased host cell viability. PMID- 11119548 TI - Expression of ExsA in trans confers type III secretion system-dependent cytotoxicity on noncytotoxic Pseudomonas aeruginosa cystic fibrosis isolates. AB - Twelve Pseudomonas aeruginosa cystic fibrosis isolates that are not able to exert a type III secretion system (TTSS)-dependent cytotoxicity towards phagocytes have been further studied. The strains, although possessing TTSS genes and exsA, which encodes a positive regulator of the TTSS regulon, showed no transcriptional activation of the exsCBA regulatory operon. The expression of exsA in trans restored the in vitro secretion of TTSS proteins and ex vivo cytotoxicity. PMID- 11119549 TI - Carboxy-terminal proteolytic processing of Helicobacter pylori vacuolating toxin. AB - The vacA gene of Helicobacter pylori strain 60190 encodes a 1, 287-amino-acid protoxin, which undergoes cleavage of a 33-amino-acid amino-terminal signal sequence and carboxy-terminal proteolytic processing to yield a mature secreted toxin. Several features of VacA suggest that it belongs to the autotransporter family of gram-negative bacterial secreted proteins. Based on matrix-assisted laser desorption ionization-time of flight mass spectrometric analysis, we calculate that the mature toxin has a mass of 88.2+/-0.2 kDa and consists of approximately 821 amino acids. PMID- 11119550 TI - Aromatic compound-dependent Brucella suis is attenuated in both cultured cells and mouse models. AB - The aroC gene of the facultative intracellular pathogen Brucella suis was cloned and sequenced. The cloned aroC gene complements Escherichia coli and Salmonella enterica serovar Typhimurium aroC mutants. A B. suis aroC mutant was found to be unable to grow in a defined medium without aromatic compounds. The mutant was highly attenuated in tissue culture (THP1 macrophages and HeLa cells) and murine virulence models. PMID- 11119551 TI - Severity of group B streptococcal arthritis in selected strains of laboratory mice. AB - The susceptibilities of C3H/HeN, BALB/c, and C57BL/6N mouse strains to group B streptococci (GBS) infection were evaluated. C3H/HeN mice developed severe polyarthitis; mild lesions and no lesions were observed in BALB/c and C57BL/6N mice, respectively. A correlation between the severity of arthritis, the number of GBS in the joints, and local interleukin-6 and interleukin-1beta production was evident. PMID- 11119552 TI - Immunity against Helicobacter pylori: significance of interleukin-4 receptor alpha chain status and gender of infected mice. AB - Vaccination of interleukin-4 (IL-4) receptor alpha (IL-4Ralpha) chain-deficient BALB/c mice with Helicobacter pylori urease and cholera toxin or with urease expressing, live attenuated Salmonella enterica serovar Typhimurium cells revealed that protection against H. pylori infection is independent of IL-4- or IL-13-mediated signals. A comparison of male and female mice suggests a sexual dimorphism in the extent of bacterial colonization that is particularly evident in the absence of the IL-4Ralpha chain. PMID- 11119553 TI - Tir tyrosine phosphorylation and pedestal formation are delayed in enteropathogenic Escherichia coli sepZ::TnphoA mutant 30-5-1(3). AB - Enteropathogenic Escherichia coli (EPEC) strain 30-5-1(3) has been reported to form attaching and effacing (A/E) lesions without Tir tyrosine phosphorylation. In this study, we show that 30-5-1(3), which has a transposon insertion within the sepZ gene, forms wild-type A/E lesions including Tir tyrosine phosphorylation, but at a slower rate. A/E lesion formation by 30-5-1(3) occurs without detectable secretion of Tir or other EPEC Esp secreted proteins. PMID- 11119554 TI - Infection of fetal feline brain cells in culture with Bartonella henselae. AB - Bartonella henselae is known to cause central nervous system (CNS) disease in humans, and neurological signs have been observed in experimentally infected cats. However, the pathogenesis of CNS disease remains unclear. This study was undertaken to determine whether B. henselae infects feline fetal brain cells in vitro. Microglial-cell- and astrocyte-enriched cultures were inoculated with B. henselae. Gimenez staining identified bacterial organisms within microglial cells by day 7 postinoculation. The viability of the intracellular bacteria was demonstrated by incubating cultures with gentamicin and plating cell lysate on agar. Electron microscopy identified intracellular organisms with characteristic Bartonella morphology but identified no ultrastructural abnormalities within infected microglial cells. No evidence of infection was seen in Bartonella inoculated astrocyte cultures. These findings suggest a role for microglia in the pathogenesis of B. henselae-associated neurological disease. PMID- 11119555 TI - High-affinity, protective antibodies to the binding domain of botulinum neurotoxin type A. AB - Monoclonal antibodies (MAbs) were prepared against the putative binding domain of botulinum neurotoxin A (BoNT/A), a nontoxic 50-kDa fragment. Initially, all fusion products were screened against the holotoxin BoNT/A and against the binding fragment, BoNT/A H(C). Eleven neutralizing hybridomas were cloned, and their specific binding to BoNT/A H(C) was demonstrated by surface plasmon resonance, with dissociation constants ranging from 0.9 to <0.06 nM. Epitope mapping by real-time surface plasmon resonance showed that the antibodies bound to at least two distinct regions of the BoNT/A H(C) fragment. These MAbs will be useful tools for studying BoNT/A interactions with its receptor, and they have potential diagnostic and therapeutic applications. PMID- 11119556 TI - Tick salivary gland extract inhibits killing of Borrelia afzelii spirochetes by mouse macrophages. AB - Salivary gland extract (SGE) from Ixodes ricinus ticks inhibited the killing of Borrelia afzelii spirochetes by murine macrophages. SGE also reduced the production of two major defense molecules of phagocytes, superoxide and nitric oxide. It is likely that the suppression of macrophage microbicidal mechanisms contributes to the inhibitory effect of tick saliva on the killing of B. afzelii spirochetes, thus facilitating the transmission of this important pathogen. PMID- 11119557 TI - Mouse toxicity and cytokine release by verotoxin 1 B subunit mutants. AB - The crystal structure of the verotoxin 1 (VT1) B subunit complexed with a globotriaosylceramide (Gb(3)) analogue showed the presence of three receptor binding sites per monomer. We wished to study the effects of altering the three sites, singly or in combination, on animal toxicity and cytokine induction in vitro. We found that while the site 1 and 2 mutants were modestly (two- to sevenfold) reduced in their ability to cause disease in BALB/c mice, the site 3 mutant, W34A, was as toxic as VT1. However, all the double-mutant proteins, irrespective of which two sites were mutated, exhibited approximately a 100-fold reduction in their 50% lethal doses for mice. These results suggest that multivalent receptor binding is important in vivo and that all three binding sites make a similar contribution to the latter process. The triple-mutant holotoxin, F30A G62T W34A, administered intraperitoneally without adjuvant, stimulated a strong antibody response in BALB/c mice, and the immune sera neutralized the activity of VT1 in vitro. Induction of tumor neurosis factor alpha release from differentiated human monocytes (THP-1 cells) was relatively impaired for site 1 and site 2 but not site 3 mutants, suggesting an auxiliary role for the latter site in mediation of cytokine release in vitro. Cytotoxicity assays on undifferentiated THP-1 cells have also demonstrated the importance of sites 1 and 2 and the relatively small role played by site 3 in causing cell death. These data suggest an association between the cytotoxicity of the protein and its ability to induce cytokine release. PMID- 11119558 TI - Genetic diversity of Pseudomonas aeruginosa strains isolated from ventilated patients with nosocomial pneumonia, cancer patients with bacteremia, and environmental water. AB - Random amplified polymorphic DNA typing was used to study the genetic diversity of Pseudomonas aeruginosa strains from (i) ventilated patients with nosocomial pneumonia who were hospitalized in intensive care units, (ii) cases of bacteremia in cancer patients with severe neutropenia, and (iii) rivers and swimming pools. Genetic diversity was determined by three phylogenetic methods and by statistical analysis of population genetics. The population studied undergoes epidemic clonality with a high rate of genetic recombination. P. aeruginosa bacteremia and pneumonia are not caused by specific clones within this species. PMID- 11119559 TI - Severe schistosomiasis in the absence of interleukin-4 (IL-4) is IL-12 independent. AB - An interleukin-4 (IL-4)-dependent Th2 response allows wild-type mice to survive infection with the parasite Schistosoma mansoni. In the absence of IL-4, infected mice mount a Th1-like proinflammatory response, develop severe disease, and succumb. Neither the Th1 response nor morbidity is IL-12 dependent in this system. PMID- 11119560 TI - Homologous and heterologous Borrelia burgdorferi challenge of infection-derived immune rabbits using host-adapted organisms. AB - We have recently found that strain B31 infection-immune rabbits are completely protected against homologous challenge with large numbers (>10(6)) of host adapted Borrelia burgdorferi (HAB) (E. S. Shang, C. I. Champion, X. Wu, J. T. Skare, D. B. Blanco, J. N. Miller, and M. A. Lovett, Infect. Immun. 68:4189-4199, 2000). In this study, we have extended these findings to determine whether B31 strain infection-immune rabbits are also protected against heterologous HAB challenge. Infection-immune rabbits challenged with large numbers (>10(6)) of homologous HAB strain B31 were completely protected from erythema migrans (EM) and skin and disseminated infection. In contrast, infection-immune rabbits challenged with heterologous HAB strains N40 and Sh-2-82 were completely susceptible to EM and skin and disseminated infection; challenge with strain 297 also resulted in EM and infection of the skin and viscera, but clearance of infection occurred 3 weeks postchallenge. These findings confirm that immunity elicited in rabbits by B31 strain infection confers complete protection against large-dose homologous HAB challenge but not against a heterologous strain. PMID- 11119561 TI - Cytosolic delivery and characterization of the TcdB glucosylating domain by using a heterologous protein fusion. AB - TcdB from Clostridium difficile glucosylates small GTPases (Rho, Rac, and Cdc42) and is an important virulence factor in the human disease pseudomembranous colitis. In these experiments, in-frame genetic fusions between the genes for the 255 amino-terminal residues of anthrax toxin lethal factor (LFn) and the TcdB(1 556) coding region were constructed, expressed, and purified from Escherichia coli. LFnTcdB(1-556) was enzymatically active and glucosylated recombinant RhoA, Rac, Cdc42, and substrates from cell extracts. LFnTcdB(1-556) plus anthrax toxin protective antigen intoxicated cultured mammalian cells and caused actin reorganization and mouse lethality, all similar to those caused by wild-type TcdB. PMID- 11119562 TI - Effect of influenza A virus infection on nasopharyngeal colonization and otitis media induced by transparent or opaque phenotype variants of Streptococcus pneumoniae in the chinchilla model. AB - Phase variation in the colonial opacity of Streptococcus pneumoniae has been implicated as a factor in bacterial adherence, colonization, and invasion in the pathogenesis of pneumococcal disease. Additionally, the synergistic effects of influenza A virus and S. pneumoniae in the development of otitis media (OM) have been reported. This study examined the ability of opaque or transparent S. pneumoniae from the same strain in combination with an antecedent influenza A virus infection to colonize the nasopharynx and invade the middle ear in the chinchilla model. Our data indicated that there was no significant difference in the level of nasopharyngeal colonization and induction of OM between the opaque and transparent variants unless there was a prior challenge with influenza A virus. Subsequent to influenza A virus infection, there was a significant difference between the variants in the ability to colonize and persist in the nasopharynx and middle ear. The concentrations of the opaque variant in nasopharyngeal-lavage samples and middle-ear fluid remained consistently higher than those of the transparent variant for 10 days postinoculation. Data from this study indicate that the effects of influenza A virus on the pathogenesis of experimental S. pneumoniae-induced OM differ depending on the opacity phenotype involved. PMID- 11119563 TI - Identification of protective epitopes by sequencing of the major outer membrane protein gene of a variant strain of Chlamydia psittaci serotype 1 (Chlamydophila abortus). AB - Protective monoclonal antibodies (MAbs) to the major outer membrane protein (MOMP) of species of the family Chlamydiaceae, which is the primary vaccine candidate antigen, recognize nonlinear epitopes conferred by the oligomeric conformation of the molecule. Protective MAbs failed to recognize oligomeric MOMP of the variant strain LLG, which bears amino acid substitutions in variable segments (VSs) 1, 2, and 4, and competed with monomer-specific MAbs mapping to these VSs in reference strain 577. The results suggest that multiple sites located in the three VSs contribute to the epitope of protective MAbs. PMID- 11119564 TI - Differential interleukin-8 response of intestinal epithelial cell line to reactogenic and nonreactogenic candidate vaccine strains of Vibrio cholerae. AB - In this study, we analyzed whether attachment of Vibrio cholerae vaccine strains to human intestinal epithelial cells can induce an interleukin-8 (IL-8) response. The IL-8 transcripts were detected by PCR amplification of reverse-transcribed mRNA, and the gene product secretion was measured by an enzyme-linked immunosorbent assay. Infection of monolayers of the undifferentiated HT29-18N2 cell line with reactogenic (JBK70 and 81) and nonreactogenic (CVD103HgR and 638) vaccine strains of V. cholerae resulted in markedly higher IL-8 expression by epithelial cells exposed to reactogenic strains than by cells exposed to the nonreactogenic strains. Additionally, epithelial cells produced IL-8 transcripts following stimulation with cholera vaccine strains in a concentration-dependent manner. These results represent a new insight into the inflammatory component of reactogenicity and could be used as a predictive marker of vaccine reactogenicity prior to human testing. PMID- 11119565 TI - T-cell responses to immunodominant LACK antigen do not play a critical role in determining susceptibility of BALB/c mice to Leishmania mexicana. AB - Although BALB/c mice develop lesions when infected with Leishmania mexicana, the mechanisms which are responsible for susceptibility to this parasite have not been elucidated. In contrast, susceptibility of BALB/c mice to Leishmania major has been shown to depend on the early production of interleukin-4 (IL-4) by T cells which react to the parasitic LACK antigen. Here, we demonstrate that the lesions induced by L. mexicana are delayed compared to those induced by L. major but rapidly develop at later time points. Interestingly, while LACK-tolerant BALB/c-derived IE-LACK transgenic mice were resistant to L. major, they were susceptible to L. mexicana and developed lesions similar to those observed in wild-type BALB/c mice. The latter result was observed despite the fact that (i) LACK was expressed by L. mexicana, (ii) splenocytes from BALB/c mice were able to stimulate LACK-specific T-cell hybridoma cells when incubated with live L. mexicana promastigotes, and (iii) LACK-specific T cells contributed to IL-4 production in L. mexicana-infected BALB/c mice. Thus, in contrast to what was observed for L. major-infected mice, LACK-specific T cells do not play a critical role in determining susceptibility to L. mexicana. Although BALB/c mice are susceptible to both L. major and L. mexicana, the mechanisms which are responsible for susceptibility to these parasites are likely to be different. PMID- 11119566 TI - Characterization of the domain of fibronectin-binding protein I of Streptococcus pyogenes responsible for elicitation of a protective immune response. AB - Fibronectin-binding protein I (SfbI) represents a major adhesin of Streptococcus pyogenes. Mice were intranasally immunized with recombinant proteins spanning different portions of SfbI to identify the minimal fragment able to elicit a protective response against a lethal challenge with S. pyogenes. The strongest cellular responses and the highest levels of antigen-specific secretory immunoglobulin A (IgA) were detected in mice immunized with the fibronectin binding region of SfbI. In contrast, animals vaccinated with a polypeptide spanning the aromatic and proline-rich regions showed the highest titers and fastest IgG response in serum. Vaccination with either SfbI without a membrane anchor and signal peptide or a polypeptide encompassing its fibronectin-binding regions resulted in efficient protection against heterologous challenge (60% and 80%, respectively), whereas the use of a polypeptide lacking this region conferred marginal protection (10%) with respect to the control group (0%). These results demonstrate that the fibronectin-binding region of SfbI is a promising candidate antigen for developing anti-S. pyogenes vaccines. PMID- 11119567 TI - Alphavirus nucleocapsid protein contains a putative coiled coil alpha-helix important for core assembly. AB - The alphavirus nucleocapsid core is formed through the energetic contributions of multiple noncovalent interactions mediated by the capsid protein. This protein consists of a poorly conserved N-terminal region of unknown function and a C terminal conserved autoprotease domain with a major role in virion formation. In this study, an 18-amino-acid conserved region, predicted to fold into an alpha helix (helix I) and embedded in a low-complexity sequence enriched with basic and Pro residues, has been identified in the N-terminal region of the alphavirus capsid proteins. In Sindbis virus, helix I spans residues 38 to 55 and contains three conserved leucine residues, L38, L45, and L52, conforming to the heptad amino acid organization evident in leucine zipper proteins. Helix I consists of an N-terminally truncated heptad and two complete heptad repeats with beta branched residues and conserved leucine residues occupying the a and d positions of the helix, respectively. Complete or partial deletion of helix I, or single site substitutions at the conserved leucine residues (L45 and L52), caused a significant decrease in virus replication. The mutant viruses were more sensitive to elevated temperature than wild-type virus. These mutant viruses also failed to accumulate cores in the cytoplasm of infected cells, although they did not have defects in protein translation or processing. Analysis of these mutants using an in vitro assembly system indicated that the majority were defective in core particle assembly. Furthermore, mutant proteins showed a trans-dominant negative phenotype in in vitro assembly reactions involving mutant and wild-type proteins. We propose that helix I plays a central role in the assembly of nucleocapsid cores through coiled coil interactions. These interactions may stabilize subviral intermediates formed through the interactions of the C-terminal domain of the capsid protein and the genomic RNA and contribute to the stability of the virion. PMID- 11119568 TI - Vaccinia virus vectors with an inactivated gamma interferon receptor homolog gene (B8R) are attenuated In vivo without a concomitant reduction in immunogenicity. AB - The vaccinia virus (VV) B8R gene encodes a secreted protein with homology to the gamma interferon (IFN-gamma) receptor. In vitro, the B8R protein binds to and neutralizes the antiviral activity of several species of IFN-gamma, including human and rat IFN-gamma; it does not, however, bind significantly to murine IFN gamma. Here we report on the construction and characterization of recombinant VVs (rVVs) lacking the B8R gene. While the deletion of this gene had no effect on virus replication in vitro, rVVs lacking the B8R gene were attenuated for mice. There was a significant decrease in weight loss and mortality in normal mice, and nude mice survived significantly longer than did controls inoculated with parental virus. This is a surprising result considering the minimal binding of the B8R protein to murine IFN-gamma and its failure to block the antiviral activity of this cytokine in vitro. Such reduction in virulence could not be determined in rats, since they are considerably more resistant to VV infection than are mice. Finally, deletion of the B8R gene had no detectable effects on humoral immune responses. Mice and rats vaccinated with the rVVs showed identical humoral responses to both homologous and heterologous genes expressed by VV. This study demonstrates that the deletion of the VV B8R gene leads to enhanced safety without a concomitant reduction in immunogenicity. PMID- 11119569 TI - Natural 2',5'-phosphodiester bonds found at the ligation sites of peach latent mosaic viroid. AB - Peach latent mosaic viroid (PLMVd) is a circular RNA pathogen that replicates in a DNA-independent fashion via a rolling circle mechanism. PLMVd has been shown to self-ligate in vitro primarily via the formation of 2',5'-phosphodiester bonds; however, in vivo the occurrence and necessity of this nonenzymatic mechanism are not evident. Here, we unequivocally report the presence of 2', 5'-phosphodiester bonds at the ligation site of circular PLMVd strands isolated from infected peach leaves. These bonds serve to close the linear conformers (i.e., intermediates), yielding circular ones. Furthermore, these bonds are shown to stabilize the replicational circular templates, resulting in a significant advantage in terms of viroid viability. Although the mechanism responsible for the formation of these 2',5'-phosphodiester bonds remains to be elucidated, a hypothesis describing in vivo nonenzymatic self-ligation is proposed. Most significantly, our results clearly show that 2',5'-phosphodiester bonds are still present in nature and that they are of biological importance. PMID- 11119570 TI - Dominance of virus over host factors in cross-species activation of human cytomegalovirus early gene expression. AB - Human cytomegalovirus (HCMV) exhibits a highly restricted host range. In this study, we sought to examine the relative significance of host and viral factors in activating early gene expression of the HCMV UL54 (DNA polymerase) promoter in murine cells. Appropriate activation of the UL54 promoter at early times is essential for viral DNA replication. To study how the HCMV UL54 promoter is activated in murine cells, a transgenesis system based on yeast artificial chromosomes (YACs) was established for HCMV. A 178-kb YAC, containing a subgenomic fragment of HCMV encompassing the majority of the unique long (UL) region, was constructed by homologous recombination in yeast. This HCMV YAC backbone is defective for viral growth and lacks the major immediate-early (IE) gene region, thus permitting the analysis of essential cis-acting sequences when complemented in trans. To quantitatively measure the level of gene expression, we generated HCMV YACs containing a luciferase reporter gene inserted downstream of either the UL54 promoter or, as a control for late gene expression, the UL86 promoter, which directs expression of the major capsid protein. To determine the early gene activation pathway, point mutations were introduced into the inverted repeat 1 (IR1) element of the UL54 promoter of the HCMV YAC. In the transgenesis experiments, HCMV YACs and derivatives generated in yeast were introduced into NIH 3T3 murine cells by polyethylene glycol-mediated fusion. We found that infection of YAC, but not plasmid, transgenic lines with HCMV was sufficient to fully recapitulate the UL54 expression program at early times of infection, indicating the importance of remote regulatory elements in influencing regulation of the UL54 promoter. Moreover, YACs containing a mutant IR1 in the UL54 promoter led to reduced ( approximately 30-fold) reporter gene expression levels, indicating that HCMV major IE gene activation of the UL54 promoter is fully permissive in murine cells. In comparison with HCMV, infection of YAC transgenic NIH 3T3 lines with murine cytomegalovirus (MCMV) resulted in lower (more than one order of magnitude) efficiency in activating UL54 early gene expression. MCMV is therefore not able to fully activate HCMV early gene expression, indicating the significance of virus over host determinants in the cross-species activation of key early gene promoters. Finally, these studies show that YAC transgenesis can be a useful tool in functional analysis of viral proteins and control of gene expression for large viral genomes. PMID- 11119571 TI - RNA sequences involved in transcriptional termination of respiratory syncytial virus. AB - RNA signals at the ends of the genes of respiratory syncytial (RS) virus direct polyadenylation and termination of viral transcription. These gene ends contain two conserved regions, a pentanucleotide and a tract of uridylate (U) residues, separated by an A/U-rich central region that is less well conserved. The U tract is thought to be the template for polyadenylation of viral mRNAs by reiterative transcription. The cis-acting requirements for termination were investigated by mutagenesis of the matrix (M) gene end (3'-UCAAUUAUUUUUU-5') in a dicistronic RNA replicon. Termination efficiencies were quantitated by intracellular metabolic labeling of monocistronic mRNAs and the dicistronic readthrough RNAs that result when termination fails to occur. All three regions of the gene end were necessary for termination. Mutation of each of the first 8 nucleotides of the M gene end to all other nucleotides showed that nucleotides 2 to 6 were important for termination and intolerant of change, whereas nucleotides 1 and 7 were tolerant of change. At position 8, A or U allowed termination, but G or C did not. Both the length and the position of the U tract were important for termination. U residues at positions 9 to 12 were necessary, while additional U residues at position 8, and especially position 13, enhanced termination efficiency. Altering the length of the central region abolished termination, suggesting that the position of the U tract with respect to the 3'-UCAAU-5' sequence was critical. The termination efficiencies of each of the 10 genes of RS virus are different. Since transcription is obligatorily sequential and termination of each gene is required for transcription of the next gene downstream, these differences may contribute to gene regulation. In agreement with our data, the naturally occurring gene ends of RS virus that terminate inefficiently have short U tracts or other sequence features that correlated with decreased termination when similar mutations were analyzed in RNA replicons. PMID- 11119572 TI - Pseudopackaging of adenovirus type 5 genomes into capsids containing the hexon proteins of adenovirus serotypes B, D, or E. AB - Adenoviruses (Ad) show promise as a vector system for gene delivery in vivo. However, a major challenge in the development of Ad vectors is the circumvention of the host immune responses to Ad infection, including both the host cytotoxic T cell response and the humoral response resulting in neutralizing antibodies. One method to circumvent the effect of neutralizing antibodies against an Ad vector is to use different Ad serotypes to deliver the transgene of interest. This approach has been demonstrated with Ad genomes of highly related members of subgroup C. However, it is not known whether an Ad5-based vector DNA molecule can be packaged into capsids of evolutionarily more divergent adenoviruses. The aim of these studies was to determine if capsids containing hexon proteins from other Ad subgroups could package the Ad5 genome. A genetic approach utilizing an Ad5 temperature-sensitive (ts) mutant with a mutation in the hexon protein was used. When grown at the nonpermissive temperature, Ad5 ts147 replicates normally, providing a source of Ad5 DNA for virus assembly, but does not produce virus particles due to the hexon protein mutation. Coinfection of Ad5 ts147 with a wild type virus of other Ad serotypes (Ad3, Ad4, or Ad9), which supply functional hexon proteins, resulted in the pseudopackaging of the Ad5 DNA genome. Furthermore, the pseudopackaged Ad5 DNA virions obtained in the coinfections were infectious. Therefore, switching hexons did not impair the infectivity or uncoating process of the pseudopackaged virion. Since hexon protein is a major antigenic determinant of the Ad capsid, this approach may prove useful to reduce the antigenicity of therapeutic Ad vectors and allow repeated vector administration. PMID- 11119573 TI - CD4(+) T cells and gamma interferon in the long-term control of persistent friend retrovirus infection. AB - We have used the Friend virus model to determine the basic mechanisms by which the immune system can control persistent retroviral infections. Previously we showed that CD4(+) T cells play an essential role in keeping persistent retrovirus in check. The present in vitro experiments with a Friend virus specific CD4(+) T-cell clone revealed that these cells produce gamma interferon (IFN-gamma), which acts with two distinct mechanisms of antiviral activity. First, IFN-gamma had a direct inhibitory effect on virus production. This inhibitory effect was noncytolytic and, interestingly, was not associated with decreased cell surface expression of viral antigens. The second mechanism of IFN gamma-mediated antiviral activity was an enhancement of CD4(+) T-cell-mediated cytolytic activity. We also found an in vivo role for IFN-gamma in the control of persistent Friend virus infections. Neutralization of IFN-gamma in persistently infected mice resulted in significantly increased levels of virus in the spleen, and a significant percentage of IFN-gamma-deficient mice were unable to maintain long-term control over Friend virus infections. PMID- 11119574 TI - Roles of disulfide linkage and calcium ion-mediated interactions in assembly and disassembly of virus-like particles composed of simian virus 40 VP1 capsid protein. AB - The simian virus 40 capsid is composed of 72 pentamers of VP1 protein. Although the capsid is known to dissociate to pentamers in vitro following simultaneous treatment with reducing and chelating agents, the functional roles of disulfide linkage and calcium ion-mediated interactions are not clear. To elucidate the roles of these interactions, we introduced amino acid substitutions in VP1 at cysteine residues and at residues involved in calcium binding. We expressed the mutant proteins in a baculovirus system and analyzed both their assembly into virus-like particles (VLPs) in insect cells and the disassembly of those VLPs in vitro. We found that disulfide linkages at both Cys-9 and Cys-104 conferred resistance to proteinase K digestion on VLPs, although neither linkage was essential for the formation of VLPs in insect cells. In particular, reduction of the disulfide linkage at Cys-9 was found to be critical for VLP dissociation to VP1 pentamers in the absence of calcium ions, indicating that disulfide linkage at Cys-9 prevents VLP dissociation, probably by increasing the stability of calcium ion binding. We found that amino acid substitutions at carboxy-terminal calcium ion binding sites (Glu-329, Glu-330, and Asp-345) resulted in the frequent formation of unusual tubular particles as well as VLPs in insect cells, indicating that these residues affect the accuracy of capsid assembly. In addition, unexpectedly, amino acid substitutions at any of the calcium ion binding sites tested, especially at Glu-157, resulted in increased stability of VLPs in the absence of calcium ions in vitro. These results suggest that appropriate affinities of calcium ion binding are responsible for both assembly and disassembly of the capsid. PMID- 11119575 TI - Evaluation of cytotoxic T-lymphocyte responses in human and nonhuman primate subjects infected with human immunodeficiency virus type 1 or simian/human immunodeficiency virus. AB - Cytotoxic T-lymphocyte (CTL) responses have been implicated as playing an important role in control of human immunodeficiency virus (HIV) infection. However, it is technically difficult to demonstrate CTL responses consistently in nonhuman primate and human subjects using traditional cytotoxicity assay methods. In this study, we systematically evaluated culture conditions that may affect the proliferation and expansion of CTL effector cells and presented a sensitive method for detection of cytotoxicity responses with bulk CTL cultures. We confirmed the sensitivity and specificity of this method by demonstration of vigorous CTL responses in a simian-HIV (SHIV)-infected rhesus macaque. The expansion of epitope-specific CTL effector cells was also measured quantitatively by CTL epitope-major histocompatibility complex tetramer complex staining. In addition, two new T-cell determinants in the SIV gag region are identified. Last, we showed the utility of this method for studying CTL responses in chimpanzee and human subjects. PMID- 11119576 TI - Protective T-cell-based immunity induced in neonatal mice by a single replicative cycle of herpes simplex virus. AB - Newborns are very susceptible to infections because their immune systems are not fully developed and react to antigen exposure preferentially with unresponsiveness. UV-inactivated herpes simplex virus type 1 (HSV-1) represents such an antigen and does not induce an immune response in neonates. In contrast, protective T cells were primed in newborn mice by a single replicative cycle of DISC HSV-1 given once within 24 h of birth. Each of the HSV-1-primed CD4(+) or CD8(+) T cells induced in wild-type or interferon-deficient mice conferred resistance to naive animals exposed to a lethal virus challenge. Inactivated HSV 1, injected at variable doses up to 10(4) times that of DISC HSV-1, was ineffective in inducing any detectable immune responses in neonates. Thus, the capacity of HSV-1 to replicate once, but not the number of virus particles per se, was decisive in inducing protective T-cell-associated immunity in newborn mice. PMID- 11119577 TI - Amino acids of Epstein-Barr virus nuclear antigen 3A essential for repression of Jkappa-mediated transcription and their evolutionary conservation. AB - Epstein-Barr virus (EBV) nuclear antigen 3A (EBNA-3A) is essential for virus mediated immortalization of B lymphocytes in vitro and is believed to regulate transcription of cellular and/or viral genes. One known mechanism of regulation is through its interaction with the cellular transcription factor Jkappa. This interaction downregulates transcription mediated by EBNA-2 and Jkappa. To identify the amino acids that play a role in this interaction, we have generated mutant EBNA-3A proteins. A mutant EBNA-3A protein in which alanine residues were substituted for amino acids 199, 200, and 202 no longer downregulated transcription. Surprisingly, this mutant protein remained able to coimmunoprecipitate with Jkappa. Using a reporter gene assay based on the recruitment of Jkappa by various regions spanning EBNA-3A, we have shown that this mutation abolished binding of Jkappa to the N-proximal region (amino acids 125 to 222) and that no other region of EBNA-3A alone was sufficient to mediate an association with Jkappa. To determine the biological significance of the interaction of EBNA-3A with Jkappa, we have studied its conservation in the simian lymphocryptovirus herpesvirus papio (HVP) by cloning HVP-3A, the homolog of EBNA-3A encoded by this virus. This 903-amino-acid protein exhibited 37% identity with its EBV counterpart, mainly within the amino-terminal half. HVP-3A also interacted with Jkappa through a region located between amino acids 127 and 223 and also repressed transcription mediated through EBNA-2 and Jkappa. The evolutionary conservation of this function, in proteins that have otherwise significantly diverged, argues strongly for an important biological role in virus mediated immortalization of B lymphocytes. PMID- 11119578 TI - Interaction between the open reading frame III product and the coat protein is required for transmission of cauliflower mosaic virus by aphids. AB - Transmission of cauliflower mosaic virus (CaMV) by aphids requires two viral nonstructural proteins, the open reading frame (ORF) II and ORF III products (P2 and P3). An interaction between a C-terminal domain of P2 and an N-terminal domain of P3 is essential for transmission. Purified particles of CaMV are efficiently transmitted only if aphids, previously fed a P2-containing solution, are allowed to acquire a preincubated mixture of P3 and virions in a second feed, thus suggesting a direct interaction between P3 and coat protein. Herein we demonstrate that P3 directly interacts with purified viral particles and unassembled coat protein without the need for any other factor and that P3 mediates the association of P2 with purified virus particles. The interaction domain of P3 is located in its C-terminal half, downstream of the P3-P2 interaction domain but overlapping a region which binds nucleic acids. Mutagenesis of P3 which interferes with the interaction between P3 and virions is correlated with the loss of transmission by aphids. Taken together, our results demonstrate that P3 plays a crucial role in the formation of the CaMV transmissible complex by serving as a bridge between P2 and virus particles. PMID- 11119579 TI - Molecular analysis of the abnormal prion protein during coinfection of mice by bovine spongiform encephalopathy and a scrapie agent. AB - Molecular features of the proteinase K-resistant prion protein (PrP res) may discriminate among prion strains, and a specific signature could be found during infection by the infectious agent causing bovine spongiform encephalopathy (BSE). To investigate the molecular basis of BSE adaptation and selection, we established a model of coinfection of mice by both BSE and a sheep scrapie strain (C506M3). We now show that the PrP res features in these mice, characterized by glycoform ratios and electrophoretic mobilities, may be undistinguishable from those found in mice infected with scrapie only, including when mice were inoculated by both strains at the same time and by the same intracerebral inoculation route. Western blot analysis using different antibodies against sequences near the putative N-terminal end of PrP res also demonstrated differences in the main proteinase K cleavage sites between mice showing either the BSE or scrapie PrP res profile. These results, which may be linked to higher levels of PrP res associated with infection by scrapie, were similar following a challenge by a higher dose of the BSE agent during coinfection by both strains intracerebrally. Whereas PrP res extraction methods used allowed us to distinguish type 1 and type 2 PrP res, differing, like BSE and scrapie, by their electrophoretic mobilities, in the same brain region of some patients with Creutzfeldt-Jakob disease, analysis of in vitro mixtures of BSE and scrapie brain homogenates did not allow us to distinguish BSE and scrapie PrP res. These results suggest that the BSE agent, the origin of which remains unknown so far but which may have arisen from a sheep scrapie agent, may be hidden by a scrapie strain during attempts to identify it by molecular studies and following transmission of the disease in mice. PMID- 11119580 TI - Alteration of zinc-binding residues of simian immunodeficiency virus p8(NC) results in subtle differences in gag processing and virion maturation associated with degradative loss of mutant NC. AB - In all retroviruses analyzed to date (except for the spumaretroviruses), the Zn(2+)-coordinating residues of nucleocapsid (NC) perform or assist in crucial reactions necessary to complete the retrovirus life cycle. Six replication defective mutations have been engineered in the two NC Zn(2+) fingers (ZFs) of simian immunodeficiency virus [SIV(Mne)] that change or delete specific Zn(2+) interacting Cys residues and were studied by using electron microscopy, reversed phase high-performance liquid chromatography, immunoblotting, and RNA quantification. We focused on phenotypes of produced particles, specifically morphology, Gag polyprotein processing, and genomic RNA packaging. Phenotypes were similar among viruses containing a point or deletion mutation involving the same ZF. Mutations in the proximal ZF (ZF1) resulted in near-normal Gag processing and full-length genomic RNA incorporation and were most similar to wild-type (WT) virions with electron-dense, conical cores. Mutation of the distal ZF, as well as point mutations in both ZFs, resulted in more unprocessed Gag proteins than a deletion or point mutation in ZF1, with an approximate 30% reduction in levels of full-length genomic RNA in virions. These mutant virions contained condensed cores; however, the cores typically appeared less electron dense and more rod shaped than WT virions. Surprisingly, deletion of both ZFs, including the basic linker region between the ZFs, resulted in the most efficient Gag processing. However, genomic RNA packaging was approximately 10% of WT levels, and those particles produced were highly abnormal with respect to size and core morphology. Surprisingly, all NC mutations analyzed demonstrated a significant loss of processed NC in virus particles, suggesting that Zn(2+) coordinated NC is protected from excessive proteolytic cleavage. Together, these results indicate that Zn(2+) coordination is important for correct Gag precursor processing and NC protein stability. Additionally, SIV particle morphology appears to be the result of proper and complete Gag processing and relies less on full-length genomic RNA incorporation, as dictated by the Zn(2+) coordination in the ZFs of the NC protein. PMID- 11119581 TI - cis-acting sequences required for coronavirus infectious bronchitis virus defective-RNA replication and packaging. AB - The parts of the RNA genome of infectious bronchitis virus (IBV) required for replication and packaging of the RNA were investigated using deletion mutagenesis of a defective RNA (D-RNA) CD-61 (6.1 kb) containing a chloramphenicol acetyltransferase reporter gene. A D-RNA with the first 544, but not as few as 338, nucleotides (nt) of the 5' terminus was replicated; the 5' untranslated region (UTR) comprises 528 nt. Region I of the 3' UTR, adjacent to the nucleocapsid protein gene, comprised 212 nt and could be removed without impairment of replication or packaging of D-RNAs. A D-RNA with the final 338 nt, including the 293 nt in the highly conserved region II of the 3' UTR, was replicated. Thus, the 5'-terminal 544 nt and 3'-terminal 338 nt contained the necessary signals for RNA replication. Phylogenetic analysis of 19 strains of IBV and 3 strains of turkey coronavirus predicted a conserved stem-loop structure at the 5' end of region II of the 3' UTR. Removal of the predicted stem-loop structure abolished replication of the D-RNAs. D-RNAs in which replicase gene 1b derived sequences had been removed or replaced with all the downstream genes were replicated well but were rescued poorly, suggesting inefficient packaging. However, no specific part of the 1b gene was required for efficient packaging. PMID- 11119582 TI - Mutagenic analysis of the 5' arm of the influenza A virus virion RNA promoter defines the sequence requirements for endonuclease activity. AB - Short synthetic influenza virus-like RNAs derived from influenza virus promoter sequences were examined for their ability to stimulate the endonuclease activity of recombinant influenza virus polymerase complexes in vitro, an activity that is required for the cap-snatching activity of primers from host pre-mRNA. An extensive set of point mutants of the 5' arm of the influenza A virus viral RNA (vRNA) was constructed to determine the cis-acting elements which influenced endonuclease activity. Activity was found to be dependent on three features of the conserved vRNA termini: (i) the presence of the 5' hairpin loop structure, (ii) the identity of residues at positions 5 and 10 bases from the 5' terminus, and (iii) the presence of base pair interactions between the 5' and 3' segment ends. Further experiments discounted a role for the vRNA U track in endonuclease activation. This study represents the first mutagenic analysis of the influenza virus promoter with regard to endonuclease activity. PMID- 11119583 TI - Envelope protein-mediated down-regulation of hepatitis B virus receptor in infected hepatocytes. AB - Entry of duck hepatitis B virus (DHBV) is initiated by specific interaction of its large envelope protein (L) with a cellular entry receptor, recently identified as carboxypeptidase D (CPD; historically gp180). In this report, we present evidence demonstrating that this receptor is down-regulated as a result of DHBV infection: (i) receptor levels determined by Western blot were much reduced in DHBV-infected duck livers and undetectable by immunostaining in infected cultured hepatocytes; (ii) results from metabolic labeling experiments indicate enhanced receptor protein turnover; (iii) the kinetics of receptor loss from newly infected cells correlated with the accumulation of newly synthesized viral protein; (iv) expression of DHBV L protein, transduced from a recombinant adenovirus, was sufficient to eliminate gp180/CPD from the Golgi compartment, its normal predominant location; (v) gp180/CPD remained absent from the Golgi compartment in infected hepatocytes, even after overexpression from a recombinant adenovirus, while residual amounts subsequently became detectable in a perinuclear compartment, containing DHBV L protein; (vi) expression of DHBV L protein in a HepG2 cell line, stably expressing gp180/CPD, leads to incomplete receptor maturation and induces its degradation. Taken together, these data are consistent with a model in which the virus receptor interacts early in the biosynthetic pathway with the viral L protein, leading to its retention in a pre Golgi compartment and to subsequent degradation, thus preventing receptor interference with the export of DHBV via the secretory pathway which it shares with its receptor. Accordingly, and analogously with receptor down-regulation in retroviral systems, DHBV receptor down-modulation may account for the much reduced efficiency of DHBV superinfection of preinfected hepatocytes. PMID- 11119584 TI - Gps2, a protein partner for human papillomavirus E6 proteins. AB - We have used the yeast two-hybrid system to screen a cDNA library prepared from normal human epidermal keratinocytes and identified protein partners for human papilloma virus (HPV) E6 proteins. A clone that encoded Gps2 interacted with E6 proteins from HPVs of high and low oncogenic risk. The specificity of these reactions was verified and the regions of E6 that were required for interaction were mapped. Steady-state and pulse-chase analyses of cells cotransfected with DNAs expressing E6 from either HPV6 or HPV18 and Gps2 demonstrated that the E6 proteins induced the degradation of Gps2 in vivo but not in vitro. Gps2 exhibited transcriptional activation activity, and high-risk E6 suppressed this activity. PMID- 11119585 TI - Duck hepatitis B virus expresses a regulatory HBx-like protein from a hidden open reading frame. AB - Duck hepatitis B viruses (DHBV), unlike mammalian hepadnaviruses, are thought to lack X genes, which encode transcription-regulatory proteins believed to contribute to the development of hepatocellular carcinoma. A lack of association of chronic DHBV infection with hepatocellular carcinoma development supports this belief. Here, we demonstrate that DHBV genomes have a hidden open reading frame from which a transcription-regulatory protein, designated DHBx, is expressed both in vitro and in vivo. We show that DHBx enhances neither viral protein expression, intracellular DNA synthesis, nor virion production when assayed in the full-length genome context in LMH cells. However, similar to mammalian hepadnavirus X proteins, DHBx activates cellular and viral promoters via the Raf mitogen-activated protein kinase signaling pathway and localizes primarily in the cytoplasm. The functional similarities as well as the weak sequence homologies of DHBx and the X proteins of mammalian hepadnaviruses strongly suggest a common ancestry of ortho- and avihepadnavirus X genes. In addition, our data disclose similar intracellular localization and transcription regulatory functions of the corresponding proteins, raise new questions as to their presumed role in hepatocarcinogenesis, and imply unique opportunities for deciphering of their still-enigmatic in vivo functions. PMID- 11119586 TI - Localization of the gD-binding region of the human herpes simplex virus receptor, HveA. AB - During virus entry, herpes simplex virus (HSV) glycoprotein D (gD) binds to one of several human cellular receptors. One of these, herpesvirus entry mediator A (HveA), is a member of the tumor necrosis factor receptor (TNFR) superfamily, and its ectodomain contains four characteristic cysteine-rich pseudorepeat (CRP) elements. We previously showed that gD binds the ectodomain of HveA expressed as a truncated, soluble protein [HveA(200t)]. To localize the gD-binding domain of HveA, we expressed three additional soluble forms of HveA consisting of the first CRP [HveA(76t)], the second CRP [HveA(77-120t)], or the first and second CRPs [HveA(120t)]. Biosensor and enzyme-linked immunosorbent assay studies showed that gD bound to HveA(120t) and HveA(200t) with the same affinity. However, gD did not bind to HveA(76t) or HveA(77-120t). Furthermore, HveA(200t) and HveA(120t), but not HveA(76t) or HveA(77-120t), blocked herpes simplex virus (HSV) entry into CHO cells expressing HveA. We also generated six monoclonal antibodies (MAbs) against HveA(200t). MAbs CW1, -2, and -4 bound linear epitopes within the second CRP, while CW7 and -8 bound linear epitopes within the third or fourth CRPs. None of these MAbs blocked the binding of gD to HveA. In contrast, MAb CW3 recognized a discontinuous epitope within the first CRP of HveA, blocked the binding of gD to HveA, and exhibited a limited ability to block virus entry into cells expressing HveA, suggesting that the first domain of HveA contains at least a portion of the gD binding site. The inability of gD to bind HveA(76t) suggests that additional amino acid residues of the gD binding site may reside within the second CRP. PMID- 11119588 TI - Reciprocal interactions between human T-lymphotropic virus type 1 and prostaglandins: implications for viral transmission. AB - Human T-lymphotropic virus type 1 (HTLV-1), the etiologic agent of adult T-cell leukemia/lymphoma, is transmitted through breast milk and seminal fluid, which are rich in prostaglandins (PGs). We demonstrate that PGE(2) upregulates the HTLV 1 long terminal repeat promoter through the protein kinase A pathway, induces replication of HTLV-1 in peripheral blood mononuclear cells (PBMC) derived from asymptomatic carriers, and enhances transmission of HTLV-1 to cord blood mononuclear cells (CBMC). Furthermore, HTLV-1 Tax transactivates a promoter for cyclooxygenase 2, a PG synthetase, and induces PGE(2) expression in PBMC or CBMC. Thus, HTLV-1 interacts with and benefits from PGs, constituents of its own vehicle for transmission. PMID- 11119587 TI - The human immunodeficiency virus type 1 gag gene encodes an internal ribosome entry site. AB - Several retroviruses have recently been shown to promote translation of their gag gene products by internal ribosome entry. In this report, we show that mRNAs containing the human immunodeficiency virus type 1 (HIV-1) gag open reading frame (ORF) exhibit internal ribosome entry site (IRES) activity that can promote translational initiation of Pr55(gag). Remarkably, this IRES activity is driven by sequences within the gag ORF itself and is not dependent on the native gag 5' untranslated region (UTR). This cap-independent mechanism for Pr55(gag) translation may help explain the high levels of translation of this protein in the face of major RNA structural barriers to scanning ribosomes found in the gag 5' UTR. The gag IRES activity described here also drives translation of a novel 40-kDa Gag isoform through translational initiation at an internal AUG codon found near the amino terminus of the Pr55(gag) capsid domain. Our findings suggest that this low-abundance Gag isoform may be important for wild-type replication of HIV-1 in cultured cells. The activities of the HIV-1 gag IRES may be an important feature of the HIV-1 life cycle and could serve as a novel target for antiretroviral therapeutic strategies. PMID- 11119589 TI - Use of intron-disrupted polyadenylation sites to enhance expression and safety of retroviral vectors. AB - Normal mRNA polyadenylation signals are composed of an AAUAAA motif and G/U box spaced 20 to 30 bp apart. If this spacing is increased further, then polyadenylation is disrupted. Previously it has been demonstrated that insertion of an intron will similarly disrupt this signal even though such introns are removed during a nuclear splicing reaction (X. Liu and J. Mertz, Nucleic Acids Res. 21:5256-5263, 1993). This observation has led to the suggestion that polyadenylation site selection is undertaken prior to intron excision. We now present results that both support and extend these observations and in doing so create a novel class of retroviral expression vector with improved qualities. We found that when an intron-disrupted polyadenylation signal is inserted within a retroviral expression vector, such a signal, although reformed in the producer cell, remains benign until transduction, where it is then preferentially used. Thus, we demonstrate that upon transduction these vectors now produce a majority of shortened subgenomic species and as a consequence have a reduced tendency for subsequent mobilization from transduced cells. In addition, we demonstrate that the use of this internal signal leads to enhanced expression from such vectors and that this is achieved without any loss in titer. Therefore, split polyadenylation signals confer enhanced performance and improved safety upon retroviral expression vectors into which they are inserted. Such split signals may prove useful for the future optimization of retroviral vectors in gene therapy. PMID- 11119590 TI - Mutations that affect dimer formation and helicase activity of the hepatitis C virus helicase. AB - Interaction between viral proteins is necessary for viral replication and viral particle assembly. We used the yeast two-hybrid assay to identify interactions among all the mature proteins of the hepatitis C virus. The interaction between NS3 and NS3 was one of the strongest viral protein-protein interactions detected. The minimal region required for this interaction was mapped to a specific subdomain of 174 amino acids in the N terminus of the helicase region. Random mutations in the minimal region were generated by PCR, and mutants that failed to interact with a wild-type minimal fragment were isolated using the yeast two hybrid assay as a screen. Three of these mutations resulted in a reduction or a loss of interaction between helicases. Analytical gel filtration showed that in the presence of an oligonucleotide, wild-type helicases form dimers whereas the mutants remain mostly monomeric. All three mutants were partially or almost inactive when assayed for helicase activity in vitro. Mixing a mutant helicase (Y267S) with wild-type helicase did not dramatically affect helicase activity. These data indicate that dimerization of the helicase is important for helicase activity. The mutations that reduce self-association of the helicase may define the key residues involved in NS3-NS3 dimerization. PMID- 11119592 TI - CrmE, a novel soluble tumor necrosis factor receptor encoded by poxviruses. AB - Cytokines and chemokines play a critical role in both the innate and acquired immune responses and constitute prime targets for pathogen sabotage. Molecular mimicry of cytokines and cytokine receptors is a mechanism encoded by large DNA viruses to modulate the host immune response. Three tumor necrosis factor receptors (TNFRs) have been identified in the poxvirus cowpox virus. Here we report the identification and characterization of a fourth distinct soluble TNFR, named cytokine response modifier E (CrmE), encoded by cowpox virus. The crmE gene has been sequenced in strains of the orthopoxviruses cowpox virus, ectromelia virus, and camelpox virus, and was found to be active in cowpox virus. crmE is expressed as a secreted 18-kDa protein with TNF binding activity. CrmE was produced in the baculovirus and vaccinia virus expression systems and was shown to bind human, mouse, and rat TNF, but not human lymphotoxin alpha, conjugates of lymphotoxins alpha and beta, or seven other ligands of the TNF superfamily. However, CrmE protects cells only from the cytolytic activity of human TNF. CrmE is a new member of the TNFR superfamily which is expressed as a soluble molecule that blocks the binding of TNF to high-affinity TNFRs on the cell surface. The remarkable finding of a fourth poxvirus-encoded TNFR suggests that modulation of TNF activity is complex and represents a novel viral immune evasion mechanism. PMID- 11119591 TI - Role of NF-kappaB and myc proteins in apoptosis induced by hepatitis B virus HBx protein. AB - Chronic infection with hepatitis B virus (HBV) promotes a high level of liver disease and cancer in humans. The HBV HBx gene encodes a small regulatory protein that is essential for viral replication and is suspected to play a role in viral pathogenesis. HBx stimulates cytoplasmic signal transduction pathways, moderately stimulates a number of transcription factors, including several nuclear factors, and in certain settings sensitizes cells to apoptosis by proapoptotic stimuli, including tumor necrosis factor alpha (TNF-alpha) and etopocide. Paradoxically, HBx activates members of the NF-kappaB transcription factor family, some of which are antiapoptotic in function. HBx induces expression of Myc protein family members in certain settings, and Myc can sensitize cells to killing by TNF-alpha. We therefore examined the roles of NF-kappaB, c-Myc, and TNF-alpha in apoptotic killing of cells by HBx. RelA/NF-kappaB is shown to be induced by HBx and to suppress HBx-mediated apoptosis. HBx also induces c-Rel/NF-kappaB, which can promote apoptotic cell death in some contexts or block it in others. Induction of c-Rel by HBx was found to inhibit its ability to directly mediate apoptotic killing of cells. Thus, HBx induction of NF-kappaB family members masks its ability to directly mediate apoptosis, whereas ablation of NF-kappaB reveals it. Investigation of the role of Myc protein demonstrates that overexpression of Myc is essential for acute sensitization of cells to killing by HBx plus TNF-alpha. This study therefore defines a specific set of parameters which must be met for HBx to possibly contribute to HBV pathogenesis. PMID- 11119593 TI - Transient mobilization of human immunodeficiency virus (HIV)-specific CD4 T helper cells fails to control virus rebounds during intermittent antiretroviral therapy in chronic HIV type 1 infection. AB - Immune control of human immunodeficiency virus (HIV) is not restored by highly active antiretroviral therapies (HAART) during chronic infection. We examined the capacity of repeated structured therapeutic interruptions (STI) to restore HIV specific CD4 and CD8 T-cell responses that controlled virus production. Eleven STI (median duration, 7 days; ranges, 4 to 24 days) were performed in three chronically HIV-infected patients with CD4 counts above 400/mm(3) and less than 200 HIV RNA copies/ml after 18 to 21 months of HAART; treatment resumed after 1 week or when virus became detectable. HIV-specific T-cell responses were analyzed by proliferation, gamma interferon (IFN-gamma) production, and enzyme-linked immunospot assays. Seven virus rebounds were observed (median, 4,712 HIV-1 RNA copies/ml) with a median of 7 days during which CD4 and CD8 counts did not significantly change. After treatment resumed, the viral load returned below 200 copies/ml within 3 weeks. Significant CD4 T-cell proliferation and IFN-gamma production against HIV p24 appeared simultaneously with or even before the virus rebounds in all patients. These CD4 responses lasted for less than 3 weeks and disappeared before therapeutic control of the virus had occurred. Increases in the numbers of HIV-specific CD8 T cells were delayed compared to changes in HIV specific CD4 T-cell responses. No delay or increase in virus doubling time was observed after repeated STI. Iterative reexposure to HIV during short STI in chronically infected patients only transiently mobilized HIV-specific CD4 T1 helper cells, which might be rapidly altered by virus replication. Such kinetics might explain the failure at delaying subsequent virus rebounds and raises concerns about strategies based on STI to restore durable HIV-specific T-cell responses in chronic HIV infection. PMID- 11119594 TI - Viral DNA synthesis defects in assembly-competent Rous sarcoma virus CA mutants. AB - The major structural protein of the retroviral core (CA) contains a conserved sequence motif shared with the CA-like proteins of distantly related transposable elements. The function of this major region of homology (MHR) has not been defined, in part due to the baffling array of phenotypes in mutants of several viruses and the yeast TY3. This report describes new mutations in the CA protein of Rous sarcoma virus (RSV) that were designed to test whether these different phenotypes might indicate distinct functional subdomains in the MHR. A comparison of 25 substitutions at 10 positions in the RSV conserved motif argues against this possibility. Most of the replacements destroyed virus infectivity, although either of two lethal phenotypes was obtained depending on the residue introduced. At most of the positions, one or more replacements (generally the more conservative substitutions) caused a severe replication defect without having any obvious effects on virus assembly, budding, Gag-Pol and genome incorporation, or protein processing. The mutant particles exhibited a defect in endogenous viral DNA synthesis and showed increased sensitivity of the core proteins to detergent, indicating that the mutations interfere with the formation and/or activity of the virion core. The distribution of these mutations across the MHR, with no evidence of clustering, suggests that the entire region is important for a critical postbudding function. In contrast, a second class of lethal substitutions (those that destroyed virus assembly and release) consists of alterations that are expected to cause severe effects on protein structure by disruption either of the hydrophobic core of the CA carboxyl-terminal domain or of the hydrogen bond network that stabilizes the domain. We suggest that this duality of phenotypes is consistent with a role for the MHR in the maturation process that links the two parts of the life cycle. PMID- 11119595 TI - Determination of coreceptor usage of human immunodeficiency virus type 1 from patient plasma samples by using a recombinant phenotypic assay. AB - We developed a recombinant virus technique to determine the coreceptor usage of human immunodeficiency virus type 1 (HIV-1) from plasma samples, the source expected to represent the most actively replicating virus population in infected subjects. This method is not subject to selective bias associated with virus isolation in culture, a step required for conventional tropism determination procedures. The addition of a simple subcloning step allowed semiquantitative evaluation of virus populations with a different coreceptor (CCR5 or CXCR4) usage specificity present in each plasma sample. This procedure detected mixtures of CCR5- and CXCR4-exclusive virus populations as well as dualtropic viral variants, in variable proportions. Sequence analysis of dualtropic clones indicated that changes in the V3 loop are necessary for the use of CXCR4 as a coreceptor, but the overall context of the V1-V3 region is important to preserve the capacity to use CCR5. This convenient technique can greatly assist the study of virus evolution and compartmentalization in infected individuals. PMID- 11119596 TI - trans-acting inhibition of genomic RNA dimerization by Rous sarcoma virus matrix mutants. AB - The genomic RNA of retroviruses exists within the virion as a noncovalently linked dimer. Previously, we identified a mutant of the viral matrix (MA) protein of Rous sarcoma virus that disrupts viral RNA dimerization. This mutant, Myr1E, is modified at the N terminus of MA by the addition of 10 amino acids from the Src protein, resulting in the production of particles containing monomeric RNA. Dimerization is reestablished by a single amino acid substitution that abolishes myristylation (Myr1E-). To distinguish between cis and trans effects involving Myr1E, additional mutations were generated. In Myr1E.cc and Myr1E-.cc, different nucleotides were utilized to encode the same protein as Myr1E and Myr1E-, respectively. The alterations in RNA sequence did not change the properties of the viral mutants. Myr1E.ATG- was constructed so that translation began at the gag AUG, resulting in synthesis of the wild-type Gag protein but maintenance of the src RNA sequence. This mutant had normal infectivity and dimeric RNA, indicating that the src sequence did not prevent dimer formation. All of the src containing RNA sequences formed dimers in vitro. Examination of MA-green fluorescent protein fusion proteins revealed that the wild-type and mutant MA proteins Myr1E.ATG-, Myr1E-, and Myr1E-.cc had distinctly different patterns of subcellular localization compared with Myr1E and Myr1E.cc MA proteins. This finding suggests that proper localization of the MA protein may be required for RNA dimer formation and infectivity. Taken together, these results provide compelling evidence that the genomic RNA dimerization defect is due to a trans acting effect of the mutant MA proteins. PMID- 11119597 TI - Successful interference with cellular immune responses to immunogenic proteins encoded by recombinant viral vectors. AB - Vectors derived from the adeno-associated virus (AAV) have been successfully used for the long-term expression of therapeutic genes in animal models and patients. One of the major advantages of these vectors is the absence of deleterious immune responses following gene transfer. However, AAV vectors, when used in vaccination studies, can result in efficient humoral and cellular responses against the transgene product. It is therefore important to understand the factors which influence the establishment of these immune responses in order to design safe and efficient procedures for AAV-based gene therapies. We have compared T-cell activation against a strongly immunogenic protein, the influenza virus hemagglutinin (HA), which is synthesized in skeletal muscle following gene transfer with an adenovirus (Ad) or an AAV vector. In both cases, cellular immune responses resulted in the elimination of transduced muscle fibers within 4 weeks. However, the kinetics of CD4(+) T-cell activation were markedly delayed when AAV vectors were used. Upon recombinant Ad (rAd) gene transfer, T cells were activated both by direct transduction of dendritic cells and by cross presentation of the transgene product, while upon rAAV gene transfer T cells were only activated by the latter mechanism. These results suggested that activation of the immune system by the transgene product following rAAV-mediated gene transfer might be easier to control than that following rAd-mediated gene transfer. Therefore, we tested protocols aimed at interfering with either antigen presentation by blocking the CD40/CD40L pathway or with the T-cell response by inducing transgene-specific tolerance. Long-term expression of the AAV-HA was achieved in both cases, whereas immune responses against Ad-HA could not be prevented. These data clearly underline the importance of understanding the mechanisms by which vector-encoded proteins are recognized by the immune system in order to specifically interfere with them and to achieve safe and stable gene transfer in clinical trials. PMID- 11119598 TI - In vivo attenuation of simian immunodeficiency virus by disruption of a tyrosine dependent sorting signal in the envelope glycoprotein cytoplasmic tail. AB - Attenuated simian immunodeficiency viruses (SIVs) have been described that produce low levels of plasma virion RNA and exhibit a reduced capacity to cause disease. These viruses are particularly useful in identifying viral determinants of pathogenesis. In the present study, we show that mutation of a highly conserved tyrosine (Tyr)-containing motif (Yxxphi) in the envelope glycoprotein (Env) cytoplasmic tail (amino acids YRPV at positions 721 to 724) can profoundly reduce the in vivo pathogenicity of SIVmac239. This domain constitutes both a potent endocytosis signal that reduces Env expression on infected cells and a sorting signal that directs Env expression to the basolateral surface of polarized cells. Rhesus macaques were inoculated with SIVmac239 control or SIVmac239 containing either a Tyr-721-to-Ile mutation (SIVmac239Y/I) or a deletion of Tyr-721 and the preceding glycine (DeltaGY). To assess the in vivo replication competence, all viruses contained a stop codon in nef that has been shown to revert during in vivo but not in vitro replication. All three control animals developed high viral loads and disease. One of two animals that received SIVmac239Y/I and two of three animals that received SIVmac239DeltaGY remained healthy for up to 140 weeks with low to undetectable plasma viral RNA levels and normal CD4(+) T-cell percentages. These animals exhibited ongoing viral replication as determined by detection of viral sequences and culturing of mutant viruses from peripheral blood mononuclear cells and persistent anti-SIV antibody titers. In one animal that received SIVmac239Y/I, the Ile reverted to a Tyr and was associated with a high plasma RNA level and disease, while one animal that received SIVmac239DeltaGY also developed a high viral load that was associated with novel and possibly compensatory mutations in the TM cytoplasmic domain. In all control and experimental animals, the nef stop codon reverted to an open reading frame within the first 2 months of inoculation, indicating that the mutant viruses had replicated well enough to repair this mutation. These findings indicate that the Yxxphi signal plays an important role in SIV pathogenesis. Moreover, because mutations in this motif may attenuate SIV through mechanisms that are distinct from those caused by mutations in nef, this Tyr-based sorting signal represents a novel target for future models of SIV and human immunodeficiency virus attenuation that could be useful in new vaccine strategies. PMID- 11119599 TI - The E1 initiator recognizes multiple overlapping sites in the papillomavirus origin of DNA replication. AB - A common feature of replicator sequences from a variety of organisms is multiple binding sites for an initiator protein. By binding to the replicator, initiators mark the site and contribute to melting or distortion of the DNA. We have defined the recognition sequence for the papillomavirus E1 initiator and determined the arrangement of binding sites in the viral origin of replication. We show that E1 recognizes a hexanucleotide sequence which is present in overlapping arrays in virtually all papillomavirus replicators. Binding of the initiator to these sites would result in the formation of a closely packed array of E1 molecules that wrap around the double helix. PMID- 11119600 TI - The vaccinia virus A33R protein provides a chaperone function for viral membrane localization and tyrosine phosphorylation of the A36R protein. AB - The products of the A33R and A36R genes of vaccinia virus are incorporated into the membranes of intracellular enveloped virions (IEV). When extracts of cells that had been infected with vaccinia virus and labeled with H(3)(32)PO(4) were immunoprecipitated with antibodies against the A33R protein, two prominent bands were resolved. The moderately and more intensely labeled bands were identified as phosphorylated A33R and A36R proteins, respectively. The immunoprecipitated complex contained disulfide-bonded dimers of A33R protein that were noncovalently linked to A36R protein. Biochemical analysis indicated that the two proteins were phosphorylated predominantly on serine residues, with lesser amounts on threonines. The A36R protein was also phosphorylated on tyrosine, as determined by specific binding to an anti-phosphotyrosine antibody. Serine phosphorylation and A33R-A36R protein complex formation occurred even when virus assembly was blocked at an early stage with the drug rifampin. Tyrosine phosphorylation was selectively reduced in cells infected with F13L or A34R gene deletion mutants that were impaired in the membrane-wrapping step of IEV formation. In addition, tyrosine phosphorylation was specifically inhibited in cells infected with an A33R deletion mutant that still formed IEV. Immunofluorescence and immunoelectron microscopy indicated that in the absence of the A33R protein, the A36R protein was localized in Golgi membranes but not in IEV. In the absence of the A36R protein, however, the A33R protein still localized to IEV membranes. These studies together with others suggest that the A33R protein guides the A36R protein to the IEV membrane, where it subsequently becomes tyrosine phosphorylated as a signal for actin tail formation. PMID- 11119601 TI - Kinetics of hepadnavirus loss from the liver during inhibition of viral DNA synthesis. AB - Hepadnaviruses replicate by reverse transcription, which takes place in the cytoplasm of the infected hepatocyte. Viral RNAs, including the pregenome, are transcribed from a covalently closed circular (ccc) viral DNA that is found in the nucleus. Inhibitors of the viral reverse transcriptase can block new DNA synthesis but have no direct effect on the up to 50 or more copies of cccDNA that maintain the infected state. Thus, during antiviral therapy, the rates of loss of cccDNA, infected hepatocytes (1 or more molecules of cccDNA), and replicating DNAs may be quite different. In the present study, we asked how these losses compared when woodchucks chronically infected with woodchuck hepatitis virus were treated with L-FMAU [1-(2-fluoro-5-methyl-beta-L-arabinofuranosyl) uracil], an inhibitor of viral DNA synthesis. Viremia was suppressed for at least 8 months, after which drug-resistant virus began replicating to high titers. In addition, replicating viral DNAs were virtually absent from the liver after 6 weeks of treatment. In contrast, cccDNA declined more slowly, consistent with a half-life of approximately 33 to 50 days. The loss of cccDNA was comparable to that expected from the estimated death rate of hepatocytes in these woodchucks, suggesting that death of infected cells was one of the major routes for elimination of cccDNA. However, the decline in the actual number of infected hepatocytes lagged behind the decline in cccDNA, so that the average cccDNA copy number in infected cells dropped during the early phase of therapy. This observation was consistent with the possibility that some fraction of cccDNA was distributed to daughter cells in those infected hepatocytes that passed through mitosis. PMID- 11119602 TI - Essential role played by the C-terminal domain of glycoprotein I in envelopment of varicella-zoster virus in the trans-Golgi network: interactions of glycoproteins with tegument. AB - Varicella-zoster virus (VZV) is enveloped in the trans-Golgi network (TGN). Here we report that glycoprotein I (gI) is required within the TGN for VZV envelopment. Enveloping membranous TGN cisternae were microscopically identified in cells infected with intact VZV. These sacs curved around, and ultimately enclosed, nucleocapsids. Tegument coated the concave face of these sacs, which formed the viral envelope, but the convex surface was tegument-free. TGN cisternae of cells infected with VZV mutants lacking gI (gI(Delta)) or its C (gI(DeltaC))- or N-terminal (gI(DeltaN))-terminal domains were uniformly tegument coated and adhered to one another, forming bizarre membranous stacks. Viral envelopment was compromised, and no virions were delivered to post-Golgi structures. The TGN was not gI-immunoreactive in cells infected with the gI(Delta) or gI(DeltaN) mutants, but it was in cells infected with gI(DeltaC) (because the ectodomains of gI and gE interact). The presence in the TGN of gI lacking a C-terminal domain, therefore, was not sufficient to maintain enveloping cisternae. In cells infected with intact VZV or with gI(Delta), gI(DeltaN), or gI(DeltaC) mutants, ORF10p immunoreactivity was concentrated on the cytosolic face of TGN membranes, suggesting that it interacts with the cytosolic domains of glycoproteins. Because of the gE-gI interaction, cotransfected cells that expressed gE or gI were able to target truncated forms of the other to the TGN. Our data suggest that the C-terminal domain of gI is required to segregate viral and cellular proteins in enveloping TGN cisternae. PMID- 11119603 TI - Interferon regulatory factor 7 mediates activation of Tap-2 by Epstein-Barr virus latent membrane protein 1. AB - Transporter associated with antigen processing 2 (Tap-2) is responsible for ATP dependent transport of peptides from the cytosol to the endoplasmic reticulum, where peptides bind to newly synthesized human leukocyte antigen (HLA) class I molecules, which are essential for cellular immune responses. Epstein-Barr virus (EBV) latent membrane protein 1 (LMP-1) has been shown to induce the expression of Tap-2. In this study, the induction of endogenous Tap-2 by LMP-1 is shown to be associated with and requires the expression of interferon regulatory factor 7 (IRF-7). In DG75 Burkitt's lymphoma (BL) cells, in which LMP-1 induces the expression of IRF-7, LMP-1 induced endogenous Tap-2, and ectopic expression of IRF-7 could enhance the induction. In Akata BL cells, in which LMP-1 could not induce IRF-7, LMP-1 could not induce Tap-2. Addition of IRF-7, which complements the defect in Akata cells, could stimulate the expression of Tap-2. Furthermore, LMP-1 and IRF-7A but not other IRF-7 splicing variants could activate endogenous Tap-2. A Tap-2 promoter reporter construct could be activated by the overexpression of IRF-7A. The activation could be specifically enhanced by LMP-1 and was dependent on an intact interferon-stimulated response element (ISRE) present in the Tap-2 promoter. Also, IRF-7 can bind to the Tap-2 promoter under physiological conditions in vivo, as shown by formaldehyde cross-linking, as well as to the Tap-2 ISRE in vitro, as shown by gel mobility shift assays. Furthermore, LMP-1 facilitates the phosphorylation and nuclear translocation of IRF-7. These data point to the role of IRF-7 as a secondary mediator of LMP-1 activated signal transduction for Tap-2 as follows: LMP-1 stimulates the expression of IRF-7 and facilitates its phosphorylation and nuclear translocation, and then the activated IRF-7 mediates the activation of the cellular Tap-2 gene. The induction of Tap-2 by IRF-7 and LMP-1 may have an important implication for the immune response to EBV and its persistence in vivo. PMID- 11119604 TI - Calpain inhibition protects against virus-induced apoptotic myocardial injury. AB - Viral myocarditis is an important cause of human morbidity and mortality for which reliable and effective therapy is lacking. Using reovirus strain 8B infection of neonatal mice, a well-characterized experimental model of direct virus-induced myocarditis, we now demonstrate that myocardial injury results from apoptosis. Proteases play a critical role as effectors of apoptosis. The activity of the cysteine protease calpain increases in reovirus-infected myocardiocytes and can be inhibited by the dipeptide alpha-ketoamide calpain inhibitor Z-Leu aminobutyric acid-CONH(CH(2))3-morpholine (CX295). Treatment of reovirus-infected neonatal mice with CX295 protects them against reovirus myocarditis as documented by (i) a dramatic reduction in histopathologic evidence of myocardial injury, (ii) complete inhibition of apoptotic myocardial cell death as identified by terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling, (iii) a reduction in serum creatine phosphokinase, and (iv) improved weight gain. These findings are the first evidence for the importance of a calpain-associated pathway of apoptotic cell death in viral disease. Inhibition of apoptotic signaling pathways may be an effective strategy for the treatment of viral disease in general and viral myocarditis in particular. PMID- 11119606 TI - Characterization of adenovirus-induced inverted terminal repeat-independent amplification of integrated adeno-associated virus rep-cap sequences. AB - Stable packaging cell lines expressing the rep and cap genes for recombinant adeno-associated virus type 2 (rAAV-2) assembly constitute an attractive alternative to transient transfection protocols. We recently characterized a stable HeLa rep-cap cell clone (HeRC32) and demonstrated that upon vector transfection and adenovirus infection, efficient rAAV assembly correlated with a 100-fold amplification of the integrated rep-cap sequence with the inverted terminal repeats (ITRs) deleted. We now report a more detailed analysis of this phenomenon and highlight the key cellular and viral factors involved. Determination of the rep-cap copy number of HeRC32 cells indicated that maximum rep-cap amplification occurred between 24 and 48 h following adenovirus infection. Analysis by pulsed-field gel electrophoresis of adenovirus-infected HeRC32 cells indicated that amplified rep-cap sequences were found in an extrachromosomal form. Amplification of the rep-cap sequence with the ITRs deleted was not dependent on adenovirus replication and still occurred when the highly specific adenovirus polymerase was inactivated. In contrast, amplification was inhibited in the presence of aphidicolin, indicating that cellular polymerases were needed. Our study also documented that among the adenovirus gene products, the DNA-binding protein (DBP) was essential, since rep-cap amplification was severely abrogated when HeRC32 cells were infected at a nonpermissive temperature with an adenovirus mutant encoding a thermosensitive DBP. Furthermore, expression of DBP alone in HeRC32 cells was sufficient to induce a sustained level of rep-cap amplification. Finally, immunofluorescence analysis showed that HeRC32 cells expressing the DBP also simultaneously expressed the Rep proteins, suggesting a possible involvement of the latter in rep-cap amplification. Indeed, the lack of detectable amplification in an adenovirus-infected stable rep-cap HeLa cell clone unable to produce Rep proteins further supported that, among the viral gene products, both the DBP and Rep proteins are necessary to induce the targeted amplification of the integrated rep cap sequences in the absence of the AAV ITRs. PMID- 11119605 TI - Functional analysis of the simian immunodeficiency virus Vpx protein: identification of packaging determinants and a novel nuclear targeting domain. AB - The vpx gene products of human immunodeficiency virus type 2 (HIV-2) and of the closely related simian immunodeficiency viruses from sooty mangabeys (SIVsm) and macaques (SIVmac) comprise a 112-amino-acid virion-associated protein that is critical for efficient virus replication in nondividing cells such as macrophages. When expressed in the absence of other viral proteins, Vpx localizes to the nuclear membrane as well as to the nucleus; however, in the context of virus replication Vpx is packaged into virions via interaction with the p6 domain of the Gag precursor polyprotein (p55(gag)). To identify the domains essential for virion incorporation and nuclear localization, site-directed mutations were introduced into the vpx gene of SIVsmPBj1.9 and functionally analyzed. Our results show that (i) mutation of two highly conserved L74 and I75 residues impaired both virion incorporation and nuclear localization of Vpx; (ii) substitution of conserved H82, G86, C87, P103, and P106 residues impaired Vpx nuclear localization but not virion incorporation; (iii) mutations of conserved Y66, Y69, and Y71 residues impaired virion incorporation but not the translocation of Vpx to the nucleus; and (iv) a mutation at E30 (predicted to disrupt an N-terminal alpha-helix) had no effect on either virion incorporation or nuclear localization of Vpx. Importantly, mutations in Vpx which impaired nuclear localization also reduced virus replication in macaque macrophages, suggesting an important role of the carboxyl terminus of Vpx in nuclear translocation of the viral preintegration complex. Analyzing this domain in greater detail, we identified a 26-amino-acid (aa 60 to 85) fragment that was sufficient to mediate the transport of a heterologous protein (green fluorescent protein [GFP]) to the nucleus. Taken together, these results indicate that virion incorporation and nuclear localization are encoded by two partially overlapping domains in the C-terminus of Vpx (aa 60 to 112). The identification of a novel 26 amino-acid nuclear targeting domain provides a new tool to investigate the nuclear import of the HIV-2/SIV preintegration complex. PMID- 11119607 TI - An Epstein-Barr virus protein interacts with Notch. AB - The Epstein-Barr virus (EBV) BamHI A mRNAs were originally identified in cDNA libraries from nasopharyngeal carcinoma, where they are expressed at high levels. The RNAs are differentially spliced to form several open reading frames and also contain the BARF0 open reading frame at the 3' end. One cDNA, RK-BARF0, included a potential endoplasmic reticulum-targeting signal peptide sequence. The RK-BARF0 protein is shown here to interact with the Notch4 ligand binding domain, using yeast two-hybrid screening, coimmunoprecipitation, and confocal microscopy. This interaction induces translocation of a portion of the full-length unprocessed Notch4 to the nucleus by using the Notch nuclear localization signal. These effects of RK-BARF0 on Notch intracellular location indicate that EBV possibly modulates Notch signaling. Unprocessed Notch4 was also detected in immunoprecipitated complexes from EBV-infected cells by using a rabbit antiserum raised against a BARF0-specific peptide. This finding provides additional evidence for expression of RK-BARF0 and its interaction with Notch during EBV infection. In EBV-infected, EBNA2-negative cells, RK-BARF0 induced the expression of EBV latent membrane protein 1 (LMP1), and this induction was dependent on the RK-BARF0/Notch interaction domain. The activation of LMP1 expression by RK-BARF0 may be responsible for expression of LMP1 in EBV latent infections in the absence of EBNA2. PMID- 11119608 TI - Genetic evidence of a role for ATM in functional interaction between human T-cell leukemia virus type 1 Tax and p53. AB - Recent evidence from several investigators suggest that the human T-cell leukemia virus type 1 Tax oncoprotein represses the transcriptional activity of the tumor suppressor protein, p53. An examination of published findings reveals serious controversy as to the mechanism(s) utilized by Tax to inhibit p53 activity and whether the same mechanism is used by Tax in adherent and suspension cells. Here, we have investigated Tax-p53 interaction simultaneously in adherent epithelial (HeLa and Saos) and suspension T-lymphocyte (Jurkat) cells. Our results indicate that Tax activity through the CREB/CREB-binding protein (CBP), but not NF-kappaB, pathway is needed to repress the transcriptional activity of p53 in all tested cell lines. However, we did find that while CBP binding by Tax is necessary, it is not sufficient for inhibiting p53 function. Based on knockout cell studies, we correlated a strong genetic requirement for the ATM, but not protein kinase dependent DNA, protein in conferring a Tax-p53-repressive phenotype. PMID- 11119609 TI - Interaction of the influenza virus nucleoprotein with the cellular CRM1-mediated nuclear export pathway. AB - Influenza virus transcription occurs in the nuclei of infected cells, where the viral genomic RNAs are complexed with a nucleoprotein (NP) to form ribonucleoprotein (RNP) structures. Prior to assembly into progeny virions, these RNPs exit the nucleus and accumulate in the cytoplasm. The mechanisms responsible for RNP export are only partially understood but have been proposed to involve the viral M1 and NS2 polypeptides. We found that the drug leptomycin B (LMB), which specifically inactivates the cellular CRM1 polypeptide, caused nuclear retention of NP in virus-infected cells, indicating a role for the CRM1 nuclear export pathway in RNP egress. However, no alteration was seen in the cellular distribution of M1 or NS2, even in the case of a mutant virus which synthesizes greatly reduced amounts of NS2. Furthermore, NP was distributed throughout the nuclei of infected cells at early times postinfection but, when retained in the nucleus at late times by LMB treatment, was redistributed to the periphery of the nucleoplasm. No such change was seen in the nuclear distribution of M1 or NS2 after drug treatment. Similar to the behavior of NP, M1 and NS2 in infected cells, LMB treatment of cells expressing each polypeptide in isolation caused nuclear retention of NP but not M1 or NS2. Conversely, overexpression of CRM1 caused increased cytoplasmic accumulation of NP but had little effect on M1 or NS2 distribution. Consistent with this, NP bound CRM1 in vitro. Overall, these data raise the possibility that RNP export is mediated by a direct interaction between NP and the cellular CRM1 export pathway. PMID- 11119610 TI - A recombinant newcastle disease virus with low-level V protein expression is immunogenic and lacks pathogenicity for chicken embryos. AB - Newcastle disease virus (NDV) edits its P-gene mRNA by inserting a nontemplated G residue(s) at a conserved editing site (3'-UUUUUCCC-template strand). In the wild type virus, three amino-coterminal P-gene-derived proteins, P, V, and W, are produced at frequencies of approximately 68, 29, and 2%, respectively. By applying the reverse genetics technique, editing-defective mutants were generated in cell culture. Compared to the wild-type virus, mutants lacking either six nucleotides of the conserved editing site or the unique C-terminal part of the V protein produced as much as 5, 000-fold fewer infectious progeny in vitro or 200,000-fold fewer in 6-day-old embryonated chicken eggs. In addition, both mutants were unable to propagate in 9- to 11-day-old embryonated specific pathogen-free (SPF) chicken eggs. In contrast, a mutant (NDV-P1) with one nucleotide substitution (UUCUUCCC) grew in eggs, albeit with a 100-fold-lower infectious titer than the parent virus. The modification in the first two mutants described above led to complete abolition of V expression, whereas in NDV-P1 the editing frequency was reduced to less than 2%, and as a result, V was expressed at a 20-fold-lower level. NDV-P1 showed markedly attenuated pathogenicity for SPF chicken embryos, unlike currently available ND vaccine strains. These findings indicate that the V protein of NDV has a dual function, playing a direct role in virus replication as well as serving as a virulence factor. Administration of NDV P1 to 18-day-old embryonated chicken eggs hardly affected hatchability. Hatched chickens developed high levels of NDV-specific antibodies and were fully protected against lethal challenge, demonstrating the potential use of editing defective recombinant NDV as a safe embryo vaccine. PMID- 11119611 TI - Kaposi's sarcoma-associated herpesvirus LANA2 is a B-cell-specific latent viral protein that inhibits p53. AB - Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is associated with three proliferative diseases ranging from viral cytokine-induced hyperplasia to monoclonal neoplasia: multicentric Castleman's disease (CD), Kaposi's sarcoma (KS), and primary effusion lymphoma (PEL). Here we report a new latency-associated 1,704-bp KSHV spliced gene belonging to a cluster of KSHV sequences having homology to the interferon regulatory factor (IRF) family of transcription factors. ORFK10.5 encodes a protein, latency-associated nuclear antigen 2 (LANA2), which is expressed in KSHV-infected hematopoietic tissues, including PEL and CD but not KS lesions. LANA2 is abundantly expressed in the nuclei of cultured KSHV-infected B cells. Transcription of K10.5 in PEL cell cultures is not inhibited by DNA polymerase inhibitors nor significantly induced by phorbol ester treatment. Unlike LANA1, LANA2 does not elicit a serologic response from patients with KS, PEL, or CD as measured by Western blot hybridization. Both KSHV vIRF1 (ORFK9) and LANA2 (ORFK10.5) appear to have arisen through gene duplication of a captured cellular IRF gene. LANA2 is a potent inhibitor of p53-induced transcription in reporter assays. LANA2 antagonizes apoptosis due to p53 overexpression in p53-null SAOS-2 cells and apoptosis due to doxorubicin treatment of wild-type p53 U2OS cells. While LANA2 specifically interacts with amino acids 290 to 393 of p53 in glutathione S-transferase pull down assays, we were unable to demonstrate LANA2-p53 interaction in vivo by immunoprecipitation. These findings show that KSHV has tissue-specific latent gene expression programs and identify a new latent protein which may contribute to KSHV tumorigenesis in hematopoietic tissues via p53 inhibition. PMID- 11119612 TI - CXCR4 is down-regulated in cells infected with the CD4-independent X4 human immunodeficiency virus type 1 isolate m7NDK. AB - Macrophages and T cells infected in vitro with CD4-dependent human immunodeficiency virus type 1 (HIV-1) isolates have reduced levels of CD4 protein, a phenomenon involved in retroviral interference. We have previously characterized the first CD4-independent HIV-1 X4 isolate m7NDK, which directly interacts with CXCR4 through its mutated gp120. We thus investigate CXCR4 expression in cells infected with this m7NDK CXCR4-dependent HIV-1 mutant. We present evidence of the down-regulation of CXCR4 membrane expression in CD4 positive or -negative cells chronically infected with the HIV-1 m7NDK, a phenomenon which is not observed in the CD4-dependent HIV-1 NDK parental strain. This down-regulation of CXCR4 was demonstrated by fluorescence-activated cell sorter analysis and was confirmed by the absence of CXCR4 functionality in m7NDK infected cells, independently of the presence of CD4 protein. Furthermore, a drastic reduction of the intracellular level of CXCR4 protein was also observed. Reduced levels of CXCR4 mRNA transcripts were found in m7NDK-infected HeLa and CEM cells, reduced levels that could not be attributed to a reduced stability of CXCR4 mRNA. Down-regulation of CXCR4 on m7NDK-infected cells may thus be explained by transcriptional regulation. PMID- 11119613 TI - Differences in affinity of binding of lymphocytic choriomeningitis virus strains to the cellular receptor alpha-dystroglycan correlate with viral tropism and disease kinetics. AB - alpha-Dystroglycan (alpha-DG) was recently identified as a receptor for lymphocytic choriomeningitis virus (LCMV) and several other arenaviruses, including Lassa fever virus (W. Cao, M. D. Henry, P. Borrow, H. Yamada, J. H. Elder, E. V. Ravkov, S. T. Nichol, R. W. Compans, K. P. Campbell, and M. B. A. Oldstone, Science 282:2079-2081, 1998). Data presented in this paper indicate that the affinity of binding of LCMV to alpha-DG determines viral tropism and the outcome of infection in mice. To characterize this relationship, we evaluated the interaction between alpha-DG and several LCMV strains, variants, and reassortants. These viruses could be divided into two groups with respect to affinity of binding to alpha-DG, dependence on this protein for cell entry, viral tropism, and disease course. Viruses that exhibited high-affinity binding to alpha-DG displayed a marked dependence on alpha-DG for cell entry and were blocked from infecting mouse 3T6 fibroblasts by 1 to 4 nM soluble alpha-DG. In addition, high-affinity binding to alpha-DG correlated with an ability to infiltrate the white pulp (T-dependent) area of the spleen, cause ablation of the cytotoxic T-lymphocyte (CTL) response by day 7 postinfection, and establish a persistent infection. In contrast, viruses with a lower affinity of binding to alpha-DG were only partially inhibited from infecting alpha-DG(-/-) embryonic stem cells and required a concentration of soluble alpha-DG higher than 100 nM to prevent infection of mouse 3T6 fibroblasts. These viruses that bound at low affinity were mainly restricted to the splenic red pulp, and the host generated an effective CTL response that rapidly cleared the infection. Reassortants of viruses that bound to alpha-DG at high and low affinities were used to map genes responsible for the differences described to the S RNA, containing the virus attachment protein glycoprotein 1. PMID- 11119614 TI - Modulation of cellular and viral gene expression by the latency-associated nuclear antigen of Kaposi's sarcoma-associated herpesvirus. AB - Kaposi's sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8 (HHV-8), is the likely etiological agent of Kaposi's sarcoma and primary effusion lymphoma. Common to these malignancies is that tumor cells are latently infected with KSHV. Viral gene expression is limited to a few genes, one of which is the latency-associated nuclear antigen (LANA), the product of ORF73. Examination of the primary sequence of LANA reveals some structural features reminiscent of transcription factors, leading us to hypothesize that LANA may regulate viral and cellular transcription during latency. In reporter gene-based transient transfection assays, we found that LANA can have either positive or negative effects on gene expression. While expression of a reporter gene from several synthetic promoters was increased in the presence of LANA, expression from the human immunodeficiency virus (HIV) long terminal repeat (LTR)-and from NF-kappaB dependent reporter genes-was reduced by LANA expression. In addition, the promoter of KSHV ORF73 itself is activated up to 5.5-fold by LANA. This autoregulation may be important in tumorigenesis, because two other genes (v cyclin and v-FLIP) with likely roles in cell growth and survival are also controlled by this element. To identify cellular genes influenced by LANA, we employed cDNA array-based expression profiling. Six known genes (and nine expressed sequence tags) were found to be upregulated in LANA-expressing cell lines. One of these, Staf-50, is known to inhibit expression from the HIV LTR; most of the other known genes are interferon inducible, although the interferon genes themselves were not induced by LANA. These data demonstrate that LANA expression has effects on cellular and viral gene expression. We suggest that, whether direct or indirect in origin, these effects may play important roles in the pathobiology of KSHV infection. PMID- 11119615 TI - Induction and inhibition of apoptosis by pseudorabies virus in the trigeminal ganglion during acute infection of swine. AB - We examined the ability of pseudorabies virus (PRV) to induce and suppress apoptosis in the trigeminal ganglion during acute infection of its natural host. Eight pigs were intranasally inoculated with a virulent field strain of PRV, and at various early times after inoculation, the trigeminal ganglia were assessed histologically. PRV-infected cells were detected by use of immunohistochemistry and in situ hybridization, and apoptosis was identified by in situ terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling. Light and electron microscopy was also used for morphological studies. Apoptosis was readily detected among infiltrating immune cells that were located surrounding PRV infected neurons. The majority of PRV-infected neurons did not show morphological or histochemical evidence of apoptosis, even including those neurons that were surrounded by numerous inflammatory cells and exhibited profound pathological changes. However, neuronal virus-induced apoptosis also occurred but at a sporadic low level. These findings suggest that PRV is able to block apoptosis of infected trigeminal ganglionic neurons during acute infection of swine. Furthermore, our results also suggest that apoptosis of infiltrating inflammatory cells may represent an important viral mechanism of immune evasion. PMID- 11119616 TI - Functional and selective targeting of adenovirus to high-affinity Fcgamma receptor I-positive cells by using a bispecific hybrid adapter. AB - Adenovirus (Ad) efficiently delivers its DNA genome into a variety of cells and tissues, provided that these cells express appropriate receptors, including the coxsackie-adenovirus receptor (CAR), which binds to the terminal knob domain of the viral capsid protein fiber. To render CAR-negative cells susceptible to Ad infection, we have produced a bispecific hybrid adapter protein consisting of the amino-terminal extracellular domain of the human CAR protein (CARex) and the Fc region of the human immunoglobulin G1 protein, comprising the hinge and the CH2 and CH3 regions. CARex-Fc was purified from COS7 cell supernatants and mixed with Ad particles, thus blocking Ad infection of CAR-positive but Fc receptor-negative cells. The functionality of the CARex domain was further confirmed by successful immunization of mice with CARex-Fc followed by selection of a monoclonal anti human CAR antibody (E1-1), which blocked Ad infection of CAR-positive cells. When mixed with Ad expressing eGFP, CARex-Fc mediated an up to 250-fold increase of transgene expression in CAR-negative human monocytic cell lines expressing the high-affinity Fcgamma receptor I (CD64) but not in cells expressing the low affinity Fcgamma receptor II (CD32) or III (CD16). These results open new perspectives for Ad-mediated cancer cell vaccination, including the treatment of acute myeloid leukemia. PMID- 11119617 TI - Structure of recombinant rabies virus nucleoprotein-RNA complex and identification of the phosphoprotein binding site. AB - Rabies virus nucleoprotein (N) was produced in insect cells, in which it forms nucleoprotein-RNA (N-RNA) complexes that are biochemically and biophysically indistinguishable from rabies virus N-RNA. We selected recombinant N-RNA complexes that were bound to short insect cellular RNAs which formed small rings containing 9 to 11 N monomers. We also produced recombinant N-RNA rings and viral N-RNA that were treated with trypsin and that had lost the C-terminal quarter of the nucleoprotein. Trypsin-treated N-RNA no longer bound to recombinant rabies virus phosphoprotein (the viral polymerase cofactor), so the presence of the C terminal part of N is needed for binding of the phosphoprotein. Both intact and trypsin-treated recombinant N-RNA rings were analyzed with cryoelectron microscopy, and three-dimensional models were calculated from single-particle image analysis combined with back projection. Nucleoprotein has a bilobed shape, and each monomer has two sites of interaction with each neighbor. Trypsin treatment cuts off part of one of the lobes without shortening the protein or changing other structural parameters. Using negative-stain electron microscopy, we visualized phosphoprotein bound to the tips of the N-RNA rings, most likely at the site that can be removed by trypsin. Based on the shape of N determined here and on structural parameters derived from electron microscopy on free rabies virus N-RNA and from nucleocapsid in virus, we propose a low-resolution model for rabies virus N-RNA in the virus. PMID- 11119618 TI - Immune response induced by airway sensitization after influenza A virus infection depends on timing of antigen exposure in mice. AB - To study which phase of viral infection promotes antigen sensitization via the airway and which type of antigen-presenting cells contributes to antigen sensitization, BALB/c mice were sensitized by inhalation of ovalbumin (OA) during the acute phase or the recovery phase of influenza A virus infection, and then 3 weeks later animals were challenged with OA. The numbers of eosinophils and lymphocytes, the amounts of interleukin-4 (IL-4) and IL-5 in the bronchoalveolar lavage fluid, and the serum levels of OA-specific immunoglobulin G1 (IgG1) and IgE increased in mice sensitized during the acute phase (acute phase group), while a high level of gamma interferon production was detected in those sensitized during the recovery phase (recovery phase group). In the acute phase group, both major histocompatibility complex class II molecules and CD11c were strongly stained on the bronchial epithelium; in the recovery phase group, however, neither molecule was detected. OA-capturing dendritic cells (DCs) migrated to the regional lymph nodes, and a small number of OA-capturing macrophages were also observed in the lymph nodes of the acute phase group. In the recovery group, however, no OA-capturing DCs were detected in either the lungs or the lymph nodes, while OA-capturing macrophages were observed in the lymph nodes. These results indicate that the timing of antigen sensitization after viral infection determines the type of immune response. PMID- 11119619 TI - The coronavirus infectious bronchitis virus nucleoprotein localizes to the nucleolus. AB - The coronavirus nucleoprotein (N) has been reported to be involved in various aspects of virus replication. We examined by confocal microscopy the subcellular localization of the avian infectious bronchitis virus N protein both in the absence and in the context of an infected cell and found that N protein localizes both to the cytoplasmic and nucleolar compartments. PMID- 11119620 TI - Cooperative transformation and coexpression of bovine papillomavirus type 1 E5 and E7 proteins. AB - Productively infected bovine fibropapillomas were examined for bovine papillomavirus type 1 (BPV-1) E7 localization. BPV-1 E7 was observed in the cytoplasm of basal and lower spinous epithelial cells, coexpressed in the cytoplasm of basal cells with the E5 oncoprotein. E7 was also observed in nucleoli throughout the basal and spinous layers but not in the granular cell layer. Ectopic expression of E7 in cultured epithelial cells gave rise to localization similar to that seen in productive fibropapillomas, with cytoplasmic and nucleolar expression observed. Consistent with the coexpression of E7 and E5 in basal keratinocytes, BPV-1 E7 cooperated with E5 as well as E6 in an anchorage independence transformation assay. While E5 is expressed in both basal and superficial differentiating keratinocytes, BPV-1 E7 is only observed in basal and lower spinous epithelial cells. Therefore, BPV-1 E7 may serve to modulate the cellular response of basal epithelial cells to E5 expression. PMID- 11119621 TI - Tissue distribution and timing of appearance of polytropic envelope recombinants during infection with SL3-3 murine leukemia virus or its weakly pathogenic SL3DeltaMyb5 mutant. AB - A time course analysis was performed to identify the sites of formation and timing of appearance of polytropic recombinant viruses following infection of NIH/Swiss mice with the murine retrovirus SL3-3 murine leukemia virus (SL3) or with a weakly pathogenic mutant termed SL3DeltaMyb5. The results indicated that (i) polytropic recombinant viruses occur initially in the thymus of SL3-infected animals, (ii) the timing of appearance of polytropic recombinants in bone marrow is not consistent with their participation in the previously reported formation of transplantable tumor-forming cells at 3 to 4 week postinoculation, and (iii) the efficient generation of recombinant virus is correlated with efficient tumor induction. PMID- 11119622 TI - The ability of integrin alpha(v)beta(3) To function as a receptor for foot-and mouth disease virus is not dependent on the presence of complete subunit cytoplasmic domains. AB - The integrin alpha(v)beta(3) has been shown to function as one of the integrin receptors on cultured cells for foot-and-mouth disease virus (FMDV), and high efficiency utilization of the bovine homolog of this integrin is dependent on the cysteine-rich repeat region of the bovine beta(3) subunit. In this study we have examined the role of the cytoplasmic domains of the alpha(v) and beta(3) subunits in FMDV infection. We have found that truncations or extensions of these domains of either subunit, including deletions removing almost all of the cytoplasmic domains, had little or no effect on the ability of the integrin to function as a receptor for FMDV. The lysosomotropic agent monensin inhibited viral replication in cells transfected with either intact or cytoplasmic domain-truncated alpha(v)beta(3). In addition, viral replication in transfected cells was inhibited by an alpha(v)beta(3) function-blocking antibody but not by function blocking antibodies to three other RGD-directed integrins, suggesting that these integrins are not involved in the infectious process. These results indicate that alterations to the cytoplasmic domains of either subunit, which lead to the inability of the integrin receptor to function normally, do not abolish the ability of the integrin to bind and internalize this viral ligand. PMID- 11119623 TI - Induction of host gene expression following infection of chicken embryo fibroblasts with oncogenic Marek's disease virus. AB - Microarrays containing 1,126 nonredundant cDNAs selected from a chicken activated T-cell expressed sequence tag database (http://chickest.udel.edu) were used to examine changes in host cell gene expression that accompany infection of chicken embryo fibroblasts (CEF) with Marek's disease virus (MDV). Host genes that were reproducibly induced by infection of CEF with the oncogenic RB1B strain of MDV included macrophage inflammatory protein, interferon response factor 1, interferon-inducible protein, quiescence-specific protein, thymic shared antigen 1, major histocompatibility complex (MHC) class I, MHC class II, beta(2) microglobulin, clusterin, interleukin-13 receptor alpha chain, ovotransferrin, a serine/threonine kinase, and avian leukosis virus subgroup J glycoprotein. PMID- 11119624 TI - Direct ex vivo measurement of CD8(+) T-lymphocyte responses to human parvovirus B19. AB - Parvovirus B19 is a common human pathogen which can cause severe syndromes, including aplastic anemia and fetal hydrops. The mapping of the first parvovirus B19-derived CD8(+) T-lymphocyte epitope is described. This HLA-B35-restricted peptide derives from the nonstructural (NS1) protein and is strongly immunogenic in B19 virus-seropositive donors. PMID- 11119625 TI - Induction of polyomavirus-specific CD8(+) T lymphocytes by distinct dendritic cell subpopulations. AB - Dendritic cells are pivotal antigen-presenting cells for generating adaptive T cell responses. Here, we show that dendritic cells belonging to either the myeloid-related or lymphoid-related subset are permissive for infection by mouse polyomavirus and, when loaded with a peptide corresponding to the immunodominant anti-polyomavirus CD8(+) T-cell epitope or infected by polyomavirus, are each capable of driving expansion of primary polyomavirus-specific CD8(+) T-cell responses in vivo. PMID- 11119626 TI - Properties of wild-type, C-terminally truncated, and chimeric maedi-visna virus glycoprotein and putative pseudotyping of retroviral vector particles. AB - We have characterized the properties of the maedi-visna virus (MVV) glycoprotein, which has a long cytoplasmic C-terminal domain, and of a panel of C-terminally truncated and C-terminally chimeric MVV-Env constructs. Cells expressing wild type MVV glycoprotein form syncytia with target cells from many different species and tissues, demonstrating that the MVV-Env cellular receptor is widely distributed. Similar to the situation with other lentiviral glycoproteins, truncation of the C-terminal domain of MVV-Env significantly increases its membrane fusion capacity. However, despite their presence in a fusogenic form at the cell surface, neither the wild-type nor any of the C-terminally modified MVV Env constructs, these latter lacking sterically inhibitory C termini, were able to successfully pseudotype murine leukemia virus- or human immunodeficiency virus derived vector particles. PMID- 11119628 TI - Benzene in the environment: an assessment of the potential risks to the health of the population. AB - OBJECTIVES: Benzene has long been recognised as a carcinogen and recent concern has centred on the effects of continuous exposure to low concentrations of benzene both occupationally and environmentally. This paper presents an overview of the current knowledge about human exposure to benzene in the United Kingdom population based on recently published data, summarises the known human health effects, and uses this information to provide a risk evaluation for sections of the general United Kingdom population. METHOD: Given the minor contribution that non-inhalation sources make to the overall daily intake of benzene to humans, only exposure from inhalation has been considered when estimating the daily exposure of the general population to benzene. Exposure of adults, children, and infants to benzene has been estimated for different exposure scenarios with time activity patterns and inhalation and absorption rates in conjunction with measured benzene concentrations for a range of relevant microenvironments. Exposures during refuelling and driving, as well as the contribution of active and passive tobacco smoke, have been considered as part of the characterisation of risk of the general population. RESULTS: Infants (<1 years old), the average child (11 years old), and non-occupationally exposed adults, receive average daily doses in the range of 15-26, 29-50, and 75-522 microg of benzene, respectively, which correspond to average ranges to benzene in air of 3.40-5.76 microg/m(3), 3.37-5.67 microg/m(3), and 3.7-41 microg/m(3) for infants, children, and adults, respectively. Infants and children exposed to environmental tobacco smoke have concentrations of exposure to benzene comparable with those of an adult passive smoker. This is a significant source of exposure as a 1995 United Kingdom survey has shown that 47% of children aged 2-15 years live in households where at least one person smokes. The consequence of exposure to benzene in infants is more significant than for children or adults owing to their lower body weight, resulting in a higher daily intake for infants compared with children or non-smoking adults. A worst case scenario for exposure to benzene in the general population is that of an urban smoker who works adjacent to a busy road for 8 hours/day-for example, a maintenance worker-who can receive a mean daily exposure of about 820 microg (equal to an estimated exposure of 41 microg/m(3)). The major health risk associated with low concentrations of exposure to benzene has been shown to be leukaemia, in particular acute non-lymphocytic leukaemia. The lowest concentration of exposure at which an increased incidence of acute non lymphocytic leukaemia among occupationally exposed workers has been reliably detected, has been estimated to be in the range of 32-80 mg/m(3). Although some studies have suggested that effects may occur at lower concentrations, clear estimates of risk have not been determined, partly because of the inadequacy of exposure data and the few cases. CONCLUSIONS: Overall the evidence from human studies suggests that any risk of leukaemia at concentrations of exposure in the general population of 3.7-42 microg/m(3)-that is at concentrations three orders of magnitude less than the occupational lowest observed effect level-is likely to be exceedingly small and probably not detectable with current methods. This is also likely to be true for infants and children who may be exposed continuously to concentrations of 3.4-5.7 microg/m(3). As yet there is no evidence to suggest that continuous exposures to these environmental concentrations of benzene manifest as any other adverse health effect. PMID- 11119629 TI - Exposure to organic solvents and personality. AB - OBJECTIVES: Although cognitive and neuropsychological changes have been found after high cumulative exposures to solvents, it is not clear whether such exposures are associated with personality characteristics. To study this two groups of British and Chinese dockyard painters who had been heavily exposed to paint solvents have been investigated. METHODS: 260 Male dockyard painters in the United Kingdom, 539 local community controls, 109 Chinese dockyard painters, and 255 dockyard controls completed the Eysenck personality questionnaire, neuroticism (N) and social conformity or dissimulation (L) scales. The non parametric Kruskal-Wallis test was used to evaluate differences in scores of personality traits between painters and controls. Adjusted relative risks for painters having high N and L scores were calculated in a Breslow-Cox regression analysis, and exposure-response relations were examined in multivariate logistic regression analysis. Non-parametric Spearman's correlation was used to examine relations between previously determined neuropsychological symptoms and personality. RESULTS: Both British and Chinese data showed that mean neuroticism scores of painters were significantly higher than controls, whereas scores of social conformity did not differ. Relative risk of being a painter increased significantly with increasing N scores, but L scores showed no such trend. In a case-control analysis, there were significant exposure-response relations for the N score. In the United Kingdom the odds ratios (ORs) (95% confidence interval (95% CI), were 2.03 (0.79 to 5.22) for 1-4 years of exposure, 2.38 (0.82 to 6.91) for 5-9 years, 7.05 (1.27 to 39.25) for 10-14 years, and 1.76 (0.63 to 4.89) for 15-41 years. In the Chinese painters, ORs were 4.66 (1.38 to 15.75) for 2-14 years, 10.03 (2.96 to 34.04) for 15-18 years, and 13.56 (3.78 to 48.59) for 19-43 years. Neuroticism was significantly positively related to neuropsychological symptoms in all subjects. Social conformity showed no association with neuropsychological symptoms in British painters and a negative relation among the Chinese painters. CONCLUSION: Increasing symptoms suggesting neuroticism seemed to relate to the duration of painting whereas scores for social conformity and dissimulation did not. The relation between exposure time and response suggests that increased neuroticism may be caused by long term occupational exposure to organic solvents. PMID- 11119630 TI - Evaluation of a modified German version of the Q16 questionnaire for neurotoxic symptoms in workers exposed to solvents. AB - OBJECTIVES: To assess sensitivity and specificity of a questionnaire designed to detect neurotoxic symptoms in workers exposed to solvents and in patients with a psycho-organic syndrome. METHODS: The Swedish Q16 is a self administered questionnaire for neurotoxic symptoms. The modified German version consists of 18 questions. The results were analysed from 1166 questionnaires which were completed by adults belonging to the following groups; 483 workers with occupational exposure to solvents and 193 non-exposed controls, 25 patients with a psycho-organic syndrome, 25 sex and age matched patients with a lung disease, and a sample of 440 people from the general population. RESULTS: The German Q18 was easy to handle and quick to perform. Workers exposed to solvents reported significantly more complaints than controls (2.9 v 2.5). All patients with a psycho-organic syndrome had five or more complaints. This was true for only 32% of patients with lung disease. These comparisons showed that chronic exposure to solvents was associated with subjective complaints related particularly to cognitive functions. In the sample of the general population, age, education level, smoking habits, and time of performance showed no significant influence on the Q18 result. Women had significantly more complaints than men (3.2 v 2.3). People who reported drinking alcohol occasionally or moderately had significantly fewer complaints than teetotalers. CONCLUSIONS: The German Q18 has an acceptable sensitivity and reliability, a reasonable specificity, and a good practicability. It is a useful instrument for screening workers exposed to solvents. A cut off point of 5 for men is recommended, and a cut off point of 6 for women is proposed. PMID- 11119631 TI - Mortality in chemical workers potentially exposed to 2,4-dichlorophenoxyacetic acid (2,4-D) 1945-94: an update. AB - OBJECTIVE: To update and add to a previously identified cohort of employees potentially exposed to the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The putative association between 2,4-D and non-Hodgkin's lymphoma has been debated for more than a decade. METHODS: Cohort members were male employees of The Dow Chemical Company who manufactured or formulated 2,4-D any time from 1945 to the end of 1994. Their mortality experience was compared with national rates and with more than 40 000 other company employees who worked at the same location. RESULTS: 330 Deaths were observed among 1517 people compared with 365 expected (standardised mortality ratio (SMR)=0.90, 95% confidence interval (95% CI) 0.81 to 1.01). There were no significantly increased SMRs for any of the causes of death analyzed. When compared with the United States rates, the SMR for non Hodgkin's lymphoma (NHL) was 1.00 (95% CI 0.21 to 2.92). The internal comparison with other Dow employees showed a non-significant relative risk of 2.63, (95% CI 0.85 to 8.33). Death was attributed to amyotrophic lateral sclerosis (ALS) for three cohort members. Compared with the other company employees, the relative risk was 3.45 (95% CI 1.10 to 11.11). The cases were employed in the manufacture or formulation of 2,4-D at different periods (1947-9, 1950-1, and 1968-86), and for varying durations of time (1.3, 1.8, and 12.5 years). CONCLUSION: There was no evidence of a causal association between exposure to 2,4-D and mortality due to all causes and total malignant neoplasms. No significant risk due to NHL was found. Although not an initial hypothesis, an increased relative risk of ALS was noted. This finding is unsupported by other animal and human studies. PMID- 11119632 TI - Exposure to silica and silicosis among tin miners in China: exposure-response analyses and risk assessment. AB - OBJECTIVES: To investigate the risk of silicosis among tin miners and to investigate the relation between silicosis and cumulative exposure to dust (Chinese total dust and respirable crystalline silica dust). METHODS: A cohort study of 3010 miners exposed to silica dust and employed for at least 1 year during 1960-5 in any of four Chinese tin mines was conducted. Historical total dust data from China were used to create a job exposure matrix for facility, job title, and calendar year. The total dust exposure data from China were converted to estimates of exposure to respirable crystalline silica for comparison with findings from other epidemiological studies of silicosis. Each worker's work history was abstracted from the complete employment records in mine files. Diagnoses of silicosis were based on 1986 Chinese pneumoconiosis Roentgen diagnostic criteria, which classified silicosis as stages I-III-similar to an International Labour Organisation (ILO) classification of 1/1 or greater. RESULTS: There were 1015 (33.7%) miners identified with silicosis, who had a mean age of 48.3 years, with a mean of 21.3 years after first exposure (equivalent to 11.0 net years in a dusty job). Among those who had silicosis, 684 miners (67.4%) developed silicosis after exposure ended (a mean of 3.7 years after). The risk of silicosis was strongly related to cumulative exposure to silica dust and was well fitted by the Weibull distribution, with the risk of silicosis less than 0.1% when the Chinese measure of cumulative exposure to total dust (CTD) was under 10 mg/m(3)-years (or 0.36 mg/m(3)-years of respirable crystalline silica), increasing to 68.7% when CTD exposure was 150 mg/m(3)-years (or 5.4 mg/m(3)-years of respirable crystalline silica). Latency period was not correlated to the risk of silicosis or cumulative dose of exposure. This study predicts about a 36% cumulative risk of silicosis for a 45 year lifetime exposure to these tin mine dusts at the CTD exposure standard of 2 mg/m(3), and a 55% risk at 45 years exposure to the current United States Occupational Safety and Health Administration and Mine Safety and Health Administration standards of 0.1 mg/m(3) 100% respirable crystalline silica dust. CONCLUSIONS: A clear exposure-response relation was detected for silicosis in Chinese tin miners. The study results were similar to most, but not all, findings from other large scale exposure-response studies. PMID- 11119633 TI - Crystalline silica exposure and lung cancer mortality in diatomaceous earth industry workers: a quantitative risk assessment. AB - OBJECTIVE: To use various exposure-response models to estimate the risk of mortality from lung cancer due to occupational exposure to respirable crystalline silica dust. METHODS: Data from a cohort mortality study of 2342 white male California diatomaceous earth mining and processing workers exposed to crystalline silica dust (mainly cristobalite) were reanalyzed with Poisson regression and Cox's proportional hazards models. Internal and external adjustments were used to control for potential confounding from the effects of time since first observation, calendar time, age, and Hispanic ethnicity. Cubic smoothing spline models were used to assess the fit of the models. Exposures were lagged by 10 years. Evaluations of the fit of the models were performed by comparing their deviances. Lifetime risks of lung cancer were estimated up to age 85 with an actuarial approach that accounted for competing causes of death. RESULTS: Exposure to respirable crystalline silica dust was a significant predictor (p<0.05) in nearly all of the models evaluated and the linear relative rate model with a 10 year exposure lag seemed to give the best fit in the Poisson regression analysis. For those who died of lung cancer the linear relative rate model predicted rate ratios for mortality from lung cancer of about 1.6 for the mean cumulative exposure to respirable silica compared with no exposure. The excess lifetime risk (to age 85) of mortality from lung cancer for white men exposed for 45 years and with a 10 year lag period at the current Occupational Safety and Health Administration (OSHA) standard of about 0.05 mg/m(3) for respirable cristobalite dust is 19/1000 (95% confidence interval (95% CI) 5/1000 to 46/1000). CONCLUSIONS: There was a significant risk of mortality from lung cancer that increased with cumulative exposure to respirable crystalline silica dust. The predicted number of deaths from lung cancer suggests that current occupational health standards may not be adequately protecting workers from the risk of lung cancer. PMID- 11119634 TI - Audiometric notch as a sign of noise induced hearing loss. AB - OBJECTIVES: To investigate the relation between different types of exposure to noise and a classic sign of noise induced hearing loss (NIHL), the audiometric notch. METHODS: The study sample had exposure to both continuous and impulse noise and was drawn from a population of electrical transmission workers. Audiograms, taken as part of a hearing conservation programme, were read by three clinicians experienced in the assessment of NIHL. Working independently and using their clinical judgment, they were asked to identify localised increases in the threshold of hearing (audiometric notches) which they would attribute to noise, had a suitable history of exposure been elicited. Prevalent cases of NIHL were identified by the presence of a notch in either ear. Risk factors for NIHL were assessed by a questionnaire which sought information about exposure to air blast circuit breaker noise; firearms; explosions, and continuous noise. The odds of exposure to these factors in those with and without hearing loss were calculated, and odds ratios (ORs) and 95% confidence intervals (95% CIs) were estimated by logistic regression. RESULTS: Of the 648 questionnaires sent out 357 were returned, a response rate of 55%. Of these, at least two out of the three assessors identified 175 (49%) people with a notch at any audiometric frequency. There was no association between these cases and the NIHL risk factors identified by the questionnaire, but a further frequency specific analysis showed a small proportion of people (15 (4%)) with notches at 4 kHz who had the expected associations with exposure to noise and a significant OR for firearms of 4.25 (95% CI 1.28 to 14.1). The much larger proportion of people with 6 kHz notches (110 (31%)) did not show these associations. CONCLUSIONS: To diagnose NIHL it is important to elicit a detailed and accurate history of exposure to noise: although the notch at 4 kHz is a well established clinical sign and may be valuable in confirming the diagnosis, the 6 kHz notch is variable and of limited importance. PMID- 11119635 TI - Mortality among women and men relative to unemployment, part time work, overtime work, and extra work: a study based on data from the Swedish twin registry. AB - OBJECTIVE: To examine mortality before 70 years of age among women and men relative to unemployment, part time work, overtime work, and extra work. Age, marital status, children, smoking and alcohol habits, use of sleeping pills and tranquilisers, stress, shift work, personality factors, and long lasting or serious illness were taken into account as potential confounding factors. METHODS: The study group comprised a subcohort of the Swedish twin registry, people born in 1926-58. Data were based on a postal questionnaire of 1973 and on information from the Swedish Causes of Death Registry. All subjects reporting a main occupation were selected, 9500 women and 11 132 men, and mortality from all causes during 1973-96 was analysed. The subjects were treated as a sample from the general population regardless of the twinning. RESULTS: Unemployment in 1973 among both women and men showed an association with increased mortality. The adjusted relative risk (RR) (95% confidence interval (95% CI)) was 1.98 (1.16 to 3.38), for women and 1.43 (0.91 to 2.25) for men. For the first 5 years of follow up, a threefold increase in risk was found for men (RR (95% CI) 3.29 (1.33 to 8.17)). The RR declined by time, but remained increased throughout the 24 year study period. In women overtime work of more than 5 hours a week was followed by an increased mortality rate (RR (95% CI) 1.92 (1.13 to 3.25)). A protective effect of moderate overtime work of a maximum 5 hours a week was shown for men (RR (95% CI) 0.58 (0.43 to 0.80)), whereas an increased mortality was indicated for part time work (RR (95% CI) 1.58 (0.91 to 2.77)) and extra work (work outside employment) of more than 5 hours a week (RR (95% CI) 1.29 (0.99 to 1.69)). CONCLUSION: Unemployment and some time aspects of work were associated with subsequent mortality, even when controlling for social, behavioural, work, and health related factors. The idea that losing a job may have less importance for women than for men is not supported by this study. PMID- 11119638 TI - Review of the fifth annual Johns Hopkins Protein Folding Meeting. PMID- 11119636 TI - Comparing performance on a simulated 12 hour shift rotation in young and older subjects. AB - OBJECTIVES: To simulate a 12 hour shift rotation and measure the difference in performance if any, between older and younger subjects. Significant reductions in neurobehavioural performance during shift work and particularly night work have long been recognised. There are conflicting reports of the effects of 12 hour shifts on performance, alertness, and safety. Furthermore, research suggests that older shift workers have more sleep disruption and maladaption to shift work. When this is combined with longer hours at work there may be considerable reductions in performance for older compared with younger workers. METHODS: Thirty two subjects were allocated to groups according to age. Group one had 16 subjects with a mean (SD) age of 21.2 (2.7) years, and group two had 16 subjects with a mean (SD) age of 43.9 (6.8) years. Subjects came to the laboratory for six consecutive days and completed a simulated 12 hour shift rotation consisting of two 12 hour day shifts (0700-1900), followed by two 12 hour night shifts (1900 0700). During the work period subjects completed a computer administered neurobehavioural performance task every hour. RESULTS: Performance for the older subjects was consistently lower than for the younger subjects. There was a significant difference in performance across the shift between older and younger subjects. There was a significant change in performance across the shifts in the older subjects, such that performance significantly increased across the day shifts and decreased across the night shifts. By contrast, the younger subjects were able to maintain performance across both day shifts and the second night shift. CONCLUSIONS: There are significant differences in performance of older and younger subjects during a simulated 12 hour shift rotation. Future studies both in the field and the laboratory would be useful in determining whether this is typical and if there are any important consequences for the older worker on 12 hour shifts. PMID- 11119639 TI - Conserved key amino acid positions (CKAAPs) derived from the analysis of common substructures in proteins. AB - An all-against-all protein structure comparison using the Combinatorial Extension (CE) algorithm applied to a representative set of PDB structures revealed a gallery of common substructures in proteins (http://cl.sdsc.edu/ce.html). These substructures represent commonly identified folds, domains, or components thereof. Most of the subsequences forming these similar substructures have no significant sequence similarity. We present a method to identify conserved amino acid positions and residue-dependent property clusters within these subsequences starting with structure alignments. Each of the subsequences is aligned to its homologues in SWALL, a nonredundant protein sequence database. The most similar sequences are purged into a common frequency matrix, and weighted homologues of each one of the subsequences are used in scoring for conserved key amino acid positions (CKAAPs). We have set the top 20% of the high-scoring positions in each substructure to be CKAAPs. It is hypothesized that CKAAPs may be responsible for the common folding patterns in either a local or global view of the protein folding pathway. Where a significant number of structures exist, CKAAPs have also been identified in structure alignments of complete polypeptide chains from the same protein family or superfamily. Evidence to support the presence of CKAAPs comes from other computational approaches and experimental studies of mutation and protein-folding experiments, notably the Paracelsus challenge. Finally, the structural environment of CKAAPs versus non-CKAAPs is examined for solvent accessibility, hydrogen bonding, and secondary structure. The identification of CKAAPs has important implications for protein engineering, fold recognition, modeling, and structure prediction studies and is dependent on the availability of structures and an accurate structure alignment methodology. Proteins 2001;42:148-163. PMID- 11119640 TI - Identification and ab initio simulations of early folding units in proteins. AB - The location of protein subunits that form early during folding, constituted of consecutive secondary structure elements with some intrinsic stability and favorable tertiary interactions, is predicted using a combination of threading algorithms and local structure prediction methods. Two folding units are selected among the candidates identified in a database of known protein structures: the fragment 15-55 of 434 cro, an all-alpha protein, and the fragment 1-35 of ubiquitin, an alpha/beta protein. These units are further analyzed by means of Monte Carlo simulated annealing using several database-derived potentials describing different types of interactions. Our results suggest that the local interactions along the chain dominate in the first folding steps of both fragments, and that the formation of some of the secondary structures necessarily occurs before structure compaction. These findings led us to define a prediction protocol, which is efficient to improve the accuracy of the predicted structures. It involves a first simulation with a local interaction potential only, whose final conformation is used as a starting structure of a second simulation that uses a combination of local interaction and distance potentials. The root mean square deviations between the coordinates of predicted and native structures are as low as 2-4 A in most trials. The possibility of extending this protocol to the prediction of full proteins is discussed. Proteins 2001;42:164-176. PMID- 11119641 TI - Stabilizing C-terminal tails on AraC. AB - We examined the effects of the metabolic stability of random sequences appended to the C-terminus of the dimerization domain of the regulatory protein of the Escherichia coli arabinose operon, AraC. Genetic scoring utilized the trans dominant negative effect of the dimerization domain on the activity of intact AraC, and physical scoring used sodium dodecyl sulfate (SDS) gel electrophoresis. We confirmed previous results obtained with Arc and lambda repressors that C terminal charged residues tend to be stabilizing and that hydrophobic residues are destabilizing. Additionally, we found that the provision of a single, charged C-terminal residue conferred significant stability that was independent of interior sequence. Hence, it appears that in the engineering of proteins, flexible tails may be freely added, with only the identity of the C-terminal amino acid being restricted. Proteins 2001;42:177-181. PMID- 11119642 TI - Three-dimensional structure of a complex of galanthamine (Nivalin) with acetylcholinesterase from Torpedo californica: implications for the design of new anti-Alzheimer drugs. AB - The 3D structure of a complex of the anti-Alzheimer drug galanthamine with Torpedo californica acetylcholinesterase is reported. Galanthamine, a tertiary alkaloid extracted from several species of Amarylidacae, is so far the only drug that shows a dual activity, being both an acetylcholinesterase inhibitor and an allosteric potentiator of the nicotinic response induced by acetylcholine and competitive agonists. The X-ray structure, at 2.5A resolution, shows an unexpected orientation of the ligand within the active site, as well as unusual protein-ligand interactions. The inhibitor binds at the base of the active site gorge, interacting with both the acyl-binding pocket and the principal quaternary ammonium-binding site. However, the tertiary amine group of galanthamine does not directly interact with Trp84. A docking study using the program AUTODOCK correctly predicts the orientation of galanthamine in the active site. The docked lowest-energy structure has a root mean square deviation of 0.5A with respect to the corresponding crystal structure of the complex. The observed binding mode explains the affinities of a series of structural analogs of galanthamine and provides a rational basis for structure-based drug design of synthetic derivatives with improved pharmacological properties. Proteins 2001;42:182-191. PMID- 11119643 TI - Localization of the C-terminus of rat glutathione S-transferase A1-1: crystal structure of mutants W21F and W21F/F220Y. AB - Twelve C-terminal residues of human glutathione S-transferase A1-1 form a helix in the presence of glutathione-conjugate, or substrate alone, and partly cover the active site. According to X-ray structures, the helix is disordered in the absence of glutathione, but it is not known if it is helical and delocalized, or in a random-coil conformation. Mutation to a tyrosine of residue 220 within this helix was previously shown to affect the pK(a) of Tyr-9 at the active site, in the apo form of the enzyme, and it was proposed that an on-face hydrogen bond between Tyr-220 and Tyr-9 provided a means for affecting this pK(a). In the current study, X-ray structures of the W21F and of the C-terminal mutation, W21F/F220Y, with glutathione sulfonate bound, show that the C-terminal helix is disordered (or delocalized) in the W21F crystal but is visible and ordered in a novel location, a crystal packing crevice, in one of three monomers in the W21F/F220Y crystal, and the proposed hydrogen bond is not formed. Fluorescence spectroscopy studies using an engineered F222W mutant show that the C-terminus remains delocalized in the absence of glutathione or when only the glutathione binding site is occupied, but is ordered and localized in the presence of substrate or conjugate, consistent with these and previous crystallographic studies. Proteins 2001;42:192-200. PMID- 11119644 TI - Crystal structure of a truncated form of porcine odorant-binding protein. AB - The odorant-binding proteins (OBPs) are a family of structurally related molecules that are found in high concentrations in the nasal mucus of vertebrates and bind with moderate affinity a large family of hydrophobic odorants. On the basis of their quaternary structure, the OBPs have been classified as monomers, homodimers, and heterodimers. Porcine OBP was believed for a long time to be a monomer under physiological conditions but there are recent data that support the existence of a monomer-dimer equilibrium. We have determined the crystal structure of a monoclinic form of porcine OBP and found that the truncated molecules, which lack the first 8 amino acids, pack in the cell as dimers that appear to have physiological relevance. The presence in the maps of electron density for an endogenous ligand has also let us identify the side chain of the amino acids that are at the ligand-binding site. In addition, an alternative way of access to the central cavity that binds the ligands is suggested by the particular packing of the molecules in this unit cell. Proteins 2001;42:201-209. PMID- 11119645 TI - GGDEF domain is homologous to adenylyl cyclase. AB - The GGDEF domain is detected in many prokaryotic proteins, most of which are of unknown function. Several bacteria carry 12-22 different GGDEF homologues in their genomes. Conducting extensive profile-based searches, we detect statistically supported sequence similarity between GGDEF domain and adenylyl cyclase catalytic domain. From this homology, we deduce that the prokaryotic GGDEF domain is a regulatory enzyme involved in nucleotide cyclization, with the fold similar to that of the eukaryotic cyclase catalytic domain. This prediction correlates with the functional information available on two GGDEF-containing proteins, namely diguanylate cyclase and phosphodiesterase A of Acetobacter xylinum, both of which regulate the turnover of cyclic diguanosine monophosphate. Domain architecture analysis shows that GGDEF is typically present in multidomain proteins containing regulatory domains of signaling pathways or protein-protein interaction modules. Evolutionary tree analysis indicates that GGDEF/cyclase superfamily forms a large diversified cluster of orthologous proteins present in bacteria, archaea, and eukaryotes. Proteins 2001;42:210-216. PMID- 11119646 TI - Energy landscape theory for Alzheimer's amyloid beta-peptide fibril elongation. AB - Recent experiments on the kinetics of deposition and fibril elongation of the Alzheimer's beta-amyloid peptide on preexisting fibrils are analyzed. A mechanism is developed based on the dock-and-lock scheme recently proposed by Maggio and coworkers to organize their experimental observations of the kinetics of deposition of beta-peptide on preexisting amyloid fibrils and deposits. Our mechanism includes channels for (1) a one-step prion-like direct deposition on fibrils of activated monomeric peptide in solution, and (2) a two-step deposition of unactivated peptide on fibrils and subsequent reorganization of the peptide fibril complex. In this way, the mechanism and implied "energy landscape" unify a number of schemes proposed to describe the process of fibril elongation. This beta-amyloid landscape mechanism (beta ALM) is found to be in good agreement with existing experimental data. A number of experimental tests of the mechanism are proposed. The mechanism leads to a clear definition of overall equilibrium or rate constants in terms of the energetics of the elementary underlying processes. Analysis of existing experimental data suggests that fibril elongation occurs through a two-step mechanism of nonspecific peptide absorption and reorganization. The mechanism predicts a turnover in the rate of fibril elongation as a function of temperature and denaturant concentration. Proteins 2001;42:217-229. PMID- 11119647 TI - The C-terminal domain of HPII catalase is a member of the type I glutamine amidotransferase superfamily. AB - Discovering distant evolutionary relationships between proteins requires detecting subtle similarities. Here we use a combination of sequence and structure analysis to show that the C-terminal domain of Escherichia coli HPII catalase with available spatial structure is a divergent member of the type I glutamine amidotransferase (GAT) superfamily. GAT-containing proteins include many biosynthetic enzymes such as E. coli carbamoyl phosphate synthetase and anthranilate synthase. Typical GAT domains have Rossmann fold-like topology and possess a catalytic triad similar to that of proteases. The C-terminal domain of HPII catalase has the GAT Rossmann fold but lacks the triad and therefore loses enzymatic activity. In addition, we detect significant sequence similarity between thiJ domains, some of which are known to have protease activity, and typical GAT proteins. Evolutionary tree analysis of the entire GAT superfamily indicates that the HPII catalase is more closely related to thiJ domains than to classical GAT domains and is likely to have evolved from a thiJ-like protein. This work illustrates the strength of sequence-based profile analysis techniques coupled with structural superpositions in developing an evolutionarily relevant classification of protein structures. Proteins 2001;42:230-236. PMID- 11119648 TI - A protein dissection study demonstrates that two specific hydrophobic clusters play a key role in stabilizing the core structure of the molten globule state of human alpha-lactalbumin. AB - The molten globule state of alpha-lactalbumin (alpha LA) has served as a paradigm for understanding the role of these partially folded states in protein folding. We previously showed that a peptide construct consisting of the A and B helices (residues 1-38) cross-linked to the D- and C-terminal 3(10) helices (residues 101 120) of alpha LA is capable of folding to a stable molten globule-like state. Here, we report the study of three peptide constructs that are designed to investigate the contribution two short hydrophobic sequences located near the C terminus of alpha LA make to the structure and stability of the alpha LA molten globule state. These regions of the protein have been shown to form stable non native structures in isolation. The three peptide constructs contain residues 1 38 cross-linked to three separate C-terminal peptides via the native 28-111 disulfide bond. The C-terminal peptides consist of residues 101-114, 106-120, and 106-114. The results of CD, fluorescence, ANS binding, and urea denaturation experiments indicate that constructs that lack either of the hydrophobic sequences (residues 101-105 and 115-120) are significantly less structured. These results highlight the importance of long-range, mutually stabilizing interactions within the molten globule state of the protein. Proteins 2001;42:237-242. PMID- 11119649 TI - Quantitative comparison of the ability of hydropathy scales to recognize surface beta-strands in proteins. AB - Methods based on the use of hydropathy scales have been used widely to ascertain the secondary structures of proteins. However, over 100 such scales have been reported in the literature, and which of these is the most successful in terms of the prediction rate of the correct structure is not clear. This article, therefore, reports a comprehensive analysis of the relative success of hydropathy scales to locate beta-strands on the surfaces of proteins. The technique we used is based on the technique proposed by Fraser and Parry, but it includes a modification that allows a higher rate of successful prediction and a lower rate of overprediction. We used as a basis for assessing the predictions a database of sequence-unique structures that we previously established. Proteins 2001;42:243 255. PMID- 11119650 TI - Efficient electrostatic solvation model for protein-fragment docking. AB - A method is presented for the fast evaluation of the binding energy of a protein small molecule complex with electrostatic solvation. It makes use of a fast preprocessing step based on the assumption that the main contribution to electrostatic desolvation upon ligand binding originates from the displacement of the first shell of water molecules. For a rigid protein, the precomputation of the energy contributions on a set of grids allows the estimation of the energy in solution of about 300 protein-fragment binding modes per second on a personal computer. The docking procedure is applied to five rigid binding sites whose size ranges from 17 residues to a whole protein of 107 amino acids. Using a library of 70 mainly rigid molecules, known micromolar inhibitors or close analogs are docked and prioritized correctly. The docking based rank-ordering of the library requires about 5 h and is proposed as a complementary approach to structure activity relationships by nuclear magnetic resonance. Proteins 2001;42:256-268. PMID- 11119651 TI - Structure of an intermediate in the unfolding of creatine kinase. AB - The homodimeric muscle isoform of creatine kinase (MM-CK) unfolds on exposure to low levels of guanidinium chloride (GdmCl) to yield a partly folded monomeric intermediate. Those regions of MM-CK that experience local unfolding were previously identified through an extensive study of antibody accessibility and protease sensitivity. Since these studies were completed, the coordinates of the rabbit isoform (MM-CK) were released. In light of this, we have determined the minimum changes to this structure required to explain our data on protease and epitope accessibility in the intermediate. We propose that the observed changes occur through (a) disruption of the monomer-monomer interface during dissociation, (b) separation and/ or unfolding of domains or subdomains, and (c) the partial unfolding of solvent-exposed helices. The proposed structure for the intermediate is consistent both with current models of unfolding intermediates and the results of independent studies pertaining to the unfolding of creatine kinase. Proteins 2001;42:269-278. PMID- 11119652 TI - Docking molecules by families to increase the diversity of hits in database screens: computational strategy and experimental evaluation. AB - Molecular docking programs screen chemical databases for novel ligands that fit protein binding sites. When one compound fits the site well, close analogs typically do the same. Therefore, many of the compounds that are found in such screens resemble one another. This reduces the variety and novelty of the compounds suggested. In an attempt to increase the diversity of docking hit lists, the Available Chemicals Directory was grouped into families of related structures. All members of every family were docked and scored, but only the best scoring molecule of a high-ranking family was allowed in the hit list. The identity and scores of the other members of these families were recorded as annotations to the best family member, but they were not independently ranked. This family-based docking method was compared with molecule-by-molecule docking in screens against the structures of thymidylate synthase, dihydrofolate reductase (DHFR), and the cavity site of the mutant T4 lysozyme Leu99 --> Ala (L99A). In each case, the diversity of the hit list increased, and more families of known ligands were found. To investigate whether the newly identified hits were sensible, we tested representative examples experimentally for binding to L99A and DHFR. Of the six compounds tested against L99A, five bound to the internal cavity. Of the seven compounds tested against DHFR, six inhibited the enzyme with apparent K(i) values between 0.26 and 100 microM. The segregation of potential ligands into families of related molecules is a simple technique to increase the diversity of candidates suggested by database screens. The general approach should be applicable to most docking methods. Proteins 2001;42:279-293. PMID- 11119659 TI - Congenital anomalies: a 25-year overview. AB - Expansion of the discipline of hand surgery and heightened interest in congenital problems have resulted in major advances in the treatment of congenital hand anomalies over the past 25 years. Increased experience with congenital anomalies of the hand has expanded the hand surgeon's knowledge of patterns and relationships between different anomalies resulting in new methods of classification and more logical approaches to treatment. The principles of treatment of the more common anomalies, such as syndactyly, established by prior generations of hand surgeons have been refined in details of technique. New technologies, such as distraction lengthening and free vascularized transfers, have allowed the surgeon to treat new problems and old problems in new ways. In spite of our successes, much remains to challenge hand surgeons in this new millennium, especially in the construction of joints and the expanding field of fetal surgery. PMID- 11119660 TI - Finger replantation in the United States: rates and resource use from the 1996 Healthcare Cost and Utilization Project. AB - Although finger replantation has been performed by hand surgeons in the United States for over 30 years the epidemiology of this procedure has not been studied. We used a national sampling database from the Agency for Health Care Policy and Research to summarize descriptive statistics surrounding finger replantation in the United States. This sample contained 304 cases of finger replantation performed in 1996. Mean hospital length of stay was 5.5 days and mean total charges were $20,330. Of the 906 hospitals included in this database, only 136 performed finger replantation during 1996. Of those, 60% performed only one case and 2% performed 10 or more cases. This descriptive analysis provides an examination of the largest sample size to date of finger replantation in the United States. Epidemiologic studies using national databases can provide researchers and policy makers with the analyses needed to examine treatment options and appropriately allocate future health care resources. PMID- 11119661 TI - Comparison between partial and minimal medial epicondylectomy combined with decompression for the treatment of cubital tunnel syndrome. AB - We have performed minimal medial epicondylectomy for cubital tunnel syndrome since 1990 to preserve the anterior medial collateral ligament. In this study we compared surgical outcomes between partial medial epicondylectomy (14 patients) and minimal medial epicondylectomy (18 patients) combined with ulnar nerve decompression for the treatment of cubital tunnel syndrome. Mean preoperative Yasutake scores were 57 +/- 17 points (+/-SD) in the partial epicondylectomy group and 60 +/- 15 points in the minimal medial epicondylectomy group. The postoperative scores were 79 +/- 19 points and 87 +/- 10 points, respectively. Both groups had significant improvement in their Yasutake scores following medial epicondylectomy. Similar improvements in motor conduction velocity were observed. There was no significant difference in improvement of either the Yasutake scores or the motor conduction velocity between the 2 groups. Valgus instability of the elbow was significantly greater in the partial epicondylectomy group. We therefore conclude that minimal medial epicondylectomy combined with ulnar nerve decompression is an effective treatment for cubital tunnel syndrome and that a larger excision of the medial epicondyle should be avoided. PMID- 11119662 TI - Rehabilitation of the medial collateral ligament-deficient elbow: an in vitro biomechanical study. AB - The purpose of this study was to determine the relative contribution of muscle activity and the effect of forearm position on the stability of the medial collateral ligament (MCL)-deficient elbow. Simulated active and passive elbow flexion with the forearm in both supination and pronation was performed using a custom elbow testing apparatus. Testing was first performed on intact specimens, then on MCL-deficient specimens. Elbow instability was quantified using an electromagnetic tracking device by measuring internal-external rotation and varus valgus laxity of the ulna relative to the humerus. Compared with the intact elbow, transection of the MCL, with the arm in a vertical orientation, caused a significant increase in internal-external rotation during passive elbow flexion with the forearm in pronation, but forearm supination reduced this instability. Overall, following MCL transection the elbow was more stable with the forearm in supination than pronation during passive flexion. In the pronated forearm position simulated active flexion also reduced the instability detected during passive flexion, with the arm in a varus and valgus gravity-loaded orientation. The maximum varus-valgus laxity was significantly increased with MCL transection regardless of forearm position during passive flexion. We concluded that active mobilization of the elbow with the arm in vertical orientation during rehabilitation is safe in the setting of an MCL-deficient elbow with the forearm in a fully supinated and pronated position. Splinting and passive mobilization of the MCL-deficient elbow with the forearm in supination should minimize instability and valgus elbow stresses should be avoided throughout the rehabilitation period. PMID- 11119663 TI - Forearm rotation alters interosseous ligament strain distribution. AB - Recent interest in reconstruction of the interosseous ligament (IOL) of the forearm has led to questions concerning optimal placement of the reconstructive graft as well as the ideal rotational position of the forearm during graft tensioning. We therefore studied the strain distribution in the IOL to determine which fibers are strained in different positions of forearm rotation. Five cadaveric human forearms were subjected to compressive axial load (simulating power grip) and the strain values across the entire IOL were measured with the forearm in neutral, supination, and pronation. The strain distribution in the IOL changed with forearm rotation. The highest overall strain was found in neutral. In neutral and pronation, higher strain was observed in the proximal region of the IOL. In supination, however, higher average strain was seen in the distal region of the IOL. These results suggest that a reconstructive graft placed in the proximal region of the IOL and tensioned in neutral rotation would provide balanced constraint in different positions of forearm rotation. A graft placed in the distal region and tensioned in forearm neutral, however, may limit forearm rotation. PMID- 11119664 TI - External fixation of the distal radius: to predrill or not to predrill. AB - Using both clinical and laboratory studies we investigated whether predrilling before insertion of external fixation pins is necessary for use in treating distal radius fractures. Our clinical study included 50 consecutive external fixators (4.0- and 2.5-mm pins) using 100 predrilled and 100 direct-drilled pins placed in a randomized manner. There was no increased incidence of pin track infection or other pin problem with the direct-drilled technique. There were, however, significantly elevated temperatures with the direct-drilled technique. We therefore recommend predrilling even though the temperature differences in this bone with this fixator were not clinically evident. PMID- 11119665 TI - Dorsal perilunate dislocations and fracture-dislocations: questionnaire, clinical, and radiographic evaluation. AB - Twenty-two consecutive patients (23 wrists) underwent open reduction internal fixation of dorsal perilunate dislocations and fracture-dislocations through combined dorsal and volar approaches. One of 5 experienced wrist surgeons performed these procedures within an average of 3 days of injury (range, 0-26 days) and intercarpal fixation was kept within the proximal carpal row. Motion was instituted an average of 10 weeks (range, 5-16 weeks) after injury. All patients were males. The average age at the time of injury was 32 years (range, 16-60 years). The average follow-up period was 37 months (range, 13-65 months). Average flexion-extension motion arc and grip strength in the injured wrist were 57% and 73%, respectively, compared with the contralateral wrist. The scapholunate angle increased and the revised carpal height ratio decreased over time, which was statistically significant for both measurements. Three patients (3 wrists) required wrist arthrodesis and a fourth patient had an immediate scaphoid excision and 4-corner arthrodesis secondary to an irreparable scaphoid fracture. One patient required a proximal row carpectomy to treat septic arthritis. Nine of the remaining 18 wrists had radiographic evidence of arthritis, most often at the capitolunate or scaphocapitate articulations. Short form-36 mental summary scores were significantly greater than age- and gender matched US population values; physical summary scores were significantly less. The disabilities of arm, shoulder, and hand evaluation, Mayo wrist score, and patient-rated wrist evaluation all reflected loss of function. Seventy-three percent of all patients had returned to full duties in their usual occupations and a total of 82% were employed. PMID- 11119666 TI - Modification of the Sauve-Kapandji procedure with extensor carpi ulnaris tenodesis. AB - The Sauve-Kapandji procedure is a useful treatment option for osteoarthritis of the distal radioulnar joint. Recent reports of a painful unstable proximal ulnar stump prompted us to develop a method of stabilizing the proximal stump of the ulna during the Sauve-Kapandji procedure by using a half-slip of the extensor carpi ulnaris. Thirteen osteoarthritic wrists (8 primary and 5 traumatic) in 8 men and 5 women with an average age of 50 years were treated by this method. The length of the follow-up periods averaged 36 months. Pain improved in all patients after surgery but pain was elicited over 1 ulnar stump by direct pressure. Both pronation/supination and flexion/extension had statistically significant improvement with the exception of flexion. Grip strength improved in all wrists after surgery. Postoperative x-rays improved alignment in both coronal and lateral planes. Stabilization of the proximal ulnar stump associated with Sauve Kapandji procedure is a useful procedure to prevent an unstable ulnar stump in the treatment of osteoarthritis of the distal radioulnar joint. PMID- 11119667 TI - Material properties of the trapezial and trapeziometacarpal ligaments. AB - Destabilization of the trapezium from its normal orientation with respect to the trapezoid, second metacarpal, and thumb metacarpal leads to incongruity at the trapeziometacarpal (TMC) joint. Abnormal shear forces may eventually result in TMC joint arthritis. By determining the relative stiffness and strength of the ligaments that stabilize this joint, one may infer their role in providing stability to the TMC joint. This study addresses the material properties of the ligaments stabilizing the trapezium and TMC joint to better understand the mechanics and kinematics of this joint. Fresh-frozen cadaveric hands (10 males and 10 females) were used to obtain bone-ligament-bone complexes from the dorsal and volar trapeziotrapezoid ligaments, dorsal and volar trapezio-second metacarpal ligaments, anterior oblique ligament, dorsoradial ligament, and trapezio-third metacarpal (T-III MC) ligament. The following material properties were derived from our data: ultimate load, ultimate stress (normalized failure load), ultimate strain (percent elongation), stiffness, toughness (energy to failure), and hysteresis. The dorsoradial ligament demonstrated the greatest ultimate load and toughness (energy to failure). The T-III MC ligament demonstrated the greatest ultimate stress (normalized failure load) and stiffness. The anterior oblique ligament demonstrated the least stiffness and the greatest hysteresis. The material properties of capsuloligamentous structures may be a good indicator of their importance to joint stability. Using these criteria we conclude that the T-III MC and dorsoradial ligaments are important stabilizers of the trapezium and TMC joint, respectively. These two ligaments were found to be the strongest, stiffest, and toughest ligaments, while the anterior oblique ligament was relatively weak and compliant. The dorsal trapezio-second metacarpal, volar trapezio-second metacarpal, and T-III MC ligaments were all relatively strong and are anatomically aligned to function as tension bands to restrain the trapezium against cantilever bending forces applied to it by the thumb during key or tip pinch. PMID- 11119668 TI - Scaphocapitate syndrome in an adolescent. AB - Scaphocapitate syndrome is a rare injury and its incidence in immature skeleton is not well documented. We describe our experience of scaphocapitate syndrome in a 12-year-old boy and report the results after a 3-year follow-up period. Treatment involved open reduction internal fixation using K-wires. The 3-year follow-up evaluation revealed no evidence of avascular necrosis. The wrist was completely asymptomatic and the patient used it normally. The difficulty in examining an injured child and the presence of open physes on radiographs can make diagnosis difficult. Awareness among orthopedic surgeons about this injury in children is needed to avoid misdiagnosis and to initiate timely treatment. PMID- 11119669 TI - Treatment of Eaton stage I trapeziometacarpal disease with thumb metacarpal extension osteotomy. AB - The current benchmark for the treatment of Eaton stage I disease of the trapeziometacarpal (TMC) joint includes palmar oblique ligament reconstruction and reflects its primary role in providing stability during lateral pinch. This study prospectively evaluates the efficacy of an alternative extra-articular approach using a 30 degrees extension osteotomy of the thumb metacarpal to redistribute trapeziometacarpal contact area and load, obviating the need for ligament reconstruction. Preoperative and postoperative subjective and objective data are reported for 12 patients enrolled in the study between 1995 and 1998. Trapeziometacarpal arthrotomy allowed accurate intra-articular assessment and verified palmar oblique ligament incompetence in each case. The average follow-up period was 2.1 years (range, 6-46 months). All osteotomies healed at an average of 7 weeks. Eleven patients were satisfied with outcome. Grip and pinch strength increased an average of 8.5 and 3.0 kg, respectively. Thumb metacarpal extension osteotomy is an effective biomechanical alternative to ligament reconstruction in the treatment of Eaton stage I disease of the trapeziometacarpal joint. PMID- 11119670 TI - The sagittal band: anatomic and biomechanical study. AB - Forty-eight digits from 12 human adult fresh-frozen and formalin-preserved cadaveric hands were used to study the anatomy and biomechanics of the sagittal band (SB) and to investigate the mechanism of its injury. The SB was observed to be part of a complex retinacular system in proximity to the metacarpophalangeal (MCP) joint collateral ligaments and the palmar plate. Dynamic changes in SB fiber orientation were observed with different positions of the MCP and wrist joints. The fibers were perpendicular (0 degrees ) to the extensor tendon in neutral position, distally angulated 25 degrees at 45 degrees of MCP flexion, and 55 degrees with full flexion. Swan-Ganz catheter measurements were obtained deep to the SB in varying positions of the MCP joint. The average pressure generation was greatest (50 mm Hg) during full MCP joint flexion and least (30 mm Hg) during 45 degrees flexion. When MCP joint radial or ulnar deviation was added the average measurement was greatest (57) in neutral MCP position and least (35 mm Hg) in 45 degrees flexion. Serial sectioning of the ulnar SB produced no extensor tendon instability. Partial proximal but not distal sectioning of the radial SB produced tendon subluxation. Complete sectioning of the radial SB produced tendon dislocation. Wrist flexion increased tendon instability after radial SB sectioning. We conclude that (1) extensor tendon instability following SB disruption is most common in the long finger and least common in the small finger; (2) ulnar instability of the extensor tendon is due to partial or complete radial SB disruption, (3) the degree of extensor tendon instability is determined by the extent of SB disruption, (4) proximal rather than distal SB compromise contributes to extensor tendon instability, (5) great forces are inflicted on the SB while the MCP joint is in full extension or less frequently in full flexion, which may be the mechanism of its injury, and (6) wrist flexion contributes to extensor tendon instability after SB disruption and may exacerbate the severity of its injury. PMID- 11119671 TI - Examination of the anatomic relationship of the proximal germinal nail matrix to the extensor tendon insertion. AB - The purpose of this study was to delineate the relationship of the terminal extensor tendon insertion to the proximal limit of the germinal nail matrix. Sixteen fresh-frozen human cadaver fingers without any evidence of trauma (average age, 55 years; 3 males and 1 female) were used for this study. Under x25 magnification the proximal limit of the germinal nail matrix and the terminal bony insertion of the extensor tendon were identified. The distance from the terminal tendon insertion to the germinal nail matrix was ascertained using precision calipers. The average distance from the terminal extensor tendon insertion to the proximal edge of the germinal nail matrix was found to be 1.2 mm. We conclude that the proximal limit of the germinal matrix is extremely close to the terminal extensor tendon bony insertion. When the extensor tendon insertion is visualized during operative exposures of the dorsum of the distal phalanx, care should be taken to avoid damaging the germinal matrix. Conversely, when the nail bed is being completely excised, visualization of the insertion of the extensor tendon will indicate that further proximal dissection is not required. PMID- 11119672 TI - Conservative management of zone II partial flexor tendon lacerations greater than half the width of the tendon. AB - Over a 5-year-period 15 patients with zone II partial flexor tendon lacerations that were larger than half the width of the tendon were treated conservatively without tendon suturing. Surgical exploration was done with a digital block and the flexor tendons were observed as the patient fully extended and flexed the finger. If present, the cause of triggering was determined and eliminated by trimming any beveled tendon edge, resection of the involved pulleys, and repair of the flexor sheath. Early protected mobilization was started the first day after injury using a dorsal splint. At 4 weeks after injury the splint was removed and exercises against resistance were started. None of the patients had triggering or rupture of the flexor tendons. Using the Strickland-Glogovac evaluation method, results were excellent in 93% of cases and good in the remaining 7%. It was concluded that conservative management of zone II partial flexor tendon lacerations larger than half the width of the tendon is safe as long as certain guidelines regarding the prevention of triggering and protected mobilization are applied. PMID- 11119673 TI - A comparison of four repair techniques for Camper's chiasma flexor digitorum superficialis lacerations: tested in an in vitro model. AB - The relative strengths of 4 methods for repair of the flexor digitorum superficialis tendon were examined in 14 fresh-frozen cadaver hands (40 tendons). All tendons underwent sharp zone II transection at Camper's chiasma. All transections were repaired with 4.0 Ethibond (Ethicon Inc, Sommerville, NJ) using modified Becker, modified Kessler, horizontal mattress, or simple sutures. Flexion of the repaired digit at a constant excursion rate was rendered up to tendon rupture. The modified Becker technique withstood breaking forces (57.9 N) significantly greater than the other techniques examined. Forces up to 34 N have been measured in vivo during unresisted active finger motion. Thus, the modified Becker technique appears to provide adequate strength for early active flexor digitorum superficialis motion. PMID- 11119674 TI - Cyclical testing of zone II flexor tendon repairs. AB - Kessler, Strickland, or modified Becker repairs, all augmented with a running circumferential epitenon suture, were performed for simulated zone II flexor tendon lacerations in the index, long, and ring fingers of 12 fresh-frozen cadaveric specimens. Each hand was tested with a tensiometer built for curvilinear testing of human flexor tendons in an intact hand. Each tendon was cycled 100 times, then examined for gapping before testing to failure. Maximum load to failure, including tendon load and pinch force, was recorded for each tendon. We propose that combining the advantages of cyclical testing and a curvilinear model is the most effective way of testing flexor tendon repairs capable of undergoing an early active motion protocol. None of the repaired tendons failed during the cyclic portion of testing. The average gapping after cycling for the 3 suture techniques was 0.12 +/- 0.35 mm for the Kessler technique, 0. 00 +/- 0.00 mm for the Strickland technique, and 0.19 +/- 0.26 mm for the modified Becker technique. The maximum tendon loads to failure were 33.8 +/- 6.8 N for the Kessler technique, 30.4 +/- 5.64 N for the Strickland technique, and 76.3 +/- 9.02 N for the modified Becker technique. There was a statistically significant difference between the modified Becker repair and the other 2 repairs for maximum tendon load and pinch force to failure. The results of this study show that all 3 tendon repair techniques can withstand forces reported with passive motion, but only the modified Becker repair allows sufficient strength above those forces that are estimated for active motion during tendon healing. PMID- 11119675 TI - Intratendinous rupture of a flexor tendon graft many years after staged reconstruction: a report of three cases. AB - Three cases of rupture of a flexor tendon graft many years after surgery are presented. Two cases occurred 12 years after reconstruction and the third case occurred 21 years after reconstruction. Each rupture was intratendinous, just proximal to the flexor tendon sheath in 2 cases and at the proximal edge of the transverse carpal ligament in the third case. Active digital flexion was restored by transfer of the flexor digitorum superficialis from an adjacent finger to the distal tendon stump or by direct end-to-end repair of the rupture site reinforced with an onlay autogenous patch graft. Patients undergoing tendon grafting should be alerted to the possibility of rupture, even many years later. PMID- 11119676 TI - Kinematic assessment of manual skill following functional hand surgery in tetraplegia. AB - To determine whether surgical key grip reinforcement actually leads to a better movement ability we developed a procedure for the kinematic analysis of manual skill following hand surgery in tetraplegia. The functional results of surgery in 5 cases were examined by the kinematic analysis of drawing movements using an electronic pen and a digitizer under 3 conditions: with eyes open, with eyes closed, and while performing a concurrent arithmetic task. Movement velocity and dysfluency (ie, the number of velocity changes per centimeter) were measured before and at several moments after surgery during subsequent rehabilitation. Both movement velocity and dysfluency showed good stability across repeated trials and were consistently affected by visual deprivation. Movement velocity showed a 39% increment between the first and last assessment. Although grip strength increased in all patients, it was not associated with the change of movement velocity. These results suggest that other factors (eg, deep sensibility, cognition, muscle coordination) play a critical role in the ability to use improved grip force for controlling drawing movements and emphasize the value of a kinematic assessment besides measuring isolated grip force in the evaluation of functional hand surgery. PMID- 11119677 TI - Surgery on the affected upper extremity of patients with a history of complex regional pain syndrome: a retrospective study of 100 patients. AB - Surgery on the extremity affected with complex regional pain syndrome (CRPS) is generally avoided because of the risk that the symptoms will recur or worsen. Perioperative sympathectomy or stellate ganglion block has previously been recommended for CRPS patients requiring surgery of the affected upper extremity. We evaluated 100 patients with a history of upper extremity CRPS undergoing surgery on the affected extremity. All signs and symptoms of CRPS had resolved before surgery. After completion of the surgical procedure half of the patients (n = 50) underwent a stellate ganglion block; the other half received no intervention. The recurrence rate of CRPS was significantly lower in those patients receiving a postoperative stellate ganglion block (n = 5; 10%) compared with those receiving no intervention (n = 36; 72%). We conclude that performing a perioperative stellate ganglion block in patients with a history of CRPS can significantly reduce the recurrence rate of this disease process. PMID- 11119678 TI - Reflex sympathetic dystrophy: misdiagnosis in patients with dysfunctional postures of the upper extremity. AB - The purpose of this case-control study was to assess the frequency of the inappropriate diagnosis of reflex sympathetic dystrophy (RSD) in patients who presented with dysfunctional postures of the upper extremity (n = 43). This group of patients with a dysfunctional posture was compared with a randomly selected control group of patients who presented with pain but no dysfunctional posture (n = 88). The patients underwent radiographic evaluation after review of previous medical records and history and physical examination. Patients with dysfunctional postures had a significantly higher frequency (63%) of a previous inappropriate diagnosis of RSD compared with the control group (6%). None of the patients in either group had objective findings consistent with a diagnosis of RSD. Patients presenting with dysfunctional postures of the upper extremity may be misdiagnosed as having RSD and rarely meet the criteria for this diagnosis. PMID- 11119679 TI - The injection of nodules of Dupuytren's disease with triamcinolone acetonide. AB - Over a 4-year period 63 patients (75 hands) with Dupuytren's nodules were treated with a series of injections with the steroid triamcinolone acetonide directly into the area of disease. The purpose of this study was to determine whether intralesional injections of triamcinolone acetonide could produce softening and flattening in nodules of Dupuytren's disease as seen in the intralesional injections of hypertrophic scars and keloids. After an average of 3.2 injections per nodule 97% of the hands showed regression of disease as exhibited by a softening or flattening of the nodule(s). Although some patients had complete resolution of the nodules, most experienced definite but incomplete resolution of the nodules in the range of 60% to 80%. Although a few patients did not experience recurrence or reactivation of the disease in the injected nodules or development of new nodules, 50% of patients did experience reactivation of disease in the nodules 1 to 3 years after the last injection, necessitating 1 or more injections. The findings of this study indicate that the intralesional injection of nodules of Dupuytren's disease with triamcinolone acetonide may modify the progression of the disease. PMID- 11119680 TI - Lymphangiosarcoma (Stewart-Treves syndrome) in postmastectomy patients. AB - Stewart-Treves syndrome (STS) is a rare but aggressive upper extremity lymphangiosarcoma in postmastectomy patients. Unfamiliarity with this disease and the innocuous appearance of the tumor often lead to delayed diagnosis. A comprehensive search of the databases at a single tertiary-care academic institution revealed only 3 cases of STS in the last 63 years. The latency time between breast cancer treatment and diagnosis of STS was 11 to 21 years. Survival after diagnosis of STS ranged from 8 to 15 months. One patient underwent radical surgery. The extensive lymphangiosarcoma in the other 2 patients precluded surgical resection and they underwent chemotherapy. All patients had adjuvant radiation therapy at the time of the original breast cancer resection. This report includes a discussion of the epidemiology, etiology, presentation, treatment, and prognosis of STS. PMID- 11119681 TI - Metastatic thymic carcinoma in a digit: a case report. AB - Acrometastases are a rare but important clinical entity. We present the case of a 54-year-old man with a metastasis to a digit from a primary thymic carcinoma. The prognostic implications of such a diagnosis are discussed. PMID- 11119682 TI - Publishing (whatever that means) neuroscience in the new millennium. AB - The World Wide Web and other forms of digital communication have led some to predict the demise of printed journals. In my experience, it is clearly demonstrable that scientific articles can be more efficiently reviewed and edited by digital document sharing. Nevertheless, high quality print journals will remain the preferred scholarly venue for authors's best works for some time to come. However, while innovative means to share raw data, validate observations, and disseminate scientific information are emerging, no system of 'publishing' will serve the community optimally unless the ethical behavior of human scientists can be maintained and appropriately rewarded. PMID- 11119683 TI - Sodium channels and their genes: dynamic expression in the normal nervous system, dysregulation in disease states(1). AB - Although classical neurophysiological doctrine rested on the concept of the sodium channel, it is now clear that there are nearly a dozen sodium channel genes, each encoding a molecularly distinct channel. Different repertoires of channels endow different types of neurons with distinct transduction and encoding properties. Sodium channel expression is highly dynamic, exhibiting plasticity at both the transcriptional and post-transcriptional levels. In some types of neurons within the normal nervous system, e.g. hypothalamic magnocellular neurosecretory neurons, changes in sodium channel gene expression occur in association with the transition from a quiescent to a bursting state; these changes are accompanied by the insertion of a different set of sodium channel subtypes in the cell membrane, a form of molecular plasticity which results in altered electrogenic properties. Dysregulation of sodium channel genes has been observed in a number of disease states. For example, transection of the peripheral axons of spinal sensory neurons triggers down-regulation of some sodium channel genes, and up-regulation of other sodium channel genes; the resultant changes in sodium channel expression contribute to hyperexcitability that can lead to chronic pain. There is also evidence, in experimental models of demyelination and in post-mortem tissue from patients with multiple sclerosis, for dysregulation of sodium channel gene expression in the cell bodies of some neurons whose axons have been demyelinated, suggesting that an acquired channelopathy may contribute to the pathophysiology of demyelinating diseases such as multiple sclerosis. The dynamic nature of sodium channel gene expression makes it a complex topic for investigation, but it also introduces therapeutic opportunities, since subtype-specific sodium channel modulating drugs may soon be available. PMID- 11119684 TI - Nitric oxide, impulse activity, and neurotrophins in visual system development(1). AB - Topographic refinement of synaptic connections within the developing visual system involves a variety of molecules which interact with impulse activity in order to produce the precise retinotopic maps found in the adult brain. Nitric oxide (NO) has been implicated in this process, as have various growth factors. Within the subcortical visual system, we have recently shown that nitric oxide contributes to pathway refinement in the superior colliculus (SC). Long-term potentiation (LTP) and long-term depression (LTD) are also expressed in SC during the time that this pathway undergoes refinement. The role of NO has been demonstrated by showing that refinement of ipsilateral fibers in the retinocollicular pathway is significantly delayed in gene knockout mice in which both the endothelial and neuronal isoforms of nitric oxide synthase (NOS) have been disrupted. The effect also depends upon Ca(2+) channels because refinement of both the ipsilateral retinocollicular and retinogeniculate pathways is disrupted in genetic mutants in which the beta3 subunit of the Ca(2+) channel has been deleted. LTD may also be involved in this process, because the time course of its expression correlates with that of pathway refinement and LTD magnitude is depressed by nitrendipine, an L-type Ca(2+) channel blocker. LTP is also expressed during early postnatal development in the LGN and SC and may contribute to synaptic stabilization. The role of neurotrophins in pathway refinement in the visual system is also reviewed. PMID- 11119685 TI - How the brain uses time to represent and process visual information(1). AB - Information theory provides a theoretical framework for addressing fundamental questions concerning the nature of neural codes. Harnessing its power is not straightforward, because of the differences between mathematical abstractions and laboratory reality. We describe an approach to the analysis of neural codes that seeks to identify the informative features of neural responses, rather than to estimate the information content of neural responses per se. Our analysis, applied to neurons in primary visual cortex (V1), demonstrates that the informative precision of spike times varies with the stimulus modality being represented. Contrast is represented by spike times on the shortest time scale, and different kinds of pattern information are represented on longer time scales. The interspike interval distribution has a structure that is unanticipated from the firing rate. The significance of this structure is not that it contains additional information, but rather, that it may provide a means for simple synaptic mechanisms to decode the information that is multiplexed within a spike train. Extensions of this analysis to the simultaneous responses of pairs of neurons indicate that neighboring neurons convey largely independent information, if the decoding process is sensitive to the neuron of origin and not just the average firing rate. In summary, stimulus-related information is encoded into the precise times of spikes fired by V1 neurons. Much of this information would be obscured if individual spikes were merely taken to be estimators of the firing rate. Additional information would be lost by averaging across the responses of neurons in a local population. We propose that synaptic mechanisms sensitive to interspike intervals and dendritic processing beyond simple summation exist at least in part to enable the brain to take advantage of this extra information. PMID- 11119686 TI - Neuroprotective signaling and the aging brain: take away my food and let me run. AB - It is remarkable that neurons are able to survive and function for a century or more in many persons that age successfully. A better understanding of the molecular signaling mechanisms that permit such cell survival and synaptic plasticity may therefore lead to the development of new preventative and therapeutic strategies for age-related neurodegenerative disorders. We all know that overeating and lack of exercise are risk factors for many different age related diseases including cardiovascular disease, diabetes and cancers. Our recent studies have shown that dietary restriction (reduced calorie intake) can increase the resistance of neurons in the brain to dysfunction and death in experimental models of Alzheimer's disease, Parkinson's disease, Huntington's disease and stroke. The mechanism underlying the beneficial effects of dietary restriction involves stimulation of the expression of 'stress proteins' and neurotrophic factors. The neurotrophic factors induced by dietary restriction may protect neurons by inducing the production of proteins that suppress oxyradical production, stabilize cellular calcium homeostasis and inhibit apoptotic biochemical cascades. Interestingly, dietary restriction also increases numbers of newly-generated neural cells in the adult brain suggesting that this dietary manipulation can increase the brain's capacity for plasticity and self-repair. Work in other laboratories suggests that physical and intellectual activity can similarly increase neurotrophic factor production and neurogenesis. Collectively, the available data suggest the that dietary restriction, and physical and mental activity, may reduce both the incidence and severity of neurodegenerative disorders in humans. A better understanding of the cellular and molecular mechanisms underlying these effects of diet and behavior on the brain is also leading to novel therapeutic agents that mimick the beneficial effects of dietary restriction and exercise. PMID- 11119687 TI - Alzheimer's disease: a dysfunction of the amyloid precursor protein(1). AB - In this review, we argue that at least one insult that causes Alzheimer's disease (AD) is disruption of the normal function of the amyloid precursor protein (APP). Familial Alzheimer's disease (FAD) mutations in APP cause a disease phenotype that is identical (with the exception that they cause an earlier onset of the disease) to that of 'sporadic' AD. This suggests that there are molecular pathways common to FAD and sporadic AD. In addition, all individuals with Down syndrome, who carry an extra copy of chromosome 21 and overexpress APP several fold in the brain, develop the pathology of AD if they live past the age of 40. These data support the primacy of APP in the disease. Although APP is the source of the beta-amyloid (Abeta) that is deposited in amyloid plaques in AD brain, the primacy of APP in AD may not lie in the production of Abeta from this molecule. We suggest instead that APP normally functions in the brain as a cell surface signaling molecule, and that a disruption of this normal function of APP is at least one cause of the neurodegeneration and consequent dementia in AD. We hypothesize in addition that disruption of the normal signaling function of APP causes cell cycle abnormalities in the neuron, and that these abnormalities constitute one mechanism of neuronal death in AD. Data supporting these hypotheses have come from investigations of the molecular consequences of neuronal expression of FAD mutants of APP or overexpression of wild type APP, as well as from identification of binding proteins for the carboxyl-terminus (C terminus) of APP. PMID- 11119688 TI - Effect of intrathecal galanin and its putative antagonist M35 on pain behavior in a neuropathic pain model. AB - There is currently some debate over a possible role of galanin in pain processing. It was recently reported that the levels of galanin in dorsal root ganglia (DRGs) seem related to development of allodynia after unilateral sciatic nerve constriction injury. In our present study, we aimed at characterizing the effect of exogenous and endogenous galanin on pain behavior in allodynic and non allodynic rats in which the levels of galanin in DRG neurons are low and high, respectively [28]. The results show that in allodynic rats, the mechanical threshold increases dose-dependently after intrathecal (i.t.) injection of galanin, while no significant changes were observed in groups treated with the putative galanin antagonist M35 or saline. In non-allodynic rats i.t. injection of M35 induced a significant mechanical allodynic state, which did not occur after injection of galanin, bradykinin, the bradykinin fragment(2-9) or saline. The results suggest that in the present experimental paradigm exogenous galanin has an anti-allodynic effect in the allodynic rats, and that endogenous galanin has a tonic inhibitory effect in the non-allodynic group. PMID- 11119689 TI - A comparison of the expression and properties of Apaf-1 and Apaf-1L. AB - Apaf-1 is a mammalian homolog of CED-4 that regulates cell death by participating in a ternary complex with cytochrome c, and procaspase-9. In the case of CED-4, two splice variants exist. The smaller (CED-4S) is proapoptotic while the larger (CED-4L) contains a short in-frame insert and is anti-apoptotic. We cloned a murine variant of apaf-1, termed apaf-1L, which contains an eleven amino acid insert similar to a recently described human apaf-1L clone. apaf-1 and apaf-1L have similar distributions in adult and fetal tissues, although apaf-1L transcripts are more abundant. Apaf-1L, undergoes homomerization and heteromerization with Apaf-1 in yeast. Apaf-1L also binds to caspase-9 and a dominant-negative isoform of caspase-9. Unlike CED-4, neither Apaf-1 variant was lethal in yeast. However, both Apaf-1 and Apaf-1L elicit cell death when cotransfected with caspase-9 into 293 EBNA cells. Although Apaf-1L was more potent than Apaf-1, their biological properties were qualitatively similar. PMID- 11119690 TI - Towards a neuroprotective gene therapy for Parkinson's disease: use of adenovirus, AAV and lentivirus vectors for gene transfer of GDNF to the nigrostriatal system in the rat Parkinson model. AB - During the last few years, recombinant viral vectors derived from adenovirus (Ad), adeno-associated virus (AAV) or lentivirus (LV) have been developed into highly effective vehicles for gene transfer to the adult central nervous system. In recent experiments, in the rat model of Parkinson's disease, all three vector systems have been shown to be effective for long-term delivery of glial cell line derived neurotrophic factor (GDNF) at biologically relevant levels in the nigrostriatal system. Injection of the GDNF encoding vectors into either striatum or substantia nigra thus makes it possible to obtain a regionally restricted over expression of GDNF within the nigrostriatal system that is sufficient to block the toxin-induced degeneration of the nigral dopamine neurons. Injection of GDNF vectors in the striatum, in particular, is effective not only in rescuing the cell bodies in the substantia nigra, but also in preserving the nigrostriatal projection and a functional striatal dopamine innervation in the rat Parkinson model. Long-term experiments using AAV-GDNF and LV-GDNF vectors show, moreover, that sustained GDNF delivery over 3-6 months can promote regeneration and significant functional recovery in both 6-OHDA-lesioned rats and MPTP-lesioned monkeys. The impressive efficacy of the novel AAV and LV vectors in rodent and primate Parkinson models suggests that the time may now be ripe to explore these vector systems as tools for neuroprotective treatments in patients with Parkinson's disease. PMID- 11119691 TI - L-type Ca(2+) channel-mediated Zn(2+) toxicity and modulation by ZnT-1 in PC12 cells. AB - In view of evidence that Zn(2+) neurotoxicity contributes to some forms of pathological neuronal death, we developed a model of Zn(2+) neurotoxicity in a cell line amenable to genetic manipulations. Exposure to 500 microM ZnCl(2) for 15 min under depolarizing conditions resulted in modest levels of PC12 cell death, that was reduced by the L-type Ca(2+) channel antagonist, nimodipine, and increased by the L-type Ca(2+) channel opener, S(-)-Bay K 8644. At lower insult levels (200 micrometer Zn(2+)+Bay K 8644), Zn(2+)-induced death appeared apoptotic under electron microscopy and was sensitive to the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-CH(2)F (Z-VAD); at higher insult levels (1000 microM+Bay K 8644), cells underwent necrosis insensitive to Z-VAD. To test the hypothesis that the plasma membrane transporter, ZnT-1, modulates Zn(2+) neurotoxicity, we generated stable PC12 cell lines overexpressing wild type or dominant negative forms of rat ZnT-1 (rZnT-1). Clones T9 and T23 overexpressing wild type rZnT-1 exhibited enhanced Zn(2+) efflux and reduced vulnerability to Zn(2+)-induced death compared to the parental line, whereas clones D5 and D16 expressing dominant negative rZnT-1 exhibited the opposite characteristics. PMID- 11119692 TI - Cellular bases of functional brain imaging: insights from neuron-glia metabolic coupling. PMID- 11119693 TI - Cerebral hemisphere regulation of motivated behavior. AB - The goals of this article are to suggest a basic wiring diagram for the motor neural network that controls motivated behavior, and to provide a model for the organization of cerebral hemisphere inputs to this network. Cerebral projections mediate voluntary regulation of a behavior control column in the ventromedial upper brainstem that includes (from rostral to caudal) the medial preoptic, anterior hypothalamic, descending paraventricular, ventromedial, and premammillary nuclei, the mammillary body, and finally the substantia nigra and ventral tegmental area. The rostral segment of this column is involved in controlling ingestive (eating and drinking) and social (defensive and reproductive) behaviors, whereas the caudal segment is involved in controlling general exploratory or foraging behaviors (with locomotor and orienting components) that are required for obtaining any particular goal object. Virtually all parts of the cerebral hemispheres contribute to a triple descending projection - with cortical excitatory, striatal inhibitory, and pallidal disinhibitory components - to specific parts of the behavior control column. The functional dynamics of this circuitry remain to be established. PMID- 11119694 TI - The hippocampal lamella hypothesis revisited. AB - We have re-examined the hippocampal lamellar organization of the CA3-to-CA1 connection. Based on a new technique with electrophysiological quantification of Schaffer collateral density, and a review of recent literature, we conclude that the lamellar organization remains a useful concept for understanding hippocampal connectivity. Using a sheet-like hippocampal preparation, containing the whole CA1 region, we mapped the distribution of Schaffer collaterals by two procedures. First, we recorded the amplitude of the Schaffer compound action potential in various parts of CA1 after stimulation of a point in CA3. Second, we charted the CA1 positions from which we could antidromically excite individual CA3 neurones. Although the Schaffer collaterals radiated from their CA3 cells of origin within a wide, fan-shaped area, covering a large part of the septo-temporal extent of CA1, the amplitude of the compound action potential was largest in a slightly oblique, transverse band across the CA1 towards the subicular region. PMID- 11119695 TI - The neurobiology of stress: from serendipity to clinical relevance. AB - The hormones and other physiological agents that mediate the effects of stress on the body have protective and adaptive effects in the short run and yet can accelerate pathophysiology when they are over-produced or mismanaged. Here we consider the protective and damaging effects of these mediators as they relate to the immune system and brain. 'Stress' is a principle focus, but this term is rather imprecise. Therefore, the article begins by noting two new terms, allostasis and allostatic load that are intended to supplement and clarify the meanings of 'stress' and 'homeostasis'. For the immune system, acute stress enhances immune function whereas chronic stress suppresses it. These effects can be beneficial for some types of immune responses and deleterious for others. A key mechanism involves the stress-hormone dependent translocation of immune cells in the blood to tissues and organs where an immune defense is needed. For the brain, acute stress enhances the memory of events that are potentially threatening to the organism. Chronic stress, on the other hand, causes adaptive plasticity in the brain, in which local neurotransmitters as well as systemic hormones interact to produce structural as well as functional changes, involving the suppression of ongoing neurogenesis in the dentate gyrus and remodelling of dendrites in the Ammon's horn. Under extreme conditions only does permanent damage ensue. Adrenal steroids tell only part of the story as far as how the brain adapts, or shows damage, and local tissue modulators - cytokines for the immune response and excitatory amino acid neurotransmitters for the hippocampus. Moreover, comparison of the effects of experimenter-applied stressors and psychosocial stressors show that what animals do to each other is often more potent than what experimenters do to them. And yet, even then, the brain is resilient and capable of adaptive plasticity. Stress-induced structural changes in brain regions such as the hippocampus have clinical ramifications for disorders such as depression, post-traumatic stress disorder and individual differences in the aging process. PMID- 11119696 TI - The AMPAR subunit GluR2: still front and center-stage. AB - Abnormal influx of Ca(2+) through AMPA-type glutamate receptors (AMPARs) is thought to contribute to the neuronal death associated with a number of brain disorders. AMPARs exist as both Ca(2+)-impermeable and Ca(2+)-permeable channels. AMPARs are encoded by four genes designated GluR1 (GluR-A) through GluR4 (GluR D). The presence of the GluR2 subunit renders heteromeric AMPA receptor assemblies Ca(2+)-impermeable. Molecular diversity of AMPARs under physiological and pathological conditions is generated by differential spatio-temporal patterns of GluR expression, by alternative RNA splicing and editing and by targeting and trafficking of receptor subunits at dendritic spines. The GluR2 gene is under transcriptional control by the RE1 element specific transcription factor, a gene silencing factor which renders it neuron-specific. GluR2 transcripts are edited by ADAR2 (double-stranded RNA-specific editase 1). AMPAR targeting and trafficking to spines are regulated by synaptic activity and are critical to synaptic plasticity. Recent studies involving animal models of transient forebrain ischemia and epilepsy show that GluR2 mRNA and GluR2 subunit expression are downregulated in vulnerable neurons prior to cell death. Ca(2+) imaging and electrical recording from individual pyramidal neurons in hippocampal slices reveal changes in AMPAR functional properties after ischemia. In slices from post ischemia animals, CA1 neurons with robust action potentials exhibit greatly enhanced AMPA-elicited rises in intracellular Ca(2+). Excitatory postsynaptic currents in post-ischemic CA1 exhibit an enhanced Ca(2+)-dependent component that appears to be mediated by Ca(2+)-permeable AMPARs. These studies provide evidence for Ca(2+) influx through AMPARs in neurons destined to die. To examine whether acute GluR2 downregulation, even in the absence of a neurological insult, can induce neuronal death, we performed knockdown experiments in rats and gerbils with antisense oligonucleotides targeted to GluR2 mRNA. GluR2 antisense oligonucleotide induced neuronal cell death of pyramidal neurons and enhanced pathogenicity of brief ischemic episodes. These observations provide evidence for Ca(2+) influx through AMPARs in neurons destined to die and implicate Ca(2+) permeable AMPARs in the pathogenesis of ischemia-induced neuronal death. PMID- 11119697 TI - Why do we sleep? AB - Slow-wave sleep consists in slowly recurring waves that are associated with a large-scale spatio-temporal synchrony across neocortex. These slow-wave complexes alternate with brief episodes of fast oscillations, similar to the sustained fast oscillations that occur during the wake state. We propose that alternating fast and slow waves consolidate information acquired previously during wakefulness. Slow-wave sleep would thus begin with spindle oscillations that open molecular gates to plasticity, then proceed by iteratively 'recalling' and 'storing' information primed in neural assemblies. This scenario provides a biophysical mechanism consistent with the growing evidence that sleep serves to consolidate memories. PMID- 11119698 TI - The intrinsic function of a motor system--from ion channels to networks and behavior. AB - The forebrain, brainstem and spinal cord contribution to the control of locomotion is reviewed in this article. The lamprey is used as an experimental model since it allows a detailed cellular analysis of the neuronal network underlying locomotion. The focus is on cellular mechanisms that are important for the pattern generation, as well as different types of pre- and postsynaptic modulation. This experimental model is bridging the gap between the molecular and cellular level to the network and behavioral level. PMID- 11119699 TI - Mechanisms of motor learning in the cerebellum. AB - How the elaborate neuronal circuit in the cerebellum operates and is involved in motor learning is a question addressed in earnest in studies on the cerebellum. During the past four decades, experimental studies have revealed circuit and module structures of the cerebellum, established long-term depression (LTD) as a unique and characteristic type of synaptic plasticity in the cerebellum, and analysed signal contents of activates of cerebellar neurons related to motor learning. In the 1990s, these studies were developed to detailed analyses of the signal transduction underlying LTD, and to uncovering the involvement of the cerebellum in cognitive function. On the other hand, theoretical studies yielded epochal Marr-Albus network models of the cerebellum around 1970, and introduced control system principles explaining the essential roles of the cerebellum in motor learning as providing internal models, both forward and inverse. The author maintains the hypothesis that reorganisation of the neuronal circuit by error driven induction of LTD constitutes the major memory and learning mechanisms of the cerebellum. In this article, we examine the validity of the hypothesis in light of currently available data in recent studies of the cerebellum. PMID- 11119700 TI - Original involvement of antimicrobial peptides in mussel innate immunity. AB - Recently, the existence and extended diversity of antimicrobial peptides has been revealed in two mussel species. These molecules are classified into four groups according to common features of their primary structure: defensins, mytilins, myticins and mytimycin. In Mytilus galloprovincialis, gene structure reveals synthesis as precursors in circulating hemocytes. Synthesised even in absence of challenge, the precursors mature and the peptides are stored in granules as active forms. The different peptides are engaged in the destruction of bacteria inside phagocytes, before being released into hemolymph to participate in systemic responses. Such involvement in anti-infectious responses is unique, and apparently more related to those of mammalian phagocytes than to those of insects. PMID- 11119701 TI - Inhibition of electron transport at the cytochrome b(6)f complex protects photosystem II from photoinhibition. AB - Photoinhibition of photosystem II (PS II) activity was studied in thylakoid membranes illuminated in the presence of the inhibitor of the cytochrome b(6)f complex 2'iodo-6-isopropyl-3-methyl-2',4, 4'-trinitrodiphenylether (DNP-INT). DNP INT was found to decrease photoinhibition. In the absence of DNP-INT, anaerobosis, superoxide dismutase and catalase protected against photoinhibition. No effect of these treatments was observed in the presence of DNP-INT. These data demonstrate that photoinhibition under these conditions is caused by reactive oxygen species which are formed most probably by the reduction of oxygen at photosystem I. The results are discussed in terms of the importance of photosynthetic control in protection against photoinhibition in vivo. PMID- 11119702 TI - 5S rRNA binding proteins from the hyperthermophilic archaeon, Pyrococcus furiosus. AB - A determination was made of the nucleotide sequence of the 2719 bp region of a ribosomal protein gene cluster (PfeL32-PfeL19-PfL18-PfS5-PfL30) containing a 5S rRNA binding protein L18 homolog of hyperthermophilic archaea Pyrococcus furiosus. The organization of the archaeal ribosomal protein gene cluster is similar to that in the spc-operon of Escherichia coli (L6-L18-S5-L30-L15) but has two additional genes, namely those encoding PfeL32 and PfeL19, which were identified as extra proteins that are apparently not present in bacterial E. coli. Using an inducible expression system, P. furiosus mature PfL18 protein and a mutant PfL18 with the basic N-terminal amino acid region deleted were produced in large amounts in E. coli and Northwestern analysis showed the N-terminal region of PfL18, including the conserved arginine-rich region, to have a significant role in 5S rRNA-PfL18 interaction. PMID- 11119703 TI - Epigenes: design and construction of new hereditary units. AB - A plasmid digene construction designed before [Tchuraev, R.N. (1982) J. Gen. Biol. 43, 79-87] has been realised, including feedback by repressing proteins with given trigger regime of gene functioning. Experimental tests of the expected epigene properties of the obtained pECPI recombinant plasmid involving lacI and cI(857) regulatory genes have shown a phenomenon of steady inheritance of two alternative epigenotypes lacI(1)cI(0) and lacI(0)cI(1), as well as an external toggle switch through metabolitic and temperature signals from one inherited functional state of the cyclic digene system into another. Thus, we have constructed a hereditary unit of a specific kind, namely, a two-component stationary epigene with preset properties. PMID- 11119704 TI - Immunocontent and secretion of S100B in astrocyte cultures from different brain regions in relation to morphology. AB - Primary astrocyte cultures prepared from neonatal hippocampus, cerebral cortex and cerebellum were morphologically distinct. Cells from hippocampus and cortex were almost entirely protoplasmic, whereas cerebellar astrocytes had many processes; in the absence of serum these differences were accentuated. We compared the immunocontent and secretion of the mitogenic protein S100B in these cultures. Immunocontent was 2.5 times higher in cerebellar astrocytes than in hippocampal or cortical astrocytes. Cells from all three regions secreted S100B under basal conditions, but the secretion rate was higher in cerebellar astrocytes. Secretion depended on protein synthesis and was increased by incubation with forskolin or lysophosphatidic acid in mechanisms which were additive. The stellate morphology induced by forskolin was reversed by lysophosphatidic acid in hippocampal but not in cerebellar cultures, suggesting that S100B secretion was not associated with a process-bearing phenotype of astrocytes. PMID- 11119705 TI - Expression of beta-arrestins in toxic and cold thyroid nodules. AB - beta-Arrestins mediate agonist dependent desensitization of G protein-coupled receptors. Somatic TSH receptor mutations were identified in the majority of hot thyroid nodules. When transiently overexpressed in COS 7 cells these mutations resulted in constitutive activation of the cAMP pathway. However, the in vivo mechanisms and the in vivo desensitization of these TSH receptor mutations are unknown. Moreover, constitutively activated beta-adrenergic receptors are known to be constitutively desensitized. Therefore, we investigated the expression of beta-arrestins in toxic thyroid nodules (TTNs) with and without somatic TSH receptor mutation and in cold thyroid nodules (CTNs) by Western blotting and ELISA. Expression of beta-arrestin 2 was increased in all TTNs while beta arrestin 2 expression was decreased in CTNs compared to their corresponding surrounding tissue. The mean beta-arrestin 1 expression was unchanged in the cytosol of TTNs, in membranes and cytosol of CTNs and decreased in the membranes of TTNs compared to their surrounding tissue. Transient coexpression of beta arrestins 1 or 2 with the TSH receptor in HEK 293 cells and subsequent determination of cAMP showed that in vitro both beta-arrestins interact with the TSH receptor and are able to desensitize the receptor. The increased beta arrestin 2 expression in TTNs and the desensitization of the TSH receptor by beta arrestin 2 in vitro suggest that the beta-arrestin 2 expression is cAMP dependent and that beta-arrestin 2 very likely desensitizes the constitutively activated TSH receptor in toxic thyroid nodules. PMID- 11119706 TI - Kidney produces a novel acylated peptide, ghrelin. AB - Ghrelin is a novel growth hormone-releasing peptide with a unique acylated structure. Here we reveal that prepro-ghrelin gene is expressed in the mouse kidney and glomerulus. We also show by reverse-phase high performance liquid chromatography coupled with radioimmunoassay that the mouse kidney does produce ghrelin. The ghrelin immunoreactivity in the mouse kidney is 6.79+/-0.48 fmol/mg (n=5), which is much more abundant than that in the mouse plasma of 0.339+/-0.029 fmol/microl (n=6). Furthermore, prepro-ghrelin gene is expressed in cultured rat mesangial cells, fibroblast-like NRK-49F cells and mouse podocytes, but not in rat epithelial cell-like NRK-52E cells. Ghrelin receptor gene is also expressed in the rat kidney. These findings demonstrate that the kidney, glomerulus and renal cells express prepro-ghrelin gene and ghrelin is produced locally in the kidney, and suggest the endocrine and/or paracrine roles of ghrelin in the kidney. PMID- 11119707 TI - P2 purinergic receptors of human eosinophils: characterization and coupling to oxygen radical production. AB - Extracellular nucleotides elicit multiple responses in eosinophils but no information on expression of purinergic receptors in these cells is available so far. In the present study we show that human eosinophils express the following P2Y and P2X subtypes: P2Y(1), P2Y(2), P2Y(4), P2Y(6), P2Y(11), and P2X(1), P2X(4), P2X(7), whose stimulation results in intracellular Ca(2+) increase and production of large amounts of reactive oxygen intermediates. These events are stimulated or inhibited, respectively, by P2 receptor agonists or antagonists. PMID- 11119708 TI - The biochemical profile of rat testicular tissue as measured by magic angle spinning 1H NMR spectroscopy. AB - The testis is the principal organ of male fertility, responsible for the production of spermatozoa and their maturation into sperm. However, the underlying biochemistry of the testis is relatively understudied. The fluidic and homogeneous nature of the testis makes it an ideal organ for high resolution magic angle spinning (MAS) 1H NMR spectroscopy. In this study we have catalogued the low molecular weight metabolites. The testis contains large amounts of creatine, of which a substantial proportion was shown to be extracellular using bipolar gradients to measure apparent diffusion coefficients. The tissue also contained relatively high amounts of uridine. PMID- 11119710 TI - Enhanced release of soluble urokinase receptor by endothelial cells in contact with peripheral blood cells. AB - The urokinase receptor (uPAR) on the cell surface plays an important role in extracellular proteolysis, cell migration and adhesion. Soluble uPAR (suPAR) has been recently discovered in plasma, but its origin is unclear. Our results now demonstrate that both unstimulated blood mononuclear and endothelial cells can release suPAR and that the release is enhanced when either mononuclear cells or thrombocytes are cultured together with endothelial cells. Co-culture without cell-cell contacts fails to enhance suPAR release. We also found suPAR fragments, known to be potent inducers of chemotaxis, in co-culture growth medium samples. Taken together, our results suggest that normal plasma suPAR can be produced by endothelial and mononuclear cells and that suPAR release in cell-cell contacts may have a regulatory role in cell adhesion. PMID- 11119709 TI - Insulin selectively stimulates nuclear phosphoinositide-specific phospholipase C (PI-PLC) beta1 activity through a mitogen-activated protein (MAP) kinase dependent serine phosphorylation. AB - Using NIH 3T3 cells, we have investigated nuclear phosphoinositide metabolism in response to insulin, a molecule which acts as a proliferating factor for this cell line and which is known as a powerful activator of the mitogen-activated protein (MAP) kinase pathway. Insulin stimulated inositol lipid metabolism in the nucleus, as demonstrated by measurement of the diacylglycerol mass produced in vivo and by in vitro nuclear phosphoinositide-specific phospholipase C (PI-PLC) activity assay. Despite the fact that nuclei of NIH 3T3 cells contained all of the four isozymes of the beta family of PI-PLC (i.e. beta1, beta2, beta3, and beta4), insulin only activated the beta1 isoform. Insulin also induced nuclear translocation of MAP kinase, as demonstrated by Western blotting analysis, enzyme activity assays, and immunofluorescence staining, and this translocation was blocked by the specific MAP kinase kinase inhibitor PD98059. By means of both a monoclonal antibody recognizing phosphoserine and in vivo labeling with [(32)P]orthophosphate, we ascertained that nuclear PI-PLC-beta1 (and in particular the b subtype) was phosphorylated on serine residues in response to insulin. Both phosphorylation and activation of nuclear PI-PLC-beta1 were substantially reduced by PD98059. Our results conclusively demonstrate that activation of nuclear PI-PLC-beta1 strictly depends on its phosphorylation which is mediated through the MAP kinase pathway. PMID- 11119711 TI - The formation of chlorophyll from chlorophyllide in leaves containing proplastids is a four-step process. AB - The time course of the different esters of chlorophyllide (Chlide) during the formation of chlorophyll a (Chl) in embryonic bean leaves containing proplastids was investigated by HPLC. After the reduction of photoactive Pchlide (Pchlide) to Chlide, three intermediates, i.e. Chlide geranylgeraniol, Chlide dihydrogeranylgeraniol and Chlide tetrahydrogeranylgeraniol were detected before the formation of Chlide phytol, i.e. authentic Chl. The transformation of Chlide to Chl was found to be much faster in leaves containing proplastids than in etiolated leaves with etioplasts. PMID- 11119712 TI - Generation and degradation of human endostatin proteins by various proteinases. AB - The angiogenesis inhibitor endostatin is a fragment of the NC1 domain of collagen XVIII. The generation of endostatin has been investigated only in murine hemangioendothelioma cell cultures and was ascribed to cathepsin L. Distinct endostatin-like fragments were detected in human tissues and serum. To identify proteinases able to generate such fragments, we incubated human NC1 with proteinases of all classes, including cathepsin L. Eleven out of 12 generate fragments with an N-terminus within the same 15 residue stretch as those occurring physiologically, indicating that this region is sensitive to many proteinases. None correspond to mouse endostatin. However, the efficiencies of these proteinases differed markedly. Some proteinases also proved to degrade endostatin, pointing to another regulatory loop of angiogenesis. PMID- 11119713 TI - NADPH oxidase activity is required for endothelial cell proliferation and migration. AB - NADPH oxidase has been shown to play an important role in cardiovascular biology. The goal of the present study was to determine whether NADPH oxidase activity is important for endothelial cell growth and migration. In proliferation assays, growth factor- or serum-induced DNA synthesis in three different types of human endothelial cells was abrogated by inhibitors of NADPH oxidase, but not by inhibitors of xanthine oxidase or nitric oxide synthase. Moreover, vascular endothelial growth factor-induced migration of human endothelial cells was suppressed in the presence of NADPH oxidase inhibitors. These results support a potential role for NADPH oxidase in mediating angiogenesis. PMID- 11119714 TI - Use of ruthenium red as an inhibitor of mitochondrial Ca(2+) uptake in single rat cardiomyocytes. AB - With the current resurgence of interest in the role of mitochondrial [Ca(2+)] in energy production and cellular Ca(2+) signalling, ruthenium red (RR) is being increasingly used as an inhibitor of mitochondrial Ca(2+) uptake. In the present study, the effects of RR on cell and mitochondrial [Ca(2+)], and on cell contractility were determined in isolated rat ventricular myocytes subjected to adrenergic and electrical stimulation. At low concentrations, 0-1 microM, RR inhibited mitochondrial Ca(2+) uptake but this was a secondary effect due to a reduced total intracellular [Ca(2+)], a conclusion supported by the ability of RR to inhibit cell shortening. 5 microM RR completely inhibited cell contraction, whereas higher concentrations, 10-25 microM, induced spontaneous Ca(2+) oscillations and contractile waves. These results indicate that great care must be taken when using RR in intact cells, and in interpreting any effects as resulting from a primary inhibition of mitochondrial Ca(2+) uptake. PMID- 11119715 TI - Peroxisome proliferator-activated receptor gamma activators inhibit interleukin 12 production in murine dendritic cells. AB - Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor superfamily. They are divided into three subtypes (alpha, beta or delta, and gamma) and are involved in lipid and glucose homeostasis and in the control of inflammation. In this study, we analyzed the expression of PPARs in murine dendritic cells (DCs), the most potent antigen presenting cells. We find that immature as well as mature spleen-derived DCs express PPARgamma, but not PPARalpha, mRNA and protein. We also show that the PPARgamma activator rosiglitazone does not interfere with the maturation of DCs in vitro nor modifies their ability to activate naive T lymphocytes in vivo. Finally, we present evidence that PPARgamma activators down-modulate the CD40-induced secretion of interleukin-12, a potent Th1-driving factor. These data suggest a possible role for PPARgamma in the regulation of immune responses. PMID- 11119716 TI - Consensus predictions of membrane protein topology. AB - We have explored the possibility that consensus predictions of membrane protein topology might provide a means to estimate the reliability of a predicted topology. Using five current topology prediction methods and a test set of 60 Escherichia coli inner membrane proteins with experimentally determined topologies, we find that prediction performance varies strongly with the number of methods that agree, and that the topology of nearly half of all E. coli inner membrane proteins can be predicted with high reliability (>90% correct predictions) by a simple majority-vote approach. PMID- 11119717 TI - Phosphorylated, but not native, tau protein assembles following reaction with the lipid peroxidation product, 4-hydroxy-2-nonenal. AB - A correlation between hyperphosphorylation of tau protein and its aberrant assembly into paired helical filaments has lead to suggestions that phosphorylation controls assembly, but lacked a mechanistic basic. In this work, we have found that phosphorylated, but not native, tau protein is able to form polymers after the reaction with 4-hydroxy-2-nonenal, a highly toxic product of lipid peroxidation. Phosphorylation of tau by both proline or non-proline directed kinases, was able to assemble it into polymers. PMID- 11119718 TI - Analysis of FAK-associated signaling pathways in the regulation of cell cycle progression. AB - Focal adhesion kinase (FAK) is an important mediator of signal transduction pathways initiated by integrins in cell migration, survival and cell cycle regulation. The ability of FAK to mediate integrin signaling in the regulation of cell cycle progression depends on the phosphorylation of Tyr397, which implies a functional significance for the formation of FAK signaling complexes with Src, phosphatidylinositol-3-kinase (PI3K) and Grb7. We have previously described a FAK mutant, D395A, that selectively disrupts FAK binding to PI3K, but allows FAK association with Src. Using this mutation in a mislocalized FAK mutant background, we show here that formation of a FAK/PI3K complex is not sufficient for cell cycle progression but the formation of a FAK/Src complex plays an essential role. We also show that mutation of D395 to A disrupted FAK association with Grb7. This suggests that a FAK/Grb7 complex is not involved in the cell cycle regulation either, which is supported by direct analysis of cells expressing a dominant negative Grb7 construct. Finally, we provide evidence that the Src-dependent association of FAK with Grb2 and p130(Cas) are both required for the regulation of cell cycle progression by FAK. Together, these studies identify important FAK downstream signaling pathways in cell cycle regulation. PMID- 11119719 TI - CREB is cleaved by caspases during neural cell apoptosis. AB - Programmed cell death, or apoptosis, is a tightly regulated process mediated by selective cleavage of proteins by caspases, resulting in ordered destruction of the cell. In addition to structural proteins, proteins that mediate anti apoptotic signal transduction are also substrates; their destruction eliminates potential futile attempts to escape execution. We asked whether cAMP response element binding protein (CREB), a transcription factor that mediates nerve growth factor (NGF) survival signals, is a target for caspases during apoptosis. CREB was specifically cleaved by caspases in neuroblastoma extracts, and in cells induced to undergo apoptosis by staurosporine. The destruction of CREB eliminates a key factor that could reverse apoptosis. PMID- 11119720 TI - Kinesin subfamily UNC104 contains a FHA domain: boundaries and physicochemical characterization. AB - By sequence analysis we show that the U104 domain found in the UNC104 subfamily of kinesins is a forkhead homology-associated domain (FHA). A combination of limited proteolysis, mass spectroscopy, and physicochemical analysis define this domain as a genuine autonomously folding domain. Our data show that the FHA domain is shorter than previously reported since the C-terminal alpha-helix is not part of its minimum core. Key amino acids postulated to recognize phosphorylated residues are conserved. These data suggest that the kinesin FHA domains are functional domains involved in protein-protein interactions regulated by phosphorylation. PMID- 11119721 TI - Ganoderma extract activates MAP kinases and induces the neuronal differentiation of rat pheochromocytoma PC12 cells. AB - The pharmacology and clinical application of traditional Chinese medicine has been extensively documented. We have used an in vitro model system, PC12 cells, to demonstrate the presence of neuroactive compounds in Ganoderma lucidum (lingzhi). Ganoderma extract induced the neuronal differentiation of PC12 cells and prevented nerve growth factor-dependent PC12 neurons from apoptosis. Moreover, these effects of ganoderma might be mediated via the ras/extracellular signal-regulated kinase (Erk) and cAMP-response element binding protein (CREB) signaling pathways, as demonstrated by the phosphorylation of Erk1, Erk2 and CREB. Thus, our data not only present the first evidence of the presence of neuroactive compounds that mediate the neuronal differentiation and neuroprotection of the PC12 cells, but also reveal the potential signaling molecules involved in its action. PMID- 11119722 TI - The T9176G mutation of human mtDNA gives a fully assembled but inactive ATP synthase when modeled in Escherichia coli. AB - A new mutation in human F(1)F(0) ATPase6, T9176G, which changes Leu 217 to an Arg, has been described in two siblings with Leigh syndrome [Carrozzo et al. (2000) Neurology, in press]. This mutation was modeled in Escherichia coli by changing Leu 259 (the equivalent residue) to Arg and the properties of the altered ECF(1)F(0) were compared to those of previously characterized ATPase6 mutants also modeled in the E. coli enzyme. The L259R change produced a fully assembled ECF(1)F(0) which had no significant ATP hydrolysis, ATP synthesis or proton pumping functions. This is very different from previously described human ATPase6 mutations. The presence of Arg at position 259 in subunit a did not make membranes permeable to protons. We conclude that the mutation inhibits functioning by blocking the rotary motor action of the enzyme. PMID- 11119723 TI - Inositol polyphosphate kinase activity of Arg82/ArgRIII is not required for the regulation of the arginine metabolism in yeast. AB - Arg82, a nuclear regulator of diverse cellular processes in yeast, is an inositol polyphosphate kinase. Some defects such as the regulation of arginine metabolism observed in an arg82Delta, result from a lack of Mcm1 and Arg80 stability. We show here that neither the kinase activity of Arg82 nor inositol phosphates are required for the control of arginine metabolism. Arg82 mutations keeping kinase active affect the expression of arginine genes, whereas mutations in the kinase domain do not impair this metabolic control. PMID- 11119724 TI - Regulation of Wee1 kinase in response to protein synthesis inhibition. AB - To investigate the mechanism coupling growth (protein synthesis) with cell division, we examined the relationship between the tyrosine kinase Wee1 that inhibits Cdc2-Cdc13 mitosis-inducing kinase by phosphorylating it, and protein synthesis inhibition in fission yeast. The wee1-50 mutant showed supersensitivity to protein synthesis inhibitor, cycloheximide. Wee1 was essential for the G(2) delay upon a partial inhibition of protein synthesis. Indeed, the protein synthesis inhibition caused an increase in the Wee1 protein by the Sty1/Spc1 MAPK dependent transcriptional and the Sty1/Spc1 MAPK-independent post-transcriptional regulations. Further, the results indicated that the post-transcriptional regulation is important for the G(2) delay. PMID- 11119725 TI - beta-agonists regulate Na,K-ATPase via novel MAPK/ERK and rapamycin-sensitive pathways. AB - We studied whether the beta-adrenergic agonist, isoproterenol (ISO), regulates Na,K-ATPase in alveolar epithelial cells (AEC) via a mitogen-activated protein kinase (MAPK)/extracellular signaling related kinase (ERK) dependent pathway. ISO increased ERK activity in AEC by 10 min via a beta-adrenergic receptor, protein kinase A (PKA)-dependent mechanism. Activation of the MAPK pathway by ISO, resulted in increased Na,K-ATPase beta1 and alpha1 subunit protein abundance in whole cell lysates, which resulted in functional Na, K-ATPases at the basolateral membranes. ISO did not change the alpha1 or beta1 mRNA steady state levels, but rapamycin, the inhibitor of the mammalian target of rapamycin, also blocked the ISO-mediated increase in Na,K-ATPase total protein abundance, suggesting a posttranscriptional regulation. We conclude that ISO, regulates the Na,K-ATPase in AEC via PKA, ERK and rapamycin-sensitive mechanisms. PMID- 11119726 TI - The human PER1 gene is transcriptionally regulated by multiple signaling pathways. AB - The mammalian period (Per) genes are components of the circadian clock and appear to be regulated via an autoregulatory feedback loop. Here we show that the human PER1 (hPER1) gene is synergistically activated by protein kinases A and C (PKA, PKC) and cAMP responsive element binding protein. Activators and inhibitors of PKA as well as PKC modulate endogenous hPER1 expression and hPER1 promoter-driven reporter gene activity in a dose-dependent manner. Our results suggest that the hPER1 promoter acts as a sensor for multiple signaling molecules thereby integrating different physiological parameters. This regulation of hPER1 appears to be significant for rapid adaptation to changing environmental conditions. PMID- 11119727 TI - Glycosylation of the calcitonin receptor-like receptor at Asn(60) or Asn(112) is important for cell surface expression. AB - The human calcitonin (CT) receptor-like receptor (hCRLR) of the B family of G protein-coupled receptors is N-glycosylated and associates with receptor-activity modifying proteins for functional interaction with CT gene-related peptide (CGRP) or adrenomedullin (ADM), respectively. Three putative N-glycosylation sites Asn(60), Asn(112) and Asn(117) are present in the amino-terminal extracellular domain of the hCRLR. Tunicamycin dose-dependently inhibited the glycosylation of a myc-tagged hCRLR and in parallel specific [(125)I]CGRP and -ADM binding. Similarly, the double mutant myc-hCRLR(N60,112T) exhibited minimal N-glycosidase F sensitive glycosylation, presumably at the third Asn(117), and the cell surface expression and specific radioligand binding were impaired. Substitution of the Asn(117) by Thr abolished CGRP and ADM binding in the face of intact N glycosylation and cell surface expression. PMID- 11119728 TI - Thyroid disease and female reproduction. AB - OBJECTIVE: To review the menstrual function and fertility in thyroid disease, mainly in hyperthyroidism and hypothyroidism. Also to register the consequences of (131)I therapy, which is used widely in the treatment of Graves' disease and thyroid cancer, on subsequent pregnancies and on fertility in these patients. DESIGN: A MEDLINE computer search was used to identify relevant studies. The type of menstrual disturbances and the status of fertility were recorded from all the studies found. Also, the fertility and genetic hazard of female patients with Graves' disease and thyroid cancer who were treated with (131)I were registered. RESULT(S): Both hyperthyroidism and hypothyroidism may result in menstrual disturbances. Menstrual abnormalities are less common now than in previous series. In a recent study, we found that only 21.5% of 214 thyrotoxic patients had some type of menstrual disturbance, compared to 50 to 60% in some older series. The most common manifestations are hypomenorrhea and oligomenorrhea. According to the results of endometrial biopsies, most thyrotoxic women remain ovulatory. Moreover, the genetic hazard incident to radioiodine therapy in Graves' disease and thyroid carcinoma is very small; exposure to (131)I does not cause reduced fecundity, and the risk of loss of fertility is not a contraindication for its use in these patients. mIn hypothyroidism, the frequency of menstrual irregularities has very recently been reported to be 23.4% among 171 hypothyroid patients studied. This is much less than that reported in previous studies, which showed that 50 to 70% of hypothyroid female patients had menstrual abnormalities. The most common manifestation is oligomenorrhea. Severe hypothyroidism is commonly associated with failure of ovulation. Ovulation and conception can occur in mild hypothyroidism. These pregnancies are, however, often associated with abortions, stillbirths, or prematurity. The latter may be of greater clinical importance in infertile women with unexplained infertility. CONCLUSION(S): These new data, mainly concerning menstrual abnormalities in hyperthyroidism and hypothyroidism, are inconsistent with what is generally believed and written in the classic thyroid textbooks and indicate that such opinions should be revised. PMID- 11119729 TI - The absolute truth. PMID- 11119730 TI - Laryngeal cytological aspects in women with surgically induced menopause who were treated with transdermal estrogen replacement therapy. AB - OBJECTIVE: To investigate the effects of estrogen replacement therapy (ERT) on laryngeal cytology in postmenopausal women. DESIGN: Prospective open clinical trial. SETTING: Outpatient menopausal clinic in the Department of Gynecology, University of Catania, Catania, Italy. PATIENT(S): Eighty-four healthy, surgically postmenopausal women, of whom 48 were treated with ERT and 36 were considered as a control group. INTERVENTION(S): Transdermal E(2) treatment by patches or gel, evaluation of laryngeal cytology with cytobrush by indirect laryngoscopy, and questionnaire for the voice history. MAIN OUTCOME MEASURE(S): Changes in cytologic aspects of laryngeal cells with respect to vaginal cytology by hematoxylin and eosin staining; subjective voice changes. RESULT(S): Sixty seven women completed the study. Ten women from the ERT group and five from the control group dropped out because of the invasive laryngoscope method; two subjects in the control group were excluded because of pathologies of the vocal cord. Hematoxylin and eosin staining confirmed similar superficial-intermediate aspects of the cells between the laryngeal and the vaginal smears in ERT-treated women. In the control group, both smears showed aspects of atrophy-dystrophy. The ERT group had a subjectively better quality of voice than the control group. CONCLUSION(S): Our study confirms that the larynx is an estrogen target, as are vaginal cells. ERT may provide prevention and treatment of dystrophic pathologies of the vocal cords in postmenopausal women. PMID- 11119731 TI - Failure of the combination of sequential oral and transdermal estradiol plus norethisterone acetate to affect plasma homocysteine levels. AB - OBJECTIVE: A high level of plasma homocysteine may be deleterious to vascular health. We therefore compared the effect of combinations of sequential oral and transdermal estradiol plus norethisterone acetate on plasma homocysteine. DESIGN: Prospective, randomized study. SETTING: Outpatient department of obstetrics and gynecology in a university hospital. PATIENT(S): Forty-two healthy, nonsmoking postmenopausal women starting hormone replacement therapy (HRT) to control climacteric symptoms. INTERVENTION(S): In a randomized order, the women started using either oral HRT (2 mg of estradiol on days 1-12, 2 mg of estradiol plus 1 mg of norethisterone acetate (NETA) on days 13-22, and 1mg of estradiol on days 23-28; n = 21) or transdermal HRT (50 microg/d of estradiol on days 1-28 and 250 microg/d of norethisterone acetate on days 15-28, n = 21) for 1 year. MAIN OUTCOME MEASURE(S): Fasting plasma levels of homocysteine were measured before the treatment and during the combined estradiol-plus-NETA phases of the sixth and 12th treatment cycles. RESULT(S): Basal homocysteine levels in the oral group (8.2 +/- 3.1 micromol/L, mean plusmn;SD) and transdermal group (8.7 plusmn; 1.8 micromol/L, mean plusmn;SD) were not affected by the estradiol-plus-NETA combination. CONCLUSION(S): Neither an oral nor a transdermal combination of sequential estradiol and NETA causes significant changes in plasma homocysteine in Finnish postmenopausal women with normal baseline homocysteine levels. PMID- 11119732 TI - Long-term follow-up after quinacrine sterilization in Vietnam. Part I: interim efficacy analysis. AB - OBJECTIVE: To determine the long-term efficacy of nonsurgical sterilization with quinacrine. DESIGN: Observational cohort study. SETTING: Rural provinces in northern Vietnam. PATIENT(S): Two thousand seven hundred and nine women who had quinacrine insertions between 1989 and 1993. INTERVENTION(S): Interviews in 1994, 1995, and 1996 and review of available medical records. Pregnancy rates were corrected for problems in detecting and confirming pregnancies. MAIN OUTCOME MEASURE(S): Pregnancy rates. RESULT(S): Over 90% of women were interviewed at least once. Uncorrected cumulative pregnancy rates were 12.9% at 5 y after two insertions and 27.3% after one insertion. Effectiveness varied by age group: the partially corrected pregnancy rates after two insertions were 6.8% in women 35 or older at the time of insertion and 13.0% in women under 35. A subgroup of women who received oral papaverine at the time of quinacrine insertion had lower pregnancy rates, with a cumulative uncorrected rate of 5.3% at 4 years among women of all ages. CONCLUSION(S): Efficacy of quinacrine appears reasonable for two insertions of quinacrine in women 35 and older. It may be possible to improve efficacy by the use of papaverine or the Hieu insertion technique. PMID- 11119733 TI - Long-term follow-up after quinacrine sterilization in Vietnam. Part II: interim safety analysis. AB - OBJECTIVE: To determine the long-term safety of nonsurgical sterilization with quinacrine. DESIGN: Observational cohort study. SETTING: Rural provinces in northern Vietnam. PATIENT(S): Two thousand eight hundred forty women who had had quinacrine insertions and an age-matched comparison group of 1,658 women who had an intrauterine device (IUD) insertion between 1989 and 1993. METHOD(S): Interviews in 1994, 1995, and 1996 and review of available medical records. This is a planned interim analysis. MAIN OUTCOME MEASURE(S): Ectopic pregnancies and the occurrence of other adverse health events. RESULT(S): Over 90% of women were interviewed at least once. Despite matching on age, the groups differed on baseline parity. The ectopic pregnancy rates were similar after either one or two insertions and were similar to the rate of ectopic pregnancies after surgical sterilization in the United States. The quinacrine group reported more gynecologic health problems than the IUD group. However, after correcting for information bias, there was no dose-response effect between the one- and two insertion quinacrine groups, suggesting the possibility of recall bias or differing baseline health status. CONCLUSION(S): Ectopic pregnancies do not appear to be increased compared with U.S. surgical sterilization rates. The data on other adverse events are more difficult to interpret. PMID- 11119734 TI - Expression of alpha4beta1 and alphavbeta3 integrins in the endometrium of women using the T200 copper intrauterine device. AB - OBJECTIVE: To investigate the expression of alpha4 and beta3 integrin subunit levels in the endometrium of healthy women and copper intrauterine device (IUD) T200 users. DESIGN: Case control study. SETTING: An academic teaching hospital and a primary care clinic. PATIENT(S): Thirteen copper IUD users and 13 normal fertile women. INTERVENTION(S): Timed endometrial biopsies during the mid secretory phase (days 20 to 24). MAIN OUTCOME MEASURE(S): Histologic dating of endometrium and immunohistochemical staining intensity of alpha4 and beta3, using the semiquantitative immunohistochemical score (HSCORE). RESULT(S): All endometrial biopsies consistent with menstrual dates were examined for integrin expression (beta3 and alpha4). No difference in alpha4 integrin expression was found between IUD users and controls in both luminal and glandular epithelium. In fertile controls, alphavbeta3 staining was present in 100% and 38.4% of glandular and luminal epithelium, respectively. In contrast, only 61.5% of the IUD users had any alphavbeta3 staining in the glandular epithelium and only 53.9% in the luminal epithelium. The intensity of immunoreactivity between the two groups (mean HSCORE) did not differ significantly. CONCLUSION(S): Proportionately, significantly fewer women using copper IUD had positive alphavbeta3 immunoreactivity in the glandular epithelium of mid-secretory endometrium. PMID- 11119735 TI - Treatment of repeated unexplained in vitro fertilization failure with intravenous immunoglobulin: a randomized, placebo-controlled Canadian trial. AB - OBJECTIVE: To evaluate the effect of intravenous immunoglobulin (IVIG) on pregnancy outcome in couples with repeated unexplained in vitro fertilization (IVF) failure. DESIGN: Prospective, randomized, double blind, placebo-controlled clinical trial. SETTING: A university-based and a free-standing IVF program. PATIENT(S): Fifty-one couples with a history of repeated unexplained IVF failure who were preparing for another fresh IVF cycle or replacement of cryopreserved embryos. INTERVENTION(S): Eligible women underwent a standard IVF stimulation using a long luteal phase GnRH analog protocol. Cryopreserved embryos were replaced after endometrial preparation with oral micronized estradiol and subsequent vaginal progesterone. The women were randomly selected to receive IVIG (500 mg/kg) or an equivalent volume of normal saline. The first infusion was given on the day of embryo transfer or during the preceding 72 hours. The second infusion was given 4 weeks later if a clinical pregnancy was confirmed by ultrasound. MAIN OUTCOME MEASURE(S): Live-birth rates. RESULT(S): Overall, the live-birth rates were 4/26 (15%) for the IVIG group and 3/25 (12%) for the placebo group (P=0. 52). There were 39 fresh IVF cycles, which yielded a clinical pregnancy rate of 28%, with live-birth rates of 4/21 (19%) for the IVIG group and 3/18 (17%) for the placebo group (P=0.59). CONCLUSION(S): In this randomized clinical trial, IVIG did not improve the live-birth rate in couples with repeated unexplained IVF failure, stringently defined by known determinants of IVF outcome. PMID- 11119736 TI - Inhibin B response to EFORT is associated with the outcome of oocyte retrieval in the subsequent in vitro fertilization cycle. AB - OBJECTIVE: To examine whether the magnitude of the rise in inhibin B levels after gonadotropin challenge is associated with subsequent response to ovarian stimulation during IVF. DESIGN: Inhibin B serum levels after EFORT (exogenous follicle-stimulating hormone ovarian reserve test). SETTING: Academic clinical practice. PATIENT(S): Serum samples from women who had undergone ovarian reserve screening with FSH in preparation for IVF. Thirteen of these women had a poor response in IVF (canceled cycle for low estradiol and/or no oocytes retrieved), and 19 had a good response (> or =10 oocytes retrieved). INTERVENTION(S): EFORT test. MAIN OUTCOME MEASURE(S): Baseline (day 3) serum E(2) (bE(2)), FSH (bFSH), and inhibin B (bInhB) levels and inhibin B and E(2) levels 24 hours after EFORT (DeltaInhB and DeltaE(2)). RESULT(S): The mean bInhB and DeltaInhB levels were significantly higher in good vs. poor responders. The odds ratio of having a good response for women with a DeltaInhB of 202 pg/mL was 51.8 times (95% CI = 6.1 1,244) the corresponding odds for women with a DeltaInhB of 49 pg/mL. As expected, DeltaE(2) was also significantly higher in good vs. poor responders; however, combination of DeltaE(2) plus DeltaInhB did not improve the odds for predicting IVF response. CONCLUSION(S): Our data suggest that DeltaInhB after EFORT may provide a method for predicting ovarian response to hyperstimulation in a subsequent IVF cycle. PMID- 11119737 TI - Bacteria in the transfer catheter tip influence the live-birth rate after in vitro fertilization. AB - OBJECTIVE: To assess the impact of individual bacteria isolated from the vagina and tip of the embryo transfer catheter on live-birth rates. DESIGN: Prospective clinical study. SETTING: Infertility outpatient clinic of a university hospital. PATIENT(S): Ninety-one women undergoing IVF-ET. INTERVENTION(S): Cultures were obtained from the vagina for aerobic and anaerobic bacteria at the time of both sonographic egg retrieval and embryo transfer and from the tip of the embryo transfer catheter. Doxycycline treatment was started after egg retrieval. MAIN OUTCOME MEASURE(S): The live birth of one or more neonates. RESULT(S): Doxycycline had no substantial impact on the recovery of individual vaginal bacteria or on bacterial vaginosis. An increase in live-birth rate was associated with the recovery of hydrogen peroxide-producing Lactobacillus from the vagina (P=0.01) and from the embryo transfer catheter (P=0.01). In contrast, a reduction in live-birth rate was associated with recovery of Streptococcus viridans (S. viridans) from the embryo transfer catheter tip (P=0.04). CONCLUSION(S): In the setting of IVF-ET, prophylactic doxycycline had little effect on vaginal bacteria. Specific bacteria recovered from the embryo transfer catheter appear associated with a detrimental or beneficial effect or with no effect on live birth rates. PMID- 11119738 TI - Effect of an in vitro fertilization program on serum CA 125, tumor-associated trypsin inhibitor, free beta-subunit of human chorionic gonadotropin, and common alpha-subunit of glycoprotein hormones. AB - OBJECTIVE: To investigate the impact of an IVF program on serum levels of tumor markers CA 125, tumor-associated trypsin inhibitor, free hCG beta-subunit, and free glycoprotein hormone alpha-subunit. DESIGN: A prospective controlled clinical study. SETTING: Outpatient university infertility clinic. PATIENT(S): Seventy-one infertile patients (with tubal occlusion, pelvic endometriosis, or unexplained infertility) undergoing IVF and nine control women with regular menstrual cycles. INTERVENTION(S): Serial blood sampling before, during, and after IVF, or during one ovulatory menstrual cycle in the controls. MAIN OUTCOME MEASURE(S): Serum levels of CA 125, tumor-associated trypsin inhibitor, hCG-beta, and glycoprotein hormone-alpha. RESULT(S): Before IVF, all tumor markers were within the normal range except for CA 125, which was elevated in patients with endometriosis. IVF led to significant increases in CA 125 and glycoprotein hormone-alpha that differed from the changes seen during normal menstrual cycles. The luteal phase increase in CA 125 correlated with levels of E(2) and P and the number of follicles. Two months after IVF, levels of CA 125 were 12% higher than levels before treatment. Tumor-associated trypsin inhibitor and hCG-beta revealed no cyclicity. CONCLUSION(S): An IVF regimen increased the release of CA 125 and glycoprotein hormone-alpha. The CA 125 elevation after IVF implies a persistent effect of ovarian hyperstimulation on CA 125 release. PMID- 11119739 TI - Basal 17beta-estradiol did not correlate with ovarian response and in vitro fertilization treatment outcome. AB - OBJECTIVE: To verify the correlation of basal 17beta-E(2) with ovarian response to stimulation and outcome of in vitro fertilization (IVF). DESIGN: Retrospective observational study. SETTING: The Assisted Conception Unit, University College London Hospitals. PATIENT(S): Three hundred five women undergoing IVF and IVF with intracytoplasmic sperm injection. INTERVENTION(S): Basal follicle stimulating hormone (FSH) and 17beta-E(2) were assessed. The cutoff level for day 2 E(2) established was 250 pmol/L. Each patient was noted for below (group A) or above (group B) the cutoff point according to her basal E(2) level. MAIN OUTCOME MEASURE(S): Basal E(2), age, duration of infertility, cycle day 2 FSH, number of ampules of gonadotropin used, number of days of stimulation, number of retrieved oocytes, fertilization rate, number of embryos transferred, number of cycles with embryo freezing, cancellation rate, clinical pregnancy rate, and implantation rate were compared between the two groups. RESULT(S): No differences were found between group A and group B in the number of oocytes retrieved (8.8 +/- 4.2 vs. 9.3 +/- 4.8), embryos transferred (2.5 +/- 0.8 vs. 2.7 +/- 0.7), cancellation (9.1% vs. 6.9%), pregnancy (24.8% vs. 30%), and implantation rate (12.3% vs. 15.6%). Correlation coefficient and coefficient of determination showed no significant correlation between basal E(2) and the number of oocytes retrieved, age, and basal FSH. CONCLUSION(S): In our study population, basal E(2) was not a sensitive predictor of ovarian response to stimulation and did not correlate with IVF outcome. PMID- 11119740 TI - Maturation and apoptosis of human oocytes in vitro are age-related. AB - OBJECTIVE: To study the morphological changes of apoptotic oocytes, and rates of in vitro maturation and apoptosis of human oocytes in relation to age. DESIGN: Prospective comparative study. SETTING: Reproductive medicine center. PATIENT(S): Women undergoing surgery for ovarian cysts. INTERVENTION(S): Oocytes were incubated in Ham's F-10 medium with 15% fetal cord serum (FCS) for 32 to 120 hours and were examined under inverted microscope every 6 to 8 hours. MAIN OUTCOME MEASURE(S): Oocyte maturation and apoptosis, Fas antigen. RESULT(S): The morphologic changes characteristic of apoptosis oocytes were shrinkage, or the occurrence of cytoplasmic condensation, membrane blebbing, fragmentation of the oocyte into "apoptotic" bodies of unequal size, or internucleosomal DNA cleavage as shown by TUNEL. The maturation rates of oocytes were highest in those from women aged 21 to 30 years, and lowest in those aged 41 to 50 years. Apoptosis occurred in 17.1% (age group 21 to 30 years), 37. 7% (31 to 40 years), and 52.3% (41 to 50 years). The rate of apoptosis of human immature oocytes cultured in vitro was significantly higher in those from older women who were 41 to 50 years old than in those women 21 to 40 years old. Fas antigen was found to be present on apoptotic oocyte membranes. CONCLUSION(S): The developmental potential of oocytes from older women decreased in vitro in a manner similar to that seen in vivo. DNA fragmentation in oocytes associated with apoptotic death might be one of the reasons for poor oocyte quality and lower fertility in older women. Fas antigen in the oocyte presumably mediates apoptosis. PMID- 11119741 TI - Effects of simulated microgravity on mammalian fertilization and preimplantation embryonic development in vitro. AB - OBJECTIVE: To study the effects of simulated microgravity on mammalian fertilization and preimplantation embryonic development in vitro with the use of a horizontal clinostat device. DESIGN: Controlled animal study. SETTING: Research laboratory at a university medical school. ANIMAL(S): B6D2F1 (C57BL/6 x DBA/2) and ICR mice between 8 and 10 weeks old. INTERVENTION(S): The first experiment was performed to investigate whether gravity is required for fertilization in vitro under three conditions: clinostat rotation, rotational control, and stationary control. In the second experiment, one-cell embryos were cultured under each condition and their morphology and viability were assessed at 96 hours. MAIN OUTCOME MEASURE(S): The fertilized numbers and embryonic numbers at the morula and blastocyst stages were recorded in each condition. RESULT(S): In the first experiment, there were no statistically significant differences in the efficiency of achieving normal fertilization in vitro among the conditions. In the second experiment, there was a statistically significant decrease in the number of embryos reaching the morula and blastocyst stages after 96 hours in culture under clinostat rotation. CONCLUSION(S): These results suggest that the process of fertilization in vitro is not sensitive to the gravitational vector. However, the possibility exists that the frequency of early embryonic lethality is increased by microgravity. PMID- 11119742 TI - Hyaluronan in follicular fluids and fertilization of oocytes. AB - OBJECTIVE: To determine the concentrations of hyaluronan, E(2), and progesterone in follicular fluids (FFs) and the incidence of apoptotic granulosa cells. Also, to examine the relationship between the concentration of hyaluronan and follicular steroids, the incidence of apoptotic cells, and the fertilizability of the oocyte in the same follicle. DESIGN: Samples of 130 follicles were retrospectively analyzed for hyaluronan and steroids and the incidence of apoptotic cells. SETTING: The reproductive center in Yamagata University Hospital. PATIENT(S): Forty women infertile because of tubal damage or unknown causes undergoing IVF treatment were selected. INTERVENTION(S): The samples were collected from follicle aspirations. MAIN OUTCOME MEASUREMENT(S): The concentrations of hyaluronan and steroids in FFs, the incidence of apoptotic granulosa cells, and oocyte fertilizability. RESULT(S): The levels of hyaluronan in FF were found to correlate positively with P (r=0.444, P<0.0001) and the incidence of apoptotic cumulus granulosa cells (r=0.387, P=0.002) and inversely with E(2) (r = -0.601, P<0.0001) and free T (r = -0.344, P=0.001). The concentration of hyaluronan in FFs containing a subsequently fertilized oocyte after insemination was significantly lower than that in FFs containing a subsequently unfertilized oocyte (P=0.0005) (fertilized, 50.0 +/- 2.6 ng/mL; triploidy, 59.1 +/- 6.8; and unfertilized, 66.9 +/- 5.9). CONCLUSION(S): The concentration of hyaluronan in FF is an indicator for estimation of oocyte viability for fertilization. PMID- 11119743 TI - In vitro maturation, fertilization, and development of human germinal vesicle oocytes collected from stimulated cycles. AB - OBJECTIVE: To evaluate whether germinal vesicle (GV) oocytes from stimulated cycles can be a source of embryos. DESIGN: In vitro model study. SETTING: Specialized laboratory of women's clinic. PATIENT(S): Women in whom oocytes were retrieved at the GV stage. INTERVENTION(S): After culture of GV oocytes in the modified TLP medium with human follicular fluid, oocytes that reached the metaphase II stage underwent ICSI. Potential of fertilization and subsequent cleavage of in vitro-matured oocytes was compared with that of an in vivo-matured control. MAIN OUTCOME MEASURE(S): Maturation rate, rate of pronuclei formation, and developmental activity. RESULT(S): The maturation rate of GV oocytes from follicles primed with gonadotropin was not affected by the presence or absence of cumulus. However, the maturation was more synchronous in oocytes with cumulus than in those without cumulus. Proportions of oocytes with two pronuclei and embryos cleaved to the 16-cell stage after ICSI were significantly lower in the oocytes matured in vitro than in the oocytes matured in vivo. CONCLUSION(S): Human GV oocytes from stimulated ovaries can be matured, fertilized, and developed in vitro. Production of embryos from GV oocytes will increase the opportunity for achieving pregnancy. PMID- 11119744 TI - Postprandial triglyceride response in patients with polycystic ovary syndrome: relationship with waist-to-hip ratio and insulin. AB - OBJECTIVE: To examine the postprandial triglyceride response to a high-fat meal in women with polycystic ovary syndrome (PCOS) compared with a matched control group. DESIGN: Controlled clinical study. SETTING: Department of Endocrinology and Pathophysiology, School of Medicine, Universidad de Los Andes. PATIENT(S): 18 Hispanic women with PCOS (nine overweight and nine nonobese) and 9 healthy control women. INTERVENTION(S): Biometric measures and blood sample collection. MAIN OUTCOME MEASURE(S): Insulin and glucose levels during a standard oral glucose tolerance test. Plasma triglyceride, cholesterol, and high-density lipoprotein cholesterol levels were measured at baseline and at 4, 5, and 6 h after a high-fat meal. RESULT(S): Both obese and nonobese PCOS women had higher waist-to-hip ratios than controls. PCOS women had higher levels of fasting and postglucose insulin and fasting triglyceride and postprandial triglyceride response and had lower levels of postprandial high-density lipoprotein cholesterol response, but no significant differences within PCOS groups were observed. CONCLUSION(S): An expanded postprandial hypertriglyceridemia in PCOS women is related to a higher waist-to-hip ratio and insulin resistance, regardless of obesity, and contributes to increase the risk for coronary artery disease. PMID- 11119745 TI - Congenital bilateral absence of the vas deferens: clinical characteristics, biological parameters, cystic fibrosis transmembrane conductance regulator gene mutations, and implications for genetic counseling. AB - OBJECTIVE: To evaluate relationships between the phenotypic and genotypic characteristics of patients with congenital bilateral absence of the vas deferens (CBAVD). DESIGN: Retrospective study. SETTING: A university hospital urology andrology department. PATIENT(S): Forty-one men with CBAVD. INTERVENTION(S): CBAVD was diagnosed during surgical and/or ultrasound exploration of the vasa deferentia (VD) (n = 39), or on the basis of impalpable scrotal VD (n = 2). MAIN OUTCOME MEASURE(S): History, clinical and seminal characteristics, and cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations including IVS 8 polyT analysis. RESULT(S): A palpable scrotal vas deferens was present as a fibrous cord or nonpermeable duct in 13% of patients undergoing surgical exploration. Seminal vesicles were bilaterally absent in 28% of patients. No CFTR gene mutation or 5T allele was detected in 24.5% of the patients. Two CBAVD patients with renal agenesis carried a CFTR gene mutation (DeltaF508/5T-9T and R117G/7T-9T). CBAVD patients who have both a semen volume of < or =1.0 mL and a semen pH of < 7.0 have a significantly higher risk of severe CFTR gene mutation (OR = 9.12 [95% CI = 1.81-49.50]). CONCLUSION(S): A palpable scrotal vas deferens was found in 13% of CBAVD patients. Semen volume of < or =1.0 mL and semen pH of < 7.0 in CBAVD patients were associated with a higher risk of severe CFTR gene mutations. Patients with CBAVD and renal agenesis should be screened for CFTR gene mutations before assisted reproductive techniques are used. PMID- 11119746 TI - Bladder endometriosis must be considered as bladder adenomyosis. AB - OBJECTIVE: To present data from a series of 17 cases of bladder endometriosis. DESIGN: Clinical study. SETTING: A university hospital department of gynecology. PATIENT(S): Seventeen patients complaining of menstrual urinary symptoms and/or pelvic pain. INTERVENTION(S): Diagnosis and resection of a bladder adenomyotic nodule. MAIN OUTCOME MEASURE(S): Histologic analysis and postsurgical outcome. RESULT(S): Seventy-six percent of the patients reported menstrual mictalgia and pollakiuria, and 88% reported dysmenorrhea and dyspareunia. Cystoscopy, intravenous pyelography, and magnetic resonance imaging revealed a nodular mass in the anterior fornix adjacent to the uterine wall, developed in the vesical muscularis and involving the vesical mucosa in all cases but one. The bladder nodule was associated with a rectovaginal nodule in six cases (35%). Because recurrence was noted soon after cessation of medical therapy, surgical excision was proposed. The vesical mucosa was found to be intact in almost all cases, so extramucosal laparoscopic excision was the method of choice. Histologic examination proved that 90% of the nodule consisted of smooth muscle hyperplasia. CONCLUSION(S): So-called bladder endometriosis is actually an adenomyotic nodule of the bladder which, from a histologic point of view, is similar to a rectovaginal adenomyotic nodule and frequently (35%) associated with it. PMID- 11119747 TI - Peritoneal fluid concentrations of interleukin-11 in women with endometriosis. AB - OBJECTIVE: To determine the peritoneal fluid concentrations of interleukin-11 (IL 11) in women with endometriosis as compared with the control group. DESIGN: A prospective, controlled study. SETTING: The obstetrics and gynecology department of a teaching hospital and a university immunology department. PATIENT(S): Sixty consecutive women undergoing laparoscopic surgery for benign gynecological indications. INTERVENTION(S): Peritoneal fluid was obtained during laparoscopy, and the concentration of IL-11 was measured. MAIN OUTCOME MEASURE(S): Concentration of IL-11 in correlation with the presence of endometriosis, its stage, and the phase of the menstrual cycle. RESULT(S): IL-11 was detectable in the peritoneal fluid of 64% of women tested. Concentrations of IL-11 showed no correlation with the presence of endometriosis, the American Fertility Society stage of the disease, or the phase of the menstrual cycle. CONCLUSION(S): We found no evidence to suggest that IL-11 is involved in the pathogenesis of pelvic endometriosis. PMID- 11119748 TI - Increase in the expression of killer cell inhibitory receptors on peritoneal natural killer cells in women with endometriosis. AB - OBJECTIVE: Malfunction of peritoneal natural killer cells (NK) may result in endometriosis. The present study was designed to determine whether the decrease in NK cytotoxicity occurs at early and advanced stages of endometriosis and is due to the increase in the NK inhibition receptors. DESIGN: A case control study. SETTING: A tertiary-care infertility center . PATIENT(S): A total of 44 women (controls, n = 11; women with early-stage endometriosis, n = 11; and women with advanced-stage endometriosis, n = 22) were included in this study. INTERVENTION(S): Laparoscopic examination. MAIN OUTCOME MEASURE(S): NK cytotoxicity was determined by assay of (51)Cr release against K562 cells, and the expression of killer cell inhibitory receptors (KIR, including NKB1, GL183, and EB6) in NK cells was examined by flow cytometry. RESULT(S): Women with endometriosis showed a decrease in peritoneal NK cytotoxicities against K562 at early and advanced stages of endometriosis. The expression of KIR (NKB1 and EB6) was significantly elevated in the peritoneal NK cells of women with advanced stage endometriosis compared with controls. KIR (NKB1) was also significantly increased in peritoneal NK cells of women with advanced-stage endometriosis, compared with those of women with early-stage endometriosis. CONCLUSION(S): The results of this study suggest that the decrease in peritoneal NK cytotoxicities against K562 is observed and that this disease may be partially due to the increased expression of KIR on these NK cells. PMID- 11119749 TI - Recurrent pregnancy loss and diminished ovarian reserve. AB - OBJECTIVE: To determine the incidence of diminished ovarian reserve (OR) in patients with recurrent pregnancy loss (RPL). DESIGN: Retrospective chart review. SETTING: Tertiary fertility center. PATIENT(S): Six hundred ninety-two women undergoing a fertility evaluation. INTERVENTION(S): Clomiphene citrate challenge test (CCCT). MAIN OUTCOME MEASURE(S): FSH concentrations measured on menstrual days 3 and 10. RESULT(S): Forty-four women were diagnosed with RPL (+RPL), and 648 women had non-RPL diagnoses (-RPL). Compared with -RPL women, women with +RPL were younger (following statistics are listed as +RPL vs. -RPL, respectively; 34 +/- 5 vs. 35 +/- 4 y) but had similar menstrual cycle length (29 +/- 4 vs. 28 +/- 4 d), and lower day 3 FSH levels (8.9 + 7 vs. 11 +/- 9 mIU/mL) and similar day 10 FSH levels (11 +/- 8 vs. 12 +/- 11 mIU/mL). Eight of 44 women with +RPL (18%) had an abnormal CCCT, compared with 117/648 (18%) of women in the -RPL group. For women with normal OR, delivery rates were similar for -RPL and +RPL patients. For women with an abnormal CCCT, delivery rates were < 5%. CONCLUSION(S): Women with RPL have a similar incidence of diminished OR as the general infertile population. Reproductive outcome for patients with an abnormal CCCT is equally poor for both groups. Ovarian reserve screening should be considered in the work up of RPL before initiation of anticoagulant or immunotherapy. PMID- 11119750 TI - Hyperhomocysteinemia and recurrent early pregnancy loss: a meta-analysis. AB - OBJECTIVE: To quantify the risk of recurrent early pregnancy loss in the presence of elevated fasting or afterload homocysteine concentrations or homozygosity for the 677C-->T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene (T/T genotype). DESIGN: Case-control studies published between January 1992 and November 1999 were identified with a MEDLINE-search. These studies were combined with a recent case-control study performed by our own research group. SETTING: Academic research environment. PATIENT(S): Studies published in the English language, concerning two or more pregnancy losses before 16 weeks' menstrual age were included. INTERVENTION(S): Meta-analysis of all of the studies included. MAIN OUTCOME MEASURE(S): The number of subjects with and without hyperhomocysteinemia or with the T/T genotype were derived, if necessary the study was supplemented by personal communication with the original authors. RESULT(S): Pooled risk estimates of 2.7 (1.4 to 5.2) and 4.2 (2.0 to 8.8) were calculated for fasting and afterload plasma homocysteine concentrations, respectively. For the MTHFR T/T genotype a pooled risk estimate of 1.4 (1.0 to 2.0) was found. CONCLUSION(S): These data support hyperhomocysteinemia as a risk factor for recurrent early pregnancy loss. Further research should be focused on the pathophysiology of this relationship and on the clinical efficacy of B vitamin supplementation. PMID- 11119751 TI - Effects of hydrogen peroxide on DNA and plasma membrane integrity of human spermatozoa. AB - OBJECTIVE: To evaluate the effects of oxidative stress on DNA and plasma membrane integrity of human spermatozoa. DESIGN: Prospective cohort study. SETTING: University-based, tertiary-care infertility center. PATIENT(S): Men (n = 10) undergoing infertility investigation. INTERVENTION(S): Purified populations of sperm with high motility were separated using Percoll density gradients. Then, spermatozoa were incubated with 0, 10, 100, and 200 microM hydrogen peroxide (H(2)O(2)) under capacitating conditions. MAIN OUTCOME MEASURE(S): Motion parameters were assessed by computer analysis. Genomic integrity was examined by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) assay. Plasma membrane integrity was evaluated by the annexin V-binding assay, a measure of phosphatidylserine translocation. RESULT(S): Under basal conditions, there was a significant and negative relationship between sperm motility and the percentages of sperm with DNA fragmentation and membrane translocation of phosphatidylserine. After a 2-h incubation, there was a significant, dose-dependent effect of H(2)O(2) on motion parameters (decrease) and DNA fragmentation (increase). The percentage of annexin V(-) live (normal) cells declined significantly as the level of oxidative stress increased. Although the percentages of annexin V(+) live cells (sperm depicting translocation of phosphatidylserine) and necrotic cells increased at the highest H(2)O(2) levels, these changes were not significant. CONCLUSION(S): In vitro sperm incubation with H(2)O(2) induces DNA fragmentation in a dose-dependent fashion. The sublethal effects of oxidative stress on motion parameters were not significantly associated with membrane translocation of phosphatidylserine. PMID- 11119752 TI - Ryudocan expression by luteinized granulosa cells is associated with the process of follicle atresia. AB - OBJECTIVE: To evaluate the presence of ryudocan in follicular fluid (FF) and its possible correlation with FF E(2) and P, and to study the levels of ryudocan in granulosa-lutein cells stimulated with hCG. DESIGN: Controlled clinical study and in vitro experiment. SETTING: University teaching hospital. PATIENT(S): One hundred seven patients undergoing IVF. INTERVENTION(S): The FF and granulosa lutein cells were aspirated from follicles 34 hours after an ovulatory gonadotropin bolus. MAIN OUTCOME MEASURE(S): FF ryudocan, E(2), and P levels as well as hCG-mediated induction of ryudocan. RESULT(S): Ryudocan was abundant in the FF; the concentration of ryudocan in human FF was estimated to be 305.5 +/- 200.8 ng/mL (mean +/- SD). Atretic follicles had higher concentrations of ryudocan (559.1 +/- 156.5 ng/mL). FF ryudocan levels were inversely correlated with FF E(2) (r = -0.5023) and P concentrations (r = -0.4459). A detectable amount of ryudocan was found in pooled granulosa-lutein cells. Ryudocan production was augmented by surge levels of hCG. CONCLUSION(S): Ryudocan is expressed in luteinized granulosa cells in vitro. The higher concentrations of ryudocan in FF of atretic follicles suggest an involvement of ryudocan in the process of atresia. PMID- 11119753 TI - Human oviductal cells reduce the incidence of apoptosis in cocultured mouse embryos. AB - OBJECTIVE: To investigate the effect of human oviductal cell coculture on the incidence of apoptosis in mouse embryos. DESIGN: Experimental laboratory study. SETTING: University gynecology unit. PATIENT(S): Fallopian tubes were obtained from patients undergoing hysterectomy. INTERVENTION(S): Mouse embryos were cocultured with human oviductal cells. MAIN OUTCOME MEASURE(S): Blastocyst development, allocation of inner cell mass (ICM) and trophectoderm (TE) in blastocyst, and apoptosis in embryos. RESULT(S): Oviductal cells significantly enhanced the blastulation (38%) and hatching rate (22%) of the cocultured zygotes. The corresponding values in medium alone culture were 21% and 9%, respectively. The cocultured embryos also had higher blastomere count at blastocyst stage (P<0. 005). This was due to increase in both the cell count of ICM (P<0. 05) and TE (P<0.001). Coculture reduced the incidence of apoptosis in the cultured morula and blastocyst from 38% and 48% to 16% (P<0. 001) and 27% (P<0.05), respectively. The number of apoptotic blastomeres per morula (1.5 +/- 0.6; P<0.005) and blastocyst (2.3 +/- 0.7; P<0.005) after coculture was also significantly lower than that of the corresponding control (morula, 2.1 +/- 0.8; blastocyst, 3.5 +/- 1.1). CONCLUSION(S): Human oviductal cells improved mouse embryo development partly by decreasing the incidence of apoptosis. PMID- 11119754 TI - Expression of vascular endothelial growth factor mRNA in human preimplantation embryos derived from tripronuclear zygotes. AB - OBJECTIVE: To detect the expression of vascular endothelial growth factor (VEGF) mRNA and/or secretion of VEGF protein by human preimplantation embryos. DESIGN: Human preimplantation embryos not suitable for uterine transfer were examined for beta-actin and VEGF mRNA expression. Culture media from normally fertilized and developing preimplantation embryos were assessed for VEGF protein secretion. SETTING: Clinics and academic research laboratories at the Departments of Obstetrics and Gynecology at the Stanford University, Palo Alto, California and the Heinrich-Heine-University, Dusseldorf, Germany. PATIENT(S): Couples undergoing IVF by intracytoplasmic sperm injection for various reasons. INTERVENTION(S): Six unfertilized oocytes and 33 pathologically fertilized (tripronucleic, 3PN) preimplantation embryos were examined for VEGF mRNA expression, and 16 embryos were examined for VEGF protein secretion. MAIN OUTCOME MEASURE(S): Embryonic expression of VEGF mRNA and VEGF protein as determined by reverse transcription (RT)/nested polymerase chain reaction (PCR) and ELISA. RESULT(S): VEGF mRNA and protein could not be detected in unfertilized oocytes. However, 30/33 preimplantation embryos did express VEGF mRNA (11/12 10-to-16-cell embryos, 3/4 morulae, 11/12 early blastocysts, 5/5 hatched blastocysts). The VEGF protein level was below the sensitivity of the ELISA. CONCLUSION(S): Production of VEGF may give the embryo the ability to induce neoangiogenesis at the implantation site, thus creating an environment necessary for its survival. PMID- 11119755 TI - Procalcitonin: does it play a role in male reproduction? AB - OBJECTIVE: To investigate the presence or absence of procalcitonin in the seminal plasma of both normal and subfertile men. DESIGN: Controlled clinical study. SETTING: Academic research environment. PATIENT(S): Healthy male volunteers and subfertile men with varicocele or infection of the male accessory glands. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Seminal plasma procalcitonin determinations. RESULT(S): Seminal plasma procalcitonin levels from subfertile men were not significantly different from those found in healthy volunteers. CONCLUSION(S): Although procalcitonin can be detected in the seminal plasma, these preliminary results indicate that it cannot be used as a diagnostic marker. PMID- 11119756 TI - Are women with polycystic ovary syndrome resistant to activated protein C? AB - OBJECTIVE: This study was performed to test the hypothesis that an increased prevalence of activated protein C (APC) resistance in women with polycystic ovary syndrome (PCOS) puts them at increased risk of miscarriage and thrombosis. DESIGN: Case control study. SETTING: A district general hospital in the United Kingdom. PATIENT(S): Forty-one women with PCOS and 25 controls. INTERVENTION(S): Clinical histories, ultrasound scans, and venepunctures. MAIN OUTCOME MEASURE(S): Diagnosis of PCOS or control, clinical histories, APC resistance according to an activated partial thromboplastin time-based assay. RESULT(S): There was no significant difference in the proportion of women with APC resistance in both groups (three women in the PCOS group [7%] vs. one woman in the control group [4%]). The prevalence of APC resistance in the entire study population was 6.5%. In the PCOS group, 29% (12/41) gave a positive family history of thrombosis compared with 8% (2/25) in the control group. None of the women with a positive family history of thrombosis had abnormal antithrombin 111, protein C, or protein S levels. CONCLUSION(S): This study suggests that women with PCOS may have the same prevalence of APC resistance as the background population and that APC resistance may not put them at a higher risk of thrombosis or miscarriage compared with the case of the general population. PMID- 11119757 TI - Association of AccI polymorphism in the follicle-stimulating hormone beta gene with polycystic ovary syndrome. AB - OBJECTIVE: To search for FSH beta-subunit gene mutations in patients with polycystic ovary syndrome (PCOS) and determine the association between the mutations and the syndrome. DESIGN: Clinical and molecular studies. SETTING: Clinics and laboratories of the National University Hospital Obstetrics and Gynecology Department in Singapore. PATIENT(S): One hundred thirty-five patients with PCOS and 105 normal control subjects. INTERVENTION(S): Exons two and three were screened for mutations by single-stranded conformational polymorphism and DNA sequencing. MAIN OUTCOME MEASURE(S): Polymerase chain reaction followed by restriction enzyme analysis. RESULT(S): No missense mutation was found in the functional units of the FSHbeta gene in patients with PCOS, but a thymine cytosine substitution in exon 3 (codon 76, TAT to TAC) was identified. The nucleotide change led to creation of an AccI digestion site. The distribution pattern of AccI polymorphism in the patients was significantly different from that in the control group, and the occurrence of homozygous carriers was significantly higher in patients (12.6%) than in the control group (3.8%). The frequency of polymorphism and prevalence of homozygosity were significantly higher in patients with PCOS with obesity (0.50% and 31.0%, respectively) than in those with menstrual disorders only (0.366% and 8.5%, respectively), which correlated with significantly higher androgen levels in the obese patients. CONCLUSION(S): The AccI polymorphism in FSHbeta gene may be associated with PCOS in some women, especially those with obesity. PMID- 11119759 TI - Dominant transmission of imperforate hymen. AB - OBJECTIVE: Imperforate hymen is an uncommon anomaly of the reproductive tract, occurring in approximately 0.1% of newborn females. The familial occurrence of imperforate hymen in a child, her mother, and her mother's monozygotic twin is reported. DESIGN: Case report. SETTING: Academic medical center. PATIENT(S): Three affected family members. MAIN OUTCOME MEASURE(S): Karyotype and pedigree analysis. RESULT(S): The proband, presenting with peritonitis, was evaluated at age 12 for imperforate hymen because this condition was diagnosed in her mother at age 14. At age 14, the mother's monozygotic twin was asymptomatic except for primary amenorrhea and was also demonstrated to have imperforate hymen. No other reproductive system abnormalities were known to be present in the remaining family members. Chromosomal structural analysis confirmed that the mother of the proband had no chromosomal abnormalities. CONCLUSION(S): The occurrence of imperforate hymen in two consecutive generations of a family is consistent with a dominant mode of transmission, either sex-linked or autosomal. Previously reported examples of siblings with imperforate hymen suggested a recessive mode of inheritance. Taken together, these cases suggest that imperforate hymen can be caused by mutations in several genes. This case highlights the importance of evaluating all family members of affected patients. Familial examples of other developmental anomalies of the female reproductive tract also suggest a multifactorial genetic etiology. PMID- 11119758 TI - A variant of the glucocorticoid receptor gene is not associated with adrenal androgen excess in women with polycystic ovary syndrome. AB - OBJECTIVE: To determine whether the frequency of the N363S variant of the glucocorticoid receptor (GRL) was increased in women with PCOS and adrenal androgen (AA) excess. DESIGN: Prospective case-control study. SETTING: University reproductive endocrinology laboratory and outpatient clinic. PATIENT(S): Consecutive patients of non-Hispanic white race diagnosed with PCOS (n = 114) and healthy controls (n = 92). INTERVENTION(S): Blood and DNA sampling before hormonal therapy. MAIN OUTCOME MEASURE(S): PCOS patient and healthy control genotypes, with the N363S allele representing a variant of GRL. RESULT(S): Fifty four PCOS patients with (DHEAS > or = 3000 ng/mL) and 55 without (DHEAS < or = 2,500 ng/mL) AA excess, respectively, were studied. Six of 109 (5.5%) patients studied were found to be heterozygous carriers of the A-->G base pair substitution at cDNA position 1220, resulting in the missense mutation N363S. Of these six, four had excessive AA secretion (i.e., excess DHEAS levels). There was no significant difference in the allele frequency of the GRL variant between PCOS patients with and without AA excess and controls (3.7% [95% confidence interval: 1.0%-5.7%], 1.8% [0.2%-6. 0%], and 3.3% [2.3%-6.0%]). None of the subjects were found to be homozygous for the N363S allele. CONCLUSION(S): The N363S variant of GRL was an uncommon occurrence in our population of healthy women and PCOS patients and did not appear to play a major role in the genetic predisposition to PCOS or to AA excess in PCOS. PMID- 11119760 TI - The activity (calcium oscillator?) responsible for human oocyte activation after injection with round spermatids is associated with spermatid nuclei. AB - OBJECTIVE: To compare the oocyte-activating ability of whole human round spermatids and their isolated nuclei. DESIGN: Prospective study using sibling oocytes from patients undergoing spermatid conception and intracytoplasmic sperm injection treatment cycles. SETTING: Private assisted reproduction laboratories and a university department. PATIENT(S): Couples with male infertility. INTERVENTION(S): Sibling oocytes were injected either with whole round spermatids or with their isolated nuclei, followed by artificial triggering of oocyte activation with calcium ionophore. Other sibling oocytes were injected either with isolated spermatid cytoplasm or with whole mature spermatozoa. MAIN OUTCOME MEASURE(S): Numbers of activated oocytes and cleaving embryos. RESULT(S): After oocyte activation was boosted with calcium ionophore, whole spermatids and isolated spermatid nuclei were equally effective in supporting oocyte activation and the formation of pronuclei, whereas no control oocyte was activated under the same conditions after injection of isolated spermatid cytoplasmic compartments. Cleavage rates were lower after the injection of isolated spermatid nuclei than after the injection of whole spermatids. CONCLUSION(S): The factor responsible for human oocyte activation after round spermatid injection is associated with spermatid nuclei. The requirement for the artificial trigger (calcium ionophore) suggests that this factor is identical to the male gamete activity previously characterized as calcium oscillator. PMID- 11119761 TI - Successful pregnancy and birth after sequential cotreatment with growth hormone and gonadotropins in a woman with panhypopituitarism: a new treatment protocol. AB - OBJECTIVE: To report a successful pregnancy in a woman with panhypopituitarism who received 3 months of pretreatment with growth hormone (GH) before ovulation induction. Prior attempts at ovulation induction had failed for this patient. DESIGN: Case report. SETTING: Department of Endocrinology. PATIENT(S): A 32-year old woman with panhypopituitarism and secondary infertility. INTERVENTION(S): GH (1 IU/day) alone for 3 months; during the next cycle, 1 IU/day of GH; 3 ampules of hMG per day during days 1-21; 1 ampule of hCG on day 21. GH was discontinued on day 35 when a pregnancy test was positive. MAIN OUTCOME MEASURE(S): Pregnancy and delivery. RESULTS: Pregnancy and birth of a normal child after a single ovulation stimulation using GH and gonadotropins. CONCLUSION(S): This case report suggests interest in a new protocol for follicular stimulation in women with hypopituitarism who are responding poorly to gonadotropin therapy. PMID- 11119762 TI - Chronic cough and infertility: a report of two cases. AB - OBJECTIVE: To report two cases of infertility caused by primary ciliary dyskinesia in patients who presented with an associated complaint of a chronic cough. DESIGN: Case report. SETTING: University teaching hospital. PATIENT(S): Two patients presenting with unexplained infertility and an associated history of long-term chronic cough. INTERVENTION(S): Patients underwent a nasal mucosal biopsy by an otolaryngologist. Electron microscopy (EM) examination of biopsy specimens was performed. MAIN OUTCOME MEASURE(S): Diagnosis and appropriate treatment for functional tubal factor infertility. RESULT(S): Both patients were diagnosed with primary ciliary dyskinesia based on EM of the nasal biopsy specimens. Given this diagnosis, they immediately underwent IVF-ET. Both patients became pregnant with their first IVF-ET cycle. CONCLUSION(S): Other investigators have shown that almost 20% of patients with a chronic cough will have EM evidence of ciliary dyskinesia. Patients presenting with idiopathic infertility and an associated unexplained chronic cough should be referred for nasal biopsy with EM evaluation to rule out primary ciliary dyskinesia. Infertility in these cases, which is due to a functional tubal factor, is best treated with IVF-ET rather than superovulation and intrauterine insemination treatments. PMID- 11119763 TI - 46,XY monozygotic twins with discordant sex phenotype. AB - OBJECTIVE: To analyze male and female sex differentiation in monozygotic twins. DESIGN: Retrospective study. SETTING: Multiple academic centers. PATIENT(S): A pair of monozygotic twins. INTERVENTION(S): Skin and blood samples were obtained for DNA analysis and karyotyping. MAIN OUTCOME MEASURE(S): Mutation within the SRY gene was analyzed by the polymerase chain reaction-single-stranded conformation polymorphism test. Monozygosity was ascertained by short tandem repeat analysis. Karyotypes were studied in blood and skin fibroblasts. RESULT(S): SRY was present in both twins, but no mutations were detected in the SRY conserved motif. Monozygosity was confirmed by the use of short tandem repeat analysis in four loci: c-fms, thyroid peroxidase, von Willebrand factor, and tyrosine hydroxylase. The karyotype was 46,XY uniformly in both twins. CONCLUSION(S): Monozygotic twins can develop discordant male and female phenotypes despite the presence of a common karyotype and despite the presence of intact testis-determining genes. In the present case, this could be due to mutation or to mosaicism involving occult 45,X cell lines in the dysgenetic gonads. PMID- 11119764 TI - Glutathione levels and miscarriage. PMID- 11119766 TI - Erratum PMID- 11119765 TI - Role of chromosome abnormalities in ectopic pregnancy. PMID- 11119767 TI - Carbon dioxide release in Coptotermes formosanus Shiraki and Reticulitermes flavipes (Kollar): effects of caste, mass, and movement. AB - Movement and carbon dioxide (CO(2)) release of individual Formosan, Coptotermes formosanus Shiraki and Eastern, Reticulitermes flavipes (Kollar) subterranean termites were recorded simultaneously in real time. Worker, soldier, and pre alate (nymph) caste termites were recorded over 1-h periods at ambient temperature and normoxia in dry, CO(2)-free air. No evidence of discontinuous gas exchange cycles (DGCs) was observed in 344 recordings. Intensity of movement was constant in video tape recordings of termites under respirometry conditions. Duration of movement did not have a significant effect on residuals of &Vdot;(CO(2)) regressed on mass. Thus, movement did not effect &Vdot;(CO(2)) for these two species. Overall CO(2) release values were calculated for all recordings resulting in mean &Vdot;(CO(2)) (ml CO(2) g(-1) h(-1)), and compared among caste, colony, and species with a nested ANOVA. There was significant interaction (P=0.0161) only for species. Mean CO(2) release was significantly greater for R. flavipes (0.507 ml CO(2) g(-1) h(-1)) than C. formosanus (0.310 ml CO(2) g(-1) h(-1)). Mass scaling of termite &Vdot;(CO(2)) was investigated by regressing log(10)&Vdot;(CO(2)) on log(10)mass. The overall model combining species gave a mass scaling coefficient of 0.861(+/-0.0791), which approximates a previously published value for the arthropods as a whole (0.825). PMID- 11119768 TI - Procathepsin and acid phosphatase are stored in Musca domestica yolk spheres. AB - Yolk spheres present in mature invertebrate oocytes are composed of yolk proteins and proteolytic enzymes. In the fly Musca domestica, yolk proteins are degraded during embryogenesis by a cathepsin-like proteinase that is stored as a zymogen. An acid phosphatase is also active in the yolk spheres during Musca embryogenesis. In this paper we show that procathepsin and acid phosphatase are initially stored by a different pathway from the one followed by yolk protein precursors. Both enzymes are taken up by the oocytes and transitorily stored into small vesicles (lysosomes) surrounding the early yolk spheres. Fusion of both structures, the early yolk spheres and lysosomes, creates the mature yolk spheres. PMID- 11119769 TI - Immunolocalization and possible effect of a moth allatotropin-like substance in a fly, Phormia regina (Diptera: Calliphoridae). AB - Insect allatotropin upregulates the biosynthesis of juvenile hormones by the corpus allatum. We raised two rabbit antisera against the allatotropin of Manduca sexta (Mas AT) using a synthetic, multiple-antigenic-peptide that contains a branching heptalysine core and eight Mas AT molecules. Both antisera recognized specifically the same neurons in the larval brain, frontal ganglion and terminal abdominal ganglion of M. sexta as previously reported by others. Immunoassay showed reactivity specific to the Mas AT. Very low or nearly no cross-reactivity was found for two Mas AT-like peptides, a myotropin from Locusta migratoria and a Mas AT-like peptide deduced from the DNA sequence of Aedes aegypti, respectively. Immunopositive neurons also were identified in adult Phormia regina, Dacus dorsalis, Oncopeltus fasciatus, and Mythimna loreyi, and in larval M. loreyi, Bombyx mori, and Andraca bipunctata. At 20 pmol per 25 ul incubation medium (i.e. 8x10(-7) M), synthetic Mas AT significantly stimulated in vitro juvenile hormone biosynthesis by the corpus allatum of adult, sugar-fed females of P. regina to 2.64-fold that of controls. Thus, this study provides the first demonstration that at the higher end of the physiological concentration range, the Mas AT has allatotropic effect in vitro to CA of non-lepidopterans. However, in vivo functions of Mas AT and/or Mas AT-like peptide in P. regina remain to be defined. PMID- 11119770 TI - The structural mechanism of trypsin-induced intrinsic motility in Manduca sexta spermatozoa in vitro. AB - Lepidopteran males produce eupyrene (nucleate) and apyrene (anucleate) spermatozoa, but in the female only eupyrene spermatozoa leave the spermatheca and fertilize the eggs. Both kinds of spermatozoa lack intrinsic motility in the male genital duct. They become motile in the spermatophore, in a process involving proteases from the male duct. In vitro, trypsin induces immotile spermatozoa to become motile. We studied the changes spermatozoa of Manduca sexta undergo during trypsin-induced motility and found that (a) they mimick rather closely those occurring in vivo during normal sperm maturation in genital ducts and (b) they are time- and dose-dependent. As in vivo, they comprise, successively, (a) disappearance of an extracellular matrix that maintains the integrity of eupyrene bundles in the seminal vesicle, (b) dispersion of the eupyrene bundles and intermingling of eupyrene and apyrene spermatozoa and (c) "hatching" of eupyrene spermatozoa from individual enclosing envelopes that are formed in the seminal vesicle. "Hatching" may not directly be related to motility since eupyrene spermatozoa become motile before "hatching" and motile apyrene spermatozoa never "hatch". Rather "hatching" may be related to the capacitation of eupyrene spermatozoa to either leave the spermatheca or fertilize the eggs, or both, as neither apyrene spermatozoa, nor those eupyrene spermatozoa that fail to "hatch", leave the spermatheca. PMID- 11119771 TI - Development of teratocytes associated with Microctonus aethiopoides Loan (Hymenoptera: Braconidae) in natural and novel host species. AB - A laboratory study investigated development of teratocytes derived from the parasitoid Microctonus aethiopoides Loan in the natural host, Sitona discoideus Gyllenhal, and in three novel hosts, the introduced weed biological control agent Rhinocyllus conicus (Froehlich), and two New Zealand native species Nicaeana cervina Broun and Irenimus stolidus Broun. Weevils were exposed to parasitoids and then examined 6, 10 and 15 days post-parasitism for parasitoid stage and size, and teratocyte number and size. In all hosts, teratocyte numbers decreased and size increased as parasitoid development progressed, although 6 days after parasitism, fewer, larger teratocytes were found in I. stolidus than S. discoideus or N. cervina. In weevils containing second-third instar parasitoid larvae, the most permissive hosts, S. discoideus and N. cervina contained more teratocytes than the least permissive hosts I. stolidus and R. conicus. Host gender influenced some aspects of parasitoid and teratocyte development. Total teratocyte volume was greater in female than male S. discoideus at all sampling times, and at 10 days post-parasitism in N. cervina. A possible relationship between host suitability and teratocyte development is discussed. PMID- 11119772 TI - Leucokinin and the modulation of the shunt pathway in Malpighian tubules. AB - Transepithelial secretion in Malpighian tubules of the yellow fever mosquito (Aedes aegypti) is mediated by active transport of Na(+) and K(+) through principal cells and passive Cl(-) transport through the shunt. Permeation through the shunt was assessed by measuring transepithelial halide diffusion potentials in isolated perfused Malpighian tubules, after first inhibiting active transport with dinitrophenol. Diffusion potentials were small under control conditions, revealing Eisenman selectivity sequence I (I(-)>Br(-)>Cl(-)>F(-)) which is the halide mobility sequence in free solution. Accordingly, electrical field strengths of the shunt are small, selecting halides for passage on the basis of hydrated size. Leucokinin-VIII (LK-VIII) significantly increased the shunt conductance from 57.1 uS/cm to 250.0 uS/cm. In parallel, the shunt selectivity sequence shifted to Eisenman sequence III (Br(-)>Cl(-)>I(-)>F(-)), revealing increased electrical field strengths in the shunt, now capable of selecting small, dehydrated halides for passage. High concentrations of peritubular F(-) (142.5 mM) duplicated the effects of LK-VIII on shunt conductance and selectivity, suggesting a role for G-protein. In the presence of LK-VIII (or F( )), coulombic interactions between the shunt and I(-) and F(-) may be strong enough to cause binding, thereby blocking the passage of Cl(-). Thus, LK-VIII increases both shunt conductance and selectivity, presumably via G-protein. PMID- 11119773 TI - Crustacean cardioactive peptide is a modulator of oviduct contractions in Locusta migratoria. AB - Crustacean cardioactive peptide (CCAP) stimulates the contractions of locust oviducts. CCAP increased the basal tonus and increased the frequency and amplitude of phasic contractions, as well as the amplitude of neurally-evoked oviduct contractions in a dose-dependent manner. Oviducts from Vth instar larvae and adult locusts aged 10 days or less, were more sensitive to CCAP than oviducts from adult locusts aged 12 days or more. This may be indicative of a differential expression of number or subtypes of CCAP receptors on the oviducts at different ages, and may be related to reproductive functions or to functions of CCAP on the oviducts during ecdysis. The oviducts appear more sensitive to CCAP when compared with previously published reports of CCAP actions on the hindgut. CCAP actions on the amplitude of neurally-evoked contractions of the oviducts are similar to those of proctolin, however, the oviducts are more sensitive to CCAP. No CCAP like immunoreactive structures were discovered in the nerves innervating the oviducts, or on the oviducts themselves, confirming the previously published suggestion (Dircksen et al., 1991) that CCAP acts as a neurohormone at the oviducts. Cells showing CCAP-like immunoreactivity were discovered in the fat body associated with the oviducts and represent a potential source of CCAP, along with CCAP released from the transverse nerve and perivisceral organs. PMID- 11119774 TI - Efficacy of native FXPRLamides (pyrokinins) and synthetic analogs on visceral muscles of the American cockroach. AB - Six pyrokinins, members of a widely distributed neuropeptide family in insects (FXPRLamides), have been identified from the American cockroach. Five of these peptides, Pea-PK-1-5, were tested in different myotropic bioassays, including hyperneural muscle, hindgut, foregut and oviduct. Among these muscles, the hyperneural muscle exhibited the highest sensitivity to pyrokinin applications. The efficacy of the different pyrokinins differed dramatically. No muscle specific effectiveness was obtained; the ranking order in all muscle assays was as follows: PK-1>PK-4>PK-3>PK-2>PK-5. Testing of synthetic analogs revealed the importance of the amino acid at the variable -4 position of the C-terminus. PK-5, the only one of the five tested peptides which is stored in abdominal perisympathetic organs, has probably no myotropic function at all. This is further evidence that these neurohemal release sites are not necessary to compensate the open circulatory system of insects but have rather specific functions which are totally unknown as yet. PMID- 11119775 TI - Changes in biosynthesis and degradation of juvenile hormone during breeding by burying beetles: a reproductive or social role? AB - Burying beetles, Nicrophorus orbicollis, depend on the location of an unpredictable resource, a small vertebrate carcass, for reproduction. When they discover a carcass, they undergo a correlated rapid rise in titers of juvenile hormone (JH) in the hemolymph and ovarian development. This study investigates the regulation of the changes in JH during breeding in both male and female burying beetles and the role of JH in ovarian development. JH biosynthesis by the corpora allata (CA), measured in vitro, increased in females within an hour of their discovery of a carcass and increased later in males. After returning to low rates as oviposition began, JH biosynthesis rose again 3 days later in females but not in males. Neither the ovaries nor testes synthesized JH. There was a concomitant fall in JH esterase activity within 12 h of discovery of the carcass in both males and females. Although the rise in JH titers and biosynthesis and the fall in JH esterase is correlated with ovarian development, application of methoprene or JH III in the absence of a carcass did not result in vitellogenin uptake by the oocytes. Therefore, we conclude that, in spite of the rapid rise in JH before oviposition, it is not sufficient to regulate vitellogenin synthesis and/or its uptake by the ovaries. We suggest that its role has been preempted to organize social behavior and coordinate parental behavior between mates. PMID- 11119776 TI - Inhibition of host-seeking response and olfactory responsiveness in Anopheles gambiae following blood feeding. AB - The effect of a single blood meal on the host-seeking response of Anopheles gambiae was investigated in the laboratory using a behavioural bioassay, whereas possible changes at the chemosensory level were monitored using electroantennogram recording (EAG). To avoid the possible confounding effect of body size, mosquitoes of a large size class only were used. Five-day old female mosquitoes were given a blood meal on a human arm and exposed to the emanations of a human hand in an olfactometer at 3, 24, 40, 48 and 72 h following the meal and their behaviour and EAG response to host stimuli were compared with that of unfed mosquitoes (controls) of corresponding age. During egg development, mosquitoes had access to glucose and an oviposition tray. The ovarian development of blood-fed mosquitoes that responded to host odours was compared with that of blood-fed mosquitoes that had not been exposed to host odours. The EAG response of blood-fed and control mosquitoes to host odour was examined upon stimulation with air led over incubated human sweat, hexanoic acid, indole and geranyl acetone. EAGs were recorded at times after a blood meal corresponding with those used in the behavioural experiment. There was no host-seeking response at 3 and 24 h post blood meal (pbm). Seven percent of the mosquitoes responded to human emanations 40-h pbm, 27% at 48 h and 68% at 72 h following a blood meal. The average response of controls to host stimuli varied from 35 (at t=40 h) to 67%. There was no ovarian development in the unfed group of mosquitoes. Of the mosquitoes that responded to host odour 48 h pbm, 12.5% (n=5) had ovaries in Christophers' stage IV and the remainder in stage V. Of the mosquitoes that responded 72 h pbm, 66.7% (n=94) had ovaries in stage V and 31.2% (n=44) had recently oviposited. Maximum EAG amplitudes recorded from blood-fed and control mosquitoes were similar for mosquitoes in Christophers' stages I-III, whereas in stage IV EAG amplitudes recorded from the blood-fed group were significantly lower than those of the corresponding control group in response to headspace of incubated human sweat and to indole. The results show that there was a strong inhibition of host seeking in An. gambiae for a period of at least 40 h following a blood meal. Host-seeking returned to pre-blood meal levels 72-h post feeding and was associated with egg maturation. The inhibition of host-seeking behaviour was accompanied by an inhibition of olfactory sensitivity to headspace of incubated sweat and indole just before the resumption of the host-seeking response. The implications of these findings for mosquito surveillance with host odours are discussed. PMID- 11119777 TI - Dealing with doubt. How patients account for non-specific chronic low back pain. AB - OBJECTIVE: To explore the ways that persons with long standing chronic low back pain respond to the problem of medical doubt about the presence of organic pathology. METHOD: Qualitative analysis of accounts provided by 12 persons attending a back pain rehabilitation clinic in NW England. RESULTS: Subjects rejected the notion that they were culpable for their pain. They were not culpable for the onset of their pain. They argued that despite their cooperation, no sensible explanation of their pain was forthcoming from health professionals. Finally, they asserted that medical scepticism had been damaging and dispiriting. CONCLUSION: Patients dealt with clinical doubt by stressing their own expertise. They constituted their beliefs about the cause and trajectory of their pain and disability as accurate accounts of their disability. They resisted the suggestion that there might be psychological factors involved in their ill-health by locating culpability among clinicians, who were confused or uncertain about diagnosis and treatment. PMID- 11119778 TI - Social desirability scores are associated with higher morning cortisol levels in firefighters. AB - OBJECTIVE: To examine the association of social desirability (SD) responding ("defensiveness") and cortisol levels. METHODS: Marlowe-Crowne SD scores and morning salivary cortisol were measured in firefighters. RESULTS: SD scores were associated with higher cortisol levels (r=.28) in 60 firefighters under age 45, but not in 25 older firefighters (or the combined sample). CONCLUSION: The results are consistent not only with SD scores being an indicator of coarctation of experience as a coping mechanism, but also (at least for younger persons) perhaps a risk factor for the psychobiological dysfunctions associated with chronic elevations of cortisol. PMID- 11119779 TI - Effects of depression and anxiety on mortality and quality-of-life 4 months after myocardial infarction. AB - OBJECTIVE: The purpose of this study was to determine the impact of depression and anxiety on mortality and quality-of-life in patients hospitalized for an acute myocardial infarction (MI). METHODS: Questionnaire measures of depression and anxiety were completed during hospitalization by 288 MI patients. The main outcomes were mortality and quality-of-life, assessed by the Dartmouth COOP charts, at 4 months. RESULTS: A total of 25 patients died, 22 from cardiac causes, during the 4-month follow-up. Symptoms of depression and anxiety did not predict either cardiac or all-cause mortality. Severity of infarction, extent of heart failure, and a longer stay in hospital predicted mortality. Symptoms of depression and anxiety predicted 4-month quality-of-life among survivors, as did gender, partner status, occupational status, living alone, previous exercise behaviour, length of hospital admission, and Peel Index scores. In a multiple regression model, depression emerged as the strongest predictor of quality-of life. State anxiety, severity of infarction, and partner status also entered the model. CONCLUSION: Neither depression nor anxiety predicted mortality 4 months after MI. Both depression and anxiety predicted quality-of-life at 4 months among survivors. PMID- 11119780 TI - Stress and airway resistance in children with asthma. AB - OBJECTIVE: The present study implements an experimental paradigm to examine airway reactivity to stress in children with asthma and controls. METHOD: 114 children with asthma and 30 controls (ages 9-15) participated. The protocol involved 5 min of baseline physiological measurements followed by a 5-min stressful task. Skin conductance (EDG), skin temperature, and heart rate were measured continuously. Airway resistance was measured at baseline and after the task. RESULTS: 110 children (76% of the sample) were significantly "stressed" as shown by physiological changes. Asthmatics and controls differed on overall airway resistance, F(1, 108)=12.3, P<.001. The entire sample demonstrated a trend toward increased airway resistance in response to stress, F(1,108)=3.1, P<. 08. A portion of asthmatics (22%) had increases of greater than 20% of baseline airway resistance. Changes in airway resistance in response to stress were unrelated to asthma severity, F(2,78)=2.0, ns. CONCLUSION: Children with asthma and controls demonstrate variation in airway function in response to stress, although increases are likely more meaningful for children with asthma. Further research is needed to examine the mechanisms underlying this response. PMID- 11119781 TI - Internalized anger, self-control, and mastery experience in inpatient anorexic adolescents. AB - OBJECTIVES: To evaluate the implications of internalized anger, self-control and experience of mastery for adolescent girls with severe anorexia nervosa (AN). METHODS: Internalized and externalized anger, internal and external control, mastery, use of methods for self-control, and severity of anorexic symptoms were measured by self-report questionnaires in inpatient anorexic teenagers (N=26), inpatient female adolescent psychiatric patients (N=24), and a normal female comparison group (N=29). RESULTS: Internalized anger was significantly higher in both the anorexic and general psychiatric patients as compared to normal controls, but this difference was significant only for the anorexic patients. Anorexic and general psychiatric patients experienced significantly less mastery than normal controls, but again this difference was significant only in the anorexic group. Within the anorexic group, severity of symptoms correlated significantly with internalized anger, low mastery, and external locus of control, and negative significant correlations among control measures and anger were found. Total length of hospitalization correlated positively with internalized anger only for the anorexics. CONCLUSION: The findings support the notion that internalized anger and defective experience of self-control are important factors in the psychopathology of adolescent anorexic inpatient females The results may have implications for the clinical management of patients with severe AN. PMID- 11119782 TI - Type D personality. A potential risk factor refined. AB - OBJECTIVE: Acute and chronic psychological distress have been associated with coronary heart disease (CHD) but little is known about the determinants of distress as a coronary risk factor. Broad and stable personality traits may have much explanatory power; this article selectively focuses on negative affectivity (NA; tendency to experience negative emotions) and social inhibition (SI; tendency to inhibit self-expression in social interaction) in the context of CHD. METHODS: The first part of this article reviews research on NA and SI in patients with CHD. The second part presents new findings on NA and SI in 734 patients with hypertension. RESULTS: Accumulating evidence suggests that the combination of high NA and high SI designates a personality subtype ("distressed" type or type D) of coronary patients who are at risk for clustering of psychosocial risk factors and incidence of long-term cardiac events. Type D and its contributing low-order traits (dysphoria/tension and reticence/withdrawal) could also be reliably assessed in a community-based sample of patients with hypertension. This finding was replicated in men and women, and in Dutch- and French-speaking subjects. Type D hypertensives reported more depressive affect than their non type D counterparts. CONCLUSIONS: There is an urgent need to adopt a personality approach in the identification of patients at risk for cardiac events. NA and SI are broad and stable personality traits that may be of special interest not only in CHD, but in other chronic medical conditions as well. PMID- 11119783 TI - Metabolic control and psychological sense of control in women with diabetes mellitus. Alternative considerations of the relationship. AB - PURPOSE: Identifying psychological strategies to buffer the adverse outcomes in people with diabetes mellitus (DM) remains a priority for many health professionals. While 'locus of control' (LOC) has been repeatedly investigated to this end, research findings are contradictory. The development of more complex appraisals of psychological control, and the utilization of control inventories deriving from such analyses, presents a way forward from such contradictions. METHODS: Employing such a measure, this study examines the relationship between metabolic control and psychological sense of control in 96 women with DM. RESULTS: Optimal metabolic control is significantly associated with overall sense of control, while poor metabolic control was significantly associated with experiences of loss of psychological control and feelings of inadequacy. Furthermore, poor metabolic control was significantly associated with reduced control in the specific domains of interpersonal relationships and bodily functions. CONCLUSIONS: Multidimensional control inventories enable a more complex appraisal of the relationship between metabolic control and psychological control, and in doing so, provide a way forward from problems arising from reliance on LOC constructs. Interventions for DM management relying on aspects of psychological control need to target domains beyond traditional issues of self and bodily functions. PMID- 11119784 TI - Animal-assisted therapy - magic or medicine? AB - A sound theoretical basis supported by scientifically measured physiological parameters is needed to gain medical support for animal-assisted therapy. Six neurochemicals associated with a decrease in blood pressure were measured in humans (n=18) and dogs (n=18) before and after positive interaction. Results (P<.05) indicated that in both species the neurochemicals involved with attention seeking or attentionis egens behavior have increased. This information can be used as a rationale for animal-assisted therapy. PMID- 11119785 TI - Asian transplant registry: 1994-1998. PMID- 11119786 TI - T-cell antigen recognition and costimulatory pathways: implications for the induction of transplantation tolerance. PMID- 11119787 TI - Delayed graft function, rejection, and long-term prognosis. PMID- 11119788 TI - New frontiers in transplantation. PMID- 11119789 TI - Meeting the challenges of transplantation. PMID- 11119790 TI - Simulect: redefining immunosuppressive strategies. PMID- 11119791 TI - Immunosuppressive therapies for the twenty-first century. PMID- 11119792 TI - Cyclosporine formulations: is there really a difference? PMID- 11119793 TI - Strategies and obstacles in an organ donation program in developing countries: Saudi Arabian experience. PMID- 11119794 TI - Microemulsion cyclosporine postmarketing surveillance in renal transplant recipients: multicenter study in Korea. PMID- 11119795 TI - Compensated living nonrelated organ donation: an Asian perspective. PMID- 11119796 TI - Cultural and societal issues in organ transplantation: examples from different cultures. PMID- 11119797 TI - Transplantation in developing countries: economics, reality, and solutions. PMID- 11119798 TI - Constraints in living donor kidney transplantation. PMID- 11119799 TI - Outcome of overseas kidney transplantation in Malaysia. PMID- 11119800 TI - Organ donation-third-party donation: expanding the living-donor pool. PMID- 11119801 TI - Vanishing distinctions in transplantation academia-industry relationships. PMID- 11119802 TI - Immunologic monitoring: implications for treatment of rejection. PMID- 11119803 TI - Posttransplant hypertension. PMID- 11119805 TI - Malignancies in renal transplant recipients. PMID- 11119804 TI - Hyperlipidemia: impact of therapy. PMID- 11119806 TI - Infections in transplant recipients in developing countries. PMID- 11119807 TI - Hepatitis in renal transplant recipients. PMID- 11119808 TI - Immunosuppression withdrawal in renal transplantation. PMID- 11119809 TI - Effective strategies to prevent cytomegalovirus disease after renal transplantation. PMID- 11119810 TI - Use of non-heart-beating donors. PMID- 11119811 TI - Surgical techniques in right lobe liver transplantation. PMID- 11119812 TI - Understanding and preventing late graft loss. PMID- 11119813 TI - Pitfalls in anesthesia for liver transplantation. PMID- 11119814 TI - Liver transplantation: late complications. PMID- 11119815 TI - Heart transplantation: the standard versus bicaval technique controversy. PMID- 11119816 TI - Finer techniques in lung transplantation. PMID- 11119817 TI - Mechanical assist device; my choice: the CardioWest total artificial heart. PMID- 11119818 TI - Problems with mechanical devices. PMID- 11119819 TI - Heart transplantation after mechanical circulatory support. PMID- 11119821 TI - Status of thoracic organ transplantation in Saudi Arabia. PMID- 11119820 TI - Management of candidates for heart transplantation in Japan. PMID- 11119822 TI - Bridge to transplantation: selection and timing. PMID- 11119823 TI - Alternatives to cadaveric lung transplantation. PMID- 11119824 TI - Transplantation of hematopoietic stem cells from unrelated volunteer donors. PMID- 11119825 TI - The role of HLA matching in unrelated donor bone marrow transplant. PMID- 11119826 TI - Approach to invasive fungal infection after blood or marrow transplantation. PMID- 11119827 TI - Non-FcR-binding, humanized anti-CD3 antibody Hu291 induces apoptosis of human T cells more effectively than OKT3 and is immunosuppressive in vivo. PMID- 11119828 TI - Physiologic barriers to xenotransplantation. PMID- 11119829 TI - Xenotransplantation and cloning: working with the World Health Organization to develop ethical guiding principles. PMID- 11119830 TI - Zoonosis in xenotransplantation. PMID- 11119831 TI - The impact of cyclosporin formulation on clinical outcomes. PMID- 11119832 TI - Neoral monitoring: limitations of trough level monitoring and the potential role of limited sampling strategies. PMID- 11119833 TI - Increasing the donor supply from the United Kingdom's Asian population: the need for further research. PMID- 11119834 TI - Refusal of consent for organ donation: from survey to bedside. PMID- 11119835 TI - Effect of triiodothyronine replacement therapy on maintenance characteristics and organ availability in hemodynamically unstable donors. PMID- 11119836 TI - Potential organ donor pool for renal transplantation in the intensive care unit and emergency room. PMID- 11119837 TI - An analysis of status of cadaver donors at The National Taiwan University Hospital: eleven-year case review. PMID- 11119838 TI - Living kidney donation made easier. PMID- 11119839 TI - Reaching out to Asia for living kidney donors. PMID- 11119840 TI - Preliminary results of selection criteria for cadaveric kidney transplantation by the Thai Red Cross. PMID- 11119841 TI - Porcine liver transplantation from non-heart-beating cadaver donor: effect of initial flushing with cold versus warm UW solution. PMID- 11119842 TI - Successful use of graft from marginal donors in non-heart-beating renal transplantation. PMID- 11119843 TI - Living liver donation for adult living-related liver transplantation. PMID- 11119844 TI - Challenges in laparoscopic donor nephrectomy and technical innovations to make it cost effective. PMID- 11119845 TI - Benefits of spiral CT angiography in preoperative assessment of living renal donors. PMID- 11119846 TI - The effect of donor age on living-related kidney transplantation. PMID- 11119847 TI - Experience with laparoscopic donor nephrectomy in India: is the open approach justified any longer? PMID- 11119848 TI - Use of horseshoe kidney as renal transplant from living donor: its surgical feasibility and pitfalls. PMID- 11119849 TI - Analysis of the renal transplant waiting list at the National Taiwan University Hospital: eleven-year case review. PMID- 11119850 TI - Kidney transplantation from living donors with renal artery disease. PMID- 11119851 TI - A case of renal transplantation received from a living donor with second generation anti-hepatitis C virus antibody positive into an anti-hepatitis C virus antibody negative recipient. PMID- 11119852 TI - The impact of donor serum creatinine level on long-term outcome of renal allografts. PMID- 11119853 TI - Kidney transplantation from a donor who is HCV antibody positive and HCV-RNA negative. PMID- 11119854 TI - Spouse donor kidney transplantation in Thailand. PMID- 11119855 TI - Transplanting horseshoe kidneys: the Maastricht experience. PMID- 11119856 TI - Does the high level of lactate dehydrogenase predict renal function and outcome after renal transplantation from non-heart-beating cadaver donors? PMID- 11119857 TI - Quality of life after cadaveric renal transplantation from a non-heart-beating donor. PMID- 11119858 TI - In situ renal cooling for kidney transplantation from non-heart-beating donors. PMID- 11119859 TI - The impact of the establishment of a multiorgan transplantation program on cold ischemia time and delayed graft function in renal transplantation. PMID- 11119860 TI - The role of an e-mail list system for communication among medical professionals, transplant candidates, recipients, and their families in Japan. PMID- 11119861 TI - Prevention of warm ischemic injury in rat liver transplantation by geranylgeranylacetone. PMID- 11119862 TI - University of Wisconsin preservation solution enhances intrarenal nitric oxide production. PMID- 11119863 TI - Furosemide prevents the inhibitory effect of cyclosporine on intrarenal nitric oxide production. PMID- 11119864 TI - Cyclosporine nephrotoxicity: the mechanisms of cell injury by cyclosporine A in renal proximal tubular cells. PMID- 11119865 TI - Effect on rat kidney preservation in the cold of various sugars added to Euro Collins solution to adjust osmolarity. PMID- 11119866 TI - Porcine liver transplantation from non-heart-beating cadaver donors: effect of passive/active venovenous bypass on graft function. PMID- 11119867 TI - New immunosuppressive reagent, FTY 720, spares immunologic memory. PMID- 11119868 TI - Rescuing acute rejection of rat cardiac allografts with FTY720. PMID- 11119869 TI - Induction of heat shock protein-70 (hsp-70) by intraarterial administration of geranylgeranylacetone. PMID- 11119870 TI - Functional integrity of the rat liver after subzero preservation under high pressure. PMID- 11119871 TI - Cytokine-mediated, deteriorative effects of brain death on porcine liver transplantation: intervention of sympathoadrenal pathway in cerebrohepatic organ interaction. PMID- 11119872 TI - FR167653 ameliorates intestinal allograft ischemic injury. PMID- 11119873 TI - Beneficial effect of machine perfusion preservation on liver transplantation from non-heart-beating donors. PMID- 11119875 TI - Role of inducible nitric oxide synthase on hepatic ischemia and reperfusion injury. PMID- 11119874 TI - Peri- and postoperative kinetics of endothelin-1/big endothelin-1 and effects of endothelin antagonist in porcine liver transplantation from non-heart-beating donors. PMID- 11119876 TI - Hypoxic preconditioning reduces ischemia/reperfusion-induced apoptosis cell death in rat kidney. PMID- 11119877 TI - Assessment of cartilage viability in the cryopreserved tracheal allograft by measurement of Na(2)(35)SO(4) incorporation. PMID- 11119878 TI - De novo protein synthesis induced by donor nutritional depletion ameliorates cold ischemia and reperfusion injury in rat liver. PMID- 11119879 TI - Improvement of modified two-layer preservation method (PFC/Kyoto solution) in islet isolation from breeder pigs. PMID- 11119880 TI - Alanylglutamine-enriched total parenteral nutrition prevents bacterial translocation after small bowel transplantation in pigs. PMID- 11119881 TI - Induction of heat shock protein-70 (hsp-70) reduces preservation injury in rat IEC-18 intestinal epithelial cells. PMID- 11119882 TI - Heme oxygenase-1 expression in rat Kupffer cells by immunomodulating agents: role in the pathogenesis of hepatic ischemia/reperfusion injury. PMID- 11119883 TI - Islet graft primary nonfunction and its prevention. PMID- 11119884 TI - An experimental study on immune response to regenerating hepatic allografts in the rat. PMID- 11119885 TI - Roles of the organ transplantation coordination team at the National Taiwan University Hospital. PMID- 11119886 TI - Current organ preservation solutions for liver preservation: experimental comparison. PMID- 11119887 TI - Measurement of tissue P(T)O(2) in reperfusion injury of isolated rat liver. PMID- 11119888 TI - Trial of steroid withdrawal in renal transplant recipients with long-term- surviving allograft. PMID- 11119889 TI - Acute rejection in renal transplant patients using cyclosporine versus microemulsion-form cyclosporine: incidence and severity. PMID- 11119890 TI - Conversion from once-daily Sandimmune cyclosporine to once-daily Neoral in renal transplant patients. PMID- 11119891 TI - Cyclosporine trough levels in diabetic and nondiabetic renal transplant patients. PMID- 11119892 TI - Optimization of cyclosporine therapy with abbreviated area under the curve method in renal transplant. PMID- 11119893 TI - Primary tacrolimus immunosuppression in kidney recipients considered "higher immunological risk". PMID- 11119894 TI - Clinical influencing factors for daily dose, trough level, and relative clearance of tacrolimus in renal transplant recipients. PMID- 11119895 TI - Successful conversion from tacrolimus to cyclosporine after kidney transplantation. PMID- 11119896 TI - Rescue therapy with tacrolimus in renal graft patients with cyclosporine A induced hepatotoxicity: a preliminary study. PMID- 11119898 TI - Primary tacrolimus-based immunosuppression in renal allograft recipients: a single center experience. PMID- 11119897 TI - Japanese single-center experience of kidney transplantation under tacrolimus immunosuppression. PMID- 11119899 TI - Tacrolimus vs CyA Neoral in combination with MMF and steroids after cadaveric renal transplantation. PMID- 11119900 TI - Clinical study of FK 506 in renal transplant recipients. PMID- 11119901 TI - Efficacy of tacrolimus in primary kidney transplant patients: multicenter, open label prospective study. PMID- 11119902 TI - Clinical experience with Prograf (tacrolimus, FK 506) in Chinese patients after renal transplantation. PMID- 11119903 TI - ABO-incompatible living donor kidney transplantation under tacrolimus immunosuppression. PMID- 11119904 TI - Conversion from cyclosporine to tacrolimus in renal allograft recipients with delayed graft function from severe acute tubular necrosis. PMID- 11119905 TI - Beneficial effect of multiple drug therapy including tacrolimus in clinical renal transplantation. PMID- 11119906 TI - Malabsorption of tacrolimus in kidney transplant recipients: a Japanese single center experience. PMID- 11119907 TI - Tacrolimus versus cyclosporine as primary prophylactic therapy after cadaveric renal transplant: two-year survival study. PMID- 11119908 TI - Outcome of tacrolimus-treated renal transplantation from elderly donors. PMID- 11119909 TI - Non-heart-beating donor kidney transplantation under tacrolimus immunosuppression. PMID- 11119910 TI - Effect of tacrolimus in renal transplant recipients with immunoglobulin-A nephropathy. PMID- 11119911 TI - Conversion of renal transplant immunosuppression from tacrolimus (FK 506) to cyclosporine: a single center experience. PMID- 11119912 TI - Impact of tacrolimus on hyperlipidemia after renal transplantation: a Japanese single center experience. PMID- 11119913 TI - An early experience with Simulect (basiliximab): an IL-2 receptor antibody. PMID- 11119914 TI - Long-term graft survival of living-related kidneys after donor-specific transfusion. PMID- 11119915 TI - Renal transplantation from non-heart-beating donors with extracorporeal membrane oxygenation: preliminary results. PMID- 11119916 TI - Effect of Deoxyspergualin on the long-term outcome of renal transplantation. PMID- 11119917 TI - Mycophenolate mofetil prevents the progression of chronic kidney allograft nephropathy. PMID- 11119918 TI - MMF-based regimen in maintenance therapy after kidney transplantation. PMID- 11119919 TI - Comparison of the safety and efficacy of mycophenolate mofetil, prednisolone and cyclosporine and conventional cyclosporine and prednisolone in kidney transplantation. PMID- 11119920 TI - Pharmacokinetic study of mycophenolate mofetil in Asian renal transplant recipients. PMID- 11119921 TI - Efficacy and safety of mycophenolate mofetil in different dosages in Asian renal allograft recipients. PMID- 11119922 TI - Safety and efficacy of mycophenolate mofetil for prophylaxis in Asian renal transplant recipients. PMID- 11119923 TI - Acute rejection and the therapeutic choice of drug. PMID- 11119924 TI - Leukocytopheresis therapy for chronic renal allograft rejection. PMID- 11119925 TI - Tacrolimus for rescue therapy in refractory renal allograft rejection. PMID- 11119926 TI - Increasing incidence of steroid resistant rejection in kidney transplantation. PMID- 11119927 TI - Immunity in maintenance immunosuppression with and without cyclosporine in long term kidney transplant recipients. PMID- 11119928 TI - Expression of interleukin-17 as a predictive parameter in acute renal allograft rejection. PMID- 11119929 TI - Immune activation gene expression in CD28 costimulated lymphocytes and its application as a diagnostic tool in kidney transplantation. PMID- 11119930 TI - Oxidative damage in renal transplant patients. PMID- 11119932 TI - Measurement of urine collagen type IV after kidney transplantation. PMID- 11119931 TI - Existence of serum p53 antibodies in cyclosporine A-treated transplant patients: possible detection of p53 protein over-expression. PMID- 11119933 TI - TGF-beta1 is upregulated in chronically deteriorated renal allografts. PMID- 11119934 TI - The significance of apoptotic changes appearing in renal allograft in acute rejection. PMID- 11119935 TI - Soluble FAS in renal allograft recipients. PMID- 11119936 TI - Collagen type IV in acute rejection of kidney. PMID- 11119937 TI - Phenotypic modulation of neointimal smooth muscle cells during the evolution of transplant renal arteriosclerosis determined via myosin heavy chain expression. PMID- 11119938 TI - Serum nitric oxide level as a prognostic parameter for chronic rejection after renal transplantation. PMID- 11119939 TI - Benefits of electron microscopy in the evaluation of renal allograft biopsies. PMID- 11119940 TI - Better clinical outcome in renal transplant recipients with peripheral blood microchimerism. PMID- 11119941 TI - Does implantation biopsy help in predicting renal allograft management and outcome? PMID- 11119942 TI - Examination of serum amyloid A protein in kidney transplant patients. PMID- 11119943 TI - Hepatitis C after renal transplantation. PMID- 11119944 TI - Economic evaluation of therapeutic drug monitoring services in renal transplant recipients treated with cyclosporine. PMID- 11119945 TI - Long-term outcome of living-unrelated donor kidney transplantation. PMID- 11119946 TI - Cadaveric organ donation at University Hospital Kuala Lumpur. PMID- 11119947 TI - Cadaveric renal transplantation at University Hospital Kuala Lumpur: a preliminary report. PMID- 11119948 TI - 30-Year experience of renal transplantation at the Catholic University of Korea. PMID- 11119949 TI - Renal replacement therapies in the elderly: renal transplantation and continuous ambulatory peritoneal dialysis. PMID- 11119950 TI - Quality of life in kidney transplant patients. PMID- 11119951 TI - Kidney transplantation from spousal donors. PMID- 11119952 TI - Clinical analysis of 100 renal transplant recipients back from the People's Republic of China to Taiwan. PMID- 11119953 TI - Clinical experience of pediatric kidney transplantation: what is the benefit? PMID- 11119954 TI - Thirteen-year results of renal transplantation in patients with systemic lupus erythematosus. PMID- 11119955 TI - Outcome of renal transplantation under cyclosporine immunosuppression in patients with lupus nephritis. PMID- 11119956 TI - Long-term outcome of IgA nephropathy in living related kidney transplantation. PMID- 11119957 TI - Both immunologic and nonimmunologic factors are risks for long-term graft survival--a multivariate analysis. PMID- 11119958 TI - Death with function: the next major hurdle. PMID- 11119959 TI - Cadaveric donor kidney transplantation: review of 39 cases. PMID- 11119960 TI - Outcome of renal transplantation for patients with long-term pretransplant dialysis longer than 15 years. PMID- 11119961 TI - Impact of CREG matching on renal allograft survival in one haplotype matched transplants. PMID- 11119962 TI - Kidney transplant using pediatric donors--effect on long-term graft and patient survival. PMID- 11119963 TI - High prevalence of hypertension in rural and urban Indian populations. PMID- 11119964 TI - Kidney transplantation in the recipient with autosomal-dominant polycystic kidney disease: a single center experience. PMID- 11119965 TI - Comparative study of urosurgical complications in renal transplantation: intravesical versus extravesical ureterocystoneostomy. PMID- 11119966 TI - Significance of antidonor blood type antibody for long-term graft acceptance in ABO-incompatible kidney transplantation. PMID- 11119967 TI - Percutaneous fine-needle ethanol injection in a renal transplant patient with enlarged parathyroid glands. PMID- 11119968 TI - A case of kidney transplantation with severe atrophic urinary bladder. PMID- 11119969 TI - Autologous blood transfusion for kidney transplant recipients. PMID- 11119970 TI - Outcome of renal transplantation in patients with renal calculus disease. PMID- 11119971 TI - Abnormal kinetics of erythrocyte Na/Li countertransport in renal transplant patients treated with cyclosporine. PMID- 11119972 TI - Comparison of HLA class I typing by serology with DNA typing in a Chinese population. PMID- 11119973 TI - Subclinical noncompliance in South African black renal allograft recipients. PMID- 11119975 TI - An in toto DNA-database encoded "ID" system: a new concept in primary antigen recognition. PMID- 11119974 TI - Evaluation of socioeconomic factors in noncompliance in renal transplantation. PMID- 11119976 TI - Pregnancy in renal transplant patients. PMID- 11119977 TI - Pregnancy after renal transplantation. PMID- 11119978 TI - Risk factors for avascular bone necrosis after renal transplantation. PMID- 11119979 TI - Pamidronate and calcitriol trial for the prevention of early bone loss after renal transplantation. PMID- 11119980 TI - Prevalence and risk factors for vertebral fractures in renal transplants. PMID- 11119981 TI - Hyperhomocysteinemia in renal transplant recipients with cyclosporine. PMID- 11119982 TI - Ileocolonic complications after kidney transplantation. PMID- 11119983 TI - Hypercalcemia in renal transplant patients with tuberculosis. PMID- 11119984 TI - Clinicopathologic correlation of patients with chronic renal allograft dysfunction. PMID- 11119985 TI - Early graft dysfunction developed within one week after renal allograft. PMID- 11119986 TI - Posttransplant diabetes mellitus in live-related renal transplantation. PMID- 11119987 TI - Clinical outcome following percutaneous transluminal angioplasty for transplant renal artery stenoses. PMID- 11119988 TI - Impact of renal transplantation on hypertension regression in recipients at different ages. PMID- 11119989 TI - Impact of heavy proteinuria (>1 g/d) following renal transplantation. PMID- 11119990 TI - Mycophenolate mofetil-induced ischemic colitis. PMID- 11119992 TI - Mycophenolate mofetil-induced hyperbilirubinemia in renal transplant recipients. PMID- 11119991 TI - Biliary tract complications in renal allograft recipients: two case reports. PMID- 11119993 TI - Primary nonfunctioning grafts in cadaveric kidney transplantation. PMID- 11119994 TI - Effect of donor age and rejection episodes on hypophosphatemia in long-term kidney transplant patients. PMID- 11119995 TI - Clinical study of malignancies after renal transplantation and impact of routine screening for early detection: a single-center experience. PMID- 11119996 TI - Embolization for arteriovenous fistula after graft biopsy in renal transplant recipients: is it essential for all cases? PMID- 11119997 TI - Spontaneous renal allograft rupture in a cohort of renal transplant recipients: a tertiary care experience. PMID- 11119999 TI - Complications associated with using nonabsorbable sutures for ureteroneocystostomy in renal transplant operations. PMID- 11119998 TI - Risk factors of acute tacrolimus nephrotoxicity in renal allografts. PMID- 11120000 TI - Effect of melatonin on renal function in cyclosporine nephrotoxicity. PMID- 11120001 TI - Impact of proteinuria in cyclosporine-treated cadaveric renal transplant recipients. PMID- 11120002 TI - Persistent proteinuria as a prognostic factor for determining long-term graft survival in renal transplant recipients. PMID- 11120003 TI - Drug fever caused by mycophenolate mofetil in a renal transplant recipient--a case report. PMID- 11120004 TI - Ureteritis due to cytomegalovirus infection in renal transplant recipient: a case report. PMID- 11120005 TI - Impact of OKT3 and polyclonal antibodies on symptomatic CMV infection in renal transplant recipients. PMID- 11120006 TI - Diagnosis and treatment of cytomegalovirus infection after renal transplantation. PMID- 11120007 TI - Association between anti-thymocyte globulin administration and cytomegalic virus infection and/or CMV disease in cadaveric renal allograft recipients. PMID- 11120008 TI - Managing hepatitis B reactivation in renal transplant recipients: a 12-year review with emphasis on early detection and early use of lamivudine. PMID- 11120009 TI - Prevalence of abnormal liver function and hepatitis B antigenemia in hepatitis B antibody positive kidney transplant patients. PMID- 11120010 TI - Impact of anti HCV (+) on renal transplantation. PMID- 11120011 TI - Clinical outcome of HCV infection after renal transplantation. PMID- 11120012 TI - Eleven-year experience of kidney transplantation in patients with hepatitis B and C infection. PMID- 11120013 TI - Older recipients who are carriers of hepatitis B and/or C and the outcome of renal transplantation. PMID- 11120015 TI - Long-term effects of hepatitis B and C infection in renal transplantation. PMID- 11120014 TI - Impact of hepatitis C virus infection in renal allograft recipients. PMID- 11120016 TI - Primary varicella virus in adult renal transplant recipients: case reports. PMID- 11120017 TI - Pure red cell aplasia due to parvovirus B19 infection in a renal transplant patient: a case report. PMID- 11120018 TI - Tuberculous abscess of the graft in a renal transplant recipient after chronic rejection: case report. PMID- 11120019 TI - Tuberculosis after renal transplantation. PMID- 11120020 TI - Paradoxical worsening of tuberculosis after anti-TB therapy in a kidney transplant recipient. PMID- 11120021 TI - Candida polyarthritis in a renal transplant patient: case report of a patient successfully treated with amphotericin B. PMID- 11120022 TI - Disseminated nocardiosis with bilateral intraocular involvement in a renal allograft patient. PMID- 11120023 TI - Discontinuation of immunosuppressive antimetabolite for parvovirus B19-associated anemia in kidney transplant patients. PMID- 11120025 TI - Infection in the renal transplant recipient. PMID- 11120024 TI - A pelvic abscess in a renal transplant recipient. PMID- 11120026 TI - Helicobacter pylori infection in renal transplant recipients. PMID- 11120027 TI - Posttransplant malignancy during 30 years at a single center. PMID- 11120028 TI - Malignancy in renal transplant recipients. PMID- 11120029 TI - A case study of adult T-cell lymphoma in a kidney transplant patient. PMID- 11120031 TI - Comparative study of renal carcinomas in renal transplant, dialysis, and general (nongrafted and nonuremic) patients. PMID- 11120030 TI - Clinicopathological study of colorectal cancers after renal transplantation. PMID- 11120032 TI - Successful treatment of Epstein-Barr virus-associated T-cell cutaneous lymphoma in a renal allograft recipient: case report and review of the literature. PMID- 11120033 TI - Spectrum of malignancies in Asian renal allograft recipients. PMID- 11120034 TI - Inhibition of apoptosis in anti-CD3-treated peripheral blood lymphocytes by immunosuppressive drugs. PMID- 11120035 TI - Correlation between proliferating cell nuclear antigen expression and phenotypic change in smooth muscle cells during the development of vasculopathy in heterotopically transplanted rat hearts. PMID- 11120036 TI - Study of the relationship between nonmuscle myosin heavy chain B (SMemb) and acute rejection of rat renal transplantation models. PMID- 11120037 TI - An ex vivo model to study the intestinal biotransformation of immunosuppressive drugs. PMID- 11120038 TI - Blockade of CD4 is more tolerogenic than blockade of Ag presentation via self MHC class II antigens. PMID- 11120039 TI - High dose DBMC associated tolerance in live-related renal allograft recipients. PMID- 11120040 TI - Adenovirus-mediated viral IL-10 gene transfer prolongs survival of allogeneic spheroidal aggregate-cultured hepatocytes. PMID- 11120041 TI - Adenovirus-mediated interleukin-10 overexpression: comparison between intraportal and intramuscular gene transfer. PMID- 11120042 TI - Total antioxidant status and antioxidant enzymes in cyclosporine treated rats. PMID- 11120043 TI - The beneficial effect of melatonin for cyclosporine hepatotoxicity in rats. PMID- 11120044 TI - Combined effect of donor-specific bone marrow transplantation via portal vein and FK506 on small bowel transplantation in the rat. PMID- 11120045 TI - Immunosuppressive effects of Tripterygium wilfordii polysaccharide on LPS stimulated human monocytes. PMID- 11120046 TI - Effects of tacrolimus on rat thymic epithelial cells. PMID- 11120047 TI - Intrathymic inoculation of donor B cells induced indefinite prolongation of fully allogeneic cardiac grafts but not islet or skin grafts. PMID- 11120048 TI - Effect of the immunosuppressants FK506 and D-allose on allogenic orthotopic liver transplantation in rats. PMID- 11120049 TI - Perioperative administration of FTY720 and CTLA4IG in rat heart transplantation. PMID- 11120050 TI - Effect of cyclosporine, mycophenolic acid, and rapamycin on the proliferation of rat aortic vascular smooth muscle cells: in vitro study. PMID- 11120051 TI - CTLA-4IG treatment induces long-term acceptance of rat small bowel allografts. PMID- 11120052 TI - Inducible expression of BTEB2, a member of the zinc-finger family of transcription factors, in cardiac allograft arteriosclerosis. PMID- 11120053 TI - Survival of fully allogeneic cardiac allografts induced by delivery of alloantigen via respiratory mucosa is abrogated by blockade of the B7 pathway. PMID- 11120054 TI - Oral delivery of alloantigen combined with non-depleting anti-CD4 monoclonal antibody induces indefinite survival of cardiac allografts and generates CD4(+) regulatory T cells. PMID- 11120055 TI - Adenoviral vectors containing the CTLA4IG-gene inhibit chronic rejection in heterotopically transplanted rat tracheas. PMID- 11120056 TI - Inhibition of CD95 ligand-mediated inflammation. PMID- 11120057 TI - The treatment of chronic rejection with mycophenolate mofetil versus azathioprine in kidney transplantation. PMID- 11120058 TI - Donor cell chimerism, host lymphocyte phenotype and allograft acceptance. PMID- 11120059 TI - Microchimerism and allograft acceptance: is it caused by microchimerism or graft antigens? PMID- 11120060 TI - Acquired tolerance is induced by MHC-incompatible test skin grafts in adult thymectomized rats with chronic drainage of thoracic duct. PMID- 11120061 TI - Induction of long-term heart graft survival by intrathymic injection of HLA class I peptide: a study with HLA class I transgenic mice model. PMID- 11120062 TI - A novel immunosuppressive agent, SQDG, derived from sea urchin. PMID- 11120063 TI - Synergistic effect of Tripterygium wilfordii Hook.F (TWHF) and cyclosporin A in rat liver transplantation. PMID- 11120064 TI - Differential suppression by Tripterygium wilfordii extracts (traditional Chinese medicine) in the allogeneic rat mixed lymphocyte reaction. PMID- 11120065 TI - Prope tolerance: a step in the search for tolerance in the clinic. PMID- 11120066 TI - Immunosuppressive effect of bactobolamine on in vitro lymphocyte alloactivation: synergism with cyclosporine. PMID- 11120067 TI - Unexpected strong immunosuppressive effect of X-ray irradiation on acute rejection of rat aortic allografts. PMID- 11120068 TI - Prolonged ischemia potentiates apoptosis formation during reperfusion by increase of caspase 3 activity and free radical generation. PMID- 11120069 TI - The thymus is essential for the development of operational tolerance induced by donor-specific transfusion plus anti-CD4 monoclonal antibody. PMID- 11120070 TI - Granulocyte colony-stimulating factor-mobilized donor monocytes facilitate heart allograft acceptance. PMID- 11120071 TI - Increase of hepatocyte growth factor after renal transplantation. PMID- 11120072 TI - Hepatocyte growth factor: a sensitive indicator for the acute rejection of renal transplants. PMID- 11120073 TI - Anti-alpha1,3-galactose-mediated hyperacute rejection of vascularized transplants in a small animal model. PMID- 11120074 TI - Dendritic cell progenitors prolong allograft survival through T-helper 2 deviation of the Th1/Th2 paradigm. PMID- 11120075 TI - Cytokine requirement for the development of rat dendritic cells by in vitro culturing of bone marrow cells. PMID- 11120076 TI - Role of natural killer cells in the rejection of transplanted hearts in the mouse model. PMID- 11120077 TI - Effects of glucocorticoids on rat thymus and apoptosis. PMID- 11120078 TI - Mice treated with anti-CD4 monoclonal antibody accept fully allogeneic thyroid grafts but reject second-donor-type thyroid grafts in maintenance phase. PMID- 11120079 TI - The regulation of natural killer-mediated swine endothelial cell lysis by HLA-G (G1 and G3). PMID- 11120080 TI - Role of natural killer cells in allograft rejection. PMID- 11120081 TI - Monocyte inhibitors as a potent immunosuppressant in rodent heart transplantation. PMID- 11120082 TI - Standard liver volume in adults. PMID- 11120083 TI - Marital adjustment after interspouse living donor liver transplantation. PMID- 11120085 TI - Gadolinium-enhanced magnetic resonance portography: application in pediatric liver transplant recipients. PMID- 11120084 TI - Intraoperative Doppler ultrasound in living-related liver transplantation. PMID- 11120086 TI - Donor fatty (steatotic) liver allografts in orthotopic liver transplantation: a revisit. PMID- 11120087 TI - Is donor obesity related to liver steatosis and liver graft dysfunction in liver transplantation? PMID- 11120088 TI - Strategy for ABO-incompatible living-related liver transplantation. PMID- 11120089 TI - Influence of HLA compatibility on living-related liver transplantation. PMID- 11120090 TI - Impact of HLA matching in living-related liver transplantation. PMID- 11120091 TI - Management of postoperative analgesia in living liver donors. PMID- 11120092 TI - Indications and efficacy of apheresis for liver transplant recipients: experience of 16 cases in 34 living-related liver transplants. PMID- 11120093 TI - Assessment of graft viability using hyaluronic acid and adenosine triphosphate in orthotopic liver transplantation from non-heart-beating donors. PMID- 11120094 TI - Postreperfusion syndrome in orthotopic liver transplantation. PMID- 11120095 TI - Use and limitations of reconditioning ischemically damaged livers from non-heart beating donors by venous oxygen persufflation. PMID- 11120096 TI - Successful conversion from cyclosporine to tacrolimus for immunosuppression. PMID- 11120097 TI - Investigation of rhIL-10 inhibition of acute rejection after liver transplantation. PMID- 11120098 TI - A prospective study comparing safety and efficacy of mycophenolate mofetil versus azathioprine in primary liver transplant recipients. PMID- 11120099 TI - Lamivudine improves the prognosis of patients with hepatitis B after liver transplantation. PMID- 11120101 TI - Liver failure following living donor liver transplantation for fulminant hepatic failure. PMID- 11120100 TI - Role of recipient CD56(+) CD3(+) cells in the graft in living-related partial liver transplantation. PMID- 11120102 TI - Strategy for chronic rejection in recipient of living-related liver transplantation. PMID- 11120104 TI - Ten years of experience of liver transplantation in Singapore. PMID- 11120103 TI - Living donor liver transplantation: the Kaohsiung experience. PMID- 11120105 TI - First successful in situ split-liver transplantation in Korea. PMID- 11120106 TI - Liver transplantation in Hong Kong--a wider application. PMID- 11120107 TI - Liver transplantation is not contraindicated in infancy. PMID- 11120108 TI - Changes in health-related quality of life and working competence before and after liver transplantation. PMID- 11120109 TI - Health status survey of adult patients undergoing living-related liver transplantation. PMID- 11120110 TI - Family perspectives of the factors facilitating Taiwanese pediatric recipients' recovery from liver transplantation: one year follow-up. PMID- 11120111 TI - Problems in living donor liver transplantation in adults: postoperative management, complications, and costs. PMID- 11120112 TI - Initial experience with right lobe living donor liver transplantation. PMID- 11120113 TI - Living-related liver transplantation in adults compared with children. PMID- 11120114 TI - Liver transplantation in Chinese patients with chronic hepatitis B. PMID- 11120115 TI - Influence of donor age on the graft function after living-related liver transplantation. PMID- 11120116 TI - Correlation between graft size and necessary tacrolimus dose after living-related liver transplantation. PMID- 11120117 TI - Long-term follow-up (more than 5 years) of patients after living-related liver transplantation. PMID- 11120118 TI - Liver transplantation for hepatocellular carcinoma. PMID- 11120119 TI - Liver transplantation in hepatocellular carcinoma. PMID- 11120121 TI - Living-related liver transplantation in patients with pulmonary vascular disease. PMID- 11120120 TI - Liver transplantation in a patient with diffuse portal venous system thrombosis. PMID- 11120122 TI - Liver transplantation in a patient with pulmonary hypertension. PMID- 11120123 TI - Use of liver graft infested with Clonorchis sinensis for living related liver transplantation: a case report. PMID- 11120124 TI - Living related partial liver transplantation for hyperammonemia due to congenital absence of the portal vein. PMID- 11120125 TI - Living related liver transplantation in two Byler disease families. PMID- 11120126 TI - Pretransplant evaluation of bone mineral density in adult patients with end-stage cholestatic liver disease. PMID- 11120127 TI - Adult-to-adult living-related liver transplantation for hepatitis B-related cirrhosis in Japan: two case reports. PMID- 11120128 TI - Factors influencing persistent hyperbilirubinemia following adult-to-adult living related liver transplantation. PMID- 11120129 TI - Serum levels of amino acids in candidates for living-related liver transplantation. PMID- 11120130 TI - Interventional radiology in the treatment of post-liver transplant complications. PMID- 11120131 TI - Ceruloplasmin, a novel candidate as a diagnostic marker for liver function after liver transplantation. PMID- 11120132 TI - Changes of serum cytokines associated with hepatic regeneration after living related liver transplantation. PMID- 11120133 TI - Liver transplantation in patients with transjugular intrahepatic portosystemic shunts. PMID- 11120134 TI - Orthotopic liver transplantation for treatment of patients with Budd-Chiari syndrome: a Singe-center experience. PMID- 11120135 TI - Living-related liver transplantation for patients with primary biliary cirrhosis. PMID- 11120136 TI - Growth of pediatric patients with biliary atresia after liver transplantation: influence of age at transplantation and steroid administration. PMID- 11120137 TI - Persistent pleural and peritoneal fluid discharge after adult-to-adult living related liver transplantation. PMID- 11120138 TI - Subclinical central pontine myelinolysis after liver transplantation. PMID- 11120139 TI - Giant intrahepatic hematoma after liver biopsy in a liver transplant recipient. PMID- 11120140 TI - Primary graft dysfunction after liver transplantation. PMID- 11120141 TI - Technique for repair of lymphocele after liver transplantation using patent blue dye. PMID- 11120142 TI - Isolated caudate lobe infarct after orthoptic liver transplantation. PMID- 11120143 TI - A case of hyperkalemic distal renal tubular acidosis secondary to tacrolimus in living donor liver transplantation. PMID- 11120144 TI - Impaired renal function after liver transplantation: role of rhabdomyoglobinuria. PMID- 11120146 TI - Fungal infection in living related liver transplantation patients. PMID- 11120145 TI - Impact of primary immunosuppression on the incidence of infectious complications after orthotopic liver transplantation. PMID- 11120147 TI - Post liver transplantation stenosis of biliary-enteric anastomoses in infancy: diagnosis and treatment. PMID- 11120148 TI - Treatment for strictures of hepatojejunostomy in living-related liver transplantation. PMID- 11120149 TI - Coagulopathy after bilirubin adsorption in a living-related partial liver transplant recipient. PMID- 11120150 TI - Porcine model of fulminant hepatic failure treated by liver transplantation. PMID- 11120151 TI - Liver transplantation for patients with hepatitis B: prevention of hepatitis B recurrence by intravenous antihepatitis B immunoglobulin and lamivudine. PMID- 11120152 TI - Prophylaxis against hepatitis B recurrence following liver transplantation in HBs Ag(+) patients. PMID- 11120153 TI - Arterial reconstruction using vein graft from the common iliac artery after hepatic artery thrombosis in living-related liver transplantation. PMID- 11120154 TI - Asymptomatic vascular complications in liver transplantation. PMID- 11120155 TI - Long-term follow up of heterotopic liver allograft survival with or without hepatic arterial reconstruction. PMID- 11120156 TI - Pseudoobstruction of the portal vein in living-related liver transplantation: a case report. PMID- 11120157 TI - Three cases of posttransplant lymphoproliferative disorder in recipients of liver transplantation. PMID- 11120158 TI - Metastatic hepatocellular carcinoma to bone in a liver transplant patient four years after liver transplantation: report of a case. PMID- 11120159 TI - Rupture of newly developed esophageal varices after adult-to-adult living-related liver transplantation. PMID- 11120160 TI - A case of esophageal variceal rupture following acute portal vein thrombosis three days after living-related liver transplantation. PMID- 11120161 TI - More effective immunosuppression with the use of FK506 after liver transplantation. PMID- 11120162 TI - Cryopreservation of primary porcine hepatocytes for use in bioartificial liver support systems. PMID- 11120163 TI - Soluble thrombomodulin antigen as a marker for endothelial damage during liver transplantation. PMID- 11120164 TI - Different changes of endothelin-1 after reperfusion in a warm ischemia/reperfusion and transplantation model in pig liver. PMID- 11120165 TI - Evaluation of angiogenesis by intrahepatic omental implantation in rat liver transplantation. PMID- 11120166 TI - Changes in hepatic microdialysate uric acid levels in porcine liver ischemia/reperfusion. PMID- 11120167 TI - Role of hepatocyte growth factor in hepatic ischemia and reperfusion injury. PMID- 11120168 TI - The role of hepatocyte transplantation to treat chronic liver failure. PMID- 11120169 TI - Analysis of the influence of portal venous pressure (shear stress) to the movement of mononuclear cells in the liver. PMID- 11120170 TI - Effects of ulinastatin on hypotension and hepatic circulation in brain dead rabbits. PMID- 11120171 TI - Effects of hypotension on hepatic circulation and function: comparison of brain death and exsanguination models. PMID- 11120172 TI - Evidence of regulatory T lymphocytes that constitute peripheral blood microchimerism following rat liver transplantation. PMID- 11120173 TI - Intrahepatic leukocytes in auxiliary partial orthotopic liver transplantation in beagle dogs. PMID- 11120174 TI - Downregulation of cytokine-induced neutrophil chemoattractants and reduction of reperfusion injury in liver allograft by interleukin-10. PMID- 11120175 TI - Protective effects of herbimycin A on hepatic reperfusion injury. PMID- 11120176 TI - Nitric oxide donor improved the impaired endothelial-dependent relaxation of canine hepatic artery after preservation with UW solution. PMID- 11120177 TI - Splenectomy and auxiliary liver transplantation. PMID- 11120178 TI - Protective effect of ischemic preconditioning on hepatic ischemia-reperfusion injury in mice. PMID- 11120179 TI - FR128998 ameliorates liver injury after ischemia-reperfusion with extended liver resection in dogs. PMID- 11120180 TI - The expression of IL-8 mRNA and infiltration of neutrophils with FR167653 administration on extended liver resection with ischemia in dogs. PMID- 11120181 TI - Protective effect of an antineutrophil antibody, Urge-8, on liver ischemia reperfusion injury in a new hepatic ischemia model. PMID- 11120182 TI - Mechanisms of immunosuppressive effects of FK 506 in light of apoptosis of hepatocytes and infiltrating lymphocytes in rat allografted livers. PMID- 11120183 TI - The effect of cyclooxygenase 2 inhibitor (FK3311) on ischemia-reperfusion injury with hepatectomy in dogs. PMID- 11120184 TI - Hepatic regeneration and ischemia/reperfusion injury in fatty-liver rats. PMID- 11120185 TI - Cold preservation of rat cultured hepatocytes: the scoparone effect. PMID- 11120186 TI - Clonal colony formation of hepatic stem/progenitor cells enhanced by embryonic fibroblast conditioning medium. PMID- 11120187 TI - Reversible immortalization of adult human hepatocytes(1). PMID- 11120188 TI - Expression of clusterin in a rat tolerogenic OLT model. PMID- 11120189 TI - Investigation of liver regeneration after rat liver transplantation. PMID- 11120190 TI - Intraoperative fluid requirements during porcine liver transplantation. PMID- 11120191 TI - Effect of leukotriene B(4) receptor antagonist (ONO4057) on hepatic allografting in rats. PMID- 11120192 TI - Stable technique for reconstruction of hepatic artery in hamster-to-rat liver transplantation. PMID- 11120193 TI - Effect of bioartificial liver in pigs with total ischemic liver failure. PMID- 11120194 TI - Assessment of viability of the liver graft in different cardiac arrest models. PMID- 11120195 TI - Main injury site of liver grafts from non-heart-beating donors in pigs. PMID- 11120196 TI - Development of bioartificial liver system using a fluidized-bed bioreactor. PMID- 11120197 TI - Immunohistochemical study of the regeneration process of extrinsic hepatic nerves following liver transplantation in rats. PMID- 11120198 TI - Development of a bioartificial liver with glutamine synthetase-transduced recombinant human hepatoblastoma cell line, HepG2. PMID- 11120199 TI - Staining of collagen type IV in liver allografts. PMID- 11120200 TI - Predictive value of CD45RC mRNA for acute rejection in hepatic allografts in rats. PMID- 11120201 TI - Donor bone marrow perioperatively administered via portal vein induced prolongation of skin allograft survival and microchimerism in liver-transplanted rats. PMID- 11120202 TI - Intrasplenic transplantation of immortalized human fetal hepatocytes prolongs the survival of 90% hepatectomized rats. PMID- 11120203 TI - Establishment of a highly differentiated immortalized adult human hepatocyte cell line by retroviral gene transfer. PMID- 11120204 TI - Proliferative and functional ability of transplanted murine neonatal hepatocytes in adult livers. PMID- 11120205 TI - Effects of combined growth factors on clonal growth and albumin secretion of murine fetal hepatocytes in low density culture. PMID- 11120206 TI - Adenovirus-mediated gene transfer of DAF and CD59 in xenogeneic pig liver perfusion. PMID- 11120207 TI - Activation of telomerase by liver transplantation in rats. PMID- 11120208 TI - Insulin gene transfer improves posthepatectomized status of diabetic rats. PMID- 11120209 TI - Rabbit antithymocyte globulin induction immunosuppression in heart transplantation. PMID- 11120210 TI - Assessment of autonomic reinnervation of cardiac grafts by analysis of heart rate variability. PMID- 11120211 TI - ABO blood types and the chance to undergo heart transplantation. PMID- 11120212 TI - Extracorporeal membrane oxygenation rescue after heart transplantation. PMID- 11120213 TI - Infection in heart transplant recipients: seven years' experience at the National Taiwan University Hospital. PMID- 11120214 TI - Is bicaval anastomosis superior to standard atrial procedure of heart transplantation? PMID- 11120215 TI - Effects of cryopreservation of aortic xenografts on graft patency and immunogenicity. PMID- 11120216 TI - Human serum induces apoptosis of isolated xenogeneic cardiomyocytes in vitro. PMID- 11120217 TI - Adenine nucleotide changes and reperfusion injury in non-heart-beating donor lungs. PMID- 11120218 TI - Cryopreserved aortic grafting in the presence of peritonitis. PMID- 11120219 TI - FK409 enhances posttransplant cardiac function following 12-hour cold preservation. PMID- 11120221 TI - The effects of FR167653 on long-term heart preservation in dogs. PMID- 11120220 TI - Effects of Celsior solution on long-term preservation of canine hearts with a new portable hypothermic perfusion apparatus: a preliminary study. PMID- 11120222 TI - In vivo gene transfer into rat hearts with Epstein-Barr virus-based episomal vectors using a gene gun. PMID- 11120223 TI - Effect of coronary perfusion with an oxygenated Celsior solution on 24-hour preservation in canine hearts. PMID- 11120224 TI - Possibility of xenotransplantation with a cryopreserved porcine heart valve in a canine model. PMID- 11120225 TI - Successful living-donor lobar lung transplantation for an end-stage bronchiectasis patient. PMID- 11120226 TI - Intermediate results in Sauropus androgynus bronchiolitis obliterans patients after single-lung transplantation. PMID- 11120228 TI - Effects of FK409 on pulmonary ischemia-reperfusion injury in rats. PMID- 11120227 TI - Energy metabolism and reperfusion injury in warm and cold ischemia of inflated and deflated lungs. PMID- 11120229 TI - Spontaneous nitric oxide (FK409) ameliorates pulmonary ischemia-reperfusion injury in dogs. PMID- 11120230 TI - Effect of FK3311 on ischemia-reperfusion injury in canine pulmonary models. PMID- 11120231 TI - Retransplantation of contralateral lung in a patient with Sauropus androgynus induced bronchobronchiolitis obliterans. PMID- 11120232 TI - Effects of FR167653 on ischemia-reperfusion injury in canine lung transplantation models. PMID- 11120233 TI - Estimation of graft blood flow in rat lung transplantation using near-infrared spectroscopy. PMID- 11120234 TI - Ultrastructural evaluation of preservation and reperfusion effects of low potassium dextran glucose solution in canine allograft lungs. PMID- 11120235 TI - Photodynamic effects of gallium-metal porphyrin on human leukemia cells in combination with high-brightness LED light: application for autologous bone marrow transplantation. PMID- 11120236 TI - Development of a novel LED apparatus for photodynamic purging of leukemia cells in hematopoietic stem-cell transplantation. PMID- 11120237 TI - NK cells expressing killer cell inhibitory receptors after allogeneic bone marrow transplantation. PMID- 11120238 TI - FK 506 inhibits severe graft-versus-host disease without mediating cytokine balance and/or cytotoxic molecules. PMID- 11120239 TI - Effect of granulocyte colony-stimulating factor administered before bone marrow transplantation on hematopoietic recovery. PMID- 11120240 TI - Increased adhesion molecule expression during graft-versus-host disease. PMID- 11120241 TI - NK/Cytotoxic T cells: major effector cells in GVHD after umbilical cord blood allotransplantation. PMID- 11120242 TI - Depletion of activated alloreactive T cells in prevention of acute graft-versus host disease. PMID- 11120243 TI - GM-CSF-independent development of dendritic cells from bone marrow cells in the GM-CSF-receptor-deficient mouse. PMID- 11120244 TI - Rapid recovery of hematopoiesis in patients given high-dose chemotherapy followed by infusion of refrigerated unprocessed autologous bone marrow and peripheral blood. PMID- 11120245 TI - Inhibition of NK cell activity induces improvement and stable chimerism after allogeneic transplantation. PMID- 11120247 TI - Four-year results of pancreas transplantation in Taiwan. PMID- 11120246 TI - High-dose chemotherapy and autologous stem cell rescue for acute myeloid leukemia remains a safe, effective, and valid option. PMID- 11120248 TI - Experience with combined pancreatic-renal transplantation using extraperitoneal placement. PMID- 11120249 TI - Comparison of pancreas transplantation outcome between the cyclosporine and tacrolimus eras. PMID- 11120250 TI - Effect of subcutaneous pancreatic tissue transplants on host pancreatic tissue levels of insulin and glucagon in the diabetic rat. PMID- 11120251 TI - Pancreatic islet cell autotransplantation in the canine model. PMID- 11120252 TI - Development of a new bioartificial pancreas possessing angiogenesis-inducing function. PMID- 11120253 TI - Sinusoidal capillaries revascularize pancreatic tissue grafts within 24 hours of transplantation. PMID- 11120254 TI - Morphologic changes in pancreatic tissue transplants in rats. PMID- 11120255 TI - FK 506 significantly improves transferred insulin gene expression in total pancreatectomized dogs. PMID- 11120256 TI - Evidence for radiosensitive regulatory T cells that constitute peripheral microchimerism after pancreaticoduodenal transplantation. PMID- 11120257 TI - Quantitative aspects of microchimerism after rat small bowel and pancreaticoduodenal transplantation. PMID- 11120258 TI - Can donor-specific transfusion via the portal vein under activated Kupffer cells enhance the beneficial effect of portal venous tolerance for rat small intestinal transplantation? PMID- 11120259 TI - Effects of insulin-like growth factor-1 on massive resection of small intestine with or without ileocecal resection in rats. PMID- 11120260 TI - Technical refinements for successful small bowel transplantation in the pig. PMID- 11120261 TI - Jejunum is preferable to ileum for transplantation when split small bowel transplantation from a single donor is performed. PMID- 11120262 TI - Combination therapy with FK 506 and RS61443 for rejection following allogeneic small bowel transplantation in rats. PMID- 11120264 TI - Pregnancy site-specific tolerance to allogeneic fetus. PMID- 11120263 TI - Effect of neovascularization-inducing bioartificial pancreas on survival of syngeneic islet grafts. PMID- 11120265 TI - Effects of four extracellular matrices associated with growth factors on clonal culture and proliferation of murine fetal hepatocytes. PMID- 11120266 TI - Human cytokine production against cattle. PMID- 11120267 TI - Characterization of bovine cells for xenotransplantation. PMID- 11120268 TI - The effective antigen presentation of human MHC on the lymphocytes of HLA DPW0401 transgenic pigs: examination with xenogenic mixed lymphocyte culture and primed lymphocyte tests. PMID- 11120269 TI - Production of transgenic pigs expressing human DAF (CD55) regulated by the porcine MCP gene promoter. PMID- 11120270 TI - An attempt to downregulate the Hanganutziu-Deicher antigen by overexpression of glycosyltransferases. PMID- 11120272 TI - Adenovirus-mediated triple gene transfer of human complement regulating proteins to the porcine endothelial cell. PMID- 11120271 TI - Effects of Gal beta 1,4 GlcNAc alpha 2,6-D-Sialyl transferase on swine xenoantigen. PMID- 11120273 TI - High efficacy of gene transfer and expression using adenovirus vector in the rat liver transplantation model. PMID- 11120274 TI - A new Cre recombinase gene based on optimal codon usage in mammals: a powerful material for organ-specific gene targeting. PMID- 11120275 TI - Carbohydrate inhibition of human serum induced porcine endothelial cell apoptosis. PMID- 11120276 TI - Effectiveness of adenovirus-mediated gene transfer with CTLA4Ig in allo and xeno transplantation. PMID- 11120277 TI - Plant lectin binding specificity to carbohydrates on porcine endothelial cells. PMID- 11120278 TI - The effect of deoxyspergualin on rat thymocytes and thymic epithelial cells. PMID- 11120279 TI - Reaching out to Asia for living kidney donors. PMID- 11120280 TI - The relationship between cytokines in MLC supernatants and acute rejection after renal transplantation. PMID- 11120281 TI - Primary tacrolimus immunosuppression in kidney recipients considered "higher immunologic risk". PMID- 11120282 TI - An ex vivo model to study the intestinal biotransformation of immunosuppressives. PMID- 11120283 TI - Intramucosal pH and serum endotoxin concentrations as early predictive parameters for primary nonfunction after experimental liver transplantation. PMID- 11120284 TI - Interferon Alfa-2A and ribavirin therapy for hepatitis C recurrence after liver transplantation. PMID- 11120285 TI - Replacement of corticosteroids by mycophenolate mofetil in liver graft recipients on initial tacrolimus immunosuppression. PMID- 11120286 TI - Donor bone marrow cell infusion is effective in inducing tolerance in dogs treated with fractionated lymphoid irradiation and FK506. PMID- 11120288 TI - Facing 21st century demographics: management of the surgical patient. PMID- 11120287 TI - Allogeneic hematopoietic stem cell transplantation: from the nuclear age into the twenty-first century. PMID- 11120289 TI - Cystic tumors of the pancreas: an update. PMID- 11120290 TI - Is there still a place for axillary dissection in breast cancer? PMID- 11120291 TI - Prevention of colonic neoplasms? Is it for real or just a hoax? PMID- 11120292 TI - What's new about CEA? PMID- 11120293 TI - Changing surgical approaches to patients with primary hyperparathyroidism. PMID- 11120294 TI - Conservative therapy of hyperparathyroidism? PMID- 11120295 TI - Another call from anesthesia complaining about a low potassium: are they right to be concerned? PMID- 11120296 TI - Clinical pathways. PMID- 11120297 TI - Geriatric surgery. PMID- 11120298 TI - Follicular thyroid cancer: an update. PMID- 11120300 TI - Radiation enteritis: sometimes the treatment is worse than the disease. PMID- 11120299 TI - Klatskin's tumor: is there new hope? PMID- 11120301 TI - Carotid endarterectomy preoperative imaging: is duplex enough? PMID- 11120302 TI - Current status of carotid angioplasty and stenting in the treatment of carotid artery stenosis. PMID- 11120303 TI - Internet and e-mail security: for sale, your vital statistics (part 1). PMID- 11120304 TI - Current management of perforated peptic ulcer. PMID- 11120305 TI - Fragment wound pseudoaneurysm presenting 54 years after injury(1). AB - A 74-year-old man presented with acute arterial insufficiency of the left leg. Surgical history was remarkable for a mortar fragment injury to his proximal medial left thigh in 1945. His wounds healed secondarily after operative debridement. He denied any knowledge of a vascular repair. He was asymptomatic until his acute presentation with a threatened limb.Physical examination demonstrated an acutely ischemic extremity with moderately impaired neurologic function and a pulsatile mass over the proximal superficial femoral artery (SFA). Arteriography demonstrated an SFA pseudoaneurysm in proximity to multiple mortar fragments with intraluminal clot and distal arterial obstruction. The extremity was successfully revascularized with embolectomy of the distal arterial thrombus and repair of the arterial injury and pseudoaneurysm with reversed contralateral saphenous vein.This case represents the longest reported delayed presentation of a traumatic false aneurysm. It emphasizes that proper assessment of underlying potential vascular injury must be performed if delayed complications are to be avoided. Comparisons between the unique challenges presented by combat associated injury versus those seen in today's civilian environment are emphasized. PMID- 11120306 TI - Patterns of appendicitis at a forward-deployed United States Army Hospital: the Korea experience(2)(2). AB - Appendicitis is both a common surgical condition in a young military active duty population and a diagnosis that is notoriously easy to miss. We reviewed all cases of appendicitis that presented to the 121st General Hospital, which is an army facility in Seoul, South Korea, during an 18-month period. Our patient population consisted of 37,000 soldiers as well as dependants and other patients eligible for care. Of 79 patients operated on for suspected acute appendicitis, 60 (76%) had either acute suppurative or perforated appendicitis. The appendix was normal in 16 patients (20%), whereas 3 patients (4%) had a normal appendix with other surgical pathology. We found a perforation rate of 24%, and of these patients, 53% had prior visits to a health care provider in which an incorrect diagnosis or no treatment was given. Only 3 patients with simple acute appendicitis had been previously seen (7%). Patients were transported to the hospital in different ways, but we were unable to find any correlation among modes of transport to the hospital and perforation rates. We believe that timely diagnosis and referral for appendicitis could be improved through focused education of all primary care providers on this disorder. PMID- 11120307 TI - The incidence of intraductal papillary mucinous tumors of the pancreas(1). AB - To review a population-controlled single institution experience with intraductal papillary mucinous tumors (IPMTs) of the pancreas treated in the United States and generate an incidence of this recently described disease process.First decribed in 1982, mucin-secreting pancreatic cancer constituted a newly recognized category of pancreatic exocrine tumors distinct from mucinous cystic neoplasms. Since that time, several small series of IPMTs of the pancreas have been reported. Most studies come from Asia and Europe, with limited data from American institutions and no description of the incidence.The authors retrospectively reviewed all patients who underwent pancreatic resections at Madigan Army Medical Center from October 1992 through November 1999. Cases suggestive of IPMTs underwent re-examination by a staff pathologist. Clinical presentation, imaging studies, treatment, histopathology, and outcomes were reviewed for those with IPMTs. The data base of all patients eligible for care was evaluated to obtain a population denominator and determine an incidence for this neoplasm.Over 7 years, 78 patients underwent pancreatic resections. Forty two had pancreatic neoplasms and, of those, 8 (19%) had IPMTs. The mean age was 67 years, with equal numbers of males and females. All patients were symptomatic (abdominal pain, 75%; jaundice, 25%; weight loss, 25%). Abnormal computed tomography scans were noted in 7/8 (88%) cases. Mucin was visualized during endoscopic retrograde cholangiopancreatography in 5/7 (71%) patients. Preoperative diagnosis of IPMT was made in 5 (62%) cases. The locations of the tumors were head (63%), tail (12%), head and body (12%), and body and tail (12%). All lesions were resectable, and procedures included 5 pancreatoduodenectomies, 2 distal pancreatectomies, and 1 total pancreatectomy. Main duct tumors were noted in 63%, whereas the remainder had both main and branch duct lesions. Tumor invasion was discovered in 2 (25%) cases. Mean follow-up was 29 months. Those without invasion were all alive (follow-up, 6 to 86 months). One patient with tumor invasion died 4 months after surgery and the other was living 20 months later. The incidence of IPMTs was 1 case/281,000 patients/year.The incidence of IPMTs of the pancreas may be higher than previously recognized. Aggressive resection is warranted based on the favorable prognosis of patients without tumor invasion. PMID- 11120309 TI - Rectal bleeding 2 years after excision of a cutaneous melanoma. PMID- 11120308 TI - Vascular trauma at a military level II trauma center(1). AB - As members of an American College of Surgeons Committee on Trauma-designated level II trauma center, we decided to review our experience with vascular trauma. In addition, we sought to characterize the vascular injuries presented and to compare our outcomes to the general trauma population.A review of all vascular trauma admissions from January 1997 through January 2000 was performed. The William Beaumont Army Medical Center (WBAMC) trauma registry data base was searched for vascular injuries utilizing 3 different search criteria: organ system, operation/procedure, and ICD-9 codes. Injuries were then characterized by age, gender, site of injury, injury severity score (ISS), mechanism, and need for surgery. Mortality rates were computed for both vascular and nonvascular trauma populations. Statistical analysis of the data was determined by Student t test and z score.Between January 1997 and January 2000, there were 1398 patients admitted to the trauma service at WBAMC. Of these, 48 patients (3.4%) had vascular injuries. The mean ISS for all nonvascular traumas was 8.4 +/- 8.9. The mean ISS for those with vascular injuries was 17.9 +/- 12.6 (p < 0.001). Blunt trauma accounted for 90% of all nonvascular admissions. Penetrating trauma accounted for 10% of all nonvascular admissions. In the vascular trauma population, blunt trauma accounted for 56% and penetrating trauma accounted for 39%. Five percent of the vascular injuries identified were iatrogenic. Surgical intervention was required in 85.4% and 44.2% of the vascular and nonvascular trauma populations, respectively. The mortality rate for nonvascular admissions was 4.8% (65/1350). Those with vascular injuries had a mortality rate of 20.8% (10/48). For trauma patients requiring an operation, the mortality rate was 4.5% (27/597). For patients with vascular injuries who required an operation, the mortality rate was 25.7% (9/35) (p = 0.007).Vascular trauma represents a small percentage of all trauma admissions. These patients have a higher ISS on admission and more of them require surgical intervention. The operative and overall mortality rates are higher in patients with vascular injuries than in the general trauma population. PMID- 11120310 TI - Combined intraoperative alcohol celiac ablation and lateral pancreaticojejunostomy for chronic pancreatitis. PMID- 11120311 TI - Effect of bead swelling on the durability of polylysine alginate microcapsules. PMID- 11120312 TI - Induction of acquired tolerance by adoptive transfer of in vivo allopeptide primed alloreactive host T cells. PMID- 11120314 TI - Heat shock inhibits the acute phase response in primary rat hepatocytes. PMID- 11120313 TI - An evaluation of a national minority community- based education program to sustain behavior modification. PMID- 11120315 TI - Neuroblastoma inhibits dendritic cell differentiation and function. PMID- 11120316 TI - GTP protein regulation of cell motility. PMID- 11120317 TI - Endogenous interleukin 6 production protects against histopathologic damage in a murine model of ischemia/reperfusion injury. PMID- 11120318 TI - p16 (cdkN2/MTS1/INK4A) expression in sporadic colorectal carcinomas. PMID- 11120319 TI - Effective treatment of pyriform sinus carcinoma: a challenge for the future. PMID- 11120320 TI - TNF-alpha in trauma patients: a touch is just enough. PMID- 11120321 TI - IL-11 is upregulated in human NEC. PMID- 11120322 TI - GTP-cyclohydrolase and cytokine gene expression in NEC. PMID- 11120323 TI - Concurrent reduction of glycogenolysis, glycolysis, and NA(+)/K(+) pump activity after hemorrhagic shock. PMID- 11120325 TI - Expression of extracellular matrix proteases by fetal and adult murine skin fibroblasts cultured in in vitro matrices. PMID- 11120324 TI - Intracoronary E- and L-selectin blockade attenuates myocardial neutrophil infiltration in cardiac ischemia/reperfusion injury. PMID- 11120326 TI - Apoptosis in newly arterialized vein grafts. PMID- 11120327 TI - External beam radiation reduces venous neointimal hyperplasia in PTFE dialysis grafts. PMID- 11120328 TI - Breast surgery of the future: will your training prepare you for credentialing? PMID- 11120329 TI - Updates on cytogenetics and molecular genetics of bone and soft tissue tumors: Ewing sarcoma and peripheral primitive neuroectodermal tumors. PMID- 11120330 TI - Recurrent chromosome changes in 62 primary gastric carcinomas detected by comparative genomic hybridization. AB - Comparative genomic hybridization (CGH) has been applied to detect recurrent chromosome alterations in 62 primary gastric carcinomas. Several nonrandom chromosomal changes, including gains of 8q (31 cases, 50%), 20q (29 cases, 47%) with a minimum gain region at 20q11. 2-q12, 13q (21 cases, 34%) with a minimum gain region at 13q22, and 3q (19 cases, 31%) were commonly observed. The regions most frequently lost included: 19p (23 cases, 37%), 17p (21 cases, 33%), and 1p (14 cases, 23%). High copy number gain (DNA sequence amplification) was detected in 6 cases. Amplification of 8q23-q24.2 and 20q11.2-q12 were observed in 3 cases. Gain of 20q and loss of 19p were confirmed by fluorescence in situ hybridization using corresponding bacterial artificial chromosomes (BAC) clones from those regions. The gain and loss of chromosomal regions identified in this study provide candidate regions involved in gastric tumorigenesis. PMID- 11120331 TI - Somatic alterations of the androgen receptor CAG repeat in human colon cancer delineate a novel mutation pathway independent of microsatellite instability. AB - The human androgen receptor gene contains a polymorphic CAG repeat region ranging from 8 to about 35 repeats in the normal human population. The repeat length is inversely related to the transactivation potential of the receptor. We have analyzed the repeat length in 50 sporadic colon cancer samples in comparison to surrounding healthy mucosa and have found somatic reductions of up to 10 repeats in 5 cases (10%), 3 of which were complex, probably involving both alleles. Alterations occurred in tumors with and without microsatellite instability indicating that they follow an independent mutation pathway. The similar repeat of the huntingtin gene did not show any somatic alterations in the same cases. No correlation to sex, tumor stage, location, or histology was evident. In the tumors that showed somatic reductions, the reduced allele was present in at least half of the cells and thus in most, if not all, of the tumor component of the sample. Somatic reductions of the androgen receptor CAG repeat thus occur frequently, through a pathway distinct from microsatellite instability and early during colon carcinogenesis. The receptor is expressed in most normal and neoplastic tissue samples analyzed. Apparent growth selection of cells bearing shortened AR alleles suggests that androgens contribute to colon carcinogenesis in a yet unknown way. PMID- 11120332 TI - Simultaneous PML/RARalpha and AML1/ETO expression with t(15;17) at onset and relapse with only t(8;21) in an acute promyelocytic leukemia patient. AB - We report a patient with acute promyelocytic leukemia with the common translocation (15;17) and PML-RARAalpha fusion gene. In relapse, blasts showed typical FAB M2 morphologic features, and the karyotype was 45,X, -Y,t(8;21). A reexamination of the leukemic cells at diagnosis revealed that an AML1-ETO fusion gene was also present at that time without cytogenetic evidence of t(8;21). In relapse, only t(8;21) was detected. Two different clones were identified by cytogenetic standard techniques. The association of two common translocations supervening in the same time in the same cells could not be established. PMID- 11120333 TI - Lack of mutations of BCL6 and BCL10 genes in mucosa-associated lymphoid tissue lymphomas of the orbital adnexa. AB - Knowledge regarding the molecular pathogenesis and progression of mucosa associated lymphoid tissue (MALT) lymphomas of ocular adnexa is limited. Eleven cases of ocular MALT lymphoma were analyzed by clonal rearrangement of antigen receptor genes using Southern blot hybridization. Polymerase chain reaction single stranded conformational polymorphism analysis and DNA sequencing was utilized to analyze the mutations of BCL6 and BCL10 genes. Clonal rearrangement of immunoglobulin heavy genes was found in all 11 patients. No point mutation was found in BCL6 or BCL10 genes in any of the samples analyzed. We suggest that mutations of BCL6 and BCL10 genes are rare in low-grade MALT lymphoma of ocular adnexa and are unlikely to be involved in the pathogenesis of the disease. But the role of alterations of both BCL6 and BCL10 genes in the disease progression of low-grade MALT lymphoma require additional study. PMID- 11120334 TI - Sister chromatid exchange frequency in B-cells stimulated by TPA in chronic lymphocytic leukemia. AB - Chronic lymphocytic leukemia (CLL) is characterised by the clonal proliferation and accumulation of neoplastic B-lymphocytes. The median age of the patients is 65 years, and more men than women are affected. The overwhelming majority of CLLs are of B-cell origin. Chromosomal aberrations have been detected in more than 50% of the B-cells obtained from peripheral blood samples after appropriate stimulation with polyclonal B-cell mitogens. The analysis of sister chromatid exchange is a cytogenetic technique used to show DNA damage due to an exchange of DNA fragments between sister chromatids. In this study, lymphocytes from 22 patients with CLL-B (7 female, 15 male; mean age 64.09 +/- 7.56 years) were stimulated by a B-cell mitogen (TPA) and BrdU added at the 24 h of the culture. Metaphase chromosomes were stained with a fluorescence plus Giemsa technique after a standard harvest procedure. The frequency of sister chromatid exchange was found to be increased significantly P =.02) in patients with CLL-B (8.24 +/- 1.36 per metaphase) compared to controls (7.25 +/- 1.42 per metaphase). We conclude that the increased frequency of sister chromatid exchange in chronic lymphocytic leukemia after stimulation with a B-cell mitogen (TPA) may reflect DNA instability and defective DNA repair in these patients. PMID- 11120335 TI - Follicular lymphoma with trisomy 18 and over-expression of BCL2 in the absence of t(14;18)(q32;q21). AB - A 66-year-old female who presented with cervical lymphadenopathy and splenomegaly was found to have large B-cell lymphoma on lymph node biopsy. However, trephine biopsy revealed involvement of the marrow by follicular lymphoma, and cytogenetic study showed an abnormal clone with 47,XX,+18. PMID- 11120336 TI - No evidence of increased chromosomal aberrations and micronuclei in lymphocytes from nonfamilial thyroid cancer patients prior to radiotherapy. AB - The relationship between the presence of high frequencies of chromosomal aberrations in peripheral lymphocytes and predisposition to cancer has been suggested for some cancer diseases. In nonfamilial thyroid cancer, the few reports available are equivocal. The aim of this study was to assess the possible chromosomal instability in peripheral blood lymphocytes from 22 patients suffering from nonfamilial thyroid cancer. For this purpose, 2 classic cytogenetic assays, the chromosomal aberrations assay and cytokinesis-blocked micronucleus assay, were chosen. The frequency of chromosomal aberrations excluding gaps (%) was 1.68 +/- 1.39 (mean value +/- SD) for the patients group versus 2.20 +/- 1.87 for the control group. The frequency of binucleated lymphocytes with micronuclei ( per thousand) was 5.41 +/- 3.51 (mean value +/- SD) for the patients group versus 5.37 +/- 3.21 for the control group. The results obtained revealed no significant differences between both groups. The present study reinforces the idea that constitutional chromosomal instability in peripheral blood lymphocytes is not visible in nonfamilial thyroid carcinomas. PMID- 11120337 TI - Molecular cytogenetics of t(X;1)(p11.2;q21) with complex rearrangements in a renal cell carcinoma. AB - We report a new case of renal cell carcinoma with the translocation (X;1)(p11;q21) and complex structural rearrangements in a female patient of 64 years of age. We analyzed abnormalities using FISH to identify chromosomal rearrangements, and wonder whether the translocation (X;1) could represent a particular subentity in renal cell carcinoma with distinct histologic features. PMID- 11120338 TI - Identification of a novel PTEN intronic deletion in Li-Fraumeni syndrome and its effect on RNA processing. AB - Germline mutations of the TP53 tumor suppressor gene account for the predisposition to cancer observed in many Li-Fraumeni syndrome (LFS) families. A causative genetic factor in the remaining families that harbor no TP53 mutations remains to be elucidated. The PTEN phosphatase tumor suppressor gene is mutated in human cancers observed in LFS. There also exists some phenotypic overlap in the occurrence of cancers in LFS and Cowden's disease (CD), for which germline PTEN mutations are believed to be responsible. We hypothesized that PTEN may be altered in some TP53-wild-type LFS families. We examined LFS primary patient lymphocytes for PTEN alterations using SSCP and sequence analysis. A novel intronic deletion was found in two unrelated individuals, adjacent to the splice acceptor site of PTEN exon 4. Based on an in vitro mRNA processing assay this alteration is predicted to be a polymorphism. The in vivo effects of this proximal splice site deletion are unknown and a genetic cause for the cancers in these families remains to be elucidated. Germline mutations of PTEN were not detected in other families, suggesting that alterations of this tumor suppressor gene do not account for the cancers observed in the subset of LFS individuals with wild-type germline TP53. PMID- 11120339 TI - The Eighth International Workshop on Chromosomes in Solid Tumors. Tucson, Arizona, January 30- February 1, 2000. Abstracts. PMID- 11120340 TI - New worldwide horizons for the International Organization of Psychophysiology (IOP). AB - This Presidential Address deals with the new challenges and horizons of the International Organization of Psychophysiology (IOP) as we are marching towards the 21st century. The importance of Clinical Psychophysiology to humanity is emphasized. The special contribution of the International Journal of Psychophysiology to the future of Psychophysiology as a world forum is underlined. The status of Psychophysiology as a leading neuroscientific discipline is described and the role and major contributions of the late Dr Herbert Jasper as Co-Founding Honorary Fellow of IOP is highlighted. PMID- 11120341 TI - Welcoming address of the Vice-President (Academic Affairs) at the 10th World Congress of Psychophysiology, IOP2000. PMID- 11120342 TI - Welcoming address of the Vice-President (External Affairs) at the 10th World Congress of Psychophysiology, IOP2000. PMID- 11120343 TI - Visual illusions and travelling alpha waves produced by flicker at alpha frequency. AB - The aim of the study was to obtain some experimental evidence of the 'scanning hypothesis' that links electroencephalogram (EEG) alpha-activity with rhythmically spreading waves in the visual cortex. The hypothesis was tested in experiments with 29 healthy adults. Under flicker stimulation through closed lids with the frequency of the individual alpha-rhythm, all subjects perceived illusory visual objects (a ring or a circle, a spiral or a spiral spring, or a grid). Most frequently noted was the perception of a ring or a circle; less frequently, a three-dimensional spiral; and even less frequently, a curved grid. It was found that the optimal stimulation frequency for this effect was tightly connected with the dominant alpha-rhythm frequency, with a correlation coefficient of 0.86. The probability of observing the ring and spiral illusion was highest at this frequency, while that for the grid illusion occurred at frequencies that differed by +/- 1-2 Hz. We observed 10 typical trajectories of travelling EEG alpha-waves on the scalp, and a significant interrelation between the occipital-frontal trajectory and illusions of the ring and spiral. The link between these effects and the propagation of the wave process through the visual cortex, as reflected by the EEG alpha-rhythm, is discussed. The data support the hypothesis of (Pitts, W. McCulloch, W.S., 1947), which proposes the scanning of the visual cortex by a spreading wave process operating at the frequency of the alpha-rhythm, which reads information from the visual cortex. PMID- 11120344 TI - Cognitive modulation of midbrain function: task-induced reduction of the pupillary light reflex. AB - The activation of processing resources has widespread effects in the nervous system. A model of pupillary control systems (Steinhauer S.R. , Hakerem G., 1992. The pupillary response in cognitive psychophysiology and schizophrenia. Ann. N.Y. Acad. Sci. 658, 182-204) had predicted that ongoing cognitive activation should result in inhibition of the light reaction at the level of the oculomotor nucleus, n. III, in the midbrain. In this study, modification of parameters of the pupillary reaction to light were examined during varying task demands. The averaged light reaction was recorded from 33 male and female healthy volunteers during the performance of a serial 7 subtraction task and compared to a 'no task' condition. For 13 subjects, an additional verbalization task with little processing demand (add 1) also was presented. Two types of effects were observed. Firstly, the tonic pupil diameter increased from the no task to the easy (add 1) task, and increased further in the more demanding condition (subtract 7). Secondly, the extent of the phasic light reaction was significantly reduced and the latency at the end of the contraction was significantly decreased in the 'subtract 7' condition compared to both the no task and easy conditions (which did not differ from each other). The locus of interference with the light reaction was the Edinger-Westphal complex of the oculomotor nucleus, which is the motor center for the pupillary sphincter muscles. Descending cortical influences inhibited the activity of the Edinger-Westphal complex. Thus, increasing activation had a tonic inhibitory effect on this center, while higher levels of processing complexity produced a separate component of inhibition that interacted with dynamic activation at this midbrain site. It was suggested that the variation in the light reaction is quantitatively responsive to varying processing loads, and may be utilized as a sensitive metric for a wide variety of cognitive operations. PMID- 11120345 TI - Synchronizing effect of clock rhythm on the 'when to move' decision in repeated voluntary movements. AB - In 13 volunteers, surface EEGs from F(z), C(z), C(3), C(4), and P(z) electrodes and EMG from the flexor digitorum communis were recorded during an experimental task in which the performance of wrist flexion was linked with the selection of a clock sound. In the epoch from 4 to 1 s before movement, more than 95% of the records exhibited a small negative shift from the baseline. The position of these shifts on the time axis made it possible to create groups of time trials. The average EEG curves from these groups showed one prominent negative shift from the baseline, which could be related to the position of particular clock sounds. The slope values of these shifts were highest under the F(z) and C(z) electrodes. We suggested that these negative shifts coincided with 'when to move' decisions, which preceded the execution of the movement at varying intervals. PMID- 11120346 TI - EEG alpha activity and the ERP to target stimuli in an auditory oddball paradigm. AB - Peak-to-peak amplitudes of the N1P2 and N2P3 components in the target ERPs from a fixed interstimulus interval auditory oddball paradigm were investigated as a function of within-subject pre-stimulus levels of alpha activity. Fourteen subjects were each presented with 600 auditory stimuli in a two-tone auditory oddball paradigm which required a button-press to targets, presented with 50% probability. Pre-stimulus alpha activity at Pz was assessed for each trial by digital filtering from 8 to 13 Hz, and alpha RMS amplitude was used to sort the ERPs at Pz and Cz. A direct relationship was obtained between component amplitudes at both Pz and Cz and pre-stimulus alpha level at Pz. Component latencies were strongly related to post-stimulus alpha peaks and troughs. These data confirm the intimate relationship between central nervous system activation, as evidenced by spontaneous EEG in the pre-stimulus period, and the ERP resulting from stimulus presentation. PMID- 11120347 TI - Updating of working memory in a running memory task: an event-related potential study. AB - The aim of the study was to identify central executive activity in the event related potential (ERP) in the time and space domain. Lists of three to eight consonants were presented sequentially. After each list the ordered recall of the three most recent items was required. In this running memory task the updating of working memory contents from the fourth letter on may be understood as a control process. ERPs elicited from each consonant presented in the lists were subtracted from those of a control condition that was also applied to the participants. The difference waveforms showed fronto-central distributed positivities, probably indicating the activity of a postulated central executive. This finding confirms those of neuroimaging studies that localize executive activity to prefrontal brain areas. PMID- 11120348 TI - The effect of data aggregation on temporal stability of cardiovascular reactivity. AB - Temporal stability of behaviorally evoked cardiovascular responses is important for theoretical (concept of activation) and practical (risk for cardiovascular diseases) reasons. As in test psychology, reliability of physiological responsivity depends on the degree of data aggregation across several measurements. This paper describes a statistical approach based on intra-class correlations. This approach is suited to define certain stability measures based on variance components representing different levels of data aggregation. An empirical investigation is presented comprised of 58 subjects, three physiological parameters (heart rate, systolic and diastolic blood pressure), two mental tasks, two sequences of the tasks within one session, and 2 days with an interval of 4 weeks between them. In addition to the finding that data aggregation can generally increase stability, the different sources of aggregation (across phases within a task, across tasks, and across task sequences) and their combinations are systematically compared with regard to their contribution to this enhancement. Finally, it will be shown how the approach can be utilized to explain aggregation effects for other psychophysiological research questions such as covariation, consistency, and ambulatory assessment of cardiovascular functioning. PMID- 11120349 TI - Frontal steady-state potential changes predict long-term recognition memory performance. AB - Converging evidence from event-related potential and functional brain imaging studies suggests that the brain activity at posterior regions of the frontal cortex can predict the strength of long-term memory traces. This study examined the relationship between posterior frontal steady-state visually evoked potential (SSVEP) latency changes and recognition memory after a delay of 7 days. Thirty five female subjects viewed an 18-min television documentary program interspersed with 12 unfamiliar television advertisements while brain electrical activity was recorded from four pre-frontal, two posterior frontal and two occipital scalp sites. After 7 days, the recognition memory was tested for images coinciding with the 20 most prominent frontal SSVEP latency minima and maxima during the viewing of ten contiguous advertisements (advertisements 2-11). We found that images coinciding with posterior frontal latency minima were more likely to be recognized (58.7% recognition) than images coinciding with SSVEP latency maxima (45.3% recognition). Furthermore, the relationship between posterior frontal SSVEP latency and recognition performance after 7 days was only apparent at the left posterior frontal site. The correlation between the recognition performance and SSVEP latency evaluated at all eight sites reached significance only at the left posterior frontal site. These findings suggest that frontal SSVEP latency variations can be used to assess the strength of long-term memory encoding for naturalistic stimuli. PMID- 11120350 TI - Mutation of Thr445 and Ile500 of initiation factor 2 G-domain affects Escherichia coli growth rate at low temperature. AB - The Escherichia coli protein synthesis initiation factor IF2 is a member of the large family of G-proteins. Along with translational elongation factors EF-Tu and EF-G and translational release factor RF-3, IF2 belongs to the subgroup of G proteins that are part of the prokaryotic translational apparatus. The roles of IF2 and EF-Tu are similar: both promote binding of an aminoacyl-tRNA to the ribosome and hydrolyze GTP. In order to investigate the differences and similarities between EF-Tu and IF2 we have created point mutations in the G domain of IF2, Thr445 to Cys, Ile500 to Cys, and the double mutation. Threonine 445 (X1), which corresponds to cysteine 81 in EF-Tu, is well conserved in the DX1X2GH consensus sequence that has been proposed to interact with GTP. The NKXD motif, in which X is isoleucine 500 in IF2, corresponds to cysteine 137 in EF-Tu, and is responsible for the binding of the guanine ring. The recombinant mutant proteins were expressed and tested in vivo for their ability to sustain growth of an Escherichia coli strain lacking the chromosomal copy of the infB gene coding for IF2. All mutated proteins resulted in cell viability when grown at 42 degrees C or 37 degrees C. However, Thr445 to Cys mutant showed a significant decrease in the growth rate at 25 degrees C. The mutant proteins were overexpressed and purified. As observed in vivo, a reduced activity at low temperature was measured when carrying out in vitro ribosome dependent GTPase and stimulation of ribosomal fMet-tRNAfMet binding. PMID- 11120351 TI - Cloning of Rac and Rho-GDI from tobacco using an heterologous two-hybrid screen. AB - To examine whether molecular similarities exist between the animal and plant Rho GTPase signaling pathways, we have developed a heterologous two-hybrid screening method. By this technique, we have cloned a cDNA encoding a tobacco Rac-like protein able to interact with a mammalian Rho-GDI. In a second screen this tobacco Rac was used as a bait and a tobacco homologue of Rho-GDI was identified. These results show that some components of the animal and plant Rac signaling pathways are similar enough to allow their interaction in an heterologous approach. Moreover these data suggest a similar regulation of Rho GTPases in animals and plants. PMID- 11120352 TI - A new model of human aortic endothelial cells in vitro. AB - Vascular endothelial cells play an important role in coagulation regulation of vascular tone and in a variety of synthetic and metabolic functions. Endothelial cells also have a pivotal role in immunological diseases atherogenesis and tumor angiogenesis. Endothelial cells are often used as system to study the pathophysiology of late complications in diabetes mellitus atherosclerotic damages and leukocyte adhesion in inflammatory diseases. Most of the studies have been performed on primary arterial and venous endothelial cell cultures with problems such as availability of autoptic material and reproducibility of cell cultures. We have isolated and characterized a novel system of proliferating long term cultures of human aortic endothelial cells that maintain their differentiated characteristics for many generations in vitro. They produce antithrombotic and thrombotic factors such as t-PA and PAI-1 and respond to TNFalpha, an important factor correlated with the inflammatory process by modifying growth characteristics by producing cytokines such as GM-CSF by expressing ICAM-1 on the surface and by producing large amounts of nitric oxide and endothelin. This new system may be very useful to understand and study the molecular mechanisms involved in many vascular alteration pathologies and in the aging process. PMID- 11120353 TI - PARP degradation in apoptotic Syrian hamster embryo (SHE) cells compared to HL60 cell line. AB - In this study, we attempted to identify apoptotic Syrian hamster embryo (SHE) cells by detecting the specific cleavage of poly(ADP-ribose)polymerase (PARP). Apoptosis was unequivocally identified in serum-deprived SHE cells. After protein electrophoresis and transfer, the anti-PARP antibody (C-2-10) was applied in order to visualize PARP degradation and the anti-polymer antibody (LP96-10) was used to identify PARP and its expected 89-kDa fragment on the membrane after renaturation and NAD+ addition. Results showed that PARP rapidly disappeared during apoptosis in SHE cells, but the resulting fragment remained undetectable with the anti-PARP antibody and no stable polymerase activity of this fragment was measured using anti-polymer antibody. Serum-starved SHE cells were compared to the etoposide-treated HL60 cell line as a control for typical apoptosis related PARP cleavage. These results underline the fact that while PARP degradation is a criterion for apoptosis, the diagnosis of apoptosis can not rely exclusively on the appearance of its 89-kDa fragment as this signal may fail to appear in some cell systems. PMID- 11120354 TI - Human homeodomain-interacting protein kinase-2 (HIPK2) is a member of the DYRK family of protein kinases and maps to chromosome 7q32-q34. AB - Here we identified the human serine/threonine kinase HIPK2 as a novel member of the DYRK kinase subfamily. Alignment of several DYRK family proteins including the kinases minibrain, MJAK, PKY, the Dictyostelium kinase YakA and Saccharomyces YAK1 allowed the identification of several evolutionary conserved DYRK consensus motifs within the kinase domain. A lysine residue conserved between all DYRK kinase family members was found to be essential for the kinase function of HIPK2. Human HIPK2 was mapped to chromosome 7q32-q34 and murine HIPK2 to chromosome 6B, the homologue to human chromosome 7. PMID- 11120355 TI - Evidence of tissue-specific, post-transcriptional regulation of NRF-2 expression. AB - Mitochondrial respiratory function requires the expression of genes both from the mitochondrial and nuclear genomes. Nuclear respiratory factor 2 (NRF-2) is a transcription factor required for the expression of several nuclear-encoded mitochondrial proteins, including the specific mitochondrial transcription factor Tfam. This makes NRF-2 a likely candidate to coordinate expression of mitochondrial components. NRF-2 is a multisubunit complex of which the alpha subunit binds DNA and the beta subunit enhances this binding, respectively. We have analysed in vivo the expression patterns of NRF-2 subunits both at the mRNA and protein level, in three rat tissues, liver, testis and brain. In contrast with Tfam or the 'housekeeping' beta-actin expressions in which a parallel gradient was observed, no correlation was found between NRF-2 mRNAs and proteins levels, thus suggesting post-transcriptional regulation. PMID- 11120356 TI - Hemoglobin binding sites on renal brush-border membranes. AB - Prolonged exposure of renal tubules to hemoglobin markedly reduces kidney function and eventually leads to acute renal failure called pigment nephropathy. Intracellular hemoglobin toxicity is one of main pathomechanisms involved in the disease development. However, the process in which hemoglobin is taken up by renal tubular epithelium has not been characterized so far. Isolated renal brush border membranes of the rat and radioiodinated rat and human hemoglobins were used. Binding properties were examined by the use of rapid filtration technique. Partial isolation of hemoglobin binding proteins was achieved by affinity chromatography. Our experiments showed that both human and rat hemoglobins can be specifically bound to renal brush-border membranes by one class of low affinity (Kd, 7.7 microM) and high capacity (Bmax, 0.18 nmol/mg protein) binding sites. The sites were relatively selective for hemoglobin. Albumin did not compete with hemoglobin. Cationic molecules cytochrome C and lysine exhibited some competition while strong competition of myoglobin was observed. The binding was affected by EGTA indicating a Ca2+ requirement for the interaction. There was a rise in binding in pH 5.4. Fall in binding activity after preincubation of the membranes with peptidases suggested the proteinaceous nature of the binding sites. Affinity chromatography of membrane proteins extract yielded heterogeneous preparation consisting of proteins with molecular masses of 110, 72, 38 and 27 kDa respectively. The existence of binding sites for hemoglobin in renal brush-border membranes strongly suggests that uptake of the protein by tubular epithelia occurs via adsorptive endocytosis. Increased binding of hemoglobin to the membranes under acidic conditions may explain exacerbation of hemoglobinuric acute renal failure in aciduric states. PMID- 11120357 TI - 2-Hydroxyacid dehydrogenase from Haloferax mediterranei, a D-isomer-specific member of the 2-hydroxyacid dehydrogenase family. AB - An NAD-dependent D-2-hydroxyacid dehydrogenase (EC 1.1.1.) was isolated and characterized from the halophilic Archaeon Haloferax mediterranei. The enzyme is a dimer with a molecular mass of 101.4 +/- 3.3 kDa. It is strictly NAD-dependent and exhibits its highest activity in 4 M NaCl. The enzyme is characterized by a broad substrate specificity 2-ketoisocaproate and 2-ketobutyrate being the substrates with the higher Vmax/Km. When pyruvate and 2-ketobutyrate were the substrates the optimal pH was acidic (pH 5) meanwhile for 2-ketoisocaproate maximum activity was achieved at basic pH between 7.5 and 8.5. The optimum temperature was 52 degrees C and at 65 degrees C there was a pronounced activity decrease. This new enzyme can be used for the production of D-2-hydroxycarboxylic acid. PMID- 11120358 TI - Cross-beam vector Doppler ultrasound for angle-independent velocity measurements. AB - Combining Doppler measurements taken along multiple intersecting ultrasound (US) beams is one approach to obtaining angle-independent velocity. Over 30 laboratories and companies have developed such cross-beam systems since the 1970s. Early designs focused on multiple single-element probes. In the late 1980s, combining multiple color Doppler images acquired from linear-array transducers became a popular modality. This was further expanded to include beam steering and the use of subapertures. Often, with each change in design, came a new twist to calculating the velocity. This article presents a review of most proposed cross-beam systems published to date. The emphasis is on the basic design, the approach used to determine the angle-independent velocity, the advantages of the design, and the disadvantages of the design. From this, requirements needed to convert the idea of angle-independent vector Doppler into a commercial system are suggested. PMID- 11120359 TI - Correlation of ultrasonographic imaging of congenital muscular torticollis with clinical assessment in infants. AB - Congenital muscular torticollis (CMT) is a common problem affecting infants and children. There is a general lack of standard clinical classification or objective assessment methods. Ultrasonographic imaging of the sternomastoid muscle (SCM) has been carried out in a consecutive series of 436 infants less than 1 y old presenting with CMT over a 5-y period. All patients were classified into three clinical groups: postural torticollis, muscular torticollis and sternomastoid tumor. The severity of the torticollis was also expressed into four subgroups according to the degree of deficits in passive rotation of the neck. The ultrasonographic image of the affected SCM included the echogenicity, texture, motility, softness and the transverse and longitudinal extent of the involvement. The disturbance in the quantitative measurement of the transverse diameter of the lower and upper third of the SCM and the ratio of the measurement to the normal side was recorded. The qualitative and quantitative changes in the SCM image were found to correlate significantly with the clinical typing and severity of rotational deficits of the neck. Ultrasonographic imaging has important potential clinical application in helping the diagnosis, prognostication and monitoring of progress of CMT longitudinally. PMID- 11120360 TI - Relevance of sonographic B-mode criteria and computer-aided ultrasonic tissue characterization in differential/diagnosis of solid breast masses. AB - We aimed to evaluate the differential diagnostic value of a method of computer assisted texture analysis in comparison to established ultrasonographic B-mode characteristics in the examination of solid breast masses. At two centers, 77 patients presenting with a solid mass on B-mode scan were studied at 7.5 MHz. Description of B-mode appearance included assessment of tumor shape, borders, presence of an echogenic rim, tissue architecture, internal echo structure, absorption and elasticity. For statistical pattern recognition, the following parameters were used: form factor, mean grey level, signal-to-noise ratio, mean gradient and correlation from the co-occurrence matrix. At center 1, the most decisive parameter for differential diagnosis was distortion of tissue architecture (sensitivity, SN, 83%; specificity, SP, 92%) and, at center 2, relation to the adjacent tissue (SN 93%, SP 92%). Among texture parameters, best discrimination was achieved for correlation from the co-occurrence matrix at center 1 (SN 58%, SP 73%) and for form factor at center 2 (SN 93%, SP 77%). Among sonographic criteria, the highest contribution to the diagnosis was found for an unsharp border (odds ratio, OR, 12.2), architectural distortion (OR, 8.6), fixation to skin or chest wall (OR, 9.0) and fixation to adjacent breast parenchyma (OR, 8.8), according to texture analysis for parameters form factor (OR 4.0) and correlation from the co-occurrence matrix (OR 4.7). Ultrasonographic texture analysis can be helpful as an additional parameter in differential diagnosis of breast tumors, but did not reach differential diagnostic accuracy of sonomorphologic features. PMID- 11120361 TI - Fetal renal volume in normal gestation: a three-dimensional ultrasound study. AB - To establish a reference chart of fetal kidneys in normal pregnancy, we performed a prospective and cross-sectional study. A total of 152 singleton fetuses ranging between 20 and 40 weeks' gestation and meeting the criteria of normal pregnancies were included. Three-dimensional ultrasound (3-D US) was used to measure the fetal renal volume. Our results revealed that both renal volumes are highly correlated with the fetal gestational age. Using gestational age (GA) as the independent variable and right renal volume (RRV) as the dependent variable, the best-fit regression equation was RRV (mL)=0.74053xGA (week)-13.318 (r = 0.89, p < 0.001). Similarly, the best-fit equation for the left renal volume (LRV) was LRV (mL)=0. 76093xGA (week)-13.421 (r = 0.86, p < 0.001). The normal growth centiles of both kidneys were established based on these two equations. There were no significant differences of the volumes between bilateral kidneys. In conclusion, our data of fetal renal volumes assessed by 3-D US may serve as a reference in evaluating fetal renal growth. PMID- 11120362 TI - Noninvasive assessment of brachial artery endothelial function with digital ultrasound and 13-MHz scanning frequency: feasibility of measuring the true inner luminal diameter using the intima-lumen interface. AB - Previous studies assessing endothelial function as flow-mediated changes in the brachial artery diameter have not been able to measure the true inner luminal diameter. This is due to the lack of image quality, which has hampered the visualisation of the lumen-intimal interface. Because increases in resolution and scanning frequency have recently led to improved ultrasound (US) image quality, we assessed the feasibility of measuring the true brachial artery diameter using digital US and 13-MHz scanning frequency. Satisfactory true inner diameter measurements were obtained in all subjects (n = 148, middle-aged men, mean age 54 +/- 7 y) participating in a risk factor study. At baseline flow, the intima to intima diameter was 4.03 +/- 0.49 and 4.67 +/- 0.52 mm measured conventionally from the anterior to the posterior media-adventitia interface (difference 0.64 +/ 0.10 mm). After hyperaemia, the intima to intima diameter was 4.23 +/- 0.46 mm and the adventitia to adventitia diameter 4.86 +/- 0.50 mm. Flow-mediated dilation (FMD) expressed as the percentage change from the baseline diameter measured 5.3 +/- 4.3% using the true inner diameters and 4.3 +/- 3.7% using the conventional outer diameters. The difference in FMD values was systematic, and there was a good linear correlation between them (r = 0.93, p < 0.0001). If FMD is presented as the percentage change from baseline to hyperaemia, this new method gives values that are approximately 1% unit higher, compared with values when brachial luminal diameter is measured in the conventional way between the adventitia-media interfaces. PMID- 11120363 TI - Flow velocity and flow volume measurements in the extracranial carotid and vertebral arteries in healthy adults: reference data and the effects of age. AB - To establish reference data and to investigate the development of haemodynamics in the extracranial carotid and vertebral arteries, we performed a prospective study in 78 age- and gender-matched healthy adults from 20 to 85 y old. Angle corrected flow velocities and luminal diameters were measured and waveform parameters and flow volumes calculated in all the arteries. Side-to-side differences and the influence of age on these parameters were also investigated. In the common carotid arteries, the internal carotid arteries and the vertebral arteries (CCA, ICA and VA, respectively) all flow velocities decreased significantly during ageing. The luminal diameter remained constant in all the carotid arteries, but increased slightly with age in the VA. An age-related decline of intravascular flow volume was observed in the ICA. Due to a pronounced decrease in end-diastolic flow velocity, the resistance index decreased in ICA and VA during ageing. There were no significant side-to-side differences in flow velocities and flow volumes in any of the extracranial arteries. The luminal diameters of the CCA, ICA and ECA were significantly smaller in women than in men. No relevant gender-related differences in flow velocities or waveform parameters were found in the extracranial arteries. There was no gender-linked difference in the flow volumes of the brain-feeding arteries and, in the ECA, flow volumes were significantly higher in men. Reference data on all flow velocities and waveform parameters, luminal diameters and flow volumes were established for different age groups between 20 and 85 y old. These data allow us to outline the development of cerebral haemodynamics during "benign ageing" and to utilise flow volume measurements in clinical practice, especially in patients with cerebrovascular diseases. PMID- 11120364 TI - Blood flow velocity using transcranial Doppler velocimetry in the middle and anterior cerebral arteries: correlation with sample volume depth. AB - The baseline transcranial Doppler studies of the middle (MCA) and anterior cerebral (ACA) arteries of 135 patients were retrospectively reviewed. 3312 data points were classified according to gender, vessel, mean blood flow velocity (MBFV), systolic blood flow velocity (SBFV), diastolic blood flow velocity (DBFV) and sample volume depth (SVD). The flow velocities at each depth were averaged, and plotted against acquisition depth. The male MCA yielded a correlation coefficient (r), between MBFV and SVD of 0.999. For the female MCA, r = 0.986. For the ACA, male, r = 0.911, and female, r = 0.894. All other data showed r values >/= 0.563, with 11 of 12 r values >/= 0.808. These data suggest that BFV reduces with SVD in a highly predictable way for the MCA and ACA, from which expected BFV values may be estimated at any given SVD. PMID- 11120365 TI - 40 MHz Doppler characterization of umbilical and dorsal aortic blood flow in the early mouse embryo. AB - Physiological study of the developing mouse circulation has lagged behind advances in molecular cardiology. Using an innovative high-frequency Doppler system, we noninvasively characterized circulatory hemodynamics in early mouse embryos. We used image-guided 43 MHz pulsed-wave (PW) Doppler ultrasound to study the umbilical artery and vein, or dorsal aorta in 109 embryos. Studies were conducted on embryonic days (E) 9.5-14.5. Heart rate, peak blood flow velocities, and velocity time integrals in all vessels increased from E9.5-14.5, indicating increasing stroke volume and cardiac output. Heart rate, ranging from 192 bpm (E9.5) to 261 bpm (E14.5), was higher than previously reported. Placental impedance, assessed by the time delay between the peaks of the umbilical arterial and venous waveforms and by venous pulsatility, decreased with gestation. Acceleration time, a load-independent Doppler index of cardiac contractility, remained constant but seemed sensitive to heart rate. High-frequency PW Doppler is a powerful tool for the quantitative, noninvasive investigation of early mouse circulatory development. PMID- 11120366 TI - Measurement of aortic arch distension wave with the echo-track technique. AB - The aim of this study was to use the echo-track method for measuring aortic arch diameter, distension waveform and elastic parameters. Data were obtained from 50 healthy volunteers of 32 +/- 15 y (mean +/- 1 SD). The aortic arch was interrogated from the suprasternal position with M-mode ultrasonography using a 3.5-MHz transducer; diameter and distension waves were determined by means of an echo-track algorithm (WTS, Pie Medical); arterial blood pressure was measured in the arm with sphygmomanometry. Aortic arch diameter, distension, distensibility and compliance were 24.55 +/- 2.99 mm, 2199 +/- 726 micrometer, 3.9 +/- 1.4. 10( 3) mmHg(-1) and 1.86 +/- 0.61 mm(2). mmHg(-1), respectively. Intrasession, interobserver and intersession variability was less than 10%, 10% and 18%, respectively. It is concluded that aortic arch distension wave can be recorded noninvasively with acceptable reproducibility, allowing assessment of aortic elastic parameters, and yielding insight into pressure wave reflection within the arterial system. PMID- 11120367 TI - Predicting the acoustic response of a microbubble population for contrast imaging in medical ultrasound. AB - Although the behavior of a bubble in an acoustic field has been studied extensively, few theoretical treatments to date have been applied to simulate the acoustic response of a real population of variably sized microbubbles in a finite width sound beam. In this paper, we present a modified Trilling equation for single bubble dynamics that has been solved numerically for different conditions. Radiated waveforms from a large number of such bubbles are combined, reflecting their size distribution and location and the shape of a real acoustic beam. The resulting time-domain pressure waveforms can be compared with those obtained experimentally. The dependence of second-harmonic radiation on incident focal amplitude at different frequencies is presented. This model is particularly suited to the study of interaction between a medical ultrasound beam and microbubble contrast agents in aqueous media. PMID- 11120368 TI - In vitro validation of volumetric blood flow measurement using Doppler flow wire. AB - Determination of any volumetric blood flow requires assessment of mean blood flow velocity and vessel cross-sectional area. For evaluation of coronary blood flow and flow reserve, however, assessment of average peak velocity alone is widely used, but changes in velocity profile and vessel area are not taken into account. We studied the feasibility of a new method for calculation of volumetric blood flow by Doppler power using a Doppler flow wire. An in vitro model with serially connected silicone tubes of known lumen diameters (1.5, 2.0, 2.5, 3.0, 3.5 and 4.0 mm) and pulsatile blood flow ranging from 10 to 200 mL/min was used. A Doppler flow wire was connected to a commercially available Doppler system (FloMap(R), Cardiometrics) for online calculation of the zeroth (M(0)) and the first (M(1)) Doppler moment, as well as mean flow velocity (V(m)). Two different groups of sample volumes (at different gate depths) were used: 1. two proximal sample volumes lying completely within the vessel were required to evaluate the effect of scattering and attenuation on Doppler power, and 2. distal sample volumes intersecting completely the vessel lumen to assess the vessel cross sectional area. Area (using M(0)) and V(m) (using M(1)/M(0)) obtained from the distal gates were corrected for scattering and attenuation by the data obtained from the proximal gates, allowing calculation of absolute volumetric flow. These results were compared to the respective time collected flow. Correlation between time collected and Doppler-derived flow measurements was 0.98 (p < 0.0001), with a regression line close to the line of equality indicating an excellent agreement of the two measurements in each individual tube. The mean paired flow difference between the two techniques was 1.5 +/- 9.0 mL/min (ns). Direct volumetric blood flow measurement from received Doppler power using a Doppler flow wire system is feasible. This technique may potentially be of great clinical value because it allows an accurate assessment of coronary flow and flow reserve with a commercially available flow wire system. PMID- 11120369 TI - Relationships of ultrasonic backscatter with ultrasonic attenuation, sound speed and bone mineral density in human calcaneus. AB - Ultrasonic attenuation and sound speed have been investigated in trabecular bone by numerous authors. Ultrasonic backscatter has received much less attention. To investigate relationships among these three ultrasonic parameters and bone mineral density (BMD), 30 defatted human calcanei were investigated in vitro. Normalized broadband ultrasonic attenuation (nBUA), sound speed (SOS), and logarithm of ultrasonic backscatter coefficient (LBC) were measured. Bone mineral density was assessed using single-beam dual energy x-ray absorptiometry (DEXA). The correlation coefficients of least squares linear regressions of the three individual ultrasound (US) parameters with BMD were 0.84 (nBUA), 0.84 (SOS) and 0.79 (LBC). The 95% confidence intervals for the correlation coefficients were 0. 69-0.92 (nBUA), 0.68-0.92 (SOS) and 0.60-0.90 (LBC). The correlations among pairs of US variables ranged from 0.63-0.79. Variations in nBUA accounted for r(2) = 62% of the variations in LBC. Variations in SOS accounted for r(2) = 40% of the variations in LBC. These results suggest that ultrasonic backscattering properties may contain substantial information not already contained in nBUA and SOS. A multiple regression model including all three US variables was somewhat more predictive of BMD than a model including only nBUA and SOS. PMID- 11120370 TI - Comparative sensitivity of human and bovine erythrocytes to sonolysis by 1-MHz ultrasound. AB - This project tested the hypothesis that human erythrocytes, being larger than bovine erythrocytes, would be the more sensitive to sonolysis induced by inertial cavitation. The rationale behind this hypothesis was an earlier demonstration that, among sized populations of erythrocytes, an inverse relation existed between erythrocyte volume and mechanically-induced shear forces in the surrounding medium; viz, the larger the cell, the less shear force required to rupture the cell's membrane. At low erythrocyte densities (i.e., approximately 5% hematocrit) the hypothesis was supported; at high cell densities (i.e., approximately 35% hematocrit) it was not supported. The data are consistent with an ultrasound (US)-induced symmetric implosion of affected gas nuclei as causing the effect at low cell densities; under such conditions there is ample spacing among cells for US-induced symmetric growth and collapse of gas nuclei and the concomitant production of radially-expanding shock waves (which lyse the cells); at high cell densities there is not sufficient spacing among cells for US-induced symmetric growth and collapse of bubbles and an alternative mechanism, possibly asymmetric bubble collapse, becomes operational. PMID- 11120371 TI - Bioeffects of positive and negative acoustic pressures in mice infused with microbubbles. AB - This study provided one test of the hypothesis that hemorrhage in tissues containing ultrasound (US) contrast agents results from inertial cavitation. The test relied on the prediction of classical cavitation theory that the response of microbubbles to negative pressures is much greater than it is for positive pressures. An endoscopic electrohydraulic lithotripter was used to generate a spherically diverging positive pressure pulse. A negative pressure pulse was produced by reflection of the positive pulse from a pressure release interface. Mice were injected with approximately 0. 1 mL of Albunex(R) and exposed to 100 pulses at either + 3.6 MPa or -3.6 MPa pressure amplitude. For comparison, mice were also exposed to the same acoustic fields without injection of contrast agents. Sham animals experienced the same protocols, with or without Albunex(R) injections, but were not exposed to the lithotripter fields. Following exposure, mice were scored for hemorrhage to various organs and tissues. When Albunex(R) was present in the vasculature, negative pressure pulses produced significantly more hemorrhage than positive pressures in tissues such as the kidney, intestine, skin, muscle, fat, mesentery and stomach. PMID- 11120372 TI - Selective transmission of a focused Doppler ultrasound beam through a plastic layer. AB - Laboratory test objects are widely used in Doppler ultrasound (US). Although the acoustic properties of in vitro materials are usually known, they are unlikely to match each other, or their in vivo counterparts, exactly. We conducted theoretical and experimental studies of a focused ultrasound beam as it passes from one fluid, through an intervening plastic layer at an oblique angle, and then into a different fluid. Dual mode propagation may occur (i.e., both longitudinal and shear waves can propagate in the plastic layer). Our calculations show that the power transmitted by either mode drops very rapidly to zero at certain critical angles. A range of angles of incidence exists within a focused beam and this, combined with the highly angle-dependent power transmission behaviour, can produce major distortions of Doppler data. These may persist even when the beam axis is not oriented exactly at the critical angle. The total power transmitted depends on all the wave speeds, may involve mode conversion, and is a very complicated function of the angle of incidence. This study reports a practical method for the calculation of power transmission though a plastic layer, and shows how the resulting power vs. angle graph can be used to avoid artefacts in in vitro Doppler studies. PMID- 11120373 TI - Comparison of finite element and heated disc models of tissue heating by ultrasound. AB - This paper compares different techniques used to model the heating caused by ultrasound (US) in a phantom containing a layer of bone mimic covered by agar gel. Results from finite element (FE) models are compared with those from two techniques based on the point-source solution to the bioheat transfer equation (BHTE): one in which the bone mimic is considered to be an absorbing disc of infinitesimal thickness and the other in which the region through which the US travels is considered to be a volume heat source. The FE results are also compared with experimental measurements. The results from the models differed by up to 40% compared with those from the FE model. Furthermore, for the intensity distribution considered, which corresponds to that in the focal zone of a single element transducer, the top hat distribution predicts a temperature rise 1.8 times greater than that for a more realistic one based on measured values. PMID- 11120374 TI - In vitro spatial compound scanning for improved visualization of atherosclerosis. AB - A new off-line multiangle ultrasound (US) compound scanner has been built with the purpose of investigating possible improvements in visualization of vascular structure. Images of two formalin-fixed human atherosclerotic plaques removed by carotid endarterectomy were recorded from seven insonification angles over a range of 42 degrees and the individual images were combined (averaged) into a single image (spatial compounding). Compared to conventional B-mode imaging, this multiangle compound imaging (MACI) method features images with reduced angle dependence, reduced random variation (speckle) and improved delineation of the plaque outline. With the MACI approach, it is, thus, easier to assess e.g., a possible residual lumen of an atherosclerotic artery as well as the level of echogenicity for the different plaque constituents. PMID- 11120375 TI - Vertebral arteries and neck rotation: Doppler velocimeter interexaminer reliability. AB - The purpose of this study was to test the interexaminer reliability of Doppler ultrasound (US) velocimeter examination of vertebral arteries during contralateral cervical rotation. Vertebral arteries from 20 adults were insonated using a bidirectional Doppler velocimeter at the suboccipital portal (standard technique) and C2 transverse process level (new technique) during contralateral cervical rotation. The data obtained by two examiners, regarding persistence or major reduction in Doppler signals, were compared. There was 93% agreement between the data from the two examiners, and the kappa score was 0.78 at p = 0.05. These results provide evidence to support the interexaminer reliability of bidirectional Doppler velocimeter examination for the purpose of assessing the effects of contralateral rotation on vertebral artery blood flow. PMID- 11120376 TI - Molecular aspects of soluble guanylyl cyclase regulation. AB - Soluble guanylyl cyclase (sGC) is a heterodimeric enzyme (comprised of alpha and beta subunits) that generates the intracellular second messenger cyclic guanosine monophosphate (cGMP) from guanosine triphosphate (GTP). cGMP is subsequently important for the regulation of protein kinases, ion channels, and phosphodiesterases. Since recent evidence has demonstrated that heterodimerization of the alpha/beta subunits is essential for basal and stimulated enzymatic activity, the existence of several types of isoforms for each of the two subunits, along with their varying degrees of expression in different tissues, implies that multiple regulatory mechanisms exist for sGC. Yet, progress in studying and clarifying the regulatory processes that can alter sGC expression and activity has only slowly started being elucidated. In the following paper, we elaborate on sGC structure, function, and distribution along with recently described signaling pathways that modulate sGC gene expression. PMID- 11120377 TI - NO-mediated MaxiK(Ca) channel activation produces relaxation of guinea pig aorta independently of voltage-dependent L-type Ca(2+) channels. AB - The role of L-type Ca(2+) channels in the relaxation to nitric oxide (NO) mediated MaxiK(Ca) channel activation was examined in guinea pig aorta. Acetylcholine (ACh) produced an endothelium-dependent relaxation of guinea pig aorta precontracted with noradrenaline (NA), which was abolished by an NO synthase inhibitor, N(G)-nitro-L-arginine (L-NNA). Both endothelium-dependent relaxation by ACh and endothelium-independent relaxation by an NO donor, (+/-) (E)-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexeneamide (NOR3), were strongly suppressed by a soluble guanylate cyclase (sGC) inhibitor, 1H-[1,2,4]-oxadiazolo [4,3-a]-quinoxalin-1-one (ODQ), suggesting that increased intracellular cGMP plays the key role in both responses. ACh- and NOR3-induced relaxations were significantly suppressed by iberiotoxin (IbTX), a selective blocker of MaxiK(Ca) channels. ACh- and NOR3-induced relaxations were greatly attenuated when arteries were precontracted with high KCl instead of NA, supporting the idea that K(+) channel activation mediates the relaxant responses. (6) NOR3-induced relaxations were not affected by a L-type Ca(2+) channel blocker, diltiazem. Furthermore, endothelium-independent relaxation by a K(ATP) channel opener, (+)-7,8-dihydro-6, 6-dimethyl-7-hydroxy-8-(2-oxo-1-piperidinyl)-6H-pyrano[2,3-f] benz-2,1, 3 oxadiazole (NIP-121) was not affected by diltiazem and nicardipine. These findings suggest that blockade of L-type Ca(2+) channels is not a major mechanism responsible for the vascular relaxation due to NO-mediated MaxiK(Ca) channel activation in guinea pig aorta. PMID- 11120378 TI - Contribution of sodium channel and sodium/hydrogen exchanger to sodium accumulation in the ischemic myocardium. AB - Contribution of sodium channels and sodium/hydrogen exchangers (NHEs) to sodium accumulation during ischemia in the ischemic/reperfused heart was examined. Ischemia increased the myocardial sodium. Reperfusion elicited a further increase in the myocardial sodium, which was associated with little recovery of the left ventricular developed pressure (LVDP) of the perfused heart. Treatment with tetrodotoxin or dimethylamirolide (DMA) dose-dependently attenuated the ischemia- and reperfusion-induced increase in myocardial sodium and enhanced the post ischemic recovery of the LVDP. There was an inverse relationship between the increase in myocardial sodium during ischemia and the post-ischemic recovery of the LVDP.The myocardial sodium accumulation during ischemia is mainly attributed to sodium influx through sodium channels and NHEs. PMID- 11120379 TI - Factors responsible for acetylcholine-induced dilatation in the isolated perfused rat kidney. AB - Mechanism of acetylcholine (ACh)-induced dilatation was investigated in isolated perfused rat kidney. Under a constant flow of 8-10 ml/min, ACh (0.001-3 microg/0.1 ml) caused a dose-dependent decrease in perfusion pressure raised by submaximum concentration of phenylephrine (PE). ACh-induced dilatations were inhibited by atropine (10(-6) mol/l), hexamethonium (10(-4) mol/l), indomethacin (10(-5) mol/l), methylene blue (10(-5) mol/l), N(G)-nitro-L-arginine (L-NOARG, 10(-4) mol/l), tetrodotoxin (TTX, 10(-6) mol/l), capsaicin (10(-6) mol/l), and glibenclamide (10(-5) mol/l). These results suggest that in the isolated perfused rat kidney, endothelium-derived hyperpolarizing factor (EDHF), nitric oxide (NO), and tachykinin neuromediators may play a role in ACh-induced dilatation via stimulation of guanylate cyclase and opening of ATP-sensitive potassium channels. PMID- 11120380 TI - Effect of acute and sub-chronic administration of the imidazoline compound S 22068 on in vivo glucose and insulin responses in normal lean CBA/Ca mice. AB - Acute S 22068 (24 mg/kg po) improved glucose tolerance and increased insulin sensitivity, assessed as the acute blood glucose response to exogenous insulin. The same acute dose did not stimulate insulin secretion or induce hypoglycemia in fed animals. Comparison of acute S 22068 to equipotent doses (with respect to effect on glucose tolerance) of gliclazide (2 mg/kg) and metformin (60 mg/kg) found S 22068 to be similar to metformin with respect to its effects on basal glucose levels (BGL) and insulin sensitivity. This also suggests that S 22068 acts by a mechanism which does not involve insulin release. Acute or sub-chronic S 22068 (14 days at 25 mg/day) had no effect on brown adipose tissue (BAT) or white adipose tissue (WAT) lipogenesis, an insulin-sensitive metabolic pathway. Sub-chronic treatment with S 22068 did not alter body weight (BW) or food intake, and resulted in tolerance to its effects on glucose metabolism and insulin sensitivity. These findings suggest that S 22068 is similar in effect to metformin, and is not insulinogenic, in contrast to the sulfonylureas or putative I(3) imidazoline site ligands. PMID- 11120381 TI - Lidocaine inhibits potassium efflux and hemolysis in erythrocytes during oxidative stress in vitro. AB - Lidocaine is a widely used local anesthetic agent. The aim of this work was to study the action of lidocaine on human red blood cells exposed to an oxidative stress in vitro. Blood was obtained from healthy volunteers. After separation from plasma, the erythrocytes were suspended in phosphate buffer. Oxidative stress was induced by incubation with a free radical generator, the 2,2' azobis (2-amidinopropane) hydrochloride (AAPH). Erythrocytes were incubated with or without lidocaine at two concentrations (36.93 and 73.85 microM) and with or without AAPH (20 mM). Electron paramagnetic resonance (EPR) spectroscopy was performed to identify the free radical species generated by AAPH using the spin trap 5-5'-dimethyl-L-pyroline-N-oxide (DMPO). Different sets of experiments were run. Potassium efflux was measured by flame photometry in each group at time 0 min and every 30 min of the experiment for 2 h. Hemolysis was studied by the Drabkin method at increasing concentrations of AAPH (20, 50, and 100 mM) and with or without lidocaine (36.93 microM). The oxygen radical absorbance capacity (ORAC) was measured by using allophycocyanin (APC) as a fluorescent indicator protein, and the antioxidant capacity of lidocaine (36.93 microM) was studied by the analysis of fluorescence of the APC. AAPH was shown to produce alkoxyl free radicals. Oxidative stress induced a marked increase in the potassium efflux and the hemolysis that was AAPH dose-dependent. Lidocaine inhibited the potassium efflux and delayed the occurrence of hemolysis. Lidocaine did not show any antioxidant properties for the free radical species generated by AAPH. In this model, lidocaine protects erythrocytes against oxidative stress. This effect is not explained by a free radical scavenging property. The results may be of great interest in clinical practice such as intravenous regional anesthesia or the prevention of ischemia-reperfusion injury. PMID- 11120382 TI - Altered endothelium-dependent responsiveness in the aortas and renal arteries of Otsuka Long-Evans Tokushima Fatty (OLETF) rats, a model of non-insulin-dependent diabetes mellitus. AB - We examined endothelium-dependent relaxation in the aortas and renal arteries of Otsuka Long-Evans Tokushima Fatty (OLETF) rats, a model of non-insulin-dependent diabetes mellitus, in comparison with non-diabetic Long-Evans Tokushima Otsuka rats as controls. Acetylcholine-induced relaxation in both arteries was attenuated, and the attenuation was restored to the control level by indomethacin. The relaxation was inhibited completely in the aortas, but only partially in renal arteries by N(G)-nitro-L-arginine methyl ester, and the degree of the latter inhibition was greater in OLETF rats than in the controls. The relaxation was inhibited by aminoguanidine in both arteries of OLETF rats but not in the controls. Serum NO(2) plus NO(3) levels significantly increased in OLETF rats. These results suggest that impairment of relaxation in OLETF rat arteries is due to increased release of contractile factors but not decreased release of nitric oxide. PMID- 11120383 TI - Effects of three different Ca(2+) pump ATPase inhibitors on evoked contractions in rabbit aorta and activities of Ca(2+) pump ATPases in porcine aorta. AB - Using vascular smooth muscle, we describe the actions of three pharmacological tools, cyclopiazonic acid (CPA), thapsigargin (TG) and 2,5-di-(tert-butyl)-1,4 benzohydroquinone (tBHQ), which are presumed to act as selective inhibitors of the sarco-endoplasmic reticulum Ca(2+)-ATPases (SERCAs). In porcine aortic smooth muscle microsomes two Ca(2+)-ATPase activities have been described, one vanadate sensitive and one vanadate-resistant, representing the Ca(2+)-ATPase activities of the plasma membrane and SERCAs, respectively. In agreement, CPA, TG and tBHQ, in the concentration range 0.1 microM to 0.1 mM, dose-dependently inhibit the Ca(2+)-ATPase activity only in the vanadate-resistant microsomes. However, 0.1 mM tBHQ also significantly inhibited the Ca(2+)-ATPase activity of vanadate sensitive microsomes. In rabbit aortic rings, all three SERCA inhibitors produced a dose-dependant inhibition of contractions evoked by 20 mM caffeine or 1 microM phenylephrine (PE) in a Ca(2+)-free physiological solution. However, in PE contracted rings, tBHQ (> or =30 microM) also significantly inhibited the ability of cromakalim to induce relaxation. In conclusion, the data suggest that CPA, TG and tBHQ can all act as selective SERCA inhibitors in both porcine and rabbit aortic smooth muscle. However, in contrast to CPA and TG, high concentrations of tBHQ can exhibit some nonspecific effects, which include inhibition of the plasma membrane Ca(2+)-ATPase and possibly K(+) channels regulated by cromakalim. PMID- 11120384 TI - Long-term effects of postnatal amphetamine treatment on striatal protein kinase A activity, dopamine D(1)-like and D(2)-like binding sites, and dopamine content. AB - The purpose of the present study was to determine whether exposure to amphetamine during the preweanling period would alter dopaminergic functioning in the dorsal striatum of adult rats. In three experiments, we assessed the effects of repeated amphetamine treatment on striatal protein kinase A (PKA) activity, dopamine (DA) D(1)-like and D(2)-like binding sites, and DA content. Rats were pretreated with saline or amphetamine (2.5 mg/kg, ip) for 7 consecutive days starting on postnatal day (PD) 11. At PD 90, rats were killed and their dorsal striata (i.e., caudate-putamen) were removed and frozen until time of assay. Amphetamine pretreatment produced long-term reductions in both striatal PKA activity and DA content. Early amphetamine exposure also resulted in an upregulation of D(2)-like binding sites, while leaving D(1)-like binding sites unaffected. It is likely that the upregulation of D(2)-like binding sites was stimulated by the persistent decline in striatal DA levels. Although speculative, it is possible that excess striatal D(2)-like receptors were responsible for inhibiting PKA activity through actions on the cAMP signal transduction pathway. The behavioral relevance of these amphetamine-induced neurochemical changes has not yet be determined. PMID- 11120385 TI - Temporal pattern in the effect of postnatal blood lead level on intellectual development of young children. AB - To determine the temporal pattern of the effect of postnatal blood lead level on the General Cognitive Index (GCI) of the McCarthy Scales of Children's Abilities, we used data from 112 children of the Mexico City Prospective Lead Study with complete evaluations from 36 to 60 months of age at 6-month intervals. We measured blood lead level every 6 months from 6 to 54 months. We controlled for 5 min Apgar, birth weight, birth order, sex, socioeconomic level, maternal IQ, and maximum maternal educational level in a repeated measures ANCOVA using child blood lead level grouped by 6-18 month (geometric mean 10.1 microg/dl, range 3.5 37.0 microg/dl), 24-36 month (geometric mean 9.7 microg/dl, range 3.0-42.7 microg/dl), and 42-54 month (geometric mean 8.4 microg/dl, range 2.5-44.8 microg/dl) averages. There were significant interactions between the 6-18 month blood lead level and age with GCI as the endpoint and between 24-36 month blood lead level and age. The regression coefficient of blood lead at 6-18 months became more negative with age until 48 months, when the rate of decline moderated (linear polynomial contrast p=0. 047). The regression coefficient of blood lead at 24-36 months with CGI became more negative as well from 36 to 48 months but then started decreasing toward zero from 48 to 60 months (quadratic polynomial contrast p=0.019). Significant between-subjects lead effects on GCI were found for 24-36 month blood lead level at 48 months (p=0.021) and at 54 months (p=0.073). The greatest effect (at 48 months) was a 5.8-point GCI decrease with each natural log unit increase in blood lead. Significant between-subjects lead effects on GCI were found for 42-54 month blood lead level at 54 months (p=0. 040) and at 60 months (p=0.060). The effect of postnatal blood lead level on GCI reaches its maximum approximately 1-3 years later, and then becomes less evident. Four to five years of age appears to be a critical period for the manifestation of the earlier postnatal blood lead level effects. PMID- 11120386 TI - The Yugoslavia Prospective Lead Study: contributions of prenatal and postnatal lead exposure to early intelligence. AB - To investigate associations between the timing of lead (Pb) exposure on early intelligence, we examined the results of psychometric evaluations at ages 3, 4, 5, and 7 years, from 442 children whose mothers were recruited during pregnancy from a smelter town and a non-lead-exposed town in Yugoslavia. We compared the relative contribution of prenatal blood lead (BPb) with that of relative increases in BPb in either the early (0-2 years) or the later (from 2 years on) postnatal period to child intelligence measured longitudinally at ages 3 and 4 (McCarthy GCI), 5 (Wechsler Preschool and Primary Scale of Intelligence-Revised, WPPSI-R IQ), and 7 (Wechsler Intelligence Scale for Children-version III, WISC III IQ), controlling for: Home Observation for Measurement of the Environment (HOME) quality; maternal age, intelligence, education, and ethnicity; and birthweight and gender. Elevations in both prenatal and postnatal BPb were associated with small decrements in young children's intelligence. PMID- 11120387 TI - Aging unmasks adverse effects of gestational exposure to methylmercury in rats. AB - The consequences of developmental exposure to methylmercury on behavior in aged animals were investigated. Methylmercury exposure was arranged by placing 0, 0.5 or 6.4 ppm Hg in the drinking water of female rats at least 4 weeks before mating and continuing until post-natal (PN) day 16. Brain Hg concentrations in cohorts of low- and high-dose offspring were 0.5 and 9.1 ppm at birth and 0.04 and 0. 52 ppm at weaning (described in another report). Lever pressing of female offspring was maintained under a Multiple Differential Reinforcement of High Rate 9:4 Extinction schedule of food reinforcement (Mult DRH 9:4 EXT). Under the DRH 9:4 schedule, a food reinforcer was delivered when nine responses occurred within 4 s. Under the Extinction schedule, responding had no programmed consequences. No exposure-related differences in reinforcement rate under the DRH schedule or discrimination between the DRH and extinction components were apparent initially. At 950 days of age, the overall response rates of controls had shown a gradual decline over the previous 500 days to about 80% of their beginning levels, but, otherwise, most controls were healthy. A gradual decline in the reinforcement rate began to appear in low- and high-dose rats at about 500 and 800 days of age, respectively. Microanalyses of the nine-response burst maintained by the DRH schedule revealed that the lever-press duration increased, the inter-response time (IRT) was unaffected, and the time between response bursts increased. Overall, the nine-response burst remained intact as a coherent response unit. The increased time between response bursts caused the decline in reinforcement rate. All rats displayed these effects as they aged, but the mercury-exposed rats did so sooner. PMID- 11120388 TI - Interaction of ethanol with retinol and retinoic acid in RAR beta and GAP-43 expression. AB - Fetal ethanol exposure has many detrimental effects on neural development, which possibly occurs through ethanol-induced disruption of the function of vitamin A. In LAN-5 neuroblastoma cells, retinol (10(-6) M) and retinoic acid (RA; 10(-5) 10(-6) M) increased RAR beta mRNA expression. Ethanol downregulated RAR beta levels, even in the presence of retinol. RAR beta mRNA expression was decreased by ethanol in the presence of 10(-6) M RA, but not 10(-5) M RA. With cycloheximide (CX), RA still stimulated RAR beta mRNA, but the effect of ethanol was abolished. The mRNA expression of GAP-43, an important factor in neural development, increased with 10(-6) M retinol and 10(-5)-10(-9) M RA. Ethanol decreased GAP-43 mRNA expression in the presence or absence of retinol. Ethanol was without effect on GAP-43 mRNA at 10(-5) M RA, but did lower the levels at 10( 6) and 10(-7) M RA. CX prevented the effects of both RA and ethanol on GAP-43 mRNA. These studies provide support for the hypothesis that retinoid function is altered by ethanol. PMID- 11120389 TI - Dose-related differences in the distribution of cocaine in the maternal fetoplacental compartments in rats. AB - Our goals were to examine whether a high dose of cocaine to causing CNS toxic manifestations in the pregnant rats influences the delivery of cocaine to the fetus, and whether the non-placental compartments have a significant role in the distribution of cocaine to the fetal tissues. Either a low or high dose of cocaine was infused intravenously to near-term pregnant rats. Arterial blood pressure and heart rate were monitored. Cardiac output and uterine and placental blood flows were measured by using radiolabeled microspheres. Plasma and tissue samples were obtained from the mother, placenta, and fetus and analyzed for cocaine and its metabolites via capillary gas chromatography/mass spectrometry. A high dose of cocaine induced convulsions that were accompanied by increased arterial blood pressure and decreased uteroplacental blood flow. However, the distribution pattern of cocaine and metabolites in the mother and fetus were similar between the high and low dose groups. Considerable amounts of cocaine and its metabolites were in the placenta. Previously ignored non-placental tissues, such as the amnion and myometrium appear to be a significant source for cocaine accumulation in the fetus. PMID- 11120390 TI - Effects of prenatal cocaine on behavioral adaptation to chronic stress in adult rats. AB - Prenatal exposure to cocaine in rats has previously been shown to alter the behavioral and hormonal responses to acute stressors, although no work has yet examined stress adaptation in these animals in adulthood, a possibility examined in this experiment. Male and female offspring of Sprague-Dawley rat dams given 40 mg/kg/3 ml subcutaneously daily from gestational days 8-20 (C40), saline injected and pair-fed dams (PF), and non-treated dams (NT) were tested in adulthood (90 120 days). Offspring were given a 5-min open field test 24 h following the last of 1 (Acute), 9 (Chronic) or 0 (control) daily 15-min intermittent footshock sessions. Substantially more behavioral adaptation was evident in NT offspring than in C40 and PF animals. The attenuated stress adaptation seen in C40 offspring extends prior work showing altered stress responsiveness in these animals, although the PF data caution against the conclusion that this lack of stress adaptation necessarily reflects gestational exposure to cocaine per se. PMID- 11120391 TI - The neurobehavioral effects of subchronic manganese exposure in the presence and absence of pre-parkinsonism. AB - Recent studies have implicated chronic elevated exposures to environmental agents, such as metals (e.g., manganese, Mn) and pesticides, as contributors to neurological disease. In particular, there is a concern that sensitive subpopulations such as the aged may be at increased risk for the onset of neurologic disorders because elevated exposures to Mn is associated with increased incidence of parkinsonism. Here, we utilized a rat model of pre parkinsonism to investigate the effects of Mn exposure on neurotoxicity and the exacerbation of parkinsonism. A pre-parkinsonism state was induced using a unilateral intrastriatal injection of 6-hydroxydopamine (6-OHDA), followed 4 weeks later by Mn exposure (4.8 mg Mn/kgx3 intraperitoneal injections/week) for 5 weeks. Female Sprague-Dawley rats (n=44) were divided among the following treatments: (A) control, saline/vehicle; (B) Mn only; (C) 6-OHDA only; and (D) 6 OHDA+Mn. Brain Mn levels were measured by ICP-MS. Neurobehavioral function was assessed following Mn exposure using a functional observational battery (FOB) consisting of 10 neurobehavioral tests. Unilateral 6-OHDA lesions produced significant ipsilateral vs. contralateral striatal dopamine depletions (60-70%), but no measurable impairment of neurobehavioral function, thereby substantiating this pre-parkinsonism (i.e., subthreshold) model. In contrast, Mn exposure resulted in significant impairment of neurobehavioral function for eight of the 10 FOB tests. No effects of Mn exposure on striatal dopamine depletion were detected, despite the 3.4-fold increase in brain Mn levels over controls. Notably, Mn exposure in the presence of a pre-parkinsonism state significantly exacerbated the neurobehavioral impairment in the reactivity to handling (P<.049) and hopping contralateral rear limb (P<.033) FOB tests. While the persistence and Mn dose-response relationship of these neurobehavioral effects were not evaluated here, these results nonetheless suggest that chronic Mn exposure may increase the risk of neurobehavioral impairment in subpopulations that are in a pre parkinsonism state. PMID- 11120392 TI - Comparative behavioural toxicity of domoic acid and kainic acid in neonatal rats. AB - Cumulative behavioural toxicity was measured in groups of male and female rat pups (n=6/sex) at different stages of postnatal development. Dose-response curves (DRCs) for toxicity produced by domoic acid (DOM) were generated using animals on postnatal days (PND) 0, 5, 14, and 22, using a behavioural rating scale. In a subsequent experiment, DRCs for toxicity generated by either DOM or kainic acid were produced in rats at PND 8 and 14 for comparison between the two toxins. DOM was found to be a very potent neurotoxin in newborn rats and the potency of DOM progressively decreased with increasing age (interpolated ED(50)=0.12, 0.15, 0.30, and 1.06 mg/kg at PND 0, 5, 14, and 22, respectively). In addition, the patterns of behavioural expression were found to differ with age. Comparisons between DOM and kainic acid revealed that DOM was approximately six-fold more potent than kainate at both PND 8 and PND 14 and that both toxins were approximately two-fold less potent in PND 14 rats, compared to PND 8. This implies that the mechanism(s) responsible for reduced potency is/are similar between the two compounds. Consistent with previous reports, however, there were both similarities and differences in the observed patterns of behavioural toxicity produced by the two toxins at both ages. PMID- 11120393 TI - The behavioral teratogenic potential of fenbendazole: a medication for pinworm infestation. AB - Fenbendazole (FBZ) is a benzimidazole currently used for anthelmintic treatment of pinworm populations in numerous animal species although it is not currently approved for laboratory rodents in the U.S. It has received considerable interest for treating rodent populations due to its low toxicity, wide safety margin and apparent absence of gross teratogenic effects. The purpose of this study was to assess the behavioral teratogenic potential of FBZ. Pregnant rats were administered either FBZ-medicated feed at a therapeutic level or normal rat chow throughout pregnancy and gestation. FBZ had no effect on pregnancy indicators such as maternal weight gain or water consumption, number of pups born or pup birth weights. Offspring were examined in a variety of paradigms including righting reflex, negative geotaxis, running wheel activity, Morris water maze (MWM) performance and digging maze performance. FBZ offspring did show delayed righting reflex, some modest changes in locomotor activity in a running wheel and minor alterations in performance during the probe session of the MWM relative to controls. However, the effects of FBZ on behavior were subtle and many of the behaviors examined were unaffected. These results suggest that FBZ may be an effective and relatively safe anthelmintic treatment for use in breeding colonies. PMID- 11120394 TI - Drug addiction, dysregulation of reward, and allostasis. AB - This paper reviews recent developments in the neurocircuitry and neurobiology of addiction from a perspective of allostasis. A model is proposed for brain changes that occur during the development of addiction that explain the persistent vulnerability to relapse long after drug-taking has ceased. Addiction is presented as a cycle of spiralling dysregulation of brain reward systems that progressively increases, resulting in the compulsive use and loss of control over drug-taking. The development of addiction recruits different sources of reinforcement, different neuroadaptive mechanisms, and different neurochemical changes to dysregulate the brain reward system. Counteradaptive processes such as opponent-process that are part of normal homeostatic limitation of reward function fail to return within the normal homeostatic range and are hypothesized to form an allostatic state. Allostasis from the addiction perspective is defined as the process of maintaining apparent reward function stability by changes in brain reward mechanisms. The allostatic state represents a chronic deviation of reward set point and is fueled not only by dysregulation of reward circuits per se, but also by the activation of brain and hormonal stress responses. The manifestation of this allostatic state as compulsive drug-taking and loss of control over drug-taking is hypothesized to be expressed through activation of brain circuits involved in compulsive behavior such as the cortico-striatal thalamic loop. The view that addiction is the pathology that results from an allostatic mechanism using the circuits established for natural rewards provides a realistic approach to identifying the neurobiological factors that produce vulnerability to addiction and relapse. PMID- 11120395 TI - Lowered serum dipeptidyl peptidase IV activity is associated with depressive symptoms and cytokine production in cancer patients receiving interleukin-2-based immunotherapy. AB - There is some evidence that treatment with interleukin-2 (IL-2) and interferon alpha (IFNalpha) frequently induces depressive symptoms and activation of the inflammatory response system (IRS). There is evidence that major depression is accompanied by lowered serum activity of dipeptidyl peptidase IV (DPP IV; EC 3.4.14.5), a membrane-bound serine protease which catalyses the cleavage of some cytokines and neuro-active peptides and which modulates T cell activation and the production of cytokines, such as IL-2. This study was carried out to examine the effects of immunochemotherapy with IL-2 and IFNalpha, alone and together, in cancer patients on serum DPP IV activity in relation to changes in depressive symptoms and the IRS. The Montgomery and Asberg Rating Scale (MADRS), serum DPP IV activity, and the serum IL-6, and IL-2 receptor (IL-2R) concentrations were measured in 26 patients with metastatic cancers before and three and five days after treatment with IL-2 and IFNalpha, alone or together. Treatment with IL-2 with or without IFNalpha significantly suppressed serum DPP IV activity. The MADRS scores were significantly elevated by treatment with IL-2 with or without IFNalpha, but not IFNalpha alone. The immunochemotherapy-induced decreases in serum DPP IV were significantly and inversely correlated with the increases in the MADRS. Treatment with IL-2 alone or combined with IFNalpha also elevated serum IL-6 and IL-2R. There were significant and inverse correlations between the immuchemotherapy-induced decreases in serum DPP IV and the elevations in serum IL 6 or IL-2R. In conclusion, treatment with IL-2/IFNalpha decreases serum DPP IV activity within 3-5 days and the immunochemotherapy-induced decreases in serum DPP IV activity are significantly and inversely related to treatment-induced increases in severity of depression and signs of activation of the IRS. PMID- 11120396 TI - Endogenous 5-HT tonically inhibits spontaneous firing activity of dorsal hippocampus CA1 pyramidal neurons through stimulation of 5-HT(1A) receptors in quiet awake rats: in vivo electrophysiological evidence. AB - The present study was performed to examine an overall effect of endogenous serotonin (5-HT) on the spontaneous firing activity of the dorsal hippocampus CA1 pyramidal neurons in quiet awake rats. A selective 5-HT(1A) antagonist N-[2-[4-(2 methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohe xanecarboxamide (WAY 100635: 0.03-0.2 mg/kg, s.c.) significantly increased the firing activity. A depletion of 5-HT with parachlorophenylalanine (PCPA: 500 mg/kg/day x 3 days) completely abolished this increasing effect of WAY-100635. The baseline spike frequency of the PCPA-treated rats (3.90 +/- 0.39 Hz) was significantly higher than that of the vehicle-treated rats (2.09 +/- 0.19 Hz). A 5-HT(2A) antagonist ritanserin (1 mg/kg, i.p.) and a 5-HT(3/4) antagonist 2-methoxy-4-amino-5-chloro benzoic acid 2-(diethylamino) ethyl ester (SDZ-205557: 3 mg/kg, s.c.) did not modify the firing activity and the increasing effect of WAY-100635. These results suggest that, in quiet awake rats, endogenous 5-HT would tonically inhibit the spontaneous firing activity of the CA1 pyramidal neurons mainly through stimulating 5-HT(1A) receptors. PMID- 11120397 TI - Opiate withdrawal-induced fos immunoreactivity in the rat extended amygdala parallels the development of conditioned place aversion. AB - Low doses of naloxone have been shown to affect the motivational aspects of opiate withdrawal in morphine-dependent rats. Conditioned place aversion to opiate withdrawal is one of the most sensitive of motivational indices of opiate withdrawal and is thought to be mediated by the basal forebrain. Expression of the transcription factor Fos is known to increase during opiate withdrawal, but its presence during low-dose antagonist-precipitated withdrawal has not previously been established. In order to determine if there is a relationship between withdrawal-induced neuronal activity and conditioned place aversion, immunocytochemical localization of Fos was examined in the basal forebrain of opiate-dependent animals receiving one of several doses of naloxone (0, 3.25, 7.5, 15, 30, or 1000 microg/kg). In separate groups of opiate-dependent animals, naloxone doses of 3.25 - 30 microg/kg were paired with a specific chamber in a single-pairing conditioned place aversion paradigm. Significant increases in both immunocytochemical detection of Fos and conditioned place aversion were seen at doses >/= 7.5 microg/kg. The shell of the nucleus accumbens and central nucleus of the amygdala were most sensitive to low doses, thus supporting the hypothesis that the extended amygdala plays a role in opiate-induced condition place aversion. PMID- 11120398 TI - Dose response of arginine vasopressin to the CCK-B agonist pentagastrin. AB - Cholecystokinin (CCK) is a peptide neurotransmitter that modulates hypothalamic pituitary-adrenal (HPA) axis activity and may be involved in fear or anxiety states. Arginine vasopressin (AVP) also modulates HPA axis activity and may play a role in fear conditioning. Few human studies have examined interactions between CCK and AVP systems. To explore relationships between CCK-B receptor activation, the HPA axis response, and AVP release, a dose-response study using the CCK-B receptor agonist pentagastrin was conducted. Adrenocorticotropin (ACTH) and cortisol results have been previously reported and AVP data is presented here. Thirty-five healthy subjects were randomly assigned to receive placebo, or 0.2, 0.4, 0.6, or 0.8 microg/kg doses of pentagastrin. AVP release appeared to increase with increasing doses of the CCK-B agonist. However, this may have been due to a greater percentage of subjects releasing AVP in the higher dose groups, rather than a direct effect of dose on magnitude of response. AVP and ACTH responses were correlated, but AVP response alone could not account for the magnitude of the ACTH response. AVP release was significantly correlated with anxiety symptom responses. These findings suggest a possible role for the CCK-B receptor in AVP release, which may be at least partially separate from its role in modulation of the HPA axis. Further work is needed to determine whether these are physiologically meaningful interactions and to determine their functional implications. PMID- 11120400 TI - Changed concentrations of tachykinins and neuropeptide Y in brain of a rat model of depression: lithium treatment normalizes tachykinins. AB - Lithium's therapeutic mechanism of action is unknown. In lithium-treated normal rats, increased striatal concentrations of neurokinin A (NKA)-like immunoreactivity (LI), substance P (SP-LI) and neuropeptide Y (NPY-LI) have been reported. To investigate whether these effects might be of therapeutic relevance, Flinders Sensitive Line rats (FSL), an animal model of depression, and control Flinders Resistant Line (FRL) rats were during a 6-week period fed chow to which either lithium or vehicle was admixed. Following sacrifice, the peptides were extracted from dissected brain regions and measured by radioimmunoassay. NKA-LI and SP-LI were markedly decreased in striatum and increased in frontal cortex in FSL compared to control FRL animals. Lithium treatment abolished these differences. Basal concentrations of NPY-LI were decreased in hippocampus of FSL rats, but unaffected by lithium. The present study suggests that changed tachykinins and NPY may underlie the characterized depressive-like phenotype of the FSL rats. It is hypothesized that altering tachykinin peptidergic neurotransmission in striatum and frontal cortex constitutes a mechanism of action of lithium and that such a mechanism might be of therapeutic relevance. PMID- 11120401 TI - Variability of erythrocyte and serum lithium levels correlates with therapist treatment adherence efforts and maintenance treatment outcome. AB - This study investigated the relationship between psychotherapeutic interventions and pharmacologic measures of pharmacotherapy treatment adherence in patients with bipolar I disorder, as well as the relationship between these measures and treatment outcome. Subjects were participating in an ongoing maintenance treatment study. Audiotaped therapy sessions were rated for frequency of psychotherapeutic interventions related to pharmacotherapy treatment adherence. Pharmacologic measures of medication adherence were compared to the tape ratings as well as to treatment outcome. Variability in log erythrocyte (RBC) lithium-a marker of probable nonadherence to the pharmacotherapy regimen-for individual patients correlated significantly with treatment adherence interventions scale ratings. This marker of nonadherence was significantly related to maintenance treatment outcome, as was variability of the serum lithium level/dose (L/D) ratio; however, no relationship was found between treatment adherence interventions scale ratings and outcome. PMID- 11120399 TI - Neurotensin gene expression and behavioral responses following administration of psychostimulants and antipsychotic drugs in dopamine D(3) receptor deficient mice. AB - Exposure to psychostimulants and antipsychotics increases neurotensin (NT) gene expression in the striatum and nucleus accumbens. To investigate the contribution of D(3) receptors to these effects we used mice with targeted disruption of the D(3) receptor gene. Basal NT mRNA expression was similar in D(3) receptor mutant mice and wild-type animals. Acute administration of haloperidol increased NT gene expression in the striatum in D(3)+/+, D(3)+/- and D(3)-/- mice. Similarly, acute cocaine and amphetamine induced NT mRNA expression in the nucleus accumbens shell and olfactory tubercle to a comparable extent in D(3) mutants and wild-type mice. Daily injection of cocaine for seven days increased NT mRNA in a restricted population of neurons in the dorsomedial caudal striatum of D(3)+/+ mice, but not in D(3)-/- and D(3)+/- animals. No differences were observed between D(3) receptor mutant mice and wild-type littermates in the locomotor activity and stereotyped behaviors induced by repeated cocaine administration. These findings demonstrate that dopamine D(3) receptors are not necessary for the acute NT mRNA response to drugs of abuse and antipsychotics but appear to play a role in the regulation of NT gene induction in striatal neurons after repeated cocaine. In addition, our results indicate that the acute locomotor response to cocaine and development of psychostimulant-induced behavioral sensitization do not require functional D(3) receptors. PMID- 11120402 TI - Delta(9)-tetrahydrocannabinol and synthetic cannabinoids prevent emesis produced by the cannabinoid CB(1) receptor antagonist/inverse agonist SR 141716A. AB - There is substantial clinical evidence that Delta(9)-tetrahydrocannabinol (Delta(9)-THC) and its synthetic analogs (nabilone and levonantradol) can prevent emesis in cancer patients receiving chemotherapy. Limited available animal studies also support the antiemetic potential of these cannabinoids. The present study investigates the mechanism of antiemetic action of cannabinoids in an established animal model of emesis, the least shew (Cryptotis parva). Since cannabinoid agonists prevent emesis, it was hypothesized that blockade of either the cannabinoid CB(1) receptor or the cannabinoid CB(2) receptor would induce vomiting. Thus, the emetic potential of SR 141716A (CB(1) receptor antagonist) or SR 144528 (CB(2) receptor antagonist) was investigated. Both intraperitoneal (0, 1, 2.5, 5, 10 and 20 mg/kg, n = 7-15 per group) and subcutaneous (0, 10, 20 and 40 mg/kg, n = 6-9 per group) administration of SR 141716A caused emesis (ED(50) = 5.52 +/- 1.23 and 20.2 +/- 1.02 mg/kg, respectively) in the least shrew in a dose dependent manner. Indeed, both the frequency of emesis and the percentage of animals vomiting increased with increasing doses of SR 141716A. Significant effects were seen at the 10- and 20-mg/kg doses for the IP route, while only the 40-mg/kg dose produced significant emesis via the SC route. The CB(2) antagonist failed to produce emesis via either route of administration. SR 141716A at an IP dose of 20 mg/kg was used to induce emesis for drug interaction studies. Thus, varying doses of three different classes of cannabinoid agonists [CP 55, 940 (0, 0.1, 0.5 and 1 mg/kg), WIN 55, 212-2 (0, 1, 5 and 10 mg/kg), and Delta(9)-THC (0, 5, 10 and 20 mg/kg)], were administered IP to different groups of shrews 10 min prior to SR 141716A injection. The frequency of emesis was recorded for 30 min following the administration of SR 141716A. The order of potency for redcing both the frequency of emesis and the percentage of shrews vomiting was CP 55, 940 > WIN 55, 212-2 > Delta(9)-THC which is consistent with an action on the CB(1) receptor. These results suggest that the antiemetic activity of Delta(9)-THC and its synthetic analogs reside in their ability to stimulate the cannabinoid CB(1) receptor. Furthermore, the antiemetic potency of CP 55, 940 is 45 times greater than Delta(9)-THC. On the other hand, blockade of CB(1) receptors can induce vomiting, which implicates an important role for endogenous cannabinoids in emetic circuits. PMID- 11120403 TI - Spatial registration of digital brain atlases based on fuzzy set theory. AB - We present a semiautomatic method based on fuzzy set theory for adjusting a computerized brain atlas to magnetic resonance images (MRIs) of the human cerebral cortex. The atlas was registered to three-dimensional MRI data sets of 10 healthy volunteers. After a global matching using the external contour of the brain, several local procedures were performed regarding selected primary furrows and cytoarchitectonic areas. The final transformation matrix was calculated with respect to these anatomical structures and to their local matrices. Evaluation revealed an increase in accuracy as expressed by a reduction of the visible mismatch with respect to the registration of cortical and subcortical brain structures. PMID- 11120404 TI - A prototype 3D CT extension for radiotherapy simulators. AB - In this paper we present a report on our custom 3D CT extension capable of producing fully 3D tomographic studies from conventional radiotherapy simulator cone-beam fluoro output. The extension consists of a common PC system on which a proprietary software toolkit provides the appropriate environment for reconstruction and acquisition of cone-beam data provided by commercial simulator fluoro video chains. The extension may compare favorably with CT options offered by simulator manufacturers: in particular, multi-slice single-scan reconstruction seems to be achievable while the current commercial solutions are based on single slice single-scan. Radiotherapy applications include on-line treatment planning, patient treatment verification and registration. PMID- 11120405 TI - Skull base tumors: evaluation with contrast-enhanced MP-RAGE sequence. AB - The purpose of this study is to assess the value of contrast-enhanced MP-RAGE sequence for evaluation of skull base tumors. A total of 36 patients were prospectively evaluated. MP-RAGE showed relatively higher CNRs than other sequences, and there was a significant difference of CNR between the MP-RAGE and contrast T1-weighted SE images when fat tissue was chosen as the background. MP RAGE was significantly superior to other sequences in the diagnosis of the extent of tumors. Contrast-enhanced MP-RAGE sequence is useful in evaluation of skull base tumors because of its higher contrast, higher spatial resolution, multiplanar capability, and suppression of the fat signal. PMID- 11120406 TI - An empirical parameter selection method for endocardial border identification algorithms. AB - Identification of the cardiac borders is a basic goal in the analysis of echocardiographic images. For this reason there has been a great deal of interest in automatic border identification methods. Numerous studies have been published, describing a wide variety of proposed methods. In spite of the large number of published studies, however, the relative merits of the various methods remain unclear. Furthermore, the optimal configuration of each method is in most cases unknown. One of the reasons for this is the lack of a well-defined, rigorous method for evaluating and comparing alternative algorithms and/or configurations. In this article we describe a systematic, empirical procedure for evaluating border identification algorithms. The method is based on a strict metrical measure of border error, and a set of test images which reflect the heterogeneity of typical clinical images. The methodology can be used to compare alternative analysis methods, or to determine the optimal configuration for any particular analysis method. We then apply this methodology to the problem of determining parameter values for several of the most commonly used analysis methods. PMID- 11120407 TI - Accurate segmentation of the breast region from digitized mammograms. AB - The segmentation of a digital mammogram into the breast region and the background is a necessary prerequisite in computer-assisted diagnosis of mammograms. By the exclusion of the background region, the accuracy of the analysis is increased and the running-time is decreased. The algorithm which segments the breast region from the background should be fully automated and give correct results for all kinds of digitized mammograms, including low-quality images. In this paper we present such an algorithm based on histogram thresholding, morphological filtering and contour modeling. Quantitative test results indicate that the computed boundary follows the estimated boundary accurately. PMID- 11120408 TI - Mesenchymal hamartoma of the liver. AB - We present a multiseptated mesenchymal hamartoma of the liver in a 10-year-old male patient, a rare benign tumor of childhood. The characteristic ultrasound and CT appearances of this unusual tumor are reviewed. A single septal calcification associated with this tumor was demonstrated, an association which has not previously been reported. The differential diagnosis for cystic liver lesions is discussed in detail. PMID- 11120409 TI - Abdominal circumference vs. estimated weight to predict large for gestational age birth weight in diabetic pregnancy. AB - Early third trimester fetal abdominal circumference and sonographic fetal weight estimates were compared to predict large for gestational age birth weight in diabetic pregnancy. Both parameters have similar sensitivity, specificity, and predictive values. However, the optimal percentile cutoff values differ. Choice of birth weight standard significantly influences test characteristics. Negative prediction of large birth weight is more accurate than positive prediction. At third trimester sonography with maternal diabetes, the abdominal circumference percentile is potentially useful and should be routinely reported. PMID- 11120410 TI - Transient colocolic intussusception. AB - We present the CT findings of a transient colocolic intussusception, related to an underlying colonic tumor, but remote from it. The resolving nature of intussusception was clearly demonstrated on delayed images and may explain the characteristic chronic clinical symptoms of intussusception in adult. PMID- 11120411 TI - Uterine arteriovenous malformations: the role of intravenous 'dual-phase' CT angiography. AB - The authors describe the use of dual-phase intravenous CT angiography of the pelvis in two female patients, who presented with ongoing excessive vaginal bleeding, to demonstrate large adnexal and uterine arteriovenous malformations (AVMs). Power Doppler was used as the initial modality to diagnose the AVMs. CT angiography, along with 3-D rendering in the form of maximum intensity projections and shaded surface display, were especially useful for anatomical conceptualization to the gynecologist. This greatly helped in the subsequent management in the form of therapeutic embolization in both patients by reducing the time, radiation dose, and contrast required for the procedure. Subsequent surgery, which was required in both patients (due to failed embolization), was also greatly aided by the demonstration of the exact extent of the AVMs on axial CT images. Thus, CT angiography emerged as an impressive non-invasive imaging modality for the complete evaluation and management of the uterine AVMs. PMID- 11120412 TI - MR imaging of a parosteal lipoma. AB - A case of a parosteal lipoma of the thigh is presented. Very little is known of the enhancement features of this tumor. In this case, mild enhancement of tissue in the region of the pedicle between reactive cortical bone hyperostosis and the lipomatous mass corresponded to fibrous tissue that outlined the hyperostotic reactive bone. PMID- 11120413 TI - Osteoid osteoma: MR imaging revisited. AB - To assess and compare with computed tomography (CT) the performance of magnetic resonance (MR) imaging in the detection of osteoid osteoma, and determine the features of this lesion on MR imaging. The prospective MR imaging and CT diagnosis of osteoid osteoma was determined from original radiology reports. MR images were assessed retrospectively with regard to the location and signal intensity of the nidus and surrounding bone marrow and soft tissue edema. These findings were correlated with the age of the patient, duration of symptoms, and drug therapy. Ten patients with histologically proven osteoid osteoma who underwent MR imaging were reviewed. All 10 lesions were correctly diagnosed at the time of MR imaging. None of the lesions was intracortical. Nine lesions were intra-articular. Two out of five patients with extracortical lesions had false negative CT preceding the MR study. Signal intensity of the nidus, marrow, and soft tissue edema on MR imaging were variable. Perinidal edema was most pronounced in younger patients and had no apparent relation to drug therapy. MR imaging reliably demonstrates the nidus of an osteoid osteoma, which has a variable appearance related to its position relative to the cortex of the bone. A predominance of cancellous osteoid osteomas are encountered in patients referred for MR imaging. Marrow edema in the vicinity of the lesion improves the conspicuity of the nidus. CT may fail to diagnose osteoid osteoma when the nidus is in a cancellous location, due to the lack of perinidal density alteration. PMID- 11120414 TI - MR findings of struma ovarii. AB - This study was performed to characterize MR findings of struma ovarii. In 10 patients, T1- and fast spin echo T2-weighted MR images were obtained in the axial, coronal, and sagittal planes using 1.5 T MR units, and they were retrospectively evaluated for the site, size, components, signal intensity, and contrast enhancement.MR images showed a unilateral complex mass with a multilobulated surface and thickened septa, corresponding pathologically to thyroid follicles and the stroma. Cystic portions had variable signal intensities on T1- and T2-weighted images. The contents of cystic components showing low signal intensities both on T1- and T2-weighted images were viscid gelatinous materials (n=4). Solid portions were relatively well-enhanced. In conclusion, struma ovarii has some characteristic MR appearance of a multilobulated complex mass with thickened septa, multiple cysts of variable signal intensities, and enhancing solid components. PMID- 11120415 TI - Leiomyoma of the ovary mimicking mucinous cystadenoma. AB - We present a case of ovarian leiomyoma of a 46-year-old woman with a history of a palpable lower abdominal mass. A multiloculated multiseptated mainly cystic mass in the left adnexa on computed tomography (CT) was initially considered to be an ovarian mucinous cystadenoma. This mass, however, was proved to be a left ovarian vascular leiomyoma on the surgical pathology. PMID- 11120416 TI - Foci of signal intensities different from fat in well-differentiated liposarcoma and lipoma: correlation between MR and histological findings. AB - To investigate the histological features of foci, which showed signal intensities different from fat by magnetic resonance (MR) imagings in well-differentiated lipoma-like liposarcomas and a case of lipoma, a retrospective review of these lipomatous tumors was performed to correlate MR and histological findings. Microscopic findings revealed that these foci were also composed of lipomatous tissue. Well-differentiated liposarcoma and benign lipoma associated with such foci should be differentiated from dedifferentiated liposarcoma based on the histological findings. PMID- 11120417 TI - MR features of a case of chronic expanding hematoma. AB - We describe a patient with chronic expanding hematoma that was pathologically confirmed by examination of the resected specimen. It has increased gradually in the right lower lung field during a period of 10 years without symptoms. The MR T2-weighted image was useful to establish a diagnosis of chronic expanding hematoma, because of the mosaic of various signal intensities we named the "mosaic sign". PMID- 11120418 TI - Abstracts PMID- 11120419 TI - Improvement in cognition associated with novel antipsychotic drugs: a direct drug effect or reduction of EPS? AB - BACKGROUND: Administration of novel, versus classic, antipsychotic agents to patients suffering from psychosis is associated both with moderately better scores on cognitive tests, and with fewer extrapyramidal symptoms (EPS). Because improved motor functioning may enable better performance on some components of cognitive test batteries, and because the advantages of the novel antipsychotics on cognitive performance are not very large, it is sometimes difficult to discern if improvement in a given cognitive task is due to a direct effect of the novel antipsychotic drug, or is secondary to the novel drug's decreased propensity to induce EPS. In an attempt to distinguish between these two possibilities, the present study examined the ability of patients suffering from schizophrenia receiving classic, versus novel antipsychotics, to perform a computerized visuo motor test (VMT). VMT assesses planning capabilities, attention and executive functions known to be impaired in schizophrenia, which are suggested to be affected by novel antipsychotics. METHODS: Seventy-six patients suffering from schizophrenia or schizophreniform disorder, receiving haloperidol (23 patients, mean dose 10.01+/-6.1mg/day), olanzapine (26 patients, mean dose 10.56 +/- 4.9 mg/day) or risperidone (27 patients, mean dose 4.35 +/- 1.7 mg/day) were assessed for EPS using the parkinsonian subscale of the Extrapyramidal Symptom Rating Subscale (ESRS), and with the VMT. RESULTS: Cognitive functioning as measured by the VMT was better for patients receiving risperidone or olanzapine, compared with those receiving haloperidol (F=6.636, df=2,67, P=0. 002), while the patients receiving haloperidol or risperidone suffered from more severe EPS compared with the patients receiving olanzapine (F=3.996, df=2,71, P=0.023). DISCUSSION: Although the patients receiving risperidone suffered from EPS similar in severity to the EPS of the patients receiving haloperidol, their performance on a task involving visuo-motor and attentional skills was similar to that of the patients receiving olanzapine. This finding implies that there is a dissociation between the antipsychotic drug's ability to affect cognitive functioning, and EPS. This dissociation indirectly suggests that the advantages offered by novel antipsychotics on cognitive performance are a direct effect, rather than being entirely mediated by improved movement abilities. PMID- 11120420 TI - Comparison of cost, dosage and clinical preference for risperidone and olanzapine. AB - BACKGROUND: Because risperidone and olanzapine have similar efficacy and tolerability in the treatment of schizophrenia, costs, physician experience, and preference become relevant considerations in making treatment decisions. The purpose of this paper is to compare daily treatment costs of risperidone and olanzapine, and to examine psychiatrists' clinical preferences. METHOD: Dosage information was obtained from a national Ministry of Health registry and a national survey of psychiatrists. In addition, psychiatrists' clinical preference of antipsychotic medication and dosage for patient subtypes were examined by the national survey. RESULTS: Data from the registry and national survey estimated the mean daily dose of risperidone to be one-third that of olanzapine, irrespective of patient subtype. Taking into account drug costs and dosage requirements, the average daily retail price was US $6.85 for risperidone and US $13.60 for olanzapine. Psychiatrists preferred risperidone for first-episode psychosis and elderly psychosis, and olanzapine for patients sensitive to EPS. They rated the drugs equally effective on positive and negative symptoms, for chronic patients, for treatment-refractory patients and relapse prevention. CONCLUSIONS: Risperidone has a substantial cost advantage over olanzapine, and was preferred by psychiatrists for more indications. PMID- 11120422 TI - Reemergence of tardive dyskinesia after discontinuation of clozapine treatment. AB - Tardive dyskinesia (TD) continues to be a significant health problem and a serious limitation to neuroleptic medication treatment. Clozapine treatment may reduce the severity of TD but it is unclear wether the medication temporarily suppresses symptoms or leads to a sustain resolution of the disorder. Herein we describe two cases with severe TD which clozapine had to be discontinued. These cases suggest that clozapine provides a temporary suppression of TD rather than a permanent resolution of the disorder. PMID- 11120421 TI - Risperidone versus haloperidol in psychotic patients with disturbing neuroleptic induced extrapyramidal symptoms: a double-blind, multi-center trial. AB - BACKGROUND: A randomized, double-blind, multi-center trial was started to compare the severity of extrapyramidal symptoms (EPS) during risperidone and haloperidol treatment in schizophrenic patients who had disturbing EPS during their previous neuroleptic treatment. Additional objectives of this trial were comparing the antipsychotic effectiveness of the two treatments and the use of antiparkinsonian medication. METHODS: Effects of flexible doses of risperidone and haloperidol were compared in 77 psychotic patients (83% with chronic schizophrenia) with disturbing neuroleptic-induced EPS (risperidone 40 patients, haloperidol 37). The trial was completed by 47 patients: 25 in the risperidone group (12 women, 13 men), and 22 in the haloperidol group (10 women, 12 men). RESULTS: An adequate antipsychotic effect was obtained in most patients by both treatments. The primary aim of this trial was comparing parkinsonism measured with the extrapyramidal syndrome rating scale (ESRS) during treatment with risperidone and haloperidol. Two primary parameters were selected: the change from baseline to the worst score during treatment of ESRS II (parkinsonism) and ESRS VI (clinical global impression of severity of parkinsonism). The CGI of severity of parkinsonism was better with risperidone (P=0.025), while the parkinsonism total score tended to be better with risperidone (P<0. 10). Before the double-blind treatment, 34 (of the 77) had used antiparkinson medication (risperidone 18, haloperidol 16). During the double-blind treatment phase, 21 patients had used antiparkinson medication (risperidone 11, haloperidol 10). The larger reduction of parkinsonism in the risperidone group was not due to a difference in the use of anti-parkinsonian medication. CONCLUSIONS: In this group of schizophrenic patients with disturbing EPS during previous neuroleptic treatment, a stronger reduction of parkinsonism was observed with risperidone than with haloperidol. PMID- 11120423 TI - Zuclopenthixol acetate in psychiatric emergencies: looking for evidence from clinical trials. AB - BACKGROUND: Case series and reviews have suggested the effectiveness of zuclopenthixol acetate in the acute management of disturbed behaviour caused by serious mental illnesses. This review investigates the trial-based evidence for these suggestions. METHODS: All randomized clinical trials comparing zuclopenthixol acetate to other 'standard' treatments for the acute management of those with serious mental illnesses were identified and, if possible, their results summated. RESULTS: Six trials were identified. All had methodological problems and one did not meet the minimal methodological inclusion criteria. The summary data do not demonstrate that zuclopenthixol acetate is better than 'standard care' for altering behaviour, decreasing the need for supplementary medication, avoiding side-effects, or postponing early discharge against medical advice. One trial, however, presented data that suggested an earlier, more intense level of sedation. CONCLUSIONS: Recommendations of reviews and open studies for the use of zuclopenthixol acetate in preference to 'standard' treatments in the psychiatric emergency are not supported by evidence from randomized controlled trials. PMID- 11120424 TI - Effects of smoking during antipsychotic withdrawal in patients with chronic schizophrenia. AB - A number of studies have shown that patients with schizophrenia smoke more than other psychiatric patients and more than the general population. Also, medicated schizophrenics who smoke present more positive symptoms of schizophrenia than non smokers. The objective of the present study was to assess the effect of smoking on ratings of psychopathology for 30 days following discontinuation of antipsychotic medication. The subjects were 101 treatment-resistant patients with schizophrenia who had been admitted to the inpatient service of Neuroscience Research Hospital (NRH), National Institute of Mental Health, between 1982 and 1994 to undergo studies involving discontinuation of antipsychotic medication. Patients were rated independently on a daily basis on the 22-item Psychiatric Symptom Assessment Scale (PSAS), an extended version of the Brief Psychiatric Rating Scale (BPRS). At baseline, ratings for Verbal Positive, Paranoia and Loss of Function were higher in smokers (n=65) than non-smokers (n=36), but a statistically significant difference was observed only for the Verbal Positive cluster. Analysis by gender revealed that male non-smokers had the lowest psychopathology ratings at baseline. There were no differences in Anxiety/depression, Behavior Positive, Deficit Symptoms or Mannerisms (a measure for abnormal involuntary movements). Following medication discontinuation, repeated-measure analysis demonstrated a 'time' effect for all the variables studied and a 'group' (smokers vs. non-smokers) effect for Verbal Positive, Paranoia, and Loss of Function. Post-hoc comparisons at individual time points showed significantly higher ratings for smokers at week 1 for Paranoia. No differences were observed at later time points. In conclusion, at baseline, smokers had more positive symptoms and were apparently more functionally impaired than non-smokers. This difference was no longer evident after a 30 day medication discontinuation period. PMID- 11120425 TI - Antipsychotic medication and smoking prevalence in acutely hospitalized patients with chronic schizophrenia. AB - The atypical antipsychotic, clozapine, has been reported to reduce smoking in schizophrenic patients. We sought to determine whether other atypical antipsychotics would also be associated with a decreased prevalence of smoking in this population. Data were obtained from three groups of chronic, hospitalized, schizophrenic patients, receiving either a typical antipsychotic (n=15), clozapine (n=6), or another atypical antipsychotic (n=18). In addition to smoking prevalence, the groups were compared with regard to demographics (age, education), medication (doses, duration of treatment, side-effects), clinical (diagnosis, duration of illness) and behavioral (Wide-Range Achievement Test, Wechsler Adult Intelligence Scale) variables. Smoking prevalence differed significantly among the three groups (P<0.001). Clozapine was associated with a significantly lower incidence of smoking than either typical drugs (P<0.003) or other atypical antipsychotics (P=0. 042). The groups did not differ on demographic or other medication variables or on any of several behavioral measures. However, a diagnosis of paranoid schizophrenia was also significantly correlated with smoking (P<0.01), but not with medication class. Although the cause is still unknown, these results are consistent with reports that clozapine reduces smoking and provide new data on smoking prevalence associated with other atypical agents. PMID- 11120426 TI - Practice-related improvement in information processing with novel antipsychotic treatment. AB - BACKGROUND: Attentional deficits are prominent in schizophrenia, and skill learning is impaired. Novel antipsychotic treatment has been reported to improve certain cognitive skills in schizophrenic patients, but no information is yet available about the effect of newer medications on skill learning. METHODS: Clinically stable patients with schizophrenia (n=16) and chronically hospitalized inpatients (n=8) were recruited while receiving conventional antipsychotic treatment. Subjects were tested at baseline on a visual continuous performance test (CPT), performed alone and simultaneously with an auditory CPT. Normal controls (n=8) were also tested at baseline. The inpatients and half of the outpatients were switched to treatment with risperidone. All patients then performed the visual CPT on a daily basis and performed the dual tasks once per week, for 4weeks. RESULTS: Patients who remained on conventional medications did not improve in their performance despite the extensive practice on the test. Both chronic and stable patients receiving risperidone treatment manifested a statistically significant (P<0.05) improvement from baseline on both single and dual-task visual CPT. Stable outpatients performed significantly better at the end of the protocol than the normal controls performance at baseline (P<0.05). IMPLICATIONS: These results suggest that practice-related improvements in the performance of information processing tests are enhanced by novel antipsychotic medications. Although the specific biological mechanism of this effect is not yet known, the results may suggest that use of newer medications will enhance skill development and perhaps facilitate rehabilitation of patients with schizophrenia. PMID- 11120427 TI - A new look at reaction time in schizophrenia. AB - Schizophrenia patients demonstrate impaired manual reaction time (RT), but not saccadic RT, when given traditional tasks. To determine whether the manual RT time impairment could be eliminated by providing imperative stimuli to the finger (thus providing stimulus-response compatibility), we tested 28 chronic schizophrenic patients on finger-lift RT to visual (VIS), tactile plus visual (TAC+VIS), and auditory plus tactile plus visual (AUD+TAC+VIS) stimuli. The patients (a) were significantly slower than controls (n=28) in all three tasks, (b) showed bimodality, with 43% of patients having means and variances nearly identical to control values, and (c) had RTs significantly closer to control values in the TAC+VIS and AUD+TAC+VIS tasks than in the VIS task. The inability to normalize finger-lift RT in schizophrenia represents a genuine slowing of this response system regardless of stimulus-response compatibility. We consider other possible explanations for the differences between manual and saccadic RT, including the notion that excess processing capacity for saccadic RT may be masking possible deficits in that system. PMID- 11120428 TI - Schizophrenia patients demonstrate a dissociation on declarative and non declarative memory tests. AB - Declarative memory refers to the recall and recognition of factual information. In contrast, non-declarative memory entails a facilitation of memory based on prior exposure and is typically assessed with priming and perceptual-motor sequencing tasks. In this study, schizophrenia patients were compared to normal comparison subjects on two computerized memory tasks: the Word-stem Priming Test (n=30) and the Pattern Sequence Learning Test (n=20). Word-stem Priming includes recall, recognition (declarative) and priming (non-declarative) components of memory. The schizophrenia patients demonstrated an impaired performance on recall of words with relative improvement during the recognition portion of the test. Furthermore, they performed normally on the priming portion of the test. Thus, on tests of declarative memory, the patients had retrieval deficits with intact performance on the non-declarative memory component. The Pattern Sequence Learning Test utilizes a serial reaction time paradigm to assess non-declarative memory. The schizophrenia patients' serial reaction time was significantly slower than that of comparison subjects. However, the patients' rate of acquisition was not different from the normal comparison group. The data suggest that patients with schizophrenia process more slowly than normal, but have an intact non declarative memory. The schizophrenia patients' dissociation on declarative vs. non-declarative memory tests is discussed in terms of possible underlying structural impairment. PMID- 11120429 TI - Event-related potentials in a working-memory task in schizophrenics and controls. AB - Event-related potentials (ERPs) were recorded from 65 channels in 12 schizophrenics and 12 age- and sex-matched controls during a delayed matching-to sample design with variation of working-memory (WM) challenge: following a 500 ms visual sample stimulus (called S1, two diamonds varying in size, rotation angle and vertical position), the same pattern was either presented throughout a 6s retention interval (no challenge) or a diamond pattern differing from the first one in at least one dimension was presented during this interval (WM challenge). The 500 ms matching stimulus (called S2) comprised one diamond, which had to be matched for identity to either the left or the right diamond of the sample stimulus. The topographical distribution of ERPs during an interval of 500 ms after S1-onset, 5s of the retention interval, a 500 ms-interval preceding the S2, and a 1s postimperative interval were evaluated. No WM challenge during the retention interval induced a right-posterior accentuation of the slow negative potentials in either group, while WM challenge evoked a tendency for left hemispheric negativity in controls, but not in patients. Patients exhibited a postimperative negative variation (PINV) with left-anterior focus irrespective of the preceding WM challenge, while in controls, the left-anterior PINV was found only following WM challenge. In schizophrenic patients the lack of a left anterior accentuation of negative ERPs under WM challenge might be related to WM dysfunction, and the condition-independent PINV might be considered either the consequence of this dysfunction or indication of processes related more to the diagnoses than to WM-challenge and -dysfunction. PMID- 11120430 TI - Memory for temporal order in patients with schizophrenia. AB - There is some evidence that memory for temporal order is a process that may be impaired independently of other forms of memory. For example, patients with Korsakoff's syndrome have been shown to have poorer temporal-order memory than other amnesic patients, despite item memory being equivalent. Patients with schizophrenia have been reported to have a variety of memory problems, although memory for the order of events has not been examined very frequently. In this study, we tested memory for temporal order in patients with schizophrenia and in control subjects. Subjects were presented with two lists of 15 words (at two different times) and were later asked to reproduce the order of each list from a random array of the words. In both versions of the test, patients with schizophrenia were impaired in placing the items in the correct temporal order. Recall and recognition of the actual words used to comprise the lists were also impaired in the schizophrenic patients. However, when recall measures were covaried, and when patients were matched with controls for recall, post-hoc group differences in temporal memory were eliminated. In contrast, covarying recognition (indexed by d' or matching for recognition) did not eliminate group differences. Therefore, although memory for temporal order is compromised in patients with schizophrenia, this deficit is highly correlated with generally poorer item-specific memory retrieval (i.e., recall). It is possible that both impairments are due to some third process that underlies and aids in the reconstruction of episodes. PMID- 11120431 TI - Reaction time, symptom profiles and course of illness in schizophrenia. AB - Schizophrenic subjects perform more poorly than normal controls on a wide range of neurocognitive tasks. Some studies suggest that patients with persistent illness may have different patterns of cognitive deficits and different associations between cognitive deficits and symptom profiles compared with patients with fluctuating illness. This study uses simple reaction time, choice reaction time and Stroop tasks to explore the relationship between reaction time and symptom profiles in patients with fluctuating illness (n=24), persistent illness (n=17) and normal controls (n=16). We tested the hypothesis that in patients with persistent illness, psychomotor poverty is associated with impaired initiation of activity, and that disorganization is associated with impaired selection in both persistent and fluctuating illness. There were no differences in the severity of symptoms in the patient groups. In the simple reaction time task, patients with persistent illness performed more poorly than patients with fluctuating illness and controls. Both patient groups performed more poorly than controls in the choice reaction time tasks. Psychomotor poverty was associated with simple reaction time in patients with persistent symptoms. Disorganization was associated with poorer performance on the choice reaction time task in both patient groups. These results suggest the possibility that persistent illness, negative symptoms, and impaired initiation may reflect dysfunction associated with enduring structural abnormalities. In the case of impaired selection processes and disorganization, our data indicate that these abnormalities occur in both persistent and fluctuating illness. This suggests that enduring brain damage might create a predisposition to impaired selection and disorganization, but these clinical features can also arise as a result of transient biochemical imbalance. PMID- 11120432 TI - IQ scores of treatment-resistant schizophrenia patients before and after the onset of the illness. AB - In this study we examined the correlations of actual pre-morbid IQ scores (obtained from routine educational assessments) and estimated current IQ scores in 27 treatment-resistant schizophrenia patients. Pre-morbid (mean = 93) and current (mean = 83) IQ scores were significantly correlated (r = 0.807, P < 0.0001), while duration of illness (10-40 years) was unrelated to the magnitude of IQ score decline (r = -0.103, P = 0.575). These data suggest that pre-morbid IQ test scores are highly predictive of post-morbid scores. PMID- 11120433 TI - Short form of the WAIS-III for use with patients with schizophrenia. AB - The recent publication of the Wechsler Adult Intelligence Scale (WAIS-III), the most widely used standard test of intelligence, requires the development of a new short form for use with patients with schizophrenia for many clinical and research purposes. We used regression analyses of complete WAIS-III data on 41 outpatients with schizophrenia and 41 education-, and age-matched healthy subjects to determine the best combination of subtests to use as a short form. Excluding three subtests that are time-consuming to administer, and requiring that the solution includes one subtest from each of the four WAIS index scores, the combination that most fully accounted for the variance in full-scale IQ (FSIQ) for both participants with schizophrenia (R(2)=0.90) and healthy controls (R(2)=0.86) included the information, block design, arithmetic, and digit symbol subtests. When the restrictions regarding which subtests could enter were relaxed, the best four-subtest solution included information, block design, comprehension, and similarities. Although the latter explained 95% of the variance in FSIQ for schizophrenia participants and 90% of the variance for healthy controls, it consistently overestimated FSIQ for the schizophrenia group. We recommend the four-factor short form for use in future research and clinical practice in which a quick, accurate IQ estimate is desired. PMID- 11120434 TI - Modification of affect perception deficits in schizophrenia. AB - This study investigated two strategies for improving facial affect perception in schizophrenia: monetary reinforcement and promoting facial feedback via mimicry of the expressions of target faces. A total of 40 inpatients with schizophrenia were administered the face emotion identification test during four phases: baseline, intervention, immediate post-test, and 1week follow-up. Subjects were randomly assigned to one of four interventions: repeated practice, monetary reinforcement, facial feedback, and a combination of reinforcement and facial feedback. Generalization of the intervention to a test of facial affect discrimination was also examined. The results showed that all groups of subjects, with the exception of those in the repeated practice group, improved in their ability to identify facial affect, with these effects showing some stability over time. There was limited evidence of these effects generalizing to the test of facial affect discrimination. PMID- 11120435 TI - Interhemispheric cooperation during word processing: evidence for callosal transfer dysfunction in schizophrenic patients. AB - Functional lateralization and interhemispheric interaction during word processing were investigated in schizophrenic patients (n=12) and matched healthy controls (n=18). Words and phonologically regular pseudowords were presented tachistoscopically either in the left or right visual field (unilateral conditions), or simultaneously in both visual hemifields (bilateral condition). Consistent with earlier findings, healthy controls showed a right visual field advantage (RVFA), indicating left-hemispheric dominance for language. The patients showed a RVFA similar to that of controls, consistent with normal left hemispheric language dominance. Importantly, controls performed much better on words presented in the bilateral condition, when two copies of the same word appeared twice, compared to stimulation in only one of the visual hemifields. This bilateral advantage, which has been interpreted as evidence for cooperation between the hemispheres, was absent in schizophrenics. These data show that schizophrenic patients can exhibit similar lateralization patterns as healthy controls. Their specific functional deficit may be a lack of cooperation between the hemispheres. PMID- 11120436 TI - The prevalence and stability of an executive processing deficit, response inhibition, in people with chronic schizophrenia. AB - The current study investigates whether an executive processing measure, response inhibition, is stable over time and across new samples of patients with schizophrenia. Two groups of patients (with and without diagnoses of schizophrenia) were followed up 6 years after baseline data collection. Another separate group of patients with less institutionalized care also completed the same measures to determine whether the response inhibition difficulties had the same prevalence in this new sample.The response inhibition measure was stable over time only in the group of patients with a diagnosis of schizophrenia. The relationship between symptoms and response inhibition difficulties changed over time but was explicable in terms of the interaction between environmental demands and information processing difficulties. The level of response inhibition difficulties (about one-third) was identical in the new sample of patients, which suggests that response inhibition is not dependent on clinical history. PMID- 11120437 TI - Is the attentional dysfunction in schizotypy related to anxiety? AB - Two factors, 'anxiety-loaded' (AL) and 'perceptual-disorganization' (PD), emerged in a factor analysis of the Schizotypal Personality Questionnaire (SPQ). Sixty of the 219 participants performed a latent inhibition (LI) task. During the pre exposure phase, one group was exposed to repeated non-reinforced presentations of an irrelevant stimulus and the other was not pre-exposed. In the subsequent test phase, learning was slower in the pre-exposed group than in the non-pre-exposed group. The LI effect was assessed, separately, as a function of SPQ, trait anxiety sub-scale (TASS) of the State and Trait Anxiety Inventory (STAI), and AL and PD factors scores. LI was disrupted in participants with either high scores on SPQ, STAI, or the AL factor. A regression analysis indicated that both TASS and SPQ independently accounted for LI disruption in high schizotypals, but that the contribution of TASS was stronger. It was suggested that the anxiety component of schizotypy, more than the perceptual-disorganization (schizophrenia like) component, accounts for the attentional dysfunction in high schizotypals, and for their greater than normal distraction by irrelevant stimuli. PMID- 11120438 TI - Executive/attentional performance and measures of schizotypy in patients with schizophrenia and in their nonpsychotic first-degree relatives. AB - Previous studies of executive/attentional functions have found impairments in nonpsychotic first-degree relatives of patients with schizophrenia. The aims of this study were: (1) to replicate these findings by three laboratory measures of attention/information processing - a continuous performance test (DS-CPT), a forced-choice span of apprehension task (SPAN), and a digit symbol substitution test (DSST), and by a series of neuropsychological tests sensitive to prefrontal cortical damage - Trail Making A and B, verbal fluency (VFT), Stroop Color and Word Test (Stroop), and Wisconsin Card Sorting Test (WCST); (2) to investigate whether such executive/attentional deficits are associated with schizotypal traits assessed using the social anhedonia, physical anhedonia, perceptual aberration and magical ideation scales (Chapman, L.J., Chapman, J.P., Raulin, M.L. 1976. Scales for physical and social anhedonia. J. Abnorm. Psychol. 85, 374 382; Chapman, L.J., Chapman, J.P., Raulin, M.L., 1978. Body-image aberration in schizophrenia. J. Abnorm. Psychol. 87, 399-407; Eckblad, M., Chapman, L.J., 1983. Magical ideation as an indicator of schizotypy. J. Consult. Clin. Psychol. 51, 215-225). In both patient and relative groups, performance was significantly poorer on the DSST, VFT and Trail B, and the reaction time on the SPAN was significantly longer. These neuropsychological impairments were present as much in siblings as in parents of schizophrenic patients; age did not appear to cancel differences between the relative and control groups. In the relative group, the four scores of schizotypy were at an intermediate level between those of patient and control groups, and the social anhedonia and perceptual aberration scores tended to be significantly different between the relative and the control groups. Only two significant correlations were found between the neuropsychological performance and the measures of schizotypy. PMID- 11120439 TI - a-b ridge count and schizophrenia. PMID- 11120440 TI - Sex differences in response to red and blue light in human primary visual cortex: a bold fMRI study. AB - Studies using a variety of investigative methods, including functional brain imaging and electroencephalography (EEG), have suggested that changes in central nervous system (CNS) dopamine function result in altered visual system processing. The discovery of abnormal retinal blue cone, but not red cone, electroretinogram in association with cocaine withdrawal and Parkinson's disease suggests that visual system response to blue light might be a marker for CNS dopamine tone. As there are numerous sex-related differences in central nervous system dopamine function, we predicted that blue and red light stimulation would produce sex-specific patterns of response in primary visual cortex when studied using the blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) technique. We analyzed the BOLD response to red and blue light in male and female human volunteers (N=20). Red and blue light responses in primary visual cortex (V1) to stepped intensities of red and blue light were compared by sex for threshold to detectable BOLD signal increase and for stimulus intensity vs. BOLD signal response. Near threshold, males and females showed similar BOLD signal change to red light, but males showed a threefold greater increase (0.52%) to blue light stimulation when compared to females (0.14%). Log-linear regression modeling revealed that the slope coefficients for the red light stimulus intensity vs. signal change curve were not significantly different for males and females (z=0.995, P=0.320), whereas the slope coefficients for the blue light stimulus intensity vs. signal change curve were significantly larger in males (z=2.251, P=0.024). These findings support a sex and color-dependent differential pattern of primary visual cortical response to photic stimulation and suggest a method for assessing the influence of specific dopamine agonist/antagonist medications on visual function. PMID- 11120441 TI - Brain electric correlates of strong belief in paranormal phenomena: intracerebral EEG source and regional Omega complexity analyses. AB - The neurocognitive processes underlying the formation and maintenance of paranormal beliefs are important for understanding schizotypal ideation. Behavioral studies indicated that both schizotypal and paranormal ideation are based on an overreliance on the right hemisphere, whose coarse rather than focussed semantic processing may favor the emergence of 'loose' and 'uncommon' associations. To elucidate the electrophysiological basis of these behavioral observations, 35-channel resting EEG was recorded in pre-screened female strong believers and disbelievers during resting baseline. EEG data were subjected to FFT-Dipole-Approximation analysis, a reference-free frequency-domain dipole source modeling, and Regional (hemispheric) Omega Complexity analysis, a linear approach estimating the complexity of the trajectories of momentary EEG map series in state space. Compared to disbelievers, believers showed: more right located sources of the beta2 band (18.5-21 Hz, excitatory activity); reduced interhemispheric differences in Omega complexity values; higher scores on the Magical Ideation scale; more general negative affect; and more hypnagogic-like reveries after a 4-min eyes-closed resting period. Thus, subjects differing in their declared paranormal belief displayed different active, cerebral neural populations during resting, task-free conditions. As hypothesized, believers showed relatively higher right hemispheric activation and reduced hemispheric asymmetry of functional complexity. These markers may constitute the neurophysiological basis for paranormal and schizotypal ideation. PMID- 11120442 TI - Psychophysiological correlates of social skills deficits in persons with schizophrenia. AB - Social skill deficits in schizophrenia profoundly affect patients' life-long outcome, although the profile of the underlying cognitive dysfunction still remains a matter of debate. In the present study, we investigated the relationship between social skills and event-related potentials (ERPs) in an auditory selective attention task, in addition to the neurocognitive indices obtained from the degraded-stimulus continuous performance test (CPT) and clinical indices, such as Brief Psychiatric Rating Scale (BRPS) and global assessment of function (GAF) scores. Social skills were assessed using a Japanese version of the structured role play test. Fourteen persons with schizophrenia participated in the study. Non-verbal skills showed a positive correlation with GAF, the performance level, N1 and N2b amplitude in the ERP task, and hit rate in the CPT, and a negative correlation with reaction time in the CPT. Verbal communication skills showed a positive correlation with GAF, the performance level and N2b amplitude in the ERP task, and hit rate in the CPT, and a negative correlation with reaction time in the CPT. Processing skills showed a positive correlation with the performance level and N1 amplitude in the ERP task and a negative correlation with reaction time in the CPT. These findings suggested that the social skill deficits of persons with schizophrenia were related to the vigilance level and controlled stimulus detection processing. PMID- 11120443 TI - Assessment of GABA concentration in human brain using two-dimensional proton magnetic resonance spectroscopy. AB - A quantitative method to assess in vivo brain gamma-aminobutyric acid (GABA) levels is proposed using a J-resolved, two-dimensional (2D) magnetic resonance spectroscopy (MRS) technique. Localized, J-resolved 2D MR spectra were obtained from a 12-cm(3) voxel in the occipital lobe of 36 healthy volunteers (18 male and 18 female, age: 25.1+/-4.8 years). Based on phantom measurements, a GABA resonance peak located at 2.94 ppm, 7.45 Hz, in J-resolved 2D MRS overlaps the least with other resonance peaks which arise from N-acetylaspartate, choline, creatine, glutamate and glutamine. Measurements of this resonance peak yield in vivo GABA concentrations of 1.01+/-0.36 micromol/cm(3) for male and 1.16+/-0.43 micromol/cm(3) for female volunteers, without correction for T1 and T2 relaxation effects. These results are in good agreement with previously reported data and suggest that, with further development, 2D MRS may provide a practical means to estimate the concentration of this important neurotransmitter. PMID- 11120444 TI - A high-beef diet alters protein kinase C isozyme expression in rat colonic mucosa. AB - We recently reported that a red meat (beef) diet relative to a casein-based diet increases protein kinase C (PKC) activity in rat colonic mucosa. The purpose of this study was to further elucidate the effects of a high-beef diet on colonic intracellular signal transduction by analyzing steady-state protein levels of different PKC isozymes as well as activities of the three types of sphingomyelinases. Male Wistar rats (n = 12/group) were fed AIN93G-based diets either high in beef or casein for 4 weeks. Rats fed the beef diet had significantly (P < 0.05) higher cytosolic PKC alpha and lower membrane PKC delta protein levels than rats fed the casein diet. The beef-fed rats also had alterations in subfractions of PKC zeta/lambda so that they had a significantly (P = 0.001) lower level of membrane 70 & 75 kDa fraction and a higher (P = 0.001) level of cytosolic 40 & 43 kDa fraction than rats fed the casein diet. Because protein levels analyzed with a PKC zeta-specific antibody were similar, these differences in PKC zeta/lambda were probably due to changes in PKC lambda expression. PKC beta2 levels did not differ between the dietary groups. Diet had no significant effect on the activity of acid, neutral, or alkaline sphingomyelinase. This study demonstrated that consumption of a high-beef diet is capable of modulating PKC isozyme levels in rat colon, which might be one of the mechanisms whereby red meat affects colon carcinogenesis. PMID- 11120445 TI - Plasma appearance and distribution of astaxanthin E/Z and R/S isomers in plasma lipoproteins of men after single dose administration of astaxanthin. AB - Appearance, pharmacokinetics, and distribution of astaxanthin E/Z and R/S isomers in plasma and lipoprotein fractions were studied in 3 middle-aged male volunteers (37-43 years) after ingestion of a single meal containing a 100 mg dose of astaxanthin. The astaxanthin source consisted of 74% all-E-, 9% 9Z-, 17% 13Z astaxanthin (3R,3'R-, 3R,3'S; meso-, and 3S,3'S-astaxanthin in a 1:2:1 ratio). The plasma astaxanthin concentration--time curves were measured during 72 hr. Maximum levels of astaxanthin (1.3 +/- 0.1 mg/L) were reached 6.7 +/- 1.2 hr after administration, and the plasma astaxanthin elimination half-life was 21 +/- 11 hr. 13Z-Astaxanthin accumulated selectively, whereas the 3 and 3'R/S astaxanthin distribution was similar to that of the experimental meal. Astaxanthin was present mainly in very low-density lipoproteins containing chylomicrons (VLDL/CM; 36-64% of total astaxanthin), whereas low-density lipoprotein (LDL) and high-density lipoprotein (HDL) contained 29% and 24% of total astaxanthin, respectively. The astaxanthin isomer distribution in plasma, VLDL/CM, LDL, and HDL was not affected by time. The results indicate that a selective process increases the relative proportion of astaxanthin Z-isomers compared to the all-E-astaxanthin during blood uptake and that astaxanthin E/Z isomers have similar pharmacokinetics. PMID- 11120446 TI - Sea buckthorn berry oil inhibits platelet aggregation. AB - A small-scale preliminary cross-over study was conducted to investigate the effects of supercritical CO(2)-extracted sea buckthorn berry oil (SBO) on some risk factors of cardiovascular disease. Special features of the oil are high proportions of palmitic (16:0), oleic (18:1n-9), palmitoleic (16:1n-7), linoleic (18:2n-6), and alpha-linolenic (18:3n-3) acids as well as vitamin E, carotenoids, and sterols. Twelve healthy normolipidemic men were recruited and each volunteer consumed SBO and fractionated coconut oil (control) 5 g per day for a period of 4 weeks in a random order (wash-out 4-8 weeks). Phospholipid fatty acids, plasma lipids, and glucose were unaffected by SBO supplementation. Instead, a clear decrease in the rate of adenosine-5'-diphosphate-induced platelet aggregation and maximum aggregation were found. This suggested the beneficial effects of SBO on blood clotting, but further studies on the dose-response effects are needed to assess the practical use of SBO supplements. PMID- 11120447 TI - Fasting does not increase mRNA levels of proteolytic systems in small intestinal mucosa of the rat. AB - Fasting results in rapid and profound wasting of the small intestine. mRNA levels of genes encoding critical components of proteolytic systems were measured in small intestinal mucosa to indirectly assess the possible role that proteolysis plays in mediating this wasting. Male Sprague-Dawley rats (120 g; n = 6 per group) were either fed or fasted for 1 or 2 days. Small intestinal mucosal mass decreased by 19% and 31% after 1 and 2 days of fasting, respectively (P < 0.05). Fasting did not significantly change mRNA levels for lysosomal (cathepsin B) or ubiquitin-proteasome-dependent (ubiquitin, 14-kDa ubiquitin-conjugating-enzyme E2, and the C8 and C9 proteasome subunits) systems. Northern hybridizations were also performed using membranes made with poly A(+) mRNA instead of total RNA. mRNA levels for these proteolytic systems and m-calpain did not significantly change with fasting. These data clearly demonstrated that fasting does not increase expression of genes encoding critical components of proteolytic systems in the small intestinal mucosa, suggesting that increased proteolysis cannot explain wasting of the small intestinal mucosa during brief fasting in young rats. PMID- 11120448 TI - Fructooligosaccharides enhance mineral apparent absorption and counteract the deleterious effects of phytic acid on mineral homeostasis in rats. AB - Phytic acid (PA) and fructooligosaccharides (FOS) such as inulin are two food components that are able to modify mineral absorption negatively or positively. The influence of PA and FOS on the cecal and apparent mineral absorption as well as on the mineral status (plasma, hepatic, and bone) were investigated in four groups of rats fed one of the experimental diets: a fiber-free (FF) diet, a FF diet containing 7 g/kg PA (FF + PA), a diet containing 100 g/kg inulin (FOS), or a FOS diet containing 7 g/kg PA (FOS + PA). The cecal enlargement together with the acidification of cecal pH in rats adapted to FOS diets led to an improved Ca and Mg cecal absorption. Mineral apparent absorption was significantly enhanced by FOS ingestion (Ca, +20%; Mg, +50%; Fe, +23%; Cu, +45%), whereas PA decreased this factor only for trace elements (Fe, -48%; Zn, -62%; Cu, -31%). These inhibitory effects of a FF + PA diet have repercussions on blood (Mg, -15%; Fe, 12%; transferrin saturation -31%), liver (Mg, -18%; Fe, -42%; Zn, -25%), and bone (Zn, -25%) variables. However, the introduction of FOS into a PA diet counteracted these observed deleterious effects by stimulating bacterial hydrolysis of PA (+60% in rats adapted to FOS + PA compared to those fed the FF + PA diet) and by improving cecal absorption of minerals. PMID- 11120449 TI - Changes in the prostaglandin levels in alcohol toxicity: effect of curcumin and N acetylcysteine. AB - The present study was undertaken to evaluate the potential role of curcumin, the antioxidant principal from Curcuma longa Linn., and the sulphur-containing amino acid N-acetylcysteine against ethanol-induced changes in the levels of prostanoids. Biochemical assessment of liver damage was done by measuring the activities of serum enzymes (i.e., aspartate transaminase and alkaline phosphatase), which were significantly increased in rats fed ethanol, whereas the elevated levels of these enzymes were decreased after curcumin and N acetylcysteine treatment to rats fed ethanol. We observed a significant increase in the levels of prostaglandins E(1), E(2), F(2alpha), and D(2) in liver, kidney, and brain. Administration of curcumin and N-acetylcysteine was shown to decrease the level of these prostanoids in the tissue studied. PMID- 11120450 TI - The immune responses to diabetes in BB rats supplemented with vitamin A. AB - A substantial amount of evidence suggests that in type I diabetes, vitamin A and zinc status could be of concern because of their impaired metabolic availability. Because both vitamin A and zinc play important roles in the regulation of immune function, the present study was undertaken to examine the immune responses to vitamin A and zinc supplements in diabetic-prone Bio-Breed rats (BBdp), and if the supplements increase the incidence of diabetes. Weanling BBdp rats were fed a NIH-07 diet supplemented with vitamin A either alone or in combination with zinc up to 120 days of age. A greater percentage of rats developing diabetes was found in rats that had supplements of vitamin A and zinc (67%) than those on the basal diet (55%) or with vitamin A supplementation alone (50%). The B cells and macrophages were all markedly increased, whereas CD(4)(+) and CD(8)(+) T cells were decreased at the onset of diabetes. However, this immune status was not changed by vitamin A and zinc supplements. The plasma vitamin A levels were significantly decreased in the presence of diabetes and the vitamin A status did not improve when the rats were given vitamin A and zinc supplements. The Natural Killer cell cytotoxicity on a per-cell basis was significantly decreased in the presence of diabetes, irrespective of supplements with vitamin A and zinc. Overall, results indicated that vitamin A and immune status are both affected by type I diabetes; these effects, however, are not responsive to supplemental intakes of vitamin A either alone or in combination with zinc. PMID- 11120451 TI - Increases in VO2max and metabolic markers of fat oxidation by caffeine, carnitine, and choline supplementation in rats. AB - We have previously shown that the combination of caffeine, carnitine, and choline supplementation decreased body fat and serum leptin concentration in rats and was attributed to increased fat utilization for energy. As a result, it was hypothesized that the supplements may augment exercise performance including physiological and biochemical indexes. Twenty 7-week-old male Sprague-Dawley rats were given free access to a nonpurified diet with or without supplementation of caffeine, carnitine, and choline at concentrations of 0.1, 5, and 11.5 g/kg diet, respectively. One half of each dietary group was exercised on a motor-driven treadmill for 3 weeks and maximal aerobic power (VO(2)max) was determined on the 18th day of exercise. Rats were killed 24-hr postexercise, and blood, regional fat pads, and skeletal muscle were collected. The VO(2)max was increased (P < 0.05) in the supplemented/exercised group; however, the respiratory quotient (RQ) was not affected. Postexercised concentrations of serum triglycerides were decreased but beta-hydroxybutyrate, acylcarnitine, and acetylcarnitine were increased in the supplemented animals. The changes in serum metabolites were complemented by the changes in the muscle and urinary metabolites. The magnitude of increase in urinary acylcarnitines (34-45-fold) is a unique effect of this combination of supplements. Cumulative evidence indicates enhanced beta-oxidation of fatty acids without a change in the RQ because acetyl units were excreted in urine as acetylcarnitine and not oxidized to carbon dioxide. For this phenomenon, we propose the term "fatty acid dumping." We conclude that supplementation with caffeine, carnitine, and choline augments exercise performance and promotes fatty acid oxidation as well as disposal in urine. PMID- 11120452 TI - A 6-year nationwide cohort study of glycaemic control in young people with type 1 diabetes. Risk markers for the development of retinopathy, nephropathy and neuropathy. Danish Study Group of Diabetes in Childhood. AB - The study aimed to identify risk markers (present at the start of the study in 1989) for the occurrence and progression of microvascular complications 6 years later (in 1995) in a Danish nationwide cohort of children and adolescents with Type 1 diabetes (average age at entry 13.7 years). Probabilities for the development of elevated albumin excretion rate (AER), retinopathy, and increased vibration perception threshold (VPT) could then be estimated from a stepwise logistic regression model. A total of 339 patients (47% of the original cohort) were studied. Sex, age, diabetes duration, insulin regimen and dose, height, weight, HbA(1c), blood pressure, and AER were recorded. In addition, information on retinopathy, neuropathy (VPT), and anti-hypertensive treatment was obtained at the end of the study. HbA(1c) (normal range 4.3-5.8, mean 5.3%) and AER (upper normal limit <20 microg min(-1)) in two, timed overnight urine collections were analysed centrally. Eye examination was performed by two-field fundus photography. Determination of VPT was assessed by biothesiometry. Increased AER (> or =20 microg min(-1)) was found in 12.8% of the patients in 1995, and risk markers for this were increased AER and high HbA(1c), in 1989 (both p<0.001). Retinopathy was present in 57.8% of patients in 1995, for which the risk markers were long duration of diabetes (p<0.0001), age (p<0.01), and high HbA(1c) (p<0.0001) in 1989. Elevated VPT (>6.5 V) was found in 62.5% of patients in 1995, for which the risk markers were male sex (p<0.05), age (p<0.0001), and increased AER (p<0.05) in 1989. This study confirms that hyperglycaemia plays a major role for the development of microvascular complications in kidneys and eyes, and emphasises the need for optimal glycaemic control in children and adolescents with Type 1 diabetes. PMID- 11120453 TI - The prognosis for type 2 diabetic patients with heart disease. A 10-year observation study of 385 patients. AB - The objective was to study the development and progression of heart disease in type 2 diabetic patients and to evaluate the influence of revascularisation procedures on its outcome. A 10-year observation study in 385 patients attending a hospital-based outpatient clinic was performed. A total of 156/385 patients developed myocardial infarction (n=68), angina (n=44), heart failure (n=34) or died (n=109). A high mortality was seen in patients with myocardial infarction (73%) and heart failure (71%), in contrast, to patients with angina (25%). Thirty patients had a coronary angiography because of angina, out of which 23 were revascularised. Four (17%) of patients with bypass surgery or angioplasty died compared with 57 (67%) of the patients with no intervention (p<0.001). The occurrence of myocardial infarction was associated with age (p<0.0001), and mean systolic (p<0.05) and diastolic (p<0.05) blood pressure and degree of albuminuria at entry (p<0.05). Heart failure was associated with age (p<0.0001), and mean HbA(1c) levels (p<0.05), while angina was associated with age only (p<0.05). Death was associated with age (p<0.0001), diabetes duration (p<0.05), mean diastolic blood pressure (p<0.05), and degree of albuminuria at entry (p<0.0001). This study shows a high incidence of heart disease in patients with type 2 diabetes. The prognosis was better in patients who had had a revascularisation procedure. Thus, a more active attitude towards revascularisation may potentially improve the prognosis for type 2 diabetic patients with atherosclerotic heart disease. PMID- 11120454 TI - Comparison of risk factors of macrovascular complications. Peripheral vascular disease, cerebral vascular disease, and coronary heart disease in Japanese type 2 diabetes mellitus patients. AB - Although macroangiopathies such as peripheral vascular disease (PVD), cerebral vascular disease (CVD), and coronary heart disease (CHD) can often be observed in patients with diabetes mellitus, they are not specific for diabetes mellitus. Moreover, it is unclear whether their progressive mechanism is different. In the present study, we compared the risk factors among the diabetic macrovascular complications. Univariate analyses showed that in all patients, age at examination, duration of diabetes, thrombin-antithrombin III complex (TAT) level, fibrinogen level, lipoprotein (a) (Lp(a)) level, total cholesterol (T-Chol) level, and existence of microagiopathy were risk factors for PVD. Age, duration of diabetes, insulin level, TAT level, fibrinogen level, HDL cholesterol (HDL Chol) level, hypertension, and nephropathy were risk factors for CVD. Only fibrinogen level was a risk factor for CHD. Moreover, Lp(a) level was a risk factor for PVD and CVD in male patients, but not in females. On the other hand, insulin level was a risk factor for CVD in female patients, but not in males. Multivariate analyses showed that TAT level, T-Chol level, and neuropathy were independent variables for PVD and that age, TAT level, and HDL-Chol level were independent variables for CVD. On the other hand, only fibrinogen level was the independent variable for CHD in males. Our results suggest that the progressive mechanism of PVD and CVD might be different from that of CHD and might differ according to gender in Japanese diabetic patients. PMID- 11120455 TI - Impact of a history of diabetes on the improvement of symptoms and quality of life during 5 years after coronary artery bypass grafting. AB - To describe the impact of a history of diabetes mellitus on the improvement of symptoms and various aspects of quality of life (QoL) during 5 years after coronary artery bypass grafting (CABG). Patients who underwent CABG between 1988 and 1991 in western Sweden were approached with an inquiry prior to surgery and 5 years after the operation. QoL was estimated with three different instruments: Physical Activity Score (PAS), Nottingham Health Profile (NHP) and Psychological General Well-Being (PGWB) index. 876 patients participated in the evaluation, of whom 87 (10%) had a history of diabetes. Symptoms of dyspnea and chest pain improved both in diabetic and non-diabetic patients. Diabetic patients scored worse than non-diabetic patients both prior to and 5 years after CABG, but without any major difference in improvement between the two groups with all three measures of QoL. PAS tended to improve more in non-diabetic than in diabetic patients, whereas improvement in NHP and PGWB was similar regardless of a history of diabetes. Diabetic patients differ from non-diabetic patients having an inferior QoL both prior to and 5 years after CABG. Both diabetic and non-diabetic patients improve in symptoms and QoL after the operation. In some aspects improvement tended to be less marked in the diabetic patients but on the whole improvement was similar compared to non-diabetic patients. PMID- 11120456 TI - Effects of behavior-modifying education in the metabolic profile of the type 2 diabetes mellitus patient. AB - To demonstrate the advantages of behavior-modifying education in the metabolic profile of the type-2 diabetes mellitus patient. A quasi-experimental study was performed with a control group. The experimental group was made up of 25 type-2 diabetic patients and the control group consisted of 24. The type of education carried out was a behavior modification. Baseline measurements and subsequent monthly measurements of serum glucose, total cholesterol and triglycerides were carried out during 9 months after the intervention. The groups were controlled according to age and sex. The statistical analysis was performed using the Student's and Wilcoxon's test to determine the difference. The experimental group in comparison with the control group in the measurement after the intervention achieved a mean difference in serum glucose of 64.2 mg/dl (p=0.001), in the cholesterol of 31.6 (p=0.008), and in the triglycerides of 50.8 (p=0.006). The behavior-modifying education is a better option than traditional intervention for metabolic control in type-2 diabetes mellitus patients. PMID- 11120457 TI - Impaired leptin response to glucocorticoid as a chronic complication of diabetes. AB - Leptin has anorectic, anti-obesity, and insulin-sensitizing properties. We recently reported subnormal responses to the leptin secretagogue dexamethasone in diabetes (DM). To determine whether this defect precedes or follows the occurrence of diabetes, we have studied 37 adults: 11 with type 2 DM diagnosed within 6 months prior to study, 16 with chronic (> or =20 years) DM, and 10 healthy controls. After baseline measurements, subjects ingested dexamethasone (4 mg), followed by blood sampling 16 and 40 h later. Nadir plasma cortisol levels (<2.5 mg/dl) occurred 16 h after dexamethasone ingestion in all study groups; this period of maximal biological action of dexamethasone was associated with peak plasma leptin levels. The peak dexamethasone-stimulated plasma leptin responses (% baseline, +/-SEM) were 188+/-18.7% among healthy controls, 180+/ 13.8% among new DM patients, and 127+/-10.5% (P<0.01) in chronic DM patients. Following dexamethasone ingestion, plasma glucose remained stable in the control and new DM groups but increased by 240% in the chronic DM patients; in contrast, plasma insulin increased significantly in controls and new DM patients but not in patients with chronic DM. These results indicate that plasma leptin responses to secretagogue are preserved in newly diagnosed diabetes patients but markedly attenuated in patients with long-standing diabetes, who also were unable to augment insulin secretion during glucocorticoid treatment. Thus, defective glucocorticoid augmentation of plasma leptin, probably related to beta-cell failure, may be a novel chronic complication of diabetes. Theoretically, such a defect could contribute to the obesity and insulin resistance associated with diabetes. PMID- 11120458 TI - Intensive diabetes management decreases Na-Li countertransport in young subjects with type 1 diabetes and enlarged kidneys. AB - In type 1 diabetes, increases in sodium-lithium countertransport (Na-Li CT), kidney volume (KV), and albumin excretion rate (AER) may precede the development of persistent microalbuminuria. Limited data exist on reversibility of these factors early in the evolution of diabetic nephropathy. A crossover design was used to study the separate effects of enalapril and intensive diabetes management (IDM) on Na-Li CT, KV and AER in 17 children and adolescents with type 1 diabetes (5-10 years duration) with large kidneys (>275 ml/1. 73 m(2)) and predominantly normoalbuminuria. Subjects were randomized to receive 3 months of either enalapril (0.25 mg/kg/day) or IDM, a 3-month washout, followed by the alternate treatment for 3 months. During IDM, HbA1c decreased 2.5% (pre 9.5+/-0.3% (mean+/ SE), post 7.0+/-0.1%, p<0.0001), but was unchanged while on enalapril (pre 8.8+/ 0.3%, post 8.5+/-0.3%, p=0.1). A significant decrease in Na-Li CT was seen with IDM (pre 0.43+/-0.05, post 0.36+/-0.04 mmol/l RBC/h, p=0.006) but not angiotensin converting enzyme inhibition (ACE-i) (pre 0.39+/-0.04, post 0.38+/-0.04 mmol/RBC/h, p=0.4). Neither ACE-i nor IDM affected KV or AER. It is concerning that kidney enlargement does not appear reversible at this early stage in the pathogenesis of diabetic nephropathy, although our conclusions are limited by the short duration of intervention and small sample size. The reduction in Na-Li CT with IDM suggests this may be a potentially modifiable risk factor for diabetic nephropathy. PMID- 11120459 TI - The role of the cytoskeleton and a molecular motor in trichome morphogenesis. PMID- 11120460 TI - Mode of action of auxin herbicides: a new ending to a long, drawn out story. PMID- 11120461 TI - Post-transcriptional gene silencing: conservation and sequences. PMID- 11120462 TI - A conduit for T-DNA through the plant cell membrane: dig it yourself! PMID- 11120464 TI - GM crops welcome in China. PMID- 11120463 TI - Green tea lowers cholesterol. PMID- 11120465 TI - More GM tomatoes. PMID- 11120466 TI - New ways for old genes. PMID- 11120467 TI - Vaccine crops. PMID- 11120468 TI - Space potato technology in Poland. PMID- 11120469 TI - PICKLE gene might hold clue to cancer. PMID- 11120470 TI - US$3.4 million project on genetics of rice. PMID- 11120471 TI - High-protein, high-yielding maize. PMID- 11120472 TI - Potato blight resistance from their mexican species. PMID- 11120473 TI - All in good time: the Arabidopsis circadian clock. AB - Biological time-keeping mechanisms have fascinated researchers since the movement of leaves with a daily rhythm was first described >270 years ago. The circadian clock confers a approximately 24-hour rhythm on a range of processes including leaf movements and the expression of some genes. Molecular mechanisms and components underlying clock function have been described in recent years for several animal and prokaryotic organisms, and those of plants are beginning to be characterized. The emerging model of the Arabidopsis clock has mechanistic parallels with the clocks of other model organisms, which consist of positive and negative feedback loops, but the molecular components appear to be unique to plants. PMID- 11120474 TI - Gibberellin metabolism: new insights revealed by the genes. AB - The identification of most of the genes involved in the metabolic pathways for gibberellin hormones has helped us to understand these pathways and their regulation. Many of these enzymes are multifunctional and therefore fewer enzymes than might be expected are required to synthesize the various gibberellin structures. However, several of the enzymes are encoded by multiple genes that are regulated differently, adding unexpected genetic complexity. Several endogenous and environmental factors modify the expression of gibberellin biosynthesis genes, including developmental stage, hormonal status and light. A future challenge will be to dissect the complex, interacting pathways that mediate the regulation of gibberellin metabolism. PMID- 11120475 TI - Plant fructokinases: a sweet family get-together. AB - Plant fructokinases are the gateway to fructose metabolism. Here, we discuss the properties of published plant fructokinases and compare the available protein sequences. In addition, we speculate on the possible function of fructokinases as sugar sensors. A proposal is presented to clarify the confusing fructokinase nomenclature. Only a few plant fructokinase genes have been cloned but the recent isolations of two such genes in tomato and three in Arabidopsis have given this research an important impulse. PMID- 11120476 TI - Phenotypic plasticity for plant development, function and life history. AB - A single genotype can produce different phenotypes in different environments. This fundamental property of organisms is known as phenotypic plasticity. Recently, intensive study has shown that plants are plastic for a remarkable array of ecologically important traits, ranging from diverse aspects of morphology and physiology to anatomy, developmental and reproductive timing, breeding system, and offspring developmental patterns. Comparative, quantitative genetics and molecular approaches are leading to new insights into the adaptive nature of plasticity, its underlying mechanisms and its role in the ecological distribution and evolutionary diversification of plants. PMID- 11120477 TI - The plant kinetochore. AB - Kinetochores are large protein complexes that bind to centromeres. By interacting with microtubules and their associated motor proteins, kinetochores both generate and regulate chromosome movement. Kinetochores also function in the spindle checkpoint; a surveillance mechanism that ensures that metaphase is complete before anaphase begins. Although the ultrastructure of plant kinetochores has been known for many years, only recently have specific kinetochore proteins been identified. The recent data indicate that plant kinetochores contain homologs of many of the proteins implicated in animal and fungal kinetochore function, and that the plant kinetochore is a redundant structure with distinct biochemical subdomains. PMID- 11120478 TI - A review of inherited cancer syndromes and their relevance to oral squamous cell carcinoma. AB - This paper examines the genetic defects associated with inherited cancer syndromes and their relevance to oral cancer. Tumour suppressor genes are now thought of as either gatekeepers or caretakers according to whether they control cell growth directly by inhibiting cell proliferation and/or promoting cell death (gatekeepers) or whether they maintain the integrity of the genome by DNA repair mechanisms (caretakers). In disorders such as xeroderma pigmentosum, ataxia telangiectasia, Bloom syndrome and Fanconi's anaemia, where there are defective caretaker genes, there is an increased incidence of second primary malignancies, including oral cancer. By contrast, with the exception of Li Fraumeni syndrome, abnormalities of gatekeeper genes do not predispose to oral cancer. Not only do Li Fraumeni patients develop second primary malignancies, but defects of the p53 pathway (p53 mutation, MDM2 over-expression, CDKN2A deletion) appear to be a ubiquitous feature of sporadic oral cancer as it occurs in the West. The findings suggest that genetic instability is of fundamental importance in the pathogenesis of oral cancer. PMID- 11120479 TI - Peripheral ameloblastoma: biological profile based on 160 cases from the literature. AB - The present profile of the peripheral ameloblastoma (PA) is based on a literature survey of 160 published tumour cases. The PA is an exophytic growth localized to the soft tissues overlying the tooth-bearing areas of the jaws, the initial diagnosis often being fibrous epulis. In most cases there is no radiological evidence of bone involvement, but a superficial bone erosion--known as cupping or saucerization--may be detected at operation. The PA accounts for 2-10% of all ameloblastomas. The overall average age is 52.1 years, slightly higher for males (52.9 years) than for females (50.6 years). Thus, the PA occurs at a significantly higher age than the intraosseous ameloblastoma (IA; 37.4 years). The male/female ratio amounts to 1.9:1, as opposed to 1.2:1 for the IA. The male/female ratio for the Japanese cases included in this survey is 2.5:1 as opposed to that of non-Japanese cases 1.4:1. As to the location of PA, the maxilla/mandible ratio is 1:2.6. The mandibular premolar region accounts for 32.6% of all sites. Five extra-gingival lesions have been reported under the term PA. As these cases most likely represent salivary gland tumours, they are not accepted under the diagnosis of PA. The odontogenic gingival epithelial hamartoma shows clinical, histological and behavioural features almost identical to the PA, and it is discussed whether this lesion and the PA should be considered one and the same entity. Pathogenetically, two major sources are discussed: remnants of the dental lamina and the oral surface epithelium. Histologically, the PA consists of proliferating odontogenic epithelium exhibiting the same histomorphological cell types and patterns as seen in the IA. The stroma is that of a mature, fibrous connective tissue. The indolent biological behaviour dictates a conservative therapeutical approach. It is discussed whether PA is a true neoplastic counterpart of the IA or rather an odontogenic hamartomatous lesion. Six cases of malignant PA have been reported. PMID- 11120480 TI - Smoking and alcohol in the etiology of oral cancer: gender-specific risk profiles in the south of Greece. AB - Oral and pharyngeal cancer (OC) mortality is very low in Greece, especially among men, compared to other European countries. We conducted a case-control study of OC in Athens, and obtained information on tobacco, alcohol use and other potential risk factors and confounding variables for 110 incident cases and 115 hospital-based controls. We used multivariate logistic regression to estimate odds ratios (ORs) and 95% confidence intervals (CIs). Tobacco smoking (pack years, P(trend)=0.01) and alcohol use (drinks/week, P(trend)=0.07) were independent risk factors, with a multiplicative effect for combined exposures (OR, 8.3; 95% CI, 2.4-29.1, for >28 alcohol drinks/week and >50 pack years of cigarette smoking). The type of alcoholic beverage also seemed important: drinking ouzo and tsipouro (liquors of high ethanol concentration) was associated with greater increased OC risk than drinking comparable amounts of wine, beer or dark spirits. While alcohol drinking is more common for male cases versus controls, few men reported regularly consuming large quantities of ethanol associated with highest risk of OC in other studies. This may partially explain the low rates of male OC mortality in Greece. Among the 38% of our cases who were women, however, neither smoking nor alcohol drinking frequencies were significantly elevated compared to controls, and so the etiology of OC risk in females requires further investigation. PMID- 11120481 TI - The diagnostic value of fine needle aspiration cytology in Waldeyer's ring lymphomas. AB - Fine needle aspiration cytology (FNAC) is a well-established diagnostic technique for tumours in the head and neck area. In recent years FNAC has been established as an accurate and useful method for the diagnosis of nodal malignant lymphoproliferative disease. The purpose of the present study was to determine and evaluate the accuracy of FNAC in the diagnosis of primary malignant lymphoma of Waldeyer's ring. The cases of 29 patients suffering from tumours of the oro- and nasopharynx, in which the diagnosis of lymphoma was established by FNAC during the years 1991-1998, were collected and analysed. Twelve of the patients had a previous history of lymphoma, and FNAC was used to establish the diagnosis of recurrent disease. In 17 patients with no previous history of malignancy FNAC was used to diagnose primary extranodal non-Hodgkin's lymphomas (NHLs). In two patients FNAC failed to diagnose NHL. In all patients cytological findings were complemented and compared with those of a histopathological examination after open biopsy. In two cases a difference in the specific histological type of the lymphoma was noted. The findings from the present study (sensitivity 93.10% and positive predictive value 100%) indicated that FNAC is a useful and accurate method in establishing diagnosis of Waldeyer's ring lymphomas. PMID- 11120482 TI - Assessment of various topical oral formulations by bone marrow transplant recipients. AB - Topical approaches to management of oral mucositis have the advantages of high local concentration and limited or no systemic absorption, reducing the risk of systemic complications. The acceptability of topical therapies in cancer patients has not been evaluated. Thirty-eight transplant patients assessed the acceptability of three formulations (rinse, thin gel, thick gel) of a new compound developed to prevent oral mucositis. The rinse was selected as the most acceptable formulation. The thick gel received the lowest rating. Consistency was a major determinant of overall acceptability. A neutral taste was desirable for most participants. Room temperature of the formulation was rated as completely acceptable. Most participants would be willing to use the rinse or thin gel several times per day, to retain each dose in the oral cavity for up to 5 min, and to swallow it. The support of caregivers may be needed to achieve compliance with such regimens. PMID- 11120483 TI - Serum level and tissue expression of c-erbB-1 and c-erbB-2 proto-oncogene products in patients with squamous cell carcinoma of the head and neck. AB - The proto-oncogene products erbB-l (EGF-Receptor) and erbB-2 (HER-2/neu), distinct members of the epidermal growth factor receptor family, are frequently overexpressed in squamous cell carcinoma of the head and neck (SCCHN). The accumulation of these transmembrane proteins may lead to significant amounts of the respective extracellular receptor domains (ECD) that are shed from the tumour cell surface and enter blood circulation, thus representing potential serum tumour markers. For erbB-l and erbB-2, we determined the ECD serum levels with enzyme-linked immunosorbent assays and evaluated the protein expression in tumour tissue by immunohistochemistry. The present study included 49 patients (37 untreated, 12 recurrences) and the same number of age- and sex-matched healthy controls. In 24 patients ECD serum levels were determined before and 6 weeks after surgery. Mean ECD serum levels for erbB-1 and erbB-2 were 54.8+/-1.6 and 153.7+/-6.1 fmol/ml in cancer patients, and 54+/-1.5 and 147.9+/-4.5 fmol/ml in healthy controls, respectively. There was no significant difference between untreated and recurrent disease. Serum ECD follow-ups 6 weeks after surgery revealed a significant 12.3% decline of erbB-1 but no change of erbB-2 values. Immunohistochemistry showed strong staining for erbB-1 in 78% and erbB-2 in 47% of the SCCHN specimens. No correlation was detectable between receptor ECD serum levels and receptor tissue expression, tumour stage, and tumour differentiation. Hence, ECD serum levels of erbB-1 and erbB-2 are not considered to be valuable tumour markers in SCCHN. PMID- 11120484 TI - The pattern of expression of the 5T4 oncofoetal antigen on normal, dysplastic and malignant oral mucosa. AB - The human 5T4 oncofoetal antigen is expressed by all types of trophoblast in pregnancy but is not detected on most adult tissues, although low levels are found on some epithelia. However, this antigen is strongly expressed by many cancers and tumour-associated labelling correlates with metastatic spread and poor clinical outcome for patients with gastric and colon cancer. Over-expression of the gene influences cell adhesion, shape and motility, which may be related to changes in the cellular localisation of the 5T4 oncofoetal antigen as malignancy develops. To establish whether the 5T4 oncofoetal antigen can serve as a tumour specific marker for oral cancer and precancer, we have evaluated the pattern of expression on biopsies of normal, inflamed and dysplastic oral mucosa using immunohistochemistry. Oral mucosa, taken from different sites in the mouth, expressed the 5T4 oncofoetal antigen with varying intensity and pattern. The majority of the immunoreactivity was detected in the basal and suprabasal layers, with expression extending into the spinous cells at fully keratinised sites and when inflammation was present. This antigen was also detected in the underlying connective tissue. Oral squamous cell carcinoma showed a variety of patterns and intensity of staining corresponding to those found for normal mucosa. However, 21 of 41 cases showed no stromal labelling, a finding also observed for dysplastic lesions. The alterations in the pattern and intensity of 5T4 oncofoetal antigen expression were not related to clinicopathological features of the tumours examined. These data show that the 5T4 oncofoetal antigen is expressed on normal oral mucosa, such that this target cannot be used for detection of neoplastic or preneoplastic cells, although altered expression may contribute to the pathogenesis of these lesions. PMID- 11120485 TI - Reverse correlation of E-cadherin and snail expression in oral squamous cell carcinoma cells in vitro. AB - The loss of E-cadherin expression has been shown to correlate to the invasion and metastasis of many types of carcinomas. We established E-cadherin positive (HOC719-PE) and negative (HOC719-NE) clones from an oral squamous cell carcinoma (SCC). HOC719-PE cells showed epithelial morphology with E-cadherin expression in the cell membrane, whereas HOC719-NE cells demonstrated fibroblastic morphology without E-cadherin expression. In invasion assay and three dimensional culture, HOC719-NE showed much higher invasive ability than HOC719-PE cells. These cells expressed similar levels of mRNAs for alpha- and beta-catenin. However, HOC719-NE cells, but not HOC719-PE cells, showed strong expression of snail, a transcription factor implicated in the differentiation of epithelial cells into mesenchymal phenotype. This reverse expression of snail and E-cadherin was further observed in other SCC cells including HOC313, and TSU cells that we previously reported to show no expression of E-cadherin protein. These results indicated that the expression of snail has a key role for the acquisition of more invasive and metastatic phenotypes of SCC and the clones we reported here will be useful tools for understanding the mechanism of the transition from epithelial to mesenchymal SCC cells. PMID- 11120486 TI - Volume-corrected mitotic index as a prognostic factor in oral squamous cell carcinomas. AB - In order to predict the prognosis of patients with oral squamous cell carcinomas, volume-corrected mitotic index (M/V index), as well as histopathological and clinical indices were estimated in 50 oral squamous cell carcinoma tumors. Histological malignancy (P = 0.0005), mode of invasion (P = 0.0168) and M/V index (P = 0.0093) showed a significant correlation with survival according to univariate survival analysis. According to multivariate survival analysis (Cox's regression model), the M/V index was the best predictor of the techniques studied of patient prognosis. The M/V index also showed significant correlation with DNA ploidy. The present results suggest that the M/V index is an important prognostic factor for oral carcinoma. Furthermore, the M/V index and DNA ploidy appear to provide similar prognostic information. PMID- 11120487 TI - Hypoxia-induced angiogenesis of cultured human salivary gland carcinoma cells enhances vascular endothelial growth factor production and basic fibroblast growth factor release. AB - The angiogenic activity of two human salivary gland tumor cell lines, ACCS from adenoid cystic carcinoma and IT-2 from mucoepidermoid carcinoma, was examined by stimulating tube formation by bovine capillary endothelial cells (BCE). ACCS and IT-2 were cultured in 20 or 3% oxygen, representing normoxic and hypoxic conditions, respectively, and conditioned medium (CM) was obtained from each culture. The BCE tubes stimulated by hypoxic CM were 1.59 (ACCS) and 1.42 (IT-2) times longer than those stimulated by normoxic CM. The tube-forming activity of CM was inhibited by preincubation with either anti-vascular endothelial growth factor (VEGF) IgG or anti-basic fibroblast growth factor (bFGF) IgG, suggesting that both VEGF and bFGF with angiogenic activity were present in the CM. This was confirmed by ELISA, which also demonstrated increased concentrations of both proteins in the hypoxic CM. Northern blot analysis showed an increased VEGF mRNA level in both carcinoma cells with hypoxia, while hypoxia did not affect the bFGF mRNA level in either cell line. The results suggest that both VEGF and bFGF are major angiogenesis factors in salivary gland tumors, and hypoxia-induced angiogenesis results from upregulation of VEGF and increased release of bFGF. PMID- 11120488 TI - Preserved salivary output and xerostomia-related quality of life in head and neck cancer patients receiving parotid-sparing radiotherapy. AB - Radiotherapy (RT) for head and neck cancers causes salivary dysfunction and diminished xerostomia-related quality of life. We have demonstrated that three dimensional treatment planning and conformational dose-delivery techniques can minimize RT doses to contralateral parotid glands while providing therapeutic doses to tumors. This study's purpose was to assess parotid salivary function up to 1 year post-RT in patients receiving bilateral neck parotid-sparing RT, and to determine if parotid preservation would significantly improve xerostomia-related quality of life. Unstimulated (UPFR) and stimulated (SPFR) parotid flow rates were collected from 20 head and neck cancer patients. All subjects completed a 15 item xerostomia-related quality of life scale (XeQoLS) prior to RT, at the completion of RT, 1, 3, 6 and 12 months post-RT. Salivary flow rates from spared and treated glands were significantly decreased at the completion of RT. After RT completion, spared UPFR and SPFR function increased and was not significantly different from baseline values. Output from treated glands remained statistically indistinguishable from zero throughout the post-RT period. Subjects had a significantly worse xerostomia-related quality of life at the completion of RT compared to baseline, and XeQoLS responses improved significantly 1 month post RT. Responses at 1 year were markedly better than at the completion of RT, but still significantly worse than baseline. These findings suggest that despite parotid-sparing RT, salivary flow rates from treated and spared glands and xerostomia-related quality of life decrease at the completion of RT. However, with the use of parotid-sparing RT, contralateral glands are preserved at 1 year post-RT with a concomitant improvement in xerostomia-related quality of life. PMID- 11120489 TI - Influence of time delay and clinical upstaging in the prognosis of head and neck cancer. AB - The delayed time period between the first diagnosis of a head and neck cancer and its treatment can affect not only the clinical staging of the disease, but also its prognosis. The time period for clinical upstaging is studied in a retrospective cohort series. A retrospective case/control series was established to assess the prognosis of head and neck cancer patients due to a delay in treatment. In the first part of the study we included 69 patients who had confirmed clinical upstage before starting the oncologic treatment (cases). The survival of this series was compared to a historical control group of 138 patients matched by tumour site and stage, having received treatment within a short period after their first medical evaluation (controls). We observed that the natural evolution of a head and neck cancer was progressively faster, from an initial clinical stage to an advanced clinical stage until it becomes untreatable. It was observed that the delay in beginning oncologic treatment influenced the prognosis of the patients (P=0.030), especially in clinical stage IV patients (P=0.001) and oral cavity cancer patients (P=0.007). PMID- 11120490 TI - Immediate knowledge increase from an oral cancer information leaflet in patients attending a primary health care facility: a randomised controlled trial. AB - The aim was to determine the immediate influence of a validated patient information leaflet (PIL) on oral cancer and knowledge in primary care attenders. Participants were patients (n=800) attending their primary health care provider from 14 general practices (eight dental and six medical) in the north west of England. Measures were a previously validated knowledge questionnaire (36 dichotomous items), self-reported dental service attendance history and demographic variables. The results showed that patients who had read the oral cancer PIL demonstrated a significant increase in knowledge regardless of clinical setting (F[1,739]=246.24, P<0.0001). Patients showed improvements in selecting the correct signs and risk factors associated with disease. Immediate knowledge gain from a simple PIL about oral cancer was found and independent of the primary care facility, where the PIL was distributed. PMID- 11120491 TI - Evaluation of surgical excision of non-homogeneous oral leukoplakia in a screening intervention trial, Kerala, India. AB - It is well established that most invasive oral cancers arise from precancerous lesions such as leukoplakia, erythroplakia and oral submucous fibrosis. One of the approaches for control of oral cancer is to detect oral precancerous lesions early in their development and prevent their malignant transformation to invasive cancer either by chemoprevention or by surgical excision of the lesions, with concurrent control of tobacco and alcohol use and other specific aetiological factors. However, the value of specific approaches such surgery in long-term control of lesions and prevention of malignant transformation is not known. We describe our experience with cold knife surgical excision of 59 cases of non homogeneous leukoplakia of the oral cavity diagnosed in the context of a community-based oral cancer cluster randomised oral cancer screening trial in Kerala, India. Two-thirds of these revealed dysplasia on histology. After a minimum follow-up of 12 months (range 12-37 months) after surgical excision, 44 (74.8%) were remaining disease free with no evidence of recurrent/new lesions; during follow-up, three (5%) developed new luekoplakic lesions, and six (10.1%) developed recurrent lesions, while six (10.1%) could not be traced after treatment. There was no event of malignant change during follow-up. The proportion of subjects remaining with no evidence of disease at 3 years by Kaplan Meier method of analysis was 62.1% (95% CI: 0.36-0.87). Accrual and long-term follow-up of large number of surgically treated cases may provide valuable leads to management policies of oral leukoplakia, since, as of now, the added value of specific treatments over and above primary prevention by tobacco and alcohol control remains to be established. PMID- 11120492 TI - Intraosseous mucoepidermoid carcinoma. Report of a long-term evolution case. AB - Primary central mucoepidermoid carcinoma (CMC) of the jaws accounts only for 2-3% of all mucoepidermoid carcinomas reported. Bhaskar in 1963 first analysed the criteria for his central origin, histology and pathogenesis. The authors report a long-term evolution case of CMC of the mandible with peculiar clinical features observed at the Department of Maxillo-Facial Surgery of the School of Medicine and Surgery of the "Federico II" University of Naples (Naples, Italy) examining histopathologic and clinical features and problems related to the treatment. PMID- 11120493 TI - Where does the dwarf stand--and whom does he thank? Presidential address. PMID- 11120494 TI - Cost-effectiveness of urodynamic testing before surgery for women with pelvic organ prolapse and stress urinary incontinence. AB - OBJECTIVE: This study was undertaken to compare cost-effectiveness between 2 preoperative testing strategies for women with pelvic organ prolapse and stress urinary incontinence symptoms. STUDY DESIGN: We developed decision-analytic models that evaluated the cost-effectiveness of basic office evaluation before surgery in women with prolapse and stress urinary incontinence symptoms and contrasted it with that of urodynamic testing. Costs were obtained from the Federal Register; effectiveness of treatment for urinary incontinence was based on the published literature. RESULTS: The strategies of basic office evaluation and urodynamic testing had the same cure rate of urinary incontinence (96%) after initial and secondary treatment. Under baseline assumptions incremental cost effectiveness (cost for single extra cure of urinary incontinence) of urodynamic testing was $328,601. According to sensitivity analyses, basic office evaluation was more cost-effective than urodynamic testing when the prevalence of pure detrusor instability was <8% or when the cost of urodynamic testing was >$103. CONCLUSION: Urodynamic testing before surgery in women with prolapse and stress urinary incontinence symptoms is not cost-effective relative to basic office evaluation. PMID- 11120495 TI - Ball urethroplasty combined with Marshall-Marchetti-Krantz urethropexy versus suburethral sling in patients with intrinsic sphincter deficiency and urethral hypermobility. AB - OBJECTIVE: It was our goal to compare the efficacy of a suburethral fascial sling with that of a combination of Marshall-Marchetti-Krantz urethropexy and Ball urethroplasty in patients with intrinsic sphincter deficiency and urethral hypermobility. STUDY DESIGN: This study consisted of a retrospective observational evaluation of patients from 2 separate practice sites. Preoperative and postoperative data were collected from patients' medical records. The long term results were based on a mailed questionnaire addressing bladder symptoms and quality-of-life issues. RESULTS: Among a total of 48 patients, 37 (77. 1%) responded in the group undergoing Marshall-Marchetti-Krantz urethropexy combined with Ball urethroplasty, and 30 out of 35 (85.7%) patients replied in the suburethral fascial sling group. The mean length of follow-up was 2. 7 years (range, 1-5 years). The patients were similar in age, hormonal status, parity, and previous bladder neck surgery. Similar cure and improvement were demonstrated in both groups (86.6% in the suburethral fascia group and 89.2% in the group with the Marshall-Marchetti-Krantz procedure combined with Ball urethroplasty). No significant differences were found in urinary incontinence types, irritable bladder symptoms, voiding difficulties, or quality-of-life measures. CONCLUSIONS: The suburethral fascial sling and a procedure consisting of Marshall-Marchetti Krantz urethropexy combined with Ball urethroplasty have similar results in patients with intrinsic sphincter and urethral hypermobility. PMID- 11120496 TI - Prevalence of perioperative complications among women undergoing reconstructive pelvic surgery. AB - OBJECTIVE: The primary aim of this study was to report on the prevalence of perioperative complications associated with reconstructive pelvic surgery. A secondary aim was to identify risk factors predictive of perioperative complications in this population. STUDY DESIGN: A retrospective chart review was performed of 100 consecutive cases of reconstructive pelvic surgery. Statistical analysis included descriptive statistics and logistic regression. RESULTS: The prevalence of perioperative complications was 46%, including 13 intraoperative complications and 33 postoperative complications. The readmission rate for complications was 15%. The number of procedures per patient was an independent risk factor for intraoperative blood loss (P <.0038). Intraoperative estimated blood loss in turn was an independent risk factor for perioperative complications (P <.0001). CONCLUSIONS: Perioperative complications associated with reconstructive pelvic surgery were increased relative to those associated with general gynecologic surgery. The number of procedures per patient and associated blood loss appeared to contribute to the increase in perioperative complications. PMID- 11120497 TI - Postoperative resolution of urinary retention in patients with advanced pelvic organ prolapse. AB - OBJECTIVE: This study was undertaken to determine whether surgery for advanced pelvic organ prolapse corrects the voiding dysfunction commonly associated with this condition and if so to evaluate the ability of preoperative voiding studies to predict such correction. STUDY DESIGN: We reviewed the records of all women who underwent surgery at our center between January 1996 and June 1999 for stage 3 or 4 pelvic organ prolapse. Patients were included in this review if they had a postvoid residual volume of >100 mL documented by catheterization on at least 2 occasions, had no normal postvoid residual volumes documented, and had undergone preoperative multichannel urodynamic testing that included an instrumented voiding study. Demographic and urodynamic data were analyzed for the ability to predict whether the elevated postvoid residual volume would be resolved after surgery. RESULTS: Thirty-five patients satisfied the criteria for inclusion in the review. Twenty-six had stage 3 pelvic organ prolapse and 9 had stage 4 pelvic organ prolapse. The mean preoperative postvoid residual volume was 226 mL (range, 105-600 mL). Thirty-one patients (89%) had normal postvoid residual volumes after surgery. As a predictor of elevated postoperative postvoid residual volumes, the preoperative voiding study (performed with the prolapse reduced) had a sensitivity of 66%, a specificity of 46%, a positive predictive value of 12%, and a negative predictive value of 93%. CONCLUSION: In our center a preoperative voiding study performed with the pelvic organ prolapse reduced most accurately predicted postoperative voiding function when results of the voiding study were normal. Most patients with advanced pelvic organ prolapse and elevated postvoid residual volume had normalization of the postvoid residual volume after surgical correction of the pelvic organ prolapse. PMID- 11120498 TI - A transvaginal approach to repair of apical and other associated sites of pelvic organ prolapse with uterosacral ligaments. AB - OBJECTIVE: The objectives of this study were (1) to describe a group of women with pelvic organ prolapse associated with apical loss of support through grading with the Baden-Walker halfway system before, during, and after the corrective operation, (2) to describe the operative repair of the support defects, (3) to report the morbidity associated with the operative repair, and (4) to assess the durability of the repair at each site. STUDY DESIGN: Between January 1, 1994, and December 31, 1998, a total of 302 consecutive women with apical and associated other support defects were evaluated before, during, and after the corrective operation by the senior author (Bob L. Shull). All patients underwent transvaginal reconstructive surgery with native tissue. Two hundred eighty-nine patients (96%) returned for at least one postoperative visit, and they constitute the group used for the follow-up data. Perioperative morbidity was considered to include hemorrhage necessitating homologous blood transfusion, visceral injury, neurologic impairment, or death. Durability was assessed by means of life-table analysis for each of 5 sites in the vagina. RESULTS: All patients had preoperative or intraoperative evidence of grade 1 or greater apical loss of support of and at least one other site of pelvic organ prolapse. Two hundred eighty-nine patients (96%) returned for at least one postoperative visit. Two hundred fifty-one patients (group 1, 87%) had optimal anatomic outcomes, with no persistent or recurrent support defects at any site. Thirty-eight patients (group 2, 13%) had one or more sites with at least grade 1 loss of support during the follow-up interval. Twenty-four of these 38 patients had grade 1 defects that were detectable only on careful pelvic examination. Fourteen of these patients (5%) had grade 2 or greater persistent or recurrent support defects. The anterior segment (bladder) was the site with the most persistent or recurrent support defects, which means that it was the site of the least durable repair. The urethra and cuff had the most durable repairs. Morbidity included a 1% transfusion rate, a 1% ureteral injury or ureteral kinking rate, and a 0.3% postoperative death rate. CONCLUSION: Careful preoperative and intraoperative evaluation of pelvic support defects and the use of native connective tissue and uterosacral ligaments are associated with excellent anatomic outcomes. The durability of the surgical correction varies according to the individual site of repair and the duration of postoperative follow-up. PMID- 11120499 TI - Low colorectal anastomosis after radical pelvic surgery: a risk factor analysis. AB - OBJECTIVE: This study was conducted to analyze our experience with low (8-12 cm above the anal verge) and very low (<6 cm above the anal verge) colorectal resection and primary anastomosis at the time of radical en bloc resection of pelvic malignancies. STUDY DESIGN: A retrospective review of 77 patients undergoing supralevator pelvic exenteration with low colorectal resection and primary anastomosis in our gynecologic oncology service was carried out. Data were obtained from patient medical records and from the tumor registry. Univariate statistical analysis of the data was used. RESULTS: The distribution of primary malignancies in this cohort was as follows: 33 (43%) recurrent or primary cervical carcinomas, 27 (35%) primary or recurrent ovarian carcinomas, 7 (9%) recurrent vaginal carcinomas, 4 (5%) endometrial carcinomas, 3 (4%) colon carcinomas, and 3 (4%) cases of stage IV endometriosis. Forty patients underwent total pelvic exenteration, and 37 patients underwent posterior exenteration. Thirty-six patients in the total pelvic exenteration group had a history of pelvic irradiation. Twelve (30%) of these patients had development of breakdown or fistulas of the anastomosis. Six of the 12 patients (50%) had undergone protective colostomy. Thirty-seven patients underwent posterior exenteration with primary anastomosis for ovarian cancer, endometrial cancer, colon cancer, or endometriosis, and only 1 of these had received pelvic irradiation. This patient did not have a protective colostomy, and a rectovaginal fistula developed. In addition, there were 3 other breakdowns in the posterior exenteration group. Finally, the presence of preoperative ascites did not appear to alter the breakdown rate of the anastomosis among the patients with ovarian cancer who underwent cytoreductive surgery. CONCLUSION: Radical resection of pelvic tissue remains a crucial part of the armamentarium of the gynecologic oncologist. Previous pelvic irradiation appears to be a major risk factor (35% vs 7.5%) for anastomotic breakdown and fistulas, independent of the presence of a protective colostomy. The overall results appear to be better for patients undergoing this procedure as part of a posterior exenteration. PMID- 11120500 TI - Abnormal spinal curvature and its relationship to pelvic organ prolapse. AB - OBJECTIVE: Intra-abdominal vector forces have been implicated in the development of genital prolapse. Because the normal spinal curvature appears to protect the pelvic cavity from direct upper abdominal forces, variations in spinal curvature may alter these vector forces and possibly potentiate the development of pelvic organ prolapse. This study was undertaken to evaluate the relationship of spinal curvature and pelvic organ prolapse, specifically, the loss of lumbar lordosis or pronounced thoracic kyphosis. STUDY DESIGN: A total of 363 patients referred for various complaints of urinary incontinence or pelvic organ prolapse were included in this multicenter, prospective, case-control study. All patients underwent a detailed history with site-specific examinations; pelvic organ prolapse was quantitatively assessed according to the POPQ (pelvic organ prolapse quantitation) staging system. Spinal curvature was measured with a flexi-curve malleable rod when patients were in a fully erect position. Spinal curvature was then transferred to graph paper by tracing the flexi-curve. Thoracic and lumbar curvatures were determined by measuring thoracic and lumbar lengths and widths, respectively. RESULTS: Ninety-two patients had abnormal spinal curvature according to the study criteria. Complete loss of lumbar lordosis was found in 69 patients. Of the 92 patients with an abnormal curvature, 84 currently had or previously had pelvic organ prolapse (sensitivity, 91%). When compared with patients with a normal curvature, patients with an abnormal spinal curvature were 3. 2 times more likely to have development of pelvic organ prolapse (odds ratio, 3.18; 95% confidence interval, 1.46 to 6.93; P =.002). There was no difference in the number of vaginal deliveries, weight of largest vaginally delivered infant, or body mass index. Only 11% (8/72) of patients with stage 0 prolapse had an abnormal spinal curvature, which increased to 30% (28/99) in patients with stage III prolapse (P =.042). CONCLUSION: An abnormal change in spinal curvature, specifically, a loss of lumbar lordosis, appears to be a significant risk factor in the development of pelvic organ prolapse. PMID- 11120501 TI - The role of vaginal apex excision in the management of persistent posthysterectomy dyspareunia. AB - OBJECTIVE: The purpose of this study was to evaluate the effectiveness of vaginal apex excision in the treatment of patients with posthysterectomy dyspareunia. STUDY DESIGN: This was a case series with an independent third-party survey of patients with posthysterectomy dyspareunia managed at the University of Utah Pelvic Pain Clinic. Thirteen patients were first treated with local injections of anesthetics into localized vaginal pain foci. Further evaluation included formal psychometric testing and a diagnostic spinal block. Nine patients underwent surgical excision of the vaginal apex. An independent interviewer who did not know the patients assessed the effects of this procedure on dyspareunia and coital frequency at a mean of 36.4 +/- 3.7 months after the operation. RESULTS: The mean coital verbal analog pain score (1-10 scale) decreased from 9.22 +/- 0.27 before excision of the vaginal apex to 3.11 +/- 0.84 after the operation (P <.001), and coital frequency improved from 5.22 +/- 2.02 episodes per month before surgery to 11.11 +/- 1.82 episodes per month after surgery (P =.02). Of the 9 patients, 5 essentially had the dyspareunia cured. Dyspareunia was decreased and coital frequency was markedly increased in all but 1 of the other 4 cases. CONCLUSION: Excision of the vaginal apex is an effective treatment for carefully selected patients with posthysterectomy dyspareunia. PMID- 11120502 TI - Pelvic muscle electromyography of levator ani and external anal sphincter in nulliparous women and women with pelvic floor dysfunction. AB - OBJECTIVE: The purpose of this study was to compare results of electromyographic assessment of muscular recruitment between nulliparous control subjects without pelvic floor dysfunction and parous subjects with genuine stress urinary incontinence and with pelvic organ prolapse. Interference pattern analysis is an electromyographic technique that reproducibly measures muscular recruitment by detecting both "turns" in the electromyographic signal produced by positive and negative peaks of the motor unit potentials and motor unit potential amplitude. Fewer turns can indicate loss of motor units or failure of central activation of contraction, whereas greater amplitude can indicate reinnervation after nerve damage. STUDY DESIGN: We performed concentric needle electrode electromyographic examinations of the levator ani and external anal sphincter in 15 nulliparous control subjects and 20 parous subjects with abnormalities (n = 9 with genuine stress urinary incontinence, n = 11 with stage III or IV pelvic organ prolapse). We made digital recordings at multiple sites at rest and with moderate and maximal contraction. Interference pattern analysis yielded the number of turns per second and the mean signal amplitude (in microvolts) for each site at each contraction level. We compared individual patient data with data from the healthy population by means of cloud analysis. Mean values of number of turns per second and mean amplitude in each group were then compared with nonparametric methods and regression models. RESULTS: Mean ages were 28.7 years (range, 20-49 years) for the control group, 54.3 years (range, 35-75 years) for subjects with genuine stress urinary incontinence, and 65 years (range, 41-77 years) for subjects with pelvic organ prolapse. Median clinical levator ani strengths were 9 (range, 5-9) in the control group, 5 (range, 2-7) in the genuine stress urinary incontinence group, and 5 (range, 2-8) in the pelvic organ prolapse group. Median external anal sphincter strengths were 9 (range, 7-9) in the control group, 5 (range, 3-9) in the genuine stress urinary incontinence group, and 8 (range, 4-9) in the pelvic organ prolapse group. The external anal sphincters of subjects with pelvic organ prolapse had the highest percentage of abnormal study results according to cloud analysis. Mean number of turns per second in levators was greater in control subjects than in subjects with abnormalities (P =.034). We found similar differences in number of turns per second for the external anal sphincter (P =.004). In contrast, we did not find differences between groups in mean amplitude in either the levator ani or the external anal sphincter. Comparison of patients with genuine stress urinary incontinence versus subjects with pelvic organ prolapse showed no significant difference in the number of turns per second in either muscle. Mean amplitude was greater in the pelvic organ prolapse group than in the genuine stress urinary incontinence group for both muscles (levator ani, P =.028; external anal sphincter, P =.048). Neither mean amplitude nor the number of turns per second could be predicted by clinically estimated levator ani strength, age, or fecal incontinence. CONCLUSION: Compared with nulliparous control subjects, patients with genuine stress urinary incontinence and pelvic organ prolapse had changes in the levator ani and external anal sphincter consistent with either motor unit loss or failure of central activation, or both. Subjects with pelvic organ prolapse had findings consistent with greater recovery than was found in those with genuine stress urinary incontinence. Measures of recruitment by interference pattern analysis correlated better with clinical external anal sphincter strength than with levator ani strength and were independent of age. PMID- 11120503 TI - Bilateral uterosacral ligament vaginal vault suspension with site-specific endopelvic fascia defect repair for treatment of pelvic organ prolapse. AB - OBJECTIVE: The anatomic and functional success of suspension of the vaginal cuff to the proximal uterosacral ligaments is described. STUDY DESIGN: Forty-six women underwent vaginal site-specific repair of endopelvic fascia defects with suspension of the vaginal cuff to the proximal uterosacral ligaments for pelvic organ prolapse. Outcome measures included operative complications, pelvic organ prolapse quantitation, and assessment of pelvic floor symptoms. RESULTS: After a median follow-up of 15.5 months (range, 3.5 months-3.4 years), 90% of patients had both resolution of vaginal bulging or prolapse symptoms and improvement of the stage of prolapse. There were improvements in all pelvic organ prolapse quantitation measurements except for total vaginal length, for which the median decrease was 0.75 cm. Intraoperatively, ureteral occlusion was noted in 11% (5/46) of patients with universal cystoscopy. In 3 patients the uterosacral suspension sutures were removed and replaced with resolution of the occlusion and in 2 patients ureteral reimplantation was required. Symptomatic prolapse (2 apical segment, 1 anterior, and 1 posterior) developed in 4 patients (10%), and 3 of them underwent reoperation. There were significant improvements in symptoms of bulging and pressure, voiding dysfunction, and vaginal and perineal splinting. CONCLUSION: Suspension of the vaginal vault to the proximal uterosacral ligaments combined with site-specific repair of endopelvic fascia defects provides excellent anatomic and functional correction of pelvic organ prolapse in most women. The risk of ureteral injury with this technique makes intraoperative cystoscopy essential. PMID- 11120504 TI - Induction of ovulation in infertile women with hyperandrogenism and insulin resistance. AB - The polycystic ovary syndrome is a common cause of anovulatory infertility. Women with severe insulin resistance are a unique subset of polycystic ovary syndrome. The syndrome of hyperandrogenism, insulin resistance, and acanthosis nigricans (HAIR-AN syndrome) is one presentation of the insulin-resistant subset of polycystic ovary syndrome. Insulin resistance and hyperandrogenism are caused by genetic and environmental factors. In women with anovulatory infertility caused by hyperandrogenism and insulin resistance, clomiphene citrate treatment often fails to result in pregnancy. For these women, weight loss and insulin sensitizers can be effective methods of inducing ovulation and pregnancy and may reduce the number of clomiphene-resistant women with polycystic ovary syndrome who are treated with gonadotropins, ovarian surgery, or in vitro fertilization embryo transfer. PMID- 11120505 TI - Negative mood changes during hormone replacement therapy: a comparison between two progestogens. AB - OBJECTIVE: The aim of this study was to compare side effects of medroxyprogesterone acetate and norethindrone acetate during postmenopausal hormone replacement therapy in women with and without a history of premenstrual syndrome. STUDY DESIGN: Fifty-one postmenopausal women were randomly selected in a double-blind crossover study. The women received 2 mg of estradiol continuously during five 28-day cycles and 10 mg of medroxyprogesterone or 1 mg of norethindrone sequentially for 12 days of each cycle. Daily symptom rating scales were kept. RESULTS: The women showed cyclic changes, with negative mood and physical symptoms culminating during the late progestogen phase and positive mood during the estrogen-only phase. Symptoms declined with time but remained after 5 months. Women with a history of premenstrual syndrome responded strongly to both progestogens. Medroxyprogesterone acetate induced less negative and more positive mood symptoms than norethindrone in women with no history of premenstrual syndrome. In both groups medroxyprogesterone caused more physical symptoms than norethindrone. CONCLUSION: The addition of medroxyprogesterone to estrogen is preferable to norethindrone with respect to mood symptoms in women without a history of premenstrual syndrome. PMID- 11120506 TI - Patients' outlook, experience, and satisfaction with hysterectomy, bilateral oophorectomy, and subsequent continuation of hormone replacement therapy. AB - OBJECTIVE: Our purposes were to investigate patients' opinions of hysterectomy, bilateral oophorectomy, and hormone replacement therapy and to evaluate whether their outlook and experience influenced the overall satisfaction and continuation of hormone replacement therapy. STUDY DESIGN: We conducted a questionnaire survey of 200 patients before and 2 years after hysterectomy with or without bilateral oophorectomy. Postoperatively all patients received long-term estradiol and testosterone replacement. The inquiries of patients' views included (1) preoperative awareness of indication and outlook, (2) postoperative recovery, symptom relief, and experiences with hormone replacement therapy, (3) perceived benefits and problems, (4) changes in physical well-being, psychologic state, and sexual activity, (5) continuation of hormone replacement therapy, and (6) overall satisfaction. RESULTS: The outlook toward hysterectomy, bilateral oophorectomy, and hormone replacement therapy was positive in 77.4%, 87.1%, and 76.3%, respectively. The experience was positive in the majority, with a satisfactory postoperative recovery (70.6%), complete symptom relief (77.9%), and minimal side effects with hormone replacement therapy (5.2%). The benefits included improved physical well-being (79.9%), lower depressive symptoms (32.0%), and better sexuality (31.4%). The continuation rate of hormone replacement therapy was 97.4%, and overall satisfaction was positive in 88.7% of patients. The independent predictors of satisfaction were outlook toward hysterectomy and incomplete symptom relief. CONCLUSION: The outcome of hysterectomy, bilateral oophorectomy, and hormone replacement therapy was satisfactory to most patients. PMID- 11120507 TI - Asymptomatic bacterial vaginosis: response to therapy. AB - OBJECTIVE: Bacterial vaginosis causes symptomatic vaginal discharge and has been associated with preterm birth and with the acquisition of human immunodeficiency virus. Half of all women with bacterial vaginosis are free of symptoms, and treatment of these women is controversial. The objective of this study was to determine the extent of poor symptom recognition in this group of women. STUDY DESIGN: Seventy-five women attending a sexually transmitted disease clinic who had asymptomatic bacterial vaginosis were entered into a randomized, double blind, placebo-controlled trial comparing metronidazole gel with placebo. Subjects' perceptions about changes in vaginal discharge and odor were determined, and treatment and placebo groups were compared by means of standard statistical analysis. RESULTS: When subjects were stratified by treatment group, there were no differences in their retrospective assessments of vaginal discharge and odor. A subset of women who had normalization of clinical parameters or of Gram stain scores did admit retrospectively to improvement; however, the difference between this group and the group without normalization was not statistically significant. Twenty-one percent of treated women subsequently had vaginal candidiasis. CONCLUSIONS: A greater percentage of women with resolution of bacterial vaginosis did retrospectively notice improvement in vaginal discharge and odor in comparison with those women without resolution; however, this was not statistically significant. These findings do not support routine treatment of women with asymptomatic bacterial vaginosis. PMID- 11120508 TI - Patient satisfaction with results of hysterectomy. AB - OBJECTIVE: The objectives of this study were to measure patient satisfaction with the results of hysterectomy and to determine factors associated with dissatisfaction. STUDY DESIGN: A total of 1299 women who underwent hysterectomy at 28 hospitals in Maryland were interviewed before and at 3, 6, 12, 18, and 24 months after the operation. RESULTS: At 12 and 24 months after the hysterectomy 95.8% and 96.0%, respectively, reported that the hysterectomy had completely or mostly resolved the problems or symptoms they had before surgery; 93.3% and 93.7%, respectively, reported that the results were better than or about what they expected; 85.3% and 81. 6%, respectively, reported that their health was better than before the hysterectomy; and 87.9% and 93.1%, respectively, reported being totally recovered. The factor most strongly and consistently associated with patient reports of negative outcomes was readmission because of a postdischarge complication. CONCLUSION: Postdischarge complication necessitating readmission plays an important role in patient dissatisfaction with the results of hysterectomy. PMID- 11120509 TI - Comparability of perioperative morbidity between abdominal myomectomy and hysterectomy for women with uterine leiomyomas. AB - OBJECTIVE: The aim of this study was to compare the perioperative morbidity associated with abdominal myomectomy with that of hysterectomy. STUDY DESIGN: This was a retrospective cohort study of 394 women at an academic medical center. Main outcome measured was perioperative morbidity, with the following secondary outcomes: febrile morbidity, hemorrhage, unintended major surgical procedures, life-threatening events, and rehospitalization. RESULTS: Morbidity was associated with myomectomy and hysterectomy in 39% and 40% of cases, respectively. The crude odds ratio for morbidity of myomectomy with respect to hysterectomy was 0.93 (95% confidence interval, 0.63-1.40). Women who underwent myomectomy were significantly younger, weighed less, and had a smaller preoperative uterine size. In a multivariable analysis that accounted for these differences the odds ratio increased to 1.46 (95% confidence interval, 0.77-2.77) but still was not statistically elevated. The study had >90% power to detect a clinically relevant 15% absolute difference in overall morbidity between the 2 groups. CONCLUSION: No clinically significant difference in perioperative morbidity between myomectomy and hysterectomy was detected. Myomectomy should be considered a safe alternative to hysterectomy. PMID- 11120510 TI - Continuous combined hormone replacement therapy and risk of endometrial cancer. AB - OBJECTIVE: Postmenopausal women who receive sequential hormone replacement therapy with estrogen combined with progestogen for 10 to 24 d/mo for a prolonged period may have an elevated endometrial cancer risk relative to those who have never received hormone replacement therapy. We investigated whether daily use of estrogen and progestogen (continuous combined hormone replacement therapy) could diminish any excess endometrial cancer risk. STUDY DESIGN: A population-based study in Washington State obtained interview data from 969 women aged 45 to 74 years with endometrial cancer diagnosed during 1985 through 1991 or 1994 through 1995 and from 1325 age-matched control subjects selected primarily by random digit dialing. Women who had received only continuous combined hormone replacement therapy were compared with women who had only received another hormone replacement therapy regimen or who had never received hormone replacement therapy. RESULTS: The risk of endometrial cancer among users of continuous combined hormone replacement therapy (n = 9 case patients, n = 33 control subjects) relative to women who had never received hormone replacement therapy was 0.6 (95% confidence interval, 0.3-1.3); the risk relative to women who received hormone replacement that included progestogen for 10 to 24 d/mo was 0.4 (95% confidence interval, 0.2-1.1). Most continuous combined hormone replacement therapy use was short-term (<72 months) or recent (in the previous 24 months). CONCLUSION: Women who had received continuous combined hormone replacement therapy for several years did not appear to be at any increased risk for endometrial cancer relative to women who had never received hormone replacement therapy and may in fact be at decreased risk for endometrial cancer. PMID- 11120511 TI - Gonadotropin-releasing hormone agonist treatment for endometriosis of the rectovaginal septum. AB - OBJECTIVE: This study was undertaken to evaluate the effectiveness of a 6-month course of gonadotropin-releasing hormone agonist treatment for patients with symptomatic endometriosis of the rectovaginal septum. STUDY DESIGN: Fifteen patients with rectovaginal endometriosis and moderate to severe pain symptoms were the subjects of the study. None of these patients had either clinical or objective evidence of ovarian endometriosis, nor was there evidence of any obstructive lesions of the intestine or ureters. All patients were given leuprolide acetate depot at 3.75 mg, 1 ampule intramuscularly every 28 days, and treatment had a planned duration of 6 months. Follow-up evaluations were set every 2 months during the treatment phase and every 3 months thereafter until the completion of 1 year after discontinuation of medical therapy. At each follow-up visit pain symptoms were recorded, and clinical exploration, transvaginal ultrasonography, and transrectal ultrasonography were performed. RESULTS: Two patients stopped the treatment early after the second and fourth leuprolide doses; in both cases the reason was persistence of pain, and both requested a surgical solution. The other 13 patients showed a marked improvement with respect to pain during the 6-month treatment course but had early pain recurrence after drug suspension; 11 of them required further treatment within the first year of follow-up. The failure rate of gonadotropin-releasing hormone agonist therapy to produce 1-year pain relief after treatment discontinuation was 87% (13/15) on an intent-to-treat basis. The endometriotic lesions showed a slight but significant reduction in size during therapy but had returned to the original volume within 6 months after cessation of the gonadotropin-releasing hormone analog treatment. CONCLUSION: Our results suggest that gonadotropin-releasing hormone analogs should not be considered a real therapeutic alternative to surgical treatment for patients with symptomatic endometriosis of the rectovaginal septum, except possibly in a limited and unpredictable number of cases. PMID- 11120512 TI - 21-Hydroxylase-deficient nonclassic adrenal hyperplasia is a progressive disorder: a multicenter study. AB - OBJECTIVE: Our aim was to determine whether the clinical features of 21 hydroxylase-deficient nonclassic adrenal hyperplasia are correlated with either age at symptom onset or age at presentation, or both, and with the degree of adrenocortical abnormality. STUDY DESIGN: In a multicenter cohort design 220 women with nonclassic adrenal hyperplasia, with a basal or adrenocorticotropic hormone-stimulated 17-hydroxyprogesterone level >30.3 nmol/L, were studied, either prospectively (n = 39) or retrospectively (n = 181). Patients were stratified by age of presentation into 5 groups: (1) <10 years (n = 25), (2) 10 to 19 years (n = 64), (3) 20 to 29 years (n = 83), (4) 30 to 39 years (n = 30), and (5) 40 to 49 years (n = 16). Two patients >50 years old were excluded from the analysis because of age. RESULTS: Ninety-two percent of patients <10 years old had premature pubarche at presentation, whereas clitoromegaly and acne were each present in only 20% of these younger subjects. With only patients > or =10 years old considered, presenting clinical features included hirsutism (59%), oligomenorrhea (54%), acne (33%), infertility (13%), clitoromegaly (10%), alopecia (8%), primary amenorrhea (4%), and premature pubarche (4%). Among the patients >/=10 years old, the prevalence but not the degree of hirsutism increased significantly with age. Basal levels of 17-hydroxyprogesterone in adolescents were significantly higher than the levels found either in children (<10 years old) or women 40 to 49 years old (P <.01 and P <.03, respectively), although no difference was noted in the stimulated 17-hydroxyprogesterone levels between age groups. The adrenocorticotropic hormone-stimulated levels but not the basal levels of 17-hydroxyprogesterone were significantly higher in patients with clitoromegaly than in women without clitoromegaly. Alternatively, there were no differences in either basal or stimulated 17-hydroxyprogesterone levels between patients with and those without hirsutism, acne, or alopecia. CONCLUSION: In children <10 years old the most common presenting complaint was premature pubarche, whereas hirsutism and oligomenorrhea were more common in older patients. The prevalence of hirsutism increased with age, suggesting the progressive nature of nonclassic adrenal hyperplasia. Furthermore, the adrenocorticotropic hormone-stimulated levels of 17-hydroxyprogesterone were higher in patients with clitoromegaly, which suggests that the degree of adrenocortical dysfunction in nonclassic adrenal hyperplasia determines, at least in part, the clinical presentation. PMID- 11120513 TI - HELLP (hemolysis, elevated liver enzymes, and low platelet count) syndrome versus severe preeclampsia: onset at < or =28.0 weeks' gestation. AB - OBJECTIVE: Our purpose was to determine whether the onset of the HELLP (hemolysis, elevated liver enzymes, and low platelet count) syndrome in women at < or =28.0 weeks' gestation is associated with an increased risk of adverse maternal and perinatal outcomes in comparison with the risk for women with severe preeclampsia but without the HELLP syndrome at a similar gestational age. STUDY DESIGN: Sixty-four patients with either the HELLP syndrome (n = 32) or severe preeclampsia but absent HELLP syndrome laboratory test results (n = 32), admitted at < or =28.0 weeks' gestation between July 1, 1992, and April 30, 1999, were studied. Maternal and perinatal outcomes were compared between the 2 groups. Statistical analysis was performed by the Student t test and the Fisher exact test. RESULTS: There were no significant differences between the 2 groups regarding African-American race (59% vs 75%), nulliparity (50% vs 56%), or the use of corticosteroids (59% vs 78%). There were no maternal deaths. One woman with the HELLP syndrome had a liver hematoma. The rate at which transfusion of blood products was required was significantly greater in women with the HELLP syndrome than in those with severe preeclampsia only (25% vs 3%; P <.05). There were no significant differences between the 2 groups with respect to eclampsia (16% vs 13%), abruptio placentae (6% vs 9%), disseminated intravascular coagulopathy (13% vs 0%), pulmonary edema (13% vs 6%), acute renal failure (3% vs 0%), pleural effusion (3% vs 3%), or ascites (6% vs 16%). No significant differences were found between the 2 groups with respect to neonatal death (11% vs 17%), respiratory distress syndrome (78% vs 86%), or composite neonatal morbidity. CONCLUSIONS: Except for the need for transfusion of blood products in women with the HELLP syndrome, onset at < or =28.0 weeks' gestation is not associated with an increased risk of adverse maternal or neonatal outcomes in comparison with the risk for women with severe preeclampsia but without the HELLP syndrome at a similar gestational age. PMID- 11120514 TI - The preterm prediction study: quantitative fetal fibronectin values and the prediction of spontaneous preterm birth. The National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network. AB - OBJECTIVE: A cervicovaginal fetal fibronectin value of >/=50 ng/mL has been used to define women at risk of having a preterm birth. We evaluated the relationship between quantitative fetal fibronectin values and spontaneous preterm birth. STUDY DESIGN: Cervical and vaginal specimens for fetal fibronectin were obtained at 24, 26, 28, and 30 weeks' gestation from 2926 women. Quantitative fetal fibronectin values were calculated by using absorbances determined by enzyme linked immunosorbent assay. The highest fetal fibronectin value (cervical or vaginal) for each woman at each visit was evaluated in relation to spontaneous preterm birth at <35 weeks' gestation. Receiver operating characteristic curves were constructed to determine the optimal cutoff point for fetal fibronectin values to predict spontaneous preterm birth at <35 weeks' gestation and within 4 weeks of testing. RESULTS: The risk of spontaneous preterm birth increased as a function of increasing fetal fibronectin values from approximately 20 to 300 ng/mL. Fetal fibronectin values > or =300 ng/mL were not associated with a further increase in spontaneous preterm birth. Examination of the receiver operating characteristic curve indicates that the optimal cutoff point for a positive fetal fibronectin test result at 24 to 30 weeks' gestation to predict spontaneous preterm birth at <35 weeks is between 45 and 60 ng/mL. CONCLUSION: Increasing levels of cervicovaginal fetal fibronectin up to 300 ng/mL are associated with an increasing risk of spontaneous preterm birth. Nevertheless, at 24 to 30 weeks, the value currently used, 50 ng of fetal fibronectin per milliliter, appears to be a reasonable cutoff point for predicting spontaneous preterm birth at <35 weeks' gestation. PMID- 11120515 TI - Beginning regular exercise in early pregnancy: effect on fetoplacental growth. AB - OBJECTIVE: Our purpose was to test the null hypothesis that beginning regular, moderate-intensity exercise in early pregnancy has no effect on fetoplacental growth. STUDY DESIGN: Forty-six women who did not exercise regularly were randomly assigned at 8 weeks either to no exercise (n = 24) or to weight-bearing exercise (n = 22) 3 to 5 times a week for the remainder of pregnancy. Outcome variables included antenatal placental growth rate and neonatal and placental morphometric measurements. RESULTS: The offspring of the exercising women were significantly heavier (corrected birth weight: 3.75 +/- 0.08 kg vs 3.49 +/- 0.07 kg) and longer (51.8 +/- 0.3 cm vs 50.6 +/- 0.3 cm) than those born to control women. The difference in birth weight was the result of an increase in both lean body mass and fat mass. In addition, midtrimester placental growth rate was faster (26 +/- 2 cm(3)/wk vs 21 +/- 1 cm(3)/wk) and morphometric indexes of placental function were greater in the exercise group. There were no significant differences in neonatal percentage body fat, head circumference, ponderal index, or maternal weight gain. CONCLUSIONS: These data indicate that beginning a moderate regimen of weight-bearing exercise in early pregnancy enhances fetoplacental growth. PMID- 11120516 TI - Vascular reactivity in patients with preeclampsia and HELLP (hemolysis, elevated liver enzymes, and low platelet count) syndrome. AB - OBJECTIVE: Early structural and functional changes in the systemic vasculature have been proposed to play a major pathogenetic role in preeclampsia and HELLP (hemolysis, elevated liver enzymes, and low platelet count) syndrome. The aim of the study was to determine vascular reactivity in patients with preeclampsia with and without HELLP syndrome with respect to those in healthy pregnant control subjects. STUDY DESIGN: Forearm blood flow was measured by strain gauge plethysmography with the venous occlusion technique in 12 hypertensive patients with HELLP syndrome, in 8 patients with preeclampsia without HELLP syndrome, and in 8 healthy normotensive pregnant control subjects. To determine vascular reactivity the forearm blood flow was measured at baseline and after forearm occlusion for a period of 5 minutes (reactive hyperemia). The investigations were repeated 4 to 6 months post partum. Forearm vascular resistance was calculated as the ratio of mean arterial pressure to forearm blood flow. RESULTS: Mean arterial pressure at rest was elevated in patients with preeclampsia (116 +/- 20 mm Hg) and in patients with HELLP syndrome (110 +/- 16 mm Hg) with respect to healthy pregnant control subjects (86 +/- 10 mm Hg; P <.05). Forearm blood flow at rest was not statistically different in patients with preeclampsia (5.1 +/- 2.6 mL/min per 100 mL) and with HELLP syndrome (4.7 +/- 1.5 mL/min per 100 mL) with respect to pregnant control subjects (5.9 +/- 3.1 mL/min per 100 mL); however, forearm vascular resistance at rest was elevated in patients with preeclampsia (25.9 +/- 9.5 units; P <.05) and in patients with HELLP syndrome(24.6 +/- 6.9 units; P <.05) with respect to healthy control subjects (17.0 +/- 6.1 units). During reactive hyperemia the peak forearm blood flow, which is an indicator of maximal vasodilatory capacity, was impaired in patients with preeclampsia (21.9 +/- 8.2 mL/min per 100 mL; P <.05) but not in patients with HELLP syndrome (37.4 +/- 17.5 mL/min per 100 mL) and healthy control subjects (44.9 +/- 15.0 mL/min per 100 mL). Consequently, minimum forearm vascular resistance was higher in women with preeclampsia (6.1 +/- 1.9 units) than in both women with HELLP syndrome (3.5 +/- 1.6 units) and the control subjects (2.8 +/- 2.4 units). CONCLUSION: Despite similarly elevated forearm vascular resistances at rest in patients with HELLP syndrome and in patients with preeclampsia, forearm vascular resistance during reactive hyperemia did not differ significantly from that in healthy control subjects in the women with HELLP syndrome but was increased in women with preeclampsia. Vasodilatory reactivity thus is reduced in preeclampsia but not in HELLP syndrome, which suggests different alterations of the vasculature. PMID- 11120517 TI - Placental expression of syndecan 1 is diminished in preeclampsia. AB - OBJECTIVE: Syndecan 1 is a cell surface heparan sulfate proteoglycan that binds growth factors and antithrombin III. The objective of this study was to examine whether placental expression of syndecan 1 in preeclampsia differs from that in normal pregnancy and whether gestational age and fetal growth affect syndecan 1 expression. STUDY DESIGN: An immunohistochemical analysis of 30 placentas of women with severe preeclampsia and 15 placentas of women without preeclampsia was performed with the monoclonal anti-syndecan 1 antibody B-B4. RESULTS: In 47% of preeclamptic placentas the immunoreactivity with antibody B-B4 was faint or absent, whereas 93% of the normal placentas exhibited strong immunoreactivity. The reduction in placental expression of syndecan 1 in preeclampsia was not associated with gestational age or impaired fetal growth. CONCLUSION: The expression of syndecan 1 on the chorionic villi is reduced in preeclampsia irrespective of gestational age or fetal growth. PMID- 11120518 TI - Delayed-interval delivery: extended series from a single maternal-fetal medicine practice. AB - OBJECTIVE: Our purpose was to review the extended experience of a single maternal fetal medicine practice with delayed-interval delivery. STUDY DESIGN: We completed a retrospective review of our maternal-fetal medicine practice database from January 1991 through March 1999. Patients were derived from both primary and consultative practices. All patients were managed with tocolysis, antibiotics, and cerclage after delivery of the first fetus(es). Retained siblings were investigated by amniocentesis to exclude intra-amniotic infection. RESULTS: Twenty-four consecutive patients had attempted delayed-interval delivery. Exclusion criteria for delayed-interval delivery included monochorionicity, abruptio placentae, severe preeclampsia, and the need for hysterotomy. The mean latency interval was 36 days, with a range of 3 to 123 days. Additionally, patients with previous cerclage(s) had significantly shorter mean latency intervals than patients without previous cerclage(s). Patients with long latency intervals (> or =49 days) had earlier births of the first fetus. CONCLUSION: Selected multichorionic pregnancies may benefit from delayed-interval delivery. Patients with previous cervical cerclage(s) during the index pregnancy are less likely to achieve significant latency intervals. Even modest intervals between births of siblings at critical gestational ages can improve neonatal survival and decrease neonatal morbidity. PMID- 11120519 TI - Association between level of delivery hospital and neonatal outcomes among South Carolina Medicaid recipients. AB - OBJECTIVE: The purpose of this study was to examine relationships between level of delivery hospital and neonatal mortality rate, length of stay, and Medicaid reimbursement. STUDY DESIGN: This was a retrospective cohort study of 2560 infants with birth weights between 500 and 1499 g who were born between 1991 and 1995 to South Carolina mothers and whose care was covered by Medicaid. RESULTS: The relative risk of neonatal death for infants born in level I and II hospitals (relative risk, 1.9; 95% confidence interval, 1.6-2.2) but not level II hospitals with 24-hour neonatology coverage (relative risk, 1.2; 95% confidence interval, 0.7-1.9) was higher than that for infants born in level III hospitals. Compared with infants born in level III hospitals mean length of stay was longer and Medicaid reimbursement was similar for infants born in level I and II hospitals. Among infants born in level II hospitals with 24-hour neonatology coverage length of stay was shorter and Medicaid reimbursement was lower. CONCLUSION: Infants born in level I and II hospitals had higher neonatal mortality rates and longer stays than did infants born in level III hospitals, despite similar Medicaid reimbursement. PMID- 11120520 TI - Differential alterations in responsiveness in particulate and soluble guanylate cyclases in pregnant guinea pig myometrium. AB - OBJECTIVE: The mechanism underlying myometrial quiescence during pregnancy is unknown. Our group has previously shown that during pregnancy myometrial cyclic guanosine monophosphate content rises to several hundred times the nonpregnant levels, only to abruptly decline days before the onset of labor. Cyclic guanosine monophosphate plays an integral role in the relaxation of smooth muscle. The aim of this investigation was therefore to determine the effects of pregnancy on both soluble and particulate guanylate cyclase enzymatic activities and messenger ribonucleic acid expressions. STUDY DESIGN: Myometrium was obtained from randomly cycling adult nonpregnant guinea pigs and near-term (50-60 days' gestation) pregnant guinea pigs of similar chronologic age. Subcellular fractions were prepared by differential ultracentrifugation. Guanylate cyclase activity was determined by the conversion of guanosine triphosphate to cyclic guanosine monophosphate under basal or stimulated conditions in either the soluble guanylate cyclase or particulate guanylate cyclase fraction. A nitric oxide donor, S-nitroso- N-penacillamine, was used to activate soluble guanylate cyclase (n = 10 animals in each group). Several natriuretic peptides (atrial natriuretic peptide, brain natriuretic peptide, and C-type natriuretic peptide) and uroguanylin were used to stimulate the different particulate guanylate cyclase isoforms guanylate cyclase A, guanylate cyclase B, and guanylate cyclase C, respectively, in pregnant (n = 8) and nonpregnant (n = 6) animals. Cyclic guanosine monophosphate content was measured by radioimmunoassay, and enzymatic activity was expressed as picomoles of cyclic guanosine monophosphate per milligram of protein per minute. Total guanylate cyclase represented the sum of soluble guanylate cyclase and particulate guanylate cyclase activities for a tissue. To investigate whether the observed changes in guanylate cyclase activity were paralleled by changes in receptor expression, messenger ribonucleic acid levels of the genes for guanylate cyclase A and guanylate cyclase B isoforms were quantified by ribonuclease protection assay (n = 5 animals in each group). RESULTS: Under basal conditions particulate guanylate cyclase represented 78% (nonpregnant state) to 88% (during pregnancy) of the total guanylate cyclase activity in the guinea pig myometrium. Pregnancy further reduced myometrial soluble guanylate cyclase (both basal and stimulated by nitric oxide) relative to the nonpregnant state. Pregnancy selectively increased atrial natriuretic peptide stimulated particulate guanylate cyclase activity (attributed to guanylate cyclase A), although it did not change basal myometrial particulate guanylate cyclase activity in general. Guanylate cyclase B (particulate guanylate cyclase stimulated by C-type natriuretic peptide) and guanylate cyclase C (particulate guanylate cyclase stimulated by uroguanylin) activities were unaltered by pregnancy. The selective increase in responsiveness of particulate guanylate cyclase to atrial natriuretic peptide during pregnancy was not paralleled by an increased in level of messenger ribonucleic acid for the gene for guanylate cyclase A. CONCLUSION: Pregnancy reduced the in vitro responsiveness of the myometrial soluble guanylate cyclase to nitric oxide while increasing the responsiveness of the particulate isoform to atrial natriuretic peptide and brain natriuretic peptide through a mechanism independent of any change in receptor expression. PMID- 11120521 TI - Preterm delivery in women with pregestational diabetes mellitus or chronic hypertension relative to women with uncomplicated pregnancies. The National institute of Child health and Human Development Maternal- Fetal Medicine Units Network. AB - OBJECTIVE: The purpose of this study was to compare the rates of indicated and spontaneous preterm delivery among women with chronic hypertension or pregestational diabetes mellitus with the rates among healthy women. STUDY DESIGN: This was a secondary analysis of data from healthy women with singleton gestations enrolled in a prospective observational study for prediction of preterm delivery (control group, n = 2738), women with pregestational diabetes mellitus requiring insulin therapy (n = 461), and women with chronic hypertension (n = 761). The two latter groups were enrolled in a randomized multicenter trial for prevention of preeclampsia. The main outcome measures were rates of preterm delivery, either spontaneous (preterm labor or rupture of membranes) or indicated (for maternal or fetal reasons), and neonatal outcomes. RESULTS: The overall rates of preterm delivery were significantly higher among women with diabetes mellitus (38%) and hypertension (33.1%) than among control women (13.9%). Rates were also significantly higher for delivery at <35 weeks' gestation. Women with diabetes mellitus had significantly higher rates of both indicated preterm delivery (21.9% vs 3.4%; odds ratio, 8.1; 95% confidence interval, 6.0-10.9) and spontaneous preterm delivery (16.1% vs 10.5%; odds ratio, 1.6; 95% confidence interval, 1.2-2.2) than did women in the control group. In addition, they had significantly higher rates of both indicated preterm delivery (odds ratio, 4.8; 95% confidence interval, 3.0-7.5) and spontaneous preterm delivery (odds ratio, 2.1; 95% confidence interval, 1.4-3.0) at <35 weeks' gestation than did control women. Compared with control women those with chronic hypertension had higher rates of indicated preterm delivery at both <37 weeks' gestation (21.9% vs 3.4%; odds ratio, 8.1; 95% confidence interval, 6.2-10.6) and at <35 weeks' gestation (12.1% vs 1.6%; odds ratio, 8.2; 95% confidence interval, 5.7-11.9), but there were no differences in rates of spontaneous preterm delivery. CONCLUSION: The increased rate of preterm delivery among women with chronic hypertension relative to control women was primarily an increase in indicated preterm delivery, whereas the rates of both spontaneous and indicated preterm delivery were increased among women with pregestational diabetes mellitus. PMID- 11120522 TI - Expression of protein kinase C isozymes in nonpregnant and pregnant human myometrium. AB - OBJECTIVE: The aim of this study was to compare the distributions of protein kinase C isozymes in human nonpregnant and pregnant myometrial tissues and primary cell cultures. STUDY DESIGN: Myometrial tissues were obtained at hysterectomy from nonpregnant women and at cesarean delivery from women both before and during early labor at term. Western immunoblot analysis was performed on homogenates of myometrial tissues and primary cell cultures with monoclonal antibodies specific for protein kinase C isozymes. Redistribution and translocation of protein kinase C were examined by means of immunocytochemical methods. RESULTS: Nonpregnant myometrial tissues contained protein kinase C isozymes alpha, gamma, delta, mu, iota, and zeta but not beta(1), beta(2), theta, or epsilon. Pregnant myometrial tissues both before and during early labor contained the same protein kinase C isozymes and also beta(1) and beta(2). The protein kinase C isozyme distribution in primary myometrial cell cultures was identical to that in the myometrial tissues. Protein kinase C redistribution and translocation were demonstrated in these cultured myometrial cells. CONCLUSION: Both human myometrial tissues and primary cell cultures expressed a broad range of protein kinase C isozymes. Protein kinase C isozymes beta(1) and beta(2) were absent in nonpregnant myometrium but were induced during pregnancy. Labor at term did not alter protein kinase C isozyme expression. PMID- 11120523 TI - Association of ABO incompatibility with elevation of nucleated red blood cell counts in term neonates. AB - OBJECTIVE: Nucleated red blood cells in the circulation in term neonates have been associated with a wide range of pathologic conditions. We sought to examine the relationship between nucleated red blood cells in the circulation of term neonates and maternal-neonatal blood type compatibility. STUDY DESIGN: We prospectively collected umbilical blood from all live-born neonates delivered at our institution. Venous blood was analyzed for nucleated red blood cells and is reported as the number of nucleated red blood cells per 100 white blood cells. We reviewed maternal and neonatal records for neonates born at > or =37 weeks' gestation for correlative clinical data. Statistical analysis was performed with the SAS statistical software package (version 6.12; SAS Institute, Inc, Cary, NC). Kruskal-Wallis analysis was used as a nonparametric test. RESULTS: We evaluated 1661 neonates delivered during the study period and found a mean (+/ SD) of 9.29 +/- 18.56 nucleated red blood cells per 100 white blood cells (range, 0-327 nucleated red blood cells per 100 white blood cells). Nucleated red blood cell counts were lower in ABO-compatible maternal-fetal dyads (mean +/- SD, 8.29 +/- 12.84 nucleated red blood cells per 100 white blood cells; range, 0-216 nucleated red blood cells per 100 white blood cells) than in ABO-incompatible dyads (mean +/- SD, 13.16 +/- 13.16 nucleated red blood cells per 100 white blood cells; range, 0-327 nucleated red blood cells/100 white blood cells; P =.006). Neonates of mothers with blood groups A and B had significantly lower nucleated red blood cell counts (P <.05). Dyads with maternal type O and neonate type B had significantly higher nucleated red blood cell counts (P <.002). Nonparametric testing determined that type O mother and type B neonate combinations had significantly higher umbilical cord nucleated red blood cell counts (P <.001). CONCLUSION: Maternal-fetal ABO incompatibility is associated with elevation of nucleated red blood cell count in term neonates. Nucleated red blood cell elevation does not always connote a serious pathologic process, however, because ABO incompatibility usually does not adversely affect neonatal outcome. The clinical significance of an elevated nucleated red blood cell count thus is limited. PMID- 11120524 TI - Elevations of group II phospholipase A2 concentrations in serum and amniotic fluid in association with preterm labor. AB - OBJECTIVE: The purpose of this study was to determine whether the elevation of secretory group II phospholipase A(2) concentration in the serum and amniotic fluid in preterm labor is associated with intrauterine inflammation. STUDY DESIGN: Serum and amniotic fluid were collected from women with preterm delivery (<37 weeks' gestation; n = 38) and term delivery (n = 20). Phospholipase activity was measured with a highly sensitive system that was based on reverse-phase high performance liquid chromatographic separation of 9-anthracenylmethyl derivatives of fatty acids released by phospholipase A(2). The concentrations of immunoreactive isozymes (group I or II) of secretory phospholipase A(2) were assayed with a radioimmunoassay kit with a monoclonal antibody against human pancreatic phospholipase A(2) and splenic IIA phospholipase A(2). Localization of immunoreactive group II phospholipase A(2) in the amniotic membrane was determined by immunostaining visualized with the Vectastain ABC (Vector Laboratories, Inc, Burlingame, Calif) method. RESULTS: Enzymatic activities of phospholipase A(2) in the serum and amniotic fluid specimens obtained from patients in preterm labor with chorioamnionitis were significantly higher than those in specimens from patients in term labor. Significant elevations of phospholipase A(2) activities were observed in patients with preterm labor without histologically evident chorioamnionitis. The activity of phospholipase A(2) was clearly correlated with the concentration of the immunoreactive group II phospholipase A(2). Group II phospholipase A(2) was localized in amniotic cells obtained from patients with a pathologically determined diagnosis of chorioamnionitis. The predictive value for chorioamnionitis of the group II phospholipase A(2) concentration was relatively higher than the predictive values of the concentrations of C-reactive protein and interleukins 6 and 8. CONCLUSION: Significant elevations of group II phospholipase A(2) concentrations were detected in the serum and amniotic fluid of women with preterm labor. Group II phospholipase A(2) concentration may be a useful indicator for preterm labor, and phospholipid metabolism is certainly activated both in preterm labor and in apparent inflammatory diseases. PMID- 11120525 TI - A randomized controlled trial of the effect of increased intravenous hydration on the course of labor in nulliparous women. AB - OBJECTIVE: One variable that has the potential to affect the course of labor but has not been evaluated previously is the adequacy of maternal hydration. Typical orders provide for 125 mL of intravenous fluids per hour in patients taking limited oral fluids. Many such patients are clinically dehydrated. Physiologists have shown that increased fluids improve skeletal muscle performance in prolonged exercise. This study was designed to determine whether increased intravenous fluids affect the progress of labor. STUDY DESIGN: Nulliparous women with uncomplicated singleton gestations at term, in spontaneous active labor with dilatation between 2 and 5 cm, and with a cephalic presentation were included. Patients who gave consent were randomly selected to receive either 125 mL or 250 mL of intravenous fluids per hour. RESULTS: One hundred ninety-five patients were randomly selected, 94 to the 125-mL group and 101 to the 250-mL group. Prerandomization variables were well matched between the 2 groups. The mean volume of total intravenous fluids was significantly greater in the 250-mL group (2008 mL vs 2487 mL; P =.002), as was the mean hourly rate (152 mL/h in the 125 mL group vs 254 mL/h in the 250-mL group; P =.001). The frequency of labor lasting >12 hours was statistically higher in the 125-mL group (20/78 [26%] vs 12/91 [13%]; P =.047). In addition, there was a trend favoring longer mean duration of the first stage and total duration of labor in patients delivered vaginally in the 125-mL group, by 70 and 68 minutes, respectively (P =.06). There was a trend toward a lower frequency of oxytocin administration for inadequate labor progress in the higher fluid rate group (61 [65%] in the 125-mL group vs 51 [49%] in the 250-mL group; P =.06). Cesarean deliveries were more frequent in the 125-mL group (n = 16) than in the 250-mL group (n = 10) but did not reach statistical significance. CONCLUSION: This study presents the novel finding that increasing fluid administration for nulliparous women in labor above rates commonly used is associated with a lower frequency of prolonged labor and possibly less need for oxytocin. Thus inadequate hydration in labor may be a factor contributing to dysfunctional labor and possibly cesarean delivery. Consideration of this factor in clinical management and in future studies considering variables that affect labor is warranted. PMID- 11120527 TI - Reductions of vascular endothelial growth factor and placental growth factor concentrations in severe preeclampsia. AB - OBJECTIVE: The aim of this study was to determine whether plasma concentrations of vascular endothelial growth factor and placental growth factor are altered in women with severe preeclampsia. STUDY DESIGN: We performed a case-control study to compare plasma concentrations of vascular endothelial growth factor and placental growth factor between women with severe preeclampsia and normotensive women admitted for delivery. Twenty-one women with severe preeclampsia were matched for gestational age and ethnicity with 21 normotensive women. Vascular endothelial growth factor and placental growth factor concentrations were measured with a specific antigen-capture enzyme-linked immunosorbent assay. RESULTS: Women with severe preeclampsia demonstrated significantly lower plasma concentrations of both vascular endothelial growth factor (6.36 +/- 3.96 pg/mL vs 18.65 +/- 5.98 pg/mL; P <.0001) and placental growth factor (138 +/- 119 pg/mL vs 531 +/- 340 pg/mL; P <.0001) than did women with normotensive pregnancy. Logistic regression analysis showed an independent association between plasma vascular endothelial growth factor concentration and plasma placental growth factor concentration and preeclampsia. CONCLUSION: Patients with severe preeclampsia had decreased maternal serum concentrations of both vascular endothelial growth factor and placental growth factor. PMID- 11120526 TI - Assessment of fetal scalp oxygen saturation determination in the sheep by transmission pulse oximetry. AB - OBJECTIVE: Electronic fetal heart rate monitoring has an unacceptable false positive nonreassuring rate, which results in an excess of operative interventions. As a more objective measure of fetal oxygenation, fetal scalp pulse oximetry has been used to assess fetal blood oxygen saturation (SO (2)). The current devices use reflectance oximetry, which has inherent limitations. These include varying depths of signal penetration, variation with position, and potential for optical interference. In this study we evaluated a newly developed transmission pulse oximetry device consisting of transmitter and receiver diodes mounted within the coil of a standard scalp electrode (Spiral O(2)CTG; Respironics Inc, Marietta, Ga). STUDY DESIGN: Six pregnant ewes at 127 to 135 days' gestation (term, 145 days' gestation) were anesthetized, intubated, and prepared with a femoral artery catheter. Fetuses were prepared with brachial artery and jugular vein catheters. Maternal inspired oxygen fraction was titrated from 21% to 3%. Oximetry O(2)CTG devices were positioned on the fetal scalp, and recordings were compared with directly determined fetal arterial pH, PO (2), and SO (2) values. RESULTS: Maternal SaO (2) and PaO (2) ranged from 102% to 16% and 110 to 18 mm Hg, respectively. Fetal SaO (2) and PaO (2) ranged from 76% to 12% and 28 to 8 mm Hg, respectively. There was excellent correlation between direct fetal SaO (2) and scalp SO (2) (r (2) = 0.90; scalp SO (2) = 0.79 SaO (2) + 6.89). With an SaO (2) of <30% as the cutoff point for assessment of fetal compromise, scalp SO (2) measurements had a 94% +/- 10% specificity and a 94% +/- 10% positive predictive value. CONCLUSION: (1) Preliminary studies of the Spiral O(2)CTG sensor demonstrated high correlation of scalp SO (2) with fetal SaO (2). (2) Although potential inaccuracies remain, transmission oximetry may offer potential advantages in consistency, ease of application, and technology with respect to the current reflection oximeter devices. PMID- 11120528 TI - Neutrophil activation in preeclampsia and isolated intrauterine growth restriction. AB - OBJECTIVE: Neutrophils have been implicated in the pathogenesis of preeclampsia. Because preeclampsia and intrauterine growth restriction result from similar placental lesions, the aim of this study was to investigate neutrophil activation in isolated intrauterine growth restriction relative to preeclampsia and uncomplicated pregnancy. Polymorphonuclear neutrophil activation was analyzed by measuring cell surface and soluble cell adhesion molecule expressions. STUDY DESIGN: L -Selectin (CD62L ) and CD11b surface expressions on polymorphonuclear neutrophils were analyzed in 13 women with preeclampsia, 11 women with isolated intrauterine growth restriction, and 17 age- and gestation-matched control women by means of a standardized quantitative flow cytometry assay. Serum levels of soluble L -selectin were measured by enzyme-linked immunosorbent assay. RESULTS: Neutrophils from women with isolated intrauterine growth restriction and women with preeclampsia displayed higher levels of CD11b and lower levels of CD62L than did neutrophils from healthy pregnant women. Soluble L -selectin serum levels were significantly increased in the preeclampsia and intrauterine growth restriction groups relative to normal values. No significant difference in the levels of CD11b, CD62L, and soluble L -selectin were observed between women with isolated intrauterine growth restriction and those with preeclampsia. Leukocyte activation was not correlated with disease severity. CONCLUSION: The observed alteration in polymorphonuclear neutrophil adhesion molecule expressions and increased serum soluble L -selectin levels are consistent with activation of peripheral blood neutrophils occurring in isolated intrauterine growth restriction in a manner similar to that seen in preeclampsia. This evidence of neutrophil activation may help to advance our understanding of the disease process in isolated intrauterine growth restriction. PMID- 11120529 TI - Delayed hypotension and subendocardial injury after repeated umbilical cord occlusion in near-term fetal lambs. AB - OBJECTIVE: This study was undertaken to determine whether myocardial injury occurs after repeated intrauterine asphyxia. STUDY DESIGN: Near-term fetal sheep with implanted instrumentation underwent either sham occlusions (n = 8) or repeated brief umbilical cord occlusions (n = 12) continued until the onset of severe (<20 mm Hg) or sustained hypotension. After 3 days of recovery, the fetal hearts were perfusion fixed. RESULTS: Repeated umbilical cord occlusions led to a severe metabolic acidosis (pH, 6.84 +/- 0.09; lactate concentration, 14.1 +/- 1.5 mmol/L) with increasing hypotension during occlusions, which were terminated after 128 +/- 38 minutes. After the occlusions, the mean arterial pressure showed a delayed fall, which resolved after 12 hours. Ultrastructural examination showed evidence of subendocardial injury, with dilatation of sarcoplasmic reticulum, margination and clumping of nuclear chromatin, and mitochondrial swelling. The most severe morphologic changes, including electron-dense mitochondrial inclusions, were found in the fetuses with delayed recovery of the fetal heart rate after the final occlusion. CONCLUSION: Subendocardial injury occurs after severe repeated intrauterine asphyxia in the late-gestation fetus, and this may contribute to cardiovascular compromise and the development of late decelerations. PMID- 11120530 TI - Vasodilatory response of fetoplacental vasculature to adrenomedullin after constriction with the thromboxane sympathomimetic U46619. AB - OBJECTIVE: This study was undertaken to determine whether adrenomedullin, a hypotensive peptide, decreases vasomotor tone in fetoplacental vasculature that has been constricted with the thromboxane sympathomimetic U46619. STUDY DESIGN: The fetoplacental vascular beds of 20 perfused human placental cotyledons were vasoconstricted with a continuous infusion of U46619 (10(-8) mol/L). The vasculature was then sequentially injected with deionized water, 30 ng adrenomedullin, 300 ng adrenomedullin, and 3000 ng adrenomedullin. Any change in perfusion pressure was noted after each dose. In a separate experiment the fetoplacental vasculature in 2 perfused cotyledons from each of 10 placentas was vasoconstricted with U46619 (10(-8) mol/L). Adrenomedullin was infused continuously at either 200 ng/min (n = 5) or 2000 ng/min (n = 5) for 40 minutes. A corresponding control cotyledon from each placenta had isotonic sodium chloride solution added to its perfusion. Perfusion pressures were recorded every minute during the infusion and for 40 minutes afterward. Analysis of variance was used to compare pressure changes in the cotyledons that received bolus doses of adrenomedullin. Paired t tests of mean percentage pressure changes were used to compare the study and control groups that received the continuous infusions. RESULTS: In the cotyledons that received bolus doses of adrenomedullin, the mean (+/-SEM) percentage perfusion pressure changes from the baseline were -6.7 +/- 0.5 for 30 ng adrenomedullin (P =.0039), -8.5+/- 0.7 for 300 ng adrenomedullin (P <.0001), and -13.1 +/- 1.0 for 3000 ng adrenomedullin (P <.0001). With the continuous adrenomedullin infusion of 200 ng/min, there was no significant difference in the mean percentage pressure change from baseline between the study and control groups (-0.57%). At 2000 ng/min there was a significant difference ( 15.34%; P <.0001). CONCLUSION: Adrenomedullin caused vasodilatation of fetoplacental vasculature previously constricted with the thromboxane sympathomimetic U46619 in the isolated perfused placental cotyledon. This vasodilatation occurred in a dose-dependent manner. PMID- 11120531 TI - Methotrexate infusion in low-risk gestational trophoblastic disease. AB - OBJECTIVES: The current study attempts to evaluate the effectiveness of methotrexate infusion therapy in the management of low-risk gestational trophoblastic disease and to find out whether an increase in the dose intensity can effect a faster remission and a shorter treatment duration. STUDY DESIGN: This is a prospective study. Between June 1990 and August 1998, 59 patients with low-risk trophoblastic disease were treated with methotrexate at a dose of 100 mg/m(2) as an intravenous bolus over 30 minutes followed by a 12-hour infusion of methotrexate at a dose of 200 mg/m(2). Folinic acid was not given unless the serum methotrexate reached a toxic level 24 hours after infusion (toxic level, 10 micromol/L). Actinomycin D was added in patients with a partial response. The follow-up period of these patients ranged from 12 to 113 months, with a median of 58.5 months and a mean of 55.7 months. RESULTS: Fifty-four patients attained a complete biochemical remission. Twenty-eight patients went into biochemical remission after one methotrexate infusion. Five patients showed a partial biochemical response. A relapse developed in 2 of the 54 complete responders at 3 months and 18 months after the initial therapy. Both patients received combination therapy consisting of methotrexate, etoposide, and bleomycin. They went into biochemical remission and have remained disease-free at the time of analysis. All of the 59 patients were in biochemical remission at the time of analysis. No significant side effects were observed except that Stevens-Johnson syndrome developed in 1 patient. CONCLUSIONS: Methotrexate infusion therapy described in this study is effective in the treatment of low-risk gestational trophoblastic disease. The omission of consolidation therapy and folinic acid rescue decreases the cost and duration of treatment. PMID- 11120532 TI - Is human myometrial sampling at the time of cesarean delivery safe? AB - OBJECTIVE: The mechanism for the initiation of human labor remains unknown and is under extensive investigation. Myometrium from patients in labor and not in labor is the ideal tissue to study structural, cellular, and molecular changes that occur during parturition. This study was designed to determine whether myometrial sampling at the time of cesarean delivery increases maternal morbidity. STUDY DESIGN: This is a prospective cohort study including 118 study and 236 control patients. A full-thickness myometrial sample was obtained from the superior edge of a transverse uterine incision at the time of cesarean delivery. Demographics and standard surgical morbidity data were collected. Statistical methods used included univariate and multivariate analysis. RESULTS: The study and control groups did not differ significantly with respect to age, gravidity, parity, birth weight, and Apgar scores. The estimated intraoperative blood loss was greater in the control group (P <.02); however, the change in hematocrit level (preoperative vs postoperative values) was not different. There were no significant differences in the rates of endometritis, wound infection, and venous thrombosis up to 6 weeks post partum. When study and control patients were stratified into term in labor, term not in labor, preterm in labor, and preterm not in labor categories and compared for maternal morbidity, there were still no significant differences for any of the outcome measures evaluated. CONCLUSION: On the basis of our data, human myometrial sampling at cesarean delivery does not increase overall maternal morbidity, irrespective of gestational age and the presence or absence of labor. PMID- 11120533 TI - Umbilical cord blood collection for transplantation: which technique should be preferred? PMID- 11120535 TI - Waveform analysis of the fetal electrocardiogram: methodologic aspects. PMID- 11120537 TI - Measurements of the pelvic floor in women and their relationship to genital prolapse. PMID- 11120539 TI - First-trimester screening for aneuploidy by nuchal translucency. PMID- 11120540 TI - First-trimester Down syndrome screening. PMID- 11120542 TI - Questions about case report on vaginal dysplasia. PMID- 11120544 TI - Increasing obesity in Brazil: predicting a new peak of cardiovascular mortality. PMID- 11120545 TI - Darier's disease: a new paradigm for genetic studies in psychiatric disorders. PMID- 11120547 TI - Hypomagnesemia in short bowel syndrome patients. AB - CONTEXT: Magnesium support to small bowel resection patients. OBJECTIVE: Incidence and treatment of hypomagnesemia in patients with extensive small bowel resection. DESIGN: Retrospective study. SETTING: Metabolic Unit of the University Hospital Medical School of Ribeirao Preto, University of Sao Paulo, Brazil. PATIENTS: Fifteen patients with extensive small bowel resection who developed short bowel syndrome. MAIN MEASUREMENTS: Serum magnesium control of patients with bowel resection. Replacement of magnesium when low values were found. RESULTS: Initial serum magnesium values were obtained 21 to 180 days after surgery. Hypomagnesemia [serum magnesium below 1.5 mEq/l (SD 0.43)] was detected in 40% of the patients [1,19 mEq/l (SD 0.22)]. During the follow-up period, 66% of the patients presented at least two values below reference (1.50 mEq/l). 40% increased their serum values after magnesium therapy. CONCLUSION: Metabolic control of serum magnesium should be followed up after extensive small bowel resection. Hypomagnesemia may be found and should be controlled. PMID- 11120546 TI - Is intra-operative gamma probe detection really necessary for inguinal sentinel lymph node biopsy? AB - CONTEXT: Sentinel node (SN) biopsy has changed the surgical treatment of malignant melanoma. The literature has emphasized the importance of gamma probe detection (GPD) of the SN. OBJECTIVE: Our objective was to evaluate the efficacy of patent blue dye (PBD) and GPD for SN biopsy in different lymphatic basins. DESIGN: Patients with cutaneous malignant melanoma in stages I and II were submitted to biopsy of the SN, identified by PBD and GPD, as part of a research project. SETTING: Patients were seen at Hospital Sao Paulo by a multidisciplinary group (Plastic Surgery Tumor Branch, Nuclear Medicine and Pathology). PATIENTS: 64 patients with localized malignant melanoma were studied. The median age was 46.5 years. The primary tumor was located in the neck, trunk or extremities. INTERVENTIONS: Preoperative lymphoscintigraphy, lymphatic mapping with PBD and intraoperative GPD was performed on all patients. The SN was examined by conventional and immunohistochemical staining. If the SN was not found or contained micrometastases, only complete lymphadenectomy was performed. MAIN MEASUREMENTS: The SN was identified by PBD if it was blue-stained, and by GPD if demonstrated activity five times greater than the adipose tissue of the neighborhood. RESULTS: Seventy lymphatic basins were explored. Lymphoscintigraphy showed ambiguous drainage in 7 patients. GPD identified the SN in 68 basins (97%) and PBD in 53 (76%). PBD and GPD identified SN in 100% of the inguinal basins. For the remaining basins both techniques were complementary. A metastatic SN was found in 10 basins. Three patients with negative SN had recurrence (median follow up = 11 months). CONCLUSION: Although both GPD and PBD are useful and complementary, PBD alone identified the SN in 100% of the inguinal lymphatic basins. PMID- 11120548 TI - Conventional chemotherapy for acute myeloid leukemia: a Brazilian experience. AB - CONTEXT: Young patients affected by acute myeloid leukemia (AML) achieve complete remission (CR) using conventional chemotherapy in about 55-85%. However, 30% of patients fail to achieve CR and the remission duration is often only about 12 months. More intensive treatment after CR seems to be necessary in order to maintain CR and obtain a definitive cure. In Brazil, few reports have been published on this important subject. OBJECTIVE: The aim of this study was to describe a Brazilian experience in the treatment of "de novo" acute myeloid leukemia (AML) in younger adult patients (age < 60 years). DESIGN: Retrospective analysis. SETTING: University Hospital, Hematology and Hemotherapy Center, State University of Campinas, Brazil. PARTICIPANTS: Newly diagnosed cases of "de novo" AML in the period from January 1994 to December 1998 were evaluated retrospectively, in relation to response to treatment, overall survival (OS) and disease free survival (DFS). Cases with acute promyelocytic leukemia (APL) were also included in this analysis. RESULTS: On the basis of an intention to treat, 78 cases of AML, including 17 cases of APL, were evaluated. The overall median follow-up was 272 days. The complete remission (CR) rate was 63.6% in the AML group (excluding APL) and 78% in the APL group. The 5-year estimated disease-free survival (DFS) was 80% for the APL group and 34% for the AML group (P = 0.02). The 5-year estimated overall survival (OS) was 52% for the APL group and 20.5% for the AML group, respectively (P = NS). Relapse was observed in 12/39 (30.7%) patients with AML and 1/11 (9%) with APL. CONCLUSIONS: These results are similar to those reported in the literature. However, relapse and mortality rates remain high, and a search for more aggressive strategies in order to prevent relapse is recommended. PMID- 11120549 TI - Calcium acetate versus calcium carbonate in the control of hyperphosphatemia in hemodialysis patients. AB - CONTEXT: Hyperphosphatemia has an important role in the development of bone and mineral abnormalities in end-stage renal disease (ESRD). OBJECTIVE: To compare the phosphorus binding power and the hypercalcemic effect of calcium acetate and calcium carbonate in hemodialysis patients. TYPE OF STUDY: Crossover, randomized, double-blind study. PLACE: A private hospital dialysis center. PARTICIPANTS: Fifty-two patients who were undergoing regular hemodialysis three times a week ([Ca++] dialysate = 3.5 mEq/L). PROCEDURES: Half of the patients were started on 5.6 g/day of calcium acetate and, after a 2 week washout period, received 6.2 g/day of calcium carbonate. The other half followed an inverse protocol. MAIN MEASUREMENTS: Clinical interviews were conducted 3 times a week to monitor for side effects. Determinations of serum urea, calcium, phosphorus, hematocrit, Kt/V and blood gas analysis were obtained before and after each treatment. RESULTS: Twenty-three patients completed the study. A significant increase in calcium plasma levels was only observed after treatment with calcium carbonate [9.34 mg/dl (SD 0.91) vs. 9.91 mg/dl (SD 0.79), P < 0.01]. The drop in phosphorus levels was substantial and significant for both salts [5.64 mg/dl (SD 1.54) vs. 4.60 mg/dl (SD 1.32), P < 0.01 and 5.89 mg/dl (SD 1.71) vs. 4.56 mg/dl (SD 1.57), P < 0.01, for calcium acetate and calcium carbonate respectively]. The percentage reduction in serum phosphorus (at the end of the study) per milliequivalent of salt administered per day tended to be higher with calcium acetate but statistical significance was not found. CONCLUSION: Calcium acetate can be a good alternative to calcium carbonate in the handling of hyperphosphatemia in ESRD patients. When calcium acetate is used, control of hyperphosphatemia can be achieved with a lower administration of calcium, perhaps with a lower risk of hypercalcemia. PMID- 11120550 TI - Breastfeeding training for health professionals and resultant changes in breastfeeding duration. AB - CONTEXT: Promotion of breastfeeding in Brazilian maternity hospitals. OBJECTIVE: To quantify changes in the breastfeeding duration among mothers served by hospitals exposed to the Wellstart-SLC course, comparing them with changes among mothers attended by institutions not exposed to this course. DESIGN: Randomized Institutional Trial. SETTING: The effects of training on breastfeeding duration was assessed in eight Brazilian hospitals assigned at random to either an exposed group (staff attending the Wellstart-SLC course) or a control group. SAMPLE: For each of the eight study hospitals, two cohorts of about 50 children were visited at home at one and six months after birth. The first cohort (n = 494) was composed of babies born in the month prior to exposure to the Wellstart-SLC course, and the second cohort (n = 476) was composed of babies born six months subsequent to this exposure. MAIN MEASUREMENTS: Kaplan-Meier curves were plotted to describe the weaning process and log-rank tests were used to assess statistical differences among survival curves. Hazard ratio (HR) estimates were calculated by fitting Cox proportional hazard regression models to the data. RESULTS: The increases in estimated, adjusted rates for children born in hospitals with trained personnel were 29% (HR = 0.71) and 20% (HR = 0.80) for exclusive and full breastfeeding, respectively. No changes were identified for total breastfeeding. CONCLUSION: This randomized trial supports a growing body of evidence that training hospital health professionals in breastfeeding promotion and protection results in an increase in breastfeeding duration. PMID- 11120551 TI - Calcified abdominal pregnancy with eighteen years of evolution: case report. AB - CONTEXT: The lithopedion (calcified abdominal pregnancy) is a rare phenomenon and there are less than 300 cases reported in the medical literature. CASE REPORT: In this case, a 40 year-old patient had had her only pregnancy 18 years earlier, without medical assistance since then. She came to our hospital with pain and tumoral mass of approximately 20 centimeters in diameter. Complementary examinations (abdominal X-ray, ultrasonography and computerized tomography) demonstrated an extra-uterine abdominal 31-week pregnancy with calcification areas. Exploratory laparotomy was performed, with extirpation of a well-conserved fetus with partially calcified ovular membranes. PMID- 11120552 TI - Bronchial oncocytoma. AB - CONTEXT: Oncocytomas are generally small and present slow growth. Finding of the tumor usually occurs incidentally. Their incidence is higher among male patients. Oncocytomas in mucous bronchial glands are extremely rare. CASE REPORT: A 35-year old male who presented bronchial oncocytoma. The tumor was found after bronchoscopy that investigated an atelectasis of the upper left lobe. Histological examination with optical microscopy revealed a mature neoplasm formed by ovoid cells with thin, granular, eosinophilic cytoplasm and small nuclei similar to oncocytes. Electron microscopy showed mitochondrial hyperplasia. A three-year follow-up after thoracotomy followed by lobectomy and removal of the bronchial tumor was uneventful. PMID- 11120553 TI - Ewing's sarcoma of the head and neck. AB - CONTEXT: Ewing's sarcoma is a rare neoplasm, which usually arises in long bones of the limbs and in flat bones of the pelvis, with the involvement of head and neck bones being very unusual. CASE REPORT: a case of Ewing's sarcoma occurring in the mandible of a 35-year-old female. Pain and swelling of the tumor were the main complaints. The early hypothesis was an undifferentiated malignant neoplasm, possibly a sarcoma. The CT scan depicted an expansive lesion, encapsulated, with septa and characteristics of soft tissue, involving the left side of the mandible and extending to the surrounding tissues. The patient underwent surgical excision of the lesion, the definitive diagnosis of Ewing's sarcoma was established, and the patient commenced on radiotherapy. PMID- 11120554 TI - Darier's disease: a new paradigm for genetic studies in psychiatric disorders. AB - CONTEXT: One strategy for identifying susceptibility genes for common disorders is to investigate Mendelian diseases, cosegregating with these common disease phenotypes. CASE REPORT: A family with seven members is described, in which three members present Darier's disease and depression. This apparent cosegregation, if true, would support the hypothesis that in some pedigrees, a gene for mood disorder may be located on chromosome 12. PMID- 11120555 TI - Identification of visual arrestin (S-antigen) in retinal pigmented epithelial cells. AB - PURPOSE: Inactivation of photolyzed rhodopsin requires phosphorylation of this receptor and binding of the 48 kDa regulatory protein arrestin (S-antigen). Arrestin is also to cause an autoimmune disease, uveoretinits, that resembles uveitis in humans. In this study we demonstrate the presence of visual arrestin in retinal pigment epithelial cells (RPE) in culture. METHODS: Bovine RPE were isolated. Mouse and rat monoclonal and rabbit polyclonal antibodies against visual arrestin, and a synthetic peptide "GFLGELTSSEVATEVPFRLM" (a pathogenic sequence corresponding to residues 340 to 359 of human visual arrestin), and rabbit polyclonal antibody against the specific peptide "EDPDTAKESFQ" for bovine visual arrestin were used to detect arrestin in RPE cells. Using visual arrestin specific primer, RT-PCR of RNA from RPE was performed. RESULTS: By western blots analysis a 48 kDa protein, corresponding to visual arrestin was detected with both mAb and polyclonal antibodies in extracts of RPE cells. RT-PCR analysis of RNA from RPE cells confirmed the presence of arrestin mRNA of predicted 377 bp and exhibited 100% homology with visual arrestin 48 kDa. CONCLUSION: Visual arrestin proteins present in RPE may be involved in the desensitization of G protein-coupled receptors in RPE cells and in arrestin uveopathogenesis. PMID- 11120556 TI - Matrix metalloproteinase (MMP) expression in experimental choroidal neovascularization. AB - PURPOSE: Matrix metalloproteinases (MMP) are a family of proteolytic enzymes that degrade basement membrane and extracellular matrix proteins. To gain information on the possible role of MMPs in choroidal neovascularization (CNV), we have analyzed the mRNA expression of MMP-2 and MMP-9, two forms of MMPs implicated in ocular neovascularization, in a rat model. METHODS: Choroidal neovascularization was induced in pigmented rats by krypton laser photocoagulation of the fundus whereafter eyes were enucleated at 1, 3, 5, 7, 10 and 60 days. Antisense and sense riboprobes were generated using DNA complementary to MMP-2 and MMP-9, and mRNA expression was analyzed using in situ hybridization. RESULTS: In the untreated eyes MMP-2 mRNA expression was weakly detected in cells within the choroid. In laser-treated eyes MMP-2 mRNA expression was markedly increased and mainly localized to macrophage-like and retinal pigment epithelial (RPE)-like cells invading the choroid, subretinal space and inner retina. This increase in MMP-2 mRNA expression peaked at day 10 whereafter a decline was detected. MMP-9 mRNA expression was low in untreated eyes and did not increase following laser treatment. CONCLUSION: The results show that MMP-2 mRNA expression is increased in experimental CNV, and support of a role for MMP-2 in the development of CNV in age-related macular degeneration. PMID- 11120557 TI - Assessment of the effect of oral clarithromycin on visual outcome following presumed bacterial endophthalmitis. AB - PURPOSE: The use of clarithromycin was assessed as a biofilm reducing agent in the management of bacterial endophthalmitis. METHODS: 84 eyes of 83 patients presenting with clinical signs highly suggestive of bacterial endophthalmitis were treated using a standard regimen of intraocular vancomycin, amikacin and systemic steroids, which in addition included oral clarithromycin. Ocular penetration of oral clarithromycin in healthy and inflamed eyes was also assessed. RESULTS: Comparing visual acuities at presentation and 6 months, 66% of patients demonstrated an improvement. Intraocular samples were culture positive in 58% of eyes. As compared to culture positive cases, more culture negative cases achieved a visual acuity of 6/12 or better (p = 0.0047). As compared to patients receiving the standard protocol but without clarithromycin, a greater number of culture negative cases demonstrated an improvement in vision of > or = + 6 Snellen lines (p = 0.023). The ocular penetration of clarithromycin into the anterior chamber of inflamed eyes appears sufficient to allow anti-biofilm activity against bacteria at the basic pH encountered in eyes with endophthalmitis. CONCLUSIONS: The ocular penetration of clarithromycin appears adequate for anti-biofilm activity in inflamed eyes. The beneficial effects of oral clarithromycin on visual outcome has been demonstrated in culture negative eyes with clinical signs highly suggestive of bacterial endophthalmitis. The final visual outcome for culture positive cases remains poor. PMID- 11120558 TI - The epithelial sodium channel (ENaC) in rodent retina, ontogeny and molecular identity. AB - PURPOSE: To assess the appearance of the epithelial sodium channels (ENaC) during retinal development and establish its molecular identity. METHODS: Photoreceptors were isolated by horizontal sectioning of rat retina with a vibratome series 1000. Intact retina and isolated photoreceptors were analyzed for the developmental appearance of ENaC using the Polymerase Chain Reaction (PCR) technique. Immunofluorescence was conducted with the aid of an antibody raised against the 14 amino acids peptide QGLGKGDKREEQGL, corresponding to the N terminal region (44-58 aa) of alpha ENaC. RESULTS: ENaC message was present in rat retina photoreceptors, isolated just one day after birth. The bipolar and ganglion cell layers, separated from whole retina by vibratome, also contained the ENaC message. The partial sequence of the photoreceptor ENaC (496 base) exhibited one hundred percent homology with the channel from rat known sources. Immunochemistry revealed that only the outer nuclear layer was positive for the ENaC in one-day-old rat; the inner segment became immunopositive at the age of 9 days. CONCLUSION: The ENaC is present in the retina and visible soon after birth. These observations suggest that ENaC may prove to have physiological importance in the retina but until now its function is unknown. PMID- 11120559 TI - Identification and characterization of a retina-specific calpain (Rt88) from rat. AB - PURPOSE: To identify and characterize a newly discovered calpain termed Rt88 from rat retina. METHODS: Rt88 in retina under normal physiological conditions was characterized in Sprague-Dawley rats of various ages by competitive RT-PCR, Northern blot analysis, cDNA cloning and sequencing. Recombinant Rt88 was expressed in the baculovirus system and characterized by casein zymography and immunoblotting. RESULTS: Rt88 was sequenced and found to be similar to muscle calpain p94 except for three differences. A different exon 1 (as in lens Lp82 calpain) was present, and exons 15 and 16 in the unique IS2 region of muscle p94 were deleted. Of eleven tissues studied, mRNA for Rt88 was found only in retina where Rt88 increased with maturation and then remained constant. Casein zymography showed that rRt88 was proteolytically active after activation by calcium, but intact rRt88 was rapidly broken due to the presence of the IS1 region in domain II. CONCLUSIONS: Rt88 is a retina-specific, calcium activated protease from the calpain superfamily (EC 3.4.22.17) of cysteine proteases. Rt88 is a recently identified member of the AX1 subfamily of calpains showing alternative exon 1 usage. So far, all AX1 subfamily members are from eye. Rt88 may perform specific proteolytic functions during development, normal turnover, or pathological degeneration of retinal proteins. PMID- 11120560 TI - Apoptosis in cytomegalovirus retinitis associated with AIDS. AB - PURPOSE: To determine the role of apoptosis in the pathogenesis of cytomegalovirus retinitis in patients with the acquired immunodeficiency syndrome. METHODS: Forty-three eyes from patients with cytomegalovirus retinitis treated before the introduction of highly active anti-retroviral therapy were examined by routine histopathology and in situ techniques to detect apoptosis (TUNEL assay). Apoptosis was graded on a scale from 0 to 3+ by quantitating the number of TUNEL positive cells per case using a standard grading procedure. Statistical analysis describing the association between apoptosis grade and the proportions of eyes with active CMV infection, with choroidal inflammation and treated with the sustained-release intravitreal ganciclovir implant was performed using the Armitage procedure. RESULTS: Apoptosis in both CMV infected and uninfected retinal cells was detected in 28 of 41 eyes (68%); 13 (45%) with active and 15 (55%) with inactive cytomegalovirus retinitis. The degree of apoptosis was mild (1+) in 14 eyes, moderate (2+) in 6 eyes and severe (3+) in 8 eyes. Apoptosis was not identified in two eyes without CMVR. An increase in apoptosis grade was positively associated with active CMVR (p = 0.014). There was no significant association between apoptosis and choroidal inflammation. The presence and the severity of apoptosis was less in eyes treated with the sustained-release intravitreal ganciclovir implant compared to those treated with systemic anti-viral therapy, however, the difference was not statistically significant. CONCLUSIONS: Apoptosis contributes to retinal cell loss in eyes with cytomegalovirus retinitis associated with AIDS but did not correlate with the progressive loss of retinal cell function in patients with treated, inactive CMVR. PMID- 11120561 TI - Alkaline protease-deficient mutants of Pseudomonas aeruginosa are virulent in the eye. AB - PURPOSE: Alkaline protease has been associated with virulence in Pseudomonas aeruginosa corneal infections. To define the role of this enzyme in such infections, isogenic mutants of P. aeruginosa deficient in alkaline protease production were constructed. This study examines the ability of these mutants to adhere to scarified corneal tissue in vitro and to establish corneal infections in vivo. METHODS: Mutants were constructed by allelic exchange in two phenotypically different wild type strains, PAO1 (invasive) and ATCC 19660 (cytotoxic). Alkaline protease-deficient mutants were characterized by zymography and western blot analysis of bacterial culture supernatants. Allelic exchange was confirmed by PCR analysis of the disrupted aprA gene of the mutants. Adherence of wild type and mutant strains to scarified corneal epithelium was assessed by an in vitro organ culture assay, while ocular virulence of the strains was determined in vivo using a mouse scarification model of bacterial keratitis. RESULTS: Being isogenic, phenotypes of mutants were identical to their respective parents with the exception of the loss of alkaline protease production. The absence of alkaline protease did not alter corneal adherence or ocular virulence of the organisms when compared to similar wild type strains. CONCLUSIONS: These data provide evidence that alkaline protease produced by P. aeruginosa is not essential in the pathogenesis of P. aeruginosa keratitis. PMID- 11120562 TI - Uveal melanoma model with metastasis in rabbits: effects of different doses of cyclosporine A. AB - OBJECTIVE: To compare the toxicity and efficacy of different doses of cyclosporine A (CsA) in a rabbit model of uveal melanoma. METHODS: We used four experimental groups: control, no CsA; group 1, 15 to 10 mg/kg/day; group 2, 15 mg/kg/day; and group 3, 20 mg/kg/day. The MKT-BR cell line was implanted in the choroid. All animals underwent ophthalmoscopic evaluation; the animals were weighed and blood levels of CsA, blood urea nitrogen (BUN), creatinine and alanine aminotransferase (ALT) were measured weekly. Necropsies and histologic study were performed to detect intraocular tumors and metastasis. RESULTS: A difference in survival rates was found between groups 2 and 3 (p = 0.0042). Differences were observed in the mean BUN and creatinine levels (p < 0.001 and p < 0.003, respectively) between groups but not in the ALT. Intraocular tumors were detected ophthalmoscopically in 50%, 65%, and 70% of the animals in groups 1, 2, and 3 respectively, and histologically in 70%, 90%, and 100% of the same groups. Lung metastases were found in 26.8% of animals with intraocular tumors. Differences were observed in mean CsA blood levels between animals with and without histologically demonstrated uveal tumors (p = 0.001) but not in animals with or without metastasis. CONCLUSIONS: Different doses of CsA affect survival, tumor development and renal toxicity. Metastatic disease is independent of CsA dose and the subsequent CsA blood levels. A blood level of CsA ranging from 500 to 1000 ng/ml and doses of 10 to 15 mg/kg/day may effectively develop this model of uveal melanoma. PMID- 11120563 TI - Effects of atrial natriuretic peptide and sodium nitroprusside on epidermal growth factor-stimulated wound repair in rabbit corneal epithelial cells. AB - PURPOSE: Treatment of rabbit corneal epithelial cells (RCEC) with epidermal growth factor (EGF) stimulates cell proliferation and wound repair in a cell culture model system. Studies have also shown that atrial natriuretic peptide (ANP) and sodium nitroprusside (SNP), a nitric oxide-generating agent, inhibit proliferation of a variety of cell types. The aim of the present work was to examine whether ANP or SNP has any effect on EGF-stimulated proliferation of RCEC involved in wound repair. METHODS: The SV-40 immortalized RCEC were cultured in 24-well plates until they became confluent. Wounds of uniform size (8 mm diameter) were created and the cells allowed to grow in the presence and absence of EGF and/or other agents. At prescribed time intervals, the cells were stained by Giemsa and the wound areas digitized and quantified by Sigma Image Scan System. The cGMP contents in RCEC, treated with or without ANP or SNP, were measured by radioimmunoassay. RESULTS: Addition of EGF (1-100 ng/ml) to RCEC stimulated cell proliferation which significantly reduced the time required for wound closure. Addition of ANP (1 nM to 10 microM) or SNP (10 microM to 1 mM), in the presence of EGF, dose-dependently inhibited the growth factor-stimulated wound closure in RCEC. When added alone to the cells, ANP or SNP increased cGMP accumulation in a dose-dependent manner. Addition of ANP (1 microM) or SNP (1 mM) to primary corneal epithelial cells, in the presence and absence of EGF, also inhibited the wound closure with a corresponding increase in cGMP contents. Treatment of the cells with ODQ (10 nM to 10 microM), a soluble guanylate cyclase inhibitor, dose-dependently decreased the SNP-induced accumulation of cGMP, and reversed the inhibitory effect of SNP on EGF-stimulated wound closure. Addition of membrane-permeable cGMP analog, 8-bromo-cGMP, to RCEC inhibited the EGF stimulated wound closure in a dose-dependent manner. Treatment of RCEC with mitomycin C (5 microM) exerted a marked inhibitory effect on wound closure in the presence and absence of EGF, and also abrogated the inhibitory effect of 8-bromo cGMP on wound closure in the EGF-treated and untreated cells. CONCLUSIONS: The results demonstrate that ANP and SNP inhibit the EGF-stimulated wound repair in RCEC. The effect of these agents is mediated via activation of guanylate cyclases that generate cGMP. Cyclic GMP appears to exert its inhibitory effect at the level of cell proliferation and not cell migration. The data suggest an important role for cGMP-dependent protein kinase in proliferation of RCEC stimulated by EGF. PMID- 11120564 TI - Age dependence of perimacular white blood cell flux during isometric exercise. AB - PURPOSE: To investigate the age dependence of perimacular white blood cell flux (WBCF) during isometric exercise. METHODS: Fourteen healthy young (age range: 21 29 years; 24 +/- 3 years, mean +/- SD, 12 male and 2 female) and 15 healthy middle-aged (age range: 45-57 years; 53 +/- 4 years, mean +/- SD; 5 male and 10 female) volunteers were studied. Subjects performed isometric handgrip for 10 minutes and squatting for 6 minutes. WBCF was assessed with the blue field entoptic technique, mean arterial pressure (MAP) was measured with an automated oscillometric device, intraocular pressure (IOP) was measured by Goldmann applanation tonometry and ocular perfusion pressure (OPP) was calculated as 2/3 MAP - IOP. RESULTS: Baseline WBCF was significantly higher in young subjects than in middle-aged subjects (191 +/- 28 vs 142 +/- 23; p = 0.001). Isometric handgrip induced a significant increase in WBCF in the middle-aged subjects (23 +/- 24%; p = 0.005), but not in the young subjects. Squatting significantly increased WBCF in both groups (young: 42 +/- 23%; p = 0.004 and middle aged: 51 +/- 27%; p < 0.001). A significant deviation from baseline WBCF was observed when OPP increased by 42 +/- 4% (p = 0.003) and 35 +/- 4% (p < 0.001) for the young and middle-aged subjects, respectively. The OPP-WBCF relationship was not different between the two study groups. CONCLUSION: Altered retinal autoregulation as observed in vascular ocular disease appears to be unrelated to the normal physiological aging process. PMID- 11120565 TI - Digital measurement of torsional eye movement due to postural change and its effect on reading performance. AB - PURPOSE: Posture induced torsional eye movements have rarely been investigated. The current study made use of digital imaging technology to measure the cycloduction resulting from postural change. The effect of cycloduction on reading performance was also investigated. METHODS: Thirty subjects were recruited and pictures of the right eye were captured using a digital camera at three postures, i.e. sitting, 90 degrees tilted to the right and 90 degrees tilted to the left. With the identification of a conjunctival landmark, torsional eye movement was measured. The subjects were then required to read a custom designed near chart while in those three postures, the reading card being rotated 90 degrees clockwise or 90 degrees anti-clockwise, to match with the tilting of the head. The reproducibility of the torsional eye movement and reading performance measure was determined in 12 of those 30 subjects. RESULTS: Incycloduction was induced when tilting to the right and excycloduction when tilting to the left. This method was found to be reproducible with the 95% confidence limits of 0.80 degrees between visits. The mean incycloduction induced was 6.50 degrees (SD 1.51 degrees ) and 6.41 degrees (SD 1.46 degrees ) for excycloduction. No significant difference in amount was demonstrated (paired t test: t = 0.624, P = 0.538). No significant difference was found in the reading scores at various postures (Repeated measures ANOVA: df = 2, F = 1.881, P = 0.162). CONCLUSIONS: The results presented here demonstrate that we have developed an objective and instantaneous method with good precision, which could be applied in other studies that require the measurement of torsional eye movements. The induced cycloduction did not affect the reading performance. PMID- 11120566 TI - Detection of female carriers of congenital color-vision deficiencies by visual pigment gene analysis. AB - PURPOSE: Congenital color-vision deficiencies are frequent among males, 4.7-8.0%, suggesting that female carriers are present at a frequency of 9-15%. The purpose of this study was to determine whether carriers could be detected by analysis of the visual pigment genes. METHODS: DNA from 29 males with congenital color-vision deficiencies, from their mothers, and from 117 randomly-selected females was analyzed. The most upstream genes, the downstream genes, and the most downstream genes in the red/green pigment gene arrays were amplified separately by PCR. Exon 5 of each gene was analyzed by single-strand conformation polymorphisms (SSCP). RESULTS: Analysis of the visual pigment genes suggests that one of the 29 mothers examined is a female protan and two others are carriers of both protan and deutan defects. The remaining 26 mothers were confirmed to be carriers of congenital color-vision deficiencies. Unusual patterns were observed in 15 (13%) of the randomly-selected females; among them, 5 appeared to be protan carriers and at least 4 to be deutan carriers. CONCLUSIONS: Female carriers of congenital color vision deficiencies can be detected by analysis of the visual pigment genes. Since the proportion of females showing unusual patterns was slightly higher than expected, some must be false-positives and require more detailed examination. PMID- 11120567 TI - Response of rabbit ciliary process cyclic nucleotides to specific phosphodiesterase inhibitors. AB - PURPOSE: To determine which cyclic nucleotide phosphodiesterase family activities can be identified in rabbit ciliary processes. METHODS: Freshly excised rabbit ciliary processes were incubated in vitro with family selective phosphodiesterase inhibitors in the absence or presence of activators of soluble or membrane bound guanylate cyclase. The resulting increases of cyclic AMP and cyclic GMP were measured by RIA. RESULTS: Rabbit ciliary process cyclic AMP levels were increased by the phosphodiesterase 1 and 4 selective inhibitors, 8-methoxymethyl-IBMX and rolipram, respectively. Cyclic GMP levels were increased by 8-methoxymethyl-IBMX; whereas the phosphodiesterase 5 and 6 selective inhibitor, zaprinast, increased cyclic GMP little. Nitric oxide donors increased cyclic AMP in addition to cyclic GMP, and inhibition of soluble guanylate cyclase eliminated the increase of cyclic AMP. The effects of sodium nitroprusside and the phosphodiesterase 3 selective inhibitor, cilostamide, on cyclic AMP were not additive suggesting that the sodium nitroprusside mediated increase of cyclic GMP, like cilostamide, inhibited phosphodiesterase 3. CONCLUSIONS: Cyclic AMP in rabbit ciliary processes is hydrolyzed primarily by phosphodiesterases 1 and 4, and, when cyclic GMP levels are low, by phosphodiesterase 3; cyclic GMP is hydrolyzed primarily by phosphodiesterase 1. PMID- 11120568 TI - P2Y(2) receptor stimulation increases tear fluid secretion in rabbits. AB - PURPOSE: The purinergic P2Y(2) receptor agonists stimulate active Cl-transport across the excised rabbit conjunctival tissue in vitro. We determined whether UTP or ATP could increase the tear volume and change tear fluid composition in normal rabbits in vivo. METHODS: Fifty mL was applied to rabbit eyes of UTP, ATP at concentrations of 0.1, 0.3, 1, 3, 8.5% (1.8-154 mM) or saline. A modified Schirmer test with topical anesthesia was performed 5, 15, 30 and 60 min after the instillation. In studies lasting 30 days, 50 microL of 0.5% UTP was applied 6 times a day for 4 weeks. Tear samples were collected from the conjunctival sac with a glass microcapillary. The protein profile of the tear fluid was analyzed by SDS-polyacrylamide gel electrophoresis and total protein was measured with the Bradford assay. The Easy-Titer rabbit IgG assay kit was used for the determination of immunoglobulin G (IgG). RESULTS: UTP had dose-dependent stimulatory effects on tear secretion. It maximally increased tear secretion about 4-fold 15 min after its application. Similar effects were obtained with ATP. Repeated treatment with UTP reproducibly increased tear volume. Furthermore, UTP did not decrease total protein and IgG concentration in tear fluid and it had no effect on the protein profile. CONCLUSION: These data indicate that activation of P2Y(2) receptor increases tear fluid secretion accompanied with some proteins in normal rabbits. The purinergic agonists, UTP and ATP, have potential therapeutic value in the treatment of dry eye. PMID- 11120569 TI - Endothelial cells analysis with the TOPCON specular microscope SP-2000P and IMAGEnet system. AB - PURPOSE: Corneal endothelium plays an important role in the maintenance of corneal transparency and stability. The purpose of this study was to evaluate the performance of the TOPCON SP-2000P and IMAGEnet system in terms of (a) the difference in results between automated endothelial cell analysis and retraced cell analysis, (b) the differences in the endothelial cell analysis when using the first image and the clearest of three images taken with the SP-2000P, (c) the repeatability and reproducibility of the IMAGEnet system in the determination of corneal endothelial cell variables, and (d) the repeatability and reproducibility of the SP-2000P in capturing endothelial cell images for endothelial cell analysis. METHODS: Two experiments were performed. Twenty male subjects participated in the first experiment in which endothelial images were captured by examiner SWC and endothelial cell analysis was performed by two examiners, SWC (twice on two different days) and PC (once only). Nineteen male subjects participated in the second experiment in which endothelial cell images were captured by examiners SWC and PC on Visit 1 and by examiner SWC on Visit 2. Endothelial cell analysis was performed by examiner SWC only. The 95% limits of agreements (95%LA) and the intraclass correlation coefficient (ICC) between tested parameters were determined. RESULTS: Automated cell analysis significantly underestimated average cell size (ACS) and hexagonality, and overestimated endothelial cell density (ECD) and coefficient of variance of cell size (CV) (p < 0.05). No statistically significant differences were found in the endothelial cell variables between the first and the clearest images (p > 0.05). There were no statistically significant intra- or inter-examiner differences in any of the endothelial variables determined (p > 0.05). For a set of endothelial cell images, the reproducibility and repeatability of the endothelial cell variables determined with the IMAGEnet system were good, with ICC > 0.9. No significant inter-visit or inter-examiner differences were found (p > 0.05). Reliability was good for the determination of ECD and ACS (ICC > 0.9) but poor or average for the determination of CV and hexagonality (ICC: 0.4-0.8). CONCLUSIONS: We suggest the use of the clearest image with the retraced method for endothelial cell analysis with the IMAGEnet system, as the variability in hexagonality results between the first and the clearest images can be clinically significant. For the same set of images captured by one examiner, the TOPCON IMAGEnet system was reliable in determining ECD and ACS, and was fairly reliable in determining hexagonality and CV. When images were captured on different days or by different examiners, the TOPCON SP-2000P with the IMAGEnet system gives repeatable and reproducible values for ECD and ACS only. Caution should therefore be exercised when using CV and hexagonality to monitor endothelial changes in comparative studies. PMID- 11120570 TI - Central and peripheral corneal thickness measured with the TOPCON specular microscope SP-2000P. AB - PURPOSE: Modern refractive surgery and follow up relies on a knowledge of corneal thickness (CT) and shape, and the reliability of modern instrumentation providing such data is important. This study sought to determine the performance of the TOPCON SP-2000P specular microscope in measuring CT. The aims of this study were: (a) to determine if there is any difference between the CT results obtained from the first image, the clearest of three images and the mean of measurements from three images; (b) to determine the correlation between central and peripheral CT and (c) to investigate the reliability (repeatability and reproducibility) of the SP-2000P in the determination of central and peripheral CT. METHODS: The central and peripheral CT measurements shown on the first (f), the clearest (c) of three images and the mean (m) CT shown in three images captured with the SP-2000P of 43 subjects were compared. All images were captured by the same examiner. Nineteen of the subjects (male) returned on another day and measurements were taken by two examiners. RESULTS: fCT, cCT and mCT were not significantly different from each other. The intraclass correlation coefficients (ICC) between the three values were> 0.9 for all corneal locations measured. However, the variability in the differences between fCT and cCT was relatively greater for peripheral CT (except temporal). The peripheral CT values obtained were all significantly greater than the central CT, and statistically significant correlations were found between the central and each of the peripheral thickness. There was no statistically significant between-visit or between-examiner differences in the central or peripheral CT. The ICC values for between-visit differences for central and temporal measurements were > 0.9 but for the other corneal locations, the ICC values were 0.81 to 0.88. For between-examiner differences, the ICC value was 0.82 for inferior CT and > 0.9 for the other four CT. CONCLUSIONS: Differences in CT measurements obtained from the first and clearest images captured by the SP 2000P were not statistically significant but can be clinically significant for peripheral CT. We therefore suggest the use of the CT value obtained from the clearest of three images. For every unit change in central CT, there is an approximately equivalent change in peripheral CT. Based on the ICC values, the SP 2000P showed very good repeatability and reproducibility in the determination of central CT. For the determination of peripheral CT, the repeatability and reproducibility of the SP-2000P were good and reasonably good respectively. These findings have relevance to the measurement of the cornea before and after refractive surgery. PMID- 11120571 TI - Ocular effects associated with the chronic administration of the adenosine A(1) agonist cyclohexyladenosine. AB - PURPOSE: To investigate changes in ocular responses associated with the chronic administration of the adenosine A(1) agonist cyclohexyladenosine (CHA). METHODS: New Zealand White rabbits were treated unilaterally twice-a-day for 30 days with CHA (165 or 500 microg) or vehicle. Intraocular pressures (IOPs) and pupil diameters (PDs) were evaluated over the course of the study. At the end of the study period, outflow facility was determined in selected animals and compared to naive vehicle- and CHA-treated animals. RESULTS: In rabbits receiving 165 microg of CHA, ipsilateral IOPs at 2 and 6 hours post-drug exhibited progressively greater reduction over the course of the study. Regression analysis demonstrated a significant correlation between study duration and lower IOP at 2 and 6 hours post-drug. In rabbits receiving 500 microg of CHA, ipsilateral IOP reductions at 2 hours post-drug were similar throughout the 30-day study. However, analysis of ipsilateral IOPs 6 hours following CHA administration, demonstrated a significant correlation between study duration and lower IOPs. Enhanced contralateral responses at 2 hours post- drug, were also measured in rabbits receiving 165 or 500 microg of CHA. In animals receiving chronic CHA treatment for 30 days, outflow facility 3 hours post-CHA was significantly elevated over that measured in naive vehicle-treated rabbits. Although mean outflow facility in chronic treatment animals was slightly elevated over CHA-induced increases in naive rabbits, this difference was not significant. No evidence of tolerance was observed for either dose during the course of these studies. No change in PD during the course of these studies was measured. CONCLUSIONS: The chronic administration of the adenosine A(1) agonist CHA twice daily produced no evidence of tolerance. Unexpectedly, the IOP response to CHA was enhanced with chronic administration. These data provide evidence that the use of adenosine A(1) agonists may be useful in the chronic treatment of ocular hypertension at doses lower than those identified in acute IOP studies. PMID- 11120572 TI - Abnormalities of retinal metabolism in diabetes or experimental galactosemia VIII. Prevention by aminoguanidine. AB - PURPOSE: Aminoguanidine has been found to inhibit the development of some retinal lesions in diabetic rats and diabetic dogs, thereby raising a possibility that the formation of glycation end products (AGEs) may be an essential step in the pathogenesis of the retinopathy. The purpose of this study is to investigate the effect of aminoguanidine administration on other metabolic abnormalities which might be involved in the development of retinopathy in two models of the retinopathy, alloxan diabetes and experimental galactosemia. METHODS: Oxidative stress, nitric oxide (NO) and the activity of protein kinase C (PKC, total activity) were measured in the retina of the rats experimentally diabetic or galactosemic for 2 months. Effect of aminoguanidine administration on the inhibition of hyperglycemia-induced retinal dysmetabolism was investigated. RESULTS: Two months of diabetes or experimental galactosemia in rats resulted in elevation of retinal oxidative stress (increase in thiobarbituric acid reactive substances, TBARS, and decrease in glutathione, GSH), NO, and PKC activity. Aminoguanidine supplementation (2.5 g aminoguanidine/kg rat diet) significantly inhibited each of these abnormalities in retinas of diabetic rats and galactosemic rats, and did so without lowering the blood hexose levels of these animals. CONCLUSIONS: The ability of aminoguanidine to normalize the hyperglycemia-induced increases in retinal oxidative stress, NO and PKC in diabetic rats and galactose-fed rats suggests that these abnormalities may be inter-related in the retina, and that the biochemical mechanism by which aminoguanidine inhibits retinal microvascular disease in diabetes may be complex. PMID- 11120573 TI - Sialomucin complex (rMuc4) expression during development of the rat cornea. AB - PURPOSE: To determine changes in sialomucin complex (SMC, rat Muc4) expression in rat ocular surface tissue during development from birth to adult. METHODS: Immunoblot and immunocytochemical analyses were performed on isolated corneal tissue and rat eyes, respectively, from animals at days 1, 9 and 15 after birth. Adult animals were used for comparison. RESULTS: SMC was detected by immunoblotting at all ages (day 1-adult) and increased into adulthood. Immunocytochemical localization also showed steadily increasing amounts of SMC to adulthood. The SMC was located at the apical surface of the corneal epithelium and in the superficial layers of the stratified corneal epithelium. CONCLUSIONS: SMC at the corneal surfaces steadily increases during development to adulthood, but does not exhibit a marked transition in expression at the time of eye opening. PMID- 11120574 TI - Functional cavity dimensions of tear lipocalin. AB - PURPOSE: We calibrated the cavity of tear lipocalin with a series of fluorescent labeled lipids of increasing chain length and varying diameter. METHODS: Cavity length was assessed with competitive fluorescent assays in which DAUDA was displaced from apo-tear lipocalin with ligands of increasing carbon chain lengths from C12-C24. The concentrations of competitors that inhibit 50% of the binding of DAUDA (IC(50)) were compared. Functional diameters of tear lipocalin and beta lactoglobulin were estimated with fatty acids bearing fluorescent labels of various diameters. The cavity dimensions of other lipocalins were derived from their published crystal structure coordinates. RESULTS: In tear lipocalin, the binding affinities of fatty acids increased up to a carbon chain length of 18 (22.5 A) but remained constant from C18-C24. The cavity length of other lipocalins in crystal form were similar to tear lipocalin in solution. Tear lipocalin showed decreased binding affinities with progressively increasing ring dimensions of the ligand. In contrast to beta-lactoglobulin and retinol binding protein, tear lipocalin bound DAUDA and cholesterol in the calyx. Neither tear lipocalin nor beta-lactoglobulin bound P646 in their respective cavities. The calculated inter-sheet distances at the mouth of the crystallized lipocalins ranged from 16-22A. CONCLUSIONS: Tear lipocalin is more promiscuous than beta lactoglobulin or retinol binding protein because of a greater functional diameter. Differences in ligand specificity of the various lipocalins can not be explained simply by variation in cavity length or the intersheet distances at the calyx mouths as determined by crystal structure. Other factors may influence ligand specificity such as size and/or dynamic motion of loops between the beta strands. PMID- 11120575 TI - Behcet's disease in Japan and in Great Britain: a comparative study. AB - Behcet's disease (BD) is an important cause of visual morbidity throughout the world, but shows striking differences in racial predilection. Despite important advances in the therapeutic management of acute intraocular inflammation, the long-term impact of these new strategies on visual outcome of BD and their efficacy in different ethnic groups is unknown. A comparative study of patient characteristics, clinical ocular features and inflammatory score, and current therapy was undertaken on all patients fulfilling the International Study Group criteria for Behcet's disease and the Behcet's Disease Research Committee of Japan, who attended the Uveitis Clinics of Moorfields Eye Hospital (n=19) and Kurume University School of Medicine (KUS) (n=35) during a continuous consecutive four-week period. Japanese patients were significantly older (43.2+/-11.8 years) than the patients seen in London (35.4+/-8. 9 years). There was a predominance of male patients in both groups. All patients seen in KUS were Japanese, while the patients in London included 12 Caucasians, five Middle Eastern, one African, and one Asian. No significant differences were seen between the two populations in the duration of systemic disease or systems affected by the disease, such as mouth ulcers, genital ulcers, skin lesions including erythema nodosum, or arthritis. The duration of ocular disease was similar in both centres: around seven years. There was, however, a significant difference in the number of eyes with active anterior uveitis (59.7% KUS vs 18.4% London (chi-square: 5.4; p=0.006)) and/or posterior uveitis (31.3% KUS vs 18.4% London (chi-square: 5.42; p<0.02)). No significant differences were found in the number of eyes with optic disc swelling or optic atrophy and in each centre the number of eyes with vision greater than 6/9 or worse than 6/60 were the same. The treatment schedules were very different between the two centres. More patients were treated with topical steroids in Japan (68.7% KUS vs 10.5% London (chi-square: 30.5; p=0.001), but a similar number used concomitant intraocular pressure-lowering agents. More patients received systemic steroids in London (84.2% London vs 17% KUS (chi square: 20.25; p<0.001)). Three patients received systemic steroids alone, five had prednisolone and cyclosporin, four had prednisolone and azathioprine, and four had triple therapy with prednisolone, cyclosporin, and azathioprine. Only one patient used colchicine. Cyclosporin use was similar in London and KUS (47.4% and 42.8%, respectively). In Japan, three patients used prednisolone alone and three tacrolimus (FK506). In addition, two patients, who were on steroids alone, took colchicine as well. More patients in Japan had undergone surgery for cataract and glaucoma (chi-square: 4.0; p=0.045). In KUS, seven of 67 eyes had cataract surgery. A further three eyes had visually significant cataract and two eyes had undergone glaucoma surgery. In contrast, no patients in London had undergone any surgery up to and including this period. PMID- 11120576 TI - Localization and characterization of immunocompetent cells in the human retina. AB - PURPOSE: Recent studies have shown that experimental uveitis can be induced by the appropriate administration of various retinal antigens. Little is known about the in-situ interactions between immune cells in the retina as a prerequisite for understanding the mechanisms involving the presentation of antigens by local antigen-presenting cells (APC) to invading T cells. The study described here was therefore designed to investigate the presence of immunocompetent cells with a focus on the characterization of retinal APCs. MATERIALS AND METHODS: Retinal wholemounts, cytospins, and ocular sections were prepared from eyebank eyes obtained within 24 hours postmortem. Immunohistochemistry (single staining and double staining) was performed on the retinal wholemounts, cytospins, and the ocular sections using monoclonal antibodies specific for HLA-DR (MHC class II), CD45 (leukocytes), CD68 (macrophages), CD22 (B cells), CD3 (T cells), and CD1a (dendritic cells). RESULTS: CD68-positive macrophages were observed in one layer of the retina, whereas HLA-DR(+) and CD45(+) cells were seen in two distinct planes: one mainly at the level of the inner nuclear layer to outer plexiform layer (deep layer) and the other mainly at the nerve fiber and ganglion cell layer (shallow layer). There was a significant difference between the different parts of the retina with regard to the density of these cells. Cell density decreased when going from the peripheral to the posterior areas of the retina. The positive cells in the deep layer were frequently associated with blood vessels, whereas the cells in the shallow layer were distributed evenly throughout the retina. Most positive cells displayed a dendritiform appearance, whereas few cells showed a pleiomorphic morphology. Very few CD1a-positive cells were noted in the retina. Neither T cells (CD3) nor B cells (CD22) could be detected in the normal human retina. Double staining showed that the majority (83. 7%) of the CD45(+) cell population was HLA-DR-positive, whereas approximately half (56.8%) the CD68(+) cell population was HLA-DR-positive. CONCLUSIONS: This study shows that the human retina contains a number of different microglia populations, some of which express HLA-DR and could thus be involved in antigen presentation. Marked differences in cell density can be observed within the retina, with the most abundant presence seen in the peripheral retina. The normal retina contains few professional antigen-presenting cells (CD1(+)). PMID- 11120577 TI - Nedocromil sodium 2% ophthalmic solution for the treatment of ragweed pollen seasonal allergic conjunctivitis. AB - PURPOSE: To determine the efficacy and safety of nedocromil sodium 2% ophthalmic solution in the treatment of seasonal allergic conjunctivitis. METHODS: A combined analysis of two multicenter, randomized, comparative, double-masked, placebo-controlled clinical trials involving 261 patients diagnosed with seasonal allergic conjunctivitis was used. Patients were randomly assigned to receive either topical 2% nedocromil sodium or placebo twice daily for eight weeks. Diary card scores and clinician assessments of allergic symptoms were recorded throughout the study; efficacy was determined by comparing symptom severity at the peak pollen period with symptom severity at baseline. Clinician and patient evaluations of treatment effectiveness were used as secondary measurements of efficacy. RESULTS: Patients treated with nedocromil sodium experienced improvement in allergy symptoms, with reductions in the summary symptom score, itch, redness, conjunctival injection, and conjunctival edema significantly (p<0.05) greater than those observed in the patients treated with placebo. Clinicians' and patients' opinions of nedocromil sodium treatment effectiveness were significantly (p<0.02) superior to those of placebo treatment effectiveness. CONCLUSION: Nedocromil sodium is effective in the management of seasonal allergic conjunctivitis. PMID- 11120578 TI - Peroxynitrite formation in the orbit of diabetics with rhinocerebral mucormycosis. AB - OBJECTIVE: To evaluate whether an intact respiratory burst exists within the orbit of diabetics with rhinocerebral mucormycosis. METHODS: Immunohistochemical detection of nitrotyrosine in the orbital tissue of diabetics requiring exenteration due to rhinocerebral mucormycosis. Nitrotyrosine is the stable product of the nitration of tyrosine residues by peroxynitrite. Peroxynitrite is a potent oxidant produced by the combination of superoxide and nitric oxide during the respiratory burst. RESULTS: Four specimens were analyzed. All showed focal areas of specific staining against nitrotyrosine of the walls and internal structures of fungal organisms. CONCLUSIONS: An intact respiratory burst is present in the orbit of diabetics during infection with rhinocerebral mucormycosis. Possible mechanisms of peroxynitrite's microbicidal effects and reasons for a deficiency in diabetics are discussed. PMID- 11120579 TI - Resident tissue macrophages within the normal rat iris lack immunosuppressive activity and are effective antigen-presenting cells. AB - Despite extensive study of the numerous immunoregulatory mechanisms that contribute to the 'immune-privileged'nature of the anterior chamber (AC) of the eye, little is known of the functional nature of antigen-presenting cells (APC) present in the tissues adjoining the AC. In the present study, we have compared the antigen-presenting capacity of dendritic cells (DC) and macrophages isolated from the normal rat iris. Whereas iris DC exhibited a potent ability to stimulate resting allogeneic T cells in MLR cultures (an in-vitro correlate of the ability to induce primary T cell responses), resident iris macrophages displayed negligible MLR-stimulatory capacity. Significantly, iris macrophages could efficiently elicit proliferation of primed antigen-specific T cells (an in-vitro correlate of the ability to act as local APC in secondary responses). This antigen-presenting activity was approximately half that of fully 'mature' iris DC and considerably greater than that of freshly isolated iris DC. A key contributor to the effectiveness of resident iris macrophage antigen presentation was considered to be the absence of lymphocytostatic control of T cell proliferation exerted by these cells. The results indicate dichotomous but complementary roles for DC (immune surveillance) and macrophages (local antigen presentation in secondary responses) in this tissue. PMID- 11120580 TI - Use of PCR in endophthalmitis. AB - In eyes with suspected endophthalmitis, early diagnosis and appropriate treatment have been noted to be associated with a better visual outcome. Currently, however, confirmation of the diagnosis of endophthalmitis (bacterial and/or fungal) is dependent on conventional techniques of microbiological isolation of organisms which require between one and twelve days. Furthermore, many samples prove to be culture-negative. In order to improve the rate of microbiological diagnosis, PCR technology has been successfully applied to the detection of bacteria and fungi in ocular samples. Specific oligonucleotide primers have been used to detect the presence of pathogens, which have been subsequently identified using RFLP analysis, DNA sequencing, and/or cloning techniques. Results demonstrated that PCR-based methods are rapidly able to confirm the presence of pathogens with high specificity and sensitivity. PCR-based techniques have also been used to rule out with confidence the presence of pathogens, a unique advantage of this methodology. The use of molecular methods has significantly increased the number of intraocular samples from which a confirmed diagnosis is made and reduced the time to laboratory diagnosis. PCR-based methods promise to be useful diagnostic tools in the management of these patients, especially those from whom ocular samples prove to be culture-negative. PMID- 11120581 TI - Granulomatous anterior uveitis in a patient with CREST syndrome. AB - The CREST syndrome is a variant form of progressive systemic sclerosis. Apart from the occurrence of keratoconjunctivitis sicca, other types of ocular involvement associated with this variant are quite rare. We present the case of a 73-year-old woman with the CREST variant of progressive systemic sclerosis who developed unilateral granulomatous anterior uveitis. Systemic and laboratory testing failed to suggest evidence for any other associated systemic disease as a possible cause of the granulomatous uveitis. The inflammation was successfully controlled with topical steroids and mydriatics. While a small number of cases of uveitis have been reported in other variant forms of progressive systemic sclerosis, to date there have been no descriptions of uveitis associated with the CREST syndrome. PMID- 11120582 TI - Alzheimer's disease: amyloid beta-peptide antibody vaccine as plaque remover. PMID- 11120583 TI - His kinase or mine? histidine kinases through evolution. PMID- 11120584 TI - Cognitive empathy in inter-disciplinary research: the contrasting attitudes of plant breeders and molecular biologists towards rice. AB - I draw attention to the perceptions of and interactions between molecular biologists and scientists engaged in plant breeding in India, who have been attempting to employ molecular biology tools to understand and intervene to improve the rice crop. The present essay suggests that the concept of cognitive empathy is crucial for enabling basic scientists and applied scientists to begin to understand phenomena from the point of view of the other and from the point of view of society at large, and in arriving at solutions that are scientifically feasible and socially acceptable. Socialization into disciplinary cultures, organizational factors and individual anxieties seem to inhibit inter disciplinary collaboration. The majority of rice breeders and a small group of molecular biologists emphasize the relative merits of marker-assisted selection (MAS) in the near term vis-a-vis the currently controversial transgenic approach for rice crop improvement in India. PMID- 11120585 TI - Enhanced expression of a calcium-dependent protein kinase from the moss Funaria hygrometrica under nutritional starvation. AB - Among the downstream targets of calcium in plants, calcium-dependent protein kinases (CDPKs) form an interesting class of kinases which are activated by calcium binding. They have been implicated in a diverse array of responses to hormonal and environmental stimuli. In order to dissect the role of CDPKs in the moss Funaria hygrometrica, a polymerase chain reaction (PCR)-based approach was adopted to clone the gene. Using degenerate PCR primers against conserved regions of CDPKs, a 900 bp amplicon was obtained from the genomic DNA of Funaria. Southern hybridization under low stringency conditions indicated the presence of several CDPK related sequences in the Funaria genome. This observation is consistent with reports of multigene families of CDPKs in other plants. The 900 bp fragment was subsequently used to isolate a 2.2 kb partial genomic clone of the CDPK gene from Funaria. The genomic clone encodes an open reading frame (ORF) of 518 amino acids. Interestingly, unlike other CDPK genes from plants, the entire 1.5 kb ORF is not interrupted by introns. The deduced amino acid sequence of the Funaria gene shows extensive homology with CDPKs from higher plants, 73% identity with the Fragaria CDPK and 71% identity with CDPK isoform 7 of Arabidopsis. Phylogenetic analysis revealed that the Funaria CDPK is closer to the CDPKs from higher plants like strawberry and Arabidopsis as compared to those from lower plants such as the liverwort Marchantia, the green alga Chlamydomonas or another moss Tortula. Northern analysis shows enhanced expression of the CDPK transcript within 24-48 h of starvation for nitrogen, phosphorus or sulphur. So far the only other kinase which is known to be induced by nutrient starvation in plants is the wpk 4 which is a snf-1 related kinase (SnRKs). To our knowledge this is the first report that implicates a CDPK in the starvation response. PMID- 11120586 TI - Tyrosine phosphorylation of the human guanylyl cyclase C receptor. AB - Tyrosine phosphorylation events are key components of several cellular signal transduction pathways. This study describes a novel method for identification of substrates for tyrosine kinases. Co-expression of the tyrosine kinase EphB1 with the intracellular domain of guanylyl cyclase C (GCC) in Escherichia coli cells resulted in tyrosine phosphorylation of GCC, indicating that GCC is a potential substrate for tyrosine kinases. Indeed, GCC expressed in mammalian cells is tyrosine phosphorylated, suggesting that tyrosine phosphorylation may play a role in regulation of GCC signalling. This is the first demonstration of tyrosine phosphorylation of any member of the family of membrane-associated guanylyl cyclases. PMID- 11120587 TI - Domain III function of Mu transposase analysed by directed placement of subunits within the transpososome. AB - Assembly of the functional tetrameric form of Mu transposase (MuA protein) at the two att ends of Mu depends on interaction of MuA with multiple att and enhancer sites on supercoiled DNA, and is stimulated by MuB protein. The N-terminal domain I of MuA harbours distinct regions for interaction with the att ends and enhancer; the C-terminal domain III contains separate regions essential for tetramer assembly and interaction with MuB protein (IIIalpha and IIIbeta, respectively). Although the central domain II (the 'DDE' domain) of MuA harbours the known catalytic DDE residues, a 26 amino acid peptide within IIIalpha also has a non-specific DNA binding and nuclease activity which has been implicated in catalysis. One model proposes that active sites for Mu transposition are assembled by sharing structural/catalytic residues between domains II and III present on separate MuA monomers within the MuA tetramer. We have used substrates with altered att sites and mixtures of MuA proteins with either wild-type or altered att DNA binding specificities, to create tetrameric arrangements wherein specific MuA subunits are nonfunctional in II, IIIalpha or IIIbeta domains. From the ability of these oriented tetramers to carry out DNA cleavage and strand transfer we conclude that domain IIIalpha or IIIbeta function is not unique to a specific subunit within the tetramer, indicative of a structural rather than a catalytic function for domain III in Mu transposition. PMID- 11120588 TI - Sequence analysis of mitochondrial 16S ribosomal RNA gene fragment from seven mosquito species. AB - Mosquitoes are vectors for the transmission of many human pathogens that include viruses, nematodes and protozoa. For the understanding of their vectorial capacity, identification of disease carrying and refractory strains is essential. Recently, molecular taxonomic techniques have been utilized for this purpose. Sequence analysis of the mitochondrial 16S rRNA gene has been used for molecular taxonomy in many insects. In this paper, we have analysed a 450 bp hypervariable region of the mitochondrial 16S rRNA gene in three major genera of mosquitoes, Aedes, Anopheles and Culex. The sequence was found to be unusually A+T rich and in substitutions the rate of transversions was higher than the transition rate. A phylogenetic tree was constructed with these sequences. An interesting feature of the sequences was a stretch of Ts that distinguished between Ae-des and Culex on the one hand, and Anopheles on the other. This is the first report of mitochondrial rRNA sequences from these medically important genera of mosquitoes. PMID- 11120589 TI - On the patterns of abundance and diversity of macrolichens of Chopta-Tunganath in the Garhwal Himalaya. AB - A total of 3211 colonies of macrolichens, from twelve 50 m x 10 m plots distributed across four macrohabitat (vegetation) types between 1500 m-3700 m in the Chopta-Tunganath landscape of the Garhwal Himalaya, yielded 13 families with 15 genera and 85 species. Lobaria retigera stood out as a broad-niched generalist species with moderate levels of abundance in all the three major microhabitats, viz. rock, soil and wood across 83% of all the plots sampled, whereas Umbilicaria indica emerged as an abundantly occurring specialist confined to rock substrates. Heterodermia incana and Leptogium javanicum appeared to be rare members of the community as they were encountered only once during the field survey. Woody microhabitats turned out to be richer than rock and soil substrates for macrolichens. Amongst the macrohabitats, middle altitude (2500-2800 m) Quercus forest was richest in species and genera followed by high altitude (2900-3200 m) Rhododendron forest, higher altitude grasslands (3300-3700 m) and then the lower elevation (1500 m) Quercus forest. Species, genus and family level alpha- as well as beta-diversities were significantly correlated with each other, implying that higher taxonomic ranks such as genera may be used as surrogates for species thus facilitating cost- and time-effective periodic monitoring of the biodiversity of macrolichens. Dynamics of the diversity of lichen communities in relation to various forms of environmental disturbance including livestock grazing and tourism as dominant land use activities in the higher Himalaya need further research. PMID- 11120590 TI - Microarthropod community structures (Oribatei and Collembola) in Tam Dao National Park, Vietnam. AB - A study on the microarthropod community with special reference to species diversity of Oribatid and Collembola communities (Microarthropoda: Oribatei and Collembola) in Tam Dao National Park of Vietnam, a subtropical evergreen broad leaf alpine forest, was undertaken with the aim to explain how they are related to forest decline, and whether they can be used as bioindicators of forest plant succession. The results have shown that microarthropod community structures, particularly species diversity of oribatid and collembolan communities, are related to forest decline. Therefore they can be used as bioindicators of forest plant succession. In Tam Dao National Park, there was an inverse relation between species diversity of the oribatid and Collembola communities. The species diversity of the oribatid community gradually decreased with forest decline whereas the species diversity of the Collembola community gradually increased. PMID- 11120591 TI - Vocal signals in a tropical Avian species, the redvented bulbul Pycnonotus cafer: their characteristics and importance. AB - Acoustic signals play an important role in the lives of birds. Almost all avian species produce vocal signals in a variety of contexts either in the form of calls or songs or both. In the present study different types of vocal signals of the tropical avian species Pycnonotus cafer were characterized on the basis of their physical characteristics and context of production. This species used six types of vocal signals: contact signals, roosting signals, alarm signals, twittering signals, distress signals and begging signals. Two types of alarm signals are produced based on predation pressure. These signals are dissimilar in all physical characteristics except for dominant frequency. Although alarm signal type I and roosting signals are phonetically similar, they have completely different sonogram characteristics. PMID- 11120592 TI - Mechanism of store-operated calcium entry. AB - Activation of receptors coupled to the phospholipase C/IP3 signalling pathway results in a rapid release of calcium from its intracellular stores, eventually leading to depletion of these stores. Calcium store depletion triggers an influx of extracellular calcium across the plasma membrane, a mechanism known as the store-operated calcium entry or capacitative calcium entry. Capacitative calcium current plays a key role in replenishing calcium stores and activating various physiological processes. Despite considerable efforts, very little is known about the molecular nature of the capacitative channel and the signalling pathway that activates it. This review summarizes our current knowledge about store operated calcium entry and suggests possible hypotheses for its mode of activation. PMID- 11120593 TI - Environmental stressors and neuroimmunotoxicological processes. PMID- 11120594 TI - Cytokine production by naive and primary effector CD4+ T cells exposed to norepinephrine. AB - We recently showed that clones of Th1 cells, but not Th2 cells, expressed a functional beta-2-adrenergic receptor (beta2AR) and that either norepinephrine or the beta2AR agonist terbutaline stimulated this receptor to modulate the level of Th1 cytokines produced. In the present study, we show that norepinephrine and terbutaline stimulate the beta2AR to decrease the level of IL-2 produced by freshly isolated murine splenic naive CD4+ T cells from either Balb/C or DO11.10 transgenic mice and activated polyclonally with anti-CD3 and anti-CD28 mAbs. In contrast, the level of cytokines produced by primary effector Th1 and Th2 cells were unaffected when norepinephrine, terbutaline, or cAMP analogs were added at the time of restimulation. These results suggest that a diversity exists among CD4+ T-cell subsets with respect to the level of adrenergic receptor expression, responsiveness to cAMP, stage of cell differentiation, or a combination of the above. PMID- 11120595 TI - Restraint-induced modulation of allergic and irritant contact dermatitis in male and female B6.129 mice. AB - Recent studies in rats have indicated that acute restraint enhances cutaneous hypersensitivity. We hypothesized that acute restraint would also modulate the development of allergic and irritant dermatitis in mice and that these restraint induced changes would be reflected in the cutaneous cytokine profile and be gender-specific. For these studies, male and female B6.129 mice were sensitized and challenged with the contact sensitizer dinitrofluorobenzene or challenged with the irritant croton oil. Two-hour restraint was applied prior to chemical challenge. Restraint combined with chemical increased ear swelling in both genders in ACD, a change that was blocked by administration of RU-486 prior to restraint. Neither restraint nor RU-486 administration modulated development of ICD; however, IL-1beta was decreased by restraint in females only. TNF-alpha and IFN-gamma production were modified in ACD; TNF-alpha in both genders and IFN gamma in female mice only. Our data demonstrate that acute restraint increases serum corticosterone in B6.129 male and female mice to comparable levels. Restraint modulated the murine ear swelling in ACD, but not ICD, in both genders, and the change in the ear swelling response and cytokine production were, at least in part, corticosterone-dependent. PMID- 11120596 TI - Quantitative analysis of the neuroendocrine-immune axis: linear modeling of the effects of exogenous corticosterone and restraint stress on lymphocyte subpopulations in the spleen and thymus in female B6C3F1 mice. AB - The effects of exogenous corticosterone and restraint stress on the number and percentage of lymphocyte subpopulations in the spleen and thymus were evaluated. The data were used to generate linear models that describe the relationship between these parameters and the area under the corticosterone concentration vs time curve (AUC). Comparison of the models revealed that the number of nucleated cells in the spleen was decreased similarly by exogenous corticosterone and restraint (at equivalent corticosterone AUC values). However, exogenous corticosterone caused a greater decrease in cell number in the thymus than it did in the spleen. Corticosterone preferentially depleted CD4+CD8+ cells in the thymus, whereas the same corticosterone exposure produced by restraint stress did not. In the spleen, cell number for all major cell types was decreased by both treatments, but there were minor differences in the change in percentage of some subpopulations induced by exogenous corticosterone as compared to restraint. The models derived here provide quantitative data that indicate the magnitude of corticosterone and stress-induced effects on lymphocyte populations in the spleen and thymus. These results have mechanistic implications, and they may be useful in future efforts to extrapolate from mouse to human by completing a risk assessment parallelogram. PMID- 11120597 TI - Age-dependent cytokine responses: trimethyltin hippocampal injury in wild-type, APOE knockout, and APOE4 mice. AB - In this study, the hippocampal neurotoxicant trimethyltin (TMT) was used to examine possible differential susceptibility associated with the apolipoprotein E genotype. Mice-wild type (C57BL6J), APOE knockout, and APOE4 transgenic-received either saline or TMT (2 mg/kg, ip) at either 21 days or 8 months of age. At both ages, similar mRNA levels were seen in the hippocampus across genotypes for ICAM 1, A20, and MAC-1. GFAP mRNA was higher in the APOE knockouts and APOE4 as compared to wild-type mice. Within 24 h, TMT produced cell death of hippocampal dentate granule neurons and mild astrogliosis in all animals. In 21-day-old mice, TMT exposure significantly increased mRNA levels for ICAM-1 and MIP-1alpha in all genotypes. EB-22, GFAP, TNFalpha, and TGF-beta1 levels were significantly elevated in both wild-type and APOE knockout mice following TMT. At 8 months of age, genotype specific differences were observed. mRNA levels for GFAP, TNFbeta, TNFalpha, and MIP-1alpha were increased in both APOE knockout and APOE4 mice compared to wild-type mice. TMT exposure significantly increased mRNA levels for GFAP and MIP-1alpha in all animals. TNFalpha mRNA levels were increased in wild type and APOE4 mice while EB22 mRNA levels were increased in both the APOE knockout and APOE4 mice but not wild-type mice. These data suggest an age dependent effect on both microglia early inflammatory responses to injury associated with the APOE genotype. PMID- 11120598 TI - Immunotoxic effects of inorganic lead on host resistance of mice with different circling behavior preferences. AB - We have observed differential immune responses in mice with different circling preferences, which are posited to reflect interindividual immune response differences influenced by brain laterality effects on neuroimmune circuits. In this study, we have investigated the influence of inorganic lead (Pb) and/or Listeria monocytogenes (LM) infection on the cytokine and corticosterone (CORT) levels of mice grouped by lateralized behavior. Pb increased the LM susceptibility of mice with both left (LC)- and right-circling (RC) preferences; however, Pb did not inhibit the host resistance of mice with no circling preference (NP mice). The basal serum IFNgamma levels were lowered in all groups after Pb exposure, which coincided with a decrease in host resistance in LC and RC mice, but not NP mice. Pb also altered the basal serum CORT levels, and these changes appear to correlate better with changes in the host resistance of all groups. The basal CORT levels were significantly lowered by Pb in mice with a circling preference, and Pb significantly suppressed the host resistance of mice with a circling preference. However, Pb slightly increased the serum CORT level of NP mice, and their host resistance was slightly improved by Pb. After infection, the increase in CORT levels was associated with an increase in the serum IL-6 levels, which may reflect cytokine influences on the hypothalamic pituitary-adrenal axis. At 3 days after infection, the serum IL-6 level seems to be a good indicator of the severity of the infection. We suggest that environmental stressors can reorder the observed differential susceptibility to LM in mice with different circling preferences, in that relatively resistant mice (RC mice) become more susceptible than NP mice after exposure to Pb. The results suggest that environmental stressors may have differential effects among individuals with endogenous differences in their neuroimmune circuits, since brain laterality is known to influence immune functions. PMID- 11120599 TI - Nuclear-cytoplasmic translocation of tRNA. PMID- 11120600 TI - Differential effects of bcl-2 on cell death triggered under ATP-depleting conditions. AB - The intracellular ATP concentration decides on the onset of either apoptosis or necrosis in Jurkat cells exposed to death stimuli. Bcl-2 can block apoptotic demise, which occurs preferably under conditions of high cellular ATP levels. Here, we investigated the effects of Bcl-2 on the necrotic type of cell demise that prevails under conditions of energy loss. ATP levels were modulated by using mitochondrial inhibitors, such as rotenone or S-nitrosoglutathione, in medium either lacking glucose or supplemented with glucose to stimulate glycolytic ATP generation. Under conditions of ATP depletion, staurosporine (STS) induced >90% necrosis in vector control-transfected cells, whereas bcl-2-transfected cells were protected. Thus, the antiapoptotic protein Bcl-2 can reduce the overall amount of cell death in ATP-depleted cells regardless whether it occurs by apoptosis or necrosis. Cytochrome c release, normally preceding STS-induced necrosis, was also inhibited by Bcl-2. However, Bcl-2 did not prevent an initial STS-induced drop of the mitochondrial membrane potential (DeltaPsi(m)). Therefore, the mechanisms whereby Bcl-2 prevents cell death and favors retention of cytochrome c in the mitochondria require neither the maintenance of mitochondrial DeltaPsi nor the maintenance of normal ATP levels. PMID- 11120601 TI - The farnesyl transferase inhibitor SCH 66336 induces a G(2) --> M or G(1) pause in sensitive human tumor cell lines. AB - SCH 66336 is a potent farnesyl transferase inhibitor (FTI) in clinical development. It efficiently prevents the membrane association of H-ras, but not K or N-ras. Yet, in soft agar, it reverts the anchorage-independent growth of human tumor cell lines (hTCLs) harboring H-ras, K-ras, and N-ras mutations, implying that blocking farnesylation of proteins besides ras may be responsible for this effect. Experiments show that SCH 66336 altered the cell cycle distribution of sensitive human tumor cells in two distinct ways. Most sensitive hTCLs accumulated in the G(2)-->M phase after the FTI treatment, but those with an activated H-ras accumulated in G(1) phase, suggesting that the biological effects induced by FTIs in cells with an activated H-ras are distinct from other sensitive cells. A careful genotypic comparison of the hTCLs revealed that those cells with wild-type p53 are especially sensitive to the FTIs. In these cells p53 and its downstream target gene p21(Cip1) are induced after treatment with SCH 66336 for 24 h. These data suggest that cell cycle effects, either G(1) or G(2)- >M accumulation, and p53 status are important for mediating the effects of FTIs on tumor cells. PMID- 11120602 TI - MMP-2 colocalizes with caveolae on the surface of endothelial cells. AB - We examined the spatial distribution of MMP-2 on the surface of human endothelial cells using immunofluorescence and confocal microscopy. Staining endothelial cells with MMP-2-specific antibodies revealed a punctate labeling at the basolateral side of the cell periphery, which colocalized with patches of caveolin-1, a major constituent of the caveolae. This colocalization was confirmed by immunogold electron microscopy. MT1-MMP, TIMP-2, and the alphavbeta3 integrin exhibited a similar pattern of staining, with pericellular patches that colocalized with either MMP-2 or caveolin-1. The presence of MT1-MMP and TIMP-2 in caveolae patches could be seen only after treatment with concanavalin A, which induced MMP-2 activation but had no noticeable effect on the pattern or intensity of MMP-2 immunostaining. In contrast, MMP-9 and TIMP-1 staining showed a pattern completely different from that of MMP-2 and TIMP-2, with positive spots uniformly distributed throughout the cell body. Our data show that MMP-2, its activator the MT1-MMP, and its proposed receptor, the alphavbeta3 integrin, are all targeted to the same membrane microdomains on the endothelial cell, thereby restricting matrix proteolysis to a limited microenvironment at the cell surface. PMID- 11120603 TI - Characterization of adriamycin-induced G2 arrest and its abrogation by caffeine in FL-amnion cells with or without p53. AB - We investigated the effect of Adriamycin on FL-amnion (FL) cells. After treatment with the drug, the cells arrested at G2, but we did not detect an increase in the p21 levels. We established a p53-deficient derivative of these cells, in which G2 arrest also occurred after treatment with Adriamycin, suggesting that the arrest we observed in these cells is independent of the p53 pathway. Low doses of Adriamycin (100-200 ng/ml) induced G2 arrest, while late S-phase arrest was observed at high doses (500-1000 ng/ml) in both FL and p53-deficient FL cells. Accumulation of cyclin B1 was detected only in cells arrested at G2, and not in those arrested at S phase, suggesting that the S-phase checkpoint functioned efficiently even in p53-deficient FL cells. In both cell lines, caffeine-induced activation of CDC2 kinase was detected only in cells arrested at G2 and CDC2 kinase-activated cells died exhibiting features of apoptosis. CDC2 kinase activation was inhibited by cycloheximide. Furthermore, cycloheximide inhibited activation of CDK2:cyclin A, which normally precedes CDC2 kinase activation in caffeine-treated cells. These results suggest that p53 and p21 do not have special roles in the S- and G2-phase checkpoints and that CDK2:cyclin A could be the target of the G2-phase DNA damage checkpoint. PMID- 11120604 TI - Cell density regulates tyrosine phosphorylation and localization of focal adhesion kinase. AB - We have investigated tyrosine phosphorylation of cellular proteins at different cell densities. A tyrosine-phosphorylated protein of 120 kDa was detected when cells were plated sparsely. The phosphorylation level of the protein gradually declined as the cells were plated at higher densities or when the sparsely plated cells approached confluence. This density-dependent phosphorylation was also associated with cell attachment since it disappeared when the cells were detached from plates or when the cells were cultured in suspension. Immunoblotting and immunoprecipitation analyses with specific antibodies revealed that the 120-kDa protein corresponded to the focal adhesion kinase (FAK) and the protein level of FAK was not altered at different cell densities. In vitro kinase assays demonstrated that the kinase activity of FAK decreased with increasing cell densities in parallel with its dephosphorylation. Cell density also affects localization of FAK associated with rearrangement of actin stress fibers. At low cell densities, FAK and actin stress fiber are distributed around the periphery of cells while they are dispersed over the ventral surface in high-density cells. Finally, the density-regulated tyrosine phosphorylation and localization of FAK appeared to be mediated by an insoluble factor produced by high-density cells. PMID- 11120605 TI - Proteomic detection of changes in protein synthesis induced by fibroblast growth factor-2 in MCF-7 human breast cancer cells. AB - Fibroblast growth factor-2 (FGF-2) is a potent regulator of breast cancer cell growth through stimulation of tyrosine kinase receptors and activation of the mitogen-activated protein kinase cascade. In the present study, we have investigated changes in protein synthesis induced by FGF-2 stimulation of the prototypic human breast cancer cell line MCF-7. Using high-resolution two dimensional electrophoresis of (35)S amino acid metabolically labeled proteins and computerized analysis of 2D autoradiograms, we found that four proteins were up-regulated within the first 12 h of FGF-2 stimulation. Mass spectrometry analysis (MALDI-TOF and MS-MS) of tryptic fragments and database searches allowed the identification of these FGF-2-regulated proteins as the heat shock proteins HSP90 and HSP70, the proliferating cell nuclear antigen (PCNA), and the transcriptionaly controlled tumor protein (TCTP). We then analyzed the distribution of these proteins in various cancerous and normal breast epithelial cells. Interestingly, the four FGF-2-regulated proteins were found to be constitutively up-regulated in ras-transfected MCF-7 cells, indicating their relevance to the up-regulation of cellular proliferation. Moreover, HSP90 and PCNA were found at higher levels in cancerous cells than in normal cells. The role of HSP90 was further investigated using the specific inhibitor geldanamycin. We showed that the functionality of HSP90 is strictly required in order to obtain FGF-2 mitogenic stimulation in MCF-7 cells, indicating the crucial role played by this molecular chaperone in the control of breast cancer cell growth. Finally, these results show that proteomic analysis is a valuable method for identifying potential markers or therapeutic targets related to cancer growth. PMID- 11120606 TI - TGFbeta and BMP-2 activation of the OPN promoter: roles of smad- and hox-binding elements. AB - Members of the transforming growth factor superfamily are known to transduce signals via the activation of Smad proteins. Ligand binding to transmembrane cell surface receptors triggers the phosphorylation of pathway-specific Smads. These Smads then complex with Smad 4 and are translocated to the nucleus where they effect gene transcription. Smads 1 and 4 were recently demonstrated to mediate BMP activation of the OPN promoter by inhibiting the interaction of Hoxc-8 protein with a Hox-binding element. While previous studies have indicated that specific DNA sequences are recognized by Smad complexes in several promoters, the role of Smad-binding elements (SBEs) in activation of the OPN promoter by members of the TGFbeta superfamily has not been previously evaluated. In this study we tested the hypothesis that a putative Smad-binding region containing the sequence AGACTGTCTGGAC is involved in the activation of the OPN promoter by members of the TGFbeta superfamily. Functional analyses demonstrated that the both the HBE- and Smad-binding region were involved in BMP-2-induced activation of the promoter, whereas, the HBE appeared to be the primary region involved in activation by TGFbeta. Deletion of the first 9 bases in the Smad-binding region substantially reduced BMP-2-mediated activation of the promoter. These results strongly suggest that both the Hox- and the Smad-binding regions play a role in BMP-2-induced activation of the OPN promoter. PMID- 11120607 TI - Antifungal drug resistance in Aspergillus. PMID- 11120608 TI - Use of polymerase chain reaction (PCR) and DNA probe hybridization to determine the Gram reaction of the infecting bacterium in the intraocular fluids of patients with endophthalmitis. AB - OBJECTIVES: To evaluate polymerase chain reaction (PCR) combined with DNA probe hybridization to determine the Gram reaction of the bacterium in intraocular specimens from patients with infectious endophthalmitis. METHODS: Fifty-seven intraocular specimens - 17 aqueous humor (AH) and 40 vitreous fluid (VF) - from 55 patients with clinically diagnosed infectious endophthalmitis and 25 control intraocular specimens from non-infectious ocular disorders (10 AH and 15 VF) were evaluated by microscopy, culture and PCR-DNA probe hybridization to detect the Gram reaction of the bacterium. RESULTS: PCR-DNA probe hybridization was specific and sensitive to detect 30 fg of both gram-positive and gram-negative bacterial DNA. None of the controls showed bacteria by microscopy, culture or PCR. Of the 57 intraocular specimens, conventional microbiological methods could detect a bacterial aetiology in 32 (56.1%), while PCR-DNA probe hybridization could detect 52 (91.2%) specimens. This difference was statistically significant (P= 0.003). In bacteriologically positive specimens, there was absolute correlation of the Gram reaction between the results of smear and culture methods and PCR-DNA probe hybridization. Of the 25 bacteriologically negative specimens, 20 (80%) were positive by PCR-DNA probe hybridization, of which seven (35%) were gram-positive, 12 (60%) gram-negative and one (5%) positive by both. Results of PCR on AH and VF were not significantly different. CONCLUSION: PCR and DNA probe hybridization to determine the Gram reaction of the bacterium in intraocular fluids is a specific and sensitive method in the diagnosis of bacterial endophthalmitis. AH is an ideal specimen for PCR, since its collection is a simple and safe office procedure. PMID- 11120609 TI - Rhodococcus equi and HIV-1 infection in Uganda. AB - OBJECTIVES: To describe three cases of Rhodococcus equi infection in a cohort of HIV-1 infected adults in Entebbe, Uganda and to compare this to the rates and presentation of tuberculosis in this cohort. METHODS: Consecutive HIV-1 infected adults registering with a community HIV/AIDS clinic in Entebbe were enrolled in a cohort between October 1995 and June 1998 as part of an intervention trial of pneumococcal polysaccharide vaccine. Participants were routinely reviewed every 6 months and had open access to the clinic when unwell. Standard protocols were followed for investigation and management of illness. Microbiological investigations followed standard procedures. RESULTS: 1372 (71% female) study participants were followed for 2141 person years of observation (pyo). Rhodococcus equi was isolated from three study participants from blood, a lymph node aspirate and stool. The individuals were undergoing investigation of acute pneumonia, acute cough with cervical lymphadenopathy and chronic fever with wasting, respectively. The clinical features of these cases are described. All had a CD4 T-cell count of <300/ml. The rate of R. equi infection in the cohort was 1.4/1000 pyo. There were 132 cases of pulmonary and extrapulmonary tuberculosis in the cohort which were diagnosed either microbiologically or clinically. The rate of laboratory confirmed mycobacterial disease was 50.1/1000 pyo. The ratio of mycobacterial disease to R. equi disease was 36:1 (95% CI 11 113:1). CONCLUSIONS: Rhodococcus equi infection occurs in HIV-1 infected adults in Africa. The infection is clinically indistinguishable from pulmonary and extra pulmonary tuberculosis in the cohort described here. Although the rate of R. equi disease is much less than that of tuberculosis, it is important to consider it in the differential diagnosis of tuberculous infection in cases which are smear negative. Rhodococcus equi infection is probably underdiagnosed in Africa due to a lack of microbiological facilities and its resemblance to common commensal organisms. PMID- 11120610 TI - Nocardial infection as a complication of HIV in South Africa. AB - OBJECTIVES: To assess the occurrence, clinical and microbiological features of nocardial infections complicating HIV in Soweto, South Africa. METHODS: A prospective study was carried out over a 2-year period. Patients were identified after isolation of Nocardia spp. from a clinical specimen. Clinical details were recorded. The nocardial isolates were identified to species level and susceptibility tests performed. RESULTS: Ten patients were identified as having nocardial disease complicating HIV. Clinical presentations were pulmonary (five patients), pulmonary and cerebral (one patient), cerebral (one patient) and skin and soft tissue infection of the lower limb (three patients). Three infections were fatal. The isolates were Nocardia asteroides (seven patients), N. farcinica (two patients) and Nocardia spp. (one). Isolates of N. farcinica demonstrated opacification of Middlebrook agar. All isolates were sensitive to amikacin and minocycline. Most nocardial isolates were susceptible to cefotaxime, imipenem and coamoxiclav. In vitro resistance to cotrimoxazole was present in five. CONCLUSIONS: Nocardial infection occurs as a complication of HIV infection in the Republic of South Africa. Pulmonary cases may be difficult to distinguish from tuberculosis. Nocardia asteroides is the most common species isolated. Nocardia farcinica has resistance to multiple antibacterial agents and demonstrates opacification of Middlebrook agar, a useful screening test for this species. Agents with good in vitro antinocardial activity were amikacin, minocycline, cefotaxime, imipenem and coamoxiclav. There was a high level of resistance in vitro to cotrimoxazole. PMID- 11120611 TI - Downward shift in serum IgM with Helicobacter pylori seropositivity. AB - OBJECTIVE: To determine whether Helicobacter pylori infection is associated with premature immune ageing, with respect to circulating immunoglobulins. METHODS: Serum immunoglobulin classes and H. pylori anti-urease antibody were measured in 205 subjects (aged 30-89 years), obeying inclusion/exclusion criteria. RESULTS: IgM decreased (P<0.001) by 0.9 (95% C.I. 0.3, 1.4)% per year, H. pylori seropositivity having an effect equivalent to 25 years of ageing (P<0.02). IgA increased by 0.5 (0.1, 0.8)% per year (P<0.007), IgG being unaffected by age. Seropositivity had no effect on IgA or IgG. CONCLUSIONS: Increasing age and H. pylori seropositivity are each associated with a downward shift in circulating IgM. If clinical extrapolation is justified, H. pylori eradication may be important in combating susceptibility to infection in old age. PMID- 11120612 TI - Genetic diversity in Proteus mirabilis isolates found in the urinary tract of rheumatoid arthritis patients. AB - OBJECTIVE: Elevated levels of anti-Proteus antibodies but not antibodies to E. coli have been reported in patients with rheumatoid arthritis (RA). The suggestion has been made that P. mirabilis may have a role in the aetiopathogenesis of rheumatoid arthritis. The aim of this study was to determine whether there are differences at the genetic level inisolates of P. mirabilis obtained from controls and RA patients. METHODS: A blind study was performed whereby P. mirabilis isolates obtained from urinary cultures of RA patients and controls were analysed using RAPD PCR. Isolates were then grouped on the basis of their DNA band profile after agarose gel electrophoresis, thereby allowing the composition of the Proteus population in the urinary tract to be analysed at the genetic level. RESULTS: Fourteen different DNA band profiles were obtained from the 93 isolates tested: 70% of these isolates fell into only five of the 14 groups and approximately 25% of all isolates fell into one group. No differences were observed in the frequency of isolates from either control or RA subjects. CONCLUSIONS: There is genetic diversity in P. mirabilis populations found in the urinary tract, but there are no differences in the frequency of these bacteria between RA patients and controls. PMID- 11120613 TI - Tuberculosis transmission in the family. AB - OBJECTIVE: To evaluate the risk of tuberculosis (TB) transmission from family members with infectious TB to other family members, and to examine whether household contact investigations had an impact on tuberculosis patterns. DESIGN: Under the direction of the Taipei Municipal Chronic Disease Hospital, 12 full time public health nurses recruited the household contacts of TB patients. Chest X-ray examination was recommended for adult contacts. Child contacts received the Mantoux tuberculin skin test, and radiography was recommended if the results were positive. SETTING: Family contacts of all TB index patients who attended one of 29 hospitals in Taipei, from July 1993 through June 1996. The medical records of index patients were obtained from the National Tuberculosis Registry. RESULTS: During the study period, the families of 3903 index patients, comprising 11873 contacts, were investigated. Among these, 4595 received radiography, for a response rate of 38.7%. Of these, 284 had active pulmonary disease: 188 (66.3%) had minimal disease, 79 (27.8%) had moderately advanced disease, and only 17 (5.9%) had far advanced disease. Overall, the index patients had more advanced TB: only 1261 (32.3%) had minimal disease, while 2022 (51.8%) had moderately advanced disease and 620 (15.9%) had far advanced disease. CONCLUSIONS: These data show a relatively high risk of intrafamily TB transmission. Our findings also show that family contact investigations may help to diagnose TB in earlier stages. Such an approach should greatly reduce the number of new TB cases and speed eradication of the disease. PMID- 11120614 TI - Highly active antiretroviral therapy normalizes the potential for MIP-1alpha production in HIV infection. AB - DESIGN: The CC chemokines RANTES, MIP-1alpha and MIP-1beta are ligands for CCR5, which has been identified as the principal co-receptor for macrophage tropic strains of HIV-1. This study investigated whether the inducible levels of RANTES, MIP-1alpha and MIP-1beta produced by cultured whole blood samples related to different rates of progression of HIV infection and to the introduction of Nelfinavir-based highly active anti-retroviral therapy (HAART). METHODS: Study subjects were HIV-positive and categorized as "slow progressors" (n= 8) or as "fast progressors" (n= 7); the latter group were treated with HAART. MIP-1alpha, MIP-1beta and RANTES production was determined using commercial ELISA kits. RESULTS: The inducible production of MIP-1alpha by whole blood cells in culture was significantly depressed in patients starting therapy compared with "slow progressors" and "normal donors". The levels of MIP-1alpha significantly increased with therapy at 12 weeks compared with pre-HAART levels (P= O.05) and became comparable to that of "normals" and "slow progressors". Differences in the inducible levels of MIP-1beta and RANTES for the separate subject groups were not significant. CONCLUSIONS: The increase in inducible MIP-1alpha production following HAART might suggest a role for the chemokines in HIV disease, either for monitoring the outcome of therapy of HIV disease, or as a direct therapeutic intervention. PMID- 11120615 TI - Non-typhoidal Salmonella bacteraemia without gastroenteritis: a marker of underlying immunosuppression. Review Of cases at St. Thomas' Hospital 1970-1999. AB - OBJECTIVES: To classify non-typhoidal salmonella bacteraemia according to clinical presentation, and to study how this correlates with the presence of underlying immunosuppression. METHODS: We analysed data collected prospectively for all 82 cases of non-typhoidal salmonella bacteraemia presenting to St. Thomas' Hospital between 1970 and 1999. RESULTS: Patients presented with one of three syndromes: diarrhoea, an extra-intestinal focus of infection, or isolated fever with no focus. Only 18% of those with diarrhoea had underlying immunosuppression, compared with 80% of those with extra-intestinal focal infections (P= 0.001) and 80% of those with no focus (P= 0.0001). There was no significant association between salmonella serotype and underlying immunosuppression. Salmonella enteritidis isolates, especially phage type 4, increased significantly during the last decade (P= 0.001). The presentation of non-typhoidal salmonella bacteraemia in the absence of diarrhoea prompted the diagnosis of HIV in two patients. CONCLUSION: Underlying immunosuppression should be excluded in patients presenting with non-typhoidal salmonella bacteraemia in the absence of gastroenteritis. This may lead to an earlier diagnosis of HIV. PMID- 11120616 TI - Intra-familial evidence of horizontal transmission of hepatitis B virus surface antigen mutant G145R. AB - OBJECTIVES: To provide intra-familial evidence on the horizontal transmission of hepatitis B virus surface antigen (HBsAg) mutant G145R. METHODS: Serum samples from family members of 10 vaccinated infants who carried this G145R mutant were collected. The presence of the mutant was analysed by polymerase chain reaction (PCR) and sequencing. RESULTS: The G145R mutant was identified in family members of three of the 10 infants. In family 1, the mutant found initially in child 1 was identified in another child and the father. In families 2 and 3, the G145R mutant detected previously in child 1 was detected in the father. Additional mutations in HBsAg were identified in at least two members in family 1 and 2, suggesting horizontal transmission of the mutant among them. The G145R mutant was found in samples with high levels of neutralizing antibody against HBV (anti HBs). In addition, liver damage was seen in one G145R carrier infant. CONCLUSIONS: The G145R mutant could be transmitted horizontally among family members, and this could occur in the presence of high levels of anti-HBs. Improvement of detection system for the G145R and other HBsAg mutant will be needed for their effective control. PMID- 11120617 TI - Zygomycosis in relapsed acute leukaemia. AB - We would like to report the use of liposomal amphotericin in eradicating mucormycosis in two patients who had relapsed acute leukaemia. The first patient with relapsed acute myeloid leukaemia developed a rapidly expanding solitary necrotic neck lesion associated with opacity of maxilliary sinus at a time when he was profoundly pancytopenic following high dose chemotherapy. The second patient was a 3-year-old boy with pre-B acute lymphoblastic leukaemia who developed a central nervous system relapse whilst on his first line treatment and was treated with more aggressive chemotherapy on the Medical Research Council Relapse Protocol. During a period of profound pancytopenia following re-induction therapy, including high dose steroids and prolonged course of antibiotics for proven septicaemia, he developed periorbital swelling and proptosis and a clinical diagnosis of rhinocerebral mucormycosis was made. Both patients were treated with high doses of liposomal amphotericin (Ambisome Nexstar). The doses were escalated to 10 and 15 mg/kg/day, resulting in successful eradication of the mucormycosis. PMID- 11120618 TI - Tuberculosis as a cause of recurrent fever of unknown origin. AB - Recurrent fever constitutes a diagnostic challenge for clinicians, due mainly to the intermittent nature of the fever that results in incomplete investigations. We describe three patients with recurrent fever thought to be due to tuberculosis, and review the 14 previously reported cases who fulfil the criteria of recurrent fever for at least 1 month's duration. The median duration of symptoms before diagnosis was 5 months, and the duration of the febrile bouts ranged from a few hours to 1 week. The most common complaints were constitutional symptoms and abdominal pain, and most patients had significant underlying conditions. The mortality rate was 31%, and was limited to the earlier cases. Routine laboratory studies are not very helpful for the diagnosis of this condition, and chest radiographs showed some alteration in half the cases at the time of diagnosis, although in some cases represented old, healed lesions. PPD testing was positive in most cases, particularly in those without underlying conditions. Empirical antituberculous therapy should be considered in cases of recurrent fever, especially in areas of high prevalence or in patients with predisposing conditions. PMID- 11120619 TI - Successful non-surgical treatment of disseminated polymicrobial fungal infection in a patient with pancytopenia and graft-versus-host disease. AB - Invasive fungal infections after bone marrow transplantation have an extremely poor prognosis. Surgical excision in combination with antifungal therapy is considered necessary for treatment, especially for central nervous system (CNS) infection. We describe successful medical management with lipid complex amphotericin B (ABLC) and itraconazole, without surgical excision, of disseminated fungal infection involving the lungs and CNS in a patient with pancytopenia and graft-versus-host disease. PMID- 11120620 TI - Candida tropicalis pacemaker endocarditis. AB - A rare case of Candida tropicalis pacemaker endocarditis was diagnosed in a 77 year-old male who presented with lethargy. The organism was isolated from cultures of blood and vegetations on the tricuspid valve, interatrial septum and the pacing wire removed at surgery. The postoperative course was stormy and he succumbed to multiorgan failure. PMID- 11120621 TI - Fatal haemoptysis induced by invasive pulmonary aspergillosis in patients with acute leukaemia during bone marrow and clinical remission: report of two cases and review of the literature. AB - We describe two patients with acute leukaemia who died of massive haemoptysis caused by invasive pulmonary aspergillosis (IPA). The fatal event occurred during the period of bone marrow remission which followed chemotherapy-induced neutropenia. This is a rare complication. We were able to find additional 17 similar cases in the English literature, which we review. Clinically, the picture consisted of unremitting fever with profound and prolonged neutropenia, cough and dyspnoea. Both our patients were treated with broad-spectrum antibiotics, fluconazole and amphotericin B. An upper lobe infiltrate in one case, and a progressive pleural effusion in the other, were late findings on chest radiographs during the period of bone marrow recovery. Both patients succumbed to sudden massive haemoptysis during the period of bone marrow and clinical improvement. In conclusion, patients with acute non-lymphoid leukaemia are at significant risk for IPA-induced fatal haemoptysis during bone marrow and clinical remission. A high index of suspicion should be sustained throughout the entire clinical course. In view of the potential fatal outcome, aggressive diagnostic and treatment efforts are mandatory. PMID- 11120622 TI - Letter in response to article in Journal of Infection (Davies et al. Length of time to laboratory diagnosis 1999; 39; 205-208). PMID- 11120623 TI - Prosthetic valve endocarditis due to Streptococcus vestibularis. PMID- 11120624 TI - Propionibacterium granulosum: a rare cause of endocarditis. PMID- 11120625 TI - Outbreak of gastroenteritis caused by echovirus type 6 in an orphanage in Japan. PMID- 11120626 TI - Clostridium septicum: a malign pathogen. PMID- 11120627 TI - Hepatic cryoablation-induced acute lung injury: histopathologic findings. AB - We have previously shown that hepatic cryoablation (cryo), but not partial hepatectomy, induces a systemic inflammatory response, with distant organ injury and overproduction of NF-kappaB-dependent cytokines. Serum tumor necrosis factor alpha (TNF-alpha) and macrophage inflammatory protein-2 (MIP-2) levels are markedly increased 1 h and beyond after cryo compared with partial hepatectomy where no elevation occurs. NF-kappaB activation (by electrophoretic mobility shift assay) is strikingly increased in the noncryo liver (but not in the lung) at 30 min and in both the liver and lung tissue 1 h after cryo, returning to the baseline by 2 h and beyond. The current study investigated the histopathologic changes associated with cryoablation-induced acute lung injury. Animals underwent 35% hepatic resection or a similar volume hepatic cryo and were sacrificed at 1, 2, 6, and 24 h. Pulmonary histologic features were assessed using hematoxylin and eosin and immunoperoxidase staining with a macrophage-specific antibody (anti lysozyme, 1:200 dilution, Dako, Carpinteria, CA). The following features were graded semiquantitatively (0-3): perivascular lymphoid cuffs, airspace edema and hemorrhage, margination of neutrophils within pulmonary vasculature, and the presence of macrophages with foamy cytoplasm in the pulmonary interstitium. Hepatic resection (n = 21) resulted in slight perivascular edema at 1, 2, 6, and 24 h post-resection, but there were no other significant changes. Pulmonary findings after hepatic cryo (n = 22) included prominent perivascular lymphoid cuffs 1 and 2 h following hepatic injury that were not present at any other time point (P 0.01). Marginating PMNs and foamy macrophages were more common after cryo at all time points (P<0.05, cryo vs resection). Severe lung injury, as evidenced by airspace edema and parenchymal hemorrhage, was present in four of six (67%) animals at 24 h (P 0.03). In follow-up studies immediate resection (n = 15) of the cryo-treated liver prior to thawing prevented the pulmonary changes. The findings of pulmonary perivascular interstitial macrophages 2 h following hepatic cryo suggests that hepatic cytokine production may induce downstream recruitment of pulmonary macrophages, which may contribute to subsequent severe lung injury. This study suggests that a soluble mediator from direct liver injury leads to neutrophilic lung inflammation and this is associated with the thawing phase of cryoablation. PMID- 11120628 TI - Radiofrequency ablation treatment of refractory carcinoid hepatic metastases. AB - BACKGROUND: Our institution has experienced excellent success using hepatic artery embolization for treating symptoms and slowing tumor progression for patients with unresectable hepatic metastases for carcinoid tumors. Our previous treatment strategies used hepatic artery embolization alone, examining control of symptoms and dependence on octreotide therapy. However, some patients exhibit hepatic metastases that are unresponsive to embolization. This report describes the use of radiofrequency ablation (RFA) as salvage treatment for these refractory metastases. METHODS: Thirteen patients with unresectable bilobar hepatic metastases from biochemically confirmed carcinoid tumors were treated with selective hepatic artery embolization using Lipiodol/Gelfoam between 1994 and 2000. Three patients developed symptoms resistant to embolization treatment resulting from progression of existing metastases or development of new metastases. These patients underwent surgical exploration and intraoperative ultrasound of their refractory lesions, followed by treatment with RFA. Tumor size, symptoms of carcinoid syndrome, and octreotide requirements were monitored postoperatively. RESULTS: Median follow-up for the three patients treated with RFA was 6 months. During the first 3-month interval following RFA, all three patients demonstrated decrease in the size of treated lesions. Using our previously developed symptom scoring system, all three patients demonstrated decreased symptoms following treatment. One patient was able to discontinue octreotide treatment, and the other two patients required decrease octreotide dosages. CONCLUSIONS: This study demonstrates that utilization of RFA treatment for carcinoid metastases refractory to hepatic artery embolization may represent a useful adjunct for symptomatic control, decreased octreotide dependence, and slowing of disease progression. PMID- 11120629 TI - Laparoscopic revision of failed antireflux operations. AB - BACKGROUND: A small number of patients fail fundoplication and require reoperation. Laparoscopic techniques have been applied to reoperative fundoplications. We reviewed our experience with reoperative laparoscopic fundoplication. METHODS: Reoperative laparoscopic fundoplication was undertaken in 28 patients, 19 F and 9 M, of mean age 56 years +/- 12. Previous antireflux procedures included 19 open and 12 laparoscopic antireflux operations. RESULTS: Symptoms were heartburn (90%), dysphagia (35%), and atypical symptoms (30%%). The mean interval from antireflux procedure to revision was 13 months +/- 4.2. The mean DeMeester score was 78+/-32 (normal 14.7). Eighteen patients (64%) had hiatal breakdown, 17 (60%) had wrap failure, 2 (7%) had slipped Nissen, 3 (11%) had paraesophageal hernias, and 1 (3%) had an excessively tight wrap. Twenty-five revisions were completed laparoscopically, while 3 patients required conversion to the open technique. Complications occurred in 9 of 17 (53%) patients failing previous open fundoplications and in 4 of 12 patients (33%) failing previous laparoscopic fundoplications and included 15 gastrotomies and 1 esophagotomy, all repaired laparoscopically, 3 postoperative gastric leaks, and 4 pneumothoraces requiring tube thoracostomy. No deaths occurred. Median length of stay was 5 days (range 2-90 days). At a mean follow-up of 20 months +/- 17, 2 patients (7%) have failed revision of their fundoplications, with the rest of the patients being essentially asymptomatic (93%). CONCLUSIONS: The results achieved with reoperative laparoscopic fundoplication are similar to those of primary laparoscopic fundoplications. Laparoscopic reoperations, particularly of primary open fundoplication, can be technically challenging and fraught with complications. PMID- 11120630 TI - Alpha-tocopherol succinate inhibits growth of gastric cancer cells in vitro. AB - BACKGROUND: Vitamin E in the form of alpha-tocopherol succinate (ATS) has been shown to inhibit growth of several cancer cell lines in vitro, including pancreas, breast, and prostate. No data exist on the effect of ATS on gastric cancer cell viability. METHODS: A gastric cancer cell line in suspension form, KATO-III, was plated in 96-well plates at 30,000 cells per well with 100 microl RPMI media. The cells were allowed to incubate for 24 h and were then treated with ATS at doses of 25, 50, or 100 microg/ml. The ATS was dissolved in 1% EtOH solution and control cells received an identical solution of EtOH without ATS. Treated cells were incubated for 24, 48, or 72 h. At the completion of the treatment period, MTT assay was performed to determine cell viability. Statistical analysis was performed using Student's t test. RESULTS: All doses of ATS resulted in inhibition of growth of the KATO-III cells. Both 100 and 50 microg/cc doses inhibited growth at all time points (P<0.005), with 48- and 72-h treatments more effective than 24-h treatment. At 24 and 48 h, 100 microg/cc was more effective at inhibition of growth than 50 microg/ml (P<0.005), but by 72 h the effects of the doses were equivalent; 25 microg/ml inhibited cell growth only at 48 and 72 h. At all time points, 50 and 100 microg/ml doses were more effective at inhibiting cell growth than 25 microg/ml. Conclusions. ATS inhibits gastric carcinoma cell growth in vitro in a dose- and time-dependent fashion. In vivo studies are indicated to further evaluate the potential benefit of this antioxidant against gastric cancer. PMID- 11120631 TI - Effects of nicotine and cotinine on porcine arterial endothelial cell function. AB - BACKGROUND. There has been a significant amount of research on the effects of nicotine on vascular biology; however, little is known about the effects of cotinine, the metabolic product of nicotine. This study used a novel vascular perfusion system to study the effects of nicotine and cotinine on the vascular endothelial cell function. METHODS: Porcine common carotid arteries were cultured in a novel vascular perfusion system with nicotine or cotinine or as controls. After 24 h, vessels were precontracted with norepinephrine and subsequently relaxed with acetylcholine. Vessel diameters were recorded and analyzed. After culture, samples were taken for en face, immunohistochemistry, and RT-PCR for eNOS. Porcine coronary arteries were incubated as controls or with nicotine or cotinine and tested on a myograph system to measure contraction and relaxation. RESULTS: Porcine carotid arteries treated with nicotine and cotinine showed a 27.2% and a 41.2% reduction in endothelial-dependent relaxation, respectively, as compared to control vessels (P<0.05). Rings of coronary arteries treated with nicotine relaxed similarly to control rings while cotinine-treated rings failed to relax to endothelial-dependent stimulation. RT-PCR for eNOS mRNA showed a 23. 2 and a 24.1% reduction in eNOS expression for nicotine- and cotinine-treated vessels, respectively (P<0.01). Additionally, immunohistochemical staining for eNOS showed less dense staining on nicotine- and cotinine-treated vessels as compared to controls. En face preparations showed normal endothelial cell morphology in all groups, but cell density decreased slightly in vessels treated with nicotine and cotinine. CONCLUSION: These results indicate that cotinine may have even more effect on the impairment of endothelial-dependent vasorelaxation than nicotine for the regulation of vessel tone in porcine carotid and coronary arteries. PMID- 11120632 TI - Short stay carotid surgery for veterans: an emerging standard. AB - We have taken the short stay approach to carotid artery surgery to our VA setting over the past 5 to 6 years. Retrospectively, we reviewed the efficacy and safety of that approach in 201 consecutive carotid operations over the recent 4-year period (January 1, 1996-December 31, 1999). In 1996 we had already begun the transition to an algorithm to (1) utilize carotid color flow Doppler duplex exams for diagnosis, (2) same-day admission (SDA), (3) intensive care unit (ICU) only when deemed medically necessary, and (4) next-day discharge. Results of this approach have been a decrease in the utilization of diagnostic arteriograms and utilization of the ICU from 100% previous to the onset of this approach to 17 and 22%, respectively. SDA increased from 24 to 89%. Mean LOS decreased from 5.13+/ 0.9 to 1.97+/-0.4 days. The percentage of patients completing the algorithm went from 15 to 72%. Stroke and/or death varied from 0 to 3.7% each year and was only 2.4% over the 4-year period. In conclusion, this approach to short stay carotid surgery in the veteran population has proven both efficacious and safe with results similar to those in university and community practices. PMID- 11120633 TI - Multi-photon microscopy in the evaluation of human saphenous vein. AB - BACKGROUND: The use of conventional fluorescence microscopy to image biological systems at the cellular level is limited by its inability to spatially resolve thick tissues. We have applied the technique of multi-photon fluorescence microscopy to study the structure and function of endothelial cells in living human saphenous vein taken from patients undergoing coronary artery bypass surgery. MATERIALS AND METHODS: Vein segments were preserved for 1-4 h to determine the temporal effects of storage. The effect of pH on endothelial and smooth muscle cell viability was examined by storing segments at pH 6.0, 7.4, and 8.0. Calcein-mediated green fluorescence and ethidium homodimer-mediated red fluorescence were used to differentiate cell viability. Increases in diaminofluorescein fluorescence were used to measure bradykinin activation of endothelial nitric oxide synthase (eNOS) with or without N-nitro-l-arginine (L NNA). Multi-photon imaging was performed with the BioRad MRC1024ES system. RESULTS: Successful imaging of endothelial and smooth muscle cells of vein segments was achieved. Cell viability was well preserved up to 3 h of storage but dramatically decreased after 4 h. Cell viability was maintained at pH 7.4, diminished at pH 8.0, and was completely lost at pH 6.0. A two- to threefold increase in eNOS activity was observed upon activation by bradykinin which was completely inhibited in L-NNA-treated samples. CONCLUSIONS: We have demonstrated the successful application of multi-photon microscopy in imaging and quantifying nitric oxide production and cell viability under various storage conditions in human saphenous veins. This imaging technique allows for the functional imaging of cellular processes and may have diagnostic potential in cardiovascular surgery for patients undergoing bypass operations. PMID- 11120634 TI - Use of rifampin-soaked gelatin-sealed polyester grafts for in situ treatment of primary aortic and vascular prosthetic infections. AB - BACKGROUND: In situ treatment of artery/graft infection has distinct advantages compared to vessel excision and extra-anatomic bypass procedures. Based on animal studies of a rifampin-soaked, gelatin-impregnated polyester graft that demonstrated prolonged in vivo antibacterial activity, this antibiotic-bonded graft was used selectively in patients for in situ treatment of low-grade Gram positive prosthetic graft infections or primary aortic infections not amenable to excision and ex situ bypass. METHODS: In a 5-year period (1995-1999), 27 patients with prosthetic graft infection (aortofemoral, n = 18, femorofemoral, n = 3; axillofemoral, n = 1) or primary aortic infection (mycotic aneurysm, n = 3; infected AAA, n = 2) underwent excision of the infected vessel and in situ replacement with a rifampin soaked (45-60 mg/ml for 15 min) gelatin-impregnated polyester graft. All prosthetic graft infections were low grade in nature, caused Gram-positive bacteria (Staphylococcus epidermidis, 16; Staphylococcus aureus, 5; Streptococcus, 1), and were treated electively. Patients with mycotic aortic aneurysm presented with sepsis and underwent urgent or emergent surgery. RESULTS: Two (8%) patients died-1 as a result of a ruptured Salmonella mycotic aortic aneurysm and the other from methicillin-resistant S. aureus infection following deep vein replacement of an in situ replaced femorofemoral graft. No amputations or late deaths as the result of vascular infection occurred in the 25 surviving patients. Two patients developed recurrent infection caused by a rifampin resistant S. epidermidis in a replaced aortofemoral graft limb and were successfully treated with graft excision and in situ autogenous vein replacement. Eighteen patients remain alive and clinically free of infection after a mean follow-up interval of 17 months. CONCLUSIONS: In situ replacement treatment using a rifampin-bonded prosthetic graft for low-grade staphylococcal arterial infection was safe, durable, and associated with eradication of clinical signs of infection. Failure of this therapy was the result of virulent and antibiotic resistant bacterial strains. PMID- 11120635 TI - Aneurysm rupture is independently associated with increased late mortality in those surviving abdominal aortic aneurysm repair. AB - PURPOSE: The aim of this study was to define whether veterans who survived repair of ruptured abdominal aortic aneurysms (AAA) experienced late survival rates similar to those surviving repair of intact AAA. METHODS: All veterans undergoing AAA repair in DRGs 110 and 111 during fiscal years 1991-1995 were identified using the Veterans Affairs (VA) Patient Treatment File (PTF). Late mortality was defined using VA administrative databases including the Beneficiary Identification and Record Locator System and PTF. Illness severity and patient complexity were defined using PTF discharge data that were further analyzed by Patient Management Category software. Veterans were followed up to 6 years after AAA repair. RESULTS: During the study, 5833 veterans underwent repair of intact AAA while 427 had repair of ruptured AAA in all VA medical centers. Operative mortality was defined as that which occurred within 30 days of surgery or during the same hospitalization as aneurysm repair. For those undergoing repair of intact AAA, operative mortality thus defined was 4.5% (265/5833). Operative mortality was 46% (195/427) after repair of ruptured AAA. Overall mortality (including operative mortality) during 2.62+/-1.61 years follow-up was 22% (1282/5833) with intact AAA versus 61% (260/427) for those with ruptured AAA (P<0.001). Further analysis of survival outcomes was performed in patients who survived AAA repair (i.e., those who were discharged alive and lived 30 days or more after surgery). Of those who initially survived repair of ruptured AAA, 28% (65/232) died during follow-up versus 18% (1017/5568) who initially survived repair of intact AAA (odds ratio 1.74; 95% confidence limits 1.30-2.34; P<0.001). In those initially surviving AAA repair, stepwise logistic regression analysis revealed that increasing age, illness severity, patient complexity, as well as AAA rupture and aortic graft complications were increasingly and independently associated with late mortality. Mean survival time was 1681 days for those who survived >30 days and who were discharged alive after repair of ruptured AAA versus 1821 days for those who initially survived repair of intact AAA (P< 0.001). CONCLUSIONS: In addition to higher postoperative mortality rates with ruptured AAA, mortality during follow-up for survivors of AAA repair was also greater for those who survived repair of ruptured AAA. The toll taken by ruptured abdominal aortic aneurysms did not end in the immediate postoperative period. PMID- 11120636 TI - Abdominal wall repair is delayed during hepatic regeneration. AB - BACKGROUND: Abdominal wall wound failure remains a common surgical problem. The signals that activate normal fibroplastic repair versus regeneration pathways are unknown. Transforming growth factor beta levels rise during incisional healing but fall during hepatic regeneration. Changes in the injured host cytokine milieu may therefore differentially effect abdominal wall repair versus hepatic regeneration. MATERIALS AND METHODS: Forty-eight rats were divided into four groups (n = 12). Groups 1-3 underwent sham celiotomy, 70% hepatectomy, or 80% enterectomy with anastamosis. Incisions from Group 4 were treated with either 1 microg of transforming growth factor beta(2) (TGF-beta(2)) or vehicle following hepatectomy. Isolated fascial and dermal incisions were harvested and tested for breaking strength on POD 7. Serum (TGF-beta(2)) and hepatocyte growth factor (HGF) levels were measured by ELISA. RESULTS: Recovery of incisional wound breaking strength was delayed following hepatectomy but not enterectomy (P<0.002). The inhibitory effect was observed in both the fascia and the dermis of the abdominal wall. TGF-beta(2) levels were depressed in hepatectomy animals on POD 7, while at the same time HGF levels were elevated. Exogenous TGF-beta(2) shifted the healing trajectory of deficient wounds back toward a control pattern. CONCLUSION: Abdominal wall fascial and dermal healing is delayed during hepatic regeneration. Elevated HGF and depressed TGF-beta(2) suggest a host mechanism that prioritizes hepatic parenchymal regeneration over fibroplastic repair (scar). Observations such as these are needed as therapeutic wound healing enters the clinical realm. PMID- 11120637 TI - The determination of cardiac surgical risk using artificial neural networks. AB - BACKGROUND: In the study presented here, an artificial neural network was used to "learn" the relationship between 11 risk factors and patient surgical outcome (survival or death). The network was then used to predict the surgical outcome of other patients. METHODS: Eleven risk factors were presented as inputs to an artificial neural network (ANN). The ANN model was developed by training and testing on 1875 patients. The results of the ANN were compared with the results from two versions of the VA surgical risk model (Denver model). Cutoffs were determined for each model in order to compare their results. RESULTS: The ANN model gave the best results when compared with the new and old Denver models. The ANN model had the lowest overall percentage of error. It predicted living patients with an error of 14% and death with an error of approximately 31.0%. The old Denver model predicted living patients with an error of 15% and deaths with an error of 31%. The new Denver model predicts living patients with an error around 18% and the deaths with an error of 31%. CONCLUSIONS: The combined predictions of the ANN model were slightly more accurate than either the new or the old Denver models. The ANN model was created from 1875 patients in about 1 month, while both of the Denver models were developed over a 3 year time period and used more than 12,000 patients. Additionally, the ANN model is easily modified, allowing instant addition or deletion of parameters to suit the users needs. PMID- 11120638 TI - Shear force regulates matrix metalloproteinase activity in human saphenous vein organ culture. AB - BACKGROUND: Development of vein graft intimal hyperplasia has been related both to shear force and to the activity of matrix metalloproteinases (MMPs). Little data are available regarding the effects of shear on MMP expression and activity. The aim of this study was to examine the relationship among shear force, metalloproteinase activity, and intimal thickening in human saphenous vein segments maintained in organ culture. MATERIALS AND METHODS: Segments of human saphenous vein were cultured under static conditions, or perfused under low-flow and high-flow conditions in a perfusion apparatus for 7 days. Metalloproteinase levels and activities were measured using ELISA and substrate gel zymography, respectively. Intimal thickening was determined by morphometric analysis. Results were compared with control vein tissue, which was not subjected to organ culture, using a one-way ANOVA. RESULTS: A 13% increase in proteolytic activity was noted on substrate gel zymography at 68-72 kDa in high-flow vein tissue. The protein content of MMP-2, MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), and TIMP-2 was increased in high-flow vein tissue by 21%, 126%, more than 100-fold, and 86%, respectively. In culture media bathing the outside of the vein, TIMP-2 was increased in high-flow specimens, while TIMP-1 was inversely related to flow rate. Intimal thickening was directly related to flow rates, and was progressively increased in the low-flow and high-flow groups by 3-fold and 4 fold, respectively. CONCLUSIONS: Metalloproteinase levels in human saphenous vein cultures are related to shear force. MMP levels and activity correlate with the degree of intimal thickening. This model may provide a valuable tool for the analysis of physical forces and their influence on intimal thickening in human saphenous vein. PMID- 11120639 TI - Early and sufficient feeding reduces length of stay and charges in surgical patients. AB - BACKGROUND: The role of perioperative nutrition in surgical patients remains controversial. We performed a Clinical Practice Improvement (CPI) study that, while controlling for severity of illness, explored the relationship between the timing and amount of parenteral or enteral nutrition, with two outcomes: length of stay (LOS) and total charges in patients undergoing open intestinal operations. MATERIALS AND METHODS: A CPI study was conducted at eight hospitals to determine which process steps were associated with shorter LOS and lower charges. Hospital charts were abstracted for over 800 components of detailed patient, process, and outcome measures. Severity of illness was measured multiple times during the stay using the Comprehensive Severity Index, a disease-specific physiologic severity measurement instrument. Data on 1007 patients undergoing intestinal operations, 183 of whom received nutritional support, were then analyzed using multiple regression procedures. Early (within 48 h of surgery) and sufficient (60% of protein and calorie goals) nutrition, patient variables, and a severity of illness measure were included as independent variables and LOS and hospital charges were used as dependent variables. RESULTS: Mean patient age was 58 years. After controlling for severity of illness, patients who received early and sufficient nutrition had significantly shorter LOS (11.9 days) and lower charges ($34,602) than patients who received early (13.3; $36,452), sufficient (14.6, $39,883), or neither early nor sufficient (14.8, $38,578) (P < or = 0.0001 for early and sufficient versus all other groups). CONCLUSIONS: CPI methodology provides a detailed view of the actual relationship between the timing and the amount of nutrition with LOS and hospital charge outcomes. PMID- 11120640 TI - Human 293 cell metabolism in low glutamine-supplied culture: interpretation of metabolic changes through metabolic flux analysis. AB - Metabolic flux analysis is a useful tool to analyze cell metabolism. In this study, we report the use of a metabolic model with 34 fluxes to study the 293 cell, in order to improve its growth capacity in a DMEM/F12 medium. A batch, fed batch with glutamine feeding, fed-batch with essential amino acids, and finally a fed-batch experiment with both essential and nonessential amino acids were compared. The fed-batch with glutamine led to a maximum cell density of 2.4x10(6) cells/ml compared to 1.8x10(6) cells/ml achieved in a batch mode. In this fed batch with glutamine, it was also found that 2.5 mM ammonia was produced compared to the batch which had a final ammonia concentration of 1 mM. Ammonia was found to be growth inhibiting for this cell line at a concentration starting at 1 mM. During the fed-batch with glutamine, the flux analysis shows that a majority of amino acid fluxes and Kreb's cycle fluxes, except for glutamine flux, are decreased. This observation led to the conclusion that the main nutrient used is glutamine and that during the batch there is an overflow in the Kreb's cycle. Thus, a fed-batch with glutamine permits a better utilization of this nutrient. A fed-batch with essential amino acid without glutamine was also assayed in order to reduce ammonia production. The maximum cell density was increased further to 3x10(6) cells/ml and ammonia production was reduced below 1 mM. Flux analysis shows that the cells could adapt to a medium with low glutamine by increasing the amino acid fluxes toward the Kreb's cycle. Adding nonessential amino acids during this feeding strategy did not improve growth further and the nonessential amino acids accumulated in the medium. PMID- 11120641 TI - Co-overexpression of RspAB improves recombinant protein production in Escherichia coli. AB - The Escherichia coli mutant CWML2 was previously reported to exhibit improved physiological characteristics, including recombinant protein production. Here we investigate the molecular basis of this phenotype by comparing the cellular level of three RNA polymerase sigma subunits by immunoblot analysis. While the level of housekeeping sigma(D) was similar in parent and mutant, the levels of the flagella synthesis regulator sigma(F) and the stationary phase regulator sigma(S) were higher in the mutant strain, indicating a different motility and stationary phase phenotype. Evidence for this conclusion was provided by the significantly higher motility of CWML2, compared to its parent. Based on these results, we hypothesized that alterations in ppGpp regulation via a homoserine lactone dependent mechanism may be relevant for the mutant phenotype. Indeed, transcription of the rspAB operon, which was previously described to be involved in the degradation of homoserine lactone, was found to be deregulated in CWML2 in a plasmid-based reporter protein assay. By overexpression of the E. coli rspAB operon, we could partly mimic the mutant phenotype and demonstrate that co overexpression of RspAB is a pertinent metabolic engineering strategy to improve recombinant protein production. PMID- 11120642 TI - Choline import into chloroplasts limits glycine betaine synthesis in tobacco: analysis of plants engineered with a chloroplastic or a cytosolic pathway. AB - The biosynthesis of the osmoprotectant glycine betaine (GlyBet) is a target for metabolic engineering to enhance stress resistance in crops. Certain plants synthesize GlyBet in chloroplasts via a two-step oxidation of choline (Cho). In previous work, a chloroplastic GlyBet synthesis pathway was inserted into tobacco (which lacks GlyBet) by expressing spinach choline monooxygenase (CMO). The transformants had low CMO enzyme activity, and produced little GlyBet (less than or = 70 nmol g(-1) fresh wt). In this study, transformants with up to 100-fold higher CMO activity showed no further increase in GlyBet. In contrast, tobacco expressing a cytosolic GlyBet synthesis pathway accumulated significantly more GlyBet (430 nmol g(-1) fresh wt), suggesting that subcellular localization influences pathway flux. Modeling of the labeling kinetics of Cho metabolites observed when [14C]Cho was supplied to engineered plants demonstrated that Cho import into chloroplasts indeed limits the flux to GlyBet in the chloroplastic pathway. A high-activity Cho transporter in the chloroplast envelope may therefore be an integral part of the GlyBet synthesis pathway in species that accumulate GlyBet naturally, and hence a target for future engineering. PMID- 11120643 TI - Metabolic flux analysis of postburn hepatic hypermetabolism. AB - The hepatic response to severe injury is characterized by a marked upregulation of glucose, fatty acid, and amino acid turnover, which, if persistent, predisposes the patient to progressive organ dysfunction. To study the effect of injury on liver intermediary metabolism, metabolic flux analysis was applied to isolated perfused livers of burned and sham-burned rats. Intracellular fluxes were calculated using metabolite measurements and a stoichiometric balance model. Significant flux increases were found for multiple pathways, including mitochondrial electron transport, the TCA and urea cycles, gluconeogenesis, and pentose phosphate pathway (PPP). The burn-induced increase in gluconeogenesis did not significantly increase glucose output. Instead, glucose-6-phosphate was diverted into the PPP. These changes were paralleled by increases in glucose-6 phosphate dehydrogenase (G6PDH) and glutathione reductase (GR) activities. Given that G6PDH and GR are the most significant NADPH producers and consumers in the liver, respectively, and that GR is responsible for recycling the free radical scavenger glutathione, these data are consistent with the notion that hepatic metabolic changes are in part due to the induction of liver antioxidant defenses. PMID- 11120644 TI - Low-copy plasmids can perform as well as or better than high-copy plasmids for metabolic engineering of bacteria. AB - Multicopy plasmids are often chosen for the expression of recombinant genes in Escherichia coli. The high copy number is generally desired for maximum gene expression; however, the metabolic burden effects that usually result from multiple plasmid copies could prove to be detrimental for maximum productivity in certain metabolic engineering applications. In this study, low-copy mini-F plasmids were compared to high-copy pMB1-based plasmids for production of two metabolites in E. coli: polyphosphate (polyP) and lycopene derived from isopentenyl diphosphate (IPP). The stationary-phase accumulation of polyP on a per cell basis was enhanced approximately 80% when either high- or low-copy plasmids were used, from 120 micromol/g DCW without augmented polyP kinase (PPK) activity to approximately 220 micromol/g DCW. The cell density of the high-copy plasmid-containing culture at stationary phase was approximately 24% lower than the low-copy culture and 30% lower than the control culture. This difference in cell density is likely a metabolic burden effect and resulted in a lower overall product concentration for the high-copy culture (approximately 130 micromol/L culture) relative to the low-copy culture (approximately 160 micromol/L culture). When the gene for DXP (1-deoxy-D-xylulose 5-phosphate) synthase, the first enzyme in the IPP mevalonate-independent biosynthetic pathway, was expressed from the tac promoter on multicopy and low-copy plasmids, lycopene production was enhanced two- to threefold over that found in cells expressing the chromosomal copy only. Cell growth and lycopene production decreased substantially when isopropyl beta-D thiogalactosidase (IPTG) was added to the high-copy plasmid-containing culture, suggesting that overexpression of DXP synthase was a significant metabolic burden. In the low-copy plasmid-containing culture, no differences in cell growth or lycopene production were observed with any IPTG concentrations. When dxs was placed under the control of the arabinose-inducible promoter (P(BAD)) on the low copy plasmid, the amount of lycopene produced was proportional to the arabinose concentration and no significant changes in cell growth resulted. These results suggest that low-copy plasmids may be useful in metabolic engineering applications, particularly when one or more of the substrates used in the recombinant pathway are required for normal cellular metabolism. PMID- 11120645 TI - Directed evolution of toluene dioxygenase from Pseudomonas putida for improved selectivity toward cis-indandiol during indene bioconversion. AB - Toluene dioxygenase (TDO) from Pseudomonas putida F1 converts indene to a mixture of cis-indandiol (racemic), 1-indenol, and 1-indanone. The desired product, cis (1S,2R)-indandiol, is a potential key intermediate in the chemical synthesis of indinavir sulfate (Crixivan), Merck's HIV-1 protease inhibitor for the treatment of AIDS. To reduce the undesirable byproducts 1-indenol and 1-indanone formed during indene bioconversion, the recombinant TDO expressed in Escherichia coli was evolved by directed evolution using the error-prone polymerase chain reaction (epPCR) method. High-throughput fluorometric and spectrophotometric assays were developed for rapid screening of the mutant libraries in a 96-well format. Mutants with reduced 1-indenol by-product formation were identified, and the individual indene bioconversion product profiles of the selected mutants were confirmed by HPLC. Changes in the amino acid sequence of the mutant enzymes were identified by analyzing the nucleotide sequence of the genes. A mutant with the most desirable product profile from each library, defined as the most reduced 1 indenol concentration and with the highest cis-(1S,2R)-indandiol enantiomeric excess, was used to perform each subsequent round of mutagenesis. After three rounds of mutagenesis and screening, mutant 1C4-3G was identified to have a threefold reduction in 1-indenol formation over the wild type (20% vs 60% of total products) and a 40% increase of product (cis-indandiol) yield. PMID- 11120646 TI - Controlled overexpression of BCKD kinase expression: metabolic engineering applied to BCAA metabolism in a mammalian system. AB - A common metabolic complication of human disease is uncontrolled muscle protein breakdown or cachexia, which occurs in patients with chronic diseases such as cancer, AIDS, renal failure, and diabetes. Increased branched-chain amino acid catabolism is implicated as causal and has stimulated the investigation of methods to regulate the metabolism of these amino acids. Here we demonstrate doxycycline-controlled overexpression of a branched-chain alpha-ketoacid dehydrogenase (BCKD) kinase transgene in mammalian cell culture. This kinase functions to inactivate the BCKD complex by phosphorylation, thus preventing the catabolism of these essential, regulatory metabolites. In this study, doxycycline treatment leads to a 10-fold increase in BCKD kinase protein. The transgene generated kinase is rapidly incorporated within mitochondria and functions correctly to inactivate the BCKD complex. The maximum reduction in basal BCKD activity achieved was 94%. Unexpectedly, total BCKD activity was also decreased by kinase overexpression despite no observable change in expression of the BCKD catalytic proteins. These results demonstrate that artificial regulation of branched-chain amino acid metabolism is possible through the controlled overexpression of a single endogenous enzyme and suggest the feasibility of clinical applications. PMID- 11120647 TI - An ESS maximum principle for matrix games. AB - Previous work has demonstrated that for games defined by differential or difference equations with a continuum of strategies, there exists a G-function, related to individual fitness, that must take on a maximum with respect to a virtual variable v whenever v is one of the vectors in the coalition of vectors which make up the evolutionarily stable strategy (ESS). This result, called the ESS maximum principle, is quite useful in determining candidates for an ESS. This principle is reformulated here, so that it may be conveniently applied to matrix games. In particular, we define a matrix game to be one in which fitness is expressed in terms of strategy frequencies and a matrix of expected payoffs. It is shown that the G-function in the matrix game setting must again take on a maximum value at all the strategies which make up the ESS coalition vector. The reformulated maximum principle is applicable to both bilinear and nonlinear matrix games. One advantage in employing this principle to solve the traditional bilinear matrix game is that the same G-function is used to find both pure and mixed strategy solutions by simply specifying an appropriate strategy space. Furthermore we show how the theory may be used to solve matrix games which are not in the usual bilinear form. We examine in detail two nonlinear matrix games: the game between relatives and the sex ratio game. In both of these games an ESS solution is determined. These examples not only illustrate the usefulness of this approach to finding solutions to an expanded class of matrix games, but aids in understanding the nature of the ESS as well. PMID- 11120648 TI - Balance of fitnesses maintaining polymorphisms in two-locus genetic systems. AB - Maintenance of a stable two-locus polymorphism is analyzed statistically by fitting a logistic regression with a quadratic function of genotypic fitnesses to the probability for a fitness set to maintain a polymorphism. The regression is fitted using a data set containing information on stable equilibria maintained by 32,00 randomly generated fitness sets with three recombination values (0. 005, 0.05, 0.5). Fitted logistic regressions discriminate with 88 to 90% accuracy between fitness sets maintaining and not maintaining a stable internal equilibrium, which implies the existence of a fitness structure (balance of fitnesses) maintaining a two-locus polymorphism. Aspects of the balance of fitnesses revealed by logistic regressions are discussed. It is demonstrated that logistic regression also discriminates between types of a stable polymorphism: globally stable polymorphism, several simultaneously stable polymorphisms, and stable equilibria in addition to a polymorphic one, which implies that different balances of fitnesses are responsible for the maintenance of different types of polymorphism. PMID- 11120649 TI - Influence of parasitized adult reproduction on host-parasitoid dynamics: an age structured model. AB - In this work, we develop an age-structured model (based on delay-differential equations) to investigate the dynamics of host-parasitoid systems in which adults are the target of parasitism. The rare previous work dealing with such interactions assumes that hosts are functionally dead as soon as they are attacked. We relax this assumption and show that low reproduction rates of parasitized hosts can promote stability at the expense of cyclic behavior (either long term or generation cycles). Higher reproduction rates make the regulation of the host population by parasitoids impossible. As it is the case in models in which adults are subjected to attacks but do not reproduce, our model generates generation cycles for a larger set of parameter values than in models in which juveniles are attacked. PMID- 11120650 TI - General theory of competitive coexistence in spatially-varying environments. AB - A general model of competitive and apparent competitive interactions in a spatially-variable environment is developed and analyzed to extend findings on coexistence in a temporally-variable environment to the spatial case and to elucidate new principles. In particular, coexistence mechanisms are divided into variation-dependent and variation-independent mechanisms with variation-dependent mechanisms including spatial generalizations of relative nonlinearity and the storage effect. Although directly analogous to the corresponding temporal mechanisms, these spatial mechanisms involve different life history traits which suggest that the spatial storage effect should arise more commonly than the temporal storage effect and spatial relative nonlinearity should arise less commonly than temporal relative nonlinearity. Additional mechanisms occur in the spatial case due to spatial covariance between the finite rate of increase of a local population and its local abundance, which has no clear temporal analogue. A limited analysis of these additional mechanisms shows that they have similar properties to the storage effect and relative nonlinearity and potentially may be considered as enlargements of the earlier mechanisms. The rate of increase of a species perturbed to low density is used to quantify coexistence. A general quadratic approximation, which is exact in some important cases, divides this rate of increase into contributions from the various mechanisms above and admits no other mechanisms, suggesting that opportunities for coexistence in a spatially variable environment are fully characterized by these mechanisms within this general model. Three spatially-implicit models are analyzed as illustrations of the general findings and of techniques using small variance approximations. The contributions to coexistence of the various mechanisms are expressed in terms of simple interpretable formulae. These spatially-implicit models include a model of an annual plant community, a spatial multispecies version of the lottery model, and a multispecies model of an insect community competing for spatially-patchy and ephemeral food. PMID- 11120651 TI - Spatial coupling in cyclic population dynamics: models and data. AB - We use a dynamic random field to model a spatial collection of coupled oscillators with discrete time stochastic dynamics. At each time step the phase of each cyclic local population is subject to random noise, incremented by a common dynamic, and pulled by a coupling force in the direction of some collective mean phase. We define asynchrony and derive expressions for its measurement in this model. We describe robust methods for phase estimation of cyclic population time series, for estimating strength of coupling between local populations, and for measuring variance of locally acting noise from field data. Proposed methods allow intermittently acting phase synchronizing events operating over large spatial scales to be distinguished from more continuous and possibly locally acting coupling, both of which could result in elevated levels of phase synchronization. We demonstrate the utility of this approach by applying it to classical spatial time series data of Canadian lynx. Analysis confirms findings of previous studies and reveals evidence to suggest that interpopulation coupling was weaker over the 20th century than for the 1800s. Analysis supports the notion that synchrony in these populations is maintained by a continuous and locally acting coupling between adjacent regions with large phase adjustments occurring only infrequently. When this coupling is absent, asynchrony develops between populations. PMID- 11120652 TI - Carl Dusing (1884) on the regulation of the sex-ratio. AB - In three publications in 1883 and 1884 Carl Dusing of Jena gave a mathematical account of the influence of natural selection on the sex ratio based on the same argument that Darwin had advanced in The Descent of Man (1871). The argument thus became quite well known, being included in the principal books on the subject around the turn of the century, as well as in the Encyclopaedia Britannica, references to Dusing being given. By 1930, when Fisher gave a verbal account of the argument in The Genetical Theory of Natural Selection, he saw no need to give references, and no other book of the period treated the subject, as a result of which Dusing's contribution became lost to view. We here give the important paragraphs of Dusing's mathematical account, translated into English. PMID- 11120653 TI - Quantitative indexes of beta-cell function during graded up&down glucose infusion from C-peptide minimal models. AB - Availability of quantitative indexes of insulin secretion is important for definition of the alterations in beta-cell responsivity to glucose associated with different physiopathological states. This is presently possible by using the intravenous glucose tolerance test (IVGTT) in conjunction with the C-peptide minimal model. However, the secretory response to a more physiological slowly increasing/decreasing glucose stimulus may uncover novel features of beta-cell function. Therefore, plasma C-peptide and glucose data from a graded glucose infusion protocol (seven 40-min periods of 0, 4, 8, 16, 8, 4, and 0 mg. kg(-1). min(-1)) in eight normal subjects were analyzed by use of a new model of insulin secretion and kinetics. The model assumes a two-compartment description of C peptide kinetics and describes the stimulatory effect on insulin secretion of both glucose concentration and the rate at which glucose increases. It provides in each individual the insulin secretion profile and three indexes of pancreatic sensitivity to glucose: Phi(s), Phi(d), and Phi(b), related, respectively, to the control of insulin secretion by the glucose level (static control), the rate at which glucose increases (dynamic control), and basal glucose. Indexes (means +/- SE) were Phi(s) = 18.8 +/- 1.8 (10(9) min(-1)), Phi(d) = 222 +/- 30 (10(9)), and Phi(b) = 5.2 +/- 0.4 (10(9) min(-1)). The model also allows one to quantify the beta-cell times of response to increasing and decreasing glucose stimulus, equal to 5.7 +/- 2.2 (min) and 17.8 +/- 2.0 (min), respectively. In conclusion, the graded glucose infusion protocol, interpreted with a minimal model of C-peptide secretion and kinetics, provides a quantitative assessment of pancreatic function in an individual. Its application to various physiopathological states should provide novel insights into the role of insulin secretion in the development of glucose intolerance. PMID- 11120654 TI - Melatonin secretion occurs at a constant rate in both young and older men and women. AB - The magnitude and duration of melatonin (MLT) secretion were measured over a period of 25 h with pharmacokinetic studies employing administration of D(7) MLT at midday and at midnight in two separate studies and two groups of subjects, 12 young and 11 older men and women. Plasma levels of endogenous MLT and D(7) MLT were quantified separately by use of a specific and sensitive method (gas chromatography-mass spectrometry) previously developed in our laboratory, enabling us to measure endogenous and exogenous MLT levels down to 0.5 pg/ml in plasma. In the two groups of subjects, MLT secretion occurred only at night: onset time of secretion was from 1915 to 2205 (Greenwich mean time), and offset was from 0305 to 0545. No MLT peak was observed in individual nocturnal MLT profiles that were similar to curves obtained for a rate-constant infusion. Modelization demonstrated the superimposition of observed data and simulated curves. MLT concentrations decreasing from the offset of secretion might correspond to the elimination of MLT present in the body at the end of nocturnal secretion. By use of the MLT clearance given by pharmacokinetics, the amount of secreted MLT was found to be 35.7 and 21.6 microg for men and women, respectively, and the rate of secretion was 4.6 and 2.8 microg/h, respectively. No significant gender difference was observed for these two parameters when normalized to body weight. No significant gender difference was observed for onset times of secretion or duration of secretion (7.6-8.6 h) within the two groups, or between young and older subjects. PMID- 11120655 TI - Gluconeogenesis in moderately and severely hyperglycemic patients with type 2 diabetes mellitus. AB - We tested the generally accepted concept that increased gluconeogenesis (GNG) and endogenous glucose production (EGP) are the main reasons for postabsorptive hyperglycemia in patients with type 2 diabetes mellitus (T2DM). GNG was measured with the (2)H(2)O method by use of both the C5-to-C2 ratio (C5/C2, with gas chromatography-mass spectrometry) and the C5-to-(2)H(2)O ratio (C5/(2)H(2)O, with isotope ratio mass spectrometry), and EGP was measured with 3-[(3)H]glucose in 27 patients with T2DM [13 with fasting plasma glucose (FPG) >10 mM and 14 with FPG <10 mM] and in 7 weight- and age-matched nondiabetic controls. The results showed 1) that GNG could be determined accurately with (2)H(2)O by using either C5/C2 or C5/(2)H(2)O; 2) that whereas after an overnight fast of 16 h, GNG was higher in the entire group of patients with T2DM than in controls (6.4 vs. 5.0 micromol. kg(-1). min(-1) or 60.4 vs. 51.4% of EGP, P < 0.02), GNG was within normal limits (less than the mean +/- 2 SD of controls or <65.3%) in 11/14 (79%) patients with mild to moderate hyperglycemia (FPG <10 mM) and in 5/13 (38%) of patients with severe hyperglycemia (FPG 10-20 mM); 3) that elevated GNG in T2DM was associated with a 43% decrease in prehepatic insulin secretion, i.e., with hepatic insulin deficiency; and 4) that FPG correlated significantly with glucose clearance (insulin resistance) (r = 0.70) and with GNG (r = 0.50) or EGP (r = 0.45). We conclude 1) that peripheral insulin resistance is at least as important as GNG (and EGP) as a cause of postabsorptive hyperglycemia in T2DM and 2) that GNG and EGP in T2DM are increased under conditions of significant hepatic insulin deficiency and thus probably represent a late event in the course of T2DM. PMID- 11120656 TI - An in vivo study of ovine placental transport of essential amino acids. AB - Under normal physiological conditions, essential amino acids (EA) are transported from mother to fetus at different rates. The mechanisms underlying these differences include the expression of several amino acid transport systems in the placenta and the regulation of EA concentrations in maternal and fetal plasma. To study the relation of EA transplacental flux to maternal plasma concentration, isotopes of EA were injected into the circulation of pregnant ewes. Measurements of concentration and molar enrichment in maternal and fetal plasma and of umbilical plasma flow were used to calculate the ratio of transplacental pulse flux to maternal concentration (clearance) for each EA. Five EA (Met, Phe, Leu, Ile, and Val) had relatively high and similar clearances and were followed, in order of decreasing clearance, by Trp, Thr, His, and Lys. The five high-clearance EA showed strong correlation (r(2) = 0.98) between the pulse flux and maternal concentration. The study suggests that five of the nine EA have similar affinity for a rate-limiting placental transport system that mediates rapid flux from mother to fetus, and that differences in transport rates within this group of EA are determined primarily by differences in maternal plasma concentration. PMID- 11120657 TI - Impaired insulin action in subcutaneous adipocytes from women with visceral obesity. AB - Visceral obesity is associated with resistance to the antilipolytic effect of insulin in vivo. We investigated whether subcutaneous abdominal and gluteal adipocytes from viscerally obese women exhibit insulin resistance in vitro. Subjects were obese black and white premenopausal nondiabetic women matched for visceral adipose tissue (VAT), total adiposity, and age. Independently of race and adipocyte size, increased VAT was associated with decreased sensitivity to insulin's antilipolytic effect in subcutaneous abdominal and gluteal adipocytes. Absolute lipolytic rates at physiologically relevant concentrations of insulin or the adenosine receptor agonist N(6)-(phenylisopropyl)adenosine were higher in subjects with the highest vs. lowest VAT area. Independently of cell size, abdominal adipocytes were less sensitive to the antilipolytic effect of insulin than gluteal adipocytes, which may partly explain increased nonesterified fatty acid fluxes in upper vs. lower body obese women. Moreover, increased VAT was associated with decreased responsiveness, but not decreased sensitivity, to insulin's stimulatory effect on glucose transport in abdominal adipocytes. These data suggest that insulin resistance of subcutaneous abdominal and, to a lesser extent, gluteal adipocytes may contribute to increased systemic lipolysis in both black and white viscerally obese women. PMID- 11120658 TI - Reference intervals for glucose, beta-cell polypeptides, and counterregulatory factors during prolonged fasting. AB - To establish reference intervals for the pancreatic beta-cell response and the counterregulatory hormone response to prolonged fasting, we studied 33 healthy subjects (16 males, 17 females) during a 72-h fast. Glucose, insulin, C-peptide, and proinsulin levels decreased (P < 0.001), and the levels of counterregulatory factors increased during the fast [P < 0.05; glucagon and free fatty acids (FFA) with a linear increase and epinephrine, norepinephrine, and cortisol with a clear underlying circadian rhythm]. Growth hormone secretion increased from the first to third day of fasting (P < 0.05) but actually decreased from the second to third day of fasting (P = 0.03). Males had higher glucose and glucagon levels and lower FFA levels during the fast (P < 0.05), whereas no effect of gender on beta cell polypeptides was observed. A high body mass index resulted in higher insulin and C-peptide levels during the fast (P < 0.05). In conclusion, we have provided reference intervals for glucoregulatory factors during a 72-h fast. We observed a diminished beta-cell response concomitant with an increased secretion of counterregulatory hormones. These results should be of clinical and scientific value in the investigation of hypoglycemic disorders. PMID- 11120659 TI - Lack of effect of incretin hormones on insulin release from pancreatic islets in the bile duct-ligated rats. AB - Hyperglycemia associated with obstructive jaundice seriously affects the prognosis of patients with hepatobiliary diseases. We investigated secretory properties of isolated islets from bile duct-ligated (BDL) rats. Pancreatic islets from BDL rats lost their secretory responses to glucagon-like peptide-1 (GLP-1), although their responses to glucose were normal. Loss of potentiation of insulin release was also observed in glucagon and glucose-dependent insulinotropic peptide (GIP), whereas modulation of the release by forskolin, dibutyryl cAMP, or epinephrine remained unaffected. cAMP production by BDL islets was not increased by these insulinotropic hormones. Serum levels of glucagon, but not GIP, were increased in BDL rats. GLP-1 levels were also elevated, although they did not reach statistical significance. Immunoblotting of trimeric G protein subunits demonstrated that G(s)alpha L and G(s)alpha S, but not G(i)alpha 1/2 and G(i)alpha 3/o alpha, were less expressed in BDL islets. Therefore, unresponsiveness of the beta-cell to cAMP-raising hormones is involved in glucose intolerance under cholestasis. It results from diminished expression of alpha subunits of the relevant G protein, G(s), and desensitization of receptors of these hormones. PMID- 11120660 TI - Diversification of cardiac insulin signaling involves the p85 alpha/beta subunits of phosphatidylinositol 3-kinase. AB - Ventricular cardiomyocytes and cardiac tissue of lean and genetically obese (fa/fa) Zucker rats were used 1) to study the role of the p85 regulatory subunit isoforms p85 alpha and p85 beta for insulin signaling through the phosphatidylinositol (PI) 3-kinase pathway, and 2) to elucidate the implications of these mechanisms for cardiac insulin resistance. Western blot analysis of cardiomyocyte lysates revealed expression of p85 alpha and p85 beta but no detectable amounts of the splice variants of p85 alpha. Essentially no p85 alpha subunit of PI 3-kinase was found to be associated with insulin receptor substrate (IRS)-1 or IRS-2 in basal and insulin-stimulated (5 min) cardiomyocytes. Instead, insulin produced a twofold increase in p85 beta associated with IRS-1, leading to a three- to fourfold increase in p85 beta-associated PI 3-kinase activity. This response was significantly reduced in obese animals. Comparable results were obtained in the intact heart after in vivo stimulation. In GLUT-4-containing vesicles, an increased abundance (3.7 +/- 0.7-fold over basal) of p85 alpha was observed after insulin stimulation of lean animals, with no significant effect in the obese group. No p85 beta could be detected in GLUT-4-containing vesicles. Recruitment of the p110 catalytic subunit of PI 3-kinase and a twofold increase in enzyme activity in GLUT-4-containing vesicles by insulin was observed only in lean rats. We conclude that, in the heart, p85 alpha recruits PI 3-kinase activity to GLUT-4 vesicles, whereas p85 beta represents the main regulator of IRS-1- and IRS-2-mediated PI 3-kinase activation. Furthermore, multiple defects of PI 3-kinase activation, involving both the p85 alpha and the p85 beta adaptor subunits, may contribute to cardiac insulin resistance. PMID- 11120661 TI - Regulatory role of arginase I and II in nitric oxide, polyamine, and proline syntheses in endothelial cells. AB - Endothelial cells (EC) metabolize L-arginine mainly by arginase, which exists as two distinct isoforms, arginase I and II. To understand the roles of arginase isoforms in EC arginine metabolism, bovine coronary venular EC were stably transfected with the Escherichia coli lacZ gene (lacZ-EC, control), rat arginase I cDNA (AI-EC), or mouse arginase II cDNA (AII-EC). Western blots and enzymatic assays confirmed high-level expression of arginase I in the cytosol of AI-EC and of arginase II in mitochondria of AII-EC. For determining arginine catabolism, EC were cultured for 24 h in DMEM containing 0.4 mM L-arginine plus [1 (14)C]arginine. Urea formation, which accounted for nearly all arginine consumption by these cells, was enhanced by 616 and 157% in AI-EC and AII-EC, respectively, compared with lacZ-EC. Arginine uptake was 31-33% greater in AI-EC and AII-EC than in lacZ-EC. Intracellular arginine content was 25 and 11% lower in AI-EC and AII-EC, respectively, compared with lacZ-EC. Basal nitric oxide (NO) production was reduced by 60% in AI-EC and by 47% in AII-EC. Glutamate and proline production from arginine increased by 164 and 928% in AI-EC and by 79 and 295% in AII-EC, respectively, compared with lacZ-EC. Intracellular content of putrescine and spermidine was increased by 275 and 53% in AI-EC and by 158 and 43% in AII-EC, respectively, compared with lacZ-EC. Our results indicate that arginase expression can modulate NO synthesis in bovine venular EC and that basal levels of arginase I and II are limiting for endothelial syntheses of polyamines, proline, and glutamate and may have important implications for wound healing, angiogenesis, and cardiovascular function. PMID- 11120662 TI - Effect of streptozotocin-induced diabetes on glycogen resynthesis in fasted rats post-high-intensity exercise. AB - It has recently been shown that food intake is not essential for the resynthesis of the stores of muscle glycogen in fasted animals recovering from high-intensity exercise. Because the effect of diabetes on this process has never been examined before, we undertook to explore this issue. To this end, groups of rats were treated with streptozotocin (60 mg/kg body mass ip) to induce mild diabetes. After 11 days, each animal was fasted for 24 h before swimming with a lead weight equivalent to 9% body mass attached to the tail. After exercise, the rate and the extent of glycogen repletion in muscles were not affected by diabetes, irrespective of muscle fiber composition. Consistent with these findings, the effect of exercise on the phosphorylation state of glycogen synthase in muscles was only minimally affected by diabetes. In contrast to its effects on nondiabetic animals, exercise in fasted diabetic rats was accompanied by a marked fall in hepatic glycogen levels, which, surprisingly, increased to preexercise levels during recovery despite the absence of food intake. PMID- 11120663 TI - Altered TNF-alpha, glucose, insulin, and amino acids in islets of Langerhans cultured in a microgravity model system. AB - The present studies were designed to determine effects of a microgravity model system upon lipopolysaccharide (LPS)-stimulated tumor necrosis factor-alpha (TNF alpha) activity and indexes of insulin and fuel homeostasis of pancreatic islets of Langerhans. Islets (1,726 +/- 117, 150 islet equivalent units) from Wistar Furth rats were treated as 1) high aspect ratio vessel (HARV) cell culture, 2) HARV plus LPS, 3) static culture, and 4) static culture plus LPS. TNF-alpha (L929 cytotoxicity assay) was significantly increased in LPS-induced HARV and static cultures; yet the increase was more pronounced in the static culture group (P < 0.05). A decrease in insulin concentration was demonstrated in the LPS-stimulated HARV culture (P < 0.05). We observed a greater glucose concentration and increased disappearance of arginine in islets cultured in HARVs. Although nitrogenous compound analysis indicated a ubiquitous reliance on glutamine in all experimental groups, arginine was converted to ornithine at a twofold greater rate in the islets cultured in the HARV microgravity model system (P < 0.05). These studies demonstrate alterations in LPS-induced TNF-alpha production of pancreatic islets of Langerhans, favoring a lesser TNF activity in the HARV. These alterations in fuel homeostasis may be promulgated by gravity-averaged cell culture methods or by three-dimensional cell assembly. PMID- 11120665 TI - pH dependence of bone resorption: mouse calvarial osteoclasts are activated by acidosis. AB - We examined the effects of HCO(3)(-) and CO(2) acidosis on osteoclast-mediated Ca(2+) release from 3-day cultures of neonatal mouse calvaria. Ca(2+) release was minimal above pH 7.2 in control cultures but was stimulated strongly by the addition of small amounts of H(+) to culture medium (HCO(3)(-) acidosis). For example, addition of 4 meq/l H(+) reduced pH from 7.12 to 7.03 and increased Ca(2+) release 3.8-fold. The largest stimulatory effects (8- to 11-fold), observed with 15-16 meq/l added H(+), were comparable to the maximal Ca(2+) release elicited by 1,25-dihydroxyvitamin D(3) [1, 25(OH)(2)D(3); 10 nM], parathyroid hormone (10 nM), or prostaglandin E(2) (1 microM); the action of these osteolytic agents was attenuated strongly when ambient pH was increased from approximately 7.1 to approximately 7.3. CO(2) acidosis was a less effective stimulator of Ca(2+) release than HCO(3)(-) acidosis over a similar pH range. Ca(2+) release stimulated by HCO(3)(-) acidosis was almost completely blocked by salmon calcitonin (20 ng/ml), implying osteoclast involvement. In whole mount preparations of control half-calvaria, approximately 400 inactive osteoclast-like multinucleate cells were present; in calvaria exposed to HCO(3)(-) acidosis and to the other osteolytic agents studied, extensive osteoclastic resorption, with perforation of bones, was visible. HCO(3)(-) acidosis, however, reduced numbers of osteoclast-like cells by approximately 50%, whereas 1,25(OH)(2)D(3) treatment caused increases of approximately 75%. The results suggest that HCO(3)(-) acidosis stimulates resorption by activating mature osteoclasts already present in calvarial bones, rather than by inducing formation of new osteoclasts, and provide further support for the critical role of acid-base balance in controlling osteoclast function. PMID- 11120664 TI - Increasing membrane-bound MCSF does not enhance OPGL-driven osteoclastogenesis from marrow cells. AB - Macrophage colony-stimulating factor (MCSF) and osteoprotegerin ligand (OPGL), both produced by osteoblasts/stromal cells, are essential factors for osteoclastogenesis. Whether local MCSF levels regulate the amount of osteoclast formation is unclear. Two culture systems, ST-2 and Chinese hamster ovary membrane-bound MCSF (CHO-mMCSF)-Tet-OFF cells, were used to study the role of mMCSF in osteoclast formation. Cells from bone marrow (BMM) or spleen were cultured with soluble OPGL on glutaraldehyde-fixed cell layers; osteoclasts formed after 7 days. Osteoclast number was proportional to the amount of soluble OPGL added. In contrast, varying mMCSF levels in the ST-2 or CHO-mMCSF-Tet-OFF cell layers, respectively by variable plating or by addition of doxycycline, did not affect BMM osteoclastogenesis: 20-450 U of mMCSF per well generated similar osteoclast numbers. In contrast, spleen cells were resistant to mMCSF: osteoclastogenesis required > or = 250 U per well and further increased as mMCSF rose higher. Our results demonstrate that osteoclast formation in the local bone environment is dominated by OPGL. Increasing mMCSF above basal levels does not further enhance osteoclast formation from BMMs, indicating that mMCSF does not play a dominant regulatory role in the bone marrow. PMID- 11120666 TI - Effects of hyperprolactinemia on rat prostate growth: evidence of androgeno dependence. AB - The effects of the polypeptide hormone prolactin (PRL) in the development and regulation of benign prostate hyperplasia (BPH) and also in prostate cancer are not very well characterized. This study examines the action of PRL, either alone or in association with androgens [testosterone (T) or dihydrotestosterone (DHT)], in the rat prostate gland. The effects of PRL and androgens were investigated after 30 and 60 days in control, castrated, castrated with a substitutive implant of T or DHT, and sham-operated Wistar rats. To enhance PRL release, we induced hyperprolactinemia by administering chronic injections of sulpiride (40 mg. kg( 1). day(-1)). Chronic hyperprolactinemia induces enlargement and inflammation of the lateral rat prostate without any histological changes on ventral and dorsal lobes. We also demonstrate that hyperprolactinemia induces Bcl-2 overexpression in the lateral rat prostate and that this could inhibit the level of apoptosis. The in vivo model established here is a useful in vivo approach for studying the hormonal regulation of normal and pathological prostate development. PMID- 11120667 TI - Leptin administration improves skeletal muscle insulin responsiveness in diet induced insulin-resistant rats. AB - In addition to suppressing appetite, leptin may also modulate insulin secretion and action. Leptin was administered here to insulin-resistant rats to determine its effects on secretagogue-stimulated insulin release, whole body glucose disposal, and insulin-stimulated skeletal muscle glucose uptake and transport. Male Wistar rats were fed either a normal (Con) or a high-fat (HF) diet for 3 or 6 mo. HF rats were then treated with either vehicle (HF), leptin (HF-Lep, 10 mg. kg(-1). day(-1) sc), or food restriction (HF-FR) for 12-15 days. Glucose tolerance and skeletal muscle glucose uptake and transport were significantly impaired in HF compared with Con. Whole body glucose tolerance and rates of insulin-stimulated skeletal muscle glucose uptake and transport in HF-Lep were similar to those of Con and greater than those of HF and HF-FR. The insulin secretory response to either glucose or tolbutamide (a pancreatic beta-cell secretagogue) was not significantly diminished in HF-Lep. Total and plasma membrane skeletal muscle GLUT-4 protein concentrations were similar in Con and HF Lep and greater than those in HF and HF-FR. The findings suggest that chronic leptin administration reversed a high-fat diet-induced insulin-resistant state, without compromising insulin secretion. PMID- 11120668 TI - Muscle fatty acid oxidative capacity is a determinant of whole body fat oxidation in elderly people. AB - In sedentary elderly people, a reduced muscle fatty acid oxidative capacity (MFOC) may explain a decrease in whole body fat oxidation. Eleven sedentary and seven regularly exercising subjects (65.6 +/- 4. 5 yr) were characterized for their aerobic fitness [maximal O(2) uptake (VO(2 max))/kg fat free mass (FFM)] and their habitual daily physical activity level [free-living daily energy expenditure divided by sleeping metabolic rate (DEE(FLC)/SMR)]. MFOC was determined by incubating homogenates of vastus lateralis muscle with [1 (14)C]palmitate. Whole body fat oxidation was measured by indirect calorimetry over 24 h. MFOC was 40.4 +/- 14.7 and 44.3 +/- 16.3 nmol palmitate. g wet tissue( 1). min(-1) in the sedentary and regularly exercising individuals, respectively (P = nonsignificant). MFOC was positively correlated with DEE(FLC)/SMR (r = 0.58, P < 0. 05) but not with VO(2 max)/kg FFM (r = 0.35, P = nonsignificant). MFOC was the main determinant of fat oxidation during all time periods including physical activity. Indeed, MFOC explained 19.7 and 30.5% of the variance in fat oxidation during walking and during the alert period, respectively (P < 0.05). Furthermore, MFOC explained 23.0% of the variance in fat oxidation over 24 h (P < 0.05). It was concluded that, in elderly people, MFOC may be influenced more by overall daily physical activity than by regular exercising. MFOC is a major determinant of whole body fat oxidation during physical activities and, consequently, over 24 h. PMID- 11120669 TI - ANG II is required for optimal overload-induced skeletal muscle hypertrophy. AB - ANG II mediates the hypertrophic response of overloaded cardiac muscle, likely via the ANG II type 1 (AT(1)) receptor. To examine the potential role of ANG II in overload-induced skeletal muscle hypertrophy, plantaris and/or soleus muscle overload was produced in female Sprague-Dawley rats (225-250 g) by the bilateral surgical ablation of either the synergistic gastrocnemius muscle (experiment 1) or both the gastrocnemius and plantaris muscles (experiment 2). In experiment 1 (n = 10/group), inhibiting endogenous ANG II production by oral administration of an angiotensin-converting enzyme (ACE) inhibitor during a 28-day overloading protocol attenuated plantaris and soleus muscle hypertrophy by 57 and 96%, respectively (as measured by total muscle protein content). ACE inhibition had no effect on nonoverloaded (sham-operated) muscles. With the use of new animals (experiment 2; n = 8/group), locally perfusing overloaded soleus muscles with exogenous ANG II (via osmotic pump) rescued the lost hypertrophic response in ACE inhibited animals by 71%. Furthermore, orally administering an AT(1) receptor antagonist instead of an ACE inhibitor produced a 48% attenuation of overload induced hypertrophy that could not be rescued by ANG II perfusion. Thus ANG II may be necessary for optimal overload-induced skeletal muscle hypertrophy, acting at least in part via an AT(1) receptor-dependent pathway. PMID- 11120670 TI - Regulation of prohormone convertase 1 (PC1) by thyroid hormone. AB - The prohormone convertases (PCs) PC1 and PC2 are key enzymes capable of processing a variety of prohormones to their bioactive forms. In this study, we demonstrated that 6-n-propyl-2-thiouracil (PTU)-induced hypothyroidism stimulated, whereas triido-L-thyronine (T(3))-induced hyperthyroidism suppressed, PC1 mRNA levels in the rat anterior pituitary. Using 5' deletions of the human PC1 (hPC1) promoter transiently transfected into GH3 (a somatotroph cell line) cells, we found that T(3) negatively regulated hPC1 promoter activity and that this regulation required the region from -82 to +19 bp relative to the transcription start site. Electrophoretic mobility shift assays (EMSAs) using purified thyroid hormone receptor-alpha1 (TR alpha 1) and retinoid X receptor beta (RXRbeta) proteins and GH3 nuclear extracts demonstrated that the region from -10 to +19 bp of the hPC1 promoter bound TR alpha 1 as both a monomer and a homodimer and bound TR alpha 1/RXR beta as a heterodimer and multimer. EMSAs with oligonucleotides containing point mutations of the putative negative thyroid response elements (TREs) exhibited diminished homodimer and loss of multimer binding. We conclude that there are multiple novel TRE-like sequences in the hPC1 promoter located from -10 to +19 bp. PMID- 11120671 TI - Impairment of endothelial nitric oxide production by acute glucose overload. AB - We examined the effects of acute glucose overload (pretreatment for 3 h with 23 mM D-glucose) on the cellular productivity of nitric oxide (NO) in bovine aortic endothelial cells (BAEC). We had previously reported (Kimura C, Oike M, and Ito Y. Circ Res, 82: 677-685, 1998) that glucose overload impairs Ca(2+) mobilization due to an accumulation of superoxide anions (O(2)(-)) in BAEC. In control cells, ATP induced an increase in NO production, assessed by diaminofluorescein 2 (DAF 2), an NO-sensitive fluorescent dye, mainly due to Ca(2+) entry. In contrast, ATP induced increase in DAF-2 fluorescence was impaired by glucose overload, which was restored by superoxide dismutase, but not by catalase or deferoxamine. Furthermore, pyrogallol, an O(2)(-) donor, also attenuated ATP-induced increase in DAF-2 fluorescence. In contrast, a nonspecific intracellular Ca(2+) concentration increase induced by the Ca(2+) ionophore A-23187, which depletes the intracellular store sites, elevated DAF-2 fluorescence in both control and high D-glucose-treated cells in Ca(2+)-free solution. These results indicate that glucose overload impairs NO production by the O(2)(-)-mediated attenuation of Ca(2+) entry. PMID- 11120672 TI - The iterative two-stage population approach to IVGTT minimal modeling: improved precision with reduced sampling. Intravenous glucose tolerance test. AB - The minimal model method is widely used to estimate glucose effectiveness (S(G)) and insulin sensitivity (S(I)) from intravenous glucose tolerance test (IVGTT) data. In the standard protocol (sIVGTT, 0.33 g/kg glucose bolus given at time 0), which allows the simultaneous assessment of beta-cell function, the precision of the individualized estimates often degrades and particularly so in the presence of reduced sampling schedules. Here, we investigated the use of a population approach, the iterative two-stage (ITS) approach, to analyze 16 sIVGTTs in healthy subjects and to obtain refined estimates of S(G) and S(I) in the population and in the individual subjects. The ITS is based on calculation of the population mean and standard deviation of the parameters at each iteration and then use of them as prior information for the individual analyses. Theoretically, the use of a prior in the ITS should improve the precision of the individual estimates. The customary approach (standard two stage, STS), where modeling is performed separately for each individual subject, does not take the population knowledge into account. We used both frequent (FSS, 30 samples) and (quasi optimally) reduced (RSS, 14 samples) sampling schedules. For the FSS, STS gave estimates (mean +/- SD) for S(G) = 2.66 +/- 1.09 x 10(-2). min(-1) and S(I) = 6.46 +/- 6.99 10(-4). min(-1). microU(-1). ml, with an average precision of 51 (range 5-176) and 33% (3-91), respectively. RSS radically worsened the precision of both S(G) and S(I). However, RSS and ITS gave S(G) = 2.59 +/- 0.73 and S(I) = 6.06 +/- 7.28, with an average precision of 23 (12-42) and 27% (), respectively. In conclusion, population minimal modeling of sIVGTT data improves the precision of individual estimates of glucose effectiveness and insulin sensitivity, as the theory predicts, and, even with reduced sampling, the improvement is substantial. PMID- 11120673 TI - Determination of muscle-specific glucose flux using radioactive stereoisomers and microdialysis. AB - The purpose of the present study was to evaluate a novel approach for determining skeletal muscle-specific glucose flux using radioactive stereoisomers and the microdialysis technique. Microdialysis probes were inserted into the vastus lateralis muscle of human subjects and perfused (4 microl/min) with a Ringer solution containing small amounts of radioactive D- and L-glucose as the internal reference markers for determining probe recovery as well as varying concentrations of insulin (0-10 microM). The rationale behind this approach was that both stereoisomers would be equally affected by the factors that determine probe recovery, with the exception of L-glucose, which is nonmetabolizable and would not be influenced by tissue uptake. Therefore, any differences in the probe recovery ratios between the D- and L-stereoisomers represent changes in skeletal muscle glucose uptake directly at the tissue level. There were no differences in probe recovery between the D- (42.3 +/- 3.5%) and L- (41.2 +/- 3.5) stereoisomers during the control period (no insulin), which resulted in a D/L ratio of 1.04 +/- 0.03. However, during insulin perfusion (1 microM), The D/L ratio increased to 1.62 +/- 0.08 and 1.58 +/- 0.07 (P < 0.05) during the two collection (0-15 and 15 30 min) periods, respectively. This was accomplished solely by an increase (P < 0.05) in D-glucose probe recovery, as L-glucose probe recovery remained unchanged. In a second set of experiments, the perfusion of 10 microM insulin did not increase the D/L ratio (1.40 +/- 0.11) above that observed during 1.0 microM (1.41 +/- 0.07) insulin perfusion. These data suggest that this method is sufficiently sensitive to detect differences in insulin-stimulated glucose uptake; thus the use of radioactive stereoisomers in conjunction with the microdialysis technique provides a novel and useful technique for determining tissue-specific glucose flux and insulin sensitivity. PMID- 11120675 TI - Bioinformatics in Australia. PMID- 11120674 TI - Increased density of glucagon receptors in liver from endurance-trained rats. AB - The binding properties of glucagon receptors were determined in plasma membranes isolated from liver of untrained (n = 6) and swimming endurance-trained Sprague Dawley male rats (n = 7; 3 h/day, 5 days/wk, for 8 wk). Plasma membranes were purified from liver by aqueous two-phase affinity partitioning, and saturation kinetics were obtained by incubation of plasma membranes (10 microg of proteins/150 microl) with (125)I-labeled glucagon at concentrations ranging from 0.15 to 3.0 nM for 30 min at 30 degrees C. Saturating curve analysis indicated no difference in the affinity of glucagon receptors (0.57 +/- 0.06 and 0.77 +/- 0.09 nM in untrained and trained groups, respectively) but a significant higher glucagon receptor density in liver from untrained vs. trained rats (3.09 +/- 0.12 vs. 4.28 +/- 0.19 pmol/mg proteins). These results suggest that the reported increase in liver glucagon sensitivity in endurance-trained subjects (Drouin R, Lavoie C, Bourque J, Ducros F, Poisson D, and Chiasson J-L. Am J Physiol Endocrinol Metab 274: E23-E28, 1998) could be partly due to an increased glucagon receptor density in response to training. PMID- 11120676 TI - Bioinorganic motifs: towards functional classification of metalloproteins. AB - The habitat of bioinorganic motifs (BIMs) is at the interface of biological inorganic chemistry and bioinformatics. BIM is defined as a common structural feature shared by functionally related, but not necessarily homologous, proteins, and consisting of the metal atom(s) and first coordination shell ligands. BIMs appear to be suitable for classification of metal centres at any level, from groups of unrelated proteins with similar function to different functional states of the same protein, and for description of possible evolutionary relationships of metalloproteins. However, they have not attracted wide attention from the bioinformatics community. Although their presence is appreciated, they are difficult to predict-therefore the current 'high-throughput' initiatives are likely to miss or ignore them altogether. The protein sequence databases do not distinguish between proteins containing different prosthetic groups (unless they have different sequences) or between apo- and holoprotein. On the other hand, the protein structure databases include data on 'hetero compounds' of various origin but these data are often inconsistent. A number of specialized databases dealing with BIMs and attempts to classify them are reviewed. SUPPLEMENTARY INFORMATION: The additional bibliography and list of Internet resources on bioinorganic chemistry are available at http://www.ebi.ac.uk/ approximately kirill/biometal/ PMID- 11120677 TI - Sequence analysis by additive scales: DNA structure for sequences and repeats of all lengths. AB - MOTIVATION: DNA structure plays an important role in a variety of biological processes. Different di- and tri-nucleotide scales have been proposed to capture various aspects of DNA structure including base stacking energy, propeller twist angle, protein deformability, bendability, and position preference. Yet, a general framework for the computational analysis and prediction of DNA structure is still lacking. Such a framework should in particular address the following issues: (1) construction of sequences with extremal properties; (2) quantitative evaluation of sequences with respect to a given genomic background; (3) automatic extraction of extremal sequences and profiles from genomic databases; (4) distribution and asymptotic behavior as the length N of the sequences increases; and (5) complete analysis of correlations between scales. RESULTS: We develop a general framework for sequence analysis based on additive scales, structural or other, that addresses all these issues. We show how to construct extremal sequences and calibrate scores for automatic genomic and database extraction. We show that distributions rapidly converge to normality as Nincreases. Pairwise correlations between scales depend both on background distribution and sequence length and rapidly converge to an analytically predictable asymptotic value. For di- and tri-nucleotide scales, normal behavior and asymptotic correlation values are attained over a characteristic window length of about 10-15 bp. With a uniform background distribution, pairwise correlations between empirically derived scales remain relatively small and roughly constant at all lengths, except for propeller twist and protein deformability which are positively correlated. There is a positive (resp. negative) correlation between dinucleotide base stacking (resp. propeller twist and protein deformability) and AT-content that increases in magnitude with length. The framework is applied to the analysis of various DNA tandem repeats. We derive exact expressions for counting the number of repeat unit classes at all lengths. Tandem repeats are likely to result from a variety of different mechanisms, a fraction of which is likely to depend on profiles characterized by extreme structural features. PMID- 11120678 TI - Information theoretical probe selection for hybridisation experiments. AB - MOTIVATION: The choice of probes is an important feature of hybridisation experiments. In this paper we present an algorithm that optimises probes with respect to a training set of sequences based on Shannon entropy as a quality criterion. The practical motivation for our algorithm is oligonucleotide fingerprinting, a method for the simultaneous identification of sequences (cDNA or genomic DNA) by their hybridisation tags according to a set of short probes such as octamers, although the algorithm is of course not restricted to that application. RESULTS: We can show that our method is superior to the selection of probes according to their frequencies, which is a widely used strategy, and to randomly chosen probe sets. The quality of probe sets is assessed by a simulation pipeline that entails the set of probes as a simulation parameter. The performance of probe sets trained on sequences from different organisms shows additionally that probes should be chosen with regard to the organism under analysis. Case studies are presented on how constraints (G+C-content, complexity of the individual probes) influence the selection process. AVAILABILITY: A description of the oligonucleotide fingerprinting pipeline is published on our web-page http://www.molgen.mpg.de/ approximately ag_onf/met.htm. An executable of the algorithm and probe lists designed for human and rodents can be downloaded from the ftp-site ftp://ftp.molgen.mpg.de/pub/mpimg/probe_design/. PMID- 11120679 TI - USAGE: a web-based approach towards the analysis of SAGE data. Serial Analysis of Gene Expression. AB - MOTIVATION: SAGE enables the determination of genome-wide mRNA expression profiles. A comprehensive analysis of SAGE data requires software, which integrates (statistical) data analysis methods with a database system. Furthermore, to facilitate data sharing between users, the application should reside on a central server and be accessed via the internet. Since such an application was not available we developed the USAGE package. RESULTS: USAGE is a web-based application that comprises an integrated set of tools, which offers many functions for analysing and comparing SAGE data. Additionally, USAGE includes a statistical method for the planning of new SAGE experiments. USAGE is available in a multi-user environment giving users the option of sharing data. USAGE is interfaced to a relational database to store data and analysis results. The USAGE query editor allows the composition of queries for searching this database. Several database functions have been included which enable the selection and combination of data. USAGE provides the biologist increased functionality and flexibility for analysing SAGE data. AVAILABILITY: USAGE is freely accessible for academic institutions at http://www.cmbi.kun.nl/usage/. The source code of USAGE is freely available for academic institutions on request from the first author. PMID- 11120680 TI - Support vector machine classification and validation of cancer tissue samples using microarray expression data. AB - MOTIVATION: DNA microarray experiments generating thousands of gene expression measurements, are being used to gather information from tissue and cell samples regarding gene expression differences that will be useful in diagnosing disease. We have developed a new method to analyse this kind of data using support vector machines (SVMs). This analysis consists of both classification of the tissue samples, and an exploration of the data for mis-labeled or questionable tissue results. RESULTS: We demonstrate the method in detail on samples consisting of ovarian cancer tissues, normal ovarian tissues, and other normal tissues. The dataset consists of expression experiment results for 97,802 cDNAs for each tissue. As a result of computational analysis, a tissue sample is discovered and confirmed to be wrongly labeled. Upon correction of this mistake and the removal of an outlier, perfect classification of tissues is achieved, but not with high confidence. We identify and analyse a subset of genes from the ovarian dataset whose expression is highly differentiated between the types of tissues. To show robustness of the SVM method, two previously published datasets from other types of tissues or cells are analysed. The results are comparable to those previously obtained. We show that other machine learning methods also perform comparably to the SVM on many of those datasets. AVAILABILITY: The SVM software is available at http://www.cs. columbia.edu/ approximately bgrundy/svm. PMID- 11120681 TI - CAST: an iterative algorithm for the complexity analysis of sequence tracts. Complexity analysis of sequence tracts. AB - MOTIVATION: Sensitive detection and masking of low-complexity regions in protein sequences. Filtered sequences can be used in sequence comparison without the risk of matching compositionally biased regions. The main advantage of the method over similar approaches is the selective masking of single residue types without affecting other, possibly important, regions. RESULTS: A novel algorithm for low complexity region detection and selective masking. The algorithm is based on multiple-pass Smith-Waterman comparison of the query sequence against twenty homopolymers with infinite gap penalties. The output of the algorithm is both the masked query sequence for further analysis, e.g. database searches, as well as the regions of low complexity. The detection of low-complexity regions is highly specific for single residue types. It is shown that this approach is sufficient for masking database query sequences without generating false positives. The algorithm is benchmarked against widely available algorithms using the 210 genes of Plasmodium falciparum chromosome 2, a dataset known to contain a large number of low-complexity regions. AVAILABILITY: CAST (version 1.0) executable binaries are available to academic users free of charge under license. Web site entry point, server and additional material: http://www.ebi.ac.uk/research/cgg/services/cast/ PMID- 11120682 TI - The reduction of large molecular profiles to informative components using a genetic algorithm. AB - MOTIVATION: Molecular profiles (DNA fingerprints) may be used to allocate an individual of unknown membership to one among the known groups of a reference population. Time and costs of profile assessment may be reduced by identifying informative profile components (markers). RESULTS: A genetic algorithm (GA) is proposed to identify promising candidate markers from a pilot experiment in which observations are supposed to be without measurement error. The analysis of simulated datasets suggests reasonable values for GA parameters and confirms that the GA finds components of the profile showing association with the considered groups. Our GA may be used to perform a first screening of candidate markers to be included in subsequent experiments. AVAILABILITY: The 32-bit executable (Windows 95, 98 and NT) is available at http://www.ds.unifi.it/ approximately stefanin/bioinformatics.htm. PMID- 11120683 TI - Finding pathogenicity islands and gene transfer events in genome data. AB - MOTIVATION: There is a growing literature on wavelet theory and wavelet methods showing improvements on more classical techniques, especially in the contexts of smoothing and extraction of fundamental components of signals. G+C patterns occur at different lengths (scales) and, for this reason, G+C plots are usually difficult to interpret. Current methods for genome analysis choose a window size and compute a chi(2) statistics of the average value for each window with respect to the whole genome. RESULTS: Firstly, wavelets are used to smooth G+C profiles to locate characteristic patterns in genome sequences. The method we use is based on performing a chi(2) statistics on the wavelet coefficients of a profile; thus we do not need to choose a fixed window size, in that the smoothing occurs at a set of different scales. Secondly, a wavelet scalogram is used as a measure for sequence profile comparison; this tool is very general and can be applied to other sequence profiles commonly used in genome analysis. We show applications to the analysis of Deinococcus radiodurans chromosome I, of two strains of Helicobacter pylori (26695, J99) and two of Neisseria meningitidis (serogroup B strain MC58 and serogroup A strain Z2491). We report a list of loci that have different G+C content with respect to the nearby regions; the analysis of N. meningitidis serogroup B shows two new large regions with low G+C content that are putative pathogenicity islands. AVAILABILITY: Software and numerical results (profiles, scalograms, high and low frequency components) for all the genome sequences analyzed are available upon request from the authors. PMID- 11120684 TI - Matching amino acid and nucleotide sequences of mouse rheumatoid factor CDRH3 FRH4 segments to other mouse antibodies with known specificities. PMID- 11120685 TI - Artemis: sequence visualization and annotation. AB - SUMMARY: Artemis is a DNA sequence visualization and annotation tool that allows the results of any analysis or sets of analyses to be viewed in the context of the sequence and its six-frame translation. Artemis is especially useful in analysing the compact genomes of bacteria, archaea and lower eukaryotes, and will cope with sequences of any size from small genes to whole genomes. It is implemented in Java, and can be run on any suitable platform. Sequences and annotation can be read and written directly in EMBL, GenBank and GFF format. AVAILABITLTY: Artemis is available under the GNU General Public License from http://www.sanger.ac.uk/Software/Artemis PMID- 11120686 TI - MAD: a suite of tools for microarray data management and processing. AB - SUMMARY: Microarray data management and processing (MAD) is a set of Windows integrated software for microarray analysis. It consists of a relational database for data storage with many user-interfaces for data manipulation, several text file parsers and Microsoft Excel macros for automation of data processing, and a generator to produce text files that are ready for cluster analysis. AVAILABILITY: Executable is available free of charge on http://pompous.swmed.edu. The source code is also available upon request. PMID- 11120687 TI - A space-efficient algorithm for aligning large genomic sequences. AB - SUMMARY: In the segment-by-segment approach to sequence alignment, pairwise and multiple alignments are generated by comparing gap-free segments of the sequences under study. This method is particularly efficient in detecting local homologies, and it has been used to identify functional regions in large genomic sequences. Herein, an algorithm is outlined that calculates optimal pairwise segment-by segment alignments in essentially linear space. AVAILABILTIY: The program is available at the Bielefeld Bioinformatics Server (BiBiServ) at http://bibiserv.techfak. uni-bielefeld.de/dialign/ PMID- 11120688 TI - MASIA: recognition of common patterns and properties in multiple aligned protein sequences. AB - SUMMARY: MASIA is a software tool for pattern recognition in multiple aligned protein sequences. MASIA converts a sequence to a properties matrix that can be scanned in both vertical and horizontal steps. Consistent patterns are recognized based on the statistical significance of their occurrence. Preset macros can be altered on-line to seek any combination of amino acid properties or sequence characteristics. MASIA output can be used directly by our programs to predict the 3D structure of proteins. AVAILABILITY: Access MASIA at http://www.scsb.utmb.edu/masia/ma sia.html. PMID- 11120689 TI - Chips ahoy: gene expression in failing hearts surveyed by high-density microarrays. PMID- 11120690 TI - Emergency polytetrafluoroethylene-covered stent implantation to treat coronary ruptures. AB - BACKGROUND: Coronary perforation is a life-threatening complication of percutaneous interventions. In the past few years, the implantation of covered stents has emerged as a strategy for treatment when the traditional conservative approach (ie, prolonged balloon inflation and reversal of anticoagulation) fails. METHODS AND RESULTS: Since May 1997 (when polytetrafluoroethylene [PTFE]-covered stents were available at our institutions), 11 of the 12 consecutive patients who had coronary ruptures that were unsuccessfully sealed with prolonged balloon inflation and reversal of anticoagulation were treated with 12 PTFE-covered stents (PTFE group). The efficacy of the PTFE-covered stent was compared with that of noncovered stents, which were used to treat 17 perforations (non-PTFE group). One patient sustained a very distal perforation that was not suitable for covered stent sealing and underwent emergency surgery. All vessel ruptures treated with PTFE-covered stent implantation were successfully sealed. The time necessary to deploy the stent was 10+/-3 minutes (range, 4 to 15 minutes). All patients but one were discharged from the hospital and had an optimal early clinical outcome. One patient underwent emergency bypass surgery and died in the intensive care unit. The occurrence of cardiac tamponade and the necessity for emergency surgery was significantly lower in the PTFE group than in the non-PTFE group. At 14+/-4 months, the 10 discharged patients had not experienced any major adverse cardiac events. CONCLUSIONS: This preliminary study supports the utility of the PTFE-covered stent for the nonsurgical treatment of vessel ruptures. PMID- 11120691 TI - Inhibition of the sodium-hydrogen exchanger with cariporide to prevent myocardial infarction in high-risk ischemic situations. Main results of the GUARDIAN trial. Guard during ischemia against necrosis (GUARDIAN) Investigators. AB - BACKGROUND: The transmembrane sodium/hydrogen exchanger maintains myocardial cell pH integrity during myocardial ischemia but paradoxically may precipitate cell necrosis. The development of cariporide, a potent and specific inhibitor of the exchanger, prompted this investigation of the potential of the drug to prevent myocardial cell necrosis. METHODS AND RESULTS: A total of 11 590 patients with unstable angina or non-ST-elevation myocardial infarction (MI) or undergoing high risk percutaneous or surgical revascularization were randomized to receive placebo or 1 of 3 doses of cariporide for the period of risk. The trial failed to document benefit of cariporide over placebo on the primary end point of death or MI assessed after 36 days. Doses of 20 and 80 mg every 8 hours had no effect, whereas a dose of 120 mg was associated with a 10% risk reduction (98% CI 5.5% to 23.4%, P=0.12). With this dose, benefit was limited to patients undergoing bypass surgery (risk reduction 25%, 95% CI 3.1% to 41.5%, P=0.03) and was maintained after 6 months. No effect was seen on mortality. The rate of Q-wave MI was reduced by 32% across all entry diagnostic groups (2.6% versus 1.8%, P=0.03), but the rate of non-Q-wave MI was reduced only in patients undergoing surgery (7.1% versus 3.8%, P=0.005). There were no increases in clinically serious adverse events. CONCLUSIONS: No significant benefit of cariporide could be demonstrated across a wide range of clinical situations of risk. The trial documented safety of the drug and suggested that a high degree of inhibition of the exchanger could prevent cell necrosis in settings of ischemia-reperfusion. PMID- 11120692 TI - Association of influenza vaccination and reduced risk of recurrent myocardial infarction. AB - BACKGROUND: Numerous studies have suggested that microbial agents may promote atherosclerosis. A smaller body of research has suggested that acute respiratory infection may be a risk factor for myocardial infarction (MI). We hypothesized that influenza vaccine might reduce the risk of recurrent MI in patients with documented coronary heart disease (CHD). METHODS AND RESULTS: A case-control study was performed on 218 CHD patients seen at Memorial Hermann Hospital during the influenza season of October 1997 through March 1998. Patients who experienced new MI were included in the case group, and those who did not experience new MI or unstable angina were assigned to the control group. Data were collected by structured review of patients' charts and through a subsequent telephone survey. Adjusted for history of influenza vaccination in previous years, multivariate logistic regression revealed risk of MI to be associated with current hypertension (OR 4.96, 95% CI 2.06 to 11.96, P<0.0001), hypercholesterolemia (OR 4.08, 95% CI 1.67 to 9.99, P=0.002), smoking (OR 3.75, 95% CI 1.76 to 7.98, P=0.001), and influenza vaccination (OR 0.33, 95% CI 0.13 to 0.82, P=0.017). Despite significant association in univariate analysis, multivitamin therapy and physical exercise were not associated with risk of reinfarction in multivariate analysis. CONCLUSIONS: In this study in patients with chronic CHD, vaccination against influenza was negatively associated with the development of new MI during the same influenza season. However, to address causal inference, examination of prospective data sets will be needed. PMID- 11120693 TI - Decreased SLIM1 expression and increased gelsolin expression in failing human hearts measured by high-density oligonucleotide arrays. AB - BACKGROUND: Failing human hearts are characterized by altered cytoskeletal and myofibrillar organization, impaired signal transduction, abnormal protein turnover, and impaired energy metabolism. Thus, expression of multiple classes of genes is likely to be altered in human heart failure. METHODS AND RESULTS: We used high-density oligonucleotide arrays to explore changes in expression of approximately 7000 genes in 2 nonfailing and 2 failing human hearts with diagnoses of end-stage ischemic and dilated cardiomyopathy, respectively. We report altered expression of (1) cytoskeletal and myofibrillar genes (striated muscle LIM protein-1 [SLIM1], myomesin, nonsarcomeric myosin regulatory light chain-2 [MLC(2)], and ss-actin); (2) genes responsible for degradation and disassembly of myocardial proteins (alpha(1)-antichymotrypsin, ubiquitin, and gelsolin); (3) genes involved in metabolism (ATP synthase alpha-subunit, succinate dehydrogenase flavoprotein [SDH Fp] subunit, aldose reductase, and TIM17 preprotein translocase); (4) genes responsible for protein synthesis (elongation factor-2 [EF-2], eukaryotic initiation factor-4AII, and transcription factor homologue-HBZ17); and (5) genes encoding stress proteins (alphaB crystallin and mu-crystallin). In 5 additional failing hearts and 4 additional nonfailing controls, we then compared expression of proteins encoded by the differentially expressed genes, alphaB-crystallin, SLIM1, gelsolin, alpha(1) antichymotrypsin, and ubiquitin. In each case, changes in protein expression were consistent with changes in transcript measured by microarray analysis. Gelsolin protein expression was also increased in cardiomyopathic hearts from tropomodulin overexpressing (TOT) mice and rac1-expressing (racET) mice. CONCLUSIONS: Altered expression of the genes identified in this study may contribute to development of the heart failure phenotype and/or represent compensatory mechanisms to sustain cardiac function in failing human hearts. PMID- 11120694 TI - Left ventricular or biventricular pacing improves cardiac function at diminished energy cost in patients with dilated cardiomyopathy and left bundle-branch block. AB - BACKGROUND: Left ventricular or biventricular pacing/stimulation can acutely improve systolic function in patients with dilated cardiomyopathy (DCM) and intraventricular conduction delay by resynchronizing contraction. Most heart failure therapies directly enhancing systolic function do so while concomitantly increasing myocardial oxygen consumption (MVO(2)). We hypothesized that pacing/stimulation, in contrast, incurs systolic benefits without raising energy demand. METHODS AND RESULTS: Ten DCM patients with left bundle-branch block (ejection fraction 20+/-3%, QRS duration 179+/-3 ms, mean+/-SEM) underwent cardiac catheterization to measure ventricular and aortic pressure, coronary blood flow, arterial-coronary sinus oxygen difference (DeltaAVO(2)), and MVO(2). Data were measured under sinus rhythm or with left ventricular or biventricular pacing/stimulation at the same heart rate. These results were then contrasted to intravenous dobutamine (n=7) titrated to match systolic changes during LV pacing. Systolic function rose quickly and substantially from LV pacing (18+/-4% rise in arterial pulse pressure, which correlates with cardiac output, and 43+/-6% increase in dP/dt(max); both P<0.01). However, DeltaAVO(2) and MVO(2) declined 4+/-2% and -8+/-6.5%, respectively (both P<0.05). Similar results were obtained with biventricular activation. In contrast, dobutamine raised dP/dt(max) 37+/-6%, accompanied by a 22+/-11% rise in per-beat MVO(2) (P<0.05 versus pacing). CONCLUSIONS: Ventricular resynchronization by left ventricular or biventricular pacing/stimulation in DCM patients with left bundle-branch block acutely enhances systolic function while modestly lowering energy cost. This should prove valuable for treating DCM patients with basal dyssynchrony. PMID- 11120695 TI - Plasma cytokine parameters and mortality in patients with chronic heart failure. AB - BACKGROUND: Inflammatory immune activation is an important feature in chronic heart failure (CHF). Little is known about the prognostic importance of tumor necrosis factor-alpha (TNF-alpha), soluble TNF-receptor 1 and 2 (sTNF-R1/sTNF R2), interleukin-6 (IL-6), and soluble CD14 receptors (sCD14) in CHF patients. METHODS AND RESULTS: In 152 CHF patients (age 61+/-1 years, New York Heart Association [NYHA] class 2.6+/-0.1, peak VO(2) 17.3+/-0.6 mL. kg(-1). min(-1), mean+/-SEM) plasma concentrations of immune variables were prospectively assessed. During a mean follow-up of 34 months (>12 months in all patients), 62 patients (41%) died. Cumulative mortality was 28% at 24 months. In univariate analyses, increased total and trimeric TNF-alpha, sTNF-R1, and sTNF-R2 (all P/=25 g/d. During a mean follow-up of 6.8 years, 414 subjects developed IC. From the lowest to the highest category of alcohol intake, the age-standardized incidence rates of IC were 5.3, 4.1, 4.2, 3.2, and 4.6 cases/1000 person-years for men and 3.4, 2.5, 1.5, 1.9, and 2.5, respectively, for women. A multivariate Cox regression model demonstrated an inverse relation, with the lowest IC risk at levels of 13 to 24 g/d for men and 7 to 12 g/d for women compared with nondrinkers; the hazard ratio (95% CI) was 0.67 (0.42 to 0.99) for men and 0.44 (0.23 to 0.80) for women. This protective effect was seen mostly with wine and beer consumption. CONCLUSIONS: Our data are consistent with a protective effect of moderate alcohol consumption on IC risk, with lowest risk observed in men consuming 13 to 24 g/d (1 to 2 drinks/d) and in women consuming 7 to 12 g/d (0.5 to 1 drink/d). PMID- 11120701 TI - Ischemia-induced coronary collateral growth is dependent on vascular endothelial growth factor and nitric oxide. AB - BACKGROUND: We hypothesized that ischemia-induced expression of vascular endothelial growth factor (VEGF) and the production of NO stimulate coronary collateral growth. METHODS AND RESULTS: To test this hypothesis, we measured coronary collateral blood flow and VEGF expression in myocardial interstitial fluid in a canine model of repetitive myocardial ischemia under control conditions and during antagonism of NO synthase. Collateralization was induced by multiple (1/h; 8/d), brief (2 minutes) occlusions of the left anterior descending coronary artery for 21 days. In controls, collateral blood flow (microspheres) progressively increased to 89+/-9 mL. min(-1). 100 g(-1) on day 21, which was equivalent to perfusion in the normal zone. Reactive hyperemic responses (a measure of the severity of ischemia) decreased as collateral blood flow increased. In N(G)-nitro-L-arginine methyl ester (L-NAME)- and L-NAME+nifedipine treated dogs, to block the production of NO and control hypertension, respectively, collateral blood flow did not increase and reactive hyperemia was robust throughout the occlusion protocol (P<0.01 versus control). VEGF expression (Western analyses of VEGF(164) in myocardial interstitial fluid) in controls peaked at day 3 of the repetitive occlusions but waned thereafter. In sham operated dogs (instrumentation but no occlusions), expression of VEGF was low during the entire protocol. In contrast, VEGF expression was elevated throughout the 21 days of repetitive occlusions after L-NAME. Reverse transcriptase polymerase chain reaction analyses revealed that the predominant splice variant expressed was VEGF(164). CONCLUSIONS: NO is an important regulator of coronary collateral growth, and the expression of VEGF is induced by ischemia. Furthermore, the induction of coronary collateralization by VEGF appears to require the production of NO. PMID- 11120702 TI - Suppression of endothelial nitric oxide production after withdrawal of statin treatment is mediated by negative feedback regulation of rho GTPase gene transcription. AB - BACKGROUND: Statins improve endothelial function by upregulating endothelial nitric oxide (NO) production that is mediated by inhibiting the isoprenylation of rho GTPase. Withdrawal of statin treatment could suppress endothelial NO production and may impair vascular function. METHODS AND RESULTS: To test this hypothesis, mice were treated for 14 days with 10 mg/kg atorvastatin per day; this led to the upregulation of endothelial NO synthase expression and activity by 2.3- and 3-fold, respectively. Withdrawal of statins resulted in a dramatic, 90% decrease of NO production after 2 days. In mouse aortas and cultured endothelial cells, statins upregulated the expression of rho GTPase in the cytosol, but statins blocked isoprenoid-dependent rho membrane translocation and GTP-binding activity. Inhibiting the downstream targets of rho showed that rho expression is controlled by a negative feedback mechanism mediated by the actin cytoskeleton. Measuring rho mRNA half-life and nuclear run-on assays demonstrated that statins or disruption of actin stress fibers increased rho gene transcription but not rho mRNA stability. Therefore, treatment with statins leads to the accumulation of nonisoprenylated rho in the cytosol. Withdrawing statin treatment restored the availability of isoprenoids and resulted in a massive membrane translocation and activation of rho, causing downregulation of endothelial NO production. CONCLUSIONS: Withdrawal of statin therapy in normocholesterolemic mice results in a transient increase of rho activity, causing a suppression of endothelial NO production. The underlying molecular mechanism is a negative feedback regulation of rho gene transcription mediated by the actin cytoskeleton. PMID- 11120704 TI - Effects of beta(-)-emitting (188)Re balloon in stented porcine coronary arteries: an angiographic, intravascular ultrasound, and histomorphometric study. AB - BACKGROUND: Restenosis within stents may be prevented by ionizing radiation from an intravascular source. METHODS AND RESULTS: A liquid beta(-) radiation ((188)Re) balloon was evaluated in a randomized and blinded porcine coronary model of stent restenosis. Group A pigs (n=17) received 0,16, 22, or 29 Gy at 0.5 mm depth, followed by stenting. Restenosis was quantified by angiography, ultrasound, and histomorphometry at 30 days. Group B (n=7) was stented first and then treated with 0 or 29 Gy with follow-up at 60 days. There was a measurable effect at 16 Gy, which improved with increasing doses. At 29 Gy, the histological stenotic area was reduced by 67% (22% versus 66% in controls, P<0.001). Radiation after stenting was equally effective; the stenotic area was reduced (21% versus 65%, P<0.001). At 16 Gy, the vessel just distal to the stent was significantly smaller than control vessels because of intimal thickening (P=0.003). Radiated vessels had distinctive histology consisting of neointimal fibrin and reduced smooth muscle cells and matrix (P<0.0001). CONCLUSIONS: (188)Re balloon brachytherapy in porcine coronary arteries results in dose-dependent and injury independent inhibition of stent restenosis for up to 60 days. Restenosis at the borders of the irradiated zone is a potential limitation and may be related to underdosing. Brachytherapy with the (188)Re balloon appears to be safe and feasible for clinical studies. PMID- 11120703 TI - Ischemia-reperfusion injury at the microvascular level: treatment by endothelin A selective antagonist and evaluation by myocardial contrast echocardiography. AB - BACKGROUND: The purpose of this study was to verify whether endothelin A antagonist administration at the time of coronary reperfusion preserves postischemic microvasculature and whether myocardial contrast echo (MCE) is able to detect pharmacologically induced changes in microvascular reflow. METHODS AND RESULTS: Twenty dogs underwent 90 minutes of LAD occlusion (OCC) followed by 180 minutes of reperfusion (RP). Five minutes before LAD reopening, an intravenous bolus (5 mg/kg) of LU 135252 was given in 10 dogs and vehicle in the remaining 10. At baseline (BSL), OCC, and 90 and 180 minutes of RP, microvascular flow (BF) was assessed by microspheres, and MCE was performed with intravenous echo contrast. MCE videointensity and BF were expressed as risk area/control ratio. Myocardial thickness of the risk area was calculated by 2D echo. No differences in BF between the 2 groups were observed at BSL, OCC, and 90 minutes of RP. At 180 minutes of RP, BF was decreased in controls (70+/-7.4% of BSL; P:<0.005 versus BSL) and preserved in LU 135252-treated animals (89+/-4% of BSL; P=NS versus BSL; P<0.05 versus controls). Videointensity at MCE closely followed the changes in BF observed in both groups throughout the protocol. Myocardial thickness at 180 minutes of RP increased to 138.6+/-9.9% of BSL in controls and remained at 108.9+/-7.4% of BSL in treated dogs (P<0.05). CONCLUSIONS: Endothelin A-antagonist treatment at the time of reperfusion significantly limited the progressive decrease in postischemic microvascular reflow and the increase in myocardial thickness. MCE allowed a reliable evaluation of pharmacologically induced changes in microvascular flow. PMID- 11120705 TI - Hibernation in noncontracting mammalian cardiomyocytes. AB - BACKGROUND: -The aim of the present study was to establish whether isolated neonatal mammalian cardiomyocytes were capable of downregulating energy-using processes other than contraction while maintaining metabolic stability when oxygen availability was reduced. METHODS AND RESULTS: Metabolic response of cardiomyocytes was investigated under moderate (5 to 6 micromol/L) and severe (2 to 3 micromol/L) forms of hypoxia. Cells exposed to oxygen concentrations of 5 to 6 micromol/L exhibited rates of oxygen consumption, which were decreased to 64% of normoxic rates. Rates of cellular energy usage were decreased because this reduced rate of oxygen consumption was not associated with either decreased intracellular ATP and phosphocreatine concentrations or a compensatory switch to glycolysis. When cells were exposed to oxygen concentrations of 2 to 3 micromol/L, rates of oxygen consumption decreased to 9% of normoxic rates. This decreased rate of oxygen consumption was associated with energetic stress, because a significant switch to glycolysis occurred and intracellular phosphocreatine concentrations were decreased by 40%. Rates of cellular energy usage were further decreased as indicated by stable intracellular ATP concentrations. CONCLUSIONS: -Our results suggest that isolated cardiomyocytes are capable of downregulating energy-consuming processes other than contraction when oxygen supply is decreased. Regions of myocardial tissue are also capable of downregulating metabolic activity during ischemia by shutting down contractile activity (myocardial hibernation). We suggest that metabolic downregulation associated with myocardial hibernation may not be exclusively due to reduced rates of contractile activity. Other energy-using processes (eg, protein synthesis, mRNA synthesis, ion channel activity, and proton leak) may also be shut down. PMID- 11120706 TI - Prostacyclin analogues differentially inhibit growth of distal and proximal human pulmonary artery smooth muscle cells. AB - BACKGROUND: Prostacyclin has proved to be a beneficial treatment for patients with severe pulmonary hypertension. We postulated that the response may reflect, at least in part, inhibition of pulmonary artery smooth muscle cell (PASMC) growth. METHODS AND RESULTS: Human PASMCs were derived from distal (<1-mm external diameter, n=8) and proximal (>8-mm external diameter, n=12) pulmonary arteries obtained at transplant surgery and pneumonectomy. The effects of the stable prostacyclin analogues on [methyl-(3)H]thymidine incorporation and cell proliferation were investigated by using immunohistochemically characterized cells. Distal cells proliferated faster than did proximal PASMCs and displayed a distinct sensitivity to cicaprost and iloprost. Both analogues inhibited thymidine uptake over 24 hours (20% to 60%, P<0.001; n=8) and abolished stimulation of DNA synthesis by platelet-derived growth factor-BB (10 ng/mL) in distal but not proximal cells. The inhibitory effect of cicaprost was mimicked by isoproterenol (10(-5) mol/L), forskolin (10(-5) mol/L), and dibutyryl cAMP (5x10( 4) mol/L) and was potentiated by the phosphodiesterase inhibitor 3-isobutyl-1 methylxanthine (5x10(-5) mol/L). Cicaprost (10(-10) to 10(-6) mol/L) inhibited the proliferation of PASMCs, which had been stimulated with either platelet derived growth factor-BB or serum, and increased cAMP production. These effects were potentiated by 3-isobutyl-1-methylxanthine and attenuated by the adenylyl cyclase inhibitor 2',5'-dideoxyadenosine (10(-5) to 10(-4) mol/L). CONCLUSIONS: ++Cicaprost and iloprost inhibit DNA synthesis and proliferation to a greater extent in distal compared with proximal human PASMCs, acting at least in part via a cAMP-dependent mechanism. The results are consistent with the hypothesis that prostacyclin analogues inhibit vascular remodeling in pulmonary hypertension and demonstrate heterogeneity among human PASMCs. PMID- 11120707 TI - Trends and disparities in coronary heart disease, stroke, and other cardiovascular diseases in the United States: findings of the national conference on cardiovascular disease prevention. AB - A workshop was held September 27 through 29, 1999, to address issues relating to national trends in mortality and morbidity from cardiovascular diseases; the apparent slowing of declines in mortality from cardiovascular diseases; levels and trends in risk factors for cardiovascular diseases; disparities in cardiovascular diseases by race/ethnicity, socioeconomic status, and geography; trends in cardiovascular disease preventive and treatment services; and strategies for efforts to reduce cardiovascular diseases overall and to reduce disparities among subpopulations. The conference concluded that coronary heart disease mortality is still declining in the United States as a whole, although perhaps at a slower rate than in the 1980s; that stroke mortality rates have declined little, if at all, since 1990; and that there are striking differences in cardiovascular death rates by race/ethnicity, socioeconomic status, and geography. Trends in risk factors are consistent with a slowing of the decline in mortality; there has been little recent progress in risk factors such as smoking, physical inactivity, and hypertension control. There are increasing levels of obesity and type 2 diabetes, with major differences among subpopulations. There is considerable activity in population-wide prevention, primary prevention for higher risk people, and secondary prevention, but wide disparities exist among groups on the basis of socioeconomic status and geography, pointing to major gaps in efforts to use available, proven approaches to control cardiovascular diseases. Recommendations for strategies to attain the year 2010 health objectives were made. PMID- 11120708 TI - Contrast-enhanced magnetic resonance angiogram of coronary artery bypass graft aneurysm. PMID- 11120709 TI - High antifibrinolytic activity of lipoprotein(a) containing small apolipoprotein(a) isoforms. PMID- 11120710 TI - Attainment and maintenance of platelet inhibition through standard dosing of abciximab in patients undergoing percutaneous coronary intervention. PMID- 11120711 TI - Use of placebo in heart failure research. PMID- 11120712 TI - Relation of contractile reserve of hibernating myocardium to myocardial structure in humans. PMID- 11120713 TI - Cardiovascular news. AMIOVIRT. PMID- 11120714 TI - An International Forum for Family Doctors. PMID- 11120715 TI - Developing a 'consultation quality index' (CQI) for use in general practice. AB - BACKGROUND: The core values of general practice include holism and patient centredness. None of the measures of quality of care in general practice presently capture the expression of these values at routine consultations. OBJECTIVES: The aim of the present study was to construct a 'consultation quality index' (CQI) which reflects the core values of general practice, using as proxies 'consultation length' and how well patients 'know the doctor' as process measures and 'patient enablement' as an outcome measure. METHODS: The CQI was constructed from data collected from 23 799 adult English-speaking patients consulting 221 doctors in four demographically contrasting areas of the UK during 2 weeks of March/April 1998. A total of 171 doctors who entered 50 qualifying consultations were allocated scores for the three component variables, and a total CQI was calculated. RESULTS: CQI scores were in the range 4-18. Validity was examined by looking at high and low scorers in greater detail and by searching for correlates with case mix, patient age and gender, and the deprivation scores of the practices concerned. Particular attention was paid to how registrars and doctors new to their practices scored. The scores of different doctors in the same practice were also noted. The results had strong face validity and were independent of case mix and deprivation. Reliability was gauged by examining similar work from a previous study which had collected information on consultation length and enablement over three time periods. High CQI scores were associated with smaller overall practice list sizes. CONCLUSIONS: We have outlined possible uses for the CQI as part of the packages assessing quality of care by doctors and practices. The measure may also have a part to play in recognizing poorly performing doctors. We suggest how CQI scores could contribute to an incentive scheme to reward good consulting practice. Further work is in hand to compare doctors' CQI scores with scores based on performance indicators constructed from routine NHS data on prescribing and preventive medicine. PMID- 11120716 TI - Variation in GP referral rates: what can we learn from the literature? AB - BACKGROUND: Variations in referral rates exist, at GP and practice level. Although the National Institute for Clinical Excellence is to produce referral guidelines, it is unclear if this variation requires regulation. A critical review of the literature on variation in referral rates was undertaken to see if existing evidence could inform the debate. OBJECTIVES: The aim of this study was to describe the variation in referral rates; to identify likely explanatory variables; and to describe the effect of GPs' decision making on the referral process. METHODS: Six bibliographic databases, the Cochrane Library, the NHS Centre for Reviews and Dissemination, and the National Research Register were searched. RESULTS: Patient characteristics explain <40% of the observed variation; practice and GP characteristics <10%. The availability of specialist care does affect referral rates, but its influence on the observed variation of referral rates is not known. Intrinsic psychological variables are important. GPs who are less tolerant of uncertainty or who perceive serious disease to be a more frequent event may refer more patients. There is a lack of consensus about what constitutes an appropriate referral, and the use of guidelines has had only limited success in altering referral behaviour. CONCLUSIONS: Variation in referral rates remains largely unexplained. Targeting high or low referrers through clinical guidelines may not be the issue. Rather, activity should concentrate on increasing the number of appropriate referrals, regardless of the referral rate. Pressure on GPs to review their referral behaviour through the use of guidelines may reduce their willingness to tolerate uncertainty and manage problems in primary care, resulting in an increase in referrals to secondary care. The use of referral rates to stimulate dialogue and joint working between primary and secondary care may be more appropriate. PMID- 11120717 TI - The effect of the full moon on general practice consultation rates. AB - BACKGROUND: The effect of the full moon on human behaviour, the so-called 'Transylvania hypothesis', has fascinated the public and occupied the mind of researchers for centuries. OBJECTIVE: The aim of the present study was to determine whether or not there was any change in general practice consultation patterns around the time of the full moon. METHOD: We analysed data from the fourth national morbidity study of general practice. The data set was split into two groups and analysed separately: consultations on ordinary weekdays and consultations on weekends and bank holidays. The data were split randomly into two equal sets, one for model building and one for model validation. The lunar cycle effect was assumed to be sinusoidal, on the grounds that any effect would be maximal at the time of the full moon and decline to the new moon, following a cosine curve (with a period of 29.54 days, the mean length of a lunar cycle). RESULTS: There was a statistically significant, but small, effect associated with the lunar cycle of 1.8% of the mean value [95% confidence interval (CI) 0.9 2.7%]. This equates to an average difference between the two extremes during the cycle of 3.6%. For this data set, this accounts for 190 (95% CI 95-285) more consultations on days at the peak of the cycle compared with those at the bottom of the cycle, or, put another way, about three consultations per practice. CONCLUSION: We can speculate neither as to what the nature of these moon-related problems may be, nor as to the mechanisms underpinning such behaviour. However, we have confirmed that it does not seem to be related to anxiety and depression. PMID- 11120718 TI - Treating patients with colorectal cancer in rural and urban areas: a qualitative study of the patients' perspective. AB - OBJECTIVES: The aim of the present study was to explore the perspectives of patients receiving treatment for colorectal cancer and compare priorities and attitudes in rural and urban areas. METHOD: A qualitative study was carried out involving four focus groups in the Aberdeen and Northern Centre for Haematology, Oncology and Radiotherapy. The sample comprised 22 patients at various stages of treatment for colorectal cancer and 10 of their relatives from different locations of Northeast Scotland and Shetland. The main themes generated by participants were identified, and similarities and differences between urban and rural patients were noted within these themes. RESULTS: Components of care that were important to rural and urban patients were speed of referral to specialists for treatment and issues of communication including test results and delivery of bad news. Tensions were perceived at the interface of primary and secondary care and these were blamed for delays in referral and communication breakdowns. For some, the referral process comprised a series of barriers to be overcome, and there were additional hurdles for remote patients referred initially to local (non-specialist) hospitals. Rural patients appeared to be less demanding than their urban counterparts when evaluating their care, and this was extended to the acceptance of some clear transport problems. CONCLUSIONS: For patients, the most important components of cancer care were similar, whether rural or urban residents. The main differences for rural patients were lower expectations of care and more hurdles before reaching specialist care. These differences might explain the trend to more advanced disease at diagnosis in rural patients if they lead to significant delays. GPs have major influences on this and all the patients' priorities. PMID- 11120719 TI - 'Strong medicine': an analysis of pharmacist consultations in primary care. AB - BACKGROUND: Patients often find it difficult to discuss their medications fully with their prescribing doctor. Little is known about what might be said about medications to another professional within the primary health care team (PHCT). Pharmacists are seeking to extend their role within primary care and are ideally placed to provide independent medication advice. OBJECTIVE: Our aim was to test the feasibility of using primary care pharmacists as medication counsellors, and to analyse the content of their consultations using qualitative methods. METHOD: Some patients were referred by their doctors, some self-referred and others were invited by the pharmacists for medication reviews. Pharmacist-patient consultations took place within GP surgeries and in patients' homes, and were audiotaped, transcribed and analysed qualitatively. The study sample consisted of 25 consultations with three primary care pharmacists conducted over a 3-month period. RESULTS: Referrals from the doctors were slow and there were no referrals from nurses. The pharmacists, who all had clinical backgrounds and were not dispensing pharmacists, experienced few problems with the consultations. Patients were willing to discuss their medications in detail with the pharmacists. A theme emerged regarding the perceived potency of medications, and this seemed to have an effect on the experience of side effects and the perceived efficacy of the medications. CONCLUSIONS: From this small study, it would seem that pharmacist consultations within primary care are a feasible extension of their current role as prescribing budget advisors. The richness of the consultations reflects the acceptability to patients. Feedback of information to other members of the PHCT, given patient consent, would be very useful for a better understanding of the patient's perspective, which in turn would facilitate concordance in the negotiation of the patient's management. PMID- 11120720 TI - Keeping families of heroin addicts together: results of 13 months' intake for community detoxification and rehabilitation at a family centre for drug users. AB - BACKGROUND: Heroin addiction is a major public health problem affecting both the addicted individuals and their children, who have been shown to have poor social, educational and health status and to be at higher risk of abuse than their peers. Whilst the antenatal effects of parental drug use and the overall poor outcomes for these children have been widely studied, there has been little emphasis on the effectiveness of treatment interventions and even less emphasis on evaluating the effect on children of the standard treatments aimed at their parents' drug use. OBJECTIVES: The aim of the present study was to evaluate the effect on heroin-addicted parents and their children of a family-based drug treatment intervention using a records-based methodology, and to identify any factors at admission which may influence outcome. This study is a pilot for a prospective Europe-wide study using a similar methodology prospectively in several treatment modalities. METHODS: A retrospective cohort study was carried out using existing clinical and residential record data. The setting was a residential family centre run by the charity Phoenix House in Sheffield, UK, offering a 6-month (180 days) family-based programme for heroin addicts including community detoxification overseen by primary care specialist doctors and residential rehabilitation. All adults and children who entered the centre between July 1997 and July 1998 were included in the study (26 adults and 33 children, in 23 family groups). An analysis was made of clinical records and records kept on the adults and children by the clinicians and staff at the centre. The main outcome measures for the adults were length of stay and reason for departure (treatment complete, early planned discharge, unplanned discharge, eviction); and for the children were reason for departure and discharge destination (with parent or taken into care). RESULTS: Mean length of stay was 110 days, and only 11 children (33%) and nine adults (35%) completed 150 days or more. Length of stay was found to be significantly correlated with parental age at admission (P < 0.01). Twelve children (37%) and nine adults (35%) were deemed to have completed treatment successfully. Of the remainder, 14 children (42%) and 11 adults (42%) left because of definite treatment failure. Successful treatment completion was found to be correlated with increased parental age (Pearson's r = 0.612, P = 0. 001). Poly-drug users were significantly less likely to complete treatment successfully (Fisher's exact test, P = 0.012). Twenty children were in the care of their parents on admission, and 24 were able to go home with their parents. There was no association between residence with parents pre- and post-admission (McNemar's chi-squared test = 1.6, P > or = 0.1). CONCLUSIONS: Whilst overall high rates of treatment success are not expected in abstinence-based programmes, the outcomes for the adults in this setting are comparable with published results in other residential settings, and there is some evidence that some families may have stayed together who might otherwise have been separated. Older adults who were not poly-drug users had significantly better outcomes. The records-based methodology proved successful, but centres need to keep detailed and preferably long-term records on children if their outcomes are to be evaluated more fully. PMID- 11120721 TI - The construction of a patient record-based risk model for recurrent falls among elderly people living in the community. AB - BACKGROUND: Predictive models of fall risk in the elderly living in the community may contribute to the identification of elderly at risk for recurrent falling. OBJECTIVES: Our aim was to investigate occurrence, determinants and health consequences of falls in a community-dwelling elderly population and the contribution of data from patient records to a risk model of recurrent falls. METHODS: A population survey was carried out using a postal questionnaire. The questionnaire on occurrence, determinants and health consequences of falls was sent to 2744 elderly persons of 70 years and over, registered in four general practices (n = 27 000). Data were analysed by bivariate techniques and logistic regression. RESULTS: A total of 1660 (60%) responded. Falls (> or =1 fall) in the previous year were reported by 44%: one-off falls by 25% and recurrent falls (> or =2 falls) by 19%. Women had significantly more falls than men. Major injury was reported by 8% of the fallers; minor injury by 49%. Treatment of injuries was by the GP in 67% of cases. From logistic regression, a risk model for recurrent falls, consisting of the risk factors female gender, age 80 years or over, presence of a chronic neurological disorder, use of antidepressants, problems of balance and sense organs and complaints of muscles and joints was developed. The model predicted recurrent falls with a sensitivity of 64%, a specificity of 71%, a positive predictive value of 42% and a negative predictive value of 86%. CONCLUSION: A risk model consisting of six variables usually known to the GP from the patient records may be a useful tool in the identification of elderly people living in the community at risk for recurrent falls. PMID- 11120722 TI - The effect of hormone replacement therapy and route of administration on selected cardiovascular risk factors in post-menopausal women. AB - BACKGROUND: There is increasing use of hormone replacement therapy (HRT) by post menopausal women. Observational epidemiological studies have shown reductions in cardiovascular risk factors in HRT users in the USA, but no randomized controlled trials of HRT have been carried out in the primary practice setting. Previous studies of cardiovascular risk factors have shown a variety of responses according to type of progestagen and oral or topical administration. None has examined the effect of route using an identical progestagen. OBJECTIVES: Our aim was to establish differences, if any, in alteration in cardiovascular risk factors with HRT in post-menopausal women according to route of administration of HRT, oral, transdermal and implant, using first oestrogen alone then oestrogen plus norethisterone, or testosterone for implant. METHODS: Subjects were recruited by letter of invitation to women aged 50-65 years from lists in general practices local to the Charing Cross Hospital Lipid Clinic in West London. Their menopausal status was confirmed and they were randomized to one of three treatment groups or acted as controls. They attended for three visits; at baseline, HRT was initiated as oestrogen alone, oral or transdermal. At the 3 month visit, HRT with the progestagen, norethisterone, was given cyclically, continuously or transdermally until the final visit at 6 months. A separate group of women from the menopause clinic at Chelsea and Westminster Hospital were studied on oestrogen implant then on implanted oestrogen and testosterone. The outcome measures studied were the separate effects of the four regimes as compared with controls on lipoproteins, glucose, insulin, fibrinogen, factor VII and E-selectin, together with weight, waist:hip ratio and blood pressure. RESULTS: The continuous combined oestrogen-progestagen therapy had similar effects on cardiovascular risk factors as oestrogen with cyclical progestagen. All regimes lowered low-density lipoprotein cholesterol, the oral route being more potent than the parenteral; the effect of transdermal HRT was similar to the implant. Lp(a) was reduced only with the oral route. Reductions in factor VII and E-selectin were observed in both the oral and transdermal routes. There was no increase in body mass index, waist:hip ratio, blood pressure or glucose and insulin levels with any of the HRT regimes used. Systolic blood pressure was reduced with the transdermal route. CONCLUSIONS: This study supports the evidence that oestrogen-progestagen HRT, both oral and transdermal, although attenuating some of the benefit of oestrogen alone on fibrinogen and high-density lipoprotein, significantly reduces cardiovascular risk factors, which should diminish post-menopausal risk of coronary disease. PMID- 11120723 TI - Hormone replacement therapy: changes in frequency and type of prescription by Dutch GPs during the last decade of the millennium. AB - OBJECTIVE: The present study was conducted in order to determine the change of frequency and type of hormone replacement therapy (HRT) regimen newly prescribed by Dutch GPs. METHODS: A comparison was made of two data sets (multi-stage random samples) collected in 1987/88 and from 1995 to 1998 concerning women 40 years and older who were newly prescribed HRT. RESULTS: Compared with 1987/88, 50% more patients were newly prescribed HRT in 1998 (2.0 in 1987/88 and 3.0 in 1998 per 1000 registered women, P < 0.01). The age distribution remained about the same, with a peak between 50 and 54 years in each year of registration. Unopposed oestrogens (including plasters) were prescribed less frequently (1.3 per thousand in 1987/88 versus 0.7 per thousand in 1998, P < 0.001), and combinations of oestrogen and progestogen more frequently in 1998 (0. 2 per thousand in 1987/88 versus 1.8 per thousand in 1998, P < 0. 01). Sequential therapy was prescribed slightly more frequently than continuous therapy (65% sequential therapy in 1995; 55% in 1998). The most frequent reason for starting HRT in 1995-1998 was climacteric symptoms (89-98%), followed by osteoporosis prevention (16-28%) and early menopause (13-25%). Rarely were preventive goals the only reason (6%) for prescribing HRT. CONCLUSIONS: The number of HRT prescriptions increased by 50% over the last decade of the millennium. The age distribution remained the same. There was a tendency to shift from prescribing unopposed oestrogens to combinations of oestrogens and progestogens. Alleviation of climacteric symptoms was the main reason for prescribing HRT throughout the registration period. Prescription of HRT for prevention of osteoporosis and/or cardiovascular disease has so far not been adopted on a large scale by Dutch GPs. PMID- 11120724 TI - Investigation and therapy in patients with different types of dyspepsia: a 3 year follow-up study from general practice. AB - BACKGROUND: Decisions by GPs on investigation and treatment are based on the symptoms presented by the patient. The relevance of dyspepsia subgroups has been questioned, but their value in general practice has not been tested. OBJECTIVE: The aim of this study was to investigate how dividing dyspepsia into different subgroups (ulcer-like, reflux-like, dysmotility-like, uncharacteristic and relapsing dyspepsia) affected the approach of GPs to patients with dyspeptic complaints. METHODS: A random sample of GPs' patients consulting for different dyspepsia subtypes were studied by postal questionnaires 3 years after the initial consultation, obtaining information from the GPs' records on investigations, prescriptions of dyspepsia medication and gastrointestinal morbidity. RESULTS: In the 3 years studied, 48% of the patients were prescribed dyspepsia medication, 14% were endoscoped and 3% were referred to a specialist. The dyspepsia subtype was significantly related to the type of drug prescribed, but not to investigations or referrals. Ulcer-like and reflux-like dyspepsia were treated in the same way. DISCUSSION: Dyspepsia subtypes significantly influenced the treatment. Danish GPs treat all acid-related dyspepsia in the same way, and differently from other types of dyspepsia. PMID- 11120725 TI - Teenage pregnancy: whose problem is it? AB - BACKGROUND: The UK has the highest rates of teenage conception in Europe. Teenage conception has been identified in medical literature as a problem for society and teenagers. However, little attempt has been made to see it from the perspective of the teenagers themselves. OBJECTIVE: To explore teenage women's attitudes to sexual health, contraception and pregnancy. METHODS: Ethnographic qualitative study based on in-depth interviews and participant observation. The study took place in young mothers' groups, young persons' clinics and general practices in Bristol. Subjects were 34 young women between the ages of 16 and 20, sampled purposefully in two groups to include young mothers and never-pregnant young women from advantaged and disadvantaged socioeconomic backgrounds. RESULTS: The two groups did not differ in their use of contraception at first intercourse. Young women from more socioeconomically advantaged backgrounds felt that motherhood would not be acceptable to them, but were more tolerant to others who became young mothers. The pregnant/ young mothers revealed more difficulties getting access to reliable contraceptive services, and dissatisfaction with sex education in schools. The pregnant/young mothers found abortion to be less acceptable than the more socially advantaged group. Both groups reported sexual behaviour that involved risks of becoming pregnant, but the more socially advantaged group were more likely to use emergency contraception. CONCLUSIONS: The study demonstrates the importance of taking the views of young people into account when planning both sex education and the provision of contraceptive services. PMID- 11120726 TI - Men's self-assessed personal health resources: approaching patients' strong points in general practice. AB - OBJECTIVE: To explore resource-oriented, gender-sensitive approaches in general practice by identifying what men perceive to be their personal health resources. METHODS: A key question was developed to invite men to tell their GPs about personal health resources during ordinary visits. The answers of 39 consecutive male patients (aged 19-84 years) visiting two female GPs were audio taped and analysed, qualitatively inspired by Giorgi's phenomenological approach, supported by theories on salutogenesis, patient-centredness and gender perspectives. The main outcome measures were personal qualities and strategies considered by men to be their health resources. RESULTS: Men considered that the following were personal health resources: optimism, good self-esteem, job satisfaction, ability to cope with stress at work, leisure activities and relaxation with friends producing energy, and fitness and lifestyle activities. CONCLUSION: A key question can give a doctor access to men's thoughts about their strong points. Self-assessed personal health resources can be identified and mobilized by the GP and support a salutogenic approach, which contrasts with the tendency of contemporary medical practice to focus on risk. Asking people about their own ideas may reveal that coping patterns are more complex than reflected in prevailing research. PMID- 11120727 TI - Physicians' attitudes towards prevention: importance of intervention-specific barriers and physicians' health habits. AB - BACKGROUND: Several studies have explored physicians' attitudes towards prevention and barriers to the delivery of preventive health interventions. However, the relative importance of these previously identified barriers, both in general terms and in the context of a number of specific preventive interventions, has not been identified. Certain barriers may only pertain to a subset of preventive interventions. OBJECTIVES: We aimed to determine the relative importance of identified barriers to preventive interventions and to explore the association between physicians' characteristics and their attitudes towards prevention. METHODS: We conducted a cross-sectional survey of 496 of the 686 (72.3% response rate) generalist physicians from three Swiss cantons through a questionnaire asking physicians to rate the general importance of eight preventive health strategies and the relative importance of seven commonly cited barriers in relation to each specific preventive health strategy. RESULTS: The proportion of physicians rating each preventive intervention as being important varied from 76% for colorectal cancer screening to 100% for blood pressure control. Lack of time and lack of patient interest were generally considered to be important barriers by 41% and 44% of physicians, respectively, but the importance of these two barriers tended to be specifically higher for counselling based interventions. Lack of training was most notably a barrier to counselling about alcohol and nutrition. Four characteristics of physicians predicted negative attitudes toward alcohol and smoking counselling: consumption of more than three alcoholic drinks per day [odds ratio (OR) = 8.4], sedentary lifestyle (OR = 3.4), lack of national certification (OR = 2.2) and lack of awareness of their own blood pressure (OR = 2.0). CONCLUSIONS: The relative importance of specific barriers varies across preventive interventions. This points to a need for tailored practice interventions targeting the specific barriers that impede a given preventive service. The negative influence of physicians' own health behaviours indicates a need for associated population-based interventions that reduce the prevalence of high-risk behaviours in the population as a whole. PMID- 11120728 TI - Improving residents' breastfeeding assessment skills: a problem-based workshop. AB - BACKGROUND: It is well documented that residents have limited knowledge about common breastfeeding problems. OBJECTIVES: The purpose of this study was to assess whether a problem-based, interactive breastfeeding workshop would improve resident skill level. METHODS: Two groups of second- and third-year family medicine residents were assigned to an intervention or control group; both groups participated in pre-and post-intervention Objective Structured Clinical Examinations (OSCEs) and completed written questionnaires. The intervention consisted of a 4.5 hour interactive workshop with didactic presentations and opportunities to work with a lactation consultant and standardized patients trained to role-play selected breastfeeding problems. RESULTS: There were no baseline differences in knowledge or performance scores on the OSCEs between the intervention and control groups. OSCE scores after intervention were significantly better in the intervention group for the content areas assessing position and latch and the evaluation of sore nipples (P < 0.001 and P = 0.05, respectively). There was a trend towards improvement in assessment of the problem of low milk supply (P = 0.31). All residents in the intervention group correctly diagnosed the cause of both the sore nipples and low milk supply at the follow-up OSCE, with P values of <0.001 and 0.068, respectively. The intervention group felt significantly more confident in their breastfeeding problem-solving (P < 0.001). CONCLUSIONS: An interactive, problem-based workshop to teach residents the basics of breastfeeding problem solving can be implemented in residency and improve clinical diagnostic skills and residents' comfort with breastfeeding. PMID- 11120729 TI - What are the characteristics of the competent general practitioner trainer? AB - BACKGROUND: Increasing attention is being given to the training of doctors to become teachers. This does not apply only to the schooling of teachers in undergraduate medical education: at the postgraduate level, general practitioner trainers (GP-trainers) receive special schooling to prepare them for their role. Yet the skills, knowledge and traits that should be expected in the competent GP trainer have not been elucidated precisely. OBJECTIVES: The aim of this research project is to determine the traits, knowledge and skills required for a competent GP-trainer. METHOD: We used a qualitative method to answer the question. Ten focus-group meetings were held involving three Departments of Vocational Training in The Netherlands. Each group consisted of GP-trainers, GP-trainees or staff members. The transcriptions of these meetings were analysed, resulting in a description of what makes a competent GP-trainer. RESULTS: Five hundred items were obtained from the focus-group meetings, each of which was formulated in the form "A good GP-trainer is/can/knows. ", etc. These items were divided into the following categories: teaching knowledge, teaching skills, teaching attitude and personality traits of the GP-trainer. A competent GP-trainer must understand basic teaching methods and be able to apply this knowledge. The skill to give good feedback was seen as an important asset for a competent GP-trainer, as were observation skills, the skill to analyse and the skill to foster reflection in the trainee. The teaching attitude of a competent GP-trainer is characterized by giving latitude to and having respect for and interest in the trainee, and being available for consultation, while the teaching approach should be individualized. Enthusiasm, flexibility, patience and self-insight were some of the personality traits identified. CONCLUSION: Many characteristics were identified as a result of this research. The next logical step will involve a Delphi consensus procedure to obtain a profile of the competent GP-trainer. This profile will then be suitable in setting the standards for curricula for future GP-trainers. PMID- 11120730 TI - Does spinal manipulation have specific treatment effects? AB - OBJECTIVE: To investigate the question whether or not spinal manipulation is associated with specific treatment effects. METHODS: Literature searches were carried out in Medline, Embase and The Cochrane Library. All sham-controlled trials of spinal manipulation were considered. RESULTS: Seven such studies were located. Their methodological quality was variable but three trials adhered to the highest standards of scientific rigour. Collectively these data do not show therapeutic effects beyond placebo. In particular, the three most rigorous studies were negative. CONCLUSION: The few sham-controlled trials that do exist show that this methodology is, in principle, applicable also to spinal manipulation. The results available to date suggest that the therapeutic success of spinal manipulation is largely due to a placebo effect. PMID- 11120731 TI - Power and influence in clinical effectiveness and evidence-based medicine. AB - BACKGROUND: The need to base clinical interventions on valid findings of research has been a dominant theme in clinical practice during the last quarter of a century. However, there is much evidence showing that research evidence reaches everyday practice slowly. Solutions to this problem include evidence-based practice and implementation by guidelines and audit. Studies of these methods have included surveys of clinicians' views, implementation projects and evaluations of educational interventions, but they have not examined their implications for the power structure of clinical organizations. This is surprising, given the emphasis placed on medical power in sociological studies of health care. METHODS: A framework derived from management theory defines and summarizes theories of power and influence under the headings: sources of power, overt methods of influence, unseen or covert methods of influence and individual response to influence. This framework is then used to analyse the power and influence possessed and exerted by general practitioners (GPs) and hospital consultants and how these are affected by evidence-based practice and guidelines and audit programmes. OUTCOMES: GPs are seen as having less expert power than consultants and to be more compliant with externally managed guidelines and audit programmes. It is pointed out that compliance with guidelines and audit programmes helps GPs to meet their contractual requirement to be involved in clinical audit activities. Evidence-based practice, which directly challenges the authority of expert opinion is seen as a threat to the power of consultants, but a potential opportunity for GPs and other clinicians whose status is traditionally lower. PMID- 11120732 TI - Nurse practitioners in primary care. AB - OBJECTIVES: Recent policy emphasizing the role of primary care has increased the workload of general practitioners (GPs) while simultaneously placing nurse practitioners (NPs) as key providers in the delivery of health care. There is need to examine the latter's work practices. The purpose of this article is to explore the role and practice of NPs in general practice. METHODS: DESIGN: Thirty-six semi-structured interviews with GPs, NPs, receptionists and patients were analysed. SETTING: Four general practices in south-east England. MAIN OUTCOME MEASURES: Data from semi-structured interviews relating to allocation, prescribing and referral practices of NPs in primary care. RESULTS: These include the differences in presenting problems of patients seen by GPs and NPs, prescribing and referral practice and legal issues of the nurse practitioner. A wide range of practice is reported. CONCLUSION: This study highlights the variation in how patients are allocated for NP consultation and in NP autonomy, prescribing and referral, which raises issues for clinical governance of protocols and risk management. PMID- 11120734 TI - An International Forum for Family Doctors. PMID- 11120733 TI - Selections from current literature. Complementary therapies for nausea and vomiting in early pregnancy. PMID- 11120735 TI - Amino acid residues outside of the pore region contribute to N-type calcium channel permeation. AB - It is widely believed that the selectivity of voltage-dependent calcium channels is mainly controlled by amino acid residues contained within four p-loop motifs forming the pore of the channel. An examination of the amino acid sequences of high voltage-activated calcium channels reveals that their domain III S5-H5 regions contain a highly conserved motif with homology to known EF hand calcium binding proteins, hinting that this region may contribute to channel permeation. To test this hypothesis, we used site-directed mutagenesis to replace three conserved negatively charged residues in the N-type calcium channel alpha1B subunit (Glu-1321, Asp-1323, and Glu-1332) with positively charged amino acids (lysine and arginine) and studied their effect on ion selectivity using whole cell and single channel patch clamp recordings. Whereas the wild type channels conducted barium much more effectively than calcium, the mutant displayed nearly equal permeabilities for these two ions. Individual replacement of residue 1332 or a double substitution of residues 1321 and 1323 with lysine and arginine, respectively, were equally effective. Disruption of the putative EF hand motif through replacement of the central glycine residue (1326) with proline resulted in a similar effect, indicating that the responses observed with the triple mutant were not due to changes in the net charge of the channel. Overall, our data indicate that residues outside of the narrow region of the pore have the propensity to contribute to calcium channel permeation. They also raise the possibility that interactions of calcium ions with a putative calcium binding domain at the extracellular side of the channel may underlie the differential permeabilities of the channel for barium and calcium ions. PMID- 11120736 TI - Control of Mung bean pectinmethylesterase isoform activities. Influence of pH and carboxyl group distribution along the pectic chains. AB - Well-characterized pectin samples with a wide range of degrees of esterification (39-74%) were incubated with the solubilized pure alpha and gamma isoforms of pectinmethylesterase, from mung bean hypocotyl (Vigna radiata). Enzyme activity was determined at regular intervals along the deesterification pathway at pH 5.6 and pH 7.6. It has been demonstrated that the distribution of the carboxyl units along the pectin backbone controls the activity of the cell wall pectinmethylesterases to a much greater extent than the methylation degree, with a random distribution leading to the strongest activity. Polygalacturonic acid was shown to be a competitive inhibitor of the alpha isoform activity at pH 5.6 and to inhibit the gamma isoform activity at both pH 5.6 and pH 7.6. Under these conditions, the drop in enzyme activity was shown to be correlated to the formation of deesterified blocks of 19 +/- 1 galacturonic acid residues through simulations of the enzymatic digestion according to the mechanisms established previously (Catoire, L., Pierron, M., Morvan, C., Herve du Penhoat, C., and Goldberg, R. (1998) J. Biol. Chem. 273, 33150-33156). However, even in the absence of inhibition by the reaction product, activity dropped to negligible levels long before the substrate had been totally deesterified. Comparison of alpha and gamma isoform cDNAs suggests that the N-terminal region of catalytic domains might explain their subtle differences in activity revealed in this study. The role of pectinmethylesterase in the cell wall stiffening process along the growth gradient is discussed. PMID- 11120737 TI - Direct interaction between endothelial nitric-oxide synthase and dynamin-2. Implications for nitric-oxide synthase function. AB - Endothelial nitric-oxide synthase (eNOS) is regulated in part through specific protein interactions. Dynamin-2 is a large GTPase residing within similar membrane compartments as eNOS. Here we show that dynamin-2 binds directly with eNOS thereby augmenting eNOS activity. Double label confocal immunofluorescence demonstrates colocalization of eNOS and dynamin in both Clone 9 cells cotransfected with green fluorescent protein-dynamin and eNOS, as well as in bovine aortic endothelial cells (BAEC) expressing both proteins endogenously, predominantly in a Golgi membrane distribution. Immunoprecipitation of eNOS from BAEC lysate coprecipitates dynamin and, conversely, immunoprecipitation of dynamin coprecipitates eNOS. Additionally, the calcium ionophore, a reagent that promotes nitric oxide release, enhances coprecipitation of dynamin with eNOS in BAEC, suggesting the interaction between the proteins can be regulated by intracellular signals. In vitro studies demonstrate that glutathione S transferase (GST)-dynamin-2 quantitatively precipitates both purified recombinant eNOS protein as well as in vitro transcribed (35)S-labeled eNOS from solution indicating a direct interaction between the proteins in vitro. Scatchard analysis of binding studies demonstrates an equilibrium dissociation constant (K(d)) of 27.6 nm. Incubation of purified recombinant eNOS protein with GST-dynamin-2 significantly increases eNOS activity as does overexpression of dynamin-2 in ECV 304 cells stably transfected with eNOS-green fluorescent protein. These studies demonstrate a direct protein-protein interaction between eNOS and dynamin-2, thereby identifying a new NOS-associated protein and providing a novel function for dynamin. These events may have relevance for eNOS regulation and trafficking within vascular endothelium. PMID- 11120738 TI - Diatom fucoxanthin chlorophyll a/c-binding protein (FCP) and land plant light harvesting proteins use a similar pathway for thylakoid membrane Insertion. AB - The light-harvesting proteins in plastids of different lineages including algae and land plants represent a superfamily of chlorophyll-binding proteins that seem to be phylogenetically related, although some of the light-harvesting complex (LHC) proteins bind different carotenoids. LHCs can be divided into chlorophyll a/b-binding proteins found in green algae, euglenoids, and higher plants and into chlorophyll a/c-binding proteins of various algal taxa. LHC proteins from diatoms are named fucoxanthin-chlorophyll a/c-binding proteins (FCP). In contrast to chlorophyll a/b-binding proteins, there is no information so far about the way FCPs integrate into thylakoid membranes. The diatom FCP preproteins have a bipartite presequence that is necessary to enable transport into the four membrane-bound diatom plastids, but similar to chlorophyll a/b-binding proteins there is apparently no presequence present for targeting to the thylakoid membrane. By establishing an in vitro import assay for diatom thylakoids, we demonstrated that thylakoid integration of diatom FCP depends on the presence of stromal factors and GTP. This indicates that a pathway involving signal recognition particles (SRP) is involved in membrane integration just as shown for LHCs in higher plants. We also demonstrate integration of diatom FCP into thylakoids of higher plants and vice versa SRP-dependent targeting of LHCs from pea and Arabidopsis into diatom thylakoids. The similar SRP-dependent modes of thylakoid integration of land plant LHCs and FCPs support recent analyses indicating a common origin of chlorophyll a/b- and a/c-binding proteins. PMID- 11120739 TI - Association of junctional adhesion molecule with calcium/calmodulin-dependent serine protein kinase (CASK/LIN-2) in human epithelial caco-2 cells. AB - We report here that junctional adhesion molecule (JAM) interacts with calcium/calmodulin-dependent serine protein kinase (CASK), a protein related to membrane-associated guanylate kinases. In Caco-2 cells, JAM and CASK were coprecipitated and found to colocalize at intercellular contacts along the lateral surface of the plasma membrane. Association of JAM with CASK requires the PSD95/dlg/ZO-1 (PDZ) domain of CASK and the putative PDZ-binding motif Phe-Leu Val(COOH) in the cytoplasmic tail of JAM. Temporal dissociation in the junctional localization of the two proteins suggests that the association with CASK is not required for recruiting JAM to intercellular junctions. Compared with mature intercellular contacts, junction assembly was characterized by both enhanced solubility of CASK in Triton X-100 and reduced amounts of Triton-insoluble JAM CASK complexes. We propose that JAM association with CASK is modulated during junction assembly, when CASK is partially released from its cytoskeletal associations. PMID- 11120740 TI - Fructose-6-phosphate aldolase is a novel class I aldolase from Escherichia coli and is related to a novel group of bacterial transaldolases. AB - We have cloned an open reading frame from the Escherichia coli K-12 chromosome that had been assumed earlier to be a transaldolase or a transaldolase-related protein, termed MipB. Here we show that instead a novel enzyme activity, fructose 6-phosphate aldolase, is encoded by this open reading frame, which is the first report of an enzyme that catalyzes an aldol cleavage of fructose 6-phosphate from any organism. We propose the name FSA (for fructose-six phosphate aldolase; gene name fsa). The recombinant protein was purified to apparent homogeneity by anion exchange and gel permeation chromatography with a yield of 40 mg of protein from 1 liter of culture. By using electrospray tandem mass spectroscopy, a molecular weight of 22,998 per subunit was determined. From gel filtration a size of 257,000 (+/- 20,000) was calculated. The enzyme most likely forms either a decamer or dodecamer of identical subunits. The purified enzyme displayed a V(max) of 7 units mg(-)1 of protein for fructose 6-phosphate cleavage (at 30 degrees C, pH 8.5 in 50 mm glycylglycine buffer). For the aldolization reaction a V(max) of 45 units mg(-)1 of protein was found; K(m) values for the substrates were 9 mm for fructose 6-phosphate, 35 mm for dihydroxyacetone, and 0.8 mm for glyceraldehyde 3-phosphate. FSA did not utilize fructose, fructose 1-phosphate, fructose 1,6-bisphosphate, or dihydroxyacetone phosphate. FSA is not inhibited by EDTA which points to a metal-independent mode of action. The lysine 85 residue is essential for its action as its exchange to arginine (K85R) resulted in complete loss of activity in line with the assumption that the reaction mechanism involves a Schiff base formation through this lysine residue (class I aldolase). Another fsa-related gene, talC of Escherichia coli, was shown to also encode fructose-6 phosphate aldolase activity and not a transaldolase as proposed earlier. PMID- 11120741 TI - The angiogenic factors Cyr61 and connective tissue growth factor induce adhesive signaling in primary human skin fibroblasts. AB - The angiogenic inducers cysteine-rich angiogenic protein 61 (Cyr61) and connective tissue growth factor (CTGF) are structurally related, extracellular matrix-associated heparin-binding proteins. Both can stimulate chemotaxis and promote proliferation in endothelial cells and fibroblasts in culture and induce neovascularization in vivo. Encoded by inducible immediate early genes, Cyr61 and CTGF are synthesized upon growth factor stimulation in cultured fibroblasts and during cutaneous wound healing in dermal fibroblasts. Recently, we have shown that adhesion of primary human fibroblasts to immobilized Cyr61 is mediated through integrin alpha(6)beta(1) and cell surface heparan sulfate proteoglycans (HSPGs) (Chen, N., Chen, C.-C., and Lau, L.F. (2000) J. Biol. Chem. 275, 24953 24961), providing the first demonstration of an absolute requirement for HSPGs in integrin-mediated cell attachment. We show in this study that CTGF also mediates fibroblast adhesion through the same mechanism and demonstrate that fibroblasts adhesion to immobilized Cyr61 or CTGF induces distinct adhesive signaling responses consistent with their biological activities. Compared with fibroblast adhesion to fibronectin, laminin, or type I collagen, cell adhesion to Cyr61 or CTGF induces 1) more extensive and prolonged formation of filopodia and lamellipodia, concomitant with formation of integrin alpha(6)beta(1)-containing focal complexes localized at leading edges of pseudopods; 2) activation of intracellular signaling molecules including focal adhesion kinase, paxillin, and Rac with similar rapid kinetics; 3) sustained activation of p42/p44 MAPKs lasting for at least 9 h; and 4) prolonged gene expression changes including up regulation of MMP-1 (collagenase-1) and MMP-3 (stromelysin-1) mRNAs and proteins sustained for at least 24 h. Together, these results establish Cyr61 and CTGF as bona fide adhesive substrates with specific signaling capabilities, provide a molecular basis for their activities in fibroblasts through integrin alpha(6)beta(1) and HSPG-mediated signaling during attachment and indicate that these proteins may function in matrix remodeling through the activation of metalloproteinases during angiogenesis and wound healing. PMID- 11120742 TI - Active (9.6 s) and inactive (21 s) oligomers of NHE3 in microdomains of the renal brush border. AB - We have previously shown that Na(+)-H(+) exchanger isoform NHE3 exists as both 9.6 and 21 S (megalin-associated) oligomers in the renal brush border. To characterize the oligomeric forms of the renal brush border Na(+)-H(+) exchanger in more detail, we performed membrane fractionation studies. We found that similar amounts of NHE3 were present in microvilli and a nonmicrovillar membrane domain of high density (dense vesicles). Horseradish peroxidase-labeled endosomes were not prevalent in the dense membrane fraction. However, megalin, which localizes primarily to the intermicrovillar microdomain of the brush border, was enriched in the dense vesicles, implicating this microdomain as the likely source of these membranes. Immunolocalization of NHE3 confirmed that a major fraction of the transporter colocalized with megalin in the intermicrovillar region of the brush border. Immunoprecipitation studies demonstrated that in microvilli the majority of NHE3 was not bound to megalin, while in the dense vesicles most of the NHE3 coprecipitated with megalin. Moreover, sucrose velocity gradient centrifugation experiments revealed that most NHE3 in microvilli sedimented with an S value of 9.6, while the S value of NHE3 in dense vesicles was 21. Finally, we examined the functional state of NHE3 in both membrane fractions. As assayed by changes in acridine orange fluorescence, imposing an outwardly directed Na(+) gradient caused generation of an inside acid pH gradient in the microvilli, indicating Na(+)-H(+) exchange activity, but not in the dense vesicles. Taken together, these data demonstrate that renal brush border NHE3 exists in two oligomeric states: a 9.6 S active form present in microvilli and a 21 S, megalin associated, inactive form in the intermicrovillar microdomain of the apical plasma membrane. Thus, regulation of renal brush border Na(+)-H(+) exchange activity may be mediated by shifting the distribution between these forms of NHE3. PMID- 11120743 TI - Roles for beta II-protein kinase C and RACK1 in positive and negative signaling for superoxide anion generation in differentiated HL60 cells. AB - beta-Protein kinase (PKC) is essential for ligand-initiated assembly of the NADPH oxidase for generation of superoxide anion (O(2)). Neutrophils and neutrophilic HL60 cells contain both betaI and betaII-PKC, isotypes that are derived by alternate splicing. betaI-PKC-positive and betaI-PKC null HL60 cells generated equivalent amounts of O(2) in response to fMet-Leu-Phe and phorbol myristate acetate. However, antisense depletion of betaII-PKC from betaI-PKC null cells inhibited ligand-initiated O(2) generation. fMet-Leu-Phe triggered association of a cytosolic NADPH oxidase component, p47(phox), with betaII-PKC but not with RACK1, a binding protein for betaII-PKC. Thus, RACK1 was not a component of the signaling complex for NADPH oxidase assembly. Inhibition of beta-PKC/RACK1 association by an inhibitory peptide or by antisense depletion of RACK1 enhanced O(2) generation. Therefore, betaII-PKC but not betaI-PKC is essential for activation of O(2) generation and plays a positive role in signaling for NADPH oxidase activation in association with p47(phox). In contrast, RACK1 is involved in negative signaling for O(2) generation. RACK1 binds to betaII-PKC but not with the p47(phox).betaII-PKC complex. RACK1 may divert betaII-PKC to other signaling pathways requiring beta-PKC for signal transduction. Alternatively, RACK1 may sequester betaII-PKC to down-regulate O(2) generation. PMID- 11120744 TI - The role of the FRE family of plasma membrane reductases in the uptake of siderophore-iron in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae takes up siderophore-bound iron through two distinct systems, one that requires siderophore transporters of the ARN family and one that requires the high affinity ferrous iron transporter on the plasma membrane. Uptake through the plasma membrane ferrous iron transporter requires that the iron first must dissociate from the siderophore and undergo reduction to the ferrous form. FRE1 and FRE2 encode cell surface metalloreductases that are required for reduction and uptake of free ferric iron. The yeast genome contains five additional FRE1 and FRE2 homologues, four of which are regulated by iron and the major iron-dependent transcription factor, Aft1p, but whose function remains unknown. Fre3p was required for the reduction and uptake of ferrioxamine B-iron and for growth on ferrioxamine B, ferrichrome, triacetylfusarinine C, and rhodotorulic acid in the absence of Fre1p and Fre2p. By indirect immunofluorescence, Fre3p was expressed on the plasma membrane in a pattern similar to that of Fet3p, a component of the high affinity ferrous transporter. Enterobactin, a catecholate siderophore, was not a substrate for Fre3p, and reductive uptake required either Fre1p or Fre2p. Fre4p could facilitate utilization of rhodotorulic acid-iron when the siderophore was present in higher concentrations. We propose that Fre3p and Fre4p are siderophore-iron reductases and that the apparent redundancy of the FRE genes confers the capacity to utilize iron from a variety of siderophore sources. PMID- 11120745 TI - Regulation of glut1 mRNA by hypoxia-inducible factor-1. Interaction between H-ras and hypoxia. AB - Oncogenic transformation and hypoxia both induce glut1 mRNA. We studied the interaction between the ras oncogene and hypoxia in up-regulating glut1 mRNA levels using Rat1 fibroblasts transformed with H-ras (Rat1-ras). Transformation with H-ras led to a substantial increase in glut1 mRNA levels under normoxic conditions and additively increased glut1 mRNA levels in concert with hypoxia. Using a luciferase reporter construct containing 6 kilobase pairs of the glut1 promoter, we showed that this effect was mediated at the transcriptional level. Promoter activity was much higher in Rat1-ras cells than in Rat1 cells and could be down-regulated by cotransfection with a dominant negative Ras construct (RasN17). A 480-base pair (bp) cobalt/hypoxia-responsive fragment of the promoter containing a HIF-1 binding site showed significantly higher activity in Rat1-ras cells than in Rat1 cells, suggesting that Ras might mediate its effect through HIF-1 even under normoxic conditions. Consistent with this, Rat1-ras cells displayed higher levels of HIF1-alpha protein under normoxic conditions. In addition, a promoter construct containing a 4-bp mutation in the HIF1 binding site showed lower activity in Rat1-ras cells than a construct with an intact HIF1 binding site. The activity of the latter construct but not the former could be down-regulated by RasN17, supporting the importance of the HIF1 binding site in regulation by Ras. The phosphatidylinositol 3-kinase inhibitor LY29004 down regulated glut1 promoter activity and mRNA levels under normoxia and also decreased HIF1alpha protein levels in these cells. Collectively these results indicate that H-Ras up-regulates the glut1 promoter, at least in part, by increasing HIF-1alpha protein levels leading to transactivation of promoter through the HIF-1 binding site. PMID- 11120746 TI - Coupling of the insulin-like growth factor-I receptor tyrosine kinase to Gi2 in human intestinal smooth muscle: Gbetagamma -dependent mitogen-activated protein kinase activation and growth. AB - Endogenous insulin-like growth factor-1 (IGF-I) stimulates growth of cultured human intestinal smooth muscle by activating distinct mitogen-activated protein (MAP) kinase-dependent and phosphatidylinositol 3-kinase-dependent signaling pathways. In Rat1 and Balb/c3T3 fibroblasts and in neurons the IGF-I receptor is coupled to an inhibitory G protein, G(i), which mediates G(beta)gamma-dependent MAP kinase activation. The present study determined whether in normal human intestinal smooth muscle cells the IGF-I receptor activates a heterotrimeric G protein and the role of G protein activation in mediating IGF-I-induced growth. IGF-I elicited IGF-I receptor tyrosine phosphorylation, resulting in the specific activation of G(i2). G(beta)gamma subunits selectively mediated IGF-I-dependent MAP kinase activation; G(alpha)i2 subunits selectively mediated IGF-I-dependent inhibition of adenylyl cyclase activity. IGF-I-stimulated MAP kinase activation and growth were inhibited by pertussis toxin, an inhibitor of G(i)/G(o) activation. Cyclic AMP inhibits growth of human intestinal muscle cells. IGF-I inhibited both basal and forskolin-stimulated cAMP levels. This inhibition was attenuated in the presence of pertussis toxin. IGF-I stimulated phosphatidylinositol 3-kinase activation, in contrast to MAP kinase activation, occurred independently of G(i2) activation. These data suggest that IGF-I specifically activates G(i2), resulting in concurrent G(beta)gamma-dependent stimulation of MAP kinase activity and growth, and G(alpha)i2-dependent inhibition of cAMP levels resulting in disinhibition of cAMP-mediated growth suppression. PMID- 11120747 TI - A genetic approach to understanding inner ear function. PMID- 11120748 TI - Pemphigus vulgaris: the other half of the story. PMID- 11120749 TI - Making matters worse for a broken heart. PMID- 11120750 TI - PAI-1, fibrosis, and the elusive provisional fibrin matrix. PMID- 11120751 TI - Animal models of emphysema: the next generations. PMID- 11120752 TI - Targeted disruption of the Kvlqt1 gene causes deafness and gastric hyperplasia in mice. AB - The KvLQT1 gene encodes a voltage-gated potassium channel. Mutations in KvLQT1 underlie the dominantly transmitted Ward-Romano long QT syndrome, which causes cardiac arrhythmia, and the recessively transmitted Jervell and Lange-Nielsen syndrome, which causes both cardiac arrhythmia and congenital deafness. KvLQT1 is also disrupted by balanced germline chromosomal rearrangements in patients with Beckwith-Wiedemann syndrome (BWS), which causes prenatal overgrowth and cancer. Because of the diverse human disorders and organ systems affected by this gene, we developed an animal model by inactivating the murine Kvlqt1. No electrocardiographic abnormalities were observed. However, homozygous mice exhibited complete deafness, as well as circular movement and repetitive falling, suggesting imbalance. Histochemical study revealed severe anatomic disruption of the cochlear and vestibular end organs, suggesting that Kvlqt1 is essential for normal development of the inner ear. Surprisingly, homozygous mice also displayed threefold enlargement by weight of the stomach resulting from mucous neck cell hyperplasia. Finally, there were no features of BWS, suggesting that Kvlqt1 is not responsible for BWS. PMID- 11120753 TI - Severe inflammatory defect and reduced viability in CD18 and E-selectin double mutant mice. AB - CD18-deficient mice (CD18(-/-) mice) have a severe leukocyte recruitment defect in some organs, and no detectable defect in other models. Mice lacking E-selectin (CD62E(-/-) mice) have either no defect or a mild defect of neutrophil infiltration, depending on the model. CD18(-/-)CD62E(-/-), but not CD18(-/ )CD62P(-/-), mice generated by crossbreeding failed to thrive, reaching a maximum body weight of 10-15 grams. To explore the mechanisms underlying reduced viability, we investigated lethally irradiated CD62E(-/-) mice that were reconstituted with CD18(-/-) bone marrow. These mice, but not single-mutant controls, showed tenfold-increased rolling velocities in a TNF-alpha-induced model of inflammation. Leukocyte adhesion efficiency in CD18(-/-)CD62E(-/-) mice was reduced by 95%, and hematopoiesis was drastically altered, including severe bone marrow and blood neutrophilia and elevated G-CSF and GM-CSF levels. The greatly reduced viability of CD18(-/-)CD62E(-/-) mice appears to result from an inability to mount an adequate inflammatory response. Our data show that cooperation between E-selectin and CD18 integrins is necessary for neutrophil recruitment and that alternative adhesion pathways cannot compensate for the loss of these molecules. PMID- 11120754 TI - Antibodies against keratinocyte antigens other than desmogleins 1 and 3 can induce pemphigus vulgaris-like lesions. AB - Pemphigus is an autoimmune disease of skin adhesion associated with autoantibodies against a number of keratinocyte antigens, such as the adhesion molecules desmoglein (Dsg) 1 and 3 and acetylcholine receptors. The notion that anti-Dsg antibodies alone are responsible for blisters in patients with pemphigus vulgaris (PV) stems from the ability of rDsg1 and rDsg3 to absorb antibodies that cause PV-like skin blisters in neonatal mice. Here, we demonstrate that PV IgGs eluted from rDsg1-Ig-His and rDsg3-Ig-His show similar antigenic profiles, including the 38-, 43-, 115-, and 190-kDa keratinocyte proteins and a non-Dsg 3 130-kDa polypeptide present in keratinocytes from Dsg 3 knockout mouse. We injected into Dsg 3-lacking mice the PV IgGs that did not cross-react with the 160-kDa Dsg 1 or its 45-kDa immunoreactive fragment and that showed no reactivity with recombinant Dsg 1. We used both the Dsg3(null) mice with a targeted mutation of the Dsg3 gene and the "balding" Dsg3(bal)/Dsg3(bal) mice that carry a spontaneous null mutation in Dsg3. These PV IgGs caused gross skin blisters with PV-like suprabasal acantholysis and stained perilesional epidermis in a fishnet like pattern, indicating that the PV phenotype can be induced without anti-Dsg 3 antibody. The anti-Dsg 1 antibody also was not required, as its presence in PV IgG does not alter the PV-like phenotype in skin organ cultures and because pemphigus foliaceus IgGs produce a distinct phenotype in Dsg3(null) mice. Therefore, mucocutaneous lesions in PV patients could be caused by non-Dsg antibodies. PMID- 11120755 TI - TNF-alpha induces osteoclastogenesis by direct stimulation of macrophages exposed to permissive levels of RANK ligand. AB - While TNF-alpha is pivotal to the pathogenesis of inflammatory osteolysis, the means by which it recruits osteoclasts and promotes bone destruction are unknown. We find that a pure population of murine osteoclast precursors fails to undergo osteoclastogenesis when treated with TNF-alpha alone. In contrast, the cytokine dramatically stimulates differentiation in macrophages primed by less than one percent of the amount of RANKL (ligand for the receptor activator of NF-kappaB) required to induce osteoclast formation. Mirroring their synergistic effects on osteoclast differentiation, TNF-alpha and RANKL markedly potentiate NF-kappaB and stress-activated protein kinase/c-Jun NH(2)-terminal kinase activity, two signaling pathways essential for osteoclastogenesis. In vivo administration of TNF-alpha prompts robust osteoclast formation in chimeric animals in which ss galactosidase positive, TNF-responsive macrophages develop within a TNF nonresponsive stromal environment. Thus, while TNF-alpha alone does not induce osteoclastogenesis, it does so both in vitro and in vivo by directly targeting macrophages within a stromal environment that expresses permissive levels of RANKL. Given the minuscule amount of RANKL sufficient to synergize with TNF-alpha to promote osteoclastogenesis, TNF-alpha appears to be a more convenient target in arresting inflammatory osteolysis. PMID- 11120756 TI - Clearance of Alzheimer's amyloid-ss(1-40) peptide from brain by LDL receptor related protein-1 at the blood-brain barrier. AB - Elimination of amyloid-ss peptide (Ass) from the brain is poorly understood. After intracerebral microinjections in young mice, (125)I-Ass(1-40) was rapidly removed from the brain (t(1/2) 100,000 RNA copies/ml plasma. Tetramer assays were compared with three functional assays: enzyme-linked immunospot (Elispot), intracellular cytokine staining, and precursor frequency (limiting dilution assay [LDA]) cytotoxicity assays. Strong positive associations were observed between cell numbers derived by the Elispot and the tetramer assay (r = 0.90). An even stronger association between tetramer-derived numbers and intracellular cytokine staining for IFN-gamma was present (r = 0.97). The majority (median 76%) of tetramer-binding cells were consistently detectable via intracellular IFN-gamma cytokine staining. Furthermore, modifications to the LDA, using a low input cell number into each well, enabled LDAs to reach equivalence with the other methods of CTL enumeration. These data together show that functionally inert CTLs do not play a significant role in chronic pediatric or adult HIV infection. PMID- 11120779 TI - Somatic mutation of the CD95 gene in human B cells as a side-effect of the germinal center reaction. AB - Somatic hypermutation specifically modifies rearranged immunoglobulin (Ig) genes in germinal center (GC) B cells. However, the bcl-6 gene can also acquire somatic mutations during the GC reaction, indicating that certain non-Ig genes can be targeted by the somatic hypermutation machinery. The CD95 gene, implicated in negative selection of B lymphocytes in GCs, is specifically expressed by GC B cells and was recently identified as a tumor suppressor gene being frequently mutated in (post) GC B cell lymphomas. In this study, the 5' region (5'R) and/or the last exon coding for the death domain (DD) of the CD95 gene were investigated in naive, GC, and memory B cells from seven healthy donors. About 15% of GC and memory, but not naive, B cells carried mutations within the 5'R (mutation frequency 2.5 x 10(-4) per basepair). Mutations within the DD were very rare but could be efficiently selected by inducing CD95-mediated apoptosis: in 22 apoptosis-resistant cells, 12 DD mutations were found. These results indicate that human B cells can acquire somatic mutations of the CD95 gene during the GC reaction, which potentially confers apoptosis resistance and may counteract negative selection through the CD95 pathway. PMID- 11120780 TI - Suppression of signal transducer and activator of transcription 3-dependent B lymphocyte terminal differentiation by BCL-6. AB - Lymphocytes usually differentiate into effector cells within days after antigen exposure, except in germinal centers where terminal differentiation is delayed while somatic hypermutation creates high-affinity antibody mutants. Here we investigate whether arrest of terminal differentiation can be mediated by BCL-6, a transcriptional repressor that is expressed by germinal center B cells and is required for this phase of B cell development. We find that BCL-6 suppresses the differentiation of transformed and primary B cells to plasma cells by inhibiting the signal transducer and activator of transcription 3-dependent expression of the major regulator of plasma cell development, the B lymphocyte-induced maturation protein (Blimp-1). This function of BCL-6 as a repressor of B lymphocyte differentiation may also underlie the association between chromosomal translocations of its gene and B cell lymphomas. PMID- 11120781 TI - Delayed expulsion of the nematode Trichinella spiralis in mice lacking the mucosal mast cell-specific granule chymase, mouse mast cell protease-1. AB - Expulsion of gastrointestinal nematodes is associated with pronounced mucosal mast cell (MMC) hyperplasia, differentiation, and activation, accompanied by the systemic release of MMC granule chymases (chymotrypsin-like serine proteases). The beta-chymase mouse mast cell protease-1 (mMCP-1) is expressed predominantly by intraepithelial MMCs, and levels in the bloodstream and intestinal lumen are maximal at the time of worm expulsion in parasitized mice. To address the in vivo functions of MMC-specific beta-chymases, we have generated transgenic mice that lack the mMCP-1 gene. They were backcrossed onto a congenic BALB/c background to investigate the response to nematode infection. The deletion of the mMCP-1 gene is associated with significantly delayed expulsion of Trichinella spiralis and increased deposition of muscle larvae in BALB/c mice despite the presence of normal and sometimes increased numbers of MMCs. Neither worm fecundity nor worm burdens were altered in Nippostrongylus-infected mMCP-1(-/)- BALB/c mice. These data demonstrate, for the first time, that the ablation of an MMC-derived effector molecule compromises the expulsion process. PMID- 11120782 TI - Will the making of plasmacytoid dendritic cells in vitro help unravel their mysteries? PMID- 11120783 TI - Cutting edge: C-C chemokine receptor 6 is essential for arrest of a subset of memory T cells on activated dermal microvascular endothelial cells under physiologic flow conditions in vitro. AB - Memory T cells (mTC) express multiple chemokine receptors (including CCR4 and CCR6) that may potentially be involved in their arrest on inflamed endothelia. Herein, we specifically addressed whether CCR6 is required for mTC to arrest on TNF-alpha-activated human dermal microvascular endothelial cells (HDMEC) in vitro under shear stress conditions. Recombinant liver and activation-regulated chemokine (LARC)/CCL20 (a CCR6 ligand) induced firm arrest of cutaneous lymphocyte Ag(+) mTC in a flow chamber system using purified substrates. Strikingly, desensitization of CCR6 with LARC, but not thymus and activation regulated chemokine/CCL17 or secondary lymphoid tissue chemokine/CCL21, caused a 50-75% decrease (p < 0. 001) in arrest of mTC on HDMEC, which was indistinguishable from the reduction observed when total mTC were treated with pertussis toxin (p > 0.5). CCR6-depleted mTC also had a markedly reduced ability to arrest on HDMEC. Our results suggest that LARC production by activated endothelial cells and CCR6 expression by mTC may be critical components in the pertussis toxin-sensitive arrest of mTC on activated HDMEC. PMID- 11120784 TI - Cutting edge: naturally occurring soluble form of mouse Toll-like receptor 4 inhibits lipopolysaccharide signaling. AB - Toll-like receptors (TLRs) are a family of proteins playing important roles in host defense. Mice defective of functional TLR4 are hyporesponsive to LPS, suggesting that TLR4 is essential for LPS signaling. Here we report the cloning of an alternatively spliced mouse TLR4 (mTLR4) mRNA. The additional exon exists between the second and third exon of the reported mTLR4 gene and contains an in frame stop codon. The alternatively spliced mRNA encodes 86 aa of the reported mTLR4 and an additional 36 aa. This alternatively spliced mTLR4 mRNA expressed a partially secretary 20-kDa protein, which we named soluble mTLR4 (smTLR4). In a mouse macrophage cell line, the exogenously expressed smTLR4 significantly inhibited LPS-mediated TNF-alpha production and NF-kappaB activation. Additionally, in mouse macrophages, LPS increased the mRNA for smTLR4. Taken together, our results indicate that smTLR4 may function as a feedback mechanism to inhibit the excessive LPS responses in mouse macrophages. PMID- 11120785 TI - Cutting edge: chromatin remodeling at the IL-4/IL-13 intergenic regulatory region for Th2-specific cytokine gene cluster. AB - During the differentiation of naive Th cells into Th2 effector cells, the entire IL-4/IL-13 locus is remodeled into an accessible chromatin conformation. Here we show that ectopic expression and activation of Stat6 or GATA-3 in Th cells developing under Th1-polarizing conditions lead to the induction of chromatin remodeling not only at the flanking regions of the IL-4 and IL-13 genes but also at the IL-4/IL-13 intergenic regulatory region for the IL-4/IL-13/IL-5 gene cluster. Furthermore, we demonstrate that GATA-3 and another Th2-specific, inducible protein complex interact with the IL-4/IL-13 intergenic DNase I hypersensitive region specifically in Th2 cells. PMID- 11120786 TI - CDK4 expression and activity are required for cytokine responsiveness in T cells. AB - Stimulation of lymphocytes through the Ag receptor can lead to cytokine responsiveness or unresponsiveness. We examined the importance of cyclin dependent kinase (CDK)4 to establish and maintain IL-2 responsiveness in human T cells. Our results show that a herbimycin A- and staurosporine-sensitive phase of CDK4 expression and activity preceded the acquisition of IL-2-responsiveness in mitogen-stimulated peripheral blood T cells. Intriguingly, CDK4 expression and activity were demonstrable in purified unstimulated peripheral blood T cells from approximately 30% (5/16) of healthy individuals examined for this study. These T cells proliferated in response to IL-2 without additional mitogens, and both the expression and activity of CDK4 and the ability to respond to cytokines were resistant to herbimycin A and staurosporine. The pattern of CDK4 expression and response to IL-2 in this subset of individuals resembled that seen in the human IL-2-dependent Kit-225 T cell line. However, in contrast to normal T cells, Kit 225 cells were rendered unresponsive to IL-2 by stimulation through the Ag receptor. In these cells, PHA, anti-CD3, or PMA induced marked reductions of CDK4 expression and activity that paralleled IL-2 unresponsiveness, and these effects were not reversible by IL-2. Furthermore, IL-2-dependent proliferation could be similarly inhibited in Kit-225 cells by overexpression of the CDK inhibitors p16/Ink4-a or p21/Waf-1a or by overexpression of a kinase-inactive CDK4 mutant. The data indicate that CDK4 expression and activity are necessary to induce and maintain cytokine responsiveness in T cells, suggesting that CDK4 is important to link T cell signaling pathways to the machinery that controls cell cycle progression. PMID- 11120787 TI - CD30 shedding from Karpas 299 lymphoma cells is mediated by TNF-alpha-converting enzyme. AB - CD30 is a costimulatory receptor on activated lymphocytes and a number of human lymphoma cells. Specific ligation of membrane-bound CD30 or cellular stimulation by PMA results in a metalloproteinase-mediated down-regulation of CD30 and release of its soluble ectodomain (sCD30). In this report, it is demonstrated that PMA-induced CD30 cleavage from Karpas 299 cells was mediated by a membrane anchored metalloproteinase which was active on intact cells following 3-[(3 cholamidopropyl)dimethylammonio]-1-propanesulfonate extraction of membrane preparations. Moreover, CD30 shedding was blocked by the synthetic hydroxamic acid-based metalloproteinase inhibitor BB-2116 (IC(50), 230 nM) and the natural tissue inhibitor of metalloproteinases (TIMP)-3 (IC(50), 30 nM), but not by the matrix metalloproteinase inhibitors TIMP-1 and TIMP-2. This inhibition profile is similar to that of the TNF-alpha- converting enzyme (TACE) and, indeed, mRNA transcripts of the membrane-bound metalloproteinase-disintegrin TACE could be detected in Karpas 299 cells. The ectodomain of TACE was expressed in bacteria as a GST fusion protein (GST-TACE) which cleaved CD30 from the surface of Karpas 299 cells and concomitantly increased the level of sCD30 in the cell supernatants. Hence, TACE does not only control the release of TNF-alpha, but also that of sCD30. PMID- 11120788 TI - MHC recognition in thymic development: distinct, parallel pathways for survival and lineage commitment. AB - The molecular events triggered by MHC recognition and how they lead to the emergence of mature CD4 and CD8 lineage thymocytes are not yet understood. To address these questions, we have examined what signals are necessary to drive the development of CD8 lineage thymocytes in TCRalpha(-) mice in which TCR/MHC engagement cannot occur. We find that the combination of constitutive Notch activity and constitutive Bcl-2 expression are necessary and sufficient to allow the appearance of mature CD8 lineage thymocytes in TCRalpha(-) mice. In addition, Notch activity alone in TCRalpha(-) mice can induce the up-regulation of HES1, suggesting that thymocytes are competent to respond to Notch signaling in the absence of MHC recognition. These data indicate that survival and lineage commitment represent distinct, parallel pathways that occur as a consequence of MHC recognition, both of which are necessary for the development of mature CD8 lineage T cells. PMID- 11120789 TI - Intestinal CD8 alpha alpha and CD8 alpha beta intraepithelial lymphocytes are thymus derived and exhibit subtle differences in TCR beta repertoires. AB - Intraepithelial lymphocytes (IEL) of the small intestine are anatomically positioned to be in the first line of cellular defense against enteric pathogens. Therefore, determining the origin of these cells has important implications for the mechanisms of T cell maturation and repertoire selection. Recent evidence suggests that murine CD8 alpha alpha intestinal IELs (iIELs) can mature and undergo selection in the absence of a thymus. We analyzed IEL origin by cell transfer, using two congenic chicken strains. Embryonic day 14 and adult thymocytes did not contain any detectable CD8 alpha alpha T cells. However, when TCR(+) thymocytes were injected into congenic animals, they migrated to the gut and developed into CD8alphaalpha iIELs, while TCR(-) T cell progenitors did not. The TCR V beta 1 repertoire of CD8 alpha alpha(+) TCR V beta 1(+) iIELs contained only part of the TCR V beta 1 repertoire of total iIELs, and it exhibited no new members compared with CD8(+) T cells in the thymus. This indicated that these T cells emigrated from the thymus at an early stage in their developmental process. In conclusion, we show that while CD8 alpha alpha iIELs originate in the thymus, T cells acquire the expression of CD8 alpha alpha homodimers in the gut microenvironment. PMID- 11120790 TI - Immortalized myeloid suppressor cells trigger apoptosis in antigen-activated T lymphocytes. AB - We described a generalized suppression of CTL anamnestic responses that occurred in mice bearing large tumor nodules or immunized with powerful recombinant viral immunogens. Immune suppression entirely depended on GM-CSF-driven accumulation of CD11b(+)/Gr-1(+) myeloid suppressor cells (MSC) in secondary lymphoid organs. To further investigate the nature and properties of MSC, we immortalized CD11b(+)/Gr 1(+) cells isolated from the spleens of immunosuppressed mice, using a retrovirus encoding the v-myc and v-raf oncogenes. Immortalized cells expressed monocyte/macrophage markers (CD11b, F4/80, CD86, CD11c), but they differed from previously characterized macrophage lines in their capacities to inhibit T lymphocyte activation. Two MSC lines, MSC-1 and MSC-2, were selected based upon their abilities to inhibit Ag-specific proliferative and functional CTL responses. MSC-1 line was constitutively inhibitory, while suppressive functions of MSC-2 line were stimulated by exposure to the cytokine IL-4. Both MSC lines triggered the apoptotic cascade in Ag-activated T lymphocytes by a mechanism requiring cell-cell contact. Some well-known membrane molecules involved in the activation of apoptotic pathways (e.g., TNF-related apoptosis-inducing ligand, Fas ligand, TNF-alpha) were ruled out as candidate effectors for the suppression mechanism. The immortalized myeloid lines represent a novel, useful tool to shed light on the molecules involved in the differentiation of myeloid-related suppressors as well as in the inhibitory pathway they use to control T lymphocyte activation. PMID- 11120791 TI - Reversal of CD8+ T cell ignorance and induction of anti-tumor immunity by peptide pulsed APC. AB - In the present report, we have studied the potential of naive and activated effector CD8(+) T cells to function as anti-tumor T cells to a solid tumor using OVA-specific T cells from TCR-transgenic OT-I mice. Adoptive transfer of naive OT I T cells into tumor-bearing syngeneic mice did not inhibit tumor cell growth. The adoptively transferred OT-I T cells did not proliferate in lymphoid tissue of tumor-bearing mice and were not anergized by the tumor. In contrast, adoptive transfer of preactivated OT-I CTL inhibited tumor growth in a dose-dependent manner, indicating that E.G7 was susceptible to immune effector cells. Importantly, naive OT-I T cells proliferated and elicited an anti-tumor response if they were adoptively transferred into normal or CD4-deficient mice that were then vaccinated with GM-CSF-induced bone marrow-derived OVA-pulsed APC. Collectively, these data indicate that even though naive tumor-specific T cells are present at a relatively high fraction they remain ignorant of the tumor and demonstrate that a CD8-mediated anti-tumor response can be induced by Ag-pulsed APC without CD4 T cell help. PMID- 11120792 TI - Targeting weak antigens to CD64 elicits potent humoral responses in human CD64 transgenic mice. AB - Previous studies have documented that targeting foreign Ags to IgG FcgammaR leads to enhanced Ag-specific responses in vitro and in vivo. However, the ability to overcome immunologic nonresponsiveness by targeting poorly immunogenic Ags to FcgammaR has not been investigated. To address this question in a simple model, we immunized transgenic mice expressing human CD64 (FcgammaRI) and their nontransgenic littermates with Fab' derived from the murine anti-human CD64 mAb m22. The m22 Fab' served as both the targeting molecule and the Ag. We found that only CD64-expressing mice developed anti-Id titers to m22. Furthermore, chemically linked multimers of m22 Fab', which mediated efficient internalization of the human CD64, were significantly more potent than monomeric m22 F(ab')(2) at inducing anti-Id responses. In all cases, the humoral responses were specific for m22 Id and did not react with other murine IgG1 Fab' fragments. Chemical addition of a second murine Fab' (520C9 anti-human HER2/neu) to m22 Fab' multimers demonstrated that IgG1 and IgG2a anti-Id titers could be generated to 520C9 only in the CD64-expressing mice. These results show that targeting to CD64 can overcome immunological nonresponsiveness to a weak immunogen. Therefore, targeting to CD64 may be an effective method to enhance the activity of nonimmunogenic tumor vaccines. PMID- 11120793 TI - Dendritic cell migration controlled by alpha 1b-adrenergic receptors. AB - Dendritic cells (DC) bring Ags into lymphoid organs via lymphatic vessels. In this study, we investigated the possibility that the sympathetic neurotransmitter norepinephrine (NE) influences DC migration. Murine epidermal Langerhans cells mobilization is enhanced by systemic treatment with the alpha(2)-adrenergic antagonist yohimbine and inhibited by local treatment with the specific alpha(1) adrenergic antagonist prazosin (PRA). Consistently, NE enhances spontaneous emigration of DC from ear skin explants, and PRA inhibits this effect. In addition, local treatment with PRA during sensitization with FITC inhibits the contact hypersensitivity response 6 days later. In vitro, bone marrow-derived immature, but not CD40-stimulated mature DC migrate in response to NE, and this effect is neutralized by PRA. NE seems to exert both a chemotactic and chemokinetic activity on immature DC. Coherently, immature, but not mature DC, express mRNA coding for the alpha(1b)-adrenergic receptor subtype. Inactivation of this adrenergic receptor by the specific and irreversible antagonist chloroethylclonidine hinders the migration of injected DC from the footpad to regional lymph nodes. Thus, besides regulating lymph flow, the sympathetic innervation of lymphatic vessels may participate in directing DC migration from the site of inflammation to regional lymph nodes. Alternatively, the chemokinetic activity of NE may enhance the ability of DC to sample local Ags, and hence increase the number of DC migrating to the draining lymph nodes. This finding might improve our understanding of the biological basis of skin diseases and allergic reactions, and opens new pharmacological possibilities to modulate the immune response. PMID- 11120795 TI - Resistance to TNF-induced cytotoxicity correlates with an abnormal cleavage of cytosolic phospholipase A2. AB - To investigate the mechanism underlying the absence of arachidonic acid (AA) release by TNF in TNF-resistant cells, we first performed comparative analysis of phospholipid pools in both TNF-sensitive (MCF7) and their equivalent resistant cells (C1001). Quantification and incorporation studies of [(3)H]AA indicated that TNF-resistant cells were not depleted in AA. Furthermore, distribution of this fatty acid in different phospholipid pools was similar in both sensitive cells and their resistant counterparts, ruling out a defect in phospholipid pools. Since phospholipase A(2) (PLA(2)) are the main enzymes releasing free AA, we investigated their relative contribution in the acquisition of cell resistance to TNF-induced cell death and AA release. For this purpose, we used two PLA(2) inhibitors, methylarachidonyl fluorophosphate (MAFP) and bromoenol lactone (BEL), which selectively and irreversibly inhibit the cytosolic PLA(2) (cPLA(2)) and the Ca(2+)-independent PLA(2), respectively. Although a significant inhibitory effect of MAFP on both TNF-induced AA release and PLA(2) activity in MCF7 was observed, BEL had no effect. The inhibitory effect of MAFP on cPLA(2) activity correlated with an inhibition of TNF-induced cell death. Western blot analysis revealed that TNF induced a differential cleavage of cPLA(2) in TNF-sensitive vs TNF-resistant cells. Although the p70 (70-kDa) form of cPLA(2) was specifically increased in TNF-sensitive cells, a cleaved form, p50 (50 kDa), was selectively observed in TNF-resistant C1001 cells in the presence or absence of TNF. These findings suggest that the acquisition of cell resistance to this cytokine may involve an abnormal cPLA(2) cleavage. PMID- 11120794 TI - Early Th1 response in unprimed nonobese diabetic mice to the tyrosine phosphatase like insulinoma-associated protein 2, an autoantigen in type 1 diabetes. AB - The insulinoma-associated protein 2 (IA-2) is a phosphatase-like autoantigen inducing T and B cell responses associated with human insulin-dependent diabetes mellitus (IDDM). We now report that T cell responses to IA-2 can also be detected in the nonobese diabetic (NOD) mouse, a model of human IDDM. Cytokine secretion in response to purified mouse rIA-2, characterized by high IFN-gamma and relatively low IL-10 and IL-6 secretion, was elicited in spleen cells from unprimed NOD mice. Conversely, no response to IA-2 was induced in spleen cells from BALB/c, C57BL/6, or Biozzi AB/H mice that express, like NOD, the I-A(g7) class II molecule, but are not susceptible to spontaneous IDDM. The IA-2-induced IFN-gamma response in NOD spleen cells could already be detected at 3 wk and peaked at 8 wk of age, whereas the IL-10 secretion was maximal at 4 wk of age and then waned. IA-2-dependent IFN-gamma secretion was induced in CD4(+) cells from spleen as well as pancreatic and mesenteric lymph nodes. It required Ag presentation by I-A(g7) molecules and engagement of the CD4 coreceptor. Interestingly, cytokines were produced in the absence of cell proliferation and IL-2 secretion. The biological relevance of the response to IA-2 is indicated by the enhanced IDDM following a single injection of the recombinant protein emulsified in IFA into 18-day-old NOD mice. In addition, IFN-gamma production in response to IA-2 and IDDM acceleration could be induced by IL-12 administration to 12-day-old NOD mice. These results identify IA-2 as an early T cell-inducing autoantigen in the NOD mouse and indicate a role for the IA-2-induced Th1 cell response in IDDM pathogenesis. PMID- 11120796 TI - The development, maturation, and turnover rate of mouse spleen dendritic cell populations. AB - Three distinct subtypes of dendritic cells (DC) are present in mouse spleen, separable as CD4(-)8alpha(-), CD4(+)8alpha(-), and CD4(-)8alpha(+) DC. We have tested whether these represent stages of development or activation within one DC lineage, or whether they represent separate DC lineages. All three DC subtypes appear relatively mature by many criteria, but all retain a capacity to phagocytose particulate material in vivo. Although further maturation or activation could be induced by bacterially derived stimuli, phagocytic capacity was retained, and no DC subtype was converted to the other. Continuous elimination of CD4(+)8(-) DC by Ab depletion had no effect on the levels of the other DC subtypes. Bromodeoxyuridine labeling experiments indicated that all three DC subtypes have a rapid turnover (half-life, 1.5-2.9 days) in the spleen, with none being the precursor of another. The three DC subtypes showed different kinetics of development from bone marrow precursors. The CD8alpha(+) spleen DC, apparently the most mature, displayed an extremely rapid turnover based on bromodeoxyuridine uptake and the fastest generation from bone marrow precursors. In conclusion, the three splenic DC subtypes behave as rapidly turning over products of three independent developmental streams. PMID- 11120797 TI - Exposure to cigarette tar inhibits ribonucleotide reductase and blocks lymphocyte proliferation. AB - Cigarette smoking causes profound suppression of pulmonary T cell responses, which has been associated with increased susceptibility to respiratory tract infections and decreased tumor surveillance. Exposure of human T cells to cigarette tar or its major phenolic components, hydroquinone and catechol, causes an immediate cessation of DNA synthesis without cytotoxicity. However, little is known of the mechanisms by which this phenomenon occurs. In this report we demonstrate that hydroquinone and catechol inhibit lymphocyte proliferation by quenching the essential tyrosyl radical in the M2 subunit of ribonucleotide reductase. PMID- 11120798 TI - HLA-B2702 (77-83/83-77) peptide binds to beta-tubulin on human NK cells and blocks their cytotoxic capacity. AB - It has been described that peptides derived from a highly conserved region of the alpha1 helix of the first domain of HLA class I Ags exhibit immunomodulatory capacity blocking both T and NK cell cytotoxicity. In vivo treatment with these peptides prolongs survival of MHC-mismatched allografts. However, the molecular bases of these effects are still unclear. In this study, we further analyze the mechanisms by which the dimeric peptide HLA-B2702 (77-83/83-77) induces suppression of NK cell cytotoxicity. This peptide inhibits natural and redirected lysis mediated by NK cells without significantly affecting effector-target cell binding. We have also isolated and sequenced a protein that binds this inhibitory peptide, which structurally corresponds to beta-tubulin. Tubulin is the major protein of microtubules and is involved in target cell killing. Furthermore, B2702 peptide promotes GTP-independent tubulin assembly, producing aggregates that cannot be depolymerized by cold. Treatment of NK cells with Taxol or demecolcine, which interfere with microtubule organization, also prevents NK cell cytotoxicity. Taken together, these results support the hypothesis that the peptide B2702 (77-83/83-77) exerts its inhibitory effect on NK cell cytotoxicity by inducing polymerization of microtubules and interfering with their normal assembly/disassembly dynamics. PMID- 11120799 TI - CD4+ Th1 and CD8+ type 1 cytotoxic T cells both play a crucial role in the full development of contact hypersensitivity. AB - The role of CD4(+) vs CD8(+) T cells in contact hypersensitivity (CHS) remains controversial. In this study, we used gene knockout (KO) mice deficient in CD4(+) or CD8(+) T cells to directly address this issue. Mice lacking either CD4(+) or CD8(+) T cells demonstrated depressed CHS responses to dinitrofluorobenzene and oxazolone compared with wild-type C57BL/6 mice. The depression of CHS was more significant in CD8 KO mice than in CD4 KO mice. Furthermore, in vivo depletion of either CD8(+) T cells from CD4 KO mice or CD4(+) T cells from CD8 KO mice virtually abolished CHS responses. Lymph node cells (LNCs) from hapten-sensitized CD4 and CD8 KO mice showed a decreased capacity for transferring CHS. In vitro depletion of either CD4(+) T cells from CD8 KO LNCs or CD8(+) T cells from CD4 KO LNCs resulted in a complete loss of CHS transfer. LNCs from CD4 and CD8 KO mice produced significant amounts of IFN-gamma, indicating that both CD4(+) and CD8(+) T cells are able to secrete IFN-gamma. LNCs from CD8, but not CD4, KO mice were able to produce IL-4 and IL-10, suggesting that IL-4 and IL-10 are mainly derived from CD4(+) T cells. Intracellular cytokine staining of LNCs confirmed that IFN gamma-positive cells consisted of CD4(+) (Th1) and CD8(+) (type 1 cytotoxic T) T cells, whereas IL-10-positive cells were exclusively CD4(+) (Th2) T cells. Collectively, these results suggest that both CD4(+) Th1 and CD8(+) type 1 cytotoxic T cells are crucial effector cells in CHS responses to dinitrofluorobenzene and oxazolone in C57BL/6 mice. PMID- 11120800 TI - Functional equivalency of B7-1 and B7-2 for costimulating plasmid DNA vaccine elicited CTL responses. AB - A costimulatory signal in addition to an Ag-specific stimulus is required for optimal activation of T lymphocytes. CD28, the primary positive costimulatory receptor on T cells, has two identified ligands, B7-1 and B7-2. Whether B7-1 and B7-2 have identical, overlapping, or distinct functions remains unresolved. In this study, we show that mice lacking B7-2 were unable to generate CTL responses following immunization with a plasmid DNA vaccine. The ability of these B7-2 deficient mice to generate CTL responses following plasmid gp120 DNA vaccination was fully reconstituted by coadministering either a plasmid expressing B7-2 or B7 1. Moreover, the ability to generate CTL responses following plasmid DNA vaccination in mice lacking both B7-1 and B7-2 could be reconstituted by administering either plasmid B7-1 or plasmid B7-2 with the vaccine construct. These data demonstrate that either B7-1 or B7-2 administered concurrently with a plasmid DNA vaccine can fully costimulate vaccine-elicited CTL responses. Functional differences between B7-1 and B7-2 observed in vivo therefore may not reflect inherent differences in the interactions of CD28 with these ligands. PMID- 11120801 TI - Ig light chain receptor editing in anergic B cells. AB - Receptor editing in the bone marrow (BM) serves to modify the Ag receptor specificity of immature self-reactive B cells, while anergy functionally silences self-reactive clones. Here, we demonstrate that anergic B cells in hen egg lysozyme Ig (HEL-Ig)/soluble HEL double transgenic mice show evidence of having undergone receptor editing in vivo, as demonstrated by the presence of elevated levels of endogenous kappa light chain rearrangements in the BM and spleen. In an in vitro IL-7-driven BM culture system, HEL-Ig BM B cells grown in the presence of soluble HEL down-regulated surface IgM expression and also showed induction of new endogenous kappa light chain rearrangements. Using a panel of soluble protein ligands with reduced affinity for the HEL-Ig receptor, the editing response was shown to correlate in a dose-dependent fashion with the strength of signaling through the B cell receptor. The finding that the level of B cell receptor cross linking sufficient to induce anergy in B cells is also capable of engaging the machinery required for receptor editing suggests an intimate relationship between these two mechanisms in maintaining B cell tolerance. PMID- 11120803 TI - Ligation of CD27 on murine B cells responding to T-dependent and T-independent stimuli inhibits the generation of plasma cells. AB - B cells can be stimulated either allogenically with the Th cell clone D10G4.1 and bone marrow-derived dendritic cells or polyclonally with LPS to proliferate and undergo terminal differentiation to Ig-secreting plasma cells in vitro. The addition of anti-CD27 to such cultures inhibits Ig secretion, and inhibition is more marked in T-dependent cultures than in T-independent cultures. Both IgM and secondary isotypes are affected, and addition of anti-CD27 even 4 days after culture initiation inhibits Ig secretion. Anti-CD27 does not affect B cell proliferation or the acquisition of activation markers by B cells, and no marked loss of B cell viability is detected in cells cultured in the presence of anti CD27, suggesting that the inhibition of Ig secretion is not due to inhibition of early activation events or to death of activated cells in vitro. However, the presence of anti-CD27 significantly inhibits the induction of Blimp-1 and J chain transcripts, which are turned on in cells committed to plasma cell differentiation. Furthermore, mice immunized under cover of anti-CD27 make less Ag-specific IgM and IgG, but have equivalent T cell responses when compared with control mice. These data suggest that ligation of CD27, a member of the TNFR family, on the B cell surface may prevent terminal differentiation of activated B cells into Ig-secreting plasma cells. PMID- 11120802 TI - Stat4 regulates multiple components of IFN-gamma-inducing signaling pathways. AB - Stat4 is activated in response to IL-12. Most functions of IL-12, including the induction of IFN-gamma, are compromised in the absence of Stat4. Since the precise role of Stat4 in IFN-gamma induction has not been established, experiments were conducted to examine Stat4 activation of IFN-gamma and other genes required for cytokine-induced expression of IFN-gamma. We first examined IL 12 signaling components. Basal expression of IL-12Rss1 and IL-12Rss2 is decreased in Stat4-deficient cells compared with that in control cells. However, IL-12 was still capable of inducing equivalent phosphorylation of Jak2 and Tyk2 in wild type and Stat4-deficient activated T cells. We have further determined that other cytokine signaling pathways that induce IFN-gamma production are defective in the absence of Stat4. IL-18 induces minimal IFN-gamma production from Stat4-deficient activated T cells compared with control cells. This is due to defective IL-18 signaling, which results from the lack of IL-12-induced, and Stat4-dependent, expression of the IL-18R. Following IL-12 pretreatment to induce IL-18R, wild type, but not Stat4-deficient, activated T cells demonstrated IL-18-induced NF kappaB DNA-binding activity. In addition, IL-12-pretreated Stat4-deficient activated T cells have minimal IFN-gamma production followed by stimulation with IL-18 alone or in combination with IL-12 compared with control cells. Thus, Stat4 activation by IL-12 is required for the function of multiple cytokine pathways that result in induction of IFN-gamma. PMID- 11120804 TI - Diminution of the AML1 transcription factor function causes differential effects on the fates of CD4 and CD8 single-positive T cells. AB - In the thymic cortex, T lymphocytes are positively selected to survive and committed either to the CD4 single-positive (SP) or the CD8 SP lineage. The SP cells then pass through a step of maturation in the medulla and are delivered to peripheral lymphoid tissues. We examined the role of AML1, the gene encoding a transcription factor, in the above processes by using the transgenic mice expressing a dominant interfering form of AML1 as well as mice targeted heterozygously for AML1. One phenotypic change seen in the AML1-diminished mice was the reduction in the numbers of both CD4 SP and CD8 SP thymocytes, reflecting the partial impairment of the transition from the double-positive to SP stage. In addition, distinct from the above abnormality, perturbed were several aspects of SP cells, including the maturation of SP thymocytes, the recent thymic emigration, and the proliferative responsiveness of peripheral T cells to TCR stimulation. Interestingly, the AML1 diminution caused inhibitory and enhancing effects on the CD4 SP and CD8 SP cells, respectively. These differential effects are most likely related to the reduction in the peripheral CD4 SP/CD8 SP ratio observed in the AML1-diminished mice. The AML1 transcription factor thus maintains the homeostasis of each SP subset by functioning at the later stages of T lymphocyte differentiation. PMID- 11120805 TI - Genetic analyses of NFKB1 and OCA-B function: defects in B cells, serum IgM level, and antibody responses in Nfkb1-/-Oca-b-/- mice. AB - Defined patterns of gene expression during cell differentiation are likely to be ensured by multiple factors playing redundant roles. By generating mice deficient in both NFKB1 and OCA-B, we show here that the two transcription factors are required for B-1 cell differentiation and serum IgM production. In addition, relative to Nfkb1(-/-) or Oca-b(-/-) mice, the Nfkb1(-/-)Oca-b(-/-) mice show a decrease in conventional B cell frequencies in the spleen and augmented reductions in T-independent and T-dependent Ab responses. These results suggest that NFKB1 and OCA-B play compensatory roles in multiple aspects of B cell differentiation. PMID- 11120806 TI - Early programming of T cell populations responding to bacterial infection. AB - The duration of infection and the quantity of Ag presented in vivo are commonly assumed to influence, if not determine, the magnitude of T cell responses. Although the cessation of in vivo T cell expansion coincides with bacterial clearance in mice infected with Listeria monocytogenes, closer analysis suggests that control of T cell expansion and contraction is more complex. In this report, we show that the magnitude and kinetics of Ag-specific T cell responses are determined during the first day of bacterial infection. Expansion of Ag-specific T lymphocyte populations and generation of T cell memory are independent of the duration and severity of in vivo bacterial infection. Our studies indicate that the Ag-specific T cell response to L. monocytogenes is programmed before the peak of the innate inflammatory response and in vivo bacterial replication. PMID- 11120807 TI - B7 requirements for primary and secondary protein- and polysaccharide-specific Ig isotype responses to Streptococcus pneumoniae. AB - The requirements for B7 costimulation during an in vivo humoral response to an intact extracellular bacteria have not been reported. In this study we immunized mice with Streptococcus pneumoniae (R36A) to determine the B7 requirements for induction of Ig, specific for two determinants on R36A, the phosphorylcholine (PC) determinant of C-polysaccharide and pneumococcal surface protein A (PspA). We show that the primary anti-PspA response, the development of PspA-specific memory, and the induction of the secondary anti-PspA response in primed mice were completely dependent upon B7 costimulation. Of note, costimulation was required only briefly after the secondary immunization compared with after the primary immunization for optimal induction of Ig. Blockade of B7 costimulation at the time of secondary immunization also completely abrogated the established state of memory, but did not induce tolerance. In contrast to the anti-PspA response, the primary anti-PC response involved only a very short period of B7 costimulation. Whereas B7-2 alone was required for induction of the primary anti-PspA and anti PC responses, a redundant role for B7-1 and B7-2 was noted for the PspA-specific secondary response. CTLA4Ig blocked both the anti-PC and anti-PspA responses equally well over a wide range of bacterial doses. These studies demonstrate a critical, but variable, role for B7-dependent costimulation during an Ig response to an extracellular bacteria. PMID- 11120808 TI - Genetic characterization of strain differences in the ability to mediate CD40/CD28-independent rejection of skin allografts. AB - Simultaneous blockade of the CD40 and CD28 T cell costimulatory pathways effectively promotes skin allograft survival in C3H/HeJ mice, extending median survival times (MSTs) beyond 100 days. This strategy is markedly less effective in C57BL/6 mice, with MSTs ranging between 20 and 30 days. In this study, we investigate the underlying genetic causes of these distinct phenotypes. Using H-2 congenic mice, we show that the genetic basis for the varied responses between these two strains is independent of the H-2 locus and T cell precursor frequency. C57BL/6 mice treated with costimulation blockade are able to generate allospecific CTL- and IFN-gamma-producing T cells within 3-4 wk posttransplant, whereas mice with a C3H background generate neither CTL- nor IFN-gamma-producing cells. Thus, differences appear to be in the generation of the immune response and not T cell homing. Strain differences in costimulation blockade-induced hyporesponsiveness persist in the absence of CD4(+) T cells, implying a direct effect on CD8(+) T cells. We demonstrate that genetic differences are important in cells of hemopoietic origin and that the costimulation blockade-resistant phenotype is dominant. Analysis of BXH recombinant inbred strains indicates that multiple loci contribute to the phenotype, and that the blockade resistance loci are preliminarily linked to 17 markers on four chromosomes. We conclude that strain variation in allograft MSTs following CD40/CD28 blockade results from the ability of CD8(+) T cells in some strains to use alternative modes of costimulation to mount an effective alloresponse. PMID- 11120809 TI - Reactivity and regulatory properties of human anti-idiotypic antibodies induced by T cell vaccination. AB - Immunization with irradiated autoreactive T cells (T cell vaccination) induces anti-idiotypic T cell responses that preferentially recognize complementarity determining region 3 sequences, contributing to clonal depletion of autoreactive T cells. However, it remains unknown whether T cell vaccination elicits anti idiotypic humoral responses and whether the anti-idiotypic Abs play a similar role in the regulatory mechanism induced by T cell vaccination. In this study we examined the occurrence, the reactivity pattern, and the regulatory role of anti idiotypic Abs elicited by T cell vaccination in patients with multiple sclerosis. We demonstrated for the first time that B cells producing anti-idiotypic Abs could be isolated from vaccinated patients. These EBV-transformed B cell lines were selected for specific reactivity to a 20-mer TCR peptide incorporating a common complementarity-determining region 3 sequence of the immunizing T cell clones. The resulting anti-idiotypic Abs were found to react with the original immunizing T cell clones and exhibit an inhibitory effect on their proliferation. The findings suggest that anti-idiotypic Ab responses can be induced by T cell vaccination in humans and that their regulatory properties are likely to contribute to the suppression of myelin basic protein-reactive T cells in vaccinated patients. The study has important implications in our understanding of the regulatory role of the anti-idiotypic humoral responses induced by T cell vaccination. PMID- 11120810 TI - Evidence for protein kinase C-dependent and -independent activation of mitogen activated protein kinase in T cells: potential role of additional diacylglycerol binding proteins. AB - Activation of mitogen-activated protein kinases (MAPK) is a critical signal transduction event for CTL activation, but the signaling mechanisms responsible are not fully characterized. Protein kinase C (PKC) is thought to contribute to MAPK activation following TCR stimulation. We have found that dependence on PKC varies with the method used to stimulate the T cells. Extracellular signal regulated kinase (ERK) activation in CTL stimulated with soluble cross-linked anti-CD3 is completely inhibited by the PKC inhibitor bisindolylmaleimide (BIM). In contrast, only the later time points in the course of ERK activation are sensitive to BIM when CTL are stimulated with immobilized anti-CD3, a condition that stimulates CTL degranulation. Surprisingly, MAPK activation in response to immobilized anti-CD3 is strongly inhibited at all time points by the diacylglycerol (DAG)-binding domain inhibitor calphostin C implicating the contribution of a DAG-dependent but PKC-independent pathway in the activation of ERK in CTL clones. Chronic exposure to phorbol ester down-regulates the expression of DAG-responsive PKC isoforms; however, this treatment of CTL clones does not inhibit anti-CD3-induced activation of MAPK. Phorbol ester-treated cells have reduced expression of several isoforms of PKC but still express the recently described DAG-binding Ras guanylnucleotide-releasing protein. These results indicate that the late phase of MAPK activation in CTL clones in response to immobilized anti-CD3 stimulation requires PKC while the early phase requires a DAG-dependent, BIM-resistant component. PMID- 11120811 TI - The c-Abl tyrosine kinase is regulated downstream of the B cell antigen receptor and interacts with CD19. AB - c-Abl is a nonreceptor tyrosine kinase that we have recently linked to growth factor receptor signaling. The c-Abl kinase is ubiquitously expressed and localizes to the cytoplasm, plasma membrane, cytoskeleton, and nucleus. Thus, c Abl may regulate signaling processes in multiple subcellular compartments. Targeted deletion or mutation of c-Abl in mice results in a variety of phenotypes, including splenic and thymic atrophy and lymphopenia. Additionally, lymphocytes isolated from specific compartments of c-Abl mutant mice have reduced responses to a variety of stimuli and an increased susceptibility to apoptosis following growth factor deprivation. Despite these observations, little is known regarding the signaling mechanisms responsible for these phenotypes. We report here that splenic B cells from c-Abl-deficient mice are hyporesponsive to the proliferative effects of B cell Ag receptor (BCR) stimulation. The c-Abl kinase activity and protein levels are elevated in the cytosol following activation of the BCR in B cell lines. We show that c-Abl associates with and phosphorylates the BCR coreceptor CD19, and that c-Abl and CD19 colocalize in lipid membrane rafts. These data suggest a role for c-Abl in the regulation of B cell proliferation downstream of the BCR, possibly through interactions with CD19. PMID- 11120812 TI - Expression and function of IL-12 and IL-18 receptors on human tonsillar B cells. AB - IL-12 activates murine and human B cells, but little information is available as to the expression and function of IL-12R on human B lymphocytes. Here we show that the latter cells, freshly isolated from human tonsils, expressed the transcripts of both beta1 and beta2 chains of IL-12R and that beta2 chain mRNA was selectively increased (4- to 5-fold) by incubation with Staphylococcus aureus Cowan I bacteria or IL-12. B cell stimulation with IL-12 induced de novo expression of the transcripts of the two chains of IL-18R, i.e., IL-1 receptor related protein and accessory protein-like. Functional studies showed that both IL-12 and IL-18 signaled to B cells through the NF-kappaB pathway. In the case of IL-12, no involvement of STAT transcription factors, and in particular of STAT-4, was detected. c-rel and p50 were identified as the members of NF-kappaB family involved in IL-12-mediated signal transduction to B cells. IL-12 and IL-18 synergized in the induction of IFN-gamma production by tonsillar B cells, but not in the stimulation of B cell differentiation, although either cytokine promoted IgM secretion in culture supernatants. Finally, naive but not germinal center or memory, tonsillar B cells were identified as the exclusive IL-12 targets in terms of induction of NF-kappaB activation and of IFN-gamma production. PMID- 11120813 TI - CpG DNA induces maturation of dendritic cells with distinct effects on nascent and recycling MHC-II antigen-processing mechanisms. AB - Murine bone marrow cultured with GM-CSF produced dendritic cells (DCs) expressing MHC class II (MHC-II) but little CD40, CD80, or CD86. Oligodeoxynucleotides (ODN) containing CpG motifs enhanced DC maturation, increased MHC-II expression, and induced high levels of CD40, CD80, and CD86. When added with Ag to DCs for 24 h, CpG ODN enhanced Ag processing, and the half-life of peptide:MHC-II complexes was increased. However, Ag processing was only transiently enhanced, and exposure of DCs to CpG ODN for 48 h blocked processing of hen egg lysozyme (HEL) to HEL(48 61):I-A(k) complexes. Processing of this epitope required newly synthesized MHC II and was blocked by brefeldin A (BFA), suggesting that reduced MHC-II synthesis could explain decreased processing. Real-time quantitative PCR confirmed that CpG ODN decreased I-A(beta)(k) mRNA in DCs. In contrast, RNase(42-56):I-A(k) complexes were generated via a different processing mechanism that involved recycling MHC-II and was partially resistant to BFA. Processing of RNase(42-56):I A(k) persisted, although at reduced levels, after CpG-induced maturation of DCs, and this residual processing by mature DCs was completely resistant to BFA. Changes in endocytosis, which was transiently enhanced and subsequently suppressed by CpG ODN, may affect Ag processing by both nascent and recycling MHC II mechanisms. In summary, CpG ODN induce DC maturation, transiently increase Ag processing, and increase the half-life of peptide-MHC-II complexes to sustain subsequent presentation. Processing mechanisms that require nascent MHC-II are subsequently lost, but those that use recycling MHC-II persist even in fully mature DCs. PMID- 11120814 TI - Negative regulation of CD8+ T cell function by the IFN-induced and double stranded RNA-activated kinase PKR. AB - The IFN-induced and dsRNA-activated kinase (PKR) mediates the antiviral and antiproliferative effects of IFN-alpha and IFN-gamma. Despite these findings, PKR:(-/-) mice have no overt immunological phenotype. Here we tested the role of PKR in cellular immunity by determining the induction and elicitation of contact hypersensitivity in PKR:(-/-) mice, a model of T cell-mediated immunity. When compared with wild type, the magnitude of contact hypersensitivity responses in PKR:(-/-) mice were 2-fold higher and of extended duration. This was also observed when naive recipients of immune CD8(+) T cells from sensitized PKR:(-/-) and CD4(+) T cells from sensitized wild-type PKR:(+/+) or PKR:(-/-) mice were challenged with hapten, indicating a regulatory defect intrinsic to the CD8(+) T cell population. Isolated lymph node T cells from PKR:(-/-) mice were hyperproliferative during Con A-mediated stimulation. These results implicate PKR for the first time in the growth control of mature T lymphocytes and give insight into the negative regulation of CD8(+) T cell-mediated immune responses. PMID- 11120815 TI - Receptor revision in peripheral T cells creates a diverse V beta repertoire. AB - In Vbeta5 transgenic mice, the age-dependent accumulation of Vbeta5(-)CD4(+) T cells expressing endogenous Vss elements represents an exception to the rule of strict allelic exclusion at the TCRbeta locus. The appearance of these cells is limited to the lymphoid periphery and is driven by a peripherally expressed tolerogen. Expression of the lymphoid-specific components of the recombinase machinery and the presence of recombination intermediates strongly suggest that TCR revision rescues tolerogen-reactive peripheral T cells from deletion. Here, we report that the appearance of Vbeta5(-)CD4(+) T cells is CD28-dependent. In addition, we find that the TCR repertoire of this unusual population of T cells in individual Vbeta5 transgenic mice is surprisingly diverse, both at the level of surface protein and at the nucleotide level within a given family of V(D)Jbeta rearrangements. This faithful recreation of the nontransgenic repertoire suggests that endogenous Vbeta-expressing populations do not arise from expansion of an initially rare subset. Furthermore, the undersized N regions in revised TCR genes distinguish these sequences from those generated in the adult thymus. The diversity of the revised TCRs, the minimal mouse-to-mouse variation in the expressed endogenous Vbeta repertoire, the atypical length of junctional sequences, and the CD28 dependence of the accumulation of Vbeta5(-)CD4(+) T cells all point to their extrathymic origin. Thus, tolerogen-driven receptor revision in peripheral T cells can expand the TCR repertoire extrathymically, thereby contributing to the flexibility of the immune repertoire. PMID- 11120816 TI - Costimulation light: activation of CD4+ T cells with CD80 or CD86 rather than anti-CD28 leads to a Th2 cytokine profile. AB - To examine the role of CD28 and CTLA-4 in Th cell differentiation, we used a novel microsphere-based system to compare the effects of CD28 ligation by Ab or CD80/CD86. One set of beads was prepared by coating with anti-CD3 and anti-CD28 Ab. Another set of beads was prepared by immobilizing anti-CD3 and murine CD80-Ig fusion protein or murine CD86-Ig fusion protein on the beads. The three sets of beads were compared in their effects on the ability to activate and differentiate splenic CD4 T cells. When purified naive CD4(+) cells were stimulated in vitro, robust proliferation of similar magnitude was induced by all three sets of beads. When cytokine secretion was examined, all bead preparations induced an equivalent accumulation of IL-2. In contrast, there was a marked difference in the cytokine secretion pattern of the Th2 cytokines IL-4, IL-10, and IL-13. The B7-Ig stimulated cultures had high concentrations of Th2 cytokines, whereas there were low or undetectable concentrations in the anti-CD28-stimulated cultures. Addition of anti-CTLA-4 Fab augmented B7-mediated IL-4 secretion. These studies demonstrate that B7 is a critical and potent stimulator of Th2 differentiation, and that anti-CD28 prevents this effect. PMID- 11120817 TI - A role for CD21/CD35 and CD19 in responses to acute septic peritonitis: a potential mechanism for mast cell activation. AB - Although it is now appreciated that mast cell-mediated release of TNF-alpha is critical for resolution of acute septic peritonitis, questions remain as to how mast cells are activated upon peritoneal bacterial infection. Clues to how this may occur have been derived from earlier studies by Prodeus et al. in which complement proteins C3 and C4 were shown to be required for survival following cecal ligation and puncture (CLP), a model for acute septic peritonitis. To evaluate the mechanism for mast cell activation in the CLP model, complement receptor CD21/CD35-deficient mice (Cr2(null)) were examined in the present study. Along with CD19-deficient (CD19(null)) mice, these animals exhibit decreased survival following CLP compared with wild-type littermates. Injection of IgM before CLP does not change survival rates for Cr2(null) mice and only partially improves survival of CD19(null) mice, implicating CD21/CD35 and CD19 in mast cell activation. Interestingly, early TNF-alpha release is also impaired in Cr2(null) and CD19(null) animals, suggesting that these molecules directly affect mast cell activation. Cr2(null) and CD19(null) mice demonstrate an impairment in neutrophil recruitment and a corresponding increase in bacterial load. Examination of peritoneal mast cells by flow cytometry and confocal microscopy reveals the expression and colocalization of CD21/CD35 and CD19. Taken together, these findings suggest that the engagement of complement receptors CD21/CD35 along with CD19 on the mast cell surface by C3 fragments may be necessary for the full expression of mast cell activation in the CLP model. PMID- 11120818 TI - Mapping the ligand of the NK inhibitory receptor Ly49A on living cells. AB - We have used a recombinant, biotinylated form of the mouse NK cell inhibitory receptor, Ly49A, to visualize the expression of MHC class I (MHC-I) ligands on living lymphoid cells. A panel of murine strains, including MHC congenic lines, was examined. We detected binding of Ly49A to cells expressing H-2D(d), H-2D(k), and H-2D(p) but not to those expressing other MHC molecules. Cells of the MHC recombinant strain B10.PL (H-2(u)) not only bound Ly49A but also inhibited cytolysis by Ly49A(+) effector cells, consistent with the correlation of in vitro binding and NK cell function. Binding of Ly49A to H-2D(d)-bearing cells of different lymphoid tissues was proportional to the level of H-2D(d) expression and was not related to the lineage of the cells examined. These binding results, interpreted in the context of amino acid sequence comparisons and the recently determined three-dimensional structure of the Ly49A/H-2D(d) complex, suggest a role for amino acid residues at the amino-terminal end of the alpha1 helix of the MHC-I molecule for Ly49A interaction. This view is supported by a marked decrease in affinity of an H-2D(d) mutant, I52 M, for Ly49A. Thus, allelic variation of MHC-I molecules controls measurable affinity for the NK inhibitory receptor Ly49A and explains differences in functional recognition in different mouse strains. PMID- 11120819 TI - The physical association of protein kinase C theta with a lipid raft-associated inhibitor of kappa B factor kinase (IKK) complex plays a role in the activation of the NF-kappa B cascade by TCR and CD28. AB - We investigated the role of protein kinase C theta (PKCtheta) in the activation of the NF-kappaB cascade in primary human CD4(+) lymphocytes. Among six or so PKC isoforms expressed in T cells, only PKCtheta participates in the assembly of the supramolecular activation clusters at the contact site of the TCR with Ag. Signaling via both the TCR and CD28 is required for optimal activation of the multisubunit IkappaB kinase (IKK) complex in primary human T lymphocytes; this activation could be inhibited by a Ca(2+)-independent PKC isoform inhibitor, rottlerin. Moreover, endogenous PKCtheta physically associates with activated IKK complexes in CD3/CD28-costimulated primary CD4(+) T cells. The same set of stimuli also induced relocation of endogenous PKCtheta and IKKs to a GM1 ganglioside-enriched, detergent-insoluble membrane compartment in primary T cells. IKKs recruited to these lipid rafts were capable of phosphorylating a recombinant IkappaBalpha sustrate. Confocal microscopy further demonstrated that exogenously expressed PKCtheta and IKKss colocalize in the membrane of CD3/CD28 costimulated Jurkat T cells. Constitutively active but not kinase-inactive PKCtheta activated IKKbeta in Jurkat T cells. Expression of dominant-active PKCtheta also had stimulatory effects on the CD28 response element of the IL-2 promoter. Taken together, these data show that the activation of PKCtheta by the TCR and CD28 plays an important role in the assembly and activation of IKK complexes in the T cell membrane. PMID- 11120820 TI - Peroxisome proliferator activator receptor-gamma agonists and 15-deoxy Delta(12,14)(12,14)-PGJ(2) induce apoptosis in normal and malignant B-lineage cells. AB - The research described herein evaluates the expression and functional significance of peroxisome proliferator activator receptor-gamma (PPAR-gamma) on B-lineage cells. Normal mouse B cells and a variety of B lymphoma cells reflective of stages of B cell differentiation (e.g., 70Z/3, CH31, WEHI-231, CH12, and J558) express PPAR-gamma mRNA and, by Western blot analysis, the 67-kDa PPAR-gamma protein. 15-Deoxy-Delta(12,14)-PGJ(2) (15d-PGJ(2)), a PPAR-gamma agonist, has a dose-dependent antiproliferative and cytotoxic effect on normal and malignant B cells as shown by [(3)H]thymidine and 3-[4,5-dimethylthiazol-2 yl]-2, 5-diphenyltetrazolium bromide assays. Only PPAR-gamma agonists (thiazolidinediones), and not PPAR-alpha agonists, mimicked the effect of 15d PGJ(2) on B-lineage cells, indicating that the mechanism by which 15d-PGJ(2) negatively affects B-lineage cells involves in part PPAR-gamma. The mechanism by which PPAR-gamma agonists induce cytotoxicity is via apoptosis, as shown by annexin V staining and as confirmed by DNA fragmentation detected using the TUNEL assay. Interestingly, addition of PGF(2alpha), which was not known to affect lymphocytes, dramatically attenuated the deleterious effects of PPAR-gamma agonists on B lymphomas. Surprisingly, 15d-PGJ(2) induced a massive increase in nuclear mitogen-activated protein kinase activation, and pretreatment with PGF(2alpha) blunted the mitogen-activated protein kinase activation. This is the first study evaluating PPAR-gamma expression and its significance on B lymphocytes. PPAR-gamma agonists may serve as a counterbalance to the stimulating effects of other PGs, namely PGE(2), which promotes B cell differentiation. Finally, the use of PGs, such as 15d-PGJ(2), and synthetic PPAR-gamma agonists to induce apoptosis in B-lineage cells may lead to the development of novel therapies for fatal B lymphomas. PMID- 11120821 TI - Crystal structure of an antibody bound to an immunodominant peptide epitope: novel features in peptide-antibody recognition. AB - The crystal structure of Fab of an Ab PC283 complexed with its corresponding peptide Ag, PS1 (HQLDPAFGANSTNPD), derived from the hepatitis B virus surface Ag was determined. The PS1 stretch Gln2P to Phe7P is present in the Ag binding site of the Ab, while the next three residues of the peptide are raised above the binding groove. The residues Ser11P, Thr12P, and Asn13P then loop back onto the Ag-binding site of the Ab. The last two residues, Pro14P and Asp15P, extend outside the binding site without forming any contacts with the Ab. The PC283-PS1 complex is among the few examples where the light chain complementarity determining regions show more interactions than the heavy chain complementarity determining regions, and a distal framework residue is involved in Ag binding. As seen from the crystal structure, most of the contacts between peptide and Ab are through the five residues, Leu3-Asp4-Pro5-Ala6-Phe7, of PS1. The paratope is predominantly hydrophobic with aromatic residues lining the binding pocket, although a salt bridge also contributes to stabilizing the Ag-Ab interaction. The molecular surface area buried upon PS1 binding is 756 A(2) for the peptide and 625 A(2) for the Fab, which is higher than what has been seen to date for Ab peptide complexes. A comparison between PC283 structure and a homology model of its germline ancestor suggests that paratope optimization for PS1 occurs by improving both charge and shape complementarity. PMID- 11120822 TI - The transcription factor Bright associates with Bruton's tyrosine kinase, the defective protein in immunodeficiency disease. AB - Binding of the transcription factor Bright to Ig heavy chain loci after B cell activation is associated with increased heavy chain transcription. We now report that Bright coprecipitates with Bruton's tyrosine kinase (Btk), the defective enzyme in X-linked immunodeficiency disease (xid). Furthermore, we observed Btk in the nucleus of activated murine B cells, and mobility shift assays suggest that it is a component of the Bright DNA-binding complex. While BRIGHT protein was synthesized in activated spleen cells from xid mice, it did not bind DNA or associate stably with Btk. These data suggest that deficiencies in BRIGHT DNA binding activity may contribute to the defects in Ig production seen in xid mice. PMID- 11120823 TI - Identification of fetal liver tyrosine kinase 3 (flt3) ligand domain required for receptor binding and function using naturally occurring ligand isoforms. AB - We used a comparative approach to identify the fetal liver tyrosine kinase 3 (flt3) ligand structure required for binding and function. Two conserved bovine flt3 ligand isoforms, which differ in a defined region within the extracellular domain, were identified and shown to be uniformly transcribed in individuals with diverse MHC haplotypes. Notably, at the amino acid level, the extracellular domain of the bovine flt3 ligand isoform 1 is 81 and 72% identical with the extracellular domains of the human and murine flt3 ligands, respectively, whereas isoform-2 has a deletion within this domain. Bovine flt3 ligand isoform 1, but not 2, bound the human flt3 receptor and stimulated murine pro B cells transfected with the murine flt3 receptor. This retention of binding and function allowed definition of key residues by identifying sequences conserved among species. We have shown that a highly conserved, 18 aa sequence within the flt3 ligand extracellular domain is required for flt3 receptor binding and function. However, a peptide representing this sequence is insufficient for receptor binding as demonstrated by its failure to inhibit the bovine flt3 ligand isoform 1 binding to the human flt3 receptor. The requirement for flanking structure was confirmed by testing bovine flt3 ligand isoform 1 constructs truncated at specific residues outside the 18 aa sequence. Overall, the flt3 ligand structure required for function is markedly similar to that of the related hemopoietic growth factors, CSF-1 and steel factor. This definition of the required flt3 ligand structure will facilitate development of agonists to enhance dendritic cell recruitment for vaccines and immunotherapy. PMID- 11120824 TI - The murine IL-2 promoter contains distal regulatory elements responsive to the Ah receptor, a member of the evolutionarily conserved bHLH-PAS transcription factor family. AB - Signaling through the TCR and costimulatory signals primarily control transcription of the IL-2 gene in naive T cells. The minimal promoter necessary for this expression lies proximal, between -300 and the transcription start site. We had previously shown that activation of the arylhydrocarbon receptor (AHR), a member of the bHLH-PAS family of transcription factors, leads to increased mRNA expression of IL-2 in murine fetal thymocytes. The AHR is abundant in the thymus and may play a role for the development of the immune system. Moreover, its overactivation by chemicals such as dioxins leads to immunosuppression and thymic involution. Binding motifs for the liganded AHR can be identified in the distal region -1300 to -800 of the mouse IL-2 promoter. We show here that these DNA motifs, the so-called dioxin response elements, after binding to the liganded AHR are sufficient to transactivate luciferase expression in a reporter gene system. The IL-2 gene can be induced by the AHR also in thymocytes in vivo after injection of 2,3,7, 8-tetrachlorodibenzo-p-dioxin, a potent ligand of the AHR. The AHR mediates the IL-2 induction as shown with AHR-deficient mice. However, in spleen cells in vitro costimulation via the TCR is necessary for optimal IL-2 gene induction. Thus, the IL-2 promoter region contains novel distal regulatory elements that can be addressed by the AHR to induce IL-2 and can cooperate with the proximal promoter in this. PMID- 11120825 TI - Conservation of Mhc class III region synteny between zebrafish and human as determined by radiation hybrid mapping. AB - In the HLA, H2, and other mammalian MHC:, the class I and II loci are separated by the so-called class III region comprised of approximately 60 genes that are functionally and evolutionarily unrelated to the class I/II genes. To explore the origin of this island of unrelated loci in the middle of the MHC: 19 homologues of HLA class III genes, we identified 19 homologues of HLA class III genes as well as 21 additional non-class I/II HLA homologues in the zebrafish and mapped them by testing a panel of 94 zebrafish-hamster radiation hybrid cell lines. Six of the HLA class III and eight of the flanking homologues were found to be linked to the zebrafish class I (but not class II) loci in linkage group 19. The remaining homologous loci were found to be scattered over 14 zebrafish linkage groups. The linkage group 19 contains at least 25 genes (not counting the class I loci) that are also syntenic on human chromosome 6. This gene assembly presumably represents the pre-MHC: that existed before the class I/II genes arose. The pre MHC: may not have contained the complement and other class III genes involved in immune response. PMID- 11120826 TI - Distinct T cell interactions with HLA class II tetramers characterize a spectrum of TCR affinities in the human antigen-specific T cell response. AB - The polyclonal nature of T cells expanding in an ongoing immune response results in a range of disparate affinities and activation potential. Recently developed human class II tetramers provide a means to analyze this diversity by direct characterization of the trimolecular TCR-peptide-MHC interaction in live cells. Two HSV-2 VP16(369-379)-specific, DQA1*0102/DQB1*0602 (DQ0602)-restricted T cell clones were compared by means of T cell proliferation assay and HLA-DQ0602 tetramer staining. These two clones were obtained from the same subject, but show different TCR gene usage. Clone 48 was 10-fold more sensitive to VP16(369-379) peptide stimulation than clone 5 as assayed by proliferation assays, correlating with differences in MHC tetramer binding. Clone 48 gave positive staining with the DQ0602/VP16(369-379) tetramer at either 23 or 37 degrees C. Weak staining was also observed at 4 degrees C. Clone 5 showed weaker staining compared with clone 48 at 37 degrees C, and no staining was observed at 23 degrees C or on ice. Receptor internalization was not required for positive staining. Competitive binding indicates that the cell surface TCR of clone 48 has higher affinity for the DQ0602/VP16(369-379) complex than clone 5. The higher binding affinity of clone 48 for the peptide-MHC complex also correlates with a slower dissociation rate compared with clone 5. PMID- 11120827 TI - Antibody repertoire development in fetal and neonatal piglets. II. Characterization of heavy chain complementarity-determining region 3 diversity in the developing fetus. AB - Since the actual combinatorial diversity in the V(H) repertoire in fetal piglets represents <1% of the potential in mice and humans, we wondered whether 1) complementarity-determining region 3 (CDR3) diversity was also restricted; 2) CDR3 diversity changed with fetal age; and 3) to what extent CDR3 contributed to the preimmune VDJ repertoire. CDR3 spectratyping and sequence analyses of 213 CDR3s recovered from >30 fetal animals of different ages showed that >95% of VDJ diversity resulted from junctional diversity. Unlike sheep and cattle, somatic hypermutation does not contribute to the repertoire. These studies also revealed that 1) N region additions are as extensive in VDJ rearrangements recovered at 30 days as those in late term fetuses, suggesting that TdT is fully active at the onset of VDJ rearrangement; 2) nearly 90% of all rearrangement are in-frame until late gestation; 3) the oligoclonal CDR3 spectratype of 30-day fetal liver becomes polyclonal by 50 days, while this change occurs much later in spleen; 4) there is little evidence of individual variation in CDR3 spectratype or differences in spectratype among lymphoid tissues with the exception of the thymus; and 4) there is a tendency for usage of the most J(H) proximal D(H) segment (D(H)B) to decrease in older fetuses and for the longer D(H) segment to be trimmed to the same length as the shorter D(H) when used in CDR3. These findings suggest that in the fetal piglet, highly restricted combinatorial diversity and the lack of somatic mutation are compensated by early onset of TdT activity and other mechanisms that contribute to CDR3 junctional diversity. PMID- 11120828 TI - Delineation of the human systemic lupus erythematosus anti-Smith antibody response using phage-display combinatorial libraries. AB - The anti-Smith (Sm) autoantibody response is highly specific for systemic lupus erythematosus and is predominantly targeted to the Sm-B/B' and -D1 polypeptides. In all animal species thus far studied, anti-Sm Abs initially recognize proline rich epitopes in the carboxyl terminus of the Sm-B/B' protein and subsequently to multiple other epitopes in B/B' and D. The absence of appropriate mAbs has limited our understanding of the genetic and structural basis of this autoimmune response. Using phage-display technology and lymphocytes from a systemic lupus erythematosus patient we have generated the first and only panel of human IgG anti-Sm mAbs thus far available. These Abs reproduced to a remarkable extent the serological reactivity of the patient. Epitope mapping and genetic studies revealed that the anti-Sm response is produced by distinct B cell clones with restricted epitope reactivity. All of the Abs in our study were exclusively encoded by different members of the V(H)4 gene family. On the aggregate, our results demonstrate that combinatorial libraries can recapitulate the immune repertoire of peripheral blood B memory cells and that epitope spreading appears to occur through the sequential recruitment of nonclonally related autoreactive B cell clones. PMID- 11120829 TI - Activation of MHC class I, II, and CD40 gene expression by histone deacetylase inhibitors. AB - Epigenetic mechanisms are involved in regulating chromatin structure and gene expression through repression. In this study, we show that histone deacetylase inhibitors (DAIs) that alter the acetylation of histones in chromatin enhance the expression of several genes on tumor cells including: MHC class I, II, and the costimulatory molecule CD40. Enhanced transcription results in a significant increase in protein expression on the tumor cell surface, and expression can be elicited on some tumors that are unresponsive to IFN-gamma. The magnitude of induction of these genes cannot be explained by the effect of DAIs on the cell cycle or enhanced apoptosis. Induction of class II genes by DAIs was accompanied by activation of a repressed class II transactivator gene in a plasma cell tumor but, in several other tumor cell lines, class II was induced in the apparent absence of class II transactivator transcripts. These findings also suggest that the abnormalities observed in some tumors in the expression of genes critical to tumor immunity may result from epigenetic alterations in chromatin and gene regulation in addition to well-established mutational mechanisms. PMID- 11120830 TI - Factors controlling the trafficking and processing of a leader-derived peptide presented by Qa-1. AB - Many leader-derived peptides require TAP for presentation by class I molecules. This TAP dependence can either be ascribed to the inability of proteases resident in the endoplasmic reticulum (ER) to trim leader peptide precursors into the appropriate epitope or the failure of a portion of the leader segment to gain access to the lumen of the ER. Using the Qa-1 binding epitope, Qdm derived from a class Ia leader as a model, we show that many cell types lack ER protease activity to trim this peptide at its C terminus. However, both T1 and T2 cells contain appropriate protease activity to process the full length D(d) leader (DL) when introduced into the ER lumen. Nevertheless, both T1 cells treated with the TAP inhibitor ICP47 and TAP(-) T2 cells fail to present this epitope from either the intact D(d) molecule or a minigene encoding the DL. This indicates that the portion of the leader containing Qdm does not gain access to the ER. However, changing the Arg at P7 of the DL to a Cys can alter its trafficking and allows for TAP-independent presentation of the Qdm epitope. PMID- 11120831 TI - HIV-1 Tat represses transcription from the mannose receptor promoter. AB - The mannose receptor is expressed on mature macrophages and immature dendritic cells, and functions to mediate phagocytosis of pathogens and capture of Ags for delivery to MHC class II-containing intracellular compartments. It has been previously reported that HIV-1-infected macrophages have reduced functions associated with the mannose receptor, including impaired Pneumocystis carinii phagocytosis and mannosylated albumin uptake. Several HIV-1-derived proteins including the Tat protein have been shown to transcriptionally repress host cell genes. The present study was undertaken to define the role of the HIV-1-derived protein Tat in HIV-mediated mannose receptor down-regulation. Cotransfection of the human macrophage cell line U937 with a Tat expression vector and a mannose receptor promoter-luciferase reporter construct resulted in down-regulation of mannose receptor promoter activity. This repression was targeted to the basal promoter. Expression of either one- or two-exon Tat resulted in decreased promoter activity. The addition of the transactivation response element (TAR) sequence enhanced the Tat-mediated repression. Down-regulation was also seen when transfected cells were treated with exogenously added Tat protein. These results are consistent with a mechanism whereby Tat reduces mannose receptor promoter activity by interfering with the host transcriptional initiation machinery, potentially resulting in decreased levels of surface mannose receptor available for Ag or pathogen capture. PMID- 11120832 TI - Two different epitopes of the signal transducer gp130 sequentially cooperate on IL-6-induced receptor activation. AB - Cytokines are key mediators for the regulation of hemopoiesis and the coordination of immune responses. They exert their various functions through activation of specific cell surface receptors, thereby initiating intracellular signal transduction cascades which lead to defined cellular responses. As the common signal-transducing receptor subunit of at least seven different cytokines, gp130 is an important member of the family of hemopoietic cytokine receptors which are characterized by the presence of at least one cytokine-binding module. Mutants of gp130 that either lack the Ig-like domain D1 (DeltaD1) or contain a distinct mutation (F191E) within the cytokine-binding module have been shown to be severely impaired with respect to IL-6 induced signal transduction. After cotransfection of COS-7 cells with a combination of both inactive gp130 mutants, signal transduction in response to IL-6 is restored. Whereas cells transfected with DeltaD1 do not bind IL-6/sIL-6R complexes, cells transfected with the F191E mutant bind IL-6/sIL-6R with low affinity. Combination of DeltaD1 and F191E, however, leads to high-affinity ligand binding. These data suggest that two different gp130 epitopes, one on each receptor chain, sequentially cooperate in asymmetrical binding of IL-6/IL-6R in a tetrameric signaling complex. On the basis of our data, a model for the mechanism of IL-6-induced gp130 activation is proposed. PMID- 11120833 TI - Fab chains as an efficient heterodimerization scaffold for the production of recombinant bispecific and trispecific antibody derivatives. AB - Due to their multispecificity and versatility, bispecific Abs (BsAbs) are promising therapeutic tools in tomorrow's medicine. Especially intermediate-sized BsAbs that combine body retention with tissue penetration are valuable for therapy but necessitate expression systems that favor heterodimerization of the binding sites for large-scale application. To identify heterodimerization domains to which single-chain variable fragments (scFv) can be fused, we compared the efficiency of heterodimerization of CL and CH1 constant domains with complete L and Fd chains in mammalian cells. We found that the isolated CL:CH1 domain interaction was inefficient for secretion of heterodimers. However, when the complete L and Fd chains were used, secretion of L:Fd heterodimers was highly successful. Because these Fab chains contribute a binding moiety, C-terminal fusion of a scFv molecule to the L and/or Fd chains generated BsAbs or trispecific Abs (TsAbs) of intermediate size (75-100 kDa). These disulfide stabilized bispecific Fab-scFv ("bibody") and trispecific Fab-(scFv)(2) ("tribody") heterodimers represent up to 90% of all secreted Ab fragments in the mammalian expression system and possess fully functional binding moieties. Furthermore, both molecules recruit and activate T cells in a tumor cell dependent way, whereby the trispecific derivative can exert this activity to two different tumor cells. Thus we propose the use of the disulfide-stabilized L:Fd heterodimer as an efficient platform for production of intermediate-sized BsAbs and TsAbs in mammalian expression systems. PMID- 11120834 TI - Induced kappa receptor editing shows no allelic preference in a mouse pre-B cell line. AB - B cell Ag receptor editing is a process that can change kappa antigen recognition specificity of a B cell receptor through secondary gene rearrangements on the same allele. In this study we used a model mouse pre-B cell line (38B9) to examine factors that might affect allelic targeting of secondary rearrangements of the kappa locus. We isolated clones that showed both productive and nonproductive rearrangements of one kappa allele, while retaining the other kappa allele in the germline configuration (kappa(+)/kappa degrees or kappa(-)/kappa degrees ). In the absence of any selective pressures, subsequent rearrangement of the germline alleles occurred at the same frequency as secondary rearrangement of the productive or nonproductive rearranged alleles. Because 38B9 cells lack Ig heavy chains, we stably expressed mu heavy chain protein in 38B9 cells to determine whether heavy-light pairing might affect allelic targeting of secondary kappa rearrangements. Although the expression of heavy chain was found to both pair with and stabilize kappa protein in these cells, it had no effect on preferential targeting Vkappa-Jkappa receptor editing compared with rearrangement of a germline allele. These studies suggest that in the absence of selection to eliminate autoreactive Vkappa-Jkappa genes, there is no allelic preference for secondary rearrangement events in 38B9 cells. PMID- 11120835 TI - Immunization with a recombinant stage-regulated surface protein from Leishmania donovani induces protection against visceral leishmaniasis. AB - Vaccination against visceral leishmaniasis has received limited attention compared with cutaneous leishmaniasis, although the need for an effective vaccine against visceral leishmaniasis is pressing. In this study, we demonstrate for the first time that a recombinant stage-specific hydrophilic surface protein of Leishmania donovani, recombinant hydrophilic acylated surface protein B1 (HASPB1), is able to confer protection against experimental challenge. Protection induced by rHASPB1 does not require adjuvant and, unlike soluble Leishmania Ag + IL-12, extends to the control of parasite burden in the spleen, an organ in which parasites usually persist and are refractory to a broad range of immunological and chemotherapeutic interventions. Both immunohistochemistry (for IL-12p40) and enzyme-linked immunospot assay (for IL-12p70) indicate that immunization with rHASPB1 results in IL-12 production by dendritic cells, although an analysis of Ab isotype responses to rHASPB1 suggests that this response is not sufficient in magnitude to induce a polarized Th1 response. Although both vaccinated and control-infected mice have equivalent frequencies of rHASPB1-specific CD4(+) T cells producing IFN-gamma, vaccine-induced protection correlates with the presence of rHASPB1-specific, IFN-gamma-producing CD8(+) T cells. Thus, we have identified a novel vaccine candidate Ag for visceral leishmaniasis, which appears to operate via a mechanism similar to that previously associated with DNA vaccination. PMID- 11120836 TI - A mechanism for the impaired IFN-gamma production in C-C chemokine receptor 2 (CCR2) knockout mice: role of CCR2 in linking the innate and adaptive immune responses. AB - We have recently shown that mice with a targeted disruption of CCR2, the receptor for monocyte chemoattractant protein-1, have markedly impaired recruitment of macrophages to sites of inflammation. An unexpected finding in the CCR2(-/-) mice was a dramatic decrease in the production of IFN-gamma after challenge with purified protein derivative of Mycobacterium bovis. In this study, we have investigated the mechanism of this cytokine production defect. In vitro, direct activation of splenocytes with CD3/CD28 Abs failed to reveal any differences in IFN-gamma production between CCR2(+/+) and CCR2(-/-) mice. However, after immunization, the number of Ag-specific, IFN-gamma-producing cells in the draining lymph nodes was decreased by 70% in the CCR2(-/-) mice, suggesting an in vivo trafficking defect. Direct measurement of cell trafficking with fluorescently labeled CFA revealed a marked decrease in the number of monocytes/macrophages migrating to the site of immunization and to the draining lymph nodes in the CCR2(-/-) mice. The data suggest that impaired trafficking of APCs in the CCR2(-/-) mice contributes to the defect in IFN-gamma production. These data support the idea that CCR2-positive monocytes/macrophages are critical in linking the innate and adaptive immune responses. PMID- 11120837 TI - The importance of exogenous antigen in priming the human CD8+ T cell response: lessons from the EBV nuclear antigen EBNA1. AB - Mouse models suggest that the processing of exogenous Ag by dendritic cells can be important for priming the CD8(+) CTL response. To study the situation in humans, we have exploited the CTL response to EBV infection. In this context EBV expresses eight latent proteins, of which EBV-encoded nuclear Ag (EBNA) 3A, 3B, and 3C appear to be immunodominant for CTL responses, whereas another nuclear Ag, EBNA1, which is completely protected from endogenous presentation via the MHC class I pathway, is thought to induce responses rarely, if ever. Here, using EBNA1 peptides and/or EBNA1 protein-loaded dendritic cells as in vitro stimuli, we have identified memory CTL responses to HLA-B*3501, -B7, and -B53-restricted EBNA1 epitopes that can be as strong as those seen in immunodominant epitopes from the "conventionally processed"" EBNA3 Ags. Furthermore, we used HLA-peptide tetramers to show that the primary response to one such EBNA1 epitope constituted up to 5% of the CD8(+) T cells in infectious mononucleosis blood, the strongest latent Ag-specific response yet detected in this setting. We conclude that exogenous protein represents a significant source of Ag for priming the human CTL response. PMID- 11120838 TI - Human CD8+ CTL specific for the mycobacterial major secreted antigen 85A. AB - The role of CD8(+) CTL in protection against tuberculosis in human disease is unclear. In this study, we stimulated the peripheral blood mononuclear cells of bacillus Calmette-Guerin (BCG)-vaccinated individuals with live Mycobacterium bovis BCG bacilli to establish short-term cell lines and then purified the CD8(+) T cells. A highly sensitive enzyme-linked immunospot (ELISPOT) assay for single cell IFN-gamma release was used to screen CD8(+) T cells with overlapping peptides spanning the mycobacterial major secreted protein, Ag85A. Three peptides consistently induced a high frequency of IFN-gamma responsive CD8(+) T cells, and two HLA-A*0201 binding motifs, P(48-56) and P(242-250), were revealed within the core sequences. CD8(+) T cells responding to the 9-mer epitopes were visualized within fresh blood by ELISPOT using free peptide or by binding of HLA-A*0201 tetrameric complexes. The class I-restricted CD8(+) T cells were potent CTL effector cells that efficiently lysed an HLA-A2-matched monocyte cell line pulsed with peptide as well as autologous macrophages infected with Mycobacterium tuberculosis or recombinant vaccinia virus expressing the whole Ag85A protein. Tetramer assays revealed a 6-fold higher frequency of peptide-specific T cells than IFN-gamma ELISPOT assays, indicating functional heterogeneity within the CD8(+) T cell population. These results demonstrate a previously unrecognized, MHC class I-restricted, CD8(+) CTL response to a major secreted Ag of mycobacteria and supports the use of Ag85A as a candidate vaccine against tuberculosis. PMID- 11120839 TI - Synergy and cross-tolerance between toll-like receptor (TLR) 2- and TLR4-mediated signaling pathways. AB - A family of Toll-like receptor (TLR) mediates the cellular response to bacterial cell wall components; murine TLR2 and TLR4 recognize mycoplasmal lipopeptides (macrophage-activating lipopeptides, 2 kDa (MALP-2)) and LPS, respectively. Costimulation of mouse peritoneal macrophages with MALP-2 and LPS results in a marked increase in TNF-alpha production, showing the synergy between TLR2- and TLR4-mediated signaling pathways. Macrophages pretreated with LPS show hyporesponsiveness to the second LPS stimulation, termed LPS tolerance. The LPS tolerance has recently been shown to be primarily due to the down-regulation of surface expression of the TLR4-MD2 complex. When macrophages were treated with MALP-2, the cells showed hyporesponsiveness to the second MALP-2 stimulation, like LPS tolerance. Furthermore, macrophages pretreated with MALP-2 showed reduced production of TNF-alpha in response to LPS. LPS-induced activation of both NF-kappaB and c-Jun NH(2)-terminal kinase was severely impaired in MALP-2 pretreated cells. However, MALP-2-pretreated macrophages did not show any reduction in surface expression of the TLR4-MD2 complex. These findings indicate that LPS-induced LPS tolerance mainly occurs through the down-regulation of surface expression of the TLR4-MD2 complex; in contrast, MALP-2-induced LPS tolerance is due to modulation of the downstream cytoplasmic signaling pathways. PMID- 11120841 TI - IL-12-activated NK cells reduce lung eosinophilia to the attachment protein of respiratory syncytial virus but do not enhance the severity of illness in CD8 T cell-immunodeficient conditions. AB - Bronchiolitis caused by respiratory syncytial virus (RSV) infection is a major cause of hospitalization in children under 1 year of age. RSV causes common colds in older children and adults, but can cause serious disease in immunodeficient patients and the elderly. Development of effective vaccines and treatments for RSV infection is therefore a priority. Because bronchiolitis and vaccine augmented disease are thought to be caused by exuberant T cell activation, attention has focused on the use of immunomodulators that affect T cell responses. In mice, IL-12 treatment down-regulates type 2 cytokine responses to the attachment protein G of RSV, reducing lung eosinophilia but further enhancing illness. We now show that CD8(+) T cells are responsible for enhanced weight loss, whereas IL-12-activated NK cells express high levels of IFN-gamma and inhibit lung eosinophilia without causing illness. Moreover, unlike immunocompetent mice, virus is detected in the mediastinal lymph nodes after elimination of both CD8(+) T cells and NK cells. These studies show that innate immune responses to viral infections direct the pattern of subsequent specific immunity and are critical to the development of nonpathogenic antiviral effects. We speculate that IL-12 treatment might be beneficial and safe in T cell deficient patients with RSV pneumonitis. PMID- 11120840 TI - Granulysin, a T cell product, kills bacteria by altering membrane permeability. AB - Granulysin, a protein located in the acidic granules of human NK cells and cytotoxic T cells, has antimicrobial activity against a broad spectrum of microbial pathogens. A predicted model generated from the nuclear magnetic resonance structure of a related protein, NK lysin, suggested that granulysin contains a four alpha helical bundle motif, with the alpha helices enriched for positively charged amino acids, including arginine and lysine residues. Denaturation of the polypeptide reduced the alpha helical content from 49 to 18% resulted in complete inhibition of antimicrobial activity. Chemical modification of the arginine, but not the lysine, residues also blocked the antimicrobial activity and interfered with the ability of granulysin to adhere to Escherichia coli and Mycobacterium tuberculosis. Granulysin increased the permeability of bacterial membranes, as judged by its ability to allow access of cytosolic ss galactosidase to its impermeant substrate. By electron microscopy, granulysin triggered fluid accumulation in the periplasm of M. tuberculosis, consistent with osmotic perturbation. These data suggest that the ability of granulysin to kill microbial pathogens is dependent on direct interaction with the microbial cell wall and/or membrane, leading to increased permeability and lysis. PMID- 11120842 TI - T cell-mediated tumor rejection displays diverse dependence upon perforin and IFN gamma mechanisms that cannot be predicted from in vitro T cell characteristics. AB - Experimental pulmonary metastases have been successfully treated by adoptive transfer of tumor-sensitized T cells from perforin knockout (KO) or Fas/APO-1 ligand(KO) mice, suggesting a prominent role for secretion of cytokines such as IFN-gamma. In the present study we confirmed that rejection of established methylcholanthrene-205 (MCA-205) pulmonary metastases displayed a requirement for T cell IFN-gamma expression. However, this requirement could be obviated by transferring larger numbers of tumor-sensitized IFN-gamma (KO) T cells or by immunosensitizing sublethal irradiation (500 rad) of the host before adoptive therapy. Extrapulmonary tumors (MCA-205 s.c. and intracranial) that required adjunct sublethal irradiation for treatment efficacy also displayed no requirement for host or T cell expression of IFN-gamma. Nonetheless, rejection of MCA-205 s.c. tumors and i.p. EL-4 tumors, but not MCA-205 pulmonary or intracranial tumors, displayed a significant requirement for T cell perforin expression (i.e., CTL participation). The capacity of T cells to lyse tumor targets and secrete IFN-gamma in vitro before adoptive transfer was nonpredictive of the roles of these activities in subsequent tumor rejection. Adoptive therapy studies employing KO mice are therefore indispensable for revealing a diversity of tumor rejection mechanisms that may lack in vitro correlation due to delays in their induction. Seemingly contradictory KO data from different studies are reconciled by the capacity of anti-tumor T cells to rely on alternative mechanisms when treated in larger numbers, the variable participation of CTL at different anatomic locations of tumor, and the apparent capacity of sublethal irradiation to provide a therapeutic alternative to host or T cell IFN-gamma production. PMID- 11120843 TI - Evidence for an accessory protein function for Toll-like receptor 1 in anti bacterial responses. AB - Members of the Toll-like receptor (TLR) family are components of the mammalian anti-microbial response, signaling with a domain closely related to that of IL-1 receptors. In this report the expression and function of TLR1, a TLR of unknown function, are examined. TLR1 is expressed by monocytes, as demonstrated using a novel mAb. Monocytes also express TLR2. TLR1 transfection of HeLa cells, which express neither TLR1 nor TLR2, was not sufficient to confer responsiveness to several microbial extracts. However, cotransfection of TLR1 and TLR2 resulted in enhanced signaling by HeLa cells to soluble factors released from Neisseria meningitidis relative to the response with either TLR alone. This phenomenon was also seen with high concentrations of some preparations of LPS. The N. meningitidis factors recognized by TLR1/TLR2 were not released by N. meningitidis mutant in the LpxA gene. Although LpxA is required for LPS biosynthesis, because cooperation between TLR1 and TLR2 was not seen with all LPS preparations, the microbial component(s) TLR1/2 recognizes is likely to be a complex of LPS and other molecules or a compound metabolically and chemically related to LPS. The functional IL-1R consists of a heterodimer; this report suggests a similar mechanism for TLR1 and TLR2, for certain agonists. These data further suggest that mammalian responsiveness to some bacterial products may be mediated by combinations of TLRs, suggesting a mechanism for diversifying the repertoire of Toll-mediated responses. PMID- 11120844 TI - V gamma 9V delta 2 T cells impair intracellular multiplication of Brucella suis in autologous monocytes through soluble factor release and contact-dependent cytotoxic effect. AB - Human Vgamma9Vdelta2 T cells are considered to play an important role in brucellosis, as this population is dramatically increased in peripheral blood of patients during the acute phase of the infection. This T lymphocyte population has been largely demonstrated to be activated by small m.w. nonpeptidic molecules from natural or synthetic origin. We recently identified a nonpeptidic fraction of Brucella suis that specifically activates human Vgamma9Vdelta2 T cells. Using a two-separate-chambers system, we showed that Brucella fraction, as well as isopentenyl pyrophosphate-activated Vgamma9Vdelta2 T cells, impaired the multiplication of B. suis in differentiated THP-1 cells through TNF-alpha and IFN gamma release. In the present study, using circulating Vgamma9Vdelta2 T cells and autologous monocytes infected with B. suis, we provide evidence that 1) intramonocytic multiplication of B. suis is impaired by supernatants of activated Vgamma9Vdelta2 T cells in part via TNF-alpha and IFN-gamma, this impairment occurring without host cell lysis; 2) unstimulated Vgamma9Vdelta2 T cells can impair intracellular bacterial multiplication after their activation by soluble factors released by infected monocytes; and 3) activated Vgamma9Vdelta2 T cells lyse Brucella-infected monocytes in a contact-dependent manner. Taken together, these results provide evidence that Vgamma9Vdelta2 T cells, in addition to being directly activated by soluble nonpeptidic molecules, can be stimulated to become highly cytotoxic in the specific presence of infected monocytes; moreover, they suggest how Vgamma9Vdelta2 T cells could be triggered and respond as antibacterial effector cells in the early stages of Brucella infection. PMID- 11120845 TI - T cell expression cloning of a Mycobacterium tuberculosis gene encoding a protective antigen associated with the early control of infection. AB - Infection of C57BL/6 mice with Mycobacterium tuberculosis results in the development of a progressive disease during the first 2 wk after challenge. Thereafter, the disease is controlled by the emergence of protective T cells. We have used this infection model in conjunction with direct T cell expression cloning to identify Ags involved with the early control of the disease. A protective M. tuberculosis-specific CD4 T cell line derived from mice at 3 wk postchallenge was used to directly screen an M. tuberculosis genomic expression library. This screen resulted in the identification of a genomic clone comprising two putative adjacent genes with predicted open reading frames of 10 and 41 kDa, MTB10 and MTB41, respectively (the products of Rv0916c and Rv0915c, respectively, in the TubercuList H37Rv database). MTB10 and MTB41 belong to the PE and PPE family of proteins recently identified to comprise 10% of the M. tuberculosis genome. Evaluation of the recombinant proteins revealed that MTB41, but not MTB10, is the Ag recognized by the cell line and by M. tuberculosis-sensitized human PBMC. Moreover, C57BL/6 mice immunized with MTB41 DNA developed both CD4- (predominantly Th1) and CD8-specific T cell responses to rMTB41 protein. More importantly, immunization of C57BL/6 mice with MTB41 DNA induced protection against infection with M. tuberculosis comparable to that induced by bacillus Calmette-Guerin. Thus, the use of a proven protective T cell line in conjunction with the T cell expression cloning approach resulted in the identification of a candidate Ag for a subunit vaccine against tuberculosis. PMID- 11120846 TI - Inhibition of TNF-alpha produced by Kupffer cells protects against the nonspecific liver toxicity of immunotoxin anti-Tac(Fv)-PE38, LMB-2. AB - LMB-2 (anti-Tac(Fv)-PE38) is a recombinant immunotoxin composed of the Fv fragment of the anti-Tac Ab fused to a 38-kDa form of Pseudomonas: exotoxin A. Recent clinical trials showed that LMB-2 is a promising agent for the treatment of patients with Tac-positive leukemia or lymphoma. One major side effect that needs to be overcome is nonspecific liver toxicity. In the current study, we have analyzed the mechanism of this toxicity using a mouse model. Mice that were injected with a lethal dose of LMB-2 showed severe hepatic necrosis. Immunohistochemistry revealed that LMB-2 accumulated in Kupffer cells in the liver, suggesting that the damage to the hepatocytes was indirect. When we examined the effects of LMB-2 on peritoneal macrophages, cells in the same lineage as Kupffer cells, we found that LMB-2 induced the production of TNF-alpha by these cells. Following LMB-2 administration to mice, the levels of TNF-alpha in the liver increased to very high levels, whereas the rise in serum levels was modest. In addition, the LMB-2-induced liver toxicity was blocked by a specific TNF binding protein (TNFsRp55). Liver toxicity was also blocked by indomethacin, which also blocked the rise of TNF-alpha in the liver. Both TNFsRp55 and indomethacin treatment protected mice against a lethal dose of LMB-2. These data indicate that TNF-alpha produced in the liver by Kupffer cells has an important causal role in the nonspecific liver toxicity of LMB-2. These findings have important clinical implications for the use of immunotoxins in the therapy of patients with cancer. PMID- 11120847 TI - Molecular mechanisms of high-dose antigen therapy in experimental autoimmune encephalomyelitis: rapid induction of Th1-type cytokines and inducible nitric oxide synthase. AB - High-dose Ag administration induces apoptotic death of autoreactive T cells and is an effective therapy of experimental autoimmune diseases of the nervous system. To explore the role of cytokines in Ag-specific immunotherapy, we analyzed mRNA induction and protein expression for the proinflammatory cytokines TNF-alpha and IFN-gamma, the anti-inflammatory cytokine IL-10, and the cytokine inducible NO synthase (iNOS) during high-dose Ag therapy of adoptive transfer experimental autoimmune encephalomyelitis (AT-EAE) in the Lewis rat. Using semiquantitative and competitive RT-PCR, we found 5- to 6-fold induction of TNF alpha mRNA and 3-fold induction of IFN-gamma mRNA in the spinal cord that occurred within 1 h after i.v. injection of Ag and was accompanied by a 2-fold increase of iNOS mRNA. Both IFN-gamma and iNOS mRNA remained elevated for at least 6 h, whereas TNF-alpha mRNA was already down-regulated 6 h after Ag injection. A comparable time course was found for circulating serum levels of TNF alpha and IFN-gamma. IL-10 mRNA levels did not change significantly following Ag injection. Neutralization of TNF-alpha by anti-TNF-alpha antiserum in vivo led to a significant decrease in the rate of T cell and oligodendrocyte apoptosis induced by high-dose Ag administration, but did not change the beneficial clinical effect of Ag therapy. Our data suggest profound activation of proinflammatory but not of anti-inflammatory cytokine gene expression by high dose Ag injection. Functionally, TNF-alpha contributes to increased apoptosis of both autoaggressive T cells and oligodendrocytes in the target organ and may thereby play a dual role in this model of Ag-specific therapy of CNS autoimmune diseases. PMID- 11120848 TI - L-selectin facilitates emigration and extravascular locomotion of leukocytes during acute inflammatory responses in vivo. AB - L-selectin has been shown to be important in mediating leukocyte recruitment during inflammatory responses. Although there are numerous in vitro studies demonstrating that engagement of L-selectin leads to the activation of several signaling pathways potentially contributing to subsequent adhesion, emigration, or even migration through the interstitium, whether this actually induces cellular events in vivo is completely unknown. Therefore, we used intravital microscopy to visualize the role of L-selectin in downstream leukocyte adhesion, emigration, and interstitial migration events in wild-type and L-selectin deficient (L-selectin(-/-)) mice. The cremaster muscle was superfused with the chemotactic inflammatory mediators platelet-activating factor or KC. Leukocyte rolling, adhesion, and emigration in postcapillary venules were examined, and the migration of emigrated leukocytes was recorded continuously using time-lapse videomicroscopy. Platelet-activating factor increased leukocyte adhesion to a similar level in both wild-type and L-selectin(-/-) mice. In contrast, both the number of emigrated leukocytes and the distance of extravascular migration were significantly reduced in L-selectin(-/-) mice. A similar pattern was observed in response to the superfusion of KC. Because superfusion of these mediators induced chemokinesis, we developed a new in vivo chemotaxis assay using slow release of KC from an agarose gel positioned 350 microm from a postcapillary venule. These experiments showed that L-selectin(-/-) leukocytes were also severely impaired in their ability to respond to a directional cue. These findings indicate that L selectin is important in enabling leukocytes to respond effectively to chemotactic stimuli in inflamed tissues. PMID- 11120849 TI - Tat protein is an HIV-1-encoded beta-chemokine homolog that promotes migration and up-regulates CCR3 expression on human Fc epsilon RI+ cells. AB - Human basophils and mast cells express the chemokine receptor CCR3, which binds the chemokines eotaxin and RANTES. HIV-1 Tat protein is a potent chemoattractant for basophils and lung mast cells obtained from healthy individuals seronegative for Abs to HIV-1 and HIV-2. Tat protein induced a rapid and transient Ca(2+) influx in basophils and mast cells, analogous to beta-chemokines. Tat protein neither induced histamine release from human basophils and mast cells nor increased IL-3-stimulated histamine secretion from basophils. The chemotactic activity of Tat protein was blocked by preincubation of FcepsilonRI(+) cells with anti-CCR3 Ab. Preincubation of Tat with a mAb anti-Tat (aa 1-86) blocked the migration induced by Tat. In contrast, a mAb specific for the basic region (aa 46 60) did not inhibit the chemotactic effect of Tat protein. Tat protein or eotaxin desensitized basophils to a subsequent challenge with the autologous or the heterologous stimulus. Preincubation of basophils with Tat protein up-regulated the level of CCR3 mRNA and the surface expression of the CCR3 receptor. Tat protein is the first identified HIV-1-encoded beta-chemokine homologue that influences the directional migration of human FcepsilonRI(+) cells and the expression of surface receptor CCR3 on these cells. PMID- 11120850 TI - Vitamin C inhibits NF-kappa B activation by TNF via the activation of p38 mitogen activated protein kinase. AB - The transcription factor NF-kappaB is a central mediator of altered gene expression during inflammation, and is implicated in a number of pathologies, including cancer, atherosclerosis, and viral infection. We report in this study that vitamin C inhibits the activation of NF-kappaB by multiple stimuli, including IL-1 and TNF in the endothelial cell line ECV304 and in primary HUVECs. The induction of a NF-kappaB-dependent gene, IL-8, by TNF was also inhibited. The effect requires millimolar concentrations of vitamin C, which occur intracellularly in vivo, particularly during inflammation. Vitamin C was not toxic to cells, did not inhibit another inducible transcription factor, STAT1, and had no effect on the DNA binding of NF-kappaB. Inhibition by vitamin C was not simply an antioxidant effect, because redox-insensitive pathways to NF-kappaB were also blocked. Vitamin C was shown to block IL-1- and TNF-mediated degradation and phosphorylation of I-kappaBalpha (inhibitory protein that dissociates from NF-kappaB), due to inhibition of I-kappaB kinase (IKK) activation. Inhibition of TNF-driven IKK activation was mediated by p38 mitogen activated protein kinase, because treatment of cells with vitamin C led to a rapid and sustained activation of p38, and the specific p38 inhibitor SB203580 reversed the inhibitory effect of vitamin C on IKK activity, I-kappaBalpha phosphorylation, and NF-kappaB activation. The results identify p38 as an intracellular target for high dose vitamin C. PMID- 11120851 TI - Endotoxin activation of macrophages does not induce ATP release and autocrine stimulation of P2 nucleotide receptors. AB - Receptors for extracellular nucleotides (P2, or purinergic receptors) have previously been implicated in the transduction of endotoxin signaling in macrophages. The most compelling evidence has been the observation that inhibitors of ionotropic nucleotide (P2X) receptors, including periodate-oxidized ATP (oATP), attenuate a subset of endotoxin-induced effects such as activation of NF-kappaB and up-regulation of inducible NO synthase. We investigated whether endotoxin induces ATP release from a murine macrophage cell line (BAC1.2F5) using sensitive on-line assays for extracellular ATP. These cells constitutively released ATP, producing steady-state extracellular concentrations of approximately 1 nM when assayed as monolayers of 10(6) adherent cells bathed in 1 ml of medium. However, the macrophages did not release additional ATP during either acute or prolonged endotoxin stimulation. In addition, cellular ecto ATPase activities were measured following prolonged endotoxin activation and were found not to be significantly altered. Although oATP treatment significantly attenuated the endotoxin-induced production of NO, this inhibitory effect was not reproduced when the cells were coincubated with apyrase, a highly effective ATP scavenger. These results indicate that activation of macrophages by endotoxin does not induce autocrine stimulation of P2 nucleotide receptors by endogenous ATP released to extracellular compartments. Moreover, the data suggest that the ability of oATP to interfere with endotoxin signaling is due to its interaction with molecular species other than ATP-binding P2 receptors. PMID- 11120852 TI - Regulation of IL-6 and IL-8 expression in rheumatoid arthritis synovial fibroblasts: the dominant role for NF-kappa B but not C/EBP beta or c-Jun. AB - Rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) produce IL-6 and IL 8, which contribute to inflammation and joint damage. The promoters of both cytokines possess binding sites for NF-kappaB, C/EBPbeta, and c-Jun, but the contribution of each to the regulation of IL-6 and IL-8 in RA FLS is unknown. We employed adenoviral-mediated gene delivery of a nondegradable IkappaBalpha, or dominant-negative versions of C/EBPbeta or c-Jun, to determine the contribution of each transcription factor to IL-6 and IL-8 expression. Inhibition of NF-kappaB activation significantly reduced the spontaneous and IL-1beta-induced secretion of IL-6 and IL-8 by RA FLS and the IL-1ss-induced production of IL-6 and IL-8 by human dermal fibroblasts. Inhibition of C/EBPbeta modestly reduced constitutive and IL-1beta-induced IL-6 by RA FLS, but not by human dermal fibroblasts, and had no effect on IL-8. Inhibition of c-Jun/AP-1 had no effect on the production of either IL-6 or IL-8. Employing gel shift assays, NF-kappaB, C/EBPbeta, and c-Jun were constitutively activated in RA FLS, but only NF-kappaB and c-Jun activity increased after IL-1beta. The reduction of cytokines by IkappaBalpha was mediated through inhibition of NF-kappaB activation, which resulted in decreased IL-6 and IL-8 mRNA. NF-kappaB was essential for IL-6 expression, because fibroblasts in which both NF-kappaB p50/p65 genes were deleted failed to express IL-6 in response to IL-1. These findings document the importance of NF-kappaB for the regulation of the constitutive and IL-1beta-stimulated expression of IL-6 and IL 8 by RA FLS and support the role of inhibition of NF-kappaB as a therapeutic goal in RA. PMID- 11120853 TI - The efficacy of immunotherapy in an experimental murine model of allergic asthma is related to the strength and site of T cell activation during immunotherapy. AB - In the present study, the relation between the efficacy of immunotherapy, and the strength and site of T cell activation during immunotherapy was evaluated. We used a model of allergic asthma in which OVA-sensitized and OVA-challenged mice display increased airway hyperresponsiveness, airway inflammation, and Th2 cytokine production by OVA-specific T cells. In this model, different immunotherapy strategies, including different routes of administration, or treatment with entire OVA or the immunodominant T cell epitope OVA(323-339), or treatment with a peptide analogue of OVA(323-339) with altered T cell activation capacity were studied. To gain more insight in how immunotherapy affects allergen specific T cells, the site of Ag-specific T cell activation and the magnitude of the T cell response induced during different immunotherapy strategies were determined using an adoptive transfer model. Our data suggest that amelioration of airway hyperresponsiveness and inflammation is associated with the induction of a strong, synchronized, and systemic T cell response, resulting in a decreased OVA-specific Th2 response. In contrast, deterioration of the disease after immunotherapy is associated with the induction of a weak nonsynchronized T cell response, resulting in the enhancement of the OVA-specific Th2 response after challenge. PMID- 11120854 TI - Chemokine production by G protein-coupled receptor activation in a human mast cell line: roles of extracellular signal-regulated kinase and NFAT. AB - Chemoattractants are thought to be the first mediators generated at sites of bacterial infection. We hypothesized that signaling through G protein-coupled chemoattractant receptors may stimulate cytokine production. To test this hypothesis, a human mast cell line (HMC-1) that normally expresses receptors for complement components C3a and C5a at low levels was stably transfected to express physiologic levels of fMLP receptors. We found that fMLP, but not C3a or C5a, induced macrophage inflammatory protein (MIP)-1ss (CCL4) and monocyte chemoattractant protein-1 (CCL2) mRNA and protein. Although fMLP stimulated both sustained Ca(2+) mobilization and phosphorylation of extracellular signal regulated kinase (ERK), these responses to C3a or C5a were transient. However, transient expression of C3a receptors in HMC-1 cells rendered the cells responsive to C3a for sustained Ca(2+) mobilization and MIP-1ss production. The fMLP-induced chemokine production was blocked by pertussis toxin, PD98059, and cyclosporin A, which respectively inhibit G(i)alpha activation, mitgen-activated protein kinase kinase-mediated ERK phosphorylation, and calcineurin-mediated activation of NFAT. Furthermore, fMLP, but not C5a, stimulated NFAT activation in HMC-1 cells. These data indicate that chemoattractant receptors induce chemokine production in HMC-1 cells with a selectivity that depends on the level of receptor expression, the length of their signaling time, and the synergistic interaction of multiple signaling pathways, including extracellular signal regulated kinase phosphorylation, sustained Ca(2+) mobilization and NFAT activation. PMID- 11120855 TI - Basophil responses to chemokines are regulated by both sequential and cooperative receptor signaling. AB - To investigate human basophil responses to chemokines, we have developed a sensitive assay that uses flow cytometry to measure leukocyte shape change as a marker of cell responsiveness. PBMC were isolated from the blood of volunteers. Basophils were identified as a single population of cells that stained positive for IL-3Ralpha (CDw123) and negative for HLA-DR, and their increase in forward scatter (as a result of cell shape change) in response to chemokines was measured. Shape change responses of basophils to chemokines were highly reproducible, with a rank order of potency: monocyte chemoattractant protein (MCP) 4 (peak at <1 nM) >/= eotaxin-2 = eotaxin-3 >/= eotaxin > MCP-1 = MCP-3 > macrophage-inflammatory protein-1alpha > RANTES = MCP-2 = IL-8. The CCR4 selective ligand macrophage-derived chemokine did not elicit a response at concentrations up to 10 nM. Blocking mAbs to CCR2 and CCR3 demonstrated that responses to higher concentrations (>10 nM) of MCP-1 were mediated by CCR3 rather than CCR2, whereas MCP-4 exhibited a biphasic response consistent with sequential activation of CCR3 at lower concentrations and CCR2 at 10 nM MCP-4 and above. In contrast, responses to MCP-3 were blocked only in the presence of both mAbs, but not after pretreatment with either anti-CCR2 or anti-CCR3 mAb alone. These patterns of receptor usage were different from those seen for eosinophils and monocytes. We suggest that cooperation between CCRs might be a mechanism for preferential recruitment of basophils, as occurs in tissue hypersensitivity responses in vivo. PMID- 11120856 TI - Up-regulation of the IL-12 receptor beta 2 chain in Crohn's disease. AB - Crohn' s disease (CD) is a chronic intestinal inflammatory disorder characterized by aberrant mucosal Th1 cell activation and production of IL-12, the major Th1 driving factor. The T cell response to IL-12 is dependent on the expression of a specific receptor composed of two subunits, termed IL-12Rbeta1 and IL-12Rbeta2. The content of IL-12Rbeta2, as measured at the mRNA level, is crucial in regulating Th1 differentiation. In this study we therefore investigated IL 12Rbeta2 RNA transcripts in CD. IL-12Rbeta2 expression was increased in active CD as well as Helicobacter pylori (HP)-associated gastritis and Salmonella colitis compared with that in inactive CD, ulcerative colitis, noninflammatory controls, and celiac disease. In contrast, IL-12Rbeta1 transcripts were expressed at comparable levels in all samples. In CD, IL-12Rbeta2 expression strictly correlated with tyrosine phosphorylation of STAT4, a key component of the IL-12 dependent Th1 polarization. This was associated with a pronounced expression of IFN-gamma. Transcripts for IL-12/p40 were detected in CD, HP-positive, and Salmonella colitis patients, but not in celiac disease, indicating that IL 12Rbeta2 up-regulation occurs only in IL-12-associated Th1 gastrointestinal diseases. Finally, we showed that stimulation of lamina propria mononuclear cells with IL-12 enhanced IL-12Rbeta2, suggesting that IL-12 regulates IL-12Rbeta2 expression in human gastrointestinal mucosa. The data show that the signaling pathway used by IL-12 to induce Th1 differentiation is increased at the site of disease in CD, further supporting the view that IL-12/IL-12R signals contribute to the inflammatory response in this condition. PMID- 11120857 TI - Evaluation of TNF-alpha and IL-1 blockade in collagen-induced arthritis and comparison with combined anti-TNF-alpha/anti-CD4 therapy. AB - We have evaluated the effects of anti-TNF-alpha, anti-IL-1, and combined anti-TNF alpha/anti-CD4 therapy in collagen-induced arthritis. Blockade of TNF-alpha or IL 1 before disease onset delayed, but did not prevent, the induction of arthritis. When treatment was initiated after onset of arthritis, anti-TNF-alpha, anti-IL 1beta, and anti-IL-1R (which blocks IL-1alpha and IL-1beta) were all found to be effective in reducing the severity of arthritis, with anti-IL-1R and anti-IL 1beta showing greater efficacy than anti-TNF-alpha. Anti-IL-1beta was equally as effective as anti-IL-1R, indicating that IL-1beta plays a more prominent role than IL-1alpha in collagen-induced arthritis. An additive effect was observed between anti-TNF-alpha and anti-IL-1R in the prevention of joint erosion and in normalization of the levels of serum amyloid P. Combined anti-TNF-alpha/anti-CD4 therapy also caused normalization of serum amyloid P levels. The therapeutic effect of anti-TNF-alpha plus anti-CD4 was comparable to that of anti-TNF-alpha plus anti-IL-1R, suggesting that combined anti-TNF-alpha/anti-CD4 therapy prevents both TNF-alpha- and IL-1-mediated pathology. Anti-TNF-alpha treatment reduced IL-1beta expression in the joint and, conversely, anti-IL-1beta treatment reduced TNF-alpha expression. Combined anti-TNF-alpha/anti-CD4 treatment almost completely blocked the expression of IL-1beta, thereby confirming the ability of this form of combination therapy to prevent IL-1ss-mediated pathology. PMID- 11120858 TI - Adenoviral transfer of cyclin-dependent kinase inhibitor genes suppresses collagen-induced arthritis in mice. AB - In rheumatoid synovial tissues, synovial fibroblasts are activated by proinflammatory cytokines and proliferate to develop hyperplastic pannus tissues, which irreversibly damage the affected joints. We recently reported that the cyclin-dependent kinase inhibitors p16(INK4a) and p21(Cip1) are not expressed in vivo in rheumatoid synovial fibroblasts, but are readily inducible in vitro. This observation was followed by the successful treatment of rat adjuvant arthritis by local p16(INK4a) gene transfer, showing that the inhibition of the cell cycle of the synovial cells ameliorates the arthritis. In this study, we show that another animal model of rheumatoid arthritis, murine collagen-induced arthritis, can be effectively treated by local gene transfer of p21(Cip1) as well as that of p16(INK4a). The anti-arthritic effects were observed even when the treatment was conducted after the arthritis had developed. Furthermore, the effects included suppression of the expression of proinflammatory cytokines such as IL-1ss, IL-6, and TNF-alpha. Our results demonstrate that the ectopic expression of cyclin dependent kinase inhibitors not only prevents synovial overgrowth but also ameliorates the proinflammatory milieu in the affected joints. The induction of p21(Cip1) in rheumatoid synovial tissues by pharmacological agents may also be an effective strategy to treat rheumatoid arthritis. PMID- 11120860 TI - Early requirement for B cells for development of spontaneous autoimmune thyroiditis in NOD.H-2h4 mice. AB - B cells are known to play an important role in the pathogenesis of several autoimmune diseases. NOD.H-2h4 mice develop spontaneous autoimmune thyroiditis (SAT) and anti-mouse thyroglobulin (MTg) autoantibodies, the levels of which correlate closely with the severity of thyroid lesions. NOD.H-2h4 mice genetically deficient in B cells (NOD.Kmu(null)) or rendered B cell-deficient by treatment from birth with anti-IgM develop minimal SAT. B cells were required some time in the first 4-6 wk after birth, because NOD.Kmu(null) or NOD.H-2h4 mice did not develop SAT when they were reconstituted with B cells as adults. The requirement for B cells was apparently not solely to produce anti-MTg autoantibodies, because passive transfer of anti-MTg Ab did not enable B cell deficient mice to develop SAT, and mice given B cells as adults produced autoantibodies but did not develop SAT. B cell-deficient mice developed SAT if their T cells developed from bone marrow precursors in the presence of B cells. Because B cells are required early in life and their function cannot be replaced by anti-MTg autoantibodies, B cells may be required for the activation or selection of autoreactive T cells. These autoreactive T cells are apparently unable to respond to Ag if B cells are absent in the first 4-6 wk after birth. PMID- 11120859 TI - Efficient simultaneous presentation of NY-ESO-1/LAGE-1 primary and nonprimary open reading frame-derived CTL epitopes in melanoma. AB - Recent studies have shown that CTL epitopes derived from tumor-associated Ags can be encoded by both primary and nonprimary open reading frames (ORF). In this study we have analyzed the HLA-A2-restricted CD8(+) T cell response to a recently identified CTL epitope derived from an alternative ORF product of gene LAGE-1 (named CAMEL), and the highly homologous gene NY-ESO-1 in melanoma patients. Using MHC/peptide tetramers we detected CAMEL(1-11)-specific CD8(+) T cells in peptide-stimulated PBMC as well as among tumor-infiltrated lymph node cells from several patients. Sorting and expansion of tetramer(+) CD8(+) T cells allowed the isolation of tetramer(bright) and tetramer(dull) populations that specifically recognized the peptide Ag with high and low avidity, respectively. Remarkably, only high avidity CAMEL-specific CTL were able to recognize Ag-expressing tumor cells. A large series of HLA-A2-positive melanoma cell lines was characterized for the expression of LAGE-1 and NY-ESO-1 mRNA and protein and tested for recognition by CAMEL-specific CTL as well as CTL that recognize a peptide (NY-ESO 1(157-165)) encoded by the primary ORF products of the LAGE-1 and NY-ESO-1 genes. This analysis revealed that tumor-associated CD8(+) T cell epitopes are simultaneously and efficiently generated from both primary and nonprimary ORF products of LAGE-1 and NY-ESO-1 genes and, importantly, that this occurs in the majority of melanoma tumors. These findings underscore the in vivo immunological relevance of CTL epitopes derived from nonprimary ORF products and support their use as candidate vaccines for inducing tumor specific cell-mediated immunity against cancer. PMID- 11120861 TI - Highly autoproliferative T cells specific for 60-kDa heat shock protein produce IL-4/IL-10 and IFN-gamma and are protective in adjuvant arthritis. AB - Previously we have shown that T cell responses to the mycobacterial 60-kDa heat shock protein (hsp60) peptide M256-270 mediated protection against adjuvant arthritis in Lewis rats. We have demonstrated now that M256-270-primed T cells become highly reactive to naive syngeneic APC upon repetitive restimulation in vitro with peptide M256-265, comprising the conserved core of peptide M256-270. These autoproliferative responses in the absence of added Ag were MHC class II restricted and resulted in the production of IL-4/IL-10 and IFN-gamma. Enhanced autoproliferation and expression of the cell surface molecule B7.2 by these T cells were observed in response to syngeneic heat-shocked APC, which indicated that the autoproliferation and expression of B7.2 resulted from the recognition of endogenously expressed and processed hsp. Despite their strong autoreactivity, upon transfer such T cells were found to induce a significant disease reduction in adjuvant arthritis. In contrast, T cells both primed and restimulated with peptide M256-270 became unresponsive toward syngeneic APC as well as toward the conserved core peptide M256-265, and they were devoid of protective capacity. This study demonstrates that the loss of self-tolerance toward hsp60 does not necessarily lead to autoimmune disease, but that hsp60-specific self-reactive and autoproliferative T cells may mediate T cell regulation in arthritis. PMID- 11120862 TI - The influence of HLA class I alleles and heterozygosity on the outcome of human T cell lymphotropic virus type I infection. AB - The inflammatory disease human T cell lymphotropic virus type I (HTLV-I) associated myelopathy (HAM/TSP) occurs in only 1-2% of HTLV-I-infected individuals and is associated with a high provirus load of HTLV-I. We hypothesize that a person's risk of developing HAM/TSP depends upon the efficiency of their immune response to the virus, which differs between individuals because of polymorphism in genes that influence this response. Previously we showed that the possession of HLA-A*02 was associated with a lower risk of HAM/TSP, and with a lower provirus load in healthy carriers of HTLV-I. However, HLA-A*02 did not account for all the observed difference in the risk of HAM/TSP. Here we present evidence, in the same study population in Japan, that HLA-Cw*08 was also associated with disease protection (probability value, two-tailed test = 0.002) and with a lower proviral load in healthy carriers. Possession of the A*02 and/or Cw*08 genes prevented 36% of potential HAM/TSP cases. In contrast, HLA-B*5401 was associated with a higher susceptibility to HAM/TSP (probability value, two-tailed test = 0.0003) and with a higher provirus load in HAM/TSP patients. At a given provirus load, B*5401 appeared to increase the risk of disease. The fraction of HAM/TSP cases attributable to B*5401 was 17%. Furthermore, individuals who were heterozygous at all three HLA class I loci have a lower HTLV-I provirus load than those who were homozygous at one or more loci. These results are consistent with the proposal that a strong class I-restricted CTL response to HTLV-I reduces the proviral load and hence the risk of disease. PMID- 11120863 TI - Direct detection and magnetic isolation of Chlamydia trachomatis major outer membrane protein-specific CD8+ CTLs with HLA class I tetramers. AB - We recently identified HLA class I-presented epitopes in the major outer membrane protein (MOMP) of Chlamydia trachomatis that elicit CTL responses in human genital tract infections. T cells possessing cytolytic activities specific for these epitopes could be detected following in vitro stimulation of peripheral blood CD8(+) T cells with peptides. In the present study we used HLA-A2 tetramers for detailed characterization of MOMP-specific CTL responses. Ex vivo tetramer analysis detected MOMP-specific T cells in the peripheral blood of infected individuals at significant frequencies (0.01-0.20% of CD8(+) T cells). After in vitro stimulation with peptides, the frequencies of MOMP peptide-specific T cells increased up to 2.34% of CD8(+) T cells in bulk cultures. In contrast, HLA A2/MOMP tetramer-binding T cells were virtually undetectable in the peripheral blood from uninfected individuals, either ex vivo or after 3 wk of in vitro peptide stimulation of their T cells. Magnetically sorted, tetramer-bound T cells specifically lysed peptide-pulsed targets as well as C. trachomatis-infected epithelial cells with nearly 50-fold greater per cell efficiency than that of unsorted populations. This study provides conclusive evidence of in vivo induction of HLA class I-restricted CD8(+) CTL responses to C. trachomatis MOMP. Direct detection of these cells with tetramers will allow their further characterization without prior manipulation and facilitate monitoring of CTL responses during infections and in immunization trials with MOMP-based vaccines. PMID- 11120864 TI - IL-12 treatment of endogenously arising murine brain tumors. AB - A number of recent studies have indicated that T cells can be stimulated to attack transplanted brain tumors in rodent models. As IL-12 has been shown to activate cytotoxic T cell responses, we tested the idea that it might stimulate a T cell response against endogenous brain tumors that arise in SV40 large T Ag transgenic mice (SV11). SV11 mice develop tumors of the choroid plexus, a specialization of the ependymal lining of the brain ventricles. They are a particularly relevant model of human disease, because they are immunocompetent but immunologically tolerant of the tumors. SV11 mice were treated with recombinant murine IL-12 for 10 days. Tumors grew more slowly than in control treated mice, and in some cases were reduced in size, as assessed by magnetic resonance imaging before and after treatment. At the end of treatment, tumors, but not brain parenchyma, exhibited extensive infiltration of activated CD8(+) and CD4(+) T cells. Tumors also showed a reduction in vascular density. Mice treated with IL-12 lived significantly longer than control mice. Tumors that progressed were nearly devoid of T cells, indicating that the T cell response was not sustained. In addition, some mice that had a substantial tumor burden at the beginning of treatment displayed evidence of immunosuppression, which might be related to TGF-ss2 detected in tumors. We conclude that IL-12 treatment can initiate an anti-tumor response even against endogenously arising brain tumors, but factors that will allow a sustained and more effective anti-tumor response need to be determined. PMID- 11120865 TI - Human and murine CD4 T cell reactivity to a complex antigen: recognition of the synthetic random polypeptide glatiramer acetate. AB - The capacity of glatiramer acetate (GA), a random copolymer of alanine, lysine, glutamic acid, and tyrosine to stimulate primary in vitro human and murine T cell proliferation was examined. PBMCs isolated from healthy humans and relapsing remitting multiple sclerosis patients and spleen cells from inbred strains of mice, expressing different H-2 haplotypes, were used as sources of non-GA-primed lymphocytes. GA functioned as a universal Ag, inducing dose-dependent proliferation of all non-GA-primed human and murine T cell populations tested. Moreover, GA stimulated PBMCs derived ex vivo from human cord blood, strongly suggesting that GA can activate both naive and memory T cells. The human T cell proliferative responses to GA were HLA class II DR-restricted by virtue of the ability of anti-class II Ab to inhibit T cell proliferation, and the demonstration that individual GA specific human T cell clones were HLA class II DR-restricted by either restriction element but not both. Furthermore, GA reactive T cells secreted Th0 cytokines and expressed a diverse repertoire of TCR. Limiting dilution analysis indicated that the T cell precursor frequency among the healthy human adults tested ranged from 1:5,000 to 1:125,000. Given that all of the T cell populations tested were isolated from non-GA-primed donors, it appears that virtually all humans and murine strains contain significant numbers of T cell populations cross-reactive with GA. These findings may explain the recent clinical finding that daily s.c. administration of GA ameliorates the progression of multiple sclerosis. PMID- 11120866 TI - Adoptive T cell immunotherapy of human uveal melanoma targeting gp100. AB - HLA-A*0201-restricted CTL against human gp100 were isolated from HLA-A*0201/K(b) (A2/K(b))-transgenic mice immunized with recombinant canarypox virus (ALVAC gp100). These CTL strongly responded to the gp100(154-162) epitope, in the context of both the chimeric A2/K(b) and the wild-type HLA-A*0201- molecule, and efficiently lysed human HLA-A*0201(+), gp100(+) melanoma cells in vitro. The capacity of the CTL to eradicate these tumors in vivo was analyzed in A2/K(b) transgenic transgenic mice that had received a tumorigenic dose of human uveal melanoma cells in the anterior chamber of the eye. This immune-privileged site offered the unique opportunity to graft xenogeneic tumors into immunocompetent A2/K(b)-transgenic mice, a host in which they otherwise would not grow. Importantly, systemic (i.v.) administration of the A2/K(b)-transgenic gp100(154 162)-specific CTL resulted in rapid elimination of the intraocular uveal melanomas, indicating that anti-tumor CTL are capable of homing to the eye and exerting their tumoricidal effector function. Flow cytometry analysis of ocular cell suspensions with HLA-A*0201-gp100(154-162) tetrameric complexes confirmed the homing of adoptively transferred CTL. Therefore, the immune-privileged state of the eye permitted the outgrowth of xenogeneic uveal melanoma cells, but did not protect these tumors against adoptive immunotherapy with highly potent anti tumor CTL. These data constitute the first direct indication that immunotherapy of human uveal melanoma may be feasible. PMID- 11120868 TI - Tyrosinase family proteins are antigens specific to Vogt-Koyanagi-Harada disease. AB - Vogt-Koyanagi-Harada (VKH) disease (and sympathetic ophthalmia) is an ocular inflammatory disease that is considered to be a cell-mediated autoimmune disease against melanocytes. The purpose of this study was to determine the Ags specific to VKH disease and to develop an animal model of VKH disease. We found that exposure of lymphocytes from patients with VKH disease to peptides (30-mer) derived from the tyrosinase family proteins led to significant proliferation of the lymphocytes. Immunization of these peptides into pigmented rats induced ocular and extraocular changes that highly resembled human VKH disease, and we suggest that an experimental VKH disease was induced in these rats. We conclude that VKH disease is an autoimmune disease against the tyrosinase family proteins. PMID- 11120867 TI - CD40 signaling replaces CD4+ lymphocytes and its blocking prevents chronic rejection of heart transplants. AB - Chronic rejection remains the major obstacle to long term survival in heart transplant recipients. The cellular and molecular mechanisms that underlie chronic rejection are not known, and their discovery can form the basis of clinical intervention. Several investigators have suggested that the development of chronic rejection in solid organ transplants is dependent on help mediated by CD4(+) lymphocytes. Importantly, the mechanism through which help is provided has not been fully delineated in transplant rejection. Using a murine heterotopic heart transplant model without immunosuppression, this study defines the functional role of CD4(+) lymphocytes in chronic rejection. In an MHC class II mismatched model, we demonstrate that chronic rejection was absolutely contingent on the presence of CD4(+) lymphocytes. Importantly, here we report that signaling through CD40 can replace the requirement of CD4(+) lymphocytes, demonstrated by the development of chronic rejection in CD4 knockout recipients treated with a CD40-activating mAb (FGK45). The return of rejection appears to be a CD8(+) lymphocyte-dependent process, noted by the absence of rejection in FGK45-treated recombinase-activated gene knockout (CD4(+) and CD8(+) lymphocyte-deficient) recipients. The CD40 signaling pathway works independently of B7-CD28 costimulation, as indicated by the development of severe chronic rejection in CD28 knockout recipients. Importantly, this study provides evidence that CD40 ligand-targeted therapies may prevent chronic rejection only in strain combinations where CD4(+) lymphocyte help is absolutely required. PMID- 11120870 TI - Ad hoc reviewers PMID- 11120869 TI - A mechanism for IL-10-mediated diabetes in the nonobese diabetic (NOD) mouse: ICAM-1 deficiency blocks accelerated diabetes. AB - Neonatal islet-specific expression of IL-10 in nonobese diabetic (NOD) mice accelerates the onset of diabetes, whereas systemic treatment of young NOD mice with IL-10 prevents diabetes. The mechanism for acceleration of diabetes in IL-10 NOD mice is not known. Here we show, by adoptive transfers, that prediabetic or diabetic NOD splenocytes upon encountering IL-10 in the pancreatic islets readily promoted diabetes. This outcome suggests that the compartment of exposure, not the timing, confers proinflammatory effects on this molecule. Moreover, injection of IL-10-deficient NOD splenocytes into transgenic IL-10-NOD.scid/scid mice elicited accelerated disease, demonstrating that pancreatic IL-10 but not endogenous IL-10 is sufficient for the acceleration of diabetes. Immunohistochemical analysis revealed hyperexpression of ICAM-1 on the vascular endothelium of IL-10-NOD mice. The finding suggests that IL-10 may promote diabetes via an ICAM-1-dependent pathway. We found that introduction of ICAM-1 deficiency into IL-10-NOD mice as well as into NOD mice prevented accelerated insulitis and diabetes. Failure to develop insulitis and diabetes was preceded by the absence of GAD65-specific T cell responses. The data suggest that ICAM-1 plays a role in the formation of the "immunological synapse", thereby affecting the generation and/or expansion of islet-specific T cells. In addition, ICAM-1 also played a role in the effector phase of autoimmune diabetes because adoptive transfer of diabetogenic BDC2.5 T cells failed to elicit clinical disease in ICAM 1-deficient IL-10-NOD and NOD mice. These findings provide evidence that pancreatic IL-10 is sufficient to drive pathogenic autoimmune responses and accelerates diabetes via an ICAM-1-dependent pathway. PMID- 11120871 TI - APO E gene and gene-environment effects on plasma lipoprotein-lipid levels. AB - Apolipoprotein E (apo E) is important in plasma lipid metabolism and is a component of several plasma lipoprotein-lipid particles. Three major apo E isoforms are encoded by three common alleles at the APO E locus. The E2 allele is associated with lower and the E4 allele with higher total plasma cholesterol and LDL cholesterol levels compared with the E3 allele. Available data generally indicate that APO E2, and possibly E3, genotype individuals reduce plasma total and low-density lipoprotein (LDL) cholesterol levels more than APO E4 individuals with statin therapy. Some evidence also indicates that APO E2 individuals are more likely to respond favorably to gemfibrozil and cholestyramine. On the other hand, it appears that with probucol, APO E4 genotype individuals may improve plasma lipoprotein-lipid profiles more than APO E3 individuals. APO E2 and E3 genotype perimenopausal women appear to improve plasma lipoprotein-lipid profiles more with hormone replacement therapy than APO E4 women. On the other hand, low fat diet interventions tend to reduce plasma LDL cholesterol and, perhaps, plasma total cholesterol levels more in APO E4 than in APO E2 or E3 individuals. Both cross-sectional and longitudinal studies generally indicate that APO E2 and E3 individuals improve plasma lipoprotein-lipid profiles more with exercise training than APO E4 individuals. Although these data are hardly definitive, they lend strong support for the possibility that in the near future individuals will be directed to what might be their optimal therapy for improving plasma lipoprotein lipid profiles and cardiovascular disease risk based partially on APO E genotype. PMID- 11120872 TI - Analysis of molecular profile data using generative and discriminative methods. AB - A modular framework is proposed for modeling and understanding the relationships between molecular profile data and other domain knowledge using a combination of generative (here, graphical models) and discriminative [Support Vector Machines (SVMs)] methods. As illustration, naive Bayes models, simple graphical models, and SVMs were applied to published transcription profile data for 1,988 genes in 62 colon adenocarcinoma tissue specimens labeled as tumor or nontumor. These unsupervised and supervised learning methods identified three classes or subtypes of specimens, assigned tumor or nontumor labels to new specimens and detected six potentially mislabeled specimens. The probability parameters of the three classes were utilized to develop a novel gene relevance, ranking, and selection method. SVMs trained to discriminate nontumor from tumor specimens using only the 50-200 top-ranked genes had the same or better generalization performance than the full repertoire of 1,988 genes. Approximately 90 marker genes were pinpointed for use in understanding the basic biology of colon adenocarcinoma, defining targets for therapeutic intervention and developing diagnostic tools. These potential markers highlight the importance of tissue biology in the etiology of cancer. Comparative analysis of molecular profile data is proposed as a mechanism for predicting the physiological function of genes in instances when comparative sequence analysis proves uninformative, such as with human and yeast translationally controlled tumour protein. Graphical models and SVMs hold promise as the foundations for developing decision support systems for diagnosis, prognosis, and monitoring as well as inferring biological networks. PMID- 11120873 TI - Integrating naive Bayes models and external knowledge to examine copper and iron homeostasis in S. cerevisiae. AB - A novel suite of analytical techniques and visualization tools are applied to 78 published transcription profiling experiments monitoring 5,687 Saccharomyces cerevisiae genes in studies examining cell cycle, responses to stress, and diauxic shift. A naive Bayes model discovered and characterized 45 classes of gene profile vectors. An enrichment measure quantified the association between these classes and specific external knowledge defined by four sets of categories to which genes can be assigned: 106 protein functions, 5 stages of the cell cycle, 265 transcription factors, and 16 chromosomal locations. Many of the 38 genes in class 42 are known to play roles in copper and iron homeostasis. The 17 uncharacterized open reading frames in this class may be involved in similar homeostatic processes; human homologs of two of them could be associated with as yet undefined disease states arising from aberrant metal ion regulation. The Met4, Met31, and Met32 transcription factors may play a role in coregulating genes involved in copper and iron metabolism. Extensions of the simple graphical model used for clustering to learning more complex models of genetic networks are discussed. PMID- 11120874 TI - Abrogation of thrombin-induced increase in pulmonary microvascular permeability in PAR-1 knockout mice. AB - We investigated the function of proteinase-activated receptor-1 (PAR-1) in the regulation of pulmonary microvascular permeability in response to thrombin challenge using PAR-1 knockout mice (-/-). Lungs were isolated and perfused with albumin (5 g/100 ml)-Krebs solution at constant flow (2 ml/min). Lung wet weight and pulmonary artery pressure (P(pa)) were continuously monitored. We determined the capillary filtration coefficient (K(fc)) and (125)I-labeled albumin (BSA) permeability-surface area product (PS) to assess changes in pulmonary microvessel permeability to liquid and albumin, respectively. Normal and PAR-1-null lung preparations received in the perfusate: 1) thrombin or 2) selective PAR-1 agonist peptide (TFLLRNPNDK-NH(2)). In control PAR-1 (+/+) mouse lungs, (125)I-albumin PS and K(fc) were significantly increased over baseline (by approximately 7- and 1.5 fold, respectively) within 20 min of alpha-thrombin (100 nM) challenge. PAR-1 agonist peptide (5 microM) gave similar results, whereas control peptide (5 microM; FTLLRNPNDK-NH(2)) was ineffective. At relatively high concentrations, thrombin (500 nM) or PAR-1 agonist peptide (10 microM) also induced increases in P(pa) and lung wet weight. All effects of thrombin (100 or 500 nM) or PAR-1 agonist peptide (5 or 10 microM) were prevented in PAR-1-null lung preparations. Baseline measures of microvessel permeability and P(pa) in the PAR-1-null preparations were indistinguishable from those in normal lungs. Moreover, PAR-1 null preparations gave normal vasoconstrictor response to thromboxane analog, U 46619 (100 nM). The results indicate that the PAR-1 receptor is critical in mediating the permeability-increasing and vasoconstrictor effects of thrombin in pulmonary microvessels. PMID- 11120875 TI - Quantitative expression analysis of the cellular specificity of HECT-domain ubiquitin E3 ligases. AB - We evaluated the expression of 28 gene sequences with homology to the carboxy terminal of HECT E3 ubiquitin ligases in nine human cell lines using RT-PCR, to determine whether gene expression could be associated with cell-specific functions (HECT is "homologous to E6AP C-terminus"). In general, HECT-domain E3 ligases are constitutively expressed at low levels with a broad range between cell types. hecth3, 21, and 23 had higher levels in three leukocytic lines (Jurkat, MM6, THP1); hecth11 was more abundant in HepG2 and A495; and hecth15 and hecth12 were differentially expressed in lung fibroblasts derived from normal and severe emphysema patients (CCD16 and CCD29, respectively). Absolute quantitation showed that most HECT E3s have about 20-100 copies of mRNA per Jurkat cell. By comparison, UBCH7 (an ubiquitin-conjugating E2) is 10-fold more abundant in Jurkat cells and 30-fold more abundant than E2 UBCH5A. We interpret the broad range of transcript levels to be consistent with the hypothesis that the concentrations of E3 are important for ubiquitination selectivity, leading us to conclude that substrate activation is necessary but not sufficient for selectivity. PMID- 11120876 TI - Gene expression profiling of mouse postnatal cerebellar development. AB - Expression patterns of 1,869 genes were determined using adapter-tagged competitive PCR (ATAC-PCR) at 6 time points during mouse postnatal cerebellar development. The expression patterns were classified into 12 clusters that were further assembled into 3 groups by hierarchical cluster analysis. Among the 1,869 genes, 1,053 known genes were assigned to 90 functional categories. Statistically significant correlation between the clusters or groups of gene expression and the functional categories was ascertained. Genes involved in oncogenesis or protein synthesis were highly expressed during the earlier stages of development. Those responsible for brain functions such as neurotransmitter receptor and synapse components were more active during the later stages of development. Many other genes also showed expression patterns in accordance with literature information. The gene expression patterns and the inferred functions were in good agreement with anatomical as well as physiological observations made during the developmental process. PMID- 11120879 TI - Shaking up glycolysis: Sustained, high lactate flux during aerobic rattling. AB - Substantial ATP supply by glycolysis is thought to reflect cellular anoxia in vertebrate muscle. An alternative hypothesis is that the lactate generated during contraction reflects sustained glycolytic ATP supply under well-oxygenated conditions. We distinguished these hypotheses by comparing intracellular glycolysis during anoxia to lactate efflux from muscle during sustained, aerobic contractions. We examined the tailshaker muscle of the rattlesnake because of its uniform cell properties, exclusive blood circulation, and ability to sustain rattling for prolonged periods. Here we show that glycolysis is independent of the O(2) level and supplies one-third of the high ATP demands of sustained tailshaking. Fatigue is avoided by rapid H(+) and lactate efflux resulting from blood flow rates that are among the highest reported for vertebrate muscle. These results reject the hypothesis that glycolysis necessarily reflects cellular anoxia. Instead, they demonstrate that glycolysis can provide a high and sustainable supply of ATP along with oxidative phosphorylation without muscle fatigue. PMID- 11120878 TI - Increased sensitivity to thyroid hormone in mice with complete deficiency of thyroid hormone receptor alpha. AB - Only three of the four thyroid hormone receptor (TR) isoforms, alpha1, beta1, and beta2, bind thyroid hormone (TH) and are considered to be true TRs. TRalpha2 binds to TH response elements on DNA, but its role in vivo is still unknown. We produced mice completely deficient in TRalpha (TRalpha(o/o)) that maintain normal serum thyroid-stimulating hormone (TSH) concentration despite low serum thyroxine (T(4)), suggesting increased sensitivity to TH. We therefore examined the effects of TH (L-3,3',5-triiodothyronine, L-T3) given to TH-deprived and to intact TRalpha(o/o) mice. Controls were wild-type (WT) mice of the same strain and mice resistant to TH due to deficiency in TRbeta (TRbeta(-/-)). In liver, T3 produced significantly greater responses in TRalpha(o/o) and smaller responses in TRbeta( /-) as compared with WT mice. In contrast, cardiac responses to L-T3 were absent or reduced in TRalpha(o/o), whereas they were similar in WT and TRbeta(-/-) mice, supporting the notion that TRalpha1 is the dominant TH-dependent TR isoform in heart. 5-Triiodothyronine (L-T3) given to intact mice produced a greater suppression of serum T(4) in TRalpha(o/o) than it did in WT mice and reduced by a greater amount the TSH response to TSH-releasing hormone. This is an in vivo demonstration that a TR deficiency can enhance sensitivity to TH. This effect is likely due to the abrogation of the constitutive "silencing" effect of TRalpha2 in tissues expressing the TRbeta isoforms. PMID- 11120880 TI - The chondrogenic transcription factor Sox9 is a target of signaling by the parathyroid hormone-related peptide in the growth plate of endochondral bones. AB - In the growth plate of endochondral bones, parathyroid hormone (PTH)-related peptide (PTHrP) regulates the rate of chondrocyte maturation from prehypertrophic chondrocytes to hypertrophic chondrocytes. Using an antibody specific for Sox9 phosphorylated at serine 181 (S(181)), one of the two consensus protein kinase A phosphorylation sites of Sox9, we showed that the addition of PTHrP strongly increased the phosphorylation of SOX9 in COS7 cells transfected with both SOX9- and PTH/PTHrP receptor-expressing vectors. PTHrP also increased the SOX9 dependent activity of chondrocyte-specific enhancers in the gene for type II collagen (Col2a1) in transient transfection experiments. This increased enhancer activity did not occur with a Sox9 mutant harboring serine-to-alanine substitutions in its two consensus protein kinase A phosphorylation sites. Consistent with these results, PTHrP also increased Col2a1 mRNA levels in rat chondrosarcoma cells as well as 10T1/2 mesenchymal cells transfected with a PTH/PTHrP receptor expressing plasmid. No phosphorylation of Sox9 at S(181) was detected in prehypertrophic chondrocytes of the growth plate or any chondrocytes of PTH/PTHrP receptor null mutants. In contrast in wild-type mouse embryos, previous immunohistochemistry experiments indicated that Sox9 phosphorylated at S(181) was detected almost exclusively in chondrocytes of the prehypertrophic zone. Sox9, regardless of the phosphorylation state, was present in all chondrocytes of both genotypes except hypertrophic chondrocytes. Our results indicated that Sox9 is a target of PTHrP signaling in prehypertrophic chondrocytes in the growth plate. We hypothesize that Sox9 mediates at least some effects of PTHrP in the growth plate and that the PTHrP-dependent increased transcriptional activity of Sox9 helps maintain the chondrocyte phenotype of cells in the prehypertrophic zone and inhibits their maturation to hypertrophic chondrocytes. PMID- 11120881 TI - Costimulation via lymphocyte function-associated antigen 1 in the absence of CD28 ligation promotes anergy of naive CD4+ T cells. AB - The mechanisms controlling induction of anergy at the level of naive CD4+ T cells are poorly understood but thought to reflect limited contact with costimulatory molecules during T cell antigen receptor (TCR) ligation. To clarify this question, naive TCR transgenic CD4+ cells were exposed to specific peptide presented by transfected antigen-presenting cells (APC) expressing MHC class II molecules with defined accessory molecules. Significantly, culturing CD4(+) cells with APC expressing MHC II plus peptide alone elicited early TCR signaling but failed to induce either proliferation or anergy. Culture with APC expressing MHC II plus B7 molecules led to strong proliferation and T cell priming but no anergy. In marked contrast, conspicuous induction of anergy occurred after T cell culture with APC expressing MHC class II and intercellular adhesion molecule-1 (ICAM-1). Thus, at the level of naive CD4(+) cells, anergy induction appears to reflect selective contact with APC expressing ICAM-1 in the absence of B7. PMID- 11120882 TI - Acid-sensing ion channel 3 matches the acid-gated current in cardiac ischemia sensing neurons. AB - Cardiac afferents are sensory neurons that mediate angina, pain that occurs when the heart receives insufficient blood supply for its metabolic demand (ischemia). These neurons display enormous acid-evoked depolarizing currents, and they fire action potentials in response to extracellular acidification that accompanies myocardial ischemia. Here we show that acid-sensing ion channel 3 (ASIC3), but no other known acid-sensing ion channel, reproduces the functional features of the channel that underlies the large acid-evoked current in cardiac afferents. ASIC3 and the native channel are both especially sensitive to pH, interact similarly with Ca(2+), and gate rapidly between closed, open, and desensitized states. Particularly important is the ability of ASIC3 and the native channel to open at pH 7, a value reached in the first few minutes of a heart attack. The steep activation curve suggests that the channel opens when four protons bind. We propose that ASIC3, a member of the degenerin channel (of Caenorhabditis elegans)/epithelial sodium channel family of ion channels, is the sensor of myocardial acidity that triggers cardiac pain, and that it might be a useful pharmaceutical target for treating angina. PMID- 11120883 TI - Antisense-induced exon skipping and synthesis of dystrophin in the mdx mouse. AB - Duchenne muscular dystrophy (DMD) is a severe muscle wasting disease arising from defects in the dystrophin gene, typically nonsense or frameshift mutations, that preclude the synthesis of a functional protein. A milder, allelic version of the disease, Becker muscular dystrophy, generally arises from in-frame deletions that allow synthesis of a shorter but still semifunctional protein. Therapies to introduce functional dystrophin into dystrophic tissue through either cell or gene replacement have not been successful to date. We report an alternative approach where 2'-O-methyl antisense oligoribonucleotides have been used to modify processing of the dystrophin pre-mRNA in the mdx mouse model of DMD. By targeting 2'-O-methyl antisense oligoribonucleotides to block motifs involved in normal dystrophin pre-mRNA splicing, we induced excision of exon 23, and the mdx nonsense mutation, without disrupting the reading frame. Exon 23 skipping was first optimized in vitro in transfected H-2K(b)-tsA58 mdx myoblasts and then induced in vivo. Immunohistochemical staining demonstrated the synthesis and correct subsarcolemmal localization of dystrophin and gamma-sarcoglycan in the mdx mouse after intramuscular delivery of antisense oligoribonucleotide:liposome complexes. This approach should reduce the severity of DMD by allowing a dystrophic gene transcript to be modified, such that it can be translated into a Becker-dystrophin-like protein. PMID- 11120885 TI - 2-Aminopurine fluorescence quenching and lifetimes: role of base stacking. AB - 2-Aminopurine (2AP) is a fluorescent analog of guanosine and adenosine and has been used to probe nucleic acid structure and dynamics. Its spectral features in nucleic acids have been interpreted phenomenologically, in the absence of a rigorous electronic description of the context-dependence of 2AP fluorescence. Now, by using time-dependent density functional theory, we describe the excited state properties of 2AP in a B-form dinucleotide stacked with guanosine, adenosine, cytosine, or thymine. Calculations predict that 2AP fluorescence is quenched statically when stacked with purines, because of mixing of the molecular orbitals in the ground state. In contrast, quenching is predicted to be dynamic when 2AP is stacked with pyrimidines, because of formation of a low-lying dark excited state. The different quenching mechanisms will result in different experimentally measured fluorescence lifetimes and quantum yields. PMID- 11120884 TI - HPP1: a transmembrane protein-encoding gene commonly methylated in colorectal polyps and cancers. AB - Adenomas are the precursors of most colorectal cancers. Hyperplastic polyps have been linked to the subset of colorectal cancers showing DNA microsatellite instability, but little is known of their underlying genetic etiology. Using a strategy that isolates differentially methylated sequences from hyperplastic polyps and normal mucosa, we identified a 370-bp sequence containing the 5' untranslated region and the first exon of a gene that we have called HPP1. Rapid amplification of cDNA ends was used to isolate HPP1 from normal mucosa. Using reverse transcription-PCR, HPP1 was expressed in 28 of 30 (93%) normal colonic samples but in only seven of 30 (23%) colorectal cancers (P < 0.001). The 5' region of HPP1 included a CpG island containing 49 CpG sites, of which 96% were found to be methylated by bisulfite sequencing of DNA from colonic tumor samples. By COBRA analysis, methylation was detected in six of nine (66%) adenomas, 17 of 27 (63%) hyperplastic polyps, and 46 of 55 (84%) colorectal cancers. There was an inverse relationship between methylation level and mRNA expression in cancers (r = -0.67; P < 0.001), and 5-aza-2-deoxycytidine treatment restored HPP1 expression in two colorectal cancer cell lines. In situ hybridization of HPP1 indicated that expression occurs in epithelial and stromal elements in normal mucosa but is silenced in both cell types in early colonic neoplasia. HPP1 is predicted to encode a transmembrane protein containing follistatin and epidermal growth factor like domains. Silencing of HPP1 by methylation may increase the probability of neoplastic transformation. PMID- 11120886 TI - Three distinct types of GnRH receptor characterized in the bullfrog. AB - It has been proposed recently that two types of GnRH receptors (GnRHR) exist in a particular species. Here we present data demonstrating that at least three types of GnRHR are expressed in a single diploid species, the bullfrog. Three different cDNAs, encoding distinct types of bullfrog GnRHR (bfGnRHR-1, bfGnRHR-2, and bfGnRHR-3), were isolated from pituitary and hindbrain of the bullfrog. BfGnRHR-1 mRNA was expressed predominantly in pituitary, whereas bfGnRHR-2 and -3 mRNAs were expressed in brain. The bfGnRHR-1, bfGnRHR-2, and bfGnRHR-3 proteins have an amino acid identity of approximately 30% to approximately 35% with mammalian GnRHRs and approximately 40% to approximately 50% with nonmammalian GnRHRs. Interestingly, bfGnRHR-2 has an 85% amino acid homology with Xenopus GnRHR. Less than 53% amino acid identity was observed among the three bfGnRHRs. All isolated cDNAs encode functional receptors because their transient expression in COS-7 cells resulted in a ligand-dependent increase in inositol phosphate production. Notably, all three receptors exhibited a differential ligand selectivity. For all receptors, cGnRH-II has a higher potency than mGnRH. In addition, salmon GnRH also has a strikingly high potency to stimulate all three receptors. In conclusion, we demonstrated the presence of three GnRHRs in the bullfrog. Their expression in pituitary and brain suggests that bfGnRHRs play an important role in the regulation of reproductive functions in the bullfrog. PMID- 11120887 TI - A multiple-site-specific heteroduplex tracking assay as a tool for the study of viral population dynamics. AB - Rapidly evolving entities, such as viruses, can undergo complex genetic changes in the face of strong selective pressure. We have developed a modified heteroduplex tracking assay (HTA) capable of detecting the presence of single, specific mutations or sets of linked mutations. The initial application of this approach, termed multiple-site-specific (MSS) HTA, was directed toward the detection of mutations in the HIV-1 pro gene at positions 46, 48, 54, 82, 84, and 90, which are associated with resistance to multiple protease inhibitors. We demonstrate that MSS HTA is sensitive and largely specific to all targeted mutations. The assay allows the accurate and reproducible quantitation of viral subpopulations comprising 3% or more of the total population. Furthermore, we used MSS HTA in longitudinal studies of pro gene evolution in vitro and in vivo. In the examples shown here, populations turned over rapidly and more than one population was present frequently. To demonstrate the versatility of MSS HTA, we also constructed a probe sensitive to changes at positions 181 and 184 of the RT coding domain. Changes at these positions are involved in resistance to nevirapine and 2',3'-dideoxy-3'-thiacytidine (3TC), respectively. This assay easily detected the evolution of resistance to 3TC. MSS HTA provides a rapid and sensitive approach for detecting the presence of and quantifying complex mixtures of distinct genotypes, including genetically linked mutations, and, as one example, represents a useful tool for following the evolution of drug resistance during failure of HIV-1 antiviral therapy. PMID- 11120888 TI - Action potential propagation in mitral cell lateral dendrites is decremental and controls recurrent and lateral inhibition in the mammalian olfactory bulb. AB - In the mammalian main olfactory bulb (MOB), the release of glutamate from lateral dendrites of mitral cells onto the dendrites of granule cells evokes recurrent and lateral inhibition of mitral cell activity. Whole-cell voltage recordings in the mouse MOB in vivo and in vitro show that recurrent and lateral inhibition together control the number, duration, and onset of odor-evoked action potential (AP) firing in mitral cells. APs in mitral cells propagate into the lateral dendrites and evoke a transient increase in dendritic calcium concentration ([Ca2+]), which is decremental with distance from the soma, and increases with AP number. These results suggest that the extent of AP propagation in lateral dendrites of mitral cells, along with the concomitant dendritic Ca(2+) transient, controls the amplitude of lateral and recurrent inhibition and thus is a critical determinant of odor-specific AP patterns in the MOB. PMID- 11120889 TI - An anthelmintic compound, nafuredin, shows selective inhibition of complex I in helminth mitochondria. AB - Infections with parasitic helminths are important causes of morbidity and mortality worldwide. New drugs that are parasite specific and minimally toxic to the host are needed to counter these infections effectively. Here we report the finding of a previously unidentified compound, nafuredin, from Aspergillus niger. Nafuredin inhibits NADH-fumarate reductase (complexes I + II) activity, a unique anaerobic electron transport system in helminth mitochondria, at nM order. It competes for the quinone-binding site in complex I and shows high selective toxicity to the helminth enzyme. Moreover, nafuredin exerts anthelmintic activity against Haemonchus contortus in in vivo trials with sheep. Thus, our study indicates that mitochondrial complex I is a promising target for chemotherapy, and nafuredin is a potential lead compound as an anthelmintic isolated from microorganisms. PMID- 11120890 TI - Glutamic and aminoadipic semialdehydes are the main carbonyl products of metal catalyzed oxidation of proteins. AB - Metal-catalyzed oxidation results in loss of function and structural alteration of proteins. The oxidative process affects a variety of side amino acid groups, some of which are converted to carbonyl compounds. Spectrophotometric measurement of these moieties, after their reaction with 2,4-dinitrophenylhydrazine, is a simple, accurate technique that has been widely used to reveal increased levels of protein carbonyls in aging and disease. We have initiated studies aimed at elucidating the chemical nature of protein carbonyls. Methods based on gas chromatography/mass spectrometry with isotopic dilution were developed for the quantitation of glutamic and aminoadipic semialdehydes after their reduction to hydroxyaminovaleric and hydroxyaminocaproic acids. Analysis of model proteins oxidized in vitro by Cu2+/ascorbate revealed that these two compounds constitute the majority of protein carbonyls generated. Glutamic and aminoadipic semialdehydes were also detected in rat liver proteins, where they constitute approximately 60% of the total protein carbonyl value. Aminoadipic semialdehyde was also measured in protein extracts from HeLa cells, and its level increased as a consequence of oxidative stress to cell cultures. These results indicate that glutamic and aminoadipic semialdehydes are the main carbonyl products of metal catalyzed oxidation of proteins, and that this reaction is a major route leading to the generation of protein carbonyls in biological samples. PMID- 11120891 TI - Loss of imprinting of the insulin-like growth factor II gene occurs by biallelic methylation in a core region of H19-associated CTCF-binding sites in colorectal cancer. AB - We hypothesize that loss of imprinting (LOI) of the insulin-like growth factor II (IGF2) gene is associated with a predisposition to sporadic colorectal cancer. We confirmed a previously known strong correlation between LOI and microsatellite instability and showed that LOI was not a consequence of microsatellite instability or mismatch repair deficiency. LOI of IGF2 correlated strongly with biallelic hypermethylation of a core of five CpG sites in the insulator region of IGF2/H19, which is a known CTCF-binding element. As this methylation-dependent LOI was present in both tumors and normal colonic mucosa, it is possible that hypermethylation creates a field defect predisposing to cancer. PMID- 11120892 TI - Mechanisms governing dendritic gamma-aminobutyric acid (GABA) release in the rat olfactory bulb. AB - In the olfactory bulb, synaptic transmission between dendrites plays an important role in the processing of olfactory information. Glutamate released from the dendrites of principal mitral cells excites the dendritic spines of granule cells, which in turn release gamma-aminobutyric acid (GABA) back onto mitral cell dendrites. Slow N-methyl-d-aspartate (NMDA) receptors on granule dendrites are particularly effective in driving this reciprocal dendrodendritic inhibition (DDI), raising the possibility that calcium influx through NMDA receptors may trigger GABA exocytosis directly. In this study, I show that NMDA receptor activation is not an absolute requirement and that DDI can be evoked solely by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors when granule cell excitability is increased or under conditions that slow AMPA receptor kinetics. In physiological extracellular Mg(2+), DDI elicited by photolysis of caged calcium in mitral dendrites is blocked by cadmium and toxins to N- and P/Q-type voltage-gated calcium channels. DDI is largely unaffected after granule dendrites have been loaded with the slow calcium chelator EGTA, suggesting a tight coupling between the site of calcium influx and the release machinery governing GABA exocytosis. These results indicate that voltage-gated calcium channels play an essential role in dendritic GABA release during reciprocal feedback inhibition in the olfactory bulb. PMID- 11120893 TI - T7 RNA polymerase transcription complex: what you see is not what you get. PMID- 11120894 TI - Do smoking parents seek the best advice for their asthmatic children? PMID- 11120895 TI - Progress in ARDS research: a protection racket? PMID- 11120896 TI - Obesity is a risk for asthma and wheeze but not airway hyperresponsiveness. AB - BACKGROUND: A study was undertaken to assess whether the recent increases in prevalence of both asthma and obesity are linked and to determine if obesity is a risk factor for diagnosed asthma, symptoms, use of asthma medication, or airway hyperresponsiveness. METHODS: Data from 1971 white adults aged 17-73 years from three large epidemiological studies performed in NSW were pooled. Doctor diagnosis of asthma ever, history of wheeze, and medication use in the previous 12 months were obtained by questionnaire. Body mass index (BMI) in kg/m(2) was used as a measure of obesity. Airway hyperresponsiveness (AHR) was defined as dose of <3.9 micromol histamine required to provoke a fall in forced expiratory volume in one second (FEV(1)) of 20% or more (PD(20)FEV(1)). Adjusted odds ratios (OR) were obtained by logistic regression. RESULTS: After adjusting for atopy, age, sex, smoking history, and family history, severe obesity was a significant risk factor for recent asthma (OR 2. 04, p=0.048), wheeze in the previous 12 months (OR 2.6, p=0.001), and medication use in the previous 12 months (OR 2.83, p=0.005), but not for AHR (OR 0.87, p=0.78). FEV(1) and forced vital capacity (FVC) were significantly reduced in the group with severe obesity, but FEV(1)/FVC ratio, peak expiratory flow (PEF), and mid forced expiratory flow (FEF(25-75)) were not different from the group with normal BMI. The underweight group (BMI <18.5 kg/m(2)) had increased symptoms of shortness of breath, increased airway responsiveness, and reduced FEV(1), FVC, PEF, and FEF(25-75) with similar use of asthma medication as subjects in the normal weight range. CONCLUSIONS: Although subjects with severe obesity reported more wheeze and shortness of breath which may suggest a diagnosis of asthma, their levels of atopy, airway hyperresponsiveness, and airway obstruction did not support the suggestion of a higher prevalence of asthma in this group. The underweight group appears to have more significant respiratory problems with a higher prevalence of symptoms, reduced lung function, and increased airway responsiveness without an increase in medication usage. This group needs further investigation. PMID- 11120897 TI - Does passive smoking increase the frequency of health service contacts in children with asthma? AB - BACKGROUND: Passive smoking is a major cause of respiratory morbidity in children. However, few studies give accurate estimates of the health effects of passive smoking in children with asthma using an objective measure of exposure. The effects of passive smoking using salivary cotinine levels to measure exposure were investigated. METHODS: A sample of 438 children aged 2-12 years with asthma who had a parent who smoked were recruited in Tayside and Fife, Scotland. Health service contacts for asthma, assessed from GP case records, were used as a proxy for morbidity. RESULTS: A weak U-shaped relationship was found between the salivary cotinine level and health service contacts for asthma: compared with low cotinine levels those with moderate cotinine levels had a reduced contact rate (relative rate (RR) = 0.91, 95% confidence interval (CI) 0.80 to 1.05), whereas high cotinine levels were associated with an increased rate of contact (RR = 1.19, 95% CI 1.05 to 1.37). In contrast, a strong association was seen with the amount the parent reported smoking in front of the child: the higher the level the fewer visits were made for asthma (RR for everyday exposure = 0.66, 95% CI 0.56 to 0.77). This effect was not seen for non-respiratory visits. Demographic factors, age of child, and number of children in the family all had a powerful effect on the number of visits for asthma. The parents' perception of asthma severity was associated with visit frequency independent of actual severity (derived from drug treatment). CONCLUSION: High levels of parental smoking in the home are associated with a reduction in health care contacts for asthma. This could be due to a lack of awareness of asthma symptoms among heavy smokers or a reluctance to visit the GP. Children with asthma who have parents who smoke heavily may not be receiving adequate management. PMID- 11120898 TI - Glutathione in induced sputum of healthy individuals and patients with asthma. AB - BACKGROUND: Glutathione is central to the antioxidant defences of the lung. The aim of this study was to determine whether sputum induction can be used for the measurement of glutathione in the respiratory tract. METHODS: Saliva and induced sputum (3% NaCl, 20 minutes) samples were collected from 10 healthy individuals and 10 patients with stable asthma receiving treatment with inhaled corticosteroids. Samples were chilled on ice and dispersed by dilution with ice cold phosphate buffered saline and pipetting. Cell-free supernatants were obtained by centrifugation of samples and filtration of supernatants and analysed for total glutathione, glutathione disulfide, and albumin content. The cells were treated with dithiothreitol and cell numbers, cell viability, and differential cell patterns were determined. RESULTS: As judged by cell viability and percentage of non-squamous cells, adequate sputum samples were obtained from nine healthy and nine asthmatic subjects. The salivary total glutathione content was low (median concentration 1.2 microM (range 0.8-1.5) in healthy subjects and 0.9 microM (0.7-1. 2) in asthmatic subjects). The sputum total glutathione content of both healthy and asthmatic subjects was within the same range (3.9 (1.0-12.3) microM and 6.4 (1.3-19.2) microM, respectively; p=0.35). Surprisingly, and in marked contrast to results obtained with bronchoalveolar lavage, sputum levels of glutathione disulfide represented more than 50% of the total glutathione in both groups (50.9% (range 24.6-83.1) and 72.3% (range 36.5-97.4), respectively; p=0.2). CONCLUSIONS: The results of this study indicate that sputum induction can be used to measure the glutathione content of bronchial secretions. Sputum glutathione levels of stable asthmatic patients did not differ significantly from healthy controls. PMID- 11120899 TI - Enhancement of goblet cell hyperplasia and airway hyperresponsiveness by salbutamol in a rat model of atopic asthma. AB - BACKGROUND: Goblet cell hyperplasia (GCH) is a prominent feature in animal models of atopic asthma produced by immunisation and following multiple challenges with antigens. The aim of this study was to examine the effect of a beta(2) agonist on the development of GCH induced by the immune response. METHODS: Brown Norway rats were immunised and challenged with an aerosol of ovalbumin for four weeks. Salbutamol (0.5 mg/kg/day) or vehicle was continuously delivered for the four weeks using a subcutaneously implanted osmotic minipump. The density of goblet cells, other morphological changes, and airway responsiveness to methacholine were evaluated 24 hours after the final challenge. RESULTS: Treatment with salbutamol induced a more than twofold increase in the mean (SE) number of goblet cells (53.7 (7.3) vs 114.5 (11.8) cells/10(3) epithelial cells, p<0.01) while it did not significantly influence airway wall thickening and eosinophilic infiltration. Airway responsiveness to methacholine expressed as the logarithmic value of the concentration of methacholine required to generate a 50% increase in airway pressure (logPC(150)Mch) was also enhanced by the beta(2) agonist (-0.56 (0. 21) vs -0.95 (0.05), p<0.05). Additional experiments revealed that the same dose of the beta(2) agonist alone did not cause GCH in non-immunised rats and that the enhancement of GCH by salbutamol was completely abolished by simultaneous treatment with methylprednisolone (0.5 mg/kg/day). CONCLUSIONS: These data suggest that salbutamol enhances goblet cell hyperplasia and airway hyperresponsiveness in this rat model of atopic asthma. PMID- 11120900 TI - Ultrastructural examination of bronchial biopsy specimens from children with moderate asthma. AB - BACKGROUND: Few studies have evaluated the asthmatic airway in childhood. The aim of this study was to assess the histopathological changes occurring in the bronchi of children with moderate asthma using light and electron microscopy. METHODS: Bronchial biopsy specimens from 10 children with moderate asthma (seven boys) of mean (SD) age 9.3 (3.8) years (range 5-14) were examined by light and electron microscopy. Patients had not had a respiratory infection for at least one month and they had not been treated with steroids or sodium cromoglycate for four weeks before the study. Bronchoscopy was performed under general anaesthesia using a Karl Storz rigid paediatric bronchoscope. Biopsy materials were stained with uracyl acetate and lead citrate and evaluated under a Zeiss-10 electron microscope and light microscope. RESULTS: The most important finding was thickening and hyalinization of the basement membrane in nine patients. The ciliated epithelial cells showed loss of cilia in some cases. Overactive fibroblasts were consistently found. Six patients had degranulating mast cells and lymphocyte infiltration in the submucosa. Eosinophils were seen in only one biopsy sample. CONCLUSION: Children with moderate asthma develop bronchial inflammation similar to the reaction observed in adults. However, in our study the inflammation was rich in lymphocytes rather than eosinophils. PMID- 11120901 TI - Sputum and plasma endothelin-1 levels in exacerbations of chronic obstructive pulmonary disease. AB - BACKGROUND: Endothelin (ET)-l is a bronchoconstrictor peptide produced in the airways. It has been implicated in the pathogenesis of asthma and virally mediated airway inflammation and may play a role in exacerbations of chronic obstructive pulmonary disease (COPD). METHODS: Seventy one patients with COPD were followed prospectively and sampled for plasma and sputum ET-1 levels when stable and during an exacerbation. Sputum was also examined for cytokines, human rhinovirus, and Chlamydia pneumoniae. RESULTS: Plasma ET-1 levels were available for 67 patients with stable COPD (mean (SD) 0.58 (0.31) pg/ml); 28 pairs of stable-exacerbation plasma samples had a mean stable ET-1 level of 0.54 (0.30) pg/ml rising to 0.67 (0.35) pg/ml at exacerbation (mean difference 0.13, 95% confidence interval (CI) 0.04 to 0.21, p = 0.004). Plasma ET-1 levels in the 67 patients with stable COPD were inversely correlated with baseline forced expiratory volume in one second (FEV(1); r = -0. 29, p = 0.022) and forced vital capacity (FVC; r = -0.38, p = 0.002). The change in plasma ET-1 levels during an exacerbation correlated with the change in oxygen saturation (SaO(2); r = -0.41, p = 0.036). In 14 stable-exacerbation pairs of sputum samples median stable ET-1 levels were 5.37 (0.97-21.95) pg/ml rising to 34.68 (13.77-51.95) pg/ml during an exacerbation (mean difference 25.14, 95% CI 3.77 to 46.51, p = 0.028). This increase in sputum ET-1 levels correlated with the increase in plasma ET-1 levels (r = 0.917, p = 0.001) and sputum interleukin (IL)-6 levels (r = 0.718, p = 0.013). CONCLUSIONS: Sputum levels of ET-1 rise in COPD patients during an exacerbation and this is reflected by a smaller rise in plasma ET-1 levels. ET-1 may have a role in mediating airway inflammatory changes during exacerbations of COPD. PMID- 11120902 TI - Relationship between airway inflammation and the frequency of exacerbations in patients with smoking related COPD. AB - BACKGROUND: Patients with more frequent exacerbations of chronic obstructive pulmonary disease (COPD) may have increased bronchial inflammation. Airway inflammation was measured in patients who had been thoroughly investigated with full pulmonary function testing, thoracic HRCT scanning, and sputum microbiology to examine further the relationship between exacerbation frequency and bronchial inflammation. METHODS: Airway inflammation (spontaneous sputum sol phase myeloperoxidase (MPO), elastase, leukotriene (LT)B(4), interleukin (IL)-8, secretory leukoprotenase inhibitor (SLPI), protein leakage) and serum levels of C reactive protein (CRP) were compared in 40 patients with stable, smoking related COPD, divided into those with frequent (> or =3/year) or infrequent (< or =2/year) exacerbations according to the number of primary care consultations during the preceding year. The comparisons were repeated after excluding eight otherwise clinically indistinguishable patients who had tubular bronchiectasis on the HRCT scan. RESULTS: Patients with frequent (n=12) and infrequent (n=28) exacerbations were indistinguishable in terms of their clinical, pulmonary function, and sputum characteristics, CRP concentrations, and all of their bronchial inflammatory parameters (p>0.05). The patients without evidence of tubular bronchiectasis (n=32) were equally well matched but the sputum concentrations of SLPI were significantly lower in the frequent exacerbators (n=8) in this subset analysis (p<0.05). CONCLUSIONS: There are several clinical features that directly influence bronchial inflammation in COPD. When these were carefully controlled for, patients with more frequent reported exacerbations had lower sputum concentrations of SLPI. This important antiproteinase is also known to possess antibacterial and antiviral activity. Further studies are required into the nature of recurrent exacerbations and, in particular, the regulation and role of SLPI in affected individuals. PMID- 11120903 TI - Methacholine responsiveness in infants assessed with low frequency forced oscillation and forced expiration techniques. AB - BACKGROUND: The contribution of the pulmonary tissues to the mechanical behaviour of the respiratory system is well recognised. This study was undertaken to detect airway and lung tissue responses to inhaled methacholine (Mch) using the low frequency forced oscillation technique (LFOT). METHODS: The respiratory system impedance (Zrs, 0.5-20 Hz) was determined in 17 asymptomatic infants. A model containing airway resistance (Raw) and inertance (Iaw) and a constant phase tissue damping (G) and elastance (H) was fitted to Zrs data. Tissue hysteresivity (eta) was calculated as eta=G/H. The raised volume rapid thoracic compression technique (RVRTC) was used to generate forced expiratory volume in 0.5 seconds (FEV(0.5)). Lung function was determined at baseline and following inhaled Mch in doubling doses (0.25-16 mg/ml) until the maximal dose was reached or a fall of 15% in FEV(0.5) was achieved (PC(15)FEV(0.5)). The response to Mch was defined in terms of the concentration of Mch provoking a change in lung function parameters of more than two standard deviation units (threshold concentration). RESULTS: At PC(15)FEV(0.5) a response in Raw, Iaw, G, and eta, but not H, was detected (mean (SE) 61.28 (12.22)%, 95.43 (34.31)%, 46.28 (22.36)%, 44.26 (25.83)%, and -6.48 (4.94)%, respectively). No significant differences were found between threshold concentrations of LFOT parameters and FEV(0.5). CONCLUSIONS: Inhaled Mch alters both airway and respiratory tissue mechanics in infants. PMID- 11120904 TI - Effects of beta-carotene supplementation for six months on clinical and laboratory parameters in patients with cystic fibrosis. AB - BACKGROUND: Patients with cystic fibrosis (CF) have significantly decreased plasma concentrations of nutrient antioxidant vitamins, especially of beta carotene, which is thought to result from fat malabsorption and chronic pulmonary inflammation. The aim of this double blind, placebo controlled study was to investigate the effect of oral beta-carotene supplementation for six months on clinical parameters. METHODS: Twenty four patients with CF were randomised to receive beta-carotene 1 mg/kg/day (maximum 50 mg/day) for three months (high dose supplementation) and 10 mg/day for a further three months (low dose supplementation) or placebo. At monthly follow up visits the plasma beta-carotene concentration, total antioxidant capacity, malondialdehyde (MDA) as a marker of lipid peroxidation, and clinical parameters (Shwachmann-Kulczycki score, body mass index (BMI), height, and lung function (FEV(1))) were assessed. The number of pulmonary exacerbations requiring antibiotic treatment (in days) three months before and during the study were evaluated. RESULTS: The plasma concentration of beta-carotene increased significantly to the normal range during the three months of high dose supplementation (baseline 0.08 (0.04) micromol/l to 0.56 (0.38) micromol/l; p<0.001) but decreased to 0.32 (0.19) micromol/l in the period of low dose supplementation. Initially raised plasma levels of MDA fell to normal levels and the total antioxidant capacity showed a non-significant trend towards improvement during high dose supplementation. Antibiotic treatment decreased significantly in the supplementation group from 14.5 (14.9) days/patient during the three months before the study to 9.8 (10.3) days/patient during high dose supplementation (p=0.0368) and to 10.5 (9.9) days/patient during low dose supplementation, but increased in the placebo group. The Shwachmann-Kulczycki score, lung function, and BMI did not show any changes in either of the treatment groups. No adverse events were observed during the study period. CONCLUSION: Oral beta-carotene supplementation in a dose of 1 mg/kg/day only was effective in normalising the plasma concentration of beta-carotene and resulted in a decrease in pulmonary exacerbations. These data suggest that patients with CF may benefit clinically from supplementation with beta-carotene and further studies are warranted. PMID- 11120905 TI - Association of alpha(1)-antichymotrypsin deficiency with milder lung disease in patients with cystic fibrosis. AB - BACKGROUND: Cystic fibrosis (CF) is characterised by an excess of free proteinases that destroy lung tissue. Despite this, previous studies have shown that patients with CF with a mild deficiency variant of the proteinase inhibitor alpha(1)-antitrypsin have less, rather than more, severe pulmonary disease. Alpha(1)-antichymotrypsin is another important serine proteinase inhibitor that protects the lung against proteolytic attack, and point mutations in the alpha(1) antichymotrypsin gene that result in plasma deficiency are associated with chronic obstructive pulmonary disease. METHODS: The effect of alpha(1) antichymotrypsin deficiency and the -15 alpha(1)-antichymotrypsin signal peptide genotype on lung function was assessed in patients with CF. RESULTS: One hundred and fifty seven patients with CF were screened and 10 were identified with a plasma deficiency of alpha(1)-antichymotrypsin (plasma concentration <0.2 g/l). In a multivariate analysis these individuals had significantly less severe lung disease than those who had normal or raised levels of alpha(1)-antichymotrypsin: forced expiratory volume in one second (FEV(1)) 69.9% predicted versus 53. 2% predicted (p=0.04) and chest radiographic score of 7.2 versus 9.7 (p=0.03) for those with and without alpha(1)-antichymotrypsin deficiency, respectively. The 15 signal peptide genotype did not affect plasma levels, but the -15 Ala/Ala signal peptide genotype was over-represented in individuals with CF compared with healthy blood donor controls. CONCLUSION: These data indicate that deficiency of alpha(1)-antichymotrypsin is associated with less severe pulmonary disease in patients with CF, and support our previous observations that mild genetic deficiency of a proteinase inhibitor is associated with an improved outcome. PMID- 11120906 TI - Prediction of pulmonary complications after a lobectomy in patients with non small cell lung cancer. AB - BACKGROUND: Although the preoperative prediction of pulmonary complications after lung major surgery has been reported in various papers, it still remains unclear. METHODS: Eighty nine patients with stage I-IIIA non-small cell lung cancer (NSCLC) who underwent a complete resection at our institute from 1994-8 were evaluated for the feasibility of making a preoperative prediction of pulmonary complications. All had either a predicted postoperative forced vital capacity (FVC) of >800 ml/m(2) or forced expiratory volume in one second (FEV(1)) of >600 ml/m(2). RESULTS: Postoperative complications occurred in 37 patients (41.2%) but no patients died during the 30 day period after the operation. Pulmonary complications occurred in 20 patients (22.5%). Univariate analysis indicated that the factors significantly related to pulmonary complications were FVC <80%, serum lactate dehydrogenase (LDH) level > or =230 U/l, and arterial oxygen tension (PaO(2)) <10.6 kPa (80 mm Hg). In a multivariate analysis the three independent predictors of pulmonary complications were serum LDH > or =230 U/l (odds ratio (OR) 10.5, 95% CI 1.4 to 77.3), residual volume (RV)/total lung capacity (TLC) > or =30% (OR 6.0, 95% CI 1.1 to 33.7), and PaO(2) <10.6 kPa (OR 5.6, 95% CI 1.4 to 22.2). CONCLUSIONS: The above findings indicate that three factors (serum LDH levels of > or =230 U/l, RV/TLC > or =30%, and PaO(2) <10.6 kPa) may be associated with pulmonary complications in patients undergoing a lobectomy for NSCLC, even though the patient group was relatively small for statistical analysis of such a diverse subject as pulmonary complications. PMID- 11120907 TI - Directed neutrophil migration to IL-8 is increased in cystic fibrosis: a study of the effect of erythromycin. AB - BACKGROUND: The aim of this study was to compare neutrophil migration in cystic fibrosis (CF) and non-CF populations and to investigate the effect of erythromycin on directed migration of neutrophils (PMNs) in CF. METHODS: PMNs were isolated and their migratory capacity in response to interleukin-8 (IL-8) or f-Met-Leu-Phe (fMLP) in the presence or absence of erythromycin (1-100 microg/ml) was assessed. RESULTS: CF derived PMNs showed significantly increased migration to IL-8 but not to fMLP compared with non-CF PMNs. Erythromycin had no significant effect on migration responses to IL-8 and in vitro exposure of PMNs to erythromycin had no effect. CONCLUSIONS: CF derived PMNs show higher migratory responsiveness to IL-8 but not to fMLP, suggesting that CF PMNs may be "primed" to IL-8 which is significantly increased in CF serum compared with non-CF serum. Treatment with erythromycin had no significant effect on PMN migration in vitro. PMID- 11120908 TI - Adult outcome of congenital lower respiratory tract malformations. PMID- 11120910 TI - Ethical problems in respiratory care: the role of the law. PMID- 11120909 TI - Enhancing physical performance in chronic obstructive pulmonary disease. PMID- 11120911 TI - Myositis associated graft-versus-host-disease presenting as respiratory muscle weakness. AB - Myositis associated with graft-versus-host-disease (GVHD) typically presents with proximal muscle weakness, myalgias, and a raised creatinine phosphokinase (CPK) level. We report a case of a 51 year old man who developed respiratory muscle weakness five years after an allogeneic bone marrow transplant for multiple myeloma. His symptoms included tachypnoea, abdominal paradox, and orthopnoea. Pulmonary function tests revealed diminished vital capacity and maximal inspiratory and expiratory pressures. Serum CPK levels were raised and a peripheral muscle biopsy specimen was consistent with GVHD. He improved with immunosuppressive therapy. PMID- 11120912 TI - Handbook of sleep medicine PMID- 11120913 TI - Effects of removal of Na(+) and Cl(-) on spontaneous electrical activity, slow wave, in the circular muscle of the guinea-pig gastric antrum. AB - In the circular muscle of the guinea-pig gastric antrum, a decrease in the external Na(+) to less than 20 mM produced depolarization of the membrane with transient prolongation of the slow wave. This was followed by a high rhythmic activity. The activity was inhibited by reapplication of Na(+) before recovery. The depolarization in Na(+)-deficient solution was prevented and rhythmic activity continued at about 5/min for at least 6 min by simultaneous removal of K(+), Ca(2+), or Cl(-). After exposure to a Na(+)- and Cl(-)-deficient solution for a few minutes, reapplication of the Na(+) in Cl(-)-deficient solution inhibited generation of the slow wave until Cl(-) reapplication. Similar results were obtained when Na(+) and Cl(-) were reapplied in the absence of K(+) after exposure to a Na(+)-, K(+)-free, and Cl(-)-deficient solution, although the inhibition was weaker than Na(+) reapplication in a Cl(-)-deficient solution. In the presence of furosemide or bumetanide, a strong inhibition of activity was produced by the reapplication of Na(+) and Cl(-) after exposure to an Na(+)- and Cl(-)-deficient solution. A hypothesis is presented that intracellular Ca(2+) concentration ([Ca(2+)](i)) is the most important factor determining the generation and frequency of the slow wave and that [Ca(2+)](i) is regulated by the Na(+) concentration gradient across the plasma membrane. The recovery of the Na(+) concentration gradient by Na(+) reapplication after removal of Na(+) and Cl(-) is mainly controlled by a Na(+)-K(+)-Cl(-) co-transport. PMID- 11120914 TI - Logistic time constant of isometric relaxation force curve of ferret ventricular papillary muscle: reliable index of lusitropism. AB - We have found that a logistic function fits the left ventricular isovolumic relaxation pressure curve in the canine excised, cross-circulated heart more precisely than a monoexponential function. On this basis, we have proposed a logistic time constant (tau(L)) as a better index of ventricular isovolumic lusitropism than the conventional monoexponential time constant (tau(E)). We hypothesize in the present study that this tau(L) would also be a better index of myocardial isometric lusitropism than the conventional tau(E). We tested this hypothesis by analyzing the isometric relaxation force curve of 114 twitches of eight ferret isolated right ventricular papillary muscles. The muscle length was changed between 82 and 100% L(max) and extracellular Ca(2+) concentrations ([Ca(2+)](o)) between 0.2 and 8 mmol/l. We found that the logistic function always fitted the isometric relaxation force curve much more precisely than the monoexponential function at any muscle length and [Ca(2+)](o) level. We also found that tau(L) was independent of the choice of the end of isometric relaxation but tau(E) was considerably dependent on it as in ventricular relaxation. These results validated our present hypothesis. We conclude that tau(L) is a more reliable, though still empirical, index of lusitropism than conventional tau(E) in the myocardium as in the ventricle. PMID- 11120915 TI - The Effect of sex steroid hormones on substrate oxidation during prolonged submaximal exercise in women. AB - In animals, female sex steroid hormones (SS, estrogens-progesterone) influence the energy substrate that is metabolized. Human research on this issue is controversial. This study examined whether changes in circulating SS hormone levels affected the carbohydrate-lipid metabolism during submaximal prolonged (60 min) exercise. Young, physically active females were studied. Four were classified as anovulatory-oligomenorrheic and four were classified as ovulatory eumenorrheic. Subject responses were pooled to form one group (n = 8) and then their responses under low (L) and high (H) pharmaceutically manipulated SS hormone conditions were examined. During exercise, the mean oxygen consumption levels were 1.70 +/- 0.10/ x min(-1) for L-SS and 1.75 +/- 0.11/ x min(-1) for H SS (p = 0.07), respectively. The respiratory exchange ratio (RER) responses were significantly different during exercise between the conditions: 0.93 +/- 0.04 for L-SS and 0.90 +/- 0.04 for H-SS (p < 0.05), respectively. RER responses were utilized to calculate substrate oxidation. Significantly less carbohydrate oxidation was found in the H-SS condition as compared to the L-SS condition (p < 0.05). Lipid oxidation was also significantly different, but for this measure, the levels of oxidation were greater in the H-SS than in the L-SS condition (p < 0.05). Finally, total energy expenditure for the 60 min of exercise was not significantly different between the hormonal conditions. Results suggest that sex steroid hormones have an impact upon substrate oxidation in women during exercise. Specifically, high circulating concentrations of the SS hormones result in an enhanced reliance upon the oxidation of lipid as an energy substrate and consequently induce a reduction in carbohydrate oxidation. The mechanism inducing this "metabolism shift" appears due to sex steroid hormones directly and indirectly increasing lipid mobilization and lipolysis. PMID- 11120917 TI - Selective recording of electroneurograms from the sciatic nerve of a dog with multi-electrode spiral cuffs. AB - Electroneurograms (ENGs) from superficial regions of the sciatic nerve of a Beagle dog were recorded selectively with a chronically implanted 33-electrode spiral cuff (cuff). By delivering stimulating pulses to groups of three electrodes (GTEs) within the cuff we could define the relative positions of the particular superficial regions that selectively innervated the tibialis anterior (TA) and gastrocnemius muscles (GM). GTEs with and without contractions of the TA and GM muscles were selected and connected to a 4-channel ENG system designed to amplify ENGs by 100,000 times and to pass frequencies between 500 Hz and 10 kHz. In our study, 12 experiments were conducted on three Beagle dogs with a cuff implanted for up to 2 years. We present the results obtained in four experiments conducted on one animal. With the implanted leg mounted in a special electronic brace we applied extending forces to the ankle, rotating it by up to 37 degrees according to the neutral position, eliciting torque to stretch the TA muscle. Only the ENG from a GTE eliciting maximum contraction of the TA muscle showed activities corresponding to the trajectory of the mechanical load of the muscle. Next, we dissected the calcanean tendon (CT) of the implanted leg and applied repetitive pull forces to the CT. Only the ENG from the GTE eliciting maximum contraction of the GM muscle was activated in correspondence to the trajectory of the mechanical load applied on the CT. The results suggest that the cuff, implanted chronically on the sciatic nerve, is useful to record ENGs of the afferent fibers from TA and GM muscles selectively and that the technique could be extended for human use in the field of rehabilitation for paralysis. PMID- 11120916 TI - Effect of acupuncture-like stimulation on cortical cerebral blood flow in anesthetized rats. AB - The effect of acupuncture-like stimulation of various areas (cheek, forepaw, upper arm, chest, back, lower leg, hindpaw, perineum) on cortical cerebral blood flow (CBF) was examined in anesthetized rats. An acupuncture needle (diameter, 340 microm) was inserted into the skin and underlying muscles at a depth of about 5 mm and twisted to the right and left once a second for 1 min. CBF of the cortex was measured using a laser Doppler flowmeter. Stimulation of the cheek, forepaw, upper arm and hindpaw produced significant increases in CBF, but stimulation of the chest, back, lower leg and perineum did not produce significant responses. Stimulation of the cheek, forepaw, and hindpaw produced an increase in mean arterial pressure (MAP), while stimulation of the back produced a decrease in MAP. Stimulation of the upper arm, chest, lower leg and perineum did not produce a significant MAP response. After spinal transection at the 1st to 2nd thoracic level, the blood pressure response to stimulation of the cheek and forepaw was suppressed, whereas an increase in CBF still took place. The increase in CBF induced by forepaw stimulation was abolished by severance of the somatic nerves at the brachial plexus. Forepaw stimulation enhanced the activity of the radial, ulnar and median nerves. Furthermore, in the present study, passing of an electric current through acupuncture needles showed that excitation of group III (Adelta) and group IV (C) afferent fibers in the somatic nerve was capable of producing an increase in CBF, whereas excitation of group I (Aalpha) and group II (Abeta) fibers was ineffective. The increase in CBF induced by forepaw stimulation was almost abolished by intravenous administration of muscarinic and nicotinic cholinergic blocking agents (atropine 5 mg/kg and mecamylamine 20 mg/kg), and by bilateral lesions in the nucleus basalis of Meynert. Acupuncture like stimulation of a forepaw increased acetylcholine release in the cerebral cortex. We concluded that the increase in CBF, independent of systemic blood pressure, elicited by acupuncture stimulation is a reflex response in which the afferent nerve pathway is composed of somatic group III and IV afferent nerves, and efferent nerve pathway includes intrinsic cholinergic vasodilators originating in the nucleus basalis of Meynert. PMID- 11120918 TI - Effects of repetitive brief ischemia on contractile efficiency and oxygen cost of contractility in dog heart. AB - It is unclear whether preceding repetitive brief ischemia causes any improvement in the energy efficiency of intracellular calcium cycling or crossbridge cycling that may lead to cardioprotection after subsequent sustained ischemia/reperfusion, a phenomenon called ischemic preconditioning. To address this issue, left ventricular (LV) contractility (E(max)) and the relation between myocardial oxygen consumption (VO(2)) and pressure-volume area (PVA, a measure of LV total mechanical energy) were assessed before (Control) and 20 min (Rep-20) and 60 min (Rep-60) after repetitive brief ischemia in 11 isolated, blood perfused dog hearts. At Rep-20, E(max) and PVA-independent VO(2) (nonmechanical energy expenditure) decreased by 23.0 +/- 19.5 and 13.9 +/- 18.0%, respectively (both p < 0.05). However, at Rep-60, both E(max) and PVA-independent VO(2) recovered to their respective control levels. The oxygen cost of contractility (the slope of the PVA-independent VO(2)-E(max) relation during CaCl(2) loading) remained constant (Control 0.0019 +/- 0.0009 vs. Rep-60 0.0018 +/- 0.0013 ml O(2) x ml x mmHg(-1) x beat(-1) x 100 g(-2), ns), suggesting unchanged efficiency in Ca(2+) cycling. Also, the contractile efficiency (the reciprocal of the slope of the VO(2)-PVA relation, reflecting the efficiency of crossbridge cycling) was the same between the Control and Rep-60 (53.7 +/- 16.7 vs. 55.4 +/- 14.4%, ns). Basal metabolism VO(2) during KCl arrest was also similar to that in the normal heart. Nonmechanical energy expenditure was reduced in proportion to the decrease in LV contractility after repetitive brief ischemia, while both the contractile efficiency and oxygen cost of contractility remained constant. These results indicate that the heart, after repetitive brief ischemia but before sustained ischemia, has normal efficiencies of crossbridge cycling and Ca(2+) cycling despite the transiently reduced contractility. PMID- 11120919 TI - Involvement of cyclin-dependent kinase 5/p35(nck5a) in the synaptic reorganization of rat hippocampus during kindling progression. AB - To test the hypothesis that a complex of cyclin-dependent kinase 5 (Cdk5) and p35(nck5a) plays an important role in sprouting in the kindling rat hippocampus, we studied the changes in kinase activity, expression level and subcellular localization during kindling progression. The kinase activity in kindling rats was significantly higher than that in normal rats. The changes in kinase activity coincided with those of the p35(nck5a) expression in kindling rats. In contrast, the expression of Cdk5 was constant at all stages of kindling. Subcellular localization of Cdk5, however, changed markedly in the hippocampal neurons during kindling progression. Cdk5 translocated from axon to soma when the kinase activity was high. The phosphorylation level of tau protein was in good agreement with the Cdk5 kinase activity. In contrast, MAP kinase activity was not correlated with tau phosphorylation during kindling progression. These findings suggest that Cdk5/p35(nck5a) plays an important role in synaptic reorganization, and the translocation of Cdk5 to soma from axons is a crucial regulatory mechanism of kinase activity. PMID- 11120920 TI - Contractility of single myofibrils of rabbit skeletal muscle studied at various MgATP concentrations. AB - A novel experimental method was developed to study the contractility of single myofibrils of skeletal muscle. Single myofibrils (ca. 1 microm in diameter) prepared from glycerinated rabbit psoas muscle were suspended between rigid and flexible microneedles by the entwining method. The length changes of the preparations applied via the rigid microneedle by an actuator and the force produced were measured by photo-electrically detecting the nanometer deflections of the flexible microneedle. Single myofibril preparations maintained uniform sarcomere striations during contraction-relaxation cycles. The isometric force produced, the velocity of unloaded shortening, and the force-velocity relationship of single myofibrils were investigated at various MgATP concentrations. The contractility of single myofibrils thus obtained in the absence of ATP regenerative systems was essentially the same as that of skinned muscle fibers under comparable conditions in the presence of ATP regenerative systems. Thus, it was found that (1) the present experimental method is useful for studying the contractility of single myofibrils, and (2) in single myofibril preparations, the MgATP concentration at actomyosin sites is well equilibrated with that in bathing solutions. PMID- 11120921 TI - 2,3-Butanedione monoxime suppresses primarily total calcium handling in canine heart. AB - Whether 2,3-butanedione monoxime (BDM, < or = 5mmol/l) suppresses primarily crossbridge cycling or total Ca(2+) handling in the blood-perfused whole heart remains controversial. Although BDM seems to suppress primarily total Ca(2+) handling in canine hearts, more evidence is lacking. We therefore analyzed the cardiac mechanoenergetics, namely, E(max) (contractility), PVA (total mechanical energy), and O(2) consumption of canine BDM-treated hearts by our recently developed integrative method to assess myocardial total Ca(2+) handling. This method additionally required the internal Ca(2+) recirculation fraction. We obtained this from the beat constant of the exponential decay component of the postextrasystolic potentiation. Our analysis indicated significant decreases in both internal Ca(2+) recirculation fraction and total Ca(2+) handling in the BDM treated heart, but virtually no change in the reactivity of E(max) to total Ca(2+) handling. This result corroborates the view that BDM suppresses primarily total Ca(2+) handling rather than crossbridge cycling in the canine blood perfused heart. PMID- 11120922 TI - Flash-related synchronization and desynchronization revealed by a multiple band frequency analysis. AB - The fast Fourier transform (FFT) is a good method to estimate power spectral density (PSD), but the frequency resolution is limited to the sampling window, and thus the precise characteristics of PSD for short signals are not clear. To relax the limitation, a multiple band-pass filter was introduced to estimate the precise course of PSDs for flash visual evoked potentials (VEPs). Signals were recorded during -200 and 600 ms using balanced noncephalic electrodes, and sampled at 1,000 Hz in 12 bits. With 1 Hz and 10 ms resolutions, PSDs were estimated between 10 and 100 Hz. Background powers at the alpha- and beta-bands were high over the posterior scalp, and powers around 200 ms were evoked at the same bands over the same region, corresponding to P110 and N165 of VEPs. Normalized PSDs showed evoked powers around 200 ms and suppressed powers following the evoked powers over the posterior scalp. The evoked powers above the 20 Hz band were not statistically significant, however, the gamma band was significantly evoked intra-individually; details in the gamma bands were varied among the subjects. Details of PSDs were complicated even for a simple task such as watching flashes; both synchronization and desynchronization occurred with different distributions and different time courses. PMID- 11120923 TI - Human beta-herpesvirus interactions in solid organ transplant recipients. AB - The replication of beta-herpesviruses-cytomegalovirus (CMV), human herpesvirus (HHV)-6, and HHV-7-and their association with CMV disease and response to antiviral therapy were prospectively investigated in 33 liver transplant recipients not given antiviral prophylaxis. CMV, HHV-6, and HHV-7 DNA were detected within 8 weeks after transplantation in 70%, 33%, and 42% of the patients, respectively. The univariate association between CMV disease and the 3 beta-herpesviruses was more significant by virus load quantitation than by qualitative detection of DNA. This association with high levels of CMV, HHV-6, and HHV-7 (P<.001,.022, and.001, respectively) occurred mainly in CMV seronegative recipients of transplants from CMV-seropositive donors. Antiviral therapy with ganciclovir (Gcv) reduced the load of CMV and HHV-6 and HHV-7. These results suggest that CMV disease in transplant recipients is related to the unique interaction of the 3 beta-herpesviruses and is ultimately reduced after intravenous Gcv treatment. PMID- 11120924 TI - Serological evidence for an inflammatory response in murine scrapie. AB - Transmissible spongiform encephalopathies (TSEs) are initiated by a novel kind of agent that produces characteristic degenerative changes in the brain without a detectable systemic inflammatory response or serological changes. A murine scrapie model was evaluated for changes in plasma concentration of serum amyloid P component (SAP), a protein that is up-regulated in infected and/or injured mice during the acute phase response (APR). C57BL10 and IRW mice inoculated with scrapie brain developed clinical scrapie 125-150 days later. At this time, concentration of plasma SAP increased in most of them. The SAP level increased > or =3-fold in >80% of the scrapie-affected C57BL10 mice and IRW male mice. A similar increase was found in <3% of respective nonscrapie control mice. The up regulation of mouse SAP during clinical scrapie provides evidence for the activation of a systemic APR in TSE, a serological change that may be clinically useful. PMID- 11120925 TI - Increased susceptibility to Kuru of carriers of the PRNP 129 methionine/methionine genotype. AB - Kuru reached epidemic proportions by the mid-twentieth century among the Fore people of New Guinea and disappeared after the abolition of cannibalistic rituals. To determine susceptibility to kuru and its role in the spread and elimination of the epidemic, we analyzed the PRNP gene coding sequences in 5 kuru patients; no germline mutations were found. Analysis of the PRNP 129 methionine (M)/valine (V) polymorphism in 80 patients and 95 unaffected controls demonstrated that the kuru epidemic preferentially affected individuals with the M/M genotype. A higher representation of M/M carriers was observed among the affected young Fore males entering the age of risk, whereas a lower frequency of M/M homozygotes was found among the survivors. M/V and V/V genotypes predisposed to a lower risk of disease development and longer incubation times. These findings are relevant to the current outbreak of variant Creutzfeldt-Jakob disease (vCJD) in the United Kingdom, because all vCJD patients tested thus far have been M/M carriers. PMID- 11120926 TI - Increased activated human T cell lymphotropic virus type I (HTLV-I) Tax11-19 specific memory and effector CD8+ cells in patients with HTLV-I-associated myelopathy/tropical spastic paraparesis: correlation with HTLV-I provirus load. AB - To discern the T cell subtype associated with T cell differentiation, the expression of CD45RA and CD27 was measured from total CD8(high) cells and from human T cell lymphotropic virus type I (HTLV-I) Tax11-19 peptide-specific CD8(+) cells in peripheral blood lymphocytes of patients with HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Phenotypically defined memory and/or effector cells (CD45RA(-)CD27(+), CD45RA(+)CD27(-), and CD45RA(-)CD27(-)) were increased in HAM/TSP CD8(+) cells, compared with those of HTLV-I seronegative healthy control subjects. The percentage of human leukocyte antigen (HLA)-DR-positive cells was also increased in CD8(+) cells of HAM/TSP, compared with those in HLA-DR(+)CD8(+) cells of healthy control subjects. HTLV-I provirus load correlated with the frequency of Tax11-19-specific CD8(+) cells. The high frequency of memory and/or effector type HTLV-I Tax11-19-specific CD8(+) cells suggests that continuous restimulation driven by HTLV-I antigens in vivo may be associated with the pathogenesis of HAM/TSP. PMID- 11120927 TI - Correlates of mother-to-child human immunodeficiency virus type 1 (HIV-1) transmission: association with maternal plasma HIV-1 RNA load, genital HIV-1 DNA shedding, and breast infections. AB - To determine the effects of plasma, genital, and breast milk human immunodeficiency virus type 1 (HIV-1) and breast infections on perinatal HIV-1 transmission, a nested case-control study was conducted within a randomized clinical trial of breast-feeding and formula feeding among HIV-1-seropositive mothers in Nairobi, Kenya. In analyses comparing 92 infected infants with 187 infants who were uninfected at 2 years, maternal viral RNA levels >43,000 copies/mL (cohort median) were associated with a 4-fold increase in risk of transmission (95% confidence interval [CI], 2.2-7.2). Maternal cervical HIV-1 DNA (odds ratio [OR], 2.4; 95% CI, 1.3-4.4), vaginal HIV-1 DNA (OR, 2.3; 95% CI, 1.1 4.7), and cervical or vaginal ulcers (OR, 2.7; 95% CI, 1.2-5.8) were significantly associated with infant infection, independent of plasma virus load. Breast-feeding (OR, 1.7; 95% CI, 1.0-2.9) and mastitis (relative risk [RR], 3.9; 95% CI, 1.2-12.7) were associated with increased transmission overall, and mastitis (RR, 21.8; 95% CI, 2.3-211.0) and breast abscess (RR, 51.6; 95% CI, 4.7 571.0) were associated with late transmission (occurring >2 months postpartum). Use of methods that decrease infant exposure to HIV-1 in maternal genital secretions or breast milk may enhance currently recommended perinatal HIV-1 interventions. PMID- 11120928 TI - Atherosclerosis in apoE knockout mice infected with multiple pathogens. AB - Cytomegalovirus (CMV) and Chlamydia pneumoniae (CP) possibly contribute to atherosclerosis. Murine CMV (MCMV) and CP increase lesion size in apoE knockout mice. In this study, apoE knockout mice were infected with MCMV and CP to determine whether infection with multiple pathogens increases lesion size to a greater extent than either pathogen alone and whether infection with MCMV changes serum cytokine levels in a manner that could increase lesion development. One group of mice received MCMV at 2 weeks of age, followed by 2 doses of CP at 6 and 8 weeks of age. Additional groups received only MCMV or CP. Animals were killed at 16 weeks of age to determine lesion area. Infection with MCMV alone, CP alone, and both MCMV and CP increased lesion size 84% (P<.001), 70% (P<.0001), and 45% (P<.01), respectively. The MCMV-induced increase in circulating levels of interferon-gamma may have contributed to this increase. PMID- 11120929 TI - Effect of azithromycin on murine arteriosclerosis exacerbated by Chlamydia pneumoniae. AB - Chlamydia pneumoniae infection can exacerbate atherosclerosis in animals. To test the hypothesis that antibiotic therapy inhibits the atherogenic effects of C. pneumoniae infection, 10-week-old apolipoprotein E (ApoE) null mice were infected with C. pneumoniae or placebo, were treated for 2 weeks after infection with azithromycin or placebo, and were killed at 20 weeks of age. Infection did not affect the size of the aortic lesion, and antibiotic treatment had no effect. Another group of mice, 12-week-old ApoE mice, were infected with C. pneumoniae or placebo, were treated for 2 weeks after infection with azithromycin or placebo, and were killed at 26 weeks of age. C. pneumoniae infection increased the size of the lesion in infected mice, but azithromycin did not reduce the size of the aortic lesion in infected mice. Therefore, immediate therapy of acute infection may be necessary to prevent the proatherogenic effects of C. pneumoniae infection. PMID- 11120930 TI - Invasive pneumococcal disease in England and Wales: vaccination implications. AB - Knowledge of the epidemiology of invasive pneumococcal disease (IPD) will aid in planning the use of pneumococcal vaccines. A United Kingdom (UK)-based surveillance in England and Wales (1995-1997) of 11,528 individuals with IPD and a local enhanced surveillance in the Oxford (UK) area (1995-1999) have been analyzed. IPD has a high attack rate in children, with 37.1-48.1 cases per 100,000 infants <1 year old per year, and in older persons, with 21.2-36.2 cases per 100,000 persons >65 years old per year, for England, Wales, and Oxford. The 7 valent conjugate vaccine includes serotypes causing < or =79% of IPD in children <5 years old, but only 66% in adults >65 years old. The data also indicate that IPD varies by serotype, age, and country, emphasizing that the epidemiology of IPD is heterogeneous and requires continued surveillance. PMID- 11120931 TI - Genes of the class II and class III major histocompatibility complex are associated with typhoid fever in Vietnam. AB - The influence of genes of the major histocompatibility complex (MHC) class II and class III loci on typhoid fever susceptibility was investigated. Individuals with blood culture-confirmed typhoid fever and control subjects from 2 distinct geographic locations in southern Vietnam were genotyped for HLA-DRB1 and HLA-DQB1 alleles, the gene that encodes tumor necrosis factor (TNF)-alpha (TNFA [-238] and TNFA [-308]), the gene that encodes lymphotoxin-alpha, and alleles of the TNF alpha microsatellite. HLA-DRB1*0301/6/8, HLA-DQB1*0201-3, and TNFA*2 (-308) were associated with susceptibility to typhoid fever, whereas HLA-DRB1*04, HLA DQB1*0401/2, and TNFA*1 (-308) were associated with disease resistance. The frequency of all possible haplotypes of the 3 individually associated loci were estimated and were found to be significantly different in typhoid case patients and control subjects (chi2=55.56, 32 df; P=.006). Haplotypes that were either protective (TNFA*1 [-308].DRB1*04) or predisposed individuals to typhoid fever (TNFA*2 [-308].DRB1*0301) were determined. This report identifies a genetic association in humans between typhoid fever and MHC class II and III genes. PMID- 11120932 TI - Association of Mycoplasma genitalium with nongonococcal urethritis in heterosexual men. AB - Chlamydia trachomatis and Neisseria gonorrhoeae are universally acknowledged as urethral pathogens, yet the etiology in the majority of cases of urethritis is unclear. Our case-control study assessed the association of Mycoplasma genitalium, Ureaplasma urealyticum, and other potential pathogens with acute nongonococcal urethritis (NGU) in heterosexual men presenting to an urban sexually transmitted diseases clinic. M. genitalium was detected in 27 (22%) of 121 NGU case patients and in 5 (4%) of 117 control subjects (P<.01). Although C. trachomatis was detected in 36 (30%) of 121 NGU case patients and in 4 (3%) of 117 control subjects (P<.01), only 3 men with NGU were infected with both C. trachomatis and M. genitalium. U. urealyticum was not associated with NGU. By multivariate analyses, controlling for age, race, history of prior urethritis, and chlamydial infection, M. genitalium was associated with a 6.5-fold increased risk of urethritis (95% confidence interval, 2.1-19.5), which supports a role of this organism in the etiology of NGU. PMID- 11120933 TI - Candida-specific systemic cell-mediated immune reactivities in human immunodeficiency virus-positive persons with mucosal candidiasis. AB - Oropharyngeal candidiasis (OPC), as opposed to vulvovaginal candidiasis (VVC), is a common opportunistic infection in human immunodeficiency virus (HIV)-positive persons that correlates with reduced CD4 T cell counts. Although cell-mediated immunity (CMI) by CD4 Th1-type cells is considered to be the predominant host defense against mucosal candidiasis, the immune factors associated with susceptibility to OPC in HIV-positive persons are not well understood. This study investigated Candida-specific systemic CMI in HIV-positive persons with OPC and/or VVC. Reductions in delayed skin test reactivity to Candida antigen were observed in HIV-positive persons with CD4 cell counts <200 cells/microL, irrespective of the presence of mucosal infection. Likewise, despite the correlate of OPC with reduced CD4 cell counts in HIV-positive persons, differences in Candida-specific peripheral blood mononuclear cell proliferation and Th1/Th2 cytokine production between HIV-positive and HIV-negative persons were not consistent in a manner to suggest that deficiencies in Candida-specific systemic CMI account solely for the susceptibility to OPC. PMID- 11120934 TI - Mutations conferring ganciclovir resistance in a cohort of patients with acquired immunodeficiency syndrome and cytomegalovirus retinitis. AB - Cytomegalovirus (CMV) retinitis is among the most common opportunistic infections in patients with acquired immunodeficiency syndrome. In a prospective study of 210 patients with CMV retinitis, 26 were identified as having either a phenotypic or a genotypic ganciclovir-resistant isolate from either blood or urine cultures. For blood culture isolates with an IC(50) >6.0 microm for ganciclovir, the sensitivity and specificity for detecting a UL97 mutation were 95% and 98%, respectively, whereas for an IC(50) >8.0 microM they were 79% and 99%, respectively. Although there were trade-offs between the 2 thresholds for blood culture isolates, for urine culture isolates an IC(50) >8.0 microM appeared to be better at identifying genotypic resistance. UL97 mutations identified in both the blood and urine cultures of individual patients were identical in 87.5% of cases. High-level ganciclovir resistance (IC(50), >30 microM) typically, but not invariably, was associated with a mutation in both the UL97 and UL54 genes. PMID- 11120935 TI - Thalidomide in low intermittent doses does not prevent recurrence of human immunodeficiency virus-associated aphthous ulcers. AB - A multicenter, double-blind, randomized, placebo-controlled study was conducted to determine the safety and efficacy of thalidomide in reduced, intermittent doses for preventing recurrences of oral and esophageal aphthous ulcers in patients with human immunodeficiency virus (HIV) infection. Forty-nine HIV infected patients whose ulcers previously had healed as a result of thalidomide therapy were randomly assigned to receive either 100 mg of oral thalidomide or placebo 3 times per week for 6 months. Ulcers recurred in 14 (61%) of 23 thalidomide-randomized patients, compared with 11 (42%) of 26 placebo-randomized patients, with no significant difference in the median time to recurrence of ulcers (P=.221). There were no changes in plasma levels of HIV RNA, tumor necrosis factor (TNF)-alpha, and soluble TNF receptor II at the time of ulcer recurrence. Adverse events among patients treated with thalidomide included neutropenia (5 patients), rash (5 patients), and peripheral sensory neuropathy (3 patients). Thalidomide in lower intermittent doses is ineffective at preventing recurrence of aphthous ulcers in HIV-infected persons. PMID- 11120936 TI - Changes in oropharyngeal colonization and infection by Candida albicans in human immunodeficiency virus-infected patients. PMID- 11120938 TI - Use of preclinical data for selection of a phase II/III dose for evernimicin and identification of a preclinical MIC breakpoint. AB - One of the most challenging issues in the design of phase II/III clinical trials of antimicrobial agents is dose selection. The choice is often based on preclinical data from pharmacokinetic (PK) studies with animals and healthy volunteers but is rarely linked directly to the target organisms except by the MIC, an in vitro measure of antimicrobial activity with many limitations. It is the thesis of this paper that rational dose-selection decisions can be made on the basis of the pharmacodynamics (PDs) of the test agent predicted by a mathematical model which uses four data sets: (i) the distribution of MICs for clinical isolates, (ii) the distribution of the values of the PK parameters for the test drug in the population, (iii) the PD target(s) developed from animal models of infection, and (iv) the protein binding characteristics of the test drug. In performing this study with the new anti-infective agent evernimicin, we collected a large number (n = 4,543) of recent clinical isolates of gram-positive pathogens (Streptococcus pneumoniae, Enterococcus faecalis and Enterococcus faecium, and Staphylococcus aureus) and determined the MICs using E-test methods (AB Biodisk, Stockholm, Sweden) for susceptibility to evernimicin. Population PK data were collected from healthy volunteers (n = 40) and patients with hypoalbuminemia (n = 12), and the data were analyzed by using NPEM III. PD targets were developed with a neutropenic murine thigh infection model with three target pathogens: S. pneumoniae (n = 5), E. faecalis (n = 2), and S. aureus (n = 4). Drug exposure or the ratio of the area under the concentration-time curve/MIC (AUC/MIC) was found to be the best predictor of microbiological efficacy. There were three possible microbiological results: stasis of the initial inoculum at 24 h (10(7) CFU), log killing (pathogen dependent, ranging from 1 to 3 log(10)), or 90% maximal killing effect (90% E(max)). The levels of protein binding in humans and mice were similar. The PK and PD of 6 and 9 mg of evernimicin per kg of body weight were compared; the population values for the model parameters and population covariance matrix were used to generate five Monte Carlo simulations with 200 subjects each. The fractional probability of attaining the three PD targets was calculated for each dose and for each of the three pathogens. All differences in the fractional probability of attaining the target AUC/MIC in this PD model were significant. For S. pneumoniae, the probability of attaining all three PD targets was high for both doses. For S. aureus and enterococci, there were increasing differences between the 6- and 9-mg/kg evernimicin doses for reaching the 2 log killing (S. aureus), 1 log killing (enterococci), or 90% E(max) AUC/MIC targets. This same approach may also be used to set preliminary in vitro MIC breakpoints. PMID- 11120939 TI - Pharmacodynamics of telithromycin In vitro against respiratory tract pathogens. AB - Telithromycin (HMR 3647) is a new ketolide that belongs to a new class of semisynthetic 14-membered-ring macrolides which have expanded activity against multidrug-resistant gram-positive bacteria. The aim of the present study was to investigate different basic pharmacodynamic properties of this new compound. The following studies of telithromycin were performed: (i) studies of the rate and extent of killing of respiratory tract pathogens with different susceptibilities to erythromycin and penicillin exposed to a fixed concentration that corresponds to a dose of 800 mg in humans, (ii) studies of the rate and extent of killing of telithromycin at five different concentrations, (iii) studies of the rate and extent of killing of the same pathogens at three different inocula, (iv) studies of the postantibiotic effect and the postantibiotic sub-MIC effect of telithromycin, and (v) determination of the rate and extent of killing of telithromycin in an in vitro kinetic model. In conclusion, telithromycin exerted an extremely fast killing of all strains of Streptococcus pneumoniae both with static concentrations and in the in vitro kinetic model. A slower killing of the strains of Streptococcus pyogenes was noted, with regrowth in the kinetic model of a macrolide-lincosamide-streptogramin B-inducible strain. The strains of Haemophilus influenzae were not killed at all at a concentration of 0.6 mg/liter due to high MICs. A time-dependent killing was seen for all strains. No inoculum effect was seen for the strains of S. pneumoniae, with a 99.9% reduction in the numbers of CFU for all inocula at both 8 h and 24 h. The killing of the strains of S. pyogenes was reduced by 1 log(10) CFU at 8 h and 2 to 3 log(10) CFU at 24 h when the two lower inocula were used but not at all at 8 and 24 h when the highest inoculum was used. For both of the H. influenzae strains there was an inoculum effect, with 1 to 2 log(10) CFU less killing for the inoculum of 10(8) CFU/ml in comparison to that for the inoculum of 10(6) CFU/ml. Overall, telithromycin exhibited long postantibiotic effects and postantibiotic sub-MIC effects for all strains investigated. PMID- 11120937 TI - Macrolide resistance conferred by base substitutions in 23S rRNA. PMID- 11120940 TI - Pharmacokinetic and pharmacodynamic study of the human immunodeficiency virus protease inhibitor amprenavir after multiple oral dosing. AB - In a dose-ranging study of amprenavir (formerly 141W94), an inhibitor of the protease enzyme of human immunodeficiency virus (HIV) type 1, single-dose and steady-state pharmacokinetic parameters were estimated from plasma samples collected on day 1 and during week 3, respectively. Amprenavir was administered on either a twice-daily (b.i.d.) or three-times-daily dosage schedule to 62 HIV infected adults, 59 of whom had pharmacokinetic data. Log-log regression analysis (the power model) revealed that the steady-state area under the curve (AUC(ss)) and the maximum, minimum, and average concentrations at steady state (C(max,ss), C(min,ss), and C(avg,ss), respectively) increased in a dose-proportional manner over the 300- to 1,200-mg dose range. Steady-state clearance was dose independent. AUC(ss)/AUC(0-->infinity) decreased linearly with dose and correlated significantly with treatment-associated decreases in alpha(1)-acid glycoprotein. After 3 weeks, the dose of 1,200 mg b.i. d. provided a median amprenavir C(min,ss) (0.280 microg/ml) that was higher than the median in vitro 50% inhibitory concentration for clinical HIV isolates (0.023 microg/ml), even after adjustment for protein binding. The median amprenavir C(min,ss) was also greater than the estimated in vivo trough concentration calculated to yield 90% of the maximum antiviral effect (0.228 microg/ml) over 4 weeks. A pharmacodynamic analysis of the relationship between steady-state pharmacokinetic parameters and safety revealed headache and oral numbness to be the only side effects significantly associated with C(max). The pharmacodynamic relationship defined in this study supports the use of 1,200 mg b.i.d. as the approved dose of amprenavir. PMID- 11120941 TI - A soxRS-constitutive mutation contributing to antibiotic resistance in a clinical isolate of Salmonella enterica (Serovar typhimurium). AB - The soxRS regulon is activated by redox-cycling drugs such as paraquat and by nitric oxide. The >15 genes of this system provide resistance to both oxidants and multiple antibiotics. An association between clinical quinolone resistance and elevated expression of the soxRS regulon has been observed in Escherichia coli, but this association has not been explored for other enteropathogenic bacteria. Here we describe a soxRS-constitutive mutation in a clinical strain of Salmonella enterica (serovar Typhimurium) that arose with the development of resistance to quinolones during treatment. The elevated quinolone resistance in this strain derived from a point mutation in the soxR gene and could be suppressed in trans by multicopy wild-type soxRS. Multiple-antibiotic resistance was also transferred to a laboratory strain of S. enterica by introducing the cloned mutant soxR gene from the clinical strain. The results show that constitutive expression of soxRS can contribute to antibiotic resistance in clinically relevant S. enterica. PMID- 11120942 TI - Clarithromycin inhibits NF-kappaB activation in human peripheral blood mononuclear cells and pulmonary epithelial cells. AB - Macrolide antibiotics modulate the production of proinflammatory cytokines in vivo and in vitro. Transcription of the genes for these proinflammatory cytokines is regulated by nuclear factor kappaB (NF-kappaB). We examined whether or not clarithromycin inhibits the activation of NF-kappaB induced by tumor necrosis factor alpha (TNF-alpha) or staphylococcal enterotoxin A (SEA) in human monocytic U-937 cells, a T-cell line (Jurkat), a pulmonary epithelial cell line (A549), and peripheral blood mononuclear cells (PBMC). Flow cytometry revealed that clarithromycin suppresses NF-kappaB activation induced by TNF-alpha in U-937 and Jurkat cells in a concentration-related manner. Western blot analysis also demonstrated that clarithromycin inhibits NF-kappaB activation induced by TNF alpha in U-937, Jurkat, and A549 cells and PBMC and by SEA in PBMC. Western blot analysis of cytoplasmic extracts of A549 cells revealed that this inhibition is not linked to preservation of expression of the IkappaBalpha protein. The chloramphenicol acetyltransferase assay indicated that NF-kappaB-dependent reporter gene expression is suppressed in U-937 cells pretreated with clarithromycin. These findings are consistent with the idea that clarithromycin suppresses the production of proinflammatory cytokines via inhibition of NF kappaB activation. PMID- 11120943 TI - Activity of gatifloxacin alone or in combination with pyrimethamine or gamma interferon against Toxoplasma gondii. AB - The activity of gatifloxacin against Toxoplasma gondii, either alone or in combination with pyrimethamine or gamma interferon (IFN-gamma), was examined in vitro and in vivo. In vitro, gatifloxacin significantly inhibited intracellular replication of tachyzoites of the RH strain with a 50% inhibitory concentration of 0.21 microg/ml at 48 h after addition of the drug to the cultures. Toxicity for host cells was not observed at this concentration. A synergistic effect (combination indices < 0.5) was demonstrated in vitro following 48 h of treatment with the combination of gatifloxacin and pyrimethamine (1:1 ratio). Doses of gatifloxacin of 100 and 200 mg/kg of body weight/day administered orally to mice for 10 days resulted in significant (P values of 0.056 and <0.0001, respectively) prolongation in time to death following infection with a lethal inoculum of tachyzoites. A dose of 400 mg/kg resulted in 20% survival (P = 0.0001). Mortality was 100% in untreated control mice and in mice treated with 25 or 50 mg/kg/day. Treatment of infected mice with a combination of gatifloxacin at 200 mg/kg/day and pyrimethamine at 12.5 mg/kg/day resulted in 85% survival, whereas 100 and 80% of mice treated with gatifloxacin alone or pyrimethamine alone, respectively, died (P < 0.0001). Moreover, a gatifloxacin dose of 200 mg/kg/day administered orally for 10 days plus 2 microg of recombinant murine IFN-gamma/day administered intraperitoneally for 10 days resulted in significant survival compared with IFN gamma alone (P < 0.0001) or gatifloxacin alone (P < 0.007). PMID- 11120944 TI - Inducible azole resistance associated with a heterogeneous phenotype in Candida albicans. AB - The development of azole resistance in Candida albicans is most problematic in patients with AIDS who receive long courses of drug for therapy or prevention of oral candidiasis. Recently, the rapid development of resistance was noted in other immunosuppressed patients who developed disseminated candidiasis despite fluconazole prophylaxis. One of these series of C. albicans isolates became resistant, with an associated increase in mRNA specific for a CDR ATP-binding cassette transporter efflux pump (K. A. Marr, C. N. Lyons, T. R. Rustad, R. A. Bowden, and T. C. White, Antimicrob. Agents Chemother. 42:2584-2589, 1998). Here we study this series of C. albicans isolates further and examine the mechanism of azole resistance in a second series of C. albicans isolates that caused disseminated infection in a recipient of bone marrow transplantation. The susceptible isolates in both series become resistant to fluconazole after serial growth in the presence of drug, while the resistant isolates in both series become susceptible after serial transfer in the absence of drug. Population analysis of the inducible, transiently resistant isolates reveals a heterogeneous population of fluconazole-susceptible and -resistant cells. We conclude that the rapid development of azole resistance occurs by a mechanism that involves selection of a resistant clone from a heterogeneous population of cells. PMID- 11120945 TI - Role of human immunodeficiency virus (HIV) type 1 envelope in the anti-HIV activity of the betulinic acid derivative IC9564. AB - The betulinic acid derivative IC9564 is a potent anti-human immunodeficiency virus (anti-HIV) compound that can inhibit both HIV primary isolates and laboratory-adapted strains. However, this compound did not affect the replication of simian immunodeficiency virus and respiratory syncytial virus. Results from a syncytium formation assay indicated that IC9564 blocked HIV type 1 (HIV-1) envelope-mediated membrane fusion. Analysis of a chimeric virus derived from exchanging envelope regions between IC9564-sensitive and IC9564-resistant viruses indicated that regions within gp120 and the N-terminal 25 amino acids (fusion domain) of gp41 are key determinants for the drug sensitivity. By developing a drug-resistant mutant from the NL4-3 virus, two mutations were found within the gp120 region and one was found within the gp41 region. The mutations are G237R and R252K in gp120 and R533A in the fusion domain of gp41. The mutations were reintroduced into the NL4-3 envelope and analyzed for their role in IC9564 resistance. Both of the gp120 mutations contributed to the drug sensitivity. On the contrary, the gp41 mutation (R533A) did not appear to affect the IC9564 sensitivity. These results suggest that HIV-1 gp120 plays a key role in the anti HIV-1 activity of IC9564. PMID- 11120946 TI - Susceptibilities of Haemophilus influenzae and Moraxella catarrhalis to ABT-773 compared to their susceptibilities to 11 other agents. AB - The activity of the ketolide ABT-773 against Haemophilus and Moraxella was compared to those of 11 other agents. Against 210 Haemophilus influenzae strains (39.0% beta-lactamase positive), microbroth dilution tests showed that azithromycin and ABT-773 had the lowest MICs (0.5 to 4.0 and 1.0 to 8.0 microg/ml, respectively), followed by clarithromycin and roxithromycin (4.0 to >32.0 microg/ml). Of the beta-lactams, ceftriaxone had the lowest MICs (32.0 microg/ml). Against 50 Moraxella catarrhalis strains, all of the compounds except amoxicillin and cefprozil were active. Time-kill studies against 10 H. influenzae strains showed that ABT-773, at two times the MIC, was bactericidal against 9 of 10 strains, with 99% killing of all strains at the MIC after 24 h; at 12 h, ABT-773 gave 90% killing of all strains at two times the MIC. At 3 and 6 h, killing by ABT-773 was slower, with 99.9% killing of four strains at two times the MIC after 6 h. Similar results were found for azithromycin, with slightly slower killing by erythromycin, clarithromycin, and roxithromycin, especially at earlier times. beta-Lactams were bactericidal against 8 to 10 strains at two times the MIC after 24 h, with slower killing at earlier time periods. Most compounds gave good killing of five M. catarrhalis strains, with beta-lactams killing more rapidly than other drugs. ABT 773 and azithromycin gave the longest postantibiotic effects (PAEs) of the ketolide-macrolide-azalide group tested (4.4 to >8.0 h), followed by clarithromycin, erythromycin, and roxithromycin. beta-Lactam PAEs were similar and shorter than those of the ketolide-macrolide-azalide group for all strains tested. PMID- 11120947 TI - Role of P glycoprotein in the course and treatment of Encephalitozoon microsporidiosis. AB - Encephalitozoon microsporidia are obligate intracellular protozoan parasites that proliferate and differentiate within a parasitophorous vacuole inside host cells that are usually epithelial in nature. Isolates of the three species of the Encephalitozoon microsporidia, E. cuniculi, E. hellem, and E. intestinalis, were obtained from AIDS patients and cultured in green monkey (E6) kidney cells. Anti P-glycoprotein (anti-Pgp) and anti-multidrug resistance-associated protein (anti MRP) monoclonal antibodies were used to probe for multidrug resistance (MDR) pump epitopes and verapamil- or cyclosporin A- and probenecid-modulated intracellular calcein fluorescence were used to assess the expression of Pgp and MRP respectively in uninfected and infected cells. Pgp, but not MRP, was detected immunocytochemically and by verapamil- and cyclosporin A-potentiated intracellular fluorescence in both host cells and parasite developing stages. When an in vitro infection assay was employed, verapamil and cyclosporin A acted as chemosensitizing agents for the antiparasitic drug albendazole. These observations suggest that inhibiting host cell and perhaps parasite MDR pumps may increase the efficacy of antiparasitic agents in these and other microsporidia species. PMID- 11120948 TI - Effects of mutations in ribosomal protein L16 on susceptibility and accumulation of evernimicin. AB - Chemical mutagenesis of Staphylococcus aureus RN450 generated two strains that displayed a stable reduction (30- to 60-fold) in susceptibility to evernimicin. Cell-free translation reactions demonstrated that the resistance determinant was located in the ribosomal fraction. Compared to ribosomes isolated from a wild type strain, ribosomes from the mutant strains displayed an 8- to 10-fold reduction in affinity for [(14)C]evernimicin. In contrast, the mutants displayed no alteration in either binding affinity or in vitro susceptibility to erythromycin. Exponential cultures of the mutant strains accumulated significantly less [(14)C]evernimicin than the wild-type strain, suggesting that accumulation is dependent on the high affinity that evernimicin displays for its binding site. Sequencing rplP (encodes ribosomal protein L16) in the mutant strains revealed a single base change in each strain, which resulted in a substitution of either cysteine or histidine for arginine at residue 51. Introduction of a multicopy plasmid carrying wild-type rplP into the mutant strains restored sensitivity to evernimicin, confirming that the alterations in rplP were responsible for the change in susceptibility. Overexpression of the mutant alleles in S. aureus RN450 had no effect on susceptibility to evernimicin, demonstrating that susceptibility is dominant over resistance. PMID- 11120949 TI - Efficacy of 2-amino-7-(1,3-dihydroxy-2-propoxymethyl)purine for treatment of vaccinia virus (orthopoxvirus) infections in mice. AB - We have previously shown that the N-7 substituted acyclic nucleoside analog 2 amino-7-[1,3-dihydroxy-2-propoxy)methyl]purine (compound S2242) is, both in vitro and in animal models, a potent inhibitor of the replication of several herpesviruses (Neyts et al., Antimicrob. Agents Chemother. 39:56-60, 1995). Here we report on the potent and selective antiviral activity of S2242 against vaccinia virus (VV), an orthopoxvirus. The 50% effective concentrations for inhibition of VV-induced cytopathic effect and viral DNA synthesis in cell culture were 2.4 and 0.2 microg/ml, respectively. We next studied the efficacy of S2242 in VV-infected mice. Immunocompetent NMRI mice that had been inoculated intravenously with VV developed tail lesions. Mice that had been treated for 5 consecutive days via the subcutaneous (s.c.) route with 100 mg of the diacetate ester prodrug of S2242 (compound H961) per kg of body weight did not develop any lesions and demonstrated no adverse effects. Severe combined immunodeficient (SCID) mice that had been inoculated intraperitoneally with VV became sick and died within 1 month after infection. Following treatment with H961 at 100 mg/kg for 10 consecutive days (either via oral gavage or s.c. injection) VV-inoculated SCID mice were completely protected, for at least 3 months, against virus-induced morbidity and mortality. At that time, no virus could be recovered from the organs of these mice (as assessed by titration for infectious virus, a DNA hybridization assay, and a PCR for VV-specific sequences). Compound S2242 and its oral prodrug H961 could be useful in treatment of orthopoxvirus infections. PMID- 11120950 TI - Complexity and diversity of Klebsiella pneumoniae strains with extended-spectrum beta-lactamases isolated in 1994 and 1996 at a teaching hospital in Durban, South Africa. AB - beta-Lactamase production was investigated in cultures of 25 Klebsiella pneumoniae isolates isolated at a hospital in Durban, South Africa, in 1994 and 1996. Twenty of these isolates gave ceftazidime MIC/ceftazidime plus clavulanate MIC ratios of >/=8, implying production of extended-spectrum beta-lactamases (ESBLs), and DNA sequencing identified an ESBL gene (bla(TEM-53)) in a further two isolates. Pulsed-field gel electrophoresis (PFGE) defined 4 distinct strains among the 12 isolates collected in 1994 and 9 distinct strains among the 13 isolates collected in 1996. In three cases, multiple isolates from single patients varied in their PFGE profiles and antibiograms, implying mixed colonization or infection. Isoelectric focusing and DNA hybridization found both TEM and SHV enzymes and their genes in all 25 isolates. Many isolates had multiple identical or different beta-lactamase gene variants, with at least 84 bla(SHV) and bla(TEM) gene copies among the 25 organisms. Sequencing identified the genes for the SHV-1, -2, and -5 enzymes and for four new SHV types (SHV-19, 20, -21, and -22). These new SHV variants had novel mutations remote from sites known to affect catalytic activity. Sequencing also found the genes for TEM-1, TEM-53, and one novel type, TEM-63. All the isolates had multiple and diverse plasmids. These complex and diverse patterns of ESBL production and strain epidemiology are far removed from the concept of an ESBL outbreak and suggest a situation in which ESBL production has become endemic and in which evolution is generating a wide range of enzyme combinations. This complexity and diversity complicates patient management and the design of antibiotic use policies. PMID- 11120951 TI - Effects of cytokines and fluconazole on the activity of human monocytes against Candida albicans. AB - This study evaluates the effects of cytokines, used singly and in combination, on the microbicidal activity of human monocyte-derived macrophages (MDM) against intracellular Candida albicans in the presence and absence of fluconazole. In the absence of fluconazole, the addition of tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), gamma interferon (IFN-gamma), or IL-4 had no effect on the growth of C. albicans. In contrast, the addition of granulocyte-macrophage colony-stimulating factor (GM-CSF) resulted in decreased growth (P < 0.05), while the addition of IL-10 resulted in increased growth (P < 0.01). In the presence of fluconazole, only the addition of IFN-gamma resulted in an increase in the growth of C. albicans. In the presence or absence of fluconazole, all cytokine combinations except IFN-gamma plus GM-CSF caused significant decreases in growth (P < 0.01). IL-10 and IL-4 did not influence the activity of TNF-alpha or IL 1beta. In the absence or presence of C. albicans the addition of fluconazole, all of the cytokines studied, and combinations of fluconazole and selected cytokines caused increases in nitric oxide (NO) production (P < 0.01). Similar observations were made for superoxide (O(2)(-)) only in the presence of C. albicans. The greatest concentrations of NO and O(2)(-) were produced when C. albicans alone was present in the assays. Our results demonstrate that in the presence of low concentrations of fluconazole (0.1 times the MIC), selected cytokines and their combinations significantly increase the microbicidal activity of MDM against intracellular C. albicans. PMID- 11120952 TI - Identification and characterization of inhibitors of multidrug resistance efflux pumps in Pseudomonas aeruginosa: novel agents for combination therapy. AB - Whole-cell assays were implemented to search for efflux pump inhibitors (EPIs) of the three multidrug resistance efflux pumps (MexAB-OprM, MexCD-OprJ, MexEF-OprN) that contribute to fluoroquinolone resistance in clinical isolates of Pseudomonas aeruginosa. Secondary assays were developed to identify lead compounds with exquisite activities as inhibitors. A broad-spectrum EPI which is active against all three known Mex efflux pumps from P. aeruginosa and their close Escherichia coli efflux pump homolog (AcrAB-TolC) was discovered. When this compound, MC 207,110, was used, the intrinsic resistance of P. aeruginosa to fluoroquinolones was decreased significantly (eightfold for levofloxacin). Acquired resistance due to the overexpression of efflux pumps was also decreased (32- to 64-fold reduction in the MIC of levofloxacin). Similarly, 32- to 64-fold reductions in MICs in the presence of MC-207,110 were observed for strains with overexpressed efflux pumps and various target mutations that confer resistance to levofloxacin (e.g., gyrA and parC). We also compared the frequencies of emergence of levofloxacin-resistant variants in the wild-type strain at four times the MIC of levofloxacin (1 microg/ml) when it was used either alone or in combination with EPI. In the case of levofloxacin alone, the frequency was approximately 10(-7) CFU/ml. In contrast, with an EPI, the frequency was below the level of detection (<10(-11)). In summary, we have demonstrated that inhibition of efflux pumps (i) decreased the level of intrinsic resistance significantly, (ii) reversed acquired resistance, and (iii) resulted in a decreased frequency of emergence of P. aeruginosa strains that are highly resistant to fluoroquinolones. PMID- 11120953 TI - Molecular basis of high-level ciprofloxacin resistance in Neisseria gonorrhoeae strains isolated in Denmark from 1995 to 1998. AB - In Denmark surveillance of the in vitro susceptibility to ciprofloxacin of Neisseria gonorrhoeae was established in 1990. The proportion of N. gonorrhoeae strains with decreased susceptibility or resistance to ciprofloxacin (MIC >/= 0.06 microg/ml) was low (0.3 to 2.3%) up to 1995. Between 1995 and 1998 the rate of less-susceptible and resistant strains rose from 6.9 to 13.2%. Among ciprofloxacin-resistant strains (MIC >/= 1 microg/ml), 81% were highly resistant (MIC >/= 4 microg/ml). Thirty-five N. gonorrhoeae strains (40 isolates) for which ciprofloxacin MICs were 4 to 32 microg/ml were investigated for the frequency and patterns of mutations within the gyrA and parC genes. The quinolone resistance determining regions of the gyrA and parC genes were amplified by PCR, and the amplicons were directly sequenced. Alterations at Ser-91 and Asp-95 in GyrA and a single or double alteration in ParC were identified in 32 strains (91%). Ser-91 to-Phe and Asp-95-to-Gly alterations in GyrA were detected in 28 strains (80%). The most common ParC alteration, Asp-86 to Asn, was found in 19 strains (54%). The strains were analyzed for genetic relationship by pulsed-field gel electrophoresis (PFGE). The analysis showed that nine strains with the same mutation pattern in the gyrA and parC genes, originating from different geographical areas over 3 years, had the same PFGE patterns after SpeI as well as NheI digestion (only one strain with one band difference in the NheI pattern), suggesting that a resistant clone had spread worldwide. The results from this study strongly suggest that double gyrA mutations plus a parC mutation(s) play an important role in the development of high-level fluoroquinolone resistance in N. gonorrhoeae. PMID- 11120954 TI - Studies of in vitro activities of voriconazole and itraconazole against Aspergillus hyphae using viability staining. AB - The minimal fungicidal concentrations (MFCs) of voriconazole and itraconazole for five clinical isolates each of Aspergillus terreus, Aspergillus fumigatus, Aspergillus flavus, and Aspergillus niger were determined by a broth macrodilution method. Conidial suspensions as inocula were compared to hyphae as inocula since the invasive form of aspergillosis is manifested by the appearance of hyphal structures. In addition, cell viability staining with the dye FUN-1 was performed to assess time-dependent damage of hyphae exposed to various concentrations of the antifungal agents. With conidial inocula the MFC ranges of voriconazole were 0.5 to 4 microg/ml and those of itraconazole were 0.25 to 2 microg/ml, whereas the MFCs (2 to >16 microg/ml) with hyphal inocula were substantially higher (P < 0.01) for both itraconazole and voriconazole. Only minor differences between the tested antifungals were observed since 16 of 20 and 17 of 20 of the isolates of Aspergillus spp. tested appeared to be killed by voriconazole and itraconazole, respectively. The results of FUN-1 viability staining correlated closely to colony counts, but various time- and dose dependent levels of viability of hyphae were also observed. In conclusion, our study demonstrates the importance of the type of inoculum used to test antifungals and the applicability of FUN-1 staining as a rapid and sensitive method for assaying the viability of hyphae. PMID- 11120955 TI - Validation of a noninvasive, real-time imaging technology using bioluminescent Escherichia coli in the neutropenic mouse thigh model of infection. AB - A noninvasive, real-time detection technology was validated for qualitative and quantitative antimicrobial treatment applications. The lux gene cluster of Photorhabdus luminescens was introduced into an Escherichia coli clinical isolate, EC14, on a multicopy plasmid. This bioluminescent reporter bacterium was used to study antimicrobial effects in vitro and in vivo, using the neutropenic mouse thigh model of infection. Bioluminescence was monitored and measured in vitro and in vivo with an intensified charge-coupled device (ICCD) camera system, and these results were compared to viable-cell determinations made using conventional plate counting methods. Statistical analysis demonstrated that in the presence or absence of antimicrobial agents (ceftazidime, tetracycline, or ciprofloxacin), a strong correlation existed between bioluminescence levels and viable cell counts in vitro and in vivo. Evaluation of antimicrobial agents in vivo could be reliably performed with either method, as each was a sound indicator of therapeutic success. Dose-dependent responses could also be detected in the neutropenic-mouse thigh model by using either bioluminescence or viable cell counts as a marker. In addition, the ICCD technology was examined for the benefits of repeatedly monitoring the same animal during treatment studies. The ability to repeatedly measure the same animals reduced variability within the treatment experiments and allowed equal or greater confidence in determining treatment efficacy. This technology could reduce the number of animals used during such studies and has applications for the evaluation of test compounds during drug discovery. PMID- 11120956 TI - Molecular modeling approach to understanding the mode of action of L-nucleosides as antiviral agents. AB - A series of unnatural L-nucleosides such as 3TC, FTC and L-FMAU have been found to be potent antiviral agents. The mode of action of L-nucleosides has been found to be similar to that of D-nucleosides as antiviral agents, despite their unnatural stereochemistry, that is, nucleotide formation by kinases followed by interaction with the reverse transcriptase (RT) of HIV or DNA polymerase. To date, the mode of action of nucleoside inhibitors at the molecular level with respect to the active conformations of the 5'-triphosphates as well as the interaction with the RT is not known. Recently, the X-ray crystal structure of the RT-DNA-dTTP catalytic complex has been reported. Computer modeling has been performed for several pairs of D- and L-nucleoside inhibitors using the HIV-1 RT model and crystal coordinate data from a subset of the protein surrounding the deoxynucleoside triphosphate (dNTP) binding pocket region. Results from our modeling studies of D-/L-zidovudine, D-/L-3TC, D-/L-dideoxycytosine triphosphates, dTTP and dCTP show that their binding energies correlate with the reported 50% effective concentrations. Modeling results are also discussed with respect to favorable conformations of each inhibitor at the dNTP site in the polymerization process. Additionally, the clinically important M184V mutation, which confers resistance against 3TC and FTC, was studied with our modeling system. The binding energy patterns of nucleoside inhibitors at the M184V mutation site correlate with the reported antiviral data. PMID- 11120957 TI - Antimalarial activities of peptide antibiotics isolated from fungi. AB - Malaria caused by Plasmodium falciparum is a major public health problem in the developing countries of the world. Clinical treatment of malaria has become complicated due to the occurrence of infections caused by drug resistant parasites. Secondary metabolites from fungi are an attractive source of chemotherapeutic agents. This work reports the isolation and in vitro antiplasmodial activities of peptide antibiotics of fungal origin. The three peptide antibiotics used in this study were efrapeptins, zervamicins, and antiamoebin. The high-performance liquid chromatography-purified peptides were characterized by nuclear magnetic resonance and mass spectral analysis. All three fungal peptides kill P. falciparum in culture with 50% inhibitory concentrations in the micromolar range. A possible mode of action of these peptide antibiotics on P. falciparum is presented. PMID- 11120958 TI - Pharmacokinetics of oral acyclovir in neonates and in infants: a population analysis. AB - Acyclovir is approved for the treatment of herpes simplex virus (HSV) and varicella-zoster virus (VZV) infections in children by the intravenous and oral routes. However, its use by the oral route in children younger than 2 years of age is limited due to a lack of pharmacokinetic data. The objectives of the present study were to determine the typical pharmacokinetics of an oral suspension of acyclovir given to children younger than 2 years of age and the interindividual variabilities in the values of the pharmacokinetic parameters in order to support the proposed dosing regimen (24 mg/kg of body weight three times a day for patients younger than 1 month of age or four times a day otherwise). Children younger than age 2 years with HSV or VZV infections were enrolled in a multicenter study. Children were treated for at least 5 days with an acyclovir oral suspension. Plasma samples were obtained at steady state, before acyclovir administration, and at 2, 3, 5, and 8 h after acyclovir administration. Acyclovir concentrations were measured by radioimmunoassay. The data were analyzed by a population approach. Data for 79 children were considered in the pharmacokinetic study (212 samples, 1 to 5 samples per patient). Acyclovir clearance was related to the estimated glomerular filtration rate, body surface area, and serum creatinine level. The volume of distribution was related to body weight. The elimination half-life decreased sharply during the first month after birth, from 10 to 15 h to 2.5 h. Bioavailability was 0.12. The interindividual variability was less pronounced when the parameters were normalized with respect to body weight. Hence, dosage adjustment by body weight is recommended for this population. Simulations showed that the length of time that acyclovir remains above the 50% inhibitory concentration during a 24-h period was more than 12 h for HSV but not for VZV. The proposed dosing regimen seems adequate for the treatment of HSV infections, while for the treatment of VZV infections, a twofold increase in the dose seems necessary for children older than age 3 months. PMID- 11120959 TI - Mechanism of action of 1-beta-D-2,6-diaminopurine dioxolane, a prodrug of the human immunodeficiency virus type 1 inhibitor 1-beta-D-dioxolane guanosine. AB - (-)-beta-D-2,6-Diaminopurine dioxolane (DAPD), is a nucleoside reverse transcriptase (RT) inhibitor with activity against human immunodeficiency virus type 1 (HIV-1). DAPD, which was designed as a water-soluble prodrug, is deaminated by adenosine deaminase to give (-)-beta-D-dioxolane guanine (DXG). By using calf adenosine deaminase a K(m) value of 15 +/- 0.7 microM was determined for DAPD, which was similar to the K(m) value for adenosine. However, the k(cat) for DAPD was 540-fold slower than the k(cat) for adenosine. In CEM cells and peripheral blood mononuclear cells exposed to DAPD or DXG, only the 5' triphosphate of DXG (DXG-TP) was detected. DXG-TP is a potent alternative substrate inhibitor of HIV-1 RT. Rapid transient kinetic studies show the efficiency of incorporation for DXG-TP to be lower than that measured for the natural substrate, 2'-deoxyguanosine 5'-triphosphate. DXG-TP is a weak inhibitor of human DNA polymerases alpha and beta. Against the large subunit of human DNA polymerase gamma a K(i) value of 4.3 +/- 0.4 microM was determined for DXG-TP. DXG showed little or no cytotoxicity and no mitochondrial toxicity at the concentrations tested. PMID- 11120960 TI - Effect of xylitol on growth of Streptococcus pneumoniae in the presence of fructose and sorbitol. AB - Xylitol is effective in preventing acute otitis media by inhibiting the growth of Streptococcus pneumoniae. To clarify this inhibition we used fructose, which is known to block similar growth inhibition observed in Streptococcus mutans. In addition, we evaluated the efficacy of sorbitol in inhibiting the growth of pneumococci, as sorbitol is widely used for indications similar to those for which xylitol is used. The addition of 5% xylitol to the growth medium resulted in marked growth inhibition, an effect which was totally eliminated in the presence of 1, 2.5, or 5% fructose but not in the presence of 1 or 5% glucose, 1% galactose, or 1% sucrose. This finding implies that xylitol-induced inhibition of pneumococcal growth is mediated via the fructose phosphotransferase system in a way similar to that in which mutans group streptococcal growth is inhibited. The addition of sorbitol at concentrations of 1, 2.5, or 5% to the growth medium did not affect the growth of pneumococci and neither inhibited nor enhanced the xylitol-induced growth impairment. Thus, it seems that xylitol is the only commercially used sugar substitute proven to have an antimicrobial effect on pneumococci. PMID- 11120961 TI - Pharmacokinetics of the new ketolide telithromycin (HMR 3647) administered in ascending single and multiple doses. AB - Telithromycin (HMR 3647) is a novel ketolide antimicrobial with good activity against both common and atypical respiratory pathogens, including many resistant strains. This randomized, three-period crossover study determined the dose proportionality of telithromycin pharmacokinetics after single and multiple dosing in healthy subjects. In each treatment period, subjects received a single oral dose of 400, 800 or 1,600 mg of telithromycin followed 4 days later by the same dose once daily for 7 days. Blood and urine samples were taken throughout the study for determination of pharmacokinetic parameters for telithromycin and RU 76363, its main metabolite. Telithromycin and RU 76363 achieved steady state within 2 to 3 days of once-daily dosing. A slight accumulation of telithromycin was observed after 7 days of therapy, with values of the area under the concentration-time curve from 0 to 24 h approximately 1.5 times higher than those achieved with the single dose. The pharmacokinetics of telithromycin and RU 76363 deviated moderately from dose proportionality. At a dose of 800 mg/day, telithromycin attained mean maximal and trough plasma concentrations of 2.27 and 0. 070 mg/liter respectively. Elimination was biphasic; initial and terminal half lives were 2.87 and 9.81 h for the 800-mg dose. Study medication was well tolerated, although adverse events tended to be more frequent at the 1,600-mg dose. This study showed that telithromycin was generally well tolerated and suggests that a once-daily 800-mg oral dose of telithromycin maintains an effective concentration in plasma for the treatment of respiratory tract infections involving the key respiratory pathogens. PMID- 11120962 TI - Effect of dosing schedule on pharmacokinetics of alpha interferon and anti-alpha interferon neutralizing antibody in mice. AB - The influences of dosing time and dosing schedule on the plasma alpha interferon (IFN-alpha) concentration and the production of anti-IFN-alpha neutralizing antibodies were investigated in ICR male mice adapted to cycles of 12 h of light and 12 h of dark. In mice pretreated with IFN-alpha for 21 days, the plasma IFN alpha concentrations were significantly lower than those in control mice (P < 0.01). The clearance of IFN-alpha and its volume of distribution obtained at steady state were significantly higher in the animals with IFN-alpha pretreatment than in the mice without IFN-alpha pretreatment. The area under the concentration time curve and the mean residence time of IFN-alpha were significantly smaller in IFN-alpha-pretreated animals than in control animals. The plasma IFN-alpha levels (measured 2 h after dosing) were significantly lower in mice treated daily with IFN-alpha, while the anti-IFN-alpha neutralizing antibody levels (measured 24 h after dosing) were significantly increased on days 15 and 21 of treatment. Plasma IFN-alpha levels were significantly decreased in association with the production of anti-IFN-alpha neutralizing antibodies in mice treated with IFN-alpha daily at either 0900 or 2100 h. By contrast, the plasma IFN-alpha levels (measured 2 h after dosing) remained stable in mice treated with IFN-alpha at 0900 h on alternate days, while they were significantly lower after 21 days of treatment in mice treated with IFN-alpha at 2100 h on alternate days. These changes were associated with a significant increase in the levels of anti-IFN-alpha neutralizing antibodies in the latter group. The present findings suggest that an appropriate dosing schedule and/or dosing time for IFN-alpha may reduce the level of production of anti-IFN-alpha neutralizing antibodies in experimental and clinical situations. PMID- 11120963 TI - Pharmacokinetics and pharmacodynamics of intravenous artesunate in severe falciparum malaria. AB - To provide novel data relating to the dispositions, effects, and toxicities of the artemisinin derivatives in severe malaria, we studied 30 Vietnamese adults with slide-positive falciparum malaria treated with intravenous artesunate. Twelve patients with complications (severe; group 1) and 8 patients without complications but requiring parenteral therapy (moderately severe; group 2) received 120 mg of artesunate by injection, and 10 patients with moderately severe complications (group 3) were given 240 mg by infusion. Serial concentrations of artesunate and its active metabolite dihydroartemisinin in plasma were measured by high-performance liquid chromatography. The time to 50% parasite clearance (PCT(50)) was determined from serial parasite densities. Full clinical (including neurological) assessments were performed at least daily. In noncompartmental pharmacokinetic analyses, group mean artesunate half-lives (t(1/2)) were short (range, 2.3 to 4.3 min). The dihydroartemisinin t(1/2) (range, 40 to 64 min), clearance (range, 0.73 to 1.01 liters/h/kg), and volume of distribution (range, 0.77 to 1.01 liters/kg) were also similar both across the three patient groups (P > 0.1) and to previously reported values for patients with uncomplicated malaria. Parasite clearance was prompt (group median PCT(50) range 6 to 9 h) and clinical recovery was complete under all three regimens. These data indicate that the pharmacokinetics of artesunate and dihydroartemisinin are not influenced by the severity of malaria. Since the pharmacokinetic parameters for both artesunate and dihydroartemisinin were similar regardless of whether injection or infusion was used, artesunate can be considered a prodrug that is converted stoichiometrically to dhydroartemisinin. Conventional doses of artesunate are safe and effective when given to patients with complications of falciparum malaria. PMID- 11120964 TI - Efficacies of lipophilic inhibitors of dihydrofolate reductase against parasitic protozoa. AB - Competitive inhibitors of dihydrofolate reductase (DHFR) are used in chemotherapy or prophylaxis of many microbial pathogens, including the eukaryotic parasites Plasmodium falciparum and Toxoplasma gondii. Unfortunately, point mutations in the DHFR gene can confer resistance to inhibitors specific to these pathogens. We have developed a rapid system for testing inhibitors of DHFRs from a variety of parasites. We replaced the DHFR gene from the budding yeast Saccharomyces cerevisiae with the DHFR-coding region from humans, P. falciparum, T. gondii, Pneumocystis carinii, and bovine or human-derived Cryptosporidium parvum. We studied 84 dicyclic and tricyclic 2,4-diaminopyrimidine derivatives in this heterologous system and identified those most effective against the DHFR enzymes from each of the pathogens. Among these compounds, six tetrahydroquinazolines were effective inhibitors of every strain tested, but they also inhibited the human DHFR and were not selective for the parasites. However, two quinazolines and four tetrahydroquinazolines were both potent and selective inhibitors of the P. falciparum DHFR. These compounds show promise for development as antimalarial drugs. PMID- 11120965 TI - Effects of amoxicillin, gentamicin, and moxifloxacin on the hemolytic activity of Staphylococcus aureus in vitro and in vivo. AB - In Staphylococcus aureus infection hemolysis caused by the extracellular protein alpha-toxin encoded by hla is thought to contribute significantly to its multifactorial virulence. In vitro, subinhibitory concentrations of beta-lactam antibiotics and fluoroquinolones increase the levels of hla and alpha-toxin expression, whereas aminoglycosides decrease the levels of hla and alpha-toxin expression. In the present study we investigated the effects of subinhibitory concentrations of amoxicillin, gentamicin, and moxifloxacin on hla and alpha toxin expression and total hemolysis of S. aureus strain 8325-4, a high-level alpha-toxin producer, and its alpha-toxin-negative mutant, DU 1090, in vitro and in a rat model of chronic S. aureus infection. The levels of expression of hla and alpha-toxin and total hemolysis did not differ significantly when amoxicillin, gentamicin, or moxifloxacin was added to cultures of S. aureus strain 8325-4. In vivo, strain 8325-4 induced a significantly increased level of hemolysis in infected pouches compared to that in uninfected control pouches, but the hemolysis was reduced to control levels by treatment with doses of amoxicillin, gentamicin, or moxifloxacin that reduced bacterial numbers by 2 orders of magnitude. Additionally, the effects of subinhibitory concentrations of the three antibiotics on total hemolysis of four methicillin-resistant S. aureus and three methicillin-sensitive S. aureus (MSSA) clinical isolates were assessed in vitro. A significant increase in total hemolysis was observed for only one MSSA strain when it was treated with amoxicillin but not when it was treated with moxifloxacin or gentamicin. When purified alpha-toxin was incubated with purified human neutrophil elastase, alpha-toxin was cleaved nearly completely. The results suggest that the penicillin-induced increases in S. aureus alpha-toxin expression are strain dependent, that reduction of bacterial numbers in vivo counteracts this phenomenon effectively, and finally, that in localized S. aureus infections alpha-toxin activity is controlled by neutrophil elastase. PMID- 11120966 TI - In vitro and in vivo antibacterial activities of L-084, a novel oral carbapenem, against causative organisms of respiratory tract infections. AB - L-084 (a prodrug of LJC 11,036 [L-036]) is a new oral carbapenem. Here we compared the in vitro and in vivo antibacterial activities of L-036 with those of imipenem, faropenem, ceditoren-pivoxil, cefdinir, amoxicillin, and levofloxacin. The MICs at which 90% of the isolates were inhibited of L-036 against methicillin susceptible staphylococci, Streptococcus pneumoniae including penicillin resistant organisms, Escherichia coli, Klebsiella pneumoniae, Haemophilus influenzae including ampicillin-resistant organisms, Legionella pneumophila, and Moraxella catarrhalis were equal to or less than 1 microg/ml. In pharmacokinetics studies of L-084 in lungs of mice, the maximum concentration in serum, half-life, and area under the concentration-time curve of this drug were 9.09 microg/g of tissue, 6.18 h, and 31.0 microg. h/ml, respectively. In murine respiratory infection models of penicillin-susceptible and -resistant S. pneumoniae and H. influenzae, the efficacies of L-084 were better than those of reference drugs. Our results indicate that the in vitro high potency and good distribution in the lungs might be the underlying mechanisms of its efficacy in the murine model of pneumonia. PMID- 11120967 TI - In vivo activity of evernimicin (SCH 27899) against methicillin-resistant Staphylococcus aureus in experimental infective endocarditis. AB - Currently, there exist few satisfactory alternatives to vancomycin for therapy of serious methicillin-resistant Staphylococcus aureus (MRSA) infections. We employed a rat model of aortic valve endocarditis to assess the potential efficacy of evernimicin (SCH 27899) compared with vancomycin against infection with a strain susceptible to both agents (MICs of 0.25 and 0.50 microg/ml, respectively). Infected animals were assigned to one of three groups: controls (no treatment), evernimicin at 60 mg/kg of body weight by intravenous (i.v.) infusion once daily, or vancomycin at 150 mg/kg of body weight per day by continuous i.v. infusion. Therapy was administered for 5.5 days. At the start of therapy, colony counts in vegetations were 6.63 +/- 0.44 log(10) CFU/g. In both treatment groups, bacterial density within vegetations was significantly reduced in comparison with control animals that had not been treated. Final colony counts were as follows (mean +/- standard deviation): controls, 10.12 +/- 1.51 log(10) CFU/g of vegetation; evernimicin, 7.22 +/- 2.91 log(10) CFU/g of vegetation; vancomycin, 5.65 +/- 1.76 log(10) CFU/g of vegetation. The difference between the evernimicin and vancomycin groups was not significant. These results confirmed the bacteriostatic activity of evernimicin in vivo in an experimental model of severe MRSA infection. PMID- 11120968 TI - In vitro antimicrobial susceptibility testing of bacterial enteropathogens causing traveler's diarrhea in four geographic regions. AB - The emergence of resistant enteropathogens has been reported worldwide. Few data are available on the contemporary in vitro activities of commonly used antimicrobial agents against enteropathogens causing traveler's diarrhea (TD). The susceptibility patterns of antimicrobial agents currently available or under evaluation against pathogens causing TD in four different areas of the world were evaluated. Pathogens were identified in stool samples from U.S., Canadian, or European adults (18 years of age or older) with TD during 1997, visiting India, Mexico, Jamaica, or Kenya. MICs of 11different antimicrobials were determined against 284 bacterial enteropathogens by the agar dilution method. Ciprofloxacin, levofloxacin, ceftriaxone, and azithromycin were highly active in vitro against the enteropathogens, while traditional antimicrobials such as ampicillin, trimethoprim, and trimethoprim/sulfamethoxazole showed high levels and high frequencies of resistance. Rifaximin, a promising and poorly absorbable drug, had an MIC at which 90% of the strains tested were inhibited of 32 microg/ml, 250 times lower than the concentration of this drug in the stools. Amdinocillin, nalidixic acid, and doxycycline showed moderate activity. Fluoroquinolones are still the drugs of choice for TD in most regions of the world, although our study has a limitation due to the lack of Escherichia coli samples from Kenya and possible bias in selection of the patients for evaluation. Azithromycin and rifaximin should be considered as promising new agents. The widespread in vitro resistance of the traditional antimicrobial agents reported since the 1980s and the new finding of resistance to fluoroquinolones in Southeast Asia are the main reasons for monitoring carefully the antimicrobial susceptibility patterns worldwide and for developing and evaluating new antimicrobial agents for the treatment of TD. PMID- 11120969 TI - Activity of moxifloxacin by itself and in combination with ethambutol, rifabutin, and azithromycin in vitro and in vivo against Mycobacterium avium. AB - Moxifloxacin activity against Mycobacterium avium complex (MAC) was evaluated in vitro against 25 strains. The MIC was determined to range from 0.125 to 2.0 microg/ml. In addition, U937 macrophage monolayers infected with MAC strain 101 (serovar 1) were treated with moxifloxacin (0.25 to 8 microg/ml) daily, and the number of intracellular bacteria was quantitated after 4 days. Moxifloxacin showed inhibitory activity at 0.5 microg/ml and higher. To assess the activity of moxifloxacin containing regimens in vivo, we infected C57BL bg(+)/bg(+) mice with 3 x 10(7) MAC strain 101 bacteria intravenously. One week later treatment was begun with the following: (i) moxifloxacin (50 mg/kg/day or 100 mg/kg/day), ethambutol (100 mg/kg/day), or a combination of moxifloxacin and ethambutol; or (ii) moxifloxacin (100 mg/kg/day), azithromycin (200 mg/kg/day), or rifabutin (40 mg/kg/day) as oral monotherapy; or (iii) all permutations of two-drug therapy or all three drugs in combination. All groups contained at least 14 animals, and the control group received the drug vehicle. After 4 weeks, quantitative blood cultures were obtained and the number of bacteria in liver and spleen was quantitated. Moxifloxacin, ethambutol, and azithromycin were active as single agents in liver, spleen, and blood. Rifabutin showed inhibitory activity only in the blood. Two-drug combinations containing azithromycin were no more active than azithromycin alone. Similarly, the three-drug combination was not more active than azithromycin alone in the spleen. Rifabutin did not add to the activity of any other single agent or two-drug combination. Moxifloxacin at both concentrations in combination with ethambutol was significantly more active than each drug alone. PMID- 11120970 TI - Unusual susceptibility of a multidrug-resistant yeast strain to peptidic antifungals. AB - The susceptibility of Saccharomyces cerevisiae JG436 multidrug transporter deletion mutant, Deltapdr5, to several antifungal agents was compared to that of JG436-derived JGCDR1 and JGCaMDR1 transformants, harboring the CDR1 and CaMDR1 genes, encoding the main drug-extruding membrane proteins of Candida albicans. The JGCDR1 and JGCaMDR1 yeasts demonstrated markedly diminished susceptibility to the azole antifungals, terbinafine and cycloheximide, while that to amphotericin B was unchanged. Surprisingly, JGCDR1 but not JGCaMDR1 cells showed enhanced susceptibility to peptidic antifungals, rationally designed compounds containing inhibitors of glucosamine-6-phosphate synthase. It was found that these antifungal oligopeptides, as well as model oligopeptides built of proteinogenic amino acids, were not effluxed from JGCDR1 cells. Moreover, they were taken up by these cells at rates two to three times higher than by JG436. The tested oligopeptides were rapidly cleaved to constitutive amino acids by cytoplasmic peptidases. Studies on the mechanism of the observed phenomenon suggested that an additive proton motive force generated by Cdr1p stimulated uptake of oligopeptides into JGCDR1 cells, thus giving rise to the higher antifungal activity of FMDP [N(3)-(4-methoxyfumaroyl)-L-2,3-diaminopropanoic acid]-peptides. PMID- 11120971 TI - Antiviral L-nucleosides specific for hepatitis B virus infection. AB - A unique series of simple "unnatural" nucleosides has been discovered to inhibit hepatitis B virus (HBV) replication. Through structure-activity analysis it was found that the 3'-OH group of the beta-L-2'-deoxyribose of the beta-L-2' deoxynucleoside confers specific antihepadnavirus activity. The unsubstituted nucleosides beta-L-2'-deoxycytidine, beta-L-thymidine, and beta-L-2' deoxyadenosine had the most potent, selective, and specific antiviral activity against HBV replication. Human DNA polymerases (alpha, beta, and gamma) and mitochondrial function were not affected. In the woodchuck model of chronic HBV infection, viral load was reduced by as much as 10(8) genome equivalents/ml of serum and there was no drug-related toxicity. In addition, the decline in woodchuck hepatitis virus surface antigen paralleled the decrease in viral load. These investigational drugs, used alone or in combination, are expected to offer new therapeutic options for patients with chronic HBV infection. PMID- 11120972 TI - Pharmacological basis for concentration-controlled therapy with zidovudine, lamivudine, and indinavir. AB - Conventional antiretroviral therapy involves administration of standard fixed doses to adults and adolescents. This approach ignores interindividual variability in pharmacokinetics and results in substantial differences in systemic concentrations among patients. Thus, variability in systemic concentrations contributes to variability in response to therapy. This study was designed to evaluate the feasibility and safety of a regimen of zidovudine, lamivudine, and indinavir designed to achieve select target concentrations versus standard dose therapy. Twenty-four antiretroviral-naive subjects completed the 24 week study; 13 received standard therapy, and 11 received concentration controlled therapy. There were no differences in baseline characteristics. Oral clearance for all three drugs was not different between weeks 2 and 28; average ratios of week 2 oral clearance to week 28 oral clearance were 0.95, 1.09, and 1.06 for zidovudine, lamivudine, and indinavir, respectively, with 95% confidence intervals including 1. The selected target concentrations were average steady state concentrations of 0.19 mg/liter for zidovudine and 0.44 mg/liter for lamivudine and a trough concentration of 0.15 mg/liter for indinavir; mean concentrations achieved at week 28 in the concentration-controlled arm were 0.20, 0.54, and 0.19 mg/liter, respectively. Concentration-controlled therapy significantly reduced interpatient variability in zidovudine concentrations and significantly increased indinavir concentrations. There was no difference in adverse drug effects or adherence. This investigation has provided a pharmacologic basis for concentration-controlled therapy by demonstrating that it is feasible and has a safety profile no different from that of standard therapy. Additional studies to evaluate the virologic effect of the concentration controlled approach to antiretroviral therapy are warranted. PMID- 11120973 TI - In vitro activity of trovafloxacin against Bacteroides fragilis in mixed culture with either Escherichia coli or a vancomycin- resistant strain of Enterococcus faecium determined by an anaerobic time-kill technique. AB - To determine the efficacy of trovafloxacin as a possible treatment for intra abdominal abscesses, we have developed an anaerobic time-kill technique using different inocula to study the in vitro killing of Bacteroides fragilis in pure culture or in mixed culture with either Escherichia coli or a vancomycin resistant strain of Enterococcus faecium (VREF). With inocula of 5 x 10(5) CFU/ml and trovafloxacin concentrations of /=6.1 (log(10) CFU/ml) was attained with all pure and mixed cultures within 24 h. With inocula of 10(8) CFU/ml, a similar E(max) and a similar concentration to produce 50% of E(max) (EC(50)) for B. fragilis were found in both pure cultures and mixed cultures with E. coli. However, to produce a similar killing of B. fragilis in the mixed cultures with VREF, a 14-fold increase in the concentration of trovafloxacin was required. A vancomycin-susceptible strain of E. faecium and a trovafloxacin-resistant strain of E. coli were also found to confer a similar "protective" effect on B. fragilis against the activity of trovafloxacin. Using inocula of 10(9) CFU/ml, the activity of trovafloxacin was retained for E. coli and B. fragilis and was negligible against VREF. We conclude that this is a useful technique to study the anaerobic killing of mixed cultures in vitro and may be of value in predicting the killing of mixed infections in vivo. The importance of using mixed cultures and not pure cultures is clearly shown by the difference in the killing of B. fragilis in the mixed cultures tested. Trovafloxacin will probably be ineffective in the treatment of infections involving large numbers of enterococci. However, due to its ability to retain activity against large cultures of B. fragilis and E. coli, trovafloxacin could be beneficial in the treatment of intra-abdominal abscesses. PMID- 11120974 TI - Kinetic study of the inflammatory response in Streptococcus pneumoniae experimental pneumonia treated with the ketolide HMR 3004. AB - Patients still die from Streptococcus pneumoniae pneumonia after initiation of antibiotic therapy, when tissues are sterile and the pneumonia is clearing. There is growing evidence that overwhelming inflammation resulting from toxin release contributes to tissue injury, shock, and death. Monitoring host response may help us understand the consequences of antibiotic therapy for the inflammatory processes that occur in bacterial pneumonia. HMR 3004 is a ketolide that displays excellent in vitro activity against S. pneumoniae. In the present experiment, we investigated the chronology of inflammatory events that occur during pneumococcal pneumonia in mice treated with HMR 3004. Infection of mice with 10(7) CFU of living S. pneumoniae resulted in 100% mortality within 5 days. HMR 3004 given at 12.5 mg/kg of body weight/dose twice daily from 48 h postinfection achieved complete bacterial clearance from lungs and blood within 36 h and ensured survival of mice. Recruitment of neutrophils and monocytes from blood to lungs was significantly reduced, and nitric oxide release was totally prevented. Interleukin-6 secretion in lungs and blood became rapidly undetectable after initiation of therapy. Histological examination of lung tissue showed protection of interstitium against edema. By controlling bacterial invasion, HMR 3004 led to rapid and profound modifications of the host response in lungs, which may protect mice from deleterious inflammatory reactions. PMID- 11120975 TI - Disruption of an Enterococcus faecium species-specific gene, a homologue of acquired macrolide resistance genes of staphylococci, is associated with an increase in macrolide susceptibility. AB - The complete sequence (1,479 nucleotides) of msrC, part of which was recently reported by others using a different strain, was determined. This gene was found in 233 of 233 isolates of Enterococcus faecium but in none of 265 other enterococci. Disruption of msrC was associated with a two- to eightfold decrease in MICs of erythromycin azithromycin, tylosin, and quinupristin, suggesting that it may explain in part the apparent greater intrinsic resistance to macrolides of isolates of E. faecium relative to many streptococci. This endogenous, species specific gene of E. faecium is 53% identical to msr(A), suggesting that it may be a remote progenitor of the acquired macrolide resistance gene found in some isolates of staphylococci. PMID- 11120976 TI - Evaluation of current activities of fluoroquinolones against gram-negative bacilli using centralized in vitro testing and electronic surveillance. AB - Given the propensity for Enterobacteriaceae and clinically significant nonfermentative gram-negative bacilli to acquire antimicrobial resistance, consistent surveillance of the activities of agents commonly prescribed to treat infections arising from these organisms is imperative. This study determined the activities of two fluoroquinolones, levofloxacin and ciprofloxacin, and seven comparative agents against recent clinical isolates of Enterobacteriaceae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Stenotrophomonas maltophilia using two surveillance strategies: 1) centralized in vitro susceptibility testing of isolates collected from 27 hospital laboratories across the United States and 2) analysis of data from The Surveillance Network Database-USA, an electronic surveillance network comprising more than 200 laboratories nationwide. Regardless of the surveillance method, Enterobacteriaceae, P. aeruginosa, and A. baumannii demonstrated similar rates of susceptibility to levofloxacin and ciprofloxacin. Susceptibilities to the fluoroquinolones approached or exceeded 90% for all Enterobacteriaceae except Providencia spp. ( 25.0 microg/ml). Structure-activity relationships of these aphidicolan derivatives are discussed. PMID- 11120980 TI - Concentrations of gatifloxacin in plasma and urine and penetration into prostatic and seminal fluid, ejaculate, and sperm cells after single oral administrations of 400 milligrams to volunteers. AB - Gatifloxacin (GTX), a new fluoroquinolone with extended antibacterial activity, is an interesting candidate for the treatment of chronic bacterial prostatitis (CBP). Besides the antibacterial spectrum, the concentrations in the target tissues and fluids are crucial for the treatment of CBP. Thus, it was of interest to investigate its penetration into prostatic and seminal fluid. GTX concentrations in plasma, urine, ejaculate, prostatic and seminal fluid, and sperm cells were determined by a high-performance liquid chromatography method after oral intake of a single 400-mg dose in 10 male Caucasian volunteers in the fasting state. Simultaneous application of the renal contrast agent iohexol was used to estimate the maximal possible contamination of ejaculate and prostatic and seminal fluid by urine. GTX was well tolerated. The means (standard deviations) for the following parameters were as indicated: time to maximum concentration of drug in serum, 1.66 (0. 91) h; maximum concentration of drug in serum, 2.90 (0.39) microg/ml; area under the concentration-time curve from 0 to 24 h, 25.65 microg. h/ml; and half life, 7.2 (0.90) h. Within 12 h about 50% of the drug was excreted unchanged into the urine. The mean renal clearance was 169 ml/min. The gatifloxacin concentrations in ejaculate, seminal fluid, and prostatic fluid were in the range of the corresponding plasma concentrations which were 1.92 (0.27) microg/ml at approximately the same time point (4 h after drug intake). The concentrations in sperm cells (0.195, 0.076, and 0.011 microg/ml) could be determined in three subjects. The good penetration into prostatic and seminal fluid, the good tolerance, and the previously reported broad antibacterial spectrum suggest that GTX may be a good alternative for the treatment of chronic bacterial prostatitis. Clinical studies should be performed to confirm this assumption. PMID- 11120981 TI - If taken 1 hour before indinavir (IDV), didanosine does not affect IDV exposure, despite persistent buffering effects. AB - Concurrent administration of indinavir and didanosine significantly reduces the level of exposure to indinavir, but it is unclear how soon after didanosine administration indinavir may be given safely. We compared indinavir pharmacokinetics and gastric pH in 12 human immunodeficiency virus-positive patients by use of 800 mg of indinavir alone versus 800 mg of indinavir administered 1 h after didanosine administration. Median gastric pH was significantly higher when indinavir was taken after didanosine administration; however, no significant difference in the maximum concentration in plasma or the area under the concentration-time curve from time zero to 8 h was observed. Indinavir may be taken with a light meal 1 h following the administration of 400 mg of didanosine. PMID- 11120982 TI - Bacteriostatic and bactericidal activities of moxifloxacin against Coxiella burnetii. AB - The in vitro activity of moxifloxacin against Coxiella burnetii was compared to those of pefloxacin, ofloxacin, and doxycycline. MICs of moxifloxacin ranged from 0.5 to 1 microg/ml for the Nine Mile, Priscilla, and Q212 strains. Moxifloxacin was not bactericidal against C. burnetii at 4 microg/ml. PMID- 11120983 TI - Affinities of beta-lactams for penicillin binding proteins of Chlamydia trachomatis and their antichlamydial activities. AB - Binding affinities of beta-lactam antibiotics for the three penicillin binding proteins (PBPs) from Chlamydia trachomatis were determined in vitro and compared with their antichlamydial activities. Mecillinam selectively inhibited PBP1, with a 50% inhibitory concentration for PBP1 binding (0.2 microg/ml) similar to the MIC (0.1 microg/ml) and minimum bactericidal concentration (0.25 microg/ml). Although the other beta-lactams inhibited a wider range of PBPs than mecillinam, their antichlamydial activities were inferior to that of mecillinam. PMID- 11120984 TI - Furazolidone- and nitrofurantoin-resistant Helicobacter pylori: prevalence and role of genes involved in metronidazole resistance. AB - The prevalence of furazolidone, nitrofurantoin, and metronidazole resistance among Helicobacter pylori strains was assessed with 431 clinical isolates. Fifty two percent were metronidazole resistant, compared to 2% (7 of 431) with resistance to furazolidone and nitrofurantoin. All seven furazolidone- and nitrofurantoin-resistant isolates were also metronidazole resistant. rdxA, frxA, and fdxB knockouts did not result in furazolidone or nitrofurantoin resistance. These data suggest that furazolidone and nitrofurantoin may be good alternatives to metronidazole for treating H. pylori infection. PMID- 11120985 TI - SHV-14, a novel beta-lactamase variant in Klebsiella pneumoniae isolates from Nijmegen, The Netherlands. AB - Four ceftazidime-resistant isolates of a Klebsiella pneumoniae strain were collected from intensive care unit patients in Nijmegen, The Netherlands. These isolates had TEM-29 and SHV-14 beta-lactamases. SHV-14 is a novel variant, with two substitutions compared with the sequence of SHV-1: Ile8Phe and Arg43Ser. Its gene also had a silent C-->T mutation at nucleotide 481. The SHV-14 enzyme had slightly higher V(max) rates than SHV-1 for oxyimino-aminothiazolyl cephalosporins, but this activity was insufficient for the enzyme to count as an extended-spectrum beta-lactamase. PMID- 11120986 TI - In vitro and in vivo efficacies of T-3811ME (BMS-284756) against Mycoplasma pneumoniae. AB - T-3811, the free base of T-3811ME (BMS-284756), a new des-F(6)-quinolone, showed a potent in vitro activity (MIC at which 90% of the isolates tested are inhibited [MIC(90)], 0.0313 microg/ml) against Mycoplasma pneumoniae. The MIC(90) of T-3811 was 4-fold higher than that of clarithromycin but was 4- to 8-fold lower than those of trovafloxacin, gatifloxacin, gemifloxacin, and moxifloxacin and was 16- to 32-fold lower than those of levofloxacin, ciprofloxacin, and minocycline. In an experimental M. pneumoniae pneumonia model in hamsters, after the administration of T-3811ME (20 mg/kg of body weight as T-3811, once daily, orally) for 5 days, the reduction of viable cells of M. pneumoniae in bronchoalveolar lavage fluid was greater than those of trovafloxacin, levofloxacin, and clarithromycin (20 and 40 mg/kg, orally) (P < 0.05). PMID- 11120987 TI - Effective combination therapy for invasive pneumococcal pneumonia with ampicillin and intravenous immunoglobulins in a mouse model. AB - Intranasal immunotherapy for Streptococcus pneumoniae invasive pneumonia with polyvalent immunoglobulins (IVIG) was effective in mice against pneumonia but failed to prevent bacteremia. The combination of subcurative doses of IVIG and of ampicillin was fully protective. Such an approach, successfully applied in the preantibiotic era, offers new perspectives for modern therapies. PMID- 11120988 TI - Mutation in 23S rRNA responsible for resistance to 16-membered macrolides and streptogramins in Streptococcus pneumoniae. AB - Streptococcus pneumoniae clinical isolate BM4455 was resistant to 16-membered macrolides and to streptogramins. This unusual resistance phenotype was due to an A(2062)C (Escherichia coli numbering) mutation in domain V of the four copies of 23S rRNA. PMID- 11120989 TI - Nitrofurantoin is active against vancomycin-resistant enterococci. AB - The activity of nitrofurantoin was tested against 300 isolates of Enterococcus faecium, Enterococcus faecalis, and Enterococcus gallinarum. No isolates tested were resistant to nitrofurantoin (MIC, >/=128 microg/ml), including vancomycin resistant E. faecium isolates with vanA- and vanB-positive genotypes and vancomycin-resistant E. gallinarum isolates. We conclude that nitrofurantoin may provide effective treatment of urinary tract infections caused by vancomycin resistant enterococci. PMID- 11120990 TI - In vitro susceptibility testing methods for caspofungin against Aspergillus and Fusarium isolates. AB - We investigated the relevance of prominent reduction in turbidity macroscopically (MIC) and formation of aberrant hyphal tips microscopically (minimum effective concentration; MEC) in measuring the in vitro activity of caspofungin against Aspergillus and Fusarium. Caspofungin generated low MICs and MECs against Aspergillus, but not for Fusarium. While MICs increased inconsistently when the incubation time was prolonged, MEC appeared as a stable and potentially relevant endpoint in testing in vitro caspofungin activity. PMID- 11120991 TI - The ovine cathelicidin SMAP29 kills ovine respiratory pathogens in vitro and in an ovine model of pulmonary infection. AB - Cathelicidins are antimicrobial peptides from sheep (SMAP29 and SMAP34), rabbits (CAP11 and CAP18), rodents (CRAMP), and humans (FALL39, LL37, and h/CAP18). In a broth microdilution assay against nine ovine pathogens, SMAP29, SMAP34, mouse CRAMP, CAP18, CAP18(31), CAP18(28), CAP18(22), and CAP18(21a) were the most active, with MICs as low as 0.6 microg/ml. Other cathelicidins were less active. In lambs with pneumonia, 0.5 mg of SMAP29 reduced the concentration of bacteria in both bronchoalveolar lavage fluid and consolidated pulmonary tissues. Hence, the antimicrobial activity of SMAP29 suggests that it has applications in the treatment of respiratory tract infections. PMID- 11120992 TI - Genomic rearrangement of the mec regulator region mediated by insertion of IS431 in methicillin-resistant staphylococci. AB - Genomic diversification of the mec regulator region mediated by IS431 was investigated for clinical isolates of methicillin-resistant staphylococci. A single rearranged form of the mecR1 gene due to IS431 insertion was detected in the three staphylococcal species, while another type of mecR1 truncation with IS431 and an IS431 located downstream of mecI were found only in Staphylococcus haemolyticus. Genetic differentiation of IS431 and staphylococcal isolates suggested transmission of mecDNA with IS431-mediated rearrangement among different staphylococcal species. PMID- 11120993 TI - Prevalence and mechanisms of macrolide resistance in Streptococcus pyogenes in Santiago, Chile. AB - Thirty-two macrolide-resistant Streptococcus pyogenes isolates were found among 594 clinical isolates collected from 1990 to 1998 in Santiago, Chile, for an overall prevalence of 7.2%. Among the 32 resistant isolates, 28 (87.5%) presented the M phenotype and 4 (12. 5%) presented the MLS(B) phenotype. Serotyping and pulsed-field gel electrophoresis analysis showed genetic diversity among the resistant isolates. PMID- 11120995 TI - Comparative in vitro activities of ABT-773 against 362 clinical isolates of anaerobic bacteria. AB - The activity of ABT-773, a novel ketolide antibiotic, against clinical isolates of anaerobic bacteria was determined and compared to the activities of other antimicrobial agents. MICs at which 90% of isolates were inhibited (MIC(90)s) were 32 microg/ml, respectively, for Eubacterium spp., Lactobacillus spp., Clostridium difficile, and Clostridium ramosum. The MIC(90) for Bilophila wadsworthia, Bacteroides ureolyticus, and Campylobacter gracilis was 1 microg/ml, and that for Prevotella bivia and other Prevotella spp. was 0.5 microg/ml. The MIC(90) for Fusobacterium nucleatum was 8 microg/ml, and that for Fusobacterium mortiferum and Fusobacterium varium was >32 microg/ml. The MIC(90)s for the Bacteroides fragilis group were as follows: for B. fragilis, 8 microg/ml; for Bacteroides thetaiotaomicron, Bacteroides ovatus, Bacteroides distasonis, and Bacteroides uniformis, >32 microg/ml; and for Bacteroides vulgatus, 4 microg/ml. Telithromycin MICs for the B. fragilis group were usually 1 to 2 dilutions higher than ABT-773 MICs. For all strains, ABT-773 was more active than erythromycin by 4 or more dilutions, and for some strains this drug was more active than clindamycin. PMID- 11120994 TI - Identification of an erm(A) erythromycin resistance methylase gene in Streptococcus pneumoniae isolated in Greece. AB - In a serotype 11A clone of erythromycin-resistant pneumococci isolated from young Greek carriers, we identified the nucleotide sequence of erm(A), a methylase gene previously described as erm(TR) in Streptococcus pyogenes. The erm(A) pneumococci were resistant to 14- and 15-member macrolides, inducibly resistant to clindamycin, and susceptible to streptogramin B. To our knowledge, this is the first identification of resistance to erythromycin in S. pneumoniae attributed solely to the carriage of the erm(A) gene. PMID- 11120996 TI - Clinical isolate of vancomycin-heterointermediate Staphylococcus aureus susceptible to methicillin and in vitro selection of a vancomycin-resistant derivative. AB - A Staphylococcus aureus strain with low-level heteroresistance to vancomycin (designated MER) but susceptible to methicillin was isolated from an outpatient with conjunctivitis who did not receive any glycopeptide antibiotics. Incubation of the parent strain, MER, with increasing concentrations of vancomycin led to rapid selection of a stable progeny homogeneously resistant to vancomycin. Electron micrographs of strain MER showed enhanced cell wall thickness and abnormal septations typically seen with methicillin-resistant S. aureus having intermediate susceptibility to vancomycin. PMID- 11120997 TI - In vitro pharmacodynamic studies of activities of ketolides HMR 3647 (Telithromycin) and HMR 3004 against extracellular or intracellular Helicobacter pylori. AB - The pharmacodynamic properties of the ketolides HMR 3647 (telithromycin) and HMR 3004 were studied against Helicobacter pylori. Both ketolides showed a pronounced concentration-dependent killing, a significant postantibiotic effect, a long postantibiotic sub-MIC effect, and a reduction of intracellular H. pylori. PMID- 11120998 TI - Use of a recombinant strain of Mycobacterium avium expressing beta-galactosidase to evaluate the activities of antimycobacterial agents inside macrophages. AB - A reliable and low-cost method that enables rapid screening of the activity exerted by new antimicrobial agents on intracellularly growing Mycobacterium avium has been developed. To this aim, a recombinant (lacZ) strain of M. avium expressing the Escherichia coli beta-galactosidase gene was used to evaluate, in murine macrophages, the susceptibility of M. avium to common antimycobacterial agents. beta-Galactosidase levels, measured in the presence of each of the antibiotics tested, were closely correlated with the number of CFU recovered from the M. avium lacZ strain-infected macrophages. PMID- 11120999 TI - In vitro antibiotic susceptibility of Neisseria gonorrhoeae in Jakarta, Indonesia. AB - Antibiotic susceptibilities were determined for 122 Neisseria gonorrheae isolates obtained from 400 sex workers in Jakarta, Indonesia, and susceptibilities to ciprofloxacin, cefuroxime, cefoxitin, cefotaxime, ceftriaxone, chloramphenicol, and spectinomycin were found. All isolates were resistant to tetracycline. A number of the isolates demonstrated decreased susceptibilities to erythromycin (MIC >/= 1.0 microg/ml), thiamphenicol (MIC >/= 1.0 microg/ml), kanamycin (MIC >/= 16.0 microg/ml), penicillin (MIC >/= 2.0 microg/ml), gentamicin (MIC >/= 16.0 microg/ml), and norfloxacin (MIC = 0.5 microg/ml). These data showed that certain antibiotics previously used in the treatment of gonorrhea are no longer effective. PMID- 11121000 TI - Therapeutic efficacy of Poly(DL-lactide-Co-Glycolide)-encapsulated antitubercular drugs against Mycobacterium tuberculosis infection induced in mice. AB - Poly(DL-lactide-co-glycolide) (PLG) microparticles were developed as carriers for isoniazid and rifampin in order to improve compliance of tuberculous chemotherapy. Antitubercular drugs encapsulated in PLG polymers and injected in a single dose subcutaneously resulted in a sustained release (up to 6 weeks) of drugs in various organs of mice. Further, Mycobacterium tuberculosis H(37)Rv infected animals given a single shot of chemotherapy in PLG microparticles exhibited a better or equivalent clearance of CFU in various organs compared to those given a daily administration of free drugs. PMID- 11121001 TI - Predictors of parent-rated credibility in a clinical psychotherapy trial for adolescent depression. AB - The authors have reported that adolescents with major depressive disorder had a higher remission rate with cognitive-behavioral therapy (CBT) than with systemic behavioral family therapy (SBFT) or nondirective supportive therapy (NST). Parent rated treatment credibility deteriorated from baseline to end of treatment if patients were treated with SBFT or NST, compared with CBT. The present study evaluated the following variables as predictors of change in parent- rated credibility over time across the three treatment cells: severity of child's and parents' depression at baseline; parent-rated family climate at baseline; clinician age, gender, and years of clinical experience; and change in severity of child's depression and in family climate. The greater the baseline depression of children treated with CBT and NST, but not SBFT, the more favorable the change in parent-rated credibility at the end of treatment. Findings suggest that any improvement (for CBT) or a supportive therapeutic contact (for NST) may appeal to parents of severely depressed children. PMID- 11121003 TI - Unwitting exposure of the therapist: transferential and countertransferential dilemmas. AB - The unwitting exposure of the therapist's private life creates an unexpected rupture in the frame and puts both the therapist and the patient off balance. The exposure introduces into the therapy a moment of human sharing of vulnerability that has the potential to enrich the treatment. Clinical vignettes are presented to help therapists anticipate possible exposures and their consequences. The discussion encourages therapists to predict their reactions, obtain needed professional support, and make the most of opportunities for full exploration of patients' material. PMID- 11121002 TI - Change in compensatory skills in cognitive therapy for depression. AB - The Ways of Responding (WOR) was developed to assess change in compensatory or metacognitive skills taught by cognitive therapists. Thus, one would expect WOR scores to change during cognitive therapy (CT) and to be associated with change in depression level. Twenty-seven patients with a DSM-III-R diagnosis of major depression who had received CT filled out the WOR and other measures of cognition. After 12 weeks of CT, the patients exhibited change in the WOR, the Attributional Style Questionnaire, the Dysfunctional Attitude Scale, and the Self Control Scale. Furthermore, there were indications that change in depression was associated with changes in these measures of cognition, including the WOR. The WOR appears to be a sensitive measure of change during CT that covaries with change in depression. It remains to be tested whether change on the WOR is specific to CT. PMID- 11121004 TI - Excellent supervision: the residents' perspective. AB - Former residents rated their videotaped psychotherapy supervision sessions on how helpful their supervisors were as teachers during the session. Residents' and experts' ratings of the same videotape were compared and found to have no significant correlation. However, male residents were less critical than either female residents or experts. Former residents were also interviewed. Supervisors were rated as excellent when they were accepting and also when they provided guidance about highly charged clinical dilemmas. Discussion of the impact of the residents' personal experiences on the clinical encounter was also rated high and is best understood from an adult developmental perspective. The findings reveal the lasting value of sympathetic supervisors acknowledging personal concerns and are likely mirrored in all clinical settings. PMID- 11121005 TI - Erotized transference in the male patient-female therapist dyad. AB - Little has been published regarding male patients' erotic transferences to female therapists. It has been suggested that male patients do not develop full erotic transferences and rarely experience erotized transferences. The author presents a case report documenting erotization in a male patient-female therapist dyad and reviews current theories on the etiology, therapeutic significance, and treatment strategies indicated for such a transference. PMID- 11121006 TI - Conducting psychotherapy with psychotherapists II: clinical practices and collegial advice. AB - This study examined psychotherapists' experiences in conducting treatment with fellow mental health professionals. 349 psychologists (35% response) rated the extent to which their therapeutic approach with psychotherapists differed from their approach with laypersons of comparable intelligence, socioeconomic status, and diagnosis. Respondents also provided recommendations for conducting effective treatment with this elite clientele. Psychologists indicated that their practices with fellow psychotherapists were in most respects similar to those used with laypersons; 55 of the 78 items were rated of equivalent frequency. Practitioners' self-characterization as "a therapists' therapist" was related to the manner in which they treated mental health professionals. Broadly speaking, two types of advice were offered: to cultivate a warm and collaborative therapeutic relationship and to maintain proper boundaries. Recommendations for clinical work and future research on psychotherapists' psychotherapy are advanced. PMID- 11121007 TI - Interpersonal psychotherapy group (IPT-G) for depression. AB - A case study of a time-limited interpersonal psychotherapy group (IPT-G) is presented to illustrate the use of interpersonal therapy (IPT) to treat patients with major depression in a group psychotherapy format. The use of individual outcome measures as a helpful adjunct to clinical psychotherapeutic practice is demonstrated. Because IPT-G has only a few exclusion criteria (active suicidality and significant borderline personality features), it can be used in a broad range of clinical settings. This clinical example demonstrates IPT-G to be a useful modality for addressing a common and difficult patient population. PMID- 11121008 TI - Psychotherapy by psychiatrists in a managed care environment: must it be an oxymoron? A forum from the APA commission on Psychotherapy by Psychiatrists. American Psychiatric Association. PMID- 11121009 TI - Development and evolution occlude: evolution of development in mammalian teeth. PMID- 11121010 TI - c-Jun N-terminal kinase (JNK) repression during the inflammatory response? Just say NO. PMID- 11121011 TI - Lipids as a common interest of microorganisms and geochemists. PMID- 11121012 TI - Imprinted expression of small nucleolar RNAs in brain: time for RNomics. PMID- 11121013 TI - Raccoon rabies in space and time. PMID- 11121014 TI - Global regulation of gene expression. PMID- 11121015 TI - DNA condensation in two dimensions. AB - We have found that divalent electrolyte counterions common in biological cells (Ca(2+), Mg(2+), and Mn(2+) ) can condense anionic DNA molecules confined to two dimensional cationic surfaces. DNA-condensing agents in vivo include cationic histones and polyamines spermidine and spermine with sufficiently high valence (Z) 3 or larger. In vitro studies show that electrostatic forces between DNA chains in bulk aqueous solution containing divalent counterions remain purely repulsive, and DNA condensation requires counterion valence Z >/= 3. In striking contrast to bulk behavior, synchrotron x-ray diffraction and optical absorption experiments show that above a critical divalent counterion concentration the electrostatic forces between DNA chains adsorbed on surfaces of cationic membranes reverse from repulsive to attractive and lead to a chain collapse transition into a condensed phase of DNA tethered by divalent counterions. This demonstrates the importance of spatial dimensionality to intermolecular interactions where nonspecific counterion-induced electrostatic attractions between the like-charged polyelectrolytes overwhelm the electrostatic repulsions on a surface for Z = 2. This new phase, with a one-dimensional counterion liquid trapped between DNA chains at a density of 0.63 counterions per DNA bp, represents the most compact state of DNA on a surface in vitro and suggests applications in high-density storage of genetic information and organo-metallic materials processing. PMID- 11121016 TI - Constant mean curvature surfaces with three ends. AB - We announce the classification of complete almost embedded surfaces of constant mean curvature, with three ends and genus zero. They are classified by triples of points on the sphere whose distances are the asymptotic necksizes of the three ends. PMID- 11121017 TI - Identifying mRNA subsets in messenger ribonucleoprotein complexes by using cDNA arrays. AB - Genomic array technologies provide a means for profiling global changes in gene expression under a variety of conditions. However, it has been difficult to assess whether transcriptional or posttranscriptional regulation is responsible for these changes. Additionally, fluctuations in gene expression in a single cell type within a complex tissue like a tumor may be masked by overlapping profiles of all cell types in the population. In this paper, we describe the use of cDNA arrays to identify subsets of mRNAs contained in endogenous messenger ribonucleoprotein complexes (mRNPs) that are cell type specific. We identified mRNA subsets from P19 embryonal carcinoma stem cells by using mRNA-binding proteins HuB, eIF-4E, and PABP that are known to play a role in translation. The mRNA profiles associated with each of these mRNPs were unique and represented gene clusters that differed from total cellular RNA. Additionally, the composition of mRNAs detected in HuB-mRNP complexes changed dramatically after induction of neuronal differentiation with retinoic acid. We suggest that the association of structurally related mRNAs into mRNP complexes is dynamic and may help regulate posttranscriptional events such as mRNA turnover and translation. Recovering proteins specifically associated with mRNP complexes to identify and profile endogenously clustered mRNAs should provide insight into structural and functional relationships among gene transcripts and/or their protein products. We have termed this approach to functional genomics ribonomics and suggest that it will provide a useful paradigm for organizing genomic information in a biologically relevant manner. PMID- 11121018 TI - Cracks in the beta-can: fluorescent proteins from Anemonia sulcata (Anthozoa, Actinaria). AB - We characterize two green fluorescent proteins (GFPs), an orange fluorescent protein, and a nonfluorescent red protein isolated from the sea anemone Anemonia sulcata. The orange fluorescent protein and the red protein seem to represent two different states of the same protein. Furthermore, we describe the cloning of a GFP and a nonfluorescent red protein. Both proteins are homologous to the GFP from Aequorea victoria. The red protein is significantly smaller than other GFP homologues, and the formation of a closed GFP-like beta-can is not possible. Nevertheless, the primary structure of the red protein carries all features necessary for orange fluorescence. We discuss a type of beta-can that could be formed in a multimerization process. PMID- 11121019 TI - Stepwise assembly of initiation proteins at budding yeast replication origins in vitro. AB - The initiation of DNA replication in the budding yeast Saccharomyces cerevisiae occurs in two sequential and mutually exclusive steps. Prereplicative complexes (pre-RCs) containing origin recognition complex (ORC), Cdc6p, and the MCM2-7 proteins assemble only under conditions of low cyclin-dependent kinase (Cdk) activity during G(1), whereas origin activation is driven by the increase in Cdk activity at the end of G(1). As a first step toward the reconstitution of this two-step process in vitro, we describe a system in which extracts prepared from G(1)-arrested cells promote sequential assembly of ORC, Cdc6p, and MCM2-7 proteins onto exogenously added origin-containing DNA. This reaction requires an intact ARS consensus sequence and requires ATP for two distinct steps. Extracts from cells arrested in mitosis also can support the binding of ORC but are unable to load either Cdc6p or MCM2-7 proteins. This system should be useful for studying the mechanism and regulation of pre-RC assembly. PMID- 11121020 TI - ATP-dependent structural change of the eukaryotic clamp-loader protein, replication factor C. AB - The eukaryotic DNA sliding clamp that keeps DNA polymerase engaged at a replication fork, called proliferating cell nuclear antigen (PCNA), is loaded onto the 3' ends of primer DNA through its interaction with a heteropentameric protein complex called replication factor C (RFC). The ATPase activity of RFC is necessary for formation of a functional PCNA clamp. In the present study, the sensitivity of RFC to partial proteolysis is used to show that addition of ATP, ATPgammaS, or ADP induces different structural changes in RFC. Direct observation by electron microscopy reveals that RFC has a closed two-finger structure called the U form in the absence of ATP. This is converted into a more open C form on addition of ATP. In contrast, the structural changes induced by ATPgammaS or ADP are limited. These results suggest that RFC adapts on opened configuration intermediately after ATP hydrolysis. We further observe that PCNA is held between the two fingers of RFC and propose that the RFC structure change we observe during ATP hydrolysis causes the attached PCNA to form its active ring-like clamp on DNA. PMID- 11121022 TI - Covalent modification of the androgen receptor by small ubiquitin-like modifier 1 (SUMO-1). AB - Modification by SUMO-1 is proposed to play a role in protein targeting and/or stability. The SUMO-1-conjugating enzyme Ubc9 interacts with androgen receptor (AR), a ligand-activated transcription factor belonging to the steroid receptor superfamily. We show here that AR is covalently modified by SUMO-1 (sumoylated) in an androgen-enhanced fashion and identify the principal acceptor site in the N terminal domain of AR. Substitutions of sumoylated Lys residues enhanced transcriptional activity of AR without influencing its transrepressing activity. Interestingly, the same Lys residues form the cores of the recently described transcriptional synergy control motifs in AR [Iniguez-Lluhi, J. A. & Pearce, D. (2000) Mol. Cell. Biol. 20, 6040-6050]. These motifs, which match perfectly with the sumoylation consensus sequence, are also present in the N-terminal domains of glucocorticoid, mineralocorticoid, and progesterone receptor. Taken together, our data suggest that reversible sumoylation is a mechanism for regulation of steroid receptor function. PMID- 11121021 TI - tau kinases in the rat heat shock model: possible implications for Alzheimer disease. AB - We have previously shown, by using the phosphate-dependent anti-tau antibodies Tau-1 and PHF-1, that heat shock induces rapid dephosphorylation of tau followed by hyperphosphorylation in female rats. In this study, we analyzed in forebrain homogenates from female Sprague-Dawley rats the activities of extracellular signal regulated kinase 1/2 (ERK1/2), c-Jun NH(2)-terminal kinase (JNK), glycogen synthase kinase-3beta (GSK-3beta), cyclin-dependent kinase 5 (Cdk5), cAMP dependent protein kinase A (PKA), and Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) at 0 (n = 5), 3 (n = 4), 6 (n = 5), and 12 (n = 5) h after heat shock and in non-heat-shocked controls (n = 5). Immunoprecipitation kinase assays at 0 h showed suppression of the activities of all kinases except of GSK-3beta, which showed increased activity. At 3-6 h, the activities of ERK1/2, JNK, Cdk5, and GSK 3beta toward selective substrates were increased; however, only JNK, Cdk5, and GSK-3beta but not ERK1/2 were overactivated toward purified bovine tau. At 3-6 h, kinase assays specific for PKA and CaMKII showed no increased activity toward either tau or selective substrates. All of eight anti-tau antibodies tested showed dephosphorylation at 0 h and hyperphosphorylation at 3-6 h, except for 12E8, which showed hyperphosphorylation also at 0 h. Immunoblot analysis using activity-dependent antibodies against ERK1/2, JNK, and GSK-3beta confirmed the above data. Increased activation and inhibition of kinases after heat shock were statistically significant in comparison with controls. Because tau is hyperphosphorylated in Alzheimer disease these findings suggest that JNK, GSK 3beta, and Cdk5 may play a role in its pathogenesis. PMID- 11121023 TI - A model of fibrin formation based on crystal structures of fibrinogen and fibrin fragments complexed with synthetic peptides. AB - A blood clot is a meshwork of fibrin fibers built up by the systematic assembly of fibrinogen molecules proteolyzed by thrombin. Here, we describe a model of how the assembly process occurs. Five kinds of interaction are explicitly defined, including two different knob-hole interactions, an end-to-end association between gamma-chains, a lateral association between gamma-chains, and a hypothetical lateral interaction between beta-chains. The last two of these interactions are responsible for protofibril association and are predicated on intermolecular packing arrangements observed in crystal structures of fibrin double-D fragments cocrystallized with synthetic peptides corresponding to the knobs exposed by the release of the fibrinopeptides A and B. PMID- 11121024 TI - An unexpectedly high excision capacity for mispaired 5-hydroxymethyluracil in human cell extracts. AB - The oxidation of thymine in DNA can generate a base pair between 5 hydroxymethyluracil (HmU) and adenine, whereas the oxidation and deamination of 5 methylcytosine (5mC) in DNA can generate a base pair between HmU and guanine. Using synthetic oligonucleotides containing HmU at a defined site, HmU-DNA glycosylase activities in HeLa cell and human fibroblast cell extracts have been observed. An HmU-DNA glycosylase activity that removes HmU mispaired with guanine has been measured. Surprisingly, the HmU:G excision activity is 60 times greater than the corresponding HmU:A activity, even though the expected rate of formation of the HmU:A base pair exceeds that of the HmU:G base pair by a factor of 10(7). The HmU:G mispair would arise from the 5mC:G base pair, and, if unrepaired, would give rise to a transition mutation. The observation of an unexpectedly high HmU:G glycosylase activity suggests that human cells may encounter the HmU:G mispair much more frequently than expected. The conversion of 5mC to HmU must be considered as a potential pathway for the generation of 5mC to T transition mutations, which are often found in human tumors. PMID- 11121025 TI - Catalysis of DNA cleavage and nucleoside triphosphate synthesis by NM23-H2/NDP kinase share an active site that implies a DNA repair function. AB - NM23/NDP kinases play an important role in development and cancer but their biological function is unknown, despite an intriguing collection of biochemical properties including nucleoside-diphosphate kinase (NDP kinase), DNA binding and transcription, a mutator function, and cleavage of unusually structured DNA by means of a covalent enzyme-DNA complex. To assess the role of the nuclease in human NM23-H2, we sought to identify the amino acid responsible for covalent catalysis. By sequencing a DNA-linked peptide and by site-directed mutagenesis, we identified lysine-12, a phylogenetically conserved residue, as the amino acid forming the covalent complex with DNA. In particular, the epsilon-amino group acts as the critical nucleophile, because substitution with glutamine but not arginine completely abrogated covalent adduct formation and DNA cleavage, whereas the DNA-binding properties remained intact. These findings and chemical modification data suggest that phosphodiester-bond cleavage occurs by a DNA glycosylase/lyase-like mechanism known as the signature of base excision DNA repair nucleases. Involvement of NM23/NDP kinase in a DNA repair pathway would be consistent with its role in normal and tumor cell development. Additionally, lysine-12, which is known in the x-ray crystallographic structure to lie in the catalytic pocket involved in the NDP kinase phosphorylation reaction, was found essential also for the NDP kinase activity of NM23-H2, suggesting that the two catalytic activities of NM23-H2 are fundamentally connected. PMID- 11121026 TI - A stable complex of a novel transcription factor IIB- related factor, human TFIIIB50, and associated proteins mediate selective transcription by RNA polymerase III of genes with upstream promoter elements. AB - Transcription factor IIIB (TFIIIB) is directly involved in transcription initiation by RNA polymerase III in eukaryotes. Yeast contain a single TFIIIB activity that is comprised of the TATA-binding protein (TBP), TFIIB-related factor 1 (BRF1), and TFIIIB", whereas two distinct TFIIIB activities, TFIIIB alpha and TFIIIB-beta, have been described in human cells. Human TFIIIB-beta is required for transcription of genes with internal promoter elements, and contains TBP, a TFIIIB" homologue (TFIIIB150), and a BRF1 homologue (TFIIIB90), whereas TFIIIB-alpha is required for transcription of genes with promoter elements upstream of the initiation site. Here we describe the identification, cloning, and characterization of TFIIIB50, a novel homologue of TFIIB and TFIIIB90. TFIIIB50 and tightly associated factors, along with TBP and TFIIIB150, reconstitute human TFIIIB-alpha activity. Thus, higher eukaryotes, in contrast to the yeast Saccharomyces cerevisiae, have evolved two distinct TFIIB-related factors that mediate promoter selectivity by RNA polymerase III. PMID- 11121027 TI - Purification and kinetic analysis of recombinant CA XII, a membrane carbonic anhydrase overexpressed in certain cancers. AB - Carbonic anhydrase XII (CA XII) is a transmembrane glycoprotein with an active extracellular CA domain that is overexpressed on cell surfaces of certain cancers. Its expression has been linked to tumor invasiveness. To characterize its catalytic properties, we purified recombinant secretory forms of wild-type and mutant CA XIIs. The catalytic properties of these enzymes in the hydration of CO(2) were measured at steady state by stopped-flow spectrophotometry and at chemical equilibrium by the exchange of (18)O between CO(2) and water determined by mass spectrometry. The catalysis of CO(2) hydration by soluble CA XII has a maximal value of k(cat)/K(m) at 34 microM(-1) small middle dots(-1), which is similar to those of the membrane-associated CA IV and to soluble CA I. The pH profiles of this catalysis and the catalyzed hydrolysis of 4-nitrophenylacetate indicate that the pK(a) of the zinc-bound water in CA XII is 7.1. His64 in CA XII acts as a proton shuttle residue, as evidenced by the reduced rate constant for proton transfer in the mutants containing the replacements His64 --> Ala and His64 --> Arg, as well as by the selective inhibition of the proton transfer step by cupric ions in wild-type CA XII. The catalytic rate of CO(2) hydration by the soluble form of CA XII is identical with that of the membrane-bound enzyme. These observations suggest a role for CA XII in CO(2)/HCO(3)(-) homeostasis in cells in which it is normally expressed. They are also compatible with a role for CA XII in acidifying the microenvironment of cancer cells in which CA XII is overexpressed, providing a mechanism for CA XII to augment tumor invasiveness and suggesting CA XII as a potential target for chemotherapeutic agents. PMID- 11121028 TI - Context-dependent anticodon recognition by class I lysyl-tRNA synthetases. AB - Lysyl-tRNA synthesis is catalyzed by two unrelated families of aminoacyl-tRNA synthetases. In most bacteria and all eukarya, the known lysyl-tRNA synthetases (LysRSs) are subclass IIb-type aminoacyl-tRNA synthetases, whereas many archaea and a scattering of bacteria contain an unrelated class I-type LysRS. Examination of the recognition of partially modified tRNA(Lys) anticodon variants by a bacterial (from Borrelia burgdorferi) and an archaeal (from Methanococcus maripaludis) class I lysyl-tRNA synthetase revealed differences in the pattern of anticodon recognition between the two enzymes. U35 and U36 were both important for recognition by the B. burgdorferi enzyme, whereas only U36 played a role in recognition by M. maripaludis LysRS. Examination of the phylogenetic distribution of class I LysRSs suggested a correlation between recognition of U35 and U36 and the presence of asparaginyl-tRNA synthetase (AsnRS), which also recognizes U35 and U36 in the anticodon of tRNA(Asn). However, the class II LysRS of Helicobacter pylori, an organism that lacks AsnRS, also recognizes both U35 and U36, indicating that the presence of AsnRS has solely influenced the phylogenetic distribution of class I LysRSs. These data suggest that competition between unrelated aminoacyl-tRNA synthetases for overlapping anticodon sequences is a determinant of the phylogenetic distribution of extant synthetase families. Such patterns of competition also provide a basis for the two separate horizontal gene transfer events hypothesized in the evolution of the class I lysyl-tRNA synthetases. PMID- 11121029 TI - Deciphering the folding kinetics of transmembrane helical proteins. AB - Nearly a quarter of genomic sequences and almost half of all receptors that are likely to be targets for drug design are integral membrane proteins. Understanding the detailed mechanisms of the folding of membrane proteins is a largely unsolved, key problem in structural biology. Here, we introduce a general model and use computer simulations to study the equilibrium properties and the folding kinetics of a C(alpha)-based two-helix bundle fragment (comprised of 66 aa) of bacteriorhodopsin. Various intermediates are identified and their free energy are calculated together with the free energy barrier between them. In 40% of folding trajectories, the folding rate is considerably increased by the presence of nonobligatory intermediates acting as traps. In all cases, a substantial portion of the helices is rapidly formed. This initial stage is followed by a long period of consolidation of the helices accompanied by their correct packing within the membrane. Our results provide the framework for understanding the variety of folding pathways of helical transmembrane proteins. PMID- 11121030 TI - Fluorescence quenching: A tool for single-molecule protein-folding study. AB - By using titin as a model system, we have demonstrated that fluorescence quenching can be used to study protein folding at the single molecule level. The unfolded titin molecules with multiple dye molecules attached are able to fold to the native state. In the native folded state, the fluorescence from dye molecules is quenched due to the close proximity between the dye molecules. Unfolding of the titin leads to a dramatic increase in the fluorescence intensity. Such a change makes the folded and unfolded states of a single titin molecule clearly distinguishable and allows us to measure the folding dynamics of individual titin molecules in real time. We have also shown that fluorescence quenching can signal folding and unfolding of a small protein with only one immunoglobulin domain. PMID- 11121031 TI - Archaeal adaptation to higher temperatures revealed by genomic sequence of Thermoplasma volcanium. AB - The complete genomic sequence of the archaeon Thermoplasma volcanium, possessing optimum growth temperature (OGT) of 60 degrees C, is reported. By systematically comparing this genomic sequence with the other known genomic sequences of archaea, all possessing higher OGT, a number of strong correlations have been identified between characteristics of genomic organization and the OGT. With increasing OGT, in the genomic DNA, frequency of clustering purines and pyrimidines into separate dinucleotides rises (e.g., by often forming AA and TT, whereas avoiding TA and AT). Proteins coded in a genome are divided into two distinct subpopulations possessing isoelectric points in different ranges (i.e., acidic and basic), and with increasing OGT the size of the basic subpopulation becomes larger. At the metabolic level, genes coding for enzymes mediating pathways for synthesizing some coenzymes, such as heme, start missing. These findings provide insights into the design of individual genomic components, as well as principles for coordinating changes in these designs for the adaptation to new environments. PMID- 11121032 TI - Ultrasensitive magnetic biosensor for homogeneous immunoassay. AB - A technique is described for specific, sensitive, quantitative, and rapid detection of biological targets by using superparamagnetic nanoparticles and a "microscope" based on a high-transition temperature dc superconducting quantum interference device (SQUID). In this technique, a mylar film to which the targets have been bound is placed on the microscope. The film, at room temperature and atmospheric pressure, is typically 40 micrometer from the SQUID, which is at 77 K in a vacuum. A suspension of magnetic nanoparticles carrying antibodies directed against the target is added to the mixture in the well, and 1-s pulses of magnetic field are applied parallel to the SQUID. In the presence of this aligning field the nanoparticles develop a net magnetization, which relaxes when the field is turned off. Unbound nanoparticles relax rapidly by Brownian rotation and contribute no measurable signal. Nanoparticles that are bound to the target on the film are immobilized and undergo Neel relaxation, producing a slowly decaying magnetic flux, which is detected by the SQUID. The ability to distinguish between bound and unbound labels allows one to run homogeneous assays, which do not require separation and removal of unbound magnetic particles. The technique has been demonstrated with a model system of liposomes carrying the FLAG epitope. The SQUID microscope requires no more than (5 +/- 2) x 10(4) magnetic nanoparticles to register a reproducible signal. PMID- 11121033 TI - What is the role of non-native intermediates of beta-lactoglobulin in protein folding? AB - The mechanism of alpha-->beta transition in folding of beta-lactoglobulin is discussed based on free energy landscape analysis of a long lattice model. It is found that helical propensity of beta-lactoglobulin is driven by conformational entropy and is intrinsically coded in its native structure. We propose a view on a role of folding intermediate, which is "on-pathway" but rich in non-native structures. The present results suggest that the native structure topology plays an important role in alpha-->beta transition. PMID- 11121034 TI - The proton to electron stoichiometry of steady-state photosynthesis in living plants: A proton-pumping Q cycle is continuously engaged. AB - A noninvasive technique is introduced with which relative proton to electron stoichiometries (H(+)/e(-) ratios) for photosynthetic electron transfer can be obtained from leaves of living plants under steady-state illumination. Both electron and proton transfer fluxes were estimated by a modification of our previously reported dark-interval relaxation kinetics (DIRK) analysis, in which processes that occur upon rapid shuttering of the actinic light are analyzed. Rates of turnover of linear electron transfer through the cytochrome (cyt) b(6)f complex were estimated by measuring the DIRK signals associated with reduction of cyt f and P(700). The rates of proton pumping through the electron transfer chain and the CF(O)-CF(1) ATP synthase (ATPase) were estimated by measuring the DIRK signals associated with the electrochromic shifting of pigments in the light harvesting complexes. Electron transfer fluxes were also estimated by analysis of saturation pulse-induced changes in chlorophyll a fluorescence yield. It was shown that the H(+)/e(-) ratio, with respect to both cyt b(6)f complex and photosystem (PS) II turnover, was constant under low to saturating illumination in intact tobacco leaves. Because a H(+)/e(-) ratio of 3 at a low light is generally accepted, we infer that this ratio is maintained under conditions of normal (unstressed) photosynthesis, implying a continuously engaged, proton pumping Q cycle at the cyt b(6)f complex. PMID- 11121035 TI - Explaining the high mutation rates of cancer cells to drug and multidrug resistance by chromosome reassortments that are catalyzed by aneuploidy. AB - The mutation rates of cancer cells to drug and multidrug resistance are paradoxically high, i.e., 10(-3) to 10(-6), compared with those altering phenotypes of recessive genes in normal diploid cells of about 10(-12). Here the hypothesis was investigated that these mutations are due to chromosome reassortments that are catalyzed by aneuploidy. Aneuploidy, an abnormal number of chromosomes, is the most common genetic abnormality of cancer cells and is known to change phenotypes (e.g., Down's syndrome). Moreover, we have shown recently that aneuploidy autocatalyzes reassortments of up to 2% per chromosome per mitosis because it unbalances spindle proteins, even centrosome numbers, via gene dosage. The hypothesis predicts that a selected phenotype is associated with multiple unselected ones, because chromosome reassortments unbalance simultaneously thousands of regulatory and structural genes. It also predicts variants of a selected phenotype based on variant reassortments. To test our hypothesis we have investigated in parallel the mutation rates of highly aneuploid and of normal diploid Chinese hamster cells to resistance against puromycin, cytosine arabinoside, colcemid, and methotrexate. The mutation rates of aneuploid cells ranged from 10(-4) to 10(-6), but no drug-resistant mutants were obtained from diploid cells in our conditions. Further selection increased drug resistance at similar mutation rates. Mutants selected from cloned cells for resistance against one drug displayed different unselected phenotypes, e.g., polygonal or fusiform cellular morphology, flat or three-dimensional colonies, and resistances against other unrelated drugs. Thus our hypothesis offers a unifying explanation for the high mutation rates of aneuploid cancer cells and for the association of selected with unselected phenotypes, e.g., multidrug resistance. It also predicts drug-specific chromosome combinations that could become a basis for selecting alternative chemotherapy against drug-resistant cancer. PMID- 11121036 TI - c-Mos forces the mitotic cell cycle to undergo meiosis II to produce haploid gametes. AB - The meiotic cycle reduces ploidy through two consecutive M phases, meiosis I and meiosis II, without an intervening S phase. To maintain ploidy through successive generations, meiosis must be followed by mitosis after the recovery of diploidy by fertilization. However, the coordination from meiotic to mitotic cycle is still unclear. Mos, the c-mos protooncogene product, is a key regulator of meiosis in vertebrates. In contrast to the previous observation that Mos functions only in vertebrate oocytes that arrest at meiotic metaphase II, here we isolate the first invertebrate mos from starfish and show that Mos functions also in starfish oocytes that arrest after the completion of meiosis II but not at metaphase II. In the absence of Mos, meiosis I is followed directly by repeated embryonic mitotic cycles, and its reinstatement restores meiosis II and subsequent cell cycle arrest. These observations imply that after meiosis I, oocytes have a competence to progress through the embryonic mitotic cycle, but that Mos diverts the cell cycle to execute meiosis II and remains to restrain the return to the mitotic cycle. We propose that a role of Mos that is conserved in invertebrate and vertebrate oocytes is not to support metaphase II arrest but to prevent the meiotic/mitotic conversion after meiosis I until fertilization, directing meiosis II to ensure the reduction of ploidy. PMID- 11121037 TI - Functional interaction between c-Abl and the p21-activated protein kinase gamma PAK. AB - A member of the p21-activated protein kinase (PAK) family, gamma-PAK has cytostatic properties and is activated by cellular stresses such as hyperosmolarity or DNA damage. We report herein that gamma-PAK is associated in vivo with the nonreceptor protein tyrosine kinase c-Abl. gamma-PAK phosphorylates c-Abl on sites located in the kinase domain, in a region that is implicated in protein-protein interactions and in subcellular localization. Activation of gamma PAK in human embryonic kidney 293T cells by cotransfection with constitutively active Cdc42 induces activation of c-Abl, resulting in increased phosphotyrosine levels. Cotransfection of c-Abl and gamma-PAK elicits phosphorylation of gamma PAK on tyrosine and down-regulation of gamma-PAK activity, promoting accumulation of inactive gamma-PAK. gamma-PAK is also phosphorylated in vitro by c-Abl. gamma PAK activity is regulated by ubiquitination and proteolysis in vivo, as shown by immunoblotting with an anti-ubiquitin antibody in the presence of proteasome inhibitors. In summary, we describe a functional interaction between gamma-PAK and c-Abl in which gamma-PAK stimulates c-Abl tyrosine kinase activity and c-Abl phosphorylates and down-regulates gamma-PAK, suggesting the existence of a negative feedback loop between c-Abl and gamma-PAK. PMID- 11121038 TI - The TACC domain identifies a family of centrosomal proteins that can interact with microtubules. AB - We recently showed that the Drosophila transforming acidic coiled-coil (D-TACC) protein is located in the centrosome, interacts with microtubules, and is required for mitosis in the Drosophila embryo. There are three known human TACC proteins that share a conserved, C-terminal, coiled-coil region with D-TACC. These proteins have all been implicated in cancer, but their normal functions are unknown. We show that all three human TACC proteins are concentrated at centrosomes, but with very different characteristics: TACC1 is weakly concentrated at centrosomes during mitosis; TACC2 is strongly concentrated at centrosomes throughout the cell cycle; and TACC3 is strongly concentrated in a more diffuse region around centrosomes during mitosis. When the C-terminal TACC domain is overexpressed in HeLa cells, it forms large polymers in the cytoplasm that can interact with both microtubules and tubulin. The full-length TACC proteins form similar polymers when overexpressed, but their interaction with microtubules and tubulin is regulated during the cell cycle. At least one of the human TACC proteins appears to increase the number and/or stability of centrosomal microtubules when overexpressed during mitosis. Thus, the TACC domain identifies a family of centrosomal proteins that can interact with microtubules. This may explain the link between the TACC genes and cancer. PMID- 11121039 TI - Activator of G protein signaling 3 is a guanine dissociation inhibitor for Galpha i subunits. AB - Activator of G protein signaling 3 (AGS3) is a newly identified protein shown to act at the level of the G protein itself. AGS3 belongs to the GoLoco family of proteins, sharing the 19-aa GoLoco motif that is a Galpha(i/o) binding motif. AGS3 interacts only with members of the Galpha(i/o) subfamily. By surface plasmon resonance, we found that AGS3 binds exclusively to the GDP-bound form of Galpha(i3). In GTPgammaS binding assays, AGS3 behaves as a guanine dissociation inhibitor (GDI), inhibiting the rate of exchange of GDP for GTP by Galpha(i3). AGS3 interacts with both Galpha(i3) and Galpha(o) subunits, but has GDI activity only on Galpha(i3), not on Galpha(o). The fourth GoLoco motif of AGS3 is a major contributor to this activity. AGS3 stabilizes Galpha(i3) in its GDP-bound form, as it inhibits the increase in tryptophan fluorescence of the Galpha(i3)-GDP subunit stimulated by AlF(4)(-). AGS3 is widely expressed as it is detected by immunoblotting in brain, testis, liver, kidney, heart, pancreas, and in PC-12 cells. Several different sizes of the protein are detected. By Northern blotting, AGS3 shows 2.3-kb and 3.5-kb mRNAs in heart and brain, respectively, suggesting tissue-specific alternative splicing. Taken together, our results demonstrate that AGS3 is a GDI. To the best of our knowledge, no other GDI has been described for heterotrimeric G proteins. Inhibition of the Galpha subunit and stimulation of heterotrimeric G protein signaling, presumably by stimulating Gbetagamma, extend the possibilities for modulating signal transduction through heterotrimeric G proteins. PMID- 11121040 TI - Yin6, a fission yeast Int6 homolog, complexes with Moe1 and plays a role in chromosome segregation. AB - The INT6 gene has been implicated in human breast cancer formation, but its function is unknown. We isolated an Int6 homolog from fission yeast, Yin6, by its binding to a conserved protein in the Ras pathway, Moe1. Yin6 and Moe1 converge on the same protein complex to promote microtubule instability/disassembly. Yin6 and Moe1 interact cooperatively: when either protein is absent, the other becomes mislocalized with decreased protein levels. Furthermore, whereas full-length human Int6 rescues the phenotypes of the yin6-null (yin6Delta) mutant cells and binds human Moe1, truncated Int6 proteins found in tumors do not. Importantly, yin6Delta alone impairs chromosome segregation weakly, but yin6Delta together with ras1Delta causes severe chromosome missegregation. These data support a model in which INT6 mutations in humans either alone or together with additional mutations, such as a RAS mutation, may contribute to tumorigenesis by altering genome stability. PMID- 11121041 TI - An alternative, nonapoptotic form of programmed cell death. AB - The term apoptosis often has been used interchangeably with the term programmed cell death. Here we describe a form of programmed cell death that is distinct from apoptosis by the criteria of morphology, biochemistry, and response to apoptosis inhibitors. Morphologically, this alternative form of programmed cell death appears during development and in some cases of neurodegeneration. Despite its lack of response to caspase inhibitors and Bcl-x(L), we show that this form of cell death is driven by an alternative caspase-9 activity that is Apaf-1 independent. Characterization of this alternative form of programmed cell death should lead to new insight into cell death programs and their roles in development and degeneration. PMID- 11121042 TI - Nitric oxide negatively regulates c-Jun N-terminal kinase/stress-activated protein kinase by means of S-nitrosylation. AB - NO, produced from l-arginine in a reaction catalyzed by NO synthase, is an endogenous free radical with multiple functions in mammalian cells. Here, we demonstrate that endogenously produced NO can suppress c-Jun N-terminal kinase (JNK) activation in intact cells. Treatment of BV-2 murine microglial cells with IFN-gamma induced endogenous NO production, concomitantly suppressing JNK1 activation. Similarly, IFN-gamma induced suppression of JNK1 activation in RAW264.7 murine macrophage cells and rat alveolar macrophages. The IFN-gamma induced suppression of JNK1 activation in BV-2, RAW264.7, or rat alveolar macrophage cells was completely prevented by N(G)-nitro-l-arginine, a NO synthase inhibitor. Interestingly, the IFN-gamma-induced suppression of JNK1 activation was not affected by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, an inhibitor of guanylyl cyclase. 8-Bromo-cGMP, a membrane-permeant analogue of cGMP, did not change JNK1 activation in intact cells either. In contrast, S-nitro-N-acetyl-dl penicillamine (SNAP), a NO donor, inhibited JNK1 activity in vitro. Furthermore, a thiol reducing agent, DTT, reversed not only the in vitro inhibition of JNK1 activity by SNAP but also the in vivo suppression of JNK1 activity by IFN-gamma. Substitution of serine for cysteine-116 in JNK1 abolished the inhibitory effect of IFN-gamma or SNAP on JNK1 activity in vivo or in vitro, respectively. Moreover, IFN-gamma enhanced endogenous S-nitrosylation of JNK1 in RAW264.7 cells. Collectively, our data suggest that endogenous NO mediates the IFN-gamma induced suppression of JNK1 activation in macrophage cells by means of a thiol redox mechanism. PMID- 11121043 TI - Ski represses bone morphogenic protein signaling in Xenopus and mammalian cells. AB - The bone morphogenic proteins (BMPs) play important roles in vertebrate development. In Xenopus, BMPs act as epidermal inducers and also as negative regulators of neurogenesis. Antagonism of BMP signaling results in neuralization. BMPs signal through the cell-surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. Here, we show that the Ski oncoprotein can block BMP signaling and the expression of BMP responsive genes in both Xenopus and mammalian cells by directly interacting with and repressing the activity of BMP-specific Smad complexes. This ability to antagonize BMP signaling results in neuralization by Ski in the Xenopus embryo and blocking of osteoblast differentiation of murine W-20-17 cells. Thus, Ski is able to repress the activity of all receptor-associated Smads and may regulate vertebrate development by modulating the signaling activity of transforming growth factor-beta family members. PMID- 11121044 TI - Widespread occurrence of structurally diverse tetraether membrane lipids: evidence for the ubiquitous presence of low-temperature relatives of hyperthermophiles. AB - Isoprenoid glycerol dialkyl glycerol tetraethers (GDGTs) and branched glycerol dialkyl diethers are main membrane constituents of cultured hyperthermophilic archaea and eubacteria, respectively, and are found in environments with temperatures >60 degrees C. Recently, we developed a new technique for the analysis of intact core tetraether lipids in cell material and sediments. The application of this technique to recent sediments shows that known and newly identified isoprenoid and branched GDGTs are widespread in low-temperature environments (<20 degrees C) and are structurally far more diverse than previously thought. Their distribution indicates the ubiquitous environmental presence of as yet uncultivated, nonthermophilic organisms that may have independently evolved from hyperthermophilic archaea and eubacteria. The structures of some of the new GDGTs point to the hybridization of both typical archaeal and eubacterial biosynthetic pathways in single organisms. PMID- 11121045 TI - Evolutionary modification of development in mammalian teeth: quantifying gene expression patterns and topography. AB - The study of mammalian evolution often relies on detailed analysis of dental morphology. For molecular patterning to play a role in dental evolution, gene expression differences should be linkable to corresponding morphological differences. Because teeth, like many other structures, are complex and evolution of new shapes usually involves subtle changes, we have developed topographic methods by using Geographic Information Systems. We investigated how genetic markers for epithelial signaling centers known as enamel knots are associated with evolutionary divergence of molar teeth in two rodent species, mouse and vole. Our analysis of expression patterns of Fgf4, Lef1, p21, and Shh genes in relation to digital elevation models of developing tooth shapes shows that molecular prepatterns predict the lateral cusp topography more than a day in advance. A heterotopic shift in the molecular prepatterns can be implicated in the evolution of mouse molar, changing locations from which historically homologous cusps form. The subtle but measurable heterotopic shifts may play a large role in the evolution of tooth cusp topographies. However, evolutionary increase in the number of longitudinal cusps in vole molar has involved accelerated longitudinal growth and iterative addition of new cusps without changes in lateral cusp topography. The iterative addition of cusps after the establishment of lateral cusp topography may limit the independence of individual morphological features used in evolutionary studies. The diversity of mammalian molar patterns may largely result from the heterotopic and iterative processes. PMID- 11121046 TI - Sexual selection and speciation in field crickets. AB - Recent theoretical work has shown that sexual selection may cause speciation under a much wider range of conditions than previously supposed. There are, however, no empirical studies capable of simultaneously evaluating several key predictions that contrast this with other speciation models. We present data on male pulse rates and female phonotactic responses to pulse rates for the field cricket Gryllus texensis; pulse rate is the key feature distinguishing G. texensis from its cryptic sister species G. rubens. We show (i) genetic variation in male song and in female preference for song, (ii) a genetic correlation between the male trait and the female preference, and (iii) no character displacement in male song, female song recognition, female species-level song discrimination, or female song preference. Combined with previous work demonstrating a lack of hybrid inviability, these results suggest that divergent sexual selection may have caused speciation between these taxa. PMID- 11121047 TI - Identification of the MATa mating-type locus of Cryptococcus neoformans reveals a serotype A MATa strain thought to have been extinct. AB - Cryptococcus neoformans is an opportunistic fungal pathogen with a defined sexual cycle involving mating between haploid MATa and MATalpha cells. Here we describe the isolation of part of the MATa mating-type locus encoding a Ste20 kinase homolog, Ste20a. We show that the STE20a gene cosegregates with the MATa mating type in genetic crosses, maps within the mating-type locus on a 1.8-Mb chromosome, and is allelic with the MATalpha locus. We identify the first MATa isolate of the most common pathogenic variety of C. neoformans (serotype A, variety grubii) which had been thought to be extinct. This serotype A MATa strain is sterile, fails to produce mating pheromone, and is less virulent than a serotype A MATalpha strain in an animal model. Our studies illustrate an association of mating type with virulence and suggest that, like Candida albicans, pathogenic isolates of C. neoformans may be largely asexual. PMID- 11121048 TI - Genetic restriction of HIV-1 pathogenesis to AIDS by promoter alleles of IL10. AB - IL10 is a powerful TH-2 cell cytokine produced by lymphoid cells that limits HIV 1 replication in vivo, ostensibly by inhibiting macrophage/monocyte and T-cell lymphocyte replication and secretion of inflammatory cytokines (IL1, TNFalpha, IL6, IL8, and IL12). A genetic epidemiological scan of patients enrolled in AIDS cohorts for candidate gene-linked short tandem repeat polymorphisms revealed significant genotype associations for HIV-1 infection and progression to AIDS with markers adjacent to and tracking (by linkage disequilibrium) common single nucleotide polymorphic variants in the IL10 promoter region. Individuals carrying the IL10-5'-592A (IL10-5'A) promoter allele possibly were at increased risk for HIV-1 infection, and once infected they progressed to AIDS more rapidly than homozygotes for the alternative IL10-5'-592 C/C (IL10-+/+) genotype, particularly in the later stages of HIV-1 infection. An estimated 25-30% of long-term nonprogressors (who avoid clinical AIDS for 10 or more years after HIV-1 infection) can be attributed to their IL10-+/+ promoter genotype. Alternative IL10 promoter alleles are functionally distinct in relative IL10 production, in retention of an avian erythroblastosis virus transcription factor recognition sequence and in binding to specific putative nuclear transcription factors, suggesting a potential mechanism whereby IL10-5'A down-regulation of inhibitory IL10 facilitates HIV-1 replication in vivo, accelerating the onset of AIDS. PMID- 11121049 TI - Transposable elements in sexual and ancient asexual taxa. AB - Sexual reproduction allows deleterious transposable elements to proliferate in populations, whereas the loss of sex, by preventing their spread, has been predicted eventually to result in a population free of such elements [Hickey, D. A. (1982) Genetics 101, 519-531]. We tested this expectation by screening representatives of a majority of animal phyla for LINE-like and gypsy-like reverse transcriptases and mariner/Tc1-like transposases. All species tested positive for reverse transcriptases except rotifers of the class Bdelloidea, the largest eukaryotic taxon in which males, hermaphrodites, and meiosis are unknown and for which ancient asexuality is supported by molecular genetic evidence. Mariner-like transposases are distributed sporadically among species and are present in bdelloid rotifers. The remarkable lack of LINE-like and gypsy-like retrotransposons in bdelloids and their ubiquitous presence in other taxa support the view that eukaryotic retrotransposons are sexually transmitted nuclear parasites and that bdelloid rotifers evolved asexually. PMID- 11121050 TI - Quantitative trait loci on chromosomes 3 and 17 influence phenotypes of the metabolic syndrome. AB - Recent research has emphasized the importance of the metabolic cluster, which includes glucose intolerance, dyslipidemia, and high blood pressure, as a strong predictor of the obesity-related morbidities and premature mortality. Fundamental to this association, commonly referred to as the metabolic syndrome, is the close interaction between abdominal fat patterning, total body adiposity, and insulin resistance. As the initial step in identifying major genetic loci influencing these phenotypes, we performed a genomewide scan by using a 10-centiMorgan map in 2,209 individuals distributed over 507 nuclear Caucasian families. Pedigree-based analysis using a variance components linkage model demonstrated a quantitative trait locus (QTL) on chromosome 3 (3q27) strongly linked to six traits representing these fundamental phenotypes [logarithm of odds (lod) scores ranged from 2.4 to 3.5]. This QTL exhibited possible epistatic interaction with a second QTL on chromosome 17 (17p12) strongly linked to plasma leptin levels (lod = 5.0). Situated at these epistatic QTLs are candidate genes likely to influence two biologic precursor pathways of the metabolic syndrome. PMID- 11121051 TI - Haploinsufficiency of steroidogenic factor-1 in mice disrupts adrenal development leading to an impaired stress response. AB - Adrenal steroids are essential for homeostasis and survival during severe physiological stress. Analysis of a patient heterozygous for the steroidogenic factor-1 (SF-1) gene suggested that reduced expression of this nuclear receptor leads to adrenal failure. We therefore examined SF-1 heterozygous (+/-) mice as a potential model for delineating mechanisms underlying this disease. Here we show that SF-1 +/- mice exhibit adrenal insufficiency resulting from profound defects in adrenal development and organization. However, compensatory mechanisms, such as cellular hypertrophy and increased expression of the rate-limiting steroidogenic protein StAR, help to maintain adrenal function at near normal capacity under basal conditions. In contrast, adrenal deficits in SF-1 heterozygotes are revealed under stressful conditions, demonstrating that normal gene dosage of SF-1 is required for mounting an adequate stress response. Our findings predict that natural variations leading to reduced SF-1 function may underlie some forms of subclinical adrenal insufficiency, which become life threatening during traumatic stress. PMID- 11121052 TI - Developmentally dynamic histone acetylation pattern of a tissue-specific chromatin domain. AB - We have defined the histone acetylation pattern of the endogenous murine beta globin domain, which contains the erythroidspecific beta-globin genes. The beta globin locus control region (LCR) and transcriptionally active promoters were enriched in acetylated histones in fetal liver relative to fetal brain, whereas the inactive promoters were hypoacetylated. In contrast, the LCR and both active and inactive promoters were hyperacetylated in yolk sac. Hypersensitive site two of the LCR was also hyperacetylated in murine embryonic stem cells, whereas beta globin promoters were hypoacetylated. Thus, the acetylation pattern varied at different developmental stages. Histone deacetylase inhibition selectively increased acetylation at a hypoacetylated promoter in fetal liver, suggesting that active deacetylation contributes to silencing of promoters. We propose that dynamic histone acetylation and deacetylation play an important role in the developmental control of beta-globin gene expression. PMID- 11121053 TI - HO endonuclease-induced recombination in yeast meiosis resembles Spo11-induced events. AB - In meiosis, gene conversions are accompanied by higher levels of crossing over than in mitotic cells. To determine whether the special properties of meiotic recombination can be attributed to the way in which Spo11p creates double-strand breaks (DSBs) at special hot spots in Saccharomyces cerevisiae, we expressed the site-specific HO endonuclease in meiotic cells. We could therefore compare HO induced recombination in a well-defined region both in mitosis and meiosis, as well as compare Spo11p- and HO-induced meiotic events. HO-induced gene conversions in meiosis were accompanied by crossovers at the same high level (52%) as Spo11p-induced events. Moreover, HO-induced crossovers were reduced 3 fold by a msh4Delta mutation that similarly affects Spo11p-promoted events. In a spo11Delta diploid, where the only DSB is made by HO, crossing over was significantly higher (27%) than in mitotic cells (5 h), however, Deltapsi(m) fell, correlating with the appearance of early apoptotic features and a decrease in cell viability. Glucose deprivation or iodoacetate treatment of cells in the presence of NO resulted in a collapse of Deltapsi(m), demonstrating involvement of glycolytic ATP in its maintenance. Under these conditions cell viability also was decreased. Treatment with oligomycin and/or bongkrekic acid indicated that the maintenance of Deltapsi(m) during exposure to NO is caused by reversal of the ATP synthase and other electrogenic pumps. Thus, blockade of complex IV by NO initiates a protective action in the mitochondrion to maintain Deltapsi(m) this results in prevention of apoptosis. It is likely that during cellular stress involving increased generation of NO this compound will trigger a similar sequence of events, depending on its concentration and duration of release. PMID- 11121063 TI - Mosaic blood vessels in tumors: frequency of cancer cells in contact with flowing blood. AB - The presence of "mosaic" vessels in which both endothelial cells and tumor cells form the luminal surface has profound implications for metastasis, drug delivery, and antivascular therapy. Yet little is known of the frequency, and thus importance, of mosaic vessels in tumors. Using CD31 and CD105 to identify endothelial cells and endogenous green fluorescent protein labeling of tumor cells, we show that approximately 15% of perfused vessels of a colon carcinoma xenografted at two different sites in mice were mosaic vessels having focal regions where no CD31/CD105 immunoreactivity was detected and tumor cells appeared to contact the vessel lumen. These regions occupied approximately 25% of the perimeter of the mosaic vessels, or approximately 4% of the total vascular surface area in these colon carcinomas. In addition, we found similar numbers of mosaic vessels in human colon carcinoma biopsies. Our results are consistent with the observation that approximately 10(6) cells are shed daily per g of tumor. More importantly, our data offer a possible explanation for the antivascular effects of cytotoxic agents and suggest potential strategies for targeting the tumor vasculature. PMID- 11121064 TI - The human hematopoietic stem cell compartment is heterogeneous for CXCR4 expression. AB - The chemokine stromal derived factor-1alpha (SDF-1alpha) has been implicated recently in the chemotaxis of primitive human hematopoietic cells, suggesting that pluripotent human stem cells express the SDF-1alpha receptor, CXCR4. By using flow cytometry and confocal microscopy, we have identified and isolated primitive subsets of human CXCR4(+) and CXCR4(-) cells. Distinctions in the progenitor content and response to SDF-1alpha in vitro indicate that CXCR4(+) and CXCR4(-) cells represent discrete populations of primitive blood cells. The i.v. transplantation of these subfractions into immune-deficient mice established that both possess comparable engraftment capacity in vivo. Human myeloid, lymphoid, and primitive CD34(+) CXCR4(+) cells were present in chimeric animals transplanted with either subset, indicating that CXCR4(+) and CXCR4(-) stem cells have equivalent proliferative and differentiative abilities. Our study indicates that the human stem cell compartment is heterogeneous for CXCR4 expression, suggesting that the relationship between CXCR4 expression and stem cell repopulating function is not obligatory. PMID- 11121065 TI - Entry and survival of Salmonella typhimurium in dendritic cells and presentation of recombinant antigens do not require macrophage-specific virulence factors. AB - Macrophages have long been regarded as the main target encountered by Salmonella typhimurium, a Gram-negative facultative intracellular pathogen that invades the intestinal mucosa. S. typhimurium, however, are first internalized by dendritic cells. To gain new insights into the interactions between Salmonella and the dendritic cells, we compared the fate of wild-type S. typhimurium and the virulence-attenuated PhoP constitutive (PhoP(c)) strain. The PhoP(c) strain is impaired for entry and survival in mammalian cells and is poorly processed by macrophages for antigen presentation on MHC class II molecules. Here, we show that bone marrow-derived dendritic cells can similarly process and present a foreign antigen expressed by the invasive wild-type and the attenuated PhoP(c) S. typhimurium. This property correlates with equivalent entry and survival efficiencies of both strains in dendritic cells. In addition, Salmonella strains mutated in mgtCB, sseC, and orfL genes required for macrophage survival showed no defect in survival in dendritic cells. Together, these results indicate that uptake of Salmonella by dendritic cells and subsequent antigen processing and presentation do not depend on virulence factors important in macrophages. PMID- 11121066 TI - Active segregation by the Bacillus subtilis partitioning system in Escherichia coli. AB - Bacterial genes required for proper partitioning consist of two transacting genes that encode proteins and a cis-acting gene that functions like a centromere. Plasmids actively partitioning by means of these genes migrate from midcell to the cell quarters and are tethered to these sites until the cells divide. Previously the partitioning genes were mainly found on plasmids and phages in Escherichia coli. However, progress in genome sequencing reveals that partitioning genes are ubiquitous in many bacterial plasmids and chromosomes. Each homologue of the two transacting genes belongs to a family, ParA or ParB. Moreover, phylogenic analysis of members of the ParA and ParB families indicates that each member falls into a chromosomal group or an extrachromosomal group. It is known that the parAB genes in the chromosomal group are located on relatively conserved chromosomal regions in several bacterial species. This suggests that the parAB genes were transferred from a chromosome to plasmids and phages, so the genes have diverged among bacterial species. To support this possibility, we show that the Bacillus subtilis Soj and Spo0J members of the ParAB families are responsible for the specific localization of plasmids at cell quarters in E. coli and can function as partition proteins. Host factors to tether actively partitioning plasmids at subcellular sites may be conserved in Gram-negative and Gram-positive bacteria so that phages and plasmids with the ParAB partitioning system can be stably inherited in host cells across bacterial species. PMID- 11121067 TI - A whole-genome microarray reveals genetic diversity among Helicobacter pylori strains. AB - Helicobacter pylori colonizes the stomach of half of the world's population, causing a wide spectrum of disease ranging from asymptomatic gastritis to ulcers to gastric cancer. Although the basis for these diverse clinical outcomes is not understood, more severe disease is associated with strains harboring a pathogenicity island. To characterize the genetic diversity of more and less virulent strains, we examined the genomic content of 15 H. pylori clinical isolates by using a whole genome H. pylori DNA microarray. We found that a full 22% of H. pylori genes are dispensable in one or more strains, thus defining a minimal functional core of 1281 H. pylori genes. While the core genes encode most metabolic and cellular processes, the strain-specific genes include genes unique to H. pylori, restriction modification genes, transposases, and genes encoding cell surface proteins, which may aid the bacteria under specific circumstances during their long-term infection of genetically diverse hosts. We observed distinct patterns of the strain-specific gene distribution along the chromosome, which may result from different mechanisms of gene acquisition and loss. Among the strain-specific genes, we have found a class of candidate virulence genes identified by their coinheritance with the pathogenicity island. PMID- 11121068 TI - Nitrogen regulatory protein C-controlled genes of Escherichia coli: scavenging as a defense against nitrogen limitation. AB - Nitrogen regulatory protein C (NtrC) of enteric bacteria activates transcription of genes/operons whose products minimize the slowing of growth under nitrogen limiting conditions. To reveal the NtrC regulon of Escherichia coli we compared mRNA levels in a mutant strain that overexpresses NtrC-activated genes [glnL(Up)] to those in a strain with an ntrC (glnG) null allele by using DNA microarrays. Both strains could be grown under conditions of nitrogen excess. Thus, we could avoid differences in gene expression caused by slow growth or nitrogen limitation per se. Rearranging the spot images from microarrays in genome order allowed us to detect all of the operons known to be under NtrC control and facilitated detection of a number of new ones. Many of these operons encode transport systems for nitrogen-containing compounds, including compounds recycled during cell-wall synthesis, and hence scavenging appears to be a primary response to nitrogen limitation. In all, approximately 2% of the E. coli genome appears to be under NtrC control, although transcription of some operons depends on the nitrogen assimilation control protein, which serves as an adapter between NtrC and final sigma(70)-dependent promoters. PMID- 11121069 TI - In vivo induction of massive proliferation, directed migration, and differentiation of neural cells in the adult mammalian brain. AB - The development of an in vivo procedure for the induction of massive proliferation, directed migration, and neurodifferentiation (PMD) in the damaged adult central nervous system would hold promise for the treatment of human neurodegenerative disorders such as Parkinson's disease. We investigated the in vivo induction of PMD in the forebrain of the adult rat by using a combination of 6-hydroxydopamine lesion of the substantia nigra dopaminergic neurons and infusions of transforming growth factor alpha (TGFalpha) into forebrain structures. Only in animals with both lesion and infusion of TGFalpha was there a rapid proliferation of forebrain stem cells followed by a timed migration of a ridge of neuronal and glial progenitors directed toward the region of the TGFalpha infusion site. Subsequently, increasing numbers of differentiated neurons were observed in the striatum. In behavioral experiments, there was a significant reduction of apomorphine-induced rotations in animals receiving the TGFalpha infusions. These results show that the brain contains stem cells capable of PMD in response to an exogenously administered growth factor. This finding has significant implications with respect to the development of treatments for both acute neural trauma and neurodegenerative diseases. PMID- 11121070 TI - Altered gating of opiate receptor-modulated K+ channels on amygdala neurons of morphine-dependent rats. AB - The molecular mechanism of tolerance to opiate drugs is poorly understood. We have used single-channel patch-clamp recordings to study opiate receptor effects on dissociated neurons from rat amygdala, a limbic region implicated in addiction processes. A 130-pS inwardly rectifying K(+)-preferring cation channel was activated by mu opioid receptors in a membrane-delimited manner. After chronic treatment of the rats with morphine, channel gating changed markedly, with an approximately 100-fold decrease in open probability at a given morphine concentration. The change in channel gating correlated both in time course and in dose of morphine treatment with the development of functional opiate dependence and appeared to arise at a step after G-protein activation and before channel permeation by K(+). This decreased receptor-channel coupling appears to be large enough to account quantitatively for opiate tolerance and may represent one of the mechanisms through which tolerance occurs. PMID- 11121071 TI - Direct isolation of human central nervous system stem cells. AB - Stem cells, which are clonogenic cells with self-renewal and multilineage differentiation properties, have the potential to replace or repair damaged tissue. We have directly isolated clonogenic human central nervous system stem cells (hCNS-SC) from fresh human fetal brain tissue, using antibodies to cell surface markers and fluorescence-activated cell sorting. These hCNS-SC are phenotypically 5F3 (CD133)(+), 5E12(+), CD34(-), CD45(-), and CD24(-/lo). Single CD133(+) CD34(-) CD45(-) sorted cells initiated neurosphere cultures, and the progeny of clonogenic cells could differentiate into both neurons and glial cells. Single cells from neurosphere cultures initiated from CD133(+) CD34(-) CD45(-) cells were again replated as single cells and were able to reestablish neurosphere cultures, demonstrating the self-renewal potential of this highly enriched population. Upon transplantation into brains of immunodeficient neonatal mice, the sorted/expanded hCNS-SC showed potent engraftment, proliferation, migration, and neural differentiation. PMID- 11121072 TI - Impaired hippocampal-dependent learning and functional abnormalities in the hippocampus in mice lacking serotonin(1A) receptors. AB - The hippocampus is a major limbic target of the brainstem serotonergic neurons that modulate fear, anxiety, and learning through postsynaptic serotonin(1A) receptors (5-HT(1A) receptors). Because chronic stress selectively down-regulates the 5-HT(1A) receptors in the hippocampus, we hypothesized that mice lacking these receptors may exhibit abnormalities reminiscent of symptoms of stress related psychiatric disorders. In particular, a hippocampal deficit in the 5 HT(1A) receptor could contribute to the cognitive abnormalities often seen in these disorders. To test whether a deficit in 5-HT(1A) receptors impairs hippocampus-related functions, we studied hippocampal-dependent learning and memory, synaptic plasticity in the hippocampus, and limbic neuronal excitability in 5-HT(1A)-knockout (KO) mice. 5-HT(1A)-KO animals showed a deficit in hippocampal-dependent learning and memory tests, such as the hidden platform (spatial) version of the Morris water maze and the delayed version of the Y maze. The performance of KO mice was not impaired in nonhippocampal memory tasks such as the visible platform (nonspatial) version of the Morris water maze, the immediate version of the Y maze, and the spontaneous-alternation test of working memory. Furthermore, paired-pulse facilitation in the dentate gyrus of the hippocampus was impaired in 5-HT(1A)-KO mice. Finally, 5-HT(1A)-KO mice, as compared with wild-type animals, displayed higher limbic excitability manifested as lower seizure threshold and higher lethality in response to kainic acid administration. These results demonstrate that 5-HT(1A) receptors are required for maintaining normal hippocampal functions and implicate a role for the 5 HT(1A) receptor in hippocampal-related symptoms, such as cognitive disturbances, in stress-related disorders. PMID- 11121074 TI - Top-down processing mediated by interareal synchronization. AB - Perception and cortical responses are not only driven "bottom-up" by the external stimulus but are altered by internal constraints such as expectancy or the current behavioral goal. To investigate neurophysiological mechanisms of such top down effects, we analyzed the temporal interactions of neurons on different levels of the cortical hierarchy during perception of stimuli with varying behavioral significance. We found that interareal interactions in a middle frequency range (theta and alpha; 4-12 Hz) strongly depend on the associated behavior, with a phase relationship and a layer specificity indicating a top-down directed interaction. For novel unexpected stimuli, presumably processed in a feed-forward fashion, no such interactions occurred but high-frequency interactions (gamma; 20-100 Hz) were observed. Thus corticocortical synchronization reflects the internal state of the animal and may mediate top down processes. PMID- 11121073 TI - Deletion of tyrosine hydroxylase gene reveals functional interdependence of adrenocortical and chromaffin cell system in vivo. AB - Catecholamines are produced in the medulla of the adrenal gland and may participate in the intraglandular regulation of its cortex. We analyzed the adrenal structure and function of albino tyrosine hydroxylase-null (TH-null) mice that are deficient in adrenal catecholamine production. Adrenal catecholamines were markedly reduced, and catecholamine histofluorescence was abrogated in 15 day-old TH-null mice. Chromaffin cell structure was strikingly altered at the ultrastructural level with a depletion of chromaffin vesicles and an increase in rough endoplasmic reticulum compared with wild-type mice. Remaining chromaffin vesicles lined up proximally to the cell membrane in preparation for exocytosis providing a "string-of-pearls" appearance. There was a 5-fold increase in the expression of proenkephalin mRNA (502.8 +/- 142% vs. 100 +/- 17.5%, P = 0.016) and a 2-fold increase in the expression of neuropeptide Y (213.4 +/- 41.2% vs. 100 +/- 59.9%, P = 0.014) in the TH-null animals as determined by quantitative TaqMan (Perkin-Elmer) PCR. Accordingly, immunofluorescence for met-enkephalin and neuropeptide tyrosine in these animals was strongly enhanced. The expression of phenylethanolamine N-methyl transferase and chromogranin B mRNA was similar in TH null and wild-type mice. In TH-null mice, adrenocortical cells were characterized by an increase in liposomes and by tubular mitochondria with reduced internal membranes, suggesting a hypofunctional state of these steroid-producing cells. In accordance with these findings, plasma corticosterone levels were decreased. Plasma ACTH levels were not significantly different in TH-null mice. In conclusion, both the adrenomedullary and adrenocortical systems demonstrate structural and functional changes in catecholamine-deficient TH-null mice, underscoring the great importance of the functional interdependence of these systems in vivo. PMID- 11121075 TI - Molecular determinants of membrane potential dependence in vertebrate gap junction channels. AB - The conductance, g(j), of many gap junctions depends on voltage between the coupled cells (transjunctional voltage, V(j)) with little effect of the absolute membrane potential (V(m)) in the two cells; others show combined V(j) and V(m) dependence. We examined the molecular determinants of V(m) dependence by using rat connexin 43 expressed in paired Xenopus oocytes. These junctions have, in addition to V(j) dependence, V(m) dependence such that equal depolarization of both cells decreases g(j). The dependence of g(j) on V(m) was abolished by truncation of the C-terminal domain (CT) at residue 242 but not at 257. There are two charged residues between 242 and 257. In full-length Cx43, mutations neutralizing either one of these charges, Arg243Gln and Asp245Gln, decreased and increased V(m) dependence, respectively, suggesting that these residues are part of the V(m) sensor. Mutating both residues together abolished V(m) dependence, although there is no net change in charge. The neutralizing mutations, together or separately, had no effect on V(j) dependence. Thus, the voltage sensors must differ. However, V(j) gating was somewhat modulated by V(m), and V(m) gating was reduced when the V(j) gate was closed. These data suggest that the two forms of voltage dependence are mediated by separate but interacting domains. PMID- 11121076 TI - Understanding and exploiting the mechanistic basis for selectivity of polyketide inhibitors of F(0)F(1)-ATPase. AB - Recently, a family of polyketide inhibitors of F(0)F(1)-ATPase, including apoptolidin, ossamycin, and oligomycin, were shown to be among the top 0.1% most cell line selective cytotoxic agents of 37, 000 molecules tested against the 60 human cancer cell lines of the National Cancer Institute. Many cancer cells maintain a high level of anaerobic carbon metabolism even in the presence of oxygen, a phenomenon that is historically known as the Warburg effect. A mechanism-based strategy to sensitize such cells to this class of potent small molecule cytotoxic agents is presented. These natural products inhibit oxidative phosphorylation by targeting the mitochondrial F(0)F(1) ATP synthase. Evaluation of gene expression profiles in a panel of leukemias revealed a strong correlation between the expression level of the gene encoding subunit 6 of the mitochondrial F(0)F(1) ATP synthase (known to be the binding site of members of this class of macrolides) and their sensitivity to these natural products. Within the same set of leukemia cell lines, comparably strong drug-gene correlations were also observed for the genes encoding two key enzymes involved in central carbon metabolism, pyruvate kinase, and aspartate aminotransferase. We propose a simple model in which the mitochondrial apoptotic pathway is activated in response to a shift in balance between aerobic and anaerobic ATP biosynthesis. Inhibitors of both lactate formation and carbon flux through the Embden-Meyerhof pathway significantly sensitized apoptolidin-resistant tumors to this drug. Nine different cell lines derived from human leukemias and melanomas, and colon, renal, central nervous system, and ovarian tumors are also sensitized to killing by apoptolidin. PMID- 11121077 TI - Highly specific, membrane-permeant peptide blockers of cGMP-dependent protein kinase Ialpha inhibit NO-induced cerebral dilation. AB - Arrays of octameric peptide libraries on cellulose paper were screened by using (32)P-autophosphorylated cGMP-dependent protein kinase Ialpha (cGPK) to identify peptide sequences with high binding affinity for cGPK. Iterative deconvolution of every amino acid position in the peptides identified the sequence LRK(5)H (W45) as having the highest binding affinity. Binding of W45 to cGPK resulted in selective inhibition of the kinase with K(i) values of 0.8 microM and 560 microM for cGPK and cAMP-dependent protein kinase (cAPK), respectively. Fusion of W45 to membrane translocation signals from HIV-1 tat protein (YGRKKRRQRRRPP-LRK(5)H, DT 2) or Drosophila Antennapedia homeo-domain (RQIKIWFQNRRMKWKK-LRK(5)H, DT-3) proved to be an efficient method for intracellular delivery of these highly charged peptides. Rapid translocation of the peptides into intact cerebral arteries was demonstrated by using fluorescein-labeled DT-2 and DT-3. The inhibitory potency of the fusion peptides was even greater than that for W45, with K(i) values of 12.5 nM and 25 nM for DT-2 and DT-3, respectively. Both peptides were still poor inhibitors of cAPK. Selective inhibition of cGPK by DT-2 or DT-3 in the presence of cAPK was demonstrated in vitro. In pressurized cerebral arteries, DT-2 and DT-3 substantially decreased NO-induced dilation. This study provides functional characterization of a class of selective cGPK inhibitor peptides in vascular smooth muscle and reveals a central role for cGPK in the modulation of vascular contractility. PMID- 11121078 TI - Completing the heterotrimer: isolation and characterization of an Arabidopsis thaliana G protein gamma-subunit cDNA. AB - Heterotrimeric G proteins consist of three subunits (alpha, beta, and gamma). alpha- and beta- subunits have been previously cloned in plants, but the gamma subunit has remained elusive. To isolate the gamma-subunit of a plant heterotrimeric G protein an Arabidopsis thaliana yeast two-hybrid library was screened by using a tobacco G-beta-subunit as the bait protein. One positive clone (AGG1) was isolated several times; it displays significant homology to the conserved domains of mammalian gamma-subunits. The predicted AGG1 protein sequence contains all of the typical characteristics of mammalian gamma-subunits such as small size (98 amino acids, 10.8 kDa), presence of a C-terminal CAAX box to direct isoprenyl modification, and an N-terminal alpha-helix region capable of forming a coiled-coil interaction with the beta-subunit. Northern and Southern analyses showed that AGG1 is a single-copy gene in Arabidopsis with a similar expression pattern to the Arabidopsis beta-subunit, AGB1 [Weiss, C. A., Garnaat, C. W., Mukai, K., Hu, Y. & Ma, H. (1994) Proc. Natl. Acad. Sci. USA 91, 9554 9558]. By using the yeast two-hybrid system, we show that AGG1 strongly interacts with tobacco and Arabidopsis beta-subunits. The in vivo results have been confirmed by using in vitro methods to prove the interaction between AGG1 and the Arabidopsis beta-subunit. As previously observed in mammalian systems, both the coiled-coil domain and the WD repeat regions of the beta-subunit are essential for AGG1 interaction. Also in agreement with previous observations, the removal of the N-terminal alpha-helix of the AGG1 greatly reduces but does not completely block the interaction. PMID- 11121079 TI - Structure-function analysis of the tobacco mosaic virus resistance gene N. AB - The tobacco N gene is a member of the Toll-interleukin-1 receptor/nucleotide binding site/leucine-rich repeat (TIR-NBS-LRR) class of plant resistance (R) genes and confers resistance to tobacco mosaic virus (TMV). We investigated the importance of specific domains of N in inducing TMV resistance, by examining various N deletion and point mutations that introduce single amino acid substitution mutants in vivo. Our deletion analysis suggests that the TIR, NBS, and LRR domains play an indispensable role in the induction of resistance responses against TMV. We show that amino acids conserved among the Toll/IL 1R/plant R gene TIR domain and NBS-containing proteins play a critical role in N mediated TMV resistance. Some loss-of-function N alleles such as the TIR deletion and point mutations in the NBS (G216A/E/V/R, G218R, G219D, K222E/N, and T223A/N) interfere with the wild-type N function and behave like dominant negative mutations. These F(1) plants mount a hypersensitive response (HR) that is indistinguishable from that of the wild-type N plants, yet TMV was able to move systemically, causing a systemic hypersensitive response (SHR). Many amino acid substitutions in the TIR, NBS, and LRR domains of N lead to a partial loss-of function phenotype. These mutant plants mount delayed HR compared with the wild type N plants and fail to contain the virus to the infection site. In addition, some partial loss-of-function alleles (W82S/A, W141S/A, G218V/S, and G219V) interfere with the wild-type N function, leading to SHR. The partial loss-of function and dominant negative mutant alleles described in this report will be useful in furthering our understanding of the TIR-NBS-LRR class of R genes. PMID- 11121081 TI - Heritable basis for choice of group size in a colonial bird. AB - Sizes of most kinds of animal groups vary considerably within a population, with group size often causing direct effects on the fitness of group members. Although the consequences of varying group size have been well studied, the causes of variation in group size remain poorly known for most animals. Groups might vary in size because different individuals perform better in differently sized groups and thus have genetic predispositions to choose large or small groups. We examined whether heritable variation for choice of group size exists in the cliff swallow (Petrochelidon pyrrhonota), a colonial bird that nests in colonies ranging from 2 to 3,700 nests. Parent-offspring regressions showed significant heritabilities for choice of colony size under natural conditions. Partial cross fostering experiments showed that individuals reared in colonies of sizes different from those of their birth returned to breed the next year in colonies that matched their birth colony in size and actively avoided those similar to their rearing colony, suggesting that choice of colony size is genetically based. Common environmental effects, maternal effects, and philopatry did not explain these results. Variation in group size probably results in part from a polymorphism in genetic preferences within the population, and the range in colony sizes is maintained by natural selection on the type of bird occupying each site. PMID- 11121086 TI - Clinical trials: New developments. PMID- 11121080 TI - Characterization of Mbb1, a nucleus-encoded tetratricopeptide-like repeat protein required for expression of the chloroplast psbB/psbT/psbH gene cluster in Chlamydomonas reinhardtii. AB - Genetic analysis has revealed that the accumulation of several chloroplast mRNAs of the green alga Chlamydomonas reinhardtii requires specific nucleus-encoded functions. To gain insight into this process, we have cloned the nuclear gene encoding the Mbb1 factor by genomic rescue of a mutant specifically deficient in the accumulation of the mRNAs of the psbB/psbT/psbH chloroplast transcription unit. Mbb1 is a soluble protein in the stromal phase of the chloroplast. It consists of 662 amino acids with a putative chloroplast-transit peptide at its N terminal end. A striking feature is the presence of 10 tandemly arranged tetratricopeptide-like repeats that account for half of the protein sequence and are thought to be involved in protein-protein interactions. The Mbb1 protein seems to have a homologue in higher plants and is part of a 300-kDa complex that is associated with RNA. This complex is most likely involved in psbB mRNA processing, stability, and/or translation. PMID- 11121087 TI - The CIHR Circulatory and Respiratory Health Institute. PMID- 11121088 TI - Introducing the fifth edition of the Canadian Tuberculosis Standards. PMID- 11121089 TI - Respirology manpower in Canada--A report for the Canadian Thoracic Society Education Committee. AB - A report on adult and pediatric respirology manpower in Canada was prepared from data supplied by the Royal College of Physicians and Surgeons of Canada (RCPSC), and from program directors (and other colleagues) at universities across Canada. The data support a significant deficiency of adult respirologists in Canada, which is estimated to be from 10%, based on a 10-year-old outdated RCPSC recommendation, to 20%, based on equalization with the 'best' province, to as high as 50%, based on long waiting lists, particularly for respiratory sleep problems, and estimates obtained from academic centres across Canada. Although there are less data available for pediatric respirology, a similar approach suggests a 50% to 100% shortfall in pediatric respirologists. Output from Canadian training programs in adult and pediatric respirology is not likely to meet this need. We recommend that steps be taken urgently to provide sufficient resources for training adult and pediatric respirologists, and to ensure that funding is provided for subspecialist positions in the community. PMID- 11121090 TI - Physician asthma management practices in Canada. AB - OBJECTIVES: To establish national baseline information on asthma management practices of physicians, to compare the reported practices with the Canadian Consensus recommendations and to identify results potentially useful for interventions that improve physician asthma management practices. DESIGN: National, stratified cross-sectional survey. SETTINGS: The 10 provinces and two territories of Canada, from 1996 to 1997. PARTICIPANTS: Questionnaires were sent to 4489 physicians stratified by province/territory and specialty group (family/general practice, respirology, internal medicine, pediatrics and allergy/immunology); 2605 responses were received. OUTCOME MEASURES: Methods for the diagnosis, treatment, education and follow-up of patients with asthma ('asthma management practices'). RESULTS: Significant variations existed among the five specialty groups in asthma management practices. A low use of objective measures of airflow limitation to assist with diagnosis was found among some respondents (mostly family physicians). Up to 40% of physicians regarded the daily fixed dosing (three or four times a day) of inhaled, short acting beta2 agonist as 'first-line therapy' for moderate to severe asthma. A minority of physicians reported using written action plans for patients or referring them to other health professionals for asthma education. Insufficient time during appointments and a perceived lack of appropriate educational materials were frequently cited as reasons for not providing asthma education. The perceived knowledge of the Canadian Consensus recommendations varied among physicians but was lowest among nonspecialists. CONCLUSIONS: The survey showed variations in certain aspects of the management of asthma by physicians. The findings will help to target specific areas for future physician education programs and other behavioural change strategies. PMID- 11121091 TI - Outpatient antibiotic therapy and short term mortality in elderly patients with chronic obstructive pulmonary disease. AB - OBJECTIVE: To determine the association between outpatient use of oral antibiotics and 30-day all-cause mortality following hospitalization in a group of elderly chronic obstructive pulmonary disease (COPD) patients. DESIGN: A population-based retrospective cohort study. SETTING: Ontario, Canada. POPULATION STUDIED: All 26,301 patients, 65 years of age or older, who were hospitalized for COPD between 1992 and 1996 in Ontario. METHODS: All elderly patients admitted at least once with a most responsible diagnosis of COPD using the Canadian Institute for Health Information database were identified. They were then linked to the Ontario Drug Benefit database to determine the use of antibiotics within 30 days of the index hospitalization and to the Ontario registered persons database to determine the 30-day mortality following their index hospitalization. RESULTS: Outpatient use of antibiotics within 30 days before the index hospitalization was associated with a significant reduction in the 30-day mortality following hospitalization (odds ratio [OR] 0.83, 95% CI 0.75 to 0.92). Use of macrolides had the lowest relative odds for mortality (OR 0.58, 95% CI 0.47 to 0.73), while use of fluoroquinolones had the highest relative odds (OR 0.98, 95% CI 0.84 to 1.15). CONCLUSIONS: Use of antibiotics before hospitalization was associated with a significant reduction in the risk of short term mortality among a group of elderly COPD patients who eventually required hospitalization for their disease. These findings support the early use of antibiotics in COPD patients who experience an acute exacerbation. PMID- 11121092 TI - Serum and red blood cell antioxidant status in patients with bronchial asthma. AB - Levels of vitamin C, ceruloplasmin, transferrin and albumin in serum, and glutathione in red blood cells were investigated in 40 patients with asthma to determine whether their antioxidant status was different from healthy subjects. Serum vitamin C and albumin levels were lower in the patient group (36.91+/-12.50 microM and 46.2+/-3.0 g/L, respectively) than in 43 healthy volunteers (53.38+/ 13.06 microM and 48.8+/-2.1 g/L, P<0.001 and P<0.05, respectively). However, erythrocyte glutathione and serum ceruloplasmin levels were higher in the patient group (0.59+/-0.11 mol/mol hemoglobin and 442+/-73 micromol/L, respectively) than in controls (0.49+/-0.09 mol/mol hemoglobin and 308+/-47 micromol/L, P<0.001 and P<0.001, respectively). No difference was observed in transferrin levels between the groups. The results suggest that reactive oxygen species may be a contributing factor in patients with asthma, causing changes in serum vitamin C, ceruloplasmin and erythrocyte glutathione levels. PMID- 11121093 TI - Thoracic complications of Lemierre syndrome. AB - Lemierre syndrome is a severe, septicemic illness most commonly caused by the anaerobic Gram-negative bacillus Fusobacterium necrophorum. It is characterized by an acute oropharyngeal infection, with secondary septic thrombophlebitis of the internal jugular vein and frequent metastatic infections. This report of a patient with the Lemierre syndrome is complemented by a review of the literature on the subject. PMID- 11121094 TI - Hydroxyurea-induced hypersensitivity pneumonitis: A case report and literature review. AB - Hydroxyurea is a cytotoxic agent indicated in the treatment of a variety of malignant and nonmalignant conditions. Apart from dose-related bone marrow suppression, this antineoplastic agent is generally well tolerated. This report describes a patient with chronic myeloid leukemia who developed severe pneumonitis within four weeks of beginning therapy with hydroxyurea. Pathological examination of a lung specimen obtained by video-assisted thoracoscopic lung biopsy revealed extensive active alveolar and interstitial inflammation, and poorly formed granulomas. After the cessation of hydroxyurea and treatment with systemic corticosteroids, both clinical and radiological resolution of pneumonitis occurred. Physicians using hydroxyurea must be aware of its potentially life-threatening pulmonary toxicity. PMID- 11121095 TI - Methyl-CpG-binding proteins. Targeting specific gene repression. AB - CpG methylation, the most common epigenetic modification of vertebrate genomes, is primarily associated with transcriptional repression. MeCP2, MBD1, MBD2, MBD3 and MBD4 constitute a family of vertebrate proteins that share the methyl-CpG binding domain (MBD). The MBD, consisting of about 70 residues, possesses a unique alpha/beta-sandwich structure with characteristic loops, and is able to bind single methylated CpG pairs as a monomer. All MBDs except MBD4, an endonuclease that forms a complex with the DNA mismatch-repair protein MLH1, form complexes with histone deacetylase. It has been established that MeCP2, MBD1 and MBD2 are involved in histone deacetylase-dependent repression and it is likely that this is also the case for MBD3. The current model proposes that MBD proteins are involved in recruiting histone deacetylases to methyl CpG-enriched regions in the genome to repress transcription. The lack of selectivity for MBD association with particular DNA sequences indicates that other mechanisms account for their recruitment to particular regions in the genome. PMID- 11121096 TI - Interaction of linear homologous DNA duplexes via Holliday junction formation. AB - Interaction of linear homologous DNA duplexes by formation of Holliday junctions was revealed by electrophoresis and confirmed by electron microscopy. The phenomenon was demonstrated using a model of five purified PCR products of different size and sequence. The double-stranded structure of interacting DNA fragments was confirmed using several consecutive purifications, S1-nuclease analysis, and electron microscopy. Formation of Holliday junctions depends on DNA concentration. A thermodynamic equilibrium between duplexes and Holliday junctions was shown. We propose that homologous duplex interaction is initiated by nucleation of several dissociated terminal base pairs of two fragments. This process is followed by branch migration creating a population of Holliday junctions with the branch point at different sites. Finally, Holliday junctions are resolved via branch migration to new or previously existing duplexes. The phenomenon is a new property of DNA. This type of DNA-DNA interaction may contribute to the process of Holliday junction formation in vivo controlled by DNA conformation and DNA-protein interactions. It is of practical significance for optimization of different PCR-based methods of gene analysis, especially those involving heteroduplex formation. PMID- 11121097 TI - Identification of two highly divergent catalase genes in the fungal tomato pathogen, Cladosporium fulvum. AB - Catalases of pathogenic micro-organisms have attracted attention as potential virulence factors. Homology-based screens were performed to identify catalase genes in the fungal tomato pathogen Cladosporium fulvum. Two highly divergent genes, Cat1 and Cat2, were isolated and characterized. Cat1 codes for a putative 566-amino-acid catalase subunit and belongs to the gene family that also encodes the mainly peroxisome-localized catalases of animal and yeast species. Cat2 codes for a putative catalase subunit of 745 amino acids and belongs to a different gene family coding for the large-subunit catalases similar to ones found in bacteria and filamentous fungi. Neither catalase had an obvious secretory signal sequence. A search for an extracellular catalase was unproductive. The Cat1 and Cat2 genes showed differential expression, with the Cat1 mRNA preferentially accumulating in spores and the Cat2 mRNA preferentially accumulating in response to external H(2)O(2). With Cat2-deleted strains, activity of the Cat2 gene product (CAT2) was identified among four proteins with catalase activity separated on non-denaturing gels. The CAT2 activity represented a minor fraction of the catalase activity in spores and H(2)O(2)-stressed mycelium, and no phenotype was observed for Cat2-deleted strains, which showed a normal response to H(2)O(2) treatment. These results indicate the existence of a complex catalase system in C. fulvum, with regard to both the structure and regulation of the genes involved. In addition, efficient C. fulvum gene-replacement technology has been established. PMID- 11121098 TI - Involvement of insulin receptor substrates in epidermal growth factor induced activation of phosphatidylinositol 3-kinase in rat hepatocyte primary culture. AB - Short-term incubation of adult rat hepatocytes with epidermal growth factor (EGF) caused tyrosine phosphorylation of insulin receptor substrate (IRS)-1 and IRS-2 when the cells had been submitted to primary culture from 1-18 h. Tyrosine phosphorylated IRS-1 and IRS-2 bound to the regulatory subunit (p85) of phosphatidylinositol (PtdIns) 3-kinase, thereby activating the enzymic activity. Tyrosine phosphorylation of the IRSs and activation of PtdIns 3-kinase in 3 h cultured hepatocytes both proceeded similarly to the same actions of insulin; the activation was rapid and transient, with peak values at 15-30 s and with similar EC(50)s in the nM range in both cases. A possible involvement of insulin receptors in these insulin-like actions of EGF was excluded by the following three lines of evidence. Insulin caused tyrosine phosphorylation of the insulin receptor beta-subunit but EGF did not. In contrast, the EGF receptor was phosphorylated by EGF, but the insulin receptor was not. The actions of EGF, but not those of insulin, were inhibited by AG1478, a selective inhibitor of EGF receptor tyrosine kinase. Cultured hepatocytes exposed to insulin or insulin-like growth factor-I (IGF-I) for a short period responded to the subsequent addition of EGF, whereas EGF-treated cells responded to insulin. The cells, however, displayed receptor desensitization under the same conditions, that is, no response was observed upon repeated addition of the same agonist, EGF, insulin or IGF-I. Thus, the EGF receptor-initiated signalling was mediated by PtdIns 3 kinase associated with tyrosine-phosphorylated IRSs in short-term cultured rat hepatocytes. PMID- 11121099 TI - Novel alpha-L-fucosidase inhibitors from the bark of Angylocalyx pynaertii (Leguminosae). AB - The extract of bark of Angylocalyx pynaertii (Leguminosae) was found to potently inhibit mammalian alpha-L-fucosidases. A thorough examination of the extract resulted in the discovery of 15 polyhydroxylated alkaloids, including the known alkaloids from seeds of this plant, 1,4-dideoxy-1,4-imino-D-arabinitol (DAB), 1 deoxymannojirimycin (DMJ) and 2,5-imino-1,2,5-trideoxy-D-mannitol (6-deoxy-DMDP). Among them, eight sugar-mimic alkaloids showed the potent inhibitory activity towards bovine epididymis alpha-L-fucosidase and their Ki values are as follows: 6-deoxy-DMDP (83 microM), 2,5-imino-1,2,5-trideoxy-L-glucitol (0.49 microM), 2,5 dideoxy-2,5-imino-D-fucitol (17 microM), 2,5-imino-1,2,5-trideoxy-D-altritol (3.7 microM), DMJ (4.7 microM), N-methyl-DMJ (30 microM), 6-O-alpha-L-rhamnopyranosyl DMJ (Rha-DMJ, 0.06 microM), and beta-L-homofuconojirimycin (beta-HFJ, 0.0053 microM). We definitively deduced the structural requirements of inhibitors of alpha-L-fucosidase for the piperidine alkaloids (DMJ derivatives). The minimum structural feature of alpha-L-fucosidase inhibitors is the correct configuration of the three hydroxyl groups on the piperidine ring corresponding to C2, C3 and C4 of L-fucose. Furthermore, the addition of a methyl group in the correct configuration to the ring carbon atom corresponding to C5 of L-fucose generates extremely powerful inhibition of alpha-L-fucosidase. The replacement of the methyl group of beta-HFJ by a hydroxymethyl group reduced its inhibitory potential about 80-fold. This suggests that there may be a hydrophobic region in or around the active site. The existence or configuration of a substituent group on the ring carbon atom corresponding to the anomeric position of L-fucose does not appear to be important for the inhibition. Interestingly, Rha-DMJ was a 70 fold more potent inhibitor of alpha-L-fucosidase than DMJ. This implies that the lysosomal alpha-L-fucosidase may have subsites recognizing oligosaccharyl structures in natural substrates. PMID- 11121100 TI - Characterization of the Co(2+) and Ni(2+) binding amino-acid residues of the N terminus of human albumin. An insight into the mechanism of a new assay for myocardial ischemia. AB - Patients suffering from myocardial ischemia reportedly exhibit reduced in vitro binding of exogenous Co(2+) to the N-terminal of human serum albumin (HSA). The purpose of our investigation was to simulate changes in the N-terminus of HSA that may account for these ischemia-induced modifications to the cobalt binding site. HPLC, LC-MS and (1)H NMR analyses have shown that the N-terminal region of HSA Asp-Ala-His-Lys binds the transition metals Co(2+) and Ni(2+). Synthetic peptides with the first 2-12 amino acids of the HSA sequence demonstrated that the first three amino acids, Asp-Ala-His, are essential for strong binding of cobalt. Modification of the N-terminus peptide of HSA by way of N-acetylation or the deletion of one or more amino acid resulted in no binding of cobalt. Because the degradation of the susceptible, specific transition metal binding site of HSA may account for the decreased cobalt binding observed during ischemic events, an assay that detects this reduced binding could be useful in the diagnosis of ischemia. PMID- 11121101 TI - Leakage and aggregation of phospholipid vesicles induced by the BH3-only Bcl-2 family member, BID. AB - BID is a BH3 domain-only member of the Bcl-2 family that acts as an apoptotic agonist in programmed cell death. After cleavage by caspase-8, the N-terminal of BID (N-BID) stays in the cytosol while the C-terminal of BID (C-BID) translocates to mitochondria, leading to cytochrome c release in vivo and in vitro. We have previously reported that BID or truncated BID (tBID) can induce the release of entrapped trypsin and cytochrome c from large unilamellar vesicles (LUVs). Further studies have been performed and are presented here; the results demonstrate that C-BID, like BID and tBID, induces vesicle leakage, whereas N-BID or the BID mutants BID (D59A) and BID (G94E) fail to have any significant effects. The affinity of the above-mentioned proteins for soybean phospholipid LUVs (SLUVs) decreased in an order similar to their leakage-inducing capability: tBID > BID > BID (D59A), while N-BID and BID (G94E) were unable to bind to the vesicles at all. BID-induced leakage was dependent on the lipid composition of vesicles. Acidic phospholipid (e.g. phosphatidic acid or phosphatidylglycerol) was necessary for BID-induced leakage while the presence of phosphatidylethanolamine or cholesterol reduced the leakage. It was also found C BID is better able to penetrate the soybean phospholipid monolayer than BID or tBID. A further finding was that tBID, but not full-length BID, could stimulate the aggregation of SLUVs. Finally, Bcl-x(L), an apoptotic antagonist in programmed cell death, can prevent the aggregation of LUVs induced by tBID, but not the release of entrapped trypsin. It is postulated that two separate domains of tBID are responsible for inducing leakage and aggregation of phospholipid vesicles. PMID- 11121102 TI - Major sites for the differentiation of V alpha 14(+) NKT cells inferred from the V-J junctional sequences of the invariant T-cell receptor alpha chain. AB - CD1d-restricted mouse NK1.1(+) TCR alpha beta(+) natural killer T (NKT) cells predominantly use an invariant TCR alpha chain encoded by V alpha 14 and J alpha 281 gene segments with a one-nucleotide N region. We found that NKT cells generated in the culture of fetal liver precursors possessed V alpha 14-J alpha 281 junctions that could be produced without the action of terminal deoxyribonucleotidyl transferase (TdT), indicating that NKT cells derived from fetal liver precursors are distinguishable from those from adult precursors with TdT expression. In fact, the frequency of the fetal-form sequences decreased with ageing. Surprisingly, the fetal-type sequences were predominantly observed in the lymphoid organs of athymic mice with the exception of bone marrow, where a sequence peculiar to the organ, with TdT-involved conversion from the invariant junction, was frequently present. These findings suggest that there are two independent sites of V alpha 14(+) NKT cell development, the hematopoietic organs throughout life (the developing liver and adult bone marrow) and, principally, the mature thymus. PMID- 11121103 TI - CREB is involved in mouse annexin A1 regulation by cAMP and glucocorticoids. AB - A cAMP and some glucocorticoid response elements have been underlined in the promoter of mouse annexin A1. To analyse the function of these DNA sequences, the role of cAMP and glucocorticoids, as well as the transcription factors involved in their activation, were investigated. A construct containing 1381 bp of the DNA 5'-flanking annexin A1 gene fused to LacZ was used. The level of activation of the reporter gene was analysed by transient transfection of the JEG3 cell line. Activation of beta-galactosidase expression was observed with both dibutyryl cAMP and dexamethasone when compared with cells treated with serum only. Simultaneous addition of dexamethasone and dibutyryl cAMP did not result in a synergistic effect but rather in a competitive one. Gel-shift assays with a probe including the cAMP response element-like element of the annexin A1 promoter revealed a main specific DNA-protein complex when cells were stimulated with dibutyryl cAMP and/or dexamethasone. In all cases CREB protein was identified by supershift analysis. We therefore conclude that this cAMP response element sequence plays a prominent role in the transactivation of the annexin A1 promoter by dibutyryl cAMP and that it is involved in the response to glucocorticoids. PMID- 11121104 TI - The dual role of thrombin's anion-binding exosite-I in the recognition and cleavage of the protease-activated receptor 1. AB - The role of thrombin anion-binding exosite-I in the recognition and cleavage of the extracellular domain of the seven transmembrane domain thrombin receptor (PAR1) was determined using site-directed mutagenesis. Basic residues in anion binding exosite-I (Arg35, Arg36, Arg67, Arg73, Arg75, Arg77A, Lys81, Lys109, Lys110 and Lys149E) were substituted with glutamines and the resultant recombinant mutant thrombins were used to determine kinetic parameters for the cleavage of a peptide (PAR38-60) based on the PAR1 extracellular domain. Compared with wild-type thrombin, replacement of Arg67 and Arg73 had a dramatic effect on the cleavage of PAR38-60 (k(cat)/K(m) = 1.8 x 10(6) and 4.6 x 10(6) vs 9.2 x 10(7) M(-1).s(-1)), whereas the remaining mutations of the anion-binding exosite I of thrombin had a less pronounced effect, with k(cat)/K(m) values ranging from 3.3 x 10(7) M(-1). s(-1) (R77(a)Q) to 5.8 x 10(7) M(-1).s(-1) (K109Q). The ability of thrombin mutants to activate platelets paralleled that of PAR38-60 cleavage, whereas their ability to clot fibrinogen differed profoundly, as did their susceptibility to hirudin inhibition. Results are interpreted with respect to known interactions of thrombin with thrombomodulin, hirudin, rhodniin and heparin cofactor II. We conclude that the basic residues of anion-binding exosite I contribute significantly to enhancing the rate of complex formation in two ways; the first (general) ensures electrostatic steering of ligands with complementary electrostatic fields, the second (specific) involves a combination of molecular contacts within the complex that is unique for each ligand. PMID- 11121105 TI - Engineering the active site of ascorbate peroxidase. AB - The oxidation of a number of thioethers, namely methyl phenyl sulphide (1), ethyl phenyl sulphide (2), isopropyl phenyl sulphide (3), n-propyl phenyl sulphide (4), p-chlorophenyl methyl sulphide (5), p-nitrophenyl methyl sulphide (6) and methyl naphthalene sulphide (7), by recombinant pea cytosolic ascorbate peroxidase (rAPX) and a site-directed variant of rAPX in which the distal tryptophan 41 residue has been replaced with an alanine (W41A) has been examined. The electronic spectrum (pH 7.0, mu = 0.10 M, 25.0 degrees C) for the ferric derivative of W41A (lambda(max)/nm = 411, 534, 560, 632) is indicative of an increased quantity of 6-coordinate, high-spin and/or 6-coordinate, low-spin haem compared to rAPX. Steady state oxidation of sulphides 1-4 and 7, gave values for kcat that are approximately 10-fold and 100-fold, respectively, higher for W41A than for rAPX. For rAPX, essentially racemic mixtures of R- and S-sulphoxides were obtained for all sulphides. With the exception of sulphide 7, the W41A variant shows substantial enhancements in enantioselectivity, with R : S ratios varying between R : S = 63 : 37 (sulphides 1 and 4) and R : S = 85 : 15 (sulphide 6). Incubation of sulphide 2 with rAPX or W41A and [(18)O] H(2)O(2) shows 95% (rAPX) and 96% (W41A) transfer of labelled oxygen to the substrate. Structure based modelling techniques have provided a fully quantitative rationalization of all the experimentally determined R : S ratios and have indicated that reorientation of the sidechain of Arg38, such that access to the haem is much less restricted, is influential in controlling the stereoselectivity for both rAPX and W41A. PMID- 11121106 TI - Scots pine expresses short-root-specific peroxidases during development. AB - Nine short-root-specific proteins from Scots pine (Pinus sylvestris L.) detected and isolated as individual spots by 2D-PAGE were identified. The similar peptide mass maps obtained for all nine polypeptide spots together with lectin-blotting results suggest that they represent forms of the same modified protein. N Terminal sequence analysis of two of the peptides showed high similarity to peroxidases. RT-PCR with oligonucleotide primers corresponding to determined peptide sequences and conserved regions in plant peroxidases led to isolation of Psyp1 cDNA which is most abundantly expressed in short roots. Psyp1 is the first peroxidase cDNA to be isolated from the genus Pinus. It encodes a 363-amino-acid class-III peroxidase with a calculated molecular mass of 35.7 kDa and theoretical pI of 4.74. The predicted PSYP1 amino-acid sequence is grouped with other class III peroxidases in phylogenetic analyses, but it has a unique amino-acid sequence which may be associated with its function in short roots or with its phylogenetic group. The presence of a signal sequence for extracellular transport indicates that PSYP1 belongs to the group of secreted class-III peroxidases. The presence of 10 tyrosine residues and putative auxin-binding regions in PSYP1 suggests that the function of the enzyme is associated with cell-wall formation in short roots. The downregulation of Psyp1 expression in symbiotic short roots hosting the ectomycorrhizal fungus Suillus bovinus is perhaps related to the change in cell wall structure necessary for ectomycorrhizal development. PMID- 11121107 TI - Proteolytic degradation of ribonuclease A in the pretransition region of thermally and urea-induced unfolding. AB - The method of limited proteolysis has proven to be appropriate for the determination of unfolding rate constants (k(U)) of ribonuclease A in the transition region of thermal denaturation [Arnold, U. & Ulbrich-Hofmann, R. (1997) Biochemistry 36, 2166-2172]. The aim of the present paper was to extend this procedure to the pretransition region of thermally and urea-induced denaturation where spectroscopic methods do not allow direct measurement of k(U). The results show that the approach can be applied successfully to denaturing (free energy of unfolding Delta G < 10 kJ.mol(-1)) and to marginally native conditions (Delta G = 10-25 kJ.mol(-1)). Under moderately (Delta G = 25-30 kJ.mol(-1)) and strongly native conditions (Delta G > 30 kJ.mol(-1)), however, the determination of kU was not possible in this way as the proteolytic degradation of ribonuclease A by thermolysin or trypsin was no longer determined by global unfolding. Here, proteolysis proceeds via the native RNase A. In the presence of low concentrations of urea, the rate constants of proteolysis were, surprisingly, smaller than in the absence of urea. As the protease activity has been taken into account, this result points to a local stabilization of the RNase A molecule. PMID- 11121108 TI - Mutational analysis of the active site of human insulin-regulated aminopeptidase. AB - Insulin-regulated aminopeptidase (IRAP) is a type II integral membrane protein belonging to the gluzincin family of metallopeptidases identified by the characteristic Zn(2+)-coordination sequence element, HEXXH-(18-64X)-E. A second conserved sequence element, the GXMEN motif, positioned 22-32 amino acids N terminal to the Zn(2+)-coordination sequence element distinguishes the gluzincin aminopeptidases from other gluzincins. To investigate the importance of the G428AMEN and H464ELAH-(18X)-E487 motifs for the activity of IRAP, mutational analysis was carried out. cDNA encoding the full-length transmembrane form of human IRAP was expressed in HEK293 cells and recombinant wild-type IRAP was shown to have biochemical and enzymatic properties similar to those reported for native IRAP and the soluble serum form of IRAP. Mutational analysis using single amino acid substitutions in the GAMEN motif (G428A, A429G, M430K, M430E, M430I, E431D and E431A) and in the Zn(2+)-binding motif (H464Y, E465D, E465Q, H468Y, E487D and E487Q) resulted in decreased or abolished aminopeptidase activity towards the leucine-para-nitroanilide substrate. The results show that conservation of residues within the GAMEN and Zn(2+)-binding motifs is important for IRAP enzyme activity. PMID- 11121109 TI - An isoform-specific interaction of the membrane anchors affects mammalian adenylyl cyclase type V activity. AB - The nine membrane-bound mammalian adenylyl cyclases (ACs) contain two highly diverged membrane anchors, M1 and M2, with six transmembrane spans each and two conserved cytosolic domains which coalesce into a pseudoheterodimeric catalytic unit. Previously, the catalytic segments, bacterially expressed as soluble proteins, were characterized extensively whereas the function of the membrane domains remained unexplored. Using the catalytic C1 and C2 domains of AC type V we employed the membrane anchors from type V and VII ACs for construction of enzymes with duplicated, inverted, fully swapped and chimeric membrane anchors. Further, in the M1 membrane domain individual transmembrane spans were removed or exchanged between type V and VII ACs. The constructs were expressed in HEK293 cells, the expression levels and membrane localization was assessed by Western blotting. Cell-free basal, forskolin-, GTP gamma S-and G(s alpha)/GTP gamma S stimulated AC activities were determined. The results demonstrate that enzymatic activities were only maintained when the M1 and M2 membrane domains were derived from either AC V or VII. Constructs with chimeric membrane domains, i.e. M1 from type V and M2 from type VII AC or vice versa, were essentially inactive although the expression levels and membrane localization appeared to be normal. The data indicate a functionally important interaction of the membrane domains of ACs in that they seem to interact in a pair-like, isoform delimited manner. This interaction directly impinges on the formation of the catalytic interface. We propose that protein-protein interactions of the AC membrane domains may constitute another, yet unexplored level of AC regulation. PMID- 11121110 TI - Thermodynamic analysis of saccharide binding to snake gourd (Trichosanthes anguina) seed lectin. Fluorescence and absorption spectroscopic studies. AB - The interaction of different saccharides with the snake gourd (Trichosanthes anguina) seed lectin (SGSL) was investigated by fluorescence spectroscopy. Binding of 4-methylumbelliferyl-beta-D-galactopyranoside (MeUmb beta Gal) to SGSL resulted in a significant increase in the fluorescence emission intensity of the sugar at 376 nm, and this change was used to estimate the association constants for the binding interaction. Interestingly, the increase in emission intensity changed with a change in temperature, increasing from 19.2% at 20 degrees C to 80.2% at 40 degrees C. At 20 degrees C the association constant, K(a), for the MeUmb beta Gal-SGSL interaction was found by fluorescence titration to be 5.8 x 10(4) M(-1). From the temperature dependence of the association constants, the changes in enthalpy (Delta H) and entropy (Delta S) associated with binding of MeUmb beta Gal to SGSL were estimated to be -80.85 kJ.mol(-1) and -184.0 J.mol( 1).K(-1), respectively. Binding of unlabeled sugars was investigated by monitoring the decrease in fluorescence intensity when they were added to a mixture of SGSL and MeUmb beta Gal. The Ka values for different sugars were determined at several temperatures, and Delta H and Delta S were determined from the van't Hoff plots. Enthalpy-entropy compensation was noticed in all cases. The results indicate that saccharide binding to SGSL is enthalpy-driven and the negative contribution from entropy is, in general, quite high. PMID- 11121111 TI - Thermally induced conformational changes in horseradish peroxidase. AB - Detailed differential scanning calorimetry (DSC), steady-state tryptophan fluorescence and far-UV and visible CD studies, together with enzymatic assays, were carried out to monitor the thermal denaturation of horseradish peroxidase isoenzyme c (HRPc) at pH 3.0. The spectral parameters were complementary to the highly sensitive but integral method of DSC. Thus, changes in far-UV CD corresponded to changes in the overall secondary structure of the enzyme, while that in the Soret region, as well as changes in intrinsic tryptophan fluorescence emission, corresponded to changes in the tertiary structure of the enzyme. The results, supported by data about changes in enzymatic activity with temperature, show that thermally induced transitions for peroxidase are irreversible and strongly dependent upon the scan rate, suggesting that denaturation is under kinetic control. It is shown that the process of HRPc denaturation can be interpreted with sufficient accuracy in terms of the simple kinetic scheme N -->k D where k is a first-order kinetic constant that changes with temperature, as given by the Arrhenius equation; N is the native state, and D is the denatured state. On the basis of this model, the parameters of the Arrhenius equation were calculated. PMID- 11121112 TI - Increasing the thermal stability of euphauserase. A cold-active and multifunctional serine protease from Antarctic krill. AB - A molecular model of Antarctic krill euphauserase based on the known crystal structure of its fiddler crab analog, collagenase I, indicates that the core structure of these enzymes is almost identical. Euphauserase is a cold-active and thermally sensitive enzyme with a high affinity for Lys, Arg and large hydrophobic amino acids. Residue Phe137 in euphauserase, localized in loop D (autolysis loop), is highly exposed on the surface of the molecule. Therefore, it appeared to be an easy target for autolysis. The broadly specific euphauserase has a low affinity for negatively charged residues. In order to increase the stability of the enzyme, two mutants were created in which residue Phe137 was replaced by a Glu and an Asp residue. Both mutations resulted in increased stability of the recombinant euphauserase towards thermal inactivation. PMID- 11121113 TI - Sulfoxidation mechanism of vanadium bromoperoxidase from Ascophyllum nodosum. Evidence for direct oxygen transfer catalysis. AB - We have previously shown that vanadium bromoperoxidase from Ascophyllum nodosum mediates production of the (R)-enantiomer of methyl phenyl sulfoxide with 91% enantiomeric excess. Investigation of the intrinsic selectivity of vanadium bromoperoxidase reveals that the enzyme catalyzes the sulfoxidation of methyl phenyl sulfide in a purely enantioselective manner. The K(m) of the enzyme for methyl phenyl sulfide was determined to be approximately 3.5 mM in the presence of 25% methanol or tert-butanol. The selectivity of the sulfoxidation of methyl phenyl sulfide is optimal in the temperature range 25-30 degrees C and can be further optimized by increasing the enzyme concentration, yielding selectivities with up to 96% enantiomeric excess. Furthermore, we established for the first time that vanadium bromoperoxidase is functional at temperatures up to 70 degrees C. A detailed investigation of the sulfoxidation activity of this enzyme using (18)O-labeled hydrogen peroxide shows that vanadium bromoperoxidase mediates the direct transfer of the peroxide oxygen to the sulfide. A schematic model of the vanadium haloperoxidase sulfoxidation mechanism is presented. PMID- 11121114 TI - A chicken hnRNP of the A/B family recognizes the single-stranded d(CCCTAA)(n) telomeric repeated motif. AB - With the aim of identifying proteins able to interact with the C-rich single stranded telomeric repeated motif, three nuclear polypeptides, CBNP alpha, CBNP beta and CBNP gamma, with apparent mobilities in SDS/PAGE of 38, 44 and 55 kDa, respectively, were isolated from mature chicken erythrocytes by affinity chromatography. In situ UV-cross-linking experiments demonstrated that CBNP alpha and CBNP gamma interact directly with the telomeric d(CCCTAA)n repeat, whereas CBNP beta does not. Moreover, they provided information on the protein components responsible for each electrophoretic mobility-shift assay signal. Ion spray and matrix-assisted laser desorption ionization MS allowed us to identify CBNP alpha with single-stranded D-box-binding factor (ssDBF), a protein previously characterized as a transcription factor belonging to the A/B family of heterogeneous nuclear ribonucleoproteins, and CBNP beta with an isoform of this protein containing an extra exon. Similarly, CBNP gamma was shown to be probably the chicken homolog of hnRNP K, a ribonuclear protein able to bind to polyC oligonucleotides. The relation of CBNP alpha (i.e. ssDBF), CBNP beta and CBNP gamma to a number of similar proteins in the protein and nucleotide sequence databank is discussed. A rather diversified spectrum of functional roles has been assigned to some of these proteins despite the strong sequence homology among them. PMID- 11121115 TI - Pyrokinin neuropeptides in a crustacean. Isolation and identification in the white shrimp Penaeus vannamei. AB - Identification of substances able to elicit physiological or behavioural processes that are related to reproduction would greatly contribute to the domestication of commercially important crustaceans that do not reproduce easily in captivity. Crustaceans are thought to release urine signals used for chemical communication involved in courtship behaviour. In contrast to insects, very little is known about the endocrinological processes underlying this phenomenon. Therefore, an extract of 3500 central nervous systems of female white shrimp Penaeus vannamei was screened for myotropic activity in order to purify pyrokinin like peptides that belong to the pyrokinin/PBAN neuropeptide family. Members of this family regulate reproductive processes in insects, including pheromone biosynthesis. Purification of these pyrokinins was achieved by a combination of reversed-phase and normal-phase chromatography. Subsequent characterization by mass spectrometry, Edman degradation and peptide synthesis resulted in the elucidation of two novel peptides. Pev-PK 1 has the primary sequence DFAFSPRL NH(2) and a second peptide (Pev-PK 2) is characterized as the nonapeptide ADFAFNPRL-NH(2). Pev-PK 1 contains the typical FXPRL-NH(2) (X = G, S, T or V) C terminal sequence that characterizes members of the versatile pyrokinin/PBAN family. Pev-PK 2 displays an Asn residue at the variable X position of the core pyrokinin sequence. These crustacean pyrokinins are the first to be found in a noninsect. The synthetic peptides display myotropic activity on the Leucophaea maderae as well as on the Astacus leptodactylus hindgut. PMID- 11121116 TI - The stability and folding process of amyloidogenic mutant human lysozymes. AB - Amyloid deposits are frequently formed by mutant proteins that have a lower stability than the wild-type proteins. Some reports, however, have shown that mutant-induced thermodynamic destabilization is not always a general mechanism of amyloid formation. To obtain a better understanding of the mechanism of amyloid fibril formation, we show in this study that equilibrium and kinetic refolding unfolding reaction experiments with two amyloidogenic mutant human lysozymes (I56T and D67H) yield folding pathways that can be drawn as Gibbs energy diagrams. The equilibrium stabilities between the native and denatured states of both mutant proteins were decreased, but the degrees of instability were different. The Gibbs energy diagrams of the folding process reveal that the Gibbs energy change between the native and folding intermediate states was similar for both proteins, and also that the activation Gibbs energy change from the native state to the transition state decreased. Our results confirm that the tendency to favor the intermediate of denaturation facilitates amyloid formation by the mutant human lysozymes more than equilibrium destabilization between the native and completely denatured states does. PMID- 11121117 TI - Soluble gp130 is the natural inhibitor of soluble interleukin-6 receptor transsignaling responses. AB - Signal transduction in response to interleukin-6 (IL-6) requires binding of the cytokine to its receptor (IL-6R) and subsequent homodimerization of the signal transducer gp130. The complex of IL-6 and soluble IL-6R (sIL-6R) triggers dimerization of gp130 and induces responses on cells that do not express membrane bound IL-6R. Naturally occurring soluble gp130 (sgp130) can be found in a ternary complex with IL-6 and sIL-6R. We created recombinant sgp130 proteins that showed binding to IL-6 in complex with sIL-6R and inhibited IL-6/sIL-6R induced proliferation of BAF/3 cells expressing gp130. Surprisingly, sgp130 proteins did not affect IL-6 stimulated proliferation of BAF/3 cells expressing gp130 and membrane bound IL-6R, indicating that sgp130 did not interfere with IL-6 bound to IL-6R on the cell surface. Additionally, sgp130 partially inhibited proliferation induced by leukemia inhibitory factor (LIF) and oncostatin M (OSM) albeit at higher concentrations. Recombinant sgp130 protein could be used to block the anti apoptotic effect of sIL-6R on lamina propria cells from Crohn disease patients. We conclude that sgp130 is the natural inhibitor of IL-6 responses dependent on sIL-6R. Furthermore, recombinant sgp130 is expected to be a valuable therapeutic tool to specifically block disease states in which sIL-6R transsignaling responses exist, e.g. in morbus Crohn disease. PMID- 11121118 TI - Genomic organization of the human phosphodiesterase PDE11A gene. Evolutionary relatedness with other PDEs containing GAF domains. AB - PDE11A is a dual-substrate, cAMP and cGMP, cyclic nucleotide phosphodiesterase (PDE). Presently four unique variants carrying distinct GAF sequences in the N terminal region have been identified. While human PDE11A3 and PDE11A4 are known to be specifically expressed in testis and prostate, respectively, PDE11A1 was mainly detected in skeletal muscle. The human PDE11A gene was investigated and revealed to span > 300 kb, contain 23 exons and be mapped on chromosome 2q31. The transcription start sites of PDE11A1, PDE11A3 and PDE11A4 were determined, and the promoter sequences were revealed. Although 5' flanking genomic regions of PDE11A1 and PDE11A3 had a consensus TATA motif, that of PDE11A4 was a TATA-less but contained CCAAT box and Sp1-binding sequence. Interestingly, we found that the exon 2 sequence for N-terminal region of PDE11A3 encoded an N-terminal sequence of the cytochrome c pseudogene in an alternate reading frame, and that C terminal region of the cytochrome c pseudogene in intron 2 was disrupted by the insertion of Alu repetitive sequence. Furthermore, we examined the exon-intron organization of the PDE2A gene and compared the exon organization among GAF-PDE family. The exon organization of the PDE11A catalytic domain was very similar to those of PDE5A and PDE6B. However, other GAF-PDEs, PDE2A and PDE10A, displayed different exon organization from PDE11A although these three PDEs are similar in their amino-acid sequences to each other. The findings suggested that PDE11A has a common ancestral gene with PDE5A and PDE6s, whereas PDE2A and PDE10A are generated separately from these three GAF-PDEs. PMID- 11121119 TI - Additional PKA phosphorylation sites in human cardiac troponin I. AB - We used mass spectrometry to monitor cAMP-dependent protein kinase catalysed phosphorylation of human cardiac troponin I in vitro. Phosphorylation of isolated troponin I by cAMP-dependent protein kinase resulted in the covalent incorporation of phosphate on at least five different sites on troponin I, and a S22/23A troponin I mutant incorporated phosphates on at least three sites. In addition to the established phosphorylation sites (S22 and S23) we found that S38 and S165 were the other two main sites of phosphorylation. These 'overphosphorylation' sites were not phosphorylated sufficiently slower than S22 and S23 that we could isolate pure S22/23 bisphosphorylated troponin I. Overphosphorylation of troponin I reduced its affinity for troponin C, as measured by isothermal titration microcalorimetry. Phosphorylation of S22/23A also decreased its affinity for troponin C indicating that phosphorylation of S38 and/or S165 impedes binding of troponin I to troponin C. Formation of a troponin I/troponin C complex prior to cAMP-dependent protein kinase treatment did not prevent overphosphorylation. When whole troponin was phosphorylated by cAMP dependent protein kinase, however, [(32)P]phosphate was incorporated only into troponin I and only at S22 and S23. Mass spectrometry confirmed that overphosphorylation is abolished in the ternary complex. Troponin I bisphosphorylated exclusively at S22 and S23 troponin I showed reduced affinity for troponin C but the effect was diminished with respect to overphosphorylated troponin I. These results show that care should be exercised when interpreting data obtained with troponin I phosphorylated in vitro. PMID- 11121120 TI - A folding variant of human alpha-lactalbumin induces mitochondrial permeability transition in isolated mitochondria. AB - A human milk fraction containing multimeric alpha-lactalbumin (MAL) is able to kill cells via apoptosis. MAL is a protein complex of a folding variant of alpha lactalbumin and lipids. Previous results have shown that upon treatment of transformed cells, MAL localizes to the mitochondria and cytochrome c is released into the cytosol. This is followed by activation of the caspase cascade. In this study, we further investigated the involvement of mitochondria in apoptosis induced by the folding variant of alpha-lactalbumin. Addition of MAL to isolated rat liver mitochondria induced a loss of the mitochondrial membrane potential (Delta Psi(m)), mitochondrial swelling and the release of cytochrome c. These changes were Ca(2+)-dependent and were prevented by cyclosporin A, an inhibitor of mitochondrial permeability transition. MAL also increased the rate of state 4 respiration in isolated mitochondria by exerting an uncoupling effect. This effect was due to the presence of fatty acids in the MAL complex because it was abolished completely by BSA. BSA delayed, but failed to prevent, mitochondrial swelling as well as dissipation of Delta Psi(m), indicating that the fatty acid content of MAL facilitated, rather than caused, these effects. Similar results were obtained with HAMLET (human alpha-lactalbumin made lethal to tumour cells), which is native alpha-lactalbumin converted in vitro to the apoptosis-inducing folding variant of the protein in complex with oleic acid. Our findings demonstrate that a folding variant of alpha-lactalbumin induces mitochondrial permeability transition with subsequent cytochrome c release, which in transformed cells may lead to activation of the caspase cascade and apoptotic death. PMID- 11121122 TI - Anti-inflammatory drugs block cytokine mRNA accumulation in the skin and improve the clinical condition of reactional leprosy patients. AB - The aim of this study was to investigate in what ways in vivo anti-inflammatory treatment affects cytokine mRNA expression in situ in both erythema nodosum leprosum and reversal reaction patients. Serial biopsies were collected from the patients undergoing leprosy reactions before and during pentoxifylline (n = 7) or thalidomide (n = 3) treatment for erythema nodosum leprosum and prednisone (n = 3) for reversal reaction. Clinical evolution of the skin lesion was assessed during the study and semiquantitative reverse transcription-polymerase chain reaction was used to investigate cytokine mRNA expression at the lesion site. Results showed expression of interferon-gamma, interleukin-6, interleukin-10, interleukin-12 p40, and tumor necrosis factor-alpha in all patients tested at the onset of reactional episodes, but interleukin-4 mRNA was rarely detected in the lesions (n = 4). Follow-up analysis showed that, irrespective of the drugs used, tumor necrosis factor-alpha mRNA was diminished in 10 of the 13 patients tested. A concomitant decrease of mRNA accumulation was also observed for interferon gamma (nine of 11 patients), interleukin-6 (nine of 11), and interleukin-12 p40 (six of eight). An inhibitory effect on interleukin-10 mRNA was likewise seen after thalidomide and pentoxifylline, but not subsequent to prednisone treatment. The data also demonstrated that cytokine mRNA inhibition correlates to the resolution of the inflammatory response in situ (n = 10), whereas the persistence/enhancement of cytokine message expression after treatment was associated with worsening of the skin condition, as seen in three erythema nodosum leprosum patients whose maintenance of local inflammation was accompanied by the appearance/persistence of interleukin-4 gene expression in situ subsequent to anti-inflammatory treatment. In summary, the participation of cytokines in leprosy inflammatory episodes seems to be directly associated with the patients' clinical evolution following therapy for reaction. PMID- 11121123 TI - Hapten-specific tolerance promoted by calcitonin gene-related peptide. AB - Calcitonin gene-related peptide has been shown to modulate inflammatory and immune responses in various systems. Recent studies in our laboratory and colleagues have shown that intracutaneously injected calcitonin gene-related peptide impairs the induction of contact hypersensitivity in mice, and participates in the pathogenesis of failed contact hypersensitivity induction after acute, low-dose ultraviolet B radiation. In this study we investigated the ability of calcitonin gene-related peptide to induce tolerance in normal and mast cell deficient mice and we examined the extent to which calcitonin gene-related peptide contributes to the tolerance induced by acute, low-dose ultraviolet B radiation. Calcitonin gene-related peptide was injected intradermally followed by application of 2,4-dinitro-1-fluorobenzene to the injected skin surface. Tolerance was assessed by re-exposing the mice 2 wk later to a second, sensitizing dose of 2, 4-dinitro-1-fluorobenzene on uninjected skin. We found that calcitonin gene-related peptide induced tolerance to 2, 4-dinitro-1 fluorobenzene in both normal and mast cell deficient mice. Calcitonin gene related peptide-induced tolerance was blocked by intradermal injection of a calcitonin gene-related peptide antagonist [CGRP-(8-37)] that selectively blocks the calcitonin gene-related peptide receptor. Tolerance was also abolished by intraperitoneally injected anti-interleukin-10, but not anti-tumor necrosis factor alpha, antibodies. When 2,4-dinitro-1-fluorobenzene was painted on skin into which splenic dendritic cells pretreated with calcitonin gene-related peptide had been injected, tolerance was observed. Calcitonin gene-related peptide- treated dendritic cells mixed with anti-interleukin-10 antibody prior to intradermal injection failed to promote tolerance. Finally, injection of CGRP-(8 37) into skin that was subsequently exposed to acute, low-dose ultraviolet B radiation partially prevented tolerance induced by local application of 2,4 dinitro-1-fluorobenzene. These results indicate that calcitonin gene-related peptide has the capacity to promote cutaneous tolerance through an interleukin-10 dependent mechanism. This mechanism, which does not require the participation of mast cells, contributes to the tolerance promoted by acute, low-dose ultraviolet B radiation. Thus, calcitonin gene-related peptide from cutaneous nerve endings plays a key role in the local immune aberrations caused by ultraviolet B radiation. PMID- 11121124 TI - Characterization of glucose transport system in keratinocytes: insulin and IGF-1 differentially affect specific transporters. AB - Skin is one of the major tissues displaying chronic diabetic complications. We have studied glucose transport following stimulation with insulin and IGF-1 in cultured mouse keratinocytes. In proliferating cells, acute stimulation with insulin and IGF-1 increased glucose uptake. Insulin translocated glucose transporters 1 and 5, whereas IGF-1 translocated glucose transporters 2 and 3. With differentiation, glucose transporter 3 expression increased and the expression of glucose transporters 1, 2, and 5 decreased. No increase in glucose uptake was observed, however, following stimulation with either hormone. These results indicate that insulin and IGF-1 differentially regulate glucose uptake as well as expression and translocation of specific transporters in skin keratinocytes. PMID- 11121125 TI - Epitopes targeted by bullous pemphigoid T lymphocytes and autoantibodies map to the same sites on the bullous pemphigoid 180 ectodomain. AB - Bullous pemphigoid is a blistering skin disease characterized by autoantibodies directed against the NC16A domain of bullous pemphigoid 180 (collagen XVII), a transmembrane protein of epidermal basal cells. Passive transfer studies in mice have shown that antibodies that bind to this immunodominant region of bullous pemphigoid 180 are capable of inducing a skin disease that closely mimics bullous pemphigoid, supporting the hypothesis that epitopes within NC16A are involved in the pathogenesis of bullous pemphigoid. In this study, we examined the autoimmune T cell response in bullous pemphigoid patients. T cells from eight of 12 bullous pemphigoid patients, all of whom had circulating anti-bullous pemphigoid 180 autoantibodies, showed a specific proliferative response to recombinant forms of NC16A. T cell lines and clones developed from four of these patients recognize the same NC16A peptides as those targeted by autoantibodies from the corresponding individuals. These NC16A-responding T lymphocytes express alpha/beta T cell receptors and CD4 memory T cell surface markers and exhibited a Th1/Th2 mixed cytokine profile that may support the production of antibodies. This new information will aid in defining the key steps involved in the development of the autoimmune response in bullous pemphigoid. PMID- 11121126 TI - Expression of glycosaminoglycans and small proteoglycans in wounds: modulation by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu(2+). AB - Glycyl-histidyl-lysine-Cu(2+) is a tripeptide-copper complex previously shown to be an activator of wound healing. We have investigated the effects of glycyl histidyl-lysine-Cu(2+) on the synthesis of glycosaminoglycans and small proteoglycans in a model of rat experimental wounds and in rat dermal fibroblast cultures. Repeated injections of glycyl-histidyl-lysine-Cu(2+) (2 mg per injection) stimulated the wound tissue production, as appreciated by dry weight and total protein measurements. This stimulation was accompanied by an increased production of type I collagen and glycosaminoglycans (assessed, respectively, by hydroxyproline and uronic acid contents of the chamber). Electrophoretic analysis of wound tissue glycosaminoglycans showed an accumulation of chondroitin sulfate and dermatan sulfate in control wound chambers, whereas the proportion of hyaluronic acid decreased with time. The accumulation of chondroitin sulfate and dermatan sulfate was enhanced by glycyl-histidyl-lysine-Cu(2+) treatment. The expression of two small proteoglycans of the dermis, decorin and biglycan, was analyzed by northern blot. The biglycan mRNA steady-state level in the chamber was maximal at day 12, whereas the decorin mRNA increased progressively until the end of the experiment (day 22). Glycyl-histidyl-lysine-Cu(2+) treatment increased the mRNA level of decorin and decreased those of biglycan. In dermal fibroblast cultures, the stimulation of decorin expression by glycyl-histidyl-lysine-Cu(2+) was also found. In contrast, biglycan expression was not modified. These results show that the expression of different proteoglycans in wound tissue are regulated in a different manner during wound healing. The glycyl-histidyl-lysine-Cu(2+) complex is able to modulate the expression of the extracellular matrix macromolecules differently during the wound repair process. PMID- 11121127 TI - IgG anti-melanocyte antibodies purified from patients with active vitiligo induce HLA-DR and intercellular adhesion molecule-1 expression and an increase in interleukin-8 release by melanocytes. AB - An immunologic hypothesis is currently proposed as a possible pathogenesis of nonsegmental-type vitiligo. IgG antibodies against melanocyte surface antigens exist in the serum of patients with vitiligo vulgaris. IgG anti-melanocyte antibodies were reported to induce melanocyte damage in vitro by a complement mediated mechanism and antibody-dependent cellular cytotoxicity. Perilesional melanocytes express major histocompatibility complex class II antigens and a higher intercellular adhesion molecule-1 compared with those in normal skin. The purpose of this study was to determine the role of IgG anti-melanocyte antibodies in the inappropriate expression of major histocompatibility complex class II antigens and intercellular adhesion molecule-1 on melanocytes. IgG anti melanocyte antibody samples were purified from the individual serum of patients with active vitiligo. After incubation of IgG anti-melanocyte antibodies with cultured melanocytes, the results revealed: (i) IgG anti-melanocyte antibody stimulated HLA-DR expression on melanocytes; (ii) intercellular adhesion molecule 1 expression on melanocytes was significantly induced by IgG anti-melanocyte antibodies; and (iii) IgG anti-melanocyte antibodies induced an increase in interleukin-8 release from melanocytes. The major histocompatibility complex class II molecules expressed in melanocytes can present antigens to CD4 helper cells as antigen-presenting cells and elicit an immune response. Intercellular adhesion molecule-1 is an important adhesion molecule involved in leukocyte and parenchymal cell interaction and thus plays an essential part in immunologic and inflammatory reactions. It is reasonable to speculate that abnormal expressions of HLA-DR and intercellular adhesion molecule-1 on melanocytes by IgG anti melanocyte antibodies would present vitiligo antigens and allow the antigen specific immune effector cell attack that results in melanocytotoxicity. PMID- 11121128 TI - Clinical, cellular, and molecular features of an Israeli xeroderma pigmentosum family with a frameshift mutation in the XPC gene: sun protection prolongs life. AB - An Ashkenazi Jewish Israeli family with two children affected with severe xeroderma pigmentosum was investigated. A son, XP12TA, developed skin cancer at 2 y and died at 10 y. A daughter, XP25TA, now 24 y old, was sun protected and began developing skin cancers at 10 y. Their cultured skin fibroblasts showed reductions in post-ultraviolet survival (11% of normal), unscheduled DNA synthesis (10% of normal), global genome DNA repair (15% of normal), and plasmid host cell reactivation (5% of normal). Transcription-coupled DNA repair was normal, however. Northern blot analysis revealed greatly reduced xeroderma pigmentosum complementation group C mRNA. A plasmid host cell reactivation assay assigned the cells to xeroderma pigmentosum complementation group C. Cells from both parents and an unaffected child exhibited normal post-ultraviolet-C survival and normal DNA repair. Sequencing the xeroderma pigmentosum complementation group C cDNA of XP12TA and XP25TA revealed a homozygous deletion of two bases (del AT 669-670) in exon 5 with a new termination site 10 codons downstream that is expected to encode a truncated xeroderma pigmentosum complementation group C protein. Sequence analysis of the xeroderma pigmentosum complementation group C cDNA in cells from the parents found identical heterozygous mutations: one allele carries both the exon 5 frameshift and an exon 15 polymorphism and the other allele carries neither alteration. Cells from the unaffected brother had two normal xeroderma pigmentosum complementation group C alleles. This frameshift mutation in the xeroderma pigmentosum complementation group C gene led to reduced DNA repair with multiple skin cancers and early death. Sun protection delayed the onset of skin cancer and prolonged life in a sibling with the same mutation. PMID- 11121129 TI - Xeroderma pigmentosum variant heterozygotes show reduced levels of recovery of replicative DNA synthesis in the presence of caffeine after ultraviolet irradiation. AB - Patients with xeroderma pigmentosum variant show clinical photosensitivity, skin neoplasias induced by ultraviolet light, and defective postreplication repair, but normal nucleotide excision repair. We recently reported an alternative, simple method for the diagnosis of xeroderma pigmentosum variant that measures by autoradiography three cellular markers for DNA repair after ultraviolet irradiation: unscheduled DNA synthesis, recovery of RNA synthesis, and recovery of replicative DNA synthesis. Among hereditary photosensitive disorders, including other xeroderma pigmentosum groups, Cockayne syndrome, and a newly established ultraviolet-sensitive syndrome, only xeroderma pigmentosum variant cells exhibited normal unscheduled DNA synthesis, normal recovery of RNA synthesis, but reduced recovery of replicative DNA synthesis (51 +/- 6% the rate relative to normal controls). This reduction of recovery of replicative DNA synthesis was enhanced in the presence of a nontoxic level of caffeine to 36 +/- 5%. In this study we assess the cellular markers in two independent families that included two photosensitive patients that were identified as xeroderma pigmentosum variant. Cells from heterozygotic parents showed normal levels of unscheduled DNA synthesis, recovery of RNA synthesis, and recovery of replicative DNA synthesis, but reduced rates of recovery of replicative DNA synthesis in the presence of 1 mM caffeine (53 +/- 8% relative to the normal control). Furthermore, with a colony-forming assay, the cells showed normal survival by ultraviolet without caffeine, but slightly reduced survival by ultraviolet with 1 mM caffeine present. In one family, we confirmed inheritance of two heterozygous mis-sense mutations. One mutation is an A-->G transition at nucleotide 1840 that generates a K535E mis-sense mutation. Another mutation is an A-->C transversion at nucleotide 2003 that generates a K589 mis-sense mutation. Each of these mutations were absent in 52 unrelated Japanese individuals. These results suggest that xeroderma pigmentosum variant heterozygotes can be identified by their sensitivity to ultraviolet irradiation in the presence of nontoxic levels of caffeine. PMID- 11121130 TI - Suppressive effects of cyclosporin A and FK-506 on superoxide generation in human polymorphonuclear leukocytes primed by tumor necrosis factor alpha. AB - Most previous studies have found no effects of cyclosporin A and FK-506 on active oxygen generation in human polymorphonuclear leukocytes. Recently various differences in biologic properties have been reported between unprimed peripheral blood human polymorphonuclear leukocytes and tissue or primed human polymorphonuclear leukocytes. In this study, we investigated the effects of cyclosporin A and FK-506 on superoxide (O(2)(-)) generation induced by the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine in human peripheral blood polymorphonuclear leukocytes primed or unprimed with tumor necrosis factor alpha. Neither cyclosporin A nor FK-506 suppressed N-formyl-L methionyl-L-leucyl-L-phenylalanine-induced O(2)(-) generation in unprimed human polymorphonuclear leukocytes at concentrations between 0.1 nM and 10 microM, as in previous studies. Only at 1 microM of cyclosporin A and 100 nM of FK-506 were marginal suppressive effects observed. On the other hand, cyclosporin A and FK 506 both suppressed N-formyl-L-methionyl-L-leucyl-L-phenylalanine-induced O(2)(-) generation in tumor-necrosis-factor-alpha-primed human polymorphonuclear leukocytes, strongly and dose dependently, at concentrations between 1 nM and 1 microM. Neither cyclosporin A nor FK-506 influenced tyrosyl phosphorylation of 115 kDa protein, which is inducible during the priming process, suggesting that neither cyclosporin A nor FK-506 influenced the tumor-necrosis-factor-alpha induced priming process itself, and instead modified the biologic response of primed human polymorphonuclear leukocytes. PMID- 11121131 TI - Concordance of dermatitis herpetiformis and celiac disease in monozygous twins. AB - Celiac disease can be defined as the classical manifestation of gluten sensitivity, which primarily affects the small intestine. Gluten sensitivity has also a skin manifestation, i.e., dermatitis herpetiformis. Both diseases have a strong genetic association with HLA DQ on chromosome 6. In this study we tried to estimate how much different clinical expressions of gluten sensitivity are determined by genetic factors, and hence how feasible they are for genetic mapping; therefore, we studied all six monozygous twin pairs found among 1292 prospectively collected patients of dermatitis herpetiformis in Finland. Three of the six twin pairs were concordant for dermatitis herpetiformis and for simultaneous enteropathy, celiac disease. Two other twin pairs were partially discordant, one of each pair had dermatitis herpetiformis and celiac disease, whereas the other had solely the gut manifestation of gluten sensitivity, i.e., celiac disease. Only one pair was found to be discordant for gluten sensitivity. All the pairs had typical risk alleles for gluten sensitivity, i.e., either HLA DQ2 or DQ8. These results demonstrate that the genetic component in gluten sensitivity as broadly defined is very strong (5/6 concordant). Genetically identical individuals can have clearly distinguished phenotypes, either dermatitis herpetiformis or celiac disease, suggesting that environmental factors determine the exact phenotype of this multifactorial disease. These findings are of importance in genetic linkage analyses, which focus to only certain phenotypic properties of a complex trait. PMID- 11121132 TI - Functional inhibitory receptors expressed by a cutaneous T cell lymphoma-specific cytolytic clonal T cell population. AB - Inhibitory receptors on natural killer cells and on a minority of T lymphocytes are major histocompatibility complex class Ia or Ib specific. We have previously reported several tumor-specific cytotoxic T cell clones infiltrating a CD4(+) V beta 13(+) cutaneous T cell lymphoma. These clones mediated a specific major histocompatibility complex class I-restricted cytotoxic activity toward the uncultured tumor cells and autologous long-term tumor T cell lines. In this study, we cultured with interleukin-2 the peripheral blood lymphocytes of the same patient a few weeks before invasion of the blood by tumor cells. We report the rapid and selective expansion of a CD8(+) V beta 13(+) lymphoid population. This population was clonal, as it expressed a unique T cell receptor-V beta junctional region. V beta 13(+) tumor cells and V beta 13(+) reactive T cells were shown to have different junctional sequences. The CD8(+) reactive clone was functional, as it had a specific autologous tumor-specific, human leukocyte antigen-A2 restricted, cytotoxic activity. This clone coexpressed high levels of CD158a, CD158b, p70, and CD94/NKG2A inhibitory receptors. Interestingly, we found that anti-CD158a and anti-CD158b monoclonal antibodies could inhibit anti-CD3 redirected cytotoxicity mediated by the reactive clonal population. Further, an anti-human leukocyte antigen-B/C monoclonal antibody enhanced the specific cytotoxic activity of the clone against autologous tumor cells. These results are the first evidence that inhibitory receptor expression can lead to the inhibition of cutaneous T cell lymphoma-specific T cell responses. PMID- 11121133 TI - Human melanoma cells secrete and respond to placenta growth factor and vascular endothelial growth factor. AB - The vascular endothelial growth factor is produced by a large variety of human tumors, including melanoma, in which it appears to play an important role in the process of tumor-induced angiogenesis. Little information is available on the role of placenta growth factor, a member of the vascular endothelial growth factor family of cytokines, in tumor angiogenesis, even though placenta growth factor/vascular endothelial growth factor heterodimers have been recently isolated from tumor cells. To investigate the role of placenta growth factor and vascular endothelial growth factor homodimers and heterodimers in melanoma angiogenesis and growth, 19 human melanoma cell lines derived from primary or metastatic tumors were characterized for the expression of these cytokines and their receptors. Release of placenta growth factor and vascular endothelial growth factor polypeptides into the supernatant of human melanoma cells was demonstrated. Reverse transcriptase polymerase chain reaction analysis showed the presence of mRNAs encoding at least three different vascular endothelial growth factor isoforms (VEGF(121), VEGF(165), and VEGF(189)) and transcripts for two placenta growth factor isoforms (PlGF-1 and PlGF-2) in human melanoma cells. In addition, placenta growth factor expression in human melanoma in vivo was detected by immunohistochemical staining of tumor specimens. Both primary and metastatic melanoma cells were found to express the mRNAs encoding for vascular endothelial growth factor and placenta growth factor receptors (KDR, Flt-1, neuropilin-1, and neuropilin-2), and exposure of melanoma cells to these cytokines resulted in a specific proliferative response, supporting the hypothesis of a role of these angiogenic factors in melanoma growth. J Invest Dermatol 115:1000-1007 2000 PMID- 11121134 TI - Inhibition of growth of melanoma cells by CD95 (Fas/APO-1) gene transfer in vivo. AB - Interaction of CD95 ligand with its cognate receptor CD95 induces apoptotic cell death. Alterations in this pathway within tumor cells can result in escape from apoptosis and from immune surveillance. Melanoma cells recently were found to escape an immune attack via high expression of CD95 ligand, thereby inducing apoptosis of activated T lymphocytes. When screening four human melanoma cell lines for expression of CD95 and CD95 ligand, respectively, an inverse correlation was found, i.e., cells expressing high levels for CD95 ligand (CD95L(high)) were almost negative for CD95 and vice versa. Since coexpression of CD95 and CD95 ligand may lead to apoptosis by autocrine suicide or fratricide, it was tested whether overexpression of CD95 in CD95L(high) melanoma cells results in apoptotic cell death. Upon transfection with a cytomegalovirus-promoter-driven expression vector encoding the CD95 gene, CD95L(high) melanoma cells underwent apoptosis at a much higher level than CD95L(low) melanoma cells. Apoptosis appeared to be due to the activation of CD95 as cell death was inhibited by cotransfection with a dominant negative mutant for the CD95 signaling protein, Fas-associated protein with death domain. Tumor progression of CD95L(high) melanoma cells transplanted into nude mice was significantly reduced when recipient animals were injected with liposomes containing the CD95 expression vector. As demonstrated by immunohistochemistry and TUNEL staining, in vivo transfected tumor cells expressed CD95 and underwent apoptotic cell death. Hence, this study indicates that delivery of the CD95 gene inhibits tumor growth in vivo and thus might be a therapeutic strategy to treat tumor cells that express high levels of CD95 ligand. J Invest Dermatol 115:1008-1014 2000 PMID- 11121135 TI - Acute effects of substance P and calcitonin gene-related peptide in human skin--a microdialysis study. AB - Upon activation nociceptors release neuropeptides in the skin provoking vasodilation and plasma protein extravasation in rodents, but only vasodilation in humans. Pivotal peptides in the induction of neurogenic inflammation comprise calcitonin gene-related peptide and substance P, the latter being suggested to act partly via degranulation of mast cells. In this study substance P and calcitonin gene-related peptide-induced vasodilation, protein extravasation, histamine release, and sensory effects were investigated simultaneously in human skin by dermal microdialysis. The vasodilatory prostaglandin E(2) and the mast cell activator codeine served as positive controls. Substance P and calcitonin gene-related peptide applied intradermally via large cut-off plasmapheresis capillaries induced dose-dependent local vasodilation, but only SP provoked protein extravasation in concentrations greater than 10(-9) M. Substance P induced (10(-8)-10(-6) M) protein extravasation was not accompanied by histamine release and was unaffected by cetirizine (histamine H1 blocker, 200 microg per ml). Only the highest concentration of substance P (10(-5) M) induced significant histamine release. Neither neuropeptide caused any axon reflex erythema or any itch or pain sensation, whereas mast cell degranulation by codeine dose dependently provoked itch, flare, protein extravasation, and histamine release. In human skin calcitonin gene-related peptide and substance P induce vasodilation by a mechanism not involving histamine. No evidence for neuropeptide-induced activation of nociceptors was obtained. Our results suggest that endogenous calcitonin gene-related peptide and substance P have no acute sensory function in human skin. The lack of neurogenic protein extravasation in humans can most probably be attributed to low local concentrations of this neuropeptide still sufficient to exert trophic and immunomodulatory effects (10(-11) M), but too low to induce protein extravasation (10(-8) M) or even mast cell degranulation (10( 5) M). J Invest Dermatol 115:1015-1020 2000 PMID- 11121136 TI - Expression of proopiomelanocortin peptides in human dermal microvascular endothelial cells: evidence for a regulation by ultraviolet light and interleukin 1. AB - Proopiomelanocortin peptides such as alpha-melanocyte-stimulating hormone and adrenocorticotropin are expressed in the epidermal and dermal compartment of the skin after noxious stimuli and are recognized as modulators of immune and inflammatory reactions. Human dermal microvascular endothelial cells mediate leukocyte-endothelial interactions during cutaneous inflammation by the expression of cellular adhesion molecules and cytokines such as interleukin-1. This study addresses the hypothesis that human dermal microvascular endothelial cells express both proopiomelanocortin and prohormone convertases, which are required to generate proopiomelanocortin peptides. Semiquantitative reverse transcriptase polymerase chain reaction and northern blot studies revealed a constitutive expression of proopiomelanocortin mRNA by human dermal microvascular endothelial cells in vitro that was time- and concentration-dependently upregulated by interleukin-1 beta. Furthermore, irradiation of human dermal microvascular endothelial cells with ultraviolet A1 (30J per cm(2)) or ultraviolet B (12.5 mJ per cm(2)) enhanced proopiomelanocortin expression as well as the production and release of the proopiomelanocortin peptides adrenocorticotropin and alpha-melanocyte-stimulating hormone. In addition to proopiomelanocortin, prohormone convertase 1 mRNA expression was detected by reverse transcriptase polymerase chain reaction in unstimulated human dermal microvascular endothelial cells and was augmented after exposure to alpha melanocyte- stimulating hormone, interleukin-1 beta, or irradiation with ultraviolet. These findings demonstrate that human dermal microvascular endothelial cells express proopiomelanocortin and prohormone convertase 1 required for the generation of adrenocorticotropin. Additionally, human dermal microvascular endothelial cells express mRNA for the prohormone convertase 2 binding protein 7B2. Taken together these findings indicate that human dermal microvascular endothelial cells upon stimulation express both proopiomelanocortin and prohormone convertases required for the generation of alpha-melanocyte stimulating hormone. As proopiomelanocortin peptides were found to regulate the production of human dermal microvascular endothelial cell cytokines and adhesion molecules and to have a variety of anti-inflammatory properties these peptides may significantly contribute to the modulation of skin inflammation. J Invest Dermatol 115:1021-1028 2000 PMID- 11121137 TI - Processing of histamine-induced itch in the human cerebral cortex: a correlation analysis with dermal reactions. AB - The subjective sensation of itch is a complex emotional experience depending on a variety of factors. In this study, the central nervous processing of pruritus was investigated in a human model. Activation of involved cerebral areas was correlated to scales of nociception and skin reactions. Six healthy male right handed subjects participated in a standardized epidermal stimulus model with nine increasing doses of histamine dihydrochloride (0.03%-8%) on their right forearms. Controls consisted of three NaCl stimuli. Cerebral activation patterns were determined by H(2)(15)O positron emission tomography 120 s after stimulation. Dermal reactions to the stimulus (wheal, flare, temperature) were coregistered during the procedure. Itch sensation was determined by visual analog scale rating. Pain was not reported during the study; all volunteers had localized itch from 0.03% histamine on. Subtraction analysis versus control revealed significant activation of the left primary sensory cortex and motor-associated areas (mainly primary motor cortex, supplementary motor area, premotor cortex). Predominantly left-sided activations of frontal, orbitofrontal, and superior temporal cortex and anterior cingulate were also observed. Correlation analysis revealed coactivation of dermal reactions and cerebral response to itch in the following Brodmann areas with a Z score greater than 5: wheal, areas 5 (bilateral) and 19 (right); flare, areas 2-5 (left); temperature, area 10 (left) and left insula. Itch intensity ratings were mainly correlated with activation of the left sensory and motor areas. Functional covariates of the itch sensation in the central nervous system were identified. The intention to pruritofensive movements is probably mirrored by the activation of motor areas in the cortex. Other areas may be involved in emotional processing of sensations. Skin reactions wheal and flare also had significantly activated covariate areas in the central nervous system.J Invest Dermatol 115:1029-1033 2000 PMID- 11121138 TI - Distinct effects of CD30 and Fas signaling in cutaneous anaplastic lymphomas: a possible mechanism for disease progression. AB - Lymphomatoid papulosis is part of a spectrum of CD30+ cutaneous lymphoproliferative disorders characterized by spontaneous tumor regression. The mechanism(s) of regression is unknown. In a recent study, a selective increase in CD30 ligand expression in regressing lesions of lymphomatoid papulosis and cutaneous CD30+ anaplastic large cell lymphoma was shown, suggesting that activation of the CD30 signaling pathway may be responsible for tumor regression, whereas no difference in Fas/Fas ligand expression was found between regressing and nonregressing lesions. Therefore we tested the effects of CD30 and Fas activation on three CD30+ cutaneous lymphoma cell lines (Mac-1, Mac-2 A, JK) derived from nonregressing tumors of two patients who had progressed from lymphomatoid papulosis to systemic anaplastic large cell lymphoma. To evaluate the effects of CD30 signaling, the cell lines were incubated with a CD30 agonistic antibody, HeFi-1. Proliferative responses, mitogen-activated protein kinase, and nuclear factor kappa B activities were determined with and without CD30 activation. Mac-1 and Mac-2 A showed increased proliferative responses to incubation with CD30 activating antibody, HeFi-1. Inhibition of the mitogen activated protein kinase activity caused growth inhibition of the Mac-1, Mac-2 A, and JK cell lines. Activation of the Fas pathway induced apoptosis in all three cell lines. Taken together, these findings suggest that resistance to CD30 mediated growth inhibition provides a possible mechanism for escape of cutaneous anaplastic large cell lymphoma from tumor regression. Mitogen-activated protein kinase inhibitors are potential therapeutic agents for the treatment of advanced cutaneous anaplastic large cell lymphoma. J Invest Dermatol 115:1034-1040, 2000 PMID- 11121139 TI - Involvement of Fas (APO-1/CD-95) during photodynamic-therapy-mediated apoptosis in human epidermoid carcinoma A431 cells. AB - Photodynamic therapy is a promising treatment modality for a variety of cutaneous neoplasms and other skin disorders. Studies suggest an involvement of multiple pathways during photodynamic-therapy-mediated cell death. A complete knowledge of the mechanisms involved in photodynamic therapy may lead to an improvement in its therapeutic efficacy. In vitro as well as in vivo studies have shown the involvement of apoptosis during photodynamic- therapy-mediated cell death. The pathways by which photodynamic therapy causes this are not fully understood. In this study, employing human epidermoid carcinoma (A431) cells and silicon phthalocyanine 4 photodynamic therapy, we show that the cell surface death receptor Fas (also known as APO-1 or CD-95) pathway is an important contributor to photodynamic-therapy-mediated apoptosis. Employ- ing flow cytometric analysis and confocal microscopy we first established that silicon phthalocyanine 4 photodynamic therapy results in a significant induction of apoptosis in A431 cells. Immunoblot analysis revealed a significant time-dependent increase in the protein expression of Fas at 5, 15, 30, and 60 min post-photodynamic therapy followed by a decrease at later time-points (2 and 3 h post-photodynamic therapy). A Fas enzyme-linked immunosorbent assay demonstrated an increase in this protein in cell culture medium starting at 1 h post-photodynamic therapy and showing a time-dependent response up to 3 h following therapy, suggesting a diffusion of soluble Fas from cells into the medium from 1 h after photodynamic therapy. Silicon phthalocyanine 4 photodynamic therapy also resulted in a time dependent increase in (i) the multimerization of Fas protein, (ii) the protein expression of Fas ligand, (iii) FADD, an adapter molecule for Fas, and (iv) the binding of FADD with Fas. Silicon phthalocyanine 4 photodynamic therapy also caused a significant activation of FLICE, as evident from the appearance of cleaved products of pro-caspase 8. Further, a pretreatment of cells with rhFas:Fc fusion protein or general caspase inhibitor Z-VAD-FMK followed by silicon phthalocyanine 4 photodynamic therapy resulted in a significantly enhanced cell survival. Taken together, our data, for the first time, delineate an involvement of the Fas pathway as an important contributor to photodynamic-therapy-mediated apoptosis of cancer cells. These observations may be important for improving the efficacy of photodynamic therapy for the treatment of skin cancer and possibly other skin disorders. J Invest Dermatol 115:1041-1046 2000 PMID- 11121140 TI - Basal-cell adhesion molecule (B-CAM) is induced in epithelial skin tumors and inflammatory epidermis, and is expressed at cell-cell and cell-substrate contact sites. AB - Basal-cell adhesion molecule (B-CAM) is a 90 kDa cell surface glycoprotein of the immunoglobulin superfamily that functions as a laminin-binding receptor. B-CAM is upregulated following malignant transformation of some cell types in vivo and in vitro, thus being a candidate molecule involved in tumor progression. As cutaneous distribution and function of B-CAM are largely unknown, we have studied its expression and regulation in normal and diseased human skin. In normal skin, B-CAM was expressed by endothelial cells of dermal blood vessels. In contrast, B CAM was strongly upregulated within the tumor tissue of both malignant and benign epithelial skin tumors, including basal cell carcinomas, squamous cell carcinomas, keratoacanthomas, and common warts. Transformation-associated upregulation was confirmed in vitro, but normal keratinocytes also expressed B CAM under culture conditions. Interestingly, the basal epidermal layer of normal appearing skin surrounding the tumors also expressed B-CAM, and B-CAM were induced on the basal and apicolateral surfaces of basal keratinocytes in inflammatory skin disorders suggesting transformation-independent mechanisms of epidermal induction of the B-CAM. Immunoelectron microscopy studies of cultured transformed keratinocytes revealed that B-CAM was expressed at cell-cell and cell substrate contact sites. Halting proliferation of transformed keratinocytes through cytostatic drugs resulted in decreased B-CAM synthesis. Likewise, inducing terminal differentiation in keratinocyte cultures by increasing the Ca(2+) concentration in the medium decreased B-CAM expression. In contrast, both ultraviolet A and B irradiation of cultured human keratinocytes resulted in significantly increased expression of the B-CAM. Overall, it appears that B-CAM expression in human skin is associated with activated states of keratinocytes, and that B-CAM may be involved in cell-cell adhesion or migration, in addition to its known function as a laminin receptor. J Invest Dermatol 115:1047-1053 2000 PMID- 11121141 TI - Decrease in epidermal CD44 expression as a potential mechanism for abnormal hyaluronate accumulation in superficial dermis in lichen sclerosus et atrophicus. AB - CD44 is a polymorphic integral membrane glycoprotein that serves as the principal cell surface receptor for hyaluronate, the major component of the extracellular matrix. CD44 is abundantly found in the skin and functions as a cell adhesion molecule. In a recent study we have observed a massive dermal accumulation of hyaluronate as a result of the in vivo selective suppression of CD44 in keratinocytes in mice expressing a keratin 5 promoter-driven CD44 anti-sense transgene. As the histologic features of the dorsal skin of these transgenic mice display some similarities to those of the skin lesions of lichen sclerosus et atrophicus, we explored the nature of the material accumulated in the dermis of genital and extragenital lesions of 14 patients with lichen sclerosus et atrophicus by Alcian Blue and human CD44 receptor globulin stainings, as well as the epidermal expression of CD44 protein and mRNA by immunohistochemistry and in situ hybridization. In this study we provide evidence that hyaluronate is accumulated in the superficial dermis of lichen sclerosus et atrophicus lesions, in particular by the use of human CD44 receptor globulin staining, which binds specifically to hyaluronate. In addition we show that the protein and mRNA expression of CD44 in the epidermis of the involved lichen sclerosus et atrophicus skin from genital and extragenital areas is significantly decreased, and in some cases completely lost. In contrast, keratinocyte CD44 expression was un-altered in the skin lesions of lupus erythematosus, scleroderma and reticular erythematous mucinosis, despite the presence of a mucinous material in the dermis. These results suggest that a decrease in CD44 in the keratinocytes may be correlated with an abnormal dermal accumulation of hyaluronate in the lesions of lichen sclerosus et atrophicus, and may play a pathogenetic role in this disease. J Invest Dermatol 115:1054-1058 2000 PMID- 11121142 TI - Elimination of CD4(+) T cells enhances anti-tumor effect of locally secreted interleukin-12 on B16 mouse melanoma and induces vitiligo-like coat color alteration. AB - CD4(+) T cells have been reported to suppress immunity against cancer in certain animal models. In this study, we investigated the role of CD4(+) T cells in the anti-tumor immune response when interleukin-12-producing melanoma cells are inoculated in mice. We found that interleukin-12-transfected B16 melanoma showed retarded tumor growth in syngeneic mice; however, all the mice developed tumors eventually. In vivo depletion of CD4(+) T cells led to complete regression of B16/interleukin-12 tumors in 12 of 20 mice (60%). Immunohistochemical analyses revealed that a number of CD8(+) T cells accumulated in close proximity to the B16/interleukin-12 tumors in the CD4(+) T cell-depleted mice, whereas CD8(+) T cells were only scarcely observed at the periphery of the tumors in control immunocompetent mice. Furthermore, 10 of 20 mice treated with both B16/interleukin-12 inoculation and CD4(+) T cell depletion exhibited vitiligo like coat color alteration. B16/interleukin-12 tumors completely regressed in all the mice with vitiligo. Histologic examination showed that CD8(+) lymphocytes accumulated around the hair bulbs of mice with vitiligo, but not in those without vitiligo. These results suggest that CD4(+) T cells have an inhibitory effect on tumor rejection by suppressing cytotoxic CD8(+) T cells in this melanoma loading model with local interleukin-12 secretion. To investigate the mechanism of enhanced anti-tumor effects by CD4(+) T cell depletion, we examined the T helper type 1/2 cytokine profile in the tumor draining lymph nodes of B16/interleukin-12 bearing mice with or without CD4(+) T cell depletion using the reverse transcription-polymerase chain reaction method. We found that CD4(+) T cell depletion eliminated T helper type 2 cells and resulted in a T helper type 1 dominant cytokine profile in tumor draining lymph nodes. We emphasize that this T helper type 1-dominant cytokine profile may generate further activated CD8(+) T cells against B16 melanoma cells, lead B16/interleukin-12 to regress, and result in the destruction of the melanocytes in hair bulbs due to cross-antigenicity between both cell types. This mouse model not only demonstrates the depletion of CD4(+) T cells as a useful strategy for cancer gene therapy with interleukin-12 but also provides a model for human melanoma-associated vitiligo.J Invest Dermatol 115:1059-1064 2000 PMID- 11121143 TI - Keloid fibroblasts resist ceramide-induced apoptosis by overexpression of insulin like growth factor I receptor. AB - Keloids are benign dermal tumors, characterized by overgrowth of lesions, invasiveness beyond the original boundary of the insult, and recurrence of lesions. The exact etiology is unknown, however. Our hypothesis is that keloids are acquired as a result of an abnormal or prolonged wound healing process, with persistent proliferation and extracellular matrix production of fibroblasts that should otherwise discontinue in normal wound healing. In this study, we examined the response of keloid fibroblasts to proapoptotic signaling. Cell-permeable ceramide, N-acetyl-D-sphingosine, induced apoptosis of dermal fibroblasts in a dose- and time-dependent manner, which was detected by phase contrast microscopy, fluorescent microscopy, the TUNEL method, flow cytometric analysis, and WST-1 assay. In contrast, keloid fibroblasts resisted apoptosis induced by N-acetyl-D sphingosine (percent survival with 40 mM ceramide treatment for 12 h, normal versus keloid: 9.6% +/- 6.6% vs 66.8% +/- 5.5%). Western blotting analysis showed insulin-like growth factor I receptor overexpression in keloid fibroblasts, but not in normal fibroblasts. Exogenously added insulin-like growth factor I enhanced the resistance of keloid fibroblasts to ceramide-induced apoptosis. Wort mannin, a phosphatidylinositol 3 kinase inhibitor, suppressed the antiapoptotic action of insulin-like growth factor I in keloid fibroblasts. Our results suggest that keloid fibroblasts overexpressing insulin-like growth factor I receptor are resistant to apoptosis, thus allowing persistent proliferation and production of excessive extracellular matrix. J Invest Dermatol 115:1065-1071 2000 PMID- 11121144 TI - Loss of normal profilaggrin and filaggrin in flaky tail (ft/ft) mice: an animal model for the filaggrin-deficient skin disease ichthyosis vulgaris. AB - Flaky tail (gene symbol ft) is an autosomal recessive mutation in mice that results in a dry, flaky skin, and annular tail and paw constrictions in the neonatal period. Previous studies demonstrated that the ft mutation maps to the central region of mouse chromosome 3, in the vicinity of the epidermal differentiation complex, a gene locus that includes many nonkeratin genes expressed in epidermis. In this study we report a detailed characterization of the flaky tail mouse. Affected homozygous ft/ft mice exhibit large, disorganized scales on tail and paw skin, marked attenuation of the epidermal granular layer, mild acanthosis, and orthokeratotic hyperkeratosis. Biochemical analysis demonstrated that ft/ft mice lacked normal high molecular profilaggrin (approximately 500 kDa), and instead expressed a lower molecular weight form of profilaggrin (220 kDa) that is not proteolytically processed to profilaggrin intermediates or filaggrin. Mutant mice lacked the large, irregular F-type keratohyalin granules that contain profilaggrin, and filaggrin was absent from the cornified layers of ft/ft epidermis. The expression of epidermal keratins was unchanged, whereas the cornified envelope proteins involucrin and loricrin were increased in ft/ft epidermis. Cultured ft/ft keratinocytes also synthesized reduced amounts of profilaggrin mRNA and protein, demonstrating that the defect in profilaggrin expression is intrinsic to epidermal cells. These findings demonstrate that flaky tail mice express an abnormal profilaggrin polypeptide that does not form normal keratohyalin F-granules and is not proteolytically processed to filaggrin. We propose that the absence of filaggrin, and in particular the hygroscopic, filaggrin-derived amino acids that are thought to function in epidermal hydration, underlies the dry, scaly skin characteristic of ft/ft mice. This animal model provides a tool for understanding the role of filaggrin in normal epidermal function and may provide insight into the molecular basis of the filaggrin-deficient human skin disorder ichthyosis vulgaris. J Invest Dermatol 115:1072-1081 2000 PMID- 11121145 TI - Intracutaneous genetic immunization with autologous melanoma-associated antigen Pmel17/gp100 induces T cell-mediated tumor protection in vivo. AB - Using the differentiation antigen Pmel17/gp100 to genetically immunize C57BL/6 mice (H-2(b)), we and colleagues noticed that only mice that had received the human homolog but not animals injected with the murine counterpart were protected against the growth of syngeneic B16 melanoma cells. The goal of this study was to determine whether the state of nonresponsiveness to the autoantigen Pmel17/gp100 can be broken by immunization with a plasmid DNA construct encoding the autologous form of the molecule. A construct containing the murine form of Pmel17 was administered intradermally to DBA/2 mice (H-2(d)), which were then investigated for the presence of Pmel17/gp100-specific immunity. We show that administration of plasmid DNA coding for the autologous melanoma-associated antigen Pmel17/gp100 protects DBA/2 mice against the growth of Pmel17-positive M3 melanoma cells but not against Pmel17-negative M3 melanoma cells or unrelated P815 mastocytoma cells. Cell depletion experiments demonstrated that this protective effect is mediated by T lymphocytes. The notion that Pmel17/gp100 represents the biologically relevant target in this system was supported by the observations (i) that recipients of Pmel17/gp100 DNA mount an antigen-specific cytotoxic T lymphocyte response and (ii) that M3 tumors growing in mice immunized with autologous Pmel17/gp100 had lost expression of this melanoma-associated antigen whereas M3 melanomas appearing in control-vector-treated animals were still Pmel17/gp100-positive. These results indicate that intracutaneous genetic immunization with autologous melanoma-associated antigen Pmel17/gp100 encoding plasmid DNA can lead to protection against melanoma cells as a result of the induction of a melanoma-associated antigen-specific and protective T-cell mediated immune response. J Invest Dermatol 115:1082-1087 2000 PMID- 11121146 TI - Mutant loricrin is not crosslinked into the cornified cell envelope but is translocated into the nucleus in loricrin keratoderma. AB - Loricrin is a major constituent of the epidermal cornified cell envelope. We have recently identified heterozygous loricrin gene mutations in two dominantly inherited skin diseases, the ichthyotic variant of Vohwinkel syndrome and progressive symmetric erythrokeratoderma, collectively termed loricrin keratoderma. In order to see whether the mutant loricrin molecules predicted by DNA sequencing are expressed in vivo and to define their pathologic effects, we raised antibodies against synthetic peptides corresponding to the mutated sequences of loricrin. Immunoblotting of horny cell extracts from loricrin keratoderma patients showed specific bands for mutant loricrin. Immunohistochemistry of loricrin keratoderma skin biopsies showed positive immunoreactivity to the mutant loricrin antibodies in the nuclei of differentiated epidermal keratinocytes. The immunostaining was localized to the nucleoli of the lower granular cell layer. As keratinocyte differentiation progressed the immunoreactivity moved gradually into the nucleoplasm leaving nucleoli mostly nonimmunoreactive. No substantial staining was observed along the cornified cell envelope. This study confirmed that mutant loricrin was expressed in the loricrin keratoderma skin. Mutant loricrin, as a dominant negative disrupter, is not likely to affect cornified cell envelope crosslinking directly, but seems to interfere with nuclear/nucleolar functions of differentiating keratinocytes. In addition, detection of the mutant loricrin in scraped horny layer could provide a simple noninvasive screening test for loricrin keratoderma. J Invest Dermatol 115:1088-1094 2000 PMID- 11121147 TI - Genetic characterization of a human skin carcinoma progression model: from primary tumor to metastasis. AB - The type and number of genetic aberrations required for a fully malignant tumor are still unclear. This study describes the genetic analysis of a series of skin squamous cell carcinomas, representing the primary tumor, two recurrences, and a metastatic lesion from a single patient and cell lines established therefrom (MET 1 to MET-4). Comparative genomic hybridization demonstrated that: (i) most of the gains and losses were common for tumors and cell lines and affected chromosomes 3 (3p loss, 3q gain), 5 (5p gain, 5q loss), 7 (7p gain), 8 (8p loss, 8q gain), 11 (11q gain), and 17 (17p loss), and (ii) only one aberration was present in a tumor but not in the cell line (10 loss in tumor 4); and only few aberrations were cell line specific. From these, 10p loss and 17q gain were shared by all lines and tumor 4, suggesting that they were already present in all tumors, although in only a subpopulation of cells, whereas 20q gain (shared by all lines), 4q loss (MET-2), and 18p gain/18q loss (MET-3) seem to be culture derived. In agreement, multiplex fluorescence in situ hybridization demonstrated a set of common translocations for all lines thereby further confirming their common origin. In addition, each cell line, exhibited one or more individual translocation chromosomes, which suggested that MET-1 was a precursor of MET-4, whereas MET-2 and MET-3 developed in parallel. Whereas MET-1 to MET-3 were hypodiploid or hyperdiploid, MET-4 was characterized by polyploidization, a set of specific aberrations (t(3;7), t(X;2), i(10q)), and increased heterogeneity (varying translocations in individual metaphases). Using sequencing and expression studies, cells from all lines were wild type for p53, did not exhibit mutations in any of the ras genes (Harvey, Kirsten, or N-ras), and expressed wild type fragile histidine triad gene (FHIT; mapped to 3p14.2, a locus underrepresented in all cells) transcripts. Thus, with the MET cell lines we present an in vivo skin carcinoma progression model that was genetically well defined, and which, despite originating from a sun-exposed site, is wild type for p53. PMID- 11121148 TI - Allelic deletion at 9p21-22 in primary cutaneous CD30(+) large cell lymphoma. AB - The genetic alterations responsible for the development of cutaneous lymphoma are largely unknown. Chromosome region 9p21 contains a gene locus encoding an inhibitor of cyclin-dependent kinase 4, and heterozygous deletions of this tumor suppressor gene (p16) have been shown in a variety of malignant tumors. We studied 11 randomly selected cutaneous CD30-positive large cell lymphomas. Several areas containing 20-50 CD30-positive lymphocytes were microdissected in each case and subjected to single-step DNA extraction. Loss of heterozygosity analysis was performed using polymorphic markers at 9p21 (IFNA, D9S171, D9S169) and 17p13 (TP53). Samples from normal cells apart from CD30-positive lymphocytes, e.g., CD30-negative lymphohistiocytic infiltrates and normal epidermal layer, were also obtained in all cases from the same slide for comparison with the tumor samples. Expression of CD30 and T-lineage antigens (CD3, CD45Ro) was confirmed in all cases. Immunohistochemical staining for p16 and p53 was performed using the monoclonal antibodies sc-1661 and DO-7, respectively. Of the 11 informative cases, seven (64%) exhibited loss of heterozygosity at least for one marker at 9p21 (p16), whereas no allelic deletions were found for the polymorphic marker at 17p13 (p53). On immunohistochemistry loss of the p16 protein was detected in two of 11 cases. Nuclear staining for p53 protein was found in four of 11 cases. Here, we provide the first evidence of the involvement of the tumor suppressor gene p16 in primary cutaneous large cell lymphoma. Whether p16 deletion in these lymphomas is associated with disease progression and whether this method could serve as an early marker to detect lymphomas at an early stage needs to be addressed in future studies. J Invest Dermatol 115:1104-1107 2000 PMID- 11121149 TI - Peroxiredoxin is ubiquitously expressed in rat skin: isotype-specific expression in the epidermis and hair follicle. AB - Peroxiredoxins are a family of peroxidases that are ubiquitously and abundantly expressed in mammalian tissues; however, comparatively less is known about their expression in the skin. In this study, we examined the expression of three isotypes of peroxiredoxins (I-III) in rat skin. Western blot analyses showed strong expression of peroxiredoxins I-III in the epidermis and dermis of intact skin. Additionally, they were expressed in cultured rat keratinocytes and fibroblasts. Confocal image analyses revealed that peroxiredoxin II was present in the cytoplasm as a diffuse, reticulated pattern. In immunohistochemical staining of rat skin, peroxiredoxin expression was mainly localized to the epidermis, hair follicles, and sebaceous glands. In the epidermis, peroxiredoxins I and II were expressed in all layers with a gradient of increasing expression to the granular layer. In contrast, peroxiredoxin III was expressed in all layers with a gradient of expression decreasing to the granular layer. In the hair follicle, peroxiredoxins I-III were mainly expressed in the outer root sheath, except peroxiredoxin II, which was strongly expressed in the inner root sheath. In situ hybridization showed that mRNA expression was commensurate with the level of protein. Ultraviolet B radiation increased peroxiredoxin II expression in rat skin within 15 min after irradiation. From this study we conclude that peroxiredoxin isoforms are ubiquitously expressed in rat skin, and expression of at least peroxiredoxin II can be regulated by ultraviolet irradiation as a peroxidase in the skin. J Invest Dermatol 115:1108-1114 2000 PMID- 11121150 TI - Differential expression of cytokine mRNA in skin specimens from patients with erythema migrans or acrodermatitis chronica atrophicans. AB - Erythema migrans, the characteristic skin manifestation of acute Lyme borreliosis, is a self-limited lesion. In contrast, acrodermatitis chronica atrophicans, the typical cutaneous manifestation of late Lyme borreliosis, is a chronic skin condition. In an effort to understand pathogenic factors that lead to different outcomes in dermatoborrelioses, skin biopsy samples from 42 patients with erythema migrans and 27 patients with acrodermatitis chronica atrophicans were analyzed for mRNA expression of five pro-inflammatory cytokines (tumor necrosis factor alpha, interleukin-1 beta, interleukin-6, interferon-gamma, and interleukin-2) and two anti-inflammatory cytokines (interleukin-4 and interleukin 10) by in situ hybridization with cytokine-specific riboprobes. Among the 27 patients who had erythema migrans alone with no associated signs or symptoms, the major cytokines expressed in perivascular infiltrates of T cells and macrophages were the pro-inflammatory cytokine interferon-gamma and the anti-inflammatory cytokine interleukin-10. In the 15 erythema migrans patients who had associated signs and symptoms, including headache, elevated temperature, arthralgias, myalgias, or fatigue, a larger number of macrophages and greater expression of macrophage-derived pro-inflammatory cytokines, tumor necrosis factor alpha, interleukin-1 beta, and interleukin-6, were also found. In comparison, infiltrates of T cells and macrophages in the skin lesions of acrodermatitis chronica atrophicans patients had very little or no interferon-gamma expression. Instead, they usually expressed only the pro-inflammatory cytokine tumor necrosis factor alpha and the anti-inflammatory cytokine interleukin-4. Thus, the activation of pro-inflammatory cytokines in erythema migrans lesions, particularly interferon-gamma, seems to be important in the control of the spirochetal infection. In contrast, the restricted pattern of cytokine expression in acrodermatitis chronica atrophicans, including the lack of interferon-gamma, may be less effective in spirochetal killing, resulting in the chronicity of this skin lesion. J Invest Dermatol 115:1115-1123 2000 PMID- 11121151 TI - RAG2-/-, I kappa B-alpha-/- chimeras display a psoriasiform skin disease. AB - Nuclear factor-kappa B, a ubiquitous transcription factor involved in inflammatory and immune responses, is inappropriately activated in several immuno related diseases, such as allograft rejection, or bronchial asthma. As nuclear factor-kappa B activity is regulated by inhibitor of kappa B (I kappa B), the gene encoding I kappa B-alpha was disrupted in mice to observe the in vivo effects of hyperactivation of nuclear factor-kappa B. I kappa B-alpha-/- mice have constitutive nuclear factor-kappa B activity, severe skin disease, and neonatal lethality. To determine the role of I kappa B-alpha deficient immunocytes in the pathogenesis of the skin disease in adult mice, we utilized the RAG2-deficient blastocyst complementation system to generate RAG2-/-, I kappa B-alpha-/- chimeras. These animals display a psoriasiform dermatitis characterized by hyperplastic epidermal keratinocytes and dermal infiltration of immunocytes, including lymphocytes. Skin grafts transferred from diseased chimeras to recipient nude mice produce hyperproliferative psoriasiform epidermal keratinocytes in response to stimulation. Furthermore, adoptive transfer of lymph node cells from diseased chimeras to RAG2-/- recipient mice recapitulates the disease. Taken together, these characterizations provide evidence to suggest that constitutive activation of nuclear factor-kappa B, due to deficiency in I kappa B alpha, can invoke severe psoriasiform dermatitis in adult mice. J Invest Dermatol 115:1124-1133 2000 PMID- 11121152 TI - Dysregulated activation of activator protein 1 in keratinocytes of atopic dermatitis patients with enhanced expression of granulocyte/macrophage-colony stimulating factor. AB - Keratinocytes of patients with atopic dermatitis produce high amounts of granulocyte/macrophage colony-stimulating factor, a factor essential for dendritic cell function and thus for the development of skin immune responses. In contrast to keratinocytes cultured from nonatopic, healthy individuals, granulocyte/macrophage colony-stimulating factor mRNA could be detected in unstimulated cultures of atopic dermatitis keratinocytes, and phorbol myristate acetate induced much greater granulocyte/macrophage colony-stimulating factor mRNA levels in these cells, although the decay kinetics were not altered. Using reporter gene (chloramphenicol acetyl transferase) analysis, a minimal granulocyte/macrophage colony-stimulating factor promoter was shown to confer constitutive and phorbol-myristate-acetate-induced regulation of transcriptional activity in keratinocytes, and significantly higher levels of chloramphenicol acetyl transferase activity were measured in lysates of unstimulated and phorbol myristate-acetate-treated atopic dermatitis keratinocytes than in control keratinocyte cultures. Electrophoretic mobility shift assays showed that low levels of NF-kappa B binding activity could be induced by phorbol myristate acetate in both normal and atopic dermatitis keratinocytes. By contrast, activator protein 1 complexes were efficiently induced, and they were invariably present at higher levels in nuclear lysates of atopic dermatitis keratinocytes. Atopic dermatitis keratinocyte nuclear lysates had higher constitutive levels of c-Jun, and phorbol myristate acetate promoted an earlier and stronger expression of c-Jun, JunB, and of the phosphorylated forms of c-Fos. A dysregulated activation of activator protein 1 may be implicated in the molecular mechanisms leading to increased granulocyte/macrophage colony-stimulating factor expression in atopic dermatitis keratinocytes. J Invest Dermatol 115:1134-1143 2000 PMID- 11121153 TI - Mosaicism for ATP2A2 mutations causes segmental Darier's disease. AB - Epidermal naevi are localized malformations of the epidermis consisting of verrucoid scaly papules and plaques following Blaschko's lines. Genetic mosaicism has been proposed to underlie the development of linear epidermal naevi. Rarely, epidermal naevi show acantholytic histology similar to Darier's disease, a dominantly inherited skin condition characterized by widespread warty papules. As patients with acantholytic dyskeratotic naevi often give a history of worsening after sun exposure and the lesions are typical of Darier's disease, numerous authors have proposed that these patients have segmental Darier's disease. The postulated relationship has not been proven, however. Recently, we identified ATP2A2, which encodes the sarco/endoplasmic reticulum Ca(2+) ATPase isoform 2 as the defective gene in Darier's disease. In this report, we investigated the involvement of ATP2A2 in acantholytic dyskeratotic naevi following Blaschko's lines in two patients. We identified a nonsense mutation (Y894X) in the first patient and a nonconservative glycine to arginine mutation at codon 769 (G769R) in the other patient. These mutations were present in affected skin, and were not detected in unaffected skin or in leukocytes. We conclude that acantholytic dyskeratotic naevi can arise from a somatic mutation in ATP2A2. These individuals are mosaics for the mutation, but the risk of transmission of generalized Darier's disease will depend on whether the germline is affected. Our findings provide further evidence that Blaschko's lines do reflect genetic mosaicism and that the term acantholytic dyskeratotic naevus might be replaced in the future by segmental Darier's disease induced by postzygotic mosaicism. J Invest Dermatol 115:1144-1147 2000 PMID- 11121154 TI - Terminal differentiation of human keratinocytes and stratum corneum formation is associated with caspase-14 activation. AB - Programmed cell death of epidermal keratinocytes (KC) results in the formation of cornified cells, which constitute the outermost skin layer, the stratum corneum. Here we show by reverse transcription-polymerase chain reaction, western blot, and immunohistochemistry that epidermal KC express caspase-14, a member of the caspase family of pro-apoptotic proteases, in a tissue-specific manner. Caspase 14 protein abundance strongly increases during terminal differentiation of KC in vivo and in vitro. Under conditions that lead to stratum corneum formation caspase-14 cleavage products, which indicate proenzyme activation, appeared in the KC lysates. Cleavage of the enzyme was also detected in lysates from normal human epidermis and in extracts of stratum corneum. Our findings demonstrate that caspase-14 is activated during KC differentiation and strongly suggest that it is involved in the formation of the human skin barrier.J Invest Dermatol 115:1148 1151 2000 PMID- 11121155 TI - Cyclical changes in rat vibrissa follicles maintained In vitro. AB - In mammals hair growth is cyclical; however, the factors that regulate the hair growth cycle are still poorly understood. The recent development of methods for culturing hair follicles in vitro has proved an important tool to investigate many aspects of the regulation of hair follicle growth. At present, however, these models are based on the culture of anagen hair follicles and have only partially been used to address the cyclical nature of hair growth. In this study we have made use of the fact that in rodents the hair growth cycle is synchronized, well characterized, and relatively short. We have isolated vibrissa follicles from 12 d old rats and confirmed by histology that these follicles are in the anagen stage of their first hair growth cycle. We have then maintained these follicles in vitro, on Gelfoam supports, for up to 23 d (35 d of age) and compared their histology with in vivo follicles from equivalent age littermates. We observed that 12 d old follicles maintained in vitro for up to 23 d show changes in morphology that suggest that cultured rat vibrissa follicles retain cyclical activity in vitro. Cyclical changes in hair follicle morphology were only seen in follicles maintained on gelfoam supports and moreover, hair follicle size appears to be a key feature in determining the ability of the follicle to cycle in vitro. All follicles that showed cyclical changes in vitro, however, appeared to remain blocked in pro-anagen. These data suggest that the vibrissa follicle is a in vitro good model system with which to investigate hair cycle control. J Invest Dermatol 115:1152-1155 2000 PMID- 11121156 TI - Congenital erythropoietic porphyria: a novel homozygous mutation in a Japanese patient. PMID- 11121158 TI - A prospective study of incident nonmelanoma skin cancer in heart transplant recipients. PMID- 11121157 TI - Ultraviolet radiation induces both full activation of ret kinase and malignant melanocytic tumor promotion in RFP-RET-transgenic mice. PMID- 11121159 TI - No modulation of circulating natural killer cell and natural killer receptor bearing memory T cell subsets in patients with atopic dermatitis. PMID- 11121160 TI - Lack of association between promoter polymorphism of the tumor necrosis factor alpha gene and psoriatic arthritis in Japanese patients. PMID- 11121162 TI - IgA antibodies of linear IgA bullous dermatosis recognize the 15th collagenous domain of BP180. PMID- 11121163 TI - Mechanisms other than shunting are likely contributing to the pathophysiology of erythromelalgia. PMID- 11121165 TI - Memory performance and the apolipoprotein E polymorphism in a community sample of middle-aged adults. AB - The apolipoprotein E genotype (APOE) is an established risk factor for Alzheimer disease, with the age-at-onset occurring earlier in individuals having at least one APOE epsilon 4 allele, relative to the APOE epsilon 3 or APOE epsilon 2 isoforms. Moreover, nondemented older adults with the APOE epsilon 4 allele also show diminished cognitive performance, particularly on tests of learning and memory, and an accelerated decline in memory performance with increasing age. The current investigation extends the study of the APOE epsilon 4 allele and cognitive performance to healthy, middle-aged adults. A community sample of 220 non-Hispanic Caucasian men and women, aged 24-60 (average age = 46), were genotyped for the APOE polymorphism and completed a battery of neuropsychological tests. Multivariate analyses were conducted on measures of verbal learning and memory (e. g., learning a list of words and recalling them 30 min later), visual memory (e.g., reproducing a previously copied figure from memory), and attention span (e.g., repeating long lists of digits), after adjustments for age, and estimated IQ. Results indicated that performance on learning and memory tasks was significantly poorer in adults having any APOE epsilon 4 allele, relative to adults with APOE epsilon 2 and epsilon 3 genotypes (P <.01). Attention span did not differ by genotype. These findings, the first in a sample of middle-aged adults, suggest that the APOE polymorphism is a marker for age-related decline in memory (detectable prior to overt, clinical manifestations of memory loss), and/or a marker for individual differences in memory ability across the life span. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:707-711, 2000. PMID- 11121166 TI - Tardive dyskinesia is not associated with the serotonin gene polymorphism (5 HTTLPR) in Chinese. AB - Neuroleptics are the mainstay of treatment for schizophrenia, but one of the complications is the development of tardive dyskinesia (TD). The pathophysiology of TD may involve dopamine-serotonin interaction. The serotonin transporter participates in the reuptake and termination of serotonin neurotransmission, and the gene that codes for this protein is thus a candidate gene for the development of TD. There is a functional polymorphism in the transcriptional control region of the serotonin transporter gene, and we investigated the association between this polymorphism and TD in Chinese schizophrenic patients. The patients who did not differ in age and sex distribution did not show variation on the rates of TD and Abnormal Involuntary Movements Scale (AIMS) scores with genotypes. Our findings suggest that 5-HTTLPR polymorphism is not a risk factor for TD in Chinese. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:712-715, 2000. PMID- 11121167 TI - Mutation and association analysis of the Fyn kinase gene with alcoholism and schizophrenia. AB - Lack of Fyn tyrosine kinase increases alcohol sensitivity. Fyn phosphorylates a component of the NMDA receptor, which may be involved in schizophrenia. The Fyn gene is located on human chromosome 6q21, to which linkage of schizophrenia has been suggested. We hypothesized that the Fyn gene is a candidate for predisposition to alcoholism and schizophrenia, and we performed a mutation study of the 5'-flanking region, all coding exons, and exon-intron junctions of the Fyn gene. The SSCP mutation analysis was performed in 48 unrelated alcoholics and 16 unrelated schizophrenics. Three polymorphisms, -93A/G in the 5'-flanking region, IVS10+37T/C in intron 10, and Ex12+894T/G in the 3'-untranslated region, were identified. A rare variant of Ex12+1162TG in the 3'-untranslated region was also detected. Neither missense nor nonsense mutations were found. Case-control studies using a larger sample of unrelated patients and controls did not reveal significant associations between these polymorphisms and alcoholism or schizophrenia. In addition, genotyping a microsatellite marker, D6S302, located in intron 10 of the Fyn gene, did not show a significant association between the marker and alcoholism or schizophrenia. Results of the present study did not provide evidence for the involvement of the genomic Fyn gene mutations in alcoholism or schizophrenia. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:716 720, 2000. PMID- 11121168 TI - Association between homozygosity at the COMT gene locus and obsessive compulsive disorder. AB - A functional polymorphism in the coding region of the catechol O methyltransferase (COMT) gene has been reported in previous studies to be associated with obsessive compulsive disorder (OCD), particularly in males [Karayiorgou et al., 1997, 1999]. Using a family-based population analysis, we attempted to replicate these findings in a group of 72 OCD patient/parent trios collected from Buffalo, New York, and Toronto, Canada. Analysis of allele and genotype frequencies using the haplotype relative risk (HRR) and transmission disequilibrium test (TDT) did not identify an association between a particular allele and OCD as had been previously reported. Furthermore, no evidence was found to support the findings of a gender-based association for COMT when the patients and the parents of the same gender were compared. However, our genotype results (n = 72) demonstrate a tendency for association between homozygosity at the COMT locus and OCD (homozygosity analysis: chi(2) = 5.66, P = 0.017; genotypic analysis: chi(2) = 5.78, P = 0.056). Although these findings do not replicate the previous reports, they do provide limited support to demonstrate a trend for homozygosity at the COMT locus in the OCD patients and, in turn, further implicate a potential role for COMT in the genetic etiology of OCD. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:721-724, 2000. PMID- 11121169 TI - Support for an association between HLA-DR1 and schizophrenia in the Japanese population. AB - An increase of HLA-DR1 has been observed in schizophrenia patients from the Japanese population. A decrease of DR4, which was reported in Caucasian patients, has also been found in some of the Japanese studies. This small study further investigated frequencies of HLA-DR1 and DR4 in unrelated Japanese patients with schizophrenia (n = 45) and healthy comparison subjects (n = 117). The number of patients possessing DR1 was higher (10 of 45, 22%) compared with the comparison group (11 of 117, 9.4%, P = 0.03). This may support the previous observation of an increased DR1 frequency in the Japanese patients. When the present data is combined with three previous studies, proportions of the Japanese subjects with DR1 were 98 of 588 schizophrenia patients (16.7%) vs. 93 of 942 comparison subjects (9.9%). However, no difference was observed in DR4 frequencies between the patients (51%) and comparison subjects (44%). Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:725-727, 2000. PMID- 11121170 TI - Search for bipolar disorder susceptibility loci: the application of a modified genome scan concentrating on gene-rich regions. AB - Conducting genome wide screens for evidence of genetic linkage has become a well established method for identifying regions of the human genome harboring susceptibility loci for complex disorders. For bipolar disorder, a number of such studies have been performed, and several regions of the genome have potentially been implicated in the disorder. The classic design for a genome screen involves examining polymorphic genetic markers spaced at regular intervals throughout the genome, typically every 10 cM, for evidence of linkage. An alternative design, based on the observation that genes do not appear to be evenly distributed, was proposed, enabling the number of markers examined in a genome wide screen to be reduced. This article describes the application of such a modified screen to a collection of 48 Irish families with bipolar disorder, comprising a total of 82 affected sib-pairs. From the results obtained a number of regions are highlighted for further study. One of these regions (17q11.1-q12) coincides with the location of a candidate gene, the serotonin transporter, whereas others concur with the findings of published studies. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:728 732, 2000. PMID- 11121171 TI - No association of an insertion/deletion polymorphism in the angiotensin I converting enzyme gene with bipolar or unipolar affective disorders. AB - A recent Japanese study on the angiotensin I converting enzyme gene (ACE) insertion/deletion polymorphism reported that both the D allele (P < 0.02) and the DD genotype (P < 0.002) were significantly more frequent in affective disorder cases than in controls [Arinami et al., 1996: Biol Psychiatry 40:1122 1127]. A replication study was performed by using 157 bipolar I affective disorder cases, 169 major depressive disorder cases, and 313 controls. No significant association with this polymorphism was found in either disorder or in a combined affective disorder group. These results do not support the ACE gene having a major role in the etiology of either bipolar or unipolar affective disorders. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:733-735, 2000. PMID- 11121172 TI - Systematic screening of the 14-3-3 eta (eta) chain gene for polymorphic variants and case-control analysis in schizophrenia. AB - The neuronal protein 14-3-3 eta is a candidate gene for schizophrenia because it maps chromosome 22q12, a region implicated in the disease by linkage analysis, and is involved in brain development. We systematically screened this gene for polymorphic variants by comparison of public EST sequence data (five cDNAs and 72 ESTs, 21,155 bp of sequence) in parallel with single-stranded conformational polymorphism analysis, and we compared these methods by using a simple power calculation. Twelve potential polymorphisms were identified from EST sequence comparison, and two of these (a 5'-VNTR and 753G/A) were confirmed by SSCP analysis and sequencing. Three additional infrequent polymorphisms (-408T/G; 177 C/G; and 989 A/G) were found by SSCP only. We next examined these variants for association with schizophrenia. One variant in untranslated region of exon 1 ( 408 T/G) was found to occur more frequently in the schizophrenic subjects (8%) than the controls (3%; P = 0.01). After fivefold correction of the P value for multiple testing, marginal association was found. Haplotype analysis of pairs of polymorphisms provided no evidence for association of this gene with schizophrenia in the population studied. Am. J. Med. Genet. (Neuropsychiatr. Genet. ) 96:736-743, 2000. PMID- 11121173 TI - Association analysis of CAG repeats at the KCNN3 locus in Indian patients with bipolar disorder and schizophrenia. AB - Bipolar affective disorder and schizophrenia are severe behavioral disorders with a lifetime risk of approximately 1% in the population worldwide. There is evidence that these diseases may manifest the phenomenon of anticipation similar to that seen in diseases caused by trinucleotide repeat expansions. A recent report has implicated a potassium channel-coding gene, KCNN3, which contains a polymorphic CAG repeat in its coding region, in schizophrenia and bipolar disorder. We have tried to confirm these findings in Indian patients suffering from bipolar disorder and schizophrenia. No statistically significant evidence for the presence of an excess of longer alleles in the patient population, as compared to ethnically matched controls, was found. However, an analysis of the difference of allele sizes revealed a significantly greater number of patients with schizophrenia having differences of allele sizes > or = 5 when compared to normal controls. This finding may be of functional significance as the KCNN3 protein is thought to act as a tetramer, and a large difference in allele sizes would result in an asymmetric molecule with a different number of glutamine residues in each monomer. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:744-748, 2000. PMID- 11121174 TI - Childhood-onset schizophrenia/autistic disorder and t(1;7) reciprocal translocation: identification of a BAC contig spanning the translocation breakpoint at 7q21. AB - Childhood-onset schizophrenia (COS) is defined by the development of first psychotic symptoms by age 12. While recruiting patients with COS refractory to conventional treatments for a trial of atypical antipsychotic drugs, we discovered a unique case who has a familial t(1;7)(p22;q21) reciprocal translocation and onset of psychosis at age 9. The patient also has symptoms of autistic disorder, which are usually transient before the first psychotic episode among 40-50% of the childhood schizophrenics but has persisted in him even after the remission of psychosis. Cosegregating with the translocation, among the carriers in the family available for the study, are other significant psychopathologies, including alcohol/drug abuse, severe impulsivity, and paranoid personality and language delay. This case may provide a model for understanding the genetic basis of schizophrenia or autism. Here we report the progress toward characterization of genomic organization across the translocation breakpoint at 7q21. The polymorphic markers, D7S630/D7S492 and D7S2410/D7S646, immediately flanking the breakpoint, may be useful for further confirming the genetic linkage for schizophrenia or autism in this region. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:749-753, 2000. Published 2000 Wiley-Liss, Inc. PMID- 11121175 TI - Influence of ascertainment strategy on finding sex differences in genetic estimates from twin studies of alcoholism. AB - Twin studies have yielded contradictory findings about sex differences in genetic influences on the etiology of alcoholism. Studies based on population registers or epidemiological samples have yielded similar estimates of heritability (50-60% of the total variance) for males and females. In contrast, studies of twins identified through treatment settings have found sizeable genetic contributions to alcoholism in males but usually negligible heritabilities for females. We investigated this discrepancy by applying a "simulated" treatment ascertainment strategy to data on alcohol-related disorders collected by structured interviews with a population-based sample of adult twins aged 18-56 years from the Mid Atlantic Twin Registry. Structural models were used to estimate heritabilities for two definitions of treatment, and these estimates were compared with those obtained from the population-based sample. In males, heritability estimates were similar across sampling methods, but the treatment ascertainment methods yielded higher estimates of common environmental influences. For females, heritability estimates based on a broad definition of treatment were similar to those obtained by using the random ascertainment design. However, estimates based on sampling women who had been in alcohol-treatment programs were (nonsignificantly) lower than those obtained with the other methods. These results provide partial support for the hypothesis that differences in sampling method may account for differences in heritability estimates for alcoholism among studies of female twins. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:754-761, 2000. PMID- 11121176 TI - The homozygous C677T mutation in the methylenetetrahydrofolate reductase gene is a genetic risk factor for migraine. AB - Increased homocysteine levels are associated with various pathological conditions in humans, including stroke and cardiovascular disorders. Homocysteine acts as an excitatory amino acid in vivo and may influence the threshold of migraine headache. Frosst et al. [1995] reported an association between the homozygous C677T mutation in the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene and serum homocysteine levels. This study was designed to determine the prevalence of the MTHFR mutation in Japanese patients with migraine and tension-type headache (TH). Seventy-four patients with migraine headaches (22 with aura and 52 without aura), 47 with THs, and 261 normal controls were recruited. Genotyping of MTHFR C677T polymorphism was performed by polymerase chain reaction-restriction fragment length polymorphism. We detected that the incidence of the homozygous transition (T/T) in migraine sufferers (20.3%) was significantly higher than that in controls (9.6%). Moreover, the frequency of the T/T genotype in individuals with migraine headaches with aura was remarkably high (40.9%). The MTHFR T allele was more frequent in the migraine group than in the control group. Our results support the conclusion that the MTHFR gene, causing mild hyperhomocysteinemia may be a genetic risk factor for migraine. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:762-764, 2000. PMID- 11121177 TI - Analysis of a 1-megabase deletion in 15q22-q23 in an autistic patient: identification of candidate genes for autism and of homologous DNA segments in 15q22-q23 and 15q11-q13. AB - We have identified a one megabase deletion in the 15q22-15q23 region in a patient with autism, developmental delay, and mild dysmorphism. Genes that map within the deletion region and genes that are interrupted or rearranged at the deletion breakpoints are candidate genes for autism. Fluroescence in situ hybridization studies in this patient revealed that part or all of the PML gene is absent from one chromosome 15 and a BAC clone containing the D15S124 gene locus hybridizes to only one chromosome 15. BAC clones containing the PTPN9, and SLP-1[hUNC24] genes showed markedly reduced hybridization in the 15q22-q23 region on one chromosome 15 in the patient. These BACs also hybridize to the 15q11-q13 region in close proximity to SNRPN and HERC2, and in this region there is equal intensity of signal on the normal and on the deleted chromosome. There are previous reports of deletions and duplications of the 15q11-q13 region in patients with autism. Our patient represents the first report of a 15q22-q23 deletion. Hybridization of the PTPN9 and Slp-1 Bac clones to the 15q11-q13 and the 15q22-q23 regions of chromosome 15 may be due to the presence of PTPN9 or SLP-1 gene sequences or to the presence of other gene sequences or to non-coding homologous DNA sequences. The PTPN9 gene encodes a non-receptor protein tyrosine phosphatase. The Slp-1 [hUNC24] gene is expressed mainly in the brain. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:765-770, 2000. PMID- 11121178 TI - Exploratory association study between catechol-O-methyltransferase (COMT) high/low enzyme activity polymorphism and hypnotizability. AB - Only recently have studies of electrocortical activity, event-related potentials, and regional cerebral blood flow begun to shed light on the anatomical and neurobiological underpinnings of hypnosis. Since twin studies show a significant heritable component for hypnotizability, we were prompted to examine the role of a common, functional polymorphism in contributing to individual differences in hypnotizability. A group of 109 subjects (51 male, 59 female) were administered three psychological instruments and tested for the high/low enzyme activity COMT val-->met polymorphism. We observed a significant correlation between hypnotizability measured by the Stanford Hypnotic Susceptibility Scale (SHSS:C), ability to partition attention (Differential Attentional Processes Inventory or DAPI), and absorptive capacities (Tellegen Absorption Scale or TAS). The effect of COMT on the various dependent variables was initially examined by multivariate analysis that corrects for multiple testing. The dependent variables were SHSS:C hypnotizability scores, four attentional subscales of the DAPI, and TAS total score grouped by the COMT genotype (val/val, val/met, met/met) as the independent variable. Hotelling's Trace statistic was significant when scores were grouped by the COMT genotype (Hotelling's T(2) = 1.88, P = 0.04. Post-hoc testing using the Bonferroni correction shows that the only significant difference is between the val/met vs. the val/val COMT genotypes on hypnotizability. This association was significant for men but not for women. As for all case-control studies, these results need to be interpreted cautiously and require replication. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:771-774, 2000. PMID- 11121179 TI - Identical distribution of the alpha 2-macroglobulin pentanucleotide deletion in subjects with Alzheimer disease and controls in a German population. AB - Recently, an association between a deletion polymorphism in the alpha 2 macroglobulin gene (A2M) and Alzheimer disease (AD) has been reported. The aim of the present study was to corroborate this association in a German population of 102 AD patients and two control samples of 191 healthy subject and 160 depressed patients. The frequency of the A2M genotype in AD patients was almost identical to that in both control samples. Logistic regression analysis revealed an effect of age and the APOE genotype on AD risk, but no effect of the A2M genotype. Our findings do not support the fact that the previously reported positive association between A2M deletion polymorphism and AD modifies the disease risk in the studied population. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:775-777, 2000. PMID- 11121180 TI - No association between the dopamine D3 receptor Bal I polymorphism and schizophrenia in a family-based study of a Palestinian Arab population. AB - Several recent meta-analyses appear to show a weak but significant effect of both forms of the gly/ser DRD3 polymorphism in conferring risk for schizophrenia. Since most studies have employed the artifact-prone case-control design, we thought it worthwhile to examine the role of this polymorphism using a robust family-based strategy in an ethnic group not previously systematically studied in psychiatric genetics, Palestinian Arabs. We failed to obtain any evidence in 129 Palestinian triads, using the haplotype relative risk (allele frequency: Pearson chi-square = 0.009, P > 0.1, df = 1, n = 258 alleles) or transmission disequilibrium test design (chi-square = 0.38, P > 0.1, n = 86 families) for association/linkage (or increased homozygosity) of the DRD3 Bal I polymorphism to schizophrenia in our sample. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:778 780, 2000. PMID- 11121181 TI - Drawbacks of GENEHUNTER for larger pedigrees: application to panic disorder. AB - Large pedigrees can pose a problem for GENEHUNTER linkage analysis software. Differences in two-point and multipoint lodscores were observed when comparing GENEHUNTER to other linkage software. Careful consideration must be given when selecting linkage analysis programs. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:781-783, 2000. PMID- 11121182 TI - Adenosine deaminase alleles and autistic disorder: case-control and family-based association studies. AB - Adenosine deaminase (ADA) plays a relevant role in purine metabolism, immune responses, and peptidase activity, which may be altered in some autistic patients. Codominant ADA1 and ADA2 alleles code for ADA1 and ADA2 allozymes, the most frequent protein isoforms in the general population. Individuals carrying one copy of the ADA2 allele display 15 to 20% lower catalytic activity compared to ADA1 homozygotes. Recent preliminary data suggest that ADA2 alleles may be more frequent among autistic patients than healthy controls. The present study was undertaken to replicate these findings in a new case-control study, to test for linkage/association using a family-based design, and to characterize ADA2 carrying patients by serotonin blood levels, peptiduria, and head circumference. ADA2 alleles were significantly more frequent in 91 Caucasian autistic patients of Italian descent than in 152 unaffected controls (17.6% vs. 7.9%, P = 0.018), as well as among their fathers. Family-based tests involving these 91 singleton families, as well as 44 additional Caucasian-American trios, did not support significant linkage/association. However, the observed preferential maternal transmission of ADA2 alleles, if replicated, may point toward linkage disequilibrium between the ADA2 polymorphism and an imprinted gene variant located in its vicinity. Racial and ethnic differences in ADA allelic distributions, together with the low frequency of the ADA2 allele, may pose methodological problems to future linkage/association studies. Direct assessments of ADA catalytic activity in autistic individuals and unaffected siblings carrying ADA1/ADA1 vs ADA1/ADA2 genotypes may provide stronger evidence of ADA2 contributions to autistic disorder. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:784-790, 2000. PMID- 11121183 TI - Obsessive and compulsive symptoms in a general population sample of female twins. AB - Obsessive-compulsive disorder (OCD) and obsessive-compulsive symptoms (OCS) exhibit a familial pattern of transmission. The different components of these conditions and the extent to which these components are inherited have not been studied well. A sample of 1,054 female twins, including both members of 527 pairs, from the Virginia Twin Registry returned questionnaires that included 20 items from the Padua Inventory of obsessive-compulsiveness. Their responses were used to estimate the heritability of the different factors of OCS in this population. Principal components analysis suggested two meaningful factors corresponding roughly to obsessions and compulsions. The best-fit model suggested heritabilities of 33 and 26%, respectively. The correlation between additive genetic effects on compulsiveness and obsessiveness was found to be +0.53. Self report symptoms of obsessions and compulsions in women from the general population are moderately heritable and due, in part, to the same genetic risk factors. An understanding of the etiology of these symptoms is relevant to the study of OCD. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:791-796, 2000. PMID- 11121184 TI - Association of 5HT2A receptor gene polymorphism and alcohol abuse with behavior problems. AB - This study investigated the association between T/C polymorphism, at position 102, of the 5-hydroxytryptamine 2A receptor gene and alcoholism with and without behavior problems. Eighty-five subjects (45 men, 40 women) with alcohol abuse, 75 subjects (51 men, 24 women) with alcohol dependence, and 70 normal control subjects (21 men, 49 women) participated in the study. The results show that the frequency of the homozygous T102 genotype was significantly lower in the group of male alcohol abuse with behavior problems than in the female group (chi(2) = 4.072, df = 1, P < 0.05) and the allele frequency of T102 was also lower in the male group than in the female group (chi(2) = 4.187, df = 1, P < 0.05). Of the male alcohol abuse subjects, the group with behavior problems was found to have lower frequencies of the T102 allele than the group without behavior problems (chi(2) = 4.328, df = 1, P < 0.05). In conclusion, this study demonstrates that alcoholism is heterogeneous and male alcohol abuse with behavioral problems was associated with T/C 102 polymorphism of the 5HT2A receptor gene. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:797-800, 2000. PMID- 11121185 TI - Association between a functional polymorphism in the monoamine oxidase A gene promoter and major depressive disorder. AB - Various polymorphisms of the X-chromosomal monoamine oxidase A (MAO-A) gene were investigated for association with affective disorders. However, none of the studied variants could consistently be associated with either major depressive or bipolar affective disorder. Recently, a positive association between panic disorder and a novel functional repeat polymorphism in the MAO-A gene promoter, with the longer alleles being more active, was reported. Since monoaminergic neurotransmission is supposed to play an important role in affective disorders, we investigated a potential association of this polymorphism with major depressive illness in a sample of 146 unrelated patients of German descent and a control group of 101 individuals with a negative life history for affective disorders. Similarly to the recent findings in panic disorder, we observed a significantly increased frequency of genotypes containing only long alleles in female patients with recurrent major depression in comparison with age- and sex matched controls. Thus, our data suggest that an excess of high-activity MAO-A gene promoter alleles resulting in an elevated MAO-A activity is a risk factor for major depressive disorder in females. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:801-803, 2000. PMID- 11121186 TI - Anticipation in bipolar affective disorder: is age at onset a valid criterion? AB - Anticipation has been suggested among the genetic mechanisms of bipolar disorder (BD), prompting the search for unstable DNA sequences. Past studies of anticipation in BD have generally relied on observed shift in the age at onset between parental and offspring generations. Such a shift, however, may be caused by a number of other factors difficult to correct for. We investigated age at onset distributions in a sample of 161 related subjects and in a sample of "pseudofamilies" consisting of 320 unrelated subjects selected from a large epidemiological cohort using Monte-Carlo simulation to mimic the family sample. Comparison of age at onset distributions in both samples shows a difference between the generations, but of a similar magnitude in each sample. This suggests that age at onset alone may not be a sufficient criterion of anticipation. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:804-807, 2000. PMID- 11121187 TI - NURR1 mutations in cases of schizophrenia and manic-depressive disorder. AB - Transgenic mice lacking the nuclear orphan transcription factor Nur-related receptor 1 (Nurr1) fail to develop mesencephalic dopamine neurons. There is a highly homologous NURR1 gene in humans (formerly known as NOT) which therefore constitutes a good candidate gene for neurologic and psychiatric disorders with an involvement of the dopamine neuron system, such as Parkinson's disease, schizophrenia, and manic-depression. By direct sequencing of genomic DNA, we found two different missense mutations in the third exon of NURR1 in two schizophrenic patients and another missense mutation in the same exon in an individual with manic-depressive disorder. All three mutations caused a similar reduction of in vitro transcriptional activity of NURR1 dimers of about 30-40%. Neither of these amino acid changes, nor any sequence changes whatsoever, were found in patients with Parkinson's disease or control DNA material of normal populations. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:808-813, 2000. PMID- 11121188 TI - Failure to replicate association between the gene for the neuronal nicotinic acetylcholine receptor alpha 4 subunit (CHRNA4) and IGE. AB - The gene for the neuronal nicotinic acetylcholine receptor alpha4 subunit (CHRNA4) was identified as a gene underlying a rare idiopathic partial epilepsy syndrome in humans, autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). In a recent study, one of four silent polymorphisms (594 C/T) in CHRNA4 showed association with the common subtypes of idiopathic generalised epilepsy (IGE). In the present study, three of these polymorphisms were investigated for association in 182 Caucasian patients with IGE, but not categorised by subtype. They were compared with 178 controls in a case/control study. Further analyses were performed using a family-based design. None of the three polymorphisms exhibited any association with IGE. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:814-816, 2000. PMID- 11121189 TI - Implications of comorbidity and ascertainment bias for identifying disease genes. AB - Comorbidity, the co-occurrence of disorders, is frequently observed to occur at higher rates in clinically ascertained samples than in population-based samples. An explanation for this finding is that subjects suffering from multiple illnesses are more likely to seek medical care and receive a diagnostic evaluation. We refer to the component of the comorbidity between illnesses due to such ascertainment bias as "spurious comorbidity." When spurious comorbidity is present, an apparent association between a candidate locus and the phenotype of interest may actually be attributable to an association between the locus and a comorbid phenotype. This phenomenon, which we call "spurious comorbidity bias," could thus produce misleading association findings. In this article, we describe this phenomenon and demonstrate that it may produce marked bias in the conclusions of family-based association studies. Because of the extremely high rates of comorbidity among psychiatric disorders in clinical samples, this problem may be particularly salient for genetic studies of neuropsychiatric disorders. We conclude that ascertainment bias may contribute to the frequent difficulty in replicating candidate gene study findings in psychiatry. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:817-822, 2000. PMID- 11121190 TI - Association of a haplotype for tumor necrosis factor in siblings with late-onset Alzheimer disease: the NIMH Alzheimer Disease Genetics Initiative. AB - Tumor necrosis factor (TNF), a proinflammatory cytokine, may be involved in the pathogenesis of Alzheimer disease (AD) based on observations that senile plaques have been found to upregulate proinflammatory cytokines. Additionally, nonsteroidal anti-inflammatory drugs have been found to delay and prevent the onset of AD. A collaborative genome-wide scan for AD genes in 266 late-onset families implicated a 20 centimorgan region at chromosome 6p21.3 that includes the TNF gene. Three TNF polymorphisms, a -308 TNF promoter polymorphism, whose TNF2 allele is associated with autoimmune inflammatory diseases and strong transcriptional activity, the -238 TNF promoter polymorphism, and the microsatellite TNFa, whose 2 allele is associated with a high TNF secretion, were typed in 145 families consisting of 562 affected and unaffected siblings. These polymorphisms formed a haplotype, 2-1-2, respectively, that was significantly associated with AD (P = 0.005) using the sibling disequilibrium test. Singly, the TNFa2 allele was also significantly associated (P = 0.04) with AD in these 145 families. This TNF association with AD lends further support for an inflammatory process in the pathogenesis of AD. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:823-830, 2000. PMID- 11121191 TI - Lack of association of serotonin-2A receptor gene polymorphism (T102C) with suicidal ideation and suicide. AB - Serotonergic dysfunction has been implicated in the pathophysiology of affective disorders and suicidality. Especially the density of the 5-HT2A receptor was claimed as being increased in suicidality, proposed as an adaptive upregulation due to reduced serotonergic transmission. Recent studies have shown an association of allele C of the 5-HT2A-T102C polymorphism with suicidal ideation in patients with major depression. The purpose of this study was to test whether this proposed marker indicates susceptibility not only to suicidal ideation in depressed patients but also to suicidality as a syndrome. We investigated the 5 HT2A-T102C polymorphism in 131 suicide victims with unknown underlying psychiatric diagnoses, 84 patients with major depression with or without suicidal ideation, and 125 healthy controls. We were unable to find any association of genotype or allele frequencies to major depression, suicidal ideation, or suicide as a syndrome. Thus, our results suggest that this polymorphism may not commonly be involved in the susceptibility to suicidality. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:831-835, 2000. PMID- 11121192 TI - Evidence for linkage by transmission disequilibrium test analysis of a chromosome 22 microsatellite marker D22S278 and bipolar disorder in a Palestinian Arab population. AB - A number of linkage studies suggest a schizophrenia susceptibility locus on chromosome 22, particularly with microsatellite marker D22S278 (22q12). In addition to some evidence for linkage to schizophrenia in this region, linkage to bipolar disorder using this marker has also been reported. We tested a group of 60 Bipolar I triads and an expanded group of 79 Bipolar I and Bipolar II triads recruited from a Palestinian Arab population for linkage with the D22S278 marker. Significant linkage was observed using the extended transmission disequilibrium test for multiallelic markers (ETDT) for both Bipolar I (P = 0.031) and the expanded group of Bipolar I and Bipolar II (P = 0.041). These weakly positive results are at least consistent with the hypothesis that this region of chromosome 22 might harbor a susceptibility locus for both major psychoses and should be considered for more intensive study. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:836-838, 2000. PMID- 11121193 TI - Case with autistic syndrome and chromosome 22q13.3 deletion detected by FISH. AB - Autism is a rare neurodevelopmental disorder with a strong genetic component. Co occurrence of autism and chromosomal abnormalities is useful to localize candidate regions that may include gene(s) implicated in autism determinism. Several candidate chromosomal regions are known, but association of chromosome 22 abnormalities with autism is unusual. We report a child with autistic syndrome and a de novo 22q13.3 cryptic deletion detected by FISH. Previously described cases with 22q13.3 deletions shared characteristic developmental and speech delay, but autism was not specifically reported. This case emphasizes a new candidate region that may bear a gene involved in autism etiopathogenesis. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:839-844, 2000. PMID- 11121194 TI - Multimarkerhaplotypes within the serotonin transporter gene suggest evidence of an association with bipolar disorder. AB - Previously we obtained modest linkage evidence implicating 17q11. 1-12 in bipolar disorder. A modified genome screen, based on gene-rich regions, on a collection of Irish sib-pair nuclear families revealed excess allele sharing at markers flanking the gene encoding the serotonin transporter (5-HTT; hSERT). Here we describe a study designed to combine the advantages of family-based association studies with the consideration of multiple polymorphic markers within a candidate gene. Ninety-two Irish families, with a total of 106 proband-parent trios, have been genotyped for 3 previously known polymorphisms within hSERT (5-HTTLPR, intron 2 VNTR, and 3' UTR G/T). Data from two and three polymorphic marker haplotypes revealed a number of marker combinations that showed evidence supportive of association; the most significant being for polymorphisms 5-HTTLPR and 3' UTR G/T (global chi(2), 12.91, df 3, P = 0.005). In addition, modest evidence of association also was observed for 5-HTTLPR alone. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:845-849, 2000. PMID- 11121195 TI - Molecular genetic studies of bipolar disorder and puerperal psychosis at two polymorphisms in the estrogen receptor alpha gene (ESR 1). AB - A number of lines of evidence point to the possible involvement of estrogen pathways in the pathophysiology of bipolar disorder in general and puerperal psychosis in particular. There is strong evidence from clinical, follow-up, and genetic studies to support the hypothesis that most cases of puerperal psychosis are manifestations of an affective disorder diathesis with a puerperal trigger and that genes influence susceptibility to both diathesis and trigger. The nature of the trigger is unknown but in view of the abrupt onset at a time of major physiological change it is widely believed that biological, probably hormonal, mechanisms are of paramount importance, with estrogen receiving the most attention to date. We have undertaken a case control association study of bipolar disorder and puerperal psychosis at two known polymorphisms within the estrogen receptor alpha gene (ESR 1) in a sample of 219 unrelated bipolar probands and 219 controls. We could exclude these polymorphisms from an important contribution to susceptibility to bipolar disorder with a high level of confidence. We found no support for the hypothesis that they contribute specific susceptibility to the puerperal trigger, but due to the small numbers of puerperal probands (n = 26) no firm conclusions can be drawn regarding their involvement in puerperal psychosis. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:850-853, 2000. PMID- 11121196 TI - Detection of expansion regions in Portuguese bipolar families. AB - We have studied 24 families with multiple affected members with bipolar disorder to test the hypothesis that in those families clinically showing genetic anticipation [Macedo et al., 1999] we would find large repeat expansions. The families meeting inclusion criteria had a minimum of two affected members over two generations and showed marked anticipation both in terms of age of onset and disease severity. We used the repeat expansion detection (RED) method to test patients (n = 24) and controls from these families and unrelated controls (n = 53). We also genotyped patients and family members from two families with large expansions at the known expansion loci on chromosomes 13, 17, and 18. The RED method revealed a higher number of large expansions in patients compared with controls (t-test; P < 0.0055: Mann-Whitney U; P = 0.02). The patients with the largest expansions were typed at the specific loci on chromosomes 13, 17, and 18 and the chromosome 18 expansion locus segregated with disease in one family, and a second family showed segregation with the expansion located at the SCA8 locus on chromosome 13. Genetic anticipation had been analyzed in this cohort of families, with correction for potential ascertainment bias, possible proband effects, cohort effects, regression to the mean, gender effects, and maternal vs. paternal transmission. None of these potential confounds appeared to account for the observed anticipation. We also identified that the presence of large expansions in affected family members derives primarily from two families from the genetically isolated Azores population. One family shows segregation with the chromosome 18 locus, whereas the other family segregates with expansions at the SCA8 locus. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:854-857, 2000. PMID- 11121197 TI - Failure to replicate an association between the catechol-O-methyltransferase polymorphism and attention deficit hyperactivity disorder in a second, independently recruited Israeli cohort. AB - Attention deficit hyperactivity disorder (ADHD) is a developmental syndrome expressed along three domains: inattention, hyperactive-impulsive, and combined type. Both environmental and genetic factors contribute to the etiology of this complex disease. We previously reported an association in 48 ADHD triads (both parents and proband) between the catechol-O-methyl- transferase (COMT) polymorphism (especially the high enzyme activity val allele) and the Diagnostic and Statistical Manual of Mental Disorders (DSM IV) combined category (excluding inattention) of ADHD (however, see erratum, Am. J. Med. Genet. [Neuropsychiatr. Genet.] 96:893). In the current report, we attempted to replicate this finding in an independently recruited group of 70 nuclear families using the haplotype relative risk design. In the current investigation, no evidence for association of the COMT polymorphism and ADHD (or any of the DSM IV subtypes) was observed in either the current cohort or the expanded cohort of 118 Israeli triads. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:858-860, 2000. PMID- 11121198 TI - Tryptophan hydroxylase gene polymorphisms are not associated with suicide. AB - Several lines of evidence suggest a serotonergic dysfunction involved in the biological susceptibility of suicide. Abnormalities of serotonergic markers such as 5-hydroxyindoleacetic acid and prolactin response to fenfluramine have been demonstrated in suicide subjects. Tryptophan hydroxylase (TPH), the rate-limiting enzyme in serotonin biosynthesis, is one of the most important regulating factors in the serotonergic system. Recently, polymorphisms of the TPH gene have been identified and some of these polymorphisms have been suggested to be associated with suicide, but the results are still inconsistent. We examined whether the 6526A/G polymorphism in the promoter region and the 218A/C polymorphism in intron 7 of the TPH gene were associated with suicide using 132 Japanese suicide victims. No significant difference in genotype distribution and allele frequencies of these polymorphisms was found between the suicide victims and the controls. We concluded neither the -6526A/G polymorphism nor the 218A/C polymorphism of the TPH gene is likely to have a major effect on the susceptibility of suicide. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:861 863, 2000. PMID- 11121199 TI - Second stage of a genome scan of schizophrenia: study of five positive regions in an expanded sample. AB - In a previous genome scan of 43 schizophrenia pedigrees, nonparametric linkage (NPL) scores with empirically derived pointwise P-values less than 0.01 were observed in two regions (chromosomes 2q12-13 and 10q23) and less than 0.05 in three regions (4q22-23, 9q22, and 11q21). Markers with a mean spacing of about 5 cM were typed in these regions in an expanded sample of 71 pedigrees, and NPL analyses carried out. No region produced significant genomewide evidence for linkage. On chromosome 10q, the empirical P-value remained at less than 0.01 for the entire sample (D10S168), evidence in the original 43 pedigrees was slightly increased, and a broad peak of positive results was observed. P-values less than 0.05 were observed on chromosomes 2q (D2S436) and 4q (D4S2623), but not on chromosomes 9q or 11q. It is concluded that this sample is most supportive of linkage on chromosome 10q, with less consistent support on chromosomes 2q and 4q. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:864-869, 2000. PMID- 11121200 TI - Human leukocyte antigen (HLA) DRB1 alleles in Kuwaiti Arabs with schizophrenia. AB - The frequency of human leukocyte anti- gen DRB1 alleles was determined in a cohort of 194 Kuwaiti Arabs consisting of 80 schizophrenia patients and 114 ethnically matched healthy controls, using a polymerase chain reaction-sequence specific primers method. A total of 12 DRB1 alleles were identified in this Kuwaiti cohort. A statistically significant difference was detected in the frequency of alleles DRB1(*)04 and DRB1(*)13 between the schizophrenia patients and controls. Allele frequency of DRB1(*)04 in schizophrenia patients was 14% compared with nearly 7% in controls (P = 0.028). For DRB1(*)13, the allele frequency was found to be 18% in schizophrenia patients compared with 9% in the controls (P = 0.015). For alleles, DRB1(*)03, DRB1(*)07, and DRB1(*)16 the frequency was higher in controls compared with schizophrenia patients. The frequency of DRB1(*)01, DRB1(*)08, DRB1(*)10, DRB1(*)11, and DRB1(*)15 alleles was almost identical in schizophrenia patients and controls. For the remaining alleles, the differences between the two groups were not statistically significant. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:870-872, 2000. PMID- 11121201 TI - Long repeat tracts at SCA8 in major psychosis. AB - Expansion at a recently identified unstable trinucleotide repeat on chromosome 13q21 has been reported as the molecular cause for spinocerebellar ataxia type 8 (SCA8). The trinucleotide repeat, which consists of a [CTA]n repeat and adjacent [CTG]n repeat, was reported to have a pathogenic range of 107-127 CTG repeats (or 110-130 combined CTA and CTG repeats) in a large ataxia kindred. This repeat region was also cloned by our group from a bipolar affective disorder (BPAD) patient, who has approximately 600 combined repeats, and large alleles (>100 repeats) were reported to be present in 0.7% of controls and 1.5% of major psychosis patients (n = 710 and n = 1,120, respectively). We have followed up these findings by screening three new samples of BPAD and schizophrenia (SCZ) patients and controls, including 272 individuals from 14 BPAD families from Sweden, 130 individuals from 32 SCZ and BPAD families/trios from the Azores Islands, and 206 SCZ individuals from the United Kingdom and Ireland, and 219 matched controls. We found large repeat alleles above the SCA8 pathogenic range in individuals from 3 of 32 Azorean pedigrees and in 1 of 206 SCZ individuals from the United Kingdom, and repeat alleles within the SCA8 pathogenic range in 1 of 14 Swedish families. Although the rarity of major psychosis patients carrying the SCA8 expansion mutation would require a much larger sample size to reach statistical significance, these results support the previously reported observation of increased occurrence of large repeats at SCA8 in major psychosis. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:873-876, 2000. PMID- 11121202 TI - No association between suicidal behavior and 5-HT2A-T102C polymorphism in alcohol dependents. PMID- 11121203 TI - No evidence for association of 5-HT2A receptor polymorphism with suicide. PMID- 11121204 TI - Linkage mapping of bipolar affective disorder. PMID- 11121205 TI - Comparison of the number of triplets in SCA1, MJD/SCA3, HD, SBMA, DRPLA, MD, FRAXA and FRDA genes in schizophrenic patients and a healthy population. PMID- 11121206 TI - Behavioral phenotype in childhood type of dystrophia myotonica. PMID- 11121207 TI - MAO-B gene intronic dinucleotide repeat polymorphism revisited. PMID- 11121208 TI - Suicidal behavior, the serotonin 2A receptor gene, and the media. PMID- 11121209 TI - Erratum PMID- 11121210 TI - Modulation of radiation-induced cytokine elevation associated with esophagitis and esophageal stricture by manganese superoxide dismutase-plasmid/liposome (SOD2 PL) gene therapy. AB - Radiation of the esophagus of C3H/HeNsd mice with 35 or 37 Gy of 6 MV X rays induces significantly increased RNA transcription for interleukin 1 (Il1), tumor necrosis factor alpha (Tnf), interferon gamma inducing factor (Ifngr), and interferon gamma (Ifng). These elevations are associated with DNA damage that is detectable by a comet assay of explanted esophageal cells, apoptosis of the esophageal basal lining layer cells in situ, and micro-ulceration leading to dehydration and death. The histopathology and time sequence of events are comparable to the esophagitis in humans that is associated with chemoradiotherapy of non-small cell lung carcinoma (NSCLC). Intraesophageal injection of clinical grade manganese superoxide dismutase-plasmid/liposome (SOD2-PL) 24 h prior to irradiation produced an increase in SOD2 biochemical activity in explanted esophagus. An equivalent therapeutic plasmid weight of 10 microgram ALP plasmid in the same 500 microliter of liposomes, correlated to around 52-60% of alkaline phosphatase-positive cells in the squamous layer of the esophagus at 24 h. Administration of SOD2-PL prior to irradiation mediated a significant decrease in induction of cytokine mRNA by radiation and decreased apoptosis of squamous lining cells, micro-ulceration, and esophagitis. Groups of mice receiving 35 or 37 Gy esophageal irradiation by a technique protecting the lungs and treating only the central mediastinal area were followed to assess the long-term effects of radiation. SOD2-PL-treated irradiated mice demonstrated a significant decrease in esophageal wall thickness at day 100 compared to irradiated controls. Mice with orthotopic thoracic tumors composed of 32D-v-abl cells that received intraesophageal SOD2-PL treatment showed transgenic mRNA in the esophagus at 24 h, but no detectable human SOD2 transgene mRNA in explanted tumors by nested RT PCR. These data provide support for translation of this strategy of SOD2-PL gene therapy to studies leading to a clinical trial in fractionated irradiation to decrease the acute and chronic side effects of radiation-induced damage to the esophagus. PMID- 11121211 TI - Estimation of oxygen distribution in RIF-1 tumors by diffusion model-based interpretation of pimonidazole hypoxia and eppendorf measurements. AB - Numerical simulations of oxygen diffusion from the capillaries in tumor tissue were used to predict the capillary oxygen supply within and near hypoxic regions of the RIF-1 tumor. A finite element method to simulate the oxygen distribution from a histology section is presented, along with a method to iteratively estimate capillary oxygen concentrations. Pathological structural data for these simulations came from sections of the tumor stained with hematoxylin and eosin and were used to define the capillary positions and shapes, while overlapping regions of low oxygen concentration were defined by the hypoxia marker pimonidazole. These simulations were used to calculate spatial maps of the oxygen concentration and were tested for their ability to reproduce Eppendorf pO(2) histograms from the same tumor line. This simulation study predicted that capillary oxygen concentrations ranged from zero to above 20 microM, with a dominant peak in the hypoxic regions showing 78% of capillaries with less than 1 microM oxygen concentration, compared to only 12% in the non-hypoxic regions. The results were not highly sensitive to the metabolic oxygen consumption rate, within the range of 2 to 16 microM/s. This numerical method for oxygen capillary simulation is readily adaptable to histology sections and provides a method to examine the heterogeneity of oxygen within the capillaries and throughout the tumor tissue section being examined. PMID- 11121212 TI - Fractionated irradiation combined with carbogen breathing and nicotinamide of two human glioblastomas grafted in nude mice. AB - This study addressed the potential radiosensitizing effect of nicotinamide and/or carbogen on human glioblastoma xenografts in nude mice. U-87MG and LN-Z308 tumors were irradiated with either 20 fractions over 12 days or 5 fractions over 5 days in air-breathing mice, mice injected with nicotinamide, mice breathing carbogen, or mice receiving nicotinamide plus carbogen. The responses to treatment were assessed using local control and moist desquamation. In U-87MG tumors, the enhancement ratios (ERs) at the radiation dose required to produce local tumor control in 50% of the treated mice (TCD(50)) with nicotinamide and/or carbogen ranged from 1.13 to 1.24 for irradiation in 20 fractions over 12 days. In LN-Z308 tumors, the ERs at the TCD(50) with nicotinamide and/or carbogen ranged from 1.22 to 1.40 for irradiation in 5 fractions over 5 days and from 1.11 to 1.30 in 20 fractions over 12 days, respectively. Skin injury was slightly enhanced, with ERs ranged from 1.06 to 1.15 when radiation was combined with carbogen and/or nicotinamide. Thus carbogen and nicotinamide can slightly improve the radiation response of human glioblastoma xenografts. PMID- 11121213 TI - Response of thyroid follicular cells to gamma irradiation compared to proton irradiation. I. Initial characterization of DNA damage, micronucleus formation, apoptosis, cell survival, and cell cycle phase redistribution. AB - The RBE of protons has been assumed to be equivalent to that of photons. The objective of this study was to determine whether radiation-induced DNA and chromosome damage, apoptosis, cell killing and cell cycling in organized epithelial cells was influenced by radiation quality. Thyroid-stimulating hormone dependent Fischer rat thyroid cells, established as follicles, were exposed to gamma rays or proton beams delivered acutely over a range of physical doses. Gamma-irradiated cells were able to repair DNA damage relatively rapidly so that by 1 h postirradiation they had approximately 20% fewer exposed 3' ends than their counterparts that had been irradiated with proton beams. The persistence of free ends of DNA in the samples irradiated with the proton beam implies that either more initial breaks or a quantitatively different type of damage had occurred. These results were further supported by an increased frequency of chromosomal damage as measured by the presence of micronuclei. Proton-beam irradiation induced micronuclei at a rate of 2.4% per gray, which at 12 Gy translated to 40% more micronuclei than in comparable gamma-irradiated cultures. The higher rate of micronucleus formation and the presence of larger micronuclei in proton-irradiated cells was further evidence that a qualitatively more severe class of damage had been induced than was induced by gamma rays. Differences in the type of damage produced were detected in the apoptosis assay, wherein a significant lag in the induction of apoptosis occurred after gamma irradiation that did not occur with protons. The more immediate expression of apoptotic cells in the cultures irradiated with the proton beam suggests that the damage inflicted was more severe. Alternatively, the cell cycle checkpoint mechanisms required for recovery from such damage might not have been invoked. Differences based on radiation quality were also evident in the alpha components of cell survival curves (0.05 Gy(-1) for gamma rays, 0.12 Gy(-1) for protons), which suggests that the higher level of survival of gamma-irradiated cells could be attributed to the persistence of nonlethally irradiated thyrocytes and/or the capacity to repair damage more effectively than cells exposed to equal physical doses of protons. The final assessment in this study was radiation-induced cell cycle phase redistribution. Gamma rays and protons produced a similar dose dependent redistribution toward a predominantly G(2)-phase population. From our cumulative results, it seems likely that a majority of the proton-irradiated cells would not continue to divide. In conclusion, these findings suggest that there are quantitative and qualitative differences in the biological effects of proton beams and gamma rays. These differences could be due to structured energy deposition from the tracks of primary protons and the associated high-LET secondary particles produced in the targets. The results suggest that a simple dose-equivalent approach to dosimetry may be inadequate to compare the biological responses of cells to photons and protons. PMID- 11121214 TI - Down-regulation of the human CDC16 gene after exposure to ionizing radiation: a possible role in the radioadaptive response. AB - We have used the method of differential display of mRNAs to search for changes in gene expression associated with radioadaptation triggered by low doses of ionizing radiation in human lymphoblasts. We isolated a cDNA designated PB13 that was down-regulated as early as 1 h after irradiation with 4 Gy in cells adapted by a pre-exposure to a dose of 2 cGy, compared to 3 h in nonadapted cells (4 Gy alone). Northern analysis confirmed the differential expression of a 2.4-kb transcript that was repressed for at least 10 h after irradiation. The major part of the PB13 cDNA was identical to the human CDC16 mRNA. When using either PB13 or CDC16 cDNA as probes, similar radiation-induced alterations in gene expression were observed. Expression of the CDC16 gene was also repressed after oxidative stress with H(2)O(2). The CDC16 protein belongs to the anaphase-promoting complex (APC) that controls progression through mitosis. The repression of expression of the CDC16 gene by ionizing radiation could result in delayed progression of damaged cells through mitosis. This cycle delay would occur earlier in adapted cells and would allow a more rapid and efficient repair that could contribute to the tolerance to subsequent irradiation. PMID- 11121215 TI - G2-phase delays after irradiation and/or heat treatment as assessed by two parameter flow cytometry. AB - Similar to what has been observed after irradiation, the fraction of G(2)-phase cells increases as a consequence of heat treatment. On the basis of cell cycle distributions alone, however, it is difficult to say whether the two results are related. In particular, comparison is complicated by the fact that the accompanying changes in the S-phase transition are different. These changes play a minor role after irradiation but constitute by far the most important cell cycle effect after heat treatment. Two-parameter flow cytometry was used here to study the proliferation of human melanoma cells in vitro. Cultures were pulse labeled with BrdU after irradiation and/or heat treatment and were fixed either immediately or after a delay of up to 36 h. DNA-synthesizing cells were identified with the help of an FITC-conjugated antibody against BrdU; DNA was quantified after staining with propidium iodide. In this way, the cell cycle distribution could be determined and the progression through the cell cycle could be analyzed. From the movement of labeled cells through the cycle, in particular the appearance of labeled cells in the G(1) compartment (after they had gone through mitosis), the delay in G(2) phase could be determined. The duration of the G(2)/M phase in control cells was about 6 h. This was increased to 12, 13 and 16 h after irradiation (4 Gy X rays), heat treatment (1 h at 43 degrees C), and a combination of the two, respectively. In all these cases, the G(2)-phase block was completely overcome within 48 h after treatment, whereas changes in the S phase were still observable at this time. As expected, the radiation-induced G(2) phase block was almost completely removed by incubating the cells with 5 or 10 mM caffeine. In the case of hyperthermia alone or in combination with radiation, however, caffeine was somewhat less effective. This does not mean, however, that the mechanisms involved are necessarily different. It can also be seen as a result of the differences in the time course of events. The long delay in S phase after heat treatment may lead to a loss of susceptibility to caffeine by the time the cells move into the G(2) phase. PMID- 11121216 TI - Comparison of F ratios generated from interphase and metaphase chromosome damage induced by high doses of low- and high-LET radiation. AB - Although biophysical models predict a difference in the ratio of interchromosomal to intrachromosomal interarm exchanges (F ratio) for low- and high-LET radiations, few experimental data support this prediction. However, the F ratios in experiments to date have been generated using data on chromosome aberrations in samples collected at the first postirradiation mitosis, which may not be indicative of the aberrations formed in interphase after exposure to high-LET radiations. In the present study, we exposed human lymphocytes in vitro to 2 and 5 Gy of gamma rays and 3 Gy of 1 GeV/nucleon iron ions (LET = 140 keV/micrometer), stimulated the cells to grow with phytohemagglutinin (PHA), and collected the condensed chromosomes after 48 h of incubation using both chemically induced premature chromosome condensation (PCC) and the conventional metaphase techniques. The PCC technique used here condenses chromosomes mostly in the G(2) phase of the cell cycle. The F ratio was calculated using data on asymmetrical chromosome aberrations in both the PCC and metaphase samples. It was found that the F ratios were similar for the samples irradiated with low- and high-LET radiation and collected at metaphase. However, for irradiated samples assayed by PCC, the F ratio was found to be 8.2 +/- 2.0 for 5 Gy gamma rays and 5.2 +/- 0.9 for 3 Gy iron ions. The distribution of the aberrations indicated that, in the PCC samples irradiated with iron ions, most of the centric rings occurred in spreads containing five or more asymmetrical aberrations. These heavily damaged cells, which were either less likely to reach mitosis or may reach mitosis at a later time, were responsible for the difference in the F ratios generated from interphase and metaphase analysis after exposure to iron ions. PMID- 11121217 TI - Chromosome translocations in T. scripta: the dose-rate effect and in vivo lymphocyte radiation response. AB - Using a whole-chromosome FISH painting probe we previously developed for chromosome 1 of the yellow-bellied slider turtle (Trachemys scripta), we investigated the dose-rate effect for radiation-induced symmetrical translocations in T. scripta fibroblasts and lymphocytes. The dose rate below which no reduction in effect per unit dose is observed with further dose protraction was approximately 23 cGy h(-1). We estimated the whole-genome spontaneous background level of complete, apparently simple symmetrical translocations in T. scripta lymphocytes to be approximately 1.20 x 10(-3)/cell projected from aberrations occurring in chromosome 1. Similar spontaneous background levels reported for humans are some 6- to 25-fold higher, ranging from about 6 x 10(-3) to 3.4 x 10(-2) per cell. This relatively low background level for turtles would be a significant advantage for resolution of effects at low doses and dose rates. We also chronically irradiated turtles over a range of doses from 0-8 Gy delivered at approximately 5.5 cGy h(-1) and constructed a lymphocyte dose-response curve for complete, apparently simple symmetrical translocations suitable for use with animals chronically exposed to radiation in contaminated environments. The best-fitting calibration curve (not constrained through the zero dose estimate) was of the form Y(as) = c + aD + bD(2), where Y(as) was the number of apparently simple symmetrical translocations per cell, D was the dose (Gy), a = (0.0058 +/- 0.0009), b = (-0.00033 +/- 0.00011), and c = (0.0015 +/- 0.0013). With additional whole-chromosome probes to improve sensitivity, environmental biodosimetry using stable chromosome translocations could provide a practical and genetically relevant measurement end point for ecological risk assessments and biomonitoring programs. PMID- 11121218 TI - Children of chernobyl cleanup workers do not show elevated rates of mutations in minisatellite alleles. AB - The disaster at the Chernobyl Nuclear Power Plant in April 1986 was accompanied by the release of large amounts of radioisotopes, resulting in the contamination of extensive regions of the Ukraine, Byelorus and the Russian Federation. Cleanup workers (liquidators) and people living on land contaminated with radioactive materials were most exposed. To assess the genetic effects of exposure to ionizing radiation after the Chernobyl accident, we have measured the frequency of inherited mutant alleles at seven hypermutable minisatellite loci in 183 children born to Chernobyl cleanup workers (liquidators) and 163 children born to control families living in nonirradiated areas of the Ukraine. There was no significant difference in the frequency of inherited mutant alleles between the exposed and control groups. The exposed group was then divided into two subgroups according to the time at which the children were conceived with respect to the fathers' work at the power plant. Eighty-eight children were conceived either while their fathers were working at the facility or up to 2 months later (Subgroup 1). The other 95 children were conceived at least 4 months after their fathers had stopped working at the Chernobyl site (Subgroup 2). The frequencies of mutant alleles were higher for the majority of loci (i.e. 1.44 times higher for CEB1) in Subgroup 1 than in Subgroup 2. This result, if confirmed, would reconcile the apparently conflicting results obtained in the chronically exposed Byelorus population and the Hiroshima-Nagasaki A-bomb survivors. PMID- 11121219 TI - T-cell responses to mitogens in atomic bomb survivors: a decreased capacity to produce interleukin 2 characterizes the T cells of heavily irradiated individuals. AB - Significant decreases in the fraction of lymphocytes that are CD4(+) and increases in serum levels of some classes of immunoglobulin have been reported to occur in atomic bomb (A-bomb) survivors and in victims of the Chernobyl nuclear plant accident. To investigate the long-term effects of nuclear radiation on cellular immunity in more detail, we used limiting dilution assays with peripheral blood mononuclear cell preparations to analyze the T-cell responses of 251 A-bomb survivors exposed to less than 0.005 Gy and 159 survivors exposed to more than 1.5 Gy. The percentages of CD2-positive cells that were capable of proliferating in response to phytohemagglutinin (PHA) in the presence of exogenous interleukin 2 (IL2) did not differ substantially between distally exposed and more heavily exposed survivors. The heavily exposed survivors appeared to possess fewer T cells that were capable of proliferating in response to concanavalin A (Con A) or of producing interleukin 2. Assuming that CD4 T cells were the ones primarily responsible for producing IL2 in response to Con A, we were able to estimate how many cells in any given CD4 T-cell population were actually producing IL2. The results indicated that peripheral blood samples from heavily exposed survivors contained significantly fewer IL2-producing CD4 T cells than did similar samples from distally exposed survivors, indicating that significant exposure to A-bomb radiation may have a long-lasting negative effect on the capacity of CD4 T-cell populations to produce IL2. PMID- 11121220 TI - Microdosimetry of a 25 keV electron microbeam. AB - Electron microbeam experiments are planned or under way to explore in part the question regarding whether the bystander effect is a general phenomenon or is restricted to high-LET radiation. Since low-LET radiations scatter more readily compared to high-LET radiations, identifying bystander cells and assessing the potential dose that they may receive will be crucial to the interpretation of radiobiological results. This paper reports on initial calculations of the basic information needed for a stochastic model of the penetration of energetic electrons in tissue-like matter; the model will be used to predict doses delivered to adjacent regions in which bystander cells may reside. Results are presented of calculations of the stochastics of energy deposition by 25 keV electrons slowing down in a homogeneous water medium. Energy deposition distributions were scored for 1-micrometer spheres located at various penetration and radial distances up to 10 micrometer from the point of origin. The energy of 25 keV was selected because experiments are planned for that energy. At 25 keV there is a high probability that the entire electron track will be contained within a typical mammalian cell. Individual tracks are scored because of their primacy; data for higher doses can be obtained by convoluting single-track distributions. The event frequency decreases approximately exponentially after the first micrometer to 1% at about 8 micrometer of penetration. Radially, the 1% contour extends to 3.5 micrometer at a penetration of 5.5 micrometer. The frequency-mean energy deposited decreases from 1.5 to 1 keV/micrometer at a penetration of 3.5 micrometer, then increases back to about 1.5 at a penetration of 6.5 micrometer. The mean energy increases to about 3 keV/micrometer at a radial distance of 8.5 micrometer. PMID- 11121221 TI - The toxicity of insoluble cerium-144 inhaled by beagle dogs: non-neoplastic effects. AB - The biological effects of inhaled beta-particle-emitting radionuclides are not well known. The non-neoplastic diseases induced by an inhaled, relatively insoluble form of cerium-144 ((144)Ce) were studied in beagle dogs exposed to graded activity levels of (144)Ce in fused aluminosilicate particles by a single, brief inhalation exposure and observed for their life span. The initial lung burdens (ILBs) achieved ranged from 0.000093-7.6 MBq (144)Ce/kg body weight. The (144)Ce was retained in the lung with an effective half-life of about 190 days. Significant (144)Ce was translocated to the tracheobronchial lymph nodes, and the concentration exceeded that of the lung at about 400 days after inhalation exposure. Significant radiation doses were delivered to the lung and tracheobronchial lymph nodes and to the heart adjacent to the tracheobronchial lymph nodes. Radiation pneumonitis was the predominant non-neoplastic disease. The dose response for radiation pneumonitis indicated that an ILB of 1.4 MBq/kg would cause death from radiation pneumonitis in 50% of the exposed dogs. This ILB resulted in a pulmonary dose to death of about 350 Gy. The tracheobronchial lymph nodes developed lesions in dogs with ILBs lower than those causing radiation pneumonitis. The overall results of this study, however, showed that (144)Ce, inhaled in an insoluble form, did not cause any unique or inexplicable biological effects in dogs or cause effects at unusually low doses that might call current radiation protection guidelines into question. PMID- 11121222 TI - Cytogenetic studies in human blood lymphocytes exposed in vitro to radiofrequency radiation at a cellular telephone frequency (835.62 MHz, FDMA). AB - Freshly collected peripheral blood samples from four healthy human volunteers were diluted with RPMI 1640 tissue culture medium and exposed in sterile T-75 tissue culture flasks in vitro for 24 h to 835.62 MHz radiofrequency (RF) radiation, a frequency employed for customer-to-base station transmission of cellular telephone communications. An analog signal was used, and the access technology was frequency division multiple access (FDMA, continuous wave). A nominal net forward power of 68 W was used, and the nominal power density at the center of the exposure flask was 860 W/m(2). The mean specific absorption rate in the exposure flask was 4.4 or 5.0 W/kg. Aliquots of diluted blood that were sham exposed or exposed in vitro to an acute dose of 1.50 Gy of gamma radiation were used as negative or positive controls. Immediately after the exposures, the lymphocytes were stimulated with a mitogen, phytohemagglutinin, and cultured for 48 or 72 h to determine the extent of genetic damage, as assessed from the frequencies of chromosomal aberrations and micronuclei. The extent of alteration in the kinetics of cell proliferation was determined from the mitotic indices in 48-h cultures and from the incidence of binucleate cells in 72-h cultures. The data indicated no significant differences between RF-radiation- and sham-exposed lymphocytes with respect to mitotic indices, incidence of exchange aberrations, excess fragments, binucleate cells, and micronuclei. In contrast, the response of the lymphocytes exposed to gamma radiation was significantly different from both RF-radiation- and sham-exposed cells for all of these indices. Thus, under the experimental conditions tested, there is no evidence for the induction of chromosomal aberrations and micronuclei in human blood lymphocytes exposed in vitro for 24 h to 835.62 MHz RF radiation at SARs of 4.4 or 5.0 W/kg. PMID- 11121223 TI - Long-term genomic instability in human lymphocytes induced by single-particle irradiation. AB - Recent evidence suggests that genomic instability, which is an important step in carcinogenesis, may be important in the effectiveness of radiation as a carcinogen, particularly for high-LET radiations. Understanding the biological effects underpinning the risks associated with low doses of densely ionizing radiations is complicated in experimental systems by the Poisson distribution of particles that can be delivered. In this study, we report an approach to determine the effect of the lowest possible cellular radiation dose of densely ionizing alpha particles, that of a single particle traversal. Using microbeam technology and an approach for immobilizing human T-lymphocytes, we have measured the effects of single alpha-particle traversals on the surviving progeny of cells. A significant increase in the proportion of aberrant cells is observed 12 13 population doublings after exposure, with a high level of chromatid-type aberrations, indicative of an instability phenotype. These data suggest that instability may be important in situations where even a single particle traverses human cells. PMID- 11121224 TI - Molecular and genetic mechanisms of prostate cancer. AB - Prostate cancer is the most commonly diagnosed malignancy in American men and is the second leading cause of cancer death in males in the United States. Despite its high incidence, the molecular and genetic events involved in progression of prostate cancer remain poorly understood. In vitro models of human prostate epithelial (HPE) cells provide a practical approach to the analysis of the molecular and genetic mechanisms underlying prostate carcinogenesis. We reported the immortalization of normal adult HPE cells by transfection of the HPV-18 DNA and the subsequent conversion of such nontumorigenic but immortalized cells (HPV 18 C-1) into tumorigenic cells by the introduction of an activated Kras oncogene. Recently, we have demonstrated the malignant transformation of HPV-18 C-1 cells after multiple exposures to the chemical carcinogen N-nitroso-N-methylurea (NMU). Such transformants showed morphological alterations and anchorage-independent growth in soft agar and induced carcinomas when transplanted into nude mice. No TP53 or RAS mutations were observed. Stepwise chromosomal changes in the progression to tumorigenicity were observed. Loss of the p arms of chromosome 8 (p10>pter) and chromosome 10(p10>pter) and gain of the q arm of chromosome 8 (p10>ptr) (the most frequent cytogenetic changes observed directly in prostate cancer patients) were observed only in the tumor outgrowths. These findings provide the first evidence of malignant transformation of HPE cells exposed to a chemical carcinogen. PMID- 11121225 TI - In vitro models to study cellular differentiation and function in human prostate cancers. AB - In Vitro Models to Study Cellular Differentiation and Function in Human Prostate Cancers. To augment the currently available models of human prostate cancer in vitro, we have established extended life-span epithelial cultures from biopsies of well-differentiated prostate cancers. The genetic identity of the target cells was assessed by allelotyping, using microsatellites located on chromosome 8p, and microdissection of tissues and primary cell cultures. Cells with an extended life span (PxE6) were derived by recombinant retrovirus infection to introduce the human papilloma virus E6 gene (epithelial cells). Immunophenotyping of the resultant cell strains confirmed retention of differentiated cell functions, and the genotype of the E6-expressing epithelial cells was stable, while SV40 immortalized cultures were more unstable, leading to tetraploidy. All PxE6 cells eventually senesced, but an immortalized epithelial culture, P4E6, was derived from one of the epithelial cultures. The properties of this cell line, which remains close to diploid, are similar to those of early prostate cancer cells, and it retains expression of many prostate-associated antigens, such as prostate specific antigen (PSA). PMID- 11121226 TI - Multiple genetic changes are required for efficient immortalization of different subtypes of normal human mammary epithelial cells. AB - Multiple Genetic Changes Are Required for Efficient Immortalization of Different Subtypes of Normal Human Mammary Epithelial Cells. Breast cancer is the second leading cause of cancer-related deaths of women in the U.S. About 180,000 new cases of breast cancer are diagnosed each year, a quarter of them fatal. Early detection is the key to the survival of these patients. However, there are no molecular markers to detect breast cancer at very early stages. A hurdle in understanding the early molecular changes in breast cancer has been the difficulty in establishing premalignant lesions and primary breast tumors as in vitro cell cultures. Normal epithelial cells grow for a finite life span and then senesce. Immortalization is defined by continuous growth of otherwise senescing cells and is believed to represent an early stage in tumor progression. To examine these early stages, we and others have developed in vitro models of mammary epithelial cell immortalization. These models have been extremely important in understanding the role of various tumor suppressor pathways that maintain the normal phenotypes of mammary epithelial cells. In this paper, we describe the establishment of these models and their relevance to understanding the molecular changes that occur in early breast cancer. These models have helped to identify molecular changes that occur in early breast cancers and appear to be well suited to identify novel markers for early diagnosis of breast cancer. PMID- 11121227 TI - The pathway of neoplastic transformation of human breast epithelial cells. AB - The morphological analysis of breast cancer development indicates this to be a multistep process that progressively evolves from ductal hyperplasia and atypical ductal hyperplasia, which represent the initial stages of neoplastic growth, to carcinoma in situ, invasive carcinoma, and ultimately metastasis, as has been documented for a number of other malignancies. The understanding of the cellular and molecular processes that lead a normal cell to malignancy requires the analysis of pure populations of human breast epithelial cells (HBEC) representing specific stages of neoplastic progression. The neoplastic transformation of HBEC in vitro represents a successful model for obtaining knowledge about the molecular and biological alterations that may contribute to the tumorigenic mechanisms. We present here a current understanding of chemically transformed HBEC in the following aspects: (1) factors affecting the transformation of HBEC such as immortalization; (2) new targets for studying the mechanism of cell immortalization such as alterations in telomerase activity, differential expression of cell cycle-dependent genes, and others recently isolated through differential cloning, such as H-ferritin, and a calcium binding protein; (3) genetic mechanisms underlying cell transformation; and (4) application of the microcell-mediated chromosome transfer technique as an approach to testing the functional role of specific genes whose dysregulation or loss of function may contribute to the ultimate cell transformation. Further efforts in this cell system will be directed to determining the roles of identified molecular changes as well as the mapping/cloning of tumor suppressor or senescence genes. PMID- 11121228 TI - Vitamin D resistance in RAS-transformed keratinocytes: mechanism and reversal strategies. AB - Human retinoid X receptor alpha (hRXRalpha) plays a critical role in DNA binding and transcriptional activity through its heterodimeric association with several members of the nuclear receptor superfamily, including the vitamin D receptor (VDR). Several cancer cell lines derived from different tissues have been shown to be resistant to the growth-inhibitory action of 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)], the biologically active metabolite of vitamin D(3). Here we show that in RAS-transformed keratinocytes, Ser260 of hRXRalpha is phosphorylated through the RAS-RAF-MAP kinase cascade. This phosphorylation event results in the inhibition of vitamin D signaling via VDR/hRXRalpha heterodimers. Strategies to reverse this resistance include the use of the MAP kinase inhibitor, PD098059, and a non-phosphorylatable hRXRalpha mutant, Ala260, which completely abolishes RXR phosphorylation and restores the function of both 1,25(OH)(2)D(3) and a specific RXR ligand, LG1069 (4-[1-(5,6,7,8-tetrahydro-3,5,5,8,8-pentamethyl-2 naphtalenyl)ethenyl]-benzoic acid). In addition, we show that a vitamin D analog with low calcemic activity (EB1089) is more potent than 1,25(OH)(2)D(3) in inhibiting cancer cell growth in this system. Targeted therapy with selective analogs such as EB1089, in combination with the inhibition of phosphorylation of the RXR, could play a critical role in the development of strategies for cancer treatment. PMID- 11121229 TI - Suppression of depleted uranium-induced neoplastic transformation of human cells by the phenyl fatty acid, phenyl acetate: chemoprevention by targeting the p21RAS protein pathway. AB - Depleted uranium is a dense heavy metal used primarily in military applications. Published data from our laboratory have demonstrated that exposure to depleted uranium in vitro can transform immortalized human osteoblast (HOS) cells to the tumorigenic phenotype (associated with aberrant RAS oncogene expression and tumor suppressor protein production). Since depleted uranium is used in military applications, it would therefore be beneficial to identify and test potential antitumor-promoting agents. Chemopreventive interventions that target deregulated signal transduction pathways may be effective strategies to prevent carcinogenesis. Since the RAS protein plays a key role in signal transduction, disruption of its signaling pathway may be particularly effective. The phenyl fatty acid, phenyl acetate, a differentiation inducer that affects post translational processing of RAS, was tested for its ability to prevent depleted uranium-induced neoplastic transformation using HOS cells. After a 24-h exposure to insoluble depleted uranium-uranium dioxide (1 mg/ml), cells were incubated for 1 day to 6 weeks with 2.5 mM phenyl acetate. Treatment with depleted uranium resulted in transformation to the tumorigenic phenotype. In contrast, HOS cells exposed to depleted uranium and then treated with phenyl acetate did not exhibit transformation to the tumorigenic phenotype. These data suggest that depleted uranium-induced neoplastic transformation in vitro can be prevented by targeting the RAS protein. PMID- 11121230 TI - RNA damage and inhibition of neoplastic endothelial cell growth: effects of human and amphibian ribonucleases. AB - Angiogenesis defines the many steps involved in the growth and migration of endothelial cell-derived blood vessels. This process is necessary for the growth and metastasis of tumors, and considerable effort is being expended to find inhibitors of tumor angiogenesis. This usually involves screening of potential anti-angiogenic compounds on endothelial cells. To this end, two candidate anti angiogenic RNA-damaging agents, onconase and (-4)rhEDN, were screened for their effects on endothelial cell proliferation using three distinct types of endothelial cells in culture: HPV-16 E6/E7-immortalized human umbilical vein endothelial cells (HUVECs), a Kras-transformed HPV-16 E6/E7 HUVEC (Rhim et al., Carcinogenesis 4, 673-681, 1998), and primary HUVECs. Onconase similarly inhibited proliferation in all three cell lines (IC(50) = 0.3-1.0 microM) while ( 4)rhEDN was more effective on immortalized HUVEC cell lines (IC(50) = 0.02-0.06 microM) than on primary HUVECs (IC(50) > 0.1 microM). Differential sensitivity to these agents implies that more than one endothelial cell type must be used in proliferation assays to screen for novel anti-angiogenic compounds. PMID- 11121231 TI - Role of stem cells and gap junctional intercellular communication in human carcinogenesis. AB - Epidemiological data, experimental animal bioassays, studies of in vitro neoplastic transformation, and molecular oncology studies have implicated a multistage, multimechanism process in human carcinogenesis. From animal carcinogenesis studies, the operational concept of a single normal cell being irreversibly altered during the first step in carcinogenesis is called initiation. The subsequent interruptible or reversible clonal expansion of these initiated cells by non-cytotoxic mitogenic stimuli, compensatory hyperplasia due to cell death by necrosis, or inhibition of apoptosis is referred to as the promotion phase. Last, when one of these clonally expanded, initiated cells acquires sufficient genetic/epigenetic alterations to become neoplastically transformed and acquire the phenotypes of promoter independence, invasiveness and metastasis, it is referred to as the progression step of carcinogenesis. This report hypothesizes that the single normal cell that is initiated is a pluripotent stem cell. By assuming that the normal pluripotent stem cell is immortal and becomes mortal when induced to terminally differentiate, initiation would be viewed as the irreversible process by which a stable alteration in a finite number of proto-oncogenes and/or tumor suppressor genes could block terminal differentiation or "mortalization". Promotion would involve the reversible inhibition of gap junctional intercellular communication (GJIC) and while progression occurs with the stable down-regulation of GJIC. PMID- 11121232 TI - Expression of the protein product of the PCPH proto-oncogene in human tumor cell lines. AB - Expression of the Protein Product of the PCPH Proto-oncogene in Human Tumor Cell Lines. Exposure of Syrian hamster embryo fibroblasts to chemical carcinogens resulted in the oncogenic activation of the PCPH proto-oncogene by induction of a single base-pair deletion that generated a truncated PCPH oncoprotein (mutated PCPH). Recently, we isolated and characterized the cDNA for the human PCPH proto oncogene and determined that in humans PCPH is a single-copy gene located in chromosome 14 (14q24.3). Pilot mRNA expression studies indicated that PCPH was expressed in the majority of normal organs tested, particularly in liver and kidney, but it appeared to be expressed either at low levels or not at all in tumor cells or cell lines derived from the high-expressing tissues. We have generated an antiserum against bacterial recombinant Syrian hamster PCPH. This antiserum recognizes both the normal and truncated, oncogenic Syrian hamster PCPH proteins and cross-reacts with the yeast, mouse, rat and human homologue proteins. Using this antibody, we have performed a study of PCPH expression in a larger sample of human neoplastic cell lines, including some derived from breast, nervous system, colon, lung and pancreas tumors. Results confirmed the frequent lack of PCPH expression in malignant cells and identified several immunoreactive forms of PCPH being differentially expressed in cells of diverse tissue origins. PMID- 11121233 TI - Telomeres and telomerase: implications for cancer and aging. AB - Maintenance of telomere stability is required for cells to escape from replicative senescence and proliferate indefinitely. Telomere length is maintained by a balance between processes that lengthen telomeres (telomerase) and processes that shorten telomeres (the end-replication problem). Telomerase is a cellular ribonucleoprotein reverse transcriptase which stabilizes telomere length by adding hexameric (TTAGGG) repeats to the telomeric ends of the chromosomes, thus compensating for the continued erosion of telomeres. Introduction of the telomerase catalytic protein component into normal telomerase negative human cells results in restoration of telomerase activity and extension of cellular life span. Human cells with introduced telomerase maintain a normal chromosome complement and continue to grow in a normal manner. Telomerase-induced manipulations of telomere length may thus be important not only for cell and tissue engineering but also for dissecting the molecular mechanisms underlying inherited genetic diseases, as well as defining the genetic pathways leading to cancer. Because almost all human tumors express telomerase activity, inhibition of telomerase may result in gradual erosion of telomeres and eventual cessation of cell proliferation or induction of apoptosis. Thus telomerase may also be a promising target for cancer therapy. PMID- 11121234 TI - Alternative lengthening of telomeres in human cells. AB - Activation of a telomere maintenance mechanism appears to be essential for immortalization. In most human tumors and tumor cell lines, the telomere maintenance mechanism involves the activity of telomerase, a reverse transcriptase holoenzyme that synthesizes telomeric repeat DNA. In some cases, telomere maintenance occurs in the absence of telomerase activity by a mechanism referred to as alternative lengthening of telomeres (ALT). The development of telomere-targeted anticancer therapies will be facilitated by an understanding of the molecular mechanisms of ALT and of the means whereby ALT is repressed in normal cells. PMID- 11121235 TI - A human breast epithelial cell type with stem cell characteristics as target cells for carcinogenesis. AB - Two types of human breast epithelial cells (HBEC) have been characterized. In contrast to Type II HBEC, which express basal epithelial cell phenotypes, Type I HBEC are deficient in gap junctional intercellular communication and are capable of anchorage-independent growth and of expressing luminal epithelial cell markers, estrogen receptors, and stem cell characteristics (i.e. the ability to differentiate into other cell types and to form budding/ductal organoids on Matrigel). A comparative study of these two types of cells has revealed a high susceptibility of Type I HBEC to immortalization by SV40 large T antigen, although both types of cells are equally capable of acquiring an extended life span (bypassing senescence) after transfection with SV40. The immortalization was accompanied by elevation of a low level of telomerase activity in the parental cells after mid-passage ( approximately 60 cumulative population doubling levels). Thus HBEC do have a low level of telomerase activity, and Type I HBEC with stem cell characteristics are more susceptible to telomerase activation and immortalization, a mechanism which might qualify them as target cells for breast carcinogenesis. The immortalized Type I HBEC can be converted to highly tumorigenic cells by further treatment with X rays (2 Gy x 2) and transfection with a mutated ERBB2 (also known as NEU) oncogene, resulting in the expression of p185(ERBB2) which is tyrosine phosphorylated. PMID- 11121236 TI - Loss of nuclear localization of the S100C protein in immortalized human fibroblasts. AB - It is well known that cancer develops through a multistep process. In vitro transformation studies of normal human cells have shown that the immortalization step is critical for neoplastic transformation of cells. Furthermore, studies of cell fusion between normal and immortalized cells have indicated that the normal phenotype is dominant and the immortal phenotype is recessive. Thus we looked for cellular proteins that were down-regulated in immortalized human cells by two dimensional gel electrophoresis to elucidate the mechanisms of immortalization of human cells. We found that the S100C protein was down-regulated in immortalized cells. This protein was localized in the cytoplasm of cells at the semiconfluent stage, while at the confluent stage it moved into the nuclei of normal cells but not into those of immortalized cells. Microinjection of an S100C antibody into normal confluent cells diminished the level of nuclear S100C protein, resulting in DNA synthesis. Taken together, loss of nuclear localization of the S100C protein, which may be related to DNA synthesis, is thought to be one of the mechanisms of immortalization. PMID- 11121237 TI - Molecular events in radiation transformation. AB - Studies of human tumor cell lines have revealed alterations in the regulation of a number of cell cycle-related genes, associated in some cases with a TP53 independent loss of the radiation-induced G(1)-phase arrest. It is not clear, however, whether these are early or late events in tumor development, or they arise in tumor cell lines during growth in culture. Since the oncogenic transformation of an individual cell is thought to be an early event in tumor development, we have used a model system of normal and radiation-transformed C3H 10T(1/2) mouse fibroblast cell clones to address this issue. Transformed clones derived from type III foci were compared with clones derived from parental, wild type cells. Approximately 25% of transformed clones showed Trp53 mutations in exon 5; however, preliminary results based on in situ immunofluorescence studies with an antibody recognizing mutant Trp53 indicate that the appearance of such mutations in transformed clones occurs late in the process of transformation and is unlikely to represent an initiating event. The remaining transformed clones and all clones derived from parental cells expressed wild-type Trp53. Radiation induced G(1)-phase arrest was either absent or significantly reduced in all of the transformed clones, independent of Trp53 status. Constitutive expression of Cdkn1a protein was significantly increased in most of the transformed clones. Also, the majority of transformed clones showed elevated levels of cyclin D1, and two clones overexpressed cyclin E. These results indicate that loss of G(1)-phase checkpoint control, independent of Trp53 status, and altered expression of cell cycle regulatory proteins may represent early events in the process of radiation induced carcinogenesis that are associated with the malignant transformation of individual cells. PMID- 11121238 TI - Neoplastic transformation and cytogenetic changes after Gamma irradiation of human epithelial cells expressing telomerase. AB - Neoplastic transformation of human epithelial cells by radiation has previously been investigated using cell lines immortalized with viral vectors. There are disadvantages to this approach, and we report here the results of studies using a human retinal pigment epithelial cell line (340RPE-T53) immortalized by treatment with telomerase. After exposure of the cells to fractionated doses of gamma radiation, there was a marked increase in anchorage-independent growth of the surviving cells. The cloned cell lines derived from these anchorage-independent cultures exhibited an increased growth rate in vitro and were serum-independent compared with the parent cell line. The parent cell line maintained a stable diploid karyotype. The cell lines cloned after irradiation with the lower doses (10 x 2 Gy) were hypodiploid with loss of chromosome 13 and a high level amplification of 10p11.2 associated with a deletion of the remaining short arm segment of chromosome 10 distal to 10p11.2. In contrast, the cell lines cloned after irradiation with the higher doses (15 x 2 Gy) were near-tetraploid with derivative chromosomes present characterized by SKY analysis. Thus this human epithelial cell line immortalized with telomerase provides an improved model to investigate mechanisms of radiation carcinogenesis. PMID- 11121239 TI - Mechanisms of radiation-induced neoplastic transformation of human bronchial epithelial cells. AB - Carcinogenesis is a multistage process with sequences of genetic events that govern the phenotypic expression of a series of transformation steps that lead to the development of metastatic cancer. To better understand the mechanisms involved in human bronchial carcinogenesis induced by alpha particles from radon, we have developed a model of neoplastic transformation based on human papillomavirus-immortalized human bronchial epithelial (BEP2D) cells. Cells exposed to alpha particles become tumorigenic after progressing through a series of sequential stages including altered growth pattern, resistance to serum induced terminal differentiation, agar-positive growth, tumorigenicity, and metastasis, with each step representing a necessary yet insufficient step toward the later, more malignant phase. Cell fusion studies indicated that the radiation induced tumorigenic phenotype in BEP2D cells can be completely suppressed by fusion with nontumorigenic BEP2D cells. Several cellular differentiation and growth regulation genes such as DCC (deleted in colorectal cancer), CDKN1A (also known as p21(C1P1)) and the gene that encodes DNA-PK were frequently found to be modulated in tumorigenic BEP2D cells and may be related to the process of carcinogenesis. PMID- 11121240 TI - Perspective: cell differentiation theory may advance early detection of and therapy for lung cancer. AB - Many of these deaths could be prevented if there were better screening methods to uncover the disease when it is limited and most responsive to intervention. Novel biomarkers of early-stage disease are therefore needed. By applying the principle of "oncology recapitulates ontogeny", we have discovered three homeobox (HOX) genes that are inappropriately expressed in the majority of lung tumors. Understanding the role of these inappropriately expressed genes in lung epithelial cell carcinogenesis may not only augment early detection, but may also offer new avenues of treatment of this disease. PMID- 11121241 TI - Neoplastic transformation in C3H 10T(1/2) cells after exposure to 835.62 MHz FDMA and 847.74 MHz CDMA radiations. AB - The effect of radiofrequency (RF) radiation in the cellular phone communication range (835.62 MHz frequency division multiple access, FDMA; 847.74 MHz code division multiple access, CDMA) on neoplastic transformation frequency was measured using the in vitro C3H 10T(1/2) cell transformation assay system. To determine if 835.62 MHz FDMA or 847.74 MHz CDMA radiations have any genotoxic effects that induce neoplastic transformation, C3H 10T(1/2) cells were exposed at 37 degrees C to either of the above radiations [each at a specific absorption rate (SAR) of 0.6 W/kg] or sham-exposed at the same time for 7 days. After the culture medium was changed, the cultures were transferred to incubators and refed with fresh growth medium every 7 days. After 42 days, the cells were fixed and stained with Giemsa, and transformed foci were scored. To determine if exposure to 835.62 MHz FDMA or 847.74 MHz CDMA radiation has any epigenetic effects that can promote neoplastic transformation, cells were first exposed to 4.5 Gy of X rays to induce the transformation process and then exposed to the above radiations (SAR = 0.6 W/kg) in temperature-controlled irradiators with weekly refeeding for 42 days. After both the 7-day RF exposure and the 42-day RF exposure after X irradiation, no statistically significant differences in the transformation frequencies were observed between incubator controls, the sham exposed (maintained in irradiators without power to the antenna), and the 835.62 MHz FDMA or 847.74 MHz CDMA-exposed groups. PMID- 11121242 TI - Radiation-induced senescence-like growth arrest requires TP53 function but not telomere shortening. AB - Suzuki, K., Mori, I., Nakayama, Y., Miyakoda, M., Kodama, S. and Watanabe, M. Radiation-Induced Senescence-like Growth Arrest Requires TP53 Function but not Telomere Shortening. Normal human diploid cells irradiated with X rays showed permanent cell cycle arrest and exhibited senescence-like phenotypes including the expression of senescence-associated beta-galactosidase (SA-beta-gal). X irradiation caused persistent phosphorylation of TP53 at Ser 15 and accumulation of the TP53 protein, followed by the induction of CDKN1A (also known as p21(Waf1/Cip1)) and CDKN2A (also known as p16), preceded the expression of SA beta-gal. NCI-H1299 human lung carcinoma cells, in which no TP53 protein was expressed, were irradiated with X rays with or without the exogenous expression of TP53 gene. Although induction of TP53 protein alone could induce SA-beta-gal expression, the frequency of SA-beta-gal-positive cells was significantly increased when TP53-induced H1299 cells were exposed to X rays. The mean terminal restriction fragment length in normal human cells was approximately 12 kb and did not change in SA-beta-gal-positive cells. These results indicate that ionizing radiation induces senescence-like growth arrest that is dependent on TP53 function but independent of telomere shortening. Our findings suggest that cells harboring irreparable DNA damage are programmed to undergo premature senescence to maintain the integrity of the genome. PMID- 11121243 TI - Culture condition-dependent senescence-like growth arrest and immortalization in rodent embryo cells. AB - Culture Condition-Dependent Senescence-Like Growth Arrest and Immortalization in Rodent Embryo Cells. We investigated the telomerase activity, telomere length, and replicative life span of cells from human embryos and rodent embryos (mouse, rat and Syrian hamster). We used two culture conditions for rodent embryo cells whereby the cells were plated at a density of 2 x 10(5) into a 25-cm(2) flask and subcultured every 3 days or every 10 days. We found that nearly 100% of the cultures of rodent embryo cells become immortal when they are subcultured using the 10-day culture protocol. These rodent embryo cells retain telomerase activity and long telomeres (19-50 kb) in the long-term cultures, whereas human embryo cells rapidly deplete telomerase activity associated with significant shortening of telomeres, and then they senesce. In contrast to the results from 10-day cultures, we found that some mouse cell cultures and most Syrian hamster cell cultures arrest cell growth after 13 and 29 population doublings, respectively, while retaining substantial levels of telomerase activity and experiencing no significant loss of telomeres when the cells were subcultured using the 3-day culture protocol. This growth arrest is phenotypically indistinguishable from cellular senescence. The present results suggest that in rodent cells the onset of senescence-like arrest can be activated without repression of telomerase, and that this activation pathway can be bypassed easily under certain culture conditions, such as the 10-day culture protocol. PMID- 11121244 TI - Gene amplification and microsatellite instability induced in tumorigenic human bronchial epithelial cells by alpha particles and heavy ions. AB - Gene amplification and microsatellite alteration are useful markers of genomic instability in tumor and transformed cell lines. It has been suggested that genomic instability contributes to the progression of tumorigenesis by accumulating genetic changes. In this study, amplification of the carbamyl-P synthetase, aspartate transcarbamylase, dihydro-orotase (CAD) gene in transformed and tumorigenic human bronchial epithelial (BEP2D) cells induced by either alpha particles or (56)Fe ions was assessed by measuring resistance to N (phosphonacetyl)-l-aspartate (PALA). In addition, alterations of microsatellite loci located on chromosomes 3p and 18q were analyzed in a series of primary and secondary tumor cell lines generated in nude mice. The frequency of PALA resistant colonies was 1-3 x 10(-3) in tumor cell lines, 5-8 x 10(-5) in transformed cells prior to inoculation into nude mice, and less than 10(-7) in control BEP2D cells. Microsatellite alterations were detected in all 11 tumor cell lines examined at the following loci: D18S34, D18S363, D18S877, D3S1038 and D3S1607. No significant difference in either PALA resistance or microsatellite instability was found in tumor cell lines that were induced by alpha particles compared to those induced by (56)Fe ions. PMID- 11121246 TI - The role of nutrition in prostate cancer prevention. PMID- 11121245 TI - Identification and characterisation of the murine homologue of the gene responsible for cystinosis, Ctns. AB - BACKGROUND: Cystinosis is an autosomal recessive disorder characterised by an intralysosomal accumulation of cystine, and affected individuals progress to end stage renal failure before the age of ten. The causative gene, CTNS, was cloned in 1998 and the encoded protein, cystinosin, was predicted to be a lysosomal membrane protein. RESULTS: We have cloned the murine homologue of CTNS, Ctns, and the encoded amino acid sequence is 92.6% similar to cystinosin. We localised Ctns to mouse chromosome 11 in a region syntenic to human chromosome 17 containing CTNS. Ctns is widely expressed in all tissues tested with the exception of skeletal muscle, in contrast to CTNS. CONCLUSIONS: We have isolated, characterised and localised Ctns, the murine homologue of CTNS underlying cystinosis. Furthermore, our work has brought to light the existence of a differential pattern of expression between the human and murine homologues, providing critical information for the generation of a mouse model for cystinosis. PMID- 11121249 TI - Neoadjuvant hormone therapy and external beam radiation for localized prostate cancer: Vancouver Island Cancer Centre experience. AB - OBJECTIVES: To evaluate the effects of neo-adjuvant hormone therapy (NAHT) given prior to radiation in patients with clinically localized adenocarcinoma of the prostate. METHODS: Six hundred nine patients were treated between 1992 and 1997 with NAHT prior to radiation therapy. Clinical stage, presenting PSA and Gleason score were examined for influence on outcome. Time to post radiotherapy failure was defined from the first assessed PSA value over 4 microg/L at follow-up. Radiation therapy was confined to the prostate and seminal vesicles. Median follow-up was 2.6 years. RESULTS: PSA disease free survival (PDFS) decreased with increasing cancer risk factors (p <.0.0001). The overall duration of NAHT was significant with patients receiving >8 months having a lower failure rate than those on therapy for <3 months (p <0.0001). The PSA prior to starting radiation correlated with outcome, a PSA <=0.1 microg?L having a better PDFS than those with a PSA >=4 microg?L (P <0.0001). NAHT for >8 months gave improved PDFS in intermediate grade Gleason score 5-7, (n = 256, p <0.0001), high grade Gleason score 8-10 (n = 80, p =0.005), but not in low grade, Gleason score <=4. CONCLUSION: Neo-adjuvant hormone therapy for >8 months offers prolonged PSA disease free survival in patients with less well differentiated tumors, Gleason score >4. Clinical trials are required to confirm this. PMID- 11121247 TI - Nutritional aspects of prostate cancer: a review. AB - OBJECTIVES: The primary prevention of prostate cancer through nutritional modification is becoming a focus of attention as important relationships between diet and cancer are becoming evident. Relevant research is reviewed, along with recent data implicating various vitamin supplements and food products in the prevention and treatment of prostate cancer. METHODS: The epidemiology of prostate cancer, and current knowledge of prevention, screening, and progression of neoplasia is discussed. The current understanding of diet and its importance in primary and secondary prevention is explored. Literature searches were performed on MedLine using relevant keywords to find studies relating to prevention and treatment of prostate cancer using dietary methods. Of these, 104 published manuscripts were used. The search was limited from the year 1975 to the present. RESULTS: Incidence rates for prostate cancer vary according to diet and lifestyle. Several double-blind placebo-controlled clinical trials have shown that supplementation with selenium reduces cancer incidence. Inhibitory effects on the growth of in vitro prostate cancer cell lines have been observed with the administration of soy isoflavones, lycopenes from tomatoes, and vitamin D. Other compounds, such as calcium and fatty acids, have been linked to higher incidences of prostate cancer. CONCLUSIONS: Evidence exists that diet may play an important role in the primary prevention of prostate cancer. Further research is necessary to define the role that nutrition plays in the prevention or promotion of prostate cancer. PMID- 11121250 TI - Long term follow-up of intravesical Bacillus Calmette-Guerin for the treatment of bladder transitional cell carcinoma. AB - OBJECTIVE: To review the long-term follow-up, in terms of recurrence and progression, of transitional cell carcinoma of the bladder treated with intravesical BCG with the following indications: CIS, Ta and T1. MATERIALS AND METHODS: Ninety-two patients who had received complete course of BCG between 1987 and 1993 were included in the study and followed for an average of 59 months (range 12 to 102). RESULTS: The recurrence and progression were looked at. Patients treated with BCG for Carcinoma in situ, 11 of 19 (53%) remained tumor free after 1 or 2 courses of BCG for the duration of the follow-up (mean 4.9 years, range 1.5 to 8.5 years). For patients treated for recurring tumors, 17 of 50 (34%) had no recurrences after 1 or 2 courses of BCG with the same follow-up. When facing multiple tumors, 10 of 23 (43%) patients did not experience recurrences. Therefore, in the 92 patients treated, 38 presented no recurrences after 1 or 2 courses of BCG, for a success rate of 41%. In terms of progression, of the 19 patients treated with BCG for CIS, 4 (21%) went on to develop muscle invasive disease. Of the 50 patients treated for recurrent tumors, 2 (4%) eventually developed lamina propria invasion (initial lesion was a Ta tumor), 4 (8%) carcinoma in situ and 7 (14%) muscle invasive disease, for an overall progression rate of 26% in this group. Of the 25 patients treated for multiple tumors, 1 (4%) developed CIS and 3 (12%) presented with muscle invasive disease, for an overall progression rate of 16% for the duration of the follow-up. Therefore, 21 of 92 (23%) patients had progression of their disease following BCG therapy. No prognostic factors for recurrence or progression could be identified in these tumors. CONCLUSION: When indications warrant its use, BCG is effective in reducing recurrences and limiting progression in TCC of the bladder. Recurrence within 2 years of treatment is, however, a sign of poor prognosis and other therapeutic options should be sought. PMID- 11121251 TI - Primary extranodal lymphoma of the scrotum. AB - Primary extranodal lymphomatous involvement of the skin or genitourinary tract is rare. We report a case of primary scrotal diffuse large cell non-Hodgkin's lymphoma in a 78 year-old male. PMID- 11121255 TI - Re: Editorial. Laurence H. Klotz, Editor-In-Chief, Can J Urol. 1999;6(6):891. PMID- 11121252 TI - Flexible ureteroscopy and lithotripsy with the Holmium: YAG laser. AB - Extracorporeal shock wave lithotripsy (ESWL), Dornier Medical Systems, remains the main form of therapy for most urinary stones. The minimally invasive nature of ESWL makes it an attractive form of therapy to both patients and physicians. Many patients however may be most effectively managed by an endoscopic procedure either percutaneously or by transurethral ureteroscopy. Recent years have seen considerable advancement in endoscope technology such that flexible, actively deflectable ureteroscopes of 7.5 F or smaller in diameter have become available for performing upper tract endoscopy. When combined with new intracorporeal lithotripsy devices such as the Holmium:YAG laser, urologists have an effective alternative to ESWL for many stone problems. In this article we review our technique of flexible ureteroscopy combined with Holmium:YAG laser lithotripsy. PMID- 11121256 TI - Re: Editorial. Laurence H. Klotz, Editor-In-Chief, Can J Urol. 1999;6(6):891. PMID- 11121259 TI - German inflection: single route or dual route? AB - The German plural system has recently become a focal point for conflicting theories of language, both linguistic and cognitive. Marcus et al. (1995) highlight the German plural as support for the dual-route account of inflectional morphology first proposed by Pinker and colleagues (Pinker & Prince, 1988). On the dual-route account, inflectional morphology is universally subserved by a symbolic rule route which deals with regular inflection and an associative memory component which deals with irregular inflection. This contrasts with single-route connectionist systems. We seek to counter supposed evidence for the dual-route account through large-scale simulations as well as through experimental data. We argue that, in its current form, the dual-route account is incapable of generating experimental data provided by Marcus et al. (1995) as support. Finally, we provide direct quantitative comparisons between single-route and dual route models of German plural inflection and find single-route performance superior on these tests. PMID- 11121261 TI - EDITORIAL. PMID- 11121260 TI - Causal status as a determinant of feature centrality. AB - One of the major problems in categorization research is the lack of systematic ways of constraining feature weights. We propose one method of operationalizing feature centrality, a causal status hypothesis which states that a cause feature is judged to be more central than its effect feature in categorization. In Experiment 1, participants learned a novel category with three characteristic features that were causally related into a single causal chain and judged the likelihood that new objects belong to the category. Likelihood ratings for items missing the most fundamental cause were lower than those for items missing the intermediate cause, which in turn were lower than those for items missing the terminal effect. The causal status effect was also obtained in goodness-of exemplar judgments (Experiment 2) and in free-sorting tasks (Experiment 3), but it was weaker in similarity judgments than in categorization judgments (Experiment 4). Experiment 5 shows that the size of the causal status effect is moderated by plausibility of causal relations, and Experiment 6 shows that effect features can be useful in retrieving information about unknown causes. We discuss the scope of the causal status effect and its implications for categorization research. PMID- 11121262 TI - Ionic Strength Effects of Electrolytes on Solubilized States of Water in AOT Reversed Micelles. AB - The limiting amounts of solubilized aqueous NaCl, NaNO(3), MgCl(2), and AlCl(3) in sodium 1,2-bis-(2-ethylhexyloxycarbonyl)-1-ethanesulfonate (aerosol OT)/isooctane solutions have been measured as a function of the ionic strength of the electrolytes. In any system, the limiting amounts increased up to the optimal ionic strength (Io(opt)), and afterward, as ionic strength increased, they decreased and were followed by a constant. These increased and decreased curves of water solubilization below and above Io(opt), called, respectively, salting-in and salting-out curves, could be interpreted, respectively, from the counteracting effects of attractive intermicellar interaction and interfacial bending stress. The effects of electrolytes on the solubilized states of water by micelles were examined using NMR and near-infrared spectroscopic techniques. Consequently, two types of waters, i.e., water bound directly to the ionic head groups of the surfactant in reversed micelles and water interacting with the hydrated head groups in swollen micelles through hydrogen bonds, were almost unaffected by any electrolytes, whereas much greater effects were observed for bulklike water in W/O microemulsions. The order of the degree of lowering of salting-out curves in W/O microemulsion regions in each system was found to agree with the Hofmeister series. Copyright 2001 Academic Press. PMID- 11121263 TI - Intersalation Reactions of Trisdiimine Metal Complexes with Montmorillonite Clay: A New Approach. AB - Intersalation reactions of the complexes [Ni(L-L)(3)]SO(4) and [Ni(L-L)(3)]Cl(2) (L-L=1,10-phenanthroline (phen), 2,2'-bipyridyl (bpy)), with Na-montmorillonite carried out under different dynamic conditions such as ultrasonic irradiation, refluxing, autoclaving, and vigorous stirring showed that under ultrasonic irradiation, maximum adsorption of the metal complexes occurred within a period of 30 min. Metal complexes containing the phen ligand showed higher adsorption than those containing bpy. Maximum adsorption up to about three times the cation exchange capacity (CEC) of the clay was observed. Aggregated (basal spacing d(001) 12.5 A) Na-montmorillonite, in treatment with a higher amount (>CEC) of metal complexes, formed monolayered (basal spacing d(001) 17.6 A) species, while with predispersed clay, bilayered (basal spacing d(001)>28 A) or pseudo-trilayer (basal spacing d(001)>32 A) species were formed. The bilayered species were thermally stable up to 250 degrees C and above this temperature monolayered species were formed. The thermal stability of the intersalated species was higher than that of the intercalated ones. X-ray diffraction, UV-visible, IR spectroscopy, thermal analyses, and surface area measurements were used for characterization of the products. Copyright 2001 Academic Press. PMID- 11121264 TI - Adsorption and Bioactivity of Protein A on Silicon Surfaces Studied by AFM and XPS. AB - The adsorption of protein A on silicon surfaces was studied by atomic force microscopy (AFM) and X-ray photoelectron spectroscopy. The deposition was made statically from various concentrations of protein A in water solution. The biological activity was checked by the immobilization of rabbit immunoglobulin G. The protein adsorption occurs in least two different phases and leads to a multilayer film. The first monolayer of proteins is rapidly adsorbed on the surface. The adsorption of the second layer of proteins occurs much more slowly (a thousand times slower) and also involves the third monolayer. The protein A of the first monolayer is denaturated and biologically inactive. On the contrary, the proteins of the second monolayer keep their natural diameter and remain biologically active. AFM artifacts such as the convolution with small objects and the resulting estimation of the coverage ratio are discussed. Copyright 2001 Academic Press. PMID- 11121265 TI - Catalytic Activity of Hexokinase in Reversed Micelles. AB - Reversed micelles can control the size of water pools and the physical property of water by changing W(0)(=[water]/[surfactant]). Hexokinase (HK) activity seems to be easily affected by the microenvironment in the neighborhood of the enzyme because it is assumed that HK binds to the outer mitochondrial membrane by insertion of its hydrophobic NH(2) tail. The catalytic activity of HK was examined in reversed micelles in order to study the effect of the microenvironment in the neighborhood of HK on the activity. Sodium bis(2 ethylhexyl)sulfosuccinate (AOT), hexadecyltrimethyl ammonium chloride (HTAC), and octaoxyethylene dodecyl ether (C(12)E(8)) were used as anionic, cationic, and nonionic surfactants, respectively. HK activity was obtained by measuring ATP and ADP amounts with HPLC. The high electrostatic inner surfaces of AOT and HTAC reversed micelles were not favorable for HK to exhibit the catalytic activity, but the activity in HTAC reversed micelles was 2-3 times higher than that in AOT reversed micelles and the activities in both reversed micelles revealed an optimum at W(0)=10. The phenomenon was discussed in connection with the location of HK, nonuniform distribution of substrates, and the size and physical properties of the water pools. On the other hand, HK activity was much higher in C(12)E(8) reversed micelles than in AOT and HTAC reversed micelles and increased with the concentration of C(12)E(8). This suggests that HK activity is easily revealed in hydrated ethylene oxide chains. In conclusion, it was demonstrated that HK activity depends on the microenvironment such as the electrostatic field, the physical properties of water, and the hydrophobicity. Copyright 2001 Academic Press. PMID- 11121266 TI - Base-Catalyzed Oxidation of Carboxymethyl-cellulose Polymer by Permanganate. AB - The kinetics and mechanism of the disproportionation of MnO(4)(-) via [CMC MnVIO(4)(2-)] intermediate complex formation during the oxidation of carboxymethyl-cellulose (CMC) polysaccharide have been investigated spectrophotometrically at pH>/=12 and various temperatures (15-35 degrees C). The rate law was suggested as rate=k(obs)[MnO(4)(-)]. Kinetic and spectrophotometric measurements revealed the base-catalyzed formation of manganate(VI) transient species. A mechanism was postulated consistent with the experimental data. Copyright 2001 Academic Press. PMID- 11121267 TI - Location of Pinacyanol in Micellar Solutions of N-Alkyl Trimethylammonium Bromide Surfactants. AB - The interaction of pinacyanol (PIN), a cationic dye formed by monomer and dimer species, with three cationic surfactants (DTAB, TTAB, and HTAB) has been studied spectroscopically and by acid-base equilibrium in the micellar concentration range. In the presence of surfactants, the absorption maximum of the two main peaks undergoes bathochromic shifts. The spectral shifts suggest a hydrophobic environment of the chromophore. The presence of micelles favors the monomer species; i.e., it reduces the extent of dimerization. The pK(a) of PIN in micellar medium is similar to the value in pure water. When acid-base equilibrium was considered, the changes in the interfacial pK(a) allowed to us to determine the constant dielectric for the interfacial region (epsilon=69). This led to the conclusion that the dye must be solubilized between the solution and the hydrocarbon chain core, i.e., in the aqueous micellar interface. This location can be explained by a cation-pi interaction between the uncharged ring system of the dye and the cationic headgroups of the surfactants. Copyright 2001 Academic Press. PMID- 11121268 TI - Interfacial Activity of Trioctyloamine in Hydrocarbon/Water Systems with Nonorganic Electrolytes. AB - Interfacial tension and surface excess isotherms for trioctylamine (TOA) were determined and interpreted. Despite its high hydrophobicity, TOA adsorbs at the hydrocarbon/water interfaces and decreases effectively the interfacial tension, especially in systems containing acidic aqueous phase. Interfacial activity of TOA rises with an increase of the aqueous phase acidity. The effect of amine protonation is clearly observed. Interfacial tension isotherms obtained experimentally can be well matched with the Szyszkowski equation. The interfacial activity of TOA is affected by the type of the organic diluent and the composition of the aqueous phase, i.e., the kind and concentration of nonorganic electrolyte present in the system. Copyright 2001 Academic Press. PMID- 11121269 TI - Evaluation of Surface Enthalpy of Porous Aluminosilicates of the MCM-41 Type Using Immersional Calorimetry: Effect of the Pore Size and Framework Si:Al Ratio. AB - Surface properties of porous aluminosilicates of the MCM-41 type have been tested by immersional calorimetry. Two series of materials, referred to as SiAlxCn, where x is the Si : Al mole ratio and n the chain length of the surfactant template, having (1) x=32 and n=8, 12, 14, 16, 18 and (2) n=14 and x=8, 32, infinity, were used. The results of thermogravimetric analysis on these samples served to evaluate the surface density of hydroxyl groups. This parameter is rather sensitive to the pore size than to the aluminium content in the solid matrix. Based on the experimentally measured enthalpies of immersion in n heptane, water, and formamide per unit BET specific surface area, estimates could be made of the apolar, Lewis acid, and Lewis base contributions to the total surface enthalpy of MCM-41 materials. The samples studied have a predominant surface acidic character, which is markedly enhanced by incorporating aluminium into the silica matrix. Surface acidity is also modified by changes in the porous structure, although the trends are less noticeable here. Nevertheless, the total surface enthalpy of MCM-41 aluminosilicates appears to be small in comparison with typical inorganic oxides, such as silica or alumina. Copyright 2001 Academic Press. PMID- 11121270 TI - Effect of 2,2'-Bipyridine on the Adsorption of Zn(2+) Ions onto Silica Surface. AB - The influence of 2,2'-bipyridine (bipy) on adsorption of zink ions onto a highly dispersed silica surface has been studied. The enhanced adsorption of zink ions onto silica surface from the solution containing 2,2'-bipyridine is explained by ternary surface complex formation, { identical withSiOH(-m)Zn(bipy)(n)((2-m)+)}. The adsorbed Zn(2+) and bipy concentrations were measured at the adsorption from solutions with different ratios of Zn : bipy. The equilibrium reaction constants of binary and ternary complexes have been calculated using the constant capacitance model. The potentiometric titration data were used in order to determine the H(+)/Zn(2+) exchange stoichiometry. The composition of formed surface complexes has been confirmed with their UV absorption spectra. Copyright 2001 Academic Press. PMID- 11121271 TI - The Adsorption-Desorption Cycle. Reversibility of the BSA-Silica System. AB - The reversibility of the adsorption-desorption cycle was established by comparing the thermostability (determined by differential scanning calorimetry) and secondary structure (obtained by circular dichroism spectroscopy) of BSA before adsorption, adsorbed on, and exchanged from silica particles. Circular dichroism was also measured as a function of temperature at a given wavelength. Adsorbed BSA presents a higher thermostability and a lower alpha-helix content than the native protein while it regains its conformation when released from the surface back into the solution; the homomolecular exchange is reversible.The changes in ellipticity (at a given wavelength) as a function of the temperature show that the thermal denaturation of native, adsorbed, and exchanged BSA proceeds in two steps. For the dissolved protein, the first step up to 50 degrees C involves a slight change in the structure while in the 50-90 degrees C temperature range the actual unfolding takes place. For the adsorbed BSA, the first step proceeds up to 60 degrees C and includes some intermolecular association between the adsorbed protein molecules, which may be responsible for the increased thermostability. The unfolding occurs in the 60-90 degrees C range; it is less cooperative and involves a lower enthalpy change than the native protein. Adsorbed BSA presents the same secondary structure as that observed for dissolved BSA that has passed a heating-cooling cycle. Copyright 2001 Academic Press. PMID- 11121272 TI - The Composition of the Milk Fat Globule Surface Alters the Structural Characteristics of the Coagulum. AB - The effects of the composition of a fat globule surface in reconstituted milks on the properties of rennet-induced coagulums were studied by rheological measurements and by front-face fluorescence spectroscopy in combination with a multivariate statistical method to investigate, at a molecular level, the evolution of the structure during the milk coagulation process. Reconstituted milks used in this study were prepared from different fat-in-water emulsions stabilized by whole skim-milk proteins, beta-casein, or beta-lactoglobulin. Coagulation of milk reconstituted with natural fat globules was also investigated. The study showed that the fat droplet/water interface influences the rheological properties (G' modulus) of the reconstituted milks during the coagulation process. The tryptophan fluorescence emission spectra of proteins were recorded during the kinetics of coagulation. The results of the principal component analysis performed on the spectral data showed a discrimination in the different systems investigated. It was shown that the fluorescence properties of protein tryptophans and, consequently, the structures of the protein networks were different for the investigated systems. The development of fluorescence transfer between protein tryptophans and fat-globule vitamin A during the coagulation kinetics agreed with the interactions between the protein network and fat globules. Copyright 2001 Academic Press. PMID- 11121273 TI - Assessment of the Surface Heterogeneity of Talc Materials. AB - The hydrophobic and hydrophilic components of the surface of talc materials in aqueous solution were determined using ionic surfactants and their polar headgroup adsorption isotherms. The hydrophilic and hydrophobic surface areas are inferred from the amount of probe molecule adsorbed and the structure of the adsorbed layer. Natural dispersion of talc shows at 298 K a pH of 9.4 and the electrophoretic measurements indicate that the particles are negatively charged. The hydrophilic surface area is estimated from the adsorption of benzyltrimethylammonium ions (BTMA(+)) through electrostatic interactions as supported by the increase of divalent ions in the bulk phase and the decrease in the exothermic displacement enthalpy. It was also observed from the adsorption isotherm of benzene sulfonate anions that the density of positive surface sites is very low and is thus neglected. The adsorption of an anionic surfactant essentially occurs through dispersive interactions between the nonpolar organic tail of the molecule and the hydrophobic surface. Furthermore, some assumptions on the structure of dodecyl sulfate surfactant aggregates at the interface allow the hydrophobic part of the talc particles to be estimated. The cationic surfactant adsorption has been investigated and found to corroborate the hydrophilic and hydrophobic area values first obtained. Copyright 2001 Academic Press. PMID- 11121274 TI - Modeling Competitive Adsorption of Molybdate, Sulfate, and Selenate on gamma Al(2)O(3) by the Triple-Layer Model. AB - Competitive adsorption of molybdate, sulfate, and selenate onto gamma-Al(2)O(3) was investigated in the present study. Binary solute systems of MoO(2-)(4)+SO(2 )(4), MoO(2-)(4)+SeO(2-)(4), and SO(2-)(4)+SeO(2-)(4) and a ternary solute system of MoO(2-)(4)+SO(2-)(4)+SeO(2-)(4) were evaluated to determine their relative effects on competitive adsorption on the gamma-Al(2)O(3) surface. Anionic competitive adsorption efficiency was pH dependent. The higher the pH, the lower the efficiency of MoO(2-)(4) in preventing SO(2-)(4) and SeO(2-)(4) adsorption; similar results were found in SeO(2-)(4) depressing SO(2-)(4) adsorption. This research found that more sites are occupied in mixed anionic adsorbate systems than when either ion is present alone. The results suggest that the gamma Al(2)O(3) surface is composed of many groups of binding sites. Because of the heterogeneity of adsorption sites, the triple-layer model (TLM) predicted the competitive effects qualitatively but not quantitatively. TLM gave reasonable descriptions of molybdate adsorption in the presence of sulfate and selenate, indicating that the model may be useful in predicting molybdate adsorption on gamma-Al(2)O(3). Copyright 2001 Academic Press. PMID- 11121275 TI - Effect of Temperature on Interfacial Adsorption of Cr(VI) on Wollastonite. AB - An extensive study on the effect of temperature on interfacial adsorption of Cr(VI) on wollastonite has been carried out. Adsorption on the wollastonite surface increased from 69.5 to 91.7% by increasing the temperature from 30 to 50 degrees C under optimum conditions. Kinetic modeling of the process of adsorption of Cr(VI) was done and various parameters were determined. The process follows a first-order kinetic equation and the rate of uptake was found to be 2.40x10(-2) min(-1) at 30 degrees C, 2.5 pH, 0.5x10(-4) M Cr(VI) concentration, and 0.01 M NaClO(4) ionic strength. Kinetic and equilibrium modeling of the process of adsorption was undertaken and the equilibrium parameters were determined. The process of adsorption follows pore diffusion and the value of the rate constant of pore diffusion was found to be 5.00x10(-3) mg g(-1) min(-1/2) at 30 degrees C and optimum conditions. The values of the coefficient of mass transfer, beta(L), were determined at different temperatures. Thermodynamic studies of the removal process were performed. The study suggests that the process is a typical example of endothermic adsorption. Copyright 2001 Academic Press. PMID- 11121276 TI - Film Formation from Nanosized Copolymeric Latex Particles: A Photon Transmission Study. AB - The photon transmission technique was used to monitor the evolution of transparency during film formation from nanosized copolymeric latex particles. The latex films were prepared from poly(methyl methacrylate-co-butyl methacrylate) (P(MMA-co-BMA)) particles which were produced by microemulsion polymerization. These films were annealed at elevated temperatures in various time intervals above the glass transition temperature (T(g)) of P(MMA-co-BMA). It is observed that the transmitted photon intensity (I(tr)) from these films increased as the annealing temperature increased. There are three different film formation stages. These stages are explained by the void closure, healing, and interdiffusion processes, respectively. The activation energies for viscous flow (DeltaH approximately 16 kcal/mol), minor chains (DeltaE(H) approximately 27 kcal/mol), and backbone motion (Delta E(b) approximately 132 kcal/mol) were obtained using various models. Void closure (tau(v), T(v)) and healing points (tau(H), T(H)) were determined. Using the time-temperature pairs, void closure and healing activation energies were measured and found to be 21 and 30 kcal/mol, respectively. Copyright 2001 Academic Press. PMID- 11121277 TI - An Experimental Test of the Ion Condensation Theory for Spherical Colloidal Particles. AB - This paper deals with the notion of ion condensation for spherical colloids and, more specifically, with a recent model developed to predict effective charges (Y. Levin, M. C. Barbosa, and M. N. Tamashiro, Europhys. Lett. 41, 123, 1998). Electrophoretic mobility measurements (carried out for a set of well characterized latexes) were used to find out to what extent this theory is able to accounts for: (i) the insensitivity of mobility to surface charge, and (ii) the small values of electrokinetic charge found at low ionic strength. As the Levin theory was developed assuming no added salt, a previous discussion about the effect of additional electrolyte was needed. Contrary to what other authors have reported, our results do not support the above-mentioned theory. Copyright 2001 Academic Press. PMID- 11121278 TI - Water Solubilization in Nonionic Microemulsions Stabilized by Grafted Siliconic Emulsifiers. AB - Microemulsions containing octanol, decanol, or dodecanol as the oil phase and oligomeric, grafted nonionic amphiphiles based on ethoxylated polymethylsiloxanes (Silwets) have been studied. It was demonstrated that significant amounts of water can be solubilized only when the hydrophobic siliconic backbone is very short (trimers). The water solubilization was evaluated using SAXS, DSC, and conductivity measurements. It was found that up to 40 wt% of water can be solubilized in dodecanol and Silwet L-7607 (MW 1000 and 75 wt% ethylene oxide (EO)). Surprisingly, no free water was detected in the aggregate core. All the solubilized water was confined in the vicinity of the interphasal region and froze at -10 degrees C and below. Up to three molecules of water can be associated with each EO headgroup. Based on SAXS measurements, the structural units of the microemulsions were interpreted to be lamellar-like, a form previously found for the related monomeric microemulsions. Copyright 2001 Academic Press. PMID- 11121279 TI - Ellipsometry and Infrared Reflection Absorption Spectroscopy of Adsorbed Layers of Soluble Surfactants at the Air-Water Interface. AB - Optical techniques play an increasingly important role in the characterization of microstructure and surface densities of thin films at various interfaces. In this study, ellipsometry and infrared reflection absorption spectroscopy (IRRAS) were used for determining the surface densities of adsorbed layers of cationic surfactants in situ at the air-water interface. The surfactants were N(alpha) lauroyl-arginine methyl ester (LAM) and N(alpha), N(omega)-bis(N(alpha)-lauroyl arginine)-alpha,omega-alkylidenediamide (C(6)(LA)(2)). In ellipsometry, the ellipsometric phase angle Delta was obtained at various surfactant concentrations and was referenced to that of the solvent. Three algorithms were used for analyzing the data. The surface densities are 3.3+/-0.3x10(-6) mol/m(2) at 1 mM for LAM and 1.5+/-0.3x10(-6) mol/m(2) at 0.1 mM for C(6)(LA)(2) by using an algorithm for which the monolayer thickness was estimated from molecular modeling. The corresponding surface densities from literature surface tension data and the Gibbs adsorption isotherm procedure are 2.2+/-0.4x10(-6) mol/m(2) and 1.2+/-0.2x10(-6) mol/m(2), respectively. In addition, IRRAS spectra were obtained from monolayers of LAM and C(6)(LA)(2) at the air-water interface. The frequencies of the methylene stretching vibration bands indicate that the monolayers are liquid-like. The surface densities were determined from the reflectance-absorbance data by using the model of either an isotropic film or an anisotropic film on the aqueous subphase. The IRRAS-based surface densities from either model, by using DPPC monolayers for calibration, are 2.4+/-0.7x10(-6) mol/m(2) at 1 mM for LAM and 1.5+/-0.6x10(-6) mol/m(2) at 0.1 mM for C(6)(LA)(2), which are in fair agreement with the ellipsometry- and the surface-tension-based surface densities. Copyright 2001 Academic Press. PMID- 11121280 TI - Monolayer Characteristics of Bacteriocin AS-48, pH Effect and Interactions with Dipalmitoyl Phosphatidic Acid at the Air-Water Interface. AB - Bacteriocin AS-48 produced by Enterococcus faecalis S-48 is a ribosomally synthesized cyclic peptide (7.4 kDa) of broad inhibitory spectrum against Gram positive and Gram-negative bacteria. Simple monolayers of AS-48 and of dipalmitoyl phosphatidic acid (DPPA) at the air-water interface are studied. The AS-48 interfacial behavior in the function of pH explains the biological activity of the peptide. The lipid monolayers show the characteristic behavior of phosphatidic acid at the mentioned interface. The interactions between AS-48 and DPPA, a majority lipid of the bacterial cell membrane, are quantitatively investigated. The results indicate that only when the lipid molecules are charged enough (pH 10.5) is an attractive interaction between AS-48 and DPPA observed, although under these experimental conditions the results seem to indicate that a deformation of the peptide helical structure could take place. Copyright 2001 Academic Press. PMID- 11121281 TI - A Revised Stern Equation for Ion Exchange at Charged Membranes. AB - We revised the classical Stern equation for ion exchange at charged membranes using the statistical mechanics concept of chemical potential and ion activity in nonhomogeneous systems. Our proposed equation does not require an iterative procedure to obtain the self-regulated surface charge at the membranes. We predict significant differences with the classical results. When compared with experimental results, our approach not only gives a better fit but also predicts dissociation constants that are physically more reasonable than those from the classical approach. Copyright 2001 Academic Press. PMID- 11121282 TI - Interactions between Local Anaesthetic Agents and Poly(N-isopropyl acrylamide) through Phase Behavior, Surface Tension, and Adsorption Measurement. AB - The interaction between the local anaesthetic agents prilocaine and lidocaine, on one hand, and poly(N-isopropyl acrylamide) (pNIPAM), on the other, is investigated through studies of the polymer phase behavior and through surface tension and adsorption measurements. In particular, the cloud points (CP) for pNIPAM in the presence of lidocaine and prilocaine under different conditions were compared to the effects of electrolytes and alcohols. It was found that the electrolytes affect the CP of pNIPAM in a lyotropic manner, whereas alcohols depress the CP of pNIPAM in an alkyl chain length dependent way; i.e., the longer the chain, the larger the decrease in CP. Lidocaine and prilocaine affect the CP of pNIPAM in a pH-dependent manner. Below the pK(a) of lidocaine and prilocaine, these cosolutes do not substantially affect the CP in the concentration range investigated, but rather behave analogous to simpler electrolytes. Above the pK(a), on the other hand, they strongly depress the CP already at low concentrations. In parallel, at low pH, the surface tension reduction due to lidocaine or prilocaine is marginal, whereas at high pH the surface tension is reduced considerably. Thus, the poor solubility of prilocaine and lidocaine at high pH causes these to become more surface active and simultaneously interact in a more pronounced way with pNIPAM. Furthermore, it was found from ellipsometry that an adsorbed pNIPAM layer contracts when lidocaine is added, presumably due to a lidocaine-pNIPAM interaction similar to that causing pNIPAM to phase separate. Analogous to this, it was demonstrated that an adsorbed pNIPAM layer shrinks and swells reversibly when the temperature is cycled above and beneath the CP. Copyright 2001 Academic Press. PMID- 11121283 TI - Circular Hydraulic Jumps Triggered by Boundary Layer Separation. AB - When a high-flow-rate circular jet impinges vertically on a horizontal plane, it flows out radially and then undergoes a distinctive hydraulic jump on the plane because of boundary layer separation induced by hydrostatic back pressure. The jump radius is shown to be 0.37 a Re(1/3) Lambda(-1/8), where Lambda=(ga(3)/nu(2)) Re(-7/3) is a modified Froude number, Re=(Q/anu) is the jet Reynolds number, a is the jet radius, and Q the liquid flow rate, which is favorably compared to experimental data in the limit of small Lambda. When Lambda exceeds 3.0x10(-4) at low flow rates, the jump radius decreases below a minimum in the film depth and our experiments detect a different jump mechanism that may be triggered by capillary pressure rather than hydrostatic pressure. Copyright 2001 Academic Press. PMID- 11121284 TI - Study of the Second Order Transitions and Acid-Base Properties of Polymers Adsorbed on Oxides by Using Inverse Gas Chromatography at Infinite Dilution. AB - The study of phenomena transitions in polymers is of vital importance in material sciences and more particularly when polymers are adsorbed on oxides. Inverse gas chromatography (IGC) at infinite dilution proved to be an excellent technique to determine not only the glass transitions, but also beta-transition and liquid liquid transitions of polymers adsorbed on solid substrates. In this paper, we used the IGC technique to determine the second order transitions of the systems PMMA/SiO(2) and PMMA/Al(2)O(3), at various covered surface fractions and for various tacticities of the polymer (atactic, isotactic, and syndiotactic). In Part I, we developed the various theories, methods, and models used in the IGC technique in order to obtain physico-chemical properties when polymers are adsorbed on oxides. Copyright 2001 Academic Press. PMID- 11121285 TI - Study of the Second Order Transitions and Acid-Base Properties of Polymers Adsorbed on Oxides by Using Inverse Gas Chromatography at Infinite Dilution. AB - In Part I, we gave the details concerning inverse gas chromatography (IGC) at infinite dilution and the methods and models that will be used to characterize solid substrates. This technique proved to be an excellent technique to determine not only the glass transitions, but also beta-transition and liquid-liquid transitions of polymers adsorbed on solid substrates. In this second part, we used the IGC technique to determine the second order transitions of the systems' polymethyl methacrylate (PMMA)/SiO(2) and PMMA/Al(2)O(3), at various covered surface fractions and for various tacticities of the polymer. The maxima of the dispersive component of the surface energy gamma(s)(d) of our two systems, obtained by IGC at infinite dilution, indicated clearly the presence of transition temperatures (glass or local transitions). In general, we observed with PMMA three principal maxima that reflect the changes in motions leading to reorganization and rearrangement of the various groups or chain segments of the polymer. The change in the retention mechanism of the probes at the transition temperatures is attributed to an increased molecular mobility of the polymer segments, allowing for the penetration of the probes into the polymer layer. The study of the chemical physical properties of PMMA/SiO(2) and PMMA/Al(2)O(3) revealed an important difference in the acidic and basic behavior, in Lewis terms, of oxide covered by various concentrations of PMMA. This study also highlighted an important effect of the tacticity of the polymer on the acidic basic character of PMMA adsorbed on oxides. Copyright 2001 Academic Press. PMID- 11121286 TI - Streaming Potential Collection and Data Processing Techniques. AB - To date, no comprehensive comparison of streaming potential coupling coefficient collection or processing techniques has been made. Here, time-varying streaming potential and dc streaming potential data collection and processing techniques are presented and compared. The time-varying streaming potential data include sinusoidal and transient data. The collection techniques include acquiring dc streaming potentials at various pressures, acquiring time-varying streaming potentials at varying pressure, acquiring streaming potentials as a function of frequency, and collecting time-varying raw data. The processing techniques include dc filtering, rms processing, cross-correlation, spectral analysis, and plotting of raw time-varying streaming potential versus raw pressure data. The results show that all processing methods yield the same coupling coefficient within 3%. The analysis also shows that if there is a good signal-to-noise ratio, all processing methods perform satisfactorily. If the signal-to-noise ratio is poor, then the spectral analysis outperforms the other processing methods. The data collection methods are all adequate, but individual applications may make one method superior to another. Copyright 2001 Academic Press. PMID- 11121287 TI - Comparison of the Use of Washburn's Equation in the Distance-Time and Weight-Time Imbibition Techniques. AB - Two experimental methods are usually employed to study liquid penetration in porous media. One of them is based on the measure of the height of the advance liquid front vs time, and the other one is based on the measure of the weight gained by the porous system due to the liquid penetration vs time. Generally, the experimental data obtained from these techniques are analyzed through Washburn's equation. However, depending on which of them is selected, different conditions, coming from the experimental method, are needed to be taken into account in order to get the correct application of Washburn's equation to the experimental data. Although these conditions are different for each method, we prove in this paper that only if these conditions are considered both techniques are equivalent to analyze imbibition experiments using Washburn's equation. Copyright 2001 Academic Press. PMID- 11121288 TI - Characterization of the Chromophore Orientation of Rhodamine B Amphiphiles in Langmuir-Blodgett Monolayers. AB - SHG (second harmonic generation) interferometry and linear dichroism measurements were applied to the characterization of Langmuir-Blodgett monolayers composed of rhodamine B derivatives on hydrophilic fused silica substrates. When excited with a Nd-YAG laser, the contribution of dimeric species composed of rhodamine B chromophores to the nonlinear optical behavior was large, even though the relative concentration of the dimers was small. This implies the formation of noncentrosymmetrically oriented dimers with a relatively higher nonlinear molecular susceptibility, beta. Therefore, a unique phenomenon was observed; that is, the relative phase of the SH light shifted depending on the incident light polarization (p and s). In addition, it was found that the relatively standing up orientation was favored for the dimer, while a strongly tilted orientation was favored for the monomer. Copyright 2001 Academic Press. PMID- 11121289 TI - Formation of Monolayer Lipid Membranes in Water and Ethanol from Bolaamphiphiles. AB - Three single-chain bolaamphiphiles containing two salicylidene units per molecule as the rigid segment were synthesized. Their aggregation behavior in water and ethanol has been studied. They can form monolayer membranes not only in water solution but also in pure ethanol. The presence of a relatively short methylene chain as a spacer at the center of the molecule has a dramatic influence on the morphology of the aggregates formed. Copyright 2001 Academic Press. PMID- 11121290 TI - Differences in binding characteristics of sex steroid binding protein in reproductive and nonreproductive female rainbow trout (Oncorhynchus mykiss), black bream (Acanthopagrus butcheri), and greenback flounder (Rhombosolea tapirina). AB - The binding characteristics of sex steroid binding protein (SBP) were investigated in vitellogenic and nonreproductive female rainbow trout (Oncorhynchus mykiss), black bream (Acanthopagrus butcheri), and greenback flounder (Rhombosolea tapirina). The binding capacity of rainbow trout and black bream SBP was significantly greater in vitellogenic than in nonreproductive-stage fish. A decrease in binding affinity was observed in male trout injected with estradiol (E(2)) compared to control fish. This difference was not observed after partial purification of the SBP by gel filtration and may have resulted from competitive inhibition of E(2) binding by vitellogenin. No differences in flounder SBP binding characteristics were observed. PMID- 11121291 TI - A generalized fecal glucocorticoid assay for use in a diverse array of nondomestic mammalian and avian species. AB - Noninvasive fecal glucocorticoid analysis has tremendous potential as a means of assessing stress associated with environmental disturbance in wildlife. However, interspecific variation in excreted glucocorticoid metabolites requires careful selection of the antibody used in their quantification. We compared four antibodies for detecting the major fecal cortisol metabolites in yellow baboons following (3)H cortisol administration, ACTH challenge, and HPLC separation of fecal glucocorticoid metabolites. The most effective antibody (ICN corticosterone RIA; Cat. No. 07-120102) demonstrated relatively high cross-reactivities to the major cortisol metabolites present in feces during peak excretion, following both radiolabel infusion and ACTH challenge. This same antibody also detected increased fecal glucocorticoid metabolites after ACTH administration in the African elephant, black rhinoceros, Roosevelt elk, gerenuk, scimitar-horned oryx, Alaskan sea otter, Malayan sun bear, cheetah, clouded leopard, longtailed macaque, and northern spotted owl. Results suggest that (1) fecal glucocorticoid assays reliably detect endogenous changes in adrenal activity of a diverse array of species and (2) where comparisons were made, the ICN corticosterone antibody generally was superior to other antibodies for measuring glucocorticoid metabolites in feces. PMID- 11121292 TI - Localization of salmon cardiac peptide (sCP) in the heart of salmon (Salmo salar L.). AB - We have previously cloned and characterized a novel cardiac hormone from the salmon (Salmo salar) which has a uniquely heart-specific distribution and a low structural similarity with any other known natriuretic peptides. Specific antibodies were raised in goat against the salmon cardiac peptide. For localization and quantification, four different methods were applied: immunohistochemistry (avidin-biotin peroxidase), transmission electron microscopy, cryoimmunoelectron microscopy (protein A-gold), and a specific radioimmunoassay. Both atrial and ventricular myocytes stained immunohistochemically. The staining was similar in all myocytes and no specific myoendocrine cells were found. Within a single myocyte, both atrial and ventricular, the staining was stronger near the nucleus. Transmission electron microscopy revealed that both the atrium and the ventricle contained small sarcoplasmic granules of similar type with a diameter of 100 to 200 nm and an electron-dense core with a clear halo. The granules were typical vesicles which can be found in secretory cells utilizing the regulatory pathway. The highest number of granules was found near the nucleus, but granules were located also near the Golgi apparatus, between myofilament bundles, and in subsarcolemmal positions. Gold particles, conjugated to antibodies raised against the salmon cardiac peptide, were deposited on similar sarcoplasmic granules found in transmission electron microscopy. Among the sarcoplasmic granules with gold particles there were granules which did not show any cardiac peptide immunoreactivity. A significantly (Student's t test, P < 0.05) higher concentration of cardiac peptide was found in the heart atrium than in the ventricle, 16.2 +/- 3.5 pmol/mg tissue (n = 8) and 4.5 +/- 1.7 pmol/mg tissue (n = 8), respectively. The findings show that the salmon cardiac peptide is localized in secretory granules in both compartments of the heart. The morphology of the granules suggests that both the atrium and the ventricle utilize the regulatory pathway to release salmon cardiac peptide. PMID- 11121293 TI - Corticosterone inhibits normal and FSH-induced testicular recrudescence in the lizard, Mabuya carinata. AB - Administration (ip) of 1, 10, or 20 microg corticosterone (alternate days for 30 days) to adult male Mabuya carinata did not affect the seasonal recrudescence of spermatogenesis whereas administration of 40 microg corticosterone did result in inhibition of spermatogenesis. Further, administration of FSH (10 IU/lizard/alternate day for 30 days) during the quiescent phase of the testicular cycle stimulated spermatogenetic and steroidogenic activity of the testis as shown by significant increases in the mean number of spermatogonia, spermatocytes, and spermatids and serum levels of testosterone. In addition there were abundant spermatozoa in the lumen of the tubules in FSH-treated lizards. Administration of 10 IU FSH + 40 microg corticosterone (per lizard on alternate days for 30 days) increased the mean number of primary and secondary spermatocytes whereas the mean number of spermatids did not show significant variation compared with that of controls. Further, the mean numbers of spermatocytes and spermatids and serum levels of testosterone were significantly less when compared to those of FSH alone treated lizards. In addition, FSH induced development of epididymis was also inhibited by corticosterone treatment. The results indicate that corticosterone inhibits FSH-induced testicular recrudescence, possibly by suppressing testosterone secretion in M. carinata. PMID- 11121294 TI - Plasma steroid-binding globulin mediation of differences in stress reactivity in alternative male phenotypes in tree lizards, Urosaurus ornatus. AB - Plasma steroid-binding globulins, for example, corticosteroid-binding globulin and sex hormone-binding globulin (SHBG), have been identified in a number of vertebrates. One possible function of these proteins is to regulate the amount of steroid delivery to target tissues, as only free steroids are believed to diffuse from the circulation to target cells. Male tree lizards, Urosaurus ornatus, exhibit alternative male reproductive tactics correlated with dewlap (throat-fan) coloration. Males with orange-blue dewlaps are aggressive and territorial, whereas males with orange dewlaps are less aggressive and employ a satellite strategy. The two types of males have similar basal levels of total plasma corticosterone and testosterone. However, testosterone levels of nonterritorial males are more sensitive than those of territorial males to negative regulation by stress-induced increases in corticosterone. We tested the hypothesis that this difference in corticosterone feedback on testosterone could be mediated, in part, by differences in binding globulin levels between the two types of males. We have identified two steroid-binding globulins in male tree lizards. The first binds androgens and estradiol with high affinity (10(-9) M) and is similar to previously described sex hormone-binding globulins. The second binds both androgens and C(21) steroids, such as progesterone and corticosterone, with higher specificity than estradiol and is best described as an androgen glucocorticoid-binding globulin (AGBG). In both types of males, the capacity of AGBG is much higher than SHBG. In addition, AGBG capacity is significantly greater in territorial than nonterritorial males, whereas the capacity of SHBG does not differ between the two types of males. Calculations of free steroid levels based on the affinity and capacity measures suggest that although most testosterone circulates bound to binding globulins, binding capacity is high enough that binding globulins are also able to bind to other steroids such as corticosterone. Thus, differences in binding capacity between the two types of males could result in higher levels of free corticosterone in nonterritorial males than in territorial males, especially during stress-induced increases in corticosterone, and may explain why testosterone levels of nonterritorial males are more sensitive to negative feedback by corticosterone. PMID- 11121295 TI - Estrogen-induced vitellogenin mRNA and protein in sheepshead minnow (Cyprinodon variegatus). AB - Many environmentally persistent xenobiotic chemicals appear to disrupt normal endocrine function by acting as ligands for endogenous steroid receptors, including the estrogen receptor. Xenobiotics that bind to the estrogen receptor may elicit several effects, one of which is activating estrogen-responsive genes, such as vitellogenin (Vtg). Primers to vitellogenin mRNA have been used to amplify a portion of the coding sequence in sheepshead minnow (SHM) (Cyprinodon variegatus). Two Vtg cDNA fragments from SHM were isolated exhibiting 72% sequence homology and corresponding to the two Vtg genes identified in the mummichog, Fundulus heteroclitus. Using these Vtg cDNA fragments as sensitive genetic probes, we evaluated the initial estrogenic response of fish exposed to natural or anthropogenic chemicals. These probes were used to study in vivo gene induction in SHM exposed to 17beta-estradiol (E(2)) and ethinylestradiol (EE(2)) under controlled laboratory conditions. Hepatic Vtg mRNA was upregulated and plasma Vtg synthesis in estrogen-induced SHM was assessed. Two in vivo time course experiments were conducted; a single injection of E(2) followed over 72 h and a double E(2) injection examined for 12 days. These two protocols provided evidence for differential hepatic Vtg mRNA regulation resulting from a single or a double injection. In a separate experiment using an aqueous flowthrough system, constant exposures to low doses of E(2) (200 ng/L) and EE(2) (100 ng/L) induced hepatic Vtg mRNA and plasma Vtg to levels comparable with the E(2) injections. Larger aqueous exposure doses (2000 ng/L E(2) or 1000 ng/L EE(2)) in the flowthrough experiment resulted in greater responses of hepatic Vtg mRNA and plasma Vtg at 7 days. Constant aqueous exposure to E(2) (2000 ng/L) or EE(2) (1000 ng/L) may thus be more effective than a single large-dose injection (5 mg/kg) to stimulate Vtg gene activation and synthesis. PMID- 11121296 TI - Novel transcripts of the estrogen receptor alpha gene in channel catfish. AB - Complementary DNA libraries from liver and ovary of an immature female channel catfish were screened with a homologous ERalpha cDNA probe. The hepatic library yielded two new channel catfish ER cDNAs that encode N-terminal ERalpha variants of different sizes. Relative to the catfish ERalpha (medium size; 581 residues) previously reported, these new cDNAs encode Long-ERalpha (36 residues longer) and Short-ERalpha (389 residues shorter). The 5'-end of Long-ERalpha cDNA is identical to that of Medium-ERalpha but has an additional 503-bp segment with an upstream, in-frame translation-start codon. Recombinant Long-ERalpha binds estrogen with high affinity (K(d) = 3. 4 nM), similar to that previously reported for Medium-ERalpha but lower than reported for catfish ERbeta. Short-ERalpha cDNA encodes a protein that lacks most of the receptor protein and does not bind estrogen. Northern hybridization confirmed the existence of multiple hepatic ERalpha RNAs that include the size range of the ERalpha cDNAs obtained from the libraries as well as additional sizes. Using primers for RT-PCR that target locations internal to the protein-coding sequence, we also established the presence of several ERalpha cDNA variants with in-frame insertions in the ligand binding and DNA-binding domains and in-frame or out-of-frame deletions in the ligand-binding domain. These internal variants showed patterns of expression that differed between the ovary and liver. Further, the ovarian library yielded a full length, ERalpha antisense cDNA containing a poly(A) signal and tail. A limited survey of histological preparations from juvenile catfish by in situ hybridization using directionally synthesized cRNA probes also suggested the expression of ERalpha antisense RNA in a tissue-specific manner. In conclusion, channel catfish seemingly have three broad classes of ERalpha mRNA variants: those encoding N-terminal truncated variants, those encoding internal variants (including C-terminal truncated variants), and antisense mRNA. The sense variants may encode functional ERalpha or related proteins that modulate ERalpha or ERbeta activity. The existence of ER antisense mRNA is reported in this study for the first time. Its role may be to participate in the regulation of ER gene expression. PMID- 11121298 TI - Molecular cloning and sequence analysis of the cDNA for thyroid-stimulating hormone beta subunit of Muscovy duck. AB - The cDNA-encoding thyroid-stimulating hormone beta subunit (TSHbeta) of the Muscovy duck (Cairina moschata) was cloned and sequenced to investigate the phylogenic diversity and evolution of TSH molecules. Oligonucleotide primers were designed and used for reverse transcription and polymerase chain reaction amplification of a TSHbeta cDNA fragment from the total cellular RNA of pituitary glands. The remaining sequence was completed by rapid amplification of cDNA ends. The nucleotide sequence of Muscovy duck TSHbeta cDNA includes 66 bp of 5' untranslated region (UTR), 402 bp of coding region, and 82 bp of 3'-UTR, followed by an 18-bp poly(A)(+) tract. The total number of amino acids deduced from the cDNA of duck TSHbeta is 134, with a signal peptide of 19 amino acids and a mature protein of 115 amino acids. The homologies of the amino acid sequence of Muscovy duck TSHbeta compared with those of chicken, quail, mammals, amphibian (frog), and teleosts are 97, 98, 68-69, 56, and 42-47%, respectively. To test for tissue specificity of the duck TSHbeta cDNA, total cellular RNA samples from brain, liver, pituitary gland, testis, and thyroid gland were analyzed by Northern blotting. A band, approximately 700 bases, was found in the pituitary gland alone. The pituitary tissue fragments were treated for 24 h in serum-free medium containing thyrotropin-releasing hormone (TRH), triiodothyronine (T(3)), or thyroxine (T(4)). TRH increased and T(3) and T(4) decreased TSHbeta mRNA levels. This study demonstrated that the amino acid sequence of TSHbeta subunits is highly conserved among birds, exhibiting a high degree of inter-order homology, and that hypothalamic TRH upregulates the TSHbeta mRNA expression in duck. PMID- 11121297 TI - Molecular cloning and characterization of alternatively spliced transcripts of the turkey pituitary adenylate cyclase-activating polypeptide. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) increases the release of growth hormone (GH) and prolactin (PRL) in mammals. However, the evolutionary and functional relationships of PACAP, GH, and PRL are not clear. To understand how PACAP is regulated in the turkey, a turkey PACAP (tPACAP) cDNA has been cloned by the combination of reverse transcription-polymerase chain reaction and the rapid amplification of cDNA 5'- and 3'-ends. The deduced amino acid sequence of tPACAP-38 and turkey PACAP-related peptide (tPRP) displayed 87-97 and 52-63% similarity when compared to a variety of known PACAP-38 and PRP sequences, respectively. Two major transcripts (1.3 and 3.0 kb) of tPACAP were detected by Northern blot analysis. The highest levels of tPACAP mRNA were shown to be expressed in the hypothalamus, the cerebellum, and the cerebrum. In contrast, most of the other tissues tested expressed relatively low steady-state levels of tPACAP mRNA. Alternative splicing of tPACAP resulted in the expression of two different isoforms. The smaller form of tPACAP was expressed in the hypothalamus during early embryonic development and decreased significantly in later stages. PMID- 11121299 TI - Localization of immunoglobulin G gamma-chain mRNA-expressing cells in the oviduct of laying and diethylstilbestrol-treated immature hens. AB - The purpose of this study was to determine the synthetic sites of IgG in the chicken oviduct by localizing IgG gamma-chain mRNA (IgGgamma mRNA)-expressing cells and the effects of estrogen on their population. Paraffin sections of oviducal tissues from laying hens (approximately 57 weeks old) and immature hens (approximately 16 weeks old) with or without diethylstilbestrol (DES) treatment were hybridized by digoxigenin-labeled IgG riboprobes or immunostained for IgG gamma-chain (IgG). Immunoreactive IgG was present in some of the mucosal epithelial cells and the plasma cell-like cells in the stromal connective tissue in all segments of the oviduct. In contrast, IgGgamma mRNA expression was observed only in the plasma cell-like cells in the stromal connective tissues, but not in the cells of mucosal epithelium. In laying hens, the lower end of the oviduct, namely the vagina and uterovaginal junction, contained more IgGgamma mRNA-expressing cells than the other segments. Treatment of immature hens with DES for 3 or 6 days increased the population of both IgGgamma mRNA-expressing cells and IgG-containing cells in the oviducal stroma. These results indicate that IgG is locally produced by plasma cell-like cells in the stroma, but not by the cells of the mucosal epithelium, and estrogen may stimulate the infiltration of IgG-producing plasma cell-like cells into the oviducal stroma. PMID- 11121300 TI - Gastroenteropancreatic hormones (insulin, glucagon, somatostatin, and multiple forms of PYY) from the pallid sturgeon, Scaphirhynchus albus (Acipenseriformes). AB - Insulin, glucagon, somatostatin-14, and three structurally related molecular forms of peptide tyrosine-tyrosine (PYY) were isolated from an extract of the combined pancreas and gastrointestinal tract of the pallid sturgeon, Scaphirhynchus albus. Pallid sturgeon insulin was identical to insulin from the Russian sturgeon, Acipenser guldenstaedti, and to insulin-2 from the paddlefish, Polyodon spathula, and was approximately twofold less potent than human insulin in inhibiting the binding of [3-[(125)I] iodotyrosine-A14] human insulin to the soluble human insulin receptor. The sturgeon glucagon (HSQGMFTNDY(10) SKYLEEKLAQ(20) EFVEWLKNGK(30)S), like the two paddlefish glucagons, contains 31 rather than 29 amino acid residues, indicative of an anomalous pathway of posttranslational processing of proglucagon. Pallid sturgeon somatostatin, identical to human somatostatin-14, was also isolated in a second molecular form containing an oxidized tryptophan residue, but [Pro(2)]somatostatin-14, previously isolated from the pituitary of A. guldenstaedti, was not identified. Sturgeon PYY (FPPKPEHPGD(10)DAPAEDVAKY(20)YTALRHYINL(30) ITRQRY.HN(2)) was also isolated in variant forms containing the substitutions (Phe(1) --> Ala) and (Ala(18) --> Val), indicative of at least two gene duplications occurring within the Acipenseriformes lineage. The amino acid sequences of the pallidsturgeon PYY peptides are appreciably different from the proposed "ancestral" PYY sequence that has otherwise been very strongly conserved among the actinopterygian and elasmobranch fish. PMID- 11121301 TI - CUMULATIVE ANIMAL INDEX Volumes 117-120. PMID- 11121302 TI - Yeast nuclear pore complex assembly defects determined by nuclear envelope reconstruction. AB - Assembly of nuclear pore complexes (NPCs) is a critical yet poorly understood cellular function. One approach to studying NPC assembly is to identify yeast mutants defective in this process. This requires robust assays for NPC assembly that can be used for phenotypic analysis. We have previously reconstructed yeast nuclei from electron micrographs of serially sectioned cells to precisely determine the number of NPCs (Winey et al., 1997). Here we report the analysis of strains mutant in either of two nucleoporin-encoding genes, NIC96 (Zabel et al., 1996) and NUP192 (Kosova et al., 1999). Using conditional alleles of either gene, we have found that the NPC number falls significantly following shift to the restrictive temperature. We conclude that the drop in NPC number results from the failure to assemble new NPCs during cell divisions, leading to the dilution of NPCs that existed when the cells were shifted to the restrictive temperature. We are also able to document a subtle defect in NPC numbers in nup192-15 cells at their permissive temperature. The data presented here quantitatively demonstrate that NPC numbers fall in nic96-1 and nup192-15 strains upon shifting to the restrictive temperature, indicating that these gene products are required for NPC assembly. PMID- 11121303 TI - ATP-induced transconformation of myosin revealed by determining three-dimensional positions of fluorophores from fluorescence energy transfer measurements. AB - The method of fluorescence resonance energy transfer (FRET) is one of the most important techniques for measuring the distance between two fluorophores and for detecting the changes in protein structure under physiological conditions. The use of green fluorescent protein is also a powerful technology that has been used to elucidate dynamic molecular events. From these we have developed a novel method to determine the three-dimensional positions of fluorophores by combining the FRET data and other structural information available. Using this method, we could determine the ATP-induced changes of three-dimensional structure of truncated Dictyostelium myosin in solution. The myosin structure with ADP in solution was found to be similar to that of the crystal structure of MgADPBeFx bound truncated Dictyostelium myosin (type I structure), whereas myosin with ATP in solution was similar to the crystal structure of MgAdPVi-bound one (type II structure). However, the crystal structure of MgADP-bound scallop myosin (type III structure) could not be explained by any of our FRET data under various conditions. This indicates that the type III crystal structure might represent a transient intermediate conformation that could not be detected using fluorescence energy transfer. PMID- 11121304 TI - Immunohistochemical demonstration of hyaluronan and its possible involvement in axolotl neural crest cell migration. AB - Hyaluronan (HA), an extracellular matrix component, is involved mainly in the control of cell proliferation, neural crest and tumor cell migration, and wound repair. We investigated the effect of hyaluronan on neural crest (NC) cell migration and its ultrastructural localization in dark (wild-type) and white mutant embryos of the Mexican axolotl (Ambystoma mexicanum, Amphibia). The axolotl system is an accepted model for studying mechanisms of NC cell migration. Using a biotinylated hyaluronan binding protein (HABP), major extracellular matrix (ECM) spaces, including those of NC cell migration, reacted equally positive on cryosections through dark and white embryos. Since neural crest derived pigment cells migrate only in subepidermal spaces of dark embryos, HA does not seem to influence crest cell migration in vivo. However, when tested on different alternating substrates in vitro, migrating NC cells in dark and white embryos prefer HA to fibronectin. In vivo, such an HA migration stimulating effect might exist as well, but be counteracted to differing degrees in dark and white embryos. The ultrastructural localization of HA was studied by means of transmission electron microscopic immunohistochemistry using HABP and different protocols of standard chemical fixation, cryofixation, embedding, and immunolabeling. The binding reaction of HA to HABP was strong and showed an equal distribution throughout ECM spaces after both standard chemical fixation/freeze substitution and cryofixation. A preference for the somite or subepidermal side was not observed. Following standard fixation/freeze substitution HABP-labeled "honeycomb"-like networks reminiscent of fixation artifacts were more prominent than labeled fibrillar or irregular net-like structures. The latter predominated in adequately frozen specimens following high-pressure freezing/freeze substitution. For this reason fibrillar or irregular net-like structures very likely represent hyaluronan in the complex subepidermal matrix of the axolotl embryo in its native arrangement. PMID- 11121305 TI - Leginon: an automated system for acquisition of images from vitreous ice specimens. AB - We have developed a system to automatically acquire cryo-electron micrographs. The system is designed to emulate all of the decisions and actions of a highly trained microscopist in collecting data from a vitreous ice specimen. These include identifying suitable areas of vitreous ice at low magnification, determining the presence and location of specimen on the grid, automatically adjusting imaging parameters (focus, astigmatism) under low-dose conditions, and acquiring images at high magnification to either film or a digital camera. This system is responsible for every aspect of image acquisition and can run unattended, other than requiring periodic refilling of the cryogens, for over 24 h. The system has been tested out on a variety of specimens that represent typical challenges in the field of cryo-electron microscopy. The results show that the overall performance of the system is equivalent to that of an experienced microscopist. PMID- 11121306 TI - Multiple restraints to the unfolding of spermidine nucleoids from Escherichia coli. AB - Bacterial DNA is largely localized in compact bodies known as nucleoids. The structure of the bacterial nucleoid and the forces that maintain its DNA in a highly compact yet accessible form are largely unknown. In the present study, we used urea to cause controlled unfolding of spermidine nucleoids isolated from Escherichia coli to determine factors that are involved in nucleoid compaction. Isolated nucleoids unfolded at approximately 3.2 M urea. Addition of pancreatic RNase reduced the urea concentration for unfolding to approximately 1.8 M urea, indicating a role of RNA in nucleoid compaction. The transitions at approximately 3.2 and approximately 1.8 M urea reflected a RNase-sensitive and a RNase resistant restraint to unfolding, respectively. Removal of the RNase-sensitive restraint allowed us to test for roles of proteins and supercoiling in nucleoid compaction and structure. The remaining (RNase-resistant) restraints were removed by low NaCl concentrations as well as by urea. To determine if stability would be altered by treatments that caused morphological changes in the nucleoids, transitions were also measured on nucleoids from cells exposed to chloramphenicol; the RNase-sensitive restraint in such nucleoids was stabilized to much higher urea concentrations than that in nucleoids from untreated cells, whereas the RNase-resistant transition appeared unchanged. PMID- 11121307 TI - Surface beta-strands in proteins: identification using an hydropathy technique. AB - From a representative set of monomeric globular proteins with known three dimensional structures, beta-strands with lengths > or = 5 amino acids have been identified and catalogued. By ascertaining the accessible surface areas of the constituent residues in these strands, and by checking whether the exposed/buried pattern is 80% or more similar to that in an idealized surface strand, a subset of structures can be delineated in which the beta-strands are all sited on the surface of the protein. The corresponding sequence data show that about 50% of the residues are apolar (Val, Ile, Leu, Phe, Tyr, Ala) and that the common occurrence of valine (14.3%), isoleucine (9.6%), and threonine (8.1%) is a characteristic feature. The frequencies of occurrence of those amino acids in the strands that face the aqueous environment and the interior have also been determined separately and show that most surface strands have a substructure of the form (apolar-X)(n), where X is approximately equally divided between apolar, charged, and hydrophilic residues. Using the frequency data thus obtained, allied with an algorithm to delineate potential surface beta-strands from characteristic hydropathy profiles, it is now possible to search through the sequences of proteins with unknown tertiary structures and make realistic predictions of the presence of this element of structure on the protein surface. In addition, new data are presented on the distribution of the various types of residues on the surface of proteins and in their interior. Significant differences were observed, not all of which have been identified previously. Furthermore, the distribution of the types of residue in a surface beta-strand was compared to that corresponding to the surfaces of all of the proteins in our database. Again, very characteristic differences were observed. These are helpful in recognizing the presence of surface beta-strands. PMID- 11121308 TI - Structure and molecular mechanism of a functional form of pneumolysin: a cholesterol-dependent cytolysin from Streptococcus pneumoniae. AB - One of the key steps in understanding human disease arising from gram-positive bacteria lies in the mechanisms of the cholesterol-dependent cytolysins (CDCs). Pneumolysin (PLY), a CDC from Streptococcus pneumoniae, is of special importance due to the severe impacts of pneumococcal infections on mortality and morbidity worldwide. We have overexpressed, purified, and characterized PLY in its fully functional complex form with the enzyme bound to its receptor activator on target cells, cholesterol. The circular dichroism studies of PLY in solution with an excess of cholesterol show a change in the far UV spectrum consistent with a decrease in the beta-sheet and an increase in the random coil structures of the enzyme. Pore formation in membranes leading to cell lysis is the functional target for this cytolysin. The sedimentation velocity and equilibrium analyses of the cholesterol-bound enzyme show hydrodynamic properties different from those of the cholesterol-free form. The soluble form of the cholesterol-free enzyme exists in solution as a mixture of monomers and dimers, whereas the cholesterol-bound form exists only as a monomer. A mechanism of formation of PLY pores in the lipid bilayer of the target cells is discussed. PMID- 11121310 TI - Mimicking lip features in free-flap reconstruction of lip defects. AB - This paper introduces a technique of lip reconstruction using free flaps in which recognisable landmarks are mimicked by strategic placement of flap junctions. The technique was applied in 15 patients undergoing reconstruction of combined cheek and lip defects using single (n= 9) or double (n= 6) free flaps. Attention to flap design and strategic placement of flap junctions successfully created the vermilion-cutaneous junction, oral commissure and labiomental groove. The presence of these distinguishing features improved the appearance of the reconstructed lip. PMID- 11121309 TI - A review of therapeutic angiogenesis and consideration of its potential applications to plastic and reconstructive surgery. AB - The use of exogenous agents to stimulate the growth of new blood vessels into ischaemic tissue is a potentially revolutionary therapy in a wide variety of clinical specialties. Therapeutic angiogenesis research has been mostly confined to ischaemia of the heart and the lower limb. There has been relatively little research into the potential applications of the technique to plastic, reconstructive and burns surgery. In this paper, relevant published work is reviewed and potential applications of therapeutic angiogenesis to our specialty are considered. PMID- 11121311 TI - Retroauricular flap: its clinical application and safety. AB - We report the application and safety of the retroauricular flap in 38 cases. The flap was used on the anterior auricular surface in 21 cases, in the peri auricular region in five cases, in the region of the preauricular sideburn area in two cases, in the malar region in six cases, in the eyelid in three cases and in the intraorbital region in one case. When this flap was used in the auricular or periauricular region, the blood circulation was safe and the appearance was aesthetically good in flaps pedicled by the postauricular vessels or by the superficial temporal vessels. However, when the defect was more remote from the auricle, the blood circulation of the flaps pedicled by superficial temporal vessels, whether subcutaneous pedicle flaps or free flaps, was unstable. In some cases there was extensive or partial necrosis of the distal area of the flap. On the other hand, the free flaps pedicled by the postauricular vessels had satisfactory blood circulation, but the vessels were sometimes short, narrow and difficult to find, especially the veins. In these cases, we were obliged to use the superficial temporal vessels. A further problem is that some of the patients, especially younger women, were dissatisfied when the retroauricular flap was used in the malar region because of the reddish colour of the flap. PMID- 11121312 TI - A new operative method of correcting cryptotia using large Z-plasty. AB - A large Z-plasty on the postauricular surface of the ear has been used for successful reconstruction of cryptotia. This technique provides extensive skin coverage of the posterior auricle without skin grafting. The technique is easy and simple. The natural hairline is not disturbed and the temporoauricular sulcus is emphasised. Almost all of the scar is hidden behind the auricle. It has been performed on 17 patients with no recurrence of the cryptotia. One patient had partial congestion in the cranial-flap tip, which improved within 1 week. There were no other complications. PMID- 11121313 TI - Lipoma extraction via small remote incisions. AB - We describe a technique for extraction of lipomata using gynaecological polyp forceps, via incisions placed in aesthetically better sites than directly over the lesion. Although this can also be achieved by liposuction, we have found that this alternative technique is valuable in those cases where liposuction has failed due to the fibrous nature of the lesion, or where equipment is not available. PMID- 11121314 TI - The posterior interosseous flap: a review of 81 clinical cases and 100 anatomical dissections--assessment of its indications in reconstruction of hand defects. AB - Based on our experience of 100 cadaveric dissections and a series of 81 clinical cases, we have assessed the indications for the posterior interosseous flap in reconstruction of the hand. Large fasciocutaneous island flaps can be harvested, even when the radial or ulnar pedicles are damaged, sacrificing only vessels of secondary importance to the perfusion of the hand. Compound flaps can be dissected based on muscular, musculoperiosteal and fascioperiosteal branches. The primary indications for using this flap are dorsal hand defects up to the metacarpal joints, reconstruction of the first web space up to the interphalangeal joint of the thumb and extensive lesions on the ulnar border of the hand. PMID- 11121315 TI - Polydactyly of the feet in children: suggestions for surgical management. AB - Polydactyly is the commonest congenital deformity of the foot, presenting as a range of defects from minor soft tissue duplications to major bony abnormalities. There is a relative paucity of information on the management of this condition in the literature compared to that concerning polydactyly of the hand. We present a consecutive series of 34 cases of polydactyly of the foot in 25 patients treated surgically at our unit and these are classified according to the protocol described by Blauth and Olason. We emphasise the importance of preoperative classification using radiographs and an individualised surgical approach giving consideration to aesthetic and functional outcome. In the literature a number of authors have expressed the view that in polydactyly of the fifth ray of the foot the most lateral digit should always be excised irrespective of whether this is the more fully formed digit. We believe this should not always be the case and we describe two cases of polysyndactyly where the more medial element of a fifth-ray polydactyly was excised to allow for better maintenance of the contour of the foot. This involved more complex surgery than excision of the lateral element but gave a superior cosmetic and functional result. PMID- 11121316 TI - A prospective quantitative comparison of breast sensation after superior and inferior pedicle mammaplasty. AB - Reduction mammaplasty techniques using the inferior pedicle have been recommended to preserve the nipple and areolar sensation after surgery. The vertical scar mammaplasty with a superior pedicle has often been criticised because of the potential for damage to the sensory supply of the nipple-areola complex. The aim of this study was to assess the breast sensation in two prospective series of patients operated upon using superior pedicle and inferior pedicle mammaplasties. Between November 1996 and February 1997, 20 consecutive patients (39 breasts) underwent breast reduction using the inferior pedicle technique with inverted T scar (Robbin's technique). This series of patients was matched with another series of 18 patients (36 breasts) who had breast reduction using a vertical scar mammaplasty with superior pedicle (Lejour's technique) in another centre. Cutaneous pressure thresholds were recorded using Semmes-Weinstein monofilaments. The values were obtained on the quadrants of the skin of the breast, the areola and the nipple. The sensitivity test was performed preoperatively, then at 3 and 6 months postoperatively. Patients' characteristics (age, weight, breast ptosis, breast mass resected and risk factors) were statistically similar between the two groups. The preoperative values of pressure sensation on the different areas tested were statistically similar between the two groups. The sensitivity decreased on almost all the tested areas of the breast at 3 months postoperatively. No patient had an insensitive area on the breast at 6 months after surgery. Some areas of the breast showed a significant difference in pressure sensitivity after one technique compared to the other: better sensation on the skin of the superior and lateral quadrants after the superior pedicle technique at 3 months (P< 0.001), poorer areolar sensation on the inferior quadrant after the superior pedicle technique at 3 and 6 months (P< 0.05) and on the superior quadrant after the inferior pedicle technique at 3 months only (P< 0.05). However, the mean value of the areolar quadrants was statistically similar after both techniques. The nipple sensation was significantly decreased in both groups at 3 months but remained comparable between the two groups. Breast innervation was damaged by breast reduction using both the inferior and the superior pedicle techniques. The breast skin had better sensation after the superior pedicle technique while the areola had slightly better sensation after the inferior pedicle technique. At 6 months, the mean value of nipple-areola complex pressure sensation was comparable in the two series of patients. PMID- 11121317 TI - Breast sensation after superior pedicle versus inferior pedicle mammaplasty: anatomical and histological evaluation. AB - Despite contradictory information about the course and distribution of the nerves supplying the breast, surgical techniques using an inferior pedicle have been recommended over those using a superior pedicle for preserving the nipple-areolar sensation after surgery. This anatomical study was designed to quantify the nerve branches preserved in inferior and superior pedicles after reduction mammaplasty performed on cadavers. Reduction mammaplasty was done on four fresh cadavers (within 48h of death) using a superior pedicle on the right and an inferior pedicle on the left in a standard way. The pedicle was cut at its base and then fixed in formalin. The base was divided in biopsy specimens and embedded in paraffin. The nerves were quantified and located in each pedicle with haematoxylin-eosin stain and light-microscopic evaluation. Histological evaluation of the pedicles showed the presence of a variable number of nerves (between one and seven) within two superior pedicles and three inferior pedicles. The nerves were located in fibrous tissue and accompanied by vessels in most cases. The nerves were always found superficially and were most likely to be located in the central part of the pedicle. Our results showed that including the nerves within the pedicle is technically uncertain regardless of the mammaplasty technique used. The final recovery of sensation in the breast after mammaplasty seems to result from the regeneration of severed cutaneous nerve branches or the remaining cutaneous innervation rather than the preserved adjacent cutaneous branches. PMID- 11121318 TI - Results of artificial inflammation in scarless foetal wound healing: an experimental study in foetal lambs. AB - Recent experimental and clinical evidence suggests that the foetus responds to injury in a fashion fundamentally different from the adult. Foetal wound healing occurs without scar formation. The mechanisms causing this difference are still not well defined but absence of inflammation may play a significant role. The aim of this study was to investigate the effects of artificially induced inflammation on scarless foetal wound healing. Twelve time-dated pregnant ewes underwent hysterotomies at 70 and 90 days' gestation. A potent chemoattractant agent (N formyl-methionyl-leucyl-phenylalanine) was injected into the upper lip of the foetus in the first operation. In the second operation, a full-thickness incisional wound on the right side and a 3-mm excisional wound on the left side of the upper lip were created and closed primarily. A control wound was created on the lower lip. Macroscopic and histologic examinations of the wounds after birth revealed visible scar lines on the upper-lip wounds but no scar line on the lower lip, and an increase in fibrous tissue and collagen content in the upper lip wounds. We have shown that injecting a chemotactic agent can stimulate inflammation in a period of gestation when tissue injury can not. Although lack of inflammation due to tissue injury in the early period of foetal life may be an important cause of scarless healing, further experimental studies should be carried out to investigate the pathways that are not activated by tissue injury, the immune status of the foetus and the growth factors involved in the inflammatory response. PMID- 11121319 TI - The use of conventional and invaginated autologous vein grafts for nerve repair by means of entubulation. AB - Nerve repair by entubulation has re-emerged recently as a possible means of enhancing the microenvironment at the site of repair by inclusion within the tube of various trophic factors. To this end, a modification of the vein-graft technique has been used by turning it inside out before repair, to expose the adventitial surface to the regenerating axons. A comparative study of standard vein grafting versus the inside-out technique was carried out in two equal-sized groups of inbred Lewis rats. Jugular vein isografts were derived from litter mates. The sciatic nerve was transected and repaired by entubulation using the standard vein graft in one group and the inside-out graft in the other group. Morpho-metric and electrophysiological assessment were carried out 3 months after repair. When the animals were assessed it was found that both the standard-vein graft group and the inside-out group exhibited a reduction in all of the morphometric and electrophysiological variables when compared to normal nerves. The mean axon diameter, fibre diameter and myelin sheath thickness were, however, found to be greater in the group that underwent the inside-out repair. The superior morphometric results seen in the inside-out group were not matched by improved electrophysiological performance. It is concluded that the use of the inside-out technique confers no functional benefit over standard vein grafting. PMID- 11121320 TI - Preconditioning of the distal portion of a rat random-pattern skin flap. AB - It has been shown that preconditioning either by proximal pedicle clamping or by pedicle intravascular drug administration, for example with adenosine, can improve flap survival. These methods, however, are not well suited to random pattern flap transfer in the clinical setting. The aim of this study was to evaluate clinically applicable preconditioning methods for random-pattern flaps. Eighteen male Sprague-Dawley rats were used. Bipedicled dorsal skin flaps (2 x 8cm) containing panniculus carnosus were elevated. In the ischaemic preconditioning group the cranial pedicle was clamped for 20min, followed by 40min reperfusion before the cranial pedicle was cut, producing a caudally based random-pattern flap. In the pharmacologic preconditioning group adenosine was locally injected in the cranial half of the flap before the cranial pedicle was cut. In the control group saline was locally injected instead of adenosine and the pedicle was cut in the same manner. Flap survival area was evaluated at day 7. Flap survival area in both preconditioning groups was significantly higher than in the control group (P<0.05). Both preconditioning methods can improve random-pattern flap survival in rats. These methods may prove useful in the clinical setting. PMID- 11121321 TI - The short head of the biceps femoris as a monitor for the free lateral thigh flap in pharyngoesophageal reconstruction. AB - Free flaps are frequently used to reconstruct the defect following radical resection of pharyngoesophageal malignancy but postoperative monitoring of buried flaps is difficult. We have designed a monitoring-muscle flap using the short head of the biceps femoris muscle when using a free lateral thigh flap. The third and fourth perforators of the profunda femoris artery, the main vascular pedicle of the lateral thigh flap, pass through the short head of the biceps femoris. Partial excision of the short head of the biceps femoris muscle does not result in any functional disturbance of the leg, and the viability of the buried lateral thigh flap can be monitored by observing the exposed muscle through a small window in the neck. Between April and October 1998 five patients underwent pharyngoesophageal reconstruction by this method. The short head of the biceps femoris was used to monitor the main flap in three patients and to obliterate the dead space after neck dissection in two patients. There were no recipient-site complications such as fistula or infection and no disturbance of thigh function. PMID- 11121322 TI - Necrotising fasciitis of the breast. AB - Necrotising fasciitis is a rare condition and to the best of our knowledge has never been reported in the breast. We report the first case in the literature of necrotising fasciitis involving the breast. PMID- 11121323 TI - Salvage of completely degloved finger with a posterior interosseous free flap. AB - The use of a free posterior interosseous skin flap should be considered in a single digit degloving injury, especially when replantation of the avulsed skin or the use of skin from the second toe, transferred as a composite-tissue flap, is not feasible. The flap is thin and pliable. It allows early mobilisation with good recovery of joint motion and attains protective sensation of the finger. PMID- 11121324 TI - An unusual schwannoma of the median nerve: effects on the motor branch. AB - An unusual case of a schwannoma of the median nerve is presented where pressure due to the tumour on the motor branch to the thenar muscles caused weakness and wasting of the abductor pollicis brevis muscle, a previously unreported phenomenon. The patient achieved a full functional recovery after enucleation, which is also unusual considering the patient's age. Aspects of schwannoma biology, differential diagnosis, investigation and treatment are discussed. PMID- 11121325 TI - Giant foot schwannoma. AB - Schwannoma of the foot is rare; only 12 cases have been reported. A schwannoma is a benign neurogenic tumour derived from Schwann cells. The diagnosis is often delayed because the symptoms are mainly those of compression disorders. We describe a 7cm schwannoma of the heel in a 30-year-old man. Ten years earlier a schwannoma was removed from the same site. The recurrent lesion was widely excised and a medial plantar flap was used to repair the heel. PMID- 11121326 TI - A challenging treatment for an ischaemic ulcer in a patient with Buerger's disease: vascular reconstruction and local flap coverage. AB - An ischaemic heel ulcer in a patient with Buerger's disease was reconstructed using an in situ saphenous vein graft combined with a local flap. The bypass was sufficient to restore blood supply to the ischaemic limb but a flap was necessary to cover the persistent heel ulcer, which remained after revascularisation. One month after bypass surgery the ulcer was debrided and the resulting defect was covered with a lateral supramalleolar flap. The postoperative course was uneventful and the flap donor site healed well. When treating ischaemic ulcers in a patient with Buerger's disease, vascular reconstruction should be considered first in order to salvage the limb. After revascularisation, a local flap can be used to cover a persistent defect but very few local flaps have been reported. This report is the first published case of successful local flap transfer after bypass surgery in a patient with Buerger's disease. We think that a local flap is one possible treatment for a non-healing ulcer after revascularisation. PMID- 11121327 TI - EBM--evidence before mortality? PMID- 11121328 TI - A simple technique to avoid inadvertent damage to monofilament core suture material during flexor-tendon repair. PMID- 11121329 TI - Head dressings for pinnaplasty: a tradition not supported by evidence. PMID- 11121330 TI - Hyperpigmentation following the use of Emla cream. PMID- 11121331 TI - Pyoderma gangrenosum: beware, it does recur. PMID- 11121332 TI - Escalator injuries to the foot. PMID- 11121333 TI - The dynamic activity work profile (DAWP): a simple way to illustrate waiting-list numbers, work load and clinic priorities for the disbelievers or the uninformed. PMID- 11121334 TI - Limb amputation and Behcet's disease. PMID- 11121335 TI - The use of stored skin grafts for keratinocyte cultures. PMID- 11121336 TI - Subcutaneous emphysema following the use of a high-pressure water jet. PMID- 11121337 TI - Sting in the tail. PMID- 11121338 TI - Stop those hopping mad surgeons. PMID- 11121339 TI - Unifying principles of locomotion: foreword. PMID- 11121340 TI - Costs of locomotion and vertic dynamics of cephalopods and fish. AB - The world's oceans are three-dimensional habitats that support high diversity and biomass. Because the densities of most of the constituents of life are greater than that of seawater, planktonic and pelagic organisms had to evolve a host of mechanisms to occupy the third dimension. Some microscopic organisms survive at the surface by dividing rapidly in vertically well mixed zones, but most organisms, small and large, have antisinking strategies and structures that maintain vertical position and mobility. All of these mechanisms have energetic costs, ranging from the "foregone metabolic benefits" and increased drag of storing high-energy, low-density lipids to direct energy consumption for dynamic lift. Defining the niches in the mesopelagic zone, understanding evolution, and applying such ecological concepts as optimal foraging require good estimates of these costs. The extreme cases above are reasonably well quantified in fishes, but the energetic costs of dynamic physiological mechanisms like swim bladders are not; nor are the costs of maintaining vertical position for the chief invertebrate competitors, the cephalopods. This article evaluates a matrix of buoyancy mechanisms in different circumstances, including vacuum systems and ammonium storage, based on new data on the metabolic cost of creating buoyancy in Sepia officinalis. PMID- 11121341 TI - Boxfishes as unusually well-controlled autonomous underwater vehicles. AB - Boxfishes (family Ostraciidae) are tropical reef-dwelling marine bony fishes that have about three-fourths of their body length encased in a rigid bony test. As a result, almost all of their swimming movements derive from complex combinations of movements of their median and paired fins (MPF locomotion). In terms of both body design and swimming performance, they are among the most sophisticated examples known of naturally evolved vertebrate autonomous underwater vehicles. Quantitative studies of swimming performance, biomechanics, and energetics in one model species have shown that (i) they are surprisingly strong, fast swimmers with great endurance; (ii) classical descriptions of how they swim were incomplete: they swim at different speeds using three different gaits; (iii) they are unusually dynamically well controlled and stable during sustained and prolonged rectilinear swimming; and (iv) despite unusually high parasite (fuselage) drag, they show energetic costs of transport indistinguishable from those of much better streamlined fishes using body and caudal fin (BCF) swimming modes at similar water temperatures and over comparable ranges of swimming speeds. We summarize an analysis of these properties based on a dynamic model of swimming in these fishes. This model accounts for their control, stability, and efficiency in moving through the water at moderate speeds in terms of gait changes, of water-flow patterns over body surfaces, and of complex interactions of thrust vectors generated by fin movements. PMID- 11121342 TI - Aquatic and terrestrial locomotory energetics in a toad and a turtle: a search for generalisations among ectotherms. AB - Murray short-necked turtles were trained to walk on a motorised treadmill and to swim in a recirculating flume. Through filmed records, the frequency of limb movement and the time that thrust was directed against the substrate were measured. The animals wore masks when walking and accessed air when swimming from a ventilated capsule placed on top of the water surface. Measurement of the exhalant O(2) and CO(2) levels from these devices enabled the measurement of metabolic rates. Equivalent data were obtained from swimming and hopping cane toads, although repeatable measures of limb frequency and contact times were not obtained due to the intermittent form of locomotion in this species. Comparing the cost of transport, the energy required to transport a mass of animal over a unit distance, with other animals showed that toads do not have a cheap form of terrestrial locomotion, but turtles do; turtles use half the cost predicted from their body mass. This economy of locomotion is consistent with what is known about turtle muscle, the mechanics of their gait, and the extremely long contact time for a limb with the substrate. Swimming in toads is energetically expensive, whereas turtles, on the basis of mass, use about the same energy to transport a unit mass as an equivalent-size fish. The data were compared with the predictions of the Kram-Taylor hypothesis for locomotory scaling, and walking turtles were found to provide a numerical fit. The data show that both terrestrial and aquatic locomotory energetics in toads are generally higher than predictions on the basis of mass, whereas in turtles they are lower. PMID- 11121343 TI - Biomechanics and energetics in aquatic and semiaquatic mammals: platypus to whale. AB - A variety of mammalian lineages have secondarily invaded the water. To locomote and thermoregulate in the aqueous medium, mammals developed a range of morphological, physiological, and behavioral adaptations. A distinct difference in the suite of adaptations, which affects energetics, is apparent between semiaquatic and fully aquatic mammals. Semiaquatic mammals swim by paddling, which is inefficient compared to the use of oscillating hydrofoils of aquatic mammals. Semiaquatic mammals swim at the water surface and experience a greater resistive force augmented by wave drag than submerged aquatic mammals. A dense, nonwettable fur insulates semiaquatic mammals, whereas aquatic mammals use a layer of blubber. The fur, while providing insulation and positive buoyancy, incurs a high energy demand for maintenance and limits diving depth. Blubber contours the body to reduce drag, is an energy reserve, and suffers no loss in buoyancy with depth. Despite the high energetic costs of a semiaquatic existence, these animals represent modern analogs of evolutionary intermediates between ancestral terrestrial mammals and their fully aquatic descendants. It is these intermediate animals that indicate which potential selection factors and mechanical constraints may have directed the evolution of more derived aquatic forms. PMID- 11121344 TI - Energetic costs of surface swimming and diving of birds. AB - The energetic costs of swimming at the surface (swimming) and swimming underwater (diving) are compared in tufted ducks (Aythya fuligula) and three species of penguins, the gentoo (Pygoscelis papua), the king (Aptenodytes patagonicus), and the emperor (Aythya forsteri). Ducks swim on the surface and use their webbed feet as paddles, whereas penguins tend to swim just below the surface and use their flippers as hydrofoils, the latter being much more efficient. Penguins are more streamlined in shape. Thus, the amount of energy required to transport a given mass of bird a given distance (known as the cost of transport) is some two to three times greater in ducks than in penguins. Ducks are also very buoyant, and overcoming the force of buoyancy accounts for 60% and 85% of the cost of descent and remaining on the bottom, respectively, in these birds. The energy cost of a tufted duck diving to about 1.7 m is similar to that when it is swimming at its maximum sustainable speed at the surface (i.e., approximately 3.5 times the value when resting on water). Nonetheless, because of the relatively short duration of its dives, the tufted duck dives well within its calculated aerobic dive limit (cADL, usable O(2) stores per rate of O(2) usage when underwater). However, these three species of penguins have maximum dive durations ranging from 5 min to almost 16 min and maximum dive depths from 155 to 530 m. When these birds dive, they have to metabolise at no more than when resting in water in order for cADL to encompass the duration of most of their natural dives. In gentoo and king penguins, there is a fall in abdominal temperature during bouts of diving; this may reduce the oxygen requirements in the abdominal region, thus enabling dive duration to be extended further than would otherwise be the case. PMID- 11121345 TI - Locomotion, respiratory physiology, and energetics of amphibious and terrestrial crabs. AB - The transition from breathing air to breathing water requires physiological and morphological adaptations. The study of crustaceans in transitional habitats provides important information as to the nature of these adaptations. This article addresses the physiology of air breathing in amphibious and terrestrial crabs and their relative locomotor abilities. Potamonautes warreni is an apparently amphibious freshwater crab from southern Africa, Cardisoma hirtipes is an air-breathing gecarcinid crab with some dependency on freshwater, and Gecarcoidea natalis is an obligate air-breathing gecarcinid endemic to Christmas Island in the Indian Ocean. All three species have well-developed lungs but retain gills and show seasonally different activity patterns that, in the gercarcinids, especially G. natalis, include long-distance breeding migrations. The three species were better at breathing air than water, but P. warreni was the best at breathing water. Cardisoma hirtipes is essentially an obligate air breather and appears to experience facultative hypometabolism during immersion. Cardisoma hirtipes has a haemocyanin with a high affinity for O(2) that facilitates loading from air but makes 30% of the Hc bound O(2) inaccessible. The gecarcinids but not P. warreni show increased diffusion limitation for O(2) over the lung during exercise. Gecarcoidea natalis outperforms C. hirtipes by virtue of a unique haemolymph shunt from the lung into the gills. Paradoxically, it is modifications of the gills for aerial O(2) uptake in G. natalis that allow for relatively greater haemolymph oxygenation. Despite showing decreased arterial venous DeltaPo(2), P. warreni increased the arterial-venous Delta[O(2)] with no recourse to anaerobiosis during 5 min exercise. In the short term, P. warreni is more adept at walking than C. hirtipes. The breeding migrations of C. hirtipes and G. natalis were completely aerobic, but G. natalis walk farther and probably faster. Seasonal changes in underlying metabolism of G. natalis are strongly implied, including variations in hyperglycaemic hormone, variable basal metabolic rates, and a diel alkalosis present only in migrating crabs. The persistent dependence on water for reproduction is a determining factor in the biology of air-breathing crabs. The annual migrations include costs other than locomotion, for example, burrow construction and intermale competition. Estimates of costs that consider walking alone will underestimate the metabolic and stored fuel requirements for successful reproduction. PMID- 11121346 TI - Unifying principles in terrestrial locomotion: do hopping Australian marsupials fit in? AB - Mammalian terrestrial locomotion has many unifying principles. However, the Macropodoidea are a particularly interesting group that exhibit a number of significant deviations from the principles that seem to apply to other mammals. While the properties of materials that comprise the musculoskeletal system of mammals are similar, evidence suggests that tendon properties in macropodoid marsupials may be size or function dependent, in contrast to the situation in placental mammals. Postural differences related to hopping versus running have a dramatic effect on the scaling of the pelvic limb musculoskeletal system. Ratios of muscle fibre to tendon cross-sectional areas for ankle extensors and digital flexors scale with positive allometry in all mammals, but exponents are significantly higher in macropods. Tendon safety factors decline with increasing body mass in mammals, with eutherians at risk of ankle extensor tendon rupture at a body mass of about 150 kg, whereas kangaroos encounter similar problems at a body mass of approximately 35 kg. Tendon strength appears to limit locomotor performance in these animals. Elastic strain energy storage in tendons is mass dependent in all mammals, but exponents are significantly larger in macropodid. Tibial stresses may scale with positive allometry in kangaroos, which result in lower bone safety factors in macropods compared to eutherian mammals. PMID- 11121347 TI - Biomechanics and physiology of gait selection in flying birds. AB - Two wing-beat gaits, distinguished by the presence or absence of lift production during the upstroke, are currently used to describe avian flight. Vortex visualization studies indicate that lift is produced only during the downstroke in the vortex-ring gait and that lift is produced continuously in the continuous vortex gait. Tip-reversal and feathered upstrokes represent different forms of vortex-ring gait distinguished by wing kinematics. Useful aerodynamic forces may be produced during tip-reversal upstroke in slow flight and during a feathered upstroke in fast flight, but it is probable that downstroke forces are much greater in magnitude. Uncertainty about the function of these types of upstroke may be resolved when more data are available on wake structure in different flight speeds and modes. Inferring from wing kinematics and available data on wake structure, birds with long wings or wings of high aspect ratio use a vortex ring gait with tip-reversal upstroke at slow speeds, a vortex-ring gait with a feathered upstroke at intermediate speeds, and a continuous-vortex gait at fast speeds. Birds with short wings or wings of low aspect ratio use a vortex-ring gait with a feathered upstroke at all speeds. Regardless of wing shape, species tend to use a vortex-ring gait for acceleration and a continuous-vortex gait for deceleration. Some correlations may exist between gait selection and the function of the muscular and respiratory system. However, overall variation in wing kinematics, muscle activity, and respiratory activity is continuous rather than categorical. To further our understanding of gait selection in flying birds, it is important to test whether upstroke function varies in a similar manner. Transitions between lifting and nonlifting upstrokes may be more subtle and gradual than implied by a binomial scheme of classification. PMID- 11121348 TI - Surface-skimming stoneflies and mayflies: the taxonomic and mechanical diversity of two-dimensional aerodynamic locomotion. AB - The best supported hypothesis for the evolutionary origin of insect wings is that they evolved from articulated, leg-derived respiratory structures of aquatic ancestors. However, there are no fossils of the immediate ancestors of winged insects, and it is difficult to imagine how a functional transition from gills to wings could have occurred. Recent studies of surface-skimming locomotion in stoneflies and mayflies offer a plausible solution by showing how rudimentary wings and muscle power can be used to accomplish two-dimensional aerodynamic locomotion on the surface of water. Here we extend that line of research by examining the phylogenetic distribution and mechanistic diversity of surface skimming in stoneflies, along with a limited examination of mayflies. These investigations reveal both a broad taxonomic occurrence and a fine gradation of mechanically distinct forms. Distinct forms of wing-flapping surface skimming include (1) stoneflies that flap their wings weakly while maintaining their body in contact with the water and undulating their abdomen laterally in a swimming like motion, (2) stoneflies that skim while elevating their body above the water and maintaining all six legs on the surface, (3) stoneflies and mayflies that skim with only four legs on the water surface, (4) stoneflies that skim with only their two hind legs on the surface, and (5) stoneflies that, beginning with a series of leg motions nearly identical to hind-leg skimmers, use their hind legs to jump from the water into the air to initiate flapping flight. Comparisons across these forms of skimming show that wing-beat amplitude, horizontal velocity, and the verticality of aerodynamic force production increase as the body orientation becomes more upright and contact with the water is minimized. These behaviors illustrate a mechanical pathway by which flying insects could have evolved from swimming ancestors via a series of finely graded intermediate stages. The phylogenetic distribution of skimming and flight in stoneflies does not indicate any clear directionality toward either greater or lesser aerodynamic abilities; however, the broad and apparently basal phylogenetic distribution of skimming taxa supports the hypothesis that the common ancestor of stoneflies was a surface skimmer. This may also be true for the common ancestor of stoneflies and mayflies, that is, the first winged insects. We combine these data with fossil evidence to form a synthetic model for the evolution of flying insects from surface skimmers. PMID- 11121349 TI - Energy metabolism during insect flight: biochemical design and physiological performance. AB - Flying insects achieve the highest known mass-specific rates of O(2) consumption in the animal kingdom. Because the flight muscles account for >90% of the organismal O(2) uptake, accurate estimates of metabolic flux rates (J) in the muscles can be made. In steady state, these are equal to the net forward flux rates (v) at individual steps and can be compared with flux capacities (V(max)) measured in vitro. In flying honeybees, hexokinase and phosphofructokinase, both nonequilibrium reactions in glycolysis, operate at large fractions of their maximum capacities (i.e., they operate at high v/V(max)). Phosphoglucoisomerase is a reversible reaction that operates near equilibrium. Despite V(max) values more than 20-fold greater than the net forward flux rates during flight, a close match is found between the V(max) required in vivo (estimated using the Haldane relationship) to maintain near equilibrium and this net forward flux rate and the V(max) measured in vitro under simulated physiological conditions. Rates of organismal O(2) consumption and difference spectroscopy were used to estimate electron transfer rates per molecule of respiratory chain enzyme during flight. These are much higher than those estimated in mammalian muscles. Current evidence indicates that metabolic enzymes in honeybees do not display higher catalytic efficiencies than the homologous enzymes in mammals, and the high electron transfer rates do not appear to be the result of higher enzyme densities per unit cristae surface area. A number of possible mechanistic explanations for the higher rates of electron transfer are proposed. PMID- 11121350 TI - Digestibility, nitrogen excretion, and mean retention time by North American porcupines (Erethizon dorsatum) consuming natural forages. AB - North American porcupines (Erethizon dorsatum) subsist predominantly on low protein, high-fiber, high-tannin diets. Therefore, we measured the porcupine's ability to digest dry matter, fiber, and protein by conducting digestion trials on eight natural forages and one pelleted ration varying in concentration of fiber, nitrogen, and tannins. On these diets, dry matter intake ranged from 5 to 234 g/kg(0.75)/d and dry matter digestibility ranged from 62% to 96%. Porcupines digested highly lignified fiber better than many large hindgut fermenters and ruminants. The porcupine's ability to digest fiber may be explained, in part, by their lengthy mean retention time of particles (38.43+/-0.56 h). True nitrogen digestibility was 92% for nontannin forages and pellets. Endogenous urinary nitrogen was 205 mg N/kg(0.75)/d, and metabolic fecal nitrogen was 2.8 g N/kg dry matter intake. Porcupines achieved nitrogen balance at relatively low levels of nitrogen intake (346 mg N/kg(0.75)/d). Tannins reduced the porcupines' ability to digest protein. However, the reduction in protein digestion was not predictable from the amount of bovine serum albumin precipitated. Like many herbivores, porcupines may ameliorate the effects of certain tannins in natural forages on protein digestibility through physiological and behavioral adaptations. PMID- 11121351 TI - Effect of metabolic acidosis on white-tailed deer antler development. AB - Metabolic acidosis can result when herbivores consume browse diets high in plant secondary compounds. One mechanism for buffering excess acid is the mobilization of calcium and other alkaline salts from the skeletal system. White-tailed deer (Odocoileus virginianus) and other cervids consuming browse during antler formation may use minerals essential for antler development as buffers, resulting in altered antler characteristics. Our research objectives were to examine the effects of metabolic acidosis on mineral metabolism, acid-base homeostasis, and antler development in white-tailed deer. Fifteen male white-tailed deer were assigned to one of three diets: 2% NH(4)Cl, 3% commercial tannic acid, or a basal ration without additive. Two feeding trials were completed on each deer to determine nutrient use. Urine pH and the percentage of urinary nitrogen excreted as NH+4 varied by diet. No significant diet or trial effects occurred for nitrogen, calcium, phosphorus, magnesium, or sodium use. Urinary calcium excretion varied between diets. No dietary differences were observed for antler characteristics. The NH(4)Cl diet induced metabolic acidosis but did not alter antler development in white-tailed deer. Skeletal mineral reserves and mineral intake appeared sufficient to buffer excess acids and support antler development. PMID- 11121352 TI - Metabolic limits on dive duration and swimming speed in the southern elephant seal Mirounga leonina. AB - The ability of air-breathing marine predators to forage successfully depends on their ability to remain submerged. This is in turn related to their total O(2) stores and the rate at which these stores are used up while submerged. Body size was positively related to dive duration in a sample of 34 adult female southern elephant seals from Macquarie Island. However, there was no relationship between body size and dive depth. This indicates that smaller seals, with smaller total O(2) stores, make shorter dives than larger individuals but operate at similar depths, resulting in less time being spent at depth. Nine adult female elephant seals were also equipped with velocity time depth recorders. In eight of these seals, a plot of swimming speed against dive duration revealed a cloud of points with a clear upper boundary. This boundary could be described using regression analysis and gave a significant negative relationship in most cases. These results indicate that metabolic rate varies with activity levels, as indicated by swimming speed, and that there are quantifiable limits to the distance that a seal can travel on a dive of a given swimming speed. However, the seals rarely dive to these physiological limits, and the majority of their dives are well within their aerobic capacity. Elephant seals therefore appear to dive in a way that ensures that they have a reserve of O(2) available. PMID- 11121353 TI - Depression of protein synthesis during diapause in embryos of the annual killifish Austrofundulus limnaeus. AB - Rates of protein synthesis are substantially depressed in diapause II embryos of Austrofundulus limnaeus. Inhibition of oxygen consumption and heat dissipation with cycloheximide indicates that 36% of the adenosine triphosphate (ATP) turnover in prediapausing embryos (8 d postfertilization [dpf]) is caused by protein synthesis; the contribution of protein synthesis to ATP turnover in diapause II embryos is negligible. In agreement with the metabolic data, incorporation of amino acids (radiolabeled via (14)CO(2)) into perchloric acid precipitable protein decreases by over 93% in diapause II embryos compared with embryos at 8 dpf. This result represents a 36% reduction in energy demand because of depression of protein synthesis during diapause. Adjusting for changes in the specific radioactivity of the free amino acid pool at the whole-embryo level yields rates of protein synthesis that are artifactually high and not supportable by the observed rates of oxygen consumption and heat dissipation during diapause. This result indicates a regionalized distribution of labeled amino acids likely dictated by a pattern of anterior to posterior cell cycle arrest. AMP/ATP ratios are strongly correlated with the decrease in rates of protein synthesis, which suggests a role for adenosine monophosphate (AMP) in the control of anabolic processes. The major depression of protein synthesis during diapause II affords a considerable reduction in energy demand and extends the duration of dormancy attainable in these embryos. PMID- 11121354 TI - Unappreciated tolerance to high ambient temperatures in a widely distributed desert rodent, Dipodomys merriami. AB - A long-held assertion has been that nocturnality is an escape mechanism for many nocturnal desert rodents because of limited tolerances to heat. To test this claim, we used a treadmill to examine the tolerances to high ambient temperatures (T(a)'s) of one subspecies of desert rodent, Merriam's kangaroo rat, Dipodomys merriami merriami, from contrasting environments. We simultaneously measured body temperature (T(b)), evaporative water loss, and metabolic rates at an ecologically relevant speed (0.6 km h(-1)) at different ambient temperatures (Ta=25 degrees -42.5 degrees C). We hypothesized that kangaroo rats from a more xeric site would have greater abilities to remain active and maintain stable T(b) than those from a more mesic site, but mesic- and xeric-site animals had comparable tolerances and were active until Tb=42 degrees C. At Ta=42.5 degrees C, however, T(b) of mesic-site animals increased more quickly than in xeric-site animals. Although most animals could not run more than 18 min at Ta=42.5 degrees C, most could run at Ta=40 degrees C for at least 30 min. Benefits of nocturnality for this species may reside more in purposes of water conservation and avoidance of predation and less on the direct regulation of T(b), as T(b) is more labile than commonly thought. PMID- 11121355 TI - Allometric scaling of maximal enzyme activities in the axial musculature of striped bass, Morone saxatilis (Walbaum). AB - It had been suggested that the activity of anaerobic enzymes in the white muscle of fish increases exponentially with body size to meet the increasing hydrodynamic costs of burst swimming. We tested whether this relationship holds across a very large size range of striped bass, spanning a nearly 3,000-fold range in body mass. We examined the scaling of marker enzymes of anaerobic (lactate dehydrogenase and pyruvate kinase) and aerobic (citrate synthase and malate dehydrogenase) metabolism in the red and white locomotor muscles. In white muscle, we found positive scaling of anaerobic enzymes only in smaller fishes. Positive scaling of anaerobic enzymes was not found among the samples that included fishes >1,000 g despite having a sufficiently large sample size to detect such scaling. The absence of positive scaling in the white muscles of large bass suggests that they are unable to generate sufficient power to sustain relative burst swimming performance. Enzymes from aerobic pathways had activities that were mass independent in both red and white muscle. Red and white muscles were metabolically distinct except among the smallest fishes. Among young of the year, the anaerobic capacity of red muscle approached that of white muscle and also showed positive scaling. This unusual pattern suggests that red muscle might augment white muscle during burst swimming and add to the total power generated by these small fish. Maximizing burst swimming performance may be critical for small fishes vulnerable to predation but unimportant for large fishes. PMID- 11121356 TI - Effects of temperature on energy cost and timing of embryonic and larval development of the terrestrially breeding moss frog, Bryobatrachus nimbus. AB - The Australian moss frog, Bryobatrachus nimbus, oviposits four to 16 large eggs in terrestrial nests constructed in moss or lichen in subalpine regions of southern Tasmania. Nidicolous larvae overwinter beneath snow, reaching metamorphosis without feeding after 395 d, the longest development time known for an endotrophic anuran. However, a few clutches develop more quickly and metamorphose before winter. This study examines the effect of temperature on development time and energy expenditure by measuring temperatures and developmental stages in field nests as well as rates of oxygen consumption (Vo2), developmental stage, body mass, and energy content in the laboratory at three relevant temperatures (5 degrees, 10 degrees, 15 degrees C). Eggs and larvae reared at 5 degrees C differentiated very slowly, and their development time far exceeded those in natural nests, but development times at 10 degrees and 15 degrees C averaged 277 and 149 d, respectively, and were shorter than field incubation times. Generally, respiration rates of aquatic hatchlings were low in comparison with other species but increased with larval age and jumped about 25% higher near metamorphosis when larvae were able to air breathe. The mean energy density was 26.0 J mg(-1) for the dry ova and 20.6 J mg(-1) for a dry gut-free froglet, and total production efficiency was 61.5%. We developed a model based on the relationships between incubation temperature and V&d2;o2 to estimate the respiratory cost of development to metamorphosis, the first such study for an amphibian. The cost was 177 J at 15 degrees C, 199 J at 10 degrees C, and at least 249 J at 5 degrees C, and we predicted that continual development at 5 degrees C would lead to premature yolk depletion because it equalled the 249 J contained in fresh ova. Continuously logged field-nest temperatures and interpolation of laboratory data provided estimates of development rates, Vo2, and respiratory energy costs in field nests. Development to metamorphosis required between 185 and 234 J when larvae overwintered, but completion of metamorphosis before winter saved 123 J. However, the advantage of emergence in warmer months, when conditions are suitable for feeding and growth, may offset the greater energy cost of overwintering. PMID- 11121358 TI - Biomedical research and the environment. PMID- 11121357 TI - Evolutionary and physiological variation in cardiac troponin C in relation to thermal strategies of fish. AB - Striated muscle contraction is initiated when troponin C (TnC) binds Ca(2+), which activates actinomyosin ATPase. We investigated (i) the variation between cardiac TnC (cTnC) primary structure within teleost fish and (ii) the pattern of TnC expression in response to temperature acclimation. There were few differences between rainbow trout (Oncorhynchus mykiss), yellowfin tuna (Thunnus albacares), yellow perch (Perca flavescens), goldfish (Carassius auratus), white sucker (Catostomus commersoni), and icefish (Chaenocephalus aceratus) in cTnC amino acid sequence. No variation existed in the regulatory Ca(2+)-binding site (site 2). The site 3 and 4 substitutions were limited to residues not directly involved in Ca(2+) coordination. Fish cTnC primary structure was highly conserved between species (93%-98%) and collectively divergent from the highly conserved sequence seen in birds and mammals. Northern blots and polymerase chain reaction showed that thermal acclimation of trout (3 degrees, 18 degrees C) did not alter the TnC isoform pattern. While cardiac and white muscle had the expected isoforms-cTnC and fast troponin C (fTnC), respectively-red muscle unexpectedly expressed primarily ftnC. Cold acclimation did not alter myofibrillar ATPase Ca(2+) sensitivity, but maximal velocity increased by 60%. We found no evidence that TnC variants, arising between species or in response to thermal acclimation, play a major role in mitigating the effects of temperature on contractility of the adult fish heart. PMID- 11121359 TI - Activities of the National Institutes of Health relating to energy efficiency and pollution prevention. AB - The National Institutes of Health (NIH) is one of the world's premier biomedical research centers. Although NIH owns and operates more than 1,300 acres and 197 buildings across the country, the main campus is in Bethesda, Maryland. This campus consists of over 312 acres and 75 laboratories and other buildings, which consume vast amounts of energy. Aware of the NIH role in setting biomedical research agendas and priorities, its administrators strive to set good examples in energy efficiency and pollution prevention. Three current projects are presented as "best practices" examples of meeting the stated commitment of NIH to leadership in environmental stewardship: a) design and current construction of a 250-bed clinical research hospital designed to allow conversion of patient care units to research laboratories and vice-versa; b) design and construction of a six-story research laboratory that combines energy-saving innovations with breakthroughs in research technologies; and c) a massive, $200-million modernization of the campus utility infrastructure that involves generation systems for steam and chilled water and distribution systems for chilled water, steam, potable water, electricity, communications and computer networking, compressed air, and natural gas. Based on introduction of energy-efficiency measures, millions of dollars in savings for energy needs are projected; already the local electric utility has granted several million dollars in rebates. The guiding principles of NIH environmental stewardship help to ensure that energy conservation measures maximize benefits versus cost and also balance expediency with efficiency within available funding resources. This is a committee report for the Leadership Conference: Biomedical Research and the Environment held 1--2 November 1999 at the National Institutes of Health, Bethesda, Maryland. PMID- 11121360 TI - Environmental practices for biomedical research facilities. AB - As a result of the Leadership Conference on Biomedical Research and the Environment, the Facilities Committee focused its work on the development of best environmental practices at biomedical research facilities at the university and independent research facility level as well as consideration of potential involvement of for-profit companies and government agencies. The designation "facilities" includes all related buildings and grounds, "green auditing" of buildings and programs, purchasing of furnishings and sources, energy efficiency, and engineering services (lighting, heating, air conditioning), among other activities. The committee made a number of recommendations, including development of a national council for environmental stewardship in biomedical research, development of a system of green auditing of such research facilities, and creation of programs for sustainable building and use. In addition, the committee recommended extension of education and training programs for environmental stewardship, in cooperation with facilities managers, for all research administrators and researchers. These programs would focus especially on graduate fellows and other students, as well as on science labs at levels K--12. PMID- 11121361 TI - Development of a pollution prevention and energy efficiency clearinghouse for biomedical research facilities. AB - This is the report of the National Association of Physicians for the Environment Committee on Development of a Pollution Prevention and Energy Efficiency Clearinghouse for Biomedical Research Facilities from the Leadership Conference on Biomedical Research and the Environment held at the National Institutes of Health in Bethesda, Maryland, on 1--2 November 1999. A major goal of the conference was the establishment of a World Wide Web-based clearinghouse, which would lend tremendous resources to the biomedical research community by providing access to a database of peer-reviewed articles and references dealing with a host of aspects of biomedical research relating to energy efficiency, pollution prevention, and waste reduction. A temporary website has been established with the assistance of the U.S. Environmental Protection Agency (EPA) Regions III and IV, where a pilot site provides access to the EPA's existing databases on these topics. A system of peer review for articles and promising techniques still must be developed, but a glimpse of topics and search engines is available for comment and review on the EPA Region IV-supported website (http://wrrc.p2pays.org/). PMID- 11121363 TI - Biomedical research leaders: report on needs, opportunities, difficulties, education and training, and evaluation. AB - The National Association of Physicians for the Environment (NAPE) has assumed a leadership role in protecting environmental health in recent years. The Committee of Biomedical Research Leaders was convened at the recent NAPE Leadership Conference: Biomedical Research and the Environment held on 1--2 November 1999, at the National Institutes of Health, Bethesda, Maryland. This report summarizes the discussion of the committee and its recommendations. The charge to the committee was to raise and address issues that will promote and sustain environmental health, safety, and energy efficiency within the biomedical community. Leaders from every important research sector (industry laboratories, academic health centers and institutes, hospitals and care facilities, Federal laboratories, and community-based research facilities) were gathered in this committee to discuss issues relevant to promoting environmental health. The conference and this report focus on the themes of environmental stewardship, sustainable development and "best greening practices." Environmental stewardship, an emerging theme within and outside the biomedical community, symbolizes the effort to provide an integrated, synthesized, and concerted effort to protect the health of the environment in both the present and the future. The primary goal established by the committee is to promote environmentally responsible leadership in the biomedical research community. Key outcomes of the committee's discussion and deliberation were a) the need for a central organization to evaluate, promote, and oversee efforts in environmental stewardship; and b) immediate need to facilitate efficient information transfer relevant to protecting the global environment through a database/clearinghouse. Means to fulfill these needs are discussed in this report. PMID- 11121362 TI - Minimization and management of wastes from biomedical research. AB - Several committees were established by the National Association of Physicians for the Environment to investigate and report on various topics at the National Leadership Conference on Biomedical Research and the Environment held at the 1--2 November 1999 at the National Institutes of Health in Bethesda, Maryland. This is the report of the Committee on Minimization and Management of Wastes from Biomedical Research. Biomedical research facilities contribute a small fraction of the total amount of wastes generated in the United States, and the rate of generation appears to be decreasing. Significant reductions in generation of hazardous, radioactive, and mixed wastes have recently been reported, even at facilities with rapidly expanding research programs. Changes in the focus of research, improvements in laboratory techniques, and greater emphasis on waste minimization (volume and toxicity reduction) explain the declining trend in generation. The potential for uncontrolled releases of wastes from biomedical research facilities and adverse impacts on the general environment from these wastes appears to be low. Wastes are subject to numerous regulatory requirements and are contained and managed in a manner protective of the environment. Most biohazardous agents, chemicals, and radionuclides that find significant use in research are not likely to be persistent, bioaccumulative, or toxic if they are released. Today, the primary motivations for the ongoing efforts by facilities to improve minimization and management of wastes are regulatory compliance and avoidance of the high disposal costs and liabilities associated with generation of regulated wastes. The committee concluded that there was no evidence suggesting that the anticipated increases in biomedical research will significantly increase generation of hazardous wastes or have adverse impacts on the general environment. This conclusion assumes the positive, countervailing trends of enhanced pollution prevention efforts by facilities and reductions in waste generation resulting from improvements in research methods will continue. PMID- 11121364 TI - Applying environmental product design to biomedical products research. AB - The principal themes for the Biomedical Research and the Environment Conference Committee on Environmental Economics in Biomedical Research include the following: healthcare delivery companies and biomedical research organizations, both nonprofit and for-profit, need to improve their environmental performance; suppliers of healthcare products will be called upon to support this need; and improving the environmental profile of healthcare products begins in research and development (R&D). The committee report begins with requirements from regulatory authorities (e.g., U.S. Environmental Protection Agency [EPA], the U.S. Food and Drug Administration), and the healthcare delivery sector). The 1998 American Hospital Association and EPA Memorandum of Understanding to reduce solid waste and mercury from healthcare facilities is emblematic of these requirements. The dominant message from the requirements discussion is to ensure that R&D organizations do not ignore customer, environmental, and regulatory requirements in the early stages of product development. Several representatives from healthcare products manufacturers presented their companies' approaches to meeting these requirements. They reported on efforts to ensure that their R&D processes are sensitive to the environmental consequences from manufacturing, distributing, using, and disposing of healthcare products. These reports describe representatives' awareness of requirements and the unique approaches their R&D organizations have taken to meet these requirements. All representatives reported that their R&D organizations have embraced environmental product design because it avoids the potential of returning products to R&D to improve the environmental profile. Additionally, several reports detailed cost savings, sustainability benefits, and improvements in environmental manufacturing or redesign, and increased customer satisfaction. Many companies in healthcare delivery are working to improve environmental performance. Fundamental to these efforts is the necessity of motivating suppliers to improve the environmental profile of new products used in the healthcare delivery sector. PMID- 11121365 TI - Reducing environmental risk associated with laboratory decommissioning and property transfer. AB - The need for more or less space is a common laboratory problem. Solutions may include renovating existing space, leaving or demolishing old space, or acquiring new space or property for building. All of these options carry potential environmental risk. Such risk can be the result of activities related to the laboratory facility or property (e.g., asbestos, underground storage tanks, lead paint), or the research associated with it (e.g., radioactive, microbiological, and chemical contamination). Regardless of the option chosen to solve the space problem, the potential environmental risk must be mitigated and the laboratory space and/or property must be decommissioned or rendered safe prior to any renovation, demolition, or property transfer activities. Not mitigating the environmental risk through a decommissioning process can incur significant financial liability for any costs associated with future decommissioning cleanup activities. Out of necessity, a functioning system, environmental due diligence auditing, has evolved over time to assess environmental risk and reduce associated financial liability. This system involves a 4-phase approach to identify, document, manage, and clean up areas of environmental concern or liability, including contamination. Environmental due diligence auditing includes a) historical site assessment, b) characterization assessment, c) remedial effort and d) final status survey. General practice standards from the American Society for Testing and Materials are available for conducting the first two phases. However, standards have not yet been developed for conducting the third and final phases of the environmental due diligence auditing process. Individuals involved in laboratory decommissioning work in the biomedical research industry consider this a key weakness. PMID- 11121366 TI - The effect of sugar-free green tea chew candies on the degree of inflammation of the gingiva. AB - The components of green tea extracts such as catechins and polyphenols gain increasing significance in tumor research and immunology. - The clinical double blind study presented here was aimed at the investigation on how green tea catechins and polyphenols in the form of green tea dragees may influence the inflammatory behaviour of the gingiva. A total of 47 test persons with a mean age of 25.76 years (23 males, 24 females) were randomly divided into two groups: one group (n = 22: 11 males, 11 females) received chew candies containing green tea extracts, the other group (n = 25: 12 males, 13 females) received placebos with the same flavour but without active substances. At the beginning of the four week investigation period, a professional dental cleaning was carried out on all test persons. Then the persons were instructed to do their usual dental cleaning and chew eight candies distributed over the day. The API (approximal plaque index) and the SBI (sulcus bleeding index) were determined after seven days (API-1, SBI 1) and after another 21 days (API-2, SBI-2). Within the verum group, a mean value of 33.2% +/- 18.3% was determined for API-1, and 29.6% +/- 17.5% for API-2. The mean SBI-1 was 5.9% +/- 7.6%, and 3.6% +/- 5.8% for SBI-2. The clinical data within the placebo group were different: The plaque index values changed from API 1 30.3% +/- 16.3% after one week to API-2 31.8% +/- 17.2% after another three weeks. The values for the inflammatory degree of the gingiva had also changed to the negative: from SBI-1 3.4% +/- 4.1% after seven days to SBI-2 4.7% +/- 6.4% after another 21 days. Whereas in the verum group a distinct improvement in both API and SBI values could be stated, slight worsening of the values were determined for the placebo group. The results indicate that the oral application of green tea catechins and polyphenols might have a positive influence on the inflammatory reaction of periodontal structures. PMID- 11121367 TI - Endotoxin-reduced milk oligosaccharide fractions suitable for cell biological studies. AB - Endotoxins (ET) are able to activate leukocytes and other cell types and thus affect cell adhesion studies with regard to biological functions of human milk oligosaccharides (HMO). In our HMO preparations we detected ET in concentrations of 1.1 to 32.7 ng/mg. However, as we found in previous tests, an ET concentration of less than 100 pg/ml was necessary in order to avoid effects on the expression of CD11b and CD62L on human neutrophil granulocytes. Therefore, we used affinity chromatography with a detoxifying gel (polymixin B) which reduces the ET concentration of 414.9 +/- 121.5 (mean +/- SEM) and 47.9 +/- 5.9 pg/mg for the acidic and the neutral HMO, respectively. As only about 50 - 100 microg HMO per ml test solution are usually used in biological assays, the residual ET concentration does not interfere with cellular functions. We conclude that despite structural similarities between ET as lipopolysaccharides and complex HMO, a one step purification of HMO preparations was sufficient to get HMO material that can be used in cell culture systems. PMID- 11121368 TI - Evidence for neuronal dysfunction in migraine: concurrence between specific qEEG findings and clinical drug response--a retrospective analysis-. AB - OBJECTIVE: The aim of this analysis was to verify objective correlates of the clinical improvement of migraine in patients by means of quantitative topographical EEG (qEEG). METHODS: Overall 40 migraine-outpatients participated in this retrospective analysis of prophylactic migraine treatment over 3-8 months with 1600 to 2000 mg cyclandelate daily. In all patients qEEG was recorded and analysed using the Fast Fourier Transformation and the determination of the Abberation Index (AI = the statistical probability of belonging to a qEEG reference group of healthy people, n = 500). RESULTS: A clinical response (> 50% reduction of migraine attack frequency and duration) was observed in 77.5% (n = 31). The number and duration of migraine attacks per month (median values) were reduced in a highly significant manner. In this observation 75% of all patients investigated showed a pathological positive Aberration Index at a single electrode or a cluster of neighbouring brain sites. This consisted in a power increase in theta, alpha and/or beta1 activity, and 73.3% of them had also a corresponding negative AI-focus (power decrease). In 15 patients EEG aberrations (positive AI clusters) within one frequency band were found, whereas 15 patients showed aberrations in more than one frequency. Positive AI s were mainly found in fronto-temporal and occipital brain areas with one single topographical difference between the patients with and without aura. CONCLUSIONS: Positive AI values were the main electrophysiological findings in these migraine patients. This was supported by the highly significant decrease of the median amount of the positive AI after clinically successful treatment with cyclandelate. Twenty four out of thirty patients with positive AI foci showed concurrence between qEEG changes and clinical response. Patients with positive AI foci involving the lower or middle frequencies (theta and/or alpha 1) seemed to show a better clinical response (84.6%) than those with AI foci with participation of only faster frequencies (alpha 2 and/or beta 1; 54.5%). A controlled study is needed to confirm these observations. PMID- 11121369 TI - Interaction of the G protein beta 3 subunit T825 allele and the IRS-1 Arg972 variant in type 2 diabetes. AB - BACKGROUND: Type 2 diabetes mellitus is a common late-onset disease with a strong genetic component. It is characterized by insulin resistance which results from alterations in insulin signal transduction. The G protein beta 3 subunit 825T allele was recently found to be associated with hypertension and obesity which makes it a sensible candidate gene for type 2 diabetes. METHODS: In a case control study on 320 male patients and 962 male healthy controls we investigated the association of two candidate genes with diabetes, i.e. (i) the GNB3 825T allele, associated with a G protein beta 3 subunit splice variant and enhanced intracellular signal transduction, and (ii) the insulin receptor substrate-1 (IRS 1) 972Arg variant, which encodes a protein variant associated with cellular insulin resistance. RESULTS: The GNB3 825T allele and the IRS-1 972Arg variant were significantly associated with diabetes (odds ratios for either variant 1.4 1.8). Odds ratios were 3 4 in males carrying both alleles. CONCLUSIONS: The results document an association of a hypertension susceptibility gene with type 2 diabetes which may partially explain the frequent coexistence of both disorders. PMID- 11121370 TI - Dermatological infectiology--Quo vadis? Symposium, Ruhr-University, September 29 30, 2000. Abstracts. AB - Infectious diseases remain a major cause of morbidity and mortality in the year 2000. 17 million deaths per year or roughly a third of all deaths are caused by infections. Infectious diseases also pose a serious economic threat. While many well-established pathogens have not been contained several new infectious agents have been discovered within the past 27 years which include rotavirus, legionella, HIV, ebola, campylobacter, helicobacter, nipah, HHV8, hepatitis C, and many others. Additionally many new pathogens have emerged as serious threats to the ever-growing number of immuno-compromised patients. Infectious etiologies have been found for many common diseases (certain leukemias, duodenal ulcers, etcetera). It is likely that infections are at least co-factors for many other diseases (transplant-associated atherosclerosis). Only specialized care and multi disciplinary collaboration will enable us to cope with current problems and the inevitable emergence of new infectious diseases. PMID- 11121371 TI - Noninvasive measurement of hydrogen and potassium ion flux from single cells and epithelial structures. AB - This review introduces new developments in a technique for measuring the movement of ions across the plasma membrane. With the use of a self-referencing ion selective (Seris) probe, transport mechanisms can be studied on a variety of preparations ranging from tissues to single cells. In this paper we illustrate this versatility with examples from the vas deferens and inner ear epithelium to large and small single cells represented by mouse single-cell embryos and rat microglia. Potassium and hydrogen ion fluxes are studied and pharmacological manipulation of the signals are reported. The strengths of the self-referencing technique are reviewed with regard to biological applications, and the expansion of self-referencing probes to include electrochemical and enzyme-based sensors is discussed. PMID- 11121372 TI - Telokin expression is restricted to smooth muscle tissues during mouse development. AB - Telokin is a 17-kDa protein with an amino acid sequence that is identical to the COOH terminus of the 130-kDa myosin light chain kinase (MLCK). Telokin mRNA is transcribed from a second promoter, located within an intron, in the 3' region of the MLCK gene. In the current study, we show by in situ mRNA hybridization that telokin mRNA is restricted to the smooth muscle cell layers within adult smooth muscle tissues. In situ mRNA analysis of mouse embryos also revealed that telokin expression is restricted to smooth muscle tissues during embryonic development. Telokin mRNA expression was first detected in mouse gut at embryonic day 11.5; no telokin expression was detected in embryonic cardiac or skeletal muscle. Expression of telokin was also found to be regulated during postnatal development of the male and female reproductive tracts. In both uterus and vas deferens, telokin protein expression greatly increased between days 7 and 14 of postnatal development. The increase in telokin expression correlated with an increase in the expression of several other smooth muscle-restricted proteins, including smooth muscle myosin and alpha-actin. PMID- 11121373 TI - Heterogeneity of calcium stores and elementary release events in canine pulmonary arterial smooth muscle cells. AB - To examine the nature of inositol 1,4,5-trisphosphate (IP(3))-sensitive and ryanodine (Ryn)-sensitive Ca(2+) stores in isolated canine pulmonary arterial smooth cells (PASMC), agonist-induced changes in global intracellular Ca(2+) concentration ([Ca(2+)](i)) were measured using fura 2-AM fluorescence. Properties of elementary local Ca(2+) release events were characterized using fluo 3-AM or fluo 4-AM, in combination with confocal laser scanning microscopy. In PASMC, depletion of sarcoplasmic reticulum Ca(2+) stores with Ryn (300 microM) and caffeine (Caf; 10 mM) eliminated subsequent Caf-induced intracellular Ca(2+) transients but had little or no effect on the initial IP(3)-mediated intracellular Ca(2+) transient induced by ANG II (1 microM). Cyclopiazonic acid (CPA; 10 microM) abolished IP(3)-induced intracellular Ca(2+) transients but failed to attenuate the initial Caf-induced intracellular Ca(2+) transient. These results suggest that in canine PASMC, IP(3)-, and Ryn-sensitive Ca(2+) stores are organized into spatially distinct compartments while similar experiments in canine renal arterial smooth muscle cells (RASMC) reveal that these Ca(2+) stores are spatially conjoined. In PASMC, spontaneous local intracellular Ca(2+) transients sensitive to modulation by Caf and Ryn were detected, exhibiting spatial-temporal characteristics similar to those previously described for "Ca(2+) sparks" in cardiac and other types of smooth muscle cells. After depletion of Ryn-sensitive Ca(2+) stores, ANG II (8 nM) induced slow, sustained [Ca(2+)](i) increases originating at sites near the cell surface, which were abolished by depleting IP(3) stores. Discrete quantal-like events expected due to the coordinated opening of IP(3) receptor clusters ("Ca(2+) puffs") were not observed. These data provide new information regarding the functional properties and organization of intracellular Ca(2+) stores and elementary Ca(2+) release events in isolated PASMC. PMID- 11121374 TI - Gender-related distinctions in protein kinase C activity in rat vascular smooth muscle. AB - Gender differences in vascular reactivity have been suggested; however, the cellular mechanisms involved are unclear. We tested the hypothesis that the gender differences in vascular reactivity reflect gender-related, possibly estrogen-mediated, distinctions in the expression and activity of specific protein kinase C (PKC) isoforms in vascular smooth muscle. Aortic strips were isolated from intact and gonadectomized male and female Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). Isometric contraction was measured in endothelium-denuded aortic strips. PKC activity was measured in the cytosolic and particulate fractions, and the amount of PKC was measured using Western blots and isoform-specific anti-PKC antibodies. In intact male WKY rats, phenylephrine (Phe, 10(-5) M) and phorbol 12,13-dibutyrate (PDBu, 10(-6) M) stimulated contraction to 0.37 +/- 0.02 and 0.42 +/- 0.02 g/mg tissue wt, respectively. The basal particulate/cytosolic PKC activity ratio was 0.86 +/- 0.06, and Western blots revealed alpha-, delta-, and zeta-PKC isoforms. Phe and PDBu increased PKC activity and caused significant translocation of alpha- and delta-PKC from the cytosolic to particulate fraction. In intact female WKY rats, basal PKC activity, the amount of alpha-, delta-, and zeta-PKC, the Phe- and PDBu-induced contraction, and PKC activity and translocation of alpha- and delta-PKC were significantly reduced compared with intact male WKY rats. The basal PKC activity, the amount of alpha-, delta-, and zeta-PKC, the Phe and PDBu contraction, and PKC activity and alpha- and delta-PKC translocation were greater in SHR than WKY rats. The reduction in Phe and PDBu contraction and PKC activity in intact females compared with intact males was greater in SHR ( approximately 30%) than WKY rats ( approximately 20%). Phe and PDBu contraction and PKC activity were not significantly different between castrated males and intact males but were greater in ovariectomized (OVX) females than intact females. Treatment of OVX females or castrated males with 17 beta-estradiol, but not 17 alpha-estradiol, subcutaneous implants caused significant reduction in Phe and PDBu contraction and PKC activity that was greater in SHR than WKY rats. Phe and PDBu contraction and PKC activity in OVX females or castrated males treated with 17 beta-estradiol plus the estrogen receptor antagonist ICI-182,780 were not significantly different from untreated OVX females or castrated males. Thus a gender-related reduction in vascular smooth muscle contraction in female WKY rats with intact gonads compared with males is associated with reduction in the expression and activity of vascular alpha-, delta-, and zeta-PKC. The gender differences in vascular smooth muscle contraction and PKC activity are augmented in the SHR and are possibly mediated by estrogen. PMID- 11121375 TI - Desmin integrates the three-dimensional mechanical properties of muscles. AB - Striated muscle is a linear motor whose properties have been defined in terms of uniaxial structures. The question addressed here is what contribution is made to the properties of this motor by extramyofilament cytoskeletal structures that are not aligned in parallel with the myofilaments. This question arose from observations that transverse loads increase muscle force production in diaphragm but not in the hindlimb muscle, thereby indicating the presence of structures that couple longitudinal and transverse properties of diaphragmatic muscle. Furthermore, we find that the diaphragms of null mutants for the cytoskeletal protein desmin show 1) significant reductions in coupling between the longitudinal and transverse properties, indicating for the first time a role for a specific protein in integrating the three-dimensional mechanical properties of muscle, 2) significant reductions in the stiffness and viscoelasticity of muscle, and 3) significant increases in tetanic force production. Thus desmin serves a complex mechanical function in diaphragm muscle by contributing both to passive stiffness and viscoelasticity and to modulation of active force production in a three-dimensional structural network. Our finding changes the paradigm of force transmission among cells by placing our understanding of the function of the cytoskeleton in the context of the structural and mechanical complexity of muscles. PMID- 11121376 TI - Oxidative stress regulates collagen synthesis and matrix metalloproteinase activity in cardiac fibroblasts. AB - Oxidative stress has been implicated in the pathophysiology of myocardial failure. We tested the hypothesis that oxidative stress can regulate extracellular matrix in cardiac fibroblasts. Neonatal and adult rat cardiac fibroblasts in vitro were exposed to H(2)O(2) (0.05-5 microM) or the superoxide generating system xanthine (500 microM) plus xanthine oxidase (0.001-0.1 mU/ml) (XXO) for 24 h. In-gel zymography demonstrated that H(2)O(2) and XXO each increased gelatinase activity corresponding to matrix metalloproteinases (MMP) MMP-13, MMP-2, and MMP-9. H(2)O(2) and XXO decreased collagen synthesis (collagenase-sensitive [(3)H]proline incorporation) without affecting total protein synthesis ([(3)H]leucine incorporation). H(2)O(2) and XXO decreased the expression of procollagen alpha(1)(I), alpha(2)(I), and alpha(1)(III) mRNA but increased the expression of fibronectin mRNA, suggesting a selective transcriptional effect on collagen synthesis. H(2)O(2), but not XXO, also decreased the expression of nonfibrillar procollagen alpha(1)(IV) and alpha(2)(IV) mRNA. To determine the role of endogenous antioxidant systems, cells were treated with the superoxide dismutase (SOD) inhibitor diethyldithiocarbamic acid (DDC, 100 microM) to increase intracellular superoxide or with the glucose-6 phosphate dehydrogenase inhibitor dehydroisoandrosterone 3-acetate (DHEA; 10 microM) to increase intracellular H(2)O(2). DDC and DHEA decreased collagen synthesis and increased MMP activity, and both effects were inhibited by an SOD/catalase mimetic. Thus increased oxidative stress activates MMPs and decreases fibrillar collagen synthesis in cardiac fibroblasts. Oxidative stress may play a role in the pathogenesis of myocardial remodeling by regulating the quantity and quality of extracellular matrix. PMID- 11121377 TI - Chromaffin-adrenocortical cell interactions: effects of chromaffin cell activation in adrenal cell cocultures. AB - Although the adrenal cortex and medulla are both involved in the maintenance of homeostasis and stress response, the functional importance of intra-adrenal interactions remains unclear. When primary cocultures of frog (Rana pipiens) adrenocortical and chromaffin cells were used, selective chromaffin cell activation dramatically affected both chromaffin and adrenocortical cells. Depolarization with 50 microm veratridine enhanced chromaffin cell neuronal phenotype, contacts with adrenocortical cells, and secretion of norepinephrine, epinephrine, and serotonin. Time-lapse video microscopy recorded the rapid establishment of growth cones on the activated chromaffin cell neurites, neurite branching, and outgrowth toward adrenocortical cells. Simultaneously, adrenocortical cells migrated toward chromaffin cells. Following chromaffin cell activation, adrenocortical cell Fos protein expression and corticosteroid secretion were increased, indicating that chromaffin cell modulation of adrenocortical cells is at the transcriptional level. These results provide evidence that intra-adrenal interactions affect cellular differentiation and modulate steroidogenesis. Furthermore, this suggests that the activity-related plasticity of chromaffin and adrenocortical cells is developmentally and physiologically important. PMID- 11121378 TI - Osteoblast expression of vascular endothelial growth factor is modulated by the extracellular microenvironment. AB - Angiogenesis, the formation of new blood vessels, is crucial to the process of fracture healing. Vascular disruption after osseous injury results in an acidic, hypoxic wound environment. We have previously shown that osteoblasts can produce vascular endothelial growth factor (VEGF) in response to a variety of stimuli. In this study we examined pH and lactate concentration, two components of the putative fracture extracellular microenvironment, and determined their relative contribution to regulation of rat calvarial osteoblast VEGF production under both normoxic and hypoxic conditions. Our results demonstrate that pH and lactate concentration do independently affect osteoblast VEGF mRNA and protein production. Acidic pH (7.0) significantly decreased VEGF production, under normoxic and hypoxic conditions (P < 0.05), compared with neutral pH (7.4). This decrease was primarily transcriptionally regulated, because the rate of VEGF mRNA degradation was unchanged at pH 7.0 vs. 7.4. Similarly, an elevated lactate concentration (22 mM) also depressed osteoblast elaboration of VEGF at both neutral and acidic pH (P < 0.001). Furthermore, the effects of increasing acidity and elevated lactate appeared to be additive. PMID- 11121380 TI - A role for PYK2 in regulation of ERK1/2 MAP kinases and PI 3-kinase by ANG II in vascular smooth muscle. AB - Abnormal vascular smooth muscle cell (VSMC) growth plays a key role in the pathogenesis of hypertension and atherosclerosis. Angiotensin II (ANG II) elicits a hypertrophic growth response characterized by an increase in protein synthesis without cell proliferation. The present study investigated the role of the nonreceptor tyrosine kinase PYK2 in the regulation of ANG II-induced signaling pathways that mediate VSMC growth. Using coimmunoprecipitation analysis, the role of PYK2 as an upstream regulator of both extracellular signal-related kinase (ERK) 1/2 mitogen-activated protein kinase and phosphatidylinositol 3-kinase (PI 3-kinase) pathways was examined in cultured rat aortic VSMC. ANG II (100 nM) promoted the formation of a complex between PYK2 and the ERK1/2 regulators Shc and Grb2. ANG II caused a rapid and Ca(2+)-dependent tyrosine phosphorylation of the adapter molecule p130Cas, which coimmunoprecipitated both PYK2 and PI 3 kinase in ANG II-treated VSMC. Complex formation between PI 3-kinase and p130Cas and PYK2 was associated with a rapid phosphorylation of the ribosomal p70(S6) kinase in a Ca(2+)- and tyrosine kinase-dependent manner. These data suggest that PYK2 is an important regulator of multiple signaling pathways involved in ANG II induced VSMC growth. PMID- 11121379 TI - Modulation of P2Z/P2X(7) receptor activity in macrophages infected with Chlamydia psittaci. AB - Given the role that extracellular ATP (ATP(o))-mediated apoptosis may play in inflammatory responses and in controlling mycobacterial growth in macrophages, we investigated whether ATP(o) has any effect on the viability of chlamydiae in macrophages and, conversely, whether the infection has any effect on susceptibility to ATP(o)-induced killing via P2Z/P2X(7) purinergic receptors. Apoptosis of J774 macrophages could be selectively triggered by ATP(o), because other purine/pyrimidine nucleotides were ineffective, and it was inhibited by oxidized ATP, which irreversibly inhibits P2Z/P2X(7) purinergic receptors. Incubation with ATP(o) but not other extracellular nucleotides inhibits the growth of intracellular chlamydiae, consistent with previous observations on ATP(o) effects on growth of intracellular mycobacteria. However, chlamydial infection for 1 day also inhibits ATP(o)-mediated apoptosis, which may be a mechanism to partially protect infected cells against the immune response. Infection by Chlamydia appears to protect cells by decreasing the ability of ATP(o) to permeabilize macrophages to small molecules and by abrogating a sustained Ca(2+) influx previously associated with ATP(o)-induced apoptosis. PMID- 11121381 TI - Role of calcium and calmodulin in ciliary stimulation induced by acetylcholine. AB - The goal of this work was to elucidate the molecular events underlying stimulation of ciliary beat frequency (CBF) induced by acetylcholine (ACh) in frog esophagus epithelium. ACh induces a profound increase in CBF and in intracellular Ca(2+) concentration ([Ca(2+)](i)) through M(1) and M(3) muscarinic receptors. The [Ca(2+)](i) slowly decays to the basal level, while CBF stabilizes at an elevated level. These results suggest that ACh triggers Ca(2+)-correlated and -uncorrelated modes of ciliary stimulation. ACh response is abolished by the phospholipase C (PLC) inhibitor U-73122 and by depletion of intracellular Ca(2+) stores but is unaffected by reduction of extracellular Ca(2+) concentration and by blockers of Ca(2+) influx. Therefore, ACh activates PLC and mobilizes Ca(2+) solely from intracellular stores. The calmodulin inhibitors W-7 and calmidazolium attenuate the ACh-induced increase in [Ca(2+)](i) but completely abolish the elevation in CBF. Therefore, elevation of [Ca(2+)](i) is necessary for CBF enhancement but does not lead directly to it. The combined effect of Ca(2+) elevation and of additional factors, presumably mobilized by Ca(2+)-calmodulin, results in a robust CBF enhancement. PMID- 11121382 TI - Activation of MT(2) melatonin receptors in rat suprachiasmatic nucleus phase advances the circadian clock. AB - The aim of this study was to identify the melatonin receptor type(s) (MT(1) or MT(2)) mediating circadian clock resetting by melatonin in the mammalian suprachiasmatic nucleus (SCN). Quantitative receptor autoradiography with 2 [(125)I]iodomelatonin and in situ hybridization histochemistry, with either (33)P or digoxigenin-labeled antisense MT(1) and MT(2) melatonin receptor mRNA oligonucleotide probes, revealed specific expression of both melatonin receptor types in the SCN of inbred Long-Evans rats. The melatonin receptor type mediating phase advances of the circadian rhythm of neuronal firing rate in the SCN slice was assessed using competitive melatonin receptor antagonists, the MT(1)/MT(2) nonselective luzindole and the MT(2)-selective 4-phenyl-2-propionamidotetraline (4P-PDOT). Luzindole and 4P-PDOT (1 nM-1 microM) did not affect circadian phase on their own; however, they blocked both the phase advances (approximately 4 h) in the neuronal firing rate induced by melatonin (3 pM) at temporally distinct times of day [i.e., subjective dusk, circadian time (CT) 10; and dawn, CT 23], as well as the associated increases in protein kinase C activity. We conclude that melatonin mediates phase advances of the SCN circadian clock at both dusk and dawn via activation of MT(2) melatonin receptor signaling. PMID- 11121383 TI - Effects of extracellular calcium and potassium on the sodium pump of rat adrenal glomerulosa cells. AB - Because the activity of the sodium pump (Na-K-ATPase) influences the secretion of aldosterone, we determined how extracellular potassium (K(o)) and calcium affect sodium pump activity in rat adrenal glomerulosa cells. Sodium pump activity was measured as ouabain-sensitive (86)Rb uptake in freshly dispersed cells containing 20 mM sodium as measured with sodium-binding benzofluran isophthalate. Increasing K(o) from 4 to 10 mM in the presence of 1.8 mM extracellular calcium (Ca(o)) stimulated sodium pump activity up to 165% and increased intracellular free calcium as measured with fura 2. Increasing K(o) from 4 to 10 mM in the absence of Ca(o) stimulated the sodium pump approximately 30% and did not increase intracellular free calcium concentration ([Ca(2+)](i)). In some experiments, addition of 1.8 mM Ca(o) in the presence of 4 mM K(o) increased [Ca(2+)](i) above the levels observed in the absence of Ca(o) and stimulated the sodium pump up to 100%. Ca-dependent stimulation of the sodium pump by K(o) and Ca(o) was inhibited by isradipine (10 microM), a blocker of L- and T-type calcium channels, by compound 48/80 (40 microg/ml) and calmidizolium (10 microM), which inhibits calmodulin (CaM), and by KN-62 (10 microM), which blocks some forms of Ca/CaM kinase II (CaMKII). Staurosporine (1 microM), which effectively blocks most forms of protein kinase C, had no effect. In the presence of A-23187, a calcium ionophore, the addition of 0.1 mM Ca(o) increased [Ca(2+)](i) to the level observed in the presence of 10 mM K(o) and 1.8 mM Ca(o) and stimulated the sodium pump 100%. Ca-dependent stimulation by A-23187 and 0.1 mM Ca(o) was not reduced by isradipine but was blocked by KN-62. Thus, under the conditions that K(o) stimulates aldosterone secretion, it stimulates the sodium pump by two mechanisms: direct binding to the pump and by increasing calcium influx, which is dependent on Ca(o). The resulting increase in [Ca(2+)](i) may stimulate the sodium pump by activating CaM and/or CaMKII. PMID- 11121385 TI - Improved oxygenation promotes CFTR maturation and trafficking in MDCK monolayers. AB - Culturing airway epithelial cells with most of the apical media removed (air liquid interface) has been shown to enhance cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl(-) secretory current. Thus we hypothesized that cellular oxygenation may modulate CFTR expression. We tested this notion using type I Madin-Darby canine kidney cells that endogenously express low levels of CFTR. Growing monolayers of these cells for 4 to 5 days with an air-liquid interface caused a 50-fold increase in forskolin-stimulated Cl(-) current, compared with conventional (submerged) controls. Assaying for possible changes in CFTR by immunoprecipitation and immunocytochemical localization revealed that CFTR appeared as an immature 140-kDa form intracellularly in conventional cultures. In contrast, monolayers grown with an air-liquid interface possessed more CFTR protein, accompanied by increases toward the mature 170-kDa form and apical membrane staining. Culturing submerged monolayers with 95% O(2) produced similar improvements in Cl(-) current and CFTR protein as air-liquid interface culture, while increasing PO(2) from 2.5% to 20% in air-liquid interface cultures yielded graded enhancements. Together, our data indicate that improved cellular oxygenation can increase endogenous CFTR maturation and/or trafficking. PMID- 11121384 TI - Defective dietary fat processing in transgenic mice lacking aquaporin-1 water channels. AB - Immunocytochemistry showed expression of aquaporin-1 (AQP1) water channels at sites involved in dietary fat processing, including intrahepatic cholangiocytes, gallbladder, pancreatic microvascular endothelium, and intestinal lacteals. To determine whether AQP1 has a role in dietary fat digestion and/or absorption, mice were placed on a diet that contained 50% fat. Whereas wild-type mice (3-3.5 wk of age, 10-12 g) gained 49 +/- 5% (SE, n = 50) body weight in 8 days, and heterozygous mice gained 46 +/- 4%, AQP1 null mice gained only 4 +/- 3%; weights became similar after return to a 6% fat diet after 6 days. The null mice on a high-fat diet acquired an oily appearance, developed steatorrhea with increased stool triglyceride content, and manifested serum hypotriglyceridemia. Supplementation of the high-fat diet with pancreatic enzymes partially corrected the decreased weight gain in null mice. Absorption of [(14)C]oleic acid from small intestine was not affected by AQP1 deletion, as determined by blood radioactivity after duodenal infusion. Lipase activity in feces and small intestine was remarkably greater in AQP1 null than wild-type mice on low- and high-fat diets. Fluid collections done in older mice (that are less sensitive to a high-fat diet) by ductal cannulation showed threefold increased pancreatic fluid flow in response to secretin/cholecystokinin, but volumes, pH, and amylase activities were affected little by AQP1 deletion, nor were bile flow rates and bile salt concentrations. Together, these results establish a dietary fat misprocessing defect in AQP1 null mice. PMID- 11121386 TI - Expression of the Na(+)/Ca(2+) exchanger in skeletal muscle. AB - The expression of the Na(+)/Ca(2+) exchanger was studied in differentiating muscle fibers in rats. NCX1 and NCX3 isoform (Na(+)/Ca(2+) exchanger isoform) expression was found to be developmentally regulated. NCX1 mRNA and protein levels peaked shortly after birth. Conversely, NCX3 isoform expression was very low in muscles of newborn rats but increased dramatically during the first 2 wk of postnatal life. Immunocytochemical analysis showed that NCX1 was uniformly distributed along the sarcolemmal membrane of undifferentiated rat muscle fibers but formed clusters in T-tubular membranes and sarcolemma of adult muscle. NCX3 appeared to be more uniformly distributed along the sarcolemma and inside myoplasm. In the adult, NCX1 was predominantly expressed in oxidative (type 1 and 2A) fibers of both slow- and fast-twitch muscles, whereas NCX3 was highly expressed in fast glycolytic (2B) fibers. NCX2 was expressed in rat brain but not in skeletal muscle. Developmental changes in NCX1 and NCX3 as well as the distribution of these isoforms at the cellular level and in different fiber types suggest that they may have different physiological roles. PMID- 11121387 TI - A proposed mechanism for the potentiation of cAMP-mediated acid secretion by carbachol. AB - Acid secretion in isolated rabbit gastric glands was monitored by the accumulation of [(14)C]aminopyrine. Stimulation of the glands with carbachol synergistically augmented the response to dibutyryl cAMP. The augmentation persisted even after carbachol was washed out and was resistant to chelated extracellular Ca(2+) and to inhibitors of either protein kinase C or calmodulin kinase II. Cytochalasin D at 10 microM preferentially blocked the secretory effect of carbachol and its synergism with cAMP, whereas it had no effect on histamine- or cAMP-stimulated acid secretion within 15 min. Cytochalasin D inhibited the carbachol-stimulated intracellular Ca(2+) concentration ([Ca(2+)](i)) increase due to release from the Ca(2+) store. Treatment of the glands with cytochalasin D redistributed type 3 inositol 1,4,5-trisphosphate receptor (the major subtype in the parietal cell) from the fraction containing membranes of large size to the microsomal fraction, suggesting a dissociation of the store from the plasma membrane. These findings suggest that intracellular Ca(2+) release by cholinergic stimulation is critical for determining synergism with cAMP in parietal cell activation and that functional coupling between the Ca(2+) store and the receptor is maintained by actin microfilaments. PMID- 11121388 TI - The Delta F508 mutation shortens the biochemical half-life of plasma membrane CFTR in polarized epithelial cells. AB - Although the biosynthetic arrest of the DeltaF508 mutant of cystic fibrosis transmembrane conductance regulator (CFTR) can be partially reversed by physical and chemical means, recent evidence suggests that the functional stability of the mutant protein after reaching the cell surface is compromised. To understand the molecular basis for this observation, the current study directly measured the half-life of Delta F508 and wild-type CFTR at the cell surface of transfected LLC PK(1) cells. Plasma membrane CFTR expression over time was characterized biochemically and functionally in these polarized epithelial cells. Surface biotinylation, streptavidin extraction, and quantitative immunoblot analysis determined the biochemical half-life of plasma membrane DeltaF508 CFTR to be approximately 4 h, whereas the plasma membrane half-life of wild-type CFTR exceeded 48 h. This difference in biochemical stability correlated with CFTR mediated transport function. These findings indicate that the Delta F508 mutation decreases the biochemical stability of CFTR at the cell surface. We conclude that the Delta F508 mutation triggers more rapid internalization of CFTR and/or its preferential sorting to a pathway of rapid degradation. PMID- 11121389 TI - Properties of voltage-gated Ca(2+) channels in rabbit ventricular myocytes expressing Ca(2+) channel alpha(1E) cDNA. AB - Using the whole-cell patch-clamp technique, we have studied the properties of alpha(1E) Ca(2+) channel transfected in cardiac myocytes. We have also investigated the effect of foreign gene expression on the intrinsic L-type current (I(Ca,L)). Expression of green fluorescent protein significantly decreased the I(Ca,L). By contrast, expression of alpha(1E) with beta(2b) and alpha(2)/delta significantly increased the total Ca(2+) current, and in these cells a Ca(2+) antagonist, PN-200-110 (PN), only partially blocked the current. The remaining PN-resistant current was abolished by the application of a low concentration of Ni(2+) and was little affected by changing the charge carrier from Ca(2+) to Ba(2+) or by beta-adrenergic stimulation. On the basis of its voltage range for activation, this channel was classified as a high-voltage activated channel. Thus the expression of alpha(1E) did not generate T-like current in cardiac myocytes. On the other hand, expression of alpha(1E) decreased I(Ca,L) and slowed the I(Ca,L) inactivation. This inactivation slowing was attenuated by the beta(2b) coexpression, suggesting that the alpha(1E) may slow the inactivation of I(Ca,L) by scrambling with alpha(1C) for intrinsic auxiliary beta. PMID- 11121390 TI - MAP kinase upregulation after hematopoietic differentiation: role of chemotaxis. AB - Mitogen-activated protein kinase (MAPK) isoform p42 is known to be active in exponentially growing cells at several points of the cell cycle. A high basal activity was present in three cell lines representative of immature myeloid cells tested: uHL-60, AML-14, and MPD. However, DMSO-induced differentiation of HL-60 cells (dHL-60) and subsequent expression of the neutrophilic phenotype occurred with a concomitant reduction on the basal level of MAPK activity. Simultaneously, extracellular stimuli like the cytokine granulocyte/macrophage colony-stimulating factor (GM-CSF) induced a fast (<10 min) and robust response. In terms of MAPK activity, the more mature the cell was, the higher the corresponding activity, in the three differentiation series considered: AML-14 < 3D10; MPD < G-MPD; uHL-60 < dHL-60 < neutrophils. Interestingly, peripheral blood neutrophils expressed the highest (16-fold) MAPK activation level in response to GM-CSF. Finally, using the specific MAPK inhibitor PD-98059, we demonstrated that MAPK activation is needed for neutrophil chemotaxis toward interleukin-8 and its priming by GM-CSF. Since neutrophils are terminally differentiated cells, GM-CSF does not serve a purpose in proliferation, and it must trigger the recruitment of selective signal transduction pathways particular to that final stage that includes enhanced physiological functions such as chemotaxis. PMID- 11121391 TI - Differential renal distribution of NHERF isoforms and their colocalization with NHE3, ezrin, and ROMK. AB - Na(+)/H(+) exchanger regulatory factor (NHERF) and NHERF2 are PDZ motif proteins that mediate the inhibitory effect of cAMP on Na(+)/H(+) exchanger 3 (NHE3) by facilitating the formation of a multiprotein signaling complex. With the use of antibodies specific for NHERF and NHERF2, immunocytochemical analysis of rat kidney was undertaken to determine the nephron distribution of both proteins and their colocalization with other transporters and with ezrin. NHERF was most abundant in apical membrane of proximal tubule cells, where it colocalized with ezrin and NHE3. NHERF2 was detected in the glomerulus and in other renal vascular structures. In addition, NHERF2 was strongly expressed in collecting duct principal cells, where it colocalized with ROMK. These results indicate a striking difference in the nephron distribution of NHERF and NHERF2 and suggests NHERF is most likely to be the relevant biological regulator of NHE3 in the proximal tubule, while NHERF2 may interact with ROMK or other targets in the collecting duct. The finding that NHERF isoforms occur in different cell types suggests that NHERF and NHERF2 may subserve different functions in the kidney. PMID- 11121392 TI - Properties of ATP-dependent K(+) channels in adrenocortical cells. AB - Bovine adrenocortical zona fasciculata (AZF) cells express a novel ATP-dependent K(+)-permeable channel (I(AC)). Whole cell and single-channel recordings were used to characterize I(AC) channels with respect to ionic selectivity, conductance, and modulation by nucleotides, inorganic phosphates, and angiotensin II (ANG II). In outside-out patch recordings, the activity of unitary I(AC) channels is enhanced by ATP in the patch pipette. These channels were K(+) selective with no measurable Na(+) or Ca(2+) conductance. In symmetrical K(+) solutions with physiological concentrations of divalent cations (M(2+)), I(AC) channels were outwardly rectifying with outward and inward chord conductances of 94.5 and 27.0 pS, respectively. In the absence of M(2+), conductance was nearly ohmic. Hydrolysis-resistant nucleotides including AMP-PNP and NaUTP were more potent than MgATP as activators of whole cell I(AC) currents. Inorganic polytriphosphate (PPP(i)) dramatically enhanced I(AC) activity. In current-clamp recordings, nucleotides and PPP(i) produced resting potentials in AZF cells that correlated with their effectiveness in activating I(AC). ANG II (10 nM) inhibited whole cell I(AC) currents when patch pipettes contained 5 mM MgATP but was ineffective in the presence of 5 mM NaUTP and 1 mM MgATP. Inhibition by ANG II was not reduced by selective kinase antagonists. These results demonstrate that I(AC) is a distinctive K(+)-selective channel whose activity is increased by nucleotide triphosphates and PPP(i). Furthermore, they suggest a model for I(AC) gating that is controlled through a cycle of ATP binding and hydrolysis. PMID- 11121393 TI - Flow-induced expression of endothelial Na-K-Cl cotransport: dependence on K(+) and Cl(-) channels. AB - Steady laminar shear stress has been shown previously to markedly increase Na-K Cl cotransporter mRNA and protein in human umbilical vein endothelial cells and also to rapidly increase endothelial K(+) and Cl(-) channel conductances. The present study was done to evaluate the effects of shear stress on Na-K-Cl cotransporter activity and protein expression in bovine aortic endothelial cells (BAEC) and to determine whether changes in cotransporter expression may be dependent on early changes in K(+) and Cl(-) channel conductances. Confluent BAEC monolayers were exposed in a parallel-plate flow chamber to either steady shear stress (19 dyn/cm(2)) or purely oscillatory shear stress (0 +/- 19 dyn/cm(2)) for 6-48 h. After shearing, BAEC monolayers were assessed for Na-K-Cl cotransporter activity or were subjected to Western blot analysis of cotransporter protein. Steady shear stress led to a 2- to 4-fold increase in BAEC cotransporter protein levels and a 1.5- to 1.8-fold increase in cotransporter activity, increases that were sustained over the longest time periods studied. Oscillatory flow, in contrast, had no effect on cotransporter protein levels. In the presence of flow sensitive K(+) and Cl(-) channel pharmacological blockers, the steady shear stress-induced increase in cotransporter protein was virtually abolished. These results suggest that shear stress modulates the expression of the BAEC Na-K-Cl cotransporter by mechanisms that are dependent on flow-activated ion channels. PMID- 11121394 TI - Generation and phenotype of cell lines derived from CF and non-CF mice that carry the H-2K(b)-tsA58 transgene. AB - Tracheal, renal, salivary, and pancreatic epithelial cells from cystic fibrosis [CF; cystic fibrosis transmembrane conductance regulator (CFTR) -/-] and non-CF mice that carry a temperature-sensitive SV40 large T antigen oncogene (ImmortoMouse) were isolated and maintained in culture under permissive conditions (33 degrees C with interferon-gamma). The resultant cell lines have been in culture for >1 year and 50 passages. Each of the eight cell lines form polarized epithelial barriers and exhibit regulated, electrogenic ion transport. The four non-CF cell lines (mTEC1, mCT1, mSEC1, and mPEC1) express cAMP-regulated Cl(-) permeability and cAMP-stimulated Cl(-) secretion. In contrast, the four CFTR -/- cell lines (mTEC1-CF, mCT1-CF, mSEC1-CF, and mPEC1-CF) each lack cAMP stimulated Cl(-) secretory responses. Ca(2+)-activated Cl(-) secretion is retained in both CF and non-CF cell lines. Thus we have generated genetically well-matched epithelial cell lines from several tissues relevant to cystic fibrosis that either completely lack CFTR or express endogenous levels of CFTR. These cell lines should prove useful for studies of regulation of epithelial cell function and the role of CFTR in cell physiology. PMID- 11121395 TI - Inhibition of JNK by overexpression of the JNL binding domain of JIP-1 prevents apoptosis in sympathetic neurons. AB - Studies in non-neuronal cells show that c-Jun N-terminal kinases (JNK) play a key role in apoptotic cell death. In some neurons JNK is also thought to initiate cell death by the activation of c-Jun. JNK inhibition has been achieved pharmacologically by inhibiting upstream kinases, but there has been no direct demonstration that inhibition of JNK can prevent neuronal death. We have therefore examined whether the JNK binding domain (JBD) of JNK-interacting protein-1 (JIP-1, a scaffold protein and specific inhibitor of JNK) can inhibit c Jun phosphorylation and support the survival of sympathetic neurons deprived of NGF. We show that expression of the JBD in >80% of neurons was sufficient to prevent the phosphorylation of c-Jun and its nuclear accumulation as well as abrogate neuronal cell death induced by NGF deprivation. JBD expression also preserved the capacity of mitochondria to reduce MTT. Interestingly, although the PTB domain of JIP was reported to interact with rhoGEF, expression of the JBD domain was sufficient to localize the protein to the membrane cortex and growth cones. Hence, JNK activation is a key event in apoptotic death induced by NGF withdrawal, where its point of action lies upstream of mitochondrial dysfunction. PMID- 11121396 TI - Isoprenylcysteine carboxyl methyltransferase deficiency in mice. AB - After isoprenylation, Ras and other CAAX proteins undergo endoproteolytic processing by Rce1 and methylation of the isoprenylcysteine by Icmt (isoprenylcysteine carboxyl methyltransferase). We reported previously that Rce1 deficient mice died during late gestation or soon after birth. We hypothesized that Icmt deficiency might cause a milder phenotype, in part because of reports suggesting the existence of more than one activity for methylating isoprenylated proteins. To address this hypothesis and also to address the issue of other methyltransferase activities, we generated Icmt-deficient mice. Contrary to our expectation, Icmt deficiency caused a more severe phenotype than Rce1 deficiency, with virtually all of the knockout embryos (Icmt-/-) dying by mid-gestation. An analysis of chimeric mice produced from Icmt-/- embryonic stem cells showed that the Icmt-/- cells retained the capacity to contribute to some tissues (e.g. skeletal muscle) but not to others (e.g. brain). Lysates from Icmt-/- embryos lacked the ability to methylate either recombinant K-Ras or small molecule substrates (e.g. N-acetyl-S-geranylgeranyl-l-cysteine). In addition, Icmt-/- cells lacked the ability to methylate Rab proteins. Thus, Icmt appears to be the only enzyme participating in the carboxyl methylation of isoprenylated proteins. PMID- 11121397 TI - rib-2, a Caenorhabditis elegans homolog of the human tumor suppressor EXT genes encodes a novel alpha1,4-N-acetylglucosaminyltransferase involved in the biosynthetic initiation and elongation of heparan sulfate. AB - The proteins encoded by the EXT1, EXT2, and EXTL2 genes, members of the hereditary multiple exostoses gene family of tumor suppressors, are glycosyltransferases required for the heparan sulfate biosynthesis. Only two homologous genes, rib-1 and rib-2, of the mammalian EXT genes were identified in the Caenorhabditis elegans genome. Although heparan sulfate is found in C. elegans, the involvement of the rib-1 and rib-2 proteins in heparan sulfate biosynthesis remains unclear. In the present study, the substrate specificity of a soluble recombinant form of the rib-2 protein was determined and compared with those of the recombinant forms of the mammalian EXT1, EXT2, and EXTL2 proteins. The present findings revealed that the rib-2 protein was a unique alpha1,4-N acetylglucosaminyltransferase involved in the biosynthetic initiation and elongation of heparan sulfate. In contrast, the findings confirmed the previous observations that both the EXT1 and EXT2 proteins were heparan sulfate copolymerases with both alpha1,4-N-acetylglucosaminyltransferase and beta1,4 glucuronyltransferase activities, which are involved only in the elongation step of the heparan sulfate chain, and that the EXTL2 protein was an alpha1,4-N acetylglucosaminyltransferase involved only in the initiation of heparan sulfate synthesis. These findings suggest that the biosynthetic mechanism of heparan sulfate in C. elegans is distinct from that reported for the mammalian system. PMID- 11121398 TI - Epilysin, a novel human matrix metalloproteinase (MMP-28) expressed in testis and keratinocytes and in response to injury. AB - We have cloned a new human matrix metalloproteinase (MMP-28, epilysin) from human keratinocyte and testis cDNA libraries. Like most MMPs, epilysin contains a signal sequence, a prodomain with a PRCGVTD sequence, a zinc-binding catalytic domain with an HEIGHTLGLTH sequence, and a hemopexin-like domain. In addition, epilysin has a furin activation sequence (RRKKR) but has no transmembrane sequence. The exon-intron organization and splicing pattern of epilysin differ from that of other MMP genes. It has only 8 exons, and 5 exons are spliced at sites not used by other MMPs. Another novel feature of epilysin is that exon 4 is alternatively spliced to a transcript that does not encode the N-terminal half of the catalytic domain. Northern hybridization of tissue RNA indicated that epilysin is expressed at high levels in testis and at lower levels in lungs, heart, colon, intestine, and brain. RNase protection assay with various cell lines indicated that epilysin was selectively expressed in keratinocytes. Recombinant epilysin degraded casein in a zymography assay, and its proteolytic activity was inhibited by EDTA and by batimastat, a selective MMP inhibitor. Immunohistochemical staining showed expression of epilysin protein in the basal and suprabasal epidermis of intact skin. In injured skin, prominent staining for epilysin was seen in basal keratinocytes both at and some distance from the wound edge, a pattern that is quite distinct from that of other MMPs expressed during tissue repair. These findings suggest that this new MMP functions in several tissues both in tissue homeostasis and in repair. PMID- 11121399 TI - Topogenesis of peroxisomal membrane protein requires a short, positively charged intervening-loop sequence and flanking hydrophobic segments. study using human membrane protein PMP34. AB - Human 34-kDa peroxisomal membrane protein (PMP34) consisting of 307 amino acids was previously identified as an ortholog of, or a similar protein (with 27% identity) to the, 423-amino acid-long PMP47 of the yeast Candida boidinii. We investigated membrane topogenesis of PMP34 with six putative transmembrane segments, as a model peroxisomal membrane protein. PMP34 was characterized as an integral membrane protein of peroxisomes. Transmembrane topology of PMP34 was determined by differential permeabilization and immunofluorescent staining of HeLa cells ectopically expressing PMP34 as well as of Chinese hamster ovary-K1 expressing epitope-tagged PMP34. As opposed to PMP47, PMP34 was found to expose its N- and C-terminal parts to the cytosol. Various deletion variants of PMP34 and their fusion proteins with green fluorescent protein were expressed in Chinese hamster ovary-K1 and were verified with respect to intracellular localization. The loop region between transmembrane segments 4 and 5 was required for the peroxisome-targeting activity, in which Ala substitution for basic residues abrogated the activity. Three hydrophobic transmembrane segments linked in a flanking region of the basic loop were essential for integration of PMP34 to peroxisome membranes. Therefore, it is evident that the intervening basic loop plus three transmembrane segments of PMP34 function as a peroxisomal targeting and topogenic signal. PMID- 11121400 TI - Substrate recognition by the collagen-binding domain of Clostridium histolyticum class I collagenase. AB - Clostridium histolyticum type I collagenase (ColG) has a segmental structure, S1+S2+S3a+S3b. S3a and S3b bound to insoluble collagen, but S2 did not, thus indicating that S3 forms a collagen-binding domain (CBD). Because S3a+S3b showed the most efficient binding to substrate, cooperative binding by both domains was suggested for the enzyme. Monomeric (S3b) and tandem (S3a+S3b) CBDs bound to atelocollagen, which contains only the collagenous region. However, they did not bind to telopeptides immobilized on Sepharose beads. These results suggested that the binding site(s) for the CBD is(are) present in the collagenous region. The CBD bound to immobilized collagenous peptides, (Pro-Hyp-Gly)(n) and (Pro-Pro Gly)(n), only when n is large enough to allow the peptides to have a triple helical conformation. They did not bind to various peptides with similar amino acid sequences or to gelatin, which lacks a triple-helical conformation. The CBD did not bind to immobilized Glc-Gal disaccharide, which is attached to the side chains of hydroxylysine residues in the collagenous region. These observations suggested that the CBD specifically recognizes the triple-helical conformation made by three polypeptide chains in the collagenous region. PMID- 11121401 TI - Equilibrium binding of single-stranded DNA to the secondary DNA binding site of the bacterial recombinase RecA. AB - The bacterial recombinase RecA forms a nucleoprotein filament in vitro with single-stranded DNA (ssDNA) at its primary DNA binding site, site I. This filament has a second site, site II, which binds ssDNA and double-stranded DNA. We have investigated the binding of ssDNA to the RecA protein in the presence of adenosine 5'-O-(thiotriphosphate) cofactor using fluorescence anisotropy. The RecA protein carried out DNA strand exchange with a 5'-fluorescein-labeled 32-mer oligonucleotide. The anisotropy signal was shown to measure oligonucleotide binding to RecA, and the relationship between signal and binding density was determined. Binding of ssDNA to site I of RecA was stable at high NaCl concentrations. Binding to site II could be described by a simple two-state equilibrium, K = 4.5 +/- 1.5 x 10(5) m(-1) (37 degrees C, 150 mm NaCl, pH 7.4). The reaction was enthalpy-driven and entropy-opposed. It depended on salt concentration and was sensitive to the type of monovalent anion, suggesting that anion-dependent protein conformations contribute to ssDNA binding at site II. PMID- 11121402 TI - alpha 2 integrin subunit cytoplasmic domain-dependent cellular migration requires p38 MAPK. AB - The alpha(2) integrin subunit cytoplasmic domain uniquely supported epidermal growth factor (EGF)-stimulated migration on type I collagen. p38 MAP kinase- and phosphatidylinositol 3-kinase-specific inhibitors, but not a MEK-specific inhibitor, eliminated EGF-stimulated and unstimulated alpha(2)-cytoplasmic domain dependent migration. Following adhesion to collagenous matrices, cells expressing the full-length alpha(2) integrin subunit, but not cells expressing a chimeric alpha(2) integrin subunit in which the alpha(2)-cytoplasmic domain was replaced by the cytoplasmic domain of the alpha(1)-subunit, exhibited sustained and robust phosphorylation of p38 MAP kinase. Expression of dominant negative p38 MAP kinase inhibited alpha(2)-cytoplasmic domain-dependent, EGF-stimulated migration as well as unstimulated migration on collagen. Expression of constitutively active Rac1(Val-12) augmented p38 MAP kinase activation and alpha(2)-cytoplasmic domain dependent migration. It also rescued the ability of cells expressing the alpha(1) cytoplasmic domain to activate p38 MAPK and to migrate. These results suggest that the alpha(2) integrin cytoplasmic domain uniquely stimulates the p38 MAP kinase pathway that is required for unstimulated and EGF-stimulated migration on type I collagen. PMID- 11121403 TI - Dissection of the ATP-binding domain of the chaperone hsc70 for interaction with the cofactor Hap46. AB - Several unrelated proteins are known that specifically interact with members of the mammalian hsp70 chaperone protein family independent of the hsp70 substrate binding site. One of these is Hap46, also called BAG-1, which binds to the ATP binding domain of hsp70 and its constitutively expressed, highly homologous counterpart hsc70, thereby affecting nucleotide binding, as well as protein folding properties, of these molecular chaperones. In an attempt to delineate the potential contact sites on hsp70/hsc70 involved in this interaction we made use of the following two independent approaches: (i) screening of membrane-bound peptide libraries based on the sequence of the ATP-binding domain and (ii) the phage-display technique with random dodecapeptides. These approaches yielded partially overlapping results and identified several possible contact regions. On the space-filling model of hsc70, the two major contact areas for Hap46 delineated in the present study are located on the same side of the molecule on either subdomain that border the central cleft harboring the nucleotide-binding site. We suggest that this bridging affects the conformation of the ATP-binding domain in a way similar to the opening of the nucleotide-binding cleft produced in the bacterial hsp70 homologue DnaK upon binding its regulatory protein GrpE. PMID- 11121404 TI - Activation of anaplastic lymphoma kinase receptor tyrosine kinase induces neuronal differentiation through the mitogen-activated protein kinase pathway. AB - Anaplastic lymphoma kinase (ALK) is a novel neuronal orphan receptor tyrosine kinase that is essentially and transiently expressed in specific regions of the central and peripheral nervous systems, suggesting a role in its normal development and function. To determine whether ALK could play a role in neuronal differentiation, we established a model system that allowed us to mimic the normal activation of this receptor. We expressed, in PC12 cells, a chimeric protein in which the extracellular domain of the receptor was replaced by the mouse IgG 2b Fc domain. The Fc domain induced the dimerization and oligomerization of the chimeric protein leading to receptor phosphorylation and activation, thus mimicking the effect of ligand binding, whereas the wild type ALK remained as a monomeric nonphosphorylated protein. Expression of the chimera, but not that of the wild type ALK or of a kinase inactive form of the chimera, induced the differentiation of PC12 cells. Analysis of the signaling pathways involved in this process pointed to an essential role of the mitogen-activated protein kinase cascade. These results are consistent with a role for ALK in neuronal differentiation. PMID- 11121406 TI - Tissue transglutaminase facilitates the polymerization of insulin-like growth factor-binding protein-1 (IGFBP-1) and leads to loss of IGFBP-1's ability to inhibit insulin-like growth factor-I-stimulated protein synthesis. AB - Insulin-like growth factor-binding protein-1 (IGFBP-1) binds to insulin-like growth factors (IGFs) and has been shown to inhibit or stimulate cellular responses to IGF-I in vitro. This capacity of IGFBP-1 to inhibit or stimulate IGF I actions correlates with its ability to form stable high molecular weight multimers. Since the ability of some proteins to polymerize is dependent upon transglutamination, we determined if tissue transglutaminase could catalyze this reaction and the effect of polymerization of IGFBP-1 upon IGF-I action. Following incubation with pure tissue transglutaminase (Tg), IGFBP-1 formed covalently linked multimers that were stable during SDS-polyacrylamide gel electrophoresis using reducing conditions. Dephosphorylated IGFBP-1 polymerized more rapidly and to a greater extent compared with native (phosphorylated) IGFBP-1. Exposure to IGF-I stimulated transglutamination of IGFBP-1 in vitro. An IGFBP-1 mutant in which Gln(66)-Gln(67) had been altered to Ala(66)-Ala(67) (Q66A/Q67A) was relatively resistant to polymerization by Tg compared with native IGFBP-1. Tg localized in fibroblast membranes was also shown to catalyze the formation of native IGFBP-1 multimers, however, Q66A/Q67A IGFBP-1 failed to polymerize. Although the mutant IGFBP-1 potently inhibited IGF-I stimulated protein synthesis in pSMC cultures, the same concentration of native IGFBP-1 had no inhibitory effect. The addition of higher concentrations of native IGFBP-1 did inhibit the protein synthesis response, and this degree of inhibition correlated with the amount of monomeric IGFBP-1 that was present. In conclusion, IGFBP-1 is a substrate for tissue transglutaminase and Tg leads to the formation of high molecular weight covalently linked multimers. Polymerization is an important post translational modification of IGFBP-1 that regulates cellular responses to IGF-I. PMID- 11121405 TI - Cholesterol depletion disrupts caveolae and insulin receptor signaling for metabolic control via insulin receptor substrate-1, but not for mitogen-activated protein kinase control. AB - Insulin exerts its cellular control through receptor binding in caveolae in plasmalemma of target cells (Gustavsson, J., Parpal, S., Karlsson, M., Ramsing, C., Thorn, H., Borg, M., Lindroth, M., Peterson, K. H., Magnusson, K.-E., and Stralfors, P. (1999) FASEB. J. 13, 1961-1971). We now report that a progressive cholesterol depletion of 3T3-L1 adipocytes with beta-cyclodextrin gradually destroyed caveolae structures and concomitantly attenuated insulin stimulation of glucose transport, in effect making cells insulin-resistant. Insulin access to or affinity for the insulin receptor on rat adipocytes was not affected as determined by (125)I-insulin binding. By immunoblotting of plasma membranes, total amount of insulin receptor and of caveolin remained unchanged. Receptor autophosphorylation in response to insulin was not affected by cholesterol depletion. Insulin treatment of isolated caveolae preparations increased autophosphorylation of receptor before and following cholesterol depletion. Insulin-increased tyrosine phosphorylation of an immediate downstream signal transducer, insulin receptor substrate-1, and activation of the further downstream protein kinase B were inhibited. In contrast, insulin signaling to mitogenic control as determined by control of the extracellular signal-related kinases 1/2, mitogen-activated protein kinase pathway was not affected. Insulin did not control Shc phosphorylation, and Shc did not control extracellular signal related kinases 1/2, whereas cholesterol depletion constitutively phosphorylated Shc. In conclusion, caveolae are critical for propagating the insulin receptor signal to downstream targets and have the potential for sorting signal transduction for metabolic and mitogenic effects. PMID- 11121407 TI - Growth factors regulate heterogeneous nuclear ribonucleoprotein K expression and function. AB - Epidermal growth factor (EGF) family of growth factors and their receptors regulate normal and cancerous epithelial cell proliferation, a process that can be suppressed by antireceptor blocking antibodies. To identify genes whose expression may be modulated by antireceptor blocking antibodies, we performed a differential display screen with cells grown in the presence or absence of antireceptor blocking antibodies; isolates from one cDNA clone were 100% identical to human heterogeneous nuclear ribonucleoprotein K (hnRNP K), a protein with a conserved KH motif and RGG boxes, has been implicated in such functions as sequence-specific DNA binding, transcription, RNA binding, and nucleocytoplasmic shuttling. Both EGF and heregulin-beta1 induced expression of hnRNP K mRNA and protein in human breast cancer cells. This growth factor-mediated hnRNP K expression was effectively blocked by pretreatment of cultures with humanized anti-EGF receptor (EGFR) antibody C225, or anti-human epidermal growth factor receptor-2 (HER2) antibody. Anti-EGFR monoclonal antibody also caused regression of human tumor xenografts and reduction in hnRNP K levels in athymic mice. Samples from grade III human breast cancer contained more hnRNP K protein than samples from grade II cancer. Finally, overexpression of hnRNP K in breast cancer cells significantly increased target c-myc promoter activity and c-Myc protein, hnRNP K protein levels, and enhanced breast cancer cell proliferation and growth in an anchorage-independent manner. These results suggested that the activity of human EGF receptor family members regulates hnRNP K expression by extracellular growth promoting signals and that therapeutic humanized antibodies against EGFR and HER2 can effectively block this function. PMID- 11121408 TI - Differential effects of leukocyte common antigen-related protein on biochemical and biological activities of RET-MEN2A and RET-MEN2B mutant proteins. AB - Protein-tyrosine-phosphatases (PTPs), in conjunction with protein-tyrosine kinases, play essential regulatory roles in diverse cellular activities by modulating the phosphorylation state of target proteins. Leukocyte common antigen related (LAR) protein is a widely expressed receptor-type protein-tyrosine phosphatase that is implicated in the regulation of intracellular signaling triggered by both cell adhesion and peptide growth factors. The gene for LAR is localized to human chromosome 1p32, a region frequently deleted in tumors of neuroectodermal origin, including neuroblastoma, pheochromocytoma, and medullary thyroid carcinoma. On the other hand, the RET gene codes for a transmembrane tyrosine kinase and is responsible for the development of multiple endocrine neoplasia (MEN) 2A and 2B. To explore the potential role of LAR in RET tyrosine kinase activity and RET-induced signal transduction, we cotransfected LAR and RET with a MEN2A or MEN2B mutation (designated RET-MEN2A or RET-MEN2B) into the NIH 3T3 cell line. Here we show that LAR reduces the constitutive tyrosine autophosphorylation and kinase activity of RET-MEN2A but not RET-MEN2B, accompanying a significant decrease of phosphorylation of phospholipase Cgamma, AKT, and ERK1/2. Interestingly, LAR expression significantly decreased the levels of disulfide-linked RET-MEN2A dimerization. Moreover, reduced oncogenic activity of RET-MEN2A by overexpression of LAR was observed both by an in vitro colony formation assay and by in vivo tumorigenicity in scid mice. These results thus suggest that LAR may contribute to deactivation of the RET-MEN2A mutant protein and reduction of its oncogenic activity in vivo. PMID- 11121409 TI - Transcytosis of lipoprotein lipase across cultured endothelial cells requires both heparan sulfate proteoglycans and the very low density lipoprotein receptor. AB - Lipoprotein lipase (LPL), the major enzyme responsible for the hydrolysis of circulating lipoprotein triglyceride molecules, is synthesized in myocytes and adipocytes but functions while bound to heparan sulfate proteoglycans (HSPGs) on the luminal surface of vascular endothelial cells. This requires transfer of LPL from the abluminal side to the luminal side of endothelial cells. Studies were performed to investigate the mechanisms of LPL transcytosis using cultured monolayers of bovine aortic endothelial cells. We tested whether HSPGs and members of the low density lipoprotein (LDL) receptor superfamily were involved in transfer of LPL from the basolateral to the apical side of cultured endothelial cells. Heparinase/heparinitase treatment of the basolateral cell surface or addition of heparin to the basolateral medium decreased the movement of LPL. This suggested a requirement for HSPGs. To assess the role of receptors, we used either receptor-associated protein, the 39-kDa inhibitor of ligand binding to the LDL receptor-related protein and the very low density lipoprotein (VLDL) receptor, or specific receptor antibodies. Receptor-associated protein reduced (125)I-LPL and LPL activity transfer across the monolayers. When the basolateral surface of the cells was treated with antibodies, only anti-VLDL receptor antibodies inhibited transcytosis. Moreover, overexpression of the VLDL receptor using adenoviral-mediated gene transfer increased LPL transcytosis. Thus, movement of active LPL across endothelial cells involves both HSPGs and VLDL receptor. PMID- 11121410 TI - A cell system with targeted disruption of the SMN gene: functional conservation of the SMN protein and dependence of Gemin2 on SMN. AB - The motor neuron degenerative disease spinal muscular atrophy is caused by reduced expression of the survival motor neuron (SMN) protein. Here we report a genetic system developed in the chicken pre-B cell line DT40, in which the endogenous SMN gene is disrupted by homologous recombination, and SMN protein is expressed from a chicken SMN cDNA under control of a tetracycline (tet) repressible promoter. Addition of tet results in depletion of SMN protein and consequent cell death, which directly demonstrates that SMN is required for cell viability. The tet-induced lethality can be rescued by expression of human SMN, indicating that the function of SMN is highly conserved between the two species. Cells expressing low levels of SMN display slow growth proportional to the amount of SMN they contain. Interestingly, the level of the SMN-interacting protein Gemin2 decreases significantly following depletion of SMN, supporting the conclusion that SMN and Gemin2 form a stable complex in vivo. This system provides a powerful setting for studying the function of SMN in vivo and for screening for potential therapeutics for spinal muscular atrophy. PMID- 11121411 TI - alpha 2beta 1 integrin is not recognized by rhodocytin but is the specific, high affinity target of rhodocetin, an RGD-independent disintegrin and potent inhibitor of cell adhesion to collagen. AB - We have recombinantly expressed a soluble form of human alpha(2)beta(1) integrin that lacks the membrane-anchoring transmembrane domains as well as the cytoplasmic tails of both integrin subunits. This soluble alpha(2)beta(1) integrin binds to its collagen ligands the same way as the wild-type alpha(2)beta(1) integrin. Furthermore, like the wild-type form, it can be activated by manganese ions and an integrin-activating antibody. However, it does not bind to rhodocytin, a postulated agonist of alpha(2)beta(1) integrin from the snake venom of Calloselasma rhodostoma, which elicits platelet aggregation. Taking advantage of the recombinantly expressed, soluble alpha(2)beta(1) integrin, an inhibition assay was established in which samples can be tested for their capability to inhibit binding of soluble alpha(2)beta(1) integrin to immobilized collagen. Thus, by scrutinizing the C. rhodostoma snake venom in this protein-protein interaction assay, we found a component of the snake venom that inhibits the interaction of soluble alpha(2)beta(1) integrin to type I collagen efficiently. N-terminal sequences identified this inhibitor as rhodocetin, a recently published antagonist of collagen-induced platelet aggregation. We could demonstrate that its inhibitory effect bases on its strong and specific binding to alpha(2)beta(1) integrin, proving that rhodocetin is a disintegrin. Standing apart from the growing group of RGD-dependent snake venom disintegrins, rhodocetin interacts with alpha(2)beta(1) integrin in an RGD-independent manner. Furthermore, its native conformation, which is stabilized by disulfide bridges, is indispensibly required for its inhibitory activity. Rhodocetin does not contain any major collagenous structure despite its high affinity to alpha(2)beta(1) integrin, which binds to collagenous molecules much more avidly than to noncollagenous ligands, such as laminin. Blocking alpha(2)beta(1) integrin as the major collagen receptor on platelets, rhodocetin is responsible for hampering collagen-induced, alpha(2)beta(1) integrin-mediated platelet activation, leading to hemorrhages and bleeding disorders of the snakebite victim. Moreover, having a widespread tissue distribution, alpha(2)beta(1) integrin also mediates cell adhesion, spreading, and migration. We showed that rhodocetin is able to inhibit alpha(2)beta(1) integrin-mediated adhesion of fibrosarcoma cells to type I collagen completely. PMID- 11121412 TI - Prostaglandin endoperoxide H synthase-1: the functions of cyclooxygenase active site residues in the binding, positioning, and oxygenation of arachidonic acid. AB - Prostaglandin endoperoxide H synthases (PGHSs) catalyze the committed step in the biosynthesis of prostaglandins and thromboxane, the conversion of arachidonic acid, two molecules of O(2), and two electrons to prostaglandin endoperoxide H(2) (PGH(2)). Formation of PGH(2) involves an initial oxygenation of arachidonate to yield PGG(2) catalyzed by the cyclooxygenase activity of the enzyme and then a reduction of the 15-hydroperoxyl group of PGG(2) to form PGH(2) catalyzed by the peroxidase activity. The cyclooxygenase active site is a hydrophobic channel that protrudes from the membrane binding domain into the core of the globular domain of PGHS. In the crystal structure of Co(3+)-heme ovine PGHS-1 complexed with arachidonic acid, 19 cyclooxygenase active site residues are predicted to make a total of 50 contacts with the substrate (Malkowski, M. G, Ginell, S., Smith, W. L., and Garavito, R. M. (2000) Science 289, 1933-1937); two of these are hydrophilic, and 48 involve hydrophobic interactions. We performed mutational analyses to determine the roles of 14 of these residues and 4 other closely neighboring residues in arachidonate binding and oxygenation. Mutants were analyzed for peroxidase and cyclooxygenase activity, and the products formed by various mutants were characterized. Overall, the results indicate that cyclooxygenase active site residues of PGHS-1 fall into five functional categories as follows: (a) residues directly involved in hydrogen abstraction from C-13 of arachidonate (Tyr-385); (b) residues essential for positioning C-13 of arachidonate for hydrogen abstraction (Gly-533 and Tyr-348); (c) residues critical for high affinity arachidonate binding (Arg-120); (d) residues critical for positioning arachidonate in a conformation so that when hydrogen abstraction does occur the molecule is optimally arranged to yield PGG(2) versus monohydroperoxy acid products (Val-349, Trp-387, and Leu-534); and (e) all other active site residues, which individually make less but measurable contributions to optimal catalytic efficiency. PMID- 11121413 TI - Mutational and X-ray crystallographic analysis of the interaction of dihomo-gamma -linolenic acid with prostaglandin endoperoxide H synthases. AB - Prostaglandin endoperoxide H synthases-1 and -2 (PGHSs) catalyze the committed step in prostaglandin biosynthesis. Both isozymes can oxygenate a variety of related polyunsaturated fatty acids. We report here the x-ray crystal structure of dihomo-gamma-linolenic acid (DHLA) in the cyclooxygenase site of PGHS-1 and the effects of active site substitutions on the oxygenation of DHLA, and we compare these results to those obtained previously with arachidonic acid (AA). DHLA is bound within the cyclooxygenase site in the same overall L-shaped conformation as AA. C-1 and C-11 through C-20 are in the same positions for both substrates, but the positions of C-2 through C-10 differ by up to 1.74 A. In general, substitutions of active site residues caused parallel changes in the oxygenation of both AA and DHLA. Two significant exceptions were Val-349 and Ser 530. A V349A substitution caused an 800-fold decrease in the V(max)/K(m) for DHLA but less than a 2-fold change with AA; kinetic evidence indicates that C-13 of DHLA is improperly positioned with respect to Tyr-385 in the V349A mutant thereby preventing efficient hydrogen abstraction. Val-349 contacts C-5 of DHLA and appears to serve as a structural bumper positioning the carboxyl half of DHLA, which, in turn, positions properly the omega-half of this substrate. A V349A substitution in PGHS-2 has similar, minor effects on the rates of oxygenation of AA and DHLA. Thus, Val-349 is a major determinant of substrate specificity for PGHS-1 but not for PGHS-2. Ser-530 also influences the substrate specificity of PGHS-1; an S530T substitution causes 40- and 750-fold decreases in oxygenation efficiencies for AA and DHLA, respectively. PMID- 11121414 TI - Rotavirus nonstructural protein NSP2 self-assembles into octamers that undergo ligand-induced conformational changes. AB - The nonstructural protein NSP2 is a component of the rotavirus replication machinery and binds single-stranded RNA cooperatively, with high affinity, and independent of sequence. Recently, NSP2 has been shown to form multimers and to possess an NTPase activity, but its precise function remains unclear. In the present study, we have characterized the solution structure of recombinant NSP2 by velocity and equilibrium ultracentrifugation, dynamic light scattering, and circular dichroism spectroscopy. We found that NSP2 exists as an octamer, which is functional in the binding of RNA and ADP. In the presence of magnesium, a partial dissociation of the octamer into smaller oligomers was observed. This was reversed by binding of ADP and RNA. We observed an increased sedimentation rate in the presence of ADP and a nonhydrolyzable ATP analogue, which suggests a change toward a significantly more compact octameric conformation. The secondary structure of NSP2 showed a high fraction of beta-sheet, with small changes induced by magnesium that were reversed in the presence of RNA. That NSP2 can exist in different conformations lends support to the previously proposed hypothesis (Taraporewala, Z., Chen, D., and Patton, J. T. (1999) J. Virol. 73, 9934-9943) of its function as a molecular motor involved in the packaging of viral mRNA. PMID- 11121416 TI - Requirements and effects of palmitoylation of rat PLD1. AB - Rat brain phospholipase D1 (rPLD1) has two highly conserved motifs (HXKX(4)D), denoted HKD, located in the N- and C-terminal halves, which are required for phospholipase D activity. The two halves of rPLD1 can associate in vivo, and the association is essential for catalytic activity and Ser/Thr phosphorylation of the enzyme. In this study, we found that this association is also required for palmitoylation of rPLD1, which occurs on cysteines 240 and 241. In addition, palmitoylation of rPLD1 requires the N-terminal sequence but not the conserved C terminal sequence, since rPLD1 that lacks the first 168 amino acids is not palmitoylated in vivo, while the inactive C-terminal deletion mutant is. Palmitoylation of rPLD1 is not necessary for catalytic activity, since N-terminal truncation mutants lacking the first 168 or 319 amino acids exhibit high basal activity although they cannot be stimulated by protein kinase C (PKC). The lack of response to PKC is not due to the lack of palmitoylation, since mutation of both Cys(240) and Cys(241) to alanine in full-length rPLD1 abolishes palmitoylation, but the mutant still retains basal activity and responds to PKC. Palmitoylation-deficient rPLD1 can associate with crude membranes; however, the association is weakened. Wild type rPLD1 remains membrane-associated when extracted with 1 m NaCl or Na(2)CO(3) (pH 11), while rPLD1 mutants that lack palmitoylation are partially released. In addition, we found that palmitoylation deficient mutants are much less modified by Ser/Thr phosphorylation compared with wild type rPLD1. Characterization of the other cysteine mutations of rPLD1 showed that mutation of cysteine 310 or 612 to alanine increased basal phospholipase D activity 2- and 4-fold, respectively. In summary, palmitoylation of rPLD1 requires interdomain association and the presence of the N-terminal 168 amino acids. Mutations of cysteines 240 and 241 to alanine abolish the extensive Ser/Thr phosphorylation of the enzyme and weaken its association with membranes. PMID- 11121415 TI - Signalling pathways regulating the dephosphorylation of Ser729 in the hydrophobic domain of protein kinase Cepsilon upon cell passage. AB - We have recently demonstrated that in quiescent fibroblasts protein kinase C (PKC) epsilon(95) is phosphorylated at Ser(729), Ser(703), and Thr(566) and that upon passage of quiescent cells phosphorylation at Ser(729) is lost, giving rise to PKCepsilon(87). Ser(729) may be rephosphorylated later, suggesting cycling between PKCepsilon(87) and PKCepsilon(95). Here we show that the dephosphorylation at Ser(729) is insensitive to okadaic acid, calyculin, ascomycin C, and cyclosporin A, suggesting that dephosphorylation at this site is not mediated through protein phosphatases 1, 2A or 2B. We demonstrate that this dephosphorylation at Ser(729) requires serum and cell readhesion and is sensitive to rapamycin, PD98059, chelerythrine, and Ro-31-8220. These results suggest that the phosphorylation status of Ser(729) in the hydrophobic domain at Ser(729) is regulated independently of the phosphorylation status of other sites in PKCepsilon, by a mTOR-sensitive phosphatase. The mitogen-activated protein kinase pathway and PKC are also implicated in regulating the dephosphorylation at Ser(729). PMID- 11121417 TI - Induction of early growth response-1 gene expression by calmodulin antagonist trifluoperazine through the activation of Elk-1 in human fibrosarcoma HT1080 cells. AB - The early growth response gene-1 (Egr-1) is a transcription factor that plays an important role in cell growth and differentiation. It has been known that Egr-1 expression is down-regulated in many types of tumor tissues, including human fibrosarcoma HT1080 cells, and introduction of the Egr-1 gene into HT1080 cells inhibits cell growth and tumorigenic potential. Trifluoperazine (TFP), a phenothiazine class calmodulin antagonist, is known to inhibit DNA synthesis and cell proliferation and potentially important in antitumor activities. To understand the regulatory mechanism of Egr-1, we investigated the effect of TFP on expression of Egr-1 in HT1080 cells. Herein, we report that Egr-1 expression was increased by TFP in synergy with serum at the transcriptional level. Both the Ca(2+)/calmodulin-dependent protein kinase II inhibitor KN62 and the calcineurin inhibitor cyclosporin A enhanced TFP-dependent increase of Egr-1, suggesting that the Ca(2+)/calmodulindependent pathway plays a role in regulation of Egr-1 expression in HT1080 cells. The TFP-stimulated increase of the Egr-1 protein was preferentially inhibited by the MEK-specific inhibitor PD98059. In addition, activation of human Egr-1 promoter and the transcriptional activation of the ternary complex factor Elk-1 induced by TFP were inhibited both by pretreatment of PD98059 and by expression of the dominant-negative RasN17. These results indicate that the Ras/MEK/Erk/Elk-1 pathway is necessary for TFP-induced Egr-1 expression. We propose that the calmodulin antagonist TFP stimulates Egr-1 gene expression by modulating Ras/MEK/Erk and activation of the Elk-1 pathway in human fibrosarcoma HT1080 cells. PMID- 11121418 TI - The cytosolic O-acetylserine(thiol)lyase gene is regulated by heavy metals and can function in cadmium tolerance. AB - Regulation of the expression of the cytosolic O-acetylserine(thiol)lyase gene (Atcys-3A) from Arabidopsis thaliana under heavy metal stress conditions has been investigated. Northern blot analysis of Atcys-3A expression shows a 7-fold induction after 18 h of cadmium treatment. Addition of 50 microm CdCl(2) to the irrigation medium of mature Arabidopsis plants induces a rapid accumulation of the mRNA throughout the leaf lamina, the root and stem cortex, and stem vascular tissues when compared with untreated plants, as observed by in situ hybridization. High pressure liquid chromatography analysis of GSH content shows a transient increase after 18 h of metal treatment. Our results are compatible with a high cysteine biosynthesis rate under heavy metal stress required for the synthesis of GSH and phytochelatins, which are involved in the plant detoxification mechanism. Arabidopsis-transformed plants overexpressing the Atcys 3A gene by up to 9-fold show increased tolerance to cadmium when grown in medium containing 250 microm CdCl(2), suggesting that increased cysteine availability is responsible for cadmium tolerance. In agreement with these results, exogenous addition of cystine can, to some extent, also favor the growth of wild-type plants in cadmium-containing medium. Cadmium accumulates to higher levels in leaves of tolerant transformed lines than in wild-type plants. PMID- 11121419 TI - Highly saturated endonuclear phosphatidylcholine is synthesized in situ and colocated with CDP-choline pathway enzymes. AB - Chromatin-associated phospholipids are well recognized. A report that catalytically active endonuclear CTP:choline-phosphate cytidylyltransferase alpha is necessary for cell survival questions whether endonuclear, CDP-choline pathway phosphatidylcholine synthesis may occur in situ. We report that chromatin from human IMR-32 neuroblastoma cells possesses such a biosynthetic pathway. First, membrane-free nuclei retain all three CDP-choline pathway enzymes in proportions comparable with the content of chromatin-associated phosphatidylcholine. Second, following supplementation of cells with deuterated choline and using electrospray ionization mass spectrometry, both the time course and molecular species labeling pattern of newly synthesized endonuclear and whole cell phosphatidylcholine revealed the operation of spatially separate, compositionally distinct biosynthetic routes. Specifically, endogenous and newly synthesized endonuclear phosphatidylcholine species are both characterized by a high degree of diacyl/alkylacyl chain saturation. This unusual species content and synthetic pattern (evident within 10 min of supplementation) are maintained through cell growth arrest by serum depletion and when proliferation is restored, suggesting that endonuclear disaturated phosphatidylcholine enrichment is essential and closely regulated. We propose that endonuclear phosphatidylcholine synthesis may regulate periodic nuclear accumulations of phosphatidylcholine-derived lipid second messengers. Furthermore, our estimates of saturated phosphatidylcholine nuclear volume occupancy of around 10% may imply a significant additional role in regulating chromatin structure. PMID- 11121420 TI - Correlation between steady-state ATP hydrolysis and vanadate-induced ADP trapping in Human P-glycoprotein. Evidence for ADP release as the rate-limiting step in the catalytic cycle and its modulation by substrates. AB - P-glycoprotein (Pgp) is a transmembrane protein conferring multidrug resistance to cells by extruding a variety of amphipathic cytotoxic agents using energy from ATP hydrolysis. The objective of this study was to understand how substrates affect the catalytic cycle of ATP hydrolysis by Pgp. The ATPase activity of purified and reconstituted recombinant human Pgp was measured using a continuous cycling assay. Pgp hydrolyzes ATP in the absence of drug at a basal rate of 0.5 micromol x min x mg(-1) with a K(m) for ATP of 0.33 mm. This basal rate can be either increased or decreased depending on the Pgp substrate used, without an effect on the K(m) for ATP or 8-azidoATP and K(i) for ADP, suggesting that substrates do not affect nucleotide binding to Pgp. Although inhibitors of Pgp activity, cyclosporin A, its analog PSC833, and rapamycin decrease the rate of ATP hydrolysis with respect to the basal rate, they do not completely inhibit the activity. Therefore, these drugs can be classified as substrates. Vanadate (Vi) induced trapping of [alpha-(32)P]8-azidoADP was used to probe the effect of substrates on the transition state of the ATP hydrolysis reaction. The K(m) for [alpha-(32)P]8-azidoATP (20 microm) is decreased in the presence of Vi; however, it is not changed by drugs such as verapamil or cyclosporin A. Strikingly, the extent of Vi-induced [alpha-(32)P]8-azidoADP trapping correlates directly with the fold stimulation of ATPase activity at steady state. Furthermore, P(i) exhibits very low affinity for Pgp (K(i) approximately 30 mm for Vi-induced 8 azidoADP trapping). In aggregate, these data demonstrate that the release of Vi trapped [alpha-(32)P]8-azidoADP from Pgp is the rate-limiting step in the steady state reaction. We suggest that substrates modulate the rate of ATPase activity of Pgp by controlling the rate of dissociation of ADP following ATP hydrolysis and that ADP release is the rate-limiting step in the normal catalytic cycle of Pgp. PMID- 11121421 TI - Phosphorylation site mutations in heterochromatin protein 1 (HP1) reduce or eliminate silencing activity. AB - HP1 is an essential heterochromatin-associated protein in Drosophila. HP1 has dosage-dependent effects on the silencing of euchromatic genes that are mislocalized to heterochromatin and is required for the normal expression of at least two heterochromatic genes. HP1 is multiply phosphorylated in vivo, and HP1 hyperphosphorylation is correlated with heterochromatin assembly during development. The purpose of this study was to test whether HP1 phosphorylation modifies biological activity and biochemical properties of HP1. To determine sites of HP1 phosphorylation in vivo and whether phosphorylation affects any biochemical properties of HP1, we expressed Drosophila HP1 in lepidopteran cultured cells using a recombinant baculovirus vector. Phosphopeptides were identified by matrix-assisted laser desorption ionization/time of flight mass spectroscopy; these peptides contain target sites for casein kinase II, protein tyrosine kinase, and PIM-1 kinase. Purified HP1 from bacterial (unphosphorylated) and lepidopteran (phosphorylated) cells has similar secondary structure. Phosphorylation has no effect on HP1 self-association but alters the DNA binding properties of HP1, suggesting that phosphorylation could differentially regulate HP1-dependent interactions. Serine-to-alanine and serine-to-glutamate substitutions at consensus protein kinase motifs resulted in reduction or loss of silencing activity of mutant HP1 in transgenic flies. These results suggest that dynamic phosphorylation/dephosphorylation regulates HP1 activity in heterochromatic silencing. PMID- 11121422 TI - Disruption of an active site hydrogen bond converts human heme oxygenase-1 into a peroxidase. AB - The crystal structure of heme oxygenase-1 suggests that Asp-140 may participate in a hydrogen bonding network involving ligands coordinated to the heme iron atom. To examine this possibility, Asp-140 was mutated to an alanine, phenylalanine, histidine, leucine, or asparagine, and the properties of the purified proteins were investigated. UV-visible and resonance Raman spectroscopy indicate that the distal water ligand is lost from the iron in all the mutants except, to some extent, the D140N mutant. In the D140H mutant, the distal water ligand is replaced by the new His-140 as the sixth iron ligand, giving a bis histidine complex. The D140A, D140H, and D140N mutants retain a trace (<3%) of biliverdin forming activity, but the D140F and D140L mutants are inactive in this respect. However, the two latter mutants retain a low ability to form verdoheme, an intermediate in the reaction sequence. All the Asp-140 mutants exhibit a new peroxidase activity. The results indicate that disruption of the distal hydrogen bonding environment by mutation of Asp-140 destabilizes the ferrous dioxygen complex and promotes conversion of the ferrous hydroperoxy intermediate obtained by reduction of the ferrous dioxygen complex to a ferryl species at the expense of its normal reaction with the porphyrin ring. PMID- 11121423 TI - The cadherin cytoplasmic domain is unstructured in the absence of beta-catenin. A possible mechanism for regulating cadherin turnover. AB - Cadherins are single pass transmembrane proteins that mediate Ca(2+)-dependent homophilic cell-cell adhesion by linking the cytoskeletons of adjacent cells. In adherens junctions, the cytoplasmic domain of cadherins bind to beta-catenin, which in turn binds to the actin-associated protein alpha-catenin. The physical properties of the E-cadherin cytoplasmic domain and its interactions with beta catenin have been investigated. Proteolytic sensitivity, tryptophan fluorescence, circular dichroism, and (1)H NMR measurements indicate that murine E-cadherin cytoplasmic domain is unstructured. Upon binding to beta-catenin, the domain becomes resistant to proteolysis, suggesting that it structures upon binding. Cadherin-beta-catenin complex stability is modestly dependent on ionic strength, indicating that, contrary to previous proposals, the interaction is not dominated by electrostatics. Comparison of 18 cadherin sequences indicates that their cytoplasmic domains are unlikely to be structured in isolation. This analysis also reveals the presence of PEST sequences, motifs associated with ubiquitin/proteosome degradation, that overlap the previously identified beta catenin-binding site. It is proposed that binding of cadherins to beta-catenin prevents recognition of degradation signals that are exposed in the unstructured cadherin cytoplasmic domain, favoring a cell surface population of catenin-bound cadherins capable of participating in cell adhesion. PMID- 11121424 TI - A 72-base pair AT-rich DNA sequence element functions as a bacterial gene silencer. AB - We have previously demonstrated that sequential activation of the bacterial ilvIH leuO-leuABCD gene cluster involves a promoter-relay mechanism. In the current study, we show that the final activation of the leuABCD operon is through a transcriptional derepression mechanism. The leuABCD operon is transcriptionally repressed by the presence of a 318-base pair AT-rich upstream element. LeuO is required for derepressing the repressed leuABCD operon. Deletion analysis of the repressive effect of the 318-bp element has led to the identification of a 72-bp AT-rich (78% A+T) DNA sequence element, AT4, which is capable of silencing a number of unrelated promoters in addition to the leuABCD promoter. AT4-mediated gene silencing is orientation-independent and occurs within a distance of 300 base pairs. Furthermore, an increased gene-silencing effect was observed with a tandemly repeated AT4 dimer. The possible mechanism of AT4-mediated gene silencing in bacteria is discussed. PMID- 11121425 TI - A molecular link between the common phenotypes of type 1 glycogen storage disease and HNF1alpha-null mice. AB - The clinical manifestations of type 1 glycogen storage disease (GSD-1) in patients deficient in the glucose-6-phosphatase (G6Pase) system (e.g. growth retardation, hepatomegaly, hyperlipidemia, and renal dysfunction) are shared by Hnf1alpha(-/-) mice deficient of a transcriptional activator, hepatocyte nuclear factor 1alpha (HNF1alpha). However, the molecular mechanism is unknown. The G6Pase system, essential for the maintenance of glucose homeostasis, is comprised of glucose 6-phosphate transporter (G6PT) and G6Pase. G6PT translocates G6P from the cytoplasm to the lumen of the endoplasmic reticulum where it is metabolized by G6Pase to glucose and phosphate. Deficiencies in G6Pase and G6PT cause GSD-1a and GSD-1b, respectively. Hnf1alpha(-/-) mice also develop noninsulin-dependent diabetes mellitus caused by defective insulin secretion. In this study, we sought to determine whether there is a molecular link between HNF1alpha deficiency and function of the G6Pase system. Transactivation studies revealed that HNF1alpha is required for transcription of the G6PT gene. Hepatic G6PT mRNA levels and microsomal G6P transport activity are also markedly reduced in Hnf1alpha(-/-) mice as compared with Hnf1alpha(+/+) and Hnf1alpha(+/-) littermates. On the other hand, hepatic G6Pase mRNA expression and activity are up-regulated in Hnf1alpha( /-) mice, consistent with observations that G6Pase expression is increased in diabetic animals. Taken together, the results strongly suggest that metabolic abnormalities in HNF1alpha-null mice are caused in part by G6PT deficiency and by perturbations of the G6Pase system. PMID- 11121426 TI - A temperature-sensitive mutation of Crygs in the murine Opj cataract. AB - In Opj, an inherited cataract in mice, opacity is associated with a mutation in Crygs, the gene for gammaS-crystallin, the first mutation to be associated with this gene. A single base change causes replacement of Phe-9, a key hydrophobic residue in the core of the N-terminal domain, by serine. Despite this highly non conservative change, mutant protein folds normally at low temperature. However, it exhibits a marked, concentration-dependent decrease in solubility, associated with loss of secondary structure, at close to physiological temperatures. This is reminiscent of processes thought to occur in human senile cataracts in which normal proteins become altered and aggregate. The Opj cataract is progressive and more severe in Opj/Opj than in Opj/+. Lens histology shows that whereas fiber cell morphology in Opj/+ mice is essentially normal, in Opj/Opj, cortical fiber cell morphology and the loss of maturing fiber cell nuclei are both severely disrupted from early stages. This may indicate a loss of function of gammaS crystallin which would be consistent with ideas that members of the betagamma crystallin superfamily may have roles associated with maintenance of cytoarchitecture. PMID- 11121427 TI - Estrogen decreases osteoclast formation by down-regulating receptor activator of NF-kappa B ligand (RANKL)-induced JNK activation. AB - The differentiation of cells of the monocytic lineage into mature osteoclasts (OC) is specifically induced by the tumor necrosis factor-related factor, RANKL (receptor activator of NF-kappaB ligand; also known as OPGL, ODF, or TRANCE). Because inhibition of osteoclastogenesis is one of the main mechanisms by which estrogen (E2) prevents bone loss, it is likely that E2 may regulate either the production of, or the target cell responsiveness to RANKL. We found that E2 decreases the differentiation into OC of both murine bone marrow monocytes and RAW 264.7 cells, a monocytic line, by down-regulating the activation of Jun N terminal kinase 1 (JNK1). Diminished JNK1 activity results in decreased nuclear levels of the key osteoclastogenic transcription factors, c-Fos and c-Jun, and lower binding of these transcriptional inducers to DNA. Thus, one novel mechanism by which E2 down-regulates osteoclastogenesis is by decreasing the responsiveness of OC precursors to RANKL. PMID- 11121428 TI - Distinct domains of CD98hc regulate integrins and amino acid transport. AB - CD98 is a cell surface heterodimer formed by the covalent linkage of CD98 heavy chain (CD98hc) with several different light chains to form amino acid transporters. CD98hc also binds specifically to the integrin beta(1A) cytoplasmic domain and regulates integrin function. In this study, we examined the relationship between the ability of CD98hc to stimulate amino acid transport and to affect integrin function. By constructing chimeras with CD98hc and a type II transmembrane protein (CD69), we found that the cytoplasmic and transmembrane domains of CD98hc are required for its effects on integrin function, while the extracellular domain is required for stimulation of isoleucine transport. Consequently, the capacity to promote amino acid transport is not required for CD98hc's effect on integrin function. Furthermore, a mutant of CD98hc that lacks its integrin binding site can still promote increased isoleucine transport. Thus, these two functions of CD98hc are separable and require distinct domains of the protein. PMID- 11121429 TI - Muscular dystrophy meets the gene chip: new insights into disease pathogenesis. PMID- 11121430 TI - Coordinate control of muscle cell survival by distinct insulin-like growth factor activated signaling pathways. AB - Peptide growth factors control diverse cellular functions by regulating distinct signal transduction pathways. In cultured myoblasts, insulin-like growth factors (IGFs) stimulate differentiation and promote hypertrophy. IGFs also maintain muscle cell viability. We previously described C2 skeletal muscle lines lacking expression of IGF-II. These cells did not differentiate, but underwent progressive apoptotic death when incubated in differentiation medium. Viability could be sustained and differentiation enabled by IGF analogues that activated the IGF-I receptor; survival was dependent on stimulation of phosphatidylinositol 3-kinase (PI3-kinase). We now find that IGF action promotes myoblast survival through two distinguishable PI3-kinase-regulated pathways that culminate in expression of the cyclin-dependent kinase inhibitor, p21. Incubation with IGF-I or transfection with active PI3-kinase led to rapid induction of MyoD and p21, and forced expression of either protein maintained viability in the absence of growth factors. Ectopic expression of MyoD induced p21, and inhibition of p21 blocked MyoD-mediated survival, thus defining one PI3-kinase-dependent pathway as leading first to MyoD, and then to p21 and survival. Unexpectedly, loss of MyoD expression did not impede IGF-mediated survival, revealing a second pathway involving activation by PI3-kinase of Akt, and subsequent induction of p21. Since inhibition of p21 caused death even in the presence of IGF-I, these results establish a central role for p21 as a survival factor for muscle cells. Our observations also define a MyoD-independent pathway for regulating p21 in muscle, and demonstrate that distinct mechanisms help ensure appropriate expression of this key protein during differentiation. PMID- 11121431 TI - Nischarin, a novel protein that interacts with the integrin alpha5 subunit and inhibits cell migration. AB - Integrins have been implicated in key cellular functions, including cytoskeletal organization, motility, growth, survival, and control of gene expression. The plethora of integrin alpha and beta subunits suggests that individual integrins have unique biological roles, implying specific molecular connections between integrins and intracellular signaling or regulatory pathways. Here, we have used a yeast two-hybrid screen to identify a novel protein, termed Nischarin, that binds preferentially to the cytoplasmic domain of the integrin alpha5 subunit, inhibits cell motility, and alters actin filament organization. Nischarin is primarily a cytosolic protein, but clearly associates with alpha5beta1, as demonstrated by coimmunoprecipitation. Overexpression of Nischarin markedly reduces alpha5beta1-dependent cell migration in several cell types. Rat embryo fibroblasts transfected with Nischarin constructs have "basket-like" networks of peripheral actin filaments, rather than typical stress fibers. These observations suggest that Nischarin might affect signaling to the cytoskeleton regulated by Rho-family GTPases. In support of this, Nischarin expression reverses the effect of Rac on lamellipodia formation and selectively inhibits Rac-mediated activation of the c-fos promoter. Thus, Nischarin may play a negative role in cell migration by antagonizing the actions of Rac on cytoskeletal organization and cell movement. PMID- 11121432 TI - Nuclear lamins A and B1: different pathways of assembly during nuclear envelope formation in living cells. AB - At the end of mitosis, the nuclear lamins assemble to form the nuclear lamina during nuclear envelope formation in daughter cells. We have fused A- and B-type nuclear lamins to the green fluorescent protein to study this process in living cells. The results reveal that the A- and B-type lamins exhibit different pathways of assembly. In the early stages of mitosis, both lamins are distributed throughout the cytoplasm in a diffusible (nonpolymerized) state, as demonstrated by fluorescence recovery after photobleaching (FRAP). During the anaphase telophase transition, lamin B1 begins to become concentrated at the surface of the chromosomes. As the chromosomes reach the spindle poles, virtually all of the detectable lamin B1 has accumulated at their surfaces. Subsequently, this lamin rapidly encloses the entire perimeter of the region containing decondensing chromosomes in each daughter cell. By this time, lamin B1 has assembled into a relatively stable polymer, as indicated by FRAP analyses and insolubility in detergent/high ionic strength solutions. In contrast, the association of lamin A with the nucleus begins only after the major components of the nuclear envelope including pore complexes are assembled in daughter cells. Initially, lamin A is found in an unpolymerized state throughout the nucleoplasm of daughter cell nuclei in early G1 and only gradually becomes incorporated into the peripheral lamina during the first few hours of this stage of the cell cycle. In later stages of G1, FRAP analyses suggest that both green fluorescent protein lamins A and B1 form higher order polymers throughout interphase nuclei. PMID- 11121433 TI - MAP1B is required for axon guidance and Is involved in the development of the central and peripheral nervous system. AB - Microtubule-associated proteins such as MAP1B have long been suspected to play an important role in neuronal differentiation, but proof has been lacking. Previous MAP1B gene targeting studies yielded contradictory and inconclusive results and did not reveal MAP1B function. In contrast to two earlier efforts, we now describe generation of a complete MAP1B null allele. Mice heterozygous for this MAP1B deletion were not affected. Homozygous mutants were viable but displayed a striking developmental defect in the brain, the selective absence of the corpus callosum, and the concomitant formation of myelinated fiber bundles consisting of misguided cortical axons. In addition, peripheral nerves of MAP1B-deficient mice had a reduced number of large myelinated axons. The myelin sheaths of the remaining axons were of reduced thickness, resulting in a decrease of nerve conduction velocity in the adult sciatic nerve. On the other hand, the anticipated involvement of MAP1B in retinal development and gamma-aminobutyric acid C receptor clustering was not substantiated. Our results demonstrate an essential role of MAP1B in development and function of the nervous system and resolve a previous controversy over its importance. PMID- 11121434 TI - Reversal of the Ras-induced transformed phenotype by HR12, a novel ras farnesylation inhibitor, is mediated by the Mek/Erk pathway. AB - We have used the selective farnesylation inhibitor HR12 [cysteine-N(methyl)valine N(cyclohexyl) glycine-methionine-O-methyl-ester] to study the role of oncogenic Ras in cytoskeletal reorganization in Ha-ras(V12)-transformed Rat1 cells (Rat1/ras). Application of HR12 resulted in complete restoration of the cytoskeleton and associated cell adhesions disrupted by oncogenic Ras. This included an increase in the number and size of focal adhesions, accompanied by massive stress fiber formation and enhanced tyrosine phosphorylation. Furthermore, HR12 induced assembly of adherens junctions and dramatically elevated the level of the junctional components, cadherin and beta-catenin. HR12 was unable to restore the nontransformed phenotype in cells expressing farnesylation-independent, myristylated Ras. Examination of the main Ras regulated signaling pathways revealed that HR12 induced a dose- and time dependent decline in Erk1&2 activation (t(1/2) approximately 6 h), which correlated with the accumulation of nonfarnesylated oncogenic-Ras. Inhibition of the Mek/Erk pathway in Rat1/ras cells, using the Mek inhibitor, PD98059, resulted in complete cytoskeletal recovery, indistinguishable from that induced by HR12. Moreover, a constitutively active Mek mimicked the effect of ras transformation in Rat1 cells, and prevented HR12-induced cytoskeletal effects in Rat1/ras cells. No such effects were observed after treatment of Rat1/ras cells with the phosphatidylinositol 3-kinase inhibitor LY294002. These findings establish the Mek/Erk pathway as the dominant pathway involved in conferring the cytoskeletal and junctional manifestations of the Ras-induced transformed phenotype. PMID- 11121435 TI - N-Cadherin extracellular repeat 4 mediates epithelial to mesenchymal transition and increased motility. AB - E- and N-cadherin are members of the classical cadherin family of proteins. E cadherin plays an important role in maintaining the normal phenotype of epithelial cells. Previous studies from our laboratory and other laboratories have shown that inappropriate expression of N-cadherin by tumor cells derived from epithelial tissue results in conversion of the cell to a more fibroblast like cell, with increased motility and invasion. Our present study was designed to determine which domains of N-cadherin make it different from E-cadherin, with respect to altering cellular behavior, such as which domains are responsible for the epithelial to mesenchymal transition and increased cell motility and invasion. To address this question, we constructed chimeric cadherins comprised of selected domains of E- and N-cadherin. The chimeras were transfected into epithelial cells to determine their effect on cell morphology and cellular behavior. We found that a 69-amino acid portion of EC-4 of N-cadherin was necessary and sufficient to promote both an epithelial to mesenchymal transition in squamous epithelial cells and increased cell motility. Here, we show that different cadherin family members promote different cellular behaviors. In addition, we identify a novel activity that can be ascribed to the extracellular domain of N-cadherin. PMID- 11121436 TI - Rab11 regulates the compartmentalization of early endosomes required for efficient transport from early endosomes to the trans-golgi network. AB - Several GTPases of the Rab family, known to be regulators of membrane traffic between organelles, have been described and localized to various intracellular compartments. Rab11 has previously been reported to be associated with the pericentriolar recycling compartment, post-Golgi vesicles, and the trans-Golgi network (TGN). We compared the effect of overexpression of wild-type and mutant forms of Rab11 on the different intracellular transport steps in the endocytic/degradative and the biosynthetic/exocytic pathways in HeLa cells. We also studied transport from endosomes to the Golgi apparatus using the Shiga toxin B subunit (STxB) and TGN38 as reporter molecules. Overexpression of both Rab11 wild-type (Rab11wt) and mutants altered the localization of the transferrrin receptor (TfR), internalized Tf, the STxB, and TGN38. In cells overexpressing Rab11wt and in a GTPase-deficient Rab11 mutant (Rab11Q70L), these proteins were found in vesicles showing characteristics of sorting endosomes lacking cellubrevin (Cb). In contrast, they were redistributed into an extended tubular network, together with Cb, in cells overexpressing a dominant negative mutant of Rab11 (Rab11S25N). This tubularized compartment was not accessible to Tf internalized at temperatures <20 degrees C, suggesting that it is of recycling endosomal origin. Overexpression of Rab11wt, Rab11Q70L, and Rab11S25N also inhibited STxB and TGN38 transport from endosomes to the TGN. These results suggest that Rab11 influences endosome to TGN trafficking primarily by regulating membrane distribution inside the early endosomal pathway. PMID- 11121437 TI - Expression of CD34 and Myf5 defines the majority of quiescent adult skeletal muscle satellite cells. AB - Skeletal muscle is one of a several adult post-mitotic tissues that retain the capacity to regenerate. This relies on a population of quiescent precursors, termed satellite cells. Here we describe two novel markers of quiescent satellite cells: CD34, an established marker of hematopoietic stem cells, and Myf5, the earliest marker of myogenic commitment. CD34(+ve) myoblasts can be detected in proliferating C2C12 cultures. In differentiating cultures, CD34(+ve) cells do not fuse into myotubes, nor express MyoD. Using isolated myofibers as a model of synchronous precursor cell activation, we show that quiescent satellite cells express CD34. An early feature of their activation is alternate splicing followed by complete transcriptional shutdown of CD34. This data implicates CD34 in the maintenance of satellite cell quiescence. In heterozygous Myf5(nlacZ/+) mice, all CD34(+ve) satellite cells also express beta-galactosidase, a marker of activation of Myf5, showing that quiescent satellite cells are committed to myogenesis. All such cells are positive for the accepted satellite cell marker, M-cadherin. We also show that satellite cells can be identified on isolated myofibers of the myosin light chain 3F-nlacZ-2E mouse as those that do not express the transgene. The numbers of satellite cells detected in this way are significantly greater than those identified by the other three markers. We conclude that the expression of CD34, Myf5, and M-cadherin defines quiescent, committed precursors and speculate that the CD34(-ve), Myf5(-ve) minority may be involved in maintaining the lineage-committed majority. PMID- 11121438 TI - Induction of terminal differentiation in epithelial cells requires polymerization of hensin by galectin 3. AB - During terminal differentiation, epithelia become columnar and develop specialized apical membrane structures (microvilli) and functions (regulated endocytosis and exocytosis). Using a clonal intercalated epithelial cell line, we found that high seeding density induced these characteristics, whereas low density seeding maintained a protoepithelial state. When cells were plated at low density, but on the extracellular matrix of high density cells, they converted to the more differentiated phenotype. The extracellular matrix (ECM) protein responsible for this activity was purified and found to be a large 230-kD protein, which we termed hensin. High density seeding caused hensin to be polymerized and deposited in the extracellular matrix, and only this form of hensin was able to induce terminal differentiation. Antibodies to hensin blocked the change in phenotype. However, its purification to homogeneity resulted in loss of activity, suggesting that an additional protein might be necessary for induction of terminal differentiation. Here, we found that a 29-kD protein specifically associates with hensin in the ECM. Addition of purified p29 restored the activity of homogenously purified hensin. Mass fingerprinting identified p29 as galectin 3. Purified recombinant galectin 3 was able to bind to hensin and to polymerize it in vitro. Seeding cells at high density induced secretion of galectin 3 into the ECM where it bundled hensin. Hence, the high density state causes a secretion of a protein that acts on another ECM protein to allow the new complex to signal the cell to change its phenotype. This is a new mechanism of inside-out signaling. PMID- 11121439 TI - Necrotic death pathway in Fas receptor signaling. AB - A caspase 8-deficient subline (JB6) of human Jurkat cells can be killed by the oligomerization of Fas-associated protein with death domain (FADD). This cell death process is not accompanied by caspase activation, but by necrotic morphological changes. Here, we show that the death effector domain of FADD is responsible for the FADD-mediated necrotic pathway. This process was accompanied by a loss of mitochondrial transmembrane potential (DeltaPsim), but not by the release of cytochrome c from mitochondria. Pyrrolidine dithiocarbamate, a metal chelator and antioxidant, efficiently inhibited the FADD-induced reduction of DeltaPsim and necrotic cell death. When human Jurkat, or its transformants, expressing mouse Fas were treated with Fas ligand or anti-mouse Fas antibodies, the cells died, showing characteristics of apoptosis. A broad caspase inhibitor (z-VAD-fmk) blocked the apoptotic morphological changes and the release of cytochrome c. However, the cells still died, and this cell death process was accompanied by a strong reduction in DeltaPsim, as well as necrotic morphological changes. The presence of z-VAD-fmk and pyrrolidine dithiocarbamate together blocked cell death, suggesting that both apoptotic and necrotic pathways can be activated through the Fas death receptor. PMID- 11121440 TI - Dynamic shuttling of TIA-1 accompanies the recruitment of mRNA to mammalian stress granules. AB - Mammalian stress granules (SGs) harbor untranslated mRNAs that accumulate in cells exposed to environmental stress. Drugs that stabilize polysomes (emetine) inhibit the assembly of SGs, whereas drugs that destabilize polysomes (puromycin) promote the assembly of SGs. Moreover, emetine dissolves preformed SGs as it promotes the assembly of polysomes, suggesting that these mRNP species (i.e., SGs and polysomes) exist in equilibrium. We used green flourescent protein-tagged SG associated RNA-binding proteins (specifically, TIA-1 and poly[A] binding protein [PABP-I]) to monitor SG assembly, disassembly, and turnover in live cells. Fluorescence recovery after photobleaching shows that both TIA-1 and PABP-I rapidly and continuously shuttle in and out of SGs, indicating that the assembly of SGs is a highly dynamic process. This unexpected result leads us to propose that mammalian SGs are sites at which untranslated mRNAs are sorted and processed for either reinitiation, degradation, or packaging into stable nonpolysomal mRNP complexes. A truncation mutant of TIA-1 (TIA-1DeltaRRM), which acts as a transdominant inhibitor of SG assembly, promotes the expression of cotransfected reporter genes in COS transfectants, suggesting that this process of mRNA triage might, directly or indirectly, influence protein expression. PMID- 11121441 TI - Spatial sensing in fibroblasts mediated by 3' phosphoinositides. AB - The directed movement of fibroblasts towards locally released platelet-derived growth factor (PDGF) is a critical event in wound healing. Although recent studies have implicated polarized activation of phosphoinositide (PI) 3-kinase in G protein-mediated chemotaxis, the role of 3' PI lipids in tyrosine kinase triggered chemotaxis is not well understood. Using evanescent wave microscopy and green fluorescent protein-tagged Akt pleckstrin homology domain (GFP-AktPH) as a molecular sensor, we show that application of a shallow PDGF gradient triggers a markedly steeper gradient in 3' PI lipids in the adhesion zone of fibroblasts. Polar GFP-AktPH gradients, as well as a new type of radial gradient, were measured from front to rear and from the periphery to the center of the adhesion zone, respectively. A strong spatial correlation between polarized 3' PI production and rapid membrane spreading implicates 3' PI lipids as a direct mediator of polarized migration. Analysis of the temporal changes of 3' PI gradients in the adhesion zone revealed a fast diffusion coefficient (0.5 microm(2)/s) and short lifetime of 3' PIs of <1 min. Together, this study suggests that the tyrosine kinase-coupled directional movement of fibroblasts and their radial membrane activity are controlled by local generation and rapid degradation of 3' PI second messengers. PMID- 11121442 TI - Endocytosis and recycling of the HIV coreceptor CCR5. AB - The chemokine receptor CCR5 is a cofactor for the entry of R5 tropic strains of human immunodeficiency viruses (HIV)-1 and -2 and simian immunodeficiency virus. Cells susceptible to infection by these viruses can be protected by treatment with the CCR5 ligands regulated on activation, normal T cell expressed and secreted (RANTES), MIP-1alpha, and MIP-1beta. A major component of the mechanism through which chemokines protect cells from HIV infection is by inducing endocytosis of the chemokine receptor. Aminooxypentane (AOP)-RANTES, an NH(2) terminal modified form of RANTES, is a potent inhibitor of infection by R5 HIV strains. AOP-RANTES efficiently downmodulates the cell surface expression of CCR5 and, in contrast with RANTES, appears to prevent recycling of CCR5 to the cell surface. Here, we investigate the cellular basis of this effect. Using CHO cells expressing human CCR5, we show that both RANTES and AOP-RANTES induce rapid internalization of CCR5. In the absence of ligand, CCR5 shows constitutive turnover with a half-time of 6-9 h. Addition of RANTES or AOP-RANTES has little effect on the rate of CCR5 turnover. Immunofluorescence and immunoelectron microscopy show that most of the CCR5 internalized after RANTES or AOP-RANTES treatment accumulates in small membrane-bound vesicles and tubules clustered in the perinuclear region of the cell. Colocalization with transferrin receptors in the same clusters of vesicles indicates that CCR5 accumulates in recycling endosomes. After the removal of RANTES, internalized CCR5 recycles to the cell surface and is sensitive to further rounds of RANTES-induced endocytosis. In contrast, after the removal of AOP-RANTES, most CCR5 remains intracellular. We show that these CCR5 molecules do recycle to the cell surface, with kinetics equivalent to those of receptors in RANTES-treated cells. However, these recycled CCR5 molecules are rapidly reinternalized. Our results indicate that AOP-RANTES induced changes in CCR5 alter the steady-state distribution of the receptor and provide the first evidence for G protein-coupled receptor trafficking through the recycling endosome compartment. PMID- 11121443 TI - p53 regulates myogenesis by triggering the differentiation activity of pRb. AB - The p53 oncosuppressor protein regulates cell cycle checkpoints and apoptosis, but increasing evidence also indicates its involvement in differentiation and development. We had previously demonstrated that in the presence of differentiation-promoting stimuli, p53-defective myoblasts exit from the cell cycle but do not differentiate into myocytes and myotubes. To identify the pathways through which p53 contributes to skeletal muscle differentiation, we have analyzed the expression of a series of genes regulated during myogenesis in parental and dominant-negative p53 (dnp53)-expressing C2C12 myoblasts. We found that in dnp53-expressing C2C12 cells, as well as in p53(-/-) primary myoblasts, pRb is hypophosphorylated and proliferation stops. However, these cells do not upregulate pRb and have reduced MyoD activity. The transduction of exogenous TP53 or Rb genes in p53-defective myoblasts rescues MyoD activity and differentiation potential. Additionally, in vivo studies on the Rb promoter demonstrate that p53 regulates the Rb gene expression at transcriptional level through a p53-binding site. Therefore, here we show that p53 regulates myoblast differentiation by means of pRb without affecting its cell cycle-related functions. PMID- 11121444 TI - Differential roles for alpha(M)beta(2) integrin clustering or activation in the control of apoptosis via regulation of akt and ERK survival mechanisms. AB - The role of integrins in leukocyte apoptosis is unclear, some studies suggest enhancement, others inhibition. We have found that beta(2)-integrin engagement on neutrophils can either inhibit or enhance apoptosis depending on the activation state of the integrin and the presence of proapoptotic stimuli. Both clustering and activation of alpha(M)beta(2) delays spontaneous, or unstimulated, apoptosis, maintains mitochondrial membrane potential, and prevents cytochrome c release. In contrast, in the presence of proapoptotic stimuli, such as Fas ligation, TNFalpha, or UV irradiation, ligation of active alpha(M)beta(2) resulted in enhanced mitochondrial changes and apoptosis. Clustering of inactive integrins did not show this proapoptotic effect and continued to inhibit apoptosis. This discrepancy was attributed to differential signaling in response to integrin clustering versus activation. Clustered, inactive alpha(M)beta(2) was capable of stimulating the kinases ERK and Akt. Activated alpha(M)beta(2) stimulated Akt, but not ERK. When proapoptotic stimuli were combined with either alpha(M)beta(2) clustering or activation, Akt activity was blocked, allowing integrin activation to enhance apoptosis. Clustered, inactive alpha(M)beta(2) continued to inhibit stimulated apoptosis due to maintained ERK activity. Therefore, beta(2)-integrin engagement can both delay and enhance apoptosis in the same cell, suggesting that integrins can play a dual role in the apoptotic progression of leukocytes. PMID- 11121445 TI - Expression profiling in the muscular dystrophies: identification of novel aspects of molecular pathophysiology. AB - We used expression profiling to define the pathophysiological cascades involved in the progression of two muscular dystrophies with known primary biochemical defects, dystrophin deficiency (Duchenne muscular dystrophy) and alpha sarcoglycan deficiency (a dystrophin-associated protein). We employed a novel protocol for expression profiling in human tissues using mixed samples of multiple patients and iterative comparisons of duplicate datasets. We found evidence for both incomplete differentiation of patient muscle, and for dedifferentiation of myofibers to alternative lineages with advancing age. One developmentally regulated gene characterized in detail, alpha-cardiac actin, showed abnormal persistent expression after birth in 60% of Duchenne dystrophy myofibers. The majority of myofibers ( approximately 80%) remained strongly positive for this protein throughout the course of the disease. Other developmentally regulated genes that showed widespread overexpression in these muscular dystrophies included embryonic myosin heavy chain, versican, acetylcholine receptor alpha-1, secreted protein, acidic and rich in cysteine/osteonectin, and thrombospondin 4. We hypothesize that the abnormal Ca(2)+ influx in dystrophin- and alpha-sarcoglycan-deficient myofibers leads to altered developmental programming of developing and regenerating myofibers. The finding of upregulation of HLA-DR and factor XIIIa led to the novel identification of activated dendritic cell infiltration in dystrophic muscle; these cells mediate immune responses and likely induce microenvironmental changes in muscle. We also document a general metabolic crisis in dystrophic muscle, with large scale downregulation of nuclear-encoded mitochondrial gene expression. Finally, our expression profiling results show that primary genetic defects can be identified by a reduction in the corresponding RNA. PMID- 11121446 TI - The cortical protein Num1p is essential for dynein-dependent interactions of microtubules with the cortex. AB - In budding yeast, the mitotic spindle moves into the neck between the mother and bud via dynein-dependent sliding of cytoplasmic microtubules along the cortex of the bud. How dynein and microtubules interact with the cortex is unknown. We found that cells lacking Num1p failed to exhibit dynein-dependent microtubule sliding in the bud, resulting in defective mitotic spindle movement and nuclear segregation. Num1p localized to the bud cortex, and that localization was independent of microtubules, dynein, or dynactin. These data are consistent with Num1p being an essential element of the cortical attachment mechanism for dynein dependent sliding of microtubules in the bud. PMID- 11121447 TI - Numb is an endocytic protein. AB - Numb is a protein that in Drosophila determines cell fate as a result of its asymmetric partitioning at mitosis. The function of Numb has been linked to its ability to bind and to biologically antagonize Notch, a membrane receptor that also specifies cell fate. The biochemical mechanisms underlying the action of Numb, however, are still largely unknown. The wide pattern of expression of Numb suggests a general function in cellular homeostasis that could be additional to, or part of, its action in fate determination. Such a function could be endocytosis, as suggested by the interaction of Numb with Eps15, a component of the endocytic machinery. Here, we demonstrate that Numb is an endocytic protein. We found that Numb localizes to endocytic organelles and is cotrafficked with internalizing receptors. Moreover, it associates with the appendage domain of alpha adaptin, a subunit of AP2, a major component of clathrin-coated pits. Finally, fragments of Numb act as dominant negatives on both constitutive and ligand-regulated receptor-mediated internalization, suggesting a general role for Numb in the endocytic process. PMID- 11121449 TI - Hutchinson Smoking Prevention Project: a new gold standard in prevention science requires new transdisciplinary thinking. PMID- 11121450 TI - Gamma-tocopherol: a new player in prostate cancer prevention? PMID- 11121451 TI - Trial data lacking for elderly patients with breast cancer. PMID- 11121453 TI - Effect of mifepristone approval on research remains to be seen. PMID- 11121455 TI - Stem cell transplant numbers decline; research continues. PMID- 11121456 TI - Why don't people heed risk warnings from science? PMID- 11121458 TI - White House commission examines research on complementary cancer therapies. PMID- 11121459 TI - Can we predict response to 5-FU? PMID- 11121460 TI - Hutchinson Smoking Prevention Project: long-term randomized trial in school-based tobacco use prevention--results on smoking. AB - BACKGROUND: No long-term impact has yet been observed with the use of the social influences approach to school-based smoking prevention for youth. However, whether this lack of impact is due to methodologic problems with the studies or to the failure of the interventions is unclear. The Hutchinson Smoking Prevention Project (HSPP), conducted from September 1984 through August 1999, aimed to attain the most rigorous randomized trial possible to determine the long-term impact of a theory-based, social-influences, grade 3-12 intervention on smoking prevalence among youth. METHODS: Forty Washington school districts were randomly assigned to the intervention or to the control condition. Study participants were children enrolled in two consecutive 3rd grades in the 40 districts (n = 8388); they were followed to 2 years after high school. The trial achieved high implementation fidelity and 94% follow-up. Data were analyzed with the use of group-permutation methods, and all statistical tests were two-sided. RESULTS: No significant difference in prevalence of daily smoking was found between students in the control and experimental districts, either at grade 12 (difference [Delta] = 0.2%, 95% confidence interval [CI] = -4.6% to 4.4%, and P =.91 for girls; Delta = 0.3%, 95% CI = -5.0% to 5.5%, and P =.89 for boys) or at 2 years after high school (Delta = -1.4%, 95% CI = -5.0% to 1.6%, and P =.38 for girls; Delta = 2.6%, 95% CI = -2.5% to 7.7%, and P =.30 for boys). Moreover, no intervention impact was observed for other smoking outcomes, such as extent of current smoking or cumulative amount smoked, or in subgroups that differ in a priori specified variables, such as family risk for smoking. CONCLUSION: The rigor of the HSPP trial suggests high credence for the intervention impact results. Consistent with previous trials, there is no evidence from this trial that a school-based social influences approach is effective in the long-term deterrence of smoking among youth. PMID- 11121461 TI - HER2 and choice of adjuvant chemotherapy for invasive breast cancer: National Surgical Adjuvant Breast and Bowel Project Protocol B-15. AB - BACKGROUND: Recent retrospective analyses have suggested that breast cancer patients whose tumors overexpress HER2 derive preferential benefit from treatment with anthracyclines such as doxorubicin. This has led some clinicians to propose that HER2 should be used as a predictive marker in choosing between anthracycline based regimens and combination chemotherapy with cyclophosphamide, methotrexate, and 5-fluorouracil (CMF). We evaluated this recommendation in a retrospective study of National Surgical Adjuvant Breast and Bowel Project Protocol B-15, in which patients received a combination of doxorubicin and cyclophosphamide (AC), CMF, or AC followed by CMF. We hypothesized that AC would be superior to CMF only in the HER2-positive patients. METHODS: Immunohistochemical detection of HER2 was performed on tumor sections from 2034 of 2295 eligible patients. We used statistical analysis to evaluate the interaction between the efficacy of the assigned treatments and HER2 overexpression. All statistical tests were two sided. RESULTS: Tumor sections from 599 patients (29%) stained positive for HER2. AC was superior to CMF in HER2-positive patients only, although differences in outcomes did not reach statistical significance. In the HER2-positive cohort, relative risks of failure (i.e., after AC treatment as compared with CMF treatment) were 0.84 for disease-free survival (DFS) (95% confidence interval [CI] = 0.65--1.07; P =.15), 0.82 for survival (95% CI = 0.63--1.06; P =.14), and 0.80 for recurrence-free survival (RFS) (95% CI = 0.62--1.04; P =.10). Tests for interaction between treatment and HER2 status were suggestive but not statistically significant (P =.19 for DFS, P =.11 for survival, and P =.08 for RFS). CONCLUSIONS: These results, together with overview results indicating minor overall superiority for anthracycline-based regimens relative to CMF, indicate a preference for the AC regimen in patients with HER2-positive tumors. Both AC and CMF regimens may be considered for patients with HER2-negative tumors. PMID- 11121462 TI - Protection of normal proliferating cells against chemotherapy by staurosporine mediated, selective, and reversible G(1) arrest. AB - BACKGROUND: A major limiting factor in human cancer chemotherapy is toxicity in normal tissues. Our goal was to determine whether normal proliferating cells could be protected from chemotherapeutic agents by taking advantage of the differential drug sensitivity of cell cycle G(1) checkpoint in normal and cancer cells. METHODS: Normal mammary epithelial cells and mammary cancer cells were initially treated with staurosporine at a cytostatic (i.e., nonlethal) concentration, which preferentially arrests normal cells in the G(0)/G(1) phase of the cell cycle without affecting the proliferation of tumor cells. After the selective arrest of normal cells in G(0)/G(1), both normal and tumor cells were treated with doxorubicin or camptothecin, two cytotoxic (i.e., lethal) chemotherapeutic agents. Cells were then allowed to recover in drug-free medium for 12 days. RESULTS: After pretreatment of both normal and tumor cells with staurosporine followed by treatment with doxorubicin or camptothecin, tumor cells were selectively killed by chemotherapeutic agents, whereas normal cells resumed proliferation after the drugs were removed. Pretreatment with staurosporine also protected normal circulating lymphocytes that had been induced to proliferate in vitro with phytohemagglutinin from chemotherapeutic agents. Staurosporine-induced arrest of normal cells in G(0)/G(1) phase was reversible, and arrested cells tolerated doses of camptothecin that were more than 100-fold higher than necessary to eradicate all tumor cells in culture. Staurosporine-mediated G(0)/G(1) arrest targets the retinoblastoma protein (pRb) pathway and was accompanied by a rapid decrease in cyclin-dependent kinase (CDK) 4 protein levels, increased binding of CDK inhibitors p21 and p27 to CDK2, and inhibition of CDK2 activity in normal cells. CONCLUSIONS: Breast cancer cells with defective checkpoints regulated by the pRb pathway can be targeted specifically with chemotherapeutic agents, following staurosporine-mediated, selective and reversible G(0)/G(1) arrest in normal cells. PMID- 11121463 TI - Racial variation in prostate cancer incidence and in hormonal system markers among male health professionals. AB - BACKGROUND: Racial variation in prostate cancer incidence in the United States is pronounced, with African-American men having the highest rates. Whether differences in the distribution of known or suspected risk factors among racial groups explain this variation is unknown. METHODS: We evaluated prospectively the relation between prostate cancer and race among 45 410 U.S. male health professionals aged 40--75 years in 1986. We used multivariable, pooled logistic regression to adjust the rate ratio (RR) for potential dietary and lifestyle risk factors. We also measured circulating levels of steroid hormones, sex hormone binding globulin, and vitamin D metabolites and length of the androgen receptor gene CAG repeat in a sample of African-American (n = 43), Asian (n = 52), and white (n = 55) participants and assessed variation by race in these possible prostate epithelial cell growth mediators by use of analysis of variance. Statistical tests were two-sided. RESULTS: The age-adjusted RR for prostate cancer was 1.73 (95% confidence interval [CI] = 1.23--2.45) for African-American men compared with white men. After multivariate adjustment, the RR increased to 1.81 (95% CI = 1.27--2.58). The rate of prostate cancer did not differ between Asians and whites. Steroid hormone and 1,25-dihydroxyvitamin D levels did not vary appreciably by race. However, the mean number of androgen receptor gene CAG repeats was lower among African-Americans (mean +/- standard deviation = 20.1 +/- 3.5) than among whites (22.1 +/- 3.1; P =.007) and Asians (22.1 +/- 3.9; P =.009). CONCLUSIONS: Our results confirm the elevated incidence of prostate cancer among African Americans and show that it is not explained by differences in the distribution of possible dietary and lifestyle risk factors in this cohort. Racial variation in length of the androgen receptor gene CAG repeat may explain a small part of the excess risk of prostate cancer among African-American men in this cohort. PMID- 11121464 TI - Association between alpha-tocopherol, gamma-tocopherol, selenium, and subsequent prostate cancer. AB - BACKGROUND: Selenium and alpha-tocopherol, the major form of vitamin E in supplements, appear to have a protective effect against prostate cancer. However, little attention has been paid to the possible role of gamma-tocopherol, a major component of vitamin E in the U.S. diet and the second most common tocopherol in human serum. A nested case-control study was conducted to examine the associations of alpha-tocopherol, gamma-tocopherol, and selenium with incident prostate cancer. METHODS: In 1989, a total of 10,456 male residents of Washington County, MD, donated blood for a specimen bank. A total of 117 of 145 men who developed prostate cancer and 233 matched control subjects had toenail and plasma samples available for assays of selenium, alpha-tocopherol, and gamma-tocopherol. The association between the micronutrient concentrations and the development of prostate cancer was assessed by conditional logistic regression analysis. All statistical tests were two-sided. RESULTS: The risk of prostate cancer declined, but not linearly, with increasing concentrations of alpha-tocopherol (odds ratio (highest versus lowest fifth) = 0.65; 95% confidence interval = 0.32--1.32; P(trend) =.28). For gamma-tocopherol, men in the highest fifth of the distribution had a fivefold reduction in the risk of developing prostate cancer than men in the lowest fifth (P:(trend) =.002). The association between selenium and prostate cancer risk was in the protective direction with individuals in the top four fifths of the distribution having a reduced risk of prostate cancer compared with individuals in the bottom fifth (P(trend) =.27). Statistically significant protective associations for high levels of selenium and alpha tocopherol were observed only when gamma-tocopherol concentrations were high. CONCLUSIONS: The use of combined alpha- and gamma- tocopherol supplements should be considered in upcoming prostate cancer prevention trials, given the observed interaction between alpha-tocopherol, gamma-tocopherol, and selenium. PMID- 11121465 TI - Androgen-receptor gene CAG repeats, plasma testosterone levels, and risk of hepatitis B-related hepatocellular carcinoma. AB - BACKGROUND: Worldwide, hepatocellular carcinoma (HCC) is more prevalent in men than in women, suggesting that sex hormones and/or X-chromosome-linked genes may be involved in hepatocarcinogenesis. We investigated the association of a trinucleotide (CAG) repeat in the androgen receptor (AR) gene (located on the X chromosome) termed "AR-CAG repeats," levels of plasma testosterone, and the risk of HCC in Taiwanese men. Chronic hepatitis B virus (HBV) infection, which is associated with risk of HCC, is hyperendemic in Taiwan. METHODS: We compared the number of AR-CAG repeats in 285 HBV carriers with HCC and in 349 HBV carriers without HCC. We also conducted a nested case--control study on participants in a cohort study. Blood was collected prospectively from 110 case patients and 239 control subjects and was used to determine the number of AR-CAG repeats and plasma testosterone level. All statistical tests were two-sided. RESULTS: The overall odds ratio (OR) for HCC was 1.72 (95% confidence interval [CI] = 1.03- 2.89) for HBV carriers with 20 or fewer AR-CAG repeats compared with those with more than 24 repeats. This association was observed only in patients with late onset HCC (OR = 2.37; 95% CI = 1.28--4.38). In the nested case-control study, HBV carriers in the highest tertile of testosterone levels had a statistically significantly increased risk of HCC (OR = 2.06; 95% CI = 1.14--3.70) compared with those in the lowest tertile. Elevated testosterone was more strongly associated with early-onset (OR = 4.67; 95% CI = 1.41--15.38) than late-onset disease. HBV carriers with 20 or fewer AR-CAG repeats and higher testosterone levels had a fourfold increase in HCC risk compared with those with more than 24 repeats and testosterone levels in the lowest tertile. CONCLUSIONS: Higher levels of androgen signaling, reflected by higher testosterone levels and 20 or fewer AR CAG repeats, may be associated with an increased risk of HBV-related HCC in men. PMID- 11121466 TI - Diffusion magnetic resonance imaging: an early surrogate marker of therapeutic efficacy in brain tumors. AB - BACKGROUND: A surrogate marker for treatment response that can be observed earlier than comparison of sequential magnetic resonance imaging (MRI) scans, which depends on relatively slow changes in tumor volume, may improve survival of brain tumor patients by providing more time for secondary therapeutic interventions. Previous studies in animals with the use of diffusion MRI revealed rapid changes in tumor water diffusion values after successful therapeutic intervention. METHODS: The present study examined the sensitivity of diffusion MRI measurements in orthotopic rat brain tumors derived from implanted rat 9L glioma cells. The effectiveness of therapy for individual brain cancer patients was evaluated by measuring changes in tumor volume on neuroimaging studies conducted 6--8 weeks after the conclusion of a treatment cycle. RESULTS: Diffusion MRI could detect water diffusion changes in orthotopic 9L gliomas after doses of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU or carmustine) that resulted in as little as 0.2 log cell kill, a measure of tumor cell death. Mean apparent diffusion coefficients in tumors were found to be correlated with and highly sensitive to changes in tumor cellularity (r =.78; two-sided P =.041). The feasibility of serial diffusion MRI in the clinical management of primary brain tumor patients was also demonstrated. Increased diffusion values could be detected in human brain tumors shortly after treatment initiation. The magnitude of the diffusion changes corresponded with clinical outcome. CONCLUSIONS: These results suggest that diffusion MRI will provide an early surrogate marker for quantification of treatment response in patients with brain tumors. PMID- 11121467 TI - Hereditary retinoblastoma and risk of lung cancer. PMID- 11121468 TI - Re: Combined treatment with buserelin and tamoxifen in premenopausal metastatic breast cancer: a randomized study. PMID- 11121469 TI - Re: Cigar smoking in men and risk of death from tobacco-related cancers. PMID- 11121471 TI - Position-specific effect of ribonucleotides on the cleavage activity of human topoisomerase II. AB - Beyond the normal DNA transactions mediated by topoisomerase II, we have recently demonstrated that the cleavage activity of the two human topoisomerase II isoforms is several-fold stimulated if a ribonucleotide rather than a deoxyribonucleotide is present at the scissile phosphodiester in one strand of the substrate. Here we show that ribonucleotides exert a position-specific effect on topoisomerase II-mediated cleavage without altering the sequence specificity of the enzyme. Ribonucleotides located within the 4 bp cleavage stagger stimulate topoisomerase II-mediated cleavage, whereas ribonucleotides located outside the stagger in general have an inhibitory effect. Results obtained from competition experiments indicate that the position-specific effect of ribonucleotides on topoisomerase II activity is caused by altered substrate interaction. When cleavage is performed with substrates containing one ribonucleotide in both strands or several ribonucleotides in one strand the effect of the individual ribonucleotides on cleavage is not additive. Finally, although topoisomerase II recognizes substrates with longer stretches of ribonucleotides, an RNA/DNA hybrid where one strand is composed entirely of RNA is not cleaved by the enzyme. The positional effect of ribonucleotides on topoisomerase II-mediated cleavage shares many similarities to the positional effect exerted by either abasic sites or base mismatches, demonstrating a general influence of DNA imperfections on topoisomerase II activity. PMID- 11121473 TI - Interaction of human DNA topoisomerase I with G-quartet structures. AB - Because of their role in the control of the topological state of DNA, topoisomerases are ubiquitous and vital enzymes, which participate in nearly all events related to DNA metabolism including replication and transcription. We show here that human topoisomerase I (Topo I) plays an unexpected role of 'molecular matchmaker' for G-quartet formation. G-quadruplexes are multi-stranded structures held together by square planes of four guanines ('G-quartets') interacting by forming Hoogsteen hydrogen bonds. Topo I is able to promote the formation of four stranded intermolecular DNA structures when added to single-stranded DNA containing a stretch of at least five guanines. We provide evidence that these complexes are parallel G-quartet structures, mediated by tetrads of hydrogen bonded guanine. In addition, Topo I binds specifically to pre-formed parallel and anti-parallel G4-DNA. PMID- 11121472 TI - Role of SR protein modular domains in alternative splicing specificity in vivo. AB - The SR proteins constitute a family of nuclear phosphoproteins which are required for constitutive splicing and also influence alternative splicing regulation. They have a modular structure consisting of one or two RNA recognition motifs (RRMs) and a C-terminal domain, rich in arginine and serine residues. The functional role of the different domains of SR proteins in constitutive splicing activity has been extensively studied in vitro; however, their contribution to alternative splicing specificity in vivo has not been clearly established. We sought to address how the modular domains of SR proteins contribute to alternative splicing specificity. The activity of a series of chimeric proteins consisting of domain swaps between different SR proteins showed that splice site selection is determined by the nature of the RRMs and that RRM2 of SF2/ASF has a dominant role and can confer specificity to a heterologous protein. In contrast, the identity of the RS domain is not important, as the RS domains are functionally interchangeable. The contribution of the RRMs to alternative splicing specificity in vivo suggests that sequence-specific RNA binding by SR proteins is required for this activity. PMID- 11121474 TI - The 26S proteasome negatively regulates the level of overall genomic nucleotide excision repair. AB - Regulation of protein expression can be achieved through destruction of proteins by the 26S: proteasome. Cellular processes that are regulated by proteolysis include cell cycle progression, stress responses and differentiation. Several nucleotide excision repair proteins in yeast and humans, such as Rad23, Rad4 and XPB, have been shown to co-purify with Cim3 and Cim5, AAA ATPases of the 19S: proteasome regulatory subunit. However, it has not been determined if nucleotide excision repair is regulated through protein destruction. We measured nucleotide excision repair in yeast mutants that are defective in proteasome function and found that the repair of the transcribed and non-transcribed strands of an RNA polymerase II-transcribed reporter gene was increased in the absence of proteasome function. Additionally, overexpression of the Rad4 repair protein, which is bound to the repair/proteolytic factor Rad23, conferred higher rates of nucleotide excision repair. Based on our data we suggest that a protein (or proteins) involved in nucleotide excision repair or in regulation of repair is degraded by the 26S proteasome. We propose that decreased proteasome function enables increased DNA repair, due to the transient accumulation of a specific repair factor, perhaps Rad4. PMID- 11121475 TI - Characterization of the B lymphocyte-induced maturation protein-1 (Blimp-1) gene, mRNA isoforms and basal promoter. AB - Blimp-1 is a transcriptional repressor that is both required and sufficient to trigger terminal differentiation of B lymphocytes and monocyte/macrophages. Here we report the organization of the mouse Blimp-1 gene, an analysis of Blimp-1 homologs in different species, the characterization of Blimp-1 mRNA isoforms and initial studies on the transcription of Blimp-1. The murine Blimp-1 gene covers approximately 23 kb and contains eight exons. There are Blimp-1 homologs in species evolutionarily distant from mouse (Caenorhabditis elegans and Drosophila melanogaster) but no homolog was found in the unicellular yeast Saccharomyces cerevisiae. The three major Blimp-1 mRNA isoforms result from the use of different polyadenylation sites and do not encode different proteins. Run-on transcription analyses were used to show that the developmentally regulated expression of Blimp-1 mRNA in B cells is determined by transcription initiation. Multiple Blimp-1 transcription initiates sites were mapped near an initiator element and a region conferring basal promoter activity has been identified. PMID- 11121476 TI - Mechanisms of DNA cleavage by copper complexes of 3-clip-phen and of its conjugate with a distamycin analogue. AB - Mechanisms of DNA oxidation by copper complexes of 3-Clip-Phen and its conjugate with a distamycin analogue, in the presence of a reductant and air, were studied. Characterisation of the production of 5-methylenefuranone (5-MF) and furfural, associated with the release of nucleobases, indicated that these copper complexes oxidised the C1' and C5' positions of 2-deoxyribose, respectively, which are accessible from the DNA minor groove. Oxidation at C1' was the major degradation route. Digestion of DNA oxidation products by P1 nuclease and bacterial alkaline phosphatase allowed characterisation of glycolic acid residues, indicating that these copper complexes also induced C4' oxidation. However, this pathway was not associated with base propenal release. The ability of the copper complex of the 3 Clip-Phen conjugate with the distamycin analogue to produce sequence-selective DNA cleavage allowed confirmation of these mechanisms of DNA oxidation by PAGE. Comparison of DNA cleavage activity showed that conjugation of 3-Clip-Phen with a DNA minor groove binder, like the distamycin analogue, decreased both its ability to perform C1' oxidation as well as the initial rate of the reaction, but this conjugate is still active after 5 h at 37 degrees C, making it an efficient DNA cleaver. PMID- 11121477 TI - A novel four zinc-finger protein targeted against p190(BcrAbl) fusion oncogene cDNA: utilisation of zinc-finger recognition codes. AB - A three zinc-finger protein that binds specifically to the cDNA representing the unique fusion gene BCR:Abl, associated with acute lymphoblastic leukaemia, has previously been characterised. At this breakpoint, a sequence homology of 8/9 bp exists between the BCR:Abl (fusion) and c-ABL: (parental) target sequences. We show that the three zinc-finger protein discriminates poorly between the fusion (BCR:Abl) and parental (ABL:) sequence (K:(d)s of 42.8 and 65.1 nM, respectively). In order to improve the discriminatory properties of this protein, and to demonstrate the utility of current zinc-finger databases, we have added a fourth zinc-finger to the original three zinc-finger protein. This fourth finger recognises a 3 bp subsite derived from the BCR: portion of the breakpoint and is not present in c-ABL: This novel four finger protein, which now recognises a 12 bp sequence, demonstrates improved specific binding to BcrAbl (K:(d )= 17 nM). More significantly we have shown that there is now enhanced discrimination between BcrAbl and ABL: sequences by the four finger protein than the original three finger protein. PMID- 11121478 TI - Misincorporation of 2'-deoxyoxanosine into DNA: a molecular basis for NO-induced mutagenesis derived from theoretical calculations. AB - A wide range of theoretical methods, including high level ab initio, density functional, self-consistent reaction field, molecular dynamics and thermodynamic integration calculations, have been used to analyze the mutagenic properties of oxanosine. The major tautomeric forms in the gas phase and aqueous solution have been determined. The ability of oxanosine to recognize thymine and cytosine in the gas phase and in the DNA environment has been compared with that of guanine. A physicochemical explanation for the mutagenic properties of oxanosine is suggested. PMID- 11121479 TI - Rapid microtiter assays for poxvirus topoisomerase, mammalian type IB topoisomerase and HIV-1 integrase: application to inhibitor isolation. AB - We have developed microtiter assays for detecting catalysis by type IB topoisomerases and retroviral integrases. Each assay employs model DNA substrates containing biotin in one strand and digoxigenin in another. In each case action of the enzyme results in the formation of a single DNA strand containing both groups. This allows the reaction product to be quantified by capturing biotinylated product DNA on avidin-coated plates followed by detection using an anti-digoxigenin ELISA. The order of addition of reactants and inhibitors can be varied to distinguish effects of test compounds on different steps in the reaction. These assays were used to screen compound libraries for inhibitors active against mammalian topoisomerase or HIV integrase. We identified (-) epigallocatechin 3-O:-gallate, as a potent inhibitor of religation by mammalian topoisomerase (IC(50) of 26 nM), potentially explaining the anti-cancer properties previously attributed to this compound. New integrase inhibitors were also identified. A similar strategy may be used to develop microtiter assays for many further DNA modifying enzymes. PMID- 11121480 TI - Local conformational variations observed in B-DNA crystals do not improve base stacking: computational analysis of base stacking in a d(CATGGGCCCATG)(2) B<-->A intermediate crystal structure. AB - The crystal structure of d(CATGGGCCCATG)(2) shows unique stacking patterns of a stable B<-->A-DNA intermediate. We evaluated intrinsic base stacking energies in this crystal structure using an ab initio quantum mechanical method. We found that all crystal base pair steps have stacking energies close to their values in the standard and crystal B-DNA geometries. Thus, naturally occurring stacking geometries were essentially isoenergetic while individual base pair steps differed substantially in the balance of intra-strand and inter-strand stacking terms. Also, relative dispersion, electrostatic and polarization contributions to the stability of different base pair steps were very sensitive to base composition and sequence context. A large stacking flexibility is most apparent for the CpA step, while the GpG step is characterized by weak intra-strand stacking. Hydration effects were estimated using the Langevin dipoles solvation model. These calculations showed that an aqueous environment efficiently compensates for electrostatic stacking contributions. Finally, we have carried out explicit solvent molecular dynamics simulation of the d(CATGGGCCCATG)(2) duplex in water. Here the DNA conformation did not retain the initial crystal geometry, but moved from the B<-->A intermediate towards the B-DNA structure. The base stacking energy improved in the course of this simulation. Our findings indicate that intrinsic base stacking interactions are not sufficient to stabilize the local conformational variations in crystals. PMID- 11121481 TI - Differential requirements for cis and trans V(D)J cleavage: effects of substrate length. AB - The assembly of productive synaptic complexes is a critical, but poorly understood, regulatory step in V(D)J recombination. Several lines of evidence suggest that there may be important differences between recombination involving sites situated in cis (on the same DNA molecule) and in trans (on separate molecules). Because biochemical experiments using both purified RAG proteins and crude extracts have failed to detect trans cleavage of plasmid substrates it has been thought that there is a substantial bias against trans synapsis. In conflict with these results are more recent studies showing that purified RAG proteins can catalyze trans cleavage of short oligonucleotide substrates. Furthermore, recent experiments have detected efficient trans cleavage of plasmid substrates in vivo. We sought to investigate why these different systems yield such divergent results. We found that, unexpectedly, the ability of both purified RAG proteins and crude extracts to cleave DNA substrates in trans is a function of substrate length. Our data raise two critical issues: first, oligonucleotides, which are the most commonly used substrates to study V(D)J recombination in vitro, do not mimic the behavior of plasmid substrates; second, in the trans cleavage reaction current purified RAG systems do not accurately reflect the in vivo situation. We propose a unifying model to explain the effects of substrate length and coniguration (cis or trans) on the efficiency of synapsis. PMID- 11121482 TI - Adenine excisional repair function of MYH protein on the adenine:8-hydroxyguanine base pair in double-stranded DNA. AB - Adenine paired with 8-hydroxyguanine (oh(8)G), a major component of oxidative DNA damage, is excised by MYH base excision repair protein in human cells. Since repair activity of MYH protein on an A:G mismatch has also been reported, we compared the repair activity of His(6)-tagged MYH proteins, expressed in Spodoptera frugiperda Sf21 cells, on A:oh(8)G and A:G mismatches by DNA cleavage assay and gel mobility shift assay. We also compared the repair ability of type 1 mitochondrial protein with type 2 nuclear protein, as well as of polymorphic type 1-Q(324) and 2-Q(310) proteins with type 1-H(324) and 2-H(310) proteins by DNA cleavage assay and complementation assay of an Escherichia coli mutM mutY strain. In a reaction buffer with a low salt (0-50 mM) concentration, adenine DNA glycosylase activity of type 2 protein was detected on both A:oh(8)G and A:G substrates. However, in a reaction buffer with a 150 mM salt concentration, similar to physiological conditions, the glycosylase activity on A:G, but not on A:oh(8)G, was extremely reduced and the binding activity of type 2 protein for A:G, but not for A:oh(8)G, was proportionally reduced. The glycosylase activity on A:oh(8)G and the ability to suppress spontaneous mutagenesis were greater for type 2 than type 1 enzyme. There was apparently no difference in the repair activities between the two types of polymorphic MYH proteins. These results indicate that human MYH protein specifically catalyzes the glycosylase reaction on A:oh(8)G under physiological salt concentrations. PMID- 11121483 TI - The SP1 sites of the human apoCIII enhancer are essential for the expression of the apoCIII gene and contribute to the hepatic and intestinal expression of the apoA-I gene in transgenic mice. AB - We have generated transgenic mice carrying wild-type and mutant forms of the apolipoprotein (apo)A-I/apoCIII gene cluster. Mutations were introduced either in one or in three SP1 binding sites of the apoCIII enhancer. In mice carrying the wild-type transgene, major sites of apoA-I mRNA synthesis were liver and intestine and minor sites were kidney and, to a lesser extent, other tissues. The major site of chloramphenicol acetyl transferase (CAT) activity (used as a reporter for the apoCIII gene) was liver and minor sites intestine and kidney. A mutation in one SP1 binding site reduced the expression of the apoA-I gene to approximately 23 and 19% in the liver and intestine, respectively, as compared to the control wild-type. The hepatic expression of the CAT gene was not affected whereas the intestinal expression was nearly abolished. Mutations in three SP1 binding sites reduced the hepatic and intestinal expression of the apoA-I and CAT genes to 14 and 4%, respectively, as compared to the wild-type control, and abolished CAT expression in all tissues. The findings suggest that the SP1 sites of the apoCIII enhancer are required for the expression of the apoCIII gene and also contribute significantly to the hepatic and intestinal expression of the apoA-I gene in vivo. PMID- 11121484 TI - Alleviating transcript insufficiency caused by Friedreich's ataxia triplet repeats. AB - Expanded GAA.TTC trinucleotide repeats in intron 1 of the frataxin gene cause Friedreich's ataxia (FRDA) by reducing frataxin mRNA levels. Insufficient frataxin, a nuclear encoded mitochondrial protein, leads to the progressive neurodegeneration and cardiomyopathy characteristic of FRDA. Previously we demonstrated that long GAA.TTC tracts impede transcription elongation in vitro and provided evidence that the impediment results from an intramolecular purine.purine.pyrimidine DNA triplex formed behind an advancing RNA polymerase. Our model predicts that inhibiting formation of this triplex during transcription will increase successful elongation through GAA.TTC tracts. Here we show that this is the case. Oligodeoxyribonucleotides designed to block particular types of triplex formation provide specific and concentration-dependent increases in full length transcript. In principle, therapeutic agents that selectively interfere with triplex formation could alleviate the frataxin transcript insufficiency caused by pathogenic FRDA alleles. PMID- 11121485 TI - Improved statistical methods reveal direct interactions between 16S and 23S rRNA. AB - Recent biochemical studies have indicated a number of regions in both the 16S and 23S rRNA that are exposed on the ribosomal subunit surface. In order to predict potential interactions between these regions we applied novel phylogenetically based statistical methods to detect correlated nucleotide changes occurring between the rRNA molecules. With these methods we discovered a number of highly significant correlated changes between different sets of nucleotides in the two ribosomal subunits. The predictions with the highest correlation values belong to regions of the rRNA subunits that are in close proximity according to recent crystal structures of the entire ribosome. We also applied a new statistical method of detecting base triple interactions within these same rRNA subunit regions. This base triple statistic predicted a number of new base triples not detected by pair-wise interaction statistics within the rRNA molecules. Our results suggest that these statistical methods may enhance the ability to detect novel structural elements both within and between RNA molecules. PMID- 11121486 TI - Molecular dynamics studies of the HIV-1 TAR and its complex with argininamide. AB - The dynamic behavior of HIV-1 TAR and its complex with argininamide is investigated by means of molecular dynamics simulations starting from NMR structures, with explicit inclusion of water and periodic boundary conditions particle mesh Ewald representation of the electrostatic energy. During simulations of free and argininamide-bound TAR, local structural patterns, as determined by NMR experiments, were reproduced. An interdomain motion was observed in the simulations of free TAR, which is absent in the case of bound TAR, leading to the conclusion that the free conformation of TAR is intrinsically more flexible than the bound conformation. In particular, in the bound conformation the TAR-argininamide interface is very well ordered, as a result of the formation of a U.A.U base triple, which imposes structural constraints on the global conformation of the molecule. Free energy analysis, which includes solvation contributions, was used to evaluate the influence of van der Waals and electrostatic terms on formation of the complex and on the conformational rearrangement from free to bound TAR. PMID- 11121487 TI - A role for MHR1, a gene required for mitochondrial genetic recombination, in the repair of damage spontaneously introduced in yeast mtDNA. AB - A nuclear recessive mutant in Saccharomyces cerevisiae, mhr1-1, is defective in mitochondrial genetic recombination at 30 degrees C and shows extensive vegetative petite induction by UV irradiation at 30 degrees C or when cultivated at a higher temperature (37 degrees C). It has been postulated that mitochondrial DNA (mtDNA) is oxidatively damaged by by-products of oxidative respiration. Since genetic recombination plays a critical role in DNA repair in various organisms, we tested the possibility that MHR1 plays a role in the repair of oxidatively damaged mtDNA using an enzyme assay. mtDNA isolated from cells grown under standard (aerobic) conditions contained a much higher level of DNA lesions compared with mtDNA isolated from anaerobically grown cells. Soon after a temperature shift from 30 to 37 degrees C the number of mtDNA lesions increased 2 fold in mhr1-1 mutant cells but not in MHR1 cells. Malonic acid, which decreased the oxidative stress in mitochondria, partially suppressed both petite induction and the temperature-induced increase in the amount of mtDNA damage in mhr1-1 cells at 37 degrees C. Thus, functional mitochondria require active MHR1, which keeps the extent of spontaneous oxidative damage in mtDNA within a tolerable level. These observations are consistent with MHR1 having a possible role in mtDNA repair. PMID- 11121488 TI - Crystal structures of Mycobacterium tuberculosis RecA and its complex with ADP AlF(4): implications for decreased ATPase activity and molecular aggregation. AB - Sequencing of the complete genome of Mycobacterium tuberculosis, combined with the rapidly increasing need to improve tuberculosis management through better drugs and vaccines, has initiated extensive research on several key proteins from the pathogen. RecA, a ubiquitous multifunctional protein, is a key component of the processes of homologous genetic recombination and DNA repair. Structural knowledge of MtRecA is imperative for a full understanding of both these activities and any ensuing application. The crystal structure of MtRecA, presented here, has six molecules in the unit cell forming a 6(1) helical filament with a deep groove capable of binding DNA. The observed weakening in the higher order aggregation of filaments into bundles may have implications for recombination in mycobacteria. The structure of the complex reveals the atomic interactions of ADP-AlF(4), an ATP analogue, with the P-loop-containing binding pocket. The structures explain reduced levels of interactions of MtRecA with ATP, despite sharing the same fold, topology and high sequence similarity with EcRecA. The formation of a helical filament with a deep groove appears to be an inherent property of MtRecA. The histidine in loop L1 appears to be positioned appropriately for DNA interaction. PMID- 11121490 TI - CHOP gene expression in response to endoplasmic-reticular stress requires NFY interaction with different domains of a conserved DNA-binding element. AB - The transcription factor CHOP/GADD153 gene is induced by cellular stress and is involved in mediating apoptosis. We report the identification of a conserved region in the promoter of the CHOP gene responsible for its inducibility by endoplasmic reticulum (ER) stress. Deletion mutants of the human CHOP promoter identify a region comprising nucleotides -75 to -104 required for both constitutive and ER-stress-inducible expression. This region of the promoter, the ER-stress element (ERSE) is sufficient to confer both increased basal activity and ER-stress inducibility to an otherwise inactive heterologous promoter. The CHOP ERSE is a novel variant of the ERSE as it contains two different functional domains, and a GA- instead of GC-rich intervening sequence. The CCAAT-box domain occupied by the constitutive transcriptional activator nuclear factor Y (NFY) is required for constitutive activation whereas the variant GCACG 'inducible' domain uniquely mediates ER-stress inducibility. By UV-crosslinking analysis NFY makes contact not only with the constitutive activator CCAAT box but also with the inducible GCACG domain. Deletions and nucleotide substitutions in the CCAAT box as well as its replacement by an SP1 site failed to support ER inducibility. These findings support the notion that NFY is not only required for constitutive activation of CHOP gene transcription, but is also an active and essential element for the assembly of an ER-stress-inducible enhanceosome that activates CHOP gene expression in response to cellular stress. PMID- 11121489 TI - Comparison of the Escherichia coli K-12 genome with sampled genomes of a Klebsiella pneumoniae and three salmonella enterica serovars, Typhimurium, Typhi and Paratyphi. AB - The Escherichia coli K-12 genome (ECO) was compared with the sampled genomes of the sibling species Salmonella enterica serovars Typhimurium, Typhi and Paratyphi A (collectively referred to as SAL) and the genome of the close outgroup Klebsiella pneumoniae (KPN). There are at least 160 locations where sequences of >400 bp are absent from ECO but present in the genomes of all three SAL and 394 locations where sequences are present in ECO but close homologs are absent in all SAL genomes. The 394 sequences in ECO that do not occur in SAL contain 1350 (30.6%) of the 4405 ECO genes. Of these, 1165 are missing from both SAL and KPN. Most of the 1165 genes are concentrated within 28 regions of 10-40 kb, which consist almost exclusively of such genes. Among these regions were six that included previously identified cryptic phage. A hypothetical ancestral state of genomic regions that differ between ECO and SAL can be inferred in some cases by reference to the genome structure in KPN and the more distant relative Yersinia pestis. However, many changes between ECO and SAL are concentrated in regions where all four genera have a different structure. The rate of gene insertion and deletion is sufficiently high in these regions that the ancestral state of the ECO/SAL lineage cannot be inferred from the present data. The sequencing of other closely related genomes, such as S.bongori or Citrobacter, may help in this regard. PMID- 11121491 TI - Quantitative analysis of globin gene induction in single human erythroleukemic cells. AB - The mechanisms involved in the normal developmental regulation of globin gene expression, and the response to pharmacological agents that elevate fetal hemoglobin, may be expected to involve either changes in each cell or a selection process affecting subsets of differentiating erythroid cells. To study these mechanisms we have developed assays to measure mRNA levels in single erythroid cells. The assay involved the use of globin-specific probes, with no detectable cross-reactivity, in real-time, fluorescence-based quantitative PCR (Q-PCR). We had previously used this Q-PCR method to measure globin mRNA levels in cultures of primary erythroid cells demonstrating that drugs like hydroxyurea, 5 azacytidine and butyric acid each yielded increases in gamma/( gamma + ss) mRNA ratios, with differential effects on ss-globin levels. We have now extended this approach to measure globin mRNA levels in single K562 cells, a human erythroleukemic cell line, with and without 30 microM hemin treatment. Hemin exposure increases total hemoglobin levels by approximately 9-fold and total alpha-, epsilon- and gamma-globin mRNA levels by 1.5-2.3-fold. Single cell analyses showed initial wide distributions of each of the three individual globin mRNA levels with most cells having detectable but very low levels of each globin transcript. Hemin induction shifted the distributions to higher levels, with a tendency to residual left skewing as some cells remained with very low expression levels despite the effect of hemin in increasing expression in most of these low expressing cells. Thus transcriptional heterogeneity remains a crucial variable, even in this extensively used model of human erythroid biology, and clearly influences strongly the response to inducing agents. These methods may enable us to define better possible molecular and/or cellular models of globin gene modulation. PMID- 11121492 TI - TE-AFLP: combining rapidity and robustness in DNA fingerprinting. AB - A new type of fingerprinting technique is presented, based on amplified fragment length polymorphism (AFLP). Rather than two endonucleases as in AFLP, we propose the use of three enzymes, hence the method is called three endonuclease (TE) AFLP. Genomic DNA is digested and two sets of adapters are selectively ligated onto the restriction fragments in a single reaction volume. No adapters complementary to the ends generated by a frequent cutter are added. Due to the addition of a third endonuclease, the TE-AFLP method provides a high discriminatory power and a reduction in the number of bands. The latter makes it especially suitable for the analysis of complex genomes. TE-AFLP fingerprints are suitable for detection by automatic fluorescent sequencers and are obtained in less than half the time and at reduced costs compared to a typical AFLP. The reliability of this method was investigated by determining the influence of varying digestion, ligation and PCR components on the fingerprint. Moreover, cross-experiments to study inheritance of loci were performed with a primitive insect and with tomato strains. The features of TE-AFLP are discussed in comparison with conventional AFLP. PMID- 11121493 TI - Non-gridded library: a new approach for BAC (bacterial artificial chromosome) exploitation in hexaploid wheat (Triticum aestivum). AB - The feasibility of exploiting non-gridded bacterial artificial chromosome (BAC) libraries and some major factors affecting the efficiency of handling such libraries were studied in hexaploid wheat. Even for a bacterial culture containing only 55% recombinants, some 2000 BAC clones with inserts ranging from 45 to 245 kb could be pooled. The pooled BAC clones could be amplified by culturing for up to 6 h without losing any target clones. These results imply that even for hexaploid wheat, which has an extremely large genome, some 250 pools are sufficient for a BAC library that should satisfy many research objectives. This non-gridded strategy would dramatically reduce the cost and make robotic equipment non-essential in exploiting BAC technology. To construct a representative library and to minimise clone competition, thawing and re-freezing ligation mixtures and bacterial cultures should be avoided in BAC library construction and application. PMID- 11121494 TI - Double-labeled donor probe can enhance the signal of fluorescence resonance energy transfer (FRET) in detection of nucleic acid hybridization. AB - A set of fluorescently-labeled DNA probes that hybridize with the target RNA and produce fluorescence resonance energy transfer (FRET) signals can be utilized for the detection of specific RNA. We have developed probe sets to detect and discriminate single-strand RNA molecules of plant viral genome, and sought a method to improve the FRET signals to handle in vivo applications. Consequently, we found that a double-labeled donor probe labeled with Bodipy dye yielded a remarkable increase in fluorescence intensity compared to a single-labeled donor probe used in an ordinary FRET. This double-labeled donor system can be easily applied to improve various FRET probes since the dependence upon sequence and label position in enhancement is not as strict. Furthermore this method could be applied to other nucleic acid substances, such as oligo RNA and phosphorothioate oligonucleotides (S-oligos) to enhance FRET signal. Although the double-labeled donor probes labeled with a variety of fluorophores had unexpected properties (strange UV-visible absorption spectra, decrease of intensity and decay of donor fluorescence) compared with single-labeled ones, they had no relation to FRET enhancement. This signal amplification mechanism cannot be explained simply based on our current results and knowledge of FRET. Yet it is possible to utilize this double-labeled donor system in various applications of FRET as a simple signal enhancement method. PMID- 11121496 TI - Cervical lymphadenopathy and adenitis. PMID- 11121495 TI - Controlling gene expression in yeast by inducible site-specific recombination. AB - An intron module was developed for Saccharomyces cerevisiae that imparts conditional gene regulation. The kanMX marker, flanked by loxP sites for the Cre recombinase, was embedded within the ACT1 intron and the resulting module was targeted to specific genes by PCR-mediated gene disruption. Initially, recipient genes were inactivated because the loxP-kanMX-loxP cassette prevented formation of mature transcripts. However, expression was restored by Cre-mediated site specific recombination, which excised the loxP-kanMX-loxP cassette to generate a functional intron that contained a single loxP site. Cre recombinase activity was controlled at the transcriptional level by a GAL1::CRE expression vector or at the enzymatic level by fusing the protein to the hormone-dependent regulatory domain of the estrogen receptor. Negative selection against leaky pre-excision events was achieved by growing cells in modified minimal media that contained geneticin (G418). Advantages of this gene regulation technique, which we term the conditional knock-out approach, are that (i) modified genes are completely inactivated prior to induction, (ii) modified genes are induced rapidly to expression levels that compare to their unmodified counterparts, and (iii) it is easy to use and generally applicable. PMID- 11121497 TI - Pancreatitis in childhood. PMID- 11121498 TI - A synopsis of the American Academy of Pediatrics' practice parameter on the management of minor closed head injury in children. PMID- 11121499 TI - Consultation with the specialist: Diagnosis and management of the newborn and young infant who have nasal obstruction. PMID- 11121501 TI - Earning CME credit-completing the PIR quiz PMID- 11121500 TI - Index of suspicion. Case #1. Diagnosis: Allergic contact dermatitis. PMID- 11121502 TI - Infant botulism. PMID- 11121503 TI - Visual diagnosis: A family that has an itchy rash. PMID- 11121504 TI - Nephrotic syndrome. PMID- 11121505 TI - Dislocations. PMID- 11121506 TI - Editor's note: pharmacological reviews to publish online before print PMID- 11121507 TI - Yeast mutants as a model system for identification of determinants of chemosensitivity. AB - The fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae have become valuable tools for the study of basic cellular functions of eukaryotic cells, including DNA repair mechanisms and cell cycle control. Since the major signaling pathways and cellular processes involved in cellular response to cytotoxic agents are conserved between yeasts and mammalian cells, these simple eukaryotic systems could be excellent models for the identification of molecular/cellular mechanisms of sensitivity to antitumor drugs. We describe relevant biological features of yeast cells and potential applications derived by their genetic manipulation. In particular, we have outlined the role of genes involved in repair processes and in checkpoint control, with specific reference to genes regulating radiation-sensitivity. Specific examples are provided concerning the use of both yeasts in understanding the mechanism of action of platinum compounds and topoisomerase inhibitors. The availability of the genomic sequence of these organisms as well as of new technologies (microarrays, proteomics) is expected to allow the identification of potential drug targets, since the drug discovery process is moving toward a genomic orientation. Among eukaryotic organisms, yeasts are suitable for easy genetic manipulations, and specific genetic alterations are exploitable for assessing the effects of chemotherapeutic agents with different mechanism of action. Although still at an early stage, this fast-moving field shows promise as a novel and potentially useful method for development of target-specific therapeutic approaches. PMID- 11121508 TI - Design of retroviral vectors and helper cells for gene therapy. AB - During the past decade, gene therapy has been applied to the treatment of disease in hundreds of clinical trials. Various tools have been developed to deliver genes into human cells; among them, genetically engineered retroviruses are currently the most popular tool for gene delivery. Most of the systems contain vectors that are capable of accommodating genes of interest and helper cells that can provide the viral structural proteins and enzymes to allow for the generation of vector-containing infectious viral particles. Retroviridae is a family of retroviruses that differs in nucleotide and amino acid sequence, genome structure, pathogenicity, and host range. This diversity provides opportunities to use viruses with different biological characteristics to develop different therapeutic applications. Currently, a variety of retroviruses that provide distinct advantages for gene delivery has been modified and used in clinical trials. In this review, the genome structures of oncoviruses, lentiviruses, and spumaviruses are reviewed and examples of vectors derived from these viruses are described. As with any delivery tool, the efficiency, the ability to target certain tissue or cell type, the expression of the gene of interest, and the safety of retroviral-based systems are important for successful application of gene therapy. Significant efforts have been dedicated to these areas of research in recent years. Various modifications have been made to retroviral-based vectors and helper cells to alter gene expression, target delivery, improve viral titers, and increase safety. The principles and design of these modifications are discussed in this review. PMID- 11121509 TI - Multiple actions of steroid hormones--a focus on rapid, nongenomic effects. AB - According to the traditional model, steroid hormones bind to intracellular receptors and subsequently modulate transcription and protein synthesis, thus triggering genomic events finally responsible for delayed effects. Based upon similarities in molecular structure, specific receptors for steroids, vitamin D(3) derivatives, thyroid hormone, retinoids, and a variety of orphan receptors are considered to represent a superfamily of steroid receptors. In addition, very rapid effects of steroids mainly affecting intracellular signaling have been widely recognized that are clearly incompatible with the genomic model. These rapid, nongenomic steroid actions are likely to be transmitted via specific membrane receptors. Evidence for nongenomic steroid effects and distinct receptors involved is presented for all steroid groups including related compounds like vitamin D(3) and thyroid hormones. The physiological and clinical relevance of these rapid effects is still largely unclear, but their existence in vivo has been clearly shown in various settings including human studies. Drugs that specifically affect nongenomic steroid action may find applications in various clinical areas such as cardiovascular and central nervous disorders, electrolyte homeostasis, and infertility. In addition to a short description of genomic steroid action, this review pays particular attention to the current knowledge and important results on the mechanisms of nongenomic steroid action. The modes of action are discussed in relation to their potential physiological or pathophysiological relevance and with regard to a cross-talk between genomic and nongenomic responses. PMID- 11121510 TI - Potassium channels: molecular defects, diseases, and therapeutic opportunities. AB - Potassium channels play important roles in vital cellular signaling processes in both excitable and nonexcitable cells. Over 50 human genes encoding various K(+) channels have been cloned during the past decade, and precise biophysical properties, subunit stoichiometry, channel assembly, and modulation by second messenger and ligands have been elucidated to a large extent. Recent advances in genetic linkage analysis have greatly facilitated the identification of many disease-producing loci, and naturally occurring mutations in various K(+) channels have been identified in diseases such as long-QT syndromes, episodic ataxia/myokymia, familial convulsions, hearing and vestibular diseases, Bartter's syndrome, and familial persistent hyperinsulinemic hypoglycemia of infancy. In addition, changes in K(+) channel function have been associated with cardiac hypertrophy and failure, apoptosis and oncogenesis, and various neurodegenerative and neuromuscular disorders. This review aims to 1) provide an understanding of K(+) channel function at the molecular level in the context of disease processes and 2) discuss the progress, hurdles, challenges, and opportunities in the exploitation of K(+) channels as therapeutic targets by pharmacological and emerging genetic approaches. PMID- 11121511 TI - The sympathetic nerve--an integrative interface between two supersystems: the brain and the immune system. AB - The brain and the immune system are the two major adaptive systems of the body. During an immune response the brain and the immune system "talk to each other" and this process is essential for maintaining homeostasis. Two major pathway systems are involved in this cross-talk: the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system (SNS). This overview focuses on the role of SNS in neuroimmune interactions, an area that has received much less attention than the role of HPA axis. Evidence accumulated over the last 20 years suggests that norepinephrine (NE) fulfills the criteria for neurotransmitter/neuromodulator in lymphoid organs. Thus, primary and secondary lymphoid organs receive extensive sympathetic/noradrenergic innervation. Under stimulation, NE is released from the sympathetic nerve terminals in these organs, and the target immune cells express adrenoreceptors. Through stimulation of these receptors, locally released NE, or circulating catecholamines such as epinephrine, affect lymphocyte traffic, circulation, and proliferation, and modulate cytokine production and the functional activity of different lymphoid cells. Although there exists substantial sympathetic innervation in the bone marrow, and particularly in the thymus and mucosal tissues, our knowledge about the effect of the sympathetic neural input on hematopoiesis, thymocyte development, and mucosal immunity is extremely modest. In addition, recent evidence is discussed that NE and epinephrine, through stimulation of the beta(2) adrenoreceptor-cAMP-protein kinase A pathway, inhibit the production of type 1/proinflammatory cytokines, such as interleukin (IL-12), tumor necrosis factor alpha, and interferon-gamma by antigen-presenting cells and T helper (Th) 1 cells, whereas they stimulate the production of type 2/anti-inflammatory cytokines such as IL-10 and transforming growth factor-beta. Through this mechanism, systemically, endogenous catecholamines may cause a selective suppression of Th1 responses and cellular immunity, and a Th2 shift toward dominance of humoral immunity. On the other hand, in certain local responses, and under certain conditions, catecholamines may actually boost regional immune responses, through induction of IL-1, tumor necrosis factor-alpha, and primarily IL-8 production. Thus, the activation of SNS during an immune response might be aimed to localize the inflammatory response, through induction of neutrophil accumulation and stimulation of more specific humoral immune responses, although systemically it may suppress Th1 responses, and, thus protect the organism from the detrimental effects of proinflammatory cytokines and other products of activated macrophages. The above-mentioned immunomodulatory effects of catecholamines and the role of SNS are also discussed in the context of their clinical implication in certain infections, major injury and sepsis, autoimmunity, chronic pain and fatigue syndromes, and tumor growth. Finally, the pharmacological manipulation of the sympathetic-immune interface is reviewed with focus on new therapeutic strategies using selective alpha(2)- and beta(2) adrenoreceptor agonists and antagonists and inhibitors of phosphodiesterase type IV in the treatment of experimental models of autoimmune diseases, fibromyalgia, and chronic fatigue syndrome. PMID- 11121512 TI - Signal transduction mechanisms mediating the physiological and pathophysiological actions of angiotensin II in vascular smooth muscle cells. AB - Until recently, the signaling events elicited in vascular smooth muscle cells by angiotensin II (Ang II) were considered to be rapid, short-lived, and divided into separate linear pathways, where intracellular targets of the phospholipase C diacylglycerol-Ca(2+) axis were distinct from those of the tyrosine kinase- and mitogen-activated protein kinase- dependent pathways. However, these major intracellular signaling cascades do not function independently and are actively engaged in cross-talk. Downstream signals from the Ang II-bound receptors converge to elicit complex and multiple responses. The exact adapter proteins or "go-between" molecules that link the multiple intracellular pathways await clarification. Ang II induces a multitude of actions in various tissues, and the signaling events following occupancy and activation of angiotensin receptors are tightly controlled and extremely complex. Alterations of these highly regulated signaling pathways in vascular smooth cells may be pivotal in structural and functional abnormalities that underlie vascular pathological processes in cardiovascular diseases such as hypertension, atherosclerosis, and post interventional restenosis. PMID- 11121513 TI - The effects of plant flavonoids on mammalian cells: implications for inflammation, heart disease, and cancer. AB - Flavonoids are nearly ubiquitous in plants and are recognized as the pigments responsible for the colors of leaves, especially in autumn. They are rich in seeds, citrus fruits, olive oil, tea, and red wine. They are low molecular weight compounds composed of a three-ring structure with various substitutions. This basic structure is shared by tocopherols (vitamin E). Flavonoids can be subdivided according to the presence of an oxy group at position 4, a double bond between carbon atoms 2 and 3, or a hydroxyl group in position 3 of the C (middle) ring. These characteristics appear to also be required for best activity, especially antioxidant and antiproliferative, in the systems studied. The particular hydroxylation pattern of the B ring of the flavonoles increases their activities, especially in inhibition of mast cell secretion. Certain plants and spices containing flavonoids have been used for thousands of years in traditional Eastern medicine. In spite of the voluminous literature available, however, Western medicine has not yet used flavonoids therapeutically, even though their safety record is exceptional. Suggestions are made where such possibilities may be worth pursuing. PMID- 11121514 TI - Design and use of electrochemical sensors in enantioselective high throughput screening of drugs. A minireview. AB - The importance of reliable detection systems for enantiomeric assays increases with the necessity of high throughput screening analysis of raw materials for the pharmaceutical industry. The utilization of electrochemical sensors in enantioselective analysis is an accurate and precise alternative to chromatographic techniques. The reliability of the response characteristics as well as of the analytical information obtained by using electrochemical sensors is strictly correlated with the design of the sensors. The designs evaluated for sensors have been based on PVC, imprinting polymers and carbon paste matrices. Among these, carbon paste sensors have been the most reliable and have been utilized for the construction of potentiometric, enantioselective membrane electrodes as well as for amperometric biosensors, and immunosensors. There are two ways to use the electrochemical sensors in enantioselective screening analysis: selective binding and catalyst selectivity. A molecule with a special chemical architecture is required for selective binding: a lock for a key. The high reliability of analytical information obtained using these sensors has made possible the automation of potentiometric and amperometric techniques by integration of enantioselective sensors as detectors in flow injection analysis and sequential injection analysis techniques. PMID- 11121515 TI - Use of capillary electrophoresis for high throughput screening in biomedical applications. A minireview. AB - Diagnosis of inherited diseases or cancer predispositions frequently involves determination of specific mutations or polymorphisms. The number of characterized monogenetic and polygenetic diseases is significantly rising every year. As a result, an increasing number of patient samples with a rising complexity of genetic diseases require molecular diagnostics. In order to apply genetic analyses to large groups of patients or population screening, automation of a sensitive and precise method is highly desirable. Capillary electrophoresis (CE) facilitates the development of methods which can rapidly process large number of patient samples in an automated fashion. In contrast, conventional techniques including Southern blotting, sequencing or standard gel electrophoresis are time consuming, cost ineffective and require substantial amounts of each specimen. Robustness, ease of operation, good reproducibility and low cost are the main advantages of CE. Currently, most protocols adapted to automated CE represent (i) analyses of DNA fragment length or DNA restriction patterns (RFLP), (ii) analyses of single-strand conformation polymorphism (SSCP) and (iii) microsatellite analyses. Recently, automated detection of variations in the FRAXA (CGG)n region (fragile X syndrome), LDL receptor gene, p53 gene, MTHFR (methylenetetrahydrofolate reductase) gene, HFE gene and others has been established on CE systems. These applications clearly demonstrate the suitability of CE for high throughput screening in medical applications. PMID- 11121516 TI - Screening procedures for simultaneous detection of several drug classes used for high throughput toxicological analyses and doping control. A review. AB - This paper reviews high throughput screening procedures for the simultaneous detection of several drug classes relevant to clinical and forensic toxicology or doping control in urine or blood using gas chromatography-mass spectrometry (GC MS), liquid chromatography coupled with a diode-array detector (LC-DAD) or mass spectrometry (LC-MS). Basic information describing these systematic toxicological analysis (STA) procedures such as the analytes, the biosample, work-up, separation column, mobile phase or separation buffer, detection mode and detection limits are summarized in tables arranged according to the analytical method. Examples of typical applications are presented in 2 figures. Analysis of alternative matrices, like sweat, saliva, nails or hair, was not reviewed. PMID- 11121517 TI - Self-organizing neural networks for screening and development of novel artificial sweetener candidates. AB - The use of Kohonen feature maps for the visualization of various aspects of molecular similarity is briefly reviewed and illustrated. It is shown that a specific feature of self-organizing maps (SOM) makes them of special interest for the screening of compounds. In particular, these methods were used to design candidates for new sweeteners, which were then synthesized. PMID- 11121518 TI - Enantioseparations using cellulose tris(3,5-dichlorophenylcarbamate) during high performance liquid chromatography with analytical and capillary columns: potential for screening of chiral compounds. AB - The appropriate selection of the mobile phase facilitated the use cellulose tris(3,5-dichlorophenylcarbamate (CDCPC) as a chiral stationary phase (CSP) during high-performance liquid chromatography (HPLC). A preliminary evaluations of this material indicated its very high chiral resolving ability toward many analytes of different chemical and pharmacological groups. Some chemicals and drugs containing two centers of chirality were also successfully resolved into all possible stereoisomers. The applicability of CDCPC for enantioseparations in capillary liquid chromatography was also shown giving promising prospects for the screening of novel biologically active compounds (which may be also synthesized based on combinatorial strategies) for their enantiomeric composition. PMID- 11121519 TI - Polymeric liquid membrane electrodes incorporated with macrocyclic hexaamines for screening adenine nucleotides. AB - Lipophilic macrocyclic hexaamines supported by a poly(vinyl chloride) PVC matrix were used for the construction of liquid membrane electrodes sensitive toward adenine nucleotide polyanions. The membrane potential strongly depended on the pH of the sample solution. This phenomenon occurs due to the ability of the ionophore to accept protons. Therefore, the optimum pH was determined based on potential pH profile. The potential measurements were carried out at pH 6.0 in the presence of 10(-2) M 2-[N-morpholino] ethanesulfonic acid (MES) buffer. The potential response of these electrodes toward ATP(-4) and/or HATP(-3) was close to the Nernstian slope. The selectivities against ADP(-3), AMP(-2), HPO(4)(-2), and monovalent inorganic anions were estimated using the matched potential method. Chloride ions slightly affected potential response of the electrodes toward ATP(-4)/HATP(-3). The influence of ionophore chemical structure on the selectivity and the sensitivity of these electrodes is briefly discussed. PMID- 11121520 TI - High throughput genetic screening for the detection of hereditary non-polyposis colon cancer (HNPCC) using capillary electrophoresis. AB - Approximately 5-10% of all colorectal carcinomas arise from cancer predisposition syndromes caused by heterozygote germline mutations in post-replicative DNA mismatch repair (MMR) genes. In contrast to gastrointestinal polyposis syndromes, carcinomas in these patients do not occur on the background of increased numbers of polyps and hence are refered to as hereditary non-polyposis colorectal cancers (HNPCC). Six different MMR genes, MSH2, MSH3, MSH6, MLH1, MLH3 and PMS2, have been identified in the human genome. In the majority of HNPCC patients, heterozygote germline mutations are present in the MSH2 or MLH1 gene. Detection of mutations by conventional sequencing technology is expensive and labor intensive due to the complex intron and/or exon structures. In this study, we therefore have explored whether capillary electrophoresis-based single strand conformation polymorphism (SSCP-CE) provides a reliable means for mutation screening. We have tested different MLH1 mutations in exons 9 and 16 and find that SSCP-CE produces reliable electrophoretic patterns that allow recognition of wild-type alleles, microdeletions and point mutations. In summary, SSCP-CE provides a rapid, automated, and cost-effective technology for MSH2 and MLH1 mutation screening and will facilitate genetic diagnostics for HNPCC patients. PMID- 11121521 TI - Pharmacological classification of drugs based on neural network processing of molecular modeling data. AB - The performance of artificial neural network (ANN) models in predicting pharmacological classification of structurally diverse drugs based on their theoretical chemical parameters was demonstrated. The classification coefficients for psychotropic agents, beta-adrenolytic drugs, histamine H(1) receptor antagonists and drugs binding to alpha-adrenoceptors were 100, 100, 95 and 86%, respectively. A set of easily accessible non-empirical molecular parameters describing the structure of xenobiotics can provide information allowing the prediction of some pharmacological properties of drugs and drug candidates employing ANN models. Since ANN analysis can help cluster as well as segregate drugs and drug candidates according to their known and expected pharmacological properties, the number of routine biological assays might be reduced. The results presented here might be used to improve the efficiency of high throughput screening programs for new drug hits by demonstrating a promising procedure for diverse combinatorial library design and evaluation. PMID- 11121522 TI - A neural network based virtual high throughput screening test for the prediction of CNS activity. AB - A virtual high throughput screening test to identify potentially CNS-active drugs has been developed. Discrimination was based on the knowledge available in databases containing CNS-active (Cipsline from Prous Science) and inactive compounds (Chemical Directory from Sigma-Aldrich). Molecular structures were represented using 2D Unit y fingerprints and a feedforward neural network was trained to classify molecules regarding their CNS activity. The parameterized network was validated by reclassification of the training set elements, by the classification of a test set preselected from the Prous database, and also by the prediction of activity for known CNS drugs not used in the training set but available in the Medchem database (Daylight). These tests revealed that our neural net recognized at least 89% of CNS-active compounds and would be suitable for use in our virtual screening protocol. PMID- 11121523 TI - Regional changes in the hippocampal density of AMPA and NMDA receptors across the lifespan of the rat. AB - The current study dissected the fascia dentata (FD) and hilar region from the CA and subicular cell fields of the rat and conducted in vitro determinations of the number of binding sites for N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy 5-methyl-4-isoxazole (AMPA) glutamate receptors across the lifespan. We determined the density of binding of [3H]-glutamate or [3H]-AMPA to NMDA or AMPA receptor sites, respectively. The changes reported might be due to either a change in receptor number or an alteration in the binding characteristics of the receptor site with aging. We found an age-related decline in the number of NMDA receptors in the CA1, CA3 and subicular cell regions of the hippocampus, but not in the FD/hilar region, and an age-related decline in the number of AMPA receptors in the FD/hilar region, but not in the CA fields. The decline in the number of NMDA or AMPA receptors that occurs with aging was not a continuous or homogeneous process. These changes in receptor number might underlie selected age associated changes in sensitivity to drugs that influence hippocampal function as well as to changes in NMDA-dependent long-term potentiation. A thorough understanding of the mechanisms underlying changes in glutamate receptor function in discrete brain regions, using combined neurochemical and electrophysiological methods, may ultimately provide insight into the fundamental substrates of age associated memory disorders related to hippocampal dysfunction. PMID- 11121524 TI - Immuno-electron microscopic localization of the alpha(1) and beta(1)-subunits of soluble guanylyl cyclase in the guinea pig organ of corti. AB - Guanylyl cyclases (GC) catalyze the formation of the intracellular signal molecule cyclic GMP from GTP. For some years it has been known that the heme containing soluble guanylyl cyclase (sGC) is stimulated by NO and NO-containing compounds. The sGC enzyme consists of two subunits (alpha(1) and beta(1)). In the present study, the alpha(1) and beta(1)-subunits were identified in the guinea pig cochlea at the electron microscopic level using a post-embedding immuno labeling procedure. Ultrathin sections of LR White embedded specimens were incubated with various concentrations of two rabbit polyclonal antibodies to the alpha(1)- and beta(1)-subunit, respectively. The immunoreactivity was visualized by a gold-labeled secondary antibody in an energy-filtering transmission electron microscope (EFTEM). Marked immunoreactivity for both antibodies was found in the inner and outer hair cells, with numerous gold particles at the border of the cuticular plates, associated with the cell nuclei or attached to electron-dense parts of the cytoplasm. In the pillar cells and apical Deiters cells, soluble guanylyl cyclase immunoreactivity was located at the rim of the cuticular plates and between the microtubuli bundles. Together with the recently identified nitric oxide synthase isoforms [Eur. Arch. Otorhinolaryngol. 254 (1997) 396; Eur. Arch. Otorhinolaryngol. 255 (1998) 483], the soluble guanylyl cyclase may be involved in signalling processes in the organ of Corti. PMID- 11121525 TI - Interferon-alpha inhibits long-term potentiation and unmasks a long-term depression in the rat hippocampus. AB - Interferons (IFN) appear to have various neuromodulatory actions. Here, we characterized the actions of IFN-alpha on the electrophysiological properties of CA1 hippocampal neurons using intracellular recordings. Superfusion of this cytokine did not alter the resting membrane potential, cell input resistance, action potentials, nor GABA-mediated fast synaptic potentials. IFN-alpha inhibited glutamate-mediated excitatory postsynaptic potentials (gEPSPs) and reversed or prevented long-term potentiation (LTP) induced by high-frequency tetanic stimulation. IFN-alpha reduced gEPSP amplitude far below its control value. Only a short-term potentiation (STP) was observed when either IFN-alpha or D-2-amino-5-phosphonovalerato (APV; NMDA receptor antagonist) were present during tetanic stimulation. After this STP in presence of APV, IFN-alpha had no effect on gEPSPs. APV had no effect on LTP when applied after tetanic stimulation and did also not prevent IFN-alpha effect on LTP. Genistein (a tyrosine kinase inhibitor) or heat inactivation prevented IFN-alpha effects. IFN-alpha also decreased the depolarization induced by local application of glutamate but did not modify those induced by NMDA. Similarly, IFN-alpha reversed the potentiation (induced by tetanic stimulation) of glutamate-induced depolarizations. IFN-alpha did not affect long-term depression (LTD) induced by low-frequency tetanic stimulation. In conclusion, IFN-alpha-induced inhibition of LTP is, at least in part, mediated by a postsynaptic effect, by tyrosine kinase activity, and by non NMDA glutamate receptors. Inhibition of LTP by IFN-alpha unmasks LTD which is induced by the same high-frequency tetanic stimulation. PMID- 11121526 TI - Changes in expressions of proinflammatory cytokines IL-1beta, TNF-alpha and IL-6 in the brain of senescence accelerated mouse (SAM) P8. AB - The senescence-accelerated mouse (SAM) is known to be a murine model for accelerated aging. The SAMP8 strain shows age-related deterioration of learning and memory at an earlier age than control mice (SAMR1). In the present study, we investigated the changes in expressions of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in the brain of SAMP8. In the hippocampus of 10 months old SAMP8, the expression of IL-1 mRNA was significantly elevated in comparison with that of SAMR1. In both strains of SAMs, increases in IL-1beta protein in the brain were observed at 10 months of age compared with 2 and 5 months. The only differences found between the strain in protein levels were at 10 months and were elevations in IL-1beta in the hippocampus and hypothalamus, and in TNF-alpha and IL-6 in the cerebral cortex and the hippocampus in SAMP8 as compared with SAMR1. However, lipopolysaccharide induced increases in the expression of these cytokines in brain did not differ between SAMP8 and SAMR1. Increases in expression of proinflammatory cytokines in the brain may be involved in the age-related neural dysfunction and/or learning deficiency in SAMP8. PMID- 11121527 TI - Electrophysiological properties of axons in mice lacking neurofilament subunit genes: disparity between conduction velocity and axon diameter in absence of NF H. AB - Neurofilament proteins (NFs) are made by co-polymerization of three intermediate filament proteins, NF-L, NF-M and NF-H and constitute the most abundant cytoskeletal element in large myelinated axons. NFs have a well-established role as intrinsic determinants of axon caliber with all the functional implications, but the role of each individual NF subunit is much less clear. The aim of our study was to examine functional properties of large myelinated axons with altered morphology from mice bearing a targeted disruption of each NF genes (NF-L -/-, NF M-/- and NF-H -/- mice). Membrane properties, action potentials and single axon refractory period were measured in isolated sciatic nerves in vitro, using intra axonal microelectrode recording in conjunction with current-clamp technique. Some results were obtained from whole nerves by sucrose-gap recording. The NF-knockout mice showed several deficits in physiological properties of low-threshold fibers. In keeping with smaller axon diameter, the conduction velocity was significantly decreased in NF-L -/- and NF-M -/- transgenic animals (control, 39.9+/-1.8 m/s, NF-M -/-; 23.5+/-1. 4 m/s, and NF-L-/-; 12.0+/-0.7 m/s, mean+/-S.E.M.; intra axonal recording; similar ratios obtained by sucrose-gap recording; 22-26 degrees C). However, in spite of their preserved caliber, large myelinated axons in NF-H /- mice also showed a significant decrease in conduction velocity (22.8+/-1.0 m/s, mean+/-S.E.M.). Although action potential amplitudes, duration and shape did not differ between control axons and transgenic animals, the refractory period was prolonged in NF-H -/- and NF-M -/- animals. Intracellular injections of 200 ms depolarizing and hyperpolarizing currents revealed outward and inward rectification in all animal groups. In comparison to control animals, NF-H -/- mice expressed a significant decrease in outward rectification. Potassium channel blockers (4AP and TEA) and cesium ions were able to block outward and inward rectification in all myelinated axons in qualitatively the same manner. These results suggest that NF-H may have a specific role in modulating ion channel functions in large myelinated fibers. PMID- 11121528 TI - DNA damage in brain mitochondria caused by aging and MPTP treatment. AB - 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment leads to marked depletion of dopamine (DA) levels in the nigrostriatal pathway and dopaminergic neuronal degeneration in caudate-putamen and substantia nigra. MPTP is believed to inhibit complex I of the electron transport system leading to the generation of reactive oxygen species. We sought to test the hypotheses that MPTP treatment: (1) leads to dopamine depletion; (2) causes extensive mitochondrial DNA damage, and (3) that these effects would be age dependent. The levels of dopamine and its metabolites, DOPAC and HVA were analyzed by HPLC equipped with electrochemical detection. DNA damage was measured by quantitative PCR in both mitochondrial and nuclear (beta-polymerase) targets from the caudate-putamen, substantia nigra and cerebellum regions of control and MPTP-treated mice. The age groups studied were 22 days and 12 months. MPTP produced no significant effect on the levels of dopamine and its metabolites in young mice whereas in old, there was a significant decrease in this neurotransmitter system after MPTP administration. These 12-month-old mice, when compared to the young mice, showed a significant increase in mitochondrial DNA damage in the caudate-putamen and cerebellum. The latter region also displayed a significant increase in DNA damage in a nuclear gene. After treatment with MPTP, there was an age-dependent increase in DNA damage in mitochondria of the caudate-putamen while there was no significant DNA damage in the nuclear target. MPTP treatment led to damage in both mitochondrial and nuclear DNA of the substantia nigra, while there was no damage in either mitochondria or nucleus in cerebellum which was used as a negative control. PMID- 11121529 TI - Early loss of synaptic protein PSD-95 from rod terminals of rhodopsin P347L transgenic porcine retina. AB - Retinitis pigmentosa (RP), a type of retinal degeneration involving first rod and then slow cone photoreceptor degeneration, can be caused by any of a number of mutations in different genes. In the cases of mutations affecting rod-specific genes such as rhodopsin, it is unclear how the mutations may cause degeneration of cones. We have used the porcine retina, which is rod-dominated and has an abundance of cones, to study the mutation-induced changes in both rod and cone photoreceptors. Like patients with the same mutation, rhodopsin P347L transgenic swine manifest rod-cone degeneration. In addition, the rod bipolar cells fail to form synaptic connections with rods; instead, they form ectopic synapses with cones. The mechanisms that prevent the formation of the rod-rod bipolar cell synaptic connection are not known. We used specific antibodies and immunocytochemistry to show that the synaptic protein, PSD-95, is present in both normal and transgenic porcine retinas. During neonatal development, however, PSD 95 is lost from rod terminals in the transgenic swine. This loss is virtually complete (90%) by postnatal day 5, at a time when greater than 80% of rod cell bodies still remain. Furthermore, the remaining rods retain their outer segments and their gross morphology appears relatively normal. In contrast, PSD-95 expression continues in cone terminals, even in 10-month-old transgenic swine, where the rods have all disappeared and the cones show signs of severe degeneration. These results suggest that loss of PSD-95 may not be a general consequence of the deteriorating cell. Rather, the very early and selective loss of PSD-95 from the rod terminals may be causally related to the absence of rod rod bipolar cell synapses in the rhodopsin P347L transgenic retina. PMID- 11121530 TI - Dietary restriction attenuates the neuronal loss, induction of heme oxygenase-1 and blood-brain barrier breakdown induced by impaired oxidative metabolism. AB - Experimental thiamine deficiency (TD) is a model of impaired oxidative metabolism associated with region-selective neuronal loss in the brain. Oxidative stress is a prominent feature of TD neuropathology, as evidenced by the accumulation of heme oxygenase-1 (HO-1), ferritin, reactive iron and superoxide dismutase in microglia, nitrotyrosine and 4-hydroxynonenal in neurons, as well as induction of endothelial nitric oxide synthase within the vulnerable areas. Dietary restriction (DR) reduces oxidative stress in several organ systems including the brain. DR increases lifespan and reduces neurodegeneration in a variety of models of neuronal injury. The possibility that DR can protect vulnerable neurons against TD-induced oxidative insults has not been tested. The current studies tested whether approximately 3 months of DR (60% of ad libitum intake) altered the response to TD. Six month-old ad libitum-fed or dietary restricted C57BL/6 mice received a thiamine-deficient diet either ad libitum, or under a DR regimen respectively for eleven days. The TD mice also received daily injections of the thiamine antagonist pyrithiamine. Control ad libitum-fed or DR mice received an unlimited amount, or 60% of ad libitum intake, respectively, of thiamine supplemented diet. As in past studies, TD produced region-selective neuronal loss (-60%), HO-1 induction, and IgG extravasation in the thalamus of ad libitum-fed mice. DR attenuated the TD-induced neuronal loss (-30%), HO-1 induction and IgG extravasation in the thalamus. These studies suggest that oxidative damage is critical to the pathogenesis of TD, and that DR modulates the extent of free radical damage in the brain. Thus, TD is an important model for studying the relationship between aging, oxidative stress and nutrition. PMID- 11121531 TI - Brainstem catecholaminergic neurons activated by hypoxemia express GR and are coordinately activated with fetal sheep hypothalamic paraventricular CRH neurons. AB - In late gestation, challenges to fetal homeostasis are accompanied by increases in adrenocorticotropin (ACTH) concentrations in fetal peripheral plasma and Fos (c-fos protein) activation in corticotropin-releasing hormone (CRH) neurons of the fetal hypothalamic paraventricular nucleus (PVN). In adults, ventrolateral brainstem catecholaminergic (CA) neurons (A1/C1, A2/C2) project to the parvocellular neurons of the PVN, possess glucocorticoid receptors (GR) and are Fos activated in parallel with CRH neurons of the PVN during hypoxia. Such observations suggest a role for the aforementioned medullary neurons in the function of the hypothalamo-pituitary-adrenal axis. The present study utilized late gestation fetal sheep, stereotaxic methodology and retrograde axon tracing and immunocytochemical techniques to investigate the relationship between activation of fetal brainstem CA neurons and activation of fetal PVN CRH immunopositive neurons in response to hypoxemia. Results indicated that: (1) the largest brainstem CA projection to PVN CRH neurons is from A1/C1 neurons, (2) brainstem neurons exhibit GR immunostaining and (3) brainstem CA neurons show a strong correlation (A1/C1 - r(2)=0.894, P<0.005; A2/C2 - r(2)=0. 848; P<0.002) of Fos activation with Fos activation in PVN CRH cells. We conclude that in late gestation the brainstem A1/C1 and A2/C2 areas are in position to influence the function of the hypothalamo-pituitary-adrenal axis during hypoxemic challenges to homeostasis in a fashion similar to that which has been demonstrated in the adult rat. PMID- 11121532 TI - Neurotrophin-3 antisense oligonucleotide attenuates nerve injury-induced Abeta fibre sprouting. AB - It is proposed that following peripheral nerve injury abnormal sprouting of Abeta fibre primary afferent neurons in the spinal cord contributes to the allodynia that often occurs with such injury. Allodynia is characterized as pain due to a stimulus which is normally non-noxious. Our recent in vivo experiments show that intrathecal administration of neurotrophin-3 (NT-3), in normal animals, induces allodynia and sprouting of Abeta-fibres. In this study, we examine whether intrathecal administration of NT-3 antisense oligonucleotides (50 microM), via an osmotic pump for 14 days, attenuates nerve injury-induced sprouting and allodynia. The oligonucleotides used in this study were phosphorothioate modified and control experiments, using an ELISA, confirm that intrathecal administration of the antisense induces a significant decrease in NT-3 levels in the spinal cord. All surgery was conducted on anaesthetized Wistar rats (sodium pentobarbitone, i.p. 50 mg/kg). Consistent with previous studies, transganglionic labelling of Abeta-fibres with choleragenoid-horseradish peroxidase (C-HRP) shows that complete transection of the sciatic nerve induces an expansion of C-HRP label into lamina II of the spinal dorsal horn. Using image analysis, we find that intrathecal administration of NT-3 antisense attenuates the density of C-HRP labelling in lamina II in nerve injured animals. A NT-3 sense oligonucleotide (50 microM) has no effect. To test the effect of NT-3 antisense on allodynia, the nociceptive flexion reflex is examined, using an Ugo Basile Analgesymeter, in animals with partial sciatic nerve ligation. Intrathecal administration of 50 microM NT-3 antisense significantly attenuates nerve injury-induced allodynia, whereas the sense oligonucleotide has no effect. These results provide further evidence that endogenous NT-3 contributes to both nerve injury-induced Abeta fibre sprouting and allodynia and demonstrates the potential of neurotrophin-3 antisense oligonucleotides as therapeutic agents for neuropathic pain. PMID- 11121533 TI - Effects of bicuculline on direction-sensitive relay cells in the dorsal lateral geniculate nucleus (LGNd) of cats. AB - The direction sensitivity of relay cells in the cat's dorsal lateral geniculate (LGNd) was measured using sinusoidal grating stimuli before and during local bicuculline administration. One hundred and twenty-eight LGNd relay cells were recorded in laminae A and A1, of which 44 relay cells (34%) were found to be sensitive to direction of stimulus movement. The direction-sensitive LGNd relay cells could be differentiated into two subgroups based on different measures of their response amplitude. Type I cells exhibited their direction sensitivity when the fundamental Fourier component (FFC) of the poststimulus time histograms (PSTHs) was used as response measure, but did not show significant direction sensitivity when mean firing rate was used. Type II cells exhibited their direction sensitivity, no matter whether the FFC or mean firing rate was used as the measure. Of 35 cells analyzed, 27 cells remained direction sensitive during bicuculline administration. At the population level, the direction bias of type I cells did not change systematically, while the direction bias of type II cells decreased significantly during bicuculline administration. These results suggest that the direction bias of these two types of relay cells are mediated by different neural mechanisms. The direction bias of type I cells may involve multiple inputs from spatio-temporally separate subunits within retinal ganglion cells receptive fields. The direction bias of type II cells may involve GABAergic neuronal circuits within the LGNd. PMID- 11121534 TI - Enhancement of persistent sodium current by internal fluorescence in isolated hippocampal neurons. AB - Following up on an earlier chance observation, voltage-dependent whole-cell currents were recorded from isolated hippocampal neurons filled with the fluorescent dyes Fluo-3 and Fura-red, that were intermittently excited by 488 nm laser light. In the absence of any ion channel blocking drugs, in most cells depolarizing voltage steps initially evoked only the 'Hodgkin-Huxley' type early, fast inward surge followed by sustained outward current. Over 5-20 min of intermittent electrical stimulation and laser-excited fluorescence pulses, a voltage-dependent, slowly inactivating inward current also appeared and grew, while sustained outward current diminished. When K(+) currents were blocked, a small persistent inward current was usually detectable immediately, and then it increased in amplitude. This current was blocked by tetrodotoxin (TTX) and it had current-voltage (I-V) characteristics of a persistent sodium current, I(Na,P). In cells not filled with dye but illuminated by laser, and in cells with dye but not illuminated, I(Na,P) remained small. There was a more than 12-fold difference in the maximal amplitude of I(Na, P) of fluorescent compared to non-fluorescent cells. Once induced, I(Na,P) decreased very slowly. Fluorescence increased the duration but not the amplitude of the transient Na(+) current, I(Na,T). With membrane potential clamped to a constant voltage, the laser-induced fluorescence did not evoke a membrane current. It is not certain whether fluorescence-induced I(Na,P) potentiation is related to photodynamic action. PMID- 11121535 TI - Potassium-induced enhancement of persistent inward current in hippocampal neurons in isolation and in tissue slices. AB - Previous work suggested a role for the voltage-dependent persistent sodium current, I(Na,P), in the generation of seizures and spreading depression (SD). Ordinarily, I(Na,P) is small in hippocampal neurons. We investigated the effect of raising external K(+) concentration, [K(+)](o), on whole-cell persistent inward current in freshly isolated hippocampal CA1 pyramidal neurons. I(Na,P) was identified by TTX-sensitivity and dependence on external Na(+) concentration. When none of the ion channels were blocked, I(Na,P) was not usually detectable, probably because competing K(+) current masked it, but after raising [K(+)](o) I(Na,P) appeared, while K(+) currents diminished. With K(+) channels blocked, I(Na,P) could usually be evoked in control solution and raising [K(+)](o) caused its reversible increase in most cells. The increase did not depend on external calcium [Ca(2+)](o). In CA1 pyramidal neurons in hippocampal slices a TTX sensitive persistent inward current was always recorded and when [K(+)](o) was raised, it was reversibly enhanced. Strong depolarization evoked irregular current fluctuations, which were also augmented in high [K(+)](o). The findings support a role of potassium-mediated positive feedback in the generation of seizures and spreading depression. PMID- 11121536 TI - Cyclooxygenase (COX)-1 expressing macrophages/microglial cells and COX-2 expressing astrocytes accumulate during oligodendroglioma progression. AB - Cyclooxygenases (COX, prostaglandin endoperoxide synthases, PGG/H synthases) are potent mediators of edema, impeding blood flow and immunomodulation in the pathologically altered brain. Two COX iso-enzymes have been associated with brain disease, the constitutively expressed COX-1 and the cytokine-inducible COX-2. We have used single and double labeling immunohistochemistry to analyse COX-1 and COX-2 expression in twenty-six primary WHO grade II oligodendrogliomas, sixteen primary WHO grade III anaplastic oligodendrogliomas, twenty-seven matched recurrences and ten neuropathologically unaltered brains. COX-1 immunoreactivity was predominantly observed in macrophages/microglial cells. The number of COX-1 expressing macrophages/microglial cells was significantly lower in primary oligodendrogliomas than in primary anaplastic oligodendrogliomas (P<0.0001) and in anaplastic oligodendroglioma relapses (P=0.011). Patients with low COX-1 labeling scores in the primary tumors had significantly longer time to progression and overall survival (P=0.0285) than those with high COX-1 labeling scores. COX-2 immunoreactivity was predominantly observed in disseminated neurons and astrocytes. In glioblastoma multiforme relapses, accumulation of COX-2 expressing astrocytes was observed surrounding areas of focal necrosis. The number of COX-2 expressing astrocytes was significantly (P=0.0471) lower in primary oligodendrogliomas than in high grade oligodendroglioma relapses. These data provide convincing evidence for the differential accumulation of cyclooxygenase isoforms during oligodendroglioma progression in vivo. PMID- 11121537 TI - Expression of CD40 in the brain of Alzheimer's disease and other neurological diseases. AB - We have investigated immunohistochemically the expression of CD40 in post-mortem human brain tissues. In control brain, the blood vessels were stained weakly for CD40. Vascular expression of CD40 was enhanced in the lesions of Alzheimer's disease and some other neurological diseases. In such diseases, reactive microglia were also positive for CD40. The results of this study suggest that CD40 expression by microglia is up-regulated upon a variety of brain insults and is not limited to lesions with amyloid beta-protein deposits. PMID- 11121538 TI - Retinal dopaminergic neurons (A17) expressing neuromedin K receptor (NK(3)): a double immunocytochemical study in the rat. AB - By using a double immunofluorescence method we examined the distribution of dopaminergic neurons (A17) expressing neuromedin K receptor (NKR, NK(3)) in the rat retina. The distribution of NKR-like immunoreactive (-LI) neurons partially overlapped that of tyrosine hydroxylase (TH)-LI neurons in the inner retina of section and flat-mount preparation. Neurons showing both TH- and NKR-like immunoreactivities were found in the retina (A17): 100% of these TH-LI neurons displayed NKR-like immunoreactivity, and they constituted about 3.5% of total NKR LI neurons. The majority of double-labeled neurons with TH- and NKR-like immunoreactivities were distributed in the proximal inner nuclear layer and the upper part of inner plexiform layer of the retina, and characterized with appearance of amacrine cells. The present study has provided morphological evidence for direct physiological modulation of dopaminergic neurons by tachykinins through NKR in the rat retina (A17). PMID- 11121539 TI - Transfection of the plasma-type platelet-activating factor acetylhydrolase gene attenuates glutamate-induced apoptosis in cultured rat cortical neurons. AB - Using an adenoviral vector, we induced overexpression of the plasma type of platelet-activating factor acetylhydrolase in cultured rat neurons. Neurons overexpressing this enzyme showed a decrease in glutamate-induced injury, mainly, apparent as decreased apoptosis. Reduction of lipid peroxidation by this enzyme and protection of mitochondrial function were demonstrated, and these may be the basis of the resistance to glutamate-induced neuronal injury that we observed. PMID- 11121540 TI - Autoradiographic analysis of dopamine receptor-stimulated [(35)S]GTPgammaS binding in rat striatum. AB - Autoradiographic analysis of [(35)S]GTPgammaS binding was used to investigate functional activation of dopamine receptors in rat striatum. Dopamine-stimulated [(35)S]GTPgammaS binding was observed with a maximal increase of 38% over basal activity. A similar stimulatory response was obtained with the D(2) agonist quinpirole, but not SKF-238393, a D(1) agonist. The effect of dopamine was blocked by the D(2) antagonist raclopride, but was unaffected by SCH-23990, a D(1) antagonist. There appeared to be a differential distribution of dopamine stimulated [(35)S]GTPgammaS binding, with the lowest activity obtained in the medial portion of the caudal striatum. These results demonstrate, using an autoradiographic approach, (i) that dopamine stimulated [(35)S]GTPgammaS binding in the rat striatum occurs through activation of D(2) receptors, and (ii) that the effects of dopamine activation vary in different areas of the rat striatum. PMID- 11121541 TI - Immunocytochemical study on the distribution of p53 in the hippocampus and cerebellum of the aged rat. AB - A role for p53-mediated modulation of neuronal viability has been suggested by the finding that p53 expression is increased in damaged neurons in models of ischemia and epilepsy. P53 gene upregulation precedes apoptosis in many cell types, and a potential role for this molecule in apoptosis of neurons has already been demonstrated in Alzheimer's disease. Recent studies suggest that p53 associated apoptosis may be a common mechanism of cell loss in several important neurodegenerative diseases. In the present study, we examined changes in p53 immunoreactive (IR) neurons in the brains of aged rats for the first time employing immunocytochemical and in situ hybridization methods. P53-IR neurons were found in the CA1 region of hippocampus, septal region and cerebellum in the aged rats, but there was no p53-IR cell in the brains of adult rats. In the hippocampus of the aged rat, p53-IR cells predominated in the stratum oriens and pyramidal layers, while the molecular layer contained relatively few p53-IR cells. The most prominent population of immunoreactive labeling in cerebellar cortex was localised within the cell bodies of Purkinje cells and dendrites in molecular layers. Upregulation of p53 in the Purkinje cells observed in this study suggests that significant loss of Purkinje cells with aging may be regulated with several apoptosis-controlling factors including p53 and oxidative stress mechanism. Further investigations are required to establish whether direct functional relations exist between p53 and the apoptotic neuronal death in normal aging or Alzheimer brains. PMID- 11121542 TI - Isolation and rapid identification of an abundant self-peptide from class II HLA DRB1*0401 alleles induced by measles vaccine virus infection. AB - Class II HLA-DR genes play an important role in the immune response to viral antigens. The effect of measles vaccine virus (MVV) infection on the induction of self-peptides presented by HLA-DR molecules during the immune response to viral infection is poorly known. Here, we describe a strategy for isolation and rapid sequence determination of an MVV-inducible class II bound peptide from a membrane protein (Leu-13). Peptides bound to HLA-DR4 (DRB1*0401 peptide complex) were eluted from immunoaffinity-purified HLA-DR4, peptides were differentially screened by MALDI-TOF-MS and subsequently sequenced by post source decay (PSD) MALDI-TOF-MS. Human B-cells infected with MVV demonstrated an enhanced pattern of self-peptide production after MVV infection. This relatively simple analytical protocol provides a sensitive method for the direct identification of peptides associated with MHC class II DR molecules. More broadly, this same approach can be used to identify sequences of specific MVV processed peptides presented by any class II MHC DR molecule. PMID- 11121543 TI - Assessment of synthetic peptides for hepatitis A diagnosis using biosensor technology. AB - In the present work we demonstrate the application of a commercial biosensor instrument (BIACORE 1000, Biacore AB, Uppsala) for the detection of antibodies against the hepatitis A virus (HAV) in human serum samples using linear and branched synthetic peptides related to the VP3 capsid protein of HAV. We also studied the conformation of the synthetic peptides by circular dichroism (CD) in order to analyse the changes in secondary structure of the constructs that could influence their recognition by antibodies. Linear and dimeric VP3(110-121) multiple antigen peptides (MAP) were the most sensitive and appropriate for serological studies of serum from HAV infected patients using BIACORE. Immobilization of tetrameric MAPs via amine groups apparently failed to preserve the active conformation of the peptide epitope since it led to lower antibody binding compared to linear and dimeric peptides. The CD analysis showed that the tetrameric MAP constructs tend to adopt a beta-sheet structure due to intermolecular aggregation, which limits epitope accessibility. Our results demonstrate the value of biospecific interaction analysis technology using synthetic peptides for the diagnosis of acute hepatitis A. PMID- 11121544 TI - Membrane assisted isoform immunoassay. A rapid method for the separation and determination of protein isoforms in an integrated immunoassay. AB - Proteins often exist as isoforms with structural microheterogenity due to, for example, small variations in their carbohydrate structure. This can give rise to differences in biological activity. Measurements of low concentrations of such isoforms are usually performed by complicated methods, which demand discrete steps of separation (chromatographic or electrophoretic) and immunodetection. In this paper a new, highly specific and rapid (<15 min) analytical technique is described which permits the quantitative determination of several types of protein isoform. The method, which is called membrane assisted isoform immunoassay (MAIIA), is based on separation (by ion-exchange or affinity chromatography) and immunoassay detection of the protein isoforms in the same lateral flow, assisted by capillary forces in a membrane device. The technique is exemplified with a method for measuring carbohydrate-deficient isoforms of the glycoprotein transferrin, where the measured isoforms constitute a minimal part (<3%) of the total amount of transferrin. The time for the measurement was about 10 min and the correlation coefficient with an established, commercial two-step procedure, which takes 4-5 h to perform, was 0. 99. The detection limit was about 1 fmol of transferrin. The anion-exchange membrane used in the test device had the ability to separate transferrin isoforms with down to 0.1 unit difference in pI. The results indicate that the technique should be useful for rapid point-of care testing of clinically interesting charged protein isoforms. PMID- 11121545 TI - A multiple transgenic mouse model with a partially humanized activation pathway for helper T cell responses. AB - Mice expressing human CD4 and human MHC II molecules provide a valuable model both for the investigation of the immunopathogenetic role of human autoantigens and for the development of therapeutic strategies based on modulating helper T cell activation in vivo. Here we present a novel mouse model expressing HLA-DR17 (a split antigen of HLA-DR3) together with human CD4 in the absence of murine cd4 (CD4/DR3 mice). Human CD4 accurately replaces murine cd4 within T cells. In particular, the preservation of cd8(+) and CD4(+) T cell subsets distinguishes CD4/DR3 mice from other multiple transgenic models in which the alternative T cell subsets are fundamentally disturbed. Moreover, human CD4 is also faithfully expressed on antigen presenting cells such as dendritic cells and monocyte/macrophages, so that the overall transgenic CD4 expression pattern resembles very closely that of humans. HLA-DR3 expression in the thymus correlates very closely to that of mouse MHC II. In contrast, only 70% of mouse MHC II positive cells in spleen, lymph node, and peripheral blood coexpress HLA DR3. No significant bias was found with regard to particular leucocytes in this respect. The stimulation of helper T cells clearly depends on the interaction between the human transgene products, since mAbs to HLA-DR and/or CD4 completely blocked in vitro recall responses to tetanus toxoid. CD4/DR3 mice represent a partially humanized animal model which will facilitate studies of DR3-associated autoimmune responses and the in vivo determination of the therapeutic potential of mAbs to human CD4. PMID- 11121546 TI - A method for investigating the role of homotypic adhesion in lymphocyte activation. AB - B cells activated via CD40 in vitro form striking homotypic aggregates, especially in the presence of costimuli such as anti-IgM, whereas those stimulated by anti-IgM alone do not. Blocking aggregation with anti-LFA-1alpha also significantly inhibits CD40-stimulated B cell proliferation, suggesting that homotypic adhesion is important for B cell activation via this receptor. To investigate this we have developed a culture system where murine B cells are stimulated in semi-solid agarose, which prevents cell-cell interactions. B cells respond to various mitogenic stimuli, including anti-CD40, in an essentially normal fashion when cultured in agarose. Furthermore, anti-LFA-1 exerts similar inhibitory effects on B cell proliferation regardless of whether the cells are in liquid, or semi-solid medium. These results indicate that homotypic aggregation is not necessary for CD40-stimulated B cell proliferation and the inhibitory effects of anti-LFA-1 could, therefore, be due to the delivery of a negative signal via this integrin, rather than as a result of inhibition of B cell clustering. Furthermore, reaggregation experiments indicated that anti-IgM stimulated B cells are attracted into anti-CD40-generated clusters, even though they do not form clusters themselves. Taken together these results indicate that clustering is a consequence of B cell activation via CD40, rather than a necessary prelude to B cell proliferation. We postulate that homotypic aggregation may involve an unknown B cell-derived chemokine. PMID- 11121547 TI - Natural anti-carbohydrate IgM in mice: dependence on age and strain. AB - Natural anti-carbohydrate antibodies in humans play a key role in natural immunity and in recognition of allogeneic and xenogeneic antigens. Presumably, natural anti-carbohydrate antibodies in mice have similar functions; but these antibodies have not been extensively characterized. An assay was developed and used to screen for anti-carbohydrate IgM in the serum of BDF-1 mice. Among the natural anti-carbohydrate IgM identified, anti-betaGlcNAc IgM were the most abundant. Anti-betaGlcNAc IgG was not detected. Levels of anti-betaGlcNAc IgM were very low in 3-week-old BDF-1 mice and increased until 5 to 7 months of age. Levels of serum anti-betaGlcNAc IgM similar to those in BDF-1 mice were found in the serum of some strains related to the BDF-1 strain (DBA and C57BL/6) and in BDF mice lacking the galactosyl transferase gene. However, in two strains unrelated to the BDF-1 strain (FVB and SJL), levels of anti-betaGlcNAc IgM were less than one-tenth of those found in BDF-1 mice. These results provide considerable insight into the effect of age on the production of natural anti carbohydrate antibodies in mice and indicate that production of those antibodies is strongly dependent on the strain of mouse. These studies will help in future development of murine models for studying the biological and medical roles of natural anti-carbohydrate antibodies. PMID- 11121548 TI - Trace detection of explosives using a membrane-based displacement immunoassay. AB - A compact membrane-based displacement immunoassay has been designed for rapid detection of explosive compounds 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5 trinitro-1,3,5-triazine (RDX) at high femtomole levels. The system consists of activated porous membranes, onto which either TNT or RDX antibodies are immobilized, that are inserted into microreactor columns, incorporated into a flow system. The assay is prepared by saturating the immobilized antibody binding sites with labeled antigen. Target analyte is introduced upstream of the microreactor, while the displacement of labeled antigen is monitored downstream using a fluorometer. The concentration of displaced labeled antigen detected is proportional to the concentration of the target analyte introduced into the system. This system provides a reusable and reagentless sensor, suitable for continuous monitoring of explosives, with an operating lifetime of over 50 positive samples. Multiple assays were performed in approximately 5 min at different flow rates, using membranes saturated with varying antibody concentrations. The membrane-based format exhibited a detection limit of approximately 450 fmol for TNT and RDX (100 microl of 1 ng/ml solution) in laboratory samples. PMID- 11121549 TI - Rapid quantitation of proinflammatory and chemoattractant cytokine expression in small tissue samples and monocyte-derived dendritic cells: validation of a new real-time RT-PCR technology. AB - The analysis of cytokine profiles plays a central part in the characterization of disease-related inflammatory pathways and the identification of functional properties of immune cell subpopulations. Because tissue biopsy samples are too small to allow the detection of cytokine protein, the detection of mRNA by RT-PCR analysis is often used to investigate the cytokine milieu in inflammatory lesions. RT-PCR itself is a qualitative method, indicating the presence or absence of specific transcripts. With the use of internal or external standards it may also serve as a quantitative method. The most widely accepted method is quantitative competitive RT-PCR, based on internal shortened standards. Recently, online real-time PCR has been introduced (LightCycler), which allows quantitation in less than 30 min. Here, we have tested its use for the analysis of cytokine gene expression in different experimental in vitro and ex vivo settings. First, we compared quantitative competitive RT-PCR with real-time RT-PCR in the quantitation of transcription levels of the CD4(+) cell-specific chemoattractant Interleukin-16 during the maturation of monocyte-derived dendritic cells, and found a good correlation between both methods. Second, differences in the amounts of IL-16 mRNA in synovial tissue from patients with rheumatoid arthritis and osteoarthritis as assessed by real-time RT-PCR paralleled differences in the level of IL-16 protein in the synovial fluid. Finally, we employed real-time RT PCR to study the cutaneous expression of several cytokines during experimental immunomodulatory therapy of psoriasis by Interleukin-10, and demonstrate that the technique is suitable for pharmacogenomic monitoring. In summary, real-time RT PCR is a sensitive and rapid tool for quantifying mRNA expression even with small quantities of tissue. The results obtained do not differ from those generated by quantitative competitive RT-PCR. PMID- 11121550 TI - Novel fusion proteins in the analysis of diabetes-associated autoantibodies to GAD65 and IA-2. AB - Assays to detect autoantibodies to glutamic acid decarboxylase (GAD65) and the protein tyrosine phosphatase-like molecule IA-2, which are both present in pancreatic islets, have been used in the diagnosis and prediction of type 1 diabetes. In this study a novel fusion protein combining the entire GAD65 molecule with the 40 kDa intracellular domain of IA-2 (GAD-IA-2) was constructed to detect autoantibodies to both antigens by one single assay. For the same purpose a truncated version of this fusion protein which contained the entire GAD65 linked to the 203 carboxy-terminal amino acids of IA-2 (GAD-dIA-2) was made. A panel of 34 diabetic sera which represented unequivocally positive or negative antibody responses to GAD65 and/or IA-2 as well as 20 serum samples from healthy controls were tested in a radioligand binding assay with the constructed fusion proteins as antigens. Nine of the samples from patients with type 1 diabetes reacted with GAD65 while being negative for IA-2. Six sera were positive for IA-2 only, 11 were double positive, and 8 negative for both antibodies using the standard in vitro transcription translation assay with single antigens. The full-length, as well as the truncated fusion protein detected all samples positive for antibodies either to GAD65 or IA-2 or both, except for one GAD65 antibody positive sample. All samples from healthy controls tested negative in all assays. We conclude that the principle of a combinatorial molecule where a fusion protein expresses both GAD65 and IA-2 epitopes is feasible, and such a fusion protein can be used instead of the single antigens to reduce time and costs of large-scale screening for clinical purposes. PMID- 11121551 TI - Expression of a human, neutralizing monoclonal antibody specific to puumala virus G2-protein in stably-transformed insect cells. AB - We cloned the heavy- and light-chain antibody genes of a human X (humanxmouse) trioma secreting a neutralizing, IgG monoclonal antibody to the G2-protein of Puumala virus. The antibody genes were inserted separately into plasmid transfer vector pIEI-4 such that the genes were under control of the baculovirus immediate early gene promoter, IEI. Trichoplusia ni (TN) cells were co-transfected with these constructs and a selection plasmid containing a neomycin-resistance gene. Cloned transformants expressing the IgG monoclonal antibody were identified by ELISA of transfected TN cell culture supernatants. TN cell lines were established from four selected clones, of which one was chosen for detailed analysis. Specificity of the insect cell-expressed human antibody was determined by ELISA with Puumala virus-infected cell lysates and by immune-precipitation of radiolabeled Puumala virus proteins. The expressed IgG retained the ability to neutralize Puumala virus in plaque-reduction neutralization assays. Using competitive polymerase chain reaction methods, multiple copies of integrated heavy- and light-chain antibody genes were detected in the insect cell genome. The transformed insect cells were stable and continuously expressed biologically active IgG. We conclude that this methodology provides an alternative eukaryotic source for the generation of human antibodies. PMID- 11121552 TI - Dendritic cell elimination as an assay of cytotoxic T lymphocyte activity in vivo. AB - We show in this paper that the survival of antigen-loaded dendritic cells in vivo may be used as a sensitive readout of CTL activity. We have previously shown that dendritic cells labeled with the fluorescent dye CFSE and injected sub cutaneously into mice migrate spontaneously to the draining lymph node where they persist for several days. In the presence of effector CTL responses, dendritic cells loaded with specific antigen rapidly disappear from the draining lymph node. In this paper we extend the above observations and set up a simple and sensitive method to reveal CTL activity in individual mice in vivo. Dendritic cells were labeled with two different fluorochromes, loaded with antigen or left untreated, and mixed together before injection into mice. We show that only the dendritic cells loaded with specific antigen were cleared from the draining lymph node, while dendritic cells not loaded with antigen remained unaffected. Cytotoxic responses generated by immunization with peptide-loaded dendritic cells, or by infection with influenza virus, could be revealed using this method. Comparison of the differential survival of dendritic cells populations mixed together also allowed us to accurately evaluate the disappearance of dendritic cells, irrespective of variability in the injection site and other parameters. Given the ability of dendritic cells to efficiently take up and present complex antigens, nucleic acids and apoptotic bodies, this method may also allow the evaluation of cytotoxic activity against antigens that are not characterized in terms of peptide epitopes. PMID- 11121553 TI - Flow immunochemical bio-recognition detection for the determination of interleukin-10 in cell samples. AB - On- and off-line heterogeneous non-competitive flow immunoassays for the determination of Interleukin-10 are described. The sample containing IL-10 is mixed, either on-line in a reaction coil or off-line in a test tube, with fluorescent labelled anti-IL-10 antibodies to form an antibody-antigen complex. The labelled unbound antibodies are trapped on an immobilized IL-10 column whereas the IL-10-antibody complexes are eluted and detected downstream by a fluorescence detector. The optimization of the systems was performed with respect to choice of affinity support, flow rate, carrier buffer additives, pH and antibody-antigen association. Both bio recognition assays were tested with a spiked cell medium and the IL-10 detection limits in this matrix was found to be 8 fmol using the off-line incubation mode and 40 fmol using the on-line incubation mode. The sample through-put was 26 and 40 samples per hour in the on line and off-line incubation modes, respectively. IL-10 identification in the sample fractions was achieved using MALDI-TOF MS. PMID- 11121554 TI - Rapid identification of local T cell expansion in inflammatory organ diseases by flow cytometric T cell receptor Vbeta analysis. AB - Oligoclonal expansion of antigen-specific T cells occurs frequently during inflammatory diseases. These cells may persist for a long time at high frequency in the body and be enriched in the affected tissues. As a screening test for expanded cell T cell populations at sites of inflammation, we developed an optimized methodology for flow-cytometry-based quantification of T cell receptor Vbeta (TCRBV) expression. We first validated the specificity of a TCRBV-specific monoclonal antibody set by direct comparison with PCR-based analysis of mono- and polyclonal T cell samples. This monoclonal antibody (mAb) panel recognized approximately two thirds of the T cell receptor alpha/beta repertoire in a group of 64 healthy donors and allowed defining TCR usage in the CD4+ and CD8+ subsets. The reliable detection of expanded Vbeta gene families in T cell populations was confirmed in experiments on superantigen-stimulated T cells. Through differential TCR analysis on T cell subpopulations in cerebrospinal fluid and blood in patients with acute encephalitis, we were able to identify locally expanded CD8+ T cells. The power of this approach affords not only high-throughput comparative TCR analysis for immunological studies in vitro, but also rapid ex vivo identification of cell populations enriched in organ compartments during inflammatory diseases. PMID- 11121555 TI - Identification of the T-cell receptor alpha variable (TRAV) gene(s) in T-cell malignancies. AB - Due to the lack of a complete range of monoclonal antibodies (mAb) it is often impossible to rapidly identify by flow cytometry the T-cell receptor variable genes in patients suffering from T-cell malignancies. This applies especially to the alpha variable genes (TRAV), since only very few anti-TcR variable alpha mAb are available. We describe a very rapid method for inverse PCR amplification of the TcR alpha chain without prior purification of the double-stranded cDNA, provide the sequences for appropriate oligonucleotides, and describe a buffer system that dramatically enhances the amplification efficiency as compared to standard conditions. PMID- 11121556 TI - Expression and purification of antigenically active soluble derivatives of the heterodimeric and homodimeric forms of the mouse CD8 lymphocyte membrane glycoprotein. AB - The T lymphocyte membrane glycoprotein CD8 enhances antigen recognition by class I-restricted T cells. There are two naturally occurring forms of CD8, an alphabeta heterodimer expressed by the majority of CD8(+) T cells, and a less abundant alphaalpha homodimer found on specialised T cell subsets. An expression strategy was developed for production of soluble CD8alphaalpha and CD8alphabeta extracellular domains for use in ligand binding studies. Mouse CD8alpha was expressed autonomously as a homodimer at 10 mg/l in mammalian fibroblasts, but CD8beta was not expressed at significant levels in the absence of CD8alpha. Co expression with CD8alpha led to significant enhancement in the level of CD8beta expression, which was secreted as a non-covalent heterodimer at 3 mg/l with CD8alpha. Despite the marked increase of CD8beta expression in the presence of CD8alpha, an excess of soluble CD8alphaalpha homodimer was also present in the supernatant of co-expressing cell clones. In order to resolve the CD8alphaalpha homodimer from the CD8alphabeta heterodimer, affinity chromatographic techniques specific for the CD8beta subunit were employed. Purification procedures requiring elution from affinity matrices at low pH led to substantial losses in the total antigenic activity and partial subunit dissociation of the soluble CD8alphabeta heterodimer. The inclusion of a hexahistidine tag at the C-terminus of CD8beta enabled affinity purification of soluble CD8alphabeta (and sCD8alphaalpha) under neutral conditions, yielding recombinant protein with the correct stoichiometry and full antigenic activity. This method may prove useful for production of other soluble recombinant heterodimeric receptor proteins whose antigenicity is affected by denaturation during immunoaffinity purification. PMID- 11121557 TI - A stable marker for specific T-cells: a TCR alpha/green fluorescent protein (GFP) fusionprotein reconstitutes a functionally active TCR complex. AB - The detection of antigen specific clonal T-cell populations in vivo during T-cell selection and an immune responses is often hampered due to the lack of suitable clonotype specific monoclonal antibodies. In order to determine the potential usefulness of green fluorescent protein (GFP) to follow specific T-cells in vivo, we decided to express and analyze the function of a T-cell receptor (TCR) alpha chain-GFP fusionprotein. The TCRalpha and beta chain cDNAs of a Leishmania major specific murine T helper 2 cell clone were cloned and inserted into the pHSE3' expression vector. Simultaneously, a TCRalpha expression vector was constructed containing a C-terminal in frame fusion with the open reading frame of the enhanced GFP (EGFP). TCRalpha/TCRbeta or TCRalpha-EGFP/TCRbeta constructs were expressed in T-cell hybridoma cells 58alpha(-)beta(-) which lack an endogenous TCR but still express CD3 components. The TCRalpha-EGFP fusionprotein was detected with the expected molecular weight by immunoprecipitation and Western Blot analysis. Surface staining of TCR components was detected in transfectants expressing the wild type TCR heterodimer and, with only a slight reduction in intensity, also in those expressing the TCR-EGFP complex. Hence, expression and transport to the outer cell membrane is possible despite the 27 kD C-terminal extension of the TCRalpha. Most importantly, the EGFP-tagged TCR was functional since the transfectants produced IL-2 in response to stimulation via their TCR. Thus, TCR-EGFP constructs represent attractive tools to study posttranslational regulation of TCR expression and ligand-induced TCR clustering as well as the fate of antigen specific T-cells during tolerance induction and immunity in transgenic mouse models. PMID- 11121558 TI - Immunophenotyping of peripheral blood leukocytes by laser scanning cytometry. AB - Many clinical situations demand repeated analyses of blood parameters but permit only minimal amounts of peripheral blood to be taken, e.g., in neonates with low birth weight, during extensive operations of young children, or in patients with restricted bone marrow function. In these cases laser scanning cytometry is the ideal tool to determine the distribution of different leukocyte-subsets. The purpose of this protocol is to describe stepwise a new method of immunophenotyping by laser scanning cytometry. In this assay nuclear DNA is stained by 7-aminoactinomycin-D (7-AAD) and surface antigens are detected by direct three-colour immunofluorescence. For data acquisition, measurements are triggered on the 7-AAD-fluorescence. Data are obtained for forward scatter, green, orange, and long red fluorescence by excitation with the argon-laser, and for far red fluorescence by excitation with the helium-neon-laser. Using this protocol the amount of peripheral blood needed is minimised to 10 microl. Specimens can be stained a second time in a different way and analysed repeatedly and archived. PMID- 11121559 TI - Preparation of human-mouse heterohybridomas against an immunising antigen. AB - The production of murine monoclonal antibodies against specific antigens by hybridomas is a well utilised technique. The production of hybridomas secreting specific human antibodies would have many advantages in therapeutic applications of monoclonal antibodies. The immortalised human lymphocytes themselves would also provide valuable tools in research on lymphocyte development. Preparation of human-human hybridomas has been limited by a lack of suitable fusion partners. This protocol paper describes the production of human-mouse heterohybridomas by two independent laboratories. The purpose of this protocol is to provide a basis for the development of heterohybridoma technology in laboratories with limited hybridoma experience. PMID- 11121560 TI - Generation of phosphorylation state-specific SRC-class kinase antibodies for analysis of kinase activation. AB - Protein phosphorylation is a major molecular mechanism by which cellular function is regulated. In order to accomplish rapid and specific biochemical changes via phosphorylation, the activity of a protein kinase must be dynamically regulated. Historically, the activity of each protein kinase has been analyzed using a unique in vitro biochemical assay with a specific substrate and detection procedure. These assays require the use of radioactivity and are often labor intensive. Upon activation, most protein kinases autophosphorylate. Thus, a technical approach to detect changes in kinase activity is to measure autophosphorylation. The purpose of this protocol is to provide a detailed stepwise procedure for measuring the regulation of Src-class kinase activity using phosphorylation state-specific antibodies. Antibodies to a phosphorylated peptide derived from the autophosphorylation site of Src-family kinases are developed and affinity purified. The purified antibodies are used to analyze the regulation of Src and Fyn activity in a mouse muscle cell line. It is anticipated that the utility of these phosphorylation state-specific antibodies will ultimately result in the development of similar antibodies useful for analyzing the activity of many different kinases. PMID- 11121561 TI - The transmission dynamics of the aetiological agent of scrapie in a sheep flock. AB - We formulate and investigate the properties of a model framework to mimic the transmission dynamics of the aetiological agent of scrapie in a sheep flock. We derive expressions for summary parameters that characterize transmission scenarios, notably the basic reproduction number R(0) and the mean generation time T(g). The timescale of epidemic outbreaks is expressed in terms of R(0) and cumulants of the generation time distribution. We discuss the relative contributions to the overall rate of transmission of horizontal and vertical routes during invasion and in endemicity. Simplified models are used to obtain analytical insight into the characteristics of the endemic state. PMID- 11121562 TI - Extensions to a procedure for generating locally identifiable reparameterisations of unidentifiable systems. AB - In this paper extensions to an existing procedure for generating locally identifiable reparameterisations of unidentifiable systems are presented. These extensions further formalise the constructive nature of the methodology and lend themselves to application within symbolic manipulation packages. The extended reparameterisation procedure is described in detail and is illustrated with application to two known non-trivial examples of unidentifiable systems of practical relevance. PMID- 11121563 TI - A note on integrals for birth-death processes. AB - A general integral for birth-death Markov processes is considered. A closed form expression is provided for its expected value in terms of the birth and death rates. A simple route for numerical evaluation of its variance is also suggested. PMID- 11121564 TI - A model for hepatocarcinogenesis with clonal expansion of three successive phenotypes of preneoplastic cells. AB - The two-stage model with clonal expansion of intermediate cells has often been used to describe the carcinogenesis process. The model hypothesizes that cells have to undergo two mutations on their way from the normal to the malignant stage. Biological experiments indicate the existence of three types of preneoplastic cells in hepatocarcinogenesis representing three successive intermediate stages in the development of malignant cells from normal cells. This finding suggests that hepatocarcinogenesis should be described by a multi-stage model with three intermediate stages, leading to a four-stage mutation model with clonal expansion of all types of intermediate cells. This model is presented and mathematical approximations for the number and size of nonextinct premalignant clones of the different cell types are derived. The model is applied to focal lesion data from a rat hepatocarcinogenesis experiment. PMID- 11121565 TI - Optimal control for a stochastic model of cancer chemotherapy. AB - Chemotherapy is useful in a number of cancers to reduce or eliminate residual disease. When used in this way the objective is to maximise the likelihood that the cancer will be eliminated. In this article, we extend a stochastic model of chemotherapy for cancer to incorporate its concomitant effect on the normal system and derive overall measures of outcome. The model includes the development of drug resistance and is sufficiently flexible to include a variety of tumour and normal system growth functions. The model is then applied to situations previously examined in the literature and it is shown that early intensification is a common feature of successful regimens in situations where drug resistance is likely. The model is also applied to data collected from clinical trials analysing the effect of adriamycin, and cyclophosphamide, methotrexate and 5 flourouracil (CMF) therapy in the treatment of operable breast cancer. The model is able to mimic the data and provides a description of the optimal regimen. PMID- 11121566 TI - The stage-structured predator-prey model and optimal harvesting policy. AB - In this paper, we establish a mathematical model of two species with stage structure and the relation of predator-prey, to obtain the necessary and sufficient condition for the permanence of two species and the extinction of one species or two species. We also obtain the optimal harvesting policy and the threshold of the harvesting for sustainable development. PMID- 11121567 TI - Unraveling the function of GABA(A) receptor subtypes. PMID- 11121568 TI - Gastric habitation by Helicobacter pylori: insights into acid adaptation. PMID- 11121569 TI - Do glucocorticoids enhance eosinopoiesis? PMID- 11121570 TI - Kinin B1 receptors, knockout mice and nociception: are kinins sensory transmitters? PMID- 11121571 TI - Benzodiazepines outside the CNS. PMID- 11121572 TI - Platelets and P2X1: is ADP really an agonist? PMID- 11121573 TI - Beta3-adrenoceptors in the cardiovascular system. AB - beta-Adrenoceptors of the beta1 and beta2 subtypes classically mediate the effects of catecholamines on the contractility of cardiac muscle and the relaxation of vascular smooth muscle. Since the molecular characterization of the beta3-adrenoceptor in 1989, most studies of this adrenoceptor subtype have focused on its control of lipolysis in adipose tissues. However, more recent studies have investigated the involvement of beta3-adrenoceptors in the physiological control of cardiac and vascular contractility. In this article, the pharmacological and molecular evidence that supports the functional role of beta3 adrenoceptors in cardiovasculartissues of various species, including humans, will be discussed. These data might provide new insights into our understanding of the abnormal responsiveness of the cardiovascular system to catecholamines in heart failure and its treatment with beta3-adrenoceptor antagonists. PMID- 11121574 TI - The molecular pharmacology of CGRP and related peptide receptor subtypes. AB - Calcitonin gene-related peptides (alpha and beta isoforms), better known as CGRPalpha and CGRPbeta, were isolated twenty years ago. In fact, these were the first peptides to be characterized using a molecular cloning strategy, which is not the traditional approach of biochemical extraction and purification. Paradoxically, progress in the characterization of CGRP receptor subtypes has been extremely slow as a result of difficulties in their cloning and the lack of selective receptor subtype agonists and antagonists. However, exciting progress has been made overthe pasttwo years and is briefly reviewed here. PMID- 11121575 TI - New windows on the mechanism of action of K(ATP) channel openers. AB - K(ATP) channel openers are a diverse group of drugs with a wide range of potential therapeutic uses. Their molecular targets, the K(ATP) channels, exhibit tissue-specific responses because they possess different types of regulatory sulfonylurea receptor subunits. It is well recognized that complex interactions occur between K(ATP) channel openers and nucleotides, but the cloning of the K(ATP) channel has introduced a new dimension to the study of these events and has furthered our understanding of the molecular basis of the action of K(ATP) channel openers. PMID- 11121576 TI - Hinges, swivels and switches: the role of prolines in signalling via transmembrane alpha-helices. AB - Extracellular signals are transduced across membranes via conformational changes in the transmembrane domains (TMs) of ion channels and G-protein-coupled receptors (GPCRs). Experimental and simulation studies indicate that such conformational switches in transmembrane (alpha-helices can be generated by proline-containing motifs that form molecular hinges. Computational approaches tested on model channel-forming peptides (e.g. alamethicin) reveal functional mechanisms in gap-junction proteins (such as connexin) and voltage-gated K+ channels. Similarly, functionally important roles for proline-based switches in TM6 and TM7 were identified in GPCRs. However, hinges in transmembrane helices are not confined to proline-containing sequence motifs, as evidenced by a non proline hinge in the M2 helix of the nicotinic acetylcholine receptor. This helix lines the pore and plays a key role in the gating of this channel. PMID- 11121577 TI - Proteins S7, S10, S16 and S19 of the human 40S ribosomal subunit are most resistant to dissociation by salt. AB - The protein components of human 40S ribosomal subunits were dissociated by centrifugation in gradients of sucrose and LiCl in the presence of 0.5 M KCl. The proteins that split off were analyzed by SDS-PAGE and 2D-PAGE. The order of dissociation of the proteins, depending on the salt concentration (from 0.8 M to 1.55 M), was established. The majority of the proteins started to split off simultaneously at a monovalent cation concentration of 0.8 M. Ten proteins were found to be more resistant; of these proteins S7, S10, S16, and S19 were retained most strongly and thereby may be considered to be core proteins. PMID- 11121578 TI - The ruv proteins of Thermotoga maritima: branch migration and resolution of Holliday junctions. AB - In homologous recombination in bacteria, the RuvAB Holliday junction-specific helicase catalyzes Holliday junction branch migration, and the RuvC Holliday junction resolvase catalyzes formation of spliced or patched structures. RuvAB and RuvC from the hyperthermophile Thermotoga maritima were expressed in Escherichia coli and purified to homogeneity. An inverted repeat sequence with unique termini was produced by PCR, restriction endonuclease cleavage, and head to-tail ligation. A second inverted repeat sequence was derived by amplification of a second template containing a three-nucleotide insertion. Reassociation products from a mixture of these two sequences were homoduplex linear molecules and heteroduplex heat-stable Holliday junctions, which acted as substrates for both T. maritima RuvAB and RuvC. The T. maritima RuvAB helicase catalyzed energy dependent Holliday junction branch migration at 70 degrees C, leading to heteroduplex linear duplex molecules with two three-nucleotide loops. Either ATP or ATP gamma S hydrolysis served as the energy source. T. maritima RuvC resolved Holliday junctions at 70 degrees C. Remarkably, the cleavage site was identical to the preferred cleavage site for E. coli RuvC [(A/T)TT(downward arrow)(G/C)]. The conservation of function and the ease of purification of wild-type and mutant thermophilic proteins argues for the use of T. maritima proteins for additional biochemical and structural studies. PMID- 11121579 TI - Cloning and functional studies of a luxO regulator LuxT from Vibrio harveyi. AB - LuxO is the central regulator integrating the quorum sensing signals controlling autoinduction of luminescence in Vibrio harveyi. We have previously purified to homogeneity a new lux regulator, LuxT, that binds to the luxO promoter. Based on the sequence of the tryptic peptides of LuxT, degenerate oligonucleotides were designed for PCR of the genomic DNA. A 273 bp PCR DNA fragment containing sequences encoding the tryptic peptides was extended by inverse PCR to obtain the complete gene (luxT) encoding a protein of 153 amino acids which shares homology with the AcrR/TetR family of transcriptional regulators. The recombinant and native LuxT gave the same footprint binding between 117 and 149 bp upstream from the luxO initiation codon. Gene disruption of luxT in V. harveyi increased luxO expression and affected the cell density dependent induction of luminescence showing that LuxT was a repressor of luxO. As LuxT also affected the survival of the V. harveyi cells at high salt concentration and homologous proteins are present in other bacterial species, including the pathogen, Vibrio cholerae, the LuxT regulatory protein appears to be a general rather than a lux-specific regulator. PMID- 11121580 TI - Differential binding and transcriptional behaviour of two highly related orphan receptors, ROR alpha(4) and ROR beta(1). AB - Nuclear receptors are ligand-inducible transcription factors that can be classified into two major groups according to their DNA-binding properties. Members of the first group bind to DNA as dimers, either homo- or heterodimers; members of the second group are also able to bind as monomers. While the first group has been extensively studied biochemically, very little is known about nuclear receptors that bind and act as monomers. In this study, we compared the binding and transcriptional behaviour of ROR alpha (NR1F1) and ROR beta (NR1F2), two representatives of the subgroup of monomer-binding receptors. We show that although they are highly related in their amino acid structures, they display remarkably different binding behaviours. Furthermore, we provide evidence that ROR beta can efficiently activate transcription in vitro as a monomer. PMID- 11121581 TI - Identification of candidate growth-regulating genes that are overexpressed in late gestation fetal liver in the rat. AB - We have shown previously that hepatocyte proliferation in the late gestation fetal rat is mediated by growth factor-independent mechanisms that are distinct from the signaling pathways that promote proliferation of adult rat hepatocytes. In the present studies, we identified six candidate growth-regulating genes that are overexpressed in fetal rat liver (embryonic day 19, 2 days pre-term) relative to adult rat liver using suppressive subtractive hybridization. These included the following: Grb10, a growth factor receptor binding protein; eps15, a growth factor receptor substrate; nuc2+, a retinoblastoma protein binding protein; cdc25B, a cell cycle tyrosine phosphatase; the peroxisome proliferator-activated receptor PPAR alpha; and a deoxyuridine triphosphatase that functions as a PPAR alpha binding partner. In every case, the ontogeny of the expression of these genes declined postnatally in a manner consistent with the transition from a fetal to an adult hepatocyte phenotype. None were found to be cell cycle dependent, in that they did not show expression that followed perinatal changes in hepatocyte cell cycle activity. Based on our identification of these genes and previous work characterizing their role in growth regulation, we conclude that they may contribute to the mitogenic signaling phenotype of fetal rat hepatocytes. PMID- 11121582 TI - Regulation of a recombinant pea nuclear apyrase by calmodulin and casein kinase II. AB - A cDNA encoding a pea nuclear apyrase was previously cloned. Overexpressions of a full-length and a truncated cDNA have been successfully expressed in Escherichia coli BL21(DE3). The resulting fusion proteins, apyrase and the C-terminus (residues 315-453) of apyrase, were used for calmodulin (CaM) binding and phosphorylation studies. Fusion protein apyrase but not the C-terminus of apyrase can be recognized by polyclonal antibody pc480. This suggested that the motif recognized by pc480 was located in the N-terminal region of apyrase. The recombinant apyrase protein also showed an activity 70 times higher than that of endogenous apyrase using ATP as a substrate. The recombinant apyrase has a preference for ATP more than other nucleoside triphosphate substrates. CaM can bind to recombinant apyrase, but not to the C-terminus of apyrase. This implies that the CaM-binding domain must be in the first 315 amino acids of the N terminal region of apyrase. We found that one segment from residue 293 to 308 was a good candidate for the CaM-binding domain. This segment 293 FNKCKNTIRKALKLNY 308 has a basic amphiphilic-helical structure, which shows the predominance of basic residues on one side and hydrophobic residues on the other when displayed on a helical wheel plot. Using the gel mobility shift binding assay, this synthetic peptide was shown to bind to CaM, indicating that it is the CaM-binding domain. Both recombinant apyrase and the C-terminus of apyrase can be phosphorylated by a recombinant human protein kinase CKII. Phosphorylation does not affect CaM binding to recombinant apyrase. However, CaM does inhibit CKII phosphorylation of recombinant apyrase and this inhibition can be blocked by 5 mM EGTA. PMID- 11121583 TI - Regulation of rat heat shock factor 2 expression during the early organogenic phase of embryogenesis. AB - A central step in the transcriptional regulation of heat shock protein (hsp) genes is the binding of the heat shock factor (HSF) to the upstream heat shock elements (HSEs). In vertebrates, HSF2 has been suggested to mediate the transcriptional regulation of hsp gene expression during development and differentiation. The expression levels of HSF2 were shown to vary widely among fully developed mouse organs. However, there exists limited information on the regulation of HSF2 expression during the inductive stage of organ formation in mammalian development. In this study, we have cloned the rat HSF2 cDNA and examined embryos for HSF2 expression from days 9.5 (E9.5) to 15.5 (E15.5) of gestation that correspond to the period when the major organ primordia are being actively established. We show that rat HSF2 has 94.6 and 96.3% identity to mouse HSF2 in nucleotide and amino acid sequences, respectively. By establishing a competitive RT-PCR, we show that about 503.6 pg of HSF2 mRNA were present per microgram of embryonic RNA in the primitive streak stage E9.5 embryos. The amounts of HSF2 mRNA then gradually decreased, resulting in an approximately 300 fold reduction in E15.5 embryos. The amounts of HSF2 mRNA in the embryos were found to be closely correlated with those of HSF2 protein and their HSE-binding activities. To our knowledge, this is the first detailed report on the structure and regulation of the rat HSF2 during the early organogenic period of mammalian embryogenesis. PMID- 11121584 TI - 2',5'-oligoadenylate synthetase gene in chicken: gene structure, distribution of alleles and their expression. AB - We have cloned the gene for chicken 2',5'-oligoadenylate synthetase (ChOAS) by the method of polymerase chain reaction with use of ChOAS cDNA sequence. The ChOAS gene is composed of five introns and six exons containing all of the sequence of the ChOAS cDNA from the start to the stop codon. The first five exons of ChOAS gene which encode the OAS catalytic domain have a similar structure to HuOAS1 gene including the exon-intron boundaries. However, the length of introns of ChOAS gene is only 1/7 of those of HuOAS1 gene. The sixth exon of the ChOAS gene encodes the ubiquitin-like (UbL) domain of two consecutive sequence (UbL1 and UbL2) homologous to ubiquitin. ChOAS encoded in a single copy gene has at least two alleles, OAS(*)A and OAS(*)B. The differences between these two alleles are in the sixth exon of the gene; a 96-nucleotide sequence in the UbL1 portion of OAS(*)A is deleted from OAS(*)B. No OAS(*)B gene was detected in nine lines of chickens tested other than Leghorns. Almost the same levels of ChOAS-A and -B proteins induced physiologically in erythrocytes were detected in infant chickens (2-week-old), but in grown-up chickens (6-month-old) the level of erythrocyte OAS B was markedly reduced in most of B/B chickens. Thus, the UbL domain of ChOAS is responsible for the maintenance of the OAS level in the tissue. PMID- 11121585 TI - Identification of a nitric oxide-regulated zinc finger containing transcription factor using motif-directed differential display. AB - We report here the isolation of human zinc finger 2 (HZF2), a putative zinc finger transcription factor, by motif-directed differential display of mRNA extracted from histamine-stimulated human vein endothelial cells. The expression of HZF2 mRNA in venous endothelial cells was verified by Northern blot analysis, which also revealed an enrichment of HZF2 mRNA in lymphocytes and monocytes. Histamine induced a time- and concentration-dependent upregulation of HZF2 level with a 6-fold peak increase of mRNA at 30 min. HZF2 upregulation was abolished by different NOS isozyme inhibitors. Guanylate cyclase inhibition resulted in a significant decrease of HZF2 expression. These observations indicate HZF2 as a potentially interesting new target for studies concerning rapid NO-mediated gene regulation. PMID- 11121586 TI - Molecular characterization of Drosophila melanogaster myo-inositol-1-phosphate synthase. AB - We have isolated and characterized a cDNA encoding Drosophila melanogaster myo inositol-1-phosphate synthase (INOS). The deduced Drosophila INOS protein is 50% identical to the Saccharomyces cerevisiae INO1 gene. The putative active site residues are well conserved in Drosophila INOS protein. Southern blot analysis shows that Drosophila INOS gene is a single copy gene. Northern blot analysis reveals that Drosophila INOS gene expresses a 2.0-kb transcript that is more abundant in the head than the body, suggesting that it may be involved in brain function. The recombinant Drosophila INOS protein was expressed in Escherichia coli and the purified protein has proved to have a myo-inositol-1-phosphate synthase activity. PMID- 11121587 TI - Alternative splicing of the human PTEN/MMAC1/TEP1 gene. AB - The human tumour suppressor gene PTEN/MMAC1/TEP1 encodes a lipid and protein phosphatase. Using RT-PCR, alternatively spliced forms of PTEN mRNA, encoding full-length PTEN and two forms of the protein truncated at the C-terminal end, were detected in normal human tissue. Cultured tumour and non-tumour cell lines show similar splicing patterns. PMID- 11121588 TI - Cloning and functional expression of the rat alpha(2) subunit of soluble guanylyl cyclase. AB - Nitric oxide-sensitive guanylyl cyclase is a heterodimeric enzyme consisting of one alpha and one beta subunit. Here, we clone the first alpha(2) subunit ortholog and functionally express the cDNA in Sf-9 cells. Our data indicate a high degree of conservation of the primary sequence and functional activity of the rat alpha(2) subunit. PMID- 11121590 TI - Pharmacogenomics: hope or hype? PMID- 11121589 TI - Cloning and characterization of glucose transporter in teleost fish rainbow trout (Oncorhynchus mykiss). AB - The facilitated diffusion of monosaccharides across the plasma membrane is mediated by glucose transporters (GLUTs). In contrast to mammals, the glucose transport system of lower vertebrates remains unexplored. We detected glucose transport activity in rainbow trout embryos. Two GLUTs sharing 83% amino acid identity were cloned from juvenile fish, these have been denoted OnmyGLUT1A and OnmyGLUT1B. In adult trout OnmyGLUT1A is predominantly expressed in the heart with low expression in other tissues. An inverse terminal repeat of a Tc1-like transposable element was found in the 3'-untranslated region of OnmyGLUT1B. Phylogenetic analysis suggested that rainbow trout genes share a common ancestor with higher vertebrate GLUT1. We also found GLUT genes in several salmonid species. PMID- 11121591 TI - Long-term outcome of coronary balloon angioplasty in diabetic patients. AB - Diabetes is recognised to increase morbidity and mortality after coronary revascularization. We compared clinical outcomes in mean 5-year-long follow-up of coronary balloon angioplasty in diabetic and non-diabetic patients. We studied 621 patients undergoing elective angioplasty from 1987 to 1996. There were 60 (9.7%) patients with diabetes who were compared with 561 non-diabetic patients. Diabetics were older, more often obese, less frequently were current smokers, and less frequently had hypercholesterolaemia. Diabetic patients in comparison with non-diabetics had lower ejection fraction and more frequently had angioplasty of complex (B2 or C) lesions, but there were no differences between both groups in the other clinical and angiographic risk factors. Clinical success of angioplasty, as well as complications rate were similar in both groups. In follow up restenosis occurred more frequently in diabetics (46.3 vs. 32.2%, P=0.03), resulting in significantly higher re-intervention rate (50.0 vs. 35.4%, P=0.03). Especially diabetic patients were more frequently referred to CABG (20.4 vs. 9. 9%, P=0.02). There were no significant differences in deaths (1.9 vs. 2.8%) and myocardial infarction (3.7 vs. 4.4%). Diabetics presented worse CCS status at the end of observation (Class 0 and I - 61.1 vs. 74.4%, P=0.037). Angioplasty proved to be a safe procedure in diabetic patients. Despite higher restenosis and re intervention rate in diabetics, mortality as well as myocardial infarction rate was the same in both groups during mean 5-year follow-up. PMID- 11121592 TI - Recent time trends in acute myocardial infarction in Stockholm, Sweden. AB - AIMS: The purpose of this study was to analyse time trends in first acute myocardial infarction (AMI) incidence in Stockholm County during 1984-1996. METHODS AND RESULTS: The study base consisted of the population of Stockholm County 30-89 years old 1984-1996. New cases of first AMI in the study base were identified by combining information regarding hospital discharges and deaths. The diagnostic quality was evaluated for 2403 First-AMI cases 45-70 years old that occurred during 1992-1994. The evaluation indicated a very high agreement with diagnostic criteria of AMI among hospital treated cases. Among fatal cases outside hospital, the autopsy rate was nearly 70%. The incidence of first AMI declined during the study period for both men and women. In men, the age-adjusted incidence of first AMI was 18% lower in 1994-1996 than in 1984-1987 and in women it was 13% lower. The average yearly decline was 2% for men and 1.4% for women. CONCLUSION: The results of this study suggest a continued decline in AMI incidence in Stockholm. A reduction in the proportion of regular smokers has probably contributed to this decline. Future trends in AMI incidence are difficult to predict in view of diverging trends in risk factors. PMID- 11121593 TI - Effects of HMG-CoA reductase inhibitor on hemostasis. AB - Clinical trials of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor (statin) therapy have demonstrated improvement in coronary atherosclerosis progression and reduction in risk of cardiovascular events. However, improvement in cardiovascular end-points is incompletely explained by the baseline or treated LDL cholesterol level. The beneficial effects of statins on clinical events may involve nonlipid mechanisms that modify hemostasis. Local activation of platelets and thrombus formation adjacent to atheromatous plaques, especially where ruptured or eroded, are now recognized to be of pathophysiological importance in the acute and chronic clinical expression of coronary heart disease. Thus, favorable effects of statins on hemostasis may be relevant to decreasing or delaying the progression and clinical manifestations of atherosclerosis. PMID- 11121594 TI - Enzymatic oxidant and antioxidants of human blood platelets in unstable angina and myocardial infarction. AB - In ischaemic heart conditions we report a remarkable increase in platelet xanthine oxidase activity and rise in the levels of malondialdehyde (MDA) with concomitant decrease in the activities of free radical scavenging enzymes - superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase. The increased levels of free radical generating system and MDA and lowered levels of free radical scavenging systems seem to have critical role in ischaemic heart conditions. PMID- 11121595 TI - Endothelin-1 is involved in the growth promotion of vascular smooth muscle cells by hyaluronic acid. AB - Proliferation of vascular smooth muscle cells (VSMC) is crucial to the progression of arteriosclerosis. In this study, we examined the role that interactions among endothelin-1 (ET-1), CD44, and hyaluronic acid (HA) play in VSMC proliferation. Co-localization of ET-1, CD44 and HA positive areas, as well as proliferating cell nuclear antigen positive nuclei, were observed in mouse neointima induced by endothelial injury. As found in intimal VSMC, cultured mouse VSMC secreted ET-1. The endothelin-converting enzyme (ECE) inhibitor, phosphoramidon, and endothelin type-A (ETA) receptor antagonist BQ-123 reduced expression of CD44 in VSMC. ET-1 reversed the reduction of CD44 expression by phosphoramidon. Because CD44 is a receptor for HA, we investigated the effects of phosphoramidon, BQ-123 or ET-1 on the mitogenic activity of HA in VSMC. Among the different molecular weights of this polysaccharide, oligosaccharides of HA (oHA) stimulated VSMC proliferation most effectively. Phosphoramidon and BQ-123 inhibited this oHA-induced DNA synthesis in VSMC. ET-1 reversed the suppression of oHA-induced proliferation by phosphoramidon. In conclusion, endogenously secreted ET-1 enhances oHA-stimulated VSMC growth via the ETA receptor in an autocrine manner. Thus it is suggested that the CD44-inducing activity of ET-1 is responsible for its stimulating effect on oHA-dependent growth of VSMC. These findings support the hypothesis that the interactions among ET-1, CD44 and HA promote the progression of arteriosclerosis. PMID- 11121596 TI - Prediction of mean pulmonary wedge pressure using doppler pulmonary venous flow variables in hypertrophic cardiomyopathy. AB - We examined whether pulmonary venous flow variables, assessed by transthoracic Doppler echocardiography, could predict mean pulmonary wedge pressure in hypertrophic cardiomyopathy. Forty-four patients with no left ventricular systolic dysfunction (left ventricular fractional shortening > or =25%) were studied. Forty patients with systolic dysfunction (dilated cardiomyopathy group) served as control. Mitral and pulmonary venous flow velocity curves were recorded with the pulsed-Doppler method and were related to mean pulmonary wedge pressure obtained by right heart catheterization. In hypertrophic cardiomyopathy group, the systolic (r=-0.15, P=0.335) and diastolic (r=0.35, P=0.022) forward flow velocity were poorly related to mean pulmonary wedge pressure, whereas the velocity of atrial reversal (r=0.68, P<0.001) correlated well with mean pulmonary wedge pressure. In dilated cardiomyopathy group, the systolic (r=-0.51, P=0.001) and diastolic (r=0.60, P<0.001) forward flow velocity were strongly related to mean pulmonary wedge pressure. With the cut-off value set at the velocity of atrial reversal >30 cm/s in hypertrophic cardiomyopathy group, the sensitivity for predicting mean pulmonary wedge pressure >15 mmHg was 79% and the specificity was 73%. In conclusion, the atrial component of the pulmonary venous flow can be used to predict mean pulmonary wedge pressure in hypertrophic cardiomyopathy. PMID- 11121597 TI - Hyperplasia of coronary intima in young males in relation to development of coronary heart disease in adults. AB - The aim of the investigation was to study structural features of coronary arteries in young males which may influence the development of stenosing coronary atherosclerosis in older age. We studied the coronary arteries from 84 males, 10 39 years old, who died from accidents in Moscow, Malmo and Riga, and 98 males aged 40 and above from Moscow who died from coronary heart disease (71 cases) or other diseases (27 cases). In children and young males from all three cities, musculo-elastic hyperplasia of the coronary intima took place constantly but with different degrees of expression; a strict relationship of the intimal thickness and age was observed. Histometric investigations of the right coronary artery showed that in young males of Riga, in comparison with those of Malmo, the intima was significantly thicker both outside (69.6+/-2.8 and 58. 2+/-2.5 microm) and within the area of cushion like thickening (118. 8+/-4.0 and 101.9+/-3.8 microm), they had more extended cushion-like thickening of intima (42.6+/-3.0 and 30.8+/ 3.3% to the length of the artery circumference) and destroyed parts of the internal elastic lamina (28.3+/-1.9 and 19.1+/-1.7% of its length). In males older than 40 years, severe coronary atherosclerosis and stenosis was also significantly more common in Riga than in Malmo. Our data indicate that with age the intimal musculo-elastic hyperplasia in the coronary arteries is transformed to a fibro-elastic layer. The thickness of this layer in the presence of stenosing plaques (>75% of arterial lumen) was much greater than in the presence of plaques with stenoses less than 50% (188.1+/- 7.3 and 69.8+/-4.5 microm, respectively). CONCLUSION: The development of stenosing coronary atherosclerosis is closely related to the degree of musculo-elastic intimal hyperplasia in childhood and young age. The formation of a fibro-elastic layer in the coronary intima decreases the ability of the artery to dilate during the development of atherosclerosis. PMID- 11121598 TI - Non-invasive coronary bypass graft imaging after multivessel revascularisation. AB - Non-invasive imaging techniques for the detection of graft patency after multivessel coronary revascularisation may be useful for follow-up after surgery. Forty consecutive asymptomatic patients (38 men, age 59.9+/-1.3 years) who had undergone coronary bypass surgery with at least three grafts were examined by spiral computed tomography or magnetic resonance angiography 24.9+/-0.3 months after surgery, using conventional angiography as reference. In total, 133 grafts (37 internal mammary artery, 96 venous grafts) were analysed. Spiral computed tomography studies were performed with a subsecond scanner; for magnetic resonance angiography, a three-dimensional contrast-enhanced gradient echo technique with ultrashort echo time during breath holding was used. For spiral computed tomography, sensitivities were 76% (internal mammary artery) and 100% (venous graft). This was compared with 100% (internal mammary artery) and 92% (venous graft) assessed by magnetic resonance angiography (P=ns). The positive predictive values were 100% for internal mammary artery and venous graft (spiral computed tomography) and 100% (internal mammary artery), 92% for venous grafts studied by magnetic resonance angiography (P=ns). Both subsecond spiral computed tomography and contrast-enhanced magnetic resonance angiography are highly accurate and relatively non-invasive approaches of assessing coronary graft patency after multivessel revascularisation and have potential for follow-up assessment in the long term. PMID- 11121599 TI - Cardiovascular disease in chinese type 2 diabetic women is associated with a prolonged QTc interval. AB - We studied the relationships between QT interval and cardiovascular disease status in 192 Chinese type 2 diabetic patients. Of these 192 subjects, 132 (68.8%) were women and 60 (31.2%) were men. The mean age (+/-S.D.) was 56.6+/ 12.9 years (range: 23-84, median: 58.0 years). Women had longer QTc interval compared to men (0.402+/-0.030 s vs. 0.387+/-0.026 s, P<0.01). Of the 192 subjects, 18 women and two men had prolonged QTc interval (QTc >0.433 s). Women with prolonged QTc interval have a 2.8-fold greater rate of cardiovascular disease as compared to those with normal QTc interval (38.9% vs. 14.0%, P<0.05). Using multiple regression analysis (stepwise) to assess the relationship with QTc interval with age, sex, body mass index, waist-hip ratio, blood pressure, fasting plasma glucose, glycated haemoglobin, lipid profiles, smoking and duration of diabetes as independent variables (R(2)=0.146, F=8.88, P<0.001), systolic blood pressure (beta=0.198, P=0.017), age (beta=0. 189, P=0.023) and female gender (beta=0.157, P=0.037) were found to be independently associated with QTc interval. In conclusion, we have shown a significant association between prolonged QTc interval, ischaemic heart disease and cardiovascular disease in Chinese type 2 diabetic women. Age, systolic blood pressure and female gender are independently correlated to QTc interval. PMID- 11121600 TI - Catecholamine cardiomyopathy in bilateral malignant pheochromocytoma: successful reversal after surgery. PMID- 11121601 TI - Haemopericardium following thrombolysis for acute myocardial infarction. PMID- 11121602 TI - Elongated mitral chordae tendinae prolapsing to the left ventricular outflow tract. PMID- 11121603 TI - Are diagnostic microbiology tests reproducible? PMID- 11121604 TI - Simultaneous detection of gfp-marked Moraxella sp. G21r and lux-marked Ralstonia eutrophas H850Lr using most-probable-number method. AB - The green fluorescent protein encoded by gfp gene and the luminescent protein encoded by luxAB genes were used as markers to detect p-nitrophenol (PNP) degrading Moraxella sp. G21r and polychlorinated biphenyl (PCB)-degrading Ralstonia eutrophas H850Lr cells, respectively, in mixed liquid cultures and in soil samples using a most-probable-number (MPN) assay. Population estimates for both gfp-marked G21r and lux-marked H850Lr by using MPN assays were similar to viable colony counts. The MPN assay with microtiter plates permitted the simultaneous detection of fluorescent and luminescent bacteria in soil samples faster than conventional plate counting. PMID- 11121605 TI - DNA extraction from coral reef sediment bacteria for the polymerase chain reaction. AB - A rapid and effective method for the direct extraction of high molecular weight amplifiable DNA from two coral reef sediments was developed. DNA was amplified by the polymerase chain reaction (PCR) using 16S rDNA specific primers. The amplicons were digested with HaeIII, HinP1I and MspI and separated using polyacrylamide gel electrophoresis and silver staining. The resulting amplified ribosomal DNA restriction analysis (ARDRA) patterns were used as a fingerprint to discern differences between the coral reef sediment samples. Results indicated that ARDRA is an effective method for determining differences within the bacterial community amongst different environmental samples. PMID- 11121606 TI - Use of substrate responsive-direct viable counts to visualize naphthalene degrading bacteria in a coal tar-contaminated groundwater microbial community. AB - A microscopy-based method was developed to distinguish naphthalene-degrading bacteria within the microbial community of a coal tar-contaminated groundwater system. Pure cultures of Pseudomonas putida NCIB 9816-4 were used to develop the substrate responsive-direct viable count (SR-DVC) method. Cells were concentrated on membrane filters, placed on agar plates of Stanier's minimal basal salts media containing antibiotics (nalidixic acid, piromidic acid, pipemidic acid, and cephalexin), and exposed to vapors of naphthalene. Following brief incubation, samples were fixed in 2% formaldehyde and examined by epifluorescent microscopy. Pure cultures displayed the expected cell elongation response to the SR-DVC assay and required a minimum incubation time of 9 h for differentiation of elongated cells. When applied to groundwater samples from the study site, naphthalene responsive cells in the groundwater community were easily distinguished from unresponsive cells and debris (350+/-180 substrate responsive cells/ml, relative to negative controls with no added growth substrate). In an attempt to reduce background counts of elongated bacteria and fungi, the SR-DVC procedure was modified by adding a wash step prior to incubation and a fungal inhibitor, cyclohexamide, to the plates. When groundwater samples were subjected to the modified procedure, only cells in washed samples showed a significant response to naphthalene (150+/-25 cells/ml), indicating the presence of inhibitory substances in the groundwater. Variations in response of the groundwater microbial community to the two SR-DVC procedures suggest that subsurface conditions (microbial and chemical composition) vary temporally. SR-DVC allows the phenotypes of individual naturally occurring cells to be assessed. PMID- 11121607 TI - An efficient cell-free protein synthesis system using periplasmic phosphatase removed S30 extract. AB - An efficient cell-free translation system was developed by removal of phosphatase localized in the periplasmic space, which hampers the translation reaction by hydrolyzing ATP. S30 extract was prepared from the spheroplast of Escherichia coli, and as much as 40% of ATP-hydrolysis activity of phosphatases could be easily removed by the spheroplast formation. The reaction period of translation using phosphatase-removed S30 extract could be prolonged and protein synthesis was enhanced by more than 30%. PMID- 11121608 TI - Estimating viability of Cryptosporidium parvum oocysts using reverse transcriptase-polymerase chain reaction (RT-PCR) directed at mRNA encoding amyloglucosidase. AB - The purpose of the present study was to determine if reverse transcriptase polymerase chain reaction (RT-PCR) directed at mRNA encoding the enzyme amyloglucosidase (CPAG) could serve as a indicator for C. parvum oocyst viability. Oocysts were stored for 1-11 months in the refrigerator and at monthly intervals extracted for total RNA for RT-PCR analysis. An aliquot of these C. parvum oocysts was inoculated into neonatal mice which were necropsied 4 days later for ileal tissue that was analyzed by semi-quantitative PCR to determine the level of parasite replication. The CPAG RT-PCR assay detected RNA from as few as 10(3) C. parvum oocysts. An effect of storage time on both RT-PCR signal and mouse infectivity was observed. RNA from oocysts stored for 1-7 months, unlike oocysts stored for 9 or 11 months, contained CPAG mRNA that was detectable by RT PCR. A gradual decrease in the RT-PCR signal intensity was observed between 5 and 7 months storage. The intensity of RT-PCR product from oocysts and the signal from semi-quantitative PCR of ileal tissue DNA from mice infected with these same aged oocysts were comparable. The RT-PCR assay of CPAG mRNA in cultured cells infected with viable C. parvum oocysts first detected expression at 12 h with highest expression levels observed at 48 h post-infection. These results indicate that CPAG RT-PCR may be useful for differentiating viable from non-viable C. parvum oocysts and for studying the expression of the gene for amyloglucosidase in vitro. PMID- 11121609 TI - Evaluation of extraction methods for recovery of fatty acids from lipid-producing microheterotrophs. AB - The effect of different extraction techniques on the recovery of fatty acids from freeze-dried biomass of two lipid-producing microheterotrophs was examined. Two procedures were used: the extraction of lipids from biomass followed by transesterification of the fatty acids (extraction-transesterification); and the direct transesterification of biomass to produce fatty acid methyl esters (i.e. without the initial extraction step). Variable factors in the extraction transesterification experiment were the sequence in which solvents were added to the samples, the relative amount of methanol in the solvent mix, and sonication of biomass while in the solvent mix. Variable factors in the direct transesterification experiment were sample size, and reaction duration. Statistical analysis of data (level of significance P<0.05) showed that: (1) extraction of total fatty acids prior to transesterification was significantly more efficient when solvents were added in the order of increasing polarity; (2) neither sonication nor increasing the proportion of methanol in the extraction solvent significantly affected extraction of fatty acids prior to transesterification; (3) efficiency of direct transesterification of fatty acids increased significantly with reaction time; (4) efficiency of direct transesterification of fatty acids was not significantly affected by sample size; (5) the most efficient method for extraction of fatty acids prior to transesterification yielded significantly less fatty acids than the most effective direct transesterification method. While the study examined only two strains, our results suggest that fatty acid analysis methodology for microheterotrophs under consideration for biotechnological exploitation requires optimisation and validation. PMID- 11121610 TI - A preliminary evaluation of a new enzyme immunoassay to detect Chlamydia pneumoniae-specific antibodies. AB - New enzyme immunoassays (EIAs) for determination of specific IgG, IgA, and IgM antibody titers to Chlamydia pneumoniae were evaluated independently in three research laboratories. Specificity of the EIAs was enhanced by removing LPS from the chlamydial antigen. The performance of these EIAs was evaluated in comparison with the microimmunofluorescence (MIF) test using specimens from: (i) a group of adult patients with community-acquired pneumonia (CAP) previously diagnosed as having an acute chlamydial infection by the complement fixation test or the whole inclusion fluorescence test; (ii) from a group of adult patients with acute respiratory tract infections; and (iii) from a group of young children consecutively presenting with acute respiratory tract infections. The MIF test and the EIAs detected acute infections in paired serum specimens from 12 of 14 patients from the first group. Eleven of these 12 patients were positive in both tests. The MIF test detected seven acute infections in single convalescence serum specimens from eight patients. Two of these were also positive in the EIAs. Paired serum specimens from the second group of adult patients (n=12) were collected during an epidemic of C. pneumoniae. The EIAs detected six acute infections. The MIF test detected two additional patients with acute infections. From the group of young children (n=30), the EIAs detected two patients with acute infections. Our conclusion from this preliminary evaluation is that these EIAs could be useful for laboratory diagnosis of acute C. pneumoniae infections. Comprehensive prospective studies should provide suitable data to calculate the sensitivity, specificity, and predictive values. PMID- 11121611 TI - A new selective medium for the isolation of glucose non-fermenting bifidobacteria from hen caeca. AB - Avian caeca contain a large and diverse population of bacteria. Certain genera, including Bifidobacterium, are thought to exert health-promoting effects. Two media were evaluated to determine their sensitivity and selectivity for bifidobacteria in the hen caecal samples: modified Wilkins-Chalgren agar (MW; Oxoid) with the addition of glacial acetic acid (1 ml/l) and mupirocin (100 mg/l) and modified TPY agar (MTPY; ADSA, Spain) with glacial acetic acid (1 ml/l) and mupirocin (100 mg/l). The colonies arising on the plates inoculated with caecal samples were Gram stained, screened for the presence of fructose-6-phosphate phosphoketolase activity, and tested for fermentation patterns using ANAEROtest (Lachema, Czech Republic) and API 50 CHL (BioMerieux, France) kits. Both agars were selective for bifidobacteria, however, MTPY agar showed higher cfu/g than MW agar. Bifidobacterial counts were higher than 10(10) cfu/g of caecal contents using MTPY agar. Most of strains isolated from this medium fermented melibiose, sucrose, and raffinose, but not glucose. Soya peptone (5 g/l; Oxoid) stimulated the growth of glucose non-fermenting strains in complex liquid media. The results suggest that the media for selective enumeration and isolation of bifidobacteria in poultry caecal samples should not contain glucose as the sole carbon source. It can be concluded that MTPY medium is highly selective and permits the growth of both glucose fermenting and glucose non-fermenting bifidobacteria. PMID- 11121612 TI - Analysis of the dynamics of fungal communities in soil via fungal-specific PCR of soil DNA followed by denaturing gradient gel electrophoresis. AB - A molecular method for profiling of fungal communities in soil was applied in experiments in soil microcosms, with two objectives, (1) to assess the persistence of two selected fungal species in soil, and (2) to analyze the response of the natural fungal community to a spill of sulphurous petrol in the same soil. To achieve the aims, two soil DNA extraction methods, one originally designed for the direct extraction of bacterial community DNA and the other one aimed to obtain fungal DNA, were tested for their efficiency in recovering DNA of fungal origin from soil. Both methods allowed for the efficient extraction of DNA from introduced Trichoderma harzianum spores as well as Arthrobotrys oligospora mycelial fragments, at comparable rates. Several PCR amplification systems based on primers specific for fungal 18S ribosomal RNA genes were tested to design strategies for the assessment of fungal communities in soil. The PCR systems produced amplicons of expected size with DNA of most fungi studied, which included members of the Ascomycetes, Basidiomycetes, Zygomycetes and Chytridiomycetes. On the other hand, the 18S rRNA genes of Oomycetes (including key plant pathogens) were poorly amplified. Plant (Solanum tuberosum), nematode (Meloidogyne sp.) and bacterial DNA was not amplified. For studies of soil fungal communities, a nested PCR approach was selected, in which the first PCR provided the required specificity for fungi, whereas the second (nested) PCR served to produce amplicons separable on denaturing gradient gels. Denaturing gradient gel electrophoresis (DGGE) allowed the resolution of mixtures of PCR products of several different fungi, as well as products resulting from mixed-template amplifications, into distinct banding patterns. The persistence of fungal species in soil was assessed using T. harzianum spores and A. oligospora hyphal fragments added to silt loam soil microcosms. Using PCR-DGGE, these fungi were detectable for about 14 days and 2 months, respectively. Both singly-inoculated soils and soils that had received mixed inoculants revealed, next to bands resulting from indigenous fungi, the expected bands in the DGGE profiles. The A. oligospora specific amplicon, by virtue of its unique migration in the denaturing gradient, was well detectable, whereas the T. harzianum specific product comigrated with products from indigenous fungi. PCR-DGGE analysis of DNA obtained from the silt loam soil treated with dibenzothiophene-containing petrol showed the progressive selection of specific fungal bands over time, whereas this selection was not observed in untreated soil microcosms. Cloning of individual molecules from the selected bands and analysis of their sequences revealed a complex of targets which clustered with the 18S rDNA sequences of the closely-related species Nectria haematococca, N. ochroleuca and Fusarium solani. Fungal isolates obtained from the treated soil on PDA plates were identified as Trichoderma sp., whereas those on Comada agar fell into the Cylindrocarpon group (anamorph of Nectria spp). PMID- 11121613 TI - Synthesis and anticonvulsant activity of 4-bromophenyl substituted aryl semicarbazones. AB - A number of 4-bromophenyl semicarbazones were synthesised and evaluated for anticonvulsant and sedative -hypnotic activities. After intraperitoneal injection to mice, the semicarbazone derivatives were examined in the maximal electroshock seizure (MES), subcutaneous pentylenetetrazole (scPTZ), subcutaneous strychnine (scSTY) and neurotoxicity (NT) screens. All the compounds showed anticonvulsant activity in one or more test models. Compound 12 showed greatest activity, being active in all the screens with very low neurotoxicity and no sedative-hypnotic activity. All the compounds except 7 had lower neurotoxicity compared to phenytoin. Three compounds (6, 11 and 14) showed greater protection than sodium valproate. The essential structural features responsible for interaction with receptor site are established within a suggested pharmacophore. PMID- 11121614 TI - Synthesis of di- and tripeptide analogues containing alpha-ketoamide as a new core structure for inhibition of HIV-1 protease. AB - Di- and tripeptide analogues containing alpha-ketoamide as a new core structure and incorporating allophenylnorstatine (Apns) as a transition state mimic, were designed and synthesized in the hope of obtaining a novel structural type of HIV 1 protease inhibitors. The immediate precursor, Apns-Thz-NHBu(t) was prepared by coupling of Boc-Apns with N-tert x butyl Thz-4-carboxamide hydrochloride. Removal of Boc group followed by coupling with the respective alpha-ketoacid residue (P2) gave the desired dipeptides (8-12) in almost quantitative yields. The alpha-keto tripeptides (18-21) were obtained by oxidation of the hydroxyl group of Apns (PI) in the appropriate tripeptide, iQOA-Val-Apns-(un)substituted Thz(Oxa)-NHBu(t) with DMSO/DCC. Preliminary evaluation of the activity of the synthesized derivatives was determined as percentage of enzyme inhibition at 5 microM and 50 nM levels of the di- and tripeptides respectively. The alpha-ketoamides displayed a significant enhanced potency relative to their parent isosteres as inhibitors of HIV-1 protease and are shown to be a promising new core structure for the development of enzyme inhibitors. A quantitative approach was attempted, using an LFE model, correlating the effect of structural modification and HIV-1 protease inhibition activity of the prepared dipeptides. The result indicates the contribution of the torsion angle by 84% to the activity of the inhibitors. PMID- 11121615 TI - Synthesis and in vitro opioid activity profiles of DALDA analogues. AB - The tetrapeptide DALDA (H-Tyr-D-Arg-Phe-Lys-NH2) is a polar and selective mu agonist showing poor penetration of the placental and blood-brain barriers. In an effort to enhance the potency of DALDA, analogues containing 2',6' dimethyltyrosine (Dmt), N,2',6'-trimethyltyrosine (Tmt), 2'-methyltyrosine (Mmt) or 2'-hydroxy,6'-methyltyrosine (Hmt) in place of Tyr1, or Orn or alpha,gamma diaminobutyric acid (A2bu) in place of Lys4, were synthesized. All compounds displayed high mu receptor selectivity in the rat and guinea pig brain membrane binding assays and most of them were more potent mu agonists than DALDA in the mu receptor-representative guinea pig ileum assay, with [Dmt1]DALDA showing the highest potency. Because of its extraordinary mu agonist potency, high mu selectivity, polar character (charge of 3 + ) and metabolic stability, [Dmt1]DALDA has potential for use in obstetrical or peripheral analgesia. PMID- 11121616 TI - Formation of micelles and liposomes from carnitine amphiphiles. AB - Esterification of the carboxy and/or the hydroxy groups of (R)-carnitine (3 hydroxy-4-trimethylammonium butanoate) produces interesting classes of (cationic or zwitterionic) surfactants whose CMC values are in general predictable from their molecular structure. In fact similar relationships between CMC and the number of carbon atoms, Cn, have been found for three classes of such surfactants. However the sensitivity of CMC to Cn for the diesters is considerably lower than that calculated from literature values for the monoesters (either in their cationic or zwitterionic forms). The CMC values for the diesters have been determined by tensiometric, conductimetric and spectrophotometric methods, both in pure water and in 0.154 M NaCl solutions, at 25 degrees C. In particular the tensiometric results provide evidence that double-chain diesters undergo self assembly into structures more complex than simple micelles if the two chains are of comparable length. EPR and electron microscopy experiments show that the aggregates spontaneously formed by these surfactants are a mixture of multilamellar vescicles. PMID- 11121617 TI - N-Benzylpiperidine derivatives of 1,2,4-thiadiazolidinone as new acetylcholinesterase inhibitors. AB - A new family of 1,2,4-thiadiazolidinone derivatives containing the N benzylpiperidine fragment has been synthesised. The acetylcholinesterase (AChE) inhibitory activity of all compounds was measured using Ellman's method and some of them turned out to be as potent as tacrine. Furthermore, compound 13 was as active as tacrine in reversing the blockade induced by tubocurarine at rat neuromuscular junction. Additionally, receptor binding studies provided new lead compounds for further development of alpha2-adrenergic and sigma-receptor antagonists. Molecular dynamic simulation using X-ray crystal structure of AChE from Torpedo californica was used to explain the possible binding mode of these new compounds. PMID- 11121618 TI - Isosterism among analogues of torasemide: conformational, electronic and lipophilic properties. AB - The structures, electronic (charges, molecular electrostatic potential, molecular orbitals) and lipophilic properties of three isostere analogues of torasemide were determined and the influence of the replacement of the sulfonyl urea group on the conformation and electronic properties of the molecules is discussed. Lipophilicity of the compounds seems to be the most discriminating property along the series and affects their pharmacological activities. PMID- 11121619 TI - 6-Substituted 1H-quinolin-2-ones and 2-methoxy-quinolines: synthesis and evaluation as inhibitors of steroid 5alpha reductases types 1 and 2. AB - A Negishi-type coupling reaction between 6-bromo-2-methoxyquinoline (1a) and various 4-bromo-N,N-dialkyl-benzamides gave access to 6-substituted 2-methoxy quinolines 1-3 and 1H-quinolin-2-ones 4-12. Most of these compound proved to be inhibitors of steroid 5alpha reductases with activity and selectivity both being strongly dependent on the features of the heterocycle and the size of the N,N dialkylamide substituent. The most active inhibitor for the human type 2 isozyme was 6-[4-(N,N-diisopropylcarbamoyl)phenyl]-1H-quinolin-2-one 4 (Ki 800 +/- 85 nM), showing mostly competitive inhibitory patterns. A type 1 selective inhibitor could be identified with 6-[4-(N,N-diisopropylcarbamoyl)phenyl]-N-methyl-quinolin 2-one (5, IC50 510 nM). PMID- 11121620 TI - Synthesis, brain antihypoxic activity and cell neuroprotection of 1-substituted 3,7-dimethylxanthines. AB - Five newly synthesised original compounds were investigated for acute toxicity, influence on hexobarbital sleeping time, effect on the locomotor activity, and brain antihypoxic activity. Two of the compounds were tested in a model of glutamate induced neurotoxicity in the brain cell culture using a cell viability test. Our studies indicate that compounds 2a-c and 4 prolonged the survival time of mice in the model of anoxic hypoxia. Only compound 3 expressed antihypoxic activity in the model of circulatory hypoxia, evaluated with a statistical significant increase of the survival time. Compound 4 (1-[3-(2,3-dihydro-3 oxobenzisosulfonazol2-yl)-propyl]-3,7-dimethylxanthine) in concentration range 0.3-3 microM statistically significantly antagonised the glutamate induced neurotoxicity. Compound 4 is important for further investigations on in vivo models of brain dementia. PMID- 11121621 TI - Triazolyl-benzimidazolones and triazolyl-benzotriazoles: new potential potassium channel activators. II. AB - This paper reports the synthesis and pharmacological evaluation of a series of 5 substituted-triazolyl benzotriazoles (2a-f) and the corresponding series of 5 substituted-triazolyl-benzimidazolones (6a-f), as potential activators of the big conductance calcium-activated potassium channels (BK(Ca)). The synthesis and structure demonstration of the stock compounds of the two series have been described in our previous works, as well as the common starting compounds 4 carboxamido-5-(4-substituted-2amino-anilino)-1,2,3-triazoles (1a-f). The triazolyl-benzotriazoles were obtained by diazotization, while the triazolyl benzimidazolones were obtained by thermal intramolecular cyclization of ethoxycarbonylamino derivatives or directly with phosgene. Benzimidazolone compounds generally showed little effect whilst the compounds with a benzotriazole ring showed full efficacy, with vasorelaxing properties and potency parameters a little lower than that of the reference compound NS 1619. These effects were significantly reduced by an increased membrane depolarization. This depolarization-sensitive response is in agreement with the pharmacodynamic hypothesis of activation of potassium channels. PMID- 11121622 TI - Anhydride modified cantharidin analogues. Is ring opening important in the inhibition of protein phosphatase 2A? AB - A series of anhydride modified cantharidin analogues have been synthesised and screened for their ability to inhibit protein phosphatase 2A. Surprisingly only analogues capable of undergoing a facile ring opening of the anhydride moiety displayed any significant inhibition. Subsequent NMR experiments indicated that 7 oxobicyclo[2.2.1]heptane-2,3-dicarboxylic acid was the major (sole) species under assay conditions. The ability of these modified anhydro-cantharidin analogues to inhibit protein phosphatase 2A varies from 4 (16) to 100% (8) at 100 microM test concentration. PMID- 11121623 TI - Concurrent chemotherapy and radiation therapy for advanced stage carcinoma of the nasopharynx. PMID- 11121624 TI - True recurrence vs. new primary ipsilateral breast tumor relapse: an analysis of clinical and pathologic differences and their implications in natural history, prognoses, and therapeutic management. AB - PURPOSE: The purpose of this study was to classify all ipsilateral breast tumor relapses (IBTR) in patients treated with conservative surgery and radiation therapy (CS+RT) as either new primary tumors (NP) or true local recurrences (TR) and to assess the prognostic and therapeutic implications of this classification. METHODS AND MATERIALS: Of the 1152 patients who have been treated at Yale-New Haven Hospital before 1990, 136 patients have experienced IBTR as their primary site of failure. These relapses were classified as either NP or TR. Specifically, patients were classified as NP if the recurrence was distinctly different from the primary tumor with respect to the histologic subtype, the recurrence location was in a different location, or if the flow cytometry changed from aneuploid to diploid. This information was determined by a detailed review of each patient's hospital and/or radiotherapy record, mammograms, and pathologic reports. RESULTS: As of 2/99, with a mean follow-up of 14. 2 years, the overall ipsilateral breast relapse-free rate for all 1152 patients was 86% at 10 years. Using the classification scheme outlined above, 60 patient relapses were classified as TR, 70 were classified as NP and 6 were unable to be classified. NP patients had a longer mean time to breast relapse than TR patients (7.3 years vs. 3.7 years, p < 0.0001) and were significantly younger than TR patients (48.9 years vs. 54.5 years, p < 0.01). Patients developed both TR and NP at similar rates until approximately 8 years, when TR rates stabilized but NP rates continued to rise. By 15 years following original diagnosis, the TR rate was 6.8% compared to 13.1% for NP. Of the patients who had been previously tested for BRCA1/2 mutations, 17% (8/52) had deleterious mutations. It is noteworthy that all patients with deleterious mutations had new primary IBTR, while patients without deleterious mutations had both TR and NP (p = 0.06). Ploidy was evenly distributed between TR and NP but NP had a significantly lower S phase fraction (NP 13.1 vs. TR 22.0, p < 0.05). The overall survival following breast relapse was 64% at 10 years and 49% at 15 years. With a mean follow-up of 10.4 years following breast relapse, patients with NP had better 10-year overall survival (TR 55% vs. NP 75%, p < 0.0001), distant disease-free survival (TR 41% vs. NP 85%, p < 0.0001), and cause specific survival (TR 55% vs. NP 90%, p < 0.0001). CONCLUSION: It appears that a significant portion of patients who experience ipsilateral breast tumor relapse following conservative surgery and radiation therapy have new primary tumors as opposed to true local recurrences. True recurrence and new primary tumor ipsilateral breast tumor relapses have different natural histories, different prognoses, and, in turn, different implications for therapeutic management. PMID- 11121625 TI - The 1993-94 patterns of care process survey for breast irradiation after breast conserving surgery-comparison with the 1992 standard for breast conservation treatment. The Patterns of Care Study, American College of Radiology. AB - PURPOSE: To determine the patterns of evaluation and treatment in the U.S. of women with early breast cancer treated with breast-conserving surgery and irradiation in 1993-94, and to compare these with a similar survey in 1983 and with the 1992 Standard for Breast Conservation Treatment. METHODS AND MATERIALS: In 1995-96, 727 randomly selected records of eligible patients treated from 1993 94 at 62 facilities representative of 3 practice types were reviewed. RESULTS: Compared with the Process Survey (PS) in 1983, patients in the 1993-94 study had an older age distribution. In the current study, 70% of patients were > or = 50 years of age, and 69% were post-menopausal, compared with 59% > or = 50 years of age and 49% post-menopausal in 1983 (p = 0.0087 and < 0.001, respectively). Work up and evaluation in the 1993-94 PS were closely aligned with the standard and were considerably improved compared with 1983. In the 1983 study, 77% of patients underwent mammography, as compared to 97% in the 1993-94 study. In 1983, pathological size documentation was performed in 83% of patients; in 1993-94, this was performed in 95% of patients. An estrogen receptor evaluation was performed in 36% of patients in 1983; in 1993-94, that increased to 76%. In 1983, 28% of patients underwent progesterone receptor evaluation; in 1993-94, this increased to 72%. Only 3% of patients in 1993-94 were enrolled in a clinical trial. Radiation treatment parameters closely adhered to standard recommendations, improving substantially from 1983. In 1983, wedge or compensator use was recommended for 64% of patients; in 1993-94, for 95% of patients. In 1983, 4-8 MV photons were recommended for breast treatment in 67% of patients; in 1993-94, 90%. In 1983, bolus was avoided in 75% of patients; in 1993-94, in 94%. In 1983, the recommended breast dose for 89% of patients was 45-50 Gy (44-51 Gy in PS); in 1993-94 this had increased to 99% of patients. In 1983, electrons were recommended for primary site boost in 70% of patients; in 1993-94, for 94% of patients. CONCLUSION: There was an extensive shift to adherence to the 1992 standard in 1993-94, compared with the 1983 PS, although there is room for improvement in some areas. PMID- 11121626 TI - Interstitial high-dose-rate brachytherapy boost: the feasibility and cosmetic outcome of a fractionated outpatient delivery scheme. AB - PURPOSE: To evaluate the feasibility, potential toxicity, and cosmetic outcome of fractionated interstitial high dose rate (HDR) brachytherapy boost for the management of patients with breast cancer at increased risk for local recurrence. METHODS AND MATERIALS: From 1994 to 1996, 18 women with early stage breast cancer underwent conventionally fractionated whole breast radiotherapy (50-50.4 Gy) followed by interstitial HDR brachytherapy boost. All were considered to be at high risk for local failure. Seventeen had pathologically confirmed final surgical margins of less than 2 mm or focally positive. Brachytherapy catheter placement and treatment delivery were conducted on an outpatient basis. Preplanning was used to determine optimal catheter positions to enhance dose homogeneity of dose delivery. The total HDR boost dose was 15 Gy delivered in 6 fractions of 2.5 Gy over 3 days. Local control, survival, late toxicities (LENT SOMA), and cosmetic outcome were recorded in follow-up. In addition, factors potentially influencing cosmesis were analyzed by logistic regression analysis. RESULTS: The minimum follow-up is 40 months with a median 50 months. Sixteen patients were alive without disease at last follow-up. There have been no in breast failures observed. One patient died with brain metastases, and another died of unrelated causes without evidence of disease. Grade 1-2 late toxicities included 39% with hyperpigmentation, 56% with detectable fibrosis, 28% with occasional discomfort, and 11% with visible telangiectasias. Grade 3 toxicity was reported in one patient as persistent discomfort. Sixty-seven percent of patients were considered to have experienced good/excellent cosmetic outcomes. Factors with a direct relationship to adverse cosmetic outcome were extent of surgical defect (p = 0.00001), primary excision volume (p = 0.017), and total excision volume (p = 0.015). CONCLUSIONS: For high risk patients who may benefit from increased doses, interstitial HDR brachytherapy provides a convenient outpatient method for boosting the lumpectomy cavity following conventional whole breast irradiation without overdosing normal tissues. The fractionation scheme of 15 Gy in 6 fractions over 3 days is well tolerated. The volume of tissue removed from the breast at lumpectomy appears to dominate cosmetic outcome in this group of patients. PMID- 11121627 TI - Randomized phase III study comparing Best Supportive Care to Biafine as a prophylactic agent for radiation-induced skin toxicity for women undergoing breast irradiation: Radiation Therapy Oncology Group (RTOG) 97-13. AB - PURPOSE: To determine if Biafine compared to Best Supportive Care (BSC) is effective in minimizing or preventing radiation-induced dermatitis in women undergoing breast irradiation. METHODS AND MATERIALS: Patients were randomized between Biafine (n = 83) vs. BSC (n = 89). The institutions identified preference for BSC at the time of randomization. A no-treatment arm was allowed (16% received no treatment). Patients were instructed to apply randomized product three times a day, but not within 4 h of their daily RT session. Application began following their first radiation treatment and continued 2 weeks postradiation. Skin dermatitis was scored weekly utilizing the RTOG and ONS (Oncology Nursing Society) skin toxicity scales, a weekly patient satisfaction and quality-of-life questionnaire. RESULTS: Using the RTOG toxicity scale there was no overall difference for maximum dermatitis during RT between Biafine and BSC (p = 0.77). There was no difference in maximum toxicity by arm or breast size. There was an interaction between breast size and toxicity, with large breasted women exhibiting more toxicity. Large-breasted women receiving Biafine were more likely to have no toxicity 6 weeks post RT. CONCLUSION: There was no overall difference between BSC and Biafine in the prevention, time to, or duration of radiation-induced dermatitis. PMID- 11121628 TI - Final report of a randomized trial on altered-fractionated radiotherapy in nasopharyngeal carcinoma prematurely terminated by significant increase in neurologic complications. AB - PURPOSE: The aim of the present study was to compare the survival, local control and complications of conventional/accelerated-hyperfractionated radiotherapy and conventional radiotherapy in nonmetastatic nasopharyngeal carcinoma (NPC). METHODS AND MATERIALS: From February 1993 to October 1995, 159 patients with newly diagnosed nonmetastatic (M0) NPC with N0 or 4 cm or less N1 disease (Ho's N stage classification, 1978) were randomized to receive either conventional radiotherapy (Arm I, n = 82) or conventional/accelerated-hyperfractionated radiotherapy (Arm II, n = 77). Stratification was according to the T stage. The biologic effective dose (10 Grays) to the primary and the upper cervical lymphatics were 75.0 and 73.1 for Arm I and 84.4 and 77.2 for Arm II, respectively. RESULTS: With comparable distribution among the T stages between the two arms, the free from local failure rate at 5 years after radiotherapy was not significantly different between the two arms (85.3%; 95% confidence interval, 77.2-93.4% for Arm I; and 88.9%; 95% confidence interval, 81.7-96.2% for Arm II). The two arms were also comparable in overall survival, relapse-free survival, and rates of distant metastasis and regional relapse. Conventional/accelerated hyperfractionated radiotherapy was associated with significantly increased radiation-induced damage to the central nervous system (including temporal lobe, cranial nerves, optic nerve/chiasma, and brainstem/spinal cord) in Arm II. Although insignificant, radiation-induced cranial nerve(s) palsy (typically involving VIII-XII), trismus, neck soft tissue fibrosis, and hypopituiturism and hypothyroidism occurred more often in Arm II. In addition, the complications occurred at significantly shorter intervals after radiotherapy in Arm II. CONCLUSION: Accelerated hyperfractionation when used in conjunction with a two dimensional radiotherapy planning technique, in this case the Ho's technique, resulted in increased radiation damage to the central nervous system without significant improvement in efficacy. PMID- 11121629 TI - Long-term survival of nasopharyngeal carcinoma following concomitant radiotherapy and chemotherapy. AB - PURPOSE: The purpose of this study is to demonstrate long-term survival of nasopharyngeal carcinoma treated with concomitant chemotherapy and radiotherapy (CCRT) followed by adjuvant chemotherapy. METHODS AND PATIENTS: One hundred and seven patients with Stage III and IV (American Joint Committee on Cancer, AJCC, 1988) nasopharyngeal carcinoma (NPC) were treated with concomitant chemotherapy and radiotherapy (CCRT) followed by adjuvant chemotherapy between April 1990 and December 1997 in Koo Foundation Sun Yat-Sen Cancer Center, Taipei. The dose of radiation was 70 Gray (Gy) given in 35 fractions, 5 fractions per week. Two courses of chemotherapy, consisting of cisplatin and 5-fluorouracil, were delivered simultaneously with radiotherapy in Weeks 1 and 6 and two additional monthly courses were given after radiotherapy. According to the AJCC 1997 staging system, 32 patients had Stage II disease, 44 had Stage III, and 31 had Stage IV disease. RESULTS: With median follow-up of 44 months, the 5-year overall survival rate in all 107 patients was 84.1%, disease-free survival rate was 74.4%, and locoregional control rate was 89.8%. The 3-year overall survival for Stage II was 100%, for Stage III it was 92.8%, and for Stage IV, 69. 4% (p = 0.0002). The 3 year disease-free survival for Stage II was 96.9%, for Stage III it was 87.7%, and for Stage IV it was 51.9% (p = 0.0001). CONCLUSION: CCRT and adjuvant chemotherapy is effective in Taiwanese patients with advanced NPC. The prognosis of AJCC 1997 Stage II and III disease is excellent, but, for Stage IV (M0), it is relatively poor. Future strategies of therapy should focus on high-risk AJCC 1997 Stage IV (M0) cohort. PMID- 11121630 TI - Salvage radiation therapy for locally recurrent nasopharyngeal carcinoma. AB - PURPOSE: To study the treatment outcome in patients with locally recurrent nasopharyngeal carcinoma (NPC) and to explore whether a combination of high-dose rate (HDR) intracavitary brachytherapy and external beam radiation therapy (ERT) could improve the therapeutic ratio. METHODS AND MATERIALS: Ninety-one patients with nonmetastatic locally recurrent NPC who were treated with curative intent during the years 1990-1999 were retrospectively analyzed. Eighty-two patients had histologically proven carcinoma. The remaining 9 had clinical and imaging features suggestive of local recurrence. The Ho's T-stage distribution at recurrence (rT) was as follows: rT1-37, rT2-14, rT3-40. Total equivalent dose (TED) was calculated by the linear-quadratic formula without a time factor correction. For those treated by combined-modality treatment (CMT), the TED was taken as the summation of the equivalent dose by ERT and the absolute dose delivered to floor of the sphenoid by brachytherapy. Eight patients were treated solely with brachytherapy, all receiving 24-45 Gy in 3-10 sessions. Forty-one patients were treated with ERT alone receiving a median TED of 57.3 Gy (range, 49.8-62.5 Gy). Forty-two patients were treated by CMT with a median equivalent dose of 50 Gy (range, 40-60 Gy) given by ERT and 14.8 Gy by brachytherapy (range, 3-29.6 Gy). Multivariate analyses were performed using the Cox regression proportional hazards model. RESULTS: The 5-year actuarial overall survival rate, disease specific survival rate and local failure-free survival (LFFS) rate for the whole group were 30%, 33. 3% and 37.8%, respectively. The 3-year LFFS rates of rT1, rT2, and rT3 diseases were 64%, 61.5%, and 18.4%, respectively (p = 0.001). Of the 8 patients treated with brachytherapy alone, 4 failed locally. Further analyses were concentrated on the ERT (41 patients) and CMT (42 patients) groups. The 3-year LFFS rates of rT1, rT2, and rT3 diseases were 66.7%, 66.7%, and 18.4%, respectively (p = 0.0008). Better local control for patients who received a TED of 60 Gy or greater was shown. The corresponding 3-year LFFS rates were 29% and 60% (p = 0.0004). Subgroup analysis on the ERT and CMT groups showed a 3-year LFFS rate of 33.5% and 57% (p = 0.003). ERT group had an excess of patients with rT3 disease. Further analysis was performed on the rT1-2 patients showing a trend toward improvement in local control in favor of the CMT group (3 year LFFS rates: CMT, 71.7%; ERT, 54%; p = 0.13). Multivariate analyses showed that rT stage (p = 0.002) and TED (p = 0.01; HR, 0.93; 95% confidence interval, 0.88-0. 98) remained significant. The 5-year major and central nervous system (CNS) complication-free rates were 26.7% and 47.8%. The following factors were found to be significant on univariate analyses for both complications in the ERT and CMT groups: (1) Modality of treatment: more complications with ERT group; and (2) rT stage. Multivariate analyses showed that the rT stage was significant for predicting the occurrence of major (p = 0.004) and CNS complications (p = 0.04). CONCLUSION: For rT1-2 local recurrences, CMT with at least 60 Gy TED is recommended. The high incidence of major late complications is of serious concern. Ways of improving the local control of Ho's rT3 disease and reducing the risk of late complications should be explored. PMID- 11121631 TI - Protective effect of amifostine on dental health after radiotherapy of the head and neck. AB - PURPOSE: The cytoprotective agent amifostine has been shown to reduce the radiation-induced acute and chronic xerostomia in head and neck cancer patients. The purpose of this study was to evaluate whether or not amifostine also reduces the incidence of dental caries associated with the radiation-induced xerostomia. METHODS AND MATERIALS: The dental status before and 1 year after radiotherapy was retrospectively compared in 35 unselected patients treated as part of the prospective randomized and multicenter open-label Phase III study (WR-38) at the University Hospitals of Heidelberg, Freiburg, and Erlangen. The WR-38 study compared radiotherapy in head and neck cancer with and without concomitant administration of amifostine. RESULTS: Patient and treatment characteristics (particularly the radiation dose and percentage of parotids included in the treatment volume) were equally distributed between the patients who received (n = 17) or did not receive (n = 18) amifostine. Fifteen patients of the amifostine group showed no deterioration of the dental status 1 year after radiotherapy as compared to 7 patients who did not receive the cytoprotector (p = 0. 015, two tailed Fisher exact test). CONCLUSION: Our data suggest a protective effect of amifostine on the dental health after radiotherapy of the head and neck. The dental status should be used as a primary endpoint in future studies on amifostine. PMID- 11121632 TI - Anemia is associated with decreased local control of surgically treated squamous cell carcinomas of the glottic larynx. AB - PURPOSE: A strong association between hemoglobin levels and tumor control exists in head and neck cancer treated with radiotherapy. This retrospective study has been performed to determine whether or not this association can also be found in the surgical setting. METHODS AND MATERIALS: Between January 1970 and December 1990, 258 patients with glottic SCC received conventional surgery only (T1/T2/T3/T4 n = 188/31/37/2, respectively). Locoregional control was calculated by the Kaplan-Meier method. The influence of hemoglobin, T stage, age, gender, performance/nutritional status, and grading was evaluated using a Cox model. RESULTS: Five-year locoregional control for T1a/T1b/T2/T3/T4 tumors was 91%/85%/76%/62%/0%, respectively (log-rank test, p < 0.0001). Anemia (male < 13, female < 12 g/dL hemoglobin) was present in 27 patients. It was associated with significantly worse 5-year locoregional control, i.e., 60% vs. 85% (log-rank test, p = 0.003). In multivariate analysis stratified for T stage, two variables were of influence: positive margins (relative risk [RR], 3.8; 95% confidence interval [CI], 1.7-8.4), anemia (RR, 3.0; 95% CI, 1.4-6.2). The largest subgroup consisted of 162 patients characterized by male gender, T1, and complete resection. In this subgroup, the significant variables were T stage (T1b vs. T1a; RR, 3.5; 95% CI, 0.96-12.4) and hemoglobin with a RR of 1.4 (95% CI, 1.0-2.1) per g/dL less analyzed as a continuous variable. CONCLUSION: Anemia is associated with a high risk of treatment failure in surgically treated glottic cancer. Hemoglobin levels might be predictive even within the normal range as indicated by subgroup analysis. PMID- 11121633 TI - Phase II, two-arm RTOG trial (94-11) of bischloroethyl-nitrosourea plus accelerated hyperfractionated radiotherapy (64.0 or 70.4 Gy) based on tumor volume (> 20 or < or = 20 cm(2), respectively) in the treatment of newly diagnosed radiosurgery-ineligible glioblastoma multiforme patients. AB - PURPOSE: To compare survivorship, and acute and delayed toxicities following radiation therapy (RT) of radiosurgery-ineligible glioblastoma multiforme (GBM) patients treated with tumor volume-influenced, high-dose accelerated, hyperfractionated RT plus bischloroethyl-nitrosourea (BCNU), using prior RTOG malignant glioblastoma patients as historical controls. METHODS AND MATERIALS: One hundred four of 108 patients accrued from June 1994 through May 1995 from 26 institutions were analyzable. Patients were histologically confirmed with GBM, and previously untreated. Treatment assignment (52 patients/arm) was based on tumor mass (TM), defined as the product of the maximum diameter and greatest perpendicular dimension of the titanium-gadolinium-enhanced postoperative MRI: Arm A, 64 Gy, TM > 20 cm(2); or Arm B, 70.4 Gy, TM < or = 20 cm(2). Both Arms A and B received BCNU (80 mg/m(2), under hyperhydration) days 1-3, 56-58, then 4 cycles, each 8 weeks, for a total of 6 treatment series. RESULTS: During the 24 months immediately post-treatment, the overall median survival was 9.1 months in Arm A (64 Gy) and 11.0 months in Arm B (70.4 Gy). Median survival in recursive partitioning analysis (RPA) Class III/IV was 10.4 months in Arm A and 12.2 months in Arm B, while RPA Class V/VI was 7.6 months in Arm A and 6.1 months in Arm B. There were no grade 4 neurological toxicities in Arm A; 2 grade 4 neurological toxicities were observed in Arm B (1 motor deficit, 1 necrosis at 157 days post treatment). CONCLUSION: This strategy of high-dose, accelerated hyperfractionated radiotherapy shortens overall RT treatment times while allowing dose escalation, and it provides the potential for combination with currently available, as well as newer, chemotherapy agents. Survival is comparable with previously published RTOG data, and toxicities are within acceptable limits. PMID- 11121634 TI - The prognostic significance of midline shift at presentation on survival in patients with glioblastoma multiforme. AB - PURPOSE: While patients with glioblastoma multiforme (GBM) who present with midline shift have a presumably worse prognosis, there is little literature evaluating the prognostic significance of this presentation in multivariate analysis in the context of other known prognostic factors. METHODS AND MATERIALS: From March 1981 to September 1993, 219 patients underwent irradiation for intracranial glioma at our institution. One hundred fourteen patients with a diagnosis of a primary GBM were analyzed for the influence of the presence of midline shift at diagnosis on survival with respect to other known prognostic factors, including age, Karnofsky performance status (KPS), and extent of surgery. Eighty-five patients (74%) presented with midline shift. Surgical treatment consisted of subtotal/total resection in 86 patients (75%). Among patients presenting with midline shift, 68 (80%) underwent subtotal/total resection before irradiation. RESULTS: Multivariate analysis of the entire cohort of patients found none of the potential prognostic factors analyzed to significantly influence survival. The overall median survival was 6 months. However, when multivariate analysis was limited to patients with a KPS of > or = 70, only the presence of midline shift and age were found to significantly influence survival. Patients with a KPS > or = 70 and with midline shift present at diagnosis had a median survival of 8 months, as compared to 14 months for those not having midline shift at presentation (p = 0.04). Patients with a KPS > or = 70 and age > 50 years had a median survival of 5 months as compared to 11 months for those < or = 50 (p = 0.02). CONCLUSION: In this series, where 80% of patients who presented with a midline shift underwent decompressive resection of GBM before irradiation, the presence of midline shift at diagnosis remained an independent prognostic factor influencing survival among good performance status patients. While the role of decompressive surgery in this setting is likely of some benefit, the extent of this benefit remains to be defined. PMID- 11121635 TI - Benign meningioma: partially resected, biopsied, and recurrent intracranial tumors treated with combined proton and photon radiotherapy. AB - PURPOSE/OBJECTIVE: To evaluate the recurrence-free survival and complications of combined proton and photon radiotherapy of patients with incompletely resected or recurrent histologically-confirmed benign meningioma. METHODS AND MATERIALS: Between May 1981 and November 1996, 46 patients with partially resected, biopsied, or recurrent meningiomas (median age of 50 years; range 11-74 years) were treated with combined photon and 160-MeV proton beam therapy at the Massachusetts General Hospital (MGH) and the Harvard Cyclotron Laboratory, using computed tomography-based conformal 3D treatment planning. Nine patients were treated after incomplete tumor resection, 8 patients after tumor biopsy only, and 29 patients after tumor recurrence following gross total (10/29 patients) or progression after subtotal (19/29 patients) resection. All patients were classified as benign meningioma on review slides at MGH. The median dose to the macroscopic gross tumor volume was 59.0 CGE (range 53.1-74.1 CGE, CGE = proton Gy x 1.1 RBE). The median follow-up was 53 months (range 12-207). RESULTS: Overall survival at 5 and 10 years was 93 and 77%, respectively, and the recurrence-free rate at 5 and 10 years was 100% and 88%, respectively. Survival without severe toxicity was 80% at 5 and 10 years. Three patients presented with local tumor recurrence at 61, 95, and 125 months. One patient developed distant intradural metastasis at 21 and 88 months. No patient died from recurrent meningioma; however, 4 patients died of other causes. A fifth patient died from a brainstem necrosis after 22 months. Eight patients developed severe long-term toxicity from radiotherapy, including ophthalmologic (4 patients), neurologic (4 patients), and otologic (2 patients) complications. All patients with ophthalmologic toxicity received doses higher than those allowed for the optic nerve structures in the current protocol. CONCLUSION: Combined proton and photon radiotherapy is an effective treatment for patients with recurrent or incompletely resected benign intracranial menigiomas. Observed toxicity appears to be dose-related; with currently employed dose constraints, toxicity should not exceed that seen in patients treated with conformal fractionated supervoltage photon radiotherapy. PMID- 11121636 TI - Comparison of intensity-modulated radiotherapy with conventional conformal radiotherapy for complex-shaped tumors. AB - PURPOSE: Conformal and intensity-modulated radiotherapy (IMRT) plans for 9 patients were compared based on characterization of plan quality and effects on the oncology department. METHODS AND MATERIALS: These clinical cases, treated originally with conformal radiotherapy (CRT), required extraordinary effort to produce conformal treatment plans using nonmodulated, shaped noncoplanar fields with multileaf collimators (MLCs). IMRT plans created for comparison included rotational treatments with slit collimator, and fixed-field MLC treatments using equispaced coplanar, and noncoplanar fields. Plans were compared based upon target coverage, target conformality, dose homogeneity, monitor units (MU), user interactive planning time, and treatment delivery time. The results were subjected to a statistical analysis. RESULTS: IMRT increased target coverage an average of 36% and conformality by 10%. Where dose escalation was a goal, IMRT increased mean dose by 4-6 Gy and target coverage by 19% with the same degree of conformality. Rotational IMRT was slightly superior to fixed-field IMRT. All IMRT techniques increased integral dose and target dose heterogeneity. IMRT planning times were significantly less, whereas MU increased significantly; estimated delivery times were similar. CONCLUSION: IMRT techniques increase dose and target coverage while continuing to spare organs-at-risk, and can be delivered in a time frame comparable to other sophisticated techniques. PMID- 11121637 TI - Conventionally fractionated stereotactic radiotherapy (FSRT) for acoustic neuromas. AB - PURPOSE: Analysis of local tumor control and functional outcome following conventionally fractionated stereotactic radiotherapy (FSRT) for acoustic neuromas. PATIENTS AND METHODS: From 11/1989 to 9/1999 51 patients with acoustic neuromas have been treated by FSRT. Mean total dose was 57.6 +/- 2.5 Gy. Forty two patients have been followed for at least 12 months and were subject of an outcome analysis. Mean follow-up was 42 months. We analyzed local control, hearing preservation, and facial and trigeminal nerve functional preservation. We evaluated influences of tumor size, age, and association with neurofibromatosis Type 2 (NF2) on outcome and treatment related toxicity. RESULTS: Actuarial 2- and 5-year tumor control rates were 100% and 97.7%, respectively. Actuarial useful hearing preservation rate was 85% at 2 and 5 years. New hearing loss was diagnosed in 4 NF2 patients. Pretreatment normal facial nerve function was preserved in all cases. Two cases of new or impaired trigeminal nerve dysesthesia required medication. No other cranial nerve deficit was observed. In Patients without NF2 tumor size or age had no influence on tumor control and cranial nerve toxicity. Diagnosis of NF2 was associated with higher risk of hearing impairment (p = 0.0002), the hearing preservation rate in this subgroup was 60%. CONCLUSION: FSRT has been shown to be an effective means of local tumor control. Excellent hearing preservation rates and 5th and 7th nerve functional preservation rates were achieved. The results support the conclusion that FSRT can be recommended to patients with acoustic neuromas where special attention has to be taken to preserve useful hearing and normal cranial nerve function. For NF2 patients, FSRT may be the treatment of choice with superior functional outcome compared to treatment alternatives. PMID- 11121638 TI - Fractionated stereotactic radiotherapy of brain metastases from renal cell carcinoma. AB - PURPOSE: To retrospectively evaluate the effectiveness of fractionated stereotactic radiotherapy (FSRT) for brain metastases from renal cell carcinoma (RCC). METHODS AND MATERIALS: From May 1983 to September 1998, 35 patients with brain metastases from RCC underwent radiotherapy at the National Cancer Center Hospital, Tokyo; 10 patients treated initially with FSRT (FSRT group); 11 with surgery followed by conventional radiotherapy (S/CR group); and 14 with conventional radiotherapy (CR group). Survival and local control rates were determined for patients who had an ECOG performance status of 0-2. RESULTS: Overall median survival rate was 18 months, and actuarial 1- and 2-year survival rates were 57.6% and 31.0%, respectively. Median survival rates were 25.6 months for the FSRT group, 18.7 months for the S/CR group, and 4.3 months for the CR group. Significant prognostic factors associated with survival were age less than 60 years and good performance status. In patients treated with FSRT, imaging studies revealed that 21 of 24 tumors (88%) were locally controlled during a median follow-up time of 5.2 months (range 0.5-68). Actuarial 1- and 2-year local control rates were 89.6% and 55.2%, respectively. No patient suffered from acute or late complications during and following FSRT. CONCLUSIONS: FSRT offers better tumor control and prolonged survival over the S/CR or CR groups, and should be considered as primary treatment for brain metastases from RCC. Patients under 60 years-old and those with a good performance status at the beginning of radiotherapy had a better prognosis. PMID- 11121639 TI - Fractionated stereotactic radiotherapy for vestibular schwannoma (VS): comparison between cystic-type and solid-type VS. AB - PURPOSE: To compare the effectiveness and complications of fractionated stereotactic radiotherapy (SRT) for cystic-type vestibular schwannoma (VS) with those of solid-type VS. METHODS AND MATERIALS: In 65 patients treated with fractionated SRT between 1991 and 1999, 20 were diagnosed with cystic VS, in which at least one-third of the tumor volume was a cystic component on magnetic resonance imaging (MRI), and 45 were diagnosed with solid VS. Thirty-six Gy to 50 Gy in 20-25 fractions was administered to the isocenter and approximately 80% of the periphery of the tumor. All cystic and solid components were included in the gross tumor volume. The mean follow-up period was 37 months, ranging from 6 to 97 months. RESULTS: The actuarial 3-year rate of no episode of enlargement greater than 2.0 mm was 55% for cystic-type and 75% for solid-type VS; the difference was statistically significant (p = 0.023). The actuarial 3-year tumor-reduction (reduction in tumor size greater than 2.0 mm) rates were 93% and 31%, respectively (p = 0.0006). The overall actuarial tumor control rate (no tumor growth greater than 2. 0 mm after 2 years or no requirement of salvage surgery) was 92% at 5 years in 44 patients with a follow-up period of 2 or more years. There was no difference in the class hearing preservation rate between cystic VS and solid VS. No permanent trigeminal or facial nerve palsy was observed in either group. CONCLUSION: Transient tumor enlargement occurs in cystic VS more frequently than in solid-type VS, but the subsequent tumor-reduction rate in cystic VS is better. PMID- 11121640 TI - Prognostic significance of the time of developing motor deficits before radiation therapy in metastatic spinal cord compression: one-year results of a prospective trial. AB - PURPOSE: To investigate prospectively the prognostic value of the time of developing motor deficits before radiation therapy (RT) for post-treatment functional outcome in metastatic spinal cord compression. METHODS AND MATERIALS: From November 1998 until October 1999, 57 patients were included. Two subgroups were formed according to the time of developing motor deficits before RT: 1-14 days (n = 29) and > 14 days (n = 28). Therapeutic effect on motor function was evaluated by an 8-point scale directly, 6, 12, and 24 weeks after RT. Patients with rapid deterioration of motor function within 48 h before RT (n = 14) were evaluated separately. RESULTS: Directly after RT, 26/28 patients (93%) of the group developing motor deficits > 14 days showed improvement of motor function, in comparison to 3/29 patients (10%) of the group 1-14 days (p < 0.001). Deterioration rates were 0% (> 14 days) and 45% (1-14 days). In patients with rapid deterioration of motor function within 48 h before RT, prognosis was poor (improvement 0%, no change 43%, deterioration 57%). Results were comparable 6, 12, and 24 weeks after RT. CONCLUSION: A slower development of motor deficits before RT predicts a better post-treatment functional outcome. In patients with rapid deterioration of motor function within 48 h before RT, prognosis was extraordinarily poor. These results support the findings of our preceding retrospective analysis. PMID- 11121641 TI - Prognostic implication of apoptosis and angiogenesis in cervical uteri cancer. AB - PURPOSE: A retrospective study was performed to investigate the relationship between spontaneous apoptosis and angiogenesis uterine cervix squamous cell carcinoma patients. The prognostic value of each (and both) of these biologic parameters was also tested. METHODS AND MATERIALS: The pathologic materials of 40 cervical uteri squamous cell carcinoma patients were examined and immunohistochemically stained to determine the tumor angiogenesis (tumor microvascular score), using factor VIII-related antigen, and their tumor apoptotic index (AI), using the TdT-mediated dUTP nick end-labeling (TUNEL) method. Three patients were Stage I, 18 were Stage II, 15 were Stage III, and 4 were Stage IV (FIGO classification). All patients were treated with radical radiotherapy and all had follow-up for more than 2 years. RESULTS: The mean AI was 15.1 +/- 12.8, with a median of 8.3. The mean tumor microvascular score was 39.7 +/- 14.4, with a median of 3 8. The patients' age and tumor grade did not seem to significantly affect the prognosis. On the other hand, AI and angiogenesis (tumor microvascular score) were of high prognostic significance. The 3-year disease-free survival (DFS) rate for the patients having AI above the median was 78% (confidence interval [CI] 69-87%), compared to 32% (CI 22-42%) for those having AI below the median. The DFS was 18% (CI 9-27%) for patients having an angiogenesis score above the median, while it was 86% (CI 78-94%) for those patients having a score below the median. CONCLUSION: Determination of both tumor microvascular score and AI can identify patients with the best prognosis of 100% DFS (with low angiogenesis score and high AI). Women with a high score and low AI had the worst prognosis (DFS = 3%, CI 1-5%). Moreover, high AI can compensate partially for the aggressive behavior of tumors showing a high rate of angiogenesis. PMID- 11121642 TI - Quantification of intracavitary brachytherapy parameters and correlation with outcome in patients with carcinoma of the cervix. AB - PURPOSE: To quantify the M. D. Anderson criteria for acceptable implant geometry; to relate our system of intracavitary radiotherapy (ICRT) prescription to Manchester and ICRU reference doses; and to correlate these parameters with outcome measures. METHODS AND MATERIALS: The relationships between intracavitary applicators and normal structures were measured directly from localization films of 808 applications performed in 396 patients who completed definitive treatment for cervical cancer between 1990 and 1994. The distances between applicators and tissue landmarks and the doses to Manchester and normal tissue reference points were correlated with outcome. RESULTS: The median distance from the tandem to the sacrum was 4.0 cm, or one-third the distance from the pubis to the sacrum. The mean distance between the vaginal ovoids and cervical marker seeds was 7 mm, and the median distance between the tandem and the posterior edge of the ovoids was 50% of the ovoid length. In 92% of insertions, vaginal packing was posterior to or within 5 mm of a line that passed through the posterior edge of the ovoids, parallel to the tandem. The median doses to Point A and rectal, bladder, and vaginal surface reference points were 87 Gy, 68 Gy, 70 Gy, and 125 Gy, respectively. Although these reference doses were not routinely used to prescribe treatment, consistent applicator geometry and source selection resulted in a relatively narrow range of delivered doses. The average ratios between the doses at bladder or rectal reference points and Point A were somewhat greater when smaller vaginal applicators were used. Patients received a median of 5600 mgRaEq h from ICRT. The total mgRaEq-h were correlated with but were not proportional to the dose at Point A. There were no significant correlations between the doses to standard reference points and the rates of central recurrence or major complications. CONCLUSION: When ICRT implants are carefully placed, relatively high paracentral doses can be delivered that yield a high rate of central disease control with an acceptable rate of complications. The narrow range of doses delivered to standard reference points and their inconsistent correlation with the maximum doses delivered to normal tissues probably contributed to a lack of correlation between reference doses and outcome. PMID- 11121643 TI - The relevance of adjuvant therapy in primary carcinoma of the fallopian tube, stages I and II: irradiation vs. chemotherapy. AB - INTRODUCTION: Primary carcinoma of the Fallopian tube (FTC) is a rare but extremely aggressive neoplasm. It must be expected to cause up to 40% of tumor related deaths even in Stage I, and up to 57% in Stage II. Due to its rarity, there exist only a few and divergent reports on the value of adjuvant therapy. Therefore the present study aims at evaluating the influence of postoperative adjuvant therapy on FTC by studying the effects of irradiation and chemotherapy on the overall survival of patients in Stages I and II. PATIENTS AND METHODS: We investigated 95 cases of FTC in Stages I (n = 66) and II (n = 29) in a retrospective multicenter study. Group I (n = 32) are patients who underwent a complete irradiation with cobalt or photon energies of 23 MV (administering a daily dose of 2 Gy resulted in a total of 45-52 Gy in the pelvic areas). Group II (n = 31) consists of those cases who received postoperative chemotherapy with platinum. Thirty-two women were excluded from this study because they had other chemotherapies, incomplete irradiation, or no adjuvant therapy at all. RESULTS: Median survival time was 57 months in Group I patients (95% confidence interval 33-81 months), compared to 73 months (95% confidence interval, 68-78 months) in the chemotherapeutically treated Group II. This difference did not prove to be statistically significant (p = 0.476).If primary surgical therapy is included in the evaluation, and patients with total abdominal hysterectomy (TAH) and bilateral salpingo-oophorectomy (BSO) are compared to those with additional radical lymphadenectomy (TAH+BSO+lymph nodes), the latter group's overall survival essentially improves but fails to reach statistical significance. Their 5-year survival rate is 83% against 58% in the TAH+BSO group (p = 0.12). CONCLUSION: Chemotherapy and irradiation are two adjuvant therapies that are similarly effective in FTC of Stages I and II, with chemotherapy being preferred at the present time. Primary surgical treatment, however, is of crucial impact on the prognosis of FTC. PMID- 11121644 TI - An analysis of the impact of pathology review in gynecologic cancer. AB - PURPOSE: To analyze the impact of pathology review in gynecologic malignancies. METHODS AND MATERIALS: For all new gynecologic patients seen between December 2, 1993 and January 4, 1996, we conducted a retrospective chart review to determine if a pathology review by the institute's consultant pathologist changed the diagnosis, and if so whether the change altered patient management. A total of 514 patients were seen, of whom 120 had cervical cancer, 226 had endometrial cancer, 122 had a primary ovarian or peritoneal malignancy, 9 had a vaginal malignancy, 28 had vulvar cancer, and 9 had a miscellaneous gynecologic malignancy. RESULTS: On pathology review the diagnosis changed for 200 of 599 specimens (33%). This altered management for 63 of 514 patients (12%). For patients with cervical cancer, the grade of tumor was the main change in pathologic diagnosis, with occasional change in the presence of lymph vascular invasion. These did not translate into patient management alterations. Eight patients (1.5%) had management alterations. The changes in depth of invasion and vascular invasion altered management for 3 patients. Changes in pap smears resulted in two management alterations, and changes in histologic diagnoses altered management for 3 cases. For endometrial primaries the changes in pathologic diagnosis included grade, depth of invasion, and the presence of cervical involvement. This did alter management in 40 cases (8%). For the ovarian malignancies, the main changes were grade, extent of disease, or histologic classification, some of which (10 patients, 2%) resulted in altered management. One patient with a vaginal lesion had the diagnosis changed, which did alter management. Of the patients diagnosed with vulvar cancer, the pathologic diagnosis changed for 11 patients. This included changes in grade and depth of invasion. This altered management of 2 patients. The remaining miscellaneous gynecologic malignancies had only two diagnosis changes that altered management. CONCLUSIONS: Pathologic review of gynecologic malignancies is justified as it can alter patient management. In addition, the process facilitates cooperation of the multidisciplinary team and provides a valuable educational forum to enhance patient care. PMID- 11121645 TI - Contrast vaginography is more accurate than the radiopaque rod for localization of the vagina. AB - PURPOSE: To compare the radiopaque vaginal rod method with contrast vaginography in localization of the vagina. METHODS AND MATERIALS: In 25 female patients who needed pelvic radiotherapy, both our standard localization procedure using the vaginal rod and a localization procedure using contrast vaginography were performed. As a rod can change the position of the vagina, contrast vaginography was considered to display the true anatomic position of the vagina. The corresponding rod and nonrod X-rays of each patient were compared. The distance from the true vaginal apex to the displaced vaginal apex (= the top of the rod) was measured in the sagittal plane. This distance was called the inaccuracy of the rod method. Furthermore, the size of the vaginal vault was measured using the contrast vaginography. RESULTS: The median inaccuracy of the rod method was 13 mm (range 2 to 24 mm). The maximal width of the vagina ranged from 24 to 68 mm in the frontal plane (median 39 mm) and from 3 to 22 mm in the sagittal plane (median 10 mm). CONCLUSION: The rod method is not accurate to localize the vagina. Furthermore, the rod gives no information on the actual size of the vaginal vault. Contrast vaginography is the method of choice to localize the vagina. PMID- 11121646 TI - Which patients with newly diagnosed prostate cancer need a radionuclide bone scan? An analysis based on 631 patients. AB - PURPOSE: Although radionuclide bone scans are frequently recommended as part of the staging evaluation for newly diagnosed prostate cancer, most scans are negative for metastases. We hypothesized that Gleason score, prostate-specific antigen (PSA), and clinical stage could predict for a positive bone scan (BS), and that a low-risk group of patients could be identified in whom BS might be omitted. METHODS: All patients who had both pathologic review of their prostate cancer biopsies and radionuclide BS at our institution between 1/90 and 5/96 were studied. Gleason score, PSA, and clinical stage (AJCC, 4th edition) were evaluated by univariate and multivariate analyses for their ability to predict a positive BS. Groups analyzed were Gleason of 2-6 vs. 7 vs. 8-10; PSA of 0-15 vs. greater than 15-50 vs. greater than 50; and clinical stage of T1a-T2b vs. T2c-T4. Univariate analysis using chi(2) and multivariate analysis using logistic regression were performed. RESULTS: Of the 631 consecutive patients, 88 (14%) had positive BS. Multivariate analysis (64 excluded due to missing PSA and/or clinical stage) showed Gleason score, PSA, and clinical stage to be significant independent predictors for positive BS (p < 0.002, p < 0.001, p < 0.001, respectively). The odds ratios were 5.25 (confidence interval [CI], 3.43-8.04) for PSA > 50 vs. 0-15; 2.25 (CI, 1.43-3.54) for Gleason of 8-10 vs. 2-6; 2.15 (CI, 1.54-2.99) for clinical stage T2c-T4 vs. T2b or less. Three of 308 (1%) had a positive BS in patients with Gleason 2-7, PSA of 50 or less, and clinical stage of T2b or less. In the subset of the same risk group with PSA of 15 or less, all 237 had negative bone scans. In patients with PSA greater than 50, 49/99(49.5%) had positive BS. CONCLUSION: Gleason score, PSA, and clinical stage were independent predictors for a positive radionuclide BS in newly diagnosed prostate cancer patients. PSA is the major predictor for positive BS. About one-half of the patients analyzed were in the low-risk group (Gleason 2-7, PSA < or = 50, clinical stage < or = T2b) and elimination of BS in these patients would result in considerable economic savings. PMID- 11121647 TI - Differential dosing of prostate and seminal vesicles using dynamic multileaf collimation. AB - PURPOSE: We have investigated the potential of applying different doses to the prostate (PTV2) and prostate/seminal vesicles (PTV1) using multileaf collimation (MLC) for intensity modulated radiation therapy (IMRT). Current dose-escalation studies call for treatment of the PTV1 to 54 Gy in 27 fractions followed by 20 Gy minimum to the PTV2. A daily minimum PTV dose of 2 Gy using a 7-field technique (4 obliques, opposed laterals, and an ant-post field) is delivered. This requires monitor unit calculations, paper and electronic chart entry, and quality assurance for a total of 14 fields. The goal of MLC IMRT is to improve efficiency and deliver superior dose distributions. Acceptance testing and commissioning of the dynamic MLC (DMLC) option on a dual-energy accelerator was accomplished. Most of the testing was performed using segmental MLC (SMLC) IMRT with stop-and-shoot sequences built within the dynamic mode of the DMLC. METHODS AND MATERIALS: The MLC IMRT fields were forward planned using a three-dimensional treatment planning system. The 14 fields were condensed to 7 SMLC IMRT fields with two segments each. In this process, steps were created by moving the leaves to the reduced field positions. No dose (<0.01%) was delivered during this motion. The monitor units were proportioned according to the planned treatment weights. Film and ionization chamber dosimetry were used to analyze leaf positional accuracy and speed, output, and depth-dose characteristics. A geometric phantom was used for absolute and relative measurements. We obtained a volumetric computerized tomography (CT) scan of the phantom, performed 3D planning, and then delivered a single treatment fraction. RESULTS: The acceptance testing and commissioning demonstrated that the leaves move to programmed positions accurately and in a timely manner. We did find an approximately 1 mm offset of the set leaf position and radiation edge (50%) due to the curved-end nature and calibration limitations. The 7-field SMLC IMRT treatment duplicated the 14-field static plan dose distribution with variations no greater than 1.5%. CONCLUSIONS: The MLC IMRT approach will improve efficiency because the number of electronic and chart entries has decreased by a factor of 2. Portal images are able to capture the initial and final MLC segments. The question of differential daily dose to the prostate and seminal vesicles remains. PMID- 11121648 TI - Factors predicting for postimplantation urinary retention after permanent prostate brachytherapy. AB - PURPOSE: Urinary retention requiring catheterization is a known complication among prostate cancer patients treated with permanent interstitial radioactive seed implantation. However, the factors associated with this complication are not well known. This study was conducted to determine these factors. METHODS AND MATERIALS: Ninety-one consecutive prostate cancer patients treated with permanent interstitial implantation at our institution from 1996 to 1999 were evaluated. All patients underwent pre-implant ultrasound and postimplant CT volume studies. Isotopes used were (125)I (54 patients) or (103)Pd (37 patients). Twenty-three patients were treated with a combination of 45 Gy of external beam radiation therapy as well as seed implantation, of which only 3 patients were treated with (125)I. Mean pretreatment prostate ultrasound volume was 35.4 cc (range, 10.0 70.2 cc). The mean planning ultrasound target volume (PUTV) was 39.6 cc (range, 16.1-74.5 cc), whereas the mean posttreatment CT target volume was 55.0 cc (range, 20.2-116 cc). Patient records were reviewed to determine which patients required urinary catheterization for relief of urinary obstruction. The following factors were analyzed as predictors for urinary retention: clinical stage; Gleason score; prostate-specific antigen; external beam radiation therapy; hormone therapy; pre-implant urinary symptoms (asymptomatic/nocturia x 1 vs. more significant urinary symptoms); pretreatment ultrasound prostate volume; PUTV; PUTV within the 125%, 150%, 200%, 250%, 300% isodose lines; postimplant CT volume within the 125%, 150%, 200%, 250%, 300% isodose lines; D90; D80; D50; ratio of post-CT volume to the PUTV; the absolute change in volume between the CT volume and PUTV; number of needles used; activity per seed; and the total activity of the implant. Statistical analyses using logistic regression and chi2 were performed. RESULTS: Eleven of 91 (12%) became obstructed. Significant factors predicting for urinary retention were the total number of needles used (p < 0.038); the pretreatment ultrasound prostate volume (p < 0.048); the PUTV (p < 0.02); and the posttreatment CT volume (p < 0.021). Two of 51 patients (3.9%) requiring 33 or fewer needles (median) experienced obstruction vs. 9 of 40 (22.5%) requiring more than 33 (p < 0.007). If the pretreatment ultrasound prostate volume was 35 cc or less (median), 3 of 43 (7%) vs. 8 of 36 (22%) with a volume greater than 35 cc experienced obstruction (p < 0.051). CONCLUSION: The number of needles required (perhaps related to trauma to the prostate) and the prostate volumes were significant factors predicting for urinary retention after permanent prostate seed implantation. PMID- 11121649 TI - Prostate volume change after radioactive seed implantation: possible benefit of improved dose volume histogram with perioperative steroid. AB - PURPOSE: To evaluate the changes in prostate volume associated with radioactive seed implantation and identify factors that influence prostate swelling. METHODS AND MATERIALS: Between June 1997 and August 1999, 161 patients implanted for prostate carcinoma at the University of California, San Francisco, had prostate volume measurements taken at 4 time points (preplan, preimplant, postimplant, postimplant dosimetry). Patient records were reviewed for treatment with perioperative steroids, hormone therapy (nHT), and external beam radiotherapy (EBRT). One and 2-way analysis of variance (ANOVA) methods were used to test differences in mean effects among patient subsets. RESULTS: A mean 20% volume increase was noted immediately postimplant overall (p < 0.0001), and even with EBRT and/or HT. Steroids were associated with a mean volume decrease of 19.9%, by 3-4 weeks post-procedure (p < 0.0001). Without steroids, only a 3.8% mean change was seen (p = ns). Steroid use resulted in a significant increase in mean dose volume histogram (DVH) (p = 0.001); however, this benefit was only observed among patients who did not receive steroid. A consistently high DVH occurred with steroid use. CONCLUSION: A significant decrease in prostate volume and improved DVH are associated with steroid use. The diminished benefit of steroid use and higher mean DVH achieved in later years suggests the existence of a significant "learning curve" for brachytherapy procedures. PMID- 11121650 TI - The definition of biochemical failure in patients treated with definitive radiotherapy. AB - PURPOSE: The American Society for Therapeutic Radiology and Oncology (ASTRO) published a definition for biochemical failure following treatment of prostate cancer. Others have noted difficulties with interpreting this definition and recommended modifications to accommodate special recurrence patterns. We have compared various modifications to the original ASTRO definition on our series of 1213 patients treated with transperineal permanent prostate brachytherapy. METHODS AND MATERIALS: The ASTRO modifications we considered adjusted for (1) early censoring of nonrecurrent patients with rising prostate-specific antigen levels (PSA), (2) cumulative rather than consecutive rises (without a decrease) as evidence of recurrence, (3) both of the above, and (4) waiting 2 years before data analysis. The Kaplan-Meier method was used to compute the effects on recurrence rate for patients treated with and without neoadjuvant hormones. RESULTS: With the original ASTRO definition, freedom from recurrence in our series of men who did not receive neoadjuvant hormones was 83% at 4 years. All of the modifications considered had statistically insignificant effects on freedom from recurrence rates, varying from 80% to 83% at 4 years. Patients treated with neoadjuvant hormones also showed very little sensitivity to the recurrence definition employed. CONCLUSION: Early censoring of equivocal patients and counting cumulative rather than consecutive rises in PSA (without a decrease) had little empiric effect on the ASTRO recurrence rates. However, we favor the addition of both these modifications to the ASTRO definition on conceptual grounds for evaluating patients following any modality (radiation or surgery), whereby a trend over multiple PSA values is used to judge failure. PMID- 11121651 TI - Recursive partitioning analysis of 1999 Radiation Therapy Oncology Group (RTOG) patients with locally-advanced non-small-cell lung cancer (LA-NSCLC): identification of five groups with different survival. AB - PURPOSE: Survival of patients with locally-advanced non-small-cell lung cancer (LA-NSCLC) is predicted by the stage of the disease and other characteristics. This analysis was undertaken to identify these characteristics in a large cooperative group patient population, as well as to define subgroups of the population with differing outcomes. PATIENTS AND METHODS: Analysis included 1,999 patients treated in 9 RTOG trials between 1983 and 1994 with thoracic irradiation (RT) with (n = 355) or without chemotherapy (CT). RESULTS: In univariate analysis, the following characteristics were significantly associated with an improved survival: use of CT, CT delivered without major deviation, abnormal pulmonary function tests, normal hemoglobin, protein, LDH and BUN, presence of dyspnea, hemoptysis, cough or hoarseness, uninvolved lymph nodes, T1 or T2 stage, no malignant pleural effusion (PE), weight loss of < 8%, Karnofsky performance status (KPS) of at least 90, adenocarcinoma histology, female gender, and age less than 70 years. Recursive partitioning analysis (RPA) was subsequently applied to identify 5 patient subgroups with significantly different median survival times (MST): Group I, KPS of > or = 90, who received chemotherapy (MST 16.2 months); Group II, KPS of > or = 90, who received no CT, but had no PE (MST 11.9 months); Group III, KPS < 90, younger than 70 years, with non-large cell histology (MST 9.6 months); Group IV, KPS > or = 90, but with PE, or KPS < 90, younger than 70 years, and with large cell histology, or older than 70 years, but without PE (MST 5.6-6.4 months); Group V, older than 70, with PE (MST 2.9 months). CONCLUSION: Cisplatinum-based CT improves survival, for excellent prognosis of LA-NSCLC patients, over RT alone. The presence of a malignant pleural effusion is a major negative prognostic factor for survival. The identification of RPA prognostic groups among patients with LA-NSCLC provides prognostic information and may serve as a basis of stratification in future trials. PMID- 11121652 TI - Adjuvant radiotherapy and concomitant 5-fluorouracil by protracted venous infusion for resected pancreatic cancer. AB - PURPOSE: To assess the toxicity and clinical benefit from adjuvant chemoradiotherapy consisting of protracted venous infusion 5-fluorouracil (5-FU) and concomitant radiotherapy in patients with resected pancreatic cancer. METHODS AND MATERIALS: Between 1994 and 1999, 52 patients who underwent pancreaticoduodenectomy received adjuvant chemoradiotherapy. The tumor bed and regional nodes received a dose of 45 Gy in fractions of 1.8 Gy followed by boost to the tumor bed if the surgical margins were involved (total dose, 54 Gy). The patients also received concomitant 5-FU by protracted venous infusion (200-250 mg/m(2)/day, 7 days/week) during the entire radiotherapy course. RESULTS: Fifty two patients (30 men, 22 women) were enrolled and treated on this protocol. The median age was 63 years (range, 38-78 years), and the median Karnofsky Performance Status was 80 (range, 70-100). Thirty-five percent had involved surgical margins and 59% had involved lymph nodes. All patients completed therapy, and there were no Grade IV/V toxicities observed. With median follow-up of 24 months (range, 3-52 months) for surviving patients, the median survival is 32 months, and 2-year and 3-year survivals are 62%, and 39%, respectively. CONCLUSION: Radiotherapy with concomitant 5-FU by protracted venous infusion as adjuvant treatment for resected pancreatic cancer is well tolerated. This approach allows for greater dose intensity with reduced toxicity. The median survival of this cohort of patients compares favorably with our earlier experience and other published series. PMID- 11121653 TI - Long-term effects in children treated with radiotherapy for head and neck rhabdomyosarcoma. AB - PURPOSE: To examine the long-term effects of treatment in children receiving radiotherapy for head and neck rhabdomyosarcoma. METHODS: From 1967 to 1994, a total of 30 children with head and neck rhabdomyosarcoma received megavoltage radiotherapy at one institution. Seventeen patients (57%) have survived and have at least a 5-year follow-up. There were 11 males and 6 females, with a median age of 5.7 years (range 2.2-11.6) at the time of radiotherapy. Tumor location was orbit in 6 patients, infratemporal fossa in 4, paranasal sinuses in 2, and supraglottic larynx in 2; the nasopharynx, pterygopalatine fossa, and parotid gland were sites for the remaining children. All but 2 patients had tumors of embryonal histology. The Intergroup Rhabdomyosarcoma Study (IRS) Group was I in 2, II in 3, and III in 11 children; 1 patient had a recurrent tumor after surgery alone. Radiotherapy volume was the primary tumor or tumor bed in 13, tumor and whole brain in 3, and tumor and craniospinal axis in 1. Median radiotherapy dose to the primary site was 5,040 cGy (range 4,140-6,500) and to the whole brain was 3,000 cGy. All but 1 were treated with 150-200-cGy fractions; 1 patient received 250-cGy fractions for a tumor in the larynx. Chemotherapy was vincristine (V), actinomycin-D (A), and cyclophosphamide (C) in 10 patients, VAC + adriamycin in 2, VA in 1, VA + ifosfamide in 1, VC + adriamycin in 1, and none in 2. One patient had salvage chemotherapy consisting of cisplatin and etoposide. Median follow-up time was 20 years (range 7.5-33). RESULTS: Late effects of treatment were seen in all patients and included facial growth retardation in 11, neuroendocrine dysfunction in 9, visual/orbital problems in 9, dental abnormalities in 7, hearing loss in 6, and hypothyroidism in 3. Intellectual and academic delays were documented in 3 patients who had received whole brain radiotherapy. While neuroendocrine, thyroid, dental, and cognitive sequelae were primarily attributed to radiotherapy, hearing loss was thought to be a direct result of tumor destruction and, in 1 case, cisplatin chemotherapy. Late effects at or beyond 10 years from radiotherapy were few, but severe, and included chondronecrosis, esophageal stenosis, second malignancy, and brain hemorrhage. CONCLUSION: Late effects of treatment in children receiving radiotherapy for head and neck rhabdomyosarcoma are frequent. Although radiotherapy is a significant contributor of neuroendocrine, dental, thyroid, and cognitive toxicity, it is not usually implicated with hearing loss. Late toxicity of treatment beyond 10 years is not as frequent as those occurring within 10 years of therapy. PMID- 11121654 TI - Prospective evaluation of delayed central nervous system (CNS) toxicity of hyperfractionated total body irradiation (TBI). AB - PURPOSE: Prospective evaluation of chronic radiation effects on the healthy adult brain using neuropsychological testing of intelligence, attention, and memory. METHODS AND MATERIALS: 58 patients (43 +/- 10 yr) undergoing hyperfractionated total body irradiation (TBI) (TBI, 14.4 Gy, 12 x 1.2 Gy in 4 days) before bone marrow or peripheral blood stem cell transplantation were prospectively included. Twenty-one recurrence-free long-term survivors were re-examined 6-36 months (median 27 months) after completion of TBI. Neuropsychological testing included assessment of general intelligence, attention, and memory using normative, standardized psychometric tests. Mood status was controlled, as well. Test results are given as IQ scores (population mean 100) or percentiles for attention and memory (population mean 50). RESULTS: The 21 patients showed normal baseline test results of IQ (101 +/- 13) and attention (53 +/- 28), with memory test scores below average (35 +/- 21). Test results of IQ (98 +/- 17), attention (58 +/- 27), and memory (43 +/- 28) showed no signs of clinically measurable radiation damage to higher CNS (central nervous system) functions during the follow-up. The mood status was improved. CONCLUSION: The investigation of CNS toxicity after hyperfractionated TBI showed no deterioration of test results in adult recurrence-free patients with tumor-free CNS. The median follow-up of 27 months will be extended. PMID- 11121655 TI - Thallium-201 scintigraphy is not predictive of late cardiac complications in patients with Hodgkin's disease treated with mediastinal radiation. AB - PURPOSE: To assess whether abnormalities depicted by Thallium-201 scintigraphy can predict the occurrence of late cardiac complications in patients with Hodgkin's disease treated with mantle field radiation therapy. METHODS AND MATERIALS: Thallium scintigraphy was performed in 49 patients at a median of 75 months after initial treatment (range 28-208 months). Initial treatment consisted in chemotherapy, given to two-thirds of the patients and mantle field radiation, delivered to all patients, using a 25-MV linear accelerator. Myocardial perfusion defects were observed in 78% of patients on thallium scintigraphy. These patients had their cardiac status reassessed at a median follow-up of 13.5 years after treatment. RESULTS: Forty-two patients were assessable, as data on the cardiac status were missing in 7 patients. The majority of patients received at least 40 Gy, and 75% of them were treated with one field per day. The median follow-up of patients is 13.5 years (range 9-24.5). Eleven cardiac complications were observed in 9 patients (coronary artery disease [n = 2], conduction-system abnormalities [n = 3], valvular defects [n = 5], and congestive heart disease [n = 1]). The median 15-year actuarial incidence of cardiac complications was 21% (95% confidence interval of 9-40%). The positive and negative predictive value of thallium scintigraphy was 19% and 77%, respectively. The univariate analysis showed that the extent of left ventricle exposure to irradiation was an adverse prognostic factor, and chemotherapy administered before mantle field irradiation was of borderline significance. CONCLUSION: Thallium scintigraphy is not predictive of late cardiac complications. The extent of left ventricle exposure to radiation and possibly chemotherapy given before radiation treatment are adverse prognostic factors. PMID- 11121657 TI - Enhancement of radiotherapy by hyperthermia-regulated gene therapy. AB - PURPOSE: Interleukin 12 (IL-12) has shown strong antitumoral effects in numerous pre-clinical studies and appears to act synergistically with radiation in murine tumors. The major impediment to its clinical use has been its systemic toxicity. While using intratumorally injected viral gene therapy vectors encoding IL-12 reduces systemic side effects substantially, elevated systemic transgene levels are still observed because adenovirus can reach the circulation. Further restricting IL-12 expression in the tumor is therefore desirable in a combined radiation and adenovirus mediated cancer gene therapy regimen. METHODS AND MATERILAS: Hyperthermia-regulated gene therapy was tested in a nonimmunogenic B16.F10 melanoma line that is syngeneic with C57BL/6 mice. For hyperthermic gene therapy, an adenoviral vector coding for IL-12 under the control of the promoter of the human heat shock protein 70B (hsp70B) was used. One week after transplantation (at a 5-7 mm diameter), tumors were irradiated with 3 x 11 Gy (mo we-fri). Adenovirus was injected at 3 x 10(8) pfu/tumor 24 h before the last radiation fraction or 3 days afterwards. Hyperthermia was performed 24 h later at 42.5 degrees C. Growth delay to reaching 3 times initial tumor volume was chosen as the biologic endpoint. IL-12 levels in tumor and serum were determined by using the enzyme-linked immunosorbant assay (ELISA). RESULTS: Adenovirus mediated intratumoral expression of IL-12 under the control of a heat inducible promoter in combination with hyperthermia is almost as effective as that under the control of a constitutive cytomegaly virus (CMV) promoter while systemic transgene levels are substantially reduced with the heat inducible promoter. The response to radiotherapy is improved considerably when combined with heat inducible gene therapy without apparent systemic toxicity. When used as a single dose, applying IL-12 gene therapy after completion of radiotherapy appears to be beneficial. CONCLUSIONS: Hyperthermia-regulated gene therapy in combination with radiation is feasible and therapeutically effective in murine tumors with no apparent systemic toxicity. PMID- 11121656 TI - Adenoviral-mediated p53 transgene expression sensitizes both wild-type and null p53 prostate cancer cells in vitro to radiation. AB - PURPOSE/OBJECTIVE: The effect of adenoviral-mediated p53 transgene expression on the radiation response of two human prostate cancer cell lines, the p53(wild type) LNCaP and p53(null) PC3 lines, was examined. The objective was to determine if this vector sensitizes cells to radiation independently of their p53 status. METHODS AND MATERIALS: A recombinant adenovirus-5 vector (RPR/INGN 201, Introgen Therapeutics, Houston, TX) containing a CMV promoter and wild-type p53-cDNA (Ad5 p53) was used to facilitate p53 transgene expression. A multiplicity of infection (MOI) of 10-40 viral particles per cell was used, based on Ad5/CMV/lacz infection and staining for the beta-galactosidase reporter gene product. Clonogenic assays were performed to evaluate the degree of sensitization to radiation of viral transduced cells compared with irradiated nontransduced controls. The relative efficacy of these treatments to induce apoptotic cell death was determined using the TUNEL assay. RESULTS: The delivery of Ad5-p53 (10 MOI) reduced control plating efficiency from 36.5% to 0.86% in the LNCaP cell line and from 75.1% to 4.1% in the PC3 cell line. After correcting for the effect of Ad5-p53 on plating efficiency, the surviving fraction after 2 Gy (SF2) of gamma-irradiation was reduced over 2.5-fold, from 0.187 to 0.072, with transgene p53 expression in the LNCaP cell line. Surviving fraction after 4 Gy (SF4) was reduced over 4.5-fold, from 0.014 to 0.003, after Ad5-p53 treatment. In the PC3 cell line, Ad5-p53 (40 MOI) reduced SF2 over 1.9-fold from 0.708 to 0.367, and SF4 over 6-fold from 0.335 to 0.056. In both the LNCaP and PC3 cell lines, the combination of Ad5-p53 plus radiation (2 Gy) resulted in supra-additive apoptosis (approximately 20% for LNCaP and approximately 15% for PC3 at 50 MOI), above that seen from the addition of the controls; control vector Ad5-pA plus RT (0.15% for LNCaP and 1.44% for PC3), Ad5-p53 alone (28.6% for LNCaP and 21.7% for PC3), RT alone (0% for LNCaP and 0.23% for PC3), or Ad5-pA alone (0.1% for LNCaP and 0.29% for PC3). CONCLUSION: The clonogenic survival and apoptosis data demonstrate that p53 transgene expression sensitizes human prostate adenocarcinoma cells in vitro to irradiation. As this effect was observed in both the p53(wild-type) LNCaP and p53(null) PC3 lines, radiosensitization was independent of p53 status. PMID- 11121658 TI - Radiosensitization of human breast cancer cells by a novel ErbB family receptor tyrosine kinase inhibitor. AB - PURPOSE: Overexpression of the ErbB family of growth factor receptors is present in a wide variety of human tumors and is correlated with poor prognosis. The purpose of this study was to determine the effects of a novel small molecule ErbB tyrosine kinase inhibitor, CI-1033, in combination with ionizing radiation on breast cancer cell growth and survival. MATERIALS & METHODS: Growth assays were performed on ErbB-overexpressing human breast cancer cells developed in our laboratory in the presence of 0.1-1.0 microM CI-1033 (Parke Davis). Clonogenic survival assays were performed in the presence of ionizing radiation with or without CI-1033. For some experiments, clonogen numbers, defined as the product of surviving fraction and total number of cells, were calculated at each time point during a course of multifraction radiation. RESULTS: CI-1033 potently inhibited the growth of ErbB-overexpressing breast cancer cells. A single 48-h exposure of 1 microM CI-1033 resulted in growth inhibition for 7 days, whereas three times weekly administration resulted in sustained growth inhibition. Clonogenic survival was modestly decreased after a 7-day exposure to CI-1033. Exposure to both CI-1033 and radiation (6 Gy) yielded a 23-fold decrease in clonogenic survival compared to radiation alone. In a multifraction experiment, exposure to CI-1033 and three 5-Gy fractions of gamma radiation decreased the total number of clonogens in the population by 65-fold compared to radiation alone. CONCLUSION: CI-1033 results in potent growth inhibition and modest cytotoxicity of ErbB-overexpressing breast cancer cells, and has synergistic effects when combined with ionizing radiation. These data suggest that CI-1033 may have excellent clinical potential both alone and in combination with radiation therapy. PMID- 11121659 TI - Changes in tumor hypoxia measured with a double hypoxic marker technique. AB - PURPOSE: Development of a double hypoxic cell marker assay, using the bioreductive nitroimidazole derivatives CCI-103F and pimonidazole, to study changes in tumor hypoxia after treatments that modify tumor oxygenation. METHODS AND MATERIALS: Both hypoxic markers were visualized by immunohistochemical techniques to detect changes in hypoxic fraction induced by carbogen breathing (95% O(2) and 5% CO(2)) or hydralazine injection. The protocol was tested in a human laryngeal squamous cell carcinoma xenograft line. Quantitative measurements were derived from consecutive tissue sections that were analyzed by a semiautomatic image analysis system. Qualitative analysis was obtained by double staining of the two hypoxic markers on the same tissue section. RESULTS: A significant correlation between the hypoxic fractions for the two markers, CCI 103F and pimonidazole, was found in air breathing animals. After carbogen breathing, the hypoxic fraction decreased significantly from 0.07 to 0.03, and after hydralazine treatment, the hypoxic fraction increased significantly. Reduction of hypoxia after carbogen breathing was most pronounced close to well perfused tumor regions. CONCLUSIONS: With this method, employing two consecutively injected bioreductive markers, changes in tumor hypoxia can be studied. A significant reduction in hypoxia after carbogen breathing and a significant increase in hypoxia after hydralazine administration was demonstrated. PMID- 11121660 TI - In-field and out-of-field effects in partial volume lung irradiation in rodents: possible correlation between early dna damage and functional endpoints. AB - PURPOSE: Recent observations have shown that there are regional variations in radiation response in mouse lung as measured by functional assays. Furthermore, there are both in-field and out-of-field effects in radiation-induced lung damage as observed by DNA assay in rats. The purpose of this work is: (a) to examine mice lethality data following partial volume lung irradiation to assess the possibility of directional or regional effects, (b) to evaluate the correlation between mice lethality data and DNA damage assayed by micronuclei production in rat lung, and (c) to re-interpret mice lethality considering the existence of directional effects in lung cellular response to partial volume irradiation. METHODS AND MATERIALS: The lethality data for mice, generated at the M. D. Anderson Cancer Center, Houston, and micronuclei yield data for rats obtained at Princess Margaret Hospital, Toronto, were used. A radiobiological model that allows for out-of-field and in-field effects for lung cell damage and lung response was developed. This model is based on the observation of DNA damage in shielded parts of rat lung that was assumed relevant to cell lethality and consequently overall lung response. RESULTS: While the experimental data indicated directional or regional volume effects, the applicability of dose and volume as sole predictors of lung response to radiation was found to be unreliable for lower lung (base) irradiation in mice. This conforms well to rat lung response where micronuclei were observed in shielded apical parts of lung following base irradiation. The radiobiological model, which was specifically developed to account for the lung response outside of primary irradiated volume, provides a good fit to mice lethality data, using parameters inferred from rat micronuclei data. CONCLUSION: Response to lung irradiation in rodents, in particular, elevated sensitivity to base irradiation, can be interpreted with a hypothesis of in-field and out-of-field effects for cellular response. If the existence of these effects for lung is subsequently proven in humans, it will require the incorporation of geometrical and directional information in normal tissue complication probability calculations for lung. These considerations are ignored in present approaches based only on conventional dose-volume histograms. PMID- 11121661 TI - Estimation of optimum dose per fraction for high LET radiations: implications for proton radiotherapy. AB - PURPOSE: For high linear energy transfer (LET) radiations, the relative biologic effect (RBE) changes with dose per fraction. Methods for calculating the optimum dose per fraction for high LET radiations should therefore include an allowance for RBE. METHODS AND MATERIALS: The linear-quadratic (LQ) model, and the associated biologic effective dose (BED) concept, has previously been extended to incorporate the RBE effect. Differential calculus is now used to calculate the optimum dose per fraction (z), when high-LET radiation is used, which is given by the solution for z of (g - LATE(alpha/beta)(L)/TUM(alpha/beta)L . RBE(M( z(2))) - 2 . f . g . K . z - (LATE)(alpha/beta)(L) . f . K . RBE(M) = 0 where g is the normal tissue sparing factor, RBE(M) is the maximum RBE value, f the mean interfraction interval, K the daily low-LET BED equivalent dose for clonogen repopulation and (LATE)(alpha/beta)(L) and (TUM)(alpha/beta)(L) are the respective late reacting normal tissue and tumor fractionation sensitivities for low-LET radiation. RESULTS: The optimum dose per fraction for proton therapy is generally lower than that calculated for photons but there is not a simple relationship between the magnitude of the reduction and the assumed value of RBE(M.) Thus(,) generic values of RBE(M) cannot always be used in such calculations. In some cases, where tumor alpha/beta ratios are low (around 5-6 Gy) and where there is good normal tissue sparing, the optimum dose per fraction is relatively large, typically 4-8 Gy. CONCLUSION: BED equations that include the RBE parameter, together with low-LET alpha/beta ratios and repopulation dose equivalents, constitute a rational model of high-LET radiotherapy. In the case of proton beam therapy, a wide range of optimum dose per fraction is predicted. PMID- 11121662 TI - Intensity modulation to improve dose uniformity with tangential breast radiotherapy: initial clinical experience. AB - PURPOSE: We present a new technique to improve dose uniformity and potentially reduce acute toxicity with tangential whole-breast radiotherapy (RT) using intensity-modulated radiation therapy (IMRT). The technique of multiple static multileaf collimator (sMLC) segments was used to facilitate IMRT. METHODS AND MATERIALS: Ten patients with early-stage breast cancer underwent treatment planning for whole-breast RT using a new method of IMRT. The three-dimensional (3D) dose distribution was first calculated for equally weighted, open tangential fields (i.e., no blocks, no wedges). Dose calculation was corrected for density effects with the pencil-beam superposition algorithm. Separate MLC segments were constructed to conform to the beam's-eye-view projections of the 3D isodose surfaces in 5% increments, ranging from the 120% to 100% isodose surface. Medial and lateral MLC segments that conformed to the lung tissue in the fields were added to reduce transmission. Using the beam-weight optimization utility of the 3D treatment planning system, the sMLC segment weights were then determined to deliver the most uniform dose to 100 reference points that were uniformly distributed throughout the breast. The accuracy of the dose calculation and resultant IMRT delivery was verified with film dosimetry performed on an anthropomorphic phantom. For each patient, the dosimetric uniformity within the breast tissue was evaluated for IMRT and two other treatment techniques. The first technique modeled conventional practice where wedges were derived manually without consideration of inhomogeneity effects (or density correction). A recalculation was performed with density correction to represent the actual dose delivered. In the second technique, the wedges were optimized using the same beam weight optimization utility as the IMRT plan and included density correction. All dose calculations were based on the pencil-beam superposition algorithm. RESULTS: For the sMLC technique, treatment planning required approximately 60 min. Treatment delivery (including patient setup) required approximately 8-10 min. Film dosimetry measurements performed on an anthropomorphic phantom generally agreed with calculations to within +/- 3%. Compared to the wedge techniques, IMRT with sMLC segments resulted in smaller "hot spots" and a lower maximum dose, while maintaining similar coverage of the treatment volume. A median of only 0.1% of the treatment volume received > or = 110% of the prescribed dose when using IMRT versus 10% with standard wedges. A total of 6-8 segments were required with the majority of the dose delivered via the open segments. The addition of the lung-block segments to IMRT was of significant benefit for patients with a greater proportion of lung parenchyma within the irradiated volume. Since August 1999, 32 patients have been treated in the clinic with the IMRT technique. No patient experienced RTOG grade III or greater acute skin toxicity. CONCLUSION: The use of intensity modulation with an sMLC technique for tangential breast RT is an efficient and effective method for achieving uniform dose throughout the breast. It is dosimetrically superior to the treatment techniques that employ only wedges. Preliminary findings reveal minimal or no acute skin reactions for patients with various breast sizes. PMID- 11121663 TI - Dosimetric improvements following 3D planning of tangential breast irradiation. AB - PURPOSE: To evaluate the dosimetric difference between a simple radiation therapy plan utilizing a single contour and a more complex three-dimensional (3D) plan utilizing multiple contours, lung inhomogeneity correction, and dose-based compensators. METHODS AND MATERIALS: This is a study of the radiation therapy (RT) plans of 85 patients with early breast cancer. All patients were considered for breast-conserving management and treated by conventional tangential fields technique. Two plans were generated for each patient. The first RT plan was based on a single contour taken at the central axis and utilized two wedges. The second RT plan was generated by using the 3D planning system to design dose-based compensators after lung inhomogeneity correction had been made. The endpoints of the study were the comparison between the volumes receiving greater than 105% and greater than 110% of the reference dose, as well as the magnitude of the treated volume maximum dose. Dosimetric improvement was defined to be of significant value if the volume receiving > 105% of one plan was reduced by at least 50% with the absolute difference between the volumes being 5% or greater. The dosimetric improvements in 49 3D plans (58%) were considered of significant value. Patients' field separation and breast size did not predict the magnitude of improvement in dosimetry. CONCLUSION: Dose-based compensator plans significantly reduced the volumes receiving > 105%, >110%, and volume maximum dose. PMID- 11121664 TI - Comparison of dose length, area, and volume histograms as quantifiers of urethral dose in prostate brachytherapy. AB - PURPOSE: To determine the magnitude of the differences between urethral dose volume, dose-area, and dose-length histograms (DVH, DAH, and DLH, respectively, or DgH generically). METHODS AND MATERIALS: Six consecutive iodine-125 ((125)I) patients and 6 consecutive palladium-103 ((103)Pd) patients implanted via a modified uniform planning approach were evaluated with day 0 computed tomography (CT)-based dosimetry. The urethra was identified by the presence of a urinary catheter and was hand drawn on the CT images with a mean radius of 3.3 +/- 0.7 mm. A 0.1-mm calculation matrix was employed for the urethral volume and surface analysis, and urethral dose points were placed at the centroid of the urethra on each 5-mm CT slice. RESULTS: Although individual patient DLHs were step-like, due to the sparseness of the data points, the composite urethral DLH, DAH, and DVHs were qualitatively similar. The DAH curve delivered more radiation than the other two curves at all doses greater than 90% of the prescribed minimum peripheral dose (mPD) to the prostate. In addition, the DVH curve was consistently higher than the DLH curve at most points throughout that range. Differences between the DgH curves were analyzed by integrating the difference curves between 0 and 200% of the mPD. The area-length, area-volume, and volume-length difference curves integrated in the ratio of 3:2:1. The differences were most pronounced near the inflection point of the DgH curves with mean A(125), V(125), and L(125) values of 36.6%, 31.4%, and 23.0%, respectively, of the urethra. Quantifiers of urethral hot spots such as D(10), defined as the minimal dose delivered to the hottest 10% of the urethra, followed the same ranking: area analysis indicated the highest dose and length analysis, the lowest dose. D(10) was 148% and 136% of mPD for area and length evaluations, respectively. Comparing the two isotopes in terms of the amount of urethra receiving a given dose, (103)Pd implants were significantly cooler than (125)I implants over most of the range of clinical interest, from 100% to 150% of mPD. CONCLUSION: Dose gradients in prostate implants result in the observed ordering of DAH, DVH, and DLH from higher to lower doses. The three histogram approaches remain in close agreement up to 100% of the mPD but diverge at higher doses. Although urethral point doses are the most easily determined, they underestimate the amount of urethra at risk at higher doses compared to dose area analysis. Because dosimetric parameters detailing high-dose regions such as D(10) show only slight differences between calculation methods, they are recommended over the corresponding geometric entities G(150) or G(175). The differences between the D(gg) entities are sufficiently small that they are unlikely to be of clinical significance or to confound analyses attempting to correlate urinary morbidity with urethral dosimetry. PMID- 11121665 TI - Real-time optimized intraoperative dosimetry for prostate brachytherapy: a pilot study. AB - PURPOSE: To assess the feasibility of real-time intraoperative treatment planning for permanent prostate brachytherapy analyzing the impact on operative time and adequacy of postimplant dosimetry. METHODS AND MATERIALS: Seventeen consecutive patients undergoing permanent brachytherapy for prostate cancer had real-time intraoperative computer-based and optimized treatment planning. The first 8 patients were implanted using a plan generated before the surgery and served to assure the team qualitatively that this could be performed without greatly increasing intraoperative time. They served as control group for expected achieved dosimetry results reviewing the D90, V100, V150 parameters from the dose volume histograms. The next 9 patients were implanted according to the real-time plan. The times needed to carry out various steps of the procedure were recorded. The achieved dosimetry was then compared to the control group to assure that accuracy was unchanged. RESULTS: The median operative time for patients undergoing intraoperative dosimetry was 57 min. Of this, 21 min were devoted to anesthesia and nursing functions. Postoperative dosimetry showed a median achieved V100 (volume of prostate receiving 100% of prescribed dose) of 97% for the control group. For the real-time dosimetry group, the median V100 was similar at 94%. The V150 (volume receiving 150%) is 49% for both groups. The D90 (dose received by 90% of the target) was normalized for easy comparison and was consistently slightly greater than the prescription dose. CONCLUSION: Treatment planning for permanent brachytherapy of prostate cancer has historically been performed as a computer-generated and optimized plan run weeks in advance of an implant, or according to a set pattern using an intraoperative nomogram. These data show that planning can now be optimized intraoperatively using widely available software without compromising the operative time or implant quality. PMID- 11121666 TI - Use of an implanted marker and real-time tracking of the marker for the positioning of prostate and bladder cancers. AB - PURPOSE: A real-time tracking radiotherapy was investigated to assess its usefulness in precise localization and verification of prostate and bladder cancers. METHODS AND MATERIALS: The real-time tracking radiation therapy (RTRT) system consists of implantation of a 2.0-mm gold marker into a clinical target volume (CTV), three-dimensional radiation treatment planning (3DRTP) system, and the use of two sets of diagnostic x-ray television systems in the linear accelerator room, image processing units, and an image display unit. The position of the patient can be corrected by adjusting the actual marker position to the planned marker position, which has been transferred from the 3DRTP and superimposed on the fluoroscopic image on the display unit of the RTRT system. The position of the markers can be visualized during irradiation and after treatment delivery to verify the accuracy of the localization. Ten patients with prostate cancer and 5 patients with bladder cancer were examined using this system for the treatment setup on 91 occasions. RESULTS: After manual setup using skin markers, the median of absolute value of discrepancies between the actual position of the marker and the planned position of the marker for prostate cancer was 3.4 (0.1-8.9) mm, 4.1 (0.2-18.1) mm, and 2.3 (0.0-10.6) mm for the lateral, anteroposterior, and craniocaudal directions, respectively. The 3D median distance between the actual and planned positions of the marker was 6.9 (1.1 18.2) mm for prostate cancer and 6.9 (1.7-18.6) mm for bladder cancer. After relocation using RTRT, the 3D distance between the actual and planned position of the marker was 0.9 +/- 0.9 mm. Median 3D distances between actual positions after treatment delivery and planned positions were 1.6 (0.0-6.3) mm and 2.0 (0.5-8.0) mm during daily radiotherapy for the marker in patients with prostate cancer and bladder cancer, respectively. CONCLUSION: We believe the new positioning system can reduce uncertainty due to setup error and internal organ motion, although further improvement is needed for the system to account for the rotational and elastic changes of the affected tissues. PMID- 11121667 TI - A geometrically based method for automated radiosurgery planning. AB - PURPOSE: A geometrically based method of multiple isocenter linear accelerator radiosurgery treatment planning optimization was developed, based on a target's solid shape. METHODS AND MATERIALS: Our method uses an edge detection process to determine the optimal sphere packing arrangement with which to cover the planning target. The sphere packing arrangement is converted into a radiosurgery treatment plan by substituting the isocenter locations and collimator sizes for the spheres. RESULTS: This method is demonstrated on a set of 5 irregularly shaped phantom targets, as well as a set of 10 clinical example cases ranging from simple to very complex in planning difficulty. Using a prototype implementation of the method and standard dosimetric radiosurgery treatment planning tools, feasible treatment plans were developed for each target. The treatment plans generated for the phantom targets showed excellent dose conformity and acceptable dose homogeneity within the target volume. The algorithm was able to generate a radiosurgery plan conforming to the Radiation Therapy Oncology Group (RTOG) guidelines on radiosurgery for every clinical and phantom target examined. CONCLUSIONS: This automated planning method can serve as a valuable tool to assist treatment planners in rapidly and consistently designing conformal multiple isocenter radiosurgery treatment plans. PMID- 11121668 TI - Intensity-modulated whole pelvic radiation therapy in patients with gynecologic malignancies. AB - PURPOSE: To evaluate the ability of intensity-modulated radiation therapy (IMRT) to reduce the volume of small bowel irradiated in women with gynecologic malignancies receiving whole pelvic radiotherapy (WPRT). METHODS AND MATERIALS: Ten women with cervical (5) or endometrial (5) cancer undergoing WPRT were selected for this analysis. A planning CT scan of each patient was obtained following administration of oral, i.v., and rectal contrast. The clinical target volume (CTV) was defined as the proximal vagina, parametrial tissues, uterus (if present), and regional lymph nodes. The CTV was expanded uniformly by 1 cm in all directions to produce a planning target volume (PTV). The bladder, rectum, and small bowel were also delineated in each patient. Two plans were created: a standard "4-field box" with apertures shaped to the PTV in each beam's eye view and an IM-WPRT plan designed to conform to the PTV while minimizing the volume of normal tissues irradiated. Both plans were normalized to deliver 45 Gy to the PTV. Isodose distributions and dose-volume histograms (DVH) were compared. RESULTS: The IM-WPRT plan reduced the volume of small bowel irradiated in all 10 patients at doses above 30 Gy. At the prescription dose, the average volume of small bowel irradiated was reduced by a factor of two (17.4 vs. 33.8%, p = 0.0005). In addition, the average volume of rectum and bladder irradiated at the prescription dose was reduced by 23% in both cases (p = 0.0002 and p = 0.0005, respectively). The average PTV doses delivered by the conventional and IM-WPRT plans were 47.8 Gy and 47.4 Gy, respectively. Corresponding maximum doses were 50.0 Gy and 54.8 Gy, respectively. However, on average, only 3.2% of the PTV received greater than 50.0 Gy in the IM-WPRT plans. CONCLUSION: Our results suggest that IM-WPRT is an effective means of reducing the volume of small bowel irradiated in women with gynecologic malignancies receiving WPRT. This approach potentially offers a method for reducing small bowel complications in patients with gynecologic malignancies. PMID- 11121670 TI - Erratum PMID- 11121669 TI - Decreasing the dosimetric effects of misalignment when using a mono-isocentric technique for irradiation of head and neck cancer. AB - PURPOSE: The purpose of this study was to quantify and develop methods to decrease inhomogeneities created with field edge mismatch when using a mono isocentric beam-split technique. METHODS AND MATERIALS: We validated techniques to determine dose across a half-blocked field edge and quantified potential sources of systematic matchline error. Then, two methods were used to evaluate matchline doses. The first used film dosimetry data from a half-beam field and a spreadsheet. Duplication and reversal provided two columns, each representing a beam-split field edge. Summation simulated perfect abutment and shifting created various gaps and overlaps. The second method involved obtaining dose profiles at midfield along the ray perpendicular to abutted, overlapped, and gapped beam split fields on six linear accelerators. To enlarge the penumbra, we designed several field edge modifiers, then re-evaluated matchline doses. The field edge modifiers applicability to a 3-field head and neck treatment technique was also examined. RESULTS: Film-determined dose profiles provide similar information across a beam-split field edge as an ionization chamber. With the mono-isocentric beam-split technique, a 4-mm overlap or gap produces inhomogeneities nearly 60% above or below the intended dose. A 2-mm overlap or gap produces inhomogeneities nearly 30% above or below the intended dose. A customized penumbra generator decreased the magnitude of these inhomogeneities to 20% and 10%, respectively. CONCLUSION: The two methods of evaluating matchline dose described above gave similar results. When using the mono-isocentric half-field technique, small misalignments produce worrisome regions of inhomogeneity. Our penumbra generator substantially decreases the magnitude of the dose inhomogeneities, although the volume receiving an inhomogeneous dose increases. PMID- 11121671 TI - Fatty acid composition of the milk lipids of Fulani women and the serum phospholipids of their exclusively breast-fed infants. AB - We previously reported that, relative to milk of women elsewhere in the world, the lipid fraction of milk of Fulani women in northern Nigeria contained relatively low proportions of alpha-linolenic acid and docosahexaenoic acid (DHA). This led us to question the essential fatty acid status of Fulani infants and the relation between the proportion of critical n-3 and n-6 fatty acids in the serum phospholipids of the mothers, their milk, and the serum phospholipids of their exclusively breast-fed infants. We were also interested in the effect de novo intermediate chain length-fatty acids (C10-C14) had on the proportions of critical and non-essential fatty acids in milk. Capillary gas-liquid chromatography was used to analyze the fatty acid content of the total milk lipids of 34 Fulani women, as well as the fatty acid content of serum phospholipids of the women and their breast-fed infants during the first 6 months of life. The proportions of critical n-3 and n-6 fatty acids in the milk of the Fulani women were adequate, but the proportions of these same fatty acids were low in their exclusively breast-fed infants. The serum phospholipids of the infants contained 18.8% linoleic acid, 0.13% alpha-linolenic acid, 12.8% arachidonic acid, and 3.40% DHA, whereas, the mean percentages of linoleic, alpha linolenic, arachidonic and DHA in the serum phospholipids of the Fulani mothers' were 21.4, 0.20, 9.79, and 1.97, respectively. There was a strong positive correlation between fatty acid content of serum phospholipids of Fulani women and the fatty acid content of their milk lipids. As the proportion of C10-C14 fatty acids in the milk lipids increased, the proportions of critical n-3 and n-6 fatty acids in milk remained relatively constant; however, proportions of three non essential fatty acids decreased dramatically. C10-C14 fatty acids do not appear to displace critical n-3 and n-6 fatty acids in milk. PMID- 11121672 TI - Proxy informed consent in pediatric research: a review. AB - This paper is aimed at discussing the issue of proxy consent for medical research with children carried out in the context of developed countries. First, requirements for valid informed consent are reviewed, and differences with clinical practice highlighted. In the second part the findings from empirical studies, and implications for improving the consent process, are discussed. Perceived benefit for their child is the most important factor motivating parents to grant consent, but also a desire to contribute to medical research and benefit others are frequently mentioned. Abstract concepts such as randomization are more difficult to grasp and remember than practical issues. The type and style of the consent-seeking process (quality of the information, physician's attitude, allocated time, readability of consent forms) have an influence on how the invitation to participate is received. Rather than as a one-sided delivery of information, consent should be viewed as a continuous, two-way communication process developing in a context of transparency and partnership between the investigator and potential research subjects. PMID- 11121673 TI - The influence of postural control on motility and hand function in a group of 'high risk' preterm infants at 1 year of age. AB - The functional outcome of a group of 75 'high-risk' preterm infants was studied at the corrected age of 12 months. Only infants with high-risk for developmental deviance with gestational ages below 32 weeks and/or birthweights less than 1500 g were included in the study. Additionally the infants were categorised according to their medical history conforming with the 'Neonatal Medical Index' (NMI I to V), with category I describing infants with few medical problems and V characterizing those with the most serious complications. In this study we included only infants with 'high-risk' as categorised in NMI III to V, since infants with 'low-risk' have been described earlier. Infants with cerebral ultrasonographic abnormalities were incorporated into the NMI categories, but also analysed separately to compare outcomes. At 12 months (corrected age) apart from pediatric follow-up, a full neurological assessment was done with emphasis on postural control, spontaneous motility and hand function. Special attention was given to symmetrical development. The infants were then categorised as having optimal, non-optimal or asymmetrical outcomes. An overall optimal outcome on postural control was found in 64% of all infants (67% in NMI III, 60% in NMI IV and 62.5% in NMI V). Too much extension interfering with postural control was found significantly more often in infants in NMI V (15%), compared to infants in NMI IV (8%) and NMI III (4.5%). Poor postural control had a significant influence on other domains of development such as motility (P=0.00), asymmetry (P=0.00) and hand function (P=0.00). Cerebral ultrasonographic abnormalities seemed to have an influence on motility (P=0.03), while no direct relationship was found with postural control, hand function or asymmetry. It is unclear whether this poor coordination of gross motor function will have consequences for appropriate visual-motor and sensorimotor integration therewith hampering later cognitive function, as often described in preterm infants. It is suggested that the poor postural control found in many infants is the result of both myogenic and/or neurogenic deviations and associated with cerebral pathology, but is also caused by the preterm birth and its nursing consequences. PMID- 11121674 TI - Gender differences in cognitive abilities at 2 years in ELBW infants. Extremely low birth weight. AB - Gender differences in cognitive abilities exist for children born at term. For very preterm infants uncertainty exists regarding the presence and extent of such differences and their relationship to perinatal brain injury and neurological impairment. This study examined gender differences in cognitive abilities in a cohort of 336 extremely low birth weight (ELBW) infants at 2 years corrected age. Infants were classified as at low or high perinatal risk at birth according to four perinatal risk factors. A subgroup of 33 neurologically impaired infants was identified. Outcome at 2 years was measured by the overall General Quotient (GQ) on the Griffiths scale and its five subscale scores. Female ELBW children were superior to male ELBW children by 4.1 GQ points (95% CI 1.0, 7.1). If the impaired subgroup was excluded, the difference in GQ was 3.2 points (95% CI 0.4, 5.6), and this difference was predominantly due to female infants being superior in the hearing and speech subscale (6.0 points, 95% CI 2.6, 9.5). These differences were relatively independent of perinatal risk status. Gender differences in the Griffiths GQ for ELBW infants are similar to expected differences for term infants and are unlikely to cause substantial bias in interpreting outcome studies for ELBW infants, unless these involve tests of specific cognitive abilities such as language. PMID- 11121675 TI - Developmental outcome, child behaviour and mother-child interaction at 3 years of age following Newborn Individualized Developmental Care and Intervention Program (NIDCAP) intervention. AB - The aim of the present pilot study was to investigate the impact of early intervention in the form of family-centred developmentally supportive care according to NIDCAP((R)) (Newborn Individualized Developmental Care and Assessment Program) on the development and behaviour of the child and on the mother-child interaction at 3 years of age. Two groups of very-low-birth-weight (VLBW) infants (< or = 1500 g) were studied. The control group (n=21) was born in 1990, i.e. prior to implementation of the NIDCAP. The intervention group, born in 1992-1993 (n=21), was subjected to formal NIDCAP observation once every 10 days. Development was assessed using the Griffiths' Developmental Scale II in conjunction with a neurological examination. Behaviour was assessed on the basis of a parental interview. Mother-child interaction was assessed according to the Parent-Child Early Relational Assessment Scale (ERA). There was no significant difference in motor development. The total developmental quotient (DQ) on the Griffiths' developmental scale was 109 (94-122) [median (range)] for the NIDCAP group and 108 (93-120) for the control group (n.s.). On the subscale hearing speech, the intervention group scored 119 (72-157) and the control group 108 (84 130) (P=0.02). The total score with respect to the Behaviour Symptom Interview was 6 (0-20) for the NIDCAP group and 16 (0-54) for the control group (P=0.03). With respect to the mother-child interaction, there was a significant difference in the child cluster 'communication', the total score being 12 (11-13) for the NIDCAP group and 10 (9-13) for the control group (P=0.03). In conclusion, care of VLBW infants according to NIDCAP appears to have certain positive long-term effects on the child's behaviour and mother-child interaction. PMID- 11121676 TI - Socio-demographic associations with digit and pacifier sucking at 15 months of age and possible associations with infant infection. The ALSPAC Study Team. Avon Longitudinal Study of Pregnancy and Childhood. AB - AIMS: To assess the prevalence of pacifier and digit sucking at 15 months of age and to investigate whether this habit adversely affects the health of 18 month old infants. STUDY DESIGN: Data collected via self-completion questionnaires from mothers forming part of the prospective, population based Avon Longitudinal Study of Pregnancy and Childhood. SUBJECTS AND METHODS: The mothers of 10006 infants gave information on their child's use of a pacifier and of digit sucking at 15 months of age and the presence of specific health symptoms at 18 months of age. Adjusted logistic regression was performed to identify any statistically significant associations between pacifier use, digit sucking or a combination of the two with possible infection. RESULTS: 36.3% of infants sucked a pacifier, 21. 3% their thumb or finger and 2.7% sucked both at 15 months. Statistically significant differences were observed among various socio-demographic variables. Mothers were more likely to give their child a pacifier if they were younger, had lower levels of education, experienced greater financial difficulties or lived in council housing (compared to owned/mortgaged). The opposite was apparent for digit suckers. After allowing for these possible confounding factors, pacifier users had a higher incidence of earache and colic compared to children with no sucking habit, however digit suckers had a lower incidence of these symptoms. Children who sucked both were significantly more likely to have reported wheezing, earache, and poor health in the past month. CONCLUSIONS: Significantly different sociodemographic characteristics were observed with pacifier suckers compared to those who sucked their thumb or finger. It is almost impossible to attribute the direction of causality between infection and a sucking habit. Further and more detailed studies are needed before any recommendations can be made based on the statistically significant associations found as they are unlikely to be of major clinical significance. PMID- 11121677 TI - Serum soluble E- and L-selectin in the very early neonatal period. AB - Both E- and L-selectin are cell adhesion molecules. E-selectin is expressed by activated endothelial cells, whereas L-selectin by quiescent leukocytes and is rapidly cleaved off after activation. Both selectins take part in the first step of the 'adhesion cascade', the 'rolling of leukocytes', leading to the extravasation of the white cells to the sites of inflammation, infection or damage. For this reason their soluble forms (sE- and sL-selectin, respectively), are considered early and reliable markers of the immune activation and response. Moreover, sE-selectin has been reported to be a potent angiogenic factor and a reliable marker of infection and sepsis in neonates, as well as endothelial activation, while sL-selectin of the leukocyte function and maturity. Following informed maternal consent, we evaluated prospectively by ELISA, sE- and sL selectin in the serum of 40 (19 females, 21 males), healthy, term, infection-free neonates, on the second and fifth day of life, and compared them with the respective values in 20 healthy adults (10 females, 10 males), with the purpose of examining the pattern of their values in the early postpartum days, and to establish reference values for both selectins. Values (mean+/-S.D.) of sE selectin both on the second (139+/-48 ng/ml) and fifth day of life (111+/-35 ng/ml) were found to be highly increased, as compared with those in controls (48+/-13 ng/ml; P<4 x 10(-11) and P<4 x 10(-10), respectively), while sL-selectin values on both the second (674+/-223 ng/ml) and the fifth day of life (684+/-221 ng/ml), were significantly lower than those in controls (938+/-181 ng/ml); P<0.0001 and P<0.0003, respectively). A significant decrease was noted in sE selectin values, from the second to the fifth day of life (P<10(-7)), while sL selectin values showed no significant change in the same time interval. A strong correlation was found between values on the second and the fifth day of life of both sE- and sL-selectin (r(P)=0.885 and r(P)=0.813, respectively; P<0.00001). Neonatal values of both sE- and sL-selectin on the second or on the fifth day of life, did not depend on the perinatal factors, neonatal sex, or birth weight, mode of delivery, and maternal age or parity. In conclusion, in the very early neonatal period, our findings of highly increased sE-selectin, while low sL selectin, suggest an immune and more specifically endothelial activation and an immature and decreased leukocyte function. PMID- 11121678 TI - Alex Comfort (1920-2000) known and unknown. A personal account. PMID- 11121679 TI - Experimental gerontology in Belgium: from model organisms to age-related pathologies. PMID- 11121680 TI - Research on aging in France. AB - This article gives some information on the French research on aging using a survey of the European Commission on research on aging in Europe and the answers to a questionnaire sent to the laboratories involved in aging research. France is involved in the European effort, but to a lesser degree than could be expected from its population size, and this picture has not changed during the last decade. The answers to the questionnaire allow us to know the opinion of colleagues on the strengths and weaknesses of French research: the lack of money and institutional recognition seem to hinder the development of experimental gerontology in France. PMID- 11121681 TI - Cellular and molecular mechanisms of stress-induced premature senescence (SIPS) of human diploid fibroblasts and melanocytes. AB - Replicative senescence of human diploid fibroblasts (HDFs) or melanocytes is caused by the exhaustion of their proliferative potential. Stress-induced premature senescence (SIPS) occurs after many different sublethal stresses including H(2)O(2), hyperoxia, or tert-butylhydroperoxide. Cells in replicative senescence share common features with cells in SIPS: morphology, senescence associated beta-galactosidase activity, cell cycle regulation, gene expression and telomere shortening. Telomere shortening is attributed to the accumulation of DNA single-strand breaks induced by oxidative damage. SIPS could be a mechanism of accumulation of senescent-like cells in vivo. Melanocytes exposed to sublethal doses of UVB undergo SIPS. Melanocytes from dark- and light- skinned populations display differences in their cell cycle regulation. Delayed SIPS occurs in melanocytes from light-skinned populations since a reduced association of p16(Ink 4a) with CDK4 and reduced phosphorylation of the retinoblastoma protein are observed. The role of reactive oxygen species in melanocyte SIPS is unclear. Both replicative senescence and SIPS are dependent on two major pathways. One is triggered by DNA damage, telomere damage and/or shortening and involves the activation of the p53 and p21(waf-1) proteins. The second pathway results in the accumulation of p16(Ink-4a) with the MAP kinase signalling pathway as possible intermediate. These data corroborate the thermodynamical theory of ageing, according to which the exposure of cells to sublethal stresses of various natures can trigger SIPS, with possible modulations of this process by bioenergetics. PMID- 11121683 TI - Postponed aging and desiccation resistance in Drosophila melanogaster. AB - Studies with the fruit fly, Drosophila melanogaster, have repeatedly shown that selection for postponed reproduction leads to increases in mean life span and increased stress resistance; including increased resistance to desiccation, starvation and ethanol vapors. We show that desiccation resistance declines with age in both short- and long-lived flies suggesting that desiccation resistance may serve as a useful biomarker for aging-related declines in physiological performance. We examined the physical basis of desiccation resistance in five replicate populations selected for postponed reproduction and five replicate control populations. The variables examined were water content, rates of water loss during desiccation, and water content at time of death due to desiccation. In the absence of desiccation stress, both the flies exhibiting postponed senescence and their controls maintained constant water content throughout their lifetimes. In the presence of desiccation stress, the short-lived flies showed significantly higher rates of water loss at all ages than did the long-lived flies. Flies from the two treatments did not differ in water content at death. Our results indicate that water loss rates are the major determinant of desiccation resistance. Water loss rates are under genetic control and covary with age in populations with genetically-determined postponed senescence. PMID- 11121682 TI - Molecular control of bone remodeling and osteoporosis. AB - Osteoprotegerin ligand (OPGL, TNFS11) and its receptor RANK (TNFRS11A) are essential for the development and activation of osteoclasts and critical regulators of physiological bone remodeling and osteoporosis. Production of OPGL by activated T cells can directly regulate osteoclastogenesis and bone remodeling. This may explain why autoimmune diseases, cancers, leukemias, asthma and chronic viral infections such as hepatitis and HIV result in systemic and local bone loss. OPGL is also the pathogenetic factor that causes bone and cartilage destruction and clinical crippling in arthritis. Inhibition of OPGL binding to RANK via the natural decoy receptor osteoprotegerin (OPG) prevents bone loss in postmenopausal osteoporosis and cancer metastases and completely blocks crippling in a rat model of arthritis. Moreover, OPG expression is induced by estrogen which provides a molecular explanation of postmenopausal osteoporosis in women. PMID- 11121684 TI - Anticipation of oxidative damage decelerates aging in virgin female medflies: hypothesis tested by statistical modeling. AB - Empirical analysis of survival data obtained from large samples of Mediterranean fruit flies shows that the trajectory of the mortality rate for virgin females departs from that for females maintained in mixed sex cages. It increases, decelerates, reaches its maximum, declines and then increases again within the reproductive interval. Non-virgin females, however, display an early-age plateau instead of this dip. We assume that these deviations are produced by the interplay between changes in oxygen consumption associated with reproductive behavior and the antioxidant defense that acts against anticipated oxidative damage caused by reproduction. Since there are no data on antioxidant mechanisms in medflies available that explain the observed patterns of mortality, we develop a model of physiological aging based on oxidative stress theory, which describes age-related changes in oxygen consumption and in antioxidative capacity during the reproductive period. Using this model, we simulate virtual populations of 25,000 virgin and non-virgin flies, calculate the respective mortality rates and show that they practically coincide with those of experimental populations. We show that the hypothesis about the biological support of reproduction used in our model does not contradict experimental data. The model explains how the early-age dip and plateau might arise in the mortality rates of female medflies and why the male mortality pattern does not exhibit such deviations. PMID- 11121685 TI - Comparative characterisation of poly(ADP-ribose) polymerase-1 from two mammalian species with different life span. AB - DNA damage induced in higher eukaryotes by alkylating agents, oxidants or ionising radiation triggers the synthesis of protein-conjugated poly(ADP-ribose) catalysed by poly(ADP-ribose) polymerase-1 (PARP-1). Previously, cellular poly(ADP-ribosyl)ation capacity has been shown to correlate positively with the life span of mammalian species [Proc. Natl. Acad. Sci. USA 89 (1992) 11,759 11,763]. Here, we have tested whether this correlation results from differences in kinetic parameters of the enzymatic activity of PARP-1. We therefore compared recombinant enzymes, expressed in a baculovirus system, from rat and man as two mammalian species with extremely divergent life span. In standard activity assays performed in the presence of histones as poly(ADP-ribose) acceptors both enzymes showed saturation kinetics with [NAD(+)]. The kinetic parameters (k(cat), k(m) and k(cat)/k(m)) of the two enzymes were not significantly different. However, in assays assessing the auto-poly(ADP-ribosyl)ation reaction, both enzymes displayed second-order kinetics with respect to [PARP-1], and up to two-fold higher specific activity was observed for human versus rat PARP-1. We conclude that the correlation of poly(ADP-ribosyl)ation capacity with life span is not reflected in the kinetic parameters, but that subtle differences in primary structure of PARP 1 from mammalian species of different longevity may control the extent of the automodification reaction. PMID- 11121686 TI - Age-related decline in mitogen-activated protein kinase phosphorylation in PTH stimulated rat enterocytes. AB - In the present study we analyzed whether parathyroid hormone (rPTH[1-34]; PTH) stimulates the tyrosine phosphorylation of the growth-related protein mitogen activated protein (MAP) kinases (p42/44-MAPK), also known as extracellular signal regulated kinases (ERK1/2), in duodenal enterocytes isolated from young (3months) and aged (24months) rats. Western blot analysis revealed that PTH rapidly stimulates MAPK phosphorylation. The hormone effects on MAPK were evident within 30s, peaking at 1min (4-fold). PTH response was dose-dependent (10(-11)-10(-7) M) with maximal stimulation achieved at 10(-9)-10(-8) M. PTH-induced MAPK phosphorylation was effectively suppressed by the tyrosine-kinase inhibitors, genistein (100microM) and herbimycin (2microM). Moreover, the tyrosine phosphorylation and activation of MAPK was dependent on Src kinase, since PP1 (10 and 20microM), a specific Src family tyrosine-kinase inhibitor, blocked PTH induced MAPK activation. With aging, the response to PTH was significantly reduced. However, The amount of basal protein expression determined by Western blot analysis for MAPK was not different in the enterocytes from young and aged rats. In conclusion, the results obtained in this work expand our knowledge on the mechanism of action of PTH in duodenal cells, revealing that protein tyrosine phosphorylation is linked to the PTH regulation of enterocyte MAPK activation, and that this mechanism is impaired with aging. Understanding the molecular mechanisms for the age-related differences in PTH signaling will require more information about the subtle mechanisms that modulate the PTH receptor-MAPK signaling pathway. PMID- 11121687 TI - Interaction of indomethacin with cytokine production in whole blood. Potential mechanism for a brain-protective effect. AB - Both Alzheimer's disease and vascular dementia are featured by inflammatory responses and it is known that non-steroidal anti-inflammatory drugs (NSAIDs) decrease the risk and severity of these diseases. To study the effect of NSAIDs on PGE2 levels and pro- and anti-inflammatory cytokine levels in the whole blood assay, blood samples from 23 elderly persons aged 85 years were stimulated with thrombin or LPS as primary stimulus. Indomethacin was added in concentrations ranging from 0.4 to 16 microg/ml and acetylsalicylic acid was added to in concentrations ranging from 0.5 to 8.0 microg/ml. Indomethacin abrogated thrombin and LPS-induced PGE2 production at all concentrations tested. In addition, indomethacin reduced the production of thrombin-induced IL-6 and IL-10 (p<0.05) at physiological concentrations. Indomethacin reduced the production of LPS induced IL-6, IL-1 beta and IL-10 (p<0.05) at the highest indomethacin concentration tested. Similar results were obtained upon incubation with acetylsalicylic acid. It is concluded that indomethacin may reduce the thrombin induced inflammatory reaction by decreasing IL-6 through inhibition of PGE2 synthesis. This IL-6 reduction may be relevant for the ability of indomethacin to reduce the risk of Alzheimer's disease. However, the decrease in IL-10 production due to indomethacin suggests a more inflammatory state. PMID- 11121688 TI - Band 3 protein clustering on human erythrocytes promotes binding of naturally occurring anti-band 3 and anti-spectrin antibodies. AB - Recognition of senescent and oxidatively stressed human erythrocytes appeared to be initiated by band 3 clustering, followed by bivalent binding of naturally occurring anti-band 3 autoantibodies (anti-band 3 NAbs), and complement deposition. The number of RBC-associated anti-band 3 NAbs was, however, low compared to the total amount of IgG that bound in vitro to RBC containing band 3 oligomers. This implied the involvement of yet other types of NAb, among which we focussed on anti-spectrin NAbs, since eluates from RBC of thalassemic patients contained these NAbs. Binding of affinity-purified anti-band 3 and anti-spectrin NAbs was studied to RBC on which band 3 oligomers were generated by exoplasmic cross-linking. This pretreatment increased binding not only of (125)I-iodinated anti-band 3, but also of anti-spectrin NAbs by 7-10-fold at 0 degrees C in the presence of nearly physiological IgG and HSA concentrations. Binding of anti spectrin NAbs was not to spectrin as judged from surface-labeling of RBCs that were pretreated with cross-linker. Binding was dose and time dependent in both cases. Moreover, binding of anti-spectrin NAbs was not competed by high concentrations of anti-band 3 NAbs and anti-spectrin NAbs even stimulated binding of anti-band 3 F(ab')(2) by 30%. This suggests that anti-spectrin NAbs bound to band 3 or a protein associated with band 3 by virtue of their inherent polyreactivity. PMID- 11121689 TI - Binding of anti-spectrin antibodies to red blood cells and vesiculation in various in vivo and in vitro ageing conditions in the rat. AB - In this study, the binding of naturally occurring antibodies as well as of induced anti-spectrin antibodies to red blood cells (RBC), in relation with different ageing conditions, was investigated in the rat. RBC from aged animals, or from rats whose RBC were age-induced either by means of hypertransfusion (which blocks erythropoiesis) or by treatment with clodronate-containing liposomes (which reduces RBC removal from circulation), were used. Attainment of RBC ageing was demonstrated by MCV reduction and by an increase of both RBC density and 4.1a/4.1b RBC membrane protein ratio. The results demonstrate an augmented anti-spectrin antibody binding to RBC in relation with their ageing condition, especially when induced by hypertransfusion. The vesiculation process was also investigated and correlated with antibody binding: vesicles were found only in the plasma of clodronate-treated rats, whose RBC showed the lowest level of anti-spectrin antibody binding with respect to the other groups. In addition, RBC preserved in vitro in different media showed a binding of anti-spectrin antibody, which inversely correlated with the vesiculation process. On the whole, the latter results suggest a protective effect of vesicles towards IgG opsonization of aged RBC. PMID- 11121690 TI - Changes in the insulin-like growth factor-system may contribute to in vitro age related impaired osteoblast functions. AB - Age-related bone loss is thought to be due to impaired osteoblast functions. Insulin-like growth factors (IGFs) have been shown to be important stimulators of bone formation and osteoblast activities in vitro and in vivo. We tested the hypothesis that in vitro osteoblast senescence is associated with changes in components of the IGF-system including IGF-I, IGF-II, IGF-binding proteins (IGFBPs) and IGFBP-specific proteases. We employed a human diploid osteoblast cell line obtained from trabecular bone explants and that exhibit typical characteristics of in vitro senescence during serial subculturing. Using a non competitive reverse-transcriptase polymerase-chain reaction (RT-PCR) assay, we found that the constitutive level of IGF-I mRNA decreased progressively to 49.9 +/- 4.9% in old osteoblasts as compared to the levels found in the young cells. No age-related change was found in IGF-II steady-state mRNA levels. Changes in IGFBPs gene expression and protein production were assessed using Northern blot analysis and Western ligand blotting (WLB), respectively. IGFBP-3 mRNA levels decreased to 30% and protein production to 16% in aged osteoblasts as compared to levels found in young cells. We also found age-related decreases in mRNA levels of both IGFBP-4 and IGFBP-5 to 70% and 60% in aged osteoblasts, respectively, compared to young cells. While IGFBP-5 protein was not detected by WLB, IGFBP-4 protein production showed a biphasic change with 50% decrease in middle-aged cells and a subsequent increase in aged osteoblasts to levels similar to those in young osteoblasts. We found an age-related increase in the immunoreactive levels of IGFBP-4 protease, however, no detectable IGFBP-4 or IGFBP-3 protease activities in conditioned media from osteoblast cultures were observed. Our findings demonstrate that osteoblast aging is associated with impaired production of the stimulatory components of the IGF-system, that may be a mechanism contributing to age-related decline in osteoblast functions. PMID- 11121691 TI - Age-related histomorphometric changes in labial salivary glands with special reference to the acinar component. AB - PURPOSE: To evaluate histomorphometric age-related changes in labial salivary glands (LSG) of healthy subjects, with special reference to the mucous and seromucous acini. METHOD: Investigation of age-related histomorphometric changes was conducted on 120 samples of LSG obtained from autopsy subjects free of salivary gland tumors/diseases. Samples were divided into young (< 30 years, n = 30), adult (30-60 years, n = 45) and old (> 60 years, n = 45) age groups. Measurements were performed on hematoxylin and eosin stained slides. Parenchymal components included acini and ducts. The acinar component was presently analyzed separately in respect to mucous acinar cells and seromucous acinar cells. Mucous acinar cells presented pale eosinophilic cytoplasm, while that of the seromucous was darker and basophilic. Stromal components included connective tissue, blood and lymphatic vessels, inflammatory infiltrate and adipose tissue. The mean volume fraction (V(v)) of all components in each age group was calculated. Statistical analysis was performed by one-way ANOVA and Tamhane tests. RESULTS: The mean V(v) of the seromucous acinar cells decreased by 49.3% (p < 0.0001) and that of the mucous acinar cells decreased only by 28.5% (p = 0.0001). Changes in the ducts were statistically non-significant (p = 0.312). Of the stromal components, the mean V(v) of the adipose tissue showed the highest increase, 2768% (p < 0.0001) followed by the inflammatory infiltrate, 512% (p < 0.0001). CONCLUSIONS: The marked decrease in the V(v) of seromucous acinar cells accompanied by a lower decrease in the V(v) of mucous acinar cells may explain age-related changes in LSG secretion. These findings could bear important implications regarding functional age-related changes of these glands. PMID- 11121692 TI - Ad libitum fed rats can keep lean or grow fat when they age. PMID- 11121694 TI - Combining independent component analysis and correlation analysis to probe interregional connectivity in fMRI task activation datasets. AB - A new approach in studying interregional functional connectivity using functional magnetic resonance imaging (fMRI) is presented. Functional connectivity may be detected by means of cross correlating time course data from functionally related brain regions. These data exhibit high temporal coherence of low frequency fluctuations due to synchronized blood flow changes. In the past, this fMRI technique for studying functional connectivity has been applied to subjects that performed no prescribed task ("resting" state). This paper presents the results of applying the same method to task-related activation datasets. Functional connectivity analysis is first performed in areas not involved with the task. Then a method is devised to remove the effects of activation from the data using independent component analysis (ICA) and functional connectivity analysis is repeated. Functional connectivity, which is demonstrated in the "resting brain," is not affected by tasks which activate unrelated brain regions. In addition, ICA effectively removes activation from the data and may allow us to study functional connectivity even in the activated regions. PMID- 11121695 TI - An investigation of the impulse functions for the nonlinear BOLD response in functional MRI. AB - Functional MRI (fMRI) based on blood oxygenation level dependent (BOLD) contrast can be used to detect hemodynamic responses to a broad range of stimuli. It however remains unclear in what fashion the BOLD response is a linear system, and how the impulse function differs with stimulation of varying duration. To address this question, fMRI using visual stimulation with a wide range of duration (0.5 12 s) was performed in six human volunteers. A strong linear correlation was shown on the full width at half maximum (r = 0.998) of the BOLD response curves and the area under the curves (r = 0.999) to the duration of stimulation. However, comparing the errors of the measured and predicted response curves, our results showed a poorer linearity at stimuli of shorter duration. By examining the impulse functions derived from different stimuli, based on the assumption that a linear convolution relationship existed, a higher differentiation was shown in the experiments with shorter stimuli (<3 s). Compared to the area under the impulse function derived from 12 s stimulation, with that obtained from 0.5, 1, 2, 3, 4, 8 s stimuli resulted in differences of 66.2, 33.5, 15.1, 5.4, 0.9, 7.9%, respectively. This study suggests a higher degree of nonlinearity in the BOLD signal changes due to stimuli of shorter duration, in agreement with earlier work. PMID- 11121696 TI - Fast 3D T(2)-weighted MRI with Hadamard encoding in the slice select direction. AB - A fast method to obtain 3-dimensional (3D) magnetic resonance imaging with long repetition times is presented. It can be used to obtain fast 3D MRI with for example T(2) or diffusion weighted imaging. The method uses a 3D multiple thin slab sequence with radio frequency encoding, preferably Hadamard encoding, in the slice select direction. The point-spread function of the Hadamard-encoded slices is close to ideal even at low encoding numbers. This allows the acquisition of 3D data volumes with tolerable image quality up to four times faster than is possible using Fourier phase encoding. The scope of the method includes both longitudinal and transverse encoding. Longitudinal encoding provides a better point spread function than transverse encoding, at the expense of having to discard one slice per slab. The method is demonstrated experimentally for 4th order longitudinal Hadamard encoding to obtain 3D T(2)-weighted images. PMID- 11121697 TI - Analysis of the Look-Locker T(1) mapping sequence in dynamic contrast uptake studies: simulation and in vivo validation. AB - An alternative to the pulse sequences at present used in dynamic contrast uptake MRI is the dynamic LL-EPI T(1) mapping method. This method generates T(1) estimates in a few seconds, thereby allowing dynamic studies. A particular advantage of the LL-EPI technique is that it provides the opportunity to generate spatial and temporal information about the paramagnetic contrast agent concentration independently of the inflow rate. This paper illustrates, by computer simulations, the accuracy of the estimated 1/T(1) value when using the LL-EPI technique in situations that are not supported by the model. The simulated situations not supported by the model are those in which the longitudinal and transversal relaxation rates change during the T(1) mapping. The most critical moment occurs during a bolus passage of contrast agent when the concentration gradient is large. The computer simulations of the LL-EPI T(1) mapping method in non-supported situations show that in normal perfused capillary tissue the error in the estimated 1/T(1) value is within the absolute error of 0.1 s(-1) in most simulated situations, although in a typical vessel the simulations do indicate that the stated absolute error tolerance of 0.5 s(-1) is exceeded relatively easily. However, this transgression can be rectified by a non-bolus injection of the contrast agent media. PMID- 11121698 TI - Sequential use of gadolinium chelate and mangafodipir trisodium for the assessment of focal liver lesions: initial observations. AB - The purpose of this study was to assess the feasibility of sequential administration of 2 different MR imaging contrast agents using a single visit protocol to image focal liver abnormalities. Twenty-one patients with known or suspected liver lesions were included in the study. All patients received a bolus intravenous injection of gadolinium chelate (Gd) and dynamically enhanced imaging performed. The patients then received an injection of mangafodipir trisodium (Mn) contrast and a second scan performed with an average delay of 62 min after the Gd bolus injection. The images were evaluated to determine the appearance of liver lesions after administration of each contrast agent, and for evidence of prior Gd administration adversely affecting evaluation of images acquired after Mn administration. Focal liver lesions were present in 19 patients, including 8 with liver metastases, 1 with liver lymphoma, 6 with hemangiomas, 3 with focal nodular hyperplasia (FNH), and 1 with hepatic abscess. In 2 other patients no liver lesions were identified in either the post-Gd or post-Gd-post-Mn scans. All malignant lesions identified on the post-Gd scan were also identified on post-Gd post-Mn scans. Although the potential benefit for increasing detection sensitivity for hepatic metastases was not demonstrated, this is a preliminary series. This study does demonstrate the practicality for use a single visit sequential Gd-Mn protocol described here, with possible application of this technique for further assessment of the utility of combining Gd and Mn for detection of liver metastases. PMID- 11121700 TI - Pretibial cyst formation after anterior cruciate ligament reconstruction using auto hamstring grafts: two case reports in a prospective study of 89 cases. AB - Eighty-nine cases after anterior cruciate ligaments (ACL) reconstruction were followed prospectively with magnetic resonance imaging (MRI). The patients were examined using axial and sagittal MRI at least twice during the postoperative evaluation of reconstructed ACL. Two cases of pretibial cyst formation were observed. At the time of cyst formation, neither patient had any subjective or objective evidence of knee instability. The cyst of one case communicated with the intra-articular. The minimum follow-up period after the surgical excision was 9 months, with no evidence of recurrence. We might speculate that the critical period for cyst formation in both patients occurred at less than 12 months after their ACL reconstruction. We concluded that the cyst formation was most likely due to incomplete graft tendon incorporation within the osseous tunnel. PMID- 11121699 TI - The influence of the resolution and contrast on measuring the articular cartilage volume in magnetic resonance images. AB - The progression of OA in patients may be followed by measuring the volume of articular cartilage from MR images. We attempted to determine the reproducibility of volume measurements of articular cartilage made from magnetic resonance images of the knees and the dependence of the reproducibility on image resolution and contrast-to-noise. A fat-suppressed 3D technique was used to generate four image sets with different image resolution. Each patient was imaged twice to obtain image pairs at each resolution. To assess the dependence of reproducibility on noise we generated six image sets for each patient by adding noise to the original images and repeating the comparison. On each image set, the femoral, tibial, and patellar cartilage were outlined by a combination of computer and manual methods, and the images were used to calculate the volume of each cartilage plate. Comparing the coefficient of variance between the volume measurements made from the two visits, the volume measurements made from images with the highest resolution (0.275 x 0.275 x 1.0 mm) had the highest reproducibility. The high resolution images of the tibia and femur had the least partial-volume averaging and, as a result, better defined the boundaries between cartilage and adjacent tissues. A different trend was evident for the patella. For studies of osteoarthritis therapies, we recommend using MR images with the highest possible in-plane spatial resolution to provide the most reproducible volume measurements of knee cartilage. PMID- 11121701 TI - Bone marrow edema in the greater tuberosity of the humerus at MR imaging: association with rotator cuff tears and traumatic injury. AB - The purpose of this study was to determine the prevalence of bone marrow edema in the greater tuberosity of the humerus on MR imaging, the association with other findings at MR imaging and the injury mechanism which can lead to this finding. SUBJECTS AND METHODS: MR reports from 863 patients referred for shoulder MRI over 74 months were reviewed to identify patients with marrow edema in the greater tuberosity. The MR images from patients with greater tuberosity marrow edema were reviewed by consensus of two radiologists for the extent of marrow edema and for associated injuries. Marrow edema in the greater tuberosity was seen in 11 of 863 patients (1.3%). Nine patients (82%) had associated rotator cuff tear by MR imaging (four full thickness and five partial thickness), one patient had avulsion of the greater tuberosity from the humerus, and one had no rotator cuff abnormality. History of trauma was reported by eight patients including fall without direct blow to the shoulder (6), car accident (1) and direct blow to the top of the shoulder (1). Marrow edema in the greater tuberosity is an infrequent finding. Marrow edema most often is associated with a history of trauma and with rotator cuff abnormalities including full thickness tears. The history of trauma without direct blow to the shoulder and the location of the edema indicates that marrow edema often results from avulsion injury by the supraspinatus tendon. PMID- 11121702 TI - Quantitative human in vivo evaluation of high resolution MRI for vessel wall morphometry after percutaneous transluminal angioplasty. AB - Visualization of the vessel wall after transluminal angioplasty is important to monitor the restenosis progress. Intravascular ultrasound proved its capabilities as an invasive procedure in many studies. The aim of this study was to evaluate the feasibility of high-resolution MRI as a non-invasive tool for follow-up after PTA. High-resolution magnetic resonance images (pixel size: 0.49 * 0.49 mm(2)) were acquired on a 1.0 T clinical scanner. Morphometry was conducted after conversion of DICOM images into TIFF format using ScionImage on a PC. In-vitro studies using a polyvinylchloride tube were evaluated by two independent investigators. Goldstandard was a caliper rule and direct radiography. Five patients were monitored before and 24 h, six weeks, three months and six months after PTA. In vivo measurements promised a good concordance for both investigators for area as well as for diameter measurements. Area measurements showed correlations up to r = 0.86 (p < 0.001) whereas the correlations of diameters were slightly inferior (r between 0.58 and 0.84; p < 0.005). Relocation of the same slice position in the follow up studies could be guaranteed using anatomic landmarks in the images. As a non-invasive tool to assess restenosis after PTA high-resolution MRI promises to be a reproducible technique. It is easy to identify the same vascular region in different studies due to neighboring anatomic landmarks. Progression of disease as well as success of pharmacologic treatment to prevent restenosis may be monitored. PMID- 11121703 TI - The mechanical state of intracranial tissues in elderly subjects studied by imaging CSF and brain pulsations. AB - The biomechanical properties of intracranial tissues influence the mechanical coupling of brain and CSF oscillations to the driving vascular pulsations. Dynamic phase contrast MRI was used to measure the transfer functions that characterize these couplings in normal elderly subjects and patients with Alzheimer's disease. The transfer functions of both groups were significantly different from the previously reported transfer functions of normal young subjects. The data show that vascular pulsations tend to cause greater spinal cord movements and smaller CSF oscillations in the older subjects than in the younger ones. These results are likely to be due to age-related changes in the mechanical state of intracranial tissues. PMID- 11121704 TI - Intratumor heterogeneity in perfusion in human melanoma xenografts measured by contrast-enhanced magnetic resonance imaging. AB - The perfusion in tumors shows substantial spatial heterogeneity compared to that in normal tissues. The aim of the present study was to evaluate the intratumor heterogeneity in perfusion in tumors of two amelanotic human melanoma xenograft lines, A-07 and R-18, grown intradermally in Balb/c nu/nu mice. A non-invasive contrast-enhanced magnetic resonance imaging method yielding results in absolute values was applied. The perfusion was determined in manually defined regions of interest, corresponding to a whole tumor or to subregions of a tumor. The mean perfusion and the intertumor heterogeneity in perfusion were similar for the two tumor lines. For whole A-07 tumors, the perfusion ranged from 0.089 mL/(g . min) to 0.20 mL/(g . min) [mean: 0.15 mL/(g . min)], and for whole R-18 tumors, from 0.030 mL/(g . min) to 0.17 mL/(g . min) [mean: 0.13 mL/(g . min)]. The intratumor heterogeneity, on the other hand, was estimated to be 6.4 times larger in A-07 tumors than in R-18 tumors. The highest perfusion values, up to 0.69 mL/(g . min), were found in subregions of A-07 tumors. The intratumor heterogeneity was substantially larger than the intertumor heterogeneity in A-07 tumors, whereas in R-18 tumors, the intratumor heterogeneity was similar to the intertumor heterogeneity. These observations imply that measurements of mean tumor perfusion may have limited value as a predictive assay for outcome of treatment. PMID- 11121705 TI - Time-course magnetic resonance imaging of rat pancreatic cyst after experimental pancreatitis. AB - Fast magnetic resonance (MR) imaging of the rat pancreas was carried out using a snapshot method to observe three-dimensional (3D) and temporal development of the pancreatic cyst after experimental pancreatitis. Acute pancreatitis was induced by a retrograde infusion of the trypsin-taurocholate solution into the pancreatic duct in 23 rats, of which seven survived for one month. Under 2% enflurane anesthesia, (1)H images of the rat abdomen were taken by a 4.7 T magnetic resonance spectrometer under spontaneous breathing. 3D images of the pancreas and cyst were reconstructed from the axial, sagittal and coronal images taken before, 24 h, 7 days, 14 days, 21 days and 28 days after the induction of pancreatitis. The 3D images reconstructed from different slice orientations at each time point showed good agreement with each other. The calculated volumes of the cyst on 7th, 14th, 21st, and 28th day were 0.3 +/- 0.1, 0.8 +/- 0.3, 2.1 +/- 0.6, 6.5 +/- 1.3 mL, respectively. The cystic fluid volume on 28th day was 6.4 +/- 1.4 mL, which confirmed reliability of volume measurement by MR imaging. Fast MR imaging (snapshot) together with 3D reconstruction allows us to understand the detailed chronological and spatial development of pancreatic cyst after acute pancreatitis in rats. PMID- 11121706 TI - Venodilatory effect of pranidipine, a calcium channel blocker, monitored with perfluorocarbon in vivo (19)F-NMR spectroscopy. AB - We previously reported that the (19)F-NMR signal intensity of perfluorocarbon was increased by a venodilator, nitroglycerine, and decreased by an arteriodilator, hydralazine. In the present study, we demonstrated that pranidipine, a calcium channel blocker, increased the intensity of the FC-43 signal, while nifedipine, a prototype of calcium channel blockers, did not. These results suggest that pranidipine dilates the venous system in contrast to nifedipine. PMID- 11121707 TI - Accuracy and validity of stereology as a quantitative method for assessment of human temporal lobe volumes acquired by magnetic resonance imaging. AB - The object of this study was to compare the accuracy and validity of stereology as a method for determining whole temporal lobe volume with the more established technique of semi-automated thresholding and tracing. Ten, fixed, post-mortem human brains, were imaged using a three dimensional (3D) acquisition protocol. The volume of the left temporal lobe, dissected from each brain, was determined by fluid displacement. Each volume was compared to measurements obtained from magnetic resonance images (MRI) of the post-mortem brain using each of the two segmentation methods. Post-acquisition processing was performed using MEASURE software. Three investigators performed each measurement three times using each method, yielding a total of 180 measurements. Stereology took, on average, half the time of thresholding/tracing. Using a clinically acceptable variation for 95% of repeat measures; both intra-observer and inter-observer variation were acceptable for each technique. However, validity, as demonstrated by graphs of agreement against water displacement showed that the "limits of agreement" using stereology were within the acceptable range, while those using the thresholding/tracing technique were not. Quantitative estimates of variation and a graphical representation of the limits of agreement show that stereology is at least as precise as the thresholding/tracing method but is superior in terms of speed and validity. This has broad implications for published estimates of brain region volumes in human diseases such as epilepsy, dementia and other neurodegenerative disorders. PMID- 11121708 TI - Hippocampal and cerebellar volumetry in serially acquired MRI volume scans. AB - In this work, we describe methodologies for serial volumetric measurements of hippocampi and cerebella. Serial scans were co-registered and intensity matched prior to the volumetric measurements. Manual drawing was used to define the boundaries of the hippocampi. For the cerebellar volumetric measurements, the brain was automatically segmented from the co-registered scans; manual drawing was used to define the boundary between the cerebellum and the cerebrum and brainstem. The operator was blinded to the nature of the subject (patient or normal control) and the chronological order of the scans. The coefficient of reliability of hippocampal volume measurements in a group of 20 controls was 0.078 cm(3) (3.1% of the mean baseline volume); for the cerebellum, the value was 3.8 cm(3) (3.0% of the mean baseline volume). We conclude that the methods presented are valid and that the software provides a useful integrated tool for the quantitative analysis of structural changes in serially acquired volume MRI data in prospective, blinded studies. PMID- 11121709 TI - Hepatic ischemia secondary to hepatic artery ligation: MRI findings. PMID- 11121710 TI - Magnetic resonance imaging appearance of metastatic Merkel cell carcinoma to the sacrum and epidural space. AB - Merkel cell carcinoma (MCC) is a rare malignant tumor of the skin and often is diagnosed histologically as lymphoma, melanoma and even metastatic small cell carcinoma of the lung (SCCL). Classified as a neuroendocrine tumor, clinically it originates in the head and neck region and may present with metastatic disease at the time of presentation [1]. Osseous involvement in the past has been described to involve regional facial bones only. We present the first reported MRI findings of distant osseous metastasis from a Merkel cell carcinoma to the lumbosacral spine with associated soft tissue and epidural involvement. Appropriate treatment and patient survival depend on prompt diagnostic imaging for establishment of metastatic disease. Previous reports have advocated CT for diagnosis and staging of distant metastases [2,3]. When spinal involvement is suspected, MRI may be a more suitable modality for assessment of the epidural space and appropriate staging and follow-up in such cases. PMID- 11121711 TI - Oxidative stress status-the fifth set PMID- 11121712 TI - Determination of salicylate hydroxylation products as an in vivo oxidative stress marker. AB - The in vivo measurement of highly reactive free radicals, such as the z.rad OH radical, is very difficult. New specific markers, which are based on the ability of z.rad OH to attack the benzene rings of aromatic molecules, are currently under investigation. The produced hydroxylated compounds can be measured directly. In vivo, radical metabolism of salicylic acid produces two main hydroxylated derivatives (2,3- and 2,5-dihydroxybenzoic acids). The latter acid can be also produced by enzymatic pathways through the cytochrome P-450 system, while the former acid is reported to be solely formed by direct hydroxyl radical attack. Therefore, measurement of 2, 3-DHBA, following oral administration of the drug acetyl salicylate, could be proposed for assessment of oxidative stress in vivo. In this paper, a sensitive method for the identification and quantification of hydroxylation products from the reaction of z. rad OH with salicylate in vivo is presented. It employs a high performance liquid chromatography and electrochemical detection system. A detection limit of < 1 pmol for the hydroxylation products has been achieved with linear response over at least five orders of magnitude. Using this technique, we measured plasma levels of 2,3- and 2,5-DHBA dihydroxylated derivatives and salicylic acid and determined the ratios following administration of 1 g acetyl salicylate in 20 healthy subjects. PMID- 11121714 TI - Accurate and sensitive measurements of pO(2) in vivo using low frequency EPR spectroscopy: how to confer biocompatibility to the oxygen sensors. AB - Within the last few years, there has been a significant amount of progress using EPR oximetry, which has resulted in the availability of instrumentation and paramagnetic materials capable of measuring pO(2) in tissues with an accuracy and sensitivity comparable to or greater than that available by any other method. While the results obtained with EPR so far indicate that criteria for the measurements of pO(2)-such as accuracy, sensitivity, repeatability, and noninvasiveness-can be met, some of the paramagnetic materials with optimum spectroscopic properties (i.e., strong simple signals which are appropriately responsive to changes in pO(2)) may have some undesirable interactions with tissues, causing reactions with and/or losing responsiveness to oxygen. In this paper, several approaches are discussed, such as encapsulation procedures, which can result in the availability of oxygen-sensitive materials in a suitable configuration for long-term studies (absence of toxicity and preservation of the responsiveness to oxygen). PMID- 11121713 TI - Urinary aldehydes as indicators of lipid peroxidation in vivo. AB - The excretion of malondialdehyde (MDA), lipophilic aldehydes and related carbonyl compounds in rat and human urine was investigated. MDA was found to be excreted mainly in the form of two adducts with lysine, indicating that its predominant reaction in vivo is with the lysine residues of proteins. Adducts with the phospholipid bases serine and ethanolamine and the nucleic acid bases guanine and deoxyguanosine also were found. Except for the adduct with deoxyguanosine (dG MDA), the excretion of these compounds increased with peroxidative stress imposed in the form of vitamin E deficiency or the administration of iron or carbon tetrachloride. Marked differences in the concentration of dG-MDA in different tissues were correlated with their content of fatty acids having three or more double bonds, the putative source of MDA. Fourteen nonpolar and eleven polar lipophilic aldehydes and other carbonyl compounds were identified as their 2,4 diphenylhydrazine derivatives in rat urine. The excretion of five nonpolar and nine polar compounds was increased under conditions of peroxidative stress. The profile of lipophilic aldehydes obtained for human urine resembled that for rat urine. Except for a reported 4-hydroxynon-2-enal conjugate with mercapturic acid, the conjugated forms of the lipophilic aldehydes excreted in urine remain unidentified. Aldehyde excretion is influenced by numerous factors that affect the formation of lipid peroxides in vivo such as energy status, physical activity and environmental temperature, as well as by wide variations in the intake of peroxides in the diet. Consequently, urinalysis for aldehydic products of lipid peroxidation is an unreliable indicator of the general state of peroxidative stress in vivo. PMID- 11121715 TI - LC-MS/MS detection of peroxynitrite-derived 3-nitrotyrosine in rat microvessels. AB - 3-Nitrotyrosine (3NT) is used as a biomarker of nitrative pathology caused by peroxynitrite (PN), myeloperoxidase (MPO)-, and/or eosinophil peroxidase (EPO) dependent nitrite oxidation. 3NT measurements in biological materials are usually based on either antibody staining, HPLC detection, or GC detection methodologies. In this report, a procedure is described for the measurement of 3NT and tyrosine (TYR) by LC-MS/MS that is simple, direct, and sensitive. Though highly specialized in its use as an assay, LC-MS/MS technology is available in many research centers in academia and industry. The critical assay for 3NT was linear below 100 ng/ml and the limit of detection was below 100 pg/ml. Regarding protein digested samples, we found that MRM was most selective with 133.1 m/z as the daughter ion. In comparison, LC-ECD was 100 times less sensitive. Basal levels of 3NT in extracted digests of rat brain homogenate were easily detected by LC MS/MS, but were below detection by LC-ECD. The LC-MS/MS assay was used to detect 3NT in rat brain homogenate that was filtered through a 180 micron nylon mesh. Three fractions were collected and examined by phase contrast microscopy. The mass ratio (3NT/TYR) of 3NT in fractions of large vessel enrichment, microvessel enrichment, and vessel depletion was 0.6 ng/mg, 1.2 ng/mg, and 0.2 ng/mg, respectively. Ultimately, we found that the basal 3NT/TYR mass ratio as determined by LC-MS/MS was six times greater in microvessel-enriched brain tissue vs. tissue devoid of microvessels. PMID- 11121716 TI - Histochemical visualization of oxidant stress. AB - Free radicals induce oxidative modification in distinct components of the living matter (lipid, proteins, and DNA). For qualitative and quantitative determination of free radical-induced modifications, different, more or less sensitive biochemical methods are available. Because of the high reactivity and short life of free radicals, ongoing oxidative damage is generally analyzed by measurement of secondary products-such as H(2)O(2), oxidized proteins, peroxidized lipids, and their breakdown products, oxidized DNA-or by fluorographic analysis in combination with fluorescent dyes such as dichlorofluorescin (DCFH). In addition, the determination of free radical-related oxidation products is usually carried out in plasma, urine, or, less frequently, in bioptic material. Consequently, biochemical data seldom reflect the effects of free radical insults in situ. The histochemical visualization of selected molecular markers of oxidative damage can often provide more valuable information concerning the in vivo distribution of oxidative processes. This review summarizes the methods currently available for histochemical detection and indirect visualization of free radical-induced alterations in tissues and isolated cells. PMID- 11121717 TI - Total antioxidant capacity as a tool to assess redox status: critical view and experimental data. AB - The measure of antioxidant capacity (AC) considers the cumulative action of all the antioxidants present in plasma and body fluids, thus providing an integrated parameter rather than the simple sum of measurable antioxidants. The capacity of known and unknown antioxidants and their synergistic interaction is therefore assessed, thus giving an insight into the delicate balance in vivo between oxidants and antioxidants. Measuring plasma AC may help in the evaluation of physiological, environmental, and nutritional factors of the redox status in humans. Determining plasma AC may help to identify conditions affecting oxidative status in vivo (e.g., exposure to reactive oxygen species and antioxidant supplementation). Moreover, changes in the plasma AC after supplementation with galenic antioxidants or with antioxidant-rich foods may provide information on the absorption and bioavailability of nutritional compounds. Consequently, this review discusses the rationale, interpretation, confounding factors, measurement limits, and human applications of the measure of plasma AC. PMID- 11121718 TI - Oxidative damage in pregnant diabetic rats and their embryos. AB - Free radical mechanisms may be involved in the teratogenesis of diabetes. The contribution of oxidative stress in diabetic complications was investigated from the standpoint of oxidative damage to DNA, lipids, and proteins in the livers and embryos of pregnant diabetic rats. Diabetes was induced prior to pregnancy by the administration of streptozotocin (45 mg/kg). Two groups of diabetic rats were studied, one without any supplementation (D) and another treated during pregnancy with vitamin E (150 mg/d by gavage) (D + E). A control group was also included (C). The percentage of malformations in D rats were 44%, higher than the values observed in C (7%) and D + E (12%) animals. D Group rats showed a higher concentration of thiobarbituric acid reactive substances in the mother's liver, however, treatment with vitamin E decreased this by 58%. The levels of protein carbonyls in the liver of C, D, and D + E groups were similar. The "total levels" of the DNA adducts measured, both in liver and embryos C groups were similar to the D groups. Treatment of D groups with vitamin E reduced the levels by 17% in the liver and by 25% in the embryos. In terms of the "total levels" of DNA adducts, the embryos in diabetic pregnancy appear to be under less oxidative stress when compared with the livers of their mothers. Graziewicz et al. (Free Radical Biology & Medicine, 28:75-83, 1999) suggested "that Fapyadenine is a toxic lesion that moderately arrests DNA synthesis depending on the neighboring nucleotide sequence and interactions with the active site of DNA polymerase." Thus the increased levels of Fapyadenine in the diabetic livers and embryos may similarly arrest DNA polymerase, and in the case of this occurring in the embryos, contribute to the congenital malformations. It is now critical to probe the molecular mechanisms of the oxidative stress-associated development of diabetic congenital malformations. PMID- 11121719 TI - Lipoic acid decreases lipid peroxidation and protein glycosylation and increases (Na(+) + K(+))- and Ca(++)-ATPase activities in high glucose-treated human erythrocytes. AB - Lipoic acid supplementation has been found to be beneficial in preventing neurovascular abnormalities in diabetic neuropathy. Insufficient (Na(+) + K(+)) ATPase activity has been suggested as a contributing factor in the development of diabetic neuropathy. This study was undertaken to test the hypothesis that lipoic acid reduces lipid peroxidation and glycosylation and can increase the (Na(+) + K(+))- and Ca(++)-ATPase activities in high glucose-exposed red blood cells (RBC). Washed normal human RBC were treated with normal (6 mM) and high glucose concentrations (45 mM) with 0-0.2 mM lipoic acid (mixture of S and R sterioisomers) in a shaking water bath at 37 degrees C for 24 h. There was a significant stimulation of glucose consumption by RBC in the presence of lipoic acid both in normal and high glucose-treated RBC. Lipoic acid significantly lowered the level of glycated hemoglobin (GHb) and lipid peroxidation in RBC exposed to high glucose concentrations. High glucose treatment significantly lowered the activities of (Na(+) + K(+))- and Ca(++)-ATPases of RBC membranes. Lipoic acid addition significantly blocked the reduction in activities of (Na(+) + K(+))- and Ca(++)-ATPases in high glucose- treated RBC. There were no differences in lipid peroxidation, GHb and (Na(+) + K(+))- and Ca(++)-ATPase activity levels in normal glucose-treated RBC with and without lipoic acid. Thus, lipoic acid can lower lipid peroxidation and protein glycosylation, and increase (Na(+) + K(+))- and Ca(++)-ATPase activities in high-glucose exposed RBC, which provides a potential mechanism by which lipoic acid may delay or inhibit the development of neuropathy in diabetes. PMID- 11121720 TI - Copper supplementation in humans does not affect the susceptibility of low density lipoprotein to in vitro induced oxidation (FOODCUE project). AB - The oxidative modification of low-density lipoprotein cholesterol (LDL) has been implicated in the pathogenesis of atherosclerosis. Copper (Cu) is essential for antioxidant enzymes in vivo and animal studies show that Cu deficiency is accompanied by increased atherogenesis and LDL susceptibility to oxidation. Nevertheless, Cu has been proposed as a pro-oxidant in vivo and is routinely used to induce lipid peroxidation in vitro. Given the dual role of Cu as an in vivo antioxidant and an in vitro pro-oxidant, a multicenter European study (FOODCUE) was instigated to provide data on the biological effects of increased dietary Cu. Four centers, Northern Ireland (coordinator), England, Denmark, and France, using different experimental protocols, examined the effect of Cu supplementation (3 or 6 mg/d) on top of normal Cu dietary intakes or Cu-controlled diets (0.7/1.6/6.0 mg/d), on Cu-mediated and peroxynitrite-initiated LDL oxidation in apparently healthy volunteers. Each center coordinated its own supplementation regimen and all samples were subsequently transported to Northern Ireland where lipid peroxidation analysis was completed. The results from all centers showed that dietary Cu supplementation had no effect on Cu- or peroxynitrite-induced LDL susceptibility to oxidation. These data show that high intakes (up to 6 mg Cu) for extended periods do not promote LDL susceptibility to in vitro-induced oxidation. PMID- 11121721 TI - Synergistic inhibition of cyclooxygenase-2 expression by vitamin E and aspirin. AB - The use of aspirin in rheumatoid arthritis is limited since inhibition of the pro inflammatory enzyme cyclooxygenase-2 occurs only at higher aspirin doses that are often associated with side effects such as gastric toxicity. Using a macrophage cell line (J774. 1A), the present study explores possible synergistic effects of aspirin and vitamin E on the expression and activity of cyclooxygenase-2. Lipopolysaccharide-induced prostaglandin E(2) formation was significantly reduced by aspirin (1-100 microM) or vitamin E (100-300 microM). When combined with vitamin E, aspirin-dependent inhibition of prostaglandin E(2) formation was increased from 59% to 95% of control. Likewise, lipopolysaccharide-induced cyclooxygenase-2 protein and mRNA expression were virtually abolished by the combined treatment of aspirin and vitamin E, whereas the two agents alone were only modestly effective. Vitamin C did not mimic the actions of vitamin E under these conditions, suggesting that redox-independent mechanisms underlie the action of vitamin E. In agreement with this, vitamin E and aspirin were without effect on lipopolysaccharide-induced translocation of the redox-sensitive transcription factor NF-kappa B. Our results show that co-administration of vitamin E renders cyclooxygenase-2 more sensitive to inhibition by aspirin by as yet unknown mechanisms. Thus, anti-inflammatory therapy might be successful with lower aspirin doses when combined with vitamin E, thereby possibly avoiding the side effects of the usually required high dose aspirin treatment. PMID- 11121722 TI - Prevention of flight activity prolongs the life span of the housefly, Musca domestica, and attenuates the age-associated oxidative damamge to specific mitochondrial proteins. AB - The purpose of this study was to explore the mechanisms by which oxidative stress affects the aging process. The hypothesis that the rate of accumulation of oxidative damage to specific mitochondrial proteins is linked to the life expectancy of animals was tested in the housefly. The rate of oxygen consumption and life expectancy of the flies were experimentally altered by confining the flies in small jars, where they were unable to fly. Prevention of flight activity decreased the rate of oxygen utilization of flies and almost tripled their life span as compared to those permitted to fly. Rate of mitochondrial H(2)O(2) generation at various ages was lower in the low activity flies than in the high activity flies. Oxidative damage to mitochondrial proteins, adenine nucelotide translocase, and aconitase, detected as carbonyl modifications, was attenuated; and the loss in their functional activity occurring with age was retarded in the long-lived low activity flies as compared to the short-lived high activity flies. The two proteins were previously identified to be the only mitochondrial proteins exhibiting age-related increases in carbonylation. Results support the hypothesis that accrual of oxidative damage to specific protein targets and the consequent loss of their function may constitute a mechanism by which oxidative stress controls the aging process. PMID- 11121723 TI - Dependence of plasma alpha-tocopherol flux on very low-density triglyceride clearance in humans. AB - To evaluate the effect of dietary fat-induced alterations in triglyceride (TG) metabolism on plasma and very low-density lipoprotein (VLDL)-alpha-tocopherol, nine healthy males (mean +/- SEM, age: 36 +/- 3 years, BMI: 24.7 +/- 1.1) consumed a 35%-fat diet (control) for one week followed by a 15% low-fat, high carbohydrate diet for 5 weeks. After each dietary phase, the subjects ingested an evening meal along with a 50 mg capsule of (2)H(6)-RRR-alpha-tocopheryl acetate; blood samples were drawn over a 24 h period while the subjects remained fasted. Low-fat feeding increased fasting plasma TG concentrations by 53% (116 +/- 27 to 178 +/- 32, mg/dl, p < 0.0001) primarily by reducing VLDL-TG clearance. Total plasma alpha-tocopherol concentrations (labeled + unlabeled) were unchanged (25.8 +/- 2.3 vs. 26.4 +/- 3.0 nmol/ml plasma) and no differences between the diets were observed for plasma (2)H(6)-alpha-tocopherol concentration (4.8 +/- 0.6 nmol/ml, for both diets) or enrichments (18.1 +/- 1.8% average for both diets). However, low-fat feeding significantly increased the amount of alpha-tocopherol in the VLDL fraction (43%, p = 0.04) in concert with elevations in VLDL-apoB and TG. The alpha-tocopherol and TG content of VLDL varied in parallel in individual subjects and fractional replacement rates and clearance of alpha-tocopherol and TG in VLDL were closely correlated. Kinetic parameters were decreased by 32-39% from high-fat to low-fat. These data suggest that vitamin E bioavailability is similar between a 15 and 35% fat diet, with a redistribution of alpha-tocopherol in lipoproteins occurring during low-fat feeding (increased in the VLDL fraction, reduced in the other lipoproteins), and transfer of alpha-tocopherol from VLDL depends upon TG removal from the particle, consistent with previous observations in vitro and in animal studies. PMID- 11121724 TI - Evidence of a defective thiol status of alveolar macrophages from COPD patients and smokers. Chronic obstructive pulmonary disease. AB - In increasing numbers of pulmonary diseases an association with a loss of intracellular thiols, mainly glutathione, is postulated. Therefore, the quantitative measurement of thiols within different viable cells is a possible metabolic parameter for cellular function and defense capacity of all pulmonary immune cells including alveolar macrophages (AM), that are highly compromised by oxidative stress. In this study the cellular thiol content was determined using fluorochrom conjugated chloromethyl derivatives (5-chloromethylfluorescein diacetate, CMFDA) in flow cytometry. The procedure was evaluated in vitro using biochemical techniques for glutathione quantification. Based on this approach, AM obtained from bronchoalveolar lavage (BAL) of smokers and patients with chronic obstructive pulmonary disease (COPD) showed a significant thiol deficiency compared to a nonsmoker/non-COPD group. The cellular thiol expression of AM from smokers and COPD patients reached only 50 and 53% of the control group. Lowest thiol concentrations (47% of control) were detected within the smoker(+)/COPD(+) group. This intracellular thiol deficiency significantly correlated with reduced lung function (FEV(1), PaO(2)). With regard to the tightly regulated thiol metabolism of immune cells, these results imply the onset of functional disturbances in thiol deficient AM. The determination of the cellular thiol content of AM, obtained from BAL by flow cytometry, presents a simple and reliable tool to monitor the effect of therapeutic measures focusing on the stabilization of the cellular thiol status. PMID- 11121725 TI - Enhanced sensitivity and long-term G2 arrest in hydrogen peroxide-treated Ku80 null cells are unrelated to DNA repair defects. AB - While the Ku complex, comprised of Ku70 and Ku80, is primarily involved in the repair of DNA double-strand breaks, it is also believed to participate in additional cellular processes. Here, treatment of embryo fibroblasts (MEFs) derived from either wild-type or Ku80-null (Ku80(-/-)) mice with various stress agents revealed that hydrogen peroxide (H(2)O(2)) was markedly more cytotoxic for Ku80(-/-) MEFs and led to their long-term accumulation in the G2 phase. This differential response was not due to differences in DNA repair, since H(2)O(2) triggered DNA damage was repaired with comparable efficiency in both Wt and Ku80( /-) MEFs, but was associated with differences in the expression of important cell cycle regulatory genes. Our results support the notion that Ku80-mediated cytoprotection and G2-progression are not only dependent on the cell's DNA repair but also may reflect Ku80's influence on additional cellular processes such as gene expression. PMID- 11121727 TI - Changes of CYP1A1, GST, and ALDH3 enzymes in hepatoma cell lines undergoing enhanced lipid peroxidation. AB - Hepatoma cells show alterations in the response to oxidative stress (decreased lipid peroxidation) and in xenobiotic metabolism enzymes (decreased P450, increased GST and ALDH3). This study examined the effect of lipid peroxidation on the expression of the above enzymes in two rat hepatoma cell lines (MH(1)C(1) and 7777). To induce oxidative stress, cells were exposed to arachidonic acid (to increase lipid peroxidation substrate) and/or to beta-naphthoflavone (to increase CYP450), and treated with one dose of iron/histidine. The cells, that were still viable after the challenge, were refed with the culture medium and CYP1A1, GST, and ALDH3 enzymes monitored for 1, 6, 12, and 24 h. Treatments that increased markers indicative of lipid peroxidation are associated with a decrease in enzyme activities, which was permanent for CYP1A1 and transient for the other enzymes. We speculate from these data that aldehydic byproducts of lipid peroxidation may be responsible for these effects. Thus, restoration of lipid peroxidation in hepatoma cells seems to induce a rapid adaptation to oxidative stress, which is achieved by a simultaneous decrease of reactive oxygen species production and an increase in the two main enzymes involved in the removal of the aldehydic products of lipid peroxidation. PMID- 11121726 TI - Melatonin directly scavenges hydrogen peroxide: a potentially new metabolic pathway of melatonin biotransformation. AB - A potential new metabolic pathway of melatonin biotransformation is described in this investigation. Melatonin was found to directly scavenge hydrogen peroxide (H(2)O(2)) to form N(1)-acetyl-N(2)-formyl-5-methoxykynuramine and, thereafter this compound could be enzymatically converted to N(1)-acetyl-5-methoxykynuramine by catalase. The structures of these kynuramines were identified using proton nuclear magnetic resonance, carbon nuclear magnetic resonance, and mass spectrometry. This is the first report to reveal a possible physiological association between melatonin, H(2)O(2), catalase, and kynuramines. Melatonin scavenges H(2)O(2) in a concentration-dependent manner. This reaction appears to exhibit two distinguishable phases. In the rapid reaction phase, the interaction between melatonin and H(2)O(2) reaches equilibrium rapidly (within 5 s). The rate constant for this phase was calculated to be 2.3 x 10(6) M(-1)s(-1). Thereafter, the relative equilibrium of melatonin and H(2)O(2) was sustained for roughly 1 h, at which time the content of H(2)O(2) decreased gradually over a several hour period, identified as the slow reaction phase. These observations suggest that melatonin, a ubiquitously distributed small nonenzymatic molecule, might serve to directly detoxify H(2)O(2) in living organisms. H(2)O(2) and melatonin are present in all subcellular compartments; thus, presumably, one important function of melatonin may be complementary in function to catalase and glutathione peroxidase in keeping intracellular H(2)O(2) concentrations at steady-state levels. PMID- 11121728 TI - Role of human CYP3A and P-glycoprotein on the absorption of drugs. PMID- 11121729 TI - Role of metabolic enzymes and efflux transporters in the absorption of drugs from the small intestine. AB - It has been established that the absorption of many drugs from the small intestine is hindered by the detoxification systems which are present in this epithelial tissue. In this article, we will summarize the significant role of small intestine in reducing the oral bioavailability of drugs, particularly focusing on the role of metabolic enzymes and efflux transporters. Since the role of cytochrome P450 3A (CYP3A) and MDR1 P-glycoprotein (P-gp) in intestinal drug disposition has been highlighted, the disposition of CYP3A substrates, P-gp substrates and CYP3A/P-gp bisubstrates are summarized. Moreover, it is plausible that conjugative enzymes and/or carboxyesterases act synergistically with efflux transporters of organic anions, affecting the intestinal availability, i.e. many xenobiotics and ester-type prodrugs are metabolized to the corresponding glucuronide and sulfate conjugates and carboxylates (active drugs), respectively, followed by cellular extrusion. The characteristics of the efflux transporters of organic anions across the apical and basal membrane of enterocytes and Caco-2 cells are also summarized from this point of view. PMID- 11121730 TI - Function and regulation of ATP-binding cassette transport proteins involved in hepatobiliary transport. AB - Hepatobiliary transport of endogenous and exogenous compounds is mediated by the coordinated action of multiple transport systems present at the sinusoidal (basolateral) and canalicular (apical) membrane domains of hepatocytes. During the last few years many of these transporters have been cloned and functionally characterized. In addition, the molecular bases of several forms of cholestatic liver disease have been defined. Combined, this has greatly expanded our understanding of the normal physiology of bile formation, the pathophysiology of intrahepatic cholestasis, as well as of drug elimination and disposition processes. In this review recent advances, with respect to function and regulation of ATP binding cassette transport proteins expressed in liver, are summarized and discussed. PMID- 11121731 TI - Structure-activity relationship of P-glycoprotein substrates and modifiers. AB - The air-water partition coefficients, K(aw), highly correlated with the corresponding lipid-water partition coefficients, K(lw), and the critical micelle concentrations, CMC, were measured for 11 compounds for which the kinetic parameters of P-glycoprotein ATPase activation (Michaelis-Menten constant, K(m), and maximal velocity, V(max)) had been determined previously in inside-out vesicles of CR1R12 Chinese hamster ovary cells. In addition, the hydrogen bond donor patterns (type I and type II) relevant for substrate recognition by P glycoprotein were determined from the energy-minimized three-dimensional structure of these compounds. A linear relation between the air-water partition coefficient, K(aw), and the inverse of the Michaelis-Menten constant, K(m), was observed such that K(m) x K(aw) approximately = 1. The maximal velocity, V(max), was shown to decrease with the number and strength of electron donor (hydrogen bond acceptor) groups in recognition patterns. If two substrates are applied simultaneously to P-glycoprotein the compound with the higher potential to form hydrogen bonds generally acts as an inhibitor. We conclude that partitioning into the lipid membrane is the rate-limiting step for the interaction of a substrate with P-glycoprotein and that dissociation of the P-glycoprotein-substrate complex is determined by the number and strength of the hydrogen bonds formed between the substrate and the transporter. PMID- 11121732 TI - The role of P-glycoprotein in determining the oral absorption and clearance of the NK2 antagonist, UK-224,671. AB - UK-224,671 has been shown to exhibit low oral bioavailability in vivo due to poor absorption from the GI tract. The purpose of this study was to investigate the underlying reason for this observation. In Caco-2 cell flux experiments, the absorptive (A to B) flux of UK-224,671 was low, consistent with poor in vivo absorption. However, flux in the B to A direction was significantly greater, suggesting that UK-224,671 can permeate the membrane of the gut wall cell. Such a Caco-2 cell flux is indicative of transporter mediated efflux, possibly by P glycoprotein. In P-glycoprotein knockout mice, the oral bioavailability of UK 224,671 was 22%, representing a significant increase over the P-glycoprotein expressing wild type mice (<2%). However, in the knockout mice absorption was still incomplete, suggesting that both P-glycoprotein mediated efflux and poor membrane permeation combine to limit the oral absorption of UK-224,671 in wild type mice. Lack of P-glycoprotein expression had no effect on the clearance of UK 224,671 in mice, which suggests that uptake from the blood into the excretory cell is mediated by a transporter other than P-glycoprotein. Bile duct cannulated rat experiments show that approximately 20% of the clearance of UK-224,671 occurs by direct secretion across the gut wall into the faeces. This clearance pathway requires UK-224,671 to cross both the basolateral and apical membranes of the gut wall cell. P-glycoprotein is likely to be involved in the passage of the compound across the apical membrane as has been observed for other P-glycoprotein substrates. PMID- 11121733 TI - Can oral midazolam predict oral cyclosporine disposition? AB - Effective cyclosporine therapy is confounded by large interindividual differences in oral bioavailability and a narrow therapeutic window. Because cytochrome P450 (CYP) 3A-mediated first-pass metabolism contributes to this unpredictable bioavailability, an in vivo oral CYP3A phenotyping probe could be a valuable tool in optimizing cyclosporine therapy. Based on similarities in the metabolic kinetics of cyclosporine and midazolam by the liver and intestinal mucosa, we evaluated whether midazolam oral clearance would predict cyclosporine oral clearance when the two drugs were administered to 20 medically stable kidney transplant recipients. Despite earlier findings in liver transplant recipients who displayed a strong correlation between the systemic clearances of midazolam and cyclosporine, there was a weak correlation between their oral clearances in the current group of subjects (r(s)=0.50, P=0.03). Differing extents of intestinal first-pass metabolic extraction between the two drugs, inhibition of midazolam metabolism by cyclosporine at the level of the intestine, and/or P glycoprotein-mediated intestinal efflux of cyclosporine (but not midazolam) may account for this poor correlation. We conclude that although oral midazolam is unlikely to be clinically useful as a probe for cyclosporine disposition, its utility in the prediction of other orally administered CYP3A substrates cannot be out ruled. PMID- 11121734 TI - Analysis of drug transport and metabolism in cell monolayer systems that have been modified by cytochrome P4503A4 cDNA-expression. AB - Human CYP3A4, the major human, intestinal, drug metabolizing cytochrome P450, has been introduced into three mammalian cell lines (Caco-2, MDCK and LLC-PK1) suitable for making drug permeability measurements. The levels and stability of expression were analyzed by enzyme assays (testosterone 6beta-hydroxylase and nifedipine oxidase). Long term, stable CYP3A4 expression/cell growth rate was obtained in MDCK cells. In the LLC-PK1 system, shorter term, stable expression was achieved. However, in Caco-2 cells, derivatives with better properties than those previously reported could not be obtained. The highest level of CYP3A4 catalytic activity was obtained in LLC-PK1 cells. In this system, CYP3A4 activity levels appeared comparable to median level human intestinal microsomes. Metabolite formation and inhibition kinetics were examined in cell monolayers. Nifedipine was found to be extensively metabolized (19%) during passage across cell monolayers. In general, affinity related parameters (apparent Km and apparent Ki) were 1.5- to three-fold higher under conditions of flux through the monolayers relative to steady-state conditions. These systems should be useful for examining the role of intestinal CYP3A4 in first-pass metabolism and drug drug interactions. PMID- 11121735 TI - P-Glycoprotein in cell cultures: a combined approach to study expression, localisation, and functionality in the confocal microscope. AB - Madin Darby canine kidney (MDCK) cells transfected with the multidrug resistance mdr1 gene, MDR1-MDCK (Pastan et al., 1988, Proc. Natl. Acad. Sci. USA 85 4486 4470), were used in a combined approach to study expression, localisation and functionality of the P-glycoprotein (P-gp) membrane transporter in the same cell culture preparations. Cells were characterised with regard to their growth curve, transepithelial electrical resistance (TEER), and cytoarchitecture. Efflux of the P-gp substrate rhodamine123 (rho123) was monitored with confocal laser scanning microscopy (CLSM). The transfected cells grew in multilayers. After reaching confluence they exhibited a complete tight junction (TJ) network. P-gp was strongly expressed at the uppermost apical surface of the multilayer already after 4 days in culture. The lower cell layers were not clearly polarised. P-gp mediated transport could be followed by efflux of the fluorescent rho123 from the cells into the apical extracellular space. Verapamil, a P-gp inhibitor, significantly decreased efflux. For MDCK parent cells the rho123 assay was negative up to about day 20, and only at later times (day 25) low P-gp activity was detected. These results clearly show that despite the fact that the transfected cells form irregular layers, they provide a good model for screening of P-gp substrates and inhibitors. PMID- 11121737 TI - Membrane fusion in a nutshell PMID- 11121736 TI - Mitochondrial tRNA import: are there distinct mechanisms? AB - Sequence information from an increasing number of complete mitochondrial genomes indicates that a large number of evolutionary distinct organisms import nucleus encoded tRNAs. In the past five years, much research has been initiated on the features of imported tRNAs, the mechanism and the energetics of the process as well as on the components of the import machinery. In summary, these studies show that the import systems of different species exhibit some unique features, suggesting that more than one mechanism might exist to import tRNAs. PMID- 11121738 TI - Where do golgi residents reside? PMID- 11121739 TI - ATM-dependent cell-cycle arrest tamed in the test tube PMID- 11121741 TI - Hints for a cure in a muscular dystrophy variant PMID- 11121740 TI - A KID's game of tug o' war PMID- 11121742 TI - Connecting to the proteasome PMID- 11121743 TI - Pollen tube targeting and axon guidance: parallels in tip growth mechanisms. AB - The growth of pollen tubes to plant egg cells and the guidance of axons to neural synapses are classic examples of targeted cell growth. Despite the evolutionary time that separates animals and plants, axon and pollen tube guidance share remarkable mechanistic similarities. In both instances, extracellular cues are transduced by intracellular signal-transduction pathways that culminate in directed tip growth. Do the mechanistic similarities extend to the molecular level? Here, we address this question by a comprehensive review of the molecules and pathways involved in pollen tube targeting and axon guidance. The emerging scenario is that similar intracellular molecules are recruited to control tip growth, while different extracellular molecules mediate guidance through the distinct plant and animal extracellular matrices. PMID- 11121744 TI - Aggresomes, inclusion bodies and protein aggregation. AB - Intracellular and extracellular accumulation of aggregated protein are linked to many diseases, including ageing-related neurodegeneration and systemic amyloidosis. Cells avoid accumulating potentially toxic aggregates by mechanisms including the suppression of aggregate formation by molecular chaperones and the degradation of misfolded proteins by proteasomes. Once formed, aggregates tend to be refractory to proteolysis and to accumulate in inclusion bodies. This accumulation has been assumed to be a diffusion-limited process, but recent studies suggest that, in animal cells, aggregated proteins are specifically delivered to inclusion bodies by dynein-dependent retrograde transport on microtubules. This microtubule-dependent inclusion body is called an aggresome. PMID- 11121745 TI - Huntington's disease: the challenge for cell biologists. AB - Huntington's disease (HD) is one of eight inherited neurodegenerative diseases caused by expansions of (CAG)(n) tracts that encode polyglutamine segments in expressed proteins. Studies of pathogenic mechanisms for all these late-onset diseases suffer from a common drawback: experimental studies require massive acceleration of a process that, in affected humans, usually takes decades. But is the rapid-onset disease of transgenic mouse models and in cells the same as the slow-onset disease in humans? We review recent work on HD, noting several issues whose significance is likely to be crucial - but which are as yet unresolved. We discuss these in light of the distinction between disease-specific pathogenic mechanisms and artifacts of polyglutamine overexpression. We suggest that the initial stages of HD result from dysfunction rather than death, and we consider the potential discovery of compounds that might interfere with early pathogenic events. PMID- 11121746 TI - An abundance of tubulins. AB - Recent data have revealed that the tubulin superfamily of proteins is much larger than was thought previously. Six distinct families within the tubulin superfamily have been discovered and more might await discovery. alpha-, beta- and gamma tubulins are ubiquitous in eukaryotes. alpha- and beta-tubulins are the major components of microtubules, and gamma-tubulin plays a major role in the nucleation of microtubule assembly. delta- and epsilon-tubulins are widespread but not ubiquitous, and zeta-tubulin has been found so far only in kinetoplastid protozoa. delta-Tubulin has an important role in flagellar assembly in Chlamydomonas, but its role in other organisms is just beginning to be investigated, as are the functions of the recently discovered epsilon- and zeta tubulins. PMID- 11121747 TI - The molecular basis of T helper 1 and T helper 2 cell differentiation. AB - Cytokines such as interleukin 12 (IL-12) and IL-4 are dominant factors in driving the development of T helper 1 (Th1) and Th2 cells, respectively, through specific signalling pathways. In addition, it has been demonstrated more recently that T helper-cell-specific transcription factors exist that determine the commitment of Th1 and Th2 cells for the production of distinct profiles of cytokines. In addition to the expression of distinct cytokine genes and transcription factors, the molecular basis for commitment to a Th1 or Th2 phenotype can probably be explained by multiple mechanisms, including differential cytokine signalling, exclusive cytokine receptor expression, differential expression of transcription factors and/or differential chromatin remodelling of Th1- and Th2-specific genes. PMID- 11121748 TI - Pictures in cell biology. A functional marker for Drosophila chromosomes in vivo. PMID- 11121749 TI - How do exported proteins and antibiotics bypass the periplasm in Gram-negative bacterial cells? PMID- 11121750 TI - Obligate intracellular bacteria and antibiotic resistance. PMID- 11121751 TI - Ralstonia solanacearum--a plant pathogen in touch with its host. PMID- 11121752 TI - Virulence and sodium bioenergetics. PMID- 11121753 TI - Erratum. PMID- 11121754 TI - Patch clamping malaria-infected red blood cells. PMID- 11121755 TI - Microbial genomics. PMID- 11121756 TI - The enlightened secrets across the ocean. PMID- 11121757 TI - Candida albicans virulence-related genes: the search continues. PMID- 11121758 TI - Genome of an opportunist. PMID- 11121759 TI - Two-component systems in Pseudomonas aeruginosa: why so many? AB - Screening the Pseudomonas aeruginosa genome has led to the identification of the highest number of putative genes encoding two-component regulatory systems of all bacterial genomes sequenced to date (64 and 63 encoding response regulators and histidine kinases, respectively). Sixteen atypical kinases, among them 11 devoid of an Hpt domain, and three independent Hpt modules were retrieved. These data suggest that P. aeruginosa possesses complex control strategies with which to respond to environmental challenges. PMID- 11121761 TI - Mitochondrial DNA inheritance in Saccharomyces cerevisiae. AB - Respiratory metabolism depends on mitochondrial DNA, yet the mechanisms that ensure the inheritance of the mitochondrial genome are largely obscure. Recent studies with Saccharomyces cerevisiae suggest that distinct factors mediate the active segregation of mitochondrial DNA during mitotic growth. The identification of the proteins required for the maintenance of the mitochondrial genome provides clues to the mechanisms of, and molecular machinery involved in, mitochondrial DNA inheritance. PMID- 11121760 TI - The origins and ongoing evolution of viruses. AB - Genome analyses of double strand DNA tailed bacteriophages argue that they evolve by recombinational reassortment of genes and by the acquisition of novel genes as simple genetic elements termed morons. These processes suggest a model for early virus evolution, wherein viruses can be regarded less as having derived from cells and more as being partners in their mutual co-evolution. PMID- 11121762 TI - The role of the CFTR in susceptibility to Pseudomonas aeruginosa infections in cystic fibrosis. AB - Recent molecular and cellular studies have shed new light on the basis for the susceptibility of cystic fibrosis (CF) patients to Pseudomonas aeruginosa infection. Changes in airway liquid composition and/or viscosity, enhanced bacterial binding to mucin and epithelial cell receptors, increased innate inflammation owing to disruptions in lipid metabolism and a role for the CFTR protein in bacterial ingestion and clearance have all been postulated. The high P. aeruginosa infection rate in CF patients can potentially be explained by the specificity of the interaction between the CFTR and P. aeruginosa. PMID- 11121763 TI - Transposon-based approaches to identify essential bacterial genes. AB - Transposons are a powerful tool for identifying genes essential for bacterial viability. The availability of many bacterial genome sequences and the large number of genes of unknown function therein have inspired the generation of a variety of different approaches. These methods are described and their advantages and disadvantages are discussed. PMID- 11121764 TI - Sensor systems for lower limb functional electrical stimulation (FES) control. AB - Two sensor systems comprising clusters of accelerometers, magnetic sensors, a rate gyroscope, and a strain gauge were designed. For one system, the clusters were located at the belt and AFO. In the other system, the clusters were located at the AFO and the thigh. The maximum cluster size was 14 cm(3) and 75 g. The clusters of each sensor system were interconnected by a single flexible wire bus, which minimized the effects of cabling. The sensors detected five phases of normal gait to a resolution of 40 ms in an able bodied test. Using a threshold method, the sensor system repeatedly predicted an incipient knee buckle in a paraplegic individual by a minimum of 30 ms. One system detected knee flexion angle analytically to an accuracy of 3.2 degrees during sit to stand trials. The second system determined knee and hip flexion angle to an accuracy of 3.8 degrees during sit to stand trials through neural networks. The signal processing of the acquired sensor signals in each system was performed on a MC68332 microcomputer in conjunction with the data sampling, and suggested the possibility for each sensor system to be used in real time control of FES. PMID- 11121765 TI - A system for monitoring the response of uniaxial strain on cell seeded collagen gels. AB - The success of cell seeded constructs for the repair of collagenous tissues may be improved by the use of mechanical stimulation in vitro. A mechanical loading apparatus, termed the cell straining system, was developed according to a set of design criteria, to enable cell seeded constructs to be cyclically loaded in tension. A suitable cell seeded collagen gel model system was used to characterise the apparatus. These gels were subjected to a cyclic strain of 10% superimposed on two separate tare loads of 2 and 10 mN, while being maintained in cell culture conditions. The computer controlled apparatus was shown to be capable of monitoring the individual loads on six specimens simultaneously, to an accuracy of 0.02 mN. Results indicated a wide variability between individual specimens. Following cyclic loading, the cell seeded collagen gels exhibited an increase in structural stiffness compared with the unloaded controls. This novel and versatile apparatus will provide a means of enhancing structural and mechanical integrity of tissue engineered repair systems. PMID- 11121766 TI - Selection of biorthogonal filters for image compression of MR images using wavelet packets. AB - We present an analysis of different filter banks for the compression of magnetic resonance (MR) images of the human brain using wavelet packets based on biorthogonal filters. Initially, peak signal to noise ratio (PSNR) and normalized root mean square (RMS) error criteria are calculated for a series of images compressed with a 33:1 ratio, using filter banks based on biorthogonal wavelet packets. The results lead us to choose a few of these filter banks as optimal for image compression. One of these filters is employed to compress several images at four different compression ratios: 12.5:1, 25:1, 37.5:1 and 50:1. The quality of these images was evaluated by visual analysis by a group of seven experts who graded image quality on a 0-7 scale. Results show that using these filters, we can compress images to a rate of around 30:1 without introducing noticeable differences. Other applications for these filters are currently under study and include the compression/fusion of MR image stacks in order to obtain even better reductions in the amount of data needed to reconstruct complete MRI studies. PMID- 11121767 TI - EEG-based discrimination between imagination of left and right hand movements using Adaptive Gaussian Representation. AB - This article uses the Adaptive Gaussian Representation (AGR) for human electroencephalogram (EEG) feature extraction aiming the discrimination among mental tasks to be used in a brain computer interface (BCI). It does not focus on the AGR time-frequency representation, but rather on their projection coefficients. Ten volunteers were asked to imagine either right or left hand movement, according to a proper visual stimulus. The features of the resulting EEG signals were characterised by extracting AGR coefficients. Classification was carried out using a Multilayer perceptron (MLP) trained with the classical backpropagation algorithm. Overall results show that AGR coefficients representation is able to reveal a significant EEG discrimination between imagination of right and left hand movement with a mean classification performance of 91%+/-5.8% achieved for female subjects and 87%+/-5.0% achieved for male subjects. PMID- 11121768 TI - Anisotropic shear behavior of the annulus fibrosus: effect of harvest site and tissue prestrain. AB - We used a shear protocol to investigate the time-independent, anisotropic structural behavior of 30 cube and 56 sheet specimens of human annulus fibrosus from six non-degenerate lumbar spines. The specimen sides were aligned with the natural collagen lamellar architecture. For the cube specimens, simple shear was applied sequentially in three orthogonal directions defined by the cube axes. The sheets were also tested in simple shear, with the stress applied in either the anatomic axial or circumferential directions. With the sheet specimens we also investigated the contribution of annular collagen to the shear modulus by applying a tensile prestrain (0, 5 or 10 percent) perpendicular to the direction of applied shear stress. Our cube data indicated that the shear modulus was anisotropic, being 56.04+/-36.3 kPa in the plane of the lamellae and approximately half that in the orthogonal directions. The sheet specimens demonstrated that shear modulus increased progressively by a factor of between 3 and 5 from the inner to outer annulus. The ratio of the axial to circumferential shear modulus for the sheets increased from being near unity for the inner annulus to near 2 for the outer annulus. Finally, the addition of a 10 percent tensile prestrain increased the shear modulus by between 1.5 and 2.5 times for the middle and outer annulus. The shear anisotropy we report is consistent with prior anatomical observations of this tissue and appears to develop through separate contributions from the matrix, the collagen fibers, and collagen fiber interactions. PMID- 11121769 TI - A comparative FEM-study of tooth mobility using isotropic and anisotropic models of the periodontal ligament. Finite Element Method. AB - Orthodontic tooth movement is usually characterized by two centres: the centre of resistance and the centre of rotation. A literature survey shows that both centres vary to a significant extent in both clinical and computational experiments. This paper reports on studies upon five different hypothetical mechanical representations of the periodontal ligament (PDL) which plays the most significant role in tooth mobility. The first model considers the PDL as an isotropic and linear-elastic continuum without fibres; it also discusses some preliminary visco-elastic aspects. The next three models assume a nonlinear and anisotropic material composed of fibres only that are arranged in three different orientations, two hypothetical that have appeared previously in the literature and one more consistent with actual morphological data. The fifth model considers the PDL as an orthotropic material consisting of both a continuum and of fibres. Results were obtained by applying the Finite Element Method (FEM) on a maxillary central incisor. It was found that the isotropic linear-elastic PDL leads to occlusal positions of both centres in comparison with those obtained through the well-known Burstone's theoretical formula, while histological anisotropic fibres locate them apically and eccentrically. PMID- 11121770 TI - Non-invasive measurement of hepatic oxygenation by an oxygen electrode in human orthotopic liver transplantation. AB - Precise evaluation of graft reperfusion is difficult in clinical liver transplantation. The oxygen electrode (OE) is a novel technique to detect blood flow indirectly by measuring the quantity of oxygen which can diffuse from the hepatic tissue to the surface electrode. Application of the surface OE does not influence the liver blood flow or parenchymal perfusion. Adequate graft oxygenation is essential to the outcome of organ transplantation and has not previously been analysed intra-operatively in liver transplant recipients. The OE was applied to the surface of the graft intra-operatively in 22 human liver grafts after restoring portal vein and hepatic artery inflow. OE readings were compared with liver blood flow using an electromagnetic flowmeter (EMF). Intra operative haemodynamics and donor organ parameters known to influence graft function were correlated with the OE readings. There was a significant correlation (r=0.89; p<0.001, n=14) between tissue oxygenation using the OE and total liver blood flow measured by EMF. The tissue oxygenation measurements were reproducible with a coefficient of variation of 5%. The hepatic tissue oxygenation increased significantly from baseline following venous reperfusion of the graft (282+/-23 vs 3107+/-288 (+/-SE) nA, p<0.001). Hepatic arterial revascularisation resulted in a significant (p<0.001) increase of 41+/-7% in liver oxygen perfusion. There was significant negative correlation (r=0.80, p<0.001, n=22) between cold ischaemic time and graft tissue oxygenation. The OE provides a reliable, cheap and non-invasive method of monitoring liver graft oxygenation and perfusion during transplantation. PMID- 11121771 TI - Web alert. Growth and development. PMID- 11121772 TI - Growth and development: signals and their transduction. PMID- 11121773 TI - Regulation of sigma factor activity during Bacillus subtilis development. AB - Progression of Bacillus subtilis through a series of morphological changes is driven by a cascade of sigma (sigma) factors and results in formation of a spore. Recent work has provided new insights into the location and function of proteins that control sigma factor activity, and has suggested that multiple mechanisms allow one sigma factor to replace another in the cascade. PMID- 11121774 TI - Control of sporulation initiation in Bacillus subtilis. AB - Recent work has provided new insights into the mechanisms by which Bacillus subtilis responds to signals that reflect high population density and nutritional limitation, the mechanisms that regulate activation of the key transcription factor Spo0A, and the physical basis for critical aspects of the Spo0A phosphorelay. PMID- 11121775 TI - Signal transduction cascades regulating pseudohyphal differentiation of Saccharomyces cerevisiae. AB - In response to nitrogen limitation, diploid cells of the yeast Saccharomyces cerevisiae undergo a dimorphic transition to filamentous pseudohyphal growth. At least two signaling pathways regulate filamentation. One involves components of the MAP kinase cascade that also regulates mating of haploid cells. The second involves a nutrient-sensing G-protein-coupled receptor that signals via an unusual G(alpha) protein, cAMP and protein kinase A. Recent studies reveal crosstalk between these pathways during pseudohyphal growth. Related MAP kinase and cAMP pathways regulate filamentation and virulence of human and plant fungal pathogens, and represent novel targets for antifungal drug design. PMID- 11121776 TI - Pheromone response, mating and cell biology. AB - Saccharomyces cerevisiae responds to mating pheromones by activating a receptor-G protein-coupled mitogen-activated protein kinase (MAPK) cascade that is also used by other signaling pathways. The activation of the MAPK cascade may involve conformational changes through prebound receptor and heterotrimeric G-protein. G beta may then recruit Cdc42-bound MAPKKKK Ste20 to MAPKKK Ste11 through direct interactions with Ste20 and the Ste5 scaffold. Ste20 activates Ste11 by derepressing an autoinhibitory domain. An underlying nuclear shuttling machinery may be required for proper recruitment of Ste5 to G beta. Subsequent polarized growth is mediated by a similar mechanism involving Far1, which binds G beta in addition to Cdc24 and Bem1. Far1 and Cdc24 also undergo nuclear shuttling and the nuclear pool of Far1 may temporally regulate access of Cdc24 to the cell cortex. PMID- 11121777 TI - Transcriptional control of cell type and morphogenesis in Candida albicans. AB - Candida albicans has a number of transcriptional regulatory circuits that control aspects of cell type and cell morphogenesis. Recent work has uncovered a cryptic mating-type locus, and a variety of transcription factors that are important in regulation of the transition from yeast growth to hyphal growth. In some cases, the signalling pathways regulating these transcription factors are becoming defined. Analysis of phenotypic switching implicates internal factors, as well as external signals, in control of cellular morphogenesis. PMID- 11121778 TI - Coordinating development with the cell cycle in Caulobacter. AB - During the Caulobacter life cycle, the timing of DNA replication, cell division and development is precisely coordinated. Recent work has begun to unravel the complex regulatory networks that couple these processes. A key aspect of these regulatory networks is the dynamic localization of multiple histidine protein kinases that control a master response regulator, thus driving downstream pathways. PMID- 11121779 TI - Signal transduction during fertilization in the unicellular green alga, Chlamydomonas. AB - Sexual reproduction in the green alga, Chlamydomonas, is regulated by environmental conditions and by cell-cell interactions. After gametogenesis, flagellar adhesion between gametes triggers gamete activation, leading to cell fusion and zygote formation. Recent studies have identified new molecular events that underlie signal transduction during Chlamydomonas fertilization, including expression of a sex-determining protein, phosphorylation of a homeodomain protein, activity of a kinesin II and regulated translocation of an aurora/Ip11 like protein kinase from the cell body to the flagella. PMID- 11121780 TI - Developmental biology in marine invertebrate symbioses. AB - Associations between marine invertebrates and their cooperative bacterial symbionts offer access to an understanding of the roots of host-microbe interaction; for example, several symbioses like the squid-vibrio light organ association serve as models for investigating how each partner affects the developmental biology of the other. Previous results have identified a program of specific developmental events that unfolds as the association is initiated. In the past year, published studies have focused primarily on describing the mechanisms underlying the signaling processes that occur between the juvenile squid and the luminous bacteria that colonize it. PMID- 11121781 TI - Vegetative incompatibility in filamentous fungi: Podospora and Neurospora provide some clues. AB - In filamentous fungi, vegetative cell fusion between genotypically distinct individuals leads to a cell-death reaction known as vegetative or heterokaryon incompatibility. Genes involved in this reaction have been characterised molecularly. We can now begin to get a better understanding of the mechanism and the biological significance of this intriguing phenomenon. PMID- 11121782 TI - Hanging by a thread: invasion of legume plants by rhizobia. AB - Nitrogen-fixing nodules on plants such as alfalfa, pea and vetch arise from the root inner cortex and grow via a persistent meristem. Thus, these nodules are defined as indeterminate. The formation of functional indeterminate nodules requires that symbiotic bacteria, collectively called rhizobia, gain access to the interior of roots and root nodules via infection threads. Recent work has begun to elucidate the important functions of the root cell cytoskeleton in infection thread formation. It has also recently become apparent that rhizobial Nod factors and rhizobial exopolysaccharides play key roles in the initiation and elongation of infection threads. PMID- 11121783 TI - Heterocyst formation in cyanobacteria. AB - When deprived of combined nitrogen, many filamentous cyanobacteria develop a one dimensional pattern of specialised nitrogen-fixing cells, known as heterocysts. Recent years have seen the identification and characterisation of some of the key genes and proteins involved in heterocyst development and spacing, including the positive regulator HetR and the diffusible inhibitor PatS. PMID- 11121784 TI - Signalling molecules involved in cellular differentiation during Dictyostelium morphogenesis. AB - GSK-3, Dd-STATa, PKA, rZIP and Ras all play important roles in cell type determination of Dictyostelium discoideum. The fact that homologs of these proteins also function in metazoan development emphasizes the importance of Dictyostelium as a model microbial organism for studying the molecular mechanisms that regulate development. The recent elaboration of the central role for GSK-3 in cell type determination has been of particular importance. The stimulatory effect of extracellular cAMP on GSK-3 activity has been shown to act through the cell surface receptor cAR3 and a tyrosine protein kinase ZAK1, which directly activates and phosphorylates GSK-3. Several proteins, including Dd-STATa, have been identified as substrates for GSK-3, and are therefore potential transducers of the signals involved in cell type determination. PMID- 11121785 TI - Cell cycle regulation in Schizosaccharomyces pombe. AB - Cdc2, a cyclin-dependent kinase, controls cell cycle progression in fission yeast. New details of Cdc2 regulation and function have been uncovered in recent studies. These studies involve cyclins that associate with Cdc2 in G1-phase and the proteins that regulate inhibitory phosphorylation of Cdc2 during S-phase and G2-phase. Recent investigations have also provided a better understanding of proteins that regulate DNA replication and that are directly or indirectly controlled by Cdc2. PMID- 11121786 TI - Pattern formation: fruiting body morphogenesis in Myxococcus xanthus. AB - When Myxococcus xanthus cells are exposed to starvation, they respond with dramatic behavioral changes. The expansive swarming behavior stops and the cells begin to aggregate into multicellular fruiting bodies. The cell-surface associated C-signal has been identified as the signal that induces aggregation. Recently, several of the components in the C-signal transduction pathway have been identified and behavioral analyses are beginning to reveal how the C-signal modulates cell behavior. Together, these findings provide a framework for understanding how a cell-surface-associated morphogen induces pattern formation. PMID- 11121787 TI - The role of the T-pilus in horizontal gene transfer and tumorigenesis. AB - The T-pilus is a flexuous filamentous appendage that is essential for Agrobacterium tumefaciens virulence. T-pilus subunits are derived from a VirB2 processing reaction that generates cyclized polypeptide subunits. The T-pilus filament has a diameter of 10 nm and contains a lumen approximately 2 nm in diameter. Biogenesis of the T-pilus requires all 11 VirB proteins, but not the VirD4 protein, which is used in conjugal plasmid transfer. VirB4 and VirB11 are two ATPases that may form homohexameric rings within the transport apparatus, which is composed of VirB6-10 proteins. PMID- 11121788 TI - How can overexpression of Na(+),Ca(2+)-exchanger compensate the negative inotropic effects of downregulated SERCA? PMID- 11121789 TI - Nucleoside diphosphate kinase: a new player in heart failure? PMID- 11121790 TI - Dysfunctional ischemic preconditioning mechanisms in aging. PMID- 11121791 TI - A role for endothelin receptor blockade in improving the endothelial function of coronary artery grafts? PMID- 11121792 TI - Mitochondrial pathology in cardiac failure. PMID- 11121793 TI - Vasoactive intestinal peptide: cardiovascular effects. AB - Vasoactive intestinal peptide (VIP) is present in the peripheral and the central nervous systems where it functions as a nonadrenergic, noncholinergic neurotransmitter or neuromodulator. Significant concentrations of VIP are present in the gastrointestinal tract, heart, lungs, thyroid, kidney, urinary bladder, genital organs and the brain. On a molar basis, VIP is 50-100 times more potent than acetylcholine as a vasodilator. VIP release in the body is stimulated by high frequency (10-20 Hz) nerve stimulation and by cholinergic agonists, serotonin, dopaminergic agonists, prostaglandins (PGE, PGD), and nerve growth factor. The VIP peptide combines with its receptor and dose-dependently activates adenylyl cyclase. The vasodilatory effect of VIP in different vascular tissues or species also may be due to increases in nitric oxide, cyclic GMP, and other signaling agents. In the heart, VIP immunoreactive nerve fibers are present not only in the epicardial coronary arteries and veins, but also the sinoatrial node, atrium, interatrial septum, atrioventricular node, intracardiac ganglia, and ventricles (right ventricle >> left ventricle). In the coronary arterial walls, VIP may contribute to the regulation of normal coronary vasomotor tone. In research animals and in humans, VIP, administered into the coronary artery or intravenously, increases the epicardial coronary artery cross-sectional area, decreases coronary vascular resistance, and significantly increases coronary artery blood flow. High frequency parasympathetic (vagal) nerve stimulation also releases endogenous VIP in the coronary vessels and heart and significantly increases coronary artery blood flow. In addition, the release of VIP in the heart is increased during coronary artery occlusion and during reperfusion where VIP may promote local blood flow and may have a free-radical scavenging effect. VIP also has a primary positive inotropic effect on cardiac muscle that is enhanced by its ability to facilitate ventricular-vascular coupling by reducing mean arterial pressure by 10-15%. In concentrations of 10(-8)-10(-5) mol, VIP augments developed isometric force and increases atrial and ventricular contractility. The presence of VIP-immunoreactive nerve fibers in and around the sinus and the atrioventricular nodes of mammals strongly suggests that this peptide can affect the heart rate. In this regard, endogenously released or exogenous VIP can significantly increase the heart rate and has a more potent effect on heart rate than does norepinephrine. The presence and significant cardiovascular effects of VIP in the heart suggests that this peptide is important in the regulation of coronary blood flow, cardiac contraction, and heart rate. Current investigations are defining the physiological role of VIP in the regulation of cardiovascular function. PMID- 11121794 TI - Overexpression of the Na(+)/Ca(2+) exchanger and inhibition of the sarcoplasmic reticulum Ca(2+)-ATPase in ventricular myocytes from transgenic mice. AB - BACKGROUND: Myocytes from failing hearts produce slower and smaller Ca(2+) transients associated with reduction in expression of sarcoplasmic reticulum (SR) Ca(2+) ATPase and an overexpression of Na(+)/Ca(2+) exchanger. Since the physiological role of both these proteins is competing for, and removing, Ca(2+) from the cytoplasm, overexpression of the exchanger may compensate for less effective SR Ca(2+) uptake. This study demonstrates this compensatory effect and provides a quantitative description of the results. METHODS: Ventricular myocytes from transgenic mice overexpressing the Na(+)/Ca(2+) exchanger (TR) and nontransgenic littermates (NON) were used. Cell shortening, cytoplasmic [Ca] (using indo-1 AM) and electrophysiological parameters were monitored. RESULTS: TR myocytes displayed faster Ca(2+) transients and twitches compared with NON myocytes. Superfusion with thapsigargin prolonged the time-course of Ca(2+) transients of TR myocytes until these were equal to the ones measured in NON myocytes. The amount of SR Ca(2+)-ATPase (SERCA) inhibition needed to obtain such transients was calculated as a function of V(max) for the Ca(2+) flux via SERCA and found to be 28%. In TR myocytes V(max) for the Ca(2+) flux via Na(+)/Ca(2+)exchange was 240% of NON myocytes. When Ca(2+) transients in TR myocytes were slowed by thapsigargin to similar values to the ones recorded in NON myocytes, SR Ca(2+) content was also correspondingly reduced. CONCLUSIONS: The results suggest that in pathophysiological conditions where there is a reduction in SERCA function, overexpression of Na(+)/Ca(2+) exchanger can compensate and allow normal Ca(2+) homeostasis to be maintained. In mouse ventricular myocytes a 2.4-fold increase in Na(+)/Ca(2+) exchange activity compensates for a reduction in SERCA function by 28% so maintaining the duration of the Ca(2+) transient. PMID- 11121795 TI - Increased activity of membrane-associated nucleoside diphosphate kinase and inhibition of cAMP synthesis in failing human myocardium. AB - OBJECTIVE: Chronic heart failure is associated with a decreased responsiveness of the heart to beta-adrenergic receptor agonists. We recently demonstrated a receptor-independent activation of G proteins and modulation of cardiac adenylyl cyclase activity by sarcolemmal membrane-associated nucleoside diphosphate kinase. We wondered whether changes in the activity of nucleoside diphosphate kinase occur in heart failure and contribute to or compensate for the impairment in myocardial receptor-mediated cAMP generation. METHODS: Sarcolemmal membranes were purified from non-failing and failing human left ventricular myocardium. The protein level and activity of nucleoside diphosphate kinase were quantified. The influence of nucleoside diphosphate kinase on adenylyl cyclase activity was determined by measuring the effect of GDP on adenylyl cyclase activity in the absence and presence of nucleoside diphosphate kinase inhibitors. RESULTS: The amount and activity of nucleoside diphosphate kinase in sarcolemmal membranes from failing hearts (n=13) were increased 3- to 4-fold compared to levels in membranes from non-failing myocardium (n=5). This increase in sarcolemmal nucleoside diphosphate kinase activity resulted in a 50% inhibition of adenylyl cyclase activity over a range of GDP and ATP concentrations. CONCLUSION: The amount and activity of nucleoside diphosphate kinase are increased in sarcolemmal membranes of failing human myocardium, resulting in a substantial receptor independent inhibition of adenylyl cyclase activity. PMID- 11121796 TI - Direct activation of mitochondrial K(ATP) channels mimics preconditioning but protein kinase C activation is less effective in middle-aged rat hearts. AB - OBJECTIVES: This study is aimed to determine whether loss of preconditioning (IP) effects in the middle-aged hearts (MA) is due to the failure of protein kinase C (PKC) activation and, if so, whether direct activation of mitochondrial ATP sensitive potassium channels (m-K(ATP)) or PKC mimics IP. BACKGROUND: PKC is a mediator and m-K(ATP) may be its downstream effector of IP in young adult hearts (YA), but we have demonstrated that IP is not effective in MA. METHODS AND RESULTS: Isolated hearts from YA (12-week) and MA (50-week) Fischer 344 rats were preconditioned by three cycles of ischemia and reperfusion (5 min each), and the translocation of PKC isoforms and the effects on reperfusion injury were assessed. In some hearts activation of m-K(ATP) or PKC by diazoxide or 1, 2 dioctanoyl glycerol (DOG) was performed before 25 min of global ischemia/30 min of reperfusion. IP could improve the recovery of LV function and resulted in higher content of ATP after reperfusion in YA but these beneficial effects of IP was not found in MA. The effects of IP in YA were abolished by 5 hydroxydecanoate. In YA but not in MA, immunohistochemical analysis revealed that IP translocated PKC-alpha and delta from the cytosolic or membrane to the perinuclear region but immunoblotting analysis showed translocation of PKC-alpha, delta and epsilon to the membrane fraction. Pretreatment with diazoxide or DOG mimicked IP and decreased the creatine kinase release in YA. Diazoxide was also effective but effects of DOG were less in MA as compared with in YA. CONCLUSIONS: IP is not effective in MA hearts partly due to failure of translocation of PKC isoforms. Moreover, less efficacy of PKC activation by DOG as compared with activities of m-K(ATP) by diazoxide in MA may suggest that defect(s) of cell signaling downstream to PKC may also be involved in the loss of IP effects in MA. PMID- 11121797 TI - PR-39, a proline/arginine-rich antimicrobial peptide, exerts cardioprotective effects in myocardial ischemia-reperfusion. AB - OBJECTIVE: PR-39, a proline/arginine-rich antimicrobial peptide, has been shown to inhibit the NADPH oxidase activity of polymorphonuclear leukocytes (PMNs) by blocking assembly of this enzyme. We hypothesized that PR-39 could attenuate PMN induced cardiac dysfunction by suppression of superoxide production. METHODS: We examined the effects of PR-39 in isolated ischemic (20 min) and reperfused (45 min) rat hearts administered PMNs at the onset of reperfusion. RESULTS: PR-39 (4 or 10 microg/ml) given i.v. 30 min prior to ischemia-reperfusion (I-R) significantly improved left ventricular developed pressure (LVDP, P<0.01) and the maximal rate of development of LVDP (i.e. +dP/dt max, P<0.01) compared to I-R hearts obtained from rats given 0.9% NaCl. PR-39-treated PMNs (10 microg/ml) also significantly attenuated cardiac contractile dysfunction after I-R (P<0.01). Superoxide release was significantly reduced (P<0.01) in N-formylmethionyl leucylphenylalanine stimulated PMNs pretreated with 4 or 10 microg/ml PR-39. PR 39 also significantly attenuated P-selectin expression on the rat coronary microvascular endothelium and CD18 upregulation in rat PMNs. In addition, PR-39 significantly reduced PMN vascular adherence and infiltration into the post ischemic myocardium. CONCLUSION: These results provide evidence that PR-39 significantly attenuates PMN-induced cardiac contractile dysfunction in the I-R rat heart at least in part via suppression of superoxide release. This cardioprotection occurred both by inhibition of PMN and endothelial NADPH oxidase. PMID- 11121798 TI - Myocardial interstitial norepinephrine and dihydroxyphenylglycol levels during ischemia and reperfusion. AB - OBJECTIVE: The aim was to elucidate the relation between norepinephrine (NE) release and intraneuronal NE kinetics in the ischemic region of the in vivo heart. METHODS: Using dialysis technique in the heart of anesthetized cats, we sampled dialysate from the ischemic region during 120-min coronary occlusion and reperfusion. Dialysate NE and dihydroxyphenylglycol (DHPG) contents were measured as indexes of myocardial interstitial NE and DHPG levels. RESULTS: Within 20 min of occlusion, interstitial NE levels increased while DHPG levels decreased. This NE increase was suppressed by omega-conotoxin GVIA and enhanced by desipramine. These data suggest that axoplasmic NE levels increased by neuronal reuptake following exocytotic release, while intraneuronal DHPG production was suppressed due to the reduced monoamine oxidase activity. After 20 min of occlusion, interstitial NE levels increased markedly, accompanied by increased DHPG levels. This NE increase was suppressed by desipramine. These findings imply that NE mobilization from stored vesicles to axoplasma exceeded outward NE transport through uptake(1) carrier in the amount of NE, and that a substantial increase in axoplasmic NE levels compensated for the reduced monoamine oxidase activity. After reperfusion, interstitial NE levels rapidly decreased while DHPG levels increased further. Both responses in NE and DHPG were suppressed by desipramine, indicating the involvement of recovered neuronal reuptake function. CONCLUSIONS: In the ischemic region, intraneuronal DHPG production was affected by alterations in monoamine oxidase activity, NE mobilization from stored vesicles, and carrier mediated NE transport. The involvement of these factors in intraneuronal NE kinetics varied with the time of occlusion and reperfusion. PMID- 11121800 TI - Altered response to ibutilide in a heart failure model. AB - OBJECTIVE: Despite the frequent use of anti-arrhythmic drugs in the general population, the electrophysiologic effects of these agents have not been elucidated in congestive heart failure (CHF). METHODS: To examine the impact of left ventricular dysfunction on actions of type III anti-arrhythmic drugs, we evaluated the actions of ibutilide in a canine model of pacing-induced dilated cardiomyopathy. Following ablation of the atrioventricular node, effects on action potential duration at 90% (APD(90)) were compared in vivo, between eight CHF animals and seven controls. Monophasic action potential recordings were obtained from right and left ventricular endocardium/epicardium during and after three doses of ibutilide (0. 01, 0.02 and 0.05 mg/kg), at pacing cycle lengths of 300-1000 ms. RESULTS: APD(90) prolongation with ibutilide (0.01 mg/kg) was significantly greater in CHF vs. controls (P=0.0026, ANOVA). However, plasma ibutilide levels at this dose, were not significantly different between the two groups. In CHF, maximal effects were observed at the lowest dose, whereas effects were gradual and dose-dependent in controls. With ibutilide administration (0.01 mg/kg), an increased dispersion of left-right ventricular APD(90) was observed in CHF, but not in controls (P=0.03). A trend was observed, for increased incidence of non-sustained polymorphic ventricular tachycardia in CHF. CONCLUSIONS: In the presence of CHF, the actions of ibutilide are altered significantly. These findings may reflect altered tissue effects, as a consequence of myocardial electrical remodeling in CHF. PMID- 11121799 TI - Potential role of eNOS in the therapeutic control of myocardial oxygen consumption by ACE inhibitors and amlodipine. AB - OBJECTIVES: Our aim was to investigate the potential therapeutic role of endothelial nitric oxide synthase (eNOS) in the modulation of cardiac O(2) consumption induced by the angiotensin converting enzyme (ACE) inhibitor ramiprilat and amlodipine. METHODS: Three different groups of mice were used; wild type, wild type in the presence of N-nitro-L-arginine methyl ester (L-NAME, 10(-4) mol/l) or genetically altered mice lacking the eNOS gene (eNOS -/-). Myocardial O(2) consumption was measured using a Clark-type O(2) electrode in an air-tight stirred bath. Concentration-response curves to ramiprilat (RAM), amlodipine (AMLO), diltiazem (DIL), carbachol (CCL), substance P (SP) and S nitroso-N-acetyl-penicillamine (SNAP) were performed. The rate of decrease in O(2) concentration was expressed as a percentage of the baseline. RESULTS: Baseline O(2) consumption was not different between the three groups of mice. In tissues from wild type mice, RAM (10(-5) mol/l), AMLO (10(-5) mol/l), DIL (10(-4) mol/l), CCL (10(-4) mol/l), SP (10(-7) mol/l) and SNAP (10(-4) mol/l) reduced myocardial O(2) consumption by -32+/-4, -27+/-10, -20+/-6, -25+/-2, -22+/-4 and 42+/-4%, respectively. The responses to RAM, AMLO, CCL and SP were absent in tissues taken from eNOS -/- mice (-7.1+/-4.3, -5.0+/-6.0, -5.2+/-5.1 and -0.4+/ 0.2%, respectively). In addition, L-NAME significantly (P<0.05) inhibited the reduction in O(2) consumption induced by RAM (-9.8+/-4.4%), AMLO (-1.0+/-6.0%), CCL (-8.8+/-3.7%) and SP (-6.6+/-4.9%) in cardiac tissues from wild type mice. In contrast, NO-independent responses to the calcium channel antagonist, DIL, and responses to the NO donor, SNAP, were not affected in cardiac tissues taken from eNOS -/- (DIL: -20+/-4%; SNAP: -46+/-6%) or L-NAME-treated (DIL: -17+/-2%; SNAP: 33+/-5%) mice. CONCLUSIONS: These results suggest that endogenous endothelial NO synthase derived NO serves an important role in the regulation of myocardial O(2) consumption. This action may contribute to the therapeutic action of ACE inhibitors and amlodipine. PMID- 11121801 TI - Greater susceptibility of failing cardiac myocytes to oxygen free radical mediated injury. AB - OBJECTIVE: Oxygen-derived free radicals can produce myocardial cellular damage, which might contribute to the ischemia-reperfusion injury and to heart failure (HF). However, the effects of oxygen radicals on myocyte structure have not been examined in the failing heart. METHODS: We examined the susceptibility of intact cardiac myocytes isolated from control (n=16) and rapid pacing (240 bpm, 4 wks) induced HF (n=8) dog hearts to an exogenous hydroxyl radical (.OH), generated from H(2)O(2) and Fe(3+)-nitrilotriacetate. The production of (.OH) was monitored by electron spin resonance with 5,5'-dimethyl-1-pyroline-N-oxide (DMPO) as a spin trap. RESULTS: The magnitude of DMPO-OH signals was not attenuated in the presence of either control or HF myocytes. (.OH) induced a time-dependent decrease in myocyte length (i.e. hypercontracture). The time to the onset of hypercontracture and that to the submaximal hypercontracture after exposure was significantly shortened in HF. Activities of superoxide dismutase, catalase, and glutathione peroxidase was not decreased in HF. CONCLUSIONS: HF myocytes were more susceptible to oxidative stress-induced cellular injury, which was not due to decreased antioxidant defense, but to the intrinsic properties of cells. PMID- 11121802 TI - Significance of timing of angiotensin AT1 receptor blockade in rats with myocardial infarction-induced heart failure. AB - OBJECTIVE: Blockade of angiotensin AT(1) receptors has been shown to prevent cardiac remodeling and improve left ventricular function and survival after myocardial infarction (MI). However, the timing of initiation of treatment has not been fully elucidated. Therefore, the purpose of the present study was to compare the effects of very early (30 min after MI), early (3 and 24 h after MI) and delayed (7 days after MI) treatments with the angiotensin AT(1) receptor antagonist fonsartan (HR 720) on cardiac morphological and hemodynamic parameters in a rat model of MI-induced heart failure and to establish the therapeutic window for the start of treatment. METHODS: Male Wistar rats underwent coronary ligation and were randomized fonsartan (HR720) treatment starting 30 min, 3 h, 24 h and 7 days after MI or no treatment. Treatment was continued up to 6 weeks post MI. RESULTS: Fonsartan (HR720) treatment attenuated cardiac hypertrophy when treatment started 30 min or later after MI, limited infarct size when treatment initiated 3 and 24 h after MI, decreased left ventricular end-diastolic pressure when treatment started 3 h to 7 days after MI, and improved dP/dt(max) when treatment commenced 24 h and 7 days after MI compared to untreated infarct group. CONCLUSION: Our results show that angiotensin AT(1) receptor blockade with fonsartan (HR720) produced the best cardioprotective effects when treatment was started 3 to 24 h after MI although a start of treatment 7 days following MI still could improve functional parameters. These results suggest an optimal time window for the start of treatment with angiotensin AT(1) receptor antagonists seems to be between 3 and 24 h post MI. PMID- 11121803 TI - Low level of sarcolemmal phosphatidylinositol 4,5-bisphosphate in cardiomyopathic hamster (UM-X7.1) heart. AB - OBJECTIVE: Phosphatidylinositol 4,5-bisphosphate (PtdIns 4,5-P(2)) is not only a precursor to inositol 1,4,5-trisphosphate (Ins 1,4, 5-P(3)) and sn-1,2 diacylglycerol, but also essential for the function of several membrane proteins. The aim of this study was to evaluate the changes in the level of this phospholipid in the cell plasma membrane (sarcolemma, SL) of cardiomyopathic hamster (CMPH) heart. METHODS: We examined the cardiac SL PtdIns 4,5-P(2) mass and the activities of the enzymes responsible for its synthesis and hydrolysis in 250-day-old UM-X7.1 CMPH at a severe stage of congestive heart failure (CHF) and in age-matched controls (Syrian Golden hamsters). RESULTS: The SL PtdIns 4,5-P(2) mass in CMPH was reduced by 72% of the control value. The activities of PtdIns 4 kinase and PtdIns 4-P 5 kinase were depressed by 69 and 50% of control values, respectively. Although, the total phospholipase C (PLC) activity was moderately, although significantly, decreased (by 18% of control), PLCdelta(1) isoenzyme activity in the SL membrane was elevated, with a concomitant increase in its protein content, whereas PLCbeta(1) and gamma(1) isoenzyme activities were depressed despite the increase in their protein levels. A 2-fold increase in the Ins 1,4,5-P(3) concentration in the cytosol of the failing heart of CMPH was also observed. CONCLUSIONS: Reduced SL level of PtdIns 4, 5-P(2) may severely jeopardize cardiac cell function in this hamster model of CHF. In addition, the profound changes in the profile of heart SL PLC isoenzyme could alter the complex second messenger responses of these isoenzymes, and elevated Ins 1,4,5-P(3) levels may contribute to intracellular Ca(2+) overload in the failing cardiomyocyte. PMID- 11121804 TI - Apoptosis of ventricular and atrial myocytes from pacing-induced canine heart failure. AB - OBJECTIVE: Rapid ventricular pacing in dogs results in a low output cardiomyopathic state similar to idiopathic dilated cardiomyopathy in man. Cell death by apoptosis may play an important role in the loss of cardiac function. This study investigates the molecular pathways involved in the regulation of apoptosis in dogs with pacing-induced heart failure. METHODS: Apoptosis was identified by terminal transferase nick end-labelling (TUNEL) in the ventricles and atria of dog hearts affected by rapid-ventricular pacing. Western blots were used to determine expression of the components involved in the initiation (Fas, Fas-Ligand, FADD), regulation (Bcl-2, Bax) and execution (caspase-2 and caspase 3) of apoptosis. RESULTS: Pacing-induced heart failure resulted in a significant increase in the number of ventricular and atrial myocyte nuclei undergoing apoptosis as measured by TUNEL. Compared to the samples from control hearts (n=6) the expression of Bcl-2, an inhibitor of apoptosis, was significantly reduced in ventricles from five dogs with pacing-induced heart failure. No change in the expression of the apoptotic inducer Bax was detected. Fas and FADD were significantly elevated in all paced ventricles, and Fas-L was only detected in the paced hearts. Both caspase-2 and caspase-3 were elevated following ventricular pacing. CONCLUSIONS: We have identified components of the signalling pathways along which apoptosis proceeds following the induction of heart failure in dogs. Apoptosis was also detected in the atria raising the possibility that, like human dilated cardiomyopathy, the molecular changes are global. PMID- 11121805 TI - Oxidized low density lipoprotein induces apoptosis via generation of reactive oxygen species in vascular smooth muscle cells. AB - OBJECTIVE: Death of vascular smooth muscle cell (VSMC) induced by oxidized LDL (oxLDL) can occur by both necrosis and apoptosis which may contribute to plaque instability and rupture. Reactive oxygen species (ROS) induces apoptosis in VSMC and is involved in oxLDL action, we tested the hypothesis here that a coupling exists between ROS generation and apoptosis of oxLDL-treated VSMC. METHODS: Cultured VSMC from rat aorta were treated with oxLDL, apoptosis and necrosis were distinguished by using FITC-annexin V label and propidium iodide stain, analyzed by flow cytometry. ROS generation of VSMCs was detected by the fluorescence intensity of DCF. Apoptosis was also determined by cleavage of procaspase-3. RESULTS: OxLDL induced apoptosis (3 h) in a dose-dependent manner and reached maximum (with near-basal necrosis) at a concentration of 300 microg/ml. At this and lower (100 microg/ml) concentration, oxLDL, but not native LDL, stimulated ROS production rapidly (< or =5 min) and ROS level remained elevated for at least 45 min. Catalase and deferoxamine reduced both oxLDL-induced apoptosis and ROS generation. Superoxide dismutase and benzoic acid neither reduced the oxLDL induced ROS generation nor inhibited apoptosis. Since oxLDL-induced ROS generation were inhibited by nordihydroguaiaretic acid and rotenone, lipoxygenase and mitochondrial pathways could be involved. In addition, catalase, deferoxamine, and N-acetylcysteine inhibited oxLDL-induced cleavage of procaspase 3 as well. CONCLUSIONS: ROS generation and apoptosis are tightly coupled in oxLDL treated VSMCs. Antioxidants that reduced ROS level inhibited apoptosis, those that did not reduce ROS level were ineffective. Both mitochondrial and lipoxygenase activities may be involved. PMID- 11121806 TI - Endothelin receptor blockade improves endothelial function in human internal mammary arteries. AB - OBJECTIVE: Endothelial dysfunction, specifically endothelium-derived contracting factors have been implicated in the development of arterial conduit vasospasm. The potent vasoconstrictor endothelin-1 (ET-1) has received much attention in this regard. The present study was designed to evaluate the role of ET-1 in the development of endothelial dysfunction in human internal mammary arteries (IMA). To this aim, we examined the effects of specific and non-specific ET-receptor antagonists on endothelial function (assessed using acetylcholine (ACh)-induced vasodilation) in segments of IMA obtained during coronary artery bypass graft (CABG) surgery. METHODS: Vascular segments of IMA were obtained from 51 patients undergoing elective coronary artery bypass graft (CABG) surgery and in vitro endothelium-dependent and -independent responses to ACh and sodium nitroprusside (SNP) were assessed. Isometric dose response curves (DRC) to ACh and SNP were constructed in pre-contracted rings in the presence and absence of bosentan (ET(A/B) receptor antagonist, 3 microM), BQ-123 (ET(A) antagonist, 1 microM) and BQ-788 (ET(B) antagonist, 1 microM) using the isolated organ bath apparatus. Percent maximum relaxation (%E(max)) and sensitivity (pEC(50)) were compared between interventions. RESULTS: ACh caused dose-dependent endothelium-mediated relaxation in IMA (%E(max) 43+/-4, pEC(50) 6. 74+/-0.12). In the presence of bosentan, BQ-123 and BQ-788 ACh-induced relaxation was significantly augmented (%E(max) bosentan 60+/-3, BQ-123 56+/-4, BQ-788 53+/-5 vs. control 43+/-4, P<0.05) without affecting sensitivity. The effects of these antagonists were endothelium-specific since endothelium-independent responses to SNP remained unaltered. Furthermore, the beneficial effects were independently and maximally mediated by ET(A) and ET(B) receptors (%E(max) BQ-123 56+/-4 vs. BQ-788 53+/-5 vs. bosentan 60+/-3, P>0. 05). CONCLUSIONS: These data uncover, for the first time, beneficial effects of ET receptor blockade on endothelial-dependent vasorelaxation in human IMA. PMID- 11121807 TI - Cysteinyl leukotrienes are involved in angiotensin II-induced contraction of aorta from spontaneously hypertensive rats. AB - OBJECTIVE: Non specific lipoxygenase inhibitors have been reported to reduce the in vitro constrictor response and the in vivo pressor effect of angiotensin II in rats. The aim of this study was to assess the role of cysteinyl leukotrienes, in the vascular response to angiotensin II in spontaneously hypertensive rats (SHR). METHODS: Rings of thoracic aorta from SHR and normotensive Wistar-Kyoto rats (WKY) were compared in terms of contractile responses and release of cysteinyl leukotrienes in response to angiotensin II. RESULTS: Pretreatment with the specific 5-lipoxygenase inhibitor AA861 10 microM reduced the efficacy of angiotensin II in intact and endothelium-denuded aorta from SHR (% inhibition vs. control: 65+/-12.6% with endothelium (n=6), P<0.05; 43+/-7.2% without endothelium (n=6), P<0.05) but not in aorta from WKY. In addition, in aorta from SHR, the CysLT(1) receptor antagonist MK571 1 microM reduced by 55+/-6.1% (n=6, P<0.05) the contractile effects of angiotensin II in rings with endothelium but not in endothelium-denuded rings. Angiotensin II induced a 8.6+/-2.1-fold increase in cysteinyl leukotriene production in aorta rings from SHR with endothelium which was prevented by the AT(1) receptor antagonist losartan 1 microM but not by the AT(2) receptor antagonist PD123319 0.1 microM. In aorta rings from WKY, cysteinyl leukotriene production remained unchanged after exposition to angiotensin II. The cysteinyl leukotrienes (up to 0.1 microM) induced contractions in aorta rings from SHR but not from WKY. CONCLUSIONS: These data suggest that cysteinyl leukotrienes, acting at least in part on endothelial CysLT(1) receptors, are involved in the contractile response to angiotensin II in isolated aorta from SHR but not from WKY. PMID- 11121808 TI - Capillary recruitment is impaired in essential hypertension and relates to insulin's metabolic and vascular actions. AB - OBJECTIVE: In patients with essential hypertension, defects in both the metabolic and vascular actions of insulin have been described. Impaired microvascular function, a well-established abnormality in essential hypertension, may explain part of these defects. In the present study we investigated whether microvascular function is impaired in essential hypertension and relates to insulin's metabolic and vasodilatatory actions. METHODS: We measured 24-h ambulatory blood pressure, capillary recruitment after arterial occlusion, and skin blood flow responses to iontophoresis of acetylcholine and sodium nitroprusside in 18 subjects with untreated essential hypertension and in 18 control subjects. Whole body insulin sensitivity and leg insulin-mediated vasodilatation were assessed with the hyperinsulinaemic clamp technique and plethysmography. RESULTS: Hypertensive, as compared to normotensive, subjects had a decreased insulin sensitivity (0.8+/-0.3 vs. 1.7+/-0. 6 mgkg(-1)min(-1) per pmoll(-1); P<0.001), capillary recruitment after arterial occlusion (21.5+/-5.8 vs. 45.9+/-10.4%; P<0.001), acetylcholine mediated vasodilatation (331+/-84 vs. 688+/-192%; P<0. 001), and insulin-mediated vasodilatation (median 29.3 vs. 47.2%; P<0.05). Correlation analyses with adjustment for sex, age, body mass index and waist-to-hip ratio showed significant relationships of capillary recruitment after arterial occlusion with blood pressure (r=-0.68; P<0.01), insulin sensitivity (r=+0.55; P<0.01) and insulin-mediated vasodilatation (r=+0.51; P<0.05), which extended from the normotensive to the hypertensive range. CONCLUSION: Skin microvascular function is associated with blood pressure and insulin's metabolic and vasodilatatory actions, both in normotensive and hypertensive subjects. These findings offer a potential mechanistic explanation of the links among insulin resistance, impaired insulin-mediated vasodilatation and hypertension. PMID- 11121809 TI - Endovascular irradiation with the liquid beta-emitter Rhenium-188 to reduce restenosis after experimental wall injury. AB - OBJECTIVE: Postinterventional irradiation is a new therapeutic concept in the prevention of restenosis. The liquid beta-emitter Rhenium-188 allows endovascular brachytherapy using a conventional balloon catheter without the problem of centering the radiation source. In an animal model of restenosis the feasibility and the dose dependent effect of intravascular brachytherapy with a Rhenium-188 filled balloon catheter was investigated. METHODS: In 68 male New Zealand White rabbits after endothelial denudation of the right common carotid artery with a Fogarty catheter, endovascular irradiation was performed with a Rhenium-188 filled 3.0-mm balloon catheter using different dosages (0, 7.5, 15, 30, 45 and 60 Gy at the surface of the vessel). Then 4 weeks after the intervention the vessels were excised and histologically analyzed. RESULTS: Whereas at 7.5 Gy the intimal area (median [first quartile; third quartile]) did not differ significantly from the control (0.46 mm(2) [0.33 mm(2), 0.75 mm(2)] vs. 0.49 mm(2) [0.34 mm(2), 0.66 mm(2)]), neointimal hyperplasia was decreased significantly at 15 Gy (0.15 mm(2) [0.04 mm(2), 0.17 mm(2)]) and 30 Gy (0.07 mm(2) [0.04 mm(2), 0. 10 mm(2)]), and completely inhibited at the highest dosages (45 Gy: 0 mm(2) [0 mm(2), 0.04 mm(2)]; 60 Gy: 0 mm(2) [0 mm(2), 0.01 mm(2)]). CONCLUSIONS: Catheter transmitted endovascular irradiation with the liquid beta-emitter Rhenium-188 after vascular injury is feasible and effectively reduced neointimal hyperplasia in hypercholesterolemic rabbits. A significant reduction of the neointimal formation could be found already at a radiation absorbed dose of 15 Gy at the vessel surface. Following a surface dosage of 45 Gy the proliferative response to the vessel injury is almost completely abolished. PMID- 11121810 TI - Transient down-regulation of L-type Ca(2+) channel and dystrophin expression after balloon injury in rat aortic cells. AB - OBJECTIVE: Migration and proliferation of arterial smooth muscle cells are critical responses during restenosis after balloon angioplasty. We investigated the changes in the expression of Ca(2+) channels and dystrophin, two determinants of contraction, after balloon injury of rat aortas. METHODS: Proliferation and migration of aortic myocytes were triggered in vivo by the passage of an inflated balloon catheter in the aortas of 12-week-old male Wistar rats. We used the whole cell patch clamp technique to investigate Ba(2+) currents (I(Ba)) through Ca(2+) channels in single cells freshly isolated from media and neointima at various times after injury (days 2, 7, 15, 30 and 45). RESULTS: No T-type Ca(2+) channel current was recorded in any cell at any time. In contrast, a dihydropyridine (DHP)-sensitive L-type I(Ba)was recorded consistently in the media of intact aorta. After aortic injury, I(Ba) decreased dramatically (at days 2 and 7) but recovered over time to reach normal amplitude on days 30 and 45. In the neointima, I(Ba) was absent on day 15 but also increased gradually over time as observed at days 30 and 45. The use of a specific antibody directed against the L type Ca(2+) channel alpha(1C) subunit showed, both by immunostaining and by Western blotting, no expression of the Ca(2+) channel protein on day 15. Parallel immunodetection of dystrophin showed that this marker of the contractile phenotype of SMCs was also not detectable at this stage in neointimal cells. Both proteins were re-expressed at days 45 and 63. Balloon injury induces a transient down-regulation of I(Ba) in arterial cells. CONCLUSIONS: Cell dedifferentiation and proliferation in vivo abolish the expression of L-type Ca(2+) channels and dystrophin in neointimal cells. These changes may be critical in the regulation of Ca(2+) homeostasis and, thereby, contraction of the arterial SMCs during restenosis following angioplasty. PMID- 11121811 TI - Transient interaction of activated platelets with endothelial cells induces expression of monocyte-chemoattractant protein-1 via a p38 mitogen-activated protein kinase mediated pathway. Implications for atherogenesis. AB - OBJECTIVE: Activated platelets induce alterations of chemotactic and adhesive properties of endothelial cells, a critical initial step in atherogenesis. We investigated the effect of transient interaction of activated platelets with cultured human umbilical vein endothelial cells (HUVECs) on secretion of monocyte chemoattractant protein-1 (MCP-1), a key molecule in monocyte chemotaxis and transmigration. METHODS AND RESULTS: Transient interaction of alpha-thrombin activated platelets with endothelial cells for 10-120 min substantially induced endothelial secretion of MCP-1, monocyte chemotaxis and adhesion to HUVECs. Platelet-induced secretion of MCP-1 and monocyte-endothelium adhesion was reduced by the MAP kinase p38-specific inhibitor SB203580, but not by other kinase inhibitors including PD98059, wortmannin, or rapamycin. In addition, activated platelets induced transcription of a luciferase reporter construct containing a MCP-1 promotor, an effect that could be inhibited by SB203580. Overexpression of dominant-negative mutants of MAP kinase p38, CSBP2-(D168A) and CSBP2 (T180E,Y182E) reduced platelet-induced expression of MCP-1. CONCLUSIONS: Activation of the p38 MAP kinase and consecutive endothelial secretion of MCP-1 induced through transient interaction of activated platelets might play an important role in atherogenesis. PMID- 11121812 TI - Regulation of endothelin-1 synthesis in human pulmonary arterial smooth muscle cells. Effects of transforming growth factor-beta and hypoxia. AB - OBJECTIVE: Endothelin-1 (ET-1) potently regulates pulmonary vascular tone and promotes vascular smooth muscle cell growth. Clinical and animal studies implicate increased ET-1 production in the pathogenesis of primary and secondary pulmonary hypertension. Although pulmonary arterial smooth muscle cells (PASMCs) synthesize ET-1 under basal conditions, it is unknown whether factors that may be important in pulmonary hypertension, such as transforming growth factor-beta (TGF beta) or hypoxia, augment ET-1 production by these cells. METHODS: We determined the effect of TGF-beta and hypoxia on ET-1 release and preproET-1 mRNA from cultured rat and human PASMCs. RESULTS: In the basal state, rat and human PASMCs synthesize, on average (mean+/-S.E.M.), 872+/-114 and 563+/-57 pg ET-1/mg cell protein over 24 h, respectively, a level that causes autocrine and paracrine effects in other tissues. TGF-beta significantly increases the expression of preproET-1 mRNA and ET-1 production by both rat and human PASMCs. Hypoxia for 24 h, however, does not affect ET-1 release from rat or human PASMCs. CONCLUSIONS: Cultured rat and human PASMCs are a source of ET-1 production. Enhanced ET-1 release from PASMCs may contribute to the pathophysiology of TGF-beta-induced pulmonary hypertension. ET-1 production by PASMCs is unlikely to contribute to the role of ET-1 in hypoxia-induced pulmonary vasoconstriction. PMID- 11121813 TI - Capillary filtration is reduced in lungs adapted to chronic heart failure: morphological and haemodynamic correlates. AB - OBJECTIVE: To determine pulmonary capillary filtration in experimental chronic heart failure and to investigate some morphological and haemodynamic mechanisms that could account for reduced filtration in lungs adapted to chronic heart failure. METHODS: We studied pulmonary capillary filtration, vascular resistances and morphology in lungs from guinea-pigs adapted to chronic heart failure. Heart failure was induced by banding of the ascending aorta (n=66) or sham control operation (n=78) in guinea-pigs which were studied at 150+/-8 days post operation. RESULTS: Reduced cardiac output, increased systemic vascular resistance and LV end diastolic pressure and increased LV and RV weight:body weight ratio (all P<0.05) indicated chronic heart failure at 5 months following aortic banding in guinea-pigs. Lung weight was increased (61%, P<0.05) in heart failure compared with controls, but lung water content was reduced (5.5%, P<0.05), a reversal of the pattern seen acutely. Studies in isolated perfused lungs demonstrated a reduced capillary filtration coefficient (0. 018+/-0.003 vs. 0.003+/-0.002 ml min(-1)mmHg(-1)g(-1), P<0.001), increased arterial (61%) and venous resistance (50%) in heart failure lungs, P<0.05. Wall thickness:lumen ratio was increased in small (<250 microm) pulmonary arterioles (0.15+/-0.02 vs. 0.08+/-0. 01) and venules (0.06+/-0.005 vs. 0.04+/-0.002) in heart failure, P<0.01. Alveolar septal volume fractions (35.2+/-5.1 vs. 23.1+/-2.7) and septal:air-space volume ratios (60.5+/-13.6 vs. 31.9+/-5.3) were also increased in heart failure, P<0.05. CONCLUSIONS: Pulmonary adaptation to chronic heart failure is associated with vascular and alveolar remodelling that contributes to increased vascular resistance and reduced capillary filtration. These changes are likely to be important in mediating resistance to pulmonary oedema in chronic heart failure. PMID- 11121814 TI - Increased alpha(1)- and alpha(2)-adrenoceptor-mediated contractile responses of human skeletal muscle resistance arteries in chronic limb ischemia. AB - OBJECTIVE: Recently, we have shown augmented contractile responses of skeletal muscle resistance arteries to noradrenaline in patients with critical limb ischemia. We investigated whether this increased sensitivity in skeletal muscle resistance arteries is due to either alpha(1)- or alpha(2)-adrenoceptor-mediated responses or both. METHODS: Skeletal muscle resistance arteries were isolated from the proximal (non-ischemic) and distal (ischemic) parts of limbs amputated for critical limb ischemia and mounted on a small vessel wire myograph. Cumulative concentration response curves of the vessel segments to noradrenaline, phenylephrine and brimonidine were obtained in the presence or the absence of the selective antagonists, prazosin and RS79948. RESULTS: Noradrenaline and phenylephrine produced almost equal maximal contractile responses. Brimonidine responses were smaller and were almost abolished by 0.1 microM RS 79948 while those of phenylephrine and noradrenaline were not affected. Prazosin reduced the maximum responses to brimonidine, shifted the concentration response curves of noradrenaline and phenylephrine rightwards giving pK(B) values of 9.86 and 9.33, respectively. Maximum responses produced by all three agonists in distal vessels were significantly higher than those obtained in proximal vessels. CONCLUSIONS: Noradrenaline contractile responses in skeletal muscle resistance arteries are predominantly mediated by alpha(1)-adrenoceptors. Both alpha(1)- and alpha(2) adrenoceptor-mediated responses are increased in the arteries from ischemic regions that may aggravate the decreased blood flow to the limbs due to arterial occlusion. PMID- 11121815 TI - Effect of gamma-melanocyte-stimulating hormones on baroreflex sensitivity and cerebral blood flow autoregulation in rats. AB - OBJECTIVE: In the present paper, we are interested in the effects of gamma melanocyte-stimulating hormones (gamma-MSHs) on cardiovascular regulatory systems. METHODS: Mean arterial pressure (MAP), cerebral blood flow (CBF) and heart rate (HR) were measured in urethane-anaesthetised rats after intravenous administration of lysgamma(2)-MSH, gamma(2)-MSH, gamma(2)-MSH(6-12) or phenylephrine. RESULTS: The gamma-MSHs caused an increase in MAP, CBF and HR, whereas phenylephrine caused an increase in MAP and CBF and baroreceptor reflex mediated bradycardia. All tested gamma-MSHs showed a significant impairment of the baroreceptor reflex sensitivity and CBF autoregulation as compared to the phenylephrine group. gamma(2)-MSH shows identical effects on the baroreceptor reflex and CBF as the endogenous occurring lysgamma(2)-MSH. In addition, the C terminal fragment of gamma(2)-MSH, gamma(2)-MSH(6-12), induced similar effects as gamma(2)-MSH. The level of increase in MAP was comparable between the gamma-MSHs and the phenylephrine group. CONCLUSIONS: The present study suggests that gamma(2)-MSH and the shorter fragment gamma(2)-MSH(6-12) impair baroreceptor reflex sensitivity, due to a strong increase in sympathetic tone and/or change in baroreceptor reflex setpoint, and induce cerebrovasodilatation, which can counteract an autoregulation-mediated cerebrovasoconstriction due to systemic pressor effects. Furthermore, the results indicate that the C-terminal site of gamma(2)-MSH is relevant for its central-mediated inhibitory effects on the baroreceptor reflex and CBF. PMID- 11121816 TI - Lymphocytes from spontaneously hypertensive rats exhibit enhanced adenylyl cyclase-Gi protein signaling. AB - OBJECTIVE: We have previously demonstrated an augmented activation of Gialpha proteins in heart and aorta from spontaneously hypertensive rats (SHRs), which was attributed to an enhanced expression of Gialpha proteins. Since immortalized lymphoblasts derived from lymphocytes of hypertensive patients have been shown to have enhanced Gi activation, the present studies were undertaken to investigate if lymphocytes from SHRs also exhibit enhanced Gi activation and whether this activation is related to enhanced expression of Gi proteins. METHODS: The levels of G-proteins and mRNA were determined by immunoblotting and Northern blotting techniques, using specific antibodies and cDNA probes, respectively. Adenylyl cyclase activity stimulated or inhibited by agonists was determined to examine the functions of G-proteins. RESULTS: The levels of Gialpha-2, Gialpha-3, Gbeta but not of Gs(alpha45) and Gs(alpha47) were significantly increased in lymphocytes from SHRs as compared to their control Wistar Kyoto (WKY) rats. Similarly the mRNA levels of Gialpha-2 and Gialpha-3 were significantly augmented in SHRs as compared to their age-matched WKYs. The increased levels of Gialpha were reflected in increased functions of Gi in SHRs as indicated by increased inhibition of forskolin-stimulated adenylyl cyclase activity by GTPgammaS. The activity of adenylyl cyclase stimulated by GTPgammaS, isoproterenol, NECA, NaF and forskolin was significantly decreased in SHRs as compared to their age matched WKY rats. On the other hand, inhibitory hormones, atrial natriuretic peptide and angiotensin II inhibited adenylyl cyclase activity to a greater extent in SHRs as compared to their age-matched WKY rats. CONCLUSIONS: These results indicate that lymphocytes from spontaneously hypertensive rats exhibit enhanced Gi activation (function) which may be attributed to the enhanced expression of Gi proteins. It may be suggested that enhanced Gi expression and associated signaling may be one of the factors responsible for enhanced lymphoblasts proliferation observed in hypertension. PMID- 11121817 TI - Horizontal versus vertical transmission of parasites in a stochastic spatial model. AB - A number of pathogens may be transmitted from parent to child at or before birth (vertically) or from one individual to another by contact (horizontally). A natural deterministic and non-spatial model, introduced by Lipsitch et al. [Proc. Roy. Soc. London Ser. B 260 (1995) 3211 shows that an epidemic is possible if the vertical transmission or the horizontal transmission is high enough. In contrast, we introduce a stochastic spatial model that shows that, on a particular graph, if the vertical transmission is not high enough, then the infected individuals disappear even for very high horizontal transmission. This illustrates the fact that introducing space may greatly change the qualitative behavior of a model. PMID- 11121818 TI - Game theory and evolution: finite size and absolute fitness measures. AB - This article is concerned with the characterization and existence of evolutionarily stable strategies (ESS) in Games against Nature, a class of models described by finite size populations and absolute fitness measures. We address these problems in terms of a new formalism which revolves around the concept evolutionary entropy, a measure of the diversity of options associated with a strategy pure - strategies have zero entropy, mixed strategies positive entropy. We invoke this formalism to show that ESS are characterized by extremal states of entropy. We illustrate this characterization of ESS by an analysis of the evolution of the sex ratio and the evolution of seed size. PMID- 11121819 TI - The effective population size of some age-structured populations. AB - It was shown in a previous paper that if generations are discrete, then the effective population size of a large population can be derived from the theory of multitype branching processes. It turns out to be proportional to the reciprocal of a term that appears in the denominator of expressions for survival probabilities when there is a supercritical positively regular branching process for which the dominant positive eigenvalue of the first moment matrix is slightly larger than 1. If there is an age-structured population with unchanging proportions among sexes and age groups, then the effective population size is shown to be also obtainable from the theory of multitype branching processes. The expression for this parameter has the same form as in the corresponding model for discrete generations, multiplied by an appropriate measure of the average length of a generation. Results are obtained for dioecious random mating populations, populations reproducing partly by selfing, and populations reproducing partly by full-sib mating. PMID- 11121820 TI - Applying the saddlepoint approximation to bivariate stochastic processes. AB - The problem of moment closure is central to the study of multitype stochastic population dynamics since equations for moments up to a given order will generally involve higher-order moments. To obtain a Normal approximation, the standard approach is to replace third- and higher-order moments by zero, which may be severely restrictive on the structure of the p.d.f. The purpose of this paper is therefore to extend the univariate truncated saddlepoint procedure to multivariate scenarios. This has several key advantages: no distributional assumptions are required; it works regardless of the moment order deemed appropriate; and, we obtain an algebraic form for the associated p.d.f. irrespective of whether or not we have complete knowledge of the cumulants. The latter is especially important, since no families of distributions currently exist which embrace all cumulants up to any given order. In general the algorithm converges swiftly to the required p.d.f.; analysis of a severe test case illustrates its current operational limit. PMID- 11121822 TI - Introduction. PMID- 11121821 TI - A mathematical model for the roles of pericytes and macrophages in the initiation of angiogenesis. I. The role of protease inhibitors in preventing angiogenesis. AB - In this paper, a simple mathematical model developed in H.A. Levine, B.D. Sleeman, M. Nilsen-Hamilton [J. Math. Biol., in press] to describe the initiation of capillary formation in tumor angiogenesis is extended to include the roles of pericytes and macrophages in regulating angiogenesis. The model also allows for the presence of anti-angiogenic (angiostatic) factors. The model is based on the observation that angiostatin can prevent the degradation of fibronectin in the basal lamina by inhibiting the catalytic action of active proteolytic enzyme. That is, it is proposed that the inhibitor 'deactivates' the protease but that it does not reduce the over all concentration of the protease. It consequently explores the possibility of preventing neovascular capillaries from migrating through the extra-cellular matrix toward the tumor by inhibiting protease action. The model is based on the theory of reinforced random walks coupled with Michaelis-Menten mechanisms which view endothelial cell receptors as the catalysts for transforming both tumor and macrophage derived angiogenic factors into proteolytic enzyme which in turn degrade the basal lamina. A simple catalytic reaction is proposed for the degradation of the basal lamina by the active proteases. A mechanism, in which the angiostatin acts as a protease inhibitor is discussed which has been substantiated experimentally. A second mechanism for the production of protease inhibitor from angiostatin by endothelial cells is proposed to be of Michaelis-Menten type. Mathematically, this mechanism includes the former as a subcase. PMID- 11121823 TI - CME information. PMID- 11121824 TI - Re-evaluating the prevalence of and diagnostic composition within the broad clinical spectrum of bipolar disorders. AB - Until recently it was believed that no more than 1% of the general population has bipolar disorder. Emerging transatlantic data are beginning to provide converging evidence for a higher prevalence of up to at least 5%. Manic states, even those with mood-incongruent features, as well as mixed (dysphoric) mania, are now formally included in both ICD-10 and DSM-IV. Mixed states occur in an average of 40% of bipolar patients over a lifetime; current evidence supports a broader definition of mixed states consisting of full-blown mania with two or more concomitant depressive symptoms. The largest increase in prevalence rates, however, is accounted for by 'softer' clinical expressions of bipolarity situated between the extremes of full-blown bipolar disorder where the person has at least one manic episode (bipolar I) and strictly defined unipolar major depressive disorder without personal or family history for excited periods. Bipolar II is the prototype for these intermediary conditions with major depressions and history of spontaneous hypomanic episodes; current evidence indicates that most hypomanias pursue a recurrent course and that their usual duration is 1-3 days, falling below the arbitrary 4-day cutoff required in DSM-IV. Depressions with antidepressant-associated hypomania (sometimes referred to as bipolar III) also appear, on the basis of extensive international research neglected by both ICD-10 and DSM-IV, to belong to the clinical spectrum of bipolar disorders. Broadly defined, the bipolar spectrum in studies conducted during the last decade accounts for 30-55% of all major depressions. Rapid-cycling, defined as alternation of depressive and excited (at least four per year), more often arise from a bipolar II than a bipolar I baseline; such cycling does not in the main appear to be a distinct clinical subtype - but rather a transient complication in 20% in the long-term course of bipolar disorder. Major depressions superimposed on cyclothymic oscillations represent a more severe variant of bipolar II, often mistaken for borderline or other personality disorders in the dramatic cluster. Moreover, atypical depressive features with reversed vegetative signs, anxiety states, as well as alcohol and substance abuse comorbidity, is common in these and other bipolar patients. The proper recognition of the entire clinical spectrum of bipolarity behind such 'masks' has important implications for psychiatric research and practice. Conditions which require further investigation include: (1) major depressive episodes where hyperthymic traits - lifelong hypomanic features without discrete hypomanic episodes - dominate the intermorbid or premorbid phases; and (2) depressive mixed states consisting of few hypomanic symptoms (i.e., racing thoughts, sexual arousal) during full-blown major depressive episodes - included in Kraepelin's schema of mixed states, but excluded by DSM-IV. These do not exhaust all potential diagnostic entities for possible inclusion in the clinical spectrum of bipolar disorders: the present review did not consider cyclic, seasonal, irritable-dysphoric or otherwise impulse-ridden, intermittently explosive or agitated psychiatric conditions for which the bipolar connection is less established. The concept of bipolar spectrum as used herein denotes overlapping clinical expressions, without necessarily implying underlying genetic homogeneity. In the course of the illness of the same patient, one often observes the varied manifestations described above - whether they be formal diagnostic categories or those which have remained outside the official nosology. Some form of life charting of illness with colored graphic representation of episodes, stressors, and treatments received can be used to document the uniquely varied course characteristic of each patient, thereby greatly enhancing clinical evaluation. PMID- 11121825 TI - A review of randomized, controlled clinical trials in acute mania. AB - This review considers the evidence supporting the use of somatic therapies (medications and electroconvulsive therapy) in the treatment of acute mania associated with bipolar disorder. Data from randomized, controlled clinical trials have established the efficacy of lithium, divalproex sodium, and carbamazepine in the treatment of acute mania. The use of combinations of mood stabilizers in the treatment of acute mania has not been well examined in controlled trials. Conventional antipsychotics and some atypical antipsychotics are frequently used as initial or adjunctive treatment. Similarly, benzodiazepines are frequently used as adjunctive agents. Preliminary data suggest that some calcium channel blockers and several anticonvulsants, e.g., lamotrigine, gabapentin, and topiramate, may have therapeutic value in the treatment of acute mania. In contrast, electroconvulsive therapy is generally accepted as being highly effective despite the lack of controlled evidence. PMID- 11121826 TI - Pharmacotherapy of depression and mixed states in bipolar disorder. AB - The treatment of bipolar depression requires the resolution of depression and the establishment of mood stability. A basic problem is that the treatments used in treating bipolar depression were developed and proven effective for other disease states: antidepressants for unipolar depression, and mood stabilizers for mania. The panel addressed four unresolved questions regarding depression in relation to bipolar disorder: (1) the relative effectiveness of different antidepressant treatments; (2) the relative likelihood of mood destabilization with different antidepressant treatments; (3) the effectiveness and role of mood-stabilizing medicines as antidepressants; and (4) the optimal approach to mixed states. The selection of an antidepressant depends both on its relative lack of mania- or hypomania-provoking potential and on its effectiveness against bipolar depression. There is little definitive evidence distinguishing effectiveness of the major groups of antidepressive agents, so side-effect profiles and pharmacokinetics are major considerations. The underlying bipolar disorder should be treated with mood stabilizers started simultaneously with any antidepressive treatments. Lithium, divalproex sodium and carbamazepine have all been found to be helpful, to some extent, in treating bipolar depressive episodes as well as for long-term mood stabilization. There is little evidence for long-term benefits of antidepressive agents in bipolar disorder, and some evidence that they may destabilize the disorder. Therefore, in contrast to the long-term use of mood stabilizers, antidepressant use is recommended on a temporary basis. The duration of antidepressant treatment is determined by past history in terms of liability for mood destabilization, and by the ability of the patient to tolerate gradual antidepressant discontinuation without return of depression. Mixed states, where symptoms of depression and mania coexist, are regarded as a predictor of relatively poor response to lithium, and divalproex has been found to be more effective. Carbamazepine may too be useful in mixed states. Most patients with mixed states in actual practice require combinations of mood stabilizers, though there is little controlled data regarding such co-prescription, especially from a long-term perspective. PMID- 11121827 TI - Maintenance therapies for classic and other forms of bipolar disorder. AB - The progressive, episodic and chronic nature of bipolar disorder dictates the need for lifelong pharmacological maintenance treatment in the majority of patients. Prophylaxis should be considered after a single episode of severe mania or after more than one episode of hypomania in bipolar II disorder, although some clinicians now consider an episode of either sufficient to warrant maintenance therapy. Lithium is efficacious as maintenance therapy, but is not as highly effective as early studies initially suggested (abrupt discontinuation of lithium probably increased placebo relapse figures). Rates of premature discontinuation of lithium are high. Divalproex sodium is used frequently in the USA and Canada for long-term treatment for bipolar disorder but an insufficient number of controlled trials have been published to assess adequately its role. Carbamazepine is also employed in maintenance treatment. Randomized studies indicate it is superior to placebo but somewhat less effective than lithium. Augmentation of any of these drugs with another mood stabilizer, an antipsychotic, or electroconvulsive therapy appears to be effective, although there are few controlled studies. Design issues that need consideration in order to achieve meaningful data are discussed. A severe manifestation of bipolar disorder is rapid cycling. It is often induced by antidepressants, although this association frequently goes unrecognized. Patients with a rapid cycling course of illness are difficult to treat effectively. Although rapid cycling is often associated with poor response to lithium, there have been no randomized, controlled treatment studies. Based on open studies and expert panel recommendations, the International Exchange on Bipolar Disorder (IEBD) recommended initial treatment with divalproex sodium, with subsequent addition of other mood stabilizers, antipsychotics or thyroid supplementation as necessary. Combination treatments are frequently required for optimal response in these patients. PMID- 11121828 TI - Current issues in the identification and management of bipolar spectrum disorders in 'special populations'. AB - Bipolar disorder is a common, lifelong condition that can present during childhood, adolescence, adulthood or later in life. It may occur alone but, more frequently, is complicated by comorbid psychiatric and medical disorders. As such, bipolar disorder presents in many different special populations, each of which warrants specific considerations of diagnosis, treatment and management. This review summarizes common issues concerning recognition of bipolar disorder, particularly in younger patients, discusses the prevalence and treatment of anxious disorder and addictive comorbidity, and considers bipolar disorder in the institutionalized and forensic populations. Treatment options and the supporting evidence are discussed. PMID- 11121830 TI - Posttest. PMID- 11121829 TI - First International Exchange on Bipolar Disorder. PMID- 11121832 TI - Fatal attractors: theoretical approaches to tumor differentiation. PMID- 11121831 TI - New potent and selective human adenosine A(3) receptor antagonists. PMID- 11121833 TI - A new enzyme: insights into mechanisms of cell trafficking. PMID- 11121834 TI - The true nature of pharmacogenomic associations? PMID- 11121835 TI - Pharmacological chaperones: a new twist on receptor folding. AB - Protein misfolding is at the root of several genetic human diseases. These diseases do not stem from mutations within the active domain of the proteins, but from mutations that disrupt their three-dimensional conformation, which leads to their intracellular retention by the quality control apparatus of the cell. Facilitating the escape of the mutant proteins from the quality control system by lowering the temperature of the cells or by adding chemicals that assist folding (chemical chaperones) can result in proteins that are fully functional despite their mutation. The discovery that ligands with pharmacological selectivity (pharmacological chaperones) can rescue the proper targeting and function of misfolded proteins, including receptors, might help to develop new treatments for 'conformational diseases'. PMID- 11121836 TI - PPAR-gamma agonists: therapeutic role in diabetes, inflammation and cancer. AB - The recent development of a novel class of insulin-sensitizing drugs, the thiazolidinediones (TZDs), represents a significant advance in antidiabetic therapy. One key mechanism by which these drugs exert their effects is by activation of the peroxisome proliferator activated receptor gamma (PPAR-gamma), a member of the nuclear receptor family. Evidence supporting this mechanism of action of the TZDs will be reviewed in this article. Recent data suggests that PPAR-gamma agonists might also have therapeutic potential in the treatment of inflammatory diseases and certain cancers. PMID- 11121838 TI - Pharmacotherapy for erectile dysfunction. AB - Erectile dysfunction (ED) is defined as the consistent inability to obtain or maintain an erection for satisfactory sexual relations. An estimated 20-30 million men suffer from some degree of sexual dysfunction. The past 20 years of research on erectile physiology have increased our understanding of the biochemical factors and intracellular mechanisms responsible for corpus cavernosal smooth muscle contraction and relaxation, and revealed that ED is predominantly a disease of vascular origin. Since the advent of sildenafil (Viagra), there has been a resurgence of interest in ED, and an increase in patients presenting with this disease. A thorough knowledge of the physiology of erection is essential for future pharmacological innovations in the field of male ED. PMID- 11121837 TI - Novel approaches to targeting neuropeptide systems. AB - Generation and/or interruption of cell signalling by neuropeptides has been shown to be essentially, although not exclusively, mediated by one or several membrane bound enzymes, giving rise to the concept of selective versus dual enzyme inhibitors. Because most of these enzymes are zinc metallopeptidases, novel inhibitors are now being designed based on the structure of these proteins. The physiological role of neuropeptides and their relationships with other peptide systems can be investigated by comparing results obtained using peptidase inhibitors and selective receptor antagonists with those obtained using mice in which genes encoding the various components of a peptide system have been deleted. The potential use of peptidase inhibitors, compared with exogenous agonists, as therapeutic agents (particularly as analgesics or antidepressants) and their use in the investigation of the neurobiology of drug abuse will be discussed with particular focus on enkephalins and cholecystokinin 8 (CCK-8). PMID- 11121839 TI - Src inhibitors: drugs for the treatment of osteoporosis, cancer or both? AB - Src was one of the first proto-oncogenes to be identified and is a prototype of non-receptor type tyrosine kinases. The role of Src in bone metabolism first became apparent in Src-deficient mice and has been confirmed using low-molecular weight Src inhibitors in animal models of osteoporosis. At the cellular level, it is well established that Src plays an important role in proliferation, and adhesion and motility. In addition, recent data indicate an involvement of Src in cell survival and intracellular trafficking in various specialized cell types. These new findings suggest that Src inhibitors might have therapeutic value in the suppression of tumor growth, tumor angiogenesis and bone resorption. PMID- 11121840 TI - Erratum. PMID- 11121842 TI - Why veterinarians should care more about parasitology. PMID- 11121841 TI - What's that buzz? Mosquitoes and fruit flies commingle. PMID- 11121844 TI - Neospora caninum in cattle. M. McAllister Et al. (2000) evidence of point-source exposure to neospora caninum and protective immunity in a herd of beef cows. J. Am. Vet. Med. Assoc. 217, 881-887 PMID- 11121843 TI - In search of the red queen. PMID- 11121845 TI - A Co-infection model of malaria and hepatitis B virus. Pasquetto, V. et al. (2000) host-virus interactions during malaria infection in hepatitis B virus transgenic mice. J. Exp. Med. 192, 529-536 PMID- 11121846 TI - Whales, worm weight and mice on the Web PMID- 11121847 TI - Why model malaria? AB - The past 30 years have seen little tangible progress in alleviating the worldwide burden of malaria. Ellis McKenzie here discusses some of the history, problems and prospects of mathematical models of malaria, and the contributions that models might make towards progress. He argues that models can be powerful tools for integrating information from different disciplines, and that advances in computer modeling can complement and extend classic approaches. PMID- 11121848 TI - Hexose transport in asexual stages of Plasmodium falciparum and kinetoplastidae. AB - The hexose sugar, glucose, is a vital energy source for most organisms and an essential nutrient for asexual stages of Plasmodium falciparum. Kinetoplastid organisms (e.g. Trypanosoma and Leishmania spp) also require glucose at certain critical stages of their life cycles. Although phylogenetically unrelated, these organisms share many common challenges during the mammalian stages of a parasitic life cycle, and possess hexose uptake mechanisms that are amenable to study using similar methods. Defining hexose permeation pathways into parasites might expose an Achilles' heel at which both antidisease and antiparasite measures can be aimed. Understanding the mode of entry of glucose also presents a good general model for substrate acquisition in multicompartment systems. In this review, Sanjeev Krishna and colleagues summarize current understanding of hexose transport processes in P. falciparum and provide a comparison with data obtained from kinetoplastids. PMID- 11121849 TI - Reaching maturity - 25 years of the TDR. AB - In its first 25 years of existence, TDR has become a key player in the development of new tools for the control of tropical diseases and the training of researchers from disease-endemic countries. In order to maintain its leading position, cope with new health challenges and profit from new avenues opened by science and technology breakthroughs, a new strategic vision is now being implemented. It aims at a closer interaction with health systems and disease control programmes, capacity strengthening based on selected research initiatives and full exploitation of scientific and technological advances in the biomedical, social and information sciences, as discussed here by Carlos Morel. PMID- 11121850 TI - The interface between epidemiology and population genetics. AB - Modern biology increasingly integrates disparate disciplines. Here, Steve Paterson and Mark Viney examine the interface between epidemiology and population genetics. They argue that infection and inheritance can be considered as analogous processes, and that epidemiology and population genetics share many common features. They consider the potential for existing population genetic theory to dissect epidemiological patterns in field studies and they consider other relationships between genetics and epidemiology that provide a research challenge for the future. PMID- 11121851 TI - Are there pros as well as cons to being parasitized? AB - The diversity of ways in which parasites reduce the fitness of their hosts has been documented during the past decades, and clearly indicates that parasites can often be considered as direct agents of selection. In natural systems, however, the outcome of a host-parasite interaction might be strongly determined by other ecological factors. Parasites can be detrimental to host fitness in one environment, whereas they can be beneficial to it in another. From an evolutionary perspective, this phenomenon is of considerable importance for understanding the dynamics of coevolution among geographically structured populations evolving under different ecological pressures. Here, Frederic Thomas and colleagues review several ecological situations in which parasitized individuals enjoy a selective advantage over unparasitized conspecifics. PMID- 11121852 TI - Molecular crosstalk in host-parasite relationships: schistosome- and leech-host interactions. AB - The host-parasite relationship is based on subtle interplay between parasite survival strategies and host defense mechanisms. In this context, parasites often use the same or similar immune signaling molecules and/or molecular mimicry to escape host immunosurveillance. Both processes represent an adaptive strategy to ensure host immunocompatibility. This bidirectional communication between parasites and their hosts includes the renin-angiotensin, opioid and opiate systems. Here, Michel Salzet, Andre Capron and George Stefano review recent work on the interaction of common signaling mechanisms in schistosomes, leeches and their host. PMID- 11121853 TI - Geographical variation in Ascaris lumbricoides fecundity and its implications for helminth control. AB - The observation by microscopy of nematode eggs in human faeces is used to diagnose a helminthic infection, while the concentration of those eggs is used to estimate the number of worms in the host. Within a community, the prevalence of infection and the mean egg count provide useful information about the extent of a public health problem, and are being used to guide the growing efforts to control disease caused by helminths. Here, Andrew Hall and Celia Holland examine data on the relationship between the worm burdens of Ascaris lumbricoides and the concentration of eggs in faeces, and discuss the implications of the variation found for using such data to plan helminth control programmes. PMID- 11121854 TI - Pathogenesis of lymphatic disease in bancroftian filariasis: a clinical perspective. AB - The pathogenesis of lymphatic filariasis has been a matter of debate for many decades. Here, Gerusa Dreyer and colleagues propose a dynamic model of bancroftian filariasis, integrating clinical, parasitological, surgical, therapeutic, ultrasonographic and histopathological data. This model has profound implications for filariasis control programs and the management of the individual patient. PMID- 11121855 TI - Automated malaria diagnosis using pigment detection. AB - Several new methods of malaria diagnosis have recently been developed, but these all rely on clinical suspicion and, consequently, an explicit clinical request. Although some methods lend themselves to automation (eg. PCR), no technique can yet be used for routine clinical automated screening. Detection of birefringent haemozoin has been used to diagnose malaria since the turn of the 20th century. A new generation of full blood count analysers, used widely in clinical laboratories, have the potential to detect haemozoin in white blood cells and probably erythrocytes. Thomas Hanscheid, Emilia Valadas and Martin Grobusch here describe this novel technique for malaria diagnosis and discuss its potential applications. PMID- 11121856 TI - Editorial. PMID- 11121857 TI - Cyclin D1 in breast premalignancy and early breast cancer: implications for prevention and treatment. PMID- 11121858 TI - Investigation of the enhancement of N-nitrosomethylbenzylamine-induced esophageal tumorigenesis by 6-phenylhexyl isothiocyanate. AB - Previous studies in our laboratory have shown that 6-phenylhexyl isothiocyanate (PHITC), enhances N-nitrosomethylbenzylamine (NMBA)-induced esophageal tumorigenesis in F344 rats while the shorter chain analogs, phenylethyl isothiocyanate (PEITC), and 3-phenylpropyl isothiocyanate (PPITC), inhibit NMBA induced esophageal tumorigenesis. To test the hypothesis that PHITC influences the promotional stage of esophageal tumorigenesis, groups of 22-27 rats were dosed with vehicle or NMBA three times a week for 5 week, and fed a modified AIN 76A diet containing PHITC at concentrations of 0.0, 1.0, and 2.5 micromol/g. At the 25th week, the rats were killed, esophagi harvested and tumors counted. In the groups that received NMBA+PHITC, apparent but statistically insignificant increases in tumor multiplicity of 32 and 42% were found in comparison to rats treated with NMBA alone. A higher frequency of dysplastic lesions was found in rats treated with NMBA+2.5 micromol/g PHITC (71%) when compared to rats treated with NMBA only (12%). To test whether PHITC increased cellular proliferation, we evaluated proliferating cell nuclear antigen (PCNA) expression by immunohistochemistry. While there were no significant increases in PCNA staining in rats treated with NMBA+PHITC compared to rats treated with NMBA only, rats treated with PHITC only had a significantly higher PCNA index compared to untreated controls. Expression of cyclin D1, another biomarker of proliferation, was analyzed by semi-quantitative reverse transcription-polymerase chain reaction. There were no significant increases in cyclin D1 expression in groups treated with NMBA+PHITC compared to the group treated with NMBA only. Thus, while the data suggest a promotional effect by PHITC as manifested by a significant increase in dysplastic leukoplakia by the high dose of PHITC and an increase in the PCNA index by PHITC alone, PHITC does not appear to have a significant effect on esophageal cell proliferation. PMID- 11121859 TI - Urine GABA levels in ovarian cancer patients: elevated GABA in malignancy. AB - The association of malignancy with elevated diamine oxidase (DAO), an enzyme producing gamma-aminobutyric acid (GABA) is well documented. In ovarian cancer, increased DAO occurs in the malignant tissues and plasma. Since higher DAO levels cause GABA accumulation, elevated GABA may occur in ovarian cancer and be reflected in urine. We tested this hypothesis and found that half the ovarian cancer patients had a clearly elevated urine GABA, a result that is in agreement with previous reports on DAO and malignancy. The published findings on DAO, GABA and malignancy suggest that elevated GABA is associated with cancer. This proposal could lead to a GABA urine monitor or new directions in cancer treatment or research. PMID- 11121860 TI - Effects of heterocyclic amines with mammary gland carcinogenic potential on estrogenic response of uterus in ovariectomized rats. AB - Heterocyclic amines (HCAs) present in cooked foods are suggested to be involved in human breast cancer development. Estrogen plays a pivotal role in mammary gland carcinogenesis. Therefore, we designed an in vivo experiment to investigate potential estrogenic effects of two HCAs, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), which induce mammary gland cancers in rodents, on the uterus of ovariectomized (OVX) Sprague Dawley (SD) rats. Female SD rats ovariectomized at 35 days of age were given intraperitoneal injections of 17beta-estradiol (E2) at doses of 0, 30 or 50 microg/kg or one of the HCAs at a dose of 50 mg/kg b.w. once a day at 47, 48, and 49 days of age. E2 dramatically increased uterine weights, stromal thickness, epithelial cell height, and 5-bromo-2'-deoxyuridine (BrdU) positive cell counts in a dose dependent manner. Intraperitoneal administration of PhIP or IQ, in contrast, did not produce any estrogenic responses in this assay system. These results indicate that the carcinogenicities of these two HCAs in mammary glands are not associated with estrogenic potential. PMID- 11121861 TI - Cyclophosphamide and 5-fluorouracil act synergistically in ovarian clear cell adenocarcinoma cells. AB - Chemosensitivity to the drugs plays a crucial role in the treatment of ovarian cancer. In this study, we evaluate the cytotoxicity of chemotherapeutic agents in six ovarian cancer cell lines; four clear cell adenocarcinoma and two serous papillary adenocarcinoma, using seven single drugs and seven sets of drug combinations with tetrazolium-based semiautomated colorimetric (MTT) assay. The drug concentration which produced 50% growth inhibition (IC50) of cisplatin was within clinically achievable range in five cell lines. The area under the curve (AUC) at IC50 of cyclophosphamide was below the clinically achievable AUC in two serous papillary cell lines. Paclitaxel was more effective in clear cells than serous papillary cells. The intensification of cytotoxicity was observed in the combinations of paclitaxel and cisplatin, and cyclophosphamide and cisplatin or 5 fluorouracil irrespective of histopathological characteristics of the original tumor. Our results indicate that ovarian cancer cell lines respond to chemotherapeutic agents heterogeneously depending upon histopathological features, indicating individualized regimens may improve survival in ovarian cancer patients. PMID- 11121862 TI - Targeting and anti-tumor efficacy of liposomal 5'-O-dipalmitoylphosphatidyl 2'-C cyano-2'-deoxy-1-beta-D-arabino-pentofuranosylcytosine in mice lung bearing B16BL6 melanoma. AB - 2'-C-cyano-2'-deoxy-1-beta-D-arabino-pentofuranosylcytosine (CNDAC) is a potent anti-cancer agent, and we previously observed that liposomal formulation of 5'-O dipalmitoylphosphatidyl derivative of CNDAC (DPP-CNDAC) is desirable for targeting. For targeting to pulmonary cancer, we investigated the in vivo behavior of liposomes containing DPP-CNDAC by a non-invasive method using positron emission tomography. Liposomes composed of DPP-CNDAC and cholesterol (DPP-CNDAC/CH liposomes) were markedly accumulated in mice lung bearing B16BL6 melanoma. In metastatic pulmonary cancer model, DPP-CNDAC/CH liposomes significantly reduced the lung colonization in a dose-dependent manner. The activity was significantly superior to conventional liposomal formulation or soluble CNDAC. These results suggest that DPP-CNDAC/CH liposomes are useful for metastatic pulmonary cancer. PMID- 11121863 TI - Simian virus 40 is present in human lymphomas and normal blood. AB - Many independent studies have demonstrated Simian virus 40 (SV40) in normal and neoplastic human tissues. Clonal integration of virus in the DNA of several thyroid and bone tumors suggests a direct role for SV40 in some cancers. However, in most cases the role of SV40 remains unclear. This study determined the presence of SV40, by amplification followed by hybridization, in 266 normal and neoplastic blood and lymphoid samples. Amplification detected SV40 in 14% of non autoimmune deficiency syndrome (AIDS) lymphomas, 28% of AIDS related lymphoma and 16% of peripheral blood lymphocytes from non-cancerous patients. No SV40 was detected in leukemia samples. Direct Southern blotting of SV40+ samples detected no virus, consistent with less than one viral genome in ten cells. Sequence analysis of SV40 in blood and lymphoid samples found sequences distinct from laboratory strains of SV40. The presence of limited quantities of SV40 in a small proportion of both normal and neoplastic tissues is suggestive of an adventitious presence with no apparent direct role in blood and lymphoid cancers. PMID- 11121865 TI - Improvement of gene transfer to cervical cancer cell lines using non-viral agents. AB - Virus-like particles (VLPs) composed of recombinant capsid protein L1 and L2 of human papillomavirus type 16 were conjugated with polylysine (PL) and gene transfer was performed using VLP-PL conjugates to allow the expression of targeted gene. When HeLa cells were incubated with VLP-PL conjugate coupled with plasmid cytomegalovirus beta-galactosidase (pCMVbeta-gal), about 10% of cells were transfected and demonstrated beta-galactosidase activity. Hence chloramphenicol acetyltransferase activity was also expressed significantly in VLP-PL-plasmid simian virus 2 chloramphenicol acetyl transferase (pSV2CAT) transfected cells, VLP-PL conjugate was tested whether it could transfer a tumor suppressor gene, pCMVp53, to HeLa cells and the exogenously provided p53 gene complexed to VLP-PL conjugate was detected from HeLa cells by polymerase chain reaction (PCR) analysis. Interestingly, additional increase of transfection efficiency was demonstrated in the presence of poloxamer 407 when C-33A cells were transfected with VLP-PL-pCMVbeta-gal complex. The result support the notion that VLP-PL conjugate may be a promising vector to transfer genetic materials into cancer cells and poloxamer 407 can be used for enhancing the transfection efficiency of VLP-PL conjugate. PMID- 11121864 TI - Quantitative relationship of the circulating tumor burden assessed by reverse transcription-polymerase chain reaction for cytokeratin 19 mRNA in peripheral blood of colorectal cancer patients with Dukes' stage, serum carcinoembryonic antigen level and tumor progression. AB - We prospectively analyzed the circulating tumor burden in colorectal cancer patients using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) for carcinoembryonic antigen (CEA) and cytokeratin 19 (CK19 ). We distinguished the mRNA levels in peripheral blood between 33 patients and 26 healthy controls with reference to SK-BR-3 cell line. We found CEA-mRNA in 88% of patients and 92% of controls, and CK19 mRNA in 64% of patients and 19% of controls. Our CK19 mRNA assay was sufficiently sensitive to detect one SK-BR-3 cell among 10(6) normal blood cells. The upper limit of CK19 mRNA among controls was exceeded by 14 patients, and 12 patients (86%) developed systemic metastases/recurrence. Significantly elevated CK19 mRNA levels appeared to originate from circulating malignant cells (P<0.0001). Of relevance, the CK19 mRNA level increased with advancing Dukes' stage and correlated directly with the serum CEA level (P=0.016). CK19 mRNA quantification may prove valuable for cancer staging and disease monitoring. PMID- 11121866 TI - Bombesin and gastrin releasing peptide increase tyrosine phosphorylation of focal adhesion kinase and paxillin in non-small cell lung cancer cells. AB - The effects of some oncogenes, growth factors and neuropeptides are mediated by tyrosine phosphorylation of focal adhesion kinase (p125(FAK)) and paxillin cytoskeletal proteins. In this study the ability of bombesin/gastrin releasing peptide (BB/GRP) to stimulate tyrosine phosphorylation of p125(FAK) and paxillin in non-small cell lung cancer (NSCLC) H1299 cells was investigated. BB, 100 nM caused increased p125(FAK) and paxillin tyrosine phosphorylation maximally after 1 min. The effect of BB on p125(FAK) and paxillin tyrosine phosphorylation was concentration-dependent, being half maximal at 4-8 nM. Also, 100 nM GRP, GRP(14 27) but not GRP(1-16) increased p125(FAK) and paxillin tyrosine phosphorylation indicating that the C-terminal of GRP is essential. BW2258U89, a GRP receptor antagonist, caused a dose-dependent inhibition of BB-stimulated p125(FAK) and paxillin tyrosine phosphorylation with an IC50 value of 3 microM. Cytochalasin D (0.3 microM), which inhibits actin polymerization, reduced the ability of BB to stimulate tyrosine phosphorylation of p125(FAK) and paxillin. Genistein (50 microM) and H-7 (50 microM), which are kinase inhibitors, reduced the tyrosine phosphorylation of p125(FAK) and paxillin stimulated by BB. Also, treatment of NCI-H1299 cells with FAK antisense resulted in decreased FAK tyrosine kinase activity and proliferation. These results suggest that p125(FAK) is an important enzyme for NSCLC proliferation. PMID- 11121867 TI - Combined analysis of MIB-1 and thyroid transcription factor-1 predicts survival in non-small cell lung carcinomas. AB - The prognostic value of combined immunohistochemical analysis for the thyroid transcription factor-1 (TTF-1) and the proliferation marker MIB-1 was assessed in a consecutive series of non-small cell lung carcinomas (NSCLC). Tumor immunoreactivity for TTF-1 and MIB-1 was classified in three groups (-,+,++) and in two groups (-,+), respectively. Comparison across groups for TTF-1 reactivity showed significantly different survival curves (P=0.04). In particular, the best prognosis was associated with a TTF-1 negative pattern, whereas the TTF-1 '++' cases showed the worst prognosis. A trend towards better prognosis was observed for MIB-1 negative cases (P=0.09). Multivariate analysis confirmed independent prognostic significance of TTF-1 (P=0.002), MIB-1 (P=0.01) and pStage (P=0.04). Accordingly, analysing TTF-1 and MIB-1 together, a better prediction of survival was obtained (P=0.02), with the poorest prognosis for the 'TTF-1++/MIB-1+' cases. PMID- 11121868 TI - Coexpression of granulocyte colony stimulating factor and its receptor in primary ovarian carcinomas. AB - Immunohistochemistry was used to determine the expression of granulocyte colony stimulating factor (G-CSF) and its receptor (G-CSFR) in primary ovarian carcinomas. The expression of G-CSFR was observed in the malignant cells of each of the 46 primary carcinomas examined; G-CSF was coexpressed in both the malignant epithelial cells and the stroma of 56.5% of the specimens. Thus the majority of ovarian carcinomas harbor both potential autocrine and paracrine G CSF axes. In 37% of the samples, G-CSF was expressed only within stromal cells, suggesting that only a potential paracrine system is in place. In a preliminary, retrospective, evaluation, the survival of patients whose tumors expressed only the apparent paracrine loop was significantly worse than patients whose tumors expressed both potential autocrine and paracrine G-CSF-based regulatory loops (14.5 vs. 42.5 months, respectively). Studies on the potential function of G-CSF were performed using the G-CSFR-expressing OVCAR-3 ovarian carcinoma line. As a single agent, rhG-CSF failed to stimulate [3H]-thymidine incorporation in these cells, but enhanced the mitogenic action of epidermal growth factor (EGF) in a dose-dependent manner. Thus, potential autocrine and/or paracrine loops involving G-CSF and its receptor occur in over 90% of primary ovarian carcinomas, and may act to modulate the action of growth factors. PMID- 11121869 TI - Tamoxifen mutagenesis and carcinogenesis in livers of lambda/lacI transgenic rats: selective influence of phenobarbital promotion. AB - Administration of tamoxifen (TAM) (20 mg/kg per day p.o.) for 6 weeks to female lambda/lacI transgenic rats caused a 4-fold increase in mutation frequency (MF) at the lacI gene locus in the livers of dosed animals compared with controls. After cessation of dosing, the MF showed a further increase with time at 2, 12 and 24 weeks, respectively. Phenobarbital promotion of similarly treated animals resulted in no increase in mutation frequency compared with TAM alone. Treatment with phenobarbital or TAM+phenobarbital resulted in time-dependent increases in liver weight compared with the corresponding controls. There was an increase in cell proliferation in the phenobarbital and TAM+phenobarbital groups, and at 24 weeks in the TAM dosed animals compared with controls. There was also a progressive increase in the number of GST-P expressing foci in the livers of TAM and TAM + phenobarbital rats compared with controls. The induction of cell proliferation and GSTP foci in the rat liver by phenobarbital is consistent with its ability to promote tamoxifen-initiated liver tumours in the rat. If the lacI gene is regarded as being representative of the rat genome in general (albeit that the gene is bacterial) the above observations suggest that promotion by tamoxifen confers selective advantage on mutated genes at loci that contribute to the tumour phenotype and that promotion of rat liver tumours by tamoxifen is not dependent simply upon the enhancement of cellular proliferation. PMID- 11121870 TI - Chromosomal mapping of human genes by radioactive hybridization of cDNAs to CEPH YAC high density gridded filter sets. AB - Chromosomal assignment of human transcribed sequences has been done mainly by high throughput genome analysis in specialized genome centres and, in a more classical fashion, by fluorescence in-situ hybridization (FISH) analysis. Not every laboratory has the ability to map cDNAs by FISH analysis. We here report a rapid mapping approach that is based on the hybridization of cDNA probes to high density gridded CEPH-YAC filters followed by subsequent computational analysis by database searches in the internet. Not only transcribed sequences but also genomic DNA could be subjected to this mapping approach. The presented approach allows to map human transcribed and genomic DNAs within 1-3 days and with a high level of resolution that will constantly increase in line with the incorporation of data deriving from high throughput genome mapping. PMID- 11121871 TI - Buthus martensi Karsch agonist of skeletal-muscle RyR-1, a scorpion active polypeptide: antinociceptive effect on rat peripheral nervous system and spinal cord, and inhibition of voltage-gated Na(+) currents in dorsal root ganglion neurons. AB - The antinociceptive effect and potential antinociceptive mechanism of Buthus martensi Karsch agonist of skeletal-muscle RyR-1 (BmK AS-1), a scorpion venom derived neurotoxic polypeptide, have been investigated in rats. The results show that: (a) the withdrawal latency to rat plantar radiant heat was increased significantly by 100 and 150% after intrathecal injection of 0.6 and 1.2 microg doses; (b) C components of rat nociceptive flexion reflex were reduced to 72, 50 and 29% after intraplantar injection of 5, 10 and 20 microg doses; (c) both central (spinal cord) and peripheral antinociceptive effects of BmK AS-1 could not be reversed by naloxone; (d) tetrodotoxin-resistant (TTX-R) Na(+) current was depressed to 83.87+/-1.64, 64.73+/-5.43 and 15.85+/-17.63%, and tetrodotoxin sensitive (TTX-S) Na(+) current was depressed to about 81.27+/-2.5, 49.08+/-8.09 and 9.03+/-12.34% with 0.2, 1.0 and 10 microg/ml BmK AS-1 measured using patch clamp recording in rat small dorsal root ganglion (DRG) neurons, respectively. The results indicate that BmK AS-1 may be a new component with potent antinociceptive activity mediated by modulating TTX-S and TTX-R Na(+) channels. PMID- 11121872 TI - Neonatal exposure to monosodium glutamate induces cell death and dendritic hypotrophy in rat prefrontocortical pyramidal neurons. AB - Monosodium glutamate was administered subcutaneously to male neonate rats, and the effects on cell number and cytoarchitecture of third-layer pyramidal neurons from the prefrontal cerebral cortex were studied in the adult. Monosodium glutamate treatment (4 mg/g of body weight, on post-natal days 1, 3, 5 and 7) resulted in fewer neurons, and shorter and less ramified dendritic processes, than those observed in control animals. Both density and proportional shapes of dendritic spines were not modified. We propose a dual effect of neonatal exposure to glutamate: an excitotoxic effect leading to cell death, and; a secondary neuroprotective effect, arising from the proliferation of glial cells and their subsequent uptake of glutamate, that favors the survival of the remaining neurons, and leads to a further hypotrophic effect on their dendritic processes. PMID- 11121873 TI - Impairment of maze learning in rats by restricting environmental space. AB - We previously reported that the restriction of environmental space attenuates spontaneous locomotor activity and hippocampal acetylcholine release. To examine the effect of the restriction of environmental space on spatial learning function, male rats were individually housed in a cylindrical large cage (diameter=35 cm) or small cage (diameter=19 cm) for 5 days. Eight-arm radial maze performance was examined to evaluate spatial learning and memory functions. The task was performed once a day between 21:00 and 22:00 h in the dark phase. Although all rats learned and performed the task, those in the small cage had lower scores and took more trial time than those in the large cage. These results suggest that the restriction of environmental space impairs spatial learning in the dark phase in rats. PMID- 11121874 TI - Choroid plexus epithelial cells in adult rats show structural alteration but not apoptosis following an exposure to hypobaric hypoxia. AB - The choroid plexus in adult rats was examined for any structural alteration or apoptotic cell death following a high altitude exposure which leads to the development of hypobaric hypoxia due to reduced oxygen tension in the atmospheric air. Caspase-3 (a protease which mediates apoptosis) immunoreactivity was absent in the choroid plexus epithelial cells in the control rats and following altitude exposure; Bcl-2 (anti-apoptotic protein) and Bax (pro-apoptotic protein) immunoreactivity were upregulated at 3 h-2 days following the altitude exposure when compared to the controls but not in longer surviving rats. At the ultrastructural level, glycogen particles and vacuoles were observed in some epithelial cells at 7 days following the altitude exposure. It is suggested that transient exposure to high altitude may not cause much damage to the choroid plexus epithelial cells except for some structural alteration which may be due to altered metabolism of the cells in response to hypobaric hypoxia. PMID- 11121875 TI - Decreased neurocalcin immunoreactivity in sympathetic and parasympathetic neurons of the major pelvic ganglion in aged rats. AB - In the rat the majority of sympathetic and parasympathetic postganglionic neurons that innervate the pelvic viscera are located together in the major pelvic ganglia. We have ascertained that it is only the sympathetic population of this ganglion that exhibits age-associated attrition. Recent immunohistochemical investigations of the distribution of calcium binding proteins in this ganglion in young adult and aged rats have demonstrated that calbindin-D28k is only present in the sympathetic neurons and that the number of calbindin immunoreactive sympathetic neurons of the aged ganglion was dramatically reduced. In the present study we have investigated the distribution of neurocalcin (NC) alpha isoform in the major pelvic ganglion. In young adults 98.7% of sympathetic neurons (identified by anti-tyrosine hydroxylase immunostaining) and 98% of parasympathetic neurons (identified by anti-nitric oxide synthase immunostaining) contained NC immunoreactivity and these figures are reduced to 68 and 45.5% in the aged group. Thus, unlike calbindin-D28k, NC is not confined to the sympathetic neuron population in the major pelvic ganglion and decreases significantly in old age in both neuronal populations. The likely effects are to impair intracellular calcium-dependent signalling in neurons of the major pelvic ganglion, possibly compounding the effects of the previously reported decrease in calbindin-D28k in the sympathetic population. PMID- 11121876 TI - Age-related changes in potentiation of evoked responses in CA1 pyramidal cells from the hamster hippocampus. AB - Hippocampal responses were compared in 16 old (15-22 month) and 14 young (2-5 month) Syrian hamsters to determine if this species showed age-dependent changes in potentiation. Population spike amplitude increased following tetanus by 84.1+/ 20.0% in slices from young animals and by 51.1+/-6.3% in slices from old animals (P<0.05). In addition, I-O curves (plots of population spike amplitude vs. intensity of Schaffer collateral excitation) were obtained before and after tetanus. While regions of I-O curves near threshold and saturation showed no significant change, the slope at the midpoint of the I-O curve increased by 152.3+/-68.4% in slices from young animals and by 13.7 +/-10.0% in slices from old animals (P<0.05). Thus, in old hamsters (as in rats) potentiation was impaired and slope changes of I/O curves clearly displayed this deficit. PMID- 11121877 TI - High-frequency oscillations in human posterior tibial somatosensory evoked potentials are enhanced in patients with Parkinson's disease and multiple system atrophy. AB - High-frequency oscillations (HFOs) and the underlying P37 primary somatosensory response evoked by posterior tibial nerve stimulation were recorded in patients with Parkinson's disease (PD) and in those with multiple system atrophy (MSA), as well as in normal controls. In order to increase the signal-to-noise ratio, we averaged a large number of responses (9998 epochs) with a high sampling rate (20 kHz per channel). HFOs were extracted by filtering the wide band-pass recording of the P37 potential with a 600-900 Hz band-pass filter. High-amplitude HFOs were observed in both the PD and MSA patients. Furthermore, the duration of illness was positively correlated with the amplitude of HFOs. The results suggest that HFOs are enhanced by dysfunction of the basal ganglia. PMID- 11121878 TI - Vasoactive intestinal peptide-stimulated adenosine 3',5'-cyclic monophosphate formation in cerebral cortex and hypothalamus of chick and rat: comparison of the chicken and mammalian peptide. AB - Chicken and mammalian (human/porcine/rat) vasoactive intestinal peptides (VIP; 0.01-3 microM), whose structures differ by four amino acid residues in 11, 13, 26 and 28 positions, were compared with respect to their ability to stimulate adenosine 3',5'-cyclic monophosphate (cyclic AMP) formation in the hypothalamus and cerebral cortex of chick and rat. In four tested biological systems, the chicken VIP appeared to be significantly more potent in evoking cyclic AMP response than its mammalian counterpart, the differences were more pronounced in the chick tissues, particularly in the hypothalamus, where the mammalian peptide produced only weak (but significant) effect at the highest used dose, i.e. 3 microM. Pituitary adenylate cyclase-activating polypeptide, a VIP-like peptide, applied as a reference drug at 0.1 microM, strongly stimulated cyclic AMP formation in all tested systems. The data demonstrate significant quantitative differences in biological activity between mammalian and non-mammalian peptides tested in brain tissue of chicks and rats, indicating that usage of the mammalian VIP in at least 'avian' studies may lead to some false conclusions. PMID- 11121879 TI - Reduced neprilysin in high plaque areas of Alzheimer brain: a possible relationship to deficient degradation of beta-amyloid peptide. AB - Neprilysin is an enzyme capable of degrading beta-amyloid protein. We measured neprilysin mRNA and protein levels in brain and peripheral organs of Alzheimer disease (AD) and control cases. Neprilysin mRNA levels were lowest in the hippocampus and temporal gyrus, which are vulnerable to senile plaque development. They were highest in the caudate and peripheral organs which are resistant to senile plaque development. Levels in AD were significantly lower than controls in the hippocampus and midtemporal gyrus but not in other brain areas or peripheral organs. We also measured levels of the mRNA for the neuronal marker microtubule-associated protein-2. They were remarkably constant in all brain areas and were not lowered in AD, indicating that the neprilysin mRNA reduction in the hippocampus and temporal gyrus was not correlated with simple neuronal loss. Relative levels of neprilysin protein generally paralleled those of the mRNA. These results suggest that deficient degradation of beta-amyloid protein caused by low levels of neprilysin may contribute to AD pathogenesis. PMID- 11121880 TI - Brain function in a patient with torture related post-traumatic stress disorder before and after fluoxetine treatment: a positron emission tomography provocation study. AB - We report positron emission tomographic measurements of regional cerebral blood flow (rCBF) in a male patient with war and torture related post-traumatic stress disorder (PTSD) during symptom provocation. The subject was exposed to war related sounds before and after treatment with a selective serotonin reuptake inhibitor (SSRI; Fluoxetine; Fontex((R))). Therapy reduced PTSD symptoms, provoked anxiety and heart rate. Before treatment trauma reminders resulted in decreased rCBF in the insula, prefrontal, and inferior frontal cortices. Increased activity was evident in the cerebellum, precuneus and supplementary motor cortex. This was normalized after SSRI administration. Prefrontal and cingulate rCBF correlated with heart rate. Hence, the anxiolytic effect of SSRI for PTSD could be mediated by prefrontal and paralimbic cortices. Data suggest that SSRI treatment normalize provocation induced rCBF alterations in areas involved in memory, emotion, attention and motor-control. PMID- 11121881 TI - Immunohistochemical localization of plasminogen activator inhibitor-1 in rat and human brain tissues. AB - We investigated immunohistochemically the localization of type 1 plasminogen activator inhibitor (PAI-1) in rat and human brain tissues. In rat, neurons and astrocytes were stained positively for PAI-1 after colchicine treatment. In post mortem human brain, neurons were stained for PAI-1 but the number of positive neurons varied greatly from case to case. PAI-1 positive astrocytes occurred in the white matter lesions of some patients. In Alzheimer's disease, weak PAI-1 labeling was seen in association with senile plaques and ghost tangles. The present results support a notion that PAI-1 and its target proteases such as plasminogen activators and thrombin are involved in a variety of physiological and pathological processes in brain. PMID- 11121882 TI - Influence of smoking and gender on diurnal variations of heart rate reactivity in humans. AB - We evaluated the influence of smoking and gender on diurnal variations of heart rate reactivity during performance of two vigilance tasks (auditory and visual) and a working memory task. Heart rate was measured hourly (08:00-21:00 h) at rest and during performance tasks in 20 smokers (ten men, ten women) and 20 non smokers (ten men, ten women). Smoking and gender influenced reactivity only at certain times, especially at the earliest and latest hours and during the post lunch period. Smokers displayed major post-lunch interference and a pattern of lowered stress in the second half of the day. Women showed greater reactivity at the first daily recording, although their levels later became similar to the men's and were even lower. The statement that women are myocardial hyperreactors must be further investigated, as it seems women may take longer than men to adapt to a task. PMID- 11121883 TI - The small molecule FKBP ligand GPI 1046 induces partial striatal re-innervation after intranigral 6-hydroxydopamine lesion in rats. AB - Extensive unilateral striatal deafferentation was produced by intranigral 6 hydroxydopamine (6-OHDA) in rats. Beginning 60 days after 6-OHDA injection animals received a 14-day course of treatment with either the small molecule FKBP ligand GPI 1046 (10 mg/kg) or its vehicle alone. Striatal dopaminergic innervation density was determined from high power image analysis of striatal tyrosine hydroxylase (TH) immunohistochemistry. GPI 1046 treatment did not alter TH fiber density in the contralateral striatum but did produce significantly higher striatal TH fiber density in the ipsilateral caudate-putamen. This striatal re-innervation occurred in the absence of increased nigral sparing, and appears to reflect the GPI 1046 induced sprouting of residual TH+ fibers spared by the 6-OHDA lesion. PMID- 11121885 TI - Human subjects exposed to a specific pulsed (200 microT) magnetic field: effects on normal standing balance. AB - Static and time-varying magnetic fields have been shown to alter animal and human behaviors, such as directional orientation, learning, pain perception (nociception or analgesia) and anxiety-related behaviors. Human volunteers (12 male, 12 female, 18-34 years old) stood on a force plate while within three square magnetic field coil pairs (2, 1.75 and 1.5 m) arranged orthogonal with the uniform magnetic field volume centered at head level. Analysis of the data shows a significant improvement of normal standing balance or center of pressure, with eyes open or eyes closed, by a specific pulsed 200 microT(pk) magnetic field (PEMF). There was no significance found in control condition testing, such as sham-sham exposure of subjects or sham/PEMF exposure of a 60 kg saline phantom. There were no significant effects of gender or age. PMID- 11121884 TI - Interference of biocytin with opioid-evoked hyperpolarization and membrane properties of rat spinal substantia gelatinosa neurons. AB - In our laboratory, preliminary whole-cell, tight seal recordings of rat spinal substantia gelatinosa neurons including biocytin in the patch pipette yielded a significantly smaller proportion of neurons hyperpolarized by selective opioid agonists compared with recordings without biocytin. Therefore, we investigated the effects of biocytin inclusion on opioid responses and other membrane properties during whole-cell, tight seal recordings of these neurons. The percentage of neurons hyperpolarized by mu-, delta(1)-, and kappa-selective opioids was significantly reduced when 1% but not < or =0.2% biocytin was included in the recording pipette, compared with neurons recorded without biocytin. However, a significantly higher proportion of neurons fired spontaneous action potentials with either 0.05-0.2 or 1% biocytin compared to no biocytin. Resting membrane potential, input impedance and the proportion of neurons displaying transient outward rectification were each significantly altered for neurons recorded with 1% but not 0.05-0.2% biocytin. These effects may be due to a relatively specific blockade of diverse potassium channel types. Because efficient labeling can be achieved with 0.1% biocytin with whole-cell recording, higher concentrations are contraindicated. PMID- 11121886 TI - The human dopamine transporter gene: the 5'-flanking region reveals five diallelic polymorphic sites in a Caucasian population sample. AB - The 5'-flanking region of the human dopamine transporter (hDAT) was systematically screened for variants by single strand conformation analysis (SSCA) between -1586 and +97 basepair (bp) relative to the transcription start site. Five diallelic polymorphisms were found, which were shown to be due to single base substitutions: T-67A, G-660C, C-839T, C-1169G, T-1476G. In a population sample of 119 unrelated Caucasians, allele frequencies of the rarer allele were 47% for -67T, 3% for -660C, 45% for -839T, 50% for -1169G, and 8% for -1476G, respectively. Among 15 observed haplotypes, seven haplotypes collected a frequency of about 96% in our sample. T-67A, C-839T, C-1169G, T-1476G were related to potential transcriptional recognition sites. These findings and the occurrence of distinct haplotypes at the hDAT promoter locus in a Caucasian population sample make this region a promising target in the context of linkage and association studies in certain diseases. PMID- 11121887 TI - Catalase in astroglia-rich primary cultures from rat brain: immunocytochemical localization and inactivation during the disposal of hydrogen peroxide. AB - The expression of catalase in cells of astroglia-rich primary cultures derived from the brains of newborn rats was investigated by double-labelling immunocytochemical staining. Strong catalase immunoreactivity was found in cells positive for glial fibrillary acidic protein and galactocerebroside, cellular markers for astroglial and oligodendroglial cells, respectively. The cells of these cultures dispose of exogenously applied hydrogen peroxide (initial concentration 200 microM) quickly with first order kinetics. In contrast, after inhibition of glutathione peroxidases by mercaptosuccinate the rate of the catalase-dependent disposal of H(2)O(2) declined with time and after about 10 min the extracellular concentration of H(2)O(2) remained almost constant at a concentration of about 100 microM. Catalase activity after 10 min of incubation under these conditions was no longer detectable. In contrast, in the absence of mercaptosuccinate catalase activity was maintained during H(2)O(2) disposal. These results demonstrate that in astroglia-rich cultures catalase is strongly expressed in the predominant astroglial cells and in the minor population of oligodendroglial cells and that the enzyme is rapidly inactivated during the disposal of H(2)O(2), if the glutathione system of the cells is compromised. PMID- 11121888 TI - The relationship between differentiation and survival in PC12 cells treated with cyclic adenosine monophosphate in the presence of epidermal growth factor or nerve growth factor. AB - We have asked whether treatment of PC12 cells with cyclic adenosine monophosphate (cAMP) and epidermal growth factor (EGF) results, like treatment with cAMP and nerve growth factor (NGF), in irreversible neuronal differentiation characterized by irreversible neurite extension, loss of serum-dependence, and death by apoptosis after trophic factor withdrawal. Although EGF alone, unlike NGF, did not cause morphological differentiation or prevent cell death, synergy between a cAMP-mediated signal transduction pathway and a pathway activated by the EGF receptor tyrosine kinase resulted in the same irreversible differentiation. EGF/cAMP-differentiated cells required cAMP to survive, but NGF, through a TrkA dependent mechanism, could substitute for cAMP. The cyclin-dependent kinase inhibitors olomoucine and roscovitine also promoted survival of the irreversibly differentiated cells, by a mechanism that must be determined, since cell death was not associated with nuclear (3)H-thymidine accumulation, an index of mitotic activity. PMID- 11121889 TI - Identification of galpha subtype(s) involved in gamma-aminobutyric acid(B) receptor-mediated high-affinity guanosine triphosphatase activity in rat cerebral cortical membranes. AB - The ability of a series of specific Galpha carboxyl-terminal antisera, (i.e. anti Gsalpha, anti-Gi1/2alpha, anti-Gi3alpha/Goalpha, anti-Goalpha/Gi3alpha, and anti Gq/11alpha) to disrupt (+/-)-baclofen-stimulated high-affinity guanosine triphosphatase (GTPase) activity was explored in rat cerebral cortical membranes to identify the Galpha subunit(s) involved in gamma-aminobutyric acid(B) (GABA(B)) receptor-mediated signal transduction. Pretreatment of the membranes with the AS/7 (anti-Gi1/2alpha) antiserum inhibited GABA(B) receptor-mediated response without affecting the basal activity. The RM/1 (anti-Gsalpha) and QL (anti-Gq/11alpha) antisera failed to inhibit GABA(B) receptor-coupled responses. The results of the EC/2 (anti-Gi3alpha/Goalpha) and GO/1 (anti-Goalpha/Gi3alpha) antisera were difficult to interpret since the basal activities were influenced by these antisera. These results, in conjunction with the data in our previous reconstitution study, indicate that Gi2alpha is a main transducer of GABA(B) receptor-mediated signaling in rat cerebral cortex. PMID- 11121891 TI - Sexual behaviour of Zebu bulls in the humid tropics of Costa Rica: single versus multiple-sire groups. AB - This study compares the courtship and mounting behaviour of Bos indicus bulls in single- and multiple-sire groups in Costa Rica (latitude 10 degrees 25'N, longitude 84 degrees 32'W, annual precipitation of 3096mm, temperature of 24 degrees C and humidity of 85.3%). Four, 3-4 year-old Brahman bulls with previous sexual experience were used to sire a group of 120 multiparous cows (average of 128 days post-partum and a body condition score of 2.5) allocated to two groups of 60 each: (1) single-sire mating group (SSM) and (2) multiple-sire mating group of three bulls (MSM). Bulls were rotated among groups every 7 days for 28 days. The frequency, type and duration of sexual activities (mounting and mounting attempts) and courtship activities (smelling and licking genital area, butting, supporting the head over a female and the sign of Flehmen) were calculated for each mating group. Descriptive and non-parametric statistics (Wilcoxon, Mann Withney tests) were used to calculate differences between mating programmes. Sexual activities tended to be more frequent in the SSM group than the MSM group (267 versus 124, P>0.05). Differences in the ratios of sexual to courtship activities between both groups were significant (1:3 in SSM and 1:6 in MSM, P>0.05). Pregnancy rate averages were 28 and 37%, respectively, (P>0.05). It is concluded that under these conditions multiple-sire mating and single-sire mating achieved similar pregnancy rates. PMID- 11121890 TI - Voltage-gated calcium channels contribute to the pattern of the resting discharge in guinea pig medial vestibular nucleus neurons. AB - In brainstem slices of guinea pigs perfused with artificial cerebro-spinal fluid (ACSF), the discharge of all the spontaneously active neurons of the medial vestibular nucleus (MVN) is regular. It has been reported that prolonged exposure to a low Ca(2+) medium could induce these neurons to fire bursts of spikes. In this study, we performed a systematic exploration of the spontaneous activity of the guinea pig MVN neurons by extracellular recordings in slices perfused either with a low Ca(2+)-high Mg(2+) medium, or with ACSF added with omega-agatoxin-IVA and with omega-conotoxin-GVIA. The percentage of recorded neurons which fired bursts, was 67% in low Ca(2+)-high Mg(2+) medium and 34% under the action of Ca(2+) channel blockers. These results show that the sensitivity of the firing properties to divalent cations is not shared by all of the MVN neurons and that the regularity of firing of a class of MVN neurons depends on the Ca(2+) channels they express in their membranes. PMID- 11121892 TI - Effect of cyclodextrin-encapsulated beta-carotene on progesterone production by bovine luteal cells. AB - Experiments were conducted to examine the effect of cyclodextrin-encapsulated beta-carotene on basal or cholesterol (cyclodextrin-encapsulated), LH and dibutyryl cyclic AMP (dbcAMP)-stimulated progesterone production by bovine corpus luteum cells isolated from mid-luteal heifer ovaries by collagenase digestion. Cells were cultured with serum-free DMEM/Ham's F12 medium in serum pre-treated plastic culture dishes for periods of up to 11 days. Medium was replaced after 24h and thereafter every 48 h. Beta-carotene was added to cultures in a carrier molecule, dimethyl-beta-cyclodextrin, to facilitate dissolution. All treatments were started on day 3 of culture. Treatment of cells with 1 or 2 micromol/l beta carotene resulted in sharp inhibition of progesterone production. On the contrary, treatment of cells with 0.1 micromol/l beta-carotene resulted in significant stimulation (P<0.05) of both basal and cholesterol-stimulated progesterone secretion. The effect of beta-carotene on LH or dbcAMP-stimulated progesterone production was also examined. Treatment of cells with LH or dbcAMP always resulted in stimulation of progesterone secretion (P<0.001). However, cells treated with LH plus beta-carotene or dbcAMP plus beta-carotene both produced significantly (P<0.01) less progesterone relative to those cells treated with LH or dbcAMP alone on days 7, 9 and 11 of culture. These results indicate that beta-carotene can enhance luteal steroidogenesis when present at low concentrations but is inhibitory at higher concentrations and that encapsulation of beta-carotene in cyclodextrin is an effective method of supplying it to cells in culture. PMID- 11121893 TI - Administration of gonadotropin-releasing hormone during metoestrus in cattle: influence on luteal function and cycle length. AB - Gonadotropin-releasing hormone (GnRH) has been used to warrant the success of artificial insemination by accurately timing occurrence of ovulation. In practical conditions, GnRH may be administered too late, after ovulation, with an eventual reduction in pregnancy rate. The aim of this study was to investigate whether GnRH administration after ovulation would have a negative effect on luteal function. Three cows and six heifers of the Finnish Ayrshire breed were used. Oestruses were synchronised. After detection of ovulation, one of the following treatments was implemented: gonadorelin (250 microg, i.m.) at either 0 24h (T1) or 24-48h (T2) post-ovulation or control (no gonadorelin, C). Every animal was assigned once to each of these three manipulations. Ultrasonography was performed on days 1, 4 or 5, 7 or 8, 11 or 12, 14 or 15 post-ovulation and daily from the beginning of the next oestrous signs until ovulation (day 0=day of ovulation). Blood samples for progesterone (P(4)) determinations were collected daily from day 1 after the occurrence of ovulation until recording of the next oestrus. Administration of GnRH during metoestrus did not induce ovulation of either large or small follicles and, thus, no accessory corpora lutea (CL) were formed. In T1, on day 14 or 15, the diameter of CL was 1.3+/-0.3mm smaller than in C (P<0.01), but no differences were found either on days 11 or 12 or on the same days of the T2 and C treatments. No significant differences in levels or profiles of P(4) curves were found between GnRH treatments and control. Neither had the treatments any effects on the length of the oestrous cycle. In conclusion, GnRH treatment during metoestrus does not seem to alter subsequent luteal function and, thus, this does not explain previous reports of reduced fertility post-treatment. PMID- 11121894 TI - The role of sub-optimal preovulatory oestradiol secretion in the aetiology of premature luteolysis during the short oestrous cycle in the cow. AB - Premature regression of the corpus luteum, following the first post partum ovulation, is often preceded by sub-optimal preovulatory oestradiol secretion and accompanied by elevated levels of oxytocin receptors early in the luteal phase. We have investigated the role of preovulatory oestradiol in the control of subsequent oxytocin receptor concentration and activity by treating ovariectomised cows, over a simulated 48 h follicular phase, with high (600 microg per day) medium (300 microg per day) or low (150 microg per day) levels of oestradiol. These doses of oestradiol generated mean+/-S.E.M. plasma oestradiol concentrations of 12.1+/-1.0, 4.9+/-0.5 and 2.9+/-0.4 pg ml(-1), respectively. In Study 1 (n=4 per group), we found that by day 4 following oestrus there was a significant (P< 0.05) effect of the level of oestradiol on the inhibition of oxytocin binding activity measured in endometrial biopsy samples. This had fallen to mean+/-S.E.M. concentrations of 25+/-2 fmol per mg protein in the high group, 47+/-8 fmol per mg protein in the medium group and 65+/-12 fmol per mg protein in the low group. In Study 2, cows (n=3 per group) were treated with the same three levels of oestradiol followed by treatment with increasing levels of progesterone from days 3 to 6 following oestrus, generating mean+/-S.E.M. plasma concentrations of 2.17+/-0.18 ng ml(-1) by day 6. On day 6, there was a significant (P< 0.01) effect of the level of oestradiol on PGF(2alpha) release in response to oxytocin challenge. High, medium and low oestradiol groups exhibiting mean+/-S.E.M., increase plasma PGF(2alpha) metabolite concentrations of 10.0+/ 2.2, 21.3+/-4.3 and 41.3+/-1.2 pg ml(-1), respectively, during the hour after oxytocin administration. From these results, we postulate that at the first post partum ovulation a low level of preovulatory oestradiol can result in the early generation of a luteolytic mechanism during the subsequent luteal phase due to impaired inhibition of oxytocin receptors allowing increased PGF(2alpha) release. PMID- 11121895 TI - Preovulatory follicular status and diet affect the insulin and glucose content of follicles in high-yielding dairy cows. AB - Insulin and glucose may be limiting factors for ovarian function in dairy cows genetically selected for high milk yield. The effects of nutrition on the intrafollicular content of insulin and glucose were investigated in Israeli Holstein dairy cattle fed a basic total mixed ration and producing 34-39kg of milk daily. In experiment 1, carried out in 11 oestrus-synchronised cows, little variation in insulin concentration was found in plasma sampled during the luteal phase, but high variation was found in plasma sampled during the follicular phase. Therefore, in order to prevent confounding the effects of diet and of phase in cycle in the following experiments, experimental diets were fed during the luteal phase of synchronised oestrus cycles. In experiment 2, designed as Latin-Square, six cows received sequentially diets containing 17.1 (control) or 19.7% of crude protein, using two sources of supplementary protein, i.e. soyabean meal (SBM) and corn gluten meal (CGM), differing in ruminal degradability and leucine content. When dry matter intake was used as covariant, plasma insulin on day 16 was 29.5 and 26.4% higher in cows fed diets containing SBM and CGM than in the control (P<0.05). In experiment 3, 17 cows were individually fed the basic diet and then switched to isoenergetic diets containing SBM (n=5), CGM (n=6) or corn grain (CG, n=6) given from day 10 to 16 of the synchronised oestrus cycle. On the eve of day 16, and in the morning of day 17, they were administered PGF(2alpha) and the content of 26 largest follicles was aspirated by using the transvaginal ovum pick-up technique. Follicles were sorted into two classes (preovulatory and subordinate) according to oestradiol concentration and the progesterone:oestradiol ratio in follicular fluid (FF). Higher concentrations of insulin (0.282 versus 0.127ng/ml, P<0.0001) and of glucose (0.614 versus 0.386g/l, P<0.002), were found in FF from preovulatory follicles. The insulin concentration in the FF of cows fed the CG diet was 26% higher than in their counterparts fed CGM (P<0.04), SBM being intermediate. Dietary effects did not reach significance in subordinate follicles. The finding that preovulatory follicular status is associated with increased intrafollicular insulin and glucose suggests that insulin is involved in follicular maturation. The nutritional effect on intrafollicular glucose and insulin may have practical implications to optimise feeding in dairy cows during phases of the oestrus cycle. PMID- 11121896 TI - Uptake of glucose and cholesterol by the ovary of sheep and cattle and the influence of arterial LH concentrations. AB - Time series analysis methods were used to evaluate the relationships between the uptake of glucose and cholesterol, and arterial luteinizing hormone (LH) concentrations. Classical arterio-venous difference methods were applied to study ovarian uptake of metabolites. Arterial and venous samples (n=20) were obtained from six cows and nine sheep every 10min. There were highly significant positive cross-correlations of 0.5 for cattle and 0.8 for sheep between the uptake of glucose and cholesterol at lag 0. All individual cross-correlations were significant for sheep. Uptake of these metabolites was not significantly associated with arterial LH concentrations in the cows. This study suggests that glucose may promote cholesterol uptake into the ovarian cells or vice versa. This study is the first to identify such a relationship. If these findings are repeated, the possibility exists that control of the oestrous cycle and fertility may be achieved by seeking a common regulator of uptake of these metabolites or by uncoupling the association between glucose and cholesterol. PMID- 11121897 TI - Nocturnal variation of prolactin secretion in the Mouflon (Ovis gmelini musimon) and domestic sheep (Ovis aries): seasonal changes. AB - Seasonal changes in nocturnal prolactin secretion and their relationship with melatonin secretion were monitored in wild (Mouflon, Ovis gmelini musimon) and domesticated sheep (breed Manchega, Ovis aries). Two groups of eleven adult females each, were maintained outdoors under natural photoperiod. Plasma concentrations of prolactin and melatonin were determined during the summer and winter solstices and the autumn and spring equinoxes. Blood samples were collected every 3h during the night hours, and 1h before and after the onset of darkness and sunrise. Maximum mean plasma concentrations of prolactin during the dark-phase in Mouflons were observed in the summer solstice, (P<0.001) and in the summer solstice and spring equinox in Manchega ewes (P<0.001). Mean plasma concentrations of prolactin were higher in the wild species (P<0.001) during the summer solstice. In contrast, during the spring equinox, mean levels of prolactin were higher in Manchega ewes than in Mouflons (P<0.05). Plasma prolactin concentrations showed a nocturnal rhythm in both breeds, with seasonal variations (P<0.001). The increase in plasma melatonin levels during the first hour after sunset was accompanied to increasing concentrations of PRL 1h after the onset of darkness, only in the autumn and spring equinox for the Mouflon, and in the summer solstice and spring equinox for the Manchega ewes. In Mouflons, the fall of plasma PRL concentrations about the middle dark-phase in all the periods studied, coincided with high levels of melatonin. A similar relation was observed in Manchega ewes only in the winter solstice and spring equinox. The current study shows that the nocturnal rhythm of prolactin secretion exhibits seasonal variation; differences in the patterns of prolactin secretion between Mouflon and Manchega sheep are taken to represent the effects of genotype. PMID- 11121898 TI - Kinase pathways in dominant and subordinate ovarian follicles during the first wave of follicular development in sheep. AB - The mechanism by which one or more dominant ovarian follicles continue development while other subordinate follicles regress is not known. The mitogen activated protein kinases (MAPKs) are a group of kinases that are activated by hormonal factors and form a cascade of processes that regulate cell growth, division and differentiation. The aim of the present experiment was to characterise the presence of the MAPKs, Erk 1/Erk 2 and Akt in healthy dominant follicles and regressing subordinate follicles. Following in vivo monitoring of ovarian follicle development, three ewes were ovariectomised and the follicular fluid and follicle wall (theca and granulosa cells) saved from the dominant and largest subordinate follicle. The dissected diameter and follicular fluid oestradiol concentration of the dominant follicle was larger (P<0.01) than the largest subordinate follicle (6.5+/-0.0mm and 41.3+/-4.9ng/ml versus 4.7+/-0.3mm and 0.6+/-0.4ng/ml). Western blot analyses showed that there was more Akt (202.7+/-6.4 versus 59.6+/-32.7 units; P<0.05) and Erk 1/Erk 2 (104.5+/-10.6 versus 0.3+/-0.2 units; P<0.01) present in follicle wall samples from the dominant compared to the largest subordinate follicles. Phosphorylated forms of Akt and Erk 1/Erk 2 were detected in samples from dominant but not subordinate follicles. We suggest that signal transduction pathways involving Akt and Erk 1/Erk 2 may play an important role in determining the outcome of ovarian follicle growth and development in sheep. PMID- 11121899 TI - Seminal carnitine and acetylcarnitine content and carnitine acetyltransferase activity in young Maremmano stallions. AB - The reproductive characteristics and seminal carnitine and acetylcarnitine content as well as carnitine acetyltransferase activity of young Maremmano stallions (n=25) are reported. The stallions were subjected to semen collection in November and January; in each trial two ejaculates were collected 1h apart. The total motile morphologically normal spermatozoa (TMMNS) and the progressively motile spermatozoa at collection and during storage at +4 degrees C were evaluated. Seminal L-carnitine (LC), acetylcarnitine (AC), pyruvate and lactate were measured using spectrophotometric methods, whereas carnitine acetyltransferase activity was measured by radioenzymatic methods. Since there were no major significant differences in seminal and biochemical characteristics between the November and January trials, data were also pooled for the first and second ejaculates. Significant differences (P<0.001) were observed between the first and second ejaculates for sperm count (0.249+/-0.025 versus 0.133+/ 0.014x10(9)/ml), total number spermatozoa by ejaculate (12.81+/-1.23 versus 6.36+/-0.77x10(9)), progressively motile spermatozoa (48.6+/-3.0 versus 52.6+/ 3.0%) and TMMNS (3.35+/-0.50 versus 2.02+/-0.37x10(9)). In the raw semen the LC and AC were significantly higher in the first ejaculate than in the second (P<0.001), whereas, pyruvate and pyruvate/lactate ratio were higher in the second ejaculate (P<0.05). Seminal plasma AC and LC concentrations resulted higher in the first ejaculate (P<0.001). The pyruvate/lactate ratio was higher in the second ejaculate (P<0.05). Both raw semen and seminal plasma LC and AC concentrations were positively correlated with spermatozoa concentration (P<0.01); in raw semen AC was also correlated to TMMNS (P<0.01). Lactate levels of raw semen was correlated to progressively motile spermatozoa after storage (P<0.01). In the second ejaculate, significant correlations were also observed among AC/LC ratio in raw semen and progressively motile spermatozoa after 48 and 72h of refrigeration. Furthermore, AC levels were correlated to lactate concentration. The positive correlation between LC, AC and spermatozoa concentration, and between AC and TMMNS indicated carnitine as potential semen quality marker. Moreover, the correlation between AC/LC ratio and progressive spermatozoa motility after refrigeration, suggests that carnitine may contribute towards improving the maintenance of spermatozoa viability during in vitro storage. PMID- 11121900 TI - The influence of blood cells and PDGF on porcine theca cell function in vitro. AB - The role of red and white blood cells in the regulation of porcine theca cell function is poorly understood. Interactions between these cell types and a potential mediator of any interaction, PDGF, were investigated using a serum-free culture system. Theca cells were collected from 6-9mm antral follicles and plated at 50x10(3) viable cells/well. In the first experiment, macrophages were removed and theca cells+/-macrophages were cultured with a range of PDGF doses (0.1, 1, and 10ng/ml)+/-IGF-1. In the second experiment, red blood cells were removed with lysing buffer. In both experiments the effect of treatment on steroidogenesis and viable cell number was examined. Macrophage removal decreased oestradiol production but increased androstenedione output irrespective of the presence of IGF-1 (oestradiol+/-IGF-1, P<0.001; androstenedione P=0.02 without IGF-1, P<0.001 with IGF-1). PDGF increased oestradiol synthesis by whole and macrophage-free theca cell preparations but only in the presence of IGF-1 (P<0.001). In contrast, androstenedione production was unaffected by PDGF dose in the presence of IGF-1 (P=0.67). Without IGF-1, 10ng/ml PDGF tended to decrease androstenedione levels (P=0.06). Macrophage removal increased viable cell number at 144h (P<0.001+/-IGF 1) as did PDGF (P<0.001+/-IGF-1). In the absence of IGF-1, there was a PDGF x cell type interaction (P=0.02). Macrophage-free cultures with 10ng/ml PDGF had twice as many viable cells as whole preparations with no PDGF. In the second experiment, red blood cell removal did not affect steroidogenesis or the number of viable cells present at 144h when cells were cultured with IGF-1. The data show that theca cell/macrophages interactions do occur, and influence both steroidogenesis and viable cell number during culture. The macrophage product(s) enhanced oestradiol synthesis but reduced androstenedione production and the number of viable cells. As all these interactions were not mimicked by PDGF, PDGF cannot be the only factor mediating the theca/macrophage interaction. When cultured under optimised conditions the presence of red blood cells was not detrimental to theca cell steroidogenesis or the number of viable cells. PMID- 11121901 TI - Review article: the role of non-acid reflux in gastro-oesophageal reflux disease. AB - The role of acid in the pathogenesis of gastro-oesophageal reflux disease (GERD) has been extensively studied and is well accepted. The role, if any, of non-acid reflux, in particular duodenogastro-oesophageal reflux, is much debated. The availability of new technology to detect non-acid reflux has heightened interest in this question. This article reviews the following: How do we define non-acid reflux? Does duodenogastro-oesophageal reflux (alone or in combination) cause oesophageal injury, symptoms or both? What is its role in complicated GERD? What methods are available to assess non-acid reflux? Does non-acid reflux need treatment and if so what modalities are available? PMID- 11121902 TI - Inflammatory bowel disease: epidemiology and management in an English general practice population. AB - BACKGROUND: Inflammatory bowel diseases have significant long-term morbidity and healthcare resource consequences. Studies based on secondary care records may have underestimated the contribution of general practitioners (GPs) to its management. AIMS: To describe the epidemiology and management of inflammatory bowel disease using GP records as the primary data source. METHODS: A systematic search of GP clinical records in northern England, identifying cases of inflammatory bowel disease, patient consultation behaviour, prescribing patterns, and extent of specialist care. RESULTS: In a population of 135 723, the incidence of ulcerative colitis was 13. 9/100 000 per year (CI: 7.5-20.3) and for Crohn's disease 8.3/100 000 per year (CI: 3.4-13.2). The age-sex adjusted point prevalence for ulcerative colitis on 1st January 1995 was 243.4/100 000 (CI: 217.4-269.4) and for Crohn's disease 144.8/100 000 (CI: 124.8-168.8). The mean number of consultations (s.d.) with specialists and GPs were similar, both in the first 12 months after referral (specialists 3.94 +/- 3.15, GPs 3.34 +/- 3.55) and in the most recent 12 months (1.02 +/- 2.02, 1.04 +/- 2.04). Only 29.9% of all patients were definitely under specialist care. CONCLUSIONS: Prevalence rates, but not incidence rates, for inflammatory bowel disease are substantially higher than previously described in UK populations. General practitioners make a significant contribution to meeting the healthcare needs of these patients. PMID- 11121903 TI - The use of 6-mercaptopurine in patients with inflammatory bowel disease after failure of azathioprine therapy. AB - INTRODUCTION: Azathioprine is a useful therapy in patients with inflammatory bowel disease that is difficult to control. However, 10% of patients are unable to tolerate azathioprine, and the best form of treatment for this group of patients is unknown. The azathioprine metabolite 6-mercaptopurine may be a useful therapy for these patients. AIM: To review our clinical experience of the use of the 6-mercaptopurine in inflammatory bowel disease patients who are intolerant of azathioprine. METHODS: All patients who were prescribed 6-mercaptopurine in a 2 year period were identified from pharmacy records. The case notes were reviewed and those who had previously been intolerant of azathioprine were included. RESULTS: A total of 19 with either ulcerative colitis and Crohn's disease were included. The reasons for discontinuing azathioprine were side-effects (13 patients), failure of efficacy (four patients) and leucopenia (two patients). Eleven of the 19 patients (68%) tolerated 6-mercaptopurine, including seven out of 13 patients (54%) who discontinued azathioprine due to side-effects. The length of follow-up of patients on 6-mercaptopurine was between 126 and 780 days (median 390 days). DISCUSSION: 6-mercaptopurine should be considered in patients with inflammatory bowel disease who require continuing immunosuppressive therapy, but are intolerant of azathioprine. PMID- 11121904 TI - A pilot study of N-acetyl glucosamine, a nutritional substrate for glycosaminoglycan synthesis, in paediatric chronic inflammatory bowel disease. AB - BACKGROUND: The breakdown of glycosaminoglycans is an important consequence of inflammation at mucosal surfaces, and inhibition of metalloprotease activity may be effective in treating chronic inflammation. AIM: To report an alternative approach, using the nutriceutical agent N-acetyl glucosamine (GlcNAc), an amino sugar directly incorporated into glycosaminoglycans and glycoproteins, as a substrate for tissue repair mechanisms. METHODS: GlcNAc (total daily dose 3-6 g) was administered orally as adjunct therapy to 12 children with severe treatment resistant inflammatory bowel disease (10 Crohn's disease, 2 ulcerative colitis). Seven of these children suffered from symptomatic strictures. In addition, similar doses were administered rectally as sole therapy in nine children with distal ulcerative colitis or proctitis resistant to steroids and antibiotics. Where pre- and post-treatment biopsies were available (nine cases), histochemical assessment of epithelial and matrix glycosaminoglycans and GlcNAc residues was made. FINDINGS: Eight of the children given oral GlcNAc showed clear improvement, while four required resection. Of the children with symptomatic Crohn's stricture, only 3 of 7 have required surgery over a mean follow-up of > 2.5 years, and endoscopic or radiological improvement was detected in the others. Rectal administration induced remission in two cases, clear improvement in three and no effect in two. In all cases biopsied there was evidence of histological improvement, and a significant increase in epithelial and lamina propria glycosaminoglycans and intracellular GlcNAc. CONCLUSIONS: GlcNAc shows promise as an inexpensive and nontoxic treatment in chronic inflammatory bowel disease, with a mode of action which is distinct from conventional treatments. It may have the potential to be helpful in stricturing disease. However, controlled trials and an assessment of enteric-release preparations are required to confirm its efficacy and establish indications for use. PMID- 11121905 TI - Factors associated with consulting medical or non-medical practitioners for dyspepsia: an australian population-based study. AB - BACKGROUND: Little is known about how many dyspeptics in the population consult medical and non-medical practitioners, or the factors associated with various consulting patterns. METHODS: A cross-sectional survey of 748 Australians with dyspepsia investigated their age, sex, dyspepsia symptoms, medical and non medical consultations, and health status on the SF-12. RESULTS: Overall, 56% had ever consulted a medical practitioner for dyspepsia. Of these, 54% consulted within 6 months of first symptoms. Non-medical practitioners were consulted by 29%. Compared to dyspeptics in all, or most, other consulting groups, subjects who did not consult (37%, group NO) were characterized by fewer symptoms, better physical health, and younger age. Those who only consulted doctors (34%, group M) were older and had better mental, but poorer physical health. Those who only consulted non-medical practitioners (7%, group N) were younger and had better physical, but poorer mental health. Dyspeptics consulting both medical and non medical practitioners (22%, group M + N), were older, more dissatisfied with medical care, had more symptoms and poorer physical and mental health. Timing of medical consultations was similar in groups M and M + N. Group M + N dyspeptics consulted similar types, but more non-medical practitioners than group N. No sex differences were found in consulting behaviour. CONCLUSIONS: Many dyspeptics do not consult; they have fewer symptoms than consulters. Consultation with non medical practitioners is common and is associated with poor mental health. Dyspeptics seeking advice from both medical and non-medical practitioners are less satisfied with their medical management than those who only consult doctors for their dyspepsia. PMID- 11121906 TI - The prevalence of gastro-oesophageal reflux symptoms in a UK population and the consultation behaviour of patients with these symptoms. AB - BACKGROUND: Patients consulting with gastro-oesophageal reflux symptoms (GORS) may differ from nonconsulters. AIM: To describe these differences in a UK population. METHODS: A postal questionnaire was sent to 4432 adults. Definitions used were GORS (either heartburn or acid regurgitation on more than six occasions during the previous year), dyspepsia (upper abdominal pain or discomfort on more than six occasions during the previous year) and irritable bowel syndrome (abdominal pain with three or more Manning criteria). Socio-economic status was identified by the Standard Occupational Classification. RESULTS: With a 71.7% response, GORS were reported by 28.7% of the sample, it was unaffected by gender and age but was more common among the socially disadvantaged (P < 0. 005). Less than 25% of GORS patients consulted during the previous year. Increasing age (chi2 for trend; P < 0.001) and coexisting upper abdominal symptoms (chi2 P < 0.001) positively influenced consultation behaviour, but it was unaffected by socio-economic status, gender, or the coexistence of irritable bowel syndrome. Dyspepsia and nausea independently predicted consultation. CONCLUSIONS: GORS are especially common among the deprived. Socio-economic variables do not affect consultation behaviour, but the patient's age and the burden (number and type) of associated symptoms do. PMID- 11121907 TI - Omeprazole 40 mg once a day is equally effective as lansoprazole 30 mg twice a day in symptom control of patients with gastro-oesophageal reflux disease (GERD) who are resistant to conventional-dose lansoprazole therapy-a prospective, randomized, multi-centre study. AB - BACKGROUND: Comparative studies of omeprazole and lansoprazole are scarce and even scarcer are comparisons of higher doses. Most of the comparative studies have assessed the effect of the two proton pump inhibitors (PPIs) on gastric acid secretion or gastric pH. Few studies have compared clinical end-points such as oesophageal healing and symptom control. AIM: To determine the clinical efficacy of omeprazole 40 mg daily as compared to lansoprazole 30 mg twice a day in symptom control of patients with severe symptomatic GERD. METHODS: Ninety-six patients who failed a standard dose of lansoprazole (30 mg once daily), were enrolled in a prospective fashion from three VA medical centres and were randomized to receive 6 weeks of either omeprazole 40 mg daily or lansoprazole 30 mg twice daily. Patients reported daily on symptom severity and frequency, antacid consumption and side-effects. RESULTS: Forty-six patients received omeprazole and 44 lansoprazole. Although not statistically significant, there was a consistent trend of better symptom control in the omeprazole group for daytime and night-time heartburn and acid regurgitation. There was no statistical difference between the two groups in mean antacid consumption overall and at the end of each of the 6 weeks of the study. In addition, there was no significant difference in the overall frequency of side-effects between the two groups nor for each individual side-effect. CONCLUSION: Omeprazole 40 mg once daily is equally effective and tolerated as lansoprazole 30 mg twice daily in symptom control of patients with GERD. PMID- 11121909 TI - Pantoprazole, azithromycin and tinidazole: short duration triple therapy for eradication of Helicobacter pylori infection. AB - BACKGROUND: Azithromycin is an acid-stable macrolide that achieves remarkably high concentrations in gastric tissue, persisting above the MIC90 for Helicobacter pylori over a period of 5-days, after a single 500 mg oral dose. AIM: To evaluate and compare the efficacy, safety, and tolerability of two eradicating regimens of pantoprazole, azithromycin and tinidazole. METHODS: A total of 100 consecutive symptomatic H. pylori-positive patients received pantoprazole 40 mg b.d. for 1 week, and were randomly assigned to either azithromycin 500 mg o.m. and tinidazole 500 mg b.d. during the first 3 days (early group, n=50) or during the last 3 days of therapy with pantoprazole (late group, n=50). H. pylori status was assessed by histology and rapid urease test at entry and by histology and 13C-urea breath test 1 month after the end of the therapy. RESULTS: Ninety-nine patients completed the study. H. pylori was eradicated in 86% of patients in the early group (intention-to-treat 86%) and in 88% of patients in the late group (intention-to-treat 88%). CONCLUSIONS: This short triple therapy is effective for H. pylori eradication. The compliance was excellent and side-effects negligible. Moreover, the pantoprazole pre-treatment did not modify the efficacy of the therapy. PMID- 11121908 TI - One-week triple therapy with esomeprazole provides effective eradication of Helicobacter pylori in duodenal ulcer disease. AB - BACKGROUND: Esomeprazole is the first proton pump inhibitor to be developed as an optical isomer for the treatment of acid-related diseases. METHODS: Four hundred and forty eight duodenal ulcer patients with Helicobacter pylori infection, confirmed by 13C-urea breath test (UBT), and no current ulcer, were randomised to double-blind treatment with esomeprazole 20 mg twice daily (b.d.) (n=224) or omeprazole 20 mg b.d. (n=224), in combination with amoxicillin 1 g b.d. and clarithromycin 500 mg b.d. for 1 week (EAC and OAC, respectively). A negative UBT at both 4 and 8 weeks after completing therapy indicated successful H. pylori eradication. RESULTS: Intention-to-treat (ITT) analysis comprised 400 patients (EAC, n=204; OAC, n=196) and per protocol (PP) analysis 377 patients (EAC, n=192; OAC, n=185). Eradication rates (95% confidence intervals) for ITT and PP populations were: EAC, 90% (85-94%) and 91% (86-94%); OAC, 88% (82-92%) and 91% (86-95%). Between-group differences in eradication rates were not statistically significant. Both regimens were well tolerated, with an adverse event profile and frequency typical of proton pump inhibitor plus antibiotic combination therapy. CONCLUSIONS: Esomeprazole-based triple therapy for 1 week is highly effective in eradicating H. pylori infection in duodenal ulcer disease, offers comparable efficacy to omeprazole-based therapy, and is well tolerated. PMID- 11121910 TI - 5-day vs. 7-day triple therapy with rabeprazole, clarithromycin and amoxicillin for Helicobacter pylori eradication. AB - AIM: To determine whether a 5-day regimen with rabeprazole, clarithromycin and amoxicillin (RCA) was as effective as a 7-day regimen. METHODS: A total of 139 H. pylori-infected patients were randomized to receive either a 5-day or 7-day course of rabeprazole 10 mg b.d., clarithromycin 400 mg b.d. and amoxicillin 750 mg b.d. Eradication was assessed by CLO test, histology and 13C-urea breath test. RESULTS: On the intention-to-treat basis, eradication rates were 66% (46 out of 70) and 84% (58 out of 69) for the 5- and 7-day regimens, respectively (P < 0.05). Using per protocol analysis, eradication rates were 70% (46 out of 66) and 91% (58 out of 64) for the 5- and 7-day regimens, respectively (P < 0.01). Adverse events, which were observed in 14 patients from each group, caused discontinuation of treatment in only two patients, resulting in excellent compliance. CONCLUSIONS: Our 5-day regimen of RCA yielded inferior results, whereas the 7-day regimen achieved an eradication rate exceeding 90% on the per protocol basis. Therefore, treatment regimens of less than 7 days for proton pump inhibitor-clarithromycin-amoxicillin therapies cannot be recommended. PMID- 11121911 TI - A lyophilized and inactivated culture of Lactobacillus acidophilus increases Helicobacter pylori eradication rates. AB - BACKGROUND: Acid suppression plus two antibiotics is considered the reference anti-Helicobacter pylori treatment. Reported eradication rates are around 65-80%. Human Lactobacillus acidophilus shows an in vitro inhibitory effect on the attachment of H. pylori to gastric epithelial cell lines. Culture supernatant of this bacillus seems to decrease the in vitro viability of H. pylori. AIM: To evaluate whether the supplementation with an inactivated preparation of L. acidophilus could improve the efficacy of a standard anti-H. pylori therapy. METHODS: One-hundred and twenty H. pylori-positive patients were randomly assigned to a 7-day triple therapy based on rabeprazole (20 mg b.d.), clarithromycin (250 mg t.d.s.) and amoxicillin (500 mg t.d.s.) (RCA group: 60 subjects), or to the same regimen supplemented with a lyophilized and inactivated culture of Lactobacillus acidophilus (t.d.s.) (RCAL group: 60 subjects). RESULTS: In the RCA group, eradication was successful in 72% (42 out of 58 patients) from a per protocol (PP) analysis, or 70% (42 out of 60 patients) using an intention to treat (ITT) analysis. In the RCAL group a significant increase in the eradication rate was observed: 88% (52 out of 59 patients) from PP analysis (P=0.03), 87% (52 out of 60 patients) from ITT analysis (P=0.02). CONCLUSIONS: These results seem to confirm the in vitro anti-H. pylori effect of L. acidophilus, suggesting that the inactivated L. acidophilus could be effective in increasing eradication rates of a standard anti-H. pylori therapy. PMID- 11121912 TI - Management of Helicobacter pylori in duodenal ulcer: a cost-effectiveness analysis. AB - BACKGROUND: Empirical eradication therapy of H. pylori has been proposed as a therapeutic alternative for duodenal ulcer. AIM: To identify the cost effectiveness of empirical eradication therapy vs. test-and-treatment for the management of patients already diagnosed with a duodenal ulcer. METHODS: A decision analysis was performed to compare the cost-effectiveness of empirical eradication therapy of H. pylori diagnosed duodenal ulcer vs. eradication therapy after confirmatory diagnosis of Helicobacter pylori infection by means of several diagnostic tests. RESULTS: The empirical eradication therapy of duodenal ulcer was found to be the most effective and cost-effective strategy of all the alternatives. Amongst the alternatives, which included the previous performance of confirmatory diagnostic tests, the best cost-effectiveness ratio used a serology test. The model was robust in the face of changes in the values of therapeutic effectiveness, sensitivity and specificity of the diagnostic tests, prevalence of H. pylori infection in duodenal ulcer, duration of the antisecretory therapy, and number of medical visits. CONCLUSIONS: Based on our cost-effectiveness analysis, a treat approach is more effective and cost effective than a test-and-treat approach in the clinical management of already diagnosed duodenal ulcer. PMID- 11121913 TI - Role of antimicrobial susceptibility testing on efficacy of triple therapy in Helicobacter pylori eradication. AB - BACKGROUND: Helicobacter pylori treatment failure may be due to resistance to macrolides and 5-nitroimidazoles. AIM: To test whether a preliminary in vitro susceptibility test of H. pylori to tinidazole and clarithromycin and a consequent specific regimen could improve the eradication rate. METHODS: A total of 109 consecutive H. pylori-positive patients with dyspeptic symptoms were included. At endoscopy, biopsy from the antrum was obtained for H. pylori culture and antimicrobial susceptibility testing. Fifty-six patients were treated with omeprazole, tinidazole and clarithromycin for 10 days (group OTC) and 53 patients received therapy on the basis of the susceptibility test (group SUSC). Treatment success was evaluated by the 13C-urea breath test 1 month after the end of therapy. RESULTS: Eight patients dropped out. Overall primary resistance to clarithromycin, tinidazole and both antibiotics was 13%, 33% and 4%, respectively. In group OTC, H. pylori was eradicated in 81% and 75% of patients by per protocol and intention-to-treat analysis, respectively. Per protocol and intention-to-treat eradication rates for group SUSC were 98% and 91% (P < 0.05 vs. group OTC). CONCLUSIONS: These data show that in H. pylori infection, antibiotic therapy based on the results of culture and susceptibility testing gives, in comparison to standard therapy, a significant improvement in eradication rate. PMID- 11121914 TI - Multicentre randomized placebo-controlled trial of ursodeoxycholic acid with or without colchicine in symptomatic primary biliary cirrhosis. AB - AIM: To establish the efficacy of combination therapy with ursodeoxycholic acid (UDCA) and colchicine in patients with symptomatic primary biliary cirrhosis (PBC), defined by the presence of liver cirrhosis, pruritus or bilirubin exceeding 2 mg/mL. METHODS: A total of 90 patients were randomly assigned to ursodeoxycholic acid 500 mg/daily plus placebo (UDCA group, n=44), or ursodeoxycholic acid at the same dosage plus colchicine, 1 mg/daily (UDCA/C group, n=46). The two groups were comparable for age, sex, stage of disease, severity of pruritus, bilirubin, and Mayo score. All patients underwent clinical, ultrasonographic, and biochemical examinations at entry and then every 6 months up to 3 years of follow-up. Patients with cirrhosis underwent endoscopy every 12 months. In a sub-group of patients without cirrhosis, who consented, liver biopsy was repeated at the end of the study. RESULTS: The number of treatment failures (i.e. dead, orthotopic liver transplantation (OLT), complications of cirrhosis, doubling of bilirubin, untreatable pruritus) was 11 (25%) in the UDCA group and four (9%) in the UDCA/C group (P < 0.05). No significant differences were observed in terms of improvement of liver enzymes related to cholestasis and cytolysis and of amelioration of pruritus. The Mayo score values increased less above the baseline values at 24 and 36 month-intervals in the UDCA/C group than in the UDCA group. Histological evaluation at baseline and at the end of the study was available for 15 patients with pre-cirrhotic stage. A significant reduction in histological grading score was observed in patients from the UDCA/C group, whereas no changes in these histological scores were observed in the UDCA group. CONCLUSIONS: The addition of colchicine to ursodeoxycholic acid in patients with symptomatic primary biliary cirrhosis results in a small but significant reduction of disease progress. PMID- 11121915 TI - Failure of a motilin receptor agonist (ABT-229) to relieve the symptoms of functional dyspepsia in patients with and without delayed gastric emptying: a randomized double-blind placebo-controlled trial. AB - INTRODUCTION: Motilin-receptor agonists are prokinetics; whether they relieve the symptoms of functional dyspepsia is unknown. We aimed to test the efficacy of the motilin agonist ABT-229 in functional dyspepsia patients with and without delayed gastric emptying. METHODS: Patients were randomized with postprandial symptoms and documented functional dyspepsia by endoscopy (n=589 in intention-to-treat analysis). Patients were assigned to either the delayed or normal gastric emptying strata, based on a validated 13C octanoic acid breath test. Patients were then further randomized within each strata, to receive one of four doses of ABT-229 (1.25, 2. 5, 5 or 10 mg b.d. before breakfast and dinner) or placebo for 4 weeks, following a 2-week baseline. The primary outcome was the assessment of change in symptom severity over the 2 weeks from baseline to final visit, based on a self-report questionnaire measuring severity on visual analogue scales. RESULTS: Baseline characteristics across the treatment arms were very similar. No significant differences in the upper abdominal discomfort severity score (maximum 800 mm) were observed for any active treatment arm vs. placebo (mean change from baseline -139, -141, -145, -160 and -134 mm for placebo, 1.25, 2.5, 5, and 10 mg, respectively, at 4 weeks by intention-to-treat). More patients on placebo reported a good or excellent global response than patients on 1.25 or 5 mg of active therapy (both P < 0.05). The results were very similar in those with and without delayed gastric emptying. Helicobacter pylori status did not predict response. Excluding patients with any baseline heartburn (total remaining n=240), ABT-229 10 mg was inferior to placebo in relief of upper abdominal discomfort. CONCLUSIONS: ABT-229 was of no value for relief of symptoms in functional dyspepsia, compared with placebo. PMID- 11121916 TI - Placebo-controlled, randomized, evaluator-blinded endoscopy study of risedronate vs. aspirin in healthy postmenopausal women. AB - BACKGROUND: Bisphosphonates are effective treatments for osteoporosis. Since some primary amino bisphosphonates are associated with oesophageal injury, we conducted a study of the upper gastrointestinal effects of risedronate, a pyridinyl bisphosphonate. METHODS: Healthy, postmenopausal women received risedronate 5 mg (n=26), aspirin 2600 mg (n=27), or placebo (n=27) daily for 14 days and underwent endoscopy at baseline, Day 8 and Day 15. RESULTS: Oesophageal erosions were noted in one subject in the aspirin group, two in the placebo group, and none in the risedronate group, and an ulcer in one aspirin-treated subject. Gastric erosions and ulcers were observed most frequently in the aspirin group. Gastric ulcers were noted in eight subjects in the aspirin group, one in the placebo group, and none in the risedronate group (P=0.010, placebo vs. aspirin; P=0.002, risedronate vs. aspirin). Duodenal erosions and ulcers were observed in the aspirin group only. Gastroduodenal erosion scores of three or more occurred more frequently in the aspirin than in the risedronate and placebo groups (P < 0.001). CONCLUSIONS: Risedronate 5 mg was not associated with oesophageal or gastroduodenal ulcers in healthy, postmenopausal women, a population representative of patients who will receive risedronate in the clinical setting. PMID- 11121917 TI - Efficacy and tolerability of a fixed combination of peppermint oil and caraway oil in patients suffering from functional dyspepsia. AB - AIM: To assess the efficacy and safety of enteric coated capsules containing a fixed combination of 90 mg peppermint oil and 50 mg caraway oil (PCC; Enteroplant) in patients with functional dyspepsia. METHODS: A total of 96 out patients received one capsule twice daily of PCC or placebo for 28 days. Primary efficacy variables were the intra-individual change in (i) pain intensity and (ii) sensation of pressure, heaviness and fullness between days 1 and 29, and the investigators' rating of (iii) global improvement (Clinical Global Impressions [CGI] item 2) on day 29. A global type I error of alpha=0.05 was controlled by a priori ordering of hypotheses. RESULTS: All patients were evaluable for efficacy and safety. On day 29, the average intensity of pain was reduced by 40% vs. baseline in the PCC group and by 22% in the placebo group. With regards to pressure, heaviness and fullness, a 43% reduction was observed for PCC vs. 22% for placebo. In CGI item 2, 67% (PCC) vs. 21% (placebo) of the patients were described as much or very much improved. In all three target parameters, the superiority of PCC over placebo was statistically significant. Six patients (PCC: 5; placebo: 1) reported adverse events, either unrelated to the trial, or attributable to an aggravation of the disease under investigation. Eructation with peppermint taste did not occur. CONCLUSION: These results demonstrate the good tolerability and the favourable risk-benefit ratio of PCC for the treatment of functional dyspepsia. PMID- 11121918 TI - A double-blind, randomized, dose response study testing the pharmacological efficacy of synthetic porcine secretin. AB - BACKGROUND: Biologically derived porcine secretin has been used as a diagnostic agent in clinical gastrointestinal practice for many years. Pure synthetic porcine secretin is now available for investigational clinical use. AIM: To compare the pharmacology of synthetic porcine secretin and biologically derived porcine secretin in healthy volunteers. METHODS: Secretin stimulation tests were performed in 12 volunteer subjects in a double-blind, randomized, Latin square crossover design study comparing three doses of synthetic porcine secretin (0.05, 0.2, and 0.4 microgram/kg) with a standard dose of biologically derived porcine secretin (1 CU/kg). Duodenal aspirates were analysed for total volume and for bicarbonate concentration. Total bicarbonate output was calculated. RESULTS: Twelve subjects completed four dosing regimens. A multiple comparison test was used to compare dosing regimens. The 0.2 and 0.4 microgram/kg doses of synthetic porcine secretin were not different from the 1 CU/kg dose of biologically derived porcine secretin for volume, bicarbonate concentration and total output from 0 to 60 min. Only one patient had an adverse event, which was mild, transient flushing after the 0.2 and 0.4 microgram/kg doses of synthetic porcine secretin and after the 1 CU/kg dose of biologically derived porcine secretin. CONCLUSIONS: Synthetic porcine secretin has identical pharmacologic effects to biologically derived porcine secretin in normal subjects. Both drugs were safe and well-tolerated. This study validates synthetic porcine secretin as a substitute for biologically derived porcine secretin. PMID- 11121919 TI - Phytomenadione or menadiol in the management of an elevated international normalized ratio (prothrombin time). AB - AIM: To evaluate the efficacy of oral menadiol compared to intravenous phytomenadione when correcting coagulopathies associated with cholestasis. METHODS: A total of 26 patients with cholestasis and an international normalized ratio (prothrombin time) greater than 1.2, were randomized to receive either 20 mg o.d. for 3 days of oral menadiol (n=12), or 10 mg o.d. of intravenous phytomenadione (n=14) prior to endoscopic retrograde cholangeopancreatography. Liver function tests and international normalized ratio were measured daily for 3 days. RESULTS: Liver function tests and international normalized ratio were comparable between groups at entry into the study (P > 0.05), but serum albumin was significantly lower in the intravenous phytomenadione group following treatment (P < 0.05). A decrease in international normalized ratio occurred in both groups following administration of vitamin K (P < 0.05). Two patients in the intravenous group required fresh frozen plasma, as failure to normalize international normalized ratio was observed. No adverse drug reactions were observed in either group, and no patient required re-admission for bleeding during a 4-week follow-up period after cholangeopancreatography. CONCLUSION: Oral menadiol appears to be an effective alternative to intravenous phytomenadione in the correction of coagulopathies associated with obstructive liver disease. This simplifies the care of patients with deranged clotting times requiring cholangeopancreatography, particularly those to be managed as out-patients. PMID- 11121920 TI - Opportunity knocks. PMID- 11121921 TI - Sequential organ scoring as a measure of effectiveness of critical care. AB - We performed an observational nonrandomised study in a critical care unit of a large district general hospital in England to establish whether sequential organ scoring could be used as a measure of effectiveness of intensive care. The degree of organ dysfunction of 75 consecutive patients admitted to the critical care unit whose duration of stay exceeded 48 h was measured using the Logistic Organ Dysfunction System score. The trends in organ dysfunction of survivors and non survivors were significantly different with function improving in survivors and remaining constant or worsening in non-survivors. In both groups, the degree of organ dysfunction decreased over the first three days of intensive care. On an individual patient basis, we achieved no change or an improvement in organ score over this period in 80% of patients. In terms of individual organ function, intensive care consistently improved scores relating to the cardiovascular, respiratory and renal systems over the first 72 h of care, but not the neurological, hepatic or haematological systems. In conclusion, daily organ scoring usefully reflects the ability of an intensive care unit to stabilise or reverse physiological dysfunction. PMID- 11121922 TI - The role of a high-dependency unit in a regional obstetric hospital. AB - The aim of our study was to review a series of critically ill patients admitted to a high-dependency unit (HDU) in a regional obstetric centre, to assess our HDU utilisation rate and to determine the indications for and rate of transfer to an intensive care unit (ICU) in a tertiary referral centre. A 4-year retrospective review of case notes and HDU/ICU registers was performed. One hundred and twenty three patients were admitted to the HDU in the 2 years following its inception, representing 1.02% of all deliveries. Obstetric complications accounted for 81.3% of admissions. Seventeen patients were admitted to an ICU during the study period; 12 (0.08%) were transferred before and five (0.04%) after the development of HDU facilities (p = 0.25). The advantages of a HDU within this setting include the concurrent availability of expert obstetric care and critical care management, the avoidance of the hazards of emergency transport and improved continuity of antenatal and postnatal care. PMID- 11121923 TI - The use of a target-controlled infusion of alfentanil to provide analgesia for burn dressing changes A dose finding study. AB - Burn dressing changes require profound analgesia for a short duration. This study aimed to determine whether an operator-adjusted target-controlled infusion of alfentanil could provide effective analgesia for burn dressing changes. Ten patients with a burn of between 5 and 50% of body surface area were studied. Pain scoring was performed before, during and following the dressing change. Target and effect site concentrations of alfentanil were recorded during and for 15 min after the dressing change. Median baseline visual analogue pain score (VAS) was 22 mm. Median (range) duration of dressing change was 35 (20-75) min. Median (range) VAS during the dressing change was 30 (14-66) mm. All patients were satisfied with their pain relief. There was no respiratory depression or cardiovascular instability. No patient became sedated and there were no episodes of nausea or vomiting. Median (range) total dose of alfentanil was 2.6 (1-10.7) mg. This study supports the efficacy of an operator-adjusted target-controlled infusion of alfentanil to provide analgesia for burn dressing changes. PMID- 11121924 TI - Menstrual cycle irregularity and the incidence of nausea and vomiting after laparoscopy. AB - Gender is an important risk factor for postoperative nausea and vomiting. Menstrual cycle phase has also been implicated as a further variable in female patients. It is not clear whether a history of an irregular menstrual cycle is a significant variable and studies to date have not examined this. The influence of an irregular menstrual cycle on nausea and vomiting after laparoscopy was evaluated in a blinded, prospective, observational study. One hundred and fifty nine patients scheduled for laparoscopy and dye investigation were included. Anaesthetic technique and postoperative analgesia were standardised. In comparison with patients who had a regular cycle, an irregular menstrual cycle increased the incidence of nausea or vomiting from 20.5% to 40.5% (absolute difference 20%; 95% CI 0.03-0.37, p = 0.01). Other variables were similar between the groups. An irregular menstrual cycle appears to increase the risk of postoperative nausea and vomiting. PMID- 11121925 TI - National obstetric anaesthetic practice in the UK 1997/1998. AB - In the United Kingdom, the Royal College of Obstetricians and Gynaecologists requires maternity units recognised for training to complete annual statistical returns. Analysis of these data revealed that anaesthetists were directly involved in more than 251 000 procedures in the peripartum period in 1997/1998. There had been an increase in the number of women delivered by Caesarean section (18. 5% of all deliveries) compared with previous reports. The proportion of Caesarean sections performed under regional anaesthesia had increased for both elective and emergency Caesarean section deliveries (85.5% and 70.2%, respectively). For pain relief in labour, there had been neither an increase nor a decrease in the uptake of regional analgesia (23.6%). There were limited training opportunities for anaesthetists in general anaesthesia for Caesarean section and for obstetricians in vaginal breech delivery. The known admissions to intensive care units equated to over 100 women per month in the United Kingdom requiring intensive care as a result of childbirth. PMID- 11121926 TI - Inadequate pre-operative evaluation and preparation: a review of 197 reports from the Australian incident monitoring study. AB - The Australian Incident Monitoring Study database was examined for incidents involving inadequate pre-operative patient preparation and/or evaluation. Of 6271 reports, 727 had appropriate keywords, of which 197 (3.1%) were used for subsequent analysis. All surgical categories were represented. In 10% of reports the patient was not reviewed pre-operatively by an anaesthetist, whilst in 23% the anaesthetist involved in the operating theatre had not performed the pre operative assessment. Death followed in seven cases, major morbidity in 23 cases, admission to a high-dependency unit or intensive care unit in 17 cases, and surgery was cancelled in nine cases. Poor airway assessment, communication problems and inadequate evaluation were the most common contributing factors. Respondents indicated that the incident was preventable in 57% of cases. Proposed corrective strategies include improved communication, quality assurance activities, development of protocols and additional training. A structured assessment of the airway, along with improvements in information exchange, patient assessment, and use of clearly defined patient management plans and pathways would prevent most of the incidents reported. PMID- 11121927 TI - Review article: cuff volume and size selection with the laryngeal mask. PMID- 11121928 TI - Evaluation of a spinal needle locking device for use with the combined spinal epidural technique. AB - Success of the 'needle-through-needle' technique for combined spinal epidural analgesia requires the immobilisation of the spinal needle during intrathecal injection. A device that achieves this was evaluated in 200 labouring women, randomly allocated to receive a combined spinal epidural using the CSEcure(R) (SIMS Portex, UK) locking needle or a conventional, non-locking technique. Data collection included the incidence of dural click as the spinal needle penetrated the dura mater, presence of cerebrospinal fluid in the spinal needle hub and the number of technical failures with the spinal component. Successful dural punctures with the spinal needle were similar for locking and non-locking needles (99.0 vs. 98.0%; p = 0.55), despite a small but significant reduction in dural click with the locking needle compared with the non-locking technique (97. 0 vs. 84.7%; p < 0.01). Although not statistically significant, there was a higher number of technical failures, mainly due to spinal needle movement, in the non locking group (9.1 vs. 3.1%; p = 0.08). The locking needle device may be a useful alternative to conventional methods for combined spinal epidural analgesia. PMID- 11121929 TI - Survival from a lethal blood concentration of cyanide with associated alcohol intoxication. AB - We present a patient with a lethal blood concentration of cyanide. Additionally, he was found to have an alcohol blood level of 270 mg. dl-1, but made a complete recovery following administration of the antidotes dimethylaminophenol and thiosulphate. It is postulated that the patient may have been able to detoxify himself as a result of metabolism of cyanide to the non-toxic form, thiocyanate. PMID- 11121930 TI - Continuing medical education by anaesthetists in scotland: activities, motivation and barriers. AB - A postal questionnaire survey was carried out to determine the activities, motivation and barriers to continuing medical education amongst career grade anaesthetists in Scotland. Four hundred and ten consultants and 49 non-consultant career grade anaesthetists were surveyed with a response rate of 84.5%. All respondents had taken part in some educational activities in the past two years. Over 80% had attended 10 or more departmental meetings and over 90% had attended meetings of a Regional society or National meetings. Less than 50% had attended for clinical experience with a colleague and only 20% had done so in another centre. There were trends of changing educational activity with increasing age. The most common motivation was to keep up to date for current clinical duties with keeping up to date for teaching second, but younger consultants were more likely to undertake continuing medical education activities in case their clinical duties changed. Perceived barriers to continuing medical education were similar for internal and external activities but funding was less of a limitation for those working in district general hospitals. There is scope for encouraging activities such as clinical experience with a colleague and a need to explore in greater detail the perception of barriers to continuing medical education and their influence on participation. PMID- 11121931 TI - Ultrasonography of the femoral vessels in the groin: implications for vascular access. AB - The femoral artery and vein are commonly used for access to the circulation. Accidental puncture of one vessel whilst attempting to cannulate the other is a common complication. Identification of relevant surface anatomical landmarks and ultrasonography of both groins was performed on 50 consecutive adult patients admitted to the intensive care unit. In most patients there was overlap of the artery over the vein far closer to the inguinal ligament than conventional anatomical texts would indicate. The frequency and degree of overlap increased as the vessels descended distally towards the knee. Surface anatomical landmarks were not useful in predicting the underlying anatomy. The side-by-side relationship of artery and vein is commonest close to the inguinal ligament. Therefore, to avoid damage to the neighbouring vessel, percutaneous access should be undertaken just below the inguinal ligament. PMID- 11121932 TI - Pharmacokinetics of transdermal fentanyl in the peri-operative period in young children. AB - The pharmacokinetics of transdermal fentanyl were assessed in eight children aged 18-60 months, weighing 11-20 kg and monitored postoperatively in the intensive care unit. A patch, delivering 25 microg.h-1 of fentanyl, was applied for 72 h from the induction of anaesthesia. Plasma fentanyl concentrations were measured over 144 h. Mean (SD) peak concentration of fentanyl was 1.7 (0.66) ng.ml-1 and time to reach maximal plasma concentration was 18 (11) h. The elimination half life was 14.5 (6.2) h, and the area under the curve for plasma fentanyl concentration (0-144 h) was 86.8 (27) ng.h.ml-1. Maximal fentanyl concentration was negatively correlated with patient age (r = - 0.71; p = 0.049) but not with body weight. These results suggest that the pharmacokinetics of transdermal fentanyl in children are similar to those in adults. PMID- 11121933 TI - Evaluation of a pilot regimen for postoperative pain control in patients receiving oral morphine pre-operatively. AB - Postoperative analgesia in patients who receive regular oral opioids pre operatively is frequently suboptimal. To improve management we introduced a regimen using subcutaneous diamorphine infusions with incremental doses. Infusion doses were calculated as half the daily pre-operative dose of oral morphine with the increments as one-sixth of the infusion dose. Results were recorded on the first two postoperative days before (n = 13) and after (n = 23) commencing the new regimen. The percentage of patients reporting severe pain at rest and on movement were significantly reduced by the new regimen (54% and 69% vs. 13% and 40%, respectively) since the opioid dose as a percentage of the pre-operative dose was significantly higher (160% vs. 352%). There were no instances of excessive sedation or slow respiratory rate in any patient. The use of the regimen has resulted in greater doses of opioids being administered with fewer patients in severe pain without significant complications. PMID- 11121934 TI - Noise in the MRI scanner. PMID- 11121935 TI - Date-expired nitrous oxide cylinders. PMID- 11121936 TI - Failure of datex AS/3 anaesthesia delivery unit. PMID- 11121938 TI - Another case of anaesthetic machine failure. PMID- 11121939 TI - Expiratory dates on anaesthetic gas cylinders. PMID- 11121940 TI - Unusual pulse oximetry waveform. PMID- 11121941 TI - A modification to the codmantrade mark microsensortrade mark skull bolt Kit. PMID- 11121942 TI - Cricothyroidotomy comments. PMID- 11121943 TI - Not all that wheezes is asthma. PMID- 11121944 TI - Swelling in the hypopharynx as a cause of difficult tracheal intubation. PMID- 11121945 TI - Retrolaryngeal extension of goitre in a morbidly obese patient leading to a difficult airway. PMID- 11121946 TI - Acute respiratory failure secondary to severe hypokalaemia. PMID- 11121947 TI - Operative tumour handling and hyperkalaemia. PMID- 11121948 TI - Anaphylaxis to rocuronium. PMID- 11121949 TI - Isolated systolic hypertension and anaesthesia. PMID- 11121950 TI - Microbial contamination of gum-elastic bougies. PMID- 11121951 TI - Ventilator trigger setting in an intensive care unit. PMID- 11121952 TI - Use of a carotid arterial line during organ harvest. PMID- 11121953 TI - Preparation of anaesthetic drugs in the obstetric theatre. PMID- 11121954 TI - Epidural anaesthesia and splanchnic blood flow. PMID- 11121956 TI - Pregnancy, anaesthesia and guillain-barre syndrome. PMID- 11121957 TI - Von Willebrand's disease and neuroaxial anaesthesia. PMID- 11121958 TI - Parenteral narcotic analgesic for labour pain. PMID- 11121959 TI - Parenteral narcotic analgesic for labour pain 2. PMID- 11121961 TI - The safest prediction of epidural analgesia. PMID- 11121962 TI - Reducing the incidence of technical failures and paraesthesia in combined spinal epidural techniques. PMID- 11121963 TI - Fixation of epidural catheters. PMID- 11121964 TI - Suturing epidural catheters. PMID- 11121965 TI - Epidural tunnelling techniques. PMID- 11121966 TI - A simple method for testing for the kinking epidural catheter. PMID- 11121967 TI - Value of attachments in A & E to intensive care training. PMID- 11121968 TI - From gasman to hospital saviour: how far should we go? PMID- 11121969 TI - 'For the times they are a-changing' - or are they? PMID- 11121970 TI - The use of electrolyzed solutions for the cleaning and disinfecting of dialyzers. AB - Recently, the use of electrolyzed solutions has attracted considerable interest in Japan. This study investigates the efficiency of electrolyzed solutions as disinfecting agents (DA) in the reuse of dialyzers and compares their efficiency to that of other disinfectants currently in use. The following 3 methods were employed. First, the rinsing time and rebound release of reused dialyzers were measured and compared after electrolyzed solutions, electrolyzed strong acid aqueous solution (ESAAS) and electrolyzed strong basic aqueous solution (ESBAS), made from reverse osmosis (RO) water (ESAAS, ESBAS; Generating apparatuses: Super Oxseed alpha 1000, Amano Corporation, Yokohama, Japan), 2% Dialox-cj (Teijin Gambro Medical, Tokyo, Japan), and 3.8% formalin were used as DAs. This involved performing dialysis with 2 types of dialyzers: a cellulose acetate membrane (CAM) dialyzer and a polysulfone membrane (PSM) dialyzer. The dialyzers were cleaned and disinfected using the different DA and left for 48 h. Next, after performing dialysis the dialyzer membranes were cleaned with a saline solution (0.9% NaCl) and RO water and then cleaned with the various DA. These membranes were observed using a scanning electron microscope (SEM) to check for the presence of physical and biological contaminants. Finally, in vitro tests were performed to determine the level of dialyzer clearance when PSM dialyzers were reused after having been cleaned and disinfected with the electrolyzed solutions. The rinsing time results for both the CAM and PSM dialyzers showed the electrolyzed solutions (ESBAS and ESAAS) as being undetectable within 10 min. With regard to the rebound release, for both the CAM and PSM dialyzers, the electrolyzed solutions were undetectable at all checking times between 30 and 240 min. Observation by SEM showed that cleaning with both ESAAS and ESBAS left the fewest contaminants, and cleaning with 2% Dialox-cj left the highest level of contaminants in the CAM dialyzers. With regard to experiments concerning use in vitro, no major changes in the dialyzer clearance were noticed after 6 uses. In every experiment, the previous investigations showed the electrolyzed solutions to be superior to 3. 8% formalin and 2% Dialox-cj DA for the reuse of dialyzers. PMID- 11121971 TI - Late symptomatic venous stenosis in three hemodialysis patients without previous central venous catheters. AB - It is well known that catheters placed in the subclavian or internal jugular veins may develop stenosis in the vein in which the catheter lies. Because the arteriovenous fistula (AVF) relies on good venous outflow, patients with ipsilateral central venous stenosis are subject to the malfunctioning of AVF. Until now, no data were published on patients showing central vein stenosis (CVS) without a previous central venous catheter (CVC) or a pacemaker. In this article, we report on 3 hemodialysis patients manifesting CVS ipslateral to AVF. None of these patients previously had undergone CVC. The stenosis observed had characteristics and symptoms similar to those observed in stenoses consequent to CVC. We concluded that CVS also may occur in subclavian or axillary veins proximal to a working AVF in hemodialysis patients who have never had a CVC and in the absence of compressive phenomena. PMID- 11121972 TI - Safe and efficient gene transfer into porcine hepatocytes using Sendai virus cationic liposomes for bioartificial liver support. AB - Establishment of a bioartificial liver support system using genetically modified hepatocytes is a potential approach to improve the treatment of severe liver failure. We describe the development of an efficient ex vivo method of gene transfer into a large number of porcine hepatocytes using hemagglutinating virus of Japan (HVJ)-liposome. The transfection efficiency of HVJ-liposome into isolated porcine hepatocytes attached to microcarrier beads was evaluated by beta galactosidase (beta-gal) staining, fluorescence activated cell sorting analysis for beta-gal and luciferase assay, respectively. To examine the function and cellular damage of transduced hepatocytes, we used enzyme-linked immunosorbent assay for porcine albumin synthesis, lidocaine clearance test (P-450 activity), aspartate aminotransferase, and lactic dehydrogenase release assays. The optimal conditions for gene transfer into the beads-attached hepatocytes using HVJ liposome included 4 microg of deoxyribonucleic acid with 200 microg of lipid/2 x 105 cells and exposure duration of 90 min. Under these conditions, beta-gal and luciferase genes were transduced to 2.5 x 108 isolated porcine hepatocytes following attachment to the beads. Positive beta-gal staining was observed in more than 30% of the beads-attached hepatocytes. The gene transfer activity of HVJ-liposome method determined by luciferase activities was about 100-fold of that of the lipofection method. Transfected porcine hepatocytes remained functional without any significant cell damage. Our results demonstrated that HVJ liposome mediated gene transfer into microcarrier-attached porcine hepatocytes is an efficient and nontoxic method suitable for a bioartificial liver support sytem. PMID- 11121973 TI - DegraPol-foam: a degradable and highly porous polyesterurethane foam as a new substrate for bone formation. AB - Bone morphogenetic protein (BMP) is known to require a suitable carrier to induce ectopic bone formation in vivo. To evaluate the suitability of DegraPol-foam, a degradable, elastic, and highly porous polyesterurethane foam as carrier for BMP induced bone formation, a fraction containing all the active BMPs (BMP cocktail) was combined with DegraPol-foam and implanted subcutaneously into rats. DegraPol BMP scaffolds were found to induce osteogenesis 2 weeks after implantation as evidenced by morphological and biochemical observations. In addition, the osteoblast-compatibility of DegraPol-foam was examined here. In vitro, primary rat osteoblasts and osteoblasts from the human cell line (HFO1) attached and proliferated preferentially on the surface of the DegraPol-foam. Both cell types exhibited relatively high attachment and low doubling time that resulted in a confluent cell multilayer with spindle-shaped morphology on the surface of the foam. Osteoblasts produced high concentrations of collagen type I and osteocalcin, and expressed increasing levels of alkaline phosphatase (ALP) activity. Taken collectively, both osteoblasts from rat tibia and from the human cell line HFO1 showed high cell attachment and growth, and preserved their phenotype. The geometrical structure of DegraPol is a suitable carrier for BMP for the induction of bone formation. PMID- 11121974 TI - Particle image velocimetry analysis of the flow field in the total cavopulmonary connection. AB - The total cavopulmonary connection (TCPC) is a common operation, meant to restore a proper pulmonary blood flow in heart defects with only one functional ventricle. It consists of the direct connection of the venae cavae to the pulmonary arteries in a cross-shaped disposition which entails a peculiar hemodynamics: Side effects can occur, such as recirculation zones and pressure drop across the connection. Our study is aimed at the quantitative investigation of the flow field of a successful Fontan-type operation, in view of the clinical importance of assuring a nearly physiological pulmonary blood flow, especially if one considers that many pediatric patients are eligible for this operation. A glass-blown TCPC phantom, realized according to nuclear magnetic resonance data, was employed in a steady-flow loop. Thus, a realistic model of this Fontan-type operation was realized using materials which enable advanced measurement techniques such as particle image velocimetry (PIV). The mean flow rates at each branch of the cavopulmonary shunt could be independently varied with a vertical shift of the corresponding upstream reservoir. The PIV technique was used successfully in identifying the flow field characteristics. The flow field in this TCPC topology was shown to be well organized and regulated by the presence of a vortex at the confluence of the venae cavae. The effect of different loading conditions, which realistically can be found in vivo, is studied with a high spatial resolution, showing the possibility to use pulmonary resistance as a parameter in designing the surgical geometry. PMID- 11121975 TI - Changes in patterns of left ventricular hypertrophy after aortic valve replacement for aortic stenosis and regurgitation with St. Jude Medical cardiac valves. AB - In this study, we analyzed the extent and pattern of regression of left ventricular (LV) hypertrophy after aortic valve replacement in patients with aortic stenosis (AS) and compared the results with those of another group of patients with aortic regurgitation (AR). Seventy patients who underwent isolated aortic valve replacement were divided into 2 groups. Group 1 was comprised of 29 patients who underwent aortic valve replacement for aortic stenosis, and Group 2 of 41 patients who underwent aortic valve replacement for aortic regurgitation. A third group of 10 healthy subjects served as a healthy control group. Echocardiographic studies were done before the operation and 5 years postoperatively. At follow-up, a significant reduction in the left ventricular mass was found in both groups, but it remained significantly greater than in the healthy control group. The ratio of LV wall thickness to radius (th/r) in Group 1 decreased significantly, and at follow-up it was within the normal value. In Group 2, the th/r ratio increased, and at follow-up it was within the normal value. After aortic valve replacement, the wall thickness remained significantly greater than normal for patients with AS, and the chamber radius remained significantly greater than normal for patients with AR. For these reasons, LV hypertrophy still existed in both groups at postoperative follow-up. The actuarial survival rate was 85.3% at 16 years for Group 1 and 83.4% at 18 years for Group 2. There was no significant difference in the long-term survival rates between the 2 groups. Actuarial freedom from valve-related events was 91.9% at 16 years for Group 1 and 82% at 18 years for Group 2. There was no significant difference in the valve-related event free curves between groups. After 5 years of follow-up, th/r reached normal for both groups, indicating remodeling of the LV geometry after aortic valve replacement. PMID- 11121976 TI - Hemodynamic performance of small-size bileaflet valves: pressure drop and laser Doppler anemometry study comparison of three prostheses. AB - Laser Doppler anemometry (LDA) is a single-point technique which is unparalleled to detect accurately the local properties of the velocity field in a turbulent flow, such as that generated by a prosthetic heart valve (PHV). We propose a correlation between the structure of the flow field in three 19 mm bileaflet PHVs (Sorin Bicarbon, St. Jude Standard, St. Jude HP), investigated at peak systole (6 L/min cardiac output [CO]) with LDA, in kinematic and geometric similarity, and the global parameter of transvalvular pressure drop measured in both steady and pulsatile conditions. The pressure transducers of the same apparatus were used to characterize pressure drops at different flow rates whereas the steady-flow case was studied with a highly accurate tester built in our laboratory. The 2 St. Jude models rank according to their internal orifice diameter (ID) with the standard model (with a smaller ID) providing higher pressure drops for each flow rate. Sorin Bicarbon, due to its leaflet geometry, generates a more complex flow field with respect to the 2 St. Jude flat-leaflet models and shows improved hemodynamical behavior in pulsatile conditions with respect to the stationary case due to differences in pressure recovery. This study can provide insights into a PHV's local flow structure and global hemodynamical parameters. PMID- 11121977 TI - Electronmicroscopic changes in selected vital organs after long-term total artificial heart pumping in animal experiments. AB - Electronmicroscopic (EM) evaluation of selected vital organs (liver, kidney, lung) informs us about otherwise hardly detectable changes during total artificial heart (TAH) pumping. In our experiments, we compared 2 groups of long surviving animals in which the TAH TNS-BRNO pneumatic device was implanted (TNS BRNO-II and VII in 45 experiments, TNS-BRNO-III in 1 experiment, and TNS-BRNO VIII in 1 experiment). In 4 experiments, the Rostock TAH (NABEL, TAH Research Center, Rostock, Germany) was implanted. One group of 22 animals with an average survival of 162 days (the longest, 293 days) was submitted to an antihypertensive treatment; another 1 of 29 calves with an average survival of 98 days (the longest, 173 days) was untreated. The evaluation was performed using a scale (0 to 3) based on very precisely fixed criteria. EM pathologic changes documented various stages of ischemic damage. Except for the liver, no significant difference was found between both groups, despite the substantially prolonged survival in the treated group. Very important was the general state of mitochondria, endoplasmic reticulum, and nucleus. Further, the state of glomerular podocytes in the kidney and the state of interalveolar septa and of pneumocytes constituting the air-blood barrier for gas exchange in the lungs are especially important. In some animals of both groups, the EM findings were completely normal, especially in the lung. PMID- 11121978 TI - Current leakage in hemodialysis machines may be a safety risk for patients. AB - During hemodialysis, the dialysis fluid is conductive and allows current to flow from or to the machine if a difference in electrical potential occurs. This may cause a health risk for the patient. We investigated the presence of such current during various conditions in 17 dialysis machines: the Gambro AK10 (n = 5) and AK100 (n = 3), and the Fresenius 2008C (n = 3), 2008E (n = 2), and 4008E (n = 4). Current leakage measurements were performed without connection of the patient to the system by using a copper tube inserted into the stream of the dialysis fluid at the point where the patient would normally be connected to the dialyzer. The current leakage (measured in microA) at the ground site was significantly higher for the AK10 and the 2008C than for the AK100, 2008E, and 4008E machines. The current leakage, at the site where the dialyzer is located, was highest for the AK10 and less for the AK100, 2008C, 2008E, and 4008E. The 2008C and 2008E dialyzers had the highest maximal current leakage when a malfunctioning external device was attached while the AK100 had less and the AK10 had the least leakage. There was a great variation in leakage current among the dialysis machines. The greatest risk occurred when the patient also was connected to other electric devices with current leakage. PMID- 11121979 TI - The effect of biofiltration on red blood cells 2.3-diphosphoglycerate and pH. AB - To investigate the effect of biofiltration (BF) on the ability of blood to supply oxygen to the peripheral tissues, a 2 week crossover study was conducted with bicarbonate hemodialysis (BcHD) and BF using 5 male patients with diabetic renal failure as subjects. BcHD and BF were performed for 4 h and 3.5 h per session, respectively. Blood gases, the pH of red blood cells (RBC-pH), and 2. 3 diphosphoglycerate in RBC (RBC-2.3DPG) were measured during each treatment. After a 2 week BF treatment, the plasma HCO3- at the beginning of BF was significantly higher than that of BcHD (p < 0.01), and the blood pH improved with an elevated plasma bicarbonate level (p < 0.05). The RBC-pH at the beginning of BF was higher than that of BcHD (p < 0.05) although the RBC-pH at the end of both therapies increased to similar levels. The RBC-2.3DPG during BcHD remained unchanged, but during BF significantly increased (p < 0.05). Metabolic acidosis was significantly improved by BF with its effect reaching to the RBC intracellular level. The improved metabolic acidosis might occur as a result of the increase in RBC-2.3DPG during BF. This increase in RBC-2.3DPG has the effect of reducing the affinity of oxygen for hemoglobin and allows more oxygen to be delivered to the peripheral tissues although the increase in RBC-pH by dialysis restricts the dissociation of oxygen from hemoglobin. PMID- 11121980 TI - Effect of electrolyzed water on wound healing. AB - Electrolyzed water accelerated the healing of full-thickness cutaneous wounds in rats, but only anode chamber water (acid pH or neutralized) was effective. Hypochlorous acid (HOCl), also produced by electrolysis, was ineffective, suggesting that these types of electrolyzed water enhance wound healing by a mechanism unrelated to the well-known antibacterial action of HOCl. One possibility is that reactive oxygen species, shown to be electron spin resonance spectra present in anode chamber water, might trigger early wound healing through fibroblast migration and proliferation. PMID- 11121981 TI - Hypotransferrinemia in chronic hemodialyzed (HD) patients. PMID- 11121983 TI - EDITOR'S COMMENT. PMID- 11121982 TI - Preliminary results of a randomized controlled trial of prophylactic shock wave lithotripsy for small asymptomatic renal calyceal stones. AB - OBJECTIVE: To report a prospective, randomized study to determine whether prophylactic extracorporeal shockwave lithotripsy (ESWL) is justified as a treatment for small, asymptomatic calyceal stones. PATIENTS AND METHODS: The study included 228 patients with small (< 15 mm total diameter) asymptomatic calyceal stones; 113 patients were randomized to undergo ESWL and 115 to the control group who were kept under observation. Outcome measurements included the stone-free rate, requirement for additional treatment, symptoms, quality of life and renal function. RESULTS: In all, 200 patients had at least one annual follow up; all outcome measurements reported were those at the most recent follow-up (mean 2.2 years, range 1-5). In the ESWL group 28 patients (28%) were stone-free, compared with 16 (17%) in the observation group (odds ratio 1.95, 95% confidence interval, CI, 0.97-3.89, P = 0.06). Additional treatment in the form of analgesics, antibiotics, ESWL, stent insertion and ureteroscopy, was required in 21 (21%) patients in the observation group and 15 (15%) in the ESWL group (odds ratio 0.66, 95% CI 0.32-1.37, P = 0.27). Ten patients in the observation group required invasive procedures, vs none in the ESWL group. There was no evidence of differences in the symptoms, quality of life or renal function tests between the arms of the trial at the final follow-up. CONCLUSIONS: Prophylactic ESWL for small asymptomatic renal calyceal stones does not appear to offer any advantage to patients in terms of stone-free rate, quality of life, renal function, symptoms or hospital admissions. However, a policy of observation is associated with a greater risk of requiring more invasive procedures. A longer follow-up is required to assess the validity of these preliminary findings. PMID- 11121984 TI - Renal artery occlusion. PMID- 11121985 TI - Are urologists prepared for latex-allergic patients? AB - OBJECTIVE: To determine how prepared operating departments are to manage latex allergic patients, and particularly urologists, because the highest incidence of latex allergy occurs in patients with spina bifida who undergo frequent urological procedures. METHODS: A standard questionnaire about the provision of latex-free equipment within operating theatres was completed by the 72 hospitals surveyed (33 district general, 27 teaching, six children's and six private hospitals). RESULTS: Anaesthetic latex-free equipment was available in five of the children's hospitals, 56% of the teaching hospitals, 45% of the district general hospitals and two of the private hospitals. A latex-free catheter only was provided in two of the children's hospitals, 30% of the teaching hospitals and 12% of the district general hospitals. In addition, one of the children's hospitals and one of the private hospitals had a comprehensive range of urological latex-free equipment available. CONCLUSION: This survey shows that many hospitals are inadequately prepared to manage latex-allergic patients, because they lack anaesthetic and urological latex-free equipment. PMID- 11121986 TI - Sacral nerve root stimulation for lower urinary tract dysfunction: overcoming the problem of lead migration. AB - OBJECTIVE: To evaluate lead migration for two different test electrodes and the response to trial stimulation of the S3 nerve root during the selection of patients for a sacral neuromodulation implant to manage lower urinary tract dysfunction. PATIENTS AND METHODS: Twelve women (mean age 49 years, range 23-79; seven with detrusor instability and five with sensory urgency) undergoing peripheral nerve evaluation for refractory lower urinary tract symptoms were recruited. Urodynamics and a urinary diary were completed before and during test stimulation. Two electrodes (the original 041830-002 and new 3057 models, Medtronics Inc, USA) were inserted under local anaesthesia into the S3 nerve roots bilaterally. The location was determined by the functional response to stimulation. Stimulation was applied for one week using the new lead; a positive response was defined as a subjective improvement (> 50%) in urinary symptoms. Lateral sacral X-rays were taken after placement and before removing the lead. The distance from the lead tip to the ventral aspect of the S3 sacral foramen was measured by two assessors. RESULTS: Ten of the women had a positive response; the mean (range) migration of the new lead (on X-ray) was 4 (2-11) mm, and of the old lead was 12 (10-45) mm (P = 0.02). CONCLUSION: The response rate to trial stimulation was greater than in previous studies, possibly reflecting reduced migration of the new lead. The new electrode may reduce the number of test failures caused by lead migration rather than no response. PMID- 11121987 TI - Pressure-flow studies in benign prostatic hyperplasia: to do or not to do for the patient? PMID- 11121988 TI - Tamsulosin in men with confirmed bladder outlet obstruction: a clinical and urodynamic analysis from a single centre in New Zealand. AB - OBJECTIVE: To evaluate the clinical and pressure-flow effects of tamsulosin 0.4 mg once daily in patients with lower urinary tract symptoms (LUTS) caused by benign prostatic obstruction (BPO), as documented by pressure-flow studies performed according to a standardized protocol in one centre. PATIENTS AND METHODS: A single-centre study was conducted with an open-label design comprising a 2-week, single-blind, placebo run-in period and a 12-week active treatment period with tamsulosin 0.4 mg once daily. Eligible patients were men (aged > or = 45 years) with LUTS (total International Prostate Symptom Score, IPSS, > or = 13) with a free-flow maximum urinary flow rate (Qmax) of > or = 4 mL/s for a voided volume of > or = 120 mL. After the 2-week placebo run-in period, only patients with BPO (i.e. in the obstructed zone of the Abrams-Griffiths, AG, nomogram) received active treatment with tamsulosin. The two primary efficacy variables were the change in detrusor pressure at maximum flow (PdetQmax) and the total IPSS, from baseline to endpoint. A standardized protocol was used for all pressure-flow studies. RESULTS: In all, 42 patients were enrolled into the 2-week placebo run-in period, after which 30 received active treatment with tamsulosin 0.4 mg once daily. All 12 patients (29%) who discontinued during the placebo run in period failed to fulfil the pressure-flow entry criterion of confirmed obstruction. The 30 patients who received tamsulosin had a high mean PdetQmax (82.1 cm H2O) and high mean AG number (68.1) at baseline, as would be expected in obstructed patients. Tamsulosin produced a significant reduction in mean PdetQmax (-10.6 cm H2O or -13%; P = 0.005 vs baseline). The mean AG number decreased accordingly (-15.5 points or -23%; P < 0.001 vs baseline). The pressure-flow mean Qmax was increased by 2.5 mL/s (36%) from 7.0 mL/s at baseline (P = 0.002 vs baseline). Urodynamic improvements were accompanied by a good symptomatic response; the mean total IPSS was reduced by 6.7 points from a baseline value of 18.1 (-37%, P < 0.001 vs baseline). At the endpoint, 67% of patients had a clinically significant symptomatic response (defined as a decrease in total IPSS of > or = 25% from baseline). Twenty-six patients (87%) were either slightly improved (13) or were much improved (13) in the opinion of the investigator. Twelve patients with LUTS who were unobstructed after the 2-week placebo run-in (PdetQmax 44.1 cm H2O) received tamsulosin treatment outside of the study protocol. Although these patients had no decrease in mean PdetQmax, the magnitude of the symptomatic response (decrease in total IPSS -6.4 or -34%, P = 0.001) was comparable with that in unobstructed patients; the free-flow Qmax was also significantly improved. Possibly or probably drug-related adverse events were reported in nine patients; one discontinued tamsulosin treatment because of a serious adverse event. CONCLUSION: Tamsulosin 0.4 mg once daily can produce a significant decrease in detrusor pressure, increase in flow rate and a symptomatic improvement in patients with LUTS and confirmed obstruction. Patients with LUTS who are unobstructed and have a low initial detrusor pressure appear to have no improvement in detrusor pressure, but have similar clinical responses to those in obstructed patients. Pressure-flow studies can be reserved for those patients with LUTS who fail to respond to these agents and in whom further diagnosis and management is warranted. PMID- 11121989 TI - Does the time of administration (morning or evening) affect the tolerability or efficacy of tamsulosin? AB - OBJECTIVE: To determine whether the time of dosing (morning or evening) affects the tolerability or efficacy of tamsulosin in the treatment of lower urinary tract symptoms. PATIENTS AND METHODS: Data were analysed from an open-label, observational study in which patients were treated with 0.4 mg tamsulosin once daily for 12 weeks. Treatment effects were determined using the Benign Prostatic Hyperplasia Impact Index, the quality-of-life question of the International Prostate Symptom Score, a similarly phrased question about sexual satisfaction, the maximum urinary flow rate, the postvoid residual urine volume, and the overall efficacy and tolerability. The results were analysed statistically for differences between dosing times, using analysis of covariance for the quantitative variables and logistic regression for the qualitative variables. RESULTS: While no specific recommendation about the dosing time was given in the trial, the retrospective analysis showed that 4420 and 2087 patients received tamsulosin in the morning and evening, respectively. Both groups had similar values for all variables before treatment. The efficacy and tolerability of tamsulosin treatment was also similar in both groups; there were small advantages for morning dosing, which were statistically significant because there were many patients. CONCLUSION: In contrast to other alpha-blockers, night-time dosing is not necessary to improve the tolerability or efficacy of tamsulosin. PMID- 11121990 TI - Urinary levels of substance P and its metabolites are not increased in interstitial cystitis. AB - OBJECTIVES: To determine whether interstitial cystitis is associated with the increased release of substance P from the bladder wall into urine, by measuring urinary excretion rates of substance P and its metabolites in women with interstitial cystitis and in a control group of women with stress incontinence and normal bladder function. PATIENTS AND METHODS: Catheter urine was collected from 13 patients and 10 controls during a water diuresis ( approximately 10 mL/min) before and after instilling the bladder with 100 mL of water. The contribution of the bladder wall to urinary substance P peptides was assessed by measuring the change in substance P peptide levels after 2 min of bladder stasis before and after instillation. RESULTS: Absolute substance P excretion rates were similar in patients with interstitial cystitis and controls; 2 min of bladder stasis reduced the substance P excretion rate (P = 0.03) and increased the excretion rate of substance P metabolites (P = 0.01). CONCLUSIONS: The release of substance P from the bladder wall was not increased in patients with interstitial cystitis. PMID- 11121992 TI - Nonsteroidal antiandrogens: a therapeutic option for patients with advanced prostate cancer who wish to retain sexual interest and function. PMID- 11121991 TI - Imaging for staging bladder cancer: a clinical study of intravenous 111indium labelled anti-MUC1 mucin monoclonal antibody C595. AB - OBJECTIVE: To investigate the clinical application of an 111In-labelled anti-MUC1 mucin monoclonal antibody (mAb) imaging for staging invasive bladder cancer. PATIENTS AND METHODS: Indirect immunohistochemistry was used to confirm the expression of the MUC1 target antigen by metastatic tumours. Twelve patients with bladder cancer (two with superficial and 10 with locally invasive/metastatic disease) underwent planar gamma-scintigraphy 48 h after an intravenous injection with 111In-labelled anti-MUC1 mucin mAb C595. RESULTS: No bladder uptake was detected in the two patients with superficial disease, but scintigraphy showed primary and recurrent bladder tumours and metastases in nine of the remaining 10 patients with invasive disease. In three patients additional staging information was obtained from the mAb imaging which would have altered patient management. There were no reported side-effects. CONCLUSION: This study confirmed the ability of the mAb technique to detect both primary and recurrent invasive bladder tumours and distant metastases. Some lesions shown by mAb imaging were not detected by other methods. The use of mAb imaging has the potential to improve clinical staging and assist in selecting those patients most likely to benefit from radical therapy. PMID- 11121993 TI - Annual changes in the clinical features of prostatic adenocarcinoma in Taiwan. AB - OBJECTIVE: To determine, using a clinical and pathological database, the annual changes in the characteristics of adenocarcinoma of the prostate diagnosed in consecutive men in Taiwan. PATIENTS AND METHODS: All prostatic adenocarcinomas newly diagnosed between 1991 and 1999 in our institution were analysed. Using logistic regression for dichotomous variables, the annual trends were assessed for changes in the percentage of T1c disease, incidental carcinoma, clinically localized disease and Gleason grade 4/5 cancers. The annual trends for changes in the pathological features of disease in patients treated by radical prostatectomy were also analysed. RESULTS: The study included 694 patients; the median age at diagnosis increased significantly (P = 0.004) and there was a significant increase in clinical T1c disease (relative risk, RR, 1.142, P = 0.022). There were also significant decreases in incidental carcinomas (RR 0.789, P < 0.001), patients with a serum prostate-specific antigen (PSA) level of > 20 ng/mL (RR 0.848, P < 0.001) and with Gleason grade 4/5 tumours (RR 0.919, P = 0.005). There was no significant change in the percentage of clinically localized disease. Radical prostatectomy was undertaken in 179 men; the annual incidence of lymph node metastases decreased significantly (P = 0.035) and there was a significant 19% increase in the RR of having organ-confined disease (pT1-2N0M0) and a 19% decrease in Gleason grade 4/5 tumours. CONCLUSIONS: These results may reflect the annual trends of prostate cancer in Taiwan in the period for which the PSA assay became available. The increasing application of PSA testing was associated with a significant increase in clinical T1c disease and decrease in incidental carcinoma. There was also a significant trend towards less aggressive cancers. Despite the dramatic increase in the annual detection rate, tumours were not detected at an earlier stage. However, for patients treated with radical prostatectomy, there was a significant change to earlier stage disease and fewer de-differentiated cancers. PMID- 11121994 TI - A comparison of radical retropubic with perineal prostatectomy for localized prostate cancer within the Uniformed Services Urology Research Group. AB - OBJECTIVE: To review and compare the outcome of patients undergoing radical retropubic prostatectomy (RRP) or radical perineal prostatectomy (RPP) for clinically localized prostate cancer. PATIENTS AND METHODS: From 1988 to 1997, 1382 men who were treated by RRP and 316 by RPP were identified from databases of the Uniformed Services Urology Research Group. The following variables were assessed; age, race, prostate-specific antigen (PSA) level before surgery, clinical stage, biopsy Gleason sum, estimated blood loss (EBL), margin-positive rate, pathological stage, biochemical recurrence rate, short and long-term complication rates, impotence and incontinence rates. To eliminate selection bias, the analysis was concentrated on pairs of patients matched by race, preoperative PSA level, clinical stage and biopsy Gleason sum. RESULTS: In the 190 matched patients there were no significant differences between the RRP and RPP groups in either organ-confined (57% vs 55%), margin-positive (39% vs 43%), or biochemical recurrence rates (12.9% vs 17.6% at a mean follow-up of 47.1 vs 42.9 months), respectively. The mean EBL was 1575 mL in the RRP group and 802 mL in the RPP group (P < 0.001). The only significant difference in complication rates was a higher incidence of rectal injury in the RPP group (4.9%) than in the RRP group (none, P < 0.05). CONCLUSIONS: In similar populations of patients, RPP offers equivalent organ-confined, margin-positive and biochemical recurrence rates to RRP, while causing significantly less blood loss. PMID- 11121995 TI - Limited value of endorectal magnetic resonance imaging and transrectal ultrasonography in the staging of clinically localized prostate cancer. AB - OBJECTIVE: To examine the role of endorectal magnetic resonance imaging (eMRI) and transrectal ultrasonography (TRUS) for clinically localized prostate cancer and to assess interobserver agreement in interpreting MRI studies. PATIENTS AND METHODS: Fifty-four patients with biopsy-confirmed prostate cancer underwent TRUS and eMRI before radical retropubic prostatectomy. The MR images were prospectively interpreted by two radiologists with special expertise in this field. The criteria evaluated prospectively in each patient were extracapsular extension (ECE) and seminal vesicle invasion (SVI). The results were correlated with the histopathological findings after radical prostatectomy. RESULTS: At pathology, 27 patients had stage pT2, 15 had stage pT3a and 12 had stage pT3b lesions. The overall accuracy of eMRI in defining local tumour stage was 93% by radiologist A and 56% by radiologist B; the overall accuracy by TRUS was 63%. There was a poor correlation for the MRI studies between observers. The eMRI was more sensitive than TRUS for detecting ECE and SVI in organ-confined prostate cancer. TRUS had a relatively high specificity for ECE and SVI, and was better than eMRI in this regard. CONCLUSION: Whereas MRI tended to over-stage, TRUS under-staged prostate cancer. This series shows the current limited value of TRUS and eMRI for planning treatment in patients with clinically localized prostate cancer. Treatment decisions should not be altered based on TRUS or eMRI findings alone. PMID- 11121996 TI - A dose-escalation study to assess the efficacy and safety of sildenafil citrate in men with erectile dysfunction. AB - OBJECTIVE: To assess the efficacy and safety of sildenafil citrate (Viagra, Pfizer Inc., USA) in a double-blind, placebo-controlled, dose-escalation study over a period of 26 weeks in men with erectile dysfunction of a broad spectrum of aetiology. PATIENTS AND METHODS: In all, 315 patients from five countries were randomized to receive treatment with placebo (156 men) or sildenafil (159 men). Significant concomitant medical conditions were hypertension (20%), a history of pelvic surgery (19%), diabetes mellitus (15%), and ischaemic heart disease (10%). Patients randomized to treatment received a starting dose of 25 mg of sildenafil or matching placebo, which could be increased to 50 mg and then to 100 mg of sildenafil, based on efficacy and tolerability. Assessments of efficacy comprised the 15-item International Index of Erectile Function (IIEF), including question three (ability to achieve an erection) and question four (ability to maintain an erection), a partner questionnaire, an overall efficacy question, and event-log data. RESULTS: After 12 weeks of treatment, 26%, 32% and 42% of patients were taking 25, 50 and 100 mg of sildenafil, respectively. A similar distribution of doses was reported after 26 weeks of treatment. Treatment with sildenafil significantly improved the patients' abilities to achieve and maintain an erection compared with treatment with placebo (P < 0.001). Scores for four of the five sexual function domains of the IIEF (erectile function, orgasmic function, intercourse satisfaction and overall satisfaction) also improved significantly (P < 0.001). There was a significant improvement in the mean score for the erectile function domain, regardless of the aetiology of erectile dysfunction (P < 0.001). After 12 weeks and 26 weeks of treatment, 82% and 79% of patients receiving sildenafil reported improved erections, compared with 24% and 23% of patients receiving placebo, respectively (P < 0.001). Treatment-related adverse events were mild to moderate and occurred in 27% of patients receiving sildenafil, compared with 8% of patients receiving placebo. CONCLUSION: Sildenafil is an effective and well-tolerated treatment for men with erectile dysfunction of a broad spectrum of aetiology. PMID- 11121997 TI - The excisional, plication and internal drainage techniques: a comparison of the results for idiopathic hydrocele. AB - OBJECTIVE: To assess the results of the excision, plication and internal drainage techniques for hydrocele repair. PATIENTS AND METHODS: Between January 1990 and June 1998, 132 patients (mean age 54.36 years, range 16-83) underwent repair for idiopathic hydrocele using one of three techniques (excision, eversion/plication or internal drainage); the complication and recurrence rates of each technique were evaluated. RESULTS: The excisional technique resulted in the highest complication rate (81%) and the internal drainage technique the lowest (7%). Postoperative scrotal oedema occurred in 74% of patients after plication and this was the highest rate among the techniques (P < 0.001). Differences in the rates of wound infection and haematoma among the three techniques were not statistically significant. The internal drainage technique had the highest recurrence rate (85%) and the excisional technique the lowest (1.3%; P < 0.001). CONCLUSIONS: Although useful, the internal drainage technique has a high recurrence rate and we suggest abandoning its use for hydrocele repair. The present results suggest that plication is better than excision, causing fewer complications, and better than internal drainage, as the results are more favourable. PMID- 11121998 TI - Expression of endothelial nitric oxide synthase in the Sertoli cells of men with infertility of various causes. AB - OBJECTIVE: To investigate how endothelial nitric oxide (eNOS) expression in the seminiferous tubules might be related to spermatogenesis, by examining eNOS expression in testicular tissue of patients infertile from various causes. PATIENTS AND METHODS: The study included five fertile men with a normal sperm concentration, nine patients with obstructive azoospermia, 20 with varicocele testes and eight with idiopathic azoospermia (Sertoli cell-only syndrome). Testicular biopsy specimens were examined by immunohistochemistry for eNOS protein expression, in addition to a routine pathological assessment. eNOS protein was detected using an eNOS monoclonal antibody. A Sertoli cell staining index (SSI) was defined as the ratio of stained Sertoli cells per total number of Sertoli cells, and was compared among the groups. RESULTS: eNOS was localized to Sertoli cells in the seminiferous tubules and Leydig cells in the interstium; although some degenerating germ cells stained, normal germ cells did not. The SSI was significantly lower in patients with Sertoli cell-only syndrome than in either fertile men or patients with obstructive azoospermia or varicocele. However, the SSI did not correlate significantly with the Johnsen score. CONCLUSION: The expression of eNOS in Sertoli cells may depend on the existence of germ cells and be associated with germ cell development. PMID- 11121999 TI - Testicular tissue bleeding as an indicator of gonadal salvageability in testicular torsion surgery. AB - OBJECTIVE: To investigate the reliability of using bleeding from the cut surface of testicular tissue during surgery for testicular torsion to assess testicular viability, compared with the duration of symptoms and preoperative findings on testicular Doppler ultrasonography (DUS). PATIENTS AND METHODS: The study comprised 19 children with testicular torsion who underwent surgery; all underwent DUS before surgery. During surgery the tunica vaginalis of the affected gonad was incised and a deep incision made through the medulla after obtaining a wedge biopsy for histological examination. After waiting up to 10 min to assess any fresh arterial bleeding from the cut surface, the patients were categorized using three grades; grade I (sufficient bleeding, i.e. bleeding or oozing when the biopsy was obtained); grade II (insufficient bleeding, no bleeding immediately after the incision but starting within 10 min); and grade III (no bleeding within 10 min). The final surgical decision on whether to save the testis was made according to the grade of testicular tissue bleeding; grade I and II testes were saved and grade III testes were removed. The biopsies were histopathologically examined and classified as haemorrhagic, necrotic or indeterminate. The patients were followed up at 15 days and at 1, 3, 6 and 12 months, with the affected testis examined using DUS. At the end of the study, the sensitivity and specificity of the duration of symptoms, characteristics of blood flow on DUS and grading of testicular tissue bleeding at surgery were calculated for predicting testis viability, using the histopathological diagnosis as the reference standard. RESULTS: The sensitivity, specificity, positive and negative predictive values were respectively 100%, 90%, 90% and 100% for a duration of symptoms of > 10 h, 78%, 80%, 78% and 80% for DUS findings, and 100%, 78%, 83% and 100% for testicular tissue bleeding in predicting gonad viability after torsion, respectively. CONCLUSION: Although the 10 h limit for the duration of symptoms seems a more accurate predictor of the fate of a twisted testis than were the other variables, testicular tissue bleeding may also be a good indicator of gonadal viability during surgery. The surgeon should wait up to 10 min after incising the testicular tissue deep to the medulla before deciding the type of surgery. In cases where bleeding from the cut surface is sufficient or insufficient (according to the proposed grading system), orchidopexy is the treatment of choice. The salvaged testes should be assessed during follow-up, especially in those who had had insufficient bleeding at surgery and/or a duration of symptoms > 10 h, to assess for any delayed damage to the untwisted testis. If no bleeding is seen during surgery the best option is to remove the affected testis. PMID- 11122000 TI - Direct comparison of radiology and nuclear medicine cystograms in young infants with vesico-ureteric reflux. AB - OBJECTIVE: To determine the sensitivity of the direct radionuclide cystogram (DRC) in detecting vesico-ureteric reflux compared with the micturating cysto urethrogram (MCU) in the same initial setting, in infants younger than one year. PATIENTS AND METHODS: The results from the dual cystograms of 62 refluxing infants < 1 year old (mean 0.58) were compared. Results from same-day renal scintigraphy with dimercaptosuccinic acid (DMSA) in 60 of the 62 infants were also compared with the reflux grades. RESULTS: Reflux was detected in 105 units, 96 detected on the DRC and 47 on the MCU, representing a sensitivity of 91% and 45%, respectively. The DRC missed half of grade 1, 20% of grade 2 and 6% of grade 3 reflux. Reflux at low bladder filling rates (DRC) represented 40% of all reflux units, and a half (52%) of scarred renal units detected by DMSA scintigraphy. CONCLUSIONS: In young infants the MCU may fail to detect significant reflux and the DRC may fail to detect the lesser grades. The combination of both cystograms in the initial investigation of reflux provides more comprehensive information. PMID- 11122001 TI - The effect of superoxide dismutase on nitric oxide-mediated and electrical field stimulated diabetic rabbit cavernosal smooth muscle relaxation. AB - OBJECTIVE: To investigate the effect of superoxide dismutase (SOD, the enzyme that accelerates the breakdown of the superoxide anion, O2- to H2O) on nitric oxide (NO)-mediated and electrical field stimulated (EFS) relaxation in diabetic rabbit cavernosal smooth muscle. Materials and methods Diabetes was induced with alloxan (65 mg/kg) in six adult New Zealand White rabbits. After 6 months, cavernosal smooth muscle strips from age-matched controls and diabetic animals were mounted in organ baths. After precontraction with phenylephrine (10 micromol/L) in the presence of atropine (1 micromol/L), guanethidine (5 micromol/L) and indomethacin (10 micromol/L), relaxation responses to EFS (1-20 Hz), carbachol (10(-8)-10(-4) mol/L) and sodium nitroprusside (SNP, 10(-9)-10(-4) mol/L) were assessed in the presence and absence of SOD (100 IU/mL). RESULTS: SNP and carbachol-mediated (endothelium-independent and -dependent, respectively) relaxations were impaired in the diabetic cavernosal smooth muscle strips compared with controls (concentration required for 50% inhibition, 1.4 micromol/L for diabetic and 0.75 micromol/L for control with SNP, and 44 micromol/L for diabetic and 0.4 micromol/L for control with carbachol). SOD significantly enhanced both SNP- and carbachol-mediated diabetic cavernosal smooth muscle relaxations (both P < 0.05). EFS-mediated relaxations were also significantly (P < 0.05) impaired in the diabetic cavernosal smooth muscle strips; these relaxations were also significantly (P < 0.05) enhanced by SOD. CONCLUSION: NO- and EFS-mediated cavernosal smooth muscle relaxation is impaired in a rabbit model of diabetes but SOD significantly reversed the impaired relaxation. Therefore, in diabetes, the generation of reactive oxygen species may play an important role in the development of erectile dysfunction. PMID- 11122002 TI - Prophylactic efficacy of a new gentamicin-releasing urethral catheter in short term catheterized rabbits. AB - OBJECTIVE: To describe an indwelling urethral catheter coated with gentamicin sulphate on the inner and outer surface of the catheter, and to evaluate the efficacy and safety of this catheter in preventing catheter-associated infections in rabbits. Materials and methods Sixty rabbits were divided equally into control and experimental groups which were then subdivided equally according to the duration of catheterization (1, 3 and 5 days). Silicone-treated latex catheters were used in the control group and gentamicin-releasing catheters in the experimental group. Urine samples and surface swabs from the catheter were cultured for bacteriological assessment, and the catheter surface examined by scanning electron microscopy to structurally analyse the biofilms. RESULTS: The gentamicin-releasing catheter reduced the incidence of bacteriuria (defined as > or = 100 c.f.u./mL) after both 3 and 5 days of catheterization (eight and 10 rabbits, respectively, for the control catheter, vs two and four rabbits for the gentamicin-releasing catheter, P < 0.05). The surfaces of the gentamicin releasing catheter were colonized less often than those of the control catheter after both 3 and 5 days (eight and 10, respectively, for the control, vs one and four for the gentamicin-releasing catheter, P < 0.05). Scanning electron microscopy showed the formation of bacterial biofilm throughout the 3-day and 5 day control catheters, but deterioration of the bacterial biofilm was visible on the surface of the gentamicin-releasing catheters. CONCLUSION: This new gentamicin-releasing catheter produced an antibacterial barrier which inhibited catheter-associated urinary tract infection with no toxicity for at least 5 days. These in vivo studies suggest that this new catheter may be useful for controlling infection, with systemic and local safety, in patients undergoing short-term indwelling urethral catheterization. PMID- 11122003 TI - Effect of experimental hyperoxaluria on renal calcium oxalate monohydrate binding proteins in the rat. AB - OBJECTIVE: To determine the functional role of calcium oxalate binding proteins in the nucleation, aggregation and retention of calcium oxalate crystals under physiological and hyperoxaluric conditions. Materials and methods Hyperoxaluria was induced in rats using 0.75% of ethylene glycol in drinking water. Calcium oxalate binding proteins were isolated and fractionated by cellulose column chromatography. Three major protein peak fractions were obtained (73 kDa in Tris HCl buffer, 20 kDa in 0.05 mol/L NaCl buffer and 23 kDa in 0.3 mol/L buffer). Oxalate binding and the inhibition of crystal nucleation and aggregation by these fractions were determined. RESULTS: The adsorption of calcium oxalate monohydrate (COM) was ubiquitous in rat tissues and subcellular organelles, but the percentage adsorption varied; maximum absorption occurred in kidneys and pancreas, with microsomes showing maximal adsorption in the kidney. Hyperoxaluric rat tissues showed a greater percentage adsorption. Microsomes were enriched with the 20 kDa protein, while nuclei contained the 23 kDa protein in higher concentrations. COM-binding proteins derived from hyperoxaluric rat kidney had a greater content of 74 kDa and 23 kDa proteins with increased oxalate-binding activities. In the crystal-growth studies, the 74 kDa protein was a promoter, while the other protein fractions inhibited crystallization. In hyperoxaluria, the crystal-growth promoting activity of the 74 kDa protein was further increased, while the inhibition by the 20 and 23 kDa proteins was decreased. The 74 kDa protein derived from control rats formed single COM crystals in a crystal growth system, while the hyperoxaluric rat fraction induced the aggregation of COM crystals. CONCLUSION: COM-binding proteins (the 74 and 23 kDa fractions) were expressed more in hyperoxaluric rats. In hyperoxaluria the 74 kDa protein tended to promote crystal nucleation and aggregation, and the 20 and 23 kDa proteins were less inhibitory, which increases the risk of stone formation. PMID- 11122004 TI - A technique for constructing an umbilicus and a concealed catheterizable stoma. PMID- 11122005 TI - Lymphoepithelioma-like carcinoma of the bladder. PMID- 11122006 TI - Clinical evidence of neuroendocrine differentiation in a patient with prostate cancer and bone marrow micrometastases. PMID- 11122007 TI - A rare complex bladder exstrophy variant. PMID- 11122008 TI - Medicolegal aspects of vesicovaginal fistula. PMID- 11122009 TI - Are men with lower urinary tract symptoms at increased risk of prostate cancer? A systematic review and critique of the available evidence. PMID- 11122010 TI - Mean time to cancer-specific death of apparently clinically localised prostate cancer: policy implications for threshold ages in prostate-specific antigen and ablative therapy. PMID- 11122011 TI - Towards a better classification of erythrokeratodermias. PMID- 11122012 TI - Drug-induced aphthous ulcers. PMID- 11122013 TI - Dermal fibroblasts are one of the therapeutic targets for topical application of 1alpha,25-dihydroxyvitamin D3: the possible involvement of transforming growth factor-beta induction. AB - BACKGROUND: Transforming growth factor (TGF) -beta has been suggested to be an effective inhibitor for abnormal keratinocyte growth in psoriasis. As a majority of the secreted TGF-beta are biologically latent complexes, activation is essential for TGF-beta-mediated cellular responses in vitro and in vivo. Objectives Here we report the response of the TGF-beta regulation system to 1alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3], an active vitamin D3 analogue Patients/methods We studied two types of fibroblasts derived from normal and psoriatic lesional skin, using an enzyme-linked immunosorbent assay and Northern blotting techniques. RESULTS: 1,25(OH)2D3 caused a dose-dependent induction of latent and active TGF-beta1 proteins in both cell cultures. The increases were significant over 72 h, but not within 48 h after stimulation. The time course of TGF-beta1 mRNA expression showed a biphasic response consisting of early ( approximately 1 h) and late phases ( approximately 96 h) of induction. Concomitant increases of TGF-beta2 and -beta3, other mammalian isoforms, were observed in the 1,25(OH)2D3-treated cells, but the kinetics were all different. Co-incubation with metabolic inhibitors, actinomycin D and cycloheximide, revealed that the early induction of TGF-beta1 mRNA by 1,25(OH)2D3 is dependent on de novo RNA synthesis, but not on RNA stabilization or protein synthesis. It seems likely to be a transient and negligible response given the absence of TGF beta1 protein production. The late induction of TGF-beta1 mRNA was partially blocked by adding isoform-specific antibodies to TGF-beta1, -beta2 and -beta3, indicating TGF-beta autoregulation. Despite these marked responses, there were no significant differences in the TGF-beta expression between normal and psoriatic fibroblasts. CONCLUSIONS: These results suggest that antiproliferative and anti inflammatory effects of 1,25(OH)2D3 on psoriatic lesional skin may be mediated, at least in part, by a complex TGF-beta regulation in local dermal fibroblasts. PMID- 11122014 TI - Lack of co-ordinate expression of the alpha1(I) and alpha1(III) procollagen genes in fibroblast clonal cultures. AB - BACKGROUND: Several extracellular matrix genes, most notably alpha1(I) and alpha1(III) procollagen, are reported to be co-ordinately expressed in cultures of dermal fibroblasts. However, it remains unclear whether the expression of these genes is truly co-ordinate or whether it may be the result of averaging the phenotypic expression of different fibroblast subpopulations present within each culture. Objectives To determine by Northern analysis the correlation between alpha1(I) and alpha1(III) procollagen mRNA levels in clonal populations of human dermal fibroblasts. METHODS: As previously described, clonal cultures were derived from parent strains of human dermal fibroblasts by a microscopically controlled dilution technique and by stimulation of single cells with low oxygen tension in the early phases of clonal growth. RESULTS: In agreement with previous reports, we found that baseline steady-state levels of alpha1(I) procollagen mRNA were co-ordinately regulated with the alpha1(III) procollagen mRNA in 26 parent strains (r = 0. 9003; P < 0.0001). However, this close correlation between the expression of these two procollagen chains was absent in a total of 40 unselected clonal strains derived from four of the parent cultures (r = 0.5745; P < 0.0001). Moreover, this intrachain heterogeneity in alpha1(I) and alpha1(III) procollagen mRNA levels in clonal cultures was statistically significant from that measured in parent strains (P = 0.0016). CONCLUSIONS: alpha1(I) and alpha1(III) procollagen mRNA levels in clonal cultures do not show the tight co-ordinate regulation observed in non-clonal cultures, suggesting that these two genes operate under different sets of regulatory controls. This clonal heterogeneity may provide additional flexibility to the process of tissue repair and fibroblast clonal expansion. PMID- 11122015 TI - C2-ceramide induces apoptosis in a human squamous cell carcinoma cell line. AB - BACKGROUND: Previous studies have demonstrated that synthetic cell-permeable analogues of ceramide promote differentiation and inhibit proliferation of keratinocytes, and that the vitamin D3 inducible sphingomyelin cycle generates ceramide in keratinocytes. Although it has been suggested that exogenous ceramide induces apoptosis of keratinocytes, which is similar to their effect on other cell types, such as leukaemia cells, only a few studies have reported ceramide induced apoptosis of keratinocytes. OBJECTIVE: To determine whether ceramide induces apoptosis of keratinocytes, we used the synthetic ceramide analogue, C2 ceramide (N-acetylsphingosine) and a human squamous cell carcinoma cell line, HSC I. METHODS: We treated HSC-I cells with C2-ceramide, followed by a viability assay, morphological observations, nick end-labelling (TUNEL), DNA electrophoresis, and electron microscopy. RESULTS: In the viability assay, C2 ceramide was toxic to HSC-I cells in a dose-dependent manner. Manifestations of apoptotic morphology occurred in the ceramide-treated cells, whereas these morphological changes did not occur in cells treated with dihydroceramide (N acetylsphinganine). TUNEL revealed that many of the ceramide-treated cells showed positive reactivity. DNA electrophoresis demonstrated that C2-ceramide caused internucleosomal fragmentation in a dose- and time-dependent manner. Electron microscopy revealed that the ceramide-treated cells manifested morphological characteristics typical of apoptosis. CONCLUSIONS: The present results demonstrate that C2-ceramide induces apoptosis of transformed human keratinocytes, whereas C2-dihydroceramide does not have such an effect. The fact that ceramide induces apoptosis of keratinocyctes raises the possibility that intracellular ceramide, which is increased with differentiation of the epidermis, might be involved in terminal differentiation, a specialized form of apoptosis of keratinocytes. PMID- 11122016 TI - Acitretin is converted to etretinate only during concomitant alcohol intake. AB - BACKGROUND: Acitretin has replaced etretinate in the treatment of various disorders of keratinization due to a considerably shorter terminal half-life. Possible esterification of acitretin to etretinate in the presence of ethanol has been reported. OBJECTIVES: To determine the plasma concentrations of etretinate as a metabolite in patients with various disorders of keratinization after multiple acitretin dosing, and to assess the influence of alcohol consumption using a questionnaire. In addition, to study the influence of alcohol consumption on the risk of metabolic formation of etretinate. PATIENTS/METHODS: Eighty-six acitretin (Neotigason(R), Roche)-treated outpatients from three centres provided pre-dose (trough) samples for determining plasma concentrations of acitretin and its metabolites 13-cis-acitretin and etretinate. Patients received acitretin doses of between 0.1 and 1.3 mg kg-1 daily. The concentrations of etretinate, acitretin and 13-cis-acitretin were determined by reverse-phase high-performance liquid chromatography. RESULTS: Of the 86 patients, 30 had detectable plasma etretinate levels. No etretinate was found in 20 patients who reported that they never drank alcohol, while etretinate was found in all 16 patients with an average weekly alcohol consumption of > 200 g ethanol, corresponding to about 15 U (1 U equals half a pint of standard beer or a wine glass of non-fortified wine). Etretinate was detected in 14 of 50 patients with a moderate weekly alcohol intake of up to 200 g ethanol. A trend linking higher alcohol intake with both higher risk of etretinate formation and higher etretinate levels was observed. The study also revealed that the ethylesterification only relates to acitretin (13-trans-) and not to the main metabolite 13-cis-acitretin, although the latter compound showed higher plasma trough concentration levels at steady state. CONCLUSIONS: Owing to the teratogenic potential and possible side-effects of oral retinoids, fertile women especially should be informed about the importance of strict alcohol abstinence during treatment and for at least 2 months after stopping therapy. In case of non-compliance with alcohol abstinence a post-therapy contraceptive period of 2-3 years should be recommended. PMID- 11122017 TI - Treatment of psoriasis with oral liarozole: a dose-ranging study. AB - BACKGROUND: Liarozole is an inhibitor of the metabolism of all-trans-retinoic acid. Systemic administration increases tissue levels of this endogenous retinoid and has been reported to improve psoriasis in an open, uncontrolled study. OBJECTIVES: A multicentre, double-blind, placebo-controlled, dose-ranging study was therefore undertaken to determine the lowest effective oral dose of liarozole in the treatment of psoriasis vulgaris. PATIENTS/METHODS: Adult male and postmenopausal female patients requiring systemic treatment for psoriasis were randomized to receive placebo or liarozole at total daily doses of 50 mg, 75 mg or 150 mg for 12 weeks. The daily doses were each divided into two equal (morning and evening) doses. Response was assessed using an eight-point global scale to assess improvement and by monitoring the Psoriasis Area and Severity Index (PASI). The primary end-point was the proportion of subjects in each treatment group demonstrating 'marked improvement' or better as assessed on the eight-point scale. The tolerability of the treatment was assessed by recording mucocutaneous effects of retinoids and all adverse events. Biochemical and haematological monitoring were also performed. RESULTS: One hundred and thirty-nine subjects were randomized (118 male and 21 female) and 116 completed the study. A marked improvement or better response was observed in 6% of subjects on placebo, 18% on liarozole 50 mg, 11% on 75 mg and 38% on 150 mg. Only in the 150-mg group was the response rate significantly different to placebo (P < 0.001). Over the treatment period the mean PASI changed from 15.9 to 15.4 on placebo, from 17.4 to 13.8 on liarozole 50 mg, from 17.5 to 14.5 on 75 mg and from 15.8 to 8.8 on 150 mg. Again, only in the group receiving 150 mg was the response significantly better than placebo (P < 0.001). Liarozole was generally well tolerated. Mucocutaneous retinoid effects were generally infrequent and mild. Five subjects were withdrawn from treatment as a result of adverse events that may have been treatment related. These events were abnormalities of liver enzymes in two cases, an episode of erythema multiforme (in a patient receiving placebo), an allergic reaction in one and a rash accompanied by deterioration of the psoriasis in another. There was mild elevation of triglycerides in the groups receiving liarozole 75 mg and 150 mg daily. In males, the serum luteinizing hormone and testosterone levels rose significantly in all the active treatment groups. CONCLUSIONS: The data confirm that liarozole is an effective treatment for psoriasis and indicate that the lowest effective dose is 75 mg twice daily. The drug seems generally to be well tolerated. PMID- 11122018 TI - HLA-C and guttate psoriasis. AB - BACKGROUND: Psoriasis is a heterogeneous disease in its clinical expression. Both genetic and environmental factors are thought to contribute to the pathogenesis of the inflammatory and hyperproliferative components of the typical skin lesions. Predisposing genetic influences include associations with human leucocyte antigens (HLA) of which that with HLA-Cw6 is the strongest. Guttate psoriasis is a specific clinical manifestation of psoriasis frequently associated with group A beta-haemolytic streptococcal throat infection. OBJECTIVES: We set out to determine whether further clinical subdivision of psoriasis is associated with tighter correlation with HLA-C alleles. PATIENTS/METHODS: We determined the HLA-C locus genotype of 29 caucasian patients with guttate psoriasis presenting consecutively with guttate psoriasis associated with a history of a sore throat and/or an antistreptolysin O titre > 200 IU mL-1. Polymerase chain reaction typing using sequence-specific primers was used to detect all known HLA-C alleles. These data were compared with a control population of 604 random caucasian cadaver donors. RESULTS: All patients (100%) with guttate psoriasis carried the Cw*0602 allele compared with 20% of the control population (odds ratio = infinity; 95% confidence limits 25.00-infinity; Pcorrected < 0.0000002). CONCLUSIONS: This result is consistent with HLA-Cw*0602 playing a part directly in the pathogenesis of guttate psoriasis. PMID- 11122019 TI - Analysis of dendritic cell populations using a revised histological staging of morphoea. AB - BACKGROUND: Recent studies have suggested that dermal dendritic cells (DDCs) may play a part in maintaining the structure of the dermis and in dermal immune modulation. Alteration in the population of DDCs has been noted in localized and systemic scleroderma, particularly a decline in the number of CD34+ DDCs. Objectives To define the alteration of the DDC populations with respect to the histological stage of morphoea. METHODS: We examined 33 biopsies of morphoea, categorized into four histological stages, and examined the DDC population (CD34+ DDCs and factor XIIIa+ DDCs), the lymphocytic infiltrate, and tenascin (extracellular matrix glycoprotein) and transforming growth factor (TGF)-beta1 expression in each biopsy. RESULTS: As the dermis became less inflammatory and more sclerotic, there was a significant decline in the number of CD34+ DDCs and an increase in the number of factor XIIIa+ DDCs. The pan-T-cell infiltrate (UCHL 1/CD45RO) and tenascin deposition exhibited a similar pattern, with elevated expression in inflammatory stages and a decrease in expression as the dermis became sclerotic. TGF-beta1 was significantly elevated in three of the four histological stages of morphoea, in both the inflammatory and sclerotic stages. The proposed four-stage histological analysis of morphoea biopsies was a useful basis for studying dendritic cells and mediators in cutaneous sclerosis. CONCLUSIONS: Our study indicates that there is a reciprocal relationship between CD34+ DDCs and factor XIIIa+ DDCs in morphoea that correlates with the relative degrees of inflammation and sclerosis. PMID- 11122020 TI - Phenotyping of epidermal dendritic cells allows the differentiation between extrinsic and intrinsic forms of atopic dermatitis. AB - Atopic dermatitis (AD) is a clinically characteristic, chronic inflammatory skin disease of unknown origin. IgE-mediated uptake and antigen focusing of environmental allergens by dendritic cells (DCs) is assumed to be a central immunopathogenetic event. A so-called intrinsic type of AD (IAD) has been delineated from the more common extrinsic AD (EAD) by normal serum IgE levels, negative RAST tests and negative immediate-type skin reactions towards environmental allergens. The recently characterized human autoantigen Hom S 1 has been proposed to play a part in the pathogenesis of IAD. OBJECTIVES: To compare clinical and laboratory data between patients with IAD and EAD, and to investigate potential differences in the inflammatory micromilieu of the epidermal compartment in IAD and EAD lesions. METHODS: Epidermal DC phenotyping, a recently validated technique based on the three-colour flow cytometric analysis of Langerhans cells and the so-called inflammatory dendritic epidermal cells from epidermal single-cell suspensions, was performed on samples from 69 patients with AD (seven with IAD and 62 with EAD) and 94 controls. RESULTS: Patients with EAD tended to have an earlier onset of disease but similar disease duration and family history of atopic diseases. Quantitative analysis of CD36 expression on DCs as a marker of inflammation, as well as the percentage of inflammatory dendritic epidermal cells in the CD1a+ epidermal DC pool, indicated a comparable disease activity in IAD and EAD. EAD was characterized by a significantly higher FcepsilonRI expression on the CD1a+ epidermal DCs than IAD. Using the FcepsilonRI/FcgammaRII expression ratio as a disease marker for AD, values for IAD fell below the diagnostic cut-off level of 1.5 for this ratio. CONCLUSIONS: While IAD is clinically similar to EAD, the inflammatory microenvironment in this condition seems different from classical EAD and can be distinguished by phenotyping of epidermal DCs. PMID- 11122021 TI - CD8-positive juvenile onset mycosis fungoides: an immunohistochemical and genotypic analysis of six cases. AB - BACKGROUND: Childhood cases of cytotoxic T-cell lymphoma have not been well described. OBJECTIVES: We have undertaken an immunohistochemical and genotypic analysis of patients presenting with juvenile onset mycosis fungoides (MF). PATIENTS/METHODS: Of 10 patients presenting over a 3-year period, six exhibited a CD8-positive phenotype. These six patients were also CD2, CD3 and TIA1 positive, but CD56 negative. Apart from the cytotoxic phenotype, these patients had clinicopathological features that were indistinguishable from ordinary cases of MF, with slowly evolving patches and plaques. Three patients were staged as 1A and three as 1B, with no evidence of nodal or systemic disease. RESULTS: Patients responded well to conventional therapy, with no evidence of disease progression after 3 years follow-up. Epidermotropism was a prominent feature in four of the six cytotoxic cases. In two patients with an equivocal histology the diagnosis was confirmed by the finding of a clonal population, using polymerase chain reaction/single strand conformational polymorphism analysis of the T-cell receptor gamma gene in lesional skin. The same technique revealed that all blood samples analysed were polyclonal. CONCLUSIONS: These data show that cytotoxic T cell lymphoma can pursue an indolent course and that cases of CD8-positive MF may be over-represented in childhood. PMID- 11122022 TI - Decreased expression of Fas (APO-1/CD95) on peripheral blood CD4+ T lymphocytes in cutaneous T-cell lymphomas. AB - BACKGROUND: The usually protracted and indolent course of cutaneous T-cell lymphoma (CTCL) is consistent with an accumulation of lymphocytes rather than being a true proliferative disorder, perhaps as the result of defective lymphocyte apoptosis. Fas (CD95) is the main signalling membrane molecule involved in postactivation T-lymphocyte apoptosis. OBJECTIVES: To evaluate expression of Fas on circulating CD4+ lymphocytes in patients with CTCL. METHODS: Fas expression on peripheral blood CD4+ T cells in 16 patients with mycosis fungoides (patch and infiltrated plaque stages) and in four patients with Sezary syndrome was compared with that in 25 matched patients with lymphocyte-mediated cutaneous benign inflammatory disorders and in 15 subjects without inflammatory cutaneous diseases. RESULTS: Fas expression on peripheral CD4+ lymphocytes was significantly lower in patients with CTCL compared with subjects with benign inflammatory cutaneous disorders and with healthy donors. CONCLUSIONS: This pattern supports the hypothesis that a defect in T-cell apoptosis may play a part in the pathophysiology of CTCL, perhaps through abnormalities of the Fas/Fas ligand system. Alternatively, this decrease could be the result of the presence of the soluble Fas ligand molecule in the sera of patients with CTCL. PMID- 11122024 TI - A clinical study of 23 cases of female anogenital carcinoma. AB - BACKGROUND: Vulval carcinoma is a relatively rare disorder that may have various aetiologies. Objectives To document the features and outcome in a series of patients with this disorder. METHODS: Retrospective analysis of patients presenting to a vulval clinic over a 5-year period. RESULTS: Twenty-one women presented with a squamous cell carcinoma (SCC) and two with a verrucous carcinoma (VC). The age range was 43-83 years. Twenty-one had well-established (1-30 years) vulval symptoms prior to developing their tumour. Specific tumour-related symptoms ranged from 3 weeks to 11 months. Eight had had a prior diagnosis of lichen sclerosus (LS) or lichen planus (LP), only two of whom were on regular treatment and follow-up. At presentation, 12 patients had clinical signs of LS, three of LP, and five had some changes of both LS and LP. Two patients had multifocal vulval intraepithelial neoplasia (VIN3). Only one had no evidence of any background vulval skin disease. The commonest histological changes noted in the epithelium either adjacent to or distant from the SCC were those of atrophic LS (n = 8), LS with squamous cell hyperplasia (n = 3), LS with hyperplastic foci and lichenoid infiltrate (n = 4), and LS with differentiated VIN3 (n = 1). Four cases demonstrated the changes of LP, and three showed VIN3. All patients were treated surgically and, in those who had lymphadenectomy, four had positive nodes. There have been two deaths due to metastatic disease, and one further patient has developed a second primary SCC at a different site. CONCLUSIONS: An underlying skin disorder prior to the development of their carcinoma was found in 22 of 23 patients with vulval SCC and is therefore an important risk factor. PMID- 11122023 TI - CD1a+, CD3+, CD4+, CD8+, CD68+ and cutaneous lymphocyte-associated antigen positive cells in Bowen's disease. AB - BACKGROUND: Bowen's disease (BD) is a squamous cell carcinoma in situ that rarely invades into the underlying dermis. However, little is known about its immunohistology. Objectives To evaluate the relationship between the cytological properties of the tumour cells in BD and the host immune response. METHODS: We examined the expression of p53, proliferating cell nuclear antigen (PCNA) and Ki67 antigen, and the number of mitotic cells, together with the number of intratumoral and dermal infiltrating CD1a+, CD3+, CD4+, CD8+, CD68+ and cutaneous lymphocyte-associated antigen (CLA)+ cells in 18 cases of genital BD. RESULTS: When compared with normal genital skin (n = 10), there was a significantly higher number of mitotic cells as well as higher expression of p53+, PCNA+ and Ki67+ cells in BD. There was significant mutual correlation between CD3+, CD4+ and CD68+ cells in the tumoral epidermis. The number of CD1a+ Langerhans cells significantly decreased in BD epidermis; however, dermal CD1a+ cells were increased. Interestingly, numbers of dermal CD1a+ cells significantly correlated with those of intratumoral CD3+, CD4+ and CD68+ cells. In situ hybridization for human papillomavirus (HPV) demonstrated that HPV-infected BD had significantly less infiltration of intratumoral CD3+ cells and CLA+ cells. CONCLUSIONS: The present data suggest that dermal CD1a+ cells may participate in the immune surveillance and that HPV infection may interfere with the intratumoral infiltration of CLA+ cells in BD. PMID- 11122025 TI - Changing trends in non-melanoma skin cancer in South Wales, 1988-98. AB - BACKGROUND: In 1988 our department carried out a population-based epidemiological study of non-melanoma skin cancer (NMSC) incidence, over a 6-month period, in West Glamorgan, South Wales. Objectives To reassess the incidence of NMSC in this defined population over a similar 6-month period 10 years after the initial study. METHODS: All cases of basal cell carcinoma and squamous cell carcinoma diagnosed in West Glamorgan are recorded by the local skin cancer registry. All cases for the relevant 6-month period were analysed. RESULTS: Using these figures, we have identified a significant rise in the crude incidence of NMSC from 173.5 10 years ago to 265.4 per 100,000 population per annum. We also applied the world standard population for age to our crude figures, demonstrating a combined male and female world population-corrected rate of 129.9 per 100,000 population. CONCLUSIONS: In our population the crude incidence of NMSC has risen significantly over 10 years. Additionally, the combined male and female world population-corrected rate appears to be the highest published standardized incidence of NMSC to date from any European country. PMID- 11122026 TI - Recurrence of port-wine stains after treatment with the flashlamp-pumped pulsed dye laser. AB - OBJECTIVES: To assess the importance of the patient's age at the start of treatment of a port-wine stain (PWS) with the flashlamp-pumped pulsed dye laser (FPDL). BACKGROUND: FDPL treatment is safe and effective for PWSs, with a low risk of scarring and pigmentary changes. The degree of clearing of the lesion is, however, unpredictable, and the ideal time to start treatment has not yet been agreed. Patients/methods By means of a questionnaire, we investigated the frequency of recurrence in PWS in 147 patients after completion of treatment with the FPDL. RESULTS: In 24 patients (16.3%), partial redarkening of their PWS was observed. The patients who had a recurrence were not different from the group who did not regarding the colour of the PWS, the response to previous treatment or the frequency of side-effects. Children under 10 years of age did not show any PWS recurrence, at least in our group of patients. CONCLUSIONS: The age at the beginning of treatment may have an influence on the recurrence rate. PMID- 11122027 TI - High levels of interleukin-8, soluble CD4 and soluble CD8 in bullous pemphigoid blister fluid. The relationship between local cytokine production and lesional T cell activities. AB - BACKGROUND: Bullous pemphigoid (BP) is an inflammatory subepidermal blistering disease associated with autoantibodies that recognize hemidesmosomal proteins. In addition to autoantibodies, the cell-mediated immune reaction is considered to play an important part in blister formation. Objectives To investigate some T cell activation markers and inflammatory cytokines in the blister fluid and sera of patients with BP. METHODS: We measured soluble CD4 (sCD4) and soluble CD8 (sCD8), which have been, respectively, associated with CD4 and CD8 T-cell activation. Enzyme-linked immunosorbent assays were also used to quantify the production of the leucocyte chemoattractant interleukin (IL) -8 and of the cytokines IL-1alpha, IL-1beta, IL-6, IL-10 and tumour necrosis factor-alpha in the blister fluid and sera of 11 patients with BP. RESULTS: The mean +/- SD level of sCD4 in patients' blisters (42.4 +/- 25.0 units mL-1) was significantly elevated (P < 0.005) compared with that in their sera (11.2 +/- 8.9) and that in the suction blisters of 10 healthy people (11.4 +/- 5.4; P < 0.005). Mean +/- SD IL-8 concentrations in BP blisters (4683.6 +/- 3878.1 pg mL-1) were much higher than those in their sera (17.1 +/- 18.9; P < 0.001), and were very significantly elevated (P < 0.005) in comparison with those in suction blisters of healthy persons (512 +/- 292). sCD4 levels in BP blisters were inversely related to IL-10 levels (P = 0. 03, r2 = 0.85), IL-8 levels were positively related to sCD8 levels (P = 0.01, r2 = 0.54), and IL-1beta levels were positively related to sCD8 concentrations (P < 0.005, r2 = 0.65). CONCLUSIONS: The correlations suggest that there is a delicately orchestrated network of cytokines and cell-mediated immunity operating in BP blisters. PMID- 11122028 TI - A comparison of real-time and store-and-forward teledermatology: a cost-benefit study. AB - BACKGROUND: Increasing use of teledermatology should be based on demonstration of favourable accuracy and cost-benefit analysis for the different methods of use of this technique. Objectives To evaluate the clinical efficacy and cost effectiveness of real-time and store-and-forward teledermatology. METHODS: Patients attended their own health centre and in the company of a general practitioner (GP) were seen by a hospital dermatologist over the videolink (real time). Before the videolink consultation commenced, the GP took instant photographs of the skin lesion and posted them along with a standard referral letter to a different hospital dermatologist (store-and-forward). In total, 96 patients were seen by both real-time and store-and-forward teledermatology. Comparative diagnoses, clinical management plans, clinical outcomes and associated costs were made between the two types of teledermatology consultation. RESULTS: There was agreement between the videolink diagnosis and the still image diagnosis in 51% of cases. The same or similar management plan was recommended at both types of consultation in 44% of cases. Following the store-and-forward consultation the dermatologist recommended that 69% of patients required at least one hospital appointment compared with 45% of those patients seen in real-time. The net societal cost of the initial real-time consultation was pound132.10 per patient compared with £26.90 per patient for the initial store-and-forward consultation. CONCLUSIONS: The store-and-forward consultation was cheaper, but less clinically efficient, compared with the real-time consultation. The absence of interaction in a store-and-forward consultation limits the dermatologist's ability to obtain clinically useful information in order to diagnose and manage a patient satisfactorily. PMID- 11122029 TI - The spectrum of skin disorders in abdominal stoma patients. AB - BACKGROUND: Skin integrity is essential for the normal usage of a stoma appliance. However, there is little published on the prevalence, nature or management of stoma-related skin disorders. Objectives To document stoma-related skin disorders in a large cohort of patients. METHODS: We sent a postal questionnaire to all surviving patients who had had abdominal stoma surgery at Hope Hospital, Salford, U.K. in the 10 years from 1 January 1989. Those reporting skin disease were invited to attend a clinic run by a dermatologist and a stoma care specialist nurse. All lesions were categorized and swabs taken for microbiological examination. RESULTS: Of 525 surviving patients, 325 (62%) replied to the questionnaire. Of these, 73% reported a skin problem that had affected normal stoma bag use. Dermatoses included irritant reactions, particularly from leakage of urine or faeces (42%); pre-existing skin diseases, principally psoriasis, seborrhoeic dermatitis and eczema (20%); infections (6%); allergic contact dermatitis (0.7%) and pyoderma gangrenosum (0.6% annual incidence). A further 15% of patients with skin problems had persistent or recurrent dermatitis not explained by allergy, frank infection or faecal irritation. This responded to short-term treatment with topical corticosteroids. Further investigation is under way into its pathogenesis. CONCLUSIONS: Skin disorders are common in stoma patients, and various patterns can be recognized and effectively treated. PMID- 11122030 TI - Role of drug exposure in aphthous ulcers: a case-control study. AB - BACKGROUND: Drug-induced aphthous ulcers have been the subject of several isolated and heterogeneous case reports for the last three decades. OBJECTIVES: To perform a case-control study to evaluate the risks linked to drug exposure in aphthous ulcers. METHODS: Eighty patients with typical clinical patterns of aphthous ulcers and 152 control patients who had had consultations for skin tumours were studied. A standardized questionnaire, concerning clinical features, life-style and medications taken during the last month, was completed for each patient. RESULTS: Case patients had a much higher intake of medications than control patients, respectively, 5.1 and 2. 8 medications per patient (P < 0.0001). Multivariate paired analysis showed an association between aphthous ulcers and two classes of drugs: non-steroidal anti-inflammatory drugs (P < 0.001) and beta-blockers (P = 0.002). Smoking could have a protective effect against aphthous ulcers (P < 0.001). CONCLUSIONS: Previous case reports and the results of this study suggest a real link between beta-blockers and aphthous ulcers. Our study did not confirm a role of other drugs but a few interesting case reports with positive reintroduction have to be considered. These results could be beneficial for patients, as healing may occur when the incriminated drug is discontinued. PMID- 11122031 TI - Anosacral cutaneous amyloidosis: a study of 10 Chinese cases. AB - BACKGROUND: Primary cutaneous amyloidoses are rare in Western countries, but are relatively common in Taiwan. Anosacral cutaneous amyloidosis is a rare type of primary cutaneous amyloidoses, first reported in Japanese patients. PATIENTS/METHODS: In the present study, we investigated the age of onset, sites of involvement, associated systemic diseases, and histopathological findings in 10 cases of anosacral cutaneous amyloidosis seen during the past 27 years. RESULTS: In previous reports the aetiology of anosacral cutaneous amyloidosis was thought to be a senile change, but half of our patients developed the disease before the age of 60 years. Based on our histopathological findings, apoptosis may be the initial event causing amyloid deposition, although the precise mechanism causing apoptosis needs further investigation. Three patients were found to have diabetes mellitus, but any relationship to anosacral cutaneous amyloidosis is unclear. CONCLUSIONS: No cases of this cutaneous disorder have been reported in the Western literature; there seems to be a racial difference accounting for the disease, although the precise factor is not clarified yet. The disease could easily be misdiagnosed as lichen simplex chronicus, postinflammatory hyperpigmentation or tinea cruris; therefore, a thorough history, a careful physical examination and a skin biopsy is needed to establish a firm diagnosis. PMID- 11122032 TI - Routine double treatments of superficial basal cell carcinomas using aminolaevulinic acid-based photodynamic therapy. AB - BACKGROUND: Superficial basal cell carcinomas of the skin (sBCC) often respond poorly to single-treatment aminolaevulinic acid-based photodynamic therapy (ALA PDT), with a number of reports indicating a relapse rate of 50% or more. OBJECTIVES: To determine whether a second treatment at seven days can improve the response. METHODS: Twenty-six lesions were treated twice with ALA-PDT, with an interval of 7 days between the two treatment sessions. RESULTS: We observed a complete response rate of 100% 1 month after treatment. Only one lesion relapsed (16 months post-PDT), a relapse rate of 4% (median follow up 27 months; range 15 45 months). Cosmetic results were excellent. CONCLUSIONS: We consider routine double treatments with ALA-PDT to be an effective approach to the management of sBCC, particularly those located in anatomically difficult, or cosmetically sensitive, sites. PMID- 11122034 TI - Frequency analysis of autoreactive T-helper 1 and 2 cells in bullous pemphigoid and pemphigus vulgaris by enzyme-linked immunospot assay. AB - BACKGROUND: Bullous pemphigoid (BP) and pemphigus vulgaris (PV) are autoimmune bullous skin diseases mediated by autoantibodies against adhesion molecules of the skin. Previous studies have identified autoreactive T cells in patients with BP and PV, which may be critical in providing B-cell help for autoantibody production. OBJECTIVES: To evaluate the frequency of autoreactive T-helper (Th) 1 and Th2 cells in patients with BP (n = 7) or PV (n = 1) and in healthy controls (n = 11). METHODS: In an enzyme-linked immunospot (ELISPOT) assay, microtitre plates were coated with antihuman interleukin (IL)-5 IgG or antihuman interferon (IFN)-gamma IgG prior to culturing human peripheral blood lymphocytes (PBL) with BP180 or desmoglein (Dsg) 3 proteins for 7 days. Cytokine-producing autoreactive T cells were visualized as spot-forming units. RESULTS: One BP patient with extensive blisters had 5.1 +/- 1.5 (mean +/- SD) BP180-reactive Th1 cells and 2.9 +/- 1.5 Th2 cells per 105 PBL. In contrast, PBL from six BP patients in remission or on immunosuppressive therapy did not form IFN-gamma- or IL-5-producing spots per Tau in a regulatory region of the COLIA1 gene encoding for the major protein of bone (type 1 collagen), and (ii) a one-base deletion in intron 4 (713-8del C) of transforming growth factor beta 1 (TGF-beta1) gene. We have found a remarkable incidence (90%) of osteopenia and osteoporosis among regularly transfused patients. Bone mass was lower in men than in women (P = 0.0023), with a more prevalent osteopenia/osteoporosis of the spine in men than in women (P = 0. 001). The sample was stratified on the basis of BMD expressed as Z-score, i.e. normal, osteopenic and osteoporotic patients, and genotype frequencies of each group were evaluated. TGF-beta1 polymorphism failed to demonstrate a statistical difference in BMD groups. However, subjects with heterozygous or homozygous polymorphism of the COLIA1 gene showed a lower BMD than subjects without the sequence variation (P = 0.012). The differences among genotypes were still present when the BMD was analysed as adjusted Z-score and when men and women were analysed separately (P = 0.022 and 0.004 respectively), with men more severely affected. Analysis of COLIA1 polymorphism could help to identify those thalassaemic patients at risk of osteoporosis and fractures. PMID- 11122086 TI - Venous thromboembolism and hypercoagulability in splenectomized patients with thalassaemia intermedia. AB - Thromboembolic phenomena have been described in patients with thalassaemia intermedia and major, although there are relatively few epidemiological data on the overall frequency of these complications. To obtain more insight into the risk and mechanism of venous thromboembolism in thalassaemia, the aims of this study were: (i) to establish retrospectively the prevalence of thromboembolic events in a large group of adults with thalassaemia intermedia and major during a follow up period of 10 years; (ii) to measure in subgroups of these patients sensitive markers of activation of coagulation and fibrinolysis enzymes; and (iii) to look for possible procoagulant mechanisms. A high prevalence of thromboembolic events was found, particularly in splenectomized patients with thalassaemia intermedia (29%). These patients had high plasma levels of markers of coagulation and fibrinolysis activation. Furthermore, thalassaemic red cells and erythroid precursors from splenectomized patients with thalassaemia intermedia had an enhanced capacity to generate thrombin. To evaluate the role of splenectomy per se on procoagulant activity, we evaluated the capacity to form thrombin in healthy individuals who had been splenectomized for trauma. They produced the same amount of thrombin as non-splenectomized controls. In conclusion, the results of this study show the existence of a hypercoagulable state in splenectomized patients with thalassaemia intermedia and that their red and erythroid cells are capable of acting as activated platelets in thrombin generation. PMID- 11122087 TI - Platelet and leucocyte activation in childhood sickle cell disease: association with nocturnal hypoxaemia. AB - We hypothesized that vaso-occlusive events in childhood sickle cell disease (SCD) may relate to inflammatory cell activation as well as interactions between sickle erythrocytes and vascular endothelium. Peripheral blood was examined from 24 children with SCD, of whom 12 had neurological sequelae and seven had frequent painful crises, and 10 control subjects. Platelet (CD62P and CD40L expression) and granulocyte (CD11b expression) activation and levels of platelet-erythrocyte and platelet-granulocyte complexes were determined by flow cytometry. Platelets (P = 0.019), neutrophils (P = 0.02) and monocytes (P = 0.001) were more activated and there were increased platelet-erythrocyte complexes (P = 0.026) in SCD patients compared with controls. Platelet-granulocyte complexes were not raised. There were no differences between the different groups of SCD. As hypoxia activates monocytes, platelets and endothelial cells and causes sickling of SCD erythrocytes, we also investigated 20 SCD patients with overnight pulse oximetry. Minimum overnight saturation correlated with the level of platelet-erythrocyte complexes (Spearman's rho -0.668, P < 0.02), neutrophil CD11b (Spearman's rho 0.466, P = 0.038) and monocyte CD11b (Spearman's rho -0.652, P = 0. 002). These findings provide important clues about the mechanism by which SCD patients may become predisposed to vaso-occlusive events. PMID- 11122088 TI - Elevated serum and bronchoalveolar lavage fluid levels of interleukin 8 and granulocyte colony-stimulating factor associated with the acute chest syndrome in patients with sickle cell disease. AB - The role of cytokines in the development of acute chest syndrome (ACS) in patients with sickle cell disease (SCD) was studied. Serum interleukin 8 (IL-8) levels were elevated in 14 episodes and undetectable in six out of 20 episodes of ACS in 19 patients with SCD. In contrast, IL-8 levels were undetectable in the sera of 29 control patients with SCD studied during routine clinic visits or hospitalization for vaso-occlusive crises. The differences in mean IL-8 levels and the proportion of patients with detectable levels between the two groups were highly significant (P < 0.0001 and 0.04 respectively). The mean IL-8 level in bronchial fluid samples from children with ACS was also significantly higher than that in sickle cell patients undergoing elective surgery (5500 +/- 1400 pg/ml vs. 1900 +/- 470 pg/ml, P = 0.03). Granulocyte colony-stimulating factor (G-CSF) (2000 +/- 1700 pg/ml) was present in five out of six samples of bronchial fluid, but not serum, from children with ACS. All but one of the patients with ACS studied were negative for the Duffy red cell antigen, which is a receptor that binds and inactivates IL-8 and other chemokines. These findings suggest that IL-8 and G-CSF may play a role in the development of the ACS and the complications associated with it. PMID- 11122089 TI - Primitive haematopoietic progenitors in the blood of patients with sickle cell disease appear to be endogenously mobilized. AB - To investigate whether haematopoietic stem cells in patients with sickle cell (SS) disease might be altered, we examined the number and cycling status of 5 week long-term culture-initiating cells (LTC-ICs) and in vitro multilineage colony-forming cells (CFCs) present in the blood of a large and clinically diverse group of SS patients. The concentrations of both of these cell types per ml of blood varied over a wide range in individual patients, but, on average, were significantly elevated above normal values ( approximately sevenfold and 15 fold respectively) and to an even greater extent than the lineage-restricted CFCs in the same samples. Wide variations in the concentration of circulating progenitors, particularly the LTC-ICs, were also seen over time (in concert with changes in the white blood cell count) in SS patients. [3H]-Thymidine suicide assays showed most of the CFCs and LTC-ICs in SS blood to be quiescent like their counterparts in normal blood. However, by comparison with historical data, the SS progenitors could be recruited into the cycle more quickly (i.e. within 2 vs. 3 d), thus showing the same kinetics of activation exhibited by 'mobilized' progenitors from patients given chemotherapy and exogenous growth factors. Taken together, these findings implicate previously documented increases in endogenous Steel factor, interleukin 3 and granulocyte-macrophage colony-stimulating factor levels in SS patients in the establishment of a chronically mobilized progenitor phenotype. PMID- 11122090 TI - Arginine therapy: a novel strategy to induce nitric oxide production in sickle cell disease. AB - To determine the effects of L-arginine (L-Arg) supplementation on nitric oxide metabolite (NOx) production, oral L-Arg was given to normal controls, sickle cell disease (SCD) patients at steady state and SCD patients hospitalized with a vaso occlusive crisis (VOC). L-Arg (0.1 g/kg) increased NOx formation by 18.8 +/- 68% in normal controls, whereas steady-state SCD patients demonstrated a paradoxical decrease in NOx of -16.7 +/- 4% (P = 0.004). In contrast, patients with VOC demonstrated a dramatic increase in NOx production by +77.7 +/- 103%, a response that was dose dependent. L-Arg appears to be the rate-limiting step in NOx production during VOC. Oral arginine may therefore benefit SCD patients by inducing an increase in NO production during VOC. PMID- 11122092 TI - The relationship between idiopathic thrombocytopenic purpura and pernicious anaemia. AB - Some reports on the simultaneous presence of chronic idiopathic thrombocytopenic purpura (ITP) and pernicious anaemia (PA) may be found in the literature. However, little is known about the coexistence of these autoimmune disorders. For this reason, we studied the prevalence of PA in a series of patients with a diagnosis of chronic ITP by means of the analysis of the concentration of the most sensitive marker of type A atrophic (autoimmune) gastritis, serum pepsinogen I. Serum pepsinogen I was low in 20.3% of the 133 patients studied. Gastrin was elevated in 15. 2% of patients, but the coexistence of both abnormalities was rather low (7.6% of patients). However, the progressive decrease in serum cobalamin as biochemical abnormalities related with atrophic gastritis appeared was noticeable. The time to progression to frank PA from type A atrophic gastritis may span some years. PMID- 11122091 TI - Characterization of osteoclast precursors in human blood. AB - Osteoclast precursors (OCPs) circulate in the mononuclear fraction of peripheral blood (PB), but their abundance and surface characteristics are unknown. Previous studies suggest that the receptor activator for NF-kappaB (RANK) on cytokine treated OCPs in mouse bone marrow interacts with osteoprotegerin ligand (OPGL/TRANCE/RANKL/ODF) to initiate osteoclast differentiation. Hence, we used a fluorescent form of human OPGL (Hu-OPGL-F) to identify possible RANK-expressing OCPs in untreated peripheral blood mononuclear cells (PBMCs) using fluorescence activated cell sorting analysis. Monocytes [CD14-phycoerythrin (PE) antibody (Ab) positive (+) cells, 10-15% of PBMCs] all (98-100%) co-labelled with Hu-OPGL-F (n > 18). T lymphocytes (CD3-PE Ab+ cells, 66% of PBMCs) did not bind Hu-OPGL-F; however, B cells (CD19-PE Ab+ cells, 9% of PBMCs) were also positive for Hu-OPGL F. All Hu-OPGL-F+ monocytes also co-labelled with CD33, CD61, CD11b, CD38, CD45 and CD54 Abs, but not CD34 or CD56 Abs. Hu-OPGL-F binding was dose dependent and competed with excess Hu-OPGL. When Hu-OPGL-F+, CD14-PE Ab+, CD33-PE Ab+, Hu-OPGL F+/CD14-PE Ab+ or Hu-OPGL-F+/CD33-PE Ab+ cells were cultured with OPGL (20 ng/ml) and colony-stimulating factor (CSF)-1 (25 ng/ml), OC-like cells readily developed. Thus, all freshly isolated monocytes demonstrate displaceable Hu-OPGL F binding, suggesting the presence of RANK on OCPs in PB; also, OCPs within a purified PB monocyte population form osteoclast-like cells in the complete absence of other cell types in OPGL and CSF-1 containing medium. PMID- 11122093 TI - Effective lysis of model thrombi by a t-PA mutant (A473S) that is resistant to alpha2-antiplasmin. AB - This study used two mutants of tissue-type plasminogen activator (t-PA) with resistance to inhibitors of fibrinolysis to define the contribution of plasminogen activator inhibitor (PAI)-1 and alpha2-antiplasmin (alpha2-AP) to the control of fibrin lysis. Wild-type t-PA was compared with KHRR296-299AAAA, which is resistant to PAI-1, and with A473S, which is resistant to alpha2-AP. We examined these forms of t-PA in model systems that are physiologically relevant. Neutralization of alpha2-AP was essential for lysis of plasma clots, irrespective of their platelet content, by either wild-type t-PA or KHRR296-299AAAA. In marked contrast, A473S lysed plasma clots without neutralization of alpha2-AP. Model thrombi, with structures similar to in vivo thrombi, were lysed slowly by wild type t-PA; the rate and extent of lysis were enhanced by the addition of antibodies to alpha2-AP or PAI-1. A473S was more effective than wild-type t-PA without the addition of antibodies by virtue of its resistance to alpha2-AP. This resistance was remarkable, in that no complex formed between A473S t-PA and alpha2-AP, even after extended incubation, when 50% of wild-type t-PA could be converted to complex. Comparison of A473S and KHRR296-299AAAA mutants showed their similar effectiveness in lysis of platelet-rich model thrombi. Thus, PAI-1 and alpha2-AP contribute approximately equally to the inhibition of thrombus lysis. This study underlines the functional significance of alpha2-AP as a direct inhibitor of t-PA and further explains the basis of the accepted role of alpha2 AP as a regulator of fibrin persistence and thrombus resistance to lysis. PMID- 11122094 TI - Myeloid leukaemic cells can lyse fibrin directly. AB - Purified preparations of circulating leukaemic blast cells from patients with acute myeloid (M1-7) or acute lymphoblastic leukaemia, and the myeloid or lymphoid cells from patients with chronic myeloid or lymphocytic forms of leukaemia, were incorporated into clots prepared from fibrinogen and plasminogen. Patterns of lysis were followed and measured by light transmission in a microtitre plate reader. Mature polymorphonuclear and mononuclear cell fractions from normal individuals were studied concurrently for comparison. Blast cells from the myeloid forms of acute leukaemia (M2-4) and 'myeloid' cell fractions from patients with chronic myeloid leukaemia were capable of lysing plasminogen containing clots; this activity was neutralized by addition of immunoglobulin against urokinase plasminogen activator (u-PA), but not by anti-tissue plasmogen activator (t-PA). Mature polymorphonuclear and mononuclear cells from normal individuals lacked lytic activity, as did the leukaemic cells from patients with acute lymphoblastic or chronic lymphocytic leukaemia. Lysed blast cells showed the presence of free plasminogen activator on sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) with overlay zymography, also neutralized by anti-u-PA, whereas normal polymorphonuclear and mononuclear cells did not. These observations suggest that mechanisms underlying some forms of severe bleeding in acute myeloid leukaemias have a critical fibrinolytic component generated by the blast cells themselves. PMID- 11122095 TI - Anticoagulation management in primary care: a trial-based economic evaluation. AB - The demand for anticoagulation management is increasing because of a widening of the indications of treatment. A primary care clinic using near-patient testing and computer decision support software is one model of care to meet this increased demand. The study aimed to determine the cost and cost-effectiveness of primary care-based anticoagulation management in comparison with 'traditional' hospital care-based provision by means of a cost-effectiveness analysis using data from a Birmingham-based multicentre randomized controlled trial. The costs per patient per year in primary care were pound170 [95% confidence interval (CI) pound149-190] vs. pound69 (95% CI pound57-81). Sensitivity analysis demonstrated that the cost in primary care could be reduced to under pound100 per patient per year under plausible changes in the variables. Primary care provides similar levels of control to secondary care for patients on anticoagulation therapy. There is an increased cost of managing patients in primary care and at no point did primary care become a lower cost option than secondary care. Local decision makers need to assess the increased cost of primary care anticoagulation management in terms of the potential reductions in high-cost serious adverse events. PMID- 11122096 TI - Abdominal venous thrombosis in neonates and infants: role of prothrombotic risk factors - a multicentre case-control study. For the Childhood Thrombophilia Study Group. AB - The factor V (FV) G1691A mutation, the prothrombin (PT) G20210A variant, the methylenetetrahydrofolate reductase (MTHFR) T677T genotype, together with fasting homocysteine (HCY) concentration, lipoprotein (Lp)(a), anti-thrombin (AT), protein C (PC), protein S (PS) and anti-cardiolipin antibodies were investigated in 65 consecutively recruited infants (neonate to < 12 months) with renal venous thrombosis (RVT; n = 31), portal vein thrombosis (PVT; n = 24) or hepatic vein thrombosis (HVT n = 10), and 100 age- and sex-matched healthy controls. FV G1691A was found in 14 babies (heterozygous: RVT n = 9, PVT n = 4; homozygous HVT n = 1) and five controls, the MTHFR TT677 genotype together with increased HCY in four infants with thrombosis (RVT n = 2; PVT n = 1; HVT n = 1) compared with one control, and the PT G20210A variant was present in one control only. PC type I deficiency was diagnosed in three patients (RVT n = 2; PVT n = 1) and AT deficiency in two patients (RVT n = 1; PVT n = 1). Three neonates with spontaneous thrombosis showed FV G1691A combined with Lp(a) and the FV G1691A was combined with the PT G20210A genotype in two infants. Additional triggering factors were reported in 27 patients (41.5%). The overall odds ratios (ORs) and 95% confidence intervals (CIs) with respect to the different thrombosis locations were: RVT (OR/CI: 10.9/3.85-31.1; P < 0.0001), PVT (5.47/1.7-17.6; P < 0.0007) and HVT (3.3/0.58-18.7; P = 0.18). The data presented here suggest that genetic prothrombotic risk factors also play an important role in abdominal venous thrombosis during infancy. PMID- 11122097 TI - Single nucleotide polymorphisms of the factor IX gene for linkage analysis in the southern Chinese population. AB - Carrier detection and prenatal testing for haemophilia B in Oriental populations have been hampered by the lack of informative markers within the factor IX (FIX) gene. We detected a T/C nucleotide variation at nucleotide 32770 in the poly-A region of the FIX gene in the mother of a haemophilia B child. Analysis of 139 unrelated alleles revealed a heterozygosity rate of 0.193, thus offering an additional marker for linkage analysis. Together with two other polymorphic sites (5' MseI and 3' HhaI) found in Chinese and Thai populations, these polymorphisms were useful in 66% of the families studied. PMID- 11122098 TI - A new polymorphism in the human factor VIII gene: implications for linkage analysis in haemophilia A and for the evolution of int22h sequences. AB - A new polymorphism in the human factor VIII gene has been localized and characterized. It is a biallelic, single nucleotide polymorphism located in intron 22 of the gene, within the 9.5 kb int22h-1 segment. The allelic forms are G (frequency 0.65) and A (frequency 0.35), giving a predicted rate of heterozygosity of 0.46. The polymorphism occurs within a CG dinucleotide and affects an MspI site (CCGG). Int22h-1 is duplicated twice extragenically at Xq28; both extragenic copies (int22h-2 and -3) are also polymorphic with respect to MspI. Investigation of 156 MspI [-] alleles, comprising 30 intragenic and 126 extragenic sites, indicated that all were due to A alleles and none had arisen by C to T transition within the CG dinucleotide. The intragenic MspI site (designated MspI A) is located 737 bases downstream of a previously described XbaI restriction fragment length polymorphism. Despite their close proximity, the polymorphisms are not in complete linkage disequilibrium; haplotype analysis in 85 factor VIII genes from a Caucasian population predicts an informativity of approximately 60% in linkage studies using both, compared with an informativity of approximately 47% in studies using either on its own. PMID- 11122099 TI - Factor IX gene sequencing by a simple and sensitive 15-hour procedure for haemophilia B diagnosis: identification of two novel mutations. AB - We have developed a simple, sensitive and cost-effective direct DNA sequencing procedure for the molecular diagnosis of haemophilia B. All factor IX gene essential regions were amplified under identical thermocycling parameters allowing mutation identification in less than 15 h from blood sample collection. Identical results in terms of accuracy and speed were obtained when using a single hair as the source of DNA. Using this approach, we have found mutations in 10 out of 10 haemophilia B patients, two of which are novel (P287T and S123C). Very suitable for individual studies, this procedure can be easily scaled up for higher throughput. PMID- 11122100 TI - Q1311X: a novel nonsense mutation of putative ancient origin in the von Willebrand factor gene. AB - Type 3 von Willebrand disease, a recessive autosomally inherited bleeding disorder, refers to complete deficiency of von Willebrand factor (VWF). The novel Q1311X mutation was detected in the homozygous state in four Spanish patients from two apparently unrelated families of gypsy origin. The lack of specific amplification of platelet VWF cDNA from two of the patients indicates reduced levels of mutated gene expression. The similar haplotype linked to mutated alleles suggests a common origin. On the basis of the two instabilities observed and the estimated mutation rate of the microsatellites of intron 40 of the VWF gene, we can estimate that this mutation could have arisen about 2300 years ago. PMID- 11122101 TI - A factor XI deficiency associated with a nonsense mutation (Trp501stop) in the catalytic domain. AB - We identified a novel mutation in an asymptomatic 65-year-old Japanese man with severe factor XI deficiency. Sequence analysis after polymerase chain reaction single-stranded conformation polymorphism (PCR-SSCP) analysis of his factor XI gene revealed a G-->A transition in codon 501 of exon 13, resulting in a substitution of Trp501 (TGG) by a stop codon (TAG) in the catalytic domain. This mutation abolished a FokI restriction site. The PCR product from normal subjects was digested with FokI and yielded two fragments, one of 223 bp and one of 47 bp. The PCR product from the patient gave a single 270-bp fragment, demonstrating possible homozygosity. PMID- 11122102 TI - Prevalence of the factor VR506Q mutation in two Irish control populations: use of a novel nested polymerase chain reaction approach. AB - The prevalence of factor V (FV) Leiden among normal populations has primarily been determined using blood donors. This control group is carefully selected and therefore may not accurately reflect the true prevalence within the population. We assessed the prevalence of FV Leiden within the Irish population using Guthrie card samples randomly selected from all newborns. We compared this result with the prevalence of FV Leiden within blood donors. A novel nested polymerase chain reaction (PCR) method for FV Leiden was developed for analysis of the Guthrie card samples. There was no significant difference between the allele frequency within the Guthrie card samples and blood donors (2.07% vs. 2.35%, P = 0.66) PMID- 11122103 TI - Effect of the angiotensin-converting enzyme gene deletion polymorphism on the risk of venous thromboembolism. AB - Allele frequencies for the insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme (ACE) gene were determined in a large case-control study of 517 unselected patients with venous thromboembolism and 478 blood donors. The D allele frequency was 0. 53 [95% confidence interval (CI) 0.50-0.56] in patients and 0.54 (95% CI 0.50-0.57) in controls, giving an odds ratio for the D allele of 0.97 (95% CI 0.81-1.16). In the same population, the odds ratio for the factor V Leiden mutation (F5G1691A) was 6.9 (95% CI 4. 0-11.9). Therefore, the ACE I/D polymorphism is not a risk factor in a representative group of unselected patients with venous thromboembolism. The possibility that the I/D polymorphism is a risk factor for venous thromboembolism specifically after hip replacement cannot be excluded. PMID- 11122104 TI - Contrasting in vitro effects for the combination of fludarabine, cytosine arabinoside (Ara-C) and granulocyte colony-stimulating factor (FLAG) compared with daunorubicin and Ara-C in P-glycoprotein-positive and P-glycoprotein negative acute myeloblastic leukaemia. AB - It has been suggested that the FLAG remission induction regimen comprising fludarabine (F-ara), cytosine arabinoside (Ara-C) and granulocyte colony stimulating factor (G-CSF) may be capable of overcoming P-glycoprotein (P-gp) related multidrug resistance (MDR) in patients with acute myeloblastic leukaemia (AML). We have investigated the in vitro response of P-gp-positive and -negative AML clones to FLAG and compared this with their response to treatment with Ara-C and daunorubicin (DNR). Twenty-four cryopreserved samples from patients with AML were studied using a flow cytometric technique for the enumeration of viable (7 amino actinomycin D negative) cells. Samples consisted of 12 P-gp-positive and 12 P-gp-negative cases, as measured by the MRK16 antibody. The results were analysed by calculating the comparative drug resistance (CDR), i.e. the percentage cell death caused by Ara-C + DNR subtracted from the percentage cell death, caused by FLAG after 48 h incubation in suspension culture. P-gp-positive clones were shown to have a significantly higher CDR than P-gp-negative clones (P = 0. 001). Furthermore, a significant positive correlation (r2 = 0.40, P < 0.01) was found between P-gp protein expression and CDR. However, P-gp function, measured using cyclosporin modulation of rhodamine 123 (R123) uptake, was not associated with the CDR, demonstrating that there are other properties of P-gp, besides its role in drug efflux, that modulate the responsiveness of AML blasts to chemotherapy. These results are consistent with a potential benefit for FLAG in P-gp-positive AML, but not P-gp-negative AML, compared with standard anthracycline and Ara-C therapy. PMID- 11122105 TI - Induction of MTG8-specific cytotoxic T-cell lines: MTG8 is probably a tumour antigen that is recognized by cytotoxic T cells in AML1-MTG8-fused gene-positive acute myelogenous leukaemia. AB - Several reports have demonstrated the persistent detection of AML1-MTG8 fusion products, representing minimal residual disease (MRD), in patients with t(8;21) acute myelogenous leukaemia (AML) who are in long-term remission. It is probable that immune-mediated mechanisms that are able to suppress the expansion of MRD may result in the continuance of remission. It was previously shown that some t(8;21) AML patients had high anti-MTG8 antibody titres. MTG8 expression in normal adult tissues is limited to the brain or heart in which human leucocyte antigen (HLA) class I cell-surface antigens are either not or are only faintly detectable. We hypothesized that the overexpression of the MTG8 gene in t(8;21) AML cells could act as a possible tumour antigen, which might be able to induce the immune-mediated suppression of the expansion of MRD. We were able to induce HLA-A0201-restricted cytotoxic T-lymphocyte (CTL) lines against an MTG8 peptide (MTG8b amino acids 182-191) using monocyte-derived dendritic cells from a healthy donor. T-cell receptor (TCR)Valpha17, TCRVbeta14 and 15, and TCRJbeta2.1 and 2.3 are predominantly used in these CTL lines. Our data, which suggest that the MTG8 protein could be one of the tumour antigens recognized by CTLs, may be helpful in further investigations of TCR analysis in t(8;21) AML patients with HLA-A0201 who are in long-term remission. PMID- 11122106 TI - Expression of tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) receptors and sensitivity to TRAIL-induced apoptosis in primary B-cell acute lymphoblastic leukaemia cells. AB - Because tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) (Apo2 ligand) preferentially kills malignant cells while sparing normal cells, it may be therapeutically useful against cancers, including those of haematopoietic origin. Although the activity of TRAIL has been studied in tumour cell lines and in a limited number of different primary tumours, its overall activity in a large number of uniform cases of primary tumours is not known. We therefore studied the activity of TRAIL in 29 primary precursor B-cell acute lymphoblastic leukaemia (ALL) samples. TRAIL was found to have a modest activity as it killed a maximum of 29% of ALL cells within 18 h compared with killing 75% of Jurkat cells. The sensitivity to TRAIL did not correlate with the pattern of TRAIL receptor expression or FLIP expression, as determined by Western blot analysis. The CD40 receptor, which can transduce survival signals in mature malignant B cells, was less frequently expressed on ALL cells, but incubation with an exogenous soluble CD40 ligand trimer did not rescue them from spontaneous apoptosis and did not mediate their resistance to TRAIL. Further, although ALL cells expressed TRAIL protein, they failed to kill target Jurkat cells in a TRAIL-dependent manner. Our data delineate major biological differences between mature and precursor malignant B cells and suggest a limited therapeutic role for TRAIL as a single agent in primary B-cell ALL. PMID- 11122107 TI - Testing Sokal's and the new prognostic score for chronic myeloid leukaemia treated with alpha-interferon. Italian Cooperative Study Group on Chronic Myeloid Leukaemia. AB - As it has been shown that alpha-interferon (alphaIFN) treatment modifies the survival of chronic myeloid leukaemia (CML) patients in comparison with conventional chemotherapy, a new prognostic score was devised with the aim of providing a treatment-adapted risk evaluation. We have tested the new prognostic score (the Euro score) in an independent series of 272 patients less than 56 years old with previously untreated, chronic phase, Philadelphia (Ph)-positive CML who had been assigned prospectively to alphaIFN treatment between 1989 and 1991. The Sokal score system was used as a reference. The new Euro score predicted the response to alphaIFN as the Sokal score. The median survival of low risk, intermediate-risk and high-risk patients was similar using the Euro score (105, 65 and 45 months) and Sokal score (105, 76 and 45 months) but, by multivariate analysis, the Euro was more potent than Sokal for predicting survival time. The new Euro score identified more low-risk cases (59% vs. 48%) and fewer high-risk cases (9% vs. 23%) than the Sokal score. The main differences between the Euro and Sokal scores concerned age (it is more important in the Euro than in Sokal), spleen size and the percentage of myeloblasts in peripheral blood (more important in Sokal than in Euro). We conclude that the new Euro score marks an improvement in the prognostic evaluation of CML treated with alphaIFN. By comparison with the Sokal score, the Euro was more potent and identified more low risk patients but left only a small number of cases in the high-risk group. PMID- 11122108 TI - Interferon alpha in combination with GM-CSF induces the differentiation of leukaemic antigen-presenting cells that have the capacity to stimulate a specific anti-leukaemic cytotoxic T-cell response from patients with chronic myeloid leukaemia. AB - Although interferon alpha (IFN-alpha) is able to induce haematological remission in 60-80% of patients with chronic myeloid leukaemia (CML) in early chronic phase, major cytogenetic remissions are only achievable in 30-40%. Recent clinical data suggest that the addition of granulocyte-macrophage colony stimulating factor (GM-CSF) to IFN-alpha therapy can significantly improve the cytogenetic response in some patients, although the mechanism remains unknown. We hypothesized that the combination of GM-CSF and IFN-alpha induces the differentiation of dendritic cells, which subsequently stimulates a specific anti leukaemic response. Monocytes from CML patients were cultured in GM-CSF and interleukin (IL)-4 (GM/IL-4)or in GM-CSF and IFN-alpha (GM/IFN-alpha). After 7 d, the number of cells exhibiting typical antigen-presenting cell (APC) morphology was equal in both groups, and fluorescence in situ hybridization (FISH) analysis confirmed that the APCs generated with GM/IFN-alpha were of leukaemic origin. Phenotypically, both sets of APCs expressed typical surface markers; however, CD86, CD83, CD11c, HLA-ABC and HLA-DR expression was significantly higher in the GM/IFN-alpha APCs, whereas CD1a expression was significantly lower. In mixed lymphocyte reactions (MLR), GM/IFN-alpha APCs stimulated the proliferation of allogeneic T cells significantly better than GM/IL-4 APCs. However, both groups of APCs stimulated autologous T-cell proliferation equally. Finally, we assessed the ability of GM/IFN-alpha APCs to induce a leukaemia-specific cytotoxic T-cell response. Some samples generated cytotoxic T lymphocytes (CTLs) that specifically lysed bcr-abl-positive target cells. These data show that the combination of GM CSF and IFN-alpha, when used in vitro, induces the differentiation of malignant APCs with potent T-cell stimulatory capacity. Although there is no in vivo evidence to support these findings, it is possible that, when administered to CML patients, GM-CSF in combination with IFN-alpha results in the generation of highly stimulatory leukaemic APCs. PMID- 11122109 TI - Autologous T lymphocytes may specifically recognize leukaemic B cells in patients with chronic lymphocytic leukaemia. AB - This study analysed a naturally occurring specific cellular immunity against tumour cells in chronic lymphocytic leukaemia (CLL) patients. Five out of eight patients had blood T lymphocytes able to recognize spontaneously and specifically the autologous tumour B cells (proliferation assay). In these five patients, detection of cytokines by real-time reverse transcription polymerase chain reaction (RT-PCR) revealed that granulocyte-macrophage colony-stimulating factor (GM-CSF) was the most abundant cytokine gene expressed by the T cells that recognized the autologous tumour B cells. Other activated cytokine genes were gamma-interferon (IFN), interleukin (IL)-2 and tumour necrosis factor (TNF) alpha, but not IL-4. This profile suggests a type 1 anti-B-CLL T-cell response. CD80 and CD54 were relatively downregulated on the native tumour B cells compared with control normal B cells. Upregulation of CD80 on the leukaemic cells was mandatory for the induction of such a specific T-cell response. CD80 and CD54 monoclonal antibodies inhibited the specific T-cell DNA synthesis proliferation. The proliferative T-cell response was either MHC class I or class II restricted (inhibition by monoclonal antibodies). The specific cytokine gene expression could be found in isolated CD4, as well as CD8, T-cell subsets. This study demonstrated the presence of a potential natural specific CD4, as well as a CD8 type 1 T-cell immunity against the leukaemic CLL tumour B cells in CLL. A further detailed analysis of the spontaneous anti-CLL T-cell immunity is warranted that may facilitate the development of effective anti-tumour vaccines in CLL. PMID- 11122110 TI - The use of real-time quantitative polymerase chain reaction and comparative genomic hybridization to identify amplification of the REL gene in follicular lymphoma. AB - Using comparative genomic hybridization (CGH), aberrations in DNA copy number were studied before and after transformation of follicular lymphoma to diffuse large B-cell lymphoma in six patients (15 lymph node biopsies in total). The most common and also the most discrete and intense amplification occurring in four out of 15 biopsies from three different patients was of 2p13-16. Using real-time quantitative polymerase chain reaction (RQ-PCR), REL amplification was found to be implicated at this locus. This technique also identified amplified REL in a further two biopsies, presumably below the detection level of CGH. REL amplification was quantified by comparing it, in most cases, with three endogenous reference genes, albumin, beta2-microglobulin and CD8alpha, that lie close to REL on 2p. There was no correlation apparent between 2p13-16 amplification or REL amplification and transformation. This study shows the usefulness of coupling CGH, for detecting recurring abnormalities, with the real time PCR technique for rapid gene dosage quantification and confirms that the REL gene is a potential candidate in the pathogenesis of a particular subset of follicular lymphomas. PMID- 11122111 TI - Insulin-like growth factor induces the survival and proliferation of myeloma cells through an interleukin-6-independent transduction pathway. AB - Multiple myeloma (MM) is a B-cell neoplasia that is associated with an increased level of bone resorption. One important mediator of bone remodelling, insulin like growth factor (IGF-I), has been shown to stimulate the proliferation of human myeloma cells. However, the mechanisms of action of IGF-I in these cells have not been determined. Using interleukin (IL)-6-dependent myeloma cell lines, we show IGF-I to be as potent a survival and proliferation factor as IL-6. We demonstrated that IGF-I functions independently of the IL-6 transducer gp130 and that these two cytokines have additive effects. Moreover, inhibition of the IGF-I pathway did not modulate the proliferative effect of IL-6. Accordingly, we found that IL-6 and IGF-I activated distinct downstream signalling molecules: IL-6 activated STAT3 phosphorylation, whereas IGF-I treatment resulted in the phosphorylation of IRS-1. Interestingly, these signalling pathways appear to converge as both cytokines activated the ras/MAPK pathway. Thus, IGF-I acts as a potent survival and proliferation factor for myeloma cells by stimulating an IL-6 independent signalling cascade. These data, together with the finding that, in vivo, IGF-I is normally expressed in close proximity to myeloma cells within the bone matrix, strongly suggest a role for this cytokine in the pathophysiology of multiple myeloma. PMID- 11122112 TI - Prospective screening by a panfungal polymerase chain reaction assay in patients at risk for fungal infections: implications for the management of febrile neutropenia. AB - Invasive fungal infections are a major cause of mortality in neutropenic cancer patients. To determine whether a polymerase chain reaction (PCR)-based assay enabled the identification of patients at risk for invasive fungal infections, a prospective monitoring once per week was performed during 92 neutropenic episodes in patients receiving chemotherapy for acute leukaemia or high-dose therapy followed by allogeneic or autologous stem cell transplantation, with the investigators blinded to clinical and microbiological data. PCR positivity was documented in 34 out of 92 risk episodes. All patients developing proven invasive fungal infection were found PCR positive, and PCR was found to be the earliest indicator of invasive fungal infection preceding clinical evidence by a mean of 5.75 d (range 0-14 d). In febrile neutropenic patients without a prior history of invasive fungal infection, a sensitivity of 100% and a specificity of 73% of the PCR assay for the development of proven or probable invasive fungal infection was documented. In conclusion, panfungal PCR performed prospectively once a week enabled the identification of patients at high risk for invasive fungal infections. PMID- 11122113 TI - Spontaneous regression of congenital leukaemia with an 8;16 translocation. AB - Congenital leukaemia (CL) is a rare disorder that presents with extramedullary infiltrates and a myeloid phenotype. CL can progress rapidly without adequate treatment, but, paradoxically, may also remit spontaneously. Because of the significant toxicity involved in delivering chemotherapy to newborns, it is important to identify those newborns who may not require treatment. We describe an infant who presented at 1 week of age with congenital myeloid leukaemia. Cytogenetic analysis revealed a t(8;16)(q11;p13) translocation. The infant's leukaemia underwent a spontaneous regression. This case further confirms the possibility of spontaneous remission in congenital leukaemia. Moreover, it suggests that the presence of a clonal cytogenetic aberration does not preclude the possibility of a spontaneous regression in CL. PMID- 11122114 TI - Novel type of BCR-ABL transcript in a chronic myelogenous leukaemia patient relapsed after bone marrow transplantation. AB - We identified a novel BCR-ABL transcript in a chronic myelogenous leukaemia (CML) patient who relapsed after bone marrow transplantation (BMT), containing a fusion between part of BCR exon 3, 44 nucleotides derived from ABL intron 1b and ABL exon 2. The breakpoints were located within BCR exon 3 on chromosome 22 and within the ABL intron 1b on chromosome 9, and the transcript derives from a splicing of ABL exon 2 to a putative splicing acceptor site 44 nucleotides downstream to the breakpoint on chromosome 9. The patient's clinical course strengthens the idea that short forms of BCR-ABL transcripts are associated with a more aggressive disease. PMID- 11122115 TI - The 8p12 myeloproliferative disorder. t(8;19)(p12;q13.3): a novel translocation involving the FGFR1 gene. AB - Translocations affecting the chromosomal locus 8p12 are hallmarks of an atypical stem cell myeloproliferative disorder. These events disrupt the fibroblast growth factor receptor 1 (FGFR1) gene and fuse the FGFR1 C-terminus catalytic domain with unrelated proteins. Here, we report on the characterization of the 19q13.3 locus as the fifth FGFR1 chromosomal partner. PMID- 11122116 TI - Differential cellular expression of the human MSH2 protein in normal and myelodysplastic haematopoiesis. AB - Loss of human MSH2 (hMSH2) protein might be involved in the multistep pathogenesis of haematological malignancies associated with genetic instability. Here, we examine cellular hMSH2 expression in bone marrow samples from 10 haematopoietically normal individuals in comparison with nine patients with myelodysplastic syndrome (MDS) [one refractory anaemia (RA), two RA with ringed sideroblasts (RARS), four RA with excess blasts (RAEB) and two RAEB in transformation (RAEB-T)]. HMSH2 protein was predominantly expressed in myeloblasts and promyelocytes. Blast cells from three patients with RAEB and one with RAEB-T displayed absent or very low hMSH2 expression. As no correlation between hMSH2 expression and chromosomal aberrations was observed, further genetic events seem to be required to induce karyotype instability. PMID- 11122117 TI - Absence of mutations in the granulocyte colony-stimulating factor (G-CSF) receptor gene in patients with myelodysplastic syndrome/acute myeloblastic leukaemia occurring after treatment of aplastic anaemia with G-CSF. AB - The development of myelodysplastic syndrome/acute myeloblastic leukaemia (MDS/AML) has been reported in patients with aplastic anaemia (AA) after administration of recombinant human granulocyte colony-stimulating factor (rhG CSF). Similarly, patients with severe congenital neutropenia (SCN) have an increased risk of developing MDS/AML after treatment with rhG-CSF. Point mutations in the G-CSF receptor gene are found in about 20% of SCN patients who are predisposed to MDS/AML. We investigated the occurrence of mutations in the G CSF receptor in eight patients with AA who developed MDS/AML. No mutations were detected around the cytoplasmic domain of the gene in our patients, indicating that the mechanisms of clonal evolution to MDS/AML in patients with AA might be different from those with SCN. PMID- 11122118 TI - Parvovirus B19 transmitted by bone marrow. AB - We describe a case of symptomatic parvovirus B19 infection transmitted by bone marrow (BM). The infection caused prolonged anaemia, thrombocytopenia, arthralgia and erythema infectiosum in a 16-year-old girl with acute myeloid leukaemia receiving a BM transplant (BMT). The BM donor was a 19-year-old asymptomatic brother who had parvovirus B19 viraemia at the time of BM harvest. Sequencing of the VP2 gene from the patient and the donor showed a perfect match of DNA sequences, confirming the mode of transmission. Parvovirus B19 represents a potential complicating factor in patients undergoing BMT, but screening by polymerase chain reaction (PCR) of donor BM may reduce the risk of infection. PMID- 11122119 TI - Encouraging preliminary results in 12 patients with high-risk haematological malignancies by omitting graft-versus-host disease prophylaxis after allogeneic transplantation. AB - Immunosuppressive therapy, routinely given after allogeneic transplantation to modulate graft-versus-host disease (GVHD) may have an adverse effect on the graft versus-tumour (GVT) effect. Twelve patients with high-risk haematological malignancies were given cyclophosphamide, total body irradiation and antithymocyte globulin followed by peripheral blood stem cell grafts from HLA identical siblings without prophylactic immunosuppression. At the earliest clinical evidence of GVHD, patients were treated with high-dose solumedrol and tacrolimus. Prompt haematological recovery [absolute neutrophil count (ANC) > 1.0 x 109/l] was observed (median time 9 d). All patients developed grade III-IV GVHD (median onset 9 d), involving the skin (11), intestine (five) and liver (three). Of nine evaluable patients, seven developed chronic GVHD [extensive (six), limited (one)]. Six patients died 1-6.5 months after transplantation. Three patients died from treatment-related complications, two from acute GVHD and one from relapsing disease. The remaining six patients are alive 5-26 months after transplantation, five in complete remission and one myeloma patient in very good partial remission. In conclusion, omission of post-transplantation GVHD prophylaxis is feasible, does not lead to graft failure or a high incidence of uncontrollable GVHD and appears to be associated with encouraging clinical responses in a group of patients with high-risk disease features. PMID- 11122120 TI - OKT-3-based reconditioning regimen for early graft failure in HLA-non-identical stem cell transplants. AB - Primary non-engraftment or early rejection after transplantation of haematopoietic stem cells represent life-threatening complications of allogeneic stem cell transplantation. Management of early graft failure has been problematic, as the risk of fatal infectious complications increases with the time of pancytopenia and as a second transplant preceded by a conventional myeloablative conditioning regimen has been associated with high rates of cumulative organ toxicity. For paediatric patients with early graft failure following the transplantation of highly purified major histocompatibility complex (MHC)-disparate haematopoietic stem cells, we have evaluated an immunosuppressive OKT-3/methylprednisolone-based reconditioning regimen with low toxicity in preparation for a secondary transplant of purified haematopoietic stem cells from the same donor. This report presents the results from a 4-year pilot study including six patients with early graft failure. The results demonstrate that this antibody-based regimen can be used effectively to prepare patients for secondary transplantation. Successful engraftment after a second transplant procedure was achieved in five of these six high-risk patients. The median interval between first and second transplant was 27 d (range 22-51 d), and the median time for engraftment was 10 d (range 9-13 d). Chimaerism analysis of microsatellite regions by polymerase chain reaction (PCR) demonstrated complete donor chimaerism in four of these patients within the first month after secondary transplant and revealed mixed chimaerism in one patient who converted to complete chimaerism after T-cell add-back. PMID- 11122121 TI - Stroma-supported progenitor production as a prognostic tool for graft failure following autologous stem cell transplantation. AB - To analyse the involvement of a possible numerical or qualitative stem cell defect in the development of sustained graft failure after autologous transplantation, we have determined the graft content of CD34+ nucleated cells, colony-forming cells and cobblestone area-forming cell subsets, as well as transplant ability to produce progenitors using the long-term culture colony forming cell (LTC-CFC) assay. We evaluated material from the graft reference ampoules of 13 graft failure patients after bone marrow transplantation (BMT), four graft failure patients and four isolated thrombocytopenia patients after peripheral blood stem cell transplantation (PBSCT). We compared these data with those from six successfully engrafted BMT patients and 20 engrafted PBSCT patients respectively. In the BMT setting, the LTC-CFC 6-week assay represented a highly significant graft failure predictor. In the PBSCT setting, the total number of 2-week and 6-week LTC-CFCs transplanted per kg bodyweight (BW) showed the highest significant difference between the engrafted and the graft failure patients, as well as between the engrafted patients and the patients suffering from isolated thrombocytopenia after transplantation. These data show that the ability of a graft to generate progenitors in vitro rather than the number of primitive progenitors transplanted can have prognostic value for post-transplant haematological reconstitution. PMID- 11122122 TI - Cytotoxic T-lymphocyte precursor frequency (CTLp-f) as a tool for distinguishing permissible from non-permissible class I mismatches in T-cell-depleted allogeneic bone marrow transplantation. AB - Matching for HLA has been the gold standard in bone marrow donor selection. But, with the ever increasing number of identified HLA alleles, it is becoming more difficult to find a fully HLA-identical donor other than a sibling. Retrospective analysis revealed that HLA mismatches do not necessarily give rise to acute graft versus-host-disease (GVHD). However, we have no means of defining these 'permissible' mismatches before bone marrow transplantation (BMT). Thus, we set out to establish whether functional matching by means of helper and cytotoxic T lymphocyte precursor frequency analysis (HTLp-f and CTLp-f respectively) can be applied to this end. Fifty-five recipient-donor pairs other than HLA-identical siblings, the recipient of which received a T-cell-depleted graft, were analysed by high-resolution HLA typing and/or HTLp-f/CTLp-f analysis. The predictive value of the CTLp-f assay for development of acute GVHD was confirmed. More importantly, our data indicate that the CTLp-f assay was able to discriminate permissible from non-permissible HLA-A, -B or -Cw mismatches, but not for DRB/DQB mismatches. The absolute number of alloreactive CTLs present in the graft correlated with the risk of acute GVHD. Although HTLp-f and CTLp-f together had a high negative predictive value, HTLp-f outcome by itself was not correlated with acute GVHD. As we have no evidence yet that HTLp-f or CTLp-f can identify permissible DRB/DQB mismatches, high-resolution matching for these antigens remains the best option. The combination of high-resolution DRB/DQB typing and the CTLp-f assay would enable the accurate prediction of the risk of acute GVHD while extending the pool of potential donors. Furthermore, it would enable adjustment of the number of T- cells in the graft accordingly to improve clinical outcome. PMID- 11122123 TI - Respiratory burst of neutrophils is not required for stem cell mobilization in mice. AB - It has been suggested that mature neutrophils may play an essential role in the cascade of events leading to egress of stem cells from the bone marrow to the peripheral blood. To investigate further the role of mature neutrophils and of reactive oxygen intermediates (ROIs), known to be involved in the signal transduction of neutrophils, we used mice deficient in respiratory burst, and thus the production of ROIs, to study the involvement of this activation pathway in stem cell mobilization. B6 mice with chronic granulomatous disease (CGD) received either cyclophosphamide (200 mg/kg) on day 1 and granulocyte colony stimulating factor (G-CSF) (250 microg/kg/d) on days 3-6 or a single dose of interleukin 8 (IL-8; 30 microg/mouse) as a mobilization regimen. On day 7, the number of stem and progenitor cells in blood and bone marrow was compared with control B6 animals (with intact respiratory burst). White blood cell counts, bone marrow cellularity and the frequency of granulocyte-macrophage colony-forming cells (GM-CFC), and cobblestone area-forming cells (CAFC) on days 7 (CAFC-7) and 28 (CAFC-28) were determined. After cyclophosphamide and G-CSF (CY + G), both mouse strains showed considerable mobilization of CAFC-7 and CFU-GM to the blood. Normal mice showed up to a 1905-fold increase in progenitors per ml blood, whereas CGD mice showed up to a 264-fold increase in blood progenitors. IL-8 also induced mobilization in both mouse strains. In addition to progenitors, primitive stem cells measured as CAFC-28 and as CAFC at day 35 were also mobilized by both mobilization protocols in normal as well as in CGD mice. In conclusion, respiratory burst and the subsequent signal transduction pathway do not appear to be required for mobilization of stem cells. Accordingly, neutrophils either are not involved in stem cell mobilization or other signalling pathways within neutrophils must exist that lead to the release of factors which activate stem cell egress from the bone marrow. PMID- 11122124 TI - The reconstitution of CD45RBhiCD4+ naive T cells is inversely correlated with donor age in murine allogeneic haematopoietic stem cell transplantation. AB - A high incidence of opportunistic infections after unrelated bone marrow transplantation has been reported. Delayed lymphocyte recovery may be associated with opportunistic infections. Immune reconstitution is influenced by recipient age and graft-vs-host disease (GVHD). In fact, children develop GVHD less frequently than adults. However, the role of donor age is largely unknown. We examined the effect of donor age on lymphocyte reconstitution after transplant. Three-month-old BALB/c recipient mice were lethally irradiated and transplanted with allogeneic haematopoietic stem cells from A/J donor mice of different ages, ranging from 0 d to 12 months. The recovery of absolute lymphocyte counts and those of CD3+ T cells, CD4+ T cells and CD45RBhi CD4+ naive T cells in the early post-transplant period correlated inversely with donor age. Recipient mice transplanted with haematopoietic stem cells from younger donors showed significantly higher survival rates and mitogenic responses than adult donors. As T cells, especially CD4+ naive T cells, play an important role in host defence, faster recovery of CD4+ naive T cells in younger donors may contribute to reduced mortality in the early post-transplant period. The results suggest that it could be better to choose a younger donor if sufficient cell dose is available. PMID- 11122125 TI - Graft-versus-tumour effect in non-small-cell lung cancer after allogeneic peripheral blood stem cell transplantation. AB - Clinical evidence of a graft-vs.-tumour effect in solid tumours after haematopoietic stem cell transplantation is lacking. We report for the first time a complete and durable regression of a stage IB non-small-cell lung carcinoma in a patient who had received an allogeneic peripheral blood haematopoietic stem cell transplant for acute myeloblastic leukaemia in first complete remission. Disappearance of the tumour coincided with development of graft-vs. -host disease. This suggests that simultaneous generation of cytotoxic T lymphocytes against lung carcinoma cells could have been responsible for the regression. This unique clinical observation broadens the possibility of using allogeneic haematopoietic stem cell transplantation in treating neoplasias lacking significant sensitivity to chemotherapy. PMID- 11122126 TI - Meisoindigo for the treatment of chronic myelogenous leukaemia. PMID- 11122127 TI - Myelodysplastic syndrome/acute myelogenous leukaemia related to adjuvant chemotherapy with oral pyrimidine anti-metabolites. PMID- 11122128 TI - Langerhans cell histiocytosis. PMID- 11122129 TI - Type C Niemann-Pick disease. PMID- 11122130 TI - The early history of haemophilia treatment: a personal perspective. PMID- 11122131 TI - Antenatal screening for fetomaternal alloimmune thrombocytopenia: an evaluation using the criteria of the UK National Screening Committee. PMID- 11122132 TI - Progress in the assessment of platelet function. PMID- 11122133 TI - Clinical course and predictive factors for cyclosporin-induced autologous graft versus-host disease after autologous haematopoietic stem cell transplantation. AB - The administration of cyclosporin A (CyA) after autologous haematopoietic stem cell transplantation (HSCT) induces a systemic autoimmune syndrome mimicking graft-vs.-host disease (GVHD). This syndrome, termed autologous GVHD has notable anti-tumour activity in animal studies. We intended to induce autologous GVHD with CyA in patients undergoing an autologous HSCT. We prospectively studied 118 patients with miscellaneous malignancies undergoing an autologous HSCT with low dose CyA to characterize the clinical syndrome, its frequency and clinical course, and to determine the factors affecting its incidence. Patients received CyA from d -1 through to d 28, first starting at 2 mg/kg intravenously and then orally as soon as feasible. The dose was adjusted to achieve pre-dose blood levels around 100 ng/ml. A skin biopsy was performed when a skin rash was observed. Thirty-three percent of the patients developed clinical GVHD: clinical stage 1 in 21 patients, stage 2 in seven patients, and stage 3 in three patients. Although total body irradiation (TBI) or high-dose cyclophosphamide were previously thought to be needed, autologous GVHD occurred in five out of 12 patients (42%) after a preparative regimen with high-dose melphalan alone. Autologous GVHD was significantly more frequent in patients older than 33 years, in patients who had received high doses of granulocyte-macrophage colony forming units (CFU-GM) and in those with a diagnosis of myeloid malignancy, compared with those with lymphoid malignancies or solid tumours. A significant negative association was also found with HLA-DR6. In lymphoma patients, GVHD occurred more frequently in advanced disease than in first or second complete remission (CR1-2) patients. All other factors studied were not predictive for GVHD. In conclusion, CyA-induced GVHD is reproducibly and safely induced with doses of CyA adapted to achieve blood levels around 100 ng/ml. In retrospective analysis, there was no survival advantage for patients with GVHD. Phase III trials with this approach are needed to evaluate its anti-tumoral effect. PMID- 11122134 TI - Late outcomes after bone marrow transplant for aplastic anaemia. AB - Allogeneic transplantation is effective in reconstituting haemopoiesis in severe aplastic anaemia (SAA). We report long-term health-related outcomes in 37 children and young adults with SAA transplanted between 1975 and 1996. The median length of follow-up was 17 years (range, 4-25 years). Using a case-control design, late social and medical outcomes in transplant recipients were compared with 146 control subjects matched for gender and age. The majority of patients received an irradiation-containing preparative regimen. There were no significant differences in the self-rating of health status between transplant recipients and controls (P = 0.8), with 71% reporting their health status as excellent and 29% as good compared with 74% and 26% of controls. They demonstrate the same normal psychosexual function as their peers and have similar educational achievements and employment history. Transplant recipients and controls are equally likely to have held a job or be currently employed and there are no significant differences in their personal income (OR = 0.60, 95% CI = 0.11-3.37). Although transplant recipients have had problems related to health insurance policies, the majority have adequate health insurance coverage. There were no differences in chronic health problems between transplant recipients and control subjects, except for expected increases in cataracts, short stature in men, hypothyroidism and gonadal dysfunction. Using self-assessment, these transplant recipients indicated an excellent level of satisfaction and social integration, showing transplantation to be an effective long-term therapy for SAA. PMID- 11122135 TI - A randomized comparison of once versus twice daily recombinant human granulocyte colony-stimulating factor (filgrastim) for stem cell mobilization in healthy donors for allogeneic transplantation. AB - To evaluate the schedule dependency of granulocyte colony-stimulating factor (G CSF) (filgrastim) for stem cell mobilization, we conducted a randomized comparison in 50 healthy donors, with one subcutaneous daily injection of 10 microg/kg G-CSF (n = 25) compared with twice injections daily of 5 microg/kg G CSF (n = 25). The two groups were well balanced for age, body weight and sex. G CSF application was performed on an out-patient basis and leukapheresis was started in all donors on day 5. The most frequent side-effects of G-CSF were mild to moderate bone pain (88%), mild headache (72%), mild fatigue (48-60%) and nausea (8%) without differences between the two groups. The CD34(+) cell count in the first apheresis was 5.4 x 10(6)/kg donor weight (range 2.8-13.3) in the 2 x 5 microg/kg group compared with 4.0 x 10(6)/kg (range 0.4-8.8) in the 1 x 10 microg/kg group (P = 0.007). The target of collecting > 3.0 x 10(6) CD34(+) cells/kg donor weight with one apheresis procedure was achieved in 24/25 (96%) donors in the 2 x 5 microg/kg group and in 17/25 (68%) donors in the 1 x 10 microg/kg group. The target of collecting > 5.0 x 10(6) CD34(+) cells/kg in the first apheresis was achieved in 64% in the 2 x 5 microg/kg group, but in only 36% in the 1 x 10 microg/kg group. The progenitor cell assay for granulocyte macrophage colony-forming units (CFU-GM) and erythroid burst-forming units (BFU E) was higher in the 2 x 5 microg/kg group than in the 1 x 10 microg/kg group (7.0 vs. 3.5 x 10(5)/kg, P = 0.01; 6.6 vs. 5.0 x 10(5)/kg; P = 0.1). Administering G-CSF (filgrastim) at a dosage of 5 microg/kg twice daily rather than 10 microg/kg once daily is recommended; this leads to a higher CD34(+) cell yield and requires fewer apheresis procedures without increasing toxicity or cost. PMID- 11122136 TI - Persistence of clonal T-cell expansions following high-dose chemotherapy and autologous peripheral blood progenitor cell rescue. AB - Analysing the regeneration of T lymphocytes after high-dose chemotherapy with autologous peripheral blood progenitor cell rescue (PBPCR) may help elucidate the mechanisms of immune recovery. The T-cell receptor variable beta chain (TCRBV) repertoire of adult patients undergoing high-dose chemotherapy was analysed by flow cytometry, before and after treatment. Four patients were found to have a stable expansion present (TCRBV3, 17, 21 and 22) ranging from 8% to 42% of the CD4(+) or CD8(+) repertoire. We demonstrated that, in these patients, following high-dose chemotherapy and autologous stem cell transplantation, the clonal expansions reappeared in peripheral blood and returned to pretransplant levels. Three expansions (CD3(+)CD8(+)TCRBV3(+), CD3(+)CD4(+)TCRBV21(+) and CD3(+)CD8(+)TCRBV22(+)) were further defined by sequence analysis of the complementarity-determining region (CDR)3 portion within the TCR rearrangements. These were shown to be predominantly clonal, with the same sequences being identified in peripheral blood before and after PBPCR, providing evidence that the overwhelming majority of T cells in these expansions arise from mature lymphocytes. This study demonstrated that patients undergoing autologous PBPCR for high-dose chemotherapy regenerate clonal expansions, consistent with pretreatment levels. They also regenerate T-cell repertoires with each TCRBV family represented to a similar level as that prior to high-dose chemotherapy. PMID- 11122137 TI - High levels of human herpesvirus 6 DNA in peripheral blood leucocytes are correlated to platelet engraftment and disease in allogeneic stem cell transplant patients. AB - The aim of this study was to correlate human herpesvirus (HHV)-6 viral load with clinical symptoms in allogeneic stem cell transplant (SCT) patients. Seventy-four patients were monitored during the first 3 months after SCT using a qualitative polymerase chain reaction (PCR) for HHV-6 DNA. HHV-6 was detected in 181 out of 494 samples (36%) from 58 (78%) patients. These 181 samples were analysed using a quantitative competitive PCR. DNA could be quantified from 146 out of 181 samples (80.6%). The HHV-6 viral load was highest at 4 weeks compared with 8 weeks (P < 0.001) and 12 weeks (P = 0.01) after SCT. Three patients had HHV-6 encephalitis and one patient had hepatitis. The HHV-6 DNA levels were higher in patients with HHV-6 than in those without HHV-6 (P = 0.01). Patients who received grafts from unrelated or HLA-mismatched family donors had significantly higher HHV-6 DNA levels than patients who received grafts from matched sibling donors (P < 0.001). In a multiple regression model, unrelated donor grafts (P < 0.001) and use of intravenous immunoglobulin prophylaxis (P = 0.04) influenced HHV-6 DNA levels. HHV-6 viral load was significantly correlated with delayed platelet engraftment in both univariate (P < 0.01) and multivariate analysis, and to the number of platelet transfusions. PMID- 11122138 TI - Extended routine polymerase chain reaction surveillance and pre-emptive antiviral therapy for cytomegalovirus after allogeneic transplantation. AB - Pre-emptive treatment strategies based on sensitive screening for cytomegalovirus (CMV) infection up to day +100 after allogeneic transplantation have been shown to reduce the incidence of CMV disease during the period of surveillance. However, the use of ganciclovir has been associated with delays in immune reconstitution and an increased incidence of late CMV disease after day +100. In the present study, 81 patients undergoing allogeneic transplantation received polymerase chain reaction (PCR)-guided pre-emptive therapy based on detection of CMV DNA by PCR on 2 consecutive weeks up to day +180. Thirty-three of the 52 high risk patients (CMV-seropositive donor or recipient) received a total of 45 treatment episodes up to day +100. Three of these patients (5.7%) developed CMV disease, with one fatality. Twelve of the surviving 44 high-risk patients (27%) required pre-emptive treatment between days +101 and +192, but none of these patients developed late CMV disease with a median follow-up of 402 d (range 117 952 d). Antiviral therapy was stopped after a single negative PCR result with no subsequent episodes of CMV disease while patients remained off antiviral treatment. As all initial episodes of CMV DNA detection occurred within 60 d of transplantation, it may be possible to discontinue monitoring beyond day +100 in patients who have remained CMV PCR negative before this. Thus, we have confirmed that PCR-guided pre-emptive therapy results in a low incidence of CMV disease before day +100 and that discontinuing treatment on the basis of viral clearance as determined by CMV PCR appears to be safe practice. In addition, we have observed no episodes of late CMV disease with an extension of surveillance to 26 weeks. PMID- 11122139 TI - Primary human keratinocytes as targets in predicting acute graft-versus-host disease following HLA-identical bone marrow transplantation. AB - Graft-versus-host-disease (GVHD) remains a major problem following allogeneic bone marrow transplantation (BMT) and manifests itself mainly by damage to epithelial cells of the skin, gut and bile ducts. Reliable tests to predict GVHD are lacking. We developed an assay in which donor T cells are stimulated by patient keratinocytes (KCs), compared that with stimulation by patient peripheral blood mononuclear cells (PBMCs) and studied the relationship to GVHD. In 27 patients undergoing HLA-identical BMT for haematological malignancies, donor T cell reactivity was determined as the helper T-lymphocyte precursor (HTLp) frequency against host PBMCs (25 patient-donor pairs) and host KCs (20 patient donor pairs). KCs were obtained by shave biopsies and cultured with interferon (IFN)-gamma to induce HLA class II expression. In assays using patient KCs and donor T cells, anti-CD28 antibody was added to compensate for the lack of co stimulatory molecules on KCs. Results were related to the occurrence of GVHD. As BMTs were performed with partially T cell-depleted grafts, GVHD was limited to grade 0 (five patients), grade I (seven patients) and grade II (12 patients). No differences were found in donor T-cell reactivity to patient PBMCs, as expressed as HTLp frequency in patients with or without GVHD. However, significant differences (P < 0.01) were found in donor T-cell reactivity to patient KCs when comparing patients with and without GVHD. Donor HTLp frequencies against patient KCs give a better prediction of GVHD than those against patient haemopoietic cells following HLA-identical BMT, which may indicate that at least some minor non-HLA histocompatibility antigens present on KCs are different from those on haemopoietic cells. PMID- 11122140 TI - Short Report: Engraftment of T-cell-depleted allogeneic haematopoietic stem cells using a reduced intensity conditioning regimen. AB - Graft-versus-host disease (GVHD) remains a significant complication in patients undergoing allogeneic stem cell transplantation (SCT) using a reduced intensity conditioning regimen. Although T-cell depletion (TCD) reduces the risk of GVHD after a myeloablative conditioning regimen, it is associated with an increased risk of graft failure. We have therefore examined whether TCD compromises engraftment using a fludarabine-based conditioning regimen. Fifteen patients have been transplanted using such a regimen of whom 13 underwent ex vivo TCD. All but one patient demonstrated durable engraftment and no patient receiving a TCD product developed severe GVHD. Thus, TCD may play a role in GvHD prophylaxis using such regimens. PMID- 11122141 TI - Additional chromosomal abnormalities in patients with acute promyelocytic leukaemia (APL) do not confer poor prognosis: results of APL 93 trial. AB - In spite of the recent improvement in the outcome of acute promyelocytic leukaemia (APL) with treatment combining all trans retinoic acid (ATRA) and chemotherapy (CT), some patients with this disease still have a poor outcome. The prognostic significance of chromosomal abnormalities in addition to t(15;17) in APL is uncertain. We examined the prognostic significance of secondary chromosomal changes in 292 patients included in a European trial who were treated with ATRA and CT. The incidence of chromosomal abnormalities in addition to t(15;17) was 26% and trisomy 8 was the most frequent secondary change (46% of the cases with secondary changes). No significant differences were seen with regard to age, sex, initial white blood cell count, % of circulating blasts, platelet count, fibrinogen level and incidence of microgranular variants between patients with or without additional rearrangements. Outcome was also similar between patients with t(15;17) alone and patients with t(15;17) and other clonal abnormalities for complete remission (92% vs. 93% respectively), event-free survival at 2 years (76.1% vs. 78.1% respectively), relapse at 2 years (16.7% vs. 11.6% respectively) and overall survival at 2 years (79.9% vs. 79.5% respectively). Analysis according to the type of induction treatment (ATRA followed by CT or ATRA plus CT) or the type of maintenance treatment (with ATRA, low-dose CT or both) also failed to show any difference between the two groups. Thus, in a large cohort of APL patients treated with ATRA and CT, additional chromosomal abnormalities had no impact on prognosis. PMID- 11122142 TI - Upregulation of lipocortin 1 inhibits tumour necrosis factor-induced apoptosis in human leukaemic cells: a possible mechanism of resistance to immune surveillance. AB - The signal transduction pathway through which tumour necrosis factor (TNF) induces apoptosis in leukaemic cells may involve activation of cytosolic phospholipase A(2) (cPLA(2)). The steroids dexamethasone (Dex) and 1,25(OH)(2) D(3) both render U937 leukaemic cells resistant to TNF-induced apoptosis. In this study, we found that Dex inhibited both spontaneous and TNF-induced activation of cPLA(2). Dex had no direct effect on cellular cPLA(2) levels, but facilitated cPLA(2) degradation upon subsequent stimulation of cells with TNF. In addition, Dex increased synthesis of the endogenous cPLA(2) inhibitor lipocortin 1 (LC1). An antisense oligonucleotide to LC1 could completely abrogate Dex-induced resistance to the cytotoxic action of TNF. Constitutive LC1 levels were relatively higher in myeloid leukaemic blasts showing resistance to TNF than TNF sensitive myeloid leukaemic cell lines. Our data suggest that Dex confers the resistance of U937 cells to TNF-induced apoptosis by upregulating intracellular levels of LC1 and by facilitating a negative-feedback loop, which is activated upon stimulation with TNF. High constitutive levels of LC1 in leukaemic blasts may protect them against immune-mediated killing. PMID- 11122143 TI - Impaired expression of the CD3-zeta chain in peripheral blood T cells of patients with chronic myeloid leukaemia results in an increased susceptibility to apoptosis. AB - In patients with myeloid malignancies, cell-mediated immunity is often suppressed, being most profound in those with advanced disease. Such immune dysfunction, as demonstrated in many patients with chronic lymphocytic leukaemia (CLL) and myelodysplastic syndrome (MDS), may, at least in part, be due to altered expression of the CD3-zeta chain, which is an important component of the T-cell receptor (TCR). We speculated that impaired expression of the TCR-zeta chain would be evident in peripheral blood T cells of patients with chronic myeloid leukaemia (CML) and that such an abnormality would result in an increased ex vivo susceptibility to apoptosis. In this study, we demonstrated that, compared with normal controls, zeta chain expression was significantly downregulated in all of the T-cell subsets (P < 0.009) in more than 90% of CML patients. In addition, there was a significantly lower expression of the CD3 epsilon chain (P < 0.001) in patients than in controls. In those patients with abnormal zeta chain expression, the proportion of lymphocytes with spontaneous DNA fragmentation, as determined by terminal deoxynucleotide transferase-mediated dUTP-biotin nick-end labelling (TUNEL) assays, was also significantly higher (P < 0.002) than controls. From all of the patients tested, it was possible to upregulate partially zeta chain expression and hence to reduce the susceptibility to apoptosis by cross-linking the T cells with interleukin (IL)-2, interferon (IFN)-alpha or immobilized CD3. In addition, such cross-linked T cells showed a significantly higher capacity to proliferate than the native CML T cells. PMID- 11122144 TI - An Asian variant of intravascular large B-cell lymphoma: clinical, pathological and cytogenetic approaches to diffuse large B-cell lymphoma associated with haemophagocytic syndrome. AB - Diffuse large B-cell lymphoma with haemophagocytic syndrome (BCL-HS) has been reported mainly in Asia and is regarded as a distinct variant of intravascular lymphoma (IVL). However, it is unclear whether all cases of BCL-HS fall within the framework of IVL and available clinical information is limited. We analysed 25 cases with BCL-HS, including 11 autopsied cases (median, 66 years; male-female ratio, 1.1:1). The patients presented with fever, anaemia, thrombocytopenia, hepatosplenomegaly, haemophagocytosis, bone marrow invasion, respiratory disturbance and disseminated intravascular coagulopathy, but usually lacked lymphadenopathy, mass formation, neurological abnormalities and skin lesions. The clinical course was aggressive with a median survival of 7 months. The morphological findings were uniform: large lymphoid cells infiltrated vessels and/or sinusoids of the liver, marrow, lung, kidney and other organs. They were positive for CD19, CD20, CD79a and HLA-DR, but negative for CD10, CD23 and CD30. CD5 was positive in five out of 17 cases. Our critical review indicates that BCL HS is the equivalent of the Asian variant of IVL. PMID- 11122145 TI - Expression and production of interleukin 10 in human myeloma cell lines. AB - Recent investigations of the cytokine network surrounding myeloma cells have disclosed the importance of gp130-related cytokines including interleukin (IL)-6 for myeloma cell survival and proliferation, identification of IL-10 as a growth factor for myeloma cells, the close relationship between IL-10 and the receptors for gp130-related cytokines, and the growth enhancement effect of IL-11 and IL-7 on myeloma cells. In this study, IL-10 production was observed in three out of seven human myeloma cell lines examined and five (including three producing lines) out of 10 lines exhibited mRNA expression of IL-10. The IL-10 mRNA expression was also enhanced in approximately one third of primary specimens, whereas the IL-10 receptor (R) expression was not changed compared with that of normal component marrow controls. However, reverse transcription-polymerase chain reaction (RT-PCR) assay of various cytokines and their receptors showed no particular association with IL-10-producing myeloma lines compared with non producing lines. Supplementing exogenous IL-10 or neutralization of the IL-10 signal by anti-IL-10 monoclonal antibody (mAb) in a culture conditions did not significantly affect myeloma cell growth regardless of expression of IL-10 or its receptor (IL-10R). However, supplement of anti-IL-10 mAb caused upregulation of certain genes such as IL-11, leukaemia inhibitory factor receptor (LIFR) and syndecan-1 in IL-10R-expressing cell lines. These findings indicate that the cytokine network surrounding myeloma cells is complicated and variable. In addition, IL-10 may modify this network and the cellular biological properties of myeloma cells rather than cell proliferation. PMID- 11122146 TI - Increased apoptosis in acquired sideroblastic anaemia. AB - Idiopathic acquired sideroblastic anaemias (IASAs) form a subgroup of the myelodysplastic syndromes and are characterized by mitochondrial iron accumulation, bone marrow erythroid hyperplasia and decreased peripheral red blood cell counts. Increased intramedullary apoptosis of erythroid precursors is presumed to constitute the pathophysiological mechanism explaining this ineffective erythropoiesis, but if and how mitochondrial dysfunction is implicated in this process is currently unknown. We therefore studied bone marrow precursor cells obtained from nine patients with IASA for (i) caspase 3 activity, (ii) numbers of Annexin V- and 7-amino-actinomycin-positive cells, (iii) numbers of cells with diminished mitochondrial membrane potential, Delta Psi(m), and (iv) numbers of cells producing reactive oxygen species (ROS), and we compared the results with those of five normal bone marrow samples. Compared with controls, we found increased caspase 3 activity in all IASA samples, which correlated with increased numbers of Annexin-V-positive cells (r = 0.7). Analysis of different subpopulations showed increased apoptosis in erythroid populations compared with myeloid and/or lymphoid populations in five out of nine cases, and increased apoptosis in the last two populations in four out of nine cases. As evidence of mitochondrial dysfunction, Delta Psi(m) was found to be diminished in the erythroid subpopulations of all cases of IASA (66.6 +/- 17% vs. 34.6 +/- 12% in normals). Delta Psi(m) decrease was correlated to Annexin V positivity (r = 0.7). Astonishingly, no difference was found between IASA and normal bone marrows with regard to the number of ROS-producing cells. In fact, both groups exhibited a similar low proportion of ROS production (10.3 +/- 7% in normals vs. 6.8 +/- 5% in IASA). Taken together, our results show that mitochondria are clearly implicated in the apoptotic process in IASA patients. Whether this is a result of an intramitochondrial defect (e.g. Fe accumulation, secondary to mitochondrial or nuclear DNA mutations) or is secondary to an extracellular stimulus [e.g. tumour necrosis factor (TNF), Fas ligand (FasL)] remains to be determined. PMID- 11122147 TI - Abnormalities of adherent layers grown from bone marrow of patients with myelodysplasia. AB - Myelodysplastic syndromes (MDS) are characterized by a clonal disorder of haemopoiesis with defective growth in vitro. The long-term culture system was used to examine aspects of stromal function in MDS patients. Primary long-term cultures of MDS bone marrow showed poor myelopoiesis with progenitors being detected for a median 3.5 weeks (n = 12) compared with 18 weeks in cultures of normal marrow (n = 10; P < 0.0001). The haemopoietic function of adherent layers was assessed in secondary co-cultures seeded with 5 x 10(6) cord blood mononuclear cells on irradiated normal (n = 27; aged 38-82 years) or MDS (n = 32; aged 41-86 years) adherent layers (> 60% confluent). The median myeloid progenitor number/cord blood co-culture was 135 in 5-week-old cultures with normal adherent layers and 22 in those with MDS layers (P < 0.0001). Myeloid colonies were detectable for a median 11 weeks with normal adherent layers and 6 weeks with MDS adherent layers (P < 0.0001); erythroid colonies were detectable for 7 weeks (normal) compared with 5 weeks (MDS) (P < 0.01). The differences in granulocyte-macrophage colony forming unit (CFU-GM) generation were not related to patient age. Cells from adherent layers of at least half of the primary normal (n = 48) and MDS (n = 26) long-term cultures expressed cytokines [interleukin (IL)-3, IL-1 beta, thrombopoietin (Tpo) and erythropoietin (Epo)] and receptors for retinoic acid (RAR alpha) [IL-2, IL-3, macrophage colony stimulating factor (M-CSF) (Fms) and Tpo (Mpl)]. Only IL-1 beta expression was reduced in week-5 MDS cultures compared with those from normal marrows (P < 0.05). There was also a highly significant decline in IL-1 beta expression in normal (but not MDS) adherent layers between week 5 and week 10. Thus, the adherent layers in cultures grown from MDS patients were haemopoietically defective and showed abnormal IL-1 beta expression. PMID- 11122149 TI - Oncogene mutation and prognosis in the myelodysplastic syndromes. AB - In a series of myelodysplastic syndrome patients, mutational status (particularly RAS mutation) was found to be prognostic for survival, independently of four previously reported scoring systems (Bournemouth, Sanz, Lille, International). PMID- 11122148 TI - Neutrophil-specific reduction in the expression of granulocyte--macrophage colony stimulating factor receptor subunits in myelodysplastic syndromes. AB - The proliferative and differentiative response of neutrophils to granulocyte macrophage colony-stimulating factor (GM-CSF) is known to be impaired in patients with myelodysplastic syndromes (MDS). To investigate the mechanisms of the defective response in MDS, we examined expression levels of GM-CSF receptor alpha (GMR alpha) and common beta (beta c) subunits on CD16(+) neutrophils, CD14(+) monocytes and CD3(+) T cells from 26 MDS patients and 10 healthy controls using flow cytometry. Expression of GMR alpha was significantly decreased on the neutrophils of five out of 26 patients and was not specific for any FAB subtype. In contrast, beta c expression on neutrophils was significantly reduced in 14 out of 26 patients with a higher proportion occurring in the advanced stages of MDS including refractory anaemia with excess of blasts (RAEB), RAEB in transformation (RAEBt) and overt leukaemia compared with refractory anaemia (RA)/RA with ringed sideroblasts (RARS) or healthy controls. Decreased beta c also correlated with the degree of hypogranular neutrophil morphology and increased infection. Expression of both subunits on T cells and monocytes in MDS was similar to normal controls. Polymerase chain reaction amplification of reverse-transcribed mRNA isolated from the affected neutrophils suggests that the reduction of beta c may result from decreased message levels. The observed reduction in GM-CSF receptor expression could account for the impaired proliferative and maturational responses in MDS. PMID- 11122150 TI - Vincristine-induced apoptosis in vivo in peripheral blood mononuclear cells of children with acute lymphoblastic leukaemia (ALL). AB - We conducted a study to demonstrate vincristine-induced apoptosis in vivo in peripheral blood mononuclear cells of children with newly diagnosed acute lymphoblastic leukaemia (ALL). In five children, apoptosis was detected by terminal deoxynucleotide transferase-mediated dUTP-digoxigenin nick-end labelling (TUNEL) assay in blood samples collected during an up-front window study of vincristine monotherapy. In one patient, we found a striking increase of apoptotic cells from 2% (SEM 0.6) before to 40.7% (SEM 2.9) 3 h after vincristine injection. In the other patients, the maximum increase ranged from 0.8% to 4.3%. Wilcoxon matched pairs analysis of all patients confirmed that vincristine induces apoptosis in vivo in peripheral blood mononuclear cells in childhood ALL (P = 0.043). PMID- 11122151 TI - Oxygen tension influences the differentiation, maturation and apoptosis of human megakaryocytes. AB - Megakaryocytes (Mks) mature adjacent to bone marrow (BM) sinus walls and subsequently release platelets within the sinusoidal space or in lung capillaries. As the sites for platelet release have higher levels of oxygen tension (pO(2)) than the core of the BM where stem and progenitor cells reside, we investigated whether pO(2) influences Mk maturation. Mks were generated from CD34(+) cells (from mobilized peripheral blood from cancer patients) under 5% and 20% O(2). At day 15, CD41(+) Mk expansion in 20% and 5% O(2) cultures was 85-fold and 31-fold respectively. Twenty percent O(2) cultures also had higher levels of high ploidy (> or = 8N, eightfold higher) and proplatelet-forming (fivefold higher) Mks. At day 21, 20% O(2) cultures had a fivefold higher number of apoptotic Mks. In contrast, 5% O(2) promoted Mk colony-forming unit (CFU-Mk) generation and maintenance. Similar results were observed in cultures initiated with CD41(+) Mks, indicating that pO(2) directly affects Mks. The change from 20% to 5% O(2) on day 5 and day 7 delayed both maturation and apoptosis, suggesting that these two processes are closely linked. These results were confirmed in CD34(+) cultures from normal BM samples. These data may provide insights into in vivo Mk maturation, such as an explanation for hypoxia-induced thrombocytopenia in animals. PMID- 11122152 TI - Generation of functionally mature dendritic cells from the multipotential stem cell line FDCP-mix. AB - Dendritic cells (DCs) are crucial components of the immune system because of their unique ability as antigen-presenting cells for the initiation of a primary immune response. DCs, macrophages (Ms) and granulocytes (Gs) are believed to originate from a common myeloid progenitor cell. However, little is known about the molecular mechanisms leading to DC sublineage commitment. To establish a cell system that allows the molecular and biochemical analysis of DC differentiation and activation, we used the murine non-leukaemic, multipotential stem cell line FDCP-mix. FDCP-mix cells were cultured in various amounts of GM-colony stimulating factor (CSF) and interleukin (IL)-4 for up to 16 d and analysed for morphology, expression of CD34, c-kit, Gr-1, Mac-1, CD40, MHC-I, MHC-II and co stimulatory molecules (CD80, CD86) using flow cytometry, and for their capacity to present foreign antigen to autologous T cells. Up to d 7, the majority of FDCP mix cells consisted of cells differentiating along the G and M lineage. Thereafter, the number of dendritic cells increased until d 13. Differentiation along the DC lineage vs. the G and M lineage was favoured when FDCP-mix cells were cultured in high concentration GM-CSF (500 U/ml) throughout the culture and IL-4 from d 9 onwards. The dendritic cells generated from FDCP-mix cells were large, non-adherent cells with veiled processes and expressed MHC II, CD40, CD80 and CD86. After pulsing with a foreign antigen (keyhole limpet haemocyanin), FDCP mix-derived dendritic cells stimulated [(3)H]-thymidine incorporation of naive T cells in an autologous mixed lymphocyte reaction (MLR). Our results show that functionally mature dendritic cells are generated from the multipotential stem cell line FDCP-mix. This cell line thus provides the unique possibility of establishing multipotential transgenic cell lines capable of differentiation along the DC lineage. The experimental system described here should prove a valuable tool for studying DC differentiation and function. PMID- 11122153 TI - Improving the diagnosis of coeliac disease in anaemic women. AB - The majority of patients with newly diagnosed coeliac disease have iron deficiency anaemia. Occult gluten enteropathy is common. We looked at blood donor volunteers, unable to donate because they were unexpectedly found to be anaemic, to determine the incidence of coeliac disease and whether this diagnosis is routinely considered. In a 4 month period, 110,973 blood donor volunteers were seen and 1% (1197 women and 183 men) were found to be anaemic. Of 483 anaemic samples selected for testing, 32 out of 483 were positive for IgA anti-endomysial antibodies. Microcytic anaemia was found in all but three of the 32 seropositive cases. Twenty-five out of 32 volunteers agreed to have endoscopic small bowel biopsies and 22 out of 25 (88%) had the typical histological appearances of coeliac disease. Twenty-one out of 22 cases were women. In no case prior to our intervention had these women been investigated for the possibility of coeliac disease. By selecting anaemic subjects for screening, there was an improved detection rate (over 6%) compared with non-anaemic volunteers (0%). There were far more anaemic women in the study population (ratio of anaemic women to anaemic men 5.5:1). We show that, especially in anaemic menstruating women, coeliac disease is unrecognized and under-investigated. PMID- 11122154 TI - Bone mineral metabolism in adults with beta-thalassaemia major and intermedia. AB - Bone disease is an important cause of morbidity in older patients with beta thalassaemia major and intermedia. We studied 27 women and 23 men with beta thalassaemia major (37) and intermedia (13) whose mean age was 32.3 +/- 9.7 years. Bone mineral density (BMD) of the lumbar spine, femoral neck and distal radius was determined by dual-energy X-ray absorbiometry (DXA). The longitudinal change in BMD over a mean of 5.6 years was determined in 19 patients. Serum 25 hydroxyvitamin D, insulin growth factor-1 (IGF-1), bone formation markers bone alkaline phosphatase, osteocalcin and the resorption marker urinary N-telopeptide cross-linked type 1 collagen (NTx) were determined. The BsmI vitamin D receptor (VDR) gene polymorphism was analysed. Reduced BMD (Z-score < -2) was present in 89%, 62% and 73% of patients in the spine, hip and radius respectively. Vitamin D deficiency was found in 62%, decreased IGF-1 in 72% and increased urinary NTx in 84% of patients. Serum IGF-1 correlated with spine and hip BMD (r = 0.4, r = 0.39, P < 0.01 respectively), and NTx correlated with the hip BMD Z-score (r = 0.35 P < 0.05). The mean annual percentage change in spine BMD was -1.36%. Patients with the VDR BB genotype had lower spine BMD than patients with the bb genotype. In conclusion, bone loss continues in adult thalassaemia patients and is associated with increased bone resorption and decreased IGF-1. The BsmI VDR gene polymorphism is associated with osteopenia in thalassaemia. PMID- 11122155 TI - Haemochromatosis in patients with beta-thalassaemia trait. AB - Severe iron overload has been reported in patients with the beta-thalassaemia trait. Studies performed before the discovery of the haemochromatosis gene (HFE) have yielded conflicting results: some suggest that iron overload might arise from the interaction of the beta-thalassaemia trait with heterozygosity for haemochromatosis, some with homozygosity for haemochromatosis and others that it was unrelated to haemochromatosis. We have studied the clinical phenotype, iron indices and HFE genotypes of 22 unrelated patients with the beta-thalassaemia trait and haemochromatosis, the inheritance of chromosome 6p and 1q haplotypes in families of non-homozygous C282Y probands and serum measures of iron status in relatives heterozygous for C282Y with or without the beta-thalassaemia trait. We demonstrate that the beta-thalassaemia trait aggravates the clinical picture of C282Y homozygotes, favouring higher rates of iron accumulation and the development of severe iron-related complications. We suggest that the coexistence of the beta-thalassaemia trait might also increase the risk of iron overload in patients with HFE genotypes at a mild risk of haemochromatosis. Our findings do not support the hypothesis that the association of the beta-thalassaemia trait with a single C282Y or H63D allele might lead to iron overload and suggest that other non-HFE-related inherited factors are present in haemochromatosis patients with incomplete HFE genotypes. PMID- 11122156 TI - Phenotypic and molecular diversity of haemoglobin H disease: a Greek experience. AB - Haemoglobin H (Hb H) disease is the severest form of alpha-thalassaemia compatible with post-natal life and occurs when alpha-thalassaemia mutations interact to reduce alpha-globin synthesis to levels approximately equivalent to the output of a single alpha-globin gene. Hb H disease has variable clinical expression, mainly related to underlying genotypes. The spectrum of alpha thalassaemia determinants in Greece appears greater than in any other population studied and, in 75 Greek Hb H disease patients, we found 12 alpha-thalassaemia mutations interacting to produce 15 Hb H disease genotypes. Evaluation of haematological, biochemical and clinical findings, and correlation with genotypes, defined genetic predictors of disease severity and factors involved in disease progression. In accordance with previous reports, patients with non deletion alpha-thalassaemia mutations had more severe clinical expression. Additionally, we found that all patients with the most severe phenotypes had alpha-thalassaemic globin variants. Phenotypic severity was not simply related to the degree of alpha-globin deficiency: high Hb H levels were found to exacerbate anaemia by negatively influencing tissue oxygenation, and both Hb H and alpha thalassaemic haemoglobin variants appear to reduce red cell survival within the bone marrow and circulation. Together with the long-term follow-up in many patients, this report provides comprehensive information for management of Hb H disease and appropriate family counselling. PMID- 11122157 TI - Rapid flow cytometric test for the diagnosis of membrane cytoskeleton-associated haemolytic anaemia. AB - The flow cytometric test measures the fluorescence intensity of intact red cells labelled with the dye eosin-5-maleimide, which reacts covalently with Lys-430 on the first extracellular loop of band 3 protein. In this study, red cells from patients with hereditary spherocytosis (HS), congenital dyserythropoietic anaemia type II, South-east Asian ovalocytosis and cryohydrocytosis have produced a greater degree of reduction of mean channel fluorescence readings than those for other patient groups and normal controls. The predictive value of this test for membrane abnormality was compared with the results obtained from the sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) method, which is currently the reference laboratory test for the identification of membrane protein deficiencies in hereditary spherocytosis and for the detection of spectrin variants in hereditary elliptocytosis. The dye method is a reliable, speedy diagnostic test (2 h from sample collection to result) for HS with a sensitivity of 92.7% and a specificity of 99.1%. Thus, it will serve well as a first-line screening test for the diagnosis of hereditary spherocytosis in routine haematology. PMID- 11122158 TI - Activation-dependent proteolytic degradation of polymorphonuclear CD11b. AB - CD11b/CD18 is the principal integrin of polymorphonuclear (PMN) leucocytes and is involved in their adhesion, migration and phagocytosis. In quiescent cells, the receptor is stored in intracellular granules from where it is translocated to the cell surface in response to a variety of stimuli. In this study, we demonstrated that strong stimulation of PMNs not only leads to the upregulation of CD11b surface expression, but also to the subsequent time-dependent apparent loss of this receptor, as detected by fluorescence-activated cell sorting (FACS) using a monoclonal antibody (mAb) against an N-terminal CD11b epitope. This epitope loss was observed following either direct stimulation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) or after multiple receptor stimulation using a combination of the agonist N-formylmethionyl-leucyl-phenylalanine (FMLP) and the priming agents granulocyte macrophage-colony stimulating factor (GM-CSF) and platelet factor (PF) 4. However, upregulation following weak stimulation with FMLP alone was not followed by subsequent epitope loss of the receptor. The increases and subsequent decreases in CD11b expression induced by PMA were paralleled by an increase and a decrease in PMN adhesion to CD11b-specific ligands, fibrinogen and intercellular adhesion molecule (ICAM)-1. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis showed that this epitope loss of PMN CD11b was the result of proteolytic degradation of the N-terminal region of the molecule. The use of a range of proteinase inhibitors indicated that this CD11b degradation involves a cell associated serine proteinase. This is the first demonstration of the proteolytic alteration of CD11b in response to strong PMN stimulation. Given the central role of CD11b/CD18 in all aspects of PMN function, this alteration of the CD11b molecule and its effect on PMN adhesion are probably of considerable pathophysiological importance. PMID- 11122159 TI - The platelet thrombopoietin receptor number and function are markedly decreased in patients with essential thrombocythaemia. AB - Essential thrombocythaemia (ET) is a relatively common myeloproliferative disorder characterized by an elevated platelet count. As thrombopoietin (TPO) and the TPO receptor (c-mpl) regulate platelet production in normal physiology, their role in ET was investigated. A well-characterized cohort of 23 ET patients was evaluated and followed for 3 years. The TPO levels in these ET patients (189 +/- 131 pg/ml) were the same as in normal subjects (179 +/- 112 pg/ml) and TPO was not produced by ET platelets. There were 5.6 +/- 5.5 TPO binding sites/ET platelet vs. 56 +/- 17 TPO binding sites/normal platelet and this was associated in ET patients with normal-sized platelet c-mpl protein and mRNA, but a 10-fold reduction in platelet c-mpl mRNA. The K(d) for the TPO receptor on ET platelets was 66 +/- 30 pmol/l vs. 163 +/- 31 pmol/l on normal platelets, but the c-mpl cDNA had a normal nucleic acid sequence. The decreased number of ET platelet TPO receptors resulted in a fourfold decrease in the platelet-dependent TPO clearance (0.30 +/- 0.14 ml/h/10(9) ET platelets vs. 1.24 +/- 0.38 ml/h/10(9) normal platelets) at a time when the platelet count in ET patients was 2.7-fold above normal. The fourfold decrease in the TPO clearance, elevated platelet mass and resulting normal total TPO clearance explain the normal TPO levels. These results also suggest that the thrombocytosis in ET may be attributed to an alteration of the normal feedback interaction between TPO and its receptor and not as a result of any defect in the structure of TPO or c-mpl. PMID- 11122160 TI - Combined defect in membrane expression and activation of platelet GPIIb--IIIa complex without primary sequence abnormalities in myeloproliferative disease. AB - Defects in glycoprotein (GP)IIb-IIIa or in its activation may cause abnormal platelet aggregation and a bleeding diathesis. We report studies in a 67-year-old man with a myeloproliferative disease and markedly abnormal platelet responses. By flow cytometry, platelet binding of two complex-specific anti-GPIIb-IIIa monoclonal antibodies (mAbs), A2A9 and 10E5, was approximately 50% of normal. An enzyme-linked immunosorbent assay (ELISA) using immobilized kistrin showed 18% of normal membrane GPIIb-IIIa complex. By immunoblot analysis, GPIIb and GPIIIa levels in platelet lysates and membranes were near normal. Activation of GPIIb IIIa, monitored with mAb PAC-1, was markedly decreased (< 20% of normal) in response to ADP, thrombin and platelet-activating factor (PAF); expression of ligand-induced binding sites (LIBS) was < or = 30% of normal. Signal transduction independent LIBS expression, induced by echistatin, was approximately 60% of normal, suggesting that the integrin present had intact ligand-binding capability. Sequence analysis of GPIIb and GPIIIa cDNA, and platelet mRNA levels for both subunits, were normal. These findings document an acquired combined defect in membrane expression (secondary to a defect in post-translational processing of the complex) and inside-out signalling-dependent activation of the GPIIb-IIIa complex. PMID- 11122161 TI - A 1063G-->A mutation in exon 12 of glycoprotein (GP)IIb associated with a thrombasthenic phenotype: mutation analysis of [324E]GPIIb. AB - We report the molecular, genetic and functional analysis of a case of thrombasthenic phenotype. The proband showed absence of platelet glycoprotein (GP)IIb and very low content of GPIIIa, and both his parents showed a marked reduction in the levels of platelet GPIIb-IIIa. Single-stranded conformational polymorphism-polymerase chain reaction (SSCP-PCR) analysis and direct sequencing of PCR-amplified GPIIb exon-12 revealed the presence of a G-->A transition at position 1063 with the expected substitution of glutamate 324 with lysine (K). This mutation did not alter the level of GPIIb mRNA. Co-expression of normal or mutant [324K] GPIIb with normal human GPIIIa in Chinese hamster ovary (CHO) cells failed to show surface exposure of [324K]GPIIb-IIIa complexes. Pulse-chase and immunoprecipitation analysis demonstrated that [324K]GPIIb cDNA was translated into proGPIIb, but neither mutant GPIIb heavy chain (GPIIbH) nor [324K]GPIIb GPIIIa complexes were detected, suggesting that this mutation is the underlying molecular basis for the thrombasthenic phenotype. Mutation analysis demonstrated that 324E of GPIIb could be replaced by other negatively charged or polar amino acids (AAs) without impairing the surface expression of GPIIb-IIIa. However, substitution of 324E of GPIIb for a positively charged AA other than K prevented the expression of GPIIb-IIIa complexes. These observations suggest that a domain encompassing 324E of GPIIb is essential for heterodimerization with GPIIIa and its substitution for a positively charged residue precludes normal subunit association. PMID- 11122162 TI - Recombinant VIIa concentrate in the management of bleeding following prothrombin complex concentrate-related myocardial infarction in patients with haemophilia and inhibitors. AB - Prothrombin complex concentrates (PCCs) and, more recently, activated prothrombin complex concentrates (APCCs), are widely used for the treatment of active bleeding in haemophiliacs with inhibitors. Myocardial infarction (MI), associated with the use of these concentrates, is a well-recognized, but uncommon, complication. We review the 14 previous cases published in the literature and describe two additional patients. MI related to the use of activated and non activated PCCs predominantly affects young patients who often have no preceding history of, or risk factors for, MI and tends to be associated with large cumulative doses of concentrate. The most frequent pathological finding is myocardial haemorrhage, with no evidence of coronary artery atheroma or thrombosis. The management of further bleeding in these patients is difficult. We have safely used recombinant factor VIIa to treat bleeding in the immediate and long-term period following PCC-related MI. PMID- 11122163 TI - Is it necessary to administer anti-D to prevent RhD immunization after the transfusion of RhD-positive platelet concentrates? AB - Serology for the presence of anti-D after RhD-incompatible platelet transfusions was performed in 24 RhD-negative patients with haematological disease and 59 RhD negative patients with non-haematological disease. None of the patients were given prophylaxis with anti-D to prevent RhD immunization. Eight out of 59 (13.5%) non-haematology patients developed detectable anti-D, whereas 0 out of 24 (0%) of the haematology patients formed anti-D (P = 0.06). The risk of alloimmunization after RhD-incompatible platelet transfusions using platelet concentrates prepared by modern technical methods appears to be small in patients with haematological disease, but is significant in non-immunocompromised patients. PMID- 11122164 TI - Bone marrow aplasia and meropenem in a paediatric patient. PMID- 11122165 TI - Thalidomide treatment in advanced refractory myeloma. PMID- 11122167 TI - Monocyte counts: an early index of haemopoietic reconstitution after peripheral blood stem cell transplantation. PMID- 11122166 TI - The effect of virus inactivation on coagulation factors in therapeutic plasma. PMID- 11122168 TI - Formation of the Northern UK Leukaemia Consortium. PMID- 11122169 TI - Non-intervention and palliative care in vascular patients. PMID- 11122170 TI - Dealing with the rejected article. PMID- 11122171 TI - Research misconduct: diagnosis, treatment and prevention. PMID- 11122172 TI - Epilogue: key considerations in surgical publishing. PMID- 11122173 TI - British journal of surgery research bursaries 2000 PMID- 11122174 TI - Liver fracture and bleeding. PMID- 11122175 TI - Neosphincters in the management of faecal incontinence. AB - BACKGROUND: Surgical treatment of end-stage faecal incontinence has its origin in the early 1950s. Interest has been revived as a result of technical advances achieved in the recent past. The purpose of this article is to review the principles that underlie the use of skeletal muscle transposition around the anal canal and of electrical stimulation in the treatment of incontinence, and to explore new methods of treatment of this condition. METHODS: A literature search was performed using Pubmed and Medline, employing keywords related to treatment of faecal incontinence by neosphincter reconstruction. Basic science and clinical aspects of neosphincter reconstruction were gathered from relevant texts, original articles and recently published abstracts. RESULTS: The electrically stimulated gracilis neoanal sphincter seems to be the popular choice of biological neosphincter. It is more likely to produce higher resting anal canal pressures than the unstimulated neosphincter, and hence improved continence. However, electrostimulator failure may result in explantation in a proportion of patients. Impairment of evacuation is a functional setback in approximately one third of patients with the gracilis neosphincter. Overall, improvement of continence may be expected in up to 90 per cent of patients according to some reports. By contrast, experience with the artificial neosphincter, which is less expensive, has been limited to a few tertiary centres across the world. Reported continence of stool is 100 per cent, and that of gas and stool 50 per cent, following implantation of the artificial sphincter. Both of the above operations have been associated with implant-related infection and impaired evacuation. CONCLUSION: Neoanal sphincter operations are technically demanding, require a considerable learning experience and should be confined to specialist colorectal centres. Patients are likely to benefit from a plan that incorporates preoperative counselling and a selective approach. PMID- 11122176 TI - Biological behaviour and clinical implications of micrometastases. AB - BACKGROUND: The most important prognostic determinant in cancer is the identification of disseminated tumour burden (metastases). Micrometastases are microscopic (smaller than 2 mm) deposits of malignant cells that are segregated spatially from the primary tumour and depend on neovascular formation (angiogenesis) to propagate. METHODS: The electronic literature (1966 to present) on micrometastases and their implications in malignant melanoma and epithelial cancers was reviewed. RESULTS: Immunohistochemical techniques combined with serial sectioning offer the best accuracy for detection of nodal micrometastases. Molecular techniques should be reserved for blood samples or bone marrow aspirates. Detection of micrometastases in regional lymph nodes and/or bone marrow confers a poor prognosis in epithelial cancers. The concept of sentinel node biopsy combined with serial sectioning and dedicated screening for micrometastases may improve staging procedures. Strategies against angiogenesis may provide novel therapies to induce and maintain micrometastatic dormancy. CONCLUSION: The concept of micrometastases has resulted in a paradigm shift in the staging of epithelial tumours and our overall understanding of malignant processes. PMID- 11122177 TI - Role of surgery in mild primary hyperparathyroidism in the elderly. AB - BACKGROUND: The appropriate management of elderly patients with mild hyperparathyroidism is the subject of much debate. METHODS: A Medline literature search was conducted using the keywords 'hyperparathyroidism', 'asymptomatic' and 'elderly'. The references of the primary sources were examined for further citations. Personally collected cullings from journals and abstracts from journals were used as an additional source of data. RESULTS AND CONCLUSION: No prospective randomized controlled trial comparing parathyroidectomy with conservative management for patients with mild or asymptomatic hyperparathyroidism has been published. There are a number of longitudinal studies that attempt to characterize the natural history of hyperparathyroidism, but most do not stratify patients according to age when assessing outcome. None the less, it is clear that elderly patients present with a different spectrum of problems, particularly indistinct neuropsychiatric and musculoskeletal symptoms, and these are likely to be improved by surgery. The evidence allows the conclusion that the truly asymptomatic elderly patient can be successfully managed conservatively; there is a good prospect of benefit from surgery for those with symptomatic disease, and such patients should not be denied an operative option. PMID- 11122178 TI - Endoscopic ultrasonography in the evaluation of idiopathic acute pancreatitis. AB - BACKGROUND: 'Idiopathic' pancreatitis may be diagnosed when gallstones are excluded by transabdominal ultrasonography and less common causes are not implicated by history or other investigations. Transabdominal ultrasonography may not, however, detect small stones responsible for acute pancreatitis. The aim of this study was to determine if endoscopic ultrasonography (EUS) is able to identify undetected gallstones in cases of 'idiopathic' pancreatitis. METHODS: Consecutive patients presenting with 'idiopathic' pancreatitis were assessed using EUS for the presence of gallstones or other potential causes of the attack. A control group was also imaged. RESULTS: Forty-four patients with 'idiopathic' pancreatitis were assessed. Ten had suffered earlier attacks of pancreatitis before this study. EUS revealed proven pathology in 18 patients. Unconfirmed pathology was evident in 14. No abnormality was seen in only nine patients. EUS failed in one patient and there were two possible false-positive results. CONCLUSION: EUS is able to identify significant pathology in patients in whom a diagnosis of 'idiopathic' pancreatitis has been made following standard investigations. Patients with untreated gallstones are at risk of recurrent attacks. Idiopathic pancreatitis should not be diagnosed unless EUS has been performed. PMID- 11122179 TI - Biliobiliary fistula associated with gallbladder carcinoma. PMID- 11122180 TI - Epidemiology of cholecystectomy and irritable bowel syndrome in a UK population. AB - BACKGROUND: Some patients with irritable bowel syndrome (IBS) may undergo unnecessary cholecystectomy. This paper describes the prevalence of cholecystectomy and IBS in a sample of British adults in Teesside. Associations between the two conditions and their relationship to consultation behaviour and socioeconomic status are described. The results are compared with those from Bristol in an attempt to determine the influence of service-related factors on the prevalence of cholecystectomy. METHODS: A postal questionnaire was sent to 4432 adults aged 20-69 years registered with six general practices. The Standard Occupational Classification was used as a proxy for socioeconomic status. RESULTS: In Teesside cholecystectomy was reported by 4.1 per cent of women and 1.3 per cent of men. Some 22.9 per cent of the women had IBS, and 10.5 per cent of men. Cholecystectomy was more common in patients with IBS (odds ratio 1.9 (95 per cent confidence interval 1.2-3.2); P < 0.01). The prevalence of cholecystectomy, of IBS and of consultation for symptoms of IBS was not influenced by socioeconomic status. CONCLUSION: Symptoms of IBS may cause diagnostic confusion and unproductive surgery. Cholecystectomy may cause IBS-like symptoms, a single underlying disorder may produce symptoms in both gastrointestinal and biliary tracts, or the associations might be due to a combination of these factors. PMID- 11122181 TI - Appendicectomy and perioperative mortality in patients with liver cirrhosis. PMID- 11122182 TI - Need for secondary interventions after endovascular repair of abdominal aortic aneurysms. Intermediate-term follow-up results of a European collaborative registry (EUROSTAR). AB - BACKGROUND: The frequency of secondary interventions after endovascular repair of abdominal aortic aneurysms (AAAs) was assessed and correlated with findings at clinical and imaging examination during follow-up. METHODS: Data were studied from 1023 patients with a follow-up of 12 months or longer, collected by 56 institutions in a multicentre data registry (EUROSTAR). Surveillance data were provided by the centres between September 1996 and November 1999. RESULTS: Overall, 186 patients (18 per cent) had a secondary intervention occurring a mean of 14 months after the initial endograft procedure. Twelve per cent of the interventions were transabdominal, 11 per cent consisted of an extra-anatomic bypass and 76 per cent involved a transfemoral procedure. The rates of freedom from intervention at 1, 3 and 4 years were 89, 67 and 62 per cent respectively. Migration (relative risk (RR) 8.9) and rupture (RR 22.6) were the most frequent causes of secondary transabdominal interventions. Graft limb thrombosis was the principal indication for extra-anatomic bypass (RR 37.5 for clinical evidence of graft limb thrombosis). Endoleak, graft kinking, stenosis or thrombosis and device migration were significant causes for secondary transfemoral interventions (RR 2.5-6.9). CONCLUSION: The high incidence of late secondary interventions is a cause for concern with regard to broad application of endovascular AAA repair, and emphasizes the need for lifelong surveillance. PMID- 11122183 TI - Gender differences in the longitudinal pressure profile of the anal canal related to anatomical structure as demonstrated on three-dimensional anal endosonography. AB - BACKGROUND: Anal canal squeeze pressure is assumed to be due to external sphincter contraction, but the contribution of other muscles has not been explored. METHODS: Ten male and ten nulliparous female asymptomatic subjects had three-dimensional anal endosonography and manometry. Incremental squeeze pressures at 0. 5-cm intervals, expressed as a percentage of the maximum pressure recorded anywhere in the canal, were related to the following anatomical levels: puborectalis, overlap between external anal sphincter (EAS) and puborectalis, external and internal anal sphincters, and external anal sphincter only. Levels were determined by coronal and sagittal endosonographic reconstructions. RESULTS: Puborectalis was the same length in men and women (median 23.9 versus 27.1 mm) but represented a greater proportion of the anal canal in women (45 versus 61 per cent; P = 0.02). At the level of puborectalis alone, the pressure generated as a proportion of maximum anal canal pressure was 71 (range 32-100) per cent in men and 62 (range 32-100) per cent in women. At the level of the EAS alone, the pressure was 60 (4-98) per cent in men and 82 (41-100) per cent in women; where the external sphincter was overlapped by puborectalis, the pressure was 98 (60 100) per cent in men and 75 (47-100) per cent in women. CONCLUSION: Maximal anal canal squeeze pressure is found where the puborectalis overlaps the EAS. This segment represents a significant proportion of anal canal length in women. PMID- 11122184 TI - Endorectal flap advancement repair and fistulectomy for high trans-sphincteric and suprasphincteric fistulas. AB - BACKGROUND: Low-lying trans-sphincteric anal fistulas respond well to simple fistulectomy or fistulotomy. However, management of high fistulas has long been a serious problem because of the necessity of preserving at least some of the sphincter mechanism. The clinical results of endorectal flap advancement and fistulectomy for complex anal fistulas were assessed. METHODS: A total of 103 consecutive patients with high trans-sphincteric (n = 91) and suprasphincteric (n = 12) fistulas undergoing endorectal advancement flap repair together with core fistulectomy were included in a prospective study. Clinical outcome was assessed in terms of continence and recurrence by an independent observer for a period of 1 year after operation. RESULTS: Successful healing was achieved in 96 patients (93 per cent). Recurrent fistula occurred in six (7 per cent) of the 91 patients in the trans-sphincteric group and in one of the 12 patients in the suprasphincteric group. Continence disturbance was noted in eight patients (8 per cent). Previous repair and the level of the fistula did not adversely affect the results obtained. CONCLUSION: Core fistulectomy associated with endorectal advancement flap repair is a safe and effective technique for any high trans sphincteric and suprasphincteric fistula, with good results in terms of recurrence and anal continence. PMID- 11122185 TI - Mitogenic effects of oestrogen mediated by a non-genomic receptor in human colon. AB - BACKGROUND: Oestrogens are important mitogens in epithelial cancers, particularly where tumours express complementary receptors. While the traditional model of oestrogen action involves gene-directed (genomic) protein synthesis, it has been established that more rapid, non-genomic steroid hormone actions exist. This study investigated the hypothesis that oestrogen rapidly alters cell membrane activity, intracellular pH and nuclear kinetics in a mitogenic fashion. METHODS: Crypts isolated from human distal colon and colorectal cancer cell lines were used as robust models. DNA replication and intracellular pH were measured by radiolabelled thymidine incorporation (12 h) and spectrofluorescence imaging respectively. Genomic protein synthesis, sodium-hydrogen exchanger (NHE) and protein kinase C (PKC) activity were inhibited with cycloheximide, ethylisopropylamiloride and chelerythrine chloride respectively. RESULTS: Oestrogen induced a rapid (less than 5 min) cellular alkalinization of crypts and cancer cells that was sensitive to NHE blockade (P < 0.01) or PKC inhibition (P < 0.01). Oestrogen increased thymidine incorporation by 44 per cent in crypts and by up to 38 per cent in cancer cells (P < 0.01), and this was similarly reduced by inhibiting the NHE (P < 0.01) or PKC (P < 0.05). CONCLUSION: Oestrogen rapidly activates cell membrane and nuclear kinetics by a non-genomic mechanism mediated by PKC but not gene-directed protein synthesis. PMID- 11122186 TI - Hyaluronic acid as prognostic marker in resectable colorectal cancer. AB - BACKGROUND: Hyaluronic acid (HA), an extracellular high molecular mass polysaccharide, is thought to be involved in the growth and progression of malignant tumours. The objective of this work was to evaluate the cytosolic HA content in resectable colorectal cancer, its possible relationship with clinicopathological parameters of tumours and its prognostic significance. METHODS: Cytosolic HA levels were examined by radiometric assay in 120 patients with resectable colorectal cancer. The mean follow-up period was 33.4 months. RESULTS: Cytosolic HA levels of tumours ranged widely, from 30 to 29 412 ng per mg protein. Intratumour HA levels were significantly correlated with Dukes stage (P < 0.005), and were higher in patients with advanced tumours (mean(s.e.m.) 2695(446), 2858(293) and 5274(967) ng per mg protein for stages A, B and C respectively). In addition, Cox multivariate analysis demonstrated that tumour HA levels higher than 2000 ng per mg protein predicted shorter relapse-free survival and overall survival periods (both P < 0.05). CONCLUSION: There is a wide variability in cytosolic HA levels in colorectal carcinomas, which seems to be related to the biological heterogeneity of these tumours. In addition, high tumour cytosolic HA levels were associated with an unfavourable outcome in patients with resectable colorectal cancer. HA may provide additional information to that given by other biochemical markers currently used in colorectal cancer. PMID- 11122187 TI - Risk factors for surgery and recurrence in 907 patients with primary ileocaecal Crohn's disease. AB - BACKGROUND: Previous studies on risk factors for resection and postoperative recurrence in Crohn's disease have given inconclusive results. The aim of this study was to assess the risk for resection and postoperative recurrence in the treatment of ileocaecal Crohn's disease and to define factors affecting the course of the disease. METHODS: A population-based cohort of 907 patients with primary ileocaecal Crohn's disease was reviewed retrospectively. RESULTS: Resection rates were 61, 77 and 83 per cent at 1, 5 and 10 years respectively after the diagnosis. Relapse rates were 28 and 36 per cent 5 and 10 years after the first resection. A younger age at diagnosis resulted in a low resection rate. The presence of perianal Crohn's disease and long resection segments increased the incidence of recurrence, and resection for a palpable mass and/or abscess decreased the recurrence rate. A decrease in recurrence rate during the study period (1955-1989) was observed. CONCLUSION: In ileocaecal Crohn's disease the probability of resection is high and the risk of recurrence moderate. Crohn's disease in childhood carries a lower risk of primary resection. Perianal disease and extensive ileal resection increase the risk of recurrence. PMID- 11122188 TI - Risk factors related to operative mortality and morbidity in patients undergoing emergency gastrectomy. AB - BACKGROUND: Emergency gastric resection for complicated peptic ulcer and gastric cancer is a major challenge for general surgeons. This study aimed to evaluate the results of emergency gastrectomy and to examine the factors that predict the operative outcome. METHODS: A total of 82 consecutive patients who underwent emergency gastrectomy were studied. The following variables were assessed: pathology, mortality rate, morbidity, reasons for reoperation and factors related to the outcome. RESULTS: There were 64 men and 18 women with a median age of 62 (range 30-90) years. The indications were bleeding and perforated gastric or duodenal ulcers in 45 and 20 patients respectively, and bleeding and perforated gastric tumours in seven and ten patients respectively. The overall mortality rate was 17 per cent (n = 14). The complication rate was 63 per cent and 11 patients (13 per cent) required reoperation. By multivariate analysis, age greater than 65 years and blood haemoglobin level less than 10 g/dl on admission were predictive of complications after emergency gastrectomy. Postoperative pulmonary and cardiac complications and hypotension on admission were independent risk factors associated with operative death. CONCLUSION: Age more than 65 years, haemoglobin level less than 10 g/dl and hypotension on admission were associated with a poor outcome after emergency gastrectomy. The operative result was not affected by the underlying gastric pathology. PMID- 11122190 TI - Clinicopathological study of multiple superficial oesophageal carcinoma. AB - BACKGROUND: Recently, the diagnosis of superficial oesophageal carcinoma has increased markedly in Japan as a result of advances in endoscopy. A number of these carcinomas have proved to be multiple. METHODS: Some 359 patients with superficial squamous cell carcinoma of the oesophagus who underwent oesophagectomy (n = 276) or endoscopic mucosal resection (EMR) (n = 83) were reviewed retrospectively. The clinicopathological features of patients with multiple superficial oesophageal carcinoma were compared with those of patients with a single superficial oesophageal carcinoma. RESULTS: Of the 359 patients, 99 (28 per cent) had multiple superficial oesophageal carcinoma. The male : female ratio in patients with multiple carcinoma was 98 : 1, compared with 5.3 : 1 for those with a single carcinoma (n = 260) (P = 0.0001). The incidence of tobacco and alcohol use was significantly higher in the patients with multiple carcinoma than in those with a single carcinoma (P = 0.04 and P = 0.03 respectively). The incidence of pharyngeal malignancy was also significantly higher in patients with multiple carcinoma (P = 0.02). CONCLUSION: The high incidence of multiple superficial oesophageal carcinoma indicates a need for careful evaluation of the oesophagus at the time of initial diagnosis, treatment and follow-up for superficial oesophageal carcinoma. Male sex, smoking, alcohol use and the presence of pharnygeal malignancy are high-risk factors for multiple superficial oesophageal carcinoma. PMID- 11122189 TI - Day-case laparoscopic fundoplication for gastro-oesophageal reflux disease. AB - BACKGROUND: Based on a series of successful outpatient laparoscopic cholecystectomies, day-case laparoscopic fundoplication for gastro-oesophageal reflux disease was introduced in January 1997. The initial results are reported. METHODS: Inclusion criteria were American Society of Anesthesiologists grade I II, living within 30 min travel from the hospital, and adult company at home. Initially only selected patients were offered day-case treatment, but later it was adopted as routine. The patients underwent general intravenous anaesthesia with propofol and remifentanil, and were given ketorolac, propacetamol, droperidol and ondansetron as prophylaxis against postoperative pain and nausea. The surgical procedure was Nissen-Rosetti fundoplication or semifundoplication depending on oesophageal manometric results. RESULTS: Forty-five patients were included. Four patients were admitted; 41 were discharged as planned 3-8 h after operation, and five of these patients were readmitted. One underwent reoperation for necrosis of the gastric fundus. A further five patients visited the outpatient department without need for admission. At follow-up 31 patients were satisfied with the day-case treatment, five were indifferent, and five were dissatisfied because of pain. If offered a similar operation in the future, 26 patients would have preferred and seven would have accepted day-case treatment, and eight would not. CONCLUSION: Outpatient laparoscopic fundoplication is safe and well tolerated by the majority of patients. PMID- 11122191 TI - Quality of life in long-term survivors after curative transhiatal oesophagectomy for oesophageal carcinoma. AB - BACKGROUND: Transhiatal resection for oesophageal cancer is a major operation with potentially severe physical, emotional and social consequences. The aim of this study was to assess various aspects of quality of life in long-term survivors following oesophageal resection for cancer. METHODS: Between January 1993 and May 1996, 100 consecutive patients with cancer of the oesophagus or oesophagogastric junction underwent a potentially curative transhiatal oesophagectomy. Patients with a minimum follow-up of 2 years and with no tumour recurrence (n = 35) were mailed questionnaires which consisted of: (a) the Short Form-36 Health Survey to assess general quality of life, (b) an adapted Rotterdam Symptom Checklist to assess disease-specific quality of life, and (c) additional questions about other effects of the operation. RESULTS: All patients returned the questionnaire. General quality of life was comparable with reference values for the same age group. However, more than half of the patients still experienced at least some early satiety, fatigue, dysphagia, heartburn and/or psychological irritability. Nine of 13 patients who worked in paid employment before operation continued to do so. CONCLUSION: Patients who survive 2 years or more after transhiatal oesophageal resection for cancer can lead satisfactory lives. Although some residual symptoms may persist, their general quality of life is similar to that of healthy individuals of the same age. PMID- 11122192 TI - Groin hernia repair in Scotland. AB - BACKGROUND: The use of mesh for groin hernia repair has dramatically changed the way this common operation is performed. The aim of this study was to survey the methods of groin hernia repair in Scotland and to assess patient satisfaction with the operation. METHODS: Between 1 April 1998 and 31 March 1999 all patients who underwent groin hernia repair in the National Health Service in Scotland were identified. As well as looking at the type of hernia repair performed and postoperative morbidity, patients were sent a Short Form-36 about 3 months after the operation to assess satisfaction and return to normal activity. RESULTS: Information was obtained on 5506 (97 per cent) of patients who underwent groin hernia repair during the study period. Eighty-five per cent of patients had an open mesh repair and 4 per cent had a laparoscopic repair. Most operations (85 per cent) were performed using general anaesthesia on an inpatient basis (78 per cent), and 8 per cent were for repair of a recurrent hernia. Potentially serious intraoperative complications were rare (seven patients), although they were significantly (P < 0. 001) more likely to be associated with a laparoscopic approach or repair of a femoral hernia: relative risk compared with open inguinal hernia repair 33 (95 per cent confidence interval (c.i.) 6-197) and 22 (95 per cent c.i. 3-152) respectively. Wound complications were common and 10 per cent of patients required a district nurse to attend the wound. Patients expressed a high degree of satisfaction; 94 per cent would recommend the same operation to someone else if required. CONCLUSION: An open mesh repair using general anaesthesia has become the repair of choice for a groin hernia in Scotland. Despite a high incidence of wound complications, patients are satisfied with this operation. PMID- 11122193 TI - Future of laparoscopic inguinal hernia surgery. PMID- 11122195 TI - BJS and the blitz PMID- 11122194 TI - The Seldinger technique for difficult transurethral catheterization: a gentle alternative to suprapubic puncture. PMID- 11122196 TI - Randomized clinical trial of laparoscopic versus open fundoplication: blind evaluation of recovery and discharge period. PMID- 11122197 TI - Authors' reply PMID- 11122198 TI - Long-term results of arteriovenous fistulas using transposed autologous basilic vein. PMID- 11122200 TI - Authors' reply PMID- 11122199 TI - Slowing the heart saves lives: advantages of perioperative beta-blockade. PMID- 11122201 TI - Surgery for colorectal liver metastases with hepatic lymph node involvement: a systematic review. PMID- 11122202 TI - New concepts in effector functions of eosinophil cytokines. PMID- 11122203 TI - Functional significance of polymorphisms of the interleukin-4 and interleukin-13 receptors in allergic disease. PMID- 11122204 TI - Airway inflammation and ion channel abnormalities in cystic fibrosis. PMID- 11122205 TI - Bee venom allergy. PMID- 11122206 TI - Nasal tolerance induction as a potential means of immunotherapy for autoimmune diseases: implications for clinical medicine. PMID- 11122207 TI - Presentation at the National Asthma Campaign International Congress, June 1999. Controlling the inflammatory response through transcriptional mechanisms. PMID- 11122208 TI - Cytokine expression during allergen-induced late nasal responses: IL-4 and IL-5 mRNA is expressed early (at 6 h) predominantly by eosinophils. AB - The production of TH2-type cytokines [interleukin-4 (IL-4) and IL-5] and tissue eosinophilia are characteristic features of allergic diseases. It was previously reported that at 24 h after allergen provocation, CD3+ T-lymphocytes were the principal cell source of IL-4 and IL-5 mRNA transcripts in both atopic asthma and rhinitis. To investigate whether IL-4 and IL-5 mRNA are expressed earlier during late nasal responses and if so, which cell(s) are responsible. Nasal biopsies were obtained at 6 h after nasal allergen challenge and following a control challenge with the allergen diluent. Sections were immunostained for T lymphocytes (CD3+, CD4+) and eosinophils (EG2+). In situ hybridization was used to detect the number of cells expressing messenger RNA (mRNA) for IL-4 and IL-5. In patients with allergic rhinitis, eosinophils (EG2+ cells P = 0. 006) but not T cells (CD3+ cells) increased in the nasal mucosa at 6 h after allergen challenge. The number of cells expressing IL-4 mRNA (P = 0.01) and IL-5 mRNA (P = 0.05) also increased at 6 h. Co-localization studies showed that 76% of IL-4 mRNA+ cells and 77% of IL-5 mRNA+ cells were eosinophils, whereas at this time point, T-cells and mast cells accounted for /= 3 mm to one of eight 1 : 20 (w/v), 50% glycerinated ('No US Standard of Potency') allergens licensed by the FDA: house dust, cat, dog, Alternaria, mixed giant/short ragweed, oak, perennial rye grass, and Bermuda grass. Survival analyses were conducted using multivariate adjusted Cox regression models to evaluate the association between atopy and all-cause, cardiovascular, and cancer mortality. RESULTS: There was no association between allergen skin test reactivity and all cause mortality: 30-44 years RR = 1.07 (95% CI 0.63-1.84); 45-59 years RR = 1.10 (0.78-1.55); 60-75 years RR = 1.07 (0.91 1.25). Results were unchanged when cancer or heart disease mortality were examined separately. The presence or absence of allergic symptoms, using the flare to define skin test reactivity, eliminating deaths in the first 5 years of follow-up, or eliminating individuals with pre-existing conditions did not alter the findings. CONCLUSIONS: Atopy, defined by allergen skin test reactivity, with or without symptoms, is not a predictor of subsequent mortality. PMID- 11122210 TI - Combined effects of aerobiological pollutants, chemical pollutants and meteorological conditions on asthma admissions and A & E attendances in Derbyshire UK, 1993-96. AB - BACKGROUND: The effect of outdoor aeroallergen exposure in asthma may be enhanced by air pollutants, including ozone, nitrogen dioxide and particulates, and by certain weather conditions. It is not yet established whether these interactions are important in determining asthma morbidity at the population level. OBJECTIVE: We have investigated the joint effects of aeroallergens, rainfall, thunderstorms and outdoor air pollutants on daily asthma admissions and Accident and Emergency (A & E) attendance using routinely collected data between 1993 and 1996 from Derby in central England. METHODS: Daily counts during the aeroallergen season of grass and birch pollen, basidiospores, Didymella, Alternaria and Cladosporium, maximum 1 hour ozone and nitrogen dioxide and daily average black smoke measurements, all made in the vicinity of the city centre, were categorized in tertiles. Rainfall was classified as dry, light ( 2 mm). The modifying effect of outdoor pollutant levels, and rainfall or the occurrence of a thunderstorm, upon the effects of individual aeroallergens on asthma admissions and A & E attendance were investigated by fitting appropriate interactions in log linear autoregression models with adjustment for potential confounders. RESULTS: We found a significant interaction between the effects of grass pollen and weather conditions upon A & E attendance, such that the increase with grass pollen count was most marked on days of light rainfall (adjusted rate ratio for >/= 50 vs < 10 grains/m3 at lag 2 days = 2.1, 95% CI 1.4, 3.3). Asthma admissions increased with Cladosporium count. We found no statistically significant interactions between effects of any individual aeroallergen and outdoor air pollutant upon either measure of asthma morbidity. CONCLUSIONS: Rainfall and thunderstorms are important effect modifiers in the relation between grass pollen and measures of acute asthma morbidity. Interactions between ambient levels of aeroallergens and chemical pollutants in the Derby area do not play a major role in determining asthma admissions and A & E attendance. PMID- 11122211 TI - Personal exposure to allergenic pollen and mould spores in inland New South Wales, Australia. AB - BACKGROUND: In inland NSW, Australia, allergic sensitization to the fungi Alternaria and Cladosporium and to pollen is common and an important risk factor for asthma. OBJECTIVE: We report the results of a series of experiments designed to assess the nature of personal exposure to these airborne allergenic particles. We have tested the effect of exposure conditions and level of activity on measurements of the personal exposure. METHOD: Personal Air Samplers (PAS) and Nasal Air Samplers (NAS) were employed. NAS are fitted just inside the nose and collect inhaled particles by impaction, while the PAS use a pump-operated filter with constant air flow (2 L/min). Thirty-three subjects (adults and children) used both NAS and PAS simultaneously for four one hour periods during which they performed activities or rested, both inside and outside their homes. Samples were analysed by light microscopy. Alternaria spores, Cladosporium spores, grass pollen and nongrass pollen were counted. RESULTS: Both samplers detected substantial variation in exposure between subjects. Between members of the same household, the intrahouse correlation coefficient ranged from < 0 - 0.38. Levels of pollen grains and fungal spores inhaled were higher during periods of activity than during rest, and higher while subjects were outdoors than indoors. During the active outdoor period, the number of Alternaria spores inhaled ranged from 4 to 794 (median 11) spores/hr, Cladosporium from 0 to 396 (median 4) spores/hr, grass pollen from 0 to 81 (median 1) grains/hr and nongrass pollen from 0 to 72 (median 5) grains/hr. CONCLUSION: This is the first study to quantify individual inhaled levels of allergenic fungal spores and pollen under normal domestic circumstances. Exposure can be substantial and highly variable between individuals. The amount of particles inhaled relates both to location of the individual and activity being performed, independent of age group. PMID- 11122212 TI - Socioeconomic status is a risk factor for allergy in parents but not in their children. AB - BACKGROUND: Allergic diseases are more prevalent in affluent countries, which has been attributed to life-style factors. Life-style habits may also differ between socioeconomic (SES) classes. The objective of this paper therefore was to evaluate if SES had an impact on the development of atopic disorders. METHODS: A total of 1314 German children were followed-up in an observational birth cohort study to 6 years of age. Parents filled in questionnaires, and had multi-allergen screening tests for sensitization. Indoor allergen concentrations were determined by ELISA. Children were examined regularly up to 6 years, specific serum IgE values were determined by CAP-Rast-Feia. RESULTS: The risk of aeroallergen sensitization (odds ratio 1.76; 95% CI 1.30-2.37), and the lifetime prevalence of hay fever (2.36; 1.76-3.17), and asthma (1.74; 1.08-2.80), but not of atopic dermatitis (AD: 0.90; 0. 54-1.51) was elevated in parents of high compared to low SES. With high SES the risk of smoking in pregnancy (0.35; 0.23-0.51), in the home (0.31; 0.21-0.46), pet ownership (0.37; 0.26-0.55), high mite (0.42; 0.25 0.74), and high cat (0.38; 0.18-0.82) allergen concentration in house dust was reduced, but elevated for breastfeeding over more than 6 months (4.67; 2.9-7.48). In children, even after controlling for other risk factors, only the risk of AD from 3 to 6 years (2.42; 1.42-4.14) was elevated in families with high SES, but not of AD in infancy or of any other atopic disorder. CONCLUSIONS: While parents of high SES have a higher prevalence of inhalative allergies, their favourable life-style prevents the development of atopic disorders in their children, except for AD beyond infancy. PMID- 11122213 TI - Association between a C+33T polymorphism in the IL-4 promoter region and total serum IgE levels. AB - Susceptibility to asthma and other atopic diseases is known to be associated with elevated total IgE levels. Several investigators have linked the interleukin-4 (IL-4) gene and nearby markers located on chromosome 5q to elevated total IgE levels. A single nucleotide polymorphism in the IL-4 gene promoter region (C+33T) has recently been identified. As part of an effort to identify genetic variants contributing to the susceptibility to elevated total serum IgE levels, an association analysis of a newly identified promoter polymorphism (C+33T) with total serum IgE levels was conducted. The study was conducted using 240 Japanese subjects (120 asthmatics and 120 healthy controls). The IL-4 C+33T polymorphism was genotyped by PCR-restriction fragment length polymorphism analysis. The frequency of the T allele was 0.675 in asthmatic subjects and 0.671 in healthy controls. An ANOVA model adjusted for age, sex and disease status suggested a genetic association of C+33T polymorphism with elevated total serum IgE levels (P < 0.05). The data suggest that IL-4 promoter C+33T polymorphism may be one of the genetic polymorphisms that explain genetic linkage or association between elevated total serum IgE levels and markers on chromosome 5q. PMID- 11122214 TI - Molecular cloning of major allergen from Cupressus arizonica pollen: Cup a 1. AB - The family Cupressaceae is a relevant source of allergens that causes winter respiratory allergies. Cloning and sequencing the major antigen of Cupressus arizonica is important for a better diagnosis and treatment of sensitized patients. To obtain a full-length complementary DNA for Cup a 1, the major allergen of Cupressus arizonica pollen. It was cloned and sequenced and the recombinant protein was expressed. Messenger RNA from Cupressus arizonica pollen was obtained and the Cup a 1 sequence was established using a 3'-RACE system and primers based on the N-terminal amino acid sequence. Recombinant Cup a 1 was cloned in pBluescript and expressed in a glycosylated form in rabbit reticulocytes. The cDNA was subcloned in pGEX-5X-1 and expressed in Escherichia coli as a fusion protein with GST. Recombinant Cup a 1 is highly homologous with the major allergens of mountain cedar (Jun a 1), Japanese cypress (Cha o 1) and Japanese cedar (Cry j 1). Cup a 1 contains three potential N-glycosylation sites that are different from those found in Jun a 1 and Cry j 1. The cloned protein contains a pectate lyase active site identical to those of Cry j 1 and Jun a 1. The IgE from patients' sera recognizes recombinant Cup a 1, and this reactivity is higher with the glycosylated protein. Cup a 1 has been cloned and sequenced. As expected, the high degree of homology with Cha o 1, Jun a 1 and Cry j 1 explains the cross-reactivity of conifer pollens. Different IgE reactivity with the glycosylated and non-glycosylated protein suggests the importance of carbohydrate moieties in the IgE binding site. PMID- 11122215 TI - Uptake of the yeast Malassezia furfur and its allergenic components by human immature CD1a+ dendritic cells. AB - Atopic dermatitis (AD) is a chronic inflammatory skin disease with increasing prevalence, though still little is known of the pathomechanisms and the causes of the disease. Patients with AD often have specific IgE reactivity to the yeast Malassezia furfur (M. furfur), present in the normal microflora on human skin. To investigate the possible interaction of immature and mature antigen-presenting dendritic cells with the yeast M. furfur and its allergenic components. Monocyte derived dendritic cells (MDDCs) generated from human peripheral blood were allowed to interact with FITC-labelled whole M. furfur yeast cells, M. furfur extract, a recombinant allergen from M. furfur designated rMal f 5 and M. furfur mannan, in the absence of IgE antibodies. Interaction and uptake were detected using flow cytometry and confocal laser scanning microscopy. Internalization of M. furfur yeast cells and yeast components by immature MDDCs was found using confocal laser scanning microscopy. Results from flow cytometric studies showed that a median of 94% (range, 65-98%) of the immature CD1a+ MDDCs were M. furfur extract positive, 81% (75-97%) rMal f 5 positive and 93% (62-98%) mannan positive. Mature CD1a+ MDDCs were significantly less efficient in this respect, with the corresponding figures only 26% (6-37%, P < 0.01), 6% (2-15%, P < 0.05) and 32% (9-50%, P < 0.01), respectively. Uptake of the non-glycosylated rMal f 5 by immature CD1a+ MDDCs was decreased to 27% (15-38%) by inhibition of pinocytosis. The binding of M. furfur extract and mannan was inhibited in a dose dependent manner by methyl-alpha-D-mannopyranoside, suggesting uptake via the mannose receptor. Human immature CD1a+ MDDCs can efficiently take up M. furfur and allergenic components from the yeast in the absence of IgE antibodies, implying that sensitization of AD patients to M. furfur can be mediated by immature dendritic cells in the skin. PMID- 11122216 TI - The relationship of eosinophil granule proteins to ions in the sputum of patients with cystic fibrosis. AB - Increased sputum levels of eosinophil granule proteins have been reported despite normal eosinophil numbers in peripheral blood and in the lung in cystic fibrosis (CF). Mechanisms of eosinophil priming and activation are still unclear in CF. In the present study we investigated whether ion concentrations in the sputa of CF patients are related to eosinophil activity. We assessed concentrations of eosinophil cationic protein (ECP), eosinophil protein X (EPX), major basic protein (MBP) and ions (Na+, Cl-, Ca2+, Mg2+) in sputum samples of 29 children with CF as well as in 10 controls with bronchial asthma. Patients with CF demonstrated significantly higher levels of ECP, Na+, Cl- and Ca2+ levels than asthmatics (P < 0.04, P < 0.0001, P < 0.0001, P < 0.02). No differences were seen between concentrations of EPX and Mg2+ in the two groups. In CF, eosinophil granule proteins correlated significantly with Ca2+ and Mg2+ concentrations (ECP, P < 0.0001, r = 0.65, P < 0.0001, r = 0.66; MBP, P < 0.03, r = 0.41, P < 0.03, r = 0.42), furthermore inversely with Cl- concentrations (ECP, P < 0. 0003, r = - 0.63; EPX, P < 0.02, r = - 0.45; MBP, P < 0.03, r = - 0. 41) but not with Na+ levels. ECP, Na+ and Cl- were also correlated with lung function variables (FVC, P < 0.04, r = - 0.38, P < 0.02, r = 0.44, P < 0.03, r = 0.41; FEV1, P < 0.007, r = - 0.49, P < 0.006, r = 0.5, P < 0.008, r = 0.48; MEF50, P < 0.003, r = - 0.54, NS, P < 0.03, r = 0.42; MEF25, P < 0.039, r = - 0.4, P < 0.005, r = 0.51, P < 0.05, r = 0.37). Our results demonstrated a significant relationship of eosinophil degranulation and ions in CF, indicating that ion composition in CF sputa may be at least partly be responsible for high levels of eosinophil products despite low eosinophil numbers. PMID- 11122217 TI - Analysis of induced sputum before and after withdrawal of treatment with inhaled corticosteroids in asthmatic patients. AB - To assess whether sputum eosinophilia predicts the recurrence of asthma symptoms after withdrawal of therapy in moderate stable asthmatics on low-dose inhaled corticosteroids. Randomized, double-blind, placebo-controlled study involving 30 subjects with stable asthma, asymptomatic, with low PEF variability measured over two run-in weeks, on treatment with low-dose inhaled beclomethasone dipropionate (BDP, 250 microgram b.i.d. in the last 3 months). At the end of the run-in, all patients underwent a methacholine challenge test and sputum induction (T1). They then stopped therapy and received either placebo (20 subjects, study group) or BDP at the same dose as in the previous 3 months (10 subjects, control group). They continued to monitor PEF and symptom score for 3 months, or until asthma symptoms recurred (diurnal and nocturnal symptom score >/=2 on two consecutive days). At the end of the study (T2), i.e., either within 5 days from the beginning of asthma symptoms or after 3 months in subjects without recurrence of asthma symptoms, all subjects repeated the methacholine challenge test and sputum induction. In the placebo-treated group, sputum eosinophils at T1 were significantly higher in subjects who subsequently developed recurrence of asthma symptoms (n = 7) after cessation of treatment than in subjects who remained asymptomatic for 3 months (8.2% [0-56.6] vs 0.9% [0-11], P < 0.05). At the time of recurrence of asthma symptoms, sputum eosinophil percentages significantly increased (from 8.2% [0-56.6] to 16.6% [5.8-73.6], P < 0.05). The positive predictive value of sputum eosinophils for the recurrence of asthma symptoms was 71%, while the negative predicting value was 84%. In the BDP-treated control group, none of the subjects experienced recurrence of asthma symptoms, and sputum eosinophil percentages measured at the beginning (T1) and at the end (T2) of the study were similar. Sputum eosinophil percentages may vary over a wide range in asthmatic subjects, although regularly treated and apparently well controlled. However, high sputum eosinophil percentages are related to early recurrence of asthma symptoms after cessation of inhaled corticosteroids. PMID- 11122218 TI - A moderate and unspecific release of cysteinyl leukotrienes by aspirin from peripheral blood leucocytes precludes its value for aspirin sensitivity testing in asthma. AB - Aspirin-induced asthma (AIA) is a clinical syndrome related to cysteinyl leukotriene overproduction in airways. The confirmation of the diagnosis requires inconvenient provocation tests with acetyl salicylic acid (ASA). A study was performed to evaluate whether measurement in vitro of cysteinyl leukotrienes (cys LTs) release by isolated peripheral blood leucocytes, stimulated with ASA, can be of use for diagnosis of AIA. A cellular allergen stimulation test, CAST, was adapted to measure leukotriene release from leucocytes of 32 aspirin-tolerant and 26 aspirin-intolerant asthmatics. The cells were stimulated with Lys-ASA, N formyl-methionyl-leucyl-phenylalanine (fMLP), or both fMLP and Lys-ASA, in a buffer containing IL-3, and results compared with human leukaemia cell line (Hl 60) response to Lys-ASA. Cys-LTs were measured in cell supernatant fluids by ELISA. ASA had a rather week stimulatory effect on cys-LTs release in both groups of patients. Contrary to some previous studies, no significant differences were found between cys-LTs release by leucocytes from AIA and ATA, or by differentiated Hl-60 cells. Measurement of cysteinyl-leukotriene release by peripheral blood leucocytes pre-treated with aspirin has no value for diagnosis of AIA. PMID- 11122219 TI - Intranasal capsaicin is lacking therapeutic effect in perennial allergic rhinitis to house dust mite. A placebo-controlled study. AB - In a recent placebo-controlled study we demonstrated that capsaicin is an efficacious substance in the treatment of non-allergic non-infectious rhinitis. In this study the therapeutic effect lasted more than 9 months. This effect was not based on modulation of inflammation. To evaluate the effect of repeated application of capsaicin to patients with a nasal allergy to house dust mites (HDM), using the same treatment protocol as recently introduced in the treatment of non-allergic patients. Twenty-six patients with rhinitis, 15 females and 11 males (range: 20-46 years; mean 30.5), allergic to HDM were treated with either capsaicin or placebo in a double-blind, placebo-controlled, parallel group design. Nasal reactivity to HDM expressed as nasal symptoms, albumin and leukotriene levels in nasal lavage fluid and responsiveness to histamine, assessed as symptoms before and 6 weeks after treatment, were used to compare both treatment groups. In addition, visual analogue scales and rhinitis quality of life (RQL) assessment before, 6 weeks after and 3 months after treatment were used as outcome variables. No significant effect of capsaicin on nasal challenge tests with HDM (nasal symptoms, albumin and leukotriene levels), on VAS or RQL outcome 6 weeks or 3 month's after treatment, was demonstrated. Capsaicin did have a small effect on the area of the curve (AUC) of histamine dose response curves (P = 0.03). Desensitization with capsaicin in doses sufficient to control symptoms in patients with severe non-allergic rhinitis is lacking therapeutic effect in perennial allergic rhinitis. PMID- 11122220 TI - Nitric oxide-dependent neutrophil recruitment: role in nasal secretion. AB - Leukotriene B4 (LTB4), an inflammatory mediator, is a potent chemoattractant for neutrophils that plays an important role in nasal secretion via release of elastase. Nitric oxide (NO) is an important modulator of leucocyte-endothelial cell interactions, endogenously produced in large quantities in the paranasal sinuses. To examine the role of NO in LTB4-stimulated nasal secretion. A newly developed method for isolating and superfusing a nasal segment in dogs was used. Instillation of LTB4 into the nasal segment caused a time-dependent increase in the volume of airway fluid and in the recruitment of neutrophils. N(G)-nitro-L arginine-methylester (L-NAME), an inhibitor of NO synthase, prevented LTB4 induced neutrophil recruitment and nasal secretion. These studies show that NO modulates LTB4-induced neutrophil recruitment and subsequent fluid secretion in the nose, and they suggest a therapeutic role for NO inhibitors in modulating neutrophil-dependent nasal secretion. PMID- 11122221 TI - Interleukin-10 generation in atopic children following oral Lactobacillus rhamnosus GG. AB - Oral Lactobacillus rhamnosus GG ingestion for 5 days to 4 weeks has been shown to alleviate clinical symptoms of gastrointestinal inflammation and atopic dermatitis. To determine whether oral Lactobacillus rhamnosus GG may act by generating immunosuppressive mediator in atopic children. Lactobacillus rhamnosus GG (ATCC 53103) at a daily dose of 2 x 1010 cfu was added for 4 weeks to the diets of nine children (mean age, 21 months) with atopic dermatitis. Blood and faecal samples were collected before supplementation and at early (2 weeks) and late stage (4 and 8 weeks from the beginning). The concentrations of interleukin 6 (IL-6), IL-10, IL-12, tumour necrosis factor-alpha (TNFalpha) and interferon gamma (IFNgamma) in sera, as well as the production of IL-2, IL-4, IL-10 and IFNgamma in mitogen-induced peripheral blood mononuclear cells, were assessed. Secretory IgA and TNFalpha were also determined in faeces. The serum IL-10 concentration differed significantly between before, early and late samples (P < 0.001) due to the elevation of serum IL-10 in the later phase of oral Lactobacillus rhamnosus GG ingestion. The enhancement of IL-10 production in mitogen-induced cultures preceded the rise in serum IL-10. The enhanced IL-10 generation in vivo substantiates the anti-inflammatory properties of specific probiotic bacteria strains, and provides an additional reason for considering such treatments for patients with intestinal inflammation. PMID- 11122223 TI - Epidemiology of atopic dermatitis. AB - Although research into atopic dermatitis (AD) has been dominated by the study of cells and chemical mechanisms over the last 40 years, the last 7 years has witnessed a respectable growth within the field of AD epidemiology. Significant advances include valid disease definitions that can be used in epidemiological studies, global prevalence studies, and studies which quantify the morbidity and economic cost of the disease. These have all helped to argue the case for more research into AD. Epidemiological studies demonstrating that AD is commoner in wealthier families, linkage with small family size, increased prevalence in migrant groups, and the increasing prevalence of the disease all argue strongly towards an important role for the environment in determining disease expression. Future research gaps include evaluation of gene-environment interactions, better studies of the natural history of AD, and better clinical trials that answer questions that are important to physicians and their patients. PMID- 11122224 TI - Atopic dermatitis: pathogenetic mechanisms. AB - Atopic dermatitis (AD) is a chronic inflammatory skin disease with increasing incidence and socio-economical relevance. The diagnosis is made on clinical grounds and different diagnostic criteria sets have been established. The majority of all AD cases is associated with a sensitization to environmental allergens and increased serum IgE (so-called extrinsic AD), but about 10--30% of all cases suffer from the so-called intrinsic AD, which obviously lacks any link to the classical atopic diathesis. The genetic background of AD has been investigated by target gene approach by different groups with mostly contradictory results for each of the genes under study. An imbalance in the spectrum of Th1/Th2 responses, a disturbed prostaglandin metabolism, intrinsic defects in keratinocyte function, delayed eosinophil apoptosis, IgE-mediated facilitated antigen presentation by epidermal dendritic cells, a two phase model of the inflammatory response and staphylococcal superantigen effects are among the currently studied pathogenetical aspects of extrinsic AD, which are reviewed in this paper. PMID- 11122225 TI - Clinical aspects of atopic dermatitis. AB - Atopic dermatitis (AD), or atopic eczema, is the most common, chronic inflammatory disease among children in industrialized countries. We still lack knowledge of its pathophysiology and in particular the role of allergy as both an eliciting factor for disease expression and for disease activity. This article describes the clinical symptoms of the disease and its qualitatively different aspects. AD cannot be understood as being induced by one factor only, e.g. allergy, and this is important when planning treatment strategies. It is also important to realize the very wide range of disease intensity: from subclinical, or latent AD, in which only a few symptoms are present and thus which prevents a clear diagnosis of AD, to its most severe forms including erythroderma. It's unknown aetiology, the wide range in symptomatology, and the fluctuating course (including the many eliciting factors) form the background for our diagnostic and therapeutic difficulties of atopic eczema. AD is prevalent in childhood, but the atopic trait continues, not only for later respiratory allergies, but also for skin diseases in adulthood (such as AD itself or the frequent irritant contact dermatitis of the hands). A child with an acute and first attack of AD is therefore a challenge to the child, its parents and - certainly - to the doctor. However, after stressing the chronicity of the disease, it is equally important to assure the parents that this disease is, in most cases, controllable through correct treatment and that it has a good prognosis: It is not a 'life sentence', but a controllable disease in an otherwise healthy child. PMID- 11122226 TI - Airborne and dietary allergens in atopic eczema: a comprehensive review of diagnostic tests. AB - Aeroallergens and food allergens are relevant eliciting factors of atopic eczema. This article focuses on the methods used for diagnosis in patients with suspected allergy to airborne or dietary allergens and who are suffering from atopic eczema. In addition to classical tests of IgE-mediated hypersensitivity (intracutaneous or in vitro testing), the role of provocation procedures is described. For aeroallergens, the atopy patch test yields the most specific results with regard to clinical history as compared with classical methods. For food allergens and pseudoallergic reactions to additives, this holds true for the double-blind, placebo-controlled food challenge. The methods and their limitations are discussed from a practical point of view. PMID- 11122227 TI - Phototherapy for atopic dermatitis. AB - The beneficial effects of ultraviolet (UV) radiation on atopic dermatitis has been appreciated for many years. While broadband UVB and psoralen UVA have been the mainstay of phototherapy for some time, the past 5 years have seen the introduction of phototherapeutic modalities, including UVA-1 and 311nm UVB. The best modality and mode of usage is dependent on the type of atopic dermatitis, severity and body site. T lymphocytes play an important role in disease pathogenesis and UV radiation has profound effects on skin and systemic immune responses. PMID- 11122228 TI - Systemic therapy of atopic dermatitis. AB - Atopic dermatitis is a chronic, relapsing, inflammatory skin disease. Topical therapy is the mainstay, but patients with widespread moderate to severe atopic dermatitis may require systemic therapy. Immunosuppressants, immune response modifiers, antihistamines and antibiotics are among the classes of systemic medications frequently used to treat extensive atopic dermatitis; the indications and scientific support for the use of these and other less commonly used medications will be reviewed in this article. PMID- 11122229 TI - New approaches to topical therapy. AB - Despite the rapid and proven efficacy of topical corticosteroids, side-effects can limit their clinical usefulness. Topically active macrolide immunosuppressants such as ascomycin and tacrolimus appear to provide comparable therapeutic potency without significant local or adverse effects. Data from ongoing studies will be crucial in determining the safety of these agents in the long term, and also their place within the current therapeutic armamentarium available for patients with atopic dermatitis. Enzyme inhibitors of PLA(2) and PDE 4 currently in the very early stages of clinical development also show potential promise as additional alternative strategies to topical treatment and may perhaps act as steroid sparing agents. Having been in the therapeutic doldrums for years, topical management of atopic dermatitis is likely to show great changes in the very near future. PMID- 11122230 TI - X-linked lymphoproliferative syndrome. PMID- 11122231 TI - Partners in crime: co-infections in the developing world. PMID- 11122232 TI - Pathogenic roles of eosinophils in guinea-pig contact sensitivity: regulation of dermal eosinophilia with remotely administered IL-5. AB - Eosinophils have a variety of functions. Although increasing evidence links the presence of eosinophils to airway damage, studies have not examined in detail if, and how, eosinophils affect skin inflammation. The purpose of this study was to determine whether eosinophil infiltration augments the contact sensitivity reaction in vivo. Guinea-pigs were sensitized with 2, 4-dinitrochlorobenzene and challenged on the dorsal skin or on the right ear lobe. The number of eosinophils and macroscopic changes of the skin lesion in the presence or absence of human recombinant IL-5 (rIL-5) administered at the remote site was assessed. The reaction on the dorsal skin was acutely eczematous with considerable basophil infiltration. In contrast, eosinophils had extensively infiltrated the right ear lobe and major basic protein was deposited in the dermis. A subcutaneous injection of rIL-5 (10 pmol/kg) at the remote site (left ear lobe) 12 h after challenge induced transient blood eosinophilia and enhanced eosinophil accumulation in the challenged ear lobe. These changes were accompanied by increased ear swelling and severe erythema. In contrast, eosinophil infiltration was significantly inhibited by rIL-5 administered at the time of challenge. Ear thickness, as well as the erythema and oedema, were also reduced. These data suggest that marked eosinophil infiltration enhances skin inflammation in allergic contact dermatitis. Moreover, locally administered IL-5 functions remotely by controlling eosinophil recruitment into the skin. The guinea-pig model of contact sensitivity may be useful for evaluating therapies and pharmaceuticals targeted at eosinophil infiltration. PMID- 11122233 TI - Early decrease in surface expression of HLA-DQ predicts the development of infection in trauma patients. AB - The behaviour of human leucocyte antigen-DR (HLA-DR) following injury has been extensively studied. However, the behaviour of other class II antigens following trauma has not been characterized as well, despite evidence that HLA-DQ genotype influences the response to several bacterial antigens. Our study attempts to characterize and analyse the behaviour of HLA-DQ after trauma in patients with and without infection. Twenty-five patients were studied following major injury. Fifteen of the 25 patients developed infection (men = 11, women = 4); 10 patients developed no infection (men = 9, women = 1). The mean age was 34 +/- 12 years for patients with no infection and 52 +/- 20 years for those with infection. Monocyte HLA-DQ surface expression was determined using FITC-labelled antibodies and flow cytometry. Expression was compared with a control population of 11 healthy volunteers. The percentage of monocytes expressing HLA-DQ following trauma was reduced in patients with infection and in those without infection, but returned to normal (days 8-14) only in those patients who did not develop infection. Monocyte HLA-DQ mean channel fluorescence was reduced on day 1, but quickly returned to normal in those patients who subsequently developed infection. Stimulated with lipopolysaccharide, the initial samples of 13 patients who developed infection showed that surface expression on these monocytes could be elevated into the normal range. We conclude that HLA-DQ is an additional early marker of outcome that may not function merely as an immune suppressor. The maintained ability of HLA-DQ to present self-antigens may be important in the initial stages of the host response to injury. PMID- 11122234 TI - Gram-negative bacteria induce proinflammatory cytokine production by monocytes in the absence of lipopolysaccharide (LPS). AB - Tumour necrosis factor-alpha (TNF-alpha), IL-1alpha and IL-6 production by human monocytes in response to a clinical strain of the Gram-negative encapsulated bacteria Neisseria meningitidis and an isogenic lpxA- strain deficient in LPS was investigated. Wild-type N. meningitidis at concentrations between 105 and 108 organisms/ml and purified LPS induced proinflammatory cytokine production. High levels of these cytokines were also produced in response to the lpxA- strain at 107 and 108 organisms/ml. The specific LPS antagonist bactericidal/permeability increasing protein (rBPI21) inhibited cytokine production induced by LPS and wild type bacteria at 105 organisms/ml but not at higher concentrations, and not by LPS-deficient bacteria at any concentration. These data show that proinflammatory cytokine production by monocytes in response to N. meningitidis does not require the presence of LPS. Therapeutic strategies designed to block LPS alone may not therefore be sufficient for interrupting the inflammatory response in severe meningococcal disease. PMID- 11122235 TI - Dendritic cell function is perturbed by Yersinia enterocolitica infection in vitro. AB - Infection with Yersinia enterocolitica is the cause of intestinal or extraintestinal diseases. We investigated the role of dendritic cells (DC), the most potent antigen-presenting cell (APC), in the course of infection with Y. enterocolitica in vitro. For these studies, DC were isolated from human peripheral blood and infected with green fluorescent protein (GFP)-labelled Y. enterocolitica. Bacteria were found within DC by FACS analysis and viable bacteria could be cultured from lysed cells. Within 24 h after infection, DC upregulated CD83 and CD86 followed at day 3, indicating maturation of DC. In contrast, for MHC class II, a marked but transient downregulation was observed at day 3 after infection, and downregulation to a lesser extent for CD80 at day 5. To assess the immunostimulatory capacity of DC, viable infected and uninfected DC were incubated with autologous T cells in the presence of phytohemagglutinin A (PHA). T cell proliferation was significantly reduced at days 4-6 after infection but not thereafter, whereas nonpathogenic Escherichia coli was not able to mimick this suppressive effect of Y. enterocolitica. The same suppression could be observed when infected DC were used in a mixed leucocyte reaction with allogeneic T cells. Thus Y. enterocolitica is able to invade DC, does not induce necrosis or apoptosis, but affects maturation of DC. However, MHC class II-molecules are downregulated initially, which coincides with a diminished immunostimulatory capacity of DC infected with Y. enterocolitica. The diminished immunostimulatory capacity of DC following infection with Y. enterocolitica in vitro might impair or delay elimination of bacteria thereby contributing to pathogenesis of bacterial enteritis or extraintestinal manifestations such as reactive arthritis. PMID- 11122236 TI - Kinetic studies of the production of nitric oxide (NO) and tumour necrosis factor alpha (TNF-alpha) in macrophages stimulated with Burkholderia pseudomallei endotoxin. AB - The mechanism by which Burkholderia pseudomallei survives in macrophages is not clearly understood. In this study, we demonstrated that the mouse macrophage cell line (RAW 264.7) treated with lipopolysaccharide (LPS) from B. pseudomallei (BP LPS) produced significantly less NO and TNF-alpha compared with those stimulated with the LPS from Escherichia coli and Salmonella typhi. The time required for the BP-LPS to trigger substantial NO and TNF-alpha release was at least 30 min, compared with < 5 min for the E. coli-LPS. A time course study of inducible nitric oxide synthase (iNOS) protein expression also indicated that the time required for macrophages stimulated with the BP-LPS to up-regulate iNOS was longer. The longer time lag for the BP-LPS to activate macrophages was probably due to the delay in up-regulation of iNOS and TNF-alpha mRNA transcription. These results indirectly suggest that the delay of the mediators' production may be due to a reduced rate of signal transduction initiated by the interaction of BP-LPS with the macrophage cell surface. The use of MoAb to phosphorylated p38 in a Western blot analysis provided data compatible with the notion that the maximum level of phosphorylated p38 from the cells activated with BP-LPS was attained at a slower rate. These results suggest that the unique structure of BP-LPS exhibits a property which may interfere with macrophage cell activation. PMID- 11122237 TI - Production of transforming growth factor-beta 1 (TGF-beta1) by blood monocytes from patients with different clinical forms of leprosy. AB - In the present study, the concentration of TGF-beta1 secreted by adherent cells isolated from human peripheral blood mononuclear cells (PBMC) and either stimulated with PGL-1 or lipopolysaccharide (LPS) or left unstimulated was determined by ELISA. The cells were isolated from untreated patients with different clinical forms of leprosy and healthy individuals. The adherent cells exhibited spontaneous release of TGF-beta1 in all clinical forms of leprosy and in healthy individuals; however, lepromatous leprosy/borderline leprosy (LL/BL) patients presenting erythema nodosum leprosum (ENL) displayed significantly higher concentrations of TGF-beta1 than either the other patients studied or the controls. These high TGF-beta1 levels were consistently observed when LL/BL ENL cells were stimulated with phenolic glycolipid (PGL-1) or LPS, and even in the absence of a stimulus (P < 0.01). The most significant differences in TGF-beta1 levels were observed when comparing the results in the presence of PGL-1 from ENL with, in order of significance: tuberculoid leprosy (TT) patients (P < 0.001), LL/BL patients without ENL (P < 0.01), healthy individuals (P < 0.01) and borderline-borderline/borderline-tuberculoid (BB/BT) patients with reversal reaction (RR) (P < 0.01). The BB/BT patients produced equivalent levels of TGF beta1 compared with LL/BL patients without ENL, for all types of stimuli (P > 0.05). In contrast, TT patients produced the lowest levels of TGF-beta1 among all the subjects studied (both patients and healthy controls), especially following PGL-1 stimulation (P < 0.001, and P < 0.05, respectively). In conjunction with our previous data regarding TGF-beta1 expression in dermal lesions, it appears that TGF-beta1 probably plays different roles in leprosy: (i) to mediate a suppressive action locally, associated with the presence of PGL-1, and (ii) to induce proinflammatory effects when secreted systemically by monocytes, thereby acting as a modulatory cytokine in the acute inflammatory reactions of ENL and associated with the Th2 immune response in multibacillary forms of leprosy. PMID- 11122238 TI - Roles of tumour necrosis factor-alpha (TNF-alpha), transforming growth factor beta (TGF-beta), and IL-10 in the modulation of intercellular adhesion molecule-1 (ICAM-1) expression by macrophages during mycobacterial infection. AB - Profiles of ICAM-1 expression on cultured murine peritoneal macrophages infected with Mycobacterium avium complex (MAC) were examined, with special reference to modulating roles of TNF-alpha, TGF-beta, and IL-10. When macrophages were infected with MAC, ICAM-1 expression, measured by microscopic counting of ICAM-1+ macrophages stained with anti-ICAM-1 antibody, ELISA, and flow cytometric analysis, was rapidly increased, peaking at day 3 (early-phase up-regulation) due to endogenous TNF-alpha, and thereafter gradually declined to the normal level within 1 week or more (late-phase down-regulation). The late-phase ICAM-1 down regulation was also seen in macrophages phagocytosing heat-killed MAC and those stimulated with lipopolysaccharide but not in macrophages phagocytosing latex beads. ICAM-1 mRNA expression was augmented markedly at day 1 after MAC infection and thereafter decreased. While TNF-alpha and IL-10 production by MAC-infected macrophages was observed during the first 3 days, TGF-beta production was initiated from day 3 and continued until day 14. Exogenously added TGF-beta strongly inhibited the early-phase increase in ICAM-1 expression by infected macrophages, and the blockade of endogenous TGF-beta with anti-TGF-beta antibody markedly inhibited late-phase ICAM-1 down-regulation. Moderate blocking effect was also observed for anti-IL-10 antibody. On the other hand, late-phase ICAM-1 down-regulation was not prevented by the addition of exogenous TNF-alpha. Therefore, TGF-beta and IL-10, especially the former, appear to play active roles in the late-phase down-regulation of ICAM-1 in MAC-infected macrophages during long-term cultivation. PMID- 11122239 TI - Influence of disease severity on nitrite and cytokine production by peripheral blood mononuclear cells (PBMC) from patients with pulmonary tuberculosis (TB). AB - Earlier studies in patients with pulmonary TB have revealed a higher production of Th1 cell type cytokines in moderate TB, with predominant Th2-like responses in advanced disease. Given the influence of IL-12 in T cell differentiation, as well as the roles of transforming growth factor-beta (TGF-beta), nitric oxide and tumour necrosis factor-alpha (TNF-alpha) in the immune response against intracellular pathogens, we decided to analyse the interferon-gamma (IFN-gamma), IL-4, IL-12, TGF-beta, TNF-alpha and nitrite concentrations in culture supernatants of PBMC from TB patients showing different degrees of lung involvement. The sample population comprised 18 untreated TB patients with either moderate (n = 9) or advanced (n = 9) disease and 12 age- and sex-matched healthy controls (total population (patients and controls) 12 women, 18 men, aged 37 +/- 13 years (mean +/- s.d.)). PBMC were stimulated with whole sonicate from Mycobacterium tuberculosis and the supernatants were collected on day 4 for measurement of cytokine and nitrite levels. Antigen-stimulated IFN-gamma, TGF beta and TNF-alpha production was found to be significantly increased in TB patients, both moderate and advanced, compared with the controls. Levels of IFN gamma were significantly higher in moderate disease than advanced cases, whereas advanced cases showed significantly higher IL-12, TGF-beta and TNF-alpha concentrations when compared with cases of moderate TB. Nitrite levels were also increased in TB patients and the increase was statistically significant when advanced cases were compared with controls. These findings may contribute to a clearer picture of the net effect of cytokine interactions in TB, essential for a better understanding of the immunopathological mechanisms underlying the distinct clinical forms of the disease. PMID- 11122240 TI - T cell activation, apoptosis and cytokine dysregulation in the (co)pathogenesis of HIV and pulmonary tuberculosis (TB). AB - Immune parameters were compared in four groups of Ugandan subjects: HIV-and HIV+ adult patients with active pulmonary TB (HIV- PTB n = 38; HIV+ PTB n = 28), patients with HIV infection only (n = 26) and PPD+ healthy controls (n = 25). Compared with healthy controls, CD4 and CD8 T cells from patients with HIV and/or PTB expressed more activation markers (HLA-DR, CD38); their CD8 T cells expressed more CD95 (pre-apoptosis) and less CD28 (co-stimulatory receptor). Peripheral blood mononuclear cells (PBMC) of patients with either HIV or PTB were impaired in interferon-gamma (IFN-gamma) production upon antigenic stimulation. PTB (with or without HIV) was characterized by monocytosis, granulocytosis, increased transforming growth factor-beta 1 production and PPD-induced apoptosis. In vivo CD4 T cell depletion, in vitro increased spontaneous CD4 T cell apoptosis and defects in IFN-gamma responses upon mitogenic stimulation were restricted to HIV+ subjects (with or without PTB). Overlapping and distinctive immune alterations, associated with PTB and HIV, might explain mutual unfavourable influences of both diseases. PMID- 11122241 TI - HIV-1 Nef protein inhibits the in vitro induction of a specific antibody response to Candida albicans by an early up-regulation of IL-15 production. AB - We have previously demonstrated that exogenous Nef protein induced activation of normal human T cells up-regulating IL-15 production by monocytes. Since HIV-1 infection results in the early impairment of immune functions we decided to evaluate if Nef is able to modulate the induction of a specific antibody response. Human peripheral blood mononuclear cells from healthy donors were induced in vitro to mount a specific antibody response to the Candida albicans antigen. We show that Nef inhibited, in a dose-dependent manner, the induction of the anti-C. albicans antibody response. The ability of an anti-Nef antibody to prevent such inhibition indicates that the effect was indeed Nef-specific. In the Nef-treated cultures an early increase of IL-15 production was observed and the addition of anti-IL-15 antibody abrogated the Nef-induced inhibitory effect. Moreover the addition of IL-15 to the cultures inhibited, as well as Nef, the induction of the specific antibody response. Thus, our results suggest that Nef may inhibit the induction of a specific antibody response by an early up regulation of IL-15 production. A better comprehension of this phenomenon may be important for unravelling some aspects of the B cell defects in HIV infection. PMID- 11122242 TI - Apoptosis and apoptosis-associated perturbations of peripheral blood lymphocytes during HIV infection: comparison between AIDS patients and asymptomatic long-term non-progressors. AB - This study was designed to compare the degree of lymphocyte apoptosis and Fas-Fas ligand (FasL) expression in AIDS patients and long-term non-progressors (LTNPs) and correlate these parameters with apoptosis-associated perturbations in lymphocyte function. LTNPs had a lower frequency of apoptotic CD4+ and CD8+ T cells compared with subjects with AIDS. This correlated with a lower frequency of cells expressing Fas and FasL. The frequency of selected lymphocyte populations exhibiting a disrupted mitochondrial transmembrane potential (DeltaPsim) and increased superoxide generation was lower in LTNPs than in patients with AIDS; these abnormalities were associated with lower levels of caspase-1 activation in LTNPs. The results indicate a significantly reduced level of apoptosis and apoptosis-associated parameters in LTNPs than in patients developing AIDS. Based on these findings, a crucial role for mitochondria can be predicted in the process of lymphocyte apoptosis during the evolution of AIDS. PMID- 11122243 TI - Activation of signal transduction and apoptosis in healthy lymphomonocytes exposed to bystander HIV-1-infected cells. AB - Persistent activation of the immune system is one of the hallmarks of HIV-1 infection. In this study we analysed the induction of factors involved in cytokine signal transduction, such as STAT 1 proteins and IRF-1 mRNA, in normal peripheral blood mononuclear cells (PBMC) exposed to HIV-infected cells, and the induction of apoptosis. Western blot analyses and reverse transcriptase polymerase chain reaction results indicate that both cells infected with a X4 strain and cells infected with a R5 strain are able to increase intracellular levels of STAT 1alpha and beta proteins as well as IRF-1 mRNA. This effect was prevented by neutralizing antibodies against interferon-alpha (IFN-alpha). HIV-1 infected cells dose-dependently induced apoptotic commitment in normal PBMC, as revealed by DNA fragmentation analysis, but this was not accompanied by an increase of caspase-3 activity, even if a slight up-regulation of IL-1beta converting enzyme mRNA was detected. Apoptosis induction could be abrogated mainly by antibodies against tumour necrosis factor-alpha (TNF-alpha) and, to a lesser extent, by antibodies against IFN-gamma. All these findings suggest that uninfected PBMC can undergo activation of signal transduction and apoptosis after exposure to bystander HIV-infected cells, subsequent to the induction of cytokines such as IFNs and TNF-alpha. PMID- 11122244 TI - Induction of CD95 ligand expression on T lymphocytes and B lymphocytes and its contribution to apoptosis of CD95-up-regulated CD4+ T lymphocytes in macaques by infection with a pathogenic simian/human immunodeficiency virus. AB - Using an established SIV/HIV-C2/1-infected cynomolgus monkey model displaying stable CD4+ T cell depletion, the kinetics of apoptosis and the levels of expression of CD95 membrane-associated CD95L on lymphocytes were investigated to test the involvement of the CD95/CD95L system in CD4+ T lymphocyte loss in vivo. Rapid depletion of CD4+ T cells occurred up to 2 weeks after infection, with chronic CD4+ T lymphopenia thereafter. During the initial CD4+ T cell loss, which was accompanied by viraemia, about 90% of the peripheral CD4+ T cell subset underwent spontaneous apoptotic cell death during 24 h of culture. Increased expression of CD95 was observed on both CD4+ and CD8+ T cell subsets, with CD95 expression on CD8+ cells declining rapidly, but high CD95 expression being maintained on CD4+ cells. Since CD95L was expressed on CD8+ T cells, B cells and to a lesser extent on CD4+ T cells, this suggests that CD95-mediated apoptosis might be controlled in an autocrine/paracrine fashion. PMID- 11122245 TI - Characterization of the T cell recognition of hepatitis B surface antigen (HBsAg) by good and poor responders to hepatitis B vaccines. AB - To study the regulation of the human cellular immune response to HBsAg we produced a series of HBsAg-specific T cell lines from good and poor responders to the hepatitis B vaccine. All T cell lines expressed CD4 on their membrane and could therefore be considered of the helper/inducer phenotype. The different HBsAg-specific T cell lines were restricted by HLA-DRB5*0101, DRB1*1201, DRB1*0701, -DRB1*0301, -DPB1*0201, -DPB1*0402, and -DPB1*0901. In good responders to the hepatitis B vaccine different HLA molecules could act as restricting element. In poor responders the diversity of HLA class II restriction determinants was more limited. This leads us to conclude that the immune response to HBsAg is multispecific and polyclonal in good responders and paucispecific and oligoclonal in poor responders to the hepatitis B vaccine. By using a panel of synthetic peptides representing selected sequences of the HBsAg, the fine specificities of each of these T cell lines could be determined. Strikingly, the majority of the identified T cell epitopes was located in and around the first hydrophobic transmembranous region of the HBsAg. This was observed in T cell lines from good and poor vaccine responders, without distinction. The remarkable T cell immunogenicity of this region may reside in its richness in binding motifs for a variety of HLA class II determinants. PMID- 11122246 TI - Hepatitis C virus (HCV) in lymphocyte subsets and in B lymphocytes expressing rheumatoid factor cross-reacting idiotype in type II mixed cryoglobulinaemia. AB - The IgMk rheumatoid factors (RF) of type II mixed cryoglobulinaemia (MC) react, in 95% of cases, with MoAbs against the cross-reactive idiotypes (CRI) Cc1 or Lc1 (corresponding to the products of the VH1 and VH4 genes). MC is closely associated with HCV infection, a virus which infects lymphocytes and may replicate in B cells. It has been suggested that HCV may induce clonal selection of B cells producing monoclonal IgMk RF in type II MC. To verify whether HCV is enriched in B cells, and in the subsets expressing Cc1 and Lc1 CRI, we studied peripheral blood lymphocytes from eight patients with MC and HCV RNA-positive sera. Seven patients had RF reacting with anti-Cc1, the other with anti-Lc1 CRI. Total lymphocytes, T cells, B cells, and Cc1+ or Lc1+, Cc1- or Lc1- B cells were purified using MoAb-coated magnetic beads. Lymphocyte subsets were then diluted to give a range of 1 x 106-1 x 103 cells and tested for HCV RNA by reverse transcriptase-polymerase chain reaction. HCV was found exclusively in B cells in seven out of eight patients. In three patients HCV was enriched in the Cc1+ cells. In one of these patients, HCV was found exclusively in Cc1+ cells, with Cc1- cells being HCV-. The data indicate that B cells from type II MC patients are almost constantly infected by HCV. In selected cases, B cell subsets expressing IgMk RF CRI are the prevalent cell type infected by HCV. Our data suggest HCV involvement in B cell dysregulation leading to cryoprecipitable IgMk RF production. PMID- 11122248 TI - Point mutations in the promoter region of the CYBB gene leading to mild chronic granulomatous disease. AB - Chronic granulomatous disease (CGD) is a clinical syndrome of recurrent bacterial and fungal infections caused by a rare disorder of phagocytic cells. In CGD, the phagocytes are unable to generate oxygen radicals after stimulation of these cells, due to a defect in the NADPH oxidase system. This NADPH oxidase is a multicomponent enzyme of at least four subunits, of which the beta-subunit of cytochrome b558, gp91-phox, is encoded by an X-linked gene (called CYBB). We report here five patients from two families; in each family we found a different mutation in the promoter region of CYBB. Both mutations prevented the expression of gp91-phox in the patients' neutrophils and thus caused inability of these cells to generate oxygen radicals. However, the mutations left the gp91-phox expression and the function of the NADPH oxidase in the patients' eosinophils intact. The relatively mild course of the CGD in these patients can probably be attributed to the fact that the eosinophils have retained their oxidative capacity. Furthermore, our results indicate that neutrophils and eosinophils differ in their regulation of gp91-phox expression. PMID- 11122247 TI - Coxsackievirus B3 infection induces anti-flavoprotein antibodies in mice. AB - Enteroviruses, the most common cause of acute myocarditis, are also supposed aetiological agents of dilated cardiomyopathy. Autoantibodies (anti-M7; Klein & Berg, Clin Exp Immunol 1990; 58:283-92) directed against flavoproteins with covalently bound flavin (alphaFp-Ab; Otto et al., Clin Exp Immunol 1998; 111:541 2) are detected in up to 30% of sera of patients with myocarditis and idiopathic dilated cardiomyopathy (IDCM). Mice inoculated with a myocarditic variant of coxsackievirus B3 (CVB3) were employed to study the occurrence of serum alphaFp Ab following viral infection. The presence of alphaFp-Ab was analysed by Western blotting with the flavoprotein antigens 6-hydroxy-D-nicotine oxidase (6HDNO) and sarcosine oxidase (SaO). Of 10 sera from CVB3-infected mice, five showed a strong reaction with both antigens. The sera were reactive also to the mitochondrial covalently flavinylated proteins dimethylglycine dehydrogenase and sarcosine dehydrogenase. Sera of non-infected mice did not react with these antigens. A 6HDNO mutant protein with non-covalently bound FAD no longer reacted on Western blots with sera of CVB3-infected mice. Preincubation with FAD abolished or reduced the reaction of the sera with the 6HDNO antigen. At 2 weeks p.i. the alphaFp-Ab were of the IgM and IgG isotypes, at 7 and 9 weeks p.i. of the IgG isotype. The sera of CVB3-infected mice reproduced closely the antigenic specificity of the anti-M7 sera of patients, lending further support to the role of coxsackieviruses in the pathogenesis of IDCM. PMID- 11122249 TI - Blood fetal microchimerism in primary biliary cirrhosis. AB - The autoimmune nature of primary biliary cirrhosis (PBC) is well established. We tested the hypothesis that fetal microchimerism indicated by the persistence of circulating fetal cells in women years after pregnancy might contribute to the aetiopathogenesis of PBC through a graft-versus-host-like response. We extracted DNA from the peripheral blood cells of 36 women carefully selected from 173 consecutive PBC patients, who were matched with 36 healthy women by age, age of last son, and number of children. Both patients and controls had to have male offspring, and no history of miscarriages or blood transfusions; they could not be twins. We tested all of the samples for the presence of two specific Y chromosome sequences (SY154 and SRY) by amplifying DNA in a nested polymerase chain reaction. Y-chromosome-specific DNA was detected in the peripheral blood cell DNA of 13 (36%) of the 36 women with PBC and in 11 (31%) of the 36 healthy controls. The two groups of PBC patients with and without male DNA sequences were similar in terms of their clinical, biochemical, and serological features. Y chromosome sequences were found in three of the four PBC women with associated systemic sclerosis. All of the 24 Y-positive samples contained SY154 sequences, but only three PBC patients and six controls showed the presence of both SY154 and SRY sequences. This discrepancy may suggest that not only fetal cells but also fragments of fetal DNA are present in maternal circulation. Overall, our data do not support the hypothesis that fetal microchimerism plays a significant role in the onset or progression of PBC. PMID- 11122250 TI - Secretory autoantibodies in primary biliary cirrhosis (PBC). AB - It is unclear how breakdown in immune tolerance to the ubiquitous self-antigen pyruvate dehydrogenase complex (PDC), seen in the autoimmune liver disease PBC, gives rise to tissue damage with such a limited distribution (restricted to the liver and salivary and lachrymal glands). One property shared by these tissues is the ability to export secretory IgA by the process of transcytosis. The aim of this study was to address whether active transcytosis of anti-PDC IgA occurs across epithelial surfaces in PBC, a finding that might implicate mucosal specific immune mechanisms in the pathogenesis of this disease. Parotid saliva was collected from PBC patients (n = 44), normal controls (n = 28) and PBC patients post-liver transplantation (n = 11). IgA and secretory component positive antibodies specific for human PDC were quantified by ELISA and immunoblotting. PBC patients (but not control subjects) had anti-PDC IgA in their saliva. The strong correlation seen between titres detected using anti-IgA and anti-secretory component antibodies suggests that this is predominantly secretory IgA reaching the saliva by the active process of epithelial transcytosis. Titres of anti-PDC IgA remain high in PBC patients saliva post-liver transplant. Findings from studies of IgA in viral infection models raise the possibility that anti-PDC IgA could, whilst undergoing transcytosis, bind to newly translated PDC components in the cytoplasm of the epithelial cells transporting them out of the cell and inducing metabolic damage. This model would, if correct, help to explain the mechanism and tropism of tissue damage in PBC and the aberrant pattern of expression of PDC on the apical surface of biliary and salivary epithelial cells reported in this disease. PMID- 11122251 TI - Expression of myelin basic protein (MBP) epitopes in human non-neural cells revealed by two anti-MBP IgM monoclonal antibodies. AB - Two monoclonal antibodies (1H6.2 and 45.30) were raised against MBP purified from human brain under experimental conditions that allowed MBP to retain binding to surrounding myelin lipids (human lipid-bound MBP (hLB-MBP)). 1H6.2 and 45.30 MoAbs were selected on the basis of their different binding properties to: hLB MBP, human lipid-free-MBP (hLF-MBP) and bovine lipid-free-MBP (bLF-MBP). Although the isotype of both MoAbs was IgM, their specificity, as tested in ELISA assays against chemical haptens and unrelated protein antigens, was restricted to MBP. 1H6.2 and 45.30 MoAbs stained MBP from human brain white matter tissue extracts, as well as bLF-MBP, in Western blot assays. Both MoAbs stained oligodendrocytes and myelin in immunohistochemical analysis of white matter from human brain. Tissue sections from human peripheral nerves were labelled by 1H6.2 only, however, demonstrating that the MoAbs recognize two different epitopes. Epitopes recognized by 1H6.2 and 45.30 MoAbs were also expressed by a wide array of human non-neural cells of either normal or pathological origin, as evidenced by cytofluorimetric assays. In particular, MBP epitopes (MEs) were expressed by lymphoid cells as well as by cells which play a pivotal role in immune homeostasis and in the immune response, such as thymic epithelial cells and professional antigen-presenting cells. Both MoAbs were efficiently internalized by cells from a human B cell line, suggesting trafficking of MEs along the endocytic pathways. These findings support hypotheses regarding the role of MEs expressed by non-neural cells in establishing self-tolerance and/or in triggering the immune response against MBP antigen. PMID- 11122252 TI - Autoantigen-pulsed dendritic cells induce tolerance to experimental allergic encephalomyelitis (EAE) in Lewis rats. AB - Dendritic cells (DC) can modulate the nature of immune responses in a stimulatory or tolerogenic fashion. Great attention has been given to the induction of immunity to tumour and infection. In this study, bone marrow-derived DC from healthy Lewis rats were pulsed in vitro with encephalitogenic myelin basic protein peptide 68-86 (MBP 68-86), and injected subcutaneously (1 x 106/rat) into normal Lewis rats. Upon observation of the rats pretreated in this way for 4 weeks, when no clinical signs of EAE occurred, these rats were immunized with MBP 68-86 and Freund's complete adjuvant. The pretreated rats failed to develop clinical EAE. This tolerance was associated with augmented proliferative responses, interferon-gamma secretion, inducible nitric oxide synthase (iNOS) expression and NO production. The frequency of apoptotic cells was increased in the rats receiving MBP 68-86-pulsed DC compared with unpulsed control DC. Few infiltrating inflammatory cells were observed in spinal cord sections from rats that had received MBP 68-86-pulsed DC. The data are compatible with the interpretation that MBP 68-86-pulsed DC induce tolerance to EAE possibly through up-regulation of iNOS expression and NO production, which mediate cell apoptosis, thereby reducing infiltration of inflammatory cells within the central nervous system. PMID- 11122254 TI - MHC class I pathway is not required for the development of crescentic glomerulonephritis in mice. AB - MHC II and CD4+ T cells are required for anti-glomerular basement membrane (GBM) globulin-initiated crescentic glomerulonephritis (GN) in mice, but the role of MHC I and CD8+ T cells is unclear. The cytolytic function of CD8+ T cells requires recognition of peptide antigens presented on MHC I. CD8+ T cells can also perform helper functions via cytokine production. The contribution of MHC I to crescentic GN was investigated using TAP-1 gene knock out (TAP-1-/-) mice, which have deficient MHC I antigen presentation. Heterozygous TAP-1 mice have normal MHC I expression and developed GN with crescents in 42 +/- 4% of glomeruli (normal 0%), proteinuria (9.1 +/- 1.6 mg/20 h, normal 1.5 +/- 0.3 mg/20 h) and impaired renal function (creatinine clearance 110 +/- 8 microl/min, normal 193 +/ 10 microl/min) following administration of sheep anti-mouse GBM globulin. TAP-1 /- mice, which have extremely low MHC I expression and reduced CD8+ T cells, developed similar GN with 39 +/- 3% crescents, proteinuria (12.7 +/- 4.3 mg/20 h) and impaired renal function (creatinine clearance 123 +/- 20 microl/min). In vivo antibody-induced CD8 depletion did not attenuate crescent formation or protect renal function in C57Bl/6 mice developing GN, although significant reduction in proteinuria (5.3 +/- 1.2 mg/20 h, P = 0. 012) and glomerular recruitment of CD4+ T cells and macrophages were observed compared with control treated mice with GN. These data demonstrate that MHC I is not required for development of crescentic GN in mice. The MHC I-independent contribution of CD8+ T cells to proteinuria and inflammatory cell recruitment suggests that they may serve a 'helper' rather than cytolytic role in this disease. PMID- 11122253 TI - The protective effects of omega-6 fatty acids in experimental autoimmune encephalomyelitis (EAE) in relation to transforming growth factor-beta 1 (TGF beta1) up-regulation and increased prostaglandin E2 (PGE2) production. AB - Polyunsaturated fatty acids are known to affect the immune response and administration of the omega-6 fatty acid linoleic acid has been reported to be beneficial in multiple sclerosis (MS) and EAE. In this study we have investigated the effects of oral feeding of plant lipid rich in the omega-6 fatty acid gamma linolenic acid from Borago officinalis on acute and relapse disease and the immune response in EAE using SJL mice. EAE was induced by an encephalitogenic peptide (92-106) of myelin oligodendrocyte glycoprotein (MOG), and mice were fed the plant lipid daily from 7 days after EAE induction to assess the effects on acute disease and from day 25 to assess the effects on disease relapse. The clinical incidence and histological manifestations of acute EAE, and the clinical relapse phase of chronic relapsing EAE (CREAE) were markedly inhibited by omega-6 fatty acid feeding. A significant increase in the production of TGF-beta1 in response to concanavalin A (Con A) at day 13 and a significant increase in TGF beta1 and PGE2 to Con A, PPD and MOG peptide (92-106) at day 21 were detected in spleen mononuclear cells from fatty acid-fed mice. There was no difference in interferon-gamma, IL-4 and IL-2 production between the fatty acid-fed and control groups. Significantly higher TGF-beta mRNA expression was found in the spleens of omega-6 fatty acid-fed mice at day 21. There were no differences in spleen cell proliferative response to Con A, PPD and MOG peptide (92-106). Biochemical analysis of spleen cell membrane fatty acids revealed significant increases in the eicosanoid precursor fatty acids dihomo-gamma-linolenic acid and arachidonic acid in response to gamma-linolenic acid feeding, indicating rapid metabolism to longer chain omega-6 fatty acids. These results show that oral feeding of gamma linolenic acid-rich plant lipid markedly affects the disease course of acute EAE and CREAE and is associated with an increase in cell membrane long chain omega-6 fatty acids, production of PGE2 and gene transcription and, on activation, secretion of TGF-beta1. PMID- 11122255 TI - Expression of cyclooxygenase-1 (COX-1) in labial salivary glands of Sjogren's syndrome. AB - COX plays an important role in inflammatory diseases such as rheumatoid arthritis. To determine the role of COX in Sjogren's syndrome (SS), we examined COX expression in the salivary glands of SS patients. We examined 15 patients with SS and two normal subjects. Labial salivary gland tissue samples were analysed immunohistochemically using anti-COX-1 and COX-2 antibodies. All biopsy samples from 15 patients with SS were stained for COX-1. In contrast, COX-1 immunostaining was not detected in normal salivary gland tissues. Co-expression of COX-1 and CD68 was confirmed by mirror section technique and double antibody immunostaining. This finding indicated that COX-1-expressing cells in SS salivary glands were infiltrating macrophages. In contrast to COX-1 staining, only a little COX-2 immunostaining was observed in salivary gland tissues from SS patients. These data suggest that COX-1 expression on infiltrating macrophages may contribute to the inflammatory process of salivary glands in SS. PMID- 11122256 TI - Increased interferon-gamma (IFN-gamma), IL-10 and decreased IL-4 mRNA expression in peripheral blood mononuclear cells (PBMC) from patients with systemic lupus erythematosus (SLE). AB - Cytokines are important regulators of lymphocyte function in SLE. However, the profile of Th1 and Th2 cytokines produced by circulating lymphocytes in human SLE has not been clearly elucidated. The aim of the present study was to characterize the gene expressions of the Th1-type cytokine IFN-gamma, and the Th2-type cytokines IL-10 and IL-4 in PBMC of 15 patients with SLE and 10 healthy individuals by a semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR). Our results showed that expression of IFN-gamma (P = 0.0004) and IL-10 (P = 0.002) transcripts were significantly increased in PBMC of patients with SLE compared with healthy controls. By contrast, expression of IL-4 transcripts in PBMC of patients with SLE was significantly decreased compared with the healthy controls (P = 0.0008). Primary sources of IL-10 were B cells and monocytes, with variable contribution of T cells as detected in various fractions of PBMC of patients with SLE (P = 0.049). These findings support the hypothesis that the enhanced production of IFN-gamma by mononuclear cells may trigger inflammatory responses, together with the enhanced production of IL-10 resulting in autoantibody production by B cells in human SLE. PMID- 11122257 TI - IL-10 stimulates production of platelet-activating factor by monocytes of patients with active systemic lupus erythematosus (SLE). AB - IL-10 displays modulatory properties on the synthesis of platelet-activating factor (PAF), a potent inflammatory mediator of vascular injury. Despite the fact that IL-10 is considered to be an anti-inflammatory cytokine, IL-10 levels correlate with disease activity in SLE. Moreover, in SLE IL-10 is unable to exert its immunosuppressive and anti-inflammatory effects. We have investigated the ability of IL-10 to stimulate PAF production from monocytes of SLE patients. Spontaneous and IL-10-stimulated PAF production by peripheral blood monocytes was measured in active (n = 13) and inactive (n = 14) SLE patients and in 15 normal control subjects. We observed that monocytes derived from patients with active SLE, but not from controls or inactive SLE, spontaneously produced significant amounts of PAF. Moreover, IL-10 enhanced the synthesis of PAF from monocytes of active SLE patients only. IL-10-induced PAF production correlated with the severity of the disease and with the extent of proteinuria. These results indicate that IL-10 only stimulates the synthesis of PAF from monocytes of SLE patients when immunologically active, suggesting that IL-10 may possess a paradoxical proinflammatory effect in SLE by promoting the production of PAF, a secondary mediator of inflammation. PMID- 11122258 TI - Parasite-mediated down-regulation of collagen-induced arthritis (CIA) in DA rats. AB - Microbial infection can impact on the course of autoimmune disease, both in disease-inducing and disease-protecting capacities. Here we investigated if infection with Trypanosoma brucei brucei (Tbb), the protozoan causative agent of African Sleeping Sickness, could ameliorate the course of CIA in the Dark Agouti rat, an experimental model which shares many features with human rheumatoid arthritis. Infection of animals with living, but not inoculation with dead Tbb resulted in complete or significant reduction of clinical arthritic symptoms. Infection prior to collagen immunization was more effective than a later treatment, and this effect was related to the level of parasitaemia. Using reverse transcriptase-polymerase chain reaction we detected an increase in interferon-gamma mRNA in the draining lymph nodes of Tbb-treated animals relative to controls at day 28 after disease induction. Transforming growth factor-beta could be detected in the lymph nodes in four out of six animals that had received Tbb. In the joints, immunohistochemistry revealed reduced production of tumour necrosis factor-alpha in Tbb-treated animals relative to controls. The most striking difference between Tbb-infected and control groups, as measured by ELISA, was the down-regulation of anti-collagen II IgG antibody responses in parasite-infected animals. We conclude that live parasites can exert an immunomodulatory and protective effect in CIA in which several mechanisms may work in parallel, although the almost complete down-regulation of the anti collagen antibody response may alone explain the protective effect in CIA. The described model may be useful in further attempts to use the mechanisms involved in parasite immune defence to prevent and treat certain autoimmune conditions. PMID- 11122259 TI - Decreased expression levels of L-selectin on subsets of leucocytes and increased serum L-selectin in severe psoriasis. AB - L-selectin is a leucocyte adhesion molecule involved in leucocyte interactions with vascular endothelial cells. Following leucocyte activation L-selectin is endoproteolytically released from the cell surface. To assess whether psoriasis vulgaris results in systemic leucocyte activation, we examined expression levels of L-selectin on subsets of peripheral blood leucocytes from patients with psoriasis (n = 25) and normal control subjects. Serum levels of soluble L selectin were quantified by ELISA in patients with psoriasis (n = 75), pustulosis palmaris et plantaris, and contact dermatitis, as well as normal control subjects. Psoriasis severity was evaluated by psoriasis area and severity index (PASI). L-selectin expression levels on CD4+ T cells, B cells, monocytes, and neutrophils from patients with severe-type psoriasis (PASI > or = 15) was significantly decreased compared with leucocytes from normal control subjects. Furthermore, L-selectin expression on CD4+ T cells showed good inverse correlation with PASI scores. Monocyte L-selectin expression was restored when the skin lesions of psoriasis were remitted. The frequencies of L-selectin+ CD4+ T cells or L-selectin+ CD8+ T cells from patients with psoriasis were almost normal. Serum L-selectin levels in patients with severe-type psoriasis were significantly higher than those in normal control subjects. These results suggest that subsets of leucocytes may be activated in psoriasis, and that L-selectin expression levels on some leucocyte subsets, especially CD4+ T cells, tend to correlate with disease severity of psoriasis. PMID- 11122260 TI - Autoantibodies to early endosome antigen (EEA1) produce a staining pattern resembling cytoplasmic anti-neutrophil cytoplasmic antibodies (C-ANCA). AB - Autoantibodies to EEA1 have been described in patients with neurological diseases, subacute cutaneous lupus and a variety of other conditions, including a patient with Wegener's granulomatosis (WG). EEA1 is a hydrophilic peripheral membrane protein transiently associated with the cytoplasmic face of early endosomes. Antibodies to EEA1 produce a staining pattern that resembles the C ANCA pattern produced by anti-proteinase 3 (PR3) antibodies in WG sera. Co localization studies show incomplete overlap of the staining produced by anti EEA1 with anti-PR3. We showed that 0/40 unselected sera, from a cohort of WG patients and antibodies to PR3, reacted with EEA1. In addition, 1/15 sera that have a C-ANCA staining pattern but do not react with PR3 in an ELISA, immunoprecipitated the recombinant EEA1 protein. We conclude that although antibodies to EEA1 produce a staining pattern that resembles anti-PR3 and C-ANCA, antibodies to EEA1 in WG are rare. However, some C-ANCA+ sera that do not react with PR3 may contain EEA1 autoantibodies. PMID- 11122261 TI - Opsonization of apoptotic neutrophils by anti-neutrophil cytoplasmic antibodies (ANCA) leads to enhanced uptake by macrophages and increased release of tumour necrosis factor-alpha (TNF-alpha). AB - Since proteinase 3 (PR3)-ANCA interact with PR3 on the surface of apoptotic polymorphonuclear neutrophils (PMN) and ingestion of apoptotic PMN is known to modulate macrophage inflammatory reactions, we raised the question whether PR3 ANCA-opsonized apoptotic PMN influence the uptake by macrophages and their state of activation. We therefore analysed the effects of PR3-ANCA-opsonized apoptotic PMN on the uptake process by enzymatic assay. We further investigated the production of TNF-alpha, IL-10, IL-12 and the secretion of lipid inflammatory mediators (TxB2, leukotriene B4 (LTB4) and prostaglandin E2 (PGE2)) by human monocyte-derived macrophages using FACS and ELISA methods. We show that PMN opsonization by PR3-ANCA substantially enhances phagocytosis by macrophages and thereby triggers the production of TNF-alpha and TxB2. These in vitro findings indicate that PR3-ANCA opsonization of apoptotic PMN might be an important mechanism in the pathogenesis of Wegener's granulomatosis (WG), prompting macrophages to produce proinflammatory mediators. These mediators, mainly TNF alpha, might prime further PMN leading to perpetuation of the known priming dependent mechanisms of ANCA action. PMID- 11122262 TI - In vitro T lymphocyte responses to proteinase 3 (PR3) and linear peptides of PR3 in patients with Wegener's granulomatosis (WG). AB - T cell-mediated immunity is thought to play an important role in the pathogenesis of WG. In previous studies a minority of WG patients as well as some healthy controls showed in vitro proliferation of their peripheral blood mononuclear cells (PBMC) to PR3, the main autoantigen in WG. The relevant peptides responsible for this in vitro proliferation have not been identified. In order to define immunogenic peptides, PBMC of 13 WG patients in remission and 10 healthy controls were tested for proliferation to linear peptides of PR3 and to whole PR3. Fifty overlapping peptides spanning the whole PR3 sequence were synthesized. Peptides were tested in pools of five peptides and as single peptide. PBMC of two WG patients and one healthy control proliferated to whole PR3 and to peptide pools. In addition, 10 WG patients and eight healthy controls that did not proliferate to whole PR3 did proliferate to pools of PR3 peptides. Although more WG patients tended to react to particular peptide pools, no significant difference was seen between lymphocyte proliferation to PR3 peptides of WG patients and that of healthy controls. The pools of peptides recognized were mainly located at the N- and C-terminus of PR3. No correlation was observed between HLA type and proliferation on particular peptide pools. No proliferation of PBMC was observed to single peptides. In conclusion, T cells of WG patients proliferate in vitro more frequently to PR3 peptides than to the whole PR3 protein. Peptides derived from the signal sequence, the propeptide or peptides located at the C-terminus of PR3 induce highest levels of proliferation. No specific PR3 sequence could be identified that was preferentially recognized by PBMC of WG patients compared with controls. PMID- 11122263 TI - Expression of IL-8 in Kawasaki disease. AB - We investigated, by Northern blotting, ELISA, and a chemotaxis assay, the expression of IL-8 mRNA, the production of IL-8 protein, and the biological activity of mononuclear cells (MNC), polymorphonuclear neutrophils (PMN) and plasma, respectively, from patients with Kawasaki disease (KD) who received intravenous immunoglobulin (IVIG). IL-8 mRNA expression by MNC and PMN, the level of IL-8 protein, and the neutrophil chemoattractant activity within plasma were all increased in the acute phase of KD, and were significantly elevated following IVIG therapy. The level of chemotactic activity of neutrophils, but not that of monocytes, in response to F-met-leu-phe was decreased in patients with KD after IVIG. The increased expression of IL-8 in PMN and MNC, the increased plasma level of IL-8 and the decreased level of neutrophil chemotactic activity of the patients who received IVIG therapy might inhibit the accumulation of neutrophils at the sites of inflammation, and may thus reduce the risk of aneurysm formation. PMID- 11122264 TI - Red cell indices as predictors of iron depletion in blood donors. AB - The objective of this study was to investigate whether red cell indices mean cell volume (MCV) and mean cell haemoglobin (MCH) were lower in frequent blood donors and hence, indirectly, able to predict impending iron depletion. Serum ferritin and/or soluble transferrin receptor levels can be used to evaluate iron status but are not practical for routinely screening blood donors prior to donation. Hb, MCV and MCH were measured on venous blood from 886 blood donors using a Sysmex E 5000. Full details were obtained for all donors of each earlier donation over the previous 3 years. MCV and MCH levels were lowest in donors with the highest frequency of previous blood donation. There was a significant negative correlation between MCV and number of donations in males and females and between MCH and number of donations in females, over the 3 year period 1995-97. Similar trends were observed when only the previous year's donations (1997) were considered with all categories showing significant negative correlations and additionally, Hb levels in females showed negative correlation with number of donations in 1997. In conclusion, increased frequency of blood donations is associated with lower MCV and MCH. These red cell indices, or more sophisticated parameters such as percentage hypochromic cells, should be used to monitor early onset of iron depletion in frequent blood donors. PMID- 11122265 TI - Malaria diagnosis with the haematology analyser Cell-Dyn 3500: What does the instrument detect? AB - The Cell-Dyn 3500 instrument could become a sensitive and specific tool in the diagnosis of malaria. The instrument appears to detect malaria-pigment within monocytes and granulocytes. A case of P. vivax malaria in a patient with increased osmotically resistant erythrocytes illustrates the potential of the instrument to detect intraerythrocytic parasites with pigment. However, in most malaria-patients with normal red cell osmotic resistance the observed phenomena seem rather to represent intraleukocytic pigment. This can remain in the circulation of clinically and parasitologically cured individuals and thus may not indicate acute infection. While the instrument can indicate those patients who have been infected a diagnosis of acute malaria must be established independently. PMID- 11122266 TI - Detection of T cell receptor delta gene rearrangements in childhood B and T lineage acute lymphoblastic leukaemia by southern blot and PCR: technical comparison of two methods of analysis. AB - Molecular analysis of antigen receptor genes (Ig and TCR) has been useful for clonal studies in acute lymphoblastic leukaemia (ALL) patients. Rearrangements of these genes can be used to track the persistence of the leukaemic clone during the therapy. The purpose of our study was to analyse the percentage and the pattern of the rearrangements at the TCR D locus in a series of ALL patients, comparing the results obtained by Southern blot and PCR. Genomic DNA was extracted from mononuclear BM cells of 40 paediatric ALL cases, digested with different restriction enzymes and hybridized to TCRDJ1 probe to study the TCR delta locus. Amplification of the rearranged TCR delta genes was performed by PCR to define the gene segments involved. The junctional region was deduced from the sequence to obtain patient-specific primers. Among the 31 B lineage ALL samples, one or two TCR delta alleles proved to be rearranged in 53% of cases. Two different types of rearrangements were chiefly detected: Vdelta2Ddelta3 and Ddelta2Ddelta3. In T-ALL patients, the predominant rearrangement involved the Vdelta1 and the Jdelta1 gene segments. PMID- 11122267 TI - Up-regulation of serine/threonine protein phosphatase type 2A regulatory subunits during methylprednisolone-induced differentiation of leukaemic HL-60 cells. AB - Serine/threonine protein phosphatase 2A (PP2A) may play a role in leukaemic cell differentiation of the HL-60 myeloid leukaemic cell-line after methylprednisolone induction. We have investigated the specific enzyme activity and expression of catalytic and regulatory subunits of PP2A. The resulting specific enzyme activity and immunoblots showed an increase in enzyme activity and the expression of regulatory subunits after methylprednisolone treatment. There was no change in the expression of PP2A catalytic subunits. It is suggested that the effect of methylprednisolone on leukaemic differentiation may be the result of PP2A upregulation. PMID- 11122268 TI - A survey of patients with acquired haemophilia in a haemophilia centre over a 28 year period. AB - Data of patients with acquired inhibitors to clotting factors seen in a haemophilia centre from 1970-98 was collected. Twenty-four patients with anti factor VIII antibodies and four with acquired von Willebrand disease case records were evaluable. Two-thirds of the patients were females and their age ranged from 2 to 92 years (median 69). There was no correlation between the inhibitor titres and the severity or the pattern of bleeding. Porcine factor VIII and activated prothrombin complex were both effective for acute bleeds. Prednisolone and cyclophosphamide proved valuable for suppression of the antibodies without adverse effects even in the elderly group of patients. Three deaths were directly related to bleeding. This review has shown the life-threatening nature of this acquired bleeding disorder, its variable clinical presentation and the importance of early referral to a specialist centre for appropriate therapy. PMID- 11122269 TI - A capillary whole blood method for measuring the INR. AB - This study describes a method of measuring the INR on native whole blood capillary samples using Innovin recombinant thromboplastin. Modification of the reagent was necessary to compensate for the nonoptimal level of calcium in the sample/reagent mixture. Ninety-five percent of results obtained by the capillary blood method were no more than 0.42 INR higher or 0.38 INR lower than the venous blood method. The effect of changes in haematocrit was minimal. Significant differences in results were found between the Innovin and Thrombotest capillary blood methods. Provided the reagent was properly stored, there was no reagent drift and satisfactory results were obtained on samples supplied by UKNEQAS (coagulation) from previous trials. The method described is a convenient, simple and accurate method of measuring the INR using native capillary whole blood and Innovin recombinant thromboplastin. PMID- 11122271 TI - INR: Intervals of measurement can safely extend to 14 weeks. AB - The number of patients taking warfarin is increasing. Each of these patients requires regular blood tests to ensure that the level of anticoagulation reaches and remains within a defined target range. INR monitoring is expensive and time consuming and the frequency of the tests has repercussions for NHS resources and patient convenience. Guidelines regarding this are available, but the research on which these are based is extremely limited. Many centres in the UK are using extended intervals between tests that exceed the current guidelines. This study shows that for selected patients the interval between INR tests can safely be extended to 14 weeks with the potential for longer intervals. PMID- 11122270 TI - Variation in inhibitor reactivity in acquired haemophilia A with different concentrates. AB - Acquired haemophilia A due to the development of auto-antibodies directed against factor VIII (FVIII) is a rare disorder that may result in serious haemorrhagic episodes. Although in many cases no associated underlying disorders are apparent, the condition has been reported in association with autoimmune disorders, haematological malignancies, treatment with certain drugs and pregnancy. The reaction kinetics of auto-antibodies to FVIII differ from those observed with allo-antibodies in congenital haemophilia. Previous studies in congenital haemophilia have raised the possibility that inhibitory antibodies vary in their reactivity with the factor VIII molecules in different concentrates used for treatment. However, the interaction of FVIII in concentrates and inhibitors in acquired haemophilia has never been previously studied. In this study, the effect of different FVIII concentrates was studied on neutralization in vitro by performing inhibitor titres using the New Oxford inhibitor assay method. The inhibitor titre in eight patients with acquired haemophilia A was assayed against five commercially available FVIII concentrates of varying purity. The intermediate purity concentrate 8Y and the high purity concentrate that contains normal amounts of von Willebrand's Factor (vWF) (Alphanate) gave lower titres than the high purity concentrates with low (Monoclate-P) or no (Kogenate) von Willebrand content. All but one antibody had very low reactivity with porcine FVIII. Further work will be required to establish whether concentrates manifesting a low level of in vitro reactivity with the inhibitor have a better haemostatic effect in vivo. PMID- 11122272 TI - Monthly haemoptysis in a woman with platelet storage pool disease. AB - A 26-year-old female with known platelet storage pool disease presented with a short history of recurrent haemoptysis. Initial investigations were unhelpful until the cyclical nature of the symptoms became apparent prompting the unusual diagnosis of pulmonary endometriosis to be made. This was subsequently confirmed on premenstrual CT scanning. The introduction of a specific hormonal therapy and multidisciplinary input was ultimately successful. PMID- 11122273 TI - Leukaemic small cell variant anaplastic large cell lymphoma during pregnancy. AB - The history of a 28-year-old woman with anaplastic large cell lymphoma (ALCL) in the first trimester of her pregnancy is reported. Investigations allowed to diagnose a T-cell CD30 positive ALCL, which appearance is rare during pregnancy. Moreover, the atypical lymphoid cells were found in the peripheral blood and were predominantly small to medium sized with nuclear irregularities and cytoplasmic azurophilic granules, which allowed the hypothesis of leukaemic presentation of a small cell variant ALCL. A variant of the t(2:5)(p23:q35) was found [del(2)(p22)]. The patient died shortly after diagnosis. PMID- 11122274 TI - Acute myeloid leukaemia with giant granules: association with t(10; 11)(p13; q14) and disseminated intravascular coagulation. AB - A 16-year-old Chinese girl presented with AML-M5a. A bone marrow examination showed that the myeloblasts which were overwhelming the marrow contained giant granules (pseudo-Chediak-Higashi anomaly). Her karyotype showed a rare translocation t(10; 11)(p13; q14). Molecular delineation of the translocation breakpoints was not possible. Nonetheless, this case further demonstrates the morphological and phenotypic heterogeneity of acute leukaemia with this translocation. In this girl it was associated with disseminated intravascular coagulation. PMID- 11122275 TI - T cell lymphocytosis associated with polymyositis, myasthenia gravis and thymoma. AB - Peripheral T cell lymphocytosis is a rare finding in association with malignant thymomas. In the majority of previous cases, the tumours have behaved aggressively with symptoms arising from local invasion. We describe a patient with ocular myasthenia gravis who presented with a rapidly progressive polymyositis and neuropathy and who was subsequently found to have a thymic mass and a mild T cell lymphocytosis. The thymoma did not give rise to local symptoms and showed no evidence of progression over a 14-month period of follow-up. The possibility of an underlying thymic tumour should be considered in any patient with chronic T cell lymphocytosis if the circulating cells show mature morphology and there is no molecular evidence of monoclonality. PMID- 11122276 TI - Otological Wegeners granulomatosis: a diagnostic dilemma. PMID- 11122277 TI - Chronic stenosing external otitis/postinflammatory acquired atresia: a review. PMID- 11122278 TI - Diagnosis and consequences of gastropharyngeal reflux. PMID- 11122279 TI - Does evidence-based medicine exist in the treatment of Meniere's disease? A critical review of the last decade of publications. AB - Debate surrounding the need for evidence-based research in all fields of otolaryngology has recently emerged. A review of English language articles dealing with the treatment of Meniere's disease published from 1989 to 1999 was performed. The papers were classified according to article type (observational studies, non-clinical studies, controlled trials, randomised controlled trials) and according to the grade of evidence that was offered. A total of 128 publications were reviewed, of which only three were randomised controlled trials and a further three were well-designed controlled studies without randomization. The majority of papers were non-experimental clinical descriptive studies or case series. Data from this review would suggest that more comprehensive evidence based studies are required in order to settle areas of confusion surrounding treatment of this relapsing condition PMID- 11122280 TI - The nasal cycle in health and disease. PMID- 11122281 TI - Systematic errors in bone conduction audiometry. AB - Air and bone conduction audiometry was carried out on two separate groups of 12 normal hearing volunteers. One group was tested at St Mary's hospital, the other at Charing Cross hospital. The results from both centres showed evidence of a systematic error in bone conduction resulting in a pattern of 'notching' at 2 kHz. We argue that the effect is likely to be more apparent in conductive deafness and that a significant distortion of the audiogram occurs in about 17% of such cases. Since the problem appears not to be restricted to the centres involved in the study, we strongly recommend that the issue be addressed by the appropriate professional bodies. PMID- 11122282 TI - Patients with neck lumps: can they be managed in a 'one-stop' clinic setting? AB - The present government has stated its intentions to expand the application of "one-stop" assessments for out-patients. To decide if this is an appropriate strategy for managing patients with neck lumps, we prospectively assessed 110 patients referred to the Neck Lump Clinic of the Otolaryngology Department of a teaching hospital. Patients were assessed clinically and with immediately reported fine needle aspiration cytology (FNAC). The accuracy of immediately reported FNAC was later compared with a final report and histology, when available. A "one-stop" visit was defined as patients who were discharged, or placed on a waiting list, after a single consultation. Eight-three (76%) patients did not have to return to the outpatient department, of which 59 (54%) were discharged. No changes occurred from immediate to final FNAC reports. If certain criteria are met, patients with neck lumps can be successfully managed in a "one stop" setting. PMID- 11122283 TI - Revision rhinoplasty: review of deformities, aetiology and treatment strategies. AB - Rhinoplasty has always been one of the most challenging aspects of facial surgery as it requires a precise assessment of the deformity, a strong grasp of nasal support mechanisms plus soft tissue skin envelope and a realistic appraisal of the outcome expected acutely and over a long period of time. This fine balance is often achieved by retrospective analysis of post-rhinoplasty results in order to highlight repeated problems and improve upon them. This study reviews 110 patients in which a total of 407 deformities were found. These are divided into upper, middle and lower thirds with a subdivision of individual deformities within each group. There is also a comparison of the results obtained from similar studies over a period of four decades to those in our study, which reveals that the post-rhinoplasty deformities noted in these studies occur with a similar frequency to those in our paper. PMID- 11122284 TI - Sinus symptom scores: what is the range in healthy individuals? AB - Patient symptom severity is an important outcome measure in the assessment and staging of chronic rhinosinusitis. Symptom scores based on a visual analogue scale (VAS) are being used increasingly for this purpose although little, if any, information is available on normal values for these scores. We assessed symptom score ratings using the ICSD symptom scores system (maximum score = 60), in a group of 100 healthy individuals who considered themselves free of nasal conditions. Scores from this healthy sample were compared with scores from a hundred patients suffering from chronic rhinosinusitis. The healthy group did not have a uniformly perfect score (mean = 8. 8) and smokers had higher sinus symptom scores than non-smokers (P = 0.02). There was a significant difference in mean values between the healthy sample and the patients with rhinosinusitis (abnormal mean score = 35.3, P = < 0.001). Despite the score difference, a marked overlap existed between the two groups. It should not be assumed that rhinology patients will achieve a zero or near zero score following treatment, but rather their scores should be expected to move closer to the score range for the healthy population. PMID- 11122285 TI - The frequency of otitis media with effusion in British pre-school children: a guide for treatment. ALSPAC Study Team. AB - Data on the prevalence of otitis media with effusion (OME) as shown by serial tympanometry is presented for young children during the first 5 years of life. The children were participants in the Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC); a randomly selected 10% (n = 1400) of whom were selected for examination at ages 8, 12, 18, 25, 31, 37, 43, 49 and 61 months. Whilst sex had no effect, there was a decrease in prevalence of OME with increasing age. There was a marked seasonal effect on the prevalence of OME. Bilateral and unilateral OME were significantly more prevalent in the winter than in the summer months (36.6% in February in children aged 8 months compared with 16% at 61 months and 16.4% in August in children aged 8 months compared with 3.1% at 61 months). The results form an important background against which to assess both the results of screening and also the indications for surgical treatment. PMID- 11122286 TI - The effect of inflammation on spiral ganglion cell density measurements in the cat cochlea. AB - The quantitative analysis of spiral ganglion cells is important in assessing the biological safety of cochlear implants. Quantitative analysis of ganglion cells in a histological section is conventionally expressed as cell density, the number of ganglion cells within Rosenthal's canal being divided by its area. The area of Rosenthal's canal conventionally excludes the area of blood vessels within it. Previous work has shown that the blood vessel area within Rosenthal's canal increases with cochlear inflammation. Consequently, excluding this area may result in an underestimate of ganglion cell loss and a tested implant parameter may be wrongly passed as safe. This study investigates whether the increase in blood vessel area with grade of inflammation has an effect on ganglion cell density measurements made by excluding blood vessel area. Eighteen implanted and stimulated cat cochleae were serially sectioned. Using computer image analysis we measured ganglion cell number, the area of Rosenthal's canal and its blood vessels. A 'blind' histologist graded the inflammation in each cochlea. Ganglion cell densities calculated by excluding and including blood vessel area showed no divergence with increasing inflammation (B = -163, P = 0.001 and B = -160, P = 0.001). The increase in the blood vessel area with inflammation has no effect on cell density measurements made by excluding that area. PMID- 11122287 TI - A randomised clinical trial of turbinectomy for compensatory turbinate hypertrophy in patients with anterior septal deviations. AB - Turbinectomy is performed at the time of nasal septal surgery by many otolaryngologists. One reason given for this procedure is the presence of a hypertrophied contralateral inferior turbinate. A randomised trial was undertaken to evaluate the relief of nasal obstruction following contralateral turbinectomy with septal surgery. Patients presenting with nasal obstruction who had a unilateral septal deviation and contralateral inferior turbinate enlargement were prospectively randomized to contralateral turbinectomy or no turbinate surgery at the time of septal surgery. Questionnaires and active anterior rhinomanometry were used for evaluation. Twenty-six patients (mean age 31 years) demonstrated a reduction in subjective and objective measures of nasal obstruction (P < 0.05) 8 weeks after operation. There was no intergroup difference, the median total decongested nasal resistance postoperatively in the non-turbinectomized patients was 0.17 kPal-1 s and 0.21 kPal-1 s in the turbinectomized patients. Contralateral inferior turbinectomy does not add to the relief of nasal obstruction beyond that attained by septal surgery in these patients. PMID- 11122288 TI - Reliable and reproducible anterior active rhinomanometry for the assessment of unilateral nasal resistance. AB - Unilateral nasal resistance is now thought to be more important than total resistance in promoting obstructive symptoms. We assessed the reproducibility of anterior active rhinomanometry in measuring unilateral nasal resistance. Ten baseline readings of unilateral nasal resistance were made over a 30-min period in seven healthy subjects (14 nostrils) using anterior active rhinomanometry performed according to the International Committee for the Standardization of Rhinomanometry (ICSR) guidelines. Baseline readings revealed that measurements using anterior active rhinomanometry had an unacceptably high coefficient of variation (19%-60%). With a more time-consuming revised protocol involving multiple recordings and the identification and exclusion of erroneous data, coefficients of variation of 7%-15% were obtained. We conclude that single anterior active rhinomanometry readings are potentially prone to large errors and each researcher using such equipment must satisfy his/herself that their methodology has an acceptable coefficient of variation in their hands. The ICSR guidelines are not always sufficient to allow reproducible measurement and specially designed protocols may be necessary to produce reliable results. PMID- 11122289 TI - Comparison between videofluoroscopy and milk-swallow endoscopy in the assessment of swallowing function. AB - The investigation of swallowing function and the detection of aspiration has evolved over the past 10 years with the introduction of the fibreoptic endoscopic assessment. Practical reasons can limit the use of videofluoroscopy in some patients. A prospective study of 20 sets of videofluoroscopic and endoscopic assessments of swallowing function were compared. Endoscopic assessment was found to be a highly sensitive and specific method of determining swallowing safety. Reduced or absent laryngeal sensation at endoscopy correlates with silent aspiration and thus a high risk of aspiration pneumonia. PMID- 11122290 TI - Changes in snoring during natural sleep identified by acoustic crest factor analysis at different times of night. AB - Sleep nasendoscopy can be used to identify the site of snoring but questions remain about how well a short assessment during drug-induced sleep reflects the natural condition. To investigate the uniformity of snoring during natural sleep we studied five patients (three men, two women) referred by their GPs for treatment of their snoring. A digital audio tape recorder captured the free-field snore sound at different times of night in hospital. Acoustic Crest Factor values were calculated on the 15 recordings made, having previously demonstrated that high crest factor values distinguish palatal from non-palatal snoring at sleep nasendoscopy. Some recordings showed reproducibility, but others showed substantial changes between recordings an hour apart. We infer that the snoring mechanism may change in some individuals during the night, with or without a change of snore site. We conclude a single recording, as in sleep nasendoscopy, may not be representative. PMID- 11122291 TI - A retrospective case-controlled study of 1490 consecutive patients presenting to a neuro-otology clinic to examine the relationship between blood lipid levels and sensorineural hearing loss. AB - This study set out to test the null hypothesis that the distribution of blood lipid levels is the same in a population with a sensorineural hearing loss (SNHL) as in a control population. Hyperlipidaemia has been implicated as an aetiological factor in SNHL; however, the majority of reports are retrospective, lack adequate controls, or are based on a series of cases which may represent incidental findings and not a true causal relationship. In all, 1490 consecutive patients who presented to a neuro-otology clinic were studied retrospectively. This group is exceptional in that all patients had had fasting lipid profiles done regardless of their presenting problem. Those with a mean SNHL > 25 dB (0.5, 1, 2 and 4 kHz) were compared with those with hearing thresholds < or = 25 dB. An analysis of variance was also done. The study group was also compared with the National Study of Hearing data set to add external validity. Simple correlations were found between hearing thresholds and many parameters such as blood pressure, fasting glucose, triglyceride or fasting cholesterol. Analysis of variance of the neuro-otology group, controlling for variables such as age and sex, showed no significant association between hearing and blood pressure, packed cell volume, mean corpuscular volume, fasting glucose, triglyceride or fasting cholesterol. However, when general linear interactive modelling was used to analyse hearing thresholds, raised total fasting cholesterol was associated with significantly better hearing threshold levels. This study leads to a rejection of the Null hypothesis that the distribution of lipid levels is the same in a population with a hearing loss as in a control population, as hearing thresholds were found to be significantly better in those with raised cholesterol levels. PMID- 11122292 TI - Origin and spread of allergic fungal disease of the nose and paranasal sinuses. AB - Although expansion of bony walls occurs in allergic fungal disease of the nose and paranasal sinuses by increased mucus secretion and fungal growth, the latter is apparently confined to the lumen and does not invade the tissues. Nevertheless, spread of the disease process from paranasal sinuses to orbit, cheek and intracranial cavity is well described. An imaging and histopathological study was carried out in 16 cases to determine how the disease originates and spreads. The infection starts in the nasal cavity, the lumen of a sinus or in a seromucinous gland or duct. A thin vascular zone of intense allergic inflammation surrounds the infected mucin. Erosion of bone takes place focally, probably by substances produced by the inflammatory tissue, allowing intromission by the thin vascular layer together with its underlying fungus-containing mucus and so extension of the disease process through the eroded bone. PMID- 11122293 TI - Residual mesenchyme in the middle ear cavity of the newborn with malformation of the kidney. AB - Microscopic sections of 37 temporal bones from 22 foetuses were examined, five with renal agenesis, six with Potter's sequence without renal agenesis and two with mild kidney pathology. Every tenth section containing the stapes were used for measuring the volume of mesenchymal tissue in the middle ear cavity. At 5% significance level, foetuses with serious renal pathology had more mesenchymal tissue in the middle ear cavity than a control group of nine foetuses with no renal or urinary tract abnormality. PMID- 11122294 TI - Reliability of electrogustometry for the estimation of taste thresholds. AB - Electrogustometry has been used as a clinical tool for diagnosis and assessment of a variety of conditions. Several studies which have estimated electrogustometric taste threshold have reported good test-retest reliability but there is some evidence that thresholds measured this way may decrease with practice. In this study, repeated testing of the right and left sides of the tongue of two subjects over 80 sessions resulted in considerable session-to session variability for both subjects, and three of the four sets of threshold estimates decreased systematically with continued testing. Two of the four data sets continued to show systematic decreases when the first 10 sessions were omitted from analysis to allow for any initial unfamiliarity with the testing procedure. This suggests that estimates based on few tests should be interpreted with caution, particularly when used for evaluation or monitoring of interventions. PMID- 11122295 TI - Identification of beta-actin as a candidate autoantigen in autoimmune inner ear disease. AB - It has been shown that sera from patients with autoimmune inner ear disease contain antibodies to several inner ear antigens. We report here the characterization of the 42-43 kDa protein against which a significant number of patients' sera react strongly. After separation of inner ear proteins from guinea pig cochleas by SDS-PAGE, the band corresponding to the 42-43 kDa protein was digested with trypsin and the peptide fragments were separated by high performance liquid chromatography. Two fractions were then subjected to amino acid sequencing by the classical automated Edman degradation. The sequence of a stretch of 15 amino acids of the first fragment was identical to that of amino acids 148-162 of beta-actin. The sequence of the 10 amino acids of the second fragment was also identical to beta-actin. On Western blots, monoclonal antibody directed against beta-actin reacted with the inner ear 42-43 kDa proteins. The serum samples from the patients and the monoclonal antibody reacted with the non muscle actin used as antigen in Western blotting. Immunoblot analysis of inner ear proteins after two-dimensional gel electrophoresis showed a spot, corresponding to the region of the 43 kDa as compared to the protein standards. On the basis of these data it is concluded that the target 42-43 kDa protein for antibodies in sera of patients with autoimmune inner ear disease is beta-actin, a molecule, which has important and numerous functions inside cells. This is the first report to identify the cytoskeletal protein beta-actin as a candidate autoantigen in autoimmune inner ear disease. PMID- 11122296 TI - Audit of stapedectomy in the north west of England for 1996 and an analysis of the criteria used to describe success. AB - A questionnaire was used to collect information related to stapedectomy from all the NHS Trusts in the North West region. The data collected included the identity and rank of the surgeon, the age and sex of the patients, the side of the operation, the audiometric values of the air conduction audiogram preoperatively and postoperatively, and an account of any complications. The results were analysed and compared to national and international standards. The criteria for analysis of success in surgery for conductive hearing loss were reviewed. PMID- 11122297 TI - Is oestrogen therapy justified in the treatment of hereditary haemorrhagic telangiectasia: a biochemical evaluation. AB - Systemic and topical oestrogen can provoke squamous metaplasia of epithelium. In Hereditary Haemorrhagic Telangiectasia (HHT) the underlying telangiectasia may be protected from trauma and epistaxis reduced. Oestrogens have been advocated but their efficacy is unclear. Recent advances have now identified two oestrogen and one progesterone receptors. The aim of this study is to analyse the sex receptor status of HHT nasal mucosa to determine if oestrogen therapy is biochemically justified. Five HHT patients (three men, two women) and eight controls (four men, four women) underwent nasal mucosa biopsy. Samples were fixed in formalin and paraffin embedded. Alpha oestrogen (ERalpha) and beta oestrogen (ERss) and progesterone (PgR) receptors were identified using mouse monoclonal antibodies by the Streptavidin-biotin peroxidase method. ERss was detected in two HHT subjects (1 M: 1F) and two control subjects. ERalpha and PgR was absent in HHT subjects. This pilot study demonstrated that a subgroup of HHT patients were ERss positive. Oestrogen therapy therefore has a potential therapeutic role on a biochemical basis in these patients. ERss status should be determined before considering oestrogen therapy. PMID- 11122298 TI - Patient and physician perspectives on the impact and management of perennial and seasonal allergic rhinitis. AB - Patient and physician perspectives on the impact and management of perennial (PAR) and seasonal allergic rhinitis (SAR) were studied. In all, 2139 subjects were questioned about their medical conditions, severity and frequency of symptoms and satisfaction with treatment. A group of general practitioners (GPs) were also invited to discuss their experiences in the management of rhinitis. In this UK survey, allergic rhinitis was more common than asthma, hypertension, skin rashes, eczema and diabetes. The prevalence of SAR and PAR was 15% and 2%, respectively. Sneezing and runny nose were the most common symptoms and GPs were the main contact for advice and treatment (54% of patients). Symptoms were well controlled in 32% of patients. Allergic rhinitis affected work, home and social life in 29%, 34% and 30% of patients, respectively. The GPs considered PAR to be more difficult to treat than SAR, and GP and patient level of satisfaction in the treatment of PAR was low. This suggests that education of patients and physicians on the benefits of allergen avoidance, and the selective use of the highly effective therapies available on prescription could improve the level of satisfaction with therapy. Adherence to current guidelines on the management of rhinitis could lead to an effective, structured treatment plan for patients. PMID- 11122299 TI - The effect of hypertonicity on nasal mucociliary clearance. AB - The effect of the tonicity of saline nasal douching solutions on mucociliary clearance was studied in order to ascertain whether hypertonicity conferred any advantage. Thirty-eight normal subjects were included in a randomised double blind crossover trial. Saline douching solutions of 0.9%, 3% and 5% tonicity were used and mucociliary clearance was measured by the saccharin clearance time (SCT). The resultant SCT after administration of 5% saline was significantly reduced compared to both 0.9% (P = 0.005) and 3% saline (P = 0.04). There was no difference between 0.9% and 3% saline administration. Thus hypertonic saline solutions improve mucociliary clearance, although this was only observed with solutions of 5% tonicity. The effect is probably brought about by changes in mucus rheology. PMID- 11122300 TI - A clinical study of vestibular schwannomas in type 2 neurofibromatosis. AB - All 13 patients with neurofibromatosis 2 (NF2) who presented over a period of 17 years at the Institute of Neurological Sciences, Glasgow were reviewed and compared to patients with sporadic vestibular schwannomas. The NF2 patients presented at a younger age than those with sporadic vestibular schwannomas. A significant number had normal pure tone audiograms and a small number also had normal auditory brainstem responses at presentation. Vestibular schwannomas in NF2 patients grow more often and more rapidly than sporadic unilateral ones. They are more liable to infiltrate the cochlear and facial nerves making hearing and facial nerve preservation more difficult to achieve. Because the relatives of these patients often have normal audiograms and normal auditory brain stem responses in the presence of a schwannoma, our recommended method of screening of relatives of NF2 patients is magnetic resonance image scanning with Gd-DTPA enhancement. PMID- 11122302 TI - Thyroid stimulating hormone increases angiogenic growth factor expression in rat thyrocytes AB - INTRODUCTION: Goitrogenesis is thought to be due to hyperstimulation of the thyroid stimulating hormone (TSH) receptor and is associated with dramatic andiogenesis within the thyroid. The angiogenic growth factors, fibroblast growth factor (FGF-2) and its receptor (FGFR-1) as well as Tie-2 (the receptor for the angiopoietins) are elevated in both the human goitre and in rat goitre. We have investigated the role of TSH in the expression of FGF-2, FGFR-1 and Tie-2 in FRTL 5 cells, which are a functional rat thyroid cell line. METHODS: Cells were cultured in medium containing 5% serum, insulin and antibodies and exposed to a variety of experimental conditions. Proteins were separated by electrophoresis and analysed by Western blotting using polyclonal antibodies to FGFR-1 (extracellular domain, Santa Cruz), Tie-2 (Santa Cruz) and FGF-2 (Sigma). RESULTS: TSH increased expression of FGF-2 reaching a maximum at 0.3 mU/ml returning to control levels at higher concentrations. A dominant fragment of its receptor (FGFR-1) at >> 57 kDa was dose-dependently increased by TSH. Tie-2 was thought to be predominantly expressed on endothelial cells but we now show that it is expressed as a full-length receptor on FRTL-5 cells with maximal expression at 0.3 mU/ml. This regulation is mediated by cAMP as forskolin and 8-Bromo-cAMP also increased Tie-2 expression. CONCLUSION: Both FGF-2 and its receptor, as well as Tie-2 are increased in rat thyroid cells by elevated TSH. These mechanisms may be central to the angiogenesis that occurs in goitrogenesis, and alteration of these growth factors may lead to new treatments for goitre. PMID- 11122301 TI - Globus pharyngeus: a postal questionnaire survey of UK ENT consultants. AB - Globus pharyngeus is a common complaint often referred to the ENT outpatient department. The precise nature of globus pharyngeus and its aetiology remains something of a mystery. There is no uniform policy of management of this condition. A postal questionnaire was sent to all UK-based ENT consultants registered with the British Association of Otorhinolaryngolgists-Head and Neck Surgeons (BAO-HNS). The aim of this study was to ascertain if there was a favoured management policy by the majority of consultants. Our results indicate that there is a lack of consensus in the investigation and management of globus pharyngeus. Fourteen per cent do not perform any investigations, but would prescribe antacid medication if clinically indicated. The remainder would investigate in a variety of ways. The most common investigation is rigid endoscopy which is performed by 61% of respondents, followed by barium swallow (56%). The combination of endoscopy and barium swallow is routinely performed by 17.5% of respondents. PMID- 11122303 TI - The quality of life impact of snoring and the effect of laser palatoplasty AB - INTRODUCTION: Snoring is a common cause of marital disharmony and social embarrassment. Obstructive sleep apnoea (OSA) have further impact on Quality of Life (QoL). AIMS: First, to compare the impact of snoring and OSA on QoL; second, to assess the impact of laser palatoplasty (LAUP) on QoL in snorers. METHODS: We conducted a prospective cohort comparison of 191 snorers (mean age 46 years; 132 men, 59 women), 57 patients with OSA (mean age 47 years; 49 men, 8 women), and 105 patients, at a mean of 12 months (range, 4-24 months) after LAUP (mean age 45 years; 82 men, 23 women). All completed the Nottingham Health Profile (NHP) and the results were compared with established NHP population norms. RESULTS: The results are shown in Table 1. CONCLUSIONS: This is the largest QoL study of snorers to date and shows that both snoring and OSA have clear impacts on all six NHP domains. The magnitude of the impact in snoring approaches that for OSA for most parameters. The impact of OSA on the sleep domain in men is significantly higher than that of snoring. Energy and emotional reaction domains are significantly improved by LAUP in both sexes, to levels approaching those in the normal population. Also pain, sleep, social isolation and mobility in habitual snorers was helped by surgery. The NHP generic QoL health status measure is a useful tool for the assessment of sleep disorders. PMID- 11122304 TI - Differing expression of bax and bcl-3 may influence the different cure rates in mouth and orophayrngeal cancer AB - AIM: To determine the reason for the differing cure rates of cancers between the mouth and oropharynx. MATERIALS AND METHODS: To achieve our aim we carried out immunohistochemistry on paraffin-embedded tissue from 31 oral and 38 oropharyngeal squamous carcinomas using the streptavidin-biotin/HRP method. RESULTS: The results from this study revealed the mean percentage of bax and bcl 2 expression was higher in the oropharyngeal samples (56.34% and 97.36%, respectively) than in the oral samples (6.6% and 82.38%, respectively). In the oral carcinomas bcl-2 was expressed in a higher percentage of cells than bax, and a high proportion of cases were bcl-2 positive in the absence of significant bax expression. In contrast the oropharyngeal cases showed a high percentage of cells expresssing both bcl-2 and bax. CONCLUSION: The predominance of anti-apoptotic bcl-2 over the pro-apoptotic bax may result in a survival advantage of the cells of oral carcinomas over the cells of the oropharynx. Tumours of the oral cavity have been found to be clinically more radioresistant than those of the oropharynx. These findings may have implications for radiotherapy treatment since the cells of oral carcinomas may already have a survival advantage and the carcinomas may be less likely to respond to treatment. PMID- 11122305 TI - A prospective retrieval study to determine how speaking valve failure is effected by colonization AB - INTRODUCTION: It has been suggested that Groningen Low Resistance (GLR) valve failure is associated with biofouling of the valve's oesophageal surface and hinge areas. However, the valve edges are responsible for efficient valve function.1 Therefore, valve edge colonization should be the most important factor determining valve failure. The null hypothesis that valve edge colonization was not associated with failure was tested using 106 GLR valves retrieved, after failure, from 41 patients. METHODS: The opening pressures, reverse opening pressures and forward resistances of the new valves were determined using apparatus validated previously.2 The pressure/flow parameters were measured again after removal and the changes calculated. The degree of colonization of each valve edge, oesophageal surface, hinge area, tracheal surface and valve lumen was scored using 100-mm linear analogue scales. The changes, in pressure/flow parameters were examined for associated with colonization of the five areas described above. RESULTS: The increase in the opening pressure and resistance, and decrease in reverse opening pressure, of the retrieved valves was significant compred with new valves. The increase in opening pressure was associated with colonization of the valve edge (rs = 0.262, P = 0. 007). The decrease in reverse opening pressure was associated with colonization of the valve edge, hinge areas and oesophageal surface (rs = 0.266, P = 0.006; rs = 0.271, P = 0.005; rs = 0.271, P = 0.004, respectively). The increase in resistance was associated with colonization in all areas (rs >/= 0.367, P = 0.0005). CONCLUSION: This study demonstrated that colonization of the valve edge is associated significantly with the changes, in pressure/flow parameters of failed valves. PMID- 11122306 TI - Are p53 and MIB-1 overexpressed in cholesteatoma? AB - INTRODUCTION: There is controversy regarding the expression of p53 and MIB-1 in cholesteatoma.1 This study was instituted to study this. p53 is an intracellular protein which plays a critical role in control of the cell cycle at the G1 check point. MIB-1 is recognized as a marker of cellular proliferation. Using deep meatal skin controls, the question addressed was, 'are p52 and MIB-1 overexpressed in cholesteatoma'? METHOD: Immunocytochemistry using the avidin biotin technique on frozen tissue sections with primary antibody to p53 (n = 17, controls = 17) and MIB (n = 17, controls = 7) was performed on cholesteatoma and deep meatal skin control specimens. Appropriate positive and negative controls were employed for each antibody. Slides were analysed in a blind fashion by two independent observers. Statistical analysis was performed using the Mann-Whitney test. RESULTS: Expression of p53 was minimal or absent in both cholesteatoma and controls. No significant difference in p53 expression was found between the two groups (P = 0.02). MIB-1 expression was higher in cholesteatoma than in controls, although this was not statistically significant (P = 0.09). CONCLUSION: This study did not demonstrate an increased expression of p53 cholesteatoma, indicating no evidence of a dysfunctional cell cycle. MIB-1 results showed a possible trend towards significance, which requires a more powerful study to evaluate this further. The probability that there is an increased rate of cellular proliferation in cholesteatoma, when compared to deep meatal skin controls, cannot be excluded. PMID- 11122307 TI - Does the bacterial DNA found in middle ear effusions come from viable bacteria? AB - INTRODUCTION: Between 20% and 50% of middle ear effusions in otitis media with effusion test positive for bacteria, i.e. H. influenzae, M. catarrhalis and S. pneumoniae. In one study 48% of effusions that tested negative by culture wre positive by polymerase chain reaction (PCR).1 This has been advanced as evidence for the presence of bacterial biofilms, as agents leading to the persistence of glue ear. There is, however, the possibility that the DNA detected is the fossilized remains of bacteria from previously cleared infections. If this is the case, then the effusioin must in some way be protecting the DNA from breakdown by DNases. METHODS: Here we demonstrate, using a viscosity assay, that middle ear effusions taken from children during myringotomy inhibit the breakdown of DNA in a concentration-dependent manner. Middle ear effusion homogenates 0.5-3.0 ml (1 : 10 Vol. vol/effusion: PBS) were incubated with 3 ml of DNA (0.5 mg/ml in PBS) plus 0.2 ml of DNase 1 (500 Kunitz) at 37 degrees C for 24 h. Viscosity measurements were taken at regular intervals and the changes in viscosity expressed as a percentage of time 0. DNase activity was inhibited by 1.5 ml and 3.0 ml of effusion 48% and 91%, respectively, after 30 min incubation. CONCLUSION: This study demonstrated that effusions have the ability to inhibit nuclease activity, and the reported presence of non-culturable bacteria based on DNA detection by PCR could still represent fossilized remains and not viable bacteria. The same could also be true of recent reports of bacterial mRNA.2 PMID- 11122308 TI - A glance at medical reading habits in Italy and England: is the writing on the wall? PMID- 11122309 TI - Effects of propofol on substance P-induced relaxation in isolated human omental arteries and veins. AB - To elucidate if an effect of propofol on endothelium-dependent relaxation could contribute to propofol-induced vasodilation, smooth muscle relaxation of isolated human omental artery and vein segments precontracted by endothelin-1 were measured. Substance P induced a concentration-dependent relaxation (mean +/- SEM) in both artery (63 +/-8.4% of precontraction, n = 9) and vein (60+/-11%, n = 7). The relaxation was enhanced by 10(-6) M propofol (artery, 72+/-9.5%, n = 9; vein, 81+/-12%, n = 7) but not affected by 10(-7), 10(-5) and 10(-4) M propofol. In the presence of Nomega-nitro-L-arginine methyl ester (nitric oxide synthase inhibitor), 10(-6) M propofol still enhanced the substance P-induced relaxation in arteries but not veins, whereas 10(-4) M propofol inhibited the relaxation in both arteries (rightward shift of the concentration-response curve) and veins (28+/-7.5%, n = 8). In the presence of potassium chloride (to prevent hyperpolarization), the enhancement of substance P-induced relaxation by 10(-6) M propofol was abolished in both arteries and veins whereas 10(-5) and 10(-4) M propofol reduced the relaxation in arteries (38+/-13% at 10(-5) M, n = 6; 30+/ 11% at 10(-4) M, n = 6) but not in veins. These results demonstrate that propofol, at lower, clinically relevant concentrations, promotes endothelium dependent relaxation mediated via hyperpolarization in human omental arteries and via both nitric oxide and hyperpolarization in human omental veins. PMID- 11122310 TI - Are diabetic patients in danger at renal transplantation? An invasive perioperative study. AB - It is assumed that diabetic patients with uraemia have more complications at renal transplantation than those who are not diabetic. We compared the preoperative ECGs, and invasive perioperative haemodynamic and oxygenation parameters in 15 diabetic and 15 non-diabetic uraemic patients undergoing renal transplantation. The number of patients with increased QT dispersion in the ECG was higher in diabetic than in non-diabetic patients (P<0.05). Before anaesthesia, heart rate and mean arterial pressure were higher (P<0.05) in the diabetic than in the non-diabetic group. After preanaesthetic volume loading all patients showed a hyperdynamic circulation, which subsided during anaesthesia. However, stroke volume index remained unchanged. Four patients in the diabetic group and six in the non-diabetic group needed additional oxygen therapy after surgery. No cardiac dysrhythmias were noted. However, the increased QT dispersion in diabetic patients calls for an adequate perioperative ECG monitoring for dysrhythmias. The diabetic and non-diabetic uraemic patients performed equally well at renal transplantation. In conclusion, renal transplantation for diabetics is justified. PMID- 11122311 TI - Evaluation of the VersaMed portable ventilator: clinical trials. AB - The VersaMed 201 is a lightweight, computer-controlled, electrically driven ventilator, designed for transport, home-ventilation and field use. We have evaluated the device in human beings with healthy and pathological lungs. In the first part of the study, the VersaMed was substituted for a standard operating room ventilator for 15-30 min during elective surgery in 20 subjects. In the second stage of the study, 20 patients with lung pathology who were being ventilated in the critical care department were transferred to the VersaMed for a 30-min period. In all the patients studied, arterial blood gases, haemodynamic parameters and other respiratory variables were maintained at similar values seen with the standard ventilators used. We conclude that the VersaMed 201 is suitable for use in human beings with healthy and pathological lungs as a reliable, portable ventilator. PMID- 11122312 TI - Haemodynamic responses to the intubating laryngeal mask and timing of removal. AB - We determined (a) the haemodynamic responses to intubating laryngeal mask (ILM) airway insertion/intubation and removal in anaesthetized patients, and (b) whether the timing of ILM removal influences these responses. One-hundred and twenty patients without cardiovascular disease were studied. ILM airway insertion/intubation was 5 min after induction with propofol 2 mg kg(-1) and maintenance of anaesthesia with sevoflurane 2% in oxygen 33% and nitrous oxide. Patients were randomly assigned for removal of the intubating laryngeal mask airway at 1, 3 and 5 min after successful intubation. Systolic and diastolic arterial pressures and heart rate were recorded preinduction (baseline), before ILM airway insertion/intubation, at 1-min intervals after insertion/intubation, and at 1-min intervals for 5 min after ILM removal. ILM insertion was successful at the first attempt in all patients, but 46 patients required more than one intubation attempt. Compared with baseline values, there were no increases in systolic or diastolic arterial pressure, but there was an increase in heart rate 1 min after ILM insertion/intubation (9%, P<0.001) and 1 min after ILM removal (8%, P<0.01). There was a significant increase in systolic and diastolic pressures and heart rate 1 min after ILM insertion/intubation (30%, 31% and 15%; all: P<0.002) compared with before ILM insertion/intubation values and 1 min after ILM removal (9%, 8% and 7%; all P<0.05) compared with 1 min after ILM insertion/intubation values. Removal of the ILM 1 min after successful intubation resulted in higher arterial pressure compared with removal at 3 min (systolic arterial pressure 10% higher for 1 min, P = 0.01) and 5 min (systolic arterial pressure 10-23% higher for 3 min, P<0.01; diastolic arterial pressure 10-20% higher for 4 min, P>0.02), but there were no differences in heart rate between groups. Systolic and diastolic arterial pressures were greater if more than one intubation attempt was required. Early removal or multiple intubation attempts did not exceed baseline haemodynamic values. We conclude that ILM insertion/intubation and removal in anaesthetized patients produces little or no haemodynamic response, even if multiple intubation attempts are required. The timing of removal exerts a small, but clinically unimportant influence on these responses. PMID- 11122313 TI - Clinical letter: epidural analgesia in a newborn with Hirschsprung's disease, associated with congenital central hypoventilation syndrome. AB - A case is presented of a neonate with Hirschsprung's disease, associated with congenital central hypoventilation syndrome. After an ileostomy (at 2 days) and a stoma revision (at 10 days), postoperative pain management was established by continuous intravenous infusion of morphine, which caused severe postoperative respiratory depression. At 6 weeks a re-exploration and stoma revision was performed using postoperative epidural analgesia with bupivacaine. This caused no respiratory depression. A colectomy under epidural analgesia at 8 months was also uneventful. Respiratory difficulties in children with congenital central hypoventilation syndrome associated with Hirschsprung's disease are discussed in relation to the technique of choice for postoperative pain management. PMID- 11122314 TI - Effects of a pneumoperitoneum in the obese patient. PMID- 11122315 TI - A reply PMID- 11122316 TI - The space mission MIR'97: operational aspects. AB - BACKGROUND: A German astronaut visited the MIR space station between 10 February and 2 March 1997. Together with his Russian colleagues, he conducted a series of scientific investigations before, during and after his stay aboard the MIR station. Research performed during this flight was part of a global space life sciences programme and focused on metabolic homeostasis, fluid balance, calcium homeostasis and cardiovascular regulatory mechanisms. The main goal of the scientific experiments was to use this mission as a milestone to establish international networks of scientific collaboration using space research as a tool for focused research in respective fields. Thus, in most cases the results obtained from the astronaut complemented a series of results obtained on ground and from other flights. In other cases, they extended previous results and opened new fields for future research. PARTICIPANTS: Human space flight with astronauts serving as operators and at the same time as test subjects is very complex. Many people, including mission control, a science management team, medical operations, ethics committees and a medical board, participated to harmonize the different requirements, thus making a maximal scientific outcome possible. CONCLUSION: In summary, this space mission may be seen as a model for focused long-term multidisciplinary international research, and demonstrates that space medicine is no longer adventure but science. PMID- 11122317 TI - Investigations of humans in space: adequate from a scientific point of view? PMID- 11122318 TI - Microgravity inhibits intestinal calcium absorption as shown by a stable strontium test. AB - BACKGROUND: Little is known about the onset and degree of biochemical and functional alterations in calcium metabolism during microgravity. OBJECTIVE: To evaluate the effect of microgravity on intestinal calcium absorption and calcium regulating hormones under metabolic ward conditions. MATERIALS AND METHODS: Fractional calcium absorption (Fc240 in percentage of dose administered) was determined pre-flight, in-flight and post-flight, by use of a stable strontium test in one cosmonaut who spent 20 days in space. Moreover, a sequence of blood samples was collected for the determination of serum parathyroid hormone (PTH), 25-hydroxyvitamin D, calcitriol and serum C-telopeptide (CTx, biomarker of bone resorption) levels. During all periods of data collection, calcium intake was held constant at a minimum level of 1.000 mg day(-1) and a daily supplement of 16.6 microg vitamin D2 was given. Personal ultraviolet (UV) light exposure was measured during the whole mission using a biologically weighting UV dosimeter. RESULTS: Fc240 was markedly reduced on flight day 19 (4.4%) as compared to pre flight and post-flight data (13.4% and 17.2%, respectively). Serum calcitriol levels fell from 40.6 pg mL(-1) (mean pre-flight level) to 1.3 pg mL(-1) on flight day 18 and returned into the normal range after recovery. Serum CTx increased during the flight, while serum PTH and 25-hydroxyvitamin D levels did not change significantly. CONCLUSIONS: Intestinal calcium absorption can be diminished after only three weeks of microgravity. Changes are associated with a severe suppression of circulating calcitriol levels, but are independent of exogenous vitamin D supply and serum PTH levels. PMID- 11122319 TI - Investigation of bone changes in microgravity during long and short duration space flight: comparison of techniques. AB - BACKGROUND: Loss of bone mass is a continuing problem in long-term space flight. Although counter-measure programmes have been developed, effective assessment of these programmes is hampered by a lack of monitoring techniques that can be used in-flight. MATERIALS AND METHODS: Three techniques were used to evaluate changes in bone during two missions of 180 and 20 days to the MIR space station, involving three subjects. Dual energy X-ray absorptiometry (DXA) was used before and after flight to measure whole body and regional bone mineral density (BMD). Ultrasonic measurements of velocity (SOS) and broadband attenuation (BUA) of the calcaneus were measured during the 180 day mission and before and after the 20 day mission. Phase velocity of flexural waves in the tibia was also measured on the same days as the ultrasonic measurements of the calcaneus. RESULTS: DXA measurements demonstrated significant variation between different sites in the body for changes in BMD, with the greatest changes occurring in the lumbar spine and proximal femur. There was a trend for increasing phase velocity in the tibia during the 180 mission, but this was not significant. BUA and SOS measurements of the calcaneus showed consistent but divergent patterns of changes during the mission. CONCLUSION: Although in-flight measurements of bone using ultrasound or phase velocity may provide information on the kinetics of bone loss in space flight, the heterogeneity of response in the skeleton means that it is difficult to predict overall bone loss from measurements at one particular site. PMID- 11122320 TI - Cardiovascular response to lower body negative pressure stimulation before, during, and after space flight. AB - BACKGROUND: It is well known that space travel cause post-flight orthostatic hypotension and it was assumed that autonomic cardiovascular control deteriorates in space. Lower body negative pressure (LBNP) was used to assess autonomic function of the cardiovascular system. METHODS: LBNP tests were performed on six crew-members before and on the first days post-flight in a series of three space missions. Additionally, two of the subjects performed LBNP tests in-flight. LBNP mimics fluid distribution of upright posture in a gravity independent way. It causes an artificial sequestration of blood, reduces preload, and filtrates plasma into the lower part of the body. Fluid distribution was assessed by bioelectrical impedance and anthropometric measurements. RESULTS: Heart rate, blood pressure, and total peripheral resistance increased significantly during LBNP experiments in-flight. The decrease in stroke volume, the increased pooling of blood, and the increased filtration of plasma into the lower limbs during LBNP indicated that a plasma volume reduction and a deficit of the interstitial volume of lower limbs rather than a change in cardiovascular control was responsible for the in-flight response. Post-flight LBNP showed no signs of cardiovascular deterioration. The still more pronounced haemodynamic changes during LBNP reflected the expected behaviour of cardiovascular control faced with less intravascular volume. In-flight, the status of an intra-and extravascular fluid deficit increases sympathetic activity, the release of vasoactive substances and consequently blood pressure. Post-flight, blood pressure decreases significantly below pre-flight values after restoration of volume deficits. CONCLUSION: We conclude that the cardiovascular changes in-flight are a consequence of a fluid deficit rather than a consequence of changes in autonomic signal processing. PMID- 11122321 TI - Water and sodium balances and their relation to body mass changes in microgravity. AB - BACKGROUND: Since the very beginning of space physiology research, the deficit in body mass that is often observed after landing has always been interpreted as an indication of the absolute fluid loss early during space missions. However, in contrast to central hypervolemic conditions on Earth, the acute shift of blood volume from the legs to the upper part of the body in astronauts entering microgravity (microG) has neither stimulated diuresis and natriuresis nor resulted in negative water-and sodium-balances. DESIGN: We therefore examined the kinetics of body mass changes in astronauts (n = 3) during their several weeks aboard the space station MIR. A continuous diet monitoring was performed during the first mission (EuroMIR94, 30 days). The second mission (MIR97, 19 days) comprised a 15-day metabolic ward period (including predefined constant energy and sodium intake). Water and sodium balances were calculated and the kinetic of changes in basal concentrations of fluid-balance-related hormones during flight were determined. CONCLUSION: The data suggest firstly that loss of body mass during space flight is rather a consequence of hypocaloric nutrition. Secondly, microG provokes a sodium retaining hormonal status and may lead to sodium storage without an accompanying fluid retention. PMID- 11122322 TI - Angiotensin II type-I receptor and ACE polymorphisms and risk of myocardial infarction in men and women. AB - BACKGROUND: Findings relating an association between an insertion/deletion polymorphism of the angiotensin converting enzyme (ACE) gene and myocardial infarction (MI) have been mixed. While other loci, such as the angiotensin II type-I receptor (AT1R), may modulate risk, few studies have adequately documented the risk in women. We aimed to study whether the findings in respect of ACE and AT1R in UK men were borne out in women. METHODS: Cases of MI (305 women, 391 men) in Belfast and Glasgow have been compared to controls (291 women, 356 men). These new samples augment the original men (200 cases, 181 controls) included from Belfast in the ECTIM study. RESULTS: Among men, the odds ratio for MI for ACE (DD vs. ID + II) was 1.03 (0.79, 1.34) and among women, 0.69 (0.47, 1.01). This heterogeneity between the risks in men and women was significant in Glasgow (P = 0.02). Among men and women the odds ratio for MI for AT1R (CC vs. AC + AA) was 1.02 (0.71, 1.47). There was a small gradient in risk, such that the odds ratio for DD genotype was 0.86 (0.63, 1.17) among subjects homozygous for the common AT1R allele (AA): 0.94 (0.67, 1.30) among heterozygotes and 1.21 (0.53, 2.77) among CC subjects; but this interaction was not statistically significant. CONCLUSIONS: Some of the contradictory findings concerning the ACE polymorphism and the risk of MI may be due to heterogeneity in the risk between men and women. The AT1R1196 polymorphism is not an independent risk factor for MI in either sex. PMID- 11122323 TI - Role of homocysteine, cystathionine and methylmalonic acid measurement for diagnosis of vitamin deficiency in high-aged subjects. AB - BACKGROUND: Intracellular B-vitamin and folate deficiency indicated by hyperhomocysteinemia is very frequent in the elderly population. Hyperhomocysteinemia increases the risk of atherothrombotic diseases and neuropsychiatric complications. Our aim was to evaluate the prevalence of increased serum metabolite concentrations in subjects of a higher age, and whether the measurement of metabolite concentrations is more effective in diagnosing B-vitamin deficiency than mere homocysteine. MATERIALS AND METHODS: Homocysteine (HCY), cystathionine (CYS) and methylmalonic acid (MMA) were investigated in serum together with vitamin B-12, B-6 and folate in 90 high-aged subjects (85-102 years), 92 seniors (65-75 years), and in 50 younger subjects (19 50 years). RESULTS: Elderly subjects (high-aged and senior) had elevated serum concentrations of metabolites. High-aged subjects had a higher frequency of pathological increases than seniors: HCY 62% vs. 24%; MMA 62% vs. 23%; CYS 81% vs. 36%. Folate and vitamin B-6 concentrations were significantly decreased in both elderly groups; vitamin B-12 was only decreased in high-aged subjects. Utilising vitamin B-6, B-12 and folate for diagnosis of intracellular vitamin deficiency, the rate was 30% in seniors and 55% in high aged subjects. However, utilising the metabolites (HCY, MMA and CYS) for the diagnosis of intracellular vitamin deficiency, there was a distinctly increased rate of 55% in seniors respective to 90% in high-aged subjects. Backward multiple regression analysis revealed that only folate, MMA, creatinine and age were independent variables influencing the HCY concentration. Furthermore, the MMA concentration was significantly and independently influenced by folate, vitamin B-12, HCY and creatinine, and the serum concentration of CYS by vitamin B-12, creatinine and age. CONCLUSION: The metabolites HCY, MMA and CYS are sensitive indicators diagnosing impaired remethylation of homocysteine to methionine with parallel activation of catabolic pathway. Compared to mere HCY or B-vitamins in serum, the efficiency of diagnosing a disturbed HCY metabolism increases very much in utilising the metabolites HCY, MMA and CYS. For differential diagnosis, parallel measurement of folate and creatinine is recommended. The early and correct diagnosis of B-vitamin deficiency in elderly subjects is of high clinical relevance. PMID- 11122324 TI - Insulin-mediated increases in renal plasma flow are impaired in insulin-resistant normal subjects. AB - BACKGROUND: Impaired vasodilatation in skeletal muscle is a possible mechanism linking insulin resistance to blood pressure regulation. Increased renal vascular resistance has been demonstrated in the offspring of essential hypertensives. We assessed whether insulin-mediated renal vasodilatation is impaired in insulin resistant normal subjects. DESIGN: In two groups of 10 insulin-resistant and 10 insulin-sensitive normal subjects, we compared the effects of sequential physiological and supraphysiological insulin dosages (50 and 150 mU kg(-1) h(-1)) on renal plasma flow (RPF) and leg blood flow using the euglycaemic clamp technique, 131I-labelled Hippuran clearances and venous occlusion plethysmography. Time-control experiments were performed in the same subjects. RESULTS: Whole-body glucose uptake amounted to 4.9 +/- 2.1 and 11.0 +/- 2.4 mg kg(-1) min(-1) in the insulin-resistant and to 12.7 +/- 2.3 and 17.4 +/- 2.6 mg kg(-1) min(-1) in the insulin-sensitive subjects during physiological and supraphysiological hyperinsulinaemia, respectively. RPF increased more in insulin sensitive compared to insulin-resistant subjects during physiological hyperinsulinaemia (13.7 vs. 6.8%, P < 0.05). RPF increased to comparable levels during supraphysiological hyperinsulinaemia. Insulin-mediated changes in leg blood flow did not differ between groups. In the combined group, we found a positive correlation between insulin-mediated glucose uptake and changes in RPF during physiological hyperinsulinaemia (r = 0.57, P = 0.009), whereas insulin mediated glucose uptake correlated with changes in leg blood flow during supraphysiological hyperinsulinaemia (r = 0.54. P = 0.017). CONCLUSIONS: Our results suggest that the sensitivities of the skeletal muscle and renal vascular bed differ for insulin's vasodilatory action. Insulin-mediated increases in RPF are impaired in insulin-resistant but otherwise normal subjects during physiological hyperinsulinaemia. PMID- 11122325 TI - Biliary lipid composition in patients with cholesterol and pigment gallstones and gallstone-free subjects: deoxycholic acid does not contribute to formation of cholesterol gallstones. AB - BACKGROUND: Four main disturbances have been attributed to cholesterol gallstone disease: hypersecretion of cholesterol from the liver with cholesterol supersaturation in bile; disturbed motility with defective absorption and secretion by the gallbladder; increased crystallisation of cholesterol in the gallbladder bile; and slow intestinal transit with increased amount of deoxycholic acid in the bile acid pool. We aimed to evaluate the biliary lipid composition in a large series of gallstone patients, with emphasis on the amount of deoxycholic acid and with respect to number of stones, compared to gallstone free subjects. MATERIALS AND METHODS: Bile was sampled during operations through puncture of the gallbladder from 145 cholesterol gallstone patients, 23 patients with pigment stones and 87 gallstone free patients undergoing cholecystectomy. Biliary lipid composition, cholesterol saturation, bile acid composition, nucleation time and cholesterol crystals were analysed. RESULTS: The patients with cholesterol gallstones showed higher molar percentage of cholesterol, lower total biliary lipid concentration, higher cholesterol saturation, shorter nucleation time and higher proportion of crystals in bile than the other groups. The nucleation time was significantly shorter in multiple cholesterol gallstone patients, but this was not due to higher cholesterol saturation. Male cholesterol gallstone patients showed higher cholesterol levels, lower total biliary lipid concentration, and higher cholesterol saturation in bile than female patients. There was no difference in biliary content of deoxycholic acid, but significantly lower content of cholic acid in gallstone patients compared to gallstone free patients. CONCLUSIONS: We conclude that deoxycholic acid does not contribute to gallstone formation in cholesterol gallstone patients. The short nucleation time in patients with multiple cholesterol stones is not due to higher cholesterol saturation. PMID- 11122326 TI - Serum tumour necrosis factor-alpha levels in cancer patients are discontinuous and correlate with weight loss. AB - BACKGROUND: Tumour necrosis factor-alpha (TNF) has been regarded as a potential mediator of cancer cachexia. Assessment of TNF circulating levels in cancer patients and their correlation with weight loss has led to controversial results. MATERIALS AND METHODS: We measured TNF circulating levels in 28 patients with gastrointestinal cancer and 29 controls with benign gastrointestinal diseases at different times (08.00 h, 14.00 h, 20.00 h) before operation. RESULTS: TNF activity was not detected in any of controls at any times. In cancer patients, TNF circulating levels were detectable in 18 cases (64.3%) and appeared to be discontinuous. TNF levels above the limit of detection were present in 15 patients (53.6%) at 08.00 h, in 14 (50%) at 014.00 h and in nine (32.1%) at 20.00 h. Mean TNF levels were 14.3 +/- 4 pg mL(-1) at 08.00 h, 16.7 +/- 4.6 pg mL(-1) at 14.00 h (P = 0.05) and 18.5 +/- 10.2 pg mL(-1) at 20.00 h (P < 0.05 vs. 08.00 h and 14.00 h). According to Spearman's analysis, the sum of TNF concentrations at the three times significantly correlated with the severity of weight loss (Spearman's correlation coefficient = - 0.420; P = 0.026). TNF concentrations were consistently and significantly higher in patients with severe weight loss than in those with moderate or light weight loss at 08.00 h (26.3 +/- 8.3, 8.9 +/ 4.2, 3.8 +/- 2.1, respectively; P = 0.04 at one-way ANOVA). TNF levels were higher in anorectic than in nonanorectic patients at any hour, but the differences were not statistically significant. CONCLUSIONS: The present study demonstrates that TNF is intermittently or discontinuously detectable in patients with gastrointestinal cancer and that its levels correlate with the severity of weight loss. PMID- 11122327 TI - The use of platelet density and volume measurements to estimate the severity of pre-eclampsia. AB - BACKGROUND: This study evaluated whether it is possible to estimate the severity of pre-eclampsia through in vitro measurements of platelet and granulocyte parameters. EXPERIMENTAL PROTOCOL: Eighteen pre-eclamptic women in the third trimester of pregnancy and 11 women in the third-trimester of normal pregnancies were included in the study. Three to 12 months after delivery, 15 patients with pre-eclampsia and all the subjects with normal pregnancies were re-examined. Before delivery, peak platelet density was determined using a specially designed apparatus. Before and 3-12 months after delivery the following were measured: platelet counts, mean platelet volume and neutrophil and monocyte counts. Furthermore, circulating P-selectin, interleukin-6 and myeloperoxidase were determined to estimate platelet, monocyte and granulocyte activities, respectively. RESULTS: Compared to their results after delivery, pre-eclamptic females demonstrated lower platelet counts (P < 0.001) and raised mean platelet volumes (P < 0.01). Both pre-eclamptic women (P < 0.01) and normal pregnancies (P < 0.05) demonstrated elevated soluble P-selectin at pregnancy. Then pre-eclamptic women had advanced neutrophil counts (P < 0.01) but normal pregnancies showed a similar phenomenon (P < 0.001). Interleukin-6 remained normal during pregnancy. Plasma myeloperoxidase levels were lower both in pre-eclampsia (P < 0.05) and in normal pregnancies (P < 0.001). In pre-eclampsia elevated blood pressure was related to higher mean platelet volumes (P < 0.05). Furthermore, a group of pre eclamptic females whose platelets had disturbed density distribution displayed elevated mean platelet volumes (P < 0.01). CONCLUSIONS: The present work demonstrates considerable platelet alterations in pre-eclampsia. We failed to show granulocyte involvement in the pathogenesis of the disease. Severe pre eclampsia is related to elevated mean platelet volumes. The latter parameter is associated with disturbed density distribution. It appears possible to estimate disease severity from measurements of platelet density and volume. PMID- 11122328 TI - Distribution of glycine receptor subunits on primate retinal ganglion cells: a quantitative analysis. AB - This study investigates the distribution of inhibitory neurotransmitter receptors on sensory neurons. Ganglion cells in the retina of a New World monkey, the common marmoset Callithrix jacchus, were injected with Lucifer yellow and Neurobiotin and subsequently processed with antibodies against one (alpha1), or against all subunits, of the glycine receptor, or against the anchoring protein gephyrin. Immunoreactive (IR) puncta representing glycine receptor or gephyrin clusters were found on the proximal and the distal dendrites of all ganglion cell types investigated. For both parasol and midget cells, the density of receptor clusters was greater on distal than proximal dendrites for all antibodies tested. In parasol cells the average density for the alpha1 subunit of the glycine receptor was 0.087 IR puncta/microm of dendrite, and for all subunits it was 0.119 IR puncta/microm of dendrite. Thus, the majority of glycine receptors on parasol cells contain the alpha1 subunit. For parasol cells, we estimated an average of 1.5 glycinergic synapses/100 microm2 dendritic membrane on proximal dendrites and about 9.4 glycinergic synapses/100 microm2 on distal dendrites. The segregation of receptors to the distal dendrites appears to be a common feature of inhibitory neurotransmitter input to parasol and midget cells, and might be associated with the receptive field surround mechanism. PMID- 11122329 TI - Endogenous BDNF is required for myelination and regeneration of injured sciatic nerve in rodents. AB - Following a peripheral nerve injury, brain-derived neurotrophic factor (BDNF) and the p75 neurotrophin receptor are upregulated in Schwann cells of the Wallerian degenerating nerves. However, it is not known whether the endogenous BDNF is critical for the functions of Schwann cells and regeneration of injured nerve. Treatment with BDNF antibody was shown to retard the length of the regenerated nerve from injury site by 24%. Histological and ultrastructural examination showed that the number and density of myelinated axons in the distal side of the lesion in the antibody-treated mice was reduced by 83%. In the BDNF antibody treated animals, there were only distorted and disorganized myelinated fibres in the injured nerve where abnormal Schwann cells and phagocytes were present. As a result of nerve degeneration in BDNF antibody-treated animals, subcellular organelles, such as mitochondria, disappeared or were disorganized and the laminal layers of the myelin sheath were loosened, separated or collapsed. Our in situ hybridization revealed that BDNF mRNA was expressed in Schwann cells in the distal segment of lesioned nerve and in the denervated muscle fibres. These results indicate that Schwann cells and muscle fibres may contribute to the sources of BDNF during regeneration and that the deprivation of endogenous BDNF results in an impairment in regeneration and myelination of regenerating axons. It is concluded that endogenous BDNF is required for peripheral nerve regeneration and remyelination after injury. PMID- 11122330 TI - Parkin expression in the adult mouse brain. AB - Mutations in a protein designated Parkin were shown to be involved in the pathogenesis of autosomal recessive juvenile parkinsonism. Nothing is known about its regional and subcellular distribution in the mouse. In order to elucidate the Parkin mRNA and protein distribution in the adult mouse, the mouse cDNA was cloned and polyclonal antisera were generated against the N-terminal part of mouse Parkin. The antibodies were shown to be specific using Western blot analysis, immunostaining of cells transfected with mouse Parkin and pre absorption tests. The Parkin protein expression profile was studied using immunohistochemistry and Western blot analysis and was compared with that of the mRNA yielded by in situ hybridization and RT-PCR analysis. Parkin protein was widely distributed in all subdivisions of the mouse brain. Low levels were found in the telencephalon and diencephalon, while the brainstem contained a large number of cells heavily expressing Parkin. Ultrastructural analysis and double immunohistochemistry revealed that the majority of Parkin-expressing cells were neurons, while only single glial cells exhibited immunostaining. The protein was distributed nonhomogeneously throughout the entire cytoplasm. A subpopulation of Parkin-immunopositive cells displayed speckled immunodeposits in the nucleus. Dopaminergic cells of the substantia nigra pars compacta exhibited high levels of Parkin mRNA but no Parkin protein, while the striatum contained immunopositive profiles but no mRNA signals. Our data indicate that Parkin is neither restricted to a single functional system nor associated with a particular transmitter system. The speckled nuclear distribution of Parkin immunoreactivity strongly suggests a role for Parkin in gene expression. PMID- 11122331 TI - High susceptibility of the anterior and posterior piriform cortex to induction of convulsions by bicuculline. AB - Accumulating evidence suggests that the piriform cortex (PC) plays a critical role in the development of limbic motor seizures. In the anterior piriform cortex (aPC), a functionally defined, discrete epileptogenic site has been previously identified by unilateral microinjection of bicuculline in Sprague-Dawley (SD) rats and termed the 'area tempestas' (AT). Compared to this site in the aPC, more posterior PC sites, particularly a site in the transition zone between the posterior and aPC (central PC) exhibited a greater susceptibility to electrical stimulation. However, it is not known whether central and posterior sites in the PC differ from the aPC, including the AT, with regard to their sensitivity to bicuculline. In the present study, unilateral focal microinfusion of picomole quantities of bicuculline induced behavioural (focal and generalized) seizures in deep layers of all parts of the PC in two rat strains, Wistar and SD. The incidence of generalized seizures was higher in the AT of SD rats, but no such difference was seen in Wistar rats, arguing against the previous proposal that the rat AT is unique in its sensitivity to induction of seizures by bicuculline compared to other locations within or outside of the PC. Injection of biotin dextran in PC seizure-sensitive sites in SD rats showed clear differences in anterograde and retrograde labelling between the different PC sites. Therefore, although it was possible to evoke generalized seizures from all parts of the PC, the anatomical connections of the bicuculline injection sites were qualitatively different. The results suggest that the deep layers of the entire PC are highly sensitive to seizure induction by bicuculline, thus substantiating the notion that the PC may be important in seizure generation and propagation. PMID- 11122332 TI - Clock genes outside the suprachiasmatic nucleus involved in manifestation of locomotor activity rhythm in rats. AB - Chronic treatment of methamphetamine (MAP) in rats desynchronized the locomotor activity rhythm from the light-dark cycle. When the activity rhythm was completely phase-reversed with respect to a light dark-cycle, 24 h profiles were examined for the clock gene (rPer1, rPer2, rBMAL1, rClock) expressions in several brain structures by in situ hybridization, and for the pineal as well as plasma melatonin levels. In the MAP-treated rats, the rPer1 expression in the suprachiasmatic nucleus (SCN) showed a robust circadian rhythm which was essentially identical to that in the control rats. Circadian rhythms in pineal as well as plasma melatonin were not changed significantly in the MAP-treated rats. However, robust circadian rhythms in the rPer1, rPer2 and rBMAL1 expressions detected in the caudate-putamen (CPU) and parietal cortex were completely phase reversed in the MAP-treated rats, compared with those in the control rats, indicating desynchronization from the SCN rhythm. Such desynchronization was not observed in the circadian rhythms of clock gene expression in the nucleus accumbens and cingulate cortex. The circadian rClock expression rhythm in the MAP treated rats was not phase-reversed in the CPU and parietal cortex. These findings indicate that the locomotor activity rhythm in rats is directly driven by the pacemaker outside the SCN, in which rPer1, rPer2 and rBMAL1 in the CPU and parietal cortex are involved. PMID- 11122333 TI - Interaction of the C-terminal tail region of the metabotropic glutamate receptor 7 with the protein kinase C substrate PICK1. AB - Group III metabotropic glutamate receptors (mGluRs) are highly enriched in the presynaptic terminals of glutamatergic synapses where they mediate feedback inhibition of neurotransmitter release. Here, we used the yeast two-hybrid system to identify a direct interaction of the C-terminal tail region of mGluR7 with the rat homologue of the protein kinase C substrate PICK1. This interaction is specifically mediated by the very C-terminal amino acids of the receptor and can be reconstituted in human embryonic kidney 293 cells by transfection of full length mGluR7 and PICK1 cDNAs. Quantitative beta-galactosidase assays revealed that among the different group III mGluRs, mGluR7 is the major PICK1 binding partner although other subfamily members can also interact with PICK1. These data indicate that PDZ domain-containing proteins might contribute to the presynaptic localization of group III mGluRs. PMID- 11122334 TI - Cooperative changes in GABA, glutamate and activity levels: the missing link in cortical plasticity. AB - Different intracortical mechanisms have been reported to contribute to the substantial topographic reorganization of the mammalian primary visual cortex in response to matching lesions in the two retinas: an immediate expansion of receptive fields followed by a gradual shift of excitability into the deprived area and finally axonal sprouting of laterally projecting neurons months after the lesion. To gain insight into the molecular mechanisms of this adult plasticity, we used immunocytochemical and bioanalytical methods to measure the glutamate and GABA neurotransmitter levels in the visual cortex of adult cats with binocular central retinal lesions. Two to four weeks after the lesions, glutamate immunoreactivity was decreased in sensory-deprived cortex as confirmed by HPLC analysis of the glutamate concentration. Within three months normal glutamate immunoreactivity was restored. In addition, the edge of the unresponsive cortex was characterized by markedly increased glutamate immunoreactivity 2-12 weeks postlesion. This glutamate immunoreactivity peak moved into the deprived area over time. These glutamate changes corresponded to decreased spontaneous and visually driven activity in unresponsive cortex and to strikingly increased neuronal activity at the border of this cortical zone. Furthermore, the previously reported decrease in glutamic acid decarboxylase immunoreactivity was found to reflect decreased GABA levels in sensory-deprived cortex. Increased glutamate concentrations and neuronal activity, and decreased GABA concentrations, may be related to changes in synaptic efficiency and could represent a mechanism underlying the retinotopic reorganization that occurs well after the immediate receptive field expansion but long before the late axonal sprouting. PMID- 11122335 TI - Up-regulation of growth-associated protein 43 mRNA in rat medial septum neurons axotomized by fimbria-fornix transection. AB - Transection of septohippocampal fibres is widely used to study the response of CNS neurons to axotomy. Septohippocampal projection neurons survive axotomy and selectively up-regulate the transcription factor c-Jun. In the present study we investigated whether these cells concomitantly up-regulate the growth-associated protein-43 (GAP-43), a potential target gene of c-Jun implicated in axonal growth and regeneration. Using in situ hybridization histochemistry (ISHH) it was demonstrated that postlesional c-jun mRNA expression is accompanied by an increased expression of GAP-43 mRNA in the medial septum 3 days following fimbria fornix transection (FFT). The increase reached a maximum at 7 days and gradually declined thereafter (17 days, 3 weeks). Retrograde prelabeling with Fluoro-Gold followed by axotomy and ISHH revealed that GAP-43 mRNA was up-regulated in septohippocampal projection neurons. Colocalization of GAP-43 mRNA and choline acetyltransferase protein showed that GAP-43 mRNA was expressed by cholinergic medial septal neurons after axotomy. Selective immunolesioning of the cholinergic component of the septohippocampal projection with 192 IgG-saporin followed by FFT demonstrated that GAP-43 mRNA was also synthesized by axotomized GABAergic neurons. These results demonstrate an up-regulation of GAP-43 mRNA in axotomized septohippocampal projection neurons independent of their transmitter phenotype which is closely correlated with c-Jun expression. Because the GAP-43 gene contains an AP-1 site, we hypothesize a c-Jun-driven up-regulation of GAP-43 in lesioned medial septal neurons that may contribute to their survival and regenerative potential following axotomy. PMID- 11122336 TI - Vascular endothelial growth factor is a neurotrophic factor which stimulates axonal outgrowth through the flk-1 receptor. AB - Vascular endothelial growth factor (VEGF) is an angiogenic factor that stimulates axonal outgrowth. Here we used in situ hybridization and immunocytochemistry to study the VEGF receptor flk-1 in cultured superior cervical ganglia (SCG) and dorsal root ganglia (DRG) from adult mice, and also the effects of VEGF on regeneration in vitro. Neurons in both ganglia contained the flk-1 receptor and showed an increased mRNA expression and immunoreactivity for flk-1 after 48 h in culture. In SCG, but not in DRG, double immunostaining for flk-1 and VEGF revealed coexpression in many neurons, implying that VEGF may exert both autocrine and paracrine actions. One proportion of the flk-1-positive neurons in DRG stained positive for the large neuron marker RT97 and another proportion expressed calcitonin gene-related peptide (CGRP). Small IB4-positive neurons were devoid of flk-1 immunoreactivity. Most flk-1-positive neurons in the DRG, but not in the SCG, were also immunoreactive to neuropilin-1. VEGF was found to stimulate axonal outgrowth from DRG, both by an action on the growing axons and the nerve cell bodies. The latter effect could be mediated by retrograde axonal transport as revealed by the use of a two compartment system to assay axonal outgrowth. We also found that the VEGF-induced axonal outgrowth was blocked by the flk-1 inhibitor SU5416. The results strongly suggest that VEGF acts as a neurotrophic factor and plays an important role during the regeneration of peripheral nerves. PMID- 11122337 TI - Retrograde signalling with nitric oxide at neocortical synapses. AB - Long-term changes of synaptic transmission in slices of rat visual cortex were induced by intracellular tetanization: bursts of short depolarizing pulses applied through the intracellular electrode without concomitant presynaptic stimulation. Long-term synaptic changes after this purely postsynaptic induction were associated with alterations of release indices, thus providing a case for retrograde signalling at neocortical synapses. Both long-term potentiation and long-term depression were accompanied by presynaptic changes, indicating that retrograde signalling can achieve both up- and down-regulation of transmitter release. The direction and the magnitude of the amplitude changes induced by a prolonged intracellular tetanization depended on the initial properties of the input. The inputs with initially high paired-pulse facilitation (PPF) ratio, indicative of low release probability, were most often potentiated. The inputs with initially low PPF ratio, indicative of high release probability, were usually depressed or did not change. Thus, prolonged postsynaptic activity can lead to normalization of the weights of nonactivated synapses. The dependence of polarity of synaptic modifications on initial PPF disappeared when plastic changes were induced with a shorter intracellular tetanization, or when the NO signalling pathway was interrupted by inhibition of NO synthase activity or by application of NO scavengers. This indicates that the NO-dependent retrograde signalling system has a relatively high activation threshold. Long-term synaptic modifications, induced by a weak postsynaptic challenge or under blockade of NO signalling, were nevertheless associated with presynaptic changes. This suggests the existence of another retrograde signalling system, additional to the high threshold, NO-dependent system. Therefore, our data provide a clear case for retrograde signalling at neocortical synapses and indicate that multiple retrograde signalling systems, part of which are NO-dependent, are involved. PMID- 11122338 TI - The central acoustic tract and audio-vocal coupling in the horseshoe bat, Rhinolophus rouxi. AB - Doppler shift compensation (DSC) behaviour in horseshoe bats is a remarkable example of sensorimotor feedback that stabilizes the echo frequency at the bat's optimum hearing range regardless of motion-induced frequency shifts in the echoes. Searching for a related neural interface, the nucleus of the central acoustic tract (NCAT) was investigated in the echolocating horseshoe bat, Rhinolophus rouxi, using various neurophysiological and tracer methods. The NCAT receives bilateral auditory input from the cochlear nuclei and sends projections to regions outside the classical acoustic pathway like the pretectal area or the superior colliculus. The binaural input is excitatory from the contralateral and inhibitory from the ipsilateral ear to 53% of the units, and auditory responses were biased to frontal and contralateral directions. The best frequencies of NCAT neurons match a narrow range above the main frequency component of the bat's species-specific echolocation call (62% of the units), and the neurons exhibit extremely sharp tuning (Q10dB up to 632). DSC is degraded by unilateral electrical or pharmacological microstimulation of the NCAT, and heavily impaired by unilateral lesion of the region. Altogether, the efferents of the NCAT to prevocal areas, the tuning of its neurons to the DSC-relevant echo frequency range, and the possibility to affect DSC by manipulation of the NCAT, support the assumption that the nucleus plays an important role in audio-vocal control in the horseshoe bat. PMID- 11122339 TI - Cloning, distribution and functional analysis of the type III sodium channel from human brain. AB - The type III voltage-gated sodium channel was cloned from human brain. The full length cDNA has 89% identity with rat type III, and the predicted protein (1951 amino acids) has 55 differences. The expression pattern of human type III mRNA was determined in adult brain tissue and, in contrast to rat, was detected in many regions, including caudate nucleus, cerebellum, hippocampus and frontal lobe. The human type III channel was stably expressed in Chinese hamster ovary (CHO) cells and its biophysical properties compared to the human type II channel using identical conditions. The voltage dependence and kinetics of activation were found to be similar to that of type II. The kinetics of inactivation of the two human subtypes were also similar. However, type III channels inactivated at more hyperpolarized potentials and were slower to recover from inactivation than type II. When expressed in human embryonic kidney (HEK293T) cells, type III channels produced currents with a prominent persistent component, which were similar to those reported for rat type II [Ma et al. (1997) Neuron, 19, 443-452]. However, unlike type II, this was prominent even in the absence of coexpressed G proteins, suggesting type III may adopt this gating mode more readily. The distinct properties of the channel, together with its wide distribution in adult brain, suggest that in humans, type III may have important physiological roles under normal, and perhaps also pathological conditions. PMID- 11122340 TI - Topographical projection from the superior colliculus to the nucleus of the brachium of the inferior colliculus in the ferret: convergence of visual and auditory information. AB - The normal maturation of the auditory space map in the deeper layers of the ferret superior colliculus (SC) depends on signals provided by the superficial visual layers, but it is unknown where or how these signals influence the developing auditory responses. Here we report that tracer injections in the superficial layers label axons with en passant and terminal boutons, both in the deeper layers of the SC and in their primary source of auditory input, the nucleus of the brachium of the inferior colliculus (nBIC). Electron microscopy confirmed that biocytin-labelled SC axons form axodendritic synapses on nBIC neurons. Injections of biotinylated dextran amine in the nBIC resulted in anterograde labelling in the deeper layers of the SC, as well as retrogradely labelled superficial and deep SC neurons, whose distribution varied systematically with the rostrocaudal placement of the injection sites in the nBIC. Topographical order in the projection from the SC to the ipsilateral nBIC was confirmed using fluorescent microspheres. We demonstrated the existence of functional SC-nBIC connections by making whole-cell current-clamp recordings from young ferret slices. Both monosynaptic and polysynaptic EPSPs were generated by electrical stimulation of either the superficial or deep SC layers. In addition to unimodal auditory units, both visual and bimodal visual-auditory units were recorded in the nBIC in vivo and their incidence was higher in juvenile ferrets than in adults. The SC-nBIC circuit provides a potential means by which visual and other sensory or premotor signals may be delivered to the nBIC to calibrate the representation of auditory space. PMID- 11122341 TI - Increased neuronal damage and apoE immunoreactivity in human apolipoprotein E, E4 isoform-specific, transgenic mice after global cerebral ischaemia. AB - Apolipoprotein E (apoE, protein; APOE, gene) is expressed as three isoforms in humans (E2, E3, E4). The APOE-epsilon4 allele is associated with a poor outcome in patients after head injury of which ischaemic brain damage is a contributor of mortality and morbidity. The aim of the study was to determine whether mice expressing human APOE-epsilon4 displayed more extensive ischaemic neuronal damage 72 h after transient global ischaemia compared with mice which express human APOE epsilon3. APOE-epsilon3 and -epsilon4 transgenic mice, under the control of a human promoter, were used which express human APOE in neurons and glia. Ischaemic neuronal damage in the CA1 pyramidal cell layer in the APOE-epsilon4 transgenic mice was significantly greater than in the APOE-epsilon3 mice after global ischaemia (36.4+/-8.9%, 18.2+/-7.3%; P<0.05). This was associated with more extensive neuronal apoE immunoreactivity in the CA1 pyramidal cell layer in the APOE-epsilon4 transgenic mice compared with APOE-epsilon3 transgenic mice. In contrast, in the caudate nucleus, there were similar levels of ischaemic neuronal damage in the APOE-epsilon3 and -epsilon4 transgenic mice (39.2 +/-10.1%; 44.6+/ 8.4%, P = 0.32). In the caudate, similar numbers of neurons were immunostained for apoE in the APOE-epsilon3 and -epsilon4 transgenic mice. The present study demonstrated that the APOE-epsilon4 allele is associated with an increased vulnerability of a specific brain region to the effects of global ischaemia, which is closely associated with an increase in neuronal apoE. The data extend previous work and are consistent with an association of the APOE-epsilon4 allele with a poor outcome after acute brain injury in humans. PMID- 11122342 TI - cDNA cloning and expression of a gamma-aminobutyric acid A receptor epsilon subunit in rat brain. AB - A cDNA encoding a GABA(A) receptor subunit was isolated from rat brain. The predicted protein is 70% identical to the human epsilon-subunit. It was recently reported [Sinkkonen et al. (2000), J. Neurosci., 20, 3588-3595] that the rodent epsilon-subunit mRNA encoded an additional sequence ( approximately 400 residues). We provide evidence that human and rat epsilon-subunit are similar in size. The distribution of cells expressing the GABA(A) epsilon-subunit was examined in the rat brain. In situ hybridization histochemistry revealed that epsilon-subunit mRNA is expressed by neurons located in septal and preoptic areas, as well as in various hypothalamic nuclei, including paraventricular, arcuate, dorsomedial and medial tuberal nuclei. The mRNA was also detected in major neuronal groups with broad-range influence, such as the cholinergic (basal nucleus), dopaminergic (substantia nigra compacta), serotonergic (raphe nuclei), and noradrenergic (locus coeruleus) systems. Immunohistochemistry using an affinity-purified antiserum directed towards the N-terminal sequence unique to the rat epsilon-subunit revealed the presence of epsilon-subunit immunoreactivity over the somatodendritic domain of neurons with a distribution closely matching that of mRNA-expressing cells. Moreover, using in situ hybridization, alpha3, theta and epsilon GABA(A) subunit mRNAs were all detected with an overlapping distribution in neurons of the dorsal raphe and the locus coeruleus. Our results suggest that novel GABA(A) receptors may regulate, neuroendocrine and modulatory systems in the brain. PMID- 11122343 TI - Differential induction of NF-kappaB activity and neural cell death by antidepressants in vitro. AB - Tricyclic antidepressants and selective serotonin reuptake inhibitors are here shown to induce cell death in a neural cell line. The exposure to these drugs led to increased generation of reactive oxygen species and a concomitant reduction of intracellular glutathione levels. Furthermore, these antidepressants induced DNA fragmentation and increased the transcriptional and DNA-binding activity of NF kappaB. In contrast, treatment with type A and B monoamine oxidase inhibitors did not induce changes in NF-kappaB activity and did not exert a detrimental influence on cell viability. These results indicate that some antidepressant drugs may cause both oxidative stress and changes in cellular antioxidative capacity, resulting in altered NF-kappaB activity and, ultimately, cell death. PMID- 11122344 TI - Changes in the firing pattern of globus pallidus neurons after the degeneration of nigrostriatal pathway are mediated by the subthalamic nucleus in the rat. AB - Changes in the neuronal activity of globus pallidus (GP) have been shown in animal models of parkinsonism. In order to study the implication of the subthalamic nucleus (STN) in these changes, the effects of STN lesions alone or in combination with 6-hydroxydopamine (6-OHDA) -induced damage to the substantia nigra compacta (SNc) were examined in rats using electrophysiological recordings of GP cells. In normal rats, the firing rate was 22.1+/-1.4 spikes/s. The pattern was regular in 45%, irregular in 49% and bursty in 6% of the cases. In rats with STN lesions, the firing rate of GP units (20.15+/-1.25 spikes/s) did not differ from that of normal rats and only regular (46%) and irregular (54%) cells were found; a bursty pattern was not observed. 6-OHDA lesions of the SNc induced no change in the firing rate of GP neurons (21.5+/-1.4 spikes/s, P>0.05) but a significant decrease in the percentage of regular cells (27%, P<0.001), a significant increase in burst cells (21%, P<0.001) with no change in the percentage of irregular units (52%) were observed. In rats with combined SNc and STN lesions, the firing pattern did not change from that of normal rats. The present results show that STN lesions induced the disappearance of bursts in normal rats and normalization of firing pattern in the GP units of rats with 6 OHDA lesions suggesting that the STN plays an important role in the modulation of the pattern of activity of GP neurons which may account for the therapeutic effect of STN lesions in Parkinson's disease. PMID- 11122345 TI - Subcellular compartmentalization of a potassium channel (Kv1.4): preferential distribution in dendrites and dendritic spines of neurons in the dorsal cochlear nucleus. AB - Voltage-dependent ion channels have specific patterns of distribution along the neuronal plasma membrane of dendrites, cell bodies and axons, which need to be unravelled in order to understand their contribution to neuronal excitability and firing patterns. We have investigated the subcellular compartmentalization of Kv1.4, a transient, fast-inactivating potassium channel, in fusiform cells and related interneurons of the rat dorsal cochlear nucleus. A polyclonal antibody which binds to a region near the N-terminus domain of a Kv1.4 channel was raised in rabbits. Using a high-resolution combination of immunocytochemical methods, Kv1.4 was localized mainly in the apical dendritic trunks and cell bodies of fusiform cells, as well as in dendrites and cell bodies of interneurons of the dorsal cochlear nucleus, likely cartwheel cells. Quantitative immunogold immunocytochemistry revealed a pronounced distal to proximal gradient in the dendrosomatic distribution of Kv1. 4. In plasma membrane localizations, Kv1.4 was preferentially present in dendritic spines, either in the spine neck or in perisynaptic locations, always away from the postsynaptic density. These findings indicate that Kv1.4 is largely distributed in dendritic compartments of fusiform and cartwheel cells of the dorsal cochlear nucleus. Its preferential localization in dendritic spines, where granule cell axons make powerful excitatory synapses, suggests a role for this voltage-dependent ion channel in the regulation of dendritic excitability and excitatory inputs. PMID- 11122346 TI - Assignment of ecto-nucleoside triphosphate diphosphohydrolase-1/cd39 expression to microglia and vasculature of the brain. AB - Extracellular nucleotides are ubiquitous extracellular mediators that interact with and activate nucleotide type 2 (P2) receptors. These receptors initiate a wide variety of signalling pathways that appear important for functional associations between neurons and glial cells and for the regulation of blood flow, haemostatic and inflammatory reactions in the brain. Ectonucleotidases are extracellular nucleotide-metabolizing enzymes that modulate P2 receptor-mediated signalling by the regulated hydrolysis of these agonists. A considerable number of ectoenzyme species with partially overlapping substrate and tissue distributions have been described. Major candidates for expression in the brain are members of the ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase or CD39) family. The production of cd39-/- mice and specific reagents have enabled us to analyse the specific cellular distribution of NTPDase1 (CD39), the prototype member of the enzyme family, in the mouse brain. Using monospecific antibodies and enzyme histochemical staining, we have identified NTPDase1 as a major ectonucleotidase associated with both microglia and the endothelial and smooth muscle cells of the vasculature. NTPDase1 is not expressed by neurons and astrocytes. Additional unidentified ectonucleotidase functional activity is observed at lower levels throughout the brain parenchyma. NTPDase1 may regulate P2 receptor-mediated functions of microglia as well as influence nucleotide signalling between neurons or astrocytes that are associated with multiple microglial ramifications. The expression of NTPDase1 by cerebrovascular endothelial and smooth muscle cells also suggests involvement in the regulation of blood flow and thrombogenesis. PMID- 11122347 TI - Differential neuronal expression and projections of melanin-concentrating hormone (MCH) and MCH-gene-overprinted-polypeptide (MGOP) in the rat brain. AB - The rat melanin-concentrating hormone (MCH) gene may produce, through alternative splicing, either the precursor of MCH and neuropeptide EI, two neuropeptides coexpressed in the zona incerta (ZI) and lateral hypothalamus (LHA), or a putative protein we named previously MCH-gene-overprinted-polypeptide (MGOP). First, we investigated the distribution and relative expression of MCH and MGOP mRNA in the rat brain by Northern blotting, RT-PCR and in situ hybridization. MGOP gene transcripts were detected mainly in the hypothalamus only by RT-PCR. Second, different antisera were raised toward the C-terminus of MGOP and used to identify the translational products. In the rat brain, no MGOP-processed peptide could be detected based on RP-HPLC coupled to specific RIA. A polypeptide of 14 kDa was found in the secretory pathway of transfected monkey COS7 cells expressing recombinant MGOP. In the rat hypothalamus, a specific protein of 12 kDa was identified by Western blot analysis. Finally, distribution of MGOP immunoreactivity (IR) was investigated in the rat brain. Colocalization studies demonstrated that 98% of the MGOP-expressing perikarya in ZI/LHA also synthesized MCH. In addition, numerous, strongly stained MGOP-containing neurons were encountered in the hypothalamic periventricular nucleus. Perikarya labelled with MGOP antiserum were also found scattered in the cortex, caudate putamen, amygdala and lateral septal nucleus. MCH was not detected in these MGOP-containing neurons. Strikingly, dense staining of terminals was observed with MGOP antiserum but not with MCH antibodies in the suprachiasmatic, ventromedial and arcuate nuclei, and also in the external layer of the median eminence. These results demonstrated that MGOP and MCH-IR overlapped in LHA/ZI but displayed a differential distribution in other areas. Based on this cerebral distribution, MGOP may act as a new secreted protein in regulating many neuroendocrine functions, such as nursing, feeding and growth control in associated behavioural components. PMID- 11122348 TI - Electrical stimulation accelerates and increases expression of BDNF and trkB mRNA in regenerating rat femoral motoneurons. AB - Electrical stimulation promotes the speed and accuracy of motor axonal regeneration. The positive effects of stimulation are mediated at the cell body. Here we characterize the effect of electrical stimulation on motoneuronal expression of BDNF and its receptor, trkB, two genes whose expression levels in motoneurons correlate with regeneration and are regulated by electrical activity in a variety of neurons. We used semiquantitative in situ hybridization to measure expression of mRNA encoding BDNF and the full-length trkB receptor at intervals of 8 h, 2 days and 7 days after unilateral femoral nerve cut, suture, and stimulation. Expression in regenerating motoneurons was compared to that of contralateral intact motoneurons. BDNF and trkB signals were not significantly upregulated 8 h and 2 days after femoral nerve suture and sham stimulation. By 7 days, there was a 2-fold increase in both BDNF and trkB mRNA expression. In contrast, stimulation of cut and repaired nerves for only 1 h led to rapid upregulation of BDNF and trkB mRNA by 3-fold and 2-fold, respectively, within the first 8 h. The stimulation effect peaked at 2 days with 6-fold and 4-fold increases in the signals, respectively. Thereafter, the levels of BDNF and trkB mRNA expression declined to equal the 2-fold increase seen at 7 days after nerve repair and sham-stimulation. We conclude that brief electrical stimulation stimulates BDNF and trkB expression in regenerating motoneurons. Because electrical stimulation is known to accelerate axonal regeneration, we suggest that changes in the expression of BDNF and trkB correlate with acceleration of axonal regeneration. PMID- 11122349 TI - Nerve growth factor expression in parasympathetic neurons: regulation by sympathetic innervation. AB - Interactions between sympathetic and parasympathetic nerves are important in regulating visceral target function. Sympathetic nerves are closely apposed to, and form functional synapses with, parasympathetic axons in many effector organs. The molecular mechanisms responsible for these structural and functional interactions are unknown. We explored the possibility that Nerve Growth Factor (NGF) synthesis by parasympathetic neurons provides a mechanism by which sympathetic-parasympathetic interactions are established. Parasympathetic pterygopalatine ganglia NGF-gene expression was examined by in situ hybridization and protein content assessed by immunohistochemistry. Under control conditions, NGF mRNA was present in approximately 60% and NGF protein was in 40% of pterygopalatine parasympathetic neurons. Peripheral parasympathetic axons identified by vesicular acetylcholine transporter-immunoreactivity also displayed NGF immunoreactivity. To determine if sympathetic innervation regulates parasympathetic NGF expression, the ipsilateral superior cervical ganglion was excised. Thirty days postsympathectomy, the numbers of NGF mRNA-positive neurons were decreased to 38% and NGF immunoreactive neurons to 15%. This reduction was due to a loss of sympathetic nerve impulse activity, as similar reductions were achieved when superior cervical ganglia were deprived of preganglionic afferent input for 40 days. These findings provide evidence that normally NGF is synthesized by parasympathetic neurons and transported anterogradely to fibre terminals, where it may be available to sympathetic axons. Parasympathetic NGF expression, in turn, is augmented by impulse activity within (and presumably transmitter release from) sympathetic axons. It is suggested that parasympathetic NGF synthesis and its modulation by sympathetic innervation provides a molecular basis for establishment and maintenance of autonomic axo-axonal synaptic interactions. PMID- 11122350 TI - Differential roles of corticotropin-releasing factor receptor subtypes 1 and 2 in opiate withdrawal and in relapse to opiate dependence. AB - The possible effects on the morphine withdrawal signs of the nonspecific corticotropin-releasing factor (CRF) receptor antagonist alpha-helical CRF, the selective CRF receptor subtype 1 antagonist CP-154,526 and the selective CRF receptor subtype 2 antagonist antisauvagine-30 (AS-30) were investigated in rats. The most withdrawal signs, including jumping, teeth chatter, writhing, shakes, lacrimation, piloerection, irritability and diarrhoea, were attenuated by pretreatment with alpha-helical CRF (10 microg i.c.v.) and CP-154,526 (30 mg/kg i.p.). However, no morphine withdrawal signs except for diarrhea were significantly affected by pretreatment with AS-30 (10 microg, i.c.v.). To investigate the possible role of different CRFR antagonists (alpha-helical CRF, CP-154,526 and AS-30) in relapse to opiate dependence, the 28-day extinction of morphine-conditioned place preference (CPP) was used. The morphine-CPP disappeared following a 28-day extinction and then was reactivated by a single injection of 10 mg/kg morphine. Pretreatment with alpha-helical CRF (10 microg, i.c.v.) and CP-154, 526 (30 mg/kg, i.p.) could significantly block this reactivation of morphine-CPP. In contrast, pretreatment with AS-30 (1 or 10 microg i. c.v.) did not affect this reactivation of morphine-CPP. The present study demonstrated that activation of the CRF receptor is involved in morphine withdrawal signs and relapse to morphine dependence, and that the role of CRF receptor subtypes 1 and 2 in withdrawal and reactivation of morphine dependence is not identical. CRF receptor subtype 1, but not subtype 2, is largely responsible for the action of the CRF system on opiate dependence. These results suggest that the CRF receptor antagonists, particularly the CRF receptor subtype 1 antagonist, might be of some value in the treatment and prevention of drug dependence. PMID- 11122351 TI - Acoustic startle and fear-potentiated startle in rats selectively bred for fast and slow kindling rates: relation to monoamine activity. AB - The acoustic startle response, prepulse inhibition, fear-potentiated startle and monoamine activity induced by either, a novel stimulus or a cue previously paired with foot-shock (fear-conditioning), were assessed in rats selectively bred for differences in amygdala excitability (Fast vs. Slow kindling epileptogenesis). Comorbid differences of anxiety, which were dependent both on the rats' behavioural style and the kind of stressor, also characterized these strains. In the present investigation, Slow rats exhibited a greater startle reflex to noise relative to Fast rats, suggesting differences in generalized anxiety, but similar rates of startle habituation and prepulse inhibition. The fear-potentiated startle, however, was greater in Fast rats. When movement of the rat was restricted in a new environment, presentation of a novel stimulus (light) increased norepinephrine, dopamine and/or serotonin activity in brain regions typically associated with stressors (e.g. locus coeruleus, paraventricular hypothalamic nucleus). Generally, these effects were more pronounced in Fast rats, and norepinephrine utilization in the central amygdala was particularly highlighted in response to a conditioned fear stimulus. Thus, while generalized anxiety appeared greater in Slow rats, behavioural and central neurochemical reactivity in response to novel stimuli and to fear-eliciting stimuli, was greater in Fast rats. Similarly, basal dopamine activity in the prefrontal cortex was greater in Fast rats, but dopamine utilization elicited by a novel stimulus was more pronounced in Slow rats. This suggested that relative to Slow rats, dopamine neurons in prefrontal cortex of Fast rats do not react normally to environmental stimuli, and this phenomenon could lead to disturbances of attention or impulsivity. PMID- 11122352 TI - Control of neurotransmission, behaviour and development, by photo-dynamic manipulation of tissue redox state of brain targets. AB - Reversible manipulation of local neurotransmission in brain areas, using controlled spatial and temporal resolution, is one of the powerful techniques used to investigate integrative aspects of brain function. We have developed a novel technique for rapidly inactivating local synaptic transmission, from outside the brain, within seconds or minutes via oxidation of target tissue using a photosensitive dye followed by photoirradiation (photo-dynamic tissue oxidation; PDTO). PDTO applied through a defined slit, sharply suppressed excitatory synaptic transmission in rat hippocampal slices and also suppressed in vivo hippocampal neurotransmission reversibly. Furthermore, we manipulated the voluntary movement of gerbils in free-field activity by application of PDTO to the striatum. Also, in freely moving kittens, the development of the visual cortex was manipulated by long-lasting application of PDTO to the eye. Thus, PDTO enables external manipulation of in vivo or in vitro neurotransmission in various clearly defined regions on the submillimeter scale. Suppression of neurotransmission occurred only within the photo-oxidized area which can be histochemically visualized. PMID- 11122353 TI - Differential DNA damage in response to the neonatal and adult excitotoxic hippocampal lesion in rats. AB - We examined the developmental profile of excitotoxin-induced nuclear DNA fragmentation using the transferase dUTP nick-end labelling (TUNEL) technique, as a marker of DNA damage and cell death in rats with neonatal and adult excitotoxic lesions of the ventral hippocampus. We hypothesized that infusion of neurotoxin may result in a differential pattern of cell death in neonatally and adult lesioned rats, both in the infusion site and in remote brain regions presumably involved in mediating behavioural changes observed in these animals. Brains of rats lesioned at 7 days of age and in adulthood were collected at several survival times 1-21 days after the lesion. In the lesioned neonates 1-3 days postlesion, marked increases in TUNEL-positive cells occurred in the ventral hippocampus, the site of neurotoxin infusion, and in a wide surrounding area. Adult lesioned brains showed more positive cells than controls only at the infusion site. In the lesioned neonates, TUNEL-labelled cells were also present in the striatum and nucleus accumbens 1 day postlesion but not at later survival times. Our findings indicate that cell death in remote regions is more prominent in immature than adult brains, that it may lead to distinct alterations in development of these brain regions, and thus may be responsible for functional differences between neonatally and adult lesioned rats. PMID- 11122354 TI - The vagus nerve mediates behavioural depression, but not fever, in response to peripheral immune signals; a functional anatomical analysis. AB - Cytokines act on the brain to induce fever and behavioural depression after infection. Although several mechanisms of cytokine-to-brain communication have been proposed, their physiological significance is unclear. We propose that behavioural depression is mediated by the vagus nerve activating limbic structures, while fever would primarily be due to humoral mechanisms affecting the preoptic area, including interleukin-6 (IL-6) action on the organum vasculosum of the laminae terminalis (OVLT) and induction of prostaglandins. This study assessed the effects of subdiaphragmatic vagotomy in rats on fever, behavioural depression, as measured by the social interaction test, and Fos expression in the brain. These responses were compared with induction of the prostaglandin-producing enzyme cyclooxygenase-2 and the transcription factor Stat3 that translocates after binding of IL-6. Vagotomy blocked behavioural depression after intraperitoneal injection of recombinant rat IL-1beta (25 microg/kg) or lipopolysaccharide (250 microg/kg; LPS) and prevented Fos expression in limbic structures and ventromedial preoptic area, but not in the OVLT. Fever was not affected by vagotomy, but associated with translocation of Stat3 in the OVLT and cyclooxygenase-2 induction around blood vessels. These results indicate that the recently proposed vagal link between the immune system and the brain activates limbic structures to induce behavioural depression after abdominal inflammation. Although the vagus might play a role in fever in response to low doses of LPS by activating the ventromedial preoptic area, it is likely to be overridden during more severe infection by action of circulating IL-6 on the OVLT or prostaglandins induced along blood vessels of the preoptic area. PMID- 11122355 TI - Role of interleukin-1beta and tumour necrosis factor-alpha in lipopolysaccharide induced sickness behaviour: a study with interleukin-1 type I receptor-deficient mice. AB - Interleukin-1 (IL-1) mediates symptoms of sickness during the host response to infection. IL-1 exerts its effects via several subtypes of receptors. To assess the role of IL-1 receptor type I (IL-1RI) in the sickness-inducing effects of IL 1, IL-1beta and the cytokine inducer lipopolysaccharide were administered to IL 1RI-deficient mice (IL-1RI-/-). Sickness was assessed by depression of social exploration, anorexia, immobility and body weight loss. IL-1RI-/- mice were resistant to the sickness-inducing effects of IL-1beta administered intraperitoneally (2 microg/mouse) and intracerebroventricularly (2 ng/mouse), but still fully responsive to lipopolysaccharide administered intraperitoneally (2.5 microg/mouse) and intracerebroventricularly (3 ng/mouse). The sensitivity of IL-1RI-/- mice to lipopolysaccharide was not due to a higher brain expression of proinflammatory cytokines other than IL-1, since lipopolysaccharide-induced expression of brain IL-1 beta, tumour necrosis factor-alpha (TNF-alpha) and IL-6 transcripts were identical in IL-1RI-/- and control mice when measured by semiquantitative reverse-transcriptase polymerase chain reaction 1 h after treatment. Blockade of TNF-alpha action in the brain by intracerebroventricular administration of a fragment of the soluble TNF receptor, TNF binding protein (3.6 microg/mouse), attenuated the depressive effects of intraperitoneal injection of lipopolysaccharide (1 microg/mouse) on behaviour in IL-1RI-/- but not in control mice. Since IL-1RI-/- mice were not more sensitive to intracerebroventricularly TNF-alpha (50 ng) than control mice, these results indicate that IL-1RI mediates the sickness effect of IL-1 and that TNF-alpha simply replaces IL-1 when this last cytokine is deficient. PMID- 11122356 TI - Effects of selective excitotoxic prefrontal lesions on acquisition of nonmatching and matching-to-place in the T-maze in the rat: differential involvement of the prelimbic-infralimbic and anterior cingulate cortices in providing behavioural flexibility. AB - The present study investigated the contributions of the medial prefrontal cortex and its major subdivisions, the dorsal anterior cingulate (ACd) and prelimbic infralimbic (PL) cortices, to spatial working memory and inhibitory control processes. In experiment 1, excitotoxic lesions centred in the ACd or PL cortex did not affect acquisition of a nonmatching-to-place task in the T-maze with a retention interval of 10 s. However, the same reinforced alternation task was impaired by larger prefrontal lesions that combined ACd and PL cortices. In experiment 2, new animals were trained on a matching-to-place task in the T-maze that uses a rule counter to the animals' innate bias to alternate spontaneously. Now, discrete lesions of both the ACd and PL cortices impaired acquisition, but in different ways. Both animals with PL and with ACd lesions perseverated by nonmatching for more sessions than the controls, but only the PL animals also showed a more general increase in perseveration reflected in a further, extended period of applying an inefficient response rule (e.g. always turn right) and a deficit at reversing from matching to nonmatching. Acquisition of the matching-to place task was also impaired by combined lesions of ACd and PL cortices. Overall, whilst spatial working memory processes appear to remain intact in those animals with discrete prefrontal lesions, the present findings provide strong evidence for the differential involvement of the prelimbic-infralimbic and anterior cingulate regions in providing behavioural flexibility. PMID- 11122357 TI - Fear conditioning in C57/BL/6 and DBA/2 mice: variability in nucleus accumbens function according to the strain predisposition to show contextual- or cue-based responding. AB - The contribution of the nucleus accumbens shell, the dorsal hippocampus, and the basolateral amygdala to contextual and explicit cue fear conditioning was assessed in C57BL/6 (C57) and DBA/2 (DBA) mice showing differences in processing contextual information associated with consistent but non-pathological variations in hippocampal functionality. Mice from both strains with bilateral ibotenic acid or sham lesions located in each area were introduced in a conditioning chamber and exposed twice to the pairing of a tone (2 x 8 s, 2000 Hz, 80 dB) with a shock (2 s, 0.7 mA). On the following day, mice were first exposed to the training context then to the tone in a different context. Freezing behaviour was scored in all situations. C57 showed more freezing to the context than to the tone whereas DBA showed more freezing to the tone than to the context. In C57, both nucleus accumbens and hippocampal lesions impaired acquisition of contextual fear conditioning but paradoxically improved acquisition of cue fear conditioning, whereas amygdala lesions disrupted performance in every task. In DBA, nucleus accumbens lesions, like amygdala lesions, impaired acquisition of both contextual and cue fear conditioning, whereas hippocampal lesions did not produce any effect. The parallelism between the effect of nucleus accumbens and hippocampus lesions in C57, and between the effect of nucleus accumbens and amygdala lesions in DBA points to a variability in nucleus accumbens function according to the strain specialization to develop context- or cue-based responding. PMID- 11122358 TI - Mapping of c-fos gene expression in the brain during morphine dependence and precipitated withdrawal, and phenotypic identification of the striatal neurons involved. AB - The c-fos gene is expressed in the central nervous system in response to various neuronal stimuli. Using in situ hybridization, we examined the effects of chronic morphine treatment and withdrawal on c-fos mRNA in the rat brain, and particularly within identified striatal neurons. Morphine dependence was induced by subcutaneous implantation of two pellets of morphine for 6 days and withdrawal was precipitated by administration of naltrexone. Placebo animals and morphine dependent rats showed a very weak c-fos mRNA expression in all the structures studied. Our study emphasized the spatial variations in c-fos mRNA expression, and also revealed a peak expression of c-fos mRNA at 1 h after naltrexone precipitated withdrawal in the projection areas of dopaminergic neurons, noradrenergic neurons and in several regions expressing opiate receptors. Interestingly, morphine withdrawal induces c-fos mRNA expression in the two efferent populations of the striatum (i.e. striatonigral and striatopallidal neurons) both in the caudate putamen and nucleus accumbens. Moreover, the proportions of activated neurons during morphine withdrawal are different in the caudate putamen (mostly in striatopallidal neurons) and in the shell and core parts of the nucleus accumbens (mostly in striatonigral neurons). The activation of striatopallidal neurons suggests a predominant dopaminergic regulation on c fos gene expression in the striatum during withdrawal. On the contrary, c-fos induction in striatonigral neurons during withdrawal seems to involve a more complex regulation like opioid-dopamine interactions via the mu opioid receptor and the D1 dopamine receptor coexpressed on this neuronal population or the implication of other neurotransmitter systems. PMID- 11122359 TI - APP is required during an early phase of memory formation. AB - The amyloid beta/A4 protein precursor (APP) has been shown to be implicated in age-associated plastic changes at synapses that might contribute to memory loss in Alzheimer's disease. As APP has previously been reported to have multiple functions during normal development, we have employed a one-trial passive avoidance task in day-old chicks to study its role in the process of memory formation. Administration of anti-APP antibodies, injected 30 min pretraining, prevented memory for a one-trial passive avoidance task in day-old chicks without effects on general behaviour or initial acquisition. Amnesia was apparent by 30 min post-training and lasted for at least 24 h. The same result was obtained by down-regulation of APP expression by APP-antisense, injected 8-12 h pretraining. However, injections of anti-APP antibodies or APP antisense at later post training time did not cause amnesia for the task. Unlike antibodies and antisense, injection of the APP328-332 pentapeptide, in either orientation, 30 min pretraining, rescued the memory and prevented antisense-induced amnesia. The post-training time within which the antibody- and antisense-induced amnesia, and within which the APP peptides prevent amnesia, correspond to that during which memory formation is vulnerable to disruption of the putative signal transduction functions of APP. These results suggest that: (i) APP is required during an early phase of memory formation, and (ii) the memory enhancing effect of secretory APP is localized within a 5-mer sequence of growth-promoting domain. PMID- 11122360 TI - Muscarinic and PACAP receptor interactions at pontine level in the rat: significance for REM sleep regulation. AB - Cholinergic and PACAPergic systems within the oral pontine reticular nucleus (PnO) play a critical role in REM sleep generation in rats. In this present work, we have investigated whether REM sleep enhancement induced by carbachol (a cholinergic agonist) or PACAP, depends on an interaction between muscarinic and PACAP receptors. This hypothesis was tested by recording sleep-wake cycles in freely moving rats injected into the PnO with PACAP in combination with the muscarinic receptor antagonist atropine, or with carbachol in combination with the PACAP receptor antagonist PACAP6-27. When administered alone, PACAP (3 pmol) or carbachol (110 pmol) induced an enhancement of REM sleep during 8 h (+61%, n = 8; +70%, n = 5), which was totally prevented by infusion of atropine (290 pmol) for PACAP, or of PACAP6-27 (3 pmol) for carbachol. Quantitative autoradiographic studies indicated that (i) PACAP (10-9-10-7 M) induced in the PnO an increase (+35%) of the specific binding of the muscarinic antagonist [3H]quinuclidinyl benzylate, which could be completely prevented by PACAP6-27 (IC50 = 8 x 10-8 M) and (ii) both carbachol and PACAP enhanced [35S]GTP-gamma-S binding in a concentration-dependent manner in the PnO. The maximal increase due to carbachol was significantly higher in the presence (+126%) than in the absence (+102%) of PACAP (0.1 microM). These data showed that interactions between muscarinic and PACAP receptors do exist within the PnO and play a role in the local mechanisms of REM sleep control in the rat. PMID- 11122361 TI - Cortical cholinergic inputs mediate processing capacity: effects of 192 IgG saporin-induced lesions on olfactory span performance. AB - An olfactory span task that required rats to discriminate an olfactory stimulus added to an increasing list of such stimuli (nonmatching-to-sample; NMTS) was employed to assess the role of the basal forebrain cholinergic system in the animals' olfactory working memory capacity. A separate group of animals was trained in a matching-to-sample (MTS) version of this task that did not tax span performance. NMTS animals required significantly more sessions to reach an olfactory span of 18 stimuli than MTS rats. Infusions of the cholino-immunotoxin 192 IgG-saporin into the basal forebrain resulted in decreases of cortical acetylcholinesterase (AChE)-positive fibre density ranging from 80% in frontodorsal and frontoparietal regions to 35% in the pyriform cortex and 24% in the olfactory bulb. Postsurgery span performance was significantly reduced in lesioned NMTS but not MTS animals. Span performance in lesioned NMTS animals recovered following 4 weeks of postoperative training; however, these animals' span remained vulnerable to the effects of increased intertrial intervals. The distribution of errors in lesioned animals indicated a recency effect. In NMTS animals, olfactory span performance during the initial two postoperative weeks correlated significantly with AChE-positive fibre density in neocortical but not olfactory areas. The privileged, automatic processing of olfactory stimuli in rats may have contributed to the transience of the lesion effect. The results support the crucial role of cortical cholinergic input in the mediation of aspects of processing capacity. PMID- 11122362 TI - Effect of feeding on Fos protein expression in sheep hypothalamus with special reference to the supraoptic and paraventricular nuclei: an immunohistochemical study. AB - The hypothalamus plays an important role in the control of food intake in different species, but there is little relevant information for ruminants like sheep. In order to study the putative role of several hypothalamic nuclei in food intake in sheep, Fos expression, a marker of cellular activity, was compared by immunohistochemistry between fed and unfed ewes. The expression of Fos protein was stimulated in the supraoptic nucleus of fed ewes, whereas it was increased in the paraventricular nucleus of unfed animals. In the latter nucleus, Fos immunoreactivity was mainly localized close to the third ventricle, an area corresponding to the parvocellular system of the nucleus, but never in the magnocellular system. In the paraventricular nucleus, the number of corticotrophin releasing factor-immunoreactive neurons and the number of Fos/corticotrophin releasing factor double-labelled neurons were not affected by feeding or lack of feeding. The number of Fos-immunoreactive neurons was higher in the lateral septum, the infundibular, the ventromedial and in the dorsomedial nuclei of unfed ewes than in those of fed ewes. Our results show for the first time that the dorsomedial and ventromedial nuclei are involved in the control of feeding in sheep as in rodents. The supraoptic nucleus of sheep is activated by the same conditions as in rodents but, conversely, the paraventricular nucleus is activated in unfed sheep, whereas in rodents and primates, this nucleus is activated by satiety as well as by fasting. In sheep, unlike in rodents, corticotrophin releasing factor did not appear to be involved in short-term regulation of food intake. PMID- 11122363 TI - Dopaminergic innervation of the pallidum in the normal state, in MPTP-treated monkeys and in parkinsonian patients. AB - The aim of the present study was to characterize the dopaminergic innervation of the pallidum in primates (humans and Cercopithecus aethiops). Firstly, in monkeys, biotin dextran amine was injected into dopaminergic areas, and the anterogradely labelled axons were reconstructed from serial sections and analysed in the pallidum. Secondly, in parkinsonian patients and MPTP-treated monkeys, the dopaminergic innervation of the pallidum was studied using tyrosine hydroxylase positive fibre quantification. Our study revealed that dopaminergic areas A8 and A9 innervated the two pallidal segments. Individual axonal arborizations displayed a great heterogeneity. Some dopaminergic axons crossed the pallidum without branching, other axons made small terminal arborizations in a restricted region of one pallidal segment, whereas others developed dense arborizations covering extended areas in the two pallidal segments. This heterogeneous organization suggests that dopamine could directly modulate the pallidum using either a point-to-point or a diffuse projection pattern. A statistically significant loss of dopaminergic fibres in the internal (-43%) and external pallidum (-39.6%) of humans, and in the internal (-54.3%) and external pallidum ( 59%) of monkeys was revealed in parkinsonian states. The consequences of this alteration are still unknown but it might participate in the triggering of motor symptoms observed in Parkinson's disease. PMID- 11122364 TI - Migrating neurons cross a reelin-rich territory to form an organized tissue out of embryonic cortical slices. AB - In this study we show that the radial migration of neuronal precursors out of cerebral cortex of embryonic brain slices cultured for 4-7 days gives rise to an organized tissue that forms de novo off developing slices. In our in vitro preparations, migrating neuronal precursors overshot the marginal zone, as did the elongation of radial glial processes out of the slices. These cells detached from radial glia at a distance from the cortex and differentiated into pyramidal and nonpyramidal profiles that expressed different neuronal markers. Glial precursors were shown to proliferate in the slice and in the neotissue, and to differentiate into astrocytes. We show that cells expressing reelin in the marginal zone of embryonic cortical slices persist after a week in culture, which implies that neuronal migration is not necessarily hindered by the presumed stop signals provided by reelin in the marginal zone. Furthermore, our results provide a new model for in vitro studies of migration and differentiation during cortical development. PMID- 11122365 TI - Vitamin E supplementation prevents spatial learning deficits and dendritic alterations in aged apolipoprotein E-deficient mice. AB - Recent studies have suggested that altered function of apolipoprotein E might lead to Alzheimer's disease via oxidative stress. In this context, the objective of this study was to determine if antioxidative treatment with vitamin E was neuroprotective in apolipoprotein E-deficient mice. For this purpose, 1-month-old control and apolipoprotein E-deficient mice received dietary vitamin E for 12 months. We showed that, compared to apolipoprotein E-deficient mice who received a regular diet, mice treated with vitamin E displayed a significantly improved behavioural performance in the Morris water maze. This improved performance was associated with preservation of the dendritic structure in vitamin E-treated apolipoprotein E-deficient mice. In addition, whilst untreated apolipoprotein E deficient mice displayed increased levels of lipid peroxidation and glutathione, vitamin E-treated mice showed near normal levels of both lipid peroxidation and glutathione. These results support the contention that vitamin E prevents the age related neurodegenerative alterations in apolipoprotein E-deficient mice. PMID- 11122366 TI - Interactions between imidazoline binding sites and dopamine levels in the rat nucleus accumbens. AB - Imidazoline binding sites are present in the striatal complex and in the extended amygdala and have been implicated in mood disorders. In this report we analysed the influence of these sites on the functional activity of the mesolimbic dopaminergic transmission, one of the major brain systems involved in the regulation of motivation and reward. We studied the effects of two imidazoline ligands, S23229 and S23230 (respectively S(+) and R(-) enantiomers of the S22687 or (5-[2-methyl phenoxy methyl] 1,3-oxazolin-2-yl) amine), on extracellular dopamine in the nucleus accumbens using microdialysis in freely moving rats. We compared these imidazoline ligands to cocaine, a dopamine uptake blocker known to increase extracellular dopamine concentrations. S23229 dose-dependently increased extracellular dopamine and locomotor activity. S23230 dose-dependently increased extracellular dopamine and produced a near-significant dose-effect on locomotor activity. S23229 had a stronger efficacy than S23230 and increased dopamine levels in the nucleus accumbens at an extent similar to the one of cocaine. These results suggest that central imidazoline binding sites could contribute to the functional regulation of the mesolimbic dopaminergic system. PMID- 11122367 TI - Increased expression of neuronal nitric oxide synthase mRNA in the accessory olfactory bulb during the formation of olfactory recognition memory in mice. AB - The mouse accessory olfactory bulb contains a high density of nitric oxide synthase and, in females, is involved in the formation of a mating and pheromone specific recognition memory. The exact role of nitric oxide in this memory model is not yet clearly understood. In this study, in situ hybridization was used to assess neuronal nitric oxide synthase mRNA expression during the critical interval associated with synaptic plasticity in the accessory olfactory bulb of female mice leading to the formation of a recognition memory for the stud male pheromones present following mating. Nitric oxide synthase mRNA was significantly increased following mating and 120-min stud male exposure compared with oestrus mice. The mRNA expression was more predominant in the anterior than the posterior regions of the bulb. These observations indicate a stimulus-specific activation of nitric oxide gene expression in the female mouse accessory olfactory bulb and support the hypothesis that nitric oxide may modulate intermediate synaptic pathways during the formation of a pheromone-dependent olfactory recognition for stud males. PMID- 11122368 TI - Differential adrenergic regulation of the circadian expression of the clock genes Period1 and Period2 in the rat pineal gland. AB - Precise temporal regulation of transcription is pivotal to the role of the mammalian pineal gland as a transducer of circadian and seasonal information. The circadian clock genes Per1 and Per2 encode factors implicated in temporally gated transcriptional programmes in brain and pituitary. Here we show that the nocturnal circadian expression of Per1 and Per2 in the rat pineal gland parallels that of serotonin N-acetyltransferase (NAT) mRNA, which encodes the rate-limiting enzyme of melatonin biosynthesis. This rhythm is dependent upon an intact sympathetic innervation. Increases in rPer1 (r indicates rat) and rPer2, as well as rNAT, expression during subjective night were blocked completely by superior cervical ganglionectomy (SCGX). In SCGX rats, the beta-adrenergic receptor agonist isoproterenol rapidly induced the rPer1 mRNA with dynamics very similar to its effect on rNAT mRNA. In contrast, isoproterenol was without effect on expression of rPer2 mRNA. These findings demonstrate that circadian pineal expression of both rPer1 and rPer2 is controlled by sympathetic afferent innervation, but whereas beta-adrenergic signalling regulates rPer1 and rNAT, an alternative route mediates sympathetic regulation over rPer2 expression. PMID- 11122369 TI - Differential maturation of motoneurons innervating ankle flexor and extensor muscles in the neonatal rat. AB - The first postnatal week is a critical period for the development of posture in the rat. The use of ankle extensor muscles in postural reactions increases during this period. Changes in excitability of motoneurons are probably an important factor underlying this maturation. The aim of this study was to identify whether variations in the maturation exist between motor pools innervating antagonistic muscles. Intracellular recordings in the in vitro brain stem-spinal cord preparation of neonatal rats (from postnatal day 0-5) were used to examine the developmental changes in excitability of motoneurons innervating the ankle flexors (F-MNs) and the antigravity ankle extensors (E-MNs). No significant difference in resting potential, action potential threshold, input resistance or rheobase was observed at birth. The age-related increase in rheobase was more pronounced for F-MNs than for E-MNs. The development of discharge properties of E MNs lagged behind that of F-MNs. More F-MNs than E-MNs were able to fire repetitively in response to current injection at birth. F-MNs discharged at a higher frequency than E-MNs at all ages. Differences in the duration of action potential afterhyperpolarization accounted, at least partly, for the differences in discharge frequency between E-MNs and F-MNs at birth, and for the age-related increase in firing rate. These results suggest that E-MNs are more immature at birth than F-MNs and that there is a differential development of motoneurons innervating antagonistic muscles. This may be a critical factor in the development of posture and locomotion. PMID- 11122370 TI - A brain slice model for in vitro analyses of astrocytic gap junction and connexin43 regulation: actions of ischemia, glutamate and elevated potassium. AB - Brain slices prepared from adult rats and maintained for up to 3 h in vitro were used to investigate the effects of pharmacological treatments on the phosphorylation state, immunolabelling characteristics and ultrastructural localization of astrocytic gap junctions and connexin43 (Cx43). Slices deprived of glucose/oxygen to mimic ischemia or those exposed to 1 mM glutamate for 1 h exhibited Cx43 dephosphorylation, epitope masking and gap junction internalization as revealed by Western blotting and Cx43 immunolocalization with various antibodies. Treatment with 15 mM K+ caused Cx43 dephosphorylation without junction internalization. The effects of glutamate and K+ were completely blocked by the N-methyl-D-aspartate (NMDA) glutamate receptor antagonist 2-amino-5 phosphonovalerate (APV), which acts largely on neuronal NMDA receptors, suggesting neuronal mediation of glial gap junction responses to these treatments. Astrocytes contained a dephosphorylated form of Cx43 with a typical migration profile at 41 kDa as well as novel, apparently dephosphorylated or partially phosphorylated, forms migrating at 43 kDa. These results indicate that slices prepared from adult brain can serve as a convenient model to investigate the molecular basis and receptor-mediated mechanisms underlying astrocytic Cx43 responses that have been observed in vivo following cerebral ischemia or neural activation. These processes can be related in part to neuronal regulation of astrocytic gap junctional coupling state, which is also amenable to analysis in brain slices. PMID- 11122371 TI - Identification of the target neuronal elements in electrical deep brain stimulation. AB - The aim of this study is to identify the primary neuronal target elements in electrical deep-brain stimulation, taking advantage of the difference in strength duration time constant (tau(sd)) of large myelinated axons ( approximately 30-200 micros), small axons ( approximately 200-700 micros) and cell bodies and dendrites ( approximately 1-10 ms). Strength-duration data were measured in patients suffering from Parkinson's disease or essential tremor and treated by high-frequency stimulation in the ventral intermediate thalamic nucleus or the internal pallidum. Threshold voltages for the elimination of tremor were determined at various pulsewidths and a pulse rate of 130 pulses per second. The tau(sd) was calculated using Weiss's linear approximation. Its mean value was 64.6+/-25.4 micros (SD) for the thalamic nucleus and 75.3+/-25.5 micros for the internal pallidum. Corrections to the mean values were made because the tau(sd) values were based on voltage-duration measurements using polarizable electrodes. Apart from this systematic error, a resolution error, due to the relatively large increment steps of the pulse amplitude, was taken into account, resulting in mean tau(sd) estimates of 129 and 151 micros for the thalamic nucleus and the internal pallidum, respectively. It is concluded that the primary targets of stimulation in both nuclei are most probably large myelinated axons. PMID- 11122372 TI - Differential calbindin-immunoreactivity in dopamine neurons projecting to the rat striatal complex. AB - The calcium-binding protein calbindin-D28K is an anatomical marker that has been associated with resistance to neurodegeneration and with the electrophysiological characteristics of neurons. In this study, we compared the presence of calbindin in dopamine neurons projecting to three distinct functional regions of the striatal complex: the striatum, and the core and the shell of the nucleus accumbens. After iontophoretic injections of Fluoro-Gold in the dopaminergic terminal fields, the presence of tyrosine hydroxylase and calbindin were immunohistochemically assessed in the mesencephalon. It was found that the proportion of cells expressing calbindin was highest in the dopamine cells projecting to the core (72%), intermediate in the cells projecting to the shell (51%) and lowest in the cells projecting to the dorsolateral striatum (2.6%). These results do not support the idea that calbindin is a sufficient condition to confer resistance to neurodegeneration because shell-projecting neurons seem the most resistant to it. The present data also raise the question of the role of calbindin in the differences in firing characteristics among dopamine neurons projecting to the striatal complex. PMID- 11122373 TI - Morphological evidence for selective modulation by serotonin of a subpopulation of dorsal horn cells which possess the neurokinin-1 receptor. AB - Serotonin selectively depresses transmission of nociceptive information through the spinal dorsal horn but the mechanisms of this depression are poorly understood. In this study we report that serotonin-containing axons form basket like clusters which are intimately woven around cell bodies and proximal dendrites of a subpopulation ( approximately 50%) of laminae III/IV neurons which possess the neurokinin-1 receptor. Statistical analysis confirms that cells belonging to this subpopulation have significantly higher numbers of serotoninergic contacts on proximal dendrites when compared with the population of neurokinin-1 cells that are not associated with clusters (mean +/- SD = 13+/ 5.8 and 5+/-2.9, respectively). Neurokinin-1 cells in laminae III/IV project to regions of the brain which are involved in nociceptive processing and are likely to be activated predominantly by nociceptive input. The concentration of serotoninergic axons around proximal regions of some of these cells indicates that serotonin may have a powerful influence on transmission through this pathway. This type of arrangement could be a morphological correlate for at least part of the selective antinociceptive actions of serotonin. PMID- 11122374 TI - Activation of ionotropic glutamate receptors reduces the production of transforming growth factor-beta2 by developing neurons. AB - Neuronal cultures derived from developing rat cerebral cortex were used to investigate the influence of glutamate receptors on the neuronal production of transforming growth factor-beta2 (TGFbeta2), a multifunctional cytokine that modulates neuronal and glial growth. Long-term exposure (48 h) of cortical neurons to selective antagonists of N-methyl-D-aspartate (NMDA) and alpha-amino-3 hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors markedly increased TGFbeta2 levels in the culture medium. Conversely, treatment with NMDA or kainate reduced TGFbeta2 to levels below those in untreated cultures. The effect of kainate did not require NMDA receptor activity. Neuronal depolarization with K+ also reduced TGFbeta2 levels by opening voltage-gated L-type Ca2+ channels. Semi-quantitative RT-PCR measurements of neuronal TGFbeta2 mRNA showed that NMDA or AMPA/kainate receptor stimulation reduced TGFbeta2 mRNA levels. These results demonstrate that tonic activation of glutamate-gated cation channels downregulates neuronal expression of the TGFbeta2 gene and provide evidence for a novel mechanism whereby excitatory amino acids could influence the development of glial and neuronal lineages. PMID- 11122375 TI - Competition between the replication initiator DnaA and the sequestration factor SeqA for binding to the hemimethylated chromosomal origin of E. coli in vitro. AB - BACKGROUND: Following replication initiation, the replication origin (oriC) in Escherichia coli enters a hemimethylated state at Dam methylation sites which are recognized by the SeqA protein. SeqA binds preferentially to hemimethylated GATC sequences of DNA in vitro. SeqA is essential for the synchronous initiation of chromosome replication from oriC copies in vivo. RESULTS: We show that: (i) purified SeqA binds AT-rich and 13-mers regions and two DnaA boxes, R1 and M, of hemimethylated oriC. (ii) SeqA inhibits the in vitro replication of a hemimethylated oriC plasmid more efficiently than the fully methylated, (iii) SeqA inhibits competitive binding of DnaA protein to the regions of the hemimethylated oriC plasmid, explaining the mechanism of its inhibitory effect. The inhibition of DnaA binding by SeqA also occurs efficiently on a small hemimethylated oriC fragment containing both R1 and M DnaA boxes, but not the 13 mer region. CONCLUSIONS: SeqA binds strongly the long region from the AT-rich region to the M DnaA box of the hemimethylated oriC DNA and releases DnaA molecules from the long region. PMID- 11122376 TI - MSI-1, a neural RNA-binding protein, is involved in male mating behaviour in Caenorhabditis elegans. AB - BACKGROUND: Neural RNA-binding proteins are thought to play important roles in neural development and the functional regulation of postmitotic neurones by mediating post-transcriptional gene regulation. RNA-binding proteins belonging to the Musashi family are highly expressed in the nervous system; however, their roles are poorly understood. RESULTS: We identified a Caenorhabditis elegans Musashi homologue, MSI-1, whose RNA-recognition motifs show extensive similarity to those of Drosophila and vertebrate Musashi proteins. We isolated a msi-1 mutant and found males with this mutation to have a mating defect. C. elegans male mating behaviour includes a distinct series of steps: response to contact, backing, turning, vulva location, spicule insertion, and sperm transfer. msi-1 is required for the turning and vulva location steps. Like other Musashi family members, MSI-1 is expressed specifically in neural cells, including male-specific neurones required for turning and vulva location. However, msi-1 was not expressed in proliferating neural progenitors in C. elegans, unlike the Musashi family genes in other systems. CONCLUSIONS: Our results suggest that msi-1 is expressed specifically in postmitotic neurones in C. elegans. msi-1 is required for full development of male mating behaviour, possibly through regulation of msi 1 expressing neurones. PMID- 11122377 TI - Expression and characterization of nonmammalian selenoprotein P in the zebrafish, Danio rerio. AB - BACKGROUND: Selenoprotein P is a protein of considerable intrigue, due to its unusual composition and requirements for its biosynthesis. Whereas most selenoproteins contain a single selenocysteine residue, the human, bovine and rodent selenoprotein P genes encode proteins containing 10-12 selenocysteines. Selenoprotein P genes have, to date, only been reported in mammals, and the function of the protein remains elusive. RESULTS: Herein, we report the identification and characterization of nonmammalian selenoprotein P in the zebrafish Danio rerio. Sequencing of the cDNA revealed the presence of 17 selenocysteine codons, the highest number reported in any protein. Two histidine rich regions present in the mammalian selenoprotein P sequences are conserved in the zebrafish protein, and two SECIS elements are present in the 3' untranslated region. Whole-mount in situ hybridization of zebrafish embryos revealed high levels of expression of selenoprotein P mRNA in fertilized eggs and in the yolk sac of developing embryos. Transient transfection of the cDNA in mammalian cells resulted in efficient expression of the full-length secreted selenoprotein. A single N-glycosylation site is predicted, and shown to be utilized. CONCLUSIONS: Discovery of selenoprotein P in the zebrafish opens a previously unavailable avenue for genetic investigation of the functions of this unusual protein. PMID- 11122378 TI - The hDLG-associated protein DAP interacts with dynein light chain and neuronal nitric oxide synthase. AB - BACKGROUND: Postsynaptic density (PSD)-95 interacts with and mediates clustering of the N-methyl-D-aspartate-receptors (NMDA-R). PSD-95 also interacts with the hDLG-associated protein DAP, which is also called Synapse-associated protein 90 associated protein (SAPAP), and Guanylate kinase-associated protein (GKAP). RESULTS: DAP interacted directly with the dynein light chain (DLC) family of proteins. DLC was contained in the NMDA-R-PSD-95-DAP-neuronal nitric oxide synthase (nNOS) complex. Furthermore, DAP interacted with nNOS and recruited it into the Triton X-100-insoluble fraction of transfected cells. CONCLUSION: DAP interacts directly with DLC and nNOS, and links these proteins to the NMDA-R-PSD 95 complex. PMID- 11122380 TI - Multiple downstream signalling pathways from ROCK, a target molecule of Rho small G protein, in reorganization of the actin cytoskeleton in Madin-Darby canine kidney cells. AB - BACKGROUND: In Madin-Darby canine kidney cells, Rho small G protein regulates formation of stress fibres, focal adhesions, and peripheral bundles through reorganization of the actin cytoskeleton. There are two morphologically distinguishable types of Rho-regulated stress fibres: parallel and stellate. Of these, effects of Rho small G protein, mDia1 regulates the formation of parallel stress fibres, whereas ROCK regulates the formation of stellate stress fibres, peripheral bundles and focal adhesions. Both mDia1 and ROCK are direct downstream targets of Rho small G protein. RESULTS: The ROCK-induced formation of stellate stress fibres is regulated mainly through the myosin light chain kinase-dependent phosphorylation of myosin light chain and the LIM-kinase-dependent phosphorylation of cofilin. The ROCK-induced formation of focal adhesions is mainly regulated through a downstream pathway of ROCK other than myosin light chain and cofilin. The ROCK-induced formation of peripheral bundles is regulated at least through ERM proteins, but not through the myosin light chain or cofilin. CONCLUSION: Our present and previous findings suggest the presence of multiple downstream signalling pathways from ROCK to reorganization of the actin cytoskeleton in Madin-Darby canine kidney cells. PMID- 11122379 TI - Extracellular matrix tenascin-X in combination with vascular endothelial growth factor B enhances endothelial cell proliferation. AB - BACKGROUND: An extracellular matrix tenascin-X (TNX) is highly expressed in muscular tissues, especially heart and skeletal muscle, and is also prominent around blood vessels. The precise in vivo role of TNX remains to be elucidated. To identify proteins that interact with TNX in the extracellular environment, we searched for TNX-binding proteins using a yeast two-hybrid system. RESULTS: We used mouse TNX-specific fibronectin type III repeats (mTNX/FNIII13-25) as a bait for the screening. We found that vascular endothelial growth factor B (VEGF-B) binds to mTNX/FNIII13-25. This interaction was confirmed by pull-down assays and co-immunoprecipitation assays. The full-length mTNX, as well as mTNX/FNIII13-25, interacted with both alternative splice isoforms VEGF-B186 and VEGF-B167. Furthermore, the full-length mTNX also bound to VEGF-A. The minimal region of TNX that interacts with VEGF-B was mapped to the FNIII repeats (FNIII13-25) but not to the other characteristic domains of TNX. The TNX-binding site of VEGF-B was located in the N-terminal 115-amino acid region. mTNX/FNIII13-25 did not prevent the interaction of VEGF-B with VEGFR-1 (VEGF receptor 1), and VEGF-B could simultaneously bind to both mTNX/FNIII13-25 and VEGFR-1. A conditioned medium from transfected 293T cells coexpressing full-length TNX and VEGF-B could promote DNA synthesis in bovine endothelial cells in which VEGFR-1 were expressed. VEGFR 1 phosphorylation triggered by VEGF-B186 were increased in cells plated with mTNX/FNIII13-25 or full-length mTNX, compared with cells plated with VEGF-B186 alone. CONCLUSION: TNX interacts with VEGF-B and enhances the ability of VEGF-B to stimulate cell proliferation. This enhanced mitogenecity is caused by increased signals mediated by the VEGFR-1 receptor. This finding suggests a role for TNX in the regulation of the development of blood vessels such as vasculogenesis and angiogenesis. PMID- 11122381 TI - In vitro analysis of STAT5 activation by granulocyte-macrophage colony stimulating factor. AB - BACKGROUND: The granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor activates multiple and complex signalling pathways in response to GM-CSF stimulation. Biochemical studies suggested that signalling pathways are transmitted through protein/protein interactions, but how these biochemical cascades are initiated and transmitted in response to cytokine stimulation is largely unknown. RESULTS: To investigate these events biochemically, we established an in vitro system leading to the GM-CSF-dependent activation of Janus kinase (JAK) 2/signal transducer and activator of transcription (STAT) 5 in cell homogenates prepared from BA/F3 cells expressing the GM-CSF receptor. Activation of STAT5 DNA binding ability requires both membrane and cytoplasmic fractions while phosphorylation of JAK2 requires only the membrane fraction. Since the addition of anti-betac or phosphotyrosine antibodies inhibited GM-CSF induced STAT5 DNA binding activity, we examined the role of tyrosine residues of betac for in vitro activation of STAT5. Addition of synthetic tyrosine phosphorylated peptides derived from betac cytoplasmic tyrosines prior to GM-CSF stimulation inhibited the in vitro activation of STAT5. The association between these tyrosine-phosphorylated peptides and STAT5 was observed by using peptide coupling beads and BA/F3 lysates. CONCLUSIONS: We established a GM-CSF-dependent in vitro system. In cases of STAT5 activation, each phosphorylated tyrosine residue of betac can act as a docking site and enhance STAT5 activation. PMID- 11122382 TI - Virucidal heat-treatment of single plasma units: a potential approach for developing countries. AB - Since HIV first burst onto the scene of transfusion medicine, the quest for viral inactivation processes for plasma and plasma products has not ceased. Sophisticated methods for improving viral safety are currently used in the industrial world. However, in developing countries, with no facilities for treating plasma, nonviral-inactivated fresh frozen plasma [FFP] continues to be used extensively, and as screening may not be optimal (or may even be absent), FFP still contributes to the spread of HIV and other infectious viruses. The feasibility of heat-treating FFP at the liquid state, in its collection bag, was explored by testing diverse conditions of temperature and duration, in the presence of biologically compatible stabilisers. Quality of the heat-treated plasma was evaluated by haematological, biochemical and animal assays. The efficiency of the method to inactivate viruses was validated using HIV and model viruses. The selected heating conditions are 50 degrees C for 3 h. The optimized combination of stabilizers is composed of 30 mM trisodium citrate, 10 g L-1 L lysine, 12 mM calcium gluconate and 150 g L-1 sorbitol. Plasma coagulability is appropriately preserved as shown by the KCT ratio (1.4). Recovery of biological activity of most coagulation factors is higher than 70% (including fibrinogen & von Willebrand factor). Electrophoretic and immunoblotting studies did not evidence protein aggregation and/or degradation. Viral validation studies of this procedure have shown complete inactivation of HIV (> 6.6 log) in less than 1 h of treatment. A viral reduction of at least 4 log for various model viruses, including those of hepatitis A and C viruses, suggests a potential contribution of the method to diminish the risk from various blood-borne viruses. The selected formulation appears to preserve plasma protein integrity and properties. The procedure does not require sophisticated equipment but it is mandatory to monitor it carefully to ensure quality and reproducibility. If properly controlled and standardized, this approach offers an opportunity to reduce the risk of transmission of HIV and other viruses, particularly in poor countries with a high incidence of HIV. PMID- 11122383 TI - Treatment of chronic hepatitis C in haemophilia patients. PMID- 11122385 TI - Treatment of children with haemophilia in Europe: a survey of 20 centres in 16 countries. AB - A survey was made of the current status of treatment of haemophilic boys at 20 centres in 16 European countries and includes approximately 1500 of the estimated 6500 haemophiliacs in the participating countries. Many mild haemophiliacs are not seen, or seen infrequently, at haemophilia centres and this requires study. Nine of 18 centres provide continuous prophylaxis to 80-100% of their patients, five centres provide it to 55-80% and the remaining four centres to 15-40% of the boys. The median dose given was 6240 U kg-1 year-1 (range 3120-7800). Four centres administered only recombinant concentrates to children with severe haemophilia A, while seven centres administered recombinant concentrates to 75 90% and the remaining centres to less than 50% of the boys (two centres < 10%). When asked for the choice of concentrate for a newly diagnosed boy with severe haemophilia A, all but one centre preferred recombinant concentrate. Most boys below 6 years received concentrates via a peripheral vein but three centres preferred a central venous line for 80-100% of the boys. Thirteen of 18 centres applied home treatment to 84-100% of the boys and the remaining five centres to 57-77% of the boys. PMID- 11122384 TI - Efficacy of a sucrose-formulated recombinant factor VIII used for 22 surgical procedures in patients with severe haemophilia A. AB - A sucrose-formulated recombinant FVIII (rFVIII-SF) was investigated under clinical trial conditions during surgical procedures in previously treated patients (PTPs). Fifteen PTPs with severe haemophilia A (FVIII < or = 1%) underwent 22 surgical procedures. The procedures performed cover a spectrum from minor to major surgery. Haemostatic outcome was assessed by the investigators to be excellent in 16 procedures and good in the remaining six procedures. It is concluded that rFVIII-SF is efficacious and safe in severe haemophilia A patients undergoing minor or major surgery. PMID- 11122386 TI - Factor VIII gene polymorphisms in the Asian Indian population. AB - Little is known about the heterozygous frequency of factor VIII gene markers in the Asian Indian population. The objective of this study was to establish the heterozygous frequency of polymorphic markers within and flanking the factor VIII gene in Indians and identify those most informative for carrier screening and prenatal diagnosis. Factor VIII gene polymorphism analysis at intragenic and extragenic sites was carried out by the polymerase chain reaction (PCR) method and Southern blot procedure. Sixty-three Asian Indian haemophiliacs and their families were screened. A control group of 150 women from nonhaemophilic families were screened for two markers, HindIII and BclI. Among the intragenic markers studied, the HindIII restriction fragment length polymorphism (RFLP) showed the highest heterozygous frequency (0.52) followed by the intron 13 (0.47) and intron 22 (0. 44) short tandem repeats (STRs). Among extragenic markers, TaqI had the highest heterozygous frequency (0.75) followed by BglII (0.54). The intron 22 inversion mutation was observed in eight (40%) of 20 severe cases. In the population studied the most diagnostic polymorphisms were the intragenic markers, intron 22 (70%) STR followed by the intron 13 (52%) STR and HindIII (52%) RFLP, and the TaqI (50%) extragenic marker. Application of HindIII, BclI and the intron 22 dinucleotide repeat combined were diagnostic in 87.2% of haemophilia A families studied. PMID- 11122387 TI - Venous thromboembolism after hip fracture surgery in a patient with haemophilia B and factor V Arg506Gln (factor V Leiden). AB - We describe a patient with mild haemophilia B who developed symptomatic venous thromboembolism after hip arthroplasty for a traumatic fracture. A deep vein thrombosis developed in the operated leg while he was receiving a high-purity factor IX concentrate. Subsequently, he was determined to be a heterozygous carrier for the factor V Arg506Gln (Leiden) mutation. This case illustrates the importance of providing thromboprophylaxis for all patients with haemophilia receiving coagulation factor replacement and who undergo surgical procedures known to be associated with a high risk of venous thromboembolism. In patients with haemophilia and a family history of venous thromboembolism, preoperative screening for the presence of the factor V Arg506Gln mutation and other thrombophilias may be useful. PMID- 11122388 TI - Frequency of inhibitor development in severe haemophilia A children treated with cryoprecipitate and low-dose immune tolerance induction. AB - The frequency of factor VIII inhibitor development was evaluated in a hundred severe haemophilia A patients < 18 years of age (mean 10.4 +/- 5.1 years); 25 were previously untreated patients (PUPs), with a mean age of 11.2 +/- 2.9 months. All were followed up for 3 years from December 1996. Immune tolerance (IT) was induced with low-dose factor VIII (FVIII); 25-50 IU kg(-1) every other day for the 10 haemophiliacs who developed persistent inhibitors. The incidence of inhibitors for PUPs was 3/25 (12%; 95% confidence interval [CI], 0. 7-24.7%) and were detected after 4, 15 and 20 exposure days (mean 13 +/- 8.2 days; 95% CI, 3.7-22.2%). Children with maximum inhibitor levels of > 40 Bethesda units (BU) per mL (n=4) received IT therapy as 25 U kg(-1) FVIII in the form of cryoprecipitate every other day for 1-4 months (mean 2.4 +/- 1.6 months; 95% CI, 0.8-3.9%), which was successful in all of them. FVIII (50 U kg(-1)) was given every other day for six patients with maximum inhibitor level > 40 BU mL(-1) for 3-9 months (mean 5.4 +/- 3.2 months; 95% CI, 2.9 -7.9%) with success in 4/6 (66.6%; 95% CI, 28.8-104.3%). Patients who showed a good IT response had an inhibitor level < or = 30 BU mL(-1), were < or = 9 years of age at inhibitor development with few exposure days to FVIII and had an early immune tolerance. In conclusion, inhibitor development in severe haemophilia A children exclusively treated with cryoprecipitate is low. Early low-dose IT induction for high responders may be achieved successfully if inhibitor level is < or = 50 BU mL( 1). PMID- 11122389 TI - The Malmo model for immune tolerance induction: impact of previous treatment on outcome. AB - Ten patients, who had been treated according to the Malmo model for immune tolerance induction (ITI), were analysed regarding treatment with clotting factors and chemotherapy during the time period between inhibitor detection and ITI. Of the patients who were successfully rendered tolerant (n=6) all but one had received treatment with FVIII, either alone (n=1), or combined with cyclophosphamide (n=4). Of the patients who did not become tolerant, three of four had received treatment with FVIII during the inhibitor period but only one with FVIII and chemotherapy. The total amount of treatment received was in general much lower in the group that did not become tolerant. In individual cases, it appeared very clear that the inhibitor level and anamnestic response was substantially reduced prior to the ITI using the Malmo treatment model. We conclude that treatment of acute bleeds during the inhibitor period may be of importance for ITI and that the different response rates published for different immune tolerance regimens most likely do not reflect the true response rate for the respective regimen. PMID- 11122390 TI - Gynaecological and obstetrical morbidity in women with type I von Willebrand disease: results of a patient survey. AB - Type 1 von Willebrand disease (vWD) is generally regarded clinically as 'mild' and the obstetrical-gynaecological features have not been fully described. We administered a patient questionnaire and provider survey of the medical and quality of life aspects of childbirth and menstruation to 99 type 1 vWD patients and compared the patients presently menstruating (n=81) to a cohort of 150 menstruating females in the general population. The following measurements had a statistically higher proportion in the vWD group: number of tampons/towels used for a typical menstrual cycle (P=0. 002); percentage reporting that clothes are stained by menses (P = 0. 001); past or present history of anaemia (P = 0.001); childbirth-related bleeding (P=0.001); and childbirth-related bleeding necessitating RBC transfusion (P=0.002). Quality of life assessment of the impact of menses in both of the above cohorts was measured by a Likert scale using seven quality of life parameters. Compared to the control group, the vWD patients had a significantly higher score, with P-values of < 0.0001 for each parameter. Hormonal interventions for menorrhagia in the vWD patients were < or = 50% effective. Menorrhagia resulted in red blood cell transfusions in 6% of patients, dilatation and curettage in 17% and hysterectomy in 13%. Despite the common connotation of type 1 vWD as clinically 'mild', childbirth and the monthly challenge to haemostasis presented by menstruation result in a substantial degree of morbidity in females with type 1 vWD. These results support the rationale for ongoing international efforts to increase awareness of vWD as a cause for menorrhagia and to improve the quality of life in females with known vWD. PMID- 11122391 TI - Joint evaluation instruments for children and adults with haemophilia. AB - With the heightened interest in protocols to prevent or treat complications of haemophilia related to recurrent haemarthroses, there is a need for sensitive joint-evaluation tools. The World Federation of Haemophilia (WFH) Physical Joint Examination instrument, which was developed for persons with haemophilia worldwide, is not sensitive enough to detect early structural or functional abnormalities. Therefore, we have expanded the WFH instrument to detect more subtle abnormalities of joint structure and function, and in addition, developed a new scale specifically tailored to the dynamic growth and gait development of children. We compared the original and three new instruments in 43 children with haemophilia. The three new scales all showed better correlation with the WFH pain instrument than did the original WFH physical examination instrument (P < 0.01 for each of the new instruments vs. P > 0.05 for the WFH instrument). In addition, results of the new child physical examination instrument best conformed to a normal distribution (P=0.35) and this instrument had better overall statistical performance. This instrument should be studied further in prospective, longitudinal clinical trials of young children. PMID- 11122392 TI - Late clinical, plain X-ray and magnetic resonance imaging findings in haemophilic joints treated with radiosynoviorthesis. AB - The clinical, plain X-ray and magnetic resonance imaging (MRI) findings were studied in 13 haemophilic joints previously treated with radiosynoviorthesis. (32)P had been injected into the joints at a median of 16 years earlier in an attempt to halt recurrent haemorrhage. Prior to (32)P injection, the majority of joints demonstrated bone damage evident on plain X-ray, secondary to recurrent haemorrhage. At the follow-up evaluation we found plain X-rays were adequate to identify cysts, erosions and cartilage loss in these very damaged joints. MRI was superior to clinical examination and plain X-ray in identifying synovial hyperplasia and effusions. PMID- 11122393 TI - The utility of the Dutch Arthritis Impact Measurement Scales 2 for assessing health status in individuals with haemophilia: a pilot study. AB - The aim of this pilot study was to examine the usefulness of the Dutch version of the Arthritis Impact Measurement Scales 2 (D-AIMS2)in assessing the health status of Dutch individuals with haemophilia. Sixty-eight individuals with mild, moderate, and severe haemophilia attending our clinic for their annual check-up participated. They first completed the Canadian Occupational Performance Measure (COPM). The D-AIMS2 was filled in afterwards at home. With the COPM, individuals rated their specific problematic activities of daily life (ADL), as well as the severity and importance of each problem. The D-AIMS2 is a comprehensive, self administered questionnaire that evaluates functional health status. Fifty-seven individuals completed and returned the D-AIMS2. Reliability analysis demonstrated good internal consistency for the scales (Cronbach's alpha=0.76-1.00), as well as for the components (alpha=0.80-0.88), except for the component 'social interaction' (alpha=0.44). Criterion validity of the D-AIMS2 was assessed by comparison with COPM outcomes; 80% of the problematic ADLs were included in the questionnaire, 20% were missing. Correlations between the D-AIMS2 components 'physical health' and 'symptoms' with predicted scores of those individuals by a highly experienced physiotherapist (r=0.63 and 0.53, respectively) substantiated its concurrent validity. Based on these results we concluded that the D-AIMS2, with minor adjustments, can be an appropriate tool for assessing the health status of Dutch haemophilia patients. PMID- 11122394 TI - Nursing roles in orthopaedic joint correction in haemophiliac patients. AB - The roles of nurses in 16 orthopaedic joint corrections of 14 haemophilia A patients (eight severe, six moderate) are described. The patients' ages ranged from 10 to 37 years with a mean age of 17 years and 4 months. The nursing tasks could be divided into three stages. The first is preoperation, the nurse acting as care provider on a team of experts involved in planning the corrective surgery for the affected joints; and as a counsellor for preparing the patients and family members to cope with the challenging operation. The second is intraoperation, the nurse having the role of care provider, giving factor concentrate either by bolus injection (seven episodes) or continuous infusion (nine episodes). The third is postoperation, as a comprehensive care provider, giving cryoprecipitate and/or factor concentrate and monitoring bleeding doses, and as a trainer, teaching the patients how to dissolve blood components and self venepuncture with aseptic technique. Home treatment for early bleeding was given for 11 episodes, while 6-month prophylaxis was given for five. The accomplishment of these different roles required good communication and nurturing skills, a well adjusted personality and a warm and positive attitude. The successful performance of nursing roles allows the haemophiliac patients to have a near-normal quality of life. PMID- 11122395 TI - Poor response to interferon treatment for chronic hepatitis C in human immunodeficiency virus-infected haemophiliacs. AB - We performed a pilot study to evaluate the factors associated with response to interferon (IFN) therapy for chronic hepatitis C (CHC) with human immunodeficiency virus (HIV) coinfected haemophiliacs. Seven haemophiliacs, coinfected with HIV and hepatitis C virus (HCV), received 9 mega-units (MU) of natural IFN-alpha daily during the first 2 weeks and then three times a week for 22 weeks, all injected subcutaneously. Six patients were receiving zidovudine (AZT) 600 mg day-1 and didanosine (ddI) 200 mg day(-1) during IFN therapy. This treatment was safe and well tolerated. Four patients had no detectable serum HCV RNA at the end of therapy, but long-term, none of the seven patients achieved a sustained response, i.e. undetectable serum HCV-RNA with persistently normal serum alanine aminotransferase (ALT) 6 months after therapy. IFN did not affect CD4-positive cell counts. Most of our patients had high HCV-RNA loads and/or low CD4 counts, both unfavourable markers for IFN therapy. In conclusion, IFN therapy did not eradicate HCV from haemophiliacs coinfected with HIV. PMID- 11122396 TI - Emotional and behavioural problems and family functioning in children with haemophilia: a cross-sectional survey. AB - A comparative study is presented about emotional and behavioural problems in haemophilia and family functioning. This cross-sectional survey looked at boys, aged between 4 and 15 years, with haemophilia and compared them with a group of their healthy school peers. A basic demographic questionnaire was used for both groups along with the Child Behaviour Checklist (CBCL) and the Family Assessment Measure (FAM). Seventeen of 24 families of boys with haemophilia participated (70.8% response). The comparison group consisted of 12 boys, i.e. 70.6% of the haemophilia sample. The groups did not differ in terms of the children's ages and family sizes but significantly fewer of the mothers of the boys with haemophilia worked outside the home. The two groups were compared for scores on the CBCL and FAM. More problems were identified in the haemophilia group on both measures, i.e. there were more emotional, behavioural and family difficulties compared with the healthy group; however, because of the small sample sizes, the differences between the groups did not reach statistical significance. A larger study would be indicated in order to explore these differences further. PMID- 11122397 TI - Massive bleeding from a gastric erosion after transcatheter arterial chemoembolization for hepatocellular carcinoma in a patient with mild haemophilia A. AB - We observed massive bleeding from a gastric erosion following transcatheter arterial chemoembolization (TAE) in a patient with mild haemophilia A. A 78-year old haemophiliac (factor VIII level over 60%) received TAE with farmorubicin and spongel. Haematemesis and melena with loss of consciousness occurred 3 days [corrected] after TAE, and endoscopy revealed superficial erosions with oozing. Toxic effects of the anticancer drug in conjunction with the bleeding disorder may have caused the massive bleeding. We should always consider the possibility of unexpected complications in patients with bleeding disorders; gastrointestinal bleeding can develop during treatment for liver tumours. PMID- 11122398 TI - Induction of immune tolerance and suppression of anaphylaxis in a child with haemophilia B by simple plasmapheresis and antigen exposure. AB - Anaphylaxis to factor IX (FIX) in patients with haemophilia B is a rare and life threatening complication that has been reported to occur in association with the development of inhibitors to FIX. Management of these patients is difficult. This report presents an 18-month-old boy with a frame-shift mutation of the FIX gene and FIX coagulant level of <1% who developed anaphylactoid reactions to low and high purity plasma-derived FIX concentration infusions and an inhibitor measuring 1.0 BU mL(-1). The patient was managed with simple plasmapheresis, a short course of corticosteroids and high-dose antigen exposure, which successfully induced long-lasting immune tolerance to FIX concentrates. PMID- 11122399 TI - Unexpected presentation of type 2N von Willebrand disease in pregnancy. AB - The finding of low factor VIII levels in pregnancy immediately raises concern of haemophilia A carriage, especially with a history of bleeding in the maternal grandfather. However, the diagnosis of type 2N von Willebrand disease (2N vWD) should also be considered as illustrated here. This is also the first reported case of the management of 2N vWD in pregnancy. PMID- 11122400 TI - Disappearance of factor VIII autoantibodies preceding autoimmune haemolytic anaemia. AB - We describe a previously healthy woman who at the age of 44 years developed a factor VIII inhibitor, that over the years increased to a maximum level of 3600 Bethesda units (BU) mL(-1) in 1978. The epitope specificity of the factor VIII inhibitor was investigated and antibodies directed against the A2 and C2 domains of factor VIII were detected. The majority of these antibodies were of subclass IgG4. Over the years, the inhibitor titre gradually decreased and in 1989, the inhibitor could no longer be detected. Shortly after, the patient developed autoimmune haemolytic anaemia. A possible link between the disappearance of factor VIII inhibitors and the development of other autoantibodies may be explained by concomitant development of anti-idiotypic antibodies that neutralize the activity of factor VIII inhibitors. We were unable to detect anti-idiotypic antibodies, which could explain the decline in factor VIII inhibitor titre in this patient. PMID- 11122401 TI - Long-term evaluation of a novel surgical approach to the pseudotumour of the ilium in haemophilia: exeresis and transposition of the omentum in the residual cavity. AB - Pseudotumour of the ilium is a rare but severe complication in haemophiliacs. Excision is often complicated by infections, fistulation and extensive pelvic bone destruction. In 1978, the first author carried out excision of the pseudotumour with transposition of the omentum in the dead cavity to avoid recurrence. This type of surgery has been carried out in three additional patients. The long follow-up of these four patients suggests that this procedure is feasible and curative; local bleeding, infection and fistulation did not recur and the patients remained ambulant with the aid of appropriate devices. PMID- 11122402 TI - Spontaneous intracranial bleeding in two patients with congenital afibrinogenaemia and the role of replacement therapy. AB - Congenital afibrinogenaemia and hypofibrinogenaemia are rare disorders of haemostasis. In this case report the problems posed in the management of two patients with fibrinogen levels less than 0.1g L(-1) and who developed intracranial bleeding are considered. The value of fibrinogen concentrate and the role of prophylaxis is also discussed. PMID- 11122403 TI - Acquired haemophilia in recipients of depot thioxanthenes. AB - We present two cases in which the occurrence of acquired haemophilia is associated with the use of depot preparations of the thioxanthenes zuclopenthixol and flupenthixol. These drugs have not previously been implicated in the aetiology of acquired haemophilia. PMID- 11122404 TI - Carrier analysis of a moderately affected haemophilia B family. AB - Here we report the successful genetic diagnosis of a pregnant caucasian female patient whose family has a history of moderate haemophilia B. While restriction fragment length polymorphism (RFLP) analysis was not informative, nucleotide sequencing of the factor IX genes of the patient's family members determined that her mother and one of her two sisters were carriers of the mutation C31008T, which causes a Thr296Met transition. In contrast, the pregnant female herself and her other sister were found to carry only normal alleles. Plasma factor IX activity and antigen levels supported these findings. PMID- 11122405 TI - Frequencies of five polymorphisms associated with the factor IX gene in the Thai population. PMID- 11122406 TI - Creutzfeldt-Jakob disease and haemophilia: prospect at the new millennium. PMID- 11122407 TI - Haemophilia and the forbidden abdomen. AB - Abdominal surgery became routinely possible over a hundred years ago, after the introduction of general anaesthesia and sterile procedures. Abdominal surgery for haemophiliacs had to wait another 60 or 70 years for adequate control of haemostasis. This paper traces its gradual achievement from the 1920s to the 1970s through a series of reports of appendectomies, gastric and intestinal operations, gall bladder operations and splenectomies in patients with haemophilia of varying degrees of severity. A short-lived flurry of interest in splenectomy as a proposed treatment for haemophilia is also mentioned. PMID- 11122408 TI - Phylogeography of the European rabbit (Oryctolagus cuniculus) in the Iberian Peninsula inferred from RFLP analysis of the cytochrome b gene. AB - We studied mitochondrial DNA variation in the European rabbit through the examination of restriction fragment length polymorphism in 526 individuals from 20 locations spread across the Iberian Peninsula. Digestion with eight enzymes of a 1120-bp fragment comprising most of the cytochrome b gene resolved 38 different haplotypes. These haplotypes were distributed in two highly divergent clades, with different but overlapping geographical distributions, and with comparable levels of within-clade variation. The overall phylogeographical pattern suggests a history of long-term regional isolation of two groups of rabbit populations, compatible with the recognition of two subspecies within the Iberian Peninsula, followed by recent contact and admixture. The underlying cause is sought in the alternation of glacial and interglacial periods in the late Pleistocene. PMID- 11122409 TI - Genetic relationships and spatial genetic structure among clonal stocks of Trypanosoma cruzi in Colombia. AB - Genetic variability in the protozoan causative agent of Chagas' disease, Trypanosoma cruzi, has been analysed in some Latin American countries; Brazil, Bolivia, Chile and Paraguay. Although Colombia is a country displaying enormous biological diversity, few studies have been conducted from the perspective of the population genetics of Trypanosoma cruzi. This study was carried out using 23 Colombian stocks of this protozoan, analysed for 13 isoenzyme loci. The main population genetic results were: (1) Colombia is one of the distribution areas where T. cruzi appears to have the highest genetic variability and heterogeneity in Latin America; (2) the Pgm locus was found in fixed heterozygosis, supporting the presence of diploidy in this organism; (3) the absence of segregating genotypes and the absence of Hardy-Weinberg equilibrium support the view of the existence of a clonal structure as claimed by Tibayrenc and Ayala. Nevertheless, one characteristic of clonal structure, that of over-representation of some identical zymodemes in vast areas of varying environmental conditions, was not found in Colombia. In this country, a strong spatial autocorrelation, with a classic structure of regional patches, was observed. PMID- 11122410 TI - Amount and structure of intra- and interspecific genetic variation in the moss genus Polytrichum. AB - Allozyme electrophoresis was used to determine amount and structure of genetic variation within and between five congeneric haploid moss species: Polytrichum formosum, P. commune, P. uliginosum, P. piliferum and P. juniperinum. For the different species, gene diversity within populations (HS) ranged from very low (0.025) to moderate values (0.127), being, on average, lower than those observed for vascular plants and many other moss species. Polytrichum piliferum and P. juniperinum showed significantly higher levels of HS than the other species examined, which possibly might be explained by sexual reproduction being more prevalent in these two species, that often live in more dynamic habitats, where turnover of individuals is more frequent. Genetic variability was observed to be structured in contrasting ways at different levels. At the intraspecific level genetic differentiation among populations of most Polytrichum species was low, FST or = 40 mm). Only one patient had tumour recurrence, but he survived 7.6 years post transplantation. The 5-year survival of patients with hepatocellular carcinoma with lymphoid stroma was better than that of the patients with other types of hepatocellular carcinomas (P = 0.04). CONCLUSIONS: Hepatocellular carcinoma with lymphoid stroma should be considered as a distinct clinicopathological and prognostic entity. PMID- 11122435 TI - Galectin-3 and carcinoembryonic antigen expression in medullary thyroid carcinoma: possible relation to tumour progression. AB - AIMS: Galectin-3 is a beta-galactoside binding protein involved in multiple biological processes through interactions with complementary glycoconjugates. We analysed the expression and coexpression of galectin-3 and carcinoembryonic antigen (CEA), one of the putative galectin-3 ligands, in medullary thyroid carcinoma (MTC). METHODS AND RESULTS: An immunohistochemical study using monoclonal antibodies was performed on paraffin sections of 20 cases of sporadic MTC comprising 10 cases without and 10 cases with lymph node metastases at the time of surgery. CEA expression was found in all tumours, distributed predominantly in the cytoplasm and occasionally at the cell surface. In the majority of cases (18/20) moderate to strong intensity of staining was found in most of the cells. Positive cytoplasmic staining for galectin-3 was found in 16/20 cases, but varied in intensity and distribution from weak/focal (7/16) to moderate (7/16) or strong (2/16). More intense staining for galectin-3 was mainly associated with MTC cases involving lymph node metastases. Eight out of these 10 cases showed moderate to strong galectin-3 expression concomitant with CEA expression throughout the tumour tissue. CONCLUSIONS: These findings suggest that galectin-3 might play a role in the pathobiology of MTC. Simultaneous expression of galectin-3 and CEA in the same tumour cells at an advanced stage of MTC indicates the possibility of their autocrine cooperation during tumour progression. PMID- 11122436 TI - High sensitivity and specificity of immunohistochemistry for the detection of hormone receptors in breast carcinoma: comparison with biochemical determination in a prospective study of 793 cases. AB - AIMS: The hormone receptor (HR) status of breast cancer is an important prognostic factor and predictive parameter of the response to hormone therapy. Enzyme immunoassay (EIA) is currently the standard for determination of HR, but immunohistochemistry (IHC) represents a potentially useful alternative. We used IHC to determine HR status in a large prospective study and compared the results to those obtained by EIA. This study was designed to determine which technique should be used in daily practice in our institution which manages a large number of patients. METHODS AND RESULTS: Oestrogen (ER) and progesterone (PgR) receptor status was evaluated in a prospective series of 793 infiltrating breast cancers by IHC in paraffin-embedded tissue sections, using antibodies 6F11 and 1A6, with a rigorous quality control of the methodology. ER were found to be significantly expressed in 81% of cases after IHC analysis and in 78% of cases by EIA. For PgR, the respective rates of positivity were 65% and 69%. The tumour HR level detected by either technique was significantly correlated with the value of tumour size, histological grade and S-phase fraction. A significant link was observed between the percentage of labelled cells after IHC analysis and the amount of protein detected by EIA. Critical analysis of discordance found that, in the group of invasive lobular carcinomas, the rate of HR positivity was higher with IHC (84%) than with EIA (45%) and that, in the overall population, IHC was more specific than EIA, since cases with nonrelevant positivity related to intraductal normal or neoplastic cells expressing HR could be discarded. The cost of IHC analysis was found to be about one-third of that of EIA. CONCLUSIONS: IHC is more sensitive, specific and economical than EIA. It should constitute the new standard technique provided that good quality assurance procedures are respected. PMID- 11122437 TI - Sequential changes in localization of repair-related proteins (heat shock protein 70, ubiquitin and vascular endothelial growth factor) in the different stages of myocardial infarction. AB - AIMS: The myocardium expresses vascular endothelial growth factor (VEGF), heat shock protein 70 (HSP70), and ubiquitin immediately after the onset of cardiac ischaemia. This study demonstrated the sequential changes in localization of these proteins, in addition to fibronectin and troponin T (TnT), in human hearts with myocardial infarction (MI). METHODS AND RESULTS: Myocardial tissues from 40 autopsied MI cases were immunostained with the five antibodies against VEGF, HSP70, ubiquitin, fibronectin and TnT. Fibronectin was recognized only in the cardiomyocytes with infarction. Although TnT, HSP70, ubiquitin and VEGF were detected in the affected myocardium in the early stages, their expression in cardiomyocytes around infarcted foci were more intense. The cardiomyocytes with coagulative myocytolysis were positive for fibronectin, but negative or weakly positive for TnT, HSP70, ubiquitin and VEGF. In contrast, the cardiomyocytes with colliquative myocytolysis were strongly positive for TnT, HSP70, ubiquitin and VEGF, but negative for fibronectin. CONCLUSIONS: Immunostaining using antibodies to fibronectin, TnT, HSP70, ubiquitin and VEGF is useful for the discrimination between infarcted myocytes and ischaemia-damaged myocytes in the human heart with MI at autopsy. PMID- 11122438 TI - Caspase-3/CPP32 immunoreactivity and its correlation with frequency of apoptotic bodies in human prostatic carcinomas and benign nodular hyperplasias. AB - AIMS: Apoptosis is mediated by apoptosis-specific genes, certain oncogenes and tumour suppressor genes. Caspase-3, a group of cystein proteases, is involved in the induction of apoptosis and has been considered to correlate with apoptosis. The aim of this study was to determine whether caspase-3 is expressed in prostatic carcinoma and benign prostatic hyperplasia, and correlated with the apoptosis. METHODS AND RESULTS: We studied the apoptotic index and caspase-3 immunoreactivity in 40 cases of benign nodular hyperplasia (BPH) and 40 cases of prostate carcinoma (PCA) by in-situ labelling and immunohistochemistry. The mean number of apoptotic bodies in cases with BPH was not significantly different from cases with PCA I (Gleason score 2-4), but samples from patients with PCA II (Gleason score 5-7) and PCA III (Gleason score 8-10) showed a significantly higher apoptotic number than cases with BPH. Positive staining for caspase-3 was seen in 42.5% (17/40) of the BPH, and 27.5% (11/40) of the PCA: PCA I was 41.7% (5/12), PCA II 14.3% (2/14) and PCA III was 28.6% (4/14). CONCLUSIONS: Based on our results, the number of apoptotic bodies was not correlated with the caspase-3 expression and there was no relationship between caspase-3 expression and Gleason score. However, the number of apoptotic bodies was significantly higher in cases with intermediate (Gleason score 5-7) and high-grade (Gleason score 8-10) PCAs than cases with BPH and low-grade PCAs (Gleason score 2-4). PMID- 11122439 TI - Mucin core proteins as differentiation markers in the gastrointestinal tract. PMID- 11122440 TI - Adenocarcinoma of colon associated with gut-associated lymphoid tissue. PMID- 11122441 TI - Adenocarcinoma of colon differentiating as dome epithelium of gut-associated lymphoid tissue. PMID- 11122442 TI - Multiple residual cysts containing deposits of AL-amyloid. PMID- 11122443 TI - Multinucleated stromal cells. PMID- 11122444 TI - Angiomyolipoma and high-grade sarcoma of the liver. PMID- 11122446 TI - Review article: role of the surrogate light chain and the pre-B-cell receptor in mouse B-cell development. PMID- 11122447 TI - Role of CD4(+) T helper 2-type cells in cutaneous inflammatory responses induced by fluorescein isothiocyanate. AB - Owing to its skin-sensitizing and fluorochromatic properties, fluorescein isothiocyanate (FITC) is employed frequently as an experimental hapten in mechanistic studies of contact allergy, particularly in the context of the role of migration and activation of Langerhans' cells. In this study we demonstrated that topical exposure of mice to FITC results in the selective development of activated lymph node cells (LNC) expressing a preferential type 2 cytokine secretion profile, with high levels of interleukin (IL)-4 and IL-10, but low levels of interferon-gamma (IFN-gamma). Negative selection (complement depletion) identified CD4(+) T helper (Th)2-type cells as the primary source in activated LNC of the type 2 cytokines IL-4 and IL-10, whereas the low levels of IFN-gamma produced were derived exclusively from CD8(+) T cytotoxic (Tc) 1-type cells. A biphasic pattern of cutaneous inflammatory reactions was elicited by exposure to FITC, the early phase of which could be transferred passively with serum (presumably immunoglobulin E [IgE] antibody), whereas adoptive transfer experiments demonstrated that Th2-type CD4(+) cells were responsible for the delayed-type component of the dermal hypersensitivity reaction. In contrast with contact allergic reactions induced by other sensitizing haptens, which are considered to be largely Th1/Tc1-mediated immune processes regulated by Th2-type cells, these results suggest therefore that the skin lesions provoked in mice by FITC are primarily a result of the activation of Th2-type cells. PMID- 11122448 TI - Differential expression of ovine CD1. AB - was examined using a combination of reverse transcription-polymerase chain reaction (RT-PCR) with sequence-specific primers and Northern and in situ hybridization techniques. The aim of the study was to establish the patterns of CD1 expression at the molecular level and address questions posed by previous studies in other species regarding expression patterns of CD1. A 'pan-CD1' probe based on the exon 4 (alpha 3) region was used in addition to isotype-specific probes for SCD1B (the exon 3 region of clone SCD1B42) and SCD1D (the exon 3 region of clone SCD1D). Widespread expression of CD1 (including thymus, peripheral blood lymphocytes, lung and intestine) was identified using both the exon 4 and SCD1D probe. SCD1B expression was more restricted, being identified in equivalent levels only in the thymus and in scattered populations of dendritic cells. These results highlight the difference in expression patterns between group 1 and group 2 CD1 family members and establish SCD1D as the CD1 family member with the widest pattern of expression, consistent with a differential role for the different CD1 family members. PMID- 11122449 TI - Natural antibody-induced intracellular signalling and growth control in C3H 10T1/2 fibroblast variants. AB - Cell-surface binding by natural antibody (NAb) places it well for controlling cell function directly through signalling. Flow cytometry revealed an instability of syngeneic NAb binding to C3H 10T1/2 fibroblast variants at 37 degrees, which could be partially reduced by H7, an inhibitor of the pivotal signalling serine/threonine kinase, protein kinase C (PKC). Cells coated with purified NAb at 4 degrees followed by a rise in temperature to 37 degrees showed an increase in membrane expression of introduced rat PKC-beta 1 and endogenous PKC-alpha, in the PKC-beta 1-overexpressing PKC-4 and v-H-ras-producing I3T2.1, respectively. Tyrosine phosphorylation of membrane-associated 60 000 MW protein including the tyrosine kinase src was markedly reduced. In addition, both the precoated NAb and numerous membrane molecules ranging from 20,000 to 220,000 MW were released into the supernatant, including the receptor-like protein tyrosine phosphatase alpha (RPTP-alpha). Furthermore, purified NAb reduced the growth of I3T2.1 cells in culture assessed as a decrease in total cell numbers and an increase in the proportion of cells in the G0/G1 phase of the cell cycle. Together, these data argue that the interaction of NAb with cell surface structures initiated a series of intracellular signalling events leading to the release of membrane molecules and over time the suppression of cell proliferation. This process could provide a biological mechanism for direct NAb control of activated cells in both physiological and pathological conditions. PMID- 11122450 TI - Distribution of, and immune response to, chicken anti-alpha Gal immunoglobulin Y antibodies in wild-type and alpha Gal knockout mice. AB - Chicken antibodies (immunoglobulin Y; IgY) to the alpha Gal epitope (galactose alpha-1,3-galactose) bind to alpha Gal antigens of mouse and porcine tissues and endothelial cells in vitro and block human anti-alpha Gal antibody binding, complement activation and antibody-dependent cell-mediated lysis mechanisms. The activities and toxicity of anti-alpha Gal IgY have not been tested in vivo. In this study, we tested the effects of multiple injections of affinity-purified anti-alpha Gal IgY (AP-IgY) in both wild-type (WT) and alpha-1,3 galactosyltransferase knockout (Gal KO) mice. WT and Gal KO mice were injected once, twice, three, or four times intravenously (i.v.) with AP-IgY and killed at 1 hr or 24 hr. Mice displayed no toxicity to four injections of AP-IgY. Heart, lung, liver, kidney, spleen and pancreatic tissue were evaluated using immunohistochemical techniques for the presence of the alpha Gal epitope using the GSI-B4 lectin, and for bound IgY, as well as mouse IgM and IgG. The binding of AP-IgY antibodies to the endothelium of WT mouse tissues was essentially identical to the pattern of binding of the GSI-B4 lectin after injection of WT mice and death at 1 hr. WT mice killed 24 hr after i.v. injection of AP-IgY showed little remaining bound IgY in their endothelia, indicating that IgY is cleared over that time period. We also evaluated the blood drawn at the time of death for the presence of anti-alpha Gal IgY, anti-IgY IgM and anti-IgY IgG by enzyme-linked immunosorbent assay. Anti-alpha Gal IgY was almost undetectable in WT mouse sera at all injection and killing times. In contrast, Gal KO mouse sera showed increasing anti-alpha Gal IgY levels until 24 hr after the fourth injection, when anti-alpha Gal IgY levels were almost undetectable. Anti-IgY IgM and IgG levels in WT and Gal KO mouse sera showed a typical increase in anti-IgY IgM 24 hr after the second injection (3 days after the first injection) and an increase in anti-IgY IgG 24 hr after the third injection (5 days after the first injection). These results show that IgY binds to alpha Gal epitopes in the WT mice and is cleared sometime over a 24-hr time period and that IgY is an expected immunogen in mice eliciting a rather typical anti-IgY IgM and IgG response. PMID- 11122451 TI - Natural killer cell-dependent immunoglobulin G2a anti-bovine serum albumin (BSA) response elicited by high molecular weight dextran-BSA conjugates associated with dextran-mediated macrophage-natural killer cell interaction. AB - The roles of the interferon-gamma (IFN-gamma) and interleukin-12 (IL-12) produced during natural killer (NK) cell interaction with macrophages (M phi) were investigated as the basis for the induction of immunoglobulin G2a (IgG2a) anti bovine serum albumin (BSA) responses by high molecular weight dextran conjugated to BSA (HMW-DEX-BSA). BALB/c mice immunized with HMW-DEX-BSA produced significantly higher levels of both IgG1 and IgG2a anti-BSA than did mice immunized with BSA alone. Both IgG1 and IgG2a anti-BSA levels were higher in mice immunized with BSA conjugated to dextran of molecular weight (MW) 5 000 000-40 000 000 compared with dextran of MW 10,000-60,000. The enhancement of anti-BSA IgG2a levels but not of anti-BSA IgG1 levels was inhibited when free BSA was added to the HMW-DEX-BSA conjugate. NK cell depletion during HMW-DEX-BSA immunization of mice resulted in significantly lower anti-BSA IgG2a levels without affecting anti-BSA IgG1 levels. Naive splenocytes or M phi + NK cell co cultures incubated with HMW-DEX or HMW-DEX-BSA produced higher IFN-gamma levels than splenocytes or co-cultures incubated with BSA alone. HMW-DEX stimulated both IFN-gamma and IL-12 production by M phi + NK cell co-cultures in a dose-dependent manner. DEX-induced IFN-gamma production by NK cells was dependent upon the presence of IL-12, and IL-12 production by M phi was dependent upon the presence of IFN-gamma in these co-cultures. Both M phi and NK cells bound DEX to their surfaces. These data demonstrate that BSA linked to HMW-DEX enhanced both T helper-1- and T-helper-2-associated antibody responses to BSA. The results also indicate an IL-12-dependent positive feedback interaction between NK cells and M phi that supports a NK cell/IFN-gamma-dependent mechanism for enhancement of anti BSA IgG2a antibody responses in mice immunized with HMW-DEX-BSA protein conjugates. PMID- 11122452 TI - Natural killer 1.1(+) alpha beta T cells in the periimplantation uterus. AB - When the developing embryo implants into the uterine wall, resident maternal immune cells may encounter antigens present on the fetal tissues. The nature and constituents of the ensuing maternal immune response, and its regulation, are of considerable interest in understanding normal and abnormal pregnancy. Here, we report the presence of natural killer (NK)1.1(+) alpha beta T cells in the murine periimplantation uterus. These cells account for a large portion of both the T cell and natural killer cell populations in early pregnancy, while their numbers in the non-pregnant uterus and later in pregnancy are greatly reduced. Phenotypically, these NK1.1+ alpha beta T cells belong to a previously described subset of cells that bear a V alpha 14-J alpha 281-encoded T-cell receptor. Unlike other organs, where both CD4(+) and CD4(-)/CD8(-) NK1.1(+) alpha beta T cells are found, the placental/decidual population appears to be entirely CD4( )/CD8(-). The V beta repertoire of the placental/decidual population is also altered from that of other organs, with a majority of cells expressing V beta 3. Together, these features suggest the possibility of local development. NK1.1(+) alpha beta T cells are known to recognize the class I-like CD1 molecule. Consistent with this association, we demonstrate CD1 expression by tissues within the pregnant uterus. Our findings define an additional organ-specific immune environment where NK1.1(+) alpha beta T cells may play a role, and continue to demonstrate the specialized nature of the maternal intrauterine immune system during pregnancy. PMID- 11122453 TI - Professional and non-professional antigen-presenting cells in the porcine small intestine. AB - We have previously presented evidence of a highly organized and compartmentalized structure of the small intestinal lamina propria of the pig. In this work, we have identified at least two major populations of cells in this site expressing high levels of major histocompatibility complex (MHC) class II antigens. One is CD45 positive and is a potent initiator of a primary immune response, this is a function usually associated with dendritic cells. These cells have characteristic dendritic morphology, but show evidence of phagocytosis as well as other phenotypic markers of immature dendritic cells. Some cells show evidence of ongoing immune maturation. We have also isolated CD45 negative endothelial cells bearing significant amounts of MHC class II, which do not trigger a mixed lymphocyte reaction. These findings have implications for the functional role of healthy gut lamina propria and clearly implicate this site as capable of differential antigen presentation by a heterogeneous population of antigen presenting cells. PMID- 11122454 TI - A critical role for alveolar macrophages in elicitation of pulmonary immune fibrosis. AB - Hapten immune pulmonary interstitial fibrosis (HIPIF) is induced by a recall cell mediated immune response against the hapten 2,4, 6-trinitrobenzene sulphonic acid (TNBS) in the lung. Studies here dissect the role of the cellular components of the bronchoalveolar lavage (BAL) cells (alveolar macrophages [AMs] versus monocytes and immature dendritic cells) in the fibrogenic inflammatory response. BAL cells from HIPIF mice were generally more activated and produced a greater amount of tumour necrosis factor-alpha (TNF-alpha) than controls. Liposome encapsulated dichloromethylene diphosphonate (Cl(2)MDP) that was inoculated intranasally (i.n.) into mice selectively depleted AMs. Following AM depletion, the number of TNF-alpha-containing cells was reduced, and both the number of immune inflammatory cells recruited into the alveolar space and the subsequent collagen deposition (hydroxyproline) were decreased in the sensitized and intratracheally (i.t.) challenged mice. In conclusion, AMs are required, in part, for the development of pulmonary fibrosis in HIPIF because AM-derived factors such as TNF-alpha are needed for initiation of chemokine and cytokine pathways and accumulation of immune inflammatory cells. PMID- 11122455 TI - Antigen processing of vesicular stomatitis virus in situ. Interdigitating dendritic cells present viral antigens independent of marginal dendritic cells but fail to prime CD4(+) and CD8(+) T cells. AB - Acute macrophage (M phi) depletion, using a liposome-mediated 'suicide technique', markedly suppressed priming of splenic CD4(+) and CD8(+) T-cell responses to vesicular stomatitis virus (VSV). However, phagocytic marginal dendritic cells (MDC), but not interdigitating dendritic cells (IDC), are now known to be also depleted by this technique. To clarify the role splenic dendritic cell (DC) subsets and M phi play in priming for a virus-specific T-cell mediated immune response, DC and M phi were purified from VSV-infected mice and assayed for the presence of epitopes recognized by VSV helper T (Th) cells and cytotoxic T lymphocytes (CTL). Antigen pulse experiments performed in situ demonstrated that VSV Th cell and CTL epitopes became transiently associated only with DC, but not M phi or B cells, indicating that DC represent the critical antigen-presenting cell (APC) population in vivo for this virus. The failure of MDC/M phi-deficient mice to become primed was not due to the complete elimination of antigen-presenting DC because VSV peptide/class I and II complexes were detected on IDC following lipsome-mediated elimination of phagocytic cells. However, the VSV-induced chemokine response was dramatically suppressed in these mice. Thus, despite the expression of VSV peptide/class I and II complexes, IDC are not sufficient to prime VSV Th cells in the absence of MDC and/or splenic M phi. PMID- 11122456 TI - Polymorphonuclear neutrophils as accessory cells for T-cell activation: major histocompatibility complex class II restricted antigen-dependent induction of T cell proliferation. AB - Polymophonuclear cells (PMN) of healthy donors do not express major histocompatibility complex (MHC) class II antigens or the T-cell costimulatory molecules CD80 or CD86. Expression of these receptors, however, is seen in patients with chronic inflammatory diseases. We now report that, by culturing PMN of healthy donors with autologous serum, interferon-gamma (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF), de novo synthesis of MHC class II, CD80 and CD86 could be induced. MHC class II-positive PMN acquired the capacity to present staphylococcus enterotoxin to peripheral T cells, apparent as induction of interleukin-2 (IL-2) synthesis and proliferation of the T cells. Moreover, the PMN also processed tetanus toxoid (TT) and induced proliferation of TT-specific T cells in a MHC class II-restricted manner. Taken together, these data indicate that PMN can be activated to function as accessory cells for T-cell activation. PMID- 11122457 TI - Expression of kininogens in the connective tissue-type mast cells of the rat. AB - The connective tissue-type mast cells present in the submandibular gland (SMG) and peritoneal cavity of rats were found to express kininogens (KGs), the expression of which was demonstrated by Western blotting, reverse transcription polymerase chain reaction (RT-PCR), RT-PCR Southern blotting, and light- and electron-microscopic immunocytochemistry. In the SMG, the analysis of cDNA amplified by RT-PCR revealed that the molecular species of mRNAs expressed were high-molecular-weight (HMW)-K KG and T-I KG. Light microscopic immunocytochemistry exclusively localized the KG protein(s) in the mast cells present in the SMG. The signals in the mast cells were very strong, but no positive reaction was observed in the granular convoluted tubular cells, acinar cells or striated duct cells. As determined by using electron microscopy, extremely strong labelling with immunogold was observed in the secretory granules of the mast cells, but no labelling in their nucleus or cytoplasm. Analysis by Western blotting and RT-PCR Southern blotting indicated that both protein and mRNA of KGs were present in the mast cells separated from the peritoneal cavity, indicating de novo synthesis of KG in these cells. Preliminary experiments implied that the connective tissue-type mast cells in other rat tissues also expressed KG. PMID- 11122458 TI - Tumour rejection by gene transfer of 4-1BB ligand into a CD80(+) murine squamous cell carcinoma and the requirements of co-stimulatory molecules on tumour and host cells. AB - NRS1 is a murine squamous cell carcinoma that constitutively expresses the co stimulatory molecule CD80 at a high level yet grows as a tumour in syngeneic C3H mice. We examined the effect of gene transfer of the 4-1BB ligand (4-1BBL) into NRS1 cells. Introduction of the 4-1BBL gene efficiently elicited anti-tumour immune responses in syngeneic mice which acquired specific immunity against wild type tumour. T-cell depletion studies showed that CD8(+), but not CD4(+) T cells were essential for tumour eradication. Our results suggest that the transduced 4 1BBL is more effective than the spontaneously expressed CD80 for generation of primary anti-tumour CD8(+) T-cell responses. In addition to CD80 and CD86, the host-derived 4-1BBL is also involved in the secondary anti-tumour responses. This study indicates the complicated contribution of 4-1BBL, CD80 and CD86 on tumour and host cells in anti-tumour immune responses and a possible therapeutic application of 4-1BBL for human tumour vaccination and gene therapy. PMID- 11122459 TI - Xenogeneic islet re-transplantation in mice triggers an accelerated, species specific rejection. AB - Xenogeneic islets could provide an unlimited source of tissue for the treatment of diabetes, and could in theory be transplanted repeatedly in a recipient. However, little is known on the consequences of islet re-transplantation in a recipient who has rejected a first graft. In this study, we investigated the functional consequence of xeno islet re-transplantation in mice sensitized with islets from different species. Sprague-Dawley (SD)-rat islets transplanted in sensitized C57/Bl6 mice that rejected either SD- or Lewis-rat islets underwent accelerated rejection. However, accelerated rejection was not found in mice sensitized with pig or human islets, suggesting that accelerated rejection was species specific. Immunohistochemistry showed increased binding of antibodies and accelerated leucocyte infiltration on re-grafted islets in sensitized mice. In situ apoptosis detection indicated that islet cell apoptosis was correlated with the time of leucocyte infiltration, but not with the time of antibody binding. In vitro experiments with cultured islet cells showed that although antibody binding was increased after incubation with sensitized mouse serum, islet cell cytotoxicity was not increased, suggesting that humoral immunity did not play a direct role in islet destruction. These results indicate that there is a cell mediated, species-specific accelerated rejection after re-transplantation of xenogeneic islets. PMID- 11122460 TI - A mimotope peptide-based vaccine against Schistosoma mansoni: synthesis and characterization. AB - A panel of four mimotopes of the epitope recognized by the highly protective monoclonal antibody against Schistosoma mansoni (152-66-9B) was obtained by screening a solid-phase 8mer random peptide library. Three of the four mimotopes (p28, p29 and p30) were efficiently recognized in an in vitro radioimmunoassay by the monoclonal antibody and by sera from infected mice and one (p30) induced in vitro proliferation of primed lymphocytes. When the mimotopes were conjugated to bovine serum albumin (BSA) and the conjugates used to immunize C57BL/6J mice, only the p30-BSA-induced antibodies which were effective at complement-mediated killing of schistosomula. The level of complement-mediated killing obtained with the anti-p30 antibodies was comparable to that seen with serum from mice immunized with the protective 9B-antigen. Furthermore, following challenge infection of mimotope-BSA-immunized mice, a greater than 40% reduction in worm burden was observed in p30-BSA-immunized mice, a level comparable to that seen following immunization with the intact 9B-antigen. These results show that a simple synthetic peptide immunogen comprising an eight-amino acid mimotope of a conformational epitope on the 9B-antigen can induce protective immune responses against S. mansoni that are comparable to those obtained following immunization with the far more complex intact antigen. This mimotope may well represent a potential component of a synthetic peptide vaccine against S. mansoni. The inclusion of other B-cell- and T-cell-stimulating synthetic epitopes in such a vaccine, together with a more appropriate carrier, adjuvant and delivery systems may well result in a level of protection even greater than that seen with the single mimotope. PMID- 11122461 TI - Virulence of Sporothrix schenckii conidia and yeast cells, and their susceptibility to nitric oxide. AB - The involvement of nitric oxide (NO) in macrophage (M phi) fungicidal activity against Sporothrix schenckii, and the relationship between NO susceptibility and the differential virulence of conidia and yeast cells, were investigated. Confirming a previously reported correlation between the length of time in culture and virulence of S. schenckii, conidia isolated from 12-day mycelial cultures (Ss-12) were less virulent to mice than conidia from 7-day cultures (Ss 7) or yeast cells. Indicative of NO production, infected animals showed a significant increase in serum levels of nitrite that was lower in mice infected with Ss-12 than in mice infected with Ss-7 or yeast. Stimulation of murine M phi with interferon-gamma (IFN-gamma) induced NO production and inhibition of fungal growth. The cytotoxic activity of M phi against Ss-12 was significantly greater than against Ss-7 or yeast cells, the highly virulent fungal forms. The addition of NO synthase inhibitors abrogated M phi cytotoxic activity against all fungal forms. The phagocytic activity of M phi against Ss-7 was significantly lower than against Ss-12 or yeast cells. Although the ingestion of fungal cells triggered the oxidative burst in M phi, the fungicidal activity was not altered in the presence of superoxide dismutase (SOD) and catalase. In addition, Ss-12 and yeast cells were more susceptible than Ss-7 to the direct fungicidal activity of the NO donors S-nitroso-N-acetyl-DL-penicillamine (SNAP), S-nitrosoglutathione (GSNO) and 3-morpholinosydnonimine (SIN-1). The results of this study indicate that NO is a key cytotoxic mediator involved in the murine M phi defence against S. schenckii, and that the virulence of Ss-7, Ss-12 and yeast cells may be related to a differential susceptibility to NO. PMID- 11122462 TI - Two independent pathways of maternal cell transmission to offspring: through placenta during pregnancy and by breast-feeding after birth. AB - Cell transmission from mother to offspring was demonstrated using mice with green fluorescent protein (GFP) transgenic markers. GFP transgene heterozygous (+/-) females were mated with GFP (-/-) males, and GFP(+) cells in the GFP (-/-) fetuses generated between them were analysed to assess maternal blood cell transmission to conceptuses in utero. The GFP+ maternal cells were observed throughout the body of the fetuses, as shown by fluorescence stereomicroscopy. Cell entrance into the fetal immune system was shown by histochemical and flow cytometric analyses of fetal organs such as thymus, spleen and liver. The GFP(+) maternal cells persisted in the offspring until postpartum. Next, GFP (-/-) neonates fed by GFP(+) foster mothers were examined to study the transfer of maternal milk leucocytes to offspring through breast-feeding. GFP(+) leucocytes that had infiltrated through the wall of the digestive tract were mainly localized in the livers of neonates. Their accumulation in the livers reached a maximum on days 5 or 6, and these cells became undetectable, as assessed by either histochemistry or flow cytometry, after day 9 of starting foster nursing. Collectively, the present results demonstrate two independent pathways of maternal cell transmission to offspring: transplacental passage during pregnancy and breast-feeding after birth. PMID- 11122463 TI - Molecular studies in insect olfaction. PMID- 11122464 TI - Immune activation upregulates lysozyme gene expression in Aedes aegypti mosquito cell culture. AB - After stimulation with heat-killed bacteria, cultured cells from the mosquito Aedes aegypti (Aag-2 cells) secreted an induced protein with a mass of approximately 16 kDa that cross-reacted with antibody to chicken egg lysozyme. To investigate whether lysozyme messenger RNA is induced in bacteria-treated cells, we used polymerase chain reaction-based approaches to obtain the complete lysozyme cDNA from Aag-2 cells. The deduced protein contained 148 amino acids, including a 23 amino acid signal sequence. The calculated mass of the precursor protein is 16 965 Da, which is processed to yield a mature lysozyme of 14 471 Da with a calculated pI of 10.1. The lysozyme from Ae. aegypti shared 50% amino acid identity with lysozymes from Anopheles gambiae and Anopheles darlingi, which in turn shared 70% identity between each other. Northern analysis with the lysozyme cDNA probe showed induction of a 1.3 kb messenger RNA during the first 3 h after treatment of Aag-2 cells with heat-killed bacteria, followed by maximal expression 12-36 h after treatment. Southern analysis suggested that the gene likely occurs as a single copy in the genome of Aag-2 cells. PMID- 11122465 TI - A locust type 1 ADP-ribosylation factor (lARF1)* is 100% identical in amino acid sequence to Drosophila ARF1 despite obvious DNA sequence divergence. AB - The cDNA of a type 1 ADP-ribosylation factor (ARF) from the desert locust, Locusta migratoria was cloned, sequenced and compared to ARF1 genes of other species. The locust ARF1 protein is 100% identical with the ARF1 protein of the fruit fly Drosophila melanogaster even though the DNA sequences are only 79% identical. The significance of this finding in relation to the considerable evolutionary distance between hemimetabolous and holometabolous insects is discussed. PMID- 11122466 TI - Secondary structure and conserved motifs of the frequently sequenced domains IV and V of the insect mitochondrial large subunit rRNA gene. AB - We have analysed over 400 partial insect mitochondrial large subunit (mit LSU) sequences in order to identify conserved motifs and secondary structures for domains IV and V of this gene. Most of the secondary structure elements described by R. R. Gutell et al. (unpublished) for the LSU were identified. However, we present structures for helices 84 and 91 that are not recognized in previous universal models. The portion of the 16S gene containing domains IV and V is frequently sequenced in insect molecular systematic studies so we have many more sequences than previous studies which focused on the complete mitochondrial LSU molecule. In addition, we have the advantage of investigating several sets of closely related taxa. Aligned sequences from thirteen insect orders and nine secondary structure diagrams are presented. These conserved sequence motifs and their associated secondary structure elements can now be used to facilitate the alignment of other insect mit LSU sequences. PMID- 11122467 TI - The tetraspanin superfamily in insects. AB - We describe four members of the tetraspanin/TM4SF superfamily of proteins that were identified in expressed sequence tag projects on the antennae of Manduca sexta moths and Apis mellifera honey bees. The three moth genes are expressed in the sensillar epithelium of male antennae, and some are expressed in female antennae, haemocytes, wing scale cell primordia and/or embryonic tissues. These proteins are probably involved in diverse cellular processes, much like their vertebrate homologues. A phylogenetic analysis of all known tetraspanins, including thirty-seven members of the superfamily revealed by the Drosophila melanogaster genome and twenty in the nematode Caenorhabditis elegans genome, reveals some phylum-specific gene amplification, in particular a contiguous array of eighteen genes in the D. melanogaster genome. PMID- 11122468 TI - Sequence, secondary structure and phylogenetic analyses of the ribosomal internal transcribed spacer 2 (ITS2) in the Timarcha leaf beetles (Coleoptera: Chrysomelidae). AB - Internal transcribed spacer 2 (ITS2) sequences of the nuclear rDNA in forty-seven specimens (thirty-four species) of the leaf beetle genus Timarcha have been studied. Timarcha ITS2 (523 bp on average) share some sequence features with other Chrysomeloidea relatives (Chrysolina, Diabrotica and Bruchus) but have no clear similarity with any other arthropod ITS2 sequences. Interspecific divergences are in the range 0.002-0.166, and 0.124-0.206 in the comparisons between subgenera. No evidence of intragenomic divergent ITS2 sequences has been found. Secondary structures are concordant with the four-domain model proposed for vertebrates and yeast, but differs from those proposed for dipterans. Phylogenetic analysis of the ITS2 data confirms the results of a previous study based on mitochondrial sequences, as the basality of the Metallotimarcha subgenus and the absence of phylogenetic support for the Timarchostoma subgenus. PMID- 11122469 TI - The piggyBac transposon mediates germ-line transformation in the Oriental fruit fly and closely related elements exist in its genome. AB - Germ-line transformation of a white eye strain of the Oriental fruit fly, Bactrocera dorsalis, was achieved with the piggyBac vector, derived from a transposon originally isolated from the cabbage looper moth, Trichoplusia ni. The vector was marked with the medfly white+ gene cDNA, and three transgenic lines were identified at a frequency of approximately 2% per fertile G0. Vector integrations were verified by Southern DNA hybridization, which also revealed the presence of endogenous genomic elements closely related to piggyBac. Approximately 10-20 elements per genome were evident in several B. dorsalis strains, and sequence analysis of 1.5 kb gene amplification products from two wild strains and the white eye host strain indicated 95% nucleotide and 92% amino acid sequence identity among resident elements and the T. ni element. PiggyBac was not evident by hybridization in other tephritid species, or insects previously transformed with the transposon. This is the first discovery of piggyBac beyond T. ni, and its existence in a distantly related species has important implications for the practical use of the vector and insects transformed with it. PMID- 11122470 TI - Differential regulation of ribosomal protein gene expression in Aedes aegypti mosquitoes before and after the blood meal. AB - In fat body of the mosquito, Aedes aegypti, a cycle of ribosome accumulation and degradation accompanies synthesis of the yolk protein precursor, vitellogenin. Here we compare the transcription and translation of ribosomal proteins rpS6, rpL8 and rpL34, relative to rRNA and vitellogenin genes in Aedes aegypti fat body after eclosion, and in response to a blood meal. Analysis using Northern blots and reverse-transcription polymerase chain reactions (RT-PCR) showed that the rpS6, rpL8 and rpL34 genes are coordinately regulated with respect to one another, and that ribosomal protein gene expression in this system was predominantly regulated by transcription during the 3-4 days between adult eclosion and blood feeding. After a blood meal, ribosomal protein mRNA levels remained similar to those in unfed females during the first 18 h, then declined to minimum levels by 48 h after the blood meal. These data indicate that transcription of ribosomal protein genes is low in vitellogenic mosquitoes, relative to previtellogenic females. Experiments with the dissected fat body, however, showed that levels of acetic acid-soluble proteins increased by approximately threefold between 12 and 24 h after the blood meal. Taken together, these observations suggest that the active translation of ribosomal proteins from stable mRNA accompanies ribosome biosynthesis after the blood meal. Thus, in the fat body of adult female mosquitoes, the expression of ribosomal protein genes is regulated at the level of transcription before the blood meal, while translational control is the predominant regulatory mechanism after the blood meal. PMID- 11122471 TI - Sperm-mediated transformation of the honey bee, Apis mellifera. AB - Our primary objective was to identify techniques to transform the genome of the honey bee (Apis mellifera) with foreign DNA constructs. The strategy we adopted was to linearize foreign DNA and introduce it with sperm during the instrumental insemination of virgin queen honey bees. We analysed extracts from larvae within the same cohort and isolated the predicted fragment by means of PCR amplification of genomic DNA. Larvae that carried the construct also expressed the introduced DNA. We propagated several transgenic lines for up to three generations, which demonstrates its heritability. Once carried by a queen, the construct can be detected in that queen's larvae over several months. However, there was no evidence of integration of the construct, at least as determined by genomic Southern analysis. Nevertheless, this demonstrates the general viability of the technique for introduction of DNA, and it should be augmented by further use of transposable elements that enhance integration. PMID- 11122472 TI - Wolbachia neither induces nor suppresses transcripts encoding antimicrobial peptides. AB - Wolbachia are intracellular maternally inherited microorganisms that are associated with reproductive abnormalities such as cytoplasmic incompatibility (CI), feminization and parthenogenesis in the various arthropod species they infect. Surveys indicate that these bacteria infect more than 16% of all insect species as well as isopods, mites and nematodes, making Wolbachia one of the most ubiquitous parasites yet described. However, nothing is known about the interactions of this bacterium with the host's immune system. We studied the expression of inducible antimicrobial markers in the adults of two Wolbachia infected insect species, Drosophila simulans and Aedes albopictus. The lack of available immune markers in the mosquito species led us to clone part of the defensin gene from this species, which was found to be very similar to the other mosquito defensins cloned from Anopheles gambiae and Aedes aegypti. Comparisons of the expression pattern of the antibacterial markers between Wolbachia-infected and cured lines, and also between bacteria-challenged and unchallenged adults indicated that Wolbachia does not either constitutively induce or suppress the transcription of these antibacterial genes. In addition, no difference in the transcription of these genes was found between double and single Wolbachia infected strains or between strains in which Wolbachia has different tissue tropisms. PMID- 11122473 TI - Heat-shock protein 90 is down-regulated during pupal diapause in the flesh fly, Sarcophaga crassipalpis, but remains responsive to thermal stress. AB - Heat-shock protein 23 (hsp23) and hsp70 are both known to be strongly up regulated during pupal diapause in the flesh fly Sarcophaga crassipalpis. This prompted us to investigate whether hsp90 was also up-regulated during diapause. To test this possibility, we developed a partial clone of a hsp90 family member for use as a probe in Northern blot hybridization. Both high and low temperature exposure up-regulated hsp90 transcripts in nondiapausing individuals. In contrast to hsp23 and hsp70, hsp90 was down-regulated following entry into diapause, and returned to prediapause levels after diapause termination. The response of hsp90 to heat shock and cold shock remained intact during diapause: both shocks evoked elevated expression. The results indicate differential regulation of hsps during diapause and in response to thermal injury inflicted on diapausing pupae. PMID- 11122474 TI - Molecular characterization of the amplified carboxylesterase gene associated with organophosphorus insecticide resistance in the brown planthopper, Nilaparvata lugens. AB - Widespread resistance to organophosphorus insecticides (OPs) in Nilaparvata lugens is associated with elevation of carboxylesterase activity. A cDNA encoding a carboxylesterase, Nl-EST1, has been isolated from an OP-resistant Sri Lankan strain of N. lugens. The full-length cDNA codes for a 547-amino acid protein with high homology to other esterases/lipases. Nl-EST1 has an N-terminal hydrophobic signal peptide sequence of 24 amino acids which suggests that the mature protein is secreted from cells expressing it. The nucleotide sequence of the homologue of Nl-EST1 in an OP-susceptible, low esterase Sri Lankan strain of N. lugens is identical to Nl-EST1. Southern analysis of genomic DNA from the Sri Lankan OP resistant and susceptible strains suggests that Nl-EST1 is amplified in the resistant strain. Therefore, resistance to OPs in the Sri Lankan strain is through amplification of a gene identical to that found in the susceptible strain. PMID- 11122475 TI - Comparison of esterase gene amplification, gene expression and esterase activity in insecticide susceptible and resistant strains of the brown planthopper, Nilaparvata lugens (Stal). AB - Organophosphorus and carbamate insecticide resistance in Nilaparvata lugens is based on amplification of a carboxylesterase gene, Nl-EST1. An identical gene occurs in susceptible insects. Quantitative real-time PCR was used to demonstrate that Nl-EST1 is amplified 3-7-fold in the genome of resistant compared to susceptible planthoppers. Expression levels were similar to amplification levels, with 1-15-fold more Nl-EST1 mRNA in individual insects and 5-11-fold more Nl-EST1 mRNA in mass whole body homogenates of resistant females compared to susceptibles. These values corresponded to an 8-10-fold increase in esterase activity in the head and thorax of individual resistant insects. Although amplification, expression and activity levels of Nl-EST1 in resistant N. lugens were similar, the correlation between esterase activity and Nl-EST1 mRNA levels in resistant individuals was not linear. PMID- 11122476 TI - Wolbachia infections in native and introduced populations of fire ants (Solenopsis spp.). AB - Wolbachia are cytoplasmically inherited bacteria that induce a variety of effects with fitness consequences on host arthropods, including cytoplasmic incompatibility, parthenogenesis, male-killing and feminization. We report here the presence of Wolbachia in native South American populations of the fire ant Solenopsis invicta, but the apparent absence of the bacteria in introduced populations of this pest species in the USA. The Wolbachia strains in native S. invicta are of two divergent types (A and B), and the frequency of infection varies dramatically between geographical regions and social forms of this host. Survey data reveal that Wolbachia also are found in other native fire ant species within the Solenopsis saevissima species complex from South America, including S. richteri. This latter species also has been introduced in the USA, where it lacks Wolbachia. Sequence data reveal complete phylogenetic concordance between mtDNA haplotype in S. invicta and Wolbachia infection type (A or B). In addition, the mtDNA and associated group A Wolbachia strain in S. invicta are more closely related to the mtDNA and Wolbachia strain found in S. richteri than they are to the mtDNA and associated group B Wolbachia in S. invicta. These data are consistent with historical introgression of S. richteri cytoplasmic elements into S. invicta populations, resulting in enhanced infection and mtDNA polymorphisms in S. invicta. Wolbachia may have significant fitness effects on these hosts (either directly or by cytoplasmic incompatibility) and therefore these microbes potentially could be used in biological control programmes to suppress introduced fire ant populations. PMID- 11122477 TI - Spin-offs from the NASA space program for tumor diagnosis. PMID- 11122478 TI - NASA/American Cancer Society High-Resolution Flow Cytometry Project-I. AB - BACKGROUND: The NASA/American Cancer Society (ACS) flow cytometer can simultaneously analyze the electronic nuclear volume (ENV) and DNA content of cells. This study describes the schematics, resolution, reproducibility, and sensitivity of biological standards analyzed on this unit. METHODS: Calibrated beads and biological standards (lymphocytes, trout erythrocytes [TRBC], calf thymocytes, and tumor cells) were analyzed for ENV versus DNA content. Parallel data (forward scatter versus DNA) from a conventional flow cytometer were obtained. RESULTS: ENV linearity studies yielded an R value of 0.999. TRBC had a coefficient of variation (CV) of 1.18 +/- 0.13. DNA indexes as low as 1.02 were detectable. DNA content of lymphocytes from 42 females was 1.9% greater than that for 60 males, with a noninstrumental variability in total DNA content of 0.5%. The ENV/DNA ratio was constant in 15 normal human tissue samples, but differed in the four animal species tested. The ENV/DNA ratio for a hypodiploid breast carcinoma was 2.3 times greater than that for normal breast tissue. CONCLUSIONS: The high-resolution ENV versus DNA analyses are highly reliable, sensitive, and can be used for the detection of near-diploid tumor cells that are difficult to identify with conventional cytometers. ENV/DNA ratio may be a useful parameter for detection of aneuploid populations. PMID- 11122479 TI - NASA/American Cancer Society High-Resolution Flow Cytometry Project - II. Effect of pH and DAPI concentration on dual parametric analysis of DNA/DAPI fluorescence and electronic nuclear volume. AB - BACKGROUND: In the present paper, we describe the effect of 4', 6-diamidino-2 phenylindole (DAPI) dihydrochloride concentration and pH on the resolution of DNA distribution histograms generated by dual-parametric simultaneous analysis of DNA content and electronic nuclear volume (ENV). METHODS: Nuclei from tissue culture cell lines and frozen human solid tumors were isolated in nuclear isolation media containing different concentrations of DAPI, at various pH levels, and analyzed on a NASA/American Cancer Society (ACS) flow cytometer. Samples stained with propidium iodide/hypotonic citrate and analyzed in a Coulter XL flow cytometer were used for comparison. RESULTS: Nuclei stained with DAPI concentration of 1-3 microg/ml, pH 6.0, gave the best resolution for the detection of the near-diploid and near-tetraploid populations. Simultaneous use of ENV and DAPI/DNA fluorescence under these conditions identified subpopulations that otherwise could not be detected by DNA analysis alone. CONCLUSIONS: Staining at 1-3 microg/ml DAPI, pH 6.0, was optimal for the detection of aneuploid populations, especially the near-diploid and/or near-tetraploid populations in human tumors. PMID- 11122480 TI - NASA/American Cancer Society High-Resolution Flow Cytometry Project - III. Multiparametric analysis of DNA content and electronic nuclear volume in human solid tumors. AB - BACKGROUND: The NASA/American Cancer Society (ACS) flow cytometer can simultaneously measure electronic nuclear volume (ENV) and DNA content of nuclei. The preceding articles in this volume ("NASA/American Cancer Society High Resolution Flow Cytometer Project-I") described the schematics, performance, and procedures used for the preparation of nuclei for analysis on this unit. In the present article, we describe the analysis of selected human tumors using the ratio of ENV/DNA content (nuclear packing efficiency [NPE]). METHODS: Tumor specimens (frozen) were minced with scalpels and stained with 1-10 microg/ml of 4',6-diamidino-2-phenylindole (DAPI) dihydrochloride at pH 6.0-7.2. Trout erythrocytes were used as internal standards. Data on ENV and DNA content were collected in list mode files. Propidium iodide-stained nuclei, analyzed on a Coulter XL cytometer, were used for comparison. RESULTS: Simultaneous measurement of ENV and DNA makes it possible to discriminate between hypodiploid or hyperdiploid tumor cells, as well as to differentiate between near-diploid aneuploid and diploid cells on the basis of their increased ENV. The NPE ratio is a valuable parameter for the detection of small quantities of tumor cells, separating overlapping diploid and aneuploid populations for cell cycle analysis and characterizing the level of differentiation in some tumors. CONCLUSION: NPE analysis provides unique measuring capabilities for the study of human solid tumors by flow cytometry. PMID- 11122481 TI - A dynamic inline sample thermoregulation unit for flow cytometry. AB - BACKGROUND: Flow cytometry is a valuable tool for the study of cellular and molecular interactions. We sought to develop instrumentation that would allow accurate and precise inline dynamic temperature control of flow cytometry samples in order to expand the analytical capabilities of flow cytometry. METHODS: Using a temperature controller, DC power supply, and a Peltier module, we designed a temperature control circuit that regulates the temperature of a heat transfer block. The heat transfer block surrounds the sample line to efficiently heat and cool the sample. We attached DNA oligomers to microspheres and observed the denaturation of fluorescently labeled complementary oligomers to verify that the sample was achieving the desired temperature. RESULTS: The inline thermoregulation unit can ramp between 30 and 95 degrees C (>2 degrees C per second) within 30 s repeatedly. The accuracy of the instrument was verified by comparing the observed melting temperatures of the oligomer sets to the predicted values. These values varied within 6% of the predicted and were reproducible over a variety of conditions. CONCLUSIONS: The apparatus we constructed accurately and dynamically controls the sample temperature of inline samples. Flow cytometers equipped with our inline thermoregulation unit will be able to conveniently use temperature as a robust experimental parameter. PMID- 11122482 TI - Cell analysis system based on immunomagnetic cell selection and alignment followed by immunofluorescent analysis using compact disk technologies. AB - BACKGROUND: Although the flow cytometer has become the standard in cell analysis, it has limitations. Recently, we introduced a new cell analysis method based on immunomagnetic selection and aligning of cells. No flow system is needed and cell analysis can be performed in whole blood. METHODS: Whole blood is incubated with fluorescent labels and immunomagnetic nanoparticles. The blood is injected into a capillary that is in a strong magnetic field. The immunomagnetic-labeled cells move upward and align themselves along ferromagnetic lines present on the upper surface of the capillary. An optical focus and tracking system analogous to that used in a conventional compact disk player focuses a 635-nm laser-diode on the magnetically aligned cells. The emitted fluorescence signals are projected on two photomultipliers. Allophycocyanin (APC)-labeled CD4 (CD4-APC) and Cyanin5.5 (Cy5.5)-labeled CD8 (CD8-Cy5.5) antibodies and Oxazine750, all red excited, are used as fluorescent labels. RESULTS: A differential white blood cell count performed in whole blood is obtained using the CD4-APC in combination with Oxazine750. The results are compared with the Technicon-H1 hematology analyzer. Correlation coefficients of 0.91 for neutrophilic granulocytes, 0.93 for lymphocytes, 0.93 for monocytes, and 0.96 for eosinophilic granulocytes were obtained. Immunofluorescence is demonstrated using CD4-APC and CD8-Cy5.5. The absolute counts obtained for CD4+ and CD8+ are compared with the Coulter Epics XL flow cytometer. Correlation coefficients of, respectively, 0.91 and 0.94 were obtained. CONCLUSION: We conclude that our system is as capable as a standard flow cytometer or hematology analyzer for a reliable routine white blood cell analysis, including immunophenotyping, and can be used as an easy-to-handle disposable white blood cell test. PMID- 11122483 TI - Caffeine dissociates complexes between DNA and intercalating dyes: application for bleaching fluorochrome-stained cells for their subsequent restaining and analysis by laser scanning cytometry. AB - BACKGROUND: Removal of the nucleic acid-bound fluorochrome is desirable when stained cells have to be reanalyzed using other fluorochromes. It is also often desirable to remove DNA-bound antitumor drugs from drug-treated cells, to improve cell staining. We have previously observed that in aqueous solutions, the methylxanthine caffeine (CFN) decreases interactions between planar aromatic molecules such as intercalating dyes or antitumor drugs and nucleic acids. The aim of this study was to explore whether this property of CFN can be utilized to remove the DNA-bound intercalating dyes propidium iodide (PI) or 7 aminoactinomycin D (7-AAD) from the cells and whether the bleached cells can be restained and reanalyzed. METHODS: HL-60 cells were fixed in 70% ethanol and their DNA was stained with PI or 7-AAD. The cells were then rinsed with a 0.05 M solution of CFN in phosphate-buffered saline (PBS) or with PBS alone. The decrease in intensity of cell fluorescence during rinsing was measured by laser scanning cytometry (LSC) to obtain the bleaching kinetics of individual cells. The bleached cells were then restained with PI, 7-AAD, or the protein-specific fluorochrome sulforhodamine 101(S101). Their fluorescence was measured again by LSC. In addition, free DNA was subjected to gel electrophoresis, DNA bands in the gels were stained with ethidium bromide (EB), and the gels were rinsed with a solution of CFN or PBS to bleach the DNA band's fluorescence. RESULTS: Rinsing the PI or 7-AAD-stained cells with solutions of CFN led to nearly complete removal of PI and a more than 75% decrease in 7-AAD fluorescence after 10 min. The rinse with PBS decreased the PI cell fluorescence intensity by less than 30% and the 7-AAD fluorescence by about 50%. The differences in kinetics of PI or 7 AAD removal by CFN from G2/M versus G1 cells suggest that these intercalators bind more strongly to DNA in chromatin of G2/M than G1 cells. The CFN-bleached cells were then successfully stained with S101 and again with PI or 7-AAD. The bivariate analysis of the LSC merged files of the cells sequentially stained with PI and S101 revealed typical DNA/protein distributions. The fluorescence of EB stained DNA bands in gels was also nearly completely removed by rinsing gels in 0.05 M CFN; PBS alone had a distinctly lesser effect. CONCLUSION: Solutions of CFN can dissociate the DNA-bound PI, 7-AAD, EB, and possibly other intercalating fluorochromes. The bleached cells can be restained and reanalyzed by LSC. This approach can also be used to remove such fluorochromes from nucleic acids immobilized in gels and perhaps in other solid matrices. Analysis of the kinetics of fluorochrome removal from cells can possibly be used to study their binding affinities to nucleic acids in situ. PMID- 11122484 TI - A strategy combining flow sorting and comparative genomic hybridization for studying genetic aberrations at different stages of colorectal tumorigenesis in ulcerative colitis. AB - BACKGROUND: DNA aneuploidy has been shown to increase the risk of developing dysplasia in ulcerative colitis (UC) and is related to tumorigenesis in the colorectum. Therefore, it is of particular interest to study genetic aberrations behind DNA aneuploidization during colorectal carcinogenesis. We wanted to elucidate further the relationship between mucosal morphology and DNA aberrations in UC. METHODS: DNA flow cytometry was applied to multiple lesions including regenerative, dysplastic, and carcinomatous mucosa from the colectomy specimen of a male patient with long-standing UC. The lesions harbored multiple DNA aneuploid stemlines that were subjected to flow sorting. We analyzed gene alterations by degenerate oligonucleotide primer (DOP; universal primers) polymerase chain reaction (PCR)-based comparative genomic hybridization (CGH) and fluorescent in situ hybridization (FISH) in diploid and aneuploid sorted cells. RESULTS: DOP-PCR based CGH shows gains and losses that can be verified by FISH. We show that with this approach one can study genetic evolution of distinct DNA diploid and aberrant subpopulations through defined stages of colorectal tumorigenesis. This includes getting information related to tumor heterogeneity that cannot be obtained by CGH with DNA extracted from nonsorted cell populations. Genetic imbalance was also detected in diploid nondysplastic flow-sorted mucosal cells from the same bowel. CONCLUSIONS: Similar gains and losses were found in aneuploid dysplasias and carcinomas at widely separated locations in the same bowel, indicating a common selection pressure in different areas of the same bowel. The common aberrations may be of importance for progression from dysplasia to carcinoma. PMID- 11122485 TI - Proof without prejudice revisited: immunofluorescence histogram analysis using cumulative frequency subtraction plus ratio analysis of means. AB - BACKGROUND: Apart from the work of Lampariello and colleagues (Cytometry 15:294 301, 1994; Cytometry 32:241-254, 1998), very little analytical work has been carried out for analysis of immunofluorescence distributions containing an overlapping mixture of labeled and unlabeled cells. The methods developed tend to rely on fitting theoretical distributions to the relevant populations. However, the method described here attempts to produce an analytical solution. METHODS: A new method for immunofluorescence histogram analysis is presented. It uses cumulative frequency distribution subtraction of the test sample from the control to predict the mean of a labeled cell component embedded within a histogram containing unlabeled cells. Ratio analysis of means (RAM) was then carried out to calculate the labeled fraction. The results were submitted to Kolmogorov-Smirnov analysis and Student's t-test for validation at a given level of probability. RESULTS: The method was developed with a data set exhibiting a small "positive" shoulder, which was predicted to contain a labeled fraction comprising 8.0% of the total at the 99% confidence limit. It was then tested with data analyzed and published previously where the Johnson Su family of distributions was used in curve fitting. CONCLUSIONS: There was good agreement between the known and predicted proportions of labeled cells. However, the method is dependent on the symmetry of the distributions. Some minor systematic errors were encountered due, in part, to skewed experimental distributions. PMID- 11122486 TI - Characterization of cytokine interactions by flow cytometry and factorial analysis. AB - BACKGROUND: Multiple cytokines are required for the growth and development of hematopoietic cells. The effect of many cytokines depends on the activity of other signaling pathways. These interactions are quantified using factorial experimental design and analysis. METHODS: Human umbilical cord blood (HUCB) CD34+ cells were cultured in fully defined media containing various combinations of recombinant cytokines as defined by resolution IV factorial (2(7-3)(IV)) or full factorial (2(4)) design experiments. The cytokines studied were stem cell factor (SCF), interleukin (IL)-3, megakaryocyte growth and development factor (MGDF), granulocyte-colony stimulating factor (G-CSF), Flt-3 ligand, IL-6, IL-11, and erythropoietin (EPO). In vitro cell divisions were tracked by staining CD34+ cells with 5-(and-6)-carboxyfluorescein diacetate, succinimidyl ester, followed by flow cytometric analysis at 4 days of culture. In separate experiments, lineage commitment and differentiation were determined at 7 days by immunophenotype. RESULTS: In addition to the main effects of single cytokines, cytokine interactions were identified. There was a negative interaction between IL-3 and MGDF that resulted in a less than additive effect of these factors on erythroid and megakaryocytic development. The effect of Flt-3 ligand and SCF factor on CD34+ cell production was also less than additive, although the response to both cytokines was greater than single cytokines. The only positive interaction that was identified was between EPO and SCF, which resulted in the synergistic production of erythroid cells. CONCLUSIONS: Factorial analysis provides a powerful methodology to study the integration of multiple signals at the cellular and molecular level. PMID- 11122487 TI - Quantitative assessment of phagocytic activity of hemocytes in the prawn, Penaeus merguiensis, by flow cytometric analysis. AB - BACKGROUND: The blood cells of crustaceans are involved in phagocytosis of invading microorganisms, contributing to their defense mechanisms. In this study, phagocytic activity of hemocytes of the prawn, Penaeus merguiensis, was quantitated by means of flow cytometric analysis. METHOD: This study was done in vitro. Hemolymph, which was extracted from prawns, was mixed with an equal volume of anticoagulant. Heat-killed Escherichia coli prestained with propidium iodide (PI) was then added. Hemocytes were fixed at various time intervals for flow cytometric analysis. This study was supplemented with electron micrographs using transmission electron microscopy (TEM), which showed three populations of hemocytes. RESULTS: It was observed that those hemocytes that were more active engulfed and digested bacteria readily, thus having higher red fluorescence intensity. The phagocytic activity was expressed as fluorescence unit or engulfed E. coli number per hemocyte. CONCLUSIONS: With this approach, the phagocytic and cellular activity of individual hemocyte populations could be studied quantitatively. PMID- 11122488 TI - A single nocturnal exposure to 2-7 millitesla static magnetic fields does not inhibit the excretion of 6-sulfatoxymelatonin in healthy young men. AB - The present study sought to evaluate possible acute effects on 6 sulfatoxymelatonin (aMT6s) excretion, a surrogate for melatonin levels in blood, in volunteers exposed to static magnetic fields with flux densities representative for workers in light metal reduction plants and operators of medical MRI in hospitals. Eleven healthy male volunteers (23-43 years) participated. Urine samples were collected for two consecutive 24 h periods from 22:00 hours day 1 (exposure day) through day 2 (day after exposure) and then for 24 h from 07:00 hours on day 7 (control day). On the day of exposure the subjects slept in the MRI room from 22:00 hours until 07:00 hours next morning, thus receiving a 9 h exposure to the magnetic field (2-7 mT). On the day after exposure and on the control day, they slept at home and otherwise performed their ordinary daily activities. Total daily urine production was collected in four parts: 22:00-07:00 hours, 07:00-11:00 hours, 11:00-18:00 hours, 18:00-22:00 hours, and the volume for each interval was measured and recorded. Samples were transferred to coded bottles and frozen for later RIA analysis of aMT6s. Pairs of values of mean hourly aMT6s excretion, both diurnal and for the four daily intervals, were compared using two-sided Wilcoxon signed ranks test. The day of exposure and the day after exposure were not significantly different from the control day, either for the total diurnal secretion or the interval data. In summary, the study shows no association between a single nocturnal exposure to a static magnetic field of strength 2-7 mT and excretion of aMT6s in urine. PMID- 11122489 TI - Cows, ground surface potentials and earth resistivity. AB - The "stray voltage" problem on dairy farms is discussed briefly. By reference to published literature it is shown that a "step voltage" (SV), i.e., a potential difference between front and hind hooves of a dairy cow, much less than the often quoted threshold value of 0.5 V, when applied for long periods of time, could possibly affect cow health and milk production. Values as low as approximately 10 mV could conceivably be significant. A measurement program carried out on 19 representative Minnesota dairy farms during the summer of 1997 is described. Nine farms had above average (HP) and 10 below average (LP) milk production. Results show that SV was 4.2 times higher on the LP than HP. However, only three farms had SV greater than 9 mV, and low milk production on these farms could possibly be due to absence of recommended vaccinations rather than high SV. Soil resistivity rho measured in the farm fields was 3.4 times larger on the LP than on HP. The possible origin of SV in relation to electric distribution systems within and to farms is discussed. Relations between SV and rho are analyzed. Conditions are specified under which SV in the barn must be related to rho measured in the field, rather than the rho of the concrete floor of the barn. It is suggested that laboratory research is necessary to establish the significance for cow health and milk production of long term exposure to low SV levels. PMID- 11122490 TI - Single, brief exposure to a 50 Hz magnetic field does not affect the performance of an object recognition task in adult mice. AB - A number of studies have shown that power frequency magnetic fields may affect spatial memory functions in rodents. An experiment was performed using a spontaneous object recognition task to investigate if nonspatial working memory was similarly affected. Memory changes in adult, male C57BL/6J mice were assessed by measuring the relative time within which the animals explored familiar or novel stimulus objects. Between initial testing and retesting, the animals were exposed for 45 min to a 50 Hz magnetic field at either 7.5 microT, 75 microT or 0.75 mT. Other animals were sham-exposed with ambient fields of less than 50 nT. No significant field-dependent effects on the performance of the task were observed at any flux density (for all measures, P > 0.05). These data provide no evidence to suggest that nonspatial working memory was affected in mice by acute exposure to an intense 50 Hz magnetic field. PMID- 11122491 TI - Review of russian literature on biological action of DC and low-frequency AC magnetic fields. AB - This review considers the Russian scientific literature on the influence of weak static and of low-frequency alternating magnetic fields on biological systems. The review covers the most interesting works and the main lines of investigation during the period 1900 to the present. Shown here are the historical roots, beginning with the ideas of V. Vernadsky and A. Chizhevsky, which led in the field of Russian biology to an increasing interest in magnetic fields, based on an intimate connection between solar activity and life on the Earth, and which determined the peculiar development of Russian magnetobiology. The variety of studies on the effects of magnetic storms and extremely low-frequency, periodic variations of the geomagnetic field on human beings and animals as well as on social phenomena are described. The diverse experiments involving artificial laboratory magnetic fields acting on different biological entities under different conditions are also considered. A series of theoretical advances are reviewed that have paved the way for a step-by-step understanding of the mechanisms of magnetic field effects on biological systems. The predominantly unfavorable influence of magnetic fields on living beings is shown, but the cases of favorable influence of magnetic fields on human beings and lower animals are demonstrated as well. The majority of Russian investigations in this area of science has been unknown among the non-Russian speaking audience for many reasons, primarily because of a language barrier. Therefore, it is hoped that this review may be of interest to the international scientific community. PMID- 11122492 TI - Osmolality dependence of erythrocyte sedimentation and aggregation in a strong magnetic field. AB - In order to specify the major determinant of the magnetic enhancement of erythrocyte sedimentation observed previously, the dependence of erythrocyte sedimentation rate (ESR) on osmolality was measured under a strong magnetic field. Even at hypotonic osmolality, an increase in ESR due to aggregation was observed in plasma solution as compared with that without aggregation in saline solution. However, the magnetic field did not enhance ESR at hypotonic osmolality, when the cell shape was an isotropic sphere (spherocyte). Thus, we narrowed our search to a mechanism that would explain the enhanced ESR found specifically in anisotropic erythrocytes. It was concluded that the major determinant can only work for anisotropic erythrocytes and is a magnetic field induced increase in an intermembrane adhesive area due to magnetic orientation of anisotropic erythrocytes. PMID- 11122493 TI - A strong constant magnetic field affects muscle tension development in bullfrog neuromuscular preparations. AB - Effects of a constant magnetic field (CMF) of 0.65 T on muscle tension over 9 h were studied in the neuromuscular preparation of the bullfrog sartorius muscle. Tension was developed every 30 min by stimulation of the sciatic nerve (nerve stimulation) or of the sartorius muscle itself (muscle stimulation). In sciatic nerve stimulation, tension decreased rapidly for the first 3-4 h at a similar rate in both test (exposed to CMF) and control muscles. However, the rate of decrease became smaller and almost leveled off after 3-4 h in the test muscles, whereas tension continued to decrease monotonically in control muscles. The slope of the decrease for these later periods was significantly different between the test and the control conditions. Accordingly, tension was larger in test than in control muscles. In muscle stimulation, tension decreased monotonically from the start of experiments in control muscles, while tension in test muscles maintained their initial values for almost 3 h. Thereafter they started to decrease with a similar rate to the control. Hence, tension was always larger in test than in control muscles. A similar pattern of temporal change was observed for the rate of rise of the maximum tension for nerve or muscle stimulation. However, a significant difference was detected only in the case of muscle stimulation. The present results showed that a strong CMF of 0.65 T had biological effects on tension development of the bullfrog sartorius muscle by stimulation of the sciatic nerve as well as by stimulation of muscle itself. The presence of a small AC magnetic field component leaves open the possibility of an AC, rather than a CMF effect. PMID- 11122494 TI - Orientation of bull sperms in static magnetic fields. AB - The orientation of bull sperm cells in static magnetic fields was investigated by microscopic observation. The bull sperm, which has a very flat head, was fixed and its motion was stopped by glutaraldehyde. It was oriented with the whole body and the flat head perpendicular to the direction of the magnetic field. The diamagnetic cell components, such as the cell membrane, the DNA in the head, and the microtubule in the tail, were thought to contribute to this orientation, because bull sperm does not have paramagnetic components. For quantitative measurement of the orientation, the intensity of transmitted light through glutaraldehyde-fixed bull sperm suspension in a photometric cell was determined. The intensity changed slightly in proportion to the mean degree of orientation of the sperms. It increased sigmoidally depending on the intensity of the magnetic field and reached 100% at just below 1 T. The magnetic orientation is very strong in comparison to that of erythrocytes or platelets. By studying the response of the bull sperm to the magnetic field, it will be possible to understand its microstructure in more detail. PMID- 11122495 TI - How might spatial nonuniformity of dose in a homogeneous biological system affect its total response? AB - The total response of a homogeneous biological system to a fixed total dose of a biological agent is modeled by dividing the system into N cubical voxels, each of which can be associated with an individual dose D(n) and an individual response R(n) =F(D(n)). Among the results shown are the following: A. (Voxel Theorem). Let the average dose D(avg) be held fixed as the dose distribution is shifted from uniform u to arbitrary a. Then, if F' > or = 0 over [D(min), D(max)] and R = summation operator (n = 1)(N) R(n), a sufficient condition that NF(D(avg)) = R(u) < or = R(a) is that F be a concave-upwards function of dose; that is, F" > or = 0 over [D(min), D(max)]. B. If F' is constant over [D(min), D(max)], then R(a) = R(u). That is, the total response is a function of D(avg) only. The applications of these (and other) results are illustrated by examples from bioelectromagnetics. PMID- 11122496 TI - Preface. PMID- 11122497 TI - Proportional hazards model with random effects. AB - We propose a general proportional hazards model with random effects for handling clustered survival data. This generalizes the usual frailty model by allowing a multivariate random effect with arbitrary design matrix in the log relative risk, in a way similar to the modelling of random effects in linear, generalized linear and non-linear mixed models. The distribution of the random effects is generally assumed to be multivariate normal, but other (preferably symmetrical) distributions are also possible. Maximum likelihood estimates of the regression parameters, the variance components and the baseline hazard function are obtained via the EM algorithm. The E-step of the algorithm involves computation of the conditional expectations of functions of the random effects, for which we use Markov chain Monte Carlo (MCMC) methods. Approximate variances of the estimates are computed by Louis' formula, and posterior expectations and variances of the individual random effects can be obtained as a by-product of the estimation. The inference procedure is exemplified on two data sets. PMID- 11122498 TI - Assessing the potential for bias in meta-analysis due to selective reporting of subgroup analyses within studies. AB - Subgroup analysis is frequently used to investigate heterogeneity in meta analysis. Subgroup data are not always available, and researchers should record what data were available for each trial. If data were not available, and it is known that the subgroup data were collected, the potential for selective reporting should be considered. Bias due to selective publishing of reports is widely recognized in meta-analysis. In contrast, selective reporting within studies is little discussed but potentially important. We explored this problem by evaluating the effect of potential bias in subgroup analysis due to within study selective reporting with an existing meta-analysis. The review addressed malaria chemoprophylaxis in pregnancy. The conclusion in the original review, that benefit is limited to primigravidae, was based on subgroup analysis using the three trials out of five which reported on subgroups. We developed a method of sensitivity analysis that imputes data for the missing subgroups to assess the robustness of the results and the conclusions drawn. In this particular example, our analysis indicates that the estimate of effect reported in the review is most likely to overestimate the true effect and the conclusion that benefit is limited to primigravidae may be false. PMID- 11122499 TI - Future directions of research in statistical genetics. AB - From a global perspective, two external developments are having dramatic effects upon the field of statistical genetics: improved genetic data, for example, human DNA sequence, and new technologies, for example, microarray technology. Meiotic mapping techniques will have to be adapted to benefit from the improved data, for example, allelic association studies have to be extended to multiple markers to profit from the new genetic map of SNP markers. Changing technology has led to ever-increasing knowledge about gene function which has enabled novel gene mapping strategies which we refer to as functional mapping. Functional mapping has great potential for mapping complex disease genes since it uses pathway fractions to intermediate between genotype and phenotype information. Methods used in whole-genome gene expression studies are used to illustrate concepts of functional mapping. PMID- 11122500 TI - Familial associations of lipid profiles: a generalized estimating equations approach. AB - Elevated plasma levels of apolipoproteins A1 (apoA1) and B (apoB) are important protective factors and risk factors, respectively, for atherosclerosis and coronary heart disease. It is well known that both apoA1 and apoB reveal strong familial aggregation. Our goal was to investigate whether exogenous variables influence these associations. We used marginal regression models for the mean and association structure (generalized estimating equations 2; GEE2) to analyse data from 1435 family members within 469 families of different sizes included in the Donolo-Tel Aviv Three-Generation Offspring Study. The usual robust variance matrix was approximated by extensions of jack-knife estimators of variance to GEE2 models. Estimation of standard errors in models with quite complex correlation structures was possible using this approach. All analyses were easily carried out using a menu-driven stand-alone software tool for marginal regression modelling. We demonstrate that a variety of hypotheses can be tested using Wald statistics by modelling regression matrices for the association structure. We show that correlation for apoB between parent-offspring pairs increased with decreasing age difference and that pairs with individuals of the same gender had more similar apoA1 levels than individuals of different gender. Associations between different relative pairs did not all agree with those expected from differences in kinship coefficients. The analysis using GEE2 models revealed structures that would not have been detected by other models and should therefore be used in addition to traditional approaches of analysing family data. GEE2 should be considered a standard method for the investigation of familial aggregation. PMID- 11122501 TI - Hierarchical models in generalized synthesis of evidence: an example based on studies of breast cancer screening. AB - Evidence regarding the potential benefits of a particular health care intervention is often available from a variety of disparate sources. However, formal synthesis of such evidence has traditionally concentrated almost exclusively on that derived from randomized studies, although for a range of conditions the randomized evidence will be less than adequate due to economic, organizational or ethical considerations. In such situations a formal synthesis of the evidence that is available from observational studies can be valuable whilst awaiting higher quality evidence from randomized trials. Consideration of randomized studies alone may be appropriate when assessing the efficacy of an intervention, but assessment of the effectiveness of such an intervention within a more general target population may be improved by consideration of evidence from non-randomized studies as well. Standard meta-analysis methods may allow for both within- and between-study heterogeneity; however when multiple sources of evidence are considered an extra level of complexity is introduced, namely study type. One possible solution to the problem of making inferences, particularly regarding an overall population effect, in such situations is to model the heterogeneity, both quantitative and qualitative, using a Bayesian hierarchical model. The hierarchical nature of such models specifically allows for the quantitative within and between sources of heterogeneity, whilst the Bayesian approach can accommodate a priori beliefs regarding qualitative differences between the various sources of evidence. The use of such methods in practice is illustrated in the context of screening for breast cancer; in this example evidence is available from both randomized clinical trials and observational studies. A particular appeal of a Bayesian approach for this type of problem lies in the prediction of future benefits likely to be observed in a target population. This approach to health service monitoring in general is discussed. PMID- 11122502 TI - A continuation ratio random effects model for repeated ordinal responses. AB - This paper presents methods of analysis for ordinal repeated measures. We use a generalization of the continuation ratio model with a random effect. We handle frailty by using a normal distribution and also a non-parametric distribution. The methodology is readily implemented in existing software and flexible enough to treat large data sets with uncommon time measurements as well as different numbers of repeated measures for each individual. The models are used to investigate how a group of explanatory variables influences the overall condition of patients treated for breast cancer. PMID- 11122503 TI - Some practical issues in the design, monitoring and analysis of a sequential randomized trial in pressure sore prevention. AB - A sequential double blind (assessor and patient) triangular design was used to compare the incidence of pressure sores following elective major surgery among patients lying on a standard foam mattress with those on a dry visco-elastic polymer pad during their operation. A total of 446 patients were recruited into the trial between 1994 and 1996. Interim analyses were carried out after 181 patients were entered into the trial and then subsequently after approximately every 100 patients recruited. The trial unexpectedly reached a stopping boundary at the first interim analysis, however the Independent Data Monitoring Committee recommended continuation of the trial. They were concerned that there was a need for a larger definitive trial and about an apparent treatment by centre interaction. They required a substudy to be undertaken to further validate the subjective endpoint, and that further sensitivity analyses of the main trail endpoint should be carried out in the second interim analysis. The trial was stopped at the third interim analysis when again a stopping boundary was crossed indicating that the gel pad was associated with significantly fewer pressure sores than the standard mattress (log odds ratio -0.7, (95 per cent confidence interval (CI), -1.28, -0.11), p=0.02) (estimate CI, p-value adjusted for group sequential conduct). The design, monitoring and analysis of this trial will be presented as an example of the practical problems or non-problems encountered for the local hospitals, for the trials unit, for the data monitoring committee and for the funding committee. PMID- 11122504 TI - Validation, calibration, revision and combination of prognostic survival models. AB - The problem of assessing the validity and value of prognostic survival models presented in the literature for a particular population for which some data has been collected is discussed. Methods are sketched to perform validation through 'calibration', that is by embedding the literature model in a larger calibration model. This general approach is exemplified for x-year survival probabilities, Cox regression and general non-proportional hazards models. Some comments are made on basic structural changes to the model, described as 'revision'. Finally, general methods are discussed to combine models from different sources. The methods are illustrated with a model for non-Hodgkin's lymphoma validated on a Dutch data set. PMID- 11122505 TI - A multilevel model framework for meta-analysis of clinical trials with binary outcomes. AB - In this paper we explore the potential of multilevel models for meta-analysis of trials with binary outcomes for both summary data, such as log-odds ratios, and individual patient data. Conventional fixed effect and random effects models are put into a multilevel model framework, which provides maximum likelihood or restricted maximum likelihood estimation. To exemplify the methods, we use the results from 22 trials to prevent respiratory tract infections; we also make comparisons with a second example data set comprising fewer trials. Within summary data methods, confidence intervals for the overall treatment effect and for the between-trial variance may be derived from likelihood based methods or a parametric bootstrap as well as from Wald methods; the bootstrap intervals are preferred because they relax the assumptions required by the other two methods. When modelling individual patient data, a bias corrected bootstrap may be used to provide unbiased estimation and correctly located confidence intervals; this method is particularly valuable for the between-trial variance. The trial effects may be modelled as either fixed or random within individual data models, and we discuss the corresponding assumptions and implications. If random trial effects are used, the covariance between these and the random treatment effects should be included; the resulting model is equivalent to a bivariate approach to meta analysis. Having implemented these techniques, the flexibility of multilevel modelling may be exploited in facilitating extensions to standard meta-analysis methods. PMID- 11122506 TI - Diagnostic strategies for the histological examination of muscle biopsy specimens for the assessment of vasculitis in rheumatoid arthritis. AB - For the diagnosis of rheumatoid vasculitis (RV), histological examination of a blindly-taken muscle biopsy is advocated. If fibrinoid necrosis (FN) is observed in one or more tissue sections of the biopsy the diagnosis of RV is confirmed. The diagnostic value of such histological investigation depends on the prevalence of FN in biopsies of RV patients, the number of tissue sections that are investigated, and the sampling design with which the tissue sections are obtained from the biopsy. In this paper we determine the mathematical relation between the sensitivity of the RV diagnosis and these three factors for four different models for the FN distribution in RV biopsies. The goodness-of-fit of these models was assessed by analysing 18 829 tissue sections of the biopsies of 56 patients, among which were 24 (otherwise histologically proven) RV patients. It appeared that the sensitivity was moderate: FN was observed in only 14 out of 24 (58 per cent) muscle biopsies of the RV patients. The prevalence of FN in the tissue sections of those 14 biopsies was low, about 17 per cent according to the best fitting model. We proved that the sampling design, maximizing the minimum distance between tissue sections (equidistant sampling) was optimal, and that with such optimal sampling, examination of about 20 tissue sections was sufficient. With practical sampling designs, however, considerably more tissue sections had to be inspected. FN appeared to cluster in the RV biopsies, and a first-order Markov model satisfactorily described the (auto)correlation between adjacent tissue sections in RV biopsies. PMID- 11122507 TI - Sequential analysis of matched dichotomous data from prospective case-control studies. AB - Sequential analysis of randomized controlled clinical trials and epidemiological prospective (matched) case-control studies can have ethical or economical advantages above a fixed sample size approach. It offers the possibility to stop early when enough evidence for an apparent effect of the risk factor or lack of the expected effect is achieved. In clinical trials it is well accepted to stop the trial early in favour of the alternative hypothesis. In epidemiological studies, in general, the need is not felt to stop early in case of a clear exposure effect. Little attention has been paid, however, to early stopping and accepting the null hypothesis. In metabolic epidemiological studies, where analysis destroys the biological material, the question of efficient use of samples, for example, those stored in a biobank, becomes crucial. Also a slow accrual of cases or the costs of follow-up of a cohort nested study can make it desirable to stop a study early once it becomes clear that no relevant exposure effect will be found. Matching can further reduce the amount of information necessary to reach a conclusion. We derived test statistics Z (efficient score) and V (Fisher's information) for the sequential analysis of studies with dichotomous data where each case can be matched to one or more controls. A variable matching ratio is allowed. These test statistics can be entered into the software PEST to monitor the course of the study. The double sequential probability ratio test and the double triangular test were evaluated with simulated data for odds ratios equal to 1.5, 2.0 and 2.5 and various type I and type II error probabilities both under H(0) and under H(1). Our simulations resulted in average and median values for the amount of information (V) that are far less than those for a fixed sample size study. Efficiency gain ranges from 32 per cent to 60 per cent. The proposed sequential analysis was applied in an investigation on the possible relationship between the polymorphism of the MTHFR gene and rectal cancer in a cohort of women with cases matched by age to one and to three controls. A sequential analysis of matched data can lead to early stopping in favour of H(0) or H(1), thus conserving valuable resources for future testing. A sequentially designed study can be more economical and less arbitrary than a study that makes use of conditional power or conditional coverage probability calculations to decide early stopping. PMID- 11122508 TI - A multi-state model for evolution of intensive care unit patients: prediction of nosocomial infections and deaths. AB - Nosocomial (hospital-acquired) infections are very frequent in intensive care units (ICU). The risk of death after severe infection is high, but the precise rate of death in ICU attributable to nosocomial infection is not known. The goal of this project was to build a statistical model to predict the occurrence of nosocomial infections in ICU and the outcome of the patients. We collected data on 676 consecutive patients admitted to an ICU for more than 24 hours between 1993 and 1996. The following data were collected for each patient: history; clinical examination at entry; subsequent infections; outcome. A multi-state heterogeneous semi-Markov model was determined and then validated; the initial data set was randomly split into two groups: two-thirds (450 patients) to build the model and one-third (226 patients) to validate it. The model defined five states: ICU admission; first simple infection; first complicated infection; death, and discharge from the ICU. Transitions between these states determined nine different events. The global model of patient histories can be divided into nine survival models, each corresponding to one of these events. The possible events from a given state were considered to be competing. Since many risk factors induced non-proportional hazard functions, piecewise exponential models were used to model event occurrence. The effect of continuous covariates on hazard functions has been described with a non-parametric method that enables non linear relations to be shown. Among other things, the model allows patients' post admission histories to be predicted from data available at ICU admission. The bootstrap estimator of the attributable risk of death due to simple or complicated nosocomial infections is 44.2 percent (95 percent CI 26.0-61.6 percent). We were also able to characterize the most highly exposed patients, those who comprise the high-risk group on whom prevention efforts must be focused. PMID- 11122509 TI - Concordance between ratings using different scales for the same variable. AB - Qualitative variables are commonly measured by means of different types of rating scales that produce ordered categorical data. The number of response categories in a scale can be chosen arbitrarily according to different operational definitions of the attribute to be measured. In the present paper classical measures based on the difference between the probabilities of concordant and discordant pairs of classifications were compared with a novel approach for assessment of order consistency between rating scales. In the new approach, illustrated by an assessment of pain, the concordant and discordant pairs are related to the pattern of total order consistency, irrespective of the scaling and the categorical distributions. The level of order consistency between a visual analogue scale (VAS) and a discrete scale for the same variable was evaluated. The non-linear properties of VAS assessments were demonstrated. The inter-scale consistency between the discrete scale and different approaches to grouping VAS responses into discrete scales showed that equidistant rescaling of VAS assessments resulted in an inter-scale bias. PMID- 11122510 TI - Baseline risk as predictor of treatment benefit: three clinical meta-re-analyses. AB - A relationship between baseline risk and treatment effect is increasingly investigated as a possible explanation of between-study heterogeneity in clinical trial meta-analysis. An approach that is still often applied in the medical literature is to plot the estimated treatment effects against the estimated measures of risk in the control groups (as a measure of baseline risk), and to compute the ordinary weighted least squares regression line. However, it has been pointed out by several authors that this approach can be seriously flawed. The main problem is that the observed treatment effect and baseline risk measures should be viewed as estimates rather than the true values. In recent years several methods have been proposed in the statistical literature to potentially deal with the measurement errors in the estimates. In this article we propose a vague priors Bayesian solution to the problem which can be carried out using the 'Bayesian inference using Gibbs sampling' (BUGS) implementation of Markov chain Monte Carlo numerical integration techniques. Different from other proposed methods, it uses the exact rather than an approximate likelihood, while it can handle many different treatment effect measures and baseline risk measures. The method differs from a recently proposed Bayesian method in that it explicitly models the distribution of the underlying baseline risks. We apply the method to three meta-analyses published in the medical literature and compare the results with the outcomes of the other recently proposed methods. In particular we compare our approach to McIntosh's method, for which we show how it can be carried out using standard statistical software. We conclude that our proposed method offers a very general and flexible solution to the problem, which can be carried out relatively easily with existing Bayesian analysis software. A confidence band for the underlying relationship between true effect measure and baseline risk and a confidence interval for the value of the baseline risk measure for which there is no treatment effect are easily obtained by-products of our approach. PMID- 11122511 TI - Silage and animal health. AB - The process of preserving crops by fermentation in silos is under the control of the farmer to a much lesser degree compared to the level of control by the manufacturer over the production of other fermented foods, such as cheese and yoghurt. Additives designed to direct the extent and pattern of the fermentation are relatively unpopular in most countries, and their use is not guaranteed to remove the risk of undesirable components in silage. Hazards to animal health associated with silage fall into three categories: (1) undesirable micro organisms e.g. Listeria, enterobacteria, clostridia and moulds; (2) undesirable chemicals, e.g. mycotoxins, and (3) excess acidity and other metabolic disorders. In some regions of Europe, the production of silage is discouraged or prohibited because of the risk of undesirable microbes. The princIpal risk in these areas is that of the secondary fermentation of cheese made from milk contaminated by bacterial spores, rather than a direct hazard of contaminated silage to animal health. With the possible exception of high dry matter silage conserved in large bales, respiratory hazards to animals from moulds and their spores generally are less from silage than hay. Mycotoxins and phytoestrogens may survive the ensiling period and constitute risks to animal health. Relatively little is known about the epidemiology of diseases that may be linked to undesirable chemicals and excess acidity in silage. Therefore, research is needed to define epidemiologically and mechanistically the risks to animal health and to the human food chain from silages contaminated with pathogenic bacteria and mycotoxins, and to understand more completely the relationships between the physical and chemical compositions of silage and metabolic disorders in animals. PMID- 11122512 TI - Analysis of the amino acid indospicine in biological samples by high performance liquid chromatography. AB - Indospicine is a hepatotoxic amino acid that accumulates in the meat of horses that consume the legume Indigofera linnaei. A method to determine indospicine concentration in biological samples using an amino acid analyser has been reported, but the analysis time is long and therefore not suited to the analysis of large numbers of samples. A rapid and reliable method was developed for the analysis of indospicine in horsemeat and serum using High Performance Liquid Chromatography. Horsemeat and serum were extracted with either water or 0.01 N hydrochloric acid, respectively, and deproteinized by ultrafiltration. Precolumn derivatization of samples with phenylisothiocyanate was followed by separation of indospicine from other amino acids on a Pico-Tag C 18 column and UV detection at 254 nm. The calibration curves for indospicine in horsemeat extract were linear over the concentration range 0.4 microg ml(-1) to 20 microg ml(-1), while for indospicine in serum, the linear range was from 0.17 microg ml(-1) to 16.67 microg ml(-1). The mean recovery of indospicine in horsemeat extract was 87.2 +/- 6.8% and in serum was 97.3 +/- 9.9%. Analysis time for indospicine in horsemeat samples was 31 min and in serum samples was 36 min. PMID- 11122513 TI - Fluorometric analysis of pectenotoxin-2 in microalgal samples by high performance liquid chromatography. AB - A rapid HPLC method with fluorescence detection of pectenotoxin-2 (PTX2), a polyether macrolide toxin, in microalgae is presented. A dienophile reagent, DMEQ TAD, was used for precolumn fluorescence labeling. PTX2 could be quantitatively detected in the range 1-200 ng. This method confirmed the occurrence of PTX2 in net haul samples mostly composed of dinoflagellates Dinophysisspp. collected in the Adriatic Sea, Italy and Mutsu Bay, Japan. PMID- 11122514 TI - A sensitive and specific determination method for azaspiracids by liquid chromatography mass spectrometry. AB - A liquid chromatography/mass spectrometry (LC/MS) method was developed for the sensitive and specific determination of azaspiracid and its two analogs, the causative toxins of azaspiracid poisoning that occurred in the Netherlands and Ireland. The LC/MS method provided a detection limit of 50 pg for azaspiracid. The sensitivity was approximately 8 x 10(4) times greater than the mouse bioassay. The method was used to confirm the presence of azaspiracids in toxic mussels collected at Arranmore Island, Ireland in 1997. PMID- 11122515 TI - Biosynthetic and genetic relationships of B-series fumonisins produced by Gibberella fujikuroi mating population A. AB - Fumonisins are mycotoxins produced by the maize pathogen Gibberella fujikuroi mating population A and frequently contaminate maize. Wild-type G. fujikuroi produces four B-series fumonisins, FB1, FB2, FR3 and FB4. These toxins are identical in structure except for the number and positions of hydroxyls along their linear carbon backbone. To elucidate the genetic and biosynthetic relationships among these fumonisins, we conducted meiotic and biochemical analyses of G. fujikuroi mutants with altered fumonisin production that resulted from defective alleles at three loci, Fum1, Fum2 and Fum3. These mutants produced either no fumonisins, only FR2 and FB4, or only FR3 and FR4. Genetic analyses revealed the orientation of the Fum loci along linkage group 1 of the fungus. The mutants were grown together in pair-wise combinations to determine if their fumonisin production phenotypes could be complemented. When FR3- and FB2 producing mutants were grown together, complementation occurred. However, when a nonproducing mutant was grown with a FR2- or FB3-producing mutant, complementation did not occur or was incomplete. When purified FR2, FR3, or FB4 was fed to mutant cultures, FR4 was converted primarily to FR2, FR3 was converted to FB1 and FB2 was not converted. The results from these assays suggest a previously unrecognized branch in the fumonisin biosynthetic pathway. PMID- 11122517 TI - Phytotoxicity of selected trichothecenes using Chlamydomonas reinhardtii as a model systemt. AB - Trichothecenes are potent inhibitors of cytoplasmic protein synthesis which can affect the severity of plant diseases such as wheat head scab. While many trichothecene-producing fungi share the initial biosynthetic intermediates, Fusarium sp. are unique in the production of trichothecenes containing an oxygen function at C-3. Although the initial trichothecene and the final products have a C-3 hydroxyl group, the intermediate steps are acetylated at C-3. By using Chlamydomonas reinhardtii, a unicellular plant with a well-defined genetic system, we were able to test the proposal that trichothecenes with a C-3 hydroxyl are more toxic to plants, as well as demonstrate that C. reinhardtii is a promising plant trichothecene bioassay system. Seven pairs of trichothecenes with either a C-3 hydroxyl or C-3 acetyl group were assayed. Our results confirm that trichothecenes acetylated at C-3 were far less toxic to Chlamydomonas than those with a C-3 hydroxyl group. PMID- 11122516 TI - Reusability of immunoaffinity columns for determination of fumonisins in maize. AB - Eighteen maize samples were assayed for fumonisin B1 (FB1) and B2 content by immunoaffinity column coupled with high performance liquid chromatography (HPLC). The FumoniTest columns were used once for the isolation of fumonisins (single-use column method). In the second part of the assay the columns were regenerated. After elution with methanol, PBS solution was left on the column for one day at room temperature to regenerate the columns (regenerated column method). The efficiency of columns regenerated twice was tested by determining FB, recovery and the reproducibility of the determinations. The recovery rate of FB1 proved to be 82% by the single-use column method (RSD: 5.7%) and 82.6% (RSD: 5.6 %) by the regenerated column method; 500-8,000 ng FB1 loaded onto the columns did not affect column performances. Nearly identical values were obtained when the FB1 content of fumonisin-containing maize samples was determined by both methods. The results indicate that the FumoniTest columns can be regenerated by the method applied at least twice without decrease in column performance. The fumonisin affinity, capacity and specificity of the regenerated columns were not changed. Thus, columns regenerated in this way can be used for determining the fumonisin content of maize samples at least three times. PMID- 11122518 TI - Winter accumulation of paralytic shellfish toxins in digestive glands of mussels from Arcachon and Toulon (France) without detectable toxic plankton species revealed by interference in the mouse bioassay for lipophilic toxins. AB - Since January 1993, neurological symptoms and rapid deaths (5 to 10 min) were typically observed in the mouse bioassay of acetone extracts of digestive glands from Arcachon and Toulon (France) during the winter season. It was assumed initially that a new lipophilic toxin was present because tests using the AOAC mouse bioassay for paralytic shellfish toxins on acid extracts of whole shellfish meat were negative, no known lipophilic toxins were detected and no toxic phytoplankton species were observed in the area during the poisoning events. In this study, however, preparative isolation of the toxic factor from toxic mussel digestive glands has revealed the presence of paralytic shellfish toxins, the principal ones being gonyautoxins-2 and -3 at Arcachon and gonyautoxins-1, -4, -2 and -3 at Toulon. The toxin concentrations recorded were below levels harmful to consumers and therefore represent a false positive in the mouse bioassay for lipophilic toxins based upon acetone extraction. The origin of the toxins remains to be determined. PMID- 11122519 TI - Immunohistochemistry of fumonisin in poultry using avidin-biotin-peroxidase system. AB - Using monoclonal anti-fumonisin B1 antibody (anti-FB1) and avidin-biotin peroxidase system, liver and kidneys of broiler chicks were evaluated for the detection and distribution of fumonisins (FBs). One hundred and fifty micrograms of FB1 or culture extract of Fusarium moniliforme str. 113F containing 150 microg of FB1 and 4 microg of FB2 were administered into the vitelline sac of 1-day old, specific pathogen-free chicks. The animals were killed 24 h after injection, and renal and hepatic tissues submitted for immunohistochemical analysis. FBs were detected in the epithelial cells of convoluted distal and proximal tubules of the kidneys, as well as in the cytoplasm of hepatocytes. This novel immunohistochemical method developed is expected to be an efficient way for monitoring the target of the FB toxins in tissues. PMID- 11122520 TI - A review of the volatile metabolites of fungi found on wood substrates. AB - The holdings of eight collections of fungi have been examined for organisms isolated from wood and/or trees. Further selection of these fungi has been made according to their reported ability to produce volatile, biologically active metabolites. It is emphasized that the isolates in the collections do not necessarily produce such metabolites. The list of fungi fulfilling these conditions is slightly augmented by reports we have found in the literature, where the fungi concerned have not yet been deposited. The biochemistry of these compounds is considered with particular emphasis on their biosynthesis including that by Homo sapiens. The physiological and toxicological activity of these metabolites is reviewed especially with reference to their potential role in the complex symbioses existent in, for example, a tree. The review concludes with a discussion of areas of botany deserving increased attention in the hope that this will stimulate further work. The statements in the review are based on 173 references. PMID- 11122521 TI - The fungal metabolite culmorin and related compounds. AB - This paper reviews the toxicology of culmorins, a family of compounds found in grains contaminated by Fusarium graminearum and related fungi. We include the results of an Ames test and studies based on Quantitative Structure-Activity Relationships. Culmorin has low toxicity in several in vitro assays and in one study in swine and is Ames test negative. Culmorin is moderately antifungal. QSAR analysis suggested that the plant compound longifolene was similar. Longifolene is a GRAS compound used in cosmetics and is also moderately antifungal. PMID- 11122522 TI - Evidence of saxitoxin derivatives as causative agents in the 1997 mass mortality of monk seals in the Cape Blanc Peninsula. AB - Monk seals in Cape Blanc (Western Sahara coast) suffered a mass mortality during May-July 1997 which was attributed to a morbillivirus. High performance liquid chromatography (HPLC) analysis on tissues of seals killed during the outbreak and on related fauna showed peaks with retention times coincident with those of some saxitoxin derivatives but their identity was not proved. Here we present results of further HPLC analyses that unambiguously prove the identity of these toxins by mass spectrometry (MS), supporting the hypothesis that this mortality of monk seals was caused by biotoxins rather than by a morbillivirus. PMID- 11122523 TI - The possible involvement of 3-methoxy-2(5H)-furanone in the etiology of Narthecium asiaticum maxim. associated nephrotoxicity in cattle. AB - Two samples of Narthecium asiaticum Maxim leaves collected in Japan were found to contain 103 microg(-1) and 160 microg g(-1) dry matter of 3-methoxy-2(5H) furanone respectively. 3-Methoxy-2(5h)-furanone was suggested to be the toxic principle of N. asiaticum causing nephrotoxicity in cattle in Japan. Two other furanones, which are thought to be non-toxic, were also isolated from the two samples. These were 4-methoxy-2(5H)-furanone and 5-hydroxy-4-methoxy-2(5H) furanone. PMID- 11122524 TI - The lichen rock tripe (Lasallia pustulata) as survival food: effects on growth, metabolism and immune function in Balb/c mice. AB - The present study was performed to investigate whether the lichen rock tripe (Lasallia pustulata) can be used as food during survival situations. The effects of 30% lichen supplementation given to female Balb/c mice were studied on growth rate, metabolism and immune functions. After 3 weeks on this diet, it was found that the lichen supplementation did not affect the growth rate or the well-being of the animals. The growth rate tended to be higher in the lichen group when compared to control mice. Food consumption was similar in both groups, but with a trend towards slightly higher intake (12%) in the lichen group. The heart, liver, kidney and lymphoid organ (spleen and thymus) weights were not affected by the lichen. Histological hematoxylin eosin staining showed that all these organs were normal. Plasma glucose levels were unchanged, but plasma urea levels decreased by 24% (p < 0.05) with the lichen diet. Red and white blood cells and the number of lymphoid cells in the thymus and spleen were normal. The activity of thymocytes and spleen T-lymphocytes were not affected by the lichen diet, but spontaneous cell-mediated cytotoxicity (NK cells) tended (n.s.) to increase and spleen B lymphocyte activity increased by 40% (p < 0.05). This study shows that the lichen rock tripe has immune stimulating effects important for host defence reactions and can be used as food in survival situations without any adverse effects on the metabolism. PMID- 11122525 TI - Aflatoxin B1 and fumosin B1 in mixed cultures of Aspergillus flavus and Fusarium proliferatum on maize. AB - Production of aflatoxin B1 and fumonisin B1 in pure and mixed cultures of Aspergillus flavus and Fusarium proliferatum were determined on irradiated maize seeds inoculated with different spore concentrations at 0.97 water activity (a(w)) and a temperature of 25 degrees C. The highest levels of aflatoxin B1 were produced by A. flavus at the lowest levels of inoculum (10(3) spore ml(-1)). There was no spore concentration influence on fumonisin B1 production after 10, 20 and 35 days of incubation. When A. flavus was co-inoculated with F. proliferatum, aflatoxin B1 production was inhibited. The higher the inocula levels of Fusarium produced, the higher the inhibition and this inhibition increased during the incubation period. Total inhibition was reached at 35 days of incubation. There was no interaction influence on fumonisin B1 production at all inoculum levels assayed. These results suggest that under optimal environmental conditions of substrate, water activity and temperature, the interaction between A. flavus and F proliferatum could produce inhibition of aflatoxin B1 and stimulation of fumonisin B1. PMID- 11122527 TI - Advances in detection methods for fungal and algal toxins. PMID- 11122526 TI - Relative inhibition of insect phenoloxidase by cyclic fungal metabolites from insect and plant pathogens. AB - The fungal metabolite kojic acid, which is produced by Aspergillus and Penicillium species fungi that may be pathogens of both insects and plants, was a significant inhibitor of phenoloxidase of different representative beetle and caterpillar insect species. Fusaric acid and picolinic acid, produced by Fusarium spp., were also significant inhibitors of phenoloxidase, while dipicolinic acid and beauvericin were ineffective at concentrations tested. Previous reports of the ability of kojic and fusaric acid to inhibit defensive enzymes of plants suggest that these compounds may be important in allowing the producing fungi to be pathogens of both insects and plants. PMID- 11122528 TI - Overview of mycotoxin methods, present status and future needs. AB - This article reviews current requirements for the analysis for mycotoxins in foods and identifies legislative as well as other factors that are driving development and validation of new methods. New regulatory limits for mycotoxins and analytical quality assurance requirements for laboratories to only use validated methods are seen as major factors driving developments. Three major classes of methods are identified which serve different purposes and can be categorized as screening, official and research. In each case the present status and future needs are assessed. In addition to an overview of trends in analytical methods, some other areas of analytical quality assurance such as participation in proficiency testing and reference materials are identified. PMID- 11122529 TI - High performance liquid chromatography coupled with post-column electrochemical oxidation for the detection of PSP toxins. AB - High Performance Liquid Chromatography (HPLC) is an important tool for the study of PSP toxins. It provides an alternative to bioassays and gives the concentration of individual toxin isomers. The current HPLC protocol uses a post column chemical reaction system (PCRS) to oxidize the saxitoxin ring system to form a fluorescent chromophore. This oxidation is sensitive to changes in the flow rate, temperature, pH and age of the reagents. We have previously shown that this oxidation can be accomplished using electrochemical techniques. Termed the electrochemical oxidation system (ECOS), this approach provides a simpler alternative to the traditional PCRS-based HPLC system. A detailed description of the construction and maintenance of an HPLC-ECOS system for the analysis of PSP toxins is presented. Comparisons of the mouse bioassay, HPLC-PCRS and HPLC-ECOS system are presented for three different sample matrices: toxic dinoflagellates (Alexandrium tamarense), geoduck (Panopea generosa) and scallops (Placopectin magellanicus). In all three cases, the correlation of the HPLC-ECOS system to the mouse bioassay is similar to that obtained using the HPLC-PCRS system for the analysis of PSP toxins. PMID- 11122530 TI - Detection of diarrhetic shellfish poisoning toxins from tropical shellfish using liquid chromatography-selected reaction monitoring mass spectrometry. AB - A negative mode liquid chromatography-selected reaction monitoring mass spectrometry (LC-SRM MS) method was developed to detect low concentrations of the diarrhetic shellfish poisoning (DSP) toxins okadaic acid and dinophysistoxin-1 (DTX-1). Detection relies upon monitoring the transition of negative precursor ions [M - H]- to a common fragment ion of m/z 255. Our limit of detection for okadaic acid with this method is 0.5 pg on column. LC-SRM MS has allowed us to detect persistent, low concentrations of DSP toxins from Singapore shellfish. PMID- 11122531 TI - HPLC/MS analysis of fusarium mycotoxins, fumonisins and deoxynivalenol. AB - Fusarium fungi are widely found in agricultural products, worldwide and can produce a great variety of mycotoxins. Fumonisins, produced by F. moniliforme, and deoxynivalenol, produced by F. graminearum, are two such mycotoxins that have received considerable attention as food safety concerns by regulatory agencies. High Performance Liquid Chromatography/Mass Spectrometry (HPLC/MS) was found to be a convenient analytical method to detect and quantify the naturally occurring fumonisin homologs and deoxynivalenol in extracts from grains and food products. The fumonisins are detected primarily as protonated molecules in the positive ion electrospray ionization (ESI) mode as they elute from a C-18 reverse phase column during a methanol water gradient containing acetic acid to facilitate chromatography. Deoxynivalenol can be detected as positive or negative ions in the atmospheric pressure chemical ionization (APCI) mode or in the negative ion ESI mode. One nanogram amounts of fumonisins or deoxynivalenol injected into the HPLC system are easily detected with signal to noise allowing detection limits of 1 microg g(-1) or better to easily be achieved with minimal clean-up of grain extracts. PMID- 11122532 TI - Fiber-optic immunosensor for mycotoxins. AB - Evanescent wave-based fiber-optic immunosensors were studied for the detection of fumonisins and aflatoxins in maize. Two formats, competitive and non-competitive, were used. A competitive format was used to measure fumonisin B1 (FB1) in both spiked and naturally contaminated maize samples. Fumonisin monoclonal antibodies were covalently coupled to an optical fiber and the competition between FB1 and FB1 labeled with fluorescein (FB1-FITC) for the limited number of binding sites on the fiber was assessed. The signal generated in the assay was inversely proportional to the FB1 concentration. For samples, the concentration causing an inhibition of binding by 50% (IC50) was dependent upon the clean-up procedure used. Simple dilution of methanolic maize extracts yielded an assay with an IC50 equivalent to 25 microg FB1 g(-1) maize with a limit of detection of 3.2 microg g(-1) maize. Affinity column clean-up yielded an assay with an IC50 equivalent to 5 microg FB1 g(-1) maize (limit of detection 0.4 microg FB1 g(-1)). An HPLC method and the immunosensor method agreed well for naturally contaminated maize samples except when large amounts of other fumonisins that cross-react with the immunosensor were present. The second sensor format, for the mycotoxin aflatoxin B1 (AFB1), was a non-competitive assay using the native fluorescence of this mycotoxin. Because the fluorescence of AFB1 itself was detected, the response of the sensor was directly proportional to the toxin concentration. The sensor, while capable of detecting as little as 2 ng ml(-1) of AFB1 in solution was technically not an immunosensor, since the attachment of aflatoxin specific antibodies was not required. Sensors of the formats described have the potential to rapidly screen individual maize samples but require coupling with a clean-up technique to be truly effective. PMID- 11122533 TI - Using an enzyme linked immunosorbent assay (ELISA) and a protein phosphatase inhibition assay (PPIA) for the detection of microcystins and nodularins. AB - Cyanotoxins produced by cyanobacteria (blue-green algae) include potent neurotoxins and hepatotoxins. The hepatotoxins include cyclic peptide microcystins and nodularins plus the alkaloid cylindrospermopsins. Among the cyanotoxins the microcystins have proven to be the most widespread, and are most often implicated in animal and human poisonings. This paper presents a practical guide to two widely used methods for detecting and quantifying microcystins and nodularins in environmental samples-the enzyme linked immunosorbant assay (ELISA) and the protein phosphatase inhibition assay (PPIA). PMID- 11122534 TI - Methodological improvement of the protein phosphatase inhibition assay for the detection of okadaic acid in mussels. AB - A simplified procedure for the enzyme inhibition assay to measure okadaic acid and DTX-1 in mussels, based on the use of a commercially available enzyme preparation, is presented. The detection limit is 10 ng of toxin per g of digestive glands. Using Certified Reference Material (MUS-2), high accuracy and good precision is demonstrated for contamination levels higher than 32 ng g(-1). Twenty samples can be processed in about 9 h by one operator, at the cost of US$ 10 per sample. Some possibilities for further enhancing the sensitivity and reducing the processing time are discussed and a monitoring example is presented. PMID- 11122535 TI - A receptor binding assay for paralytic shellfish poisoning toxins: recent advances and applications. AB - We recently described a high throughput receptor binding assay for paralytic shellfish poisoning (PSP) toxins, the use of the assay for detecting toxic activity in shellfish and algal extracts, and the validation of 11-[3H] tetrodotoxin as an alternative radioligand to the [3H]-saxitoxin conventionally employed in the assay. Here, we report a dramatic increase in assay efficiency through application of microplate scintillation technology, resulting in an assay turn around time of 4 h. Efforts are now focused on demonstrating the range of applications for which this receptor assay can provide data comparable to the more time consuming, technically demanding HPLC analysis of PSP toxins, currently the method of choice for researchers. To date, we have compared the results of both methods for a variety of sample types, including different genera of PSP toxin producing dinoflagellates (e.g. Alexandrium lusitanicum, r2 = 0.9834, n = 12), size-fractioned field samples of Alexandrium spp. (20-64 microm; r2 = 0.9997, n = 10) as well as its associated zooplankton grazer community (200-500 microm: r2 = 0.6169, n = 10; >500 microm: r2 = 0.5063, n = 10), and contaminated human fluids (r2 = 0.9661, n = 7) from a PSP outbreak. Receptor-based STX equivalent values for all but the zooplankton samples were highly correlated and exhibited close quantitative agreement with those produced by HPLC. While the PSP receptor binding assay does not provide information on toxin composition obtainable by HPLC, it does represent a robust and reliable means of rapidly assessing PSP-like toxicity in laboratory and field samples. Moreover, this assay should be effective as a screening tool for use by public health officials in responding to suspected cases of PSP intoxication. PMID- 11122536 TI - A yeast bioassay for trichothecenes. AB - Like all eucaryotic cells, yeasts are sensitive to trichothecenes, especially T-2 toxin and verrucarin A. Based on this sensitivity, a yeast bioassay was developed to evaluate the toxicity of corn samples. The bioassay was optimized using spiked maize extracts. The toxicity of samples was defined as toxicity equivalent to a certain concentration of T-2 toxin standards. The assay can be performed on crude extracts, but the results are more precise after column clean-up. The test can also be used for the screening of trichothecene toxicity in general. The relative standard deviation (RSD) at 85 % growth inhibition (EC85) was 4.5% for the T-2 toxin standards (n = 8). This corresponds to an initial T-2 toxin concentration of approximately 58 ppb in the corn sample. Samples containing 188 and 113 ppb T 2 toxin caused a growth inhibition higher than 85%, whereas samples with toxin concentrations of 56 and 19 ppb had a growth inhibition less than 85%. Therefore the test can be used for the qualitative evaluation of corn samples up to a level of 58 ppb +/- 2.8 ppb. The bioassay is easy to perform with minimum requirements for equipment. Results can be obtained within 24 h and a large number of samples can be analysed daily. The costs are low and the results obtained are repeatable. With some modifications this test can be used for toxicity studies on trichothecene metabolites as well as for extracts with unknown compounds with properties similar to trichothecenes. PMID- 11122537 TI - Alteration in sphingolipid metabolism: bioassays for fumonisin- and ISP-I-like activity in tissues, cells and other matrices. AB - The first discovered naturally occurring inhibitor of de novo sphingolipid biosynthesis was fumonisin B1. There are now 11 identified fungal inhibitors of ceramide synthase or 'fumonisin B1-like' compounds. With the exception of the australifungins, all other fungal ceramide synthase inhibitors are structurally sphingoid-like. There are several recently discovered fungal inhibitors of another enzyme in the de novo sphingolipid biosynthesis pathway: serine palmitoyltransferase (SPT). One of the SPT inhibitors is named ISP-I. While ceramide synthase inhibitors are toxic to animals, plants and fungi, the SPT inhibitors are not known to cause animal or plant disease, but are potent inhibitors of fungal growth. Very little is known about their toxicity in animals. There are at least 24 fungal SPT inhibitors produced by a variety of fungi. Given that the fungal inhibitors of sphingolipid biosynthesis are chemically and biologically diverse, two bioassays have been developed to screen for fumonisin-like or ISP-I-like activity in naturally contaminated products or fungal culture materials. These bioassays are based on the changes in free sphingoid base concentration that occur when the ceramide synthase or SPT are inhibited. The bioassays have the advantage that they are functionally rather than chemically specific and thus will detect ceramide synthase and SPT inhibitors regardless of their chemical structure. PMID- 11122538 TI - Reporter gene assays for algal-derived toxins. AB - We have modified the cell-based directed cytotoxicity assay for sodium channel and calcium channel active phycotoxins using a c-fos-luciferase reporter gene construct. In this report we describe the conceptual basis to the development of reporter gene assays for algal-derived toxins and summarize both published and unpublished data using this method. N2A mouse neuroblastoma cells, which express voltage-dependent sodium channels, were stably transfected with the reporter gene c-fos-luc, which contains the firefly luciferase gene under the transcriptional regulation of the human c-fos response element. The characteristics of the N2A reporter gene assay were determined by dose response with brevetoxin and ciguatoxin. Brevetoxin-1 and ciguatoxin-1 induced c-fos-luc with an EC50 of 4.6 and 3.0 ng ml(-1), respectively. Saxitoxin caused a concentration-dependent inhibition of brevetoxin-1 induction of c-fos-luc with an EC50 of 3.5 ng ml(-1). GH4C1 rat pituitary cells, which lack voltage-dependent sodium channels but express voltage-dependent calcium channels, were also stably transfected with the c-fos-luc. GH4C1 cells expressing c-fos-luciferase were responsive to maitotoxin (1 ng ml(-1)) and a putative toxin produced by Pfiesteria piscicida. Although reporter gene assays are not designed to replace existing detection methods used to measure toxin activity in seafood, they do provide a valuable means to screen algal cultures for toxin activity, to conduct assay-guided fractionation and to characterize pharmacologic properties of algal toxins. PMID- 11122539 TI - Small-molecule glycoprotein IIb/IIIa inhibitors as adjunctive therapy in percutaneous coronary interventions. PMID- 11122540 TI - Oral platelet IIb/IIIa inhibitors: from attractive theory to clinical failures. PMID- 11122541 TI - Left circumflex occlusion--underrecognized and undertreated. PMID- 11122542 TI - Association of endogenous tissue plasminogen activator (t-PA) with clinical characteristics of the insulin resistance syndrome. PMID- 11122543 TI - Argatroban and alteplase in patients with acute myocardial infarction: the ARGAMI Study. AB - ARGAMI was designed to assess safety and efficacy of argatroban compared with heparin as adjunctive treatment to alteplase in the treatment of patients with acute myocardial infarction. ARGAMI consisted of an open-dose finding study (35 patients) followed by a placebo-controlled study with double dummy technique and 2:1 (argatroban:heparin) randomization. An argatroban dosage of 100 microg/kg bolus plus 3 microg/kg/min infusion for 72 hours was selected for the randomized study in which 82 patients were allocated to argatroban and 45 to heparin (5000 U intravenous bolus, 1000 U/h infusion). Patency of the infarct-related artery (Thrombolysis in Myocardial Infarction [TIMI] grade 2 or 3 flow) after 90 minutes was obtained in 62 patients (76%) allocated to argatroban versus 37 patients (82%) allocated to heparin (p=ns). Angiograms after 24 hours and 5 to 10 days showed low reocclusion rates in both groups. Bleeding complications were observed in 16 patients allocated to argatroban (19.5%) and in 9 patients allocated to heparin (20.0%). One patient allocated to heparin suffered from hemorrhage stroke. Argatroban, given as adjunctive treatment to alteplase, is tolerated well in patients with acute myocardial infarction. Safety and efficacy of the combination alteplase and argatroban (with this dose regimen) are similar to those of alteplase and heparin. PMID- 11122544 TI - The TETAMI trial: the safety and efficacy of subcutaneous enoxaparin versus intravenous unfractionated heparin and of tirofiban versus placebo in the treatment of acute myocardial infarction for patients not thrombolyzed: methods and design. AB - Patients with acute myocardial infarction (AMI) who do not receive early reperfusion therapy are at high risk of reinfarction or death, and the efficacy and safety of antithrombotic therapy in this group of patients has not been evaluated. Enoxaparin is a low-molecular-weight heparin (LMWH) that has previously been shown to reduce the incidence of ischemic events in patients with unstable angina or non-Q-wave MI. The principal aims of the TETAMI study are to investigate the efficacy and safety of treatment with enoxaparin or tirofiban (a glycoprotein IIb/IIIa receptor antagonist) alone or in combination for 2 to 8 days in patients with AMI who are not eligible for early reperfusion therapy. In this 2 by 2 factorial design study approximately 900 patients will be randomly assigned, in a blinded manner, to one of four treatments: enoxaparin alone, enoxaparin plus tirofiban, unfractionated heparin (UFH), or UFH plus tirofiban, with appropriate matched placebos. The primary end point is the composite of death, recurrent AMI, and recurrent angina, analyzed at 30 days after AMI. The design and methods of the TETAMI study are described in this article. PMID- 11122545 TI - Impaired culprit vessel flow in acute coronary syndromes ineligible for thrombolysis. AB - The majority of patients with acute myocardial infarction and other acute coronary syndromes (ACS) are considered ineligible for thrombolysis and do not routinely receive reperfusion therapy. We hypothesized that predictors and outcomes of angiographically impaired culprit vessel flow can be identified and compared. This trial evaluated the outcomes following triage angiography in acute coronary syndromes ineligible for thrombolytic therapy. Eligible patients (n=201) with<24 hours of symptoms were randomized to early triage angiography and subsequent therapies based on the angiogram versus conventional medical therapy. This analysis was performed in 165 patients, from experimental and control arms, in whom angiography was performed on the index hospitalization with the outcome of interest being target vessel flow (Thrombolysis In Myocardial Infarction [TIMI] grades 0 to 2) on initial angiography. Patients with and without impaired culprit lesion flow were similar with respect to age, gender, diabetes, and prior coronary disease. A family history of premature coronary disease was more common in those with impaired flow, 50.0 versus 28.5% (p=0.02). Abnormal culprit vessel flow was found in 19.2% of patients who underwent angiography within 6 hours of symptom onset; however, after 24 hours this rate was reduced to 11.7%. Impaired culprit lesion flow can be expected in approximately 20% of patients presenting with ACS who are ineligible for reperfusion therapy by conventional guidelines and therefore represents an opportunity for early intervention within 6 hours of the onset of symptoms in these patients. PMID- 11122546 TI - Activation of blood platelets after percutaneous transluminal coronary angioplasty and coronary artery bypass graft surgery. AB - We have evaluated the activation of platelets in blood samples taken from patients with stable angina undergoing balloon angioplasty (percutaneous transluminal coronary angioplasty [PTCA]) (n=11) or coronary artery bypass grafting (CABG) under hypothermic (n=11) or normothermic conditions (n=11). We have found that surface expression of P-selectin on platelets in whole blood from PTCA patients upon thrombin treatment was significantly reduced, as compared with control platelets from healthy subjects. This effect was partially reversed when platelets washed from the same blood sample were used, but even then P-selectin expression was significantly lower in PTCA patients than it was in control subjects. There was a significant increase in basal expression of P-selectin in blood platelets taken from patients who underwent CABG under normothermic conditions (warm blood cardioplegia) as opposed to hypothermic patients (cold crystalloid cardioplegia). These platelets retain the ability to respond to agonists, although to a much lower extent than do those from healthy control donors. The surface exposure of P-selectin on resting and thrombin-treated platelets isolated from CABG surgery patients was not different from that of the control platelets. The adhesion to fibrinogen of resting and thrombin-treated platelets from patients who underwent balloon angioplasty as well as CABG surgery under normothermic and hypothermic conditions was significantly reduced when compared with the fibrinogen of the control platelets. These results suggest that the function of platelet fibrinogen receptor is impaired in patients with stable angina pectoris and that PTCA and CABG surgery activates platelets. PMID- 11122547 TI - Ticlopidine and aspirin fail to suppress the increased platelet aggregability that follows percutaneous coronary interventions. AB - Previous studies indicate that percutaneous transluminal coronary angioplasty (PTCA) is associated with platelet activation. It is not well-established whether enhanced platelet aggregability after PTCA is prevented by the association of ticlopidine with aspirin. The aim of this study was to evaluate whole blood platelet aggregability before and after elective PTCA in patients with chronic stable angina receiving ticlopidine and aspirin. We studied 16 patients referred for elective PTCA, treated for > or = 72 hours with oral aspirin and ticlopidine (group 1), and 10 patients referred for diagnostic coronary angiography, treated with oral aspirin alone (group 2). An intravenous bolus of heparin was administered at the start of PTCA. In both groups, platelet aggregability was assessed at baseline and 24 hours after the procedure, using the PFA 100(R) system. This method measures the time required for flowing whole blood to occlude a collagen and adenosine diphosphate (ADP)-coated ring, shorter times indicating greater aggregability. In both groups, platelet aggregability after the procedure was significantly increased compared with baseline: 104+/-30 seconds before versus 88+/-24 seconds at 24 hours in group 1 (p=0.03) and 84+/-16 seconds before versus 69+/-14 seconds at 24 hours in group 2 (p=0.004). Group 1 patients, compared with group 2, showed a trend toward reduced aggregability at baseline (p=0.06) and significantly lower aggregability 24 hours after the procedure (p=0.03). Ticlopidine and aspirin reduce whole-blood platelet aggregability compared with aspirin alone but fail to suppress the increased aggregability that occurs 24 hours after PTCA. PMID- 11122548 TI - The treatment and outcome of cancer patients with thromboses on central venous catheters. AB - Thromboses are a common complication of central venous catheters in cancer patients. This study was performed to analyze retrospectively the treatment and outcome of all patients with venous thromboses related to central venous catheters at a major cancer center. From 1992 through 1995, 319 oncology patients with central venous catheters underwent radionuclide venography (RNV) at the Dana Farber Cancer Institute for suspected catheter or venous thrombosis. The treatment and outcome of patients found to have venous thromboses were evaluated. Of the 319 patients, 112 were found to have evidence of venous thrombosis. The median age and platelet counts were not significantly different between the patients with and without thromboses. The most common indication for obtaining RNV was difficulty in aspirating or infusing material through the catheter. Patients who had pain or edema, or both, of the neck or upper extremity were more likely to have a venous thrombosis. Regardless of therapeutic intervention, including anticoagulation with heparin or coumadin, or both; line removal or replacement; or a combination thereof, no patients had a major adverse outcome such as pulmonary embolism, compromise of limb, or death. Only 4 patients did not have resolution of their presenting symptoms, all of whom were treated with line replacement. The overall survival of patients with and without thromboses was not significantly different. Either anticoagulation or removal of the central venous catheter (or both) appears to be adequate treatment for catheter-related thrombosis. A prospective trial to evaluate these approaches may be worthwhile so that the use of unnecessary procedures may be avoided in this patient population. PMID- 11122549 TI - The antithrombotic effects of CI-1028, an orally bioavailable direct thrombin inhibitor, in a canine model of venous and arterial thrombosis. AB - Direct thrombin inhibitors represent a new class of drug that may offer a therapeutic alternative for the treatment and prevention of thrombembolic conditions, especially on the venous side of the systemic circulation. CI-1028 (PD 172524/LB30057) is a potent, highly selective inhibitor of thrombin that is orally bioavailable. The efficacy of this compound has been demonstrated in animal models in which intra-venous administration was used. The objective of this study was to evaluate the efficacy of CI-1028 after oral administration in a canine electrolytic injury model of venous and arterial thrombosis. CI-1028 was administered via oral gavage, and animals received either saline or 10, 15, 20, or 30 mg/kg of drug. Fifteen minutes later, the dogs were anesthetized and a femoral artery and vein were exposed and instrumented to induce electrolytic injury and thrombosis while continuously monitoring blood flow in the vessels. Maximum blood CI-1028 concentrations of 0.88+/-0.27, 1.8+/-0.3, 2.2+/-0.5, and 3.2+/-0.5 microg/mL were generally achieved 15 to 30 minutes after administering the compound in the 10-, 15-, 20-, and 30-mg/kg groups, respectively. Administration of CI-1028 increased the time to occlusion (TTO), the principal efficacy end point, in a dose-dependent manner in both arteries and veins. The TTO in the control group (n=8) averaged 66+/-11 minutes in the arteries and 69+/ 6 minutes in the veins. In dogs treated with 10 mg/kg (n=8), the TTO was not significantly different from that of the control group. In the 15-mg/kg group (n=9) TTO averaged 140+/-27 minutes in the arteries (p=not significant) and 125+/ 15 minutes (p<0.05) in the veins. In the 20-mg/kg group (n=8), TTO was significantly longer than controls in both types of vessels, averaging 168+/-30 minutes in the arteries (p=0.05) and 155+/-21 minutes (p<0.05) in the veins. Likewise, at 30 mg/kg (n=8) both the arterial (179+/-17 minutes) and venous (188+/-15 minutes) TTO was significantly prolonged compared with controls. Surgical blood loss and template bleeding times tended to increase in a dose dependent manner but a statistically significant elevation was detected for template bleeding time only at the highest dose. Dramatic changes in thrombin time were detected, consistent with the CI-1028 mechanism of action. Virtually no changes were detected in prothrombin time. Maximum activated partial thromboplastin time (aPTT) and activated clotting time changes were detected approximately 30 minutes after dosing, and they were approximately twofold and fivefold baseline values, respectively, at the highest dose. In conclusion, these results demonstrate that CI-1028 provides dose-dependent antithrombotic efficacy after oral administration in a canine model of venous and arterial thrombosis. PMID- 11122550 TI - Intramyocardial injection of DNA encoding vascular endothelial growth factor in a myocardial infarction model. AB - In a previous study, we observed that one injection of 500 microg of DNA for the plasmid encoding for vascular endothelial growth factor (ph VEGF(165)) into one site in a rat myocardial infarction model resulted in neovascularization confined to angiomatous structures that did not contribute to regional myocardial blood flow. The purpose of the present study was to determine whether a lower dose (125 microg DNA), which is the same as that being used in some clinical trials, injected into four separate sites could enhance collateral flow and vascularity to the ischemic bed without inducing angiomas. Rats received injections of 125 microg DNA of the plasmid encoding phVEGF(165) or control DNA at four separate sites within the anterior free wall of the left ventricle (LV) supplied by the left coronary artery. The left coronary artery was ligated and hearts analyzed at 4 weeks. In vitro studies confirmed that the phVEGF(165) used was capable of producing VEGF polypeptide in mammalian cells. The infarct size (percentage of endocardial circumference that infarcted) was similar in controls (42+/-6%) and treated hearts (39+/-7%); the LV cavity area did not differ between groups. The number of vascular structures per high-power field within the infarct scar was 10.50+/-0.68 in controls and 10.00+/-0.85 in phVEGF(165)-treated rats. Relative regional myocardial blood flow determined by radioactive microspheres and expressed as a ratio of radioactive counts within the scar divided by radioactive counts in the noninfarcted ventricular septum was similar in control (0.74+/ 0.25) and treated hearts (0.88+/-0.30) (p=not significant). No angiomatous structures were observed. Injections of 125 microg of DNA of phVEGF(165) into myocardium to become ischemic had no effect on infarct size or LV cavity size. Unlike higher doses of 500 microg of DNA, it did not cause gross angiomatous structures; however, it failed to improve neovascularization or regional myocardial blood flow in this rodent model of acute myocardial infarction. PMID- 11122551 TI - Nutrition and ageing. PMID- 11122552 TI - Nutrition assessment in the elderly. AB - The prevalence of malnutrition, which is relatively low in free-living elderly persons (5-10%), is considerably higher (30-60%) in hospitalized or institutionalized elderly persons. As a result, nutritional assessment should be part of routine clinical practice in elderly patients who are frail, sick or hospitalized. A comprehensive screening tool for assessment of nutritional status is needed that is clinically relevant and cost-effective to perform. A number of simple and rapid tests for detecting or diagnosing malnutrition in the elderly have recently been developed. If malnutrition is suggested by such screening tests, then they should be supplemented by conventional nutritional assessment before treatment is planned. PMID- 11122553 TI - Anorexia, body composition, and ageing. AB - Over the lifespan there is a decline in food intake. This has been termed the physiological anorexia of aging. It has many causes, including alterations in the gastrointestinal satiating system, the effect of elevated leptin levels, especially in men, and a variety of changes in central nervous system neurotransmitters. Beyond the age of 70 years body mass declines. This includes both loss of adipose tissue and muscle mass. The loss of muscle mass in older individuals is termed sarcopenia. There is increasing evidence that this is caused, in men, partly by the decline in testosterone. Illness results in an increase of cytokines that produce both anorexia and cause protein wasting. Many of the causes of cachexia in older individuals are treatable. Depression is the most common cause of weight loss in older individuals. Dieting in older individuals is associated with a loss of skeletal tissue as well as fat mass. This can place older individuals at risk of becoming the 'fat frail'. PMID- 11122554 TI - The contribution of nutritional factors to osteopenia in the elderly. AB - Calcium and vitamin D deficiency increase age-related bone loss by causing secondary hyperparathyroidism. Reduced endogenous vitamin D synthesis exacerbates the problem of dietary deficiency and involves elderly people living in their own homes, who are just as much at risk as those living in institutionalized care. The effects of secondary hyperparathyroidism may be offset by hypercalcaemia of the increased bone turnover of immobility, which has a direct adverse effect on the skeleton causing osteoporosis. Active vitamin D analogues are effective in suppressing secondary hyperparathyroidism caused by vitamin D deficiency. However, simple deficiency is optimally treated with parent vitamin D, which has a greater safety margin than active vitamin D therapy (1,25 dihydroxyvitamin D), which requires close monitoring in the elderly. PMID- 11122555 TI - Oral supplements in the elderly. AB - Under-nutrition is common in elderly patients, especially for those in hospital. Hospital stay is often associated with further weight loss. Many authors have considered nutritional supplementation. A recent meta-analysis of the trials of nutritional supplementation in all age groups and pathologies found benefits from supplementation but suggested that further work was required. This review considers the work done since the meta-analysis until the end of 1999 and in particular considers the benefits evident to elderly patients using oral supplements. PMID- 11122556 TI - Herbal-drug therapy interactions: a focus on dementia. AB - Older people with dementia are often prescribed numerous medications. Use of herbal therapies in addition to these conventional drug therapies may lead to interactions that result in an adverse drug event. We have conducted a systematic review to identify all studies that examined interactions between herbal and conventional drug therapies (i.e. prescription or over-the-counter). Using a MEDLINE search of English-language studies published between 1980 and 2000, we limited our search to those herbal therapies most likely to be used for the treatment of dementia (memory loss and decreased concentration) and related symptoms. We identified 28 articles that describe interactions between these herbal (i.e. St. John's wort, ginkgo biloba, kava, valerian, and ginseng) and conventional drug therapies. Of these articles, 11 examined St. John's wort, four examined ginkgo biloba, five examined kava, one examined valerian, and seven examined ginseng. We identified a series of potential interactions between herbal and conventional drug therapy that place older people at risk for an adverse drug event. Health care professionals need to be aware of these potential interactions. PMID- 11122557 TI - Protein metabolism at the crossroads? PMID- 11122558 TI - Regulation of protein synthesis by branched-chain amino acids. AB - Historically, amino acids have been viewed as precursors for protein synthesis as well as metabolic substrates. Recently, a new role for amino acids as regulators of mRNA translation has been identified. In this role, they modulate the phosphorylation state of proteins that represent important control points in translation initiation, including the translational repressor 4E-BP1 and the ribosomal protein S6 kinase S6K1. When administered orally to fasted rats the branched-chain amino acids are particularly effective in stimulating translation initiation. Of the branched-chain amino acids, leucine is most potent. Interestingly, leucine administration stimulates global rates of protein synthesis in skeletal muscle but not in liver. However, in liver, branched-chain amino acids enhance the translation of a particular set of mRNAs typified by those encoding the ribosomal proteins and translation elongation factors, suggesting that branched-chain amino acids upregulate the capacity of the tissue to synthesize protein. PMID- 11122559 TI - Regulation of proteolysis. AB - The mechanisms of proteolysis remain to be fully defined. This review focuses on recent advances in our understanding of the ubiquitin-proteasome-dependent pathway, which is involved in the control of many major biological functions. The ubiquitinylation/deubiquitinylation system is a complex machinery responsible for the specific tagging and proof-reading of substrates degraded by the 26S proteasome, as well as having other functions. The formation of a polyubiquitin degradation signal is required for proteasome-dependent proteolysis. Several families of enzymes, which may comprise hundreds of members to achieve high selectivity, control this process. The substrates tagged by ubiquitin are then recognized by the 26S proteasome and degraded into peptides. In addition, the 26S proteasome also recognizes and degrades some non-ubiquitinylated proteins. In fact, there are multiple ubiquitin- or proteasome-dependent pathways. These systems presumably degrade specific classes of substrates and single proteins by alternative mechanisms and could be interconnected. They may also interfere or cooperate with other proteolytic pathways. PMID- 11122560 TI - Of flux and flooding: the advantages and problems of different isotopic methods for quantifying protein turnover in vivo : II. Methods based on the incorporation of a tracer. AB - The most common methods for measuring the incorporation of tracer amino acids into tissue protein are the constant tracer infusion and the flooding dose. The flooding dose is an attractive method for measuring tissue protein synthesis because of its convenience and precision. A primary assumption of the method, that the free amino acid precursor pools are equilibrated with the true precursor pool, aminoacyl-transfer RNA, has recently been validated. When short labelling periods are involved, the large dose of amino acid does not appear to alter protein synthesis. The constant tracer infusion is a satisfactory method from a theoretical point of view, but its use requires the measurement of the protein synthetic precursor pool. The best estimate of the aminoacyl-tRNA precursor pool for the constant infusion method appears to be the acid-soluble tissue pool in muscle and VLDL apolipoprotein B-100 in the liver. The experimental approach chosen for measuring tissue protein synthesis should be dictated by the question being addressed. PMID- 11122561 TI - Nucleotides as immunomodulators in clinical nutrition. AB - Dietary nucleotides, like glutamine, have attracted attention as a key ingredient missing from nutritional formulae for many years. They are the building blocks of tissue RNA and DNA and of ATP and their presence in breast milk has stimulated research in babies which has indicated that supplementation of infant formula milk leads to improved growth and reduced susceptibility to infection. Animal studies have confirmed some of these data. In particular, dietary nucleotides modulate immune function, promote faster intestinal healing and have trophic effects on the intestine of parenterally-fed rats which are similar to those resulting from glutamine supplementation, but at much lower intakes. Nucleotide supplementation has also been shown to improve some aspects of tissue recovery from ischaemia/reperfusion injury or radical resection. There is, however, a fundamental paradox. The intestine and liver possess powerful homeostatic mechanisms which degrade intake of purines and pyrimidines (i.e. salvage) and replace it with de novo synthesised output. It is possible that peripheral tissues receive only small amounts of nucleotides of dietary origin. Previously, nucleotides have been proposed as being conditionally-essential nutrients that provide an adequate supply of purines and pyrimidines for nucleic acid synthesis in neonates or in the stressed patient. This review explores this puzzle in the light of recent data from nutritional studies and from research into purinergic signalling in the intestine, heart and cells of the immune system. We propose that dietary nucleotides should be considered within a pharmacological and metabolic framework. PMID- 11122562 TI - Glutathione in disease. AB - Altered glutathione metabolism in association with increased oxidative stress has been implicated in the pathogenesis of many diseases. However, whether strategies aimed at restoring glutathione concentration and homeostasis are effective in ameliorating or modifying the natural history of these states is unknown. In this review we discuss the pathogenic role for altered glutathione metabolism in such diseases as protein energy malnutrition, seizures, Alzheimer's disease, Parkinson's disease, sickle cell anaemia, chronic diseases associated with ageing and the infected state. In addition, we discuss the efficacy of glutathione precursors in restoring glutathione homeostasis both in vitro and in vivo. PMID- 11122564 TI - JAMA 100 years ago: THE ENGLISH LANGUAGE AND MEDICINE PMID- 11122563 TI - A piece of my mind: lost by degrees. PMID- 11122565 TI - JAMA 100 years ago: THE METRIC SYSTEM PMID- 11122566 TI - Helsinki discord? A controversial declaration. PMID- 11122567 TI - Fast, aggressive treatment for myocardial infarction urged. PMID- 11122569 TI - Health agencies update: genes and HIV infection PMID- 11122571 TI - Health agencies update: leishmaniasis protection? PMID- 11122570 TI - Health agencies update: brain-powered prosthetics PMID- 11122572 TI - From the Health Care Financing Administration: Medicare + Choice plans. PMID- 11122573 TI - Treatment of mild depression in elderly patients. PMID- 11122574 TI - Treatment of mild depression in elderly patients. PMID- 11122575 TI - Treatment of mild depression in elderly patients. PMID- 11122576 TI - Treatment of mild depression in elderly patients PMID- 11122577 TI - Quality of care at teaching and nonteaching hospitals. PMID- 11122578 TI - Quality of care at teaching and nonteaching hospitals PMID- 11122579 TI - Health consequences of eclipse cigarettes. PMID- 11122580 TI - Health consequences of eclipse cigarettes PMID- 11122582 TI - Non-english reports of medical research PMID- 11122581 TI - Non-English reports of medical research. PMID- 11122583 TI - Gender vs sex. PMID- 11122584 TI - Gender vs Sex PMID- 11122585 TI - Acute myocardial infarction and prior antibiotic use. PMID- 11122586 TI - Handheld cellular telephone use and risk of brain cancer. AB - CONTEXT: A relative paucity of data exist on the possible health effects of using cellular telephones. OBJECTIVE: To test the hypothesis that using handheld cellular telephones is related to the risk of primary brain cancer. DESIGN AND SETTING: Case-control study conducted in 5 US academic medical centers between 1994 and 1998 using a structured questionnaire. PATIENTS: A total of 469 men and women aged 18 to 80 years with primary brain cancer and 422 matched controls without brain cancer. MAIN OUTCOME MEASURE: Risk of brain cancer compared by use of handheld cellular telephones, in hours per month and years of use. RESULTS: The median monthly hours of use were 2.5 for cases and 2.2 for controls. Compared with patients who never used handheld cellular telephones, the multivariate odds ratio (OR) associated with regular past or current use was 0.85 (95% confidence interval [CI], 0.6-1.2). The OR for infrequent users (<0. 72 h/mo) was 1.0 (95% CI, 0.5-2.0) and for frequent users (>10.1 h/mo) was 0.7 (95% CI, 0.3-1.4). The mean duration of use was 2.8 years for cases and 2.7 years for controls; no association with brain cancer was observed according to duration of use (P =.54). In cases, cerebral tumors occurred more frequently on the same side of the head where cellular telephones had been used (26 vs 15 cases; P =.06), but in the cases with temporal lobe cancer a greater proportion of tumors occurred in the contralateral than ipsilateral side (9 vs 5 cases; P =.33). The OR was less than 1.0 for all histologic categories of brain cancer except for uncommon neuroepitheliomatous cancers (OR, 2.1; 95% CI, 0.9-4.7). CONCLUSIONS: Our data suggest that use of handheld cellular telephones is not associated with risk of brain cancer, but further studies are needed to account for longer induction periods, especially for slow-growing tumors with neuronal features. PMID- 11122587 TI - Marital stress worsens prognosis in women with coronary heart disease: The Stockholm Female Coronary Risk Study. AB - CONTEXT: Psychosocial stress has been associated with incidence of coronary heart disease (CHD) in men, but the prognostic impact of such stress rarely has been studied in women. OBJECTIVE: To investigate the prognostic impact of psychosocial work stress and marital stress among women with CHD. DESIGN AND SETTING: Population-based, prospective follow-up study conducted in the city of Stockholm, Sweden. PARTICIPANTS: A total of 292 consecutive female patients aged 30 to 65 years (n = 279 working or cohabiting with a male partner) who were hospitalized for acute myocardial infarction or unstable angina pectoris between February 1991 and February 1994. Patients were followed up from the date of clinical examination until August 1997 (median, 4.8 years). MAIN OUTCOME MEASURES: Recurrent coronary events, including cardiac death, acute myocardial infarction, and revascularization procedures, by marital stress (assessed using the Stockholm Marital Stress Scale, a structured interview) and by work stress (assessed using the ratio of work demand to work control). RESULTS: Among women who were married or cohabiting with a male partner (n = 187), marital stress was associated with a 2.9-fold (95% confidence interval [CI], 1.3-6. 5) increased risk of recurrent events after adjustment for age, estrogen status, education level, smoking, diagnosis at index event, diabetes mellitus, systolic blood pressure, smoking, triglyceride level, high-density lipoprotein cholesterol level, and left ventricular dysfunction. Among working women (n = 200), work stress did not significantly predict recurrent coronary events (hazard ratio, 1.6; 95% CI, 0.8 3.3). CONCLUSIONS: Our results indicate that marital stress but not work stress predicts poor prognosis in women aged 30 to 65 years with CHD. These findings differ from previous findings in men and suggest that specific preventive measures be tailored to the needs of women with CHD. PMID- 11122588 TI - Longitudinal study of moderate weight change and sleep-disordered breathing. AB - CONTEXT: Excess body weight is positively associated with sleep-disordered breathing (SDB), a prevalent condition in the US general population. No large study has been conducted of the longitudinal association between SDB and change in weight. OBJECTIVE: To measure the independent longitudinal association between weight change and change in SDB severity. DESIGN: Population-based, prospective cohort study conducted from July 1989 to January 2000. SETTING AND PARTICIPANTS: Six hundred ninety randomly selected employed Wisconsin residents (mean age at baseline, 46 years; 56% male) who were evaluated twice at 4-year intervals for SDB. MAIN OUTCOME MEASURES: Percentage change in the apnea-hypopnea index (AHI; apnea events + hypopnea events per hour of sleep) and odds of developing moderate to-severe SDB (defined by an AHI > or =15 events per hour of sleep), with respect to change in weight. RESULTS: Relative to stable weight, a 10% weight gain predicted an approximate 32% (95% confidence interval [CI], 20%-45%) increase in the AHI. A 10% weight loss predicted a 26% (95% CI, 18%-34%) decrease in the AHI. A 10% increase in weight predicted a 6-fold (95% CI, 2.2-17.0) increase in the odds of developing moderate-to-severe SDB. CONCLUSIONS: Our data indicate that clinical and public health programs that result in even modest weight control are likely to be effective in managing SDB and reducing new occurrence of SDB. PMID- 11122589 TI - Gender disparities in the receipt of home care for elderly people with disability in the United States. AB - CONTEXT: Projected demographic shifts in the US population over the next 50 years will cause families, health care practitioners, and policymakers to confront a marked increase in the number of people with disabilities living in the community. Concerns about the adequacy of community support are particularly salient to women, who make up a disproportionate number of disabled elderly people and who may be particularly vulnerable because they are more likely to live alone with limited financial resources. OBJECTIVE: To address gender differences in receipt of informal and formal home care. DESIGN, SETTING, AND PARTICIPANTS: Nationally representative survey conducted in 1993 among 7443 noninstitutionalized people (4538 women and 2905 men) aged 70 years or older. MAIN OUTCOME MEASURE: Number of hours per week of informal (generally unpaid) and formal (generally paid) home care received by survey participants who reported any activity of daily living (ADL) or instrumental activity of daily living (IADL) impairment (n = 3109) compared by gender and living arrangement and controlling for other factors. RESULTS: Compared with disabled men, disabled women were much more likely to be living alone (45.4% vs 16.8%, P<.001) and much less likely to be living with a spouse (27.8% vs 73.6%, P<.001). Overall, women received fewer hours of informal care per week than men (15.7 hours; 95% confidence interval [CI], 14.5-16.9 vs 21.2 hours; 95% CI, 19. 7-22.8). Married disabled women received many fewer hours per week of informal home care than married disabled men (14.8 hours; 95% CI, 13.7-15.8 vs 26.2 hours; 95% CI, 24.6 27.9). Children (>80% women) were the dominant caregivers for disabled women while wives were the dominant caregivers of disabled men. Gender differences in formal home care were small (2.8 hours for women; 95% CI, 2.5-3.1 vs 2.1 hours for men; 95% CI, 1.7-2.4). CONCLUSION: Large gender disparities appear to exist in the receipt of informal home care for disabled elderly people in the United States, even within married households. Programs providing home care support for disabled elderly people need to consider these large gender disparities and the burden they impose on families when developing intervention strategies in the community. PMID- 11122590 TI - Influence of hospital procedure volume on outcomes following surgery for colon cancer. AB - CONTEXT: Survival following high-risk cancer surgery, such as pancreatectomy and esophagectomy, is superior at hospitals where high volumes of these procedures are performed. Conflicting evidence exists as to whether the association between hospital experience and favorable health outcomes also applies to more frequently performed operations, such as those for colon cancer. OBJECTIVE: To determine whether hospital procedure volume predicts survival following colon cancer surgery. DESIGN, SETTING, AND PARTICIPANTS: Retrospective cohort study of data from the Surveillance, Epidemiology and End Results-Medicare linked database on 27 986 colon cancer patients aged 65 years and older who had surgical resection for primary adenocarcinoma diagnosed between 1991 and 1996. MAIN OUTCOME MEASURES: Thirty-day postoperative mortality and overall and cancer-specific long term survival, by hospital procedure volume. RESULTS: We found small differences in 30-day postoperative mortality for patients treated at low- vs high-volume hospitals (3. 5% at hospitals in the top-volume quartile vs 5.5% at hospitals in the bottom-volume quartile). However, the correlation was statistically significant and persisted after adjusting for age at diagnosis, sex, race, cancer stage, comorbid illness, socioeconomic status, and acuity of hospitalization (P<.001). The association was evident for subgroups with stage I, II, and III disease. Hospital volume directly correlated with survival beyond 30 days and also was not attributable to differences in case mix (P<.001). The association between hospital volume and long-term survival was concentrated among patients with stage II and III disease (P<.001 for both). Among stage III patients, variation in use of adjuvant chemotherapy did not explain this finding. CONCLUSION: Our data suggest that hospital procedure volume predicts clinical outcomes following surgery for colon cancer, although the absolute magnitudes of these differences are modest in comparison with the variation observed for higher risk cancer surgeries. PMID- 11122591 TI - Contraindicated use of cisapride: impact of food and drug administration regulatory action. AB - CONTEXT: Cisapride, a gastrointestinal tract promotility agent, can cause life threatening cardiac arrhythmias in patients susceptible either because of concurrent use of medications that interfere with cisapride metabolism or prolong the QT interval or because of the presence of other diseases that predispose to such arrhythmias. In June 1998, the US Food and Drug Administration (FDA) determined that use of cisapride was contraindicated in such patients and informed practitioners through additions to the boxed warning in the label and a "Dear Health Care Professional" letter sent by the drug's manufacturer. OBJECTIVE: To evaluate the impact of the FDA's 1998 regulatory action regarding contraindicated use of cisapride. DESIGN AND SETTING: Analysis of data for the 1 year periods before (July 1997-June 1998) and after (July 1998-June 1999) the regulatory action from the population-based, pharmacoepidemiology research databases of 2 managed care organizations (sites A and B) and a state Medicaid program (site C). PARTICIPANTS: Patients with at least 180 days of prior enrollment in 1 of the 3 sites who were prescribed cisapride at least once in the period before (n = 24 840) or after (n = 22 459) regulatory action. Patients could be included in both cohorts. MAIN OUTCOME MEASURES: Proportion of cisapride users in each period for whom cisapride use was contraindicated by the product label, based on computerized patient medical encounter records. RESULTS: In the year prior to regulatory action, cisapride use was contraindicated for 26%, 30%, and 60% of users in study sites A, B, and C, respectively. In the year after regulatory action, use was contraindicated for 24%, 28%, and 58% of users, a reduction in contraindicated use of approximately 2 per 100 cisapride users at each site. When the analysis was restricted to new users of cisapride after regulatory action, only minor reductions in contraindicated use were found. CONCLUSION: The FDA's 1998 regulatory action regarding cisapride use had no material effect on contraindicated cisapride use. More effective ways to communicate new information about drug safety are needed. PMID- 11122592 TI - Hepatitis B vaccination and hepatocellular carcinoma rates in boys and girls. AB - CONTEXT: Hepatocellular carcinoma (HCC) has a male predominance and is closely related to hepatitis B virus (HBV) infection. Hepatitis B virus vaccination was launched in 1984 in Taiwan for neonates of mothers carrying hepatitis B e antigen, resulting in a decreased incidence of HCC in children. The effect on boys vs girls is not known. OBJECTIVE: To evaluate the association between a HBV vaccination program with incidence of childhood HCC by sex. DESIGN AND SETTING: Analysis of data collected from Taiwan's National Cancer Registry System and the Taiwan Childhood Hepatoma Study Group between 1981 and 1996. PARTICIPANTS: Children aged 6 to 14 years who were diagnosed as having HCC (201 boys and 70 girls). MAIN OUTCOME MEASURE: Incidence of HCC in boys and girls before and after implementation of the vaccination program. RESULTS: The boy-girl incidence ratio decreased steadily from 4.5 in 1981-1984 (before the program's introduction) to 1.9 in 1990-1996 (6-12 years after the vaccination program was launched). The incidence of HCC in boys born after 1984 was significantly reduced in comparison with those born before 1978 (relative risk [RR], 0.72; P =.002). No significant decrease in HCC incidence was observed in girls born in the same periods (RR, 0.77; P =.20). The incidence of HCC in boys remained stable with increasing age, while an increase of HCC incidence with age in girls was observed. These age and sex effects remained the same regardless of birth before or after the vaccination program. CONCLUSION: Our results suggest that boys may benefit more from HBV vaccination than girls in the prevention of HCC. PMID- 11122593 TI - World Medical Association Declaration of Helsinki: ethical principles for medical research involving human subjects. PMID- 11122594 TI - Perspectives on the Fifth Revision of the Declaration of Helsinki. PMID- 11122595 TI - Drug labeling revisions-guaranteed to fail? PMID- 11122596 TI - The patient-physician relationship. Ensuring competency in end-of-life care: communication and relational skills. AB - Physician competence in end-of-life care requires skill in communication, decision making, and building relationships, yet these skills were not taught to the majority of physicians during their training. This article presents a 7-step approach for physicians for structuring communication regarding care at the end of life. Physicians should prepare for discussions by confirming medical facts and establishing an appropriate environment; establish what the patient (and family) knows by using open-ended questions; determine how information is to be handled at the beginning of the patient-physician relationship; deliver the information in a sensitive but straightforward manner; respond to emotions of the patients, parents, and families; establish goals for care and treatment priorities when possible; and establish an overall plan. These 7 steps can be used in situations such as breaking bad news, setting treatment goals, advance care planning, withholding or withdrawing therapy, making decisions in sudden life-threatening illness, resolving conflict around medical futility, responding to a request for physician-assisted suicide, and guiding patients and families through the last hours of living and early stages after death. Effective application as part of core end-of-life care competencies is likely to improve patients' and families' experiences of care. It may also enhance physicians' professional fulfillment from satisfactory relationships with their patients and patients' families. PMID- 11122684 TI - Therapeutic impact of statin therapy in patients with low HDL cholesterol. PMID- 11122685 TI - Reducing atherothrombotic events in high-risk patients: recent data on therapy with statins and fatty acids. AB - Traditional treatment of atherosclerotic coronary heart disease by cardiovascular specialists, which has focused on "critical" stenoses, may be less effective in reducing morbidity and mortality than therapies that stabilize plaques and reduce thrombosis and sudden death. Recent data from clinical trials of 3-hydroxy-3 methylglutaryl coenzyme A reductase inhibitor (statin) therapy and modification of dietary fat composition demonstrate that both these approaches can reduce clinical events. Although revascularization therapy is effective in reducing angina caused by high-grade stenotic lesions, this therapy is incomplete because the more-numerous "smaller" plaques that typically cause clinical events remain untreated. Two recent trials suggest that statin therapy may have benefits on stabilizing plaques in high-risk patients within a year. Additional benefit may also be provided by increasing dietary consumption of monounsaturated or omega-3 polyunsaturated fatty acids. Both statin therapy and diets high in monounsaturated or omega-3 fatty acids appear to improve morbidity and mortality by modifying the underlying atherothrombotic disease process. PMID- 11122686 TI - Prevention of cardiovascular events in elderly hypercholesterolemic patients. AB - The management of hyperlipidemia in the elderly patient is a major problem, given the frequency of dyslipidemias and cardiovascular disorders in this age group. Therapy must take current uncertainties into account and, in the absence of therapeutic studies carried out in the elderly, is typically based upon a case-by case approach. Raised cholesterol levels remain a significant risk factor for coronary heart disease (CHD) in the elderly. Although the relative risk of CHD tends to diminish with increasing age, this reduction is accompanied by an increase in absolute risk (ie, the number of events) as the frequency of the illness increases markedly with age. The results of major outcome studies with 3 hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins), when analyzed according to patient age, indicate that the benefits of these agents are not merely confined to younger individuals. However, the elderly form a unique patient population--the proportion of women is greater and the profile of cardiovascular illnesses is characterized, among others, by a greater incidence of cerebrovascular accidents. Problems relating to poor tolerability and comorbidity (which may give rise to drug-drug interactions) also occur more frequently in this age group. Moreover, the potential widespread treatment of hyperlipidemia in the elderly has profound economic implications. Under these circumstances, the clinical practice recommendations depend upon a reasonable extrapolation of epidemiologic and therapeutic data obtained from middle-aged men. At present, treatment is therefore aimed at patients with the most severe forms of hyperlipidemia, generally in the secondary prevention setting, taking into account the patient's life expectancy. The results of ongoing studies will determine the benefits of lipid-lowering therapy for primary prevention of CHD in the elderly. PMID- 11122687 TI - Treatment of dyslipidemia: genetic interactions with diet and drug therapy. AB - Coronary heart disease (CHD) is multifactorial, and its manifestation is determined by multiple gene loci and their interaction with a cohort of environmental factors. Variation at several candidate gene loci has already been shown to have a significant effect over the spectrum of plasma lipid levels observed in the population. Moreover, some variants are known to influence the interindividual variability in response to dietary and pharmacologic interventions aimed to reduce atherogenic lipoproteins. The continuous progress in this area of research is getting us closer to the development of genetic screening panels that will allow a more precise assessment of individual CHD risk and response to therapeutic interventions. PMID- 11122688 TI - New developments in the treatment of low high-density lipoprotein cholesterol. AB - Reduced levels of high-density lipoprotein (HDL) cholesterol represent an important risk factor for the development and progression of coronary artery disease. In recent years, clinical outcome studies have verified that statin therapy may reduce the risk of initial or recurrent cardiovascular events in subjects with elevated or "normal" cholesterol levels. Subgroup analysis has also revealed that patients with low HDL benefit from this therapy. Two recently presented outcome trials using fibrate therapy also demonstrated a potential role for these medications in subjects with low HDL. The use of various HDL raising agents, singly or in combination on arteriographic progression and their potential mechanisms of action are reviewed. The latter may be an important consideration in the treatment of high-risk patients with low HDL. PMID- 11122689 TI - Recent developments in the treatment of hypertriglyceridemia. AB - Hypertriglyceridemia is now recognized as an independent risk factor of coronary artery disease (CAD). A recent secondary prevention study of CAD with a statin suggested that it may be prudent to target fasting triglycerides to less than 150 mg/dL. Secondary prevention trials of CAD with drugs acting primarily on triglycerides (fibrates) have shown that reducing triglycerides and increasing high-density lipoprotein (HDL) cholesterol, without significantly affecting low density lipoprotein cholesterol slows down coronary artery luminal narrowing (Lopid Coronary Angiography Trial [LOCAT], Bezafibrate Coronary Atherosclerosis Intervention Trial [BECAIT], Bezafibrate Infarction Prevention [BIP]). Furthermore, Veterans Administration-HDL Intervention Trial (VA-HIT) and Gruppo Italiano per lo Studio della Sopravvivenza nell'Infarto-1 (GISSI) studies recently showed that gemfibrozil and fish oils, respectively, decreased CAD mortality in secondary prevention trials. Statins are also capable of significantly reducing high triglyceride levels. Further clinical studies are necessary to confirm in terms of mortality the beneficial effect of reducing triglycerides and increasing high-density lipoprotein cholesterol in secondary CAD prevention; whereas, in primary prevention the beneficial effect of drastically reducing triglycerides by the way of pharmacology needs to be proved. PMID- 11122690 TI - The Air Force/Texas Coronary Atherosclerosis Prevention Study: implications for preventive cardiology in the general adult US population. AB - Atherosclerotic cardiovascular diseases are the most common causes of death in the United States. Fibrous plaques develop in 77% to 78% of men before age 30. The mean low-density lipoprotein-cholesterol (LDL-C) level in men aged 25 to 30 years is 117 mg/dL. In the Air Force/Texas Coronary Atherosclerosis Prevention Study, lovastatin therapy was associated with a 40% reduction in risk of fatal and nonfatal myocardial infarction. The reduction in risk of myocardial infarction was independent of baseline LDL-C level with baseline LDL-C levels as low as 125 mg/dL. The National Cholesterol Education Program guidelines should be simplified by recommending that LDL-C levels be less than 100 mg/dL in all adults in the United States. PMID- 11122691 TI - Combination drug therapy for dyslipidemia. AB - Effective treatment of dyslipidemia improves prognosis. Statin therapy has been documented to decrease the cardiovascular event rate in the setting of elevated low-density lipoprotein (LDL) cholesterol levels and coronary heart disease, but most patients are not treated to the target (LDL